PMID- 12108317 TI - [Nutritional treatment of acute diarrhea in an infant and young child]. AB - This paper written by the Comite de nutrition de la Societe francaise de pediatrie is specially devoted to the nutritional treatment of infant and child acute diarrhea, i.e. oral rehydration with salts solution and feeding. It complements an article on drug therapy of child acute diarrhea written by the Groupe francophone d'hepatologie, gastroenterologie et nutrition pediatriques, and published in this same issue of the Archives de pediatrie. PMID- 12108318 TI - [Drug treatment of acute infectious diarrhea in infants and children]. AB - In this paper written by the Groupe francophone d'hepatologie, gastroenterologie et nutrition pediatriques, recommendations are given on the indications of drugs in infant and child infectious acute diarrhea, based upon the current scientific knowledge on their effectiveness and tolerance. This paper complements an article on nutritional treatment of acute diarrhea written by the Comite de nutrition de la Societe francaise de pediatrie, and published in the same issue of the Archives de Pediatrie. PMID- 12108319 TI - [Efficacy and tolerance of vaccinations in premature infants]. AB - Immunization may prevent an enhanced risk of infectious diseases, providing that it is completed on time. A review of the literature summarizes several studies on effectiveness, safety and duration of protection in preterm infants. Immune maturation depends on chronological age rather than gestational age. Then immunization against diphteria-tetanus-pertussis-poliomyelitis-Haemophilus influenzae b should be initiated at 2 months of age and completed prior than 6 months. The youngest preterm infants, still hospitalized at 8 weeks of age should be monitored following the first immunization as they may develop apnea episodes, probably linked with the pertussis component of the vaccine. In premature, BCG vaccination induces a delayed hypersensitivity to tuberculin less important than in full-term neonates, and should not be given right after birth in newborns less than 33 weeks of gestational age. Hepatitis B vaccination should be offered as soon as two months of age and even at birth to children born from HBs Ag carriers. Neither duration of immunity, nor safety are modified by prematurity. PMID- 12108320 TI - [Periodic fevers]. AB - Periodic fever is defined as a series of unexplained febrile episodes, most often starting during childhood. The febrile episodes last usually few days, are of fixed or variable duration, and regress spontaneously, the intervals between episodes being asymptomatic. Fever is accompanied by clinical manifestations affecting peritoneal, pleural and/or mucous membranes, joints and skin. Four different etiologies are presently known. Three are hereditary diseases: familial mediterranean fever and periodic fever with hyperimmunoglobulinemia D which have a recessive autosomal transmission, and TNF receptor associated periodic syndrome or TRAPS which has a dominant autosomal transmission. One is sporadic: periodic fever with aphthous stomatitis, pharyngitis and adenopathy or PFAPA. Other etiologies are yet to be identified as many cases of periodic fever remain unexplained. PMID- 12108321 TI - [Pneumatic otoscopy in pediatrics]. AB - Pneumatic otoscopy is based upon the reaction of the tympanic membrane after sending a small air volume. A normal mobility means that air is present in the middle ear. Instead absent or reduced mobility means that effusion is present. This technique improves the quality of ears clinical examination and is particularly useful for the diagnosis of suppurative and serous otitis media. PMID- 12108322 TI - [Extraosseous Ewing's sarcoma]. PMID- 12108323 TI - [Pediatrics in French hospitals is severely threatened]. PMID- 12108324 TI - [Mortality factors in premature infants in the Department of Pediatrics of the Owendo-Libreville Hospital (Gabon)]. PMID- 12108325 TI - [Open letter from a grandmother on guard duty at the Paris Public Assistance Hospital]. PMID- 12108326 TI - [Quality control 2001]. AB - The quality control comprises of accounts and the identification of ten taxas on blind blades. The article gives the list of these pollens and details the answers. The rate of exact recognitions is 81.34% and 52 analysts out of 54 took part in the quality control. PMID- 12108327 TI - [Allergo-Pollen Bulletin. "National Bulletin" assessment 2001]. AB - The article gives the assessment of the weekly results published in the National Bulletin of RNSA: 35 bulletins were published, 46 sites sent results. Of the tables indicate: the dates of pollen traps departure to record counting for each site 2001, the number of bulletins published over four years, a comparison over six years and one 28 weeks principal period of number of weeks lost for the publication. A histogram shows the evolution of the number of results published per week in 2000 and 2001. PMID- 12108328 TI - [Temporal and geographic development of the main allergenic pollens in France: 1987-2001]. AB - At various occasions, we presented the temporal and geographical evolution of pollination during ten to fifteen last years on various sites of the French network. The year 2000 had shown often different characteristics for some tax for part of the towns of France. An hypothesis had been suggested blaming the great storm at the end of December 1999. Work presented has pure goal to prolong of one year (2001) the curves of evolution of all taxa and those of main allergenic pollens so, the share to consolidate or not the assumption 2000 and the another share, to present the assessment of pollination during the season 2001. PMID- 12108329 TI - [Aeropalynological Surveillance Rhone-Alpes (ASRA: a new network of surveillance and information]. AB - The spread of ragweed in the Lyon region and Rhone valley has lead two associations regarding this problem: Association Francaise d'Etudes Des Ambroisies (AFEDA) and Groupement d'Allergologie et d'Immunologie Clinique du Rhone-Moyen (GAICRM) to join their competences and create a new group: Surveillance Aeropalynologique Rhone-Alpes (SARA). SARA manages six pollen traps in the Lyon area and central Rhone valley and has set up an original and effective information network. PMID- 12108330 TI - [Study of the coverage of pollen capture in Lyon over three seasons (1999, 2000, 2001)]. AB - For the third consecutive year, the two sensors of the type HIRST of the National Network of aerobiology monitoring (RNSA) worked on the agglomeration of Lyon. The primary trap (Lyon 1) is located at 26 m height on a roof in southern zone of Lyon (district of Gerland), the second (Lyon 2) is located in northern zone of the city (district of Vaise) on a roof at 15 m height compared to the ground. The study of the daily variations of the pollinic counts over the first two years had shown a perfect parallelism for pollens of trees, Poaceae and Urticaceae. Only the curves of pollens of ambrosia presented different layouts between the two pollen traps. The study of this third year makes it possible to consolidate the proceeding results and to appreciate the value of the cover of a Hirst pollen trap in urban implantation within the framework of the allergo-pollinic monitoring. PMID- 12108331 TI - [Pollen composition of the air in Annaba (north-eastern Algeria)]. AB - The originality of the city of Annaba is that it is situated between two big ecological surroundings, to know, the Mediterranean sea to the north and the forest covering the massif of the Edough to the west. It incited us to determine to what point the vegetation of massif, influence it atmospheric pollen of the city of Annaba, notably in pollens allergisants. Of our survey, it comes out again that the region of Annaba is submitted to the influence of the massif of the Edough, because pollinic specter has reveled that nearly the half of pollens atmospheric identified belongs to the forest plants. The remainder of pollen belongs to humid, swampy, uncultivated zone species and of culture plants. In the end we can said that: the topography, the climatic conditions, and the speed of wind are factors determining the richness of the atmospheric in pollen. PMID- 12108332 TI - [Influence of the height position of two pollen traps in Amiens]. AB - The object of this study is to compare the results obtained in 2001 with those of the previous years and to continue the study started two years ago, making it possible to compare the daily pollinic accounts observed on two pollen traps located at the same place (Faculty of Science in Amiens), but at different heights. The primary trap is located on the roof of faculty (approximately 15 m of the ground) and the other on the ground of the same building. PMID- 12108333 TI - [Local immune tolerance mechanisms in kidney cancer]. AB - Many arguments suggest that renal tumours are immunogenic. However, the immune cells present around or within the tumour are unable to induce tumour rejection and the results of immunotherapy in metastatic renal cancer remain disappointing regardless of the protocols used. The objective of this study was to review the main mechanisms by which a renal tumour can escape immune destruction. These mechanisms can concern: tumour antigens, antigen-presenting molecules on the cell surface or defects of the cell machinery leading to the preparation of these molecules. Defects may also concern intercellular communications, especially adhesion and co-stimulation molecules. The immune cells present may also be defective, presenting qualitative or quantitative deficits, abnormalities of the T receptor, defect of cytokine production and these defects may concern both effector cells and antigen-presenting cells. The capacity of tumour cells to release anergic substances, i.e. substances which paralyze the immune system, also constitutes another very powerful immunosuppressive mechanism. These substances are cytokines, especially TGF-b. This anergy can also be mediated by intercellular contacts between tumour cells and lymphocytes, especially via the Fas system. It is important to study these mechanisms for several reasons: 1/Understanding of anergy mechanisms in order to discover new therapeutic targets or to short-circuit these mechanisms in vitro; 2/Definition of an "immune phenotype" of the tumour which should be evaluated as a prognostic marker both for survival after radical surgery of localized tumours as a prognostic factor for response to immunotherapy in metastatic forms. PMID- 12108334 TI - [Interleukin-2 and interferon in metastatic kidney cancer. Experience of the French Immunotherapy Group]. AB - The authors report the experience of the investigators of the Groupe Francais d'Immunotherapie in the treatment of metastatic renal cancer by cytokines based on several clinical trials conducted successively between 1991 and 1998. This group initially conducted a large-scale randomized trial reported in the New England Journal of Medicine in 1999, comparing three treatment regimens: intravenous interleukin-2, interferon alone and a combination of these two treatment modalities. The two cytokines demonstrated an additive effect allowing doubling of the response rate and the one-year progression-free survival. Unfortunately, this effect was not maintained in the long term and no overall survival difference was observed. Patients resistant to one of the two cytokines also failed to respond to the other cytokine administered subsequently. A subgroup of patients was identified with a probability of progression during treatment greater than 70% and a median survival of 6 months. These patients concomitantly presented initial metastases during the year following the diagnosis of renal cancer, involving several organs including the liver. Several subsequent trials testing the two cytokines by s.c. injection provided poor results, with low response rates, even in combination with 5-fluorouracil. In the French experience, s.c. cytokine regimens are therefore fairly disappointing. These results were taken into account in the ongoing evaluation programme (Percy programme) based on patient selection as a function of prognostic factors. PMID- 12108335 TI - [Nadir PSA and kinetics of PSA decline between the 3rd and 6th month after external beam radiotherapy for T1 T2 Nx M0 localized prostate cancer: value of the prediction of the risk of biological progression]. AB - INTRODUCTION: This study analyses the results of external beam radiotherapy in stage T1 T2 Nx M0 prostate cancer, with reference to the nadir PSA and the kinetics of PSA decline. MATERIAL AND METHODS: 65 patients with T1 T2 Nx M0 localized prostate cancer were treated by external beam radiotherapy (conventional or conformal) between 1990 and 1999. Two populations of 22 and 25 patients were distinguished according to the nadir PSA: population A with a nadir < or = 0.5 ng/ml or not yet reached, but with PSA < or = 0.5 ng/ml and population B with a nadir > 0.5 ng/ml. The various clinical and laboratory parameters and the kinetics of PSA decline (calculation based on the course of PSA between the first 3 and 6 months after irradiation) were compared by statistical tests (Chi square, Student t test). According to the ASTRO criteria, the results in terms of absence of biochemical progression were evaluated by non-parametric Kaplan-Meier estimate. RESULTS: No biochemical progression was observed in population A with a mean follow-up of 29.5 months. The absence of biochemical progression in population B at 42 months was 52.77%. The baseline PSA (p = 0.009), the dose delivered (p = 0.027), and the kinetics of PSA decline (p = 0.0069) were identified as predictive factors. The patient with a zero kinetic developed biochemical progression, while 91.3% of patients with a kinetic < 0.35 ng/ml/month remained free of biochemical progression. A group of patients (median nadir: 0.8 ng/ml, baseline PSA < 10 ng/ml and kinetic < 0.35 ng/ml/month) was distinguished by its good prognosis. CONCLUSION: In stage T1-T2 prostate cancer, the value of the nadir PSA is an essential prognostic factor. The kinetics of PSA decline appear to have an early predictive role. PMID- 12108336 TI - [Epidemiology of prostate cancer in the Limousin area]. AB - OBJECTIVE: Prostate cancer is the commonest cancer in men in France, and its incidence increases with age. It occupies 4th place in terms of mortality. The Limousin region, with the oldest population in France, established a regional registry of new cases of cancers in 1998. The objective of this study was to analyse the prostate cancer incidence and mortality data in the Limousin region for 1998. MATERIAL AND METHODS: Based on the cancer registry data and martality data of the department of information on medical causes of death, a registry was established for all new of prostate cancer and all cases of prostate cancer death in patients living in the Limousin region in 1998. Standardized incidence and mortality rates were compared with French data. RESULTS: The crude incidence rate detected in the Limousin region was 149.5 per 100,000 inhabitants and the standardized rate compared to the French population was 124. These rates were significantly higher than those reported in France. The mortality rate from prostate cancer in the Limousin region was 29.2 deaths per 100,000 inhabitants and was higher than that reported for France as a whole. CONCLUSION: In view of the data for 1998, prostate cancer is a major public health problem in the Limousin region. This excess incidence could be related to the methodology used, the increased awareness of general practitioners of the problems of prostate cancer, who do not hesitate to request individual screening by prostatic-specific antigen (PSA) or the presence of specific risk factors in the Limousin region. Further studies must be conducted in the Limousin region. PMID- 12108337 TI - [Prognostic factors of prostate cancer treated with first-line hormone therapy]. AB - OBJECTIVE: Endocrine therapy of prostate cancer is designed to eliminate the action of androgens to prevent the growth of hormone-sensitive cancer cells. The duration and quality of the response to this treatment vary from one patient to another. The objective of this study was to evaluate the prognostic factors of patients treated by first-line endocrine therapy for prostate cancer. MATERIAL AND METHODS: From September 1988 to September 1999, 170 patients receiving first line endocrine therapy for prostate cancer were included. Endocrine therapy consisted of LHRH analogues alone in 59.4% of cases, complete androgen suppression in 21.2% of cases, antiandrogens alone in 17.6% of cases and oestrogens in 1.8% of cases. Clinical, laboratory, scintigraphic and histological data were collected. The overall survival was studied by univariate analysis as a function of pretreatment criteria using Kaplan-Meier survival curves. RESULTS: The mean survival of the population was 33 +/- 26 months (range: 2-126 months). The following parameters were associated with a significant reduction of overall survival: age greater than 70 years, high ECOG score, mode of presentation of the cancer by alteration of general state, presence of bone pain, presence of bone metastases particularly in the appendicular skeleton, dilatation of renal cavities, Gleason score > 5, PSA level > 100 ng/ml, haemoglobin < 13.5 g/dl, serum creatinine > 105 mumol/l, low plasma testosterone, high alkaline phosphatase, poor response to treatment assessed by PSA greater than 4 ng/ml at 3, 6 and 12 months. CONCLUSION: Determination of these prognostic factors can be used to predict the patient's response to endocrine therapy and survival. These factors can also be used to stratify patients in comparable groups for clinical trials. PMID- 12108338 TI - [Study of intermittent endocrine therapy in patients presenting with biologic recurrence after radical prostatectomy or radiotherapy]. AB - INTRODUCTION: Study of the efficacy of intermittent endocrine therapy after failure of local treatment. MATERIAL AND METHODS: 74 patients were treated for biochemical recurrence after radical prostatectomy (n = 30), radiotherapy (n = 28) or radical prostatectomy followed by radiotherapy (n = 16). Treatment (63 patients were treated by antiandrogens alone, 8 by LHRH analogue and 3 by complete androgen suppression) was continued for 6 months after obtaining undetectable PSA levels for patients after radical prostatectomy (and restarted when PSA > 4 ng/ml) or a PSA nadir < 4 ng/ml for the other patients (and restarted for PSA > 10 ng/ml). RESULTS: The duration of periods without treatment represented 50% of the total treatment cycle. With a mean follow-up of 43.8 months, the overall 5-year biochemical progression-free survival rate was 54.6%. On multivariate analysis, factors predictive of biochemical progression were age less than 70 years (p = 0.05), Gleason score greater than or equal to 8 (p = 0.038) and the presence of lymph node metastases (p = 0.05). CONCLUSION: Intermittent endocrine therapy is a treatment option for patients with recurrence after local treatment. Candidates for intermittent endocrine therapy must be over the age of 70, with localized adenocarcinoma and a Gleason score less than or equal to 7. PMID- 12108339 TI - [Can urinary corpuscular volume using automated hematology contribute the diagnostic etiology of hematuric microscopy?]. AB - OBJECTIVES: To assess the value of measuring urinary corpuscular volume (UCV) with a haematology automat for determination of the glomerular or non-glomerular origin of isolated microscopic haematuria (MHi). MATERIAL AND METHODS: Forty-five fresh urine samples taken from 45 consecutive patients, with a median age of 59 years, consultant for MHi on urinary dip-sticks were studied with a haematology automat. After each analysis, MHi was classified into one of the following three groups: High MHi (glomerular, UCV < 50 fl), low MHi (non-glomerular, UCV > 50 fl), MHi of unknown origin (mixed UCV). A complete aetiological assessment of MHi was performed in each case. RESULTS: The aetiological diagnosis of MHi was established in 28 cases (62%), with a glomerular origin in 16 cases and a non glomerular origin in 12 cases. UCV was of the glomerular (19 cases), non glomerular (3 cases) or mixed type (6 cases). When UCV indicated a glomerular or non-glomerular origin for MHi (22/28 = 78.6%), the result was exact in 77% of cases (17/22). The sensitivity to diagnose glomerular haematuria was 100%, the specificity was 37.5%, the PPV was 73.7% and the NPV waas 100%. CONCLUSION: The study of UCV can suggest the origin of MHi, but cannot confirm this origin due to its poor specificity. PMID- 12108340 TI - [Postoperative osteitis of the pubis: diagnosis, treatment and results]. AB - OBJECTIVE: Osteomyelitis of the pubis is a possible complication of urological and gynaecological surgery. This article reviews this disease which is still a source of controversy. MATERIAL AND METHODS: Between 1984 and 1997, five patients with osteomyelitis of the pubis, following complementary investigations, underwent resection of the symphysis pubis with histological and bacteriological examination, combined with antibiotic therapy. The follow-up was 1.5 months to 4 years. RESULTS: The time to onset of first symptoms was 2 weeks to 5 years. Pain initially presented inflammatory features during the early postoperative period and subsequently became mechanical as the disease became chronic, with intermittent acute episodes in 3 cases. Radiation of the pain and the abnormalities observed on imaging were predominantly unilateral in all patients. Fever and laboratory signs of inflammation were inconstant. Bacteria were identified in 4 cases in the absence of antibiotics and lesions of chronic osteomyelitis were observed in every case. Antibiotics were administered by intravenous injection for 3 to 7 weeks, then orally for 1.5 to 3 months. Good functional and infectious results were obtained in 4 patients, while the remaining patient presented posterior instability. CONCLUSION: Postoperative pubic osteomyelitis is an infectious disease. Acute osteomyelitis, which can be treated by antibiotics in the absence of a collection and bone sequestrum, is distinguished from chronic osteomyelitis, in which every effort must be made to identify the bacteria responsible. Infected tissues must be widely excised in combination with prolonged antibiotic therapy. Posterior instability is the main complication. PMID- 12108341 TI - [Prevalence of erectile dysfunction in France: results of an epidemiological survey conducted on a representative sample of 1004 men]. AB - OBJECTIVE: Several studies have recently reported the prevalence rates of erectile dysfunction (ED) that vary according to the country. The authors report the results of a survey conducted in France in 2001. MATERIAL AND METHODS: This study was conducted by telephone conversation with a representative sample of 1,004 men over the age of 40. The prevalence of ED was measured by the patient's self-assessment and by the IIEF-5 index. RESULTS: The results show that about 1 man in 3 (31.6%) suffers from ED in France, which is essentially mild to moderate (according to IIEF-5). The majority of men reporting ED declared to be dissatisfied with their relationship with their partner. Finally, a large proportion of men declared that they would be willing to consult a practitioner. CONCLUSION: This survey confirms the high prevalence of erectile dysfunction in France. It is responsible for a distress that is still difficult to express and assess. This disease needs to be more accurately evaluated by rigorous and standardized methods, especially as simple and effective treatments are now available. PMID- 12108342 TI - [Relationship between the Area Under the Curve of detrussor contraction during micturition and the usual parameters of flow-pressure rate in women]. AB - OBJECTIVES: The aim of this work was to assess the relationship between a new urodynamic parameter Area Under Curve of detrusor pressure during voiding (AUCdet) and the usual pressure flow study parameters. MATERIAL AND METHODS: 103 women with various urinary symptoms (incontinence, dysuria, chronic bladder pain or frequent urinary infection) had a urodynamic evaluation with pressure flow analysis. The intravesical pressure was measured through a 6CH Porges Neoplex and the intra-abdominal pressure through a 6CH rectal catheter in an empty rectum. Pressure-flow study was performed one time in each patient. The maximum flow rate (Qmax), the detrusor pressure at maximal flow (PdetQmax), the vesical pressure at maximal flow (PvesQmax), the urethral resistance defined as the ratio between the PvesQmax and Qmax2 (RU) and the AUCdet/Vol were recovered. RESULTS: 85 women were included and mean age was 55 years old. The averages were 19.42 for Qmax, 28.91 for PdetQmax, 63.85 for PvesQmax, 2.10 for RU and 7.83 for AUCdet/Vol. The correlation with AUCdet/Vol were for Qmax -0.80 p < 0.0001, for PdetQmax 0.71 p < 0.0001, for PvesQmax 0.42 p < 0.001 and for RU 0.84 p < 0.0001. CONCLUSIONS: AUCdet/Vol is the first urodynamic parameter which takes into account the whole detrusor contraction during voiding. AUCdet/Vol is relatively close to the urethral resistance. The very high degree of correlation between RU and AUCdet/Vol confirms clinically this definition of the RU. Further studies are needed to assess the validity of AUCdet/Vol. PMID- 12108343 TI - [Auto-reeducation of female stress urinary incontinence]. AB - The Pelvia device is the fruit of the latest anatomic, functional and physiotherapeutic research into the pelvic floor. Thanks to its sophisticated technology, Pelvia is a user-friendly device fully suited to today's lifestyle of women, allowing them to do self-retraining exercises of the perineal muscles as an adjunct to the other therapies available. It is a reliable method to make patients aware of the strength and improvement of th perineal muscles contraction. It has nothing to do with "vaginal weight cones". This method is based on feedback, but the weight of the device itself does not only increase awareness but allows patients to increase their muscular strength. 30 women with stress urinary incontinence followed the protocol of self-retraining. All of them presented the following symptoms: urinary leakage, urethral urinary incontinence due to urethral hypermobility or sphincteral insufficiency, a perineum muscular testing below 4. After retraining the results were as follows: 17 cases of cure (57%); 9 cases (30%) were improved without full disappearance of incontinence; 4 cases failed (13%). The average follow-up period is 10 months (2-22). The results observed in 30 women seem promising. Of course this is only a small series and results need confirming by studying a larger series with a longer follow-up. PMID- 12108344 TI - [Obstructive ureteral calculi in children. Retrospective analysis of 24 cases]. AB - OBJECTIVE: To specify the characteristics of symptomatic ureteric stones in children and to define therapeutic indications for these stones. MATERIAL AND METHODS: Between 1993 and 2000, 24 children were treated for obstructive ureteric stones. The assessment comprised ultrasound and plain x-ray of the abdomen. A metabolic work-up and cystography +/- IVU were performed secondarily. First-line treatment consisted of medical treatment only in 34% of cases (8/24), piezoelectric extracorporeal lithotripsy (ESWL) (EDAP LT02) in 58% of cases (14/24) and ureteroscopy with ballistic fragmentation in 8% of cases (2/24). RESULTS: This series consisted of 15 boys and 9 girls with a mean age of 6 years (range: 1 month to 15 years). The most frequent presenting complaints were renal colic in 14 cases and acute pyelonephritis in 7 cases. The obstructive ureteric stone was associated with urinary tract infection in 1/3 of cases. 12 children had a single stone and 12 children had multiple stones. The stone was situated in the pelvic ureter in 15 cases, the iliac ureter in 4 cases, and the lumbar ureter in 6 cases. The aetiological survey revealed hypercalciuria in 6 cases, cystinuria in 2 cases, and 4 urinary tract malformations. In 34% of cases (8/24), the stone was eliminated spontaneously (phi < or = 5 mm). 78% (11/14) of stones treated by ESWL (14/24) (mean phi = 8.3 mm) left no residual fragments with this treatment alone. The re-treatment rate was 36% (4/11). Three failures of ESWL were treated by ureteroscopy in 2 cases and surgery in 1 case. 92% (22/24) of these children were stone-free with a mean follow-up of 16 months. DISCUSSION: Obstructive ureteric stones in children are associated with a high risk of infectious complications, justifying rapid and appropriate treatment. A metabolic abnormality must always be investigated and is present in 1/3 of cases. A stone diameter of 5 mm corresponds to the upper limit allowing spontaneous elimination during medical treatment. ESWL is the treatment of choice for ureteric stones greater than 5 mm in diameter, especially in the pelvic ureter. Ureteroscopy can be performed in children after failure of ESWL and for complicated stones. PMID- 12108345 TI - [Ureteral multiplicity: report of 3 cases]. AB - The ureteric multiplicity is a rare malformation with about one hundred cases described in the literature. The rarity of this anomaly and the complexity of possible associated anomalies often make the preoperative diagnosis difficult. The authors report 1 case of partial quadruplicate ureter associated with bladder duplication and 2 cases of triplicate ureter, corresponding to types I, II and III of Smith's classification. Based on their experience and a review of the literature, they discuss the diagnostic approach and treatment of these anomalies. PMID- 12108346 TI - [Laparoscopic treatment of the nonpalpable testis. Results]. AB - OBJECTIVE: The purpose of laparoscopy in the management of the non palpable testis is to provide information regarding testicular presence and location to facilitate overall surgical management. MATERIALS AND METHODS: We report our experience with laparoscopic orchiopexy to treat 39 nonpalpable testes in 32 patients, patient age ranged from 2.3 years to 14 years (average 4.18 years). We retrospectively reviewed the medical records of all patients for a 5-year period. RESULTS: At laparoscopy 11 testes were at the internal inguinal ring, these patients underwent one-stage laparoscopic orchiopexy without division of the spermatic vessels. 18 high intra-abdominal testes underwent two-stage Fowler Stephens orchiopexy. One Patient underwent laparoscopically assisted orchiectomy for atrophy, 9 testes were absent. At follow-up 6, 12 and 24 months after 26 of 29 (89%) testes are without atrophy, and in acceptable scrotal position. CONCLUSIONS: The low incidence of complications and 89% success rate underscore the feasibility of laparoscopic orchiopexy. It is our procedure of choice for the management and treatment of nonpalpable testis. PMID- 12108347 TI - [E coli spondylitis after ureteroscopy for lithiasis]. AB - The authors report a rare complication of ureteroscopy for stones of the lumbar ureter: Escherichia coli spondylitis. PMID- 12108348 TI - [Vaginal delivery in a patient with an artificial urinary sphincter and augmentation enterocystoplasty]. AB - Few cases of management of pregnancy in patients with an artificial sphincter (AS) have been reported in the literature, but this situation is very likely to become increasingly frequent with the more frequent use of AS in children. The authors report the case of a pregnancy in a 36-year-old woman with lumbosacral spina bifida, treated by artificial urinary sphincter and augmentation enterocystoplasty. The pregnancy and vaginal delivery did not raise any particular problems. Vaginal delivery is possible provided several precautions are observed: pelvimetry, MRI at the 8th month of pregnancy and close collaboration between urologists and obstetricians near term. Emergency caesarean section should be avoided due to the risk of lesions caused by elements of the AS and the surgical difficulties related to the presence of the augmentation enterocystoplasty. PMID- 12108349 TI - [Secondary tumors of the kidney]. AB - The frequency of secondary tumours of the kidney is underestimated, especially as their diagnosis is often delayed, after nephrectomy or autopsy. The authors present the case of a 59-year-old patient presenting with renal metastases from lung cancer operated two years previously and review about forty cases published in the literature over the last twelve years. The diagnosis by percutaneous needle biopsy can avoid useless surgery and improve patient management. The specificity and sensitivity of this method need to be evaluated. PMID- 12108350 TI - [Can ofloxacin constitute an alternative antitubercular treatment for BCG hepatitis?]. AB - The authors report a case of severe BCG hepatitis resistant to tuberculostatic therapy and treated with ofloxacin. Modern antibiotics, such as certain fluoroquinolones, can constitute an alternative to tuberculostatic therapy in the management of serious adverse effects of BCG therapy. PMID- 12108351 TI - [Linitis plastica type of primary signet cell adenocarcinoma of the bladder]. AB - Primary adenocarcinoma represent 0.5 to 2% of all bladder tumours and are classified according to whether or not they are derived from the urachus, although, histologically, this classification now appears to be obsolete. The authors report a very rare case of linitis plastica type of primary signet cell adenocarcinoma of the bladder in a 53-year-old patient. This carcinoma, with very unusual histological features, needs to be distinguished. Due to the delayed diagnosis, it has a poor prognosis despite the most aggressive treatment modalities, as reported in the literature. The elevated CA 19-9 observed in the present case may be a useful marker for follow-up. PMID- 12108352 TI - [Lympho-epithelioma of the bladder. Discussion of pathogenesis]. AB - Lympoepithelioma, originally described within the nasopharynx is an undifferenciated malignant epithelial tumor with a prominent lymphoid stroma. We report a case of lymphoepithelioma of the bladder after intravesical BCG treatment for carcinoma in situ. PMID- 12108353 TI - [Fibromyxoid tumor of the bladder: report of 3 cases]. AB - Fibromyxoid tumours are rare inflammatory bladder tumours. These benign tumours are responsible for minimal symptoms. The diagnosis is based on histological examination showing spindle cells infiltrating the detrusor associated with more or less abundant inflammatory infiltrates. It is important to demonstrate the myofibroblastic nature of these cells, which constantly express specific muscle actin and smooth muscle actin on immunohistochemistry. The differential diagnoses are high-grade sarcomatoid spindle cell urothelial carcinoma, leiomyosarcoma and embryonal rhabdomyosarcoma in children. Treatment is surgical, comprising complete endoscopic resection or partial or total cystectomy. The authors report 3 new cases of fibromyxoid tumours treated conservatively by complete endoscopic resection or partial cystectomy. No recurrence was observed. The authors present a review of the literature and a discussion of the differential diagnoses. PMID- 12108354 TI - [Pseudo-tumoral colic metaplasia of the urinary bladder]. AB - Colic metaplasia of the urinary bladder is a rare disease, secondary to a chronic irritative factor. In its minor form, it has the same clinical features as simple cystitis, but its major pseudoneoplastic form may be mistaken for bladder tumor. The diagnosis is essentially histological. Treatment is based on eradication of the irritative factor and of the resection of pseudoneoplastic form. Surgery is performed in the case of complications of this disease. The clinical course is unclear, requiring long-term surveillance. We report one case of colic-type glandular metaplasia of the urinary bladder in a 50 years-old patient. The clinical symptomatology was dominated by hematuria and pollakuria. A bladder neoplasm was highly suspected in ultrasound and endoscopic findings. The patient underwent a transurethral resection of the bladder tumor. Histological examination of of resection shavings revealed a colic-type glandular metaplasia. PMID- 12108355 TI - [A rare entity: cystadenoma of the prostate]. AB - The authors report the case of a 40-year-old man with cystadenoma of the prostate. This rare disease is difficult to diagnose despite a complete radiological assessment (transrectal and bladder ultrasound and MRI of pelvis). Only histological examination of the operative specimen demonstrated absence of malignancy. However, follow-up is required in view of the risk of recurrence of this prostatic lesion. PMID- 12108356 TI - [Buschke-Lowenstein penile tumor]. AB - The Buschke-Lowenstein genital tumour is a poorly defined, uncommon tumour. The distinction between benign lesions, potentially malignant lesions and carcinomatous lesions is difficult. The authors report a case of Human Papillomavirus (HPV) 11-associated Buschke-Lowenstein tumour with an area of micro-invasive carcinoma on histological examination of the surgical resection specimen. A 34-year-old patient was operated for recurrent condylomatous lesions of the penis with scrotal extension. Histological examination of the complete operative specimen confirmed the presence of Buschke-Lowenstein tumour as well as an area of dermal micro-invasion on one section. Molecular hybridization revealed the presence of HPV 11 DNA and immunohistochemistry showed basal cells weakly expressing mutant p53. The classification of Buschke-Lowenstein tumours is controversial. Some authors consider these tumours to be benign tumours or giant condylomata (non-metastatic, associated with HPV 6-11), while others consider these tumours to be borderline malignant (local extension and risk of progression to invasive carcinoma). The role of HPV as cofactor involved in carcinomatous transformation also remains controversial. The authors emphasize the need for surgical resection of this type of tumour with histological examination of the entire operative specimen looking for areas of micro-invasion. In the presence of micro-invasion with healthy resection margins and staging by clinical examination and complementary investigations, treatment essentially consists of regular surveillance. PMID- 12108357 TI - [Surgical treatment of penile-scrotal elephantiasis]. AB - Penoscrotal elephantiasis is a rare disease outside of filariasis endemic regions. The authors report a case of primary penoscrotal elephantiasis treated by complete surgical resection of the pathological tissue and penoscrotal reconstruction, with a good functional and cosmetic result. The complementary investigations indicated and the various treatment options are analysed in the light of a review of the literature. PMID- 12108358 TI - [Control of the development of Onuf's spinal nucleus]. AB - Onuf's nucleus is located at the sacral level of the spinal cord and is formed by the motoneurons innervating the perineal muscles. Both the number and the size of these neurons vary between males and females creating a so-called sexual dimorphism. This dimosphism is mediated by androgens. In absence of these hormones, the motoneurons die by apoptosis. This androgenic effect is not mediated directly on the motoneurons but on perineal muscles. These muscles survive by the action of androgens and they secrete factors that act to promote motoneuronal survival. One of these factors could be CNTF (Ciliary Neurotrophic Factor). Later during development, androgens act directly on motoneurons either by binding their own receptor or by intracellular transformation into estrogens due to the action of the aromatase. PMID- 12108359 TI - [Anatomic and experimental basis for nerve grafts after radical retropubic prostatectomy]. AB - A nerve graft technique has been developed to restore erectile function after radical prostatectomy, following bilateral resection of the neurovascular bundles. The objective of this review of the literature is to describe this new surgical technique and analyse its anatomical and experimental bases. According to the current data of the literature, there is little scientific evidence to support the preliminary results. PMID- 12108361 TI - [Reaction to the rubric, "Controversies and points of view"]. PMID- 12108360 TI - [Predictive factors for technical difficulties during retropubic radical prostatectomy using external pelvimetry]. AB - External pelvimetry can be used to predict the technical difficulty of retropubic radical prostatectomy and the results should be taken into account in the treatment plan. PMID- 12108362 TI - [Reaction to the article, "Cost-effectiveness analysis of dental implantation"]. PMID- 12108363 TI - [Letter to the editor]. PMID- 12108364 TI - Smoke free. PMID- 12108365 TI - Learning curve. PMID- 12108366 TI - Self-preservation. PMID- 12108367 TI - Prisoners at work. PMID- 12108368 TI - Could it happen here? PMID- 12108369 TI - A dying art. PMID- 12108370 TI - Fact or fiction. PMID- 12108371 TI - Day in the life. PMID- 12108372 TI - Never fear. PMID- 12108373 TI - Diary of a nurse with cancer. Double whammy. PMID- 12108374 TI - Day in the life. An occupational health nurse at a nuclear power station. PMID- 12108381 TI - Gleanings about dentistry from the world of literature (twenty-sixth in a series). PMID- 12108382 TI - Dental questions in Frank Norris's novel "McTeague". AB - Frank Norris's McTeague, one of the first novels in which the main character is a dentist and dentistry is central to the plot, has been a minor member of the American literary canon for over a century. Possibly because Norris was such a committed and practicing realist/naturalist, his characterization of McTeague as a believable dentist has not until now been questioned. Nor has his use or misuse of the California Dental Licensure Legislation of 1885 to bring about McTeague's downfall been subjected to historical analysis. Such analysis may cast doubt on the credibility and reputation of the novel. PMID- 12108383 TI - Five dental comedy films produced by the Three Stooges (1938-1951). PMID- 12108384 TI - [A new proposal for uniform hepatic segmental anatomic nomenclature]. PMID- 12108385 TI - [Effects of TNFalpha and dexamethasone on the expression of intercellular cell adhesion molecule-1, P-selectin in patients with intrahepatic bile duct obstruction]. PMID- 12108386 TI - [The relationship between liver fibrosis stages and the diameters of broadening main portal vein and spleen vein, splenectasis, and hematological tests]. PMID- 12108387 TI - [The immunohistochemistry of adhesion molecules in the myocardium in experimental cirrhotic rats]. PMID- 12108388 TI - [Suggestions for the diagnostic criteria of alcoholic hepatopathy]. PMID- 12108389 TI - [Apoptosis effects of human liver carcinoma cells induced by 5-fluorouracil]. PMID- 12108390 TI - [The expression of bcl-xL mRNA in the tissues of normal liver and hepatocarcinoma]. PMID- 12108391 TI - [Lamivudine treatment consensus from relative experts]. PMID- 12108392 TI - [The interaction between 5-fluorouracil and marini on QGY cell line in vitro]. PMID- 12108393 TI - Mitochondrial encephalomyopathies:gene mutation. PMID- 12108394 TI - [Abstracts of the 19th Congress of the Hungarian Federation of Endocrine Societies and the Symposium of the European Federation of Endocrine Societies. 15 17 May 2002, Gyula]. PMID- 12108396 TI - Managed care outlook. Small employers plan to shift increased costs to workers. PMID- 12108395 TI - Compensation monitor. Radiologists, anesthesiologists in demand. PMID- 12108397 TI - List of education centers and teachers of ultrasound in the world. PMID- 12108398 TI - Smallpox: mass vaccinations or not? PMID- 12108399 TI - Law. The right to choose. PMID- 12108400 TI - Law. Traveller's checks. PMID- 12108401 TI - Law. Race against time. PMID- 12108402 TI - Eating on the run. PMID- 12108403 TI - Automated dispensing machines need approval. PMID- 12108404 TI - GPOs: good or evil? Studies show answer depends on approach of researchers. PMID- 12108405 TI - [Creutzfeldt-Jakob disease/BSE special issue]. PMID- 12108406 TI - Eggs of the pig whipworm Trichuris suis. PMID- 12108407 TI - Information for patients. What you should know about constipation. PMID- 12108408 TI - WHO to support new National Institutes of Health research programme. PMID- 12108410 TI - [Abstracts of the 51st Meeting of the North-German Association of Ophthalmology. Schwerin, 25-26 May 2002]. PMID- 12108409 TI - WHO medicines expert calls for new rules against commercial bias in medical research. PMID- 12108411 TI - Redescription of the fish parasite Lernanthropus polynemi Richiardi, 1881 (Copepoda: Siphonostomatoida) and relegation of two congeners to synonymy. AB - Lernanthropus polynemi Richiardi, 1881 is described, for the first time since its original discovery, based on the type-material found in the Museum fur Naturkunde in Berlin, Germany. Morphological features of this parasite, which infects the commercially-caught fish Eleutheronema tetradactylum (Shaw), are described and illustrated, including those of the male, which is described for the first time. The types of another species from the same host, L. lappaceus Wilson, 1912, and deposited in the National Museum of Natural History in Washington DC, are also examined based on the type-specimens. The types of a third congener, L. trifoliatus Basset-Smith, 1898, also from the same host, are not available, but the original illustrations are studied. It is considered that the three nominal species described from E. tetradactylum represent a single taxon, and that L. lappaceus and L. trifoliatus be relegated to synonymy with L. polynemi. PMID- 12108413 TI - Abstracts of the 37th Congress of the European Society for Surgical Research (ESSR). Szeged, Hungary, May 23-25, 2002. PMID- 12108412 TI - Davisia hexagrammi n. sp. (Myxosporea: Sinuolineidae) parasitic in the urinary bladder of marine fishes from the coasts of the Yellow Sea and Bohai Bay, China. AB - A new species of myxosporean (Sinuolineidae), Davisia hexagrammi n. sp. from marine fishes collected from the coasts off Qingdao (Tsingtao) and Rongcheng on the Yellow Sea coast and Laizhou Bay, China, is described. Both spores and plasmodia were found in the urinary bladder of two hosts, Hexagrammos otakii Jordan & Starks and Agrammus agrammus (Temminck & Schlegel). The diagnostic features of D. hexagrammi n. sp. are: trophozoite monosporous or disporous; spore body spherical, 9.5-10.5 (9.9 +/- 0.5) x 10-12 (10.8 +/- 1.0) microm in size and with a straight or slightly sinuous sutural line; two shell valves each with one long, hollow lateral appendage of 44-59 (52.4 +/- 6.9) microm in length; two spherical polar capsules arranged anteriorly, well-separated from each other and 3.5-4 (3.8 +/- 0.2) microm in diameter, coelozic; marine habitat. PMID- 12108414 TI - Fracture Treatment in Elderly and Osteoporotic Patients. Symposium of the International Society for Fracture Repair. Bologna, Italy, 16-18 May 2002. Abstracts. PMID- 12108415 TI - Association of Surgeons of Great Britain and Ireland and Society of Academic and Research Surgery joint annual meeting. Dublin, Ireland, 22-24 May 2002. Abstracts. PMID- 12108416 TI - [108th Congress of the French Society of Ophthalmology. 11-15 May 2002, Paris, France. Abstracts]. PMID- 12108417 TI - Caribbean Health Research Council 47th Annual conference. April 24-27, 2002, Guyana. Abstracts. PMID- 12108418 TI - 2002 Congress on In Vitro Biology and 10th IAPTC&B (International Association for Plant Tissue Culture and Biotechnology) Congress, June 25-29, 2002. Orlando, Florida, USA. Abstracts. PMID- 12108420 TI - Master index to volumes 81-90. PMID- 12108419 TI - Prevention and control of occupational infections in health care. An infection control guideline. Canada Communicable Disease Report. PMID- 12108421 TI - Abstracts of the XIII International Congress of Histocompatibility and Immunogenetics. Seattle, Washington, USA. 18-22 May 2002. PMID- 12108423 TI - Index 1982-2001, volumes 1-20. PMID- 12108422 TI - FOCIS 2002. Federation of Clinical Immunology Societies Meeting. San Francisco, California, USA. June 28-July 1, 2002. Abstracts. PMID- 12108424 TI - Abstracts of the Society for Reproduction and Fertility Annual Conference. University of Leeds, July 2002. PMID- 12108425 TI - Bleeding peptic ulcers: what's new? PMID- 12108426 TI - Association of Veterinary Teachers and Research Workers. Current Topics in Veterinary Science 56th Annual Conference. Scarborough, United Kingdom, 25-27 March 2002. Abstracts. PMID- 12108427 TI - Abstracts of the 4th Central European Congress for Rheumatology. Baden, Austria, May 10-12, 2002. PMID- 12108428 TI - Abstracts of the 5th International Diabetes Federation Western Pacific Region Congress. Beijing, China, 4-7 May 2002. PMID- 12108430 TI - Abstracts of the XXX European Muscle Conference. Pavia, Italy, 9-12 September 2001. PMID- 12108429 TI - Developmental differences in the role of interleukins in hyperoxic lung injury in animal models. AB - Interleukins (IL) are part of the group of immune mediators known as cytokines. IL are produced by many different cells and possess a wide spectrum of biological activities. This review will be focused on the role of IL-1 to 6, 8, 10-13 as it pertains to the effects of hyperoxia on the adult and newborn lung in animal models. Hyperoxic exposure to the adult and newborn lung had variable effects on the expression of IL-1alpha and IL-1beta. Increased IL-6 levels were seen in adult lungs by day 3 and in the newborn lungs by day 10 of exposure to hyperoxia. IL-8 also peaked around day 10 in the newborn lung but there were no significant changes in IL-10. Pretreatment with IL-1, endotoxin, rhSOD, lidocaine, lisofylline, pentoxifylline and overexpression of IL-6, 11, and 13 seemed to attenuate hyperoxic lung injury in the adult. This protection was accompanied by increased pulmonary MnSOD, VEGF expression and decreased apoptosis. It is clear that IL have a significant role to play in hyperoxic lung injury. Increased IL expression and release has a cascade effect and appears to predate the influx of inflammatory cells. There are significant differences in the type and timing of IL expression and release in the adult and newborn lung in response to hyperoxia. Designing a therapeutic approach to counteract oxygen toxicity in the immature lung first needs understanding of the unique responses in the newborn. PMID- 12108431 TI - Abstracts of the 18th UICC International Cancer Congress. 30 June-5 July 2002. Oslo, Norway. PMID- 12108432 TI - Commentary on Barro's 'First interview and one session with "Ana"'. PMID- 12108433 TI - Transplant 2002. American Transplant Congress. April 26-May 1, 2002. Washington, DC, USA. Abstracts. PMID- 12108434 TI - Abstracts of the 11th International Congress on Cardiovascular Pharmacotherapy. Montreal, Canada, 18-21 May 2002. PMID- 12108435 TI - The evaluation of renal transplantation candidates: clinical practice guidelines. PMID- 12108436 TI - Transplant 2001. The ASTS and AST Joint American Transplant Meeting. May 11-16, 2001. Chicago, Illinois, USA. Abstracts. PMID- 12108437 TI - Videoconferencing for continuing medical education: from pilot project to sustained programme. AB - Videoconferencing has been used to provide distance education for medical students, physicians and other health-care professionals, such as nurses, physiotherapists and pharmacists. The Dalhousie University Office of Continuing Medical Education (CME) has used videoconferencing for CME since a pilot project with four sites in 1995-6. Since that pilot project, videoconferencing activity has steadily increased; in the year 1999-2000, a total of 64 videoconferences were provided for 1059 learners in 37 sites. Videoconferencing has been well accepted by faculty staff and by learners, as it enables them to provide and receive CME without travelling long distances. The key components of the development of the videoconferencing programme include planning, scheduling, faculty support, technical support and evaluation. Evaluation enables the effect of videoconferencing on other CME activities, and costs, to be measured. PMID- 12108438 TI - Introduction. Peat bog archives of atmospheric metal deposition. PMID- 12108439 TI - A chemical and mineralogical reconstruction of Zn-smelter emissions in the Kempen region (Belgium), based on organic pool sediment cores. AB - The atmospheric pollution history of a former Belgian Zn-smelter complex is preserved in organic sediments of a nearby peat bog pool. The stratigraphy of trace metals, Pb-isotope ratios and mineralogy indicate extreme heavy metal pollution in recent sediments. In the pollutant trend, extremes coincide with maxima in 20th century metal production, minima during major war conflicts and the final shutdown of the smelter. Peak pollution concentrations measure up to 4.7 wt.% Zn, 1.1 wt.% Pb and 0.1 wt.% Cd, which correspond to calculated atmospheric deposition rates of 9.0, 1.6 and 0.16 g m(-2) yr(-1), respectively. 206Pb/207Pb isotope ratios show higher values in the polluted interval (1.135 1.162) relative to local geogenic values deeper down-core (1.194-1.215). Within the polluted interval, three significantly different 208Pb/206Pb plateaus are recognized and suggested to indicate changes in the origins of processed ores. Microprobe analyses on sediment thin sections show extensive in situ FeS2 and ZnS precipitation, which suggests that anoxic processes are responsible for the immobilization of the atmospheric metal inputs. The occurrence of oxidized smelter dusts in an independent surface soil sample indicates a rapid diagenetic transformation of metal oxides into sulfides. Morphology and chemical characteristics allow the distinction between smelter related and diagenetic mineral deposits, and give evidence for dust from open-air ore repositories, as well as smelter operation without dust filters. PMID- 12108440 TI - Behavior of selected micro and trace elements and organic matter in sediments of a freshwater system in south-east Brazil. AB - The Cima Lake (CL) is a freshwater system located in the Northern Region of Rio de Janeiro State. It has a history of agricultural and cattle raising activities. The original Atlantic forest has been deforested and replaced by pasture and sugar cane crops. The objectives of this study were to understand the vertical distribution of organic matter and heavy metals associated with lacustrine sediments and to evaluate the influence of land use on the relative contribution of organic matter sources and heavy metals to the lake sediments. For these purposes, sediment cores were collected at three stations: inlet, center and outlet of the CL. It was possible to date sediments buried over the last 150 years. The results revealed that the changes caused by human intervention in the CL watershed influenced sediment sources and composition, and led to increasing sedimentation rates over the past century. The (C/N)a ratios ranged between 5.7 and 47.4 and showed that between the 1950s and 1960s there was an alteration in the sources of organic matter to CL sediments, indicating a period marked by the expansion of sugar-cane and alcohol industries in the region and an intense deforestation in the lake watershed. The heavy metal concentrations obtained in this study were similar to those described for natural systems, in spite of the system being influenced by anthropogenic sources. The Me+/Ti ratios indicate the existence of human intervention processes in the lake, showing that the origin of metals is not only from natural processes. PMID- 12108441 TI - An analytical protocol for the determination of total mercury concentrations in solid peat samples. AB - Traditional peat sample preparation methods such as drying at high temperatures and milling may be unsuitable for Hg concentration determination in peats due to the possible presence of volatile Hg species, which could be lost during drying. Here, the effects of sample preparation and natural variation on measured Hg concentrations are investigated. Slight increases in mercury concentrations were observed in samples dried at room temperature and at 30 degrees C (6.7 and 2.48 ng kg(-1) h(-1), respectively), and slight decreases were observed in samples dried at 60, 90 and 105 degrees C (2.36, 3.12 and 8.52 ng kg(-1) h(-1), respectively). Fertilising the peat slightly increased Hg loss (3.08 ng kg(-1) h( 1) in NPK-fertilised peat compared to 0.28 ng kg(-1) h(-1) in unfertilised peat, when averaged over all temperatures used). Homogenising samples by grinding in a machine also caused a loss of Hg. A comparison of two Hg profiles from an Arctic peat core, measured in frozen samples and in air-dried samples, revealed that no Hg losses occurred upon air-drying. A comparison of Hg concentrations in several plant species that make up peat, showed that some species (Pinus mugo, Sphagnum recurvum and Pseudevernia furfuracea) are particularly efficient Hg retainers. The disproportionally high Hg concentrations in these species can cause considerable variation in Hg concentrations within a peat slice. The variation of water content (1.6% throughout 17-cm core, 0.97% in a 10 x 10 cm slice), bulk density (40% throughout 17-cm core, 15.6% in a 10 x 10 cm slice) and Hg concentration (20% in a 10 x 10 cm slice) in ombrotrophic peat were quantified in order to determine their relative importance as sources of analytical error. Experiments were carried out to determine a suitable peat analysis program using the Leco AMA 254, capable of determining mercury concentrations in solid samples. Finally, an analytical protocol for the determination of Hg concentrations in solid peat samples is proposed. This method allows correction for variation in factors such as vegetation type, bulk density, water content and Hg concentration in individual peat slices. Several subsamples from each peat slice are air dried, combined and measured for Hg using the AMA254, using a program of 30 s (drying), 125 s (decomposition) and 45 s (waiting). Bulk density and water content measurements are performed on every slice using separate subsamples. PMID- 12108442 TI - The chronology of anthropogenic, atmospheric Pb deposition recorded by peat cores in three minerogenic peat deposits from Switzerland. AB - Peat cores were collected from three minerogenic peatlands in Switzerland: La Tourbiere des Genevez (TGE), in Canton Jura; Gola di Lago (GDL), in Canton Ticino; and Mauntschas (MAU), in Canton Grisons. Chemical analyses of the pore waters and the peats document the increasingly minerogenic character with depth. In particular, the Sr concentration profile in the peats reflects the growing importance of mineral sediment dissolution with increasing distance below the peat surface. Despite this circumstance, Pb concentrations are greatest in the surface layers, and decrease with depth. Thus, dissolution of the basal sediments at these sites appears to be an unimportant source of Pb, compared with atmospheric deposition. Twenty-one new radiocarbon age dates for the three cores provide a chronology of Pb enrichment, and these are consistent with the record of atmospheric Pb deposition recorded by the ombrotrophic peat bog 'Etang de la Gruere' (EGR). Sediment samples from TGE and EGR were reacted at pH 5 with ammonium acetate to dissolve the carbonate fraction, filtered, and the residues analyzed using XRF. These analyses show that much of the Ca, Mg, Sr and Mn is lost when the carbonate fraction is dissolved, but this process does not measurably affect Pb. This finding suggests that carbonate dissolution dominates the weathering of the sediments underlying these peat deposits, but this fraction does not contain significant concentrations of Pb. Minerogenic peat deposits such as those described here, therefore, can serve as reliable archives of atmospheric Pb deposition, provided that mineral dissolution in the underlying sediments does not contribute measurably to the Pb inventory of the peat profile. PMID- 12108443 TI - Atmospheric Pb deposition in Spain during the last 4600 years recorded by two ombrotrophic peat bogs and implications for the use of peat as archive. AB - Two ombrotrophic peat bogs in Northwestern Spain provided a history of 4600 years of Pb accumulation. Highest Pb concentrations (84-87 microg g(-1)) were found near the bogs' surface, but there were also other significant peaks (6-14 microg g(-1)), indicating pre-industrial atmospheric pollution. The enrichment factors (EFs) in both cores show a remarkably similar record. Atmospheric Pb pollution dates back to at least approximately 2500 years ago, reaching a first maximum during the Roman period. For the last 300 years, Pb EFs significantly increased due to industrial development, but the uppermost samples of the bogs show decreasing Pb EFs, probably due to the phasing out of leaded gasoline. These results are also supported by 206Pb/207Pb isotope ratios, as they continuously decrease from ca. 3000 BP until 2000 BP (from 1.275 at 4070 14C years BP to 1.182), indicating the growing importance of nonradiogenic Pb released from Iberian ores by ancient mining. Peat samples at a 3-5-cm depth are even less radiogenic (206Pb/107Pb = 1.157), indicating the strong influence of leaded gasoline. Despite the common history shared by the two bogs, striking differences were found for Pb enrichment, whether this was calculated by normalising to the Pb/Ti ratio of the upper continental crust or to the Pb/Ti ratios of peats from pre-anthropogenic times. This effect seems to be related to differences in Ti accumulation in both bogs, possibly due to physical fractionation of the airborne dust during wind transport. Enrichment has to be carefully considered when comparing the results obtained for different bogs, since our results suggest that normalising to crustal proportions is meaningless when the bulk of the deposition in an area is strongly influenced by short- and medium-range dust transport. PMID- 12108444 TI - Stable lead isotopes and lake sediments--a useful combination for the study of atmospheric lead pollution history. AB - Analysis of stable lead isotopes and lead concentrations in lake-sediment deposits, not least in varved (annually-laminated) sediments, is a useful method to study lead pollution history. This paper presents details from a study of 31 lakes in Sweden. Using a strong acid digestion of sediment samples and ICP-MS analyses, we have found that Swedish lake sediments have a high natural (pre pollution) 206[Pb]207[Pb] ratio (mean 1.52+/-0.18, range 1.28-2.01, n=31 lakes). In contrast, atmospheric lead pollution derived from metal smelting processes, coal burning and from alkyl-lead added to petrol has a lower ratio (< 1.2). Consequently, when pollution lead deposition began approximately 3500 years ago, the lead isotope ratio of the sediments started to decline, and in modern sediments it is typically < 1.2. Using the isotope and concentration values and a mixing model, the relative contribution of pollution and natural lead in sediment samples can be calculated. The pollution lead records of the Swedish lake sediments show a consistent picture of the atmospheric lead pollution history. Some noticeable features are the Roman peak (approx. 0 AD), the large and permanent Medieval increase (approx. 1000 AD), peaks at approximately 1200 and 1530 AD, the rapid increase after World War II, the peak in the 1970s, and the large modern decline. PMID- 12108445 TI - Stable lead isotopic characterisation of the historical record of environmental lead contamination in dated freshwater lake sediment cores from northern and central Scotland. AB - Sediment cores from three Scottish freshwater lakes, Loch Ness in the remote north and Loch Lomond and the Lake of Menteith, much closer to the heavily populated and industrialised central belt were analysed for 210Pb, 137Cs, Pb and stable Pb isotopic composition (206Pb/207Pb). The radionuclide data were used to establish chronologies for the Loch Ness and Loch Lomond cores, but a chronology could not be developed for the Lake of Menteith core, in which the surface sediment had been subject to intense mixing. Although Pb concentrations generally started increasing during the mid-17th Century, a small peak occurred for Loch Ness in the early 16th Century, perhaps attributable to the influence of medieval mining and smelting in mainland Europe. Temporal trends in the pattern of Pb accumulation were similar for Loch Ness and both sites in Loch Lomond, with 40 50% of the anthropogenic Pb deposited prior to the 20th Century. Fluxes of anthropogenic Pb to the lake sediments peaked during the 1950s at all locations where chronologies could be established. The 5-fold increase in anthropogenic Pb inventory for the southern basin of Loch Lomond relative to Loch Ness reflected geographical proximity to the main polluting sources. The 206Pb/207Pb data for anthropogenic Pb in the sediments from Loch Ness and Loch Lomond exhibited largely similar trends related to five different time periods. Pre-1820, the 206Pb/207Pb ratio was close to that for coal (1.181). From 1820 to 1900, a fairly constant 206Pb/207Pb ratio of approximately 1.17 probably resulted from a combination of emissions from the smelting of indigenous Pb ore (1.170) and coal burning (1.181) in Scotland, and industrial activity to the south in England, where Australian Pb of characteristically low 206Pb/207Pb ratio (1.04) was already in use. From 1901 to 1930, the 206Pb/207Pb ratio declined by <0.01, due to the increasing influence of Australian Pb. From 1931 to 1975/1985, the 206Pb/207Pb ratio of anthropogenic Pb declined by a further 0.03 to 0.04, to minimum values from approximately 1975 to 1985, primarily a consequence of car exhaust emissions of Pb arising from the introduction of alkyl Pb petrol additives (206Pb/207Pb approximately 1.06-1.09). From 1975/1985 to the mid-1990s, the 206Pb/207Pb ratio of anthropogenic Pb increased by up to 0.015, a consequence of a reduction in car-exhaust emissions of Pb, resulting from reductions in the maximum permitted concentration of Pb in petrol, and the introduction and increasing uptake of unleaded petrol. Source apportionment calculations, on the basis of 206Pb/207Pb values in surface sediment, suggested that the contribution of Pb emissions from the use of leaded petrol was 27-40% of the atmospheric burden by the mid-1990s, in line with estimates from rainwater 206Pb/207Pb data. PMID- 12108446 TI - Geochemical evidence for atmospheric pollution derived from prehistoric copper mining at Copa Hill, Cwmystwyth, mid-Wales, UK. AB - This paper presents geochemical data from a blanket peat located close to a Bronze Age copper mine on the northern slopes of the Ystwyth valley, Ceredigion, mid-Wales, UK. The research objective was to explore the possibility that the peat contained a geochemical record of the pollution generateD by mining activity. Four peat monoliths were extracted from the blanket peat to reconstruct the pollution history of the prehistoric mine. Three different geochemical measurement techniques were employed and four copper profiles have been reconstructed, two of which are radiocarbon-dated. The radiocarbon dates at one profile located close to the mine confirm that copper enrichment occurs in the peat during the known period of prehistoric mining. Similar enrichment of copper concentrations is shown in one adjacent profile and a profile within 30 m away. In contrast, copper was not enriched in the other radiocarbon-dated monolith, collected approximately 1.35 km to the north of the mine. Whilst other possible explanations to explain the copper concentrations are discussed, it is argued that the high copper concentrations represent evidence of localised atmospheric pollution caused by Bronze Age copper mining in the British Isles. The results of this study suggest that copper may be immobile in blanket peat and such deposits can usefully be used to reconstruct atmospheric pollution histories in former copper mining areas. PMID- 12108447 TI - Comparative study of the temporal evolution of atmospheric lead deposition in Scotland and eastern Canada using blanket peat bogs. AB - The temporal evolution of atmospheric lead deposition and its possible sources were assessed in eastern Canada and in western Scotland, using blanket peat bogs as geochemical archives. Short cores were taken from two remote sites located close to the sea. Significant lead enrichments in the upper layers at both sites reflect the increasing emission of lead into the atmosphere due to anthropogenic activities during the last century. At the Scottish site, a region under aeolian influence from Europe, anthropogenic derived lead could be recognized by the distinctive unradiogenic composition (206Pb/207Pb ratios down to approximately 1.115), being clearly different from the pre-industrial values (206Pb/207Pb approximately 1.166). In contrast, the lead pollution in eastern Canada (influenced by North American sources) is identified by a more radiogenic lead isotope composition (206Pb/207Pb ratios up to approximately 1.199) compared to preindustrial values (206Pb/207Pb approximately 1.161). Emission inventories and isotope characteristics suggest that industrial (coal burning, mining) and traffic (leaded gasoline) outputs are the most likely sources during the first and the second half of the 20th century, respectively, in both, western Scotland and eastern Canada alike. The Scottish record is in line with previous studies of past atmospheric lead deposition. However, the Canadian deposit suggests that the wind derived, pre-industrial lead, is less radiogenic as previously implied using sediment archives. These results are thus the first to report pre-industrial lead isotope ratios and concentrations of atmospheric derived aerosols in North America. PMID- 12108448 TI - Enrichment of Cu, Ni, Zn, Pb and As in an ombrotrophic peat bog near a Cu-Ni smelter in southwest Finland. AB - The accumulation of selected trace elements (Cu, Ni, Zn, Pb, As) in the surface peat layer of an ombrotrophic bog 2.4 km from a Cu-Ni smelter at Harjavalta, Finland was studied using a peat core. A reference core was taken from an ombrotrophic bog at a background site, Hietajarvi, in eastern Finland. Element concentrations were analysed from 1-cm slices and enrichment factors (EF) were calculated. The enrichment factors of both Cu and Ni in the Harjavalta peat bog are extremely high compared to the Hietajarvi site. However, only the 6-cm surface peat Pb values are higher in Harjavalta compared to Hietajarvi. Precipitation was collected during 1992-1996, in the vicinity of the Harjavalta smelter, in order to estimate the current atmospheric deposition load. Comparison between the precipitation and peat data reveals that at Harjavalta the surface peat is relatively much more polluted than the current precipitation. The variation in EF of the Harjavalta peat core with respect to depth shows two patterns: Cu and Pb are similar, as are Ni, Zn and As. The vertical gradient in Harjavalta Cu EF suggests that Cu supplied to the peat by atmospheric deposition is very well preserved by the bog. PMID- 12108449 TI - Phasing out cadmium and lead--emissions and sediment loads in an urban area. AB - This paper examines how fluxes in the aquatic environment reflect the reduced use of cadmium (Cd) and lead (Pb) in Stockholm, Sweden, between 1975 and 1995. The sediment deposition of Cd and Pb in the waters around Stockholm was investigated using laminated sediment cores, which facilitated reconstructions of historical annual metal deposition to the sediments. The resulting reconstructions were compared to independent estimations of the emissions to the aquatic environment during the phase-out period. The loads of Cd and Pb from sewage treatment plants, storm water and in atmospheric deposition were studied using literature data. The data set indicates a reduced load of Cd and Pb on the aquatic surroundings of Stockholm. The reduction is, however, not as pronounced in the sediment deposition as in the calculated emissions. This indicates that emissions may be delayed on their way to the sediments or that there are other sources, e.g. resuspension of older sediments. It is therefore argued that sediment investigations are an essential component in environmental monitoring, in order to get a complete picture of the metal fluxes to and in the environment in urban areas. PMID- 12108450 TI - Practical approaches to long oligonucleotide-based DNA microarray: lessons from herpesviruses. PMID- 12108451 TI - The major histopathologic pattern of pulmonary fibrosis in scleroderma is nonspecific interstitial pneumonia. AB - BACKGROUND: The prognosis of pulmonary fibrosis associated with scleroderma (PF SSc) has been reported to be significantly better than that of IPF. Because the nonspecific interstitial pneumonia-pattern (NSIP), a newly defined subgroup of idiopathic interstitial pneumonias (IIP), has better prognosis than the usual interstitial pneumonia pattern (UIP), we postulated that NSIP may occur more frequently than UIP in patients with scleroderma who develop fibrosis. METHOD: We reviewed the pathologic, radiologic and clinical outcomes in 19 patients with PF SSc. Two pulmonary pathologists reclassified the histopathology of surgical lung biopsies (SLBx) and consensus diagnosis was achieved in all patients. RESULTS: Thirteen patients had NSIP, five had UIP, and remained one showed only nondiagnostic honeycombing. No significant difference was noted in the initial pulmonary function test (PFT), bronchoalveolar lavage (BAL) findings, or other clinical parameters between UIP and NSIP groups. Comparison of the clinical outcome of 12 patients who were followed for more than 12 months (mean: 34.5 +/- 26.0 months) suggested a better prognosis for NSIP than UIP. Five of the eight NSIP patients improved and three were stable, whereas in patients with UIP, one worsened and three were stable. CONCLUSION: NSIP seems to be the major histopathologic pattern in patients with PF-SSc. PMID- 12108452 TI - [Special issue: the prospect of education activities for space life sciences]. PMID- 12108453 TI - IOM says: X-ray mammography remains the gold standard in breast cancer screening technology. PMID- 12108454 TI - Draft CDC environmental infection control guidelines released. PMID- 12108455 TI - Emergency department pediatric guidelines on absolute necessities. PMID- 12108456 TI - How-to manual gives nuts and bolts of trauma center performance improvement. PMID- 12108457 TI - From poison gas to wonder drug. PMID- 12108458 TI - The 'sheer thrill of tackling problems of human disease'. PMID- 12108459 TI - Clinical governance and risk management revisited. PMID- 12108460 TI - The pleasure of clinical practice: job satisfaction and the infrastructure. PMID- 12108461 TI - Careers in academic medicine: the clinician scientist scheme. PMID- 12108462 TI - Professional attitudes: why we should care. PMID- 12108463 TI - Flexibility and diversity in consultant careers. PMID- 12108464 TI - Multisystem diseases and the kidney. PMID- 12108465 TI - Cardiovascular risk factors in progressive renal disease. PMID- 12108466 TI - Cardiorenal failure: pathophysiology, recognition and treatment. PMID- 12108467 TI - Epidemiology of end-stage renal disease. PMID- 12108468 TI - Common electrolyte problems. PMID- 12108469 TI - A small earthquake? AB - In recent years, concerns about health and healthcare have been expressed by the public, the profession, patients and politicians. These are neither new nor confined to the UK. Doctors were not always held in high public esteem; that had been earned over the last 150 years through scientific discoveries such as anaesthesia which revolutionised patient care. The turbulence of the last few years has a number of causes, including increasing patient expectations. Perhaps inevitably it has again called into question the relationship between government, the public and the profession. The resulting debate has been more widespread and better informed than previous episodes of dissent, perhaps indicating a greater willingness on the part of all three parties to assess their relationship anew. Any such assessment would no doubt have to accept the current workforce difficulties experienced by many doctors and other healthcare professionals practising in unsatisfactory circumstances. PMID- 12108470 TI - Professional attitudes: can they be taught and assessed in medical education? AB - The medical profession is under increasing scrutiny with regard to the undesirable attitudes and behaviours of some of its members. Despite the setting of objectives for professional attitudes, it remains unclear how these can be taught and assessed. Having defined 'attitudes', we consider some of the influences upon the development of professional attitudes within medicine. We then review possible ways of encouraging desirable attitudes and behaviours. Finally, we review and critique the main types of attitude assessment. We conclude that attitudes are complex, that the influence of medical culture is crucial, and that feasible assessment tools have yet to be developed. PMID- 12108471 TI - The European Union: open to the practising physician? AB - The European Union (EU) has powers and responsibilities in the area of public health, while the organisation of healthcare facilities remains the concern of member states. However, a series of directives has, since the late 1970s, sought to ensure the right of individual practitioners to travel freely throughout the EU, both to obtain specialist training and to practise in all member states on the same basis as doctors working in the member states where they were first registered. More recently, there has been mutual recognition of completion of specialist training across the EU. A new directive concerning the mobility of medical practitioners across the EU is under discussion. PMID- 12108472 TI - The expert patient: a new approach to chronic disease management for the twenty first century. AB - The expert patient: a new approach to chronic disease management for the twenty first century, produced by the Department of Health, recommends the introduction of 'user-led self management' for chronic diseases to all areas of the NHS by 2007. The premise is that many patients are expert in managing their disease, and this could be used to encourage others to become 'key decision makers in the treatment process'. Furthermore, these expert patients could 'contribute their skills and insights for the further improvement of services'. It is hypothesised that self-management programmes could reduce the severity of symptoms and improve confidence, resourcefulness and self-efficacy. It is stressed that this is more than just patient education to improve compliance. Instead there should be 'a cultural change...so that user-led self management can be fully valued and understood by healthcare professionals'. I point out that these ideas, while welcome, are not particularly new. Achieving the desired culture change will not be easy. PMID- 12108473 TI - National clinical guidelines for stroke: a concise update. AB - The National Clinical Guidelines for Stroke (1) cover the management of stroke from the acute illness through to transfer of care from hospital to the community, to longer-term problems including carer support and secondary prevention. They are designed to be read by all health and social service professionals, including those working in primary care. Since the guidelines were first published there have been some major developments in stroke research. These have now been incorporated into an updated supplement to the guidelines (2). The new updates include: The recommendation that specialist stroke services should include a neurovascular clinic to enable patients with transient ischaemic attack (TIA) and minor stroke, (where the patient has not been admitted to hospital), to be investigated and treated within a maximum of two weeks. Changes in the recommendations about the management of blood pressure after stroke following the publication of the HOPE and PROGRESS trials. Although advances in therapy research do not warrant radical alterations to practice, two changes have been made. These recommend the use of resisted exercise to improve motor function in targeted muscles and that patients should be given as much opportunity to practice tasks as possible. More precise recommendations on the management of depression. The withdrawal of some recommendations concerning the management of shoulder pain, deep venous thromboses and biofeedback. With the research evidence evolving at a rapid rate a new version of the complete guidelines will be published in 2003. PMID- 12108475 TI - To stand on one's own legs. AB - A fundamental human expectation is to stand upright. This exposes the cardiovascular system to gravitational forces, with a fall in pressure above heart level exposing organs such as the brain to impaired perfusion if adequate adaptive mechanisms are not activated. The autonomic nervous system plays an important role in the initial response to standing upright, and can be affected by several disorders, some rare, some common. Autonomic failure can result in orthostatic hypotension with hypoperfusion of vital organs, causing a variety of symptoms including syncope. Thus, it is important to recognise orthostatic hypotension, determine its aetiology, evaluate and treat it. Intermittent autonomic dysfunction (such as neurally mediated syncope without chronic neurogenic failure) also results in falls and syncope; various forms include the 'common faint' (vasovagal syncope) and carotid sinus hypersensitivity (especially in the elderly). Orthostatic intolerance without orthostatic hypotension is increasingly recognised as due to an autonomic disturbance. New techniques are helping to unravel the functional anatomy of cerebral autonomic centres and their pathways in the causation of orthostatic intolerance. PMID- 12108474 TI - Confidential reporting: from aviation to clinical medicine. AB - The spectacular nature of accidents involving aircraft has obliged the aviation industry to introduce a process of confidential reporting to identify situations that may lead to poor work or even an accident. The information so gathered can then be used to reduce the likelihood of such problems occurring. Confidential reporting allows an individual to bring information relevant to safety to the attention of a body that is completely independent of management. They can be assured that their identity will remain confidential to that body. It is, as far as the individual is concerned, a confidential but not an anonymous procedure, which will be subjected to careful scrutiny. Confidentiality and independence from management are the essential features. The introduction of such a programme into the NHS would materially enhance its safety profile. PMID- 12108476 TI - Teaching the humanities to medical students. AB - The decoding of the human genome offers great promise for the understanding and treatment of chronic human diseases at the last frontier. There is a widely recognised hazard that an exaggerated emphasis on molecular reductionism may lead to the loss of the essential humanitarian instincts of young doctors. To counteract this danger it is now accepted by many leading figures of the medical establishment that the undergraduate curriculum must evolve to incorporate a variety of subjects conventionally taught in the faculty of humanities at our great universities. In this article, the case is argued that the study of 'medical humanities' will enhance the empathy, communication skills, ethical standing and, paradoxically, the scientific literacy of the next generation of young doctors. As a clinical scientist, I cannot prove these assertions with an evidence base, but offer up arguments as qualitative or hypothesis generation. PMID- 12108477 TI - To PEG or not to PEG. AB - Patients with adequate intestinal function who are unable to eat may benefit from enteral tube feeding. Percutaneous endoscopic gastrostomy (PEG) is preferred when prolonged treatment is envisaged. PEG feeding will reduce morbidity and mortality in many such patients by reversing malnutrition. The increasing numbers of elderly patients with chronic diseases have resulted in an increased demand for PEG placement that has stretched resources. Many patients who are referred for PEGs are frail and the procedure is associated with complications. Careful management and support for the carers in the community are essential. Not all patients benefit from PEG feeding. The aim must be an improvement in the quality of life, not a prolongation of terminal disease. PMID- 12108478 TI - Science and medicine 2001. PMID- 12108479 TI - Medicine and ageing--an agenda for progress. PMID- 12108480 TI - Lessons from nostalgia. PMID- 12108481 TI - Norman Rockwell visits a family doctor. PMID- 12108482 TI - Informed consent in medical research. AB - That people should only be enrolled in medical research if they have given free and informed consent is now an unquestioned principle of research ethics. It is however a recent innovation. Prior to the prominence given to consent to participation in research in the condemnation of German doctors arraigned at Nuremberg in 1945, informed consent had appeared in American litigation, but only as an issue in clinical malpractice suits. Informed consent as an ethical requirement in medical research had arisen in some earlier European contexts. Despite the Nuremberg judgement, informed consent by participants in research was not widely recognised as ethically mandatory until the early 1970s. This delay seems to have been due in part to scepticism about the practicability of truly informed consent, but medical paternalism and the circumstances surrounding military research during the Cold War period may have contributed. PMID- 12108483 TI - The consultation in art: Henry Tonks. PMID- 12108484 TI - Dying with dignity. PMID- 12108485 TI - Consultant nurses. PMID- 12108486 TI - NHS direct: growing awareness and use. PMID- 12108487 TI - Troponin I as a risk stratification tool in the district general hospital. PMID- 12108488 TI - Patterns of childbearing amongst female hospital doctors and GPs. PMID- 12108489 TI - The elderly with unrecognised chronic disease: do they provide clues to good health? PMID- 12108490 TI - Characterization of paroxysmal and persistent atrial fibrillation in the human left atrium during initiation and sustained episodes. AB - INTRODUCTION: Atrial fibrillation (AF) in the left atrium (LA) is poorly defined in terms of regional differences in the degree of organization, characteristics of paroxysmal and persistent variants, and electrophysiologic events that develop at the onset of episodes. METHODS AND RESULTS: The study population consisted of 21 patients (15 men and 6 women; mean age 58+/-9.4 years) with paroxysmal (10 patients) or persistent (11 patients) AF. Mapping of the LA during sustained episodes and the onset of AF was performed with a 64-electrode basket catheter. At the onset of AF, repetitive beats starting with atrial premature complexes and ending with generation of the earliest fibrillatory activity were defined as intermediary rhythm. Patients with paroxysmal AF had longer AF cycle lengths and more pronounced regional differences than patients with persistent AF. In total, AF cycle lengths in the LA in patients with persistent AF were 20% shorter than in patients with paroxysmal AF. Initiation of AF was preceded by an intermediary rhythm of 5.5+/-2.5 cycles (6.3+/-2.7 cycles in paroxysmal AF vs 4.2+/-1.0 cycles in persistent AF; P = 0.026). At the onset of AF, the earliest generators of fibrillatory activity were located more frequently in the posterior wall of the LA. CONCLUSION: AF in the LA displays substantial regional differences in terms of AF cycle lengths and degree of organization. Patients with persistent AF have shorter cycle lengths and a higher degree of disorganized activity than patients with paroxysmal AF. Intermediary rhythms play an important role in initiation of AF via activation of generator regions in the LA. PMID- 12108491 TI - About electrograms and their structural origin: clues to more effective ablation strategies for atrial fibrillation. PMID- 12108492 TI - Quantitative assessment of the recovery property of atriofascicular/atrioventricular-type Mahaim fiber. AB - INTRODUCTION: One of the characteristics of the Mahaim fiber is that it possesses a decremental property related to the slow rate of recovery of its excitability similar to that of AV node. The aim of this study was to evaluate the recovery property of the atriofascicular/atrioventricular-type Mahaim fiber and compare it with that of the AV node. METHODS AND RESULTS: Nine patients with a Mahaim fiber were studied; 8 of the patients had atriofascicular/atrioventricular-type fiber. Different models were used to analyze the relationship between conduction time for the Mahaim fiber and the corresponding coupling intervals. The simplest model with the best fit was found to be the linear regression of the natural log of conduction time on corrected coupling intervals. The individual R2 values ranged between 0.43 and 0.98 for the Mahaim fiber and between 0.79 and 0.98 for AV node. The final model chosen for the log transformed data for the Mahaim fiber and for the AV node was the line with parameter estimates defined as a weighted average, over patients, of the corresponding individual line parameters. The weight used for each parameter was the inverse of its variance. The slopes of the lines of the transformed data were not significantly different between the Mahaim fiber and the AV node. Thus, the best fitting curve for the recovery property of the AF type Mahaim fiber is a simple exponential curve similar to that of the AV node. CONCLUSION: The atriofascicular/atrioventricular-type Mahaim fiber has a quantitative recovery property very similar to that of the AV node. PMID- 12108493 TI - Atrial fibrillation after DDDR pacemaker implantation. AB - INTRODUCTION: Newer implantable pulse generators have data storage capabilities that permit detection of multiple episodes of atrial fibrillation (AF). This study evaluated the clinical predictors and time course of AF development in a general pacemaker population. METHODS AND RESULTS: Patients (n = 231) received DDDR pacemakers with features that permit detection and storage of information about the date, time of onset, and duration of multiple, sequential episodes of AF. Patients were followed for 718+/-383 days. Time to first occurrence of AF, interval between first and second episode of AF, and total AF burden were determined at each follow-up visit. AF occurred more often in patients (68%) with sinus node disease than in patients with AV block (37%; P < 0.001). Time to first occurrence of AF was 21.2 days (95% confidence interval [CI] 14.7 to 30.6 days) after pacemaker implantation. AF burden initially decreased significantly in patients (0.8 hours/day, 95% CI 0.7 to 0.9 at 8 weeks after implant vs 0.6 hours/day, 95% CI 0.4 to 0.8 at 12 months after implant; P = 0.005) but then increased significantly during long-term follow-up (2.0 hours/day, 95% CI 1.0 to 3.7 at 48 months after implant; P = 0.008). The long-term increase in AF burden was seen predominantly in patients with sinus node disease. A prior history of AF and the duration of follow-up were independent predictors of AF occurrence. CONCLUSION: AF develops frequently after dual-chamber pacemaker implantation. AF burden increases progressively over the long term. PMID- 12108494 TI - Atrial fibrillation detected by automatic mode switching: from fool's gold toward a gold standard. PMID- 12108495 TI - How to diagnose, locate, and ablate coronary cusp ventricular tachycardia. AB - INTRODUCTION: Although radiofrequency energy usually is applied to the most favorable endocardial site in patients with outflow tract ventricular tachycardia, there are still some patients in whom the tachycardia can be ablated only from an epicardial site. We established the characteristics and technique of catheter ablation from both the left and right coronary cusps to cure left ventricular outflow tract ventricular tachycardia. METHODS AND RESULTS: We studied 15 patients in whom VT was thought to originate from the coronary cusp by both activation and pace mapping after precise mapping of the right ventricle, left ventricle, pulmonary artery, coronary cusps, and anterior interventricular vein. Twelve-lead ECG analysis revealed an S wave on lead I, tall R wave on leads II, III, and aVF, and no S wave on either lead V5 or V6. Precordial R wave transition occurred on leads V1 and V2. The earliest ventricular electrogram at a successful ablation site was recorded 35+/-12 msec before QRS onset and 19+/-15 msec earlier than the earliest ventricular electrogram recorded from the anterior interventricular vein. Almost identical pace mappings were obtained from the coronary cusp. Catheter tip temperature was maintained at 55 degrees C during energy delivery, and the distance from the tip to the ostium of each left and right coronary artery was > 1.0 cm by coronary angiography. CONCLUSION: Left ventricular outflow tract VT that could not be ablated from an endocardial site could be safely eliminated by radiofrequency application to the left and right coronary cusps. PMID- 12108496 TI - Impaired negative chronotropic response to adenosine in patients with inappropriate sinus tachycardia. AB - INTRODUCTION: Adenosine is an endogenous nucleoside that has an important role in the diagnosis and treatment of several cardiac arrhythmias. However, its effects on inappropriate sinus tachycardia (IST) are not well established. METHODS AND RESULTS: In this study, the response to intravenous adenosine (0.1 to 0.15 mg/kg) was studied in 18 patients (age 46+/-15 years) with IST. In a subset of patients (n = 5), the direct effects of adenosine were assessed during pharmacologic beta adrenergic and cholinergic blockade. Atrial cycle length (ACL) was measured before adenosine injection, at the time of the greatest cycle length prolongation, and during the maximum rebound acceleration of heart rate. Eighteen subjects (age 46+/-11 years) with normal sinus rhythm undergoing clinically indicated electrophysiologic study served as controls. Adenosine did not terminate IST in any patient. The maximum dose of adenosine prolonged the sinus interval significantly, from 780+/-128 msec to 985+/-225 msec (P < 0.001) in the control subjects. In contrast, adenosine caused no significant lengthening of atrial cycle length (527+/-69 msec vs 590+/-148 msec; P = NS) in the patients with IST. Similar difference in the response to adenosine was seen during the pharmacologic autonomic blockade. The reflex increase of the sinus rate (rebound effect) was greater in the control subjects than in the patients with IST (21.2%+/-9.7% vs 8.5%+/-8.8%; P < 0.001). CONCLUSION: The usual negative chronotropic effect of adenosine was impaired in the patients with IST. This may have important diagnostic implications and provide new insight into the mechanism(s) of IST. PMID- 12108497 TI - Outcome of women versus men with ventricular tachyarrhythmias treated with the implantable cardioverter defibrillator. AB - INTRODUCTION: Important sex differences in the incidence and outcome of patients with ischemic heart disease, the leading cause of ventricular tachyarrhythmias, have been identified. Implantable cardioverter defibrillator (ICD) therapy has become the treatment of choice for patients with ventricular tachycardia (VT) and ventricular fibrillation (VF), but little is known about gender differences in the outcome of ICD-treated patients. METHODS AND RESULTS: In this retrospective study, we compared arrhythmic events and survival of 376 women and 1,654 men treated with an ICD as part of prospective evaluations of transvenous devices or lead systems. Women were younger (62+/-14 years vs 65+/-12 years, P = 0.0005), had higher left ventricular ejection fraction (0.36+/-0.15 vs 0.32+/-0.13, P < 0.0001), were more likely to present with VF (34% vs 19%, P < 0.001), and had lower implantation defibrillation threshold (11+/-6 vs 13+/-6 J, P < 0.0001). Implant complication rates were similar in men and women (2.6% vs 3.5%, P = 0.46). The 1-year and 2-year cumulative rates of appropriate ICD therapies were 31.4% and 38.4% for men and 32.6% and 40.8% for women, respectively (P = 0.63). The unadjusted 1-year and 2-year cumulative survival rates were 95.6% and 93.7% for men and 95.7% and 94.3% for women, respectively (P = 0.98). Adjusted total (P = 0.61), sudden (P = 0.82), and cardiac (P = 0.34) death-free survivals also were similar in men and women. CONCLUSION: Despite clinical differences suggesting women are at lower risk than men, men and women with VT/VF who are treated with an ICD have similar arrhythmic event and survival rates. These factors should be considered when determining risk and prescription of ICD therapy for women. PMID- 12108498 TI - Sex, shocks, and survival: sudden cardiac death prevention with implantable cardioverter defibrillators in women versus men. PMID- 12108499 TI - Simultaneous biatrial computerized mapping during permanent atrial fibrillation in patients with organic heart disease. AB - INTRODUCTION: Activation patterns during permanent atrial fibrillation (AF) in patients with organic heart diseases are unclear. METHODS AND RESULTS: We studied six patients with permanent AF and organic heart diseases undergoing surgery. The duration of AF averaged 4.9+/-7.6 years. Computerized epicardial mappings of the right atrial (RA) free wall and the left atrial (LA) posterior wall were simultaneously performed with 224 bipolar electrodes at 3-mm spatial resolution. In the RA, large wavefronts and conduction blocks were frequently observed. The lines of block correlated with the crista terminalis and large pectinate muscles. In contrast, the LA had rapid repetitive activities originated from corners of the electrode plaque, near the four pulmonary veins (PVs). On average, 2.8+/-1.2 sites of rapid repetitive activities were identified per patient. They activated continuously, intermittently, or alternately during AF. The mean activation cycle length in the RA (196+/-22 msec) was significantly longer than that in the LA (179+/-26 msec; P = 0.004). The maximum dominant frequency in the LA was higher than that in the RA (6.41+/-1.18 Hz vs 5.66+/-0.55 Hz; P = 0.049). The maximum dominant frequency was consistently located in areas with rapid repetitive activations near the PVs. CONCLUSION: During human permanent AF associated with organic heart diseases, the activation cycle length was shorter in the LA posterior wall than in the RA free wall. Rapid repetitive activities are consistently observed in the LA posterior wall, at or near the PVs. PMID- 12108500 TI - Mechanism(s) of chronic atrial fibrillation. PMID- 12108502 TI - Short-term rapid atrial pacing produces electrical remodeling of sinus node function in humans. AB - INTRODUCTION: Depression of sinus node function occurs in dogs and in patients after cessation of atrial flutter and fibrillation. We tested whether transient atrial pacing might produce similar changes in humans. METHODS AND RESULTS: We studied the impact of short-term rapid atrial pacing, simulating atrial tachyarrhythmias, on sinoatrial conduction time (SACT) and corrected sinus node recovery time (CS-NRT) in 10 patients undergoing electrophysiologic study. None had recognizable structural heart disease, history of atrial fibrillation or flutter, autonomic dysfunction, or any tachycardia for at least 24 hours before study. All cardiac drugs were discontinued >5 half-lives prior to study. No patient had significant hypotension during atrial stimulation. SACT and CSNRT were measured at baseline, and sinus node reset zone was determined. Right atrial pacing was performed for 10 to 15 minutes, after which SACT and CSNRT were measured again. Both parameters increased significantly, from 423+/-208 msec to 491+/-214 msec and from 80+/-50 msec to 96+/-53 msec, respectively (P = 0.02 and P < 0.001, respectively). CONCLUSION: Rapid atrial pacing for only 10 to 15 minutes, simulating transient atrial tachyarrhythmias, alters sinus node function in humans. Additional studies are needed to evaluate the mechanism, but the clinical implication is that even transient episodes of atrial tachyarrhythmias can cause sinus node remodeling in patients. PMID- 12108501 TI - Survival of antiarrhythmic or implantable cardioverter defibrillator treated patients with varying degrees of left ventricular dysfunction who survived malignant ventricular arrhythmias. AB - INTRODUCTION: The relative effectiveness of the implantable cardioverter defibrillator (ICD) and antiarrhythmic drugs (AADs) varies with left ventricular ejection fraction (LVEF). However, once an ICD or AAD treatment strategy is chosen, the degree to which the LVEF influences survival is unknown. This article addresses that question. METHODS AND RESULTS: Using patient data from the Antiarrhythmics Versus Implantable Defibrillators (AVID) trial, the impact of LVEF on prognosis of patients who were treated with either an ICD or AAD was assessed. Survival within each quintile of LVEF was estimated by the method of Kaplan-Meier for patients treated with either the ICD or AADs. The Cox proportional hazards model was used to investigate the prognostic value of LVEF for estimating survival after adjustment for other baseline covariates among all patients in the subgroups treated by ICD or AAD. In the highest two quintiles of LVEF, survival was comparable in AAD-treated and ICD-treated patients. In the AAD treated patients, higher LVEF was significantly and independently associated with survival free of all-cause mortality and arrhythmic death. In the ICD-treated patients, however, the statistical significance of the association was lost and only a trend toward greater survival was present. Death due to congestive heart failure remained independently and significantly associated with survival in both AAD-treated and ICD-treated patients. CONCLUSION: In patients treated with AADs but not patients treated with ICDs, survival is strongly associated with LVEF. The absence of a statistically significant association in the ICD patients is likely related to the effectiveness of the ICD in treating malignant ventricular arrhythmias, but a chance lack of association cannot be excluded. PMID- 12108503 TI - Beta-adrenergic stimulation of pig myocytes with decreased cytosolic free magnesium prolongs the action potential and enhances triggered activity. AB - INTRODUCTION: Heart failure results in chronic beta-adrenergic stimulation, repolarization lability, and arrhythmias associated with early afterdepolarizations (EADs) and delayed afterdepolarizations (DADs). Having described a significant reduction in intracellular free magnesium ([Mg2+]i) in experimental heart failure, we asked whether a reduction in [Mg2+]i would delay repolarization or facilitate EADs and/or DADs. METHODS AND RESULTS: Left ventricular myocytes were isolated from Yorkshire swine. Cytosolic free [Mg2+] was set at 0.12 mM (LoMg) or 1.2 mM (HiMg) through pipette dialysis. Action potentials (AP), Ca current (I(Ca)), and sodium/calcium exchange current (I(NCX)) were measured in the presence or absence of isoproterenol (2 microM) at 37 degrees C. Under basal conditions (0.1-Hz stimulation, 2 mM external [Ca2+]), reducing [Mg2+]i had no effect on AP duration and I(Ca) but did significantly enhance I(NCX). In contrast, during superfusion with isoproterenol, reduced [Mg2+]i caused a significant increase in AP duration at both 50% and 90% repolarization (APD50 and APD90) compared with HiMg (P < 0.05). LoMg cells manifested a high incidence of triggered activities, including spontaneous AP, EADs, and DADs (83.3% in LoMg, n = 12 vs 38.3% in HiMg, n = 13; P < 0.05). I(Ca) and I(NCX) were significantly increased in LoMg cells compared with HiMg cells (P < 0.05). CONCLUSION: Decreased cytosolic free magnesium prolongs AP duration and increases the incidence of triggered activity during beta-adrenergic stimulation. These effects may be due to increased I(Ca) and I(NCX) in the presence of reduced intracellular [Mg2+]. A magnesium-dependent increase in triggered activity coupled with delayed repolarization during beta-adrenergic stimulation could contribute to the arrhythmogenic substrate in heart failure. PMID- 12108504 TI - Microvolt T wave alternans inducibility in normal newborn puppies: effects of development. AB - INTRODUCTION: The cause of sudden infant death syndrome is unknown, but increased cardiac vulnerability due to repolarization instability may be a contributing factor. The QT interval normally is long at birth and increases further during the first few postnatal months. Although excessive QT intervals indicate increased cardiac vulnerability in the long QT syndrome, the impact of less pronounced QT prolongation during this developmental period is unclear. In adults and older children, the ease of inducing microvolt-level T wave alternans (TWA) is used as a measure of repolarization instability and arrhythmia vulnerability. The aim of this study was to determine if TWA is inducible in normal newborn puppies. METHODS AND RESULTS: Atrial pacing was performed in 15 anesthetized beagle puppies 7 to 35 days old. The pacing drive cycle length was systematically decreased in 20-msec steps from baseline until AV conduction blocked. Pacing was performed for 8 minutes at each cycle length. Three-lead ECGs were recorded continuously during the last 5 minutes of pacing at each cycle length. The recordings were analyzed off-line for the presence of microvolt-level TWA using a sensitive spectral analysis technique. Microvolt-level TWA was present in all puppies. TWA was not present at baseline but developed and increased in amplitude as heart rate increased. The threshold heart rate for TWA did not correlate with age. However, due to age-dependent changes in baseline heart rate, the 7- to 14 day-old animals needed a 50% to 78% increase in heart rate to reach threshold heart rate, whereas the oldest animals needed only a 5% to 25% increase. CONCLUSION: These data suggest that developmentally dependent dynamic repolarization instability exists in puppies as manifest by the inducibility of TWA. PMID- 12108505 TI - Arrhythmogenesis of T wave alternans associated with surface QRS complex alternans and the role of ventricular prematurity: observations from a canine model of LQT3 syndrome. AB - INTRODUCTION: T wave alternans (TWA) is characterized by cycle-to-cycle changes in the QT interval and/or T wave morphology. It is believed to amplify the underlying dispersion of ventricular repolarization. The aim of this study was to examine the mechanisms and arrhythmogenesis of TWA accompanied by QRS complex and/or blood pressure (BP) waveform alternans, using transmural ventricular electrogram recordings in an anthopleurin-A model of long QT syndrome. METHODS AND RESULTS: The cardiac cycle length was gradually shortened by interruption of vagal stimulation, and TWA was induced in six canine hearts. Transmural unipolar electrograms were recorded with plunge needle electrodes from endocardial (Endo), mid-myocardial (Mid), and epicardial (Epi) sites, along with the surface ECG and BP. The activation-recovery interval (ARI) was measured to estimate local refractoriness. During TWA, ARI alternans was greater at the Mid than the Epi/Endo sites, and it was associated with the development of marked spatial dispersion of ventricular repolarization. As TWA increased, ventricular activation of the cycles associated with shorter QT intervals displayed delayed conduction at the Mid sites as a result of a critically longer ARI of the preceding cycle and longer QT interval, while normal conduction was preserved at the Epi site. Delayed conduction at the Mid sites manifested as surface ECG QRS and BP waveform alternans, and spontaneous ventricular tachyarrhythmias developed in absence of ventricular prematurity. In other instances, in absence of delayed conduction during TWA, ventricular premature complexes infringed on a prominent spatial dispersion of ventricular repolarization of cycles with long QT intervals and initiated ventricular tachyarrhythmia. CONCLUSION: TWA accompanied by QRS alternans may signal a greater ventricular electrical instability, since it is associated with intramural delayed conduction, which can initiate ventricular tachyarrhythmia without ventricular premature complexes. PMID- 12108507 TI - Complete elimination of incessant polymorphic ventricular tachycardia in an infant with MIDAS syndrome: use of endocardial mapping and radiofrequency catheter ablation. AB - Experience with radiofrequency catheter ablation of ventricular tachycardia (VT) in pediatric patients is limited. A 5-month-old white female infant (body weight 5.5 kg) with MIDAS syndrome who suffered from incessant polymorphic VT (ventricular rates 250 to 300 beats/min) and was unresponsive to medical treatment resulting in significantly depressed left ventricular (LV) function underwent a total of four catheter ablation procedures during a 5-month period. Each of the procedures reduced the number of morphologies and the rate of the tachycardia, but VT returned after each of the first three procedures, despite concomitant medical therapy. Activation mapping and pace mapping were used to identify the anatomic substrates, which were found at different locations at the LV septum and LV free wall. All forms of VT finally were ablated successfully. There were no significant complications. After the fourth procedure, the patient was in continuous sinus rhythm. Follow-up examination 29 months after the last procedure while the child was not taking any medication showed normal sinus rhythm and normal LV function. This report demonstrates the usefulness and safety of radiofrequency catheter ablation in an infant with polymorphic VT who was unresponsive to medical therapy. PMID- 12108506 TI - Intrapericardial therapeutics: a pharmacodynamic and pharmacokinetic comparison between pericardial and intravenous procainamide delivery. AB - INTRODUCTION: Procainamide delivery into the pericardial space may produce a greater and more prolonged electrophysiologic effect, particularly in thin superficial atrial tissue, compared with intravenous delivery. METHODS AND RESULTS: Swine were randomized to sequential procainamide doses delivered intravenously (n = 6) or into the pericardial space (n = 7). The cumulative pericardial doses were 0.5, 1.5, and 3.5 mg/kg, and the intravenous doses were 2, 10, and 26 mg/kg. Pericardial procainamide prolonged right atrial effective refractory period from baseline by 22% (P < 0.01) but only at the 3.5 mg/kg cumulative dose. This dose slowed interatrial conduction time by 14% (P < 0.05) and raised atrial fibrillation threshold by 70 mA (P < 0.05). Pericardial procainamide had minimal effect on ventricular electrophysiology. Similar results occurred with a single 2 mg/kg pericardial dose in a closed chest model. Intravenous 10 and 26 mg/kg cumulative doses prolonged atrial effective refractory period from baseline by 24% and 18% (P < 0.01), respectively. The 26 mg/kg cumulative intravenous dose slowed interatrial and atrial-ventricular conduction times by 27% and 17%, respectively (P < 0.05), raised atrial fibrillation threshold, and slowed ventricular conduction time by 29% (P < 0.05). Pericardial procainamide produced pericardial fluid concentrations ranging from 250 to 1,500 microg/mL, but plasma concentrations were <1 microg/mL. Intravenous procainamide doses produced pericardial fluid concentrations similar to plasma trough concentrations 0 to 12 microg/mL. CONCLUSION: The single 2 mg/kg and 3.5 mg/kg cumulative pericardial procainamide doses prolonged atrial refractoriness and raised atrial fibrillation threshold similar to the 26 mg/kg cumulative intravenous dose, but the duration of effect was similar between delivery methods. Pericardial procainamide did not affect global or endocardial ventricular electrophysiology nor was it associated with ventricular proarrhythmia. PMID- 12108508 TI - Novel cause of spurious mode switching in dual-chamber pacemakers: atrioventricular desynchronization arrhythmia. AB - Spurious mode switching is a common problem in DDDR pacing and implantable cardioverter defibrillator systems. It may produce symptoms indistinguishable from classic pacemaker syndrome or paroxysmal atrial arrhythmias due to loss of AV synchrony. The dominant cause of spurious mode switching is oversensing of far field R waves. This report describes a novel cause of spurious mode switching, AV desynchronization arrhythmia. Clinical manifestations, recognition, and management are discussed. PMID- 12108509 TI - Ectopic tachycardia originating from the superior vena cava. AB - We report a 65-year-old female patient with a 3-year history of symptomatic paroxysmal supraventricular tachycardia. Electroanatomic and basket catheter mapping revealed a focal tachycardia originating in the superior vena cava (SVC), 5 cm above the SVC-right atrium (SVC-RA) junction. An area of fractionated potentials and slow conduction was found on the anterior wall of the SVC. A line of conduction block extending downwardly and obliquely from the anteroseptal aspect to anterolateral aspect of the SVC forcing the impulse to enter the RA via the posterior aspect of SVC-RA junction was observed. Entrainment attempts from multiple sites within the SVC failed to demonstrate reentry as a mechanism of arrhythmia. The ablation approach consisted of isolation of the arrhythmogenic area from the rest of the SVC. PMID- 12108510 TI - Diversity of connexin expression patterns in the atrioventricular node: vestigial consequence or functional specialization? PMID- 12108511 TI - Widening of the QRS complex during atrial pacing: what is the mechanism? PMID- 12108512 TI - Amiodarone-induced giant T wave alternans hastens proarrhythmic response. PMID- 12108513 TI - Real-time, three-dimensional, nonfluoroscopic localization of the Lasso catheter. PMID- 12108514 TI - Effect of hypoxia on parathyroid hormone in lactating and neonatal rats: interaction with halothane. AB - Low oxygen in the blood (hypoxemia) may occur in the neonate or women in the postpartum period. Administration of inhalation anesthetic may be required in this period. The purpose of this study was to evaluate the effect of 7 d of hypoxia on the neonatal rat pup and lactating dam without or with acute halothane anesthesia on serum calcium and calciotropic hormones. Ionized calcium was not altered by hypoxia or halothane administration. Hypoxia from birth had no effect on serum parathyroid hormone (PTH) in 7-d-old rat pups (48+/-4 pg/mL). Halothane increased PTH in rat pups (74+/-8 pg/mL). The effect of halothane was not augmented in hypoxic pups. Hypoxia for 7 d had no effect on serum PTH in lactating dams (23+/-3 pg/mL). Halothane resulted in an increase in PTH (106+/-17 pg/mL). When halothane was administered to hypoxic lactating dams, a striking increase in serum PTH was observed (401+/-50 pg/mL). We hypothesize that halothane and hypoxia alter parathyroid gland function by a direct effect on cellular calcium dynamics. This interaction may have clinical significance in hypoxic patients requiring general anesthesia. PMID- 12108515 TI - Estradiol and luteinizing hormone regulation of insulin-like growth factor binding protein production by bovine granulosa and thecal cells. AB - To determine the effects of estradiol and luteinizing hormone (LH) on insulin like growth factor-binding protein (IGFBP) production by bovine granulosa and thecal cells, both cell types were collected and cultured in serum-free medium with various hormone treatments, arranged in three experiments. In thecal cells, insulin stimulated (p < 0.05) production of IGFBP-2 and IGFBP-5, but had no effect (p > 0.10) on IGFBP-3 and IGFBP-4 production; LH stimulated (p < 0.05) production of IGFBP-2 and IGFBP-3 but had no effect (p > 0.05) on IGFBP-4 and IGFBP-5. Estradiol had no effect (p > 0.10) on IGFBP-2, IGFBP-3, IGFBP-4, and IGFBP-5 production by thecal cells. Production of IGFBP-2/-5 by granulosa cells from small follicles was inhibited (p < 0.05) by insulin, but estradiol and LH did not influence (p > 0.10) insulin's inhibitory effect on basal IGFBP-2/-5 production. Insulin, LH, and estradiol each inhibited IGFBP-4 production by small follicle granulosa cells, but their effects were not additive. IGFBP-3 was not produced by small-follicle granulosa cells. In large-follicle granulosa cells, insulin and LH inhibited (p < 0.05) production of IGFBP-2/-5 and IGFBP-3, whereas estradiol had no effect. Insulin alone had no effect (p > 0.10) on production of IGFBP-4, but estradiol and LH inhibited (p < 0.05) production by large-follicle granulosa cells, and their effects were not additive. These results suggest that production of IGFBP-2, IGFBP-3, IGFBP-4, and IGFBP-5 by granulosa and thecal cells is differentially affected by hormonal stimuli. PMID- 12108516 TI - Evidence of free leptin in human seminal plasma. AB - Leptin is an adipose tissue-secreted hormone that actively participates in the regulation of energy homeostasis. Besides this principal role, leptin has been implicated in a large variety of neuroendocrine, paracrine, and autocrine actions involved in the regulation of reproductive function in both experimental animals and humans. Although the participation of leptin in female reproduction is well established, any role in male reproductive function is at best tenuous. The aim of this study was to ascertain whether true leptin is present in human seminal fluid and the tissue of its production. Pooled human seminal plasma obtained from healthy donors showed by direct radioimmunoassay (RIA) the presence of radioimmunoassayable leptin. Serial dilutions of unextracted semen paralleled the RIA standard curve, also devoid of interference in the assay. To prove that this activity was true leptin, seminal plasma was subjected to size-exclusion chromatography, which showed that leptin immunoreactivity eluted with the same partition coefficient as cold leptin, 125I-leptin, and 125I-leptin preincubated with seminal plasma. The results demonstrate that true leptin was present in semen in a free form, i.e., without binding proteins. The presence of leptin charge variants in seminal plasma was assessed by anion-exchange chromatography, which showed two peaks of leptin inmunoreactivity, while 125I-leptin eluted as a single peak. Preincubation of 125I-leptin with seminal fluid converted the single peak into a double peak, indicating that components of the seminal fluid introduce a charge variation in leptin. Leptin levels in seminal plasma of 40 healthy men were 0.95+/-0.19 ng/mL while in 5 vasectomized men the levels were 0.92+/-0.25 ng/mL, suggesting that testicular tissues were not the source of seminal leptin. No correlation was observed between leptin concentrations in semen and the physical characteristics of semen samples or physical characteristics of spermatozoids, such as concentration, motility, vitality, or morphology. In conclusion it was unambiguously demonstrated that human leptin is present in seminal fluid, with at least two charge variants and no binding proteins, the most likely source being either seminal vesicles or prostate tissue. The role of seminal fluid leptin in the male reproductive function or sperm capacitation is at present unknown. PMID- 12108518 TI - Glucose intolerance induced by a high-fat/low-carbohydrate diet in rats effects of nonesterified fatty acids. AB - We examined the time course of effects of a high-fat/low-carbohydrate (HF/LC) diet on the impairment of glucose tolerance in rats, clarified whether insulin secretion and sensitivity were impaired by the HF/LC diet, and investigated the relationship between the increased nonesterified fatty acids (NEFA) after HF/LC diet feeding and insulin secretion and sensitivity. We found that glucose tolerance and the postglucose-loading insulin secretion were impaired after 3 and 7 d on the HF/LC diet. The glucose intolerance was accompanied by a rise in the fasting plasma NEFA level. When stimulated with 15 mmol/L of glucose, the insulin secretion was impaired in pancreatic islets from rats fed the HF/LC diet. Rats fed the HF/LC diet showed insulin resistance in vivo. The glucose-stimulated insulin secretion was inhibited in the islets following 24-h culture with palmitic acid. The 24-h infusion of palmitic acid decreased whole-body insulin sensitivity. In summary, at least 3 d on a HF/LC diet is needed to induce glucose intolerance in rats, and the impairment may be induced by decreased insulin secretion and sensitivity, which is related to the increase in the plasma NEFA level. PMID- 12108517 TI - Pulsatile leptin secretion is independent of luteinizing hormone secretion in prepubertal sheep. AB - Many studies have suggested that leptin modulates the gonadal axis. A synchronicity of luteinizing hormone (LH) and leptin has been described in humans, suggesting that leptin may modulate the episodic secretion of LH. The objective of this study was to establish whether episodic leptin secretion depends on the episodic LH secretion in prepubertal sheep. We used two different approaches. The first consisted of blocking the release of LH using a long-acting LH-releasing hormone (LHRH) agonist and analyzing the episodic LH and leptin secretions. The second method stimulated the pituitary gland with pulses of LHRH and again LH and leptin secretions were analyzed. Spring-born 20-wk-old Suffolk ewe lambs (n = 5) received intramuscularly a long-acting LHRH agonist (Decapeptyl). Treatment was repeated at 24 and 28 wk of age. Control lambs (n = 6) received the vehicle of Decapeptyl. Diurnal and nocturnal pulsatilities of LH and leptin were studied at 20 (before Decapeptyl injection), 26, and 30 wk. Blood samples were taken at 10-min intervals for 6 h, beginning at 10:00 AM (diurnal sampling) and at 10:00 PM (nocturnal sampling). In all samples, LH and leptin were measured by radioimmunoassay, and pulsatile hormone secretion characteristics were assessed by the CLUSTER program. To characterize further the synchronicity between LH and leptin pulses, LHRH (10 ng/kg body wt) was injected at 60-min intervals, six times, to another five 30-wk-old ewe lambs, for the same time period as the pulsatility study. In the control group, LH secretion did not change between lambs of 20 and 30 wk of age. In LHRH agonist-treated lambs, LH secretion diminished from 20 to 30 wk of age and was lower than in control lambs at 26 and 30 wk of age (p < 0.05). The transversal mean (ng/[mL x 6 h]) of leptin concentrations was different between control lambs of 20 wk of age and 26 and 30 wk of age (p < 0.01). Contrary to the findings in LH secretion, in LHRH agonist treated lambs, mean plasma leptin concentrations did not decrease. Furthermore, the mean diurnal and nocturnal leptin concentrations and the pulse amplitude were higher at 26 and 30 wk than at 20 wk in LHRH agonist-treated lambs (p < 0.05). There were no differences between diurnal and nocturnal parameters of leptin secretion in both groups. There was no synchronicity between LH and leptin pulses. LHRH pulses significantly increased plasma LH concentrations, producing discernible LH pulses; however, leptin amplitude and leptin pulse frequency were not modified by the exogenous LHRH pulses, exhibiting no coincidence with LH pulses. The data suggest that pulsatile leptin secretion is independent of LH secretion in ewe lambs. PMID- 12108519 TI - Endogenous excitatory amino acid neurotransmission regulates thyroid-stimulating hormone and thyroid hormone secretion in conscious freely moving male rats. AB - The role of neurotransmission of endogenous excitatory amino acid (EAA) on serum thyroid hormones and thyroid-stimulating hormone (TSH) levels was examined in conscious and freely moving adult male Sprague-Dawley rats. The rats were cannulated at the third ventricle 2 d before the experiments. Several glutamate receptor agonists, such as kainic acid and domoic acid, and antagonists, such as 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) and dizocilpine (MK-801) were administered into the third ventricle. Serum TSH levels were assesed by radioimmunoassay, and serum thyroid hormone levels were assessed by enzyme immunoassay. The results showed that the administration of CNQX and MK-801 produced a decrease in serum levels of TSH and thyroid hormones. The administration of kainic acid and domoic acid increased TSH concentrations, whereas CNQX completely blocked the release of TSH induced by kainic acid and domoic acid. These results suggest the importance of endogenous EAA in the regulation of hormone secretion from the pituitary-thyroid axis, as well as the role of the N-methyl-D-aspartate (NMDA) and non-NMDA receptors in the stimulatory effect of EAAs on the pituitary-thyroid axis. PMID- 12108522 TI - Growth hormone-releasing hormone and gonadotropin-releasing hormone stimulate nitric oxide production in 17beta-estradiol-primed rat anterior pituitary cells. AB - It was reported that neuronal nitric oxide synthase (nNOS) was expressed only in gonadotrophs and folliculo-stellate cells in the anterior lobe of the pituitary gland. However, recent studies have demonstrated the occurrence of nNOS in the somatotrophs and lactotrophs. In the present study, we investigated effects of growth hormone-releasing hormone (GHRH), gonadotropin-releasing hormone (GnRH), and 17beta-estradiol on nitric oxide (NO) release in cultured rat anterior pituitary cells in vitro. The NO2- level in the incubation medium of the rat anterior pituitary cells was dependent on the cell density. Pretreatment with 10 microM 17beta-estradiol resulted in an increase in medium NO2- level. GHRH and GnRH failed to change medium NO2- levels, but they elicited increases in medium NO2- levels in estrogen-treated cells. The GHRH-induced increase in NO2- level was inhibited by Nomega-nitro-L-arginine methyl ester, a NOS inhibitor. These findings suggest that GnRH and GHRH could activate nNOS in the gonadotrophs and the somatotrophs, respectively. PMID- 12108520 TI - Injection of antiangiogenic agents into the macaque preovulatory follicle: disruption of corpus luteum development and function. AB - Ovulation and conversion of the follicle into the corpus luteum involve remarkable changes in vascular permeability and neovascularization of the luteinizing granulosa layer. To evaluate the importance of these vascular events in follicle rupture and luteal development, sequential experiments were designed in which vehicle or angiogenic inhibitors (TNP-470 or angiostatin) were injected directly into the preovulatory follicle of rhesus monkeys during spontaneous menstrual cycles. After control injections, 13 of 14 animals exhibited serum levels of progesterone (P) during the subsequent luteal phase that were comparable to untreated animals in our colony. Following low-dose (400 pg/mL) TNP 470, serum P levels increased normally until d 8 of the luteal phase, but then declined prematurely by d 9 (p < 0.05 compared to controls) and remained below controls until menses. Following high-dose (2 microg/mL) TNP-470, serum P levels were diminished in the early luteal phase (d 3-5; p < 0.05 compared to controls), but reached typical levels at mid luteal phase, only to decline prematurely by d 9 (p < 0.05) and remain low until menses. Control ovaries displayed indices of follicle rupture (protruding stigmata) and luteinization. TNP-470-treated ovaries exhibited signs of distension (torn surface epithelium/tunica albuginea) and luteinization; however, a well-formed stigmata was not observed. A "trapped" oocyte was not observed in serial sections of developing corpora lutea from control or TNP-470-treated animals. However, the early corpus luteum of TNP-470 injected ovaries contained pockets of excessive numbers of blood cells that were absent in controls. Angiostatin did not alter serum P levels or ovarian morphology compared to controls. These data suggest that acute exposure to the antiangiogenic agent TNP-470 impairs the development and functional capacity of the primate corpus luteum in a dose-dependent manner. The results are consistent with a critical role for angiogenesis in cyclic ovarian function in primates. PMID- 12108524 TI - A meal stimulation test in the diagnosis of pancreatic endocrine tumors in multiple endocrine neoplasia type 1. AB - BACKGROUND: The diagnostic value of the determination of the serum pancreatic polypeptide (PP) and gastrin concentrations after a standard meal for early diagnosis of patients with multiple endocrine neoplasia type 1 (MEN 1) is controversial. The aim of this study was to clarify this issue. Thirteen patients with MEN 1, seven healthy family members, and eight healthy controls were studied. Plasma PP and serum gastrin were measured before and after the ingestion of a standardized meal. The meal caused a statistically significant (p < 0.05) increase of both PP and gastrin in all three groups studied. Concerning PP, no statistically significant difference was observed between patients and controls. In family members, the values were significantly (p < 0.05) lower than in the other two groups. On the whole, no significant differences in gastrin levels were noted between patients and controls; in family members, the values were significantly (p < 0.05) lower than in patients. All patients who had abnormally high postprandial values of PP and gastrin also had abnormally high basal values of these two peptides. The determination of serum PP and gastrin levels after a meal stimulation test in patients with MEN 1 adds no information about the presence of pancreatic endocrine tumors over that provided by basal values of the two peptides. PMID- 12108521 TI - Expression of estrogen and progesterone receptors in the bovine ovary during estrous cycle and pregnancy. AB - The objective of the study was to demonstrate the mRNA expression of estrogen receptor a (ERa), ERbeta, and progesterone receptor (PR) by block reverse transcription-polymerase chain reaction(RT-PCR) and real-time RT-PCR (LightCycler) in bovine ovarian follicles and in corpus luteum during the estrous cycle and pregnancy. The mRNA expression of ERalpha and ERbeta mRNA in theca interna tissue (TI) (lower pg/microg RNA) increased continuously and significantly during final growth of follicles, with much higher levels for ERalpha. The mRNA expression of ERalpha and ERbeta in granulosa cells (GC) (fg/microg RNA) increased continuously during follicle growth but without any significant change. The expression of mRNA for PR in follicles (lower fg/microg RNA) increased continuously to maximum level in preovulatory follicles with a significant change only in TI. The highest mRNA expression for ERalpha (fg/microg RNA) was detected in corpus luteum (CL) during the early luteal phase, following by a significant decrease of expression during the mid, late, and regression phases. In contrast, ERbeta mRNA expression is relatively high during the early stage, decreased during the late early and mid luteal phase, and increased significantly again during the late luteal phase and after CL regression. During pregnancy (>3 mo), low levels of ERalpha and ERbeta mRNA expression (<25 fg/microg RNA) with no significant changes were measured. No significant change in PR mRNA expression (levels <13 fg/microg RNA) during the estrous cycle and pregnancy in bovine CL were found. The results suggest an autocrine/paracrine role of steroid receptors in the regulation of final follicle growth and corpus luteum formation and function. PMID- 12108523 TI - Reciprocal regulation by estradiol 17-beta of ezrin and cadherin-catenin complexes in pituitary GH3 cells. AB - The antiestrogen, ICI 182780, and estradiol-17beta (E2) regulate cadherin mediated cell adhesion in pituitary GH3 cells. Using a cDNA expression array to screen for E2-regulated genes that are associated with the cytoskeleton, we observed that E2 stimulated ezrin gene expression and confirmed that ezrin gene expression is regulated pretranslationally by ICI 182780 versus E2. E2 increased ezrin protein levels in whole-cell lysates and in the cytoskeletal-associated, detergent-insoluble fraction. Confocal microscopy revealed that ezrin was associated with free apical membranes of E2-treated cells. E2 decreased N cadherin and beta-catenin levels and induced a redistribution of p120ctn to the cytoplasm. In GH3 transfectants overexpressing E-cadherin, E2 had no effect on adhesiveness or on E-cadherin and p120ctn distribution, but increased levels of active ezrin. Ezrin was concentrated at free and apical membranes. These studies provide the first demonstration of the regulation of ezrin by E2 and show that the ER signaling pathway coordinately regulates two cytoskeletal-associated protein complexes, with mutually exclusive cellular distributions, in a reciprocal manner. These findings indicate that E2 enriches the cell membrane with ezrin-membrane protein complexes by both increasing ezrin expression and by enlarging the relative area of nonadhesive membrane to which ezrin is targeted. PMID- 12108525 TI - Estrogen stimulates expression of p21Waf1/Cip1 in mouse uterine luminal epithelium. AB - p21Waf1/Cip1 was originally identified as an inhibitor of the cell cycle. Recent evidence suggests that it can act as a positive regulator of the cell cycle under the influence of some growth stimulators. We investigated the effects of ovarian steroids on the expression of p21, DNA synthesis, and mitosis in the uterus. Capsules containing 17beta-estradiol (E2) were subcutaneously implanted in ovariectomized mice that were sacrificed on different days. Their uteri were collected for p21 immunohistochemical staining. To study mitosis and DNA synthesis, colchicine and bromodeoxyuridine (BrdU) were injected into mice 3 or 5 h before sacrifice. The results showed that p21 expression, BrdU incorporation, and the mitotic index in uterine luminal epithelium increased 1 to 2 d after E2 stimulation and then declined to basal levels between d 3 and 6. Furthermore, cotreatment with progesterone (P4) and E2 suppressed both p21 expression and the DNA synthesis stimulated by E2 alone in uterine epithelial cells. Our results show that estrogen stimulates p21 expression and cell proliferation in uterine luminal epithelium and that cotreatment with P4 prevents both effects, suggesting that p21 may act as a positive cell-cycle regulator. PMID- 12108527 TI - Changing clinical practice--and perceptions. PMID- 12108526 TI - Isolated neurosarcoidosis--a diagnostic enigma: case report and discussion. AB - Neurosarcoidosis is a rare, but well-recognized cause of hypopituitarism with a predilection for the hypothalamus. We describe a case of panhypopituitarism in a 57-yr-old Asian lady, associated with an infiltrating hypothalamo-hypophyseal lesion, and other intracranial deposits, initially diagnosed as cerebral tuberculomata. Despite antituberculous therapy, the intracranial lesions progressed with significant clinical deterioration. Repeated lumbar puncture, magnetic resonance imaging scans, liver biopsy and Gallium scan were noncontributory, and the diagnosis of isolated neurosarcoidosis was established only following biopsy of an intracranial lesion. The lesion regressed on steroid and azathioprine therapy. Isolated neurosarcoidosis poses a considerable management problem. We review recent advances in the investigation, diagnosis, and treatment of this condition. PMID- 12108528 TI - The Leeds University Maternity Audit Project. AB - OBJECTIVES: To measure levels of and changes in compliance with evidence-based recommendations in obstetrics in the UK. To identify barriers to and factors associated with compliance. DESIGN: A quantitative case-note audit for 1988 and 1996, and a qualitative interview study of key staff. SETTING: Twenty maternity units, selected at random from all UK units SUBJECTS: Fifty consecutive cases of pre-term delivery (PTD), Caesarean section (CS), instrumental delivery (ID), and perineal repair (PR) operations in each period in each unit. The lead clinician, midwifery manager, a senior midwife, neonatologist, and middle-grade obstetrician in each unit. MAIN OUTCOME MEASURES: Maternal steroid use in PTD, antibiotic use in CS, use of the ventouse (vacuum extractor) rather than forceps as instrument of first choice for ID, and use of polyglycolic acid (PGA) sutures for PR in each time period. Facilities for implementing, staff attitudes to, and the degree of planning to follow each recommendation. MAIN RESULTS: The median proportion of ventouse as instrument of first choice in each unit was 8% (range 0-32%) in 1988, rising to 64% (range 0-98%) in 1996. PGA use for PR was 0% (range 0-30%) in 1988, and 72% (range 0-100%) in 1996. Steroid use for eligible PTD was median 0% (range 0-23%) in 1988, rising to 82% (range 63-95%) in 1996. Antibiotic use for CS was 7% (range 0-25%) rising to 84% (range 10-100%) in 1996. There was no relationship between unit size, type of unit, facilities, staff attitudes or degree of planning, and compliance with the recommendations, nor was the level of adherence to one standard typically correlated with adherence to the others. However, there was a positive correlation (R = 0.6, P < 0.005) between local availability of the Cochrane database of perinatal trials and unit compliance with the audit standards in the latter time period. CONCLUSIONS: We have documented a massive shift in practice in line with the evidence, although many units still have substantial room for improvement. About 2000 wound infections, 200 deaths due to prematurity, nearly 8000 women in pain from catgut sutures, and 1500 cases of severe perineal trauma from forceps remain preventable. The reasons why units vary remain obscure, although the qualitative interviews often revealed local factors such as key enthusiastic staff. There was no sign of evidence being positively driven into practice by any systematic managerial process. The relationship between Cochrane availability and high-standard care may be simply a marker of commitment to the evidence, but it remains plausible that if senior staff make Cochrane available for their juniors, audit compliance improves. PMID- 12108529 TI - Preventing drug-related morbidity--determining valid indicators. AB - OBJECTIVE: To describe the process that is being undertaken to validate a series of indicators for preventable drug-related morbidity - originally developed in the US - for application in the UK health care system. DESIGN: A two-round Delphi questionnaire survey after a preliminary validation of the indicators within the University of Manchester School of Pharmacy. SETTING: A primary care study set in the UK. STUDY PARTICIPANTS: A purposively selected sample of general practitioners with a specific responsibility for prescribing-related issues (n = 6) and pharmacists actively involved in medication review in primary care (n = 10). MAIN OUTCOME MEASURES: The establishment of consensus among the participants that an indicator reflected preventable drug-related morbidity in primary care. RESULTS: After preliminary validation, 37 of the original 57 US indicators were retained. The Delphi panel generated 16 additional new indicators in the first round. At the end of the second round, the pre-defined level of consensus was reached for 29 indicators (19 of the US generated indicators; 10 generated by the panel in the first round). CONCLUSIONS: The Delphi results highlighted differences in both the clinical perspective and, possibly, philosophical viewpoints of health professionals practising in the UK and US health care systems. Further work, located in both primary and secondary care, is now in progress to operationalize the indicators. This process will form a key part of the refining, and hence further validation, of the indicators. The future development of prospective medical-record-based indicators should facilitate a reduction in the human, clinical, and economic burden of drug-related morbidity. PMID- 12108530 TI - Using an explicit guideline-based criterion and implicit review to assess antipsychotic dosing performance for schizophrenia. AB - OBJECTIVE: Using structured implicit review as the gold standard, this study assessed the sensitivity and specificity of an explicit antipsychotic dose criterion derived from schizophrenia guidelines. DESIGN: Two psychiatrists reviewed medical records and made consensus-structured implicit review ratings of the appropriateness of discharge antipsychotic dosages for hospitalized patients who participated in a schizophrenia outcomes study. Structured implicit review ratings were compared with the explicit criterion: whether antipsychotic dose was within the guideline-recommended range of 300-1000 chlorpromazine milligram equivalents (CPZE). In addition, reasons for deviation from guideline dose recommendations were examined. SETTING AND STUDY PARTICIPANTS: A total of 66 patients hospitalized for acute schizophrenia at a Veterans Affairs medical center or state hospital in the southeastern US. MAIN OUTCOME MEASURES: The sensitivity and specificity of the explicit dose criterion at hospital discharge were determined in comparison with the gold standard of structured implicit review. RESULTS: At hospital discharge, 61% of patients (n = 40) were receiving doses within the guideline-recommended range. According to structured implicit review ratings, antipsychotic dose management was appropriate for 80% (n = 53) of patients. When the 300-1000 CPZE dose criterion (dosage within or outside the recommended range) was compared with structured implicit review, it demonstrated 84.6% sensitivity and 71.7% specificity for detecting inappropriate antipsychotic dose. CONCLUSIONS: The explicit antipsychotic dose criterion may provide a useful and efficient screen to identify patients at significant risk for quality of care problems; however, the relatively low specificity suggests that the measure may not be appropriate for quality measurement programs that compare performance among health plans. PMID- 12108531 TI - Staff satisfaction and its components in residential aged care. AB - OBJECTIVE: The purpose of this study was to assess the direction and magnitude of the effects among the components of staff satisfaction in residential aged care and to examine whether the relationships among satisfaction components vary according to facility type (i.e. nursing homes and hostels). A hostel is a low care facility in which residents are more independent, have a lower level of care needs, and receive personal but not nursing care. DESIGN: A cross-sectional survey design was adopted to collect the required information, and a stratified random sampling approach was utilized to select facilities. Structural equation modeling was used to examine relationships among satisfaction components. SETTING: Seventy residential aged care facilities in Western Australia. STUDY PARTICIPANTS: The sample includes 610 nursing home and 373 hostel care staff. RESULTS: The relationships among satisfaction components are different for nursing home and hostel staff. Professional support is found to have a strong and positive effect on all other aspects of staff satisfaction. CONCLUSION: The findings lead to an improved understanding of the interrelationship among staff satisfaction components, which has important implications through enhancing professional support. This needs to be recognized and emphasized by managers, care providers, and policy makers so as to maintain stable personnel and continuity of care. PMID- 12108532 TI - Technical quality control practices in mammography screening programs in 22 countries. AB - OBJECTIVE: To assess current technical quality control (QC) practices within breast cancer screening or surveillance programs internationally. MATERIALS AND METHODS: The International Breast Cancer Screening Network (IBSN) conducted an extensive survey of quality assurance (QA) activities in developed countries known to have population-based breast cancer screening or surveillance programs in place. Twenty-three countries were sent questionnaires that included items about QA and QC requirements at screening sites, the minimum frequencies of QC test performance, and the personnel responsible for performing QC tests. RESULTS: All 23 countries in the IBSN completed general information on their QA practices. Twenty-two countries responded with complete details on their technical QC practices. The responses indicated a pattern of consistently high-quality control practices among population-based breast cancer screening and surveillance programs. Most programs performed the great majority of QC tests. Variations were observed in the performance frequencies of QC tests and in the personnel responsible for performing QC tests. CONCLUSION: QC practices among population based breast cancer screening and surveillance programs are highly evolved, with the great majority of responding countries following prescribed QC protocols. Further research is needed on appropriate performance frequencies for mammography QC tests. PMID- 12108533 TI - Evaluation of quality improvement interventions to reduce inappropriate hospital use. AB - OBJECTIVE: To assess the impact of process analyses and modifications on inappropriate hospital use. DESIGN: Pre-post comparison of inappropriate hospital use after process modifications. SETTING: The Department of Internal Medicine of the Geneva University Hospitals, Switzerland. PARTICIPANTS: A random sample of 498 patients. INTERVENTIONS: Two processes of care (i.e. non-urgent admissions and transfer to a rehabilitation hospital), which influenced inappropriate hospital use, were identified and modified. The impact of these modifications was then assessed. MAIN OUTCOME MEASURES: The proportion of inappropriate hospital admissions and inappropriate hospital days. RESULTS: As a baseline assessment before quality improvement interventions, the appropriateness of hospital use (admissions and hospital days) was evaluated using the Appropriateness Evaluation Protocol (AEP) in a sample of 500 patients (5665 days). After modification of the two processes through a quality improvement program, inappropriate hospital use was reassessed in a sample of 498 patients (6095 days). Inappropriate hospital admissions decreased from 15 to 9% (P = 0.002) and inappropriate hospital days from 28 to 25% (P = 0.12). CONCLUSION: Using the AEP as a criterion, the quality improvement interventions significantly reduced inappropriate hospital use due to the process of non-urgent admissions, but the reduction of inappropriate hospital days specifically attributed to the transfer to the rehabilitation hospital did not reach statistical significance. PMID- 12108534 TI - Translating research into practice: the future ahead. AB - OBJECTIVE: To summarize and analyze the focus and methodologies of the Translating Research into Practice (TRIP) projects funded in 1999-2000 by the US Agency for Healthcare Research and Quality (AHRQ). DATA SOURCES AND STUDY DESIGN: An analysis of the successful applications for the TRIP I and II requests for applications in 1999 and 2000 was produced from the data collected. DATA COLLECTION: The following items were abstracted from each of the successful applications: provider focus, patient population, vulnerable populations, methodologies, interventions for change, outcomes measured, and conceptual framework used. PRINCIPAL FINDINGS: AHRQ funded 27 TRIP grants in 1999 and 2000. A wide variety of health care providers, settings, and patients were the target of the grants. The most common study design was a randomized controlled trial. The most common TRIP interventions were educational and the most common frameworks were either adult learning theory or organizational theory. More than half of the projects planned to use information technology and half the projects had a focus on reducing errors. CONCLUSIONS: The TRIP projects encompass a broad range of providers, environments, patients, and interventions. The field of applied research and quality improvements should be considerably enhanced by these research projects. PMID- 12108535 TI - Assessing doctors' competence: application of CUSUM technique in monitoring doctors' performance. AB - BACKGROUND: Quality assurance of medical practice requires assessment of doctors' performance, whether informally via a system such as peer review or more formally via one such as credentialing. Current methods of assessment are, however, subjective or implicit. More objective methods of assessment based on statistical process control technique such as cumulative sum (CUSUM) procedure may be helpful. OBJECTIVE: To determine the usefulness and acceptability of CUSUM charting for assessing doctors' performance. METHOD: We applied CUSUM charting to assess doctors' performance of endoscopic retrograde pancreatography, renal and breast biopsies, thyroidectomy, and instrumental deliver. A CUSUM chart is a graphical representation of the trend in the outcome of a series of consecutive procedures. At acceptable levels of performance, the CUSUM curve is flat, while at unacceptable levels of performance, the curve slopes upward and eventually crosses a decision interval. When this occurs, the CUSUM chart indicates unsatisfactory performance. Thus, it provides an early warning of an adverse trend. RESULTS: All participating doctors found the technique useful to objectively measure their proficiency. CUSUM charts showed the progress of trainees in acquiring new skills. As they become more skillful with training, their CUSUM curves flatten. Among consultants, level CUSUM curves demonstrated ongoing maintenance of competence. All participants found the technique acceptable as a self-assessment tool. They were, however, less certain of its acceptability as a basis for credentialing. CONCLUSION: We recommend the use of CUSUM charting as a tool for personal audit at an individual level. It may also be used to show proof of technical competence for the purpose of credentialing. PMID- 12108536 TI - Breaks in care in the ambulatory care setting: the risks to patient safety. PMID- 12108537 TI - Structural and functional diversity of connexin genes in the mouse and human genome. AB - Gap junctions are clustered channels between contacting cells through which direct intercellular communication via diffusion of ions and metabolites can occur. Two hemichannels, each built up of six connexin protein subunits in the plasma membrane of adjacent cells, can dock to each other to form conduits between cells. We have recently screened mouse and human genomic data bases and have found 19 connexin (Cx) genes in the mouse genome and 20 connexin genes in the human genome. One mouse connexin gene and two human connexin genes do not appear to have orthologs in the other genome. With three exceptions, the characterized connexin genes comprise two exons whereby the complete reading frame is located on the second exon. Targeted ablation of eleven mouse connexin genes revealed basic insights into the functional diversity of the connexin gene family. In addition, the phenotypes of human genetic disorders caused by mutated connexin genes further complement our understanding of connexin functions in the human organism. In this review we compare currently identified connexin genes in both the mouse and human genome and discuss the functions of gap junctions deduced from targeted mouse mutants and human genetic disorders. PMID- 12108538 TI - Congopain from Trypanosoma congolense: drug target and vaccine candidate. AB - Trypanosomes are the etiological agents of human sleeping sickness and livestock trypanosomosis (nagana), which are major diseases in Africa. Their cysteine proteases (CPs), which are members of the papain family, are expressed during the infective stages of the parasites' life cycle. They are suspected to act as pathogenic factors in the mammalian host, where they also trigger prominent immune responses. Trypanosoma congolense, a major pathogenic species in livestock, possesses at least two families of closely related CPs, named CP1 and CP2. Congopain, a CP2-type of enzyme, shares structural and functional resemblances with cruzipain from T. cruzi and with mammalian cathepsin L. Like CPs from other Trypanosomatids, congopain might be an attractive target for trypanocidal drugs. Here we summarise the current knowledge in the two main areas of research on congopain: first, the biochemical properties of congopain were characterised and its substrate specificity was determined, as a first step towards drug design; second, the possibility was being explored that inhibition of congopain by host-specific antibodies may mitigate the pathology associated with trypanosome infection. PMID- 12108539 TI - Biosynthesis of lysosomal proteinases in health and disease. AB - Proteolytic maturation of lysosomal proteinases is initiated after receptor mediated targeting to prelysosomal compartments, while terminal processing occurs upon delivery to lysosomes. These late processing events are impaired in patients suffering from inherited lysosomal disorders, such as sialic acid storage disease and mucolipidosis II (I-cell disease). Lysosomes in the affected cells display marked changes in their physiological and morphological properties, with features reminiscent of prelysosomal compartments. This indicates that the absence of mature lysosomes interferes with the final processing steps during the biosynthesis of lysosomal proteinases. Thus, impaired proteinase maturation reflects an incompetent lysosomal apparatus and as such can be seen as a hallmark of lysosomal storage diseases. PMID- 12108540 TI - Calpain function in the differentiation of mesenchymal stem cells. AB - Calpain is a calcium-activated non lysosomal neutral thiol protease (EC 3.4.22.17) present in a wide variety of eukaryotic cells. Calpain is usually present as an inactive form and is activated by calcium ions and phospholipids. The ability of calpain to alter, by limited proteolysis, the activity or function of numerous cytoskeletal proteins, enzymes, and receptors suggests its involvement in various Ca2+-regulated cellular functions. In this review we focus on the differentiation of mesenchymal stem cells, such as the myoblastic, osteoblastic, chondrocytic, and adipocytic lineages, and the biological significance of calpain in its regulation. Calpain has been implicated in the differentiation of myoblasts through the turnover of glycoproteins. In preosteoblastic cells, calpain is important in mediating the proliferative and prodifferentiating effects of parathyroid hormone and bone morphogenetic proteins. For the differentiation of chondrocytes, calpain is involved in cartilage-matrix mineralization. Furthermore, calpain is required for the differentiation of 3T3-L1 preadipocytes into adipocytes, involving the transcriptional activation of the C/EBP alpha gene and the degradation of the cyclin-dependent kinase inhibitor p27 during the mitotic clonal expansion phase of adipocyte differentiation. We summarize these regulatory effects of calpain on the differentiation of mesenchymal stem cells and speculate on the function and location of calpain in the differentiation processes. PMID- 12108542 TI - Thyroid stimulating hormone upregulates secretion of cathepsin B from thyroid epithelial cells. AB - Constant levels of thyroid hormones in the blood are principal requirements for normal vertebrate development. Their release depends on the regulated proteolysis of thyroglobulin which is extracellularly stored in the follicle lumen under resting conditions. Thyroglobulin is proteolytically degraded to a major part in lysosomes, but in part also extracellularly leading to the release of thyroxine. Extracellularly occurring lysosomal enzymes are most probably involved in the proteolytic release of thyroxine. In this study we have analyzed the secretion of cathepsin B by thyroid follicle cells (primary cells as well as FRTL-5 cells) and its regulation by thyroid stimulating hormone, which stimulated the secretory release of the proenzyme as well as of mature cathepsin B. Within one to two hours of stimulation with thyroid stimulating hormone, the cathepsin B activity associated with the plasma membrane increased significantly. This increase correlated closely with the localization of lysosomes in close proximity to the plasma membrane of cultured thyrocytes as well as with the thyroxine liberating activity of thyrocyte secretion media. These observations indicate that thyroid stimulating hormone induces the secretion of cathepsin B, which contributes to the extracellular release of thyroxine by thyrocytes. PMID- 12108541 TI - Ku antigen supports termination of mammalian rDNA replication by transcription termination factor TTF-I. AB - A replication fork barrier at the 3'-end of mouse ribosomal RNA genes blocks bidirectional fork progression and limits DNA replication to the same direction as transcription. This barrier is an inherent property of a defined DNA-protein complex including transcription termination factor I, and specific protein protein interactions occur between this factor and protein(s) of the replication machinery. Here we report that a second DNA-binding protein is essential for barrier activity. We have purified and functionally characterised the protein from HeLa cells. The final preparation contained two polypeptides with molecular masses of 70 and 86 kDa, respectively. Both polypeptides interact with a GC stretch adjacent to the binding site of transcription termination factor I. The specificity of binding to the barrier DNA was demonstrated in an electrophoretic mobility shift assay. The biochemical properties of this protein resemble that of Ku antigen, a human nuclear DNA-binding heterodimer that is the target of autoimmune-antibodies in several autoimmune diseases. Recombinant Ku protein, purified as heterodimer from co-infected insect cells, is able to partially rescue the barrier activity in Ku-depleted HeLa cell extracts. These data demonstrate that transcription termination factor I and Ku act synergistically to prevent head-on collision between the replication and the transcription machinery. PMID- 12108543 TI - Selective release of calpain produced alphalI-spectrin (alpha-fodrin) breakdown products by acute neuronal cell death. AB - Activation of calpain results in the breakdown of alpha II spectrin (alpha fodrin), a neuronal cytoskeleton protein, which has previously been detected in various in vitro and in vivo neuronal injury models. In this study, a 150 kDa spectrin breakdown product (SBDP150) was found to be released into the cell conditioned media from SH-SY5Y cells treated with the calcium channel opener maitotoxin (MTX). SBDP150 release can be readily quantified on immunoblot using an SBDP150-specific polyclonal antibody. Increase of SBDP150 also correlated with cell death in a time-dependent manner. MDL28170, a selective calpain inhibitor, was the only protease inhibitor tested that significantly reduced MTX-induced SBDP150 release. The cell-conditioned media of cerebellar granule neurons challenged with excitotoxins (NMDA and kainate) also exhibited a significant increase of SBDP150 that was attenuated by pretreatment with an NMDA receptor antagonist, R(-)-3-(2-carbopiperazine-4-yl)-propyl-1-phosphonic acid (CPP), and MDL28170. In addition, hypoxic/hypoglycemic challenge of cerebrocortical cultures also resulted in SBDP150 liberation into the media. These results support the theory that an antibody-based detection of SBDP150 in the cell-conditioned media can be utilized to quantify injury to neural cells. Furthermore, SBDP150 may potentially be used as a surrogate biomarker for acute neuronal injury in clinical settings. PMID- 12108544 TI - Altered storage of proteases in mast cells from mice lacking heparin: a possible role for heparin in carboxypeptidase A processing. AB - Heparin-deficient mice, generated by gene targeting of N-deacetylase/N sulfotransferase-2 (NDST-2), display severe mast cell defects, including an absence of stored mast cell proteases. However, the mechanism behind these observations is not clear. Here we show that NDST-2+/+ bone marrow-derived mast cells cultured in the presence of IL-3 synthesise, in addition to highly sulphated chondroitin sulphate (CS), small amounts of equally highly sulphated heparin-like polysaccharide. The corresponding NDST-2-/- cells produced highly sulphated CS only. Carboxypeptidase A (CPA) activity was detected in NDST+/+ cells but was almost absent in the NDST-/- cells, whereas tryptase (mouse mast cell protease 6; mMCP-6) activity and antigen was detected in both cell types. Antigen for the chymase mMCP-5 was detected in NDST-2+/+ cells but not in the heparin-deficient cells. Northern blot analysis revealed mRNA expression of CPA, mMCP-5 and mMCP-6 in both wild-type and NDST-2-/- cells. A approximately 36 kDa CPA band, corresponding to proteolytically processed active CPA, as well as a approximately 50 kDa pro-CPA band was present in NDST-2+/+ cells. The NDST-2-/- mast cells contained similar levels of pro-CPA as the wild-type mast cells, but the approximately 36 kDa band was totally absent. This indicates that the processing of pro-CPA to its active form may require the presence of heparin and provides the first insight into a mechanism by which the absence of heparin may cause disturbed secretory granule organisation in mast cells. PMID- 12108545 TI - Clustering-induced signaling of CEACAM1 in PC12 cells. AB - Carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1), an Ig-like transmembrane protein, functions in cell adhesion, angiogenesis and epithelial cell morphogenesis, and has been identified as a tumor suppressor. For all of these functions, CEACAM1 requires signaling capabilities. However, the mechanisms of CEACAM1-mediated signaling are only poorly understood. Here we characterized for the first time CEACAM1 expression and signaling in the neuroendocrine rat pheochromocytoma PC12 cell line. Stimulation of CEACAM1 by ligation on the cell surface with antibodies induced formation of large CEACAM1 clusters and a rapid and transient CEACAM1 tyrosine dephosphorylation. Functionally, this dephosphorylation correlated with a reduced association between CEACAM1 and the tyrosine phosphatase SHP2. Clustering also stimulated binding of CEACAM1 to the actin cytoskeleton, measured by a partial translocation of CEACAM1 into the insoluble fraction after detergent extraction. Both tyrosine dephosphorylation and interaction with the cytoskeleton were sensitive to neuronal differentiation of PC12 cells. The first detected downstream activation of the mitogen-activated protein kinases ERK1 and ERK2, but not of JNK or p38, describes a novel target of CEACAM1-mediated signaling and contributes to the understanding of how CEACAM1 regulates cellular function. PMID- 12108546 TI - Spin adducts of superoxide, alkoxyl, and lipid-derived radicals with EMPO and its derivatives. AB - The compound 5-(ethoxycarbonyl)-5-methyl-1-pyrroline N-oxide (EMPO) is a hydrophilic cyclic nitrone spin trap, which, in contrast to DMPO, forms a relatively stable superoxide adduct (t(1/2)=8.6 min) with an EPR spectrum similar to the respective DMPO adduct. In order to find the optimal degree of lipophilicity of this novel type of spin trap with respect to the detection of radicals formed during lipid peroxidation, the ethoxy group of EMPO was replaced by alkoxy substituents of increasing chain length, leading to the methoxy- (MeMPO), 1-propoxy- (PrMPO), 1-butoxy- (BuMPO), and 1-octyloxy- (OcMPO) derivatives of EMPO. The stability of their superoxide adducts was found to be strongly dependent on the size of the alkoxycarbonyl group. Increasing chain length of the alkoxyl substituent decreased the stability of alkoxyl radical adducts of MeMPO, EMPO, and PrMPO, but increased the stability of OcMPO adducts. The stability of alkoxyl radical adducts of BuMPO, on the other hand, were practically independent of the size of the alkoxyl group. Detection of lipid alkoxyl radicals formed by peroxidizing linoleic acid in a stationary system was therefore only possible with the most lipophilic spin trap, OcMPO. However, with the more hydrophilic spin traps MeMPO, EMPO, PrMPO, and BuMPO optimal EPR signal intensity could be obtained when a slow-flow system was used. Thus, within this series EMPO is the best spin trap for the detection of superoxide; OcMPO, on the other hand, is most suitable for the detection of lipid alkoxyl radicals. PMID- 12108547 TI - Glutathione S-transferase of the malarial parasite Plasmodium falciparum: characterization of a potential drug target. AB - Glutathione S-transferases (GSTs), which occur abundantly in most organisms, are essentially involved in the intracellular detoxification of numerous substances including chemotherapeutic agents, and thus play a major role in the development of drug resistance. A gene encoding a protein with sequence identity of up to 37% with known GSTs was identified on chromosome 14 of the malarial parasite Plasmodium falciparum. It was amplified using gametocyte cDNA and expressed in Escherichia coli as a hexahistidyl-tagged protein of 26 kDa subunit size. The homodimeric enzyme (PfGST) was found to catalyse the glutathione (GSH)-dependent modification of 1-chloro-2,4-dinitrobenzene and other typical GST substrates such as o-nitrophenyl acetate, ethacrynic acid, and cumene hydroperoxide. The Km value for GSH was 164+/-20 microM. PfGST was inhibited by cibacron blue (Ki=0.5 microM), S-hexylglutathione (Ki=35 microM), and protoporphyrin IX (Ki=10 microM). Hemin, a most toxic compound for parasitised erythrocytes, was found to be an uncompetitive ligand of PfGST with a Ki of 6.5 microM. Based on the activity of PfGST in extracts of P. falciparum, the enzyme represents 1 to 10% of cellular protein and might therefore serve as an efficient in vivo buffer for parasitotoxic hemin. Destabilising ligands of GST are thus expected to be synergistic with the antimalarial drug chloroquine, which itself was found to be a very weak inhibitor of PfGST (IC50>200 microM). X-ray quality crystals of PfGST (250x200x50 microm) will serve as starting point for structure-based drug design. PMID- 12108548 TI - Analysis of the structural determinants for RNA binding of the human protein AUF1/hnRNP D. AB - The protein AUF1/hnRNP D was one of the first factors identified that binds to the AU-rich region of certain mRNAs and mediates their fast degradation. Here we describe experiments to address the structural determinants for the binding of AUF1 to the RNA by combining comparative molecular modeling with gel shift assays. From our model of the RNA binding region of AUF1 we predicted that it interacts with RNA predominantly through stacking interactions that do not provide base-specific recognition. Only two RNA positions bound by AUF1 show base preferences: one for pyrimidine bases and the second for a conserved adenine residue. Gel shift assays with a panel of RNA oligonucleotides largely confirmed these model-based binding determinants. An alignment with proteins of the hnRNP family demonstrated that the amino acids involved in the stacking interactions are conserved whereas those that confer a base-specific recognition in AUF1 are variable. PMID- 12108549 TI - Effect of cysteine proteinase inhibitors on murine B16 melanoma cell invasion in vitro. AB - Various types of proteinases are implicated in the malignant progression of human and animal tumors. Proteinase inhibitors may therefore be useful as therapeutic agents in anti-invasive and anti-metastatic treatment. The aims of this study were (1) to estimate the relative importance of proteinases in B16 cell invasion in vitro using synthetic, class-specific proteinase inhibitors and (2) to assess the inhibitory effect of some naturally occurring cysteine proteinase inhibitors. Serine proteinase inhibitor reduced invasiveness by up to 24%, whereas inhibition of aspartic proteinases reduced invasion by 11%. Synthetic inhibitors of cysteine proteinases markedly impaired invasion: cathepsin B inhibitors, particularly Ca 074Me, inhibited invasion from 20-40%, whereas cathepsin L inhibitor Clik 148 reduced invasion by 11%. The potato cysteine proteinase inhibitor PCPI 8.7 inhibited invasion by 21%, whereas another potato inhibitor, PCPI 6.6, and the mushroom cysteine proteinase inhibitor clitocypin had no effects. As the inhibitors that inhibited cathepsin B were in general more efficient at impairing the invasiveness, we conclude that of the two cysteine proteinases, cathepsin B plays a more important role than cathepsin L in murine melanoma cell invasion. PMID- 12108550 TI - Stage-specific antimalarial activity of cysteine protease inhibitors. AB - Cysteine proteases of the malaria parasite Plasmodium falciparum, known as falcipains, are promising targets for antimalarial chemotherapy. We evaluated cultured parasites for the stage-specific expression of cysteine proteases and sensitivity to cysteine protease inhibitors. Protease activity and inhibitor sensitivity varied markedly over time. Cysteine protease activity was greatest in early trophozoites, while sensitivity to cysteine protease inhibitors was greatest in mature trophozoites. Our results indicate the importance of considering the stage-specific effects of antimalarials and are consistent with the conclusion that the principal antimalarial activity of cysteine protease inhibitors is due to a block in hemoglobin hydrolysis. PMID- 12108552 TI - Design of inhibitors for human tissue kallikrein using non-natural aromatic and basic amino acids. AB - We explored the unique substrate specificity of the primary S, subsite of human urinary kallikrein (hK1), which accepts both Phe or Arg synthesizing and assaying peptides derived from Phenylacetyl-Phe-Ser-Arg-EDDnp, a previously described inhibitor with analgesic and anti-inflammatory activities [Emim et al., Br. J. Pharmacol. 130 (2000), 1099-1107]. Phe was substituted by amino acids containing larger aliphatic or aromatic side chains as well as by non-natural basic amino acids, which were designed to combine a large hydrophobic and/or aromatic group with a positively-charged group at their side chains. In general, all peptides with basic amino acids represented better inhibitors than those with hydrophobic amino acids. Furthermore, the S1 subsite specificity proved to be much more selective than the mere distinction between Phe and Arg, for minor differences in the side chains of the non-natural amino acids resulted in major differences in the Ki values. Finally, we present a series of peptides that were assayed as competitive inhibitors for human tissue kallikrein that may lead to the development of novel peptides, which are both more potent and selective. PMID- 12108551 TI - Epoxysuccinyl peptide-derived cathepsin B inhibitors: modulating membrane permeability by conjugation with the C-terminal heptapeptide segment of penetratin. AB - Besides its physiological role in lysosomal protein breakdown, extralysosomal cathepsin B has recently been implicated in apoptotic cell death. Highly specific irreversible cathepsin B inhibitors that are readily cell-permeant should be useful tools to elucidate the effects of cathepsin B in the cytosol. We have covalently functionalised the poorly cell-permeant epoxysuccinyl-based cathepsin B inhibitor [R-Gly-Gly-Leu-(2S,3S)-tEps-Leu-Pro-OH; R=OMe] with the C-terminal heptapeptide segment of penetratin (R=epsilonAhx-Arg-Arg-Nle-Lys-Trp-Lys-Lys NH2). The high inhibitory potency and selectivity for cathepsin B versus cathepsin L of the parent compound was not affected by the conjugation with the penetratin heptapeptide. The conjugate was shown to efficiently penetrate into MCF-7 cells as an active inhibitor, thereby circumventing an intracellular activation step that is required by other inhibitors, such as the prodrug-like epoxysuccinyl peptides E64d and CA074Me. PMID- 12108553 TI - Amyloid fibril formation by human stefin B in vitro: immunogold labelling and comparison to stefin A. AB - The mechanism by which proteins form amyloid fibrils is of high interest to the scientific community as its understanding could resolve questions relevant to conformational diseases. The structural and energetic basis of the process is still largely unknown. The main controversial issue is the co-existence of several protein conformations. Three models for the mechanism of protein fibrillogenesis have been proposed which need to be tested by experiments. In this report, amyloid fibrils grown from human stefin B (type I cystatin) are described. This physiologically relevant protein readily forms fibrils in vitro, in contrast to the homologue--human stefin A--which forms fibrils under extreme conditions only. In order to specifically label stefin B fibrils in vitro, rabbit polyclonal antibody and mouse monoclonal antibody A6/2 against human stefin B were used for immunogold labelling. Samples were examined by transmission electron microscopy. Fibrils of stefin B were strongly labelled using polyclonal antibody and Protein A gold, whereas no positive reaction was observed with monoclonal antibody A6/2. PMID- 12108554 TI - Lysosomal peptidases and glycosidases in rheumatoid arthritis. AB - Lysosomal serine and cysteine proteases are reported to play a role in collagen degradation. In this study, the activities of the lysosomal cysteine proteases cathepsin B and H, dipeptidyl peptidase I, and the serine protease tripeptidyl peptidase I and dipeptidyl peptidase II, all ascribed a role in collagen digestion, were compared with those of the aspartate protease cathepsin D, and lysosomal glycosidases in leukocytes from rheumatoid arthritis patients at different stages of the disease. In all patients the activities of cysteine protease cathepsin B, dipeptidyl peptidase I, aspartate protease cathepsin D, and two glycosidases were elevated, but the activities of the serine proteases tripeptidyl peptidase I, dipeptidyl peptidase II, and the cysteine protease cathepsin H was unchanged. The magnitude of the increased activity was correlated with the duration of the disease. Patients with long-standing RA (10 years or more) had higher cysteine protease activity in their leukocytes than did those with disease of shorter duration. This tendency suggests that elevated lysosomal cysteine protease activities, together with aspartate protease cathepsin D and lysosomal glycosidases (but not serine proteases), are associated with progression of rheumatoid arthritis. PMID- 12108555 TI - The psychological best interest of the child is not the legal best interest. PMID- 12108556 TI - Killing [editorial]. PMID- 12108557 TI - The perceived coerciveness of involuntary outpatient commitment: findings from an experimental study. AB - This study examines self-reported coercion in subjects with severe mental illness who were randomly assigned in an experimental study to continue under, or be released from, involuntary outpatient commitment (OPC) subsequent to hospital discharge. After review of bivariate relationships, multivariable analyses demonstrated significantly higher levels of reported coercion among subjects who experienced longer periods of OPC; who were African American; who were single and not cohabiting; and who had ongoing substance abuse problems, poor insight into illness, and severe symptoms. Case managers' verbal reminders to subjects about the consequences of nonadherence to treatment partially account for higher reports of coercion. Previous reports from this study have found that OPC, if sustained and combined with frequent outpatient mental health services, can improve some outcomes. The current analyses demonstrate that a consequence of OPC is increased perceptions of coercion in the treatment process, which is partially explained by the increased attention by case managers to noncompliance with treatment. PMID- 12108558 TI - Commentary: the search for a formula to relate competence, coercion, and mandated treatment. PMID- 12108559 TI - Stalking, threatening, and harassing behavior by psychiatric patients toward clinicians. AB - The authors surveyed hospital staff to determine how often they had been the target of stalking, threatening, or harassing behavior (STHB) by patients, what strategies they had used to manage the behavior, and their evaluation of various interventions. A written survey about STHB by patients was sent to all clinical staff (N = 82) of the adult inpatient psychiatric service of an urban university hospital. Clinicians who had been the target of such behavior were interviewed about their experiences. Of the 62 staff members who completed the survey, 33 (53%) had experienced some type of STHB during their career. Seventeen of these 33 individuals agreed to be interviewed and provided information about 28 cases of STHB. Staff often rated the behavior as upsetting and disruptive. The frequency with which staff used various management strategies and their perceived effectiveness are described. The results suggest that although severe cases are relatively rare, milder forms of STHB are experienced by a substantial proportion of clinicians and have significant adverse consequences. A variety of management options are available to the clinician when confronted with this situation. PMID- 12108560 TI - "I could tell you, but then I'd have to kill you": classified information in the psychiatric evaluation. AB - Psychiatrists and other mental health professionals are presented with special challenges when their patients are involved in covert operations or other matters of national security. The patients' involvement may, by legal necessity, limit disclosures during the evaluation. Such situations may be encountered with varying degrees of frequency by military psychiatrists or consultants to various federal or law enforcement agencies involved in classified or undercover activities. The need to assess relevant psychosocial stressors while avoiding prohibited disclosure, the legal requirements to report potentially adverse information, or the procedure to gain legal permission to discuss classified details may present novel challenges for therapists in such evaluations. In this article, we present a case report illustrating these challenges and review applicable regulations and public law governing the disclosure of classified information. We also discuss common pitfalls and strategies for handling such situations. PMID- 12108561 TI - Violent behavior, impulsive decision-making, and anterograde amnesia while intoxicated with flunitrazepam and alcohol or other drugs: a case study in forensic psychiatric patients. AB - It is known that many male juvenile delinquents commit violent crimes while intoxicated with flunitrazepam (FZ), often in combination with alcohol or other drugs. We have also noted the combined abuse of FZ with, for example, alcohol in male forensic psychiatric patients. Our objective was to study violent behavior, impulsive decision-making, and amnesia in male forensic psychiatric patients who were intoxicated predominantly with FZ, to increase knowledge of the abuse of FZ in vulnerable subjects. We studied five forensic psychiatric patients, all of whom were assessed in 1998. All of the subjects reported earlier reactions to FZ, including hostility and anterograde amnesia. At the time of their crimes they were all intoxicated with FZ, often in combination with alcohol or other drugs, such as amphetamine or cannabis. In contrast to their behavior based on their ordinary psychological characteristics, their crimes were extremely violent, and the subjects lacked both the ability to think clearly and to have empathy with their victims. Our observations support the view that FZ abuse can lead to serious violent behavior in subjects characterized by vulnerable personality traits, and that this effect is confounded by the concurrent use of alcohol or other drugs. It is evident that FZ causes anterograde amnesia. Previous research and the results presented herein allow us to draw the following conclusion: on the basis of the neuropsychopharmacologic properties of FZ, legal decisions, such as declaring FZ an illegal drug, are needed in countries where it is now legal. PMID- 12108562 TI - Competence-to-stand-trial evaluations of geriatric defendants. AB - This descriptive study compares geriatric defendants (n = 57) found competent to stand trial (n = 36) with those found incompetent (n = 21). A review of the records of 57 consecutive pretrial geriatric detainees who underwent competence to-stand-trial evaluation was conducted. The review included comparison of demographic and historical variables, mental status examination (MSE) elements, and trial abilities. Incompetent subjects were older and more frequently had dementia, but did not necessarily have other psychiatric illnesses. Deficits in orientation, memory, abstraction, concentration, calculation, and thought process were associated with incompetence. Deficits in orientation and memory correlated most highly with incompetence. Trial-related deficits associated with incompetence included failure to understand Miranda warnings, legal charges, potential penalties, roles of court officers, pleas, and plea-bargaining and inability to consult with an attorney and be self-protective. The ability to maintain appropriate courtroom behavior was not different between groups. The inability to consult with an attorney and understand Miranda was most predictive of incompetence-to-stand-trial opinions. PMID- 12108563 TI - Animal cruelty and psychiatric disorders. AB - Animal cruelty in childhood, although generally viewed as abnormal or deviant, for years was not considered symptomatic of any particular psychiatric disorder. Although animal cruelty is currently used as a diagnostic criterion for conduct disorder, research establishing the diagnostic significance of this behavior is essentially nonexistent. In the current study, investigators tested the hypothesis that a history of substantial animal cruelty is associated with a diagnosis of antisocial personality disorder (APD) and looked for associations with other disorders commonly diagnosed in a population of criminal defendants. Forty-eight subjects, criminal defendants who had histories of substantial animal cruelty, were matched with defendants without this history. Data were systematically obtained from the files by using four specifically designed data retrieval outlines. A history of animal cruelty during childhood was significantly associated with APD, antisocial personality traits, and polysubstance abuse. Mental retardation, psychotic disorders, and alcohol abuse showed no such association. PMID- 12108564 TI - On the psychiatric abuse of Falun Gong and other dissenters in China: a reply to Stone, Hickling, Kleinman, and Lee. PMID- 12108565 TI - Tarasoff at twenty-five. PMID- 12108566 TI - Penry revisited: is execution of a person who has mental retardation cruel and unusual? PMID- 12108567 TI - Preserving children's protection while enhancing justice for parents in abuse and neglect evaluations. PMID- 12108568 TI - Reduced punishment for murder in Israel: what constitutes a severe mental disorder? PMID- 12108569 TI - Hendricks v. People: forcing the insanity defense on an unwilling defendant. AB - At least 17 jurisdictions permit insanity defenses to be entered over the objections of defendants. Those jurisdictions believe that society's interest in a just determination of the charges outweighs a competent defendant's choice. If competency includes the ability to rationally choose a plea, competent defendants should not be forced to enter insanity defenses against their wills. PMID- 12108570 TI - The ethical use of psychology in criminal investigations. PMID- 12108571 TI - A British psychiatrist objects to the dangerous and severe personality disorder proposals. PMID- 12108572 TI - A British psychiatrist objects to the dangerous and severe personality disorder proposals. PMID- 12108573 TI - Ghrelin, growth and obesity. AB - The objective of this paper is to review the current evidence of a novel gastric hormone ghrelin. Ghrelin is an endogenous ligand for the growth hormone secretagogue receptor (GHS-R) that potently stimulates growth hormone (GH) release. The discovery of ghrelin opens up a new regulatory system for the GH secretion. Surprisingly, recent studies in rodents suggest that peripherally or centrally administrated ghrelin, independent of GH, decreases fat oxidation and increases food intake and adiposity. In addition, plasma ghrelin levels are lower in obese human subjects. Ghrelin might participate in meal initiation and may signal to the hypothalamus when an increase in the metabolic efficiency is needed. However, the role of ghrelin in the regulation of body weight in humans is unknown. Whether ghrelin's somatotrophic effect surpasses its adipogenic effect in the prolonged administration determines its effect on energy balance. New interesting implications for ghrelin have recently been suggested. For instance, ghrelin's cardiovascular effects might have clinical relevance. Furthermore, ghrelin might be involved in the growth of some neoplasms. In conclusion, ghrelin, a new somatotrophic, orexigenic and adipogenic peptide hormone, links the regulatory systems for growth and energy balance. PMID- 12108574 TI - The enigma of fetal alcohol neurotoxicity. AB - The neurotoxic effects of ethanol on the human fetal brain (fetal alcohol syndrome, FAS) have been recognized for three decades, but the underlying mechanisms have remained elusive. Recently, we discovered that a single episode of ethanol intoxication lasting for several hours can trigger a massive wave of apoptotic neurodegeneration in the developing rat or mouse brain. The window of vulnerability coincides with the developmental period of synaptogenesis, also known as the brain growth-spurt period, which in rodents is a postnatal event, but in humans extends from the sixth month of gestation to several years after birth. We propose that the N-methyl-D-aspartate (NMDA) antagonist and gamma aminobutyric (GABA)mimetic properties of ethanol are responsible for its apoptogenic action, in that we have found that other drugs that block NMDA glutamate receptors or mimic GABA at GABA(A) receptors also trigger apoptotic neurodegeneration in the developing brain. Our findings have clinical significance, not only because they can explain the reduced brain mass and neurobehavioral disturbances associated with the human FAS, but because many agents in the human environment, other than ethanol, have NMDA antagonist or GABAmimetic properties. Such agents include drugs that may be abused by pregnant mothers [phencyclidine (angel dust), ketamine (Special K), nitrous oxide (laughing gas), barbiturates, benzodiazepines], and many medicinals used in obstetric and pediatric neurology (anticonvulsants), and anesthesiology (all general anesthetics are either NMDA antagonists or GABAmimetics). PMID- 12108575 TI - Features of ST segment and T-wave in exercise-induced myocardial ischemia evaluated with multichannel magnetocardiography. AB - BACKGROUND AND AIM: Magnetocardiography (MCG) is a novel, non-contact mapping technique to record cardiac magnetic field. We evaluated MCG criteria for myocardial ischemia in stress testing. METHODS: Multichannel MCG over frontal chest was performed in 44 patients with coronary artery disease (CAD) and 26 healthy controls during supine bicycle exercise test. Of the 44 patients 16 had anterior, 15 posterior, and 13 inferior ischemia documented by coronary angiography and exercise thallium scintigraphy. ST amplitude, ST slope, T-wave amplitude, and ST-T integral were measured. The optimal sites for detecting the ischemia-induced changes on MCG were sought. The orientation of the magnetic field was also determined. RESULTS: The optimal sites for the decrease of ST slope, ST amplitude, T-wave amplitude, and ST-T integral were over the abdomen. The reciprocal increase of these parameters was found over the left parasternal area. The optimal sites were approximately the same for all patient groups. In single-vessel disease patients without previous myocardial infarction (MI), ST slope increase and ST elevation performed the best (area under the receiver operating characteristic curve 92% and 90%, respectively). In post-MI patients with triple-vessel disease the decrease of T-wave amplitude and ST slope performed the best (area under curve 91%, for both). The magnetic field orientation at ST segment performed equally well as the other ST parameters. In stepwise logistic regression analysis, by use of the presence of CAD as the dependent parameter, ST slope increase and ST peak gradient orientation entered the model. CONCLUSIONS: Various ST segment and T-wave parameters detect ischemia in MCG. ST amplitude performs especially well in non-MI patients with less severe CAD. In advanced CAD late development of T-wave amplitude might be more sensitive to ischemia than ST amplitude. PMID- 12108576 TI - Differences in the activation patterns between sustained and self-terminating episodes of human ventricular fibrillation. AB - BACKGROUND: Experimental studies have suggested that R-R interval dynamics during ventricular fibrillation (VF) have organized features, but whether dynamic behavior of non-sustained VF differs from sustained VF is unknown. AIM: The purpose of this study was to investigate whether the dynamics of R-R intervals during non-sustained VF differs from the dynamics during sustained VF. METHODS: A group of 67 patients undergoing routine implantable cardioverter defibrillator (ICD) testing was studied. Forty-three VF events containing mean of 38 local cardiac activation intervals before the termination by ICD shock were analyzed. From intracardiac electrogram recordings, the ratio between the short and long term variability (SD1/SD2) and fractal scaling exponent (alpha) were analyzed. After the initial analyses, data sets were randomized and reanalyzed. Local activation dynamics were then also compared in seven patients with both sustained and spontaneously terminating VF episodes. RESULTS: Randomization caused a change in the VF dynamics from organized toward less organized dynamics (alpha) from 1.08 +/- 0.57 to 0.81 +/- 0.45, P < 0.05 and SD1/SD2 from 0.80 +/- 0.23 to 1.04 +/- 0.20, P < 0.01). Spontaneously terminating VF showed more organized dynamics than sustained VF terminated by shock (P < 0.05). CONCLUSIONS: Local cardiac activation dynamics during initial phase of human VF shows organized dynamics. Spontaneously terminating VF episodes have more structured dynamics than sustained VF. Thus, the dynamic behavior of local cardiac activation intervals may be related to the maintenance of ventricular tachyarrhythmias. PMID- 12108577 TI - Assessing airway inflammation: from guessing to quantitative measurements. AB - Diagnosing asthma and following the response to treatment have relied on lung function measurements. Improved knowledge of the cellular events leading to airflow limitation has led to new clinical methods to assess the inflammatory component of the disease. Induced sputum analysis and exhaled nitric oxide (NO) measurements are already tools for clinical practice. New cell-specific inflammatory markers and further innovations in the testing of exhaled air, e.g. breath condensates and technical development of simple methods and devices, will also benefit the busy practitioner in near future. Assessing airway inflammation by quantitative measurements, instead of guessing, will also strengthen the role of anti-inflammatory medication as first-line treatment of asthma, even in its mildest forms. PMID- 12108578 TI - Molecular mechanisms of adhesion, colonization, and invasion of group A streptococci. AB - The initial step in establishing a bacterial infection is adhesion of the organism to the epithelium of the host. Group A streptococci use multiple adhesins to attach to host cells and the types of adhesins expressed by a particular strain will determine its tissue specificity. Expression of adhesins is regulated in response to changing environmental and growth conditions. Thus, the array of adhesins expressed by a group A streptococcus will depend on the complement of its adhesin genes and on the environment. Expression of some adhesins may trigger internalization of the streptococci by host cells, which may enable the streptococci to evade antibiotics and to facilitate the penetration of deeper tissues. In this review, we present the different molecular mechanisms of adhesion utilized by group A streptococci and how these interactions lead to colonization and invasion. PMID- 12108579 TI - New strategies for identifying gene-gene interactions in hypertension. AB - Essential hypertension is a common disease that has a complex multifactorial etiology. For this reason, it is not surprising that studies of the effects of single genes on hypertension have often failed to replicate the original findings. We propose, as a working hypothesis, that the failure to replicate some single locus results is because the impact of single alleles on the risk of hypertension is dependent on genetic variations at other loci (i.e. gene-gene interactions) and on environmental factors (i.e. gene-environment interactions). Thus, studies that do not consider the appropriate genetic and/or environmental contexts may not identify important susceptibility loci. The identification and characterization of such gene-gene and gene-environment interactions have been limited by a lack of powerful statistical methods and/or a lack of large enough sample sizes. Here, we review the general problem of identifying gene-gene interactions and describe several traditional and several newer methods that are being used to assess complex genetic interactions in essential hypertension. PMID- 12108580 TI - Role of defensins in inflammatory lung disease. AB - The human airways are protected from invading micro-organisms by the highly efficient innate immune system. Antimicrobial peptides that are produced by inflammatory cells and airway epithelial cells are key elements in this innate immune system. A major subgroup of the antimicrobial peptides is the family of defensins--small non-enzymatic and cationic peptides. Besides their extensively studied role in antimicrobial defense, recent studies have demonstrated that defensins are also able to modulate inflammatory responses, to stimulate adaptive immunity and contribute to tissue repair. In line with these observations, increased defensin levels were observed in inflammatory lung diseases, such as cystic fibrosis (CF), diffuse panbroncheolitis (DPB), idiopathic pulmonary fibrosis (IPF) and acute respiratory distress syndrome (ARDS), and in infectious diseases. In the past decade much has been learnt about the activity of defensins and there is abundant evidence for their presence in human inflammatory lung disease. Future studies are required to elucidate their role in the pathogenesis of these diseases. PMID- 12108581 TI - National EMS Research Agenda. AB - Now, more than ever before, the spirit of the emergency services professional is recognized by people everywhere. Individuals from every walk of life comprehend the reality of the job these professionals do each day. Placing the safety of others above their own is their acknowledged responsibility. Rescue and treatment of ill and injured patients are their purpose as well as their gratification. The men and women who provide prehospital care are well aware of the unpredictable nature of emergency medical services (EMS). Prehospital care is given when and where it is needed: in urban settings with vertical challenges and gridlock; in rural settings with limited access; in confined spaces; within entrapments; or simply in the street, exposed to the elements. Despite the challenges, EMS professionals rise to the occasion to do their best with the resources available. Despite more than 30 years of dedicated service by thousands of EMS professionals, academic researchers, and public policy makers, the nation's EMS system is treating victims of illness and injury with little or no evidence that the care they provide is optimal. A national investment in the EMS research infrastructure is necessary to overcome obstacles currently impeding the accumulation of essential evidence of the effectiveness of EMS practice. Funding is required to train new researchers and to help them establish their careers. Financial backing is needed to support the development of effective prehospital treatments for the diseases that drive the design of the EMS system, including injury and sudden cardiac arrest. Innovative strategies to make EMS research easier to accomplish in emergency situations must be implemented. Researchers must have access to patient outcome information in order to evaluate and improve prehospital care. New biomedical and technical advances must be evaluated using scientific methodology. Research is the key to maintaining focus on improving the overall health of the community in a competitive and cost-conscious health care market. Most importantly, research is essential to ensure that the best possible patient care is provided in the prehospital setting. The bravery and dedication of EMS professionals cannot be underestimated. Images of firefighters, EMS personnel, and others going into danger while others are evacuating will remain burned in our collective consciousness. These professionals deserve the benefit of research to assist them in providing the best possible care in the challenging circumstances they encounter. With this document, we are seeking support for elevating the science of EMS and prehospital care to the next level. It is essential that we examine innovative ways to deliver prehospital care. Strategies to protect the safety of both the patient and the public safety worker must be devised and tested. There are many questions that remain to be asked, many practices to be evaluated, and many procedures to be improved. Research is the key to obtaining the answers. PMID- 12108582 TI - Study of the healing process after transplantation of pasteurized bone grafts in rabbits. AB - Different bone allografts (pasteurized, autoclaved, and frozen) were compared based on their osteoinductive properties. Our primary purpose was to examine the biologic qualities of pasteurized allografts, as pasteurization inactivates most viruses transmitted by transplantation. Frozen, pasteurized, and autoclaved allografts were packed into a standard defect of rabbit ulna. The animals were sacrificed at 2 and 4 weeks after surgery. The parts of bones with experimental defects were explored en bloc, and a roentgenogram was carried out. Ulna bone samples were then embedded in methyl-methacrylate. Roentgenograms showed that after 2 weeks, calluses were well-formed, but irregular in shape in all 3 types of allografts. After 4 weeks, the calluses were regular in shape in all but the autoclaved grafts. After 2 weeks, the healing processes had begun in the frozen and pasteurized grafts, with the reaching approximately the same stage, while in the autoclaved grafts these processes were not seen and the bone particles were surrounded by connective tissue without any changes. After 4 weeks, osteoinductive processes were very strong, with the first signs of complete bone remodeling at the bone edges of the defect in pasteurized and frozen allografts. The osteoinductive values of these 2 types were very high and similar. Autoclaved allografts, on the other hand, had very low osteoinductive values, as they were still at the very beginning of the healing process. Histomorphometric analysis revealed a significant difference in both newly formed osteoid thickness and osteoblast number per microm of bone surface in all experimental groups (P < 0.005). Values of osteoid thickness and osteoblast number were significantly higher in both frozen and pasteurized grafts when compared with the autoclaved ones (P < 0.005). Osteogenic properties of pasteurized bone allografts were preserved, and the allografts have been gradually replaced with newly formed bone. As such, pasteurized bone grafts from a bone bank have approximately the same biologic validity as frozen grafts, while autoclaved grafts impair bone healing. PMID- 12108583 TI - Comparison of chemosensitivity tests: clonogenic assay versus MTT assay. AB - When the development of chemotherapeutic agents reaches the clinical trial stage, it is necessary to perform drug sensitivity tests quickly in order to select the most promising agents for the treatment of cancer. In order to assess the possibility of using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay as a substitute for the human tumor clonogenic assay (HTCA), we evaluated the correlation between the results obtained by these 2 assays in 5 human lung cancer cell lines. The correlation coefficient between the results of the HTCA and the MTT assay was 0.673, indicating a relatively good correlation. The correlation was most prominent in platinum analogues (r = 0.939) and good in anthracyclines/anthracenedione (r = 0.611). However, no significant correlation was observed in vinca alkaloids, etoposide, irinotecan, SN-38 (an active metabolite of irinotecan), and rhizoxin. The results of the MTT assay showed a high degree of correlation with those of the HTCA in predicting the sensitivity of cancer cell lines to platinum analogues, and anthracyclines/anthracenedione. These results suggest that the MTT assay may be more convenient and quickly performed than the HTCA and can replace HTCA in evaluating the effects of anticancer agents, especially the platinum analogues and anthracyclines/anthracenedione. PMID- 12108584 TI - Development of syngeneic monoclonal anti-idiotype antibodies to mouse monoclonal anti-asialoglycoprotein receptor antibody. AB - Anti-idiotype antibodies (Ab2) play an important role in the homeostasis of immune responses and are related to the development and the disease activity of certain autoimmune diseases. The asialoglycoprotein receptor (ASGPR) is considered one of the target antigens in the pathogenesis of autoimmune chronic active hepatitis (AIH). We previously developed a mouse monoclonal antibody (clone 8D7) which recognizes rat and human ASGPR. In this study, to help investigate the anti-ASGPR antibody-anti-idiotype antibody network in patients with AIH, we developed a syngeneic mouse monoclonal Ab2 to the 8D7 anti-ASGPR antibody (Ab1). One clone, designated as 3C8, tested positive for specific reactivity to 8D7-Ab1 and did not bind to other irrelevant immunoglobulins. By competitive inhibition assays, the binding of 8D7-Ab1 to liver membrane extracts, i.e., the crude antigen preparation, was inhibited by 3C8-Ab2 in a dose-dependent manner, and the binding of 8D7-Ab1 to 3C8-Ab2 was inhibited by the liver membrane extracts. In the immunohistochemical analysis, 3C8-Ab2 blocked the specific staining of sinusoidal margins of rat hepatocytes by 8D7-Ab1. These results suggest that 3C8 anti-idiotype antibody recognizes the specific idiotypic determinants within the antigen-binding site of 8D7-Ab1. PMID- 12108585 TI - Hepatitis C virus quasispecies in cancerous and noncancerous hepatic lesions: the core protein-encoding region. AB - We have shown that highly proofreading DNA polymerase is required for the polymerase chain reaction in the genetic analysis of hepatitis C virus (HCV). To clarify the status of HCV quasispecies in hepatic tissue using proofreading DNA polymerase, we performed a genetic analysis of the HCV core protein-encoding region in cancerous and noncancerous lesions derived from 4 patients with hepatocellular carcinoma. In contrast to the previously published data, we observed neither deletions nor stop codons in the analyzed region and no significant difference in the complexity of HCV quasispecies between cancerous and noncancerous lesions. This result suggests that the HCV core gene is never structurally defective in hepatic tissues, including cancerous lesions. However, in 3 of the patients, the consensus HCV species differed between cancerous and noncancerous lesions, suggesting that the predominant replicating HCV species differs between these 2 types of lesions. Moreover, during the course of the study, we obtained several interesting variants possessing a substitution at codon 9 of the core gene, whose substitution has been shown to induce the production of the F protein synthesized by a - 2/+1 ribosomal frameshift. PMID- 12108586 TI - Effect of change in body mass index on morbidity in non-obese university graduates. AB - To establish the actual serial changes in body weight in Japanese people and to elucidate the influence of changes in BMI on morbidity, we conducted a historical cohort study of university graduates from 1955 to 1990 using questionnaires and BMI data. The subjects of this study were 3,675 university graduates aged 26-62 years in whom BMI was determined at the time of enrollment in the university (Pre BMI), 5 to 40 years earlier. Morbidity (one or more system diseases or obesity related system diseases) was analyzed according to current age, sex, current BMI, deltaBMI (difference between current BMI and pre-BMI), and various lifestyle variables. The proportion of overweight subjects at enrollment to university was higher in recent male students compared to old students, but not in female graduates, and the BMI in both genders increased progressively after graduation, especially in recent male graduates. Pre-BMI correlated negatively and significantly with deltaBMI. The percentages of obese (BMI > or = 30 kg/m2) males and females were 1.6% and 0.5%, respectively, and high morbidity was observed in 56.1% and 42.2% of males and females, respectively. Stepwise regression analysis showed that in subjects with normal BMI at enrollment, prospective morbidity was dependent on ABMI in addition to age. Our results indicate that in subjects with normal body weight, prospective morbidity is determined by increment of ABMI, and suggest that maintenance of BMI at the late adolescence level is an important factor in preventing future disease. PMID- 12108587 TI - A radiographic study of hip-joint alignment after prosthetic hip arthoplasty. AB - Postoperative hip alignment was studied on radiographs in cases of total hip arthroplasty (THA) and of Bipolar Head Prosthesis(BHP), both with MX-1. Postoperative anteroposterior-view radiographs of hip joints of patients with a normal hip joint on the unoperated side and without pelvic tilt were used. Thirty nine THA patients (femoral neck fracture), 26 THA patients (osteonecrosis of the femoral head and osteoarthritis of the hip joint), and 34 BHP patients were selected for this study. Lines and points for measurement of 9 parameters were established on radiographs. The position of the greater trochanter upper edge is 6.5 mm (mean) superior to the femoral head center in the normal hip joint of Japanese, unlike in Caucasians. A femoral head prosthesis should be inserted so that its center and the greater trochanter upper edge are level in order to equalize leg lengths. In BHP cases, the insertion is made so that the greater trochanter upper edge is approximately 4-mm superior to the center of the prosthesis. For further securing of the stem and to equalize leg lengths, stems should be available in 11 diameters from 5-15 mm in 1-mm increments. Postoperative hip alignment in MX-1 THA cases was found to be satisfactory. PMID- 12108588 TI - An unusual cause of hypercalcemia in polycythemia vera: parathyroid adenoma. AB - In this paper we describe a patient with polycythemia vera (PV), who presented with hypercalcemia due to a parathyroid adenoma. In November 1999, the patient was admitted to our hospital with meteorism and constipation. Her physical examination revealed plethora and hepatosplenomegaly. Laboratory data revealed hyperparathyroidism in addition to PV: Rbc 8 x 10(6)/mm3, Hct 63.7%, serum calcium 13.4 mg/dl, serum phosphorus 1.2 mg/dl, albumin 4.25 mg/dl, and alkaline phophatase activity 433 U/l. Intact Parathyroid Hormone level (iPTH) was 376 pg/ml (n.v.12-72 pg/ml). Twenty-four hour urinary calcium excretion was higher than normal (900 mg). A parathyroid adenoma was detected with Tc-99m sesta-MIBI scanning under the left lobe of the thyroid gland and an ultrasonographic examination of the neck also supported the diagnosis. The patient was recommended for surgery. The histopathological examination confirmed the diagnosis. Postoperatively, iPTH dropped to 53.4 pg/ml at the 15 th minute and to 33.5 pg/ml at the first hour. The calcium level was 7.5 mg/dl one hour after the operation. Five days later, Hct was 40.8%. This case represents a rare association between PV and primary hyperparathyroidism, and may provide evidence for a causal link between PTH and polycythemia vera in our patient. In conclusion, this case indicates that the differential diagnosis of hypercalcemia and polycythemia vera should also include the possibility of a parathyroid tumor in addition to malignancy. PMID- 12108589 TI - Quantitative modelling of crude oil toxicity using the approach of cybernetics and structured mechanisms of microbial processes. AB - Cybernetics and structured approach to biochemical processes in microbial cells offer the status of using various cell components of Azotobacter as molecular markers in toxicity assay of environmental toxicants. The intra-linked dynamic physicochemical reactions of the basic macromolecules--DNA, protein and membrane lipids--with respect to the crude oil in the growth environment, results in an 'impulse transfer function'. The structured effects are reduction in both DNA and protein levels, and an elevated level of lipid peroxidation products. Toxicity index (Ti) of the crude oil, at a given concentration, is the percent ratio of the summation of the products of these effect parameters' and their respective weightings, relative to control. The EC50 is the effective percent (w/v) concentration of the crude oil at which a toxicity index of 50% was recorded, and this corresponded with about 44% loss in nitrogen fixation of the diazotrophic bacterium. PMID- 12108590 TI - A terrain-based paired-site sampling design to assess biodiversity losses from eastern hemlock decline. AB - Biodiversity surveys are often hampered by the inability to control extraneous sources of variability introduced into comparisons of populations across a heterogenous landscape. If not specifically accounted for a priori, this noise can weaken comparisons between sites, and can make it difficult to draw inferences about specific ecological processes. We developed a terrain-based, paired-site sampling design to analyze differences in aquatic biodiversity between streams draining eastern hemlock (Tsuga canadensis) forests, and those draining mixed hardwood forests in Delaware Water Gap National Recreation Area (USA). The goal of this design was to minimize variance due to terrain influences on stream communities, while representing the range of hemlock dominated stream environments present in the park. We used geographic information systems (GIS) and cluster analysis to define and partition hemlock dominated streams into terrain types based on topographic variables and stream order. We computed similarity of forest stands within terrain types and used this information to pair hemlock-dominated streams with hardwood counterparts prior to sampling. We evaluated the effectiveness of the design through power analysis and found that power to detect differences in aquatic invertebrate taxa richness was highest when sites were paired and terrain type was included as a factor in the analysis. Precision of the estimated difference in mean richness was nearly doubled using the terrain-based, paired site design in comparison to other evaluated designs. Use of this method allowed us to sample stream communities representative of park wide forest conditions while effectively controlling for landscape variability. PMID- 12108591 TI - Organochlorine insecticide residues in drinking and ground water in and around Delhi. AB - A multiresidue method was developed for the estimation of 15 organochlorine pesticides in water. 50 samples of drinking water supplied by the Municipal Corporation to the residential areas of Delhi and 20 ground water samples from nearby villages used for irrigation were monitored for the presence of organochlorine insecticides by the method developed. Although, organochlorine pesticides were detected in the ground water and irrigation water samples, the levels of pesticides were below the Maximum Contaminant Level as prescribed by WHO. No organochlorine insecticides were detected in any of the drinking water samples. PMID- 12108592 TI - Time dependent influx and efflux of phenol by immobilized microbial consortium. AB - Phenol, like many other organic solvents, is toxic to micro-organisms even at low concentrations. However, some micro-organisms can withstand this toxicity to a certain concentration. To observe the uptake mechanism of phenol, bacteria were isolated from a petroleum refinery effluent and identified. Study was carried out to understand the effect of varying sub-lethal concentrations of phenol, on all the isolated individual bacterial cultures. Out of the bacteria isolated, Serratia liquefaciens was found to tolerate phenol concentration up to 1500 mg l( 1). A microbial consortium of the isolated bacteria was formulated and immobilized. Individual cultures were also immobilized and uptake of phenol by the immobilized micro-organisms was observed in a nutrient-rich and nutrient stressed medium containing phenol as a sole source of carbon. A time-dependent uptake of phenol was exhibited by the micro-organisms in nutrient-stressed medium, after which a sudden increase in phenol concentration occurred in the extracellular medium, till it reached back to the initial concentration. This was attributed to an active efflux mechanism adopted by the micro-organisms to withstand the toxic shock. PMID- 12108593 TI - Losses of nitrogen and phosphorus from agricultural and forest areas in Finland during the 1980s and 1990s. AB - The temporal changes and spatial variability of phosphorus and nitrogen losses and concentrations in Finland during the period 1981-1997 were studied in 15 small agricultural and forested catchments. In addition, four coastal river basins with high agricultural land use located in southern Finland were included in the study in order to assess the representativeness of agricultural loss estimates from small agricultural catchments. The mean annual loss specific for agricultural land was estimated to be on average 110 kg km(-2) a(-1) for total phosphorus and 1500 kg km(-2) a(-1) for total nitrogen. The results from small agricultural catchments were in agreement with the corresponding loss estimates from rivers, with an average of 137 kg km(-2) a(-1) for total phosphorus and 1800 kg km(-2) a(-1) for total nitrogen. The results from the studied agricultural catchments and rivers during the period 1981-1997 suggest that weather-driven fluctuation in discharge was usually the main reason for changes in nutrient losses, and little or no impact of changes in agricultural production or management practises can be observed. In forested areas the total phosphorus loss (average 9 kg km(-2) a(-1)) and total nitrogen loss (average 250 kg km(-2) a(-1)) were lower than in agricultural areas. In forested catchments the impact of forestry operations, such as clear-cutting and fertilization, and the impact of atmospheric nitrogen deposition can be seen in changes in nutrient losses. PMID- 12108594 TI - Behaviour and fluxes of trace and toxic elements in creek sediment near Mumbai, India. AB - Studies on marine sediments are extremely important since they act as ultimate sink of anthropogenic pollutants. The present study was conducted near Mumbai city of India to understand and assess the behaviour and fluxes of trace and toxic elements in creek sediment. Seven sediment core samples were collected and analysed for trace and toxic elements such as Fe, Cu, Pb, Zn, Rb and Sr in different sections of the core using EDXRF technique. The fluxes of the elements in each section of the core were calculated using the mass sedimentation rates derived from 210Pb dating technique and the sediment density at each location. The estimated depositional fluxes of Fe, Rb and Sr in Zone-1 and Zone-3 are in the ranges of 0.4-0.5% cm(-2) yr(-1); 4-6 microg cm(-2) yr(-1) and 10-20 microg cm(-2) yr(-1) respectively, where as they were about 3-4 times higher in zone-2 for the same elements. The depositional fluxes of elements Cu (40-60 microg cm( 2) yr(-1)), Zn (35-43 microg cm(-2) yr(-1)) and Pb (6-12 microg cm(-2) yr(-1)) were also found to be higher in zone-2 compared to zone-1 and zone-3 which can be attributed to the release from the newly developed chemical zone of Thane-Belapur industrial belt. PMID- 12108595 TI - Kary B. Mullis--Nobel Laureate for procedure to replicate DNA. PMID- 12108596 TI - Images and reflections from Mayo Clinic heritage. PMID- 12108597 TI - Bioterrorism preparedness for the public health and medical communities. PMID- 12108598 TI - Risk indices and osteoporosis screening: scope and limits. PMID- 12108599 TI - Detection of vaccinia virus, herpes simplex virus, varicella-zoster virus, and Bacillus anthracis DNA by LightCycler polymerase chain reaction after autoclaving: implications for biosafety of bioterrorism agents. AB - OBJECTIVE: To determine whether autoclaving suspensions of vaccinia virus, herpes simplex virus (HSV), varicella-zoster virus (VZV), and Bacillus anthracis inactivate infectivity of these agents but allow detection of target DNA by LightCycler polymerase chain reaction (PCR). MATERIAL AND METHODS: Swabs were inserted into tubes containing serial 10-fold dilutions (10(-1) to 10(-5); 500 microL; 6 samples per dilution) of vaccinia virus, HSV, VZV, or a single suspension of 10(8) colony-forming units of B anthracis (2 samples). One half of the samples were autoclaved, and the remainder were not. An aliquot of each not autoclaved sample served as a control for infectivity. RESULTS: Autoclaving swabs saturated with suspensions of vaccinia virus, HSV, or VZV eliminated the infectivity of these agents; however, DNA was detectable in most autoclaved samples in dilutions of 10(-1) to 10(-4) by LightCycler PCR. All not autoclaved specimens were detected by culture (infectivity) except for VZV and, in most dilutions of 10(-1) to 10(-3), by assay of target DNA by LightCycler PCR. Similarly positive results were obtained for PCR assessment of sporulated B anthracis. CONCLUSIONS: Standard autoclaving procedures eliminated the infectivity of viruses (and B anthracis), but target DNA was often retained for detection by LightCycler PCR. Current recommendations indicate that the laboratory diagnosis of smallpox virus infection be performed only within Biosafety Level 4 facilities. We suggest that, in addition to the requirement for immediate coordination with public health officials, the federal government consider expanding the existing guidelines for processing these specimens to encourage immediate collection, autoclaving, and testing by LightCycler PCR to differentiate smallpox virus from other dermal pathogens such as HSV and VZV by specific qualified laboratories. PMID- 12108600 TI - Performance of risk indices for identifying low bone density in postmenopausal women. AB - OBJECTIVE: To examine the ability of 4 published osteoporosis risk indices to identify women with low bone density. SUBJECTS AND METHODS: Subjects included postmenopausal women 45 years and older consecutively recruited from US clinics, women from general practice centers in The Netherlands (age range, 50-80 years), women in the Rotterdam Study (The Netherlands) 55 years and older, and women aged 55 to 81 years old screened for a clinical trial of alendronate. Bone mineral density (BMD) was measured at the femoral neck or lumbar spine; T scores represent the number of SDs below the mean for young healthy women. One risk index was calculated from age and weight; the other risk indices included up to 4 additional variables obtained by questionnaire. We calculated the sensitivity and specificity for identifying women with BMD T scores of -2.5 or less or -2.0 or less in the US clinic sample and created 3 risk categories, using each of the 4 indices. RESULTS: Data were available for 1102 women from the US clinic sample, 3374 women in the Rotterdam Study, 23,833 women screened for a clinical trial of alendronate, and 4204 women from general practice centers in The Netherlands. Specificity for identifying BMD T scores of -2.5 or less ranged from 37% to 58% (depending on risk index) when sensitivity was approximately 90%. The prevalence of osteoporosis (defined as T scores < or = -2.5) differed widely across the 3 risk categories, ranging from 2% to 4% for the low-risk category to 47% to 61% for the high-risk category in the US clinic sample. For spine BMD in the US clinic sample, the prevalence of T scores of -2.5 or less ranged from 7% (low risk) to 38% (high risk). The large differences in prevalence across risk categories were consistent across the other 3 samples of postmenopausal women in the United States and The Netherlands for all 4 risk indices. CONCLUSIONS: We recommend measuring BMD in women who are classified as having an increased risk of osteoporosis by using any of these risk indices because all 4 indices appear to predict low bone mass equally well. The Osteoporosis Self-assessment Tool index is easiest to calculate and therefore may be most useful in clinical practice. PMID- 12108601 TI - Sex differences in evaluation and outcome after stress testing. AB - OBJECTIVE: To examine sex differences in evaluation and outcome after stress testing for coronary artery disease (CAD) in a geographically defined cohort. SUBJECTS AND METHODS: Subjects were residents of Olmsted County, Minnesota, who underwent an initial stress test between January 1, 1987, and December 31, 1990. End points included referral for coronary angiography, death, and cardiac events, defined as cardiac death, nonfatal myocardial infarction, or congestive heart failure. RESULTS: A total of 2276 men and 1270 women under went stress tests. Women were older and had more risk factors and comorbidities (P < .05). Among persons without documented CAD (86% of the cohort), the median probability of CAD was 11% (interquartile range, 5%-25%) for men and 8% (interquartile range, 2% 31%) for women (P < .001). Within 6 months after stress testing, 9% of men and 7% of women underwent coronary angiography. Among persons without documented CAD, there was no sex difference in referral for angiography when the stress test result was negative. When the test result was positive, men were more likely to be referred for angiography (adjusted odds ratio [OR] for male sex, 2.02; 95% confidence interval [CI], 1.21-3.38; P = .008). After adjusting for the predicted probability of CAD, this association was no longer detected (adjusted OR for male sex, 0.67; 95% CI, 0.26-1.73; P = .41). Among persons with documented CAD, no sex difference was noted. After a mean +/- SD follow-up of 7.6 +/- 2.7 years and among persons without documented CAD, male sex was associated with a higher adjusted risk of death (relative risk for male sex, 1.40; 95% CI, 1.05-1.86; P = .02) and cardiac events (relative risk for male sex, 1.67; 95% CI, 1.24-2.26; P < .001). Among persons with documented CAD, no sex difference in outcome was noted. CONCLUSION: These population-based data indicate that, when the diagnosis of CAD was not established, there was a greater use of angiography among men with positive stress test results, which could be attributed to the increased probability of CAD in men. In the absence of documented CAD, men fared worse than women, with an increase in the risk of death and cardiac events. Among persons with documented CAD, no sex difference in use of angiography and outcome was noted. PMID- 12108602 TI - Spectrum of reoperations after repair of aortic coarctation: importance of an individualized approach because of coexistent cardiovascular disease. AB - OBJECTIVE: To determine the indications for and spectrum of late reoperations in adults who had previously undergone coarctation repair. PATIENTS AND METHODS: We reviewed clinical, cardiac catheterization, and echocardiographic data and criteria for reoperation, surgical procedures, and outcome in 43 patients who underwent 54 reoperations between 1972 and 1996. RESULTS: Of the reoperations for recoarctation or associated cardiovascular disease (or both), 20% were performed in asymptomatic patients and 80% in symptomatic patients. Associated cardiovascular disease included bicuspid aortic valve in 36 patients (84%), aortic arch hypoplasia in 12 (28%), true or false aortic aneurysm in 6 (14%), mitral valve disease in 6 (14%), and subvalvular aortic stenosis in 5 (12%). Surgical procedures included 22 recoarctation repairs and 32 other cardiovascular interventions. Simultaneous repair of recoarctation and associated cardiovascular disease was performed as a single-stage repair in 5 reoperations through a median sternotomy using an extra-anatomic, ascending-to-descending aortic bypass, with no complications. One patient died (surgical mortality, 1.9%) of preexisting severe pulmonary vascular obstructive disease. CONCLUSIONS: After coarctation repair, associated cardiovascular diseases are the most common cause for reoperation. An individualized surgical approach is important and may range from valve replacement or recoarctation surgery to extra-anatomic bypass combined with other cardiovascular procedures, enabling simultaneous repair of recoarctation and associated lesions. Despite complex surgical techniques and multiple reoperations, morbidity and mortality were low in our series. PMID- 12108603 TI - Audit of nutrition support for hematopoietic stem cell transplantation at a single institution. AB - OBJECTIVE: To analyze experience with total parenteral nutrition (TPN) for hematopoietic stem cell transplantation (HSCT) at our institution compared with reports in the literature. PATIENTS AND METHODS: We reviewed medical records of 100 patients (53 men and 47 women) who underwent HSCT from 1992 to 2001. Data were abstracted on demographics, primary diagnosis, type of transplantation, myeloablative regimen, length of hospital stay, time to engraftment, 1- and 5 year survival, initiation and duration of TPN, and TPN-related complications. RESULTS: Seventy-one transplantations were autologous, 27 allogeneic, and 2 syngeneic. The median age of the patients was 51 years (range, 19-71 years). We initiated TPN when patients' oral caloric intake was less than 50% of their estimated needs (4 to 7 days after the start of myeloablative therapy; median, 1.2 days after HSCT; range, 8 days before HSCT to 13 days after HSCT). We discontinued TPN when oral intake was more than 50% of estimated needs (median duration, 16 days for autologous and 24 days for allogeneic transplantations, with the shortest duration in breast cancer patients and the longest duration in those treated with cyclophosphamide). Mean weight loss was less than 2%. No differences in patient characteristics, myeloablative regimen, or diagnosis were observed between patients who required and those who did not require TPN. Infection, hospital stay, time to engraftment, and mortality were comparable to published reports. CONCLUSION: In patients undergoing HSCT, TPN should not be initiated until oral caloric intake is less than 50% of estimated needs. During the period of inadequate oral intake, TPN maintains stable body weight with longer duration of support needed for patients undergoing allogeneic than for those undergoing autologous transplantations. PMID- 12108604 TI - Confronting bioterrorism: physicians on the front line. AB - The events surrounding September 11, 2001, and its aftermath have compelled the public health and medical community to face the hitherto unfamiliar reality of bioterrorism. Physicians and public health personnel are frontline soldiers in this new form of warfare. This article provides a general overview of the pathophysiology, clinical presentation, diagnosis, and management of patients infected with the 6 highest priority agents that could potentially be used in bioterrorism. The diseases discussed include anthrax, smallpox, tularemia, plague, botulism, and viral hemorrhagic fevers. Despite the unpredictable nature of bioterrorism, disaster preparedness and knowledge of essential diagnostic and epidemiological principles can contribute substantially toward combating this new threat. PMID- 12108605 TI - Application of rapid-cycle real-time polymerase chain reaction for the detection of microbial pathogens: the Mayo-Roche Rapid Anthrax Test. AB - Rapid-cycle real-time polymerase chain reaction has immediate and important implications for diagnostic testing in the clinical microbiology laboratory. In our experience this novel testing method has outstanding performance characteristics. The sensitivities for detecting microorganisms frequently exceed standard culture-based assays, and the time required to complete the assays is considerably shorter than that required for culture-based assays. We describe the principle of real-time polymerase chain reaction and present clinical applications, including the detection of Bacillus anthracis, the causative agent of anthrax. This latter test is commercially available as the result of a collaborative venture between Mayo Clinic and Roche Applied Science, hence the designation The Mayo-Roche Rapid Anthrax Test. PMID- 12108606 TI - Achieving and maintaining cognitive vitality with aging. AB - Cognitive vitality is essential to quality of life and survival in old age. With normal aging, cognitive changes such as slowed speed of processing are common, but there is substantial interindividual variability, and cognitive decline is clearly not inevitable. In this review, we focus on recent research investigating the association of various lifestyle factors and medical comorbidities with cognitive aging. Most of these factors are potentially modifiable or manageable, and some are protective. For example, animal and human studies suggest that lifelong learning, mental and physical exercise, continuing social engagement, stress reduction, and proper nutrition may be important factors in promoting cognitive vitality in aging. Manageable medical comorbidities, such as diabetes, hypertension, and hyperlipidemia, also contribute to cognitive decline in older persons. Other comorbidities such as smoking and excess alcohol intake may contribute to cognitive decline, and avoiding these activities may promote cognitive vitality in aging. Various therapeutics, including cognitive enhancers and protective agents such as antioxidants and anti-inflammatories, may eventually prove useful as adjuncts for the prevention and treatment of cognitive decline with aging. The data presented in this review should interest physicians who provide preventive care management to middle-aged and older individuals who seek to maintain cognitive vitality with aging. PMID- 12108607 TI - Female sexual dysfunction. AB - Female sexual dysfunction (FSD) was recently recognized as arising from multiple organic etiologies; it is not primarily a psychological symptom as believed previously. A symptom-related complex resulting in physiologic changes, FSD can respond to either treatment of the underlying condition or supportive measures. A new diagnostic classification allows physicians to perform a clinical evaluation of women with FSD, and recently validated FSD question naires allow monitoring of treatment efficacy. This article details the clinical evaluation and physical examination of women with FSD and outlines the fledgling research and treatment options. PMID- 12108608 TI - 68-year-old man with generalized weakness. PMID- 12108609 TI - Septic arthritis due to Streptococcus sanguis. AB - We report an unusual case of septic arthritis due to Streptococcus sanguis, a member of the viridans group of streptococci that are part of the normal flora of the mouth and upper respiratory tract. Our patient had severe underlying periodontal disease, which likely contributed to his joint sepsis through hematogenous spread. Although viridans streptococci are rare causes of septic arthritis in native joints, they should be considered in the setting of severe periodontal disease. PMID- 12108610 TI - Toxic effects associated with consumption of zinc. AB - A 27-year-old man with a history of acne presented to his primary care physician because of fatigue and dyspnea on exertion of 4 weeks' duration. He was remarkably pale, orthostatic pulse changes were noted, and a systolic ejection murmur was heard. The patient had profound anemia (hemoglobin concentration, 5.0 g/dL) and neutropenia (neutrophil count, 0.06 x 10(9)/L); he was admitted for further evaluation. A detailed inquiry into his medication history revealed that he was taking several vitamins and zinc gluconate, 850 to 1000 mg/d for 1 year (US recommended daily allowance, 15 mg), as therapy for acne. A zinc toxic and copper-deficient state was confirmed by laboratory studies. The patient was treated with intravenous copper sulfate, followed by 3 months of oral therapy. The complete blood cell count, serum copper level, and serum zinc level returned to normal. PMID- 12108611 TI - Locally invasive cutaneous Apophysomyces elegans infection acquired from snapdragon patch test. AB - Apophysomyces elegans is an environmental fungus related to other well-known agents of zygomycosis. We report a case of locally invasive A elegans soft tissue infection resulting from the application of a skin patch to test for snapdragon sensitivity. The infection was limited to skin and soft tissue, and treatment consisted of local debridement and liposomal amphotericin B. Outcome was successful. PMID- 12108612 TI - Acute tumor lysis syndrome in a patient with mixed small cell and non-small cell tumor. AB - Tumor lysis syndrome, caused by massive tumor cell death, is an infrequent occurrence in solid tumors, and only a few cases of tumor lysis syndrome occurring in patients with lung cancer have been reported. We present a case of tumor lysis syndrome in a patient with mixed small cell and non-small cell lung cancer complicated by Listeria sepsis. Despite aggressive supportive measures with fluids, electrolytes, antibiotics, pressor agents, ventilation, and alkalinization of the urine, multiorgan failure developed, and the patient died on day 5 in the intensive care unit. Physicians should be aware of this infrequent but potentially fatal complication occurring in critically ill patients with bulky solid tumors so that early and aggressive therapeutic measures can be initiated and appropriate monitoring can be performed. PMID- 12108613 TI - Ma huang toxicity. PMID- 12108614 TI - Complex regional pain syndrome. PMID- 12108615 TI - Chronic sympathetic activation. PMID- 12108616 TI - Efficacy of porcine secretin in children with autism and pervasive developmental disorder. AB - Secretin, a gastrointestinal (GI) hormone, was reported in a preliminary study to improve language and behavior in children with autism/pervasive developmental disorder (PDD) and chronic diarrhea. To determine the efficacy of secretin, we completed a double-blind, placebo-controlled, crossover (3 weeks) study in children with autism/PDD and various GI conditions using a single dose of intravenous porcine secretin. Children with chronic, active diarrhea showed a reduction in aberrant behaviors when treated with the secretin but not when treated with the placebo. Children with no GI problems are unaffected by either secretin or placebo. The improvement seen with secretin in children with autism/PDD and chronic diarrhea suggests that there may be a subtype of children with autism/PDD who respond to secretin. PMID- 12108617 TI - Double-blind placebo-controlled trial of secretin: effects on aberrant behavior in children with autism. AB - Secretin has been proposed as a treatment alternative for autistic spectrum disorders, but empirical support is lacking. A double-blind placebo-controlled study examined the effect of a single dose of synthetic human secretin on aberrant behavior. Parent and teacher data from the Aberrant Behavior Checklist for eight male children were analyzed for reliable change in a clinical replication series. By parent and teacher report, the majority of change occurred either on the placebo trial or reflected deterioration subsequent to secretin infusion. Repeated-measures multivariate analysis of variance results were similar. Results are consistent with other studies, suggesting that secretin may not be an effective treatment option. PMID- 12108618 TI - Perception of human and nonhuman facial age by developmentally disabled children. AB - Human and nonhuman faces were shown to clinical controls, autistic, mentally retarded, and language-disordered children to assess their ability to detect and draw inferences about facial age. Children were asked to select from sets of three faces the one that appeared youthful or to select faces that would be associated with some age-related characteristic. In two studies, it was found that, relative to other children, autistic children had more difficulty perceiving youthfulness in nonhuman faces compared with human faces. These data are discussed with respect to differences in mechanisms and processes that may underlie facial information processing in autistic and nonautistic children. PMID- 12108619 TI - A factor analytic study of the Autism Behavior Checklist. AB - The factor structure of the Autism Behavior Checklist (ABC) (Krug, Arick, & Almond, 1980a, 1980b), a 57-item screening instrument for autism, was examined on a sample of 383 individuals with autism spectrum disorders (i.e., autistic disorder, Asperger syndrome, and other autism-like conditions) aged 5-22 years. A five-factor model accounted for 80% of the total variance in the checklist. Thirty-nine of the 57 items had factor loadings of 0.4 or more, with 13 items loading on Factor 1, 11 items on Factor 2, 6 items on Factor 3, 5 items on Factor 4, and 4 items on Factor 5. No support was found for classifying the 57 items into the five subscales proposed by Krug et al. (1980a, 1980b) or for the three factor solution suggested by Wadden, Bryson, and Rodger (1991). PMID- 12108620 TI - Reading the mind in the voice: a study with normal adults and adults with Asperger syndrome and high functioning autism. AB - People with high functioning autism (HFA) and Asperger syndrome (AS) have deficits in theory of mind (ToM). Traditional ToM tasks are not sensitive enough to measure ToM deficits in adults, so more subtle ToM tests are needed. One adult level test, the Reading the Mind in the Eyes test has shown that AS and HFA subjects have measurable deficits in the ability to make ToM inferences. Here we introduce a test that extends the above task into the auditory domain and that can be used with adults with IQ Scores in the normal range. We report the use of the test with an adult sample of people with AS/HFA and with two adult control groups. Results suggest that individuals with AS/HFA have difficulty extracting mental state information from vocalizations. These results are consistent with previous results suggesting that people with HFA and AS have difficulties drawing ToM inferences. PMID- 12108621 TI - Changes in social impairment for people with intellectual disabilities: a follow up of the Camberwell cohort. AB - The skills and social impairments of a total population of children with severe intellectual disabilities and/or autism from Camberwell, South London (Wing and Gould, 1978 and 1979), were assessed using the Handicaps, Behaviours and Skills schedule, and they were reassessed when they were adolescents and young adults (Shah, 1986). Changes in social impairment over time are presented here. As Shah (1986) had found with a smaller sample, social impairment remained relatively stable over time: on a simple "socially impaired" versus "sociable" dichotomous grouping, 93% did not change social group. Within the socially impaired group, there was a significant increase in impairment over time (i.e., people who were passive at Time 1, were aloof at Time 2). Implications of these results and predictions for a further follow-up study are discussed. PMID- 12108622 TI - The changing prevalence of autism in California. AB - We conducted a population-based study of eight successive California births cohorts to examine the degree to which improvements in detection and changes in diagnosis contribute to the observed increase in autism prevalence. Children born in 1987-1994 who had autism were identified from the statewide agency responsible for coordinating services for individuals with developmental disabilities. To evaluate the role of diagnostic substitution, trends in prevalence of mental retardation without autism were also investigated. A total of 5038 children with full syndrome autism were identified from 4,590,333 California births, a prevalence of 11.0 per 10,000. During the study period, prevalence increased from 5.8 to 14.9 per 10,000, for an absolute change of 9.1 per 10,000. The pattern of increase was not influenced by maternal age, race/ethnicity, education, child gender, or plurality. During the same period, the prevalence of mental retardation without autism decreased from 28.8 to 19.5 per 10,000, for an absolute change of 9.3 per 10,000. These data suggest that improvements in detection and changes in diagnosis account for the observed increase in autism; whether there has also been a true increase in incidence is not known. PMID- 12108623 TI - Descriptive epidemiology of autism in a California population: who is at risk? AB - We investigated the association between selected infant and maternal characteristics and autism risk. Children with autism born in California in 1989 1994 were identified through service agency records and compared with the total population of California live births for selected characteristics recorded on the birth certificate. Multivariate models were used to generate adjusted risk estimates. From a live birth population of more than 3.5 million, 4381 children with autism were identified. Increased risks were observed for males, multiple births, and children born to black mothers. Risk increased as maternal age and maternal education increased. Children born to immigrant mothers had similar or decreased risk compared with California-born mothers. Environmental factors associated with these demographic characteristics may interact with genetic vulnerability to increase the risk of autism. PMID- 12108624 TI - Brief report: increasing communication skills for an elementary-aged student with autism using the Picture Exchange Communication System. AB - The purpose of this study was to examine the effects of the Picture Exchange Communication System (PECS) on the spontaneous communication skills of a 6-year old girl with autism across her home and school environments. The effects of the PECS were also examined for social interaction. Results indicated increases in spontaneous language (i.e., requests and comments) including use of the icons and verbalizations across those settings in which PECS was implemented. Intelligible verbalizations increased in two of three settings, and changes in peer social interaction were noted in one of the two school settings. PMID- 12108625 TI - Brief report: postural reactivity to fast visual motion differentiates autistic from children with Asperger syndrome. AB - The aim of the present study was to search for a sensorimotor marker (i.e., visuopostural tuning) that could be correlated with the severity of motor impairments in children with autistic spectrum disorders. Given that autistic children were previously reported to be posturally hyporeactive to visually perceived environmental motion in comparison with normal control children (Gepner et al., 1995), we sought to determine whether children with Asperger syndrome (AS) would share the same postural hyporeactivity to visual motion. Three autistic children with mild to severe motor impairments, three AS children with soft motor signs, and nine normal control children were tested for overall postural instability and postural reactivity to environmental motion. Results indicate, first, that overall postural instability is significantly reduced in autistic children compared with both AS and normal children. Second, although postural oscillations in the fore-aft axis become more attuned to the oscillation frequency of an immersive dynamic visual display as visual speed is increased, in both control and AS subjects, this is not the case in autistic children. Despite the small number of subjects tested in this study, our data confirm the existence of a visuopostural detuning in autistic children. Third, they argue for a correlation between visuopostural tuning and severity of motor signs in children with autistic spectrum disorders. Finally, they suggest a differentiation between children with autism and children with AS with regard to postural reactivity to fast visual motion. Neurophysiological implications of these results are discussed. In particular, a visuocerebellar pathway deficit hypothesis in autism is proposed. PMID- 12108626 TI - Is exposure to alcohol during pregnancy a risk factor for autism? PMID- 12108627 TI - Structure-chiroptical properties relationship in oxabicyclic beta-lactam derivatives. AB - The relationship between molecular structure of 5-dethia-5-oxacephams and clavams and their chiroptical properties was investigated by means of X-ray diffraction analysis, molecular modeling calculations and circular dichroism spectroscopy. It was found that the amide chromophore of the beta-lactam unit in these compounds is nonplanar with nitrogen atom having a pyramidal configuration. It was also found that the helicity of the lactam moiety in investigated oxacephams and clavams is controlled by the absolute configuration at the C-6 and C-5 carbon atom, respectively. Thus, the applicability of helicity rule correlating a positive (negative) torsional angle of the beta-lactam subunit O=C-N-C with a negative (positive) sign of the n-->pi* CE, previously applied to oxacephams, is now extended to clavams. PMID- 12108628 TI - The first circular dichroism observation for organic radical cations: chiroptical properties of neomenthyloxy- and isobornyloxyanisole radical cations. AB - The first unequivocal circular dichroism spectra were obtained for organic radical cations, which were prepared from p-(1S,2S,5R)-neomenthyloxyanisole (1) and p-(1S,2S,4S)-isobornyloxyanisole (2) by single electron oxidation with triethyloxonium hexachloroantimonate in dichloromethane. The chiroptical properties of the radical cations 1*+ and 2*+ were discussed in comparison with those of neutral species 1 and 2. PMID- 12108629 TI - Strategic placement of stereogenic centers in molecular materials for second harmonic generation. AB - Basic aspects of the nonlinear optical phenomenon of second harmonic generation (SHG) and the assembly of molecular materials for SHG are reviewed. Extensive use of chirality as a convenient tool to generate noncentrosymmetricity in molecular lattices, an essential requirement for the development of quadratic nonlinear optical materials, is noted. An overview of our investigations of chiral diaminodicyanoquinodimethanes is presented, which provides insight into a systematic approach to the effective deployment of chirality to achieve optimal molecular orientations for enhanced solid state SHG. Extension of these ideas to the realization of strong SHG in materials based on helical superstructures is outlined. PMID- 12108630 TI - Synthesis of calix[4]arene and porphyrin tethering four chiral five-membered cyclic carbonates. AB - Calix[4]arene tethering four chiral five-membered cyclic carbonates 1c was synthesized by O-glycidylation of p-tert-butylcalix[4]arene 1a with (R)-glycidyl tosylate, followed by carbonation with carbon dioxide catalyzed by lithium bromide. Porphyrin tethering chiral five-membered cyclic carbonates 2c was similarly synthesized. PMID- 12108631 TI - Straightforward synthesis of two pairs of enantiomeric butenolides 3-tetradecyl- and 3-hexadecyl-5-methyl-2(5H)-furanone. AB - Based on the synthetic method of optically active 3-alkyl-5-methyl-2(5H) furanones from lactic acid, two pairs of chiral butenolides 3-tetradecyl- and 3 hexadecyl-5-methyl-2(5H)-furanone have been straightforwardly synthesized from methyl stearate and methyl palmitate respectively by aldol condensation with (R)- or (S)-O-tetrahydropyranyl lactal prepared from corresponding ethyl lactate and beta-elimination. PMID- 12108632 TI - Evaluation of the origins of the selectivity of polymers imprinted with a HIV protease inhibitor using infrared spectroscopy and high performance liquid chromatography. AB - This work investigates the origins of enantioselectivity of polymers imprinted with the HIV protease inhibitor, Indinavir. For the preparation of imprints of the drug, the critical interactions between the functional monomer, methacrylic acid, and Indinavir were characterized by infrared (IR) spectroscopy to explore the optimum functional monomer concentration for the polymerization. It was shown that a polymer with high selectivity and minimum non-selective binding for Indinavir was obtained when prepared with enough functional monomer to hydrogen bond with all of the functional groups of the drug without using an excess of monomer. This observation is explained in terms of a balance that is achieved in the monomer-template equilibrium during the polymerization that yields a polymer with highly selective sites and minimal non-selective sites. This paper further demonstrates that IR spectroscopy can be a valuable tool in the design and syntheses of molecular imprinted polymers. PMID- 12108633 TI - Separation, conformation in solution and absolute configuration of ethopropazine enantiomers. AB - Enantiomers of ethopropazine x HCl (10-(2-diethylaminopropyl)phenothiazine hydrochloride) were prepared by fractional crystallization of diastereomeric dibenzoyltartaric acid salts, and their optical purity (enantiomeric excess, ee) determined by HPLC on Chiralcel OJ column. With a solvent mixture n-hexane/t butanol/triethylamine (100:3:0.5) as eluent a very good enantioseparation (alpha = 1.68) for racemic ethopropazine was obtained. Enantiomeric purity for (-) enantiomer was 99.1% and for (+)-enantiomer 97.9%. Combined data from NMR and CD spectra of both enantiomers, along with previously reported X-ray structure analyses of racemic ethopropazine, revealed skewed conformation of tricyclic system in solution, and (S)-configuration on the stereogenic center for (-) enantiomer, and (R)-configuration for (+)-enantiomer. PMID- 12108634 TI - CD of single-stranded, double-stranded, and G-quartet nucleic acids in complexes with a single-stranded DNA-binding protein. AB - We review CD studies of a single-stranded DNA binding protein, g5p, of the Ff group of bacterial viruses. The CD spectrum of the g5p is dominated by a positive tyrosine La band at 229 nm, to which all five of the protein tyrosines contribute. The La band becomes much less positive upon binding of g5p to nucleic acids. CD spectra of mutant proteins identified a single tyrosine, Y34, that is largely responsible for this CD perturbation. At >250 nm, CD perturbations of nucleic acids can be monitored during g5p binding, and CD titrations have identified two distinct modes of binding of the g5p at physiological ionic strength (0.2 M NaCl). SELEX selection of sequences bound preferentially by g5p yielded a G-rich sequence that is closely related to telomere sequences and has CD properties of a G-tetraplex. CD spectroscopy showed that the presumed G quadruplex form is maintained within saturated g5p x DNA complexes. PMID- 12108635 TI - Chiral inclusion crystals constructed with achiral host and achiral guest molecules. AB - Achiral molecules of tetrakis(p-bromophenyl)ethylene and 2,3,6,7,10,11 hexahydroxy triphenylene were found to arrange in a chiral form in their inclusion crystal with an achiral guest molecule. The chiral arrangement of these achiral molecules was studied by X-ray analysis and CD spectral measurement in the solid state. PMID- 12108636 TI - The supramolecular structure of self-assembly formed by capsanthin derivatives. AB - Carotenoids form structured self-assembly upon aqueous dilution of their organic solutions. In order to test the proposal predicting carotenoid aggregates to be organized in closely packed H-type (card-pack) manner by intermolecular hydrogen bonds, the trihydroxy derivative of capsanthin (1), (6'R)-capsanthol (2) ((all E,3R,3'S,5'R,6'R)-beta,kappa-carotene-3,3',6'-triol) was acetylated to obtain all varieties of mono-, di- and triacetates and the corresponding supramolecules were studied by UV/Vis- and CD spectroscopy. It was verified that derivatives lacking hydroxyl functions at either of the end-groups form the loosely organized J-type (head-to-tail) aggregates. A model for the structure of the J-type self-assembly is proposed. Evidence was also obtained suggesting that close contacts of carotenoid molecules are not confined to hydrogen bonding. PMID- 12108638 TI - The use of benzamide derivatives of secondary amines for stereochemical studies by circular dichroism. AB - The benzamide chromophore is widely used as a Cottonogenic derivative of primary amines for stereochemical studies by circular dichroism. The assignments based on the exciton chirality method are reliable since the benzamide group has well defined geometry and conformation. A recent report (J.D. Chisholm, J. Golik, B. Krishnan, J.A. Matson, D.L. Van Vranken, J. Am. Chem. Soc. 1999, 121: 3801-3802) claimed a caveat in the application of the exciton chirality method to benzamides derived from secondary amines. By the use of benzoyl derivatives of amino alcohols (1-4) and diamines (5, 6) of known absolute configuration we demonstrate that the 250-210 nm range exciton Cotton effects due to secondary and tertiary benzamides are generally of opposite sign. The origin of such disparity is traced to different conformational equilibria of the amide C-N bond in secondary and tertiary benzamides, as shown by semiempirical molecular modelling and NMR data. This feature can be useful in the determination of absolute configuration by analysis of the CD spectra due to exciton coupling of tertiary benzamides. PMID- 12108637 TI - Determination of the absolute configuration of rubroflavin by comparison of measured and calculated CD spectra of its thermolysis product 3-methanesulfinyl-5 methylmercaptophenol. AB - The absolute configuration of rubroflavin has been determined indirectly by comparison of the measured and the calculated CD spectrum of its thermal decomposition product 3-methanesulfinyl-5-methylmercaptophenol. Performing geometry optimizations at the HF/6-31+G* level we found fifteen local minima for the (R)-isomer of 3-methanesulfinyl-5-methylmercaptophenol. The CD spectrum of the compound was then obtained as a superposition of the Boltzmann-weighted spectra for each structure calculated with the non-empirical CIS method. The corresponding Boltzmann factors have been calculated employing the relative energies of these minima determined at the ZPE + MP2/6-31+G*//HF/6-31+G* level of ab initio theory. Comparing the signs of the observed and calculated longest wave length Cotton effect we assign an absolute configuration to the thermolysis product. Since additional calculations revealed that the tricoordinate sulfur atom in rubroflavin and in its decomposition product is configurationally stable under the conditions of thermolysis we conclude that the absolute configuation at the corresponding sulfur atom of rubroflavin is the same. PMID- 12108639 TI - Synthesis and characterization of new chiral calix[4]arenes. AB - The unique chiral calixarenes were successfully synthesized by the following two methods: the Williamson ether synthesis and a stepwise ether cleavage with a mild Lewis acid. The high regioselectivity is recognized by the latter stepwise method. The ionophores having oligoethylene glycol unit efficiently extracted larger alkali metal ions like K+, Rb+, and Cs+ than smaller ones like Li+ and Na+. Their ion selectivity apparently changed by the chain length of crown ether. All racemates obtained could be resolved to each enantiomer by HPLC using a chiral column. The calixarenes with planar chirality recognized the chirality of guest molecules. Thus, the (-)-receptor resolved strongly forms 1:1 complex with (R)-(+)-alpha-phenylethylammonium picrate. PMID- 12108640 TI - Rapid and direct determination of iodide in seawater by electrostatic ion chromatography. AB - An electrostatic ion chromatographic (IC) method for rapid and direct determination of iodide in seawater is reported. Separation was achieved using a reversed-phase ODS packed column (250x4.6 mm I.D.) modified by coating with Zwittergent-3-14 micelles, with an eluent comprising an aqueous solution containing 0.2 mM NaClO4 and 0.3 mM Zwittergent-3-14 and using UV detection at 210 nm. Samples prepared by dissolving NaIO3, NaNO2, NaBr, NaBrO3, NaNO3, NaI, and NaSCN in artificial or real seawaters were analyzed using this IC system. Nitrite, iodate, bromide, bromate, and nitrate showed very little or no retention, while iodide and thiocyanate were well separated, being eluted within 6 and 16 min, respectively. The detection limit for iodide obtained by injecting 400 microL of sample was 0.011 microM (S/N = 3), and the precision values obtained by analyzing samples containing 0.1 or 0.3 microM iodide in real seawater samples were 2.3% RSD and 1.2% RSD, respectively. Direct determination of iodide in real seawater samples was possible using this proposed IC system. PMID- 12108641 TI - Ion chromatographic analysis of anions in ammonium hydroxide, hydrofluoric acid, and slurries, used in semiconductor processing. AB - In this study, ion chromatography (IC) with suppressed conductivity detection was used for the determination of trace anions in 29% (w/w) ammonium hydroxide, 49% (w/w) hydrofluoric acid and slurries. For these samples, various sample pretreatment methods were applied to eliminate matrix interferences. For concentrated ammonium hydroxide, an on-line electrochemical neutralizer (SP10 AutoNeutralization module) was used to neutralize the base prior to the IC analysis. For concentrated hydrofluoric acid, a heart cutting technique with an ion-exclusion column was used to separate the anions of interest prior to an IC separation. A method was also developed to analyze chloride in silica slurries by IC. PMID- 12108642 TI - Automated trace anion determinations in concentrated electronic grade phosphoric acid by ion chromatography. AB - Modifications have been made to the method of ion-exclusion pre-separation followed by ion exchange with conductivity detection for the determination of trace levels of chloride, sulfate and nitrate in concentrated phosphoric acid. Ion-exclusion separation and pre-concentration of impurity anions is performed using Dionex AS6-ICE and AS11-HC (4 mm) columns, respectively, with water eluent. Final separation is performed using Dionex AG11-HC and AS11-HC (2 mm) columns, KOH gradient elution, and suppressed conductivity detection. Improvements to the method include addition of an autosampler and eluent generator, and use of external standard calibration. These instrumental and procedural changes significantly improve the method's throughput, while the method's capability relative to phosphoric acid specifications is maintained, as verified through statistical evaluation of control sample analyses. Detection limits of 60, 680, and 4,0 ppb (w/w) are obtained vs. semiconductor-grade phosphoric acid specifications of 1000, 12,000, and 5000 ppb for chloride, sulfate, and nitrate, respectively. PMID- 12108643 TI - Determination of anions at trace levels in power plant water samples by ion chromatography with electrolytic eluent generation and suppression. AB - An ion chromatographic method was developed for the determination of nine inorganic and organic acid anions at sub- to low-microg/l levels in power plant water samples. In this method, samples were injected using a large-volume direct injection technique, the analyte anions were separated on a hydroxide-selective anion-exchange column using high-purity hydroxide eluents generated by an on-line electrolytic eluent generator and detected using the suppressed conductivity detection method. The method performance was evaluated by analyzing synthetic water samples containing additives encountered in the power plant water samples and four water samples from a fossil fuel power plant. The relative standard deviations of retention times of analyte ions separated on the hydroxide selective anion-exchange column were less than 0.4%. The recoveries of analyte ions spiked into the synthetic water samples at concentrations of 0.13-1.0 microg/l were in the range of 70-120%. The method detection limits for analyte ions in deionized water were 0.0099, 0.0056, 0.019, 0.057, 0.0084, 0.023, 0.067, 0.037, and 0.079 microg/l for fluoride, acetate, formate, chloride, nitrite, sulfate, bromide, nitrate, and phosphate, respectively. PMID- 12108644 TI - Determination of total acidity and of divalent cations by ion chromatography with n-hexadecylphosphocholine as the stationary phase. AB - An ion chromatographic (IC) method is reported for simultaneous determination of total acidity (H+), Ba2+, Ca2+, and Mg2+ in aqueous samples. A standard ODS silica column modified by coating with n-hexadecylphosphocholine was used as the separation column. Water alone was used as the eluent, with conductivity detection of the sample ions. An excess of sodium iodide was added to each sample so that both H+ and divalent cations were always eluted with iodide as the counterion. The elution order was Ba2+, Mg2+, Ca2+, and H+ with H+ being eluted much later than the divalent cations. Acetic acid and several other weak acids could also be separated because all the protons were transposed from acetic acid (HAc) to HI by the sodium iodide. Detection limits for 100 microl injection, S/N=3 were in the low micromolar range for the divalent cations and approximately 0.3 mM for H+/I-. This method was used successfully for simultaneous determination of total acidity, magnesium and calcium in HCl-type of hot-spring water. PMID- 12108645 TI - Simultaneous separation of common mono- and divalent cations on a calcinated silica gel column by ion chromatography with indirect photometric detection and aromatic monoamines-oxalic acid, containing crown ethers, used as eluent. AB - The application of unmodified silica gel (Super Micro Bead Silica Gel B-5, SMBSG B-5) as a cation-exchange stationary phase in ion chromatography with indirect photometric detection (IC-IPD) for the separation of common mono- and divalent cations (Li+, Na+, NH4+, K+, Mg2+ and Ca2+) was carried out using various aromatic monoamines [tyramine [4-(2-aminoethyl)phenol], benzylamine, phenylethylamine, 2-methylpyridine and 2,6-dimethylpyridine] as eluents. When using these amines as eluents, the peak resolution between these mono- and divalent cations was not quite satisfactory and the peak shapes of NH4+ and K+ were largely destroyed on the SMBSG B-5 silica gel column. Hence, the application of SMBSG B-5 silica gel calcinated at 200, 400, 600, 800 and 1000 degrees C for 5 h in the IC-IPD was carried out. The peak shapes of the monovalent cations were greatly improved with increasing calcination temperature and, as a result, symmetrical peaks of these mono- and divalent cations were obtained on the SMBSG B-5 silica gel calcinated at 1000 degrees C as the stationary phase. In contrast, the peak resolution between these mono- and divalent cations was not improved. Therefore, crown ethers [18-crown-6 (1,4,7,10,13,15-hexaoxacyclooctadecane), 15 crown-5 (1,4,7,10,13-pentaoxacyclopentadecane)] were added to the eluent for the complete separation of these mono- and divalent cations. Excellent simultaneous separation and highly sensitive detection at 275 nm were achieved in 25 min on a column (150x4.6 mm I.D.) packed with SMBSG B-5 silica gel calcinated at 1000 degrees C by elution with 0.75 mM tyramine-0.25 mM oxalic acid at pH 5.0 containing either 1.0 mM 18-crown-6 or 10 mM 15-crown-5. PMID- 12108646 TI - Recent advances in ion chromatography suppressor improve anion separation and detection. AB - The Alltech DS-Plus suppressor improves and simplifies anion analysis by suppressor-based ion chromatography (IC). In addition to suppressing the mobile phase and enhancing the analyte signal, the DS-Plus suppressor removes carbonic acid from the suppressor effluent when carbonate-hydrogencarbonate mobile phases are used, reducing the water dip that interferes with early eluting peaks, increasing detection sensitivity, and enabling carbonate-hydrogencarbonate gradients. The ability to operate with carbonate-hydrogencarbonate gradients places exceptional separating power in the mobile phase, forgoing the need for expensive columns with special selectivity. Continuous operation and solid-phase chemistry eliminates the need for external regenerants, switching valves, and multiple operating modes. PMID- 12108647 TI - Retention behavior of common mono- and divalent cations on calcinated silica gel columns in ion chromatography with conductimetric detection and the use of nitric acid, containing crown ethers, as eluents. AB - Ion chromatographic behavior of common mono- and divalent cations (Li+, Na+, NH4+, K+, Mg2+ and Ca2+) on columns packed with silica gels (Super Micro Bead Silica Gel B-5, SMBSG B-5) calcinated at 200, 400, 600, 800 and 1000 degrees C for 5 h was investigated using nitric acid containing crown ethers [18-crown-6 (1,4,7,10,13,15-hexaoxacyclooctadecane) and 15-crown-5 (1,4,7,10,13 pentaoxacyclopentadecane)] as eluent. When using 0.5 mM HNO3 as the eluent, the calcination had almost no effect on the improvement of peak resolution between these mono- and divalent cations. In contrast, when using 0.5 mM HNO3 containing crown ethers as the eluent, with increasing the calcinating temperature, the amount of crown ethers adsorbed on the corresponding calcinated SMBSG B-5 silica gels columns increased and, as a consequence, peak resolution between these mono- and divalent cations was quite improved. Excellent simultaneous separation of these mono- and divalent cations was achieved on column (150x4.6 mm I.D.) packed with the SMBSG B-5 silica gel calcinated at 1000 degrees C by elution with 0.5 mM HNO3 containing either 1.0 mM 18-crown-6 or 5.0 mM 15-crown-5. PMID- 12108648 TI - Simultaneous separation of common mono- and divalent cations on an acid-treated silica gel column by ion chromatography with indirect photometric detection and tyramine-oxalic acid, containing 18-crown-6 as eluent. AB - The application of unmodified silica gel (Super Micro Bead Silica Gel B-5, SMBSG B-5) as cation-exchange stationary phase in ion chromatography with indirect photometric detection for common mono- and divalent cations (Li+, Na+, NH4+, K+, Mg2+ and Ca2+) was carried out using 0.75 mM tyramine [4-(2-aminoethyl)phenol] 0.25 mM oxalic acid at pH 5.0 as the eluent. Although complete group separation between these mono- and divalent cations was achieved on the SMBSG B-5 column (150x4.6 mm I.D.) in 12 min, peak shapes of NH4+ and K+ were strongly tailed. Hence, the application of SMBSG B-5 silica gel treated with conc. hydrochloric acid at reflux-temperature for 12 h for the simultaneous separation of these cations was carried out. Although the retention volumes of these cations decreased on the acid-treated SMBSG B-5 silica gel column, the peak shapes of NH4+ and K+ were quite improved. Excellent simultaneous separation and highly sensitive detection at 275 nm [detection limits (signal-to-noise ratio of 3 and injection volume of 20 microl), 0.34 microM for Li+, 0.47 microM for Na+, 0.39 microM for NH4+, 0.59 microM for K+, 0.24 microM for Mg2+ and 0.28 microM for Ca2+] were achieved in 15 min on the acid-treated SMBSG B-5 column using 0.5 mM tyramine-0.2 mM oxalic acid-10 mM 18-crown-6 (1,4,7,10,13,15 hexaoxacyclooctadecane) at pH 5.5 as the eluent. PMID- 12108649 TI - Determination of inorganic cations and ammonium in environmental waters by ion chromatography with a high-capacity cation-exchange column. AB - While alkali and alkaline earth cations are commonly determined by using spectrometric techniques such as atomic absorption spectrometry or inductively coupled plasma, ammonium cation in the same sample must be measured separately by a wet chemical technique such as colorimetry, titrimetry, or ammonia-selective electrode. In a single 25-min run ion chromatography can determine all of the important inorganic cations including lithium, sodium, ammonium, potassium, magnesium and calcium. In this paper, we describe the use of ion chromatography with a new high-capacity cation-exchange column (the IonPac CS16), an electrolytically-generated methanesulfonic acid eluent and suppressed conductivity detection to determine dissolved alkali and alkaline earth cations and ammonium in drinking water wastewater and aqueous soil extracts. The IonPac CS16 is a high-capacity cation-exchange column that incorporates recent advances in polymer chemistry to enable trace-level determinations of cations even in high ionic-strength matrices. We discuss the linear range, method detection limits, and analyte recoveries obtained with this column, and evaluate the effect of potential interferences on method performance during the analysis of typical environmental samples. PMID- 12108650 TI - Applications of ion chromatography in cane sugar research and process problems. AB - Ion chromatography (IC) offers the sugar technologist a simple, reliable technique for the simultaneous separation and determination of inorganic and organic ions in complex process mixtures. Identification and measurement of ions present in sugar liquors are important factors in understanding and maximising sugar recovery. Choice of IC column packing, eluent composition and pH, and detection mode (suppressed and non-suppressed) are shown to be useful variables when analysing sugar factory products, especially the multivalent ions such as phosphate and organic acids (aconitic and oxalic acid). The IC methods show good agreement with conventional analysis methods (potentiometric titration, atomic emission and absorption). IC also produced excellent repeatability and recovery from these matrices. Examples of the application of IC analysis in applied sugar research and processing problems include clarification studies, degradation of molasses, sugar solution colour removal and factory process problems. PMID- 12108651 TI - Qualitative analysis of some carboxylic acids by ion-exclusion chromatography with atmospheric pressure chemical ionization mass spectrometric detection. AB - A simple, selective and sensitive method for the determination of carboxylic acids has been developed. A mixture of formic, acetic, propionic, valeric, isovaleric, isobutyric, and isocaproic acids has been separated on a polymethacrylate-based weak acidic cation-exchange resin (TSK gel OA pak-A) based on an ion-exclusion chromatographic mechanism with detection using UV-photodiode array, conductivity and atmospheric pressure chemical ionization mass spectrometry (APCI-MS). A mobile phase consisting of 0.85 mM benzoic acid in 10% aqueous methanol (pH 3.89) was used to separate the above carboxylic acids in about 40 min. For LC-MS, the APCI interface was used in the negative ionization mode. Linear plots of peak area versus concentration were obtained over the range 1-30 mM (r2=0.9982) and 1-30 mM (r2=0.9958) for conductimetric and MS detection, respectively. The detection limits of the target carboxylic acids calculated at S/N=3 ranged from 0.078 to 2.3 microM for conductimetric and photometric detection and from 0.66 to 3.82 microM for ion-exclusion chromatography-APCI-MS. The reproducibility of retention times was 0.12-0.16% relative standard deviation for ion-exclusion chromatography and 1.21-2.5% for ion-exclusion chromatography APCI-MS. The method was applied to the determination of carboxylic acids in red wine, white wine, apple vinegar, and Japanese rice wine. PMID- 12108652 TI - Vacancy ion-exclusion chromatography of carboxylic acids on a weakly acidic cation-exchange resin. AB - In this preliminary study, a new approach to ion-exclusion chromatography is proposed to overcome the relatively poor conductivity detection response which occurs in ion-exclusion chromatography when acids are added to the eluent in order to improve peak shape. This approach, termed vacancy ion-exclusion chromatography, requires the sample to be used as eluent and a sample of water to be injected onto a weakly acidic cation-exchange column (TSKgel OApak-A). Vacancy peaks for each of the analytes appear at the retention times of these analytes. Highly sensitive conductivity detection is possible and sharp, well-shaped peaks are produced, leading to efficient separations. Retention times were found to be affected by the concentration of the analytes in the eluent, and also by the presence of an organic modifier such as methanol in the eluent. Detection limits for oxalic, formic, acetic, propionic, butyric and valeric acids were 0.1, 0.2, 0.3, 0.3, 0.4 and 0.5 microM, respectively, and linear ranges for some acids extended over two orders of magnitude. Precision values for retention times were 0.21% and for peak areas were <1.90%. The vacancy ion-exclusion chromatography method was found to give detection responses four to 10 times higher than conventional ion-exclusion chromatography using sulfuric acid eluent and two to five times higher than conventional ion-exclusion chromatography using benzoic acid eluent. PMID- 12108653 TI - Organic analysis by ion chromatography. 1. Determination of aromatic amines and aromatic diisocyanates by cation-exchange chromatography with amperometric detection. AB - A method has been developed for the simultaneous determination of a range of aromatic amines using cation-exchange chromatography performed on a standard ion chromatography column using d.c. amperometric detection. The analytes separated were 2,4- and 2,6-toluenediamine (2,4- and 2,6-TDA), aniline, o-toluidine, benzidine, p-chloroaniline, 4,4'-diaminodiphenyl (4,4'-DDP), m-nitroaniline and 1 naphthylamine. A Dionex CS12 column was used with gradient elution from an initial eluent of 5% CH3CN+35 mM H2SO4 to 27% CH3CN+35 mM H2SO4 (at 35 min). Detection limits in the range 2.6-22.6 microg/l were observed for all analytes except m-nitroaniline, for which the detection limit was 201 microg/l. Linear calibrations and good precision were observed and the method was applied to the determination of benzidine, p-chloroaniline and 1-naphthylamine in wastewater samples. Further, the separation was also used (after some modification of the eluent conditions) for the determination of 2,4- and 2,6-toluene diisocyanate (2,4- and 2,6-TDI) and 4,4'-methylenediphenyl diisocyanate (4,4'-MDI) after their hydrolysis to 2,4-TDA, 2,6-TDA and 4,4'-DDP. Detection limits for 2,6- and 2,4 TDI and 4,4'-MDI were 3.8, 8.2, and 11.2 microg/l, respectively. The method was applied to the determination of diisocyanates in air. PMID- 12108654 TI - Determination of vanadium as 4-(2-pyridylazo)resorcinol-hydrogen peroxide ternary complexes by ion-interaction reversed-phase liquid chromatography. AB - The separation and determination of the vanadium(V) ternary complex formed with 4 (2-pyridylazo)resorcinol (PAR) and hydrogen peroxide using ion-interaction reversed-phase high-performance liquid chromatography on a C18 column has been investigated. The optimal mobile phase was a methanol-water solution (32:68, v/v) containing 3 mM tetrabutylammonium bromide, 5 mM acetic acid and 5 mM citrate buffer at pH 7, with absorbance detection at 540 nm. The stoichiometry of the ternary complex of vanadium at pH 6 in 10 mM acetate buffer using the mole ratio and Job's method by HPLC indicated that the mole ratio of V(V):PAR:H2O2 was 1:1:1. The optimal conditions for precolumn formation of the ternary complex were 10 mM acetate, 7 mM H2O2, 0.3 mM PAR, and pH 6. The method gave relative standard deviations of retention time, peak area and peak height for the ternary complex of 0.187, 0.45 and 0.57%, respectively. The detection limit (at a signal-to-noise ratio of 3) for V(V) was 0.09 ng/ml in the digested sample using a 100-microl injection loop (or 0.09 microg/g in the solid fertiliser sample). The method was applied to the analysis of fertilisers (phosphate rocks and nitrogen, phosphorus and potassium fertiliser). The results for vanadium obtained by the HPLC method agreed well with those from magnetic sector inductively coupled plasma MS analysis. PMID- 12108655 TI - Anion-exchange chromatography of metal cyanide complexes with gradient separation and direct UV detection. AB - In the regulatory analysis of water samples, cyanide content is usually expressed in various forms as free cyanide, total cyanide, weak-acid dissociable cyanide, available cyanide, and cyanide amenable to chlorination. Concentration of individual metal cyanide complex, not furnished in any of these methods, is useful in meaningful assessment of toxicity due to cyanide. In the present work, two macroporous anion-exchange columns, having high and intermediate hydrophobicity, were evaluated to achieve separation of cyano complexes of silver, iron, gold, copper, nickel, and cobalt. On the QS-Al SC column, of high hydrophobicity, the monovalent cyano complexes of silver and gold eluted last while the multivalent cyano complexes, e.g. iron and copper, eluted early. It is suggested that the retention order on this column is due to relative hydrophobicity of the metal cyanide complex, and its affect on ion exchange. In contrast, on the QS-A2 SC column of intermediate hydrophobicity, with the exception of cyano complex with Fe, the separation of the cyano complexes of five other metals closely followed an anion-exchange mechanism. Under gradient conditions, the six metal cyanide complexes were well resolved on the QS-A2 SC column and the method with direct UV detection at 215 nm was accurate (spike recovery of 99.8-118.8%) and precise (RSD of 1.0-2.6%). PMID- 12108656 TI - Reproducible digestion method for ion-chromatographic analysis of anions in 30% hydrogen peroxide. AB - Semiconductor-grade hydrogen peroxide (30%) is analyzed for anion contamination to be certain the levels of each analyte do not exceed 30 ppb (w/w). This paper presents a reproducible, platinum-decomposition approach that uses ion chromatography to quantify the various analytes (fluoride, chloride, sulfate, bromide, nitrate, and phosphate). Important to the success of the method are: (1) use of disposable HDPE bottles for the digestion, (2) immersion of the bottles in a water bath, (3) careful re-mixing of the digesting peroxide after two (of six total) hours, and (4) careful clean-up and sample-handling procedures (to avoid contamination). Except for fluoride and nitrate, all analytes exhibited recoveries from 89.6 to 98.3%, with +/- prediction intervals (at the 95% confidence level) from 1.5 to 3.0 ppb. Fluoride's recovery was low (74.9%), but reproducible (+/- prediction interval at 95% confidence=2.0 ppb). Nitrate reeovery was 99.1%, but noisy (+/- prediction interval at 95% confidence=8.7 ppb); this imprecision was thought to be due to contamination from atmospheric nitrite. A Dionex DX 500 microbore ion chromatograph with AS15 column and 1-ml sample loop were used for all determinations; detection was by conductivity. Statistical analyses were performed using JMP software. PMID- 12108657 TI - Determination of zwitterionic and cationic surfactants by high-performance liquid chromatography with chemiluminescenscent nitrogen detection. AB - The use of chemiluminescent nitrogen specific detection (CLND) combined with an HPLC separation allows for the identification and quantification of cationic and zwitterionic surfactants. The CLND provides equimolar responses, based on the amount of nitrogen within any compound. This allows for the detection of any nitrogen containing surfactant. Reversed-phase separation methods using cyano columns are developed for cationic and zwitterionic (sulfobetaine) surfactant mixtures. The limits of detection for these surfactants are in the single micromolar range (1 ng N). A linear response was obtained (R2=0.9981) between 50 microM and 5 mM. The methodology was then applied to the determination of an industrial zwitterionic surfactant, Rewoteric AM CAS U [coco(amidopropyl)hydroxyldimethylsulfobetaine]. PMID- 12108658 TI - Applications of ion chromatography with electrospray mass spectrometric detection to the determination of environmental contaminants in water. AB - Ion chromatography (IC) is widely used for the compliance monitoring of common inorganic anions in drinking water. However, there has recently been considerable interest in the development of IC methods to meet regulatory requirements for analytes other than common inorganic anions, including disinfection byproduct anions, perchlorate, and haloacetic acids. Many of these new methods require the use of large injection volumes, high capacity columns and analyte specific detection schemes, such as inductively coupled plasma mass spectrometry or postcolumn reaction with UV-Vis detection, in order to meet current regulatory objectives. Electrospray ionization mass spectrometry (ESI-MS) is a detection technique that is particularly suitable for the analysis of permanently ionized or polar, ionizable compounds. The combination of IC with MS detection is emerging as an important tool for the analysis of ionic compounds in drinking water, as it provides increased specificity and sensitivity compared to conductivity detection. This paper reports on the application of IC-ESI-MS for the confirmation and quantitation of environmentally significant contaminants, i.e. compounds with adverse health effects which are either regulated or being considered for regulation, such as bromate, perchlorate, haloacetic acids, and selenium species, in various water samples. PMID- 12108659 TI - Determination of hexavalent chromium at the level of the California Public Health Goal by ion chromatography. AB - Chromium is a primary drinking water contaminant in the USA with hexavalent chromium, Cr(VI), being the most toxic form of the metal. As a required step in developing a revised state drinking water standard for chromium, the California Department of Health Services recently issued a new Public Health Goal (PHG) of 2.5 microg/l for total chromium and 0.2 microg/l for Cr(VI). Hexavalent chromium can be determined (as chromate) by ion chromatography, as described in US Evironmental Protection Agency Method 218.6; however, the method as originally published does not allow sufficient sensitivity for analysis at the California PHG level of 0.2 microg/l. Modification of the conditions described in Method 218.6, including the use of a lower eluent flow-rate, larger reaction coil, and larger injection volume, significantly increases the method sensitivity. The modified method, which uses IonPac NG1 and AS7 guard and analytical columns, an eluent of 250 mM ammonium sulfate-100 mM ammonium hydroxide operated at 1.0 ml/min, a 1000 microl injection volume, and postcolumn reaction with 2 mM diphenylcarbazide-10% methanol-0.5 M sulfuric acid (using a 750 microl reaction coil) followed by UV-Vis detection at 530 nm, permits a method detection limit for chromate of 0.02 microg/l. This results in a quantitation limit of 0.06 microg/l, which is more than sufficient for analysis at the California PHG level. Calibration is linear over the range of 0.1-10 microg/l and quantitative recoveries (>80%) are obtained for chromate spiked at 0.2 microg/l in drinking water. The modified method provides acceptable performance, in terms of chromate peak shape and recovery, in the presence of up to 1000 mg/l chloride or 2000 mg/l sulfate. PMID- 12108660 TI - Ion chromatographic determination of trace hydroxylamine in waste streams generated by a pharmaceutical reaction process. AB - Hydroxylamine is a key raw material used in a synthetic drug process at Pharmacia. Since hydroxylamine is harmful to microorganisms, concentrations above 5 ppm could interfere with the biological sewage plant performance. This necessitated the development of a sensitive analytical method for detecting low levels of hydroxylamine in the waste streams generated from the pharmaceutical process. The present report describes a cation-exchange chromatographic method coupled with pulsed amperometric detection at a gold electrode for trace analysis of hydroxylamine. This method was evaluated by generating data on the parameters of specificity, precision, linearity, recovery and sensitivity. PMID- 12108661 TI - Semi-permanent surfactant coatings for inorganic anion analysis in capillary electrophoresis. AB - Capillary electrophoretic separations of inorganic anions are performed using a capillary coated with a mixture of the cationic surfactant didodecyldimethylammonium bromide (DDAB) and the zwitterionic surfactant 1,2 dilauroyl-sn-phosphatidylcholine (DLPC). These double-chained surfactants form semi-permanent coatings on the capillary wall, which allows the excess surfactant to be removed from the buffer prior to separation. Interactions between surfactant aggregates in the buffer and analyte anions are thus eliminated. The electroosmotic flow (EOF) can be altered from fully reversed (100% DDAB) to near zero (100% DLPC) using different ratios of DDAB and DLPC. Controlling the EOF allows for improved resolution of the anions while maintaining a rapid, co-EOF separation, free from analyte-surfactant additive interactions. PMID- 12108662 TI - Capillary electrophoretic analysis of inorganic anions in atmospheric hailstone samples. AB - A new application of capillary electrophoresis (CE) for measuring inorganic anions in hailstones was carried out. Five hailstone specimens were collected from large blocks of ice that fell in January 2000 in some provinces of Spain. Sample handling and preparation procedures were performed with care. CE analysis of anions was carried out using indirect UV detection at 254 nm with a negative power supply (-15 kV) and hydrostatic injection (10 cm for 30 s) at 35 degrees C. Anion separation was completed in less than 4 min. The working electrolyte consisted of 4.7 mM sodium chromate, 4.0 mM OFM-OH (tetradecyltrimethylammonium hydroxide), 10 mM CHES [2-(N-cyclohexylamino)ethanesulfonic acid], and 0.1 mM calcium gluconate (pH 9.1). Good repeatability of migration times after eight injections (<0.7% RSD), adequate linearity responses (r2>0.9) as well as satisfactory detection limits (<0.35 ppm) were achieved. The analytical results provided by CE were compared with those obtained by traditional wet-chemical (WCH) methods. Most of the results obtained by CE were consistent with those of WCH, except for one sample. PMID- 12108663 TI - Recent developments in ion chromatography. AB - This paper summarizes how ion chromatography is now a multimode technique suitable for solving analytical problems in all areas of interest. Current and more recent applications will be overviewed within the new trends. PMID- 12108664 TI - Adjustable-capacity anion-exchange separator. AB - A cryptand-based anion exchanger has been developed in which the capacity and to a lesser degree, selectivity are adjustable simply by the choice of the mobile phase. Although much work has been done in the past using cryptand-based anion exchangers, these stationary phases were based on adsorbed cryptands rather than covalently bound cryptands. These phases suffered from the usual problems associated with adsorbed systems. A novel styrene-based cryptand has been synthesized which can be covalently attached to a solid support. A brief review of cryptands and binding constants as well as comparisons of adsorbed phases versus covalently bound phases will be discussed. Some of the unique chromatographic properties of this prototype column will be illustrated as well. PMID- 12108665 TI - The membrane suppressor: a historical perspective. AB - The membrane suppressor for ion chromatography was first demonstrated in 1971. However, development of the membrane suppressor was delayed by the ease of preparation and ruggedness of a suppressor column. By 1981, a practical membrane suppressor had been developed but its excessive band broadening characteristics were not compatible with improved columns that were then being developed. The packed membrane suppressor solved the band broadening problem by 1982. PMID- 12108666 TI - New suppressor technology improves trace level anion analysis with carbonate hydrogencarbonate mobile phases. AB - Sodium carbonate-hydrogencarbonate mobile phases are preferred over sodium hydroxide for anion analysis by suppressor-based ion chromatography (IC). Unlike hydroxide, carbonate-hydrogencarbonate has strong eluting power and its buffering capacity can be used as a selectivity tool for controlling separations. However, carbonate-hydrogencarbonate mobile phases fell out of favor for trace level analysis because the carbonic acid suppressor effluent has some background conductivity, which reduces sensitivity compared to sodium hydroxide. This paper describes a new suppressor technology that improved anion analysis with carbonate hydrogencarbonate mobile phases. In addition to converting the carbonate hydrogencarbonate buffer to carbonic acid like other traditional IC suppressors, the new DS-Plus suppressor also removed carbonic acid from the suppressor effluent. Anions are now detected in water background, just like when using sodium hydroxide as the mobile phase. The lower background conductivity improves sensitivity and reduces detection limits. The water-dip often seen with conventional suppressors is greatly reduced, improving fluoride quantification. PMID- 12108667 TI - Ion chromatographic system with a novel switching suppression device. AB - Ion chromatography (IC) has been a powerful tool for measuring ionic species in environmental samples such as tap, river and drain waters. Suppressor modules (membrane and disposable column types) have been used for reducing the background of a baseline. A new type of suppressor device, which has a suppressor resin and switching valve was developed. Fresh ionic resin is introduced into a groove for each analysis to perform the suppression of the working eluent. The eluent composition for obtaining higher sensitivity and better resolutions among ionic species and carbonate ion was also investigated. Although carbonate buffers are used for ion separation in general, the separation of carbonate ion from other ions was not achieved. A borate eluent resulted in good resolutions and higher sensitivity. A new column was also developed for obtaining higher column efficiency and resolution. The optimization of anion separation using a new IC system (IC-2001) that consists of a new suppressor device, an anion-exchange column (TSKgel SuperIC-Anion, 150x4.6 mm), an autosampler, a conductivity cell and a pump in a compact module is described. PMID- 12108668 TI - New conductivity detection response equation for anions eluted with fully and partially ionised eluents in non-suppressed ion chromatography. AB - A response equation for conductivity detection in ion chromatography has been derived. This equation is applicable to non-suppressed ion chromatography using both fully ionised and partially ionised eluents. A prime assumption of this equation is that when partially ionised eluents are used (such as benzoic acid), both the ionised and neutral components of the eluent species contribute to the detector response of anionic analytes. Experimental evidence is provided to support this assumption in that pH changes accompanying the elution of an analyte have been measured. These pH changes are proportional to the concentration of analyte injected onto the column, in accordance with predictions from the response equation. Furthermore, it is shown that protonated eluents (such as benzoic acid) give more sensitive detection than equivalent ionised eluents (such as potassium benzoate) and the signal enhancement achieved using a protonated eluent species is in accordance with theory. PMID- 12108669 TI - Effects of common metal ions on the determination of anions by suppressed ion chromatography. AB - The effects of common metal ions (Na+, K+, Ca2+, Mg2+, Al3+ and Fe3+) on the determination of nitrate, phosphate and sulfate by suppressed ion chromatography were investigated. The responses of nitrate within 1-10 mg/l were not affected in the presence of six metal ions at 10 mg/l. The peak areas of 1.0 mg/l sulfate increased significantly at 10 mg/l Fe3+ or Al3+ up to 51.34 and 48.37%, respectively. In the presence of 10 mg/l Fe3+, the peak area of 1 mg/l and 10 mg/l phosphates decreased by 91.44 and 33.71%, respectively. At 10 mg/l Al3+ the peak areas of phosphate also increased slightly. The adverse effect of Fe3+ on the determination of phosphate could be overcome effectively by reducing the pH of the samples. The method to overcome the adverse effect was studied. PMID- 12108670 TI - Solving complex anion separation problems with a carbonate-hydrogencarbonate gradient. AB - This paper describes the application of a new suppressor to solve complex anion separation problems with carbonate-hydrogencarbonate gradients. In addition to suppressing the mobile phase and enhancing the analyte signal like any other ion chromatography suppressor, the new DS-Plus suppressor removes carbonic acid from the suppressor effluent when carbonate-hydrogencarbonate mobile phases are used. This greatly reduces the background signal, enabling carbonate-hydrogencarbonate gradients. Since carbonate-hydrogencarbonate mobile phase has stronger eluting power than hydroxide or tetraborate, only dilute concentrations are needed to separate most anions. By adjusting the carbonate-hydrogencarbonate ratio in the mobile phase, various anion mixtures including those containing organic and inorganic anions with -1 to -3 charges can be separated on one or two columns, eliminating the needs for columns with special selectivity. PMID- 12108671 TI - Rapid ion chromatography of L-ascorbic acid, nitrite, sulfite, oxalate, iodide and thiosulfate by isocratic elution utilizing a postcolumn reaction with cerium(IV) and fluorescence detection. AB - Rapid separation and determination of mixtures of L-ascorbic acid, nitrite, sulfite, oxalate, iodide and thiosulfate by conventional ion chromatography is often difficult due to incomplete separation of L-ascorbic acid and nitrite from the water peak when using eluents giving short elution times for iodide and thiosulfate. Separation of the six species within about 15 min has been achieved by isocratic elution using a resin-based ion-exchange column with a carbonate eluent containing a trace amount of 1,3,5-benzenetricarboxylic acid (BTA) and fluorescence measurement of cerium(III) formed via postcolumn reactions of the separated sample species with cerium(IV). Calibration plots of peak height versus concentration were linear up to 10.0 microM (1.76 ppm) for L-ascorbic acid, 8.0 microM (0.37 ppm) for nitrite, 8.0 microM (0.70 ppm) for oxalate, 80.0 microM (10.2 ppm) for iodide and 25.0 microM (2.80 ppm) for thiosulfate, whilst the sulfite calibration was linear up to 25.0 microM (2.00 ppm) when peak area was plotted against concentration. Detection limits (defined as S/N = 3) were 18 ppb for L-ascorbic acid, 4 ppb for nitrite, 16 ppb for sulfite, 7 ppb for oxalate, 72 ppb for iodide and 37 ppb for thiosulfate. The proposed method was applied successfully to the determination of L-ascorbic acid, nitrite, sulfite, oxalate, iodide or thiosulfate in water samples. PMID- 12108672 TI - Determination of trace level bromate and perchlorate in drinking water by ion chromatography with an evaporative preconcentration technique. AB - A simple sample preconcentration technique employing microwave-based evaporation for the determination of trace level bromate and perchlorate in drinking water with ion chromatography is presented. With a hydrophilic anion-exchange column and a sodium hydroxide eluent in linear gradient, bromate and perchlorate can be determined in one injection within 35 min. Prior to ion chromatographic analysis, the drinking water sample was treated with an OnGuard-Ag cartridge to remove the superfluous chloride and concentrated 20-fold using a PTFE beaker in a domestic microwave oven for 15 min. The recoveries of the anions ranged from 94.6% for NO2 to 105.2% for F-. The detection limits for bromate, perchlorate, iodate and chlorate were 0.1, 0.2, 0.1 and 0.2 microg/l, respectively. The developed method is applicable for the quantitation of bromate and perchlorate in drinking water samples. PMID- 12108673 TI - US Environmental Protection Agency Method 326.0, a new method for monitoring inorganic oxyhalides and optimization of the postcolumn derivatization for the selective determination of trace levels of bromate. AB - The development of US Environmental Protection Agency (EPA) Method 317.0 provided a more sensitive, acceptable alternative to EPA Method 300.1 to be proposed as one of the recommended compliance monitoring methods for Stage II of the Disinfectants/Disinfection By-Products (DBP) Rule. This work was initiated to evaluate other postcolumn reagents (PCRs) that might be utilized to provide an additional, alternative method in order to augment compliance monitoring flexibility for inorganic oxyhalide DBP anions. Modifications of the method reported by Salhi and von Gunten, which included adjustment and optimization of flow-rates, reaction temperature, and delivery of the PCR, improved the method performance. Method 326.0 incorporates an acidic solution of potassium iodide containing catalytic amounts of molybdenum(VI) as the PCR and provides acceptable precision and accuracy for all analytes and a postcolumn bromate detection limit in reagent water of 0.17 microg/l. PMID- 12108674 TI - Animal models of inflammatory bowel disease. PMID- 12108675 TI - Telomerase activity and expression of human telomerase catalytic subunit gene in esophageal tissues. AB - BACKGROUND: Telomerase, the ribonucleoprotein enzyme that synthesizes telomeric DNA, is thought to be necessary for cellular immortality and carcinogenesis. Telomerase activity is associated with the majority of malignant human cancers. The mRNA that encodes the telomerase catalytic subunit (human telomerase repeat transcriptase; hTERT) has recently been identified, and the expression of the hTERT gene is thought to regulate the activation of telomerase. However, the expression of hTERT mRNA in esophageal tissues has not been reported. We investigated hTERT gene expression in cancerous and noncancerous esophageal tissues, and determined the relationship between hTERT mRNA expression and telomerase activity. METHODS: Tissues from esophageal carcinomas in 14 patients, reflux esophagitis in 12 patients, esophageal acanthosis in 2 patients, esophageal papilloma in 1 patient, radiation esophagitis in 1 patient, and normal esophageal epithelium in 11 patients (including 3 specimens of normal epithelium from patients with esophageal carcinoma) were examined. All specimens were taken endoscopically. hTERT gene expression was investigated using reverse transcription-polymerase chain reaction (RT-PCR). Quantitative analysis of telomerase activity was analyzed by fluorescence telomeric repeat amplification protocol (F-TRAP) assay. RESULTS: Thirteen of the 14 (93%) esophageal carcinoma specimens expressed hTERT mRNA and revealed detectable telomerase activity. Noncancerous esophageal lesions had not only hTERT mRNA expression with a high frequency (14 of 16 cases; 88%) but also detectable telomerase activity (12 of 13 cases; 92%). Normal esophageal epithelium also highly expressed hTERT mRNA (10 of 11 cases; 91%) and revealed detectable telomerase activity (all 9 cases; 100%). In 32 of the 35 specimens analyzed for both hTERT mRNA and telomerase activity (91%), the expression of hTERT mRNA was consistent with detectable telomerase activity. CONCLUSIONS: The expression of hTERT mRNA was detected not only in cancerous but also in noncancerous esophageal tissues at a high frequency. This result was different from that reported for other gastrointestinal epithelium. Moreover, telomerase activity in esophageal carcinoma was significantly stronger than that in reflux esophagitis and normal epithelium. In addition, there was a strong relation ship between the detection of telomerase activity and the expression of hTERT mRNA in cancerous and noncancerous esophageal tissues. Thus, the qualitative analysis of hTERT mRNA expression may not be useful as a biomarker of carcinoma in esophageal tissues. Nevertheless, the quantitative analysis of telomerase activity may be somewhat useful. PMID- 12108676 TI - Immunological and molecular analysis of B lymphocytes in low-grade MALT lymphoma of the stomach. Are there any useful markers for predicting outcome after Helicobacter pylori eradication? AB - BACKGROUND: Although Helicobacter pylori eradication is effective in treating low grade gastric mucosa-associated lymphoid tissue (MALT) lymphoma, the condition in some patients deteriorates even after the eradication. Therefore, it is important to predict the disease outcome before starting H. pylori eradication. We investigated the usefulness of flow cytometry, quantifying CD19- and CD20 positive B lymphocytes in MALT lymphoma tissue, for predicting the disease outcome after H. pylori eradication. METHODS: Tissue specimens from 14 patients with H. pylori-positive low-grade gastric MALT lymphoma were examined by histology, Southern blotting, and flow cytometry before therapy. Serum levels of soluble interleukin (IL)-2 receptor were also measured. The relationship between the data and the prognosis after H. pylori eradication was analyzed. RESULTS: Remission occurred in 10 of the 14 patients. The condition in the 4 remaining patients deteriorated even after H. pylori eradication. The percentages of CD19- and CD20-positive cells in MALT lymphoma tissue from the patients in remission were both significantly lower than those in the tissue from patients not in remission. Indeed, 4 of the 5 patients in whom both CD19- and CD20-positive cells accounted for more than 50% of the total number of lymphocytes had gastrectomy, whereas all patients in whom both CD19- and CD20-positive cells accounted for less than 50% of the total number of lymphocytes achieved remission. Although immunoglobulin gene rearrangement was present in all patients operated on, there were also 6 patients whose MALT lymphoma was ameliorated in spite of the presence of gene rearrangement. The serum level of soluble IL-2 receptor was in the normal range in all patients tested. CONCLUSIONS: Analysis of mature B-cell markers in MALT lymphoma tissue is more useful than the examination of immunoglobulin gene rearrangement or serum levels of soluble IL-2 receptor in predicting the outcome of low-grade gastric MALT lymphoma after H. pylori eradication. PMID- 12108677 TI - Luminal polyamines upregulate transmural glucose transport in the rat small intestine. AB - BACKGROUND: Polyamines, which are contained in many foods, play an important role in the growth and differentiation of the enterocyte, but their role in glucose transport is unclear. Using isolated rat small intestine and a nonrecirculating perfusion system, we studied the effect of luminal polyamines on glucose uptake and on the concentration of sodium-glucose transporter 1 (SGLT1) and glucose transporter 5 (GLUT5) proteins. METHODS: In the control group, 300mg glucose solution was administered through the jejunum, and the glucose concentration in the portal vein was measured for 15 min. In treatment groups, various concentrations of polyamine (putrescine [Put] or spermine [Spm]) were administered simultaneously with the glucose. At the end of the perfusion period, the amount of SGLT1, GLUT5, and aminopeptidase N (APN) in the brush border membrane was subjected to Western blot analysis. RESULTS: Glucose concentration in the portal vein increased after the simultaneous administration of glucose and polyamines, and the area under the curve (AUC) after the 15-min perfusion was enhandced to 188%, 196%, 132%, and 192% by 0.5mM Spm, 4mM Spm, 1 mM Put, and 8 mM Put, respectively. The brush border membrane concentration of SGLT1 protein 15 min after polyamine administration was also enhanced in all treatment groups, and it correlated with the AUC. The concentration of GLUT5, on the other hand, was reduced by 4mM Spm, and the concentration of APN was not affected by polyamine administration. CONCLUSIONS: Luminal polyamines increase glucose absorption in the small intestine via the rapid enhancement of SGLT1 protein in the brush border membrane. PMID- 12108678 TI - Efficacy of lactulose plus 13C-acetate breath test in the diagnosis of gastrointestinal motility disorders. AB - BACKGROUND: We designed a new method of measuring gastric emptying and orocecal transit time (OCTT) at the same time to assess the influence of gastric emptying upon OCTT. METHODS: Twenty-five dyspeptic patients (6 men, 19 women) with a mean age of 64.8 years (range, 25-80 years) were studied. The patients received a liquid test meal, containing 100 mg of 13C-acetate and 12g of lactulose, while they were in the sitting position after an overnight fast. Breath samples were collected at 10-min intervals of 120 min and both 13CO2 and hydrogen (H2) levels were measured. Subsequently, H2 concentrations were measured at 30-min intervals, for a total of 240 min. RESULTS: The results of gastric emptying were expressed as the time of peak 13CO2 excretion. OCTT was defined as the period between the ingestion of lactulose and a H2 peak rise of 5 ppm above the baseline value. The onset of H2 enrichment in the breath began at 90-110 min, whereas 13CO2 levels increased from the beginning, with peak enrichment values being reached after 60 80 min. OCTT was related to 13CO2 peak time. In 5 of the 25 patients, H2 breath enrichment in the 10-min sample was more than 5 ppm over the baseline value. All these 5 patients had double or triple peaks in serial breath H2 concentrations. CONCLUSIONS: The combination of the lactulose hydrogen breath test (LHBT) with the 13C-acetate breath test, which requires only breath samples, provides us with much information on the gastrointestinal tract; gastric emptying, OCTT, bacterial overgrowth in the small intestine, colonic fermentation, and oropharyngeal flora. The 13C-acetate breath test can be useful as an adjuvant test when LHBT is performed for measuring OCTT. PMID- 12108679 TI - Molecular cloning, and characterization and expression of dihydrolipoamide acetyltransferase component of murine pyruvate dehydrogenase complex in bile duct cancer cells. AB - BACKGROUND: The association between the dihydrolipoamide acetyltransferase component (E2) of pyruvate dehydrogenase complex (PDC) and primary biliary cirrhosis (PBC) is clinically established. However, the detailed pathological function of the PDC-E2 gene is as yet unclear. In order to study the gene function in knockout and transgenic mouse models, we cloned and characterized the mouse PDC-E2 (mPDC-E2) gene. Because the expression level of PDC-E2 was elevated in PBC bile duct cells, we tried to construct a bile duct carcinoma cell line that overexpressed PDC-E2 as a PBC cell model. METHODS: The mPDC-E2 cDNA was obtained by the 3'Race method. We overexpressed this gene in KMBC cells, using a retrovirus vector. The transcript and translated protein of mPDC-E2 were detected by Northern blot and Western blot, respectively. RESULTS: The deduced amino-acid sequence from the cloned cDNA indicated that the fully mature protein consisted of 557 amino-acid residues, with a calculated molecular mass of 59kD. This mature protein was highly consistent with those of previously reported rat and human PDC E2, which possessed three structurally identifiable regions: the lipoyl-bearing domain, the E3-binding site, and the catalytic domain. Mouse fibroblast NIH3T3 cells expressed one species of mPDC-E2 mRNA, 3.5kb in length. We also successfully constructed a stable KMBC cell line overexpressing the PDC-E2. CONCLUSIONS: This is the first report of the mPDC-E2 sequence and is valuable for further investigation of PDC-E2 gene function in transgenic or knockout mouse models. The PDC-E2 overexpressing KMBC cell line can be used to study alterations in signal transduction or gene expression profiles in PBC bile duct. PMID- 12108680 TI - The relationship between gallbladder disease and smoking and drinking habits in middle-aged Japanese. AB - BACKGROUND: Few studies have investigated the association between smoking and ultrasonographically diagnosed gallbladder (GB) disease, and their results were uncertain. This study was conducted to examine the association between smoking and drinking and GB diseases. METHODS: A total of 9,947 subjects (age, 30-69 years; 4,953 men and 4,994 women) voluntarily received a paid medical check-up at our center in Yamanashi Prefecture in Japan. All of the subjects underwent abdominal ultrasonographic (US) examination, a demographic check, and a biochemical test, and answered a self-administered questionnaire asking about smoking habits and alcohol consumption. Of the 9,947 subjects, 483 had gallstones, 819 had gallbladder polyps, and 169 were in a state of postcholecystectomy. We compared the findings in this group with the findings in 8,417 people (4,144 males and 4,273 females) with normal gallbladder. RESULTS: Multiple regression analysis among males showed that cigarette smoking was inversely related to GB polyps (odds ratio, [OR], 0.76; 95% confidence internal [CI], 0.59-0.98 and OR, 0.74; 95% CI, 0.56-0.98, respectively, for current and ex smokers). Ex-smokers a showed positive association with the postcholecystectomy state (OR, 2.56; 95% CI, 1.18-5.52). Light drinkers showed an inverse relation to GB stones (OR, 0.69; 95% CI, 0.49-0.99), and heavy drinkers showed an inverse relation to GB polyps (OR, 0.68; 95% CI, 0.51-0.90). Current drinkers showed an inverse relation to the postcholecystectomy state (OR, 0.48; 95% CI, 0.28-0.83). CONCLUSIONS: Cigarette smoking was inversely related to gallbladder polyps in males and was positively related to the postcholecystectomy state. Drinking was inversely related to gallstones, GB polyps, and the postcholecystectomy state in males. PMID- 12108681 TI - Cap polyposis cured by Helicobacter pylori eradication therapy. AB - The pathogenesis of cap polyposis remains unknown. Here, we report a patient with cap polyposis that developed simultaneously in the colon and stomach, and which regressed after Helicobacter pylori eradication. A 63-year-old man was diagnosed as having cap polyposis with mucoid stool, diarrhea, and bleeding on defecation. Following 5 weeks of total parenteral nutrition, his symptoms and hypoproteinemia improved and he was discharged, although follow-up colonoscopic findings revealed no improvement. Subsequent gastroscopy revealed diffusely erosive polyps with cap like "fur" from the angle to the antrum of the stomach, similar to the lesions observed in the colon. Because H. pylori infection was demonstrated in the stomach, eradication therapy was administered. After this treatment, his symptoms immediately disappeared, and the polypoid lesions in the colon and stomach had completely disappeared 8 months later. Because there have been no previous reports of a relationship between H. pylori and cap polyposis, this case is of great interest. PMID- 12108682 TI - Serrated adenoma developing into advanced colon cancer in 2 years. AB - We report the case of a 76-year-old woman with a flat elevated lesion (laterally spreading tumor; LST) in the transverse colon that had been identified 2 years before the current presentation. Pathologically, the lesion had been diagnosed as a serrated adenoma with severe dysplasia. The patient had refused endoscopic resection because she was afraid of the risk of colonic perforation; she did not come to follow-up until 2 years after the diagnosis, when she had advanced colon cancer in the transverse colon where the LST had been detected. Histopathological examination confirmed a poorly differentiated adenocarcinoma with no adenomatous component such as had been found in the LST lesion. PMID- 12108683 TI - Small-intestinal sarcomatoid carcinoma with superior vena cava syndrome. AB - A 56-year-old man was hospitalized because of swelling of the right upper extremity and anemia. A diagnosis of superior vena cava (SVC) syndrome caused by lymphogenous metastasis was made after chest computed tomography (CT) scan and biopsy of cervical lymph nodes were carried out. Standard examinations, such as abdominal CT scan and endoscopies of the upper and lower gastrointestinal tract, failed to find the primary lesion. However, selective angiography of the superior mesenteric artery (SMA) showed a clear stain of bleeding vessels in the small intestine. Laparotomy was performed, and immunohistochemical findings revealed sarcomatoid carcinoma in the small intestine (a rarely seen neoplasm). This aggressive carcinoma, which showed negative reactivity with CD34, CD117 (c-kit), and S-100 was clearly distinguished from other mesenchymal tumors, such as malignant gastrointestinal stromal tumor (GIST) and malignant fibrous histiocytoma (MFH). PMID- 12108684 TI - Unusual presentation of Bouveret's syndrome. AB - Isolated gastric outlet and ileal obstruction are rare complications of gallstones. We report what we believe to be the first case of a gallstone causing gastric outlet obstruction while eroding into the duodenum, yet once in the duodenum, passing into the small bowel, causing ileal obstruction. The endoscopic appearance of the eroding gallstone was so unusual that it was initially mistaken for a fibrotic duodenal ulcer. PMID- 12108685 TI - A vanishing pseudocyst in the remnant pancreas after pylorus-preserving pancreatoduodenectomy. AB - We report a 74-year-old Japanese woman with a spontaneously vanishing pseudocyst in the remnant pancreas after pylorus-preserving pancreatoduodenectomy for intraductal papillary-mucinous adenoma of the pancreas. A cystic lesion appeared in the remnant pancreas 6 months after the operation and had disappeared 3 months later. When a cystic lesion is encountered in the remnant pancreas after pancreatectomy for mucin-hypersecreting tumor of the pancreas, pseudocyst, as well as recurrence, should be considered in the differential diagnosis. Additional resection would likely cause considerable morbidity, with loss of endocrine and exocrine functions. PMID- 12108686 TI - Possible relationship between deficiency of interstitial cells of Cajal and colonic inertia in a patient with colon malfixation. PMID- 12108687 TI - A patient with multiple endocrine neoplasia type 1 with a novel MEN1 gene mutation (237delC). PMID- 12108688 TI - Telomerase activity and human telomerase RNA component in gastrointestinal diseases. PMID- 12108689 TI - Regulation of monosaccharide transporter proteins in the small intestine. PMID- 12108690 TI - Cloning of murine PDC-E2 gene and generation of PDC-E2 overexpressing cell line: a key step to clarify the pathogenesis of PBC? PMID- 12108691 TI - Determination of pentachlorophenol (PCP) in waste wood--method comparison by a collaborative trial. AB - Two independently developed and validated procedures for the determination of pentachlorophenol (PCP) in waste wood were compared by means of a collaborative trial. Both methods foresee quantification of PCP by gas chromatography (GC-ECD) after acetylation and differ with regard to the use of methanol or toluene/sulphuric acid, respectively, as solvent in the sonication extraction step. Test samples with established analyte homogeneity were prepared from a ground "real life" starting material. A total of 23 participating laboratories with experience in wood preservative analysis were instructed to apply both methods to three levels of content in the range of 0.5-20 mg PCP/kg. In case of the toluene/sulphuric acid extraction, lower recoveries and higher interlaboratory dispersion of results at the higher PCP contents were observed. Seen against the background of the Horwitz equation a reproducibility standard deviation of approximately 19% for the methanol extraction at the 4.5 mg/kg level meets the requirement for a sound analytical method. Thus, the sonication extraction procedure with methanol has been annexed as a reference method to the German waste wood regulation. PMID- 12108692 TI - Multiple regression models: a methodology for evaluating trihalomethane concentrations in drinking water from raw water characteristics. AB - The presence of trihalomethanes (THMs) in drinking water has attracted the attention of both researchers and professionals, because of the harmful effects of these substances on human health. A multiple regression model was developed to estimate THM concentrations in finished drinking water, using data from the Menidi Treatment Plant of Athens. A number of routinely measured characteristics- including chlorine dose, chlorophyll a, temperature, pH and bromide--of raw water, were used to generate a reliable methodology for predicting both total THM and individual species concentrations. Seasonality effects were also considered during the analysis. In general, these models were found to give acceptable fits, estimating accurately lows and highs over the annual cycle. PMID- 12108693 TI - Photocatalytic oxidation of methyl orange in a three-phase fluidized bed reactor. AB - The photocatalytic oxidation of methyl orange under weak illumination conditions was carried out in two different types of fluidized bed reactors. TiO2 powder was employed as the photocatalysts and a 15 W low pressure mercury lamp was used as the light source and the reactor volume was 2.5 l. The parametric study of photocatalytic oxidation of methyl orange in two different types of fluidized beds was investigated; effect of catalyst loadings, pH, air flow rate, initial concentration and oxygen concentration on the oxidation reaction rate. The experimental results were analyzed in conjunction with the characteristics of fluidized bed and the reactor geometry effect on the reaction rate was analyzed. PMID- 12108694 TI - Herbicides and nitrogen in precipitation: progression from west to east and contribution to the Marne river (France). AB - Herbicides and nitrogen fallout were studied in France at six sites from west to east. Atrazine, a herbicide widely used in France and forbidden in some European countries was found. Its fallout was quantified to 400 kg on the river Marne catchment (13 500 km2), an agricultural region located east of Paris. The average fallout contribution to the Marne river contamination was estimated at 20 ng/l from March to August. Independently of the wind direction, only a small part of this atmospheric contamination can reach adjacent countries. PMID- 12108695 TI - Biodegradability of aged pyrene and phenanthrene in a natural soil. AB - A study was conducted to evaluate the biodegradability of pyrene (PYR) and phenanthrene (PHE) aged in a natural soil. Both the single and binary systems were either biostimulated via a nutrient amendment or bioaugmented via an inoculation of the enriched bacteria and nutrients. Aging resulted in higher concentration of both compounds and smaller bacterial activity in the solution phase. Surprisingly, the total biodegraded extent was greater in the aged soil system than in the freshly spiked system. As anticipated, biostimulation was not appropriate to attain an effective biodegradation in this study, and bioaugmentation achieved a substantial increase the total biodegradation extent. The above findings were attributed to indigenous Pseudomonas aeruginosa entering a stationary-phase during the 200-day aging and producing rhamnolipid biosurfactants. In addition, a different sampling technique (i.e., after vigorous hand-shaking) revealed a 15 times higher microbial population than the normal sampling from the stagnant solution. Therefore, PAH bioavailability in the aged soils can be underestimated when the microbial activity is determined only from the stagnant solution. Furthermore, cometabolism enhanced PYR degradation when PHE was present as a primary substrate. PMID- 12108696 TI - Effect of concentration, moisture and soil type on the dissipation of flufenacet from soil. AB - Effect of concentration, moisture and soil type on dissipation of flufenacet from soil has been studied under laboratory condition. The treated soil samples (1 and 10 microg/g levels) were incubated at 25+/-1 degrees C. The effect of moisture was studied by maintaining the treated soil samples (10 microg/g level) at field capacity and submerged condition. In general, flufenacet persisted for 60-90 days at lower and beyond 90 days at high rate. The dissipation of flufenacet from soil followed first order kinetics with half-life (DT50) values ranging from 10 to 31 days. The dissipation of flufenacet was faster at low rate than high rate of application. The slow dissipation at high rate could be attributed to inhibition of microbial activity at high rate. There was little overall difference in rate of dissipation in Ranchi and Nagpur soil maintained at field capacity and submerged condition moisture regimes. In Delhi soil net dissipation was faster under field capacity moisture than submerged condition. Soil types greatly influenced the dissipation of flufenacet. Dissipation was fastest in Delhi soil (DT50 10.1-22.3 days) followed by Ranchi soil (DT50 10.5-24.1 days) and least in Nagpur soil (DT50 29.2-31.0 days). The difference in dissipation could be attributed to the magnitude of adsorption and desorption of flufenacet in these soils. PMID- 12108697 TI - Mercury removal from incineration flue gas by organic and inorganic adsorbents. AB - Experiments were performed to investigate various adsorbents for their mercury removal capabilities from incineration flue gases. Four different materials were tested; Zeolite, Bentonite, activated carbon (AC), and wood char. Real incineration off-gas and in-lab simulated combustion flue gases (N2 + Hg) were used. Three cylindrical-shaped sorbent columns with 5 cm in diameter and 20 cm in length were used. The gas flow rate was fixed at 660 l/h at all times. Concentrations of NO, CO, O2, CO2, SO2, H2O, HCl, and mercury were continuously monitored. Mercury removal efficiencies of natural Zeolite and Bentonite were found to be much lower than those of the referenced AC. Amount of Hg removed were 9.2 and 7.4 microg/g of Zeolite and Bentonite, respectively. Removal efficiencies of each layer consisted of inorganic adsorbents were no higher than 7%. No significant improvement was observed with sulfur impregnation onto the inorganic adsorbents. Organic adsorbents (wood char and AC) showed much higher mercury removal efficiencies than those of inorganic ones (Zeolite and Bentonite). Mercury removal efficiency of wood char reached over 95% in the first layer, showing almost same effectiveness as AC which currently may be the most effective adsorbents for mercury. Amount of mercury captured by wood char was approximately 0.6 mg/g of wood char, close to the amount captured by AC tested in this study. Hence, wood char, made from the waste woods through a gasification process, should be considered as a possible alternative to relatively expensive AC. PMID- 12108698 TI - Kinetic modeling of fenton oxidation of phenol and monochlorophenols. AB - A kinetic model, consisting of 28 reactions, was proposed to understand the key mechanism of the Fenton oxidation of phenol and o-, m-, and p-chlorophenols. Particular attention is paid to the interactions of the organic intermediates with the Fe species. The proposed model reasonably predicts the decomposition kinetics and by-product formation for the different phenols at widely varying levels of Fe2+, H2O2, and the phenols. For the phenols and intermediates, change in concentrations with time was predicted within 20-30% deviation from the measured data. The single model predicts the overall kinetics of Fenton oxidation of all the tested phenols by adjusting the rate constant of hydroxyl radical for each phenol. Sensitivity analysis indicates that the key reactions are those that directly govern the levels of OH radical and Fe2+. Both the model prediction and the experimental results show that the decomposition rate could be complicated particularly by the availability of Fe2+. Understanding the interactions of the organic intermediates with Fe2+ is thus of critical importance to improve the decomposition performance. PMID- 12108699 TI - Kinetics of chemical decolorization of the azo dye C.I. Reactive Orange 96 by sulfide. AB - The mechanism of decolorization of azo dyes based on the extracellular chemical reduction with sulfide (H2S, HS-, S2-) was postulated for sulfate reducing environments. To design technical decolorization processes of textile wastewater treatment with sulfide produced by sulfate reducing bacteria (SRB), kinetics is of great significance. Batch experiments were made in order to investigate the kinetics of abiotic decolorization of the reactive mono-azo dye C.I. Reactive Orange 96 (RO 96) with sulfide, with varying pH. The decolorization of RO 96 by sulfide under the exclusion of O2 corresponded to first-order kinetics with respect to both dye and sulfide concentration. The decolorization of RO 96 with sulfide at neutral pH (7.1) was advantageous compared with that at pH for 4.1, 6.3, and 6.5. This is attributed to an increase in the fraction of HS- of total sulfide species at neutral pH. The rate constants k for the decolorization at 37 degrees C were obtained as 0.01 for pH = 4.1, 0.06 for pH = 6.3, 0.08 for pH = 6.5, and 0.09 for pH = 7.1 in mM(-1) min(-1). The high rate constants for sulfide at pH 6.5-7.1 support that the decolorization through SRB (i.e. by bio-sulfide) can be effective in anaerobic bacterial systems with sulfate. PMID- 12108700 TI - Determination of the temperature dependence of water solubilities of polycyclic aromatic hydrocarbons by a generator column-on-line solid-phase extraction-liquid chromatographic method. AB - An improved dynamic coupled column liquid chromatographic (DCCLC) technique for determining water solubility data of hydrophobic compounds is presented. The technique is based on pumping water through a thermostated generator column in order to generate emulsion-free, saturated aqueous solutions of the compound under study. Through a switching valve system the solute in the aqueous solution is extracted and concentrated by an on-line solid-phase extraction process and subsequently eluted and analyzed by high performance liquid chromatography (fluorescence detection coupled to photodiode array detection). The improvements carried out to the original DCCLC technique have given rise to savings in time for the experimental work and increased sensitivity during the detection and quantification stage. Applicability of the method for studying highly hydrophobic substances is demonstrated by determining water solubility of anthracene and pyrene in the temperature range of 8.9-49.9 and 8.5-32.2 degrees C, respectively. The measured water solubilities are in good agreement with the best available literature data. The method has also been applied to the determination of water solubility of m-terphenyl, 9, 10-dihydrophenanthrene and guaiazulene, in the temperature range of 4.8-49.9, 4.8-25.0, and 4.5-29.9 degrees C, respectively. The uncertainty in the Sw values determined in this work ranged from 0.7% to 4.6%. The experimental water solubility data, as a function of temperature, are fitted to the equation In Sw = A + B/T; where Sw and T are given in mole fraction and Kelvin, respectively. PMID- 12108701 TI - Screening of pesticides for environmental partitioning tendency. AB - The partitioning tendency of chemicals, in this study pesticides in particular, into different environmental compartments depends mainly on the concurrent relevance of the physico-chemical properties of the chemical itself. To rank the pesticides according to their distribution tendencies in the different environmental compartments we propose a multivariate approach: the combination, by principal component analysis, of those physico-chemical properties like organic carbon partition coefficient (Koc), n-octanol/water partition coefficient (Kow), water solubility (Sw), vapour pressure and Henry's law constant (H) that are more relevant to the determination of environmental partitioning. The resultant macrovariables, the PC1 and PC2 scores here named leaching index (LIN) and volatality index (VIN), are proposed as preliminary environmental partitioning indexes in different media. These two indexes are modeled by theoretical molecular descriptors with satisfactory predictive power. Such an approach allows a rapid pre-determination and screening of the environmental distribution of pesticides starting only from the molecular structure of the pesticide, without any a priori knowledge of the physico-chemical properties. PMID- 12108702 TI - Transformation of [14C] trichloroethylene by poplar suspension cells. AB - Trichloroethylene (TCE) is one of the most prevalent environmental contaminants, and it poses an expensive remediation problem. Phytoremediation has been investigated as a potential tool for the removal of TCE from ground water and soil, and has shown promise in preliminary trials. However, the fate of TCE in plants is largely unknown. Radiolabel studies showed that once taken up and transformed, most of the TCE is incorporated into plant tissue as a non-volatile, un-extractable residue. We describe here an assay for TCE transformation by poplar suspension cells. Using this assay, it was shown that two different activities contribute to the fixation of TCE by poplar cells, one associated with cell walls and insoluble residues, the other associated with a high molecular weight, heat labile fraction of the cell extract. It appears that plant enzymes catalyze some of the transformations. PMID- 12108703 TI - Comparison of hydrolysis/coagulation behavior of polymeric and monomeric iron coagulants in humic acid solution. AB - Fluorescence quenching is commonly used to study the extent of metal binding to humic acid or fulvic acid. By studying this phenomenon, the hydrolysis and precipitation behaviors of polymeric and monomeric iron coagulants in the coagulation of humic acid were evaluated. Combined measurements of fluorescence intensity and dissolved organic carbon were performed to distinguish the hydrolysis and organic matter binding of polymeric and monomeric iron salts in coagulating high molecular weight organic compounds. Experimental results showed that ferric chloride had lower fluorescence quenching than polyferric sulfate (PFS), indicating more rapid hydrolysis of the monomeric ferric ion in the coagulation and dilution process. The less fluorescence quenching was caused by the lower residual concentration of Fe-humic acid complexes in the filtrate. There was no clear difference in adsorption ability and fluorescence quenching, which indicated that the physical and chemical properties of Fe(OH)3 flocs formed from PFS and FeCl3 coagulation were similar. From the pH study, we found that sludge formed from PFS coagulation was more stable than that from FeCl3 coagulation. PMID- 12108704 TI - Dechlorination of pentachloroethane by commercial Fe and ferruginous smectite. AB - Short-term experiments were conducted to investigate the effect of a commercial Fe and an iron-bearing clay mineral, ferruginuous smectite (SWa-1), on the degradation of pentachloroethane (PCA). After 3 h of contact time, SWa-1 catalyzed PCA dehydrochlorination to tetrachloroethene (PCE, 65% conversion), whereas commercial Fe promoted PCA stepwise dechlorination via dehydrochlorination (approximately 40% conversion) and subsequent PCE hydrogenolysis to trichloroethene (TCE). The addition of unaltered SWa-1 to commercial Fe led to a complete inhibition on TCE production, whereas the addition of reduced SWa-1 barely resulted in a 30% decrease. PMID- 12108705 TI - Effect of dissolved metal ions on PCE decomposition by gamma-rays. AB - This study investigates the effect of initial tetrachloroethylene (PCE) concentration, irradiation dose and dissolved metal ions such as Cr3+, Mn2+, Fe3+, Co2+, Ni2+, Cu2+ and Zn2+ on removal of PCE by gamma irradiation. The amount of removed PCE decreased with increase in initial PCE concentration and increased with increase in irradiation dose. PCE removal reached a maximum in the presence of Fe3+, while Cu2+ strongly hindered PCE decomposition. Except for Cu2+, the amount of removed PCE in the presence of metal ions was linearly dependent on the standard reduction potential of the metal ions. The extraordinary inhibition of Cu2+ in PCE removal was caused by the action of Cu2+ as a strong *OH scavenger, that was directly confirmed by electron paramagnetic resonance spectroscopy. PMID- 12108706 TI - Adsorption of phenolic compounds from aqueous solutions by a water-compatible hypercrosslinked polymeric adsorbent. AB - A water-compatible hypercrosslinked polymeric adsorbent (NJ-8) for adsorbing and removing phenolic compounds from their aqueous solutions was prepared. This product can be used directly without a wetting process. Its adsorption property toward four phenolic compounds, phenol, p-cresol, p-chlorophenol, and p nitrophenol was tested using the commercial Amberlite XAD-4 as a reference. The capacities of equilibrium adsorption for all four phenolic compounds on the NJ-8 from their aqueous solutions are around two times as high as that of Amberlite XAD4 within the temperature range 283-323 K, which may contribute to their micropore structure and the partial polarity on the network. Freundlich isotherm equations, as well as relative adsorption capacities and isosteric adsorption enthalpies for the four phenolic compounds, indicate that the adsorption of phenolic compounds on the NJ-8 resin is a physical adsorption process. Mini column adsorption studies for phenol on Amberlite XAD4 and NJ-8 resins show that the breakthrough adsorption capacities are 0.54 and 0.99 mmol/ml, and the total capacities are 0.62 and 1.37 mmol/ml, while no extra acetone was needed to remove the adsorbed phenol from NJ-8 as from Amberlite XAD4. PMID- 12108707 TI - Riboflavin-photosensitized degradation of atrazine in a freshwater environment. AB - The effect of the photosensitizer riboflavin (0, 10, 50, 100 microM) on the fate of atrazine (10 mg/l) in a freshwater environment was studied. It was found that at 100 microM riboflavin significantly enhanced the degradation of atrazine and more than 80% of atrazine in a natural water environment was depleted in 72 h. The relative contribution of microbial assemblages and the freshwater matrix to the degradation of atrazine and the degradation kinetics of atrazine were compared under different experimental conditions. The products and pathways of atrazine transformation were studied with GC-MS and HPLC with a photodiode array detector. The results show that dealkylation and alkyl chain oxidation are involved in the degradation of atrazine. PMID- 12108708 TI - Enhanced photo-degradation of contaminants in petroleum refinery wastewater. AB - In order to rise efficiency of the wastewater treatment in a refinery plant, several oxidation experiments were done, testing their applicability as an additional pretreatment method. The influence of treatment with low concentrations of H2O2 combined with stirring and UV light on degradation of organic compounds present in the refinery wastewater was studied. Oxidation of the total petroleum hydrocarbons occurs at relatively low concentrations of H2O2, additional UV irradiation slightly accelerates the process due to the increased formation of hydroxyl radicals. 1,2-dichloroethane and t-butyl methyl ether degrade in the similar manner and except for the lowest H2O2 concentration used (1.17 mM), the reduction after 24 h is total. The degradation rate for dichloromethane is the lowest one, depending both on hydrogen peroxide concentration and the presence of UV. Its maximum reduction of 83% was obtained using the highest applied peroxide concentration of 11.76 mM. PMID- 12108709 TI - The importance of light intensity in algal tests with coloured substances. AB - Current algal test protocols do not distinguish between chemical toxicity and growth inhibition due to physical shading when coloured substances like dyes are evaluated. To eliminate the interfering physical effect, we investigated the importance of light intensity and culture volume in the algal growth inhibition tests with Desmodesmus subspicatus and three selected dyestuffs. A photon flux density above light saturation is needed to prevent inhibition of algal growth solely due to the light absorption by dyes. Furthermore, a smaller culture volume and moderate shaking of the test flasks is advantageous in that these conditions decrease the influence of the light absorption. At 120 microE s(-1) m(-2), the shading effect of three tested dyes could be eliminated, up to a concentration of 100 mg l(-1). In a mixture with a blue dyestuff, the model toxicant potassium dichromate showed nearly the same EC50 as if applied alone, indicating that the impact of the dye was negligible. Also a small culture volume decreases the inhibition of the average growth rate of Desmodesmus. Thus, by increasing the light intensity and optionally also reducing the culture volume, this method allows a more direct measure of a potential inherent chemical toxicity of coloured substances to algae. Additionally, high light intensities and lower culture volumes result in lower coefficients of variation, which improves the sensitivity of the test for the statistical calculation of toxicity data. PMID- 12108710 TI - The behavior of polar aromatic sulfonates during drinking water production: a case study on sulfophenyl carboxylates in two European waterworks. AB - First investigations are reported on the efficiency of individual purification steps at two waterworks to eliminate sulfophenyl carboxylates (SPC) originating from biodegradation of linear alkylbenzene sulfonate surfactant. The average SPC concentrations in the waters taken from the Llobregat river, Spain and the Rhine river, Germany amounted to 5.0 and 1.8 microg L(-1), respectively. In the Spanish waterworks, neither prechlorination nor flocculation followed by rapid sand filtration had an impact. After ozonation, granular activated carbon filtration, and final chlorination the SPC level was about 2 microg L(-1) in such processed drinking water. In the German waterworks already the rapid sand filtration diminished the SPC concentration by >85%. Subsequent subsoil passage resulted only in a slight elimination, but once again a slow sand filtration prior to the closing chlorination substantially removed the polar micro-pollutants down to a level of <0.05 microg L(-1) SPC. PMID- 12108711 TI - The direct and indirect photolysis of 4,4'-dichlorobiphenyl in various surfactant/solvent-aided systems. AB - Surfactant and organic solvents have individually been shown useful in assisting the solubilization of hydrophobic organics out of contaminated soil and promoting UV-induced photodecay at a succeeding treatment process. A newly proposed process is to use the mixtures of surfactant and solvents together in achieving better performance. This study was to explore some basic variables and conditions that may be useful in optimizing the performance of surfactant/solvent-aided UV systems through the selection of surfactant micelles, the adjustment of the reaction pH levels, and the addition of photosensitizer and hydrogen sources. A PCB monomer 4,4'-dichlorobiphenyl (DCB) was used as the target compound for this study; about 1.3 to 2.3 times greater DCB photodecay rate enhancements were observed by using surfactant/acetone or surfactant/triethylamine (TEA) combinations at their optimal conditions. Overdose of these additives (acetone and TEA) will result in retardation of the reactions. However, the use of surfactant with a multisolvents system (surfactant/acetone/TEA) shows an add-on effect in amplifying the overall decay rate for more than 12%, suggesting that a more complicated mechanism is involved than the simple parallel-reaction assumption. PMID- 12108712 TI - Removal of suspended solids by coagulation and foam separation using surface active protein. AB - By using several kinds of surface-active proteins as a chemical agent that combined collector with frother, removal of suspended substances by coagulation and foam separation with dispersed air was examined. Milk casein showed the greatest capability of suspension removal, and coagulating flocs formed by clay particles and iron hydroxide were almost perfectly recovered in foam generated from the liquid, even in the case of freshwater and seawater suspension at neutral pH. In contrast, the removal efficiency was extremely low using sodium dodecyl sulfate (SDS). Casein had a much greater capability for removing solids than SDS as a result of the high adsorptive activity of casein on the floc. For municipal wastewater treatment, the removal efficiency of turbidity and suspended solids was over 98% with the condition of iron coagulant (FeCl3) 20 mg-Fe/L and casein 3 mg/L and pH 5-6. Moreover, this method proved to be an effective treatment for polluted saline water (salinity 1.5%), and the suspended solids were almost perfectly recovered in foam. Here, we show a new method for quickly removing (within 7 min) suspended solids from polluted wastewater utilizing casein and bubbles. PMID- 12108713 TI - Combined chemical and biological degradation of tannery wastewater by a periodic submerged filter (SBBR). AB - The paper reports on the results of an investigation aimed to evaluate the performances of an innovative tannery wastewater process based on the combining biological degradation, carried out in a sequencing batch biofilm reactor, with chemical oxidation, performed by ozone. The combined treatment was carried out at the laboratory scale on real primary effluent coming from a centralised plant treating the wastewater from a large tanning district in Northern Italy. SBBR performances with and without ozonation were compared resulting to be very satisfactory only in the latter instance where recorded COD, NH4-N and TSS average removals were 97%, 98% and 99.9%, respectively. Such efficiencies correspond to specific concentrations in treated effluent well below the limit values fixed by the in-force Italian regulations. Furthermore, it was proved that the combined process is characterised by a very low sludge production. In fact, the measured specific sludge production (0.03 kg TSS/kg COD(removed)) resulted unexpectedly much more lower than the value reported for conventional biological systems (i.e., 0.3-0.5 kg TSS/kg COD(removed)). PMID- 12108714 TI - Photoelectrocatalytic degradation of humic acid in aqueous solution using a Ti/TiO2 mesh photoelectrode. AB - Humic acid (HA) is one of natural organics existing in water supply as a precursor of trihalomethanes formation in chlorination. The photo-degradation of HA in aqueous solution by photoelectrocatalytic (PEC) oxidation using a Ti/TiO2 mesh electrode was investigated in terms of UV absorbance at 254 nm, colour and TOC concentration. The key factors affecting the PEC oxidation efficiency were studied, including the concentration of electrolyte, electrical bias applied. pH value of HA solution, the intensity of incident light and the area of Ti/TiO2 mesh photoelectrodes. The first-order kinetic model was applied to describe the PEC oxidation, in which the kinetic constant k was verified by the experimental data as a function of the concentration of electrolyte, light intensity, the area of Ti/TiO2 mesh electrode and the voltage of electrical bias applied. It was found that there was an optimal bias voltage of 1.63 V and low pH value was favourable for TOC removal in HA solution. Our investigation showed that PEC oxidation was a convenient way to mineralise the organic matters with high efficiency. PMID- 12108715 TI - Treatment of domestic sewage in a two-step anaerobic filter/anaerobic hybrid system at low temperature. AB - The treatment of domestic sewage at low temperature of 13 degrees C was investigated in a two-step system consisting of an anaerobic filter (AF) +an anaerobic hybrid (AH) reactor operated at different hydraulic retention times (HRTs). The AF reactor was efficient in the removal of suspended COD, viz. 81%, 58% and 57% at an HRT of, respectively, 4, 2 and 3 h. For optimisation of the removal of suspended COD and dissolved COD, an HRT of 4 + 4 h is required for the AF + AH system. For additional optimisation of colloidal COD removal, the AH reactor needs an HRT of 8 h. The AF + AH system operated at an HRT of 4 + 8 h at 13 degrees C provided a high removal efficiency for all COD fractions. The achieved total COD removal was as high as 71% which is similar to values found in tropical areas. Moreover, 60% of the removed COD was converted to methane. PMID- 12108716 TI - Copper and zinc adsorption onto poorly humified Sphagnum and Carex peat. AB - Peat generally has a high adsorption capacity and has been suggested as an adsorbent for metals in polluted waters. However, the adsorption potential of peat can be expected to be strongly dependent on the chemical properties of the water. In this study, the effect of pH, ionic strength (CaCl2 and NaCl concentrations), and metal concentration on Cu and Zn adsorption onto poorly humified Sphagnum and Carex peat was investigated in batch experiments using a fractional factorial experimental design. The pH value was varied between 4 and 8, the CaCl2 and NaCl concentrations between 1.2-6.2 and 0.4-43 mM, respectively, and the Cu and Zn concentrations between 0.05 and 0.5 mM. The amount of Zn adsorbed increased more with increasing pH than the amount of Cu adsorbed. The effect of NaCl/CaCl2 concentration was minor. It was found that Zn adsorption in particular, but also Cu adsorption, increased more with pH onto Carex peat than onto Sphagnum peat. In the pH interval 4-8, the removal of Zn from the solution increased from 0% to 80% using Carex peat and from 10% to 65% using Sphagnum peat as the adsorbent. The Carex peat sample decreased the Cu concentration by 80% at pH 4 and by 95% at pH 8 and for Sphagnum peat a decrease of 85% was maintained in the pH range investigated. The differences between Sphagnum and Carex peat were attributed to the habitat conditions at the time of peat formation. Carex peat has higher ash, nitrogen and sulphur concentrations, while Sphagnum peat contains a higher amount of uronic acid. In treating polluted waters with peat, a higher degree of metal removal can be expected at high pH values than at low. The removal of Cu, which has a high affinity to the peat surface, was less dependent on pH than Zn removal. Poorly humified Carex peat should be chosen in treating wastewaters high in pH. PMID- 12108717 TI - Electrolytic oxygen generation for subsurface delivery: effects of precipitation at the cathode and an assessment of side reactions. AB - This research investigated the oxygen-generating characteristics and side reactions of an electrolytic cell assembly that could be used to remediate sites with contaminants that are amenable to aerobic biodegradation. The oxygen generating capabilities of new electrolytic cells and cells with light and heavy calcium carbonate precipitates on the cathode were evaluated in the laboratory under current densities ranging from 0.5 to 5.0 mA/cm2. Higher current densities resulted in higher mass transfer coefficients (K(L)a) and greater saturation oxygen concentrations (Csat). As the cathodic deposits increased, the K(L)a tended to decrease and the Csat tended to increase. The oxygen transfer efficiency (OTE) did not vary as a function of current density or cathode coating, while the average OTE for all the tests was 67%. Laboratory column tests showed that chlorine production increased with current density and depended on chloride levels in the water. Hydrogen peroxide was generated at low concentrations (< 1 mg/L) and at higher levels when chloride was absent in the feed solution. Calcium removal from solution increased with current density and resulted in a decrease in solution pH. Tests at a field monitoring well showed average Csat levels of 16.9 mg/L after 14 days of operation, no chlorine production because of low chloride levels in the well, artificially elevated hydrogen peroxide levels because of background interferences, and a pH decrease of 2.4 units. With passive venting, the average hydrogen gas levels at the headspace of the well were less than 1%. PMID- 12108718 TI - NMR spectroscopy for determination of cationic polymer concentrations. AB - Organic polyelectrolytes are utilized extensively in wastewater treatment, but their fate after use is poorly understood. Analytical methods used for polymer determination in less complex systems appear to fail in application to wastewater systems, contributing to the lack of knowledge. Thus, the development of 1H NMR spectroscopy is reported here for specifically quantitating certain cationic flocculant polymers in environmental samples. Proton observe frequencies of 250 or 400 MHz proton were used. A copolymer of acrylamide and acryloyloxyethyltrimethylammonium chloride was used, representative of cationic flocculant polymers possessing quaternary ammonium groups with terminal methyls that provide a sharp singlet at a chemical shift of approximately 3.06 ppm. A strong linear relationship was demonstrated between polymer concentration and either height or area of this peak. Recoveries were up to 96% at higher concentrations (250 mg/L), and were greater than when using viscosity or charge titration methods for polymer determination. Lesser recoveries at lower concentrations (70% at 5 mg/L) were attributed to adsorptive losses. The detection limit of this method was determined to be <0.5 mg/L. Use of the method was exemplified by analysis of anaerobically digested sludges for residual polymer following a range of dosages, showing the resulting isotherm. PMID- 12108719 TI - Diagnosing reservoir water quality using self-organizing maps and fuzzy theory. AB - Since trophic status assessment of water quality is very important for the water resources management, the assessment results obtained from using only one parameter may easily mislead or bias the decision makers or managers. Even when using a multivariable index, how to determine the weights of all factors is debatable. In this research, one complementary evaluation method, self-organizing map (SOM), for diagnosing water quality has been used to develop a trophic state classifier and is illustrated with a case study of trophic status assessment for Fei-Tsui Reservoir in Taiwan. The historical database was collected from the management agency of Fei-Tsui Reservoir from 1987 to 1995. The results of SOM are compared with those of the Carlson index and Fuzzy synthetic evaluation, showing that the inconsistent records can be mapped to the conflicting data zone of the SOM output map. In addition, SOM creates a diagnostic axis on the map to express the trophic status of the water body. As long as the SOM model is well-trained, new records can be assessed and classified as either one of three trophic levels or speciarcases. If special water quality conditions are expressed on the SOM output, those data can reveal that either total phosphorus (TP) or chlorophyll A (chl a) is higher than usual. PMID- 12108720 TI - Occurrence and alteration of organic contaminants in seepage and leakage water from a waste deposit landfill. AB - Organic-geochemical analyses have been applied to seepage water and leakage water samples of a waste deposit landfill in order to give a comprehensive view on the composition of the organic contaminants. Based on intense GC/MS screening analyses a wide variety of organic substances were identified and attributed to natural or xenobiotic waste components. Apart from plant material-derived compounds and degradation products of peptides, carbohydrates and lignin, numerous xenobiotic substances were identified and attributed to the groups of pharmaceuticals, plasticizers, pesticides or chlorinated aromatics. Not all of the substances identified in the seepage water samples were recovered in the leakage water sample due to degradation processes or dilution by uncontaminated water. Quantitative analysis of selected contaminants was used to discriminate substances affected by degradation processes and persistent compounds. PMID- 12108721 TI - Microbial degradation of quinoline by immobilized cells of Burkholderia pickettii. AB - A quinoline-biodegrading microorganism was isolated from activated sludge of coke oven wastewater treatment plant using quinoline as sole carbon and nitrogen source. It is a gram negative, rod-shaped and aerobic strain, which was identified as Burkholderia pickettii. The biodegradation of quinoline was carried out with this isolated strain. Analysis by high performance liquid chromatography and gas chromatography/mass spectrum (GC/MS) revealed that 2-hydroxyquinoline (2 OH-Q) was the first intermediate in the course of quinoline biodegradation. A novel immobilization carrier, that is, polyvinyl alcohol (PVA)-gauze hybrid carrier, was developed. The isolated strain was immobilized by two different immobilizing techniques and used for the quinolinerdegradation. It was found that biodegradation rate of quinoline by the microorganisms immobilized on PVA-gauze hybrid carrier was faster than that by the microorganisms immobilized in PVA gel beads. Kinetics of quinoline biodegradation by cells of Burkholderia pickettii immobilized on PVA-gauze hybrid carrier was investigated. The results demonstrate that quinoline degradation could be described by zero-order reaction rate equation when the initial quinoline concentration was in the range of 50-500 mg l(-1). PMID- 12108722 TI - Catalytic degradation of CH2O and C6H5CH2OH in wastewaters. AB - The heterogeneous oxidation of benzyl alcohol and formaldehyde in aqueous medium has been investigated. The catalytic oxidation of these compounds is carried out in the presence of Ni-oxide system at ambient temperature. The results show that under studied conditions, a 90% conversion of CH2O to CO2 and a complete oxidation of C6H5CH2OH to C6H5COOH is achieved. Addition of H2SO4 to adjust pH to 2 further precipitates benzoic acid. The precipitate is filtered and the resulting filtrate is free of organic substances (COD is below 10 mg O2 dm(-3)). Based on the results obtained, a technology for purification of wastewaters containing benzyl alcohol as well as a catalytic method for degradation of CH2O in aqueous solutions have been developed. PMID- 12108723 TI - Single- and multi-component adsorption of cadmium and zinc using activated carbon derived from bagasse--an agricultural waste. AB - The use of low-cost activated carbon derived from bagasse, an agricultural waste material, has been investigated as a replacement for the current expensive methods of removing heavy metals from wastewater. With a view to find a suitable application of the material, activated carbon has been derived, characterized and utilized for the removal of cadmium and zinc. The uptake of cadmium was found to be slightly greater than that of zinc and the sorption capacity increases with increase in temperature. The adsorption studies were carried out both in single- and multi-component systems. Adsorption data on derived carbon follows both the Freundlich and Langmuir models. The data are better fitted by the Freundlich isotherm as compared to Langmuir in both the single- and multi-component systems. Isotherms have been used to obtain the thermodynamic parameters. The kinetics of adsorption depends on the adsorbate concentration and the physical and chemical characteristics of the adsorbent. Studies were conducted to delineate the effect of temperature, initial adsorbate concentration, particle size of the adsorbent and solid-to-liquid ratio. On the basis of these studies, various parameters such as mass transfer coefficient, effective diffusion coefficient, activation energy and entropy of activation were evaluated to establish the mechanisms. It was concluded that the adsorption occurs through a film diffusion mechanism at low as well as at higher concentrations. PMID- 12108724 TI - Analysis of the antioxidant butylated hydroxytoluene (BHT) in water by means of solid phase extraction combined with GC/MS. AB - The antioxidant 3,5-di-tert-butyl-4-hydroxy-toluene (BHT) is widely used as an additive to increase the tenability of food and plastics. BHT is degraded to 3,5 di-tert-butyl-4-hydroxybenzaldehyde (BHT-CHO) in mammals, as well as in the natural environment such as in water and soils. BHT-CHO has been studied extensively in terms of their potential toxicities. The present investigation was carried out to quantify BHT and BHT-CHO in river, ground, rain and drinking water obtained from several locations in Germany. Apart from the compounds mentioned above, 1,2-bis-(3,5-di-tert-butyl-4-hydroxyphenyl)ethane (2-BHT), which is a dimer of BHT, was also detected in the extracts of some ground water samples. The applied analytical method is based on solid phase extraction (SPE) to concentrate trace compounds from water samples followed by gas chromatography/mass spectrometry (GC/MS) of the extracts. A total of 51 of the respective water samples were used for extraction purposes and analyte recoveries were all > or = 80%. The determination limit for BHT was 5 ng l(-1) and for BHT-CHO 16 ng l(-1). The standard deviations for the analytical procedure were 6% for BHT and 10% for BHT-CHO. The use of the antioxidant BHT in Germany has resulted in water concentrations of 7-791 ng l(-1) in the rivers Rhine, Elbe, Main, Oder, Nidda and Schwarzbach. The degradation product BHT-CHO was also detected in the river water samples at concentrations between 29 and 223 ng l(-1). The concentrations of BHT measured in German rivers are lower compared to values measured in the USA and Japan 20 years ago. In ground water, levels for BHT varied from non-detectable up to 2156 ng l(-1) and for BHT-CHO from non-detectable up to 674 ng l(-1). Both compounds were also detected in rain water in Frankfurt/Main at a concentration of 1797 ng l(-1) for BHT and 59 ng l(-1) for BHT-CHO. PMID- 12108725 TI - Adsorption of natural organic matter oxidized with ClO2 on granular activated carbon. AB - The paper describes the influence of the oxidation of natural organic matter (NOM) molecules with chlorine dioxide (ClO2) on granulated activated carbon (GAC) adsorption. In order to determinate the influence of ClO2 dosage on the NOM adsorption on GAC two parallel pilot scale experiments were performed. The raw water was treated respectively with 0.2 and 0.4 mg ClO2 L(-1) followed by the adsorption on GAC filters. Experiments were total organic carbon (TOC) measurements and size exclusion chromatography (SEC) controlled. The molecular weight distribution of NOM in the filtration bed outlet demonstrates that the low molecular weight molecules are less retained than the higher molecular weight components of NOM. It is shown that low molecular weight NOM causes less ClO2 demand. The oxidation of NOM molecules and very high capacity of GAC filter bed for NOM components can be used to control high ClO2 demand. PMID- 12108726 TI - Operating conditions for the determination of the biochemical acidogenic potential of wastewater. AB - The aim of this work was to study the test conditions for the determination of the biochemical acidogenic potential (BAP) of wastewater, which should be useful to predict the performance of enhanced biological phosphorus removal (EBPR). Proposed operating conditions for a simple and reproducible BAP test in 250 ml serum bottles (equipped with black butyl stoppers and magnetic bars) are: use of either frozen or fresh water, no inoculum addition, fermentation carried out in the dark during 15 days, addition of 1 mM bromo-ethane sulfonate (BES) and 2 mM barium chloride (BaCl2), stirring speed strong enough to maintain vortex conditions, no pH control and controlled temperature of 20 degrees C. PMID- 12108727 TI - The transient-state, multiple-species biofilm model for biofiltration processes. AB - We describe the transient-state, multiple-species biofilm model (TSMSBM), which is a novel synthesis of key modeling features needed to describe multiple-species biofilms that experience time-varying conditions, particularly including periodic detachment by backwashing. The TSMSBM includes six features that are essential for describing multiple-species biofilms that undergo changes over time: (1) four biomass types: heterotrophs, ammonia oxidizers, nitrite oxidizers, and inert biomass; (2) seven chemical species: input biodegradable organic material (BOM), NH4(+)-N, NO2(-)-N, NO3(-)-N, utilization-associated products, biomass-associated products, and dissolved oxygen; (3) eight reactions that describe the rates of consumption or production of the different species, as well as the stoichiometric linkages among the rates; (4) reaction with diffusion of all the soluble species in the biofilm; (5) growth, decay, detachment, and flux of each biomass type by location in the biofilm; and (6) constant or periodic detachment of biofilm, both of which allow for protection of biomass deep inside the biofilm. The last two features of the TSMSBM provide novel additions to biofilm modeling, and the synthesis of all features is a unique advancement. A series of examples illustrates insights that the TSMSBM can provide about the transient development of multiple-species biofilms; the roles of soluble microbial products and detachment in controlling the distribution of biomass types and process performance; and how backwashing affects the biofilm in drinking-water biofiltration. PMID- 12108728 TI - A comparison of surface water natural organic matter in raw filtered water samples, XAD, and reverse osmosis isolates. AB - This research compared raw filtered waters (RFWs), XAD resin isolates (XAD-8 and XAD-4), and reverse osmosis (RO) isolates of several surface water samples from McDonalds Branch, a small freshwater fen in the New Jersey Pine Barrens (USA). RO and XAD-8 are two of the most common techniques used to isolate natural organic matter (NOM) for studies of composition and reactivity; therefore, it is important to understand how the isolates differ from bulk (unisolated) samples and from one another. Although, any comparison between the isolation methods needs to consider that XAD-8 is specifically designed to isolate the humic fraction, whereas RO concentrates a broad range of organic matter and is not specific to humics. The comparison included for all samples: weight average molecular weight (Mw), number average molecular weight (Mn), polydispersity (rho), absorbance at 280 nm normalized to moles C (epsilon280) (RFW and isolates); and for isolates only: elemental analysis, % carbon distribution by 13C NMR, and aqueous FTIR spectra. As expected, RO isolation gave higher yield of NOM than XAD-8, but also higher ash content, especially Si and S. Mw decreased in the order: RO > XAD-8 > RFW > XAD-4. The Mw differences of isolates compared with RFW may be due to selective isolation (fractionation), or possibly in the case of RO to condensation or coagulation during isolation. 13C NMR results were roughly similar for the two methods, but the XAD-8 isolate was slightly higher in 'aromatic' C and the RO isolate was slightly higher in heteroaliphatic and carbonyl C. Infrared spectra indicated a higher carboxyl content for the XAD-8 isolates and a higher ester:carboxyl ratio for the RO isolates. The spectroscopic data thus are consistent with selective isolation of more hydrophobic compounds by XAD-8, and also with potential ester hydrolysis during that process, although further study is needed to determine whether ester hydrolysis does indeed occur. Researchers choosing between XAD and RO isolation methods for NOM need to consider first the purpose of the isolation; i.e., whether humic fractionation is desirable. Beyond that, they should consider the C yield and ash content, as well as the potential for alteration of NOM by ester hydrolysis (XAD) or condensation/coagulation (RO). Furthermore, the RO and XAD methods produce different fractions or isolates so that researchers should be careful when comparing the compositions and reactivities of NOM samples isolated by these two different techniques. PMID- 12108729 TI - Natural organic matter (NOM) in a limed lake and its tributaries. AB - The chemistry of a limed lake and its main tributaries were studied for 3 years (1992-94) with an emphasis on natural organic matter (NOM). Increased transparency and decreased water colour indicated a general reduction of NOM in the lake. Increased A(254 nm)/A(410 nm) ratios in the epilimnion during summer and early autumn suggested degradation of higher molecular size organic matter into low molecular size NOM. Increase in ammonium and dissolved inorganic carbon concentrations in the lake was possibly due to the NOM degradation. Using budget calculations and the literature values, photodegradation and microbial activity were estimated to be the main mechanisms of the NOM removal. These mechanisms accounted for about 30-35% and 60-65% of the total loss of organic matter, respectively, in the summer and early autumn period. Low sedimentation rates indicate that co-precipitation of organic matter with calcium, aluminium and/or iron was of minor importance in these seasons. PMID- 12108730 TI - Marine environmental pollution stress detection through direct viable counts of bacteria. AB - Direct viable counts (DVC) of bacteria were quantified from polluted and relatively less/non-polluted coastal locations during different seasons to assess whether they can be routinely monitored for an understanding of environmental stress(es) that may impede the full functioning of bacterial communities in situ. Most notably, DVC were quite low during pre-monsoon (March-May) in pollution affected locations when compared to relatively less/non-polluted ones. In contrast, their abundance was significantly higher (up to or > 10%) suggesting a substantially higher microbial activity (thus, a larger turnover of organic matter) during monsoon (June-September) and post-monsoon (October-February) even in pollution-affected locations. The ease of reliably measuring DVC was useful in realising decreased metabolic functioning of bacteria during pre-monsoon, a season where dispersion of land discharges/effluents is much lower. From laboratory and field analyses of this study it is ascertained that DVC are direct indices of potential bacterial metabolic activity, reliable for sensing metabolic stress experienced by bacterial communities in situ and can be useful for evaluating risks in marine environment through human (industrial) activities. PMID- 12108731 TI - Poorly humified peat as an adsorbent for metals in wastewater. AB - Metal adsorption and surface charge determinations were performed previously on well-characterised Sphagnum and Carex peat samples. The aim of this investigation was to determine metal adsorption from complex wastewaters onto these peat samples and compare it to the adsorption onto peat granules, clinoptilolite, glauconite and a flue dust from steel production. A sulphide mine leachate, a landfill leachate and a laundry wastewater were chosen, giving a variation in pH, ionic strength, total organic carbon and concentrations of metals. Metal adsorption was determined in batch and column experiments. The wastewater composition was of great importance for metal removal efficiency, mainly due to the difference in dominating metal species. In the sulphide mine leachate, containing free metal ions, a high metal adsorption was observed onto both peat and inorganic adsorbents. In the landfill leachate the metals formed carbonate and organic complexes and a low metal removal was achieved. Contrary to the leachates, the laundry wastewater contained suspended particles. The high amount of metals removed, 80% of the Cu and 30-60% of the Zn concentration, was probably withdrawn bound to the particle fraction. The highest removal of metal ions was obtained in the sulphide mine leachate with Carex peat, removing 97-99% of the Zn and 85-100% of the Cu content. The Sphagnum peat sample removed 37-77% of the Zn and 80-100% of the Cu content. The differences found between Sphagnum and Carex peat were attributed to the original chemistry of the plant material and the habitat conditions at the time of peat formation. Generally, the combination of glauconite or clinoptilolite with the peat samples in column experiments gave a minor improvement in metal removal. PMID- 12108732 TI - Effect of mass transfer on concentration wave propagation during anaerobic digestion of solid waste. AB - A distributed model of anaerobic digestion of solid waste was developed to study effects of mass transfer on the rate of propagation of initiation methanogenic area. The diffusion and advection of volatile fatty acids (VFA) and methanogenic biomass were taken into account in the model of a one-dimensional (ID) reactor. It was considered that VFA inhibits both polymer hydrolysis and acetoclastic methanogenesis. This approach allows to view the bioreactor as an active medium that provokes concentration waves from some area of methanogenic initiation (local VFA depression) to the total reactor volume. The model shows that mass transfer-based acceleration of methane production in the reactor is possible when the intensity of VFA utilization in the methanogenic area is sufficient for a complete digestion of the incoming VFA. Otherwise, initiation methanogenic area will be suppressed by increasing concentration of VFA. The obtained results emphasize the importance of considering spatial heterogeneity of the reaction for the analysis of solid anaerobic digestion in bioreactors and landfills. The digestion of solid waste can be optimized by setting the low rate of mass transfer (mixing or leachate recirculation) during lag-phase of the reaction with subsequent increase in the mass transfer rate in parallel with the propagation of methanogenic population. In this case, the rate of concentration waves substantially increases. PMID- 12108733 TI - Fate of Cryptosporidium oocysts in an immobilised titanium dioxide reactor with electric field enhancement. AB - We have undertaken simple proof of principle experiments to find out if electric field enhanced photo-oxidation using immobilised titanium dioxide will damage Cryptosporidium oocysts. Using a simple Petri dish reactor and two forms of immobilised titanium catalyst (sol-gel and thermal-film) we have tested the ability of this technology to affect Cryptosporidium oocysts permeability assessed by propidium iodide exclusion. Test and control reactor runs were significantly different (P = 0.007). The thermal-film reactor had the greatest effect (approximately 27% of the seed) and was statistically distinguishable from the sol-gel reactor and the controls. The sol-gel reactor showed an increase in oocyst permeability, but was not statistically distinguishable from one of the controls. The enhanced performance of the thermal film reactor is attributed to the superior conversion of photochemical holes to hydroxyl radicals at the surface of this catalyst. PMID- 12108734 TI - Flocculation of river silt using chitosan. AB - Flocculation of silt in river water using chitosan was studied in the pH range 4 9, and suspended solid concentrations in the range 20-80 mg/L. Chitosan effectively reduces turbidity due to silt by flocculation and settling. Flocculation efficiency is very sensitive to pH, and reaches a maximum at pH 7. The optimal chitosan concentration required to effect flocculation is 0.5 mg/L and is independent of silt concentration within the range examined. Restabilisation of the suspension is observed at higher concentrations of chitosan, and the amount required for restabilisation increases with increasing concentration of suspended solids. Flocculation is faster at higher concentrations of silt and the flocs are large and fibrous. PMID- 12108735 TI - F-specific RNA coliphages: occurrence, types, and survival in natural waters. AB - A small, well-defined watershed was investigated over a 2-year period to determine the prevalence of F-specific RNA coliphage (F + RNA) serotypes as indicators of animal fecal contamination. Sampling sites collected runoff from areas of urban and agricultural land use patterns. F-specific coliphages were concentrated from 2-L freshwater samples by polyethylene glycol precipitation, isolated using the double agar layer (DAL) method, confirmed as F + RNA by RNAse suppression, and serotyped. A subset of serotyped F + RNA were confirmed by genotyping. To determine relative survival, 10 confirmed F + RNA field isolates and 5 prototypic F + RNA were spiked into surface water and incubated at 25 degrees C for 36 days. F-specific coliphage isolation was strongly associated with rainfall events and was infrequent from primarily animal impacted surface waters. Field isolates were predoffiinantly Type I F + RNA (81%) and raw sewage isolates were predominantly Type III F + RNA (57%). Genotyping from either the watershed or raw sewage samples never positively identified Type IV F + RNA. Results from laboratory studies showed that F + RNA differ in their survival in water and that Type IV strains were the least persistent. Type III F + RNA were found to be reliably related to the release of uncontrolled human fecal material in the watershed, but the results of this study suggest that further study is required before utilizing for fecal source identification in natural waters. PMID- 12108736 TI - The clinical applicability of automated gait analysis systems. PMID- 12108737 TI - Medication: a way forward from zero tolerance to irrelevant plasma concentrations. PMID- 12108738 TI - Immunological and haematinic consequences of feeding a standardised Echinacea (Echinacea angustifolia) extract to healthy horses. AB - This study was undertaken to compile new data on the efficacy of Echinacea in stimulating the immune system of the horse. Use of Echinacea is becoming widespread in horses, despite an absence of controlled laboratory research into its effectiveness or safety. This paper documents results of a double-blind, placebo-controlled, cross-over trial investigating the effect of standardised Echinacea extract on 8 horses. Animals were supplemented with Echinacea or placebo for 42 days, and their response to supplements recorded. Treatment with Echinacea increased phagocytic ability of isolated neutrophils, boosted peripheral lymphocyte counts and appeared to stimulate neutrophil migration from peripheral circulation into the tissues. Echinacea supplement also increased the size and concentration of peripheral red blood cells, and the concentration of haemoglobin and packed cell volume. It was concluded that Echinacea effectively stimulates equine immunocompetence, and the plant extract behaves, in equine subjects, as a haematinic agent, i.e. one which improves the quality of blood by increasing haemoglobin levels and the number of erythrocytes and which, by virtue of their effects on oxygen transport cells, are considered to improve parameters of exercise physiology and performance. PMID- 12108739 TI - A study of the ultrastructure and staining characteristics of the 'dental star' of equine incisors. AB - The objective of this study was to examine the diameter, extent, orientation and contents of dentinal tubules in order to validate the hypothesis of pigment penetration into the dental star of equine incisival occlusal surfaces. The time of appearance and the configuration of the dental star on the incisival occlusal surface are macroscopically visible features that, along with other more reliable parameters, are used for the determination of horses' age. Although dental stars are an integral part of the equine incisor occlusive surface, the exact nature and microstructure of the dental star are poorly documented. Therefore, equine incisor dentine was examined macroscopically and by scanning electron microscopy to elucidate numerical density, diameter and 3-dimensional organisation of the dentinal tubules in the dental star. The dental star is surrounded by primary dentine and consists of a central core of tertiary dentine, an intermediate ring of pale secondary dentine and a peripheral rim of darker, yellowish-brown secondary dentine. The central core of tertiary dentine contains relatively few dentinal tubules (<8000/mm2) that have small diameters (mean +/- s.d. 1.67 +/- 034 microm) and are arranged in an irregular pattern. The surrounding pale ring of secondary dentine comprises manifestly more and wider tubules that lie almost parallel to the occlusal surface. The dark peripheral rim of the dental star contains high numbers of tubules (28,000-58,000/mm2) that have wide luminal diameters (mean +/- s.d. 3.09 +/- 0.31 microm) and open perpendicular to the occlusal surface. In contrast, the primary dentine surrounding the dental star is made up by a lower number of dentinal tubules (<25,000/mm2). The tubules of primary dentine, which are initially mean +/- s.d. 5.15 +/- 0.80 microm wide, are narrowed by circumferential deposits of peritubular dentine and are obliquely exposed at the occlusal surface. From these observations, it was concluded that the regional differences in numerical density, diameter and spatial orientation of the dentinal tubules may influence the penetration of food pigments into the equine occlusal surface and result in the particular staining of the dental star. PMID- 12108740 TI - Repeatability of back kinematics in horses during treadmill locomotion. AB - We tested the hypothesis that repeatability of a standardised protocol for quantifying back kinematics is sufficiently high not to prevent its use in the clinical evaluation of horses with back problems. We investigated the extent to which differences between laboratories may affect the results when a standardised protocol is used. As a clinical tool, movement analysis techniques are helpful for the objective and quantitative assessment of kinematics. Knowledge about the repeatability of the kinematic data is very important. The present study investigates the repeatability of back kinematics in 10 sound horses over 5 successive days and in 2 laboratories (5 horses at each location). Measurements were performed on the treadmill during the walk and the trot. The between-stride, between-day and between-horse repeatability were determined. A high degree of between-stride and between-day repeatability was observed in the spatiotemporal parameters and in the time-angle diagrams of thoracic and lumbar vertebrae, the sacrum and the hindlimb during both the walk and trot. Much more variability was found between horses, with the highest degree of dissimilarity in the lateral bending rotation of the L1 vertebra. For range of motion values, the between-day coefficient of variability was <14% and the between-horse coefficient of variability was up to 4 times higher. Small differences were found in range of motion values between the 2 laboratories. It is concluded that an analysis of back kinematics in the horse can provide highly repeatable data, warranting clinical use. PMID- 12108741 TI - Pharmacokinetic/pharmacodynamic approach to assess irrelevant plasma or urine drug concentrations in postcompetition samples for drug control in the horse. AB - The current performance of analytical techniques used for drug control in horses lead the Regulatory Authorities to decide whether trace levels of drugs legitimately used for therapeutic medication should or should not be reported. Here, we propose a well-ordered and nonexperimental pharmacokinetic/pharmacodynamic approach for the determination of irrelevant drug plasma (IPC) and urine concentrations (IUC). The published plasma clearance is used to transform an effective (marketed) dose into an effective concentration (EPC). EPC is transformed into an IPC by applying a safety factor (SF). This method is based on several assumptions (eg, drug effects reversibly driven by plasma concentration, linearity of drug disposition). The suitability of the computed IPC and IUC can be checked by calculating the residual amount of drug at IPC and computing a minimal drug withdrawal time. It is concluded that controlling the drug effect (using drug or any analyte concentration as a marker) rather than the drug exposure will be more demanding and also makes urine a less than ideal matrix. PMID- 12108742 TI - Parenteral nutrition for horses with gastrointestinal disease: a retrospective study of 79 cases. AB - Parenteral nutrition is advocated for sick foals and horses, although there is no report which has critically evaluated its benefits in this species. Therefore, the hypothesis that parenteral nutrition (PN) is beneficial for horses with severe gastrointestinal disease was investigated in a retrospective study. Records from 79 treatment courses in horses with gastrointestinal disease were reviewed for the years 1992-2000. The fatality rate (48.1%) was attributed to the severity of the primary disease. Complications due to PN were recorded and hyperglycaemia was the most common complication associated with PN. The number of laparatomies, proportion of horses that received lipid emulsion and the daily cost were higher in nonsurvivors than in survivors. The length of PN course was longer in survivors, although glucose peak was observed later in nonsurvivors after initiation of PN. The content of vitamins in PN solution and plasma protein at the beginning of PN were higher for survivors, while the volume of plasma administered was significantly larger in nonsurvivors. In horses with inflammatory diseases, the rate of glucose infusion and the amount of vitamins administered were higher than in horses with nonstrangulated obstruction. The proportion of horses that received lipid emulsion was higher among those with nonstrangulated and strangulated obstruction than in the group with inflammatory conditions. Because of the diverse group of diseases and the variability in the clinical signs, evaluating the effect of PN on disease outcome was not possible. A prospective study to compare horses with similar clinical conditions treated and not treated with PN is needed to evaluate fully the benefits of PN, and to establish guidelines for patient selection for PN. PMID- 12108743 TI - Normal ultrasonographic anatomy and injury of the patellar ligaments in the horse. AB - The purposes of this study were to investigate the ultrasonographic appearance of the patellar ligaments in clinically normal horses and to describe the clincical features, diagnosis and outcome of patellar ligament injury. The medial, middle and lateral patellar ligaments of 5 Thoroughbred and 5 Warmblood horses, free from lameness and in full work, were examined ultrasonographically. The ligaments were all of uniform echogenicity and each ligament was fairly consistent in its shape. The size of the ligaments of the Warmblood horses tended to be bigger than the lighter bodyweight Thoroughbred horses. The bone surfaces were smooth at the ligament insertions on the patella and tibia. Nine horses, including 7 showjumpers, were identified with a unilateral hindlimb lameness associated with ultrasonographic evidence of damage to one or more patellar ligaments. Four horses had primary desmitis of the middle patellar ligament, one of which had concurrent intermittent upward fixation of the patella and a second had abnormal movement of the patella. Two additional horses had desmitis of the middle patellar ligament associated with previous medial patellar desmotomy. Two horses had desmitis of both the middle and lateral patellar ligaments, and one horse had desmitis of the lateral patellar ligament alone. None of the 9 horses were able to return consistently to their former level of competition. Careful clinical evaluation and ultrasonographic examination of the patellar ligaments should be considered in horses with hindlimb lameness of otherwise undetermined cause. PMID- 12108744 TI - Development of biochemical heterogeneity of articular cartilage: influences of age and exercise. AB - The objective of this study was to document the development of biochemical heterogeneity from birth to maturity in equine articular cartilage, and to test the hypothesis that the amount of exercise during early life may influence this process. Neonatal foals showed no biochemical heterogeneity whatsoever, in contrast to a clear biochemical heterogeneity in mature horses. The process of formation of site differences was almost completed in exercised foals age 5 months, but was delayed in those deprived of exercise. For some collagen-related parameters, this delay was not compensated for after an additional 6 month period of moderate exercise. It is concluded that the functional adaptation of articular cartilage, as reflected in the formation of biochemical heterogeneity in the horse, occurs for the most part during the first 5 months postpartum. A certain level of exercise seems essential for this process and withholding exercise in early life, may result in a delay in the adaptation of the cartilage. PMID- 12108745 TI - Retrospective study of primary intention healing and sequestrum formation in horses compared to ponies under clinical circumstances. AB - In accidental wounds, trauma and infection can result in dehiscence of primarily closed wounds and in sequestrum formation when cortical bone is exposed. In experimental studies, it has been shown that second intention healing is faster and occurs with less complications in ponies than in horses. Also, a greater initial inflammatory response was seen in ponies. Based on these experimental data, it was hypothesised that accidental wounds in ponies would heal with a lower incidence of wound dehiscence and/or sequestrum formation compared to horses. A retrospective study of 89 ponies and 422 horses with traumatic wounds was performed. The animals, wounds and treatments were categorised and related to the success rate of primary closure and to the incidence of sequestrum formation. The ponies and horses were of similar age and sex. The wounds that were treated were comparable for localisation, duration, degree of contamination and depth in both groups of animals, but there were significantly more cases with ruptured extensor tendons in ponies. Antibiotics and NSAIDs were administered significantly less often to ponies. The success rate of primary closure was significantly higher in ponies than in horses, and sequestra were formed significantly less often in ponies. It was concluded that the results of healing were better in ponies although the external conditions were less favourable. This may be associated with the differences in the initial inflammatory response after injury as found in earlier experimental work, which may result in a better local defence against wound infection. PMID- 12108746 TI - Age-related morphometry of equine calcified cartilage. AB - Although there are many studies in the equine literature focused on articular diseases and the aetiology of osteoarthritis, few have concentrated on normal articular structures and how they change with age. The objective of this investigation was to study the thickness and morphology of the calcified cartilage layer of the distal metacarpus over a range of ages. A parasagittal slab of bone was sectioned from the region of sesamoid contact on the medial condyle of the metacarpi from 34 horses. The slab of bone was preserved, dehydrated and embedded, undecalcified, in methylmethacrylate and then stained with toluidine blue. Six repeatable fields of interest from the distal aspect of each metacarpus were digitised and examined to determine the morphology of the calcified cartilage layer. The thickness of the calcified cartilage, range 88-426 microm, was estimated using a method of integration. The results indicate an age related influence on the thickness of the calcified cartilage layer, generally increased in older horses. While this finding is significant, perhaps more importantly a positional relationship was also identified, indicating that pressures endured by different regions within a joint may dictate morphological development of the tissues. This study has begun to lay the groundwork to determine whether the calcified layer of the hyaline cartilage could be involved in the development of osteoarthritis. PMID- 12108747 TI - Chemoattractant properties of conditioned medium from equine corpora lutea collected at various stages of the oestrous cycle. AB - This study investigated the chemotactic activity of equine CL at different stages of the oestrous cycle. The purpose of this was to ascertain whether luteal tissue itself contributes to the massive influx of leucocytes around the time of natural and induced luteal regression. Corpora lutea were collected at different stages of dioestrus and after treatment with PGF2alpha. Culture medium harvested after incubation of luteal tissue for 20 h was chemotactic for both polymorphonuclear and mononuclear cells in late dioestrus (before functional regression) as well as after natural and induced luteal regression. By contrast, midluteal tissue showed no chemotactic activity. This is the first report of the ability of equine luteal tissue actively to recruit inflammatory cells in vitro and supports our earlier findings that this infiltration starts prior to functional luteolysis. We hypothesise that this early influx of inflammatory cells may play an active role in luteal regression. Further research is needed to identify the specific chemotactic factor(s). PMID- 12108748 TI - Prednisone per os is likely to have limited efficacy in horses. AB - Based on its efficacy for the treatment of human asthma, the corticosteroid prednisone is commonly used in horses for treatment of recurrent airway obstruction. However, recent studies have failed to show any benefit of prednisone tablets for the treatment of this condition. The purpose of this study was to determine why oral prednisone has poor efficacy for the treatment of heaves in horses. In a crossover study, 5 horses were given the following treatments: prednisone tablets, prednisone liquid, prednisolone tablets, prednisolone liquid and i.v. prednisolone sodium succinate (positive control). Blood samples were taken before drug administration and at selected time points during a 24 h period. Serum concentrations of prednisone and prednisolone were determined in order to evaluate gastrointestinal absorption and hepatic metabolism. Serum concentrations of the endogenous glucocorticoid hydrocortisone were also determined as an indicator of the biological activity of the drugs. Both prednisolone tablets and liquid were absorbed rapidly, with prednisolone detectable in serum within 15 min of administration and with peak concentrations occurring within 45 min. Small amounts of prednisone were detected in the serum samples after administration of both prednisone tablets and liquid. Prednisolone was not detected in serum samples after administration of prednisone liquid and was detected in serum samples from only one horse after administration of prednisone tablets. Endogenous hydrocortisone production was suppressed when horses received prednisolone. The results of these studies indicate that prednisone has poor efficacy for the treatment of heaves because it is poorly absorbed and the active metabolite prednisolone is rarely produced. In contrast, prednisolone tablets have excellent bioavailability and should be useful as a therapeutic agent in horses. PMID- 12108749 TI - Cytological analysis of equine bronchoalveolar lavage fluid. Part 1: Comparison of sequential and pooled aliquots. AB - The aim of this study was to investigate whether initial equine bronchoalveolar lavage fluid (BALF) aliquots were more representative of bronchial cytology that bronchiolar and alveolar cytology. Cell viability and total nucleated (TCC), differential (DCC) and absolute cell counts of cytocentrifuged preparations of 3 sequentially collected BALF aliquots (Aliquots 1-3) were compared with those of pooled BALF (Aliquot 4) to assess whether all aliquots were representative of the lavaged lung segment. BALF samples (n = 21) were collected from control horses (n = 5) or heaves-affected horses (n = 5). There were nonsignificant trends of increasing TCC and absolute macrophage count from Aliquot 1 to Aliquot 3 and significant differences in macrophage (P<0.05) and lymphocyte (P<0.01) DCC among aliquots of all horses; however, no linear trend in this DCC data was observed. There was a significant decrease in mast cell DCC (P<0.01) from Aliquot 1 to Aliquot 3 in control horses. Cell viability did not differ significantly among aliquots. There was no diagnostically significant difference in TCC, DCC, absolute cell counts or cell viability, among sequential and pooled BALF aliquots and, therefore, all aliquots can be considered to represent the cytology of the lavaged lung segment. This indicates that even if BALF recoveries are very low, cytological analysis of samples will be of diagnostic value. PMID- 12108750 TI - Cytological analysis of equine bronchoalveolar lavage fluid. Part 2: Comparison of smear and cytocentrifuged preparations. AB - The aim of this study was to develop a diagnostically useful smear method for preparation of equine bronchoalveolar lavage fluid (BALF) for use by practitioners. A smear method for equine BALF preparation which included the addition of serum was developed, and cell morphology, differential cell counts (DCC) and repeatability of counting DCC compared with those of cytocentrifuged BALF preparations. BALF samples (n = 21) were collected from 5 control horses and 5 heaves-susceptible horses. Smear preparations of BALF produced smaller, darker, staining cells, making cytological identification more difficult than on cytocentrifuged preparations. There was a significantly higher (P<0.01) macrophage DCC and lower lymphocyte DCC on cytocentrifuged compared to smear preparations. Mast cell and eosinophil DCC were significantly higher (P<0.05) on cytocentrifuged compared to smear preparations of BALF. Smear preparations were shown to be reliable for the cytological diagnosis of equine neutrophilic pulmonary disease and offer practitioners an alternative to sending equine BALF to a laboratory for processing and cytological analysis. PMID- 12108751 TI - Cytological analysis of equine bronchoalveolar lavage fluid. Part 3: The effect of time, temperature and fixatives. AB - Bronchoalveolar lavage fluid (BALF) samples are often subject to time delays, possibly with temperature fluctuations, between collection and processing. The aim of this study was to evaluate the effects of time, temperature and 2 different fixatives on equine BALF cytology, in order to develop guidelines for optimal equine BALF storage conditions. Total nucleated cell count (TCC), differential cell counts (DCC), absolute cell counts (ACC), cell viability, cell morphology and bacterial growth of BALF samples stored at 4, 18 (+/- addition of formalin- or alcohol-based fixatives) and 38 degrees C were monitored serially over a 72 h period. The time taken for a significant reduction in TCC and cell viability of unfixed BALF samples decreased as the storage temperature increased. There was no diagnostically significant difference in DCC or ACC over this time course at any temperature. Unfixed BALF samples showed significant bacterial growth by 24 h at 4 degrees C, and 8 h at 18 and 38 degrees C; and poor morphology by 48 h at 4 degrees C, 24 h at 18 degrees C and 8 h at 38 degrees C. Fixed BALF samples showed poor morphology with Leishman's stain compared to unfixed samples. PMID- 12108752 TI - Dietary soyabean oil depresses the apparent digestibility of fibre in trotters when substituted for an iso-energetic amount of corn starch or glucose. AB - The aim of the present study was to establish whether the inhibitory effect of fat feeding on fibre digestion has been underestimated due to the substitution of fat for corn starch. A high fat intake has been shown to lower total intestinal tract apparent digestibility of crude fibre in horses but, since fat was substituted for nonstructural carbohydrates, including starch, the specific effect of fat could not be ascertained. The possibility could not be excluded that starch also inhibits fibre digestibility, so that the fat effect observed earlier would have been underestimated. In this study, the intakes of iso energetic amounts of soyabean oil, corn starch or glucose were compared as to fibre digestibility. Unlike starch, glucose is fully absorbed by the small intestine and, therefore, is not expected to influence fibre fermentation in the caecum and colon. Six trotters were fed rations high in soyabean oil (158 g/kg dry matter), corn starch (337 g/kg dry matter) or glucose (263 g/kg dry matter) according to a 3 x 3 Latin square design. Apparent crude fibre digestibility was similar for the rations with corn starch (mean +/- s.d., 70.7 +/- 3.06% of intake, n = 6) or glucose (71.0 +/- 1.90%), but was significantly depressed by fat feeding (56.5 +/- 7.65%). Similar observations were made for apparent digestibilities of neutral and acid detergent fibre and of cellulose. It was concluded that the addition of fat to the feed ration of horses has a specific inhibitory effect on fibre utilisation and, therefore, reduces the amount of energy provided by dietary fibre. PMID- 12108753 TI - The effect of intra-articular methylprednisolone acetate and exercise on equine carpal subchondral and cancellous bone microhardness. AB - Dorsal carpal osteochondral injury is a major cause of lameness in horses undergoing high intensity training. Intra-articular corticosteroid treatment is used commonly to manage exercise-associated articular pain, but its use remains highly controversial in the equine athlete. This project, therefore, aimed to compare the mechanical properties of intra-articular MPA and diluent-treated middle carpal subchondral and cancellous bone in horses undergoing a short-term treadmill exercise programme. It was hypothesised that subchondral and cancellous bone mechanical properties are influenced by intra-articular administration of methylprednisolone acetate (MPA). Eight 2-year-old female horses had MPA or diluent administered into contralateral middle carpal joints at 14 day intervals, for a total of 4 treatments per horse. Horses underwent a standard treadmill exercise protocol until euthanasia (Day 70). Standard sites were located on the dorsal aspect of third, radial and intermediate carpal bones. Osteochondral samples from each test site were divided into subchondral bone and cancellous bone portions. These were dried, resin-embedded and gold-coated. Microhardness measurements were obtained at each test site. No significant effect of intra articular treatment was detected. At each site, cancellous bone trabecular struts had an 18-19% higher microhardness value than the overlying subchondral bone. These findings indicate that intra-articular administration of MPA at this dose has no effect on subchondral or cancellous bone adaptation to short-term exercise and, therefore, on the propensity of carpal bones to injury. Further investigation into the calcified cartilage layer, effect of different corticosteroid preparations and diffusion of medication are required. PMID- 12108754 TI - Folate deficiency during treatment with orally administered folic acid, sulphadiazine and pyrimethamine in a horse with suspected equine protozoal myeloencephalitis (EPM). PMID- 12108755 TI - Bilateral nonossifying fibromas in the proximal tibiae of a yearling Thoroughbred filly. PMID- 12108756 TI - Saliva and gastrointestinal functions of taste, mastication, swallowing and digestion. AB - Saliva has multiple essential functions in relation to the digestive process taking place in the upper parts of the gastrointestinal (GI) tract. This paper reviews the role of human saliva and its compositional elements in relation to the GI functions of taste, mastication, bolus formation, enzymatic digestion, and swallowing. The indirect function of saliva in the digestive process that includes maintenance of an intact dentition and mucosa is also reviewed. Finally, pathophysiological considerations of salivary dysfunction in relation to some GI functions are considered. PMID- 12108757 TI - Current issues in Sjogren's syndrome. AB - Sjogren's syndrome is a chronic autoimmune and rheumatic disorder with prominent sicca complaints from the mucous membranes because of lack of proper exocrine secretions. There is no straightforward and simple diagnostic test for Sjogren's syndrome, although several classification criteria have been designed including several oral diagnostic tests. A new set of classification criteria in a joint effort by research groups in Europe and USA has recently been presented. A large number of autoantibodies have been reported in Sjogren's syndrome where, in some cases, the antibodies are correlated with the extent and severity of disease. The finding of serum autoantibodies directed against the muscarinic M3 receptor is an important advance in understanding the pathogenesis of not only the impaired glandular function but also associated features of autonomic dysfunction in some patients. The treatment of primary Sjogren's syndrome is still mainly symptomatic. PMID- 12108758 TI - The enigmatic mechanism of irradiation-induced damage to the major salivary glands. AB - Irradiation is a central treatment modality administered for head and neck malignancies. Its major and most devastating side-effect is an induced damage to the major salivary glands. This article aims at suggesting a comprehensive explanation for the underlying mechanism of this damage, which has been considered as enigmatic throughout the 90 years since it was first described in 1911. The mechanism suggested is based on the considerable literature concerning this enigma in rat salivary glands. According to this proposed mechanism, the irradiation results in a sublethal DNA damage, which manifests and becomes lethal at a delayed phase. Thus, when the acinar progenitor cells are going through a reproductive phase when parenchymal replenishment is required, they die. The injurious agents, which result in this delayed reproductive cell death, appear to be highly redox-active transition metal ions, such as iron and copper. These metal ions, which seem to be associated with secretion granules, are not necessarily contained within the granules as previously suggested, but rather are probably located at sites more proximal to the DNA. PMID- 12108759 TI - Regulatory mechanisms of osteoblast and osteoclast differentiation. AB - Bone is continuously destroyed and reformed to maintain constant bone volume and calcium homeostasis in vertebrates throughout their lives. Osteoblasts and osteoclasts are specialized cells responsible for bone formation and resorption, respectively. Recent developments in bone cell biology have greatly changed our conceptions of the regulatory mechanisms of the differentiation of osteoblasts and osteoclasts. Bone morphogenetic proteins (BMPs) play critical roles in osteoblast differentiation. The discovery of Smad-mediated signals revealed the precise functions of BMPs in osteoblast differentiation. Transcription factors, Runx2 and Osterix, are found to be essential molecules for inducing osteoblast differentiation, as indicated by the fact that both Runx2-null mice and Osterix null mice have neither bone tissue nor osteoblasts. Smad transcriptional factors are shown to interact with other transcription regulators, including Runx2. Also, the recent discovery of receptor activator of NF-kappaB ligand (RANKL)-RANK interaction confirms the well-known hypothesis that osteoblasts play an essential role in osteoclast differentiation. Osteoblasts express RANKL as a membrane associated factor. Osteoclast precursors that express RANK, a receptor for RANKL, recognize RANKL through the cell-cell interaction and differentiate into osteoclasts. Recent studies have shown that lipopolysaccharide and inflammatory cytokines such as tumor necrosis factor receptor-alpha and interleukin I directly regulate osteoclast differentiation and function through a mechanism independent of the RANKL-RANK interaction. Transforming growth factor-beta super family members and interferon-gamma are also shown to be important regulators in osteoclastogenesis. These findings have opened new areas for exploring the molecular mechanisms of osteoblast and osteoclast differentiation. PMID- 12108761 TI - Enhancement of transmucosal permeation of cyclosporine by benzalkonium chloride. AB - OBJECTIVES: Permeation of cyclosporin A (CsA) through intact and de epithelialized human vaginal mucosa in the presence and absence of benzalkonium chloride (BZCl) was tested. MATERIALS AND METHODS: Human vaginal mucosa (snap frozen in liquid nitrogen, stored at -85 degrees C) had been used for permeability experiments. CsA permeation through thawed frozen intact and de epithelialized vaginal mucosa was determined using a flow-through diffusion apparatus (20 degrees C, 24 h). Flux rates for CsA across these two mucosae were determined in the presence and absence of 0.01% BZCl. ANOVA and Duncan's multiple range test were used to test for steady-state and an unpaired t-test with Welch's correction was used to test for differences between the mean flux values at each time point (significance level of 5%). A piece of thawed tissue from each patient, before and after de-epithelialization, was placed in formalin and histologically examined. RESULTS: Flux rates of CsA across intact vaginal mucosa tended to increase by 28-46% in the presence of 0.01% BZCl, and CsA across de epithelialized mucosa by approximately 28%. The latter differences were statistically significantly higher after 10 h. Flux rates across de epithelialized mucosa were 52-140% higher in the presence of 0.01% BZCl (statistically significantly higher after 12 h). CONCLUSIONS: The permeation of CsA through intact and de-epithelialized human vaginal mucosa can be enhanced by 0.01% BZCl. PMID- 12108760 TI - Inhibitory function of secretory leukocyte proteinase inhibitor (SLPI) in human saliva is HIV-1 specific and varies with virus tropism. AB - OBJECTIVE: Secretory leukocyte proteinase inhibitor (SLPI) is an endogenous mucosa associated protein that has been proposed to possess anti-human immunodeficiency virus type 1 (HIV-1) activity. The aim of this study was to investigate the biological function of SLPI in salivary mediated inhibition of HIV infection and in addition the inhibitory effect of SLPI using isolates of varied virus tropism. MATERIAL AND METHODS: The inhibitory effect of HIV-1 infection in vitro, mediated by 60 different saliva samples was analyzed with respect to levels of SLPI. Salivary samples depleted from IgA and SLPI, respectively, were further analyzed for anti-HIV activity. The antiviral effect of recombinant SLPI was investigated within an in vitro system of HIV-1 infection of target cells using a panel of viral isolates with distinct coreceptor usage. Furthermore we tested a panel of overlapping synthetic peptides, representing the amino acids in SLPI, for their capacity to inhibit HIV-1 infection of human peripheral blood mononuclear cells (PBMCs). RESULTS AND CONCLUSIONS: These experiments show that elevated levels of salivary SLPI can be associated with an increased inhibitory effect of the whole saliva sample, and that this inhibitory effect is decreased with broad coreceptor usage of the virus. PMID- 12108762 TI - Minor recurrent aphthous stomatitis and smoking: an epidemiological study measuring plasma cotinine. AB - OBJECTIVE: To compare blood cotinine levels between a group of patients with minor recurrent aphthous stomatitis (RAS) and a group representative of the general population, matched for age and gender. SETTING: Although smoking has been implicated as a protective factor in RAS, few studies have compared the prevalence of smoking in RAS patients with population controls. Cotinine is a smoking derivative with a long half-life in the circulation. Measurement of plasma cotinine levels provides an accurate and objective measurement of an individual's cigarette consumption. METHODS: Blood samples were taken from 84 RAS patients and 81 controls. A microplate enzyme immunoassay (EIA) was used to detect plasma cotinine levels. RESULTS: The number of RAS patients [2 (2.4%)] who were smokers (plasma cotinine > 25 microg ml(-1) was significantly lower than in the control group [12 (14.8%); P = 0.00102]. Furthermore, the mean cotinine level amongst smokers in the RAS group was significantly lower than in smokers in the control group (P = 0.0068). CONCLUSION: This study shows that a group of RAS patients is significantly less likely to contain smokers than a matched control population, and among smokers the level of cigarette use was significantly lower in RAS patients than the control population. The perceived negative association between RAS and smoking was supported by this epidemiological study. PMID- 12108763 TI - Biosocial studies of antisocial and violent behavior in children and adults: a review. AB - Despite increasing knowledge of social and biological risk factors for antisocial and violent behavior, we know surprisingly little about how these two sets of risk factors interact. This paper documents 39 empirical examples of biosocial interaction effects for antisocial behavior from the areas of genetics, psychophysiology, obstetrics, brain imaging, neuropsychology, neurology, hormones, neurotransmitters, and environmental toxins. Two main themes emerge. First, when biological and social factors are grouping variables and when antisocial behavior is the outcome, then the presence of both risk factors exponentially increases the rates of antisocial and violent behavior. Second, when social and antisocial variables are grouping variables and biological functioning is the outcome, then the social variable invariably moderates the antisocial-biology relationship such that these relationships are strongest in those from benign home backgrounds. It is argued that further biosocial research is critical for establishing a new generation of more successful intervention and prevention research. PMID- 12108764 TI - Response to commentary on the multimodal treatment study of ADHD (MTA): mining the meaning of the MTA. AB - In the December 2000 issue of the Journal of Abnormal Child Psychology, we published a set of papers presenting secondary analyses of the Multimodal Treatment Study of ADHD (MTA), and R. A. Barkley (2000) provided a commentary. A critique of the design of the study (MTA Cooperative Group, 1999) was presented based on a theoretical perspective of a "behavioral inhibition" deficit that has been hypothesized as the core deficit of ADHD (R. A. Barkley, 1997). The commentary questioned the design and analysis of the MTA in terms of (1) the empirical criteria for selection of components of behavioral (Beh) intervention, (2) the effectiveness of the Beh intervention, (3) the methods for analyses at the group and individual level, (4) implications of the MTA findings for clinical practice, (5) the role of genetics in response to treatment, and (6) the lack of a nontreatment control group. In this response, we relate the content of the papers to the commentary, (1) by reviewing the selection criteria for the Beh treatment, as outlined by K. C. Wells, W. E. Pelham, et al. (2000), (2) by addressing the myth that the MTA Beh treatment was ineffective (Pelham, 1999), (3) by describing the use of analyses at the level of the individual participant, as presented by J. S. March et al. (2000) and W. E. Pelham et al. (2000) as well as elsewhere by J. M. Swanson et al. (2001) and C. K. Conners et al. (2001), (4) by relating some of the suggestions from the secondary analyses about clinically relevant factors such as comorbidity (as presented by J. S. March et al., 2000) and family and parental characteristics (as presented by B. Hoza et al., 2000, S. P. Hinshaw et al., 2000, and K. C. Wells, J. N. Epstein, et al., 2000), (5) by discussing the statistical concept of heritability and the lack of a significant difference in the presence of ADHD symptoms in parents of the MTA families compared to parents in the classmate-control families (as presented by J. N. Epstein, et al., 2000), and (6) by acknowledging that an ethically necessary weakness of the MTA design is that it did not include a no-treatment control group. We discuss the use of secondary analyses to suggest how, when, and for what subgroups effectiveness of the Beh treatment may have been manifested. Finally, we invite others to use the large and rich data set that will soon be available in the public domain, to perform secondary analyses to mine the meaning of the MTA and to evaluate theories of ADHD and response to treatments. PMID- 12108765 TI - Adolescent outcomes of childhood conduct disorder among clinic-referred boys: predictors of improvement. AB - Much remains to be learned about the adolescent outcomes of clinic-referred boys whose childhood conduct problems are serious enough to meet diagnostic criteria for conduct disorder (CD). Six structured diagnostic assessments were conducted over 7 years of 73 clinic-referred 7-12-year-old boys who met criteria for CD in Wave 1. There were substantial individual differences in the adolescent outcomes of CD, ranging from worsening to sustained recovery, with most boys showing persistent, but fluctuating levels of CD. Improvement in CD was not accounted for by treatment or incarceration, but more positive outcomes over Waves 2-7 were predicted prospectively with substantial accuracy, using a combination of baseline predictors: less initial severity of CD, fewer symptoms of attention deficit hyperactivity disorder, higher child verbal intelligence, greater family socioeconomic advantage, and not having antisocial biological parents. PMID- 12108766 TI - Suspensions and detentions in an urban, low-income school: punishment or reward? AB - Disciplinary records for 3rd through 8th grade students (n = 314) in an inner city, public school were examined for one school year to assess students' variation in response to discipline. Rates of disciplinary referrals were compared for students who received no detentions or suspensions throughout the year ("never group" n = 117), students who received one or more detention or suspension in the fall but not in the spring ("fall group" n = 62), and students who received one or more detention or suspension in the fall and one or more detention or suspension in the spring ("fall + spring group" n = 75). Results indicated that during the fall, the "fall group" had nearly equivalent rates of referrals to the "fall + spring group"; however, the "fall group" exhibited significantly lower rates of referrals during winter and spring that were nearly equivalent to the "never group," as would be expected for a punishment procedure. In contrast, the "fall + spring group" evidenced increases in referrals across the year, suggesting the possibility that detentions and suspensions were functioning as rewards for this group. The "fall + spring group" was rated by teachers and peers at mid-year as highly aggressive, lacking social skills, and high on hyperactivity, whereas the "fall group" and the "never group" were statistically equivalent on teacher and peer ratings. Implications for mental health programs for urban schools are discussed, especially the need for alternatives to detention and suspension for the subset of students who account for the majority of school discipline. PMID- 12108767 TI - Interparental conflict and risk behaviors among Mexican American adolescents: a cognitive-emotional model. AB - This study used a cognitive-emotional model to examine the relations between multiple dimensions of interparental conflict and health risk behaviors among young adolescents. Participants were 151 Mexican American adolescents and their parents. At initial individual interviews, parents reported on conflict with their spouses, and adolescents reported on their parents' conflict, their appraisals of the conflict, their emotional distress, and their acculturation level. At 6-month follow-ups, adolescents reported on their risk behaviors, including substance use and sexual activity. In general, adolescents' acculturation level was not related to their risk behaviors. More frequent conflict, more conflict about the adolescent, more adolescent involvement in the conflict, and poor conflict resolution were related to greater emotional distress. More conflict about the adolescent, mothers being more demanding/dominating during conflict, and more adolescent involvement in the conflict were related to greater risk behaviors. Adolescents' cognitions mediated the link between two dimensions of parental conflict, frequency and resolution, and emotional distress. Adolescents' emotional distress mediated the association between adolescent involvement in parental conflict and adolescents' risk behaviors. PMID- 12108768 TI - Fear of abandonment as a mediator of the relations between divorce stressors and mother-child relationship quality and children's adjustment problems. AB - This study examines whether fear of abandonment mediates the prospective relations between divorce stressors and mother-child relationship quality and adjustment problems of children of divorce. Participants were 216 children, ages 8-12, and their primary residential mothers. Children reported on divorce stressors and fear of abandonment; mothers and children reported on mother-child relationship quality and internalizing and externalizing problems. Structural equation models indicated that Time 1 fear of abandonment mediated the relation between Time 1 divorce stressors and Time 2 internalizing and externalizing problems. Time 1 fear of abandonment also mediated the relation between Time 1 mother-child relationship quality and Time 2 internalizing and externalizing problems. Implications of these results for understanding variability in children's postdivorce adjustment problems and interventions for divorced families are discussed. PMID- 12108769 TI - Deviant peer affiliations, crime and substance use: a fixed effects regression analysis. AB - The present study was designed to assess the influence of deviant peer affiliations on crime and substance use in adolescence/young adulthood. Data were used from a 21-year longitudinal study of health, development, and adjustment of a birth cohort of 1,265 New Zealand children. Annual assessments of deviant peer affiliations were obtained for the period from age 14-21 years, together with measures of psychosocial outcomes including, violent crime, property crime, alcohol abuse, cannabis abuse, and nicotine dependence. Affiliating with deviant peers was found to be significantly associated with each of these outcomes (p < .0001). Statistical control for confounding by both fixed and time dynamic factors reduced the strength of association between deviant peer affiliations and outcome measures. Nevertheless, deviant peer affiliations remained significantly associated (p < .0001) with all outcomes. For violent/property crime, cannabis and alcohol abuse there was significant evidence of age-related variation in the strength of association with deviant peer affiliations, with deviant peer affiliations having greater influence on younger participants (14-15 years) than older participants (20-21 years). These results suggest that deviant peer affiliations are associated with increased rates of a range of adjustment problems in adolescence/young adulthood with deviant peer affiliations being most influential at younger ages. PMID- 12108770 TI - Experimental validation of the DPM Monte Carlo code using minimally scattered electron beams in heterogeneous media. AB - A comprehensive set of measurements and calculations has been conducted to investigate the accuracy of the Dose Planning Method (DPM) Monte Carlo code for electron beam dose calculations in heterogeneous media. Measurements were made using 10 MeV and 50 MeV minimally scattered, uncollimated electron beams from a racetrack microtron. Source distributions for the Monte Carlo calculations were reconstructed from in-air ion chamber scans and then benchmarked against measurements in a homogeneous water phantom. The in-air spatial distributions were found to have FWHM of 4.7 cm and 1.3 cm, at 100 cm from the source, for the 10 MeV and 50 MeV beams respectively. Energy spectra for the electron beams were determined by simulating the components of the microtron treatment head using the code MCNP4B. Profile measurements were made using an ion chamber in a water phantom with slabs of lung or bone-equivalent materials submerged at various depths. DPM calculations are, on average, within 2% agreement with measurement for all geometries except for the 50 MeV incident on a 6 cm lung-equivalent slab. Measurements using approximately monoenergetic, 50 MeV, 'pencil-beam'-type electrons in heterogeneous media provide conditions for maximum electronic disequilibrium and hence present a stringent test of the code's electron transport physics; the agreement noted between calculation and measurement illustrates that the DPM code is capable of accurate dose calculation even under such conditions. PMID- 12108771 TI - Monte Carlo simulations of pinhole imaging accelerated by kernel-based forced detection. AB - Pinhole collimation can provide both higher sensitivity and resolution than parallel hole collimation when used to image small objects. When objects are placed close to the pinhole, small pinhole diameters combined with high magnification pinhole geometries yield ultra high resolution images. With Monte Carlo (MC) calculations it is possible to simulate accurately a wide range of features of pinhole imaging. The aim of the present work is to accelerate MC simulations of pinhole SPECT projections. To achieve speed-up, forced detection (FD), a commonly used acceleration technique, is replaced by a kernel-based forced detection (KFD) step. In KFD, instead of tracing individual photons from the source or last scatter position to the detector, a position dependent kernel (point spread function (PSF)) is projected on the detector. The PSFs for channel and knife edge pinhole apertures model the penetration effects through the aperture material. For simulations, the PSFs are pre-calculated and stored in tables. The speed-up and accuracy achieved by using KFD were validated by means of digital phantoms. MC simulations with FD and with KFD converge to almost identical images. However, KFD converges to an equal image noise level one to four orders of magnitude faster than FD, depending on the number of photons simulated. A simulator accelerated by KFD could serve as a practical tool to improve iterative image reconstruction. PMID- 12108772 TI - Intensity-modulated radiation therapy using only jaws and a mask: II. A simplified concept of relocatable single-bixel attenuators. AB - Intensity-modulated radiation therapy (IMRT) can be delivered inefficiently using the movable jaws of a linac. The efficiency can be much improved by delivering IMRT with the movable jaws together with a relocatable mask below the jaws (Webb 2002 Phys. Med. Biol. 47 257-75). This paper extends the modelling work by showing that a much simpler relocatable mask than previously conceptualized can lead to very similar improvements in monitor-unit efficiency and a decrease in the number of field components compared with the use of jaws only (JO). The new concept comprises a set of relocatable single-bixel attenuators (SBAs) which can be moved into the field components otherwise collimated by jaws only. Typically for a 15 x 15 bixel2 2D matrix of fluence with a peak value of Ip = 10 MUs (or equivalently ten stratified fluence levels) and using just four SBAs the MU efficiency is nearly three times that of the JO technique and the number of field components is reduced to about 0.6 the number required by the JO technique. These gains become greater by using more SBAs or for larger Ip values. Component reordering was achieved to minimize the total delivery time including intersegment deadtimes. Practicalities are discussed. PMID- 12108773 TI - Magnetization transfer imaging for polymer gel dosimetry. AB - Off-resonance RF pre-saturation was used to obtain contrast in MRI images of polymer gel dosimeters irradiated to doses up to 50 Gy. Two different polymer gel dosimeters composed of 2-hydroxyethyl-acrylate or methacrylic acid monomers mixed with N, N'-methylene-bisacrylamide (BIS), dispersed in an aqueous gelatin matrix were evaluated. Radiation-induced polymerization of the co-monomers generates a fast-relaxing insoluble polymer. Saturation of the polymer using off-resonance Gaussian RF pulses prior to a spin-echo readout with a short echo time leads to contrast that is dependent on the absorbed dose. This contrast is attributed to magnetization transfer (MT) between free water and the polymer, and direct saturation of water was found to be negligible under the prevailing experimental conditions. The usefulness of MT imaging was assessed by computing the dose resolution obtained with this technique. We found a low value of dose resolution over a wide range of doses could be obtained with a single experiment. This is an advantage over multiple spin echo (MSE) experiments using a single echo spacing where an optimal dose resolution is achieved over only very limited ranges of doses. The results suggest MT imaging protocols may be developed into a useful tool for polymer gel dosimetry. PMID- 12108774 TI - Automated planning volume definition in soft-tissue sarcoma adjuvant brachytherapy. AB - In current practice, the planning volume for adjuvant brachytherapy treatment for soft-tissue sarcoma is either not determined a priori (in this case, seed locations are selected based on isodose curves conforming to a visual estimate of the planning volume), or it is derived via a tedious manual process. In either case, the process is subjective and time consuming, and is highly dependent on the human planner. The focus of the work described herein involves the development of an automated contouring algorithm to outline the planning volume. Such an automatic procedure will save time and provide a consistent and objective method for determining planning volumes. In addition, a definitive representation of the planning volume will allow for sophisticated brachytherapy treatment planning approaches to be applied when designing treatment plans, so as to maximize local tumour control and minimize normal tissue complications. An automated tumour volume contouring algorithm is developed utilizing computational geometry and numerical interpolation techniques in conjunction with an artificial intelligence method. The target volume is defined to be the slab of tissue r cm perpendicularly away from the curvilinear plane defined by the mesh of catheters. We assume that if adjacent catheters are over 2r cm apart, the tissue between the two catheters is part of the tumour bed. Input data consist of the digitized coordinates of the catheter positions in each of several cross-sectional slices of the tumour bed, and the estimated distance r from the catheters to the tumour surface. Mathematically, one can view the planning volume as the volume enclosed within a minimal smoothly-connected surface which contains a set of circles, each circle centred at a given catheter position in a given cross-sectional slice. The algorithm performs local interpolation on consecutive triplets of circles. The effectiveness of the algorithm is evaluated based on its performance on a collection of soft-tissue sarcoma tumour beds within various anatomical structures. For each of 15 patient cases considered, the algorithm takes approximately 2 min to generate the planning volume. Although the tumour shapes are rather different, the algorithm consistently generates planning volumes that visually demonstrate smooth curves compactly encapsulating the circles. This general-purpose contouring algorithm works well whether the catheters are all close together, spread far apart in the plane or arranged in a convoluted way. The automatic contouring algorithm significantly reduces labour time and provides a consistent and objective method for determining planning volumes for soft tissue sarcoma. Further studies are needed to validate the significance of the resulting planning volumes in designing treatment plans and the role that sophisticated brachytherapy treatment planning optimization may have in producing good plans. PMID- 12108775 TI - Bio-acoustic thermal lensing and nonlinear propagation in focused ultrasound surgery using large focal spots: a parametric study. AB - It is well known that the acoustic properties of soft tissue have a dependence on tissue temperature. This is of particular interest in focused ultrasound surgery since the mechanism of action of focused ultrasound surgery is to kill targeted tissue by inducing localized heating by ultrasound absorption, and hence cautery of that tissue. However, the act of localized heating induces a change in the acoustic properties of the targeted tissue and tissue surrounding it. This phenomenon distorts the incoming acoustic wavefront, and has been termed the thermal lens effect for this reason. Furthermore, nonlinear effects in acoustic propagation become non-negligible at the ultrasound intensities required for therapeutic action. This paper examines the importance of the thermal lens effect and nonlinear tissue properties by simulating a variety of clinically applicable phased array transducer configurations that have not yet been appropriately analysed using a full three-dimensional nonlinear treatment of acoustic propagation. The significance of the thermal lens effect is characterized by comparing the simulation of coupled acoustic and thermal propagation with an uncoupled treatment; neglecting thermal lensing typically produces a movement of 1 to 2 mm in the predicted position of the focus towards the transducer. The results also show that the classical methods of acoustic propagation can produce grossly erroneous results under certain clinically relevant transducer configurations and that an acoustic field scan with a hydrophone may not accurately predict therapeutic effect. PMID- 12108777 TI - Modified ionic models of cardiac tissue for efficient large scale computations. AB - Recirculation of excitation, or re-entry, is one of the most important mechanisms of life-threatening cardiac arrhythmias and fibrillation. Modelling these phenomena requires large scale computations in two and three-dimensional slabs of cardiac tissue. Because of computational constraints, most of the studies use simplified (non-ionic) models of cardiac tissue, which are electrophysiologically less accurate than the detailed ionic models. In this paper, we propose a method to modify ionic models of cardiac tissue into an intermediate class of models, which are almost as efficient for computations as simplified models, and retain most of the properties of the original ionic models, such as the shape of the action potential, the restitution of action potential duration and of the conduction velocity, as well as unchanged description of most of the ionic currents. PMID- 12108776 TI - Spectral variation of the infrared absorption coefficient in pulsed photothermal profiling of biological samples. AB - Pulsed photothermal radiometry can be used for non-invasive depth profiling of optically scattering samples, including biological tissues such as human skin. Computational reconstruction of the laser-induced temperature profile from recorded radiometric signals is sensitive to the value of the tissue absorption coefficient in the infrared detection band (muIR). While assumed constant in reported reconstruction algorithms, muIR of human skin varies by two orders of magnitude in the commonly used 3-5 microm detection band. We analyse the problem of selecting the effective absorption coefficient value to be used with such algorithms. In a numerical simulation of photothermal profiling we demonstrate that results can be markedly impaired, unless the reconstruction algorithm is augmented by accounting for spectral variation muIR(lambda). Alternatively, narrowing the detection band to 4.5-5 microm reduces the spectral variation muIR(lambda) to a level that permits the use of the simpler, unaugmented algorithm. Implementation of the latter approach for depth profiling of port wine stain birthmarks in vivo is presented and discussed. PMID- 12108778 TI - In vivo metabolism and partitioning of 6-[18F]fluoro-L-meta-tyrosine in whole blood: a unified compartment model. AB - Physiological quantification of dynamic PET data requires the determination of an input function, preferably from plasma. A compartmental model relating a parent radiotracer, its radiolabelled metabolites and their exchange between plasma and erythrocytes is presented. This model allows for the time course of radioactivity measured in whole blood to be transformed into the time course of the radiotracer in plasma. The utility of this approach is illustrated with blood data collected on 30 human subjects injected with 6-[18F]fluoro-L-meta-tyrosine (FmT), a pre synaptic dopaminergic radiotracer. A three-compartment four-parameter model is shown to yield significantly better fits to the blood data than related lower and higher order models. This model is found to be robust to measurement noise, and yet sensitive to metabolic changes induced by pretreatment with carbidopa. For FmT, the between-subject variations are shown to be small enough to warrant the use of a population-based correction; tissue time-activity curves were simulated to verify that this correction does not significantly affect the precision and accuracy of the derived rate constants. The unified blood model can be adapted for radiotracers other than FmT as long as the blood partition ratio of the parent radiotracer differs from that of its metabolites and/or the rate at which they equilibrate between plasma and erythrocytes is different. PMID- 12108779 TI - Imaging of in vitro chicken leg using time-resolved near-infrared optical tomography. AB - Near-infrared optical imaging gains much attention because of its noninvasiveness and deep penetration depths into tissue. Here, we report near-infrared optical tomographic imaging of an in vitro chicken leg from time-resolved measurements. The in vitro chicken leg, dipped in a cylindrical container filled with diluted Intralipid-10% solution, was imaged with a multichannel time-resolved imaging system. A two-dimensional reconstruction algorithm based on a modified generalized pulse spectrum technique has been developed to reconstruct the images of both the absorption and reduced scattering coefficients simultaneously and quickly. The results demonstrate that a simultaneous reconstruction of absorption and reduced scattering coefficients from time-resolved measurement has a potential to reveal the changes in the optical properties associated with not only the physiological information but also the anatomical structure of the organ. PMID- 12108780 TI - Scavenging is no magic. PMID- 12108781 TI - Investigation of photon beam output factors for conformal radiation therapy- Monte Carlo simulations and measurements. AB - The purpose of this study was to investigate beam output factors (OFs) for conformal radiation therapy and to compare the OFs measured with different detectors with those simulated with Monte Carlo methods. Four different detectors (diode, diamond, pinpoint and ionization chamber) were used to measure photon beam OFs in a water phantom at a depth of 10 cm with a source-surface distance (SSD) of 100 cm. Square fields with widths ranging from 1 cm to 15 cm were observed; the OF for the different field sizes was normalized to that measured at a 5 cm x 5 cm field size at a depth of 10 cm. The BEAM/EGS4 program was used to simulate the exact geometry of a 6 MV photon beam generated by the linear accelerator, and the DOSXYZ-code was implemented to calculate the OFs for all field sizes. Two resolutions (0.1 cm and 0.5 cm voxel size) were chosen here. In addition, to model the detector four kinds of material, water, air, graphite or silicon, were placed in the corresponding voxels. Profiles and depth dose distributions resulting from the simulation show good agreement with the measurements. Deviations of less than 2% can be observed. The OF measured with different detectors in water vary by more than 35% for 1 cm x 1 cm fields. This result can also be found for the simulated OF with different voxel sizes and materials. For field sizes of at least 2 cm x 2 cm the deviations between all measurements and simulations are below 3%. This demonstrates that very small fields have a bad effect on dosimetric accuracy and precision. Finally, Monte Carlo methods can be significant in determining the OF for small fields. PMID- 12108782 TI - Measurement of skin dose variations produced by a silicon-based protective dressing in radiotherapy. AB - Variations in skin dose caused by a silicon-based burn dressing used in radiotherapy during treatment have been investigated. Measurement of these variations in skin dose has been achieved using thermoluminescent dosimeters (TLDs) and Gafchromic film. For a 6 MV x-ray beam results have shown that an approximately 0.4 mm thick silicon mesh dressing increases the average surface dose by approximately 12.5% to 14% of the maximum and average dose at 1 mm depth and by 4% to 6% of the maximum for field sizes ranging from 5 cm x 5 cm up to 40 cm x 40 cm at 100 cm source to surface distance (SSD). The radiation effective thickness of the silicon dressing was calculated to be 0.5 mm +/- 0.05 mm water equivalent. TLDs of various thicknesses provide point-dose assessment and Gafchromic film can provide a detailed two-dimensional dose map with a high spatial resolution. Results have shown that a large variation in skin dose is delivered under the dressing depending on the amount of material directly above it as defined by the silicon mesh outline. PMID- 12108783 TI - Modified ferrous ammonium sulfate benzoic acid xyelenol orange (MFBX) and thermoluminescent dosimeters--a comparative study. AB - Radiation dosimetry deals with the determination of absorbed dose to the medium exposed to ionizing radiation. Chemical dosimetry depends on oxidation or reduction of chemicals by ionizing radiation. A ferrous ammonium sulfate benzoic acid xyelenol orange (FBX) dosimeter based on this principle is being used as a clinical dosimeter at present. Certain modifications were carried out in the preparation and storage of the FBX dosimeter to increase its shelf life. The resulting dosimeter was called a modified FBX (MFBX) dosimeter and has been used in our department for the past few years. An extensive study of the dose, dose rate and energy response of the dosimeter was carried out and compared with a thermoluminescent (LiF7) dosimeter. The results obtained were found to be comparable to the thermoluminescent (LiF7) dosimeter. Hence it was concluded that the MFBX dosimeter could be used for phantom dosimetry, data collection and in vivo measurements. Easier preparation and availability of the reagents are added advantages of using MFBX as a clinical dosimeter in small radiotherapy departments. PMID- 12108784 TI - Expression and regulation of gp130 messenger ribonucleic acid in cultured immature rat Sertoli cells. AB - The expression of glycoprotein 130 (gp130) was studied in rat primary Sertoli cells by Northern blot analysis. Gp130 mRNA of 9.0 and 7.5 kb were detected in a variety of rat tissues including the testis. Gp130 mRNAs were detected in isolated, immature Sertoli cells. The levels of gp130 were significantly stimulated by the addition of interleukin-1-beta (IL-1-beta) or interleukin-6 (IL 6) but not by the addition of follicle-stimulating hormone (FSH). IL-6 activates intracellular signaling by binding with a receptor consisting of an 80-kDa ligand binding protein, IL-6 receptor (IL-6R), and a second 130-kDa protein, gp130. As previously reported, expression of IL-6R mRNA in rat Sertoli cells was stimulated not only by IL-1-beta and IL-6 but also by FSH. In contrast, gp130 mRNA expression was not stimulated by FSH in our analysis. These data suggest that gp130 expression may be regulated by more than 1 mechanism and that production of gp130 and IL-6R, the 2 components of the IL-6 receptor system, may be regulated, at least in part, by a different pathway. PMID- 12108785 TI - Effect of compound CDRI 84/35 and synthetic estrogen on the seminiferous epithelium of immature rat. AB - Compound CDRI 84/35 (a piperazine derivative--a potent antispermatogenic agent) has been shown to cause significant inhibition in testicular spermatogenesis without affecting Leydig cell and accessory sex organ function in adult rats. The present study was conducted to determine its effect on the germ cell population and Leydig cell morphology in immature rats (40-50 gm) administered CDRI 84/35 (100 mg/kg/day p.o.), synthetic estradiol benzoate (EB; 5 microg/rat/day) and vehicle at the age of 21 days. Animals were killed 24 h later following 7 and 14 days' treatments. Bouin's fixed testes were sectioned (at 5 microm) and stained with PAS-hematoxylin. Quantitative determination of Sertoli Cell-Germ Cell ratio was carried out in 150 round seminiferous tubules in each group of 5 rats. Results revealed a significant decrease in number of the spermatocytes (non pachytene and pachytene) and early (round) spermatids in step 1-8 of spermiogenesis without affecting Leydig cell morphology in rats administered CDRI 84/35 for 7 and 14 days as compared to corresponding controls. In contrast, the testes of rats injected with synthetic EB, caused a marked inhibition in these meiotic and post-meiotic germ cell types, as well as in the diameters of round seminiferous tubules, and Leydig cells nuclei (only in 14 days treatment), and testicular weight on autopsy days 8 and 15 as compared to CDRI 84/35-treated rats. While the number of pre-meiotic spermatogoniae was observed to be slightly decreased after only 14 days treatment in both CDRI 84/35 and EB treatment groups, the Sertoli cell number did not show any significant change as compared to controls. The present investigation confirms the antispermatogenic effect of compound CDRI 84/35 in immature rats similar to that reported in adult rats. Marked inhibition in pachytene spermatocytes and other testicular parameters following synthetic estrogen treatment might be due to its antiandrogenic action, contrasting with the non-hormonal profile of CDRI compound. PMID- 12108786 TI - Influence of hypertension on tetrahydropapaveroline-induced vasorelaxation in rat thoracic aorta. AB - Tetrahydropapaveroline (THP), a condensation product of ethanol-derived acetaldehyde, has been shown to elicit a vasorelaxant response in rat thoracic aorta. This study examined the influence of hypertension on the THP-induced vasorelaxant responsiveness. Ring segments of thoracic aorta were isolated from Wistar-Kyoto (WKY) and spontaneously hypertensive rats (SHR) and isometric tension development was measured with a force transducer. In aorta, with or without intact endothelium, the contractile responses to potassium chloride (0 120 mM) were comparable between the WKY and the SHR groups. Hypertension did not affect the vasoconstrictive response to norepinephrine (0-10 microM) in vessels with intact endothelium, whereas it attenuated the norepinephrine-induced response in vessels without endothelium. THP (0.1-100 microM) elicited endothelium-intact as well as -denuded vasorelaxation in aorta from both WKY and SHR groups. Interestingly, the THP-induced endothelium-dependent vasorelaxation was significantly enhanced, whereas the THP-induced endothelium-independent vasorelaxation was not affected by hypertension. These data indicate that the THP induced endothelium-dependent vasorelaxant response is altered by the hypertensive state. PMID- 12108787 TI - Plasma ghrelin levels in patients with short bowel syndrome. AB - Ghrelin is a novel peptide hormone which was identified as an endogenous growth hormone secretagogue. It is mainly secreted in the stomach, but important sites of its secretion are other parts of the gastrointestinal tract. Ghrelin is thought to be involved not only in regulation of growth hormone secretion but also in regulation of food intake and nutritional status. This study was aimed to investigate the changes in plasma ghrelin levels in patients with short bowel syndrome. Twenty-four patients with malnutrition due to short bowel syndrome and eleven healthy controls were included in the study. They underwent clinical examination and assessment of plasma or serum levels of ghrelin leptin, soluble leptin receptor, IGF-I, IGFBP-1 and IGFBP-3. Plasma ghrelin levels were decreased in patients with short bowel syndrome (p<0.01). Furthermore, decreased serum levels of IGF-I (p<0.01) and IGFBP-3 (p<0.001) were found in patients with short bowel syndrome. Other laboratory differences between both groups were not significant. No relationship between ghrelin and other determined variables was found. We conclude that plasma ghrelin levels are decreased in the group of patients with short bowel syndrome. It is probably because of a decrease in the tissue mass that is able to secrete ghrelin. PMID- 12108788 TI - Comparison of the secretory properties of four insulin-secreting cell lines. AB - The insulin-secretory responsiveness of four popular and widely used insulin secreting cells lines (RINm5F, HIT-T15, INS-1 and BRIN-BD11 cells) to a range of stimuli including glucose, amino acids, neurotransmitters, peptide hormones and sulphonylureas was studied. Differences were seen in the pattern of responsiveness of the cell lines to the various modulators of insulin release. While these studies revealed that INS-1 cells had the highest insulin content, only BRIN-BD11 cells exhibited a significant step-wise insulin secretory response to increasing glucose concentrations. BRIN-BD11 cells also showed pronounced insulin responses to leucine, KIC, L-arginine, L-alanine and palmitic acid. All the cell lines tested gave significant responses to the neurotransmitters carbachol and glibenclamide with increased insulin release. A comparison was made between the functional characteristics of the various cell lines with those of freshly isolated rat islets. This illustrated the general value of each cell line as a model for studies of insulin secretion. Electrofusion-derived BRIN-BD11 cells appeared to closely mimic the glucose sensitivity and overall secretory performance of normal rat islets. PMID- 12108789 TI - The impact of vitamin C on diabetes induced alterations at murine neuromuscular junction. AB - Physiological functions of skeletal muscle are compromised in diabetes. This may involve free radical mechanisms and may be reversed by antioxidants. We have studied effects of vitamin C on twitch tension, resting membrane potential (RMP) and miniature endplate potentials (MEPPs) frequencies in dorsiflexor muscle of diabetic murine. Forty mice were divided randomly into 2 groups (n = 20 each). One group served as control and the other was injected once with streptozotocin (STZ) solution (60 mg/kg, i.p.) to induce diabetes. The animals were then divided further into two subgroups (n = 10 each). Vitamin C (200 mg/kg, i.p.) was administered daily to one control and one diabetic group for three weeks prior to recording day. Experiments were conducted four weeks following diabetes induction. Isometric twitch tension (evoked directly by muscle stimulation and indirectly by nerve stimulation) was measured in urethane anesthetized (2 mg/g, i.p.) mice via a transducer connected to computer system. Utilizing intracellular recording method, resting membrane potential RMP and MEPPs frequencies were also measured. Compared to control, diabetic mice showed reduced twitch tension (4.2 +/- 0.5 g control versus 2.6 +/- 0.2 g diabetic) and demonstrated delayed half time of decay. Diabetic flexor muscle also displayed significant reduction in MEPPs frequencies with no changes in RMP. Vitamin C reversed tension reduction in diabetic mice (from 2.6 +/- 0.2 g to 3.9 +/- 0.3 g), impacted delayed half time of decay and increased MEPPs frequencies. Vitamin C improves diabetes-induced nerve and muscle dysfunction possibly via a free radical scavenging mechanism. PMID- 12108790 TI - Investigation of red blood cell carbonic anhydrase, glucose 6-phosphate dehydrogenase, hexokinase enzyme activities, and zinc concentration in patients with hyperthyroid diseases. AB - We have recently reported on the activities of glucose-6-phosphate dehydrogenase (G6PD), hexokinase (HK) and carbonic anhydrase I (CA-I) and II (CA-II) isoenzymes obtained from erythrocytes of healthy subjects and untreated patients with hyperthyroid diseases. Also, erythrocyte zinc concentrations were measured. Red blood cell (RBC) zinc (Zn) concentration was measured by atomic absorption spectrophotometry. Activities of carbonic anhydrase II and I isoenzymes were determined with CO2-hydratase activity method by using selective inactivation with bromopyruvate. G6PD and HK enzyme activities were measured spectrophotometrically via absorbance change (at 340 nm) in NADPH formed as a result of the reactions catalysed by these enzymes. In statistical analysis of all these parameters, activity of CA-I was 4388 +/- 207 (EU/gHb) and 2881 +/- 869 (EU/gHb) in healthy and untreated hyperthyroid subjects, respectively. The activity values for CA-II were 5391 +/- 257 (EU/gHb) and 4688 +/- 12.6 (EU/gHb) in healthy and untreated hyperthyroid subjects. Glucose 6-phosphate dehydrogenase (G6PD) activity was 10.19 +/- 1.87 (EU/gHb) in healthy group and 4.92 +/- 2.49 (EU/gHb) in patient group. While hexokinase enzyme activity was 1.575 +/- 0.898 in healthy subjects, it was 0.651 +/- 0.418 (EU/gHb) in the patient group. While erythrocyte zinc concentration in the healthy subjects was 49.32 +/- 23.5 (mg/gHb), this concentration for patients with uncontrolled hyperthyroid diseases was significantly decreased to 29.62 +/- 4.26 (mg/gHb). As a conclusion, CA-I isoenzyme, G6PD, hexokinase activities and erythrocyte zinc concentration had decreased in untreated patients carrying hyperthyroid diseases as compared to those of the healthy subjects. PMID- 12108791 TI - Rapid action of testosterone and diethylstilbestrol on enzymes of osmoregulation in a freshwater fish Oreochromis mossambicus. AB - The rapid action of sex steroids on enzymes of osmoregulation has not been studied in teleosts. In vivo administration of 0.1 microg/g body wt. of testosterone and diethylstilbestrol (DES) significantly enhanced branchial Na+-K+ ATPase, Ca2+ ATPase, and Na+, K+, and Ca2+ ions as early as 30 minutes in a freshwater teleost Oreochromis mossambicus. Treatment with 10(-6) M testosterone significantly stimulated Na+-K+ ATPase as early as 5 minutes in vitro. The same dose of DES also stimulated Na+-K+ ATPase activity within 10 minutes of incubation in a gill culture study. Administration of the transcriptional inhibitor, actinomycin D (0.1 microg/g body wt. in vivo and 10(-6) M in vitro), prior to hormone treatment, did not prevent the steroid-induced ATPase activity both in vivo and in vitro. It seems that stimulation of Ca2+ ion may be responsible for the hormonal effects to increase enzyme activity. A possible nongenomic mode of action for testosterone and DES is suggested in Oreochromis. PMID- 12108792 TI - Adrenomedullary hormonal and glycemic modulation following gonadotropin and sex hormone administration in the soft-shelled turtle Lissemys punctata punctata. AB - The aim of the current investigation was to ascertain whether gonadotropins and sex hormones can modulate adrenomedullary and glycemic functions in turtles. Exogenous FSH (3.0 microg/100 gm body wt daily for 10 days) stimulated adrenomedullary activity by increasing nuclear diameter and cytoplasmic degranulation, and elevating adrenal norepinephrine and epinephrine, and blood glycemic levels in turtles. LH (3.0 microg/100 gm body wt daily for 10 days) had no significant effect and the combined treatment of FSH and LH (3.0 microg each/100 gm body wt daily for 10 days) failed to show further adrenomedullary stimulation and hyperglycemia beyond that of FSH alone. Estrogen treatment in low dose (25 microg/100 gm body wt daily for 10 days) had no significant effect on adrenomedullary activity or glycemia, but at higher doses (50 microg or 100 microg/100 gm body wt daily for 10 days) showed dose-dependent adrenomedullary stimulation by inducing the same manifestations as those of FSH. Progesterone in low dose (25 microg/100 gm body wt daily for 10 days) had no perceptible effect, but in other doses (50 microg/100 microg per 100 gm body wt daily for 10 days), suppressed dose-dependent adrenomedullary activity by reversing the changes to those of estrogen. Simultaneous treatment with estrogen and progesterone (50 microg each/100 gm body wt daily for 10 days) failed to show further changes beyond that of estrogen alone. PMID- 12108793 TI - Prolactin stimulation in multiple sclerosis--an indicator of disease subtypes and activity? AB - Prolactin (PRL) belongs to the growth and lactogenic hormone family and has potent immunomodulating properties. Mild hyperprolactinemia has been found to enhance several autoimmune diseases and increased PRL plasma levels have been described in the experimental multiple sclerosis (MS) model while the PRL antagonist bromocriptine was able to suppress the disease. As studies of PRL serum levels in MS have led to conflicting results we investigated further the question of prolactin alterations in MS. We correlated PRL baseline values in a large sample of 132 MS patients with disease course and activity. Furthermore, inhibitory (bromocriptine) and stimulatory (metoclopramide) tests were performed in a subsample (n = 39) to gain functional information. We found no correlation of baseline values with disease course or activity. Nevertheless in the regression analysis of stimulatory test results, 14% of the variance was attributable to disease activity. In conclusion PRL does not seem to be relevant as an activity marker in the whole MS population. PMID- 12108794 TI - Diabetic ketoacidosis depletes plasma tryptophan. AB - Diabetic ketoacidosis (DKA) is a severe metabolic disturbance of insulin dependent diabetes mellitus (IDDM) which has a significant effect on amino acid metabolism. Amino acids serve as precursors for various neurotransmitters which are involved in affective disorders, and patients with IDDM are known to have an increased prevalence of affective disorders. We monitored the plasma concentrations of 23 amino acids in six adolescents prior to treatment of DKA and at 6, 24 and 120 hours after initiation of treatment. The well-known increase in the concentrations of the glucogenic amino acids and the decrease in the branched chain amino acids were observed in response to treatment of DKA. Low levels of tryptophan were found prior to treatment of DKA. Treatment increased the plasma tryptophan levels, but the mean concentration remained low throughout the sampling period. Only the glutamate-derived amino acids (glutamate, proline and glutamine) from the Krebs cycle pool were significantly affected by treatment. Glutamine declined initially, but recovered as the plasma pH normalized. Our results indicate that DKA causes a depletion of plasma tryptophan. This depletion may predispose some patients with IDDM to have affective disorders secondary to a neurotransmitter imbalance. PMID- 12108795 TI - The case of Matthew. PMID- 12108796 TI - Attempted abuse of concerta. PMID- 12108797 TI - Modified multiple-monitored electroconvulsive therapy. PMID- 12108798 TI - Priapism associated with trazodone in an adolescent with autism. PMID- 12108799 TI - The NIMH blueprint for change report: research priorities in child and adolescent mental health. AB - The National Institute of Mental Health established a special subgroup of its National Advisory Mental Health Council to review major research findings on child and adolescent psychiatric disorders over the past decade and to recommend research priorities for the next decade. This Workgroup on Child and Adolescent Mental Health Intervention Development and Deployment published its report, titled Blueprint for Change: Research on Child and Adolescent Mental Health, in August 2001, and several new research announcements reflecting these new directions have been issued since that time. This article summarizes the rationale for and background to the report, its major conclusions, and the reasons why interdisciplinary and translational approaches to research questions in child and adolescent mental health will help to maximize scientific advances. PMID- 12108800 TI - School-based treatment for anxious african-american adolescents: a controlled pilot study. AB - OBJECTIVE: To evaluate the feasibility and effectiveness of a school-based group cognitive-behavioral treatment (CBT) for anxiety disorders with African-American adolescents. METHOD: Twelve adolescents (mean age = 15.6 years) with anxiety disorders were randomly assigned to CBT (n = 6) or a group attention-support control condition (AS-Control; n = 6). Both groups met for 10 sessions in the same high school. Key treatment ingredients in CBT involved exposure, relaxation, social skills, and cognitive restructuring. Key ingredients in AS-Control involved therapist and peer support. At pre- and posttreatment, diagnostic interviews were conducted, and adolescents completed self-report measures of anxiety. RESULTS: At posttreatment and among those who attended more than one treatment session, 3/4 adolescents in CBT no longer met diagnostic criteria for their primary anxiety disorder, compared with 1/5 in AS-Control. Clinician ratings of impairment and self-report levels of overall anxiety were significantly lower at posttreatment in CBT compared with AS-Control. Teenagers in both groups reported lower levels of social anxiety from pre- to posttreatment. CONCLUSIONS: Findings support the feasibility of implementing a manual-based CBT in an urban school setting. Responder rates among African American adolescents were similar to those found in studies with white youths. PMID- 12108801 TI - Atomoxetine and methylphenidate treatment in children with ADHD: a prospective, randomized, open-label trial. AB - OBJECTIVE: To assess the comparability of atomoxetine, a new therapy for attention-deficit/hyperactivity disorder (ADHD) and methylphenidate. (Atomoxetine was originally called tomoxetine. The name was recently changed in order to avoid any potential confusion with tamoxifen that might lead to errors in dispensing drug.) METHOD: Children with ADHD were randomized to open-label atomoxetine or methylphenidate for 10 weeks. Response was assessed with the ADHD-IV Rating Scale. RESULTS: Two hundred twenty-eight patients were randomized (atomoxetine n = 184, methylphenidate n = 44). Both drugs were associated with marked improvement in inattentive and hyperactive-impulsive symptom clusters as assessed by parents and investigators. No statistically significant differences between treatment groups were observed on the primary outcome measure (investigator-rated ADHD-IV Rating Scale total score: atomoxetine baseline: 39.4 [8.5], endpoint: 20.0 [13.9]; methylphenidate baseline: 37.6 [9.7], endpoint: 19.8 (16.6); p = .66). Safety and tolerability were also similar between the 2 drugs. Discontinuations due to adverse events were 10/184 (5.4%) for atomoxetine and 5/44 (11.4%) for methylphenidate; p = .175. CONCLUSION: These data provide preliminary evidence that atomoxetine is associated with therapeutic effects comparable to those of methylphenidate. PMID- 12108802 TI - A study of compliance with FDA recommendations for pemoline (Cylert). AB - OBJECTIVE: To assess compliance with product labeling recommendations to use pemoline as second-line therapy for attention-deficit/hyperactivity disorder (ADHD) and to obtain baseline and biweekly liver enzyme tests. METHOD: Retrospective cohort study using administrative claims data to identify first line therapies and liver enzyme tests among pemoline users between January 1, 1998, and March 31, 2000. Prescriptions for first-line therapy were searched for 90 days prior to the first pemoline claim. Liver enzyme testing (baseline and follow-up) was compared between two groups (the prerecommendation cohort October 1,1998, to March 31, 1999, and the postrecommendation cohort October 1,1999, to March 31,2000). RESULTS: 1,308 patients received at least one pemoline prescription during the study period; 76% of patients < or = 20 years were male. ADHD was the claims-identified indication for 688 patients (52%). Despite the labeling recommendation for use as second-line therapy, only 237 ADHD patients (34%) received a first-line therapy prior to pemoline. Only 12% and 11% of the pre- and post-cohort patients, respectively, received baseline liver enzyme tests; 9% in the pre- and 12% in the post-cohort received at least one liver enzyme follow-up test. CONCLUSIONS: Compliance with product labeling was low for both recommendations. Understanding the reasons for this finding could help improve risk management strategies. PMID- 12108803 TI - Symptom factors in early-onset psychotic disorders. AB - OBJECTIVE: To examine the factor structure of symptom ratings in early-onset psychotic illnesses. METHOD: Subjects were drawn from a 2-year prospective study of early onset psychotic disorders. Principal components analysis with orthogonal (varimax) rotation was used to create factors from baseline ratings on the Schedule for Positive Symptoms, the Schedule for Negative Symptoms, and the Brief Psychiatric Rating Scale for Children. RESULTS: Youths with schizophrenia (n = 27), bipolar disorder (n = 22), and psychosis not otherwise specified (n = 20) were included. Four symptom factors were identified: negative symptoms, positive symptoms, behavioral problems, and dysphoria. Negative symptoms were predictive of the diagnosis of schizophrenia and treatment with antipsychotic medications. Neither behavior problems nor dysphoria were predictive of diagnosis. In subjects who completed follow-up assessments at year 1 (n = 49) and year 2 (n = 39), negative symptoms and behavioral problems predicted poorer functioning. CONCLUSIONS: The four factors are clinically relevant, with both treatment planning and prognostic implications. Negative symptoms best differentiated schizophrenia from the other disorders. Behavior problems and dysphoria were nonspecific problems that occurred in all three disorders, which likely leads to misdiagnosis in community settings. PMID- 12108804 TI - Association of attention-deficit/hyperactivity disorder symptoms with levels of cigarette smoking in a community sample of adolescents. AB - OBJECTIVE: Research on the association of attention-deficit/hyperactivity disorder (ADHD) with cigarette smoking has primarily occurred within samples of clinically referred youths. This paper reports the association of ADHD symptoms with smoking practices in a community sample of adolescents. METHOD: Confidential self-report surveys were completed by 1,066 tenth-grade students enrolled in five public high schools who were taking part in a longitudinal study of biobehavioral predictors of adolescent smoking adoption. A well-standardized measure of ADHD inattention and hyperactivity-impulsivity symptoms, as well as demographic and social risk factors, were examined in relation to three levels of cigarette smoking: (1) never having smoked, (2) ever having smoked, and (3) current smoking (having smoked a cigarette within the past 30 days). RESULTS: Regarding lifetime cigarette use, approximately 43% of students had ever smoked. Among those who had ever smoked, approximately 31% of students were current smokers. Ever having smoked was associated with family (odds ratio [OR] = 2.49, confidence interval [CI] = 1.85, 3.36) and peer smoking (OR = 4.05, CI = 3.07, 5.33) and clinically significant ADHD inattention symptoms (OR = 3.39, CI = 1.53, 7.54). Current smoking was also associated with peer smoking (OR = 2.99, CI = 1.72, 5.20) and clinically significant ADHD inattention symptoms (OR = 2.80, CI = 1.20, 6.56). CONCLUSION: Clinically significant ADHD symptoms should be taken into account when identifying adolescents at risk to smoke, since those with problematic inattention may be more likely to experiment with smoking and to become regular tobacco users. PMID- 12108805 TI - Cytogenetic abnormalities in attention-deficit/hyperactivity disorder. AB - OBJECTIVE: To systematically assess the prevalence of fragile X syndrome, velocardiofacial syndrome, and other cytogenetic abnormalities in a group of children with attention-defict/hyperactivity disorder (ADHD). METHOD: Blood samples were obtained from 100 children (64 boys) with combined type ADHD and normal intelligence and analyzed for the presence of fragile X mutation expansions, the 22q11.2 microdeletion associated with velocardiofacial syndrome, and cytogenetic abnormalities that would be detected with high resolution chromosomal banding. RESULTS: One girl with ADHD had a sex chromosome aneuploidy (47,XXX). One boy had a premutation-sized allele for fragile X; no subjects showed the full mutation. Testing for 22q11.2 microdeletion was negative for all subjects with ADHD screened. None of these differences exceeded those expected by chance. CONCLUSIONS: In the absence of clinical signs or positive family history, these relatively expensive laboratory assessments are not clinically indicated for children with ADHD and normal intelligence, and are not recommended as a component of other genetic investigations of this disorder. PMID- 12108806 TI - Altered cortical activity in children with attention-deficit/hyperactivity disorder during attentional load task. AB - OBJECTIVE: To evaluate whether cortical activity recorded during attentional load in children with ADHD is different compared with controls. METHOD: Quantitative electroencephalography (QEEG) was performed at open eyes and during performance of the Continuous Performance Task. RESULTS: Children with ADHD showed an altered pattern of QEEG activity, especially during the attentional load task, with increased slow cortical activity (mainly over the frontal areas) and decreased fast cortical activity. CONCLUSIONS: The findings indicate a different arousal level in children with ADHD, which could be due to a delay in functional cortical maturation. To evaluate the clinical importance of these findings, a longitudinal follow-up will be conducted. PMID- 12108807 TI - Discrimination of DSM-IV and latent class attention-deficit/hyperactivity disorder subtypes by educational and cognitive performance in a population-based sample of child and adolescent twins. AB - OBJECTIVE: Despite the general use of DSM-IV attention-deficit/hyperactivity disorder (ADHD) subtypes, there is controversy over the optimal phenotyping strategy for this disorder.This report contrasts two ADHD subtyping approaches on the prediction of cognitive function and educational achievement. METHOD: ADHD subtypes were determined using DSM-IV and latent class approaches for a population sample of 1,154 child and adolescent twins using parent report data. Twins completed cognitive and achievement testing and parents reported on school grades, special education placement, and history of being held back in school. RESULTS: The DSM-IV primarily inattentive and combined subtype ADHD groups showed significant deficits in cognitive and achievement testing, worse grades, and increased use of special education resources compared with the primarily hyperactive/impulsive subtype and no-ADHD groups. Clinically relevant and less severe latent class ADHD subtypes were also associated with deficits in cognitive and achievement testing, grades, and special education use. CONCLUSIONS: DSM-IV primarily inattentive and combined subtypes of ADHD have similar significant patterns of cognitive and academic dysfunction in the general population. Latent class-defined ADHD subtypes also have patterns of serious cognitive and achievement deficits. PMID- 12108808 TI - Emotional and behavioral problems among female twins: an evaluation of the equal environments assumption. AB - OBJECTIVE: To evaluate the equal environments assumption (EEA) of the twin method for mother-reported symptoms of child and adolescent emotional and behavioral problems. METHOD: Four emotional and behavioral problem scales (symptoms of separation anxiety disorder, attention-deficit/hyperactivity disorder, oppositional defiant disorder, and conduct disorder) and four environmental similarity measures (sharing friends, sharing classes, dressing alike, and perceived zygosity) were assessed by telephone interviews with the biological mothers of 1,948 female adolescent and young adult twin pairs. The effect of environmental similarity on the magnitude of the monozygotic and dizygotic twin correlations and on the parameter estimates from genetic model-fitting was examined for each symptom scale. RESULTS: The measures of environmental similarity were not strongly or consistently related to the similarity of twins for mother-reported emotional and behavioral problems. On average, controlling for environmental similarity did not substantially affect estimates of genetic and environmental influences. CONCLUSIONS: These results lend support for the validity of the EEA and suggest that estimates of genetic and environmental influences obtained from twin studies of mother-reported child and adolescent emotional and behavioral problems are not unduly biased by the greater environmental similarity of monozygotic than dizygotic twins. PMID- 12108809 TI - A developmental analysis of sociodemographic, family, and peer effects on adolescent illicit drug initiation. AB - OBJECTIVE: To examine the effects of sociodemographic, family, and peer predictors on the developmental patterns of illicit drug initiation from ages 12 to 21 years. METHOD: A gender-balanced, ethnically diverse urban sample of 808 children in Seattle was surveyed at age 10 in 1985 and followed prospectively to age 21 in 1996. Discrete-time survival analysis was used to assess the effects of sociodemographic, family, and peer factors on the risk of initiation. RESULTS: The risk for initiating illicit drug use increased steadily from ages 12 to 21. High family conflict, low family bonding, and high peers' antisocial activities predicted higher risk of initiation across this developmental period. The effect of family bonding began to decline after age 18, while the effect of peers' antisocial activities began to increase after age 15. Few gender and ethnic differences were found. CONCLUSIONS: Prevention programs need to include family and peer factors as important targets. Parents should create a warm and supportive family environment with appropriate supervision and control throughout adolescence. Association with antisocial peers should be reduced, especially in high school. Interventions addressing these family and peer factors should have beneficial effects across gender and ethnic groups. PMID- 12108810 TI - Smoking, getting drunk, and engaging in bulimic behaviors: in which order are the behaviors adopted? AB - OBJECTIVE: To assess the relation between beginning to binge-eat or purge, beginning to smoke, and getting drunk for the first time. METHOD: Prospective study of 11,358 girls and boys, 10 to 15 years of age, in an ongoing cohort study who completed questionnaires in 1997 and 1998. The outcome measures were beginning to engage in bulimic behaviors, beginning to smoke, and getting drunk for the first time between 1997 and 1998. RESULTS: During 1 year, 4.3% of girls and 3.8% of boys started smoking, 5.3% of girls and 4.8% of boys started getting drunk, and 2.4% of girls and 0.8% of boys started engaging in bulimic behaviors. Among the girls, weight concerns in 1997 were predictive of beginning to smoke (odds ratio [OR] = 2.2), get drunk (OR = 1.7), purge (OR = 3.8), and binge-eat (OR = 2.6). Adolescents who reported smoking in 1997 were more likely than nonsmokers to get drunk for the first time (girls: OR = 5.7; boys: OR = 7.1). The reverse association, getting drunk as a predictor of starting to smoke, was of lesser magnitude (OR = 2.3-2.6). CONCLUSIONS: The three unhealthy behaviors were associated prospectively with each other. The results suggest that prevention efforts should target weight concerns and multiple risk behaviors. PMID- 12108811 TI - Predictors of female criminality: findings from the Northern Finland 1966 birth cohort. AB - OBJECTIVE: Females still commit fewer criminal offenses than males, but the percentage of female offending has been increasing during the past few decades. Thus there is a need for original studies into the perinatal contribution to the etiology of female offending. METHOD: A large, prospectively collected birth cohort database of female members (N = 5,056) was available. Information on perinatal biological and psychosocial risks as well as data from the National Crime Registers up to 32 years of age were collected and analyzed by logistic regression and a chi2 automatic interaction detection (CHAID) analysis. RESULTS: The absence of the father during childhood was the strongest risk factor in predicting female criminality (odds ratio 2.5; 95% confidence interval 1.4-4.3). Furthermore, in the families in which the father was present, maternal smoking during pregnancy together with being born unwanted correlated with an increased prevalence of criminal offending significantly up to 7.2%. CONCLUSIONS: CHAID analysis proved to be a useful statistical method in predicting female adult criminality after preceding perinatal risks. It revealed that the risk factors were mainly familial, the paternal factor being the most important one in determining the probability of daughters committing criminal offenses. PMID- 12108812 TI - Gender differences in psychopathology, functional impairment, and familial risk factors among adjudicated delinquents. AB - OBJECTIVE: To test the hypotheses that female juvenile delinquents would have higher rates of psychological symptoms, DSM-IVpsychiatric and substance use disorders, functional impairment, and familial risk factors than male juvenile delinquents. METHOD: A stratified random sample of adjudicated delinquents (n = 513 males, n = 112 females) was drawn from San Diego County administrative databases. Of those sampled youths who could be located, 65.7% completed interviews. Psychological symptoms, DSM-lVdiagnoses, and familial risk factors were assessed between October 1997 and January 1999. RESULTS: Female delinquents scored higher on parent and self-report measures of psychological symptoms and had higher rates of DSM-IVmental disorders than did male delinquents. Girls also experienced greater incidences of physical, emotional, and sexual abuse; physical neglect; and family history of mental illness than their male counterparts. No gender differences were found on parental ratings of youth functional impairment, substance use disorders, comorbidity, or parental history of antisocial behavior. CONCLUSIONS: Findings indicated that female adjudicated delinquents have significantly higher rates of psychopathology, maltreatment history, and familial risk factors than males and suggest that the mental health needs of girls in juvenile justice deserve increased attention. PMID- 12108813 TI - Four-year follow-up of multisystemic therapy with substance-abusing and substance dependent juvenile offenders. AB - OBJECTIVE: Although several treatments for adolescent substance abuse have been identified as promising by reviewers and federal agencies, treatment effects extending beyond 12 months have not been demonstrated in randomized clinical trials. The primary purpose of this report was to examine the 4-year outcomes of an evidence-based treatment of substance-abusing juvenile offenders. METHOD: Eighty of 118 substance-abusing juvenile offenders participated in a follow-up 4 years after taking part in a randomized clinical trial comparing multisystemic therapy (MST) with usual community services. A multimethod (self-report, biological, and archival measures) assessment battery was used to measure the criminal behavior, illicit drug use, and psychiatric symptoms of the participating young adults. RESULTS: Analyses demonstrated significant long-term treatment effects for aggressive criminal activity (0.15 versus 0.57 convictions per year) but not for property crimes. Findings for illicit drug use were mixed, with biological measures indicating significantly higher rates of marijuana abstinence for MST participants (55% versus 28% of young adults). Long-term treatment effects were not observed for psychiatric symptoms. CONCLUSIONS: Findings provide some support for the long-term effectiveness of an evidenced based family-oriented treatment of substance-abusing juvenile offenders. The clinical, research, and policy implications of these findings are noted. PMID- 12108814 TI - Posttraumatic stress disorder among adolescent earthquake victims in Taiwan. AB - OBJECTIVE: To assess the exposure experience and prevalence of posttraumatic stress disorder (PTSD) among adolescent victims in the worst-affected region (Chungliao) near the epicenter of a severe earthquake (7.3 on the Richter scale) that occurred on September 21, 1999, in Taiwan. METHOD: The experience of exposure to the earthquake and subjective symptoms of junior high school students aged 12 to 14 who remained in the area were assessed with self-rated questionnaires. Psychiatrists made independent diagnoses for PTSD by using the Children's Interview for Psychiatric Syndromes. RESULTS: Six weeks after the earthquake, 21.7% of 323 students demonstrated PTSD. Those with PTSD showed significantly more psychiatric symptoms than did those without PTSD. Being physically injured and experiencing the death of a close family member with whom they had lived were the 2 major risk factors for PTSD. CONCLUSIONS: This study demonstrates that PTSD among adolescent victims of a severe earthquake in Taiwan is not as high as that reported in other studies. Methodological differences in the investigations are discussed, along with differences in symptom manifestations. However, long-term follow-up of these victims is recommended to prevent the development of other psychiatric complications. PMID- 12108815 TI - Genetics of childhood disorders: XL. Stem cell research, part 4: neural horticulture. PMID- 12108816 TI - Depression, anxiety, and the cardiovascular system: the cardiologist's perspective. AB - Up to one fifth of patients with cardiovascular disease, including those who have experienced a myocardial infarction, may have concomitant major depression. Studies have suggested that the relative risk of major depression with cardiovascular disease ranges from 1.5 to 4.5. Further information is required to establish a dose-response relationship between depression and coronary artery disease (CAD); however, such a relationship has been shown between anxiety and CAD. Development of a conceptual model of the pathophysiologic actions of stress in CAD will assist in the understanding of this relationship. In patients with angiographic evidence of CAD, the presence of major depressive disorder was the best single predictor of cardiac events during the 12 months following diagnosis. Significantly, 6-month cumulative mortality following diagnosis of myocardial infarction has been shown to be higher in depressed patients than in nondepressed patients. A decrease in heart rate variability may mediate the deleterious effect of depression on post-myocardial infarction prognosis. Other factors such as mental stress and altered platelet function may also predispose depressed patients to a heightened risk of cardiac events. With an increased understanding of the relationship between depression and heightened risk of cardiovascular mortality, it is necessary to assess current overall treatment for cardiac patients. PMID- 12108817 TI - Depression, anxiety, and the cardiovascular system: the psychiatrist's perspective. AB - It is becoming clear that the comorbidity of depression and cardiovascular disease does not occur by chance but rather is an inevitable consequence of the relationship between the conditions. Depression in patients with cardiovascular disease is a significant risk factor for developing symptomatic and fatal ischemic heart disease. Moreover, depressed patients have a higher than expected rate of sudden cardiovascular death. Therefore, appropriate treatment of patients with depression and cardiovascular disease cannot be restricted to considerations of either depression or cardiovascular disease in isolation. The tricyclic antidepressants (TCAs) have various effects on the cardiovascular system, including Type IA antiarrhythmic activity that has been associated with an increased risk of mortality in post-myocardial infarction patients. The selective serotonin reuptake inhibitors (SSRIs) are not associated with adverse cardiac effects. The SSRI paroxetine was compared with a therapeutic level of the TCA nortriptyline in a randomized, controlled study and demonstrated a benign cardiovascular profile, while the TCA induced a significantly higher rate of serious adverse cardiovascular events. On the basis of this favorable cardiovascular profile, the SSRIs should therefore be the preferred choice for the treatment of most patients with comorbid depression and cardiovascular disease. Investigation of putative pathophysiologic mechanisms linking depression and cardiovascular mortality, such as the role of platelet activation, will form the basis for further investigation of antidepressant treatments in order to establish if the antidepressants have a beneficial effect on the prognosis of cardiovascular diseases. PMID- 12108818 TI - Consensus statement on depression, anxiety, and cardiovascular disease. PMID- 12108819 TI - Depression, anxiety, and the gastrointestinal system. AB - Functional disorders of the digestive system, such as irritable bowel syndrome, are often associated with affective disorders, such as depression, anxiety, panic, and posttraumatic stress disorder (PTSD). Some of these associations are observed not only in clinical populations, but also in population-based samples, suggesting a relationship with pathophysiologic mechanisms underlying both gastrointestinal (GI) dysfunction and certain affective disorders. Sustained and acute life-threatening stressors play an important role in the onset and modulation of GI symptoms as well as in the development of affective disorders and PTSD. A neurobiological model is proposed that attempts to explain the development of visceral hypersensitivity, the neuroendocrine and autonomic dysfunction characteristic of functional GI disorders, as well as the overlap with affective disorders. PMID- 12108820 TI - Irritable bowel syndrome, anxiety, and depression: what are the links? AB - Irritable bowel syndrome (IBS) is a common and potentially disabling functional gastrointestinal disorder characterized by abdominal pain and altered bowel patterns. A significant amount of clinical and research data suggest the importance of the brain-gut interaction in IBS. This review examines the observed high prevalence of psychiatric disorders in patients with IBS. The published literature indicates that fewer than half of individuals with IBS seek treatment for it. Of those who do, 50% to 90% have psychiatric disorders, including panic disorder, generalized anxiety disorder, social phobia, posttraumatic stress disorder, and major depression, while those who do not seek treatment tend to be psychologically normal. Both physiologic and psychosocial variables appear to play important roles in the development and maintenance of IBS. Recent information suggests that the association of IBS and psychiatric disorders may be more fundamental than was previously believed. A brain-gut model for IBS is presented, and the role of traumatic stress and corticotropin-releasing factor as modulators of the brain-gut loop is discussed. Finally, the rationale for the use of psychotropic agents in the treatment of IBS with or without psychiatric symptoms is presented. PMID- 12108821 TI - The burden of depression and anxiety in general medicine. AB - Psychological illness is responsible for considerable disability worldwide. The World Health Organization Global Burden of Disease Survey estimates that by the year 2020, major depression will be second only to ischemic heart disease in the amount of disability experienced by sufferers. Although different measures of disability have been used in different studies, they have consistently demonstrated that individuals with depression and anxiety disorders experience impaired physical and role functioning, more days in bed due to illness, more work days lost, increased impairment at work, and high use of health services. The disability caused by depression and anxiety is just as great as that caused by other common medical conditions, such as hypertension, diabetes, and arthritis. Comorbidity of depression with anxiety or medical illness further increases the disability experienced by sufferers. Recognition and treatment, however, relieve the burden imposed by untreated depression on the individual, society, and health services. PMID- 12108822 TI - Consensus statement on depression, anxiety, and functional gastrointestinal disorders. PMID- 12108823 TI - Depression and anxiety in oncology: the oncologist's perspective. AB - Depression and anxiety frequently occur in oncology patients and have a significant impact on patient quality of life, health care utilization, and even disease outcome. Depression and anxiety are eminently treatable, and therefore psychiatric assessment and appropriate intervention should form an integral component of management strategy in patients with cancer. It is essential that patients are recognized at an early stage, so that resources can be targeted effectively at those most at risk of developing psychiatric morbidity. Evaluation techniques that can identify signs or symptoms of depression and anxiety and can be incorporated into the program of a busy oncology clinic or in the primary care setting are therefore needed. Diagnosis of depression and anxiety may be facilitated by using primary screening tools, such as the Hospital Anxiety and Depression Scale questionnaire, and by considering factors such as family psychiatric history, levels of family support, and degrees of pain suffered by the patient. In this article, the issues surrounding diagnosis of depression and anxiety in cancer patients and the benefits of early intervention are considered from the point of view of the oncologist. PMID- 12108824 TI - Depression and anxiety in oncology: the psychiatrist's perspective. AB - Depression and anxiety are frequently undiagnosed and untreated in cancer patients, resulting in a significant negative impact on quality of life and disease outcome. Furthermore, although effective therapies for depression and anxiety are available, there have been few clinical trials of pharmacotherapy in cancer patients. This article explores how the diagnosis of depression and anxiety can be improved in cancer patients and reviews current treatment options in the oncology setting. The Hospital Anxiety and Depression Scale, a simple, patient-administered questionnaire, is effective in screening for psychiatric symptoms at an early stage of cancer treatment, particularly if vulnerable individuals can be identified and targeted. Selective serotonin reuptake inhibitors are currently first-choice therapy for depression, and many of the drugs in the class are effective in anxiety disorders. Further studies of these agents in the oncology setting are warranted. PMID- 12108825 TI - Consensus statement on depression, anxiety, and oncology. PMID- 12108826 TI - Elements of a genetics counseling service. AB - Providing genetic information and counseling can be complex and time consuming. A Genetic Counseling Service consists of individuals that are certified by the American Board of Medical Genetics and others. These include Masters Degree trained genetic counselors, physicians that have completed special training in clinical genetics, nurses with special skills in genetics, and support staff. A team approach is required in order to provide patients comprehensive genetic counseling services. PMID- 12108827 TI - A general approach to genetic counseling. AB - Genetic counseling can best be performed if a systematic approach is taken. The geneticist or genetic counselor has the difficult task of conveying complex information to patients in an understandable form. At the same time, this must be done in a nondirective manner, affording the patient the right to make his or her own decisions. Obtaining accurate family and medical history information is crucial to the genetic counseling process. Given the nature of genetic information, multiple sessions may be necessary to ensure that the patient understands the risks involved in his or her particular situation. PMID- 12108828 TI - Variables that underlie cost efficacy of prenatal screening. AB - As genetic research and technology continues to expand, carrier testing for an increasing number of single gene disorders is becoming available. Tay-Sachs disease and cystic fibrosis are two common recessive conditions with large-scale health implications. Tay-Sachs disease was the first genetic disorder for which community-based screening efforts were utilized and has provided a foundation for the development of other screening programs. Cystic fibrosis testing, on the other hand, has additional complexities and the implementation of population based screening has been under debate. The many issues (technical, educational, social, psychological and economical) which must be considered as preconceptional and prenatal genetic screening is incorporated into clinical practice are discussed here in the context of Tay-Sachs disease and cystic fibrosis. PMID- 12108829 TI - Tay-Sachs disease screening and counseling families at risk for metabolic disease. AB - Carrier testing for Tay-Sachs disease should be offered to couples when at least one individual is of Ashkenazi Jewish (carrier frequency 1/30), Pennsylvania Dutch, Southern Louisiana Cajun, or Eastern Quebec French Canadian descent. Ideally, testing is done prior to conception. For Ashkenazi Jews, in whom DNA testing identifies 99.9% of carriers, DNA testing is the preferred method to ascertain carriers [14]. For non-Jewish individuals seeking carrier testing, enzyme assay should be done initially and positive or indeterminate results should be confirmed by DNA mutation analysis. If only one partner is descended from a high-risk group, that person should be tested first; only if he/ she is a carrier should the other partner be tested. If the couple is pregnant at the time carrier testing is requested, both partners should have enzyme testing (leukocyte assay for the pregnant woman and serum assay for the father) and DNA testing sent concomitantly to expedite counseling and action. Carriers are individuals with a disease causing DNA mutation or carrier range enzyme analysis results on both serum and leukocytes with no detectable mutation and no pseudodeficiency alleles. Noncarriers are individuals with normal enzyme results or carrier range enzyme results and a pseudodeficiency allele on DNA mutation analysis. If both partners are found to be carriers they should be counseled of a 25% risk of having an affected child with each pregnancy. Options to modify this risk include prenatal diagnosis by amniocentesis or chorionic villus sampling, egg or sperm donation, preimplantation diagnosis or adoption. PMID- 12108830 TI - Canavan disease prenatal diagnosis and genetic counseling. AB - Canavan disease is a severe leukodystrophy more common among Ashkenazi Jews. The enzyme defect, apartoacylase, has been identified, and the gene cloned. Only two mutations account for over 98% of all Jewish alleles with Canavan disease. The carrier frequency among healthy Jews is 1:37-58. Carrier detection and prenatal diagnosis can be accurately carried out using molecular analysis. When mutations are unknown, analysis of amniotic fluid for NAA using stable isotope dilution technique can be used for prenatal diagnosis. PMID- 12108831 TI - Screening for thalassemia: a model of success. AB - Programs of prospective carrier screening and genetic counseling for beta thalassemia among couples planning marriage, preconception, or during early pregnancy are ongoing in several at-risk populations in the Mediterranean area, including Greeks, Greek Cypriots and Continental Italians. Carrier detection is carried out by haematological analysis followed by mutation detection by DNA analysis. Once carrier couples are identified, prenatal diagnosis is accomplished by mutation analysis on PCR amplified DNA from chorionic villi. These programs have been very effective, due to education programs and subsequent acceptance of screening. Future prospects include automation of the process of mutation detection by microchips analysis, introduction of preconception and preimplantation diagnosis and hopefully fetal diagnosis by analysis of fetal cells in maternal circulation. PMID- 12108832 TI - Genetic screening for cystic fibrosis. AB - The importance of the recent recommendations that address cystic fibrosis carrier screening cannot be overemphasized. For the first time, a systematic approach to offering or making screening available to all pregnant women in the hopes of providing refined risk estimates for a genetic disease has been established. Caucasian of European or Ashkenazi-Jewish descent should be offered screening. Within the proposed guidelines are ethnic-specific carrier frequencies (1/29) used to establish who should be offered testing and to whom testing should be made available. Recent recommendations have made clear that in a pan-ethnic population a frequency of 1/1,000 is required for inclusion into the cystic fibrosis mutation panel. A general framework for screening during pregnancy has been established (either concurrent or sequential). It will be interesting to watch as the fruits of the human genome project are inspected and applied to everyday clinical practice. No doubt the cost of screening will be reduced through advances in technology. The combined efforts of NIH, ACOG, and ACMG have provided the first set of comprehensive standards for screening of recessive diseases. How time changes these guidelines deserves following. PMID- 12108833 TI - Counseling the at risk patient in the BRCA1 and BRCA2 Era. AB - The 5-year experience since the identification of the BRCA1 and BRCA2 genes has shown that genetic evaluation and testing can increase understanding of cancer risks for individuals with a personal or family history of early onset breast cancer and ovarian cancer. However, testing needs to be undertaken in a clinical setting where pretest counseling, including likelihood of identifying a mutation and risks and benefits of the process are provided. Identifying women who carry mutations in either BRCA1 or BRCA2 has implications for prevention, screening and treatment of these cancers. PMID- 12108834 TI - Fragile X and other trinucleotide repeat diseases. AB - Hereditary unstable DNA is composed of strings of trinucleotide repeats, in which three nucleotides are repeated over and over (ie CAGCAGCAGCAG). These repeats are found in several sites within genes; depending on their location, the number of triplet repeats in a string can change as it is passed on to offspring. When the number of repeats increases to a critical size, it can have a variety of affects on gene function. The repeats may cause a loss in gene function (as in Fragile X) or may result in the gain of a new, abnormal protein and thus a new function (as in myotonic dystrophy and Huntington disease). Although a variety of trinucleotide repeat diseases have been reported and merit consideration, this discussion will focus primarily on Fragile X syndrome, myotonic dystrophy, and Huntington disease. PMID- 12108835 TI - A numerical study of blood flow in coronary artery bypass graft side-to-side anastomoses. AB - PURPOSE: When sequential grafts are used in multivessel coronary artery bypass grafting, the graft first supplies blood to one or more coronary arteries via a side-to-side anastomosis. We studied hemodynamics in idealized models of "parallel" and "diamond" side-to-side anastomoses, identifying features that might promote restenosis. METHODS: Blood flow was computed in three representative anastomosis configurations: parallel side-to-side, diamond side-to side, and end-to-side. We compared configurations and the effect of host-graft diameter ratio. RESULTS: Hemodynamic patterns depended strongly on anastomosis geometry and graft/host diameter ratio. In the distal graft, the diamond configuration had large areas of low wall shear stress (WSS) and high spatial WSS gradients. In the proximal graft the unfavorable WSS patterns were comparable for all models, while the distal portion of the host artery the diamond model was best. Models with smaller host arteries had smaller regions of low WSS. CONCLUSIONS: The parallel configuration was preferred over the diamond for maintaining graft patency, while the diamond configuration appeared best for maintaining host artery patency. Since graft patency is critical, parallel configurations seem hemodynamically advantageous. Larger graft/host ratios have better hemodynamic performance than smaller ones. PMID- 12108836 TI - Model-based assessment of cardiovascular health from noninvasive measurements. AB - Cardiovascular health is currently assessed through a variety of hemodynamic parameters, many of which can only be determined by invasive measurement often requiring hospitalization. A noninvasive method of evaluating several of these parameters such as systemic vascular resistance (SVR), maximum left ventricular elasticity (E(LV)), end diastolic volume (VED), and cardiac output, is presented. The method has three elements: (1) a distributed model of the human cardiovascular system (Ozawa et aL, Ann. Biomed. Eng. 29:284-297, 2001) to generate a solution library that spans the anticipated range of parameter values, (2) a method for establishing the multidimensional relationship between features computed from the arterial blood pressure and/or flow traces (e.g., mean arterial pressure, pulse amplitude, mean flow velocity) and the critical hemodynamic parameters, and (3) a parameter estimation method that yields the best fit between measured and computed data. Sensitivity analyses were used to determine the critical parameters, and the influence of fixed model parameters. Using computer-generated brachial pressure and velocity profiles (which can be measured noninvasively), the error associated with this method was found to be less than 3% for SVR, and less than 10% for E(LV) and V(ED). Simulations were also performed to test the ability of the approach to predict changes in SVR and E(LV) from an initial base line state. PMID- 12108837 TI - Mechanical properties of dilated human ascending aorta. AB - Dilation of the ascending aorta, associated with Marfan Syndrome, bicuspid aortic valve, or advanced age, may lead to aortic dissection and rupture. Mathematical models can be used to assess the relative importance of increased wall stresses and decreased strength in these mechanical failures. To obtain needed inputs for such models, mechanical properties of dilated human ascending aorta were measured in vitro. Specimens for opening angle, biaxial elastic, and uniaxial circumferential strength tests were cut from excised tissue obtained from 54 patients (age 18-81 years) undergoing elective aortic graft replacement surgery. Opening angle was significantly greater in patients older than 50 years (262 degrees + 76 degrees, n = 21) compared to younger patients (202 degrees +/- 70 degrees, n = 13). All biaxial elastic specimens (n = 40) exhibited nonlinear stress-strain behavior. Rapid increases in circumferential and axial stresses occurred at lower strains in the older patient group than in the younger. Mean strength was significantly lower in older patients (1.35 +/- 0.37 MPa, n= 14) than younger (2.04 +/- 0.46 MPa, n = 11, age <50 years). These changes in mechanical properties suggest that age may influence the risk of aortic dissection or rupture of dilated ascending aorta. PMID- 12108838 TI - Approach to quantify the mechanical behavior of the intact embryonic chick heart. AB - The heart undergoes remarkable changes during embryonic development due to genetic programing and epigenetic influences such as mechanical loads. An important goal is to develop mathematical models that describe and predict the influence of mechanics on cardiac development, yet the data needed to develop such models remain scant. In particular, prior data from embryonic hearts have come from one-dimensional tests, which reveal well the general characteristic behaviors but are not sufficient for quantifying the complex material behavior exhibited by most soft tissues. We developed a computer-controlled system for quantifying in vitro the multiaxial, regionally dependent mechanical response of the intact embryonic chick heart to controlled distension pressures; such tests have not been accomplished heretofore but promise to contribute to our understanding for they better mimic the native loading conditions and geometry. Calibration of the device indicated that distending the hearts using low flow rates avoids significant viscous losses and thereby allows pressure to be measured directly over small ranges (0-2 mmHg) with good resolution (0.01 mm Hg). Likewise, an on-line video system allows two-dimensional strains to be measured regionally by tracking the motions of triplets of closely spaced (100 microm) microspheres to reasonable resolution (2.5 microm/pixel). Illustrative data from 18 hearts show the utility of the new system, confirm previous findings with regard to the strong viscoelastic response of the stage 18 embryonic chick heart, and provide guidance for the design of future experiments. PMID- 12108840 TI - Blood pressure measurement in noise intensive environments using adaptive interference cancellation. AB - The traditional auscultatory technique and current methodologies for measuring human blood pressure are limited when used in situations where extreme vibration or acoustic noise is present. In this study, human subjects were used to establish the effectiveness of a novel adaptive blood pressure monitoring (ABPM) system in determining systolic-diastolic pressure in vibration and noise intense environments. To remove the effects of noise and vibration from the audible Korotkoff sounds, the proposed ABPM system employs two acoustic sensors in the pressure cuff. The primary acoustic sensor is placed on the brachial artery to record the Korotkoff sounds while the secondary acoustic sensor is placed away from the artery to record background noise and vibrations. The signals from the two acoustic sensors are provided to the input of an adaptive interference canceller to remove the noise effects in the signal of the primary acoustic sensor. In two phases of clinical testing, the ABPM system was first employed in a noiseless environment and then near and onboard search and rescue helicopters. The results from both phases deliver a successful demonstration of the ABPM system's capabilities to provide blood pressure estimates in noisy environments where the conventional auscultatory and other techniques have limited use. PMID- 12108839 TI - Endothelial cell dynamics under pulsating flows: significance of high versus low shear stress slew rates (d(tau)/dt). AB - Shear stress modulates endothelial cell (EC) remodeling via realignment and elongation. We provide the first evidence that the upstroke slopes of pulsatile flow, defined as shear stress slew rates (positive d(tau)/dt), affect significantly the rates at which ECs remodel. We designed a novel flow system to isolate various shear stress slew rates by precisely controlling the frequency, amplitude, and time-averaged shear stress (tau(ave)) of pulsatile flow. Bovine aortic endothelial cell (BAEC) monolayers were exposed to three conditions: (1) pulsatile flow (1 Hz) at high slew rate (293 dyn/cm2 s), (2) pulsatile flow (1 Hz) at low slew rate (71 dyn/cm2s), and (3) steady laminar flow at d(tau)/dt = 0. All of the three conditions were operated at tau(ave) = 50 dyn/cm2. BAEC elongation and alignment were measured over 17 h. We were able to demonstrate the effects of shear stress slew rates ((tau)/dt) on EC remodeling at a fixed spatial shear stress gradient (d(tau)/dx). We found that pulsatile flow significantly increased the rates at which EC elongated and realigned, compared to steady flow at d(tau)/dt = 0. Furthermore, EC remodeling was faster in response to high than to low slew rates at a given tau(ave). PMID- 12108841 TI - Quantification of lung microvascular injury with ultrasound. AB - Quantification of water and solute exchange rates across the lung microvascular barrier (LMB) may be an important early-warning indicator of pulmonary microvascular diseases such as acute respiratory distress syndrome. Our objective was to determine the degree to which osmotic water movement across the LMB induced by injection of hypertonic solutions of NaCl and glucose could be detected downstream from the lung with a specialized ultrasonic velocity (USV) transducer manufactured by Transonic Systems. We hypothesized that mathematical modeling of the osmotic transients (OT) would yield estimates of osmotic exchange parameters that were sensitive to microvascular injury. Two groups of six dogs were studied under baseline conditions and after injury with high dose (HD) or low dose (LD) oleic acid. Osmotic conductances (sigmaK(1), sigmaK(2)), and volumes (V(1), V(2)) of two extravascular spaces were estimated by fitting the mathematical model to the OT data. HD results (mean +/- standard error) indicated a significant decrease (by paired t test) in sigmaK(1) from 1.59 +/- 0.09 to 1.04 +/- 0.015 [ml h(-1) (mosm/l)(-1) g(-1) WLW)], an increase in sigmaK(2) from 0.20 +/- 0.08 to 0.32 +/- 0.12 [ml h(-1) (mosm/l)(-1) g(-1)], and a significant increase in V2 from 23.26 +/- 2.51 to 78.0 +/- 15.23 (ml) for NaCl injections. LD V2 estimated from NaCl increased significantly from 21.57 +/- 2.15 to 37.59 +/- 2.36 (ml), sigmaK2 increased from 0.09 +/- 0.03 to 0.17 +/- 0.04 and no significant change in sigmaK(1) was found. Baseline glucose sigmaK(1), sigmaK(2), and V1 in the LD series were 2.08 +/- 0.18, 0.64 +/- 0.19 [ml h(-1) (mosm/l)(-1) g(-1)] and 13.08 +/- 1.89 (ml), respectively, and did not change significantly with injury. We conclude that OT data measured by USV is a sensitive and informative indicator of LMB osmotic properties, and may be useful for quantification of LMB permeability changes due to acute injury. We further conclude that V(1) represents microvascular endothelial volume and V(2) is an estimate of interstitial volume. PMID- 12108842 TI - Quantifying fractal dynamics of human respiration: age and gender effects. AB - We sought to quantify the fractal scaling properties of human respiratory dynamics and determine whether they are altered with healthy aging and gender. Continuous respiratory datasets (obtained by inductive plethysmography) were collected from 40 healthy adults (10 young men, 10 young women, 10 elderly men, and 10 elderly women) during 120 min of spontaneous breathing. The interbreath interval (IBI) time series were extracted by a new algorithm and fractal scaling exponents that quantify power-law correlations were computed using detrended fluctuation analysis. Under supine, resting, and spontaneous breathing conditions, both healthy young and elderly subjects had scaling exponents for the IBI time series that indicate long-range (fractal) correlations across multiple time scales. Furthermore, the scaling exponents (mean +/- SD) for the IBI time series were significantly (p < 0.03) lower (indicating decreased correlations) in the healthy elderly male (0.60 +/- 0.08) compared to the young male (0.68 +/- 0.07), young female (0.70 +/- 0.07), and elderly female (0.67 +/- 0.06) subjects. These results provide evidence for fractal organization in physiologic human breathing cycle dynamics, and for their degradation in elderly men. These findings may have implications for modeling integrated respiratory control mechanisms, quantifying their changes in aging or disease, and assessing the outcome of interventions aimed toward restoring normal physiologic respiratory dynamics. PMID- 12108843 TI - The pathway of bone fluid flow as defined by in vivo intramedullary pressure and streaming potential measurements. AB - The pathway for intracortical fluid flow response to a step-load was identified in vivo using intramedullary pressure (ImP) and streaming potential (SP) measurements, and allowed the development of a load-induced flow mechanism which considers mechanotransduction and mechanoelectrotransduction phenomena. An avian model was used for monitoring, simultaneously, ImP and SP under axial loading which generated peak strains of approximately 600 microstrain (microepsilon). ImP response to step-load decayed more quickly than SP relaxation, in which multiple time constants were observed during the relaxations. While the initial relaxation of SP showed a decay on the order of 200 ms, ImP decayed on the order of approximately 100 ms. After the initial decay (approximately 200 ms after loading), ImP quickly relaxed to base line, while SP continued to dominate relaxation. It appears that the decay of ImP is indicative of resistive fluid flow occurring primarily in the vasculature and other intraosseous channels such as lacunar-canalicular pores, and that SP represents the fluid flow in the smaller porosities, i.e., lacunar-canalicular system or even microspores. These results suggest that SP and ImP decays are determined by a hierarchical interdependent system of multiple porosities, and that the temporal dynamics of load-bearing define the manner in which the fluid patterns and pressures are distributed. PMID- 12108844 TI - Quantification of varying adhesion levels in chondrocytes using the cytodetacher. AB - The potential to promote cell adhesion must be evaluated in the development of tissue-engineered implants and scaffolds. One measure of cell adherence is the force necessary to detach the cell. The objective of this study was to evaluate the cytodetacher (Athanasiou, K. A., et al. Development of the cytodetachment technique to quantify cellular adhesiveness. Biomaterials 20: 2405-2415, 1999), modified to test cells grown on substrata, as a means of calculating cell adhesion forces. Live and formalin-fixed bovine and rabbit chondrocytes underwent cytodetachment to verify that the cytodetacher provides satisfactory resolution to differentiate between live and fixed cells. Fixed cells had significantly greater mechanical adhesiveness than those prepared live: the values for the fixed rabbit and bovine chondrocytes were 1.01 and 1.56 microN, respectively, versus 0.14 and 0.17 microN for the live cells (p<0.05). The sensitivity of the cytodetacher was also gauged by detaching live rabbit chondrocytes seeded for varying amounts of time (40, 80, and 120 min). For the 40, 80, and 120 min time points the maximum detachment forces were found to be significantly different: 2.87 x 10(-2), 6.75 x 10(-2), and 14.30 x 10(-)2 microN, respectively. This study validates the use of the modified cytodetacher as an effective means of evaluating the strength of adhesion of cells attached to a substratum. PMID- 12108845 TI - The effect of anterior cruciate ligament injury on knee joint function under a simulated muscle load: a three-dimensional computational simulation. AB - Understanding the biomechanical effect of various factors on knee behavior after anterior cruciate ligament (ACL) injury or reconstruction is instrumental for the development of an optimal surgical treatment of ACL injury that can better restore normal knee function. This paper presents the application of a three dimensional (3D) computational knee model for parametric studies of knee kinematics in response to simulated muscle loads. The knee model was constructed using the magnetic resonance images and biomechanical experimental data of the same cadaveric human knee specimen. The kinematics of the knee predicted by the computational model was compared with that measured from different specimens in a wide range of loading conditions and flexion angles. In general, the model predictions were within the range of experimental data. The model was then used to predict knee motion, ligament forces, and contact pressure in response to a simulated quadriceps force when the knee was ACL deficient. Partial ACL injury was simulated by reducing the stiffness of the ACL in the model. The results demonstrated that even with a reduction of 75% of the ACL stiffness, the ACL still carried a significant amount of the load (more than 58%) carried by an intact ACL. The kinematics (both tibial translation and rotation) varied less than 20% compared to that of the knee with intact ACL. The 3D computational model can be a powerful tool to simulate different variables that would influence knee function after ACL reconstruction, such as the initial tension of the ACL graft, the insertion sites of the graft, multibundle grafts, graft materials, and various physiological loading conditions. PMID- 12108846 TI - Free radical profiles in an encapsulated pancreatic cell matrix model. AB - The survival of encapsulated pancreatic cells or islets is often limited because of nutrient deficiency, fibrotic overgrowth, and immune attack. Activated immune cells, such as macrophages, release nitric oxide (NO) and superoxide (O2-). These species or their reactive intermediates, such as peroxynitrite, can be cytotoxic, mutagenic, and/or carcinogenic. The transport of these free radicals to encapsulated pancreatic cells cannot be impeded by the present immunoisolation technology. A model has been developed simulating free radical profiles within an encapsulation matrix due to macrophage immune cells attached to the surface of an encapsulation matrix. The model incorporates the transport and reactions of NO, O2- , O2, and total peroxynitrite (PER). The model predictions of NO, O2-, and PER concentrations to which pancreatic cells are potentially exposed are in the range of 8-42 microM, 0.5-8 nM, and 0.1-0.8 microM, respectively, for a 100-500 microm radius encapsulation matrix. The results demonstrate that the potential exists for free radical damage of encapsulated pancreatic cells and also demonstrates that additional exposure studies may be necessary for assessing free radical effects on pancreatic cell function. Also, care must be taken in assuming that encapsulated cell systems are completely protected from immunological action. PMID- 12108847 TI - Engineering analysis of ex vivo retroviral transduction system. AB - The overall dynamics of the retrovirus-cell encounter under a static retroviral transduction system can be described in terms of the process of uptake (adsorption/ internalization), decay, and diffusion. In this study, a mathematical model illustrating these processes was derived assuming a semi infinite domain and solved analytically using the Laplace transform. The closed form solutions for retroviral concentrations and time course profile of transduced cell colonies are presented to clarify the contributions of the processes involved in the retroviral transduction system. To manifest the usefulness of the closed-form solutions, the neomycin-resistant gene encoding retroviruses produced by two different packaging cells (human 293 cells and murine GP+ E86/LNCX cells) were employed to transduce NIH 3T3 cells, which formed neomycin-resistant colonies after G418 selection. The experimental results were curve fitted with the model-derived analytical solutions to quantitatively determine transduction rate constant k and initial concentration of infectious retrovirus C0. Our study showed that the vesicular stomatitis virus G protein pseudotyped retrovirus produced from 293 packaging cells exhibited much higher transduction rate (k = 0.0480 cm/h) than the ecotropic retrovirus (k = 0.0102 cm/h) produced from GP + E86/LNCX cells. The fitted values of C0 are approximately two orders of magnitude higher than the experimentally estimated titers for both retroviruses. PMID- 12108848 TI - Supramaximal flow in asthmatic patients. AB - In normal subjects supramaximal flows (SF) are known to be correlated with flow limitation. To further understand the mechanisms involved in SF this correlation and the influence of salbutamol and methacholine administration on SF have been investigated in asthmatic subjects. Protocol A involved obtaining basal maximal expiratory flow/volume curves and interrupted curves through a fast valve from 36 asthmatic patients. Maximal flow at 50% of forced vital capacity (V'max50) increase (deltaV'max50) was compared between basal curves and envelope curves passing through SF peaks obtained after each interruption. Protocol B involved the study of 33 asthmatic patients after salbutamol administration and 12 asthmatic patients after methacholine challenge. DeltaV'max50 between basal versus interrupted curves were analysed (deltaV'max50B). Similar procedures were performed after salbutamol (deltaV'max50S) and after methacholine administration (deltaV'max50M). A significant negative correlation between forced expiratory volume in one second and deltaV'max50 was observed. SF decreased significantly after salbutamol administration and increased significantly after methacholine. The results of this study suggest that in asthmatic patients participation of "pendelluft" in SF increases as airflow limitation increases. PMID- 12108849 TI - Effects of ventilation, humidity and temperature on airway responsiveness to methacholine in rats. AB - Exercise-induced bronchoconstriction is associated with heat and water loss from the airways. It is not known whether these conditions can influence the response to bronchoactive agonists. The effects of different degrees of alveolar ventilation on the pulmonary response to methacholine and the role of humidity and temperature in this response were evaluated. Wistar rats were anaesthetized, tracheostomized and mechanically ventilated. Increasing doses of methacholine were infused intravenously and respiratory system resistance (Rrs) and elastance (Ers) were measured. The rats were ventilated with dry air at 13 degrees C, dry air at 37 degrees C, humid air at 13 degrees C and humid air at 37 degrees C. These four groups were further divided into three subgroups with a respiratory frequency adjusted to reach a carbon dioxide tension in arterial blood of 30, 40 and 50 mmHg. Temperature, humidity and level of alveolar ventilation did not influence the position of the dose/response curve to methacholine. However, the maximal changes in Ers were significantly lower in the rats ventilated with humid air. In addition, maximal changes in Ers were significantly higher in the rats with lower alveolar ventilation. These differences were not observed for maximal values of Rrs. The pulmonary response to methacholine in normal rats is significantly affected by the humidity of inspired air and the level of alveolar ventilation. This influence is more intense in the small airways and/or distal airspaces. This suggests that exercise or hyperventilation can change the behaviour of airway smooth muscle. PMID- 12108850 TI - Changes of exhaled nitric oxide during steroid treatment of childhood asthma. AB - Exhaled nitric oxide (eNO) is elevated in several inflammatory airway diseases and is significantly reduced by anti-inflammatory treatment with inhaled steroids. The aim of this randomized, open clinical trial was to evaluate eNO in relation to conventional lung function parameters at rest and after exercise during sequential changes of inhaled steroids in children with persistent asthma. The study consisted of a 4 week run-in period, a 4 week washout phase and a randomized treatment period during which only one group was treated again with inhaled budesonide. After run-in, eNO was reduced to normal values, and rose again during washout. In the patients randomized to steroid treatment, eNO was again decreased, whereas it remained unchanged in the untreated patients. Forced expiratory volume in one second and forced vital capacity at rest and after exercise improved significantly after run-in, but showed no difference after randomization. However there was a strong correlation of eNO with patient compliance. Exhaled nitric oxide was able to differentiate between children briefly treated with or without steroids, the conventional lung-function variables however could not. In practice exhaled nitric oxide may thus be a valuable parameter to monitor adherence to steroids, but less suitable to describe physiologically relevant impairments of lung function. PMID- 12108851 TI - Respiratory impedance response to a deep inhalation in asthmatic children with spontaneous airway obstruction. AB - The aim of the study was to determine whether the bronchomotor effect of a deep inhalation (DI) may be detected during tidal breathing in asthmatic children with spontaneous airway obstruction (AO). Two groups of children aged 5-15 yrs were studied. AO was mild in group 1 (n=12, forced expiratory volume in one second (FEV1) > or = 75% predicted) and moderate-to-severe in group 2 (n=9, FEV1 > or = 70% pred). The forced oscillation technique at 12 Hz using a head generator allowed the determination of respiratory resistance in inspiration (Rrsi) and expiration (Rrse) before and after DI, at baseline and after salbutamol. At baseline, Rrsi but not Rrse was found to decrease significantly after DI in group 1 but not in group 2. The change induced by DI was significantly different in group 1 (-1.5+/-0.5 hPa x s x L(-1)) compared to group 2 (0.5+/-0.5 hPa x s x L( 1)) and exhibited significant negative correlation to FEV1 % pred. After salbutamol, DI had no effect. In conclusion, asthmatic children show a bronchomotor response to deep inhalation that depends on the degree of airway obstruction. The effect is more readily demonstrated in inspiration than in expiration. PMID- 12108852 TI - Asthma prevalence in children living in villages, cities and refugee camps in Palestine. AB - Previous studies have suggested that asthma prevalence is generally lower in the Middle East than in more developed countries. The aim of this study was to investigate the prevalence and severity of asthma and asthma symptoms in schoolchildren in the Ramallah District in Palestine. In the autumn of 2000, 3,382 schoolchildren aged 6-12 yrs were surveyed in 12 schools, using the International Study for Asthma and Allergies in Childhood (ISAAC)-phase III, parents-administered translated questionnaire. The crude prevalence rates for "wheezing-ever", "wheezing in the previous 12 months", and "physician-diagnosed asthma" were 17.1, 8.8 and 9.4% respectively, with urban areas having higher prevalence rates than rural areas. Within urban areas, refugee camps had higher prevalence rates than cities. Yet, within the rural areas, the 12-month prevalence was lower in the deprived villages than other residences. Place of residence remained significant for asthma and asthma symptoms, after adjusting for sex, age, and place of birth. To conclude, children from refugee camps appear to be at higher risk of asthma than children from neighbouring villages or cities. The prevalence of asthma and asthma symptoms in Palestine appears to be close to that of Jordan, but it is much lower than Israel, and lower than some other countries in the region, such as Kuwait and Saudi Arabia, and more developed countries. This initial study is a baseline for a study on lifestyle and environmental determinants for asthma among Palestinian children. PMID- 12108853 TI - Hospitalization trends for paediatric asthma in eastern Finland: a 10-yr survey. AB - The prevalence of childhood asthma has increased during the past 20 yrs in eastern Finland. The use of regular medication for asthma has, consequently, also risen at the same time. The aim of the present study was to assess how these factors have influenced hospitalization for asthma in children. Data concerning the annual number of children aged <16 yrs treated for asthma, their number of hospital periods, and length of hospitalization were collected using patient specific medical records for the years 1988-1997. Admissions for asthma more than doubled (from 1.2 in 1,000 to 2.7 in 1,000) (p<0.001), whereas re-admissions decreased from 28.1% to 8.7% (p<0.001) during the study period. The greatest increase was seen among children aged <2 yrs, whereas during recent years the admission rate among children aged >2 yrs has declined by one-third. Despite the increased prevalence of asthma, the decline in admissions seen in all but the youngest children may, at least partly, be attributed to the decrease in re admissions, resulting from the early onset and increased use of inhaled steroids. PMID- 12108854 TI - Trends in prevalence of atopic diseases and allergic sensitization in children in Eastern Germany. AB - Trends in prevalence of atopic diseases and allergic sensitization in children from Eastern Germany during the 1990s were analysed. The study consisted of three regional cross-sectional surveys of a total of 7,632 children (aged 5-14 yrs) in 1992-1993, 1995-1996, and 1998-1999. Information was gathered on atopic diseases and potential predictors by a parental questionnaire. Allergic sensitization for birch, grass, mite, cat, and cladosporium were assessed by radioallergosorbent test (RAST). After adjustment for age, sex and the study area of the participants, prevalence increased between the first and third survey for hay fever, for asthma and for atopic eczema. The adjusted prevalence of allergic sensitization (RAST > 0.35 kU x L(-1)) showed a decrease, whereas the prevalence of strong sensitization (RAST > or = 17.5 kU x L(-1)) increased significantly, specifically in cohorts born after 1989. Further adjustment for possible determinants of these atopic diseases did not change the trend estimates. A clear increase in the prevalence of atopic diseases, with the exception of hay fever, was observed as well as a shift towards a stronger allergic sensitization, which might affect the onset of clinical manifestations of atopic diseases. Differences in the epidemiology of respiratory symptoms, illnesses and allergies between populations living in the former East Germany and those living in the former West Germany have been reported. Among East German children, lower prevalence rates of asthma, and positive skin-prick tests were observed compared to West German children in the early 1990s. Similarly, among East German adults, lower specific immunoglobulin (Ig) E levels and lower prevalence rates of asthma, wheezing, positive methacholine-challenge tests, allergic rhinitis, and positive skin-prick tests have been reported compared to those of West German adults. PMID- 12108855 TI - Dyspnoea perception during clinical remission of atopic asthma. AB - Symptoms of atopic asthma often disappear around puberty. The authors recently demonstrated that this clinical remission is accompanied with ongoing airways inflammation in most subjects. The discrepancy between lack of symptoms and persistent airway inflammation suggests that perception of the symptoms is unclear. In the present study, young adults in clinical remission of atopic asthma assigned themselves a modified Borg score during methacholine and adenosine-5'-monophosphate induced bronchoconstriction. Borg scores of subjects in clinical remission were compared with those of symptomatic asthmatic subjects. A marked variation in the Borg scores at a 20% fall in the forced expiratory volume in one second was found. Significant differences in Borg scores between remission patients and asthmatics could not be detected. It was concluded that perception of dyspnoea, induced with methacholine and adenosine challenge, is similar in young adults in clinical remission of atopic asthma compared to that of patients with symptomatic asthma. Hence, an unclear perception seems to be an unlikely explanation for the discrepancy between lack of symptoms and ongoing inflammation. Other factors, including both physical and psychological ones, may play a role in the apparent absence of symptoms, thereby potentially leading to undertreatment. PMID- 12108856 TI - Role of secretory leukocyte protease inhibitor in the development of subclinical emphysema. AB - Secretory leukocyte protease inhibitor (SLPI) is a potent inhibitor of human leukocyte elastase. In some chronic airway diseases, the level of SLPI is decreased in sputum, leading to the continuation of neutrophil inflammation. In this study, the role of SLPI in subclinical pulmonary emphysema was examined. Sequential bronchoalveolar lavage was performed in an attempt to separately evaluate the levels of SLPI in the large airways and in the peripheral airways for two groups of smokers. One group had subclinical emphysema by computed tomography scans and one group did not. SLPI localized in alveolar macrophages (AM) was also assessed. The level of SLPI was significantly elevated in the peripheral airways from the subjects with emphysema compared to those without emphysema (1589.2+/-353.9 versus 729.1+/-31.0 ng x mL(-1)), although it was similar in the large airways (3442.3+/-499.6 versus 2535.7+/-578.8 ng x mL(-1)). There was a trend for higher amount of SLPI to be released from AM in subjects with subclinical emphysema, although this did not reach statistical significance. In conclusion, there is compensatory upregulation of secretory leukocyte protease inhibitor in peripheral airways in subclinical pulmonary emphysema, which is in sharp contrast to the decreased level of secretory leukocyte protease inhibitor reported in some chronic airway diseases. PMID- 12108857 TI - Bone mineral density in patients with chronic obstructive pulmonary disease treated with budesonide Turbuhaler. AB - There is a need for studying the effects of long-term inhaled corticosteroid therapy on bone mineral density (BMD) and vertebral fracture rates in patients with mild chronic obstructive pulmonary disease (COPD). Patients (n=912, mean age 52 yrs) with mild COPD (mean forced expiratory volume in one second (FEV1) 77% of predicted; mean FEV1/slow vital capacity ratio 62%) were randomized to receive budesonide 400 microg, or placebo twice daily via Turbuhaler. BMD was measured at the L2-L4 vertebrae and the femoral neck, trochanter and Ward's triangle by dual energy X-ray absorptiometry at baseline and after 6, 12, 24 and 36 months (n=161). Radiographs of the thoracic and lumbar spine were obtained at the beginning and end of treatment (n=653). Previous fractures were present at baseline in 43 budesonide-treated patients (13.4%) and 38 placebo-treated patients (11.5%). New fractures occurred in five budesonide-treated patients, compared with three in the placebo group (p=0.50). There were no significant changes in BMD at any site in budesonide-treated patients, compared with the placebo group, during the course of the study. Budesonide treatment was associated with a slight but statistically significant decrease in the area under the concentration-time curve for serum osteocalcin. In the present study, involving a large group of patients with chronic obstructive pulmonary disease, long-term treatment with budesonide 800 microg x day(-1) via Turbuhaler had no clinically significant effects on bone mineral density or fracture rates. PMID- 12108858 TI - Alterations in gene expression in hamster diaphragm after emphysema and lung volume reduction surgery. AB - The authors have demonstrated previously that emphysema and lung volume reduction surgery (LVRS) resulted in a significant shift of type IIx/b to type IIa fibres in the diaphragm of hamsters with elastase-induced emphysema. To explore the mechanisms leading to this fibre switching, the mRNA expression of the myogenic regulatory factors, the inhibitors of DNA binding proteins (Id-proteins) and insulin-like growth factor-I were examined. Ribonucleic acid was extracted from the diaphragm of control, emphysematous, emphysematous and sham operated and LVRS hamsters and subjected to reverse transcriptase polymerase chain reaction. Compared to control, the ratio MyoD to myogenin declined with emphysema, sham and even more after LVRS, due to a decrease in MyoD mRNA and an increase in myogenin mRNA. Similarly, compared to control, Id-1 protein mRNA levels decreased significantly in sham and even more in LVRS. Id-2 protein mRNA levels decreased in all groups, but reached statistical significance in LVRS only, compared to control. IN CONCLUSION: 1) the reduced MyoD/myogenin ratio may be the mechanism of the shift to a slower fibre type, 2) the decreased MyoD/myogenin ratio in lung volume reduction surgery animals suggests that lung volume reduction surgery enhances rather than decreases the load placed on the diaphragm and 3) the observed down-regulation of the inhibiting factors may facilitate the diaphragm adaptation to overload. PMID- 12108859 TI - Resistance versus endurance training in patients with COPD and peripheral muscle weakness. AB - The effects of endurance training on exercise capacity and health-related quality of life (HRQL) in chronic obstructive pulmonary disease (COPD) patients have been studied thoroughly, while resistance training has been rarely evaluated. This study investigated the effects of resistance training in comparison with endurance training in patients with moderate to severe COPD and peripheral muscle weakness (isometric knee extension peak torque <75% predicted). Forty-eight patients (age 64+/-8 yrs, forced expiratory volume in one second 38+/-17% pred) were randomly assigned to resistance training (RT, n=24) or endurance training (ET, n=24). The former consisted of dynamic strengthening exercises. The latter consisted of walking, cycling and arm cranking. Respiratory and peripheral muscle force, exercise capacity, and HRQL were re-evaluated in all patients who completed the 12-week rehabilitation (RT n=14, ET n=16). Statistically significant increases in knee extension peak torque (RT 20+/-21%, ET 42+/-21%), maximal knee flexion force (RT 31+/-39%, ET 28+/-37%), elbow flexion force (RT 24+/-19%, ET 33+/-25%), 6-min walking distance (6MWD) (RT 79+/-74 m, ET 95+/-57 m), maximum workload (RT 15+/-16 Watt, ET 14+/-13 Watt) and HRQL (RT 16+/-25 points, ET 16+/-15 points) were observed. No significant differences in changes in HRQL and 6MWD were seen between the two treatments. Resistance training and endurance training have similar effects on peripheral muscle force, exercise capacity and health-related quality of life in chronic obstructive pulmonary disease patients with peripheral muscle weakness. PMID- 12108860 TI - Contribution of respiratory acidosis to diaphragmatic fatigue at exercise. AB - The factors that may modulate ventilatory muscle fatigue during exercise are controversial. In this study the contribution of acidosis to exercise-induced diaphragmatic fatigue was investigated, using measurements of the twitch mouth pressure response (tw,Pmo) to cervical magnetic stimulation. After learning sessions, 14 healthy subjects performed two cycling tests (at 60% of maximal aerobic power for 16 min), one while breathing spontaneously (mean minute ventilation (V'E) 67.9 L x min(-1)) and the other while hypoventilating voluntarily (mean V'E 53.8 L x min(-1)). Exercise was voluntarily set at a moderate power to avoid a fatiguing effect of exercise per se. As compared with spontaneous breathing (SB), voluntary hypoventilation (VHV) significantly increased mean carbon dioxide tension in arterial blood (Pa,CO2) (51 mmHg versus 41 mmHg) and significantly decreased arterial pH (7.28 versus 7.34). After 10 min of SB test, tw,Pmo was unchanged compared to the baseline value (19.1 versus 18.5 cmH2O) whereas tw,Pmo fell significantly as compared to baseline (17.1 versus 18.5 cmH2O) and to SB (17.1 versus 19.1 cmH2O) after the VHV test. The results of this study suggest that exposure to hypercapnia may impair respiratory muscle function. This impairment could be more clinically relevant in patients with chronic obstructive lung disease. PMID- 12108861 TI - Prospective population-based study on the survival of patients with lung cancer. AB - The purpose of this study was to evaluate the change, over 20 yrs, in the survival of lung cancer patients in a population-based study. Information on all patients with lung cancer in a defined geographical area during 1990-1992 (n=602) was prospectively gathered. The survival of these patients was assessed and also compared with the results of a similar study in the same area during the years 1968-1971 (n=446). The 5-yr survival had improved during 20 yrs from 4% to 12%. The 5-yr survival of the patients with squamous cell carcinoma had increased from 6% to 16%, and adenocarcinoma from 4% to 19%, whereas the survival of small cell carcinoma had remained the same (2% and 3%, respectively). Even though the recent patients were older than those of the earlier series the proportion of surgically treated patients had remained the same (16% and 20%), but the 5-yr survival of patients who had been operated on had increased significantly from 23% to 48%. The differences in survival in the second cohort (1990-1992) between histological types (Chi-squared logrank=59.2), tumour, node, metastasis stages (Chi-squared logrank=199.6), symptomatic stages (Chi-squared logrank=120, p<0.001) and treatment (Chi-squared logrank=277) were significant. Based on this study the independent prognostic factors for better survival of lung cancer patients are tumour, node, metastasis stages I and II, surgical treatment and Feinstein's symptomatic stages I and II. PMID- 12108862 TI - The correlation of emphysema or airway obstruction with the risk of lung cancer: a matched case-controlled study. AB - A matched case-controlled study was conducted to determine if airway obstruction or emphysema were associated with an increased risk of lung cancer. Lung cancer cases (n=24) were identified through a low-dose spiral computed tomography (CT) screening trial from 1,520 participants. Four controls without lung cancer were selected for each case from the participants and matched by sex, age and smoking history. Emphysema was assessed by quantitative CT analysis. Conditional logistic regression was employed to assess results of spirometry and CT quantitative analysis as potential risk factors for lung cancer. The likelihood of lung cancer was found to be significantly increased for those with forced expiratory volume in one second (FEV1) < or = 40% of predicted. The results suggested that a lower percentage of predicted FEV1 was indicative of lung cancer. No compelling evidence was found to suggest that the percentage of emphysema was associated with lung cancer. These results suggest an increased risk of lung cancer associated with airway obstruction. However, percentage of emphysema as determined by computed tomography was not associated with an increased risk of lung cancer. PMID- 12108864 TI - Socioeconomic outcome of subjects experiencing asthma symptoms at work. AB - The aim of this study was to investigate the socioeconomic outcomes of subjects who experienced work-related asthma symptoms in the absence of demonstrable occupational asthma (OA) and to compare these outcomes with those found in subjects with documented OA. Subjects (n=157) who were being investigated for work-related asthma, were surveyed. Of these 86 had OA, ascertained by a positive specific inhalation challenge (SIC), and 71 subjects had a negative SIC response. After a median interval of 43 months (range 12-85 months), the subjects were interviewed to collect information on employment status, income changes, and asthma-related work disability. Rates of work disruption and income loss at follow-up were similar in subjects with negative SIC (46% and 59%, respectively) and in those with OA (38% and 62%). The median loss as a percentage of initial income was 23% in subjects with negative SIC and 22% in subjects with OA. Asthma related work disability, defined as any job change or work loss due to asthma, was slightly more common in subjects with OA (72%) than in those with negative SIC (54%). This study shows that, even in the absence of demonstrable occupational asthma, work-related asthma symptoms are associated with considerable socioeconomic consequences. PMID- 12108863 TI - Lung function of school children with low levels of alpha1-antitrypsin and tobacco smoke exposure. AB - Exposure to environmental tobacco smoke (ETS) and other air pollutants has been associated with small decrements in lung function. The susceptibility to pollution exposure may, however, vary substantially between individuals. Children with an impaired protease-antiprotease balance may be particularly vulnerable. Therefore this study aimed to investigate the effects of ETS exposure on children with reduced levels of alpha1-antitrypsin (alpha1-AT). Random samples of school children (aged 9-11 yrs) (n=3,526) were studied according to the International Study of Asthma and Allergies in Childhood (ISAAC) phase II protocol, including parental questionnaires, pulmonary function and allergy testing. Blood samples were obtained to measure plasma levels of alpha1-AT and to genotype for pleomorphic protein inhibitor (Pi)Z and PiS alleles. Children with low levels of alpha1-AT (< or = 116 mg x dL(-1)) showed significant, albeit small decrements in baseline lung function. When exposed to ETS, pronounced decrements of pulmonary function, particularly in measures of mid- to end-expiratory flow rates, were seen in these children as compared to exposed children with normal levels of alpha1-AT. The mean levels of % predicted+/-SE in both groups were: maximum expiratory flow at 50% of vital capacity 79.4+/-7.2 versus 99.0+/-1.5, maximum expiratory flow at 25% of vital capacity 67.4+/-10.0 versus 100.3+/-2.1, maximal midexpiratory flow 73.7+/-8.6 versus 99.9+/-1.7. These findings suggest that school children with low levels of alpha1-antitrypsin are at risk of developing pronounced decrements in pulmonary function, particularly if they are exposed to environmental tobacco smoke. Parents of children with heterozygous alpha1 antitrypsin deficiency resulting in significantly reduced blood concentrations should be advised to prevent their children from being exposed to environmental tobacco smoke and dissuade them from taking up smoking. PMID- 12108865 TI - Survival of patients with biopsy-proven usual interstitial pneumonia and nonspecific interstitial pneumonia. AB - This is the first Australian study to examine survival and clinical characteristics in biopsy-proven idiopathic interstitial pneumonia. A cohort of 70 patients from a single institution between January 1990 and December 1999 was reviewed. All patients were Caucasian, 23 (33%) female. Mean age+/-SD at diagnosis was 60+/-12 yrs for males and 54+/-14 yrs for females. A total 24% of patients had never smoked. The histopathological diagnoses were usual interstitial pneumonia (UIP) (n=59), nonspecific interstitial pneumonia (NSIP) (n=7), desquamative interstitial pneumonia (n=3) and acute interstitial pneumonia (n=11). Clinical and functional characteristics of the two main histological subgroups of UIP and NSIP showed significantly older patients in the UIP group and a significantly lower mean forced expiratory volume in one second (FEV1) in the NSIP group. Median survival for UIP was 78 months compared with 178 months for NSIP. No survival difference between treated and untreated patients with UIP was found. Multivariate analysis revealed smoking alone to be predictive of poorer survival. This study demonstrates the best median survival for usual interstitial pneumonia of available series and confirms a survival difference between usual interstitial pneumonia and nonspecific interstitial pneumonia. Furthermore, the reported results may have implications for treatment timing using conventional protocols currently recommended. PMID- 12108866 TI - Glutathione deficiency of the lower respiratory tract in patients with idiopathic pulmonary fibrosis. AB - Idiopathic pulmonary fibrosis (IPF) is a disease of unknown aetiology. Increased oxidant burden and antioxidant, e.g. glutathione (GSH), deficiency in the lower respiratory tract have been thought to play a role in the progression of IPF. Sputum induction is a safe noninvasive tool to study inflammation in the respiratory tract. The aim of the present study was to evaluate the direct measurement of GSH in induced sputum supernatant. Sixteen IPF patients and 15 healthy, nonsmoking subjects underwent sputum induction. Total GSH in sputum, saliva and plasma was measured spectrophotometrically. Sputum GSH was decreased more then four-fold in IPF patients when compared to healthy subjects (mean GSH 1.4+/-0.34 microM versus 5.8+/-0.98 microM). Salivary GSH was generally low or undetectable in all subjects. Plasma GSH levels were lower in IPF patients (0.26+/-0.1 versus 0.74+/-0.16 microM). In IPF patients, there was a borderline correlation of sputum GSH levels with disease duration and lung-function impairment. These data confirm the established role of oxidant/antioxidant imbalance in the pathogenesis of idiopathic pulmonary fibrosis, and show the potential of induced sputum to directly study inflammatory processes and surrogate markers in interstitial lung diseases like idiopathic pulmonary fibrosis. PMID- 12108867 TI - The rapamycin analogue SDZ RAD attenuates bleomycin-induced pulmonary fibrosis in rats. AB - Pulmonary fibrosis is characterized by excessive deposition of extracellular matrix proteins within the pulmonary interstitium. The new macrolide immunosuppressant SDZ RAD, a rapamycin analogue, inhibits growth-factor dependent proliferation of mesenchymal cells and might therefore be of therapeutic interest for the treatment of fibrotic lung disease. In this study the effect of SDZ RAD on lung-collagen accumulation in the bleomycin model of pulmonary fibrosis in rats was investigated. SDZ RAD (2.5 mg x kg(-1) x day(-1)) or drug vehicle were administered orally by daily gavage. Successful dosing was confirmed by measuring splenic weight. Total lung-collagen content was measured by high-performance liquid chromatographic quantitation of hydroxyproline. In animals given bleomycin and drug vehicle, total lung collagen was increased by 182+/-11% (mean+/-SEM) compared with saline controls at 14 days (p<0.001). The increase in lung-collagen accumulation was reduced by 75+/-12% (p<0.01) in animals given SDZ RAD and was accompanied by a concomitant 56+/-6% (p<0.001) reduction in lung weight. SDZ RAD is currently in clinical trials for the prevention of solid organ graft rejection, another condition characterized by excessive extracellular matrix production. The authors propose that SDZ RAD warrants evaluation as a novel therapeutic agent for fibrotic lung disease. PMID- 12108868 TI - Surfactant apoprotein A modulates interleukin-8 and monocyte chemotactic peptide 1 production. AB - Previous studies have shown that surfactant apoprotein A (SP-A) and natural or synthetic surfactant can modulate the release of pro-inflammatory cytokines from alveolar mononuclear phagocytes. The aim of this study was to assess whether SP-A or Surfactant (Surf) from patients with pulmonary alveolar proteinosis (PAP) can affect the release of two chemokines (interleukin (IL)-8 and monocyte chemtactic peptide (MCP)-1) from human monocytes and rat lung type-II cells. In addition IL 8 and MCP-1 levels were assessed in the brochoalveolar lavage fluid (BALF) of seven patients with PAP and compared with those in a group of control subjects (n=5). SP-A, tested over a wide range of concentrations, significantly increased IL-8 and MCP-1 release from monocytes. SP-A retained its activity after collagenase digestion, but was not active after heat treatment. The release of IL 8 by monocytes was also stimulated by Surf. Finally, median BALF IL-8 and MCP-1 levels in PAP patients were significantly higher than in controls (9.50 and 9.51 pg x mL(-1) in controls versus 151.95 and 563.70 pg x mL(-1) in PAP, respectively) and significantly correlated with SP-A concentrations in BALF. Overall the results of this study support the view that the high content of alveolar surfactant apoprotein A may contribute to the upregulation of chemokine release in pulmonary alveolar proteinosis, thus contributing to airway inflammation. PMID- 12108869 TI - Colistin stimulates the activity of neutrophil elastase and Pseudomonas aeruginosa elastase. AB - Nonantimicrobial effects of antibiotics may contribute to their activity in the treatment of infective airway disease. The aim of this study was to identify antibiotics used for the treatment of infection in cystic fibrosis that may alter the activity of human neutrophil elastase (HNE) and Pseudomonas aeruginosa elastase (PE). The effect of antibiotics on the activity of purified HNE and PE, and HNE in sputum was assessed using colourimetric and fluorescent substrate assays by kinetic measurements, and by examining the interaction of HNE with inhibitors. Ceftazidime, tobramycin, and gentamycin slightly inhibited purified HNE activity whereas erythromycin and colistin significantly stimulated purified HNE and PE (395 and 557%, respectively). However, only colistin increased HNE activity in sputum (+102%) and was therefore studied in more detail. This increase in activity was not due an interference with the specific inhibition of HNE by alpha1-antitrypsin but colistin was found to reverse the inhibitory effects of small molecular weight molecules like heparin. Colistin increases the activity of human neutrophil elastase and Pseudomonas aeruginosa elastase, two proteases that contribute to the pathogenesis of cystic fibrosis airway disease. PMID- 12108870 TI - Role of alveolar epithelial ICAM-1 in lipopolysaccharide-induced lung inflammation. AB - Intercellular adhesion molecule-1 (ICAM-1) is known to play a central role in lung inflammation. Limited information, however, is available regarding the expression and biological function of ICAM-1 in the alveolar epithelial compartment. The current report analyses the expression pattern of ICAM-1 in primary cultures of rat alveolar epithelial cells (AECs) and in the rat lung following instillation of bacterial endotoxin (lipopolysaccharide (LPS)) in order to better define the role of alveolar epithelial ICAM-1. AECs stimulated in vitro with LPS were evaluated for ICAM-1 and ICAM-1 messenger ribonucleic acid content. Adherence assays with neutrophils and macrophages were performed. Endotoxin induced ICAM-1 upregulation on AECs was demonstrated in vivo by immunofluorescence staining. In addition, the effect of intratracheally-instilled anti-ICAM-1 was assessed. Significant upregulation of ICAM-1 occurred in vitro and in vivo on AECs after LPS stimulation. Adherence assays showed a 114% increase in adhesion of neutrophils to AECs. Antibody directed against ICAM-1 reduced this adhesion by 40%. A significant reduction in the number of neutrophils in bronchoalveolar lavage fluid and whole lung was seen under airway ICAM-1 blockade. These data indicate that intercellular adhesion molecule-1 participates in the inflammatory response to lipopolysaccharide-induced lung injury in the distal airways by interacting mainly with neutrophils. PMID- 12108871 TI - Role of biomarkers for early detection of lung cancer and chemoprevention. AB - Lung cancer is the leading cause of cancer deaths in developed countries. The poor prognosis associated with this disease is closely related to the fact that most lung cancer patients are not identified until their malignancy has reached an advanced stage. Recent advances have added to the understanding of the morphological and molecular characteristics of preinvasive bronchial lesions and early lung cancers. Such information is being used to provide new tests for the detection of lung cancer at early or preinvasive stages, and for identifying targets for therapeutic intervention that can prevent progression to advanced disease. Laser induced fluorescence endoscope bronchoscopy has improved the sensitivity with which preinvasive dysplastic bronchial lesions and early invasive malignancies can be detected. Morphological features of such lesions have been described and can be monitored by follow-up bronchoscopies in order to validate potential chemoprevention treatments. Distinct morphological characteristics such as angiogenic squamous dysplasia also suggest that processes like angiogenesis are present early in the development of lung cancer. Furthermore, tissue obtained from these early lesions has been used to describe alterations in the expression of a number of factors that distinguish these early lesions from normal bronchial epithelium. This could provide molecular markers and targets for the detection and treatment of early lung cancer. Studies to detect these alterations by polymerase chain reaction and/or immunhistochemical analyses of easily obtained specimens such as sputa are helping identifing molecular markers that could be utilized in effective screening programmes. The current article reviews new findings regarding the molecular biology of preinvasive bronchial lesions and early lung cancers, and describes new developments regarding their application in the early detection and chemoprevention of lung cancer. PMID- 12108872 TI - Where to perform noninvasive ventilation? AB - Noninvasive positive-pressure ventilation (NPPV) has been shown to be a means of reducing the need for endotracheal intubation, which when effective reduces the complication rate and improves outcome. Because paralysis and sedation are not needed and because the patient is not necessarily dependent upon a machine for respiration, ventilation outside the intensive care unit (ICU) is an option. A number of studies have shown that NPPV for acute exacerbations of chronic obstructive pulmonary disease (COPD) can be effective in the non-ICU environment, though usually in patients with less severe exacerbations. However, there have been no direct comparisons of the application of NPPV in different locations. The likelihood of success of the technique is an important factor in deciding there NPPV should be performed. Ready access to invasive ventilation is important when NPPV is not indicated from the outset or fails after an initial trial. In acute exacerbations of COPD, NPPV is less likely to be successful the more severe the exacerbation, as measured by the severity of acidosis. Good tolerance of NPPV, which translates into an improvement in pH and a fall in respiratory rate, predicts a successful outcome and is a useful way of monitoring progress. NPPV has been shown to be cost effective both in the ICU and when performed on general wards. A dedicated intermediate care unit with particular expertise in noninvasive modes of ventilation may provide the best environment, both in terms of outcome, but also cost effectiveness. The ideal location for noninvasive positive-pressure ventilation will vary from country to country and indeed from hospital to hospital, depending upon local factors. However, the most important factor is that staff be adequately trained in the technique and be available throughout the 24-h period. PMID- 12108873 TI - Evaluating the effects of asthma therapy on childhood growth: principles of study design. AB - Inhaled corticosteroids have been established as the most effective treatment for childhood asthma. However, concerns persist regarding their potential effects on growth and, most importantly, final height. To assess their effects on growth, inhaled corticosteroids can be compared with placebo (type 1 study), nonsteroidal anti-asthma therapy (type 2 study), another inhaled corticosteroid (type 3 study) or "real-life" anti-asthma therapy (type 4 study). Owing to the difficulties in obtaining final height data, several different surrogate measures have often been used: short-term lower leg growth, longer-term statural height growth velocity, childhood height and predicted final height. This paper discusses the choice of end point, key trial design issues (including selection and number of subjects in the active and control populations) duration of assessments and methods for measuring height and data analysis, in the context of the different study types. Specific study design recommendations have been developed after consideration of these factors, and these principles will be used to guide the interpretation of previously published growth studies. PMID- 12108874 TI - Evaluating the effects of asthma therapy on childhood growth: what can be learnt from the published literature? AB - The difficulties of assessing the effects of asthma therapy on childhood growth were explored in the first part of this review. In this part of the review growth studies with inhaled corticosteroids were selected that included a control group, measured height by stadiometry and were of > or = 1 yr duration. The studies were classified as type 1 (placebo control), type 2 (nonsteroidal therapy control), type 3 (comparator inhaled corticosteroid control) or type 4 ("real-life" studies with dose adjustment). The design attributes of these studies were then compared with the recommendations described in the first part of this review. Of the 18 studies identified, 17 were susceptible to one or more important confounding factors. Nevertheless, the outcomes of all 18 studies were mostly consistent. At recommended doses, beclomethasone dipropionate and budesonide demonstrated a small degree of growth suppression over 1-2 yrs (study types 1 and 2), but there was little evidence of such an effect with fluticasone propionate. Studies comparing different inhaled corticosteroids at recommended doses indicated more rapid growth with fluticasone propionate than with beclomethasone dipropionate or budesonide. However, none of the inhaled corticosteroids appeared to affect final height. In conclusion, the results from the majority of published growth studies with inhaled corticosteroids must be interpreted with a degree of caution owing to their potential susceptibility to important confounding factors. Further well designed studies are needed to establish whether different inhaled corticosteroids have different effects on growth in the long term. PMID- 12108875 TI - Sleep and breathing in neuromuscular disease. AB - Respiratory muscle weakness in neuromuscular disease causes significant morbidity and mortality. The published data on respiratory muscle activity and breathing during sleep in normal subjects, the impact of respiratory muscle weakness on sleep and breathing and the relations to daytime respiratory function in neuromuscular disease are reviewed here. In normal subjects during sleep upper airway resistance increases, chemosensitivity is reduced and the wakefulness drive to breathe is lost, resulting in a fall in ventilation. During rapid eye movement (REM) sleep, ribcage and accessory breathing muscles are suppressed, particularly during bursts of eye movements, and breathing is more irregular, rapid and shallow, with a further fall in ventilation. In subjects with respiratory muscle weakness sleep is fragmented, with shorter total sleep time, frequent arousals, an increase in stage 1 sleep and a reduction in, or complete suppression of, REM sleep. Sleep-disordered breathing and nocturnal desaturation are common and most severe during REM sleep. Correlations between daytime respiratory function and nocturnal desaturation are moderate or weak, but daytime respiratory function has greater prognostic value than nocturnal measurements. Noninvasive ventilation improves sleep quality and breathing in subjects with respiratory muscle weakness. However, the optimal criteria for initiation of ventilation and its role in rapidly progressive neuromuscular diseases are unclear. PMID- 12108876 TI - Pulmonary medicine and (adult) critical care medicine in Europe. PMID- 12108877 TI - Acute adrenal crisis in asthmatics treated with high-dose fluticasone propionate. AB - Four cases of asthma (one adult, three children) developing acute adrenal crisis after introduction of high-dose inhaled fluticasone proprionate are presented. The three children, aged 7-9 yrs, had been prescribed inhaled fluticasone, dosage 500-2,000 microg x day(-1) and duration 5 months-5 yrs. All presented with convulsions due to hypoglycaemia (blood glucose 1.3-1.8 mM). The fourth case was a male of 33 yrs with difficult-to-control asthma and had been taking fluticasone propionate 1,000-2,000 microg x day(-1) for 3 yrs. He presented with fatigue, lethargy, nausea and postural hypotension. Acute adrenal crisis in each case was confirmed by investigations which included measurement of acute phase cortisol levels, short and long Synacthen stimulation tests and glucagon stimulation tests. Other cases of hypthoalamic-pituitary-adrenal axis suppression were excluded. PMID- 12108878 TI - Cough, fever and weight loss in a young male. PMID- 12108879 TI - Evaluation of impulse oscillation system: comparison with forced oscillation technique and body plethysmography. PMID- 12108880 TI - Total energy expenditure in children with obstructive sleep apnoea syndrome. PMID- 12108881 TI - Nasal continuous positive airway pressure for sleep apnoea following stroke. PMID- 12108883 TI - Polysomnography for the management of progressive neuromuscular disorders. PMID- 12108882 TI - Inhaled steroids in children: adrenal suppression and growth impairment. PMID- 12108884 TI - Studying human airway pharmacology in microsections: application of videomicrometry. AB - The influence of endogenously-released mediators and activated eosinophils on the airway lumen and the effect of passive sensitization on anti-immunoglobulin (Ig) E-induced contractile responses was investigated by videomicrometry. Human bronchial sections of 2-3 mm internal diameter, placed in 250 microL Hank's balanced salt solution on microtitre plates, were monitored and recorded by digitized image analysis. Airway preparations exhibited a spontaneous narrowing (mean+/-SEM -33+/-5% of the luminal area). Removal of the bronchial epithelium almost completely prevented the development of spontaneuous narrowing (-6+/-3%; p<0.001). The addition of platelet-activating factor stimulated human eosinophils to the bronchial sections led to significant narrowing of the airway lumen (-39+/ 9%; p<0.05). Passive sensitization induced hyperresponsiveness to polyclonal anti IgE (-35+/-8%; p<0.01). It is concluded that videomicrometry is suitable for studying interactions between human airways and inflammatory cells, as well as the effect of passive sensitization on smooth muscle reactivity in vitro, without the imposition of preload. Under these conditions, human airways exhibited a spontaneous decrease of the airway lumen over time suggesting a role for epithelium-derived mediators because the development of spontaneous tone was epithelium dependent. PMID- 12108885 TI - Effect of inhaled fluticasone on bronchial responsiveness to neurokinin A in asthma. AB - Neurokinin (NK) A causes airway narrowing in patients with asthma through direct and indirect mechanisms. The effects of the inhaled glucocorticosteroid fluticasone propionate (FP) on the bronchial responsiveness to NKA and methacholine were studied. Patients (n=11) with mild asthma participated in a randomized, double-blind, placebo-controlled crossover trial. FP (500 microg b.i.d.) or matched placebo was administered via Diskhaler for 14 days. Bronchial challenges were performed on days 1 and 13 (methacholine) and 0 and 14 (NKA) for each treatment period. At the active treatment period, the mean log2 provocative concentration causing a 20% fall in the forced expiratory volume in one second (PC20)+/-SEM for NKA was -12.72+/-0.63 at the beginning and -9.77+/-0.49 at the end of the period (p<0.0001), while under placebo, it was -12.16+/-0.82 and 12.19+/-0.51 respectively (NS). At the active treatment period, the mean log2 PC20 for methacholine was -5.25+/-0.40 at the beginning and -4.22+/-0.31 at the end of the period (p=0.012), while under placebo, it was -5.47+/-0.47 and -5.24+/ 0.42 respectively (NS). The reduction in response to NKA was significantly larger than that for methacholine. A 2-week course of an inhaled steroid reduces bronchial responsiveness to neurokinin A, an effect more pronounced than the reduction in bronchial responsiveness to methacholine. PMID- 12108886 TI - Cure in prostate cancer patients. PMID- 12108887 TI - The sentinel node in colorectal carcinoma. Mapping technique, pathologic assessment, and clinical relevance. AB - One of the most important prognostic factors in colorectal cancer is the presence or absence of regional lymph node metastases. In many instances, micrometastatic disease may not be found on routine pathologic analysis using hematoxylin and eosin staining, but may be discovered only with immunohistochemical methods or polymerase chain reaction assay. Lymphoscintigraphy with biopsy of the sentinel nodes, defined as the first nodal basin in the drainage pathway of a tumor, was developed to provide accurate staging without the morbidity associated with the classic lymph node dissections performed for melanoma or breast cancer. This concept has recently been applied to colorectal cancers, but the method used is unique because oncologic principles of resection are still adhered to for the primary tumor along with en bloc resection of the locoregional mesenteric nodes, some of which are sentinel nodes. Sentinel nodes are ideal for sensitive pathologic techniques of detecting micrometastatic disease, as they often reflect the status of the entire locoregional nodal basin. Gross metastatic nodes reveal significant prognostic information and guide the use of adjuvant therapy in affected patients. However, the detection of micrometastatic disease in sentinel nodes by sensitive pathologic methods has not been proven to result in poor prognosis or benefit from adjuvant therapy for colorectal cancer. PMID- 12108888 TI - Controversies in early-stage Hodgkin's disease. AB - Early-stage Hodgkin's disease accounts for approximately 60% of all cases of the illness. Because of its excellent cure rate (80% to 90%) and high salvage rate, it is difficult to demonstrate survival advantages for different management options. Currently, there is no consensus as to the optimal staging and treatment strategy for early-stage Hodgkin's disease. With the increasing recognition of the late consequences of Hodgkin's disease therapy, the focus of recent clinical trials has been on exploring treatment modifications to reduce these late effects. Areas of controversy that are being explored include extent of staging work-up, radiation field size and dose (and as part of combined-modality therapy), optimal chemotherapy regimen, number of cycles of chemotherapy, and limited- vs extended-field radiation therapy and dose. In addition, several studies are investigating the feasibility of chemotherapy alone in early-stage patients. Along with the evaluation of modified treatments, long-term follow-up efforts should continue in patients who are cured in order to confirm the long term safety of such therapies. PMID- 12108889 TI - Clinical trials referral resource. Targeted therapy in squamous cell cancers of the head and neck. PMID- 12108890 TI - Current management of childhood ependymoma. AB - Radiation therapy has long been a mainstay in the treatment of ependymoma. Concerns about the long-term effects of radiation therapy have made many parents and caregivers wary of this treatment modality. However, with the advent of conformal radiation and evidence supporting its use in younger children (ie, < 3 years old), the standard of care for childhood ependymoma is rapidly evolving to include immediate postoperative radiation therapy for all pediatric patients. The role of chemotherapy in the treatment of ependymoma has diminished recently because (1) chemotherapy fails to delay radiation therapy for a meaningful period of time; (2) tumors that progress during chemotherapy do not respond as well to subsequent irradiation; and (3) the combination of chemotherapy and irradiation does not improve overall survival. However, chemotherapy may make residual tumor more amenable to a second resection. Fewer than 50% of pediatric patients with ependymoma undergo complete resection before receiving radiation therapy. Because the extent of resection is one of the most important prognostic factors in the treatment of this disease, increasing the rate of complete resections is a significant means of increasing long-term survival. By incorporating current concepts of ependymoma, a more uniform approach to the treatment of this disease can be developed. In addition, by combining the best available means of detecting and managing side effects, the future for pediatric patients with ependymoma remains optimistic. This review presents historical and current practices used to treat ependymoma, and is intended to provide an information framework for caregivers so that they can assist parents in the decision-making process. PMID- 12108891 TI - AIDS malignancies in the era of highly active antiretroviral therapy. AB - The introduction of highly active antiretroviral therapy (HAART) has had a dramatic impact on the morbidity and mortality of individuals living with human immunodeficiency virus (HIV). In addition to contributing to dramatic declines in the incidence of several opportunistic infections, HAART is affecting the incidences of several acquired immunodeficiency syndrome (AIDS)-defining malignancies. The incidence of Kaposi's sarcoma (KS) and primary central nervous system lymphoma (PCNSL) has dropped precipitously since the introduction of HAART in 1995. Systemic non-Hodgkin's lymphoma (NHL) appears to be declining in incidence as well, but to a lesser degree than KS and PCNSL. On the contrary, the incidence of invasive cervical carcinoma has not significantly changed in the HAART era. The impact of HAART on the epidemiology of other HIV-associated malignancies, including Hodgkin's disease and anal carcinoma, remains unclear. Data regarding the impact of HAART on the natural history and treatment outcomes of HIV-associated malignancies are limited. The possibility of direct and indirect roles of HIV in HIV-related carcinogenesis suggests that antiretroviral therapy may be an important component of the treatment strategy for several HIV related malignancies. Patients with HIV-NHL treated with HAART in addition to chemotherapy experience fewer intercurrent opportunistic infections. Furthermore, the simultaneous administration of HAART and chemotherapy does not appear to significantly increase toxicity. Whether the combination of HAART and standard therapy results in improved survival remains uncertain. This two-part article, which began in the April 2002 issue, analyzes the impact of HAART on the incidence, clinical course, and outcomes of each of the AIDS-related malignancies. PMID- 12108892 TI - Oral complications of cancer therapy. AB - The mouth is a frequent site of complications arising from drug or radiation cancer therapy, with mucositis, xerostomia, osteoradionecrosis, and local infections being the most common. From the stand-point of dose limitation, treatment breaks, quality of life, and health economic outcomes, mucositis is the most significant acute oral toxicity. Xerostomia, a chronic side effect of radiation, involves the salivary gland tissue, and results in changes in taste, tissue resilience, and an increased risk of caries and periodontal disease. While the incidence of osteoradionecrosis seems to be decreasing, the chronicity and symptoms of this festering bony condition are especially difficult for patients. Local oral infections resulting from the overgrowth of opportunistic organisms or the activation of latent viruses are so common as to warrant a prophylactic approach in many cases. A surge of investigational interest has been directed at understanding the mechanisms of these stomatotoxicities and at developing treatment strategies to combat them. PMID- 12108893 TI - Docetaxel in the management of advanced or metastatic urothelial tract cancer. AB - Phase II studies of single-agent docetaxel (Taxotere) yielded promising results in advanced or metastatic transitional cell carcinoma (TCC) of the urothelium. Antitumor responses have been demonstrated in previously treated and chemotherapy naive TCC patients, as well as in a subgroup of patients with renal impairment unable to receive traditional cisplatin-based regimens. Investigations of docetaxel-containing doublet and triplet drug combinations in the first-line setting have been pursued. When combined with cisplatin in previously untreated patients, response rates of 52% to 60% have been achieved, with median overall survival of 8 to 13.6 months. Triplet drug combinations of the docetaxel/platinum base have been investigated, when the addition of an anthracycline (doxorubicin or epirubicin [Ellence]) or of gemcitabine (Gemzar) has proven feasible, and has resulted in favorable efficacy findings. Non-platinum-containing regimens such as docetaxel/gemcitabine are also being studied. Randomizedphase III trials of a docetaxel-containing regimen in comparison with established regimens are ongoing, and additional studies are warranted to determine if overall long-term survival of TCCpatients can be improved. PMID- 12108894 TI - The current status of docetaxel for metastatic breast cancer. AB - Docetaxel (Taxotere) has been intensively investigated for the treatment of metastatic breast cancer, where it has proved to be one of the most active agents. Initial phase II studies in anthracycline-resistant metastatic breast cancer demonstrated impressive response rates that have been confirmed in phase III randomized trials. Docetaxel remains the only single agent to demonstrate a survival benefit in anthracycline-resistant patients. More recently, the combination of docetaxel with capecitabine (Xeloda) has demonstrated additional improvement in survival over docetaxel alone in a randomized phase III trial. In patients previously treated with an alkylating agent, docetaxel is the only single drug to demonstrate improved efficacy over doxorubicin in a randomized trial. Docetaxel has been investigated in combination with the anthracyclines doxorubicin and epirubicin in randomized trials. The docetaxel-containing regimens have consistently demonstrated improvement over the non-docetaxel containing regimens. The efficacy and safety of weekly docetaxel has extended the line of investigation for combinations with agents normally administered on a weekly basis, such as vinorelbine [Navelbine], gemcitabine [Gemzar], and trastuzumab [Herceptin], with promising findings. In addition, the results of the triple-drug combination of docetaxel, a platinum salt (cisplatin or carboplatin), and trastuzumab have resulted in impressive response rates and time to progression in a population of metastatic breast cancer patients with HER2/neu positive tumors. The consistent demonstration of a high level of efficacy with manageable toxicity ensures the continued widespread investigation of docetaxel in metastatic breast cancer. PMID- 12108895 TI - Integration of docetaxel into adjuvant breast cancer treatment regimens. AB - Adjuvant chemotherapy is an integral component of the multidisciplinary curative treatment of primary breast cancers. The experience of the last 3 decades indicates that anthracycline-containing regimens provide the most effective cytotoxic treatment for this purpose. The development of the taxanes over the past 10 years represents substantial progress, and taxane-containing regimens are widely used for the treatment of metastatic breast cancer. Most ongoing clinical trials of adjuvant chemotherapy include a taxane-related question. The docetaxel (Taxotere)-containing regimens were developedfollowing one of three important strategies: (1) the sequential administration of docetaxel to existing, commonly used combinations; (2) the simultaneous addition of docetaxel to an existing regimen; and (3) the substitution of docetaxelfor one of the drugs included in standard combinations. These three approaches are under intense investigation by large multicenter, multinational clinical trials. The results of these phase III, prospective, randomized trials will establish the contribution of docetaxel to the curative treatment of breast cancer and will determine the optimal method of incorporating this drug into standard adjuvant therapy. PMID- 12108896 TI - Primary chemotherapy with docetaxel for the management of breast cancer. AB - Several clinical trials have explored the efficacy of docetaxel (Taxotere) as primary chemotherapy for breast cancer. Docetaxel has been evaluated as single agent therapy, sequentially as a single agent following anthracycline-containing regimens, and in combination with anthracyclines, cisplatin, and trastuzumab (Herceptin) in patients with high-risk early breast cancer. Two large, randomized phase III trials have demonstrated significant improvements in clinical and pathologic response rates with the sequential addition of docetaxel to an anthracycline-containing preoperative regimen. A trial conducted in the United Kingdom demonstrated that docetaxel sequential to CVAP (cyclophosphamide [Cytoxan, Neosar], vincristine, doxorubicin [Adriamycin], prednisolone) produced a higher overall clinical response rate (94% vs 66%, P = .001) and pathologic complete response rate (34% vs 18%) compared to additional cycles of CVAP as primary chemotherapy. This translated into a survival advantage for docetaxel treated patients, whose 3-year disease-free and overall survival were significantly improved (97% vs 84%; 90% vs 77%, P = .03). The results of the National Surgical Adjuvant Breast and Bowel Project (NSABP) protocol B-27 demonstrated that primary doxorubicin/cyclophosphamide followed by docetaxel significantly increased the clinical complete response (65% vs 40%, P < .001) and pathologic complete response rates (25.6% vs 13.7%, P < .001) and decreased the rate of positive axillary nodes (40.5% vs 48.5%, P = .01). Final analysis of NSABP B-27 may also potentially demonstrate improved disease-free and overall survivals. Additional phase II and phase III randomized trials have compared docetaxel/anthracycline combinations with standard anthracycline-based regimens. Preliminary results have shown that incorporation of docetaxel can improve the rate of breast preservation surgery and the overall clinical and pathologic complete response rates. PMID- 12108897 TI - Docetaxel for previously treated non-small-cell lung cancer. AB - Two phase III trials were conducted using docetaxel (Taxotere), administered every 3 weeks, as second-line treatment of non-small-cell lung cancer (NSCLC) in patients previously treated with platinum-based chemotherapy. In the TAX 317 trial, 204 patients were randomized to receive either docetaxel (49 received 100 mg/m2 and 55 received 75 mg/m2) or best supportive care (100 patients). Median survival was 7.5 months with docetaxel at 75 mg/m2 (D75) vs 4.6 months for best supportive care (P = .010); and 1-year survival was 37% for D75 vs 11% for best supportive care (P = .010). Quality-of-life analysis also showed statistically significant improvement in disease-related symptoms with docetaxel vs best supportive care. In the TAX 320 study, 373 patients were randomized to receive docetaxel at 100 mg/m2 (D100), docetaxel at 75 mg/m2 (D75), or a control arm of either vinorelbine (Navelbine) or ifosfamide (Ifex) (V/I). Partial response rates were 11.9% with D100 and 7.5% with D75 vs 1% with V/I (P values: .001 [D100] and .036 [D75]). Median response duration was over 7 months. One-year survival was 32% with D75 vs 19% in V/I (P = .025). Prior paclitaxel exposure had no bearing on the response rate and survival advantage of second-line treatment with docetaxel. Response rates to docetaxel were equivalent in the cohort of patients who had received prior paclitaxel (10.5%) and the group ofpatients who had not received prior paclitaxel (8.5%). The 1-year survival rates for patients with no prior paclitaxel therapy were 33% (D75) vs 20% (V/I); and the 1-year survival rates for patients who had received prior paclitaxel were 30% (D75) vs 17% (V/I). In conclusion, two large randomized phase III trials of second-line chemotherapy for NSCLC have shown significant differences favoring docetaxelfor response rate, time to progression, survival, and quality of life. Prior paclitaxel did not decrease the likelihood of response to docetaxel, nor did it lessen the survival advantage seen with docetaxeL Docetaxel offers a clinically meaningful benefit in this setting, with manageable toxicity. Based upon the observed response rates, survival, impact on quality of life, and toxicity profile, the optimal dose of docetaxel in this pretreated population is 75 mg/m2 every 3 weeks. PMID- 12108898 TI - Docetaxel for locally advanced or metastatic non-small-cell lung cancer. Current data and future directions as front-line therapy. AB - Docetaxel (Taxotere) has shown activity both as a single-agent and in combination with multiple other cytotoxic agents in the front-line therapy of advanced, metastatic non-small-cell lung cancer (NSCLC). A randomized, phase III trial demonstrated a survival advantage for docetaxel over best supportive care in the front-line setting, with docetaxel achieving a 2-year survival of 12% vs 0% for best supportive care. Combinations of docetaxel with the platinum agents have been the most extensively studied in the front-line setting and have produced notably high response rates and encouraging median survivals. When compared to the paclitaxel/cisplatinum combination in a large, phase III randomized trial, the combination of docetaxel and cisplatin resulted in similar response, median survival, and 1-year survival rates. Another randomized phase III trial compared docetaxel/platinum combinations to the US Food and Drug Administration (FDA) approved vinorelbine (Navelbine)/cisplatin regimen. The docetaxel/cisplatin combination produced a superior overall survival, 2-year survival, and overall response rates compared to vinorelbine/cisplatin. The combination of docetaxel and carboplatin (Paraplatin) demonstrated similar survival and response, and was associated with quality-of-life benefits over the vinorelbine/cisplatin arm. Docetaxel has been successfully combined with gemcitabine in multiple trials with impressive response and survival rates, and an acceptable toxicity profile. A large phase IIb trial demonstrated therapeutic equivalence and lesser toxicities for the docetaxel/gemcitabine combination compared to the combination of docetaxel and cisplatin. The docetaxel/gemcitabine combination therefore represents a viable nonplatinum regimen for first-line treatment of NSCLC. Other combinations that have been tested include docetaxel with vinorelbine, and docetaxel with irinotecan (CPT-11, Camptosar). Docetaxel is active in NSCLC and should be investigated further in combination with novel molecularly targeted drugs such as tyrosine kinase inhibitors, andfarnesyl transferase inhibitors. PMID- 12108899 TI - Docetaxel in the integrated management of prostate cancer. Current applications and future promise. AB - Docetaxel (Taxotere)-based regimens can be included among the most effective treatment options for the management of patients with advanced, androgen independent prostate cancer. Results with docetaxel as a single agent and in combination regimens with estramustine (Emcyt) have consistently achieved a palliative response, reduced serum PSA levels by > or = 50%, and produced objective responses in patients with measurable disease. In addition, encouraging survival data have been demonstrated in several phase II trials. The ability to administer docetaxel on a weekly basis has substantially enhanced research efforts for treatment in prostate cancer patients. The results of ongoing phase III randomized trials evaluating docetaxel regimens in androgen-independent prostate cancer are eagerly awaited for their potential to definitively demonstrate a beneficial impact on overall patient survival. Docetaxel-containing regimens are likely to demonstrate a substantial role in the management of early stage prostate cancer patients in the adjuvant and neoadjuvant settings, where clinical investigations are under way. In addition, study results from ongoing trials that integrate docetaxel with hormonal therapies for patients with biochemical recurrence following definitive local treatments will be important in refining the future role of chemotherapy for prostate cancer in general. The preliminary findings from studies conducted with docetaxel are encouraging and await final analysis. Finally, preliminary results from studies exploring combination regimens of docetaxel and novel agents that possess completely different mechanisms of action (eg, proapoptotic agents, angiogenesis inhibitors, and vitamin D analogs) have demonstrated the regimens to be feasible and safe, with promising early response data. These types of investigational studies will likely occupy a dominant position in future research initiatives for patients with advanced prostate cancer. PMID- 12108900 TI - Docetaxel in the treatment of ovarian cancer. AB - Docetaxel (Taxotere) has extended the armamentarium of agents with significant activity in the treatment of ovarian cancer. As a single agent in advanced ovarian cancer patients previously treated with a platinum agent, docetaxel at 100 mg/m2 every 3 weeks yields a 30% overall response rate and a 6-month duration of response. In vitro data demonstrate a lack of complete cross-resistance between docetaxel and paclitaxel. As a result, antitumor activity has also been demonstrated in patients refractory to a paclitaxel-containing regimen. In both platinum- and paclitaxel-pretreated patients, the highest response rates were obtained in patients with the longest interval of time since receipt of prior chemotherapy. Docetaxel has been successfully combined with the platinum salts for the first-line treatment of ovarian cancer patients. In combination with cisplatin, response rates of 69% were reported. In an effort to minimize hematologic toxicities and asthenia associated with the cisplatin/docetaxel combination, investigators have substituted carboplatin (Paraplatin) for cisplatin. Several phase II studies and the Scottish Randomized Trial in Ovarian Cancer (SCOTROC), a large phase III randomized trial, of the docetaxel/ carboplatin combination have been completed. The most frequent toxicity noted is neutropenia, which is generally of brief duration, predictable, and manageable. The docetaxel/carboplatin combination has a notably low rate of neurotoxicity. Therefore, the SCOTROC comparative trial, demonstrating equivalent overall response rates and progression-free survival rates, suggests that the docetaxel/carboplatin combination may represent a new alternative to paclitaxel/carboplatin as first-line chemotherapy for advanced ovarian cancer. PMID- 12108901 TI - The role of docetaxel in the management of squamous cell cancer of the head and neck. AB - The activity of docetaxel (Taxotere) as a single agent (overall response rates, 24%-45%) in the treatment of patients with recurrent squamous cell cancer of the head and neck has resulted in the investigation of docetaxel-based doublet and triplet combinations in both the recurrent and neoadjuvant settings. When combined with cisplatin, with or without fluorouracil (5-FU), in the treatment of recurrent disease, response rates of 33% to 44% have been observed for docetaxel, with median survival ranging from 9.6 to 11 months. In the neoadjuvant setting, response rates have been typically greater than 90%, with promising disease-free and overall survival results. Randomized trials are now under way to assess the value of docetaxel-based therapy relative to that of the standard cisplatin/5-FU combination in both the neoadjuvant and recurrent settings. Preclinical data indicate that docetaxel is a potent radiosensitizer and its initial evaluation with concurrent radiation in patients with locally advanced unresectable squamous cell cancer of the head and neck suggests feasibility. Phase II evaluation of this approach is in progress. PMID- 12108902 TI - Docetaxel for gastric and esophageal carcinomas. AB - Docetaxel (Taxotere) has been successfully investigated in the therapy for advanced gastroesophageal tumors as both a single agent and in combination regimens. As a single agent, phase II study results demonstrate an overall response rate of 17% to 24%, with occasional complete responses in a disease in which complete responses are rare. These figures classify docetaxel among the most active agents for the disease. Further research initiatives in gastric cancer have evaluated the combined use of docetaxel with traditionally established agents, such as cisplatin and fluorouracil (5-FU). The rationales for the combined use of docetaxel with these agents include the in vitro demonstration of a lack of complete cross-resistance and nonoverlap-ping side effect profiles. Phase II study results of docetaxel-based combinations demonstrate high overall response rates and progression-free survival, comparable with results obtained with established three- and four-drug regimens. Therapy is generally well tolerated, with a predominant toxicity of hematologic neutropenia. Docetaxel-based combination regimens are currently undergoing evaluation in randomized phase III trials in comparison with established standard regimens. While previous combination chemotherapy regimens have failed to improve survival over single-agent therapy, the aim for incorporation of docetaxel with other active agents is to improve palliation and possibly survival of patients with gastric cancer. PMID- 12108903 TI - The current status of docetaxel in solid tumors. An M. D. Anderson Cancer Center Review. AB - In less than a decade, docetaxel (Taxotere) has progressed from initial studies in anthracycline-refractory metastatic breast cancer to several large, phase III randomized trials evaluating its efficacy as adjuvant, neoadjuvant, and first line therapy for metastatic breast cancer, non-small-cell lung cancer (NSCLC), and ovarian cancer. In other tumor types, including prostate, head and neck, gastric, and bladder cancer, ongoing phase III trials are comparing docetaxel containing regimens to previously established regimens. For the seven tumor types reviewed in this supplement, phase III study information for docetaxel or docetaxel-based combinations are presented. Impressive results have been consistently demonstrated in the trials reported to date. In early-stage and metastatic breast cancer, NSCLC, and ovarian cancer, randomized trials have shown that docetaxel-containing therapies are superior to or as effective as established standard chemotherapeutic regimens and are often associated with an improved safety profile. Trials of docetaxel as adjuvant and neoadjuvant therapy for breast cancer are under way or have been completed. Docetaxel has demonstrated a survival benefit in many settings that previously had not achieved such a benefit. For example, a survival benefit was demonstrated in anthracycline resistant breast cancer and second-line NSCLC cancer phase III comparative trials. PMID- 12108904 TI - Docetaxel and radiation as combined-modality therapy. AB - Combined-modality approaches for the treatment of non-small-cell lung cancer (NSCLC), head and neck cancer, and esophageal cancer offer survival benefits by improving locoregional control and treating micrometastatic disease. The taxanes are active, tolerable drugs in these solid tumors and have radiation-sensitizing activity. Docetaxel (Taxotere) has been studied in combination with radiation with favorable results. In phase II trials, docetaxel combined with radiation therapy resulted in response rates of up to 80%, with the most commonly used schedule being docetaxel at 20 to 30 mg/m2 per week with concomitant radiation administered at fractions of 1.8 to 2.0 Gy, 5 days a week over 5 to 6 weeks. Studies of docetaxel and platinum combinations have been conducted predominantly in patients with NSCLC. Early results show good activity and acceptable toxicity, with esophagitis or mucositis being dose-limiting. Doses of docetaxel at 20 mg/m2 per week combined with cisplatin at 25 mg/m2 or carboplatin (Paraplatin) at an area under the concentration-time curve (AUC) of 2 with concomitant radiation appear to be well tolerated and active. Future investigations, including phase III trials in patients with locally advanced NSCLC, are encouraged. Current trials are studying various design schedules, including induction chemotherapy with radiation followed by consolidation chemotherapy. PMID- 12108905 TI - Tei-index in symptomatic patients with primary and secondary mitral regurgitation. AB - Significant mitral regurgitation (MR) may result from primary valve dysfunction or develop secondary to ischemic or dilated cardiomyopathy. The index 'isovolumic contraction time and isovolumic relaxation time divided by ejection time' (ICT + IRT/ET, 'Tei-index') is a well established measure of global cardiac function in patients with dilated cardiomyopathy and cardiac amyloidosis. We sought to define the diagnostic value of the Tei-index in patients with significant MR of various origin. Sixteen asymptomatic control subjects (8 male (m)/8 female (f), age 62+/ 8 years, control group), 12 patients with primary MR (PMR) (mean grade 3.1+/-0.3, due to rupture of the chordae tendineae (n = 2), flail leaflet (n = 1), valve prolapse (n = 6) or rheumatic degeneration (n = 3), 6 m/6 f, age 58+/-18 years, NYHA class 2.5+/-0.3, PMR group) and 25 patients with secondary MR (SMR) (mean grade 3.1+/-0.3; due to ischemic (n = 14) or dilated cardiomyopathy (n = 10), 19 m/6 f, age 60+/-11 years, NYHA class 3.1+/-0.5, SMR group) underwent conventional two-dimensional (2D) and Doppler echocardiographic examination including measurement of the Tei-index. In the SMR group, left ventricular ejection fraction was reduced compared to the control and the PMR group (29+/-13% vs. 59+/ 8% and 59+/-8%, p < 0.001 for both comparisons). The E/A ratio was elevated in PMR and SMR groups in comparison to the control group (1.74+/-0.44 and 1.70+/ 0.45 vs. 1.09+/-0.28, p < 0.05). The Tei-index was easily and reproducibly measured in all study subjects. The mean value of the index was significantly elevated in the SMR group compared to control and PMR groups (0.87+/-0.3 vs. 0.42+/-0.07 and 0.38+/-0.05, p < 0.001). The difference between the control group and the PMR group did not reach statistical significance. In MR patients, receiver operating characteristic curve analysis for the Tei-index yielded an area under the curve of 0.96+/-0.03 for separating the PMR and the SMR group. Using a Tei-index > 0.51 as a cutpoint, SMR was identified with a sensitivity of 92% and a specificity of 88%. In MR patients, a significant correlation between left ventricular end-systolic volume and the Tei-index was observed (r = 0.71, p < 0.01). The Tei-index is a feasible and sensitive indicator of overall cardiac dysfunction in severely symptomatic patients with significant MR secondary to ischemic or dilated cardiomyopathy. The index is in the normal range in symptomatic patients with PMR and preserved systolic function. The Tei-index differentiates between patients with SMR and PMR and may be useful in the work-up of such patients. PMID- 12108906 TI - Three-dimensional echocardiographic measurement of left and right ventricular mass and volume: in vitro validation. AB - INTRODUCTION: Three-dimensional (3D) echocardiography has been shown to offer highly accurate measurements of left ventricular (LV) volume and mass. The present study evaluated the accuracy of 3D surface reconstruction by the piecewise smooth subdivision method in measuring volume and mass not only in the LV but also in the more complexly shaped right ventricle (RV). METHODS: 3D echo scans were obtained of in vitro LV's (n = 15) and RVs (n = 10). From digitized images, ventricular borders were traced and used in surface reconstructions. Mass and volume determined from the reconstructions were compared to true volume and mass determined prior to imaging. Additionally casts of two RVs were made and laser-scanned. Distances between the laser-identified points on the RV surface and the corresponding 3D echo reconstructions were measured. RESULTS: 3D LV volume agreed well with the true volume (y = 0.99x + 1.73, r = 0.99, SEE = 3.35 ml, p < 0.0001), as did 3D LV mass (y = 0.99x - 4.71, r = 0.99, SEE = 9.85 g, p < 0.0001). 3D RV volume overestimated true volume (y = 1.11x + 1.77, r = 0.99, SEE = 3.36 ml, p < 0.001) by 6.23+/-3.70 ml (p < 0.0001). 3D mass agreed well with RV mass (y = 0.78x + 17.32, r2 = 0.93, SEE = 3.54 g, p < 0.0001). 3D echo reconstructions matched the laser-scanned RV closely with residual distances of 1.1+/-0.9 and 1.4+/-1.2 mm, respectively. CONCLUSIONS: 3D echo using freehand scanning combined with surface reconstruction by the piecewise smooth subdivision surface method enables accurate determination of LV mass and volume, of RV mass and volume, and of the RV's complex shape. PMID- 12108907 TI - Carotid turbulent flow observed by convergent color Doppler flowmetry in silent cerebral infarction. AB - AIM: A newly developed convergent color Doppler (CCD) was used for evaluating the possible relationship of the flow dynamics of the internal carotid artery to silent cerebral infarction (SCI). METHODS: In 108 patients (65+/-8 years) with stroke risk factors, the CCD simultaneously images information, on both flow direction and Doppler signal energy. The relation between turbulent flow and the incidence of brain lesions of SCI as identified by magnetic resonance imaging was investigated in 212 vessels, excluding four occluded vessels. Percent area stenosis was measured as (vessel area - lumen area)/(vessel area) on cross sectional echo image of stenotic site. RESULTS: Incidence of turbulent flow in SCI patients with 50-70% or 70-90% stenosis was higher (76.5 or 59.1%) than that in non-SCI patients (17.4 or 33.3%, both p < 0.0001). CONCLUSION: Non-invasive assessment of flow dynamics by CCD imaging can be useful for proposing the early stages of brain damage even in patients free from neurological deficits. PMID- 12108908 TI - Long-term prognostic value of stress-redistribution-reinjection Tl-201 imaging in patients with severe left ventricular dysfunction and coronary artery bypass surgery. AB - OBJECTIVES: This study sought to evaluate the long-term prognostic significance of stress-redistribution-reinjection Tl-201 imaging in patients with severe left ventricular (LV) dysfunction and coronary artery bypass surgery. BACKGROUND: Preoperative stress-redistribution-reinjection Tl-201 imaging detects viable but asynergic segments which show functional improvement postoperatively and is considered as a valuable noninvasive method in selection of patients with severe LV dysfunction for revascularization. The long-term prognostic value of the reinjection technique remains unclear. METHODS: Fifty-two patients with severe LV dysfunction (mean ejection fraction (EF) 0.32+/-0.03) who underwent coronary artery bypass surgery in 1993-1994 were included in the study. Patients had follow-up 49+/-12 months. LV function was assessed by two-dimensional echocardiography. Perfusion was assessed by Tl-201 SPECT imaging and was graded on a four-point scale (0 = normal, 3 = absent uptake) using the 20 segment model. Perfusion index was derived by adding the score of all segments and dividing these by 20. Patients were divided into two groups. Group A comprised patients with seven and more dysfunctional viable myocardial segments. Group B included patients with less than seven dysfunctional but viable segments. RESULTS: Mean EF increased from 0.32+/-0.03 to 0.46+/-0.04. Mean perfusion index did not show a significant difference as a whole during follow-up compared to the early postoperative values (0.9+/-0.4 and 1.1+/-0.4, p = NS). When adequacy of revascularization was considered, the predictive value of a positive preoperative viability test for functional improvement was 82%. Nineteen cardiac events occurred in group B patients and six in group A patients: six deaths (four from cardiac and two from noncardiac causes), 13 myocardial infarctions (MI). Multivariate Cox survival analysis identified the number of viable segments detected preoperatively (chi2 = 7.2, p = 0.002), postoperative improvement in Tl uptake (chi2 = 6.6, p = 0.01) and functional improvement (chi2 = 5.3, p = 0.03) postoperatively as independent predictors of cardiac events. Preoperative EF and functional capacity were not associated with cardiac events in long-term prognosis. CONCLUSION: These data suggest that preoperative stress-redistribution reinjection Tl-201 imaging, specifically the number of viable segments detected preoperatively and postoperative improvement in Tl-201 uptake provide important long-term prognostic information in patients with severe LV dysfunction who had coronary artery bypass surgery. PMID- 12108910 TI - Strategies for cardiac CT imaging. AB - We review the scanning techniques for cardiac CT imaging with single slice and multislice scanners. Combined with prospective triggering for transaxial scanning and retrospective gating for helical scanning the potential advantages and the basic limitations are discussed. Based on those theoretical considerations, the major conclusion is that high resolution data sets with isotropic spatial resolution can be acquired with quadslice, spiral scanning, only. First clinical results support this conclusion. PMID- 12108909 TI - Measurement of left ventricular volume after anterior myocardial infarction: comparison of magnetic resonance imaging, echocardiography, and radionuclide ventriculography. AB - We have compared echocardiography (echo) and radionuclide ventriculography (RNV) with magnetic resonance imaging (MRI) for the measurement of left ventricular (LV) volume and ejection fraction. Seventy asymptomatic patients were studied up to 12 days after first Q wave anterior myocardial infarction and again after 6 months. Each patient had LV volume measured by all three techniques within 24 hours of each other on each occasion. LV end-systolic and end-diastolic volume index (LVESVI and LVEDVI) and LV ejection fraction (LVEF) were measured using the modified Simpson formula (echo), a counts-based method (RNV), and a multislice area summation method (MRI). Radionuclide volumes were measured both with and without correction for attenuation of isotope. Echocardiography overestimated LV volume compared with MRI. Mean (SD) differences (echo-MRI) were: LVEDVI + 10.6 ml/m2 (16.8), LVESVI + 13.7 ml/m2 (12.9), LVEF -8.5% (11.2). RNV underestimated both volume and ejection fraction compared with MRI. Mean differences (RNV-MRI) were: LVEDVI -25.4 ml/m2 (23.8), LVESVI -5.0 ml/m2 (18.6), LVEF -13.8% (10.4). Variability in the difference between echo and MRI and between RNV and MRI was very similar for LVEF (coefficient of variation 23.9% echo, 22.2% RNV) but there was greater variability in the radionuclide than the echo measurements of absolute volume. Variability of the radionuclide measurements was reduced by not correcting for attenuation, and this finding may improve the radionuclide technique for serial measurements of percentage change in volume. Long-term inter study reproducibility of MRI for LVEF (coefficient of reproducibility) was 10.9%, for echo it was 10.6%, and for RNV it was 14.6%. We conclude that measurements of LV volume depend on the method used and are not interchangeable. Echocardiography agrees more closely with MRI than RNV for the measurement of absolute volume, but the two techniques are similar for the measurement of LVEF. PMID- 12108911 TI - How good are experienced cardiologists at predicting the hemodynamic severity of coronary stenoses when taking fractional flow reserve as the gold standard. AB - BACKGROUND: Coronary angioplasty should be based on documented ischemia. However, in daily clinical practice the indication for angioplasty is often based on eyeball assessment of the severity of the stenosis. This study was performed to assess the accuracy of eyeball estimation of coronary stenosis when taking functional flow reserve (FFR) as gold standard. METHODS: Study lesions were where no mutual agreement on the severity of the stenosis was obtained. The procedure consisted of a repeat control angiogram, FFR measurement and in case of FFR<75% percutaneous coronary intervention. The eyeball assessment of the stenosis was written down before further execution of the procedure. FFR was measured with a pressure monitoring guide. Maximal myocardial hyperemia was induced by intravenous adenosine infusion. RESULTS: Fifty-two patients were studied. Agreement between eyeball assessment and FFR existed in a total of 36 cases (69.2%). Over estimation of hemodynamic severity occurred in six cases (11.5%) and under estimation in 10 cases (19.2%). Consequently, the positive predictive value of eyeball assessment for pressure-derived FFR was 63% and the negative predictive value 76%. CONCLUSION: The assessment of the hemodynamic severity of intermediate coronary stenosis should not be based on eyeball assessment even by experienced interventional cardiologists. PMID- 12108912 TI - The 6sigma quality approach for quantitative arteriography performance improvement. AB - Quantitative angiography is a medical application where the user requires tools whose operational results in term of accuracy, precision, robustness and reliability must be extensively assessed and validated for clinical use. In this study, the 6sigma methodology has been applied to analyze the performances of the General Electric QA software method. In particular, the catheter calibration procedure was identified as the weakest function in term of sensitivity to procedure parameters, like point-spread-function, field-of-view, catheter dimensions. It was therefore improved by following a design for 6sigma scheme, and the main parameters that govern the QA accuracy and precision were put under quantifiable control. PMID- 12108913 TI - Noninvasive prediction of coronary artery disease progression by comparison of serial exercise electrocardiography and dipyridamole stress echocardiography. AB - BACKGROUND: The possibility of noninvasive prediction of angiographically assessed coronary artery disease (CAD) progression by comparison of serial studies of exercise electrocardiography (EET) and dipyridamole stress echocardiography (DET) is not known. AIM: To assess the relative value of EET and DET in predicting angiographically assessed progression of CAD. METHODS: From the Institute of Clinical Physiology, National Research Council, Pisa Italy stress echo data bank (1983-1998), we selected 46 patients with two repeated EET, DET and coronary angiography (CA) in two different hospital admissions (46+/-30 months). A priori, angiographic progressors were defined as any stenosis progression to occlusion and/or any stenosis >30% with >20% stenosis progression measured by visual and quantitative CA. EET progressors were defined as a previous negative test becoming positive or as a positive test with decrease in ischemic threshold response in the second test. DET progressors were defined as previous negative test becoming positive or as a positive test with a more severe ischemic response in the second test. RESULTS: Angiographic progressors were 31/46 patients (67%) and angiographic nonprogressors were 15/46 (33%). When angiography was taken as the gold standard, there were no differences in sensitivity for EET and DET (87 vs. 87%). Specificity was significantly higher for DET (93 vs. 40% p =< 0.001). By kappa statistics DET had a good concordance (kappa = 0.768) and EET a poor concordance (kappa = 0.299) with angiographic progression. CONCLUSION: DET is more accurate than EET at predicting angiographically assessed CAD progression. PMID- 12108914 TI - Clinical research: the best or worst of times? PMID- 12108915 TI - Reporting of adverse event data in hematopoietic stem cell transplantation clinical trials involving investigational new drugs or devices: a report from the William Guy Forbeck Foundation 2001 focus meeting on clinical trials in hematopoietic stem cell transplantation. AB - The William Guy Forbeck Foundation was established in 1984 in memory of William Guy Forbeck, an 11-year-old boy who died of neuroblastoma. The objectives of the Forbeck Foundation are to promote advances and shorten the research timetable in the field of oncology, particularly pediatric oncology. The Foundation's centerpiece activity is an annual scientific forum held at Hilton Head Island, South Carolina, where 12 to 15 leading scientists from a variety of disciplines associated with a specific topic are invited to participate in a private "think tank" environment, where they can freely exchange ideas in the hope of building on each other's knowledge and experience. Additionally, the Foundation sponsors grants for Focus Meetings, which are designed to give other researchers an opportunity to conduct their own meetings along the lines of the Foundation's annual forum. The idea for this Focus Meeting was born during the 2000 Annual Forbeck Forum in South Carolina, which considered the current status of allogeneic hematopoietic stem cell transplantation (HSCT). Participants considered various obstacles to conducting clinical trials in this area and decided to bring together experts from academia, industry, and government to discuss ways in which these obstacles might be overcome. Topics included efficient clinical trial designs, issues of monitoring and reporting adverse events, and appropriate definitions and grading systems for transplantation specific outcomes. This article summarizes the issue of adverse event reporting in HSCT. PMID- 12108916 TI - Leucyl-leucine methyl ester-treated haploidentical donor lymphocyte infusions can mediate graft-versus-leukemia activity with minimal graft-versus-host disease risk. AB - L-leucyl-L-leucine methyl ester (LLME) prevents GVHD in several animal models by depleting dipeptidyl peptidase I (DPPI)-expressing cytotoxic cellular subsets. However, clinical application has been hampered by difficulties in stem cell engraftment following treatment of donor bone marrow inocula with LLME at the concentrations necessary to purge DPPI-expressing T-cells. Noting that T-cells can mediate graft-versus-leukemia (GVL) responses via both perforin (usually co expressed in cytotoxic granules with DPPI) and Fas ligand pathways in a murine model, we hypothesized that LLME might be useful for treatment of delayed DLIs for potential GVL activity with a decreased risk of GVHD induction. In regard to the clinical setting, the ex vivo use of LLME for this purpose would circumvent any toxicity issues for donor stem cells, because by that time patients would have already achieved successful engraftment. For our preclinical studies, we used the haploidentical C57BL/6 (B6) (H2b) --> ((B6 x DBA/2)F1 (H(2b/d)) murine model with lethally irradiated hosts that had received transplants of T-cell depleted bone marrow cells and were challenged with the MMD2-8 myeloid leukemia line (H2d) of DBA/2 origin. A DLI of LLME-treated donor splenocytes, from B6 mice presensitized to recipient alloantigens, was administered in varying doses 14 days post-marrow transplantation, and the potential for both GVHD and GVL activity was assessed. All mice that received any dose of LLME-treated DLI survived indefinitely, without evidence of cachexia nor B-cell hypoplasia, in contrast to the severe and lethal GVHD induced by mock-treated DLI. Histological analysis largely correlated with the symptomatic findings and revealed no GVHD like lesions in the spleens of LLME-treated DLI recipients, although some mice displayed various degrees of hepatic mononuclear infiltration. Most notably, mice given LLME-treated DLI also experienced DLI dose-dependent increases in survival against the challenge with the MMD2-8 leukemia. LLME-treated splenocytes remained immunocompetent, as these cells could proliferate in response to mitogens and to restimulation with ovalbumin when used as a recall antigen. In conclusion, LLME treated DLI possesses immune potential and, in particular, GVL activity without inducing clinically evident GVHD. PMID- 12108917 TI - Administration of either anti-CD40 or interleukin-12 following lethal total body irradiation induces acute lethal toxicity affecting the gut. AB - Interleukin (IL)-12 and antibodies against CD40 have demonstrated antitumor effects in a variety of in vivo model systems. However, both agents can also mediate significant toxicities either when used following lethal TBI or when administered in combination with other agents such as IL-2. In this study, we assessed the effects of anti-CD40 monoclonal antibody (MoAb) and IL-12 in lethally irradiated mice. Acute lethal toxicity was observed following the administration of either 10 microg anti-CD40 MoAb (FGK45) or 0.5 microg of recombinant murine (rm)IL-12 that resulted in 100% mortality of all mice within 4 to 6 days. Histological evaluation revealed destruction of the normal gut architecture in both anti-CD40 MoAb- and rmIL-12-treated mice. Analysis of serum cytokine levels in the lethally irradiated mice receiving anti-CD40 MoAb demonstrated a marked increase of interferon (IFN)-gamma and IL-12 p40, whereas mice receiving rmIL-12 demonstrated a marked increase of IFN-gamma. Lethally irradiated IL-12 p40 knock-out mice were resistant to anti-CD40-induced toxicity, suggesting that the lack of IL-12 p40 with no possibility of making functional IL 12 p70 is key for this toxic reaction. Similarly, lethally irradiated IFN-gamma knock-out mice were completely resistant to rmIL-12-induced toxicity, suggesting that IFN-gamma is a major player in IL-12-mediated toxicity. These results suggest that both anti-CD40 MoAb and rmIL-12 induce an acute fatal toxicity characterized by similar intestinal pathology and mediated in part by IFN-gamma. PMID- 12108918 TI - Dose-intensive therapy for extensive-stage small cell lung cancer and extrapulmonary small cell carcinoma: long-term outcome. AB - Treatment for extensive-stage small cell lung cancer (ES SCLC) or extrapulmonary small cell carcinoma (EPSC) is typically palliative. We set out to determine progression-free survival (PFS) and overall long-term survival of ES SCLC and EPSC patients, physiologically aged < or = 60 years, responding to first-line chemotherapy followed by high-dose combination alkylating agents with hematologic stem cell support. Patients in first-line chemotherapy response underwent stem cell collection (marrow, peripheral blood progenitor cells, or both) followed by high-dose therapy with 1 of 2 regimens: CBP (cyclophosphamide, cisplatin, and carmustine) or ICE (ifosfamide, carboplatin, and etoposide) with or without etanidazole. Involved-field radiotherapy was given to selected patients with oligometastatic disease distributed in sites allowing for reasonable radiation ports, and prophylactic cranial radiotherapy was given upon recovery to patients in complete response (CR) or near-CR. A total of 36 patients were treated. Of 29 patients with ES SCLC, 6 (21%) had achieved CR, 18 near-CR, and 5 partial response prior to high-dose therapy. Of 7 patients with EPSC, 3 (43%) had achieved CR, 3 had achieved near-CR, and 1 had progression of disease prior to high-dose therapy. Thirteen ES SCLC patients received high-dose CBP. Of the remaining 23 patients with SCLC or EPSC, 17 were treated with ICE and 6 with ICE plus etanidazole, a hypoxic cell sensitizer. Treatment-related mortality was 11% (4 of 36 patients). For all patients, the median event-free survival (EFS) was 5 months. The 2- and 5-year survivals after intensification were 12% (95% confidence interval [CI], 5%-31%) and 9% (95% CI, 3%-27%), respectively. Of the 30 patients in or near CR prior to high-dose therapy, 5 remain continuously progression-free (2 ES SCLC, 3 EPSC) for a median of 55 months (range, 1-96 months) after high-dose therapy. By multivariate analysis, factors associated with more favorable EFS were the use of a more aggressive induction regimen (ICE), and the EPSC histology. These factors were also associated with more favorable overall survival. Other factors associated with more favorable overall survival were the use of short induction therapy (< or = 4 cycles) and younger age (<50 years). Except for high-dose ICE with etanidazole, the use of high-dose systemic therapy in ES SCLC and EPSC was associated with low treatment-related morbidity and mortality over the past 5 years. Late complications were infrequent, and most patients returned to full-time work and activity, barring disease recurrence. Nonetheless, few patients with ES SCLC have progression-free long-term survival. We conclude that high-dose therapy is not indicated as an approach for ES SCLC, except as part of an investigative trial. Conversely, 3 of the 7 patients with EPSC remain relapse-free (range, 1-96 months), warranting further phase II evaluation of this approach in this population. PMID- 12108919 TI - Increased pulmonary toxicity results from a 1-day versus 2-day schedule of administration of high-dose melphalan. PMID- 12108920 TI - Preconditioning modulates susceptibility to ischemia-induced arrhythmias in the rat heart: the role of alpha-adrenergic stimulation and K(ATP) channels. AB - A new concept of cardioprotection based on the exploitation of endogenous mechanisms is known as ischemic preconditioning (IPC). It has been hypothesized that substances released during brief ischemic stress (e.g. catecholamines) stimulate the receptors and trigger multiple cell signaling cascades. Opening of ATP-sensitive K+ channels [K(ATP)] has been suggested as a possible final step in the mechanisms of protection. In this study, the role of adrenergic activation was tested in Langendorff-perfused rat hearts subjected to test ischemia (TI; 30 min occlusion of LAD coronary artery) by: 1) mimicking IPC (5 min ischemia, 10 min reperfusion) with short-term (5 min) administration of norepinephrine (NE, 1 microM), 15 min prior to TI; 2) blockade with beta- or alpha1-receptor antagonists, propranolol (10 microM) and prazosin (2 microM), respectively, applied 15 min prior to TI during IPC. The role of K(ATP) opening was examined by perfusion with a K(ATP) blocker glibenclamide (10 microM) during IPC. Both IPC and NE-induced PC effectively reduced the incidence of ventricular tachycardia (VT) to 33% and 37%, respectively, vs 100% in the non-PC controls, whereby ventricular fibrillation (VF) was totally abolished by IPC and markedly suppressed by PC with NE (0% and 10%, respectively, vs 70% in the non-PC hearts; P < 0.05). The severity of arrhythmias (arrhythmia score, AS) was also markedly attenuated by both interventions (IPC: AS 1.7 +/- 0.4; NE-PC: AS 1.8 +/- 0.3 vs AS 4.1 +/- 0.2 in the controls; P < 0.05). Protection was not suppressed by propranolol (VT 28%; VF 14%; AS 2.2 +/- 0.6), whereas prazosin reversed the protective effect of PC (VT 83%; VF 67%; AS 4.0 +/- 0.8). Antiarrhythmic protection afforded by NE-PC was abolished by pretreatment of rats with pertussis toxin (25 microg/kg, i.p.) given 48 h prior to the experiments. Glibenclamide did not suppress the IPC-induced protection. In conclusion, the sensitivity of the rat heart to ischemic arrhythmias can be modulated by IPC. Protection is mediated via stimulation of alpha1-adrenergic receptors coupled with Gi-proteins but glibenclamide-sensitive K(ATP) channels do not appear to be involved in the mechanisms of antiarrhythmic protection in this model. PMID- 12108921 TI - QT dispersion and T-loop morphology in late pregnancy and after delivery. AB - The aim of the study was to detect changes of both the QT dispersion and T-loop morphology resulting from the changed spatial position of the heart during pregnancy. Electrocardiographic and vectorcardiographic recordings were obtained from 37 healthy women 19-36 years old in the 36th to 40th week of physiological pregnancy and 2 to 6 days after delivery. The same recordings were obtained from 18 healthy women of the same age. The average QT dispersion (+/- S.D.) in normal subjects was significantly lower (34 +/- 12 ms) than in those in late pregnancy (73 +/- 18 ms) (P < 0.001). The average amplitude of T-loop (Ta) in women in late pregnancy was significantly (P < 0.001) smaller (532 +/- 98 microV) and the width of T-loop (Tw) was wider (21.24 +/- 11.48 deg) than in the control group (793 +/- 114 microV and 7.17 +/- 3.02 deg, respectively). The partial post-partum restoration of all parameters was not significant. In all groups, the QT dispersion was significantly correlated with Tw but not with Ta. According to these results we can conclude that the QT dispersion is an indirect reflection of the complete process of ventricular repolarization, reflected in the morphology of the T-loop. PMID- 12108922 TI - Constitutive inhibitory action of muscarinic receptors on adenylyl cyclase in cardiac membranes and its stereospecific suppression by hyoscyamine. AB - Muscarinic acetylcholine receptors in the heart have been shown to display agonist-independent spontaneous (constitutive) activity which causes changes in the opening of cardiac ion channels and in the activity of G proteins. We investigated whether an inhibition of the constitutive activity of muscarinic receptors induced by the binding of antagonist brings about a change in the synthesis of cyclic AMP in rat cardiac membranes, and whether the action ofthe antagonist is stereospecific. Atropine and S-(-)-hyoscyamine were indeed found to enhance the forskolin-stimulated synthesis of cyclic AMP in rat cardiac (both atrial and ventricular) membranes by up to 24%. The effect was stereospecific and the potency of R-(+)-hyoscyamine was 30 fold lower than that of the S-(-) enantiomer, confirming that the action of hyoscyamine is receptor-mediated. The effect did not depend on the presence of endogenous acetylcholine in the system used. The results strongly suggest that the adenylyl cyclase in the heart is exposed to continuous mild inhibition by constitutively active muscarinic receptors in the membranes of cardiomyocytes. PMID- 12108923 TI - Post-exercise decrease of plasma hyaluronan: increased clearance or diminished production? AB - The exercise-induced increase and post-exercise decrease of plasma hyaluronan concentration were studied in human subjects. Six well trained men performed incremental exercise until exhaustion (MAX), intensive (submaximal, SUB) and extensive exercise (moderate, MOD) on a bicycle ergometer, defined as work at 100, 77 and 50% of maximal oxygen consumption. Hyaluronan was analyzed using a high-sensitivity, proteoglycan-dependent time-resolved immunoassay and hemoglobin, hematocrit and plasma protein levels were assessed using standard laboratory procedures. Compared to resting control levels, the plasma hyaluronan concentration (pHA) increased (p < 0.05) by 76% (65.0 +/- 6.1 vs. 37.0 +/- 1.0 microg/l) during 15 min MAX, by 44% (56.4 +/- 2.6 vs. 39.2 +/- 3.8 microg/l) during 30 min SUB and by 27% (46.3 +/- 7.8 vs. 36.4 +/- 4.3 microg/l) during 90 min MOD. The increase with time averaged 4.03%.min(-1) during MAX, 1.35%.min(-1) during SUB and 0.35%.min during MOD. After exercise (15 and 30 min), pHA decreased by 43% below resting levels after MAX (p < 0.05) and by 36% after SUB, respectively. In conclusion, pHA steadily rose with time during physical exertion, with a non-linear increase of concentration/time slope with exercise intensity; second, the magnitude of the post-exercise pHA decrease was proportional to the exercise-induced pHA increase, suggesting elevated hyaluronan clearance with rising plasma levels after physical exertion. PMID- 12108924 TI - The effects of cardiopulmonary bypass with hollow fiber membrane oxygenator on blood clotting measured by thromboelastography. AB - In cardiac surgical patients we investigated the effects of cardiopulmonary bypass (CPB) with a hollow fiber membrane oxygenator on blood clotting measured by thromboelastography (TEG). We found only a minimal change in the strength of blood clot described either by the TEG parameter MA (maximum amplitude) or by the shear modulus G calculated from MA. After CPB there was also a significant tendency towards hypercoagulation as defined by shortened parameters R, K and increased a-angle. After comparison with published data obtained in cardiac surgical patients using a bubble oxygenator we conclude that currently used extracorporeal technology exerts a less negative influence on blood clotting than had been conceived previously. PMID- 12108925 TI - Purification and properties of ornithine carbamoyltransferase from loggerhead turtle liver. AB - Ornithine carbamoyltransferase has been purified from the liver of the loggerhead turtle Caretta caretta by a single-step procedure using chromatography on an affinity column to which the transition-state analogue, delta-N-(phosphonoacetyl) L-ornithine (delta-PALO), was covalently bound. The procedure employed yielded an enzyme which was purified 373-fold and was judged to be homogeneous by nondenaturing and sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS PAGE). The enzyme showed a specific activity of 224. The molar mass of the C. caretta enzyme was approximately 112 kDa, the single band obtained by SDS-PAGE indicated a subunit molar mass of 39.5 kDa; hence, the enzyme is a trimer of identical subunits. It catalyzes an ordered sequential mechanism in which carbamoyl phosphate binds first, followed by L-ornithine. The Michaelis constants were 0.858 mM for L-ornithine and 0.22 mM for carbamoyl phosphate, the dissociation constant of the enzyme-carbamoyl phosphate complex was 0.50 mM. PMID- 12108926 TI - Concentrations of free mg2+, pH and 31P MR metabolite ratios in calf muscles of healthy controls and patients with primary juvenile hypertension. AB - 31P MR spectroscopy was used to measure the signal intensity ratios of high energy metabolites for the calculation of free cytosolic magnesium concentration [fMg(2+)] and pH in the calf muscles of patients with primary juvenile hypertension and of healthy controls. Surface coil and spectroscopic imaging techniques were used. In patients with hypertension, the concentrations of [fMg(2+)] was 788 +/- 33 micromol/l and intracellular pH was 7.05 +/- 0.02; these values were not significantly different from the results obtained in healthy controls ([fMg(2+)], 776 +/- 21 micromol/l and pH, 7.06 +/- 0.01). Biochemical assays of magnesium in the serum (S-Mg) and in urine (DU-Mg) confirmed this finding. Significant differences in the relative signal intensities of high energy phosphates between patients with primary juvenile hypertension and healthy controls were observed: a) signal intensity ratios of PCr/Pi, PCr/PbetaATP, PDE/PbetaATP were increased, and b) Pi/PDe, Pi/PATP were decreased. The results were the same irrespective of whether the surface coil method or 31P spectroscopic imaging were employed. PMID- 12108927 TI - Improvement of the accuracy by the measurement of the electrical cell membrane parameters. AB - The electrical parameters of the cell membrane are mostly estimated employing ac methods. The measurement is based on the analysis of the current(s) flowing through an access resistance and the membrane. A current/potential transducer is used at the input of the device. The parameters of this transducer, especially its feedback capacity, degrades the accuracy of the measurement and hence diminishes the suppression of mutual influences of the individual parameters. The paper suggests a possible software correction and is supplemented by remarks for practical application. PMID- 12108928 TI - Augmentation of analgesic effect of ibuprofen by alprazolam in experimental model of pain. AB - The reports of analgesic effects of benzodiazepines are inconsistent. There is evidence of a hyperalgesic effect induced by activation of supraspinal GABAA receptors and an antinociceptive effect induced by activation of receptors located in the spinal cord (dorsal horns). The aim of the study was to discover whether the systemic administration of a benzodiazepine agent alprazolam increases the systemic analgesic efficacy of non-opioid analgesic ibuprofen. Experimental studies combining these agents have not yet been published. We used three experimental methods - writhing test (with acetic acid), tail-flick test and plantar test to assess analgesic action. The drugs were administered orally. Augmentation of the analgesic effect of ibuprofen by alprazolam was proved for the writhing test at a dose of 30 mg/kg of ibuprofen and alprazolam 1 mg/kg. The reaction time of the combination was significantly prolonged in comparison with ibuprofen alone. The results of the tail-flick test and plantar test were negative. The effect of ibuprofen was not enhanced by alprazolam in tests of acute thermal pain. Our results have demonstrated that the analgesic action of ibuprofen is only weakly enhanced by alprazolam. PMID- 12108929 TI - Are acute changes after status epilepticus in immature rats persistent? AB - Early consequences of lithium-pilocarpine convulsive status epilepticus (SE) were studied six days after this status had been induced in rat pups at the age of either 12 or 25 days. Studies of spontaneous EEG activity demonstrated the presence of epileptic phenomena (isolated spikes) in both hippocampus and cortex (cortical spikes were more expressed in the older group). There were no marked behavioral correlates of spikes and transition into the ictal phase was exceptional. The motor performance on a rotorod and a horizontal bar was the same in experimental and control rats of both ages. Behavior in the open field was changed in a reverse manner in the two age groups: the locomotor activity of rats with induced seizures at the age of 12 days was significantly lower than that of their control siblings, whereas animals undergoing status at the age of 25 days were hyperactive. In addition, they also exhibited increased exploratory activity (rearing) and their habituation to the open field was deranged. Nissl-stained brain sections demonstrated extensive brain damage in the older group in contrast to the negative findings in younger animals. EEG, behavioral and morphological changes induced by status epilepticus in developing rats persisted for 6 days after the status. They markedly differed according to the age of animals. PMID- 12108930 TI - Suramin affects capsaicin responses and capsaicin-noxious heat interactions in rat dorsal root ganglia neurones. AB - The effect of suramin, an inhibitor of G protein regulated signalling, was studied on the membrane currents induced by noxious heat and by capsaicin in cultured dorsal root ganglia neurones isolated from neonatal rats. Whole-cell responses induced by a heat ramp (24-52 degrees C) were little affected by suramin. The noxious heat-activated currents were synergistically facilitated in the presence of 0.3 microM capsaicin 13.2-fold and 6.3-fold at 40 degrees C and 50 degrees C, respectively. In 65% of neurones, the capsaicin-induced facilitation was inhibited by 10 microM suramin to 35 +/- 6% and 53 +/- 6% of control at 40 degrees C and 50 degrees C (S.E.M., n = 15). Suramin 30 microM caused a significant increase in the membrane current produced by a nearly maximal dose (1 microM) of capsaicin over the whole recorded temperature range (2.4-fold at 25 degrees C and 1.2-fold at 48 degrees C). The results demonstrate that suramin differentially affects the interaction between capsaicin and noxious heat in DRG neurones and thus suggest that distinct transduction pathways may participate in vanilloid receptor activation mechanisms. PMID- 12108931 TI - Visual event-related potentials to moving stimuli: normative data. AB - Visual cognitive responses (P300) to moving stimuli were tested in 36 subjects with the aim to find the normal range of P300 parameters. Concomitantly, the circadian intra-individual variability of the P300 was studied in a subgroup of 6 subjects. Visual stimuli consisted of either coherent (frequent stimulus) or non coherent motion (random stimulus). The oddball paradigm was applied for recording cognitive responses. P300 to rare stimuli had an average latency of 447.3 +/- 46.6 ms and amplitude of 12.9 +/- 6.0 microV. The average reaction time was in the range from 322 to 611 ms and there was no correlation between the reaction time and P300 latency. We did not find any significant circadian changes of the P300 parameters in the 6 subjects tested four times during the same day. Cognitive (event-related) responses (P300) displayed distinctly greater inter individual variability (S.D. of 50 ms) when compared with pattern-reversal and motion-onset VEPs (S.D. of 6.0 ms and 14 ms, respectively). For this reason, the clinical use of P300 elicited by this kind of visual stimuli seems to be rather restricted and the evaluation of its intra-individual changes is preferable. PMID- 12108932 TI - Model of spike propagation reliability along the myelinated axon corrupted by axonal intrinsic noise sources. AB - We investigated how selected electromorphological parameters of myelinated axons influence the preservation of interspike intervals when the propagation of action potentials is corrupted by axonal intrinsic noise. Hereby we tried to determine how the intrinsic axonal noise influences the performance of axons serving as carriers for temporal coding. The strategy of this coding supposes that interspike intervals presented to higher order neurons would minimally be deprived of information included in interspike intervals at the axonal initial segment. Our experiments were conducted using a computer model of the myelinated axon constructed in a software environment GENESIS (GEneral NEural SImulation System). We varied the axonal diameter, myelin sheath thickness, axonal length, stimulation current and channel distribution to determine how these parameters influence the role of noise in spike propagation and hence in preserving the interspike intervals. Our results, expressed as the standard deviation of spike travel times, showed that by stimulating the axons with regular rectangular pulses the interspike intervals were preserved with a microsecond accuracy. Stimulation with pulses imitating postsynaptic currents, greater changes of interspike intervals were found, but the influence of implemented noise on the jitter of interspike intervals was approximately the same. PMID- 12108934 TI - QUOROM and systematic reviews. PMID- 12108933 TI - Cytosolic free Ca2+ concentration in canine aortic endothelial cells lining the polyester arterial prosthesis. AB - A rise in baseline cytosolic free Ca2+ in canine vascular endothelial-like cells (VEC) lining the luminal surface of the polyester arterial prosthesis is described. In one, three and six month implantation experiments we employed six adult mongrel dogs, polyester arterial prostheses Arteknit Ra K, fluorescent Ca2+ indicator Fura-2 and digital imaging microscopy to study cytosolic free Ca2+ in cultured VEC. The electron microscopy scanning of the luminal surface in different regions of the graft were also performed. A rise in cytosolic free Ca2+ in the VEC lining the luminal surface of the prosthesis is probably the result of the immunologic reaction and mechanical stress which stimulate the proliferation activity of the endothelial cells. It seems that the baseline cytosolic free Ca2+ reflects the course of the endothelization process on the polyester arterial prosthesis. PMID- 12108935 TI - Spontaneous regression of bilateral dentigerous cysts. PMID- 12108936 TI - Tongue bar bells. PMID- 12108937 TI - Ingested foreign bodies. PMID- 12108938 TI - Oral ulceration. PMID- 12108939 TI - General anaesthesia. PMID- 12108940 TI - HIV patients seek better dental care. PMID- 12108941 TI - Gingival recession due to trauma caused by a lower lip stud. AB - Body piercing seems to be increasing in popularity, with more patients attending for their routine check-up having had a tongue or lip stud placed. Many complications have been documented, some of them, particularly involving tongue piercing, can be life threatening. There appears to be fewer problems associated with lip piercing; however this case study illustrates the damage that a lip stud can cause to previously healthy gingival tissue. PMID- 12108942 TI - Crowns and other extra-coronal restorations: provisional restorations. AB - The important role of provisional restorations is often overlooked. This may be because they are left until the end of an appointment when time for construction is short or because they generally do not need to last for long. However, not only can good provisional restorations help produce better final restorations, they can also save a lot of time and expense at subsequent appointments. In fact time spent in their construction will be more than repaid in time saved doing additional procedures, adjustments and remakes later on. PMID- 12108943 TI - Trends in oral surgery in England and Wales 1991-2000. AB - OBJECTIVE: To investigate trends in oral surgery in England and Wales 1991-2000. METHODS: Oral surgery procedure data were derived from Dental Practice Board and Department of Health Hospital Episode Statistics. RESULTS: There was a 6% increase in minor oral surgery (MOS) procedures, including ordinary extractions, extractions of special difficulty, apicectomies and third molar removals, carried out in the General Dental Services (GDS) but the number of third molars removed fell by 32% after 1997. General anaesthetics (GA) administered in the GDS fell by 77% and the number of sedations rose 54% after 1998. There was concentration of minor oral surgery in practices: in the year 2000, 88% of practitioners carried out less than five third molar removals. In the Hospital Dental Service (HDS) there was a 98% increase in day surgery, and a 53% decrease in ordinary admissions for minor oral surgery. HDS waiting times remained constant over the ten year period. CONCLUSIONS: The principal trends were substantial decreases in apicectomies, third molar removals after 1997 and GAs after 1998; increases in extractions of special difficulty and concentration of MOS in the GODS. Numbers of ordinary extractions did not change. In the HDS there was a large shift from in-patient to daycase provision which has facilitated expansion of maxillofacial surgery. This is an important example of NHS reconfiguration. Perhaps the most important implication of these changes concerns the place of MOS in vocational training. PMID- 12108944 TI - The changing patterns of drinking, illicit drug use, stress, anxiety and depression in dental students in a UK dental school: a longitudinal study. AB - OBJECTIVE: To investigate alcohol and illicit drug use in a cohort of dental undergraduates through to VT year. SETTING: A UK dental school (with a medical school comparison group). SUBJECTS AND METHODS: A cohort of dental students anonymously completed a lifestyle questionnaire about drinking and smoking, illicit drug use, stress, anxiety and depression in the spring of 1995 and 1998 as second and final year undergraduate students respectively, and in the summer of 1999 after one year working as qualified dentists. A parallel cohort of medical students also anonymously completed the questionnaire at the same time points in their undergraduate course as for the dental students, and at the end of a year working as Pre-Registration House Officers (PRHOs). RESULTS: The proportion of dental students in Newcastle drinking above the recommended low risk limits of alcohol declined from 47% as second year students to 25% as final year students and then it increased to 41% as qualified dentists, whilst in medical students it steadily increased over the three time points of the survey (33% to 43% to 54%). A greater proportion of dental students were drinking at hazardous levels at all three time-points, compared with medical students. Experimentation with illicit drugs ranged from 47% as second year students to 54% as final year students and to 51% as dentists. The prevalence of illicit drug use in medical students was similar to that in dental students. Forty seven per cent of the dental student cohort as second year students, 67% as final year students and 16% as dentists suffered from possible pathological anxiety, compared with 47%, 26% and 30% in the medical student cohort. The proportion of dentists suffering from stress decreased from 72% as final year students to 19% as dentists. In the medical student group, the proportion increased from 32% as final year students to 39% as PRHOs. CONCLUSION: This longitudinal study revealed that a high proportion of dental students from Newcastle continue to drink excessively and experiment with illicit drugs both as undergraduates and as practising dentists. A significant proportion also suffer from anxiety and stress. Further measures are needed in order to reduce alcohol and substance misuse and stress and anxiety among dental students and dentists. PMID- 12108945 TI - Toothache tales: Part 1. AB - This selection of literary extracts deals with the subject of toothache from the point of view of the sufferer. Circumstances, effects and remedies, orthodox and otherwise, are described. PMID- 12108946 TI - The promise and obstacle of p53 as a cancer therapeutic agent. AB - p53 is a tumor suppressor gene that is mutated in greater than 50% of human cancers. The action of p53 as a tumor suppressor involves inhibition of cell proliferation through cell cycle arrest and/or apoptosis. Loss of p53 function therefore allows the uncontrolled proliferation associated with cancerous cells. While design of most anti-cancer agents has focused on targeting and inactivating cancer promoting targets, such as oncogenes, recent attention has been given to restoring the lost activity of tumor suppressor genes. Because the loss of p53 function is so prevalent in human cancer, this protein is an ideal candidate for such therapy. Several gene therapeutic strategies have been employed in the attempt to restore p53 function to cancerous cells. These approaches include introduction of wild-type p53 into cells with mutant p53; the use of small molecules to stabilize mutant p53 in a wild-type, active conformation; and the introduction of agents to prevent degradation of p53 by proteins that normally target it. In addition, because mutant p53 has oncogenic gain of function activity, several approaches have been investigated to selectively target and kill cells harboring mutant p53. These include the introduction of mutant viruses that cause cell death only in cells with mutant p53 and the introduction of a gene that, in the absence of functional p53, produces a toxic product. Many obstacles remain to optimize these strategies for use in humans, but, despite these, restoration of p53 function is a promising anti-cancer therapeutic approach. PMID- 12108947 TI - Malignant hyperthermia: a pharmacogenetic disease of Ca++ regulating proteins. AB - Malignant hyperthermia (MH) is a pharmacogenetic, life-threatening hypermetabolic syndrome in genetically predisposed individuals exposed to certain anesthetic agents. Discovered by Denborough and Lovell [1] in 1960, MH was associated with high mortality and morbidity as the cause was unknown and an effective treatment was unavailable. There is no classic clinical presentation of the syndrome, and the onset and signs of MH are dependent upon known and unknown environmental and genetic factors. Initial theories involved central temperature regulation defects or uncoupling of oxidative phosphorylation in mitochondria [2], but later investigations targeted skeletal muscle as the affected organ. Subsequently freshly biopsied skeletal muscle was used for in vitro pharmacologic contracture testing to discriminate between normal and MH-affected muscle and remains the "gold standard" for MH diagnosis. Spontaneous, genetic models for MH were discovered in pigs and dogs and substantial knowledge about MH was gained from these valuable resources. The abnormal contracture response of MH skeletal muscle evoked a focus on calcium regulation, and abnormalities in calcium release (as opposed to calcium sequestration) mechanisms were discovered. About this same time the major calcium release channel in the skeletal muscle sarcoplasmic reticulum membrane was purified and named the ryanodine receptor [3]. Although the ryanodine receptor represents one of the largest functional proteins, the enormous gene encoding the 5021 amino acids comprising the ryanodine receptor subunit was eventually cloned [4,5]. Patient and dedicated work on the ryanodine receptor gene has found linkage to MH in the pig [6], dog [7], and among several different mutations and MH in unrelated human families [8,9]. Expression of these mutations in HEK cells has resulted in abnormal calcium release [10,11], supporting but not proving a causal basis for MH. In this review each of the areas mentioned above is discussed in detail revealing a wonderful success story that changed the anesthesiologist's "worst nightmare" from a syndrome with high mortality and morbidity to a reasonably well managed disease today. This success story includes unraveling the molecular basis for the disease and brings its pathoetiologic and diagnostic aspects toward molecular genetic resolution. PMID- 12108948 TI - Critical role for IL-15 in innate immunity. AB - Although some functional activities of interleukin (IL)-15 on NK and T cells overlap with those of IL-2, recent findings obtained from gene-targeted mice deficient in components of IL-2/IL-15 system demonstrate distinct roles of IL-15 for the activation of innate immune system. IL-15 is a pivotal cytokine for the development and survival of NK cells, NKT cells, TCRydelta+ intestinal intraepithelial lymphocytes (ilEL), and for the functional maturation of dendritic cells and macrophages. IL-15 is also important for memory T cell maintenance in vivo. In this review, I summarize recent progress of studies in the IL-15/IL-15R system. PMID- 12108950 TI - Immunity to systemic Salmonella infections. AB - Salmonella infections are a serious public health problem in developing countries and represent a constant concern for the food industry. The severity and the outcome of a systemic Salmonella infection depends on the "virulence" of the bacteria, on the infectious dose as well as on the genetic makeup and immunological status of the host. The control of bacterial growth in the reticuloendothelial system (RES) in the early phases of a Salmonella infection relies on the NADPH oxidase-dependent anti-microbial functions of resident phagocytes and is controlled by the innate resistance gene Nramp1. This early phase is followed by the suppression of Salmonella growth in the RES due to the onset of an adaptive host response. This response relies on the concerted action of a number of cytokines (TNFalpha, IFNgamma, IL12, IL18, and IL15), on the recruitment of inflammatory phagocytes in the tissues and on the activation of the recruited cells. Phagocytes control bacterial growth in this phase of the infection by producing reactive nitrogen intermediates (RNI) generated via the inducible nitric oxide synthase (iNOS). Clearance of the bacteria from the RES at a later stage of the infection requires the CD28-dependent activation of CD4+ TCR alphabeta T-cells and is controlled by MHC class II genes. Resistance to re infection with virulent Salmonella micro-organisms requires the presence of Th1 type immunological memory and anti-Salmonella antibodies. Thus, the development of protective immunity to Salmonella infections relies on the cross-talk between the humoral and cellular branches of the immune system. PMID- 12108949 TI - The role of STATs in apoptosis. AB - Signal transducers and activators of transcription (STATs) are transcription factors that mediate cytokine and growth factor induced signals that culminate in various biological responses, including proliferation and differentiation. Recent studies indicate a role for STATs in apoptosis as well. Depending upon the particular stimulus or cell type, STATs can mediate either pro-apoptotic signals or anti-apoptotic signals. STAT1 and, under some circums-tances. STAT3 are important for transducing pro-apoptotic signals whereas STAT3 and STAT5 have been implicated in promoting cell survival. Recent studies demonstrate that regulation of apoptotic pathways by STATs is largely due to transcriptional activation of genes that encode proteins that mediate or trigger the cell death process, such as Bcl-xL, caspases, Fas and TRAIL as well as those that regulate cell cycle progression, such as p21waf1. Interestingly, STAT proteins may also regulate apoptosis through a non-transcriptional mechanism by inhibiting the anti apoptotic protein NF-kappaB. Considering that dysregulation of the STAT signaling pathway is commonly found in clinical tumor samples, understanding the mechanisms underlying STAT regulation of cell survival may lead to successful strategies for targeting STATs in cancer therapy. PMID- 12108951 TI - Rate limiting steps of AAV transduction and implications for human gene therapy. AB - Despite the fact that adeno-associated virus type 2 (AAV2) is an extremely attractive gene therapy vector, its application has been limited to certain tissues such as muscle and the brain. In an attempt to broaden the array of target organs for this vector, molecular studies on the mechanism(s) of AAV transduction have expanded over the past several years. These studies have led to the development of innovative strategies capable of overcoming intracellular barriers to AAV2 transduction. The basis of these technologic breakthroughs has stemmed from a better understanding of the molecular processes that control AAV entry and intracellular trafficking to the nucleus. This review will focus on the identification of molecular components important for recombinant AAV (rAAV) transduction while highlighting the techniques used to discover them and potential clinical application of research findings. PMID- 12108952 TI - Adenoviral vector-mediated gene transfer for human gene therapy. AB - Human gene therapy promises to change the practice of medicine by treating the causes of disease rather than the symptoms. Since the first clinical trial made its debut ten years ago, there are over 400 approved protocols in the United States alone, most of which have failed to show convincing data of clinical efficacy. This setback is largely due to the lack of efficient and adequate gene transfer vehicles. With the recent progress in elucidating the molecular mechanisms of human diseases and the imminent arrival of the post genomic era, there are increasing numbers of therapeutic genes or targets that are available for gene therapy. Therefore, the urgency and need for efficacious gene therapies are greater than ever. Clearly, the current fundamental obstacle is to develop delivery vectors that exhibit high efficacy and specificity of gene transfer. Recombinant adenoviruses have provided a versatile system for gene expression studies and therapeutic applications. Of late, there has been a remarkable increase in adenoviral vector-based clinical trials. Recent endeavors in the development of recombinant adenoviral vectors have focused on modification of virus tropism, accommodation of larger genes, increase in stability and control of transgene expression, and down-modulation of host immune responses. These modifications and continued improvements in adenoviral vectors will provide a great opportunity for human gene therapy to live up to its enormous potential in the second decade. PMID- 12108953 TI - Imaging methods in gene therapy of cancer. AB - Clinical gene therapy needs non invasive tools to evaluate the efficiency of gene transfer. This includes the evaluation of infection efficiency as well as the verification of successful gene transfer in terms of gene transcription. These informations can be used for therapy planning, follow up studies in treated tumors and as an indicator of prognosis. Therapy planning is performed by the assessment of gene expression for example using radiolabeled specific substrates to determine the activity of suicide enzymes as the Herpes Simplex Virus thymidine kinase or cytosine deaminase. Furthermore, other in vivo reporter genes as receptors, antigens or transport proteins may be used in bicistronic vector systems for the evaluation of gene transduction and expression. This is done using radiolabeled ligands, antigens or substrates. Follow up studies with magnetic resonance imaging, single photon emission tomography or positron emission tomography may be done to evaluate early or late effects of gene therapy on tumor volume, metabolism or proliferation. Finally, enhancement of radioactive isotope accumulation in tumors by transfer of the appropriate genes may be used for the treatment of malignant tumors. PMID- 12108954 TI - Viral based gene therapy for prostate cancer. AB - In the last few years, significant advances in gene therapy have been made as a result of advances in many areas of molecular and cell biology, including the improvement of both viral and nonviral gene delivery systems, discovery of new therapeutic genes, better understanding of mechanism of disease progression, exploration of tissue specific promoter, receptor- and antibody-mediated targeting delivery, and development of better prodrug enzyme/prodrug systems. In this article, viral based gene therapy for prostate cancer will be reviewed and discussed. The areas of emphasis in this review are: choice of viral vectors, comparison of delivery routes, development of prostate-targeted viruses, choice of therapeutic genes and strategies including corrective gene therapy (tumor suppressor gene and anti-oncogene gene approaches), suicide gene therapy, programmed cell death therapy, immunomodulation therapy, and conditional oncolytic virus approach. Among them, several examples will be discussed in detail for the scientific basis and therapeutic applications. In addition, prostate cancer gene therapy clinical trials, unresolved problems and future directions in this field will also be described. PMID- 12108955 TI - Nonviral gene therapy. AB - The last 10 years have seen substantial progress in the development and application of nonviral vectors in gene therapy. Several novel nonviral methods have been developed that approach viruses with respect to transfection efficiency. A variety of nonviral delivery systems that can be used for gene therapy in different clinical settings are also available. In this review article, we will detail all of the major nonviral vectors that are currently used in gene therapy while highlighting some recent developments, particularly the progress towards the understanding of the cellular and in vivo barriers in gene transfer. Recent advancement in achieving sustained and regulated gene expression will also be addressed. Finally, this review will briefly cover targeted gene repair using nonviral delivery systems. Their impact in gene therapy will also be discussed. PMID- 12108956 TI - Transplantation-based gene therapy for inflammatory diseases: focus on glomerulonephritis. AB - Over the past decade, bone marrow transplantation has come to be considered an ideal therapeutic strategy for the treatment of certain diseases affecting the hematopoietic system such as hemophilia, and several clinical trials have been performed. Although traditionally used for the treatment of lethal diseases, it is speculated that this approach could also be used in the treatment of non lethal but much more common diseases, which are resistant to conventional therapies, and affect a large number of patients physically and even financially. Inflammation may be one target for transplantation-based gene therapy, since macrophages and neutrophils, which are basically derived from hematopoietic stem cells, have been identified as key determinants in the development of diseases. This article focuses on the glomerulonephritis as a model of local inflammation and reviews recent investigations on transplantation-based gene therapy for inflammatory diseases. PMID- 12108957 TI - The dialysis facility's rights, responsibilities, and duties when there is conflict with family members. AB - The stressful effect that a chronic illness may have on a patient and his or her family can make the requirements of a chronic dialysis treatment program most difficult. There will be times when tensions arise between staff, patients, and family members, especially when the needs of the families and the patients are perceived as being unmet, unacknowledged, or otherwise lacking in importance to the staff. Frustrations, if not addressed and successfully resolved, can lead to conflict and the threat of or incidences of violence. Questions that dialysis unit administrators/managers should ask when considering or revoking visitation privileges are: What are the causes and contributory factors that resulted in the unresolved issues and conflict? Did we clearly communicate our policies and procedures and the consequences for failing to follow them to those accompanying the patient? Did we fail to maintain proper boundaries with the invitee (Allowing the family members to engage in inappropriate behaviors such as threats, cursing, name-calling)? Have we attempted to resolve the issues with the family members? If so, how and what was the outcome? Did we document our actions and the family members' responses? Is the revocation of the visitation privilege the only viable option? If so, why? If not, what else could be done? What steps should be taken to minimize the disruptive effect on the patient, whose family member is being disruptive? What are the reasonable and the required steps that a facility should take to ensure that the patients are properly cared for and free from abuse and neglect? What did we learn from this situation that we could use in future situations? The uncomfortable position that Sacred Heart Hospital found itself in is a difficult one and one that many other clinics have or may experience. The focus on continuing to meet the needs of a patient while dealing with issues, such as those presented here, depicts and represents professionalism at work. Answering these questions before problems arise will help unit administrators be prepared to deal effectively with conflict at the dialysis facility. Having a plan of action in place ensures a suitable, uniform response to unacceptable situations and reduces the risk of reacting inappropriately when those situations arise. PMID- 12108958 TI - How U.S. health plans can be more proactive about identifying and treating CRI. PMID- 12108959 TI - The nursing shortage and its effect on nephrology practice. Interview by Barbara Bednar. PMID- 12108960 TI - Advanced practice nurses in nephrology: collaborative solutions to manpower shortages. AB - As the shortages of both nephrologists and experienced staff nurses increase, innovative solutions will be necessary. The most common CPM will continue to be the nephrology APN hired by and working in collaborative practice with nephrologists to provide care across all settings (hospital, office and dialysis unit). Creative arrangements between dialysis corporations, nephrologists, and APNs may also provide for a team approach for treating renal disease patients within the dialysis unit. The team would include the nephrologist, APN, dialysis staff, dietitian, and social worker. Each team member would fulfill roles appropriate to their training, experience, and demonstrated level of competence with the common goal of improved patient outcomes. This would provide the best use of limited resources. PMID- 12108961 TI - Disease management for Medicaid's kidney failure patients: the Florida initiative. PMID- 12108962 TI - Does pre-dialysis care management reap rewards for patients, renal staff, and payers. AB - This article describes one renal disease management organization's experience in implementing a pre-dialysis care management program to improve outcomes in patients who develop end-stage renal disease (ESRD). Optimal Renal Care, a national renal disease management company, has implemented care management programs for 600+ ESRD patients and 600+ pre-ESRD patients in Oregon (1998) and Hawaii (1999). This article describes our Oregon pre-ESRD program's start-up experience. PMID- 12108963 TI - Improving URRs in the dialysis unit. PMID- 12108964 TI - Evolution of the frequency of hemodialysis for acute and chronic renal failure. PMID- 12108965 TI - Is it time for Medicare to provide reimbursement for daily hemodialysis? PMID- 12108966 TI - Nocturnal: thinking "out of the chair" about dialysis. PMID- 12108967 TI - A patient's perspective on nocturnal dialysis. PMID- 12108968 TI - Short daily HD: making an in-center program work. PMID- 12108969 TI - Heparin therapy in routine hemodialysis. PMID- 12108970 TI - Genetic chemoprotection with mutant O6-alkylguanine-DNA-alkyltransferases. AB - One of the main barriers to more efficacious use of modern chemotherapeutic agents, is the collateral toxicity exhibited in normal, highly proliferative tissues, primarily the haemopoietic, gastrointestinal and pulmonary tissues. Drug resistance of tumours to these drugs compounds this problem. This review discusses the role of O6-alkylguanine-DNA alkyltransferase (ATase) in conferring protection against O6-alkylating agents in normal tissue, focusing mainly on the haemopoietic compartment. The development of mutant forms of ATase, which are resistant to the effects of soluble analogues of O6-alkylation such as O6 benzylguanine, is examined and the gene therapy approach of combining these two strategies to confer chemoprotection to vulnerable tissues whilst sensitising malignant tissue is reviewed. PMID- 12108971 TI - Applications of muscle electroporation gene therapy. AB - Muscle is a convenient and accessible site for non-viral gene delivery, which can manufacture gene products and provide a long-duration of gene expression. The level of gene expression after administration of naked DNA plasmid or polymer formulated DNA plasmid containing a reporter gene to muscle via syringe injection, however, is very low. As a result, no significant therapeutic effect can be detected after saline- or polymer-mediated gene delivery into muscle. In 1998, investigators published a striking new approach--electrotransfection--for intramuscular gene delivery (now commonly referred to as electroporation or electroinjection). Electroporation of a non-viral gene into the muscles of small animals has increased the level of gene expression by as much as two orders of magnitude, which is comparable to levels achieved with adenoviral gene delivery. Three years later, intramuscular electroporation gene delivery technology has blossomed. Treatments for different diseases using this approach in animal models have been reported. In this review, I discuss the applications of intramuscular electroporation gene therapy to treat malignancies, renal disease, and anemia, and to prevent drug toxicity to sensory nerves. PMID- 12108972 TI - Lentiviral vectors for gene therapy of HIV-1 infection. AB - Lentiviral vectors based on HIV-1, HIV-2, or SIV have the ability to transduce dividing and non-dividing T cells, dendritic cells, hematopoietic stem cells and macrophages, which are the main target cells for gene therapy of HIV-1 infection. Besides their function as gene delivery vehicles, lentiviral vector backbones containing the cis-acting sequences necessary to perform a complete replication cycle in the presence of viral proteins provided in trans, have the ability to inhibit HIV-1 replication by several mechanisms that include sequestration of the regulatory proteins Tat and Rev, competition for packaging into virions and possibly by inhibition of reverse transcription in heterodimeric virions. Expression of anti-HIV-1 genes in these vectors would strengthen the potency of this inhibition. To avoid self-inhibition of the vector packaging system, lentiviral vectors have to be modified to become resistant to the anti-HIV-1 genes encoded by them. This review discusses the different genetic intervention strategies for gene therapy of HIV-1 infection focusing in the use of lentiviral vectors as the main agents to mediate inhibition of HIV-1 replication. It also discusses possible strategies to adapt HIV-1 or HIV-2 vectors to express the different classes of anti-HIV-1 genes and approaches to improve in vivo vector mobilization. PMID- 12108973 TI - Gene transfer into hematopoietic stem cells using lentiviral vectors. AB - Gene transfer into hematopoietic cells is currently being used to modulate immune responses, to protect hematopoietic cells against cytotoxic drugs or viral genes, and to restore gene deficiencies due to either inborn genetic defects or acquired loss of regular gene function. In particular, gene addition strategies for inherited severe combined immunodeficiencies (SCID) due to adenosine deaminase (ADA) deficiency or defects of the interleukin-2 receptor gamma-chain represent potentially curative strategies based on gene transfer into hematopoietic cells using recombinant retroviral vectors. Since long-term correction of genetic defects in hematopoietic cells often requires transduction of hematopoietic stem cells, an effective gene transfer into stem cells with efficient long-term and multi-lineage transgene expression is the desired goal for these therapeutic strategies. However, gene transfer strategies with retroviral vectors unable to integrate into non-cycling cells are limited by the quiescent state of the stem cells that have to be stimulated by cytokines to induce cell cycle progression. To circumvent these barriers, lentiviral vector systems based on HIV-1 have recently been developed which are able to deliver and express genes in non dividing cells both in vitro and in vivo. This review outlines the development and improvement of lentivirus-based gene transfer protocols and discusses the use of lentiviral vectors in preclinical gene therapy studies. PMID- 12108974 TI - Cancer immunotherapy using gene-modified dendritic cells. AB - Gene-engineered dendritic cells (DC) are being tested in cancer immunotherapy. DC are the best equipped antigen-presenting cells (APC) to overcome tolerance/ignorance to self antigens presented by cancer cells. Genetic immunotherapy with DC engineered to express tumor antigens has the potential advantages of endogenous epitope presentation by both major histocompatibility complex (MHC) class I and II molecules. DC can also be gene-modified to express immunostimulatory molecules that further enhance their antigen-presenting function. Review of the literature provided 52 manuscripts where gene-modified DC were being tested in murine models of immunotherapy for cancer. Review of the antitumor effects of gene-modified DC in these preclinical studies provides valuable information on the optimal methods of gene transfer into DC, the schedule of administration, the route, dose and the underlying immunological mechanisms of the antitumor effects. These data may help in the translation of this promising approach to the clinic. PMID- 12108975 TI - Cytokine gene transfer into dendritic cells for cancer treatment. AB - Bone marrow-derived dendritic cells have been used to treat established experimental tumors by unleashing a cellular immune response against tumor antigens. Such antigens are artificially loaded onto dendritic cells' antigen presenting molecules by different techniques including incubation with synthetic antigenic determinants, tumor lysates or nucleic acids encoding for those relevant antigens. Ex vivo gene transfer with viral and non-viral vectors is frequently used to obtain expression of the tumor antigens and thereby to formulate the therapeutic vaccines. Efficacy of the approaches is greatly enhanced if dendritic cells are transfected with a number of genes which encode immunostimulating factors. In some cases, such as with IL-12, IL-7 and CD40L genes, injection inside experimental malignancies of thus transfected dendritic cells induces complete tumor regression in several models. In this case tumor antigens are captured by dendritic cells by still unclear mechanisms and transported to lymphoid organs where productive antigen presentation to T-cells takes place. Many clinical trials testing dendritic cell-based vaccines against cancer are in progress and partial clinical efficacy has been already proved. Transfection of genes further strengthening the immunogenicity of such strategies will join the clinical club soon. PMID- 12108976 TI - Genetic engineering in allotransplantation of vascularized organs. AB - Transplantation offers a unique opportunity for gene transfer into allografts before grafting. After organ retrieval, the cold ischemic period renders organs available for manipulation and gene transfer. Local expression of protective or immunomodulatory molecules within the graft environment offers a better local bioavailability of bioreagents and potentially less systemic side effects. Protection against ischemia-reperfusion injury, acute and/or chronic rejection without significant side effects would be a major breakthrough in transplant research. However, protocols of transfection adapted to the transplant setting and control of gene expression must be clearly evaluated before going to clinical trials. The first part of this review deals with gene transfer techniques into the allograft, emphasizing particular transplant conditions that are encountered and that must be respected when designing protocols for gene transfer experiments. The second part deals with specific therapeutic strategies to protect and prolong allograft survival. PMID- 12108978 TI - Atom transfer radical cyclisation reactions mediated by copper complexes. AB - This article describes recent advances in the use of copper complexes in mediating atom transfer radical cyclisation reactions (ATRC). Recent developments have included the design of activated complexes which mediate the cyclisation of tri-, di-, and mono-halo derived substrates at ambient temperatures. Using this methodology, cyclisation to give a variety of ring sizes (4-18 membered rings) has been demonstrated. In addition tandem and radical-polar crossover reactions have also been developed. The design of solid supported and perfluorous complexes that mediate cyclisations may make this approach to the synthesis of rings more attractive towards industrial applications. PMID- 12108977 TI - Improvement of adoptive cellular immunotherapy of human cancer using ex-vivo gene transfer. AB - A variety of adoptive cellular strategies, aimed at boosting the immune system, have been tested in the management of metastatic diseases. Despite the drawbacks associated with ex vivo cell manipulation and upscaling, several such approaches have been assessed in the clinic. The use of lymphokine-activated killer (LAK) cells, auto-lymphocyte therapy (ALT) and tumor-infiltrating lymphocytes (TIL) have been the best studied and further trials are ongoing. Thus far, these approaches have not consistently shown benefit when compared to standard immune based treatment with biologic response modifiers, notably, high-dose interleukin 2 (IL-2). More recently, it has been shown, in various animal models, that the ex vivo transfer of genes to cells of the immune system can have a dramatic impact on cancer immunotherapy. The application of gene transfer techniques to immunotherapy has animated the field of cell-based cancer therapy research. A wide variety of viral and non-viral gene transfer methods have been investigated in this context. Ex vivo strategies include gene delivery into tumor cells and into cellular components of the immune system, including cytotoxic T cells, NK, macrophages and dendritic cells (DC). Several of these approaches have already been translated into cancer therapy clinical trials. In this review, we focus on the rationale and types of ex vivo gene-based immunotherapy of cancer. Finally, the use of genetically modified DC for tumor vaccination and its prospects are discussed. PMID- 12108979 TI - Nucleophilic transition metal based cyclization of allenes. AB - Allenes bearing a pro-nucleophile can be cyclized on treatment with a wide variety of transition metal catalysts and reagents: palladium, cobalt, ruthenium, silver, rhodium, lanthanides, gold. The nucleophilic groups can be nitrogen, oxygen or carbon based and can form rings of various sizes, often with good control of stereochemistry. A variety of mechanisms can be proposed for these reactions and the metal complex can be used to introduce a variety of functional groups during cyclization. Several heterocyclic natural products have been prepared using a selection of these reactions. PMID- 12108981 TI - Laser control of chemical reactions. AB - Chemical reactions are at the heart of chemistry and the dream of controlling the outcome of these reactions is an old one. Thus, with given reactants, a solvent and perhaps assisted by a catalyst, we would like to 'steer' the reactants into a particular desired product. This review focuses on how to control the dynamics of chemical reactions, beyond traditional temperature control, with the emphasis on unimolecular reactions. The electromagnetic radiation of lasers can induce so called coherent dynamics. The recent theoretical and experimental results on this coherent control are explained and illustrated with computational and experimental examples. PMID- 12108980 TI - Fullerene-porphyrin architectures; photosynthetic antenna and reaction center models. AB - Fullerenes and porphyrins are molecular architectures ideally suited for devising integrated, multicomponent model systems to transmit and process solar energy. Implementation of C60 as a 3-dimensional electron acceptor holds great expectations on account of their small reorganization energy in electron transfer reactions and has exerted a noteworthy impact on the improvement of light-induced charge-separation. This article describes how the specific compositions of porphyrin chromophores linked to C60--yielding artificial light harvesting antenna and reaction center mimics--have been elegantly utilized to tune the electronic couplings between donor and acceptor sites and the total reorganization energy. Specifically, the effects that these parameters have on the rate, yield and lifetime of the energetic charge-separated states are considered. PMID- 12108982 TI - Celastraceae sesquiterpenoids: biological activity and synthesis. AB - Plant extracts of the Celastraceae have been used for centuries throughout South America and China as insect repellents and insecticides in traditional agriculture, and also for the treatment of a plethora of medical ailments from stomach complaints and fever to rheumatoid arthritis and cancer. Many of the medicinally interesting properties associated with these crude preparations have now been attributed to a large family of highly oxygenated sesquiterpenoids based on a tricyclic dihydroagarofuran skeleton. In this article, the structural diversity and range of biological activities associated with this intriguing class of natural products are examined with a view to stimulating interest in their total synthesis. Existing synthetic endeavours towards their synthesis are also evaluated. PMID- 12108984 TI - HHS, OIG look at value of donor registries. PMID- 12108983 TI - Methods and techniques for the selective extraction and recovery of oxoanions. AB - An important goal in environmental chemistry is the extraction of metals that are toxic or radioactive from soils and waters. For many such metals, the problem is solved by designing compounds with coordination sites that are specific for that particular metal. For cases such as oxoanions, however, where the inorganic center is already fully coordinated by oxygens, different strategies need to be used. Chromate, phosphate, selenate, pertechnetate, and aluminate are such anions. These species are problematic contaminants in soils and waters because they are either toxic, environmentally undesirable, or radioactive. Furthermore, under both acidic and basic conditions, the coordination positions of these oxoanions are occupied by oxygens. Chromium(VI) as either chromate or dichromate presents a particular problem because it is a strong oxidizing agent. In this review the different methods of extracting this group of pseudotetrahedral oxoanions are discussed, along with the individual advantages and limitations of each strategy. PMID- 12108985 TI - Can registries make a difference? PMID- 12108986 TI - Viral load monitoring in BK virus-associated nephropathy. PMID- 12108987 TI - Maintaining the transplant waiting list. AB - The renal transplant waiting list now has over 51,000 persons hoping for a new kidney. Median waiting time is 18 to 32 months. This article reviews some consideration in managing this population during the waiting period. This article reviews who is responsible for maintenance of the list, which patients should be assessed and at what intervals, and how patients should be removed from the list. Consideration is also given to the financial aspect of continuing evaluation. PMID- 12108988 TI - The transplant evaluation process: a case study. PMID- 12108989 TI - Making the numbers work. URREA has added Social Security Death Master File statistics to supplement missing data from transplant facility follow-up reports. PMID- 12108990 TI - Show me the donors! Failed efforts by mass media to increase donation signal time for bold new initiatives. PMID- 12108991 TI - What does it mean for patients? The expanded Medicare coverage of immunosuppressive medications. PMID- 12108992 TI - A center-based approach to a transplant employment program. AB - The return to work after transplantation has been actively discussed in the transplant community for years. However, despite the desire for transplant practitioners to have recipients return to being healthy, contributing members of society as well as return to work, practitioners often passively support the sick role. We discovered that patients who are returning to work after transplantation may have been out of the work force for several years, and require assistance that we as health care providers were unable or untrained to provide. An employment specialist was added to the transplant staff and became a vital part of our attempts to create a proactive employment atmosphere and enhance our patients' quality of life. Adding an advocate for employment in our center has also facilitated the shift in our thinking and approach to care from the sick role to one of rehabilitation and from reactive to proactive. This change in attitude has assisted in empowering our recipients to feel as if they can truly resume a normal life. PMID- 12108993 TI - Remove price controls to relieve the shortage. PMID- 12108994 TI - Improve the organ recovery system. AB - Following the revised 1987 AGA, at least to the extent of correct interaction with donor families, can reduce organ waiting times to nominal within a decade, and could save about $360 million per year. Fear of lawsuits is no excuse for not implementing the revised AGA guidelines. Following the 1987 AGA revision in its entirety is non-controversial and its improvements are easy to implement. Several other system improvements which could also help were suggested. To optimize the system, the concept of 'intestate as to organs' should be implemented. It has precedent and is ethical. Compensation for burial expenses is not recommended. PMID- 12108995 TI - Back to basics. How to negotiate contracts with health plans. PMID- 12108996 TI - Continuous flow peritoneal dialysis. PMID- 12108997 TI - Bone disease: a primer/Part I. Clinical differences among vitamin D hormone compounds. PMID- 12108998 TI - Chronic inflammation and water quality in hemodialysis patients. AB - Chronic inflammation, the associated cardiovascular disease, and attendant high mortality remain huge problems in the HD population. Determining the predominant causal factors in the triggering and maintenance of this inflammatory state is of paramount importance in formulating treatment strategies for individuals and for centers. In most cases, causation is multifactorial and several different areas of the HD process need to be considered. In this respect, the water system in HD centers is certainly one parameter that demands careful design, rigorous monitoring and strict adherence to effective disinfection protocols. In this way it may be possible not only to reduce levels of inflammatory markers, but to improve performance in other clinical areas, such as EPO therapy, and perhaps outcomes in the HD population. PMID- 12108999 TI - The needlestick act: getting the point. Part II. PMID- 12109000 TI - Splitting fibers: understanding how dialyzer differences can impact adequacy. PMID- 12109001 TI - Annual Network/HCFA Report on clinical indicators shows more patients getting better care. But catheter use is up, serum albumin shows little change. PMID- 12109002 TI - Improving hematocrits. In the dialysis unit. PMID- 12109003 TI - Does MFD fit under Medicare coverage? PMID- 12109004 TI - Teaching psychology as a laboratory science in the age of the Internet. AB - For over 30 years, psychologists have relied on computers to teach experimental psychology. With the advent of experiment generators, students can create well designed experiments and can test sophisticated hypotheses from the start of their undergraduate training. Characteristics of new Net-based experiment generators are discussed and compared with traditional stand-alone generators. A call is made to formally evaluate the instructional effectiveness of the wide range of experiment generators now available. Specifically, software should be evaluated in terms of known learning outcomes, using appropriate control groups. The many inherent differences between any two software programs should be made clear. The teacher's instructional method should be fully described and held constant between comparisons. Finally, the often complex interaction between the teacher's instructional method and the pedagogical details of the software must be considered. PMID- 12109005 TI - Fitting mixed-effects models for repeated ordinal outcomes with the NLMIXED procedure. AB - This paper presents an analysis of repeated ordinal outcomes arising from two psychological studies. The first case is a repeated measures analysis of variance; the second is a mixed-effects regression in a longitudinal design. In both, the subject-specific variation is modeled by including random effects in the linear predictor (inside a link function) of a generalized linear model. The NLMIXED procedure in SAS is used to fit the mixed-effects models for the categorical response data. The presentation emphasizes the parallel between the model specifications and the SAS statements. The purpose of this paper is to facilitate the use of mixed-effects models in the analysis of repeated ordinal outcomes. PMID- 12109006 TI - Multidimensional data visualization. AB - Historically, data visualization has been limited primarily to two dimensions (e.g., histograms or scatter plots). Available software packages (e.g., Data Desk 6.1, MatLab 6.1, SAS-JMP 4.04, SPSS 10.0) are capable of producing three dimensional scatter plots with (varying degrees of) user interactivity. We constructed our own data visualization application with the Visualization Toolkit (Schroeder, Martin, & Lorensen, 1998) and Tcl/Tk to display multivariate data through the application of glyphs (Ware, 2000). A glyph is a visual object onto which many data parameters may be mapped, each with a different visual attribute (e.g., size or color). We used our Multi-Dimensional data Viewer to explore data from several psycholinguistic experiments. The graphical interface provides flexibility when users dynamically explore the multidimensional image rendered from raw experimental data. We highlight advantages of multidimensional data visualization and consider some potential limitations. PMID- 12109007 TI - Improving computer security for authentication of users: influence of proactive password restrictions. AB - Entering a username-password combination is a widely used procedure for identification and authentication in computer systems. However, it is a notoriously weak method, in that the passwords adopted by many users are easy to crack. In an attempt to improve security, proactive password checking may be used, in which passwords must meet several criteria to be more resistant to cracking. In two experiments, we examined the influence of proactive password restrictions on the time that it took to generate an acceptable password and to use it subsequently to long in. The required length was a minimum of five characters in Experiment 1 and eight characters in Experiment 2. In both experiments, one condition had only the length restriction, and the other had additional restrictions. The additional restrictions greatly increased the time it took to generate the password but had only a small effect on the time it took to use it subsequently to long in. For the five-character passwords, 75% were cracked when no other restrictions were imposed, and this was reduced to 33% with the additional restrictions. For the eight-character passwords, 17% were cracked with no other restrictions, and 12.5% with restrictions. The results indicate that increasing the minimum character length reduces crackability and increases security, regardless of whether additional restrictions are imposed. PMID- 12109008 TI - Implications of document-level literacy skills for Web site design. AB - The proliferation of World Wide Web (Web) sites and the low cost of publishing information on the Web have placed a tremendous amount of information at the fingertips of millions of people. Although most of this information is at least intended to be accurate, there is much that is rumor, innuendo, urban legend, and outright falsehood. This raises problems especially for students (of all ages) trying to do research or learn about some topic. Finding accurate, credible information requires document-level literacy skills, such as integration, sourcing, corroboration, and search. This paper discusses these skills and offers a list of simple ways that designers of educational Web sites can help their visitors utilize these skills. PMID- 12109009 TI - Personality research on the Internet: a comparison of Web-based and traditional instruments in take-home and in-class settings. AB - Students, faculty, and researchers have become increasingly comfortable with the Internet, and many of them are interested in using the Web to collect data. Few published studies have investigated the differences between Web-based data and data collected with more traditional methods. In order to investigate these potential differences, two important factors were crossed in this study: whether the data were collected on line or not and whether the data were collected in a group setting at a fixed time or individually at a time of the respondent's choosing. The Visions of Morality scale (Shelton & McAdams, 1990) was used, and the participants were assigned to one of four conditions: in-class Web survey, in class paper-and-pencil survey; take-home Web survey, and take-home paper-and pencil survey. No significant differences in scores were found for any condition; however, response rates were affected by the type of survey administered, with the take-home Web-based instrument having the lowest response rate. Therefore, researchers need to be aware that different modes of administration may affect subject attrition and may, therefore, confound investigations of other independent variables. PMID- 12109010 TI - Using latent semantic analysis to assess reader strategies. AB - We tested a computer-based procedure for assessing reader strategies that was based on verbal protocols that utilized latent semantic analysis (LSA). Students were given self-explanation-reading training (SERT), which teaches strategies that facilitate self-explanation during reading, such as elaboration based on world knowledge and bridging between text sentences. During a computerized version of SERT practice, students read texts and typed self-explanations into a computer after each sentence. The use of SERT strategies during this practice was assessed by determining the extent to which students used the information in the current sentence versus the prior text or world knowledge in their self explanations. This assessment was made on the basis of human judgments and LSA. Both human judgments and LSA were remarkably similar and indicated that students who were not complying with SERT tended to paraphrase the text sentences, whereas students who were compliant with SERT tended to explain the sentences in terms of what they knew about the world and of information provided in the prior text context. The similarity between human judgments and LSA indicates that LSA will be useful in accounting for reading strategies in a Web-based version of SERT. PMID- 12109012 TI - Open courseware and shared knowledge in higher education. AB - Most college and university campuses in the United States and much of the developed world today maintain one, two, or several learning management systems (LMSs), which are courseware products that provide students and faculty with Web based tools to manage course-related applications. Since the mid-1990s, two predominant models of Web courseware management systems have emerged: commercial and noncommercial. Some of the commercial products available today were created in academia as noncommercial but have since become commercially encumbered. Other products remain noncommercial but are struggling to survive in a world of fierce commercial competition. This article argues for an ethics of pedagogy in higher education that would be based on the guiding assumptions of the non-proprietary, peer-to-peer, open-source software movement. PMID- 12109011 TI - Web-based experiments controlled by JavaScript: an example from probability learning. AB - JavaScript programs can be used to control Web experiments. This technique is illustrated by an experiment that tested the effects of advice on performance in the classic probability-learning paradigm. Previous research reported that people tested via the Web or in the lab tended to match the probabilities of their responses to the probabilities that those responses would be reinforced. The optimal strategy, however, is to consistently choose the more frequent event; probability matching produces suboptimal performance. We investigated manipulations we reasoned should improve performance. A horse race scenario in which participants predicted the winner in each of a series of races between two horses was compared with an abstract scenario used previously. Ten groups of learners received different amounts of advice, including all combinations of (1) explicit instructions concerning the optimal strategy, (2) explicit instructions concerning a monetary sum to maximize, and (3) accurate information concerning the probabilities of events. The results showed minimal effects of horse race versus abstract scenario. Both advice concerning the optimal strategy and probability information contributed significantly to performance in the task. This paper includes a brief tutorial on JavaScript, explaining with simple examples how to assemble a browser-based experiment. PMID- 12109013 TI - Using NetCloak to develop server-side Web-based experiments without writing CGI programs. AB - Server-side experiments use the Web server, rather than the participant's browser, to handle tasks such as random assignment, eliminating inconsistencies with JAVA and other client-side applications. Heretofore, experimenters wishing to create server-side experiments have had to write programs to create common gateway interface (CGI) scripts in programming languages such as Perl and C++. NetCloak uses simple, HTML-like commands to create CGIs. We used NetCloak to implement an experiment on probability estimation. Measurements of time on task and participants' IP addresses assisted quality control. Without prior training, in less than 1 month, we were able to use NetCloak to design and create a Web based experiment and to help graduate students create three Web-based experiments of their own. PMID- 12109014 TI - A server-side program for delivering experiments with animations. AB - A server-side program for animation experiments is presented. The program is capable of delivering an experiment composed of discrete animation sequences in various file formats, collecting a discrete or continuous response from the observer, evaluating the appropriateness of the response, and ensuring that the user is not proceeding at an unreasonable rate. Most parameters of the program are controllable by experimenter-edited text files or simple switches in the program code, thereby minimizing the need for programming to create new experiments. A simple demonstration experiment is discussed and is freely available. PMID- 12109015 TI - Toward an Experimental Timing Standards Lab: benchmarking precision in the real world. AB - Much discussion has taken place over the relative merits of various platforms and operating systems for real-time data collection. Most would agree that, provided great care is taken, many are capable of millisecond timing precision. However, to date, much of this work has focused on the theoretical aspects of raw performance. It is our belief that researchers would be better informed if they could place confidence limits on their own specific paradigms in situ and without modification. To this end, we have developed a millisecond precision test rig that can control and time experiments on a second presentation machine. In this paper, we report on the specialist hardware and software used. We elucidate the importance of the approach in relation to real-world experimentation. PMID- 12109016 TI - A multimodal data collection tool using REALbasic and Mac OS X. AB - This project uses REALbasic 3.5 in the Mac OS X environment for development of a configuration tool that builds a data collection procedure for investigating the effectiveness of sonified graphs. The advantage of using REALbasic with the Mac OS X system is that it provides rapid development of stimulus presentation, direct recording of data to files, and control over other procedural issues. The program can be made to run natively on the new Mac OS X system, older Mac OS systems, and Windows (98SE, ME, 2000 PRO). With modification, similar programs could be used to present any number of visual/auditory stimulus combinations, complete with questions for each stimulus. PMID- 12109017 TI - ePsych: interactive demonstrations and experiments in psychology. AB - ePsych (http://epsych.msstate.edu), a new Web site currently under active development, is intended to teach students about the discipline of psychology. The site presumes little prior knowledge about the field and so may be used in introductory classes, but it incorporates sufficient depth of coverage to be useful in more advanced classes as well. Numerous interactive and dynamic elements are incorporated into various modules, orientations, and guidebooks. These elements include Java-based experiments and demonstrations, video clips, and animated diagrams. Rapid access to all material is provided through a layer based navigation system that allows users to visit various "Worlds of the Mind." Active learning is encouraged, by challenging students with puzzles and problems and by providing the opportunity to "dig deeper" to learn more about the phenomena at hand. PMID- 12109018 TI - WEXTOR: a Web-based tool for generating and visualizing experimental designs and procedures. AB - WEXTOR is a Javascript-based experiment generator and teaching tool on the World Wide Web that can be used to design laboratory and Web experiments in guided step by-step process. It dynamically creates the customized Web pages and Javascripts needed for the experimental procedure and provides experimenters with a print ready visual display of their experimental design. WEXTOR flexibly supports complete and incomplete factorial designs with between-subjects, within-subjects, and quasi-experimental factors, as well as mixed designs. The software implements client-side response time measurement and contains a content wizard for creating interactive materials, as well as dependent measures (graphical scales, multiple choice items, etc.), on the experiment pages. However, it does not aim to replace a full-fledged HTML editor. Several methodological features specifically needed in Web experimental design have been implemented in the Web-based tool and are described in this paper. WEXTOR is platform independent. The created Web pages can be uploaded to any type of Web server in which data may be recorded in logfiles or via a database. The current version of WEXTOR is freely available for educational and noncommercial purposes. Its Web address is http://www.genpsylab.unizh.ch/wextor/index.html. PMID- 12109019 TI - On-line Homework/Quiz/Exam applet: freely available Java software for evaluating performance on line. AB - The Homework/Quiz/Exam applet is a freely available Java program that can be used to evaluate student performance on line for any content authored by a teacher. It has database connectivity so that student scores are automatically recorded. It allows several different types of questions. Each question can be linked to images and detailed story problems. Three levels of feedback are provided to student responses. It allows teachers to randomize the sequence of questions and to randomize which of several options is the correct answer in multiple-choice questions. The creation and editing of questions involves menu selections, button presses, and the typing of content; no programming knowledge is required. The code is open source in order to encourage modifications that will meet individual pedagogical needs. PMID- 12109021 TI - Measuring keyboard response delays by comparing keyboard and joystick inputs. AB - The response characteristics of PC keyboards have to be identified when they are used as response devices in psychological experiments. In the past, the proposed method has been to check the characteristics independently by means of external measurement equipment. However, with the availability of different PC models and the rapid pace of model change, there is an urgent need for the development of convenient and accurate methods of checking. The method proposed here consists of raising the precision of the PC's clock to the microsecond level and using a joystick connected to the MIDI terminal of a sound board to give the PC an independent timing function. Statistical processing of the data provided by this method makes it possible to estimate accurately the keyboard scanning interval time and the average keyboard delay time. The results showed that measured keyboard delay times varied from 11 to 73 msec, depending on the keyboard model, with most values being less than 30 msec. PMID- 12109020 TI - Computer program to generate operant schedules. AB - A computer program for programming schedules of reinforcement is described. Students can use the program to experience schedules of reinforcement that are typically used with nonhuman subjects. A cumulative recording of a student's responding can be shown on the screen and/or printed with the computer's printer. The program can also be used to program operant schedules for animal subjects. The program was tested with human subjects experiencing fixed ratio, variable ratio, fixed interval, and variable interval schedules. Performance for human subjects on a given schedule was similar to performance for nonhuman subjects on the same schedule. PMID- 12109022 TI - Full-screen ultrafast video modes over-clocked by simple VESA routines and registers reprogramming under MS-DOS. AB - Fast full-screen presentation of stimuli is necessary in psychological research. Although Spitczok von Brisinski (1994) introduced a method that achieved ultrafast display by reprogramming the registers, he could not produce an acceptable full-screen display. In this report, the author introduces a new method combining VESA routine calling with registers reprogramming that can yield a display at 640 x 480 resolution, with a refresh rate of about 150 Hz. PMID- 12109023 TI - A Web-accessible database of characteristics of the 1,945 basic Japanese kanji. AB - In 1981, the Japanese government published a list of the 1,945 basic Japanese kanji (Jooyoo Kanji-hyo), including specifications of pronunciation. This list was established as the standard for kanji usage in print. The database for 1,945 basic Japanese kanji provides 30 cells that explain in detail the various characteristics of kanji. Means, standard deviations, distributions, and information related to previous research concerning these kanji are provided in this paper. The database is saved as a Microsoft Excel 2000 file for Windows. This kanji database is accessible on the Web site of the Oxford Text Archive, Oxford University (http://ota.ahds.ac.uk). Using this database, researchers and educators will be able to conduct planned experiments and organize classroom instruction on the basis of the known characteristics of selected kanji. PMID- 12109024 TI - Visual-word identification thresholds for the 260 fragmented words of the Snodgrass and Vanderwart pictures in Spanish. AB - Word difficulty varies from language to language; therefore, normative data of verbal stimuli cannot be imported directly from another language. We present mean identification thresholds for the 260 screen-fragmented words corresponding to the total set of Snodgrass and Vanderwart (1980) pictures. Individual words were fragmented in eight levels using Turbo Pascal, and the resulting program was implemented on a PC microcomputer. The words were presented individually to a group of 40 Spanish observers, using a controlled time procedure. An unspecific learning effect was found showing that performance improved due to practice with the task. Finally, of the 11 psycholinguistic variables that previous researchers have shown to affect word identification, only imagery accounted for a significant amount of variance in the threshold values. PMID- 12109025 TI - Using Internet search engines to estimate word frequency. AB - The present research investigated Internet search engines as a rapid, cost effective alternative for estimating word frequencies. Frequency estimates for 382 words were obtained and compared across four methods: (1) Internet search engines, (2) the Kucera and Francis (1967) analysis of a traditional linguistic corpus, (3) the CELEX English linguistic database (Baayen, Piepenbrock, & Gulikers, 1995), and (4) participant ratings of familiarity. The results showed that Internet search engines produced frequency estimates that were highly consistent with those reported by Kucera and Francis and those calculated from CELEX, highly consistent across search engines, and very reliable over a 6-month period of time. Additional results suggested that Internet search engines are an excellent option when traditional word frequency analyses do not contain the necessary data (e.g., estimates for forenames and slang). In contrast, participants' familiarity judgments did not correspond well with the more objective estimates of word frequency. Researchers are advised to use search engines with large databases (e.g., AltaVista) to ensure the greatest representativeness of the frequency estimates. PMID- 12109026 TI - [Antibiotic therapy: state-of-the-art]. PMID- 12109027 TI - [Probiotics]. PMID- 12109029 TI - [Rational use of oral antibiotics in children and young people. Findings of an expert commission of the Paul Ehrlich Society for Chemotherapy ]. PMID- 12109030 TI - [On the paper, "Dieting for weight reduction", MMP 2002(25): 52-62]. PMID- 12109028 TI - [Rational use of oral antibiotics. Findings of an expert commission of the Paul Ehrlich Society for Chemotherapy]. PMID- 12109031 TI - [Initial experiences with adjuvant locoregional radioimmunotherapy using 131I labeled monoclonal antibodies against tenascin (BC-4) for treatment of glioma (WHO III and IV)]. AB - AIM: None of the established treatments (surgery, radiotherapy, chemotherapy) for malignant glioma has improved its very poor prognosis. Adjuvant locoregional radio-immunotherapy (RIT) represents a new therapeutic approach. We present our initial experience with this therapeutic tool with respect to adverse effects, biokinetics and clinical follow-up. METHODS: Following surgery and radiotherapy, 12 patients with glioma (4, WHO stage III; 8, WHO stage IV) underwent 1-5 RIT cycles (average dose 1100 MBq 131labelled monoclonal BC-4 antibodies) at six week intervals. Follow-up included serial FDG-PET and MRI investigations. Evaluation of biokinetics included whole body scans, together with analysis of blood, urine and fluid from the tumor cavity. RESULTS: Following RIT, four patients experienced temporary seizures, which, in one case, were associated with temporary aphasia. Eight patients developed HAMA (human anti-mouse antibodies) during follow-up. Mean biologic half-life of the radiopharmaceutical in the resection cavity was 3.9 d (range: 1.0-10.2 d) and remained stable intraindividually during further RIT-cycles. The antibody/radionuclide conjugate remained stable in the tumor cavity for at least 5 d. Median survival presently stands at 18.5 months compared to 9.7 months in a historical patient group (n = 89) undergoing conventional therapeutic strategies. Five patients show no signs of recurrence. In three patients with post-surgical evidence of residual tumor, one patient showed partial remission, one stable disease, and one progressive disease during RIT. Four patients without evidence of residual tumor mass at the beginning of RIT developed recurrence during therapy. CONCLUSIONS: Initial experience demonstrates that locoregional RIT is a well tolerated treatment modality that may represent a promising new approach in the management of patients with malignant glioma. Advantages of local application include passage of the blood-brain barrier, high concentration of activity within the resection cavity and low systemic toxicity. PMID- 12109032 TI - Minimal contribution of cell-bound antibodies to the immunoscintigraphy of inflamed joints with 99mTc-anti-CD4 monoclonal antibodies. AB - AIM: The cellular joint infiltrate in rheumatoid arthritis patients is rich in CD4-positive T-helper lymphocytes and macrophages, rendering anti-CD4 monoclonal antibodies (mAbs) suitable for specific immunoscintigraphy of human/experimental arthritis. Following intravenous injection, however, mAbs are present both in the free form and bound to CD4-positive, circulating monocytes and T-cells. Thus, the present study aimed at analyzing the relative contribution of the free and the cell-bound component to the imaging of inflamed joints in experimental adjuvant arthritis (AA). METHODS: AA rat peritoneal macrophages or lymph node l-cells were incubated in vitro with saturating amounts of 99mTc-anti-CD4 mAb (W3/25) and injected i.v. into rats with AA. RESULTS: In vitro release of 99mTc-anti-CD4 mAb from the cells was limited (on average 1.57%/h for macrophages and 0.84%/h for T cells). Following i.v. injection, whole body/joint scans and tissue measurements showed only negligible accumulation of radioactivity in inflamed ankle joints (tissue: 0.22 and 0.34% of the injected activity, respectively), whereas the radioactivity was concentrated in liver (tissue: 79% and 71%, respectively), kidney, and urinary bladder. Unlike macrophages, however, anti-CD4 mAb-coated T cells significantly accumulated in lymphoid organs, the inflamed synovial membrane of the ankle joints, as well as in elbow and knee joints. CONCLUSION: While the overall contribution of cell-bound mAbs to the imaging of arthritic joints with anti-CD4 mAbs is minimal, differential accumulation of macrophages and T-cells in lymphoid organs and the inflamed synovial membrane indicates preferential migration patterns of these 2 cell populations in arthritic rats. Although only validated for 99mTc-anti-CD4 mAbs, extrapolation of the results to other anticellular mAbs with similar affinity for their antigen may be possible. PMID- 12109033 TI - [Diagnosis of initial changes in the hand of patients with rheumatoid arthritis- comparison between low-field magnetic resonance imaging, 3-phase bone scintigraphy and conventional x-ray]. AB - Besides conventional X-rays, in the diagnostic work up of initial changes in patients suffering from rheumatoid arthritis (RA), 3-phase bone scintigraphy (3P Sz) is as well established as magnetic resonance imaging (MRI). The aim of this study was to compare the diagnostic value of the newly developed low field MRI with the proven methods X-rays and 3P-Sz. METHODS: 65 patients (47f, 18m; 20-86 yrs) were studied on a one day protocol with 3P-Sz (550 MBq Tc-99m DPD), MRI and X-rays of the hands. Images were visually analysed by two blinded nuclear medicine physicians and radiologists and classified as a) RA-typical, b) inflammatory, non-RA-typical and c) non inflammatory changes. All methods were compared to 3P-Sz as golden standard. RESULTS: In comparison to 3P-Sz, low field MRI presents with almost equal sensitivity and specificity in rheumatoid-typical and inflammatory changes. Conventional X-rays revealed in arthritis-typical changes as well as in inflammatory changes a significantly lower sensitivity and also a lower negative predictive value while specificity equals the one of MRI. Quantitative analysis of 3P-Sz using ROI-technique unveiled significantly higher values in patients with rheumatoid arthritis than in those with no inflammatory changes. CONCLUSION: MRI represents an equally sensitive method in the initial diagnosis of rheumatoid-typical and inflammatory changes in the region of the hands as compared to the 3P-Sz. Besides the basic diagnosis with conventional X rays, 3P-Sz is still the recommended method of choice to evaluate the whole body when RA is suspected. Additionally, quantitative analysis of the 3P-Sz using the ROI technique in the region of the hands reveals statistically significant results and should therefore be taken into account in the assessment of inflammatory changes. PMID- 12109034 TI - Value of tumour marker S-100B in melanoma patients: a comparison to 18F-FDG PET and clinical data. AB - AIM: The S-100B protein is commonly used in the immunohistochemical diagnosis of malignant melanoma and its metastases and has recently been introduced as a tumor marker in peripheral blood, whereas 18F-FDG PET is currently the most sensitive in-vivo imaging method for melanoma staging. Thus, the efficiency of serum S-100B and 18F-FDG PET in the detection of metastatic disease in melanoma patients are compared. METHODS: Serum S-100B was measured with a commercially available immunoradiometric assay. As part of primary tumour staging whole-body positron emission tomography (PET) with 18F labeled fluorodeoxy-D-glucose (18F-FDG) was performed in 67 patients suffering from cutaneous melanoma with a tumour thickness > 0.75 mm and a Clark-level III-V. Final diagnosis based on histology, morphologic imaging results and/or clinical follow-up after at least six months. RESULTS: No evidence of disease was seen in 43 of 67 patients (64.2%), 11 patients (16.4%) presented with lymph node metastases, 13 patients (19.4%) had one or more distant metastases. Alltogether, 18 of 67 patients showed S-100B values > 0.2 microgram/l, including two patients without metastatic disease, 3 of 11 patients with lymph node metastases, and the 13 patients with distant metastases. One patient showed false-positive FDG-uptake in the mediastinum, but presented with S-100B values off curve. CONCLUSION: Our data indicate that serum S-100B determination might be helpful in identifying melanoma patients with distant metastases. In comparison to 18F-FDG PET, the value of serum S-100B for lymph-node staging is limited. PMID- 12109036 TI - Myocardial fatty acid utilisation during exercise induced ischemia in patients with coronary artery disease. AB - AIM: Reversible or irreversible myocardial damage due to ischemia correlates with altered membrane functions of the cells. To compare myocardial free fatty acid (FFA) metabolism and flow during exercise induced ischemia we studied ten patients with coronary artery disease but without previous myocardial infarction. METHODS: A series of post-exercise single-photon emission computed tomography (SPECT) measurements was performed after injection of 123I labelled heptadecanoic acid (HDA). Myocardial perfusion was estimated from the separately performed exercise-redistribution thallium study. Fatty acid metabolic rate, thallium uptake and washout were calculated for anterior, lateral, posterior and septal segments. RESULTS: The more reduced post-exercise FFA metabolic rate (-63 +/- 18%, mean +/- 1 SD) compared to flow (-36 +/- 16%) was related to the severity of myocardial ischemia and wall motion abnormalities. CONCLUSION: In this small group of patients, the reduced post-exercise FFA metabolic rate tentatively suggests a parsimonious workload of the exercising myocardium by reducing oxygen consumption in patients with coronary artery disease. PMID- 12109035 TI - [Breast scintigraphy using 99mTc-sestamibi--use and limitations]. AB - AIM: Until now scintimammography did not achieve any definite role in the assessment of breast lesions. Purpose of this study was to elaborate its use as well as the limitations of scintimammography after 500 examinations completed. METHODS: Scintigraphic findings were correlated with the histopathologic outcome of 219 patients, who underwent surgery or biopsy for histopathological confirmation. The results were determined with respect to palpability of the lesion and tumour size. Additionally, a distinct analysis was performed for the patient subpopulation with indeterminate results of previously performed physical examination, mammography, and sonography. RESULTS: Overall sensitivity for scintimammography was 82.1% at a specificity of 87.5%. For palpable lesions sensitivity was 91.7% which was evidently higher as compared to 64.9% for non palpable lesions. For palpable lesions specificity was 81.1% and 88.6% for non palpable lesions. According to tumour size sensitivity ranged between 65.2% for carcinoma with a diameter < 1 cm and 93.7% for carcinoma > 1 cm. In the patients subgroup with indeterminate preliminary diagnosis (n = 143) sensitivity decreased to 71.7% at a specificity of 87.8%. Patients undergoing neoadjuvant chemotherapy showed decreasing sestamibi uptake as early as 8 days after therapy if tumour response was evident. However, small residual invasive tumours in patients with complete remission could not be visualised. CONCLUSION: Scintimammography is neither suited for screening, nor early diagnosis of breast cancer, nor for the further evaluation of small and unclear mammographic findings. Scintimammography should not be used whenever histopathological clarification of a suspicious lesion is necessary. It is useful to further investigate patients with unclear or probably benign findings in physical examination and/or mammography and to monitor tumour response to neoadjuvant chemotherapy. PMID- 12109037 TI - [Certification of an RIA laboratory according to DIN EN ISO 9001-2000]. AB - The university hospital Freiburg intends to establish step by step a total quality management system (TQMS) in all facilities with an external certification as its final aim. This has been reached already in the central laboratory. Therefore, it is effective for the department of nuclear medicine to do the same for its own lab specialized in thyroid hormones. The TQMS has been built up within six months on the basis of DIN EN ISO 9001:2000. This internationally accepted standard is of uttermost economical importance for an institution situated near the French and Swiss border. This review describes the procedure in detail. Responsible for the effort were the engagement of the coworkers in the lab as well as an adequate choice of the external adviser and the authority for certification. The procedure is formalized to a high degree and much understanding for analytic thinking and systematization is needed. Now it remains to be seen, if the desired effect (increased understanding for quality within the department, more efficient performance of the lab, better acceptance by the clients) will arise. PMID- 12109038 TI - [Recurrence of a renal cell carcinoma mimicking a multiple endocrine neoplasia with thyroid gland involvement]. PMID- 12109039 TI - [Multimodal therapy concept in advanced distant metastasis of a follicular thyroid carcinoma]. PMID- 12109040 TI - [On the unusual cae: SAPHO syndrome--results of 2-phase scintigraphy (Ncklearmedizin 2001; 40: N48-50)]. PMID- 12109041 TI - From toxic precursors to safe drugs. Mechanisms and relevance of idiosyncratic drug reactions. AB - In adverse drug reactions, effects accounted for by the pharmacological profile of the drug may be distinguished from sometimes bizarre, unpredictable events ("idiosyncrasies"). In the majority of cases, the latter are due to reactive intermediates formed by members of the cytochrome P450 family of enzymes. These products are usually formed at very low quantities. However, genetic disposition (e.g. by enhanced expression of the catalyzing enzyme or failure of protective factors such as glutathione) or certain external conditions (for example comedication of inducers of drug metabolism) may lead to toxic intermediates being available at relevant quantities. Once generated, these metabolites will react with macromolecules and finally cause cell necrosis. Necrosis may result from direct damage to functions essential to the cell or be secondary to harmful immune reactions activated by recognition of the new structures as neoantigens. As the principal site of drug metabolism, the liver is most frequently affected by idiosyncratic reactions, which often enough are recognized only after large numbers of patients have been treated. The idiosyncratic potential of a drug relates to its chemical structure rather than its pharmacological mechanism. Risks are evident if biotransformation yields products with chemical substructures such as quinones, phenoles, acyl halides, aromatic and hydroxyl amines. There are numerous cases of precursor drugs with idiosyncratic potential that have been replaced or marginalized by agents with more favorable chemical properties. Examples include beta 1-preferential beta-adrenoceptor antagonists (toxic precursor: practolol) and the choline esterase inhibitors used in treatment of Alzheimer's disease. In the latter group, tacrine (CAS 321-64-2) gives rise to high liver enzyme levels in about 30% of the patients, an effect not shared with more recent, chemically different entities. A similar case can be made for the insulin sensitizer troglitazone (CAS 97322-87-7) (marketed, now withdrawn in the USA) and its successors rosiglitazone (CAS 122320-73-4) and pioglitazone (CAS 111025-46-8). Due to its different chemical structure, only troglitazone can be transformed into a reactive quinone metabolite, a process that may be accelerated by the drug inducing its own metabolism. In agreement with this view, there is no evidence of specific hepatic toxicity with the two successor drugs. Ongoing progress in molecular toxicology may aid in minimizing idiosyncratic hazards already at early stages of drug development. PMID- 12109043 TI - Effects on the reactive oxygen species of erdosteine and its metabolite in vitro. AB - Erdosteine (CAS 84611-23-4) and its active metabolite (M1) decrease the luminol dependent chemiluminescence (LDCL). In this study, the scavenging effects of erdosteine and M1 on each reactive oxygen species, hypochlorous acid (HOCl), hydrogen peroxide (H2O2) and superoxide anion (O2-), were investigated. M1 interacted with HOCl at high activity, whereas erdosteine exhibited low activity. Although erdosteine interacted with H2O2 only at 1 mmol/l, M1 could significantly decrease H2O2 (> 0.1 mmol/l). Erdosteine and M1 did not interact with O2-. These results suggest that M1 possesses scavenging activities against H2O2 and HOCl in vitro. PMID- 12109042 TI - Pharmacokinetics of pipamperone from three different tablet formulations. AB - Twenty-four (24) Caucasian male subjects completed a single-blind, randomised, three-treatment, three-period, cross-over study. In each treatment phase, subjects received a single dose of 144 mg pipamperone dihydrochloride (CAS 2448 68-2) (equivalent to 120 mg pipamperone; CAS 1893-33-0) as either the reference product (3 x 40 mg tablets), test product A (3 x 40 mg tablets) or test product B (1 x 120 mg tablet). Each consecutive dosing was separated by a washout period of 14 days. Following each dosing, venous blood samples were collected over a period of 120 h for the determination of plasma pipamperone concentrations by high performance liquid chromatography. The most common drug related adverse events, ranging from mild to moderate in intensity, were bloodshot eyes, nasal congestion, dry mouth, hypotension and dizziness. The geometric mean Cmax of pipamperone for both the reference product and test product A was 266 ng/ml and for test product B 263 ng/ml. The geometric mean AUC0-infinity was 3107 ng.h/ml for the reference product, 3229 ng.h/ml for test product A and 3108 ng.h/ml for test product B. The two test products were shown to be bioequivalent to the reference product with respect to all pharmacokinetic variables investigated. PMID- 12109044 TI - Mechanism of the antitussive effect of azelastine in guinea pigs. AB - In order to clarify the mechanism of the antitussive effects of azelastine hydrochloride (azelastine, CAS 790307-93-0), the cough responses to inhaled capsaicin and substance P (SP) were evaluated before and after the administration of azelastine in conscious guinea pigs. The concentrations of SP were also measured before and after the administration of azelastine by radioimmunoassay in anesthetized guinea pigs. Capsaicin and SP caused coughing in conscious guinea pigs in a dose-dependent fashion. After the treatment with azelastine, capsaicin induced cough decreased significantly, and the dose-response curve to capsaicin was shifted to a higher concentration in comparison with the the controls. SP induced cough was not inhibited by the treatment with azelastine, and the dose response curves to SP did not change. The concentrations of SP recovered in bronchoalveolar lavage fluid and in the trachea were decreased statistically significantly in a dose-dependent manner after the treatment with azelastine, while the SP concentrations of the subjects not treated with azelastine were not inhibited. These results suggest that the antitussive effect of azelastine might be partly due to the inhibition of SP release from sensory nerves in guinea pigs. PMID- 12109045 TI - Synthesis and activity of phenyl derivatives containing 5,6-dimethylthieno[2,3 d]pyrimidin-4(1H)-one or 4H-pyrimido[5,4-b]indol-4-one heterocyclic system as potential nonsteroidal anti-inflammatory drugs. AB - The synthesis, the analgesic and anti-inflammatory activities of two series of phenyl derivatives containing 5,6-dimethyl-thieno[2,3-d]pyrimidin-4(1H)-one and 4H-pyrimido[5,4-b]indol-4-one system, respectively, are reported. Two of these derivatives, 6A and 9B, showed interesting activities. The results of the pharmacological assays are discussed. PMID- 12109046 TI - Studies on new cyclic imides obtained from aminophenazone with analgesic properties. Potent effects of a 3,4-dichloromaleimide derivative. AB - This paper describes the synthesis of new cyclic imides obtained by reaction with aminophenazone (CAS 58-15-1, 4-aminoantipyrine) and different anhydrides with further cyclization with acetic acid under reflux. Their structures were confirmed by spectral data (IR and NMR) and elemental analysis. The analgesic activity of the synthesized compounds was investigated initially with the writhing test in mice and the most promising compound, a 3,4-dichloromaleimide derivative (3), was analyzed using other models of nociception. The results indicated that compound 3 exerts potent analgesic activity in mice, being more active than some reference drugs. The analgesia caused by this compound was not reversed by naloxone in the writhing test. In the hotplate test, compound 3 did not increase the latency period of pain induced by thermal stimuli, confirming that it does not interact with opioid systems. PMID- 12109047 TI - Evaluation of the analgesic and anti-inflammatory activities of some thiazolo[4,5 d]pyrimidines. AB - Some thiazolo[4,5-d]pyrimidine-7(6H)-one derivatives were evaluated in vivo for their analgesic and anti-inflammatory activities. The results were compared with that of acetyl salicylic acid and phenylbutazone. Compounds 3b and 3h were the most active in the anti-inflammatory paw edema inhibition test. In terms of the analgesic activity (acetic acid writhing test), the most active compound was 2a followed by 31. The most active members of the series were investigated for their ED50 values and ulcerogenic potential. PMID- 12109048 TI - Effects of gold-chloroquine complexes on respiratory burst of polymorphonuclear leukocytes. AB - The effects of gold-chloroquine derivatives with the formula [Au(PR3)(CQ)]PF6 (where R = Ph (1), Et or Me) on the superoxide anion production by human neutrophils (PMNs, polymorphonuclear cells) were investigated. When these complexes (0.1-3 mumol/l) were added to PMNs prior to the activators formyl methionyl-leucyl-phenylalanine (fMLP) or phorbol myristate acetate (PMA), they inhibited isoluminol-horseradish peroxidase-dependent chemiluminescence (CLisol). The inhibition was a direct result of effects on PMNs since chemiluminescence in the cell-free system (horseradish peroxidase-hydrogen peroxidase-isoluminol) was not affected. The above mentioned concentrations of the complexes did not show in vitro toxicity on the cells. On the other hand, when 1 mumol/l of complex 1 was added to cells after stimulation, the chemiluminescence of PMNs stimulated by PMA was inhibited, but not the chemiluminescence stimulated by fMLP. The gold chloroquine binding was essential for the referred activity as chemiluminescence was not influenced by the precursors chloroquine (CAS 54-05-7) and AuCl(PPh3). Furthermore, the extent of inhibition of chemiluminescence in PMNs activated by PMA did not increase with the duration of preincubation in presence of 1 mumol/l of 1. Extensive washing of cells after preincubation with 1 mumol/l of 1 reversed the aforementioned inhibition. All these results show that the gold-chloroquine binding can lead to compounds with specific properties that could make them useful in the treatment of some inflammatory diseases. PMID- 12109049 TI - Synthesis and antitumour activity of a series of cyclic amidines of 2,3-dihydro 1H-1,5-benzodiazepines. AB - 2,3-Dihydro-1H-1,5-benzodiazepine amidines were prepared by nucleophilic substitution of 2,3,4,5-tetrahydro-1H-1,5-benzodiazepine-2-thiones. Evaluation of the 13 synthesised amidines as antitumour agents was carried out in vitro against 60 human tumour cell lines at the National Cancer Institute, Bethesda, USA. The screening revealed a moderate cell growth inhibition of two derivatives on all cell lines at concentrations ranging from 10(-5) to 10(-4) mol/l. Log P values were theoretically calculated. The more active derivatives were found to exhibit a higher lipophilicity. PMID- 12109050 TI - Synthesis and antimalarial activity of substituted pyrazole derivatives. AB - The development of new antimalarial drugs is an urgent priority considering the increasing prevalence of drug-resistant Plasmodium falciparum parasites. A series of pyrazoles are described as part of efforts directed toward the synthesis of some potent antimalarial agents. The replacement of the ester group as a substituent in the pyrazole ring by nitrile group caused a precipitous loss of activity as antimalarial due to the lack of hydrogen-bond formation. Further modification of the heterocyclic ring to give substituted aryl derivatives afforded potent antimalarial derivatives: methyl 5-amino-3-anisidinepyrazole-4 carboxylic acid 3a (IC50: 0.149 mumol/l) and methyl 5-amino-3-(m anisidin)pyrazole-4-carboxylic acid 3c (IC50: 0.15 mumol/l). The synthesis, structure-activity relationships (SAR), X-ray crystallography and pharmacological activity associated with these series of compounds are discussed. PMID- 12109051 TI - Effects of amidine derivatives on parasite-macrophage interaction and evaluation of toxicity. AB - In this work, the effect of amidine derivatives (with bromine and methoxy as substituents) in the "in vitro" parasite-macrophage interaction was evaluated. The potential toxicity was also analyzed. The results show that the methoxy derivative was able to decrease the percentage of "in vitro" infection, being not hazardous to the host cell. Furthermore, experiments using Balb/c mice showed that this compound was very effective in avoiding infection in these animals. On the other hand, the compound with bromine as substituent was toxic to macrophages and unable to prevent infection in Balb/c mice. Pentamidine isethionate, used as reference drug, was not efficient in both experiments. PMID- 12109053 TI - Guiding principles for human immunodeficiency virus (HIV) testing of women during pregnancy--2002. PMID- 12109052 TI - Comparison between the mechanisms of action of plant toxins ricin and viscumin on the stage of intracellular dissociation. AB - Pharmacological effects of mistletoe extracts are determined by the concentration of three toxic lectins: mistletoe lectin I (MLI, or viscumin), MLII, MLIII. These proteins, as well as ricin, belong to ribosome-inactivating proteins type 2 (RIP2). However, the extracts from the plant Ricinus communis, containing ricin, are highly toxic. Ricin is about 30 times more effective in cell culture than viscumin. The dissociation of subunits and the transmembrane transport of catalytic subunit into the cytoplasm are needed to obtain the cytotoxic effect of RIP2. In this paper, hybridomas producing monoclonal antibodies against catalytic subunits of ricin and viscumin are described. Monoclonal antibodies against different epitopes, including one localized in intra-subunit area of catalytic subunits of ricin and viscumin, do not inhibit the enzymatic activity of these proteins in cell-free system. These hybridomas are resistant to the cytotoxic action of native toxins. Protective effect of antibodies are about the same for both toxins, though the dissociation of the subunits of ricin is more effective. The causes of the differences in activity of plant toxins as pharmacological agents, and the importance of above mentioned epitopes for neutralizing antibodies at the clinical applications of mistletoe extracts are discussed. PMID- 12109054 TI - Neurologic illness associated with eating Florida pufferfish--2002. PMID- 12109055 TI - [Skin manifestations in kidney diseases]. AB - Skin irritations in conjunction with renal disorders are often specific. Therefore a close inspection of the integument may point to an internal organ pathology. A selection of hereditary syndromes and a summary table are shown including acquired diseases displaying symptoms of the skin and the kidneys. A special emphasis is laid on specific and unspecific skin alterations under chronic renal failure. Instead of a detailed list of therapeutic possibilities, priority is given to the exact description of macroscopic and microscopic skin alterations including pathophysiologic mechanisms. PMID- 12109056 TI - [Specific and nonspecific skin manifestations in leukemia]. AB - The cutaneous manifestations of leukemias are conventionally divided into nonspecific benign lesions and specific malignant lesions. Specific lesions (leukemia cutis) are localized or disseminated infiltrations of the skin by malignant leukemic cells which may involve all layers of the skin. The clinical appearance of leukemia cutis is variable and may range from papules and nodules to a generalized cutaneous eruption and erythroderma. The histopathologic examination of the skin lesion is essential for diagnosis of leukemia cutis. Specific skin lesions are usually observed in patients with an aggressive clinical course and are associated with a poor prognosis. However, an overall survival of patients with specific skin lesions of chronic lymphocytic leukemia is significantly better, as compared with other types of leukemia. Rarely, skin lesions containing leukemic cells are present before evidence of leukemia cutis can be detected in the peripheral blood and bone marrow (aleukemic leukaemia cutis). Leukemic skin lesions should be differentiated from numerous nonspecific lesions, which may be present in up to 80% of all patients with leukemia. PMID- 12109057 TI - [Adverse chemotherapy effects on skin and mucous membranes]. AB - Mucocutaneous side effects of chemotherapeutic agents are frequent, especially alopecia and stomatitis. The latter can be dose limiting or can endanger the patient in case of secondary infection. Nail changes and hyperpigmentation are cosmetically disturbing. For the diagnosis of the acral erythrodysesthesia syndrome and the neutrophilic eccrine hidradenitis knowledge of the clinical presentation is required. Interaction of chemotherapeutic agents with radiotherapy or UV light can aggravate or cause skin changes like the radiation recall. Despite the completely different mode of action of new substances or immunotherapeutic agents like monoclonal antibodies the same range of side effects can be observed. PMID- 12109058 TI - [Unusual etiology of erythema nodosum, pleural effusion and reactive arthritis: Giardia lamblia]. PMID- 12109059 TI - Gene therapy to overcome drug resistance in cancer: targeting key regulators of the apoptotic pathway. AB - A better understanding of the molecular events responsible for the development of drug resistance in cancer cells has emerged in recent years. It is now established that tumor cells can acquire drug resistance by alterations of pathways involved in the regulation of apoptosis and that failure to activate this pathway in cancer cells may confer resistance to chemotherapy. This resistance to drug-induced apoptosis is likely to play an important role in tumors that are refractory to chemotherapy. The identification of points in the apoptotic pathway at which dysregulation occurs opens up new therapeutic opportunities in situations where conventional cytotoxic chemotherapy approaches fail. Although these gene therapy-based strategies are still in their infancy they will likely lead to more effective treatments for human cancers. This review will focus on gene therapy strategies developed to specifically target the apoptotic pathway and how these strategies can affect the sensitivity of tumor cells to chemotherapy. PMID- 12109060 TI - T lymphocytes as targets of gene transfer with Moloney-type retroviral vectors. AB - Peripheral T lymphocytes are a target of choice for many gene therapeutic strategies. Retrovirus-mediated transduction allows genomic integration and long term expression of transgenes in target cells. Over many years, low transduction efficiency into primary T lymphocytes has limited clinical application of existing protocols. Recently, gene transfer rates > 50% have been achieved facilitating clinical studies. More attention is thus being focused on the ability of gene-modified cells to carry out innate as well as conferred functions in vivo and the influence of culture conditions, retroviral vector and host response thereon. PMID- 12109061 TI - Herpes simplex virus vectors for the nervous system. AB - Herpes simplex virus type 1 (HSV1) has a number of properties which could potentially be exploited in the development of vectors for the delivery of genes to the nervous system. These include a natural tropism for neurons, a large viral genome allowing the insertion of multiple exogenous genes, and the ability to establish asymptomatic life-long latent infections. Despite these inherent advantages, the development of HSV vectors successfully exploiting all these properties has been problematical. This has mainly been due to either vector toxicity or an inability to maintain transgene expression in the long term. Recent progress towards overcoming these problems and several applications of the technology are discussed. PMID- 12109062 TI - Chemoprotection and selection by chemotherapy of multidrug resistance-associated protein-1 (MRP1) transduced cells. AB - Utilization of chemotherapy for treatment of tumors is mainly limited by its hematological toxicity. Because of the low level expression of drug resistance genes, transduction of hematopoietic progenitors with multidrug resistance 1 (MDR1) or multidrug resistance-associated protein 1 (MRP1) genes should provide protection from chemotherapy toxicity. Successful transfer of drug resistance genes into hematopoietic cells might allow the administration of higher doses of chemotherapy and, therefore, increase regression of chemosensitive tumors. In addition, this approach can be used to select in vivo transduced cells by their enrichment after administration of cytotoxic drugs. Our group has studied the potential value of MRP1 to protect hematopoietic cells. The interest in the use of MRP1 as an alternative to MDR1 gene transfer for bone marrow protection lies in its different modulation. Indeed, classical P-gp reversal agents, tested in clinic to decrease MDR1 tumor resistance, have little or no effect on MRP1 function. This would allow, in the same patient, the use of reversal agents to decrease P-gp tumor resistance without reversing bone marrow protection of the transduced hematopoietic cells provided by MRP1. We constructed two different MRP1-containing vectors with either the Harvey retroviral long terminal repeat (LTR) or phosphoglycerate kinase (PGK) as promoters and generated ecotropic producer cells. MRP1 transduced fibroblasts were more resistant to doxorubicin, vincristine, and etoposide and their chemoprotection was increased after selection with chemotherapeutic agents in the presence of glutathione, a co factor for MRP1 function. Lethally irradiated mice were engrafted with bone marrow (BM) cells transduced with MRP1 vectors (PGK promoter). We demonstrated that high expression of MRP1 in murine hematopoietic cells reduces doxorubicin induced leukopenia and mortality. In addition, in vivo selection of MRP1 transduced BM cells was achieved following doxorubicin administration and allowed a better chemoprotection after the second chemotherapy cycle. PMID- 12109063 TI - Vector delivery methods and targeting strategies for gene therapy of brain tumors. AB - Efficient virus and non-virus vector systems for gene transfer to tumor cells have been developed and tested in cell culture and in animal experiments. With some of the earliest and most comprehensively evaluated vectors, such as retroviruses, advanced clinical trials were performed in tumor patients. Malignant primary brain tumors (gliomas) have been chosen for the first clinical studies on novel gene therapy approaches because these tumors are non-metastatic and develop on the largely postmitotic background of normal glial and neuronal tissue. However, the human cancer gene therapy studies performed so far were not as successful as preclinical animal experiments. Furthermore, the clinical studies did not address major limiting factors for in vivo gene therapy, such as insufficient gene transfer rates to the tumor with the used local delivery modalities, and the resulting inability of a particular transgene-prodrug system to confer permanently eradicating cytotoxicity to the whole neoplasm. Critical evaluation of gene transfer and therapy studies has led to the conclusion that, even using identical vectors, the anatomical route of vector administration can dramatically affect both the efficiency of tumor transduction and its spatial distribution, as well as the extent of intratumoral and intracerebral transgene expression. This review concentrates on different physical methods for vector delivery to malignant primary brain tumors in experimental or clinical settings: stereotactic or direct intratumoral injection or convection-enhanced bulk-flow interstitial delivery; intrathecal and intraventricular injection; and intravascular infusion with or without modification of the blood-tumor-barrier. The advantages and drawbacks of the different modes and delivery routes of in vivo vector application, and the possibilities for tumor targeting by modifications of the native tropism of virus vectors or by using tissue-specific or inducible transgene expression are summarized. PMID- 12109064 TI - Genetic approaches for antigen-selective cell therapy. AB - Attempts to improve the efficacy of adoptive T-cell therapies have led to the development of innovative strategies that combine the high specificity of antibody molecules with the efficient trafficking properties and effector functions of immune cells. These antigen-selective cell therapies are designed to convert therapeutically important native antigens expressed on the cell surface (tumor associated antigens, viral envelope proteins) into recruitment points of effector functions, and address the goal of major histocompatibility complex- and exogenous cytokine-independent activation of mature effector T-cells. The most promising and best characterized antigen-selective strategy is based on the genetic manipulation of the recognition specificity of T-cells by grafting the recognition specificity of a monoclonal antibody onto a lymphocyte triggering molecule (TCR-associated polypeptides, Fc epsilon RI-gamma chain). Upon encountering specific antigen, cells harboring chimeric immune receptors (CIRs) are able to undergo specific stimulation and kill antigen bearing cells in both in vitro and in vivo model systems. Initial studies have focused on terminally differentiated effector cell-based protocols. However, recent data indicate that progenitor cell-based therapies allow the permanent generation of stable populations of CIR-expressing cells of multiple lineages, leading to long-term persistent systemic immunity. Emerging gene therapy strategies are based on the use of biespecic antibody fragments. The advantages of these biespecic antibody mediated immune recruitment (BIR) approaches (trans-recruitment and multieffector activation) could complement conventional CIR-based immunotherapies. Although further scientific progress is required regarding the selection of the ideal effector cell/s and the definition of the optimal targeting and recruitment systems, clinical trials recently initiated in patients with advanced cancer and human immunodeficiency virus infection should help us to determine the real efficacy of these approaches. The relevance of these and other emerging concepts to cell-mediated immunotherapy is discussed. PMID- 12109066 TI - An unprecedented example of a cis-phosphonodithioato nickel(II) complex built by an extensive hydrogen bonding supramolecular network. AB - The P-N bond hydrolysis of the 4-methoxyphenyl-ammonium ethylamido phosphonodithioato ligand during its complexation to NiII leads to the first example of phosphonodithioato nickel(II) complex having a cis configuration; this complex is stabilised in the solid state by an extensive and intricate network of hydrogen bondings involving the released ethylenediamine and a water molecule. PMID- 12109065 TI - Morphine, the Proteus of organic molecules. AB - This feature article encapsulates the senior author's longstanding interests in opiate chemistry and attempts to place it within an historical context and against the backdrop of related work by others who have viewed morphine as one of the pinnacles of natural product synthesis. Biomimetic and 'bioanalogous' routes to the morphine skeleton are discussed followed by approaches based on the elaboration of phenanthrene platforms. The latter include an asymmetric synthesis of ent-morphine developed in our laboratory. PMID- 12109067 TI - A novel synthetic route to fused propenochlorin and benzochlorin photodynamic therapy probes. AB - Reaction of meso-(2-formylvinyl)octaethylporphyrin with (CH3)3SiCN-Cu(OTf)2 produced unexpected 10(3)-trimethylsiloxyl and 10(3)-hydroxyl fused propenochlorins which, in H2SO4, underwent subsequent migration of the 8-ethyl group to the 10(3)-position of the exocyclic benzene ring to form a novel benzochlorin. PMID- 12109068 TI - The role of solvent in sonochemical reactions: the case of acetic acid. AB - Among the sonolysis products of acetic acid, oxidizing intermediates can play a direct role in reactions run in this solvent, illustrating the fact that organic sonochemistry can originate in organic solvents as aqueous sonochemistry can in water. PMID- 12109069 TI - Alkoxide binding in inverse crown chemistry: rational synthesis of a series of composite alkali metal-magnesium-alkoxide-diisopropylamides. AB - A series of composite lithium- or sodium-magnesium-alkoxide-diisopropylamides of general formula [(MMg[N(Pri)2]2OR)2] (where M = Li, R = Octn; M = Na, R = Bun or Octn), has been synthesised rationally by treating synergic amide mixtures with the appropriate alcohol. PMID- 12109070 TI - Heteropoly acid as a novel efficient catalyst for Fries rearrangement. AB - Heteropoly acid H3PW12O40 is a very efficient and environmentally benign catalyst for the Fries rearrangement of phenyl acetate in homogeneous or heterogeneous liquid-phase systems at 100-150 degrees C. PMID- 12109071 TI - Synthesis and crystal structure of the first scandium-containing open framework solid. AB - A novel open framework scandium sulfate phosphate is prepared hydrothermally in the presence of the azamacrocycle cyclen (1,4,7,10-tetraazacyclododecane) in which secondary building units of formula Sc7(S,P)12O48 are linked to give a structure with supercages. PMID- 12109072 TI - Oxidation of lignin model compounds by organic and transition metal-based electron transfer mediators. AB - We have studied the oxidation of lignin model compounds by organic and transition metal-based mediators using either an enzyme or an electrolysis cell as the mediator oxidizing agent. Electrolysis of inorganic mediator seems a promising technology for pulp delignification. PMID- 12109073 TI - Breakthrough in the direct conversion of methane into c1-oxygenates. AB - The partial oxidation of methane to CH3OH and HCHO (C1-oxygenates) was evaluated over a low surface area V2O5/SiO2 catalyst. The introduction of low amounts of NO (0-2.92% vol) to the reaction feed strongly enhanced both the conversion of methane and selectivity to C1-oxygenates. In the presence of NO, both the reaction temperature and the CH4/O2 ratio affected selectivity to CH3OH and HCHO. Selectivity values of C1-oxygenates as high as 40% at a methane conversion close to 40% were obtained. PMID- 12109074 TI - Microwave assisted template removal of siliceous porous materials. AB - Organic templates of meso- and macro-porous siliceous materials can be completely removed within minutes by microwave digestion, resulting in highly ordered inorganic frameworks with higher surface areas, larger pore volumes, lower structural shrinkage and richer silanol groups compared with those from conventional template removal methods. PMID- 12109075 TI - The isolation and secondary functionalisation of fac-tris-2,2'-bipyridine complexes of ruthenium(II). AB - fac-Ruthenium(II) tris-(5-carboxy-2,2'-bipyridine) has been synthesised as a single geometric isomer for the first time, and proves to be a good 'building block' to introduce new functionality with retention of the isomeric integrity. PMID- 12109076 TI - First successful reaction of a silyl anion with hafnium tetrachloride. AB - Hafnium tetrachloride reacts with the tris(trimethylsilyl)silyl potassium tmen adduct (1) to form a [tris(trimethylsilyl)silyl]trichlorohafnium tmen complex (2); reaction of 2 with 2,6-dimethylphenylisonitrile leads to insertion into the silicon hafnium bond (4). PMID- 12109077 TI - The stable radical cation of thiophene annelated with bicyclo[2.2.2]octene and its reaction with triplet oxygen to give a protonated cation of 2-butene-1,4 dione derivative. AB - The first isolable salt of the thiophene radical cation was prepared from the derivative annelated with two bicyclo[2.2.2]octene units, and its reaction with triplet oxygen was found to give a novel cation of a proton-chelating 2-butene 1,4-dione derivative with remarkable stability. PMID- 12109078 TI - Interplay of anions and ligands on the nature and reducibility of NiOx/Al2O3 catalysts prepared by impregnation. AB - When NiOx/Al2O3 catalysts (Ni wt% = 1.5) are prepared by impregnation using [NiL2(H2O)2]X2 as precursors (L = diamine, X = Cl- or NO3-), a supported oxidic or metallic phase can be selectively obtained after thermal treatment in N2 depending on the nature of the ligand and counter anion; the oxidic phase can be reduced at a lower temperature than the classical nickel aluminate phase obtained from [Ni-(H2O)6](NO3)2. PMID- 12109079 TI - Synthesis and characterisation of the first carbene and diazabutadiene-indium(II) complexes. AB - The reactions of IMes [:CN(Mes)C2H2N(Mes), Mes = mesityl] and DAB [(ArN=CH)2, Ar = C6H3Pri2-2,6] with indium(I) halides have afforded the first carbene and diazabutadiene indium(II) complexes, [In2Br4(IMes)2] and [In2Cl2(DAB.)2], both of which have been crystallographically characterised. PMID- 12109080 TI - Highly saddle shaped (porphyrinato)iron(III) iodide with a pure intermediate spin state. AB - Combined analyses using NMR, EPR and Mossbauer spectroscopy as well as SQUID magnetometry have revealed that highly saddle shaped Fe(OETPP)I adopts an essentially pure intermediate spin state in spite of the coordination of an iodide ligand. PMID- 12109081 TI - Novel tungsten calix[8]arene complexes. AB - p-tert-Butylcalix[8]areneH8 reacts with WCl6 forming a number of new metallocalix[8]arenes, one of which is reduced to give the first example of a metal-metal triple bond supported by a large calixarene ligand; the degree of metallation and conformations adopted by the newly generated metallocalix[8]arene systems are determined by single crystal X-ray diffraction. PMID- 12109082 TI - Hierarchical interlinked structure of titanium oxide nanofibers. AB - A novel hierarchical interlinking structure of titanium oxide nanofibers with high surface area has been prepared by a soft hydrothermal chemical process via the reactions of amorphous TiO2 gel (or commercial TiOSO4 particles) and NaOH solution. PMID- 12109083 TI - A one-step synthesis of fused pentathiepins. AB - Treatment of nucleophilic heterocycles like pyrroles and thiophene, and their tetrahydro derivatives, with S2Cl2 and a base in chloroform at room temperature provides a simple one-pot synthesis of heterocyclic fused mono and bis pentathiepins such as 2, 3, 4, 5, 9, and 11. PMID- 12109084 TI - New efficient catalyst for ammonia synthesis: barium-promoted cobalt on carbon. AB - Barium-promoted cobalt catalysts supported on carbon exhibit higher ammonia activities at synthesis temperatures than the commercial, multipromoted iron catalyst and also a lower ammonia inhibition. PMID- 12109085 TI - A well-defined magnesium enolate initiator for the living and highly syndioselective polymerisation of methylmethacrylate. AB - The reaction of (BDI)MgiPr [BDI = HC(C(Me)=N-2,6-iPr2C6H3)2] with 2',4',6' trimethylacetophenone in toluene affords the enolate complex [(BDI)Mg(mu-OC(=CH2) 2,4,6-Me3C6H2)]2 which is found to be an excellent initiator for the living, syndioselective (sigma r > 0.95) polymerisation of methyl methacrylate. PMID- 12109086 TI - A novel dizinc bridged hydroxamate model for hydroxamate inhibited zinc hydrolases. AB - Reaction of Zn(OAc)(2).2H2O with tmen leads to the formation of [Zn(tmen)(OAc)2] (I) which reacts with benzohydroxamic acid to form Zn(BA)2.H2O (II) and the novel dizinc hydroxamate bridged complex [Zn2(mu-OAc)2(OAc)(mu-BA)(tmen)] (III), which may also be prepared by self-assembly and whose structure closely mimics that of the native hydroxamate inhibited Aeromonas proteolytica aminopeptidase. PMID- 12109087 TI - Double helical silica fibrils by sol-gel transcription of chiral aggregates of gemini surfactants. AB - Silica fibrils with a novel double stranded helical structure are prepared by sol gel transcription of twisted bilayer ribbons formed by cationic gemini surfactants. PMID- 12109088 TI - The total synthesis of alkaloids (-)-histrionicotoxin 259A, 285C and 285E. AB - The first total syntheses of three "unsymmetrical" (i.e. different terminal groups in the side chains) members of the histrionicotoxin family of alkaloids have been accomplished via stepwise introduction of the two side chain moieties onto a common tricyclic core. PMID- 12109089 TI - Simultaneous introduction of chemical and spatial effects via a new bimodal catalyst support preparation method. AB - Nano-sized zirconia-silica bimodal catalyst supports are prepared by direct introduction of zirconia sol into silica gel, which improved supported cobalt catalyst activity significantly via a spatial effect and a chemically promotional effect of zirconia in liquid-phase Fisher-Tropsch synthesis (FTS). PMID- 12109090 TI - Anion sensing with luminescent lanthanide complexes of tris(2 pyridylmethyl)amines: pronounced effects of lanthanide center and ligand chirality on anion selectivity and sensitivity. AB - Luminescent lanthanide complexes with tris(2-pyridylmethyl)amine ligands exhibited anion-specific sensory functions, and their anion selectivity and response sensitivity were modulated by a combination of lanthanide center and chiral ligand. PMID- 12109091 TI - First direct observation of a CO-bridged primary photoproduct of [Ru3(CO)12] by picosecond time-resolved IR spectroscopy. AB - For the first time, a CO-bridged primary photoproduct was observed for [Ru3(CO)12] by using picosecond time-resolved IR spectroscopy (ps-TRIR). PMID- 12109092 TI - Electric field induced cis-to-trans isomerization of polyphenylacetylene in solid state. AB - A field induced isomerization from cis to trans form in stereoregular cis-rich polyphenylacetylenes (PPAs) was found, and it provides an alternate method to control the order of chromophores in thin solid films. PMID- 12109093 TI - Self-assembly of an organometallic side-by-side double helix. AB - The first polymeric organometallic double helix has been synthesized by self assembly through hydrogen bonding by using a biomimetic strategy and a new side by-side structural motif. PMID- 12109094 TI - The fluorine gauche effect. Langmuir isotherms report the relative conformational stability of (+/-)-erythro- and (+/-)-threo-9,10-difluorostearic acids. AB - (+/-)-Erythro- and (+/-)-threo-9,10-difluorostearic acids, which differ only by a stereogenic interconversion of a single C-F bond, have significantly different conformational stabilities. PMID- 12109095 TI - Calix[4]arene based dendrimers. AB - The synthesis of calix[4]arene based dendrimers containing up to seven calix[4]arene moieties is described, including the X-ray crystal structure of a tris-calix[4]arene branching derivative. PMID- 12109097 TI - N2O decomposition over Fe/ZSM-5: reversible generation of highly active cationic Fe species. AB - Fe-oxide species in Fe/ZSM-5 (prepared by chemical vapor deposition of FeCl3)- active in N2O decomposition--react with zeolite protons during high temperature calcination to give highly active cationic Fe species, this transformation being reversible upon exposure to water vapor at lower temperature. PMID- 12109096 TI - Synthesis of planar chiral (1,3-disubstituted arene) Mn(CO)3+ cations via addition of nucleophiles to (oxocyclohexadienyl)Mn(CO)3 in the presence of chiral ligands. AB - The first example of the synthesis of planar chiral (1,3-disubstituted arene)Mn(CO)3+ cations (3) has been demonstrated by a reaction of (p cresol)Mn(CO)3+ with KOBut followed by addition of nucleophiles and subsequent quenching with electrophiles in the presence of (S)-binaphthol in CH2Cl2. PMID- 12109098 TI - Nanotectonic approach of the texturation of CeO2 based nanomaterials. AB - Original and homogeneous macrotextures shaped with coral-like, helical or macroporous sieves morphologies are obtained following a nanotectonic approach based on the template-directed assembly by poly-gamma-benzyl-L-glutamate (PBLG) of organically functionalised CeO2 crystalline nanoparticles. PMID- 12109099 TI - Highly selective amorphous Ni-Cr-B catalyst in 2-ethylanthraquinone hydrogenation to 2-ethylanthrahydroquinone. AB - A nanosized amorphous Ni-Cr-B catalyst prepared by the chemical reduction method exhibited superior thermal stability and selectivity in hydrogen peroxide synthesis via the anthraquinone route. PMID- 12109100 TI - Direct synthesis and catalytic evaluation of AlSBA-1. AB - The direct synthesis of the new mesoporous molecular sieve AlSBA-1 containing exclusively tetrahedrally coordinated aluminium and the catalytic activity of the novel material in the isomerization of n-decane are reported. PMID- 12109101 TI - Photocurrent responses at dye sensitised ultrathin polyelectrolyte multilayers supported on gold electrodes. AB - The photoelectrochemical behaviour of ionic conducting ultrathin multilayers formed by sequential deposition of poly-L-lysine and poly-L-glutamic acid on modified gold electrodes is investigated upon sensitisation by zinc mesotetrakis(p-sulfonatophenyl)porphyrin. PMID- 12109102 TI - Diels-Alder and ene reactions of singlet oxygen, nitroso compounds and triazolinediones: transition states and mechanisms from contemporary theory. AB - Singlet oxygen, nitroso compounds and triazolinediones have similar electronic structures: they share a low lying LUMO, making them powerful electrophiles, and a high lying HOMO, orthogonal to the LUMO and consisting of an antibonding combination of lone paris. This bestows some nucleophilic character on these species. We describe a number of studies employing the best levels of theory currently available for systems of this size and demonstrate that the Diels-Alder and ene reactions of these three species are calculated to show subtle changes in mechanism. The calculations have been calibrated, wherever possible, by comparison to experimental observations including measured activation and reaction energies, regio- and stereo-selectivities, intermediates observed either spectroscopically or by trapping, and kinetic isotope effects. PMID- 12109103 TI - Complexed bridging ligand, Cu(bptap)2, as a ferromagnetic coupler. AB - A novel complexed bridging ligand [Cu(bptap)2] which acts as a ferromagnetic coupler forms one dimensional chain consisting of tri-copper(II) [Cu2(Cu(bptap)2)]4+ units in which adjoining copper(II) ions are ferromagnetically coupled. PMID- 12109104 TI - Insertion of a strongly pi-pi stacked chloranilate pair into an M4 arrangement preorganized within a large macrocyclic ligand (M = Zn2+ and Cu2+). AB - Four Zn(II) ions arranged within a pyridine-modified large phenolate-containing macrocyle Lpy2- encapsulate two chloranilate ions in a double bis-didentate bridging fashion; the ligands are strongly pi-pi stacked with each other with a short distance of 3.27 A and are electrochemically reduced at the same potential of -1.00 V to produce a reasonably stable biradical species with T1/2 = 60 min at 25 degrees C. PMID- 12109105 TI - Molecular control of recombination dynamics in dye sensitised nanocrystalline TiO2 films. AB - Modification of the structure of a porphyrin dye shows a significant change in the rate of charge recombination between injected electrons in the TiO2 and the oxidized dye anchored to it following optical excitation, offering an insight into fundamental understanding of processes occurring at the dye/semiconductor interface. PMID- 12109106 TI - Orientation dependent electrocatalysis using self-assembled molecular films. AB - Surface orientation of self-assembled molecular films of 2,9,6,23-tetraamino cobalt phthalocyanine on gold and silver is shown to determine the nature and the products of the electrocatalytic reduction of oxygen. PMID- 12109107 TI - Formation and manipulation of supramolecular structures of oligo(p phenylenevinylene) terminated poly(propylene imine) dendrimers. AB - A third generation poly(propylene imine) dendrimer modified with pi-conjugated oligo(p-phenylenevinylene)s forms spherical and rod-like aggregates that can be manipulated by optical tweezers. PMID- 12109108 TI - Changes in motion vs. bonding in positively vs. negatively cooperative interactions. AB - From a consideration of the interactions between non-covalent bonds, it is concluded that positively cooperative binding will occur with a benefit in enthalpy and a cost in entropy, and that negatively cooperative binding will occur with a cost in enthalpy and a benefit in entropy; experimental data support these conclusions. PMID- 12109109 TI - Synthesis and reversible thermo-induced conformational transitions of a stable nitroxide biradical based on calix[4]arene. AB - A stable paramagnetic calix[4]arene(III) bearing two nitroxide groups on the upper rim is synthesized, which exhibits a strong intramolecular spin-spin exchange interaction and is found to be able to undergo reversible conformational transitions upon heating. PMID- 12109111 TI - Discontinuous pressure effect upon enantiodifferentiating photosensitized isomerization of cyclooctene. AB - A hydrostatic pressure of up to 750 MPa induced discontinuous changes in the enantiomeric excess of the (E)-isomer obtained in the enantio-differentiating photoisomerization of (Z)-cyclooctene and (Z,Z)-cycloocta-1,5-diene, sensitized by chiral benzene-1,2,4,5-tetracarboxylates; indicating a switching of the enantio-differentiation mechanism, which is attributable to dramatic conformational changes of chiral alkoxycarbonyl auxiliaries at a specific pressure. PMID- 12109110 TI - Synthesis of amino acid derivatives via enantio- and diastereoselective Pd catalyzed allylic substitutions with a non-stabilized enolate as nucleophile. AB - Diastereomer ratios of up to 95:5 and enantiomeric excesses of up to 95% were achieved in Pd-catalyzed asymmetric alyllic substitutions with zinc enolates of glycine esters as nucleophiles; a remarkable effect of the ligand on the diastereoselectivity of the substitution was found. PMID- 12109112 TI - 5,5-Fused thiophene gamma-lactams as templates for serine protease inhibition. AB - Novel 5,5-fused thiophene lactams are potent inhibitors and acylating agents of HNE and PPE. PMID- 12109113 TI - Synthesis of a deca-lithium cage containing an [(RN)2As(mu-NR)As(NR)2]4- tetraanion; a homologue of group 15 trianions of the type [E(NR)3]3-. AB - The novel, deca-lithium cage [(mtaNHLi)(As2(Nmta)5)-Li(4).2thf]2 (1) (mtaN = 5 methylthiazolyl, C4H4N2S) contains an imido-bridged tetraanion [(mtaN)2As(mu Nmta)-As(Nmta)2]4-, which represents a new type of multi-functional imido group 15 ligand framework (homologous with group 15 anions of the type [As(NR)3]3-). PMID- 12109114 TI - Dehydrodimerization of iodobenzenes to iodinated biaryls. AB - The molybdenum pentachloride-mediated oxidative coupling of iodo-substituted electron rich benzenes without the loss of the iodo-substituents is reported. The presented methodology is an environmentally friendly alternative to known thallium(III)- and lead(IV)-reagents. Even 2,2',6,6'-tetraiodobiphenyl derivatives are easily accessible. PMID- 12109116 TI - Pseudo-polyrotaxanes based on a protonated version of the 1,2-bis(4,4' bipyridinium)ethane-24-crown-8 ether motif. AB - Protonated 1,2-bis(4,4'-bipyridinium)ethane axles and dibenzo-24-crown-8 ether wheels thread to form [2]pseudorotaxanes which associate in the solid state to form pseudopolyrotaxanes by hydrogen bonding or pi-stacking. PMID- 12109117 TI - Rhodium-catalyzed approach to Mannich-type products using aldimine, alpha,beta unsaturated ester, and hydrosilane. AB - A rhodium-catalyzed method for the synthesis of beta-amino esters was accomplished in a one-pot procedure from aldimine, alpha,beta-unsaturated ester and hydrosilane. PMID- 12109118 TI - Three 2-oxazolinyl rings on one quaternary carbon atom: preparation of a novel tripodal tris(oxazolinyl) ligand and the tetrameric molecular structure of its CuI complex. AB - Reaction of the lithium salt of 1,1-bis[2-(4,4-dimethyl)oxazolinyl]ethane (1) with 2-bromo-4,4-dimethyloxazoline yielded the novel tripodal ligand, 1,1,1 tris[2-(4,4-dimethyl)-oxazolinyl]ethane (trisox-Me2 2) which was reacted with one mol equivalent of [Cu(NCCH3)4](BF4) in dichloromethane to give [Cu(trisox Me2)]n(BF4)n (3); while monomeric in solution, an X-ray diffraction study of 3 established a cyclic, centrosymmetric tetramer in the solid state. PMID- 12109115 TI - Total synthesis of (-)-stevastelin B. AB - The total synthesis and an unambiguous structure confirmation of stevastelin B 1, a novel 15-membered cyclic depsipeptide, are described; the fatty acid moiety in 1, prepared stereoselectively from L-quebrachitol was converted into the amino carboxylic acid, whose macrolactamization by Shioiri's procedure effectively constructed the cyclic structure of 1. PMID- 12109119 TI - High conversion of olefins to cis-diols by non-heme iron catalysts and H2O2. AB - Efficient and highly stereoselective oxidation of olefins to cis-diols as the major product is obtained by using biomimetic non-heme FeII catalysts in combination with H2O2. PMID- 12109120 TI - A novel precursor for synthesis of pure boron nitride nanotubes. AB - A novel precursor, a mixture of B2O2 and Mg which is generated in situ by reacting B and MgO at 1300 degrees C, can be used to effectively synthesize bulk amounts of pure BN nanotubes with Mg evaporated from the final product; transmission electron microscope observation for the synthesized BN nanotubes indicates that defects present strongly depend on the tube diameter. PMID- 12109121 TI - Operando Raman study of alumina-supported Sb-V-O catalyst during propane ammoxidation to acrylonitrile with on-line activity measurement. AB - Operando Raman spectra during propane ammoxidation show partially reversible structural transformations of the active phases as a function of reaction environment. PMID- 12109122 TI - Addition of malonyl radicals to glycals with C-1 acceptor groups: remarkable influence of the substituents on the product distribution. AB - The ceric(IV) ammonium nitrate (CAN)-mediated radical addition of dimethyl malonate to glycals 1 affords methyl glycosides 2 and ortho esters 3 as main products; the product distribution strongly depends on the substitution pattern at the 1-position, which can be rationalized in terms of the oxidation potentials of the intermediary anomeric radicals. PMID- 12109123 TI - Solid supported fluoronitroaryl triazenes as immobilized and convertible Sanger reagents--synthesis and SNAr reactions towards a novel preparation of 1-alkyl-5 nitro-1H-benzotriazoles. AB - Synthesis of novel fluoronitroaryl triazenes in liquid phase and on solid support have been described; mild displacement of the fluoride ion with various nucleophiles provides access to substituted arenes which in turn can be cleaved to provide a unique access to 1-alkyl-5-nitro-1H-benzotriazole. PMID- 12109124 TI - Living ethylene/norbornene copolymerisation catalyzed by titanium complexes having two pyrrolide-imine chelate ligands. AB - Ethylene/norbornene copolymerisation behaviour of titanium complexes with two pyrrolide-imine chelate ligands is described. PMID- 12109125 TI - Synthesis of silver dendritic nanostructures protected by tetrathiafulvalene. AB - Silver dendritic nanostructures protected by tetrathiafulvalene (TTF) were synthesized via reduction of silver ions with TTF in acetonitrile. PMID- 12109126 TI - Sex pheromone biosynthesis in the female olive fruit-fly. Double labelling from [18O2]-dioxygen into 1,7-dioxaspiro[5.5]undecane. AB - The demonstration that both oxygen atoms of 1,7-dioxaspiro[5.5]undecane (1), the sex-pheromone of the female olive fly, originate from dioxygen, strongly implicates monooxygenase mediated processes in assembly of (1), and reveals unexpected complexity in the formation of its nine-carbon precursor. PMID- 12109128 TI - Crystalline CrV0.95P0.05O4 catalyst for vapor-phase oxidation of picolines. AB - CrV0.95P0.05O4 prepared as a pure crystalline form was found to be highly active for the vapor-phase oxidation of picolines to the corresponding aldehydes and acids in the presence of water. PMID- 12109127 TI - Interplay of phenyl-perfluorophenyl stacking, C-H...F, C-F...pi and F...F interactions in some crystalline aromatic azines. AB - Analysis of phenyl-perfluorophenyl stacking synthon, C-H...F, C-F...pi interactions, and F...F tetramer in three closely related azine crystal structures shows the dominance of Ar-ArF synthon while other interactions are turned on/off depending on the H/F stoichiometry in the molecule. PMID- 12109129 TI - A one-pot access to cyclopropanes from allylic ethers via hydrozirconation deoxygenative ring formation. AB - A synthetic method for the direct transformation of allylic ether into mono-, di- and trisubstituted cyclopropanes is presented. PMID- 12109131 TI - High-pressure CO2-induced reduction of the melting temperature of ionic liquids. AB - An in situ ATR-IR spectroscopic study has shown that high-pressure CO2 reduces melting temperature of ionic liquids such as [C16mim][PF6]. PMID- 12109130 TI - Catalytic one-pot synthesis of N-phenyl alkyl amides from alkene and aniline in the presence of cobalt on charcoal under carbon monoxide. AB - N-Phenyl alkyl amides were synthesized by the cobalt on charcoal-catalyzed one pot reaction of alkene and aniline under carbon monoxide; this is the first heterogeneous catalytic formation of N-phenyl alkyl amides. PMID- 12109132 TI - Assembly of supermolecular complexes from the tripodal ligand titmb: assembly of a large M6L8 cage from 14 components. AB - The coordinatively saturated, nanometer-sized M6L8 complex [Pd6(titmb)8]Cl(12).2H2O (titmb = 1,3,5-bis(imidazol-1-ylmethyl)-2,4,6 trimethylbenzene) was obtained by assembly of six Pd(II) ions with eight flexible titmb tripodal ligands; structural analysis shows that these eight titmb are in a disordered cube configuration and the six Pd atoms are in a disordered octahedral configuration; the inner cavity of the cage is estimated to have a volume of 1000 A3, large enough to encapsulate eight Cl- anions. PMID- 12109133 TI - Progress toward vector design for hematopoietic stem cell gene therapy. AB - Hematopoietic stem cells (HSCs) are attractive targets for gene therapy because of their capacity for self renewal and the wide systemic distribution of their progeny. Sustained expression of transgenes at clinically relevant levels in the progeny of HSCs would provide novel and potentially curative treatments for a wide range of inherited and acquired blood diseases. Recent improvements in retroviral transduction protocols have resulted in the first successful amelioration of a human hematologic disease--a form of severe combined immunodeficiency--by HSC gene transfer. However, continued advances in gene transfer technology are necessary if the inherent promise of HSC gene therapy is to be fully realized. Ongoing efforts are focused on modifying oncoretroviral vector designs and pseudotyping with alternative envelope proteins. In addition, because of their ability to transduce non-divided cells, safety-modified human immunodeficiency virus-1-based lentiviral vectors have emerged as promising tools for gene modification of HSCs, which reside primarily in the G0/G1 phase of the cell cycle. Irrespective of these advances, accumulated data indicate that stably integrated transgenes are frequently subject to position-effect variegation and extinction of expression. Therefore, the extent to which genetic control elements such as chromatin domain insulators and scaffold/matrix attachment regions in conjunction with posttranscriptional regulatory elements will result in enhanced probability and level of transgene expression is under active investigation. Collectively, these developments increase the likelihood that HSC gene transfer will ultimately become an effective therapeutic strategy. PMID- 12109134 TI - Cancer gene therapy by adenovirus-mediated gene transfer. AB - Cancer arises as a direct result of genetic mutations. It therefore stands to reason that cancer should be well suited for the correction through gene therapy. Recent advances in the understanding of the molecular pathogenesis of cancer and the rapid development of recombinant DNA technology have made cancer gene therapy feasible in the clinical setting. The current efforts for cancer gene therapy mainly focus on immunogene therapy, chemogene therapy, restoration of tumor suppressor gene function, and oncolytic virus therapy. Central to all these therapies is the development of efficient vectors for gene delivery--this remains a work in progress. These vectors can be classified as viral and non-viral vectors. This paper will concentrate on viral vectors because of their practical advantages over non-viral vectors. Of the viral vectors, by far the most important are the human adenoviruses as is reflected by the enormous data and literature accumulated by studies relating to animal tumor models and clinical trials. In this review, we examine the recent progress in adenovirus-mediated cancer gene therapy with regard to cytokine gene, tumor suppressor gene, chemogene, and oncolytic adenovirus. We also discuss the current limitations of the adenoviral vector system and how they may be circumvented in future developments relating to targeted gene delivery. PMID- 12109135 TI - The use of Fas Ligand, TRAIL and Bax in gene therapy of prostate cancer. AB - Prostate cancer is the second leading cause of cancer death in the United States. Treatment options for confined disease are generally successful in prolonging life but long-term cures (10-15 years) are elusive for the majority of patients. The prognosis for advanced extra-capsular prostate cancer is grim. However, we are now entering the era of gene therapy options for treatment of prostate cancer. The human genome project coupled with genomics and protemics are providing information that will lead to selection of genes for treatment of prostate cancer. The problem is the science of delivery lags behind knowledge of gene function. Thus, it is important to develop therapies that do not require delivery to 100% of tumor cells but which nevertheless kills the entire cancer by virtue of the bystander effect or other means. This review covers the use, in gene therapy, of apoptotic inducing molecules such as Fas Ligand, and TRAIL which are believed to induce bystander killing activity and Bax which also may function in a similar way. PMID- 12109136 TI - Alphavirus vectors for gene therapy applications. AB - High-titer alphavirus vectors have been generated for efficient gene delivery both in vitro and in vivo. Studies on CNS infection via intranasal and peripheral injections with virulent and avirulent replication-competent Semliki Forest virus (SFV) strains has demonstrated the potential of gene delivery. Replication deficient alphavirus particles have shown high local transgene expression of a transient nature in rodent brain. Alphavirus vectors have been demonstrated to induce apoptosis in infected human tumor cell lines and SFV vectors expressing interleukin-12 resulted in tumor regression in a B16 murine melanoma model. Repeated SFV injections led to stronger anti-tumor effects without immunogenic response detected against SFV. It has also been shown that intra-tumoral SFV injections into nude mice with implanted human lung carcinomas led to tumor regression. Likewise, injection of replicative SFV-LacZ RNA resulted in tumor response as well as prophylactic protection against tumor formation. Alphaviruses have also showed potential in vaccine production. Additionally, modifications in the envelope structure of Sindbis virus resulted in substantial change in host range and demonstrated the feasibility of targeting alphavirus vectors. Moreover, SFV has been used as an expression vector for the generation of high-titer retrovirus-like particles. Recent alphavirus vector development has introduced novel non-cytopathogenic vectors, tightly temperature-regulated vectors as well as replication-persistent forms that should prolong the duration of expression. Alphavirus vectors can therefore be considered as highly potential gene delivery vehicles for future gene therapy applications, especially where only short-term expression is required, or even preferred. PMID- 12109137 TI - Biomaterials for gene delivery: atelocollagen-mediated controlled release of molecular medicines. AB - Over the last decade, increasing attention has been paid to the development of systems to deliver drugs for long periods at controlled rates. Some of these systems can deliver drugs continuously for over one year. However, little effort has been given to developing systems for the controlled release of nucleic acids. Recently, a novel gene transfer method which allows prolonged release and expression of plasmid DNA in vivo in normal adult animals was established. In this system, a biocompatible natural polymer such as collagen or its derivatives acts as the carrier for the delivery of DNA vectors. The biomaterial carrying the plasmid DNA was administered into animals and, once introduced, gradually released plasmid DNA in vivo. A single injection of plasmid DNA/biomaterial produced physiologically significant levels of gene-encoding proteins in the local/systemic circulation of animals and resulted in prolonged biological effects. These results suggest that the biomaterials carrying plasmid DNA may enhance the clinical potency of plasmid-based gene transfer, facilitating a more effective and long-term use of naked plasmid vectors for gene therapy. Furthermore, the biomaterials can be removed surgically, minimizing the effect of gene products if some unexpected side effects should be observed after application. The application of these systems to expand the bioavailability of molecular medicine, including antisense oligonucleotides and adenovirus vectors, and to aid in stem cell transplantation in the context of DNA-based tissue engineering will be discussed. PMID- 12109139 TI - The molecular genetics of lentiviral vectors--current and future perspectives. AB - Gene therapy is a promising endeavor for the treatment of disease in the 21st century. The key to capitalize on this venture lies in the availability of efficient gene transfer and expression tools. Viral vectors are useful vehicles for the delivery of foreign genes into target cells, and retroviral vectors have been popular because of their ability to integrate into the host cell genome and maintain persistent gene expression. Recent studies of the human immunodeficiency virus (HIV) have demonstrated that lentiviruses, members of the retroviral family, have the ability to infect cells at both mitotic and post-mitotic stages of the cell cycle. This article aims to analyze the molecular genetics, review existing systems and applications, and address problems as well as potential future developments of the lentiviral vector systems. PMID- 12109138 TI - Genetic vaccination for the active immunotherapy of cancer. AB - Molecular biology techniques have given novel impetus to the immunotherapy of cancer because they have catalyzed the identification of several potential tumor antigens, and permitted the generation of vectors for the delivery of genetic material encoding these antigens. Vaccines can be defined "genetic" when the antigen they enclose is present as DNA or RNA. Microrganisms used as vectors can deliver the genetic information, but naked nucleic acids have also been shown to be effective immunogens thanks to built-in adjuvants that activate professional antigen presenting cells. Although gene-based cancer vaccines have been tested in mouse models and selected for pilot clinical trials, enthusiasm has somewhat waned due to an apparently major drawback of cancer vaccination: tumor antigens are weak, and therefore fail to stimulate a sterilizing immune response in tumor bearing patients. Mouse studies, however, have shown that cancer vaccines are extremely efficacious in establishing a state of active immunosurvellance against tumor growth. This review reconsiders the findings emerging from preclinical studies in the context of our current knowledge of the cellular and molecular bases of the immune responses to vaccines, in an attempt to approach critically the use of genetic vaccination for the treatment of cancer. PMID- 12109140 TI - Life or death of T cells with antigen-specific receptors--using T cells for cancer adoptive immunotherapy/gene therapy. AB - A promising strategy for cancer treatment is adoptive gene therapy/immunotherapy by genetically modifying T lymphocytes with a chimeric receptor (ch-TCR) so that cytotoxic T lymphocytes (CTL) can target and lyse tumors in a MHC-non-restricted manner. It is, however, not clear whether non-MHC-restricted tumor cell recognition by T cells will result in activation-induced apoptosis (AICD). This review discusses the factors that affect the development of AICD or CTL proliferation, and how such factors should be considered in the design of clinical trials using ch-TCR. PMID- 12109141 TI - The use of adenoviral vectors for genetic manipulation and analysis of primitive hematopoietic cells. AB - Gene transfer into stem cells has long been studied as a means by which primitive hematopoietic cells could be characterized and manipulated. While a variety of strategies have been attempted, it still remains relatively difficult to perform direct stem cell analysis. In this review, we examine recent studies using adenovirus-based vectors as a means to achieve high-level gene transfer into primitive hematopoietic cell types. PMID- 12109142 TI - Possibilities of non-viral gene transfer to improve cutaneous wound healing. AB - Enhancement of dermal and epidermal regeneration represents a crucial goal for the treatment of acute, e.g. burn and trauma wounds, and chronic wounds, e.g. diabetic, autoimmune, arterial and venous wounds. Studies defining molecular mechanisms of the complex cascade of wound healing have shown that growth factors represent a new therapeutic strategy. The clinical application of growth factors in the form of proteins has been shown to be of little benefit. Therefore new delivery systems and therapeutic strategies needed to be developed to improve dermal and epidermal regeneration, one of which is gene therapy. For successful gene delivery the selection of an appropriate vector has been shown to be paramount. Because Retroviruses, Adenoviruses and Adeno-Associated Viruses can cause immunologic reactions and mutations, non-viral delivery systems for gene therapy, such as liposomal gene transfer appear advantageous over viral gene therapy. This review discusses the success, potential and limitations of non viral gene transfer to improve regeneration of dermal and epidermal structures. PMID- 12109143 TI - Human neural stem and progenitor cells: in vitro and in vivo properties, and potential for gene therapy and cell replacement in the CNS. AB - The generation of unlimited quantities of neural stem and/or progenitor cells derived from the human brain holds great interest for basic and applied neuroscience. In this article we critically review the origins and recent developments of procedures developed for the expansion, perpetuation, identification, and isolation of human neural precursors, as well as their attributes. Factors influencing their in vitro properties, both under division and after differentiation conditions, are evaluated, with the aim of identifying properties common to the different culture systems reported. This analysis suggests that different culture procedures result in cells with different properties, or even in different cells being isolated. With respect to in vivo performance, present evidence obtained in rodents indicate that cultured human neural precursors, in general, are endowed with excellent integrative properties. Differentiation of the implanted cells, in particular in the case of adult recipients, seems not to be complete, and functionality still needs to be demonstrated. In relation to gene transfer and therapy, aspects currently underexplored, initial data support the view that human neural stem and progenitor cells may serve a role as a platform cell for the delivery of bioactive substances to the diseased CNS. Although a large deal of basic research remains to be done, available data illustrate the enormous potential that human neural precursors isolated, expanded, and characterized in vitro hold for therapeutic applications. In spite of this potential, maintaining a critical view on many unresolved questions will surely help to drive this research field to a good end, that is, the development of real therapies for diseases of the human nervous system. PMID- 12109144 TI - Viral vector-mediated gene therapy for hemophilia. AB - Hemophilia A and B are hereditary coagulation disorders that result from functional deficiencies of factor VIII (FVIII) or factor IX (FIX), respectively. Current treatment consists of injections with plasma-derived or recombinant clotting factors. Despite the significant clinical benefits of protein replacement therapies, these do not constitute a cure and patients are still at risk of bleeding. Significant progress has been made recently in the development of gene therapy for hemophilia. This has been primarily due to the technical improvements of existing vector systems and the development of new gene delivery methods. Therapeutic and sometimes physiologic levels of FVIII and FIX could be achieved in FVIII- and FIX-deficient mice and hemophilic dogs using different types of viral vectors. In these preclinical studies, long-term correction of the bleeding disorders and in some cases a permanent cure has been realized. However, complications related to the induction of neutralizing antibodies or viral promoter inactivation often precludes stable phenotypic correction. Several gene therapy phase I clinical trials have been initiated in patients suffering from severe hemophilia A or B. The results from the extensive pre-clinical studies and the preliminary clinical data are encouraging. It is likely that successful gene therapy for hemophilia will become a reality at the beginning of this new millennium, serving as the trailblazer for gene therapy of other diseases. PMID- 12109145 TI - Current systematics of Penicillium and Aspergillus and the implication to fungal biodiversity and applied research. PMID- 12109146 TI - High-level expression of industrial enzymes originated from plants in fungal hosts. PMID- 12109147 TI - The smut fungi of the world. A survey. AB - After a short historical review, our current knowledge about the smut fungi of the world, their number, classification, occurrence on host plant groups and their relationship to other groups of fungi are presented. Surprising results of the new classification of smut fungi are shown. The estimated number of existing species, expected results and trends in smut fungus taxonomy, the necessity and modalities of the conservation of smut fungi and the preparation of an illustrated world monograph of smut fungi are discussed. PMID- 12109148 TI - Mycological characterization and comparison of climax forest associations in the Mecsek Mountains. AB - Deciduous forests, developed in Submediterrean climate, differ essentially from those in Central and Western Europe. Present paper concerns fungistical characterization of four Submediterranean climax associations based on 8 years investigation, as well as on literary data. Near common and rare species differential and local character ones are considered, those with indicative value, too. In addition, functional distribution of these associations is discussed. PMID- 12109149 TI - Lignicolous macrofungi of the Kekes North forest reserve in the Matra Mountains, Hungary. AB - A three-year mycological investigation of the Kekes North forest reserve in the Matra mountains. Hungary proved the richness of the area in lignicolous macrofungi. Diversity of macrofungi was in close correlation with the developmental phases of the forest as well as with the amount of dead wood of different quality (diameter, stage of decomposition, etc.). PMID- 12109150 TI - Studies on the root associations of the truffle Terfezia terfezioides. AB - The paper contains an overview of the results of the studies made on the truffle Terfezia terfezioides, particularly the investigations related to the associations of this fungus with plants. Twelve plant species originated from a natural habitat of the fungus were supposed to be connected with T. terfezioides based on the anatomy of the endogenous fungal structures in their roots. Aseptic experiments were carried out on modified MMN substrates with different phosphate concentrations to study the interaction of T. terfezioides with Robinia pseudoacacia and Helianthemum ovatum. The colonization of the roots of black locust was always weaker than that of Helianthemum. The main characteristics were the intracellular coiled, branched, frequently septated hyphae in dead root cells. The intercellular hyphae formed Hartig-net with finger like structures only in Helianthemum, the interactions could not be considered unambiguously as mycorrhizae. There was no difference between the RFLP profiles of the nr DNA ITS of nineteen fruit bodies collected at the same time from the habitat and the ITS of three randomly chosen specimens were identical on sequence level, too. These invariability makes to design species specific PCR primers possible to check unambiguously the host plants. PMID- 12109151 TI - The role of mycorrhizae in afforestation (a review). AB - Hungary is facing to perform intensive afforestation. New forests will be planted on dry, poorly fertile soils nonprofitable for agricultural use. Applying artificially mycorrhized seedlings may considerably increase the effectivity of afforestation and decrease costs. PMID- 12109152 TI - The aerobiology of the ascospores. AB - Atmospheric ascospores have been monitored using volumetric spore trap. Spore concentration data were analysed using Spearman's correlation. Our results show that the meteorological factor with the greatest effect on spore concentration was the duration of rain. Temperature increase strongly reduced the ascospore concentration; but the length of windless periods resulted in an increase in spore count. The only measurable effect wind perse actually had on spore count, was registered when a strong wind blew after a long windless period. We observed that the count of ascospores during wet weather could surpass the total concentration of dry conidia measured on a typical, highly polluted summer day. Using selected air samples to study the effect of storms, certain aspects of long distance spore transport were elucidated. We describe here three main strategies for long-range ascospore transport, "splash-off", "secondary emission" and "sporematrix projectiles". PMID- 12109153 TI - Is the widely used medicinal fungus the Ganoderma lucidum (Fr.) Karst. sensu stricto? (A short review). AB - The identification of Ganoderma species is usually based on classical morphological criteria. The objectives of this review were to collect available information on Ganoderma lucidum and to utilize them in exact identification of Ganoderma lucidum (Fr.) Karst. sensu stricto. A lot of taxonomical confusion has always been associated with G. lucidum and allied species. Species circumscription, phylogenetic relationships, host range and distribution of species of the G. lucidum complex are unclear even among the few taxa living in temperate climate. Several methods have been proposed to identify the species as examination of cultural characteristics, isozymes, secondary metabolites, DNA sequences and interfertility. Although G. lucidum sensu stricto has been reported world-wide accumulated evidence supported the suggestion that it seems restricted to Europe. The strains used in the medicine are usually collected in Asia. There is little likelihood that any one belongs to the G. lucidum sensu stricto. The strains labelled as G. lucidum in the medicinal and pharmacological literature encompass a broad range of species which produce different medicinally active compounds and have significantly different pharmacological effects. PMID- 12109154 TI - Advances in the identification of emerging powdery mildew fungi using morphological and molecular data. AB - Recently, the classical morphological criteria and host range data used in the identification of powdery mildew fungi were supplemented with scanning electron microscope (SEM) and molecular phylogenetic analyses. This paper discusses the joint use of these methods in the identification of powdery mildew anamorphs causing new or emerging plant diseases in different parts of the world. PMID- 12109155 TI - Canker and wilt of black locust (Robinia pseudoacacia L.) caused by Fusarium species. AB - From the pathological material of black locust trees showing symptoms of wilting of the foliage or canker of the bark the following Fusarium species were isolated: Fusarium avenaceum (Fr.) Sacc., Fusarium lateritium Nees., Fusarium semitectum Berk. & Rav., Fusarium solani (Mart.) Sacc., Fusarium sulphureum Schlecht. (syn.: Fusarium sambucinum Fuckel f. 6 Wollenw.) The results of the provocation infections of one-year-old black locust seedlings showed that all of the species--except Fusarium solani--are able to cause considerable necrosis in living bark and phloem. Fusarium sulphureum had by far the highest pathogenecity among the tested species. Fusarium semitectum isolated from withered black locust tree also caused necrosis on significant bark area. In the course of the penetration assay Fusarium sulphureum and Fusarium avenaceum were the most successful, and these species can cause cankers on the stem and twigs of black locust without frost effect. PMID- 12109156 TI - Carbon catabolite repression in the regulation of beta-galactosidase activity in Aspergillus nidulans. PMID- 12109157 TI - Investigation of glutathione metabolism in filamentous fungi (a short auto review). PMID- 12109158 TI - Molecular mechanisms of action of chromium compounds in yeast (a short communication). PMID- 12109159 TI - Genetics of sulphate assimilation in Schizosaccharomyces pombe (a short review). AB - Sulphur plays an important role in yeasts, especially in the biosynthesis of methionine and cysteine. The inorganic sulphur source, sulphate, is taken up by the cells via the sulphate-permease(s). After its transport, it is activated and subsequently reduced to sulphide or serves as a donor for sulphurylation reactions. Selenate anion (SeO4(2-)), which has the same metabolic pathway as sulphate, is toxic for the cells of Schizosaccharomyces pombe. We isolated selenate resistant mutants which cannot utilize sulphate, therefore they need organic sulphur source for growth. One of the selenate resistant mutants was successively transformed with S. pombe genomic libraries and the gene complementing the selenate resistance was identified as that of coding for the ATP-sulphurylase enzyme. PMID- 12109160 TI - Isolation and characterization of fission yeast genes involved in transcription regulation of cell cycle events (a short communication). PMID- 12109161 TI - Regularities and irregularities in the cell cycle of the fission yeast, Schizosaccharomyces pombe (a review). AB - In an exponentially growing wild-type fission yeast culture a size control mechanism ensures that mitosis is executed only if the cells have reached a critical size. However, there is some scattering both in cell length at birth (BL) and in cycle time (CT). By computational simulations we show here that this scattering cannot be explained solely by asymmetric cell division, therefore we assume that nuclear division is a stochastic, asymmetric process as well. We introduce an appropriate stochastic variable into a mathematical model and prove that this assumption is suitable to describe the CT vs. BL graph in a wild-type fission yeast population. In a double mutant of fission yeast (namely wee1-50 cdc25 delta) this CT vs. BL plot is even more curious: cycle time splits into three different values resulting in three clusters in this coordinate system. We show here that it is possible to describe these quantized cycles by choosing the appropriate values of the key parameters of mitotic entry and exit and even more the clustered behavior may be simulated by applying a further stochastic parameter. PMID- 12109162 TI - Cloning and sequence analysis of Mucor circinelloides glyceraldehyde-3-phosphate dehydrogenase gene. AB - A genomic library of Mucor circinelloides ATCC 1216b has been constructed in Lambda Fix II vector. The library has an average insert site of 10 kb and covers the genome 12 times. The M. circinelloides gene encoding glyceraldehyde-3 phosphate dehydrogenase (gpd) was isolated from this library by hybridization of the recombinant phage clones with a gpd-specific gene probe generated by PCR reaction. The complete nucleotide sequence encodes a putative polypeptide chain of 339 amino acids interrupted by 3 introns. The predicted amino acid sequence of this gene shows a high degree of sequence similarity to the GPD proteins from other filamentous fungi. The promoter region, containing a consensus TATA and CAAT box and a 298 nucleotid long termination region were also determined. PMID- 12109163 TI - Structure and function of mating-type genes in Fusarium species. PMID- 12109164 TI - Diversity of extrachromosomal genetic elements in yeasts (a rewiev). PMID- 12109166 TI - Role of mobile introns in mitochondrial genome diversity of fungi (a mini review). PMID- 12109165 TI - Identification of Fusarium species by isozyme analysis. AB - Cellulose-acetate electrophoresis (CAE) was used to investigate isozyme polymorphisms among different isolates of Fusarium cerealis, F. culmorum, F. graminearum and F. pseudograminearum. After initial testing of 18 enzymes in three buffer systems for activity and resolution of bands, 12 proved to be appropriate for analysis of the full sample set. Comparing the different electrophoretic types (ETs), adenylate kinase (AK), NADP dependent glutamate dehydrogenase (NADP GDH), peptidase B (PEP B), peptidase D (PEP D) and phosphoglucomutase (PGM) proved to be diagnostic for at least one species examined. However, only PEP D was useful alone as a marker to distinguish the four taxa studied providing a rapid and simple CAE based diagnostic protocol. PMID- 12109167 TI - Identification and incidence of fungal strains in chronic rhinosinusitis patients. AB - The fungal revolution taking place in otorhinology inspired us to study the frequency of occurrence of fungi in the nasal mucus of chronic rhinosinusitis (CRS) patients (with or without polyposis) in order to evaluate the incidence of eosinophilic fungal sinusitis in CRS patients. Ninety-six samples were examined from patients with CRS. In 74 cases mucus was collected non-invasively, and in 22 cases during operation. The Gram-stained direct smears of all samples were also evaluated. Bacteria and fungi colonizing in the mucus were detected by culturing method. The control group consisted of 50 healthy volunteers. Typical aerobic pathogenic bacteria could be isolated from 34 patients. Fifty-seven aerobic bacteria were isolated, i.e. 1.6 bacteria/positive patient with a maximum of 3 different bacteria/sample. The most frequently isolated bacteria were Staphylococcus aureus, Escherichia coli, Klebsiella pneumoniae, Streptococcus pneumoniae, and Haemophilus influenzae. Yeasts and moulds could be detected from 79 patients (83%): Candida albicans, Candida spp., Aspergillus spp., Cladosporium spp, and Penicillium spp. were isolated most frequently. Altogether 237 yeasts and moulds were isolated, i.e. 3.0 different fungi/positive patient, with a maximum of 5 different fungi/sample. In the control group aerobic pathogens were not isolated, only apathogenic species. Fungi were isolated from 22 healthy patients (44%). These data indicate that fungi are frequently involved in the aetiology of CRS. IgE-medicated hypersensitivity to fungal allergens could not be proven in our patients. PMID- 12109168 TI - Onychomycosis and its impact on quality of life. PMID- 12109169 TI - Distribution and susceptibility of Candida species isolated in the Medical University of Debrecen. AB - Data of Candida albicans and non-albicans Candida species isolated during the 1997-2000 period in the Medical and Health Science Center of the University of Debrecen are analysed. The number of yeast isolates increased from 408 to 1213 per year during this period. Dominance of C. albicans has been persistent, but a slight increase of C. glabrata and C. krusei could be observed. Distribution of different Candida species isolated from 16 body sites indicates that C. albicans seems to be still the most aggressive Candida species. Investigation of 244 urinary Candida isolates (parallel with bacterial cultures) suggests that tha aetiological role of Candida species in the pathogenesis of urinary tract infections can be hypothesized if colony forming unit (CFU) number of yeasts is higher than 10(4)/ml and bacteria are present in low CFU number or are absent. Antifungal susceptibility testing of C. albicans, C. glabrata, C. tropicalis and C. krusei against Flucytosine, Amphotericin-B, Miconazole, Ketoconazole and Fluconazole suggests that Amphotericin-B is still the most effective antifungal agent. Finally, the problems in judging the aetiological role of isolated Candida species in the pathogenesis of different types of diseases are critically discussed. PMID- 12109170 TI - Pathological and clinical aspects of the diseases caused by Malassezia species. AB - From veterinary point of view Malassezia pachydermatis has the greatest significance. It has been standing in the focus of interest since the early 1990s, mostly because of the frequency of otitis externa and dermatitis caused by this yeast in dogs. This is the only lipid-independent species in the genus Malassezia. It can be found in very large proportion on the skin of healthy animals, but can be isolated in much greater number from diseased dogs. It often causes illness together with other pathogens (e.g. Staphylococcus intermedius). Some breeds are predisposed. In addition to the treatment of the accidental concurrent diseases, therapy consists of systemic and/or topical antimicrobial treatment. Ketoconazole is used most frequently. Malassezia pachydermatis plays also a role in the skin disorders of other carnivores. It has little zoonotic potential, it can be dangerous to immunocompromised humans. The other Malassezia species have little veterinary importance, although M. sympodialis and M. globosa were isolated from asymptomatic animals (mostly cats) and from mixed infections. PMID- 12109171 TI - Research priorities in school nursing: a Delphi process. PMID- 12109172 TI - Evaluation of the North Carolina "Baby Think It Over" project. AB - The North Carolina "Baby Think It Over" (BTIO) evaluation was conducted during spring semester 2000. Data were collected from participating teachers, students, and parents. Twenty-five teachers were selected randomly from different counties in North Carolina. Each randomly selected teacher coordinated the evaluation in conjunction with the intervention. Student surveys determined whether using the BTIO doll changed perceptions and attitudes toward teen parenting. Information also was collected on the veracity of each student's participation based on data provided from the computer in the baby. Surveys were sent home to the parents of participating students so information on communication, disruption of the household, and parenting perceptions could be obtained. Each teacher completed a survey that sought information on topics discussed throughout the intervention, hours of the program, and perceptions of program effectiveness. Parents and teachers were offered participation incentives. Overall, support existed for the BTIO intervention by parents and teachers. Most teachers and parents felt the program was effective at increasing communication about parenting and changing teens' attitudes in a desired direction. Most teachers reported that the intervention was not disruptive to their classes. However, results from student surveys did not reveal the same support. Student changes in attitudes and beliefs about parenting after the intervention were minimal. PMID- 12109173 TI - Urban elementary school students' perceptions of fighting behavior and concerns for personal safety. AB - This study assessed urban elementary school students' experience with weapon carrying and violence, concerns for personal safety, and perceptions of passive and direct interventions in resolving fights. The survey was completed by 1,912 urban students in the fourth and fifth grades. This cross-sectional study found that one in 12 students reported weapon carrying one or more times during the past month. One-third indicated that they would hit peers back if struck by them. One-quarter of students did not feel safe going to or from school, and 23%-43% worried about being physically attacked in or around school. Adjusted odds ratios and 95% confidence intervals were calculated using logistic regression. Significant associations were found between the independent variables of age, race, gender, and academic success (grades) and the dependent variables of weapon carrying, hitting a peer back, concerns of safety, and passive solutions or direct interventions for peer fighting. PMID- 12109174 TI - Association of language spoken at home with health and school issues among Asian American adolescents. AB - The study examined the association of language spoken at home with the school and health risks and behaviors of Asian American adolescents. Using the United States component of the 1997-1998 World Health Organization Study of Health Behavior in School Children, bivariate and multiple logistic regression analyses were conducted of records for Asian children to explore the relationship between language spoken at home and outcome variables regarding health behaviors, psychosocial and school risk factors, and parental factors. Compared to those who usually speak English at home, adolescents who usually speak another language, or who speak two languages equally, face a greater risk for health risk factors, psychosocial and school risk factors, and parental risk factors. Not speaking English at home was associated with higher health risks, including not wearing seat belts and bicycle helmets; higher psychosocial and school risk factors, including feeling that they do not belong at school, difficulty making new friends, and lacking confidence; and higher parental risks, including reporting that parents were not ready to help them or willing to talk to teachers. Adolescents less acculturated to the United States experience a variety of physical and psychosocial risks. School-based interventions such as early identification and outreach, needs assessment, and counseling and support services should be provided to immigrant students and their families. PMID- 12109175 TI - Evaluation of an upper elementary school program to prevent body image, eating, and weight concerns. AB - Preparing young students to resist the sociocultural pressures that contribute to body image and eating problems in American culture poses a monumental challenge. This project determined if the 11-lesson "Healthy Body Image: Teaching Kids to Eat and Love Their Bodies Too!" curriculum would have a positive effect when presented to upper elementary school children. A controlled study with 415 students measured changes from pretest to posttest related to: 1) body image; 2) knowledge about the biology of size, shape, and restricted hunger or dieting; 3) body size prejudice; 4) media awareness; 5) self-image; and 6) lifestyle behavior. Boys and girls who completed the curriculum showed significant or notable improvement compared to a control group not exposed to the curriculum. Results suggest it is possible to provide children with a knowledge base to use as they face increasing pressures about appearance, weight, and eating in the critical middle school years. PMID- 12109176 TI - Truancy, grade point average, and sexual activity: a meta-analysis of risk indicators for youth substance use. AB - Society increasingly holds schools responsible for the effectiveness of health promotion activities, such as drug abuse prevention efforts funded through the federal Safe and Drug-Free Schools program. Consequently, school districts use student surveys as a method for assessing trends and evaluating effects of programs on behavior. Because cost and practical concerns often preclude consistent population-based school survey sampling, risk indicators can provide an essential tool in analyzing needs assessment and program evaluation data. In this paper, three risk measures associated with substance use were selected from among commonly used school surveys. These measures--truancy, grade point average, and recent sexual intercourse--were compared, using meta-analysis techniques, to assess the reliability of risk measures across different survey instruments, different communities, and different points in time. Truancy was judged superior, because of its strong predictive value, particularly among younger students, and because rates can be compared to school records to assess sampling validity over time. PMID- 12109177 TI - Using the Kindergarten Health Assessment Report as a health report card. PMID- 12109178 TI - [Videothoracosopy in diagnosis and surgical treatment of tuberculosis]. AB - From 1993 to May 2001, 795 psychiatric patients were treated including 563 with pleural effusion, 98 with multiple or solitary tuberculoma, 69 with tuberculous empyema, 14 with fibrotic-cavitary tuberculosis, 51 with disseminated tuberculosis. Mean age of the patients was 32 years. Survey of pleural cavity with pleural or lung biopsy was performed in 691 patients. In tuberculoma 14 lobectomies were performed, 23 patients underwent atypical lung resection without stapler (including with precise technique). Bilateral on-stage interventions were made in 32 patients. In empyema necrectomy and lung decortication were performed. 14 patients underwent videocavernoscopy with sequestrectomy and cavity drainage. In disseminated tuberculosis lung biopsy without staplers was usually performed. Aerohemostasis was achieved with plasma stream. One patient with empyema and one patient with pleural effusion died (lethality was 0.25%). Rate of postoperative complications was 7.5% in tuberculoma and 1.5% in disseminated tuberculosis. Conversion to thoracotomy was necessary in 3 (3%) patients with tuberculoma and 12 (17%) patients with empyema. Mean hospital stay was 4 days after diagnostic surgery and 7 days after lung resection. In pleural effusion diagnosis was verified in 98% cases, in disseminated tuberculosis--in 100%. Videothoracoscopy is the best diagnostic method for pleural effusion and disseminated forms of lung tuberculosis and operation of choice in tuberculoma and empyema. Videothoracoscopy in tuberculosis is highly effective and associates with low rate of postoperative complications and lethality. PMID- 12109179 TI - [Pancreatoduodenal resection and total duodenopancreatectomy in surgery for stomach cancer]. AB - Experience with combined operations for stomach cancer in combination with pancreatoduodenal resection or total duodenopancreatectomy is presented. Immediate and long-term results of 10 pancreatoduodenal resections and 5 total duodenopancreatectomies are analyzed. There were 2 lethal outcomes. Complicated postoperative period was seen in 7 patients. During the first year of follow-up 6 of 13 operated patients died due to dissemination. After total pancreatectomy severe homeostatic disorders were seen. Prognosis after pancreatoduodenal resection is better when the tumor involves the duodenum than when it invades the pancreas. Subtotal resection of the stomach and gastrectomy in combination with duodenopancreatosplenectomy lead to unfavorable functional results and have bad long-term prognosis. PMID- 12109181 TI - [Diagnosis of postoperative peritonitis]. AB - 10-year experience of diagnosis of postoperative peritonitis (POP) in 106 patients is presented. Clinical deterioration, dryness in the mouth and thirst, tachycardia, bloated abdomen, hyperthermia, leukocytosis, positive Blumberg Shchetkin symptom, intestinal paresis and rigidity of abdominal muscles were the main symptoms of POP which were revealed in 83-100% patients. Absence of drainage secretions and pneumoperitoneum don't exclude POP. Ultrasonic examination is a highly effective additional method in diagnosis of POP. When POP is suspected it makes sense to take into account opinion of surgeons who have not operated this patient. PMID- 12109180 TI - [Comparative evaluation of the motor-evacuation function of the stomach after functional operations for complicated duodenal ulcers]. AB - Motor-evacuatory disorders (MED) of the stomach in 114 patients who have undergone functional operations for duodenal ulcers were analyzed. Clinical picture, barium contrast and ultrasonic examinations, fibrogastroduodenoscopy, pH monitoring were used for evaluation. Many patients have MED of different degree in early postoperative period. Some patients have severe disorders that require drug therapy. US and endoscopy role in complex assessment and differential analysis of stomach evacuatory disorders is emphasized. Isolated elements of study methods are discussed. PMID- 12109182 TI - [Laparoscopic sanitation of the abdominal cavity in combined treatment of peritonitis]. AB - Standard methods of treatment were analyzed in 228 patients with general peritonitis of different etiology: sanitation and drainage of abdominal cavity, peritoneal lavage, programmed laparotomy compared with laparoscopic sanitation (LS) performed in 60 patients. A significant decrease in the number of purulent inflammatory complications, intestinal fistulas and broncho-pulmonary complications were seen after LS. It was most effective for treatment of peritonitis with Mannheim peritoneal index from 16 to 29 points (lethality 11.8%) compared with conventional treatment (p < 0.05). Indications and contraindications for LS, original device "BRUSAN" were developed. Abdominal LS is a highly effective and mini-invasive method of local treatment in general peritonitis. PMID- 12109184 TI - [Variants of surgical treatment of lateral postoperative abdominal hernia by a combined method]. AB - 616 patients with postoperative abdominal hernias (AH) were treated, 124 (20%) of them had lateral hernias. By M. Yatsentyuk classification, there were 41 (33.1%) patients with small hernias, 37 (29.8%)--with middle-size, 35 (28.2%)--with big, 6 (4.3%)--with very large, 5 (4%)--with giant hernias. Two operative techniques for lateral AH based on an original method of combined plastic surgery were applied in 37 patients. Good immediate and long-term results were achieved in all the patients. There were no recurrences. PMID- 12109183 TI - [Endovascular intrahepatic portacaval shunting]. AB - Pilot experience in Russia of transjugular intrahepatic portosystemic shunting is presented. Results of endovascular shunting in 14 patients with cirrhosis of the liver were analyzed. Four patients underwent emergency endovascular shunting when conservative treatment was not effective. Ten patients underwent elective surgery. Repeated esophagealgastric bleedings were indications for shunting. Relapse of bleeding was seen in 1 patient 5 days after surgery. Regular dopplerography permitted to reveal timely stenotic process. When stenosis of portosystemic shunt was diagnosed, ambulatory balloon dilatation of anastomosis through transjugular approach was performed. It was enough for opening of anastomosis lumen and decrease of pressure gradient. PMID- 12109185 TI - [Local esophagoplasty in patients with burn strictures of the esophagus]. AB - Local plastic reconstruction of short burn strictures of the esophagus (SBSE) was performed in 14 patients aged from 22 to 37 years, 9 were men, 5--women. Decreased body mass was revealed in all the patients, mass deficit over 20% was in 4 (28.6%) of them. SBSE located in the upper third of the esophagus in 9 (64.3%) patients, in middle third--in 5 (35.7%). Local esophagoplasty was performed from cervical approach in 9 (64.3%) patients, from right-sided thoracotomy--in 5 (35.7%). Dissection of SBSE with 3/4 anastomosis was performed in 5 (35.7%) patients; circulatory resection of esophageal stricture with end-to end anastomosis--in 7 (50.0%); detour side-to-side anastomosis without stricture resection--in 2 (14.3%). There was no hospital lethality. Postoperative complications were seen in 14.2% cases. Long-term results were good in 10 (71.4%) patients, satisfactory (strictures of anastomosis)--in 3 (21.4%), unsatisfactory- in 1 (7.2%). Local esophagoplasty is indicated for patients with SBSE when bouginage and dilatation of stenosis are not effective. PMID- 12109186 TI - [Ultrasonic method of monitoring the course of the wound process in the anterior abdominal wall]. AB - Method of ultrasonic monitoring of wound process was used in 102 patients with acute surgical abdominal diseases undergone surgery from middle laparotomy. On 7th and 10th day after operation in phase of regeneration patients with uncomplicated wound process demonstrated reduce of hypoechogenic zone with appearance of zones with high echogenic structure, or transformation of hypoechogenic zones in hyperechogenic. Increase of hypoechogenic zone width in. 5 7 days after operation testifies high risk of purulent-necrotic complications in wound. PMID- 12109187 TI - [Comparative characteristics of Liga Sure and Auto Sonix devices in laparoscopic surgery]. AB - Use of Liga Sure and Auto Sonix devices in laparoscopic surgery on the stomach, colon and rectum for gastric, duodenal ulcers and colorectal cancer was analyzed. These devices make easier surgery and improved postoperative period. In test group of patients time of surgery, blood loss, smoke in surgical field were less, scope of laparoscopic stage increased. PMID- 12109188 TI - [Scintigraphic evaluation of the efficacy of nonstandard methods of treating critical ischemia of the lower limbs]. AB - In 77 patients with critical limb ishaemia treated surgically (arterialization of venous blood flow, creation of arteriovenous fistulas, transplantation of great omentum on microvascular anastomosis) regional perfusion of both legs was evaluated with radionuclide angiography. High perfusion of the ishaemic limb was revealed in the majority of patients. In these patients positive results were achieved most often, particularly in creation of arteriovenous fistulas on the leg low third and foot. PMID- 12109189 TI - [Surgical treatment of older patients with critical ischemia of the lower limps in occlussive disease of aorta and iliac arteries]. AB - 17-year experience in surgical treatment of 302 elderly and old patients with occlusive disease of aorta and iliac arteries and critical limb ishaemia is analyzed. Different surgical policy depending on the type and length of atherosclerotic lesion and somatic state is offered. For persons in especially grave somatic state extraanatomical bypass operations (90 patients) were recommended, their technique was discussed in detail. Causes of postoperative complications were analyzed and prophylactic measures suggested. Prevention and treatment of infectious complications were considered. PMID- 12109190 TI - [Diagnosis and treatment of abdominal aortic aneurysm in elderly and older patients]. AB - 83 patients over 60 years with abdominal aortic aneurysms (AAA) were treated from 1991 to 2000. 87.9% patients were men. Stable AAA were seen in 22 (26.5%) patients, 14 of them with small AAA were observed in terms from 1 month to 3 years. Three patients required surgery due to an increase of AAA size. Surgical treatment was performed in 83.1% patients, it was elective in 8 (11.6%), urgent- in 27 (39.1%), clinical picture of rupture was in 34 (49.3%) patients. There were no lethal outcomes in elective operations, lethality in urgent operations was 18.5%, in emergency--85.3%. Screening examinations in aged patients promote early diagnosis and improve results of treatment. PMID- 12109191 TI - [Abscessed cyst of the urachus, complicated by peritonitis]. PMID- 12109192 TI - [Endovascular treatment of giant aneurysm of the left renal artery complicated by its rupture]. PMID- 12109193 TI - [Prospects of computer tomography in diagnosing adrenal tumors]. AB - Computed tomography (CT) is one of main topical methods of adrenal tumors diagnosis. Medical histories of 68 patients treated from 1995 to 2000 for various adrenal tumors were analyzed. CT was used in all the patients. These data were compared with results of morphological study. It is demonstrated that CT has high sensitivity in diagnosis of adrenal tumors (96.55% cases). It permits to predict the type of tumor with high probability. The degree of tumor size mistake made at CT was estimated. Mean coefficients to assess true sizes of tumors before surgery were calculated. This is important for choice of treatment policy in hormonal unactive adrenal tumors. PMID- 12109194 TI - The Wisconsin Cardiovascular Health Program: a partnership effort to improve the health of Wisconsin residents. AB - The CHP is actively engaging citizens, health professionals, and organizations as partners to help change community and patient behaviors and risk factors that may lead to heart disease and stroke and to develop environmental strategies and cardiovascular policy initiatives. We welcome additional physician involvement throughout the state in this important initiative. PMID- 12109195 TI - Regional variations in coronary heart disease mortality in Wisconsin, 1979-1998. AB - BACKGROUND: Although coronary heart disease mortality has declined in Wisconsin during recent decades, progress in reducing mortality was not equal in all counties. METHODS: Expected and observed numbers of coronary heart disease deaths from 1994 to 1998 were calculated for all 72 Wisconsin counties using data from the Centers for Disease Control and Prevention. The estimated percent decline in coronary heart disease mortality from 1979 to 1998 was also calculated for each county. RESULTS: Significant regional disparities are apparent in both recent coronary heart disease morality rates and declines in coronary heart disease mortality during the past two decades in Wisconsin. Counties that experienced the smallest declines in reducing coronary heart disease mortality during 1979-1998 tended to have the highest rates at the end of the period. Counties with lower income, lower education levels, less population density, and more blacks had higher rates of coronary heart disease. CONCLUSIONS: These results may be useful in targeting resources to reduce regional disparities in coronary heart disease mortality in Wisconsin. PMID- 12109196 TI - Decline in mortality of coronary heart disease among whites and blacks in Wisconsin 1979-1998. AB - OBJECTIVES: 1) To examine overall trends in Coronary Heart Disease (CHD) mortality in Wisconsin from 1979 to 1998 to assess progress toward Wisconsin's "Public Health Agenda for the Year 2000" goal; and 2) to compare the trends in mortality rates due to CHD by race, gender, and age groups between the periods of 1979-1983 and 1994-1998. METHODS: Mortality data for CHD (ICD codes 410-414, 429.2) were retrieved from the Center for Disease Control and Prevention's WONDER database. Age-specific CHD mortality rates were calculated as averages over the 5 year periods, 1979-1983 and 1994-1998. Both the percentage change in mortality rates between the two periods and the number of lives saved during the 1994-1998 period as a result of change were obtained. To look at racial disparities in mortality, relative rates, that is, ratios between mortality rates of blacks and whites in the 1994-1998 period, were calculated. RESULTS: Between the two time periods (1979-1983 and 1994-1998) there was a 40% reduction of CHD mortality in most age groups for white men and women. The slowest decline in mortality occurred among black men: 11% in the 35-44 age group and less than 30% among those 55 years and older. Although the mortality rate declines for black women over the study period were similar to those for whites overall, in the 1994-1998 period mortality rates among younger black women aged 35 to 54 were 2 to 4 times higher than those for white women in the corresponding age groups. CONCLUSION: Wisconsin achieved its year 2000 objective for CHD mortality reductions in 1998, experiencing a 49% decline in CHD mortality from 1979 to 1998. Despite this overall decline, however, there was a noticeable difference in the trends among demographic groups, suggesting a further focus on racial disparities in CHD mortality. PMID- 12109197 TI - Regional variation in stroke mortality in Wisconsin, 1989-1998. AB - OBJECTIVE: To evaluate the regional variation in stroke mortality and determine the level of excess stroke mortality in Wisconsin over the period 1989-1998. METHODS: Wisconsin stroke mortality (ICD-9 430-438) data from the Center for Disease Control and Prevention (CDC WONDER) were analyzed by county for the period 1989-1998 using indirect age standardization. RESULTS: Estimates of observed and excess number of deaths associated with cerebrovascular disease in Wisconsin varied considerably by county during the 10-year time frame studied. Twenty-five counties had an observed number of stroke deaths that were statistically significantly different from what was expected given the age structure of their population. Of these, 14 had significantly more deaths than expected while 11 had significantly fewer. DISCUSSION: There is substantial variation in stroke mortality between Wisconsin counties. Potential reasons for regional variation are unknown and warrant further research. This paper may give Wisconsin counties a benchmark of their progress in preventing stroke mortality to date and give direction for future public health efforts. PMID- 12109198 TI - Progress in reducing stroke mortality in Wisconsin, 1984-1998. AB - BACKGROUND: Despite recent advances in the prevention and treatment of stroke, it remains one of the leading causes of death in Wisconsin and the nation. This report examines trends in stroke mortality in Wisconsin over the past two decades and assesses progress toward reaching the Healthier People in Wisconsin 2000 objective of a 33% reduction in stroke mortality. METHODS: Trends in stroke mortality for Wisconsin were examined across gender, age, and racial subgroups. Stroke mortality rates (ICD-9 430-438) were extracted from the Centers for Disease Control and Prevention's database via WONDER, for the period 1979-1998. RESULTS: Overall stroke mortality rates in Wisconsin decreased 11% from 1984-1986 to 1996-1998. Blacks had higher stroke mortality rates than whites, and males had higher rates than females. From 1984 to 1998, stroke mortality decreased more in men than in women for both races. In blacks under 65, both males and females experienced an increase in stroke mortality from 1984 to 1998. CONCLUSION: This analysis indicates mixed progress in the reduction of stroke mortality in Wisconsin. As of 1998, Wisconsin had not made enough progress to enable it to meet the public health agenda goal for the year 2000. PMID- 12109199 TI - Second case of infective endocarditis caused by Gemella sanguinis. AB - BACKGROUND: A 69-year-old man presented with a history of spiking fevers and symptoms of transient expressive aphasia and a left hemispheric cerebrovascular accident. A transthoracic echocardiogram revealed thickened mitral leaflets. A subsequent transesophageal echocardiogram demonstrated vegetations on both mitral leaflets and severe mitral regurgitation by color flow imaging. A gram-positive coccus was isolated from the blood culture of this patient. METHODS: The bacterium was identified by polymerase chain reaction (PCR) amplification of its 16S rRNA gene with the broad range eubacterial primers, FD1 and RD1, followed by sequencing of the PCR product. The obtained sequence was compared to all sequences deposited in GenBank and the ribosomal database project II. A phylogenetic tree was created to determine the relatedness of this bacterium to other bacterial species. RESULTS: The phylogenetic tree created from the 1389 bp 16S rDNA sequence indicated that the endocarditis was due to Gemella sanguinis, a member of the normal oral flora and a rare cause of bacteremia. CONCLUSION: This report describes the second case of endocarditis caused by G. sanguinis. A history of periodontal disease and tooth abscess in this patient suggests that the oropharynx was the probable origin of the Gemella bacteremia. We propose that G. sanguinis should be added to the list of Gemella species that can cause endocarditis. PMID- 12109200 TI - Low back pain therapy ineffective? PMID- 12109201 TI - A review of clinically relevant cardiac biochemical markers. AB - Acute coronary syndromes remain the leading cause of mortality in the United States and represent an enormous cost to the health care system. Despite decades of investigation into the diagnosis of acute myocardial infarction (MI), the diagnostic process is still quite complex because the majority of patients with chest pain fall in the low or medium risk category with atypical symptoms and nonspecific electrocardiogram (ECG) changes. Cardiac biochemical markers play an important role in helping physicians make the diagnosis and stratify the risk to patients. However, the ideal cardiac marker and the best diagnostic approach to patients with chest pain in the Emergency Department (ED) remain elusive. Currently, among many cardiac markers, cardiac troponin I seems the most cardiac specific in the diagnosis of acute MI. This article is focused on reviewing the characteristics of different cardiac markers and comparison of their usages in the diagnosis of acute MI. However, since this field is large and rapidly expanding, it is impossible to cover every aspect of cardiac markers. As the search for the most efficacious, specific and cost-effective means to approach patients with chest pain continues, further prospective, randomized, multicenter trials are needed to confirm the value of troponins and other diagnostic strategies in the early diagnosis of acute MI. PMID- 12109203 TI - Dilemmas in geriatrics: scenario 3 results. PMID- 12109202 TI - Evolving clinical applications of cardiac markers: a review of the literature. AB - Ischemic coronary syndrome is still the most frequent cause of mortality in the United States. Despite extensive investigation into the diagnosis of acute myocardial infarction (MI), this process remains quite complex because the majority of patients with chest pain have atypical symptoms and nonspecific electrocardiogram (ECG) changes and fall in the low or medium risk category. The biochemical cardiac markers play an important role in helping physicians make the diagnosis of acute MI and stratify patients for risk modifications. The use of cardiac markers in the diagnosis of acute MI is discussed extensively on pages 36 44 of this issue. This article focuses on reviewing other clinical applications. This field is large and rapidly expanding, making it impossible to cover every single clinical application of cardiac markers. Several research efforts are underway to find better cardiac markers and develop new applications for them. We review the literature on the use of cardiac markers, particularly troponin, with the risk stratification of acute coronary syndromes, the detection of reperfusion, perioperative MI, periprocedural MI, myocarditis, cardiac contusion, and MI associated with sepsis. PMID- 12109204 TI - The spectroscopy of molecular reaction intermediates trapped in the solid rare gases. AB - The trapping of neutral and electrically charged molecular reaction intermediates in the solid rare gases and the characterization of these intermediates by vibrational and electronic spectroscopy are surveyed. Spectral data for reaction intermediates trapped in solid neon and argon are compared with the corresponding data obtained from gas phase studies and from quantum chemical calculations. Emphasis is placed on recent progress, including the production, stabilization, and spectroscopic study of highly reactive small molecular ions and the use of ab initio and density functional calculations for the identification of reaction intermediates. PMID- 12109205 TI - Rigidization, preorientation and electronic decoupling--the 'magic triangle' for the design of highly efficient fluorescent sensors and switches. AB - One of the key interests in the recent development of fluorescent molecular sensors and switches is the realization of systems that show strong signal changes as a response to the chemical trigger. Aiming at rational probe design, this article compiles and compares different promising strategies to extract those supramolecular and photophysical features that allow the construction of molecular devices suitable for efficient signaling. The examples comprise fluorescence 'OFF'-'ON' as well as 'ON'-'OFF' operative systems and the mechanisms, properties, and limitations of the different design concepts are discussed. PMID- 12109206 TI - The emerging chemistry of blood product disinfection. AB - Given the importance and limitations of the blood supply worldwide, widely applicable procedures for the inactivation of pathogens (viruses, bacteria, protozoa etc.) in donated blood and blood products are now required. Conventional treatments such as ultraviolet irradiation cause damage to therapeutic components in the blood, so more targeted approaches are being sought. These include targeted chemotherapy, photochemotherapy and photodynamic antimicrobial chemotherapy (PACT). PMID- 12109207 TI - Catalysis inside dendrimers. AB - Catalytic sites can be placed at the core, at interior positions or at the periphery of a dendrimer. There are many examples of the use of peripherally functionalized dendrimers in catalysis and this subject has been thoroughly reviewed in the recent literature. This review is concerned only with dendrimer based catalysis involving catalytic sites at the core of a dendrimer and within the interior voids. In covering the significant achievements in this area, we have concentrated on examples that highlight key features with respect to positive and/or negative catalytic activity. PMID- 12109208 TI - Bringing time resolution to EXAFS: recent developments and application to chemical systems. AB - The currently available methodologies for applying extended X-ray absorption fine structure (EXAFS) spectroscopy to the interrogation of dynamic chemical systems are critically reviewed. Particular regard is paid to applicable experimental timescales, extraction of structural information, sample presentation, and simultaneous pairing with complementary techniques; the potential for further developments in this area is also assessed. PMID- 12109209 TI - Mechanically interlocked molecules incorporating cucurbituril and their supramolecular assemblies. AB - Mechanically interlocked molecules incorporating cucurbituril (CB[6]) as a molecular 'bead' and their supramolecular assemblies are described. An efficient synthesis of 1D, 2D and 3D polyrotaxanes with high structural regularity and molecular necklaces has been achieved by a combination of self-assembly and coordination chemistry. The functional aspects of these interlocked molecules and their supramolecular assemblies, including molecular machines and switches based on [2]rotaxanes, a 2D polyrotaxane with large cavities and channels, pseudorotaxane-terminated dendrimers, and interaction of pseudorotaxanes containing polyamines and CB[6] with DNA are also described. PMID- 12109210 TI - Separating fact from fiction: assessing the potential of modified adenovirus vectors for use in human gene therapy. AB - One of the major hurdles to successful gene therapy of genetic and/or acquired disease is the ability to efficiently introduce a foreign gene into the tissue of interest and, in the case of some genetic diseases, achieve long-term expression of the transgene. Due to their ability to transduce a wide variety of cell types in a cell-cycle independent fashion, adenovirus (Ad)-based vectors have received considerable attention in recent years as delivery vehicles for multiple gene therapy applications. Effective use of early "first-generation" versions of these vectors was hampered by not only the induction of strong immune responses in the host to the Ad vector and transduced cells, but also to direct acute and chronic toxicity caused by the vector itself. Furthermore, transgene expression was typically transient, lasting only a few weeks. Despite these limitations, these vectors have been used in a number of human clinical trials, eliciting both interesting as well as controversial results, some of which are summarized herein. Because of these limitations, a number of advances in adenovirus "vectorology", manifested primarily as the development of multiply attenuated Ads and vectors deleted of all viral protein coding sequences, has resulted in vectors which retain all of the advantages of Ad vectors and, in addition, do not exhibit the deleterious characteristics associated with [E1-]deleted Ads. This review focuses on the current state of the art regarding the potential for human use of Ad-based vectors, and how the use of this vector continues to offer the potential for successful use as a gene delivery tool for the treatment of a great number of human genetic and non-genetic diseases. PMID- 12109211 TI - Adenovirus as an integrating vector. AB - Recombinant adenoviral vectors have served as one of the most efficient gene delivery vehicles in vivo thus far. Multiply attenuated or completely gutless adenoviral vectors have been developed to achieve long-term gene expression in animal models by overcoming cellular immunity against de novo synthesized adenoviral proteins. However, since adenovirus lacks native integration machinery, the goal of gene therapy obtaining permanent expression cannot be realized with current adenoviral vector systems. Recent studies have shown that replication-incompetent adenoviral vectors randomly integrate into host chromosomes at frequencies of 0.001-1% of infected cells. To improve the integration frequencies of adenoviral vectors, a variety of hybrid vectors combining the highly efficient DNA delivery of adenovirus with the integrating machinery of retroviruses, adeno-associated viruses, and transposons, have been emerging. These hybrid vectors have shown promise, at least in in vitro systems. Furthermore, adenoviral vectors have shown potential as gene targeting vectors. These developments should eventually lead to more effective gene therapy vectors that can transduce a myriad of cell types stably in vivo. PMID- 12109212 TI - Second generation adeno-associated virus type 2-based gene therapy systems with the potential for preferential integration into AAVS1. AB - Adeno-associated virus type 2 (AAV-2) is a non-pathogenic human parvovirus that is being developed as a gene therapy vector for the treatment of numerous diseases. One property of wild-type AAV-2, that is highly desirable in a gene therapy vector, is its ability to preferentially integrate its DNA into a 4 kilobase region of human chromosome 19, designated AAVS1. One disadvantage of AAV 2 is its relatively small packaging capacity, approximately 4.7 kilobases. Because of this size limitation, the AAV-2 rep and cap genes were removed from first-generation AAV-2-based gene therapy vectors to make room for the therapeutic or marker gene. It was later discovered that the rep gene, or at least one of its products, the Rep68 or Rep78 protein, is required for preferential integration of AAV-2. Recent developments in AAV-2 gene therapy vector construction allow the inclusion of the rep gene into a second generation of AAV-2-based gene therapy systems. These new systems fall into four major categories: plasmid-based systems, co-transduction with multiple AAV-2 vectors, incorporation of the AAV-2 vector into a larger virus, and in vitro packaging. These systems not only allow the inclusion of the rep gene, they also allow the delivery of larger therapeutic genes. PMID- 12109213 TI - Advances in the development of non-human viral DNA-vectors for gene delivery. AB - Within the last two decades, various vectors based on human viruses have been developed as gene transfer vehicles for gene therapy applications and vaccination. However, one yet unresolved problem connected to the use of viral vectors in humans is the pre-existing immunity to most of these vectors in the vast majority of the population which can result in impaired gene transfer efficiency and increased secondary toxicity. One approach to solve this problem is the development of recombinant viruses of non-human origin as vectors for gene transfer. The major rationale for using such vectors is the avoidance of vector neutralization by pre-existing antibodies directed against the virus on which the vector is based. Use of vectors based on non-human viruses may therefore allow the use of lower initial vector doses to achieve efficient gene transfer. Side effects caused by interactions between vectors derived from human viruses with a primed immune system or with blood components could also be reduced. Furthermore, these vectors might show new cell type tropisms and could therefore infect tissue and organs that are not accessible to current viral vectors. This review outlines some of the problems inherent in the human origin of current viral vectors and describes features and progress with non-human adenovirus and baculovirus-derived vectors that may provide alternatives. PMID- 12109214 TI - CFTR gene transfer to lung epithelium--on the trail of a target cell. AB - Cystic fibrosis (CF) is a lethal inherited disease that afflicts up to 1 in 2,500 people in the western world. Since 1989, when mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene were identified as responsible for the disease, intense effort has been applied to the development of replacement gene therapy strategies to cure CF. Problems with basic gene delivery techniques along with limited knowledge of the pathogenesis of CF have hindered progress so far. However, recent insights into the expression patterns and functions of CFTR in developing and adult lungs are now advancing our understanding of this disease. It is becoming apparent that progress in gene delivery to cure CF may be best served by identification of a target cell(s) around which gene transfer strategies can be specifically tailored to most closely reproduce the effects of normal CFTR expression. In fact, accurate restoration of endogenous expression patterns may be crucial, not only for disease reversal, but also to avoid potentially deleterious effects of inappropriate expression. This approach is in turn confounded however, by ill defined stem and progenitor cell pathways within the lung epithelium. Nonetheless, studies to date suggest that these pathways are relatively plastic and may respond differently during homeostasis compared with repair following injury. It may therefore be feasible to target the lung epithelium in a non-cell specific manner and allow endogenous differentiation pathways to subsequently establish correct CFTR distribution patterns. In this review, emerging information on CFTR expression and function in developing and adult lungs is discussed in the context of putative stem cell populations and their potential for current gene delivery approaches. PMID- 12109215 TI - Intracellular barriers to non-viral gene transfer. AB - Non-viral vector mediated gene transfer, compared to viral vector mediated one, is a promising tool for the safe delivery of therapeutic DNA in genetic and acquired human diseases. Although the lack of specific immune response favor the clinical application of non-viral vectors, comprising of an expression cassette complexed to cationic liposome or cationic polymer, the limited efficacy and short duration of transgene expression impose major hurdles in the widespread application of non-viral gene therapy. The trafficking of transgene, complexed with chemical vectors, has been the subject of intensive investigations to improve our understanding of cellular and extracellular barriers impeding gene delivery. Here, we review those physical and metabolic impediments that account, at least in part, for the inefficient translocation of transgene into the nucleus of target cells. Following the internalization of the DNA-polycation complex by endocytosis, a large fraction is targeted to the lysosomal compartment by default. Since the cytosolic release of heterelogous DNA is a prerequisite for nuclear translocation, entrapment and degradation of plasmid DNA in endo lysosomes constitute a major impediment to efficient gene transfer. Only a small fraction of internalized plasmid DNA penetrates the cytoplasm. Plasmid DNA encounters the diffusional and metabolic barriers of the cytoplasm, further decreasing the number of intact plasmid molecules reaching the nuclear pore complex (NPC), the gateway of nucleosol. Nuclear translocation of DNA requires either the disassembly of the nuclear envelope or active nuclear transport via the NPC. Comparison of viral and plasmid DNA cellular trafficking should reveal strategies that viruses have developed to overcome those cellular barriers that impede non-viral DNA delivery in gene therapy attempts. PMID- 12109216 TI - Bone marrow stromal cells as targets for gene therapy. AB - The bone marrow (BM) is composed of the non-adherent hematopoietic and adherent stromal cell compartment. This adherent BM stromal cell fraction contains pluripotent mesenchymal stem cells (MSCs) and differentiated mesenchymal BM stromal cells. The MSCs self-renew by proliferation while maintaining their stem cell phenotype and give rise to the differentiated stromal cells which belong to the osteogenic, chondrogenic, adipogenic, myogenic and fibroblastic lineages. A more primitive adherent stem cell was recently identified, the multipotent adult progenitor cell (MAPC) or mesodermal progenitor cell, which co-purifies with MSCs. These MAPCs differentiate into MSCs, endothelial, epithelial and even hematopoietic cells. BM stroma cells, including the primitive pluripotent MSCs and MAPCs, are attractive targets for cell and gene therapy. The BM stromal cell population and its multipotent stem cells can be engineered to secrete a series of different proteins in vitro and in vivo that could potentially treat a variety of serum protein deficiencies and other genetic or acquired diseases, including bone, cartilage and BM stromal disorders or even cancer. PMID- 12109217 TI - The recombinant T cell receptor strategy: insights into structure and function of recombinant immunoreceptors on the way towards an optimal receptor design for cellular immunotherapy. AB - A promising approach in adoptive immunotherapy is based on the induction of a specific cellular anti-tumor response by antigen-specific, cytolytic T cells. Due to difficulties in isolating tumor-specific T cells in sufficient amounts, it was proposed to graft cytolytic T cells with an antigen-specific, recombinant T cell receptor. The antigen binding domain of the receptor consists of a single-chain antibody fragment (scFv) that is derived from a monoclonal antibody and binds to a tumor associated antigen. The intracellular signalling domain is derived from the cytoplasmic part of a membrane bound receptor to induce cellular activation, e.g., the Fc epsilon RI receptor gamma-chain or the CD3 zeta-chain. By use of this type of recombinant receptor, the strategy combines the advantages of MHC independent, antibody-based antigen binding with efficient T cell activation upon specific binding to the receptor ligand. The modular composition of the receptor, moreover, facilitates modification of both the antigen binding and signalling properties. Accordingly, we and others have generated a panel of recombinant T cell receptors with specificities for malignantly or virally transformed cells. Receptor grafted effector cells were demonstrated to mediate a highly efficient immune response towards antigen expressing target cells. However, little is known about the impact of the recombinant receptor modules on recognition of highly heterologous target antigens and on cellular activation in a complex immunological context. This review summarizes the current knowledge about the generation and function of recombinant immunoreceptors and discusses the limitations and perspectives of the methodology for use in cellular immunotherapy. PMID- 12109218 TI - Restoration of transgene expression in hematopoietic cells with drug-selectable marker genes. AB - Somatic gene therapy is supposed to cure life-threatening hematopoietic disorders but is limited by unstable transgene expression. Efficient gene transfer to hematopoietic progenitor cells does not ensure long-term gene expression. It would therefore be advantageous if the expression of transgenes could be restored in bone marrow. Transfer of drug resistance genes such as the multidrug resistance (MDR1) or mutated dihydrofolate reductase (DHFR) genes to hematopoietic cells protects them from the toxicity of anticancer drugs. In addition, transduced cells obtain a selective growth advantage in the presence of anticancer drugs. This can be used to introduce and enrich otherwise non selectable genes by cotransfer to target cells. Bicistronic vectors have been constructed for coexpression of drug resistance genes and non-selectable, therapeutic genes with the use of an internal ribosomal entry-site (IRES). With the use of bicistronic vectors, expression and function of therapeutic genes have been increased in tissue culture and in animal models. Further preclinical investigations are needed to identify optimal conditions for selection. PMID- 12109219 TI - Genetic vaccination for the immunotherapy of B-cell malignancies. AB - Vaccination protocols based on targeting of the idiotype expressed on malignant B cells have so far provided encouraging results in clinical trials. The essential requirement to induce an immune response is the inclusion of carriers to overcome T-cell tolerance. Chemical cross-linking of idiotypic protein is so far the method of choice to induce protective responses in human studies. Meanwhile, a flurry of alternative strategies to simplify vaccine production is being tested in murine model. Thanks to the advance in antibody engineering the two relevant antigenic domains of the lymphoma immunoglobulin can be assembled into an appropriate format, genetically linked to molecules that act as immunological adjuvants and directly delivered as plasmid DNA. Upon immunization, rejection of tumor cells may depend on cellular or humoral mechanisms, whose relative importance has not been entirely estimated. We have recently analyzed the specificity of anti-idiotypic antibodies induced by DNA vaccination and characterised the elements contributing to optimal anti-idiotypic responses. PMID- 12109220 TI - Oncolytic viruses: programmable tumour hunters. AB - Despite significant improvements in early detection and refinements of therapeutic protocols over the last several decades, cancer remains one of the leading causes of death in North America. In particular, treatment of metastatic cancers is a highly desirable and yet still elusive goal of the oncologist. One strategy which holds promise is the use of self replicating viral strains with the ability to specifically kill tumour but not normal cells. These so-called "oncolytic viruses" are in general, attenuated for growth in normal cells but are able to exploit tumour specific, genetic defects to gain a growth advantage. In this review, we will discuss the virus:host cell interactions which help form the niche occupied by oncolytic viruses. The current and potential clinical applications/limitations will be discussed for oncolytic viruses from the herpesvirus, adenoviruses, picornavirus, rhabdovirus, and paramyxovirus families. PMID- 12109221 TI - Cancer gene therapy with tissue inhibitors of metalloproteinases (TIMPs). AB - Matrix metalloproteinases (MMPs) are of crucial importance for the invasive behavior of primary tumors and their metastases. MMP activity is regulated by the four naturally occurring tissue inhibitors of metalloproteinases (TIMPs). It has been shown that overexpression of TIMPs in tumors of various origins leads to reduced tumor growth and formation of metastases. More recently, antitumor efficacy by in vivo gene transfer of TIMPs has been reported in several clinically relevant animal models. This review analyses the therapeutic potential of the TIMPs from a cancer gene therapeutic point of view with particular emphasis on cell culture and in vivo data. PMID- 12109222 TI - [Laparoscopic colostomy: experience in patients with ovarian or ano-rectal cancer, non-operable or with rectovaginal fistula]. AB - Between August 1995 and May 2001 laparoscopic colostomy was successfully carried out in 23 patients with advanced ovarian cancer, inoperable carcinoma of the anorectum or rectovaginal fistulas. There were no intraoperative or postoperative complications and postoperative recovery was rapid with all patients having function of the colostomy within 24 hrs and regaining their preoperative state of mobility on the second postoperative day. The laparoscopic approach allows the careful selection of the colostomy site, easy mobilisation of the colon, causing only little disruption to the intestinal function hence improving postoperative recovery. From Authors' experience, laparoscopic colostomy is a simple and safe operation in most cases and can be used as the preferred technique of intestinal diversion. PMID- 12109223 TI - [Color-Doppler sonographic mapping for the choice of treatment in variceal disease]. AB - The Authors, after having reported Huch's classification concerning both large and small saphenas varixes, make a short historical excursus about the most significant operative methods. Then they start explaining how a correct therapeutic approach can't be done without a careful hemodynamic study. After having given data about their case-report, they conclude affirming how today the duplex-scanner permits to obtain an excellent radicality also with not very invasive techniques. PMID- 12109224 TI - [Surgical treatment of varicocele in day surgery]. AB - Aim of this study is the comparison between spermatic vein ligature (Ivanissevich technique) and low intrafunicular ligature in the surgical treatment of varicocele. The Authors present two groups of patients (total 61) operated between 1995 and 1999 with the above-mentioned surgical techniques. Apart from the utilized method, they didn't ascertain any improvements in the spermiograms of the patients older than 30 years. The low ligature of the spermatic veins demonstrated the advantages of surgery treatment and lower percentage of relapses. PMID- 12109225 TI - NM23 expression as prognostic factor in colorectal carcinoma. AB - BACKGROUND: The current most important prognostic indicator in colorectal carcinoma is tumor stage at the moment of diagnosis. The role of NM23 gene as prognostic factor is controversial. The aim of this study was to investigate NM23 expression. PATIENTS AND METHODS: The study population included 104 unselected patients who underwent surgery for colorectal carcinoma between 1992 and 1997. NM23 expression was quantified by estimating the percentage of tumor cells with unequivocal reactivity. The percentage was scored: 0 when no tumor cells showed immunoreactivity; 1 when less than 10% of cells showed immunoreactivity; 2 when 11-50% of cells were positive; 3 when more than 51% of cells were positive. RESULTS: Four cases belonged to group 0.21 to group 1.55 to group 2 and 24 to group 3. CONCLUSION: NM23 cannot be considered an independent prognostic variable. PMID- 12109226 TI - [Polyembrioma of the testis: case report following chemotherapy for non-Hodgkin's lymphoma]. AB - Testicular tumours represent 2% of all male malignancies, mostly concerning young men (20-40 years old). The polyembryoma is one of the uncommonest lesions and just recently it has been identified as autonomous nosographic entity. The reported case is peculiar because the patient was older than the most ones described in the literature and the tumour arose after polychemotherapy for non Hodgkins' disease. The Authors analyse some aspects concerning etiology, pathology and clinical approach to such rare neoplasm. PMID- 12109227 TI - [Primary gastric lymphoma: a case report]. AB - In this case report the Authors describe a case of primary gastric lymphoma in a 62 years old patient who presented with dyspepsia and weigh loss. Primary gastric lymphoma is a rare neoplasm which of 1-10% of the malignant gastric neoplasms in the gastroenteric tract. The clinic presentation is usually aspecific. The infection by H. pylori is a factor of predisposition for this kind of disease. The diagnostic pathway consists in x-ray examination of the gastrointestinal tract, the endoscopy with biopsies, the computerized tomography and the echo endoscopy. However obtaining a preoperative diagnosis is often difficult because of the submucosal localization of the lymphoma. There is not a common strategy among the Authors for the treatment of the disease, which can be surgical, radiotherapic or chemotherapic. PMID- 12109228 TI - [Sarcoidosis and clear cell carcinoma of the kidney: the sixth case?]. AB - The Authors report a very rare case of clear cell renal cancer associated with sarcoidosis, incidentally discovered in a 39 year-old man, admitted for a not correlated pathology (multiple left rib fractures due to automobile crash). Problems related to a proper assessment of sarcoidosis are discussed as well as potential arising of a neoplasm during the entire follow-up period: for that, it must always be complete and accurate. PMID- 12109229 TI - [Results of surgical treatment of acute cholecystitis. Prospective study of 280 cases]. AB - The Authors reported the results of surgical treatment of acute gallstone cholecystitis (AGC) in patients in whom different selection criteria have been applied. Two-hundred-eighty patients with a clinical and/or ultrasonographic diagnosis of ALC were admitted to the 1st Division of General Surgery-University of Verona Italy between January 1992 and June 2001, the patients were divided into five groups according to clinical features, laboratory tests and echographic signs. A specific approach was used in the different groups. An urgent laparoscopic cholecystectomy was performed in 67 patients. Elective laparoscopic treatment after urgent US guided percutaneous cholecystostomy (US-PC) was performed in 119 and after US-PC and ERCP in 50 patients. Laparoscopic cholecystectomy was performed in 236 patients with a conversion rate of 7.6%. No mortality, 6.7% morbidity and a mean hospital stay of 7.5 days. A selective therapeutic approach to AGC allow immediate treatment in all cases and correct diagnosis of associated diseases treatment. This approach makes it possible to reduce the conversion rate of laparoscopic cholecystectomy, morbidity and mortality. PMID- 12109230 TI - [Retroperitoneal leiomyosarcoma: clinical case]. AB - The Authors have reported a case of retroperitoneal leiomyosarcoma. The retroperitoneal localization is quite unusual and early diagnosis is difficult. Only surgery operation and radio-chemotherapy can improve the prognosis. Tumor size is the major prognostic factor. PMID- 12109231 TI - [Complications of inguinal hernia repair]. AB - It's shown by literature and confirmed by Author's experience that, on account of the excellent results, prosthetic repair of inguinal hernia is more effective than "conventional" (Bassini, Mc Vay, Shouldice). Between January 1993 and December 2000 were observed 875 patients with inguinal hernia (814 monolateral, 61 bilateral); all patients underwent a Lichtenstein repair both in the primary version and in its variations (internal ring plastic, trasversalis plicate, plug repair). The patients were discharged from hospital within 24 hours after surgery in 90% of cases. No important intraoperative complications were observed; the patients restarting work varied from 3 to 15 days after the discharging in relation to patient anxiety, onset of complications and to the type of work. The complications observed were: urine retention (1.6%), superficial haematoma (1.3%), superficial infection (1%), wound suppuration (0.5%), serous effusion (0.7%), postsurgery pain (2.1%), scrotal edema (1.7%), persistent inguinal neuralgia (0.6), local hypoesthesia (4.3%), ischemical orchitis (0.1%), recurrence (0.2%). In conclusion Authors assert that "tension free" repair allows optimal results both for the surgery point (easiness of the technique, repeatability, less invasivity, scanty incident of recurrences, low frequency of postoperative complications) and in economic terms, allowing an early mobilization of the patients. A further improvement would be obtained with more care in surgical and patient management, with more excellent results. PMID- 12109232 TI - Presacral myelolipoma. A case report. AB - Presacral myelolipoma is a rare benign tumour composed of fat and haemopoietic tissue. Ultrasound, computed tomography and magnetic resonance imaging are of help to achieve the diagnosis, but pathologic confirmation is mandatory. The Authors report an asymptomatic case whose diagnosis has been achieved by means of CT scan-guided percutaneous needle biopsy. Unnecessary surgical treatment was avoided in this case. Clinical approach and role of surgery are discussed. PMID- 12109233 TI - [Apudoma of Vater's ampulla: case report and review of the literature]. AB - The Authors report a case of Vater's ampulla apudoma and after having examined the characteristics of these neoplasms they discuss clinical presentation, diagnostic and treatment problems of islet cell adenomas. They review the literature and make some remarks. PMID- 12109234 TI - A comparison of the effects of manual and ventilator hyperinflation on static lung compliance and sputum production in intubated and ventilated intensive care patients. AB - BACKGROUND AND PURPOSE: Lung hyperinflation is a technique used by physiotherapists to mobilize and remove excess bronchial secretions, reinflate areas of pulmonary collapse and improve oxygenation. Hyperinflation may be delivered by the ventilator or manually, by use of a manual resuscitation circuit, depending upon the respiratory and cardiovascular status of the patient. The effects of manual hyperinflation, with respect to excess bronchial secretions and static lung compliance, have been well-established. There is, however, only limited evidence as to the efficacy of ventilator hyperinflation as a physiotherapy treatment technique. The purpose of the present study was to compare the effects of manual hyperinflation and ventilator hyperinflation on static pulmonary compliance and sputum clearance in stable intubated and ventilated patients. METHOD: Twenty patients who met the inclusion criteria were studied. This was a double crossover study where all patients were randomly allocated to one of two treatment sequences over two days. The first sequence involved manual hyperinflation followed two hours later by ventilator hyperinflation and the order was reversed on the second day. In the second sequence, ventilator hyperinflation preceded manual hyperinflation. The variables of static pulmonary compliance and sputum wet weight were analysed by use of an analysis of variance (ANOVA) for repeated measures. Statistical significance was set at p < 0.05. RESULTS: There was no significant difference in sputum wet weight production between either technique or on either day of treatment. Static pulmonary compliance improved with both hyperinflation techniques (p < 0.05). CONCLUSIONS: Hyperinflation as part of a physiotherapy treatment can be performed with equal benefit using either a manual resuscitation circuit or a ventilator. Both methods of hyperinflation improve static pulmonary compliance and clear similar volumes of pulmonary secretions. PMID- 12109235 TI - Feedback withdrawal and changing compliance during manual hyperinflation. AB - BACKGROUND AND PURPOSE: The performance of manual hyperinflation by physiotherapists can be improved by the availability of a pressure manometer. The present study aimed to test whether these benefits could be maintained when the manometer is withdrawn and whether the availability of a manometer affects the pressures delivered under changing respiratory compliances. METHOD: Manual hyperinflation breaths were delivered to a test lung by student physiotherapists, with a target peak airway pressure of 30 cm H2O under control, feedback and feedback-withdrawal conditions. The breaths were delivered for three trials under each testing condition at each of three respiratory compliance settings. RESULTS: The availability of augmented feedback increased the accuracy and reduced the variability of performance; however, these improvements were not maintained when feedback was withdrawn. Changing respiratory compliance significantly affected the accuracy and variability during the control and withdrawal conditions, but the availability of a manometer negated these differences. CONCLUSIONS: The availability of a pressure manometer negates the influence of respiratory compliance on the achievement of target peak airway pressures during manual hyperinflation in the laboratory environment, however these benefits are not retained when feedback is withdrawn. Therefore, it is recommended that a pressure manometer should be routinely available during manual hyperinflation in clinical practice to optimize treatment safety and effectiveness. PMID- 12109236 TI - Instructing pelvic floor contraction facilitates transversus abdominis thickness increase during low-abdominal hollowing. AB - BACKGROUND AND PURPOSE: Low abdominal hollowing in four-point kneeling is used clinically to test and rehabilitate transversus abdominis (TrA) but many people find this exercise difficult to perform. Contracting pelvic floor muscles (PF) during low abdominal hollowing may facilitate contraction of TrA. Thickness increase in the abdominal muscles during low abdominal hollowing has been measured with real-time ultrasound scanning and may indicate muscle contraction. The present study investigated the effect of instructing PF contraction on TrA thickness increase during low abdominal hollowing. METHOD: Twelve females and eight males with no reported pelvic floor dysfunction or low back pain in the last two years were taught low abdominal hollowing in four-point kneeling. Subjects performed low abdominal hollowing with and without instruction to contract PF in random order. Transversus abdominis, obliquus internus (OI) and obliquus externus (OE) thickness were measured with ultrasound scanning at rest and during both tests. RESULTS: Mean increase in TrA thickness during low abdominal hollowing was 49.71% (SD 26.76%), during low abdominal hollowing with PF it was 65.81% (SD 23.53%). Paired Student's t-tests indicated a significant difference between tests (p = 0.015). There were no significant differences between tests for OE or OI thickness increase. CONCLUSIONS: Instructing healthy subjects to co-contract PF results in greater increase in TrA thickness during low abdominal hollowing in four-point kneeling. This may indicate greater contraction of TrA and thus be useful for clinicians training TrA. Further research could investigate the validity of change of thickness as a measure of abdominal muscle contraction, investigate the effect of instructing PF co contraction on TrA in patients with low back pain and measure PF and TrA activity simultaneously. PMID- 12109237 TI - Neurological rehabilitation: a science struggling to come of age. AB - Over the last few decades, there have been considerable improvements in the outcome of stroke patients both as regards mortality and disability. At least some of these improvements can be attributed to better organization of services and improved rehabilitation. Many patients, however, remain severely disabled and we will need to develop new strategies in which the focus will be on reversing impairments rather than simply helping patients to adapt to unaltered impairments. For this to happen, neurological rehabilitation research will have to develop therapies that have a clearly defined rationale and are rooted in neurosciences, are clinically described, are addressed to a well-characterized target population and are evaluated using appropriate outcome measures. Few studies at present meet all these criteria. The recent revolution in our understanding of the nervous system as being soft-wired, of the potential for recovery through reorganization and of the central role of afferent information associated with normal activity is ground for optimism and indicates the direction in which future therapies should be sought. The paper considers some approaches to providing appropriate afferent information, including inputs such as that from electrotherapy, novel approaches to assisted activity and constraint induced therapy. We are on the verge of a revolution in neurological rehabilitation. If we exploit the new understanding of the nervous system arising from basic neurosciences in developing and evaluating therapies we should be able to build on the achievements of the last few decades so that fewer of our patients have to carry the burden of severe disability. PMID- 12109238 TI - Correlation of impairment and activity limitation after wrist fracture. AB - BACKGROUND AND PURPOSE: During physiotherapy rehabilitation, judgements about activity limitation are often based on measurements of impairment. The present paper reports an investigation into the relationship between measurements of impairment and activity limitation in people with a fracture of the distal radius. METHOD: Twenty people with a fracture of the distal radius were referred to physiotherapy after conservative management of their fractures. Measurements of impairment (grip strength and range of wrist extension) and activity limitation (Jebsen Test of Hand Function) were taken at the second physiotherapy appointment after cast removal. The scores between these measurements were correlated. RESULTS: There were strong and significant correlations (-0.51 < rs < -0.76) between grip strength and the tasks of the Jebsen Test of Hand Function. The relationship between range of wrist extension and the Jebsen Test of Hand Function was weaker (-0.17 < rs < -0.55). However, subgroup analysis of those subjects with a Colles' type fracture of the distal radius demonstrated significant relationships between wrist extension and three of the seven activity tasks (-0.74 < rs < -0.84). CONCLUSIONS: Preliminary evidence was found to support the physiotherapy practice of use of measurement of impairment to monitor progress in a person with a fracture of the distal radius. PMID- 12109239 TI - Reflections on the direction of research and PRI. PMID- 12109240 TI - Flexible learning: a new approach to public health instruction for APACPH. Asia Pacific Academic Consortium for Public Health. PMID- 12109241 TI - Broadcasting health programmes among audience and professionals in Korea: perceptions and needs. AB - Mass media has become an essential part of modern society, and it should be noted that mass media plays an important role in delivering information even in the area of health. This study was intended to investigate the perceptions of and needs for broadcasting health programmes among audience and related professionals in Korea, which will serve to help in the development of valuable health programmes. Data were collected through a series of self-administered questionnaire surveys conducted on two types of subjects sampled from residents in the areas around Seoul, Korea, and health or broadcasting professionals working on health programmes. There were some discrepancies in the perceptions of and needs for health programmes between audience and professionals, especially the main audience age group and image, needs for topics and mode of health programme. In conclusion, in order to enhance current health programmes which focus on aspects of diagnosis and treatment of diseases, the range of health programmes should be expanded to fulfill various needs. PMID- 12109242 TI - Evaluating health information sites on the Internet in Korea: a cross-sectional survey. AB - Through the internet the public in South Korea has access to a growing supply of information on health and disease. In South Korea an estimated 13.93 million people used the internet in 2000. The number has increased rapidly compared to 1.63 million in 1997. Health information is often said to be one of the most retrieved types of information on the internet. However, a concern has emerged for the quality of health information documents contained on the World Wide Web. Lack of evaluation and oversight, and ease of publication, have led to inaccurate and misleading health-related publications on the Internet. For those seeking easy ways to identify high-quality and reliable information, rating systems to evaluate the quality of health information on the internet should be provided and developed. Given this background, the purpose of this research was to evaluate health information web sites on the internet. In this study we aimed to survey websites providing health information. 440 websites were selected using four search engines, YahooKorea (http://www.yahoo.co.kr), LycosKorea (http://www.lycos.co.kr), Empas (http://www.empas.co.kr), and Naver (http://www.naver.com), to conduct searches in December 2000. General quality criteria were used for the evaluation. These included ownership, currency, authorship, source, feedback mechanism, links, and functionality. More than 50% of web sites did not provide the date of publication or update of information, author and author credentials, references to source, etc. Websites of universities and universities' hospitals were more likely to provide name and type of provider (p < .01), author's name (p < .001), and references to source (p < .01) than other service providers. There is a need for better evidence-based health information as well as a need to develop simple criteria that ordinary people can understand and use. In addition, gateway services that operate a selective process and provide links to other organizations that provide high quality health information should be offered and developed. PMID- 12109243 TI - Assessing public understanding of the policy of separating dispensing and prescribing in Korea: a survey of college students. AB - We investigated the level of public understanding of the 'separation of dispensing and prescribing' health policy in Korea and its associated factors. A questionnaire survey was conducted upon 700 college students at four months after the introduction of the policy. The understanding level was measured using four question items describing the goal and motivation of the policy, and ten items describing its operational rules. For each item, respondents were asked to mark whether the description was true or false. While the goal and motivation of the policy was relatively well informed (mean understanding score: 69.58 out of 100), people did not have a good understanding of the operational details of the policy (mean score: 32.52). The results of regression analyses showed that personal interest and agreement with the need of the policy were the most significant factors affecting the understanding level (p < 0.01). It is concluded that, for other public policies in the future, policy makers in Korea need to develop more effective media communication strategies to effectively inform the public of the practical details of the policy. PMID- 12109244 TI - Development and evaluation of an Internet-based distance learning system for public health in Korea. PMID- 12109245 TI - Viewpoints of Korean senior high school students on school-based sex education. AB - The percentage of sexually experienced young persons in South Korea has increased and the age of first intercourse has lowered significantly. However, these marked changes in the sexual behavior of young people have been taking place when schools in the country still generally lack a realistic sex education program. Given this situation, the aim of this study is to gather student opinions on current school-based sex education in Korea. A self-administered questionnaire survey was conducted in Seoul, Incheon, and Kyunggi Province to assess the status and needs of high school sexuality education. Survey data was obtained from 1,160 senior high school students. In this study 68.8% of boys and 94.4% of girls had some school-based sex or sexuality education. The mean hours of sex education instruction of boys and girls were 2.29 and 3.39 hours respectively. There are several reasons for not being satisfied with sex education in schools. Lack of information was the first reason in all categories. About seventy-four percent of respondents felt that the sex education taught in schools did not cover the information that they want. The second reason was that there was no trained sex education teacher. Lack of materials, interactive teaching method, and time were the other reasons given. They advocated starting sex education in elementary school and covering all topics by the age-appropriate level. More than half supported that sex education should be made compulsory in schools. PMID- 12109246 TI - Patient confidentiality, ethics and licensing in telemedicine. AB - Telemedicine is fast becoming popular in many countries in the world. It has several advantages such as being cost saving and providing better access to health care in the remote areas in many parts of the world. However, it has some disadvantages as well. One of the major problems is the problem of patients' rights and confidentiality in the use of telemedicine. There are no standard guidelines and procedures in the practice of telemedicine as yet. Both the patient and the physician are unsure of the standard of practice and how to maintain confidentiality. The patient is uncertain as to how to protect her/his rights in the use of telemedicine. The issue of litigation is also unclear as to where the physician is practicing when he/she uses telemedicine. Is she/he practicing in the country where the patient is or is the physician practicing in the country of her/his origin? These issues need to be addressed urgently so that telemedicine will have standards of ethical practice and the patient's rights and confidentiality will be protected. PMID- 12109247 TI - Teaching information technology and research skills for public health. AB - The acquisition of information technology and research skills is fundamental for all students in Public Health because of its fundamental emphasis on population studies. In the Curtin University, Division of Health Sciences the School of Public Health has the responsibility of introducing communication and IT skills to all students, both undergraduate and postgraduate. A special program of information technology skills is offered. The age of the Internet has meant added temptation for students in terms of plagiarism and all students and staff must be aware of their ethical obligations in this area. The advent of flexible learning strategies will provide many opportunities for Schools of Public Health, both in the education of their students and in the continuing education of public health practitioners. APACPH member institutions could usefully pool their resources to develop flexible learning resources. PMID- 12109248 TI - Improved interactive medical and public health education in Japan and Korea. AB - We have carried out case based comparative studies in Japan, US and Korea with special reference to the medical education programme using standardized patients (SPs). The schools in US we have visited are still in the process further reforming interactive medical education. In Daegu, Korea, five medical schools have jointly started a working group for the betterment of medical education on a regular basis from the year 1999. The emphasis is on cooperative research on standardized patient training and its application. In Japan, the substantial activities for training and implementing SPs were later followed by several citizens who are active in training and providing SPs to medical schools and hospitals in need. In comparison to the established research oriented framework and flexible application of SPs in US and new & dynamic developmental discussions of SPs in Korea, the Japanese reality is a bit behind with less opportunity for research oriented trials. PMID- 12109249 TI - An evaluation of use of information technology equipment among Japanese elderly women--relation between health status and the preferred input device for the Internet. AB - This study was conducted to determine how mobile phones (MP) and the Internet through personal computers (IPC) have been used among Japanese elderly women. It also aims to examine the relationship between the input devices and factors such as age, health status, and future needs for a MP and the IPC. The magnitude of the relation was evaluated by the odds ratio (OR). The survey was done in December 2000 and January 2001. The study sample was forty women, 78.5 +/- 4.5 years of age (70-89), who lived in their own houses in Y Town near Himeji City, Hyogo, Japan. Findings reveal that use of a MP was 10.0% and none used the IPC. On the other hand, 60.0% of the subjects were interested in using a MP in the future and 27.5% in using the IPC. The touch screen was the most preferred input device among the elderly with high age [OR = 3.86, 95% confidence interval (CI) 0.83-18.98]. It was also preferred by subjects with more difficulties (OR = 5.00, 95% CI 1.05-25.41) and less future need for a MP and the IPC (OR = 7.22, 95% CI 1.34-43.88). On the other hand, the Japanese kana syllabary (JKS) was the most preferred input device among those with a low age (OR = 4.33, 95% CI 0.68-35.00) and with a more future need for a MP and the IPC (OR = 6.33, 95% CI 1.14-39.59). Considering these results, we intend to create a type of keyboard that combines the features of touch screen and JKS for the elderly women. PMID- 12109250 TI - Health development experience in North and South Korea. AB - The purpose of this study is to compare the difference in health status between South Koreans and North Koreans and to identify factors responsible for the remarkable improvements in the health status of South Koreans. In order to examine the causes of the difference in health level, the health indices and their determinants of two Koreas were analyzed in time order. As of the year 2000, the average life expectancy at birth is 71.0 years for men and 78.6 years for women in South Korea, which is longer than that of North Korea by 8.1 for men and 11.2 for women. Infant mortality rate in 1998 was 9.0 per 1,000 live births in South Korea and 54.0 in North Korea. Since being liberated from Japanese ruling in 1945, South Korea has achieved remarkable economic growth under democracy and a market economy system. On the other hand, North Korea has maintained a socialistic system. North Korea has suffered from economic crisis since the 1990s. From this point it could be said that economic status is the major factor for the differences in health level between the two Koreas. Economic status not only directly influences health level but also indirectly affects it through influences on nutrition, hygiene, health resources, and other intervening factors. The South Korean government has concentrated its limited resources on public health activities such as tuberculosis control, family planning (FP), and maternal and child health (MCH) programmes whereas the private sector has taken charge of constructing the health delivery system including health facilities and human resources. In order to solve the problem, which might occur in the private oriented medical care system, the South Korean government has introduced the national health insurance programme and enforced regulation policies. Many developing countries which are suffering from poverty and disease, can learn from the experience of Korea that had suffered from similar problems up to the early 1970s. PMID- 12109251 TI - Teleprimary care in Malaysia: a tool for teleconsultation and distance learning in health care. AB - Malaysia enjoys a comprehensive range of health services, the government being committed to the principles of universal access to high quality health care, which the Ministry of Health provides through a wide variety of nation wide network of clinics and hospitals. One of the major problems is the availability of comprehensiveness and quality of health care in remote health centres. When patients are transferred from the health centres to the hospitals for further treatment, this not only incurs inconvenience to the patients and their family but also increases the cost to the health care system. Teleprimary care is one of the tools to overcome this problem. The doctors in the remote clinics are able to discuss the problem cases through teleconsultation with the doctors and specialist in the hospitals using an audiovisual system to provide better care in the health centers without transferring the patients to the hospitals. Only the essential and needy patients are referred to the hospitals. This has not only reduced the number of patients referred to the hospitals but it has reduced the cost to the health care system. It has also provided a more comprehensive care to the patients in the health centres. The doctors in the health centers are also provided training and are also updated on the latest in medicine. This method of training has made doctors in the health centers more efficient and satisfied. PMID- 12109252 TI - A Web-based interactive education programme on vaccination for public health officials. AB - Health and Welfare Training Center of Korean National Institute of Health (KNIH) has developed various education curricula for officials who are involved in the public health or welfare sectors. Still almost all education programmes are off line based. In off-line settings, both the lecturers and the students should come to NIH from their counties to join the education programmes, and it is impossible for the students to review or to re-practice the education contents. We developed a prototype on-line setting education programmes to provide a more convenient and more effective service to the officials. This web-based interactive education programme consisted of the specific objectives on vaccination for the public health officials who were working for the national immunization programmes. It became a prototype of web-based education and was evaluated by the participants in April, 2001. From the evaluation, we improved the contents and tried to develop more subjects for the officials' training. PMID- 12109253 TI - Challenges and strategies for improving public health in countries undergoing rapid socioeconomic transition--lessons from Beijing and China. PMID- 12109254 TI - Influence of women's knowledge on maternal health care utilization in southern Laos. AB - This study aims to clarify women's knowledge on obstetric care and to analyse the situation of maternal health care (MHC) utilization in southern Laos. Face to face structured questionnaire and focus group discussions were carried out for 205 mothers aged 29.6 +/- 6.7 who had children under the age of five. Three measurements of MHC utilization comprised: 1) antenatal care (ANC), 2) tetanus toxoid (TT) inoculation, and 3) attendance delivery by health professional. Women's knowledge on obstetric care was scored. Women's knowledge was positively correlated with ANC and TT inoculation. It was found that the women with strong superstitious belief were less likely to utilize all three types of MHC than the others. Accessibility to health care facilities strongly affected ANC and attendance delivery. Enhancing women's knowledge on obstetric care and reducing barriers related with sociocultural beliefs are essential to improving maternal health in southern Laos. PMID- 12109255 TI - Health beliefs and Pap smears among Thai women in Brisbane, Australia. AB - Although a number of studies have assessed the use of Pap smear among Thai women in Thailand, little is known about factors influencing the use of this cervical cancer screening among potentially high risk Thai migrant women. We related health belief model (HBM) factors and sociodemographic variables to the use of Pap smears among migrant Thai women in Brisbane, Australia. A cross-sectional study was conducted in Brisbane, Queensland, Australia. A snowball sampling method was used to recruit 145 women. Thirty-nine percent reported regular Pap smears. Summary HBM index and self-efficacy index were positively associated with Pap smears. Barriers to screening were negatively associated. The HBM appears to be a useful framework for planning cervical cancer prevention. Strategies that reduce barriers to the screening and increase the confidence of women and their self-efficacy are likely to increase their participation. PMID- 12109256 TI - Aerosol particle and organic vapor concentrations at industrial work sites in Malaysia. AB - The objective of this study was to establish baseline data about air pollutants potentially related to nasopharyngeal carcinoma (NPC) in the Federal Territory and Selangor, Malaysia. During 1991-1993, ambient air quality was monitored at 42 work sites representing ten industrial sectors: adhesive manufacturing, foundries, latex processing, metalworking, plywood/veneer milling, ricemilling, rubber tire manufacturing, sawmilling, shoemaking, and textile related industries. At each work site, aerosol particle size distributions and concentrations of formaldehyde, benzene, toluene, isopropyl alcohol, and furfural were measured. Mean aerosol particle concentrations ranged from 61 micrograms/m3 in foundries to 5,578 micrograms/m3 in ricemills, with five industries (adhesives, metalworking, ricemilling, sawmilling, and shoemaking) exceeding the US EPA 24-hr ambient air standard for PM-10. Formaldehyde concentrations exceeded the threshold limit value (TLV) in adhesives factories. Other vapours and elements measured were well below TLVs. PMID- 12109257 TI - The role of AIDS volunteers in developing community-based care for people with AIDS in Thailand. AB - The present study analyses the effectiveness of AIDS volunteers in mitigating the stigma attached to People With AIDS (PWAs) within the context of developing community-based care (CBC) in Thailand. A total of 86 trained village health volunteers (T-VHVs) and 99 non-trained village health volunteers (N-VHVs) were enrolled in the study. In addition, 58 villagers in the T-VHV's intervention area and 72 villagers in the non-intervention area were also enrolled. Both T-VHVs and N-VHVs as well as villagers were assessed to determine their level of knowledge with respect to HIV/AIDS and attitudes toward PWAs. Furthermore, we also determined the village health volunteers' level of activity in distributing knowledge of HIV/AIDS in order to prevent and reduce stigma in the community. Although T-VHVs showed a greater depth of knowledge of HIV/AIDS than N-VHVs (p < 0.05), positive attitudes toward PWAs and the level of practice of village health volunteers did not differ significantly between T-VHVs and N-VHVs. While the level of health knowledge of villagers did not differ significantly between the T VHV's intervention and control areas, a significant difference was observed between the two areas in terms of the villagers' attitudes towards PWAs (p < 0.01). Villagers in the intervention area attached less stigma to PWAs; therefore, T-VHVs played a role in providing basic information on AIDS to the villagers and in mitigating the stigma attached to PWAs. However, these volunteers need to undergo further training through a well-organized training programme in order to obtain a greater depth of knowledge. This is essential for the development of community-based care for PWAs. PMID- 12109258 TI - Sero-epidemiology and risk factors of positive hepatitis B surface antigen amongst Chinese adolescents. AB - This paper reports the prevalence rate of hepatitis B antigen (HBsAg) amongst the Hong Kong Chinese adolescents (age 11 to 19), and the risk factors associated with HBsAg positive. The study is cross sectional and 1,580 students were randomly selected from 12 secondary schools in four regions of Hong Kong. For those subjects who agreed to participate and were randomly selected, their blood was tested for HBsAg and anti-HBs. The overall prevalence of HBsAg positive was reported to be 5.8% (7.9% in male and 4.1% in female), lower than 8.1% in 1978. Males, those born in Mainland China and family history of carriers had higher prevalence of HBsAg positive (7.9% vs 4.1%, 12.2% vs 4.7%, 52.9% vs 3.8% respectively) with statistical significance. Males and those born in mainland China were found to have significantly higher odds ratio 1.8 (95% CI. 0.98-3.52) and 4.4 (95% CI. 2.2-8.8) respectively of HBsAg positive by multivariate analysis. Findings suggest that family history of carriers and those born in endemic area are at a higher risk. Therefore it is worthwhile to consider vaccination programme for adolescents to reduce the carrier rate, and to also reduce the injection amongst the adults by horizontal transmission. PMID- 12109259 TI - Measurement of thyroid volume in children using a portable ultrasound machine: a technical note. AB - The data on thyroid volume measurements that determines prevalence of goitre in children is very important for public health consideration as the presence of goitre in children effectively reflects the status of iodine deficiency disorders (IDD) in the general population. Ultrasound is an excellent modality to evaluate thyroid size. Local experience in using a portable ultrasound machine to measure thyroid volume is presented. The thyroid anatomy and techniques of ultrasound assessment are highlighted. Proper training of public health doctors to perform thyroid gland ultrasonography is crucial to ensure that the thyroid volume data collected would be more accurate and reliable for the planning of health programmes to eliminate IDD in the particular areas. PMID- 12109260 TI - Assessing quality of life for adolescents in western Australia. AB - This study investigates the quality of life (QOL) for adolescents residing in Perth, Western Australia. The Quality of Life Profile-Adolescent Version (QOLPAV), a generic self-reported questionnaire, was administered to 363 adolescents aged between ten and 18 years who were enrolled in 20 high schools within metropolitan Perth. Stepwise regression analysis of the data showed that age, control, opportunity and perceptions of health have significant associations with QOL. It was also found that adolescents with a chronic condition and those without have similar QOL scores. This suggests that, in terms of QOL, the chronically ill adolescents do not view themselves as different from their healthy counterparts. PMID- 12109261 TI - Oral health condition and endemic goitre, in an iodine-deficient area in Bali, Indonesia. AB - Iodine deficiency is a major determinant of endemic goiter which affects bone growth. The purpose of this study was to clarify the relationship between iodine deficiency and oral health. A cross-sectional survey was carried out on 181 inhabitants aged 17 to 40 years in two villages in an iodine-deficient area in Bali, Indonesia. Thyroid swelling, serum level of thyroid-stimulating hormone, DMFT index, plaque index, gingival index, dietary habits and socioeconomic status were examined. Thirty two subjects (21%) had thyroid swelling considered to be endemic goiter, with the highest prevalence in females aged 21 to 30 years (40 41%). The DMFT, food habits, and socioeconomic status were different between both villages. Multiple regression analysis adjusted for age, sex, and location (village) did not detect any association between DMFT and serum thyroid stimulating hormone level or thyroid swelling. Further studies will be required to confirm these findings. PMID- 12109262 TI - Primary care research--a blueprint for action for Singapore. AB - Primary care research, hitherto overshadowed by research in the tertiary institutions, lacks impetus due to image, limitations of resources and poor infrastructure. This article looks into the current state of research development and the barriers. These hurdles can be overcome by a paradigm shift through the initiation of regular research meetings, training, infrastructure support and a collaborative research network to optimize time and resources. This will open the door to a wide field of primary care research, from basic, clinical research to health service and health system analysis and evaluation. Family physicians can tap on this opportunity to sharpen their critical appraisal skills through research, which can form an integral part of their continuous professional development. A network of pro-research family physicians will foster a vibrant research culture and pave the way to a renaissance of primary care research. PMID- 12109263 TI - Evaluation of the Seventh Malaysian Health Plan: a new approach. AB - Since the First Malaysia Plan (1966-1970) many reviews have been done on the five year health plans of the Ministry of Health (MOH). These included the Mid-Term Reviews and the review done at the end of the respective five-year plan period. There was no structured evaluation method carried out until the Seventh Malaysia Plan (7MP) period (1996-2000), among others because of the complexity of the MOH health plans. The evaluation of the 7MP was the first one conducted using a better-structured process. The findings and recommendations of the evaluation were used and incorporated in the subsequent 5-year health plan, under the Eighth Malaysia Plan (2001-2005). PMID- 12109264 TI - Should community-managed drug stores be phased out? AB - The existence of community-managed drug stores (CDS), operated by trained village health workers under the supervision of health officials, is under threat in the Philippines. This is to determine if there is an association between households' access to medicines and three types of CDS, namely, Botika Binhi (BB), Family Health Management By and For Urban Poor Settlers Pharmacy (FAMUSCY) and Botika ng Barangay (BnB) in selected urban and rural communities in a province where all three types operate. The study design was case-control using structured interviews and observations. A total of 1,710 households or 90% of the total were interviewed, and the medicines at home were inspected. Indicators for the five dimensions of access are affordability, availability, A1 quality, acceptability and appropriateness. The village with the longest-running CDS for each type was selected. RESULT: There is an association between the presence of any of the three CDS and villagers' perception of affordability of medicines, between FAMUSCY or BnB and availability of unbranded medicines at home as well as perception of counselling provided by prescribers and dispensers to be more than expected. On the contrary, there is an inverse association between the presence of FAMUSCY and availability of herbal plants at home, between BnB and percent of correct uses of medicines at home, and between either FAMUSCY or BnB and perceptions of availability of medicines and quality of services by dispensers at patronized drug outlets. CONCLUSION: Phasing out CDS should be weighed against the benefit of making medicines affordable at the village level, promoting the use of unbranded medicines and herbal plants with recognised therapeutic value, and keeping the clients informed through drug counselling provided by prescribers and dispensers. PMID- 12109265 TI - Rotating spiral waves in fertilized ascidian eggs. AB - Excitable systems modelled by reaction-diffusion equation may be expected to produce quite complex spatial patterns. Winfree [1974] demonstrated experimentally, in the Belousov-Zhabotinskii reaction, the existence of particular waves called rotating spiral waves. Later Keener and Tyson [1986] presented a thorough analysis of these waves in excitable systems. Spiral waves can also be observed in brain tissue (Shibata and Bures [1974]), while it seems that the precursor to cardiac fibrillation is the appearance of rotating waves of electrical impulses (Winfree [1983]). In this work we suppose the appearance of Ca++ spiral waves in the vegetal pole of ascidian egg cells after the first ooplasmic segregation. Previously we observed that (Ballaro and Reas [2000a]), when the myoplasm is completely localized in the vegetal region (excitable stage) and the ascidian egg cell is perturbed by an increase of Ca++ concentration in the culture medium, the cell reacts by showing persistent mechanical waves of contraction which exist as long as the cell is perturbed. Experimentally we observed the production of a polar lobe located in the vegetal region and the change of the inclination of mitotic furrow, after the appearance of a myoplasmic spiral wave in the vegetal pole. So we suppose that the myoplasmic spiral wave is due to a Ca++ spiral wave, and the myoplasmic spiral wave then causes the changes in the shape of the cell (polar lobe, inclination of mitotic furrow, etc.). Moreover we give a simple geometrical description of a spiral wave. PMID- 12109266 TI - Theses on Darwin. AB - The received view that teleology has been successfully eliminated from the modern scientific worldview is challenged. It is argued that both the theory of natural selection and molecular biology presuppose the existence of natural teleology, and so cannot explain it. A number of other issues in the foundations of biology are briefly examined, while stress is laid throughout on empirical evidence of the rational agency inherent in life. It is urged that teleology be rehabilitated and that the reigning functionalist philosophy be replaced by a realistic view of biological functions as emergent properties of living matter within a broad, selforganization framework. PMID- 12109268 TI - [The cosmos in art, architecture, and ecology in the new millennium]. PMID- 12109267 TI - Conditions for a theoretical biology: is biology truly an autonomous science? AB - In a line of a previous paper, the conditions for a theoretical biology were discussed and it was pointed out that the primary condition is that biology is an autonomous science. This statement is connected to the problem of reductionism. A discussion of the autonomy of biology shows that reductionism cannot be maintained, although particularly in physiology often physics and mathematics are used. Development, organization and evolution of biological systems are typical areas of autonomous biological researches and of autonomous theoretical developments. A sort of reduction to history seems today a nonsensical attempt to reduce the area of free theoretical biological activity. PMID- 12109269 TI - Towards a morphogenetic perspective on cancer. AB - The purpose of this paper is to present a critique of the current view that reduces cancer to a cellular problem caused by specific gene mutations and to propose, instead, that such a problem might become more intelligible, if it is understood as a phenomenon which results from the breakdown of the morphological plan or Gestalt of the organism. Such an organism, in Aristotelian terms, is characterised for presenting a specific morpho or logos (form) and for having a telos (end) to fulfill. A malignant tumour represents an entity separated from both, the organic logos and the organic telos. According to the basic postulates of Semiophysics--a blend of Aristotelian physics and Catastrophe Theory developed by Rene Thom--an organism is a source (original) form individuated by a dominant pregnancy which corresponds to its morphogenetic field. Here it is suggested that cancer in aged individuals may result from the progressive exhaustion of the developmental constraints that regulate the process of ontogeny, that is expected to go from the fertilised non-differentiated zygote to the mature fully-developed organism, because there is no further point ahead in the developmental pathway past the reproductive age. Cancer in young individuals (before their reproductive maturity) may then be consequence of premature derangement of such fundamental developmental constraints. In all cases the result is the loss of morphological coherence within the organism. Thus representing a conflict between an organised morphology (the organism) and a part of such a morphology that drifts towards an amorphous state (the tumour). PMID- 12109270 TI - "Darwin is a prime number". PMID- 12109271 TI - Geophysiological modeling: new ideas on modeling the evolution of ecosystems. AB - Living organisms evolve within ecological associations (from ecosystems to the biosphere) that are constituted by a biological component and a physico-chemical component. It is generally supposed that interactions such as competition and predation between the biological components of ecosystems are the main cause for the observed organization of ecosystems. We believe that in the search for a more comprehensive theory of evolution a much greater attention should be paid to the ways in which living organisms interact with the physico-chemical environment. To test our ideas, we develop a mathematical model to study the evolution of ecosystems, and we apply it to the study of hydrothermal vents. The model proposed is still a qualitative model. It tries to study, in a first approximation, the behaviours of the biological and chemical components. In the following we hope to develop and to improve it so to give a more realistic model. PMID- 12109272 TI - The concept of species and the foundations of biology, a case study: the Callithrix jacchus group (Primates-Platyrrhini). AB - The axiomatic theory presented in Galleni and Forti [1999], being part of the foundational programme of Ennio De Giorgi, is based on the fundamental notions of quality, relation, operation and collection, and provides a very general axiomatization of the biological notions of living object, generation, species and speciation. Within this theoretical framework we consider here a difficult case of classification of species: the Callithrix jacchus group of the New World monkeys. Although the morphological analysis strongly suggests the individuation of six different species, nevertheless several experiments of crossing give evidence to fertility of hybrids. Since both the morphological and the hybridological criteria have shown to be of enormous importance in actual classification of species, this apparent contradiction seems very disappointing. Our axiomatization of speciation processes as operations acting in special time intervals permits to avoid the contradiction by allowing for individuals which, during such special periods, may belong to more than one species. Therefore we assume that one or more speciation processes are developing, starting from a unique protospecies and differentiating in six new ones. In order to obtain more evidence of these processes, new observations and suitable experiments are needed. PMID- 12109273 TI - [What is your ultrasound diagnosis? Medullary nephrocalcinosis with proximal ureteral calculus and moderate hydronephrosis left]. PMID- 12109274 TI - [Psychosomatic research in primary health care--ideal concept and reality]. AB - The cooperation between research institutions and primary care doctors can be seen as a possible model for practical research in psychosomatic medicine. Cooperation experience with a study in the field of sexual medicine are reported as well as recommendations regarding the planning and implementation of similar studies are given. 7455 Swiss physicians specialized in psychiatry, internal or general medicine were asked to fill out a survey on "sexual dysfunction and anti depressants". They were also asked if they were willing to interview their patients on the same issue. 1100 physicians agreed to answer the doctors survey and 62 were interested in interviewing their patients. However, only 12 physicians actually did. In 55 telephone interviews doctors mentioned as their main reasons for not participating: lack of time and their hesitation to discuss sexual issues with depressive patients. CONCLUSION: The collaboration in research projects means a considerable effort on the side of primary care doctors. Therefore, their cooperation should be rewarded appropriately, e.g. time could be counted as further training. PMID- 12109275 TI - [Accident-induced tooth avulsion--consequences for dental pulp and periodontium]. PMID- 12109276 TI - [Hypophyseal incidentaloma in a patient with autosomal dominant polycystic kidney disease]. AB - The prevalence of incidentally discovered lesions within the pituitary (pituitary incidentalomas) is about 10%. The most common form of sellar mass are clinically nonfunctioning adenomas (less than 10 mm); functioning adenomas, however, are rare. Incidentally discovered pituitary microadenomas causing growth hormone hypersecretion are uncommon. In addition, the association of autosomal dominant polycystic kidney disease with acromegaly is exceptional and has not yet been reported to our knowledge. PMID- 12109277 TI - [Exposure to open pulmonary tuberculosis. 30-year-old asymptomatic patient]. PMID- 12109278 TI - [Uncertain neurological status in a depressed patient]. PMID- 12109279 TI - [Skin manifestations of diabetes mellitus]. AB - Cutaneous manifestations of diabetes mellitus are common and manifold. In the course of their disease, 30-70% of type I and type II diabetics are affected. At a closer look, the majority of all diabetics would show preclinical alterations of the vessel walls or of the connective tissue. In the diabetic patient, it is important to be familiar with and to look for the possible cutaneous alterations, which have to be diagnosed on mainly clinical grounds. Some of them can be a consequence or an accompanying symptom of a poorly controlled diabetes and therefore they can potentially help the responsible physician to prevent more serious complications. Some of these dermatoses are so called marker lesions and can precede the manifestation of the diabetes by years. The exact pathogenesis of most of these dermatoses is unknown. It is reasonable to assume though that vessel and connective tissue alterations as well as the impairment of the immune system and other associated metabolic changes caused by the diabetes play an important role. PMID- 12109280 TI - [Skin and hair, marker organs for thyroid diseases]. AB - Skin and cutaneous appendages are target organs for thyroid hormones. Thus, a variety of changes of skin, hair and nails occurs in association with thyroid diseases. Most of these cutaneous changes are unspecific, but in their entirety they may nontheless be indicative of thyroid diseases. In this review the specific cutaneous lesions as thyroglossal duct cysts and cutaneous metastases of thyroid carcinoma, the unspecific cutaneous changes of hypo- respectively hyperthyroidism, the typical skin lesions of Grave's disease, and selected skin diseases and syndromes associated with thyroid abnormalities are discussed. PMID- 12109281 TI - [Skin manifestations of diseases of the gastrointestinal tract]. AB - Diseases of the gut frequently show skin symptoms. These can give first and important clues in regard to diagnosis. In general the etiology can be divided into genetic disorders, chronic inflammation, drug reaction, infectious diseases or related to malignancy. In genetic disorders increasing knowledge about the involved genes is available, allowing prenatal diagnosis and screening of clinically not affected family members. Especially in cancer prone syndromes early diagnosis and preventive treatment is crucial. Inflammatory bowel diseases show a high prevalence, therefore necessitating the knowledge of skin complications such as pyoderma gangrenosum, Sweet syndrome and erythema nodosum. Gastrointestinal malignancies may metastasize into the skin or may produce typical paraneoplastic changes. PMID- 12109282 TI - [Skin changes in liver diseases]. AB - The liver is the second largest organ in the human body. An association between the skin and the liver has been recognized for centuries. Indeed, there is not always a correlation between the severity of the liver disease and the skin changes and they often are not specific. Several types of interaction between the skin and the liver are encountered: 1. Liver disease may cause skin changes. 2. The skin and the liver may be involved by the same pathologic process. 3. Skin disease may cause-liver abnormalities. 4. The liver may be damaged by drugs used to treat skin disease. In the following, the first two conditions are discussed. PMID- 12109283 TI - [Experimental and clinical rationale for complex treatment of mental disorders in clean-up workers of the Chernobyl nuclear plant accident]. AB - Piracetam is an effective drug for the treatment of asthenic and intellectual mnestic disorders. However, this drug induces superfluous stimulation which reduces the therapeutic effect in rescuers after Chernobyl nuclear power disaster. The results of experimental and clinical investigations show that the administration of gidazepam--an atypical benzodiazepine tranquilizer--in combination with piracetam produces correction of the undesired stimulant action of the latter drug, while potentiating the nootropic (antiamnesic) and anxiolytic effects. This drug interaction led to an improvement in the therapy. PMID- 12109284 TI - [Comparative study of dilceren (nimodipine) on the cerebral circulation in the intact animals and animals subjected to ischemic brain damage]. AB - The influence of the calcium entry blocker dilceren (nimodipine) on the local and regional circulation was studied in rats and cats. In the majority of experiments on intact animals, dilceren increased the local and regional blood flow. However, in some cases, the circulation decreased or remained unchanged, which was accompanied by pronounced hypotension. Administered after global brain ischemia, dilceren improved the cerebral hemodynamics in all experiments. PMID- 12109285 TI - [Effect of Cerebral on the immune system in experimental hemorrhagic stroke]. AB - Data on the development of three experimental models of hemorrhagic stroke in mice are reported. The models differ in the extent of damage. From the standpoint of the immunopharmacological investigation, most adequate model is provided by the light acute brain circulation disorder (LABCD). In the LABCD model, the hemorrhagic stroke is characterized by reduction in the thymus weight, inhibition of the antibody (hemolysin) synthesis, and enhancement of the delayed type hypersensitivity response. The new antistroke drug cerebral decreased (especially after intranasal administration) the level of lethality in experimental animals and improved the immunological indices. PMID- 12109286 TI - [Comparative study of the anti-arrhythmic activity of cardiocyclide and nibentan in hypersympathotonia and acute myocardial ischemia]. AB - The antiarrhythmogenic and antifibrillar action of cardiocyclide and nibentan- class III antiarrhythmogens (according to the classification of Vaughan Williams) was experimentally studied on narcotized cats. The hypersympathotonia model was induced by a 30-sec stimulation of the left stellate cervical ganglion on the background of acute myocardial infarction. On this model of a ventricular disorder in the heart rhythm, cardiocyclide (2.4 mg/kg) and nibentan (0.2 mg/kg) exhibited pronounced antiarrhythmic activity. It is suggested that both drugs can be used under clinical conditions for the prophylaxis of sudden coronary death in patents with acute myocardial infarction. PMID- 12109287 TI - [Effect of mexidol, emoxypine, and dimephosphon on electrophysiological effects of nibentan]. AB - Mexidol, emoxypine, and dimephosphon decrease the acute toxicity of nibentan in mice. In the experiments on cats, these drugs reduced the negative dromotropic action of nibentan and prevented an increase in the effective refractory period of the atrioventricular node and ventricles, in the QT interval length, and in the ventricular excitation threshold. PMID- 12109288 TI - [Effect of the novel dipeptide nootropic agent noopept and its metabolite cyclo-L prolylglycine on the transcallosal evoked potential in the rat brain]. AB - The effect of new nootropic dipeptides--noopept (N-phenylacetyl-L-prolylglycine, GVS-111) and its metabolite (cyclo-L-prolylglycine)--and a standard nootrope piracetam on the transcallosal evoked potential (TEP) in rat brain was studied. In the dose range from 150 to 300 mg/kg, piracetam increased the TEP amplitude, which exhibited a maximum after 1.5-2 h and then gradually decreased. Both noopept and cyclo-L-prolylglycine also increased the TEP amplitude, which attained a plateau and retained this level over the entire observation time (above 3.5 h). All the nootropes studied increased both components of the evoked potential. Piracetam and cyclo-L-prolylglycine led to an approximately equal increase in both waves, while noopept induced a somewhat greater increase in the negative TEP wave amplitude. It is suggested that the positive effect of noopept and cyclo-L-prolylglycine upon the interhemispheric signal transfer (indicated by the improved transcallosal response) can be considered as a potential neurophysiological basis for a positive drug influence on the behavioral level. PMID- 12109289 TI - [Hepatoprotective properties of liproxol]. AB - The experiments on rats with a model toxic liver damage (tetrachloromethane hepatitis) showed evidence of a high hepatoprotector activity of liproxol, representing a combination of eplir and lokhein mixed in a 1-12 ratio. Liproxol inhibits lipid peroxidation, stimulates the excretory and detoxicant functions of liver, reduces hyperfermentation, and normalized membrane phospholipid composition. PMID- 12109291 TI - [Effect of sulodexide on hemocoagulation and lipid peroxidation in athletes during physical load]. AB - High physical loads lead to an increase in the blood coagulation rate and inhibit fibrinolysis in athletes. At the same time, the fibrin-stabilizing factor activity and the fibrinogen concentration tend to increase. Sulodexide produces an anticoagulant effect, thus preventing from hypercoagulation. An increase in the working capacity of athletes caused by suloxide administration is accompanied by the antioxidant action. PMID- 12109290 TI - [Multicomponent antithrombotic effect of the neuroprotective prolyl dipeptide GVS 111 and its major metabolite cyclo-L-prolylglycine]. AB - The experiments in vivo showed that the new nootropic prolyl-containing GVS-111 produces an antithrombotic effect, influencing various stages of the blood coagulation process. GVS-111 exhibits anticoagulant and fibrinolytic properties and enhances fibrin destabilization by reducing the XIIIa factor activity. These effects are manifested upon both intraperitoneal (1 mg/kg) and peroral (10 mg/kg) administration of GVS-111 (in both cases, a single daily treatment over a period of 10 days). The same effects (anticoagulant, fibrinolytic, antifibrin stabilizing) were observed in in vitro experiments with both GVS-111 (10(-3)-10( 6) M) and its main metabolite cyclo-L-prolylglycine (up to 10(-10) M). In addition, the latter metabolite exhibited an antiaggregant effect. The antithrombotic activity of GVS-111, together with previously established neuroprotector properties, low toxicity, and the absence of complications, makes this compound a promising antistroke drug. PMID- 12109292 TI - [Effect of dimephosphon and xidifon on parameters of lipid peroxidation and the antioxidant system in rats subjected to the long-term administration of prednisolone]. AB - Experiments in rats with an osteoporosis model induced by the chronic administration of prednisolone (50 mg/kg, p.o., over 14 days) showed that the drug differently affects the lipid peroxidation (LPO) rate and the antioxidant system activity. Xydiphone (45 mg/kg, p.o.) administered over 14 days exhibited a perooxidant action. Dimephosphon (208 mg/kg, p.o.) administered over the same period of time produced an antioxidant effect as manifested by a growth in the catalase activity, an increase in the content of ceruloplasmin and glutathione, and a decrease in the content of LPO products in the blood serum and liver tissues. The results of combined administration of prednisolone and phosphonates indicate good prospects of the dimephosphon treatment as a means of normalization of the LPO and antioxidant system functioning disturbed by prolonged use of glucocorticoids. PMID- 12109294 TI - [New approaches to prognosis of risk of development of gastropathies induced by nonsteroid anti-inflammatory agents]. AB - A pharmacological test with indomethacin is suggested for predicting the risk of NSAID-induced gastropathy. It is shown that patients with diagnoses of rheumatoid arthritis and osteoarthrosis can be divided into two groups with respect to the indomethacin test--"vulnerable" and "resistant", characterized by a high and low risk of NAIDS-induced gastropathy development, respectively. The proposed index of resistance to this disorder, together with the glutathione redox buffer test index, can be used as reliable criteria for predicting the NSAID-induced gastropathy. PMID- 12109293 TI - [Pharmacological efficacy of delta sleep-inducing peptide in experimental acute pancreatitis]. AB - The development of experimental acute pancreatitis (EOP) in rats is accompanied by (i) intensification of the lipid peroxidation (LPO) process and the accumulation of malonic dialdehyde (MDA, an LPO product) in the tissues of pancreas, liver, and kidney and in the blood serum, (ii) destabilization of membranes and reduction of the osmotic resistance of erythrocytes (ORE), (iii) increase in the concentration of extracorpuscular hemoglobin (ECH) and medium molecular-weight molecules (MWM) in the blood serum, and intensification of protein autolysis in tissues. Preliminary triple intraperitoneal administration of a delta-sleep-inducing peptide (DSIP) in a dose of 12 micrograms/100 g body weight to the test rats with EOP stabilized LPO, improved the erythrocyte membrane structure, reduced the MDA level in tissues and blood serum, increased ORE, reduced the ECH and MWM level in the blood, and decreased the protein autolysis rate in tissues. PMID- 12109296 TI - [Dependence of galodif pharmacokinetics on duration of its administration]. AB - Pharmacokinetics of the new antiepileptic drug galodif was experimentally studied on rats. A single administration of the anticonvulsant is followed by slow elimination. A five-day administration leads to a significant acceleration of the drug elimination due to activation of the cytochrome P-450 dependent monooxygenase system of liver. After a 14-day administration, the elimination rate decreases as compared to that for the five-day treatment, while being still higher than in the case of single administration. PMID- 12109295 TI - [Anti-inflammatory properties of noopept (dipeptide nootropic agent GVS-111)]. AB - It is established that single intravenous (0.5 and 5 mg/kg, p.o.) or single peroral (10, 50, 100 mg/kg) and prolonged peroral (5 mg/kg, over 10 days) administration of noopept produces a dose-dependent inhibition of the model inflammatory response to concanavaline A in CBA mice. Intravenously injected (5 mg/kg) noopept suppressed the acute nonimmune carrageenan-induced foot inflammation in rats by 62.2% within 3 h. The most pronounced antiinflammatory effect of dipeptide was observed on the model of adjuvant arthritis in rats, where the drug administered over 25 days in a daily dose of 0.5 mg/kg (i.m.) or 5 mg/kg (p.o.) significantly reduced the chronic immune inflammation (on the 12th day, by 94.0 and 74.1%, respectively). The in vitro experiments with neutrophilic leukocytes of F1(CBA.C57BL/6) mice treated with noopept in a single dose of 5 mg/kg (i.v.) showed a 5- to 6-fold suppression of the hemiluminescence stimulated by opsoinized zymosan or phorbolmyristate acetate. It is suggested that the antiinflammatory activity of noopept is probably related to its antioxidant properties. PMID- 12109297 TI - [Efficacy of a preparation based on Lactobacillus acidophilus for protection from myelotoxic effects of ionizing radiation and antineoplastic cytostatics]. AB - The ability of a Lactobacillus acidophilius (LA) based preparation to prevent from a myelotoxic damage was studied on (CBA.C57BL/6)F1 mice upon total gamma irradiation or single intraperitoneal injection of cyclophosphan. The antitumor and antimetastatic activity of LA-based preparation was studied in experiments with C57BL/6 mice bearing Lewis lung carcinoma or melanoma B16. The preparation obtained from heat-killed Lactobacillus acidophilius species provides protection of hemopoietic stem cells, as evidenced by the amount of endogenous CFUs increased 3.4-6.7 times. The LA-based preparation favored accelerated overcoming the radiation- and cyclophosphan-induced leukopenia. At the same time, the drug did not affect the proliferative activity and apoptosis of tumor cells, their growth rate, and metastatic activity. Nor did it reduce the efficacy of the experimental tumor radiotherapy. PMID- 12109298 TI - [Effect of carbamazepine on the structure-function relations in the development of the epileptic structure]. AB - The experiments on white outbred male rats with cobalt-induced epileptogenic focus in the left sensomotor cortical region showed that the anticonvulsant effect of carbamazepine (20 mg/kg) depends on the functional state of the epileptic system (ES). In various stages of the ES development, the drug effect is related to the influence upon the determinant focus generating the epileptic activity. In the initial stage, carbamazepine primarily affects epileptic activity of the cortical foci (ipsi- and contralateral hemisphere). In the second (generalization) stage, the drug decreases the epileptic activity in all structures, but to a greater extent, in the dorsal hippocampus and lateral hypothalamus. PMID- 12109299 TI - [Efficacy of galstena in liver damage induced by antitubercular agents]. AB - It is established that galstena possesses hepatoprotector activity and is capable of reducing pathological changes in animals treated with toxic doses of antituberculous drugs. Galstena prevents from the development of cytolysis, as evidence by inhibition of the activity of aspartate aminotransferase, alanine aminotransferase, and lactate dehydrogenase. The anti cholestatic effect of galstena is confirmed by suppression of the growth in the levels of total bilirubin, alkaline phosphatase, and gamma-glutarate transpeptidase. Galstena was also found to possess antiinflammatory properties. Moreover, a growth in the activity of glutathione-dependent reductase (related to inhibited growth of the iron level in the blood serum) is evidence of the antioxidant activity. In addition, galstena prevents from an increase in the content of creatinine and urea, which is evidenced of decreasing endogenous intoxication. PMID- 12109300 TI - [The novel immunodepressant rapamune]. PMID- 12109301 TI - [Opioid receptors--current state and prospects]. PMID- 12109302 TI - [Licopin--prospects for clinical use]. PMID- 12109303 TI - [Neurotropic activity of lambertian acid and its amino derivatives]. AB - The neurotropic activity of the originally synthesized methyl lambertianate (II), as well as lambertianic acid (I) and three amino derivatives (III-V) was studied. All compounds exhibit equally low toxicity, but differ in the type of influence upon CNS. The most pronounced action was observed for esters II and V. Compound II exhibited a strong antidepressant effect with stimulating action. Introduction of the amino group led to an opposite tendency: compound V exhibited an antipsychotic and sedative (calming) effect upon CNS, without any anticonvulsant action. PMID- 12109304 TI - [Signs of miliary tuberculosis: which diagnostic tests and when to treat?]. AB - Three patients, men aged 21, 57 and 53 years, presented with variable non specific symptoms such as general malaise, weight loss, elevated temperature, abdominal pain, cough, pulmonary crepitations and elevated liver enzymes. Diffuse fine nodular infiltration was seen on chest radiography in the last two cases. The first patient refused to be tentatively treated with tuberculostatics and died. Mycobacterium tuberculosis complex grew on Lowenstein medium a week later. The two other patients received tuberculostatic treatment. The second patient recovered, while the third patient suffered a cerebrovascular accident on top of emaciation and respiratory insufficiency and died. In the Netherlands, currently more than one hundred patients with tuberculosis disease die each year. The disease is mostly seen in people from the high-risk groups for tuberculosis such as asylum seekers and immigrants. Even after extensive diagnostic procedures it can be difficult to obtain rapid bacteriological confirmation. When miliary tuberculosis is suspected it is important to carry out the complete range of tests (Ziehl Neelsen microscopy, PCR, Lowenstein cultivation) and to start therapy immediately and not to await the results of the diagnostic tests. However, in many cases this may still be too late, with an estimated mortality of 20%. PMID- 12109306 TI - [Cardiovascular risk factors. I. Review of possible causes of heart and vascular diseases]. AB - Despite the decline in the past 30 years in age-adjusted mortality, cardiovascular disease is still the most important cause of morbidity and mortality in the Western world and increasingly so in the non-Western world. The decreased mortality rate is attributed to increased knowledge of the risk factors, a concomitant healthier life style and improved treatment for the risk factors. It has been clearly demonstrated that increased serum levels of cholesterol, increased blood pressure, the presence of diabetes, physical inactivity and smoking increase the risk of cardiovascular disease. The role of estrogens, homocysteine, thrombotic and inflammatory factors is currently the subject of considerable research. It seems likely that a certain genetic constitution enhances the susceptibility to the effects of other risk factors. New candidate genes are being suggested daily. Until now, no clearly decisive results have been achieved in determining the relations between the investigated genetic polymorphisms and the occurrence of cardiovascular disease. The consumption of foodstuffs that are rich in anti-oxidants seems to reduce the risk of cardiovascular disease, but randomised research into the effects of anti oxidants as food supplements suggests that this does not affect the development of cardiovascular disease. PMID- 12109305 TI - [The treatment of chronic inflammatory diseases with monoclonal antibodies against tumor necrosis factor: side effects, contraindications and precautions]. AB - Monoclonal antibodies are increasingly used to modulate immunologically mediated diseases such as rheumatoid arthritis, psoriatic arthritis, systemic lupus erythematosus, Crohn's disease, multiple sclerosis and systemic vasculitis. Constructs of monoclonal antibodies to tumour necrosis factor (TNF) alpha differ with respect to their structure, effects and immunogenic side effects. Clinical experience with TNF alpha-neutralizing therapy has revealed several other side effects over the past few years. The most important is increased infection rates, especially the activation of (latent) tuberculosis, although other opportunistic infections such as listeriosis, Pneumocystis carinii pneumonia, histoplasmosis, candidiasis and aspergillosis have also been reported. Furthermore, results from clinical studies indicate that TNF alpha-neutralizing therapy should not be given to patients with cardiac failure (NYHA class III or IV) or a history of demyelinating disease. An increased incidence of malignancies has not been observed up to now, but data from the long-term follow-up are not yet available. PMID- 12109307 TI - [Cardiovascular risk factors. II. Signs of vascular damage: practical implications]. AB - In recent years, an increasing amount of information has become available on new indicators of cardiovascular damage, their determinants and the value of these indicators in the prediction of the development of cardiovascular disease. These indicators include carotid intima-media thickness, ECG characteristics, endothelial function and the measurement of coronary calcium. These indicators are currently used predominantly in cardiovascular disease research. At present, only limited information is available as to whether these indicators are useful in clinical practice for estimating the absolute risk of cardiovascular disease in an individual. The relative value of these newer indicators compared to the routinely collected risk factor information such as medical history, lifestyle, anthropometric variables, blood pressure and lipids is also not yet well established. The recent Dutch guidelines for the initiation of treatment with blood pressure lowering and lipid lowering drugs as a function of the absolute risk of cardiovascular disease within 10 years for the individual patient point toward the urgent necessity for research in the area of new risk indicators of vascular damage. PMID- 12109308 TI - [Diagnostic image (93). A boy with a painful leg swelling. Osteoid osteoma]. AB - A 13-year-old boy presented with a swelling on his left lower leg and suffered from pain, which was almost immediately relieved by acetylsalicylic acid. Roentgenographic and histologic appearance demonstrated an osteoid osteoma in the tibia. PMID- 12109309 TI - [From gene to disease; from mutations in the Von Willebrand factor gene to hemorrhagic diathesis and thrombocytopenia]. AB - The broad spectrum of both clinical presentation and severity of bleeding complications observed in Von Willebrand's disease is the result of a high number of mutations. Most are clustered in specific functional domains, leading to quantitative (types 1 and 3) or qualitative (types 2: A, B, M and N or Normandy) defects in the Von Willebrand factor (VWF) protein. Unlike all the other subtypes, type 2B is the result of a 'gain of function' phenotype characterised by an enhanced binding affinity of the VWF protein to the glycoprotein (Gp) Ib complex of platelets, which causes the platelets to disappear from the circulation, resulting in thrombocytopenia. The thrombocytopenia can be triggered by stress or desmopressin, among other causes, and therefore the latter is contraindicated. Type 2B is characterised in vitro by enhanced platelet agglutination with ristocetin. All 2B mutations are located in exon 28, in a small region encoding the Gp-Ib binding site within the A1 domain of VWF. These mutations inactivate a regulatory function of the A1 domain which facilitates Gp Ib binding. PMID- 12109310 TI - [Treatment adherence amongst HIV-infected patients in Rotterdam: poorer prognosis independent of nationality in heterosexual infected patients, intravenous drug users, Negroid and Latin-American patients]. AB - OBJECTIVE: To assess adherence to antiretroviral therapy (ART) in Dutch and non Dutch HIV-infected patients. DESIGN: Observational, cross-sectional study. METHODS: Consecutive HIV-infected patients taking ART and visiting the internal medicine outpatients' clinic at the Rotterdam Dijkzigt University Hospital between 1 February until 30 April 2001 were interviewed by a multilingual interviewer using a standard questionnaire. Classification of adherence was based on the interview data. Multivariate analysis was used to determine independent predictors of adherence. Nationality was defined as 'Dutch' if the person was born in the Netherlands, and otherwise as 'non-Dutch'. RESULTS: The 203 patients included in this study comprised 131 men and 69 women with an average age of 42 years. There were no data available on treatment adherence for 3 of the patients. Of the 81 Dutch patients, 60 (74%) adhered to the treatment compared with 68 (57%) of the 119 non-Dutch patients. However, after correction for sex, risk group, race and duration of treatment, there was no difference in treatment adherence between these two groups (OR: 0.6; 95% CI: 0.2-1.9). Failure to adhere to treatment was seen most frequently in the 109 heterosexually infected patients (OR: 2.6; 0.98-6.7), the 22 intravenous drug users (OR: 3.3; 1.04-10.1), as well as in the group of Negroid patients (OR: 3.5; 1.1-11.3) and Latin-American patients (OR: 8.5; 1.7-42.7). PMID- 12109311 TI - [Intermittent thrombocytopenia as a manifestation of Von Willebrand's disease]. AB - A 38-year-old man with Von Willebrand's disease type 2 came to us for treatment advice in relation to a trip abroad, and also a 53-year-old woman with bleeding treated as idiopathic thrombocytopenic purpura (ITP). In the man, a trial dose of desmopressin led to severe thrombopenia, and in the woman, treatments, such as splenectomy and prednisone in the past, had been ineffective. In both patients further investigations led to the diagnosis 'Von Willebrand-disease type 2B'. Despite the fact that Von Willebrand's disease type 2B has distinctive laboratory characteristics, such as thrombocytopenia, a positive Ristocetin Induced Platelet Agglutination (RIPA) test at a low ristocetin concentration, and an abnormal multimer pattern, some cases have incomplete or atypical presentations which can be misleading for the diagnosis. A wrong diagnosis can lead to ineffective and potentially dangerous therapeutic interventions. Molecular genetic analysis of type 2B mutations is simple and can help in making the correct diagnosis in cases of familial thrombocytopenia or for a further characterisation of Von Willebrand type 2. PMID- 12109312 TI - [Fatal miliary tuberculosis during treatment with infliximab]. AB - A 73-year-old woman was admitted to hospital with fever whilst under treatment with infliximab for rheumatoid arthritis. Despite repeated and specific testing, tuberculosis was only diagnosed post mortem. During infliximab therapy, latent tuberculosis can reactivate in subacute form with a possible fatal outcome. For infliximab therapy to be administered safely, the risk that the patient concerned is latently infected with tuberculosis has to be estimated beforehand; if necessary a prophylactic anti-tuberculosis treatment may be given. PMID- 12109313 TI - [Transition of health care for adolescents with chronic diseases: from pediatrics to specialities for adults]. PMID- 12109314 TI - [Clinical thinking and decision making in daily practice. An old man with hyponatremia]. PMID- 12109315 TI - [Airway fire, a serious complication during laryngeal laser surgery]. PMID- 12109316 TI - Feeding disorders in infants and toddlers: diagnosis and treatment. AB - Each of the six feeding disorders described presents with specific symptoms, has a different origin, and responds to different interventions. Consequently, it is important to establish an accurate diagnosis and use the appropriate intervention for the specific feeding disorder. As the study by Benoit et al [7] demonstrates, an intervention that may be helpful for one feeding disorder can be completely ineffective for another. PMID- 12109317 TI - Eating disorders in school-aged children. AB - It is widely accepted that eating disorders do occur in children. There is a growing literature on childhood-onset AN, and it seems that the core behavioral, psychologic, and physical features are similar to those in adults. The differences between children and adults also must be taken into account, however. Because children have lower levels of body fat, they tend to become emaciated and suffer the effects of starvation for more quickly than adults, which must be taken into account when considering treatment. Although cases of childhood-onset BN have been reported, they are so rare that empirical research is difficult. Clinical features reported regarding the atypical childhood-onset eating disorders generally concur, although empirical testing of these features has yet to be developed. Theories as to why children develop these disorders need further development. The general consensus is that all childhood-onset eating disorders must be considered using a multidimensional model that takes into account physical, psychologic, social, and family factors in origin, assessment, and treatment. PMID- 12109318 TI - Eating disorders in adolescents and young adults. AB - Adolescence and young adulthood mark the convergence of substantial biologic and social change. Individuals differ in their innate capacity to tolerate change. Research suggests that some of the personality characteristics that are fundamental to individuals with eating disorders may render them particularly vulnerable to the impact of a changing body and changing social demands. The fact that eating disorders emerge primarily during adolescence and often run a protracted course can deprive the afflicted individual of the core social, emotional, and biologic developmental processes that normally occur during this time of life. PMID- 12109319 TI - Childhood and adolescent anorexia nervosa. AB - Eating disorders are common during childhood and adolescence. Early intervention is associated with the best prognosis. Treatment interventions that focus on achieving ideal body weight and use various family therapy approaches are most likely to be effective. Much remains to be learned about the origin of AN, but there are promising recent advances. PMID- 12109320 TI - Bulimia in children and adolescents. AB - Although there have been many recent advances in research, much work remains to be done in the area of child and adolescent eating disorders. More data are needed regarding the normal development of eating behavior, resilience and risk factors for eating pathology, and treatment studies in children and adolescents. The best studied areas include epidemiology, short-term treatment for bulimia nervosa (BN), and outcome in anorexia nervosa. A case report of the single blind use of repetitive transcranial magnetic stimulation in a patient with BN has been reported, but its safe use in children and adolescents remains to be established. PMID- 12109321 TI - Childhood obesity. AB - The prevalence of childhood obesity is rising, and pediatric obesity has become an important public health issue. It can be defined as BMI more than 95th percentile for age and sex, whereas overweight is defined as BMI more than 85th percentile. Using these cut points, more than one quarter of all children and adolescents are either overweight or obese. The recent increases in the prevalence of childhood obesity result from the interaction between a strong genetic predisposition that facilitates storage of fat, easy access to calorically dense foods, and the low levels of physical activity that characterize modern societies. Childhood obesity has pervasive psychosocial and medical consequences in the short term and the long term. Many of these consequences may not be apparent for decades, but the metabolic complications of obesity and the insidious effects of early psychosocial stigmatization are sometimes observed even in young children. For pediatric obesity, behavioral approaches seem moderately efficacious in younger children. Comprehensive programs generally include modification of dietary and activity habits and some degree of parental involvement. Medication for pediatric obesity cannot be recommended currently outside the context of clinical trials. Gastric bypass surgery is effective but should be considered a last resort for the child with life-threatening complications of obesity. PMID- 12109322 TI - Inpatient and partial hospital treatment for adolescent eating disorders. AB - The eating disorders are complex illnesses that tend to have a chronic relapsing course with severe morbidity and high mortality rates. Outcome seems to be best when the disorders are recognized early, brought to treatment quickly, the family is involved, and the first episode of care results in full return and maintenance of weight and menstruation. Adolescents who reach the point of needing hospitalization should be treated aggressively. In this article the authors have tried to outline some key treatment principles not just for the hospital stay, but elements that should be carried throughout the entire program of recovery for adolescents with eating disorders: from medical evaluation, through inpatient stay, partial hospitalization, intensive outpatient program, and follow-up outpatient therapy. Recent reductions in insurance authorizations and decreased lengths of stay in the psychiatric hospital make the already difficult challenge of recovery from AN and BN even more daunting. Despite these difficulties, we are still able to get a high proportion of youths better and eventually fully recovered. PMID- 12109323 TI - Individual psychotherapy for children and adolescents with an eating disorder from historical precedent toward evidence-based practice. AB - This article reviews the place of individual therapy in the suite of treatment services required by children and adolescents with an eating disorder. Individual therapy is defined and traced from historical origins in psychoanalytic practice with later important modifications by Hilda Bruch and Arthur Crisp. More recent developments, based primarily on cognitive and learning theory are discussed, as is the timing of individual therapy with respect to illness stage and patient motivation. Evidence for therapy effectiveness is reviewed where possible. At present, treatment evidence in the pediatric mental health field must be inferred from adult research. PMID- 12109324 TI - Treating adolescents with eating disorders in the family context. Empirical and theoretical considerations. AB - The author described the Maudsley approach for family treatment of adolescent AN and the empirical evidence supporting its use in this population. This treatment focuses on the family as a resource for recovery and puts the patients in charge of re-feeding their affected child. Its success seems to depend on the successful motivation of parents to take on this task and see it through while simultaneously supporting the processes of adolescent development as they reemerge. Although this treatment is promising, substantial data to support its being the best approach for adolescents with AN are lacking. The author also described a theoretical model for involving parents in CBT for adolescents with BN. Although CBT is accepted as the most efficacious treatment for adults with BN, it has not been tested systematically in adolescents. At the same time, it is clear that the adult models of CBT for BN are unlikely to be successful without modifications that take into account the realities of adolescence. Although CBT as a model is likely to be acceptable to adolescents, parents are needed to promote motivation, provide a supportive milieu for behavioral change, and provide guidance and support in stressful periods that lead to relapse. It is important that CBT that is appropriately modified to include parents be tested for its efficacy in adolescents with BN. Preliminary, uncontrolled results are promising. PMID- 12109325 TI - Art therapy, psychodrama, and verbal therapy. An integrative model of group therapy in the treatment of adolescents with anorexia nervosa and bulimia nervosa. AB - Anorexia nervosa and bulimia nervosa typically afflict individuals in adolescence. Given the intractability of these diseases in combination with the natural recalcitrance of adolescence, treatment with this population presents a daunting challenge. Traditional group therapy that focuses on verbal therapy is often not effective with this population, particularly in the acute stages of the diseases. A group therapy approach that integrates art therapy, psychodrama, and verbal therapy offers an innovative alternative to traditional group therapy. PMID- 12109326 TI - Pharmacotherapy and medical complications of eating disorders in children and adolescents. AB - In this article, the authors cover two areas of interest regarding eating disorders in childhood and adolescence: (1) the detection of eating disorders in medical practice and their medical complications and (2) the psychopharmacologic treatment of patients with eating disorders. PMID- 12109328 TI - Eating disorders in males. AB - Eating disorders are one of the rare psychiatric disorders with a large preponderance of female patients. The other articles in this issue review eating disorders in children and adolescents and focus primarily on female patients. This article reviews the eating disorders that occur in male children and teenagers, including anorexia nervosa, bulimia nervosa, binge eating disorder, a subtype of body dysmorphic disorder named muscle dysmorphobia, and obesity. This article reviews subgroups of boys who are at higher risk for developing eating disorders. The article commences with the difference in male perceptions of body image and dieting behaviors. PMID- 12109327 TI - The human genetics of eating disorders lessons from the leptin/melanocortin system. AB - Genetic analysis of eating disorders is complex phenotypically and genotypically. As seen in the leptin/melanocortin system, however, the results can lead to a deeper understanding and to new therapies. Benefits are expected for eating disorders that stem from genetic and psychologic causes. Finally, an awareness of possible genetic causes of eating disorders will help determine the causes--and thus the treatments--in children and adolescents with eating disorders, as exemplified by obese patients with mutations in the POMC, PC1, leptin, and MC4R loci. PMID- 12109329 TI - Eating disorders and health insurance understanding and overcoming obstacles to treatment. AB - Eating disorders are complex in cause and course, with biomedical, psychiatric, and psychosocial components. To maximize the likelihood of recovery, patients require skillful and coordinated care from various health care professionals, including medical professionals, mental health professionals, and nutritionists. All too often, at some point in treatment, clinicians discover that their patients' health insurance status has been compromised or that further treatment is denied because of maximization of mental health benefits. It is important for patients, families, and health care professionals to understand the health insurance issues that are involved in the treatment of eating disorders and learn strategies that may help to overcome the obstacles that these issues present. PMID- 12109330 TI - Beyond bioterrorism. PMID- 12109331 TI - Effects of polyethylene glycol molecular weight and concentration on lactate dehydrogenase activity in solution and after freeze-thawing. AB - Lactate dehydrogenase (LDH) is a tetrameric enzyme that has been used as a model for labile protein drugs. Polyethylene glycols (PEGs) have been proposed as excipients to stabilize labile proteins in solution and during the freezing portion of the lyophilization cycle. The aim of this study was to determine the effects of PEG molecular weight and concentration on the activity of LDH. In general, PEG protection increases with PEG molecular weight and concentration. PEGs slow the loss of activity in LDH solutions stored at 4 degrees C, but are not sufficiently effective to allow for a solution product. PEGs 8000, 10,000, and 20,000 show full freezing protection at less than 0.01%, while lower molecular weight PEGs need higher concentrations to produce protection upon freezing. Circular dichroism (CD) studies of LDH solutions before and after freezing with PEG 400 and PEG 8000 confirm the activity studies. The CD spectrum of LDH before freezing shows the classic alpha helix pattern. After unprotected LDH solution is frozen and thawed, the CD spectrum erodes. Low concentrations of PEG 8000 (1% or less) preserve the alpha helix profile after freezing of the samples. PEG 400 preserves the alpha helix CD profile in a stepwise fashion with increasing concentrations. The CD and activity data suggest that PEGs can protect alpha helix structures and activity of LDH through the freezing process. PMID- 12109332 TI - Pharmaceutical vials with extremely high chemical inertness. AB - Traditional type I glass containers are becoming more unsuitable for packaging modern diagnostics and drugs. When using certain drugs (high-pH formulations, complexing agents, drugs with low dosage, sensitivity to pH-shift or ion release), product degradation can occur, reducing the drug's performance, potency, and shelf-life. To prevent chemical interaction between the container and the content, a coating technology applying a silicon dioxide layer to the internal container surface can bring the solution. PMID- 12109333 TI - In defense of filters in pharmaceutical water systems. PMID- 12109334 TI - The Devonport incident, the Clothier Report, and related matters--30 years on. PMID- 12109335 TI - Method for qualifying microbial removal performance of 0.1 micron rated filters. Part IV: Retention of hydrogenophaga pseudoflava (ATCC 700892) and Ralstonia pickettii (ATCC 700591) by 0.2 and 0.22 micron rated filters. AB - Ralstonia pickettii has emerged as a bioburden microorganism of considerable importance in pharmaceutical processes utilizing conventional 0.2 or 0.22 micron rated "sterilizing grade" filters. In this article, we re-evaluated and studied the retention efficiencies of 0.2 micron rated nylon 6.6 and 0.22 microns rated modified polyvinylidene fluoride (PVDF) filters for Hydrogenophaga pseudoflava (ATCC 700892) and R. pickettii (ATCC 700591). Out of a total of forty-four 0.2/0.22 micron rated filters discs tested in this study (spanning different challenge fluids, different challenge conditions, and different filter types), H. pseudoflava penetration was observed for every filter disc tested. Log titer reduction (LTR) values ranged from 0.3 to 2.0 logs for 20-48 hour challenges conducted in Water for Injection (WFI), and 3.8-7.1 logs for 6-hour challenges conducted in Minimal Media Davis (MMD). For 0.2 micron nylon 6.6 filter discs, penetration by R. pickettii was observed only in WFI challenges and was dependent on the culture and challenge conditions used. Penetration by R. pickettii was also restricted to only those membrane discs that were very close to the filter manufacturer's production integrity test (the Quantitative Bubble Point, QBP, test) limit. Where R. pickettii penetration was observed, LTR values were significantly higher than those observed for H. pseudoflava with the same filter discs. This study: 1) supports the use of H. pseudoflava as a worst-case challenge model for R. pickettii in process- and product-specific bacterial retention testing; 2) provides experimental evidence, for the first time, for the need to include filter membrane lots that have a physical integrity test value at or near the filter manufacturer's production (lower) limit in these tests; and 3) demonstrates how a standardized membrane integrity test (such as the QBP test) can be used select such "worst-case" membranes and to verify the inclusion of such "worst-case" membranes in these tests, thus serving as the link between the membrane disc used in bacterial retention validation testing and the production process filter. PMID- 12109336 TI - Prevalence of complementary medicine in urologic practice. A review of recent studies with emphasis on use among prostate cancer patients. AB - The use of complementary and alternative medicine (CAM) has been used since antiquity for the relief of symptoms related to many diseases including urologic ailments. CAM is typically defined as those healing philsophies, approaches and therapies that are generally not taught in medical schools and are not usually reimbursed by medical insurance companies. Complementary and alternative medicine therapies are distinguished as those used alone (alternative) or in addition to conventional therapies (complementary). Few studies have focused on the urologic population. In this article, we summarize recent studies assessing the prevalence rate of CAM use with special emphasis in prostate cancer patients. PMID- 12109337 TI - Vitamin E and prostate cancer. AB - Descriptive epidemiologic evidence suggests that environmental exposures, such as diet, have an important causative role in the progression of prostate cancer. Increasing evidence also suggests that vitamin E may be beneficial in preventing prostate cancer or delaying disease progression. Clinical trials have been initiated that should definitively determine the therapeutic benefit, if any, of vitamin E. PMID- 12109338 TI - Dietary supplements and other alternative medicines for erectile dysfunction. What do I tell my patients? AB - Dietary supplements and other alternative medicines have enjoyed a tremendous amount of popularity and use over the past decade. Although, the prevalence of these therapies for erectile dysfunction (ED) is unknown at this time, numerous media outlets and alternative medicine publications seem to support the utilization of these therapies for ED. The placebo effect is approximately 25% (1 out of 4 benefit) from past randomized trials of FDA approved medications for ED. Adequate clinical trials are needed for dietary supplements for ED to access whether or not a benefit beyond a placebo effect exists. Clinicians should become aware of these supplements and the current research espousing or discouraging their use, and they should understand the adverse effects associated with them in order to effectively discuss these products with any patient inquiring about them. PMID- 12109339 TI - Lifestyle/dietary supplement partial androgen suppression and/or estrogen manipulation. A novel PSA reducer and preventive/treatment option for prostate cancer? AB - There is a large interest in prostate cancer prevention and/or slowing the progression of this disease via dietary/lifestyle/supplement interventions. Numerous mechanisms have been suggested as to how these interventions may lower PSA levels. However, it is possible that the primary mechanism of action is partial androgen suppression and/or estrogen manipulation. PMID- 12109340 TI - The placebo effect and randomized trials: analysis of conventional medicine. AB - Randomized controlled trials are generally regarded as the gold standard of study designs to determine causality. The inclusion of a placebo group in these trials, when appropriate, is critical to access the efficacy of a drug or supplement. The placebo response itself has received some attention in the medical literature over the past fifty years. The recent increasing utilization of dietary supplements and herbal medications by patients makes it imperative to reevaluate the placebo response in conventional and alternative medicine. This article will review a whole series of unique conditions (allergies/asthma, alopecia, BPH, erectile dysfunction, osteoporosis, weight loss...) and the placebo response associated with them from conventional medical randomized trials. PMID- 12109341 TI - The placebo effect and randomized trials: analysis of alternative medicine. AB - Randomized controlled trials are generally regarded as the gold standard of study designs to determine causality. The inclusion of a placebo group in these trials, when appropriate, is critical to access the efficacy of a drug or supplement. The placebo response itself has received some attention in the medical literature over the past fifty years. The recent increasing utilization of dietary supplements and herbal medications by patients makes it imperative to reevaluate the placebo response in conventional and alternative medicine. This article will review some of the negative and positive results from randomized trials utilizing dietary supplements (androstenedione, beta-carotene, CoQ10, garlic, soy, vitamin C and E...) for a number of non-urologic and urologic conditions, including cancer. PMID- 12109342 TI - Chemoprevention of bladder cancer. AB - The data presented herein, although highly supportive for a protective role of various nutrients against bladder cancer, are far from definitive. Many authorities question the validity of current recommendations for nutritional chemoprevention against bladder cancer. The reason for the wide variations reported in epidemiologic studies lies in the nature of observational studies. Dietary studies are limited in their conclusions because the protection afforded by the consumption of a particular nutrient may be multifactorial, with different components of the food exerting potential chemopreventive effects. Furthermore, measuring levels of nutrients in the food intake of populations is confounded by factors that might affect these levels and also the incidence of cancer. For example, vitamin A can come from animal or vegetarian sources. Because animal fat has been identified as a potential carcinogen in man, depending on the source of the vitamin, varying levels of protection might be deduced. In addition, chemoprevention studies using dietary supplements are expected to have mild effects, and large studies would be required to confirm statistical significance. Even with agents such as intravesical chemotherapy, only half the studies achieve statistical significance [29]. Prospective randomized trials with a large sample size, longer follow-up, and an extended duration of treatment are needed to clarify the association between micronutrients and cancer protection. With these caveats in mind, several recommendations can be made. Simple measures, such as drinking more fluids (especially water), can have a profound impact on the incidence of bladder cancer. Vitamins are being extensively studied in chemopreventive trials for different cancers. There is strong evidence for a chemoprotective effect of vitamin A in bladder cancer. The authors recommend 32,000 IU/day of vitamin A initially, with lower doses (24,000 IU) for persons less than 50 kg. Because liver toxicity is a possibility with long-term administration, the dose should be decreased to 16,000 IU after 3 years. High doses of beta-carotene should be avoided based on a large clinical trial reporting a 25% increase in the number of cases of prostate cancer and a statistically significant increase in the incidence of lung cancer. Vitamin B6 has been studied in several clinical trials in bladder cancer. The US-based Veterans Administration cooperative study found benefit for vitamin B6 when given as a single agent. Data for vitamins C and E are insufficient to recommend either agent as stand-alone treatment. Nonetheless, each of these vitamins is known to have beneficial effects, including improved function of the immune system. It is possible that only a small percentage of patients with bladder cancer respond to vitamins B6, C, or E, yet each is safe, nontoxic, and inexpensive. In an effort to pool the efficacy of individual agents and to increase the power of study, the authors evaluated the combination of vitamins A, B6, C, and E in a double-blind trial. The observed 50% 5-year reduction in tumor recurrence was highly significant and greater than would be expected for any of the individual ingredients and suggests that combinations of nutritional agents may be most appropriate. A large-volume study along similar lines is being conducted. Among the numerous other compounds and dietary substances purported to have chemopreventive effect, soybeans, garlic, and green tea stand out as having the greatest promise and can freely be recommended to patients. For synthetically synthesized agents such as celecoxib, piroxicam, or DFMO, recommendations must be deferred until the results of clinical trials are conclusively in favor of their use. Many of the dietary factors found to be protective against bladder cancer are being investigated in other cancers and are beneficial to general health. Although naturally occurring nutrients are ideal, especially because the delicate balance of various micronutrients might be impossible to synthesize in the laboratory, the general population finds it easier to take vitamin supplements. Unfortunately, dietary changes such as decreasing fat and increasing fruit and vegetable intake are more difficult to initiate. There is a mistaken notion that simply because an agent is naturally occurring, it cannot be as beneficial as taking a substance synthesized in the laboratory. Even in a high-risk group such as nuclear-bomb survivors in Japan, high consumption of vegetables and fruit is protective against bladder cancer [44]. Encouraging patients to follow an essentially healthy food habit lifestyle will be a significant contribution in the fight against cancer. PMID- 12109343 TI - Prostate cancer trends in the era of prostate-specific antigen. An update of incidence, mortality, and clinical factors from the SEER database. AB - Concurrent with the successful life-saving efforts in terms of prostate cancer diagnosis and treatment, some men who do not need treatment are receiving it. These are men destined to die of causes other than prostate cancer. Unfortunately, at diagnosis, men needing treatment for prostate cancer cannot be differentiated from men who do not. To make such decisions correctly for individual patients would require extremely precise measures of the time to death from prostate cancer versus when the patient would die from a competing cause. Predictive tools with this level of accuracy will never be available given the inherent uncertainty of life. At the time of prostate cancer diagnosis, the date and the cause of death for the patient are matters of weak statistical speculation. Unless the date of death from prostate cancer and the date of death from non-prostate cancer causes can be precisely determined for each patient, some men will always be overtreated or undertreated. Conservative strategies result in the undertreatment of some patients who would benefit from treatment while sparing other patients unneeded treatment. Aggressive strategies result in the overtreatment of patients who do not need therapy while curing other men of prostate cancer. Both strategies are correct, but only some of the time. Better methods of determining the length of life and cause of death may improve this situation, but not by much. [figure: see text] Dramatic shifts in the incidence, grade, stage, and age of men with prostate cancer have been observed with the advent of widespread PSA-based cancer detection in the United States. Grade and stage trends suggest that more biologically relevant (the shift from well differentiated to moderately differentiated tumors) and yet therapeutically amenable (earlier stage) tumors have been identified in large numbers of patients during the PSA era. Clearly many men have been diagnosed and treated who will not benefit from such treatment. The relative mix of these two groups of men is not known. Given the long delay between treatment and mortality that is inherent in prostate cancer (Fig. 14), the full effects of treatment on prostate cancer mortality are probably not yet seen in prostate cancer mortality data. PMID- 12109344 TI - Racial differences in prostate carcinogenesis. Histologic and clinical observations. AB - Racial differences in the prevalence of HGPIN and in the Gleason score of local stage cancers indicate that clinically observed racial differences in cancer incidence and stage at diagnosis reflect racial variability in prostate carcinogenesis. Exploration of genetic, hormonal, nutritional, and behavioral differences in black and white men may provide insight into the fundamental mechanisms of prostatic carcinogenesis and cancer progression. PMID- 12109345 TI - Clinical usefulness of assays for complexed prostate-specific antigen. AB - Tests for the tumor marker prostate-specific antigen (PSA) vary widely in specificity and sensitivity. Use of an assay that tests the complexed form of PSA (cPSA) results in improved specificity and sensitivity. By improving specificity, similar results are obtained by cPSA compared with use of the free-to-total PSA ratio but are measured by a single analyte instead of two. Assays for cPSA can serve as a substitute for total PSA in all current applications. PMID- 12109346 TI - Prostate biopsy strategies: past, present, and future. AB - Endorectal sonography for prostatic imaging was first described in 1968 and gained wide acceptance, particularly since the 1980s. Presently, extended biopsies consist of the sextant biopsy pattern plus various combinations of anteriorly directed biopsies and posterolateral sampling that includes the anterior horn of the peripheral zone. The cancer detection rate for transrectal ultrasound-guided prostate biopsies has fallen, and the repeat biopsy rate has risen. The future will most likely see fewer biopsies performed but with wiser guiding systems. PMID- 12109347 TI - Prediction of tumor heterogeneity in localized prostate cancer. AB - Clinical T1 and T2 prostatic carcinoma is a heterogeneous tumor with respect to pathologic stage and outcome. In the authors' experience, 60% of patients have a pT2 prostatic carcinoma, and 2% to 4% have tumors less than 0.5 cm3 in volume. The latter group cannot be predicted by the use of preoperative parameters with a sufficient sensitivity and specificity. Quantitative analysis of six systematic biopsies, that is, reporting the number of biopsies with any Gleason grade 4 or 5 cancer or the number of biopsies with more than 50% Gleason grade 4 and 5 cancer, together with preoperative PSA levels can be used to predict the different pathologic stages and risk groups of patients with T1 or T2 prostatic carcinoma. CART analysis that using these preoperative parameters can predict the lymph node stage and the capsular penetration on each side of the prostate with a sufficient positive and negative predictive value and a sufficient specificity to avoid routine lymphadenectomy in approximately 80% of the patients classified as a low risk group for having lymph nodes positive for disease. CART analysis also allows a solid identification of patients in whom the unilateral or bilateral nerve may be spared during surgery. These algorithms may be improved further by determining the HK-2 level in the blood or by including other molecular biologic markers in the analysis of the biopsies. Clinical T1 or T2 prostatic carcinoma is a heterogeneous but fairly predictable tumor. PMID- 12109348 TI - Timing hormonal therapy in prostate cancer. AB - The history of the use of hormonal therapy to treat prostrate cancer is outlined, focusing on such treatments and disease scenarios as diethylstilboestrol therapy, androgen ablation alone or in conjunction with radiation, and the treatment of minimal disease burdens. Studies have pointed to the benefits of initiating androgen ablation therapy earlier in the treatment of prostate cancer; the greatest benefits of anti-androgen therapy may be gained when only tiny amounts of prostate cancer are present. PMID- 12109349 TI - Current concepts of tumor markers in bladder cancer. AB - The critical issue in characterizing the usefulness of tumor markers in screening or monitoring for urothelial cancer and in determining prognosis revolves around the question, "What do we need to know?" During surveillance of patients with low risk disease, highly sensitive markers may detect disease persistence or early recurrence (on the basis of field-effect changes or tumor cell implantation); however, they may not indicate the possibility of progression. If cystoscopy is negative in these instances, is it appropriate to treat? Does earlier treatment actually prevent recurrence or progression? Some investigators have suggested a positive answer to the former question, although lead-time bias may a role. The issue in the latter question may be moot because low-risk disease is by definition unlikely to progress. A corresponding question is whether one should treat the patient if the marker has a low specificity. In such instances, a marker with low specificity may incur more frequent instrumentation and a panoply of additional costs. A marker may have some value in providing earlier diagnosis in low-risk disease if treatments are available to prevent clinical expression and recurrence. A tumor marker would have additional value if it could detect biologic change from low-risk to high-risk disease and permit a corresponding change in treatment. Moreover, if a marker could provide earlier diagnosis in high-risk disease, it might permit identification of those patients likely to progress at earlier and curable stages of disease. To be successful, such markers must have high sensitivity and specificity in diagnosis. It has been suggested that "a simple noninvasive highly sensitive and specific method for detecting bladder cancer would decrease the morbidity associated with current surveillance methods, improve quality of life for patients, and reduce costs by substituting a less expensive test for the more expensive endoscopic procedures" (Dr. M. Soloway, personal communication 2000). In considering this statement, it is tempting to question some of its implications. The term "simple" suggests that point-of-care tests may be the easiest, but tests that are available may be inconsistent and not sufficiently sensitive or specific. The need for a "highly sensitive and specific" test is of critical importance for applicability in the individual, even when such tests are based on the results in large populations. When considering the "morbidity of current methods," it is tempting to question whether cystoscopy is that morbid a procedure. Given the information it provides, it seems improbable that it can be readily replaced. Currently, no markers provide information over and above that provided by cystoscopy and histology- or cytology-based understanding of the biology of the disease especially in considering distinctions between low-risk and high-risk disease. The ability to "improve quality of life" will depend on the use of the predictive value made on the basis of a test's sensitivity and specificity. A false-positive test may lead to anxiety and extensive unnecessary evaluation, whereas a false-negative test may lead to missed diagnosis, delayed treatment, and the possibility of disease progression until accurate diagnosis is obtained. In each case, quality of life is compromised. "Substituting a less expensive test" depends on the benefits of that substitution and the practicality of omitting or delaying standard assessment. Costs depend on the frequency of testing, those incurred by false positive tests, and those incurred by false-negative tests. Several caveats emerge when taking into account these various considerations. In screening for low-risk disease, the urgency of diagnosis may be less important. High sensitivity is of less importance because the risk of a missed diagnosis has less ominous repercussions. Nevertheless, high specificity is important to avoid unnecessary frequent instrumentations. In considering surveillance for low-risk disease, cytology may be more useful in indicating a change from negative (persistent or recurrent low-risk disease) to positive (high-risk disease) with the consequent need for more aggressive treatment approach. No urinary "screening" markers reliably provide this information. In considering surveillance for high-risk disease, sensitive markers would detect persistence or early recurrence. If such markers are negative, intervals between repeated cystoscopies may be prolonged; however, a high specificity is needed to ensure the validity of a truly negative interpretation. Otherwise, diagnosis may be missed and the risk from progression and the development of incurability increased. Markers for urothelial cancer must be expressed exclusively as a consequence of the presence of cancer cells. The test must be sensitive in detecting disease and must be validated in nonselected populations. It must be sensitive in excluding nondisease, and noncancer conditions must have little influence on its accuracy. There must be little or no interassay or intra-assay variability. Interpretation of an assay for a tumor marker should be all or none, or based on a threshold level. The assays must have sufficient sensitivity and specificity in populations and in the individual to have practical clinical applicability. PMID- 12109350 TI - Herbal medications in the treatment of benign prostatic hyperplasia (BPH). AB - Herbal medications are used widely in the treatment of BPH. Recent studies suggest a benefit for some of these products with few side effects. The results of these studies are summarized and the possible mechanism of action of these medications are reviewed. PMID- 12109351 TI - Management of chronic prostatitis-chronic pelvic pain syndrome. AB - Although the neurobiologic basis of CPPSs in men remains unclear, therapeutic interventions should continue to be improved. Invasive or destructive modalities should be avoided when possible. Electrical neuromodulation techniques seem to be a promising, among other multimodal approaches. Physicians must learn from patients in attempt to relieve symptoms. PMID- 12109352 TI - New concepts in the pathogenesis of urinary tract infections. AB - Random amplified polymorphic DNA fingerprinting was used to distinguish among Escherichia coli bacterial strains creating urinary tract infections (UTIs) in women. Bacteria bound more avidly to cells from postmenopausal donors with history of UTIs (PK) compared with cells from women without history of UTIs (AO). Nonpiliated bacterial strains did not adhere to the cell lines. Increasing S-IgA concentrations has no effect on AO cell bacterial binding, whereas bacterial adhesion to PK cell epithelium increased with increasing S-IgA concentration. PMID- 12109353 TI - Prospective study of urinary tract infections and urinary antibodies after radical prostatectomy and cystoprostatectomy. AB - The authors have prospectively documented that men who undergo orthotopic bladder substitution more frequently experience bacteriuria than do normal men [19] or men with carcinoma of the prostate scheduled to have radical prostatectomy (see Table 1). Because the frequency of bacteriuria in men after prostatectomy was also lower than that after orthotopic bladder substitution (see Table 1), removal of the prostate and any of its presumed antibacterial properties probably does not account for this difference. Furthermore, the authors' data (see Table 5 ) and that of Woodside and associates [23] demonstrate that intestine incorporated into the genitourinary tract generates a local antibody response against urinary bacteria. Although others have suggested that the incorporation of bowel in the urinary tract may be associated with increased bacteriuria, this effect has never been documented prospectively. The mechanism of this increased frequency of bacteriuria is unknown. Because the anatomy of the male secretory genitourinary system may be altered after radical prostatectomy and orthotopic bladder substitution, the authors evaluated local antibody production before and after these operations. More than 20 years ago, Burdon [5] found that the initial portion of the VB1 sample in men had significantly higher levels of IgA compared with the VB2 specimen, whereas the levels of IgG were similar in the two portions. This latter finding was confirmed by Shorliffe and co-workers [22] when they examined prostatic secretion. Other investigators have found high levels of IgA in human prostatic tissue and fluid. [24,25]. On the basis of these findings, it was believed that, in men, the prostate produces most of the urinary IgA, whereas the bladder or upper urinary tracts make most of the urinary IgG. Although the authors' study confirms that most local urinary tract IgG is produced by the bladder or upper urinary tracts, this study documents that the prostate is not the only source of urethral IgA in men. Despite almost complete removal or prostate secretory epithelium by radical prostatectomy, as evidenced by a dramatic fall in postoperative VB1 and VB2 PSA compared with preoperative levels (Table 3). men who had this operation had only slightly decreased IgA levels after the operation (Table 4, Fig I). The source of this IgA must be urethral because the VB1 urinary stream contains more IgA than the VB2 urine even after radical prostatectomy. The authors have not determined whether the urinary IgA concentrations observed after radical prostatectomy are the true baseline values for a man without a prostate, or whether they actually reflect abnormal production of local IgA stimulated by radical prostatectomy. Because post prostatectomy bacteriuris occurred frequently during urethral catheter drainage, the authors screened for postoperative IgA titers to mix 1 and mix 2 to determine whether specific production of antibody against gram-negative organisms might account for some of the postoperative IgA measured. Postradical prostatectomy mix 1 and mix 2 titers were not elevated, compared with preoperative measurements. Because urethral glandular tissue other than prostatic tissue is present in the male urethra, these glands also might be responsible for significant local antibody production. The high levels of urinary IgA and IgG after cystoprostatectomy with ileal orthotopic bladder substitution document that intestine incorporated into the urinary tract is still capable of producing local antibody. This observation corresponds with the findings of Mansson and associated [26] of elevated IgA and IgG in ileal reservoir urine compared with normal urinary tracts. It has been estimated that 1 m of intestine may secrete up to 780 mg/d of IgA [27], indicating that normal intestine production of antibody alone can account for the high IgA and IgG levels found in the patients who underwent bladder substitution. Interestingly, the ratio of IgA to IgG concentration in smal intestine fluid is 2:129, similar to the ratio of IgA to IgG in bladder substitution urine (2.92.1:52, Table 4). Because mix 1 and mix 2 IgA concentrations were elevated in VB1 and VB2 urine after ileal bladder substitution (see Table 5), some of this antibody was produced by the ileal bladder substitution in response to the inevitable bacteriuria that occurs during the prolonged postoperative catheter drainage. The findings is absent after radical prostatectomy alone. In addition, some of this increased antibody might be a result of the increased bacteriuria noted in the patients who underwent ileal bladder substitution after the initial postoperative period. The significance of the increased bacteriuria and elevated antibody levels after ileal bladder substitution is unclear. Because most of these episodes of bacteriuria were asymptomatic, whether they represent clinical infections that should be treated is not known. Bishop and associates [28] found that the bacterial flora of ileal conduits with asymptomatic bacteriuria had bacterial counts of 1000 or fewer colonies, and they noted that the healthy ileum in situ may contain more than 10,000 organisms per milliliter [29]. Because the normal urinary tract is usually sterile, it is possible that the bacteriuria found by the authors after ileal bladder substitution represents some form of bowel colonization more commonly associated with the bowel rather than clinical urinary tract infection and has limited clinial importance. Trinchieri and associated [30] found that urinary from patients with ileocystoplasty prevented attachment of E. coli to human uroepithelial cells more effectively that urine from patients with recurrent urinary infections. This observation suggests that the relatively large quantities of Iga produced by the ileal bladder substitution may, in fact, prevent clinical infection by preventing tissue invasion by the bacteria. Only long-term follow-up of patients with ileocystoplasty or ileal bladder substitution will determine the clinical significance of the bacteriuria. The authors' study had documented an increased incidence of bacteriuria in men after ileal bladder substitution and no such increase after radical prostatectomy. Analysis of the data shows that male sources other than the prostate--probably urethral glands-- must produce significant quantities of local urinary tract IgA. After ileal bladder substitution, the incorporated ileum may produce volumes of local antibody that may exceed the amounts ordinarily produce by the normal urinary tract. The clinical significance of the increased incidence of bacteriuria and elevated antibody levels in patients after illeal bladder substitution is unclear. PMID- 12109354 TI - Potential lifestyle and dietary supplement options for the prevention and postdiagnosis of bladder cancer. AB - Apart from smoking, certain occupational exposures, and schistosomiasis, little is known about other potential lifestyle risk factors for bladder cancer. Other investigations thus far have also been important because of the large number of individuals who are diagnosed with this cancer that apparently have no known risk factors. Preventing the recurrence of bladder cancer has generated some interest because several preliminary trials have found that a combination dietary supplement of vitamins and minerals or a probiotic agent (Lactobacillus casei) may impact this outcome favorably. Advising patients on some of these lifestyle modifications is currently recommended because the majority of them are also currently recommended for cardiovascular disease reduction. PMID- 12109355 TI - Green tea and prostate cancer. AB - Many laboratory studies and human epidemiological data suggest that most prostate cancer deaths are attributable to lifestyle, including nutritional factors where diet plays a major role in initiation as well as subsequent progression of the disease. Under these circumstances, chemoprevention seems to be a logical and obvious strategy. Because of its long latency and high incidence, prostate cancer is an ideal disease for chemoprevention. The suitable agent(s) for prostate cancer chemoprevention should be the one(s) that has efficacy in the laboratory experiments on one hand, and also possesses proven epidemiological basis on the other hand. In this article, we address the use of green tea for prostate cancer chemoprevention. Experimental as well as the epidemiological basis for this possibility is provided. PMID- 12109356 TI - PC-SPES and prostate cancer. AB - PC-SPES (Botanic Lab, Brea, CA) is a combination of eight herbal therapies with activity against prostate cancer, both in vitro and in vivo. Studies in human prostate cancer cell lines demonstrate significant dose-dependent decreases in cellular viability after exposure to extracts of PC-SPES. Clinical studies suggest that PC-SPES can reduce specific antigen levels in both androgen dependent and androgen-independent prostate cancer patients. Toxicity is mild, thought there is an approximately 5% risk of thromboembolic events with treatment. Although PC-SPES has apparent estrogenic activity, controversy exists over the exact mechanism of its effects. PMID- 12109357 TI - Prostate cancer and selenium. AB - The important progress achieved in the treatment of prostate cancer comes by exacting significant costs [11, 16-18, 20, 23, 25]. Currently, there is incomplete evidence that the radical interventions at hand significantly reduce the human costs of the disease. Surgery and radiotherapy induce substantial risks of incontinence and impotence. The PSA test has probably decreased the stage at which prostate cancer is diagnosed [15]. Nonetheless, the PSA is a means of earlier detection; it does not elucidate quantitatively distinct modes of treatment. The PSA test is not a means of prostate cancer prevention. The continuing incidence, morbidity, and mortality imposed by this disease strongly indicate that preventive strategies for its control are necessary. Chemoprevention with selenium and other agents offers a promising approach that is undergoing intensive investigation. Randomized trials underway at the authors' center are building on the important clinical trial results reported by Dr. Larry C. Clark. These studies will evaluate the activity of selenium at several points along a continuum ranging from cancerous prostatic tissue in men with diagnosed cancer to premalignant tissue in men with high-grade PIN to healthy tissue in high-risk men with negative biopsy to long-term effects on cancerous tissue in men with frank cancer. These trials will also offer an opportunity for preliminary evaluation of the mechanisms by which selenium treatment could result in the slower development or progression of prostate cancer. PMID- 12109358 TI - The role of soy phytoestrogens in prostate cancer. AB - Epidemiological data of phytoestrogens and prostate cancer strongly supports the cancer protective effects of isoflavones found in soy products. Inhibition of cell proliferation via hormone-dependent and hormone-independent mechanisms by soy phytochemicals has been studied extensively in cell culture and animal studies. Herein, we review the current literature on the epidemiology and effects of two soy phytoestrogens, genistein and daidzein, and would stress the need for controlled human trials to assess the true preventive and therapeutic effects of these compounds. PMID- 12109360 TI - Vitamin D and prostate cancer. AB - Current approaches to the management of prostate cancer include surgery, radiation therapy or hormonal manipulation either individually or in combination. Diet is increasingly being recognized as playing a role in many cancers including that of the prostate. There is now considerable evidence suggesting a role for vitamin D in prostate cancer. In this article, we have reviewed the current evidence supporting the use of vitamin D in the prevention and treatment of prostate cancer. PMID- 12109359 TI - Tomato products, lycopene, and prostate cancer risk. AB - Several case-control and large prospective studies focusing on dietary assessment suggest that the intake of tomatoes and tomato products may be associated with a lower risk of prostate cancer [18]. Although less certain at present, the accumulated data suggest that the benefit may be most pronounced in the protection against more advanced or aggressive prostate cancer. It is possible that lycopene is one of the compounds in raw and processed tomato products that may contribute to a lower risk of prostate cancer; however, this hypothesis remains to be further investigated. Other carotenoids and phytochemicals in tomato products may also contribute to the proposed health benefits. Food processing does not seem to reduce the benefits but may, in fact, enhance the bioavailability of beneficial components. The reported correlations or associations between the consumption of tomato products and prostate cancer risk should not be interpreted as causal until additional data are available from a variety of studies in different populations. Ideally, randomized controlled intervention studies would provide an ultimate test of the tomato/lycopene hypothesis; however, the expense, long duration of exposure, and the near universal consumption of tomato products among Americans make a dietary intervention study difficult to undertake. It is reasonable to recommend to the general population the consumption of tomato products at approximately one serving per day or five servings per week as part of an overall healthy dietary pattern that may reduce the risks of prostate cancer, other malignancies, or other chronic diseases. This recommendation is consistent with current dietary guidelines to increase fruit and vegetable consumption to lower the risk of heart disease and many types of cancer [38]. Nutritional prevention of prostate cancer is very different from the use of dietary or nutritional treatments for established prostate cancer. The use of lycopene and other extracts for the treatment of prostate cancer is a separate issue that warrants individual attention and investigation. PMID- 12109361 TI - Precursors to onset clusters in acquisition. AB - Two lawful relationships involving word-initial onset clusters have been advanced in the acquisition literature; namely, that clusters imply affricates (Lleo & Prinz, 1996, 1997), and that liquid clusters imply a liquid distinction (Archibald, 1998). This study evaluated and extended the validity of these implicational laws in a population of 110 children (aged 3;0 to 8;6) with functional phonological delays who contributed extended speech samples for computational analyses. Results indicated that, for the most part, the composition of children's sound systems were in compliance with the proposed laws; however, there were noted asymmetries and apparent exceptions in the data. The asymmetries motivated an integration of the two laws to reveal a pattern of segmental-prosodic cyclicity consistent with deterministic models of phonological acquisition. The apparent exceptions highlighted the relevance of independent methodologies and offered a potential theoretical alternative with the Resolvability Principle as directions for future research. PMID- 12109362 TI - Processes in language acquisition: the roles of gender, attention, and maternal encouragement of attention over time. AB - This longitudinal study including 87 infant-mother dyads examined the relation between infant temperamental attention, maternal encouragement of attention, language, and the effects of gender. At ages 0;4, 0;8, and 1;0, global attention was assessed from Rothbart's (1981) IBQ; manipulative exploration was assessed with the Bayley (1969) IBR; and maternal verbal, visual and physical encouragement of attention were coded from 5 minutes of mother-infant free-play. At 1;0, language was measured using language items from the Bayley Mental Scale and parent-report items from Hendrick, Prather & Tobin's (1984) SICD-Revised. Multiple regressions indicated that gender, infants' manipulative exploration and maternal physical encouragement of attention at 0;4, and maternal verbal encouragement of attention at 1;0, were all positively related to language at 1;0. Interactions indicated that girls high in 0;8 or 1;0 manipulative exploration had more advanced language skills than girls low in manipulative exploration or than boys, regardless of their attention level. Additionally, maternal verbal encouragement of attention appears to be particularly salient in the development of language for boys. PMID- 12109363 TI - Language-general and language-specific influences on children's acquisition of argument structure: a comparison of French and English. AB - This research investigates language-general and language-specific properties of the acquisition of argument structure. Ten French preschoolers enacted forty sentences containing motion verbs; sixteen sentences were ungrammatical in that the syntactic frame was incompatible with the standard argument structure for the verb (e.g. *Le tigre va le lion = *The tiger goes the lion). Previous work (Naigles, Fowler & Helm, 1992, 1995; Naigles, Gleitman & Gleitman, 1993) indicated that English-speaking two-year-olds faced with such ungrammatical sentences consistently altered the usual meaning of the verb to fit the syntactic frame (frame compliance) whereas adults faced with the same sentences altered the syntax to fit the meaning of the verb (verb compliance). The age at which children began to perform Verb Compliantly varied by frame and by verb. The current study finds that the level of Verb Compliance in French five-year-olds largely mirrors that of English-speaking five-year-olds. The sole exception is the intransitive frame with an added prepositional phrase (e.g. *Le tigre amene pres de la passerelle = *The tiger brings next to the ramp), which elicits a higher level of Verb Compliance among French kindergarteners than among their English learning peers. This effect may be due to the unambiguous interpretation of French spatial prepositions (i.e. next to has both locative and directional interpretations whereas pres de supports only the locative interpretation). These data support the conclusion that the acquisition of argument structure is influenced by both language-general mechanisms (e.g. uniqueness, entrenchment) and language-specific properties (e.g. prepositional ambiguity). PMID- 12109364 TI - German children's productivity with tense morphology: the Perfekt (present perfect). AB - Two nonce-word studies examined German-speaking children's productivity with the Perfekt (present perfect) from 2;6 to 3;6. The German Perfekt consists of the past participle of the main verb and an inflected form of an auxiliary (either haben 'have' or sein 'be'). In Study 1, nonce verbs were either introduced in the infinitival form, and children (seventy-two children, aged 2;6 to 3;6) were tested on their ability to produce the Perfekt, or introduced in the Perfekt, and children were tested on their ability to produce the infinitive. In Study 2 twenty-four children aged 3;6 were given the past participle form of nonce verbs to see if they could supply the appropriate auxiliary (based mainly on verb semantics). The results were that many children as young as 2;6 used past participles productively (more than used infinitival forms productively), but all children had much difficulty in supplying both auxiliaries appropriately. The current findings suggest that mastery of the Perfekt construction as a whole does not take place before the age of four and that frequency of exposure is an important factor in determining the age at which children acquire grammatical constructions. PMID- 12109365 TI - The development of regular and irregular verb inflection in Spanish child language. AB - We present morphological analyses of verb inflections produced by 15 Spanish speaking children (age range: 1;7 to 4;7) taken from longitudinal and cross sectional samples of spontaneous speech and narratives. Our main observation is the existence of a dissociation between regular and irregular processes in the distribution of errors: regular suffixes and unmarked (non-alternating) stems are over-extended to irregulars in children's inflection errors, but not vice versa. We also found that overregularization errors at all ages are only a small minority of the children's irregular verbs, that the period of overregularization is preceded by a stage without errors, and that the onset of over-regularizations is connected to the emergence of obligatory finiteness markings. These findings are explained in terms of the dual-mechanism model of inflection. PMID- 12109366 TI - Learning to construct verbs in Navajo and Quechua. AB - Navajo and Quechua, both languages with a highly complex morphology, provide intriguing insights into the acquisition of inflectional systems. The development of the verb in the two languages is especially interesting, since the morphology encodes diverse grammatical notions, with the complex verb often constituting the entire sentence. While the verb complex in Navajo is stem-final, with prefixes appended to the stem in a rigid sequence, Quechua verbs are assembled entirely through suffixation, with some variation in affix ordering. We explore issues relevant to the acquisition of verb morphology by children learning Navajo and Quechua as their first language. Our study presents naturalistic speech samples produced by five Navajo children, aged 1;1 to 4;7, and by four Quechua-speaking children, aged 2;0 to 3;5. We centre our analysis on the role of phonological criteria in segmentation of verb stems and affixes, the production of amalgams, the problem of homophony, and the significance of distributional learning and semantic criteria in the development of the verb template. The phenomena observed in our data are discussed in light of several proposals, especially those of Peters (1983, 1995), Pinker (1984), Slobin (1985), and Hyams (1986, 1994). PMID- 12109367 TI - The development of complex verb constructions in British Sign Language. AB - This study focuses on the mapping of events onto verb-argument structures in British Sign Language (BSL). The development of complex sentences in BSL is described in a group of 30 children, aged 3;2-12;0, using data from comprehension measures and elicited sentence production. The findings support two interpretations: firstly, in the mapping of concepts onto language, children acquiring BSL overgeneralize the use of argument structure related to perspective shifting; secondly, these overgeneralizations are predicted by the typological characteristics of the language and modality. Children under age 6;0, in attempting to produce sentences encoded through a perspective shift, begin by breaking down double-verb constructions (AB verbs) into components, producing only the part of the verb phrase which describes the perspective of the patient. There is also a prolonged period of development of non-manual features, with the full structure not seen in its adult form until after 9;0. The errors in the use of AB verbs and the subsequent protracted development of correct usage are explained in terms of the conceptual-linguistic interface. PMID- 12109368 TI - Relevance and early word learning. AB - Several theorists have proposed that children may interpret an ambiguous word by attending to the dimension that is most relevant in the immediate discourse context. The current study offers a direct test of this hypothesis. Children aged 2;6 and 3;4 (N = 24 in each group) were presented with a novel object with an unusual shape and texture and were told 'This is a dacky one'. In the Shape Relevant condition, two other objects' shapes were described before the target object was labelled ('This is a round one; this is a square one'). In the Texture Relevant condition, two preceding objects' textures were described ('This is a smooth one; this is a fuzzy one'). Subsequent comprehension tests indicated that, in extending the novel adjective to other exemplars, children attended to the dimension that was most relevant to the preceding discourse context. PMID- 12109369 TI - Naturalistic language sampling in typically developing children. AB - This study compared naturalistic samples of three features of language in 30 two year-olds--total utterances, word roots, and MLU--in the home in three contrasting situations: the child observed playing by her/himself with mother near by, the child and mother observed in direct play interaction, and the child and mother unobserved at a time the mother judged would provide a sample of the child's 'optimal' language. Children produced more utterances and word roots and expressed themselves in longer MLU when in interaction than when playing 'alone', but children's utterances, word roots, and MLU were greatest in the 'optimal' language production situation. Girls used more word roots and spoke in longer MLU (especially in the 'optimal' language situation) than boys. Despite mean level differences, children maintained their rank orders across the three situations in use of word roots and in MLU. These findings have implications for understanding children's language and the representativeness of sampling child language. PMID- 12109370 TI - The acquisition of compound vs. phrasal stress: the role of prosodic constituents. AB - This paper investigates the acquisition of compound vs. phrasal stress (hot dog vs. hot dog) in English. This has previously been shown to be acquired quite late, in contrast to recent research showing that infants both perceive and prefer rhythmic patterns in their own language. Subjects (40 children in four groups the averages ages of which are 5;4, 7;2, 9;3 and 11;6 and 10 adults) were shown pairs of pictures representing a compound word and the corresponding phrase. They heard a prerecorded tape with the names of the items, and were asked to indicate which one they heard. In addition to 9 real compounds and corresponding phrases, 9 novel compounds were presented (redcup = invented type of flower vs. red cup). A gradual increase in overall correct scores until age twelve was found along with a significant effect of real vs. novel compounds (p < 0.001), and an overwhelming tendency for the younger children to prefer compounds regardless of stress. We conclude that the results are due to the slow development of the ability to use prosodic information to override a strong lexical bias. PMID- 12109371 TI - Restructuring of similarity neighbourhoods in the developing mental lexicon. AB - Previous evidence suggests that the structure of similarity neighbourhoods in the developing mental lexicon may differ from that of the fully developed lexicon. The similarity relationships used to organize words into neighbourhoods was investigated in 20 pre-school children (age 3;7 to 5;11) using a two alternative forced-choice classification task. Children classified the similarity of test words relative to a standard word to determine neighbourhood membership. The similarity relationship between the test and standard words varied orthogonally in terms of type of similarity and position of overlap. Standard words were drawn from neighbourhoods differing in density. Results showed that dense neighbourhoods were organized by phoneme similarity in the onset + nucleus or rhyme positions of overlap. In contrast, sparse neighbourhoods appeared to be organized by phoneme similarity in the onset + nucleus, but manner similarity in the rhyme. These results are integrated with previous findings from infants and adults to propose a developmental course of change in the mental lexicon. PMID- 12109372 TI - Between-word junctures in early multi-word speech. AB - Most children aged 1;6 to 2;0 begin to use utterances of two words or more. It is therefore important for child phonologists to consider the development of phonetic and phonological phenomena that characterize connected speech. The longitudinal case study reported here investigated three juncture types- assimilation, elision and liaison--in the speech of a typically-developing child between the ages of 2;4 and 3;4. Attempts at production of these adult juncture types occurred from the onset of two-word utterances. However, for some juncture types, the child still had to perfect the intergestural relationships and gestural articulations that the adult between-word junctures demand. This process of phonetic development was largely accomplished by the age of 3;4. With one exception, between-word junctures appear not to be the result of learned phonological rules or processes. The exception is liaison involving /r/, which did not occur until the child was three years old. PMID- 12109373 TI - Grammatical and situational aspect in French: a developmental study. AB - We study the ability of children to provide an appropriate continuation for a stimulus sentence, taking into account the joint demands of situational aspect and grammatical aspect. We hypothesize that the aspectual transitions required by some aspectual combinations play a role in the difficulty of providing an approrpriate continuation for them. We tested 130 French-speaking children of 5;06 to 9;0. In general, the data are consistent with the idea that the ability of children to construe an appropriate continuation for a stimulus clause is a function of both the situational aspect of the clause and the grammatical aspect provided by the verbal morpheme. There is a significant tense x situational aspect interaction in the number of continuations that children are able to provide in answer to the stimuli. Contrary to our expectations, there is no significant tense x situational aspect in the number of appropriate continuations, this being perhaps due to the small number of continuations for each stimulus type, but there are trends in the expected direction, which further studies may be able to confirm. PMID- 12109374 TI - Overt subjects in English: evidence for the marking of person in an English Italian bilingual child. AB - Data from one English-Italian bilingual child (1;10-3;1) are presented in this study which challenge the hypothesis that the consistent realization of overt subjects in English is caused by the emergence of finite verbal morphology in the child's grammar. The argument is made for the emergence of subjects as an independent grammatical property of English, namely the marking of person deixis. Throughout the period of observation there is a significant proportion of overt subjects in the child's English utterances appearing both with finite and non finite verb forms. Production of subjects stabilizes at 90% of obligatory contexts when no morphological correlates of finiteness have been acquired yet. While subjects are produced at significantly lower rates in Italian, we observe the consolidation of a number of inflected forms marking person agreement. The emergence of overt subjects in English on the one hand, and of subject-verb agreement in Italian on the other suggest that this bilingual child is grammaticalizing the all-important function of person deixis in language-specific ways: the same function is expressed by different forms in the child's two languages. PMID- 12109375 TI - Basic-level nouns: first learned but misunderstood. AB - It is commonly believed that first-learned words correspond with first-learned categories (both described as 'basic level') leading to the belief that language acquisition is a reasonably good indicator of early cognition. However, toddlers often overextend their first words. Do these errors reflect their comprehension? Two experiments were conducted in order to examine two-year-olds' production and comprehension of basic-level terms. The results showed overextensions both in production (e.g. children labelled a rocket 'airplane') and comprehension (e.g. they pointed to a rocket when airplane was requested). One reason toddlers extend labels to a wider conceptual domain is because they have not clearly differentiated basic-level concepts from related conceptual categories. PMID- 12109376 TI - The influence of exposure to phonological neighbours on preschoolers' novel word production. AB - We investigated the influence of exposure to phonologically similar words on four year-olds' acquisition of novel object words. In Experiment 1, hearing phonological neighbours before learning a new word did not influence children's novel word productions. In Experiment 2, when children heard the phonological neighbours of a novel word after learning a new word, they correctly produced the target word more often than children who did not receive this exposure. These findings suggest that exposing children to similar sounding words after a novel word was introduced may have helped maintain a representation of that word in working memory, leading to enhanced word learning. PMID- 12109377 TI - Negative commands in Spanish-speaking children: no need for recourse to relativized minimality (a reply to Grinstead, 2000). PMID- 12109378 TI - A study of relative clauses in Williams syndrome. AB - Despite growing empirical evidence to the contrary, claims continue to be made that the grammar of people with Williams syndrome (WS) is intact. We show that even in a simple elicited imitation task examining the syntax of relative clauses, older children and adults with WS (n = 14, mean age = 17;0 years) only reach the level of typical five-year-old controls. When tested systematically in a number of different laboratories, all aspects of WS language show delay and/or deviance throughout development. We conclude that the grammatical abilities of people with WS should be described in terms of relative rather than absolute proficiency, and that the syndrome should no longer be used to bolster claims about the existence of independently functioning, innately specified modules in the human brain. PMID- 12109379 TI - Developing linguistic literacy: a comprehensive model. AB - This is a position paper modelling the domain of linguistic literacy and its development through the life span. It aims to provide a framework for the analysis of language development in the school years, integrating sociolinguistic and psycholinguistic notions of variation, language awareness, and literacy in a comprehensive model. The paper focuses on those aspects of literacy competence that are expressed in language as well as aspects of linguistic knowledge that are affected by literacy competence, tracing the route that children take in appropriating linguistic literacy as part of their cognitive abilities and examining the effect of literacy on language across development. Our view of linguistic literacy consists of one defining feature: control over linguistic variation from both a user-dependent ('lectal') and a context-dependent (modality, genre, and register) perspective; of one concomitant process: metalanguage and its role in language development; and of one condition: familiarity with writing and written language from two aspects: written language as discourse style--the recognition that the kind of language used for writing is essentially different from the one used for speech; and written language as a notational system--the perception and growing command of the representational system that is used in the written modality. Linguistic literacy is viewed as a constituent of language knowledge characterized by the availability of multiple linguistic resources and by the ability to consciously access one's own linguistic knowledge and to view language from various perspectives. PMID- 12109380 TI - Literacy and development as discourse, cognition or as both? PMID- 12109381 TI - Developing linguistic literacy: perspectives from corpus linguistics and multi dimensional analysis. PMID- 12109383 TI - Lexical learning in school-age children, adolescents, and adults: a process where language and literacy converge. PMID- 12109382 TI - Sentence processing studies and linguistic literacy. PMID- 12109384 TI - Oral language, written language and language awareness. PMID- 12109385 TI - The foundations and development of metalinguistic knowledge. PMID- 12109386 TI - Sociocultural and cognitive constraints on literacy development. PMID- 12109388 TI - Rabies: dealing with dog bites. PMID- 12109387 TI - A crisis and its solution: setting up the UK clinical virology network. PMID- 12109389 TI - Tuberculosis and poverty in the ethnic minority population of West Yorkshire: an ecological study. AB - The rising incidence of TB in the UK in the last decade has been attributed mainly to the persistently high TB rates amongst ethnic minority groups. The epidemiology of TB in these groups is presently poorly understood. The aim of this ecological study was to investigate the relationship between TB and poverty in the South Asian ethnic groups. TB notification rates in South Asians residing in Kirklees (West Yorkshire) were correlated with eight indices of deprivation. Spearman's correlation coefficient analysis suggested a strong relationship between notification rates and material deprivation (r = 0.58 [p value = 0.009 for a two-tailed test]). A similar association was also observed for unemployment (r = 0.51 [p value = 0.02 for a two-tailed test]), but not for overcrowding. This study suggests a link between TB and deprivation in ethnic minority groups in the UK. PMID- 12109390 TI - Hepatitis B prevalence and risk factors for HBsAg carriage amongst Somali households in Liverpool. AB - The prevalence of hepatitis B core antibody (anti-HBc) and surface antigen (HBsAg) in the Somali population in Liverpool is described and groups who may benefit from vaccination are identified. A cross-sectional descriptive study was undertaken. A total of 439 subjects, aged between 10 months and 80 years, from 151 households, were screened for anti-HBc and HBsAg. One hundred ninety-four (44.2%) were children aged less than 15 years. Three hundred and nine (69%) of enrolled subjects were born in Somalia, 122 (27.2%) were born in the UK and 8 were born elsewhere. Of the study population, 5.7% were carriers of HBsAg, with the highest prevalence in adults aged 20 to 44 years (9.4%). A history of circumcision (RR 95% CI; 1.2, 1.1-1.5) was the most significant risk factor for HBsAg carriage, but was not significant on multivariate analysis. Prevalence of anti-HBc was 27.5%, and increased with age over the first four decades. Univariate analysis showed that a history of living in a refugee camp (RR 95% CI; 3.1, 1.7-5.7), receiving an injection in Somalia (2.1, 1.7-2.5), a history of circumcision or other surgical procedure in Somalia (1.4, 1.3-1.6) and being born in Somalia (1.3, 1.2-1.4) were significant risk factors for anti-HBc positivity. On multivariate analysis, only circumcision (OR 95% CI; 4.3, 1.8-10.3) and receiving an injection in Somalia (2.5, 1.5-4.4) remained significant. Seven of 80 (8.7%) children born in the UK and aged five years or less had evidence of exposure to hepatitis B, of whom only one had a close family member identified to be HBsAg seropositive. Previous infection with hepatitis B is common in this population. Horizontal transmission may be continuing at an early age within the UK, suggesting a population of at risk individuals who would benefit from surveillance and immunisation. Community circumcision is a risk factor for hepatitis B transmission and best practice should be followed when this procedure is undertaken. PMID- 12109391 TI - Re-emerging syphilis in the UK: a behavioural analysis of infected individuals. AB - In-depth interviews with 27 individuals infected with syphilis in a recent UK outbreak (out of a total of 58 diagnosed between May 1999 and August 2000 in three city hospitals) were carried out to examine behaviour and attitudes. Most (23/27) participants were homosexual men, seven of whom were HIV positive. Between them, the 23 gay men had 1,494 different contacts in the twelve months prior to their awareness of having syphilis, but only 10% of these contacts could be named. While oral sex (usually unprotected) was the most prevalent behaviour (median = 30 partners per year), only 39% perceived unprotected oral sex as a syphilis risk (c.f. 70% for anal sex). Many gay men (61%) used gamma hydroxybutyrate (GHB) during sex as an aphrodisiac. This syphilis-infected subset of the population had high levels of unprotected and anonymous sex, which brings into question the usefulness of contact tracing to control syphilis outbreaks. The majority of partners were casual oral sex partners. More awareness is urgently needed around syphilis symptoms and risks, and risks of using drugs to reduce sexual inhibitions. PMID- 12109392 TI - General outbreaks of infectious intestinal disease linked with red meat, England and Wales, 1992-1999. AB - Between 1992 and 1999, 1,426 foodborne general outbreaks of infectious intestinal disease (IID) were reported to the Public Health Laboratory Service (PHLS) Communicable Disease Surveillance Centre (CDSC). Sixteen percent were linked with the consumption of red meat. Over 5,000 people were affected, with 186 hospital admissions and nine deaths. Beef (34%) and pig meat (32%) were the most frequently implicated meat types, with lamb implicated in 11% of outbreaks. The organisms most frequently reported were Clostridium perfringens (43.4%) and salmonellas (34.3%). During the summer, outbreaks were mainly of Salmonella spp. and attributed to the consumption of pig meat. In December, outbreaks of C. perfringens linked with beef predominated. Most outbreaks occurred as a result of food cooked on commercial catering premises (46%). The highlight of this surveillance period is a fall in the number of outbreaks linked with foods containing red meat. This corresponds with a steady decline in red meat consumption over the last two decades, as well as a transient though marked decline in the purchase and consumption of red meat in the UK during the BSE crisis in the early to mid 1990s. As cited in the Pennington Report, further reducing the morbidity and mortality from red meat outbreaks means targeting meat production at various points along the food chain from abattoir and butchering, to cooking and holding of cooked food, especially on commercial catering premises. PMID- 12109393 TI - Improving the quality of communicable disease control: the example of meningococcal disease. AB - Improving the quality of health services is central to current attempts to reform the National Health Service. The main approaches to quality improvement in communicable disease control to date have been the development of practice guidelines and audit projects. Descriptions of the impact of quality initiatives in public health generally and communicable disease in particular have been limited, objective measures of quality are rare and few reports outline improvements that have been achieved in practice. We describe a project to develop a set of quality indicators for meningococcal disease control. A set of candidate indicators was developed and screened using standard approaches based on local consensus. We outline how they could be further tested and used to promote quality improvement. PMID- 12109394 TI - Second dose of MMR vaccine: health professionals' level of confidence in the vaccine and attitudes towards the second dose. AB - Public concerns have been raised about the safety of MMR vaccine following the publication of two studies linking the vaccine to inflammatory bowel disease and to a syndrome of Ileal-lymphoid-nodular hyperplasia, non-specific colitis and pervasive developmental disorder. Our study had two aims, to determine whether health professionals' confidence in MMR vaccine was affected and to assess professional knowledge and attitudes towards the second dose of MMR. In July 1998 we undertook a questionnaire survey of general practitioners, practice nurses and health visitors in an inner city area. A significant fall was found in professional confidence following the two publications and the subsequent media coverage (from 59.4% to 40.9%). Forty percent of respondents were unsure about the need for a second dose of MMR vaccine and more than one in ten professionals stated that it was not necessary. It is reasonable to assume that this reduced confidence in the safety of MMR and the professional uncertainty about the second dose have contributed to the observed decline in MMR uptake rates. More professional and public education appears to be needed. PMID- 12109395 TI - Investigation of postanginal sepsis and Lemierre's syndrome in the South West Peninsula. AB - We describe a retrospective case series of postanginal sepsis and Lemierre's syndrome (LS) identified from laboratory records of Fusobacterium necrophorum isolates and from clinical case note review. Some patients presented with sore throat, tonsillitis, quinsy or a septicaemic illness, whereas others presented with symptoms related to metastatic septic lesions with later recognition of the significance of the preceding sore throat. Patients with otitis media and mastoiditis are included in the study. The incidence of postanginal sepsis and LS appears to have increased over the study period (1994-99). The population of patients who had received antibiotics pre-admission has decreased in recent years. Attention is drawn to features which may assist in differentiating this condition from simple viral sore throats not requiring antibiotic therapy. A prospective study of the incidence of this rare but life-threatening condition mainly affecting young people is required in view of the more restricted use of antibiotic treatment for sore throat now recommended. PMID- 12109396 TI - Code of practice for funeral workers: managing infection risk and body bagging. AB - There is substantial variation in the advice given to funeral workers on handling bodies with infection risk. Inconsistent advice results in inappropriate practice. A model code of practice is presented that uses risk assessment in response to statutory and executive responsibilities to provide health and safety advice to funeral workers. The code of practice should increase compliance with safety requirements, avoid unnecessary bagging and allow bereaved families freer access to the deceased. PMID- 12109397 TI - Evaluation of the notification of hepatitis C risk to children who received unscreened blood or blood products. AB - Parents of children who received blood or blood products between 1984 and 1990 were notified about the potential risk of hepatitis C virus (HCV) infection. Data were collected about knowledge, attitudes and intended behaviours to determine the acceptability of the notification process. Demographic variables that may predict responses to notification were also recorded and analysed. Recipients were sent couriered letters explaining HCV risk, and the survey questionnaire. Sera were screened for HCV antibody and reactive samples confirmed with a recombinant immunoblot assay (RIBA). Four letter recipients were RIBA positive for a prevalence of 1.1% (4/358) in the notification group. Thirty-two percent of respondents did not know their child had been transfused and 58% did not know about the potential risk of HCV infection. Although 90% (165/185) felt the notification was valuable, 65% reported emotional distress (fear, worry, anger, very depressed). Responders were similar to non-responders except for HCV testing rate (76.2% v. 59.8%, p < 0.0002). Parents of children at risk of transfusion acquired HCV virus approved of notification programs, but experienced some emotional distress. Awareness of transfusion history or risk of HCV was not universal, indicating the need to address notification to individuals, rather than through public education campaigns alone. PMID- 12109398 TI - Microbiological examination of ready-to-eat burgers sampled anonymously at the point of sale in the United Kingdom. AB - During May and June 1999 a microbiological study of ready-to-eat burgers purchased anonymously from burger outlets (combined take-away and burger restaurants, take-away-only fixed premises, mobile vendors, temporary stalls and other burger outlets) was undertaken. The intention was to determine the microbiological quality of ready-to-eat burgers as purchased by customers of take away premises and to ascertain, where information was available, whether the Chief Medical Officer's advice on cooking burgers was being followed. Examination of 3,128 ready-to-eat burgers found that 2,868 (92%) were of acceptable quality and 260 (8%) were of unsatisfactory quality. Unsatisfactory results were mostly due to high aerobic colony counts (ACCs). Salmonella spp., Campylobacter spp. and Escherichia coli O157 were not detected in any of the samples examined. Acceptable microbiological quality of ready-to-eat burgers was associated with outlets, such as combined take-away and burger restaurants and in particular national franchise outlets, which had management food hygiene training and hazard analysis in place. Poor microbiological quality was associated with undercooking and local outlets as indicated by Local Authority Inspectors' Consumers at Risk scores. PMID- 12109399 TI - Outbreak of Salmonella indiana associated with egg mayonnaise sandwiches at an acute NHS hospital. AB - An outbreak of Salmonella indiana infection in December 2000 affected 17 staff, relatives and patients at an acute NHS Hospital in Swansea. Epidemiological investigation identified egg mayonnaise sandwiches as the vehicle of infection. It was not possible to definitively determine the source of the infection or how the prepared sandwiches became contaminated. The most likely explanation was a pasteurisation failure of a batch of the egg roll used to make these sandwiches. Sandwiches are the most frequently identified vehicle of infection in foodborne outbreaks of salmonella infection in hospitals in England and Wales. The process of sandwich preparation has inherent risks because it involves considerable handling of food, which is consumed without further cooking. Care is required in all stages of preparation including the sourcing of materials used to produce the sandwiches. NHS Trusts should review their Hazard Analysis Critical Control Point plans for sandwich production. PMID- 12109400 TI - Outbreak of Norwalk-like virus infection associated with salad provided in a restaurant. AB - Preliminary enquiries following the reporting of some cases of suspected food poisoning by members of the public revealed that they had all eaten at the same restaurant during the preceding weekend. Subsequent investigation, including a merged cohort study, identified 49 people with gastrointestinal symptoms, six of whom showed evidence of infection with Norwalk-like virus (NLV). Over a four day period all had eaten at the restaurant. Eating salad was strongly associated with infection. One of the chefs, who handled salad in the food preparation area and who had been absent from work with symptoms of gastrointestinal illness, had returned to work the day before the four-day period, reporting that he was 48 hours symptom free. He was subsequently shown to have been infected with NLV by reverse transcriptase polymerase chain reaction (RT-PCR) assay. The NLV belonged to the same genogroup as that infecting the cases who had eaten at the restaurant. PMID- 12109401 TI - Mumps outbreak among young people in Northern Ireland. AB - An outbreak of mumps occurred in the Mid-Ulster area of Northern Ireland between 1st November 1999 and 31st August 2000, with 729 cases notified. Salivary test results were obtained for 430 (59%) reported cases, of which 332 (77%) were positive for mumps IgM antibody. Three hundred and sixteen (95.2%) of these confirmed cases were in the age range 9 to 19. One hundred and eighty-four (55.4%) confirmed cases had received one dose of MMR (measles, mumps and rubella) vaccine, but only 3 (0.9%) confirmed cases had received 2 doses of MMR. The low incidence of mumps among children who had received two doses of MMR illustrates the importance of the second dose of MMR for full protection. PMID- 12109402 TI - Cluster of meningococcal disease in rugby match spectators. AB - Four adults among three unconnected groups of spectators at a rugby match developed invasive meningococcal disease four to five days after the match. Two died. All four cases were caused by serogroup C serotype 2a strains, genotype P1.5, P1.2. Although the route of transmission remains uncertain, the most likely explanation is that an asymptomatic carrier disseminated meningococci to at least four others in the course of an afternoon. Clusters among spectators at an event without other links between cases are very unusual. PMID- 12109403 TI - Continued role for HNIG in hepatitis A post-exposure prophylaxis. PMID- 12109404 TI - Flu from the grave. PMID- 12109405 TI - Effect of psychological climate in the microbiology laboratory on the quality of service. PMID- 12109406 TI - [Qualify quantity]. PMID- 12109407 TI - [Commentary on the occasion of, "Willingly remove M3's to benefit the breech in M2's"]. PMID- 12109408 TI - [Commentary on the occasion of, "Control photo of apex resection sometimes difficult to judge"]. PMID- 12109410 TI - An evaluation plan to assess the process and outcomes of a learner-centered training program for clinical research. AB - In 1994, the National Center for Research Resources' Committee on Addressing Career Paths for Clinical Research reported that insufficient training in research methods, inadequate mentoring, and inappropriate timing of training presented major barriers to the development of clinical researchers. The National Institutes of Health responded to the need for additional training programs by supporting 57 institutions through the Clinical Research Curriculum Award (K30) Program. The ability to assess the success of these programs depends on the nature and extent of their evaluation plans. Evaluation plans for clinical research training programs should include means of assessing both process and outcomes of a program's implementation in its formative and summative stages. This article describes the University of Wisconsin-Madison's Clinical Investigator Preparatory Program and an evaluation plan that incorporates process and outcome assessments based on a theoretical framework of adult and professional education. PMID- 12109409 TI - [Ultra-quick detoxification of opioid addicted patient: a supplement to the article fy Fontaine et al. (Encephale 2001; XXVII: 187-93)]. PMID- 12109411 TI - Mammography: consensus in sight? PMID- 12109412 TI - Europe against cancer: whats next? PMID- 12109413 TI - Use of the common gas outlet for the administration of supplemental oxygen during Caesarean section under regional anaesthesia. AB - A postal survey investigating the administration of supplemental oxygen to women undergoing Caesarean section under regional anaesthesia was sent to 262 lead consultant obstetric anaesthetists in the UK. Two hundred and fifteen (82) completed questionnaires were returned. In 139 units (65) supplemental oxygen was administered routinely to all Caesarean sections under regional techniques, while in 71 (33), supplemental oxygen was given only if the procedure is an emergency or if there was evidence of fetal or maternal compromise. In 196 units (91), the common gas outlet was used as the source of supplemental oxygen, with the standard anaesthetic breathing circuit disconnected in 194 (90) and the vaporisers left on the back bar in 191 (89). Critical incidents had occurred in 39 (18) of units using the common gas outlet as a source of supplemental oxygen and 63 (30) had experience of critical incidents with this practice in a non obstetric setting. We suggest that supplemental oxygen is more safely administered from a separate and dedicated source. PMID- 12109414 TI - Closed loop control of sedation for colonoscopy using the Bispectral Index. AB - Sixteen patients undergoing colonoscopy were sedated with propofol using a closed loop control system guided by the Bispectral Index (BIS). Propofol administration, via a target-controlled infusion, was controlled by a proportional-integral-differential control algorithm. The median (range) propofol target concentration during closed-loop control was 2.3 (1.7-3.6) microg.ml(-1). The performance characteristics of the system were excellent, with a median absolute performance error of 7 (1-15). Patients were drowsy yet rousable, with a median (range) BIS set-point of 80 (75-85). No patient became apnoeic, required airway support or became haemodynamically unstable whilst sedated. Eight patients moaned or moved during colonoscopy and four had recall. Median (range) time to full consciousness was 4 (2-20) min after the end of closed-loop control. Patient and surgeon satisfaction were high. We conclude that BIS may be a suitable control variable for closed-loop control of sedation with propofol. PMID- 12109415 TI - Patient feedback on the anaesthetist's performance during the pre-operative visit. AB - A questionnaire was devised from guidelines published by the Royal College of Anaesthetists and the Association of Anaesthetists of Great Britain and Ireland for the conduct of the pre-operative interview and conduct of anaesthesia. The responses to the questionnaire formed the basis of an accumulative record of patient feedback on individual anaesthetist's performance and used as one component of annual appraisal. The median "desired answer" and "overall dissatisfaction" percentages, and the "desired : undesired answer" ratio for consultants and non-training grade doctors from 835 patients at a large Acute District General Hospital were 92%, 0.6% and 12:1, respectively. PMID- 12109416 TI - The effect of dexamethasone upon patient-controlled analgesia-related nausea and vomiting. AB - Ninety female patients were enrolled in this randomised, double-blind, placebo controlled study to compare the anti-emetic effect of intravenous dexamethasone 8 mg with saline control in preventing patient-controlled analgesia-related nausea and vomiting following major orthopaedic surgery. The prophylactic administration of dexamethasone 8 mg significantly reduced the overall incidence of patient controlled analgesia-related nausea and vomiting (p<0.001) and the need for rescue anti-emetics (p<0.01). Furthermore, patients who received dexamethasone showed a higher incidence of complete responses (no vomiting or need for rescue anti-emetic for a 24-h postoperative period) than those who received saline (p<0.05). We conclude that dexamethasone 8 mg may be valuable for preventing patient-controlled analgesia-related nausea and vomiting in women undergoing major orthopaedic surgery. PMID- 12109417 TI - Will competency assessment improve the training and skills of the trainee anaesthetist? PMID- 12109418 TI - Anions and the anaesthetist 2. PMID- 12109419 TI - Anions and the anaesthetist 1. PMID- 12109420 TI - When is a white cell count not a white cell count? PMID- 12109421 TI - Chronic pain in the UK. PMID- 12109423 TI - The Trachlight compared to laryngeal mask airway assisted intubation. PMID- 12109422 TI - Cardiac output determination using vascular compliance. PMID- 12109424 TI - Oxytocin during Caesarean section. PMID- 12109425 TI - Dose of oxytocin after Caesarean section. PMID- 12109426 TI - Low-dose remifentanil infusions for major spinal surgery. PMID- 12109427 TI - Midazolam--an anti-emetic? PMID- 12109428 TI - Alternative coupling systems for regional anaesthetic equipment. PMID- 12109429 TI - Pulmonary oedema and race. PMID- 12109430 TI - Factors affecting intubation conditions. PMID- 12109431 TI - The Pro-seal laryngeal mask airway. PMID- 12109432 TI - Gum elastic bougies. PMID- 12109433 TI - Postoperative pain relief in the day surgery unit. PMID- 12109434 TI - An uncommon cause of pacemaker malfunction. PMID- 12109435 TI - Use of the "T-bag" oxygen enhancement device to improve oxygenation during single lung ventilation. PMID- 12109437 TI - On predicting difficult intubation. PMID- 12109436 TI - Inappropriate drug labelling. PMID- 12109438 TI - Tracheostomy pilot tube failure. PMID- 12109439 TI - Further evidence for the rising incidence of childhood Type 1 diabetes in Kuwait. AB - AIMS: To provide age-gender standardized incidence rate, temporal trend and seasonal variation of Type 1 diabetes in Kuwaiti children aged < or = 14 years. METHODS: Data were prospectively collected over a period of 6 years (1992-1997) according to the DiaMond Project protocol using the capture-recapture method of ascertainment. RESULTS: Data ascertainment varied between 90% and 96%. The incidence rate of Type 1 diabetes was 20.1 per 100,000 children 0-14 years (95% confidence interval (CI) 18.0-22.1); age-standardized incidence rate 20.9 (95% CI 18.8-23.0). The incidence rate among boys, 21.1 per 100,000 (95% CI 18.1-24.1) was slightly higher than that among girls, 19.0 per 100,000 (95% CI 16.1-21.8). The age-standardized incidence rate was 21.9 (95% CI 18.9-24.8) in boys, and 19.9 (95 CI 17.1-22.8) in girls. Incidence rates increased with age in both sexes (boys chi(2) for linear trend = 13.5, P < 0.001; and for girls chi(2) = 27.8, P < 0.0001). There was a significant trend towards increase in overall incidence during the 6-year period (chi(2) = 6.210, P = 0.013), and in age group 5-9 (chi(2) = 10.8, P = 0.001). Seasonality was demonstrated overall, in boys and girls (P < 0.001). CONCLUSION: The incidence of Type 1 diabetes in Kuwait is high compared with the neighbouring Arab countries, and it appears to be increasing as in many European populations. PMID- 12109440 TI - Activation of human T cells with NK cell markers by staphylococcal enterotoxin A via IL-12 but not via IL-18. AB - We have reported recently that mouse liver NK cells and NK1 x 1+ T cells were activated by bacterial superantigens via the IL-12 production from Kupffer cells. In the present study, we examined the effect of staphyloccoccal enterotoxin A (SEA) on human T cells with NK cell markers, CD56 or CD57 (NK-type T cells). After stimulating peripheral blood mononuclear cells (PBMC) with SEA, PBMC produced a large amount of IFN- and acquired a potent antitumour cytotoxicity. The in vitro depletion of either CD56+ TCR NK cells, CD56+ T cells or 57+ T cells from PBMC significantly inhibited the IFN- production from PBMC. When purified NK type T cells, NK cells and regular T cells were cultured with monocytes and SEA they all produced IFN-, while the IFN- amounts produced by both NK-type T cells were greater than those produced by NK cells. NK cells as well as CD56+ T cells showed cytotoxicity against NK-sensitive K562 cells, whereas both NK-type T cells showed a more potent cytotoxicity against NK-resistant Raji cells than did NK cells. The IFN- production from each population as well as from whole PBMC was greatly inhibited by anti-IL-12 antibody but not by anti-IL-18 antibody. The antitumour cytotoxicity of whole PBMC was also significantly inhibited by anti-IL 12 antibody while the SEA-induced proliferation of PBMC was not affected by anti IL-12 antibody. Furthermore, SEA-activated NK-type T cells as well as NK cells showed cytotoxicities against vascular endothelial cells. Our findings suggest that human NK-type T cells are thus involved in bacterial superantigen-induced immune response. PMID- 12109441 TI - Macrophage migration inhibitory factor activates antigen-presenting dendritic cells and induces inflammatory cytokines in ulcerative colitis. AB - The level of macrophage migration inhibitory factor (MIF) and the functions of dendritic cells (DC) are up-regulated in the peripheral blood, and the numbers of MIF-expressing cells and mature DC are increased at the colonic mucosa from patients with ulcerative colitis (UC). However, a functional relationship between MIF and DC, and the role of MIF in the pathogenesis of UC, are not clear. In this study, we showed that a pure population of peripheral blood DC is a new and still unknown source of MIF. DC from UC patients produced significantly higher levels of MIF (17 x 5 +/- 9 x 8 ng/ml, n = 10) compared with patients with Crohns disease (CD) (4 x 6 +/- 2 x 5 ng/ml, n = 5, P< 0 x 01) and control subjects (5 x 0 +/- 2 x 6 ng/ml, n = 10, P< 0 x 01). A double immunofluorescence study revealed the expression of MIF by CD83-positive mature DC at the colonic mucosa from UC patients. Blood DC treated with high amounts of MIF (500 ng/ml) showed a significantly higher stimulatory capacity (43287 +/- 5998 CPM, n = 5) in an allogenic mixed leucocyte reaction compared with untreated DC (27528 +/- 8823 CPM, n = 5, P< 0 x 05). Study of intracellular cytokine expression showed that MIF induced significant levels of interleukin (IL)-1 and IL-8 in monocytes and DC from UC and CD patients. These results showing the capacity of MIF to induce increased functional capacity of DC, and to produce IL-1 and IL-8 from monocytes and DC, indicate a role of MIF in the induction and/or perpetuation of the inflammatory environment in UC. PMID- 12109442 TI - [Guideline for proper use of antineoplastic agents. Cancer of the digestive system--malignant cancers (stomach, colonic, and pancreatic cancers)]. PMID- 12109443 TI - [Postoperative adjuvant therapy of gastric and colonic cancers]. PMID- 12109444 TI - [Guidelines for proper use of antineoplastic agents. Breast cancer]. PMID- 12109445 TI - [Guidelines for proper use of antineoplastic agents. Gynecologic cancer]. PMID- 12109446 TI - [Guideline for proper use of antineoplastic agents. Urologic cancer]. PMID- 12109447 TI - [Guidelines of proper use of antineoplastic agents. Brain tumors]. PMID- 12109448 TI - [Guidelines for proper use of antineoplastic agents. Skin cancer]. PMID- 12109449 TI - [Guideline for proper use of antineoplastic agents. Malignant tumors of bones and soft tissue]. PMID- 12109450 TI - [Guideline for correct use of antineoplastic agents (draft). General theory]. PMID- 12109451 TI - [Guideline for proper use of antineoplastic agents. Hematopoietic tumors]. PMID- 12109453 TI - [Question of the month. Retinoid-induced osteoporosis?]. PMID- 12109452 TI - [Guideline for chemotherapy of malignant lung cancer]. PMID- 12109454 TI - [Meeting of the Tuberculosis Network. Clinical and molecular epidemiology of tuberculosis, with vaccinal perspectives (Bobigny, Feb. 7 2002)]. PMID- 12109455 TI - Evolution of student assessment in McMaster University's MD Programme. AB - In response to the competitive examination culture that pervaded medical education in the 1940s and 1950s the founders of McMaster University's new MD Programme created an assessment system based on group functioning within the tutorial. While the tutorial has served the educational process well, 30 years of experience has highlighted its deficiencies as an assessment tool. This paper describes the accumulation of evidence that led to the awareness of the weakness of tutorial assessment and the attempts to provide reliable assessment by the reintroduction of examinations, but in novel formats which would not alter the goals of the curriculum. PMID- 12109456 TI - Students' evolution of a learning method: a comparison between problem-based learning and more traditional methods in a specialist university training programme in psychotherapy. AB - Two educational groups of psychotherapy students were compared regarding their educational training in a three-year programme. One of these groups attended a traditional psychotherapy programme based on conventional lectures (1993-96), while the other attended a PBL programme (1994-97). In an initial study, the students in both groups answered a questionnaire composed to evaluate relevant sub-areas of the training programme. The traditionally trained group reported a significantly higher level of knowledge than the PBL group. On the other hand, the PBL group reported more positive general opinions of their education than did the traditional group. Based on the initial questionnaire, a follow-up questionnaire was designed to investigate the professional situation and the former students' view of their education two to three years later. Statistically, there were no significant differences between the two groups in their evaluations of the practical use. PMID- 12109457 TI - Participants' perception of impact of a workshop on their preventive practices. AB - A 90-minute interactive workshop, offered to small groups on request, was developed to help physicians include evidence-based preventive interventions in their practices. Between 25 September 1996 and 10 December 1997, 593 family physicians throughout the Province of Quebec (Canada) participated in one of the 40 workshops presented in all the regions of Quebec. Almost all participants (98%) completed the self-administered questionnaire. Their opinion of the achievement of three workshop objectives were evaluated using a seven-point Likert scale (-3 to +3)as their perception of the direct impact of the workshop on their practice. The workshop objectives were reached to a high degree: 2.1 (sd 0.90) for prescribing a proper check-up for adults; 1.83(sd 1.02) for explaining to the patient the reasons motivating his/her choice to include or exclude certain tests; 2.09 (sd 0.93)for using concrete and useful tool facilitating the integration of preventive measures in his/her professional practice. Female physicians and those under 40 perceived that the objectives were reached to a greater degree. Participants indicate their intention to modify their practice according to the clinical practice guidelines presented in the workshop. PMID- 12109458 TI - Problem-based learning, work demands and psychological distress in pre registration house officers: a preliminary study. AB - There has been little work examining the longer term occupational and psychological outcomes of problem-based learn-ing (PBL). The authors aimed to investigate the relationship between psychological distress, occupational factors and experience of PBL in the first cohort of UK pre-registration house officers to be trained exclusively using problem-based methods. Questionnaires were sent to al pre-registration house officers working in the three teaching hospitals in the Manchester area. Measures included demographic items, hours of actual work and the GHQ-12 as a measure of psychological distress, as well as validated Likert scales of job satisfaction, perceived work demands, support from superior,and experience and attitudes towards PBL. The response rate was 87% (66/76). Almost one-third of respondents (20/66) worked more than 72 hours a week. The occupational and psychological measures were significantly interrelated. The multiple regression analysis revealed that work demands and job satisfaction together accounted for over 45% of the variance in psychological distress. The other demographic, occupational and PBL-related factors did not make an independent contribution to distress. Scores were similar for respondents with high and low levels of exposure to PBL, and similar to scores obtained from a previous cohort of house officers graduating from a traditional course. These findings should be interpreted cautiously because of the small size of the current study and the setting. It is possible that as staff and students grow accustomed to new styles of teaching, and work intensity for junior doctors reduces, the effect of PBL will become more evident. Further research is needed examining the occupational and psychological outcomes of medical students taught by problem-based methods indifferent settings. PMID- 12109459 TI - JAMA patient page. Endocarditis. PMID- 12109460 TI - [Are antioxidants effective in the prevention of coronary events?]. PMID- 12109461 TI - [Vasovagal syncope and beta-blockers]. PMID- 12109463 TI - Genetic discrimination--an overblown fear? PMID- 12109464 TI - The need for comprehensive and consistent treatment of the nitrogen cycle in nitrogen cycling and mass balance studies: I. Terrestrial nitrogen cycle. AB - A review of conceptual models that scientists use to characterize the nitrogen (N) cycle and to conduct N mass balance studies at global, regional and local scales is presented. Large uncertainties in processes and process rates make it difficult to conduct precise N mass balances and the dominant conceptual model has changed in recent decades. An earlier conceptual model recognized explicitly that human activities, especially agriculture, have both depleted terrestrial N and increased the fixation of atmospheric N in biologically available forms. The current conceptual model does not include adequate treatment of the depletion of the terrestrial N reservoir, the resulting transfer of N to the hydrosphere and atmosphere, or the cycling of terrestrial N below the plow layer. Thus, it delivers an unrealistically limited view of human influences on the N cycle. It is recommended that a comprehensive and consistent treatment of terrestrial N cycling be developed to better facilitate scientific explanation of historical N related environmental changes and more closely balance N budgets on a range of geographical and temporal scales. Improved N-cycle models will provide an improved scientific basis for answering important resource management and policy questions. PMID- 12109465 TI - On the correlation between wind speed, coarse aerosol concentration and the electrical state in the ground atmospheric layer in semi-arid areas. AB - Our general work has been dedicated to the problems connected with the influence of short-term changes of the atmospheric state on some medically unexplained human responses. What are the meteorological stimuli eliciting the transition between health and disease? There are many reports on the impact of humidity, winds or pollution on feelings of disease or discomfort. We suppose that the main influencing climatic factor, which could excite multiform biological reactions, is atmospheric electricity, E, whose parameters depend strongly on the content of the air column and on the meteorological conditions in this volume, especially along its ground path. A clear connection between wind speed Vw and coarse aerosol concentration N(T) with electric field intensity E exists only for limited intervals of Vw, and N(T). In these ranges, the mean correlation coefficients p(E; Vw) and p(E; N(T)) are usually up to 0.45. In quiet weather, rho is significantly larger than in the equinox seasons or in transferred weather situations. Strong winds or a considerably increased pollution level lead to fast and deep fading of E and even to principal changes in the form of its diurnal distribution from what is usual for stable atmosphere conditions, while weak air streams or a clear atmosphere have almost no influence on electric state variations. PMID- 12109466 TI - Antifouling paint booster biocides in UK coastal waters: inputs, occurrence and environmental fate. AB - This study considered the inputs of antifouling paint booster biocides into the aquatic environment directly from painted hulls and high pressure hosing operations, the occurrence of booster biocides in marinas, harbours and docks, and the influence of degradation and water-sediment partition on their environmental fate. Irgarol 1051, the Irgarol 1051 degradation product GS26575, diuron, and the diuron degradation products 1-(3-chlorophenyl)-3,1-dimethylurea (CPDU), 1-(3,4-dichlorophenyl)-3-methylurea (DCPMU) and 1-(3,4 dichlorophenyl)urea (DCPU) were all detected at measurable concentrations in surface waters. Irgarol 1051, GS26575 and diuron were also detected in bottom sediments. A preliminary study of biocide input during both normal use and foreshore hull hosing showed that hosing may be a significant point source input and also be a cause for future concern since much of this input is in the form of paint particles. Field based measurements and laboratory experiments showed that Irgarol 1051 and diuron persist in the water column, due to a low affinity to partition onto sedimentary material and high resistance to degradation. Other biocides such as chlorothalonil, dichlofluanid, and Sea-Nine 211 were all found to be rapidly removed from the water column and be less persistent. PMID- 12109467 TI - Selenium in soils, spermatophytes and bryophytes around a Zn-Pb smelter, New South Wales, Australia. AB - Selenium concentrations and its spatial distribution in soils, spermatophytes and bryophytes (mosses) around the Cockle Creek Zn-Pb smelter, New South Wales were studied from May to November 2000. Selenium was determined by ICP-MS on soils digested in nitric and hydrochloric acid (HNO3:HCl = 3:1), and plant samples digested in distilled concentrated nitric acid. At distances greater than 3 km, selenium in soils, spermatophytes and bryophytes converge to uniform values, which are considered to represent the background value. Mean selenium in soils around the smelter is two times higher than the background value. Mean concentration of selenium in plants around the smelter is three times greater than that for background plants. Selenium from 'in site' bryophyte is twice that found in the background bryophyte. The transfer coefficients of selenium between plants and soils are low, especially at a distance less than 1 km from the smelter. However, mosses show the reverse trend. This study indicates that the Zn Pb smelter is one of the anthropogenic point sources of selenium pollution in the Lake Macquarie district. PMID- 12109468 TI - Changes in blood lead of a recreational shooter. AB - We have measured the concentration and isotopic composition of lead in blood over a 15-month period for a subject who undertook recreational shooting in outdoor and indoor firing ranges on an irregular basis. We have also measured the isotopic composition in cast lead, Cu-jacketed and Teflon-coated bullets, propellant and primer from which he assembled the cartridges. Blood lead concentration increased from 3.2 to 6.7 microg/dl with use of dominantly cast lead bullets in the outdoor range. In two intervals when no firing was undertaken for 3-4 months, the blood lead concentrations either decreased towards a baseline value in the case where only Cu-jacketed bullets were fired or remained elevated when dominantly cast lead bullets were fired. The propellants contained <2 ppm Pb and contribute negligibly to blood lead. The isotopic composition of the primer used for all bullets was consistent with a source from the US. The bullets were of different materials and made in Australia and the US, with lead from sources of different geological age and hence different isotopic signatures. Variations in blood lead concentration and isotopic composition appear most strongly influenced by the bullets. Although more expensive, the use of Cu-jacketed bullets, non-lead primers and well-ventilated indoor firing ranges would lessen the health impacts of recreational shooting. PMID- 12109469 TI - Can flooded organic matter from sediments predict mercury concentrations in zooplankton of a perturbed lake? AB - Mercury (Hg) and the composition of organic matter (OM) in sediments and large zooplankton (> 190 microm) have been analysed in a perturbed lake (Lake 154) in northern Quebec, Canada. We investigated whether methylmercury concentrations ([MeHg]) in flooded sediments can predict [MeHg] in zooplankton. [MeHg] at the uppermost sediment layers located immediately downstream of an inundated system (LA-40) are 10 times higher than at other sediment stations. Zooplankton organisms sampled downstream of LA-40 contain higher [MeHg] (183+/-50 ng g(-1) dry wt.) than those of neighbouring natural lakes (66+/-37 ng g(-1) dry wt.). OM in sediment and zooplankton samples has been characterised applying atomic carbon/nitrogen ratios (C/N) and stable carbon isotopic ratios (delta13C). While C/ N ratios in zooplankton from Lake 154 (5.6+/-0.4) do not differ from samples of natural lakes (5.7+/-0.1), delta13C signatures are more depleted at stations close to the discharge of LA-40 (-33+/-0.4/1000) with respect to zooplankton from unperturbed lakes (-30.6+/-0.7/1000). Results of this tracer study suggest that the biochemical composition of OM in flooded sediments did not modify the biochemical composition of zooplankton and can therefore not predict [MeHg] in zooplankton. PMID- 12109470 TI - First flush analysis of urban storm runoff. AB - Stormwater runoff was monitored on 13 separate urban watersheds, which were chosen to represent distinct types of residential and industrial development, along with various watershed characteristics. A total of 38 storm events were monitored to investigate the first flush phenomenon. The first flush phenomenon may be defined as the initial period of stormwater runoff during which the concentration of pollutants is substantially higher than during later stages. The magnitude of the first flush phenomenon, and if it actually occurs, was calculated using a method of data analysis which results in determining the 'event mean concentration' (EMC). The magnitude of the first flush phenomenon was found to be greater for some pollutants (e.g. suspended solids from residential areas) and less for others (e.g. chemical oxygen demand from industrial areas). No correlation was observed between the first flush phenomenon and the antecedent dry weather period, however, the first flush phenomenon was greater for smaller watershed areas. PMID- 12109471 TI - Sedimentation rates and pollution history of a dried lake: Al-Oteibeh Lake. AB - Sediment accumulation rates as well as the distribution of selected elements in a dried Syrian lake (Al-Oteibeh Lake), near Damascus City, are reported. Five core samples from different locations of the lake were collected, and four major elements (Fe, K, Mg and Na) and six trace metals (Co, Ni, Cr, Pb, Zn, U and Cu) were analyzed. Sedimentation rates were determined applying the 210Pb dating method and found to vary between 0.100 and 0.793 cm year(-1). The results showed that the constant flux constant sedimentation rate (CF:CS) simple dating model is applicable for dating recent dried sediment and recording the past historical pollution of the last 100 years. However, the method was found to be only applicable for dating trace and major elements, which cannot be leached to deeper layers by rainwater. In addition, the obtained records can be used to verify the date of water level declining. PMID- 12109472 TI - Transport of fallout radiocesium in the soil by bioturbation: a random walk model and application to a forest soil with a high abundance of earthworms. AB - It is well known that bioturbation can contribute significantly to the vertical transport of fallout radionuclides in grassland soils. To examine this effect also for a forest soil, activity-depth profiles of Chernobyl-derived 134Cs from a limed plot (soil, hapludalf under spruce) with a high abundance of earthworms (Lumbricus rubellus) in the Olu horizon (thickness=3.5 cm) were evaluated and compared with the corresponding depth profiles from an adjacent control plot. For this purpose, a random-walk based transport model was developed, which considers (i) the presence of an initial activity-depth distribution, (ii) the deposition history of radiocesium at the soil surface, (iii) individual diffusion/dispersion coefficients and convection rates for the different soil horizons, and (iv) mixing by bioturbation within one soil horizon. With this model, the observed 134Cs-depth distribution at the control site (no bioturbation) and at the limed site could be simulated quite satisfactorily. It is shown that the observed, substantial long-term enrichment of 134Cs in the bioturbation horizon can be modeled by an exceptionally effective diffusion process, combined with a partial reflection of the randomly moving particles at the two borders of the bioturbation zone. The present model predicts significantly longer residence times of radiocesium in the organic soil layer of the forest soil than obtained from a first-order compartment model, which does not consider bioturbation explicitly. PMID- 12109473 TI - The effect of environmental conditions and electrical charge on the weighing accuracy of different filter materials. AB - Different filter materials and electrical charge elimination methods were used to investigate the weighing accuracy of filter papers under different environmental conditions. The results show that the standard deviations (S.D.) of weight data for glass fiber and MCE filters were substantial without environmental control, whether or not the electrical charge eliminators were used. Values of 0.157 and 0.349 mg were determined for glass and MCE filters, respectively. The accuracy of weighing was substantially improved and the S.D. was reduced to 0.01 and 0.09 mg for glass fiber and MCE filters, respectively, after applying the environmental control conditions. For PVC and Teflon filters, the accuracy of weighing was good, even in the uncontrolled environmental conditions, whether or not the electrical charge eliminators were used. The S.D. values of weighing data of PVC and Teflon filters were 0.007 and 0.011 mg, respectively. PMID- 12109474 TI - Antimicrobial multiresistance in bacteria isolated from freshwater Chilean salmon farms. AB - The intensive use of antimicrobial agents, mainly oxytetracycline, to prevent and control bacterial pathologies in Chilean salmon culture is a frequent practice. A total of 103 gram-negative oxytetracycline-resistant bacteria recovered from various sources of 4 Chilean freshwater salmon farms were identified and investigated for their susceptibility patterns to various antibacterial agents, by using an agar disk diffusion method. Antibacterial resistance patterns of isolates were not correlated with bacterial species or strain source. A high number of bacteria resistant to amoxicillin, ampicillin. erythromycin, and furazolidone, as well as an important frequency of bacterial resistance to florfenicol, chloramphenicol, cefotaxime and trimethoprim-sulfamethoxazole was found. On the contrary, the proportion of bacteria resistant to gentamicin, kanamycin, flumequine and enrofloxacin was rather low. Resistant microflora showed a high taxonomic variability and mainly consisted of non-fermenting bacteria (77.7%). These strains mainly belonged to the species Pseudomonas fluorescens (29), Aeromonas hydrophila (10), Stenotrophomonas maltophilia (6), isolated from salmon fingerlings, and Acinetobacter lwoffii (5) isolated from pelletized feed. The occurrence of simultaneous resistance to various antibacterials was frequent. We observe a high frequency of bacteria resistant to 6-10 antibacterials (74 strains), and antibiotic resistance index (ARI) values ranging from 0.38 to 0.48 for the four salmon farms studied. These results suggest that Chilean salmon farms might play a role as reservoirs of antibacterial multiresistant bacteria, thus prompting the necessity for a more restrictive attitude towards the intensive use of antibacterials in salmon farming. PMID- 12109475 TI - Bioaerosol exposure during refuse collection: results of field studies in the real-life situation. AB - To determine the bioaerosol exposure of refuse collectors, field measurements were performed under real working conditions within the framework of a research project. Influencing variables such as different types of refuse, community structure, collection interval and season were taken into account. Overall, 1612 samples were taken in towns of Westfalia, Germany. With workplace levels on a scale of 10(3) to less than 10(4) CFU/m3 for the loader, the results show a surprisingly low total fungi concentration in comparison with earlier studies. Total bacteria concentrations, in contrast, were largely on a scale of 10(4) CFU/m3, with 10(5) CFU/m3 being registered sporadically, especially in apartment block districts. Endotoxin levels were high especially in the summer months, occasionally reaching values of more than 50 EU/m3, whereas they were normally below 10 EU/m3 in autumn and winter. Inside the cab, the exposure level for the entire spectrum was at least one power of ten lower. The factors believed to account primarily for the low total fungi concentration were workplace hygiene, the prevailing 1-week collection interval, and the low in-process exposure time resulting from the effective deployment of automatic lifting devices. In contrast, the type of refuse was not found to have a significant influence. PMID- 12109476 TI - Distribution of selected anthropogenic radionuclides (137Cs, 238Pu, (239,240)Pu and 241Am) in marine sediments with emphasis on the Spitsbergen-Bear Island area. AB - Measurements of anthropogenic radionuclides in marine sediments can provide good estimates of past and present radioactive contamination of the marine environment. The Barents Sea is one of the world's richest ocean areas, and it is therefore of major interest for Norwegian fisheries to document the levels of radioactive contamination of this and adjacent ocean areas. In this study, concentrations of 137Cs, 238Pu, (239,240)Pu and 241Am were measured in marine sediments collected mainly in the Spitsbergen-Bear Island region. Additional samples collected in the deep Norwegian Sea and near the Greenland ice-edge have been included for comparison. The highest radionuclide levels were found near Spitsbergen, reaching over 50% higher than in adjacent areas. Determinations of the mineral content and particle size distribution indicate a relation between the content of clay minerals and radionuclide levels. The present study confirms earlier observations of elevated levels of 137Cs in sediments in the Spitsbergen Bear Island area and documents elevated levels of 238Pu, (239,240)Pu and 241Am in these sediments. Using an exponential curve fit based on determinations of unsupported 210Pb in sediment cores, sedimentation rates of 0.28, 0.05 and 0.26 cm year(-1) were calculated at locations south of Spitsbergen, near the Greenland ice-edge and in the deep Norwegian Sea, respectively. PMID- 12109477 TI - Forest fire indicators and mercury deposition in an intense land use change region in the Brazilian Amazon (Alta Floresta, MT). AB - Black carbon, charcoal and mercury fluxes were measured from sediment cores taken in an artificial water dam in an intense land use change area in the Alta Floresta district in the Brazilian Amazon, in order to characterize the differences in the evolution of human occupation patterns in the region during the last 18 years. A positive correlation between the black carbon and charcoal particle fluxes and the evolution of the Brazilian gross domestic production (GDP) was observed. Mercury fluxes showed a positive correlation with gold production and exhibited a distinct evolution pattern when compared to in relation to the forest fires indicators and Brazilian GDP. The fluxes of forest fires markers showed an increase in deforestation activities in the region after 1993. Mercury deposition showed a substantial decrease after 1994. The patterns of distribution in both forest fires tracers and gold mining tracers indicate substitution of the regional economic model. It also marked different antropogenic impact type in the ecosystem. PMID- 12109478 TI - Analysis and implications of aircraft disinsectants. AB - Aircraft disinsection is required by various countries. In-flight spraying with a 2% phenothrin aerosol exposes passengers and crew directly. Residual spaying uses a permethrin emulsions in the absence of passengers and crew and results in dermal and oral exposures. Exposed passengers and crew often complain of, skin rashes, respiratory problems, tingling and numbness in fingertips and lips and burning eyes. A number of formulations were analyzed for their constituents using GLC-Mass. spec. Volatile organic compounds (VOCs) were found in all aerosol preparations including, ethyl benzene and xylene isomers along with phenothrin. Residual sprays contained, cis-, and trans-, permethrins, palmidrol, and occasionally naphthalene. Headspace analysis found methylene chloride and hexene derivatives but not the active ingredients. The known synergistic effects between organophosphates and pyrethrins, based on carboxyesterases inhibition, can be expected in the presence of Tricresylphosphates (TCPs), constituents found in jet engine oils and in some hydraulic fluids. During oil seal failure, the presence of TCP in the ventilation air could explain the increased sensitivity of some crew members and passengers to disinsectants. PMID- 12109479 TI - Mercury levels in lichens from different host trees around a chlor-alkali plant in New Brunswick, Canada. AB - Mercury concentrations were determined in the epiphytic lichen Hypogymnia physodes along five transects starting from a chlor-alkali plant located at Dalhousie, New Brunswick, a landfill site and a nearby electricity generating station. Lichen samples were collected from white birch (Betula papyrifera) and spruce (Picea sp.) or balsam fir (Abies balsamea). Average lichen background mercury values were 0.088+/-0.005 microg/g from white birch and 0.148+/-0.046 microg/g from spruce trees, with a detection limit of 0.05 microg/g. The chlor alkali plant and a power plant were identified, respectively, as a major source and a minor source of elevated mercury levels in lichens. At 125 m north-west of the New Brunswick Power plant, 0.28 microg/g Hg were found in Hypogymnia physodes from spruce trees, while at 250 m west (downwind) of the chlor-alkali plant, 3.66 microg/g of mercury were determined. High values, 0.98 microg/g in lichens from spruce trees and 0.79 microg/g in lichen samples from white birch were also measured at 125 m south of the chlor-alkali plant and decreased exponentially with distance. The sphere of influence of the chlor-alkali plant with respect to mercury deposition was estimated to extend 2.4-3.4 km from the plant. The mercury concentrations in Hypogymnia physodes collected from white birch were significantly lower than the concentrations in the same lichen from spruce trees in areas with elevated levels of mercury, but not in areas with low mercury levels. The magnitude of this difference dropped with distance from the source. PMID- 12109480 TI - Platinum levels in natural and urban soils from Rome and Latium (Italy): significance for pollution by automobile catalytic converter. AB - Platinum concentrations in topsoil samples collected in 1992 (48) and in 2001 (16) from the urban area of Rome have been determined by ICP-MS. Concentrations in 47 soil samples collected in 1992 from natural sites of Latium (an area around Rome) have been determined for a first assessment of natural background levels. The Pt concentrations in Rome urban soils collected in 1992 range from 0.8 to 6.3 ng/g (mean = 3.8 +/- 1.0) overlapping the concentration range of natural soils from Latium (mean = 3.1 +/- 2.1 ng/g). No significant correlation has generally been found between Pt contents in the 'natural' soils and related bedrock or major pedogenetic parameters. These results suggest that there is no evidence of Pt pollution in Rome urban soils at that time, because the massive use of the automobile catalytic converter has only just started. Higher (up to six times more) Pt concentrations, than those measured in the 1992 samples, have been measured, in some cases, in Rome urban soils collected in 2001, suggesting a possible start of Pt accumulation because of the large-scale use in the last decade of automobile catalytic converters. At the same time, a clear decrease of lead levels in Rome urban soils with respect to the levels measured in 1992 has been observed, paralleling the decreasing number of lead gasoline-fuelled cars. Here we present one of the first systematic studies for defining background levels of Pt in Italian natural soils, thus allowing for monitoring, in the future, should any possible Pt pollution caused by the use of automobile catalytic converter, especially in urban soils, occur. PMID- 12109481 TI - 129I/127I ratio measurements in bovine thyroids from the North Cotentin area (France). AB - Iodine-129 is routinely released in the gaseous and liquid low-level radioactive effluents of nuclear spent fuel reprocessing plants. Environmental impact assessment of these discharges are performed based on monitoring samples of different types of natural indicators. Thyroid is considered as a relevant organ to monitor radioactive iodine isotopes. In this study, bovine thyroids were collected in herds located in the Cotentin area under the influence of authorized low level gaseous effluents of the La Hague nuclear fuel-reprocessing plant. For low level 129I/127I ratios, characteristic of environmental samples (10(-12) to 10(-7)), RNAA or SMA that includes important radiochemical preparation steps are used. This paper demonstrates the interest of direct gamma-X spectrometry to measure 129I activities, associated to INAA to measure 127I to attain higher ratios levels (10(-6) to 10(-4)). This study shows the interest of monitoring bovine thyroids considered as sentinel organs to characterize the dispersion in space and time of 129I discharged in low level radioactive gaseous effluents by the La Hague reprocessing plant. PMID- 12109482 TI - Detection of estrogenic activity in sediment-associated compounds using in vitro reporter gene assays. AB - Sediments may be the ultimate sink for persistent (xeno-)estrogenic compounds released into the aquatic environment. Sediment-associated estrogenic potency was measured with an estrogen receptor-mediated luciferase reporter gene (ER-CALUX) assay and compared with a recombinant yeast screen. The ER-CALUX assay was more sensitive to 17beta-estradiol (E2) than the recombinant yeast screen, with an EC50 of 6 pM E2 compared to 100 pM in the yeast screen. Yeast cells were unable to distinguish the anti-estrogens ICI 182,780 and (4-hydroxy)tamoxifen, which were agonistic in the yeast. Acetone-soluble fractions of hexane/acetone extracts of sediments showed higher estrogenic potency than hexane-soluble extracts in the ER-CALUX assay. Sediments obtained from industrialized areas such as the Port of Rotterdam showed the highest estrogenic potency of the 12 marine sediments tested (up to 40 pmol estradiol equivalents per gram sediment). The estrogenic activity of individual chemicals that can be found in sediments including: alkylphenol ethoxylates and carboxylates; phthalates; and pesticides, was tested. Increasing sidechain length of various nonylphenol ethoxylates resulted in decreased estrogenic activity. Of the phthalates tested, butylbenzylphthalate was the most estrogenic, though with a potency approximately 100,000 times less than E2. The organochlorine herbicides atrazine and simazine failed to induce reporter gene activity. As metabolic activation may be required to induce estrogenic activity, a metabolic transformation step was added to the ER-CALUX assay using incubation of compounds with liver microsomes obtained from PCB-treated rats. Results indicate that metabolites of E2, NP and bisphenol A were less active than the parent compounds, while metabolites of methoxychlor were more estrogenic following microsomal incubations. PMID- 12109483 TI - Historical changes in lead concentrations in tree-rings of sycamore, oak and Scots pine in north-west England. AB - Lead concentrations in tree rings of sycamore (Acer pseudoplatanus L.), oak (Quercus robur L.) and Scots pine (Pinus sylvestris L.) sampled at a parkland in north-west England were measured in wood formed since the mid-1800s. Concentrations of Pb in Scots pine and oak peaked in wood formed between 1900 and 1940, most likely because of Pb accumulation in heartwood, indicating that oak and Scots pine are unsuitable for monitoring temporal changes in Pb deposition at the study site. In contrast, Pb concentrations in sycamore, a species that has similar heartwood and sapwood chemistry, were relatively constant in wood formed between the mid-1800s and 1950. Lead concentrations decreased steadily in sycamore tree rings formed after the 1950s, and decreased more abruptly in wood formed after 1985. This sharp decrease in wood Pb cannot be due to decreases in soil Pb concentration. Stable Pb isotope analysis was used to further investigate Pb patterns in sycamore wood. Excess 206Pb/207Pb ratios in tree-rings of sycamore were relatively constant, approximately 1.17, in wood formed prior to the 1930s, but decreased steadily thereafter reaching a minimum value of approximately 1.16 in wood formed between 1975 and 1985 after which time 206Pb/207Pb ratios increased. This pattern is consistent with changes in Pb isotope ratios measured in peat, sediment and aerosol samples in the UK. However, the magnitude of the decrease in 206Pb/207Pb (largely due to gasoline Pb) is considerably lower than in other studies and our estimates indicate that less than 20% of the total Pb in sycamore wood measured since the mid-1800s is derived from gasoline emissions. A more likely explanation for the pattern of Pb observed in sycamore tree rings is that soil Pb accumulates within rings of the diffuse porous wood over a number of years. Such uptake patterns would result in lower Pb concentrations in the outer (more recently formed) tree rings, which coincide with recent reductions in Pb deposition in the UK. Overall, this study indicates that tree ring chemistry is unsuitable for monitoring historical changes in Pb deposition at the study site. PMID- 12109484 TI - Distributions of rare earths and heavy metals in field-grown maize after application of rare earth-containing fertilizer. AB - Rare earths are widely applied in Chinese agriculture to improve crop nutrition through the use of fertilizers, yet little is known of their accumulation in field-grown crops. We have studied the distribution of 16 rare earths (Sc, Y and 14 lanthanide elements) in field-grown maize and the concentration of heavy metals in the grains after application of rare earth-containing fertilizer. When maize entered the vigorous vegetation growth stage (e.g. early stem-elongation stage), rare earth-containing fertilizer was applied to the soil with irrigation water. At 10 days after application of the rare earths, significantly dose dependent accumulative effects of individual rare earth concentrations in the roots and the plant tops of maize were observed, with the exception of Sc and Lu. At the level of 2 kg rare earths ha(-1), accumulative concentrations of most light rare earths (e.g. La, Ce, Pr and Nd) and Gd in the plant tops were much larger than those in the control. Concentrations of individual rare earths in a field-grown maize after application of rare earths decreased in the order of root>>leaf>stem>grain. During the maize growth period, selective accumulation of individual rare earths (e.g. La, Ce) in the roots seemed to be in dynamic equilibrium, and the distribution of these elements in the plant tops was variable. At a dosage of less than 10 kg rare earths ha(-1), no apparent accumulative concentrations of individual rare earths appeared in the maize grains. Under the experimental conditions, application of rare earth-containing fertilizer did not induce an increase in the concentrations of heavy metals in the grains. We conclude that the present dosage of rare earths (<0.23 kg ha(-1) year(-1)) currently applied in China can hardly affect the safety of maize grains in arable soil, even over a long period. PMID- 12109485 TI - RE: Shen P, Cai F, Nowicki A, Vincent J, Reynolds EC (2001). Remineralization of enamel subsurface lesions by sugar-free chewing gum containing phosphopeptide amorphous calcium phosphate. J Dent Res 80:2066-2070. PMID- 12109486 TI - Michael Phelps: nuclear medicine innovator awarded Cassen Prize. PMID- 12109487 TI - CPT code recommendations for nuclear medicine therapy. PMID- 12109488 TI - Observed classroom behavior of children with ADHD: relationship to gender and comorbidity. AB - Examined hypothesized gender and comorbidity differences in the observed classroom behavior of children with attention deficit hyperactivity disorder (ADHD). The behavior of 403 boys and 99 girls with ADHD, ages 7-10, was compared (a) to observed, sex-specific classroom behavior norms, (b) by sex, and (c) by comorbid subgroups. Boys and girls with ADHD deviated significantly from classroom norms on 15/16 and 13/16 categories, respectively. Compared to comparison girls, girls with ADHD had relatively high rates of verbal aggression to children. Boys with ADHD engaged in more rule-breaking and externalizing behaviors than did girls with ADHD, but the sexes did not differ on more "neutral," unobtrusive behaviors. The sex differences are consistent with notions of why girls with ADHD are identified and referred later than boys. Contrary to hypothesis, the presence of comorbid anxiety disorder (ANX) was not associated with behavioral suppression; yet, as hypothesized, children with a comorbid disruptive behavior disorder (DBD) had higher rates of rule-breaking, and impulsive and aggressive behavior, than did children with ADHD alone and those with ADHD+ANX. Elevated rates of ADHD behaviors were also observed in children with comorbid DBD, indicating that these behaviors are truly present and suggesting that reports of higher ADHD ratings in this subgroup are not simply a consequence of negative halo effects and rater biases. PMID- 12109490 TI - Ischemic stroke and active migraine. PMID- 12109491 TI - Does the weather influence stroke incidence? PMID- 12109489 TI - Mother-child relationships of children with ADHD: the role of maternal depressive symptoms and depression-related distortions. AB - We investigated the Depression-->Distortion hypothesis by examining the effects of maternal depressive symptoms on cross-informant discrepancies in reports of child behavior problems and several measures of parent-child relationship. The sample included ninety-six 6 to 10-year-old children diagnosed with ADHD-Combined Type, and their mothers, who provided baseline data before participating in a randomized clinical trial. Measures incorporated child characteristics, self reports of maternal depressive symptoms, parenting practices, and laboratory mother-child interactions. Elevations in maternal depressive symptoms were associated with maternal reports of negative parenting style but not with observed laboratory interactions. Mothers' levels of depressive symptoms predicted negative biases in their reports of their child's ADHD symptoms, general behavior problems, and their own negative parenting style. Whereas levels of depressive symptoms did not predict observed parenting behaviors, maternal distortions did predict problemaTic parent-child interactions. Exploratory analyses showed a marginally significant mediation effect of the relationship between maternal depressive symptomatology and reports of negative parenting by depressive distortions. We discuss implications of linkages between depressive symptoms in mothers, depression-related distortions, and mother-child relationships for research and intervention in developmental psychopathology. PMID- 12109492 TI - Intra-arterial thrombolytic therapy for acute stroke: the debate continues. PMID- 12109493 TI - [Biological warfare]. PMID- 12109494 TI - [Medicine on the banks of the Arno]. PMID- 12109495 TI - [Medical marvels of the Tantra manuscripts]. PMID- 12109497 TI - Drama and discovery: the story of histoplasmosis. PMID- 12109496 TI - [The remarkable odyssey of the potato]. PMID- 12109498 TI - Scientific method discourses in the construction of 'EMF' science: interests, resources and rhetoric in submissions to a public inquiry. AB - Since the late 1970s, there has been considerable debate surrounding the question of whether or not exposures to non-ionizing radiation and electric and magnetic fields (EMF), produced by powerlines and electrical and telecommunications technologies, are harmful to health. Whilst there has been some recent evidence of regulatory fatigue, and attempts to enforce closure, the EMF debate nevertheless still continues. This paper will explore the role played by competing images of scientific method in the argumentative strategies used by two of the main protagonists in an Australian public inquiry (held in 1990-91) which investigated the EMF issue: 'Inquiry into Community Needs and High Voltage (132kv and above) Transmission Line Development', the so-called Gibbs Inquiry. Apart from documenting some of the epistemologically intricate features of the EMF controversy, the following discussion will also consider the way scientific method discourses can contribute to enhancing the durability of knowledge claims in legal and regulatory settings. PMID- 12109499 TI - Style and substance in psychology: are influential articles more readable than less influential ones? AB - The results from four studies are reported separately to test the idea that influential articles in psychology will be more readable than less influential ones. This idea is upheld when the papers involved are ones that have been highly rated by fellow colleagues (Studies 1 and 2) but it is not supported when the papers involved are highly-cited journal articles (Studies 3 and 4). PMID- 12109500 TI - Generations do not write books: a sociological autobiography of my medical humanities career. PMID- 12109501 TI - The place of the humanities in an academic health science center: the double edged sword of a required ethics curriculum. PMID- 12109502 TI - The medical humanities as an elephant seen by blind men. PMID- 12109503 TI - A journey in the borderlands of medicine and the humanities. PMID- 12109504 TI - Welfare state and infant mortality. AB - This article seeks to understand the effects of welfare-state spending on infant mortality rates. Infant mortality was chosen for its importance as a social indicator and its putative sensitivity to state action over a short time span. Country fixed-effects models are used to determine that public health spending does have a significant impact in lowering infant mortality rates, net of other factors, such as economic development, and that this effect is cumulative over a five-year time span. A net effect of health spending is also found, even when controlling for the level of spending in the year after which the outcome is measured (to account for spurious effects or reverse causation). State spending effects infant mortality both through social mechanisms and through medical ones. This article also shows that the impact of state spending may vary by the institutional structure of the welfare state. Finally, this study tests for structural breaks in the relationship between health spending and infant mortality and finds none over this time period. PMID- 12109505 TI - Do we owe our intelligence to a predatory past? AB - As I am not a neuroscientist, it is an unexpected pleasure for me to contribute a lecture to the James Arthur series on the Evolution of the Human Brain. By contrast, I am an African naturalist, and what I have to say will be very much from the perspective of African cave taphonomy, a recent and rather macabre discipline that uses fossils in an attempt to reconstruct the circumstances of death of the animals involved, as well as to gain insights into their behavior and the paleoecology of the time. The lecture's focus will be on predation, to which I am largely indebted to Professor Raymond Dart, who provoked me into devoting many years of my life developing the principles of cave taphonomy and interpreting the bone accumulations in southern African caves where hominid fossils have been found. PMID- 12109506 TI - Searching for the magic bullet: veterinary experiences with the first antibiotic. PMID- 12109507 TI - Animal therapy over the ages: 7. acetum, alcohol, allium, and aloes. PMID- 12109508 TI - Same bed, different dreams: veterinary medicine and the American Animal Protection Movement in the 19th century. PMID- 12109509 TI - Henry Stockton Lewis, Sr. (1858-1922): an early African-American veterinarian. PMID- 12109510 TI - Mark L. Morris, Sr., DVM pioneer and innovator. PMID- 12109511 TI - [Vaccination for smallpox in Madrid in the last third of the 19th century]. PMID- 12109512 TI - [The Bowlby revolution in psychopathology]. PMID- 12109513 TI - New perspectives for tumor pathology provided by comparative genomic hybridization. AB - Comparative genomic hybridization (CGH) allows rapid screening for DNA copy number gains and losses across the entire genome. CGH analyses have revealed a number of common aberrations and characteristics associated with specific tumor cells or pathogeneses. Recurrent aberrations suggest that tumor-related gene(s) may be located in such regions. Furthermore, some specific aberrations may serve as novel diagnostic features. Quantitative chromosomal analyses based on CGH have also provided stimulating information associated with chromosomal instability, genetic pathways, telomerase activity status, and DNA ploidy and have yielded valuable insights into tumor pathology. This review focuses on scientific advances facilitated by this technique. PMID- 12109514 TI - Control of cell proliferation kinetics of tumor in neoadjuvant chemotherapy for advanced oral squamous cell carcinoma and its prognostic implications. AB - BACKGROUND: Few attempts have so far been made at studies of the cell proliferation kinetics of the tumor in neoadjuvant chemotherapy for head and neck cancer. We examined the effects of neoadjuvant chemotherapy for advanced oral squamous cell carcinoma in terms of the cell proliferation kinetics of the tumor, and attempted to correlate them with patients' survival. METHODS: Fifty-two patients with advanced oral squamous cell carcinoma who received neoadjuvant chemotherapy followed by surgery participated in this study. Cellular DNA content and mitotic index (MI) in tissue samples were measured before and after chemotherapy, using a cell image analyzer. RESULTS: A decrease in both mean DNA content (mean DNA) and MI (left-shift type of change in cell growth kinetics), indicating an accumulation of cancer cells in the G0-G1 phase, was found in 25 patients. An increase in mean DNA with decreased MI (right-shift type), which was found in 13 patients, appeared to be correlated with an accumulation of cancer cells in the S-G2 phase. Neither of these two types of change, which were considered to be a favorable effect, were found in 14 patients (ineffective type). Excellent survival rates were obtained in patients who showed favorable changes in cell growth kinetics (79% for patients with left-shift type and 92% for patients with right-shift type), whereas the survival rate for patients with the ineffective type was extremely poor (14%). The type of change in cell proliferation kinetics was a powerful independent prognostic indicator. CONCLUSION: Analysis of cell growth kinetics appears to be useful not only as a diagnostic tool to predict patient outcome but also as a means to infer the chemotherapeutic effects in oral squamous cell carcinoma. PMID- 12109516 TI - Immunohistochemical demonstration of apoptosis-regulated proteins, Bcl-2 and Bax, in resected non-small-cell lung cancers. AB - OBJECTIVE: Bcl-2 and Bax proteins regulate apoptosis independently, cooperatively, or both in vitro. The purpose of this study was to clarify the association between their expression with spontaneous apoptosis and various clinicopathologic features in patients with non-small-cell lung cancer (NSCLC). METHODS: Bcl-2 protein, Bax protein, and spontaneous apoptosis were evaluated retrospectively in 70 resected specimens from NSCLC patients. Immunohistochemical (IHC) tests were used to assess the expression of Bcl-2 and Bax in the samples. The apoptotic index (AI) was also measured in these samples by the terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL) method. We then evaluated the clinicopathologic features of all 70 samples and their relationships with malignancy. RESULTS: Bcl-2 overexpression was significantly associated with male sex and squamous cell carcinoma (SCC). Bax overexpression was not associated with any clinicopathologic features. AI was significantly associated with SCC and negative nodal metastasis. No clear associations were found among Bcl-2 expression, Bax expression, and AI. Bcl-2/Bax ratios were not associated significantly with AI. Bcl-2, Bax, Bcl-2/Bax ratio, and the grading of AI did not have prognostic values. CONCLUSION: Bcl-2 overexpression was significantly associated with SCC. Spontaneous apoptosis was significantly associated with SCC and with negative nodal metastasis. Apoptosis regulated proteins, Bcl-2 and Bax, and AI lack significant associations with each other and prognostic values in patients with resected NSCLC. PMID- 12109515 TI - Fuzzy logic-based tumor-marker profiles improved sensitivity in the diagnosis of lung cancer. AB - BACKGROUND: The aim of this study was to improve the diagnostic efficiency of tumor markers in the diagnosis of lung cancer, by the mathematical evaluation of a tumor marker profile employing fuzzy logic modelling. METHODS: A panel of four tumor markers, i.e., carcinoembryonic antigen (CEA), cytokeratin 19 antibody (CYFRA 21-1), neuron-specific enolase (NSE), squamous cell carcinoma-related antigen (SCC) and, additionally, C-reactive protein (CRP), was measured in 175 newly diagnosed lung cancer patients with different histological types and stages. Results were compared with those in 120 control subjects, including 27 with chronic obstructive pulmonary diseases (COPD), 65 with pneumoconiosis, and 11 persons with acute inflammatory lung diseases. A classificator was developed using a fuzzy-logic rule-based system. RESULTS: Application of the fuzzy-logic rule-based system to the tumor marker values of CYFRA 21-1, NSE, and CRP yielded an increase in sensitivity of approximately 20%, i.e., 92%, compared with that of the best single marker, CYFRA 21-1(sensitivity, 72%). The corresponding specificity was 95%. The fuzzy classificator significantly improved the sensitivity of the tumor marker panel in stages I and IIIa for non-small-cell lung cancer, as well as in "limited disease" status for small-cell lung cancer. Also, the diagnosis of other stages of lung cancer was enhanced. CONCLUSION: Fuzzy-logic analysis was proven to be more powerful than the measurement of single markers alone or combinations using multiple logistic regression analysis of all markers. Therefore, fuzzy logic offers a promising diagnostic tool to improve tumor marker efficiency. PMID- 12109517 TI - Histological features of hypovascular or avascular renal cell carcinoma: the experience at four university hospitals. AB - BACKGROUND: We aimed to evaluate the pathological features of hypovascular or avascular renal cell carcinoma (RCC), by the retrospective review of the histological features that coincide with hypovascular or avascular RCC. METHODS: Seven hundred and ninety-one patients who underwent both preoperative angiography and nephrectomy were examined. Of these patients, the 126 patients (15.9%) who showed hypovascular or avascular angiographic features were selected. Patients with hemorrhage or cyst(s), or with at least 50% necrosis or more in the tumor, and those with renal tumor metastatic to the kidney were excluded. The criteria proposed by the World Health Organization (1998) were adopted for the histological classification. RESULTS: Papillary RCC was the most frequently observed hypovascular or avascular renal tumor (44 cases; 34.9%). The vascularity differed among the variants, i.e., some cases had a basophilic and solid variant with avascular features, while the remaining cases had wide stromal organization showing hypovascular features. The second most frequently observed hyporascular or avascular RCC was chromophobe cell carcinoma (35 cases; 27.8%). No difference in vascularity was detected between variants, except for 2 cases with sarcomatoid changes (avascular features). The third most frequently observed hypovascular or avascular RCC was cyst-associated RCC (29 cases; 23%). All of the 7 RCCs originating in a cyst showed avascular features, and the remaining 22 cystic RCCs showed hypovascular features. The remaining hypovascular or avascular RCCs were cases of clear cell carcinoma accompanied by sarcomatoid changes (8 cases; 6.3%), spindle cell carcinoma (5 cases; 4.0%), and collecting-duct carcinoma (5 cases; 4.0%). CONCLUSION: Hypovascular or avascular RCC can be categorized as non-clear cell carcinoma and some clear cell carcinoma accompanied by sarcomatoid changes. PMID- 12109518 TI - BCG effects on telomerase activity in bladder cancer cell lines. AB - BACKGROUND: Although intravesical bacillus Calmette-Guerin (BCG) therapy is very effective in the treatment of and prophylaxis against superficial bladder cancer, its exact mechanism of action is not clear. In this study, the effect of BCG on telomerase activity was examined in the T24 and J82 bladder cancer cell lines, the ACHN human renal cell carcinoma cell line, and the PC-3 human prostate cancer cell line. METHODS: T24 and J82 cells were cocultured with BCG for 5 days. Telomerase activity was measured by telomeric repeat amplification protocol. Cell cycle phase was determined by FACS analysis. RESULTS: Telomerase activity in all cell lines provided high absorbance. Telomerase activity in BCG-treated T24 and J82 cells was significantly decreased when compared with that in the nontreated cells. On the other hand, telomerase activity did not change in ACHN and PC-3 cells after they had been cocultured with BCG. In BCG-treated T24 cells and J82 cells, apoptotic cells were markedly increased compared with those in the nontreated cells. CONCLUSION: These results suggest that the reduction of telomerase activity is related to the mechanism of BCG effects. Possible mechanisms to be considered are either that BCG inhibits telomerase first, or that it induces apoptosis and decreases telomerase activity as a result of this induction. PMID- 12109519 TI - Antitumor and antivascular effects of AC-7700, a combretastatin A-4 derivative, against rat liver cancer. AB - BACKGROUND: Unlike the many chemotherapeutic agents that do not effectively stop blood flow or induce necrosis in hepatocellular carcinoma, AC-7700 has been shown to inhibit tubulin polymerization and selectively stop tumor blood flow. The aim of this study was to elucidate the antivascular and antitumor effects of AC-7700 on rat hepatoma. METHODS: AH-130 cells, a rat hepatoma cell line, were solidified and implanted into the liver of Donryu rats. Vascularity of the liver tumor was directly identified by in-vivo fluorescence microscopy from 0 to 60 min after the injection of 10 mg/kg AC-7700. To observe the antivascular effect of AC-7700, the vascular density of the tumor was measured and assessed as the ratio of preinjection to postinjection values. The antitumor effects were evaluated with histopathologic findings and analysis of animal survival. RESULTS: In-vivo microscopic observation showed that tumor perfusion diminished within 30 min after AC-7700 administration. Vascular density in the AC-7700 group was significantly less than that in the control group at 60 min (AC-7700, 26.3 +/- 16.4%; control, 88.5 +/- 9.2%; P < 0.001). After AC-7700 injection, marked necrosis of tumor cells was observed histologically, and tumor area was decreased significantly (AC-7700, 11.5 +/- 15.4 mm2; control, 43.5 +/- 18.3 mm2; P < 0.05). The survival rate (50%) of the AC-7700 group animals was better than that of the control group (0%; P < 0.01). CONCLUSION: Markedly decreased tumor perfusion was induced by AC-7700 within 30 min, and this decrease may have contributed to the tumor necrosis and favorable outcome in the treatment group. AC-7700 appears to be a promising agent for the treatment of hepatocellular carcinoma. PMID- 12109520 TI - Phase I studies of nogitecan hydrochloride for Japanese. AB - BACKGROUND: SmithKline Beecham synthesized camptothecin analogs and identified nogitecan hydrochloride (topotecan) with a broad spectrum of antitumor activity and less toxicity than camptothecin. Because preclinical and overseas clinical data indicated the antitumor effect of nogitecan hydrochloride with a 5-day repeat-dose schedule, we carried out phase I studies in Japan to determine the maximum tolerated dose (MTD), pharmacokinetics, and antitumor effect of nogitecan hydrochloride. METHODS: Phase I studies of nogitecan hydrochloride given by single and 5-day repeat dosing were carried out in patients with various solid tumors at 15 medical institutions in Japan. Pharmacokinetic evaluations were performed for both single and 5-day repeated dosing. RESULTS: The dose-limiting factor (DLF) was reversible leucopenia, and the maximum tolerated dose (MTD) was higher than 22.5 mg/m2 in the single-dose study. In the 5-day repeat-dose study, the DLF was also reversible leucopenia, and the MTD was estimated to be 1.5 mg/m2 per day. The plasma concentration of nogitecan hydrochloride increased with increasing dose, and the half-life after single dosing ranged from 3 to 5h. There was no evidence of accumulation or delayed excretion during 5-day repeat dosing. CONCLUSION: Based on these results and the finding that there were responders among patients treated at 1.5 mg/m2 per day by 5-day repeat dosing in overseas studies, 5-day repeat dosing of 1.2mg/m2 per day, one dose level lower than the MTD, was selected for phase II studies in Japan. PMID- 12109521 TI - Two pedigrees of hereditary prostate cancer. AB - We encountered two pedigrees of hereditary prostate cancer. In one family, the father and his two sons had prostate cancer, and in the other family, three brothers developed prostate cancer. The mean age of these six individuals at the first examination was 65.3 years. Two individuals had stage B disease; three individuals, stage D disease; and one individual, disease of unknown stage. Histopathologically, two, one, and three individuals had well-, moderately, and poorly differentiated adenocarcinoma, respectively. As of September 28,2000, five of the six individuals were still alive. In a search of the literature, these were found to be the seventh and eighth pedigrees in Japan that met the criteria of hereditary prostate cancer proposed by Carter and colleagues in 1993. PMID- 12109522 TI - Complete regression of esophageal cancer with concomitant liver metastasis achieved by concurrent chemoradiation therapy. AB - We report a patient with esophageal cancer with concomitant liver metastasis in whom complete response was achieved by chemoradiation therapy. A 66-year-old man was diagnosed as having stage IVB esophageal cancer with synchronous metastasis in the liver and cardiac lymph node, and concurrent chemoradiation therapy was started. The chemotherapy, consisting of 5-fluorouracil (300 mg/body per day, continuous infusion) and low-dose cisplatin (5 mg/body per day on 1-5 days every week), was performed for 7 weeks. In addition, radiation therapy (2 Gy/day on 1-5 days every week) was employed for both the local and the metastatic lesions, along with the chemotherapy. Throughout the course of this therapy, the patient did not experience severe toxicity, and this chemoradiation therapy resulted in complete regression of both the local and the metastatic diseases. Subsequently, he was followed-up as an outpatient without any maintenance therapy, and he has been free of disease for 38 months after completion of the therapy. This concurrent chemoradiation therapy may be effective for esophageal cancer even with visceral metastasis. PMID- 12109524 TI - Efficacy of induction chemoradiotherapy in thymic cancer: report of a successful case and review of the literature. AB - Thymic cancer is a rare tumor, the optimal treatment of which remains controversial. The efficacy of induction therapy in thymic cancer is unclear. A 51-year-old man was diagnosed as having a poorly differentiated carcinoma of the thymus with lymph node metastasis (stage IVb according to the Masaoka staging system), through an echo-guided biopsy. The patient was administrered cisplatin (CDDP) combined with paclitaxel once per week for 4 weeks, under concurrent local radiation. Once a partial response was achieved, the residual tumor was completely resected, and bilateral mediastinal lymph nodes were dissected. Histopathological examination of the resected specimen showed no evidence of viable malignant cells. At present, 15 months after surgery, the patient is doing well and shows no signs of recurrence. This case demonstrates that induction chemoradiotherapy with CDDP and paclitaxel may be well tolerated and useful for patients with advanced thymic cancer. PMID- 12109523 TI - Complete remission of uterine endometrial cancer with multiple lung metastases treated by paclitaxel and carboplatin. AB - Endometrial cancer is believed to have a better prognosis than cervical cancer. However, this is not necessarily true for cases beyond International Federation of Gynecology and Obstetrics (FIGO) stage III, and advanced endometrial cancer with distant metastases in particular has a poor prognosis. Moreover, there is no established therapy for advanced endometrial cancer. Recently, we treated two patients with endometrial cancer with multiple lung metastases (FIGO stage IVb). Both patients had massive uncontrollable genital bleeding and eventually progressed to anemia. The imminent severe bleeding was considered to be a major reason for exacerbation of their general condition. Therefore, hysterectomy was performed as a counter-measure to improve their general condition. In their postoperative course, the two patients successfully underwent T-J chemotherapy [paclitaxel: 210 m/m2 over 3h; carboplatin: area under the curve (AUC) 5]. Six courses of the regimen were given every 3-4 weeks. Multiple lung shadows in chest X-P and computed tomography (CT) were reduced in number and size after two courses of T-J chemotherapy. The multiple lung metastases either disappeared or just remained as scars after six courses. There has been no evidence of recurrence for 28 months in one patient and 7 months in the other patient. PMID- 12109525 TI - Predictors of cat allergen (Fel d 1) in house dust of German homes with/without cats. AB - BACKGROUND: Exposure to cat allergen is a major risk factor for sensitization and the development of asthma in many parts of the world. The study was designed to examine the levels of cat allergen (Fel d 1) in homes of two German cities and to detennine predictors of Fel d 1 exposure. METHODS: We collected dust samples from 405 randomly selected homes in Hamburg (n = 201) and Erfurt (n = 204). In each apartment dust samples were taken from living room floor (LR), bedroom floor (BR), and mattress surface (MA) using vacuum sampling and analyzed by two-site monoclonal antibody ELISA. Environmental variables were assessed by questionnaire to obtain information on factors supposed to have an impact on the Fel d 1 levels in house dust. The effects of possible predictors of Fel d 1 were assessed by multiple linear regression models. RESULTS: Fel d 1 was present in 98% of the homes (395/405), ranging from 0.015 to 7.278 microg/g dust (GM 0.486 microg/g). The levels were similar in floors (GM for LR 0.575 microg/g and BR 0.469 microg/g) and in MA (GM 0.424 microg/g). Higher allergen concentrations (> 2 microg/g) were detected in 99% of the homes keeping cats, in 55% of homes that had had a cat during the last year, and in 28% of the homes without a cat. Fel d 1 levels were significantly higher in homes with a cat (628-682-fold, MA, floor) and in homes in which a cat had previously lived (11-12-fold, floor, MA) than in homes that never had a cat. Furthermore, after controlling for possible other confounders, Fel d 1 levels were significantly higher in summer and in homes with low frequency of cleaning and low ventilation rate. CONCLUSION: It could be confirmed that keeping a cat has the highest impact on Fel d 1 concentration. Besides, a continuous influx of Fel d 1 in homes without cat, as a result of direct or indirect cat contact outside the home, is likely. High frequencies of dusting and ventilation might reduce cat allergens in homes with and without cats. PMID- 12109526 TI - The mechanisms, causes, and treatment of anaphylaxis. AB - Anaphylaxis is a severe life-threatening systemic reaction that offers many challenges to the clinician. The incidence of anaphylaxis is significant in the general population and an important cause of morbidity and mortality. While the most common causes of anaphylaxis include drugs, foods, and venoms, other important etiologies must be considered. The etiology of anaphylaxis is classically based on IgE mediated hypersensitivity but multiple mechanisms may be involved. The clinical presentation of anaphylaxis may be extremely variable with a broad differential diagnosis which will be outlined. Although the diagnosis of anaphylaxis can many times be based on a careful history and physical examination, there are laboratory and skin tests which may be helpful in establishing a diagnosis in some cases. The cornerstone of treatment of anaphylaxis remains epinephrine. Other supportive therapies will be discussed. PMID- 12109527 TI - Conjunctival provocation tests in the diagnosis of Anisakis simplex hypersensitivity. AB - INTRODUCTION: The prevalence of positive skin tests to Anisakis simplex is high compared with the low incidence of true gastroallergic Anisakiasis. There is little information about the prevalence of positive conjunctival tests in A. simplex-sensitized individuals. This study assesses the predictive value of conjunctival provocation test in two groups of A. simplex-sensitized subjects and negative controls. METHODS: Group A consisted of 28 individuals with a clinical history strongly suggestive of gastroallergic anisakiasis and group B, of 32 individuals without such history. The 60 individuals had positive skin tests and in vitro specific IgE determinations to A. simplex. Skin and conjunctival provocation tests were performed with an extract of A. simplex at a concentration of 1 mg of lyophilized material per ml. RESULTS: Conjunctival provocation tests were positive in 21/28 (75%) of group A patients and in 10/32 (31%) of group B patients. This difference was statistically significant (p < 0.015); the odds ratio was 6.6 (2.11-20.5; p < 0.05). The diagnostic value of the conjunctival provocation test was expressed by the following statistical indicators: sensitivity 75% (55.8%-88.6%), false positive 25% (11.4%-45.2%), specificity 68.7% (49.9%-83.2%), false negative 31.2% (16.7%-50%), positive predictive value 67.7% (48.5%-82.7%), negative predictive value 75.9% (56.1%-89%), Yuden index 1.43. There were no statistical differences related to age, sex, or atopic status in patients with a positive or negative conjunctival challenge. Total and specific IgE levels to A. simplex were significantly greater (p < 0.05 and p < 0.001, respectively) in the group of patients with a positive challenge. CONCLUSIONS: The results demonstrate that positive conjunctival provocation tests and high specific and total IgE levels are more prevalent in patients with a clinical history strongly suggestive of gastroallergic anisakiasis. Despite the significant differences obtained in both groups, this test has a limited diagnostic value to clinically discriminate patients with a history consistent with gastroallergic anisakiasis. PMID- 12109528 TI - Extracts of Anacardium occidentale (cashew) pollen in patients with allergic bronchial asthma. AB - Allergic reactions to the pollen of trees is among the most prevalent allergic sensitivities. The cashew tree grows in abundance in the northeast region of the Brazil, mainly in Fortaleza city, in state of the Ceara. It flowers once a year between August and October. This is the first study conducted to establish the possible role of the cashew pollen extract in causing skin test reactivity in patients with allergic asthma. A stock solution of pollen extract was prepared with the standard weight/volume method for intradermal skin tests and for the protein content of the extract, estimated with the use of Folin phenol reagent and a spectrophotometer. Ten nonallergic volunteers and 80 subjects with allergic asthma, as documented by previous positive skin test reactions to various pollens, were studied. All of the 80 patients (100%) had positive test reactions (grade III and grade IV reactions). None of the control subjects (n = 10) had positive responses to the intradermal tests. This study provided us with knowledge of an additional pollen extract of the Anacardium occidentale, which could provoke skin test reactivities in asthmatic individuals from the northeastern area of Brazil. The results suggest a relationship between the period of flowering of the cashew tree and the increased number of allergic asthma cases. PMID- 12109529 TI - Clinical experience with specific immunotherapy to horse dander. AB - OBJECTIVE: This open clinical study was designed to investigate the occurrence of adverse reactions of specific immunotherapy (SIT) with horse dander and to recognize signs of efficacy regarding horse-induced cutaneous and respiratory symptoms. METHODS: Twenty-four patients attending our Outpatient Clinic in Huelva (Spain) with horse-induced allergy were selected for receiving a standardized extract of horse dander (Alutard SQ). RESULTS: Local and systemic reactions (five adverse reactions in four patients) were observed during the induction phase of treatment when administering the doses with the highest allergen concentration. Systemic side effects were mild, one immediate case consisting of wheezing and prompt response to treatment and one mild conjunctival hyperemia. Doses were reduced in all cases, and good tolerance to SIT was maintained. The sensitivity of patients to horse dander as assessed by deliberate natural exposure to horse decreased significantly when patients reached maintenance dose: Conjunctivitis symptoms were reduced in all patients, rhinitic symptoms in 93%, asthmatic symptoms in 90%, and cutaneous symptoms in 87% of the patients. Subjective assessment of a patient's allergic disease on a visual analog scale (VAS) showed an improvement. Ninety-five percent of the patients were highly satisfied with the treatment (efficacy, suitability, and troubles). CONCLUSION: Specific immunotherapy with standardized horse-allergen extract is a safe treatment with the doses of allergen extract administered in the present study. In addition, it seems to have beneficial effects in patients with moderate-severe rhinoconjunctivitis and/or asthma. PMID- 12109530 TI - International Study of Asthma and Allergies in Childhood (ISAAC): validation of the written questionnaire (eczema component) and prevalence of atopic eczema among Brazilian children. AB - Although Hanifin and Rajka's criteria have been used for the diagnosis of atopic eczema (AE), there is no instrument destined for epidemiological studies on AE that actually uses them. Written questionnaires (WQ) have generally been used, but when translated into another language they must be validated. The Intemational Study of Asthma and Allergies in Childhood (ISAAC) WQ was previously validated in a comprehensive study, but its validation in Brazil had not been done. Our objective was to validate the eczema component of the self-applicable ISAAC's WQ following its translation into Portuguese. The group of 10 pediatricians and 10 pediatric allergologists graded the questions from 0 to 2 and established the maximum score for each question. The WQ was answered by parents or guardians of children with atopic dermatitis (AE), aged 6-7 years (n = 23) and of non-AE control children of the same age (n = 46) as well as by AE (n = 24) and non-AE (n = 48) adolescents, aged 13-14 years. In order to evaluate the reproducibility of the ISAAC WQ, half of these individuals answered the same questionnaire after 2 to 4 weeks. The maximum possible global scores were 13 for the children aged 6-7 years and 11 for the adolescents, and the cutoff level for both groups was 3. In both age periods the WQ was reproducible (Kappa and McNemar tests) in a significant way (6-7 years, Kw = 0.79; 13-14 years, Kw = 0.73). The prevalence of AE, using the validated WQ, was then studied. The WQ was applied to the parents of 3,005 children aged 6-7 years and to 3008 children aged 13-14 years. Response rates were 72% and 94% for the 6-7-year-old children and the 13 14-year-old children, respectively. There was a slight predominance of male children in the studied population. In the group of the 6-7-year-old children, the cumulative prevalence of AE was 13.2% for boys and for girls; in the group of the 13-14-year-old children, it was 12.5% and 15.4%, respectively. AE severity was similar for both age groups. Using the criteria of global cutoff score, in the group of the 6-7-year-old children, the prevalence of AE was 12.6% for boys and 13.8 for girls; in the group of the 13-14-year-old children, it was 11.7% and 12.4%, respectively. There were no significant differences between them. In conclusion, the AE component of the ISAAC WQ proved to be reproducible, adequate, and able to discriminate between AE and control children. A significant concordance was observed between the criteria utilized in this study (ISAAC x global cutoff score). PMID- 12109531 TI - Simplified local nasal immunotherapy in mite dust allergic rhinitis. AB - The present work aimed at evaluating the efficacy and tolerance of an alternative schedule of local nasal immunotherapy for the treatment of mite dust allergic rhinitis. The authors suggest the nasal administration of the maximum tolerated dosage chosen on the basis of nasal provocation test threshold, comparing allergen extracts in micronized powder and watery solution. Forty-five patients (25 men and 20 women), aged 18 to 66 years, affected by allergic rhinitis to Dermatophagoides (Dpt) were selected and treated either by local immunotherapy in watery solution (15) or in powder form (15) or by parenteral specific hyposensitizing treatment (15). Before and one year after the beginning of the study, the clinical diaries and the total and specific IgE variation were evaluated. The monthly symptoms and drugs use are comparable among the three treatment groups. No significant difference was found, with the exception of local symptomatology, which improved more in patients undergoing local immunotherapy (p > 0.05); and oral antihistamines use, which was lower in patients treated with the watery solution (p < 0.05). Thus, local simplified hyposensitizing treatment is able to combine the absence of symptomatological worsening with the decrease of both local and systemic drugs use. The advantages of the LNIT protocol proposed herein are as follows: simplified schedule for self administration; improved patient compliance; reduction of local side effects; clinical efficacy comparable with subcutaneous specific immunotherapy. PMID- 12109532 TI - Respiratory infection in asthma. AB - Ever since the first decades of the 20th century, some authors have given respiratory infection triggered by bacteria an etiologic role in bronchial asthma, focusing on infection and the asthmatic response. In 1995 our group already presented a study in this sense on nasal secretion cultures and the relationship between IgE and sensitization to allergens. There is a significant association between patients with sensitization to Dermatophagoides, high levels of total IgE, and positive culture to Staphylococcus aureus. Following studies by Norn, we performed a study with 40 children, aged 2-14 years, where we observed that children with sensitization to mites and a positive culture had higher levels of histamine release than did children with negative culture and controls, the differences being significant. We also found, like other authors, that the joint presence of Staphylococcus aureus and Derrmatophagoides pteronyssinus potentiates antigen-specific histamine release. In recent years, with the increasing prevalence of bronchial asthma being studied, the role that infection could play in this increase is being considered again among other factors. As participants of the ISAAC project and using the same methods as in this study, we performed a simultaneous questionnaire with questions related with triggering and contributing factors, etc., including respiratory infection. We found an association between having had more than three episodes of "bronchitis" with fever and lasting for longer than seven days in the last year and having ever had asthma (OR 29.09). This association is still greater with having had wheezing in the last 12 months (OR 43.26), a finding that it is also associated with requiring attention in an emergency room (OR 30.65). From these results, we concluded that respiratory infection is an aggravating factor of asthma, something we already knew. In order to have our own experience, we studied serum interleukin 4 (IL-4) and interferon gamma (IFNgamma) in a sample of 41 children aged 3 to 17 years. The most frequent values of IL-4 ranged between 0.25 and 0.40 ng, and very low dispersion was found in the sample, which did not allow correlation with other parameters. Regarding IFNgamma, we found values between < 5 pg/ml and 605 pg/ml. When we studied children under treatment with antigen specific immunotherapy, we observed mean values of IFNgamma of 115.86 pg/ml, whereas the ones who did not follow this treatment or had followed it for less than one year had a mean of 66.06 pg/ml, these differences being significant (p = 0.035), and proving a Th1 response to immunotherapy. This significance is not found if children who have been under immunotherapy for less than one year are included. When we studied children with bacterial immunotherapy, we found that the mean IFNgamma value in children under immunotherapy for longer than one year was 56.4 pg/ml, whereas in children with no immunotherapy it was 101.75 pg/ml (p = 0.034). We conclude that bacterial immunotherapy modifies the Thl response, inhibiting it in those children with greater susceptibility to infections. PMID- 12109533 TI - Changes in inflammatory and clinical parameters and in bronchial hyperreactivity asthmatic children sensitized to house dust mites following sublingual immunotherapy. AB - In allergic patients with rhinoconjunctivitis (RNC), sublingual immunotherapy (SLIT) is effective in reducing both clinical symptoms and immuno-mediated local inflammatory responses. The study evaluates whether SLIT could reduce upper airway inflammation and improve clinical parameters also in children with rhinoconjunctivitis and asthma from house dust mite sensitization. Ten children with mild to intermittent asthma, monosensitized to house dust mites, received SLIT (Der p 1 monthly dose = 48 microg) for 2 years, in addition to "as needed" pharmacologic treatment. Before (T0) and after treatment (T2), changes were evaluated in (1) nasal eosinophilia, (2) intercellular adhesion molecule (ICAM)-1 expression by nasal epithelial cells, (3) RNC, asthma, and drug-intake scores, assessed by diary card, (4) pulmonary function parameters, and (5) bronchial reactivity to methacholine (MCh). RESULTS: All children completed the study without side or adverse effects. After the treatment period, we found no modification in nasal eosinophil values in mean fluorescence channel percentages, but a significant downregulation in ICAM-1 expression by nasal epithelial cells [mean (mfc): T0: 13.5 +/- 3.8 mfc; T2:4.6 +/- 0.5 mfc; p = 0.04]. These changes were associated with a reduction in RNC score [median values: T0: 3.4 (1.4-6.2); T2: 1.1 (0.6-2.4) p = 0.012], asthma score [median values: T0: 0.5 (0.4-1.0); T2: 0.3 (0.1-0.5); p = 0.005] and drug-intake score [median values: T0: 4.2 (3.1 5.3); T2: 1.1 (0-3.0); p = 0.005]. These clinical effects were not associated with changes in pulmonary function parameters (p > 0.05), but with improvement in bronchial reactivity to MCh [mean values: T0: 338.8 (91.5-1255.5); T2: 1698.2 (1,110.5-2,597.1); p = 0.02]. To what extent these observations may be related to the natural improvement of respiratory allergy symptoms with age remains to be determined. PMID- 12109534 TI - Recurrent urticaria as a rare manifestation of familial Mediterranean fever. AB - Familial Mediterranean fever (FMF) is a genetic disorder characterized by acute episodes of fever with some combination of severe abdominal pain, pleurisy, arthritis, and skin rash. The case of a patient with recurrent urticaria referred for study of drug allergy is presented. After allergy had been ruled out, the urticaria was attributed to previously undiagnosed symptoms of an underlying systemic disease: FME. Urticaria is the least frequent cutaneous manifestation of this disease, and genetic analysis was required to confirm the diagnosis. PMID- 12109535 TI - Allergic contact dermatitis to temporary tattoo by p-phenylenediamine. AB - Temporary tattoos are widely applied today all over the world. The tattoo makers explain that they use "natural henna paint," although in fact they use "black henna," which includes a mixture of many substances, among them p phenylenediamine (PPD). There have recently been many reports of allergic contact dermatitis because of temporary tattoo with PPD sensitization. We are adding a new case of temporary tattoo with black henna with an extensive reaction, in which a 12-year-old white boy showed contact dermatitis from PPD, followed by cutaneous eruption after corticosteroid topical treatment. PMID- 12109536 TI - Vocal cord dysfunction: more morbid than asthma if misdiagnosed. AB - Vocal cord dysfunction may lead to respiratory distress, which in turn may cause the misdiagnosis of asthma. We present two adolescents who were misdiagnosed as asthmatic and aggressively treated for asthma with resulting iatrogenic complications. Vocal cord dysfunction simulating or coexisting with asthma should be considered especially in patients with frequent emergency visits for episodic dyspnea despite aggressive treatment. PMID- 12109537 TI - Anaphylactoid reaction caused by moxifloxacin. AB - We report on a patient who suffered an anaphylactoid reaction because of moxifloxacin. PMID- 12109538 TI - Sjogren's syndrome: viewpoint on pathogenesis. One of the reasons I was never asked to write a textbook chapter on it. AB - OBJECTIVE: To critically consider the public opinion/consensus on SS formulated by opinion leaders and textbook chapters. RATIONALE: Although our clinical work is based on evidence-based medicine, it is obvious that we do not have evidence based solutions to the etiology and pathogenesis of autoimmune rheumatic diseases. In spite of this, consensus if often taken as a truth, which may hamper the production, funding and/or publication of new and original ideas and views. METHODS: Comparison of the classic view with one of the many other possible views. RESULTS: The consensus view states that 1) SS is initiated and/or caused by an exogenous agent, probably some type of retrovirus, and 2) after initiation, a straightforward sequence of events follows: a) salivary gland epithelial cells are disrupted, b) T lymphocytes migrate to and are activated in the glands, c) B cells get the help they need and start to produce SS and RF autoantibodies, which processes lead to structural destruction and loss of acinar cells and, thus, to sicca symptoms (an example of the linear, step-by-step "computer" logic). The problems inherent to this view include: 1) why women? (gender aspect), 2) why at the age of 50? (chronobiologic aspect), 3) is the normal immune system in SS only responding to normal (formely sequestrated) autoantigens? Is the loss of exocrine gland function really caused by "autoimmune" destruction? - or do the SS autoantibodies and lymphocyte infiltrates only represent markers in an appropriate HLA background? (autoimmune aspect), 4) are the retroviral diseases really similar to SS? (exogenous rs endogenous causes), 5) is our current view compatible with unexpected, future findings? Is the textbook interpretation the final truth (evolutionary aspect of our view on pathogenesis). CONCLUSION: The tubuloalveolar exocrine glands may be seen as 1) locus minoris resistentiae for normal oral microbial flora and immune-inflammatory attacks at the normal environment-host interface. Apoptotic and/or necrotic cells are released into the intraluminal space and pass in normal glands through normal, immunologically competent lymphocyte foci and/or ectopic lymphatic tissue. Acinar cell degeneration/death may increase upon 2) aging and acinar cell renewal and well being may be hampered by age-dependent deficiencies in the trophic 3) neuro endocrine support. In a proper immunogenetic setting, a) marker autoantibodies (e.g. RF, SS-A/Ro, SS-B/La), useful in the diagnosis, are produced. However, sicca symptoms/SS develop only if muscarinic receptor or other b) pathogenetic autoantibodies disrupting the normal neuronal-to acinar cell communication are also produced. c) Systemic symptoms could be produced on neuroendocrine, chronobiologic and autoimmune basis. Other professionals are invited to entertain their own views on the pathogenesis of SS - make your own one! PMID- 12109539 TI - Collaborative research into outcome measures in Sjogren's syndrome. Update on disease assessment. AB - Sjogren's syndrome (SS) is a multisystem immune-mediated disorder characterized by inflammation of exocrine glands leading to clinical symptoms of dryness, particularly of the eyes and mouth which can be severe and disabling. It can occur in association with other rheumatic disorders or as a primary entity (PSS), often associated with B-cell hyperreactivity manifested by hypergammaglobulinaemia and anti-Ro and/or anti-La autoantibodies. These patients are more likely to have systemic involvement, for example of the pulmonary, neurological or haematological systems, and have a 44 times increased risk of non Hodgkins lymphoma, this being the major adverse outcome in this disorder. Clinical trials of new therapies in disorders such as rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE) have been advanced by the development of internationally agreed assessment and outcome measures such as the American College of Rheumatology/EULAR core datasets of disease activity in RA and measures of activity and damage in SLE such as BILAG, SLEDAI, SLICC etc. No such equivalent consensus exists for SS and as a result clinical trials have all used different, unvalidated, ad hoc measures of disease assessment and outcome. One attempt to bring a coherent approach to this issue is the "Copenhagen model" or disease assessment wheel that places each disease manifestation within specific categories depending on their presumed pathogenesis. Attempts have also been made to develop this into a severity tool by grading specific manifestations according to severity. In March 2000, interested specialists from around Europe met to develop a consensus on the broad principles underpinning disease assessment in SS (particularly PSS). It was agreed to adopt the international approach of dividing assessment into: exocrine and-nonexocrine disease activity (potentially reversible and including sicca symptoms and objective measures and systemic symptoms (particularly fatigue) and clinical features), damage (present for over 6 months), health-related and generic quality of life, and standard approaches to adverse events/toxicity and health economic aspects. The workshop also began the process of developing specific measures of sicca symptoms and systemic activity and damage. In order to convert this approach into detailed, experimentally validated assessment tools, a UK-based collaboration has focused on the development of systemic and sicca symptom questionnaires and a prospective evaluation in Europe of systemic activity and damage measures is planned. PMID- 12109540 TI - Case report I. A patient with sicca features and arthritis. PMID- 12109541 TI - Sjogren's syndrome: mechanisms of pathogenesis involve interaction of immune and neurosecretory systems. AB - Although biopsies of salivary and lacrimal glands from patients with Sjogren's syndrome (SS) have focal lymphocytic infiltrates and partial destruction of glandular secretory units (acinar and ductal structures), the degree of dryness is beyond that expected for the level of glandular destruction. The failure to exhibit adequate secretory function is not due simply to the destruction of neural innervation to the residual glandular elements or the absence of receptors for acetylcholine on the glandular cells. It is likely that release of cytokines by lymphocytes and glandular cells (especially interleukin-1, interleukin-6 and tumor necrosis factor alpha), autoantibodies and metalloproteinases lead to decreased release of neurotransmitters and decreased response of the residual glandular cells to available neurotransmitters. The ability to modulate immune response and stimulate residual glandular elements provides new therapeutic opportunities for Sjogren's patients. PMID- 12109542 TI - Case report II. A former healthy 58-year old woman suddenly developed acute, symmetrical polyarthritis. PMID- 12109543 TI - Flaw localization using the reassigned spectrogram on laser-generated and detected Lamb modes. AB - Rapid, accurate inspection of metallic plates using broadband guided waves has traditionally been limited by the multi-mode, highly dispersive nature of these waves; current practice typically restricts either the type of mode generated or detected, and/or its frequency range. The current study presents an effective alternative procedure by combining a novel digital signal processing technique, the reassigned spectrogram, with laser generated and detected Lamb waves. The reassigned spectrogram is used to characterize the modal and frequency content of a single ultrasonic signal as a function of time, enabling a procedure to locate flaws in an aluminum plate specimen. PMID- 12109544 TI - Mass spring lattice modeling of the scanning laser source technique. AB - The scanning laser source (SLS) technique is a promising new laser ultrasonic tool for the detection of small surface-breaking defects. The SLS approach is based on monitoring the changes in laser generated ultrasound as a laser source is scanned over a defect. Changes in amplitude and frequency content have been observed for ultrasound generated by the laser over uniform and defective areas. In this paper, the SLS technique is simulated numerically using the mass spring lattice model. Thermoelastic laser generation of ultrasound in an elastic material is modeled using a shear dipole distribution. The spatial and temporal energy distribution profiles of typical pulsed laser sources are used to model the laser source. The amplitude and spectral variations in the laser generated ultrasound as the SLS scans over a large aluminum block containing a small surface-breaking crack are observed. The experimentally observed SLS amplitude and spectral signatures are shown to be captured very well by the model. In addition, the possibility of utilizing the SLS technique to size surface-breaking cracks that are sub-wavelength in depth is explored. PMID- 12109545 TI - Evaluation of the acoustic intensity of new ultrasound therapy equipment. AB - International safety standards recommend a limit below 30% variation in acoustic intensity for ultrasound therapy (UST) equipment. In view of this question, the purpose of this work was to evaluate the intensity of new UST equipment in the Brazilian market. An evaluation was performed of eight models manufactured by six different national manufacturers; under continuous and pulsed conditions, at frequencies of 1.0-3.0 MHz, for a total of 48 items of equipment. The intensities were analysed according to the technical standards IEC 601-2-5, in the range 0.01 3.0 Wcm(-2), using a radiation pressure scale UPM-DT-10 (Ohmic Instruments), previously calibrated. The results demonstrated that the models Sonacel, Sonacel plus, Sonacel III, Avatar I, and Sonamed I, although they were new (unused) presented calibration errors of over 30% in more than one intensity checked, and the models SONOPULSE, PRO-SEVEN and SONOMASTER ST. are within the standards proposed. The results show that industry must improve quality control on their production lines, as well as that there is a need for a supervising body at national level. Published by Elsevier Science B.V. PMID- 12109546 TI - Numerical analysis of the hybrid transducer ultrasonic motor: comparison of characteristics calculated by transmission-line and lumped-element models. AB - In this paper, a hybrid transducer ultrasonic motor is numerically analyzed by using two equivalent electrical circuit models. A transmission-line model for the torsional vibration in the stator, which can model any torsional vibration mode and their combinations, was introduced and compared with a lumped-element model, which modeled the fundamental torsional resonance mode in the stator. The calculation result by using the transmission-line model demonstrated that the second harmonic torsional vibration increased either with the static spring force by which the rotor was pressed to the stator or with the load torque placed on the rotor. The difference in the calculated motor performance between the two models appeared when the second harmonic torsional vibration became large at a sufficient static spring force. PMID- 12109547 TI - Ultrasonic characterization of orthotropic elastic bovine bones. AB - The aim of the present study was to determine the mechanical properties of bovine bones. An ultrasonic method was used to determine acoustical parameters such as the longitudinal and transverse velocities in the longitudinal and two radial directions of compact bone, i.e., in all directions of the plane. Waves propagating through bovine femoral bones were studied using an ultrasonic scanner for linear and sectorial scanning. The mechanical parameters of compact bone, such the Young's modulus and Poisson's ratio in the orthotropic case, were then determined from the measured velocities. The results are in line with those in the literature. PMID- 12109549 TI - A simple scattering model for measuring particle mass fractions in multiphase flows. AB - In this paper we present a simple theoretical model of how pulsed ultrasound is attenuated by the particles in a solid/liquid flow. The theoretical model is then used to predict the attenuation of sound, given the mass fraction, the density, and the size distribution of the solid particles. The model is verified experimentally for suspensions of 0-10% (by mass) Dolomite ((Ca,Mg)CO3) particles and water. The experimental results show that the attenuation of sound due to particles varies linearly with mass fraction, and that the proposed theoretical model can be used to predict this attenuation. In all experiments the transmitter and receiver array were clamped onto the pipe wall, thus providing a completely non-invasive and non-intrusive measurement technique. PMID- 12109548 TI - Theoretical determination of Rayleigh wave acoustoelastic coefficients: comparison with experimental values. AB - The characterization of stress states in materials is often necessary in some industrial application. The ultrasonic methods can be potentially convenient since stress states inside materials can be obtained even if materials are opaque. Nevertheless, the knowledge of acousto-elastic coefficients is generally necessary to estimate residual stresses by ultrasonic methods, but the experimental determination of these acousto-elastic coefficients can be difficult in some cases. In this paper, Rayleigh wave (RW) acousto-elastic coefficients of an orthotropic material are theoretically determined according to its characteristics, i.e. the density and the secondand third-order elastic constants. Then, these RW acousto-elastic coefficients are directly measured during an experimental stage and a comparison between calculated and measured coefficients is realized. This study allows on the one hand to check the theoretical development and on the other hand to show that it is possible to calculate acousto-elastic coefficients theoretically from intrinsic characteristics of the material rather than measuring them directly during a calibration phase which is sometimes long and difficult to realize. PMID- 12109550 TI - Miniaturization of surface acoustic waves rotary motor. AB - This paper presents the experimental study of a miniaturized surface acoustic waves (SAWs) rotary motor and the theoretical calculation. After the first success in SAW rotary motor operating at 9.85 MHz, a smaller rotary motor is designed. With the operating frequency of 30 MHz and the driving voltage of 120 V(p-p), the motor can rotate at a speed of 270 rpm. PMID- 12109551 TI - Interdigital transducer analysis using equivalent PSpice model. AB - Interdigital transducers generating surface acoustic waves have found numerous applications. The present paper deals with an equivalent model of a single interdigital transducer as well as an SAW filter complying with the PSpice programme for the analysis of electronic systems. The suggested model makes it possible to take into account such effects as interelectrode reflections, losses along the acoustic path and the electromagnetic coupling between the transducers. The obtained results of the analysis of an interdigital transducer comply with the results obtained basing on an admittance model and delta-function model. The final aim is to model an SAW oscillator. PMID- 12109552 TI - Can health promotion at the workplace help prevent cancer? PMID- 12109553 TI - Health promotion trials at worksites and risk factors for cancer. AB - Studies of worksite health promotion have frequently reported larger effects than those at the community level. Many of these studies have serious methodological problems. Forty-five worksite health promotion trials following specific quality criteria were selected and estimated for behavioral changes in cancer risk factors and the effectiveness of different intervention components. Tobacco control programs found quit rates of about 5% with relapse rates of 40% to 80% at 6 months after the intervention. Effectiveness increased with the duration of the intervention for at least 6 months, repeated contacts with the participants, continuous support, and tailored messages. There was less evidence for the long term effectiveness of incentives. Trials on diet, alcohol, physical activity, overweight, and solar radiation showed the same positive trends. The overall evidence indicates a modest but positive effect of health promotion trials at worksites and the effect, for smoking cessation trials, is slightly larger than that of community-based trials. Many of the recommendations made to increase participation and effectiveness were not based on empirical data. PMID- 12109554 TI - Self-experienced physical workload and risk of breast cancer. AB - OBJECTIVES: The association between the risk of breast cancer and the physical load of work was studied because physical activity may reduce breast cancer risk via hormonal mechanisms. METHODS: Occupational physical activity was estimated from a self-determined rating [scale 1 (low)-5 (high)] of occupational physical load for 1800 randomly selected women born in 1930-1969. The medians of the ratings were used as occupation-specific indices of occupational physical activity. All 65 occupations with at least 5 ratings, covering 75% of the economically active female population in Finland, were included in further analyses. The occupation-specific numbers of observed and expected cases of breast cancer during 1971-1995 among women born in 1906-1945 (17,986 cases) were grouped according to the index for occupational physical activity. Expected rates were calculated with the social-class-specific population and the entire Finnish female population as reference populations. The relative risks (RR) of breast cancer for categories 3-5, in comparison with categories 1-2 were calculated using Poisson regression models. The occupation-specific mean number of children and mean age at first childbirth were adjusted for. RESULTS: The RR was lower for occupations in category 5 than for those in categories 1-4, especially in the youngest (25-39 years) age group (RR 0.51, 95% confidence interval 0.44-0.58). Adjustment for social class and reproductive factors raised the RR (95% confidence interval 0.56-0.74) for category 5 in different age strata, all the RR values still being statistically significant. CONCLUSIONS: The results support the hypothesis that occupational physical activity, if high enough, markedly reduces breast cancer risk. PMID- 12109555 TI - Ten-year update on mortality among mild-steel welders. AB - OBJECTIVES: This study is an update on the lung cancer risk of mild-steel welders with no asbestos exposure using a cohort of nonwelders for comparison. METHODS: The subjects came from three United States (US) plants that manufactured heavy equipment. The follow-up was extended from 1988 to 1998. The welders were not exposed to asbestos (typical of shipyard welders) or to chromium or nickel (present in stainless steel). RESULTS: There were 108 lung cancer deaths among the welders and 128 such deaths among the nonwelders (double the previous number of lung cancer deaths). The standardized mortality ratio (SMR) for lung cancer was 1.46 [95% confidence interval (95% CI 1.20-1.76] for the welders and 1.18 (95% CI 0.98-1.40) for the nonwelders, both in comparison with the US general population. Direct comparison between the welders and nonwelders yielded a rate ratio of 1.22 (95% CI 0.93-1.59). Analyses using a 15-year lag time did not differ greatly from those of an unlagged analysis. There were no marked trends for lung cancer risk by duration of exposure or latency. Evidence from cross sectional data from a sample of the cohort indicated that the welders smoked somewhat more than the US population and more than the nonwelders. An approximate adjustment of the rate ratios for possible confounding by smoking suggested that smoking may have accounted for about half of the excess lung cancer observed among the welders versus that of either reference population. CONCLUSIONS These data provide suggestive but not conclusive evidence of a modest lung cancer risk from mild-steel welding. PMID- 12109556 TI - Physical workload of student nurses and serum markers of collagen metabolism. AB - OBJECTIVES: This study explored the association between biomarkers of type I collagen metabolism and exposure to physical workload. METHODS: In a prospective cohort study, serum concentrations of markers of type I collagen synthesis and degradation were assessed monthly for student nurses who worked as nurses for a period of 6 months and compared with those of a reference group. The number of patient-handling activities was estimated from observations at the workplace. Linear generalized estimating equations were used to analyze differences in the serum concentrations of the biomarkers between the exposed group and reference group, as well as to analyze whether the number of patient-handling activities was associated with serum concentrations of the biomarkers. RESULTS: Serum concentrations of the biomarkers were found to differ between the groups. The biomarkers reflected a higher anabolism of type I collagen in the exposed group when compared with that of the reference group. An analysis of the effect of the number of patient-handling activities revealed that a higher exposure was associated with higher effective type I collagen synthesis within the exposed group. CONCLUSIONS: These results indicate that serum concentrations of these biomarkers of type I collagen metabolism can reflect differences in exposure between contrasting groups, and also varying levels of exposure between persons within an occupation. PMID- 12109557 TI - Individually fitted sports shoes for overuse injuries among newspaper carriers. AB - OBJECTIVES: The aim of the study was to determine the effectiveness of new, individually fitted sports shoes against overuse injuries to the lower limb among newspaper carriers. METHODS: Patients (N = 176) with lower-limb overuse injuries were randomly assigned to use new, individually adjusted footwear with good shock absorbing properties (test group = 86) or the subjects' own, used footwear (control group = 90). The main outcome measurements were lower-limb pain intensity during walking, as rated on a visual analogue scale (0-100), number of painful days, subjective assessment of global improvement, foot fatigue, number of hyperkeratotic skin lesions and diagnosed overuse injuries, and costs of foot care as compared between the treatment groups. RESULTS: At the 6-month follow-up there was a difference in favor of the test group with respect to lower-limb pain intensity and number of painful days, when compared with the control group. At 1 year, 53% and 33% of the test and control groups, respectively, thought they were better than at the time of the baseline examination (number needed to treat being 5 between the test and control groups). The test subjects had less foot fatigue and fewer hyperkeratotic skin lesions. There was no difference in the number of diagnosed overuse injuries between the groups. During the year of follow-up, the all-inclusive mean costs of foot care were USD 70 and USD 158 in the test and control groups, respectively. CONCLUSIONS: Individually adjusted shock-absorbing shoes offer slight health benefits for lower-limb overuse injuries. Proper shoes may decrease the need to use health care resources. PMID- 12109558 TI - Physical and psychosocial prerequisites of functioning in relation to work ability and general subjective well-being among office workers. AB - OBJECTIVES: The purpose of the study was to investigate the physical and psychological prerequisites of functioning, as well as the social environment at work and personal factors, in relation to work ability and general subjective well-being in a group of office workers. METHODS: The study was a descriptive cross-sectional investigation, using path analysis, of office workers. The subjects comprised 88 volunteers, 24 men and 64 women, from the same workplace [mean age 45.7 (SD 8.6) years]. The independent variables were measured using psychosocial and physical questionnaires and physical measurements. The first dependent variable, work ability, was measured by a work ability index. The second dependent variable, general subjective well-being, was assessed by life satisfaction and meaning of life. The variables were structured according to a modified version of the International Classification of Functioning, Disability and Health. RESULTS: Forward flexion of the spine, intensity of musculoskeletal symptoms, self-confidence, and mental stress at work explained 58% of work ability and had indirect effects on general subjective well-being. Self confidence, mood, and work ability had a direct effect on general subjective well being. The model developed explained 68% of general subjective well-being. Age played a significant role in this study population. CONCLUSIONS: The prerequisites of physical functioning are important in maintaining work ability, particularly among aging workers, and psychological prerequisites of functioning are of even greater importance in maintaining general subjective well-being. PMID- 12109559 TI - A new occupational health agenda for a new work environment. AB - At the beginning of the 21st century, the emergence of new forms of work organization are transforming what had become standard types of work arrangements in industrialized countries. In this new labor market environment, new firms, types of workers, and risk factors are powerfully emerging. Contrary to common belief, emergent occupational health hazards should not be approached only as "technical" or "economic" value-free problems. Instead, many of the challenges faced by occupational health policy makers are predominantly related to professional values and to the political ideologies and economic interests of key stakeholders in the decision-making process. In this paper some of the key principles leading to efficient and equitable occupational health policies in the new work environment are discussed. An alternative is also proposed for dealing with the conditions and settings needed to meet the new challenges related to establishing an effective occupational health policy. PMID- 12109560 TI - Evidence-based medicine for occupational health. AB - OBJECTIVES: This study attempted to determine the feasibility and utility of methods used in evidence-based medicine for some common questions in the practice of occupational medicine. METHODS: The following clinical questions were generated that were representative of the type of problems encountered by occupational health physicians: is work a cause of health problems and is impaired health a cause of diminished work capacity for a specific job? Answers were generated according to the method used in evidence-based medicine by formulating an answerable question, searching the literature, critically appraising the results, and applying the results to the clinical question. RESULTS: Answers were found to all the questions in a reasonable amount of time. The searches revealed a need for more systematic reviews and studies that use work-related health outcomes like return to work. However, there is more evidence available in Medline than is generally assumed by occupational health physicians. Using this evidence led to better clinical decisions. Pitfalls during the literature search were typing mistakes, problems in using medical subject headings, and unreliable search strategies. With the use of the abstracts only, most clinical questions could be answered satisfactorily, but concrete risk estimates were often lacking. The lack of availability of full text journals decreased the reliability of the critical appraisal and risk estimation. CONCLUSIONS: Evidence-based medicine is a feasible and useful method for occupational medicine. Instruction and training is needed for most occupational health physicians to increase their searching and critical appraisal skills. More research is needed to determine the information needs of occupational health physicians and to develop tools that facilitate literature searches. PMID- 12109561 TI - Inhalation of decomposed chlorodifluoromethane (freon-22) and myocardial infarction. AB - After exposure to decomposed chlorodifluoromethane (freon-22), a 65-year-old man developed respiratory symptoms such as cough, blood-stained sputum, and increasing dyspnea. Three weeks later, his family doctor diagnosed infectious bronchitis. Another week later he died due to myocardial infarction. The discussion focuses on an inflammatory process caused by the inhalation of decomposed freon and its possible association with myocardial infarction. PMID- 12109562 TI - Simultaneous determination of parathion and p-nitrophenol in vegetable tissues by derivative spectrophotometry. AB - A first derivative spectrophotometric method has been developed for the determination of parathion and p-nitrophenol in vegetable tissues. Ethanol was used as solvent for extracting the compounds from the tissues and subsequently the samples were evaluated against a vegetable tissue blank, directly by derivative spectrophotometry. The simultaneous determination of these compounds can be carried out using the zero-crossing approach for parathion at 253.0 nm and for p-nitrophenol at 273.1 nm. In the samples each analyte was determined in the presence of one another in the ranges between 4.9 to 3883.5 microg g(-1) for parathion and 4.9 to 3285.3 microg g(-1) for p-nitrophenol. The detection limits (3a) were found to be 1.5 and 1.4 microg g(-1) for parathion and p-nitrophenol, respectively. The relative standard deviations were in all instances less than 1.8%. The proposed method was applied to the determination of the analytes in spiked leaves of corn. The results show a good recovery and they are in agreement with those obtained by polarography. PMID- 12109563 TI - Patchy distribution fields: a zigzag survey design and reconstruction adequacy. AB - A mathematical model was used to examine the effects of a zigzag survey design on the adequacy of reconstructing patchy distribution fields. The model simulates fish or plankton patches (or gaps) of different shapes and spatial orientations, and an acoustic survey by a set of transects forming a zigzag. Adequacy of the reconstructed fields to those originally generated was evaluated by calculating their correlations (r). A priori information on the autocorrelation radius for the field in the direction of a survey (R) allows optimization of survey design and the algorithm of data analysis. A patchy field can be reconstructed properly (r > 0.80) if the distance between transects D < (1.0-1.5)R. If a priori information on the field is not available, the autocorrelation radius should be determined when reconstructing the field, i.e. a posteriori. In cases of field movement, the criterion for choosing a survey direction is based on the relationship between the dimension of moving patches in the direction of movement and that of the surveyed area. The results obtained indicate that, for a fixed transect spacing, zigzag pattern allow less adequate reconstruction of an original distribution field (in cases of both immovable and movable fields) than corresponding parallel pattern. PMID- 12109564 TI - Effects of exposure of crocodiles to sublethal concentrations of Petroleum waste drilling fluid in the Niger Delta basin of Midwestern Nigeria. AB - Static bioassay were carried out using two aquatic crocodiles (the short nosed crocodile, Osteolemus tetraspis and the Nile crocodile, Crocodilus niloticus) as test organisms in soft natural dilution water, with Petroleum waste drilling fluid as the test material, at 28 +/- 2 degrees C. Comparison of results for the control and different concentrations of the waste drilling fluid were made by means of the F-statistic method. Both crocodile species exhibited a high insensitivity to the undiluted waste drilling fluid and the different dilutions. Differences in concentration of waste drilling fluid did not influence the response of crocodiles to the potential toxicant. Percentage of deaths which was never greater than 0.2% in control tanks was not significantly different from that in test tanks where mortality values of organisms was typically 1.6% or less in most cases. There was a delay toxicant-induced mortality effect. PMID- 12109565 TI - Multi-level screening of a proposed hazardous waste treatment and disposal facility site--a case study. AB - A comprehensive environmental assessment (EA) is a pre-requisite before the site for developing a hazardous waste treatment and disposal facility (HWTDF) is selected. However, the resource limitations, especially for developing countries, often dictate that the detailed EA be restricted to those sites, which are constraint free and have low hazard potentials. Thus, a preliminary screening exercise for assessing the suitability of site for developing the HWTDF needs to be carried out to avoid huge costs involved in detailed EA. While screening a HWTDF site, various factors such as present land use, ecologically sensitive areas, geology and hydrogeology of the area, the quality and quantity of wastes, engineered safeguards, and the operational procedures that need to be adopted. are required to be addressed. In this paper, a multi-level screening criteria employing Remote Sensing, Constraint Mapping, Groundwater Pollution Potential Index (DRASTIC Index), and the Site Ranking was used to assess the suitability of a proposed site for the development of HWTDF. The study helped to identify various constraints at the proposed site and their significance on the development of the HWTDF. PMID- 12109566 TI - A ten year summary of concurrent ambient water column and sediment toxicity tests in the Chesapeake Bay watershed: 1990-1999. AB - The goal of this study was to identify the relative toxicity of ambient areas in the Chesapeake Bay watershed by using a suite of concurrent water column and sediment toxicity tests at seventy-five ambient stations in 20 Chesapeake Bay rivers from 1990 through 1999. Spatial and temporal variability was examined at selected locations throughout the 10 yr study. Inorganic and organic contaminants were evaluated in ambient water and sediment concurrently with water column and sediment tests to assess possible causes of toxicity although absolute causality can not be established. Multivariate statistical analysis was used to develop a multiple endpoint toxicity index (TOX-INDEX) at each station for both water column and sediment toxicity data. Water column tests from the 10 yr testing period showed that 49% of the time, some degree of toxicity was reported. The most toxic sites based on water column results were located in urbanized areas such as the Anacostia River, Elizabeth River and the Middle River. Water quality criteria for copper, lead, mercury, nickel and zinc were exceeded at one or more of these sites. Water column toxicity was also reported in localized areas of the South and Chester Rivers. Both spatial and temporal variability was reported from the suite of water column toxicity tests. Some degree of sediment toxicity was reported from 62% of the tests conducted during the ten year period. The Elizabeth River and Baltimore Harbor stations were reported as the most toxic areas based on sediment results. Sediment toxicity guidelines were exceeded for one or more of the following metals at these two locations: arsenic, cadmium, chromium, copper, lead, nickel and zinc. At the Elizabeth River stations nine of sixteen semi-volatile organics and two of seven pesticides measured exceeded the ER-M values in 1990. Ambient sediment toxicity tests in the Elizabeth River in 1996 showed reduced toxicity. Various semi-volatile organics exceeded the ER-M values at a number of Baltimore Harbor sites; pyrene and dibenzo(a,h)anthracene were particularly high at one of the stations (Northwest Harbor). Localized sediment toxicity was also reported in the Chester, James, Magothy, Rappahannock, and Potomac Rivers but the link with contaminants was not determined. Both spatial and temporal variability was less for sediment toxicity data when compared with water column toxicity data. A comparison of water column and sediment toxicity data for the various stations over the 10 yr study showed that approximately half the time agreement occurred (either both suite of tests showed toxicity or neither suite of tests showed toxicity). PMID- 12109567 TI - Knowledge and attitude assessment and education of prehospital personnel in child abuse and neglect: report of a National Blue Ribbon Panel. PMID- 12109568 TI - Law Enforcement Agency Defibrillation (LEA-D): proceedings of the National Center for Early Defibrillation Police AED Issues Forum. AB - Why does LEA-D intervention seem to work in some systems but not others? Panelists agreed that some factors that delay rapid access to treatment, such as long travel distances in rural areas, may represent insurmountable barriers. Other factors, however, may be addressed more readily. These include: absence of a medical response culture, discomfort with the role of medical intervention, insecurity with the use of medical devices, a lack of proactive medical direction, infrequent refresher training, and dependence on EMS intervention. Panelists agreed that successful LEA-D programs possess ten key attributes (Table 6). In the end, the goal remains "early" defibrillation, not "police" defibrillation. It does not matter whether the rescuer wears a blue uniform--or any uniform, for that matter--so long as the defibrillator reaches the victim quickly. If LEA personnel routinely arrive at medical emergencies after other emergency responders or after 8 minutes have elapsed from the time of collapse, an LEA-D program will be unlikely to provide added value. Similarly, if police frequently arrive first, but the department is unwilling or unable to cultivate the attributes of successful LEA-D programs, efforts to improve survival may not be realized. In most communities, however, LEA-D programs have tremendous lifesaving potential and are well worth the investment of time and resources. Law enforcement agencies considering adoption of AED programs should review the frequency with which police arrive first at medical emergencies and LEA response intervals to determine whether AED programs might help improve survival in their communities. It is time for law enforcement agency defibrillation to become the rule, not the exception. PMID- 12109569 TI - Prehospital care and outcome of pediatric out-of-hospital cardiac arrest. AB - Cardiac arrest in children outside the hospital is associated with high mortality rates. Recent investigations have suggested that the use of advanced life support (ALS) measures by emergency medical services (EMS) personnel may decrease survival. These studies have used the pediatric Utstein style of defining ALS and basic life support (BLS) measures. The pediatric Utstein style defines BLS as "an attempt to restore effective ventilation and circulation" using noninvasive means to open the airway but specifically excludes the use of bag-valve-mask devices. Advanced life support is defined as the "addition of invasive maneuvers to restore effective ventilation and circulation." The authors of the study described below believe that using this definition would categorize some patients into an ALS group who would otherwise be categorized as having received BLS (i.e., "bag-valve-mask only"). OBJECTIVE: To compare survival rates among children receiving BLS or ALS following out-of-hospital cardiac arrest using amended definitions of prehospital life support measures. Specifically, the definition of BLS was expanded to include the use of bag-valve-mask devices only. METHODS: This was a retrospective chart review in an urban, pediatric emergency department. Patients included all children presenting to the emergency department between January 1, 1986, and December 31, 1999, following out-of-hospital cardiac arrest. The main outcome measure was survival to hospital discharge. RESULTS: Two hundred ten children were identified. Twenty-one patients were excluded from further analysis because of absent or incomplete medical records. One hundred eighty-nine patients were studied. Five children (2.6%) survived to discharge from the hospital. Of 189 children, 39 (20.6%) were provided BLS measures by prehospital personnel; 150 (79.4%) received ALS. There was no significant difference between groups in survival to hospital discharge. Patients who survived to hospital discharge were more likely to be in sinus rhythm upon arrival in the emergency department (p < 0.001) and to have received fewer doses of standard-dose epinephrine in the emergency department (p < 0.001). CONCLUSION: The use of ALS by prehospital personnel for children with out-of-hospital cardiac arrest did not improve survival to discharge from the hospital when compared with the use of BLS. PMID- 12109570 TI - Amiodarone and rural emergency medical services cardiac arrest patients: a cost analysis. AB - OBJECTIVE: Recent American Heart Association guidelines suggest amiodarone as an antiarrhythmic in refractory ventricular fibrillation (VF) and pulseless ventricular tachycardia (VT). The authors sought to assess the impact of amiodarone use on outcomes and cost associated with this practice in a rural emergency medical services (EMS) state. METHODS: Statewide EMS records were reviewed for the calendar year 1999. Data reviewed included prehospital diagnosis, medications given by prehospital providers to patients with cardiac arrest, and procedures performed, including cardiopulmonary resuscitation (CPR) and defibrillation. Cost-benefit analysis assumed the cost of amiodarone treatment to be $137.65 per patient encounter. Absolute risk reduction (ARR) and number needed to treat (NNT) analysis utilized resuscitation rates published in the ARREST and ALIVE trials. RESULTS: During the study period, EMS providers diagnosed 2,189 patients as having cardiac arrest. Five hundred thirty-five (24.4%) cardiac arrest patients were defibrillated. One hundred sixty patients (7.3%), including 15 who did not receive defibrillation, were given lidocaine during resuscitation efforts. The annual cost increase from current practice for a statewide amiodarone VF/VT protocol was $21,822.40 (10,572.87%). The initial cost to stock EMS vehicles for this protocol would be $50,115.52. The cost benefit analysis yielded a potential for one additional patient survival to hospital discharge in Maine per 3.125 years of system-wide practice at a cost of $68,840.00. CONCLUSION: Based on current data, instituting amiodarone treatment for refractory VF and pulseless VT in a rural EMS setting requires the investment of substantial resources, relative to current treatment strategies, for any potential survival benefit. PMID- 12109571 TI - Automated external defibrillation by very young, untrained children. AB - For patients with sudden cardiac death (SCD), the time interval to defibrillation is the main determinant of survival. As such, the American Heart Association has attempted to promote public-access defibrillation (PAD). Previous studies have shown that automated external defibrillators (AEDs) can be used successfully by untrained adults. OBJECTIVE: To determine whether very young, untrained children could use AEDs. METHODS: Third-grade students from an elementary school participated in this study representing a convenience sample of volunteers. They were given no formal training, but were shown how to peel off the backing from the electrode pads, like a sticker. Students were then given a mock code situation using a training manikin. The time to delivery of first shock was recorded. Students were then trained during a 2-minute review of the process, one on one with an instructor, and the study was then repeated. Data were analyzed using a paired Student's t-test comparing pre- and post-training. RESULTS: Thirty one children participated in the study, with a median age of 9 years. For untrained children, the mean time for delivery of the first shock was 59.3 +/- 13.6 seconds, 95% CI = 54.3 to 64.3. Following training, the mean time for delivery of the first shock was 35.2 +/- 6.0 seconds, 95% CI = 33.0 to 37.4, p = 0.001. CONCLUSION: Although this study suggests that even very young, untrained children can successfully perform automated external defibrillation, training does significantly decrease the time to delivery of first shock. PMID- 12109572 TI - ATLS practices and survival at rural level III trauma hospitals, 1995-1999. AB - OBJECTIVE: To determine whether Advanced Trauma Life Support (ATLS) practices characterizing initial resuscitation and interfacility transfer at rural trauma hospitals are associated with risk-adjusted survival. METHODS: Retrospective, observational analysis of rural injured patient survival. Process-of-care variables were associated with TRISS (trauma and injury severity score)-derived Z statistics (95% confidence intervals) for high-risk population subsets (defined below). INCLUSION CRITERIA: all patients > or = 12 years of age entered into a statewide trauma system, January 1, 1995, to December 31, 1999, and initially presenting to Level III trauma centers (N = 4,961). EXCLUSION CRITERIA: pronounced dead on arrival (n = 26), directly admitted to hospital (n = 3), and unknown disposition at first hospital (n = 2). Process variables include: intubation in emergency department (ED) given Glasgow Coma Scale (GCS) score < 9 [INTUB], administration of blood products in ED given systolic blood pressure (SBP) < 90 mm Hg [BLOOD], trauma surgeon presence within 5 minutes of patient arrival given GCS < 9 mm Hg or SBP < 90 mm Hg [UNSTABLE-TS], trauma surgeon presence within 5 minutes of patient arrival given injury severity score (ISS) > 15 [ISS-TS], transfer to higher level of care given ISS > 20 and no hypotension [TRAN], transfer to higher level of care given GCS < 9 [TRAN-GCS]. RESULTS: For the high-risk subpopulations, the following Z-scores (with and without an intervention) were found: CONCLUSIONS: Some ATLS interventions (BLOOD, TRAN, and TRAN-GCS) are associated with improved survival for selected high-risk subgroups in these 21 rural Level III trauma hospitals. PMID- 12109574 TI - An emergency medical services program of alternate destination of patient care. AB - OBJECTIVE: The emergency department (ED) is ideally reserved for urgent health needs. The ED, however, is often the site of care for nonurgent conditions. The authors investigated whether emergency medical technicians could decrease ED use by patients with nonurgent concerns who use 911 by appropriately identifying and triaging them to alternate care destinations. METHODS: From August 2000 through January 2001, two King County fire-based emergency medical services (EMS) agencies participated in an alternate care destination program for patients with specific low-acuity diagnosis codes (intervention group). Eligible patients were offered care at a clinic-based destination as an alternate to the ED (n = 1,016). The frequency of the destination of care (ED, clinic, or home) for the intervention group was compared with a matched control group that was comprised of a preintervention historical cohort of EMS encounters from the same two fire based agencies and with the same acuity and diagnosis criteria and seasonal interval (n = 2,617). RESULTS: Compared with the preintervention group, a smaller proportion of patients in the intervention group received care in the ED (44.6% vs. 51.8%, p = 0.001), while a greater proportion of patients in the intervention group received clinic care (8.0% vs. 4.5%, p = 0.001) or home care (no transport) (47.4 vs. 43.7%, p = 0.043). Results were comparable when adjusted for other patient characteristics. Similar relationships were not evident among nonparticipating King County EMS agencies. Based on physician review and patient interview, the alternate care intervention appeared to be safe and satisfactory. CONCLUSION: An EMS-based program may represent one approach to limiting nonurgent ED use. PMID- 12109573 TI - Helicopter transport of pediatric versus adult trauma patients. AB - OBJECTIVE: Conflicting reports exist regarding the appropriate utilization of helicopter transport for victims of trauma. It has been suggested that adult patients are more severely injured compared with pediatric patients when transported by helicopter. The purpose of this study was to determine whether injury severity and survival probability in pediatric trauma patients were similar to those for adults when helicopter transport was utilized at a suburban trauma center. METHODS: The authors conducted a retrospective review of all trauma patients transported by helicopter from the accident scene. Patients were identified from the Christiana Care Health System trauma registry from January 1995 to November 1999. Pediatric patients were defined as those aged 15 years and younger. Data collected were utilized to determine injury severity score (ISS), revised trauma score (RTS), and survival probability. RESULTS: Nine hundred sixty nine patients were transported; 143 were pediatric. There was no statistical difference noted in ISS (14.21 adult, 12.76 pediatric; p = 0.1506) and RTS (7.23 adult, 7.31 pediatric; p = 0.1832). Mean length of stay was less for the pediatric group (7.5 days adult, 5.2 days pediatric; p = 0.008). Survival probabilities were likewise similar for the two groups, yet the difference met statistical significance (0.92 adult, 0.95 pediatric; p = 0.03). CONCLUSION: Pediatric patients transported from the accident scene by helicopter have similar ISSs and RTSs compared with adults. These data suggest that prehospital selection criteria for the two groups are similar. PMID- 12109575 TI - The California 500: medical care at a NASCAR Winston Cup race. AB - BACKGROUND: Stock car racing is America's fastest-growing professional sport. With more than 5.5 million paid admittances and another 148 million watching the 34-race NASCAR Winston Cup series on television, emergency physicians are increasingly called upon to organize medical support for such events. Currently, little reliable information is available to assist in determining what specific personnel and equipment are necessary to optimally support a race event. OBJECTIVE: To characterize the spectrum of presenting injuries and illnesses at a NASCAR Winston Cup event. METHODS: This study was a retrospective review of all patients presenting to nine on-site first aid stations from June 19 to 22, 1997, for the inaugural California 500 race weekend at California Speedway in Fontana, California. Staffing of the nine first aid stations was provided by 20 paramedics, 25 emergency nurses, five emergency physicians, nine advanced life support (ALS) ambulances with two crew members each, and a medically configured helicopter with flight crew. RESULTS: Of the 923 patients seen, 38 were drivers/crew, 230 were track employees, and 644 were spectators. One hundred thirty-six of the patients were treated in the two infield facilities, while 787 were treated in the grandstand first aid stations. Patients seen per hour peaked just before the start of the race at 73 patients seen. Of the ten patients transported to the hospital, three required admission. No deaths occurred. CONCLUSION: These data may assist individuals planning medical support for large motorsports venues. PMID- 12109576 TI - Improvements in prehospital medication storage practices in response to research. AB - Previously the authors showed that prehospital medications were stored outside their recommended temperature range. In response, the state office of emergency medical services (EMS) issued regulations regarding temperature control and monitoring of prehospital medications. OBJECTIVE: To determine the impact of previous research (on medication storage conditions) on current practices among the mobile intensive care units (MICUs) within the state. METHODS: A statewide, structured telephone survey of MICU directors was conducted between April and December 2000. Questions focused on changes in storage and monitoring practices (including modifications to vehicles, medication boxes, and the use of temperature monitoring devices) since the authors' previous research. RESULTS: Thirty-three of 35 (94%) programs (100 vehicles) participated in the survey. Eighty-five percent changed their practices since the research five years ago. Of the five that did not change, three already had temperature control measures in place, while two have not made any changes. Twenty-one (63%) of the programs reported changing specifically because of state regulations. Eighty-one percent of the programs have taken some measure to control temperature. Currently, 63% of the 100 vehicles in use have both heating and cooling devices specifically for the medications, whereas 14% have only a heater and 23% have neither. Thirty-one (94%) MICUs monitor the temperature in some manner: 42% in the vehicle, 58% in the medication box. Of these, 68% are using 30-day electronic temperature data recorders, whereas 32% are using non-recording digital thermometers. CONCLUSIONS: This survey demonstrates a positive impact from previous research. Most of the MICUs in the state have changed their practices in controlling and monitoring prehospital medication storage temperature. PMID- 12109577 TI - Tactical EMS: an emerging opportunity in graduate medical education. AB - OBJECTIVES: Modification of traditional emergency medical services (EMS) principles and procedures for use in the tactical law enforcement setting is emerging as a subspecialty of emergency medicine. Few opportunities exist to train physicians in the principles of tactical medicine, and no studies demonstrate the effectiveness of physician-level training in tactical EMS. METHODS: A standardized eight-hour CONTOMS (Counter Terrorism Operations Medical Support) Physician Awareness course was presented to a group of emergency physicians. The physicians completed an anonymous survey before and after the course, and again four months later. A five-point Likert scale (04) was utilized to measure knowledge and comfort levels among six areas specific to tactical EMS. Changes were measured from precourse scores. RESULTS: Of 39 eligible physicans, 38 completed surveys before and 35 completed surveys after the course, while 16 returned four-month follow-up surveys. Mean scores rose in all areas specific to tactical EMS (p < 0.0005 compared with pretest scores). Mean scores for each topic were: [table: see text] CONCLUSIONS: The COMTOMS Physician Awareness course is effective in increasing physicians' knowledge and comfort levels in areas related to tactical EMS. Knowledge retention at four months is very good. PMID- 12109578 TI - Automated external defibrillator training and skill retention at a ski patrol. AB - OBJECTIVES: To determine whether members of a ski patrol, most of whom have no off-season medical responsibilities, can successfully complete an automated external defibrillator (AED) training program prior to the ski season, and retain AED skills at the end of the season and at the beginning of the following season. METHODS: A prospective educational study was conducted with 61 ski patrol personnel: 51 (84%) had no other medical training, 44 (72%) had no off-season medical duties, and 57 (93%) had no prior exposure to AEDs. Prior to the ski season (December 1, 1998), all members were trained and tested using the standard American Heart Association (AHA) AED training package and a Life-Pak 500 AED and AED Trainer donated by the Medtronic Physio-Control Corporation. Both after the ski season (April 1, 1999) and prior to the following season (October 30, 1999), with no refresher training, participants were retested with the same written and practical exams. Cochrane's linear trend test was used to compare scores on the practical and written tests over time. RESULTS: For the three testing sessions, practical test pass rates were 95%, 92%, and 97%, and written test pass rates were 100%, 98%, and 98%. There was no change in individuals' scores on either the written test (p = 0.914) or the practical test (p = 0.413) over time. CONCLUSIONS: A heterogeneous group of ski patrollers can successfully complete an AED training course, with good skill retention both after the ski season and at the beginning of the following season. PMID- 12109579 TI - Characteristics of twice-transferred, rural trauma patients. AB - OBJECTIVE: Undertriage has seldom been evaluated in the trauma population. In rural states patients often go to the nearest hospital first, where they are evaluated and, if necessary, transferred to another hospital. If they are undertriaged when transferred to the second hospital, they will require a second transfer to a higher-level trauma center. METHODS: The authors retrospectively reviewed the charts of all trauma patients at a level I trauma center from 1996 to 1999 who were seen at two acute care facilities because of a single acute traumatic event before reaching the trauma center. Ninety-three patient charts were analyzed. RESULTS: Forty-six percent of the patients were victims of a motor vehicle crash. Patients were mostly transferred to the level I trauma center for non-spine orthopedic injuries (28%), followed by spine injuries (14%) and head injuries (13%). These patients were stable, as manifested by an average trauma score of 11.6. However, there was a significant positive interaction between injury severity score and time to definitive care. CONCLUSIONS: The authors infer from the data analysis that more serious or complex injuries took longer to evaluate. Since these patients were physiologically stable, reducing the number of twice-transferred trauma patients will involve refining transfer protocols concerning the need for specialty care. PMID- 12109581 TI - Patient restraint in emergency medical services systems. PMID- 12109580 TI - Emergency medical services workforce in the city of Santo Domingo. AB - Santo Domingo is the largest city in the Dominican Republic. In recent years this city has experienced a significant increase in ambulance services. OBJECTIVE: To describe the current emergency medical services (EMS) workforce trends in Santo Domingo. METHODS: This was a cross-sectional descriptive study. Emergency medical services providers working within the city of Santo Domingo filled out a nine item self-administered questionnaire. RESULTS: A total of 101 providers, 48 volunteer and 53 paid, returned the survey (response rates of 75% and 91.4%, respectively). The volunteers showed a mean of 7.48 +/- 2.02 years of involvement in EMS, whereas the paid providers had a mean of 3.58 +/- 2.91 years. When asked about planned long-term involvement in EMS, 93.8% of the volunteers responded positively, compared with 58.5% of the paid group. Nine (16.98%) of the paid providers were not satisfied with their jobs, whereas all the volunteers (100%) stated that they were satisfied; 46 (86.8%) and 27 (50.9%) of the paid providers agreed that better salary and better work hours, respectively, would influence their satisfaction. Among the volunteers, 40 (83.3%) stated that occupational health insurance would improve job satisfaction; and 39 (81.3%) stated that life insurance would bring satisfaction to their job. Logistic regression analysis showed no predictive association between job satisfaction and years of service (p = 0.342). CONCLUSION: Volunteers appeared to be less educated but with a higher long-term interest in EMS. Salary and better work hours seem to be important factors affecting satisfaction of paid providers, whereas occupational health and life insurance appear to affect the satisfaction of volunteers. PMID- 12109582 TI - Law Enforcement Agency Defibrillation (LEA-D): position statement and best practices recommendations from the National Center for Early Defibrillation. PMID- 12109583 TI - Food-dependent exercise-induced anaphylaxis. PMID- 12109584 TI - Street drug toxicity resulting from opiates combined with anticholinergics. PMID- 12109585 TI - Trauma helicopter emergency medical services transport: annotated review of selected outcomes-related literature. AB - Based on its roots in military air evacuation, helicopter emergency medical services (HEMS) has always been emphasized as a tool for trauma transportation. Despite much discussion regarding resource allocation for HEMS, a literature search found little recent systematic review of pertinent studies. As HEMS utilization is subject to increased scrutiny in a health care dollar-conscious environment, it was felt that a compendium of available outcomes-related literature could assist those assessing utility of HEMS trauma transport. The current study utilized a Medline search to identify outcomes studies relative to HEMS trauma transport. The goal of this review is to provide a useful resource for those interested in pursuing systematic review of the HEMS trauma outcomes literature. The primary purpose of the review is bibliographic, but there is editorial comment after each paper's summary. The initial article in this two part series focused on HEMS outcomes literature covering noninjured patients as well as papers assessing outcome in mixed trauma-nontrauma HEMS study groups. PMID- 12109586 TI - An easy, safe, and sure way of open stent grafting: chain-stitch bonding with a balloon catheter. AB - A modified transaortic graft insertion technique with a nephrostomy balloon catheter is presented herein. The graft, which has a Z stent at its end, is bound to the catheter with a chain stitch and then is inserted into the descending aorta under transesophageal echographic observation. Unlacing the chain stitch easily deploys the stented graft. This technique is safer and more reliable than other current methods. PMID- 12109587 TI - Lester M. Crawford, Jr, DVM, PhD. PMID- 12109588 TI - May 2, 2002, statement by David A. D'Alessio, MD, AFMR Secretary-Treasurer to House Appropriations Subcommittee regarding FY 2003 NIH and AHRQ budgets. American Federation for Medical Research. Agency for Healthcare Research and Quality. PMID- 12109590 TI - Classified research is a very small part of NIH bioterrorism effort, Kirschstein tells C-SPAN. PMID- 12109589 TI - Human Research Subject Protections Bills being developed in both houses of Congress. PMID- 12109591 TI - Nitric oxide and chronic gut inflammation: controversies in inflammatory bowel disease. AB - One of the most consistent and dramatic findings in both experimental and human inflammatory bowel disease (e.g., Crohn's disease, ulcerative colitis) is the enhanced expression of the inducible isoform of nitric oxide synthase (iNOS) and the sustained overproduction of the free radical nitric oxide (NO). The role that iNOS-derived NO plays in the pathophysiology of inflammatory bowel disease remains the subject of intense investigation and active debate. Although several different studies using a variety of animal models of acute and chronic gut inflammation suggest that NO may promote intestinal inflammation, an equally impressive number of investigations suggest that iNOS may play no role or may act to attenuate or to limit the extent of inflammatory tissue injury. This review discusses some of the basic concepts related to the immunoregulation of chronic gut inflammation and summarizes the current state of knowledge of the role that NO may play in modulating inflammatory tissue injury. PMID- 12109592 TI - Effect of smoking on hemoglobin A1c and body mass index in patients with type 2 diabetes mellitus. AB - BACKGROUND: Smoking is associated with a decrease in body weight in patients without diabetes mellitus and an increase in insulin resistance and hemoglobin A1c (HbA1c) levels in patients with type 1 diabetes mellitus. Whether smoking is associated with an increase in HbA1c and/or a decrease in body mass index (BMI) in type 2 diabetes mellitus is unresolved, however. Therefore, the objective of this study was to determine the effect of smoking on HbA1c levels and BMI in a cross-section of outpatients with type 2 diabetes mellitus. METHODS: A questionnaire was completed by 102 sequential outpatients (32 men, 70 women) with type 2 diabetes mellitus to assess age, sex, duration of diabetes mellitus, medications, exposure to nicotine, medical complications as a result of type 2 diabetes mellitus, and BMI In addition, a urine sample was obtained from each patient to assess the microalbumin-creatinine and cotinine-creatinine ratios. HbA1c level were also obtained from each volunteer as a measure of glucose control. RESULTS: Smokers had significantly higher cotinine-creatinine ratios than nonsmokers. Smokers and nonsmokers did not differ significantly with regard to HbA1c level, BMI, or duration of diabetes mellitus. Smokers were younger than nonsmokers. Smokers and nonsmokers did not differ significantly in terms of microalbumin-creatinine ratio, amount of diabetic medications, or medical complications. CONCLUSION: The results of this study suggest that smoking does not have a significant direct effect on BMI or HbA1c in patients with type 2 diabetes mellitus. This conclusion suggests that the relationship between these factors is much more complex than in people without diabetes or in patients with type 1 diabetes mellitus. PMID- 12109593 TI - 11beta-Hydroxysteroid dehydrogenase types 1 and 2 are up- and downregulated in cortisol-secreting adrenal adenomas. AB - BACKGROUND: 11beta-Hydroxysteroid dehydrogenase types 1 and 2 (11betaHSD1 and 11betaHSD2) are two isoenzymes that convert inactive glucocorticoids (e.g., cortisone) to their active forms (e.g., cortisol) and vice versa. Abundant evidence indicates that 11betaHSD2 is expressed as mRNA and protein in both adrenal cortex and adrenal tumors. In contrast, 11betaHSD1 has been investigated to a much lesser degree. We therefore studied and compared the expression and activity of the two isoenzymes in the human adrenal cortex (HAC) and cortisol secreting adenomas (CSAs). METHODS: Six HAC and six CSA specimens were studied. 11betaHSD1 and 11betaHSD2 gene expression was studied by conventional and semiquantitative reverse transcription-polymerase chain reaction. 11betaHSD1 and 11betaHSD2 activity was assayed by measuring the capacity of both microsomal fraction and tissue fragments to convert [3H]cortisone to [3H]cortisol and vice versa. Steroid hormones were separated and purified by high-performance liquid chromatography, and cortisol concentration was measured by radioimmunoassay. RESULTS: Semiquantitative reverse transcription-polymerase chain reaction and enzymatic assay demonstrated higher 11betaHSD1 expression and activity and lower 11betaHSD2 expression and activity in CSAs than in HACs. CSA slices secreted larger amounts of cortisol than did HAC specimens, and the cholesterol side-chain cleaving enzyme inhibitor aminoglutethimide, by blocking the early step of steroid synthesis, reduced cortisol secretion by approximately 70%. Aminoglutethimide decreased [3H]cortisol production from [3H]cortisone and increased [3H]cortisone production from [3H]cortisol in both tissues, thereby annulling differences in 11betaHSD1 and 11betaHSD2 activity between HACs and CSAs. CONCLUSION: Collectively, our findings indicate that 1) both 11betaHSD isoenzymes are expressed as mRNA and proteins in the HAC and CSA, with 11betaHSD1 upregulated and 1betaHSD2 downregulated in CSAs; and 2) 11betaHSD1 and 11betaHSD2 activity is positively and negatively correlated with the intracellular concentration of steroid hormones. Hence, 11betaHSD isoenzymes could act as amplifiers of the secretagogue effect of agonists and could contribute to the elevated hormonal secretion of CSAs. PMID- 12109594 TI - Human brain nascent polypeptide-associated complex alpha subunit is decreased in patients with Alzheimer' s disease and Down syndrome. AB - Nascent polypeptide-associated complex (NAC) protein, a heterodimeric complex of alpha- and beta-subunits, prevents mistargeting of nascent polypeptide chains to the endoplasmic reticulum membranes. alpha-NAC has sequence similarities with transcription-regulating proteins and has been reported to function as a transcriptional coactivator potentiating c-Jun-mediated transcription. Performing gene hunting using differential display-polymerase chain reaction, a downregulated sequence in the frontal cortex of patients with Alzheimer's disease (AD) and Down syndrome (DS) with AD-like neuropathology was identified as a-NAC with 100% homology. The significant decrease in alpha-NAC mRNA was shown by semiquantitative reverse transcription-polymerase chain reaction, and in parallel, the significant decrease of alpha-NAC protein, which was even more pronounced when related to either actin or neuron-specific enolase levels, was also observed in both disorders. Linear regression analysis revealed a strong, significant correlation between alpha-NAC protein and mRNA expression. In fetal DS brain, however, mRNA levels of alpha-NAC were comparable between DS and controls, suggesting that the decrease in alpha-NAC might be involved in the pathology of neurodegenerative diseases. The decrease in alpha-NAC as a transcriptional coactivator could contribute to the characteristic decline of the c-Jun-mediated transcriptional machinery and could function as the complementary mechanism in c-Jun-mediated apoptosis. Decreased alpha-NAC may result in the mistargeting, mistranslation, and proteolysis of proteins by affecting overall NAC function. PMID- 12109595 TI - Infectious hepatitis C, hepatitis G, and TT virus: review and implications for dentists. AB - In the past 10 years, hepatitis C and G viruses have been identified, and in the last two years a further parenterally transmitted agent, termed TT virus (TTV), has been discovered. These viruses have a worldwide distribution and frequently cause chronic infection. The purpose of this article was to promote an understanding of these viral agents and their relevance in dental practice. Infected patients may develop a chronic carrier state without clinical disease or may develop liver disease, and may have related oral conditions. Dental providers will see a growing number of patients with HCV/HGV and possibly TTV infection. All of these patients require appropriate infection control measures during dental treatment. PMID- 12109596 TI - Mechanical and chemical home plaque control: a study of Brazilian children and adolescents with disabilities. AB - The purpose of this study was to evaluate the compliance of children and adolescents with disabilities with a home plaque control program. A random sample of 52 students between 7 and 21 years old was solicited from a special school, in Belo Horizonte, Brazil. Measurement of the Debris Index and the presence of bleeding on light probing of the buccal gingival papillae were included as part of the initial clinical examination. Plaque control instructions were given during the first and subsequent clinical sessions. During the fourth clinical session, the patients who had more than 25% sites with bleeding papillae on light probing were placed in a separate group (Group 2) which used a mechanical and chemical plaque control protocol. Group 1 consisted of persons who used mechanical plaque control only. All of the patients were followed up for two more recalls (days 51, 81) during which they were given plaque control instruction and had another gingival examination. On day 111, the final Debris Index and gingival examination were carried out. A significant reduction (p < 0.02) was found between the first and final Debris Index recordings in both groups. Subjects in Group 1 had a significant reduction (p < 0.001) when the first gingival examination was compared with days 21, 51, 81, and 111. Group 2 showed significant reduction (p < 0.01) in gingival bleeding when day 21 was compared with days 81 and 111. Our study suggests that it is possible for these children/adolescents and their parents to learn and comply with mechanical and chemical plaque control at home. PMID- 12109597 TI - Prevalence of dental caries in an adult population with mental disabilities in Spain. AB - The objective of this study was to analyze the prevalence of dental caries and the possible influence of extraoral factors in a group of adults with a mild to moderate degree of developmental delay. A total of 166 subjects ranging in age from 20 to 40 years were examined. All subjects were institutionalized, and 70.4% (n = 117) participated in a preventive program designed by the educators of the center. This consisted of weekly fluoride mouthrinses (0.2%) and control of dental plaque by toothbrushing with a fluoride toothpaste. The mean DMFT index for the whole sample was 5.97. Subjects in the oldest age group had the highest DMFT index and the lowest filled component. There were no significant differences in the DMFT index and its components between the subjects who received a weekly fluoride mouthrinse (n = 117) and those who did not (n = 49). Comparison of subjects who had cerebral palsy, Down Syndrome, and idiopathic developmental delay (n = 157) found a significantly lower DMFT index (p < 0.05) in the persons with Down Syndrome compared with the other special-needs groups. Other factors which could influence the results-such as fluoride use, oral hygiene, and administration of benzodiazepines-were also analyzed. An analysis of the results revealed a moderately high prevalence of caries in the whole sample that was lower than that in a national survey. This could be attributed to strict dietary control of sugar in the study population. PMID- 12109598 TI - Sickle cell anemia and dental caries: a literature review and pilot study. AB - The purpose of this cohort study was to determine whether individuals with sickle cell anemia (SCA) were more susceptible to dental caries than non-sickle-cell control subjects. A review of the literature suggests several reasons why individuals with SCA may be at increased risk. Thirty-five cases of SCA aged 6 years and older were identified from a screening of 15,900 current patient files at the Howard University College of Dentistry Dental Clinic. A total of 140 non SCA control subjects (four per case), frequency-matched on enrollment period (+/- 5 yrs) and age (+/- 2 yrs if under age 21, or +/- 5 yrs if 21 or over), was selected by a nonbiased method from the same dental clinic files. SCA cases and controls were identical on mean age (30.4 +/- 19 yrs, ranging from 5 to 92 yrs) and were similar in sex distribution (males: 34% of SCA cases, 40% of controls). The mean number of permanent teeth present was very similar for SCA cases and controls (23.4 +/- 6.4 vs. 24.2 +/- 6.4). The mean DMFT was 21% higher in the SCA cases (12.0 +/- 8.4 vs. 9.9 +/- 6.9), and the mean DMFS was 26% higher in the SCA cases (33.0 +/- 32.3 vs. 26.2 +/- 27.7). While there was virtually no difference in DMFS between SCA cases and controls for 6- to 19-year-olds, for subjects aged 20 and older, the DMFS was 30.4% higher in the SCD cases. For all ages, the M component for SCA cases was 40.7% higher, and the D component was 20.0% higher, while the F component was only 3.5% higher than for controls. Untreated decay (the D/D+F surfaces ratio) was 24.4% higher in the SCA cases. The findings from this pilot study suggest that SCA cases have a higher susceptibility to dental caries and/or that SCA patients may have different treatment pathways once caries is detected. While none of the observed differences was statistically significant, these findings were of clinical interest and should be pursued in future large-scale studies. PMID- 12109599 TI - A comparison of perceived injection pain with and without anesthetic between elderly and young female subjects. AB - Several reports have indicated that the perception toward pain declines with age. However, since pain thresholds have been reported to be lower in females than in males, studies to detect pain perception may be more sensitive in females. In dental care, the injection of local anesthetics with a needle is used to abrogate subsequent pain from treatment. However, the anomaly of using one potentially painful procedure to prevent subsequent pain often hinders patients from seeking dental care. The purpose of this study was to evaluate the perception of pain from needle-sticks with and without anesthetic between young and elderly female subjects. We compared the perception of pain stimulated by needle-sticks without (NS) and with (NS-I) 0.3 mL of local anesthetic injected into the oral mucosal tissues of a group of 20 young females (aged 19-24 yrs) and a group of 18 elderly females (aged 70-75 yrs). A Verbal Pain Scale (VPS; descriptors assigned, 0-4) and a Visual Analogue Scale (VAS; 0-10 cm) were used to measure pain intensity. Statistical analyses by ANOVA and Fisher's Exact Test were used where appropriate. The VPS and VAS scores for perception of pain in elderly females were lower than the scores from the younger subjects. No differences between Ns and Ns-I in either age group were found. Study limitations included potential periodic hormonal imbalances in both groups of females as well as untested diminished cognitive function in elderly subjects. However, we conclude, with the noted cautions, that pain perception between the two groups was not significantly different. PMID- 12109600 TI - Relative foster parents of HIV-affected children. AB - The HIV epidemic is one of the reasons for an increase in relative foster care. Most children of HIV-infected parents are not infected, but both the children and their caregivers are affected by the parent's illness. Twenty-eight caregivers participated in an exploratory, qualitative study of the permanency planning needs of HIV-affected relative foster parents. Of the caregivers, 17 disclosed that the foster child's parent was HIV-infected; 11 caregivers did not report HIV infection in the immediate family. HIV-affected caregivers' greatest need was a more adequate response from social workers and therapy services for the children; nonaffected caregivers needed financial assistance most. More HIV-affected caregivers were considering permanency than nonaffected caregivers. Child welfare workers and relative foster parents need training on HIV's effect on the families so that appropriate services can be accessed as early as possible. PMID- 12109601 TI - Problem-solving communication in foster families and birthfamilies. AB - This study assessed child behavior problems and parent-child communication behaviors during problem solving in three groups of families with adolescent children: foster families, birthfamilies with a child at risk for behavior problems, and birthfamilies with a child not at risk. Levels of positive and negative communication behaviors in the foster families were similar to those in the lower-risk families and were significantly related to foster child behavior problems. PMID- 12109602 TI - Parental visiting and family reunification: could inclusive practice make a difference? AB - This study examines whether inclusive practice, or parental involvement in foster children's lives while in placement, is correlated with more frequent visiting and a greater likelihood of reunification. This hypothesis was tested among a random sample of 230 twelve- and thirteen-year-olds placed in traditional family foster care. Results suggest that mothers who visit their child and are involved in case reviews and child care activities visit more frequently than mothers who visit in settings such as agency offices and have no other types of involvement. In addition, visiting frequency is highly predictive of reunification. These associations were not explained by maternal substance abuse, mental illness, or the child's placement history. PMID- 12109604 TI - Barrett's oesophagus--are British gastroenterologists denying their patients prevention of malignant change? AB - Adenocarcinoma of the lower oesophagus is rapidly increasing in industrialised countries. The importance of Barrett's oesophagus is because of the potential for it to progress to oesophageal adenocarcinoma. It has a strong correlation to chronic GORD. Symptomatic patients or those with a long segment, if dysplasia is present or the patient is under 50 years of age, should be offered anti-reflux surgery. Patients may be denied the procedure by some gastroenterologists PMID- 12109603 TI - Services for maltreated children: variations by maltreatment characteristics. AB - Mental health services were the most frequently recommended service for maltreated children known to a child protective service agency, but more than one fourth of the children with serious behavioral problems did not receive referrals for such services. Children with long histories of maltreatment were more likely to be referred for some services, especially out-of-home care. Gaps in services remain, underscoring a need for research focused on the need for interventions targeted to maltreated children involved with child protective service agencies. PMID- 12109606 TI - Whipple's resection-proximal pancreaticoduodenectomy (PD). AB - Pancreatic resection offers the potential for long-term cure in 15% of patients with pancreatic cancer. This article describes the author's technique of pancreaticoduodenectomy (PD), together with guidelines for disease staging, pre operative work-up and patient selection. The role of neo-adjuvant and adjuvant chemotherapy is currently under evaluation and all patients who have a curative resection should be considered for entry into the ESPAC 3 trial that aims to establish the definitive role of adjuvant chemotherapy in pancreatic cancer. PMID- 12109605 TI - Endovascular repair of abdominal aortic aneurysm: current status. AB - INTRODUCTION: Endovascular aneurysm surgery (EVAR) was introduced a decade ago. Early results are promising, however, there remain concerns regarding the longer term durability of this technique. Consequently, the national multi-centre EVAR trial has been commenced to define the role of endovascular surgery in the management of abdominal aortic aneurysm. DISCUSSION: Successful EVAR requires accurate pre-operative assessment of aneurysm morphology. Current stent-grafts allow 60% of all infra-renal AAA to be treated. Reduced physiological stress and low peri-operative morbidity and mortality rates have been demonstrated with this technique when compared to open repair. Endoleak is an Achilles heel of EVAR, although in itself does not accurately predict outcome. First and second generation devices are estimated to have a 1% per year risk of rupture. CONCLUSIONS: Increased understanding of the issues surrounding aneurysm morphology and successful stent-grafting have allowed a major reduction of early type I endoleak. Late endoleak and graft migration remain problematic. Type I and III endoleaks are risk factors for subsequent rupture although the significance of type II endoleak remains uncertain. More robust indicators of outcome success/failure are required so that follow-up may be rationalised. PMID- 12109607 TI - Internal fixation of intracapsular fractures. AB - The main indications for internal fixation of intracapsular fractures are undisplaced and minimally displaced fractures and displaced intracapsular fractures in those aged less than about 70 years. For displaced fractures, closed reduction is to be preferred to open reduction. Numerous different implants may have to be used; with current practice favouring two or three parallel cannulated cancellous screws. These may be inserted either percutaneously or with minimal surgical exposure. Attention to surgical details of fracture reduction and implant positioning will minimise the risk of fracture healing complications. Post-operative care should generally be unrestricted mobilization with weight bearing as tolerated. PMID- 12109608 TI - Short and long-term outcome of pancreatic surgery in a district general hospital. AB - Pancreatic surgery is a formidable undertaking with historically high mortality and poor prognosis for periampullary lesions. This has led to recommendations that all pancreatic surgery should be performed in specialist centres. There is no doubt from large series that a low mortality can be achieved in these centres, but there has been no direct comparison between results from these specialist centres and district general hospitals with an interest in pancreatic disease. We present a retrospective, seven-year experience with a 3% 30 day mortality, 39% morbidity and 14 month median survival for malignant disease. Comparison with the UK survey of specialist pancreatic units shows that pancreatic surgery can be safely performed in the setting of a district general hospital with low morbidity and mortality, and good long-term outcome. PMID- 12109609 TI - Predictors of excessive blood loss during operative treatment of hip fractures. AB - OBJECTIVES: To determine the total blood loss and transfusion needs during operative treatments of hip fracture, and identify predictors of excessive blood loss. METHOD: A prospective study of 242 consecutive patients operated for hip fractures over a 6 month period. The main outcome measure of blood loss was assessed by blood volume in the drainage system and swab weight. A loss of more than 480 mls was considered as excessive blood loss. RESULTS: The study consisted of 190 women and 52 men, mean age was 81.6 years (range 44-99). More than one third of patients (34%) lost more than 480 mls of blood, and mean units transfused per patient was 2.3. Univariate predictors of increased blood loss were patients of American Society of Anaesthesiology (ASA) grade III and IV, patients at risk of cerebral or cardiac ischemia from volume depletion as defined by the American College of Physicians, patients with two or more pre-existing medical conditions and patients who had a hemiarthroplasty carried out. However, with subsequent multivariate analysis, patients who had undergone a hemiarthroplasty and those at risk predicted increased blood loss. CONCLUSION: Pre-operative characteristics can help determine which patients should have either blood requested on the day of surgery (group and cross-match) or the customary group and save policy. PMID- 12109610 TI - Right hemicolectomy: mechanical bowel preparation is not required. AB - BACKGROUND: Mechanical bowel preparation before colonic surgery is widely advocated but remains controversial. Recent guidelines published by the Clinical Standards Board for Scotland recommend mechanical bowel preparation prior to surgery for all colorectal cancers but this may be inappropriate. This study examines the outcome of a policy of no mechanical preparation before elective right hemicolectomy. METHOD: Data on 102 consecutive patients undergoing elective right or extended right hemicolectomy for colonic adenocarcinoma were extracted from a prospective database. RESULTS: No clinical anastomotic leaks were observed. Two patients developed wound infections and one patient died with no autopsy evidence of anastomotic leak. CONCLUSION: Mechanical bowel preparation can safely be omitted prior to right hemicolectomy in patients with colonic cancer. PMID- 12109611 TI - Results of inguinal canal repair in athletes with sports hernia. AB - AIMS: To evaluate the role of surgical exploration and repair of the inguinal canal in athletes suspected of having a sports hernia. METHODS: Thirty-five (34 males, 1 female) athletes with a suspected sports hernia underwent surgical exploration and inguinal hernia repair. After six months, all athletes were sent questionnaires to assess any improvement in analogue pain scores, return to sport, recurrence of symptoms and the overall result of surgery. RESULTS: Operative findings revealed a tear in the external oblique aponeurosis with or without a significant posterior bulge (n=20), a lone significant posterior bulge (n=10), a tear in the conjoint tendon with dilated superficial ring (n=3), small direct hernial sac (n=1) and lipoma of the spermatic cord (n=1). Surgery consisted of repair of external oblique tear (when present) and prolene darn or lichtenstein mesh repair of the posterior inguinal canal. Twenty-seven patients replied to the questionnaire giving a response rate of 78%; of these, 25 patients (93%: 95% CI 83-100) had returned to normal athletic activities at pre-injury level. There was a marked improvement in level of pain (median pain level=8 pre operative vs 2 post-operative, p<0.001). Eleven patients (41%) rated the results as excellent, eleven (41%) as good and five (18%) as fair and none worse. Six patients complained of occasional discomfort related to the scar. Three patients complained of recurrence of their symptoms after 4 to 5 months following strenuous exercise. CONCLUSION: Sports hernia presents with a spectrum of surgical findings. Athletes with sports hernia should be considered for routine hernia repair, as the majority of the patients benefit from surgery. It is important to offer a structured rehabilitation programme to maximise the benefits of surgery. PMID- 12109612 TI - The Myles Gibson military lecture: surgery in the Napoleonic Wars. AB - Throughout the twenty-two year period of the Napoleonic Wars, campaigns under extremes of climate cost the Allies much in terms of mortality and morbidity. Although Bonaparte brought about many sound political and national improvements, when France had been brought to its knees by the bloody Revolution, his ambitions became excessive and his military forays difficult to support. Following early successes in the field, he underestimated the determination, persistence and the ability of some opposing commanders. The French medical services profited greatly from the innovations of the post-revolutionary period, and the efforts of men such as Larrey and Percy. The British Army medical support was scanty, and, initially lacked experience. To some extent, this latter defect was corrected by Sir James McGrigor during the Peninsular War. Each campaign brought it's own perils and most men died of deprivation, disease and effects of climate, rather than battle injury. There were technically able surgeons who were inevitably hampered by lack of antiseptic technique, anaesthesia and the lack of understanding of the fundamental aspects of hygiene, adequate diet and good nursing care. PMID- 12109613 TI - Cocoon abdomen in a liver transplant patient. AB - Cocoon abdomen is a rare cause of intestinal obstruction and its pathogenesis remains unclear. This is probably the first report of the development of such a condition in a liver transplant recipient. The management of this patient is discussed and the current literature is reviewed PMID- 12109614 TI - Anterior sacral meningocoele presenting as a peri-anal abscess. AB - Anterior sacral meningoceole is a rare occurrence and presentation as a perianal abscess has not been previously reported. The case is presented and the condition discussed. The potential risks of failing to establish the diagnosis, prior to surgery, are outlined. PMID- 12109615 TI - Potentially fatal oro-facial infections: five cautionary tales. AB - Five cases of oro-facial infection leading to life-threatening complications are reported. Although all had been treated with antibiotics prior to maxillofacial referral, lack of surgical intervention had allowed progressive infection. The importance of resuscitation, supportive therapy, extraction of involved teeth to remove the source of infection and drainage of pus is emphasised PMID- 12109616 TI - Risk group-based management of differentiated thyroid carcinoma. PMID- 12109617 TI - GH/IGF-I axis in severe systemic disorders. PMID- 12109618 TI - Thyroid function in breast-fed infants whose mothers take high doses of methimazole. AB - Recently, a few studies have shown the safety of methimazole (MMI) therapy of thyrotoxic lactating mothers on thyroid function of their infants. However, it is not known whether the effect of moderately high doses of MMI therapy on lactating mothers can be dangerous for breast-fed infants. Eighty-eight thyrotoxic lactating mothers and their infants were studied. 46 received 20 mg MMI and 42 were given 30 mg MMI during the first month, 10 mg for the second and 5-10 mg for additional 10 months of therapy. Serum T4, T3 and TSH concentrations and in hyperthyroid MMI treated mothers and their RT3U were measured in hyperthyroid MMI treated mothers and their infants, before and at 1, 2, 6, and 12 months after initiation of therapy. Serum MMI was measured in the infants of thyrotoxic mothers taking 20-30 mg MMI. Mean+/-SD of free T4 index (FT4I) in thyrotoxic mothers treated with 20 and 30 mg MMI for one month decreased from 20.1+/-4.2 to 9.7+/-1.5 (p<0.001) and from 20.6+/-4.8 to 8.6+/-3.0 (p<0.001), respectively. Values for free T3 index (FT3I) decreased from 587+/-53 to 180+/-39 (p<0.001) and from 610+/-49 to 151+/-31 (p<0.001) in those treated with 20 and 30 mg MMI, respectively. By the end of one month 5 had elevated FT4I or FT3I or both and 12 had elevated TSH. The dose of MMI was adjusted and thyroid function remained normal up to 12 months of MMI therapy in thyrotoxic lactating mothers. Serum T4, T3 and TSH concentrations of breast-fed infants were normal before and up to 12 months of MMI therapy of their breast-feeding mothers. The lowest T4 and T3 and the highest TSH values were 101 nmol/l, 1.8 nmol/l and 4.1 mU/l, respectively. Serum MMI levels were <0.03 in 7 and 0.03, 0.034 and 0.035 microg/ml in the other 3 infants. We conclude that the treatment of hyperthyroid lactating mothers with doses of 20-30 mg MMI day does not cause deleterious effects on thyroid function of their breast-fed infants. PMID- 12109619 TI - Three-year prospective evaluation of glucose tolerance, beta-cell function and peripheral insulin sensitivity in non-diabetic patients with thalassemia major. AB - The aim of this prospective study was to evaluate the evolution of glucose tolerance (GT), insulin secretion and peripheral insulin sensitivity during a 3 yr follow-up in a homogenous population consisting of fourteen non-diabetic adults with thalassemia major (TM). All the patients underwent 2 OGTTs with a 3 yr interval and random measurements of fasting glycemia during the entire follow up. At the time of both OGTTs, peripheral insulin sensitivity was assessed by both homeostatic model assessment (HOMA) index and a novel index derived from the OGTT. At the second OGTT patients exhibited both significantly higher fasting glucose concentrations and enhanced glycemic responses, with greater average glucose areas. GT deterioration over time was accompanied by a reduction of insulin sensitivity, with no concomitant change of insulin secretion. No patient developed diabetes mellitus (DM) during follow-up. To conclude, the natural history of glycometabolic status in TM adults seems to be characterized by a GT deterioration over time, which may probably reflect an increase of insulin resistance. GT deterioration is more evident in patients with the highest responses to the 1st OGTT and particularly in those with pre-existing impaired GT. PMID- 12109620 TI - Effects of two different somatostatin analogs on glucose tolerance in acromegaly. AB - Impaired glucose tolerance is present in many acromegalic patients and treatment with somatostatin analogs has variable effects on glycemic control. The aim of this study was to compare the effects of 2 somatostatin analogs on glucose metabolism, lanreotide slow release (L-SR) and octreotide long acting release (O LAR), in 10 patients with acromegaly (2 of whom with overt Type 2 diabetes mellitus). Glucose and insulin levels in fasting conditions and in response to OGTT, evaluated as AUC, insulin resistance (IR) evaluated by homeostatic model assessment (HOMA-IR), glycosylated hemoglobin (HbA1c), GH, IGF-I, were assessed during L-SR and O-LAR treatment. Mean fasting glucose, glucose response to OGTT and HbA1c levels in 8 non-diabetic patients did not significantly change after L SR therapy while they all increased after O-LAR treatment (p<0.05 vs baseline and L-SR). Mean HOMA-IR values calculated in acromegalic patients before medical therapy were higher than in normal subjects (p<0.005) and showed a significant decrease during both treatments (p<0.05). In the 2 diabetic acromegalic patients a worsening in glucose metabolism was observed during O-LAR treatment but not during L-SR. GH and IGF-I levels significantly decreased with both drugs and normalized respectively in 38% and 12% with L-SR, 50% and 25% with O-LAR. In conclusion, both drugs decreased IR in acromegalic patients; O-LAR seems to be more detrimental to glucose metabolism than L-SR, despite being more effective in reducing GH and IGF-I levels. PMID- 12109621 TI - MEN1 gene alterations do not correlate with the phenotype of sporadic primary hyperparathyroidism. AB - Loss of heterozygosity (LOH) in the MEN1 region on chromosome 11q13 and MEN1 gene mutations have been found in a subset of sporadic parathyroid tumors. The question of whether these genetic abnormalities in the parathyroid tumors might influence the clinical and biochemical characteristics of primary hyperparathyroidism (PHPT) remains to be elucidated. The aim of the present study was to correlate the presence of MEN1 gene alterations in PHPT tumors with the clinical phenotype. Using microsatellite analysis for LOH at 11q13 and DNA sequencing of the coding exons, the MEN1 gene was studied in 38 parathyroid tumors of patients with sporadic PHPT. Fourteen tumors showed LOH at 11q13, and mutations of MEN1 gene were detected in 7 cases. The clinical and biochemical characteristics of patients were unrelated to the presence or absence of LOH and/or MEN1 gene mutations. In conclusion, MEN1 gene alterations are rather common in sporadic PHPT and their presence does not correlate with the clinical manifestations of the disease. PMID- 12109622 TI - Post-partum thyroiditis in a mediterranean population: a prospective study of a large cohort of thyroid antibody positive women at the time of delivery. AB - Post-partum thyroiditis (PPT) is a common disease with important clinical sequelae. Its clinical features, association with antimicrosomal antibodies (TPOAb) and long-term prognosis are not thoroughly described in Mediterranean populations. The aims of this study were: 1) to describe clinical features of PPT in a Greek population, 2) to associate TPOAb with PPT presence, and 3) to identify long-term prognosis of PPT. 1,594 consecutive post-partum women were studied within 3 days of their delivery. Women with positive TPOAb were followed up for one yr for clinical and biochemical evidence of PPT. Incidence of positive TPOAb was 5.2% whereas incidence of PPT was 2.4%. PPT was presented as hyperthyroidism alone (18%), hypothyroidism alone (40%) and hyperthyroidism followed by hypothyroidism (42%). A titer of 1:1600, immediately post-partum, could predict PPT with 97% sensitivity and 91% specificity. History of PPT in previous pregnancies was associated with PPT in the current pregnancy. This association did not exist with maternal age, gestation duration, parity, abortions, sex of the newborn, birth-weight or smoking. Long-term hypothyroidism developed in 21% of women. Observation and post-partum thyroid function testing, where PPT is suspected, seems to constitute the best clinical approach. There is need for long-term follow-up in PPT patients. PMID- 12109623 TI - Effect of long-term strength training on glucose metabolism. Implications for individual impact of high lean mass and high fat mass on relationship between BMI and insulin sensitivity. AB - The aim of this study was to examine the independent effect of high lean mass on glucose metabolism, as well as its consequences on the classic relationship between BMI and insulin sensitivity (SI) in 3 groups: 1) 8 strength-trained males with BMI >27 kg/m2 (athletes); 2) 10 sedentary males with BMI >27 kg/m2 (obese); and 3) 12 sedentary males with BMI 22-25 kg/m2 (control). Body composition was measured with impedance analysis. Iv glucose tolerance test was performed at 09:00 h after overnight fast. Estimation of insulin sensitivity and glucose effectiveness by Minimal Model Approach. Plasma glucose and insulin determination by glucose-oxidase and RIA respectively. BMI and lean mass (LM) were greater in athletes than in controls, but there were no differences in fat mass (FM), basal glucose (Gb), basal insulin (Ib), glucose tolerance (Kg), SI, glucose effectiveness (Sg), acute insulin response to glucose (AIRG) and leptin. Obese showed greater FM, leptin, lb and AIRG than athletes, while SI was lower; BMI, LM, Gb, Kg and Sg were similar. BMI, FM, LM, Ib, AIRG and leptin were lower in controls than in obese, while SI index was greater; Gb, Sg and Kg were similar. We found that: 1) Resistance exercise does not modify glucose effectiveness, but can improve insulin sensitivity through FM reduction (LM augmentation alone has no effect on glucose metabolism); and 2) High BMI causes insulin resistance only if it depends on adipose tissue hypertrophy. PMID- 12109624 TI - Impact of cervical lymph node dissection on serum TG and the course of disease in TG-positive, radioactive iodine whole body scan-negative recurrent/persistent papillary thyroid cancer. AB - In the management of papillary thyroid cancer (PTC), surgery is indicated for locoregional recurrent/persistent disease. In this study, we examined the effect of such surgery on serum TG and the course of the disease in 21 patients with PTC (mean age 38.5 yr), who after the initial surgery and radioactive iodine (RAI) ablation developed high TG (>10 ng/ml) and negative 123I whole body scan (DxWBS). All patients had neck persistent/recurrent PTC that was confirmed by ultrasound guided fine needle aspiration. Prior to neck re-exploration, radiological studies (chest X-rays, CT scan of the chest, and fluoro-18-deoxyglucose positron emission tomography [FDG-PET]) showed no evidence of distant metastases. TG autoantibodies were negative in 19 patients. Second surgery consisted of unilateral (13 patients) or bilateral (8 patients) modified neck dissection. The mean+/-SE TG prior to neck re-exploration was 184.8+/-79.0 ng/ml and declined after surgery to 127.5+/-59.0 ng/ml (p=0.25). The corresponding TSH values were 150.6+/-23.0 and 143.4+/-20.0 mU/l, respectively (p=0.34). After a mean follow-up of 20.7+/-3 months, TG increased to 168+/-68.0 ng/ml. This increase, however, was NS (p=0.67). The corresponding TSH values were 143.4+/-20.0 and 132.0+/-22.0 mU/l (p=0.27). Following second surgery, only 4 patients achieved remission, the other 17 patients received one or more of the following therapies; RAI (10 patients), third surgery (5 patients), and/or external radiation (7 patients). Thirteen patients continued to have persistent disease and 4 patients showed progressive course of their disease (distant metastases or grossly palpable neck disease). In conclusion, second surgery for recurrent/persistent PTC leads to remission in only a minority of cases but the course of the disease tends to be stable in most cases. PMID- 12109625 TI - Gender- and age-related differences in the endocrine parameters of acromegaly. AB - Acromegaly is a severe slow-developing disease associated with a poor prognosis for cardiovascular disease. To evaluate the impact of age and gender on the severity of the disease, 151 de novo patients with acromegaly (79 women, 72 men, age range 19-77 yr) were included in this open retrospective multi-center cohort study. Basal GH and IGF-I levels, GH response after glucose load and maximal tumor diameter at MRI were measured in all patients at diagnosis. Fasting GH levels and maximal tumor diameter were similar in women and men, while serum IGF I levels were lower (664.9+/-24.9 vs 755.9+/-32 microg/l; p=0.02) and GH nadir after glucose load was higher (27.5+/-3.7 vs 18.5+/-2.2 microg/l; p=0.04) in women than in men. In both sexes, patients' age was negatively correlated with basal and nadir GH, IGF-I levels and tumor size; fasting GH levels were positively correlated with IGF-I levels and nadir GH after glucose. No interaction between age and gender was found on biochemical and morphological parameters. At diagnosis, elderly patients with acromegaly have lower GH and IGF I levels, lower GH nadir after glucose load and smaller adenomas than young patients. Women have lower IGF-I levels but higher GH nadir after glucose load than men. These age and gender differences should be considered to appropriately evaluate the activity of acromegaly throughout a life-span. PMID- 12109626 TI - Bisphosphonates influence the proliferation and the maturation of normal human osteoblasts. AB - The key pharmacological action for the clinical use of bisphosphonates lies in the inhibition of osteoclast-mediated bone resorption. Osteoblasts could be other target cells for bisphosphonates. We studied the effects of bisphosphonates on the proliferation and the differentiation of normal human bone trabecular osteoblastic cells (hOB). We tested 4 different compounds: clodronate, pamidronate and 2 newer compounds: ibandronate, a nitrogen-containing bisphosphonate and zoledronate, which is a heterocyclic imidazole compound. Ibandronate and zoledronate stimulated hOB cell proliferation by up to 30% (p<0.05) after 72 h for concentrations ranging from 10(-8) M to 10(-5) M. Clodronate transiently enhanced hOB cell survival after only 24 h (+60%, p<0.001) whereas pamidronate had no effect. Longer time course studies, in presence of fetal calf serum, revealed that cell growth was finally reduced by all 4 bisphosphonates (40% after 7 days). Type I collagen synthesis was transiently increased by all 4 bisphosphonates after only 48 h incubation (+17% to +67%, p<0.05). Clodronate increased ALP activity by up to 1.7-fold after 4 days of culture (p<0.05) whereas ibandronate or zoledronate exhibited lesser stimulatory effects (+17 to +30%), and pamidronate had no significant effect. In conclusion, we found that different bisphosphonates, currently used or tested in various clinical conditions, transiently stimulated the growth of preosteoblastic cells and thereafter increased their differentiation according to sequential events (type I collagen synthesis first, then ALP activity to a lesser extent). Our data suggest that the beneficial effects of bisphosphonate treatment on bone mass and integrity could be partly mediated through a direct action on osteoblasts. PMID- 12109627 TI - Individual effect of E2 and dydrogesterone on insulin sensitivity in post menopausal women. AB - The aim of this study was to evaluate the impact of a three-month continuous administration of oral E2, alone, or combined with 2 different dosages of dydrogesterone, on the glucose tolerance and insulin sensitivity in postmenopausal women. In a prospective placebo-controlled study, 43 normal weight and normoinsulinemic women were randomized to receive either 2 mg of oral 17beta E2 daily (group A), or 2 mg E2 daily plus 5 mg daily oral dydrogesterone, from day 14 to 28, in a sequentially combined regimen (group B), or 2 mg of E2 and 10 mg dydrogesterone in the same sequentially combined regimen (group C) or placebo for 12 weeks. An OGTT and a euglycemic hyperinsulinemic clamp were performed before and after treatment. Serum glucose and insulin concentrations were measured both in fasting conditions and after OGTT. C-peptide pancreatic secretion was tested only in fasting conditions. Total body glucose utilization (M), for insulin sensitivity evaluation, was determined in each subject. Postmenopausal women treated with unopposed 17beta E2 (group A) showed a slight but statistically significant decrease of insulin sensitivity (p<0.05). A more marked deterioration of the same parameter was observed in the 2 groups treated with E2 plus dydrogesterone (group B and group C: p<0.01). Post hoc testing for the percent change from baseline indicated that group A significantly differed from group C (p<0.05) and all treated groups significantly differed from the placebo group (p<0.01). Finally, after treatment in group C, a significant reduction of insulin and an increase of glucose responses to OGTT (p<0.01) were observed. These results indicate that, in a short-term period, the use of 17beta E2 and overall 17beta E2 plus dydrogesterone, even with the reduction of insulin plasma levels, might cause a decrease in insulin sensitivity in normal weight and normoinsulinemic post-menopausal women. PMID- 12109629 TI - A novel splicing defect (IVS6+1G>T) in a patient with pseudovitamin D deficiency rickets. AB - A 15-month-old boy with severe rickets, that by clinical analysis was diagnosed as affected by hereditary pseudovitamin D deficiency rickets (PDDR), was evaluated for mutations in the 25OHD3 1alpha-hydroxylase gene. Molecular analysis showed a double heterozygous state for a novel splicing mutation in the invariant dinucleotide of the donor site of IVS6 and a 7 nucleotide insertion in the exon 8, which is common in different ethnical backgrounds. PMID- 12109630 TI - Post-partum hypocalcemia: idiopatic hypoparathyroidism manifested early in lactation. AB - Hypocalcemia associated with labor and lactation is a rare condition reported previously in patients with hypovitaminosis D. We here describe a case of a young woman in whom symptomatic severe hypocalcemia appeared after her second delivery, early in lactation. At the end of lactation the condition worsened. We review all previously reported cases and suggest a possible physiologic explanation for the association between pregnancy, lactation and the appearance of symptomatic hypocalcemia. PMID- 12109628 TI - Glucagon administration elicits blunted GH but exaggerated ACTH response in obesity. AB - Reduction in both spontaneous and stimulated GH secretion in obesity has been clearly demonstrated. Mild hyperactivity of hypothalamus-pituitary-adrenal (HPA) axis has been also reported. Glucagon, at least after im administration, induces clear increase in either GH or ACTH and F levels but its effect on somatotroph and corticotroph secretion in obesity has never been studied. In 7 patients with abdominal obesity (OB, aged 24-42 yr, BMI: 29.1-43.9 kg/m2, waist/hip ratio [WHR]: 0.86-1.00) we studied the GH, ACTH and F responses to the im administration of glucagon (0.017 mg/kg at 0 min). The results in OB were compared with those in a group of 6 age-matched controls normal subjects (Ns aged 26-32 yr, BMI 19.7-22.5 kg/m2). In Ns glucagon administration induced clear increase in GH (peak vs baseline, mean+/-SE: 11.6+/-3.4 vs 3.3+/-0.7 microg/l, p<0.02), and ACTH (52.9+/-15.2 vs 19.0+/-1.5 pg/ml, p<0.02) levels which peaked at +150 and +165 min, respectively. Increase in F levels (222.3+/-23.8 vs 158.3+/ 7.0 ng/ml, p<0.05) was also recorded but peaked at +180 min. In OB glucagon administration induced GH response (7.4+/-2.3 vs 0.8+/-0.6 microg/l) lower (p<0.05) than that recorded in Ns; when the GH responses were evaluated by co variance analysis, a significant difference between the 2 groups was recorded in term of peaks but not of AUCs. On the other hand, the ACTH response to glucagon in OB was higher than that in Ns (11452.6+/-2447.7 vs 4892.2+/-719.4 pg/ml x min, p<0.05). The F response to glucagon in OB and Ns was, however, similar (24057.9+/ 4109.1 vs 29835.9+/-1566.0 ng/ml x min). In conclusion, this study demonstrates that in obese patients the im administration of glucagon elicits blunted GH response but exaggerated ACTH increase which is uncoupled with the adrenal response. These findings agree with the existence of concomitant GH insufficiency and altered corticotroph function in obesity. PMID- 12109631 TI - Breast pain and mammographic density increase as a consequence of raloxifene therapy. AB - A 55-yr-old post-menopausal woman with osteopenia and no history of breast disease is presented. She was placed on raloxifene HCl 60 mg and soon after developed severe breast pain. The follow-up mammogram, performed prematurely at 6 months, showed a marked increase in breast density. Therapy was accordingly stopped and mastodynia subsided. The patient's mammogram regressed to pre treatment status after 6 months off-therapy. PMID- 12109632 TI - Diagnosis and management of pheochromocytoma during pregnancy. AB - Although the presence of pheochromocytoma in pregnancy is extremely rare, this association deserves much attention as the tumor constitutes a very high risk for both mother and fetus. Any pregnant woman with hypertension, especially if paroxysmal or labile, or with so far unexplained "spells", should induce the clinician to consider the possibility of a pheochromocytoma. Maternal and fetal survival depend a lot on an early diagnosis, a correct medical therapy and a correct timing of delivery and surgery. In this respect, a strict collaboration between obstetricians, endocrinologists, anesthesiologists and surgeons is pivotal. PMID- 12109634 TI - GUSTO V: should it affect clinical practice? Global Use of Strategies to Open Occluded Coronary Arteries. PMID- 12109633 TI - Simvastatin normalizes qtc dispersion and reduces ventricular electrical instability in isolated hypercholesterolemia. AB - This study aimed at evaluating a possible relationship between cholesterol levels and ventricular electrical instability in human beings. Forty subjects (26 males and 14 females, mean age+/-SD 50.3+/-3.7 yr) with isolated hypercholesterolemia (> or =240 mg/dl) were selected from a population of 250 patients who attended the outpatient department of our institution for symptomatic extrasystolic activity (ventricular premature complexes >3,000/24 h). Subjects were randomly assigned to receive either simvastatin 40 mg/d or placebo for 3 consecutive months. After treatment, subjects in the simvastatin group presented a significant decrease of total cholesterol and LDL-cholesterol (p<0.001) and an increase of HDL-cholesterol levels (p<0.01), associated with a reduction of both QTc dispersion (p<0.001) and ventricular premature complexes (p<0.001). None of these changes were observed in the placebo group. At baseline, there was a relationship between cholesterol levels, ventricular premature complexes (VPC) (r=0.33, p<0.05) and QTc dispersion (r=0.41, p<0.01). After treatment, reductions in serum cholesterol levels correlated with decreases of both VPCs (r=0.37, p<0.01) and QTc dispersion (r=0.49, p<0.01). In subjects with isolated hypercholesterolemia simvastatin may reduce the cardiovascular risk associated with ventricular electrical instability. PMID- 12109635 TI - A traveler with nonhealing skin nodules. PMID- 12109636 TI - Malignant melanoma: treatments emerging, but early detection is still key. AB - Although we are beginning to develop treatment options for malignant melanoma, earlier recognition of potential primary melanomas remains the most effective way to increase survival in this highly malignant disease. This article reviews prominent risk factors for melanoma, key physical findings in at-risk patients, new melanoma staging guidelines, and recent and emerging therapy options. PMID- 12109637 TI - Why shouldn't we use warfarin alone to treat acute venous thrombosis? PMID- 12109638 TI - A 76-year-old man with septic arthritis. PMID- 12109639 TI - GUSTO V: combination drug treatment of acute myocardial infarction. Global Use of Strategies to Open Occluded Coronary Arteries. AB - The combination of abciximab in full doses and reteplase in half doses did not significantly reduce the rate of mortality at 30 days in patients with acute ST segment elevation myocardial infarction (MI) when compared with reteplase in full doses in the Global Use of Strategies to Open Occluded Coronary Arteries (GUSTO V) trial. However, subgroup analysis indicates that the combined regimen reduced the complications of acute MI, representing an important alternative strategy for pharmacologic reperfusion. PMID- 12109640 TI - Aromatase inhibitors in breast cancer: current and evolving roles. AB - The aromatase inhibitors are established first-line drugs for treating metastatic breast cancer in postmenopausal women, and are gaining acceptance as adjuvant treatment as well. PMID- 12109641 TI - Coxibs supplement: has CCJM sold out? PMID- 12109642 TI - Acute renal failure in hospitalized patients. AB - Distinguishing among the three categories of acute renal failure is important, as the evaluation and management are tailored to the particular cause. Most cases are due to acute tubular necrosis. To minimize the risk, we should give hospital patients adequate hydration, use potentially nephrotoxic drugs with caution, keep the use of radiographic contrast agents to a minimum, and give patients at risk a nonionic instead of ionic contrast agent when undergoing radiographic procedures. PMID- 12109643 TI - Catastrophic antiphospholipid syndrome in a patient with Behcet's disease. AB - We report on a patient who had a life-threatening relapse of Behcet's disease associated with a catastrophic antiphospholipid syndrome. The patient experienced over a short time a recurrent acute myocardial infarction, multiple venous thromboses, uveitis, and erythema nodosum. Search for thrombophilic factors was positive only for lupus anticoagulant (LAC) and criteria for the diagnosis of the antiphospholipid antibody syndrome were fulfilled. LAC was not found three months after the discharge. At that time the patient had no evidence of clinically active disease or thrombosis. We suggest that LAC was the main triggering factor for the repeated thromboses in this patient. PMID- 12109645 TI - Multicentric reticulohistiocytosis. The long course of a rare disease. AB - Multicentric reticulohistiocytosis is a rare systemic disease. The patients usually develop an erosive polyarthritis in a few years. The most effective treatment is still unclear due to the few reports published of this disease. We report a case of multicentric reticulohistiocytosis with clinical, histopathologic and immunohistochemical study. The patient was treated with cyclophosphamide and nonsteroidal antiinflammatory drugs with an infrequent long term survival. PMID- 12109644 TI - Renal cell carcinoma mimicking polymyalgia rheumatica. Clues for a correct diagnosis. AB - Proximal musculoskeletal symptoms mimicking the clinical picture of polymyalgia rheumatica may herald the onset of solid malignancies. We report on three patients presenting with polymyalgia-like symptoms, who had renal cell carcinoma. The review of the literature and our cases suggest that the absence of prolonged morning stiffness, the atypical clinical findings, the inefficacy of corticosteroids, and the absence of shoulder sonographic pathologic findings may help to facilitate the proper diagnosis. PMID- 12109646 TI - Catastrophic antiphospholipid syndrome precipitated by bowel surgery. PMID- 12109647 TI - Diagnosis and definition of primary Sjogren's syndrome. AB - During the last years, several sets of criteria for the diagnosis and classification of primary Sjogren's syndrome (SS) have been promulgated. So far none has achieved universal acceptance. It is conceivable that the lack of pathognomonic features of SS and the frequent coexistence of SS with other connective tissue di-seases partly explain the lack of agreement. Regardless of criteria preference the final diagnosis of SS should reflect the definition of the syndrome, being an inflammatory and autoimmune rheumatic disease. It is therefore suggested that any criteria employed should include a mandatory combination of measurements of exocrine dysfunction. histological confirmation of inflammation, serological evidence of autoimmunity and exclusion or diseases mimicking primary Sjogren's syndrome. PMID- 12109648 TI - Protein deposition diseases: when structurally altered proteins assemble out of place. With special emphasis on monoclonal immunoglobulin deposition disease causing erosive polyarthropathy. PMID- 12109649 TI - Increased cancer incidence in a Swedish cohort of patients with systemic lupus erythematosus. AB - OBJECTIVE: To assess the risk of cancer in patients with systemic lupus erythematosus (SLE). METHODS: A population-based cohort of 5 715 hospitalised SLE patients was followed 1964-1995 through linkage of the Hospital Discharge Register to the National Swedish Cancer Register. RESULTS: In all, 443 malignancies occurred during the observation-period. The overall risk was increased by 25% (SIR= 1.25, CI 95% 1.14-1.37) and lymphomas constituted the major excess risk. The risk of non-Hodgkin's lymphoma (NHL) was nearly 3-fold increased (SIR = 2.86, CI 95% 1.96-4.04). There was also an increased risk of lung cancer (SIR= 1.73, CI 95% 1.25-2.32) and squamous cell skin cancer (SIR= 1.53, CI 95% 0.98-2.28), which was most pronounced at more than 15 years of follow-up. CONCLUSION: The major finding was a bimodal incidence pattern with an increased risk of lymphoma, mainly NHL, early during follow-up, but lung cancer and squamous skin cancer later on. PMID- 12109650 TI - Gammalinolenic acid treatment of fatigue associated with primary Sjogren's syndrome. AB - OBJECTIVE: To evaluate the efficacy of the essential omega-6 fatty acid Gammalinolenic acid (GLA) on fatigue associated with primary Sjogren's syndrome. METHODS: Ninety patients with primary Sjogren's syndrome (with or without signs of autoimmunity) entered a 6-month double blind placebo-controlled randomised trial with high dose GLA (extracted from Evening Primrose Oil) or corn oil. The primary outcome parameter was fatigue; secondary endpoints were eye dryness, mouth dryness, muscle and joint pain. RESULTS: No statistically significant improvement was found in fatigue assessed by Visual Analogue Scale (VAS) or in the time needed for sleeping/resting during a 24-hour period. No differences were found between the treatment and placebo group. The same applies to the secondary endpoints: no differences in VAS for eye and mouth dryness or pain, no significant changes in Schirmer-1-test, van Bijsterveld score, unstimulated whole sialometry (UWS), or use of artificial tears or analgesics. Only mild side effects were observed. CONCLUSION: According to our study results GLA (Evening Primrose oil) treatment for fatigue in primary Sjogren's syndrome is ineffective. PMID- 12109651 TI - Tc-99m-labeled human polyclonal immunoglobulin G (HIG) scintigraphy in Sjogren's syndrome. AB - OBJECTIVE: To evaluate the usefulness of Tc-99m-HIG scintigraphy in patients with Sjogren's syndrome. METHODS: Twelve consecutive patients with verified secondary Sjogren's syndrome were included in this prospective study. The control group consisted of seven patients with Lupus erythematosus; none of them showed clinical signs of Sjogren's syndrome. Planar and SPECT images of the head were performed six hours after i.v. administration of Tc-99m HIG. RESULTS: Eleven out of twelve patients with secondary Sjogren's syndrome showed a positive result, while one was false negative. Tracer accumulation in patients with positive scintigraphy varied. All patients of the control group were negative. CONCLUSION: Our data in a limited number of patients suggest that Tc-99m HIG scintigraphy could be a modality with high sensitivity and specificity for the diagnosis of Sjogren's syndrome and can provide objective information on the severity of the disease. PMID- 12109652 TI - Carotid and femoral artery stiffness in Takayasu's arteritis. An ultrasound study. AB - PURPOSE: To determine the stiffness of large arteries in patients with Takayasu's arteritis (TA). MATERIALS AND METHODS: A noninvasive ultrasonic method was used to measure Peterson's elastic modulus (Ep), Young's elastic modulus (YM). and the stiffness constant beta of the common carotid arteries in 13 and of the common femoral arteries in 16 patients with TA and in healthy control subjects. RESULTS: The median carotid artery Ep, YM, and stiffness index beta were statistically significantly higher in patients with TA than in controls (p=0.019, 0.013, and 0.004, respectively). The median femoral artery Ep and YM were also higher in patients with TA than in controls (p=0.005 and 0.039, respectively), and the stiffness constant beta (p=0.127) also tended to be higher in patients with TA relative to healthy persons. CONCLUSIONS: Carotid artery compliance is strongly diminished in patients with TA. A comparable but somewhat less conspicuous change is seen in the femoral arteries. PMID- 12109654 TI - The prevalence of H. pylori is still substantial in rheumatic patients. AB - The separate contribution of NSAIDs and H. pylori in the pathogenesis of peptic ulcer disease has not been fully elucidated. The aim of this study was to investigate the seroprevalence of H. pylori in patients with rheumatic diseases and chronic NSAID treatment. Patients with a rheumatic disease, age 40-80 years, and regular use of NSAIDs (at least 3 times a week) were included (n= 1214). IgG antibodies to H. pylori were found in 39% and increased gradually with age: from 25% in patients in the 40-50 years age group to 48% in patients aged 70-80 years (p<0.0001). No difference was observed between men and women, or between the three centres. In our population of rheumatic patients treated with NSAIDs the seroprevalence of H. pylori is substantial (39%), but seems to be lower than in previous reports, which may be due to a cohort effect. PMID- 12109653 TI - Abnormal regional cerebral blood flow in primary antiphospholipid antibody syndrome patients with normal magnetic resonance imaging findings. A preliminary report. AB - Technetium-99m ethyl cysteinate dimer (Tc-99m ECD) brain single photon emission computed tomography (SPECT) was used to detect abnormal regional cerebral blood flow (rCBF) in primary antiphospholipid antibody syndrome (PAPS) patients. The patients had mild neuropsychiatric manifestations and normal brain magnetic resonance imaging (MRI) findings in this preliminary study. Fifteen such female PAPS patients were examined with Tc-99m ECD brain SPECT in order to evaluate the rCBF. In addition, serum anti-cardiolipin antibodies (ACA) and lupus anticoagulant (LA) were simultaneously measured for comparison. RESULTS: (a) Tc 99m ECD brain SPECT revealed hypoperfusion brain lesions in 12 (80%) of the PAPS patients. Pariental lobes were the most commonly involved areas. (b) 11 (73%) and 9 (60%) cases had positive ACA and positive LA, respectively. In addition, ACA and LA results were correlated to the Tc-99m ECD brain SPECT findings. Tc-99m ECD brain SPECT is a sensitive tool for detecting abnormal rCBF in PAPS patients with mild neuropsychiatric manifestations and normal brain MRI findings. PMID- 12109655 TI - Ureaplasma urealyticum as a possible cause of reflex sympathetic dystrophy syndrome. AB - We describe the cases of two patients with clinical and radiological findings of the reflex sympathetic dystrophy syndrome (RSDS) in whom the history of a previous genito-urinary inflammation and high levels of ESR lead us to suspect a hidden reactive arthritis. However, instrumental examinations showed a characteristic picture of RSDS without evident signs of arthritis. In both patients we decided a treatment with quinolones because of detection of an ureaplasma urealyticum genito-urinary infection. This brought to complete remission of the joint symptoms in a few days. Our findings suggest that ureaplasma urealyticum can cause and sustain a RSDS picture, maybe with a reactive arthritis-like mechanism, and that an antibiogram-driven antimicrobial treatment can be rapidly effective against this disorder. PMID- 12109656 TI - Analysis of p53 mutations and Helicobacter pylori infection in human and animal models. AB - BACKGROUND: p53 gene mutations are believed to play a critical role in the development of gastric carcinoma. We examined the relation between Helicobacter pylori infection and p53 gene mutations of the gastric mucosa in human and animal models. METHODS: To detect the original p53 DNA sequences of the Japanese monkey and Mongolian gerbil, the p53 genes of these animals were amplified using the nested polymerase chain reaction method with primers for the human p53 gene. Direct DNA sequencing of exons 5, 6, 7, and 8 of the p53 genes was performed by the dideoxy terminator method for gastric mucosa of humans, the Japanese monkey, and the Mongolian gerbil. The expression of p53 was examined immunohistochemically in a Japanese monkey model. RESULTS: Mutations of the p53 gene were identified in 52.4% of human H. pylori-positive mucosa and in 100% of monkey H. pylori-positive mucosa. However, no mutations of the p53 gene were found in the H. pylori-positive gastric mucosa of Mongolian gerbils. There were no mutations in H. pylori-negative gastritis mucosa of humans, monkeys, or Mongolian gerbils. Nuclear staining of p53 was seen in the glandular cells of the H. pylori-infected mucosa of Japanese monkeys, especially in the neck region of the glands. CONCLUSIONS: These findings demonstrate that the H. pylori infection can induce p53 point mutations in humans and the Japanese monkey and appear to be involved in the pathway leading to dysplasia or carcinoma. However, our direct DNA sequencing method showed no p53 mutations in the Mongolian gerbil model at present. Further studies with this model are needed. PMID- 12109658 TI - Virtual endoscopy: current perspectives. AB - Virtual endoscopy is a new method for evaluating the gastrointestinal tract using either thin-section computed tomography (CT) or magnetic resonance imaging (MRI). The acquired data are then subjected to computer manipulation to render images simulating the conventional endoscopic view. CT and MR imaging data can provide information that is not accessible endoscopically. These important features include information on tissue extending through and beyond organ walls and the anatomic context of the entire gastrointestinal tract, which permits correct anatomic localization of the lesion. Many clinical studies have shown that it is a safe, noninvasive, well-tolerated alternative to conventional endoscopy. Virtual endoscopy may have potential as a method of screening for colorectal cancer. This review describes the technique, reviews reported results, and discusses the present and future applications of this technique, focusing on CT colography (CTC). PMID- 12109657 TI - Helicobacter pylori strain-specific modulation of gastric mucosal cellular turnover: implications for carcinogenesis. AB - Helicobacter pylori colonization induces inflammation in essentially all hosts, a persistent process that increases the risk of developing distal gastric adenocarcinoma. However, only a small percentage of persons carrying H. pylori develop neoplasia; enhanced risk may be related to differences in expression of specific bacterial products, differences in the host response to the bacteria, or the interaction between host and microbe. H. pylori strains that have the cag pathogenicity island are associated with further increased risk for developing distal gastric cancer; however, host responses to H. pylori, such as altered epithelial cell proliferation and apoptosis, also may be important in lowering the threshold for carcinogenesis. H. pylori cag+ strains selectively enhance proliferation and attenuate apoptosis in human mucosa compared to cag- strains. However, cag+ strains also induce more severe gastritis, suggesting that host inflammatory mediators such as cytokines, prostaglandins, and hormones may modulate H. pylori-induced alterations in cellular turnover. In the Mongolian gerbil model of gastric carcinogenesis, apoptosis increases early and transiently following H. pylori infection, but scores progressively decline despite worsening gastric inflammation. Epithelial cell proliferation peaks later and is significantly related to increased gastrin levels, suggesting that epithelial cell growth in H. pylori-colonized mucosa may be mediated by gastrin-dependent mechanisms. An emerging model invoked by these data is one in which H. pylori cag+ strains, in conjunction with host mediators, enhance gastric epithelial cell proliferation but not apoptosis in vivo. The combination of increased proliferation without a concordant increase in apoptosis may therefore contribute to the heightened retention of mutagenized cells, which over decades may increase the subsequent risk for gastric cancer. PMID- 12109659 TI - Very-high-flow injection rate for upper abdominal CT angiography. AB - BACKGROUND: The purpose of this study was to compare a very-high-flow injection rate method (group A) and a conventional injection-rate method (group B) for visualization of upper abdominal arteries by multidetector helical computed tomography (MDHCT). METHODS: The subjects were 240 patients suspected to have abdominal lesions. They were randomly assigned to group A (120 patients) and group B (120 patients). In group A, the bilateral medial cubital veins were punctured, and contrast medium was infused at a rate of 8.6-9.6 ml/s. In group B, the unilateral medial cubital vein was punctured, and contrast medium was infused at a rate of 2.0-3.0 ml/s. The quality of vascular visualization was graded as poor, good, or excellent by three radiologists. RESULTS: All visualizations of the celiac trunk (CE) and superior mesenteric artery (SMA) were graded as excellent in both group A and group B. Visualization grades of the subsegmental branches of the hepatic artery (HA), right gastric artery (RGA), cystic artery, dorsal pancreatic artery (DPA), and superior pancreaticoduodenal artery (SPDA) were good or excellent, in 75% (paging method)/53.3% (three-dimensional method), 85%/30%, 77.7%/18.3%, 76.7%/28.3%, and 88.3%/42.5%, respectively, in group A, and 33.3%/11.7%, 46.7%/3.4%, 41.6%/5%, 55%/4.2%, and 72.5%/14.2%, respectively, in group B. The appearance rate of intrahepatic portal branches was 28.3% in group A and 66.7% in group B in the arterial dominant phase. CONCLUSION: Group A showed better visualization results than Group B in upper abdominal arteries according to MDHCT. PMID- 12109660 TI - Gastric mucosa: long-term outcome after cure of Helicobacter pylori infection. AB - The histopathological changes due to chronic Helicobacter pylori infection are well characterized. Nevertheless, the clinical and pathological outcomes resulting from the cure of this infection remain incompletely described. In particular, the influence of eradication of H. pylori on nonulcer dyspepsia, the long-term effects of H. pylori eradication on gastric atrophy and intestinal metaplasia, and the role of H. pylori eradication in the prevention of gastric cancer are still unclear. We reviewed 38 studies reported between 1993 and 1999 on the outcome of various disorders related to H. pylori infection after successful eradication. There is general agreement concerning the regression of chronic gastritis, lymphoid follicles, and limited-stage low-grade MALT lymphomas of the gastric mucosa after eradication of H. pylori infection. Conversely, there are still major questions on whether H. pylori eradication improves the outcome of premalignant lesions, such as atrophy, intestinal metaplasia, and dysplasia. Finally, some extragastric idiopathic diseases seem to improve in consequence of the eradication of the infection, although there are still no definitive data to support this. PMID- 12109661 TI - Long-term Helicobacter pylori infection and the development of atrophic gastritis and gastric cancer in Japan. AB - The incidence of gastric cancer and the prevalence of Helicobacter pylori are high in Japan, so it is an important issue whether long-term H. pylori infection leads to chronic atrophic gastritis, considered one of the precursors of gastric cancer. We have reported that the grade of atrophy was higher in H. pylori positive subjects than in H. pylori-negative subjects. It has also been reported that the atrophy of gastric mucosa increased in H. pylori-infected monkeys compared with control monkeys in a 5-year follow-up study. Most H. pylori infections occur in children, and atrophy of the gastric mucosa progresses during aging. Long-term data show that H. pylori infection can lead to gastric atrophy and may play an important role in the development of gastric cancer. Interestingly, there was no difference in the prevalence of H. pylori between patients with chronic gastritis and gastric cancer in Japan, but the prevalence of H. pylori in young Japanese gastric cancer patients was significantly higher than in the control group. These data clearly show that H. pylori infection is one of the risk factors of gastric cancer in young Japanese people, There is no answer to whether curing H. pylori infection can reverse the atrophy of the gastric mucosa and decrease the risk of gastric cancer development. To clarify this issue, an intervention study must be done. A large clinical trial called the Japanese Intervention Trial of H. pylori is in progress. PMID- 12109662 TI - Pathological issues of gastric and lower esophageal cancer: helicobacter pylori infection and its eradication. AB - Gastric carcinoma is thought to develop via the actions of inducers and promoters of carcinogenesis. Tryptophan in charred fish or animal meat, ultraviolet rays, and irradiation, which damage genes of normal cells, have long been regarded as inducers of carcinoma, and agents such as alcohol, tobacco, aflatoxin, and nitrosoamine as promoters, with tobacco having both activities. The interaction between these environmental factors, principally diet, and Helicobacter pylori (Hp) is important in the genesis of gastric carcinoma. In this report, the histopathological feature of the Hp gastritis-carcinoma sequence is outlined, and the pathological characteristics of gastroesophageal reflux disease (GERD) and endoscopically negative reflux disease (ENRD) and the risk factors for lower esophageal carcinoma after Hp eradicated status in particular are discussed regarding aspects of cell cycle-associated factors. We conclude that (1) Infection with Hp increases the risk of gastric cancer in two histological phenotypes (i.e., diffuse undifferentiated type and intestinal differentiated type). Excessive cell replication and interrupting the mucus secretion mechanism may result in a large proportion of cells with genetic abnormalities. (2) Genetic alterations in gastric carcinogenesis may differ from those in colonic carcinogenesis. (3) The degree of GERD in Japanese patients is milder than that in patients from Western countries, although the incidence of GERD increases the status after successful eradication of Hp. It is also possible that accumulation of genetic abnormalities increases the number of cardiac and lower esophageal cancers. Investigation of cell cycle factors in GERD including ENRD can be expected to reveal the risk of carcinogenesis. PMID- 12109663 TI - Indications for Helicobacter pylori eradication therapy and first-line therapy regimen in Japan: recommendation by the Japanese Society for Helicobacter Research. AB - In November 2000, eradication therapy for Helicobacter pylori infection was approved under the present Japanese system of health insurance. Before the approval, the Japanese guideline of "Diagnosis and Treatment of H. pylori infection" was made by the guideline committee of the Japanese Society for Helicobacter Research. Indications for H. pylori eradication therapy were classified into three groups: (A) recommended: gastric and duodenal ulcers; (B) recommended and managed at a specialized institution: low-grade gastric mucosa associated lymphoid tissue lymphoma; (C) current studies of the significance of the therapy: following endoscopic mucosal resection of early gastric cancer and gastric surgery for gastric cancers, hyperplastic gastric polyp, atrophic gastritis, and nonulcer dyspepsia. The first-line therapy regimen recommended is 1 week of triple therapy with a proton pump inhibitor standard dose twice a day, amoxicillin 750 mg bid, and clarithromycin 200 or 400 mg bid. The reasons for preferring this regimen are to avoid extensive use of metronidazole, which is allowed for treatment of Trichomonas infection in Japan, and the low rate of emergence of clarithromycin-resistant H. pylori with amoxicillin co-therapy. For second-line therapy patients should be referred to specialists, who can examine the susceptibility of H. pylori isolates against antibiotics and have much experience with this therapy. PMID- 12109664 TI - Gastric cancer: pathogenesis, risks, and prevention. AB - Intestinal (IGCA) and diffuse (DGCA) gastric adenocarcinomas, the two main microscopic subtypes, are dissimilar regarding their epidemiological and demographic characteristics. Both tumor types comprise approximately 40% of all gastric adenocarcinomas. The DGCAs more often occur in young age groups, more often affect the corpus, and are less infrequently associated with atrophic gastritis and intestinal metaplasia than the IGCAs. The risk of both DGCA and IGCA is increased in the presence of Helicobacter pylori infection, and the risk rises with increases in grade and extent of atrophic gastritis and intestinal metaplasia. It is likely that the development of up to 80% of the DGCAs and IGCAs can be prevented with early eradication of the H. pylori infection. The pathogenesis and morphogenesis of DGCAs are unknown, but the morphogenesis of IGCAs includes identifiable precancerous conditions such as atrophic gastritis and intestinal metaplasia as well as identifiable precancerous lesions (adenomas, dysplasias). Atrophic gastritis is a direct result of the H. pylori infestation. Atrophic gastritis, for unknown reasons, appears in more than half of the infected subjects during their lifetime. H. pylori gastritis triggers a variety of reactions, with the reaction cascades resulting in errors of the cell genome and ending up as neoplastic tumors. PMID- 12109665 TI - Helicobacter pylori infection and molecular changes in gastric carcinogenesis. AB - In recent years there has been much progress in understanding the pathogenesis of gastric cancer. The role of individual factors in gastric carcinogenesis continues to be debated and is also subject to further analysis. In addition to the activation of oncogenes and the inactivation of tumor suppressor genes, alteration of adhesion molecules seems to be critical for the development of gastric cancer. Helicobacter pylori has been linked to an increased risk of developing gastric cancer, and the molecular changes induced by H. pylori are currently being investigated. Recent studies indicate that H. pylori induces cell proliferation and apoptosis during the early phase of chronic inflammation of the gastric mucosa, whereas in the malignant transformation of the gastric mucosa apoptosis is inhibited and adhesion of gastric epithelial cells is impaired. This review focuses on the role of H. pylori in the development of molecular changes in gastric cancer and its preneoplastic lesions. PMID- 12109666 TI - Hepatitis C virus and its roles in cell proliferation. AB - Hepatitis C virus (HCV) causes chronic hepatitis and is linked to the development of hepatocellular carcinoma (HCC). The role of HCV infection in the development of HCC remains to be clarified. We analyzed the effect of HCV core protein on modulation of cell proliferation. HCV core protein was shown to have at least two functions: activation of the Ras/Raf signaling pathway and anti-apototic function. PMID- 12109667 TI - Hepatocarcinogenesis in hepatitis viral infection: lessons from transgenic mouse studies. AB - Over the past decade, genetically engineered mouse models have been used for studies of the mechanisms underlying human diseases. One advantage of these models is that the targeted protein executes its function in normal cells in their natural tissue microenvironments. Transgenic mouse models for human viral hepatitis have also been established and have provided new insights into the pathogenesis of hepatitis and hepatocellular carcinoma (HCC). In the search for the mechanism of hepatocarcinogenesis in hepatitis viral infection, two viral proteins, the core protein of hepatitis C virus (HCV) and the HBx protein of hepatitis B virus (HBV), have been shown to possess oncogenic potential through transgenic mouse studies, indicating the direct involvement of the hepatitis viruses in hepatocarcinogenesis. The presence of the hepatitis C virus core or HBx protein, which has an oncogenic potential, may allow some of the steps in multistep hepatocarcinogenesis to be skipped. This may explain the very high frequency of HCC in patients with HCV or HBV infection. PMID- 12109668 TI - Development of an H. pylori-infected animal model and gastric cancer: recent progress and issues. AB - Gastric cancer is one of the most common malignancies in the world. Gastric carcinogenesis is multifactorial. Although some environmental factors, such as excessive intake of salt and N-nitroso compounds in foods, are involved in this process, Helicobacter pylori has been closely linked to an increased risk of the development of gastric adenocarcinoma. The IARC/WHO in 1994 concluded that H. pylori is a definite carcinogen in humans based on seroepidemiological evidence. H. pylori infection is associated with both intestinal and diffuse types of gastric cancer. Strong evidence has accumulated that H. pylori infection has been active in the development of gastric adenocarcinoma in the Mongolian gerbil (MG) model after long term infection and with or without treatment using low-dose chemical carcinogens. There are several criticisms of these reports. The most troublesome issue is the diagnostic criteria for gastric cancer in MGs, especially for well-differentiated cancers induced by H. pylori infection. Hence, common criteria for the diagnosis of gastric cancer in MGs is required. Another scientific approach is to find genetic-evidence to confirm the diagnosis. Such evidence could indicate directly that H. pylori infection itself is a promoter, initiator, or both in the development of gastric adenocarcinoma. PMID- 12109669 TI - Hepatitis C virus superinfection in patients with chronic hepatitis B virus infection. AB - Because hepatitis B virus (HBV) and hepatitis C virus (HCV) have the same transmission routes, dual infection may occur and even persist in the same patient. The reported series on seroprevalence of HCV indicate that HCV is found in more than 10% of HBV-infected patients worldwide. HCV superinfection in patients with chronic HBV infection tends to cause severe and progressive liver disease that is resistant to interferon therapy. Paradoxically, HCV exerts a suppressive effect on HBV and may enhance seroclearance of HBV antigens, or even usurp the role of HBV as the agent for continuing hepatitis. In view of the complex dynamism of viral interaction, the importance of HCV assay and the necessity of monitoring patients with chronic HBV infection in clinical studies cannot be overemphasized. The basic mechanisms that regulate the viral interactions largely remain to be investigated. PMID- 12109670 TI - Management of chronic hepatitis C and prevention of hepatocellular carcinoma. AB - Hepatitis C virus (HCV) infection is responsible for more than a half of the cases of chronic viral hepatitis in Japan. About 20% of patients who are chronically infected with the virus develop cirrhosis about 20-30 years after the infection, with hepatocellular carcinoma developing in about 5% of patients a year. The only drug that effectively reduces the virus is interferon, but complete eradication of the virus can be obtained in only 30%-40% of treated patients. Reevaluation of the predictive factors to eradicate the virus by 24 week interferon therapy showed that the genotype other than 1b, a low virus load, and multiple amino acid substitutions in the interferon sensitivity determining region (ISDR) of genotype 1b are statistically significant predictive factors. Amino acid substitution in the PePHD domain of the E2 protein was rare and was unrelated with the outcome of interferon therapy. The fluctuation of the virus titer measured by branched DNA during a 2-year observation period was less that 10-fold in most patients, and amino acid substitutions in the ISDR were rare in such patients, suggesting that one point measurement of these parameters may be useful to select candidates for interferon therapy. A comparison of patients treated with interferon and untreated patients from the viewpoint of cancer prevention showed only a slight decrease in the risk of treated patients developing hepatocellular carcinoma. However, patients who showed normal alanine amino-transaminase (ALT) irrespective of virus clearance showed a significantly reduced risk of liver carcinogenesis. Similarly, a retrospective study to evaluate the long-term preventive effect of glycyrrhizin on hepatocellular carcinoma development showed that the therapy was effective in lowering the ALT value and in preventing liver carcinogenesis. PMID- 12109671 TI - Indications and criteria for liver transplantation for fulminant hepatic failure. AB - Liver transplantation is the only effective treatment for potentially fatal cases of fulminant hepatic failure. However, it is very difficult to predict which cases will be fatal. The mortality may depend on alternative medical therapies. According to a nationwide survey of patients with fulminant hepatic failure presenting with encephalopathy of a coma grade greater than II within 8 weeks from the first symptoms of illness with a prothrombin time less than 40% of normal value, there were 93 patients in 311 hospitals between January and December 1998 in Japan. During this period, there were 11 patients with late onset hepatic failure. The etiology was HAV infection in 4%, HBV infection in 44%, and nonA-nonB in 41%. Specific therapies were intensively used in all patients. The mean survival rate was 41%, with differences depending on the etiology. Six patients underwent liver transplantation, and 5 survived. In animal experiments, sinusoidal fibrin deposition caused massive liver necrosis. Activation of Kupffer cells and hepatic macrophages was a major contributing factor of this development. There were different mechanisms of such fibrin deposition. Tumor necrosis factor-alpha and superoxide anions released from hepatic macrophages after endotoxin administration destroyed endothelial cells, and then coagulopathy occurred in the sinusoids in rats given Propionibacteriom acnes, while a tissue factor from Kupffer cells played that role in rats undergoing partial hepatectomy. The prognosis of fulminant hepatic failure may depend on the etiology. The indication for liver transplantation for this disease must be carefully decided by analyzing the etiology, pathological conditions, and response to therapies in each case. PMID- 12109672 TI - Liver transplantation for fulminant hepatic failure. AB - The etiology and prognosis of individuals with various forms of fulminant hepatic failure are reviewed. Special techniques of clinical management and decision making as to when and to whom to transplant in cases of fulminant hepatic failure are reviewed. PMID- 12109673 TI - Liver transplantation for fulminant hepatitis at Stanford University. AB - BACKGROUND: To review the clinical characteristics and outcomes of 26 patients evaluated for liver transplantation for fulminant hepatic failure at Stanford University and Lucile Packard Children's Hospital in an attempt to identify risk factors and prognostic predictors of survival. METHODS: A retrospective review of the records of 26 consecutive patients who were evaluated for possible liver transplantation for acute liver failure from May 1, 1995, to January 1, 2000. Pretransplant patient demographics and clinical characteristics were collected, and the data were analyzed by univariate and multivariate analysis. RESULTS: Clinical assessment of encephalopathy did not predict outcome. Patients with abnormal computed tomography (CT) of the brain had a twofold increase in mortality compared with those patients with normal studies (p = 0.03). Patients requiring mechanical ventilation and continuous venovenous hemofiltration (CVVH) also had a poor prognosis. CONCLUSION: Predictors of poor outcome after fulminant hepatic failure include abnormal CT scan, mechanical ventilation, and requirement for hemofiltration. PMID- 12109674 TI - Auxiliary partial orthotopic liver transplantation (APOLT) in the treatment of acute liver failure. AB - BACKGROUND: Auxiliary partial orthotopic liver transplantation (APOLT) has been developed in order to benefit from the efficacy of orthotopic liver transplantation (OLT) in the treatment of fulminant hepatic failure (FHF), but to avoid the negative counterpart of OLT which is to eliminate the possibility of native liver (NL) regeneration and which consequently implies a life-long immunosuppression. METHODS: In our institution we performed 16 consecutive APOLTs in 15 patients between October 1992 and December 1999. Patients' mean age was 30 years (range 0.5-65 years). The causes of FHF were viral (HAV = 3; HBV = 3), drugs (n = 4), or others (n = 5). None of the patients had a history of chronic liver disease. The decision to transplant was taken when the patients met well defined criteria. All but one of the patients were in a coma. RESULTS: Five patients died, 10 patients are alive (66.7%). Regeneration of the NL occurred in 11 of the 15 patients (73.3%) and in 8 of the 10 survivors. Six of these 8 patients have permanently stopped immunosuppressive therapy. These results can be favorably compared with those of OLT for FHF. In the European Transplant Registry, the survival rate is 57% at 5 years (2612 patients receiving OLT for FHF between 1988 and 1998). In our experience the survival rate is 59% at 5 years (42 patients receiving OLT for FHF between 1987 and 1999). CONCLUSIONS: APOLT is feasible in both adults and children; it rapidly restored liver function and reversed encephalopathy. Right APOLT seems more advisable since the right liver provides more functional hepatocytes; however, left APOLT harvested in an adult appears sufficient for a child. APOLT should be proposed only to patients with high chances of liver regeneration: age of recipient, etiology of liver failure, interval between onset of jaundice and occurrence of encephalopathy, and quality of liver graft are early prognostic indicators. Better results have been observed with younger patients (less than 40 years old) presenting with FHF (rather than subfulminant hepatic failure (SHF)) and due to HAV, HBV, or paracetamol. PMID- 12109675 TI - Colorectal cancer screening: the potential role of virtual colonoscopy. AB - Virtual colonoscopy is a promising new technique that combines rapid spiral CT scanning of the abdomen with advanced computer programs capable of re-creating two- and three-dimensional views of the colon and rectum. Recent studies comparing this method with conventional colonoscopy show that virtual colonoscopy already is more accurate than barium enema X-ray studies for the detection of colorectal polyps, and that it approaches the accuracy of colonoscopy for diagnosing advanced lesions. Before virtual colonoscopy can be promoted for population-based screening for colorectal cancer, a number of issues discussed in this review need to be addressed. These include questions of accuracy, availability, acceptability, and cost-effectiveness. PMID- 12109676 TI - Biliopancreatic ascariasis: endoscopic approach. AB - From June 1985 to June 1999, 120 patients were diagnosed by means of panendoscopy, choledocoscopy, and endoscopic retrograde cholangiopancreatography as having biliary and pancreatic ascariasis. All duodenoscopic procedures were performed under fluoroscopic control. Endoscopic extraction was successful in 99 of 108 (92%) patients, some of whom had had previous endoscopic sphincterotomy. The endoscopic accessories used were a basket catheter, polypectomy snare, and balloon catheter. Because no significant complications were observed after this endoscopic treatment, we recommend endoscopic extractions for biliary ascariasis. PMID- 12109677 TI - Diverse cellular and apoptotic responses to variant shapes of UHMWPE particles in a murine model of inflammation. AB - The wear of orthopaedic prostheses results in the release of a markedly heterogeneous assortment of particulate debris, with respect to both size and shape. Although particle size has been extensively examined, the role of particle shape in adverse inflammatory reactions to debris remains unclear. Using an in vivo murine model of inflammation, we assessed tissue responses to globular and to elongated ultra-high molecular weight polyethylene (UHMWPE) particles with a similar surface area, and investigated whether inflammation and cellular apoptosis varied with particle shape in the debris-tissue interaction. Histological changes of UHMWPE-stimulated pouch membrane were assessed using a computerized image analysis system. Quantitative real time PCR and ELISA were performed to assess mRNA expression and protein level of the cytokines, and TUNEL assays were conducted to quantify apoptotic cells. The data revealed that elongated particles generated more active inflammatory air pouches, stimulated more severe membrane proliferation and the inflammatory cellular infiltration compared to globular particles. Increased levels of IL-1beta and TNFalpha were detected in the lavage and homogenate of pouches stimulated with elongated particles in comparison to pouches with globular particles, and the apoptotic assay indicated more severe apoptotic changes within the inflammatory membrane provoked with elongated particles. Our results suggest that cellular responses to UHMWPE wear debris are dependent on the shape of the particles. PMID- 12109678 TI - Soft tissue response to microtextured silicone and poly-L-lactic acid implants: fibronectin pre-coating vs. radio-frequency glow discharge treatment. AB - From in vitro studies it is known that a plasma-treatment can enhance cell spreading. Similar effects can be observed after pretreatment of the surface with a protein coating, to mediate cell adhesion. The aim of the current study was to evaluate the in vivo effects of these surface modifications, in a three-month experiment in a goat model. We made silicone and poly-L-lactic acid implants with double-sided parallel micro-grooves (depth 1.0 microm, width 10.0 microm), a random surface roughness, or a smooth surface. Implants either received a radio frequency glow discharge (RFGD) treatment, a fibronectin (Fn) pre-coating, or no pre-treatment. Subsequently, they were inserted into subcutaneous pockets created on the flanks of goats for 1, 3 or 12 weeks. Histological analysis showed that a fibrous tissue capsule had formed around all implants. Histomorphometrical analysis was performed on capsule thickness, capsule quality and the implant tissue interface quality. Fn-treated surfaces showed a considerable early inflammatory reaction. Besides this, RFGD treatment or Fn pre-coating did not further influence any of the measured parameters. In conclusion, pre-treatment of polymer implant surfaces with Fn or RFGD treatment did not significantly influence tissue reaction around implants with micro-grooved, roughened or smooth surfaces. PMID- 12109679 TI - PEG modulated release of etanidazole from implantable PLGA/PDLA discs. AB - In this work, etanidazole (one type of hypoxic radiosensitizer) is encapsulated into spray dried poly(D),L-lactide-co-glycolide) (PLGA) microspheres and then compressed into discs for controlled release applications. Etanidazole is characterized by intracellular glutathione depletion and glutathione transferases inhibition, thereby enhancing sensitivity to radiation. It is also cytotoxic to tumor cells and can chemosensitize some alkylating agents by activating their tumor cell killing capabilities. We observed the release characteristics of etanidazole in the dosage forms of microspheres and discs, subjected to different preparation conditions. The release characteristics, morphology changes, particle size, and encapsulation efficiency of microspheres are also investigated. The release rate of etanidazole from implantable discs (13 mm in diameter, 1 mm in thickness, fabricated by a press) is much lower than microspheres due to the reduced specific surface. After the initial burst of 1% release for the first day, the cumulative release within the first week is less than 2% until a secondary burst of release (caused by polymer degradation) occurs after one month. Some key preparation conditions such as drug loadings, disc thickness and diameter, and compression pressure can affect the initial burst of etanidazole from the discs. However, none of them can significantly make the release more uniform. In contrast, the incorporation of polyethylene glycol (PEG) can greatly enhance the release rate of discs and also reduces the secondary burst effect, thereby achieving a sustained release for about 2 months. PMID- 12109680 TI - pH-stabilization of predegraded PDLLA by an admixture of water-soluble sodiumhydrogenphosphate--results of an in vitro- and in vivo-study. AB - Aim of the study was to examine if the addition of buffering sodiumhydrogenphosphate to poly(D,L)lactide(PDLLA) would stabilize the pH-value in the in vivo environment of implanted material and whether this improves its biocompatibility. The material was predegraded just to the point of viscous disintegration to test the PDLLA in the moment of its most aggressive effect on the surrounding tissue. Racemic amorphous PDLLA was injection-molded with or without the admixture of 1 mol NaP per 100 mol lactate, the degradation product of PDLLA (=1 mol%) to form 20mm x 3 mm x 2mm rods. Predegradation was performed by storing the rods at 55 degrees C for 14 days, just to the point of beginning dissolution. Predegraded PDLLA or PDLLA + NaP samples were used for in vitro incubation tests, as well as for the in vivo study, where the rods were implanted into the spinal muscles of 30 male Wistar rats. Repeatedly, measurements of the pH-value were made in the incubation solutions in vitro. The surrounding tissue of the implanted samples as well as the normal contralateral muscle tissue was checked for its pH-value in a group of 3 rats, respectively, anaesthesized at various time intervals after implantation. After these measurements the implants and their surrounding tissues were excised for histological examination. In Ringer's solution pH-values dropped immediately within the first week of incubation of both predegraded materials reaching -4 pH units after 4 weeks in the PDLLA containing medium, after 6 weeks in the PDLLA + NaP containing medium. Soerensen buffer slowed the pH decrease with significant differences between the material groups up to the 28th week. In vivo, the pH of the surrounding tissue was influenced by the implanted PDLLA material up to the 4th week, while the admixture of NaP resulted in a significant pH stabilization. A higher quantity of macrophages and giant cells were seen between the 2nd and 6th week after the implantation in the environment of pure PDLLA compared with PDLLA + NaP. Complete resorption of predegraded pure PDLLA or PDLLA + NaP from the extracellular space was reached 28 weeks postimplantation in vivo. Thus, sodiumhydrogenphosphate improves the biocompatibility of degrading PDLLA at the point of viscous disintegration by stabilizing the pH-value in the environment of the implants for several weeks and reducing adverse tissue reactions. PMID- 12109681 TI - Viscoelastic memory and self-expansion of self-reinforced bioabsorbable stents. AB - The possibility to decide the speed and rate of expansion of stents is of great clinical importance by reason of the varying requirements for different indications to use stents. Self-reinforced bioabsorbable stents can be made self expanding owing to the viscoelastic memory of the material. Stents are stable at room temperature and expansion occurs at body temperature. The level at which the expansion stops depends on the material, crystallinity, initial diameter of spira and annealing temperature. The expansion rate can be estimated by logarithmic equation, if material, draw ratio and diameter of stent wire are constant. This is, however, possible only if processing parameters are constant. Based on the present results annealing temperature and expansion time were seen to be directly proportional to the expansion rate of the stent. PMID- 12109682 TI - Identification of apolipoprotein A-I as a major adsorbate on biomaterial surfaces after blood or plasma contact. AB - Apolipoprotein A-I is the major protein component of HDL. It is reported in this communication that apo A-I is a significant component of the protein layer adsorbed from blood or plasma to a variety of biomaterial surfaces, including hydrophobic and hvdrophilic polymers, liposomes, a heparinized surface, and a polysulfone hemodialyzer membrane. Evidence in support of this conclusion from SDS-PAGE and immunoblots of proteins eluted from the surfaces after blood or plasma contact is presented. Whether the presence of apo A-I on these surfaces results from the uptake of the free protein or HDL particles remains to be determined. It appears that apolipoprotein A-I and/or HDL deposition may be an important effect in blood-biomaterial interactions generally, and one that has been largely overlooked. PMID- 12109683 TI - A dynamic mechanical method for determining the silicone elastomer solubility of drugs and pharmaceutical excipients in silicone intravaginal drug delivery rings. AB - The silicone elastomer solubilities of a range of drugs and pharmaceutical excipients employed in the development of silicone intravaginal drug delivery rings (polyethylene glycols, norethisterone acetate, estradiol, triclosan, oleyl alcohol, oxybutynin) have been determined using dynamic mechanical analysis. The method involves measuring the concentration-dependent decrease in the storage modulus associated with the melting of the incorporated drug/excipient, and extrapolation to zero change in storage modulus. The study also demonstrates the effect of drug/excipient concentrations on the mechanical stiffness of the silicone devices it 37 degrees C. PMID- 12109684 TI - Differential response to chemically altered polyethylene by activated mature human monocyte-derived macrophages. AB - Macrophages and polyethylene (PE) particulate are currently recognized as being the two common denominators in the development of chronic inflammation, periprosthetic osteolysis, and subsequent implant failure. In this study, the effect of PE particulate surface chemistry on mature human monocyte-derived macrophage (MDM) function was investigated. Virgin high-density PE (HDPE: 4-10 microm) and HDPE oxidized by irradiation, thermal and chemical treatment were characterized by FT-IR and suspended in soluble type I collagen, which was subsequently solidified on glass coverslips. Human MDMs, derived from differentiating monocytes on polystyrene for 14 days, were trypsinized and cultured on collagen-particle substrata and collagen controls for 31 days. Analysis of conditioned media collected at 24h incubation showed a significantly higher level of IL-1beta secretion in virgin HDPE over oxidized HDPE or collagen controls, and a significant inhibition of IL-6 secretion in both virgin and oxidized samples. Esterase activity was increased in the medium at a significantly higher level in the virgin HDPE versus controls with the highest activity observed in oxidized HDPE at 31 days. These results illustrate the effect of PE particle surface chemistry (oxidation) on MDM cytokine secretion and esterase activity, and highlight the need to further investigate the potential of PE surface chemistry on modulating MDM function. PMID- 12109685 TI - A microshear test to measure bond strengths of dentin-polymer interfaces. AB - The microbond test. a single fiber shear test, has been adapted to be a microshear test for the measurement of the adhesion of resin-based dental materials to dentin and enamel. The objective of this study is to improve the design of this microshear test so that it can provide accurate and reliable shear bond strength data. In the current design of the microshear test apparatus, the bonding diameters of the specimens have been as small as 0.70 mm. The smaller diameters give researchers the ability to test several bonded specimens on one flat dentin or enamel surface, thus allowing both for the regional mapping of the mineralized surface and the conservation of extracted teeth needed to provide the necessary substrates. The test corfiguration used in earlier studies has been modified through finite element analysis to address concerns in the test methodology. The results of this study show that the microshear bond test can be a useful tool in helping to understand the complex interactions that occur at the interface between dental composites and dentin and/or enamel surfaces, especially at interfacial sites not amenable to macroshear testing. PMID- 12109686 TI - FTIR/ATR study of protein adsorption and brushite transformation to hydroxyapatite. AB - Previous study has demonstrated that brushite (CaHPO4 x 2H2O), modified by partial potassium substitution for calcium, can transform quickly into hydroxyapatite (HA, Ca5(PO4)3OH) when exposed to aqueous salt solutions at room temperature. Analyses techniques used in those studies required sample retrieval from solution, which may alter the sample surface. In this work FTIR/ ATR was used in analysis, enabling in situ study of the transformation within the aqueous environment. To test the biocompatibility of this brushite, cellular response to the transformation needs to be understood. Cellular response was initiated by bovine serum albumin adsorption on the brushite surface. The response was studied by monitoring the conformation of the adsorbed protein, which is critical to cellular reaction. This required monitoring the brushite transformation and surface adsorbed protein conformation simultaneously which can be realized using FTIR/ATR. Based on band fitting and second derivative results from the spectra it was found that the conformation of the adsorbed BSA changes during the brushite transformation to HA. This study also demonstrated that the deposition of the brushite could be monitored in real time which offers the possibility for studying surface bonding during electrodeposition. PMID- 12109687 TI - Synthesis and characterization of photocrosslinkable, degradable poly(vinyl alcohol)-based tissue engineering scaffolds. AB - Hydrogels have many advantages that make them prime candidates for tissue engineering applications: high water content, tissue-like elasticity, and relative biocompatibility. We aim to tissue engineer heart valves using a hydrogel scaffold based on poly(vinyl alcohol) (PVA), and the design parameters for a suitable tissue engineering scaffold are quite stringent. In this research, we develop degradable and photocrosslinkable poly(lactic acid)-g-PVA multifunctional macromers that can be reacted in solution to form degradable networks. The mass loss profiles and bulk properties of the resulting scaffolds are easily tailored by modifying the structure of the starting macromers. Specifically, altering the number of lactide repeat units per crosslinking side chain, percent substitution, molecular weight of PVA backbone, and macromer solution concentration, the rate of mass loss from these degradable networks is controlled. In addition, by increasing the network's hydrophobicity, valve interstitial cell adhesion is improved. PMID- 12109688 TI - Immobilization of lipase using hydrophilic polymers in the form of hydrogel beads. AB - The purpose of this study was to immobilize lipase (triacylglycerol ester hydrolase, E.C. 3.1.1.3) from Candida rugosa using various polymers in the form of beads, to evaluate enzyme loading, leaching, and activity; and to characterize the beads. Agarose, alginate, and chitosan were the polymers selected to immobilize lipase by entrapment. Agarose beads exhibited undesirable swelling in the leaching and activity medium and the polymer was not used further. Alginate or chitosan beads were prepared by ionic gelation using calcium chloride or sodium tripolyphosphate, respectively, as the cross-linking agent in the gelling solution. Some hatches of beads of each polymer were freeze dried. The results show that alginate beads leached substantially more enzyme than did chitosan beads. Entrapment efficiency, however, was the same for different chitosan levels as well as different alginate levels (43-50%). Activity in alginate was low at 240 +/- 33 and 220 +/- 26, compared to 1,110 +/- 51 and 1,150 +/- 11 units/ml in chitosan, for fresh and freeze-dried beads, respectively. The higher lipase activity in chitosan beads compared to that in alginate beads could be attributed to an alginate-enzyme interaction. It can be concluded that chitosan is a polymer worthy of pursuit to immobilize lipase. PMID- 12109689 TI - Molecular weight and degree of deacetylation effects on lipase-loaded chitosan bead characteristics. AB - The effects of the molecular weight (MW) and degree of deacetylation (DD) of chitosan on chitosan hydrogel beads were characterized, and the entrapment efficiency, release of entrapped lipase, and activity of immobilized Candida rugosa lipase were investigated. Fresh and freeze-dried beads were characterized. A solution of lipase was prepared in a 1.5% (w/v) chitosan and 1% (v/v) acetic acid medium, and then dropped into a tripolyphosphate solution to prepare the beads. The release studies were performed over 36 h. The enzyme activity was assayed using the Sigma lipase activity method. Chitosan with high MW and DD resulted in a higher loading. A lower activity was observed for beads produced with high DD chitosan. MW did not have a marked effect on the activity. The release study revealed that enzyme release increased to a maximum when the bead was manufactured with a low MW and a moderate to high DD chitosan sample. Freeze drying did not affect the release or the activity of the lipase. Chitosan with a high MW and DD can thus improve loading and reduce the release of lipase in these beads. The choice of chitosan can affect the activity normalized for lipase loading, and beads with desirable qualities can be produced. PMID- 12109690 TI - Cytotoxicity and MMP-9 activation induced in human mononuclear cells by UHMWPE oxidation. AB - Ultra high molecular weight polyethylene (UHMWPE) used in orthopedic prosthesis can be oxidized during sterilization processes such as gamma-ray irradiation. Oxidation alters UHMWPE structure and mechanical properties and it has been suggested that this alteration is the first step in the loosening process. Direct effects of UHMWPE oxidation on the cellular and tissue response to the implant have not been previously investigated. We used heat-oxidized UHMWPE, whose oxidative state is comparable to the one induced in gamma-ray irradiated polymer. in order to observe the peripheral blood mononuclear cells (PBMCs) behavior after 24 and 48 h exposure to the oxidized and non-oxidized polymers. Flow cytometric analysis showed a cytotoxic effect of oxidized UHMWPE after 48 h compared with the control samples. Moreover, gelatin zymography of PBMCs conditioned media showed a strong increase in MMP-9 (gelatinase B) release and activation in oxidized UHMWPE samples. This first evidence of a direct effect of the UHMWPE oxidative status on the cellular behavior suggests that oxidation alters not only the UHMWPE physical-mechanical properties. but it can also be responsible for altered tissue response. PMID- 12109691 TI - In vitro effects on MG63 osteoblast-like cells following contact with two roughness-differing fluorohydroxyapatite-coated titanium alloys. AB - The chances of integration between an implant and the surrounding bone tissue depend on the surface characteristics of the implant itself. Particularly, chemical composition and surface roughness of the material have emerged as crucial factors in affecting the behaviour of cells in contact with the material. Among various surfaces, calcium phosphate coatings seem to favour a rapid initial integration, but their dissolution by extracellular fluids raises some concern about the long-term stability at the bone-implant interface. Fluorinated apatites are known to be more stable than other ceramic coatings, but, at present, little is known on their effects on human cells. In this study, MG63 osteoblast-like cells were seeded onto two fluorohydroxyapatite (FHA)-coated titanium alloy (Ti6Al4V) materials differing in roughness, respectively, LR-FHA (Ra = 5.6 microm) and HR-FHA (Ra = 21.2 microm). Quantification of the cells in contact with the FHA-coated materials by conventional methods involved some technical difficulties, on which we report. Only the indirect esteem by the measure of total content of proteins and a procedure based on cell count, following a double enzymatic treatment to detach the cells, offered plain results, indicating no significant differences between cellular growth in contact with test materials and with plastic control. Differentiation and functionality of the cells were comparatively evaluated by analysis of alkaline phosphatase activity and osteocalcin production. As far as osteocalcin release is concerned, only slight variations were detected on FHA-coated materials in comparison with the control. Both types of coatings showed a significant increase in alkaline phosphatase activity with respect to the control, the roughest surface exhibiting a more prolonged effect on the time. PMID- 12109693 TI - A new method to produce macropores in calcium phosphate cements. AB - A new way to create macropores in calcium phosphate cements has been developed. The method consists in adding NaHCO3 to the starting cement powder (Biocement D) and using two different liquids: first a basic liquid to form the paste and later an acid liquid to obtain CO2 bubbles. Mercury intrusion measurements showed a dramatic increase both in macropores with an average size of 100 m and in the total porosity (even higher than 50% with respect to the Biocement D). This method does not change in any significant way the final reaction products of the starting material after being soaked 3 days in Ringer solution. Only, due to the increase of the porosity. the compressive strength of the porous cement decreases significantly. PMID- 12109692 TI - Glycosaminoglycan hydrogel films as bio-interactive dressings for wound healing. AB - Chemically-crosslinked glycosaminoglycan (GAG) hydrogel films were prepared and evaluated as bio-interactive wound dressings. Hyaluronan (HA) and chondroitin sulfate (CS) were first converted to the adipic dihydrazide derivatives and then crosslinked with poly(ethylene glycol) propiondialdehyde to give a polymer network. The crosslinking occurred at neutral pH in minutes at room temperature to give clear, soft hydrogels. After gelation, a solvent-casting method was used to obtain a GAG hydrogel film. A mouse model was used to evaluate the efficacy of these GAG films in facilitating wound healing. Full-thickness wounds were created on the dorsal side of Balb/c mice and were dressed with a GAG film plus Tegaderm' or TegadermT' alone. A significant increase in re-epithelialization was observed on day 5 (p < 0.001) and day 7 (p < 0.05) for wounds treated with a GAG film plus Tegaderm versus those treated with Tegaderm alone. While no significant differences in wound contraction or inflammatory response were found, wounds treated with either HA or CS films showed more fibro-vascular tissue by day 10. The GAG hydrogel films provide a highly hydrated, peri-cellular environment in which assembly of other matrix components. presentation of growth and differentiation factors, and cell migration can readily occur. PMID- 12109695 TI - Optical grating coupler biosensors. AB - By incorporating a grating in a planar optical waveguide one creates a device with which the spectrum of guided lightmodes can he measured. When the surface of the waveguide is exposed to different solutions, the peaks in the spectrum shift due to molecular interactions with the surface. Optical waveguide lightmode spectroscopy (OWLS) is a highly sensitive technique that is capable of real-time monitoring of these interactions. Since this integrated optical method is based on the measurement of the polarizability density (i.e., refractive index) in the vicinity of the waveguide surface, radioactive, fluorescent or other kinds of labeling are not required. In addition, measurement of at least two guided modes enables the absolute mass of adsorbed molecules to be determined. In this article, the technique will be described in some detail, and applications from different areas will be discussed. Selected examples will be presented to demonstrate how monitoring the modification of different metal oxides with polymers and the response of the coated oxides to biofluids help in the design of novel biomaterials; how OWLS is useful for accurate bioaffinity sensing, which is a key issue in the development of new drugs; and how the quantitative study of protein-DNA/RNA and cell surface interactions can enhance the understanding of processes in molecular and cellular biology. PMID- 12109694 TI - Thermomechanical behavior of virgin and highly crosslinked ultra-high molecular weight polyethylene used in total joint replacements. AB - Three series of uniaxial tension and compression tests were conducted on two conventional and two highly crosslinked ultra-high molecular weight polyethylenes (UHMWPEs) all prepared from the same lot of medical grade GUR 1050. The conventional materials were unirradiated (control) and gamma irradiated in nitrogen with a dose of 30 kGy. The highly crosslinked UHMWPEs were gamma irradiated at room temperature with 100 kGy and then thermally processed by either annealing below the melt transition at 100 degrees C or by remelting above the melt transition at 150 degrees C. The true stress-strain behavior of the four UHMWPE materials was characterized as a function of strain rate (between 0.02 and 0.10 s(-1)) and test temperature (20-60 degrees C). Although annealing and remelting of UHMWPE are primarily considered as methods of improving oxidation resistance, thermal processing was found to significantly impact the crystallinity, and hence the mechanical behavior, of the highly crosslinked UHMWPE. The crystallinity and radiation dose were key predictors of the uniaxial yielding, plastic flow, and failure properties of conventional and highly crosslinked UHMWPEs. The thermomechanical behavior of UHMWPE was accurately predicted using an Arrhenius model, and the associated activation energies for thermal softening were related to the crystallinity of the polymers. The conventional and highly crosslinked UHMWPEs exhibited low strain rate dependence in power law relationships, comparable to metals. In light of the unifying trends observed in the true stress-strain curves of the four materials investigated in this study, both crosslinking (governed by the gamma radiation dose) and crystallinity (governed by the thermal processing) were found to be useful predictors of the mechanical behavior of UHMWPE for a wide range of test temperatures and rates. The data collected in this study will be used to develop constitutive models based on the physics of polymer systems for predicting the thermomechanical behavior of conventional and crosslinked UHMWPE used in total joint replacements. PMID- 12109696 TI - Development of hydrogel implants for urinary incontinence treatment. AB - Hydrogel implants for urinary incontinence treatment based on HEMA supplemented with 10% methacrylic acid have been developed. The swelling properties of implants were tested in vitro and in vivo after implantation to laboratory mice. Biocompatibility has been determined by incubation of implants in tissue culture, by histological examination of adjacent tissues after subcutaneous application of implants to laboratory mouse in a long-term experiment, and by flow cytometry examination of blood cells. The swelling of hydrogel implants was completed in 6 24 h. There was no effect on in vitro growth of cells incubated with implants. In mice, implants were well tolerated without any sign of inflammatory reaction. The material allows an elastic compression of urethra compensating a damaged sphincter after trans-urethral sub-mucosal implantation of hydrogel cylinders. PMID- 12109697 TI - Enhanced fibrin remodeling in vitro with TGF-beta1, insulin and plasmin for improved tissue-equivalents. AB - The aim of this study was to better understand how to increase collagen content in and enhance mechanical properties of tissue-equivalents formed by entrapping tissue cells in fibrin gels, with the ultimate goal of developing mechanically robust artificial tissues. The two main areas of focus were cell culture medium composition and replacement frequency relative to a base case of incubating constructs in medium supplemented with just serum and replaced weekly. The optimal condition involved a combination of all three factors investigated-TGF beta, insulin, plasmin-with medium replacement three times a week. Compared to a base case without these three factors, the combination case resulted in 20 times more collagen and a ten-fold increase in tensile strength. The high strain modulus and tensile strength were within an order of magnitude of cardiovascular tissues. This study indicates great potential for fibrin-based tissue-equivalents in soft connective tissue applications. PMID- 12109698 TI - Effect of metal alloy surface stresses on the viability of ROS-17/2.8 osteoblastic cells. AB - In this study we compared the effect of structural stresses and surface roughness on biocompatibility of NiTi- and Ti-alloy for ROS-17/2.8 osteoblastic cells. We suggest here that cell viability and cell attachment are linear functions of internal (structural) stress and subgrain size of the implant alloy. However, this is not the case with surface roughness. The two-phase state in these materials is characterized by different mean values of structural stresses (sigma) in alpha-martensite and beta-phase. We found a straight correlation between cell viability and sigma(beta)/sigma(alpha) ratio. Atomic force microscopy revealed that, even after equal surface polishing treatments, roughness varied significantly between the different alloys. The effect of the surface structure of the alloy on the osteoblastic ROS-17/2.8 cell survival rate was studied with combined calcein-ethidium-homodimer fluorescence labeling. The possible effects on cell attachment to substrate were studied by staining the focal contacts with paxillin antibody. All the NiTi surfaces were tolerated well and the cells attached most abundantly to the roughest NiTi surface but the smoothest Ti-alloy surface. However, other parameters of the material state, such as the surface stresses created by hot rolling seem to be responsible for some of the attachment and cell survival features observed in this study. PMID- 12109699 TI - Oral keratinocytes cultured on dermal matrices form a mucosa-like tissue. AB - Oral reconstructions for cleft palate repair are often complicated by a shortage of mucosal tissue. This shortage causes scar tissue formation leading to impaired growth of the dento-maxillary complex. The overall aim of our research is to develop a substitute. which limits the iatrogenic effects of cleft palate surgery. This study describes the culture and characterization of mucosal substitutes containing keratinocytes. Epidermal and oral keratinocytes from a beagle dog were cultured on several skin-derived and collagen-based substrates. Oral keratinocytes cultured on the skin-derived substrates closely resembled normal oral epithelium of the dog. A multi-layered epithelium was formed showing parakeratosis, expression of cytokeratin 16 and the formation of a basement membrane. Epidermal keratinocytes cultured on the skin-derived substrates formed an epithelium which was similar to dog epidermis. In contrast, keratinocytes cultured on the collagen-based substrates invaded the substrate without the formation of a multi-layered epithelium. In conclusion, this study shows that oral canine keratinocytes cultured on skin-derived substrates exhibit a tissue organization that resembles normal oral mucosa. This type of mucosal substitute will therefore be used in further studies for implantation on the palate of beagle dogs. These studies might eventually lead to an improvement of cleft palate surgery in humans. PMID- 12109700 TI - In vitro behavior of silicate glass coatings on Ti6A14V. AB - The in vitro response in simulated body fluid (SBF) of silicate glass coatings on Ti6A14V was evaluated. Glasses belonging to the SiO2-CaO-MgO-Na2O-K2O-P2O5 system were used to prepare 50-70 m thick coatings on Ti6Al4V, employing a simple enameling technique. Glasses with silica content higher than 55 wt% can be used to prepare coatings that do not crack or delaminate and exhibit good adhesion to the alloy. It has been found that coatings with silica content lower than 60 wt% are more susceptible to corrosion and precipitate carbonated hydroxyapatite on their surface during in vitro tests. However, these coatings have a higher thermal expansion than the metal and are under tension. After 2 months in SBF cracks grow in the coating that reach the glass/metal interface and initiate delamination. Glasses with silica content higher than 60 wt% are more resistant to corrosion and have lower thermal expansion. These coatings do not crack but they do not precipitate apatite even after 2 months in SBF. PMID- 12109701 TI - Effects of Ti ions and particles on neutrophil function and morphology. AB - We compared the cytotoxicity of soluble and particulate titanium (Ti), vanadium (V) and nickel (Ni) by biochemical functional analysis and by microscopic morphology with micro-area elemental analysis in vitro using human neutrophils as probes and in vivo in animals. The biochemical analyses of LDH, superoxide anion, TNF-alpha and scanning electron microscopy (SEM) showed that Ni in solution destroys the cell membrane of neutrophils, whereas Ti and V in solution stimulate neutrophils and increase the quantity of released superoxide anions. Fine Ti particles (1-3 microm), which smaller than neutrophils (about 5 microm), were phagocytized by the cells and the results were similar in vivo. These results showed that the cytotoxic effect of Ti particles is size dependent, and that they must be smaller than that of cells. The present study demonstrated that the biochemical functional tests are useful for evaluating the biocompatibility of materials. PMID- 12109702 TI - Polyphosphoester microspheres for sustained release of biologically active nerve growth factor. AB - Controlled delivery of neurotrophic proteins to a target tissue by biodegradable polymer microspheres has been explored widely for its potential applications in the treatment of various disorders in the nervous system. We investigated in this study the potential of polyphosphoester microspheres as carriers for the sustained release of nerve growth factor (NGF), a water-soluble neurotrophic protein. Two polyphosphoesters (PPEs), P(BHET-EOP/TC) and P(DAPG-EOP), as well as poly(lactide/glycolic acid) (PLGA), were used to fabricate microspheres by a W/O/W emulsion and solvent evaporation/extraction method. With bovine serum albumin as a model protein to optimize processing parameters. P(DAPG-EOP) microspheres exhibited a lower burst effect but similar protein entrapment levels and efficiencies when compared with those made of PLGA. Bioactive NGF could be released for at least 10 weeks from the P(DAPG-EOP) microspheres, as confirmed by a neurite outgrowth assay of the PC12 cells. These NGF containing microspheres were incorporated into the nerve guide conduits that were implanted to bridge a 10 mm gap in a rat sciatic nerve model. Two weeks after implantation, immunostaining with an antibody against the neurofilament protein confirmed the presence of axons at the distal end of regenerated cables within the NGF microsphere-loaded conduits. These results demonstrated the feasibility of using biodegradable PPEs for microencapsulation of NGF and provided a basis for future therapeutic application of the microspheres. PMID- 12109703 TI - Mechanical integrity of compression-moulded ultra-high molecular weight polyethylene: effects of varying process conditions. AB - Ultra-high molecular weight polyethylene (UHMWPE) bearing surfaces in knee and hip prostheses are frequently manufactured by direct compression moulding of the as-polymerised powder. A study was made of the important role of the temperature time sequence in the melt state during processing, in determining the mechanical integrity of mouldings at 37 degrees C. Structural features were determined by calorimetry (for the degree of crystallinity), infra-red spectroscopy (for the degree of oxidation), density measurement, and scanning electron microscopy. Mechanical integrity was assessed by tensile tests at a constant nominal strain rate of 10(-3) s(-1), with post-failure microscopic examination. For the whole range of melt temperatures 145-200 degrees C and times 10-90 min, essentially the same stress-strain path was followed, reflecting invariance of the degree of crystallinity. However, there were dramatic changes in elongation-to-break, from ca 10% for some mouldings at 145 degrees C to a mean of 560% at 175 degrees C where, at the 86% confidence level, there was evidence for a peak. The rise was explained by microscopy, that revealed two distinct types of fusion defect, of reducing severity with increasing temperature. Type 1 defects were voids arising from incomplete powder compaction, and persisted up to 165 degrees C. Type 2 defects were regions of enhanced deformability at inter-particle boundaries in apparently fully compacted mouldings, evidenced microscopically by localised relative displacements at particle interfaces, during the plastic deformation at 37 degrees C. They persisted up to 200 degrees C. Type 2 defects may be attributed to the slow self-diffusion of UHMWPE in the melt, leading to incomplete homogenisation. even after compaction is complete. The level of oxidation in the mouldings was small but rose with melt temperature, explaining the fall in elongation-to-break at temperatures higher than 175 degrees C. PMID- 12109704 TI - Influence of a novel object in the home range of the cockroach, Blattella germanica. AB - We investigated whether exploratory behaviour of the cockroach, Blattella germanica (L.) (Dictyoptera: Blattellidae), was modified by the introduction of a novel object, such as a bait station, into their familiar home range. In particular, we measured the attractiveness of boxes, as novel objects, in relation to their complexity and to their contents. The presence of a novel object induced an increase of exploratory behaviour on and inside the object and its environs. Time spent around the object depended on the complexity of the object (closed box or with holes). The more complex the object, the longer cockroaches spent exploring it. When the object contained food, its attractiveness was greatly increased and it affected cockroaches from further away. PMID- 12109705 TI - Landing responses of Anopheles gambiae elicited by oxocarboxylic acids. AB - A wind tunnel bioassay and video system were used to observe Anopheles gambiae Giles sensu stricto (Diptera: Culicidae) landing on glass cylinders, heated to human skin temperature (34 degrees C) and treated with aqueous solutions of oxocarboxylic acids. Six of nine compounds tested: 2-oxobutanoic, 2-oxo-3 methylbutanoic, 2-oxopentanoic, 2-oxo-3-methylpentanoic, 2-oxo-4-methylpentanoic and 2-oxohexanoic elicited significant landing responses in comparison to a water control. Landing responses appeared to be restricted to C4-C6, 2-oxocarboxylic acids. A solution of 1 microg/microL of 2-oxopentanoic acid elicited the highest level of response that was temperature dependent: significant numbers of landings occurred only within +/-2 degrees C of human skin temperature. Chemical analysis by linked gas-liquid chromatography/mass spectrometry of methyl-oxime, trimethylsilyl derivatized samples of human sweat extracts revealed the presence of 2-oxopropanoic (pyruvic) acid and three behaviourally active, branched chain acids: 2-oxo-3-methylbutanoic, 2-oxo-3-methylpentanoic and 2-oxo-4 methylpentanoic. PMID- 12109706 TI - Validity of species status of the parasitic mite Otodectes cynotis. AB - A combined molecular and phenotypic approach was used to determine whether ear mites of the genus Otodectes (Acari: Psoroptidae) belong to a single species. The second internal transcribed spacer (ITS 2) of the rDNA of 16 isolates from 11 cats, two dogs, one arctic fox and two ferrets originating from four different continents was characterized. In addition, mites from dog, cat and arctic fox were investigated morphologically. Sequence comparisons revealed five different, but closely related genotypes which did not segregate according to host species or geographical origin. Morphologically, mites of the three host species did not differ significantly in their body or leg sizes. These investigations support the view that ear mites of the genus Otodectes from different hosts and geographical origins belong to a single species, Otodectes cynotis (Hering). PMID- 12109708 TI - Effect of temperature on the transmission of orbiviruses by the biting midge, Culicoides sonorensis. AB - The influence of temperature on the likelihood of Culicoides sonorensis Wirth & Jones (Diptera: Ceratopogonidae) transmitting African horse sickness virus (AHSV) serotypes 4 and 6, bluetongue virus (BTV) serotypes 10 and 16 and epizootic haemorrhagic disease of deer virus (EHDV) serotype 1 was investigated. Extrinsic incubation periods (EIP), vector competence and vector survival were determined at 15, 20, 25 and 30 degrees C. The effect of humidity on vector survival was also investigated by maintaining adult C. sonorensis at 40, 75 and 85% r.h. at each temperature. Higher temperatures were associated with a shorter EIP for all virus serotypes except AHSV6, to which C. sonorensis was orally refractory, increased vector competence for AHSV4 and EHDV1, but not for BTV10 or BTV16, and a reduction in vector survival. Humidity interacted with temperature in influencing vector survival, such that at low temperatures, lower humidity (40 and 75% r.h.) was detrimental for survival (up to 18% reduction in longevity), whereas at high temperatures, high humidity (85% r.h.) was detrimental (up to 36% reduction in longevity). In general, the transmission potential of C. sonorensis for AHSV4, EHDV1, BTV10 and BTV16 was greater at higher temperatures, because although vector survival was reduced, this was more than compensated for by the accompanying decrease in duration of the EIP. PMID- 12109707 TI - Phylogenetic analysis of the mitochondrial cytochrome oxidase subunit I gene of five species of the Culicoides imicola species complex. AB - The phylogenetic status of members of the Culicoides imicola Kieffer (Diptera: Ceratopogonidae) species complex of haematophagous midges is unknown, and simple means to identify the members using all life stages are unavailable. In this study, the status of three confirmed (C. imicola s.s., C. bolitinos Meiswinkel and C. loxodontis Meiswinkel) and two provisional (C. tuttifrutti Meiswinkel and C. kwagga Meiswinkel) members of the complex from South Africa was assessed using phylogenetic analysis of partial DNA and amino acid sequences of the mitochondrial cytochrome oxidase subunit I (COI) gene. The four or five individuals of each species analysed contained one or two haplotypes each. Interspecific divergence was significant and characterized by strong A <--> T transversion bias. Phylogenetic trees constructed using neighbour-joining, parsimony and maximum likelihood showed each species to be distinct. Combinations of sites for two restriction enzymes in the COI sequences were species-specific and could form the basis of a diagnostic PCR assay. PMID- 12109709 TI - Toxicity and behavioural effects of diet-borne alkaloids on larvae of the black blowfly, Phormia regina. AB - Larvae of the black blowfly, Phormia regina (Meigen) (Diptera: Calliphoridae) were exposed for 24 h to artificial diets that contained one of the following alkaloids: arecoline, caffeine, nicotine, quinine, sparteine or strychnine at either 1000 or 100 p.p.m. Each of the alkaloids caused reduced weight gain, relative to a control population in a no-choice bioassay and, with the exception of quinine, all alkaloids caused reduced larval weights in a choice bioassay. Larvae were unable to move away from diets containing arecoline (1000 and 100 p.p.m) and congregated away from diets containing 1000 p.p.m. quinine. Arecoline (1000 p.p.m) and both concentrations of nicotine caused significant mortality of larvae. Over a longer period (120 h), 10 and 1 p.p.m. nicotine resulted in significant numbers of larvae congregating away from a treated diet. Ten p.p.m. nicotine caused reduced weight gain over 120 h, although larvae provided with a choice were less affected. Exposure of larvae to dried residues of nicotine for 2 h did not affect subsequent development. PMID- 12109711 TI - Seasonal variations in the distribution and abundance of the tsetse fly, Glossina morsitans morsitans in eastern Zambia. AB - The seasonal changes in the distribution of Glossina morsitans morsitans Westwood (Diptera: Glossinidae) and its main host, cattle, were examined in a cultivated area of the plateau of eastern Zambia. During four consecutive years, the tsetse and cattle populations were monitored along a fly-round transect traversing the two main vegetation types in the study area. These were miombo, a one-storied open woodland with the genera Brachystegia and Julbernardia dominant, and munga, a one- or two-storied woodland where the principal tree genera were Acacia, Combretum and Terminalia. Concurrently, a capture/mark/release/recapture (CMRR) exercise was conducted along two other transects also traversing both vegetation types. The index of apparent abundance of tsetse (IAA) in miombo increased at the beginning of the rainy season (November), reached its peak at the end of the rainy season (April) and was low during the cold season (May to late August), but especially the hot dry season (September to late October). The IAA of tsetse in munga showed a pattern that was the reverse of that in miombo. The seasonal changes in the IAA of tsetse in both vegetation types were in accordance with changes in the movement patterns of tsetse between the two vegetation type as observed using CMRR. The distribution and abundance of cattle along the transect also showed a seasonal trend. This was especially so in munga, during the first three years of observations, where cattle abundance increased gradually from June onwards, reached a maximum at the end of the hot dry season (October-November) and declined steeply at the start of the rainy season (November-December). In both vegetation types, the monthly mean IAA of tsetse was positively correlated with the abundance of cattle in the previous month. It is concluded that the distribution of tsetse in cultivated area of the eastern plateau of Zambia undergoes substantial seasonal changes, which can partly be attributed to changes in the distribution of cattle. The implications of these observations for the control of tsetse are discussed. PMID- 12109710 TI - Background odour induces adaptation and sensitization of olfactory receptors in the antennae of houseflies. AB - The presence of background odour was found to have a small but significant effect on the sensitivity of the antennal olfactory system of houseflies, Musca domestica Linnaeus (Diptera: Muscidae), to new pulses of odour. We show that cross-adaptation and cross-sensitization between a background odour of (+/-)-1 octen-3-ol and pulses of (+/-)-1-octen-3-ol, 2-pentanone and R-(+)-limonene can occur, confirming that olfactory receptor cells are sensitive to different odours. Background odour can increase the responses to low concentration odour pulses and decrease the responses to higher concentration odour pulses. It is suggested that background odour has a larger effect on olfactory receptor cells that respond with a tonic increase of spike frequency, giving information about the level of odour concentration, i.e. the 'static' environment. Cells that respond in a phasic way only provide information on the dynamics of the olfactory environment. PMID- 12109712 TI - A comparison of the feeding behaviour of tsetse and stable flies. AB - In Zimbabwe, observations were made of the behaviour of individual stable flies (Stomoxys spp.) (Diptera: Muscidae) and tsetse (Glossina spp.) (Diptera: Glossinidae) feeding on cattle during the wet (Stomoxys and tsetse) and dry (tsetse only) seasons. For Stomoxys landing on adult cattle, only 27% took a full meal (mean feeding time = 147 s). Most Stomoxys left the host before completing their meal, largely due to disturbance by the host's defensive behaviour (24%, mean time = 59 s) or other flies (44%, 71 s). The probability of a Stomoxys leaving the host progressively increased with time. Simultaneous observations of tsetse showed that, compared to Stomoxys, their feeding success was lower (15%), feeding was interrupted earlier (33 s) and the time taken to complete a meal was shorter (109 s). Further studies of tsetse across different seasons and hosts showed that feeding success varied according to host age (adult = 7%; calf = 3%) and was negatively correlated with the frequency of host defensive behaviour and the relative abundance of non-biting Diptera. Disturbances were more often caused by host behaviour (69%) than other flies (31%) and the probability of tsetse leaving decreased with time on the host. Overall, these results suggest that tsetse and Stomoxys have different feeding strategies. In particular, tsetse appear to be more responsive to host defensive behaviour, which reduces their feeding success relative to Stomoxys. These behavioural differences are consistent with the respective life-history characteristics of Stomoxys and tsetse. PMID- 12109713 TI - IgG responses to salivary gland extract of Ixodes ricinus ticks vary inversely with resistance in naturally exposed sheep. AB - Enzyme-linked immunosorbent assay (ELISA) was used to investigate the antibody responses of control sheep, and sheep naturally exposed to Ixodes ricinus Linne (Acari: Ixodidae) ticks, to salivary gland extract (SGE) proteins of partially fed, adult I. ricinus. Comparisons between responses of control sheep and naturally infested sheep by Western blot analysis suggested that variations in IgG responses of I. ricinus-exposed sheep were mostly associated with specific responses to I. ricinus SGE antigens. Sheep IgG responses were positively related to the numbers of adult ticks feeding per sheep at the time samples were collected, were greater during the spring than the autumn periods of I. ricinus activity and were inversely related to sheep resistance to ticks measured by the weights of nymphal I. ricinus that engorged on the sheep. These findings suggest that sheep lose their resistance to ticks due to polarization of a Th1 type response to some tick antigens towards a Th2 type response when sheep are exposed to high, natural tick infestations, or to seasonal conditions of relative nutritional stress. Potential consequences for the epidemiology of tick-borne diseases are discussed. PMID- 12109714 TI - Vector potential of houseflies for the bacterium Aeromonas caviae. AB - Houseflies, Musca domestica Linnaeus (Diptera: Muscidae), have been implicated as vectors or transporters of numerous gastrointestinal pathogens encountered during feeding and ovipositing on faeces. The putative enteropathogen Aeromonas caviae (Proteobacteria: Aeromonadaceae) may be present in faeces of humans and livestock. Recently A. caviae was detected in houseflies by PCR and isolated by culture methods. In this study, we assessed the vector potential of houseflies for A. caviae relative to multiplication and persistence of the bacterium in the fly and to contamination of other flies and food materials. In experimentally fed houseflies, the number of bacteria increased up to 2 days post-ingestion (d PI) and then decreased significantly 3 d PI. A large number of bacteria was detected in the vomitus and faeces of infected flies at 2-3 d PI. The bacteria persisted in flies for up to 8 d PI, but numbers were low. Experimentally infected flies transmitted A. caviae to chicken meat, and transmissibility was directly correlated with exposure time. Flies contaminated the meat for up to 7 d PI; however, a significant decrease in contamination was observed 2-3 d PI. In the fly-to-fly transmission experiments, the transmission of A. caviae was observed and was apparently mediated by flies sharing food. These results support houseflies as potential vectors for A. caviae because the bacterium multiplied, persisted in flies for up to 8 d PI, and could be transmitted to human food items. PMID- 12109715 TI - Comparison of three pyrethroid treatments of top-sheets for malaria control in emergencies: entomological and user acceptance studies in an Afghan refugee camp in Pakistan. AB - Insecticide-treated bedding materials (sheets and blankets) could be protective against vectors of malaria and leishmaniasis--especially in complex emergencies, epidemics and natural disasters where people are more likely to sleep in exposed situations. Comparison of cotton top-sheets impregnated with different pyrethroids (permethrin 500 mg/m2, deltamethrin 25 mg/m2 or alphacypermethrin 25 mg/m2) for effectiveness against mosquitoes (Diptera: Culicidae) was undertaken in a refugee camp in Pakistan. Predominant species encountered were Anopheles stephensi Liston, An. pulcherrimus Theobald, An. nigerrimus Giles, Culex quinquefasciatus Say, Cx. tritaeniorhynchus Giles and other culicine mosquitoes. All three pyrethroid treatments performed significantly better than the untreated sheets in deterrence and killing of mosquitoes. No significant differences were found between the three insecticides tested in terms of entomological effect. Washing of the treated sheets greatly reduced their effectiveness. In a user acceptance study conducted among 88 families (divided into four groups), six families complained of irritation of the skin and mucous membranes. Of these reports, one was from the placebo group (using untreated sheets) and the other five (5/22=23%) from families using deltamethrin-treated sheets. All families allocated to permethrin and alphacypermethrin groups declared an appreciation for the intervention and reported no side-effects. Ten of the placebo group disliked the intervention, citing no prevention of mosquito biting as the reason. Side effects associated with deltamethrin indicate that alphacypermethrin and permethrin are more appropriate first choice insecticides for treatment of sheets and blankets. PMID- 12109716 TI - Host selection by Anopheles arabiensis and An. quadriannulatus feeding on cattle in Zimbabwe. AB - In the Zambezi valley, mosquito females of the Anopheles gambiae Giles complex (Diptera: Culicidae) were collected from a hut containing pairs of cattle distinguishable by known DNA markers. DNA was extracted from the blood-fed mosquito abdomens and primer sets for ungulate and mosquito DNA loci were used to identify the mosquito sibling species and individual host source(s) of their bloodmeals. The 67 mosquitoes comprised a mixture of An. arabiensis Patton (31%) and An. quadriannulatus Theobald (69%). DNA from one or both of the cattle present in the hut was detected in 91% of samples. When the hut contained an adult and a calf, the percentage of bloodmeals from the adult, the calf and adult + calf were 58%, 27% and 15%, respectively; the trend towards meals from the adult host was consistent but not always significant. When the pair of cattle comprised two adults of roughly equal size and age, then mosquitoes generally showed no significant bias towards feeding from one individual. There was no significant difference in the pattern of host selection made by An. arabiensis and An. quadriannulatus but the former had a significantly higher percentage (20%) of mixed meals than An. quadriannulatus (9%). These two members of the An. gambiae complex appear to be less selective in their choice of cattle hosts compared to day-active Diptera such as tsetse and Stomoxys, possibly because the hosts are generally asleep when Anopheles are active and there is therefore less selective pressure to adapt to host defensive behaviour. The slight bias of Anopheles towards older and/or larger cattle may be related to the host's larger surface area. PMID- 12109717 TI - PCR assay for identification of Anopheles quadriannulatus species B from Ethiopia and other sibling species of the Anopheles gambiae complex. AB - Sibling species A and B of Anopheles quadriannulatus (Theobald) are recognized as allopatric members of the Anopheles gambiae Giles complex of Afrotropical mosquitoes (Diptera: Culicidae). Species A represents An. quadriannulatus sensu stricto, widespread in southern Africa, whereas An. quadriannulatus species B occurs in Ethiopia. Because of difficulty of identification, distribution of An. quadriannulatus sensu lato remains poorly known. Cytotaxonomy and the standard DNA polymerase chain reaction (PCR) assay do not distinguish between species A and B of An. quadriannulatus. By optimizing the standard PCR assay (Scott et al., 1993) for identification of members of the An. gambiae complex, we identified two discriminant fragments of 153 bp and 900 bp from DNA of An. quadriannulatus species B, whereas only the 153 bp fragment was amplified for species A from South Africa. This modified PCR assay can therefore be used to distinguish between species A and B of An. quadriannulatus s.l. as well as other members of the An. gambiae complex. PMID- 12109718 TI - Evidence for p-glycoprotein modification of insecticide toxicity in mosquitoes of the Culex pipiens complex. AB - Pesticide resistance has parallels with multi-drug resistance syndrome of tumours in clinical medicine, which has been linked to an ATP-dependent pump, p glycoprotein (P-gp). P-gps pump drugs out of the cell, thereby reducing cellular concentrations of the chemical. P-gps have been found in several invertebrate species and have been shown to provide a defence against environmental xenobiotics, including pesticides. This study used a model cell culture system to investigate the interaction of pesticides with P-gp. Ivermectin and endosulfan were shown to be strong inhibitors of dye transport out of cells, which is a standard measure of P-gp modulation. We then investigated the action of a P-gp inhibitor, verapamil (calcium channel blocker), on insecticide toxicity to fourth instar mosquito larvae of the Culex pipiens L. complex (Diptera: Culicidae). Verapamil increased toxicity to examples of three insecticide classes (cypermethrin, endosulfan, ivermectin), but not to chlorpyrifos (organophosphate). The discovery of a novel protective mechanism in mosquitoes, with a wide substrate range, has implications for the control of important pest and vector species. PMID- 12109719 TI - Qualitative tokenism. PMID- 12109720 TI - Health-seeking behaviors and social change: the experience of the Hong Kong Chinese elderly. AB - In this article, the author outlines how the Hong Kong Chinese elderly revise, shift, and modify their health-seeking behaviors to adapt to rapid social change, presenting data drawn from three studies undertaken with elderly Chinese conducted in Hong Kong during the period 1993 to 1998. The primary data source is a qualitative survey involving interviews with 47 elderly Hong Kong Chinese men and women to ascertain their health-seeking beliefs and behaviors. The author analyzes how public policy and Confucian rhetoric constitute social guidelines, which are perceived in light of an individual's resources, gender and generational experiences and are manifest in health-seeking behaviors: seeking religious solace, preparing special food, visiting formal and informal healers, and shifting expectations of what constitutes family support. The article highlights the interface between public interpretations of old age, morality, religion, filial support, and personalized meaning as manifest in health behaviors. PMID- 12109721 TI - Immigrant women: making connections to community resources for support in family caregiving. AB - The purpose of this ethnographic study was to understand how immigrant women caregivers accessed support from community resources and identify the barriers to this support. The study included 29 Chinese and South Asian women caring for an ill or disabled child or adult relative. All experienced barriers to accessing community services. Some possessed personal resources and strategies to overcome them; others remained isolated and unconnected. Family and friends facilitated connections, and a connection with one community service was often linked to several resources. Caregivers who failed to establish essential ties could not initiate access to resources, and community services lacked outreach mechanisms to identify them. These findings contribute new understanding of how immigrant women caregivers connect with community resources and confirm the impact of immigration on social networks and access to support. PMID- 12109722 TI - The personal development of mothers of terminal cancer patients: how Japanese women change through the experience of caring for and losing their children to cancer. AB - The author interviewed 57 mothers who had lost children to cancer about their experiences concerning their children's illness and death. These mothers became their children's main caretakers because they felt responsible and unable to count on others. They maintained emotional stability while interacting with their children and worked to protect their children from mistakes made by health care professionals. These experiences made many mothers tougher. After their children died, they had to form a different kind of relationship to their children to overcome their grief. This process compelled the mothers to reconsider issues concerning life and death and changed their fundamental values. PMID- 12109723 TI - "That feeling of not feeling": numbing the pain for substance-dependent African American women. AB - Using ethnographic methodology, the author uncovered the meanings and expressions of recovery care for substance-dependent African American women residing in an inner-city transitional home for substance abuse. A convenience sample of 12 key and 18 general participants revealed emotional pain associated with negative life experiences, including overt and covert racism, primarily within society but also within their family networks; and physical, sexual, and emotional abuse from parents, siblings, and male relationships. The women described feelings of abandonment associated with the death of loved ones, particularly mothers. They had attempted to numb their emotional pain with alcohol and drugs. As they moved through treatment and recovery, they began to work through past and current painful life experiences without using alcohol and drugs. PMID- 12109724 TI - Diabetes: the layperson's theories of causality. AB - The authors examine laypersons' perspectives of illness: the content of causal explanations of diabetes and differences in explanations according to gender. Qualitative research was carried out in Guadalajara, Mexico. A nonprobabilistic sample of 20 diabetic individuals participated in interviews, and the content of the interviews was analyzed. On the origin of their condition, participants offered explanations that match neither the biomedical model nor any other formal causal theory. Participants attributed the onset of diabetes to socioemotional circumstances linked to their life experiences and practices. Men attributed causality to work and social circumstances outside the home; women attributed it to family life and domestic circumstances. The authors discuss how lay theories can be useful for the reorganization of health services. PMID- 12109725 TI - Ethical decision making in international nursing research. AB - Researchers in international settings must continually examine cross-cultural ethical issues to ensure that their work is ethically sound. In this article, the authors discuss the process of ethical decision making in international nursing research. They draw on research in Ghana during 1999 involving HIV seropositive women to document this process. Referring to this experience, they argue that international research involving human beings must meet two related but distinct ethical tests: It must meet international ethical standards for the protection of human participants but, at the same time, acknowledge the ethical standards related to the institutional setting and cultural environment in which the research takes place. PMID- 12109726 TI - Research in black and white. AB - The authors consider the methodological, interpretative, and practical issues that arise when there is a difference in ethnicity between researcher and informant in qualitative research by drawing on the academic literature and their fieldwork experiences as White researchers undertaking studies with individuals of African/Caribbean and South Asian descent. Some contemporary issues raised by "researching the other" in the context of pragmatic health services research are highlighted, including access to same-ethnicity researchers, the involvement of interpreters, and the potential for ethnocentric interpretation. The authors believe that qualitative research should be judged by the plausibility of the findings and by a critical evaluation of the way in which the research was conducted and the reflexivity of the researcher. PMID- 12109727 TI - Focus groups in rural Gujarat, India: a modified approach. AB - Focus groups have become increasingly popular in health research. However, their feasibility depends on the context of such research. Through discussion of focus groups they conducted in rural India, the authors argue that successful focus groups in rural contexts must be culturally sensitive, with a research team that goes beyond the mere technicalities of collecting data. A culturally competent focus group can result when the research team has geographic, political, economic, and sociocultural knowledge related to the research area and its population. With extensive local collaboration, foreign researchers are better able to conduct data collection respectfully. The authors provide recommendations for future studies toward increasing the cultural appropriateness of focus groups in areas such as rural India. PMID- 12109728 TI - Crossed wires: interpreters, translators, and bilingual workers in cross-language research. AB - Increasingly, researchers are undertaking studies involving people who do not speak the same language as they do. Sociologists have long argued that language constructs the social world at the same time as it describes it. However, the implications of this for cross-language research are rarely considered. Employing interpreters/translators and "cultural brokers" in research raises methodological issues around the meanings of concepts and how to convey difference. Using a project that employed two Asian mental health workers, the author teases out some of the implications for research of language difference. She focuses both on the value of a biographical approach and on the problems such an approach presents. PMID- 12109729 TI - Matrix analysis as a complementary analytic strategy in qualitative inquiry. AB - In the current health care environment, researchers are asked to share meaningful results with interdisciplinary professional audiences, concerned community members, students, policy makers, planners, and financial officers. Emphasis is placed on effective health care outcomes and evidence, especially for underserved and vulnerable populations. Any research strategy that facilitates the clear, accurate communication of findings and voices will likely benefit groups targeted for intervention with scarce resources. In this example, matrix analysis contributes to the display, interpretation, pragmatic evaluation, and dissemination of findings in a study of rural elders. The author proposes matrix analysis as a strategy to advance knowledge and enhance the development of evidence in qualitative research. PMID- 12109730 TI - Measurement of triclosan in wastewater treatment systems. AB - The objective of this study was to investigate the fate and removal of triclosan (TCS; 5-chloro-2-[2,4-dichloro-phenoxy]-phenol), an antimicrobial agent used in a variety of household and personal-care products, in wastewater treatment systems. This objective was accomplished by monitoring the environmental concentrations of TCS, higher chlorinated derivatives of TCS (4,5-dichloro-2-[2,4-dichloro-phenoxy] phenol [tetra II]; 5,6-dichloro-2-[2,4-dichloro-phenoxy]-phenol [tetra III]; and 4,5,6-trichloro-2-(2,4-dichloro-phenoxy)-phenol [penta]), and a potential biotransformation by-product of TCS (5-chloro-2-[2,4-dicholoro-phenoxy]-anisole [TCS-OMe]) during wastewater treatment. These analytes were isolated from wastewater by using a C18 solid-phase extraction column and from sludge with supercritical fluid CO2. Once the analytes were isolated, they were derivatized to form trimethylsilylethers before quantitation by gas chromatography-mass spectrometry. Recovery of TCS from laboratory-spiked wastewater samples ranged from 79 to 88% for influent, 36 to 87% for final effluent, and 70 to 109% for primary sludge. Field concentrations of TCS in influent wastewater ranged from 3.8 to 16.6 microg/L and concentrations for final effluent ranged from 0.2 to 2.7 microg/L. Removal of TCS by activated-sludge treatment was approximately 96%, whereas removal by trickling-filter treatment ranged from 58 to 86%. The higher chlorinated tetra-II, tetra-III, and penta closans were below quantitation in all of the final effluent samples, except for one sampling event. Digested sludge concentrations of TCS ranged from 0.5 to 15.6 microg/g (dry wt), where the lowest value was from an aerobic digestion process and the highest value was from an anaerobic digestion process. Analysis of these results suggests that TCS is readily biodegradable under aerobic conditions, but not under anaerobic conditions. The higher chlorinated closans were near or below the limit of quantitation in all of the digested sludge samples. Based on results from this study, the chlorinated analogues and biotransformation by-product of TCS are expected to be very low in receiving waters and sludge-amended soils. PMID- 12109731 TI - Fate and effects of triclosan in activated sludge. AB - Triclosan (TCS; 5-chloro-2-[2,4-dichloro-phenoxy]-phenol) is a widely used antimicrobial agent. To understand its fate during sewage treatment, the biodegradation and removal of TCS were determined in activated sludge. In addition, the effects of TCS on treatment processes were assessed. Fate was determined by examining the biodegradation and removal of TCS radiolabeled with 14C in the 2,4-dichlorphenoxy ring in laboratory batch mineralization experiments and bench-top continuous activated-sludge (CAS) systems. In batch experiments with unacclimated sludge, TCS was mineralized to 14CO2, but the total yield varied as a function of test concentration. Systems that were redosed with TCS exhibited more extensive and faster mineralization, indicating that adaptation was a critical factor determining the rate and extent of biodegradation. In a CAS study in which the influent level of TCS was incrementally increased from 40 microg/L to 2,000 microg/L, removal of the parent compound exceeded 98.5% and removal of total radioactivity (parent and metabolites) exceeded 85%. Between 1.5 and 4.5% of TCS in the influent was sorbed to the wasted solids, whereas >94% underwent primary biodegradation and 81 to 92% was mineralized to CO2 or incorporated in biomass. Increasing levels of TCS in the influent had no major adverse effects on any wastewater treatment process, including chemical oxygen demand, biological oxygen demand, and ammonia removal. In a subsequent experiment, a CAS system, acclimated to TCS at 35 microg/L, received two separate 4-h shock loads of 750 microg/L TCS. Neither removal of TCS nor treatment processes exhibited major adverse effects. An additional CAS study was conducted to examine the removal of a low level (10 microg/L) of TCS. Removal of parent equaled 94.7%, and biodegradation remained the dominant removal mechanism. A subsequent series of CAS experiments examined removal at four influent concentrations (7.5, 11, 20, and 50 microg/L) of TCS and demonstrated that removal of parent ranged from 98.2 to 99.3% and was independent of concentration. Although TCS removal across all experiments appeared unrelated to influent concentration, removal was significantly correlated (r2 = 0.87) with chemical oxygen demand removal, indicating that TCS removal was related to overall treatment efficiency of specific CAS units. In conclusion, the experiments show that TCS is extensively biodegraded and removed in activated-sludge systems and is unlikely to upset sewage treatment processes at levels expected in household and manufacturing wastewaters. PMID- 12109732 TI - Aquatic toxicity of triclosan. AB - The aquatic toxicity of triclosan (TCS), a chlorinated biphenyl ether used as an antimicrobial in consumer products, was studied with activated-sludge microorganisms, algae, invertebrates, and fish. Triclosan, a compound used for inhibiting microbial growth, was not toxic to wastewater microorganisms at concentrations less than aqueous solubility. The 48-h Daphnia magna median effective concentration (EC50) was 390 microg/L and the 96-h median lethal concentration values for Pimephales promelas and Lepomis macrochirus were 260 and 370 microg/L, respectively. A no-observed-effect concentration (NOEC) and lowest observed-effect concentration of 34.1 microg/L and 71.3 microg/L, respectively, were determined with an early life-stage toxicity test with Oncorhynchus mykiss. During a 96-h Scenedesmus study, the 96-h biomass EC50 was 1.4 microg/L and the 96-h NOEC was 0.69 microg/L. Other algae and Lemna also were investigated. Bioconcentration was assessed with Danio rerio. The average TCS accumulation factor over the five-week test period was 4,157 at 3 microg/L and 2,532 at 30 microg/L. Algae were determined to be the most susceptible organisms. Toxicity of a TCS-containing wastewater secondary effluent to P. promelas and Ceriodaphnia was evaluated and no observed differences in toxicity between control and TCS treated laboratory units were detected. The neutral form of TCS was determined to be associated with toxic effects. Ionization and sorption will mitigate those effects in the aquatic compartment. PMID- 12109733 TI - Dynamic equilibrium dissolution of complex nonaqueous phase liquid mixtures into the aqueous phase. AB - Human health risks posed by hazardous substances seeping from a pool of nonaqueous phase liquids (NAPLs) into groundwater change over time because the more soluble compounds such as benzene, toluene, ethylbenzene, and xylene (BTEX) dissolve faster into the aqueous phase than less soluble compounds such as polycyclic aromatic hydrocarbons (PAH). Long-term dissolution from diesel fuel into the aqueous phase was determined experimentally in a continuous flow-through system using the slow-stirring method. The data obtained are interpreted using a dynamic equilibrium dissolution model based on Raoult's law. The predicted temporal development of aqueous concentrations are in good agreement with the experimental results. When a compound in the NAPL approaches complete depletion, a tailing behavior is observed, which is assigned to nonequilibrium effects, such as mass transfer limitations in the NAPL phase. The model predicted an increase of the mean molar mass of the diesel fuel of 1.5% over the entire experimental period. It should be noted that, if the dissolution process were to proceed further, the change in the mean molar mass could become significant and render the simple model inaccurate. Yet the simple model supports the assessment of initial action after a contamination event as well as the planning of long-term remedial strategies. PMID- 12109734 TI - Isolation of compounds from bleached kraft mill recovery condensates associated with reduced levels of testosterone in mummichog (Fundulus heteroclitus). AB - Previous studies have identified chemical recovery condensates as a primary source of hormonally active substances within a Canadian bleached kraft pulp mill. Although reverse osmosis treatment of condensates raises the exposure threshold that alters circulating levels of testosterone in mummichog (Fundulus heteroclitus), the responsible chemicals have not been characterized. In this study, a solid-phase extraction (SPE) method was developed to isolate chemical recovery condensate extractives for evaluation of hormonal activity. Condensates generated during softwood and hardwood pulp production were investigated for their relative potential to affect both circulating and gonadal production of sex steroids in mummichog. Mummichog were exposed to whole condensates, extracts from suspended particulates (> 1 microm), two fractions from SPE, and residual condensates after SPE. The distribution of bioactivity among condensate fractions was similar for both wood furnishes. In both sexes, significant depressions in circulating testosterone and in in vitro gonadal testosterone production were associated with exposure to whole condensates, particulate extracts, one SPE fraction, and residual material after SPE. An optimized SPE method subsequently demonstrated complete recovery of polar, bioavailable chemicals that reduced testosterone levels in both sexes. Characterizations of active fractions by gas chromatography-mass spectrometry (GC-MS) showed the presence of extractives with molecular masses > or = 240 amu possessing functionalities consistent with lignin degradation products. This study provides the first isolation of chemicals derived from pulp production associated with impaired reproductive performance in fish. PMID- 12109735 TI - Organochlorine concentrations in the Saimaa ringed seal (Phoca hispida saimensis) from Lake Haukivesi, Finland, 1981 to 2000, and in its diet today. AB - Organochlorine and mercury (Hg) pollution has been one possible reason for a decline in the size of the population of the Saimaa ringed seal (Phoca hispida saimensis) in Lake Haukivesi, Finland. In this study, we analyzed changes in the concentrations of organochlorine compounds (OCls) and extractable organic halogen (EOX) in the blubber of Saimaa ringed seals that died from 1981 to 2000. In addition, the present concentrations and biomagnification factors of OCls and Hg from the main prey fish of ringed seal and pike (Esox lucius) were determined. Concentrations of pollutants in bream (Abramis brama) also were analyzed. The decline in the OCl concentrations in blubber was, on average, 25% during the two decades since polychlorinated biphenyls and DDT compounds were banned. Today, concentrations in blubber of Saimaa ringed seals are similar of those in ringed seals from the Arctic. Correspondingly in fish, the OCl and Hg concentrations were, on average, similar to those found in other studies on Finnish and Arctic areas. The low diet concentrations and metabolism of OCls also reflect the lower age correlation of OCls in the male ringed seals in the 1990s than in the 1980s. Biomagnification was considerably higher from fish to the ringed seal than from prey fish to pike because of higher feeding rate of ringed seals and differences in the metabolisms of these animals. PMID- 12109736 TI - Modeling sorption isotherms of volatile organic chemical mixtures in model and natural solids. AB - Parameters from single-component isotherm models were used in multicomponent isotherm models to predict the aqueous phase sorption of trichloroethylene (TCE) in the presence of tetrachloroethylene (PCE) in four zeolites, Tenax, and three natural solids. The Langmuir, the Polanyi-Dubinin, and the Freundlich or the Langmuir-Freundlich isotherm models were used to simulate single-component sorption in zeolites. The Langmuir two-site, the Polanyi-Dubinin two-site, and the Freundlich or the Langmuir-Freundlich isotherm models were used to simulate single-component sorption in Tenax and natural solids. Two-site models have been used previously to model sorption in soils and sediments, and they combine an adsorption component (e.g., Langmuir) with a linear partitioning component. By using parameters from the different single-component isotherm models, the multicomponent Langmuir, the ideal adsorbed solution theory, and the Polanyi theory were each used to predict multicomponent sorption. In general, the ability to predict TCE sorption in the presence of PCE depended more on the choice of the single-component model than the multicomponent model, and better results were obtained when the Freundlich or the Langmuir-Freundlich isotherm was used for single-component sorption. This suggests that the more mechanistically based Langmuir and Polanyi-type models may not adequately describe the distribution of adsorption sites in some model and natural solids. The Freundlich or the Langmuir Freundlich model, although empirical, has greater flexibility in characterizing sorbent heterogeneity and results in better multicomponent model predictions. However, this last statement is tenuous, because more solids must be tested against various model combinations. PMID- 12109737 TI - Effect of adsorption to elemental iron on the transformation of 2,4,6 trinitrotoluene and hexahydro-1,3,5-trinitro-1,3,5-triazine in solution. AB - The effect of adsorption to elemental iron on the reductive transformation of 2,4,6-trinitrotoluene (TNT) and hexahydro-1,3,5-trinitro-1,3,5-triazine (royal demolition explosive [RDX]) in aqueous solution was studied with scrap iron and high-purity iron. In batch experiments with the same total iron surface area and a mixing rate of 100 rpm, TNT and RDX were removed from the solution within 30 min. With high-purity iron, adsorbed TNT was reduced to 2,4,6-triaminotoluene (TAT) rapidly, with little accumulation of intermediates at the surface. With scrap iron, the extent of adsorption of TNT and its daughter products was more significant and reduction of these adsorbed molecules to TAT was slower. Distribution of the intermediates indicated that the reaction with scrap iron occurred primarily through reduction of the ortho nitro group. Kinetic analysis suggests that mass transfer or adsorption of TNT controlled the overall rate of TNT reduction to TAT with pure iron, whereas with scrap iron, the rate of TAT formation was probably limited by other processes. Compared to TNT, transformation of adsorbed RDX was more rapid and less affected by iron type. The RDX was reduced to an unidentified, water-soluble intermediate and NH4+, which accounted for approximately 50% of the RDX nitrogen. No total organic carbon reduction was observed before and after RDX transformation with scrap iron. PMID- 12109738 TI - Adsorption behavior of toxic tributyltin to clay-rich sediments under various environmental conditions. AB - The adsorption and desorption behavior of tributyltin (TBT) from aqueous solution to clay-rich sediments was studied under various conditions (pH, salinity) using the batch technique. Sediments containing illite, kaolinite, and montmorillonite in different proportions were used as sorbent materials. Several physicochemical parameters of the sediments (e.g., Brunauer-Emmett-Teller [BET] surface area, cation exchange capacity [CEC], total organic carbon [TOC]) were evaluated to assess the influence of sediment characteristics to the adsorption capacity for TBT Adsorption isotherms were linear over the concentration range of 100 to 1,000 ng(Sn)/ml. The adsorption coefficient (Kd) values range from 29 to 70 at the pH value generally found in marine systems (pH 8). The adsorption capacity shows a maximum in the range of pH 6 and 7. Salinity is also an important factor in controlling TBT adsorption. The strongest adsorption was observed at salinity of 0/1000, and it strongly decreases with increasing salinity. The adsorption mechanism is controlled by the properties of the clay minerals as well as the aquatic chemistry of TBT. Desorption takes place over the studied pH range (4-8) when contaminated samples interact with TBT-free water at given experimental conditions. PMID- 12109739 TI - Resistance to cadmium and parasite infection are inversely related in two strains of a freshwater gastropod. AB - Phenotypes that are either resistant or susceptible to infection by the trematode parasite Schistosoma mansoni exist in the tropical freshwater snail Biomphalaria glabrata. We tested the hypothesis that a cost of parasite resistance in B. glabrata is greater sensitivity to cadmium toxicity, using parasite-resistant and parasite-susceptible strains exposed to cadmium in the laboratory. Survival analysis showed that time to death for cadmium was significantly shorter in eggs, juveniles, and adults of the parasite-resistant BS90 strain in comparison with the parasite-susceptible NMRI strain. Cadmium exposure increased time to hatch in both strains, but the effect was greater in BS90. Percentage hatch decreased with increased cadmium; BS90 was again more sensitive than NMRI. Comparison of the median effective concentration (EC50) for hatching and median lethal concentrations (LC50s) for survival of juveniles and adults showed that the order for cadmium resistance was adults > juveniles > eggs in NMRI and adults > eggs > juveniles in BS90. Cadmium resistance of F1 and F2 progeny of BS90/NMRI crosses was intermediate to that of parental strains. Numerical estimates indicated that a single genetic factor was responsible for the difference in cadmium resistance in the two strains. These findings were consistent with the hypothesis that greater sensitivity to cadmium is a cost of resisting parasitic infection. PMID- 12109740 TI - Reproductive characteristics of male mosquitofish (Gambusia affinis) inhabiting a small southeastern U.S. river receiving treated domestic sewage effluent. AB - A population of western mosquitofish (Gambusia affinis) living below a wastewater treatment plant in the vicinity of Birmingham (AL, USA) was studied for evidence of exposure to estrogens. Mosquitofish are sexually dimorphic live-bearing fish. Males have an elongated and modified anal fin, called a gonopodium, used in mating. It has been hypothesized that exposure to estrogens and/or anti-androgens in treated wastewater might inhibit the androgen-dependent development of the gonopodium. The population in this study showed no evidence of having been exposed to endocrine-disrupting chemicals. The difference in adjusted (for overall fish size via analysis of covariance) mean gonopodium length between effluent-exposed and control populations was not significant. No detectable levels of vitellogenin were observed in the blood of any of the male mosquitofish, either from the effluent-exposed or the control population. Testes and livers were weighed and examined histologically. The fish exposed to treated wastewater effluent had significantly larger adjusted mean weights for both testis and liver, but no histological changes indicating exposure to estrogens were observed. PMID- 12109741 TI - Dose-response and time course relationships for vitellogenin induction in male western fence lizards (Sceloporus occidentalis) exposed to ethinylestradiol. AB - The long-term goal of this research is to develop and validate an in vivo reptile model for endocrine-mediated toxicity using fence lizards (Sceloporus spp.). One of the best defined estrogenic responses in oviparous vertebrates is induction of the yolk precursor protein, vitellogenin (Vtg). In this study, dose-response and time course relationships for Vtg induction were determined in male western fence lizards (Sceloporus occidentalis) given intraperitoneal injections of 17alpha ethinylestradiol (EE2). Plasma Vtg was quantified directly with an antibody capture enzyme-linked immunosorbent assay (ELISA) and indirectly using plasma alkaline-labile phosphate (ALP) in order to compare these two methods. Both ELISA and ALP predicted similar median effective dose (ED50 [dose causing a 50% maximal response]) values for plasma Vtg induction (0.167 mg/kg for ELISA and 0.095 mg/kg for ALP). In addition, both ELISA and ALP detected significant Vtg induction at a dose of 0.0003 mg/kg of EE2, which was the lowest dose used in our study. A decrease in body weight at the highest dose (10 mg/kg) and an increase in hepatosomatic index at the four highest doses were observed. Serial dilutions of plasma from an EE2-exposed male revealed a high correlation between plasma Vtg and ALP determinations in this species. In conclusion, our data show that plasma ALP may be a suitable alternative for measuring plasma Vtg compared with developing a Vtg ELISA in fence lizards exposed to estrogenic compounds. PMID- 12109742 TI - The effects of polychlorinated biphenyls (Aroclor 1242) on thyroxine, estradiol, molt, and plumage characteristics in the American kestrel (Falco sparverius). AB - The purpose of this experiment was to determine the effects of Aroclor 1242, a mixture of polychlorinated biphenyls (PCBs), on plumage characteristics and molt in the American kestrel, Falco sparverius. Several characteristics of plumage, including color and molt schedule, are modulated by hormonal signals and hence may be modified by endocrine-active contaminants. If so, the functions of plumage (e.g., communication for mating or territorial defense) may be compromised by exposure to such compounds. Captive American kestrels were fed Aroclor 1242 at 0, 6.0, and 60.0 ppm (n = 6 males and 6 females per treatment) mixed in their normal diet. Concentrations of plasma estradiol and thyroxine were measured weekly from the beginning of treatment. Measured plumage characteristics included width of the black subterminal band on the tail, color (a composite index of hue and saturation), reflectance from 230 to 800 nm. pattern of feather loss and regrowth on the tail and wing, and timing of onset and duration of molt. Aroclor 1242 depressed plasma thyroxine. Plasma estradiol levels remained low due to the phase of the breeding cycle. Treatments did not disrupt the measured plumage characteristics. This may be due to timing or dose of exposure or to genetic factors. PMID- 12109743 TI - A field validation of two sediment-amphipod toxicity tests. AB - A field validation study of two sediment-amphipod toxicity tests was conducted using sediment samples collected subtidally in the vicinity of a polycyclic aromatic hydrocarbon (PAH)-contaminated Superfund site in Elliott Bay (WA, USA). Sediment samples were collected at 30 stations with a 0.1 m2 grab from which subsamples were taken for sediment toxicity testing and geochemical and macrofaunal analyses. Standard 10-d sediment-amphipod toxicity tests were conducted with Rhepoxynius abronius and Leptocheiros plumulosus. Sediments were analyzed for 33 PAHs, pentachlorophenol, polychlorinated biphenyls, acid-volatile sulfide, simultaneously extracted metals (Cd, Cu, Zn, Pb, Ni), total organic carbon, and grain size. Sediment temperature, oxygen-reduction potential, water depth, and interstitial water salinity were also measured. Polycyclic aromatic hydrocarbons, quantified as total PAH toxic units (TU(PAH)), were confirmed to be an important common causal agent of the changes in the two toxicity test (% survival R. abronius, % survival L. plumulosus) and five macrofaunal community (number of species, S; numerical abundance, A: total biomass, B: Swartz's dominance index, SDI; Brillouin's index, H) endpoints. Two other macrofaunal community metrics (the complement of Simpson's index, 1 - SI, and McIntosh's index, MI) were less sensitive to TU(PAH) than the two toxicity test endpoints. The sensitivities of R. abronius and L. plumulosus to TU(PAH) were statistically indistinguishable. Field validations were conducted by testing the association between or among each toxicity test endpoint, each of seven macrofaunal community metrics (S, A, B, SDI, H, 1 - SI, MI), and TU(PAH) by (1) Spearman's coefficient of rank correlation, (2) Kendall's coefficient of concordance, (3) G tests of independence, and (4) regression analysis. Some field validations based on multivariable tests of association (e.g., points 2 and 3) among toxicity test, field, and stressor endpoints produced false positive results. Both toxicity test endpoints were validated as indicators of changes in S, A, SDI, and H by all the methods tested. The resolution power of the relationships between the laboratory toxicity test and macrofaunal field endpoints was low (< or = three classes) but sufficient to discriminate ecologically important effects. We conclude that standard sediment-amphipod toxicity tests are ecologically relevant and that, under the proper conditions, their results can be used for lab-to-field extrapolation. PMID- 12109744 TI - Hydrocarbon composition and toxicity of sediments following the Exxon Valdez oil spill in Prince William Sound, Alaska, USA. AB - An 1-year study of the 1989 Exxon Valdez oil spill found that spill residues on the oiled shorelines rapidly lost toxicity through weathering. After 1990, toxicity of sediments remained at only a few heavily oiled, isolated locations in Prince William Sound (AK, USA), as measured by a standard amphipod bioassay using Rhepoxynius abronius. Data from 648 sediment samples taken during the 1990 to 1993 period were statistically analyzed to determine the relationship between the total concentration of 39 parent and methyl-substituted polycyclic aromatic hydrocarbons (defined as total polycyclic aromatic hydrocarbons [TPAH]) and amphipod mortality and the effect of oil weathering on toxicity. A logistic regression model yielded estimates of the lower threshold, LC10 (lethal concentration to 10% of the population), and LC50 (median lethal concentration) values of 2,600, 4,100, and 10,750 ng/g TPAH (dry wt), respectively. Estimates of the threshold and LC50 values in this field study relate well to corresponding sediment quality guideline (SQG) values reported in the literature. For sediment TPAH concentrations >2,600 ng/g, samples with high mortality values (>90%) had relatively high fractions of naphthalenes and those with low mortality (<20%) had relatively high fractions of chrysenes. By 1999, the median sediment TPAH concentration of 117 ng/g for the post-1989 worst-case sites studied were well below the 2,600 ng/g toxicity threshold value, confirming the lack of potential for long-term toxic effects. Analysis of biological community structure parameters for sediment samples taken concurrently found that species richness and Shannon diversity decreased with increasing TPAH above the 2,600 ng/g threshold, demonstrating a correspondence between sediment bioassay results and biological community effects in the field. The low probability of exposure to toxic concentrations of weathered spill residues at the worst-case sites sampled in this study is consistent with the rapid overall recovery of shoreline biota observed in 1990 to 1991. PMID- 12109745 TI - Accumulation of tributyltin in the blood of fish: its application for monitoring in the marine environment. AB - Accumulation of tributyltin (TBT) in serum, liver, muscle, and gill of olive flounder (Paralichthys olivaceus) was examined in a 30-d static-renewal exposure. Tributyltin accumulated rapidly in the serum of the olive flounder and to a greater extent than in the other tissues. The accumulated TBT concentrations in tissues were in the order of serum > gill > liver > muscle on a dry-weight basis. Tributyltin also was detected in the serum of feral fine-spotted flounder, Pleuronichthys cornutus, collected from the coastal area. The mean TBT concentration in serum (2,470 ng Sn/g) of the fine-spotted flounder was about 40 times higher than that in the liver (60 ng Sn/g) and 200 times higher than that in the muscle (27 ng Sn/g) on a dry-weight basis. The TBT concentrations in serum and sediment demonstrated a positive correlation. The percent TBT composition to total butyltin was much higher in the serum (71%) than in the other tissues and sediment (<47%). These results suggest that the analysis of fish blood serum could be a useful tool for monitoring exposure to TBT in the marine environment. PMID- 12109746 TI - An assessment of in vitro androgenic activity and the identification of environmental androgens in United Kingdom estuaries. AB - Environmental androgens are a group of compounds that to date have received very little attention. In this study, a yeast-based androgen screen (YAS) was used to determine the level of in vitro androgenic activity in seven United Kingdom estuaries. Surface water, sediment pore water, and sediment particulate material solvent extracts collected from Southampton Water, the Thames, Mersey, Tees, Tyne, Clyde, and Forth were tested for in vitro androgenic activity. Eleven of the 41 surface water samples collected displayed androgenic activity >2 ng dihydrotestosterone (DHT) equivalents/L (3-9 ng DHT/L), while eight of the 39 sediment pore waters collected showed activity >45 ng DHT/L (51-187 ng DHT/L). High levels of androgenic activity were determined in the solvent extracts of sediments, with 10 of 39 samples exhibiting a level of androgenic activity >454 ng DHT/kg (1,020-15,300 ng DHT/kg). In vitro YAS testing of five selected sewage treatment works (STW) effluents entering these estuaries showed that measurable levels (34-635 ng DHT/L) of androgenic activity were observed in those receiving only primary treatment (Howdon STW and Irvine Valley Sewer) at the time of the survey. A toxicity identification evaluation (TIE) study of Irvine Valley Sewer effluent using the YAS assay was used to identify the natural steroids/steroid metabolites dehydrotestosterone, androstenedione, androstanedione, 5beta androstane-3alpha,11beta-diol-17-one, androsterone, and epi-androsterone as responsible for 99% of the in vitro activity determined in the effluent. PMID- 12109747 TI - Application of postexposure feeding depression bioassays with Daphnia magna for assessment of toxic effluents in rivers. AB - A bioassay that used postexposure feeding depression in Daphnia magna Straus as an endpoint previously had been developed under laboratory conditions. Laboratory studies revealed that this was a sensitive, robust endpoint, which could potentially be used in an in situ bioassay. This study adapted the laboratory bioassay for use in the field and deployed D. magna in situ at four known or suspected contaminated and reference sites. The bioassay was demonstrated to be reliable for use in the field because more than 90% of test organisms were recovered live from the test chambers after exposure allowing feeding rates to be measured after exposure. At each of the contaminated sites, significant depressions in postexposure feeding rates were recorded. Although depressions in postexposure feeding rates were apparent at all contaminated sites, with the exception of Langholm, no impacts were detected on the benthic macroinvertebrate community, when using the Biological Monitoring Working Party scoring system. This demonstrated that during this study, post-exposure feeding depression was a more reliable and sensitive endpoint to use to detect toxicity than were changes in community structure. Therefore, the postexposure feeding depression bioassay can offer a sensitive, robust, ecologically relevant diagnostic endpoint for use in water-quality assessment schemes. PMID- 12109748 TI - Acute toxicity and cholinesterase inhibition in larval and early juvenile walleye exposed to chlorpyrifos. AB - Application of organophosphorus (OP) insecticides to agricultural fields during the spring often overlaps the period of spawning, egg incubation, and larval stages of fish species such as walleye (Stizostedion vitreum). Because life stage can affect uptake, distribution, and effects of contaminants, our objective was to identify the most sensitive life stage of walleye tochlorpyrifos, a broad spectrum OP insecticide. Prolarvae (yolk sac, endogenous feeding stage) were least sensitive to chlorpyrifos (median lethal concentration [LC50] = 225-316 microg/L), and postlarvae I (oil globule, exogenous feeding stage) were less sensitive (LC50 = 24-29 microg/L) than postlarvae II (oil globule absent; LC50 = 12-13 microg/L). The differences in sensitivity of the larval stages coincide with stages of gill development. Gill filaments were absent until the end of the prolarval stage, and development of secondary lamellae did not occur until the end of the postlarval I stage. Juvenile fish were less sensitive than postlarval stages, but did not differ significantly among the juvenile ages tested. The LC50s ranged from 37 to 45 microg/L for 30- and 90-d-old juvenile walleye, respectively. Larval walleye survived when cholinesterase (ChE) activity was inhibited by as much as 90%; however, 60- and 90-d-old juvenile walleye did not survive when ChE activity was inhibited more than 71%. PMID- 12109749 TI - Toxicity of the explosives 2,4,6-trinitrotoluene, hexahydro-1,3,5-trinitro-1,3,5 triazine, and octahydro-1,3,5,7-tetranitro-1,3,5,7-tetrazocine in sediments to Chironomus tentans and Hyalella azteca: low-dose hormesis and high-dose mortality. AB - The toxicity of the explosives 2,4,6-trinitrotoluene (TNT); hexahydro-1,3,5 trinitro-1,3,5-triazine (royal demolition explosive [RDX]); and octahydro-1,3,5,7 tetranitro-1,3,5,7-tetrazocine (high-melting explosive [HMX]), was evaluated in spiked sediment with two freshwater invertebrates. The midge Chironomus tentans and the amphipod Hyalella azteca demonstrated significant toxic effects after exposure to TNT and its degradation products, 1,3,5-trinitrobenzene (TNB) and 2,4 diamino-6-nitrotoluene (2,4-DANT). Significant reductions in survival of C. tentans exposed to TNT, TNB, and 2,4-DANT were observed at nominal sediment concentrations as low as 200 mg/kg. Hyalella azteca was more sensitive to TNT, TNB, and 2,4-DANT than the midge, where significant reductions in survival were observed at nominal concentrations of 50, 100, and 200 mg/kg, respectively. Survival of the midge and the amphipod was unaffected after exposure to RDX or HMX at the highest concentrations of 1,000 and 400 mg/kg, respectively. Growth of the midge, measured as total weight, was significantly reduced by 2,4-DANT. However, significantly increased growth was observed after exposure to sublethal concentrations of RDX and HMX. Although significant reductions in amphipod survival were observed at high concentrations of TNB, growth was significantly increased at sublethal concentrations. The results of the current investigation suggest that organisms exposed to explosives at contaminated sites may be affected at concentrations less than 25 mg/kg through hormetic growth enhancement and at higher concentrations through increased mortality. PMID- 12109750 TI - Accumulation and elimination of lanthanum by duckweed (Lemna minor L.) as influenced by organism growth and lanthanum sorption to glass. AB - Lanthanide emissions to the environment increase as a result of the growing industrial applications of these elements. However, robust data to evaluate the environmental fate of lanthanides are scarce. This article describes the accumulation and elimination of lanthanum (La) by common duckweed (Lemna minor L.). Speciation modeling was performed to assure that solubility products were not exceeded. It also showed that La was predominantly associated with ethylenediaminetetraacetic acid (EDTA). Lanthanum concentrations in plants and medium and the amounts sorbed to glass vessels were quantified by using the radioisotope 140La. The amount of La adsorbed on the glass reached values of 25% of the total La present. A model was formulated to describe La uptake in exponentially growing duckweed in the presence of an adsorptive surface. Growth induced dilution appeared more efficient in lowering plant La concentrations than actual elimination. An elimination study revealed two compartments, of which the smallest eliminated 50 times faster than the bigger compartment, which eliminated mainly by growth dilution. The average bioconcentration factor was 2,000 L/kg fresh weight and 30,000 L/kg dry weight, comparable with those of other higher plants. At the applied concentration of 10 nM, no effects were observed on duckweed growth. However, the high bioconcentration factor warrants monitoring of lanthanide emissions. PMID- 12109751 TI - Ecological impact and environmental fate of perfluorooctane sulfonate on the zooplankton community in indoor microcosms. AB - There is presently a substantial amount of information being gathered concerning the environmental risk associated with the perfluorooctane sulfonate (PFOS) compound. The U.S. Environmental Protection Agency (U.S. EPA) is requiring that more research be completed before making definitive decisions concerning the regulatory issues covered in the significant new use rule (18/10-2000) under the Toxic Substance Control Act. However, there are no risk assessment requirements under seminatural conditions in microcosms. The PFOS can enter, and has been found in, the aquatic environment through different pathways, including spills associated with use of fire-fighting foams containing PFOS, leaching from washing Scotchgard-treated clothes with the wastewater, leaching from various coatings, discharges as residual waste from fluorochemical production, or volatilization and transportation atmospherically. The biota is the sink of PFOS rather than the sediment or soil. The aim of this article is to determine a 35-d community no observable-effect concentration (NOECcommunity) for freshwater zooplankton and the fate of PFOS during the course of study. The PFOS persisted in the water phase with only slight reductions over the study; only the decrease from 33.9 mg/L at day 1 to 29.8 mg/L at day 35 was significant. A 90 to 100% reduction (p < 0.01) of the total zooplankton population was found after one week of exposure to 30 mg PFOS/L and a similar reduction after two weeks at 10 mg PFOS/L. The Daphnia magna 21-d NOECsurvival of 12 mg/L has previously been found in a standard laboratory bioassay by 3M. The rank order of susceptibility for the test community was Copepoda > Cladocera > Rotifera, assuming all adverse direct effects. PMID- 12109752 TI - Estimating the probability of bird mortality from pesticide sprays on the basis of the field study record. AB - The outcome of avian field studies was examined to model the likelihood of mortality. The data were divided into clusters reflecting the type of pesticide application and bird guilds present on site. Logistic regression was used to model the probability of a bird kill. Four independent variables were tested for their explanatory power: a variable reflecting acute oral toxicity and application rate; a variable reflecting the relative oral to dermal toxicity of the pesticides; Henry's law constant; and a variable reflecting possible avoidance of contaminated food items, the hazard factor (HF). All variables except for HF significantly improved model prediction. The relative dermal to oral toxicity, especially, was shown to have a major influence on field outcome and clearly must be incorporated into future avian risk assessments. The probability of avian mortality could be calculated from a number of current pesticide applications and the conclusion was made that avian mortality occurs regularly and frequently in agricultural fields. PMID- 12109753 TI - Increased toxicity to invertebrates associated with a mixture of atrazine and organophosphate insecticides. AB - This study examined the joint toxicity of atrazine and three organophosphate (OP) insecticides (chlorpyrifos, methyl parathion, and diazinon) exposed to Hyalella azteca and Musca domestica. A factorial design was used to evaluate the toxicity of binary mixtures in which the lethal concentration/lethal dose (LC1/LD1, LC5/LD5, LC15/LD15, and LC50/LD50) of each OP was combined with atrazine concentrations of 0, 10, 40, 80, and 200 microg/L for H. azteca and 0, 200, and 2,000 ng/mg for M. domestica. Atrazine concentrations (> or = 40 microg/L) in combination with each OP caused a significant increase in toxicity to H. azteca compared with the OPs dosed individually. Acetylcholinesterase (AChE) activity also was examined for the individual OPs with and without atrazine treatment. Atrazine in combination with each of the OPs resulted in a significant decrease in AChE activity compared with the OPs dosed individually. In addition, H. azteca that were pretreated with atrazine (> or = 40 microg/L) were much more sensitive to the OP insecticides compared with H. azteca that were not pretreated with atrazine before being tested. Topical exposure to atrazine concentrations did not significantly increase OP toxicity to M. domestica. The results of this study indicate the potential for increased toxicity in organisms exposed to environmental mixtures. PMID- 12109754 TI - Better bootstrap estimation of hazardous concentration thresholds for aquatic assemblages. AB - The introduction of species sensitivity distribution (SSD) approaches to ecological risk assessment offers the potential for a more transparent scientific basis for the derivation of predicted no-effect concentrations. However, conventional SSD methodologies have relied on standard distributions (e.g., log logistic, log normal) that are not necessarily based on sound ecological or statistical grounds. More recently, bootstrap resampling techniques that do not rely on distributional assumptions have been applied to the problem. Here we describe how a more advanced bootstrap methodology may be applied to derive better point estimates and confidence intervals for SSD estimates of safe environmental concentrations. Motivated by the fact that the true SSD may not fit any standard model category, we go on to consider a hybrid bootstrap regression approach. This can yield a substantially different estimate for the SSD when compared with both the basic bootstrap and the more frequently used parametric curve approaches. With increasing use of SSDs in ecological risk assessment, it is now imperative that the scientific community develops agreement over appropriate methods for their derivation. PMID- 12109755 TI - Predicting the occurrence of genetic adaptation to dioxinlike compounds in populations of the estuarine fish Fundulus heteroclitus. AB - A population of the nonmigratory estuarine fish species Fundulus heteroclitus (mummichog) indigenous to a polychlorinated biphenyl (PCB)-contaminated Superfund site (New Bedford Harbor, MA, USA) demonstrated an inherited tolerance to local, dioxinlike contaminants (DLCs). These findings suggest that DLCs have acted as selective agents, allowing the survival of only the most tolerant individuals, forming DLC-adapted populations. We hypothesized that DLC-tolerant mummichog populations would reside where local conditions are toxic to sensitive individuals, and that toxic environmental conditions could be predicted based on responses of sensitive early life stages to laboratory exposures of DLCs. As a measure of DLC tolerance, progeny of field-collected fish were tested in the laboratory with a dioxinlike PCB congener, 3,3',4,4',5-pentachlorobiphenyl (PCB 126). Mummichog populations were collected from sites with sediment PCB concentrations predicted to range from nontoxic to toxic. Consistent with predictions, tolerant populations were indigenous to sites with elevated sediment PCB concentrations. Also, as predicted, DLC-tolerant populations were resident to sites far less contaminated than the Superfund site. These results suggest that exposures to persistent, bioaccumulative, and toxic contaminants may produce evolutionary effects on a geographic scale larger than previously envisioned. This study presents an approach and describes a model system that may improve understanding of the scale of occurrence for these potentially irreversible ecological effects. PMID- 12109756 TI - The influence of word frequency on recency effects in directed free recall. AB - Leading theoretical explanations of recency effects are designed to explain the reported absence of a word frequency effect on recall of words from recency serial positions. The present study used a directed free-recall procedure (J. J. Dalezman, 1976) and manipulated the frequency composition of the word lists (pure and mixed). Overall, with pure lists, a greater proportion of high-frequency (HF) words were recalled than low-frequency (LF) words, and with mixed lists, a greater proportion of LF words were recalled than HF words. Of importance, this recall advantage for one frequency over the other as a function of list composition was evident across the last three serial positions, indicating an influence of word frequency on recency effects that is dependent on the frequency composition of the lists. These results challenge one of the major assumptions on which several theories of recency effects have been based. PMID- 12109757 TI - List composition and the word-frequency effect for recognition memory. AB - The attention/likelihood theory (ALT; M. Glanzer & J. K. Adams, 1990) and the retrieving effectively from memory (REM) theory (R. M. Shiffrin & M. Steyvers, 1997) make different predictions concerning the effect of list composition on word recognition. The predictions were empirically tested for two-alternative forced-choice, yes-no, and ratings recognition tasks. In the current article, the authors found that discrimination of low-frequency words increased as the proportion of high-frequency words studied increased. The results disconfirm the ALT prediction that recognition is insensitive to list composition, and they disconfirm the predictions of the REM model described by R. M. Shiffrin and M. Steyvers (1997). The current authors discuss a slightly modified version of REM that can better predict our findings, and we discuss the challenges the present findings pose for ALT and REM. PMID- 12109758 TI - When is schematic knowledge used in source monitoring? AB - Source monitoring involves judgments regarding the origin of information (M. K. Johnson, S. Hashtroudi, & D. S. Lindsay, 1993). When participants cannot remember the source in a source-monitoring task, they may guess according to their prior schematic knowledge (U. J. Bayen, G. V. Nakamura, S. E. Dupuis, & C.-L. Yang, 2000). The present study aimed at specifying conditions under which schematic knowledge is used in source monitoring. The authors examined the time course of schema-based guesses with a response-signal technique (A. V. Reed, 1973), and multinomial models that separate memory and guessing bias. Use of schematic knowledge was observed only when asymptotic old-new recognition was low. The time course of schematic-knowledge retrieval followed an exponential growth function. Implications for theories of source monitoring are discussed. PMID- 12109759 TI - The dynamics of intention retrieval and coordination of action in event-based prospective memory. AB - Event-based prospective memory requires responding to cues in the environment that are associated with a previously established intention. Some researchers believe that intentions reside in memory with an above baseline level of activation, a phenomenon called the intention superiority effect. The authors of this study predicted that intention superiority would be masked by additional cognitive processes associated with successful event-based prospective memory. These additional processes include noticing the cue, retrieving the intention, and coordinating intention execution with the ongoing activity. In 3 experiments, intention superiority was demonstrated by faster latencies to the ongoing activity on failed prospective trials and the existence of the additional processes was demonstrated by slower latencies on successful trials. This study demonstrates the importance of investigating the microstructure of the cognitive components involved with processing and responding to an event-based prospective memory cue. PMID- 12109760 TI - The length of the retention interval, forgetting, and subjective similarity. AB - A long retention interval tends to result in the poor retention known as forgetting. A high subjective similarity between stimuli frequently produces their poor retention. Thus, a long retention interval may increase the subjective similarity between stimuli (the RIISS hypothesis), and this increase may produce forgetting. To examine this hypothesis, college students made speeded same different discriminations between two lines or tones of different lengths or frequencies that were 400 ms or 3,300 ms apart, and they rated the similarity of these stimuli. The long interval produced poorer overall performance as expected, but also produced poorer performance on different than same stimuli, implying that it increased the subjective similarity between the initial and subsequent stimuli, and it also increased rated similarity, in support of the RIISS hypothesis. The position that stored stimuli lose less common information than distinctive information explains RIISS evidence better than does perturbation theory. PMID- 12109761 TI - Representational change and analogy: how analogical inferences alter target representations. AB - The ways that analogy alters the representation of target information was investigated in 4 experiments. Participants read information about a target, followed by a potential source analog. Participants later completed a recognition test in which some of the sentences were old, some novel, and some analogical inferences that were not seen before. Participants who read the description of a source analog erroneously recognized analogical inferences as being in the target description. The effect occurred with different delays between study and test and with an unfamiliar target domain. It also occurred when source and target shared few superficial features. Reading-time data suggest that participants were drawing analogical inferences when encoding the source. Overall, these experiments show that analogical inferences are incorporated in the representation of the target and cannot be differentiated from information actually presented. PMID- 12109762 TI - Ambiguity and synonymy effects in lexical decision, naming, and semantic categorization tasks: interactions between orthography, phonology, and semantics. AB - In this article, ambiguity and synonymy effects were examined in lexical decision, naming, and semantic categorization tasks. Whereas the typical ambiguity advantage was observed in lexical decision and naming, an ambiguity disadvantage was observed in semantic categorization. In addition, a synonymy effect (slower latencies for words with many synonyms than for words with few synonyms) was observed in lexical decision and naming but not in semantic categorization. These results suggest that (a) an ambiguity disadvantage arises only when a task requires semantic processing, (b) the ambiguity advantage and the synonymy disadvantage in lexical decision and naming are due to semantic feedback, and (c) these effects are determined by the nature of the feedback relationships from semantics to orthography and phonology. PMID- 12109763 TI - Flexible use of recent information in causal and predictive judgments. AB - Associative and statistical theories of causal and predictive learning make opposite predictions for situations in which the most recent information contradicts the information provided by older trials (e.g., acquisition followed by extinction). Associative theories predict that people will rely on the most recent information to best adapt their behavior to the changing environment. Statistical theories predict that people will integrate what they have learned in the two phases. The results of this study showed one or the other effect as a function of response mode (trial by trial vs. global), type of question (contiguity, causality, or predictiveness), and postacquisition instructions. That is, participants are able to give either an integrative judgment, or a judgment that relies on recent information as a function of test demands. The authors concluded that any model must allow for flexible use of information once it has been acquired. PMID- 12109764 TI - Within- and between-language priming differ: evidence from repetition of pictures in Spanish-English bilinguals. AB - In the current study, the authors used an immediate repetition paradigm with pictures to observe whether repetition enhances word production in bilinguals. In Experiment 1, participants were asked to name pictures that were named previously in the same language (Spanish-Spanish or English-English) or in the opposite language (Spanish-English or English-Spanish). Results revealed a repetition effect both within languages and between languages. Furthermore, there was an asymmetry within language, with repetition priming being larger in Spanish than in English. Experiments 2 and 3 revealed that lag interacted with language for both within- and between-language priming. However, lag resulted in a decrease in the asymmetry for within- but not between-language priming. The results are consistent with the view that within- and between-language repetition priming are mediated by different mechanisms. PMID- 12109765 TI - The influence of phonological similarity neighborhoods on speech production. AB - The influence of phonological similarity neighborhoods on the speed and accuracy of speech production was investigated with speech-error elicitation and picture naming tasks. The results from 2 speech-error elicitation techniques-the spoonerisms of laboratory induced predisposition technique (B. J. Baars, 1992; B. J. Baars & M. T. Motley, 1974; M. T. Motley & B. J. Baars, 1976) and tongue twisters-showed that more errors were elicited for words with few similar sounding words (i.e., a sparse neighborhood) than for words with many similar sounding words (i.e., a dense neighborhood). The results from 3 picture-naming tasks showed that words with sparse neighborhoods were also named more slowly than words with dense neighborhoods. These findings demonstrate that multiple word forms are activated simultaneously and influence the speed and accuracy of speech production. The implications of these findings for current models of speech production are discussed. PMID- 12109766 TI - The processing role of structural constraints on the interpretation of pronouns and anaphors. AB - The authors report 6 self-paced word-by-word reading studies of how morphosyntactic agreement, focus status, and the structural constraints of binding theory apply and interact during the online interpretation of pronouns (e.g., him, her) and anaphors (e.g., himself, each other). Previous studies held that structural conditions on coreference work as interpretive filters that impose exceptionless limits on which antecedent candidates can be evaluated by subsequent, content-based processes. These experiments instead support an interactive-parallel-constraint model, in which multiple weighted constraints (including constraints on binding) simultaneously influence the net activation of a candidate during preselection stages of antecedent evaluation. Accordingly, structurally inaccessible candidates can interfere with antecedent selection if they are both prominent in focus structure and gender-number compatible with the pronoun or anaphor. PMID- 12109767 TI - Temporal order relations in language comprehension. AB - The role of temporal orientation (chronological or reverse) and chronological distance (close, intermediate, or distant) in general event knowledge on language comprehension was examined. Experiment 1 used a relation-recognition paradigm in which the comprehension of a target event could be facilitated or disrupted by the temporal orientation implied by the prior information. Experiments 2 and 3 used a sentence-probe-recognition paradigm in which the temporal orientation, the stimulus onset asynchrony, and the chronological distance between the sentence event and the probe event were manipulated. The results demonstrated that readers used temporal information conveyed by their knowledge to construct situation models while comprehending sentences. The internal temporal dimension appeared to be directional and reflected the chronological distance between everyday events. PMID- 12109768 TI - Retrieval of concepts in script-based texts and narratives: the influence of general world knowledge. AB - The goal of the present experiments was to examine the accessibility of concepts embedded within text. J. A. Albrecht and E. J. O'Brien (1991) found that with narrative texts central concepts are more quickly retrieved than peripheral concepts. In contrast, F. R. Yekovich and C. H. Walker (1986) found that when concepts are embedded within scripts, peripheral concepts are more quickly retrieved than central concepts. Over 13 experiments, central concepts were generally more quickly accessed than peripheral concepts. The only exception occurs when (a) script-based texts are used, (b) the number of mentions or elaboration of central and peripheral concepts is held constant, and (c) the measure of centrality is theme relatedness rather than degree of interconnectedness. Under those conditions, central concepts are more difficult to access than peripheral concepts. PMID- 12109769 TI - Dynamics and constraints in insight problem solving. AB - This article reports 2 experiments that investigated performance on a novel insight problem, the 8-coin problem. The authors hypothesized that participants would make certain initial moves (strategic moves) that seemed to make progress according to the problem instructions but that nonetheless would guarantee failure to solve the problem. Experiment 1 manipulated the starting state of the problem and showed that overall solution rates were lower when such strategic moves were available. Experiment 2 showed that failure to capitalize on visual hints about the correct first move was also associated with the availability of strategic moves. The results are interpreted in terms of an information processing framework previously applied to the 9-dot problem. The authors argue that in addition to the operation of inappropriate constraints, a full account of insight problem solving must incorporate a dynamic that steers solution-seeking activity toward the constraints. PMID- 12109770 TI - Distinguishing prototype-based and exemplar-based processes in dot-pattern category learning. AB - The authors contrast exemplar-based and prototype-based processes in dot-pattern categorization. In Experiments 1A and 1B, participants provided similarity ratings of dot-distortion pairs that were distortions of the same originating prototype. The results show that comparisons to training exemplars surrounding the prototype create flat typicality gradients within a category and small prototype-enhancement effects, whereas comparisons to a prototype center create steep typicality gradients within a category and large prototype-enhancement effects. Thus, prototype and exemplar theories make different predictions regarding common versions of the dot-distortion task. Experiment 2 tested these different predictions by having participants learn dot-pattern categories. The steep typicality gradients, the large prototype effects, and the superior fit of prototype models suggest that participants refer to-be-categorized items to a representation near the category's center (the prototype), and not to the training exemplars that surround the prototype. PMID- 12109771 TI - Current trends in research into the waterborne parasite Giardia. AB - The waterborne flagellated parasite Giardia intestinalis continues to be the most frequent protozoan agent of intestinal disease world-wide, causing an estimated 2.8 x 10(8) cases per annum. Severe symptoms of diarrhea and sickness can be persistent and even life threatening in the immunocompromised, in infants, and in the aged, although self-limiting in the majority of patients. Despite a growing awareness and intensified research many uncertainties remain, especially with respect to the risk of potential zoonotic transmission. Water supplies can be monitored for cysts using automated cytofluorimetric immunoassays, but this does not measure infectivity. Filtration provides the best protection, because cysts are highly resistant to chlorine and ozone. Other incompletely elucidated aspects include mechanisms of pathogenicity, host reaction to infection, immunity and parasite control using vaccines or antigiardial compounds; the 5-nitroimidazole metronidazole is the most effective of these. Molecular typing of various isolates indicates that most animal parasites are not infective to humans, but those that are can be genotypically classified as assemblage A or B. The phylogeny of the organism remains uncertain, but there is a growing opinion that Giardia is not an ancient primitive eukaryote, but that it is derived from a more complex mitochondria-containing protozoon. PMID- 12109772 TI - Ribonucleases from T2 family. AB - Ribonucleases are ubiquitous in distribution. Ribonucleases that hydrolyse RNA to 3' mononucleotides via 2', 3' cyclic nucleotides are classified into three groups, RNase A, RNase T1, and RNase T2 families. Apart from salvage of cellular or extracellular RNAs, RNases participate in vital cellular functions such as DNA replication, transcription and RNA processing, splicing and editing, and control of translation by determining the turnover of RNA. T2 family RNases have been implicated in nutrition, phosphate remobilization, self-incompatibility, senescence, and defense against pathogens. They are important analytical enzymes and have been exploited for the structural determination of RNAs. Although considerable information is available on RNase A and T1 family RNases, less information is available on RNases from T2 family except RNase Rh from Rhizopus niveus and RNase LE from tomato. However, during the last few years, the primary structure, active site nature based on sequence homology, and probable mechanism of action have been postulated for some of these enzymes. RNases of T2 family, their occurrence, purification, characteristics, biological role, and applications have been reviewed. PMID- 12109773 TI - Icatibant blocks but does not reverse ACE inhibitor renal effect in Goldblatt rabbit. AB - This study examined the proposition that kinins are involved in the renal hemodynamic effect of an ACE inhibitor in Goldblatt (GB) hypertension. The effects of the ACE inhibitor enalaprilat were compared in two groups of anesthetized two-kidney one-clip GB rabbits. One group (n = 11) was given enalaprilat (10 mg/kg, i.v.) while a second group (n = 10) received the kinin B2 receptor antagonist, icatibant (2.5-5 microg/kg/min, i.v.) prior to enalaprilat. Enalaprilat caused a 40% rise in renal blood flow and 11 mm Hg decrease in blood pressure in the untreated, but no significant renal effect in the icatibant treated group. Blood pressure was reduced to the same degree in both groups. The results indicate that kinins play a major role in the renal hemodynamic, but not the blood pressure effect of ACE inhibition in the GB rabbit. PMID- 12109774 TI - Antioxidant therapy potentiates antihypertensive action of insulin in diabetic rats. AB - BACKGROUND: Poorly controlled longstanding diabetes frequently results in sustained hypertension (HTN) which plays a major role in the pathogenesis of diabetic nephropathy. In addition, hyperglycemia, per se, causes a reversible rise in blood pressure (BP) and increases production of reactive oxygen species (ROS). Increased ROS activity may raise BP by promoting inactivation of nitric oxide (NO) and/or nonenzymatic generation of vasoconstrictive prostaglandins from peroxidation of arachidonic acid. Therefore, we hypothesized that antioxidant therapy may enhance the BP-lowering effect of glycemia control with insulin replacement in diabetes. METHODS: Male Sprague-Dawley rats were rendered diabetic by streptozotocin administration and randomized to untreated, antioxidant-treated (vitamin E-fortified food, tocopherol 5000 U/kg chow and vitamin C-fortified H2O, 1000 mg/L), insulin-treated and insulin plus antioxidant-treated groups. Normal rats fed a regular diet or antioxidant-fortified diet served as controls and monitored for 4 weeks. RESULTS: The diabetic animals showed marked hyperglycemia, HTN, proteinuria, depressed tissue glutathione level and elevated plasma lipid peroxidation product, malondialdehyde (MDA) denoting increased ROS activity. Insulin therapy alone resulted in significant, but incomplete reduction in BP and plasma MDA but not proteinuria. Antioxidant therapy alone had no effect on the measured parameters in either the diabetic or control animals. However, combined insulin and antioxidant therapies normalized BP, plasma MDA and urinary protein in the diabetic animals. As expected, uncontrolled diabetes resulted in glomerular hyperfiltration which was partially reversed by insulin therapy, but was unaffected by antioxidant therapy. CONCLUSION: Uncontrolled hyperglycemia in the early phase of diabetes was associated with elevated plasma MDA, HTN and proteinuria. Insulin therapy alone resulted in significant but incomplete reduction of plasma MDA and BP. Antioxidant therapy which was ineffective when given alone, normalized plasma MDA, BP and reduced urinary protein excretion when combined with insulin treatment. PMID- 12109775 TI - Association of the G protein beta3 subunit T allele with insulin resistance in essential hypertension. AB - A polymorphism (C825T) in the gene encoding the G protein beta3 subunit (GNB3) has recently been associated with hypertension and obesity in several populations. The aim of the study was to analyse the relationship between this polymorphism and insulin sensitivity, an hypothesised unifying factor for hypertension and obesity. One hundred thirty unrelated patients with essential hypertension, 70 female and 60 male, aged 58 +/- 1 years with systolic blood pressure of 173 +/- 2 mm Hg and diastolic blood pressure of 105 +/- 1 mm Hg, were genotyped for the GNB3 polymorphism by PCR and restriction digestion with BseDI, and classified in two groups according to the genotypes CC and CT + TT. Body mass index (BMI) was significantly higher in patients with the T allele as compared with patients without the T allele (29.3 +/- 0.4 vs. 26.7 +/- 0.6 kg/m2, p<0.001). On the contrary, there were no differences in the level of systolic or diastolic blood pressure among the genotypes. Insulin sensitivity was measured in a subgroup of 35 patients by means of an euglycemic hyperinsulinemic clamp test. In this subgroup, patients with the T allele displayed lower insulin sensitivity index (1.6 +/- 0.3 vs. 2.7 +/- 0.3 mg/kg/min, p = 0.022), higher fasting serum insulin (121 +/- 16 vs. 77 +/- 11 pmol/L, p = 0.032), higher serum glucose 120 min after 75 g load (9.8 +/- 1.2 vs. 7.0 +/- 0.5 mmol/L, p = 0.038), and higher glycosilated haemoglobin (5.7 +/- 0.4 vs. 4.7 +/- 0.2%; p = 0.042) as compared with patients without the T allele. A regression analysis showed that the association between the T allele and insulin sensitivity was independent of BMI (beta coefficient -0.386, p = 0.022). These results suggest a relationship between the 825T allele of GNB3 and insulin resistance in the essential hypertensive patients studied, which seems to be independent of BMI. PMID- 12109776 TI - Zn deficiency aggravates hypertension in spontaneously hypertensive rats: possible role of Cu/Zn-superoxide dismutase. AB - Using spontaneously hypertensive rats (SHR) fed a standard or a Zn-deficient diet for 4 weeks, we examined whether Zn deficiency affects systemic blood pressure (BP) levels in a genetically hypertensive state through a fall in the activity of Cu/Zn-superoxide dismutase (SOD). SHR fed a Zn-deficient diet had a progressive increase in systolic BP during the dietary conditioning. Consequently, SHR fed a Zn-deficient diet exhibited significantly increased levels of systolic BP by 2 weeks after the start of dietary treatment when compared with SHR fed a standard diet. Similarly, levels of basal mean arterial pressure (MAP) observed at the end of dietary treatment were SHR fed a Zn-deficient diet > SHR fed a standard diet. Administration of the nitric oxide synthase (NOS) inhibitor, L-NAME, caused an increase in MAP levels in the two groups of rats, demonstrating the involvement of the vasodilator, nitric oxide (NO), in the regulation of systemic BP in a genetically hypertensive state. The expression of endothelial (e) NOS mRNA and protein in the thoracic aorta paralleled basal MAP levels in the two groups of rats, suggesting the counter-regulation of eNOS against the developed hypertensive state in SHR fed a Zn-deficient diet. On the other hand, administration of the superoxide scavenger, tempol (a SOD mimetic compound), led to a decrease in MAP levels in the two groups of rats, indicating the participation of the oxygen free radical, superoxide, in an increase in systemic BP in a genetically hypertensive state. As reported recently, the mechanism involved is due likely to a decrease in the action of the vasodilator, NO, based on the formation of peroxynitrite coming from the non-enzymatic reaction of superoxide and NO. In addition, tempol treatment completely restored MAP levels in SHR fed a Zn-deficient diet to levels comparable to those observed in SHR fed a standard diet, indicating that a further increase in systemic BP levels seen in SHR fed a Zn-deficient vs. a standard diet is presumably brought by a reduction in the action of the vasodilator, NO, resulting from an increase in the action of superoxide. The activity of the superoxide scavenger, Cu/Zn-SOD, in the thoracic aorta was significantly decreased in SHR fed a Zn-deficient diet relative to SHR fed a standard diet. It appears that a decrease in the activity of Cu/Zn-SOD observed in the thoracic aorta of SHR fed a Zn-deficient diet at least in part plays a role in an increase in the action of superoxide in this model. Thus, Zn deficiency may be a factor to develop genetic hypertension presumably through the oxidative stress caused by superoxide. PMID- 12109777 TI - Nephrin expression in the post-natal developing kidney in normotensive and hypertensive rats. AB - Nephrin is a slit diaphragm protein and its expression in the developing kidney is largely unknown. In this study, we explored the expression of nephrin in the developmental kidney in spontaneously hypertensive (SHR) and in Wistar-Kyoto (WKY) rats at different time points, from day 5 after birth to adulthood. Real time RT-PCR, in situ hybridization and immunohistochemistry were used to assess and quantify gene and protein expression of nephrin in the kidney. SHR had hypertension at week 10 and albuminuria at week 20. Nephrin expression in both SHR and WKY increased from day 5 to adulthood. Furthermore, both gene and protein expression of nephrin were significantly lower in SHR after birth when compared to WKY at the same age. These findings suggest that both in normotensive and hypertensive rats, nephrin expression increased from birth to the adult age and that down-regulation of nephrin in SHR evident from the early developmental kidney to adulthood may contribute to the development of albuminuria in adult SHR. PMID- 12109778 TI - Intestinal dopaminergic activity in obese and lean Zucker rats: response to high salt intake. AB - The present study examined intestinal dopaminergic activity and its response to high salt (HS, 1% NaCl over a period of 24 hours) intake in obese (OZR) and lean Zucker rats (LZR). The basal Na+,K+-ATPase activity (nmol Pi/mg protein/min) in the jejunum of OZR was higher than in LZR on normal salt (NS) (OZR-NS = 111.3 +/- 6.0 vs. LZR-NS = 88.0 +/- 8.3). With the increase in salt intake, the basal Na+,K+-ATPase activity significantly increased in both animals (OZR-HS = 145.9 +/ 11.8; LZR-HS = 108.8 +/- 6.7). SKF 38393 (10 nM), a specific D1-like dopamine receptor agonist, inhibited the jejunal Na+,K+-ATPase activity in OZR on HS intake, but failed to inhibit enzyme activity in OZR on NS intake and LZR on NS and HS intakes. The aromatic L-amino acid decarboxylase (AADC) activity in OZR was lower than in LZR on NS intake. The HS intake increased AADC activity in OZR, but not in LZR. During the NS intake the jejunal monoamine oxidase (MAO) activity in OZR was similar to that in LZR. The HS intake significantly decreased MAO activity in both OZR and LZR. The jejunal COMT activity in OZR was higher than in LZR on NS intake. The HS intake reduced COMT activity in OZR but not LZR. It is concluded that inhibition of jejunal Na+,K+-ATPase activity through D1 dopamine receptors is dependent on salt intake in OZR, whereas in LZR, the enzyme failed to respond to the activation of D1 dopamine receptors irrespective of their salt intake. PMID- 12109780 TI - Human papillomavirus update with a particular focus on cervical disease. AB - Human papillomavirus (HPV) is a common viral infection of squamous epithelial tissues, but its importance has only recently been recognised by the medical community. HPVs are now realised to consist of many genotypes and are associated with a diverse spectrum of clinical manifestations. Within the genital tract, some diseases have been recognised since antiquity; for example, genital warts which are caused by HPV types distinct from those causing genital cancer. However, others (such as cervical cancer), although recognised centuries ago as linked to sexual activity, have only been associated with oncogenic HPVs relatively recently, with the tools of molecular biology. We now understand that genital HPV infections are the most common sexually transmitted viral infections, are largely transient, asymptomatic and of no consequence. This virus manifests as more than just benign warts. Chronic carriage of with oncogenic genotypes (over years and in a minority of patients), together with other cofactors (host and/or exogenous) in complex pathways not totally understood, result in severe dysplasia or, ultimately, carcinogenesis. As it takes time for precursor lesions to develop and there are effective screening programmes for their detection and treatment, HPV-related neoplastic disease of the cervix is largely a preventable reproductive health issue of women. Yet, on a global scale, cervical cancer is the second most common cancer of women, with the majority of cases occurring in developing countries. Although HPV is noncultivatable by traditional diagnostic virological methods, successfully applied molecular biology techniques have underpinned development of vaccines which are now in phase II/III clinical trials. Successful vaccination ultimately has the greatest potential to impact upon the global burden of disease from genital HPV infection. However, the outcome from reduction in incidence of dysplasia and neoplasia will take years to eventuate; consequently, various cervical cancer prevention strategies still need to be endorsed and maintained in the meantime. PMID- 12109779 TI - Increased ability of erythrocytes to aggregate in spontaneously hypertensive rats. AB - The development of hypertension is accompanied by changes in the rheological properties of blood, particularly by increased red blood cell (RBC) aggregation leading to further pathological complications. However, it is not clear whether these changes in aggregation are caused only by increased concentrations of plasma adhesion proteins or if alterations in RBC membranes are also involved. The aim of the present study was to determine if RBC aggregability is altered during hypertension and if these changes correlate with alterations in RBC membrane protein concentrations. Aggregability changes were evaluated by comparing fibrinogen (Fb)-induced aggregation of RBCs from spontaneously hypertensive rats (SHR) with RBCs from age matched normotensive Wistar Kyoto (WKY) rats. ANOVA showed a significant increase in dose-dependent Fb-induced aggregation of RBCs in the SHR group. Analysis of Coomassie-stained gels of RBC membrane proteins using SDS-PAGE showed a significant increase in the amount of a protein at 110 kD in the SHR group. These results show that increased RBC aggregability is accompanied by alterations in RBC membrane protein composition during hypertension development. PMID- 12109781 TI - Banking for the future: an Australian experience in brain banking. AB - The New South Wales (NSW) Tissue Resource Centre (TRC) has been set up to provide Australian and international researchers with fixed and frozen brain tissue from cases that are well characterised, both clinically and pathologically, for projects related to neuropsychiatric and alcohol-related disorders. A daily review of the Department of Forensic Medicine provides initial information regarding a potential collection. If the case adheres to the strict inclusion criteria, the pathologist performing the postmortem examination is approached regarding retention of the brain tissue. The next of kin of the deceased is then contacted requesting permission to retain the brain for medical research. Cases are also obtained through donor programmes, where donors are assessed and consent to donate their brain during life. Once the brain is removed at autopsy, the brain is photographed, weighed and the volume determined, the brainstem and cerebellum are removed. The two hemispheres are divided, one hemisphere is fresh frozen and one fixed (randomised). Prior to freezing, the hemisphere is sliced into 1-cm coronal slices and a set of critical area blocks is taken. All frozen tissues are kept bagged at -80 degrees C. The other hemisphere is fixed in 15% buffered formalin for 2 weeks, embedded in agar and sliced at 3-mm intervals in the coronal plane. Tissue blocks from these slices are used for neuropathological analysis to exclude any other pathology. The TRC currently has 230 cases of both fixed and frozen material that has proven useful in a range of techniques in many research projects. These techniques include quantitative analyses of brain regions using neuropathological, neurochemical, neuropharmacological and gene expression assays. PMID- 12109782 TI - Vimentin expression does not assist in predicting survival in ductal carcinoma of the breast. AB - AIMS: To establish the value of vimentin expression in predicting survival in patients with breast cancer. METHODS: Five-year follow-up data were obtained for 68 patients with ductal carcinoma (NOS) of the breast in whom vimentin expression had been studied in fresh frozen and formalin-fixed, paraffin-embedded tissue. The predictive value on survival of tumour size, growth fraction (as assessed using the Ki67 monoclonal antibody), oestrogen receptor status and Bloom and Richardson grade of the primary tumour, and the presence or absence of lymph node metastases in axillary samples, were also studied. RESULTS: Twenty-two patients died of their disease within 5 years of diagnosis. Vimentin expression either on frozen or paraffin sections did not provide a statistically significant prediction of survival. On univariate analysis tumour grade, size and the presence of lymph node metastases provided prognostic information. Only lymph node status was of independent prognostic importance on multivariate analysis. CONCLUSIONS: Whilst these results confirm the value of established prognostic factors, they do not support the use of vimentin expression in either fresh or fixed tissue for the prediction of survival in ductal carcinoma (NOS) of the breast. PMID- 12109783 TI - Expression of E-cadherin in prostate cancer in formalin-fixed, paraffin-embedded tissues: correlation with pathological features. AB - AIMS: E-cadherin has been studied recently as a potential marker for tumour progression. The present study aimed to assess the expression of E-cadherin in formalin-fixed and paraffin-embedded radical prostatectomy specimens and to compare its expression with the pathological stage and Gleason score. METHODS: This study comprised a total of 58 men who were selected on the basis of the negative surgical margins of surgical specimens from radical retropubic prostatectomies and concomitant pelvic lymph node dissections. Indirect immunoperoxidase staining was performed as described previously using HECD-1 monoclonal antibody with the retrieval of antigen by treatment of the paraffin embedded tissue with microwaves in citrate buffer. RESULTS: Aberrant staining patterns of E-cadherin were observed in 18 (64%) and in 25 (83%) of cases with pathological stages pT2 and pT3a, respectively (P>0.05). Immunohistochemical examination also revealed aberrant staining patterns of E-cadherin in 16 (89%) of specimens with Gleason score > or =7 and in 27 (68%) of specimens with Gleason score <7 (P>0.05). Biochemical recurrence was identified in three (5%) patients and immunohistochemical examination of their specimens revealed aberrant staining patterns of E-cadherin molecule in all of them. CONCLUSION: Our preliminary results indicate that, even though we could not demonstrate any significant correlation between E-cadherin staining pattern and tumour invasion and Gleason scores, aberrant staining patterns of E-cadherin may be a significant predictor for disease recurrence following radical prostatectomies if supported by large scale studies. PMID- 12109784 TI - Differentiated thyroid carcinomas with vascular invasion: a comparative study of follicular, Hurthle cell and papillary thyroid carcinoma. AB - AIM: Non-medullary thyroid carcinomas arise from follicular cells. The purpose of this study is to correlate clinical and pathological properties of these tumours with the rate of distant metastasis from a series of thyroid tumours excised at one institution. METHODS: A total of 311 non-medullary thyroid tumours were identified and divided into: 29 follicular carcinoma (FC), 12 Hurthle cell carcinoma (HC), 13 Hurthle cell papillary thyroid carcinoma (HPTC) with vascular invasion (VI), 32 papillary thyroid carcinoma (PTC) with VI and 225 PTC without VI. The mean follow-up was 6.5 years with a range of 1-17 years. The tumours were histologically subdivided into minimal or wide invasion for FC and HC and focal or extensive invasion for PTC and HPTC, and stratified according to status of VI. RESULTS: The rate of distant metastasis was similar for FC, malignant Hurthle cell tumours and PTC with VI, and increased with extent of invasion. VI was seen in 12% of all PTC and 0% of HPTC in this study. PTC without VI were associated with a much lower potential of distant metastasis, were smaller in size and occurred in patients of younger age than PTC with VI. In addition, there was a tendency for increased potential for distant metastases with increased tumour size and patient age for all groups of tumours in the study. Patient age and tumour size appeared to play a smaller role than that of VI in predicting distant metastasis. CONCLUSIONS: Our study suggests that the rate of distant metastasis relates to VI, patient age and tumour size, regardless of Hurthle cell, FC or PTC differentiation. PTC of large size, and in patients older than 45 years, have a high propensity for vascular invasion. PMID- 12109785 TI - Epstein-Barr virus-associated smooth muscle tumour: a distinctive mesenchymal tumour of immunocompromised individuals. AB - immunosuppressed patients are predisposed to the development of smooth muscle tumours which show near consistent association with Epstein-Barr virus (EBV). This report describes a 37-year-old patient with acquired immunodeficiency syndrome who initially presented with two masses in the liver. Image-guided core biopsy revealed a spindle cell tumour with histological and immunological features of smooth muscle neoplasm which was shown by in situ hybridisation for EBV early RNAs to be EBV-associated. The literature on this uncommon entity is critically reviewed and the differential diagnosis is also discussed. PMID- 12109786 TI - A possible mouse model for spontaneous cholangitis: serological and histological characteristics of MRL/lpr mice. AB - AIMS: MRL/Mp-lpr/lpr (MRL/lpr) mice spontaneously develop lymphadenopathy, hypergammaglobulinaemia, serum auto-antibodies, and a generalised auto-immune disease including glomerulonephritis and arthritis, and have been used as a model for the study of systemic lupus erythematosus. Recently, MRL/lpr mice were also reported as a potentially suitable animal model of primary biliary cirrhosis (PBC). The aim of this study was to determine the suitability of MRL/Mp-lpr/lpr (MRL/lpr) mice as an experimental auto-immune-mediated cholangitis model for PBC. METHODS: We investigated the serum hepatobiliary enzymes, histopathological findings, and the target antigen of antimitochondrial antibodies (AMA) in MRL/lpr mice. RESULTS: Serum levels of total bilirubin and hepatobiliary enzymes including alanine aminotransferase (ALT), leucine aminopeptidase (LAP), and gamma glutamyl transpeptidase (G-GTP) in older-aged (over 20 weeks old) MRL/lpr or MRL/Mp-+/+ (MRL/+) mice were not significantly higher than those in younger (8-12 weeks old) MRL/lpr, MRL/+, or older-aged control mice (C3H/HeJ and BALB/C mice). Histopathologically, 24 of 47 (51%) older-aged MRL/lpr mice showed evidence of cholangitis, compared with two of 20 (10%) younger MRL/lpr mice. Especially, epithelioid granuloma and/or bile duct loss were seen in 11 out of 47 (23%) older aged MRL/lpr mice, whereas such findings were seen in only one of 20 (5%) younger MRL/lpr mice. None of the MRL/+, C3H/HeJ, and BALB/C mice developed cholangitis. The target antigens of AMA were not pyruvate dehydrogenase complex but 2 oxoglutarate dehydrogenase complex and/or branched-chain oxo-acid dehydrogenase complex as confirmed by immunoblotting. There was no significant correlation between the presence of AMA and severity of histological lesions in older-aged MRL/lpr mice, and there were no significant differences in these biochemical data, the proportion of mice with portal inflammation, cholangitis and AMA between male and female MRL/lpr mice. CONCLUSION: Although several clinical features were incompatible with PBC, the serological and histopathological features of MRL/lpr mice indicate that these mice can be used as an experimental immune-mediated cholangitis model for PBC. PMID- 12109787 TI - An autopsy approach to bee sting-related deaths. AB - Although severe reactions to the sting of the common honey bee (Apis mellifera) are a common problem in Australia, reported deaths are uncommon, with the estimated mortality varying from one to four persons each year. The following study presents the postmortem findings in three cases of bee sting fatality, including one in which no observable sting was found. An autopsy approach to such cases is detailed. Overreporting of bee sting-related deaths may occur due to the inclusion of deaths unrelated to a reaction to bee venom, while under-reporting may be due to unexplained deaths where a history of a bee sting is not available or apparent at autopsy. A classification of bee sting-related deaths is proposed, which would allow more accurate reporting of bee sting-related fatalies. A serum tryptase and specific IgE to bee venom on serum obtained at autopsy can assist in confirming anaphylactic reaction to bee venom as the cause of death, particularly in the absence of observable stings. Although there are limitations to the usefulness of serum tryptase tests in the postmortem situation, it may still be useful to confirm suspected anaphylaxis in autopsy cases with an undetermined cause of death. PMID- 12109788 TI - Neutrophil dysplasia characterised by a pseudo-Pelger-Huet anomaly occurring with the use of mycophenolate mofetil and ganciclovir following renal transplantation: a report of five cases. AB - AIM: The pseudo-Pelger-Huet (PH) anomaly has been associated with a variety of primary haematological disorders, infections and drugs. Recently, the development of dysgranulopoiesis characterised by a pseudo-PH anomaly has been reported in two patients with the use of mycophenolate mofetil (MMF) in the setting of heart and/or lung transplantation. We present a further five cases of MMF-related dysgranulopoiesis characterised by a pseudo-PH anomaly occurring after renal transplantation. METHODS: All patients were receiving standard immunosuppression protocols for renal transplantation, including a combination of MMF, steroids and either cyclosporin or tacrolimus. Oral ganciclovir was also used for cytomegalovirus prophylaxis in each case. RESULTS: Development of dysplastic granulopoiesis occurred a median of 96 days (range 66-196 days) after transplantation. Moderate or severe neutropaenia (<1.0 x 10(9)/l) developed in three cases, and appeared to be directly correlated with the percentage of circulating neutrophils present with dysplastic morphology. Resolution of dysgranulopoiesis occurred in all cases only after dose reduction and/ or cessation of both MMF and ganciclovir. CONCLUSIONS: In our series, the observed dysplastic granulopoiesis appeared related to the combination of MMF and ganciclovir, rather than MMF alone. Further study is required to determine the exact incidence and pathogenesis of this pattern of bone marrow toxicity. PMID- 12109789 TI - Amiodarone-induced bone marrow granulomas. AB - We describe a case of a 76-year-old man with indolent myeloma who developed extensive bone marrow granulomas whilst on amiodarone therapy. The granulomas resolved on cessation of amiodarone. PMID- 12109790 TI - The detection of Chlamydia pneumoniae in atherosclerotic plaques of Australian subjects. AB - AIM: To determine whether the common respiratory pathogen, Chlamydia pneumoniae, was associated with atherosclerotic plaques in Australian subjects. METHODS: A total of 29 coronary atherosclerotic lesions and 18 normal coronary arterial samples were tested for the presence of C. pneumoniae by PCR and immunofluorescence methods. RESULTS: Chlamydia pneumoniae was detected in 15 of the atheromatous lesions as well as in three of the normal tissues; the immunofluorescence assay was more sensitive (P=0.028) than PCR (P=0.26). CONCLUSIONS: These findings contradict previous Australian studies which did not detect C. pneumoniae in atherosclerotic plaques, thereby discounting the speculation that its absence was likely due to geographical variation. The detection of the bacterium in some of the normal tissues suggests that C. pneumoniae infection might be an initial trigger of atherosclerotic development. PMID- 12109792 TI - Web-based self-assessments in pathology with Questionmark Perception. AB - Formative assessment with appropriate feedback is an effective method of promoting learning. Software tools are now available to facilitate the delivery of formative self-assessments via the World Wide Web. For the past 5 years, we have offered on-line formative self-assessments in pathology to undergraduate medical students. These assessments, which have been authored using Questionmark Perception, have proved particularly popular and have received high ratings in course evaluation surveys. Furthermore, they appear to have been effective in promoting student learning, with evidence that students made a genuine attempt to answer the questions in the assessment and that they learnt from doing so. The potential of on-line formative self-assessments for postgraduate training and continuing education in pathology so far remains untapped. PMID- 12109793 TI - Test and Teach. A hyperechoic intracranial lesion in a foetus. Large congenital cerebral primitive neuroectodermal tumour (PNET). PMID- 12109791 TI - Chromosome 2p, 3p, 5q and 18q status in sporadic gastric cancer. AB - AIM: The genetic make-up of gastric cancers in low-risk population groups from South Africa is largely unknown. The purpose of this study was to ascertain the incidence of microsatellite instability and loss of heterozygosity in this population. METHODS: Thirty-seven gastrectomy specimens for sporadic gastric cancer were analysed for the following clinicopathological parameters: age, gender, race, histopathological type, size of tumour, lymph node status and the presence/absence of Helicobacter pylori. DNA was then extracted from paraffin embedded tissue and seven microsatellite markers in 2p, 3p, 5q and 18q loci were examined using automated DNA fluorescent technology. RESULTS: Only eight cases showed microsatellite instability (MSI) for one marker and were thus categorised as MSI-low. In the 3p region, loss of heterozygosity (LOH) was detected in 21.7 38.3% of informative cases, whilst in the 18q region CLOH ranged from 25 to 38.4%. LOH was not seen in the 2p locus and only one case showed LOH in the 5q region. When the molecular changes were compared with clinicopathological parameters, a statistically significant relationship did not emerge with any single parameter. CONCLUSIONS: This study shows that sporadic gastric cancer from a low-risk population in South Africa is MSI-low or MSI-stable, and that LOH in the 3p and 18q regions is similar to that seen in other low-risk populations from different geographical regions. PMID- 12109794 TI - Early purulent meningococcal pericarditis due to an unusual strain of Neisseria meningitidis. PMID- 12109795 TI - Soft tissue amyloidoma. PMID- 12109796 TI - Miliary blastomycosis developing in an immunocompromised host with chronic lymphocytic leukaemia. PMID- 12109797 TI - Diffuse glomerulonephritis associated with rifampicin treatment for tuberculosis. PMID- 12109798 TI - Surgically correctable hypertension. PMID- 12109799 TI - Irinotecan, cisplatin, and radiation in esophageal cancer. AB - The limited effectiveness of currently available chemotherapy in the treatment of advanced esophageal cancer, and the poor survival achieved in locally advanced disease with combined chemoradiotherapy with or without surgery, have prompted the evaluation of new agents. Irinotecan (CPT-11, Camptosar) has promising single agent activity in gastrointestinal cancers. In phase II evaluation of weekly irinotecan plus cisplatin, response rates have exceeded 30% in esophageal and gastric cancers. Irinotecan is an active radiosensitizer in preclinical studies and clinical trials in lung cancer. We performed a phase I trial of weekly irinotecan, cisplatin, and concurrent radiotherapy in locally advanced esophageal cancer. Induction chemotherapy with irinotecan and cisplatin was given prior to radiotherapy, over 6 weeks, cycled on a 2-week-on, 1-week-off schedule to relieve dysphagia. Radiotherapy was given subsequently in 180-cGy daily fractions to a total dose of 5,040 cGy. Doses of chemotherapy, when given with concurrent radiotherapy, were cisplatin at 30 mg/m2 followed by irinotecan at escalated doses (40, 50, 65, and 80 mg/m2), on days 1, 8, 22, and 29. Among 18 patients entered in the trial, minimal toxicity has been observed, with no grade 3/4 esophagitis or diarrhea. Hematologic toxicity has been minimal. Dose-limiting toxicity (ie, requiring more than a 2-week delay in radiotherapy) has been seen in one of three patients at the 80-mg/M2 irinotecan dose level, and accrual continues at this dose level. Among 13 evaluable patients, five complete responses have been seen (38%), including three pathologic complete responses in 10 patients undergoing surgery (30%). Asymptomatic pulmonary emboli were noted on the posttreatment computed tomography scan in 3 of 15 patients, prompting the addition of warfarin sodium (Coumadin) prophylaxis on protocol. Full doses of weekly irinotecan (65 mg/ m2) and cisplatin (30 mg/m2) can be combined safely with concurrent radiotherapy in patients with locally advanced esophageal cancer. PMID- 12109800 TI - Irinotecan/cisplatin in advanced, treated gastric or gastroesophageal junction carcinoma. AB - We conducted a phase II study to assess the response rate and toxicity profile of the irinotecan (CPT-11, Camptosar) plus cisplatin combination administered weekly to patients with at least one previous chemotherapy for advanced adenocarcinoma of the stomach or gastroesophageal junction. Patients with histologic proof of adenocarcinoma of the stomach orgastroesophageal junction with adequate liver, kidney, and bone marrow functions were treated with 50 mg/m2 of irinotecan plus 30 mg/m2 of cisplatin, both administered intravenously 1 day a week for 4 consecutive weeks, followed by a 2-week recovery period. Response rate, time to disease progression, survival, and toxic effects were analyzed. Of 32 registered patients, 29 were assessable. Nine patients (31%) achieved a partial response. Median time to disease progression was 7 weeks (range: 5-48+ weeks) and median survival time was 5 months (range: 2.5-31 months). There were no treatment related deaths. Major toxic effects included diarrhea, neutropenia, and fatigue. Of 260 doses administered in 65 6-week courses, 46 doses were either delayed or missed. Most delayed or missed doses occurred in the third or fourth week of the cycle. We concluded that the combination of irinotecan and cisplatin is an active second-line therapy for patients with advanced gastric or gastroesophageal adenocarcinoma in whom one previous chemotherapy has failed. Modifications in doses and schedule are warranted to increase the tolerability of the regimen. Phase III trials are necessary to establish the clinical utility of irinotecan/cisplatin in these patient populations. PMID- 12109801 TI - Irinotecan and gemcitabine in patients with solid tumors: phase I trial. AB - Using a day 1 and 8, every-3-week schedule, our purpose was to determine the maximum tolerated dose of irinotecan (CPT-11, Camptosar) that can be administered immediately after gemcitabine (Gemzar) at a dose of 1,000 mg/m2 IV. In this phase I trial, the maximum tolerated dose was defined as the dose level immediately below the level in which two of the first three patients in any cohort, or at least two of six patients in any expanded cohort, experienced dose-limiting toxicity. Dose-limiting toxicity pertained only to toxicity during the first cycle of treatment. Escalation of irinotecan was planned in groups of three patients, with three additional patients added at the first indication of dose limiting toxicity. A total of 19 patients have been enrolled. Grade 4 diarrhea was the dose-limiting toxicity at the irinotecan dose of 115 mg/m2. Hematologic toxicity was not dose limiting. Three patients required canceling of the day 8 dose due to grade 3 myelosuppression. Three patients, two with pancreatic cancer and one with metastatic carcinoma of unknown primary, had a partial response. The maximum tolerated dose of irinotecan in this combination was 100 mg/m2/dose. The dose-limiting toxicity was diarrhea. The maximum tolerated dose is the recommended starting dose for phase II studies. PMID- 12109802 TI - Irinotecan/gemcitabine followed by twice-weekly gemcitabine/radiation in locally advanced pancreatic cancer. AB - Early clinical studies combining irinotecan (CPT-11, Camptosar) and gemcitabine (Gemzar) have yielded encouraging results. Gemcitabine administered via a twice weekly schedule results in an enhanced radiation-sensitizing effect. This multi institution phase II trial of induction irinotecan/gemcitabine followed by twice weekly gemcitabine and upper abdominal radiation has been initiated to determine the activity of this regimen in patients with unresectable pancreatic cancer. Patients received two cycles of induction irinotecan (100 mg/ m2 IV) and gemcitabine (1,000 mg/m2 IV) on days 1 and 8 of each 3-week cycle. Following the induction therapy, patients without disease progression received twice-weekly gemcitabine at 40 mg/m2 and radiation. Nine patients have been enrolled in the study to date. Median patient age was 71 years (range: 65-85 years). The major toxicity observed thus far was grade 3/4 neutropenia. Grade 3/4 nonhematologic toxicity was rarely observed and included dehydration (12%) and diarrhea (12%), which were likely related to the irinotecan. No treatment-related deaths have occurred. These preliminary data suggest that this regimen is well tolerated. Although the data are limited, tumor progression during the induction chemotherapy has not been observed thus far (radiographically or biochemically [CA-19-9]). PMID- 12109803 TI - Irinotecan in epithelial ovarian cancer. AB - Ovarian cancer, the second most common gynecologic malignancy, accounts for approximately 14,000 deaths annually in the United States. Disease relapse after primary treatment, which consists mainly of surgery followed by platinum-based therapy, occurs in more than 60% of ovarian cancer patients overall, and in more than 80% of those diagnosed initially with advanced-stage disease. Responses to second-line chemotherapy agents range from 15% to 25%, and are usually partial responses of short duration. Irinotecan (CPT-11, Camptosar), a camptothecin derivative that inhibits topoisomerase I, is undergoing assessment for the treatment of ovarian cancer. Preclinical studies demonstrated antitumor activity in ovarian cancer. Early phase I or II trials showed response rates of 20% to 25% in patients with recurrent or refractory disease, with some responses noted in the relatively chemoresistant mucinous and clear cell tumors. Irinotecan has also been studied in combination regimens, most commonly irinotecan plus cisplatin. In a phase II trial of irinotecan plus cisplatin in platinum-sensitive or -resistant patients, response rates were 75% and 33%, respectively. Irinotecan seems to be relatively well tolerated; dose-limiting toxicities appear to be diarrhea, nausea and vomiting, and leukopenia/neutropenia. PMID- 12109805 TI - Combined-modality therapy for rectal cancer using irinotecan. AB - Preoperative or postoperative pelvic radiation plus concurrent fluorouracil-based chemotherapy is standard adjuvant treatment for patients with T3 and/or N1/2 rectal cancer. Newer chemotherapeutic regimens have been developed for the treatment of patients with metastatic disease. Irinotecan (CPT-11, Camptosar) based regimens have improved survival in patients with metastatic disease and are being actively investigated in combination with pelvic radiation therapy for patients with rectal cancer. PMID- 12109804 TI - Irinotecan for the treatment of cervical cancer. AB - Topoisomerase inhibitors have been widely studied for the treatment of refractory or recurrent cervical cancer. Various schedules have been used, with response rates ranging from 13% to 20%. The combination of cisplatin and irinotecan (CPT 11, Camptosar) is being studied in cervical cancer. Hematologic and gastrointestinal toxicities have been observed. Gastrointestinal toxicities have been particularly problematic at high irinotecan doses. Further studies of irinotecan in combination with other drugs and radiotherapy are warranted. PMID- 12109807 TI - Future directions in adjuvant therapy for rectal cancer. AB - The US National Cancer Institute Gastrointestinal Intergroup has contributed to the development of chemotherapy and radiation regimens for the treatment of stage II and III rectal cancer. The first Intergroup trial demonstrated improvement in relapse-free and overall survival for patients who received protracted venous infusion fluorouracil (5-FU) with radiation compared to those treated with bolus 5-FU. The second trial, INT 0114, demonstrated that there was no difference among four regimens, including 5-FU with or without leucovorin or levamisole (Ergamisol) vs three drugs given as concomitant therapy. High-risk (T3, N+; T4) and low-risk (T1/2, N+; T3, N0) tumor categories were identified, revealing significant differences in 5- and 7-year survival rates. Risk of local failure was also greater in the high-risk group. Pathologic assessment of lymph nodes in the surgical specimens had an important impact on these trial results. Relapse free and overall survival were significantly different among the lymph node negative patients, as determined by the number of lymph nodes examined. The National Surgical Adjuvant Breast and Bowel Project trials (R-01, R-02) have shown that the addition of radiation does not affect disease-free or overall survival but improves local recurrence rates. Future trials will explore use of newer agents, including capecitabine (Xeloda), irinotecan, and oxaliplatin. In addition, prospective evaluation of laboratory correlates, including molecular markers, will be integrated into trial design. PMID- 12109806 TI - Preoperative chemoradiation for locally advanced rectal cancer: emerging treatment strategies. AB - Over the past decade, patients with locally advanced rectal cancer at The University of Texas M. D. Anderson Cancer Center have been managed with preoperative chemoradiation. Patients achieving a complete clinical response to preoperative chemoradiation have had better pelvic tumor control, sphincterpreservation, and overall survival than those with gross residual disease. Some patients achieving a complete clinical response have even had rectal-preserving surgery (full-thickness local excision). These results emphasize the importance of maximizing tumor response. Further improvement in response and survival could be achieved by using novel chemotherapeutic agents or through tumor-selective molecular targeting strategies that enhance the effects of chemotherapy, radiotherapy, or both. Irinotecan (CPT-11, Camptosar) is a novel chemotherapy agent being evaluated clinically as a radiosensitizing agent in rectal cancer. Inhibition of several molecular targets-such as epidermal growth factor receptor, ras oncogene activation, the cyclooxygenase-2 (COX-2) enzyme, and neoangiogenesis-appears to be tumor-selective in preclinical models. COX-2 expression has been shown to enhance cytotoxic therapy in preclinical models. In vitro and in vivo studies show that selective COX-2 inhibition enhances the effects of radiotherapy as well as chemotherapy. COX-2 is also markedly upregulated in human colorectal cancer and appears to be associated with adverse patient prognosis. Thus, integration of molecular targeting, such as COX-2 selective inhibition with existing chemoradiation approaches, may provide selective tumor radiosensitization and chemosensitization, resulting in improved pelvic control, sphincter preservation, and overall survival. PMID- 12109808 TI - Effects of sodium fluoride on total serum protein levels and transaminase activity in rats. AB - Transaminase activity and serum total protein level were investigated in adult rats after oral treating with sodium fluoride at three doses, 10, 20 and 30 mg/kg daily for 90 days. After 90 days, the average total serum protein level of the rats in the treatment group decreased significantly compared with that in the control [1.9 +/- 0.1 (mean +/- S.D., n = 140) vs. 3.1 +/- 0.2] mg/dl, P< 0.05. Serum transaminase activity in the treatment group increased compared with that in the control [5.3 +/- 0.4 (mean +/- S.D., n = 140) vs. 3.2 +/- 0.3] micromol/min per ml, P < 0.05. PMID- 12109809 TI - Role of selenium in reducing hypoxia-induced oxidative stress: an in vivo study. AB - At high altitudes, the reactive oxygen species are continuously generated as a consequence of low oxygen partial pressure (hypoxia), which causes tissue damage. The body's defence system to combat the oxidative stress (e.g., anti-oxidant enzymes, free radical scavengers such as vitamin C, vitamin E, beta-carotene, reduced glutathione and minerals such as selenium, etc.) may diminish. In the present study, the antioxidant effect of selenium (Se) in reducing the hypoxia induced oxidative stress was evaluated by exposing male albino rats to hypoxic stress in a decompression chamber. Exposure to hypoxia resulted in an increase in malondialdehyde (MDA) levels in plasma and tissues and a concurrent decrease in blood glutathione (GSH), glutathione peroxidase (GPx), plasma protein and plasma selenium content when compared with controls. Haemoglobin concentration (Hb%), red blood corpuscles (RBC) and white blood corpuscles (WBC) count were also increased in the hypoxia-exposed group. Selenium supplementation to animals reversed the trend. There was a significant decrease (P < 0.001) in MDA and subsequent increase in plasma and tissue GSH levels. Similarly the blood and tissue GPx and plasma protein also increased significantly in the Se supplemented animals compared with control animals. The Hb%, RBC and WBC counts showed no significant difference between Se-fed and control rats. These results suggest that selenium may help in reducing the lipid peroxidation during hypoxia. PMID- 12109810 TI - Antidiabetic effect of a nitrosamine-free dephostatin analogue, methoxime-3,4 dephostatin, in db/db mice. AB - Et-3,4-dephostatin, a protein-tyrosine phosphatase (PTPase) inhibitor, potentiates insulin-dependent signal transduction and shows an antidiabetic effect in mice. However, it contains a nitrosamine moiety that is often mutagenic and carcinogenic. Therefore, we previously designed and synthesized methoxime-3,4 dephostatin as a nitrosamine-free analogue of dephostatin. In the present paper, we studied in situ and in vivo antidiabetic effects of this PTPase inhibitor. Methoxime-3,4-dephostatin induced 2-deoxyglucose transport by mouse 3T3-L1 adipocytes and rat L6 myocytes without insulin. It also inhibited glucagon induced glucose release from primary culture rat hepatocytes. When hepatocytes were prepared from starved rats, methoxime-3,4-dephostatin did not inhibit the release of glucose, indicating that the chemical may act on glycogenolysis. Oral administration of methoxime-3,4-dephostatin for 3-7 days inhibited the increase in the blood glucose level in type-2 diabetes model db/db mice. It also decreased food and water intakes of mice, but showed no liver or blood toxicity. PMID- 12109811 TI - Mitochondrial permeability transition can be directly monitored in living neurons. AB - Mitochondria have been suggested as key players in apoptotic cell death of neurons and many other tissues, since the release of proapoptotic molecules from mitochondria is implicated in caspase activation. As a potential release mechanism, the occurrence of a large pore opening in the inner membrane (mitochondrial permeability transition pore, PTP) has been proposed, but has not yet been observed directly in neurons. We investigated whether the calcein/Co2+ quenching technique introduced by Petronilli et al. [Biofactors 8 (1998) 263], which allows direct observation of PTP opening, can be applied to neurons. Exposure of calcein-loaded neurons to Co2+ ions resulted in the fading of diffuse cytoplasmic calcein fluorescence, with organelle-restricted fluorescent spots remaining. These spots were colocalized with mitochondrially-entrapped tetramethylrhodamineethylester (TMRE) fluorescence and corresponded to colocalization of calcein and TMRE fluorescence in digitonin-permeabilized neurons. Importantly, extensive neuronal calcium loading, which is assumed to induce PTP opening, resulted in significant fading of mitochondrial fluorescence, suggesting the occurrence of permeability transition. This fluorescence decrease could be completely prevented by the PTP blocker cyclosporin A. PMID- 12109812 TI - Thalidomide, an antiangiogenic agent with clinical activity in cancer. AB - Despite the teratogenic past of thalidomide, there is recent evidence indicating the drug's efficacy in the management of various diseases from immune disorders to cancers. The history, pharmacodynamic and pharmacokinetic properties of thalidomide in the clinic are discussed in this review article. PMID- 12109813 TI - Polyphenols: do they play a role in the prevention of human pathologies? AB - Polyphenols are the most abundant antioxidants in our diets. The main classes of polyphenols are phenolic acids (mainly caffeic acid) and flavonoids (the most abundant in the diet are flavanols (catechins plus proanthocyanidins), anthocyanins and their oxidation products), which account for one- and two thirds, respectively. Polyphenols are reducing agents, and together with other dietary reducing agents, such as vitamin C, vitamin E and carotenoids, referred to as antioxidants, protect the body's tissues against oxidative stress and associated pathologies such as cancers, coronary heart disease and inflammation. The biological properties, bioavailability, antioxidant activity, specific interactions with cell receptors and enzymes, are related to the chemical structure of polyphenols. It is, therefore, essential to know the nature of the main polyphenols ingested, their dietary origin, the amounts consumed in different diets, their bioavailability and the factors controlling their bioavailability. PMID- 12109815 TI - Significance of fluid regime and wetted area in biofilm reactors. AB - Mass transfer within microbial films was described using Monod-type biological kinetics in terms of properties of filter media and feed solution. The performance characteristics of a trickling filter were thus modeled. The model enables one to consider the effect of inlet substrate concentration and flow rate upon the removal efficiency. For this purpose a second-order partial differential equation describing the dispersion phenomena inside the liquid layer was solved under special boundary conditions and used to determine substrate flux into the biofilm. A uniform biofilm thickness was considered. The model is based on computer techniques and the numerical evaluation of the normalized biofilm mathematical model. A design procedure was also given to calculate biological filters. The numerical model was also applied to experimental data to demonstrate its validity. PMID- 12109814 TI - Tetanus in a renal transplant recipient exhibiting the presence of circulating antitetanus antibodies determined by ELISA. PMID- 12109816 TI - Enzyme kinetics and glycan structural characterization of secreted alkaline phosphatase prepared using the baculovirus expression vector system. AB - Secreted human alkaline phosphatase (SEAP, a model protein containing a single N glycan chain) was expressed in Spodoptera frugiperda Sf-9 (Sf-9) and Trichoplusia ni BTI-Tn-5B1-4 (Tn-5B1-4) insect cell lines infected with recombinant Autographa californica multiple nuclear polyhedrovirus expressing SEAP under control of the polyhedrin promoter. SDS-PAGE showed that both systems expressed fairly pure rSEAP products. The rSEAP expression level was 7.0 U/mL in Tn-5B1-4, higher than the 4.1 U/mL produced by Sf-9. Kinetic analysis showed that Vmax and Km of human placental SEAP were approx 10-fold higher than that of rSEAP, whereas the Vmax and Km of rSEAP prepared using both insect cell lines were comparable. To characterize the recombinant SEAP (rSEAP) glycosylation, the purified rSEAP was digested with PNGase F to release the N-glycan chains. Glycan analysis showed the presence of oligomannose-type N-linked glycans (i.e., Man(2-8)GlcNAc2 and FucMan(3 or 4)GlcNAc2) in rSEAP from Sf9 and Tn-5B1-4 cell lines. The proportions of these oligosaccharide structures were different in the two cell lines. Man4GlcNAc2 and FucMan4GlcNAc2 were the major rSEAP N-glycans produced in Sf-9 cells, while Man2GlcNAc2 was the major rSEAP N-glycan produced in Tn-5B1-4 cells. PMID- 12109817 TI - Scale-up of microbubble dispersion generator for aerobic fermentation. AB - A laboratory-scale microbubble dispersion (MBD) generator was shown to improve oxygen transfer to aerobic microorganisms when coupled to the conventional air sparger. However, the process was not demonstrated on a large scale to prove its practical application. We investigated the scale-up of a spinning-disk MBD generator for the aerobic fermentation of Saccharomyces cerevisiae (baker's yeast). A 1-L spinning-disk MBD generator was used to supply air for 1- and 50-L working volume fermentation of baker's yeast. For the two levels investigated, the MBD generator maintained an adequate supply of surfactant-stabilized air microbubbles to the microorganisms at a relatively low agitation rate (150 rpm). There was a significant improvement in oxygen transfer to the microorganism relative to the conventional sparger. The volumetric mass transfer coefficient, kLa, for the MBD system at 150 rpm was 765 h(-1) compared to 937 h(-1) for the conventional sparger at 500 rpm. It is plausible to surmise that fermentation using larger working volumes may further improve the kLa values and the dissolved oxygen (DO) levels because of longer hold-up times and, consequently, improve cell growth. There was no statistically significant difference between the cell mass yield on substrate (0.43 g/g) under the MBD regime at an agitation rate of 150 rpm and that achieved for the conventional air-sparged system (0.53 g/g) at an agitation rate of 500 rpm. The total power consumption per unit volume of broth in the 50-L conventional air-sparged system was threefold that for the MBD unit for a similar product yield. Practical application of the MBD technology can be expected to reduce power consumption and therefore operating costs for aerobic fermentation. PMID- 12109818 TI - Influence of Tween 80 on lipid metabolism of an Aspergillus niger strain. AB - Addition of 0.1% of nonionic surface-active Tween 80 to a medium optimized for pectolytic enzyme production of Aspergillus niger increased the amount of enzymes excreted by 70%. In the presence of Tween 80 the amount of sterol esters and triacylglycerols was increased. During the course of cultivation the amounts of precursors for ergosterol biosynthesis diminished with an increase of ergosterol. A. niger incorporated cholesterol from the medium, partly converting it to cholesterol esters. Sterol esters were formed only with selected fatty acids. Oleic acid, the hydrophobic part of Tween 80, was mainly incorporated in phospholipids and glycolipids. PMID- 12109820 TI - Dendritic cells for the immunotherapy of cancer. PMID- 12109819 TI - pH-sensitive chitosan films for baker's yeast immobilization. AB - Dried baker's yeast cells were immobilized on a chitosan film, which is a natural polymer. Prepared chitosan films were treated with glutaraldehyde to facilitate the immobilization of the cells. The effects of the amount of glutaraldehyde, incubation time, pH, and temperature on immobilization were investigated. The amount of glutaraldehyde was chosen to be 0.01% (weight). The highest amount of yeast immobilization was obtained with 5 h incubation. It was determined that optimum temperature for immobilization is 25 degrees C, and the optimum pH for immobilization is 6. Immobilized cells were allowed to stand for 3 d in distilled water and buffer solution (pH 6) to investigate the desorption, but no desorption was found. The maximum immobilization capacities were found to be 90 microg protein cm(-2) film in optimum conditions. PMID- 12109821 TI - The use of vaccines in the prevention and treatment of cervical cancer. AB - The close association between high risk HPV infection and cervical carcinoma has provided the impetus for the development of prophylactic and therapeutic vaccination schedules. An effective prophylactic vaccine would obviate the need for population-based cervical screening programmes, while therapeutic vaccination might provide an effective adjunct to or replacement for conventional treatment for benign and malignant cervical disease. While the challenges associated with the design and implemention of immunotherapies are numerous, optimism remains high and it is expected that the next few decades will witness a revolutionary change in the way we treat cervical cancer and its premalignant lesions. A papillomavirus vaccine that prevented HPV infection on the one hand and acted against established disease on the other, would have a profound impact on one of the major cancers affecting women globally. PMID- 12109822 TI - Chemoradiation: the new gold standard for non-surgical treatment of cervical cancer. PMID- 12109823 TI - Concurrent cisplatin-based chemotherapy plus radiotherapy for cervical cancer--a meta-analysis. AB - PURPOSE: To evaluate the role of concurrent cisplatin plus radiotherapy in the treatment of cervical cancer. METHODS: A systematic review of randomized trials of cisplatin administered concurrently with external beam radiotherapy versus radiotherapy without cisplatin for cervical cancer was combined with a meta analysis of results abstracted from published reports of the trials. RESULTS: Pooled survival rates from eight randomized trials that evaluated the role of cisplatin, alone or in combination with other chemotherapy agents, administered concurrently with external beam radiotherapy to patients with cervical cancer demonstrated a statistically significant effect in favour of cisplatin-based chemotherapy plus radiotherapy compared with radiotherapy without cisplatin (relative risk [RR] of death, 0.74; 95% confidence interval [CI], 0.64 to 0.86). The pooled RR of death among the six trials that enrolled only women with locally advanced cervical cancer was 0.78 (95% CI, 0.67 to 0.90). The pooled relative risk for the two trials in high-risk early-stage disease also demonstrated a statistically significant benefit for the addition of cisplatin-based chemotherapy to radiotherapy (RR=0.56; 95% CI, 0.41 to 0.77). CONCLUSION: This meta-analysis confirms that treatment with concurrent cisplatin-based chemotherapy plus radiotherapy improves overall survival over various controls in women with locally advanced cervical cancer, large stage IB tumours (prior to surgery) and high-risk early-stage disease (following surgery). The variation in control treatments and the quality of their delivery among the randomized trials makes interpretation difficult. Nonetheless, the meta-analysis supports the use of concurrent cisplatin with radical radiotherapy in the treatment of cervical cancer. PMID- 12109824 TI - Intra-uterine death resulting from placental metastases in adenocarcinoma of unknown primary. AB - A thirty-five year old woman presented with bilateral neck, chest wall and back masses. She was 16 weeks pregnant. Lymph node excision revealed metastatic poorly differentiated adenocarcinoma of unknown primary. Abdominal ultrasound showed a mildly enlarged spleen and a 2-3 cm porta hepatis node. All other investigations were negative. The lymph node and cutaneous metastases progressed rapidly so it was decided to initiate systemic chemotherapy with a view to delivery at 28 weeks gestation by Caesarean section. Shortly after the second 3-weekly cycle of cisplatinum chemotherapy the patient suffered severe lower back and hip pain with MRI scan showing multiple bony metastases in the pelvic girdle. Ultrasound revealed the fetus to have been dead for at least 10 days. The products of conception were delivered following medical induction of labour. Two days later the patient suffered a cardiac arrest from which she could not be resuscitated. Placental histology revealed extensive metastases. With the exception of melanoma this has rarely been reported in solid adult malignancy. As a cause of fetal death, placental metastases are extremely rare. PMID- 12109825 TI - Measuring the quality of cancer care: how do we judge 'appropriate' clinical practice? PMID- 12109826 TI - Factors influencing the use of thoracic radiotherapy in lung cancer--an analysis of the 1995 Scottish lung cancer audit. AB - AIMS: To determine the frequency of delivery of thoracic radiotherapy (TRT) to patients with lung cancer in Scotland in 1995, and identify patient, disease and process variables affecting the probability of receiving TRT. METHODS: Retrospective case note audit of all patients with lung cancer diagnosed in Scotland in 1995. RESULTS: 1118 (30.8%) of 3,855 patients diagnosed with lung cancer in Scotland in 1995 for whom the medical records could be traced received TRT. In those who did not have small cell lung cancer, multivariate analysis indicated that diagnosis by a lung cancer specialist, clinical extent of disease and microscopic verification of cancer (all P<0.0001) and age (P=0.0005) were associated with an increased chance of receiving TRT. There was also a wide variation between different Health Boards (HB) of residence in the proportion of patients receiving TRT (P<0.0001). There was no association between the presence of local symptoms (cough, chest pain or haemoptysis) and the delivery of TRT. Of 351 patients with limited stage small cell lung cancer, 51 (14.5%) received chemotherapy and TRT, and 19 (5.4%) chemotherapy and cranial irradiation. CONCLUSIONS: TRT was delivered to fewer than one-third of patients with lung cancer in Scotland in 1995. This is lower than reported in other international audits. The chance of receiving TRT seemed to be associated with service issues rather than clinical need. PMID- 12109827 TI - Radiological stenting provides effective palliation in malignant central venous obstruction. AB - Malignant superior vena caval obstruction (SVCO) due to lung cancer carries a poor prognosis and response to palliative radiotherapy is variable. Eighteen patients presenting with the appearance of malignant SVCO (thirteen due to primary bronchial tumours) in a four year period from June 1995 to July 1999 underwent venography with a view to placement of an expanding metal stent under radiological control. Fourteen patients were male and four female with a mean age at presentation of 65 years (range 44-84 years). At venography only three patients had isolated SVC stenosis, fivealso had brachiocephalic, internal jugular or subclavian vein stenoses and one had an isolated right brachiocephalic stenosis. Venography demonstrated thrombus in eight of the remaining patients, seven in association with stenosis. In one patient stenosis of the SVC was thought to be due to radiation fibrosis as no mass was visible. All underwent Seldinger catheterization of the great veins via a right femoral puncture with deployment of the stent over a guide wire. Balloon angioplasty was performed prior to stent placement in eight patients. The eight patients with thrombus underwent thrombolysis using rt-PA (recombinant tissue-type plasminogen activator) with subsequent stent placement in seven. Radiological relief of obstruction with abolition of collateral flow was observed in all cases. One patient required further stenting for recurrent obstruction after 160 days of palliation. The median duration of palliation after stenting was 87 days. No procedure-related morbidity was observed. Thrombolysis was of value in patients with occlusive thrombus, either in isolation or complicating a malignant stricture, where radiotherapy may not have been effective. Radiological stenting is a safe technique which offers rapid palliation of SVCO. PMID- 12109828 TI - External beam radiotherapy: a treatment option for massive haemoptysis caused by mycetoma. AB - Five patients with life threatening haemoptysis secondary to a mycetoma were treated with external beam radiotherapy (EBRT). External beam radiotherapy of 3.5 Gy was given once a week continuing for one fraction after the haemoptysis stopped. Three patients required 7 Gy, one required 10.5 Gy and the fifth patient required 14 Gy before the haemoptysis had completely stopped. Irradiation was successful in achieving haemostasis with no side effects being observed after treatment in all five patients. Radiation therapy is an effective modality of treatment for life-threatening haemoptysis due to a mycetoma with no significant acute or late side effects. PMID- 12109829 TI - Accelerated radiation therapy, seven fractions per week, for advanced head and neck cancer--a feasibility study. AB - AIM: The feasibility and improved efficacy of six conventional fractions per week has previously been proven in a Danish randomized trial. We tested the tolerance and efficacy of seven conventional fractions per week using a concomitant boost technique. METHODS: From September 1996 to May 1998, 20 patients with squamous cancer of the head and neck were treated with radiation alone. The site of disease was oropharynx in 35%, larynx in 30%, oral cavity 20%, and hypopharynx in 15%. All patients had stage III (10%)/IV (90%) disease. The planned total dose to gross disease was 66 Gy delivered in 33 fractions of 2 Gy each in 31 days. Large volumes were treated to 46 Gy, 2 Gy per fraction, once each morning, Monday Friday. Boosts to gross disease consisted of 20 Gy in 10 fractions > or = 6 h after the morning dose on Tuesday and Thursday. RESULTS: Acute toxicity > or = grade 3 was mucous membrane 75%, pharynx 60%, skin 65%, and larynx 35%. One acute toxicity was fatal. Chronic toxicity > or = grade 3 (three patients) was mucous membrane 5%, pharynx 10%, skin 5%, salivary 15%, and larynx 5%. All patients with grade III or greater late toxicity had grade III acute toxicity in each toxicity category. At 30 months Kaplan-Meier survival is 55%, and local control is 39%. CONCLUSIONS: Without increasing resource utilization this scheme accelerates treatment by 30%. As expected acute toxicity is high but manageable. Chronic toxicity appears comparable to other altered fractionation strategies however the median follow up is only 30 months more toxicity may emerge as the data matures. We plan further trials using 1.8 Gy fractions to reduce toxicity. PMID- 12109830 TI - Intensity modulation technique using the complementary boost-fields for ethmoid sinus cancer. AB - PURPOSE: To explore a static intensity-modulated radiation therapy (IMRT) technique of a more homogeneous isodose distribution to an irregular-shaped tumour of the ethmoid sinus, with concomitantly sparing the adjacent critical normal organs including the orbit. METHODS AND MATERIALS: We conducted a static IMRT technique adding 2 or smaller complementary boost-fields to the underdosed volume in the PTV, which resulted from complete blocking of the orbits in all coplanar or non-coplanar main ports of the standard 3-D CRT. The standard 3-D CRT plans (Plan A) and IMRT plans adding complementary boost fields (Plan B) were established for 10 patients with ethmoid sinus cancer. Two sets of different plans for each patient were compared using isodose distribution, dose statistics, and dose volume histogram (DVH) of the planning target volume (PTV) and also using dose statistics and DVH of the adjacent critical structures. RESULTS: The IMRT plans adding 2 or more complementary boost-fields (Plan B) for each patient demonstrated better coverage and improved dose homogeneity of the PTV compared to the standard 3-D CRT plan (Plan A). Moreover, the radiation doses to adjacent normal tissue organs, such as the orbits, optic nerves, brain stem and optic chiasm were similarly spared in both plans. CONCLUSION: With concomitantly sparing the surrounding visual pathway structures, our IMRT technique using the complementary boost-fields was quantitatively better than current standard 3-D CRT technique with respect to the dose homogeneity within the PTV. Therefore, we believe that our technique, though still not ideal, is thorough enough to be used routinely in treatment of ethmoid sinus tumour. PMID- 12109831 TI - Homeopathy: what is it and is it of value in the care of patients with cancer? AB - The underlying principles of homeopathy include treating 'like with like' by remedies which are potentized by serial dilution and succussion. These distinguish homeopathy from other forms of alternative or complementary therapy. Conventional scientific wisdom dictates that homeopathy should have no effect above and beyond placebo but experiments on ultra-high dilutions of solutes together with some clinical data suggest the intriguing possibility that it might do in some circumstances. However, most clinical evidence comes from treating relatively minor self-limiting diseases and little comes from treating life threatening disorders such as cancer. This paper explains the principles of homeopathy and reviews the data on its use in cancer care. PMID- 12109832 TI - Homoeopathy: inaccuracies, misunderstandings and half-truths in allopathic doses. PMID- 12109833 TI - Bilateral mental nerve neuropathy. PMID- 12109834 TI - Cerebral hemorrhage in the course of stereotactic radiosurgery. PMID- 12109835 TI - Management of malignant phaeochromocytoma. PMID- 12109836 TI - Chemotherapy associated arterial thrombosis. PMID- 12109837 TI - Doxorubicin and BOOP. A possible near fatal association. PMID- 12109838 TI - An application of the thresholds of change model to the analysis of mental health data. AB - The threshold of change model (TCM) is a statistical technique for analyzing ordered stages of change variables. TCM focuses on the thresholds that separate the ordered stages, and the effects of explanatory variables are evaluated in terms of raising or lowering the thresholds. TCM also allows the explanatory variables to exert differential influence on each threshold. In this paper, we use TCM to analyze the data from a clinical trial that compared assertive community treatment (ACT) with standard case management (SCM) for patients with co-occurring severe mental illness and substance use disorder. Endpoint data (36 month follow up) were used for this analysis. The response variable is the recoded Substance Abuse Treatment Scale with three ordered levels (engagement/persuasion, active treatment, and recovery/relapse prevention), and hence two thresholds. The explanatory variables are gender and group (ACT vs. SCM). The results indicate that gender exerts constant and significant effects on both thresholds. The group effect is somewhat mixed: ACT lowers the first threshold (active treatment), but raises the second threshold (recovery/relapse prevention). PMID- 12109839 TI - How capitated mental health care affects utilization by youth in the juvenile justice and child welfare systems. AB - Examine the impact of Colorado's Medicaid mental health carve-out program on children in child welfare and juvenile justice systems. Medicaid claims and encounter data for two experimental managed care sites and one comparison fee-for service site are used to estimate a two-part model of inpatient, outpatient, and residential treatment center utilization, controlling for patient characteristics. The study finds that juvenile justice and child welfare populations were more strongly affected by managed care than the general youth population, regarding reduced utilization of inpatient and outpatient services. Increases in Residential Treatment Centers use were greater for juvenile justice than either the child welfare sample or the total sample. Youth in child welfare increase utilization of outpatient services. Most utilization effects are stronger for not-for-profit than for-profit managed care organizations. The experience of Colorado implies that a mental health carve-out affects patterns of care for youth and differentially so for youth in juvenile justice and child welfare systems. Controlling for population characteristics, the effects are stronger for not-for-profit than for-profit managed care organizations. PMID- 12109841 TI - Interpreting results in mental health research. AB - It is often difficult to interpret the clinical or policy significance of findings from mental health research when results are presented only in terms of statistical significance. Results expressed in terms of p values or as a metric corresponding to a mental health status scale are seldom intuitively meaningful. To help interpret the significance of research results, we demonstrate a social validity approach that relates scores on mental health status scales to four subsequent major life events. A logistic regression model is used to estimate the relation between mental health status scores and the probability of subsequent major life events, using data obtained on Medicaid beneficiaries with schizophrenia from an evaluation of the Utah Prepaid Mental Health Plan. Using this relatively simple approach will demonstrate to policy makers, clinicians, and researchers the social impact of an outcome, thereby aiding in the interpretation of the significance of results. PMID- 12109840 TI - Treated prevalence, incidence, and pharmacotherapy of child and adolescent mood disorders in an HMO. AB - This study examined the "treated" prevalence, incidence, and pharmacotherapy of child and adolescent mood disorders in a managed care setting. General prevalence patterns across age and sex were similar to those reported in community epidemiology studies, although, not unexpectedly, the overall rates were somewhat lower. Primary care providers identified a substantial proportion of the youth with a mood disorder. Antidepressant medication was used more often by youth identified with a mood disorder in medical care settings than by those youth identified in specialty mental health care settings. These results are valuable in determining if youth with mood disorders receiving medication prescriptions across settings are treated according to current best practice guidelines (such as the adult depression guidelines [U.S. Department of Health and Human Services, 1993]), that is, primarily with SSRI medications. PMID- 12109842 TI - Trade-offs in the conduct of economic evaluations of child mental health services. AB - Childhood psychological morbidity places both short-term and long-term costs on families, health services, social services, voluntary organizations, and society as a whole. Internationally, there have been few economic evaluations alongside clinical trials of population-based interventions to improve child mental well being. This paper sets out a number of trade-offs relevant to the evaluation of the costs and benefits of multisectoral interventions to improve child mental well-being. These trade-offs have relevance for those who fund research, those who conduct research, and those who make policy decisions. The paper sets out a conceptual model for the economic evaluation of population-based interventions to improve child mental health. Finally, the paper argues that with the advent of "joined up" public policy and the breakdown of traditional ring-fenced budgets for health and social care, there is now, more than ever before, an opportunity for the research community to generate guidance for multisectoral collaborative interventions to promote the mental well-being of children and their families. PMID- 12109843 TI - Practical considerations in the pathologic diagnosis of needle core biopsies of breast. AB - The success of needle core biopsy procedures and the validity of pathologic diagnoses made on material from the procedures are key determinants in planning the optimal management of a wide variety of breast diseases. The most common diagnostic problems encountered in these biopsy specimens include lobular and ductal proliferations, sclerosing and papillary lesions, cellular fibroepithelial tumors, and minimally invasive or microinvasive carcinoma. This review provides practical guidance to help narrow the differential diagnosis of lesions on needle core biopsies and offers guidelines for the pathologic reporting of these specimens. PMID- 12109844 TI - Welcome innovation in erythrocyte sedimentation testing. PMID- 12109845 TI - Vanilloid (capsaicin) receptors in health and disease. AB - The cloned vanilloid (capsaicin) receptor subtype 1 (VR1) integrates multiple noxious stimuli on peripheral terminals of primary sensory neurons. The initial excitation of these neurons is followed by a lasting refractory state, traditionally termed desensitization, that has clear therapeutic potential. Capsaicin is used to relieve neuropathic pain, uremic pruritus, and bladder overactivity. The ultrapotent vanilloid resiniferatoxin, now in phase 2 clinical trials, has improved tolerability. A less recognized human exposure to high capsaicin concentrations may occur by pepper sprays used in law enforcement. Evidence is mounting that VR1 expression is not restricted to sensory neurons. From the olfactory bulb to the cerebellum, VR1-expressing neurons are present in a number of brain nuclei, where they might be activated by anandamide. VR1 presence also was demonstrated in nonneuronal tissues. These discoveries place VR1 in a much broader perspective than pain perception and enhance the potential for unforeseen side effects, especially following prolonged vanilloid therapy. The expression of VR1 is plastic and down-regulated during vanilloid therapy, which might have a pivotal role in desensitization. Good evidence suggests altered VR1 expression in various disease states. This recognition not only may provide novel insights into pathogenesis but also may prove useful in diagnosis. PMID- 12109846 TI - Performance characteristics of four immunoassays for antiepileptic drugs on the IMMULITE 2000 automated analyzer. AB - Carbamazepine, phenobarbital, phenytoin, and valproic acid are commonly used antiepileptic drugs that show complicated pharmacokinetic behavior Nonisotopic immunoassays are used routinely to monitor these drugs, and assay specificity is important to obtain accurate results. By using samples from subjects receiving each of these antiepileptic medications, competitive immunoassays for them were evaluated on an IMMULITE 2000 automated chemiluminescent analyzer (Diagnostic Products, Los Angeles, CA). Phenytoin assays were evaluated using an additional set of samples from patients with abnormal renal function. All 4 methods were linear, had imprecision of less than 10%, and compared well with other commercial immunoassays. A positive bias was observed for phenytoin measured in samples from uremic patients compared with a high-performance liquid chromatography reference method. The molar cross-reactivity of carbamazepine-10,11-epoxide was 12% in the carbamazepine assay. Phenytoin metabolites and fosphenytoin had substantial cross reactivity in the phenytoin assay. All antiepileptic drug assays performed well and are suitable for use in monitoring patients receiving antiepileptic drug therapy. One possible exception is the phenytoin assay with samples from patients with renal insufficiency. PMID- 12109847 TI - Endogenous and exogenous digoxin-like immunoreactive substances: impact on therapeutic drug monitoring of digoxin. AB - Endogenous digoxin-like immunoreactive substance (DLIS) was first reported in volume-expanded dogs. Its presence has been confirmed in blood, urine, and other body fluids. Elevated DLIS concentrations are encountered in patients with volume expanded conditions such as uremia, essential hypertension, liver disease, and preeclampsia. DLISs cross-react with antidigoxin antibodies and falsely elevate serum digoxin concentrations, interfering in interpretation of results for therapeutic digoxin monitoring. Falsely lower digoxin values due to the presence of DLISs have been reported. The association of DLISs with volume expansion led to speculation that they could be natriuretic hormones. Several structures have been proposed for DLISs, including nonesterified fatty acid, phospholipid, lysophospholipid, bile acid, bile salt, and steroid. Exogenous DLISs can be found in serum after ingestion of various Chinese medicines and therapy with spironolactone, canrenone, or potassium canrenoate. Like endogenous DLISs, exogenous DLISs interfere with serum digoxin assays, complicating therapeutic digoxin monitoring. However, most reported endogenous and exogenous DLISs are strongly protein-bound while digoxin is weakly protein-bound. Therefore, interference of both endogenous and exogenous DLISs in serum digoxin measurement can be eliminated by monitoring digoxin concentrations in the protein-free ultrafiltrates. PMID- 12109848 TI - Validation of the Diesse Mini-Ves erythrocyte sedimentation rate (ESR) analyzer using the Westergren ESR method in patients with systemic inflammatory conditions. AB - The Diesse Mini-Ves (DMV) erythrocyte sedimentation rate (ESR) analyzer was designed to improve efficiency in determining the ESR. We compared the Westergren ESR method with the 4-sample DMV ESR analyzer for performance and clinical correlation. This prospective observational study, conducted at a 450-bed tertiary medical center, evaluated 291 paired samples from subjects with various systemic inflammatory conditions. Linear regression analysis revealed a statistically significant correlation between the 2 methods. Satisfactory precision of the DMV analyzer was obtained for high and mid-range ESR values. The 4-sample DMV ESR analyzer was precise and comparable in results to the Westergren ESR method. This DMV ESR analyzer is now used at our medical center based on quality control improvements that include a faster, safer, and more standardized ESR method. Hospital or office-based clinical laboratories should consider using the 4-sample DMV ESR analyzer in place of the Westergren method. PMID- 12109849 TI - Targeted therapeutics and implications for pathologists. PMID- 12109851 TI - Evaluation of a single-tube multiplex polymerase chain reaction screen for detection of common alpha-thalassemia genotypes in a clinical laboratory. AB - We prospectively compared a single-tube multiplex polymerase chain reaction (PCR) for detecting alpha-thalassemia with our current approach using 452 blood samples. Initial evaluation of 89 specimens revealed sensitivity and specificity, respectively, for the hemoglobin H inclusion body test (HbH prep) vs PCR for detecting alpha0-thalassemia carriers of 0.79 and 0.96 and for a mean corpuscular volume (MCV) of 82 microm3 (82 fL) or less, 1.0 and 0.45. Detection of all alpha thalassemia genotypes was significantly lower for HbH prep and MCV (sensitivity and specificity, respectively: HbH prep, 0.48 and 0.96; MCV, 0.87 and 0.47). In a follow-up evaluation of patients with positive HbH prep results or suspected alpha-thalassemia prescreened by low MCV, the sensitivity and specificity, respectively, of HbH prep vs PCR increased to 0.97 and 0.93 for alpha0 thalassemia and 0.83 and 0.92 for any alpha-thalassemia. PCR detected alpha thalassemia in 37.2% of 298 suspected alpha-thalassemia cases with suggestive indices but negative HbH prep results and no detectable hemoglobinopathy. This multiplex approach was more sensitive than the HbH prep for detecting all alpha thalassemia genotypes, particularly alpha+-thalassemia; was particularly valuable for identifying carriers of alpha0-thalassemia at risk for offspring with hemoglobin Bart hydropsfetalis, regardless of other diagnosed hemoglobinopathies; and is an ideal adjunct to standard clinical screening protocols for detecting alpha-globin deletions. PMID- 12109850 TI - A clarification. PMID- 12109852 TI - Hodgkin disease in adult and juvenile groups from two different geographic regions in Brazil: characterization of clinicopathologic aspects and relationship with Epstein-Barr virus infection. AB - We analyzed clinicopathologic data, immunophenotype, and Epstein-Barr virus (EBV) status in 96 cases of Hodgkin disease (HD) in juveniles (younger than 20 years) and adults (20 years or older) from 2 distinctive states in Brazil. We studied 34 juvenile (group 1) and 16 adult (group 2) cases from Ceara and 31 juvenile (group 3) and 15 adult (group 4) cases from Sao Paulo. Ceara has a socioeconomic profile similar to a developing country; Sao Paulo is in better economic condition. Mixed cellularity (MC) was the major histologic subtype among groups 1 (22 [65%]), 3 (21 [68%]), and 4 (7 [47%]); nodular sclerosis (NS) was more frequent in group 2 (8 [50%]). EBV infection was observed in 61 cases (64%), including the following (among others): group 1, MC, 22 (65%) and NS, 4 (12%); group 2, NS, 3 (19%) and MC, 2 (12%); group 3, MC, 16 (52%) and NS, 1 (3%); and group 4, MC, 7 (47%). There was predominance of EBV+ HD cases in group 1 compared with group 3. HD in Brazilian patients is highly associated with EBV infection, but geographic differences reflect histologic subtypes and age distribution. PMID- 12109854 TI - Dual sampling of the endocervix and its impact on AutoCyte Prep endocervical adequacy. AB - We compared satisfactory for evaluation but limited by (limited by) and unsatisfactory gynecologic cytologic diagnoses for samples collected by conventional smearing with those generated with the AutoCyte Prep in a population with a historic squamous intraepithelial lesion (SIL) rate of less than 1%. Results from 18,819 AutoCyte Preps were compared with 53,835 conventional cervical smears. Furthermore, 23 women ages 18 to 65 years undergoing annual Papanicolaou tests underwent sequential sampling with the AutoCyte Prep and the Surgipath C-E brush. Comparison of the AutoCyte Prep with conventional cytologic diagnoses revealed the following: unsatisfactory rate, down 97%; limited by rate, down 67%; low-grade SIL rate, up 86%; cervical cancer rate, up 300%; and high grade SIL rate, unchanged. Examination of unsatisfactory and limited by cases for the AutoCyte Prep showed that 88% were due to absence of endocervical cells (ECs). Dual sampling showed no improvement in EC recovery over the AutoCyte collection device. Compared with conventional Papanicolaou smears, the AutoCyte Prep significantly decreased the rate of unsatisfactory and limited by specimens while increasing low-grade SIL and cancer detection and EC recovery. The majority of limited by specimens with the AutoCyte Prep were due to absence of ECs, but use of a brush-type device for better endocervical sampling did not enhance EC recovery. PMID- 12109853 TI - Expression of CD117 and CD11b in bone marrow can differentiate acute promyelocytic leukemia from recovering benign myeloid proliferation. AB - The morphologic characteristics of bone marrow aspirates from patients recovering from acute agranulocytosis may be closely similar to the pattern observed in cases of acute promyelocytic leukemia (APL). The clinical manifestation also can be ambiguous in a substantial number of cases. The immunophenotypic features of bone marrow from 5 patients recovering from acute agranulocytosis, showing an increase in the percentage of promyelocytes (26%-66%), were compared with the immunophenotype of 31 consecutive patients with APL whose diagnosis was confirmed by PML-RAR alpha gene rearrangement. All markers were similarly expressed, except for CD117 and CD11b. CD117 was positive in 24 (77%) of the APL cases and in none of the acute agranulocytosis cases. On the other hand, CD11b was positive in 5 (100%) of the acute agranulocytosis cases and in only 2 (6%) of the APL cases. Thus, the CD117-CD11b+ phenotype was detected in all patients recovering from agranulocytosis and in only 1 (3%) of 31 APL cases. Therefore, we suggest that the combination of both markers is helpful in the differentiation of APL from recovering benign myeloid proliferation. PMID- 12109855 TI - MUC1 expression is correlated with nuclear grade and tumor progression in pT1 renal clear cell carcinoma. AB - We studied, by immunohistochemical analysis, the expression of MUC1 and epithelial membrane antigen in 44 stage pT1 renal cell carcinomas (RCCs). Six patients had a metastatic evolution. The percentage of stained cells was determined for each tumor. All tumors and normal adjacent renal parenchyma were stained. In normal kidney, distal convoluted tubules and collecting ducts stained strongly with an apical distribution. In tumors, there was a significant statistical correlation of the MUC1 expression level with the nuclear grade and with tumor progression. High-grade tumors had more stained cells than did low grade tumors. Metastatic tumors also were more stained than nonmetastatic lesions. By using the Kaplan-Meier method and the log-rank test, we observed that patients with fewer than 10% of stained cells had no metastatic evolution. In contrast, patients with 70% or more stained cells had significantly lower metastasis-free survival rates. We conclude that MUC1 is expressed in RCC and is associated with tumor progression in pT1 RCC. PMID- 12109856 TI - Immunohistochemical validation of a novel epithelial and a novel stromal marker of pancreatic ductal adenocarcinoma identified by global expression microarrays: sea urchin fascin homolog and heat shock protein 47. AB - We extended the results of a previous microarray analysis by immunohistochemical validation of differential protein expression in a series of 57 surgically resected infiltrating ductal pancreatic adenocarcinomas. Two representative genes were examined: sea urchin fascin homolog (overexpressed in both cell lines and primary tumors) and heat shock protein 47 (HSP47; overexpressed in primary tumors only). Protein expression also was evaluated in the precursor lesions of pancreatic cancer pancreatic intraepithelial neoplasia (PanIN), and normal ductal epithelium. Fascin expression was seen in the neoplastic cells of 54 (95%) of 57 ductal adenocarcinomas but not in 49 (94%) of 52 adjacent nonneoplastic epithelium. In the multistep pathogenesis of ductal adenocarcinomas, fascin expression seemed to be a late event, usually present in PanINs 2 and 3. HSP47 expression was almost universal and most intense in the ductal adenocarcinoma associated stromal desmoplasia (57/57), although 37 cases (65%) also expressed HSP47 in the neoplastic epithelium. HSP47 expression was absent in the majority of nonneoplastic pancreata (46 [88%]). Fascin and HSP47 are novel tumor markers with potential diagnostic and therapeutic implications for pancreatic carcinoma. These results establish the usefulness of global expression platforms to identify novel tumor markers. PMID- 12109857 TI - Gene amplification and protein overexpression of c-erb-b2 in Barrett carcinoma and its precursor lesions. AB - We examined 39 samples of metaplastic specialized epithelium (SE), 27 of low grade dysplasia (LGD), 27 of high-grade dysplasia (HGD), and 46 of adenocarcinoma (CA) derived from Barrett esophagus for c-erb-b2 gene amplification using differential polymerase chain reaction and for overexpression of c-erb-b2 protein using immunohistochemical analysis. Amplification of the c-erb-b2 gene was as follows: SE, 0.0%; LGD, 0.0%; HGD, 11.1%; and CA, 13.6%; and protein overexpression was as follows: SE, 0.0%; LGD, 7.4%; HGD, 18.5%; and CA, 21.7%. In 8 (89%) of 9 samples, c-erb-b2 gene amplification correlated with protein overexpression. The reverse was true in 8 (47%) of 17 samples: c-erb-b2 protein overexpression was proved with simultaneous gene amplification. Amplification of c-erb-b2 is a late event in the carcinogenesis of Barrett esophagus. In contrast, protein overexpression appears more often and earlier Besides gene amplification, other mechanisms to induce protein overexpression must exist. PMID- 12109858 TI - Clinicopathologic significance of expression of CD44s and CD44v6 isoforms in squamous cell carcinoma of the supraglottic larynx. AB - CD44 splice variants are assumed to have a critical role in the malignant progression of many human tumors. However, the clinical significance of CD44 expression is not yet understood. The aim of this study was to investigate the prognostic significance of expression of CD44s and CD44v6 isoforms in squamous cell carcinomas of the supraglottic larynx. CD44s and CD44v6 expression was determined by immunohistochemical analysis of paraffin-embedded tissue specimens from 101 patients. There was a significant correlation between decreased CD44s or CD44v6 expression and a poorer histologic differentiation. No relationship was observed with T stage or nodal metastasis. Decreased CD44s expression, but not CD44v6 expression, correlated with increased recurrence rates. There was no correlation between the decreased expression of any isoform tested and survival. These data confirm a reduction of CD44s and CD44v6 expression in poorly differentiated tumors. However, these changes do not offer a useful adjunct to current prognostic indicators. PMID- 12109859 TI - Role of liver biopsy in the diagnosis of hepatic iron overload in the era of genetic testing. AB - We studied hepatic iron overload (HIOL) patterns in 32 patients who underwent liver biopsies and testing for HFE mutations (C282Y, H63D). Iron-stained biopsy specimens were examined for patterns of iron deposits: hereditary hemochromatosis (HH) pattern or non-HH pattern. Visual iron grade based on amount of cellular and lobular iron was evaluated. We found the HH pattern in 17 biopsy specimens (53%) and the non-HH pattern in 6 specimens (19%). HH with superimposed non-HH was noted in 9 cases (28%). In 25 patients with HFE mutations, HH alone and combined with non-HH patterns was noted in 22 specimens (88%). Visual iron grade correlated approximately with the hepatic iron index. Heavy HIOL was noted in C282Y homozygotes and 1 patient with cirrhosis without either HFE mutation. Mild steatohepatitis was found in 21 specimens (66%); it was associated with the non HH pattern in 80% (12/15) and the HH pattern in 62% (16/26) of cases. Liver biopsy can identify pattern and grade of HIOL and associated pathology for diagnosis and management of patients with abnormal iron studies and elevated liver function test results. Genetic tests for HFE mutations and liver biopsies are complementary in the workup of these patients. PMID- 12109860 TI - Can mucinous lesions of the breast be reliably diagnosed by core needle biopsy? AB - Lesions of the breast containing extravasated mucin span a continuum from benign mucoceles to invasive mucinous (colloid) carcinoma. It is well known that distinguishing benign from malignant mucinous lesions is difficult infine-needle aspiration material. Whether these lesions also are difficult to distinguish in core needle biopsy material is not known. To address this, I reviewed the results of 4,297 breast core needle biopsies. Mucinous lesions were identified in 22 cases (0.51%), and excisional biopsy material was available for 15 of these. At excision, 0 of 8 benign mucinous lesions showed carcinoma, while 7 of 7 mucinous lesions associated with carcinoma at the time of core needle biopsy showed carcinoma at excision. The vast majority of mucinous lesions of the breast can be diagnosed accurately by core needle biopsy. Whether all such lesions require excision is not known at this time. PMID- 12109861 TI - Detection of malignant epithelial cells in effusions using flow cytometric immunophenotyping: an analysis of 92 cases. AB - We compared the efficiency of immunophenotyping using flow cytometry (FCM) and a combination of morphologic and immunocytochemical studies for detecting malignant cells in 92 effusions. Cytologic results were as follows: carcinoma cells, 43 specimens; benign, 42 specimens; suggestive of nonepithelial malignancy, 7 specimens. After immunocytochemical analysis, 5 benign specimens were reclassified as malignant and 4 malignant epithelial specimens as benign. With FCM, cells positive for Ber-EP4, B 72.3, AH6, and HB-TN were detected in 28 to 36 (64%-82%) of 44 carcinomas but only 2 to 12 (5%-29%) of 41 benign specimens. Significant association was seen for coexpression. Ber-EP4 and AH6 were the most sensitive; Ber-EP4 was the most specific. The presence of cells positive for 3 of 4 markers strongly suggested malignancy (34/44 carcinoma specimens [77%]; 3/41 reactive specimens [7%]). The presence of cells positive for all 4 markers was diagnostic of malignancy (17/44 malignant specimens [39%]; 0/41 reactive effusions [0%]). FCM and immunocytochemical resultsfor Ber-EP4 expression showed excellent association. FCM is a powerful tool for diagnosing difficult effusions and can quantify coexpression of various markers in fresh specimens. By using established cellular markers coupled with biological markers, FCM also has great promise for experimental purposes. PMID- 12109862 TI - Kinetic profiles by topographic compartments in muscle-invasive transitional cell carcinomas of the bladder: role of TP53 and NF1 genes. AB - We evaluated 71 muscle-invasive transitional cell carcinomas (TCCs) of the bladder by tumor compartments. Kinetic parameters included mitotic figure counting, Ki-67 index, proliferation rate (DNA slide cytometry), and apoptotic index (in situ end labeling [ISEL] of fragmented DNA using digoxigenin-labeled deoxyuridine triphosphate and Escherichia coli DNA polymerase [Klenow fragment]). At least 50 high-power fields per compartment were screened from the same tumor areas; results are expressed as percentage of positive neoplastic cells. Mean and SD were compared by tumor compartment. DNA was extracted from microdissected samples (superficial and deep) and used for microsatellite analysis of TP53 and NF1 by polymerase chain reaction-denaturing gradient gel electrophoresis. Significantly higher marker scores were revealed in the superficial compartment than in the deep compartment. An ISEL index of less than 1% was revealed in 63% (45/71) of superficial compartments and 86% (61/71) of deep compartments. Isolated NF1 alterations were observed mainly in superficial compartments, whereas isolated TP53 abnormalities were present in deep compartments. Lower proliferation and down-regulation of apoptosis define kinetically the deep compartment of muscle-invasive TCC of the bladder and correlate with the topographic heterogeneity, NF1-defective in superficial compartments and TP53 defective in deep compartments. PMID- 12109863 TI - Clinical endpoints. PMID- 12109864 TI - Prevention of coronary restenosis with radiation therapy: a review. AB - The problem of restenosis after percutaneous transluminal coronary angioplasty remains the major limiting factor of the procedure. Over the last 10 years, investigators have been studying the use of radiation therapy for preventing restenosis after angioplasty or stent placement. Since radiotherapy has been proven in other cases to be effective in disrupting the cell cycle regulatory proteins and thereby slowing or stopping growth, it was decided to apply the same principle to neointimal hyperplasia. To review the data that have emerged regarding vascular radiation with an emphasis on irradiated stents, 65 articles were reviewed and both preclinical and clinical experiments were included. Overall, studies with gamma and beta radiation show promising results. Endovascular gamma radiation has been shown effective in randomized trials, even at 3-year follow-up. Beta radiation is preferred because of greater safety and localization, and because it has also shown encouraging results in initial clinical trials, as well as in larger randomized studies. Consequently, the Federal Drug Administration has approved the use of both. In both types of endovascular brachytherapy, it seems the greater the dose, the better the initial response. Safety concerns include an increased incidence of late thrombosis and greater restenosis at margins. With irradiated stents, however, the situation is not as clear. At times, animal models have presented confusing results. These have ranged from significant suppression of hyperplasia to outright adverse effects of radiation on the vessel wall. While some clinical trials have been encouraging, others have not. Follow-up of up to 1 year has been disappointing so far. Many issues, such as the "candy wrapper" effect and rebound hyperplasia, must be dealt with before this becomes a viable form of therapy. It has become clear that radiation therapy in this setting, while having potentially great benefits, can cause deleterious effects as well. However, the mixed bag of positive and negative results seen so far, and the attractiveness of stents or percutaneous transluminal coronary angioplasty being "restenosis-proofed," eventually is cause for cautious optimism. PMID- 12109865 TI - Low-molecular-weight heparin for the treatment of patients with mechanical heart valves. AB - BACKGROUND: The interruption of oral anticoagulant (OAC) administration is sometimes indicated in patients with mechanical heart valves, mainly before noncardiac surgery, non-surgical interventions, and pregnancy. Unfractionated heparin (UH) is currently the substitute for selected patients. Low-molecular weight heparin (LMWH) offers theoretical advantages over UH, but is not currently considered in clinical guidelines as an alternative to UH in patients with prosthetic valves. HYPOTHESIS: The aim of the present study was to review the data accumulated so far on the use of LMWH in this patient population and to discuss its applicability in common practice. METHODS: For this paper, the current medical literature on LMWH in patients with mechanical heart valves was extensively reviewed. RESULTS: There were eight series and six case reports. None of the studies was randomized, and only one was prospective. Data to establish the thromboembolic risk were incomplete. After excluding case reports, the following groups were constructed: (a) short-term administration, after valve insertion (n = 212); (b) short-term, perioperative (noncardiac)/periprocedural (n = 114); (c) long-term, due to intolerance to OAC (n = 16); (d) long-term, in pregnancy (n = 10). The incidence rate of thromboembolism was 0.9% for all the studies and 0.5, 0, 20, and 0% in groups a, b, c, and d, respectively; for hemorrhage, the overall rate was 3.4% (3.8, 2.6, 10, and 0% for the respective groups). CONCLUSIONS: In patients with mechanical heart valves, short-term LMWH therapy compares favorably with UH. Data on mid- and long-term LMWH administration in these patients are sparse. Further randomized studies are needed to confirm the safety and precise indications for the use of LMWH in patients with mechanical heart valves. PMID- 12109866 TI - Sustained ventricular tachycardia as a marker of inadequate myocardial perfusion during the acute phase of myocardial infarction. AB - BACKGROUND: Sustained ventricular tachycardia (VT) complicating the acute phase of myocardial infarction (AMI) is a quite rare event but with short-term unfavorable prognosis. The clinical characteristics as well as the therapeutic implications have not yet been well defined. HYPOTHESIS: This paper attempts to prove that VT may be considered a marker of inadequate myocardial perfusion after thrombolysis. METHODS: To assess the clinic-electroangiographic characteristics and prognosis of patients with VT occurring within the first 4 days of an AMI, a case-control study was carried out in 23 patients from a total of 1,100 patients (1.9%) hospitalized with AMI between March 1993 and July 1997. These patients were compared with a control group of 131 patients hospitalized consecutively. A statistical analysis was made using the chi-square test, t-test, and logistic regression. RESULTS: There were no differences among groups with regard to age, gender, and area of necrosis. Average time for the onset of VT was 26 h (range 0 92 h). Sixteen patients underwent coronary angiography: 4 patients had left main coronary artery disease, 2 had single-vessel disease, 8 had lesions in two vessels, and 2 had triple-vessel disease. Univariate analysis showed that patients with VT had a higher incidence of creatine phosphokinase (CPK)-MB peak > 300 UI/l (61 vs. 30%; p<0.001), more frequent occurrence of previous AMI (48 vs. 17%; p<0.001), and acute intraventricular conduction disorders (26 vs. 4%; p<0.001). Furthermore, these patients suffered ischemia previous to VT more frequently (65 vs. 11%; p<0.0001), and had a greater mortality rate than that in the control group (35 vs. 4%; p<0.0001). In the multivariant analysis, the variables related to the occurrence of VT were CPK-MB peak > 300 IU/l (OR 5.9; 95% CI 1.6-21), acute intraventricular conduction disorders (OR 9.02; 95% CI 1.7 48), and ischemia immediately prior to VT (odds ratio [OR] 19.64; 95% confidence interval [CI] 5.3-73). CONCLUSIONS: Ventricular tachycardia may be considered a marker of inadequate myocardial perfusion after thrombolysis; therefore, a more aggressive revascularization treatment in these patients would be advisable. The profile of patients with AMI, hospitalized in the coronary care unit, who will likely suffer from VT is previous AMI, CPK-MB peak > 300, acute intraventricular conduction disorders, Killip > I, and ischemia previous to VT. PMID- 12109867 TI - T-peak to T-end interval may be a better predictor of high-risk patients with hypertrophic cardiomyopathy associated with a cardiac troponin I mutation than QT dispersion. AB - BACKGROUND: Patients with hypertrophic cardiomyopathy (HCM) associated with a deletion of lysine 183 (K183del) in the cardiac troponin I (cTnI) gene suffer sudden cardiac death at all ages. However, the correlation between QT variables and sudden cardiac death in these patients remains uncertain. HYPOTHESIS: We evaluated the correlation between QT variables and sudden cardiac death and/or ventricular tachyarrhythmia (SCD/VT) in patients with HCM associated with the cTnI mutation. METHODS: We analyzed 10 probands with HCM associated with the cTnI gene K183del and their family members. The subjects were divided into three groups: Group A (n = 7), mutation carriers with SCD/VT; Group B (n = 16), mutation carriers without SCD/VT; Group C (n = 24), no mutation carriers. QT intervals were corrected using Bazett's formula. RESULTS: Maximum QTc and corrected QT dispersion were significantly longer in Groups A and B than in Group C. However, there were no differences in either parameter between Groups A and B. On the contrary, the peak-to-end interval of T wave/QT interval in V5 (Tpe) in Group A was significantly longer than that in Groups B and C. Logistic regression analysis revealed that Tpe was a good clinical predictor for SCD/VT in patients with HCM in this study. CONCLUSIONS: These results suggest that Tpe rather than QT dispersion may be one of the best predictors for SCD/VT in patients with HCM associated with the K183del mutation in the cTnI gene. PMID- 12109868 TI - Coronary collaterals during single-vessel coronary angioplasty: effects of nitroglycerin. AB - BACKGROUND: Although the protective role of collaterals in coronary artery disease (CAD) is well known, the influence of drugs on collateral function remains controversial. HYPOTHESIS: We aimed to investigate prospectively the prevalence of spontaneously visible and recruitable coronary collaterals in consecutive patients with single-vessel CAD and the effect of systemic administration of nitroglycerin on these types of collaterals during percutaneous transluminal coronary angioplasty (PTCA). METHODS: Ipsi- and contralateral coronary artery contrast injections were performed before and during PTCA. Simultaneously with balloon occlusion, we measured coronary artery occlusion pressure via the balloon catheter. All measurements were repeated after administration of 0.5 mg of nitroglycerin intravenously. RESULTS: Of 101 consecutive patients, 24% had spontaneously visible and 30% had recruitable collaterals. Contralateral collaterals were five times more frequent than ipsilateral collaterals. Presence of collaterals was highly associated with the degree of coronary stenosis. Coronary occlusion pressure was higher in patients with than in those without collaterals. Collaterals prevented pain and ischemia during PTCA, and in this respect spontaneously visible collaterals were more effective than recruitable collaterals. There was no effect of systemic administration of nitroglycerin on appearance or occlusion pressure of coronary collaterals. CONCLUSION: Coronary collaterals were found in more than half of patients with single-vessel CAD, as the prevalence of recruitable collaterals was slightly higher than that of spontaneously visible collaterals. Nitroglycerin did neither recruit nor augment coronary collaterals. PMID- 12109869 TI - Images in cardiology: Loeffler's endocarditis resulting from acute lymphoblastic leukemia. PMID- 12109870 TI - Prolonged low-dose thrombolytic therapy: a novel adjunctive strategy in the management of an infected right atrial thrombus. AB - An 81-year-old man presented with a large, infected right atrial thrombus that was refractory to anticoagulants and several courses of antibiotics. The risk of surgical removal of the thrombus, which was associated with a pacemaker electrode, was considered prohibitive. The patient was treated for 7 days with low-dose (40 mg/day) tissue-type plasminogen activator (t-PA). Hemostatic monitoring during infusion revealed (1) a plasma t-PA antigen that was approximately 5% of that achieved during short-course t-PA for acute myocardial infarction, (2) biochemical evidence of prolonged clot lysis, and (3) no significant depletion of fibrinogen or plasminogen. Nearly complete dissolution of the thrombus was observed. His bacteremia was eradicated by intravenous penicillin despite the presence of the pacemaker lead. This case highlights the benefits of combined antibiotic and thrombolytic therapy and documents for the first time the response of the human hemostatic system to prolonged t-PA infusion and the plasma t-PA levels attained when thrombolytic therapy is administered in this manner. Prolonged courses of fibrinolytic agents may be a good alternative to surgical intervention in selected patients with infected, right-sided intracardiac thrombi. PMID- 12109871 TI - Pavlos K. Toutouzas. PMID- 12109873 TI - Mechanisms underlying cytokine-mediated cell-fate regulation in the nervous system. AB - Neurons, astrocytes, and oligodendrocytes, the three major cell types in the nervous system, are generated from common neural stem cells during development. Recent studies have provided evidence that neural stem cells are preserved in the adult brain, where, until recently, neurogenesis had not been considered to take place. The mechanisms that gOvern the fate of neural stem-cell determination have yet to be clarified. It is becoming apparent that soluble protein mediators referred to as cytokines play critical roles in cell-fate determination. For instance, bone morphogenetic proteins (BMPs) alter the fate of developing brain cells from a neurogenic differentiation to an astrocytic one. Different types of cytokines sometimes cooperate to modulate differentiation. For example, the interleukin-6 (IL-6) family cytokines and the BMP family cytokines act in synergy to elaborate astrocyte differentiation. In this review, we focus on recent progress that addresses the molecular mechanisms whereby cytokines regulate the fate of cells in neural lineages. We also discuss possible clinical applications of these findings to minimize the undesirable gliogenesis that occurs after neural stem-cell implantation and nerve injury. PMID- 12109872 TI - Tau function and dysfunction in neurons: its role in neurodegenerative disorders. AB - Alzheimer's disease (AD) is the most usual neurodegenerative disorder leading to dementia in the aged human population. It is characterized by the presence of two main brain pathological hallmarks: senile plaques and neurofibrillary tangles (NFTs). NFTs are composed of fibrillar polymers of the abnormally phosphorylated cytoskeletal protein tau. PMID- 12109874 TI - The role of astroglia on the survival of dopamine neurons. AB - Glial cells play a key role in the function of dopamine (DA) neurons and regulate their differentiation, morphology, physiological and pharmacological properties, survival, and resistance to different models of DA lesion. Several studies suggest that glial cells may be important in the pathogenesis of Parkinson's disease (PD), a common neurodegenerative disorder characterized by degeneration of the nigrostriatal DA system. In this disease the role of glia could be due to the excessive production of toxic products such as nitric oxide (NO) or cytokines characteristic of inflammatory process, or related to a defective release of neuroprotective agents, such as small antioxidants with free radical scavenging properties or peptidic neurotrophic factors. PMID- 12109877 TI - Use of PCR-RFLP for genotyping 16S rRNA and characterizing bacteria cultured from halibut fry. AB - Small subunit ribosomal genes were explored using PCR-RFLP to facilitate the characterization of bacteria cultured from reared fry of the Atlantic halibut (Hippoglossus hippoglossus). Concern has been expressed about pathogen invasion in larvae lacking a counteracting normal flora that may aid the immune system in producing robust noninfected individuals. In this study, pure cultured representatives of normal flora that were previously found to be antagonistic towards a pathogenic Vibrio sp. were subjected to a whole cell PCR protocol amplifying approximately 1500 bp of 16S rDNA. Amplified DNA was digested by AluI, BstUI, CfoI, and RsaI, to generate restriction profiles. Before the isolates were characterized, a survey was performed to test the discriminative efficiency of the RFLP. Efficient detection of polymorphism and the resolution of species and subspecies were achieved. Using the RFLP on 103 isolates generated as many as 22 genotypes. Based on the restriction profiles, a taxonomic tree incorporating 19 reference strains was constructed. Partial sequencing found this tree to be dominated by gamma-Proteobacteria in clusters of Vibrio-, Pseudomonas-, and Alteromonas-affiliated species. Only nine isolates fell outside these genera, including the three isolates Shewanella alga, Deleya marina, and Marinomonas protea. These species have not previously been reported as halibut flora. The most frequently isolated genotype resembled Vibrio salmonicida. PMID- 12109878 TI - Molecular characterization of DNA encoding 16S-23S rRNA intergenic spacer regions and 16S rRNA of pectolytic Erwinia species. AB - Sequences of 16S rDNAs and the intergenic spacer (IGS) regions between the 16S and 23S rDNA of bacterial strains from genus Erwinia were determined. Comparison of 16S rDNA sequences from different species and subspecies clearly revealed intraspecies-subspecies homology and interspecies heterogeneity. Phylogenetic analyses of 16S rDNA sequence data revealed that Erwinia spp. formed a discrete monophyletic clade with moderate to high bootstrap values. PCR amplification of the 16S-23S rDNA regions using primers complementary to the 3' end of 16S and 5' end of 23S rRNA genes generated two DNA fragments. The small 16S-23S rDNA IGS regions of Erwinia spp. examined in this study varied considerably in size and nucleotide sequence. Multiple sequence alignment and phylogenetic analysis of small IGS sequence data showed a consistent relationship among the test strains that was roughly in agreement with the 16S rDNA data that reflected the accepted species and subspecies structure of the taxon. Sequence data derived from the large IGS resolved the strains into coherent groups; however, the sequence information would not allow any phylogenetic conclusion, because it failed to reflect the accepted species structure of the test strains. PMID- 12109879 TI - Physiological diversity and trehalose accumulation in Schizosaccharomyces pombe strains isolated from spontaneous fermentations during the production of the artisanal Brazilian cachaca. AB - Twenty-seven Schizosaccharomyces pombe isolates from seven cachaca distilleries were tested for maximum temperature of growth and fermentation, osmotolerance, ethanol resistance, invertase production, and trehalose accumulation. Two isolates were selected for studies of trehalose accumulation under heat shock and ethanol stress. The S. pombe isolates were also characterized by RAPD-PCR. The isolates were able to grow and ferment at 41 degrees C, resisted concentrations of 10% ethanol, and grew on 50% glucose medium. Four isolates yielded invertase activity of more than 100 micromol of reducing sugar x mg(-1) x min(-1). The S. pombe isolates were able to accumulate trehalose during stationary phase. Two isolates, strains UFMG-A533 and UFMG-A1000, submitted to a 15 min heat shock, were able to accumulate high trehalose levels. Strain UFMG-A533 had a marked reduction in viability during heat shock, but strain UFMG-A1000 preserved a viability rate of almost 20% after 15 min at 48 degrees C. No clear correlation was observed between trehalose accumulation and cell survival during ethanol stress. Strain UFMG-A1000 had higher trehalose accumulation levels than strain UFMG-A533 under conditions of combined heat treatment and ethanol stress. Molecular analysis showed that some strains are maintained during the whole cachaca production period; using the RAPD-PCR profiles, it was possible to group the isolates according to their isolation sites. PMID- 12109875 TI - Microarray technology and its application on nicotine research. AB - Since its development, microarray technique has revolutionized almost all fields of biomedical research by enabling high-throughput gene expression profiling. Using cDNA microarrays, thousands of genes from various organisms have been examined with respect to differentiation/development, disease diagnosis, and drug discovery Nevertheless, research on nicotine using cDNA microarrays has been rather limited. Therefore, it is our intention in this article to report the findings of our cDNA microarray study on nicotine. We first present an overview of the microarray technology, particularly focusing on the factors related to microarray design and analysis. Second, we provide a detailed description of several newly identified biological pathways in our laboratory, such as phosphatidylinositol signaling and calcium homeostasis, which are involved in response to chronic nicotine administration. Additionally, we illustrate how comparisons between microarray studies help identify candidate genes that potentially may explain the observed inverse association between smoking and schizophrenia. Lastly, given the early stage of microarray research on nicotine, we elaborate on the need for an efficient analysis of genetic networks to further enhance our understanding of the mechanisms involved in nicotine abuse and addiction. PMID- 12109880 TI - Diversity of antifungal actinomycetes in various vegetative soils of Korea. AB - Diversity of actinomycetes and their antifungal activities against some plant pathogenic fungi were examined in various vegetative soils from 14 different sites in the western part of Korea. Actinomycete counts ranged from 1.17 x 10(6) to 4.20 x 10(6) cfu x g(-1) dried soil. A total of 1510 actinomycetes were isolated from the soil samples. Streptomyces was predominant in soils with a pH range of 5.1-6.5, 9.1-13.0% moisture, and 9.1-11.0% organic matter. Most Micromonospora, Dactylosporangium, and Streptosporangium were distributed in soils with pH 4.0-5.0, 2.0-9.0% moisture, and 4.0-7.0% organic matter. Actinomadura and nocardioform actinomycetes were abundant in soils with pH 4.0 5.0 and 13.1-20.0% moisture and with 9.1-11.0 and 4.0-7.0% organic matter, respectively. Populations of Streptomyces were predominant in all the soils, but were highest in grassland and lowest in mountain-forest soils. Micromonospora was most abundant in pepper-field soil and nocardioform actinomycetes were highest in rice paddy field soil. Dactylosporangium was predominant in lake-mud sediments and pepper-field soil, Streptosporangium in lake-mud sediments, and Actinomadura in mountain-forest soil. Antifungal actinomycetes were abundant in orchard soil and lake mud. More than 50% of antifungal isolates from most soils were classified as genus Streptomyces. Actinomycete isolates that showed strong antifungal activity against Alternaria mali, Colletotrichum gloeosporioides, Fusarium oxysporum f.sp. cucumerinum, and Rhizoctonia solani were predominant in pepper-field soils, whereas those against Magnaporthe grisea and Phytophthora capsici were abundant in radish-field soils. PMID- 12109876 TI - Molecular advances in understanding inherited prion diseases. AB - The prion diseases are neurodegenerative disorders that have attracted great interest because of the possible link between bovine spongiform encephalopathy (BSE) and variant Creutzfeldt-Jakob disease (CTD) in humans. Possible transmission of these diseases has been linked to a single protein termed the prion protein. This protein is an abnormal isoform of a normal synaptic glycoprotein. The majority of prion diseases does not appear to be caused by transmission of an infectious agent but occur spontaneously with no known cause. The strongest supporting evidence that the prion protein is the causative agent in prion disease comes from specific inheritable forms of prion disease which are linked to single point mutations in the prion protein gene. Paradoxically, these point mutations, although autosomal dominant with 100% penetrance do not lead to disease until late in life. Molecular techniques are now being used extensively to determine how these point-mutations alter the prion protein's normal structure and activity. This review deals with the latest insights into how inherited mutations in the prion protein gene lead to neurodegenerative disease. PMID- 12109881 TI - Characterization of two novel yeast strains used in mediated biosensors for wastewater. AB - After isolation from a pulp mill wastewater treatment facility, two yeast strains, designated SPT1 and SPT2, were characterized and used in the development of mediated biochemical oxygen demand (BOD) biosensors for wastewater. 18S rRNA gene sequence analysis revealed a one nucleotide difference between the sequence of SPT1 and those of Candida sojae and Candida viswanthii. While SPT2 had the highest overall homology to Pichia norvegensis, at only 73.5%, it is clearly an ascomycete, based on BLAST comparisons and phylogenetic analyses. Neighbor joining dendrograms indicated that SPT1 clustered with several Candida spp., and that SPT2 clustered with Starmera spp., albeit as a very deep branch. Physiological tests, microscopic observations, and fatty acid analysis confirmed that SPT1 and SPT2 are novel yeast strains. Physiological tests also indicated that both strains had potential for use in mediated biosensors for estimation of BOD in wastewater. The lower detection limits of SPT1- and SPT2-based K3Fe(CN)6 mediated biosensors for a pulp-mill effluent were 2 and 1 mg BOD/L, respectively. Biosensor-response times for effluents from eight different pulp mills were in the range of 5 min. Reliability and sensitivity of the SPT1- and SPT2-based biosensors were good, but varied with the wastewater. PMID- 12109882 TI - Population structure and mating-type genes of Colletotrichum graminicola from Agrostis palustris. AB - Eighty-seven isolates of Colletotrichum graminicola, mostly from Agrostis palustris, were collected in grass fields, most of which were in Ontario, Canada. Specific primers were designed to amplify the mating-type (MAT) genes and, among 35 isolates tested, all yielded a band of the expected size for MAT2. For six isolates, the MAT2 PCR products were sequenced and found to be similar to that reported for MAT2 of C. graminicola from maize. Based on 119 polymorphic bands from 10 random amplified polymorphic DNA primers, analyses of genetic distances were found to generally cluster isolates by host and geographic origin. Among 42 isolates from a grass field in Ontario, significant spatial autocorrelation was found to occur within a 20-m distance, implying that this is the effective propagule dispersal distance. Although clonal propagation was observed in the 87 isolates with 67 unique genotypes, the extent of genetic variation in local populations implies some occurrence of sexual or asexual recombination. PMID- 12109883 TI - Utilization and cell-surface binding of hemin by Histoplasma capsulatum. AB - Histoplasma capsulatum, a dimorphic fungus capable of causing severe respiratory illness in immunocompromised individuals, resides in macrophages during mammalian infection. Previous studies suggest that siderophore-mediated iron transport may be important for the acquisition of iron from transferrin while the organism resides in macrophages. However, iron is also present as hemin in the intracellular environment of the macrophage and may serve as a major source of iron during infection. Thus the ability of H. capsulatum to use hemin and heme containing compounds was examined. Histoplasma capsulatum G217B was iron-starved by adding the iron chelator deferoxamine mesylate to the culture. The addition of 10 microM hemin in the presence of deferoxamine mesylate restored growth to the levels seen in the absence of the chelator. Histoplasma capsulatum was also cultivated in an iron-limited, chemically defined medium without the addition of chelators and it was determined that the organism could also use hemoglobin as a sole source of iron. The method of iron internalization from heme was examined by measuring hemin binding to the yeast-cell surface. The ability of H. capsulatum to bind hemin was related to the nutritional status of the cells. Cells grown under iron-limited conditions bound more heme to the cell surface than did cells grown in medium without chelator. Pretreatment of iron-starved cells with proteinase K eliminated the ability of the organism to bind hemin. Additionally, the pre-incubation of iron-starved H. capsulatum with hemin eliminated the ability of these cells to remove hemin from the solution, although pre-incubation of cells with the iron-free form of hemin, protoporphyrin IX, only modestly affected the ability of the organism to bind hemin. These results suggest that H. capsulatum uses hemin as a sole source of iron and that one mechanism of iron acquisition involves a cell-surface receptor for hemin. PMID- 12109884 TI - Studies on exudate-depleted sclerotial development in Sclerotium rolfsii and the effect of oxalic acid, sclerotial exudate, and culture filtrate on phenolic acid induction in chickpea (Cicer arietinum). AB - Exudate depletion from developing sclerotia of Sclerotium rolfsii Sacc. in culture caused reduced size and weight of sclerotia. Germination of exudate depleted sclerotia was delayed on Cyperus rotundus rhizome meal agar medium when compared with that of control sclerotia. The exudate-depleted sclerotia caused infection in chickpea (Cicer arietinum) plants in a glasshouse. Different temperatures and incubation periods had no effect on the germination ability of the exudate-depleted sclerotia. Oxalic acid, sclerotial exudate, and culture filtrate of S. rolfsii induced the synthesis of phenolic acids, including gallic, ferulic, chlorogenic, and cinnamic acids, as well as salicylic acid, in treated chickpea leaves. Gallic acid content was increased in treated leaves compared with the untreated controls. Maximum induction of gallic acid was seen in both leaves treated with oxalic acid followed by exudate and leaves treated with culture filtrate. Cinnamic and salicylic acids were not induced in exudate treated leaves. Ethyl acetate fractionation indicated that the sclerotial exudates consisted of gallic, oxalic, ferulic, chlorogenic, and cinnamic acids, whereas the culture filtrate consisted of gallic, oxalic, and cinnamic acids along with many other unidentified compounds. PMID- 12109885 TI - Some factors affecting the adherence of probiotic Propionibacterium acidipropionici CRL 1198 to intestinal epithelial cells. AB - Adhesion to the intestinal mucosa is generally considered an important property of probiotic microorganisms and has been related to many of their health benefits. This study investigated some factors that could affect or be involved in the adherence of Propionibacterium acidipropionici CRL 1198, a dairy strain with suggested probiotic effects and high adherence in vitro and in vivo to intestinal epithelial cells. In vitro adhesion of propionibacteria was decreased by gastric digestion but not affected by bile and pancreatic enzymes. Adherence was also decreased by pretreatment of bacterial cells with protease, sodium metaperiodate, and trichloroacetic acid, revealing that different features of the cell surface, like protein factors, carbohydrates, and teichoic acids, are involved in the process. Adherence to intestinal epithelial cells was enhanced by calcium and was dependent on other divalent cations. Adhesion to intestinal mucus was also demonstrated. The results should explain the metabolic effects in the host previously obtained with this strain and support the potential of Propionibacterium for development of new probiotics. PMID- 12109886 TI - Effect of fur mutation on acid-tolerance response and in vivo virulence of avian septicemic Escherichia coli. AB - The Fur (ferric uptake regulator) protein is a master regulator of iron metabolism in gram-negative bacteria. In the present study, the effect of a partial deletion of the fur gene on the acid-tolerance response and in vivo virulence of avian Escherichia coli was examined. The fur mutant was unable to trigger the acid-tolerance response as observed in the wild-type parent strain. However, the mutant was as virulent as the wild-type parent strain when tested in 1-day-old chickens by subcutaneous inoculation. These data indicate that the fur gene is involved in the acid-tolerance response but not involved in the virulence of E. coli, as detected by the ability to cause septicemia in our experimental infection. PMID- 12109887 TI - Cryptococcus tephrensis, sp.nov., and Cryptococcus heimaeyensis, sp.nov.; new anamorphic basidiomycetous yeast species from Iceland. AB - Two new species from Iceland are described on the basis of physiological profiles and sequence data from the D2 region of LSU rDNA: Cryptococcus tephrensis (type ICE99-IToM Y5, ATCC MYA-1765, CBS 8935, GenBank AF317208) and Cryptococcus heimaeyensis (type ICE99-IToM Y8, ATCC MYA-1759, CBS 8933, GenBank AF370717). The two new species are identifiable from sequence data and can be distinguished from their closest relative, Cryptococcus victoriae, by their higher maximum temperatures for growth, failure to utilize nitrate as sole nitrogen source, and utilization of cadaverine and ethylamine as sole nitrogen sources. Cryptococcus tephrensis is distinguishable from C. heimaeyensis by failure to grow on saccharate as sole source of carbon and energy. PMID- 12109888 TI - PCR and blot hybridization for rapid identification of Haloferax species. AB - Based on the amplification of a 16S rDNA, a PCR assay for the identification of species of Haloferax to genus level was performed. Two variable regions of the 16S rDNA in Haloferax spp. were selected as genus-specific primers for the PCR assay and hybridization probe. Five genera of halophilic Archaea and Escherichia coli were examined as outside groups. Using this approach, all strains of Haloferax spp. were positive. In contrast, all species belonging to the most closely related genera, including Natrinema, Halorubrum, Halobacterium, and Haloarcula, were negative. In addition, the mass bloom of halophilic Archaea that develops in the El-Mallahet saltern of Alexandria City was positive using the same approach. This assay, which does not require pure cultures of microorganisms, is a specific and rapid method for identifying Haloferax spp. in hypersaline environments. PMID- 12109889 TI - Transposition of pGh9:ISS1 is random and efficient in Streptococcus thermophilus CNRZ368. AB - Streptococcus thermophilus bacteria are used as a starter in the fermentation of yogurts and many cheeses. To construct mutants of S. thermophilus CNRZ368, the use of the plasmid pGh9:ISS1 was considered. This plasmid is known to be a good tool for insertional mutagenesis in gram-positive bacteria, owing to its ability to integrate in the genome by a mechanism of replicative transposition. However, the presence of three endogenous ISS1 copies in the genome of S. thermophilus CNRZ368 and the possible occurrence of homologous recombination could reduce the efficiency of pGh9:ISS1 as a tool for generating mutants. To address this question, the ability of pGh9:ISS1 to transpose randomly in the genome of strain CNRZ368 was investigated. The results of our experiments indicated that: (i) the frequency of transposition of ISS1 was high, approximately 2 x 10(-2), in S. thermophilus CNRZ368; (ii) the integration of multiple tandem copies of the plasmid was frequent; (iii) homologous recombination events between ISS1 were not predominant; and (iv) plasmid pGh9:ISS1 transposed randomly around the S. thermophilus CNRZ368 chromosome. In addition, we describe the strategy used to localize the pGh9:ISS1 insertion locus on the physical map of strain CNRZ368 and the method used to clone the regions flanking this insertion site, especially when multiple copies of the plasmid were integrated in tandem. PMID- 12109891 TI - Stress-like responses to common procedures in male rats housed alone or with other rats. AB - The objective of this study was to assess the cardiovascular function and behavior of male Sprague-Dawley rats housed individually or with one or three cagemates during resting conditions and when subjected to common husbandry and experimental procedures and potentially stressful olfactory stimuli. Heart rate (HR), mean arterial blood pressure (MAP), and movement in the cage were collected by using radiotelemetry for 24 h on an experiment-free day and for 2 h before and 3 h after the following acute procedures: cage change, restraint and subcutaneous injection, restraint and tail-vein injection, exposure to the odor of urine and feces from stressed rats, and exposure to the odor of dried rat blood. Home cage behaviors (sleeping, awake, moving, rearing, and grooming) were scored once each minute for 15 min before and 45 min after the acute procedures. Resting HR and MAP values consistently were lower in rats housed four per cage than animals housed alone or with one cage mate. Compared to that of animals housed individually, general activity was higher during the light phase and lower during the dark phase in rats housed four per cage. Rats housed four per cage showed significantly lower HR and MAP in response to acute husbandry and experimental procedures than rats housed alone, and the HR and MAP of rats housed in pairs were not consistently lower than those of rats housed alone. Procedure-induced arousal behaviors were observed in all housing groups after the acute husbandry and experimental procedures, but rats housed four per cage returned to sleeping behavior more quickly than did rats in the other housing groups. In light of these results, we concluded that under resting conditions, rats housed four per cage were less stressed than were rats housed alone, that common procedures induce noteworthy stress-like responses in male rats, and that the magnitude and duration of these responses are reduced by group housing. PMID- 12109892 TI - Stress-like responses to common procedures in rats: effect of the estrous cycle. AB - The objective of this study was to assess cardiovascular and behavioral responses of Sprague-Dawley female rats subjected to or witnessing common husbandry and experimental procedures at various points during the estrous cycle. Heart rate (HR) and mean arterial blood pressure (MAP) were determined using radiotelemetry for 2 h before and 3 h after: cage change, restraint and subcutaneous (s.c.) injection, restraint and tail-vein injection, witnessing decapitation of other rats, smelling urine and feces from stressed rats, and smelling dried rat blood. Home cage behaviors (sleeping, awake, moving, rearing, and grooming) were scored once each minute for 15 min before and 45 min after the incidents. Being subjected to cage change, restraint and injections, and odors significantly increased HR and MAP 60-120 beats per min and 10-20 mm Hg over baseline respectively for 45 to 90 min. Witnessing these procedures also significantly increased HR and MAP but the magnitude was reduced by 20% to 30% compared to that seen in rats subjected to the procedures. Witnessing decapitation of other rats induced cardiovascular responses which were not different from those of rats witnessing other procedures. The cardiovascular responses were not enhanced during proestrus-estrus compared to metestrus-diestrus. There were also no significant effects of the estrous cycle on home cage behavior after these procedures. We conclude that cycling female rats show stress-like effects when they are subjected to or witness common husbandry and experimental procedures, but there is little to no effect of the estrous cycle. PMID- 12109893 TI - Use of molecular methods for genetic monitoring of an institutional mouse breeding colony. AB - It is important to perform genetic quality control on inbred mouse strains to minimize variability of genetic backgrounds. We used sensitive molecular methods to examine the genetic integrity of inbred mouse substrains maintained at an academic institution. Our goal, in part, was to compare the different molecular genetic monitoring methods to determine which were most sensitive, efficient, and beneficial in our genetic monitoring program. We examined the sensitivity and efficiency of simple sequence length polymorphism (SSLP) analysis of microsatellites and restriction fragment length polymorphism (RFLP) analysis of minisatellites with commercial, human, and synthetic mouse minisatellite probes. Although no polymorphisms were detected with the microsatellite analysis, certain minisatellite probes detected a small degree of polymorphism between our mouse substrains and the commercially available strains used as controls. Minisatellite probes also detected intra-substrain variation within our colonies; this variation probably represents mutations in highly unstable loci rather than genetic variation. Our analysis indicated that the genetic integrity of in-house C57BL/Ka, BALB/cKa, and C3H/Km inbred substrains had remained intact over 35 generations. Genetic monitoring by RFLP minisatellite analysis was more sensitive and efficient in detecting substrain differences than was SSLP microsatellite analysis. On the basis of these results, we established a strategy for future analysis of the in-house breeding colony. PMID- 12109894 TI - Assessment of static isolator cages with automatic watering when used with conventional husbandry techniques as a factor in the transmission of mouse hepatitis virus. AB - This study evaluated protection against mouse hepatitis virus (MHV) afforded by static filter-top caging when automatic watering was used with conventional husbandry techniques as a labor-saving option. We fitted one side of a double sided 72-cage rack with valves external to each cage; cages on the other side were fitted with shielded internal valves. More than 50% of the mice were breeding mice, and 30% were genetically altered. One cage of mice on each shelf on both sides of the rack was infected with MHV-A59. Each row of cages also contained one standard cage (no filter top) of uninoculated mice at various distances from the infected cage. At 2, 4, and 6 weeks after infection of the mice in the test cages, uninoculated mice in 22 cages were tested by serology, and at 8 weeks the uninoculated mice in 54 cages were tested by serology and those in 24 cages were tested by polymerase chain reaction (PCR) amplification of fecal samples to assess transmission of infection. At 8 weeks post-infection, mice in one uninoculated cage (which had a filter top and an internal valve and was adjacent to a cage of inoculated mice) was seropositive. Examination of feces by PCR revealed MHV shedding in mice in nine uninoculated cages (three lacking filter tops but with internal valve cages; two with filter tops and internal valve cages and adjacent to non-filter top cages; two non-filter-top cages with external valves; and two filter-top cages with external valves, of which one was adjacent to a non-filter-top cage). Routine husbandry using either automatic water valve system prevented (with one exception) transmission among filter-top cages for at least 6 weeks. The 10 cages where transmission occurred were non filter-top cages (n = 5) and filter-top cages adjacent to non-filter top, infected, or sentinel cages (n = 5). These results suggest that the use of filter top-caging with automatic watering may limit MHV transmission for 6 weeks, during which immunocompetent mice would be expected to clear the virus. Our findings also suggest that long-term use of automatic watering in static filter-top cages handled using conventional husbandry techniques may not prevent transmission in the vicinity of high virus concentrations or open caging. PMID- 12109895 TI - Herpes B virus-specific pathogen-free breeding colonies of macaques: serologic test results and the B-virus status of the macaque. AB - The demand by the biomedical research community for B virus-free macaques is growing. The availability of B virus-free macaques has never been greater but still falls short of the current demand. Providing resources depends on the definitions of a specific pathogen-free (SPF) macaque, a B virus-free macaque, and a serologically negative macaque. Many colony managers define a B virus-free macaque in light of the serologic status of the breeding colony, because virus identification is a more complex issue. The lack of a standardized definition of a B virus-free macaque is compounded by the fact that all assays inherently result in some false antibody-positive or -negative data, and reconciliation is equivocal. Long-term follow-up of a colony in which elimination of B virus is actively being attempted will necessarily result in a relatively small sample within the population being incorrectly categorized with regard to antibody status. The goal of this study was to determine whether serially collected serologic data demonstrated patterns that could be used to predict whether an animal was genuinely in the process of B virus antibody seroconversion. The best indication of seroconversion was an ELISA titer of > 1:500 or a positive result on a confirmatory test. Patterns of seroreactivity are more useful in identifying B virus-positive macaques during the screening phase of SPF colony development than during the maintenance phase. PMID- 12109896 TI - Laboratory diagnosis of Trypanosoma cruzi infection in a colony-raised pigtailed macaque. AB - An adult (57 months), male, pigtailed macaque (Macaca nemestrina) was screened for an AIDS-related research project and found to be positive on a B-cell lymphocyte culture for trypanosomes. Characterization of the trypanosome was done by using hemocultures, serology, and molecular techniques. Nucleic-acid amplification and sequencing of the nuclear repetitive sequence, kinetoplast DNA, and a fragment of the small subunit RNA gene confirmed that the trypanosome isolated was Trypanosoma cruzi and not T. rangeli, the only other trypanosome known to infect mammals in the New World. This represents the first naturally occurring case of Chagas' disease (American trypanosomiasis) in M. nemestrina, an Old World macaque, and may present an opportunity for development of an animal model of this human disease. The presence of under-recognized but enzootic T. cruzi infection in colony-raised M. nemestrina may have a significant impact on research programs, mandating effective screening of high-risk animals prior to protocol assignment. PMID- 12109897 TI - The effects of uncomplicated miscarriages and stillbirths on the ability of baboons to return to cyclicity and subsequently conceive. AB - Pregnant baboons are used regularly in medical research studies. Occasionally these studies have resulted in stillbirths and/or miscarriages. In addition, pregnant animals can spontaneously undergo stillbirths or miscarriages unrelated to any medical or research procedure. In the absence of identifiable inflammatory, infectious, or pathologic processes, it generally was assumed that these events had no bearing on the baboon's future ability to return to cyclicity and conceive. However, these assumptions were based on observational and anecdotal evidence. To test the validity of these assumptions, we established two data groups: baboons that had uncomplicated stillbirths (Gp-1; n = 11) and those that had uncomplicated miscarriages (Gp-2; n = 12). The mean number of days from first detectable postpartum estrus (i.e., perineal swelling/turgescence) to conception was 49 days for Gp-1 and 53 days for Gp-2. In addition, for Gp-1 animals we determined that the mean number of days to the first indication of estrus was 29 days; these data were unavailable for Gp-2 because of the lack of parturition as a reference point. Control baboons (lactating mothers) required approximately 59 days from first detectable estrus to conception, and our findings for Gp-1 and Gp-2 were consistent with this value. Therefore, within the limits of our study parameters, we suggest that uncomplicated stillbirth and miscarriage had no profound effects on a baboon's future ability to return to cyclicity and conceive. PMID- 12109898 TI - Femoral fracture repair and postoperative management in new zealand white rabbits. AB - Low bone density and large muscle mass predispose rabbits to femoral fractures. However, there are few reports describing treatment and prognosis. Two New Zealand White rabbits presented with unilateral left rear limb abduction and lateral rotation of the distal left rear limb 2 and 17 days after experimental surgery to create a "stair step" in the patellar groove of the left medial femoral chondyle. This procedure was performed after approval by the Institutional Animal Care and Use Committee. Radiography revealed a spiral oblique mid-shaft fracture of the left femur in both rabbits. Open fracture reduction was undertaken. Because of the presence of screws and Kirschner-wires in the medial femoral condyle, a lateral approach to surgical correction was chosen. Intramedullary fixation was used to reduce and stabilize the fractures. A 0.062" Kirschner wire was selected for the intramedullary device, because it was sufficiently flexible to allow easy passage into the femoral canal while being sufficiently stiff to promote reduction of the fracture. In addition, the ends of the fracture were secured with a 0.032" Kirschner cerclage wire to provide additional control of rotation and angulation. Then we assessed the range of motion of the knee joint to determine fracture stability and ensure that the hardware did not impinge on soft-tissue elements. After closure and application of sterile dressing, the hind legs were hobbled proximal to the hock by using elastic veterinary wrap in a figure-eight pattern to maintain limb alignment and prevent formation of pressure ulcers. Intraoperative fluoroscopic evaluation and postoperative radiographs confirmed fracture reduction. Bruising and seroma formation occurred at the surgical site, and transient anorexia developed. Rabbits were treated with fluids, analgesics, antibiotics, and fitted with Elizabethan collars. They were housed in isolation to limit excessive environmental stimulation, which could alarm them and provoke "thumping" of the rear legs. Muscular weakness and atrophy developed in the affected legs, but the fractures remained immobilized. Radiographs obtained 21 days after surgery confirmed marked callus formation and integrity of the implanted hardware. Four weeks after surgical fixation, both rabbits showed increased muscle development in the repaired leg and were ambulating normally. The long-term prognosis was excellent. These cases demonstrate that repair of femoral fractures in rabbits can be achieved by using basic orthopedic techniques and diligent post-operative management. PMID- 12109899 TI - Transmission pattern of parainfluenza 3 virus in guinea pig breeding herds. AB - In searching for the cause of experimental variations in respiratory research data, serology revealed the prevalence of antibodies against parainfluenza virus type 3 (PIV 3) in guinea pigs. The aim of the present study was to explore the transmission rate, course, and kinetics of enzootic PIV 3 infection in guinea pig breeding units. In the first part of the study, blood samples to be analyzed for PIV 3 antibodies were collected from guinea pigs of a PIV 3-positive breeding colony at different times after birth. In the same breeding unit, 6 of 12 2-week old guinea pigs were relocated and separately housed. The PIV 3 serum antibody titers of the two groups were compared at various times from birth to 13 weeks after birth. In the second part of the study, the spread of infectious virus and virus persistence were explored by housing seronegative sentinel animals together with 2- to 3-week-old guinea pigs from three different PIV 3-positive breeding units. The guinea pigs remaining in the breeding colony as well as those removed and housed separately showed declining serum antibody titers for about 1 month after birth, thereafter the titers were stable until about 8 weeks after birth. Five weeks later, the mean antibody titer of the guinea pigs remaining in the breeding colony had increased to a markedly higher level than that of the relocated, separately housed guinea pigs. Seroconversion was demonstrated in 7 of the 14 sentinels housed with the 2- to 3-week-old guinea pigs from PIV 3-positive breeding units. Sentinels housed together with PIV 3-positive guinea pigs 24 weeks after the start of the experiment did not seroconvert. We conclude that young guinea pigs born to PIV 3-positive mothers were protected by maternal immunity against infection with PIV 3 during their first 14 days of life. The guinea pig offspring became infected during the period from about 2 weeks until 8 weeks after birth, as demonstrated by seroconversion of sentinel animals and an increasing mean antibody titer seen beyond 8 weeks of age. The study did not reveal any indication of virus persistence or prolonged carrier status. PMID- 12109900 TI - Behavioral management: it's everyone's job. AB - In this day and age of regulatory demands and with the ever-increasing flow of environmental enhancement data and opinions, it can become very confusing for animal programs to create and maintain a successful behavioral management program. Behavioral management as a concept provides a common ground from which animal facilities may start to build a successful facility behavioral program. In addition, the implementation of a behavioral management program can help to break down barriers between members of the various disciplines within the biomedical community. It is everyone's responsibility and duty to work together with this common goal: to provide the best care and environment we can for the animals in our charge. Working together to improve animal behavior can help us to achieve this goal. PMID- 12109901 TI - SAR studies of piperidine-based analogues of cocaine. 4. Effect of N-modification and ester replacement. AB - A series of novel N- and 3alpha-modified piperidine-based analogues of cocaine were synthesized and tested for their ability to inhibit reuptake of DA, 5-HT, and NE by the DA, 5-HT, and NE transporters. N-Demethylation of trans-(+)-3alpha piperidine-based ligands leads to improved activity at the SERT and NET and modest changes at the DAT. Replacement of the N-methyl group in trans-(+)-ester 1a with phenylalkyl groups leads to a modest 2.3-fold improvement in activity at the SERT (K(i) < or = 3.27 microM), insignificant changes at the NET, and a 3.5 fold loss in activity at the DAT (K(i) > or = 810 nM); however, such replacement in cis-(-)-ester 4, the more potent isomer of 1a, leads, in general, to a significant decrease in activity at all monoamine transporters (K(i) > 1 microM). Other N-modified ligands, including the ligands with polar groups incorporated in the N-alkyl substituent (3e-g) and ligands lacking the basic nitrogen (3i and 6d), show decreased activity at all monoamine transporters, though ligands 3e-g are similar in potency at the NET to 1a. N-Norester 2a, a possible metabolite of the lead compound 1a, and alcohol 1c, a compound with a 3alpha-substituent that is more stable to metabolism than 1a, were selected for further behavioral tests in animals. Alcohol 1c and ester 2a are similar in potency at the DAT to cocaine, ester 1a, and oxadiazole 1b, and both fully substitute for cocaine and have potency similar to that of cocaine in drug discrimination tests. Like cocaine, 1c increased locomotor activity (LMA) monotonically with time, whereas 2a produces biphasic effects consisting of initial locomotor depression followed by delayed locomotor stimulation. An interesting difference between cocaine, ester 1a, alcohol 1c, and N-norester 2a is that 1c and 2a are significantly longer acting in LMA tests. Although this result was anticipated for alcohol 1c, it is rather surprising for 2a which has an ester function susceptible to hydrolysis, a pathway of in vivo deactivation of cocaine and its ester analogues. The present results may have important implications for our understanding of the pharmacological mechanisms underlying the behavioral actions of cocaine and of the structural features needed for the design of the new leads in the discovery of a cocaine abuse medication. PMID- 12109902 TI - Common and selective molecular determinants involved in metabotopic glutamate receptor agonist activity. AB - Several potent and group selective agonists of metabotropic glutamate receptors (mGluRs) have been docked at mGlu1,2,4R binding sites in the closed conformation of the bilobate extracellular domain. Quisqualic acid and (S)-3,5 dihydroxyphenylglycine (3,5-DHPG) were selected for mGlu1R, dicarboxycyclopropylglycine (DCG-IV), LY354740, (S)-4-carboxyphenylglycine (4CPG) for mGlu2R, and (S)-2-amino-4-phosphonobutyric acid (AP4), 1-aminocyclopentane 1,3,4-tricarboxylic acid (ACPT-I), (S)-4-phosphonophenylglycine (PPG) for mGlu4R. The models show a conserved binding pattern for the glycine moiety (alpha-amino and alpha-acidic functions) and group specific bindings for the distal acidic function. The best agonists allow optimized interaction with both lobes of the binding domain. Interlobe connections around the ligand are also described and participate in stabilizing the closed form of the amino-terminal domain. Altogether, the docking models support the proposal that the stabilization of a closed state represents a key step in agonist activation of mGluRs. PMID- 12109903 TI - Structure activity of 3-aryl-1,3-diketo-containing compounds as HIV-1 integrase inhibitors. AB - The 4-aryl-2-hydroxy-4-oxo-2-butenoic acids and their isosteric tetrazoles are among an emerging class of aryl beta-diketo (ADK)-based agents which exhibit potent inhibition of HIV-1 integrase (IN)-catalyzed strand transfer (ST) processes, while having much reduced potencies against 3'-processing (3'-P) reactions. In the current study, L-708,906 (10e) and 5CITEP (13b), which are two examples of ADK inhibitors that have been reported by Merck and Shionogi pharmaceutical companies, served as model ADK leads. Structural variations to both the "left" and "right" sides of these molecules were made in order to examine effects on HIV-1 integrase inhibitory potencies. It was found that a variety of groups could be introduced onto the left side aryl ring with maintenance of good ST inhibitory potency. However, introduction of carboxylic acid-containing substituents onto the left side aryl ring enhanced 3'-P inhibitory potency and reduced selectivity toward ST reactions. Although both L 708,906 and 5CITEP show potent inhibition of IN in biochemical assays, there is a disparity of antiviral activity in cellular assays using HIV-1-infected cells. Neither 5CITEP nor any other of the indolyl-containing inhibitors exhibit significant antiviral effects in cellular systems. Alternatively, consistent with literature reports, L-708,906 does provide antiviral protection at low micromolar concentrations. Interestingly, several analogues of L-708,906 with varied substituents on the left side aryl ring, while having good inhibitory potencies against IN in extracellular assays, are not antiviral in whole-cell systems. PMID- 12109904 TI - Synthesis and effects on chloroquine susceptibility in Plasmodium falciparum of a series of new dihydroanthracene derivatives. AB - To suggest a mechanism of action for drugs capable to reverse the chloroquine resistance, a new set of 9,10-dihydro-9,10-ethano and ethenoanthracene derivatives was synthesized and compounds were tested with the aim to assess their effect on chloroquine susceptibility in Plasmodium falciparum resistant strains. With respect to this, reversal of resistance and change in drug accumulation were compared. Structure-activity relationship and molecular modeling studies made it possible to define a pharmacophoric moiety for reversal agents and to propose a putative model of interaction with some selected amino acids. PMID- 12109905 TI - 4D-QSAR analysis of a set of propofol analogues: mapping binding sites for an anesthetic phenol on the GABA(A) receptor. AB - A training set of 27 propofol (2,6-diisopropylphenol) analogues was used to construct four-dimensional (4D) quantitative structure-activity relationship (QSAR) models for three screens of biological activity: loss of righting reflex (LORR) in tadpoles, enhancement of agonist activity at the gamma-aminobutyric acid type A (GABA(A)) receptor, and direct (agonist-independent) activation of the receptor. The three resulting 4D-QSAR models are almost identical in form, and all suggest three key ligand-receptor interaction sites. The formation of an intermolecular hydrogen bond involving the proton of the ligand -OH group is the most important binding interaction. A hydrophobic pocket binding interaction involving the six-substituent is the second most significant binding site, and a similar hydrophobic pocket binding interaction near the two-substituent is the third postulated binding site from the 4D-QSAR models. A test set of eight compounds was used to evaluate the tadpole LORR 4D-QSAR model. Those compounds highly congeneric to the training set compounds were accurately predicted. However, compounds exploring substituent sites and/or electronic structures different from the training set were less well-predicted. Overall, the results show a striking similarity between the models of the sites responsible for anesthesia and those mediating effects of the training set of propofol analogues on the GABA(A) receptor; it follows that the GABA(A) receptor is therefore the likely site of propofol's anesthetic action. PMID- 12109906 TI - Structure-based approach for binding site identification on AmpC beta-lactamase. AB - Beta-lactamases are the most widespread resistance mechanism to beta-lactam antibiotics and are an increasing menace to public health. Several beta-lactamase structures have been determined, making this enzyme an attractive target for structure-based drug design. To facilitate inhibitor design for the class C beta lactamase AmpC, binding site "hot spots" on the enzyme were identified using experimental and computational approaches. Experimentally, X-ray crystal structures of AmpC in complexes with four boronic acid inhibitors and a higher resolution (1.72 A) native apo structure were determined. Along with previously determined structures of AmpC in complexes with five other boronic acid inhibitors and four beta-lactams, consensus binding sites were identified. Computationally, the programs GRID, MCSS, and X-SITE were used to predict potential binding site hot spots on AmpC. Several consensus binding sites were identified from the crystal structures. An amide recognition site was identified by the interaction between the carbonyl oxygen in the R1 side chain of beta lactams and the atom Ndelta2 of the conserved Asn152. Surprisingly, this site also recognizes the aryl rings of arylboronic acids, appearing to form quadrupole dipole interactions with Asn152. The highly conserved "oxyanion" hole defines a site that recognizes both carbonyl and hydroxyl groups. A hydroxyl binding site was identified by the O2 hydroxyl in the boronic acids, which hydrogen bonds with Tyr150 and a conserved water. A hydrophobic site is formed by Leu119 and Leu293. A carboxylate binding site was identified by the ubiquitous C3(4) carboxylate of the beta-lactams, which interacts with Asn346 and Arg349. Four water sites were identified by ordered waters observed in most of the structures; these waters form extensive hydrogen-bonding networks with AmpC and occasionally the ligand. Predictions by the computational programs showed some correlation with the experimentally observed binding sites. Several sites were not predicted, but novel binding sites were suggested. Taken together, a map of binding site hot spots found on AmpC, along with information on the functionality recognized at each site, was constructed. This map may be useful for structure-based inhibitor design against AmpC. PMID- 12109907 TI - Synthesis and pharmacological characterization of novel analogues of the nicotinic acetylcholine receptor agonist (+/-)-UB-165. AB - (+/-)-UB-165 (1) is a potent neuronal nicotinic acetylcholine receptor (nAChR) ligand, which displays functional selectivity between nAChR subtypes. Using UB 165 as a lead structure, two classes of racemic ligands were synthesized and assessed in binding assays for three major nAChR subtypes (alpha4beta2, alpha3beta4, and alpha7). The first class of compounds comprises the three pyridine isomers 4-6, corresponding to the 3-, 2-, and 4-substituted pyridine isomers, respectively. Deschloro UB-165 (4) displayed a 2-3-fold decrease in affinity at alpha4beta2 and alpha3beta4 nAChR subtypes, as compared with (+/-)-UB 165, while at the alpha7 subtype a 31-fold increase in affinity was observed. At each of the nAChR subtypes, high affinity binding was dependent on the presence of a 3-substituted pyridine, and the other isomers, 5 and 6, resulted in marked decreases in binding affinities. The second class of compounds is based on replacing the pyridyl unit of 1 with a diazine moiety, giving pyridazine (7), pyrimidine (8), and pyrazine (9), which retain the "3-pyridyl" substructure. Modest reductions in binding affinity were observed for all of the diazine ligands at all nAChR subtypes, with the exception of 7, which retained potency comparable to that of 4 in binding to alpha7 nAChR. In functional assays at the alpha3beta4 nAChR, all analogues 4-9 were less potent, as compared with 1, and the rank order of functional potencies correlated with that of binding potencies. Computational studies indicate that the 3-substituted pyridine 4 and 2 substituted pyridine 5, as well as the diazine analogues 7-9, all conform to a distance-based pharmacophore model recently proposed for the alpha4beta2 receptor. However, the nicotinic potencies of these ligands vary considerably and because 5 lacks appreciable nicotinic activity, it is clear that further refinements of this model are necessary in order to describe adequately the structural and electronic demands associated with this nAChR subtype. This rational series of compounds based on UB-165 presents a systematic approach to defining subtype specific pharmacophores. PMID- 12109908 TI - Discovery of aminopyridine-based inhibitors of bacterial enoyl-ACP reductase (FabI). AB - Bacterial enoyl-ACP reductase (FabI) catalyzes the final step in each cycle of bacterial fatty acid biosynthesis and is an attractive target for the development of new antibacterial agents. Our efforts to identify potent, selective FabI inhibitors began with screening of the GlaxoSmithKline proprietary compound collection, which identified several small-molecule inhibitors of Staphylococcus aureus FabI. Through a combination of iterative medicinal chemistry and X-ray crystal structure based design, one of these leads was developed into the novel aminopyridine derivative 9, a low micromolar inhibitor of FabI from S. aureus (IC(50) = 2.4 microM) and Haemophilus influenzae (IC(50) = 4.2 microM). Compound 9 has good in vitro antibacterial activity against several organisms, including S. aureus (MIC = 0.5 microg/mL), and is effective in vivo in a S. aureus groin abscess infection model in rats. Through FabI overexpressor and macromolecular synthesis studies, the mode of action of 9 has been confirmed to be inhibition of fatty acid biosynthesis via inhibition of FabI. Taken together, these results support FabI as a valid antibacterial target and demonstrate the potential of small-molecule FabI inhibitors for the treatment of bacterial infections. PMID- 12109909 TI - Three-dimensional quantitative structure-activity relationship analysis of ligand binding to human sequence antidigoxin monoclonal antibodies using comparative molecular field analysis. AB - The present study indicates that the newly generated human sequence antidigoxin monoclonal antibody (mAb), 1B3, binds digoxin with a different fine specificity binding than our previously obtained human sequence monoclonal antibodies (mAbs) (Ball, W. J.; et al. J. Immunol. 1999, 163, 2291-2298). Uniquely, 1B3 has a higher affinity for digitoxin than digoxin, the immunizing hapten, and a strong requirement for at least one sugar residue linked to the aglycone (-genin). By means of comparative molecular field analysis (CoMFA), the results of competition binding studies for 56 cardiotonic and hormonal steroids were employed to develop three-dimensional quantitative structure-activity relationship (3D-QSAR) models for ligand binding to 1B3 and to three additional human sequence mAbs, as well as the murine antidigoxin mAb 40-50 (Mudgett-Hunter, M.; et al. Mol. Immunol. 1985, 22, 447-488). All five 3D-QSAR models yielded cross-validated q(2) values greater than 0.5, which indicates that they have significant predictive ability. The CoMFA StDevCoeff contour plots, as well as the competition results, indicate that 1B3 binds ligands in a manner distinct from the other four mAbs. The CoMFA contour plots for 40-50 were also compared with the known X-ray crystallographic structure of the 40-50-ouabain complex (Jeffrey, P. D.; et al. J. Mol. Biol. 1995, 248, 344-360) in order to identify correlations between residues in the mAb binding site and specific contour plot regions. These 3D-QSAR models and their respective contour plots should be useful tools to further understand the molecular nature of antibody-antigen interactions and to aid in the redesign or enhancement of therapeutic antibodies. PMID- 12109910 TI - N(6)-alkyl-2-alkynyl derivatives of adenosine as potent and selective agonists at the human adenosine A(3) receptor and a starting point for searching A(2B) ligands. AB - A series of N(6)-alkyl-2-alkynyl derivatives of adenosine (Ado) have been synthesized and evaluated for their affinity at human A(1), A(2A), and A(3) receptors and for their potency at A(2B) adenosine receptor subtypes. The corresponding 2-(1-alkynyl) derivatives of 5'-N-ethylcarboxamidoadenosine (NECA) and Ado are used as reference compounds. Binding studies demonstrated that the activities of 2-alkynylAdos were slightly increased for the adenosine A(1) receptor and slightly decreased for both A(3) and A(2B) subtypes compared to those of their corresponding NECA derivatives, whereas the A(2A) receptor affinities of the two series of nucleosides were similar. The presence of a methyl group on N(6) of the 2-alkynyladenosines, inducing an increase in affinity at the human A(3) receptor and a decrease at the other subtypes, resulted in an increase in A(3) selectivity. In particular, 2-phenylethynyl-N(6)-methylAdo (8b) showed an A(3) affinity in the low nanomolar range (K(i)(A(3)) = 3.4 nM), with a A(1)/A(3) and A(2A)/A(3) selectivity of about 500 and 2500, respectively. These findings motivated us to search for the preparation of new selective radioligands for the A(3) subtype; hence, a procedure to introduce a tritiated alkylamino group in these molecules was carried out. As far as the potency at the A(2B) receptor, the type of 2-alkynyl chain and the presence of the ethylcarboxamido group on the sugar seem to be very important; in fact, the (S)-2 phenylhydroxypropynylNECA [(S)-PHPNECA, 1e, EC(50)(A(2B)) = 0.22 microM] proved to be one of the most potent A(2B) agonist reported so far. On the other hand, the (S)-2-phenylhydroxypropynyl-N(6)-ethylAdo (9e, EC(50)(A(2B)) = 0.73 microM) showed a significantly increase of potency at the A(2B) subtype in comparison with the N(6)-methyl, N(6)-isopropyl, and the unsubstituted adenosine derivatives, although it resulted in being less potent than (S)-PHPNECA (1e, EC(50)(A(2B)) = 0.22 microM). These observations suggest that the introduction of an ethyl group in the N(6)-position and an ethylcarboxamido substituent in the 4' position of (S)-2-phenylhydroxypropynyladenosine could lead to a compound endowed with high potency at the A(2B) receptor. PMID- 12109911 TI - Synthesis and pharmacological testing of 1,2,3,4,10,14b-hexahydro-6-methoxy-2 methyldibenzo[c,f]pyrazino[1,2-a]azepin and its enantiomers in comparison with the two antidepressants mianserin and mirtazapine. AB - The synthesis and resolution of 1,2,3,4,10,14b-hexahydro-6-methoxy-2 methyldibenzo[c,f]pyrazino[1,2-a]azepin (6-methoxymianserin, 6) are described. Furthermore, the in vitro and in vivo effects of 6 and its enantiomers are presented. 6 displayed high affinity for the 5-HT2A/2C receptors, only moderate affinity for the adrenoceptors, and no affinity for the NA reuptake site. Surprisingly, 6 also showed moderate to high affinity for the dopamine D2 receptor, an effect that resides in the (R)-(-)-enantiomer. PMID- 12109912 TI - Structure-activity relationships of methoctramine-related polyamines as muscular nicotinic receptor noncompetitive antagonists. 3. Effect of inserting the tetraamine backbone into a macrocyclic structure. AB - The present article expands on the study of another aspect of structure-activity relationships of the polymethylene tetraamines, namely, the effect of inserting the tetraamine backbone into a macrocyclic structure. To this end, compounds 8-12 were designed by linking the two terminal nitrogen atoms of prototype methoctramine 2 to an aryl moiety. Alternatively, 2 was first modified to achieve compounds 6 and 7, which in turn were cyclized by linking the two terminal primary amine functions to a polyphenyl spacer, affording 13-20. All the compounds were tested on muscle-type nAChRs and most of them as well on AChE. Furthermore, selected compounds were tested also on peripheral M(2) and M(3) mAChRs. All these cyclic derivatives, like prototypes, were potent noncompetitive antagonists at both frog and Torpedo nAChRs, suggesting that polyamines do not need to be linear or in extended conformation to optimally interact with the nicotinic channel; rather, they may bind in a U-shaped conformation. Relative to muscarinic activity, macrocyclic compounds 10, 13, 14, and 20, in contrast with the profile displayed by 2, were almost devoid of affinity. It is derived that an aryl spacer is detrimental to the interaction of polyamines with mAChRs. Finally, all the diamine diamides investigated in this study were much less potent in inhibiting AChE activity than prototype 3, suggesting that a macrocyclic structure may not be suitable for AChE inhibition. PMID- 12109914 TI - Rational design and synthesis of a novel thyroid hormone antagonist that blocks coactivator recruitment. AB - Recent efforts have focused on the design and synthesis of thyroid hormone (T(3)) antagonists as potential therapeutic agents and chemical probes to understand hormone-signaling pathways. We previously reported the development of novel first generation T(3) antagonists DIBRT, HY-4, and GC-14 using the "extension hypothesis" as a general guideline in hormone antagonist design.(1-3) These compounds contain extensions at the 5'-position (DIBRT, GC-14) of the outer thyronine ring or from the bridging carbon (HY-4). All of these compounds have only a modest affinity and potency for the thyroid hormone receptor (TR) that limits studies of their antagonistic actions. Here, we report the design and synthesis of a novel series of 5'-phenylethynyl derivatives sharing the GC-1 halogen-free thyronine scaffold.(4) One compound (NH-3) is a T(3) antagonist with negligible TR agonist activity and improved TR binding affinity and potency that allow for further characterization of its observed activity. One mechanism for antagonism appears to be the ability of NH-3 to block TR-coactivator interactions. NH-3 will be a useful pharmacological tool for further study of T(3) signaling and TR function. PMID- 12109913 TI - Structure-activity relationships on phenylalanine-containing inhibitors of histone deacetylase: in vitro enzyme inhibition, induction of differentiation, and inhibition of proliferation in Friend leukemic cells. AB - Inhibitors of histone deacetylases (HDACs) are a new class of anticancer agents that affect gene regulation. We had previously reported the first simple synthetic HDAC inhibitors with in vitro activity at submicromolar concentrations. Here, we present structure-activity data on modifications of a phenylalanine containing lead compound including amino acid amides as well as variations of the amino acid part. The compounds were tested for inhibition of maize HD-2, rat liver HDAC, and for the induction of terminal cell differentiation and inhibition of proliferation in Friend leukemic cells. In the amide series, in vitro inhibition was potentiated up to 15-fold, but the potential to induce cell differentiation decreased. Interestingly, an HDAC class selectivity was indicated among some of these amides. In the amino acid methyl ester series, a biphenylalanine derivative was identified as a good enzyme inhibitor, which blocks proliferation in the submicromolar range and is also a potent inducer of terminal cell differentiation. PMID- 12109915 TI - Synthesis and antiviral activity of new anti-HIV amprenavir bioisosteres. AB - Starting from the chemical structure of the recent FDA-approved anti-HIV drug Amprenavir (Agenerase), a potent HIV-protease inhibitor, we have designed new series of Amprenavir bioisoteres in which the methylene group of the benzyl group was replaced by a sulfur atom. This structural modification has required an original multistep synthesis. Unfortunately, introduction of the sulfur atom abolished or drastically decreased both inhibitory activity on recombinant HIV protease and HIV infection protection on MT4 cell cultures. PMID- 12109916 TI - Methods for claims-based pharmacoeconomic studies in psychosis. AB - Pharmacoeconomic studies using claims data are frequently employed to compare the healthcare costs associated with competing drugs. Different methodological approaches with varying limitations for evaluating claims data are reviewed within the context of psychosis. Intent-to-treat paradigms and mirror-image designs have limitations that can bias comparisons of health resource use and can be addressed through the use of drug treatment episodes. Health resource use is better measured in financial rather than volume amounts, to account for service intensity. However, financial measures reported in claims records need to be carefully chosen. Between-group comparisons are frequently affected by selection bias, and within-group comparisons are often limited by period bias. While selection bias can be addressed by controlling for patient factors such as disease severity, and period bias by controlling for trend, devising appropriate control measures from the limited information in claims data can be challenging. Healthcare data are often skewed, which affects statistical testing. While skewed data are commonly handled through log transformation, this method has serious limitations, potentially distorting pharmacoeconomic comparisons. Determining the most appropriate methods for claims-based data involves careful evaluation of the alternate approaches to best achieve the goals of a pharmacoeconomic investigation. PMID- 12109918 TI - Ready-to-use injection preparations versus conventional reconstituted admixtures: economic evaluation in a real-life setting. AB - OBJECTIVE: To measure, in a real-life setting, the benefits of using ready-to-use (RTU) injection preparations compared with conventional reconstituted admixtures (Admix) in terms of cost savings. DESIGN AND PERSPECTIVE: An economic model was developed, based on a randomised study. The perspective of the economic evaluation was that of the hospital administration. A microcosting approach was used to determine costs. SETTING: Department of Cardiac Surgery at the Charleroi University Hospital in Belgium. STUDY PARTICIPANTS: Fifty-eight patients undergoing cardiac surgery under cardiopulmonary bypass were randomised to Admix dobutamine or to the RTU dobutamine group and were followed up during 24 hours after initiation of dobutamine therapy. MAIN OUTCOME MEASURES AND RESULTS: Nursing time was reduced by 32% in the RTU group compared with the Admix group. Material cost was also reduced and the overall cost savings in the RTU group amounted to a 60% reduction in the cost of the conventional Admix process (p<0.001). When drug cost was included in the equation, cost savings varied from 1.60 euros (EUR) to EUR21.40 per patient depending on dosage. There was no difference between the two groups in terms of safety and efficacy. A user satisfaction survey showed that medical staff especially welcomed improved ease of preparation and potential for prevention of errors and risks of handling. CONCLUSION: This study confirmed the potential for RTU forms to reduce nursing time associated with preparation and administration of intravenous admixtures and to enable overall cost savings. PMID- 12109917 TI - Cost-effective approaches to the treatment of community-acquired pneumonia in the era of resistance. AB - Community-acquired pneumonia (CAP) infects upwards of four million people in the US each year, of which 20% require subsequent hospitalisation. Consequently, it is a large contributor to excessive healthcare resource consumption and cost. Since the aetiology of CAP is not identified in a majority of patients, treatment is often empiric, aimed at the most common causes, Streptococcus pneumoniae, Haemophilus influenzae, Moraxella catarrhalis, and the atypical pathogens (Mycoplasma pneumoniae, Chlamydia pneumoniae and Legionella pneumophila). A variety of pharmaceutical agents exist for the treatment of CAP, most notably the cephalosporin and penicillin derivatives, the macrolide/azalide antibacterials, the newer tetracyclines, and most recently the respiratory fluoroquinolones. Choosing an agent is usually related to issues such as patient compliance, adverse event profiles, and the presence of resistance. Of these, resistance seems to be the main factor today. S. pneumoniae, the most common cause of CAP, is steadily acquiring resistance to a majority of the currently available antibacterials, thus further increasing costs due to prolonged hospitalisation, treatment of relapses and the use of more expensive antibacterials. Understanding and maximising the pharmacodynamic properties of the available antibacterials will not only prevent the emergence of resistance, thus prolonging their clinical utility, but also reduce the costs associated with treating the infection through rapid symptom improvement and earlier patient discharge. Numerous methods for reducing costs in patients with bacterial infections are documented in the literature and can be applied to CAP. Choosing monotherapy instead of combination therapy can reduce costs associated with the administration of the antibacterial. Agents with longer half-lives allow for once-daily administration, which in turn, leads to improved compliance, successful outcomes, and decreased costs. Administering antibacterials to maximise their pharmacodynamics, such as with continuous infusion of beta-lactams, reduces the amount of drug needed in addition to savings associated with administration and supplies. Finally, transitioning patients to oral therapy as soon as they are clinically stable can significantly reduce the length of hospital stay, which is the major contributing factor of healthcare costs. The use of a clinical pathway in an institution is the most effective way to apply these cost-saving approaches in the treatment of CAP. These pathways should be specific to each institution, thus considering the resistance rates in the area and encouraging the use of the most active, cost effective agents to produce rapid, positive clinical outcomes. PMID- 12109919 TI - Stroke rehabilitation services to accelerate hospital discharge and provide home based care: an overview and cost analysis. AB - BACKGROUND: Limited information exists on the best way to organise stroke rehabilitation after hospital discharge and the relative costs of such services. OBJECTIVE: To review the evidence of the cost effectiveness of services that accelerate hospital discharge and provide home-based rehabilitation for patients with acute stroke. METHODS: A systematic review with economic analysis of published randomised clinical trials (available to March 2001) comparing early hospital discharge and domiciliary rehabilitation with usual care in patients with stroke was conducted. From included studies, data were extracted on study quality; major clinical outcomes including hospital stay, death, institutionalisation, disability, and readmission rates; and resource use associated with hospital stay, rehabilitation, and community services. The resources were priced using Australian dollars ($A) healthcare costs. The outcomes and costs of the new intervention were compared with standard care. RESULTS: Seven published trials involving 1277 patients (54% men; mean age 73 years) were identified. The pooled data showed that overall, a policy of early hospital discharge and domiciliary rehabilitation reduced total length of stay by 13 days [95% confidence interval (CI): -19 to -7 days]. There was no significant effect on mortality (odds ratio = 0.95; 95% CI: 0.65 to 1.38) or other clinical outcomes making a cost minimisation analysis for the economic analysis appropriate. The overall mean costs were approximately 15% lower for the early discharge intervention [$A16 016 ($US9941) versus $A18 350] ($US11 390)] compared with standard care. CONCLUSIONS: A policy of early hospital discharge and home based rehabilitation for patients with stroke may reduce the use of hospital beds without compromising clinical outcomes. Our analysis shows this service to be a cost saving alternative to conventional in-hospital stroke rehabilitation for an important subgroup of patients with stroke-related disability. PMID- 12109920 TI - Economic benefits of amlodipine treatment in patients with coronary artery disease. AB - OBJECTIVE: To estimate the potential savings in overall cardiovascular disease (CVD) treatment costs for the US population with coronary artery disease (CAD) resulting from the use of amlodipine. STUDY DESIGN AND METHODS: Using patient level data from a retrospective analysis of the Prospective Evaluation of the Vascular Effects of Norvasc Trial (PREVENT), a randomised, placebo-controlled clinical trial (n = 825), we constructed a Markov cohort simulation model to estimate the health economic outcomes of patients with CAD treated with either amlodipine or placebo. PERSPECTIVE: Healthcare payer perspective. RESULTS: The expected number of CVD events for amlodipine recipients was significantly lower than the number of CVD events in the placebo cohort (p < 0.01). The net present value of the cost per patient for CVD treatment was estimated to be $US14 117 for amlodipine recipients and $US16 683 (1999 values, assuming a 3% discount rate) for placebo recipients over 3 years of follow-up with cost savings realised in the amlodipine cohorts after 6 months. CONCLUSIONS: According to the model, amlodipine results in an expected per patient cost savings of $US2566 over a 3 year period, mainly due to a reduction in hospitalisations for cardiovascular related events and procedures. PMID- 12109921 TI - Gene therapy for restenosis: current status. AB - Atherosclerosis is a major cause of morbidity and mortality in Western world. Vascular occlusion caused by atherosclerosis usually requires invasive treatment, such as surgical bypass or angioplasty. However, bypass graft failure and restenosis limit the usefulness of these procedures, with 20% of patients needing a new revascularisation procedure within 6 months of angioplasty. Numerous pharmacological agents have been investigated for the prevention of restenosis but none has shown undisputed efficacy in clinical medicine. Gene transfer offers a novel approach to the treatment of restenosis because of easy accessibility of vessels and already existing gene delivery methods. It can be used to overexpress therapeutically important proteins locally without high systemic toxicity, and the therapeutic effect can be targeted to a particular pathophysiological event. Promising results have been obtained from many pre-clinical experiments using therapeutic genes or oligonucleotides to prevent restenosis. Early clinical trials have shown that plasmid- and adenovirus-mediated vascular gene transfers can be conducted safely and are well tolerated. Ex vivo gene therapy with E2F decoy succeeded in reducing graft occlusion rate after surgical bypass in a randomised, double-blind clinical trial. In the future, further development of gene delivery methods and vectors is needed to improve the efficacy and safety of gene therapy. Also, better knowledge of vascular biology at the molecular level is needed to find optimal strategies and gene combinations to treat restenosis. Provided that these difficulties can be solved, gene therapy offers an enormous potential for clinical medicine in the future. PMID- 12109922 TI - Single-agent immunosuppression after liver transplantation: what is possible? AB - Orthotopic liver transplantation is a life saving and life enhancing procedure. The development of immunosuppressive drugs has contributed to the high rate of success in terms of both patient and graft survival. However, the considerable adverse effects of these therapies are affecting long-term outcomes of transplant recipients. Complications related to immunosuppression are responsible for the majority of deaths in patients surviving more than 1 year. Therefore, the search for an optimal immunosuppressive regimen has become of paramount importance. The liver has proved to be an 'immunologically privileged' organ, capable in several animal models to be accepted as an allograft without any intervention on the immune system of the recipient. In some human liver allografts acceptance of the new organ is recognised after withdrawal of immunosuppressants, but prior identification of such individuals is not yet possible, thus negating this management option. Graft-recipient interaction is peculiar in liver transplantation: acute cellular rejection does not always need to be treated, and if it is not severe, appears to be associated with a better survival of both patient and graft. In the last decade there has been an evolution of immunosuppressive protocols, driven by empirical observation and a deeper understanding of immunological events after transplant. However, most modifications have been made because of the necessity to reduce long-term drug related morbidity and mortality. Withdrawal of corticosteroids has proven to be safely achievable in most patients, with no deleterious effects on patient or graft survival but with a great benefit in terms of reduction of incidence of metabolic and cardiovascular complications. Long-term 'steroid-free' regimens are therefore now widely used. Patients with stable graft function can be easily maintained using a single drug usually after 6 or 12 months and usually with a calcineurin inhibitor. The more evolved step of using monotherapy ab initio has also proven to be effective in a few studies and needs to be explored further. In the future new strategies will be designed to help the development of tolerance of the allograft, selectively stimulating instead of suppressing the immune reaction of the recipient. PMID- 12109923 TI - Cardiovascular risk profile of antirheumatic agents in patients with osteoarthritis and rheumatoid arthritis. AB - Several new drugs have become available for the treatment of patients with osteoarthritis and rheumatoid arthritis (RA). These agents include selective cyclooxygenase (COX)-2 inhibitors, leflunomide and anti-tumour necrosis factor (TNF)-alpha antagonists. COX-2 inhibitors have a more favourable gastrointestinal adverse effect profile than conventional non-steroidal anti-inflammatory drugs (NSAIDs). However, the COX-2 inhibitors are also associated with hypertension, oedema and congestive heart failure, the well-known adverse effects of conventional NSAIDs. Patients with treated hypertension should be monitored regularly when conventional NSAIDs or COX-2 inhibitors are administered. At present, there is a considerable debate regarding the risk of cardiovascular events with the COX-2 inhibitors. The available literature gives no unequivocal answers. This matter can only be solved by an appropriate trial assessing the cardiovascular risk of these agents. Patients with RA appear to have an enhanced cardiovascular risk which might be related to an unfavourable lipid profile. Corticosteroids induce hypercholesterolaemia in patients other than those with RA. It was recently shown that total and high-density lipoprotein (HDL) cholesterol were low in patients with RA who had a high disease activity. Contrary to the expectation, combination therapy with prednisolone rapidly improved the atherogenic index (total/HDL cholesterol). Ongoing studies investigating this topic are underway. It is not known to what extent corticosteroids induce hypertension in patients with RA. Hence, we advocate blood pressure control for these patients. A small percentage of patients with RA develop hypertension when taking leflunomide, and no other serious cardiovascular adverse effects have been reported in the literature. Blood pressure monitoring is recommended especially in the first months of treatment. TNFalpha antagonists are contraindicated in patients with RA who have congestive heart failure. No specific cardiovascular adverse effects have been reported with the use of these agents in the non-cardiovascular compromised patient. TNFalpha antagonists are the most powerful anti-inflammatory drugs presently available. As inflammation plays an important role in RA as well as in cardiovascular disease and, in view of the increased cardiovascular risk in RA, it is tempting to expect that suppression of inflammation ultimately will lower the cardiovascular morbidity and mortality in patients with RA. PMID- 12109925 TI - Generalised anxiety disorder: treatment options. AB - In recent years generalised anxiety disorder (GAD) has become a much better defined disorder, with specific criteria distinguishing it from the other anxiety disorders; however, it still lacks the same public and scientific interests as some of the other anxiety disorders such as panic and social phobia. Nevertheless, refinement in the treatment of GAD is becoming more evident through the conduct of clinical trials. Up until the mid-1980's, treatment consisted primarily of benzodiazepines. However, as a result of growing characterisation of their abuse potential, other therapeutic options were explored. Benzodiazepines became seen as an effective short-term therapy, and buspirone and some of the newer antidepressants have become the treatment of choice for patients with GAD requiring long-term treatment. Buspirone was the first available alternative to the benzodiazepines in the US; however, the initial excitement over this agent was somewhat dampened because of its mild efficacy combined with a slow onset of action. The antidepressants were seen as beneficial for the treatment of GAD because of the high comorbidity with depression, thus allowing a better outcome for these patients. The antidepressants that offer both a good adverse effect profile and efficacy are the selective serotonin reuptake inhibitors including paroxetine, and the serotonin-norepinephrine reuptake inhibitors such as venlafaxine. Clinicians should also consider the potential benefits of psychotherapy as an adjunct to medication. There are a number of potentially new pharmacotherapies being investigated, including newer serotonin 5-HT1A receptor agonists, cholecystokinin receptor antagonists, neurokinin receptor antagonists, gabapentin and its analogues, and gamma-aminobutyric acid (GABA)A receptor modulators. However, these compounds are all in the early stages of investigation, and there are no new therapies expected to be released in the near future. Nonetheless, in the search for the ideal anxiolytic, a more positive outlook is allowed by imminent future research for new treatment options in patients with GAD. PMID- 12109927 TI - Fondaparinux sodium. AB - black triangle Fondaparinux sodium, a selective factor Xa inhibitor, is the first in a new class of antithrombotics. It binds selectively with high affinity to antithrombin III and specifically catalyses the inactivation of factor Xa. The elimination half-life of fondaparinux sodium permits once daily treatment. black triangle A randomised, double-blind, parallel-group, dose-ranging, multicentre phase IIb study in 933 eligible patients established that a subcutaneous dose of between 1.5 and 3mg of fondaparinux sodium has the optimum efficacy and safety profile for prophylaxis of venous thromboembolism in patients undergoing major orthopaedic surgery. black triangle Fondaparinux sodium, given to more than 3600 patients undergoing major orthopaedic surgery who participated in prospective, randomised, double-blind, multicentre phase III clinical trials, significantly reduced the incidence of venous thromboembolism, with an overall risk reduction of 55.2% compared with enoxaparin. black triangle Fondaparinux sodium was well tolerated by patients undergoing major orthopaedic surgery, and at the recommended clinical dose of 2.5mg has a similar tolerability profile, including bleeding events, to standard enoxaparin regimens. Fondaparinux sodium has not been reported to cause antibody-induced thrombocytopenia. PMID- 12109926 TI - Antipsychotic-related QTc prolongation, torsade de pointes and sudden death. AB - Sudden unexpected deaths have been reported with antipsychotic use since the early 1960s. In some cases the antipsychotic may be unrelated to death, but in others it appears to be a causal factor. Antipsychotics can cause sudden death by several mechanisms, but particular interest has centred on torsade de pointes (TdP), a polymorphic ventricular arrhythmia that can progress to ventricular fibrillation and sudden death. The QTc interval is a heart rate-corrected value that represents the time between the onset of electrical depolarisation of the ventricles and the end of repolarisation. Prolongation of the QTc interval is a surrogate marker for the ability of a drug to cause TdP. In individual patients an absolute QTc interval of >500 msec or an increase of 60 msec from baseline is regarded as indicating an increased risk of TdP. However, TdP can occur with lower QTc values or changes. Concern about a relationship between QTc prolongation, TdP and sudden death applies to a wide range of drugs and has led to the withdrawal or restricted labelling of several. Among antipsychotics available in the UK, sertindole was voluntarily suspended, droperidol was withdrawn, and restricted labelling introduced for thioridazine and pimozide. The degree of QTc prolongation is dose dependent and varies between antipsychotics reflecting their different capacity to block cardiac ion channels. Significant prolongation is not a class effect. Among currently available agents, thioridazine and ziprasidone are associated with the greatest QTc prolongation. Virtually all drugs known to cause TdP block the rapidly activating component of the delayed rectifier potassium current (I(kr)). Arrhythmias are more likely to occur if drug-induced QTc prolongation coexists with other risk factors, such as individual susceptibility, presence of congenital long QT syndromes, heart failure, bradycardia, electrolyte imbalance, overdose of a QTc prolonging drug, female sex, restraint, old age, hepatic or renal impairment, and slow metaboliser status. Pharmacodynamic and pharmacokinetic interactions can also increase the risk of arrhythmias. Further research is needed to quantify the risk of sudden death with antipsychotics. The risk should be viewed in the context of the overall risks and benefits of antipsychotic treatment. It seems prudent, where possible, to select antipsychotics that are not associated with marked QTc prolongation. If use of a QTc-prolonging drug is warranted, then measures to reduce the risk should be adopted. PMID- 12109930 TI - Balsalazide: a review of its therapeutic use in mild-to-moderate ulcerative colitis. AB - The aminosalicylate balsalazide is a prodrug which is metabolised by bacterial azo reductases in the colon to release its therapeutically active moiety mesalazine [mesalamine (US) or 5-aminosalicylic acid] and an inert carrier molecule. The systemic absorption of balsalazide and its metabolites is not required for the therapeutic efficacy of the drug, and has been demonstrated to be limited. Data from well designed trials with a duration of 8 to 12 weeks show that oral balsalazide 6.75 g/day is as effective as (two trials) or more effective than (one trial) oral delayed-release (pH-dependent) mesalazine 2.4 g/day and appears to be as effective as oral sulfasalazine 3 g/day in the treatment of active mild-to-moderate ulcerative colitis. In addition, balsalazide appears to have a faster onset of action than mesalazine. Furthermore, balsalazide was as effective as delayed-release mesalazine (dosages used were 1.2 and 1.5 g/day, where 1.6 g/day is recommended) and oral sulfasalazine 2 g/day (recommended dosage) in the prevention of relapse in ulcerative colitis in remission after 6 to 12 months of treatment; the balsalazide dosage was 3 g/day versus mesalazine and 2 g/day versus sulfasalazine. Although not well established, additional benefits may be achieved with balsalazide dosages up to 6 g/day. Data from well designed, 2- to 12-month trials show that balsalazide is well tolerated by patients with ulcerative colitis in both acute and maintenance indications, and is better tolerated than standard formulations of sulfasalazine at therapeutically relevant dosages. CONCLUSION: Balsalazide is a well tolerated and effective first-line therapeutic option for patients with ulcerative colitis, both for the treatment of active mild-to-moderate disease and as maintenance therapy to prevent disease relapse. PMID- 12109931 TI - The epidemiology of smoking: health consequences and benefits of cessation. AB - Tobacco use is the single most important preventable health risk in the developed world, and an important cause of premature death worldwide. Smoking causes a wide range of diseases, including many types of cancer, chronic obstructive pulmonary disease, coronary heart disease, stroke, peripheral vascular disease, and peptic ulcer disease. In addition, smoking during pregnancy adversely affects fetal and neonatal growth and development. Recent decades have seen a massive expansion in tobacco use in the developing world and accelerating growth in smoking among women in the developed world. Globally, smoking-related mortality is set to rise from 3 million annually (1995 estimate) to 10 million annually by 2030, with 70% of these deaths occurring in developing countries. Many of the adverse health effects of smoking are reversible, and smoking cessation treatments represent some of the most cost effective of all healthcare interventions. Although the greatest benefit accrues from ceasing smoking when young, even quitting in middle age avoids much of the excess healthcare risk associated with smoking. In order to improve smoking cessation rates, effective behavioural and pharmacological treatments, coupled with professional counselling and advice, are required. Since smoking duration is the principal risk factor for smoking-related morbidity, the treatment goal should be early cessation and prevention of relapse. PMID- 12109924 TI - Ocular allergy guidelines: a practical treatment algorithm. AB - The treatment of ocular allergy requires a better understanding of the spectrum of clinical disorders involving various components of the immune system, and of interactions at the conjunctival surface. The immune response focuses primarily on the different levels of activity of Th2 lymphocytes and various other immune cells associated with allergic disorders, including mast cells, eosinophils, fibroblasts, and epithelial and endothelial cells. Ocular allergic disorders include seasonal allergic conjunctivitis (SAC), perennial allergic conjunctivitis (PAC), vernal keratoconjunctivitis (VKC), giant papillary conjunctivitis (GPC) and atopic keratoconjunctivitis (AKC), which, through immunopathological and molecular immunological techniques, can all be better appreciated as being part of a larger spectrum of an atopic disease state. In SAC, pathological changes, such as increased mast-cell activation, the presence of migratory inflammatory cells, and early signs of cellular activation at the molecular level, are minimal. In PAC, these changes are more pronounced in line with the increased duration of allergenic stimulation. In more chronic forms of allergic conjunctivitis, such as VKC in children and AKC in adults, the following changes are evident: a persistent state of mast cell, eosinophil and lymphocyte activation; noted switching from connective-tissue to mucosal-type mast cells; increased involvement of corneal pathology; and follicular development and fibrosis. The treatment of acute and more chronic forms of allergic conjunctivitis has focused in the past on symptomatic relief of symptoms, but with a better understanding of the mechanisms involved we can now provide interventional therapeutic strategies and symptomatic relief. Our advances in the basic understanding of these conditions are providing the foundation for guidelines that improve the ocular health of patients with ocular allergies. PMID- 12109932 TI - Pharmacokinetic optimisation of sustained-release bupropion for smoking cessation. AB - Sustained-release bupropion (bupropion SR) is a unique, non-nicotine smoking cessation aid that is hypothesised to act upon neurological pathways involved in nicotine dependence. Pharmacokinetic and metabolism studies reveal that bupropion SR is metabolised by multiple pathways with no single pathway predominating. When one pathway is inhibited, others are available to compensate. Therefore, only a few clinically relevant drug-drug interactions involving bupropion SR have been observed, although the potential for interactions exists, as with any extensively metabolised drug. Population pharmacokinetic/pharmacodynamic analyses of data from patients receiving daily oral doses of 100mg, 150mg, or 300mg reveal that the anti-smoking efficacy of bupropion SR is directly related to dose. The incidences of dry mouth and insomnia were directly related to bupropion plasma concentrations while the incidence of anxiety was inversely proportional to bupropion plasma concentrations. To maximise efficacy (with an acceptable safety profile), the optimal daily dose for the majority of patients is 300mg. PMID- 12109933 TI - Clinical efficacy of bupropion in the management of smoking cessation. AB - Nicotine addiction is a chronic relapsing condition that can be difficult to treat. Until recently, pharmacological options for the treatment of tobacco dependence were primarily limited to nicotine replacement therapy (NRT). Sustained-release bupropion (bupropion SR) is the first non-nicotine pharmacological treatment approved for smoking cessation. Bupropion SR is recommended for first-line pharmacotherapy alongside NRT in the updated US Clinical Practice Guidelines and the UK Health Education Authority Guidelines. The UK National Institute of Clinical Excellence recommends NRT and bupropion SR for smokers who have expressed a desire to quit smoking. This review presents evidence that bupropion SR is an effective first-line therapy for smoking cessation in a wide range of patient populations. It is associated with significantly higher smoking cessation rates compared with placebo in patients with or without a history of prior bupropion SR or NRT use, and its effect is independent of gender. Bupropion SR treatment is effective in the prevention of relapse to smoking in those patients who have successfully quit, and re-treatment is effective in smokers who recommence smoking after a previous course of bupropion SR. Bupropion SR treatment relieves the symptoms of craving and nicotine withdrawal, and attenuates the weight gain that often occurs after smoking cessation. Data collected from motivational support programmes and employer-based studies provide strong evidence of the effectiveness of bupropion SR as an aid to smoking cessation in 'real life' situations, and confirm the efficacy seen in clinical trials. PMID- 12109934 TI - Use of sustained-release bupropion in specific patient populations for smoking cessation. AB - Smoking cessation trials of sustained-release bupropion (bupropion SR) were initially conducted in a general population of smokers who were motivated to quit smoking. Bupropion SR has also been found to be a useful treatment of tobacco dependence in various special populations of smokers who often experience difficulty in overcoming tobacco addiction. Point-prevalence quit rates at 6 months were higher in those treated with bupropion SR than in those receiving placebo in studies on smokers with chronic obstructive pulmonary disease (23% vs 16%) and in those with cardiovascular disease (34% vs 12%). Abstinence from smoking after treatment with bupropion SR was not affected by a history of major depression or alcoholism. Women treated with bupropion SR were just as likely as men to abstain from smoking. Approximately one-third of a study population who were initially unwilling or unable to quit smoking were able to reduce their smoking by 50% or more during therapy with bupropion SR; 14% of these went on to achieve abstinence. Bupropion SR was well tolerated in these trials; importantly, it had no clinically significant effect on mean blood pressure in smokers, including those with hypertension, and attenuated the weight gain associated with smoking cessation, particularly in women. PMID- 12109935 TI - Tolerability and safety of sustained-release bupropion in the management of smoking cessation. AB - Sustained-release bupropion (bupropion SR) was first launched in the US in 1997 as an aid to smoking cessation and has since been launched in many other countries. Adverse events associated with the use of bupropion SR at the recommended dosage of 150mg twice daily in clinical trials most commonly included insomnia, headache, dry mouth, nausea and anxiety; insomnia and anxiety are also recognised as symptoms of nicotine withdrawal. Only insomnia and dry mouth occurred significantly more frequently with bupropion SR than with placebo. Relative to placebo, no significant changes in mean values for heart rate, blood pressure or routine laboratory parameters have been reported in smokers using bupropion SR alone in clinical trials. When bupropion SR was compared with a nicotine transdermal patch in a clinical trial, insomnia predominated in the bupropion SR group, while dream abnormalities were more common in smokers using the nicotine patch. Bupropion SR and the nicotine transdermal patch in combination can be used safely (with appropriate monitoring) as an aid to smoking cessation. Infrequent but clinically important adverse reactions to bupropion SR include seizures and hypersensitivity reactions: in controlled clinical trials of bupropion SR (300 mg/day), where smokers were carefully screened for risk factors for seizure, the incidence of both seizures and severe hypersensitivity reactions was approximately 0.1% for each event. In order to avoid a risk of seizure of greater than 0.1%, smokers should be screened for predisposing risk factors and adhere to the manufacturer's dosage recommendations (maximum daily dose of 300mg). Thus, bupropion SR is generally well tolerated, as seen by the low discontinuation rate due to an adverse event in clinical trials (6 to 12%). The most common adverse events (insomnia and dry mouth) are generally transient and often resolve quickly without therapeutic intervention; they can be managed if necessary by a reduction in bupropion dose. PMID- 12109938 TI - Critical care apheresis. PMID- 12109936 TI - Current approaches to the management of smoking cessation. AB - Smoking remains a widespread intractable behaviour and is a significant cause of morbidity and mortality worldwide. Effective approaches to smoking cessation include behavioural intervention and pharmacotherapy, in particular nicotine replacement therapy (NRT) and sustained-release bupropion (bupropion SR). Pharmacotherapy remains a popular choice of smoking cessation intervention for many smokers, and both NRT and bupropion SR, combined with behavioural interventions, achieve 1.5- to >2-fold increases in smoking cessation rates. Various national and international smoking cessation guidelines have been published recommending effective implementation of smoking cessation strategies. Recommendations include the systematic identification of smokers, assessment of their willingness to quit smoking, provision of advice promoting a cessation attempt, and administration of approved first-line therapies. PMID- 12109939 TI - Double filtration plasmapheresis in critical care. AB - Many kinds of technologies have been introduced and successfully developed for therapeutic apheresis. Furthermore, several kinds of these technologies have also been applied in critical care. Double filtration plasmapheresis (DFPP), however, is rarely applied in this field in comparison with other treatments such as continuous hemofiltration, continuous hemodiafiltration, single filtration plasmapheresis, and plasma adsorption therapies. In this paper, the characteristics of the DFPP treatments for critical care are summarized. During the DFPP treatments, the patient's blood volume (BV) often decreases with time due to albumin loss induced by inadequate albumin infusion in a supplementation fluid. We examined the change of BV by a continuous hematocrit monitor, Crit Line, during an in vivo study for 9 patients. As a result, albumin loss fairly occurred in DFPP treatments. The decrease of patient BV was induced by an oncotic pressure drop due to albumin loss and often resulted in a blood pressure drop. This is a serious problem for DFPP in critical care. We should avoid inadequate albumin infusion if the patient is suffering from these adverse effects. In order to determine the optimal concentration C(S) and volume V(S) values of a supplemented albumin solution, we introduced a variable blood volume model for albumin transport in DFPP. PMID- 12109937 TI - Pharmacoeconomic considerations in the management of smoking cessation. AB - Smokers are more likely to develop a variety of serious diseases than non smokers; the morbidity and mortality from these diseases place a great resource burden on society in respect of demands on healthcare resources and lost productivity. The cost to the healthcare system of treating the consequences of smoking is high. Given the availability of inexpensive pharmaceutical therapies such as sustained-release bupropion (bupropion SR) and nicotine replacement therapy (NRT), which are proven to be effective, there is economic advantage for governments and the medical profession in encouraging patients to quit. The cost effectiveness of effective smoking cessation support is far superior to that of many other potentially life-saving interventions and, indeed, the UK National Institute for Clinical Excellence (NICE) stated in April 2002 that "both bupropion and NRT are considered to be amongst the most effective of all healthcare interventions". Recent economic analyses have confirmed that the use of bupropion SR as an aid to smoking cessation is a highly cost-effective intervention. PMID- 12109940 TI - Plasma adsorption in critical care. AB - Plasmapheresis therapies such as plasma exchange (PE), double filtration plasmapheresis (DFPP), or immunoadsorption plasmapheresis (IAPP) have become therapeutic tools in critical care. PE or DFPP are limited by their non- or semiselective removal of all plasma components. Replacement fluids such as fresh frozen plasma and albumin are necessary during PE or DFPP. There is the risk of infection and allergic reactions whenever such fluids are used. On the other hand, IAPP is superior to PE and DFPP because it does not require any replacement fluid. There has been development of many adsorbent columns used for removing specific pathogenic substances, and patients with various kinds of critical illness have been treated with IAPP. However, IAPP can be applied only for certain diseases because of the limitations of the commercially available columns. It is concluded that the development of new adsorption therapy may improve the high mortality and morbidity rate in critically ill patients. PMID- 12109941 TI - Hemoadsorption in critical care. AB - This paper concerns the results of endotoxin hemoadsorption therapy using a PMX column in patients with perforative peritonitis complicated by multiple organ failure. The subjects were 31 patients aged 68 +/- 12 years. When systolic arterial pressure decreased to less than 90 mm Hg, endotoxin hemoadsorption was initiated and continued for 2 h. At the completion of endotoxin hemoadsorption, systolic arterial pressure, diastolic arterial pressure, and mean arterial pressure were significantly increased. Platelet count decreased to less than 50,000/mm(3) in 30% of patients. As for cytokines and vascular endothelial cell function markers, interleukin-6 and plasminogen activator inhibitor-1 significantly decreased. These results suggest favorable effects of endotoxin hemoadsorption on the hemodynamic and pathophysiological conditions in patients with septic shock although attention should be given to the decrease in platelet count. PMID- 12109942 TI - Continuous hemofiltration/hemodiafiltration in critical care. AB - Continuous hemofiltration and continuous hemodiafiltration (CHF/CHDF) were developed as continuous renal replacement therapy for patients with severe conditons and has come to be widely performed mainly in critical care, taking the place of intermittent hemodialysis. The membrane pore size of a hemofilter used for CHF/CHDF allows passage of substances ranging from 30,000 to 50,000 Da, and the method for solute removal in CHF/CHDF employs the principle of convection, which is advantageous for removing middle- to high-molecular-weight substances. As apheresis therapy to remove pathogenic substances in blood, CHF/CHDF is therefore being investigated for its possible effect on various morbid conditions. It has recently been found that CHF/CHDF removes humoral mediators including cytokines, particularly in severe systemic inflammatory response syndromes such as septic shock and severe acute pancreatitis. CHF/CHDF is thus beginning to be performed for the prevention and treatment of organ dysfunction secondary to septic shock, trauma, or acute pancreatitis. CHF/CHDF is also efficacious as artificial liver support in preventing adverse effects caused by plasma exchange (PE) and for continuous removal of hepatic coma-inducing substances. CHF/CHDF is effective for various morbid conditions not only as renal replacement therapy, but also as apheresis therapy and is expected to be applied more widely in critical care in the future. PMID- 12109943 TI - On-line hemodiafiltration in critical care. AB - On-line products of substitution fluid permits virtually unlimited fluid volume exchange during continuous hemodiafiltration (CHDF) to critical care. In on-line hemodiafiltration (HDF), endotoxin free dialysate obtained using pyrogen cut filters is infused into the blood circuit, and HDF is automatically performed using the closed-loop balancing system of the dialysis machine. On-line CHDF is the application of this on-line HDF to continuous renal replacement therapy in the critical care field. We performed on-line CHDF on 376 acute renal failure patients during a 5 year period, and the mean survival rate was 62.5%. We concluded that the on-line CHDF system is safe and effective at maintaining acute renal failure patients. PMID- 12109944 TI - Critical care by cytapheresis. AB - We report our experience of cytapheresis using a nonwoven polyester fiber filter to treat critical states of immune diseases. In 7 critical states of ulcerative colitis (UC), cytapheresis was effective in improving symptoms of UC. Administration of steroids was important in some cases. In 3 cases of renal transplantation, cytapheresis was also effective in controlling rejection. IgA nephropathy of transplanted cases was well controlled. Furthermore, an original disease such as focal segmental glomerulosclerosis (FCGS) in a transplant patient was well treated by extracorporeal immune modulation of the cytapheresis. PMID- 12109945 TI - Long-term survivors with artificial liver support in fulminant hepatic failure. AB - Clinical ability of artificial liver support (ALS) has been improved greatly in recent years which has allowed us to encounter long-term survivors with fulminant hepatic failure (FHF) whose liver function has been almost completely lost. This suggests that application of ALS in patients with FHF gains time while awaiting transplantation as well as time for functional recovery and regeneration of the liver graft following receipt of the graft with marginal function and/or size. Thus, ALS will contribute greatly to extending the indications for liver transplantation and increase the number of patients receiving and benefiting from this treatment. On the other hand, introduction of ALS prolongs the duration of intensive treatment which increases the risk of infection and increases medical costs. In addition, when to discontinue intensive treatment of patients whose level of consciousness is maintained only by ALS is controversial. Thus, further investigation will be needed to establish a consensus on indications for long term ALS in FHF. PMID- 12109946 TI - Affinity hemodialysis for antiviral therapy. I. Removal of HIV-1 from cell culture supernatants, plasma, and blood. AB - We tested an affinity hemodialysis technique designed to efficiently remove HIV and toxic viral proteins from blood. Miniature polyethersulfone hollow-fiber dialysis cartridges (200-500 nm pore) were packed with anti-HIV antibodies covalently coupled to agarose beads and sealed inside the cartridge. Cell culture fluids, plasma, or infected blood (7-15 ml) containing HIV-1 were circulated over the cartridge at 0.7-10 ml/min and the rate of removal of HIV measured by PCR and p24 ELISA. The technique removed up to 98% of HIV-1 particles from cell culture supernatants. Affinity hemodialysis also efficiently captured cultured HIV from human blood plasma (90%) and native HIV from infected blood (83% to 100%). Viral capture followed first-order kinetics (t(1/2) = 2.8 h). Variations in antibody type, matrix linkage (protein G versus direct coupling), bead pore size, and temperature of operation (25-37 degrees C) had only small effects. Although some binding was nonspecific, direct binding to the immobilized antibodies appeared to be the predominant mechanism. PMID- 12109947 TI - A patient with severe acute pancreatitis successfully treated with a new critical care procedure. AB - It has been accepted widely that excessive humoral mediators play important roles in the pathogenesis of organ failure in patients with severe acute pancreatitis (SAP) and that infection of the pancreas due to bacterial translocation (BT) is the most frequent cause of death in SAP. On the other hand, it has been reported that continuous hemodiafiltration (CHDF) removes humoral mediators on hypercytokinemic patients such as those with systemic inflammatory response syndrome. Furthermore, several clinical studies have demonstrated that selective digestive decontamination (SDD) effectively eliminates aerobic Gram-negative bacteria from the intestinal tract and reduces the incidence of septic complications in SAP. Herein we report a case of SAP who was treated successfully with intensive care including CHDF and SDD. Thus, this case report suggests that CHDF aimed at removing causative humoral mediators and SDD for the prevention of BT are useful new tools for the management of SAP. PMID- 12109948 TI - Plasma exchange in patients with toxic epidermal necrolysis. AB - We describe our experience with plasma exchange (PE) therapy in 13 patients with drug-induced toxic epidermal necrolysis (TEN), 4 of whom had malignant disorders. Skin lesions covered 17% to 100% of total body surface area and 1 to 4 mucous membranes were involved. None of the patients was hospitalized in a burn unit. The patients underwent from 2 to 5 PE sessions (mean 3.4 +/- 0.2 standard error of mean [SEM], median 3) exchanging 6.6 to 17.6 L of plasma (mean 10.1 +/- 0.7 SEM, median 10). PE sessions were carried out every other day in 8 patients and daily in 5. Three patients died (23%) while the remaining 10 (77%) had a full recovery. Plasmapheresis may be an effective treatment in patients with drug induced TEN hospitalized outside a burn unit. PMID- 12109949 TI - Influence of single low-density lipoprotein apheresis on the adhesion molecules soluble vascular cellular adhesion molecule-1, soluble intercellular adhesion molecule-1, and P-selectin. AB - The aim of our study was to investigate the influence of single low-density lipoprotein apheresis (heparin extracorporeal low-density lipoprotein precipitation [HELP]procedure) on plasma concentrations of soluble adhesion molecules (sAMs) such as soluble vascular cellular adhesion molecule-1 (sVCAM-1), soluble intercellular adhesion molecule-1 (sICAM-1), and P-selectin in patients with familial heterozygous hypercholesterolemia and documented coronary artery disease enrolled in a chronic weekly HELP apheresis. Before HELP apheresis, the mean plasma concentration of sVCAM-1 was 515 +/- 119 ng/ml, 204 +/- 58 ng/ml for sICAM-1, and 112 +/- 45 ng/ml for P-selectin. After single HELP apheresis, plasma concentrations of sAM declined significantly by 32 +/- 7%, 18 +/- 15%, and 33 +/- 25% for sVCAM- 1,sICAM-1 and P-selectin, respectively. After a 1 week interval, sAM concentrations rose to approximately the initial values. The concentrations of all sAMs studied were significantly lower in the plasma leaving than entering the filter. Due to filtration, the decline in plasma level of sVCAM-1, sICAM-1, and P-selectin was 62 +/- 19%, 51 +/- 39%, and 67 +/- 22%, respectively. In addition to lipid reduction, single HELP apheresis significantly lowers plasma concentrations of sVCAM-1, sICAM-1, and P-selectin. PMID- 12109950 TI - Continuous hemodiafiltration with polymyxin-B immobilized fiber is effective in patients with sepsis syndrome and acute renal failure. AB - The aim of this study was first, to evaluate the effects of continuous hemodiafiltration (CHDF) alone or combined with CHDF and polymyxin-B immobilized fiber (PMX) on survival rates of patients with sepsis and acute renal failure, and second, to evaluate the changes in plasma levels of inflammatory cytokines before and after treatment with CHDF and PMX and CHDF alone in these patients. Forty-eight patients with septic shock and acute renal failure were enrolled in this study. The survival rate of all patients at 28 days was 25% for those with CHDF and 75% for those with PMX and CHDF treatment. Combination treatment produced a significant reduction of plasma levels of endotoxin and interleukin-6 compared to the basal values and to the treatment with CHDF alone. From these data, it is suggested that the combined therapy with PMX and CHDF is effective in improvement of survival rate of patients with septic shock and acute renal failure. PMID- 12109951 TI - The neurochemistry of waking and sleeping mental activity: the disinhibition dopamine hypothesis. AB - This paper describes a hypothesis related to the neurochemical background of sleep-waking mental activity which, although associated with subcortical structures, is principally generated in the cerebral cortex. Acetylcholine, which mainly activates cortical neurons, is released at the maximal rate during waking and rapid eye movement (REM) sleep dreaming stage. Its importance in mental functioning is well-known. However, brainstem-generated monoamines, which mainly inhibit cortical neurons, are released during waking. Both kinds of influences contribute to the organized mentation of waking. During slow wave sleep, these two types of influence decrease in intensity but maintain a sufficiently high level to allow mental activity involving fairly abstract pseudo-thoughts, a mode of activity modelled on the diurnal pattern of which it is a poor reply. During REM sleep, the monoaminergic neurons become silent except for the dopaminergic ones. This results in a large disinhibition and the maintained dopamine influence may be involved in the familiar psychotic-like mental activity of dreaming. Indeed, in this original activation-disinhibition state, the increase of dopamine influence at the prefrontal cortex level could explain the almost total absence of negative symptoms of schizophrenia during dreaming, while an increase in the nucleus accumbens is possibly responsible for hallucinations and delusions, which are regular features of mentation during this sleep stage. PMID- 12109952 TI - Effects of low-dose ketamine on neuropathic pain: An electroencephalogram electrooculogram/behavioral study. AB - The aim of the present study was to clarify the neurophysiological changes associated with analgesic and behavioral effects of low-dose ketamine HCl in patients suffering from chronic neuropathic pain. Ten in-patients with neuropathic pain participated in this single-blind, placebo-controlled study after giving written informed consent. Following intravenous injections of a saline solution (placebo), three bolus injections of 5 mg ketamine HCl were administered at 5 min intervals. Changes in pain perception were assessed using a numerical rating scale for pain. Behavioral changes, including psychotomimetic effects, were assessed using the Brief Psychiatric Rating Scale (BPRS). Electroencephalograms (EEG) and electrooculograms (EOG) were recorded continuously throughout the testing period. One minute EEG and closed-eye eye movements were quantified. The effects of ketamine were evaluated by comparing the neurophysiological and behavioral parameters obtained from the placebo and ketamine trials. Pain reduction was significantly correlated with ketamine induced changes in hallucinatory behavior and excitement as measured by the BPRS. Ketamine injections caused a significant decrease in the EEGalpha amplitude without an accompanying reduction in EEG frequency. The EEGalpha amplitude reduction at the right central electrode was significantly related to subjective pain relief. Subanesthetic doses of ketamine significantly decreased rapid eye movements, but did not initiate slow eye movements. In conclusion, the present EEG-EOG/behavioral results indicate that ketamine-induced failure of neural integration between cortical-subcortical regions induces psychotic symptoms and alters pain perception on neuropathic pain. PMID- 12109953 TI - Psychosocial problems in attention-deficit hyperactivity disorder with oppositional defiant disorder. AB - The purpose of this study is to clarify psychosocial characteristics of the comorbidity of attention-deficit hyperactivity disorder (ADHD) and oppositional defiant disorder (ODD) in comparison with ADHD or ODD alone. Thirty-one patients with ADHD comorbid with ODD were compared with 23 ADHD alone and 10 with ODD alone, in terms of various examination items including objective assessment scales. The comorbid group demonstrated higher Children Depression Inventory score and State-Trait Anxiety Inventory for Children (state-anxiety) score than the ADHD or the ODD group, possessing more problems in the relationship with teachers than the ADHD group, with friends more than the ADHD or the ODD group, and with their mothers more than the ADHD group and less than the ODD group. School refusal occurred more frequently in the comorbid group than the ADHD group and less than the ODD group. The comorbid group had more psychosocial problems than the ADHD group and the ODD group. These problems could be classified into three types: (i) those derived from ODD, problems in the relationship with teachers; (ii) those derived from ODD but reduced by the coexistence of ADHD, problems in the relationship with their mothers; and (iii) those resulting from the comorbidity of ADHD and ODD, problems in the relationship with friends and anxious and depressive tendency. The difficulties in the relationship with teachers and friends observed in the comorbid group may lead to school refusal. PMID- 12109955 TI - Frustration and fulfillment of needs in dissociative and conversion disorders. AB - We reviewed all patients with dissociative disorders (nine patients with dissociative amnesia or dissociative fugue) and conversion disorders (10 patients) who were admitted and treated during the past 15 years. Needs frustrated at the appearance of the symptoms and those fulfilled at discharge were studied in both groups using Maslow's hierarchy of needs. The patients of both groups who encountered troubles in their life events were found to have frustrated needs. These symptoms tended to be accompanied more often by frustrations regarding a 'need for love' in the dissociative disorders group and by frustration in the need for 'self-esteem and self-actualization' in the conversion disorders group. In addition, needs of lower orders were already threatened at onset in many patients. The symptoms disappeared in patients in whom the situation completely improved (needs were fulfilled), but the symptoms were alleviated or unchanged in those in whom the problems remained unresolved. PMID- 12109954 TI - Supporting by a nurse teacher in a school infirmary using collage therapy. AB - The objective of this study is to verify the effects of collage therapy when applied in the school infirmary, without time restriction, to children with problems such as school refusal. Collage therapy was used with two students who refused to attend school for a period of 1 year and 2 months to 1 year and 7 months. The results showed that both cases were successful in recovering self insight and starting to pursue a course leading to the establishment of their identity. For students with various mental health problems, the school, as a community of school friends of the same generation, has been suggested to be one of the most important places for the recovery of energy. The performance in the school infirmary of collage therapy without any time restraints makes use of this environment, and appears to foster self-insight and self-understanding, and hence promote recovery from mental health problems. PMID- 12109956 TI - Bizarre delusions and DSM-IV schizophrenia. AB - The present study investigated whether schizophrenia patients with and without DSM-IV bizarre delusions, categorized as bizarre delusions of Schneiderian first rank symptoms (SBD) and as non-Schneiderian bizarre delusions (non-SBD), differed on demographic or clinical features, in view of the weight given to bizarre delusions in the diagnosis of schizophrenia. One hundred and twenty-nine in patients with schizophrenia were assessed systematically for both types of bizarre delusions on the five domains of psychopathology of the Positive and Negative Syndrome Scale (PANSS; delusions/hallucinations, thought disorder/disorganization, excitement, negative symptoms and depressive symptoms) and for extrapyramidal side-effects. Inter-rater reliabilities for SBD and non SBD were assessed and were exceptionally high (kappa value 0.85 and 0.92, respectively). Neither SBD nor non-SBD were associated with any demographic or non-PANSS clinical characteristics tested. However, the presence of non-SBD was significantly associated with more severe psychopathology in all five domains of the PANSS, whereas the presence of SBD was significantly associated with more severe psychopathology in three domains only: delusions/hallucinations, thought disorder/disorganization and depressive symptoms. However, patients with only SBD did not differ from patients with only non-SBD on any demographic or clinical variables, including five psychopathological domains. These findings suggest that, despite showing more severe symptoms, patients with DSM-IV bizarre delusions do not constitute a clinically distinguishable subgroup. PMID- 12109957 TI - Efficacy of day care treatment against readmission in patients with schizophrenia: A comparison between out-patients with and without day care treatment. AB - The present study examined the efficacy of day care (DC) treatment against readmission to a mental hospital. Subjects were 65 out-patients with chronic schizophrenia after discharge. Day care treatment was defined as positive if patients were under DC treatment constantly for 4 months or more with a frequency of at least one visit per week. Information regarding demographic and disease related factors was obtained from medical records at the time of discharge. Logistic regression analysis was used to calculate odds ratios (OR) and 95% confidence intervals (CI) with adjustment for confounding variables. The modestly preventive efficacy of DC treatment against readmission was observed within 2 years after discharge (adjusted OR 0.52; 95% CI 0.12-2.35). The present findings suggest that DC treatment may be preventive against readmission for schizophrenic out-patients. PMID- 12109958 TI - Wisconsin Card-Sorting Test performance does not discriminate different patterns of premorbid behavioral abnormalities in schizophrenic patients. AB - It is well documented that premorbid behavior abnormalities precede the onset of schizophrenia in a large number of patients. The research findings suggest that there are differences in the type and severity of these premorbid dysfunctions. Another research field has shown impairment of preschizophrenic patients in several cognitive domains. The present study reports retrospective childhood and adolescence neurobehavioral assessment in 31 patients with schizophrenia by the Child Behavior Checklist (CBCL) and current Wisconsin Card-Sorting Test (WCST) evaluation in order to investigate whether specific behavioral abnormality (BA) patterns are related to executive function as evaluated by the WCST. Cluster analysis was conducted on the childhood premorbid behavior ratings for the schizophrenic patients and two subgroups emerged: (i) Cluster I with an initial low level of BA that increased over the years; and (ii) Cluster II with a high level of BA that remained relatively stable until early adulthood. Furthermore, Cluster II showed more severe current negative and total symptoms, but the two groups did not differ in WCST performance. Our results show that the patterns and severity of CBCL upon retrospective evaluation are not related to WCST performance, which seems to be a feature inherent to the disease process. Different factors could be responsible for cognitive and behavioral disturbances in schizophrenia. PMID- 12109959 TI - Eye movements during the Rorschach test in schizophrenia. AB - In order to understand relationships between scanning behaviors, characteristics of visual stimuli and the clinical symptoms in schizophrenia, eye movements of 37 schizophrenic patients and 36 controls were recorded using an eye-mark recorder during a free-response period in a Rorschach test. Four cards (I, II, V and VIII) were used. Data were analyzed during 15 s from the presentation of each card. For all cards, the number of eye fixations and the number of eye fixation areas were fewer, and total scanning length and mean scanning length were shorter for schizophrenic patients than for controls. For card II, in the non-popular response group, eye fixation frequency upon area 5 + 6 (red) was higher for schizophrenic patients. For card VIII, in the popular response group, eye fixation frequency upon area 5 + 6 (pink) was lower for schizophrenic patients. For cards II and VIII, the number of eye fixations was inversely correlated with negative symptoms. For card II, total scanning length tended to be inversely correlated with negative symptoms, and mean eye fixation time was correlated with negative symptoms. The number of eye fixation areas was inversely correlated with positive symptoms. For card VIII, eye fixation frequency in a stimulative area tended to be correlated with positive symptoms. Scanning behaviors in schizophrenic patients are affected by characteristics of visual stimuli, and partially by clinical symptoms. PMID- 12109960 TI - Attentional dysfunction of chronic schizophrenia: No association with long-term institutionalization. AB - Attentional processes play a central role in information selection, which is impaired in schizophrenic patients. In the present study, we attempted to characterize the attentional performance of chronic schizophrenics using a computerized assessment of the multiple components of attentional function. Two comparable samples, consisting, respectively, of out-patients and in-patients, were tested in order to assess the effect of chronic institutionalization. Twenty four subjects (half in-patients and half out-patients) fulfillling DSM-IV criteria for schizophrenia were examined with a standard computerized battery for the assessment of attention, namely Testbatterie zur Aufmerksamkeitsprufung (TAP). Both groups were impaired on all measures of attentional processing (in terms of both reaction times and number of errors). There were no significant differences in attentional performance between in- and out-patients. In conclusion, the present findings confirm the presence of pervasive attentional dysfunction in chronic schizophrenia; the lack of significant differences in performance between in- and out-patients supports the hypothesis that the cognitive deficits are inherently associated with the illness and cannot be attributed to environmental/social factors. PMID- 12109961 TI - The flunitrazepam abuse prevention program at a general hospital in Taiwan: a descriptive study. AB - The Bureau of Controlled Drugs at Ministry of Health, Executive Yuan in Taiwan announced, on 1 April 2000, the schedules of controlled drugs with abuse potential and implemented a policy on 1 October 2000 to control them. Flunitrazepam (FM2), along with other two benzodiazepines (triazolam and brotizolam), is placed on Schedule III. The aim of the present study was to analyze the pattern of flunitrazepam prescriptions across all medical subspecialty departments at Taipei Medical University-Wan Fang Hospital (TMU WFH), Taiwan. We analyzed 1170 prescriptions over 12 month period from 1 July 2000 to 31 May 2001. All prescription data were divided into three 4 month periods: period I was when the flunitrazepam prescription was not controlled, period II represented the time when flunitrazepam was placed on Schedule III and when physicians were required to use a special duplicated prescription form and period III was when the TMU-WFH started to set a stricter control for the prescription of flunitrazepam. The results indicated that the number of flunitrazepam prescriptions during period III had decreased significantly compared with period I (P < or = 0.05). Eventually, 45.7% of flunitrazepam medicated patients were followed up monthly with a restriction of their flunitrazepam supply to no more than 14 days, 22.9% of patients were followed up fortnightly at clinics with a 14 day supply of flunitrazepam, 15.7% were followed up fortnightly with a 14 day restriction of flunitrazepam plus a non flunitrazepan benzodiazepine supplement, 10.7% were referred to clinics within the Department of Psychiatry and 5% were switched from flunitrazepam to other drugs. PMID- 12109962 TI - Quality of life and its correlates in a community population in a Japanese rural area. AB - Correlations of three aspects of quality of life (QOL) (health perception, life satisfaction, and self-confidence) with personality traits and early experiences were examined. Quality of life aspects were examined using 220 inhabitants in a rural community in Japan. Health perception was better among men than among women. Life satisfaction and self-confidence were better in people aged 55 or over than in those under 55. Among the current predictor variables, the Eysenck Personality Questionnaire neuroticism score was correlated with poor life satisfaction in the younger women; the extraversion score with the older women's health perception, the older men's life satisfaction, and the women's self confidence; and the psychoticism score with the older men's life satisfaction. Among early life predictors, self-confidence was lower among those older men who had reported early parental loss. Childhood paternal overprotection was correlated with poor health perception in younger people and with good health perception in older women. Some negative life events experienced during childhood were correlated with poorer QOL measures in some subgroups, while positive life experiences were correlated with the older women's life satisfaction. These findings suggest that the three aspects of the QOL are discrete in their psychosocial correlates and that interventions on health education and care should take into account individual's psychosocial attributes. PMID- 12109963 TI - Minnesota Multiphasic Personality Inventory profile characteristics of schizotypal personality disorder. AB - The goal of the present study was to determine whether precursors for psychopathology can be found in personality dimensions of the general population. Two hundred and 62 university students were compared with 41 schizophrenic patients and 18 patients with schizotypal personality disorder (SPD) on the Minnesota Multiphasic Personality Inventory (MMPI). Schizotypal personality disorder patients showed significantly elevated Pt and Si scales compared with the schizophrenic patients. Schizophrenia and SPD groups generally produced two point codetypes of 6-8/8-6, 2-6/6-2, 7-8/8-7, and 7-8/8-7, 2-7/7-2, 6-8/8-6. A total of 77.5% of students had no codetype with a T-value of > or = 70, although the frequency of codetypes of spike 5, spike 0 and 2-7/7-2 was relatively high in the student group compared with the general population. Discriminant function analysis of the MMPI profiles revealed significant variance among the three groups. The overall rate of correct classification of the subjects into schizophrenia, SPD or university students was 90.3%. The first coefficient, mainly defined by a negative weight on the Sc scale, best distinguished the patients with either schizophrenia or SPD from the students. The second coefficient, defined by negative weights on the Sc and Si scales, and positive weights on the F and Ma scales identified patients with schizophrenia and SPD patients. The Harris-Lingoes subscales, which are supposed to provide the profile patterns characteristic of schizotypy, well discriminated the three groups. These results suggest the usefulness of the MMPI subscales for the detection of subjects with the SPD trait. PMID- 12109964 TI - Human leukocyte antigen alleles in patients with bipolar disorder in the Korean population. AB - We performed an association study between human leukocyte antigen (HLA) alleles and bipolar disorder to evaluate the potentiality of HLA as a genetic marker in bipolar disorder. HLA class I and class II allele frequencies were assessed in 87 bipolar patients and were compared with those of 206 normal controls in the Korean population. HLA class I typing was performed using the microlymphocytotoxicity method, whereas class II (DRB1 and DQB1) genotyping was performed with polymerase chain reaction-sequence specific oligonucleotide probes. When the allele frequency of HLA in bipolar patients was compared with that in normal controls, there were some significant differences. Bipolar patients showed statistically significant increased allele frequencies of HLA-A29 and B54. Allele frequencies of HLA-B51 and DRB1*02 were significantly higher in normal controls. However, these results were no longer significant after correcting for the number of alleles. The results of the present study suggest that HLA alleles may not confer susceptibility to bipolar disorder in the Korean population. To clarify the genetic influence of HLA on bipolar disorder, we should conduct a consecutive study with a larger cohort of subjects. PMID- 12109965 TI - Characteristics of women using a mental health clinic in a gynecologic out patient setting. AB - A special mental health clinic was set up in the gynecologic out-patient setting of a medical center. The present study examined the demographic and clinical characteristics of patients seen at the clinic and evaluated the need for mental health services in a gynecologic setting. Participants were 136 consecutive patients who visited the clinic during a 6 month period. Their ages ranged from 20 to 74 years, with a mean age of 41.5 +/- 12.3 years. Twenty-three percent of women were referred by gynecologists. All subjects were interviewed by experienced psychiatrists using the structured Mini-International Neuropsychiatric Interview (MINI), supplemented by the DSM-IV criteria for premenstrual dysphoric disorder (PMDD) and other diagnoses. The most common diagnosis was major depressive disorder (36.0%), followed by generalized anxiety disorder (29.4%), PMDD (16.2%), dysthymic disorder (14.7%) and others. Patients were categorized into four major diagnostic categories: depressive disorders, anxiety disorders, PMDD and others. No significant differences were found in years of education, employment status, living situations or referral pattern among the four major diagnostic groups. Most patients with anxiety disorders were married. Our results suggest that the gynecologic department may be a good setting to help women with mental disorders. PMID- 12109966 TI - No evidence for an association between the 5-hydroxytryptamine 5-HT2a receptor gene and schizophrenia in Kuwaiti Arabs. AB - The prevalence of T102C polymorphism of the 5-hydroxytryptamine 5-HT2a receptor gene has been investigated using a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method in 80 Kuwaiti Arabs with schizophrenia and in 109 normal healthy controls with a similar ethnic background. There was no significant difference in the frequency of T102 polymorphism in the Kuwaiti cohort of schizophrenia patients and the controls (P = 0.23). The data from Kuwaiti Arabs (although our sample size is relatively small) support the findings from some other populations (Caucasians, Japanese), in which a lack of association has been found between T102C polymorphism and the onset of schizophrenia. PMID- 12109967 TI - Association of the Ser9Gly polymorphism in the dopamine D3 receptor gene with tardive dyskinesia in Korean schizophrenics. AB - Tardive dyskinesia (TD) is usually regarded as one of the most serious side effects of the long-term usage of neuroleptics due to its high prevalence and potentially irreversible nature. Previously, several genetic polymorphisms were investigated for an association with TD in various ethnic populations. Among them, the Ser9Gly variant in the MscI restriction site of the dopamine D3 receptor gene was reported to be associated with TD. We have investigated the association of Ser9Gly polymorphism of the dopamine D3 receptor gene with TD in Korean schizophrenics. The frequencies of the genotypes of Ser/Ser, Ser/Gly and Gly/Gly in 54 schizophrenic patients without TD were 21 (38.9%), 33 (61.1%) and 0 (0%), while the corresponding frequencies in 59 schizophrenic patients with TD were 25 (42.4%), 28 (47.5%) and 6 (10.1%). We have found a significant genotypic association of the Gly/Gly genotype with TD in Korean schizophrenics (P = 0.028, two-tailed Fisher's exact test). However, there was no significant allelic association of the Ser9Gly allele with TD (chi2 = 0.288, d.f. = 1, P = 0.591) and there was no significant difference in the Abnormal Involuntary Movement Scale score between the three genotypic groups (P = 0.071, anova). In conclusion, we suggest that Gly/Gly homozygotes in the MscI polymorphic site of the dopamine D3 receptor gene may cause some change in the function of the dopamine D3 receptor and may be involved the pathogenesis of TD. PMID- 12109968 TI - Long-term rectal temperature measurements in a patient with menstrual-associated sleep disorder. AB - The international classification of sleep disorders has proposed menstrual associated sleep disorder. However, few studies have investigated its pathophysiological mechanism. A 34-year-old woman complaining of insomnia in the late luteal phase underwent continuous rectal temperature measurements and simultaneous actigraphic monitoring for 146 days. The acrophase of rectal temperature rhythm was delayed in the luteal phase, compared with that in the follicular phase. Her bedtime and risetime did not differ across the menstrual cycle. These results suggest that her insomnia in the luteal phase may have been a consequence of desynchronization between her temperature rhythm and sleep phase in the luteal phase. PMID- 12109969 TI - Extrapyramidal side-effect due to drug combination of risperidone and donepezil. PMID- 12109973 TI - Developing research questions in medical education: the science and the art. PMID- 12109970 TI - Improving depth and maintenance of sleep with intramuscular calcitonin: A case report. PMID- 12109975 TI - A problem shared. PMID- 12109976 TI - Facing up to the realities of global medical education in the 21st century. PMID- 12109977 TI - Point: Global standards in medical education - what are the objectives? PMID- 12109978 TI - Global standards in medical education - an instrument in quality improvement. PMID- 12109979 TI - The global pipeline: too narrow, too wide or just right? AB - BACKGROUND: Access to a well-trained workforce is one of many factors underscoring the global health divide that separates industrialized and developing nations. This paper describes the distribution and physician output of the world's medical schools, compares regional physician to population ratios, examines population trends and points out potential mismatches between output and anticipated demographic changes. METHOD: This paper has used multiple data sources in published and electronic form from organized medicine, international health institutions and the medical literature. In addition, a stratified, random survey of 130 medical schools was conducted to determine annual numbers of graduates. RESULTS: Tracking the number and distribution of medical schools and their student capacity is a complex task. The number of medical schools and the estimated number of graduates per population vary by region. In areas of predicted substantial population growth, the production of physicians is neither adequate to meet future needs, nor sufficient to overcome low physician population ratios. Regions with high physician-population ratios and either expected population decline or small population gains over the next 50 years appear to have an over-capacity to train medical students. CONCLUSION: This paper emphasizes the need for new methods of tracking the global pipeline of medical education and of establishing ways of sharing expertise. The growing interdependence of nations, accentuated by globalization of the world's economies, our shared goal of achieving health for all and the migration of physicians across borders highlight the need to understand the global capacity to educate the next generation of physicians. PMID- 12109980 TI - A pre-employment programme for overseas-trained doctors entering the Australian workforce, 1997-99. AB - OBJECTIVES: Overseas-trained doctors (OTDs) have limited access and formal interaction with the Australian health care system prior to joining the Australian medical workforce. A pre-employment programme was designed to familiarize OTDs with the Australian health care system. METHOD: All OTDs who had passed their Australian Medical Council (AMC) exams and were applying for a pre registration year in New South Wales were invited to participate in the voluntary, free programme. A 4-week full-time programme was developed consisting of core group teaching and a hospital attachment. The curriculum included communication, health and workplace skills; and sessions on culture shock and the role of junior doctors. A pilot programme was run in 1997. The programme was repeated in 1998 and 1999. The OTDs' confidence regarding the general duties of internship, and attitudes towards hospital workplace skills were examined. RESULTS: The 66 OTDs reported greater understanding of staff and communication issues and familiarization with the hospital environment. They reported a more realistic understanding of the role of a junior doctor, the need for separation of workplace and personal responsibilities and knowledge of pathways for future professional development. The course structure, with a focus on hospital attachments, establishment of a peer network, and workplace familiarization facilitated entry into the hospital workforce. CONCLUSION: The pre-employment programme enabled the OTDs to have a more equitable entry into the public hospital system, resulting in a more integrated, confident and functional workforce. PMID- 12109982 TI - Psychiatric education: a survey of Greek trainee psychiatrists. AB - OBJECTIVE: This survey investigates psychiatric trainees' attitudes regarding specific educational fields, in order to detect possible divergence from the priorities specified by the European Board of Psychiatry (EBP). It also explores certain factors which influence these attitudes. METHOD: A structured questionnaire containing 80 randomized educational objectives, which corresponded to 18 basic areas of psychiatric knowledge and competence, was given to 60 psychiatric trainees in 12 psychiatric training institutions in Greece. For the analysis of data, we used descriptive statistics, correlations, and one-way analysis of variance. RESULTS: Despite the variance which existed, the majority of trainees considered nearly all the fields in question as essential for their training, with great priority given to psychopharmacology and biological psychiatry by 91.7% and 91.4% of the trainees, respectively. On the other hand, we found that stage of training, gender, training centre and medical school of graduation played an important role in the formation of the trainees' priorities. CONCLUSIONS: The investigation of trainees' attitudes towards educational fields in psychiatry can be very useful in the development of a training plan, and at the same time constitute a valuable index for monitoring the responsiveness of training programmes to the European guidelines for psychiatric training. PMID- 12109983 TI - No debt for medical students in Greece. AB - BACKGROUND: Medical education in Greece is government-funded by constitution. Tuition fees are therefore not payable by students. This investigation aims to identify the sources of income of medical students in Greece. METHODS: This is a questionnaire-based analysis of a sample of 488 students who were asked to state their sources of income. RESULTS: Mean contributions to total student income are parental financial support (93.2%), part-time employment (2.4%), scholarships (2.5%) and other sources including loans (1.9%). CONCLUSION: The income of medical students in Greece is mostly dependent on parental contributions. Other sources do not represent popular options. This situation differs to that in other countries, where loans make significant contributions to the total income of medical students. PMID- 12109984 TI - An innovative model for teaching and learning clinical procedures. AB - CONTEXT: Performing a clinical procedure requires the integration of technical clinical skills with effective communication skills. However, these skills are often taught separately. OBJECTIVES: To explore the feasibility and benefits of a new conceptual model for integrated skills teaching. : Design A qualitative observation and interview-based study of undergraduate medical students. METHODOLOGY: Medical students performed technical and communication skills in realistic clinical scenarios (urinary catherization and wound closure), using latex models connected to simulated patients (SPs). Procedures were observed, videorecorded and assessed by tutors from an adjoining room. Students received immediate feedback from tutors and SPs, before engaging in a process of individual feedback through private review of their videotapes. Group interviews explored the response of students, SPs and tutors. Data were analysed using standard qualitative techniques. SUBJECTS: Fifty-one undergraduate students were recruited from the Faculty of Medicine, Imperial College, London. RESULTS: The scenarios provided a realistic simulation of two common clinical situations and proved feasible in terms of time, facilities and resources within this institution. Students found the opportunity to integrate communication and technical skills valuable, challenging and an appropriate learning experience. Immediate feedback was especially highly valued. Some students found difficulty integrating technical and communications skills, but benefited from conducting two procedures in the same session. CONCLUSION: The integrated model was feasible and was perceived to be valuable. Benefits include the opportunity to integrate, within a safe environment, skills which are often taught separately. Promoting reflective practice may enable the successful transfer of these integrated skills to other procedures. PMID- 12109985 TI - Improving learning of a clinical skill: the first year's experience of teaching endotracheal intubation in a clinical simulation facility. AB - BACKGROUND: All medical practitioners should be able to manage the airway of an unconscious patient. Endotracheal intubation is the most effective method of securing the airway but is a complex skill requiring much practice. Traditionally, endotracheal intubation has been taught on patients, but this is not ideal. METHODS: We have developed a short course on endotracheal intubation taught in a clinical simulation unit (CSU). This unit has a large range of airway trainers and patient simulators, some of which can be manipulated to make intubation more difficult. Endotracheal intubation is taught in a series of steps in order to avoid cognitive overload. Each step is taught on an airway trainer that has no difficult features. Once this is mastered, more difficult situations are presented which require application of new techniques and/or equipment. In this way, students learn useful schemas to apply clinically. RESULTS: In 1 year, over 100 students and trainees were taught endotracheal intubation in the CSU. The ideal group size was found to be two students and one trainer. It took 75 to 90 minutes for most students to reach a standard where they could be expected to safely perform the technique on a patient. All comments on learning endotracheal intubation in this setting were positive. Many students felt more comfortable learning on a model than on a patient. CONCLUSION: Learning clinical procedures on simulators is becoming an essential part of medical education. More than one airway trainer may be needed to give students the expertise to perform endotracheal intubation on patients. PMID- 12109986 TI - Support for students with academic difficulties. AB - CONTEXT: The human and financial costs of academic failure amongst medical students are extremely high. Often, remedial support is infrequently available or is available only for students failing their final examinations. We describe the design, implementation and preliminary evaluation of a remedial programme (RP) for students who experience academic difficulties. METHODS: A total of 24 students were identified from Years 4 and 5 of the undergraduate medical course at our medical school. Students invited to participate were identified following failure in summative and/or continuous assessment processes. Students underwent an individual educational diagnosis by means of free discussion and a semi structured interview. Each negotiated a problem list, action plan and learning contract with course tutors. Of the 24 students, 16 received academic support and tutorials. Tutorial content was determined by individual students and took place over a 6-12-month period, initially individually and subsequently in pairs. Regular feedback and appraisal occurred. Information was delivered to the medical school academic progress review system. Student satisfaction was investigated and subsequent examination scores reviewed. RESULTS: The causes of academic failure were widespread and ranged from deficient study skills to financial, domestic and emotional problems. In contrast, the subjects in which students had difficulty were remarkably similar, in that they all related to core clinical skills and frequently involved communication skills. Students enjoyed the programme, reported improved motivation both for study and for their chosen career and demonstrated a greatly improved pass rate in subsequent examinations. CONCLUSIONS: The causes of academic failure in undergraduate medical students are diverse and are often not academic in origin. Students can benefit from an individually tailored remedial programme and go on to success in subsequent parts of the curriculum. Provision of individually tailored remedial teaching is labour intensive and requires full faculty support. PMID- 12109987 TI - The Pastoral Pool: an evaluation of a new system of pastoral care provision. AB - CONTEXT: Medical students experience significant stress and stress-related problems. The benefits of support during training are described in this paper. Recently, student support systems have become increasingly stretched as medical schools merge, courses become modular and staff face increasing clinical and research pressures. The pastoral support system at St Bartholomew's and the Royal London School of Medicine has been changed in response to an evaluation of the old system of personal tutors. Pastoral care is now provided by a group of willing staff members known to have an interest in student welfare. The group is known as the 'Pastoral Pool' (PP). METHODS: Students and staff PP members completed similar questionnaires addressing their expectations of the PP and identifying the need for support systems in general. Pastoral Pool activity was investigated using confidential record sheets stating the frequency, duration and content of each PP encounter. RESULTS: Students were aware of the PP and many would consider using it. Staff and students agreed over the functions of the PP. Students frequently expressed concerns over confidentiality within the system. Uptake was low, with only 3% of students approaching the PP and the majority of PP work performed by only two staff members. The content of meetings was often multifactorial and usually on a subject within the PP remit. CONCLUSIONS: The PP is beneficial because pastoral care is provided only by dedicated staff members with an interest in student welfare. Membership is small enough to enable staff training and development. However, the system relies on self-referral by students. Rather than preventing problems arising, it therefore provides secondary support to those students insightful enough to identify their problems and motivated enough to seek help. PMID- 12109988 TI - Sci45: the development of a specialty choice inventory. AB - OBJECTIVE: To devise a valid career selection instrument to help doctors in training choose from a range of specialties that match their attributes and aspirations and to help selection interviewers focus on the key issues pertaining to the suitability of candidates for particular training opportunities. DESIGN: A psychometric instrument of 130 4-response choice items was developed to match individual personal and professional preferences to possible career specialty choices. The development process involved semi-structured interviews with consultants in 35 specialties, a national postal survey of consultants in 45 specialties, factor analysis of the results, design of the pilot instruments, testing on 450 senior house officers (doctors in basic specialist training within 2-5 years of leaving medical school), and further item analysis to derive the final instrument. A scoring system and software were developed to indicate the best and worst fit specialties for the respondent. PARTICIPANTS: The participants were hospital consultants, general practitioners and senior house officers (SHOs) in basic specialist training. OUTCOME MEASURE: The successful construction of a valid and accessible career choice instrument (Specialty Choice Inventory/Sci45). CONCLUSIONS: This project has yielded a psychometrically valid computer- or paper based instrument that can be used by doctors at any stage of training to assist in career choice. It can be used as part of the selection process, for careers guidance, for analysis of career problems, for research or to validate a particular range of career options. PMID- 12109990 TI - Interactive logbooks for medical students: are they useful? AB - OBJECTIVES: The use of logbooks in the education of medical undergraduate students is not well-established. Traditionally, logbooks are used simply as a means for students to document their activities. This report examines whether logbooks used as an interactive vehicle between students and tutors can assist both student learning and Faculty teaching. METHOD: As part of the New Medical Curriculum implemented by the Faculty of Medicine, The University of Hong Kong, all third year students beginning their formal hospital and community health clerkships were given pocket-sized logbooks to document and monitor their learning activities. The logbooks were specially designed to mirror the activities of the teaching blocks, including bedside teaching, tutorials, teaching clinics, health care projects, and whole class sessions, etc. RESULTS: At the end of each teaching block, effort, accuracy of the notes, appropriateness of the notes and the assessor's overall impression of logbook entries formed the basis of 20-point assessment. Randomly-selected logbooks were reviewed at the end of every rotation and compared with course outlines in order to evaluate if, according to the students' notes, the learning objectives were being met. Throughout each teaching block, the logbook process identified students who could benefit from counselling and/or remediation. The logbook feedback mechanism was immediate and therefore, remediation was timely and appropriate. CONCLUSIONS: The logbooks were effective in 3 ways: logbooks were a means of continuous assessment of small group learning; logbooks encouraged immediate and ongoing interaction between tutors and students; and they provided a feedback loop for the evaluation of learning activities. PMID- 12109989 TI - Appraisal of doctors: problems with terminology and a philosophical tension. AB - The term appraisal lacks useful definition. It is used to describe both the summative assessment which forms part of performance management, as well as an educational process sometimes referred to as formative assessment. In this analysis we trace the purpose of each of these components as applied to career grade doctors. In the first, authority for the process, and the motive for applying it, lies with management. Within a publicly funded health service the purpose of this is directed towards equality of health care, and to obtain the greatest performance available with the resources available. By contrast, appraisal which supports a doctor's self-directed learning is likely to address issues of quality. The two processes exist in tension with each other, and are mutually informative. We argue that in the appraisal of doctors the two processes and purposes should be made explicit. PMID- 12109991 TI - Developing a new line of patter: can doctors change their consultations for sore throat? AB - BACKGROUND AND OBJECTIVES: Doctors report pressure from peers to reduce prescribing of antibiotics for minor respiratory illnesses, and from patients to do the opposite. It has been suggested that doctors adopt a more patient-centred consulting style in order to encourage patient satisfaction and shared decision making. No evidence exists that such changes are achievable. We developed a new, on-site method for training postgraduates and used this for teaching patient centred intervention. Here, we examine whether this training method is associated with changes in consulting patterns in consultations for sore throat with children, among doctors from a single group practice. METHODS: Audiotaped consultations (simulated and real) conducted before and after training were analysed and interviews were carried out with participants about the impact of training. SETTING: A general practice in South Wales. PARTICIPANTS: Four general practitioners who consulted with 25 real and simulated patients participated in the study. MAIN OUTCOME MEASURES: Four patient-centred skills used by doctors and 2 patient behaviours measured before and after training were identified. RESULTS: Three out of 4 practitioners produced clear evidence of changes in patient centred consulting skills. These changes were evident in simulated and real consultations 2 and 4 weeks later, respectively. Prior to training the doctors produced only five examples of patient-centred skills in 10 consultations. After training they produced 39 examples in 15 consultations. CONCLUSIONS: Evidence from both consultations and interviews indicated that the intervention and training were well received and had been put into practice. PMID- 12109996 TI - Professor Georges Bordage. PMID- 12109997 TI - Update on vitamin D and its newer analogues: actions and rationale for treatment in chronic renal failure. AB - Vitamin D is an important hormone for mineral homeostasis and the proper formation and maintenance of bone. In addition, vitamin D has broader functions in the body that expand its traditionally known role in mineral balance. In chronic renal failure, calcitriol deficiency contributes to the development and progression of secondary hyperparathyroidism, bone disorders, and altered mineral metabolism. Recent revelations of the broader role of vitamin D also suggest calcitriol deficiency may contribute to decreased cardiac and immune function in chronic renal failure patients. Research on vitamin D has led to a more complete understanding of the actions of vitamin D at the transcriptional level and with respect to the clinical use of vitamin D and its analogs to control parathyroid hormone overactivity and to replace the other D hormone-dependent actions in patients with renal failure. Limitations of vitamin D and its metabolites include hypercalcemia, hyperphosphatemia and suppression of bone turnover with the risk of adynamic bone disease. Vitamin D analogs may offer greater selectivity and potentially greater safety as compared to calcitriol because of their altered relative potency on calcium and phosphorus metabolism. This review focuses on the current understanding of the biological actions of vitamin D and its analogs and the rationale for treating patients with chronic renal failure. PMID- 12109998 TI - Functional domains in the renal type IIa Na/P(i)-cotransporter. AB - The proximal tubular brush border membrane type IIa Na/P(i)-cotransporter is an important element in overall phosphate (Pi) homeostasis. Its regulation is tightly associated with membrane retrieval/reinsertion mechanisms. Specific molecular domains are involved in its internalization (predicted third intracellular loop) and in its apical expression (carboxy-terminus). Regulation and apical expression require a correct ('proximal tubular') cellular context and interaction with specific cellular proteins (scaffolding). Basic cotransport function is via a 3 Na+ to 1 P(i)-coupling ratio, also including the possibility of a Na+-leak, and is strongly affected by changes in pH. This function can be assigned to monomeric transporter molecules. The predicted first intracellular and third extracellular loops contribute important functional characteristics. It is suggested that they may form "re-entrant loops" and thereby a "permeation pore." Sequences in this region determine also pH-sensitivity and affinities in P(i)- and in Na+-interaction, respectively. PMID- 12109999 TI - Novel collagen glomerulopathy in a homotrimeric type I collagen mouse (oim). AB - BACKGROUND: Oim/oim mice [osteogenesis imperfecta model; homozygous null for the proalpha2(I) collagen gene] synthesize exclusively the homotrimeric type I collagen isotype, alpha1(I)3, and are unable to synthesize the normal heterotrimeric type I collagen isotype, alpha1(I)2alpha2(I). Previous studies of the oim/oim mouse have focused on the musculoskeletal system, with no systematic evaluation of other organ systems. METHODS: Multiple tissues from oim/oim, oim/+ (heterozygous) and +/+ (wild-type) mice were examined for gross and histological abnormalities. Tissues were stained with (1) hematoxylin and eosin (to assess lesion formation), (2) picrosirius red (collagen content), and (3) periodic acid methenamine silver (basement membrane). Kidneys were further evaluated ultrastructurally by electron microscopy and immunohistochemically with anti alpha1(I) and anti-alpha1(III) collagen antibodies. RESULTS: Histological analyses revealed accumulations of picrosirius red-positive material, consistent with collagen, in glomeruli of 28/29 oim/oim mice, with no evidence of mesangial cell proliferation. Only the most severely affected animals had evidence of increased capillary basement membrane thickening or mild inflammation around the affected glomeruli. Electron microscopy confirmed the presence of fibrillar collagen. Immunohistochemistry with anti-alpha1(I) collagen antibodies confirmed accumulation of type I collagen in the oim/oim glomeruli. The +/+ and oim/+ kidneys had normal mesangium with no evidence of infiltration of collagenous material. CONCLUSIONS: This study demonstrates the first evidence, to our knowledge, of abnormal glomerular collagen deposition associated with a type I collagen defect. Further in vivo and in vitro studies are necessary to elucidate the mechanistic, functional, and pathological significance of the oim/oim collagen glomerulopathy. PMID- 12110000 TI - Potential mechanisms of marked hyperoxaluria not due to primary hyperoxaluria I or II. AB - BACKGROUND: Hyperoxaluria may be idiopathic, secondary, or due to primary hyperoxaluria (PH). Hepatic alanine:glyoxylate aminotransferase (AGT) or glyoxylate/hydroxypyruvate reductase (GR/HPR) deficiency causes PHI or PHII, respectively. Hepatic glycolate oxidase (GO) is a candidate enzyme for a third form of inherited hyperoxaluria. METHODS: Six children were identified with marked hyperoxaluria, urolithiasis, and normal hepatic AGT (N = 5) and GR/HPR (N = 4). HPR was below normal and GR not measured in one. Of an affected sibling pair, only one underwent biopsy. GO mutation screening was performed, and dietary oxalate (Diet(ox)), enteric oxalate absorption (EOA) measured using [13C2] oxalate, renal clearance (GFR), fractional oxalate excretion (FE(ox)) in the children, and urine oxalate in first-degree relatives (FDR) to understand the etiology of the hyperoxaluria. RESULTS: Mean presenting age was 19.2 months and urine oxalate 1.3 +/- 0.5 mmol/1.73 m2/24 h (mean +/- SD). Two GO sequence changes (T754C, IVS3 - 49 C>G) were detected which were not linked to the hyperoxaluria. Diet(ox) was 42 +/- 31 mg/day. EOA was 9.4 +/- 3.6%, compared with 7.6 +/- 1.2% in age-matched controls (P = 0.33). GFR was 90 +/- 19 mL/min/1.73 m2 and FE(ox) 4.2 +/- 1.4. Aside from the two brothers, hyperoxaluria was not found in FDR. CONCLUSIONS: These patients illustrate a novel form of hyperoxaluria and urolithiasis, without excess Diet(ox), enteric hyper-absorption, or hepatic AGT, GR/HPR deficiency. Alterations in pathways of oxalate synthesis, in liver or kidney, or in renal tubular oxalate handling are possible explanations. The affected sibling pair suggests an inherited basis. PMID- 12110001 TI - Nitric oxide down-regulates connective tissue growth factor in rat mesangial cells. AB - BACKGROUND: Nitric oxide (NO) exerts complex regulatory actions on mesangial cell (MC) biology, such as inhibition of proliferation, adhesion or contractility and induction of apoptosis. In our previous studies the NO-donor S-nitroso glutathione (GSNO) was found to be a potent inhibitor of MC growth. This effect was mediated at least in part by inhibitory effects of GSNO on the transcription factor early growth response gene-1 (Egr-1) [10]. We therefore were interested in the regulation of gene expression in MC after treatment with NO. METHODS: To identify the genes that are regulated by NO in MC, gene expression was analyzed by representational difference analysis. Expression of connective tissue growth factor (CTGF) was studied by Northern and Western blot analyses. RESULTS: Cultured rat MCs treated with GSNO for 8 hours were compared with unstimulated MCs and the CTGF mRNA was found to be down-regulated. The down-regulation was dose-dependent and transient, with a maximum inhibition seen after 6 hours. In parallel, down-regulation of CTGF protein by GSNO was observed by Western blot analysis. Other NO-donors such as S-nitroso-N-acetyl-D,L-penicillamine and spermine-NO showed similar effects. The induction of the inducible NO-synthase by TNF-alpha, IL-1beta and LPS provoked a transient down-regulation of CTGF mRNA, an effect that could be partially overcome by pretreatment with the NOS-inhibitor Nomega-nitro-l-arginine methyl ester. The observed NO-effect could be simulated by treatment with the stable cGMP analog 8br-cGMP, and was abolished by blocking the guanylyl cyclase with the inhibitor NS2028. CONCLUSION: NO acts as a strong repressor of CTGF expression in cultured rat MC. Thus, in addition to its antiproliferative effects, NO potentially exerts antifibrotic activity by down regulation of CTGF. PMID- 12110002 TI - Inhibition of IGF-I-induced Erk 1 and 2 activation and mitogenesis in mesangial cells by bradykinin. AB - BACKGROUND: The beneficial effects of therapeutic angiotensin-converting enzyme (ACE) inhibitor treatment against the worsening of glomerulosclerosis during the course of diabetic nephropathy have been widely documented. ACE inhibitors inhibit both angiotensin II formation and bradykinin (BK) degradation, thereby reducing angiotensin II type 1 (AT1) receptor activity and favoring B2-kinin receptor (B2 receptor) activation. Since the involvement of growth factors such as insulin-like growth factor (IGF-I) has been implicated in the early steps of diabetic nephropathy, we investigated the effect of BK on Erk 1 and 2 activation and cell proliferation by IGF-I. METHODS: The activation of Erk 1 and 2 in mesangial cells (MCs) and isolated glomeruli (IG) was investigated by immunoprecipitation and Western blotting during activation of the IGF-I receptor in the presence or absence of BK and of protein kinase C (PKC), tyrosine-kinase and phosphatase selective inhibitors. Mesangial cell proliferation was assessed in vitro by cell counting. RESULTS: In untreated MCs and IG, when added separately, BK and IGF-I both activated Erk 1 and 2. In contrast, in MCs and IG pretreated with BK, the IGF-I-induced Erk 1 and 2 activation was dose-dependently reduced. The inhibitory effect of BK on IGF-I-induced activation of Erk 1 and 2 was completely abolished by addition of a B2 antagonist, by chelation of intracellular calcium and by tyrosine phosphatase inhibition. Additionally, BK reduced MC proliferation induced by IGF-I. CONCLUSIONS: A new inhibitory pathway of the early steps of IGF-I signaling by the B2 receptor is found both in cultured MCs and in IG, which involves a calcium-dependent tyrosine phosphatase activity. Recruitment of this mechanism may account for the beneficial effects of ACE inhibitor treatment on glomerulosclerosis associated with diabetic nephropathies. PMID- 12110003 TI - Angiotensin II regulation of vascular endothelial growth factor and receptors Flt 1 and KDR/Flk-1 in cyclosporine nephrotoxicity. AB - BACKGROUND: Vascular endothelial growth factor (VEGF) is involved in angiogenesis, wound healing and inflammation. VEGF exerts its effect via the tyrosine kinase receptors Flt-1 and KDR/Flk-1. We have previously shown that VEGF is up-regulated in a chronic cyclosporine (CsA) nephrotoxicity model. Our current study examined the role of angiotensin II (Ang II) blockade with enalapril (E) or losartan (L) on VEGF in this model. METHODS: Pair-fed salt-depleted rats were administered vehicle, CsA, CsA + nilvadipine, CsA + hydralazine/hydrochlorthiazide (HCTZ), CsA + E or CsA + L, and were sacrificed at 7 or 28 days. Physiologic and histologic changes were studied in addition to the mRNA expression of VEGF and its receptors Flt-1 and KDR/Flk-1 by Northern blot, and the protein expression of VEGF by Western blot. RESULTS: While all groups achieved similar blood pressures and creatinine clearances, the amelioration in nephrotoxicity was observed only with Ang II blockade. VEGF mRNA and protein expressions increased with CsA and became significantly reduced with Ang II blockade. Flt-1 expression was similar in all groups; it decreased early and remained low. On the other hand, KDR/Flk-1 mRNA expression was higher at seven days in all groups, except in the +E and +L groups where it was significantly lower, and then became further down-regulated at 28 days. CONCLUSIONS: The increased VEGF expression in chronic CsA nephrotoxicity seems to be related to up regulation of Ang II. In addition, VEGF probably exerted its effect via the KDR/Flk-1 receptor. The actions of VEGF in this model remain speculative, but may be related to its effect on macrophage infiltration or matrix deposition. PMID- 12110004 TI - The luminal membrane of rat thick limb expresses AT1 receptor and aminopeptidase activities. AB - BACKGROUND: Endogenous intratubular angiotensin II (Ang II) supports an autocrine tonic stimulation of NaCl absorption in the proximal tubule, and its production may be regulated independently of circulating Ang II. In addition, endogenous Ang II activity may be regulated at the brush border membrane (BBM), by the rate of aminopeptidase A and N (APA and APN) activities and the rate of Ca2+-independent phospholipase A2 (PLA2-dependent endocytosis and recycling of the complex Ang II subtype 1 (AT1) receptor (AT1-R). The aim of the present study was to look for subcellular localization of AT1-R, and APA and APN activities in the medullary thick ascending limb of Henle (mTAL), as well as search for an asymmetric coupling of AT1-R to signal transduction pathways. METHODS: Preparations of isolated basolateral membrane (BLMV) and luminal (LMV) membrane vesicles from rat mTAL were used to localize first, AT1-R by 125I-[Sar1, Ile8] Ang II binding studies and immunoblot experiments with a specific AT1-R antibody, and second, APA and APN activities. Microfluorometric monitoring of cytosolic Ca2+ with a Fura-2 probe was performed in mTAL microperfused in vitro, after apical or basolateral application of Ang II. RESULTS: AT1-R were present in both LMV and BLMV, with a similar Kd (nmol/L range) and Bmax. Accordingly, BLMV and LMV preparations similarly stained specific AT1-R antibody. APA and APN activities were selectively localized in LMV, although to a lesser extent than those measured in BBM. In the in vitro microperfused mTAL, basolateral but not apical Ang II induced a transient increase in cytosolic [Ca2+]. CONCLUSIONS: Besides the presence of basolateral AT1-R in mTAL coupled to the classical Ca2+-dependent transduction pathways, AT1-R are present in LMV, not coupled with Ca2+ signaling, and co-localized with APA and APN activities. Thus, apical APA and APN may play an important role in modulating endogenous Ang II activity on NaCl reabsorption in mTAL. PMID- 12110005 TI - Activin A: an autocrine regulator of cell growth and differentiation in renal proximal tubular cells. AB - BACKGROUND: Activin A is involved in tubular regeneration after ischemia/reperfusion injury. The present study was conducted to examine the role of activin A in cell growth, apoptosis and differentiation of tubular cells. METHODS: We performed cell proliferation assays (MTT assay, [3H]-thymidine incorporation) and apoptosis detection assays (nuclear staining, DNA ladder formation, TUNEL staining) using LLC-PK1 cells. Expression of activin and activin receptor in LLC-PK1 cells also were examined by real-time polymerase chain reaction (PCR) and immunostaining. Stable cell lines expressing the truncated type II activin receptor were generated and the phenotype of these cells was analyzed. RESULTS: Activin A inhibited DNA synthesis and cell growth in a dose dependent manner and induced apoptosis in LLC-PK1 cells. The expression level of mRNA for the activin betaA subunit was markedly increased when the growth was stimulated. The expression of the type II activin receptor was observed in LLC PK1 cells. The growth rate of cells expressing dominantly negative activin receptor was significantly faster than that of non-transfected cells. The expression level and pattern of cytokeratin and vimentin in these cells were quite different compared to non-transfected cells. When cultured in collagen gel, these cells formed multiple processes, which was not observed in non-transfected cells. Finally, the expression of Pax-2 was markedly elevated in these cells. CONCLUSIONS: Activin A acts as an autocrine inhibitor of cell growth, an inducer of apoptosis, and an important modulator of differentiation in cultured proximal tubular cells. PMID- 12110006 TI - Effects of different PPARgamma-agonists on MCP-1 expression and monocyte recruitment in experimental glomerulonephritis. AB - BACKGROUND: Activators of peroxisome proliferator activated receptor gamma (PPARgamma) have been shown to modulate chemokine expression in isolated monocytes/macrophages (M/M) and to exert anti-inflammatory effects in some models of experimental inflammatory diseases. We evaluated the effects of different forms of PPARgamma activators in a model of experimental glomerulonephritis (GN) in rats. METHODS: GN was induced in rats by application of an anti-thymocyte antibody (ATS). Nephritic rats were treated with two synthetic PPARgamma ligands of the thiazolidinedione (TZD) group, troglitazone (200 mg/kg/day) and ciglitazone (100 mg/kg/day), and with a natural ligand 15d-PGJ2 (1.5 mg/day). Twenty-four hours after induction of the GN, the glomerular mRNA expression of the chemokine monocyte chemoattractant protein-1 (MCP-1) and the cognate chemokine receptor CCR-2 were examined by Northern blotting and RT-PCR. The glomerular M/M infiltration was determined by immunohistology. The activation of the transcription factors PPARgamma, nuclear factor-kappaB (NF-kappaB) and activator protein-1 (AP-1) in glomeruli was analyzed by electrophoretic mobility shift assay. RESULTS: Induction of GN up-regulated glomerular nuclear protein binding of NF-kappaB and AP-1. Treatment of nephritic rats with troglitazone and ciglitazone augmented nuclear PPARgamma and AP-1 DNA binding but did not affect NF-kappaB binding. TZD enhanced glomerular MCP-1 expression and increased glomerular M/M recruitment. In contrast, 15d-PGJ2 attenuated NF-kappaB activation and did not affect AP-1 activity or MCP-1 expression. CONCLUSION: Our data show that PPARgamma activators of the TZD group, but not 15d-PGJ2, enhance MCP-1 expression and M/M infiltration in the induction phase of experimental GN. The results demonstrate that TZD and 15d-PGJ2 may exert different effects in the immune response in experimental GN. Our study underscores the need to critically evaluate whether PPARgamma ligands will have beneficial or possibly deleterious effects in GN. PMID- 12110007 TI - Interactions of human mesangial cells with IgA and IgA-containing immune complexes. AB - BACKGROUND: IgA nephropathy (IgAN) is characterized by IgA1-containing immune complexes in mesangial deposits and in the circulation. The circulating immune complexes (CIC) are composed of galactose- (Gal) deficient IgA1 and IgG or IgA1 antibodies specific for the Gal-deficient IgA1; interactions of these CIC with mesangial cells (MC) were studied. METHODS: Binding, internalization, and catabolic degradation of myeloma IgA1 protein as a standard control and the isolated CIC were studied using human MC, hepatoma cell line HepG2 expressing the asialoglycoprotein receptor (ASGP-R), and monocyte-like cell line U937 expressing the Fc(alpha)-R (CD89). Biochemical and molecular approaches were used to assess expression of CD89 and ASGP-R by MC. RESULTS: At 4 degrees C, radiolabeled IgA1 bound to MC and HepG2 cells in a dose-dependent and saturable manner. The binding was inhibited by IgA-containing CIC or excess IgA1 or its Fc fragment but not by the Fab fragment of IgA1. At 37 degrees C, the cell-bound IgA1 was internalized and catabolized. In addition to IgA1, HepG2 cells also bound (in a Ca2+-dependent manner), internalized, and catabolized asialoorosomucoid (ASOR), other asialo (AS)-glycoproteins, and secretory component (SC). The binding by MC appeared to be restricted to IgA1 or AS-IgA1 and was not Ca2+-dependent. Furthermore, MC and HepG2 cells internalized and catabolized IgA1-containing CIC. Using RT-PCR with ASGP-R- or CD89-specific primers, mRNAs of the two respective genes were not detected in MC. CONCLUSIONS: The data showed that the ability of MC to bind IgA1 and IgA1-containing CIC in vitro was mediated by an IgA receptor that was different from CD89 or ASGP-R and had a higher affinity for IgA-CIC than for uncomplexed IgA. PMID- 12110008 TI - The renal segmental distribution of claudins changes with development. AB - BACKGROUND: Permeability properties of mammalian nephron are tuned during postnatal maturation. The transepithelial electrical resistance (TER) and complexity of tight junctions (TJs) vary along the different tubular segments, suggesting that the molecules constituting this structure change. We studied the differential expression of occludin and several claudins in isolated renal tubules from newborn and adult rabbits. METHODS: Isolated renal tubules from newborn and adult rabbits were processed for occludin, claudin-1 and claudin-2 immunofluorescence, and Western blot detection of claudin-1 and -2. Claudin-5 was detected in whole kidney frozen sections. RT-PCR from isolated tubules was performed for claudins-1 to -8. RESULTS: Immunofluorescence revealed that occludin, claudin-1 and -2 were present at the cell boundaries at the neonatal stage of development. Claudin-1 was detected in the tighter segments of the nephron (distal and collecting duct), while claudin-2 was found in the leaky portions (proximal). Claudin 5 was found in the kidney vasculature. PCR amplification revealed the presence of claudins-1 to -4 in tubules of newborns. In adults, claudins-1, -2 and -4 were present in proximal, Henle's loop and collecting segments; claudin-3 was in proximal and collecting tubules, while claudins-5 and -6 were absent from all tubular portions. Claudin-7 was restricted to proximal tubules, while claudin-8 was present in proximal and Henle's segments. CONCLUSIONS: The pattern of occludin distribution is present from the neonatal age. Claudins-7 and -8 are up-regulated after birth. Each tubular segment expresses a peculiar set of claudins that might be responsible for the permeability properties of their TJs. PMID- 12110009 TI - Expression of the fractalkine receptor (CX3CR1) in human kidney diseases. AB - BACKGROUND: CX3CL1 (fractalkine) is a membrane bound chemokine that can function as an adhesion molecule for cells expressing the receptor CX3CR1. This receptor is involved in the recruitment of inflammatory cells in a rat model of crescentic glomerulonephritis, where blockade of CX3CR1 has been shown to be of benefit. Here we describe the distribution of CX3CR1 positive cells in a variety of kidney diseases and renal development. METHODS: A total of 84 formalin-fixed, paraffin embedded specimens including fetal kidneys (N = 12), normal areas of kidneys uninvolved by neoplasia from tumor nephrectomies (N = 4), renal transplant nephrectomies (N = 5), renal transplant biopsies (N = 19), and kidney biopsies from patients with crescentic glomerulonephritis (N = 7), membranous nephropathy (N = 7), membranoproliferative glomerulonephritis (N = 8), focal and segmental glomerulosclerosis (N = 10), collapsing glomerulopathy (N = 6), and minimal change disease (N = 6) were studied. Immunohistochemistry was performed on consecutive tissue sections for CD3 positive T cells, CD68 positive monocyte/macrophages, CCR5 positive cells and CX3CR1 positive cells. RESULTS: The majority of inflammatory leukocytes infiltrating the kidney expressed CX3CR1. The distribution pattern was consistent with expression by both T cells and monocytes/macrophages. In contrast to the distribution of CCR5, which was expressed on a subset of infiltrating cells predominantly localized in the interstitium, CX3CR1 was present on both interstitial and glomerular infiltrating leukocytes. In developing kidneys CX3CR1 positive cells formed a small, scattered population of cells, consistent with the distribution of infiltrating leukocytes. CONCLUSIONS: The high number of CX3CR1-positive inflammatory cells in various disease entities is consistent with its having a role in the accumulation of intrarenal inflammatory cells, but does not provide evidence of specificity of leukocytes bearing this receptor for specific types of injury. Other chemokine gradients, like those created by the ligands for the chemokine receptor CCR5, might subsequently guide leukocyte subsets to specific microenvironments. PMID- 12110010 TI - Renal structural abnormalities following recovery from acute puromycin nephrosis. AB - BACKGROUND: Rats that recover from acute puromycin nephrosis later develop widespread glomerular and tubulointerstitial injury. The current study sought to identify structural changes present in the recovery phase that could precipitate progressive renal disease. METHODS: Stereologic studies were performed 10 weeks after administration of puromycin (PAN) or saline (Cont). Serial sections were examined to assess glomerular structure. RESULTS: Rats receiving puromycin developed heavy proteinuria that returned nearly to control levels at 10 weeks. Kidneys in these animals were moderately enlarged and exhibited expansion of the interstitium (PAN, 254 +/- 47 mm3; Cont, 152 +/- 23 mm3; P < 0.05). The average glomerular volume was not different from control (PAN, 1.90 +/- 0.38 x 10(6) microm3; Cont, 2.07 +/- 0.47 x 10(6) microm3), but a subpopulation of glomeruli of about half normal size was found in PAN rats. Serial sections revealed that most of these glomeruli were not connected to normal tubule segments. Serial sections also revealed that more than 90% of glomeruli in rats recovering from nephrosis had synechias joining the tuft to Bowman's capsule. Synechias occupied an average of 8 +/- 11% of the Bowman's capsule surface in PAN animals versus less than 1% of the surface in controls. The appearance of synechias was not associated with a reduction in the mean number of visceral or parietal epithelial cells per glomerulus. CONCLUSIONS: Acute puromycin nephrosis does not cause a notable reduction in visceral epithelial cell number. However, widespread glomerular injury characterized by synechia between the tuft and Bowman's capsule is present following remission of proteinuria. Progression of this residual glomerular injury could contribute to the late development of glomerular segmental sclerosis following recovery from acute nephrosis. PMID- 12110011 TI - Nitric oxide, enhanced by macrophage-colony stimulating factor, mediates renal damage in reflux nephropathy. AB - BACKGROUND: Reflux nephropathy (RN) is a major cause of end-stage renal failure in children and young adults. Nitric oxide (NO) is an important mediator of tissue injury and inflammation. NO production is enhanced by hematopoietic growth factor including macrophage colony stimulating factor (M-CSF). M-CSF plays a pivotal role in the development of nephritis via macrophage activation. The aim of this study was to investigate the expression of inducible NO synthase (iNOS) and M-CSF in the refluxing kidney, in order to further understand the pathogenesis of RN. METHODS: The kidney specimens from 6 patients with severe RN and 6 controls were examined by NADPH-diaphorase histochemistry and immunohistochemistry using ABC method with anti-M-CSF antibody. Double staining using NADPH-diaphorase histochemistry/M-CSF immunohistochemistry and M-CSF/iNOS fluorescence immunohistochemistry also was performed. In situ hybridization was performed using digoxigenin labeled M-CSF specific probe. RT-PCR was performed to evaluate the relative amount of iNOS mRNA expression. Apoptosis was determined using the in situ end-labeling technique. RESULTS: The most striking difference between tissues from RN patients and controls was the marked increase in NADPH-d activity, iNOS immunoreactivity and mRNA and M-CSF immunoreactivity and mRNA expression in the kidneys of RN patients, particularly in the distal tubules, collecting system. Apoptotic cells were markedly increased in RN compared to controls. CONCLUSIONS: Our findings suggest that the increase in M-CSF-stimulated local production of nitric oxide may be a major mediator in the development of reflux nephropathy. PMID- 12110012 TI - Down-regulation of rat organic cation transporter rOCT2 by 5/6 nephrectomy. AB - BACKGROUND: In rat kidneys, the organic ion transporters rOCT1, rOCT2, rOAT1 and rOAT3 are considered to mediate the basolateral uptake of various ionic compounds. However, their changes in chronic renal failure (CRF) are poorly understood. The present study examined the renal handling of organic ions and the expression of these transporters under CRF. METHODS: 5/6 Nephrectomized rats were used as the animal model of CRF. Renal handlings of cimetidine and paraaminohippuric acid (PAH) were examined by in vivo experiments. rOAT1, rOAT3, rOCT1 and rOCT2 expressions were determined by Western blotting. RESULTS: The tubular secretion rates of both PAH and cimetidine were markedly decreased in CRF rats. Although the distribution rates of PAH into the kidney cortex and medulla, and of cimetidine into the kidney cortex were maintained, the distribution rate of cimetidine into the kidney medulla was significantly decreased in CRF rats. The expression level of the rOCT2 protein was markedly depressed in CRF rats, but those of rOCT1, rOAT1 and rOAT3 were maintained. In addition, the plasma concentration of testosterone, a regulator of rOCT2 expression, was significantly reduced by CRF. Both the renal clearance of cimetidine and rOCT2 expression were recovered by the exogenous administration of testosterone in CRF rats. CONCLUSIONS: The levels of urinary excretion of cationic drugs, especially substrates for rOCT2, were reduced under CRF partly due to the reduced expression of rOCT2, and the lowered plasma level of testosterone was suggested to be responsible for the depressed rOCT2 expression in CRF. PMID- 12110013 TI - Role of mesangial cells and gap junctions in tubuloglomerular feedback. AB - BACKGROUND: Tubuloglomerular feedback (TGF) is a process whereby the resistance of the afferent arterioles delivering blood to the glomeruli is regulated by the NaCl concentration of the forming urine in the lumen of the macula densa. Intraglomerular mesangial cells are located between capillaries within the glomerulus, while extraglomerular mesangial cells are located between the macula densa and the afferent arteriole. They are electrically and chemically coupled via gap junctions. The purpose of this study was to investigate the role of mesangial cells and gap junctions in TGF using the isolated, perfused juxtaglomerular apparatus. METHOD: Juxtaglomerular apparatuses were dissected from male New Zealand white rabbits and perfused in vitro. The NaCl concentration at the macula densa was changed from 17/2 to 65/50 Na/Cl to initiate a TGF response. Afferent arterioles were perfused at 60 mm Hg throughout the experiment. Changes in luminal diameter caused by increasing the NaCl concentration at the macula densa were taken as the TGF response. TGF was measured before and after disrupting the gap junctions or damaging the mesangial cells in paired experiments. RESULTS: During the control period, TGF decreased afferent arteriole diameter by 2.9 +/- 0.2 microm. After mesangial cells were damaged by perfusing Thy 1-1 antibody and complement into the afferent arteriole, the TGF response was completely eliminated. Separate experiments showed no statistically significant change in TGF response with time, or when antibody and complement were perfused into the macula densa lumen. The presence of Thy 1-1 antibody and complement in the afferent arteriole perfusate did not alter the ability of norepinephrine to constrict or acetylcholine to dilate the afferent arteriole. To investigate the role of gap junctions in TGF, we used heptanol to disrupt them. During the control period, TGF decreased afferent arteriole diameter by 2.9 +/- 0.4 microm. After perfusing heptanol into the lumen of the afferent arteriole, the TGF response was completely eliminated. When heptanol was added to the bath, it had no significant effect on TGF response. DISCUSSION: The data show that after mesangial cells were selectively damaged, the constriction of the afferent arteriole induced by increasing the NaCl concentration at the macula densa was eliminated. However, such treatment had no effect when Thy 1-1 was perfused into the macula densa lumen, and did not alter the response of the afferent arteriole to norepinephrine or acetylcholine. Disruption of the gap junctions also eliminated the TGF response. These data indicate that the mesangial cells play a key role in mediating the TGF response, and that gap junctions among mesangial cells and between mesangial cells and vascular smooth muscle cells communicate the TGF signal to the afferent arteriole. PMID- 12110014 TI - Reduced nitric oxide synthase activity in rats with chronic renal disease due to glomerulonephritis. AB - BACKGROUND: Animal studies with systemic nitric oxide synthase (NOS) inhibition and renal ablation, suggest that NO deficiency is both a cause and a consequence of chronic renal disease (CRD). METHODS: This study examined a glomerulonephritis (GN) model of CRD to determine if NO is deficient. In addition to measuring indices of renal function (proteinuria, creatinine clearance, structural damage), indices of total and renal nitric oxide production also were assessed (total NO(X) excretion, renal NOS activity, renal NOS protein abundance, plasma levels of NOS substrate and endogenous inhibitor). RESULTS: Rats developed increasing proteinuria 12 to 20 weeks after induction of GN (with anti-glomerular basement membrane, GBM, antibody) and at 20 weeks exhibited reduced creatinine clearances and increased glomerulosclerosis relative to age-matched controls. Total NO(X) excretion was reduced and the renal cortical NOS activity and neuronal NOS (nNOS) abundance was decreased relative to controls. There was no impact on renal or aortic endothelial NOS expression or cerebellar nNOS. The plasma l-arginine (Arg) concentration was well maintained but plasma asymmetric dimethylarginine (ADMA) concentration increased in GN versus control animals. CONCLUSIONS: Total and renal NOS activity is reduced in the GN model of CRD due to increased circulating endogenous NOS inhibitors and decreased renal nNOS abundance. PMID- 12110015 TI - Regulation of endothelin synthesis by extracellular matrix in human endothelial cells. AB - BACKGROUND: Vascular diseases are characterized by the presence of structural changes and the progressive loss of endothelial function. Although the biochemical basis of these structural changes have started to be outlined, it seems that accumulation of normal extracellular matrix proteins as well as the appearance of interstitial collagens, mainly collagen type I, characterize this process. On the other hand, a role for endothelial vasoactive factors has been proposed in the genesis of endothelial dysfunction, and it is generally accepted that changes in extracellular matrix composition may modify cell behavior. METHODS: Experiments were designed to test the influence of the supporting matrix on endothelin-1 (ET-1) synthesis by endothelial cells. Northern blot experiments were performed to analyze the prepro-endothelin-1 (prepro-ET-1) mRNA expression. ET-1 production was measured by ELISA. RESULTS: Cells grown on collagen type I (Col I) showed an increase of prepro-ET-1 mRNA level when compared with cells cultured on collagen type IV (Col IV). According to these results, the release of ET-1 to culture medium was also higher in Col I-grown cells than in those cultured on Col IV. Treatment of cells with a peptide that interferes with Col I integrins (D6Y), or with protein tyrosine kinase inhibitors such as genistein and herbimycin, completely abolished the effect of Col I. Moreover, experiments with antibodies against integrins suggest that these cell surface receptors could be involved in the modulation of ET-1 system by extracellular matrix. CONCLUSIONS: These results suggest that the presence of an abnormal extracellular matrix could stimulate endothelin synthesis by human endothelial cells, through integrin activation. PMID- 12110016 TI - Mechanism involved in bradykinin-induced efferent arteriole dilation. AB - BACKGROUND: There is evidence that kinins play a role in the regulation of renal hemodynamics. The balance of vascular resistance in afferent and efferent arterioles (Af-Art and Ef-Art) is a crucial factor in controlling glomerular filtration. We have previously reported that bradykinin has a biphasic effect on the Af-Art and that dilation and constriction are due to cyclooxygenase products, not nitric oxide (NO). The present study was designed to examine (1) the direct effect of bradykinin on the Ef-Art and (2) the mechanisms that mediate bradykinin induced Ef-Art dilation. METHODS: Isolated Ef-Arts were microperfused retrograde while maintaining the Ef-Art pressure at 30 mm Hg. Isolated Ef-Arts were preconstricted with norepinephrine. RESULTS: Perfusing the Ef-Art lumen with bradykinin caused dose-dependent vasodilation, increasing diameter from 6.9 +/- 0.7 to 8.0 +/- 0.8 (0.01 nmol/L), 8.3 +/- 0.7* (0.1 nmol/L), 10.3 +/- 0.7* (1 nmol/L) and 11.5 +/- 0.8* microm (10 nmol/L; N = 8; *P < 0.05 vs. NE). Neither L NAME nor indomethacin blocked the vasodilator effect of bradykinin; the diameter increased from 8.1 +/- 0.9 to 12.9 +/- 0.6 microm (10 nmol/L; P < 0.05 vs. control; N = 6) in the L-NAME-treated group and from 7.4 +/- 0.9 to 11.0 +/- 1.0 microm (10 nmol/L; P < 0.05 vs. control; N = 6) in the indomethacin-treated group. However, 25 micromol/L 17-ODYA, a cytochrome cP450 inhibitor, blocked the vasodilator effect of 10-8 mol/L bradykinin, leaving diameter unchanged (from 7.9 +/- 0.8 to 7.7 +/- 0.7 microm; N = 6). Finally, 0.1 micromol/L icatibant, a B2 receptor antagonist, completely blocked the vasodilation induced by bradykinin, and the diameter went from 7.8 +/- 0.7 to 8.3 +/- 0.8 microm (10 nmol/L). CONCLUSIONS: Bradykinin dilates Ef-Arts, but in contrast to Af-Arts its effect is not biphasic. The vasodilator effect of bradykinin in Ef-Arts via B2 receptors is mediated by cP450 metabolites (probably EETs), but not by NO or cyclooxygenase products. PMID- 12110017 TI - Statin therapy improves brachial artery endothelial function in nephrotic syndrome. AB - BACKGROUND: Patients with nephrotic syndrome have impaired endothelial function probably related to dyslipidemia. This study evaluated the effects of statin therapy on dyslipidemia and endothelial function in patients with nephrotic syndrome. METHODS: A sequential, open-label study of the effects of statins on endothelial dysfunction in 10 nephrotic patients treated with an angiotensin converting enzyme (ACE) inhibitor or angiotensin II (Ang II) receptor antagonist. Endothelial function was assessed at baseline, after 12 weeks of treatment with statins, and after an 8-week washout. Brachial artery endothelial function was measured as post-ischemic flow-mediated dilation (FMD) using ultrasonography. Endothelium-independent, glyceryl trinitrate-mediated vasodilation (GTNMD) also was measured. RESULTS: Serum lipids were significantly lower following statin: total cholesterol mean 8.2 +/- 0.4 (standard error) mmol/L versus 5.2 +/- 0.3 mmol/L, triglycerides 2.6 +/- 0.4 mmol/L versus 1.6 +/- 0.2 mmol/L, non-HDL cholesterol 6.7 +/- 0.4 mmol/L versus 3.7 +/- 0.2 mmol/L (all P < 0.001). There was a trend to an increase in serum albumin (31.0 +/- 1.3 g/L vs. 33.8 +/- 1.5 g/L; P = 0.078) and FMD improved significantly following treatment (3.7 +/- 1.1% vs. 7.0 +/- 0.8%, P < 0.01). After washout, FMD deteriorated significantly to 3.5 +/- 1.4% (P < 0.05) versus week 12 FMD. GTNMD was unchanged. In multivariate regression, reduction in non-high-density lipoprotein (HDL)-cholesterol (beta - 0.736, P = 0.027) and increase in serum albumin (beta 0.723, P = 0.028), but not the on-treatment level of non-HDL-cholesterol, were significant independent predictors of improvement in FMD after adjusting for change in resting brachial artery diameter. Changes in serum lipoprotein and albumin concentrations off treatment were not associated with deterioration in FMD. CONCLUSION: Statin therapy significantly improves dyslipidemia and brachial artery endothelial function in patients with nephrotic syndrome. Improvement in brachial artery endothelial function may be in part related to a non-lipid effect of statins. The findings also suggest a role for dyslipidemia in endothelial dysfunction and the risk for cardiovascular disease in nephrotic syndrome. PMID- 12110018 TI - Transient receptor potential channels in rat renal microcirculation: actions of angiotensin II. AB - BACKGROUND: This study assessed the calcium-activating mechanisms mediating glomerular arteriolar constriction by angiotensin II (Ang II). METHODS: Immunohistochemical and physiological studies were carried out, using antibody against transient receptor potential (TRP)-1 and an isolated perfused kidney model. RESULTS: Immunohistochemical experiments demonstrated that TRP-1 proteins were transcribed on both afferent and efferent arteriolar myocytes. In the first series of physiological experiments, Ang II (0.3 nmol/L) considerably constricted afferent (20.2 +/- 0.9 to 14.9 +/- 0.7 microm) and efferent arterioles (18.4 +/- 0.7 to 14.0 +/- 0.7 microm). The addition of nifedipine (1 micromol/L) restored decrements in afferent (to 20.0 +/- 0.8 microm) but not efferent arteriolar diameters. Further administration of SKF-96365 (100 micromol/L), a TRP channel blocker, reversed efferent arteriolar constriction (to 16.2 +/- 0.8 micromol/L). In the second group, although 2-aminoethoxydiphenyl borate (100 micromol/L), an inhibitor of inositol trisphosphate-induced calcium release (IP3CR), did not alter glomerular arteriolar diameters, it prevented Ang II-induced afferent arteriolar constriction and attenuated efferent arteriolar constriction (18.8 +/- 0.8 to 16.9 +/- microm). Subsequent removal of extracellular calcium abolished residual efferent arteriolar constriction (to 19.1 +/- 0.8 microm). CONCLUSIONS: Our data provide evidence that Ang II elicits IP3CR, possibly inducing a cellular response that activates voltage-dependent calcium channels on afferent arterioles. The present results suggest that Ang II-induced efferent arteriolar constriction involves IP3CR and calcium influx sensitive to SKF-96365. PMID- 12110019 TI - Renal dysfunction in allogeneic hematopoietic cell transplantation. AB - BACKGROUND: Allogeneic hematopoietic cell transplantation (HCT), formerly called bone marrow transplantation, can potentially cure various malignant and non malignant diseases, but it is associated with a high risk of toxicity. We have previously shown an overall 21% incidence of severe acute renal failure in patients undergoing autologous HCT. The present study evaluated renal dysfunction in patients undergoing allogeneic HCT. METHODS: The clinical course of 88 adult patients who received allogeneic HCT at the University of Colorado Health Science Center was analyzed. Renal dysfunction was classified as follows: Grade 0 = normal renal function; Grade 1 =>25% decrement in GFR but twofold increase in serum creatinine; Grade 3 =>twofold increase in serum creatinine and need for dialysis. RESULTS: Of the 88 patients, 81 (92%) patients had some degree of renal dysfunction (Grade 1, 20 patients; Grade 2, 32 patients; Grade 3, 29 patients). Severe nephrotoxicity (Grade 2 and Grade 3 renal dysfunction) was associated with significantly higher frequencies of sepsis, hepatic toxicity and hepatic veno-occlusive disease (VOD), and lung toxicity. The overall mortality rate at the end of 6 months was 58%. Grade 3 renal dysfunction was associated with a significantly increased risk of mortality (82.6%). CONCLUSION: A 92% incidence of renal dysfunction in allogeneic HCT patients was found. Lung and liver toxicities were significantly correlated with developing renal dysfunction, and the mortality rates for patients with Grade 3 renal failure exceeded 80%. PMID- 12110020 TI - Functional effects of renal artery stent placement on treated and contralateral kidneys. AB - BACKGROUND: This study examined the effects of stent placement for renal artery stenosis on the function of treated and contralateral kidneys. METHODS: Eighteen patients who underwent stent placement for unilateral renal artery stenosis presenting with hypertension and/or renal failure were studied before angiography and stent placement and at their one-year follow-up. Renal vein blood samples were taken at both sides, at each side simultaneously with a sample from the aorta, to measure the plasma renin concentration and the concentrations of 131I hippuran and 125I-thalamate during constant systemic infusion of these radiochemicals. This allowed an assessment of the single-kidney contributions to the total renin secretion, effective renal plasma flow (131I-hippuran clearance) and glomerular filtration rate (125I-thalamate clearance). RESULTS: At the one year follow-up, the vein-to-artery renin ratio at the treated side had decreased to normal, from 1.65 +/- 0.131 to 1.23 +/- 0.076 (mean +/- SEM; P = 0.011), indicating an improved renal blood flow. Contralaterally it rose from 1.09 +/- 0.042 to 1.17 +/- 0.029 (P = 0.055) at follow-up. The extraction ratio of 131I hippuran improved at the treated side (0.48 +/- 0.049 to 0.62 +/- 0.034; P = 0.003) and contralaterally (0.67 +/- 0.033 to 0.73 +/- 0.026; P = 0.043). The extraction ratio of 125I-thalamate, which equals filtration fraction, improved at both sides (0.12 +/- 0.014 to 0.17 +/- 0.012 at the treated side, P = 0.001; 0.18 +/- 0.013 to 0.22 +/- 0.011 contralaterally, P = 0.002). Two-kidney effective renal blood flow and glomerular filtration rate remained unchanged. CONCLUSION: Renal artery stenting was capable of causing improvement of glomerular filtration rate of the treated kidney, although the overall glomerular filtration rate did not change. PMID- 12110021 TI - Electrocardiogram with prolonged QT interval in Gitelman disease. AB - BACKGROUND: Potassium and magnesium deficiency prolong the QT interval on a standard electrocardiogram and predispose the patient to dangerous cardiac arrhythmias. No information is available on QT interval in patients diagnosed with Gitelman disease. METHODS: The QT interval was assessed on lead II in 27 patients with biochemically and genetically defined Gitelman disease, who had discontinued medical treatment for at least four weeks. They included 15 female and 12 male subjects, aged 6.7 to 40 years old, median 20 years old. The corrected QT interval was calculated from the measured QT interval and heart rate using the Bazett formula. RESULTS: The corrected QT interval was normal (between 391 and 433 msec) in 16 and prolonged in the remaining 11 patients (between 444 and 504 msec). Patients with prolonged and patients with normal QT interval did not significantly differ with respect to female to male ratio, plasma potassium, plasma total magnesium, and plasma ionized calcium. Plasma sodium and chloride values were slightly but significantly lower and bicarbonate levels higher in patients with a prolonged than in those with a normal QT interval. CONCLUSIONS: The corrected QT interval is often pathologically prolonged in patients with Gitelman disease, suggesting that there is an increased risk for development of dangerous arrhythmias. Further investigations are required in patients with a prolonged QT interval to assess the true hazard of dangerous arrhythmias. PMID- 12110022 TI - Modest serum creatinine elevation affects adverse outcome after general surgery. AB - BACKGROUND: Modest preoperative serum creatinine elevation (1.5 to 3.0 mg/dL) has been recently shown to be independently associated with morbidity and mortality after cardiac surgery. It is important to know if this association can be applied more broadly to general surgery cases. METHODS: Multivariable logistic regression analyses of 46 risk variables in 49,081 cases from the Veterans Affairs National Surgical Quality Improvement Program, undergoing major general surgery from 10/1/96 through 9/30/98. RESULTS: Thirty day mortality and several cardiac, respiratory, infectious and hemorrhagic morbidities were significantly (P < 0.001) higher in patients with a serum creatinine>1.5 mg/dL. With multivariable analysis, the adjusted odds ratio for mortality for patients with a serum creatinine of 1.5 to 3.0 mg/dL was 1.44 [95% confidence interval (95% CI) 1.22 to 1.71] and for creatinine>3.0 mg/dL was 1.93 (95% CI 1.51 to 2.46). The adjusted odds ratio for morbidity (one or more postoperative complications) for patients with a serum creatinine of 1.5 to 3.0 mg/dL was 1.18 (95% CI 1.06 to 1.32) and for creatinine>3.0 mg/dL was 1.19 (95% CI 0.99 to 1.43). Further stratification and recursive partitioning of creatinine levels revealed that a serum creatinine level>1.5 mg/dL was the approximate threshold for both increased morbidity and mortality. CONCLUSIONS: Modest preoperative serum creatinine elevation (>1.5 mg/dL) is a significant predictor of risk-adjusted morbidity and mortality after general surgery. A preoperative serum creatinine of 1.5 mg/dL or higher is a readily available marker for potential adverse outcomes after general surgery. PMID- 12110023 TI - Malnutrition in hemodialysis diabetic patients: evaluation and prognostic influence. AB - BACKGROUND: This work aimed to evaluate the role of malnutrition in the increased mortality rate of hemodialysis diabetic patients from a French cooperative series. METHODS: Body mass index (BMI), serum albumin, prealbumin, cholesterol, and pre-dialysis creatinine, normalized protein catabolic rate and lean body mass (LBM) were measured in 734 diabetic and 6389 non-diabetic patients (aged 63.4 +/- 12.2 and 62.0 +/- 15.9 years; 1.01 male to 1.40 female ratio). The outcome of 1610 of these patients, including 170 diabetics, was assessed during a 30-month follow-up. RESULTS: Diabetic as compared to non-diabetic patients showed a significant (P < 10-4) increased BMI (25.9 +/- 5.2 vs. 23.1 +/- 4.3) and cholesterol (5.5 +/- 1.6 vs. 5.3 +/- 1.5 mmol/L), and decreased albumin (37.8 +/- 5.4 vs. 38.9 +/- 5.3 g/L), prealbumin (317 +/- 91 vs. 340 +/- 94 mg/L), creatinine (711 +/- 184 vs. 816 +/- 217 micromol/L) and LBM (76 +/- 18 vs. 87 +/- 21%). Normalized protein catabolic rate was similar in the two groups (1.11 +/- 0.31 vs. 1.13 +/- 0.32 g/kg/L). One and two-year survival was 83.7 +/- 2.9% and 65.5 +/- 3.8% in diabetic patients versus 90.3 +/- 0.8% and 79.9 +/- 1.1% in non diabetics (relative risk 1.26, P < 0.01). Independent predictors of survival were age, albumin and prealbumin in non-diabetics and only age in diabetics. CONCLUSION: Diabetic patients compared to non-diabetics were characterized by an increased incidence of protein malnutrition and decreased survival. However, the higher death risk associated with diabetes was not related to malnutrition. PMID- 12110024 TI - Vitamin E-modified filters modulate Jun N-terminal kinase activation in peripheral blood mononuclear cells. AB - BACKGROUND: The generation during hemodialysis of activated complement fragments and reactive oxygen species, including nitric oxide (NO), may affect peripheral blood mononuclear cell (PBMC) function. Currently, little is known about signal transduction pathways involved in PBMC activation. Jun N-terminal kinase (JNK) is a novel mitogen-activated protein (MAP) kinase phosphorylated and activated in response to oxidative stress and directly involved in cell activation. METHODS: The present study evaluated the activation of JNK in PBMCs isolated from eight uremic patients undergoing, in a randomized manner, three month-subsequent periods of hemodialysis with a low-flux cellulose acetate (CA) and a vitamin E modified cellulose membrane (CL-E). After each period of treatment, PBMCs were harvested before (T0), during (T15) and after three hours (T180) of dialysis. At the indicated time points, plasma C5b-9 generation by ELISA and inducible NO synthase (iNOS) gene expression by in situ hybridization were evaluated also. The activation of JNK was studied by Western blotting using a specific monoclonal anti-phospho-JNK antibody, which recognizes the activated form of JNK. RESULTS: At T0, a significant increase in plasma C5b-9 levels was found in CA patients compared to CL-E-treated patients. During hemodialysis, C5b-9 levels rose more significantly in CA patients than in CL-E patients and returned to baseline values only in CL-E patients. At the same time, in CA patients an increased iNOS gene expression was observed at T180 together with a striking activation of JNK. By contrast, PBMC from CL-E-treated patients showed undetectable levels of phospho-JNK and a significant reduction in iNOS expression. Interestingly, incubation of PBMCs with normal human plasma (10%), activated by contact with a cellulosic membrane, induced a time-dependent increase in JNK phosphorylation that was completely inhibited by blocking complement cascade activation. CONCLUSION: Our data suggest that JNK phosphorylation is strikingly increased in PBMCs obtained from CA-treated patients and may represent a key cellular event in PBMC activation during dialysis with bioincompatible membranes. The activation of this signaling enzyme, mediated by active complement fragments and PBMC-dialyzer interaction, can be significantly reduced by the use of vitamin E-coated membrane. PMID- 12110025 TI - Bile acid binding to sevelamer HCl. AB - BACKGROUND: Clinical studies have shown sevelamer HCl (Renagel) to be effective for the reduction of serum phosphate in hemodialysis patients. These studies also consistently have demonstrated a significant reduction of low-density lipoprotein (LDL) cholesterol following treatment with sevelamer. METHODS: Equilibrium binding of bile acids and oleic acid was determined by incubating sevelamer with ligand containing buffer. Aliquots of the solution were filtered and the free ligand concentrations quantitated by high-pressure liquid chromatography (HPLC). Flow kinetics were determined using a cylindrical flow cell containing trapped sevelamer. Bile acid and oleic acid were pumped through the stirred cell in a manner designed to mimic the in vivo situation. Binding was monitored by HPLC. RESULTS: Sevelamer binds bile acids cooperatively and with high capacity. At low binding densities, the presence of the more hydrophobic bile acids enhances the binding of the less hydrophobic bile acids, and the presence of oleic acid enhances the binding of all bile acids. At saturating oleic acid concentrations, the bile acid binding capacity of sevelamer is reduced by only a factor of two. Moreover, the presence of oleic acid dramatically diminishes the release rate of bile acids from sevelamer. CONCLUSIONS: The favorable bile acid binding characteristics of sevelamer provide a compelling explanation for its ability to lower LDL cholesterol in hemodialysis patients and in healthy volunteers. PMID- 12110026 TI - Vascular access and increased risk of death among hemodialysis patients. AB - BACKGROUND: Hemodialysis with a venous catheter increases the risk of infection. The extent to which venous catheters are associated with an increased risk of death among hemodialysis patients has not been extensively studied. METHODS: We conducted a retrospective cohort study of 7497 prevalent hemodialysis patients to assess the association between dialysis with a venous catheter and risk of death due to all causes and to infection. RESULTS: A tunneled cuffed catheter was used for access in 12% of the patients and non-cuffed, not tunneled catheter in 2%. Younger age (P = 0.0005), black race (P = 0.0022), female gender (P = 0.0004), short duration since starting dialysis (P = 0.0003) and impaired functional status (P = 0.0001) were independently associated with increased use of catheter access. The proportion of patients who died was higher among those who were dialyzed with a non-cuffed (16.8%) or cuffed (15.2%) catheter compared to those dialyzed with either a graft (9.1%) or a fistula (7.3%; P < 0.001). The proportion of deaths due to infection was higher among patients dialyzed with a catheter (3.4%) compared to those dialyzed with either a graft (1.2%) or a fistula (0.8%; P < 0.001). The adjusted odds ratio (95% CI) for all-cause and infection-related death among patients dialyzed with a catheter was 1.4 (1.1, 1.9) and 3.0 (1.4, 6.6), respectively, compared to those with an arteriovenous (AV) fistula. CONCLUSION: Venous catheters are associated with an increased risk of all-cause and infection-related mortality among hemodialysis patients. PMID- 12110028 TI - Intercurrent clinical events are predictive of plasma C-reactive protein levels in hemodialysis patients. AB - BACKGROUND: In chronic hemodialysis (HD) patients, the repetitive induction of the acute phase response (APR) may induce a chronic micro-inflammatory state, leading to various long-term complications. METHODS: The present prospective study was designed to assess the alterations in the APR in 74 patients who were randomized to HD with a high-flux polysulfone (PS; F 60S), a super-flux PS (F 500S), or a super-flux cellulosic tri-acetate (CTA and CTA with filtered dialysate, CTA(f)) dialyzer. Blood samples collected at the start of the study and after twelve weeks were analyzed for interleukin-6 (IL-6) and C-reactive protein (CRP). In addition to the microbiological quality of the dialysate, the appearance of a "clinical event" was assessed. RESULTS: At baseline, mean IL-6 levels were within the reference range whereas mean CRP levels were slightly elevated. Mean values did not change after 12 weeks of HD with either modality. After subdividing the patients in quartiles with increasing change in plasma CRP, 23.0% of the patients showed a change of more than 8.0 mg/L. In a multiple regression analysis, CRP levels appeared to be independent of the degree of dialysate contamination, the material and the flux characteristics of the devices. In fact, the variable "clinical events" was the only significant predictor of the plasma CRP levels (P < 0.001). CONCLUSIONS: Based on these results, both PS and CTA super-flux dialyzers appear safe for clinical use. Whether changes in CRP values, which are associated with intercurrent clinical events, influence the long-term prognosis of chronic HD patients remains to be established. PMID- 12110027 TI - Regulatory functions of alloreactive Th2 clones in human renal transplant recipients. AB - BACKGROUND: Chronic allograft rejection is the major clinical problem in organ transplantation. There is evidence that indirect T cell recognition of donor specific HLA peptides may play an important role in the immunopathogenesis of chronic allograft rejection. We have recently shown that HLA allopeptide-specific T cell clones generated from renal transplant recipients with chronic allograft nephropathy are of the Th1 phenotype, while those from stable patients are Th2. There is evidence in experimental animal models of autoimmunity and transplantation that Th2 cells may function to regulate immune responses, but the biological relevance of these observations in humans has not been reported. METHODS: The purpose of this study was to investigate the putative regulatory functions of alloreactive human Th2 clones. HLA-DR allopeptide-specific Th1 and Th2 cell clones were generated from peripheral blood lymphocytes of human renal allograft recipients with chronic allograft nephropathy (CAN) or with stable renal function (SRF), respectively. RESULTS: An in vitro co-culture system showed that the proliferative responses of Th1 clones from patients with CAN were significantly inhibited by the Th2 clones in response to the donor-derived HLA allopeptides. In addition, co-culture of the Th2 clones inhibited cytokine production (IFN-gamma) by the Th1 clones in response to the donor-specific peptides. The regulatory functions of Th2 clones were antigen-specific since they only occurred when both the Th1 and Th2 clones were reactive to the same HLA-DR allopeptide, and were mediated by IL-4 and IL-10. CONCLUSIONS: This is the first demonstration, to our knowledge, indicating that Th2 cells may function to regulate indirect Th1 alloimmune responses that are critical for the progression of CAN in humans. PMID- 12110029 TI - A novel association between residual renal function and left ventricular hypertrophy in peritoneal dialysis patients. AB - BACKGROUND: Left ventricular hypertrophy (LVH) and dialysis adequacy are both important predictors for mortality in dialysis patients. This study evaluated the association between residual renal function (RRF) and the severity of LVH in endstage renal failure (ESRF) patients undergoing long-term continuous ambulatory peritoneal dialysis (CAPD). METHODS: A cross-section study was performed with left ventricular mass index (LVMi), determined in 158 non-diabetic CAPD patients using echocardiography and its relationship with residual glomerular filtration rate (GFR), peritoneal dialysis (PD) and total weekly urea clearance (Kt/V) and other known risk factors for LVH was evaluated. RESULTS: Twelve patients had no LVH (group I). The remaining 146 patients were stratified [group II (lowest), III and IV (highest)] according to the LVMi (median 207 g/m2; range 103 to 512 g/m2). Across the four groups of patients with increasing LVMi, there was significant decline in GFR (2.27 +/- 1.98 vs. 1.49 +/- 1.58 vs. 1.61 +/- 1.91 vs. 0.80 +/- 1.42 mL/min/1.73 m2; P = 0.011) and total weekly Kt/V (1.98 +/- 0.44 vs. 1.96 +/- 0.38 vs. 1.92 +/- 0.42 vs. 1.71 +/- 0.42; P = 0.037); however, PD Kt/V was similar for all four groups. Patients with better-preserved residual GFR not only had significantly higher total Kt/V, but were less anemic and hypoalbuminemic and had a trend toward lower systolic blood pressure and arterial pulse pressure. Multiple regression analysis showed that other than age, gender, body weight, arterial pulse pressure, hemoglobin and serum albumin, known factors for LVH, residual GFR (estimated mean -7.94; 95% confidence interval -15.13 to -0.74; P = 0.031) was also independently associated with LVMi. CONCLUSIONS: Other than anemia, hypoalbuminemia and arterial pulse pressure, this study demonstrates an important, novel association between the degree of RRF and severity of LVH in ESRF patients undergoing long-term CAPD. Prospective studies are needed to define if indeed there is a cause-effect relationship between this association, to evaluate if a decline in residual GFR is independently associated with an increase in LVMi, and to determine whether treatment directed at preserving RRF will reduce the severity of LVH, improve cardiac performance and hence survival of these patients. PMID- 12110030 TI - Cardiovascular disease in pediatric chronic dialysis patients. AB - BACKGROUND: Little information is available regarding cardiac morbidity and mortality in children with end-stage renal disease. We sought to determine the incidence of cardiac morbidity and mortality in pediatric chronic dialysis patients. METHODS: Medicare incident pediatric (0 to 19 years) dialysis patients from 1991 to 1996 were identified from the United States Renal Data System. Study endpoints included development of arrhythmia, valvular heart disease, cardiomyopathy, or cardiac arrest, all causes of death, and cardiac-related death. Statistical analyses were performed using the Poisson regression model and chi-square test. RESULTS: A total of 1454 children were eligible for inclusion, 452 (31.1%) of whom developed a cardiac-related event. Arrhythmia was the most common event (19.6%) compared with valvular disease (11.7%), cardiomyopathy (9.6%), and cardiac arrest (3%). Arrhythmia and valvular heart disease incidence were increased in 15- to 19-year-olds (P < 0.0001 for both), females (P = 0.004, P = 0.03) and blacks (P < 0.0001, P = 0.002). Cardiomyopathy incidence was increased in blacks (P = 0.001) and tended to be increased in females (P = 0.053). The adjusted annual cardiomyopathy rate during the first 3 years increased between 1991 and 1996 (P = 0.003). Death occurred in 107 patients, and 41 (38%) were cardiac deaths. CONCLUSIONS: Cardiovascular disease is a significant cause of morbidity and mortality in pediatric chronic dialysis patients. Cardiomyopathy incidence is increasing. Black, female, and adolescent children have increased risk for cardiovascular disease. PMID- 12110031 TI - Polyethylene glycol reduces the inflammatory injury due to cold ischemia/reperfusion in autotransplanted pig kidneys. AB - BACKGROUND: The conditions of storage of donor kidney may influence the deleterious consequences of ischemia/reperfusion injury (IRI) on delayed graft function. Since polyethylene glycol (PEG) can protect renal tubule cells against cold injury, we tested the effects of adding PEG 20 kD to ice-cold preservation solutions on the IRI of autotransplanted pig kidneys. METHODS: The pigs' left kidneys were removed, cold-flushed with University of Wisconsin (UW) or simplified high K+ or high Na+ solutions with or without 30 g/L PEG 20M and stored for 48 hours at 4 degrees C. The kidneys were then autotransplanted and the contralateral kidneys were removed. Kidney biopsies were then performed and renal function parameters were analyzed over 8 to 12 weeks following surgery. RESULTS: The kidneys cold-flushed with PEG-supplemented solutions on day 7 post transplantation were better preserved and exhibited less marked nuclear tubular cell damage than the kidneys cold-flushed with the UW solution alone. PEG also almost completely inhibited the overexpression of major histocompatibility complex class II that was detected in epithelial tubule cells from kidneys cold flushed with the UW solution. PEG also significantly reduced the number of CD4+ T lymphocytes and limited the infiltration of macrophages/monocytes and the progression of interstitial fibrosis in the 8- to 12-week post-transplanted kidneys. Moreover, pigs autotransplanted with kidneys flushed with PEG supplemented solutions had the best renal function and the lowest levels of proteinuria. CONCLUSIONS: These findings indicate that PEG inhibits the early inflammatory response due to IRI, improves renal function, and may prevent the progression of interstitial fibrosis in the long-term autotransplanted pig kidney. PMID- 12110032 TI - Experimental diabetes induces functional and structural changes in the peritoneum. AB - BACKGROUND: Peritoneal dialysis (PD) is an established renal replacement therapy in diabetic patients, but the influence of diabetes on the peritoneal membrane (PM) remains debated. We have used functional, biochemical and molecular studies in vivo and in vitro to substantiate the changes induced by diabetes and hyperglycemia in the PM. METHODS: Peritoneal equilibration tests were performed 2, 4, and 6 weeks after induction of diabetes with streptozotocin (STZ) in rats. Morphological analyses, determination of nitric oxide synthase (NOS) activities, and expression studies for NOS isoforms and advanced glycation end products (AGE) were performed in parallel. Additional studies were conducted in diabetic rats treated with insulin, non-diabetic rats fed with urea, and cultured bovine aortic endothelial cells (BAEC). RESULTS: In comparison with controls, diabetic rats were characterized by: increased permeability for small solutes and decreased sodium sieving; capillary proliferation; increased endothelial NOS (eNOS) and AGE immunoreactivity; up-regulation of eNOS and down-regulation of neuronal NOS; and increased NOS activity in the PM. The changes, which culminated at week 6, were prevented by chronic insulin treatment in diabetic rats. In contrast to hyperglycemia, hyperosmolality alone did not induce functional or structural changes in the PM. Studies in BAEC showed that high glucose incubation led to increased activity and expression of eNOS, a prerequisite for vascular proliferation. CONCLUSIONS: These data demonstrate that chronic hyperglycemia is associated with functional and structural changes in the peritoneum that parallel with selective regulation of NOS isoforms and AGE deposits. The alterations are prevented by insulin treatment, which suggests that adequate control of diabetes can preserve PM integrity in diabetic patients prior to PD. PMID- 12110033 TI - Efficient in vitro lowering of carbonyl stress by the glyoxalase system in conventional glucose peritoneal dialysis fluid. AB - BACKGROUND: Reactive carbonyl compounds (RCOs) present in heat-sterilized peritoneal dialysis (PD) fluid have been incriminated in the progressive deterioration of the peritoneal membrane observed in long-term PD patients. The present study utilized the glyoxalase I (GLO I) system as a new approach to lower in vitro the peritoneal fluid content of RCOs such as methylglyoxal (MGO), glyoxal (GO) and 3-deoxyglucosone (3-DG). METHODS: GO, MGO, and 3-DG solutions or conventional glucose PD fluids were incubated in vitro with various RCO lowering compounds. The evolution of GO, MGO, and 3-DG levels was monitored by high performance liquid chromatography. The tested compounds included aminoguanidine and glutathione (GSH), alone or together with GLO I. The human GLO I gene was overexpressed in Chinese hamster ovary (CHO) cells, or ubiquitously in transgenic mice. Cell supernatant of the CHO transfectant and protein extracts of various organs of the transgenic mice were also tested. RESULTS: Aminoguanidine incubated with MGO/GO/3-DG mixtures, promptly reduced RCO levels. GSH alone had a similar but milder and slower effect. Together with GLO I, it promptly decreased GO and MGO levels but was less efficient toward 3-DG. After incubation with glucose PD fluid, GSH together with GLO I had the same effect on MGO, GO, and 3-DG levels. Addition of transfected cell supernatant or tissue extracts overexpressing GLO I, together with GSH to either GO, MGO, or 3-DG solutions, promptly and markedly reduced GO and MGO but not 3-DG levels. CONCLUSIONS: GLO I together with GSH efficiently lowers glucose-derived RCOs, especially GO and MGO, both in conventional glucose PD fluids and in RCO solutions. The fact that genetically manipulated cells overexpressing GLO I activity have a similar effect suggests that maneuvers raising GLO I activity in peritoneal cells or in the peritoneal cavity might help prevent the deleterious effects of the peritoneal carbonyl stress in PD patients. The clinical relevance of this approach is yet to be documented. PMID- 12110034 TI - Recombinant human growth hormone post-renal transplantation in children: a randomized controlled study of the NAPRTCS. AB - BACKGROUND: Growth retardation persists in renal allograft recipients despite successful transplantation. The etiology is multi-factorial including the adverse effects of corticosteroids, suboptimal allograft function, and perturbations of the GH/GF axis. Recombinant human growth hormone (rhGH) has been effective in improving growth velocity; however, allograft dysfunction has been reported. Therefore, a randomized controlled study was undertaken. METHODS: Sixty-eight growth retarded pediatric renal allograft recipients were enrolled in a one-year randomized controlled study to determine the efficacy and safety of rhGH. A protocol biopsy was performed prior to enrollment. RESULTS: After one year, the delta SDS (standardized height) was +0.49 +/- 0.10 in the treatment group (N = 30) compared to -0.10 +/- 0.08 in the control group (N = 22; P < 0.001). During the first year, there were no rejection episodes in the treatment group and three in the control group. After the first year, when all recipients were receiving rhGH, there were three patients in the treatment group and two patients in the control group who experienced an acute rejection episode. Prior to enrollment, more than one acute rejection episode was predictive of a subsequent rejection following enrollment. There was no difference in adverse events between the two groups. CONCLUSION: In conclusion, rhGH is effective in improving the growth velocity of pediatric renal allograft recipients and is not associated with an increase in adverse events. PMID- 12110035 TI - 3,4-Dideoxyglucosone-3-ene (3,4-DGE): a cytotoxic glucose degradation product in fluids for peritoneal dialysis. AB - BACKGROUND: Bioincompatible glucose degradation products (GDPs) in fluids for peritoneal dialysis (PD) develop during sterilization and storage. Their biological activity has successfully been monitored through the use of various in vitro methods but their molecular and chemical nature is less well understood. Many GDPs are highly reactive carbonyl compounds. Although some of the identified GDPs are extremely cytotoxic, none of them actually possess cytotoxicity at the concentrations found in PD fluids. Thus, the GDP responsible for the toxicity in PD fluids has not yet been identified. The intention of the present work was to investigate to what extent the unsaturated dicarbonyl compound, 3,4 dideoxyglucosone-3-ene (3,4-DGE) was present in PD fluids, and if it could be responsible for the in vitro effects on L-929 fibroblast cells. METHODS: A commercial preparation of 3,4-DGE and two different liquid chromatography methods were used for the chemical identification and quantification. In vitro bioincompatibility was determined as inhibition of cell growth using the L-929 fibroblast cell line. RESULTS: 3,4-DGE was present in conventionally manufactured PD fluids at a concentration of 9 to 22 micromol/L. In the newly developed PD fluid, Gambrosol trio, the concentrations were 0.3 to 0.7 micromol/L. When added as synthetic 3,4-DGE to cell growth media at the concentrations measured in conventional PD fluids, the inhibition of cell growth was significantly lower than for that seen with the conventional fluids. However, in the conventional PD fluids the total amount of 3,4-DGE available for toxic reactions most probably was higher than that measured, because 3,4-DGE was freshly recruited from a molecular pool when consumed. The speed of this recruitment was high enough to explain most of the growth inhibition seen for heat-sterilized PD fluids. CONCLUSION: 3,4-DGE is present in conventional PD fluids at a concentration between 9 and 22 micromol/L, and is the most biologically active of all GDPs identified to date. Thus, it is the main candidate to be held responsible for the clinical bioincompatibility caused by conventionally manufactured PD fluids. PMID- 12110036 TI - Five preventable causes of kidney graft loss in the 1990s: a single-center analysis. AB - BACKGROUND: Despite improvements in immunosuppressive protocols and patient care, kidney allografts continue to fail. We studied causes of graft loss for primary kidney transplants in the 1990s to determine major causes and potential interventions. METHODS: Causes of graft loss were reviewed for 1467 primary kidney transplants done at our institution between January 1, 1990, and December 31, 1999. Graft loss for that entire population was studied and then the causes of loss selectively examined at <1 year, 1 to 5 years, and>5 years post transplant. Finally, causes of loss in the 1990s versus the 1980s were compared. RESULTS: Five major causes of graft loss were noted in the 1990s: thrombosis, acute rejection (either alone or combined with delayed graft function or infection), chronic rejection, death with function, and noncompliance. In the first year post-transplant, thrombosis (25%) and death with function (41%) were the major causes of graft loss. After the first year, chronic rejection and death with function predominated. For recipients dying with graft function, cardiovascular disease was the major cause of death. CONCLUSIONS: This study identified the five major causes of kidney graft loss in the 1990s. Different interventions are required to decrease loss from each of these causes. Future research needs to be directed at such interventions. PMID- 12110037 TI - The identification of IgA receptors in human mesangial cells: in the search for "Eldorado". PMID- 12110039 TI - Novel erythropoiesis-stimulating protein in the management of the anemia of chronic renal failure. PMID- 12110038 TI - Identifying and addressing potentially preventable causes of renal allograft loss. PMID- 12110040 TI - Revision of the 1992-edition of the WHO histological typing of odontogenic tumours. A suggestion. AB - Classification of odontogenic tumours is an academic exercise that has developed over the last 150 years. It was not until 1971 when a 5-year collaborated effort, organized by the World Health Organization (WHO), resulted in the first consensus on taxonomy of odontogenic tumours. The appearance of this first authoritative guide to the classification of odontogenic tumours marked the start of an era of quite intensive interest for studying this particular field of oral pathology. An updated 2nd edition of the WHO classification was published in 1992. PMID- 12110041 TI - Proliferation and differentiation markers in snuff-induced oral mucosal lesions. AB - BACKGROUND: Regular use of snuff is known to cause whitish oral mucosal lesions of variable severity at the usual quid placement site. The main aim of this study was to elucidate cellular mechanisms involved in snuff-induced epithelial changes. METHODS: Expression patterns for markers of cell proliferation (PCNA, Ki 67), cell cycle regulation (p53, p21), keratin changes (pankeratin, CK18, CK19), cell stress (HSP 70) and collagen type IV in 14 snuff-induced oral mucosal lesions and 12 control samples were analyzed by immunohistochemistry (IHC). RESULTS: On light microscopy, all snuff-induced lesions were characterized by a hyperkeratinized and thickened epithelium. Some vacuolized cells, markers of cell degeneration, were frequently seen (in 9/14 of the samples) in the superficial layers in epithelia. Expression of PCNA and Ki-67 was found in a statistically significantly fewer cells in snuff-induced lesions (P < 0.001) than in the controls. This indicates that epithelia in snuff-induced lesions are not thickened as a result of increased cellular proliferation, but by protracted turnover of differentiating cells. Of cell cycle markers, p21 was found be up regulated in 4/14 snuff-induced lesions, probably by p53-independent pathways. Only two snuff-induced lesions showed p53 positivity. However, the number of stained cells with p53 and p21 was not statistically different from that in controls. Expression of CK18, but not any alterations in CK19 expression, was seen in 5 of 14 snuff-induced lesions. Snuff also seems to stimulate the expression of collagen type IV, possibly by basal cells, as indicated by the thickened staining of the basal lamina. CONCLUSIONS: The findings of this study showing suppressed cellular proliferation and infrequent p53 dysfunction in snuff lesions may partly explain why dysplastic changes are seldom seen in mucosal lesions induced by the Scandinavian type of snuff. PMID- 12110042 TI - Burning mouth syndrome (BMS): double blind controlled study of alpha-lipoic acid (thioctic acid) therapy. AB - BACKGROUND: Burning mouth syndrome (BMS) has features of a neuropathy and could be related to the production of the toxic free radicals that are released in stress situations. Alpha-lipoic acid is an antioxidant able to increase the levels of intracellular glutathione and eliminate free radicals. This study aimed to examine the effectiveness of alpha-lipoic acid in the therapy of BMS. METHOD: This was a double blind, controlled study conducted for two months on 60 patients with constant BMS. Comparing alpha-lipoic acid (test) with cellulose starch (placebo), there was no laboratory evidence of deficiencies in iron, vitamins or thyroid function and no hyperglycaemia. RESULTS AND CONCLUSION: Following treatment with alpha-lipoic acid, there was a significant symptomatic improvement, compared with placebo, with the majority showing at least some improvement after 2 months, thus supporting the hypothesis that burning mouth syndrome is a neuropathy. This improvement was maintained in over 70% of patients at the 1 year follow-up. PMID- 12110043 TI - Infrequent genetic alterations of the tumor suppressor gene PTEN/MMAC1 in squamous cell carcinoma of the oral cavity. AB - BACKGROUND: Fifty tumor specimens from primarily untreated patients were analyzed to elucidate the involvement of the tumor suppressor gene PTEN/MMAC1 in the development of oral squamous cell cancer. METHODS: Eight microsatellite markers, spanning 10 cM of genomic DNA located centromeric, telomeric or intragenic of PTEN/MMAC1 were used for loss of heterozygosity (LOH) and breakpoint analysis. The microsatellite panel within or in close proximity (1 cM) to the 10q23.3 locus showed a LOH rate of 12%. Complete sequence analysis of the genes coding region was performed in all 10 cases that exhibited LOH in one of the eight microsatellite markers within or around the PTEN/MMAC1 gene. Comparative multiplex PCR reactions served to screen for homozygous deletions. RESULTS: There was no association between allelic loss of the gene, overall patient survival and recurrence-free survival. Sequencing did not reveal any mutation in the coding region of PTEN/MMAC1. Differential PCR analysis failed to detect any homozygous deletion. CONCLUSIONS: We conclude that PTEN/MMAC1 gene alterations do not play a key role in tumorigenesis of oral squamous cell cancers. PMID- 12110044 TI - Progressive bulbar palsy: a case report diagnosed by lingual symptoms. AB - The aim of this report is to show a case of Progressive Bulbar Palsy (PBP), diagnosed by oral medicine specialists, from oral symptoms of the disease. We have found no more than two published cases of PBP diagnosed by lingual alterations. We have followed the patient for almost four years, which is remarkable considering that the normal survival period for these patients is up to three years. We would like to emphasize the role of general dentists in the diagnosis of systemic conditions based on an oral examination that should include the oro-facial muscles. PMID- 12110045 TI - Lymphangioma involving the mandible: immunohistochemical expressions for the lymphatic proliferation. AB - We report a case of lymphangioma involving oral mucosa and mandible of an elderly female. The surgical and radiological examinations indicated that the lymphangioma was mainly distributed in the labial mucosa tissue, but had gradually extended into the periosteum and intrabony space of mandible. Immunohistochemical staining was also performed using antiseras of alpha-smooth muscle actin (alpha-SMA), von Willebrand factor (vWF), angiogenin, vascular endothelial growth factor (VEGF), and proliferating cell nuclear antigen (PCNA) to elucidate the pathogenetic implications of the intraosseous lymphangioma. The present case of lymphangioma showed strong immunohistochemical reactivity of angiogenin and vWF, while it showed weak reactions of VEGF and PCNA. The immunostaining of alpha-SMA disclosed an abnormally thinned and discontinuous smooth muscle layer in the lymphatics. Both the X-rays and histological examination showed that the lymphangioma lesion was gradually extending into the adjacent osteoporotic marrow space of mandible. Therefore, we believe that the present case of intraosseous lymphangioma, which showed the harmatomatous growth of the lymphatics into the marrow space of mandible, is closely related to osteoporotic changes of old age. PMID- 12110046 TI - The treatment of oral aphthous ulceration or erosive lichen planus with topical clobetasol propionate in three preparations. A clinical study on 54 patients. PMID- 12110049 TI - A rhesus monkey model of respiratory syncytial virus infection. AB - Respiratory syncytial virus (RSV) is a major cause of lower respiratory tract disease in infants and young children worldwide. To date, there is no single animal model that adequately reproduces all human disease states. Here, we have developed a model of experimental infection with human RSV in infant Rhesus macaques. Infected animals demonstrated mild clinical disease including increased respiratory rates, fever and adventitious lung sounds. While more severe disease was not observed, preliminary virological and histopathological findings are promising. It is anticipated that with further optimization, this model will provide a useful system with which to study disease due to RSV infection and evaluate candidate vaccines. PMID- 12110050 TI - Clinical and histopathological evaluation of 13 cases of adenocarcinoma in aged rhesus macaques (Macaca mulatta). AB - In recent years, the emphasis on aging research, has led to an increase in the number of aged macaques being maintained in some research facilities with a subsequent increase in the occurrence of age-related diseases. One of the most commonly reported age related diseases is intestinal adenocarcinoma. At the University of Illinois at Chicago (UIC), which maintains a colony of approximately 55 aged rhesus macaques 13 cases of intestinal adenocarcinoma were diagnosed within a 25-month period. This report provides a comprehensive description of the clinical findings for intestinal adenocarcinoma in aged rhesus macaques, including results from physical examinations, laboratory tests, radiographic evaluations, gross and histopathologic findings as well as a comparison with the disease condition in humans. The use of carcinoembryonic antigen as a potential tumor marker was evaluated by immunohistochemical analysis of tissue specimens in 10 cases. Intestinal adenocarcinoma is a disease condition that should be of concern to individuals responsible for the care of aged rhesus macaques. PMID- 12110052 TI - Exposure of cynomolgus monkey embryos to retinoic acid causes thymic defects: effects on peripheral lymphoid organ development. AB - We have previously reported that exposure of monkey embryos to 13-cis-retinoic acid (cRA) results in thymic defects. In this study, we analyzed lymphocyte and antigen-presenting cell populations at gestational days (GDs) 80-100 in the thymus, spleen, mesenteric lymph nodes, and gut-associated lymphoid tissue following a teratogenic dosing regimen of cRA (2.5 and 5 mg/kg) at GD14-27. Tissue sections were immunostained for T-cells (anti-CD3), B-cells (anti-CD20), dendritic cells (p55), and major histocompatibility class II (anti-HLA-DR). Digital images of spleen sections were analyzed to obtain the relative area occupied by the cell subsets within the white pulp (WP). Compared with controls, the T-cell dependent compartment of the spleen WP in specimens with perturbed thymic development (aplasia and severe hypoplasia) showed a reduction in size and proportion of CD3(+) T cells. Our findings indicate that cRA-induced thymic defects result in disrupted development of the splenic T-cell dependent compartment. PMID- 12110051 TI - Spontaneous pancreatic islet amyloidosis in 40 baboons. AB - Spontaneous amyloidosis occurs in many nonhuman primate species but remains difficult to diagnose and treat. Nonhuman primates continue to offer promise as animal models in which to study amyloidosis in humans. Amyloidosis was not diagnosed clinically but was found histologically in four male and 36 female baboons. The baboons averaged 18 years of age at death (range, 7-28 years). Clinical signs, if present, were hyperglycemia and cachexia. Blood glucose values were elevated in 12 of 30 baboons with available clinical pathology data. Four baboons had been clinically diagnosed as diabetic and three were treated with insulin. Amyloid was found in the islets of Langerhans of the pancreas in 40 baboons; 35 baboons had amyloid only in the islets of Langerhans. Amyloid was found in nonislet tissue of baboons as follows: five, nonislet pancreas; four, intestine and adrenal; three, kidney; two, prostate and spleen; and one each, lymph node, liver, gall bladder, stomach, tongue, urinary bladder, and salivary gland. Sections of paraffin-embedded tissues were evaluated for amyloid with hematoxylin and eosin (HE) and congo red (CR) staining, and using immunohistochemistry for human islet amyloid polypeptide (IAPP), calcitonin gene related peptide (CGRP), glucagon, pancreatic polypeptide (PP), somatostatin (SS), and porcine insulin. Islet amyloid was positive with HE in 40 baboons, with CR in 39 baboons, and with IAPP and CGRP in 35 baboons. IAPP and CGRP only stained islet amyloid. PP, SS, glucagon, and porcine insulin did not stain amyloid. Islet amyloidosis in the baboon appears to be difficult to diagnose clinically, age related, and similar to islet amyloidosis in other species. The baboon may be a good model for the study of islet amyloidosis in humans. PMID- 12110053 TI - Two cases of digital defects in Macaca mulatta infants and a survey of the literature. AB - Although congenital digital defects, particularly polydactyly, have been reported frequently in humans, their occurrence in rhesus macaques is relatively rare. We observed two cases of spontaneous digital defects in male rhesus monkey infants recently born at the California Regional Primate Research Center. One infant exhibited bilateral postaxial polydactyly and the other infant had bilateral oligodactyly with unilateral phalangeal duplication. In this report, we present the clinical/pathological details of these cases as well as discuss the embryology of normal and abnormal limb development. We will also summarize a variety of spontaneous and experimentally induced digital defects that have been reported in several nonhuman primate species. PMID- 12110054 TI - Fatal infection with human pinworm, Enterobius vermicularis, in a captive chimpanzee. AB - A fatal infection with human pinworms, Enterobius vermicularis, was found in a 26 year-old chimpanzee kept in a zoo. Grossly, the animal was highly emaciated, and had severe enteritis with cecal multifocal nodules and severe cholelithiasis. Histopathologically, a large number of human pinworms were observed in the nodular lesions in the cecum and intestinal wall. These migrating worms were surrounded by an inflammatory cell infiltration which lacked eosinophils. There were areas of multifocal hyperemia and/or hemorrhages in various organs including the entire gastrointestinal tract. Pinworms were also observed in the portal venule and parenchyma of the liver. A light infection with Strongyloides cf. stercoralis was also observed. PMID- 12110056 TI - The effect of body mass index on complications from cardiac surgery in the oldest old. AB - OBJECTIVES: Overweight (body mass index (BMI)>25) and obesity (BMI>30) are associated with adverse outcomes in the general population. In older people, an increased risk of adverse events was observed in leaner and overweight older subjects. We evaluated the association between BMI and cardiac surgery complications in subjects aged 75 and older. DESIGN: Retrospective review of complication rates compiled as part of the New York State Department of Health Adult Cardiac Surgery Report. SETTING: Academic tertiary care medical center. PARTICIPANTS: One thousand four hundred forty-eight subjects aged 75 and older who underwent cardiac surgery in an academic hospital between 1991 and 1999. MEASUREMENTS: Subjects were divided into tertiles based on their BMI. Logistic regression was used in multivariate analyses examining the association between tertiles of BMI (<23, 23-26,>26) and complications from cardiac surgery, adjusting for age and gender or using a full model adjusting for history of diabetes mellitus, hypertension, myocardial infarction (MI), congestive heart failure, smoking, chronic obstructive pulmonary disease, peripheral vascular disease, renal disease, surgical priority, age, and gender. RESULTS: Mean age was 79 (range 75-94); 58% of the subjects were male. The incidence of any postoperative complication, respiratory failure, or death was U-shaped, with higher incidence in the first tertile of BMI, followed by the last tertile and then the middle tertile. Subjects in the lowest tertile of BMI in the full model had a higher adjusted risk of stroke (odds ratio (OR) = 1.58, 95% confidence interval (CI) = 0.96-2.59), bleeding (OR = 1.71, 95% CI = 0.79-3.67), respiratory failure (OR = 1.65, 95%CI = 0.95-2.87), cardiovascular complications (stroke, MI, arrhythmia) (OR = 1.59, 95% CI = 0.93-2.73), and all complications (OR = 1.45, 95% CI = 1.05-2.00) than subjects in the middle tertile. The risk of most complications in subjects in the highest tertile of BMI was similar to that of subjects in the middle tertile except for wound infections (OR = 3.51, 95% CI = 0.92-13.33). The risk of death was higher in subjects in the lower tertile of BMI (OR = 1.97, 95% CI = 1.12-3.44) than in subjects in the middle tertile. The association between BMI and adverse cardiac surgical outcomes was stronger in men than women and decreased with advancing age. CONCLUSIONS: In the geriatric population, subjects with lower BMI have a higher risk of complications and death from cardiovascular surgery than subjects with higher BMI. An increased BMI does not increase the risk of complications from cardiovascular surgery, with the exception of wound infections. PMID- 12110057 TI - Inappropriate prescribing before and after nursing home admission. AB - OBJECTIVES: To compare the prevalence of inappropriate prescribing before and after nursing home admission and to determine which patient and physician characteristics are associated with inappropriate prescribing in the nursing home setting. DESIGN: A pre/post retrospective, cohort study. SETTING: All licensed nursing homes in Ontario, Canada. PARTICIPANTS: Nineteen thousand nine hundred eleven individuals aged 66 and older, newly admitted to nursing homes in Ontario between April 1, 1997, and March 31, 1999. MEASUREMENTS: For each patient in the cohort, a subset of the Beers criteria was used to characterize and compare the prevalence of inappropriate prescribing (as indicated by the prescription of one of 49 inappropriate drugs) before and after nursing home admission. A logistic regression model was used to study the association between inappropriate prescribing and patient and physician characteristics. RESULTS: The proportion of patients receiving a prescription for at least one inappropriate drug decreased from 25.4% before nursing home admission to 20.8% afterward (P <.001). Most patients who had been prescribed an inappropriate agent before nursing home entry had that agent discontinued after admission. The most commonly prescribed inappropriate drugs after nursing home admission were strongly anticholinergic antidepressants (6.4%) and long-half-life benzodiazepines (5.9%). Patients younger than 85 were more likely to receive inappropriate drug therapy (odds ratio (OR) = 1.25, 95% confidence interval (CI) = 1.15-1.35) than those aged 85 and older. Other significant predictors were having more than one prescriber (OR = 1.40, 95% CI = 1.29-1.51), having a physician aged 50 or older (OR = 1.14, 95% CI = 1.05-1.23), having a male physician (OR = 1.20, 95% CI = 1.05-1.37), having a nonspecialist physician (OR = 1.23, 95% CI = 1.01-1.49), having a nonurban physician (OR = 1.13, 95% CI = 1.03-1.24), and having a physician practicing outside the greater Ontario metropolitan area (OR = 1.31, 95% CI = 1.19-1.51). CONCLUSIONS: Although a substantial number of nursing home residents receive inappropriate drug therapy, the prevalence of inappropriate prescriptions in our cohort declined after nursing home admission despite an overall increase in drug use. Patient and physician characteristics were associated with inappropriate prescribing. Targeted interventions such as regionally based education programs or drug use restrictions may reduce the prevalence of inappropriate prescribing. PMID- 12110058 TI - Inappropriate medication prescribing in residential care/assisted living facilities. AB - OBJECTIVES: To identify the extent to which inappropriately prescribed medications (IPMs) are administered to older patients in residential care/assisted living (RC/AL) facilities and to describe facility and resident factors associated with receipt of one or more IPMs. DESIGN: Cross-sectional study of a stratified, representative sample of 193 facilities in four states. SETTING: We identified representative geographic regions within Florida, New Jersey, North Carolina, and Maryland and drew from within them a stratified random sample of 193 RC/AL facilities. Three subtypes of facilities were included in the sample: small homes (<16 beds), larger "new-model" homes, and larger "traditional" homes. PARTICIPANTS: Within each larger home, a random sample of residents aged 65 and older was approached for consent; in smaller homes all residents were approached. The overall enrollment rate was 92%; 2,078 residents were enrolled. MEASUREMENTS: Questionnaires and on-site observations were used to gather data on facility administration and staffing and resident characteristics. All prescription and nonprescription medications taken at least 4 of the 7 days before data collection were taken from medication administration records and coded for analysis. IPM designation was based on modification of a list developed by Beers et al. and currently used by nursing home surveyors. RESULTS: The majority of RC/AL patients were taking five or more medications; 16.0% of these patients were receiving IPMs. The most common IPMs were oxybutynin, propoxyphene, diphenhydramine, ticlopidine, doxepin, and dipyridamole. In multivariate analyses, using generalized estimating equations, IPM use was associated with the number of medications received, smaller facility bed size, moderate licensed practical nurse turnover, absence of dementia, low monthly fees, and absence of weekly physician visits. CONCLUSIONS: IPMs remain a problem in long-term care, but rates in these RC/AL settings compare favorably with those reported for other frail older populations, suggesting that use of medications with severe adverse effects may be waning. Regular physician facility visits may improve prescribing, as will attention to high-risk groups such as individuals on multiple medications. PMID- 12110059 TI - Is the use of benzodiazepines associated with incident disability? AB - OBJECTIVES: This study examined the association between benzodiazepine use and incident disability with an emphasis on elucidating whether the underlying health conditions that result in benzodiazepine use (confounding factors) or intrinsic adverse effects of benzodiazepine use were responsible for functional decline. DESIGN: Cohort study with follow-up of 4 to 5 years. SETTING: A health maintenance organization (HMO) in western Washington. PARTICIPANTS: Individuals aged 65 and older from a random sample of HMO enrollees who participated in a health promotion intervention trial (n = 1,519). MEASUREMENTS: Benzodiazepine use was ascertained from computerized pharmacy records. Self-reported functional status was assessed using a six-item physical function scale ranging from vigorous activity to self-care activities of daily living (ADLs). Two outcomes were examined: decline in overall physical function and limitations in self-care ADLs. Multivariate models were examined that included demographic characteristics, health status, and health behaviors that were likely to be confounders. Several analyses were conducted to examine whether benzodiazepine use or confounding factors were responsible for functional decline. RESULTS: Benzodiazepine use was significantly associated with incident loss of physical function (hazard ratio (HR) = 1.51, 95% confidence interval (CI) = 1.02-2.24) in the fully adjusted model. Although use of benzodiazepines was associated with limitations in ADLs, it was not significant when adjusting for other factors (HR = 1.71, 95% CI = 0.87-3.34). Several of our findings suggest that the health conditions leading to benzodiazepine use may partly or fully explain these associations: (1) use of anxiolytic benzodiazepines (HR = 1.95, 95% CI = 1.24 3.07), but not hypnotic agents (HR = 1.21, 95% CI = 0.73-2.00), was associated with functional decline; (2) adjustment for health status variables minimized these associations; and (3) there was little evidence of dose response. CONCLUSIONS: A modestly increased risk for decline in physical function was associated with benzodiazepine use, especially of anxiolytic agents. The health conditions that result in benzodiazepine use may be more important in the pathogenesis of disability than benzodiazepine use itself. Although there are many reasons for avoiding benzodiazepines in older adults, it is still unclear whether use contributes independently to functional decline. PMID- 12110060 TI - Endothelial function in healthy younger and older hyperhomocysteinemic subjects. AB - OBJECTIVES: To compare endothelium-dependent vasomotor response in healthy younger and older subjects without classic cardiovascular risk factors, with high and normal fasting homocysteine (tHcy) levels. DESIGN: We compared endothelium dependent vasodilatation, using ultrasound, in healthy younger (aged 18-40) and older (> or =70) people with normal (<13 micromol/L) and high (>15 micromol/L) tHcy levels. Exclusion criteria were smoking, personal history of cardiovascular disease, hypertension, chronic diseases, vitamin intake, obesity, abnormal serum lipids levels, and creatinine higher than 130 micromol/L. SETTING: Research laboratory. MEASUREMENTS: In addition to tHcy levels, serum folate and vitamin B12 levels were measured. RESULTS: We studied 17 younger and 12 older hyperhomocysteinemic subjects and respective aged-matched normohocysteinemic subjects. Endothelium-dependent vasodilatation was lower in the hyperhomocysteinemic older people (P <.01) than in all younger subjects and in normohomocysteinemic older people. Serum vitamin B12 levels were higher in younger and older normal controls. Folic acid levels were higher in younger controls and in both older groups. CONCLUSIONS: This study shows an effect of high circulating tHcy on vascular reactivity in older people. Because serum levels of tHcy are associated with nutritional status of vitamin B12 and folic acid, prospective studies are necessary to demonstrate the effects of a long-term nutritional supplementation with vitamins on vascular function and global cardiovascular risk. PMID- 12110061 TI - Tai Chi Chuan training is associated with enhanced endothelium-dependent dilation in skin vasculature of healthy older men. AB - OBJECTIVES: The vascular endothelium modulates vascular tone by synthesizing and metabolizing vasoactive substances. Endothelium-dependent vasodilation declines with age. This study investigated whether Tai Chi Chuan (TCC) training could enhance endothelial function in the skin vasculature of older men. SETTING: Community setting. DESIGN: Basic hemodynamic characteristics and skin vascular response to endothelium-dependent and -independent vasodilators were studied. PARTICIPANTS: Ten older men who practiced TCC, 10 older healthy sedentary men, and 12 younger healthy sedentary men. The older TCC subjects had practiced classical Yang TCC for a mean +/- standard deviation of 11.2 +/- 3.4 years; mean attendance was 5.1 +/- 1.8 times weekly. Sedentary subjects had not participated in any regular exercise training for at least 5 years. MEASUREMENTS: Different doses of 1% acetylcholine (ACh) and 1% sodium nitroprusside (SNP) were iontophoretically applied to the skin of subjects' lower legs, and cutaneous microvascular perfusion responses were determined by laser doppler measurements. Additionally, arterial and venous hemodynamic variables were measured by impedance plethysmograph. RESULTS: The older TCC group had higher lower leg arterial blood flow (LABF); LABF in response to reactive hyperemia; and lower leg venous capacity, tone and blood flow than their sedentary counterparts, but the older TCC group displayed similar arterial and venous hemodynamic variables to the younger sedentary group. The younger sedentary group had a higher ACh-induced cutaneous perfusion and a higher ratio of ACh- to SNP-induced cutaneous perfusion than the two older groups. The older TCC group showed a higher ACh-induced cutaneous perfusion and a higher ratio of ACh- to SNP-induced cutaneous perfusion than the older sedentary group. Skin vascular responses to SNP did not differ significantly between the three groups. CONCLUSIONS: Regular practice of TCC is associated with enhanced endothelium-dependent dilation in skin vasculature of older individuals. Moreover, TCC training may delay the age-related decline of venous compliance and hyperemic arterial response. PMID- 12110062 TI - Identifying postmenopausal women at high risk of fracture in populations: a comparison of three strategies. AB - OBJECTIVES: To describe the prevalence of risk factors for women at high risk of fracture in a population-based sample of postmenopausal women who were not using hormone replacement therapy (HRT), to demonstrate how the estimated prevalence of women at high risk of future fracture is affected by the different criteria used for classification, and to characterize the populations identified and missed by each of the criteria. A key study objective was to compare the proportion of postmenopausal women at high risk of fracture in a managed care population using several different definitions of who is at high risk. DESIGN: The Osteoporosis Population-based Risk Assessment study, a randomized trial of three screening strategies. SETTING: Conducted at Group Health Cooperative in western Washington state. PARTICIPANTS: Women aged 60 to 79 who had not used HRT for at least 12 months were chosen at random. MEASUREMENTS: In one of the trial arms, 428 women had their bone mineral density (BMD) measured at the hip and spine (L1-L4) using dual energy x-ray absorptiometry. Minimum t scores and z scores at all sites were used for classification. Risk factors for fractures were assessed at the time of the BMD scan. RESULTS: Guidelines based on the Study of Osteoporotic Fractures classified 25.1% of the women as being at high risk of fracture, compared with 30.0% and 68.0% using World Health Organization (WHO) recommendations and National Osteoporosis Foundation guidelines, respectively. Classification based on low BMD alone (WHO) failed to include more than 50% of women who had already experienced a clinical fracture. CONCLUSIONS: Prevalence of women at high risk of fracture not using HRT varies notably depending on the criteria used for identification. The criteria used to identify women to target for primary and secondary prevention of osteoporotic fractures has major implications for population-based prevention strategies. PMID- 12110063 TI - Pain in cognitively impaired nursing home residents: how well are physicians diagnosing it? AB - OBJECTIVES: This study examined the reliability and validity of geriatricians' assessments of pain in cognitively impaired nursing home residents. DESIGN: Cross sectional analysis. SETTING: A large suburban nursing home. PARTICIPANTS: Seventy nine nursing home residents participated in the study. Of these, 31 had mild/moderate cognitive impairment (average Mini-Mental State Examination (MMSE) = 16.04) and 48 were severely cognitively impaired (average MMSE = 1.91). More than 80% of the participants were female, and the average age was 87. MEASUREMENTS: Two geriatricians from outside the nursing home examined laboratory results, performed a physical examination, and completed a detailed assessment of pain. The personal geriatricians of 42 of the participants also completed the same assessment. RESULTS: Intergeriatrician agreement rates were statistically significant and moderate in magnitude. When examined by subgroup, the correlations were significant only for those with mild/moderate impairment. Some of the geriatricians' ratings of pain correlated significantly with residents' self-reports. All relationships were weaker in the severely cognitively impaired group. Ratings of greater pain were significantly correlated with higher cognitive functioning. CONCLUSIONS: The results validate geriatricians' evaluations of pain during a medical examination for moderately impaired persons and question their ability to evaluate pain in the severely cognitively impaired. There is a need for increased awareness of pain in this population and a need for improved methodologies to identify it. PMID- 12110064 TI - Cognitive impairment, depressive symptoms, and functional decline in older people. AB - OBJECTIVES: Although cognitive impairment and depressive symptoms are associated with functional decline, it is not understood how these risk factors act together to affect the risk of functional decline. The purpose of this study is to determine the relative contributions of cognitive impairment and depressive symptoms on decline in activity of daily living (ADL) function over 2 years in an older cohort. DESIGN: Prospective cohort study. SETTING: A U.S. national prospective cohort study of older people, Asset and Health Dynamics in the Oldest Old. PARTICIPANTS: Five thousand six hundred ninety-seven participants (mean age 77, 64% women, 86% white) followed from 1993 to 1995. MEASUREMENTS: Cognitive impairment and depressive symptoms were defined as the poorest scores: 1.5 standard deviations below the mean on a cognitive scale or 1.5 standard deviations above the mean on validated depression scales. Risk of functional decline in participants with depressive symptoms, cognitive impairment, and both, compared with neither risk factor, were calculated and stratified by baseline dependence. Analyses were adjusted for demographics and comorbidity. RESULTS: Eight percent (n = 450) of subjects declined in ADL function. In participants who were independent in all ADLs at baseline, the relative risk (RR) of 2-year functional decline was 2.3 (95% confidence interval (CI) = 1.7-3.1) for participants with cognitive impairment, 1.9 (95% CI = 1.3-2.6) for participants with depressive symptoms, and 2.4 (95% CI = 1.4-3.7) for participants with cognitive impairment and depressive symptoms. In participants who were dependent in one or more ADLs at baseline, RR of 2-year functional decline was 1.9 (95% CI = 1.2-2.8) for participants with cognitive impairment, 0.6 (95% CI = 0.3-1.3) for participants with depressive symptoms, and 1.5 (95% CI = 0.8-2.6) for participants with cognitive impairment and depressive symptoms. CONCLUSIONS: In participants with no ADL dependence at baseline, cognitive impairment and depressive symptoms are risk factors for decline, but that, in participants with dependence in ADL at baseline, cognitive impairment, but not depressive symptoms, is a risk factor for additional decline. PMID- 12110065 TI - Predicting cognitive impairment in high-functioning community-dwelling older persons: MacArthur Studies of Successful Aging. AB - OBJECTIVES: To examine whether simple cognitive tests, when applied to cognitively intact older persons, are useful predictors of cognitive impairment 7 years later. DESIGN: Cohort study. SETTING: Durham, North Carolina; East Boston, Massachusetts; and New Haven, Connecticut, areas that are part of the National Institute on Aging Established Populations for Epidemiological Studies of the Elderly. PARTICIPANTS: Participants, aged 70 to 79, from three community-based studies, who were in the top third of this age group, based on physical and cognitive functional status. MEASUREMENTS: New onset of cognitive impairment as defined by a score of less than 7 on the Short Portable Mental Status Questionnaire (SPMSQ) in 1995. RESULTS: At 7 years, 21.8% (149 of 684 subjects) scored lower than 7 on the SPMSQ. Using multivariate logistic regression, three baseline (1988) cognitive tests predicted impairment in 1995. These included two simple tests of delayed recall-the ability to remember up to six items from a short story and up to 18 words from recall of Boston Naming Test items. For each story item missed, the adjusted odds ratio (AOR) for cognitive impairment was 1.44 (95% confidence interval (CI) = 1.16-1.78, P <.001). For each missed item from the word list, the AOR was 1.20 (95% CI = 1.09-1.31, P <.001). The Delayed Recognition Span, which assesses nonverbal memory, also predicted cognitive impairment, albeit less strongly (odds ratio = 1.06 per each missed answer, 95% CI = 1.003-1.13, P =.04). CONCLUSIONS: This study identifies measures of delayed recall and recognition as significant early predictors of subsequent cognitive decline in high-functioning older persons. Future efforts to identify those at greatest risk of cognitive impairment may benefit by including these measures. PMID- 12110067 TI - Reproducibility of blood pressure variation in older ambulatory and bedridden subjects. AB - OBJECTIVES: We investigated the influence of ambulation on the reproducibility of circadian blood pressure variation in older nursing home residents. DESIGN: Ambulatory blood pressure monitoring was performed twice in 37 older nursing home residents. SETTING: Nursing home in Japan. PARTICIPANTS: Subjects included 18 ambulatory nursing home residents who had no limitation on physical activity and 19 bedridden residents who did not participate in physical activity. MEASUREMENTS: Twenty-four-hour, daytime, and nighttime blood pressure levels and their variability. RESULTS: The 24-hour and daytime variability of systolic blood pressure (SBP) was significantly greater in ambulatory than in bedridden subjects, whereas nighttime variability was similar. Significant correlations in SBP averaged for the whole day, daytime, and nighttime were observed between the two examinations in ambulatory (r =.80-.83) and bedridden (r =.83-.91) subjects, but the variabilities of SBP for the whole day and during the daytime of the first measurement were correlated with those of the second measurement in bedridden (r =.67 and r =.47, respectively) but not in ambulatory (r =.39 and r =.28, respectively) subjects. Significant correlations were found between the nocturnal SBP changes at two occasions in both ambulatory (r =.50) and bedridden (r =.51) subjects, but the dipper versus nondipper profiles, defined as reduction in SBP of greater than 10% versus not, showed low reproducibility in ambulatory subjects; five ambulatory (28%) and one bedridden (5%) subjects showed divergent profiles between the two examinations. CONCLUSIONS: The reproducibility of blood pressure variation in nursing home residents is influenced by ambulation. PMID- 12110066 TI - The effects of mammographic detection and comorbidity on the survival of older women with breast cancer. AB - OBJECTIVES: To determine an upper age limit or quantifiable level of comorbidity that would render mammography screening ineffectual in decreasing mortality in women aged 65 and older. DESIGN: Retrospective cohort study. SETTING: Upper midwestern United States. PARTICIPANTS: Five thousand one hundred eighty-six predominantly Caucasian women aged 65 to 101 diagnosed with invasive breast cancer from 1986 through 1994. Data were obtained from The Upper Midwest Tumor Registry System, a regional consortium database in Minnesota, North Dakota, and South Dakota. MEASUREMENTS: Relative risks (RRs) of death were computed for patients with mammographically detected tumors, stratified by age and comorbidity. Survival analysis was performed, stratified by level of comorbidity and method of tumor detection. RESULTS: Patients with mammographically detected tumors and no comorbidity experienced significantly lower RRs of death in every age group (range P <.001 to P =.039). Women with mammographically detected tumors and mild to moderate comorbidity had RRs of death as follows: age 65 to 69 (RR = 0.32, 95% confidence interval (CI) = 0.15-0.69), age 70 to 74, (RR = 0.45, 95% CI = 0.22-0.91); age 75 to 79 (RR = 0.47, 95% CI = 0.25-0.88), age 80 and older (RR = 0.52, 95% CI = 0.33-0.80). Women with severe or multiple comorbidities experienced no improvement in survival with mammographically detected tumors. CONCLUSIONS: Mammographic detection of breast cancer may be associated with a significantly decreased risk of death for older women of all ages, even for women with mild to moderate levels of comorbidity, but for older women with severe or multiple comorbidities, mammography is not associated with improvement in overall survival. PMID- 12110068 TI - Prevalence of and association between silent myocardial ischemia and new coronary events in older men and women with and without cardiovascular disease. AB - OBJECTIVES: To investigate the prevalence of silent ischemia (SI) in older men and women detected by 24-hour ambulatory electrocardiograms (AECGs) and the association between SI and new coronary events. DESIGN: In a prospective study, the prevalence of SI detected by 24-hour AECGs and the incidence of new coronary events in 915 older men and 1,874 older women with coronary artery disease (CAD); with hypertension, valvular disease, or cardiomyopathy without CAD; and with no cardiovascular disease were investigated. SETTING: Large long-term healthcare facility. PARTICIPANTS: Nine hundred fifteen men, mean age 80, and 1,874 women, mean age 81. MEASUREMENTS: The prevalence of SI and the incidence of new coronary events in older men and women. RESULTS: SI was present in 34% of men and 33% of women with CAD; 15% of men and 14% of women with hypertension, valvular disease, or cardiomyopathy without CAD; and 6% of men and 5% of women with no cardiovascular disease. At 45-month follow-up in men and 47-month follow-up in women, SI significantly increased the incidence of new coronary events by 2.0 times in men and women with CAD (P <.001); by 1.8 times in men and 1.7 times in women with hypertension, valvular disease, or cardiomyopathy without CAD (P <.001); and by 6.3 times in men (P =.018) and 4.4 times in women (P =.008) with no cardiovascular disease. CONCLUSIONS: SI increases the incidence of new coronary events in older men and women with CAD, with hypertension, valvular disease, or cardiomyopathy without CAD, and with no cardiovascular disease. PMID- 12110069 TI - Caregiver attitudes and hospitalization risk in michigan residents receiving home and community-based care. AB - OBJECTIVES: To study a cohort of participants in home- and community-based services (HCBS) in Michigan to evaluate the relationship between (1) caregiver attitudes and participant characteristics and (2) the risk of hospitalization. SETTING: HCBS programs funded by Medicaid or state/local funds in Michigan. PARTICIPANTS: Five hundred twenty-seven individuals eligible for HCBS in Michigan were studied. These HCBS participants were randomly selected clients of all agencies providing publicly funded HCBS in Michigan from November 1996 to October 1997. MEASUREMENTS: Data for this study were collected using the Minimum Data Set for Home Care. Assessments were collected longitudinally, and the baseline (initial admission assessment) and 90-day follow-up assessments were used. Key measures were caregiver attitudes (distress, dissatisfaction, and decreased caregiving ability) and HCBS participant characteristics (cognition, functioning, diseases, symptoms, nutritional status, medications, and disease stability). Multinomial logistic regression was used to evaluate how these characteristics were associated with the competing risks of hospitalization and death within 90 days of admission to HCBS. RESULTS: We found a strong association between caregiver dissatisfaction (caregiver dissatisfied with the level of care the home care participant was currently receiving) and an increased likelihood of hospitalization. HCBS participant cancer, chronic obstructive pulmonary disease, pain, and flare-up of a chronic condition were also associated with increased hospitalization. Poor food intake and prior hospitalization were associated with hospitalization and death. CONCLUSIONS: We conclude that, within a cohort of people receiving HCBS who are chronically ill, highly disabled, and at high risk for hospitalization and death, interventions addressing caregiver dissatisfaction, pain control, and medical monitoring should be evaluated for their potential to decrease hospitalization. PMID- 12110070 TI - Screening for Alzheimer's disease: the memory impairment screen versus the conventional three-word memory test. AB - OBJECTIVES: To improve screening for Alzheimer's disease (AD) with the Memory Impairment Screen (MIS), a 4-minute, four-item delayed free and cued recall memory test with controlled learning and high discriminative validity. To assess the discriminative validity of the MIS for AD and to compare it with the conventional three-word memory test, a delayed free recall task, widely recommended as a dementia-screening test in clinical practice. DESIGN: Cross sectional validation study nested within a longitudinal study of aging and dementia. The MIS and the standard three-word memory task were administered as part of a comprehensive neurological and neuropsychological evaluation. SETTING: Einstein Aging Study at the Albert Einstein College of Medicine, Bronx, New York. PARTICIPANTS: Two hundred forty community-dwelling older adults. MEASUREMENTS: Sensitivity, specificity, and positive predictive value (PPV) were calculated for the MIS and three-word memory test as screening tests for AD. RESULTS: In comparison with the three-word memory task, the MIS had higher sensitivity (.86 vs.65), higher specificity (.97 vs.85), and greater PPV (.80 vs.37) as a screen for AD. CONCLUSIONS: The MIS had high discriminative validity as a screening test for AD and substantially outperformed the three-word memory task. Given its validity and brevity, the MIS has important advantages as an AD screen for use in primary care. PMID- 12110071 TI - Screening for depression in patients in long-term care facilities: a randomized controlled trial of physician response. AB - OBJECTIVES: To determine the effect of a screening protocol using the Geriatric Depression Scale (GDS) on the frequency of primary care physicians' decisions to prescribe drug therapy or refer long-term care patients with possible depression to mental health care. DESIGN: Case-finding phase, followed by a randomized controlled trial of the effect of a physician-targeted intervention on antidepressant prescribing or referral to mental health services. SETTING: Twenty two nonacademic long-term care facilities. PARTICIPANTS: One hundred three of 1,602 patients aged 65 and older who met criteria for cognitive function and untreated symptoms of depression. INTERVENTION: The 77 physicians of these patients were randomized as clusters into an early notification (experimental) or a delayed notification (control) group. MEASUREMENTS: Frequency of physician response (mental health consult or antidepressant therapy) at 4 and 8 weeks from notification, physician follow-up, and factors associated with physician response. RESULTS: Frequency of physician response in the early group (25%) was greater than in the delayed group (2%) (P <.005) 4 weeks from baseline. Physician response rate when the groups were combined was 36% (95% confidence interval (CI) = 26%-46%) 8 weeks from notification. Overall, there was evidence of physician action after letters of notification in 69% (95% CI = 60%-78%) of cases. Univariate logistic regression suggested that physicians' decisions were primarily associated with physician-related characteristics. CONCLUSIONS: Screening of long-term care patients for depression can increase the frequency of treatment or referral by primary care physicians. PMID- 12110072 TI - Resistance exercise and physical performance in adults aged 60 to 83. AB - OBJECTIVES: This investigation examined the effect of 6 months of high- or low intensity resistance exercise on muscular strength and endurance and stair climbing ability in adults aged 60 to 83. DESIGN: A randomized controlled trial. SETTING: University of Florida Center for Exercise Science. PARTICIPANTS: Sixty two men and women completed the study protocol. Subjects were matched for strength and randomly assigned to a control (n = 16), low-intensity (LEX, n = 24), or high-intensity (HEX, n = 22) group. INTERVENTION: Six months of progressive, whole-body resistance training. Subjects trained at 50% of their one repetition maximum (1RM) for 13 repetitions (LEX) or 80% of 1RM for eight repetitions (HEX) three times per week for 24 weeks using resistance machines. One set each of 12 exercises was performed. MEASUREMENTS: One-repetition maximum was measured for eight different exercises. Muscular endurance was measured using leg press and chest press machines. Low back strength was measured using a lumbar extension machine. Stair climbing ability was assessed as the time to ascend one flight of stairs. RESULTS: 1RM significantly increased for all exercises tested for the HEX and LEX groups (P < or =.050). The increases in total strength (sum of all eight 1RMs) were 17.2% and 17.8% for the LEX and HEX groups, respectively. Muscular endurance improved by 79.2% and 105.0% for the leg press, and 75.5% and 68.0% for the chest press for the LEX and HEX groups, respectively. The time to ascend one flight of stairs significantly decreased for both the LEX and HEX groups (P < or =.050). Lumbar extension strength increased by 62.6% and 39.5% for the LEX and HEX groups, respectively. CONCLUSIONS: These data indicate that significant and similar improvements in strength, endurance, and stair climbing time can be obtained in older adults as a consequence of high- or low-intensity resistance exercise training. These findings may have an effect on how resistance exercise is prescribed to older adults. PMID- 12110073 TI - Profiles of older medicare decedents. AB - OBJECTIVES: To evaluate the usefulness of a clinical scheme to classify older decedents to better understand the issues associated with healthcare use and costs in the last year of life. DESIGN: We analyzed Medicare claims data for a random sample of 0.1% of all Medicare beneficiaries with expenditures between 1993 and 1998. This sample yielded 7,966 deaths. SETTING: Medicare claims data. PARTICIPANTS: Medicare beneficiaries. MEASUREMENTS: We classified decedents into groups representing four trajectories at the end of life: sudden death, terminal illness, organ failure, and frailty. RESULTS: Ninety-two percent of decedents were captured by the profiling strategy. The four trajectory groups had distinct patterns of demographics, care delivery, and Medicare expenditures. Frailty was a dominant pattern, with 47% of all decedents, whereas sudden death claimed only 7%; cancer claimed 22%, and organ system failure, 16%. CONCLUSIONS: The clinical scheme to classify decedents appears to fit most decedents and to form groups with substantial clinical differences. Acknowledging the differences among these groups may be a fruitful way to evaluate expenditures and develop strategies to improve care at the end of life. PMID- 12110074 TI - Racial and ethnic differences in place of death: United States, 1993. AB - OBJECTIVES: To examine racial and ethnic differences in place of death, adjusting for likely confounders. DESIGN: A retrospective cohort analyzed using multinomial logistic regression. SETTING: United States in 1993. PARTICIPANTS: A nationally representative sample of 22,658 deaths in 1993 from the National Mortality Followback Survey. MEASUREMENTS: Place of death as determined on the death certificate, with controls for age, sex, income, education, and cause of death. The outcomes of interest were death in a hospital during an inpatient stay, death in a nursing home, death in a private residence, or death in some other place. RESULTS: After adjustment, 43% of whites die after an inpatient hospital stay, as do 50% of blacks and 56% of Mexican Americans. Twenty percent of whites, 22% of Mexican Americans, and 14% of blacks die in nursing homes. Twenty-two percent of whites, 18% of blacks, and 9% of Mexicans die in a private residence. CONCLUSIONS: There are substantial differences between whites, blacks, and Mexican Americans in place of death that cannot be explained by differences in age, sex, income, education, and causes of death between the groups. PMID- 12110075 TI - Prevalence of impaired swallowing in institutionalized older people in taiwan. AB - OBJECTIVES: To investigate the prevalence of impaired swallowing in residents at long-term care facilities (LTCFs) in Taiwan. DESIGN: A chart review, a structured questionnaire completed at interview, a neurological examination, and a timed swallowing test were used to assess impairment and to gather demographic data. SETTING: Nine skilled nursing facilities and nine intermediate-care facilities in metropolitan Taipei. PARTICIPANTS: One thousand two hundred twenty-one conscious and unconscious residents with a mean age of 77.07. MEASUREMENTS: Impaired swallowing was defined when a subject met two or more of the following criteria: self-report of swallowing difficulty, a score of 2 or more derived from a swallowing questionnaire combined with a neurological examination investigating symptoms and signs of impairment, and coughing/choking during a timed swallowing test or a measured swallowing rate (volume swallowed per second) below the 10th percentile as derived from a gender-based study of an older community in Taipei. RESULTS: Of the 1,221 subjects, 356 (29.2%) were fed by tube. The prevalence rates for impaired swallowing as estimated were 97.5% and 31.9% for tube-fed and non-tube-fed subjects respectively, whereas the overall prevalence for tube-fed and non-tube-fed subjects altogether was 51.0%. CONCLUSIONS: The findings may serve to increase awareness of this problem among healthcare professionals in LTCFs. Further research is contemplated to investigate whether early identification makes a difference in treatment choices and outcomes. PMID- 12110076 TI - Deaths between bedrails and air pressure mattresses. AB - OBJECTIVES: To describe how patients die by becoming trapped between therapeutic air pressure mattresses and bed rails. DESIGN: A retrospective review of all voluntary reports deaths in beds with air mattresses that can be found in the Food and Drug Administration's on-line databases of adverse medical events that cover 1994 to 2001. SETTING: Death reports come from manufacturers, medical staff, and coroners and describe deaths in hospitals, nursing homes, and home care, although type of care site is often not given. MEASUREMENTS: Event descriptions were reviewed to determine how the person became entrapped in the rail and how responsibility for the event was allocated. RESULTS: There were 35 deaths involving many product lines. Twenty-one deaths involved overlay air mattresses placed on top of a regular mattress. Thirteen patients died in beds with built-in air mattresses. Compression of the mattress allowed an off-center person to slide against the rail where reexpansion of the mattress kept the person compressed against the rail. Two patterns were seen. In one, the mattress bunched up behind a person who was lying on the side of the bed, pushing the neck against a bedrail. In the second type, a patient died after sliding off the bed and having the neck or chest compressed between the rail and bed. Manufacturers attributed the deaths to poor clinical decision-making or inadequate monitoring. CONCLUSIONS: Lethal asphyxiation in beds with air pressure mattresses is a variant of bedrail-mattress entrapment that requires redesign by bed manufacturers and risk awareness by clinicians. PMID- 12110077 TI - Profiling nursing homes using Bayesian hierarchical modeling. AB - OBJECTIVES: New methods developed to improve the statistical basis of provider profiling may be particularly applicable to nursing homes. We examine the use of Bayesian hierarchical modeling in profiling nursing homes on their rate of pressure ulcer development. DESIGN: Observational study using Minimum Data Set data from 1997 and 1998. SETTING: A for-profit nursing home chain. PARTICIPANTS: Residents of 108 nursing homes who were without a pressure ulcer on an index assessment. MEASUREMENTS: Nursing homes were compared on their performance on risk-adjusted rates of pressure ulcer development calculated using standard statistical techniques and Bayesian hierarchical modeling. RESULTS: Bayesian estimates of nursing home performance differed considerably from rates calculated using standard statistical techniques. The range of risk-adjusted rates among nursing homes was 0% to 14.3% using standard methods and 1.0% to 4.8% using Bayesian analysis. Fifteen nursing homes were designated as outliers based on their z scores, and two were outliers using Bayesian modeling. Only one nursing home had greater than a 50% probability of having a true rate of ulcer development exceeding 4%. CONCLUSIONS: Bayesian hierarchical modeling can be successfully applied to the problem of profiling nursing homes. Results obtained from Bayesian modeling are different from those obtained using standard statistical techniques. The continued evaluation and application of this new methodology in nursing homes may ensure that consumers and providers have the most accurate information regarding performance. PMID- 12110078 TI - Androgen supplementation in older women: too much hype, not enough data. AB - Androgen supplementation in women has received enormous attention in the scientific and lay communities. That it enhances some aspects of cognitive function, sexual function, muscle mass, strength, and sense of well-being is not in question. What is not known is whether physiological testosterone replacement can improve health-related outcome in older women without its virilizing side effects. Although it is assumed that the testosterone dose-response relationship is different in women than in men and that clinically relevant outcomes on the above-mentioned effects can be achieved at lower testosterone doses, these assumptions have not been tested rigorously. Androgen deficiency has no clear-cut definition. Clinical features may include impaired sexual function, low energy, depression, and a total testosterone level of less than 15 ng/dL, the lower end of the normal range. Measurement of free testosterone is ideal, because it provides a better estimate of the biologically relevant fraction. It is not widely used in clinical practice, because some methods of measuring free testosterone assay are hampered by methodological difficulties. In marked contrast to the abrupt decline in estrogen and progesterone production at menopause, serum testosterone is lower in older women than in menstruating women, with the decline becoming apparent a decade before menopause. This article reviews testosterone's effects on sexual function, cognitive function, muscle mass, body composition, and immune function in postmenopausal women. PMID- 12110079 TI - Attitudes toward working on interdisciplinary healthcare teams: a comparison by discipline. AB - Interdisciplinary teams are important in providing care for older patients, but interdisciplinary teamwork is rarely a teaching focus, and little is known about trainees' attitudes towards it. To determine the attitudes of second-year post graduate (PGY-2) internal medicine or family practice residents, advanced practice nursing (NP), and masters-level social work (MSW) students toward the value and efficiency of interdisciplinary teamwork and the physician's role on the team, a baseline survey was administered to 591 Geriatrics Interdisciplinary Team Training participants at eight U.S. academic medical centers from January 1997 to July 1999. Most students in each profession agreed that the interdisciplinary team approach benefits patients and is a productive use of time, but PGY-2s consistently rated their agreement lower than NP or MSW students. Interprofessional differences were greatest for beliefs about the physician's role; 73% of PGY-2s but only 44% to 47% of MSW and NP trainees agreed that a team's primary purpose was to assist physicians in achieving treatment goals for patients. Approximately 80% of PGY-2s but only 35% to 40% of MSW or NP trainees agreed that physicians have the right to alter patient care plans developed by the team. Although students from all three disciplines were positively inclined toward medical interdisciplinary teamwork, medical residents were the least so. Exposure to interdisciplinary teamwork may need to occur at an earlier point in medical training than residency. The question of who is ultimately responsible for the decisions of the team may be an "Achilles heel," interfering with shared decision-making. PMID- 12110080 TI - Similarities and differences in attitudes toward long-term care between Japanese Americans and Caucasian Americans. AB - The purpose of this study was to compare attitudes toward the use of long-term care between older Japanese Americans (n = 1,244) and older Caucasian Americans (n = 1,354). When presented with a hypothetical situation in which they have dementia, 39% of older Japanese Americans and 42% of older Caucasians intended to be cared for at home, whereas 53% versus 38%, respectively, intended to use nursing home care (P <.001). If the hypothetical situation was hip fracture, 81% of older Japanese Americans and 72% of older Caucasians intended to be cared for at home, with 13% of both groups intending to use nursing home care (P = NS). The subjects' perceptions of what their families, friends, ministers, and communities would want them to choose differed, with more uncertainty among Caucasians (P <.001). For provision of home care, Japanese Americans were more likely to rely on loved ones than Caucasians, who were more likely to rely on paid providers. Multivariate logistic regression showed ethnicity to be independently related to intention to use nursing home care in the dementia scenario, controlling for demographic variables. Being married lowered the odds of intending to use nursing homes in any situation. We conclude that Caucasian Americans intend to use paid home health care at higher rates than Japanese Americans if they become disabled by dementia. Japanese Americans demonstrated more certainty about the influences of others on their opinions, suggesting a more stable cultural norm in this population, and intended to use more nursing home care in the event of permanent debility (dementia). PMID- 12110081 TI - Is the verdict out? A systematic review of pharmacotherapy for hypertension in the elderly. PMID- 12110082 TI - Paradigm of vascular aging: conceptual strides slowed by technical limitations. PMID- 12110083 TI - Can we identify women at high risk of osteoporotic fractures? PMID- 12110084 TI - Drug use in older people. PMID- 12110085 TI - Depressive symptoms in older Estonians: prevalence and models. PMID- 12110086 TI - Risk of delirium with concomitant use of tolterodine and acetylcholinesterase inhibitors. PMID- 12110087 TI - Not all clock-drawing tasks are the same. PMID- 12110088 TI - Falls' guidelines and osteoporosis assessment. PMID- 12110090 TI - Predictive value of renal histological changes for postoperative renal function improvement in children with congenital ureteropelvic junction stenosis. AB - BACKGROUND: The aim of this study was to evaluate the relationship between renal function, as measured by diuretic radionuclide renography, and the outcome of pyeloplasty. A study was designed to evaluate renal parenchymal biopsy specimens derived from children undergoing corrective surgery for ureteropelvic junction (UPJ) stenosis, and compare these to preoperative and postoperative renal function status. METHODS: Thirty-five children with congenital unilateral UPJ stenosis were evaluated. In addition to all conventional diagnostic procedures for UPJ stenosis, differential renal functional (DRF) activity was assessed in each of these children by obtaining 99mTc diethylenetriaminepentaacetic acid renogram curves. All children underwent dismembered pyeloplasty, and follow-up renogram evaluation was conducted 6 and 12 months after surgical repair. Biopsy specimens from renal cortical regions obtained during the surgical correction of UPJ stenosis were evaluated, and changes in renal histology were graded from I to V according to their severity. Spearman's correlation test was used to compare the histological evaluation results and the basal, 6- and 12-month follow-up DRF findings. A Wilcoxon paired test was used to evaluate statistical differences between values. RESULTS: The findings showed a positive correlation between the severity of histological changes and DRF activity. All kidneys (22) with a DRF activity value of < 40% preoperatively demonstrated at least grade III changes when biopsy specimens were examined. Of children with a DRF activity value > 40% (13), only three showed severe histological changes. Histological grades were correlated between basal (r = -0.4; P = 0.019), 6-month (r = 0.54; P = 0.002) and 12-month (r = 0.54; P = 0.02) findings. In the Wilcoxon paired test, there was a statistically significant difference between basal and 6-month values (P < 0.05), and also between basal and 12-month values (P < 0.01). There was no statistically significant difference between 6- and 12-month values (P > 0.20). CONCLUSION: Comparative evaluation of postoperative renal function with DRF activity and renal parenchymal histological alterations revealed a close correlation in terms of renal function improvement potential following reconstructive surgery in children with UPJ stenosis. PMID- 12110091 TI - Involvement of adrenomedullin induced by hypoxia in angiogenesis in human renal cell carcinoma. AB - BACKGROUND: Adrenomedullin (AM) has pluripotent activities and is involved in the regulation of vasomotor tone, cell differentiation and embryogenesis. However, the expression and pathophysiological role of AM has not been determined in human renal cell carcinoma (RCC). METHODS: Twenty-six RCC specimens and three cultured human RCC cell lines (A498, SN12C and KPK-13) were analyzed. Expression of AM was determined by immunohistochemistry and reverse transcription-polymerase chain reaction (RT-PCR) analysis. The correlation between AM expression and microvessel count (MVC) in RCC specimens was examined to determine if AM plays a role in tumor angiogenesis. The correlation between the expression of AM and vascular endothelial growth factor (VEGF) was also investigated. Lastly, the effect of hypoxia upon the mRNA expression of AM, VEGF and hypoxia inducible factor-1 (HIF 1) by RCC cell lines was determined. RESULTS: Immunohistochemistry indicated that AM and VEGF were primarily localized in the cytosol of RCC cells. AM and VEGF mRNA were detected in all RCC specimens and cultured RCC cell lines analyzed by RT-PCR. There was a positive correlation between AM mRNA expression and MVC (r = 0.516, P = 0.0062), and between VEGF mRNA expression and MVC (r = 0.485, P = 0.0111). We also observed a positive correlation between AM mRNA expression and VEGF mRNA expression (r = 0.552, P = 0.0029). Hypoxia significantly induced AM and VEGF mRNA expression, although the increase of the AM mRNA level (10.6-26.7 fold) was markedly greater than that of the VEGF mRNA level (1.5-1.9 fold). CONCLUSION: These results suggest that hypoxia-induced AM plays a part in tumor angiogenesis in conjunction with VEGF and facilitates human RCC growth under hypoxic conditions. PMID- 12110093 TI - Factors associated with failure of extracorporeal shock-wave lithotripsy for ureteral stones using Dornier lithotripter U/50. AB - BACKGROUND: In the present series of 170 patients who underwent extracorporeal shock-wave lithotripsy (SWL) treatment for ureteral stones, the authors determine which patients with ureteral stones had an unsuccessful outcome. METHODS: The records of 170 patients with ureteral stones who were treated with SWL using the Dornier lithotriptor U/50 (EMSE 140) between January 1998 and December 1999 were retrospectively investigated. One hundred and thirty-one patients were treated with SWL alone (single session, n = 98; multiple session, n = 33) and 39 patients required auxiliary treatment due to failure of SWL (33 with transurethral ureterolithotripsy (TUL), one with open lithotomy, and five with residual fragments who were followed up). These two groups were compared using multivariate logistic regression analysis. RESULTS: Lower ureteral stones and stones more than 12 mm in diameter were associated with a poor outcome of SWL. There were no significant differences in age, gender, number of stones, JJ stent placement, and degree of ureteral obstruction due to the stone between the two groups. The odds ratios of lower ureteral stones and stones > or = 12 mm were 4.18 and 2.57, respectively. CONCLUSION: Patients with distal ureteral stones and/or stones more than 12 mm in diameter were difficult to treat successfully with SWL. Alternatives such as TUL should possibly be considered as a first-line therapy for these stones. PMID- 12110094 TI - Clinical efficacy and safety of sildenafil citrate (Viagra) in a multi-racial population in Singapore: A retrospective study of 1520 patients. AB - BACKGROUND: Sildenafil citrate (Viagra), a selective inhibitor of cGMP-specific phosphodiesterase type-5, has been used as an oral therapeutic drug for erectile dysfunction. The present paper is a clinical study of the success rate and side effects of the use of sildenafil in a multi-racial population in Singapore. METHODS: From April 1999 to May 2000, 1520 patients were given sildenafil citrate. Of these, 912 patients (mean age, 54.6 years; age range, 22-99 years) were followed up and evaluated for clinical efficacy and safety of the drug. The mean duration of erectile dysfunction (ED) and follow-up periods were 31.5 and 3.0 months, respectively. RESULTS: Satisfactory erections assessed by single global efficacy question (GEQ) occurred in 83% of patients, major side-effects in the form of flushing (3.48%), headache (1.97%), blurred vision (1.25%), giddiness (1.18%), warmth (1.11%) and others (4.92%) were recorded in 127 patients (13.9%). Racially, Chinese men with ED had higher efficacy (85.7%), compared to Indian men (74.2%) and Malay men (72.8%). With respect to comorbid profiles, an efficacy of 77.8% (n = 271), 83.9% (n = 292), 86.4% (n = 44) and 83.3% (n = 199) was recorded in diabetic, hypertensive, ischemic heart disease patients and in benign prostatic hyperplasia patients, respectively. Patients who smoked (n = 135) and drank alcohol (n = 118) showed an efficacy of 80%. Baseline hormonal profiles of luteinizing hormone, follicle stimulating hormone, testosterone and prolactin did not affect the success rates of sildenafil citrate. Many patients had earlier received other forms of treatment (medicated urethral suppository for erection (MUSE; 84.9%); vacuum devices (86.8%), traditional medicines (100%) and other oral medications (89.2%)), but this did not influence the success rate of sildenafil citrate. But patients previously treated with prostaglandin-E intracavernosal injections were less successful on sildenafil citrate (77.3%). In the total cohort, 50 mg sildenafil citrate was an effective dose in 49% of patients and 46.5% patients needed 100 mg sildenafil citrate, while 4.1% of the total cohort needed only 25 mg sildenafil citrate. CONCLUSION: Oral sildenafil citrate has been shown to be an effective, safe and well tolerated drug in Singaporean men with ED, as in men from other parts of the world. PMID- 12110092 TI - Urinary adrenomedullin levels are increased and correlated with plasma concentrations in patients with Behcet's syndrome. AB - BACKGROUND: The objective was to measure urinary adrenomedullin (AM) levels in patients with active or inactive Behcet's syndrome and compare them to levels in healthy control subjects. METHODS: Forty-five consecutive patients with Behcet's syndrome (20 men and 25 women with a mean age of 37.7 +/- 10.8 years) and 20 age- and sex-matched healthy hospital staff volunteers as control subjects (nine men and 11 women with a mean age of 36.2 +/- 10.4 years) were studied. Urinary and plasma AM concentrations were measured by high-performance liquid chromatography. We also investigated whether disease activity correlates with urinary and plasma AM levels. The Mann-Whitney U-test was used in statistical analysis and the values were expressed as mean +/- SD. RESULTS: Urinary excretion of AM (pmol per mg urinary creatinine) in patients with Behcet's syndrome (81.3 +/- 35.1) was significantly higher (P < 0.001) than in control subjects (31.2 +/- 16.1). Plasma AM levels (pmol/L) in patients with Behcet's syndrome and controls were 69.1 +/- 19.2 and 20.7 +/- 11.8, respectively; the difference was significant (P < 0.001). Although active Behcet's syndrome patients (n = 22) had higher urinary AM levels (92.1 +/- 41.1) compared to inactive (n = 23; 70.8 +/- 32.2), the difference was not significant (P > 0.05). Plasma AM levels in active Behcet's syndrome patients (77.5 +/- 21.2) were also higher than in inactive (61.6 +/- 17.3), but the difference was not significant (P > 0.05). CONCLUSION: Urinary AM levels were higher in Behcet's patients than in control subjects. Urinary AM levels were correlated with plasma AM levels. The results suggest that the higher AM levels found in the urine may be produced by the kidney as a result of the stimulation of inflammation during the course of Behcet's syndrome, or may come from plasma, as plasma AM levels were increased. However, the exact sites of AM synthesis by the kidney (e.g. glomeruli, blood vessels and/or tubular cells) could not be determined in this study. Further studies in this respect are necessary. PMID- 12110095 TI - Usefulness of prostate-specific antigen velocity in screening for prostate cancer. AB - BACKGROUND: The cut-off value of prostate-specific antigen velocity (PSAV) was investigated in relation to the initial prostate-specific antigen (PSA) value in subjects with initial values of 1.0-4.0 ng/mL, and the usefulness and limitations of PSAV as a screening test for prostate cancer were examined. METHODS: In this study, 4883 men who underwent mass screening for prostate cancer two or more times between 1987 and 1998 and had initial PSA levels of 1.0-4.0 ng/mL were investigated. The subjects ranged in age from 42 to 96 years (mean: 68.0 +/- 6.6 years). The cut-off value of PSAV was set at 0.1-1.5 ng/mL per year, and the sensitivity, specificity, efficiency and positive predictive value (PPV) of PSAV for detecting prostate cancer were determined according to the initial PSA value. A similar examination of the average PSAV was carried out in 2888 subjects with three or more visits for mass screening for prostate cancer. RESULTS: The diagnostic efficiency of PSAV was optimal with cut-off values of 0.3 and 0.75 ng/mL per year in those subjects with initial PSA levels of 1.0-1.9 and 2.0-4.0 ng/mL, respectively, but the PPV was low at 1.8% in subjects with initial PSA levels of 1.0-1.9 ng/mL. When the cutoff value of PSAV was set at 1.2 ng/mL per year in individuals with initial PSA levels of 1.0-1.9 ng/mL, the PPV increased to 7.3% and the sensitivity was 40%. The diagnostic efficiency of the average PSAV was optimal at the cut-off values of 0.2 and 0.4 ng/mL per year in subjects with initial PSA levels of 1.0-1.9 and 2.0-4.0 ng/mL, respectively, but the PPV was low at 2.2% in the subjects with initial PSA values of 1.0-1.9 ng/mL. When the cut-off value of PSAV was set at 0.75 ng/mL per year in individuals with initial PSA levels of 1.0-1.9 ng/mL, the PPV was 9.8% and the sensitivity was 46%. CONCLUSION: It is possible to improve the diagnostic accuracy of prostate cancer screening using the cut-off value of PSAV and average PSAV in subjects with initial PSA levels of 1.0-4.0 ng/mL. The cut-off values of PSAV should be set at 1.2 and 0.75 ng/mL per year in individuals with initial PSA levels of 1.0 1.9 and 2.0-4.0 ng/mL, respectively. The cut-off values of the average PSAV should be set at 0.75 and 0.4 ng/mL per year in individuals with initial PSA levels of 1.0-1.9 and 2.0-4.0 ng/mL, respectively. PMID- 12110097 TI - Comparison of surgical stress between laparoscopy and open surgery in the field of urology by measurement of humoral mediators. AB - BACKGROUND: Recently, laparoscopic procedures have become more common in urological surgery, and are suggested to be less stressful compared with open surgery; however, little data on objective evaluation of surgical stress after laparoscopic surgery are available. The objective of this study was to compare surgical stress between laparoscopic and open surgery in the field of urology by measurement of humoral mediators. METHODS: Blood samples were obtained from 25 patients who underwent laparoscopic surgery (laparoscopic radical prostatectomy, 10; retroperitoneoscopic nephrectomy or nephroureterectomy, seven; laparoscopic adrenalectomy, five; and hand-assisted laparoscopic radical nephrectomy, three) and 10 who underwent open surgery (retropubic radical prostatectomy, four; radical cystectomy, three; and radical nephrectomy, three), 48 h before surgery, during surgery, and 24, 48 and 96 h after surgery. Serum levels of interleukin (IL)-6, IL-10 and granulocytic elastase in these samples were measured, and the results were analyzed with respect to several clinical factors. RESULTS: In both groups, IL-6 and granulocytic elastase levels increased during surgery and reached maximum levels 24 h after surgery; the increase in granulocytic elastase tended to be prolonged compared with that of IL-6. Changes in IL-10 levels in the open surgery group were similar to those of IL-6 and granulocytic elastase levels, whereas IL-10 levels in the laparoscopic surgery group reached a maximum level during surgery and then decreased to the same level as at 48 h before surgery, on the first postoperative day. The maximum levels of these three mediators in the laparoscopic surgery group were significantly lower than those in the open surgery group. IL-6 was closely associated with the interval of fasting and duration of hospitalization after surgery. CONCLUSION: Although the present study was based on retrospective and non-randomized analysis, the findings suggest that serum levels of IL-6, IL-10 and granulocytic elastase are useful as objective markers of surgical stress, and that laparoscopic surgery is markedly less stressful than open surgery. PMID- 12110098 TI - Annual PSA tests are not necessary for men with a PSA level below 2 ng/mL: findings of the Imari prostate cancer screening program. AB - BACKGROUND: Annual changes in prostate specific antigen (PSA) levels detected by the Imari prostate cancer screening program were evaluated to establish a more efficient and cost-saving screening system, especially for men with low PSA levels. METHODS: Prostate specific antigen-based annual mass screenings for prostate cancer were conducted for men aged 60-69 in the Imari district, Saga, Japan. Between 1992 and 2000, 1822 men had their PSA levels tested. A total of 4661 PSA tests were conducted. Changes in PSA levels over the following 1 to 5 years were analyzed in men with PSA levels of 3 ng/mL or less, a range in which the detection rate of prostate cancer would seem to be negligibly low. RESULTS: The overall detection rate of prostate cancer between 1992 and 2000 was 0.73%. The detection rate in men with a PSA level between 3.1 and 3.9 ng/mL, and between 4 and 9.9 ng/mL was 1.6% and 8.3%, respectively. Of 4661 determinations of PSA, 2553 (54.8%) were found to be < or = 1 ng/mL, 1273 (27.3%) were between 1.1 and 2 ng/mL, and 401 (8.6%) were between 2.1 and 3 ng/mL. Four hundred and thirty-four men (9.3%) had PSA levels > or = 3.1 ng/mL, with possible indications for prostate biopsy. Of the men tested, 1.4% with an initial PSA level of < or = 2 ng/mL and 22.3% with an initial level between 2.1 and 3 ng/mL had a PSA level of > or = 3.1 ng/mL after 1 year. Almost the same rate of PSA increase was observed between the two PSA tests conducted at 2 to 5-year intervals. Of the men tested, 2.2% with an initial PSA level of < or = 2 ng/mL, and 21.9% with an initial level between 2.1 and 3 ng/mL, had a level of > or = 3.1 ng/mL after 5 years. CONCLUSION: Levels of PSA in men with an initial level below 2 ng/mL remained stable for up to 5 years. Levels of PSA in 97.8- 98.8% of men remained below 3 ng/mL after 1 to 5 years. In contrast, 18-35.3% of men with an initial PSA level between 2.1 and 3 ng/mL showed PSA progression to 3.1 ng/mL or more within 5 years. Our present data suggest that annual PSA testing is not necessary for men with a PSA level below 2 ng/mL. Prostate specific antigen testing could therefore be conducted at longer intervals in such individuals. PMID- 12110099 TI - Dendritic cell immunotherapy for patients with metastatic renal cell carcinoma: University of Tokyo experience. AB - BACKGROUND: Dendritic cells (DC) are the most potent antigen-presenting cells and induce host antitumor immunity through the T-cell response. A clinical study of immunotherapy using cultured DC loaded with tumor antigen, for patients with metastatic renal cell carcinoma (RCC) was performed. METHODS: Dendritic cells were generated by culturing monocytes from peripheral blood for 7 days in the presence of granulocyte-macrophage colony-stimulating factor and interleukin-4. On day 6 the DC were pulsed with lysate from autologous tumor as the antigen and with keyhole limpet hemocyanin (KLH) as immunomodulator. The patients were given four doses of lysate-pulsed DC by intradermal injection with a 2-week interval between doses. Clinical effect and immune response were, respectively, evaluated by radiological examination and delayed-type hypersensitivity (DTH) test. RESULTS: Three patients were enrolled and the immunotherapy was well tolerated without significant toxicity. The vaccination induced a positive DTH reaction to tumor lysate in two patients and to KLH in all patients. Clinical responses consisted of one case of no change and two cases of progression of disease. However, we did not see a significant reduction of tumor volume in any case. CONCLUSION: Dendritic cell vaccination can safely induce an immunological response against RCC. Further trials are needed to fully evaluate its efficacy. PMID- 12110100 TI - Stable maintenance of 5alpha-reductase activity in long-term subcultures of fibroblasts derived from the foreskin. AB - BACKGROUND: There is up to a 50-fold variation in control subjects in current assays of 5alpha-reductase activity which makes interpretation difficult. It was therefore attempted in this study to establish an assay method which produced stable 5alpha-reductase activity in long-term subcultured foreskin fibroblasts. METHODS: Foreskin fibroblasts were obtained from three boys with phimosis (control subjects), three patients with Reifenstein syndrome and one patient with 5alpha-reductase deficiency (due to mutation L113P in exon 2 of the SRD5A2 gene). To maximize the number of cells in the DNA synthesis phase, cells were subcultured consistently to approximately 70% confluency. Thawed cells, frozen after the third subculture, were incubated for 24 h with [1beta,2beta-3H] testosterone. 5alpha-Reductase activity was expressed as the sum of formed [3H] 5alpha-reduced metabolites (separated by thin-layer chromatography). RESULTS: The full range of 5alpha-reductase activity in controls and patients with Reifenstein syndrome was 3.44-15.59 pmol/h per mg protein: a 4.53-fold variation. The activity in the patient with 5alpha- reductase deficiency was 0.52 pmol/h per mg protein. CONCLUSION: By the cell culture methods used in this study, which aimed to increase the number of cells in the DNA synthesis phase, foreskin fibroblasts maintained a considerably stable level of 5alpha-reductase activity during long term subculture. Therefore, this assay method can be used for differential diagnosis of 5alpha-reductase deficiency from other relevant entities. PMID- 12110101 TI - Sarcomatoid carcinoma and carcinosarcoma of the urinary bladder. AB - Two cases, sarcomatoid carcinoma and carcinosarcoma, of the urinary bladder are reported. A 68-year-old man with sarcomatoid carcinoma underwent total cystectomy and was alive and had had no recurrence after 21 months. A 78-year-old woman with carcinosarcoma underwent total cystectomy, but she died from increasing multiple lung metastases 4 months after surgery. The histopathological characteristics of both neoplasms are reported and discussed. PMID- 12110102 TI - Failure to maintain the suppressed level of serum testosterone during luteinizing hormone-releasing hormone agonist therapy in a patient with prostate cancer. AB - A 75-year-old man with metastatic prostate cancer had been treated with goserelin acetate, and prostate specific antigen (PSA) had decreased, but 11/2 years after beginning the treatment of goserelin acetate, PSA was markedly elevated and serum testosterone was at normal level. After castration the serum testosterone was at castrate level and PSA decreased. In the present case, leuprorelin acetate 1 month depot suppressed the luteinizing hormone level in 1 month, even after the patient underwent castration. PMID- 12110103 TI - Crown fractures in the permanent dentition: pulpal and restorative considerations. AB - Crown fractures account for the highest percentage of all traumatic injuries in the permanent dentition. This review paper will discuss the different types of crown fracture, from the uncomplicated to complicated, including crown-root fractures. It will focus on two different aspects: the pulp, with an attempt to correlate epidemiological, experimental, histopathological and clinical studies, so that the clinician can better understand the underlying processes accounting for success or failure to maintain pulp vitality. Also, we will consider the restoration: knowledge about bonding to dentin and new material is evolving extremely quickly making it difficult for the clinician to keep up with the developments. If handled properly, prognosis of the pulp, after traumatic crown fracture, is good. Prognosis of the restoration has also improved considerably over the last few years, and it appears that this trend will continue in the future. PMID- 12110104 TI - Effect of treatment delay upon pulp and periodontal healing of traumatic dental injuries -- a review article. AB - Based on an analysis of the literature concerning parameters influencing the prognosis of traumatic dental injuries, few studies were found to have examined possible relationships between treatment delay and pulpal and periodontal ligament healing complications. It has been commonly accepted that all injuries should be treated on an emergency basis, for the comfort of the patient and also to reduce wound healing complications. For practical and especially economic reasons, various approaches can be selected to fulfill such a demand, such as acute treatment (i.e. within a few hours), subacute (i.e. within the first 24 h), and delayed (i.e. after the first 24 h). In this survey the consequences of treatment delay on pulpal and periodontal healing have been analyzed for the various dental trauma groups. Applying such a treatment approach to the various types of injuries, the following treatment guidelines can be recommended, based on our present rather limited knowledge of the effect of treatment delay upon wound healing. Crown and crown/root fractures: Subacute or delayed approach. Root fractures: Acute or subacute approach. Alveolar fractures: Acute approach (evidence however questionable). Concussion and subluxation: Subacute approach. Extrusion and lateral luxation: Acute or subacute approach (evidence however questionable). Intrusion: Subacute approach (evidence however questionable). Avulsion: If the tooth is not replanted at the time of injury, acute approach; otherwise subacute. Primary tooth injury: Subacute approach, unless the primary tooth is displaced into the follicle of the permanent tooth or occlusal problems are present; in the latter instances, an acute approach should be chosen. These treatment guidelines are based on very limited evidence from the literature and should be revised as soon as more evidence about the effect of treatment delay becomes available. PMID- 12110105 TI - Long-term calcium hydroxide as a root canal dressing may increase risk of root fracture. AB - It has been proposed (Cvek 1992) that immature teeth are weakened by filling of the root canals with calcium hydroxide dressing and gutta-percha. The aim of the present study was to test the hypothesis that dentin in contact with calcium hydroxide would show a reduction in fracture strength after a certain period of time. Immature mandibular incisors from sheep were extracted and divided into two experimental groups. Group 1: the pulps were extirpated via the apical foramen. The root canals were then filled with calcium hydroxide (Calasept) and sealed with IRM(R) cement, and the teeth were then stored in saline at room temperature for 0.5, 1, 2, 3, 6, 9, or 12 months. Group 2: the pulps were extirpated and the root canals were filled with saline and sealed with IRM(R) cement. The teeth were then stored in saline for 2 months. Intact teeth served as controls and were tested immediately after extraction. All teeth were tested for fracture strength in an Instron testing machine at the indicated observation periods. The results showed a markedly decrease in fracture strength with increasing storage time for group 1 (calcium hydroxide dressing). The results indicate that the fracture strength of calcium hydroxide-filled immature teeth will be halved in about a year due to the root filling. The finding may explain the frequent reported fractures of immature teeth filled with calcium hydroxide for extended periods. PMID- 12110106 TI - Internal root resorption studied by radiography, stereomicroscope, scanning electron microscope and computerized 3D reconstructive method. AB - AIM AND METHODOLOGY: Two cases of internal tooth resorption were examined. A mandibular premolar and a mandibular canine were studied after they were extracted using radiographs, a stereomicroscope (SM) and a scanning electron microscope (SEM). Lastly, 3D images of the sectioned teeth were obtained(3D). RESULTS: Radiographically, internal root resorption was shown as a uniform radiolucency. By SM examination, an extensive destruction of dentin was seen, while, by SEM examination, a disappearance of dentinal tubules was clear. The 3D reconstructive method revealed a circumscribed, oval-shaped defect that did not perforate the cemental layer. CONCLUSIONS: Internal root resorption is a rare remodeling process that can be studied using different experimental methods. PMID- 12110107 TI - Survey on the use of complementary and alternative medicine among patients with headache syndromes. PMID- 12110108 TI - Relationship between sleep apnoea syndrome, snoring and headaches. AB - There is still a controversy regarding the relationship between sleep apnoea syndrome and headaches, especially morning headaches. Our objectives were: (i) to compare the prevalence and the clinical data of headaches in sleep apnoea syndrome (SAS) and control (snorers) groups defined by polysomnographic recording; (ii) to analyse the clinical improvement of headaches with appropriate treatment; and (iii) to correlate headaches with mood disorders, and nocturnal respiratory and architectural sleep parameters in order to understand the underlying pathophysiological mechanisms. This is a prospective study of 324 consecutive patients referred to our sleep centre for snoring. Of these, 312 patients who underwent sleep polysomnography were finally included. Patients and controls were interviewed about their medical past, headache history and clinical characteristics, their daytime sleepiness (Epworth's sleepiness scale) and their mood disorders (Zerssen's scale). Follow-up of patients with headaches (SAS and control groups), treated or not, was also assessed. According to our definition of SAS, patients were dissociated in SAS (n=164) and snorers (n=148). Fifty-three SAS patients had headaches, of whom 58.5% (n=30) suffered from morning headaches. However, there was no statistical difference between the two groups concerning the prevalence and the clinical characteristics of headaches. In addition, headaches and morning headaches were not correlated with nocturnal respiratory and architectural sleep parameters, nor with excessive daytime sleepiness, but were strongly correlated with mood disorders. In 36 SAS patients, headaches improved under treatment, but this was not statistically different from what was found among untreated snorers. Headaches and morning headaches are common in patients with SAS but may be considered as a non-specific symptom. The underlying mechanisms are not fully elucidated but depression could play an important role. Despite this absence of specificity, the treatment of SAS, especially nasal continuous positive airway pressure, leads to an improvement in headaches in several cases. PMID- 12110109 TI - Muscle tenderness in pericranial and neck-shoulder region in children with headache. A controlled study. AB - Increased pericranial muscle tenderness is connected with tension-type headache in adults. In children, the importance of muscle tenderness in the pericranial or neck-shoulder region in the pathogenesis of different types of headache is unknown. The present study evaluated muscle tenderness in the pericranial and neck-shoulder region in children with migraine, those with tension-type headache and those without headache. An unselected population-based questionnaire study concerning headache was carried out in 1135 Finnish schoolchildren aged 12 years. Of them, 183 children were randomly selected for a face-to-face interview and a clinical examination. Muscle tenderness was recorded by manual palpation and dolorimeter. Children with migraine had increased overall tenderness, recorded by manual palpation, compared with those without headache. They also self-reported tenderness in the neck-shoulder region during daily activities more often than the children of the other groups. Muscle tenderness was not associated with paediatric tension-type headache. The mean pressure pain thresholds did not differ among the three groups. However, a negative correlation between the total tenderness score and the dolorimeter score was found in each group. In conclusion, children with migraine had increased muscle tenderness at palpation of the pericranial and neck-shoulder muscles and they also reported pain symptoms in the neck-shoulder region most frequently. Instead, increased pericranial and neck-shoulder muscle tenderness was not associated with tension-type headache in children. PMID- 12110110 TI - Intravenous magnesium sulphate in the acute treatment of migraine without aura and migraine with aura. A randomized, double-blind, placebo-controlled study. AB - Magnesium sulphate has been used in the acute treatment of migraines; some studies found it to be a highly effective medication in the acute control of migraine pain and associated symptoms. This randomized, double-blind, placebo controlled study assesses the effect of magnesium sulphate on the pain and associated symptoms in patients with migraine without aura and migraine with aura. Sixty patients in each group were assigned at random to receive magnesium sulphate, 1000 mg intravenously, or 0.9% physiological saline, 10 ml. We used seven parameters of analgesic evaluation and an analogue scale to assess nausea, photophobia and phonophobia. In the migraine without aura group there was no statistically significant difference in the patients who received magnesium sulphate vs. placebo in pain relief. The analgesic therapeutic gain was 17% and number needed to treat was 5.98 at 1 h. There was also no statistical difference in relief of nausea. We did observe a significant lower intensity of photophobia and phonophobia in patients who received magnesium sulphate. In the migraine with aura group patients receiving magnesium sulphate presented a statistically significant improvement of pain and of all associated symptoms compared with controls. The analgesic therapeutic gain was 36.7% at 1 h. A smaller number of patients continued to have aura in the magnesium sulphate group compared with placebo 1 h after the administration of medication. Our data support the idea that magnesium sulphate can be used for the treatment of all symptoms in migraine with aura, or as an adjuvant therapy for associated symptoms in patients with migraine without aura. PMID- 12110111 TI - Sudden onset headache: a prospective study of features, incidence and causes. AB - Sudden onset headache is a common condition that sometimes indicates a life threatening subarachnoid haemorrhage (SAH) but is mostly harmless. We have performed a prospective study of 137 consecutive patients with this kind of headache (thunderclap headache=TCH). The examination included a CT scan, CSF examination and follow-up of patients with no SAH during the period between 2 days and 12 months after the headache attack. The incidence was 43 per 100 000 inhabitants >18 years of age per year; 11.3% of the patients with TCH had SAH. Findings in other patients indicated cerebral infarction (five), intracerebral haematoma (three), aseptic meningitis (four), cerebral oedema (one) and sinus thrombosis (one). Thus no specific finding indicating the underlying cause of the TCH attack was found in the majority of the patients. A slightly increased prevalence of migraine was found in the non-SAH patients (28%). The attacks occurred in 11 cases (8%) during sexual activity and two of these had an SAH. Nausea, neck stiffness, occipital location and impaired consciousness were significantly more frequent with SAH but did not occur in all cases. Location in the temporal region and pressing headache quality were the only features that were more common in non-SAH patients. Recurrent attacks of TCH occurred in 24% of the non-SAH patients. No SAH occurred later in this group, nor in any of the other patients. It was concluded that attacks caused by a SAH cannot be distinguished from non-SAH attacks on clinical grounds. It is important that patients with their first TCH attack are investigated with CT and CSF examination to exclude SAH, meningitis or cerebral infarction. The results from this and previous studies indicate that it is not necessary to perform angiography in patients with a TCH attack, provided that no symptoms or signs indicate a possible brain lesion and a CT scan and CSF examination have not indicated SAH. PMID- 12110112 TI - An epidemiological survey of hemiplegic migraine. AB - The objective of the present study was to use systematic nation-wide case-finding methods to establish the prevalence and sex ratio of hemiplegic migraine (HM) in the entire Danish population of 5.2 million inhabitants. Affected patients were identified from three different recruitment sources: the National Patient Register, case records from private practising neurologists and advertisements. Based on the observed number of affected patients from each case-finding method, it was attempted to estimate the total number of affected patients by means of the statistical method known as capture-recapture. Two hundred and ninety-one affected patients were identified; 147 were familial HM from 44 different families, 105 were sporadic HM and 39 were unclassifiable HM. The HM sex ratio (M:F) was 1:3. Based on the identified number of affected patients the prevalence of HM at the end of 1999 was estimated to be 0.01% in Denmark, where the familial and sporadic form were equally frequent. PMID- 12110113 TI - Zolmitriptan is effective and well tolerated in Japanese patients with migraine: a dose-response study. AB - This phase II study investigated the efficacy, tolerability and dose-response relationship of oral zolmitriptan in the treatment of a single migraine attack in Japanese patients. A bridging analysis then assessed the validity of extrapolating western clinical data to these Japanese patients. In this multicentre, randomized, double-blind, placebo-controlled study, patients received a single dose of placebo or zolmitriptan 1, 2.5 or 5 mg. The primary endpoints were 2-h headache response and the tolerability of zolmitriptan. A statistically significant dose-response relationship was observed for the 2-h headache response (P=0.003). The 2.5 mg group had significantly greater 2-h headache response than the placebo group (P=0.032). The adverse event profile was similar to that reported in western patients, and no adverse events unique to the Japanese population were observed. The bridging analysis report confirmed similar efficacy and tolerability of zolmitriptan in Japanese and western populations. In the Japanese patients, the estimated response rates were 34.3%, 45.2%, 57.7% and 66.2% for placebo, and zolmitriptan 1, 2.5 and 5 mg, respectively, while in the western population the corresponding rates were 39.9%, 49.6%, 61.2% and 71.7%. Zolmitriptan is effective and well tolerated in the acute treatment of migraine in Japanese patients. The optimal dose was 2.5 mg, although the 5 mg dose may provide further benefit for some patients. The bridging analysis supports extrapolation of data from western to Japanese patients. PMID- 12110114 TI - 5-Hydroxytryptamine(1B/1D) and 5-hydroxytryptamine1F receptors inhibit capsaicin induced c-fos immunoreactivity within mouse trigeminal nucleus caudalis. AB - In order to investigate the c-fos response within the trigeminal nucleus caudalis (Sp5C) after noxious meningeal stimulation, capsaicin (0.25, 0.5, 1 and 5 nmol) was administered intracisternally in urethane (1 g/kg) and alpha-chloralose (20 mg/kg) anaesthetized male mice. Capsaicin induced a robust and dose-dependent c fos-like immunoreactivity (c-fos LI) within Sp5C. C-fos LI was observed within laminae I and II of the entire brain stem from the area postrema to C2 level, being maximum at the decussatio pyramidum level. The area postrema, solitary tract, medullary and lateral reticular nuclei were also labelled. The 5 hydroxytryptamine(1B/1D/1F) receptor agonist sumatriptan (0.01, 0.1, 1 and 10 mg/kg), administered intraperitoneally 15 min before capsaicin stimulation (1 nmol), decreased the c-fos response within Sp5C, but not within solitary tract. The novel specific 5-hydroxytryptamine1F agonist LY 344864 (0.1 and 1 mg/kg, i.p.) significantly decreased the c-fos LI within the Sp5C as well. These findings suggest that intracisternally administered capsaicin activates the trigeminovascular system and that the pain neurotransmission can be modulated by 5-hydroxytryptamine(1B/1D/1F) receptors in mice. Thus, the availability of this model in mice, taken together with the possibility of altering the expression of specific genes in this species, may help to investigate further the importance of distinct proteins in the neurotransmission of cephalic pain. PMID- 12110115 TI - Survey on the use of complementary and alternative medicine among patients with headache syndromes. AB - The objective was to determine headache patients' knowledge, prevalence of use and perceived effectiveness of complementary and alternative medicine. Seventy three patients with headache syndromes attending a head and neck pain clinic were interviewed using a standardized questionnaire. Alternative medical therapies were used by 85% of surveyed patients for the relief of their head pain. In 60%, the therapies were perceived to have a benefit. Almost 100% of the patients were familiar with one or more of the presented alternative treatments. Eighty-eight per cent perceived at least one of the complementary treatments to be an effective remedy for headache pain. Exposure to and interest in alternative treatments are common among patients with headache syndromes, despite the lack of scientific evidence of benefit and assessments of risks for many of the treatments. Neurologists and general physicians should be aware of the increasing role of alternative medicine in the healthcare system. There is still an urgent need for objective, integrative and critical research with regard to complementary and alternative medicine. PMID- 12110116 TI - Determinants of tension-type headache in children. AB - The objective of this study was to study the prevalence, characteristics and predisposing factors of tension-type headache in children. An unselected population-based questionnaire study was carried out in 1409 Finnish schoolchildren aged 12 years. Of them, 1135 (81%) returned an acceptably completed questionnaire. The prevalence of episodic tension-type headache in children was 12% (138 of 1135). Children with episodic tension-type headache also often reported characteristics of pain typical for migraine. Children with frequent and persistent episodic tension-type headache reported stabbing and severe occipital pain, phonophobia and abdominal pain significantly more often than children with infrequent episodic tension-type headache. Neck-shoulder symptoms, symptoms of depression and oromandibular dysfunction were each independently associated with episodic tension-type headache. The father's occupation of a lower-level white-collar worker put the child at a four-fold risk for episodic tension-type headache. We conclude that episodic tension-type headache is as common as migraine in children. It can be associated with depression, oromandibular dysfunction and muscular stress. Especially children with frequent and persistent episodic tension-type headache report characteristics of pain typical for migraine. PMID- 12110117 TI - Strictly unilateral headache reminiscent of hemicrania continua resistant to indomethacin but responsive to gabapentin. PMID- 12110119 TI - Perspectives for TNF-alpha-targeting therapies. AB - Rheumatoid arthritis (RA) is the most common chronic autoimmunopathy, clinically leading to joint destruction as a consequence of the chronic inflammatory processes. The pathogenesis of this disabling disease is not well understood, but molecular events leading to tissue inflammation with cartilage and bone destruction are now better defined. Therapy with slow-acting, disease-modifying antirheumatic drugs (DMARDs), such as low-dose methotrexate, which is generally accepted as a standard, leads to a significant amelioration of symptoms but does not stop joint destruction. Due to these disappointing treatment options and the identification of certain inflammatory mediators as therapeutic targets, novel therapeutic agents such as monoclonal antibodies, cytokine-receptor/human immunoglobulin constructs or recombinant human proteins have been tested in RA with some success. Clinical trials testing anti-TNF-alpha agents, alone or in combination with methotrexate, have convincingly shown the feasibility and efficacy of these novel approaches to the therapy of RA. A clinical trial testing combination therapy with chimeric (mouse/human) anti-TNF-alpha monoclonal antibody infliximab and methotrexate showed, for the first time in any RA trial, that there was no median radiological progression in the groups given infliximab plus methotrexate over a 12-month observation period. Similar encouraging results might arise from trials employing other TNF-alpha-directed agents, such as the fully human monoclonal antibody D2E7, the p75 TNF-alpha-receptor/Ig construct, etanercept, or others, as discussed in this review. Combination partners other than methotrexate will be established as suitable cotreatment along with anti-TNF alpha biologicals. Forthcoming new indications for TNF-alpha-targeted therapies are discussed. PMID- 12110120 TI - Therapy of systemic lupus erythematosus: a look into the future. AB - The prognosis for patients with systemic lupus erythematosus has greatly improved over the past two decades. However, therapies that are more effective and that have fewer sequelae are needed to rescue patients from organ failure and further reduce mortality. Research under way, including that into induction of tolerance to self-antigens, prevention of the consequences of pathogenic autoantibody production, interference with the cytokine network and signal transduction, the identification and treatment of any infectious triggers, and stem cell therapy, offers hope of improved remedies or even of cure. Given the fact that a number of biological therapies for rheumatologic disease are already in use or are in the development stage, such progress may come soon. PMID- 12110121 TI - Genes and environment in arthritis: can RA be prevented? AB - Understanding of how interactions between genes and environment contribute to the development of arthritis is a central issue in understanding the etiology of rheumatoid arthritis (RA), as well as for eventual subsequent efforts to prevent the disease. In this paper, we review current published data on genes and environment in RA as well as in certain induced animal models of disease, mainly those in which adjuvants only or adjuvants plus organ-specific autoantigens are used to induce arthritis. We refer to some new data on environmental and genetic factors of importance for RA generated from a large case-control study in Sweden (1200 patients, 1200 matched controls). We found an increased risk of seropositive but not of seronegative RA in smokers, and there are indications that this effect may be due to a gene-environment interaction involving MHC class II genes. We also found an increased risk of RA in individuals heavily exposed to mineral oils. This was of particular interest because mineral oils are strong inducers of arthritis in certain rodent strains and because polymorphisms in human genetic regions syntenic with genes predisposing for oil-induced arthritis in rats have now been shown to associate with RA in humans. Taken together, our data support the notion that concepts and data on gene-environment interactions in arthritis can now be taken from induced animal models of arthritis to generate new etiological hypotheses for RA. PMID- 12110118 TI - Immunotherapy of type 1 diabetes: lessons for other autoimmune diseases. AB - The nonobese diabetic (NOD) mouse is a well-recognised animal model of spontaneous autoimmune insulin-dependent diabetes mellitus. The disease is T-cell mediated, involving both CD4 and CD8 cells. Its progress is controlled by a variety of regulatory T cells. An unprecedented number of immunological treatments have been assessed in this mouse strain. This chapter systematically reviews most of these therapeutic manoeuvres, discussing them in the context of their significance with regard to the underlying mechanisms and the potential clinical applications. The contrast between the surprisingly high rate of success found for a multitude of treatments (more than 160) administered early in the natural history of the disease and the few treatments active at a late stage is discussed in depth. Most of the concepts and strategies derived from this model apply to other autoimmune diseases, for which no such diversified data are available. PMID- 12110122 TI - Matrix metalloproteinases in arthritic disease. AB - The role of matrix metalloproteinases in the degradative events invoked in the cartilage and bone of arthritic joints has long been appreciated and attempts at the development of proteinase inhibitors as potential therapeutic agents have been made. However, the spectrum of these enzymes orchestrating connective tissue turnover and general biology is much larger than anticipated. Biochemical studies of the individual members of the matrix metalloproteinase family are now underway, ultimately leading to a more detailed understanding of the function of their domain structures and to defining their specific role in cellular systems and the way that they are regulated. Coupled with a more comprehensive and detailed study of proteinase expression in different cells of joint tissues during the progress of arthritic diseases, it will be possible for the future development and application of highly specific proteinase inhibitors to be directed at specific key cellular events. PMID- 12110124 TI - Tissue engineering: chondrocytes and cartilage. AB - Tissue engineering offers new strategies for developing treatments for the repair and regeneration of damaged and diseased tissues. These treatments, using living cells, will exploit new developments in understanding the principles in cell biology that control and direct cell function. Arthritic diseases that affect so many people and have a major impact on the quality of life provide an important target for tissue engineering. Initial approaches are in cartilage repair; in our own programme we are elucidating the signals required by chondrocytes to promote new matrix assembly. These principles will extend to other tissues of the musculoskeletal system, including the repair of bone, ligament and tendon. PMID- 12110123 TI - Engineering of tissue inhibitor of metalloproteinases mutants as potential therapeutics. AB - Matrix metalloproteinases (MMPs) play a central role in many biological processes such as development, morphogenesis and wound healing, but their unbalanced activities are implicated in numerous disease processes such as arthritis, cancer metastasis, atherosclerosis, nephritis and fibrosis. One of the key mechanisms to control MMP activities is inhibition by endogenous inhibitors called tissue inhibitors of metalloproteinases (TIMPs). This review highlights the structures and inhibition mechanism of TIMPs, the biological activities of TIMPs, the unique properties of TIMP-3, and the altered specificity towards MMPs achieved by mutagenesis. A potential therapeutic use of TIMP variants is discussed. PMID- 12110125 TI - Insights into integrin-ligand binding and activation from the first crystal structure. AB - Integrin receptors transduce bidirectional signals between extracellular adhesion molecules and intracellular cytoskeletal and signalling molecules. The structural basis of integrin signalling is unknown, but the recent publication of the first crystal structure of the extracellular domain of integrin alphaVbeta3 has provided a number of insights. In this review, previous structure-function analyses of integrins that have employed biochemical and molecular biological approaches are placed in the context of the crystal structure, and novel routes to the development of integrin antagonists are discussed. PMID- 12110126 TI - Angiogenesis in rheumatoid arthritis. AB - The expansion of the synovial lining of joints in rheumatoid arthritis (RA) and the subsequent invasion by the pannus of underlying cartilage and bone necessitate an increase in the vascular supply to the synovium, to cope with the increased requirement for oxygen and nutrients. The formation of new blood vessels - termed 'angiogenesis' - is now recognised as a key event in the formation and maintenance of the pannus in RA. This pannus is highly vascularised, suggesting that targeting blood vessels in RA may be an effective future therapeutic strategy. Disruption of the formation of new blood vessels would not only prevent delivery of nutrients to the inflammatory site, but could also lead to vessel regression and possibly reversal of disease. Although many proangiogenic factors are expressed in the synovium in RA, the potent proangiogenic cytokine vascular endothelial growth factor (VEGF) has been shown to a have a central involvement in the angiogenic process in RA. The additional activity of VEGF as a vascular permeability factor may also increase oedema and hence joint swelling in RA. Several studies have shown that targeting angiogenesis in animal models of arthritis ameliorates disease. Our own study showed that inhibition of VEGF activity in murine collagen-induced arthritis, using a soluble VEGF receptor, reduced disease severity, paw swelling, and joint destruction. Although no clinical trials of anti-angiogenic therapy in RA have been reported to date, the blockade of angiogenesis - and especially of VEGF - appears to be a promising avenue for the future treatment of RA. PMID- 12110127 TI - Interactions between leukocytes and endothelial cells in gout: lessons from a self-limiting inflammatory response. AB - Interactions with endothelium are necessary for leukocytes to pass from the blood into extravascular tissues, and such interactions are facilitated in inflammation by the coordinated expression of endothelial adhesion molecules and chemoattractants. Although the general mechanisms and intracellular pathways of endothelial activation are now fairly well characterised in vitro, relatively little detailed information exists on how endothelial activation changes during the course of inflammatory responses and how such change influences the amount of leukocyte recruitment and the types of leukocytes recruited. Having developed a radiolabelled-antibody-uptake technique for quantifying the expression of endothelial adhesion molecules in relation to leukocyte trafficking, we have analysed the acute, self-limiting inflammatory response to injection of monosodium urate (MSU) crystals. Our studies have supported the view that endothelial activation is closely paralleled by leukocyte recruitment at the onset of the response and have highlighted separate vascular and extravascular stages of downregulation. More recent studies addressing the extravascular contribution to downregulation point to an important role for monocyte-macrophage differentiation in limiting further endothelial activation as a consequence of phagocytosis of MSU crystals. PMID- 12110128 TI - VEGF and imaging of vessels in rheumatoid arthritis. AB - Angiogenesis is a prominent feature of rheumatoid synovitis. Formation of new blood vessels permits a supply of nutrients and oxygen to the augmented inflammatory cell mass and so contributes to perpetuation of joint disease. Vascular endothelial growth factor (VEGF) is a potent endothelial cell-specific growth factor that is upregulated by proinflammatory cytokines and by hypoxia. Serum VEGF concentrations are elevated in rheumatoid arthritis (RA) and correlate with disease activity. Furthermore, serum VEGF measured at first presentation in RA is highly significantly correlated with radiographic progression of disease over the subsequent year. Power Doppler ultrasonography is a sensitive method for demonstrating the presence of blood flow in small vessels and there is a very close relation between the presence or absence of vascular flow signal on power Doppler imaging and the rate of early synovial enhancement on dynamic gadolinium enhanced magnetic resonance imaging (MRI) of joints with RA. Images obtained by both dynamic enhanced MRI and power Doppler ultrasonography correlate with vascularity of synovial tissue as assessed histologically. In early RA, there is a striking association between joint erosions assessed on high-resolution ultrasonography and vascular signal in power Doppler mode. Collectively, these findings implicate vascular pannus in the erosive phase of disease and strongly suggest that proangiogenic molecules such as VEGF are targets for novel therapies in RA. Animal model data supports this concept. It seems likely that serological and imaging measures of vascularity in RA will become useful tools in the assessment of disease activity and response to therapy. PMID- 12110129 TI - Endothelial activation: intracellular signaling pathways. AB - Tumor necrosis factor (TNF) is the prototypic proinflammatory cytokine and endothelial cells are the principal cellular targets of its actions. Here I review the responses of endothelial cells to TNF, with emphasis on the induction of endothelial leukocyte adhesion molecules. I focus on the biochemistry and cell biology of signal transduction in TNF-treated endothelial cells that lead to the expression of adhesion molecules. PMID- 12110131 TI - The instructive role of dendritic cells on T-cell responses. AB - Immune responses are initiated in the T-cell areas of secondary lymphoid organs where naive T lymphocytes encounter dendritic cells (DCs) that present antigens taken up in peripheral tissues. DCs represent the interface between the universe of foreign and tissue-specific antigens and T lymphocytes, and they are the key players in the regulation of cell-mediated immunity. We discuss how the nature of the DC maturation stimuli and the density and quality of DCs present in the T cell areas of secondary lymphoid organs determine the magnitude and class of the T-cell response. PMID- 12110130 TI - The immunological synapse. AB - T-cell activation requires interaction of T-cell antigen receptors with proteins of the major histocompatibility complex (antigen). This interaction takes place in a specialized cell-cell junction referred to as an immunological synapse. The immunological synapse contains at least two functional domains: a central cluster of engaged antigen receptors and a surrounding ring of adhesion molecules. The segregation of the T-cell antigen receptor (TCR) and adhesion molecules is based on size, with the TCR interaction spanning 15 nm and the lymphocyte-function associated antigen-1 (LFA-1) interaction spanning 30-40 nm between the two cells. Therefore, the synapse is not an empty gap, but a space populated by both adhesion and signaling molecules. This chapter considers four aspects of the immunological synapse: the role of migration and stop signals, the role of the cytoskeleton, the role of self-antigenic complexes, and the role of second signals. PMID- 12110132 TI - Humanized mice as a model for rheumatoid arthritis. AB - Genetic susceptibility to rheumatoid arthritis (RA), a common autoimmune disease, is associated with certain HLA-DR4 alleles. Treatments are rarely curative and are often tied to major side effects. We describe the development of a humanized mouse model wherein new, less toxic, vaccine-like treatments for RA might be pretested. This model includes four separate transgenes: HLA-DR*0401 and human CD4 molecules, a RA-related human autoantigenic protein (HCgp-39), and a T-cell receptor (TCRalphabeta) transgene specific for an important HCgp-39 epitope, eliciting strong Th1 responses in the context of HLA-DR*0401. PMID- 12110134 TI - HLA-B27: natural function and pathogenic role in spondyloarthritis. AB - The human leukocyte antigen HLA-B27 is strongly associated with development of a group of inflammatory arthritides collectively known as the spondyloarthritides. We have set out to define the natural immunological function of HLA-B27, and then to apply this knowledge to understand its pathogenic role. Human leukocyte antigen class 1 molecules bind antigenic peptides for cell surface presentation to cytotoxic T lymphocytes. HLA-B27 binds and presents peptides from influenza, HIV, Epstein-Barr virus, and other viruses. This leads to vigorous and specific cytotoxic T lymphocyte responses, which play an important role in the body's immune response to these viruses. HLA-B27 thus carries out its natural function highly effectively. Although many theories have been proposed to explain the role of HLA-B27 in the pathogenesis of spondyloarthropathy, we favour those postulating that the pathogenic role of HLA-B27 stems from its natural function. For example, the 'arthritogenic' peptide hypothesis suggests that disease results from the ability of HLA-B27 to bind a unique peptide or a set of antigenic peptides. Additionally, a number of lines of evidence from our laboratory and other laboratories have suggested that HLA-B27 has unusual cell biology. We have recently demonstrated that HLA-B27 is capable of forming disulfide-bonded homodimers. These homodimers are expressed on the cell surface and are ligands for a number of natural killer and related immunoreceptors, expressed on a variety of cell types including natural killer cells, T lymphocytes and B lymphocytes, and members of the monocyte/macrophage lineage. We are currently investigating the possibility that such interactions could be involved in disease pathogenesis. PMID- 12110133 TI - Multiple roles for tumor necrosis factor-alpha and lymphotoxin alpha/beta in immunity and autoimmunity. AB - Tumor necrosis factor (TNF)-alpha and lymphotoxin (LT) alpha/beta play multiple roles in the development and function of the immune system. This article focuses on three important aspects of the effects of these cytokines on the immune response and on autoimmunity. In several experimental systems (Jurkat T cells, murine T-cell hybridomas), TNF-alpha appears to cause a downregulation of signaling through the TCR, revealed by changes in calcium flux, activation of p21, p23 and ZAP70, and a decrease in nuclear activation of NF-kappaB. Previous and present results suggest that TNF-alpha interferes in some manner with signaling through the TCR, at a locus yet to be delineated. Transgenic expression of LTbetaR-Fc in nonobese diabetic (NOD) transgenic mice results in prevention of type 1 diabetes in NOD mice as long as the level of expression of the fusion protein (under the control of the cytomegalovirus promoter) remains above a level of 2-3 microg/ml. Once the expression levels of the fusion protein have dropped below this critical level, the diabetic process resumes and the animals become diabetic at 40-50 weeks of age, whereas nontransgenic littermates develop diabetes by 25-30 weeks of age. The paradoxical effects of neonatal TNF-alpha administration in NOD mice in increasing incidence of and hastening onset of type 1 diabetes, while neonatal anti-TNF administration completely prevents all signs of islet cell autoimmunity, are due partly to the low levels of CD4+CD25+ T cells in NOD mice. These low levels are reduced by a further 50% on neonatal administration of nontoxic levels of TNF-alpha. In contrast, neonatal administration of anti-TNF-alpha results in a dramatic increase in the levels of CD4+CD25+ regulatory T cells, to levels beyond those seen in wild-type untreated NOD mice. TNF-alpha and LTalpha/beta thus have pleomorphic regulatory effects on the development and expression of autoimmunity. PMID- 12110135 TI - The contrasting roles of IL-2 and IL-15 in the life and death of lymphocytes: implications for the immunotherapy of rheumatological diseases. AB - Interleukin-15 (IL-15) is a 14-15-kDa member of the 4alpha helix bundle family of cytokines that stimulate T and NK (natural killer) cells. IL-15 and IL-2 utilize heterotrimeric receptors that include the cytokine-specific private receptors IL 2Ralpha and IL-15Ralpha, as well as two receptor elements that they share, IL 2Rbeta and gammac. Although IL-2 and IL-15 share two receptor subunits and many functions, at times they provide contrasting contributions to T-cell-mediated immune responses. IL-2, through its pivotal role in activation-induced cell death (AICD), is involved in peripheral tolerance through the elimination of self reactive T cells. In contrast, IL-15 in general manifests anti-apoptotic actions and inhibits IL-2-mediated AICD. IL-15 stimulates the persistence of memory phenotype CD8+ T cells, whereas IL-2 inhibits their expression. Abnormalities of IL-15 expression have been described in patients with rheumatoid arthritis or inflammatory bowel disease and in diseases associated with the retrovirus HTLV-I (human T-cell lymphotropic virus I). Humanized monoclonal antibodies that recognize IL-2Ralpha, the private receptor for IL-2, are being employed to inhibit allograft rejection and to treat T-cell leukemia/lymphoma. New approaches directed toward inhibiting the actions of the inflammatory cytokine, IL-15, are proposed for an array of autoimmune disorders including rheumatoid arthritis as well as diseases associated with the retrovirus HTLV-I. PMID- 12110137 TI - Cytokine regulation in RA synovial tissue: role of T cell/macrophage contact dependent interactions. AB - Several groups have documented the expression of cytokines in rheumatoid arthritis synovial tissue over the past 15 years or so. These studies have indicated that most cytokines examined are expressed at the mRNA levels at least, and many other cytokines are found in abundance as proteins. Our attention has recently focused on the mechanisms that induce and regulate tumour necrosis factor and IL-10. Other workers and ourselves have found that cell-cell contact is an important signal for the induction of cytokines, and our work has demonstrated that tumour necrosis factor and IL-10 production in rheumatoid arthritis synovial joint cells cultures is dependent on T cell/macrophage interaction. In this chapter, we review recent advances in this area and also highlight areas where new therapeutic intervention opportunities arise. PMID- 12110136 TI - The role of human T-lymphocyte-monocyte contact in inflammation and tissue destruction. AB - Contact-mediated signaling of monocytes by human stimulated T lymphocytes (TL) is a potent proinflammatory mechanism that triggers massive upregulation of the proinflammatory cytokines IL-1 and tumor necrosis factor-alpha. These two cytokines play an important part in chronic destructive diseases, including rheumatoid arthritis. To date this cell-cell contact appears to be a major endogenous mechanism to display such an activity in monocyte-macrophages. Since TL and monocyte-macrophages play a pivotal part in the pathogenesis of chronic inflammatory diseases, we investigated the possible ligands and counter-ligands involved in this cell-cell interaction. We also characterized an inhibitory molecule interfering in this process, apolipoprotein A-I. This review aims to summarize the state of the art and importance of contact-mediated monocyte activation by stimulated TL in cytokine production in rheumatoid arthritis and mechanisms that might control it. PMID- 12110138 TI - Chemokine receptors on dendritic cells promote autoimmune reactions. AB - This paper presents a brief review of several lines of evidence suggesting that chemokine receptors on dendritic cells play an important role in breaking tolerance to self and in inducing autoimmunity. First, we have shown that an idiotypic self-antigen obtained from malignant murine lymphomas, when covalently linked to selected chemokines or defensins that interact with receptors on immature dendritic cells (iDCs), has the capacity to break tolerance to self and induce humoral or cell-mediated anti-tumor responses. Since unlinked antigens mixed with the same chemokines or defensins or antigens fused with a mutant ligand deficient in receptor-binding capacity were not immunogenic, we propose that delivery of an antigen coupled to a ligand for receptors on iDCs promotes the processing and subsequent presentation of the antigen, resulting in immunoadjuvant effects. In a second study, we observed that two of five aminoacyl tRNA synthetases (aaRSs) - which act as autoantigens to which some patients with myositis have autoantibodies - were chemotactic for activated monocytes, T cells, and iDCs. These aaRSs interacted with either CC chemokine receptor (CCR)5 or CCR3, as was shown by desensitization with chemokines and the response of cell lines transfected with the chemokine receptor. Presumably, these autoantigens therefore have the capacity to attract inflammatory cells, including iDCs, to infiltrate affected muscle cells. These observations suggest the hypothesis that antigens delivered to receptors on iDCs are potent immunogens capable of breaking self-tolerance to tumor antigens to induce autoimmune diseases. PMID- 12110139 TI - How are the regulators regulated? The search for mechanisms that impose specificity on induction of cell death and NF-kappaB activation by members of the TNF/NGF receptor family. AB - Signals emanating from receptors of the tumor necrosis factor/nerve growth factor (TNF/NGF) family control practically all aspects of immune defense and, as such, constitute potential targets for therapeutic intervention through rational drug design. Indeed, arrest of these signals by blocking ligand-receptor interactions enables effective suppression of a variety of activities that are implicated in various pathologies, such as T and B lymphocyte activation and growth, inflammation, fibroblast proliferation, and cell death. To be therapeutically useful, however, inhibition of signaling should be restricted by determinants of specificity, at least to the same degree observed when blocking activation of individual receptors. In spite of their broad range of functions, receptors of the TNF/NGF family are known to activate just a few signaling pathways. Of these, the most extensively studied are the activation of the caspase protease cascade, which leads to cell death, and the activation of NF-kappaB (nuclear factor kappaB) transcription factors through protein phosphorylation cascades. Until recently, most studies of the two pathways have solely focused on the core signaling complexes that are shared by the different receptors: death-inducing complexes containing the cysteine proteases caspase-8 and caspase-10, bound to the adapter protein MORT1/FADD (mediator of receptor-induced toxicity/Fas associated DD protein), and the NF-kappaB-activating complex, composed of the protein kinases IKK1 (IkappaB kinase 1) and IKK2 (IkappaB kinase 2) and the regulatory subunit NEMO (NF-kappaB essential modulator; the 'IKK signalosome'). Knowledge has begun to emerge of additional molecules and mechanisms that affect these basic signaling complexes and impose specificity on their function. PMID- 12110140 TI - Studies of T-cell activation in chronic inflammation. AB - The strong association between specific alleles encoded within the MHC class II region and the development of rheumatoid arthritis (RA) has provided the best evidence to date that CD4+ T cells play a role in the pathogenesis of this chronic inflammatory disease. However, the unusual phenotype of synovial T cells, including their profound proliferative hyporesponsiveness to TCR ligation, has challenged the notion that T-cell effector responses are driven by cognate cartilage antigens in inflamed synovial joints. The hierarchy of T-cell dysfunction from peripheral blood to inflamed joint suggests that these defects are acquired through prolonged exposure to proinflammatory cytokines such as tumour necrosis factor (TNF)-alpha. Indeed, there are now compelling data to suggest that chronic cytokine activation may contribute substantially to the phenotype and effector function of synovial T cells. Studies reveal that chronic exposure of T cells to TNF uncouples TCR signal transduction pathways by impairing the assembly and stability of the TCR/CD3 complex at the cell surface. Despite this membrane-proximal effect, TNF selectively uncouples downstream signalling pathways, as is shown by the dramatic suppression of calcium signalling responses, while Ras/ERK activation is spared. On the basis of these data, it is proposed that T-cell survival and effector responses are driven by antigen-independent, cytokine-dependent mechanisms, and that therapeutic strategies that seek to restore T-cell homeostasis rather than further depress T cell function should be explored in the future. PMID- 12110141 TI - High-efficiency gene transfer into nontransformed cells: utility for studying gene regulation and analysis of potential therapeutic targets. AB - The elucidation of the signalling pathways involved in inflammatory diseases, such as rheumatoid arthritis, could provide long sought after targets for therapeutic intervention. Gene regulation is complex and varies depending on the cell type, as well as the signal eliciting gene activation. However, cells from certain lineages, such as macrophages, are specialised to degrade exogenous material and consequently do not easily transfect. Methods for high-efficiency gene transfer into primary cells of various lineages and disease states are desirable, as they remove the uncertainties associated with using transformed cell lines. Significant research has been undertaken into the development of nonviral and viral vectors for basic research, and as vehicles for gene therapy. We briefly review the current methods of gene delivery and the difficulties associated with each system. Adenoviruses have been used extensively to examine the role of various cytokines and signal transduction molecules in the pathogenesis of rheumatoid arthritis. This review will focus on the involvement of different signalling molecules in the production of tumour necrosis factor alpha by macrophages and in rheumatoid synovium. While the NF-kappaB pathway has proven to be a major mediator of tumour necrosis factor alpha production, it is not exclusive and work evaluating the involvement of other pathways is ongoing. PMID- 12110142 TI - Signaling crosstalk between RANKL and interferons in osteoclast differentiation. AB - Regulation of osteoclast differentiation is an aspect central to the understanding of the pathogenesis and the treatment of bone diseases such as autoimmune arthritis and osteoporosis. In fact, excessive signaling by RANKL (receptor activator of nuclear factor kappaB ligand), a member of the tumor necrosis factor (TNF) family essential for osteoclastogenesis, may contribute to such pathological conditions. Here we summarize our current work on the negative regulation of osteoclastogenesis by unique signaling crosstalk between RANKL and interferons (IFNs). First, activated T cells maintain bone homeostasis by counterbalancing the action of RANKL through production of IFN-gamma. This cytokine induces rapid degradation of the RANK (receptor activator of nuclear factor kappaB) adapter protein TRAF6 (TNF-receptor-associated factor 6), resulting in strong inhibition of the RANKL-induced activation of NF-kappaB and JNK (c-Jun N-terminal kinase). Second, RANKL induces the IFN-beta gene but not IFN-alpha genes, in osteoclast precursor cells, and that IFN-beta strongly inhibits the osteoclast differentiation by interfering with the RANKL-induced expression of c-Fos. The series of in vivo experiments revealed that these two distinct IFN-mediated regulatory mechanisms are both important to maintain homeostasis of bone resorption. Collectively, these studies revealed novel aspects of the two types of IFN, beyond their original roles in the immune response, and may offer a molecular basis for the treatment of bone diseases. PMID- 12110144 TI - Signaling for survival and apoptosis in the immune system. AB - Signal transduction induced by tumor necrosis factor (TNF) family members and their receptors has been an intensive area of research for several years. The major impact of these studies has been the delineation of apoptotic and cell survival signaling pathways. These discoveries, coupled with major advances in the study of mammalian apoptotic machinery, constitute a promising blueprint of the molecular network governing the fate of all living cells. In this review, we concentrate on the fate of cells in the immune system, where regulation of cell death and cell survival is a frequent and important exercise. A small imbalance in favor of either fate can result in disastrous pathological outcomes, such as cancer, autoimmunity or immune deficiency. It is an insurmountable task to discuss all molecules reported in the literature that are implicated in lymphocyte death or survival. We have therefore focused on discoveries made by mouse gene targeting, as these studies provide the most physiologically relevant information on each molecule. We begin with a description of signaling channels initiated by TNF receptor type 1 engagement, which can lead to either cell survival or to cell death. The point of bifurcation of this pathway and the decision-making molecules FADD, TRAF2 and RIP are discussed. We then follow apoptotic and survival pathways from upstream to downstream, describing many important players involved in signal transduction. Molecules important for NF kappaB and JNK/stress-activated protein kinase activation such as IKKbeta, NEMO, MAP3K and TRAF6 are discussed, as is the impact of BAFF and its receptors on B cell survival. Mouse mutants that have helped to define the mammalian apoptosis execution machinery, including animals lacking Apaf-1, caspase-3 and caspase-9, are also described. We conclude with a brief analysis of the potential therapeutic options arising from this body of work. PMID- 12110143 TI - The paradigm of IL-6: from basic science to medicine. AB - IL-6 is a pleiotropic cytokine with a wide range of biological activities in immune regulation, hematopoiesis, inflammation, and oncogenesis. Its activities are shared by IL-6-related cytokines such as leukemia inhibitory factor and oncostatin M. The pleiotropy and redundancy of IL-6 functions have been identified by using a unique receptor system comprising two functional proteins: an IL-6 receptor (IL-6R) and gp130, the common signal transducer of cytokines related to IL-6. Signal transduction through gp130 is mediated by two pathways: the JAK-STAT (Janus family tyrosine kinase-signal transducer and activator of transcription) pathway and the Ras mitogen-activated protein kinase pathway. The negative regulators of IL-6 signaling have also been identified, although the physiological roles of the molecules are not yet fully understood. The pathological roles of IL-6 have also been clarified in various disease conditions, such as inflammatory, autoimmune, and malignant diseases. On the basis of the findings, a new therapeutic approach to block the IL-6 signal using humanized anti-IL-6R antibody for rheumatoid arthritis, Castleman's disease, and multiple myeloma has been attempted. PMID- 12110145 TI - Susceptibility genes in the pathogenesis of murine lupus. AB - Systemic lupus erythematosus (SLE) is the paradigm of a multisystem autoimmune disease in which genetic factors strongly influence susceptibility. Through genome scans and congenic dissection, numerous loci associated with lupus susceptibility have been defined and the complexity of the inheritance of this disease has been revealed. In this review, we provide a brief description of animal models of SLE, both spontaneous models and synthetic models, with an emphasis on the B6 congenic model derived from analyses of the NZM2410 strain. A hypothetical model of disease progression that organizes many of the identified SLE susceptibility loci in three distinct biological pathways that interact to mediate disease pathogenesis is also described. We finally discuss our recent fine mapping analysis, which revealed a cluster of loci that actually comprise the Sle1 locus. PMID- 12110146 TI - Epidemiology and genetics of rheumatoid arthritis. AB - The prevalence of rheumatoid arthritis (RA) is relatively constant in many populations, at 0.5-1.0%. However, a high prevalence of RA has been reported in the Pima Indians (5.3%) and in the Chippewa Indians (6.8%). In contrast, low occurrences have been reported in populations from China and Japan. These data support a genetic role in disease risk. Studies have so far shown that the familial recurrence risk in RA is small compared with other autoimmune diseases. The main genetic risk factor of RA is the HLA DRB1 alleles, and this has consistently been shown in many populations throughout the world. The strongest susceptibility factor so far has been the HLA DRB1*0404 allele. Tumour necrosis factor alleles have also been linked with RA. However, it is estimated that these genes can explain only 50% of the genetic effect. A number of other non-MHC genes have thus been investigated and linked with RA (e.g. corticotrophin releasing hormone, oestrogen synthase, IFN-gamma and other cytokines). Environmental factors have also been studied in relation to RA. Female sex hormones may play a protective role in RA; for example, the use of the oral contraceptive pill and pregnancy are both associated with a decreased risk. However, the postpartum period has been highlighted as a risk period for the development of RA. Furthermore, breastfeeding after a first pregnancy poses the greatest risk. Exposure to infection may act as a trigger for RA, and a number of agents have been implicated (e.g. Epstein-Barr virus, parvovirus and some bacteria such as Proteus and Mycoplasma). However, the epidemiological data so far are inconclusive. There has recently been renewed interest in the link between cigarette smoking and RA, and the data presented so far are consistent with and suggestive of an increased risk. PMID- 12110147 TI - Single nucleotide polymorphisms and disease gene mapping. AB - Single nucleotide polymorphisms are the most important and basic form of variation in the genome, and they are responsible for genetic effects that produce susceptibility to most autoimmune diseases. The rapid development of databases containing very large numbers of single nucleotide polymorphisms, and the characterization of haplotypes and patterns of linkage disequilibrium throughout the genome, provide a unique opportunity to advance association strategies in common disease rapidly over the next few years. Only the careful use of these strategies and a clear understanding of their statistical limits will allow novel genetic determinants for many of the common autoimmune diseases to be determined. PMID- 12110149 TI - Why do we not have a cure for rheumatoid arthritis? AB - There are currently unprecedented opportunities to treat rheumatoid arthritis using well-designed, highly effective, targeted therapies. This will result in a substantial improvement in the outcome of this disorder for most affected individuals, if they can afford these therapies. Yet our lack of understanding of the basic mechanisms that initiate and sustain this disease remains a major obstacle in the search for a definitive cure. It is possible, if not likely, that our best approach will be to identify individuals at risk and devise reliable, safe methods of preventing the disease before it occurs. The means to do this are currently unknown but should serve as a major focus of research. PMID- 12110151 TI - Magnetic resonance imaging: opportunities for rheumatoid arthritis disease assessment and monitoring long-term treatment outcomes. AB - Early diagnosis of rheumatoid arthritis (RA) combined with early initiation of an appropriate treatment regimen is acknowledged as an important factor in improving clinical outcomes in patients with RA. Early diagnosis allows treatment intervention to occur sooner in order to inhibit the progression of structural joint damage as well as providing improved patient quality of life. Unfortunately, early diagnosis has been challenging due to the non-specific signs and symptoms associated with many polyarthropathies and the lack of accurate definitive diagnostic tests that can accurately classify RA at presentation. The emphasis on early diagnosis has fueled the need for powerful, sensitive, non invasive imaging techniques that not only accurately define RA and give an indication of prognosis, but can also serve as a tool to monitor long-term treatment outcomes. This article reviews the potential uses of magnetic resonance imaging as a tool for the classification, documentation, and clinical monitoring of RA. PMID- 12110150 TI - Autoantibodies in rheumatoid arthritis and their clinical significance. AB - Autoantibodies are proven useful diagnostic tools for a variety of rheumatic and non-rheumatic autoimmune disorders. However, a highly specific marker autoantibody for rheumatoid arthritis (RA) has not yet been determined. The presence of rheumatoid factors is currently used as a marker for RA. However, rheumatoid factors have modest specificity (~70%) for the disease. In recent years, several newly characterized autoantibodies have become promising candidates as diagnostic indicators for RA. Antikeratin, anticitrullinated peptides, anti-RA33, anti-Sa, and anti-p68 autoantibodies have been shown to have >90% specificity for RA. These autoantibodies are reviewed and the potential role of the autoantibodies in the pathogenesis of RA is briefly discussed. PMID- 12110148 TI - Complement and systemic lupus erythematosus. AB - Complement is implicated in the pathogenesis of systemic lupus erythematosus (SLE) in several ways and may act as both friend and foe. Homozygous deficiency of any of the proteins of the classical pathway is causally associated with susceptibility to the development of SLE, especially deficiency of the earliest proteins of the activation pathway. However, complement is also implicated in the effector inflammatory phase of the autoimmune response that characterizes the disease. Complement proteins are deposited in inflamed tissues and, in experimental models, inhibition of C5 ameliorates disease in a murine model. As a further twist to the associations between the complement system and SLE, autoantibodies to some complement proteins, especially to C1q, develop as part of the autoantibody response. The presence of anti-C1q autoantibodies is associated with severe illness, including glomerulonephritis. In this chapter the role of the complement system in SLE is reviewed and hypotheses are advanced to explain the complex relationships between complement and lupus. PMID- 12110152 TI - The determination and measurement of functional disability in rheumatoid arthritis. AB - Although functional outcome is frequently discussed and written about, it is often not clear what functional outcome is and how it can be measured. This paper introduces the concept of latent and observed measures of functional disability, and distinguishes between disability as a process measure and disability as an outcome measure. Using the Health Assessment Questionnaire as the main functional outcome measure in rheumatoid arthritis, we propose and discuss several methods for determining disability, and describe the implications of altering the disability course. PMID- 12110153 TI - Current and future management approaches for rheumatoid arthritis. AB - With the introduction of new disease-modifying antirheumatic drugs (DMARDs) and other therapeutic agents, the management of rheumatoid arthritis (RA) has shifted toward earlier, more aggressive therapy. The ultimate goal is to prevent structural joint damage that leads to pain and functional disability. Early diagnosis of RA is therefore essential, and early DMARD treatment combined with nonsteroidal anti-inflammatory drugs is recommended. Combination DMARD regimens and new biologic agents (anti-tumor necrosis factor [TNF] therapies [infliximab, etanercept] and the interleukin [IL]-1 antagonist [anakinra]) have emerged as viable options for early treatment of RA patients. These new biologic agents and future nonbiologic agents that target proteins in signaling cascades are likely to change the landscape of RA treatments. PMID- 12110154 TI - How does infliximab work in rheumatoid arthritis? AB - Since the initial characterization of tumor necrosis factor alpha (TNFalpha), it has become clear that TNFalpha has diverse biologic activity. The realization that TNFalpha plays a role in rheumatoid arthritis (RA) has led to the development of anti-TNF agents for the treatment of RA. Infliximab, a chimeric monoclonal antibody that specifically, and with high affinity, binds to TNFalpha and neutralizes the cytokine, is currently approved for the treatment of RA and Crohn's disease, another immune-inflammatory disorder. In addition to establishing the safety and efficacy of infliximab, clinical research has also provided insights into the complex cellular and cytokine-dependent pathways involved in the pathophysiology of RA, including evidence that supports TNFalpha involvement in cytokine regulation, cell recruitment, angiogenesis, and tissue destruction. PMID- 12110155 TI - Structural damage in rheumatoid arthritis as visualized through radiographs. AB - Several agents show an effect on reducing radiographic progression in rheumatoid arthritis. It is tempting to retrospectively compare the effects of these agents on radiographic progression across clinical trials. However, there are several limitations in interpreting and comparing radiographic results across clinical trials. These limitations, including study designs, patient characteristics, durations of follow-up, scoring methodologies, reader reliability, radiograph sequence, handling of missing data, and data presentation, will be discussed. The consequences are illustrated with several examples of recent clinical trials that show an effect on radiographic progression. A guide in the interpretation and clinical relevance of radiographic results is presented, with the Anti-TNF Trial in Rheumatoid Arthritis with Concomitant Therapy used as an example. PMID- 12110156 TI - Emerging role of anti-tumor necrosis factor therapy in rheumatic diseases. AB - Tumor necrosis factor alpha (TNF-alpha) is an inflammatory cytokine that has been implicated in a variety of rheumatic and inflammatory diseases. New understanding of the importance of TNF-alpha in the pathophysiology of rheumatoid arthritis and Crohn's disease led to the development of a new class of targeted anti-TNF therapies. Anti-TNF-alpha agents including etanercept (a fusion protein of the p75 TNF receptor and IgG1) and infliximab (a chimeric monoclonal antibody specific for TNF-alpha) have been approved for the treatment of rheumatoid arthritis. In addition, infliximab has been approved in the treatment of patients with active or fistulating Crohn's disease. A new appreciation of the importance of TNF-alpha in other rheumatic and inflammatory diseases has led to a broadening of the application of anti-TNF agents. Both etanercept and infliximab have been used in open-label and randomized studies in patients with psoriatic arthritis. Although larger randomized trials are needed to confirm early results, both these anti-TNF-alpha agents, etanercept and infliximab, have demonstrated activity in improving the signs and symptoms of psoriatic arthritis and psoriasis. Infliximab has also been shown to be effective in patients with other rheumatic diseases, including ankylosing spondylitis, and may be effective in adult-onset Still's disease, polymyositis, and Behcet's disease. Further investigations will fully elucidate the role of infliximab in these and other rheumatic diseases. PMID- 12110157 TI - Application of the development stages of a cluster randomized trial to a framework for valuating complex health interventions. AB - INTRODUCTION: Trials of complex health interventions often pose difficult methodologic challenges. The objective of this paper is to assess the extent to which the various development steps of a cluster randomized trial to optimize antibiotic use in nursing homes are represented in a recently published framework for the design and evaluation of complex health interventions. In so doing, the utility of the framework for health services researchers is evaluated. METHODS: Using the five phases of the framework (theoretical, identification of components of the intervention, definition of trial and intervention design, methodological issues for main trial, promoting effective implementation), corresponding stages in the development of the cluster randomized trial using diagnostic and treatment algorithms to optimize the use of antibiotics in nursing homes are identified and described. RESULTS: Synthesis of evidence needed to construct the algorithms, survey and qualitative research used to define components of the algorithms, a pilot study to assess the feasibility of delivering the algorithms, methodological issues in the main trial including choice of design, allocation concealment, outcomes, sample size calculation, and analysis are adequately represented using the stages of the framework. CONCLUSIONS: The framework is a useful resource for researchers planning a randomized clinical trial of a complex intervention. PMID- 12110159 TI - BiP binding keeps ATF6 at bay. AB - A study by, in this issue of Developmental Cell shows that transport to the Golgi complex and subsequent proteolytic activation of the stress-regulated transcription factor ATF6 is initiated by the dissociation of the ER chaperone BiP from ATF6. This demonstrates that BiP is a key element in sensing the folding capacity within the ER and provides mechanistic insights on how the activation of membrane-bound transcription factors can be regulated. PMID- 12110158 TI - Avian papillomaviruses: the parrot Psittacus erithacus papillomavirus (PePV) genome has a unique organization of the early protein region and is phylogenetically related to the chaffinch papillomavirus. AB - BACKGROUND: An avian papillomavirus genome has been cloned from a cutaneous exophytic papilloma from an African grey parrot (Psittacus erithacus). The nucleotide sequence, genome organization, and phylogenetic position of the Psittacus erithacus papillomavirus (PePV) were determined. This PePV sequence represents the first complete avian papillomavirus genome defined. RESULTS: The PePV genome (7304 basepairs) differs from other papillomaviruses, in that it has a unique organization of the early protein region lacking classical E6 and E7 open reading frames. Phylogenetic comparison of the PePV sequence with partial E1 and L1 sequences of the chaffinch (Fringilla coelebs) papillomavirus (FPV) reveals that these two avian papillomaviruses form a monophyletic cluster with a common branch that originates near the unresolved center of the papillomavirus evolutionary tree. CONCLUSIONS: The PePV genome has a unique layout of the early protein region which represents a novel prototypic genomic organization for avian papillomaviruses. The close relationship between PePV and FPV, and between their Psittaciformes and Passeriformes hosts, supports the hypothesis that papillomaviruses have co-evolved and speciated together with their host species throughout evolution. PMID- 12110160 TI - How to grab a microtubule on the move. AB - In migrating cells, Rho family GTPases and their effectors play a central role in polarizing and in organizing the actin and microtubule cytoskeletons. A study by Fukata et al. in the June 28th issue of Cell now shows that the Rac1/Cdc42 effector IQGAP1 captures microtubules by binding to CLIP170. PMID- 12110161 TI - Top-SUMO wrestles centromeric cohesion. AB - Sister chromatid cohesion at the centromere is distinct from cohesion at the chromosome arms. In the June issue of Molecular Cell, Bachant et al. have shown that centromeric cohesion in budding yeast is specifically regulated by SUMO-1 modification of Topoisomerase II. PMID- 12110162 TI - Genetics leads the way to the accomplices of presenilins. AB - Presenilins mediate the gamma-secretase cleavage of Notch transmembrane receptors as well as the transmembrane beta-amyloid precursor protein (betaAPP), but they are not thought to accomplish this alone. Recent genetic screens in C. elegans, presented in this issue of Developmental Cell, identify two genes that are essential to gamma-secretase activity and may interact with presenilins. PMID- 12110163 TI - Dynamic analysis of dorsal closure in Drosophila: from genetics to cell biology. AB - Throughout development a series of epithelial bendings, sweepings, and fusions occur that collectively give shape to the embryo. These morphogenetic movements are driven by coordinated assembly and contraction of the actomyosin cytoskeleton in restricted populations of epithelial cells. One well-studied example of such a morphogenetic episode is dorsal closure in Drosophila embryogenesis. This process is tractable at a genetic level and has recently become the focus of live cell biology analysis because of the availability of flies expressing GFP-fusion proteins. This marriage of genetics and cell biology is very powerful and is allowing the dissection of fundamental signaling mechanisms that regulate the cytoskeletal reorganizations and contractions underlying coordinated tissue movements in the embryo. PMID- 12110164 TI - Stephen Jay Gould (1941-2002): a wonderful life. PMID- 12110165 TI - Adipose tissue selective insulin receptor knockout protects against obesity and obesity-related glucose intolerance. AB - Insulin signaling in adipose tissue plays an important role in lipid storage and regulation of glucose homeostasis. Using the Cre-loxP system, we created mice with fat-specific disruption of the insulin receptor gene (FIRKO mice). These mice have low fat mass, loss of the normal relationship between plasma leptin and body weight, and are protected against age-related and hypothalamic lesion induced obesity, and obesity-related glucose intolerance. FIRKO mice also exhibit polarization of adipocytes into populations of large and small cells, which differ in expression of fatty acid synthase, C/EBP alpha, and SREBP-1. Thus, insulin signaling in adipocytes is critical for development of obesity and its associated metabolic abnormalities, and abrogation of insulin signaling in fat unmasks a heterogeneity in adipocyte response in terms of gene expression and triglyceride storage. PMID- 12110166 TI - E2Fs regulate adipocyte differentiation. AB - When preadipocytes reenter the cell cycle, PPAR gamma expression is induced, coincident with an increase in DNA synthesis, suggesting the involvement of the E2F family of cell cycle regulators. We show here that E2F1 induces PPAR gamma transcription during clonal expansion, whereas E2F4 represses PPARg amma expression during terminal adipocyte differentiation. Using a combination of in vivo experiments with knockout and chimeric animals and in vitro experiments, we demonstrate that the absence of E2F1 impairs, whereas depletion of E2F4 stimulates, adipogenesis. E2Fs hence represent the link between proliferative signaling pathways, triggering clonal expansion, and terminal adipocyte differentiation through regulation of PPAR gamma expression. This underscores the complex role of the E2F protein family in the control of both cell proliferation and differentiation. PMID- 12110167 TI - tej defines a role for poly(ADP-ribosyl)ation in establishing period length of the arabidopsis circadian oscillator. AB - In a genetic screen for altered circadian period length in Arabidopsis, we isolated a mutant with a long free-running period. The tej mutation acts independently of light quality and quantity. It affects clock-controlled transcription of genes in Arabidopsis and alters the timing of the photoperiod dependent transition from vegetative growth to flowering. Map-based cloning of TEJ identified a poly(ADP-ribose) glycohydrolase (PARG). An inhibitor of poly(ADP ribosyl)ation rescued the period phenotype of tej mutant and shortened the period length of wild-type plants. Posttranslational poly(ADP-ribosyl)ation of an oscillator component may contribute to setting the period length of the Arabidopsis central oscillator. PMID- 12110168 TI - Periodic Lunatic fringe expression is controlled during segmentation by a cyclic transcriptional enhancer responsive to notch signaling. AB - A molecular oscillator regulates the pace of vertebrate segmentation. Here, we show that the oscillator (clock) controls cyclic initiation of transcription in the unsegmented presomitic mesoderm (PSM). We identify an evolutionarily conserved 2.3 kb region in the murine Lunatic fringe (Lfng) promoter that drives periodic expression in the PSM. This region includes conserved blocks required for enhancing and repressing cyclic Lfng transcription, and to prevent continued expression in formed somites. We also show that dynamic expression in the cycling PSM is lost in the total absence of Notch signaling, and that Notch signaling acts directly via CBF1/RBP-Jkappa binding sites to regulate Lfng. These results are consistent with a model in which oscillatory Notch signaling underlies the segmentation clock and directly activates and indirectly represses Lfng expression. PMID- 12110169 TI - Clock regulatory elements control cyclic expression of Lunatic fringe during somitogenesis. AB - Somitogenesis requires a segmentation clock and Notch signaling. Lunatic fringe (Lfng) expression in the presomitic mesoderm (PSM) cycles in the posterior PSM, is refined in the segmenting somite to the rostral compartment, and is required for segmentation. We identify distinct cis-acting regulatory elements for each aspect of Lfng expression. Fringe clock element 1 (FCE1) represents a conserved 110 bp region that is necessary to direct cyclic Lfng RNA expression in the posterior PSM. Mutational analysis of E boxes within FCE1 indicates a potential interplay of positive and negative transcriptional regulation by cyclically expressed bHLH proteins. A separable Lfng regulatory region directs expression to the prospective rostral aspect of the condensing somite. These independent Lfng regulatory cassettes advance a molecular framework for deciphering somite segmentation. PMID- 12110171 TI - ER stress regulation of ATF6 localization by dissociation of BiP/GRP78 binding and unmasking of Golgi localization signals. AB - ATF6 is an endoplasmic reticulum (ER) stress-regulated transmembrane transcription factor that activates the transcription of ER molecular chaperones. Upon ER stress, ATF6 translocates from the ER to the Golgi where it is processed to its active form. We have found that the ER chaperone BiP/GRP78 binds ATF6 and dissociates in response to ER stress. Loss of BiP binding correlates with the translocation of ATF6 to the Golgi, which was slowed in cells overexpressing BiP. Two Golgi localization signals (GLSs) were identified in ATF6. Removal of BiP binding sites from ATF6, while retaining a GLS, resulted in its constitutive translocation to the Golgi. These results suggest that BiP retains ATF6 in the ER by inhibiting its GLSs and that dissociation of BiP during ER stress allows ATF6 to be transported to the Golgi. PMID- 12110170 TI - aph-1 and pen-2 are required for Notch pathway signaling, gamma-secretase cleavage of betaAPP, and presenilin protein accumulation. AB - Presenilins are components of the gamma-secretase protein complex that mediates intramembranous cleavage of betaAPP and Notch proteins. A C. elegans genetic screen revealed two genes, aph-1 and pen-2, encoding multipass transmembrane proteins, that interact strongly with sel-12/presenilin and aph-2/nicastrin. Human aph-1 and pen-2 partially rescue the C. elegans mutant phenotypes, demonstrating conserved functions. The human genes must be provided together to rescue the mutant phenotypes, and the inclusion of presenilin-1 improves rescue, suggesting that they interact closely with each other and with presenilin. RNAi mediated inactivation of aph-1, pen-2, or nicastrin in cultured Drosophila cells reduces gamma-secretase cleavage of betaAPP and Notch substrates and reduces the levels of processed presenilin. aph-1 and pen-2, like nicastrin, are required for the activity and accumulation of gamma-secretase. PMID- 12110172 TI - SRC-1 and Wnt signaling act together to specify endoderm and to control cleavage orientation in early C. elegans embryos. AB - In early C. elegans embryos, signaling between a posterior blastomere, P2, and a ventral blastomere, EMS, specifies endoderm and orients the division axis of the EMS cell. Although Wnt signaling contributes to this polarizing interaction, no mutants identified to date abolish P2/EMS signaling. Here, we show that two tyrosine kinase-related genes, src-1 and mes-1, are required for the accumulation of phosphotyrosine between P2 and EMS. Moreover, src-1 and mes-1 mutants strongly enhance endoderm and EMS spindle rotation defects associated with Wnt pathway mutants. SRC-1 and MES-1 signal bidirectionally to control cell fate and division orientation in both EMS and P2. Our findings suggest that Wnt and Src signaling function in parallel to control developmental outcomes within a single responding cell. PMID- 12110173 TI - sonic hedgehog and vascular endothelial growth factor act upstream of the Notch pathway during arterial endothelial differentiation. AB - The appearance of molecular differences between arterial and venous endothelial cells before circulation suggests that genetic factors determine these cell types. We find that vascular endothelial growth factor (vegf) acts downstream of sonic hedgehog (shh) and upstream of the Notch pathway to determine arterial cell fate. Loss of Vegf or Shh results in loss of arterial identity, while exogenous expression of these factors causes ectopic expression of arterial markers. Microinjection of vegf mRNA into embryos lacking Shh activity can rescue arterial differentiation. Finally, activation of the Notch pathway in the absence of Vegf signaling can rescue arterial marker gene expression. These studies reveal a complex signaling cascade responsible for establishing arterial cell fate and suggest differential effects of Vegf on developing endothelial cells. PMID- 12110175 TI - Destabilizing influences in apoptosis: sowing the seeds of IAP destruction. AB - Inhibitor of apoptosis proteins (IAPs) can block apoptosis through interactions with members of the caspase family of cysteine proteases. Recent developments suggest that ubiquitin-proteasome mediated destruction of the Drosophila IAP, DIAP1, is a key event during the initiation of programmed cell death in the fly. PMID- 12110174 TI - Myeloid or lymphoid promiscuity as a critical step in hematopoietic lineage commitment. AB - We demonstrate here that "promiscuous" expression of myeloid or lymphoid genes precedes lineage commitment in hematopoiesis. Prospectively purified single common myeloid progenitors (CMPs) coexpress myelo-erythroid but not lymphoid genes, whereas single common lymphoid progenitors (CLPs) coexpress T and B lymphoid but not myeloid genes. Genes unrelated to the adopted lineage are downregulated in bipotent and monopotent descendants of CMPs and CLPs. Promiscuous gene expression does not alter the biological potential of multipotent progenitors: CMPs with an activated endogenous M lysozyme locus yield normal proportions of myelo-erythroid colonies, and CLPs expressing the pre-T cell receptor alpha gene differentiate into normal numbers of B cells. Thus, the accessibility for multiple myeloid or lymphoid programs promiscuously may allow flexibility in fate commitments at these multipotent stages. PMID- 12110176 TI - RNA chaperones exist and DEAD box proteins get a life. AB - The RNA chaperone hypothesis suggests the existence of proteins that resolve misfolded RNA structures in vivo. A recent study has found an RNA-dependent ATPase that functions in this manner. PMID- 12110177 TI - Histone methylation: dynamic or static? AB - Methylation of histones mediates transcriptional silencing at heterochromatin sites and affects regulated transcription at euchromatic loci. So is the methyl group a permanent mark on histones, or can it be removed by an active process necessary for regulated gene expression? PMID- 12110178 TI - Bidirectional gene organization: a common architectural feature of the human genome. PMID- 12110179 TI - Unrepaired DNA breaks in p53-deficient cells lead to oncogenic gene amplification subsequent to translocations. AB - Amplification of large genomic regions associated with complex translocations (complicons) is a basis for tumor progression and drug resistance. We show that pro-B lymphomas in mice deficient for both p53 and nonhomologous end-joining (NHEJ) contain complicons that coamplify c-myc (chromosome 15) and IgH (chromosome 12) sequences. While all carry a translocated (12;15) chromosome, coamplified sequences are located within a separate complicon that often involves a third chromosome. Complicon formation is initiated by recombination of RAG1/2 catalyzed IgH locus double-strand breaks with sequences downstream of c-myc, generating a dicentric (15;12) chromosome as an amplification intermediate. This recombination event employs a microhomology-based end-joining repair pathway, as opposed to classic NHEJ or homologous recombination. These findings suggest a general model for oncogenic complicon formation. PMID- 12110180 TI - Requirement of yeast RAD2, a homolog of human XPG gene, for efficient RNA polymerase II transcription. implications for Cockayne syndrome. AB - In addition to xeroderma pigmentosum, mutations in the human XPG gene cause early onset Cockayne syndrome (CS). Here, we provide evidence for the involvement of RAD2, the S. cerevisiae counterpart of XPG, in promoting efficient RNA polymerase II transcription. Inactivation of RAD26, the S. cerevisiae counterpart of the human CSB gene, also causes a deficiency in transcription, and a synergistic decline in transcription occurs in the absence of both the RAD2 and RAD26 genes. Growth is also retarded in the rad2 Delta and rad26 Delta single mutant strains, and a very severe growth inhibition is seen in the rad2 Delta rad26 Delta double mutant. From these and other observations presented here, we suggest that transcriptional defects are the underlying cause of CS. PMID- 12110181 TI - Yph1p, an ORC-interacting protein: potential links between cell proliferation control, DNA replication, and ribosome biogenesis. AB - Immunoprecipitation of the origin recognition complex (ORC) from yeast extracts identified Yph1p, an essential protein containing a BRCT domain. Two Yph1p complexes were characterized. Besides ORC, MCM proteins, cell-cycle regulatory proteins, checkpoint proteins, 60S ribosomal proteins, and preribosome particle proteins were found to be associated with Yph1p. Yph1p is predominantly nucleolar and is required for 60S ribosomal subunit biogenesis and possibly for translation on polysomes. Proliferating cells depleted of Yph1p arrest in G(1) or G(2), with no cells in S phase, or significantly delay S phase progression after release from a hydroxyurea arrest. Yph1p levels decline as cells commit to exit the cell cycle, and levels vary depending on energy source. Yph1p may link cell proliferation control to DNA replication, ribosome biogenesis, and translation on polysomes. PMID- 12110182 TI - Noc3p, a bHLH protein, plays an integral role in the initiation of DNA replication in budding yeast. AB - Initiation of eukaryotic DNA replication requires many proteins that interact with one another and with replicators. Using a yeast genetic screen, we have identified Noc3p (nucleolar complex-associated protein) as a novel replication initiation protein. Noc3p interacts with MCM proteins and ORC and binds to chromatin and replicators throughout the cell cycle. It functions as a critical link between ORC and other initiation proteins to effect chromatin association of Cdc6p and MCM proteins for the establishment and maintenance of prereplication complexes. Noc3p is highly conserved in eukaryotes and is the first identified bHLH (basic helix-loop-helix) protein required for replication initiation. As Noc3p is also required for pre-rRNA processing, Noc3p is a multifunctional protein that plays essential roles in two vital cellular processes. PMID- 12110183 TI - The dsRNA binding protein RDE-4 interacts with RDE-1, DCR-1, and a DExH-box helicase to direct RNAi in C. elegans. AB - Double-stranded (ds) RNA induces potent gene silencing, termed RNA interference (RNAi). At an early step in RNAi, an RNaseIII-related enzyme, Dicer (DCR-1), processes long-trigger dsRNA into small interfering RNAs (siRNAs). DCR-1 is also required for processing endogenous regulatory RNAs called miRNAs, but how DCR-1 recognizes its endogenous and foreign substrates is not yet understood. Here we show that the C. elegans RNAi pathway gene, rde-4, encodes a dsRNA binding protein that interacts during RNAi with RNA identical to the trigger dsRNA. RDE-4 protein also interacts in vivo with DCR-1, RDE-1, and a conserved DExH-box helicase. Our findings suggest a model in which RDE-4 and RDE-1 function together to detect and retain foreign dsRNA and to present this dsRNA to DCR-1 for processing. PMID- 12110184 TI - Rac1 and Cdc42 capture microtubules through IQGAP1 and CLIP-170. AB - Linkage of microtubules to special cortical regions is essential for cell polarization. CLIP-170 binds to the growing ends of microtubules and plays pivotal roles in orientation. We have found that IQGAP1, an effector of Rac1 and Cdc42, interacts with CLIP-170. In Vero fibroblasts, IQGAP1 localizes at the polarized leading edge. Expression of carboxy-terminal fragment of IQGAP1, which includes the CLIP-170 binding region, delocalizes GFP-CLIP-170 from the tips of microtubules and alters the microtubule array. Activated Rac1/Cdc42, IQGAP1, and CLIP-170 form a tripartite complex. Furthermore, expression of an IQGAP1 mutant defective in Rac1/Cdc42 binding induces multiple leading edges. These results indicate that Rac1/Cdc42 marks special cortical spots where the IQGAP1 and CLIP 170 complex is targeted, leading to a polarized microtubule array and cell polarization. PMID- 12110185 TI - MICALs, a family of conserved flavoprotein oxidoreductases, function in plexin mediated axonal repulsion. AB - Members of the semaphorin family of secreted and transmembrane proteins utilize plexins as neuronal receptors to signal repulsive axon guidance. It remains unknown how plexin proteins are directly linked to the regulation of cytoskeletal dynamics. Here, we show that Drosophila MICAL, a large, multidomain, cytosolic protein expressed in axons, interacts with the neuronal plexin A (PlexA) receptor and is required for Semaphorin 1a (Sema-1a)-PlexA-mediated repulsive axon guidance. In addition to containing several domains known to interact with cytoskeletal components, MICAL has a flavoprotein monooxygenase domain, the integrity of which is required for Sema-1a-PlexA repulsive axon guidance. Vertebrate orthologs of Drosophila MICAL are neuronally expressed and also interact with vertebrate plexins, and monooxygenase inhibitors abrogate semaphorin-mediated axonal repulsion. These results suggest a novel role for oxidoreductases in repulsive neuronal guidance. PMID- 12110186 TI - Recruitment of Nck by CD3 epsilon reveals a ligand-induced conformational change essential for T cell receptor signaling and synapse formation. AB - How membrane receptors initiate signal transduction upon ligand binding is a matter of intense scrutiny. The T cell receptor complex (TCR-CD3) is composed of TCR alpha/beta ligand binding subunits bound to the CD3 subunits responsible for signal transduction. Although it has long been speculated that TCR-CD3 may undergo a conformational change, confirmation is still lacking. We present strong evidence that ligand engagement of TCR-CD3 induces a conformational change that exposes a proline-rich sequence in CD3 epsilon and results in recruitment of the adaptor protein Nck. This occurs earlier than and independently of tyrosine kinase activation. Finally, by interfering with Nck-CD3 epsilon association in vivo, we demonstrate that TCR-CD3 recruitment of Nck is critical for maturation of the immune synapse and for T cell activation. PMID- 12110187 TI - Bacterial adhesion to target cells enhanced by shear force. AB - Surface adhesion of bacteria generally occurs in the presence of shear stress, and the lifetime of receptor bonds is expected to be shortened in the presence of external force. However, by using Escherichia coli expressing the lectin-like adhesin FimH and guinea pig erythrocytes in flow chamber experiments, we show that bacterial attachment to target cells switches from loose to firm upon a 10 fold increase in shear stress applied. Steered molecular dynamics simulations of tertiary structure of the FimH receptor binding domain and subsequent site directed mutagenesis studies indicate that shear-enhancement of the FimH-receptor interactions involves extension of the interdomain linker chain under mechanical force. The ability of FimH to function as a force sensor provides a molecular mechanism for discrimination between surface-exposed and soluble receptor molecules. PMID- 12110189 TI - A highly efficient synthesis of an octasaccharide, the repeating unit of the cell wall mannan of Trichophyton mentagrophytes and T. rubrum. AB - A highly concise and effective synthesis of the mannose octasaccharide repeating unit of the cell-wall mannan of Trichophyton mentagrophytes and T. rubrum was achieved via 6-O-glycosylation of a tetrasaccharide acceptor with a tetrasaccharide donor, followed by deprotection. The key tetrasaccharide (11) was constructed by selective 6-O-glycosylation of allyl 3,4-di-O-benzoyl-alpha-D mannopyranosyl-(1-->6)-2,3,4-tri-O-benzoyl-alpha-D-mannopyranoside with 6-O acetyl-2,3,4-tri-O-benzoyl-alpha-D-mannopyranosyl trichloroacetimidate, then with 2,3,4,6-tetra-O-benzoyl-alpha-D-mannopyranosyl trichloroacetimidate. The tetrasaccharide acceptor (13) was obtained by selective 6-O-deacetylation of 11, while the tetrasaccharide donor 12 was obtained by deallylation of 11, followed by trichloroacetimidation. PMID- 12110190 TI - Unexpected alpha-stereochemical outcomes of attempted beta-glycosylations. AB - In an effort to prepare complex oligosaccharide derivatives, a series of unexpected alpha glycosides were predominantly formed in the presence of neighboring group participation using imidates or thioglycosides as glycosyl donors under standard glycosylation conditions. The observations are especially suitable in the case of alpha-(1-->3) glycosidic bond formation. PMID- 12110191 TI - Convenient syntheses of symmetrical and unsymmetrical glycosyl carbodiimides and N,N-bis(glycosyl)cyanamides. AB - Reaction of glycosyl trimethylphosphinimides with carbon disulfide under mild conditions (room temperature, short reaction time) leads to symmetrical glycosyl carbodiimides. Addition of bis(trimethylsilyl)carbodiimide to peracetylated aldoses under the influence of SnCl(4) afforded N,N-bis(glycosyl)cyanamides for the first time. Readily accessible unsymmetrical N,N'-bis(glycosyl)thioureas can be desulfurated and transformed into the corresponding carbodiimides using HgO in CHCl(3)/water at room temperature. PMID- 12110192 TI - Effect of fatty acid chain length on initial reaction rates and regioselectivity of lipase-catalysed esterification of disaccharides. AB - In a reaction medium mixture of 9:11 t-BuOH and pyridine (v/v) the effect of fatty acid chain length (C-4-C-12) on C. antarctica lipase B (Novozym 435, EC 3.1.1.3) catalysed esterification was studied. alpha and beta maltose 6'-O-acyl esters in an anomeric molar ratio of 1.0:1.1 were synthesised independently of the chain length, but the initial specific reaction rate increased with decreasing chain length of the acyl donor. The product yield followed the same trend with a lauryl ester yield of 1.1% (mol/mol) and a butyl ester yield of 27.6% (mol/mol) after 24 h of reaction. With sucrose as the acyl acceptor the 6' O-acyl and 6-O-acyl monoesters were formed with fatty acids of chain length C-4 and C-10 while the 6',6-O-acyl diester was formed only with butanoic acid (C-4:0) as acyl donor. The 6'-O-acyl and 6-O-acyl monoesters and the 6',6-O-acyl diester of butanoic acid were produced in a molar ratio of 1.0:0.5:0.2 and with decanoic acid (C-10:0) the 6'-O-acyl and 6-O-acyl monoesters were formed in the ratio of 1.0:0.3. The highest initial reaction rate and yield were obtained with the shortest chain length of the acyl donor. Initial reaction rates and ester yields were affected by the solubility of the disaccharide, with higher reaction rates and yields with maltose than with sucrose, while no formation of esters were observed with either cellobiose or lactose as acyl acceptors. PMID- 12110193 TI - Chemical formation of 4-hydroxy-2,5-dimethyl-3[2H]-furanone from D-fructose 1,6 diphosphate. AB - The selective chemical formation of 4-hydroxy-2,5-dimethyl-3[2H]-furanone (HDF) from D-fructose 1,6-diphosphate in the presence of reduced nicotinamide-adenine dinucleotides (NAD(P)H) was investigated by means of HPLC-DAD and HPLC-UV-MS/MS. The temperature optimum for HDF formation was 30 degrees C, whereas the pH value (pH 3-10) and chemical nature of the buffer had no significant influence. A linear correlation of reaction time and D-fructose 1,6-diphosphate concentration with the obtained HDF yield was observed. Proteins appeared to have a stabilizing effect. The NAD(P)H were mandatory, even in the presence of protein, implying a non-enzymatic hydride-transfer to an unknown intermediate which finally leads to the selective formation of HDF. The hydride-transfer was confirmed by the application of selectively pro-4R or pro-4S deuterium labeled NADH resulting in each case in the formation of HDF exhibiting a deuterium labeling of approx 30% and employment of [4R,S-(2)H(2)]-NADH led to a deuterium labeling of approx 66%. The incubation of [1-(13)C]-D-fructose 1,6-diphosphate with [4R,S-(2)H(2)]-NADH revealed that the hydride is transferred to C-5 or C-6 of the D-fructose 1,6 diphosphate skeleton. Thus, a chemical HDF formation from D-fructose 1,6 diphosphate under physiological reaction conditions was shown and for the first time to our knowledge a non-enzymatic hydride-transfer from NADH to a carbohydrate structure was demonstrated. PMID- 12110194 TI - Novel lipochitin oligosaccharide structures produced by Rhizobium etli KIM5s. AB - The novel lipochitin oligosaccharide (LCOs) structures produced by Rhizobium etli KIM5s were characterized using a nanoHPLC reverse-phase system coupled to an ion trap mass spectrometer. This technique was shown to be more sensitive for structural elucidation of LCOs than previously used mass spectrometric methods. The structures of the LCOs of R. etli KIM5s, the majority containing six monosaccharide residues, differed from those synthesized by all other rhizobia analyzed to date. In addition, novel structures in which the chitin backbone was deacetylated at one or more GlcNAc moieties were found as minor compounds. The difference in host range of this strain compared to that of other known bean microsymbionts is discussed. PMID- 12110195 TI - New exopolysaccharides produced by Aureobasidium pullulans grown on glucosamine. AB - The polysaccharides produced by Aureobasidium pullulans, grown using glucosamine as the carbon source, were investigated by means of methylation analysis, affinity chromatography and NMR spectroscopy. The results indicated that, besides a small amount of pullulan, this micro-organism was capable of producing-in low yields-mixtures of at least two different complex polysaccharides containing mainly mannose and galactose. (1)H NMR spectra of two fractions obtained by lectin affinity chromatography indicated that one polymer was constituted exclusively of mannose residues while the other contained both galactofuranosyl and mannopyranosyl residues. PMID- 12110196 TI - Examination of molar-based distribution of A, B and C chains of amylopectin by fluorescent labeling with 2-aminopyridine. AB - A method for determination of a molar-based distribution of A, B and C chains of amylopectin was developed. Labeling with fluorescent 2-aminopyridine was proportional to the number-average degree of polymerization (dp(n)) of the chains in the range of 6-440. Number-average chain lengths (cl(n)) of amylopectins from six different plant sources (rice, maize, wheat, potato, sweet potato and yam) determined by the labeling method were in good agreement with values obtained by determination of non-reducing residues. The molar-based distributions were polymodal (A, B(1) and B(2)+B(3) fractions) and characteristic to botanical sources. Amylopectins from starches with A-crystalline type had higher amount of A+B(1) chains (90-93% by mole) than starches with B-type (68-87%). Molar ratios of (A+B(1))/(B(2)+B(3)) were 8.9-12.9 for the A-type starches and 2.1-6.5 for the B-type starches, suggesting that amylopectins of A-type starches had 1.5-2 times more branches per cluster than B-type. The distributions of C chains, except for amylomaize, showed a broad, asymmetrical profile from dp approximately 10 to approximately 130 with a peak at dp approximately 40 and were very similar among botanical sources, suggesting that the biosynthetic process for C chains is similar in different plant species. PMID- 12110197 TI - Application of mass spectrometry to determine the activity and specificity of pectin lyase A. AB - Electrospray ionization (ESI) with quadrupole ion-trap mass spectrometry was used to assess the activity and specificity of the enzyme pectin lyase A (PLA) (EC 4.2.2.10) on model pectins with varying degrees and patterns of methyl esterification. PLA is a pectinase which cleaves alpha-(1-->4)-glycosidic linkages in pectin by a trans-elimination process. Using pectins with different degrees and patterns of methyl esterification, there was a significant variation in the activity rate of PLA. The enzymatic products generated at various time intervals were structurally analyzed by mass spectrometry to determine the specificity of PLA. Although the preferred substrate for PLA is fully methyl esterified polygalacturonate, cleavage was also observed with a non-methyl esterified galacturonic acid residue on either the non-reducing end or the reducing end. The current study shows that although PLA prefers fully methyl esterified substrates it can also accept partially esterified ones. It also demonstrates the suitability of ESI ion-trap mass spectrometry in determining enzyme specificities. PMID- 12110198 TI - Structure of the complex of heptakis(2,6-di-O-methyl)-beta-cyclodextrin with (2,4 dichlorophenoxy)acetic acid. AB - The structure of the complex formed by heptakis(2,6-di-O-methyl)-beta cyclodextrin and (2,4-dichlorophenoxy)acetic acid was studied by X-ray diffraction. The dichlorophenyl moiety of the guest molecule was found outside the host hydrophobic cavity in the primary methoxy groups region whereas the oxyacetic acid chain penetrates the cavity from the primary face. The host molecules stacks along the a crystal axis forming a column. In the space between three successive hosts of the column, a guest molecule is accommodated. PMID- 12110199 TI - Syntheses of the 4-nitrophenyl glycosides of hyalobiuronic acid and chondrosine. AB - 4-Nitrophenyl [sodium beta-D-glucopyranosyluronate]-(1-->3)-2-acetamido-2-deoxy beta-D-glucopyranoside (1) and 4-nitrophenyl [sodium beta-D glucopyranosyluronate]-(1-->3)-2-acetamido-2-deoxy-beta-D-galactopyranoside (2) were prepared from the zwitterions hyalobiuronic acid [beta-D-glucopyranuronic acid-(1-->3)-2-amino-2-deoxy-D-glucopyranose] and chondrosine [beta-D glucopyranuronic acid-(1-->3)-2-amino-2-deoxy-D-galactopyranose], respectively. Compounds 1 and 2 were not hydrolysed by bovine testicular hyaluronidase. PMID- 12110200 TI - Anticoagulant activity of a sulfated chitosan. AB - Chitin prepared from the shells of rice-field crabs (Somanniathelphusa dugasti) was converted into chitosan with a degree of acetylation of 0.21 and then sulfated with chlorosulfonic acid in N,N-dimethylformamide under semi heterogeneous conditions to give 87% of water-soluble sulfated chitosan with degree of substitution (d.s) of 2.13. 1H NMR revealed the sulfate substitution at C-2, C-3 and C-6. Gel filtration on Sepharose CL-6B of the sulfated chitosan gave three fractions with average molecular weights of 7.1, 3.5, and 1.9 x 10(4). The three sulfated chitosan preparations showed strong anticoagulant activities, with the same mechanism of action observed for standard therapeutic heparin. PMID- 12110201 TI - Can MMTV exploit TLR4? AB - The recognition of microbial pathogens based on their molecular patterns is essential for host defense. Recently, Toll-like receptors have been shown not only to recognize viruses as well as bacteria and fungi, but also to trigger an efficient immune response. A recent publication proposed that the retrovirus mouse mammary tumor virus exploits the pattern-recognition receptor Toll-like receptor 4 to achieve more efficient infection. PMID- 12110203 TI - Viral suppression of systemic silencing. AB - RNA silencing in plants is a form of antiviral defense that was originally discovered from the anomalous effects of transgenes. The process is associated with a systemic signal, presumed to be RNA, and is suppressed by plant virus encoded proteins. One of these proteins, the 2b protein of cucumber mosaic virus, prevents systemic spread of the signal molecule but, curiously, is located in the nucleus of infected cells. The antiviral role of silencing might also apply in animals because a suppressor of silencing encoded by an insect virus was identified recently. PMID- 12110204 TI - Genome giants. PMID- 12110208 TI - Recombinant HIV-1 spreads. PMID- 12110209 TI - Virulence and pathogenesis. AB - Why do viruses cause disease? As intracellular parasites they grow at the expense of the host, yet many infections are non-virulent. We tend to focus on unusual outcomes of infection that are important to the individual but trivial for host parasite evolution, for example, paralytic polio or viral cancer. The assumption that the features of disease help onward transmission of the virus is true for, say, rabies, but not for AIDS or neurodegenerative diseases. Moreover, minor host differences can result in major changes in pathogen virulence. Although viral burden relates to disease severity, pathogenesis is not necessarily coupled with transmission dynamics. PMID- 12110210 TI - Herpesvirus genetics has come of age. AB - The genetic analysis of the large and complex herpesviruses has been a constant challenge to herpesvirologists. Elegant methods have been developed to produce mutants in infected cells that rely on the cellular recombination machinery. Bacterial artificial chromosomes (BACs), single copy F-factor-based plasmid vectors of intermediate insert capacity, have now enabled the cloning of complete herpesvirus genomes. Infectious virus genomes can be shuttled between Escherichia coli and eukaryotic cells. Herpesvirus BAC DNA engineering in E. coli by homologous recombination requires neither restriction sites nor cloning steps and allows the introduction of a wide variety of DNA modifications. Such E. coli based technology has provided a safe, fast and effective approach to the systematic mining of the information stored in herpesvirus genomes as a result of their intimate co-evolution with their specific hosts for millions of years. Use of this technique could lead to new developments in clinical virology and basic virology research, and increase the usage of viral genomes as investigative tools and vectors. PMID- 12110211 TI - Picornavirus-receptor interactions. AB - Many picornaviruses use cell-surface molecules belonging to the immunoglobulin superfamily (IgSF) as their cellular receptors. These molecules usually consist of tandem repeats of between two and five Ig-like domains whose amino-terminal domains (D1) interact with invading viruses, with their carboxy-terminal sections comprising a transmembrane and a short cytoplasmic region. Most rhino- and enteroviruses, belonging to the Picornavirus family, use a canyon-like feature on their surface to attach to cellular receptors. Binding into the canyon destabilizes the virus and thus initiates the uncoating process. By contrast, non IgSF molecules, when used by picornaviruses as receptors, bind outside the canyon and do not cause viral instability. PMID- 12110212 TI - Immunomodulation by cytomegaloviruses: manipulative strategies beyond evasion. AB - Human cytomegalovirus (CMV) remains the major infectious cause of birth defects as well as an important opportunistic pathogen. Individuals infected with CMV mount a strong immune response that suppresses persistent viral replication and maintains life-long latency. Loss of immune control opens the way to virus reactivation and disease. The large number of immunomodulatory functions encoded by CMV increases the efficiency of infection, dissemination, reactivation and persistent infection in hosts with intact immune systems and could contribute to virulence in immunocompromised hosts. These functions modulate both the innate and adaptive arms of the immune response and appear to target cellular rather than humoral responses preferentially. CMV encodes a diverse arsenal of proteins focused on altering and/or mimicking: (1) classical and non-classical major histocompatibility complex (MHC) protein function; (2) leukocyte migration, activation and cytokine responses; and (3) host cell susceptibility to apoptosis. Evidence that the host evolves mechanisms to counteract virus immune modulation is also accumulating. Although immune evasion is certainly one clear goal of the virus, the pro-inflammatory impact of certain viral functions suggests that increased inflammation benefits viral dissemination. The ability of such viral functions to successfully 'face off' against the host immune system ensures the success of this pathogen in the human population and could provide key insights into disease mechanisms. PMID- 12110213 TI - The continued pandemic threat posed by avian influenza viruses in Hong Kong. AB - In 1997, a highly pathogenic avian H5N1 influenza virus was transmitted directly from live commercial poultry to humans in Hong Kong. Of the 18 people infected, six died. The molecular basis for the high virulence of this virus in mice was found to involve an amino acid change in the PB2 protein. To eliminate the source of the pathogenic virus, all birds in the Hong Kong markets were slaughtered. In 1999, another avian influenza virus of H9N2 subtype was transmitted to two children in Hong Kong. In 2000-2002, H5N1 avian viruses reappeared in the poultry markets of Hong Kong, although they have not infected humans. Continued circulation of H5N1 and other avian viruses in Hong Kong raises the possibility of future human influenza outbreaks. Moreover, the acquisition of properties of human viruses by the avian viruses currently circulating in southeast China might result in a pandemic. PMID- 12110215 TI - Institute profile: the Uganda Virus Research Institute. PMID- 12110214 TI - Institute profile: Institute of Virology, Glasgow. PMID- 12110216 TI - Counseling molecular diagnostics. PMID- 12110217 TI - Glyxins to treat neurological disorders. PMID- 12110218 TI - Peptide provides three-in-one protection. PMID- 12110219 TI - Tailoring vaccines to individual lymphomas. PMID- 12110222 TI - The commercialization of biotechnology in Japan. PMID- 12110223 TI - 'This is your captain speaking'. PMID- 12110224 TI - The use of imaging to accelerate drug development. PMID- 12110225 TI - High throughput in drug discovery. PMID- 12110227 TI - Probes for chemical genomics by design. AB - Chemical genomics represents a convergence of biology and chemistry in the era of global approaches to target identification and intervention. The success of genomics has led to a bottleneck in target validation that could be overcome by using small diverse organic compounds to interfere with biological processes. Because of the limitations of existing compound collections, this diversity can only fully be exploited using in silico design techniques to guide the selection of molecules with optimal binding properties. Structure-based design is used to create structures de novo that can be synthesized for use as chemical probes and drug leads. PMID- 12110228 TI - Virus-based vectors for human vaccine applications. AB - Vaccinology has experienced a dramatic resurgence recently, as traditional methodologies of using attenuated live pathogens or inactivated whole pathogens have been either ineffective or are not an acceptable risk for several disease targets, including HIV and Hepatitis C. Gene-based vaccines can stimulate potent humoral and cellular immune responses, and viral vectors might be an efficient strategy for both delivery of antigen-encoding genes, as well as facilitating and enhancing antigen presentation. Vectors derived from diverse viruses with distinct tropism and gene expression strategies have been developed, and are being evaluated in preclinical and clinical vaccine studies. Virus-based vaccines represent a promising approach for vaccines against infectious and malignant disease. PMID- 12110232 TI - Bioinformatics: a holistic approach to drug discovery. PMID- 12110229 TI - Toxicogenomics: a new revolution in drug safety. AB - New drugs are screened for adverse reactions using a laborious, costly process and still some promising therapeutics are withdrawn from the marketplace because of unforeseen human toxicity. Novel higher throughput methods in toxicology need to be developed. These new approaches should provide more insight into potential human toxicity than current methods. Toxicogenomics, the examination of changes in gene expression following exposure to a toxicant, offers the potential to identify a human toxicant earlier in drug development and to detect human specific toxicants that cause no adverse reaction in rats. PMID- 12110233 TI - Starving cancer cells. PMID- 12110234 TI - Chip-based diagnostics for meningitis and food poisoning. PMID- 12110235 TI - Bacteria prove gutsy against inflammatory bowel disease. PMID- 12110238 TI - Sir Richard Sykes contemplates the future of the pharma industry. Interview by Rebecca N Lawrence. PMID- 12110239 TI - MCRs: changing the chemists' mindset. PMID- 12110240 TI - Is X-ray crystallography a viable tool for lead discovery? PMID- 12110241 TI - Drug discovery technology for ion channels. PMID- 12110242 TI - NF-kappa B: arresting a major culprit in cancer. AB - Nuclear factor-kappa B (NF-kappa B) has long been known to play a central role in the immune system by regulating the expression of key genes. Moreover, activation of this transcription factor helps a wide variety of cell types survive damage induced by pro-apoptotic stimuli. Because of its crucial role in the regulation of pro-inflammatory genes, NF-kappa B is a promising target for the discovery of anti-inflammatory drugs. More recently, NF-kappa B has also emerged as a major culprit in a variety of human cancers mainly because of its ability to protect transformed cells from apoptosis. The pharmaceutical industry should, therefore, seriously consider testing inhibitors of NF-kappa B, identified as part of their anti-inflammatory drug discovery programs, in combination with other chemotherapeutic drugs in models of cancer. PMID- 12110243 TI - Therapeutic vaccines: realities of today and hopes for the future. AB - Vaccines are by definition prophylactic, but in recent years an interest has developed in therapeutic vaccines for infectious diseases such as AIDS and tuberculosis, as well as gastric ulcers, cancer (with different approaches to combat various types of malignancy) and autoimmune diseases (a definite success was the development of a vaccine against multiple sclerosis) and there are potential vaccines in development for myasthenia gravis, lupus and diabetes. Therapeutic vaccines are also being developed against cognitive diseases such as Alzheimer's disease, prion diseases and Huntington's disease. All of these efforts are based on the therapeutic vaccine being closely related chemically to the etiological agent that causes the disease. PMID- 12110244 TI - Neural transplantation for treatment of Parkinson's disease. AB - Neural transplantation has emerged as an efficacious experimental treatment for CNS disorders, especially Parkinson's disease. However, logistical and ethical issues impede large-scale clinical trials. To this end, alternatives to human fetal cells as donor cell grafts have been examined, including xenografts, stem cells, genetically engineered cells, immortalized cell lines, or paraneural cells that secrete specific neurotrophic or growth factors. Accumulating evidence also suggests that exogenous treatment with neurotrophic or growth factors, immunosuppressants, free radical scavengers, and anti-apoptotic agents can enhance survival and functional effects of the grafts. This article will review recent studies demonstrating the potential of these alternative cell graft sources and novel drugs for treating Parkinson's disease. PMID- 12110247 TI - World Health Organization, 50 years of influenza surveillance: a challenge for the 21st century. Meeting 17-19 February 1999, WHO, Geneva. PMID- 12110249 TI - Influenza pandemics: lessons for the future. PMID- 12110248 TI - Influenza and the work of the World Health Organization. AB - Before World War I, influenza was not considered a particularly serious problem. The great pandemic of 1918-1919 changed all that, and the possibility that such a catastrophe could occur again has conditioned all subsequent developments. In epidemiological terms, the hallmark of an influenza is the excess mortality that it causes combined with an enormous burden of ill-health that saps the energy of individuals, families and communities throughout the whole world. In order to engage in influenza prevention and control, the global influenza surveillance network was set up by World Health Organization (WHO) in 1948 as a worldwide alert system for the identification of new influenza viruses, gathering information from 110 participating laboratories in 82 countries and four WHO Collaborating Centers for Influenza reference and research: Centers for Disease Control and Prevention, Atlanta (USA), National Institute for Medical Research, London (UK), WHO Collaborating Centre for Influenza Reference and Research, Melbourne (Australia) and the National Institute for Infectious Diseases, Tokyo (Japan). This network helps WHO to monitor influenza activity all over the world and provides the organization with the viral isolates and information it requires to decide which new virus strains will be used to produce influenza vaccines during the following season. Each year, information about the isolates over the previous 12 months is analyzed and used to determine the composition of the influenza vaccine to be administered during the coming influenza season both for the northern and southern hemisphere. If necessary, the recommendations for the southern hemisphere differ from the ones formulated for the northern hemisphere vaccine. The information supplied by this network enables the organization to regularly update its World Wide Web (WWW) site (FluNet), which reports on the situation of diseases. This network will also enable the WHO to detect a new influenza pandemic as early as possible. PMID- 12110250 TI - The importance of animal influenza for human disease. AB - Influenza is a zoonotic disease caused by a constantly varying RNA virus resulting in a need for continuous surveillance to update human vaccines. Our knowledge indicates that the intermittent pandemics of influenza originate from influenza viruses or gene segments from influenza viruses in lower animals and birds. These pandemics can be mild to catastrophic. While we have learned a great deal about the ecology and molecular properties of "animal" influenza viruses, we do not have a system for comprehensive international surveillance. The 1918 Spanish influenza pandemic that originated from lower animals and the recent H5N1 bird flu incident in Hong Kong serves to remind us that influenza is an emerging disease. The challenge for the 21st century is to accumulate the necessary epidemiological data on animal influenza viruses so that an international surveillance system can be devised. This epidemiological data may provide clues on how to reduce interspecies transmission of influenza. The separation of aquatic birds from other "land based" domestic poultry in Hong Kong after the H5N1 bird flu incident indicates that animal husbandry practices could influence the interspecies transmission of influenza viruses. PMID- 12110251 TI - Cost effectiveness of influenza vaccination for healthy persons between ages 65 and 74 years. AB - Healthy persons between 65 and 74 years of age represent a large proportion of the population in this age group. Internationally, there is a substantial variation in whether these people are included among the recommendations for routine influenza vaccination. We therefore conducted this study, updating an earlier analysis, to assess the health and economic benefits of routine influenza vaccination of healthy persons between 65 and 74 years of age. The health benefits associated with vaccination were estimated using the administrative data bases of a large HMO in the Minneapolis, St. Paul, Minnesota area. Multivariate models were used to estimate reductions in hospitalization and death associated with vaccination. The economic analysis took the societal perspective and presented the results as net cost or saving per 10,000 persons vaccinated and per death prevented. Direct and indirect monetary costs were included in the models and were estimated from the published literature. Monte Carlo simulation was used to conduct probabilistic sensitivity analysis in order to derive probability intervals for each estimate of net costs or savings. Over the six consecutive study seasons, 1990-1991 to 1995-1996, vaccination of healthy elderly person was associated with a 36% reduction in hospitalization for pneumonia or influenza (95% CI, 2-39%), an 18% reduction in hospitalization for all respiratory conditions (95% CI, -6 to 37%) and a 40% reduction in death (95% CI, 14-38%). Vaccination was also associated with cost savings in all scenarios evaluated. The findings of this study again affirm the value of an age-based strategy for routine influenza vaccination of all elderly persons including healthy elderly persons between 65 and 74 years. PMID- 12110252 TI - Perspectives on influenza surveillance in Europe: European scientific working group on influenza, Brussels. AB - Although influenza surveillance has reached a high level of sensitivity, it is not given the same priority in all European countries. A recent survey has shown the diversity of the surveillance and diagnostic methods. Suggestions are made to encourage homogenisation of techniques, assistance to less active laboratories, training of personnel, evaluation, and co-operation in the field of animal influenza. PMID- 12110253 TI - Influenza activity in China: 1998-1999. AB - During 1989-1999, influenza A H3N2 and H1N1 subtypes and B type viruses were still co-circulating in human population in China, while influenza A (H3N2) virus was predominant strain. The two antigenically and genetically distinguishable strains of influenza B virus were also still co-circulating in men in southern China. The antigenic analysis indicated that most of the H3N2 viruses were A/Panama/2007/99 (H3N2)-like strain, the most of the H1N1 viruses were antigenically similar to A/Beijing/262/95 (H1N1) virus. However, most of the influenza B viruses were B/Beijing/184/93-like strain, but few of them were antigenically similar to B/Shandong/7/97 virus. In the summer of 1998, the influenza outbreaks caused by H3N2 subtype of influenza A virus occurred widely in southern China. Afterwards, during 1998-1999 influenza season, a severe influenza epidemic caused by H3N2 virus emerged in northern China. The morbidity was reached as high as 10% in Beijing area. It was interesting that during influenza, surveillance from 1998 to 1999, five strains of avian influenza A (H9N2) virus were isolated from outpatients with influenza-like illness in July August of 1998, and another one was repeatedly isolated from a child suffering from influenza-like disease in November of 1999 in Guangdong province. The genetic analysis revealed that the five strains isolated in 1998 were genetically closely related to H9N2 viruses being isolated from chickens (G9 lineage virus), whereas, A/Guangzhou/333/99 (H9N2) virus was a reassortant derived from reassortment between G9 and G1 lineage of avian influenza A (H9N2) viruses due to its genes encoding the HA, NA, NP and NS proteins, closely related to G9 lineage virus, the rest of the genes encoding the M and three polymerase (PB2, PB1 and PA) were closely related to G1 lineage strain of H9N2 virus. However, no avian influenza A (H5N1) virus has so far been isolated neither from in or outpatients with influenza-like disease in mainland China. Unfortunately, where did the reassortment occur and how did the reassortant transmit to men? These questions are still unknown. PMID- 12110254 TI - Perspectives on the characteristics and achievements of recent surveillance of influenza activity in Japan. PMID- 12110255 TI - Regional perspectives on influenza surveillance in North Africa (PISNA). PMID- 12110256 TI - Regional perspectives on influenza surveillance in Africa. AB - Africa possesses little capacity for influenza surveillance-Senegal and South Africa being the only countries in the WHO African region who regularly pursue active surveillance and characterize influenza isolates. South Africa has three sites-in Cape Town, Durban and the largest in Johannesburg at the National Institute for Virology (NIV). The NIV antigenically and molecularly characterizes influenza viruses isolated from specimens provided by a sentinel network of approximately 50 clinical sites. This information, together with the isolates themselves are supplied to WHO International Influenza Centres in London and Melbourne. In addition, proxy markers of influenza severity such as school absenteeism and doctor/clinic visits are monitored to assess the severity of epidemics. Although, influenza exacts a heavier toll of the illness burden in developing countries already beset with underlying chronic medical conditions and also has a more severe impact on economies largely dependent on single income earners and subsistence farmers, influenza surveillance and vaccination awareness is woefully lacking on the African continent, and this urgently needs to be remedied. PMID- 12110257 TI - Regional perspectives on influenza surveillance in South America. AB - The implementation of influenza vaccination programs emphasize the necessity of an extended influenza surveillance in the countries of the region. As it is based on the activity of National Influenza Centers we intend to make a description of their work, their historical background and further development. Technical capacities in influenza South America laboratories, and networks in Argentina, Brazil and Chile are shown. Examples of different viral circulation in a unique country or in countries with common borders illustrate the importance of the information obtained by means of the virological surveillance. Specific characteristics of the region as long distances and the lack of modern information systems require a suitable fitting of the systems that are working in Northern Hemisphere countries. The contribution of motivated physicians and public health workers must not to be disregarded. The following actions are proposed: optimizing technical capacities of National Influenza Centers in order to improve the quality of data available; improving the communication of the information obtained by surveillance activities to all the participants; increasing the cooperation among the countries; motivate health authorities to become aware of influenza impact in public health. PMID- 12110258 TI - Influenza haemagglutinin. AB - Studies are described of the processing and structural modifications of haemagglutinin required for its membrane fusion activity. PMID- 12110259 TI - Neuraminidase inhibitors as antivirals. AB - The description of the three-dimensional structure of the influenza virus neuraminidase in 1983 opened a new phase in the search for inhibitors of the enzyme. Two compounds in late development (February 1999) have similar levels of potency against influenza A viruses, different routes of administration and, surprisingly, different resistance profiles both in vitro and in vivo. PMID- 12110260 TI - Quantification of neuramidase (NA) protein content. AB - An ELISA-immunocapture assay was developed for quantification of influenza neuraminidases (N2 and B), with a sensitivity about 10 ng neuramidase(NA)/ml. This sensitive and specific assay was applied to several trivalent vaccine preparations (1997/1998 and 1998/1999). It was shown capable to detect 3.5-6.9 microg NA/ml according to the influenza strain and the vaccine origin. PMID- 12110261 TI - Reverse genetics. PMID- 12110262 TI - Preparing for pandemic influenza: the need for enhanced surveillance. AB - In the US, planning for the next influenza pandemic is occurring in parallel at the national, state and local levels. Certain issues, such as conducting surveillance and purchasing pandemic vaccine, require co-ordination at the national level. However, most prevention and control actions will be implemented at the state and local levels, which vary widely in terms of population demographics, culture (e.g. rural versus urban) and available resources. In 1995, a survey by the Council of State and Territorial Epidemiologists (CSTE) found that only 29 (59%) states perceived a need to develop a specific influenza pandemic plan for their jurisdiction. Since then, the process of developing state and local plans has gained considerable momentum. Integration of these efforts with the national planning process has been facilitated by: (1) the mutual involvement of state and federal staff in both processes; (2) the sharing of draft documents; (3) the ongoing occurrence of local and national co-ordinating meetings; (4) the provision of financial resources by the federal government. So far, approximately 12 states either have drafted or begun drafting a state and local influenza pandemic plan. One of the benefits of the collaborative planning process has been the development of new working relationships and partnerships among several agencies at the state, local and national levels. Such efforts will improve our collective ability to rapidly investigate and control other emerging or re-emerging public health threats in the 21st century, be it a bioterrorist event, pandemic influenza, or any other catastrophic health event. PMID- 12110263 TI - The role of live influenza vaccines in children. AB - Live attenuated cold-adapted influenza vaccines (CAIVs) have been developed over the past two decades by taking advantage of the segmented RNA genome of influenza and creating attenuated reassortants containing contemporary hemagglutinin (HA) and neuraminidase (NA) genes. These vaccines have been shown to be easily administered, safe and immunogenic in adults and children. Recent trials of a trivalent live attenuated CAIV (CAIV-T, tradename FluMist, Aviron, Mt. View, CA) in children have demonstrated greater than 85% efficacy against culture positive H3N2 and B influenza illness and complications, such as otitis media. CAIV-T also prevented shedding of H1N1 virus in 83% of vaccinated subjects after a monovalent CAIV challenge. Nasal IgA and serum HA inhibition (HAI) antibody produced by these vaccines have been associated with protection against infection, but protection may exist even in the absence of identifiable antibody response. Work to date documenting phenotypic and genetic stability, low likelihood of reactogenicity, infrequent transmissibility and attenuating properties of reassortants heralds promise for the broad use of this vaccine. Targeting children to receive this vaccine may now prove practical and may serve to reduce overall influenza morbidity, given the significant contribution of the pediatric age group of children to influenza illness burden and community spread. Studies of vaccine use in community settings will aid in determining the public health future of this approach. PMID- 12110264 TI - Influenza DNA vaccines. AB - DNA vaccines have been the subjects of much effort over the past decade due to their ability to induce broad-based immune responses and protection in various animal models of infectious and non-infectious diseases. In particular, influenza DNA vaccines have been well studied. This brief review highlights some of this early work that helped establish the DNA vaccine technology as a potential new mode of vaccination. PMID- 12110265 TI - Influenza A (H5N1) in Hong Kong: an overview. AB - Worldwide pandemics of human influenza virus caused extensive morbidity and mortality around the world had been documented in the 20th century. However, the mechanisms involved in the emergence of novel influenza virus and the epidemiological factors leading to pandemics are unpredictable. Southern China is postulated as the epicentre of influenza epidemics due to its agricultural-based communities and high population density. Pandemic influenza viruses are through to arise from avian viruses through genetic reassortment among influenza viruses. An influenza virus (H5N1) known to infect only birds previously was found to infect human causing disease and death in Hong Knog in 1997 and the outbreak involved 18 patients with six deaths. Prior to the human outbreak, the H5N1 virus was found to cause extensive death in chickens in three farms in Hong Kong. The significance of this outbreak raised worldwide concern on the possibilities that such an influenza virus may become the next influenza pandemic strain. Investigations were initiated to find the source of the virus. In addition the extend of spread in individuals in contact with the index case and infected poultry was studied by H5-specific serology. Results demonstrated that individuals in close contact with the index case or with exposure to poultry were at risk of being infected. Out of the 18 cases of human infection, eleven had severe infection with symptoms of pneumonia and multi-organ failure. All severe cases presented with lower respiratory infection and lymphopenia and six eventually died. Case-fatality ratio was high among patients over 12 years of age (five out of nine). Control measures aimed at reducing exposure of human to potential H5-positive poultry were instituted which included culling of all poultry in Hong Kong, the segregation of water fowls and chicken, and the introduction of import control measures for chickens. Such measures had successfully controlled the outbreak and continuous surveillance of the poultry in Hong Kong of H5N1 infection is maintained to minimize future human exposure. PMID- 12110266 TI - H5N1 influenza in Hong Kong: virus characterizations. AB - In 1997, 18 people were infected in Hong Kong with an avian influenza A(H5N1) virus from chicken. This type of interspecies transmission was never detected before and could have resulted in the development of a pandemic strain. The occurrence suggests that the pig is not needed for the emergence of pandemic influenza virus strains. Characteristics of the strains involved are discussed in relation to the question why, on the one hand, these strains were able to infect humans but on the other hand were not able to start an epidemic. PMID- 12110267 TI - Preparation of vaccines against H5N1 influenza. AB - In response to the pandemic warning provided by the highly pathogenic H5N1 influenza virus infections in Hong Kong, there were world-wide attempts to develop vaccines. Three strategies were followed and although each was associated with some success, there were also some problems. Pre-clinical vaccine efficacy results are presented from one such strategy, that of using an apathogenic H5N3 avian strain for vaccine production. PMID- 12110268 TI - Pandemic planning. AB - During the previous century, three influenza A pandemics occurred with a variable severity. The two latter were explained by a genetic re-assortment and false alarms without pandemic spread were observed later by the same mechanism or by direct animal infection. The likelihood that such an event occurs again is high and each country has to be prepared for facing what could be a catastrophe. The last event in Hong Kong in 1997 where six persons died, has allowed refining the definitions and phasing a pandemic threat from the moment that a novel virus is discovered. WHO implemented 50 years ago a large network of surveillance with five collaborating centres, including the animal influenza centre of Memphis, and 110 national influenza centres. These centres are encouraged to prepare or improve a national contingency plan that could reduce importantly medical and socio-economic consequences of an influenza A pandemic. Countries or regions are recommended to use these guidelines that provide a framework for preparing an appropriate and proportionate response. PMID- 12110270 TI - In vitro cytotoxic effect of bread wheat gliadin on the LoVo human adenocarcinoma cell line. AB - The pathogenesis of celiac disease is not completely understood but, although the initial step of the process is still unclear, an altered immune response seems to play a major role. Previous studies of the biological properties of gliadin have highlighted its cytotoxic effects, and the aim of this study was to develop an in vitro technique to study them. The LoVo (human colon adenocarcinoma) cell line grown in two-dimensional cultures was exposed to different concentrations of digested bread wheat gliadin (62, 125, 250, 500 and 750 microg/ml) for 48 h, after which cell growth and oxidative balance (the content of reduced glutathione (GSH), and peroxidase, transferase and reductase activity) was evaluated. Other food proteins were used as controls. Our data revealed a statistically significant inhibition of cell growth in proportion to the gliadin concentration (from 26 to 100%), combined with a decrease in GSH content (-38% at 500 microg/ml) and reduced enzymatic activity (-30% at 500 microg/ml). The controls did not show any noxious effect. Our results confirm the usefulness of LoVo cells in evaluating gliadin cytotoxicity and that they can be used to investigate the biological properties of gliadin. PMID- 12110271 TI - Damaging effects of advanced glycation end-products in the murine macrophage cell line J774A.1. AB - The interaction of reducing sugars, such as aldose, with proteins and the subsequent molecular rearrangements, produces irreversible advanced glycation end products (AGEs), a heterogeneous class of non-enzymatic glycated proteins or lipids. AGEs form cross-links, trap macromolecules and release reactive oxygen intermediates. AGEs are linked to aging, and increase in several related diseases. The aim of this study was to assess, in a murine macrophage cell line, J774A.1, the effects of 48 h of exposure to glycated serum containing a known amount of pentosidine, a well-known AGE found in the plasma and tissues of diabetic and uremic subjects. Fetal bovine serum was incubated with ribose (50 mM) for 7 days at 37 degrees C to obtain about 10 nmol/ml of pentosidine. The cytotoxic parameters studied were cell morphology and viability by neutral red uptake, lactate dehydrogenase release and tetrazolium salt test. In the medium and in the intracellular compartment, bound and free pentosidine were evaluated by HPLC, as sensitive and specific glycative markers, and thiobarbituric acid reactive substances (TBARs), as index of the extent of lipid peroxidation. Our results confirm that macrophages are able to take up pentosidine. It is conceivable that bound pentosidine is degraded and free pentosidine is released inside the cell and then into the medium. The AGE increase in the medium was combined with an increase in TBARs, meaning that an oxidative stress occurred; marked cytotoxic effects were observed, and were followed by the release of free pentosidine and TBARs into the culture medium. PMID- 12110272 TI - Ultraviolet B radiation induces activation of neutral and acidic sphingomyelinases and ceramide generation in cultured normal human keratinocytes. AB - The sphingomyelin pathway is an ubiquitous, evolutionary conserved signaling system which transduces an extracellular signal into the cell. During the past few years increasing evidence has shown that the sphingolipid ceramide may play a role as a second messenger in intracellular signal transduction. The ceramide generation via sphingomyelinase (SMase) is followed by three major cellular responses: cell growth arrest, induction of cell differentiation and/or induction of programmed cell death or apoptosis. The aim of this study is to investigate whether activation of SMases and generation of ceramide can be induced by UVB radiation in normal human keratinocytes. The present data show that exposure to UVB radiation results in rapid generation of ceramide. The ceramide accumulation starts 15 min after UV exposure and progressively increases up to 24 h. In vitro measurement of SMase activity following exposure to UVB evidences an activation of both neutral and acidic SMases. Moreover, UVB induces apoptosis in normal human keratinocytes as shown by TUNEL technique and FACS analysis. These data indicate that UVB induced ceramide generation and activation of both neutral and acidic SMases, suggesting that sphingolipids metabolism may be involved in the UVB signaling pathway. PMID- 12110273 TI - Caspase activation and death induced by yessotoxin in HeLa cells. AB - We have studied the death response induced by yessotoxin (YTX) in cultured HeLa cells, and have compared it to that triggered by okadaic acid (OA) in the same experimental system. Sub-nanomolar concentrations of YTX were found to induce HeLa cell death after a 48-96-h incubation. YTX caused loss of intact poly(ADP ribose)-polymerase (PARP) in HeLa cells, and detection of the 85kDa fragment, which is indicative of proteolytic attack by caspases. Measurements of caspase activities using extracts prepared from YTX-treated cells and substrates of the caspase-3/7 and caspase-2 isoforms, showed that the relative proteolysis of caspase-3/7 substrate was about eight-fold higher than that of caspase-2, the levels of which were about twice those measured with extracts from control cells. These findings were matched by Western blot analyses of caspase-2, -3 and -7 in HeLa cell extracts, which showed that the levels of pro-caspase-2 were not greatly affected by YTX treatment, whereas pro-caspase-3 and -7 were activated in YTX-treated cells. Taken together, these data complement others previously obtained with OA, and support the notion that caspase isoforms involved in cell death induced by OA and YTX are cell- and toxin-specific. PMID- 12110274 TI - Response of rainbow trout (Oncorhynchus mykiss) D-11 cell line to 3 methylcholanthrene (3MC) exposure. AB - The rainbow trout cytochrome P4501A gene subfamily consists of two members, CYP1A1 and CYP1A3, which are induced by polycyclic aromatic hydrocarbons (PAHs). In this study, we investigated the induction of cytochrome P4501A3 in the rainbow trout (Onchorhynchus mykiss) D-11 cell line after 3-methylcholanthrene (3MC) exposure by generating chimeric constructs in which a 2.3 kb fragment or portion of the 5'-flanking region of the trout cytochrome CYP1A3 gene was fused to the firefly luciferase (Luc) gene. The constructs were then transiently transfected into the trout D-11 cells and their transcriptional activity measured by luciferase assay after treatment with different 3MC concentrations. Maximal induction following exposure to 2 microM 3MC was 2.2-fold after 72 h. Deletion of the region specifying the 5' untranslated region (5'UTR) of the mRNA encoding the CYP1A3 gene increased unstimulated luciferase activity but also led to a loss of response to 3MC treatment. This finding suggests that the region specifying the 5'UTR contains a negative element that is also involved in the transcriptional response to 3MC. PMID- 12110276 TI - Inter-individual variation in urothelial DNA repair gene expression in vitro. AB - DNA repair efficiency may play a significant role in individual susceptibility to bladder cancer, the third most common cancer in Europe. Bladder cancer arises from the urothelial cell layer which lines the urinary tract. As DNA repair gene expression levels should reflect DNA repair capacity, we investigated the expression of genes from the base excision, nucleotide excision and mismatch repair pathways in normal human urothelial (NHU) cells in vitro. RNA was extracted from six independent NHU cell lines and expression of 26 DNA repair genes was determined by ribonuclease protection assay. The results show that all the genes analysed were detected in NHU cells in vitro with a similar expression pattern in most cell lines. However, there was some variation between cell lines, with one expressing base excision repair genes very strongly, but another having weak expression of mismatch repair genes. These results suggest that DNA repair genes are constitutively expressed by NHU cells and that there is some inter individual variation. Prospective studies are required to determine whether these differences in gene expression may play a role in susceptibility to bladder cancer. PMID- 12110275 TI - Time-dependent variations of drug-metabolising enzyme activities (DMEs) in primary cultures of rabbit hepatocytes. AB - In the present study, time-dependent variations of drug-metabolising enzyme activities (DMEs) in primary cultures of rabbit hepatocytes, a species of economic importance in Mediterranean countries, were investigated. Cross-bred rabbits were anesthetised and their livers perfused in situ by a two-step collagenase technique; cells suspensions were filtered, seeded in collagen-coated dishes and cultivated at 37 degrees C in a controlled atmosphere for 24 and 72 h. Cytochrome P450 and b(5) contents as well as the catalytic activity of some P450 dependent monooxygenases were measured in subcellular fractions obtained by differential ultracentrifugation; microsomal proteins were also subjected to immunoblotting, using antibodies to rat P4501A, 2B, 2E1 and 3A isoforms. The activity of some microsomal hydrolytic enzymes was also determined. As regards conjugative enzymes, glutathione content and activities of glutathione S transferase, uridindiphosphoglucuronosyl-transferase, acetyl-transferase and 1,2 epoxibuthane glutathione transferase were assayed. An overall reduction of the catalytic activity was observed 72 h after plating, reaching in certain instances the level of statistical significance. On the whole, our data confirm those previously reported with hepatocytes obtained from other species; however, the evidence that DMEs were still measurable after 72 h supports the usefulness of this in vitro method for drug metabolism studies in the rabbit as well. PMID- 12110277 TI - Role of the lung resistance-related protein (LRP) in the drug sensitivity of cultured tumor cells. AB - Drug resistance, one of the major obstacle in the successful anticancer therapy, can be observed at the outset of therapy (intrinsic resistance) or after exposure to the antitumor agent (acquired resistance). To gain a better insight into the mechanisms of intrinsic resistance we have analyzed two human cell types derived from untreated tumors: MCF-7 breast cancer and A549 non small cell lung cancer (NSCLC). We have examined: the cytotoxic effect induced by doxorubicin (DOX); the time course of drug accumulation by flow cytometry and intracellular drug distribution by confocal microscopy; the expression and distribution of proteins related to anthracycline resistance, such as P-gp (P-glycoprotein), MRP1 (multidrug resistance-associated protein) and LRP (lung resistance-related protein). The cytotoxicity assays showed that A549 cells were less sensitive than MCF-7 cells to the DOX treatment in agreement with the different DOX uptake. Moreover, while in A549 cells DOX was mostly located in well defined intracytoplasmic vesicles, in MCF-7 cells it was mainly revealed inside the nuclei. The analysis of P-gp and MRP expression did not show significant differences between the two cell lines while a high expression of LRP was detected at the nuclear envelope and cytoplasmic levels in A549 cells. These findings suggest that the lower sensitivity to DOX treatment showed by lung carcinoma cells could be ascribed to drug sequestration by LRP inside the cytoplasmic compartments. PMID- 12110278 TI - Iron and copper alter tight junction permeability in human intestinal Caco-2 cells by distinct mechanisms. AB - Human intestinal Caco-2 cells differentiated for 15-17 days on transparent filter inserts were treated for up to 3 h with 50 and 100 microM CuCl(2) or FeSO(4) in the AP compartment at pH 6.0. Trans-epithelial electrical resistance (TEER) showed a progressive decrease during the course of the experiment that was slower in cells treated with 50 microM CuCl(2) than in those treated with 100 microM CuCl(2). Both 50 and 100 microM FeSO(4) produced a similar decrease in TEER over time, tailing off after 120 min. F-actin localization by fluorescent phalloidin binding in control cells and in cells treated for 3 h with 50 microM CuCl(2) or FeSO(4) highlighted striking differences in the two treatments. Cu(II) led to an overall reduction in F-actin staining with extensive depolymerization in areas of the monolayer, in the absence of cellular loss. Conversely, Fe(II) treatment produced disorganization of F-actin and decreased staining of the perijunctional actin filaments. No changes in the localization and intensity of staining of the junctional proteins ZO1, occludin and E-cadherin were observed after treatment with 100 microM FeSO(4) in analogy with previous observations in Cu(II)-treated cells. The data presented suggest that different mechanisms are responsible for the changes to tight junction permeability produced by the two metals. PMID- 12110279 TI - Epithelial cells and expression of the phagocytic marker CD68: scavenging of apoptotic bodies following Rho activation. AB - Macropinocytosis is a ruffling-driven process which drives the ingestion of large particles by both macrophages and epithelial cells. In this context, we have previously described a Rho-activating bacterial toxin from E. coli, the cytotoxic necrotizing factor 1 (CNF1), which allows epithelial cells to macropinocytose not only latex beads and bacteria, but also apoptotic cells in a fashion similar to that of professional phagocytes. We herein report that (i) epithelial cells express the typical phagocytic marker CD68, (ii) Rho activation by CNF1 varies the intracellular localization of CD68, which appears to be co-distributed, as in macrophages, with the homologous lysosomal protein Lamp-1. Together with the capability of digesting apoptotic cells following their internalization, our findings indicate that Rho-activated epithelial cells behave in most respects as professional phagocytes. PMID- 12110280 TI - Synthesis of a new platinum(II) complex: anticancer activity and nephrotoxicity in vitro. AB - New mixed dithiocarbamate-amino Pt(II) complex ([Pt(ESDT)(Py)Cl]) has been recently synthesised with the aim to produce potential anticancer drug able to conjugate cytostatic activity with lack of nephrotoxicity. This complex contains: (1) an amino ligand; (2) a good leaving group (halide); and (3) an S-containing chelating agent potentially able to protect the metal centre from its interaction with S-containing protein-legating sites that are believed to be at the basis of the nephrotoxicity of Pt(II)-based drugs. This complex has been found to be effective as an antiproliferative agent (more active than cis-platin) towards a normal human adenocarcinoma cell line and the corresponding cis-platin-resistant C13 strain. Toxicity tests on the kidney were performed by means of a renal cortical slice model. The slices, prepared with a Brendel-Vitron slicer, were incubated with different doses (0.125-5.0 x 10(-4) M, final concentration) of [Pt(ESDT)(Py)Cl] or cis-platin dissolved in methyl sulphoxide. The platinum(II) complex showed very low renal cytotoxicity as compared with cis-platin; in particular, lipid peroxidation induced by cis-platin appeared about five-fold higher than that induced by [Pt(ESDT)(Py)Cl]. In conclusion, besides being less toxic for the kidney, the results showed that the new synthesised platinum(II) complex appeared in vitro more effective than cis-platin when tested on sensitive and resistant cis-platin tumour cell lines. PMID- 12110281 TI - Epithelial cells challenged with a Rac-activating E. coli cytotoxin acquire features of professional phagocytes. AB - Activation of Rho, Rac and Cdc42 GTPases by an Escherichia coli cytotoxin (CNF1) has been reported to induce a phagocytic-like activity by epithelial cells in terms of a ruffle-driven capture and ingestion of large material. More recently, it has been reported that treatment with CNF1 induces superoxide anion release by these cells following a phagocytic stimulus. We herein show that in epithelial cells both transfection with the dominant form of Rac (RacV12) and treatment with the Rac-activating epidermal growth factor (EGF) may increase the secretion of superoxide anions on challenge with latex beads. Moreover, exposure to CNF1 induces a significant augmentation of acidic vesicles where the internalized particles were detectable. Our results indicate that (i) Rac is a pivotal GTPase for inducing in epithelial cells superoxide anion generation and (ii) the internalized material travels trough acidic compartments in CNF1-treated epithelial cells. Altogether this suggests a novel role for epithelial cells that, following Rac activation, might share with professional phagocytes the task of eliminating unwanted pathogens. PMID- 12110282 TI - Use of differential display-polymerase chain reaction to identify genes selectively modulated by chemical allergens in reconstituted human epidermis. AB - In the screening of topical drugs, cosmetics and other chemicals for human use, it is very important, both from a safety and an economic point of view, to have biological markers to discriminate irritant and allergic contact dermatitis that have different impacts on human health. Owing to their anatomical location, keratinocytes are among the first cells to be exposed to various antigens and the use of these cells as a simplified in vitro model to evaluate the potential toxicity of chemicals destined for cutaneous application is amply justified. The purpose of this work was to identify new genes selectively modulated by skin toxicants. Commercially available reconstituted human epidermis (Epiderm) was treated for 18 h with sodium dodecyl sulfate (SDS) 0.4 mg/ml, as reference irritant, or with dinitrochlorobenzene (DNCB) 0.2 mg/ml, as reference allergen, or with vehicle control. Differential display PCR (DD-PCR) was performed. Results identified adipose differentiation-related protein (ADRP) as up-regulated by both irritant and allergen, and KIAA0368 as selectively up-regulated by contact allergen. These data indicate the enormous potential of functional genomic techniques, which allow the identification of genes not immediately connected with the immune response, or even novel genes with unknown functions, which nevertheless may be potential markers of skin irritation and allergy. PMID- 12110283 TI - The relative embryotoxicity of 1,3-dichloro-2-propanol on primary chick embryonic cells. AB - 1,3-Dichloro-2-propanol (1,3-DCP) is a chlorinated compound used in the fabrication of industrial products such as hard resins, celluloid or paints. It has also been detected in instant soups and soy sauce. 1,3-DCP has been associated with major necrosis of the liver in humans [Chem.-Bio. Interact. 80 (1991) 73]. In humans and laboratory animals, 1,3-DCP is metabolised to dichloroacetone (1,3-DCA) by cytochromes P450 2E1 and 1A2 [J. University Occup. Environ. Health 14 (1992) 13]. 1,3-DCA is a hepatotoxin. We suggest that 1,3-DCA could be embryotoxic at doses that do not cause adverse maternal hepatic damage. To investigate the embryotoxic effects of 1,3-DCA, we have adapted a micromass culture method from Atterwill and colleagues [1992. A tiered system for in vitro neurotoxicity testing. In: Zbinden, G. (Ed.), The Brain in Bits and Pieces. Verlag M.T.C., Vollikon, pp. 89-91], using chick midbrain cells and from Wiger et al. [Pharmacol. Toxicol. 62 (1988) 32] using chick mesenchymal cells. The basis of the micromass system is that embryotoxins in vitro are likely to affect development and differentiation of disaggregated neuronal and limb bud micromass cultures. The endpoints chosen for the midbrain assay are resazurin reduction (viability), total protein content (cell number), morphological quantification of neuronal cultures (neuronal projection number) and of limb bud cultures (cartilage nodule number). Preliminary results using chick whole embryo cultures indicated that 1,3-DCA had an inhibitory effect on whole chick embryo development. We also found that embryonic derived cells were sensitive to 1,3-DCA but not 1,3-DCP at concentrations above 1 microM, suggesting a potential teratogenic effect of 1,3-DCA. The exposure to 1,3-DCP is not limited to industrial settings, and hence a better knowledge of its effects and tissue specific actions on embryonic-derived cells would be beneficial. PMID- 12110284 TI - Comparison of embryotoxicity of ESBO and phthalate esters using an in vitro battery system. AB - Epoxidized soy bean oil (ESBO) and phthalate esters have been used as a plasticizer in polyvinyl chloride products. In this study, the embryotoxicity of ESBO and phthalate esters, namely, diethyl hexyl phthalate (DEHP), butylbenzyl phthalate (BBP) and dibutyl phthalate (DBP) was evaluated using short-term in vitro battery system, such as the whole embryo, midbrain and limb bud culture systems. Whole embryos at gestation day 9.5 were cultured for 48 h and the morphological scoring was measured. The cytotoxic effect and differentiation for mid-brain (MB) and limb bud (LB) cell were assessed by 50% inhibition concentration (IC(50)) with neutral red uptake and hematoxylin-stained foci (MB) or Alcian Blue staining (LB), respectively. In the whole embryo culture assay, ESBO (83, 250 and 750 microg/ml) exerted no toxic effect on growth and development of the embryo, whereas phthalate esters (1, 10, 100 microg/ml for DEHP, 10, 100, 1,000 microg/ml for BBP and DBP) inhibited growth and development dose dependently. In mid-brain and limb bud culture, the IC(50) of differentiation and cytotoxicity in BBP was 412.24 and 231. 76 microg/ml for mid brain, and 40.13 and 182.38 microg/ml for limb bud, respectively. The IC(50) of differentiation and cytotoxicity in DBP was 27.47 and 44.53 microg/ml for mid brain, and 21.21 and 25.54 microg/ml for limb bud cells, respectively. The lower IC(50) in both cells was obtained from DBP when compared to BBP. From these results, limb bud cells responded more sensitively to BBP and DBP than mid-brain cells. The IC(50) of limb bud cell differentiation and cytotoxicity in DBP is 1.9 and 7.1 less than that of BBP. However, any alteration in cytotoxicity and differentiation was observed with ESBO treatment. These studies suggested that ESBO is not embryotoxic; however, DEHP, BBP and DBP exhibit embryotoxic potential at high concentration. PMID- 12110285 TI - Phototoxic effect of fluoroquinolones on two human cell lines. AB - Photosensitization induced by the fluoroquinolone ofloxacin (OFLX) has been studied using two human cell lines, HL60 and K562, two UV wavelengths, 290 and 330 nm, and two different exposure protocols, acute and protracted. The examined endpoints are the cellular lethality and recovery and the membrane changes produced by the oxidative damage, studied using cloning and counting techniques and the measurement of the generalized polarization (GP) of the fluorescent membrane probe 2-dimethylamino-6-lauroyl-naphthalene (Laurdan). The results show that: (i) the photosensitizing effect is detectable at concentrations similar to those found in patients treated with OFLX only when the cells are irradiated with 330 nm; (ii) the amount of photodamage is a function of the drug concentration and of UV dose and persists also after the removal of the drug; (iii) during the first 24 h after OFLX treatment, a large decrease of the cell number can be observed due to cell lysis; (iv) the OFLX is inserted in the cell membranes at concentrations directly related with the drug concentration and incubation time; (v) the OFLX produces an increase in the GP values similar to that produced by membrane lipid oxidation which persists for hours after the removal of the drug. The overall results suggest the cell membrane as the main target of the OFLX adverse action, with a possible mechanism involving the formation of reactive oxygen species (ROS), which triggers, in turn, the lipid peroxidation chain reaction. PMID- 12110286 TI - Antioxidant potential and gap junction-mediated intercellular communication as early biological markers of mercuric chloride toxicity in the MDCK cell line. AB - In this study, the early nephrotoxic potential of mercuric chloride (HgCl(2)) has been evaluated in vitro, by exposing a renal-derived cell system, the tubular epithelial Madin-Darby canine kidney (MDCK) cell line, to the presence of increasing HgCl(2) concentrations (0.1-100 microM) for different periods of time (from 4 to 72 h). As possible biological markers of the tubular-specific toxicity of HgCl(2) in exposed-MDCK cultures we analysed: (i) critical biochemical parameters related to oxidative stress conditions and (ii) gap-junctional function (GJIC). HgCl(2) cytotoxicity was evaluated by cell-density assay. The biochemical analysis of the pro-oxidant properties of the mercuric ion (Hg(2+)) was performed by evaluating the effect of the metal salt on the antioxidant status of the MDCK cells. The cell glutathione (GSH) content and the activity of glutathione peroxidase (Gpx) and catalase (Cat), two enzymes engaged in the H(2)O(2) degradation, were quantified. HgCl(2) influence on MDCK GJIC was analysed by the microinjection/dye-transfer assay. HgCl(2)-induced morphological changes in MDCK cells were also taken into account. Our results, proving that subcytotoxic (0.1-10 microM) HgCl(2) concentrations affect either the antioxidant defences of MDCK cells or their GJIC, indicate these critical functions as suitable biological targets of early mercury-induced tubular cell injury. PMID- 12110287 TI - Induction of the CYP2B genes by triphenyldioxane treatment in the rat liver. AB - Triphenyldioxane (TPD) is a potent phenobarbital-type (PB) inducer of the CYP2B cytochrome isoforms, the inducing effect of which is one order of magnitude higher than PB. The fact that TPD is unable to induce CYP2B genes having the proximal promoter disrupted (mouse Cyp2b10) suggests an existence of the proximal promoter-dependent mechanism of the CYP2B induction. So a TPD-dependent activation of the nuclear proteins to the binding with Barbie-box sequence (the most conservative part of the proximal promoter) was studied. In the nuclear extracts from the intact rat liver there were detected five proteins that could be activated to the Barbie-box binding by the TPD treatment in vitro (II, III, NI, NII and NIII). The first three were effected also by another PB-like inducers tested (PB and TCPOBOP), when NII and NIII complexes were formed under the influence of TPD only. It is possible that a direct activation of the NII and NIII proteins by TPD exists as (3)H-labeled TPD was detected in the composition of NII and NIII complexes. However, both of them disappeared from the nuclear extracts after the long exposure time with TPD (6 h or more). A short induction by the direct intra-liver delivery of TPD (15-30 min) led to the stabile activation of one TPD-specific protein. Apart from the activation of the Barbie box-binding protein, the short TPD treatment caused the activation of three nuclear proteins being able to interact with the NR1 sequence of the distal promoter PBREM element. These findings suggest that TPD is really the first member of the PB-like inducers family for which a special mechanism of CYP2B induction may exist. PMID- 12110288 TI - Estrogenic activity of procymidone in primary cultured rainbow trout hepatocytes (Oncorhynchus mykiss). AB - It has been shown that procymidone, a dicarboximide fungicide, alters sexual differentiation in vivo and in vitro. The aim of this study was to evaluate the estrogenic activity of this fungicide using the synthesis of vitellogenin (Vtg) in rainbow trout hepatocyte as a biological marker. The cells were treated for 24 h with procymidone 150 microM, using 17beta-estradiol 20 microM as a positive control. The doses were chosen on the basis of cell viability (Neutral Red and MTT tests) and solubility. The results show that procymidone leads to a qualitative and quantitative increase in Vtg synthesis. In Western immonoblots, the 170 and 30 kDa bands, which respectively correspond to the monomeric form of Vtg and posvitine, were brighter in cells treated with procymidone and 17beta estradiol than those corresponding to the negative controls (cells treated for 24 h with DMSO 0.1% alone); ELISA showed that the cells treated with the fungicide and 17beta-estradiol had a 48 and 76%, respectively, higher Vtg concentration than the negative controls (P<0.01). Western blotting also revealed the induction of HSP27 (27 KDa), which further confirms the estrogenic acitivity of procymidone as it is known that the 3' region of HSP27/28 containing the gene mRNAs is induced by estrogen treatment. Procymidone increased also the production of both HSP70 protein (70 KDa) and free oxygen radicals. This last finding is in agreement with the toxic mechanism of dicarboximide fungicides. It can therefore be presumed that the estrogenic activity of procymidone in primary cultured trout hepatocytes is related to oxidative damage which, as many other studies have shown, can increase the levels of estrogens such as 17beta-estradiol, and thus increase Vtg synthesis PMID- 12110289 TI - Cysteine dioxygenase: modulation of expression in human cell lines by cytokines and control of sulphate production. AB - Cysteine dioxygenase (CDO) is the initial and rate-limiting enzyme involved in the oxidative degradation of cysteine to inorganic sulphate. It is believed to be the major source of sulphate in vivo. Inflammatory conditions such as rheumatoid arthritis have been linked with high plasma cysteine:sulphate ratios in patients. The cytokines tumour necrosis factor-alpha (TNF-alpha) and transforming growth factor-beta (TGF-beta) have been shown to inhibit the expression of CDO in neuronal (TE671) and hepatic (Chang) human cell lines at nanomolar concentrations. Cytokine release may therefore modulate sulphate production and hence regulate formation of sulphated biocomponents. PMID- 12110290 TI - A protein from the cabbage looper, Trichoplusia ni, regulated by a bacterial infection is homologous to 3-dehydroecdysone 3beta-reductase. AB - During the screening of immune-regulated genes from the cabbage looper, Trichoplusia ni, a 3-dehydroecdysone 3beta-reductase homologue (DERH) was cloned. In the course of development, 3-dehydroecdysone 3beta-reductase mediates the conversion of 3-dehydroecdysone (3dE) secreted from the prothoracic glands to ecdysone (E), which is subsequently converted to 20-hydroxyecdysone (20E), the major insect molting hormone. The cloned gene is upregulated in fat body during development and is strongly induced after the larva is challenged with bacteria. The gene codes for a 308 amino acid residue protein which shows 42.5% identity to Spodoptera littoralis 3-dehydroecdysone 3beta-reductase. Using the baculovirus expression system, the recombinant DERH was expressed. The purified protein mediates the reduction of 3-dehydromakisterone A to makisterone A, and requires NADPH as a cofactor. Western blots using an antiserum to T. ni DERH revealed the presence of the protein in larval hemolymph and integument. The data indicate that the protein is regulated developmentally and is induced after a challenge with bacteria. Immunohistochemical studies localized the enzyme exclusively in the epidermis and the cuticle. PMID- 12110291 TI - Evolution of noctuid pheromone binding proteins: identification of PBP in the black cutworm moth, Agrotis ipsilon. AB - Male black cutworm moths (Agrotis ipsilon, Lepidoptera, Noctuoidea, Noctuidae), which are attracted by a three-component pheromone blend ((Z)-7-dodecenyl acetate, Z7-12:Ac; (Z)-9-tetradecenyl acetate, Z9-14:Ac; (Z)-11-hexadecenyl acetate, Z11-16:Ac), express diverse antennal pheromone binding proteins (PBPs). Two PBP isoforms (Aips-1 and Aips-2) that show 46% identity were cloned from antennal cDNA of male A. ipsilon. The protein Aips-1 displays a high degree of identity (70-95%) with PBPs of other noctuiids, but shows only 42-65% identity with the PBPs of more phylogenetically distant species. The other protein, Aips 2, represents a distinct group of PBP that includes proteins from Sphingidae and Yponomeutidae. These differences observed suggest that each of the two PBPs may be tuned to a specific pheromone ligand. PMID- 12110292 TI - Cyclic nucleotide-independent phosphorylation of vitellin by casein kinase II purified from Rhodnius prolixus oocytes. AB - In this study we show that Vitellin (VT) phosphorylation in chorionated oocytes of Rhodnius prolixus is completely inhibited by heparin (10 microg/ml), a classical casein kinase II (CK II) inhibitor. VT phosphorylation is not affected by modulators of cyclic nucleotide-dependent protein kinases such as c-AMP (10 microM), H-8 (1 microM) and H-89 (0.1 microM). We have obtained a 3000-fold VT free enriched preparation of CK II. Autophosphorylation of this enzyme preparation in the presence of (32)P-ATP demonstrated that it lacks any endogenous substrates. Rhodnius CK II is strongly inhibited by heparin (Ki = 9 nM) and uses ATP (Km = 36 microM) or GTP (Km = 86 microM) as phosphate donors. Incubation of VT with purified Rhodnius CK II and (32)P-ATP led to the incorporation of 2 mols of phosphate/mol VT. However, the total number of phosphorylation sites available can be altered by previous incubation of VT with alkaline phosphatase. These data show that an insect yolk protein contain phosphorylation sites for a cyclic nucleotide-independent protein kinase such as CK II. PMID- 12110293 TI - Cyp6a8 of Drosophila melanogaster: gene structure, and sequence and functional analysis of the upstream DNA. AB - In Drosophila, the insecticide resistant 91-R strain is an overproducer and susceptible 91-C and ry(506) strains are the underproducers of CYP6A8 mRNA encoded by a cytochrome P450 gene, Cyp6a8. Low expression of Cyp6a8 in the underproducer strains is due to a downregulatory effect of a putative repressor locus, which is thought to be mutant in the overproducer strain. In the present investigation, organization of Cyp6a8 and promoter activity of its upstream DNA were analyzed. Cyp6a8 has two introns of which intron II is similar to the introns of other insect CYP genes with respect to its length and position. Intron I is only 36 bp long and lacks consensus splice sites. It is also in-frame with the CYP6A8 open reading frame. Therefore, inefficient splicing of intron I may produce two isoforms of CYP6A8. Analysis of Cyp6a8 upstream DNA of the overproducer 91-R strain showed that DNA sequences between -199 and -761 bp are required for the highest constitutive and barbital-induced expression of Cyp6a8. This region has six barbie boxes and binding sites for various transcription factors. Promoter activity of the -11/-761 DNA of the overproducer 91-R strain was found to be 4-fold lower in the genome of underproducer ry(506) strain, which is wild type for the putative repressor gene, than in the genome of F1 hybrids of 91-R and ry(506) strains. These results suggest that -11/-761 Cyp6a8 DNA of the 91-R strain can respond to the active repressor present in the hybrid genome and further support our previous findings that overexpression of Cyp6a8 is a result of mutation of a repressor gene rather than mutation of the cis-regulatory sequences. PMID- 12110294 TI - Oocyte fertilization triggers acid phosphatase activity during Rhodnius prolixus embryogenesis. AB - Acid phosphatase activity, previously identified in Rhodnius prolixus oocytes, was studied during egg development. Fertilized eggs exhibited a five fold increase of total acid phosphatase activity during the first days of development. In contrast non-fertilized oviposited eggs showed no activation of this enzyme. An optimum pH of 4.0 for pNPP hydrolysis in a saturable linear reaction and a strong inhibition by lysosomal acid phosphatase inhibitors such as NaF (10 mM) and Na(+)/K(+) tartrate (0.5 mM) are the major biochemical properties of this enzyme. Fractionation of egg homogenates through gel filtration chromatography revealed a single peak of activity with a molecular mass of 94 kDa. The role of this enzyme in VT dephosphorylation was next evaluated. Western blots probed with anti-phosphoserine polyclonal antibody demonstrated that VT phosphoaminoacid content decreases during egg development. In vivo dephosphorylation during egg development was confirmed by following the removal of (32)P from (32)P-VT in metabolically labeled eggs. Vitellin was the only phosphorylated molecule able to inhibit pNPPase activity of partially purified acid phosphatase. These data indicate that acid phosphatase activation follows oocyte fertilization and this enzyme seems to be involved in VT dephosphorylation. PMID- 12110295 TI - Protein purification, cDNA cloning and gene expression of attacin, an antibacterial protein, from eri-silkworm, Samia cynthia ricini. AB - Attacin was isolated from immunized larval hemolymph of the wild silkmoth, Samia cynthia ricini. The antibacterial effect of the attacin was limited to some species of Gram-negative bacteria. Two cDNA clones encoding attacin A and B, respectively, were isolated by screening the cDNA library from immunized fat body. The two cDNAs encoded the same length of precursor protein with 233 amino acid residues. The 46-residue prepropeptides of the two attacins were identical to each other, but 4 out of 187 residues of the mature proteins were different in each other. The two attacins show 98% identity at the amino acid level, while 92% identity at the nucleotide level. Both of the mature proteins were highly homologous to cecropia basic attacin with identity of 96%. The attacin transcripts were detected at significant level in fat body, hemocytes and Malpighian tube after injection with peptidoglycan, but not in the midgut and the silkgland. The induction of attacin gene expression was elicited most effectively by peptidoglycan and UV-killed bacteria in the fat body. PMID- 12110296 TI - Partial purification of a farnesyl diphosphate synthase from whole-body Manduca sexta. AB - Farnesyl diphosphate synthase (FPP synthase) is a ubiquitous enzyme that is required for the biosynthesis of sesquiterpenes, dolichols ubiquinones, and prenylated proteins in insects. We report on the partial purification and characterization of an FPP synthase, obtained from whole-body preparations of the lepidopteran insect, Manduca sexta. The larval enzyme was separated from isopentenyl diphosphate (IPP) isomerase, phosphatase, and GGPP synthase by preparative isoelectric focusing, and was further purified by DEAE Sepharose, hydroxyapatite, and size exclusion chromatography. Whole-body M. sexta FPP synthase has a native molecular weight of 60.5+/-3.5 kDa and consists of two subunits of 28.5+/-0.5 kDa. As seen with other prenyltransferases, the enzyme has an absolute requirement for divalent cation and both Mn(2+) and Mg(2+) stimulated activity, although the former was inhibitory at higher concentrations. Insect FPP synthase catalyzes the condensation of IPP (K(m)=2.9+/-1.2 microM) with both dimethylallyl diphosphate and geranyl diphosphate (K(m)=0.8+/-0.4 microM). The enzyme requires the presence of detergent, glycerol, and non-specific protein protein interactions for stability and maximum catalytic activity. PMID- 12110297 TI - A new member of the PBAN family in Spodoptera littoralis: molecular cloning and immunovisualisation in scotophase hemolymph. AB - In this article, we report evidence suggesting that the immunoreactive factor previously detected in Spodoptera littoralis scotophase hemolymph is PBAN, which supports a humoral route of the hormone to the pheromone gland. Western blot after native-PAGE of prepurified scotophase hemolymph extracts yielded an immunoreactive band with the same mobility as S. littoralis Br-SOG factor and the expected mobility for a noctuid PBAN. This band was not detected in photophase hemolymph extract. The identity of S. littoralis Br-SOG factor as PBAN was obtained from cDNA cloning using RT-PCR strategy. This allowed us to deduce the amino acid sequence of Spl-PBAN, which is highly homologous to other known PBANs. Moreover, we found that the PBAN encoding cDNA also encoded four other putative amidated peptides (Spl-DH homologue, Spl-alpha-NP, Spl-beta-NP and Spl-gamma-NP) that are identical or highly conserved among noctuids, and two non amidated peptides of unknown function. This cDNA organization is common to all known cDNAs encoding PBANs, leading to the release of different peptides after putative enzymatic cleavage of the preprohormone. PMID- 12110298 TI - Structure-activity relations of the dark-colour-inducing neurohormone of locusts. AB - The dark-colour-inducing effect of several peptides in comparison to that of the dark-colour-inducing neurohormone (DCIN, [His(7)]-corazonin) of locusts was investigated by a bioassay based on nymphs of a DCIN-deficient albino mutant of Locusta migratoria. The study was aimed at elucidating the active part of the DCIN and to explore the contribution of its amino acids to the activity. Graded doses of all peptides were injected in oil. [Arg(7)]-corazonin and DCIN were equally effective. Certain arthropod neuropeptides having the -SXGW- partial sequence (a part of the DCIN and of [Arg(7)]-corazonin; X=His and X=Arg, respectively) yielded the following findings: Scg-AKH-II (adipokinetic hormone II of Schistocerca gregaria X=Thr), Grb-AKH ( adipokinetic hormone of Gryllus bimaculatus X=Thr) and RPCH (red pigment concentrating hormone of crustaceans X=Pro) evoked a moderate darkening response, but Lom-AKH-II (adipokinetic hormone II of L. migratoria X=Ala) was ineffective. Step by step shortening of the sequence of the DCIN at the N-terminal, from pGlu-3-11DCIN to pGlu-9-11DCIN, resulted in a decreasing activity, but even pGlu-9-11DCIN induced a weak response with high doses. Shortening of the DCIN from the C-terminal revealed a moderate activity of 1-7DCIN-NH(2) and a weak activity of 1-5DCIN-NH(2). An octadecapeptide which induces dark colour in moth larvae, having the pentamer FTPRL-NH(2) at its C-terminal, evoked no darkening in the albino locusts. We conclude that although the -SXGW- partial sequence has some role in induction of darkening, for obtaining maximal effect the whole sequence of the DCIN (or of [Arg(7)]-corazonin) is necessary. PMID- 12110299 TI - Insect hemolymph clotting: evidence for interaction between the coagulation system and the prophenoloxidase activating cascade. AB - Here we describe a novel approach to isolate proteins involved in insect hemolymph coagulation. In order to avoid problems in purifying clot proteins after they had been crosslinked, we performed an in vitro coagulation reaction with cell-free hemolymph from the lepidopteran Galleria mellonella and used the resulting complexes to produce a specific antiserum. The antiserum reacted with a subset of hemolymph proteins as well as with granular cells, but not with other hemocyte types of Galleria. Screening expression libraries identified some positive clones, which turned out to code for some previously characterized components of immune cascades, as well as some novel candidates for clotting factors. Known components include members of both the coagulation system and the prophenol-activating cascade, lending support to the idea that both systems work together during the formation of a hemolymph clot. Novel candidates for insect clotting factors include a mucin-like protein, a glutathione-S-transferase, and a distant member of the alpha-crystallin/small heat shock protein family. Using assays measuring the activity of transglutaminase, a key enzyme in clotting reactions in both vertebrates and invertebrates, we found a partial overlap between transglutaminase substrates and proteins recognized by the antiserum against the in vitro-induced clot. PMID- 12110300 TI - Purification and characterization of the plasma membrane glycosidases of Drosophila melanogaster spermatozoa. AB - Previous studies from our laboratory have demonstrated the presence of two integral proteins with glycosidase activity in the plasma membrane of Drosophila melanogaster spermatozoa and we have suggested that these enzymes might have a role in sperm-egg binding. In this study the glycosidases have been purified and characterized. We have evidenced the presence of three distinct enzymes, two beta N-acetylhexosaminidase isoforms, named HEX 1 and HEX 2, and an alpha-mannosidase. The molecular size of the native enzymes estimated by gel filtration was 158 kDa for beta-hexosaminidases and 317 kDa for alpha-mannosidase. SDS-PAGE showed that HEX 1 and HEX 2 are dimers formed by subunits with different molecular sizes, whereas alpha-mannosidase consists of three subunits with different molecular weights. All the enzymes are terminally glycosylated. Characterization of the purified enzymes included their 4-methylumbelliferyl-substrate preferences, kinetic properties, inhibitor constants and thermal stability. On the basis of substrate specificity, kinetics and the results of inhibition studies, beta hexosaminidases appear to differ from each other. HEX 1 and HEX 2 are similar to mammalian isoenzyme A and isoenzyme B, respectively. These findings represent the first report on the characterization of sperm proteins that are potentially involved in interactions with the egg in Insects. PMID- 12110301 TI - Isolation, characterization, and functional expression of kynurenine aminotransferase cDNA from the yellow fever mosquito, Aedes aegypti(1). AB - This study describes the molecular and biochemical characterization of kynurenine aminotransferase (KAT) from Aedes aegypti mosquitoes. Through screening an A. aegypti larval cDNA library, a 1695-bp full-length cDNA clone with a 1434-bp open reading frame (ORF) was isolated. Its deduced amino acid sequence consists of 477 amino acid residues with a predicted molecular mass of 53,490 and the amino acid sequence shares 47 and 42% identity with KATs from Homo sapiens and Rattus norvegicus, respectively. This putative A. aegypti KAT (AeKAT) has four potential N-glycosylation sites (Asn-Xxx-Trp/Ser) and a typical mitochondrial leader sequence consisting of 49 amino acids at its NH(2)-terminus with a putative cleavage site between Met-49 and Ser-50. A consensus pyridoxal 5'-phosphate (PLP) binding domain (Gly-Ser-Ala-Gly-Lys-Thr-Phe-Ser) is present in the central region of the ORF. Using a baculovirus/insect cell expression system, a full-length AeKAT and a truncated AeKAT, lacking the mitochondrial leader sequence, were expressed. The full-length AeKAT expressed in Sf9 insect cells is insoluble and has no detectable KAT activity, but the truncated AeKAT is soluble and capable of catalyzing the transamination of kynurenine to kynurenic acid in the presence of pyruvate as an amino group acceptor. However, the expressed truncated protein is not active to 3-hydroxykynurenine. Northern analysis indicates that transcription of the AeKAT occurs at all stages during mosquito development, but higher levels of mRNA are observed during the pupal and adult stages. Results indicate that a specific KAT is present in mosquitoes and the enzyme may play an important physiological role in A. aegypti. PMID- 12110302 TI - Transport of the highly charged myo-inositol hexakisphosphate molecule across the red blood cell membrane: a phase transfer and biological study. AB - To address the problem of delivering highly charged small molecules, such as phytic acid (InsP(6) or IHP), across biological membranes, we investigated an approach based on a non-covalent interaction between transport molecule(s) and IHP. Thus, we synthesized a collection of compounds containing IHP ionically bound to lipophilic (but non-lipidic) ammonium or poly-ammonium cations. First, we assessed the ability of these water-soluble salts to cross a biological membrane by measuring the partition coefficients between human serum and 1 octanol. In view of the ability of IHP to act as potent effector for oxygen release, the O(2)-hemoglobin dissociation curves were then measured for the most efficient salts on whole blood. From both the biological and the physical properties of IHP-ammonium salts we determined that cycloalkylamines (or poly amines) were the best transport molecules, especially cycloheptyl- and cyclooctylamine. Indeed, the octanol/serum partition coefficient of IHP undecacyclooctylammonium salt, is superior to 1, which is very favorable for potential uptake into the red blood cell membrane. A qualitative correlation was found between the partitioning experiments and the biological evaluations performed on whole blood. PMID- 12110303 TI - DNA damage and biological effects induced by photosensitization with new N(1) unsubstituted furo[2,3-h]quinolin-2(1H)-ones. AB - New furoquinolinones unsubstituted at the N(1) position were prepared and their photobiological activities were studied in comparison with 4,6,8,9 tetramethylfuro[2,3-h]quinolin-2(1H)-one (HFQ) and 8-MOP. The anti-proliferative activity of furoquinolinones 3a-f was tested upon UVA irradiation in mammalian cells, studying DNA synthesis and clonal growth capacity, and in micro-organisms, evaluating T2 infectivity. Almost all compounds appeared to be more active than 8 MOP, and free of any mutagenic activity and skin phototoxicity. Among them, compound 3b was the most effective one. Similarly to HFQ, compound 3b appeared to be very active also in DNA damaging, forming monoadducts and DPC(L=0), but no ISC and DPC(L>0), both responsible for furocoumarin genotoxicity and phototoxicity. Moreover, Ehrlich ascites cells, photoinactivated by the new furoquinolinone 3b and injected into recipient mice, proved to be capable of inducing protection against a successive challenge performed with the same tumor cells. For all these features, 3b seemed to be a new promising potential drug for PUVA therapy and photopheresis. PMID- 12110304 TI - Synthesis and cytotoxic evaluation of analogues of the marine pyridoacridine amphimedine. AB - 4-Substituted-7H-pyrido-[4,3,2-de][1,8] or [1,9]-phenanthroline-7-ones and 9 methyl-1,4-diazanaphtacene-3,10-dione, analogues of the marine pyridoacridine amphimedine were synthesised from isoquinoline-5,8-dione. The first compounds were obtained starting from a Diels-Alder reaction whereas the synthesis of the last compound was initiated by a reaction of condensation with 2 aminoacetophenone. The different tetra- and pentacyclic compounds were evaluated for in vitro cytotoxic activities against six distinct human cancer cell lines. All the compounds exhibit cytotoxic activity with IC(50) values (i.e., the drug concentration inhibiting the mean growth value of the six cell lines by 50%)<10( 7)M for two of them. PMID- 12110305 TI - Sesquiterpene lactones as inhibitors of human neutrophil elastase. AB - Human neutrophil elastase (HNE) is a serine protease that has been implicated in the abnormal turnover of connective tissue proteins and has been described as an important pathogenic factor in several inflammatory diseases such as rheumatoid arthritis or cystic fibrosis. Here we investigated 17 sesquiterpene lactones (SLs) for their ability to inhibit human neutrophil elastase in an in vitro assay. Podachaenin was the most active compound with an IC(50) value of 7 microM. SLs do not covalently bind to the amino acids of the catalytic triad, thus differing from other elastase inhibitors with a lactone moiety. In contrast to most other biological activities of SLs HNE inhibition is not mediated by alpha,beta-unsaturated carbonyl functions. Ligand binding calculations using the X-ray structure of HNE and the program FlexX revealed structural elements which are a prerequisite for their inhibitory activity. PMID- 12110306 TI - (Z)-1,4-diamino-2-butene as a vector of boron, fluorine, or iodine for cancer therapy and imaging: synthesis and biological evaluation. AB - Polyamine vectors are attractive for tumor targeting. We envisaged (Z)-1,4 diamino-2-butene (Z-DAB), an unsaturated analogue of putrescine as vector of (10)B, (18)F and (131)I for boron neutron capture therapy (BNCT), and tumor imaging by positron emission tomography or scintigraphy respectively. In the present work, the synthesis and characterization of new derivatives of Z-DAB were reported. Z-DAB was actively transported in cells via the polyamine transport system and converted into the spermidine analogue.(E)-2-iodo-1,4-diamino-2-butene (E-I-DAB) was not taken up by the polyamine transport system and may not be suitable for tumor imaging. In contrast, (Z)-2-[4-(5,5-dimethyl-dioxaborinan-2 yl)phenyl]methyl-1,4-diamino-2-butene (Z-4-Bbz-DAB) was a substrate of the transport system and allowed significant boron accumulation in 3LL cells. Its potential in BNCT will be evaluated. PMID- 12110307 TI - Pyridazines. Part XXIX: synthesis and platelet aggregation inhibition activity of 5-substituted-6-phenyl-3(2H)-pyridazinones. Novel aspects of their biological actions. AB - A series of 6-phenyl-3(2H)-pyridazinones with a diverse range of substituents in the 5-position have been prepared and evaluated in the search for new antiplatelet agents. A significant dependence of the substituent on the inhibitory effect has been observed. The pharmacological study of these compounds confirms that modification of the chemical group at position 5 of the 6-phenyl 3(2H)-pyridazinone system influences both variations in the antiplatelet activity and the mechanism of action. PMID- 12110308 TI - Molecular docking and 3-D-QSAR studies on the possible antimalarial mechanism of artemisinin analogues. AB - Artemisinin (Qinghaosu) is a natural constituent found in Artemisia annua L, which is an effective drug against chloroquine-resistant Plasmodium falciparum strains and cerebral malaria. The antimalarial activities of artemisinin and its analogues appear to be mediated by the interactions of the drugs with hemin. In order to understand the antimalarial mechanism and the relationship between the physicochemical properties and the antimalarial activities of artemisinin analogues, we performed molecular docking simulations to probe the interactions of these analogues with hemin, and then performed three-dimensional quantitative structure-activity relationship (3-D-QSAR) studies on the basis of the docking models employing comparative molecular force fields analysis (CoMFA) and comparative molecular similarity indices analysis (CoMSIA). Molecular docking simulations generated probable 'bioactive' conformations of artemisinin analogues and provided a new insight into the antimalarial mechanism. The subsequent partial least squares (PLS) analysis indicates that the calculate binding energies correlate well with the experimental activity values. The CoMFA and CoMSIA models based on the bioactive conformations proved to have good predictive ability and in turn match well with the docking result, which further testified the reliability of the docking model. Combining these results, that is molecular docking and 3-D-QSAR, together, the binding model and activity of new synthesized artemisinin derivatives were well explained. PMID- 12110309 TI - Synthesis of new 2,5-disubstituted-1,3,4-thiadiazoles and preliminary evaluation of anticonvulsant and antimicrobial activities. AB - Two new series of 2,5-disubstituted-1,3,4-thiadiazoles were synthesized for their possible anticonvulsant, antibacterial and antifungal activities. The degree of protection afforded by these compounds at a dose of 100mg/kg i.p. against pentylenetetrazole-induced convulsions in mice ranged from 0 to 90%. Among these compounds, 2a (90%) and 2g (70%) showed maximum protection. Antimicrobial tests showed that the MIC value of 3j against Pseudomanas aeruginosa was equal to that of penicillin. PMID- 12110310 TI - In vitro antitumor activities of 2,6-di-[2-(heteroaryl)vinyl]pyridines and pyridiniums. AB - The in vitro antitumor activities of 2,6-di-[2-(heteroaryl)vinyl]pyridines versus the standard National Cancer Institute 60 cell lines panel and of 2,6-di-[2 (heteroaryl)vinyl] pyridinium cations versus MCF7 (human mammary carcinoma) and LNCap (prostate carcinoma) cell lines are reported. Antiproliferative effects in both series are particularly evident for MCF7 mammary adenocarcinoma cells. Multivariate analysis of DNA microarray data for responsive tumor cell lines suggest a mechanistic pathway involving polyamine biosynthesis and prolactin signal transduction. PMID- 12110311 TI - Novel potent substance P and neurokinin A receptor antagonists. Conception, synthesis and biological evaluation of indolizine derivatives. AB - Exploration of SAR around dual NK(1)/NK(2) antagonist Cbz-Gly-Leu-Trp OBzl(CF(3))(2) and its derivatives disclosed the essential requirements for more potent dual NK(1)/NK(2) binding. We report here the synthesis and the biological properties of a novel series of indolizine including pharmacophoric elements. PMID- 12110312 TI - QSAR study on competition binding of rodenticides (PATs) to H(1) receptor in rat and guinea pig brain. AB - A quantitative structure-activity relationship (QSAR) study on binding activity for a series of rodenticides (PAT analogues) to the [(3)H]-mepyramine-labeled H(1) receptor in rat and guinea pig brain is attempted topologically. From the pool of molecular descriptors we, observed that the most significant descriptor is the molecular redundancy index (MRI). Multiple regression analysis gave excellent results with MRI upon introduction of some dummy parameters (indicator parameters). Predictive ability of the proposed models is discussed using cross validation parameters. PMID- 12110313 TI - QSAR studies on biological activity of piritrexim analogues against pc DHFR. AB - Quantitative structure-activity relationship (QSAR) studies for 2,6-substituted 2,4-diaminopyrido[3,2-d] pyrimidine analogues of piritrexim (PTX) as inhibitors of dihydrofolate reductase (DHFR) are now made using topological indices. The results have shown that best models are obtained by multiparametric analysis. The predictive potential of the model is discussed on the basis of a cross-validation method. PMID- 12110314 TI - Design, synthesis, and biological evaluation of a cephalosporin monohydroguaiaretic acid prodrug activated by a monoclonal antibody-beta lactamase conjugate. AB - A novel cephalosporin derivative of monohydroguaiaretic acid (cephem-M(3)N, 7) was synthesized and found to possess anticancer activity against human leukemia (K562), breast carcinoma (MCF7), human lung cancer (A549), human colon cancer (Colo205) and pancreatic cancer cells (Capan2 and MiaPaCa2). A tumor targeting fusion protein (dsFv3-beta-lactamase) was also used in conjunction with cephem based M(3)N 7 and its potency toward K562, MCF7, A549, Colo205, Capan2, and MiaPaCa2 was found to approach that of the free M(3)N (4). In the presence of dsFv3-beta-lactamase, tumor cells were found to be much more susceptible to conjugate 7 than normal human embryonic lung (HEL) cells and normal fibroblasts (Hef522). These notions provide a new approach to the use of nordihydroguaiaretic acid (NDGA) and its derivatives for antitumor therapy. PMID- 12110315 TI - Nucleosides and nucleotides. Part 214: thermal stability of triplexes containing 4'alpha-C-aminoalkyl-2'-deoxynucleosides. AB - In order to develop novel antigene molecules forming thermally stable triplexes with target DNAs and having nuclease resistance properties, we synthesized oligodeoxynucleotides (ODNs) with various lengths of aminoalkyl-linkers at the 4'alpha position of thymidine and the aminoethyl-linker at the 4'alpha position of 2'-deoxy-5-methylcytidine. Thermal stability of triplexes between these ODNs and a DNA duplex was studied by thermal denaturation. The ODNs containing the nucleoside 2 with the aminoethyl-linker or the nucleoside 3 with the aminopropyl linker thermally stabilized the triplexes, whereas the ODNs containing the nucleoside 1 with the aminomethyl-linker or the nucleoside 4 with the 2-[N-(2 aminoethyl)carbamoyl]oxy]ethyl-linker thermally destabilized the triplexes. The ODNs containing 2 were the most efficient at stabilizing the triplexes with the target DNA. The ODNs containing 4'alpha-C-(2-aminoethyl)-2'-deoxy-5 methylcytidine (5) also efficiently stabilized the triplexes with the target DNA. Stability of the ODN containing 5 to nucleolytic hydrolysis by snake venom phosphodiesterase (a 3'-exonuclease) was studied. It was found that the ODN containing 5 was more resistant to nucleolytic digestion by the enzyme than an unmodified ODN. In a previous paper, we reported that the ODNs containing 2 were more resistant to nucleolytic digestion by DNase I (an endonuclease) than the unmodified ODNs. Thus, it was found that the ODNs containing 4'alpha-C-(2 aminoethyl)-2'-deoxynucleosides were good candidates for antigene molecules. PMID- 12110316 TI - Novel furano analogues of duocarmycin C1 and C2: design, synthesis, and biological evaluation of seco-iso-cyclopropylfurano[2,3-e]indoline (seco-iso-CFI) and seco-cyclopropyltetrahydrofurano[2,3-f]quinoline (seco-CFQ) analogues. AB - The design, synthesis and biological evaluation of novel seco-iso cyclopropylfurano[2,3-e]indoline (seco-iso-CFI) and the seco cyclopropyltetrahydrofurano[2,3-f]quinoline (seco-CFQ) analogues of the duocarmycins are described. These novel analogues (4-7) were designed on the premise that the lone pair of electrons on the furano-oxygen atom could enter into conjugation with the isocyclopropylfurano[e]indolone (iso-CFI) alkylating moiety, formed from the loss of HCl in compounds 4-7. The seco-iso-CFI DNA alkylating pharmacophore was synthesized through a well precedented approach of 5 exo-trig aryl radical cyclization with a vinyl chloride. In our studies, in addition to the formation of the seco-iso-CFI product, an equal amount of an unexpected seco-CFQ product was also generated during the radical cyclization reaction. Like CC-1065 and adozelesin, using Taq DNA polymerase stop and thermal cleavage assays, the seco-iso-CFI compounds (4 and 6) and the seco-CFQ compounds (5 and 7) were shown to preferentially alkylate the adenine-N3 position within the minor groove of long stretches of A residues. A MM2 energy optimized molecular model of a 1:1 complex of compound 6 with DNA reveals that the iso-CFI compound fits snugly within the minor groove. Using a MTT based experiment, the cytotoxicity of compounds 4-7 were determined against the growth of murine leukemia (L1210), mastocytoma (P815) and melanoma (B16) cell lines. The concentrations of compounds required to inhibit the growth of these tumor cells by 50% is in the range of 10(-8)M. These compounds were also tested against a panel of human cancer cells by the National Cancer Institute, demonstrating that the compounds exhibited a high level of activity against selected solid tumors. At a concentration of 0.0084 microM (based on the IC(50) of compound 17 (seco-CBI TMI) against the growth L1210 cells), while compounds 4 and 17 were toxic against murine bone marrow cells as judged by a colony forming study of freshly isolated murine progenitor hematopoeitic cells, compound 5, a seco-CFQ compound, was significantly less toxic. Flow cytometric analysis of P815 cells that had been incubated for 24h with compounds 4 and 5 at their cytotoxic IC(50) concentrations indicated the induction of apoptosis in a large percentage of cells, thereby suggesting that this might be the mechanism by which the iso-CFI compounds kill cells. PMID- 12110317 TI - Molecular modeling of 3-arylisoquinoline antitumor agents active against A-549. A comparative molecular field analysis study. AB - A series of 58 3-arylisoquinoline antitumor agents were investigated for defining the pharmacophore model using comparative molecular field analysis (CoMFA) program. The studied compounds related to bioisostere of benzophenanthridine alkaloid were synthesized and evaluated for antitumor cytotoxicity against human lung tumor cell (A 549). In order to perform the systematic molecular modeling study of these compounds, the conformational search was carried out based on the single X-ray crystallographic structure of 7,8-dimethoxy-3-phenylisoquinolin-(2H) one (2). Interestingly, two types of structures having different dihedral angles between the isoquinoline ring and 3-aryl ring were found in the crystals. Therefore, CoMFA was performed two different, overlapping ways. The alignments of the structures were based on the common isoquinoline ring and 3-aryl ring. The 3 D-quantitative structure-activity relationship study resulted in significant cross-validated, conventional r(2) values equal to 0.715 and 0.927, respectively. PMID- 12110318 TI - Structure-activity relationships of benzylidene anabaseines in nicotinic acetylcholine receptors of cockroach nerve cords. AB - Ten analogues of 6'-chloro-3-benzylideneanabaseine (CBA) bearing substituents at the ortho- and the para-positions of the phenyl group were synthesized, together with two related compounds. The affinity of the synthesized compounds for nicotinic acetylcholine receptors (nAChRs) in the nerve cord of the American cockroach (Periplaneta americana L.) was examined by the radioligand binding assay using [(3)H]epibatidine (EPI), a nAChR agonist. All 12 tested compounds inhibited [(3)H]EPI binding, showing K(i) values ranging from 14.6 to 6830nM. The potency variation of para-substituted CBA analogues was explained by the steric (Delta B(1)) and electronic (sigma(p)) parameters of the para-substituents, or by the steric parameter and the charge of the N1 nitrogen atom (qN(1)). Among the CBA analogues, only two compounds containing a dimethylamino group and a methoxy group at the para-position showed high insecticidal activity against the German cockroach (Blattella germanica) when injected after pretreatment with metabolic inhibitors. High-affinity analogues of CBA might be suitable probes for use in classifying and characterizing insect nAChR subtypes. PMID- 12110319 TI - Adenine and deazaadenine nucleoside and deoxynucleoside analogues: inhibition of viral replication of sheep MVV (in vitro model for HIV) and bovine BHV-1. AB - A series of N(6)-cycloalkyl-2',3'-dideoxyadenosine derivatives has been prepared by coupling of 2,6-dichloropurine to protected 2,3-dideoxyribose, followed by reaction with appropriate cycloalkylamines. Synthesized compounds, along with other purine nucleoside analogues previously synthesized in our laboratory, have been tested for their antiviral activity against Bovine herpesvirus 1 (BHV-1) and sheep Maedi/Visna Virus (MVV), the latter being an in vitro and in vivo model of Human Immunodeficiency Virus (HIV). All compounds showed good antireplicative activity against MVV, with the N(6)-cycloheptyl-2',3'-dideoxyadenosine (5b) being the most active [effective concentration (EC(50)) causing 50% reduction of cytopatic effects (CPE)=27 nM]. All compounds showed also a from low to very low cell toxicity, resulting in a cytotoxic dose 50 (CD(50))/EC(50) ratio in some cases higher than 1000. PMID- 12110320 TI - CYP3A4 inhibitory activity of new bisalkaloids, dipiperamides D and E, and cognates from white pepper. AB - Two new bisalkaloids, dipiperamides D and E, were isolated as inhibitors of a drug metabolizing enzyme cytochrome P450 (CYP) 3A4 from the white pepper, Piper nigrum. Their structures were elucidated by spectroscopic methods. Dipiperamides D and E showed potent CYP3A4 inhibition with IC(50) values of 0.79 and 0.12 microM, respectively, and other metabolites from the pepper were moderately active or inactive. PMID- 12110322 TI - QSAR studies on carbonic anhydrase inhibitors: a case of ureido and thioureido derivatives of aromatic/heterocyclic sulfonamides. AB - QSAR studies on modelling of biological activity (hCAI) for a series of ureido and thioureido derivatives of aromatic/heterocyclic sulfonamides have been made using a pool of topological indices. Regression analysis of the data showed that excellent results were obtained in multiparametric correlations upon introduction of indicator parameters. The predictive abilities of the models are discussed using cross-validation parameters. PMID- 12110321 TI - Hydrazinocurcumin, a novel synthetic curcumin derivative, is a potent inhibitor of endothelial cell proliferation. AB - Curcumin and some of its derivatives were known as in vivo inhibitors of angiogenesis. In present study, a novel curcumin derivative, named hydrazinocurcumin (HC) was synthesized and examined for its biological activities. HC potently inhibited the proliferation of bovine aortic endothelial cells (BAECs) at a nanomolar concentration (IC(50)=520 nM) without cytotoxicity. In vivo and in vitro angiogenesis experiments showed HC as a new candidate for anti-angiogenic agent. PMID- 12110323 TI - Structural studies on bioactive compounds. Part 36: design, synthesis and biological evaluation of pyrimethamine-based antifolates against Pneumocystis carinii. AB - As part of a research effort to improve the quality of current chemotherapy of Pneumocystis carinii pneumonia, we report a structure-based design project to optimise activity, species selectivity and pharmaceutical properties of the triazenyl-pyrimethamine TAB (4) (IC(50)=0.17 microM; rat liver DHFR IC(50)/P. carinii DHFR IC(50)=114). This has led us to design, synthesise and evaluate four new series of pyrimethamine derivatives bearing triazole, triazolium, triazinium and amino moieties at the 3'-position of the p-chlorophenyl ring. Such stabilised 'triazene' derivatives address the potentially compromised pharmaceutical profile of TAB and the 3'-amine substituted agents afford conformationally flexible substitutes. The benzylamino-pyrimethamine derivative (24a) (IC(50)=0.12 microM, rat liver DHFR IC(50)/P. carinii DHFR IC(50): 5.26) was the most potent and the only P. carinii-selective antifolate of the new series. PMID- 12110324 TI - Superoxide dismutase mimetics: synthesis and structure-activity relationship study of MnTBAP analogues. AB - Carboxylic ester and amide-substituted analogues of [5,10,15,20-tetrakis(4 carboxyphenyl)-porphyrinato]manganese(III) chloride (MnTBAP) were synthesized and assayed as potential superoxide dismutase (SOD) mimetics. The tetraester analogues 4a and 4b were found to have comparable SOD activity to the known SOD mimetic MnTBAP, while amides 4c-4e exhibited reduced SOD activity. In the substituted methyl benzoate/acid and disubstituted porphyrin series, analogues 12c, 12f, and 12m were found to have comparable to improved SOD activity relative to MnTBAP and analogues 12j, 13a, and 13d exhibited improved activity in both the SOD and thiobarbituric acid reactive species (TBARS) assays relative to MnTBAP. PMID- 12110325 TI - Synthesis and characterization of a catalytic antibody-HPMA copolymer-Conjugate as a tool for tumor selective prodrug activation. AB - Selective chemotherapy remains a key issue for successful treatment in cancer therapy. The use of targeting approaches like the enhanced permeability and retention (EPR) effect of macromolecules, is consequently needed. Here, we report the preparation of a novel catalytic antibody-polymer conjugate for selective prodrug activation. HPMA copolymer was conjugated to catalytic antibody 38C2 through an amide bond formation between epsilon-amino group of lysine residue from the antibody molecule and a p-nitrophenyl ester of the polymer. The conjugate was purified over a size exclusion column using FPLC. In the isolated fraction, one or two molecules of polymer were conjugated to one molecule of antibody based on gel analysis. The resulting conjugate retained most of its catalytic activity (75-81%) in comparison to the free antibody. The activity was monitored with a fluorogenic substrate and a prodrug activation assay using HPLC. Furthermore, the conjugate was evaluated in vitro for its ability to activate an etoposide prodrug using two different cancer cell lines. Cells growth inhibition using the prodrug and the conjugate was almost identical to inhibition by the free antibody and the prodrug. For the first time, a catalytic antibody was conjugated to a passive targeting moiety while retaining its catalytic ability to activate a prodrug. The conjugate described in this work can be used for selective activation of prodrug in the PDEPT (polymer directed enzyme prodrug therapy) approach by replacing the enzyme component with catalytic antibody 38C2. PMID- 12110326 TI - Synthesis and biological evaluation of MAO-A selective 1,4-disubstituted-1,2,3,6 tetrahydropyridinyl substrates. AB - Many mammalian tissues express both the A and B forms of monoamine oxidase (MAO), flavoenzymes that catalyze the oxidative deamination of a variety of endogenous and exogenous amines and the ring alpha-carbon oxidative bioactivation of neurotoxic 1,4-disubstituted-1,2,3,6-tetrahydropyridinyl derivatives. Substrates selective for MAO-A that display good kinetic and spectroscopic properties would be of value for developing quantitative assays for MAO-A in tissues that express both the A and B forms of the enzyme. This paper describes the synthesis of several 1-substituted-4-(1-methylpyrrol-2-yl)-1,2,3,6-tetrahydropyridinyl derivatives. Kinetic parameters and MAO-A selectivity indicate that 1-allyl- and 1-propyl-4-(1-methylpyrrol-2-yl)-1,2,3,6-tetrahydropyridine should be good candidates to develop a robust spectrophotometric-based assay that is selective for MAO-A. PMID- 12110327 TI - Synthesis and biological activity of novel neuroprotective diketopiperazines. AB - The cyclic dipeptide cyclo[His-Pro] (CHP) is synthesized endogenously de novo and as a breakdown product of thyrotropin-releasing hormone (TRH), a tripeptide with known neuroprotective activity. We synthesized two isomeric compounds based on the structure of CHP, in which the histidine residue was replaced by 3,5-di-tert butyltyrosine (DBT), a phenolic amino acid that traps reactive oxygen species. These novel diketopiperazines prevented neuronal death in an in vitro model of traumatic injury. In addition, they dose-dependently prevented death caused by the direct induction of free radicals, and by calcium mobilization through an agent that evokes rapid, necrotic death. The drugs showed activity in the latter system at picomolar concentrations. The neuroprotective profile of these compounds suggests that they may be useful as treatments for neuronal degeneration in vivo, potentially through several different mechanisms. PMID- 12110328 TI - Novel substrates for nitric oxide synthases. AB - Enzymatic generation of nitric oxide (NO) by nitric oxide synthase (NOS) consists of two oxidation steps. The first step converts L-arginine to N(G)-hydroxy-L arginine (NOHA), a key intermediate, and the second step converts NOHA to NO and L-citrulline. To fully probe the substrate specificity of the second enzymatic step, an extensive structural screening was carried out using a series of N-alkyl (and N-aryl) substituted-N'-hydroxyguanidines (1-14). Among the eleven N-alkyl-N' hydroxyguanidines evaluated, N-n-propyl (2), N-iso-propyl (3), N-n-butyl (4), N-s butyl (5), N-iso-butyl (6), N-pentyl (8) and N-iso-pentyl (9) derivatives were efficiently oxidized by the three isoenzymes of NOS (nNOS, iNOS and eNOS) to generate NO. N-Butyl-N'-hydroxyguanidine (4) was the best substrate for iNOS (K(m)=33 microM) and N-iso-propyl-N'-hydroxyguanidine (3) was the best substrate for nNOS (K(m)=56 microM). When the alkyl substituents were too small (such as ethyl 1) or too large (such as hexyl 10 and cyclohexyl 11), the activity decreased significantly. This suggests that the van der Waals interaction between the alkyl group and the hydrophobic cavity in the NOS active site contributes significantly to the relative reactivity of compounds 3-11. Moreover, five N-aryl N'-hydroxyguanidines were found to be good substrates for iNOS, but not substrates for eNOS and nNOS. N-phenyl-N'-hydroxyguanidine was the best substrate among them (K(m)=243 microM). This work demonstrates that N-alkyl substituted hydroxyguanidine compounds are novel NOS substrates which 'short-circuit' the first oxidation step of NOS, and N-aryl substituted hydroxyguanidine compounds are isoform selective NOS substrate. PMID- 12110329 TI - Effects of 4,4-dimethyl-5,8-dihydroxynaphtalene-1-one and 4,4-dimethyl-5,8 dihydroxytetralone derivatives on tumor cell respiration. AB - A set of structurally related compounds incorporating a carbonyl group in the ortho position with regard to a phenol function were tested against the TA3 mouse carcinoma cell line and its multidrug-resistant variant TA3-MTX-R. The series consists of 2'-hydroxyacetophenone, 4'-hydroxyacetophenone 2',5' dihydroxyacetophenone, 4-acetyl-3,3-dimethyl-5-hydroxy-2-morpholino-2,3 dihydrobenzobfuran, five 4,4-dimethyl-5,8-dioxygenated naphtalene-1-ones and three 4,4-dimethyl-5,8-dioxygenated tetralones. A tentative structure-activity relationship was found for this family of substances, suggesting that a coplanar ortho-carbonyl-1,4-hydroquinone motif is able to cause inhibition of cellular respiration. PMID- 12110330 TI - Cytotoxic T lymphocyte epitope analogues containing cis- or trans-4 aminocyclohexanecarboxylic acid residues. AB - In order to improve the immunotherapeutical potential of H-Cys-Leu-Gly-Gly-Leu Leu-Thr-Met-Val-OH (CLG) peptide, an Epstein-Barr virus (EBV) subdominant epitope derived from the membrane protein LMP2, we have synthesized and tested CLG analogues containing cis- and/or trans-4-aminocyclohexanecarboxylic acid (ACCA) replacing Gly-Gly and/or Thr-Met dipeptide units. All pseudopeptides were tested for metabolic stability and for their capacity to bind HLA-A2 molecules and to sensitize target cells to lysis. All new compounds exhibited higher enzymatic resistance compared to the original CLG and some trans-ACCA-derivatives were able to associate HLA-A2 and to efficiently stimulate CTL responses directed against the CLG natural epitope. PMID- 12110331 TI - Synthesis and anti-inflammatory activity of phthalimide derivatives, designed as new thalidomide analogues. AB - This paper describes the synthesis and anti-inflammatory activity of new N-phenyl phthalimide sulfonamides (3a-e) and the isosters N-phenyl-phthalimide amides (4a e), designed as hybrids of thalidomide (1) and aryl sulfonamide phosphodiesterase inhibitor (2). In these series, compound 3e (LASSBio 468), having a sulfonyl thiomorpholine moiety, showed potent inhibitory activity on LPS-induced neutrophil recruitment with ED(50)=2.5mg kg(-1), which was correlated with its inhibitory effect on TNF-alpha level. PMID- 12110333 TI - The aggressive nature of the odontogenic keratocyst: is it a benign cystic neoplasm? Part 3. Immunocytochemistry of cytokeratin and other epithelial cell markers. AB - Numerous studies of keratin expression by the more common odontogenic cysts were done to determine whether patterns of cytokeratin staining could provide accurate diagnostic markers for the different varieties; to see whether comparative studies with oral mucosa and developing odontogenic epithelium could explain the pathogenesis of the cysts; and whether cytokeratin patterns could provide clues in elucidating the aggressive nature of the OKC. This review was a complex task with a range of at least 19 different cytokeratins being studied and also a broad range of antibodies in use for the same cytokeratin or group of cytokeratins. Moreover, there was not always standardisation of laboratory techniques in the selection and preparation of material. These difficulties were, in general, recognised by the different workers in the field, particularly when there was disagreement on results and caution was expressed about drawing conclusions from some positive findings. It would be fair to conclude that cytokeratin immunocytochemistry has not advanced to any meaningful extent, its use as a diagnostic marker for the OKC nor in eludidating its pathogenesis. With regard to OKC behaviour, it has been pointed out that there was strong reaction of OKC lining for keratin 16, a cytokeratin that has been associated with high proliferative activity. Yet other studies have also shown keratin 16 expression in dentigerous and radicular cysts. Differences in cytokeratin, EMA and CEA immunocytochemical reactivity between the parakeratinised and orthokeratinised varieties of cyst were demonstrated and the suggestion made that the orthokeratinised type has a considerably less aggressive behaviour, is a different entity and should bear a different name. Furthermore, Ki67 positive cells in the parakeratinised OKC linings were considerably more frequent than in the orthokeratinised linings.OKC, dentigerous and radicular cyst epithelium reacted positively for epithelial growth factor receptor (EGFr) but a trend indicating the most intense staining in the OKCs, followed by the dentigerous and then the radicular cyst linings led to the conclusion that the OKCs have an intrinsic growth potential not present in other odontogenic cysts. PMID- 12110334 TI - Maryland family physicians' knowledge, opinions and practices about oral cancer. AB - The objective of this study was to assess family physicians' knowledge, opinions and practices regarding oral cancers in the state of Maryland, USA. A 40-item, self-administered questionnaire was mailed to all members of the Maryland Academy of Family Physicians. Unweighted data (n=240) were analyzed using SAS and SUDAAN software; results were evaluated using an alpha < or =0.05. Family physicians (FPs) were aware of the major risk factors for oral cancers, but misinformation existed about the non-risk factors. Approximately 77% asked their patients the eight questions related to risk factors for oral cancer when taking a medical history but less than 24% provided an oral cancer examination to patients 40 years of age and over. Nearly 64% were interested in a continuing education course about oral cancer. This survey identified gaps in knowledge and practices among FPs but it is encouraging that they expressed interest in continuing education courses on this topic. PMID- 12110335 TI - Elevated vimentin expression in buccal mucosal fibroblasts by arecoline in vitro as a possible pathogenesis for oral submucous fibrosis. AB - Areca quid chewing is strongly correlated with oral submucous fibrosis (OSF) in Taiwan. The cytotoxicity of arecoline, a major areca nut alkaloid, on human oral fibroblasts has been extensively studied. To date, however, there has been little research exploring the possible effects of arecoline on cytoskeleton components. In this study, in addition to conducting a cytotoxicity assay, we examine the effect of arecoline on vimentin, an intermediate filament, and its expression in human buccal mucosal fibroblasts on exposure to various levels of arecoline (0 200 microg/ml) for 48 h. At a concentration above 50 microg/ml, arecoline demonstrated dose-dependent cytotoxicity (P<0.05) for cultured fibroblasts. Using sodium dodecyl sulphate-polyacrylamide gel electrophoresis, we demonstrated dose dependent elevation of 57 kDa cytoskeletal-protein levels for arecoline. Evidence from immunoblotting assay indicated this 57 kDa cytoskeletal protein was vimentin. The increase in vimentin with arecoline exposure corresponded to that noted for fibroblasts cultured from OSF patients. Immunohistochemical assay also revealed that vimentin expression was much higher for OSF specimens than for normal buccal mucosa. We suggest these results may advance understanding of the possible pathogenesis for submucous fibrosis through the transformation of normal buccal mucosa as a result of areca quid chewing. PMID- 12110336 TI - Garlic induces apoptosis during 7,12-dimethylbenz[a]anthracene-induced hamster buccal pouch carcinogenesis. AB - The apoptosis-inducing capacity of aqueous garlic extract during 7,12 dimethylbenz[a]anthracene-induced hamster buccal pouch (HBP) carcinogenesis was investigated in male Syrian hamsters using DNA fragmentation and the apoptosis associated proteins, tissue transglutaminase (tTG) and Bcl-2. Hamsters were divided into four groups of six animals each. Animals in group 1 were painted with a 0.5% solution of DMBA in liquid paraffin on the right buccal pouches three times a week for 14 weeks. Group 2 animals painted with DMBA as in group 1, in addition received 250 mg/kg body weight aqueous garlic extract orally on days alternate to DMBA application. Group 3 animals received garlic extract as in group 2. Group 4 animals received neither DMBA nor garlic extract and served as the control. The experiment was terminated at the end of 14 weeks. Administration of aqueous garlic extract (250 mg/kg body weight) to animals painted with DMBA inhibited DMBA-induced oral carcinogenesis as revealed by the absence of neoplasms, induction of tTG and inhibition of Bcl-2 expression. The results of the present study suggest that garlic may exert its chemopreventive effect by inducing apoptosis. PMID- 12110337 TI - Immunohistochemical distinction of high-grade mucoepidermoid carcinoma and epidermoid carcinoma of the parotid region. AB - The correct diagnosis of high-grade mucoepidermoid (MEC), which is composed of solid islands of intermediate and squamous cells, may be challenging, due to its similarity to other tumours, mainly with squamous cell carcinoma (SCC). The present report employed immunohistochemical technique against different cytokeratins (CKs), in order to differentiate these two entities. : Six high grade MEC and six SCC of the parotid region, retrieved from the files of both Oral Pathology Department of the School of Dentistry of University of Sao Paulo and Pathology Department of A.C. Camargo Hospital, were submitted immunohistochemical technique against Cks 7,8, 10, 13, 14 and 19. : High-grade MEC was positive for Cks 7, 8, 13, 14 and 19. The cases of SCC showed strong positivity for CK14, and CK10 was present only in focal areas. Our results highlight the use of CKs (especially CK14) to differentiate high-grade MEC and SCC. PMID- 12110338 TI - Cellular basis and clinical implications of biological markers in salivary tissues: their topological distribution in murine submandibular gland. AB - Cell proliferation and apoptosis as well as cell-cell adhesion and communication are essential processes that assure cell survival, renewal and coordination. Since junctional proteins have a tumor suppressor activity, their immunohistochemical characterization has diagnostic and prognostic value. The purpose of this report is to review the role played by junctional and proliferation-related proteins in the salivary glands and to illustrate their immunohistochemical localisation in normal murine submandibular gland. Normal salivary gland tissue was obtained from normal adult male BALB/c mice. After immediate fixation in formalin and ethanol, the samples were immunohistochemically stained for E-cadherin (HECD-1), Bcl-2, Ki67 (MIB-1), connexin26 and connexin 32, beta-catenin and gamma-catenin. Their topological distribution and reactivity were evaluated by light microscopy. The nuclei of submandibular acinar cells exhibited low to moderate staining for Ki67, but no reaction was observed in ductal cells. Murine Bcl-2 was light to moderately expressed in the latero-basal domain of cells of submandibular acini but was only lightly expressed in striated and eosinophilic ducts. The lateral domain of acinar cells were heavily stained with anti-E-cadherin, while only low levels were expressed at the cellular surface of ducts. beta-Catenin was consistently and evenly distributed along the latero-apical boundaries of eosinophilic secretory duct cells as well as on the lateral domain of acinar cells. On the contrary, gamma-catenin was generally expressed at lower levels than beta catenin, was not expressed in ductal cells and was only lightly stained on the lateral membranes of acinar cells. No expression of connexin 32 was observed in ducts but it was significantly expressed in a spotted pattern along the plasma membrane of acinic cells. Connexin 26 showed similar localization to that of connexin 32 but the staining was much more intense. Since these proteins have been reported to play key roles in maintaining homeostasis via control of cell growth, differentiation and death, their analysis in normal salivary tissue will hopefully contribute to the study of salivary tumorigenesis. PMID- 12110339 TI - Rhabdomyosarcoma of the head and neck in children. AB - OBJECTIVE: The purpose of this study was to determine the rhabdomyosarcoma types involving the head and neck (H&N) region in children and their immunophenotype. DESIGN: Anatomic pathology archives at Texas Children's Hospital were searched for all rhabdomyosarcoma cases over a 20-year period. One-hundred and thirty seven cases were identified, with 50 being H&N cases. The cases were typed according to the Intergroup Rhabdomyosarcoma Study (IRS) criteria. Immunocytochemistry for myogenic and non-myogenic markers was performed on all H&N cases. Electron micrographs from cases (n=32) where ultrastructural examination had been performed at the time of original diagnosis were reviewed. RESULTS: Children with H&N rhabdomyosarcomas had a mean age of 5.3 years (median 4 years). There was a male predilection (1.7M:1.0F). Primary tumor sites were: face NOS (18%), orbit/periorbital (16%), nasal cavity/nose (14%), lymph nodes (12%), paranasal sinuses (10%), parameningeal (10%), parotid gland (6%), neck (6%), infratemporal fossa/zygoma (2%), buccal mucosa (2%), palate (2%), and larynx (2%). Metastatic disease at diagnosis (33% of all cases) occurred in the bone marrow (11%), cerebrospinal fluid (6%), peritoneal fluid (6%), lung (4%), parietal pleura (2%), pleural fluid (2%) and pericardial fluid (2%). Rhabdomyosarcoma types (IRS criteria) were: embryonal (60%), alveolar (classic and solid subtypes, 28%), botryoid (4%), undifferentiated (4%), spindle cell (2%) and anaplastic (2%). Immunocytochemical findings were: polyclonal desmin (96%); myogenin (96%); muscle-specific actin (74%), smooth muscle actin (12%). Nonmyogenic markers included: vimentin (100%), CD99 (16%), p53 (16%), pancytokeratin (10%), NSE (8%), LCA (6%), CD20 (6%), EMA (2%), and NB-84 (0%, neuroblastoma). Undifferentiated sarcoma expressed only vimentin. By ultrastructural examination, 44% had readily identified z-bands and myofilaments, 37% had infrequent to rare myofilaments and z-bands, and 19% had myotubular intermediate filaments. CONCLUSIONS: Distribution of H&N rhabdomyosarcoma IRS types is similar to that for all primary sites, with the exception that embryonal types are modestly increased while alveolar type is mildly decreased. There are many non-myogenic immunocytochemical markers that cross-react with rhabdomyosarcoma. Differentiation between favorable and unfavorable rhabdomyosarcoma types is important for appropriate therapy, and predicting prognosis and survival. PMID- 12110340 TI - Intralesional vinblastine vs. 3% sodium tetradecyl sulfate for the treatment of oral Kaposi's sarcoma. A double blind, randomized clinical trial. AB - In this double-blind, randomized trial, we compared the clinical efficacy of intralesional vinblastine (VNB) and 3% sodium tetradecyl sulfate (STS) in the treatment of oral Kaposi's sarcoma (OKS). Subjects with OKS were randomly assigned to receive a single intralesional injection of either VNB or STS, at a standard dose (0.2 mg/cm(2)). Differences were evaluated by the Mann-Whitney U and Fisher's exact tests. Sixteen HIV-infected patients were included, eight received VNB and eight received STS; clinical response was evaluated at days 7, 14, and 28 following treatment. Tumor size reduction was 0.68 and 0.61 cm in the VNB and STS groups, respectively (P=0.80). Two VNB patients had complete or partial response whereas four STS subjects had partial responses (P=0.61). Patients in both groups experienced minimal toxicity. We conclude that intralesional vinblastine or STS are adequate for the management of OKS. The benefits of STS are its low cost and ease of use. PMID- 12110341 TI - Overexpression of extracellular-signal regulated kinases on oral squamous cell carcinoma. AB - Mitogen-activated protein kinases (MAPKs) consist of major three subfamilies, extracellular-signal regulated kinases (ERK MAPKs), the c-Jun N-terminal kinases/stress activated protein kinases (JNK MAPKs/SAP MAPKs), and p38 MAPKs. ERK MAPKs pathway is one of the most important pathways for cell proliferation. ERK MAPKs are located at downstream of a lot of growth factors (epidermal growth factor (EGF), nerve growth factor (NGF), platelet-derived growth factor (PDGF), etc.), the overexpressions and activation of which are frequently detected on a number of cancers including oral squamous cell carcinoma (OSCC). These data indicate that overexpression and activation of ERK MAPKs play an important role in cancer progression. On the contrary, JNK MAPKs are possible regulators of cell death induced by chemotherapeutic agents. p38 MAPKs are activated by pro inflammatory cytokines and inflammatory drugs (non-steroidal anti-inflammatory drug), which are known to suppress cancer growth. These findings imply that each MAPKs can be molecular targets for cancer therapy in OSCC and its investigation is very important things in OSCC. PMID- 12110342 TI - XPD/ERCC2 EXON 8 Polymorphisms: rarity and lack of significance in risk of squamous cell carcinoma of the head and neck. AB - BACKGROUND: Inherited polymorphisms of DNA repair genes may contribute to genetic susceptibility to squamous cell carcinoma of the head and neck (SCCHN). The objective was to assess whether two polymorphisms in the nucleotide excision repair gene XPD (ERCC2) are markers of SCCHN risk. METHODS: We performed a hospital-based case-control study of 180 SCCHN patients and 400 cancer-free controls frequency matched on age, sex, smoking, and alcohol use. All subjects were non-Hispanic whites. XPD alleles 23047 and 23051 were assessed by digestion with the restriction enzymes XhoII and SphI after PCR amplification. RESULTS: The XPD 23047 G and XPD 23051 T alleles were extremely rare among both the cases and controls (allele frequencies<1.0%), and not statistically different between groups (P>0.6). CONCLUSIONS: The 23047 and 23051 variants of the DNA repair gene XPD are extremely rare and do not contribute significantly to the risk of SCCHN in the non-Hispanic white population. PMID- 12110343 TI - Exposure of oral mucosa to bioactive milk factors reduces severity of chemotherapy-induced mucositis in the hamster. AB - A biologically active extract containing bovine whey proteins, whey growth factor extract-A (WGFE-A) was administered topically to the oral mucosa of hamsters and its ability to prevent and treat chemotherapy-induced oral mucositis investigated. Oral mucositis was induced in Syrian golden hamsters through a combination treatment of the antimetabolite chemotherapy drug 5-fluorouracil (5 FU), and mild abrasion of the cheek pouch. WGFE-A administered to the oral mucosa via hydrogel and liquid treatments, pre and concurrent to 5-FU therapy, resulted in significantly reduced mucosal ulceration. The protective effect was dose dependent with greatest benefit from WGFE-A doses applied at 4.2 mg/ml gel and 14 mg/ml mouthwash (P<0.01). The protective activity of WGFE-A also appeared related to mode of delivery. Administration of WGFE-A from an alternate vehicle Orabase(R) did not alleviate mucositis compared to WGFE-A applied in hydrogel. When administered continuously after the chemotherapy schedule, WGFE-A failed to reduce ulcer area when applied over a 12-day period. In a separate study, cell cycle staining indicated that cheek pouch mucosal epithelial cells pre-exposed to WGFE-A in-vivo showed a reduced rate of proliferation, measured as a 21% reduction in the bromodeoxyuridine (BrdU) cell labelling index (P<0.04). This was consistent with a protective mode of WGFE-A action against anti-metabolites such as 5-FU which target rapidly dividing cells. The results were also consistent with recent in vitro data showing protective properties from WGFE-A administered to epithelial cells given pre/concurrent to chemotherapy exposure. WGFE-A is known to contain mitogens which stimulate cells of mesenchymal origin and inhibit epithelial cell growth in culture. Several WGFE-A constituents are likely to confer protective effects on the cheek mucosa, including anti-proliferative, anti apoptotic and anti-microbial factors. WGFE-A provides a potentially valuable source of topically delivered proteins for clinical application in preventing severe oral mucositis caused by chemotherapy. PMID- 12110344 TI - Genetic polymorphisms of drug-metabolizing enzymes and susceptibility to oral cavity cancer. AB - We investigated the association of polymorphisms of drug-metabolizing enzymes and susceptibility to oral cavity cancer. Polymerase chain reaction (PCR)-based analyses were performed on genomic DNA of 94 Caucasian patients in Germany and 92 healthy German controls to determine genotypes of polymorphisms in CYP1A1, GSTM1 and NAT2. For CYP1A1, the homozygous mutant genotype Val/Val did not occur. The heterozygous genotype Ile/Val (6.5% cases versus 4.3% controls) and the homozygous wild-type Ile/Ile (95.7% cases versus 93.5% controls) showed no statistically significant differences between groups (X(2)=0.47; P=0.534, Fisher's exact test, two-sided). The GSTM1 homozygous null genotype occurred more frequently in cancer patients (59.6%) compared to controls (53.3%) but this difference remained insignificant in X(2)-analysis (X(2)=1.07; P=0.587). Almost identical genotype distributions between cases and controls were found for all three NAT2 acetylators. Hence, these three genetic polymorphisms are unlikely to be associated with oral cavity cancer in the population studied. PMID- 12110345 TI - Prognostic factors in betel and tobacco related oral cancer. AB - Paucity of well-defined prognostic molecular markers severely hampers prediction of the clinical course of squamous cell carcinoma (SCC) of oral cavity. The aim of the study was to evaluate the prognostic significance of impairments in the expression of proteins involved in cell cycle regulation and locoregional spread in oral SCC of habitual betel and tobacco chewers. A prospective study was performed in 105 betel and tobacco consumers with oral SCCs during the period 1988-1999, to evaluate the prognostic relevance of impairments in the expression of proteins involved in cell cycle regulation and locoregional spread. Alterations in the expression of p53, pRb, p16, MDM2, p21, and Ets-1 proteins were determined by immunohistochemical analysis in formalin fixed, paraffin embedded tissue sections from oral SCCs. Analysis of multiple molecular biological factors showed overexpression of p53 in 69/105 (66%) cases, MDM2 in 72/105 (69%) cases, p21 in 57/105 (54%) cases and Ets-1 in 64/105 (61%) cases. Loss of pRb was observed in 58/105 (55%) cases and p16 loss was observed in 72/105 (69%) cases. Interestingly, multivariate analysis revealed loss of pRb as the most significant predictor of advanced tumour stage [P=0.001; Odd's Ratio (OR)=3.5] and overexpression of Ets-1 protein was an independent risk factor for lymph node metastasis (OR: 10.1; P<10(-6)). Multiple logistic regression models showed that pRb loss [Hazard's Ratio (HR): 3.93] and p53 overexpression (HR: 2.97) may serve as adverse prognosticators for disease free survival of the patients. The data demonstrate multiple impairments in p53/MDM2/p21/Ets-1 and p16/pRb pathways in betel and tobacco related oral tumourigenesis. PMID- 12110346 TI - Treatment failure and margin status in head and neck cancer. A critical view on the potential value of molecular pathology. AB - Molecular pathology may demonstrate tumour cells not detected by histology. The idea has emerged that these cells influence the prognosis negatively and that their detection will lead to more appropriate treatment and improved patient survival. We theorized that tumour cells at surgical margins overlooked by the pathologist should demonstrate their clinical significance by causing recurrences at the primary site in the patients reported to have tumour-free margins by histology. To assess this assumption, we investigated the prognostic influence of the histologically determined status of the surgical margins. The material that formed the basis of this study consisted of 394 patients that underwent resection for their primary tumour during the years 1990-1995. In 207 patients, initial treatment was complete as assessed by conventional histopathological examination of the surgical specimen. In 187 patients, initial treatment was incomplete, defined as tumour in or close to the margin, or mild, moderate or severe dysplasia or in situ cancer at the margin. Causes for treatment failure were recorded for both groups separately. In the group with tumour-free margins, 16.9% had a second primary head and neck cancer, 8.2% had a second tumour in the lung, 10.6% had recurrent disease in the neck, 2.9% had distant metastasis, and 3.9% had local recurrence at the same site as the primary cancer. For the group without tumour-free margins, these figures were the following: second primary in the head and neck area: 17.1%, second primary in the lung: 7.0%, recurrent disease in the neck: 11.8%, distant metastasis: 8.0% and local recurrence at the primary site: 21.9%. Local recurrences were rare in patients in which the pathologist reported the resection to be complete. Although there may be tumour cells in surgical margins that evade histological detection, their clinical impact appears to be almost negligible. PMID- 12110347 TI - Odontogenic carcinoma: a functional genomic comparison with oral mucosal squamous cell carcinoma. AB - Intraosseous squamous cell carcinomas of the mandible arise de novo or secondary to a tumor or transformed cyst epithelium. Current diagnostic tests frequently fail to distinguish between these tumors, leading to confusing classification schemes. We report the functional genomic analysis of a mandibular odontogenic carcinoma. Malignant keratinocytes from the lesion were isolated using laser capture microdissection. Target sample generated from the total RNA of the LCM procured cells was used to hybridize high-density oligonucleotide arrays. Functional genomic analysis of the odontogenic carcinoma database compared with four oral mucosal squamous cell carcinoma gene expression databases was performed. Preliminary results suggest a small subset of genes distinguish this odontogenic carcinoma from oral mucosal epidermoid carcinomas. PMID- 12110348 TI - Orofacial rhabdomyosarcoma in neonates and young children: a review of literature and management of four cases. AB - Rhabdomyosarcoma (RMS) is an aggressive malignant skeletal muscle neoplasm arising from embryonal mesenchyme. It accounts for over 50% of all pediatric soft tissue sarcomas. The head and neck region is the most common site for this tumor in children. Neonatal presentation of this tumor is rare. We present the management of one neonatal case and three additional cases of orofacial RMS in children under the age of 7 years. All four patients were seen in the department of oral and maxillofacial surgery at Children's Hospital and Regional Medical Center (CHRMC) in Seattle between 1992-2000. Three of the four cases were alveolar RMS and one was botryoid sub-type of embryonal RMS. Three patients were treated with a combination of surgery, chemotherapy and radiation, while the patient with botryoid RMS was treated with surgery and chemotherapy only. The patient with congenital RMS died at 2.5 years of age due to recurrent metastatic disease. The other three patients are alive without evidence of recurrent with a mean follow up was 5.5 years (range 2.5-8.5 years). We discuss the current management, diagnosis, biological behavior, histopathology, prognosis and survival of head and neck RMS in neonates and young children. PMID- 12110349 TI - Minimally differentiated acute myelogenous leukemia (AML-M0) granulocytic sarcoma presenting in the oral cavity. AB - Acute myelogenous leukemia with minimal differentiation (AML-M0) is a rare subtype of acute leukemia in which blasts fail to show morphologic differentiation and conventional cytochemical stains and myeloid markers are negative. Acute myelogenous leukemia (AML) presents primarily with peripheral blood and/or bone marrow involvement. Presentation in extramedullary sites, including the head and neck region, is not uncommon. Acute myelomonocytic leukemia (AML-M4) and acute monocytic leukemia (AML-M5) have had the highest incidence of associated oral infiltrates. We report a case of a 58-year-old gentleman, with no prior history of acute leukemia, presenting with a solitary palatal swelling. Initial morphologic examination favored high-grade non Hodgkin's lymphoma (NHL). Conventional cytochemical and immunohistochemical stains were negative for lymphoid and myeloid markers. Subsequent immunophenotyping via flow cytometry performed on peripheral blood and bone marrow aspirate demonstrated myeloid lineage without lymphoid differentiation, confirming the diagnosis of AML-M0.To our knowledge, this subtype of AML-M0 has not been previously reported involving the oral cavity. With absence of morphologic differentiation, and negative findings on conventional cytochemical and immunohistochemical stains, this subtype of leukemia may be misdiagnosed as non-Hodgkin's lymphoma (NHL). Flow cytometry is useful in detecting the myeloid lineage of this leukemia. PMID- 12110350 TI - Eye movements during visual mental imagery. AB - It has long been debated whether eye movements play a functional role in visual mental imagery. A recent paper by Laeng and Teodorescu presents new evidence that eye movements are stored as a spatial index that is used to arrange the component parts correctly when mental images are generated. PMID- 12110351 TI - A neural mechanism for non-verbal discourse comprehension. AB - Sometimes, a thousand words isn't worth a picture. A recent study by West and Holcomb used event-related potentials to examine non-verbal, discourse-level processing of stories presented as picture sequences. These complex pictures were presented at a rapid rate, thereby minimizing linguistic mediation. The ERP showed N300 and N400 deflections, hypothesized to reflect image activation and amodal semantic integration, respectively. PMID- 12110353 TI - Crossing the chasm of consciousness. AB - 'Toward a Science of Consciousness 2002' was held at Tucson, AZ, USA, on 8-12 April 2002 PMID- 12110352 TI - Joint explorations. AB - The 6th Joint Conference on Information Sciences was held in Research Triangle Park, North Carolina, USA, on 8-13 March 2002. PMID- 12110354 TI - The evolution of language comes of age. AB - The Fourth International Conference on the Evolution of Language was held at Harvard University, Cambridge, MA, USA, on 27-30 March 2002. PMID- 12110356 TI - Placebo effect really is all in your mind. PMID- 12110355 TI - Sniffing out a link with Alzheimer's. PMID- 12110357 TI - Grasping the point. PMID- 12110359 TI - What is the plot in intelligence? PMID- 12110360 TI - Temporal synchrony in perceptual grouping: a critique. AB - It has been hypothesized that the human visual system can use temporal synchrony for the perceptual grouping of image regions into unified objects, as proposed in some neural models. It is argued here, however, that previous psychophysical evidence for this hypothesis is due to stimulus artifacts, and that earlier studies do not, therefore, support the claims of synchrony-sensitive grouping mechanisms or processes. PMID- 12110361 TI - Visual illusions affect planning but not control. AB - Much debate has arisen over how to account for the pattern of effects of visual illusions on action - that is, the findings that illusions affect actions in some circumstances but not others. I propose that this pattern can best be explained by postulating that visual illusions affect the planning of actions but do not affect the on-line control of actions. Strong evidence for this viewpoint comes from recent studies that show 'dynamic illusion effects': a large illusion effect early in a movement, but a decreasing effect as the hand approaches the target. These findings pose difficulties for other models of illusion effects on action. PMID- 12110362 TI - Models of habituation in infancy. AB - Research on infant cognition using habituation methods has sparked considerable controversy in recent years. At the core of the debates is the issue of whether infants have early (and possibly innate) conceptual understandings. This article reviews a range of computational models of habituation that might provide insights into such discussions. The models are assessed against key behavioral and neural features of habituation: temporal unfolding, exponential decrease, familiarity-to-novelty shift, habituation to repeated testing, discriminability of habitual items, selective inhibition and cortical-subcortical interactions. The review suggests that current models fail to offer comprehensive explanations of the behavioral phenomena. PMID- 12110363 TI - Inhibitory processes and the control of memory retrieval. AB - People are often confronted with reminders of things they would prefer not to think about. When this happens, they often attempt to put the unwanted memories out of awareness. Recent research shows that the capacity to suppress distracting traces is mediated by executive-control processes that are analogous to those involved in overriding prepotent motor responses, and it is these processes that cause persisting memory failures for the suppressed items. There is evidence that memory retrieval and motor tasks that are likely to demand executive control recruit overlapping neural mechanisms, suggesting that a common process mediates control in these domains. Together, these findings indicate that memory failures often arise from the mechanisms that lie at the heart of our capacity to influence the focus of thought. PMID- 12110364 TI - Amygdala oscillations and the consolidation of emotional memories. AB - The amygdala receives multi-modal sensory inputs and projects to virtually all levels of the central nervous system. Via these widespread projections, the amygdala facilitates consolidation of emotionally arousing memories. How the amygdala promotes synaptic plasticity elsewhere in the brain remains unknown, however. Recent work indicates that amygdala neurons show theta activity during emotional arousal, and various types of oscillations during sleep. These synchronized neuronal events could promote synaptic plasticity by facilitating interactions between neocortical storage sites and temporal lobe structures involved in declarative memory. PMID- 12110365 TI - Emotion is a moral issue. PMID- 12110366 TI - Carbohydrate responsive element-binding protein (ChREBP): a key regulator of glucose metabolism and fat storage. AB - Feeding a high carbohydrate diet induces transcription of more than 15 genes involved in the metabolic conversion of glucose to fat. A new transcription factor binding to a glucose response element of the pyruvate kinase and lipogenesis enzyme genes was discovered recently. This factor, termed carbohydrate responsive element-binding protein (ChREBP), is activated in response to high glucose and up-regulates these genes. Cyclic AMP and a high fat diet inhibit ChREBP and slow down glucose utilization. ChREBP is able to control transcription of lipogenic enzyme genes in response to nutritional and hormonal inputs, and may play an important role in disease states such as diabetes, obesity, and hypertension. PMID- 12110367 TI - Identification of catalase in human livers as a factor that enhances phenytoin dihydroxy metabolite formation by human liver microsomes. AB - We have reported previously that the formation of a 3',4'-dihydroxylated metabolite of phenytoin (3',4'-diHPPH) by human liver microsomal cytochrome P450 (P450) is enhanced by the addition of human liver cytosol [Komatsu et al., Drug Metab Dispos 2000;28:1361-8]. The enhancing factor was determined in this study. The addition of cytosolic proteins precipitated by 50% ammonium sulfate to incubation mixtures increased the rate of microsomal 3',4'-diHPPH formation. This fraction was separated further by diethylaminoethyl-, carboxymethyl-, and hydroxyapatite-column chromatography. The amino acid sequence of the purified protein of approximately 55kDa by electrophoresis revealed this protein to be a catalase. The addition of purified or authentic catalase to the incubation mixtures increased the rates of microsomal 3',4'-diHPPH formation from 3'- and 4' hydroxylated metabolites and from phenytoin in a concentration-dependent manner. In reconstituted systems containing CYP2C9, CYP2C19, and CYP3A4, the formation of 3',4'-diHPPH was also enhanced by catalase to different extents. This is the first report that catalase in livers enhances drug oxidation activities catalyzed by P450 in human liver microsomes. PMID- 12110368 TI - Identification and characterization of a novel protein histidine kinase in the islet beta cell: evidence for its regulation by mastoparan, an activator of G proteins and insulin secretion. AB - Using insulin-secreting cells, we previously demonstrated that specific proteins associated with the cytosolic, secretory granule, and mitochondrial fractions undergo a novel type of phosphorylation on their histidine residues. Subsequently, we identified these proteins as the nucleoside diphosphate kinase (NDPK) [Kowluru and Metz, Biochemistry 1994;33:12495-503], the beta subunit of trimeric GTP-binding proteins [Kowluru et al., Biochem J 1996;313:97-107], and the alpha subunit of succinyl-CoA synthetase [Kowluru, Diabetologia 2001;44:89 94], respectively. Since several other enzymes of intermediary metabolism (e.g. ATP-citrate lyase and glucose-6-phosphatase) also undergo histidine phosphorylation, these initial findings may have a more generalized significance to beta cells. Herein, we characterized a novel protein histidine kinase in pancreatic beta cells, and determined it to be acid- and heat-labile as well as alkali-resistant in its phosphorylation of histone 4. Such an activity was detected in normal rat islets, human islets, and clonal beta (HIT-T15 and INS-1) cells, and could utilize either ATP or GTP as a phosphoryl donor (with K(m) values in the range of 60-100 microM). On a size-exclusion column, its molecular mass was estimated to be in the range of 60-70 kDa. It was stimulated by divalent cations (Mg(2+)>Mn(2+)>control=Ca(2+)=Zn(2+)=Co(2+)), but was resistant to polyamines. It was inactivated by known in vitro inhibitors of protein histidine phosphorylation (e.g. UDP or cromoglycate). Mastoparan, a global activator of G proteins and insulin secretion from isolated beta cells, but not mastoparan-17, its inactive analog, stimulated histidine kinase activity and histidine phosphorylation of G(beta) subunit and insulin secretion from isolated rat islets. These studies identify, for the first time, a protein kinase activity in the pancreatic beta cell that does not act on traditional -Ser, -Tyr, or -Thr residues. They also establish a possible link between histidine kinase activity and G(beta) phosphorylation in isolated beta cells. PMID- 12110369 TI - DNA sequence of butyrylcholinesterase from the rat: expression of the protein and characterization of the properties of rat butyrylcholinesterase. AB - The rat is the model animal for toxicity studies. Butyrylcholinesterase (BChE), being sensitive to inhibition by some organophosphorus and carbamate pesticides, is a biomarker of toxic exposure. The goal of this work was to characterize the purified rat BChE enzyme. The cDNA sequence showed eight amino acid differences between the active site gorge of rat and human BChE, six clustered around the acyl binding pocket and two below the active site serine. A prominent difference in rat was the substitution of arginine for leucine at position 286 in the acyl pocket. Wild-type rat BChE, the mutant R286L, wild-type human BChE, and the mutant L286R were expressed in CHO cells and purified. Arg286 was found responsible for the resistance of rat BChE to inhibition by Triton X-100. Replacement of Arg286 with leucine caused the affinity for Triton X-100 to increase 20-fold, making it as sensitive as human BChE to inhibition by Triton X 100. Wild-type rat BChE had an 8- to 9-fold higher K(m) for the positively charged substrates butyrylthiocholine, acetylthiocholine, propionylthiocholine, benzoylcholine, and cocaine compared with wild-type human BChE. Wild-type rat BChE catalyzed turnover 2- to 7-fold more rapidly than human BChE, showing the highest turnover with propionylthiocholine (201,000 min(-1)). Human BChE does not reactivate spontaneously after inhibition by echothiophate, but rat BChE reactivates with a half-life of 4.3hr. Human serum contains 5mg/L of BChE and 0.01mg/L of AChE. Male rat serum contains 0.2mg/L of BChE and approximately 0.2mg/L of AChE. PMID- 12110370 TI - Reduced (+/-)-3,4-methylenedioxymethamphetamine ("Ecstasy") metabolism with cytochrome P450 2D6 inhibitors and pharmacogenetic variants in vitro. AB - "Ecstasy" [(+/-)-3,4-methylenedioxymethamphetamine or MDMA] is a CNS stimulant, whose use is increasing despite evidence of long-term neurotoxicity. In vitro, the majority of MDMA is demethylenated to (+/-)-3,4-dihydroxymethamphetamine (DHMA) by the polymorphic cytochrome P450 2D6 (CYP2D6). We investigated the demethylenation of MDMA and dextromethorphan (DEX), as a comparison drug, in reconstituted microsomes expressing the variant CYP2D6 alleles (*)2, (*)10, and (*)17, all of which have been linked to decreased enzyme activity. With MDMA, intrinsic clearances (V(max)/K(m)) in CYP2D6.2, CYP2D6.17, and CYP2D6.10 were reduced 15-, 13-, and 135-fold, respectively, compared with wild-type CYP2D6.1. With DEX, intrinsic clearances were reduced by 37-, 51-, and 164-fold, respectively. It was evident that CYP2D6.17 displayed substrate-specific changes in drug affinity (K(m)). Compounds potentially used with MDMA [fluoxetine, paroxetine, (-)-cocaine] demonstrated significant inhibition of MDMA metabolism in both human liver and CYP2D6.1-expressing microsomes. These data demonstrate that individuals possessing the CYP2D6(*)2, (*)17, and, particularly, (*)10 alleles may show significantly reduced MDMA metabolism. These individuals, and those taking CYP2D6 inhibitors, may demonstrate altered acute and/or long-term MDMA-related toxicity. PMID- 12110371 TI - A critical role for a tyrosine residue in the cannabinoid receptors for ligand recognition. AB - Previous mutation and modeling studies have identified an aromatic cluster in the transmembrane helix (TMH) 3-4-5 region as important for ligand binding at the CB(1) and CB(2) cannabinoid receptors. Through novel mixed mode Monte Carlo/Stochastic Dynamics (MC/SD) calculations, we tested the importance of aromaticity at position 5.39(275) in CB(1). MC/SD calculations were performed on wild-type (WT) CB(1) and two mutants, Y5.39(275)F and Y5.39(275)I. Results indicated that while the CB(1) Y5.39(275)F mutant is very similar to WT, the Y5.39(275)I mutant shows pronounced topology changes in the TMH 3-4-5 region. Site-directed mutagenesis studies of tyrosine 5.39 to phenylalanine (Y-->F) or isoleucine (Y-->I) in both CB(1) and CB(2) were performed to determine the functional role of this amino acid in each receptor subtype. HEK 293 cells transfected with mutant receptor cDNAs were evaluated in radioligand binding and cyclic AMP assays. The CB(1) mutant and WT receptors were also co-expressed with G-protein-coupled inwardly rectifying channels (GIRK1 and GIRK4) in Xenopus oocytes to assess functional coupling. The Y-->F mutation resulted in cannnabinoid receptors with subtle differences in WT binding and signal transduction. In contrast, the Y-->I mutations produced receptors that could not produce signal transduction or bind to multiple cannabinoid compounds. However, immunofluorescence data indicate that the Y-->I mutation was compartmentalized and expressed at a level similar to that of the WT cannabinoid receptor. These results underscore the importance of aromaticity at position CB(1) 5.39(275) and CB(2) 5.39(191) for ligand recognition in the cannabinoid receptors. PMID- 12110372 TI - Coordinate regulation of UDP-glucuronosyltransferase UGT1A6 induction by 3 methylcholanthrene and multidrug resistance protein MRP2 expression by dexamethasone in primary rat hepatocytes. AB - Concentration-dependent regulation of 3-methylcholanthrene (MC) inducibility of UDP-glucuronosyltransferase UGT1A6 by the synthetic glucocorticoid, dexamethasone (DEX) was studied. Treatment of cultured rat hepatocytes with MC, 0.1, 1, and 10 microM DEX, and MC combined with DEX, resulted in different induction patterns measured in the intact cells compared to that observed in the microsomes prepared from the same cells. DEX treatment in various concentrations caused a concentration-dependent increase in p-nitrophenol (p-NP) conjugation in intact cells (3-, 4-, and 5-fold over control, respectively), and it positively regulated MC induction (4-, 5-, and 6-fold over control, respectively). In contrast, DEX had smaller effect on microsomal p-NP conjugation (115, 200, 220% of control, respectively) and although MC induction was increased significantly by 0.1 microM DEX (520% of control), but higher concentrations of DEX (10 microM) decreased the degree of induction to 410%. Similar results obtained from in vivo experiments showed that at high DEX concentration (100mg/kg), the rate of MC induction (540%) decreased (420%). Permeabilization of the plasma membrane resulted in a 15-fold increase of p-NP conjugation indicating the importance of transport in the rate of overall p-NP elimination, and the induction pattern was similar to that observed in microsomes isolated from cells. Hyper-osmolarity (405 mOsmol/L) led to a 3-fold decrease of p-NP conjugation, the loss of DEX inducibility and reduction of the MRP2 protein level. Our results suggest coordinated regulation of UGT1A6 inducibility and substrate or product transport by DEX. PMID- 12110373 TI - (-)-Epicatechin effects in rat liver epithelial cells: stimulation of gap junctional communication and counteraction of its loss due to the tumor promoter 12-O-tetradecanoylphorbol-13-acetate. AB - Gap junctional intercellular communication (GJIC) is a direct signaling pathway for neighboring cells. Disturbances in GJIC are suggested to play a role in carcinogenesis and may be involved in cardiac arrhythmia. Tumor promoters like 12 O-tetradecanoylphorbol-13-acetate (TPA) are capable of inhibiting GJIC, whereas GJIC is stimulated by several micronutrients like genistein, retinoids or carotenoids. (-)-Epicatechin (4-40 microM), a major flavonoid in cocoa and green tea, exhibited stimulatory effects on GJIC in WB-F344 rat liver epithelial cells after 24-72hr of incubation; no change was observed after 90 min. However, treatment of cells for 90 min with TPA (5 or 10nM) led to complete loss of GJIC, whereas 40% loss was found with 1nM. These inhibitory effects of TPA were largely suppressed when (-)-epicatechin or genistein (40 microM) were present during the incubation. In communicating WB-F344 cells, most of the major gap junction protein connexin43 (Cx43) was located in the plasma membrane. When the cells were exposed to TPA, considerably less protein was found in the membrane. Such a delocalization of Cx43 proteins was not observed when TPA was coincubated with the flavonoids, (-)-epicatechin or genistein. It is concluded that TPA affects Cx43 trafficking between cellular compartments, and that this effect is counteracted by (-)-epicatechin or genistein. PMID- 12110374 TI - Dopamine D(2) receptor-induced COX-2-mediated production of prostaglandin E(2) in D(2)-transfected Chinese hamster ovary cells without simultaneous administration of a Ca(2+)-mobilizing agent. AB - We have earlier demonstrated that dopamine stimulates the liberation of the prostaglandin E(2) (PGE(2)) precursor, arachidonic acid, in Chinese hamster ovary cells transfected with the rat dopamine D(2) receptor (long isoform), also without concomitant administration of a Ca(2+)-releasing agent [Nilsson et al., Br J Pharmacol 1998;124:1651-8]. In the present report, we show that dopamine, under the same conditions, also induces a concentration-dependent increase in the production of PGE(2), with a maximal effect of 235% at approximately 100 microM, and with an EC(50) of 794 nM. The effect was counteracted by the D(2) antagonist eticlopride, pertussis toxin, the inhibitor of intracellular Ca(2+) release TMB 8, incubation in Ca(2+)-free experimental medium, and PKC desensitization obtained by chronic pretreatment with the phorbol ester TPA. It was also antagonized by the non-specific cyclooxygenase (COX) inhibitor, indomethacin, and by the selective COX-2 inhibitor, NS-398, but not by the specific COX-1 inhibitor, valeryl salicylate. Both the non-specific phospholipase A(2) inhibitor, quinacrine, and an inhibitor of cPLA(2) and iPLA(2), AACOF3, counteracted the effect; in contrast, a selective iPLA(2) inhibitor, BEL, and a selective sPLA(2) inhibitor, TAPC, were ineffective. No effects of dopamine were obtained in control cells mock-transfected with the p3C vector only. The results reinforce previous assumptions that dopamine may interact with eicosanoid metabolism by means of D(2) receptor activation, and implicate an involvement of cPLA(2) and COX-2 in this effect. It is suggested that measurement of dopamine induced PGE(2) production may serve as a convenient way to study D(2) receptor function in vitro. PMID- 12110375 TI - Differences in caffeine and paraxanthine metabolism between human and murine CYP1A2. AB - For the characterisation of murine models of CYP1A2 mediated metabolism in humans we compared the metabolism of caffeine and paraxanthine in human liver microsomes (LM) (two samples) and in LM from CYP1A2-null and wild-type mice. Inhibition experiments were carried out with the quinolones norfloxacin and pefloxacin and the substrate, caffeine. Additionally, in vivo pharmacokinetics of paraxanthine was determined in CYP1A2-null and wild-type mice. All LM produced the primary metabolites of caffeine and paraxanthine. In human LM, the main metabolite of caffeine was paraxanthine (K(M) 0.4 and 0.5 mmol L(-1)). In wild-type and CYP1A2 null mice LM, the main caffeine metabolite was 1,3,7-trimethylurate, but formation was not saturable. Apparent K(M) for paraxanthine formation from caffeine in wild-type and CYP1A2-null murine LM were 0.2 and 4.9 mmol L(-1), respectively. The main metabolite of paraxanthine was 1-methylxanthine in human (K(M) 0.13 and 0.2 mmol L(-1)) and in wild-type mice LM (K(M) 0.53 mmol L(-1)). In CYP1A2-null murine LM, the main paraxanthine metabolite was 7-methylxanthine. The quinolones competitively inhibited caffeine metabolism in human but not in wild-type or CYP1A2-null murine LM. No obvious differences were seen for blood pharmacokinetics and urinary metabolite excretion of paraxanthine between CYP1A2 null and wild-type mice. Thus, for paraxanthine, norfloxacin and pefloxacin interaction with CYP1A2 there were clear differences between mice and man. Our results suggest that an interspecies comparison is required for the metabolism of individual xenobiotics interacting with CYP1A2 prior to the use of mice models to predict its toxicity and/or pharmacological activity in man. PMID- 12110376 TI - Elevated uptake of low density lipoprotein by drug resistant human leukemic cell lines. AB - Overexpression of a 170kD membrane glycoprotein, P-glycoprotein (Pgp), which acts as an energy dependent efflux pump for cytotoxic drugs is believed to be one of the factors that is responsible for clinical drug resistance. Recent studies suggest that Pgp is also responsible for the intracellular transport of cholesterol from the plasma membrane to the endoplasmic reticulum. Leukemic cells from patients with acute myelogenous leukemia have an elevated uptake of low density lipoprotein (LDL) when compared with white blood cells from healthy individuals. Since elevated LDL receptor expression and multidrug resistance are both common events in leukemic cells, we investigated LDL receptor expression in sensitive and drug resistant human leukemic cell lines. We found a 2- to 10-fold higher uptake of LDL in five out of five drug resistant K562 cell lines. All three drug resistant HL60 cell lines studied also had higher uptake than the parental cells. The LDL receptor expression in vincristine resistant Pgp positive K562 cells was less sensitive to downregulation by sterols than in parental cells. There was no selective effect of the Pgp inhibitor PSC-833 or other Pgp modulators on LDL receptor activity in Pgp positive cells. Since also resistant Pgp, multidrug resistance protein 1, and breast cancer resistance protein negative cells exhibited an elevated LDL receptor activity, we conclude that overexpression of these proteins is not the mechanism behind the elevated LDL uptake in the drug resistant leukemic cell lines. The findings are of interest for the concept of using lipoproteins as carriers of cytotoxic drugs in cancer treatment. PMID- 12110377 TI - Structure-activity relationship of neurokinin A(4-10) at the human tachykinin NK(2) receptor: the effect of amino acid substitutions on receptor affinity and function. AB - A structure-activity study of the neurokinin A (NKA) fragment NKA(4-10) was performed to investigate the importance of amino acid residues for receptor efficacy, potency and affinity at the NK(2) receptor in human colon circular muscle. Fourteen analogs of NKA(4-10) were produced with substitutions at positions 4, 5, 7, 9 and/or 10 of NKA. Their potencies were determined by in vitro contractile responses and affinities by radioligand binding using [125I]NKA. Functional potency was enhanced 8-fold by single amino acid substitutions with Lys(5) and MeLeu(9) but not significantly altered by substitutions Glu(4), Arg(5), His(5) and Nle(10). The multiply-substituted analogs [MeLeu(9),Nle(10)]NKA(4-10), [Lys(5),MeLeu(9),Nle(10)]NKA(4-10) and [Lys(5),(Tyr(7)),MeLeu(9),Nle(10)]NKA(4-10) displayed 6-9-fold increase in potency. Although [Arg(5),Nle(10)]NKA(4-10) was similar in potency to NKA(4-10), it was the only analog to show significantly reduced efficacy. All analogs were able to compete fully for [125I]NKA binding. [Lys(5),MeLeu(9)]NKA(4-10), [MeLeu(9),Nle(10)]NKA(4-10), [Lys(5),Nle(10)]NKA(4-10) and analogs containing single substitutions with Glu(4), Arg(5), Lys(5) and MeLeu(9) displayed significantly higher affinity, whereas those with Nle(10) and [Glu(4),Nle(10)] substitutions showed significantly lower affinity than NKA(4-10). There was a positive correlation (r=0.63) between binding affinity and functional potency, which was markedly improved (r=0.95) by removal of three analogs: [Lys(5),MeLeu(9),Nle(10)]NKA(4-10), [Lys(5),Tyr(7),MeLeu(9),Nle(10)]NKA(4-10) and [Lys(5),Tyr(I(2))(7),MeLeu(9),Nle(10)]NKA(4-10). These exhibited similar binding affinities to that of NKA(4-10) but were more potent in functional studies, possibly indicating a different mechanism of receptor interaction. In conclusion, substitution of Ser(5) with Lys, and/or N-methylation of Leu(9), were the most effective changes to increase functional and binding potency of NKA(4-10) at the human colon NK(2) receptor. PMID- 12110378 TI - Sequence and chemistry requirements for a novel aptameric oligonucleotide inhibitor of EGF receptor tyrosine kinase activity. AB - We have previously identified a phosphorothioate oligonucleotide (PS-ODN) that inhibited epidermal growth factor receptor tyrosine kinase (TK) activity both in cell fractions and in intact A431 cells. Since ODN-based TK inhibitors may have anti-cancer applications and may also help understand the non-antisense mediated effects of PS-ODNs, we have further studied the sequence and chemistry requirements of the parent PS-ODN (sequence: 5'-GGA GGG TCG CAT CGC-3') as a sequence-dependent TK inhibitor. Sequence deletion and substitution studies revealed that the 5'-terminal GGA GGG hexamer sequence in the parent compound was essential for anti-TK activity in A431 cells. Site-specific substitution of any G with a T in this 5'-terminal motif within the parent compound caused a significant loss in anti-TK activity. The fully PS-modified hexameric motif alone exhibited equipotent activity as the parent 15-mer whereas phosphodiester (PO) or 2'-O-methyl-modified versions of this motif had significantly reduced anti-TK activity. Further, T substitutions within the two 5'-terminal G residues of the hexameric PS-ODN to produce a sequence, TTA GGG, representing the telomeric repeats in human chromosomes, also did not exhibit a significant anti-TK activity. Multiple repeats of the active hexameric motif in PS-ODNs resulted in more potent inhibitors of TK activity than the parent ODN. These results suggested that PS-ODNs, but not PO or 2'-O-methyl modified ODNs, containing the GGA GGG motif can exert potent anti-TK activity which may be desirable in some anti-tumor applications. Additionally, the presence of this previously unidentified motif in antisense PS-ODN constructs may contribute to their biological effects in vitro and in vivo and should be accounted for in the design of the PS-modified antisense ODNs. PMID- 12110379 TI - Pharmacologically distinct binding sites in rat brain for [3H]thyrotropin releasing hormone (TRH) and [3H][3-methyl-histidine(2)]TRH. AB - We have used a directed peptide library, in which the histidyl residue of thyrotropin-releasing hormone (TRH) was systematically replaced by a series of 24 natural and unnatural amino acids, to characterise TRH binding sites in rat brain cortex. This was achieved by measuring the ability of library peptides to compete with [3H][3-Me-His(2)]TRH or [3H]TRH binding to rat cortical homogenates. [3H][3 Me-His(2)]TRH was observed to bind to a single population of high-affinity, low capacity sites (K(d): 4.54+/-0.62 nM, N=5; B(max): 4.38+/-0.21 fmol/mg wet weight tissue, N=5), consistent with them being central TRH receptors. Displacement studies showed TRH to bind to these sites with an apparent K(i) of 22 nM. K(i) values for the library peptides at [3H][3-Me-His(2)]TRH-labelled sites varied from 10(-3) to 10(-9)M; the potency order was: [3-Me His(2)]>His>Thi>Leu,Phe,Asn>Gln, Arg, Thr, Ala, HomoPhe. All other replacements had K(i) values >10(-4)M. [3H]TRH was observed to label a single population of low-affinity, high-capacity sites (K(d): 7.55+/-1.23 microM, N=6; B(max): 3.40+/ 0.63 pmol/mg wet weight tissue, N=6). The affinities of the synthetic peptides for [3H]TRH-labelled sites did not correlate with their affinities for [3H][3-Me His(2)]TRH-labelled sites (r=0.33, N=18, P>0.1). They did, however, correlate significantly with previously reported binding affinities for TRH-degrading ectoenzyme (r=0.72, N=12, P<0.01). These results strongly indicate that the identity of the low-affinity, [3H]TRH-labelled site is the membrane-bound enzyme, TRH-degrading ectoenzyme, not a subpopulation of TRH receptors. They also provide the first comprehensive description of the influence of the histidyl residue in TRH on binding of TRH to brain receptors. PMID- 12110380 TI - Proceedings of the 9th International Meeting of Recent Developments in Pharmaceutical Analysis (RDPA '01). Lipari, Italy, June 3-8, 2001. PMID- 12110381 TI - Separation of delta-, gamma- and alpha-tocopherols by CEC. AB - In this study capillary electrochromatography (CEC) was used for the separation of three tocopherols (TOHs), namely delta-, gamma- and alpha-TOH and the antioxidant compound, butylated hydroxytoluene (BHT). The CEC experiments were carried out using an octadecylsilica (ODS) stationary phase packed, in our laboratory, in a fused-silica capillary (100 microm I.D., 365 microm O.D. x 33 cm of total length and 24.6 or 8.4 cm effective length). The mobile phase was composed by a mixture of methanol (MeOH) and acetonitrile (ACN), at different concentrations and 0.01% (w/v) of ammonium acetate. Retention time (t(R)), retention factor (k), resolution (R(s)) of the three TOHs were strongly influenced by the organic solvent composition of the run buffer and by the effective length of the capillary. Optimum experimental conditions were found even employing the short effective length of the capillary achieving the baseline separation of the studied analytes in a relatively short time (less than 5 min). The optimized method was applied to the qualitative analysis of vitamin E (alpha TOH) present in a human serum extract. PMID- 12110382 TI - Determination of losartan and hydrochlorothiazide in tablets by CE and CEC. AB - Capillary Electrophoresis (CE) and Capillary Electrochromatography (CEC) have been used to determine losartan and hydrochlorothiazide. The CE separation was carried out in an uncoated capillary filled with a 100 mM sodium borate pH 9 solution containing trimethyl-beta-cyclodextrins. CEC was performed using a capillary packed with a RP-18 stationary phase. The mobile phase was a mixture of 50 mM ammonium acetate pH 7, water, acetonitrile (1/1.5/7.5). By CE and CEC suitable methods to determine simultaneously losartan and hydrochlorothiazide in working standard mixture or pharmaceutical form were obtained. The proposed methods are very simple and both gave accurate and precise results. PMID- 12110383 TI - An experimental design methodology applied to the enantioseparation of a non steroidal anti-inflammatory drug candidate. AB - An experimental design methodology has been applied to the enantioseparation of a new synthesized aryl propionic acid of pharmaceutical interest, namely 2-[(4' benzoyloxy-2'-hydroxy)phenyl-propionic acid] (DF-1770y) by chiral capillary zone electrophoresis (CCZE). The chiral separation of the studied compound has been achieved employing vancomycin as the chiral selector. The partial filling-counter current method has been used in order to avoid the presence of the absorbing chiral selector in the path length of the detector and to increase the method sensitivity. A central composite design has been employed to optimize the experimental conditions for a fast separation of the enantiomers of the new synthesized aryl propionic acid. Critical parameters such as chiral selector concentration, pH and temperature have been studied to evaluate how they affected responses such as resolution and migration times. The desirability function approach has been employed in order to find the best compromise between the different experimental responses. The proposed CCZE method provided the baseline enantioseparation of the investigated drug. A Britton-Robinson buffer at pH 6.4 supplemented with 7 mM of vancomycin at 22 degrees C and -20 kV were the optimum experimental conditions allowing to achieve the highest enantioresolution of DF 1770y in less than 8.5 min. PMID- 12110384 TI - Chiral separation of methoxamine and lobeline in capillary zone electrophoresis using ethylbenzene-deactivated fused-silica capillary columns and cyclodextrins as buffer additives. AB - The complete chiral separation of methoxamine and lobeline was achieved by capillary zone electrophoresis on an ethylbenzene-deactivated fused-silica capillary column and with cyclodextrins (CDs) as buffer additives. Among the CDs investigated in this study, i.e. alpha-CD, beta-CD, dimethyl-beta-CD, hydroxypropyl-beta-CD and gamma-CD, all the three beta-type CDs showed chiral recognition on the two drugs investigated. Under the investigated conditions, the baseline chiral separation of methoxamine can be achieved with 90 mM Tris-H3PO4 (pH 2.5) containing 11.5 mM of the three beta-type CDs, with dimethyl-beta-CD giving the best resolution, whereas the baseline chiral separation of lobeline can be realized by using 90 mM Tris-H3PO4 buffer (pH 2.5) containing 5.8 mM dimethyl-beta-CD or 29.5 mM hydroxypropyl-beta-CD. PMID- 12110385 TI - Study of flavonoids/beta-cyclodextrins inclusion complexes by NMR, FT-IR, DSC, X ray investigation. AB - Flavonoids are natural substances with a lot of biological activities, including the antioxidant one. Their use in pharmaceutical field is, however, limited by their aqueous insolubility. As the formation of the inclusion complexes can improve their solubility in water, the flavonoids hesperetin, hesperidin, naringenin and naringin have been complexed with beta-cyclodextrin (beta-CD) by the coprecipitation method and studied in solution and in solid state by NMR, FT IR, differential scanning calorimetry and X-ray techniques. The effects of complexation on the chemical shifts of the internal and external protons of beta CD in the presence of each flavonoid were observed. PMID- 12110386 TI - Characterization of physicochemical properties of naproxen systems with amorphous beta-cyclodextrin-epichlorohydrin polymers. AB - Ground mixtures of naproxen with amorphous beta-cyclodextrin-epichlorohydrin soluble (betaCd-EPS) or insoluble cross-linked (betaCd-EPI) polymers were investigated for both solid phase characterization (Differential Scanning Calorimetry, powder X-ray Diffractometry) and dissolution properties (dispersed amount method). The effect of different grinding conditions and of drug-to carrier ratio was also evaluated. Co-grinding induced a decrease in drug crystallinity to an extent which depended on the grinding time, and was most pronounced for the cross-linked insoluble polymer, particularly in combinations at the lowest drug content. Both cyclodextrin polymers were more effective in improving the naproxen dissolution properties, not only than the parent betaCd but also than hydroxyalkyl-derivatives, and their performance was almost comparable to that of methyl-derivatives, previously found as the best carriers for naproxen. Dissolution efficiencies of naproxen from physical mixtures with betaCd-EPS, thanks to the high water solubility of this Cd-derivative, were up to three times higher than those from the corresponding products with betaCd-EPI. However this difference in their performance became much less evident in co ground products and tended to progressively diminish with increasing the polymer content in the mixture, according to the better amorphizing power shown by betaCd EPI during the co-grinding process. The 10/90 (w/w) drug-carrier co-ground products exhibited the best dissolution properties, giving dissolution efficiencies about 30 times higher than that of naproxen alone. PMID- 12110387 TI - Temperature-rate profiles by polarimetric variable-temperature kinetic experiments to study racemization reactions. AB - The racemization of (-)-adrenaline was followed by polarimetric variable temperature kinetic experiments obtaining activation parameters and k(obs)(T) profile in one tenth of the time usually spent for traditional kinetic runs. A polarimeter connected to a computer for the acquisition and processing of the analytical data was used. The kinetic profiles were processed by both an integral method and a differential method. The results are in good agreement with each other and with those obtained by constant-temperature kinetics. PMID- 12110388 TI - Kinetic analysis of thermal decomposition for penicillin sodium salts: model fitting and model-free methods. AB - A kinetic study on decomposition processes of some penicillin salts was carried out. Both isothermal and dynamic thermogravimetric curves were used. As expected by their complex structures, several steps with different energies were involved in decomposition processes. Model-fitting and -free kinetic approaches were applied to nonisothermal and isothermal data. The kinetic triplet (f(alpha),A and E(a)) related to model-fitting method that defines a single step reaction resulted to be at variance with the multi-step nature of salts-decomposition. The model-free approach represented by the isothermal and nonisothermal isoconversional methods, gave different dependencies of the activation energy on the extent of conversion. The complex nature of the multi-step process of the studied compounds was more easily revealed using a broader temperature range in nonisothermal isoconversional method. The failure in the model-fitting method did not allow calculating shelf life and half-life times. PMID- 12110390 TI - GC-MS analysis of the lipophilic principles of Echinacea purpurea and evaluation of cucumber mosaic cucumovirus infection. AB - An analytical GC-MS method based on nonpolar fused silica capillary column was developed to analyze the lipophilic constituents, mainly alkamides, from the root extracts of Echinacea purpurea (L.) Moench. In particular, the proposed method was applied to evaluate the phytochemical impacts of cucumber mosaic cucumovirus (CMV) infection on the plant's lipophilic marker phytochemicals. Methanolic (70% v/v) extracts, obtained from root materials by ultrasonic treatments, were subjected to liquid-liquid extraction with n-hexane-ethyl acetate (1:1 v/v) to recover the lipophilic, volatile to semivolatile, principles. Seventeen components, including the 11 alkamides known to E. purpurea roots, were identified in the GC-MS traces of the analyzed fractions and efficiently separated in a turnaround time of 25 min. CMV infection was found to be responsible for significant variations in the relative compositions of the major constituents, in particular germacrene D, Dodeca-2E, 4E, 8Z, 10Z(E)-tetraenoic acid isobutylamide cis/trans isomers, Undeca-2Z, 4E-diene-8, 10-diynoic acid isobutylamide and Dodeca-2E, 4Z-diene-8, 10-diynoic acid isobutylamide. PMID- 12110389 TI - A comparison between different immobilised glucoseoxidase-based electrodes. AB - Biosensors obtained by immobilising glucose oxidase 'unentrapped' and 'entrapped in liposomes', both with a classical H2O2 amperometric electrode and with screen printed electrochemical sensor, were compared. Electrode response, linearity range and the influence of some parameters as phospholipid nature, temperature and measurement techniques were investigated. Experimental results showed that, while with the unentrapped enzyme the output current is linear only up to about 4 mM glucose concentration, the linearity range increases up to about 20 mM using enzyme-loaded liposomes; however the low permeability of the lipid bilayer decreases the electrode sensitivity to very low values (200 nA/M for palmitoylolelyl phosphatidylcholine liposomes). The approach with screen-printed sensors showed a better performance and gave biosensors with higher sensitivity (about 14500 nA/mM). A mathematical model, useful to compare the behaviour of the different analytical systems and to design electrodes with the required properties, was also proposed. PMID- 12110391 TI - In vitro metabolism of a nitroderivative of acetylsalicylic acid (NCX4016) by rat liver: LC and LC-MS studies. AB - The metabolism of a nitroderivative of acetylsalicylic acid, benzoic acid, 2 (acetyloxy)-3-[(nitrooxy)methyl]phenyl ester (NCX4016), the lead compound of a new class of NO-releasing non steroidal-antiinflammatory drugs has been studied in vitro in rat liver subcellular fractions (S 9000xg, microsomes, cytosol). Samples were extracted with CH3CN (2 vol.) containing 1% H3PO4 (2 M), vortexed for 3 min and then centrifuged for 5 min at 5000 rpm. Supernatants were diluted with 0.02 M phosphoric acid and analysed by reverse-phase LC. Linearity of calibration for NCX4016 and metabolites was observed over the range 0.25-50 microg/ml with coefficients of determination greater than 0.9996. Extraction efficiency from spiked liver samples ranged from 85 to 95% for all the analytes. In the S 9000xg fraction, NCX4016 undergoes rapid metabolization, with the formation of salicylic acid (SA) and [3-(nitrooxymethyl)phenol] (HBN). HBN is then rapidly metabolised to 3-hydroxybenzylalcohol (HBA), and mainly to a new metabolic species, whose formation takes place specifically in the liver cell cytosol. LC-MS analysis (electrospray ionisation) of the cytosol extract in negative and positive-ion modes furnished deprotonated [M-H]- and protonated [M+H]+ molecular ions at m/z 412 and 414, respectively, accompanied by the typical clusters with sodium. MS/MS analysis in negative-ion mode, by selection and collision of the ion at m/z 412, gave a fragmentation pattern characterized by the ions at m/z 272 and 254, which allowed to assign the structure of 1 (glutathion-S-yl)methylene-3-hydroxy-benzene, a conjugated product between GSH and the benzyl carbon atom of HBN. In rat liver cytosol HBN is completely metabolised to this thioether adduct within 30 min incubation; the process is enzymatically mediated by GSH transferase and strictly dependent on GSH availability. The relevance of this new metabolic pathway in NCX4016 detoxification by rat liver is discussed. PMID- 12110392 TI - Determination of amphetamines in human whole blood by capillary electrophoresis with photodiode array detection. AB - A capillary electrophoresis (CE) with photodiode array detection (DAD) method for the analysis of amphetamines in human whole blood samples is described. Amphetamines were applied to CE without any derivatization procedure and detected at 200 nm for a rapid and simple analysis. The UV-spectra are show in. Amphetamines were separated within 7 min through an uncoated fused-silica capillary (50 cm x 50 microm ID) using a 100 mM phosphate buffer (pH 2.5). A simple and fast extraction method of amphetamines from human whole blood was developed using acetonitrile. Very clean extracts were obtained in one step. Whole blood drugs free samples were spiked with amphetamine standard solution of known concentration. Linear calibration plots were obtained over a large concentration range, with correlation coefficients higher than 0.998. Recoveries between 81 and 99% were obtained. Limit of detection (LOD) was from 10 to 30 ng ml(-1) for most of amphetamines, except for MDMA, for which it was 80 ng ml(-1). PMID- 12110393 TI - Simultaneous determination of zidovudine and nevirapine in human plasma by RP-LC. AB - A simple method for the simultaneous determination of zidovudine and nevirapine in human plasma by reversed-phase liquid chromatography with UV detection at 265 nm was developed. A solid-liquid extraction procedure with internal standard was applied to the samples prior to analysis. The system requires a Zorbax SB-C18 column, 250 x 4.6 mm I.D. and a mobile phase composed of potassium dihydrogen phosphate (10 mM; pH 6.5)-acetonitrile (83:17, v/v). Peak-areas are linear; correlation coefficients are better than 0.999; both inter- and intra-day accuracy and precision are lower than 15%. Extraction recoveries are higher than 90% for both zidovudine and nevirapine. The method proposed was employed to determine the levels of the two retroviral drugs in plasma from HIV infected human subjects. PMID- 12110395 TI - Validation of a LC method for the analysis of oxaliplatin in a pharmaceutical formulation using an experimental design. AB - A rapid and sensitive RP-HPLC method with UV detection for routine control of oxaliplatin in a pharmaceutical formulation (Eloxatin) was developed. Quantitation was accomplished with the internal standard method. The procedure was validated by linearity (correlation coefficient=0.999948), accuracy, robustness and intermediate precision. Experimental design was used during validation to calculate method robustness and intermediate precision. For robustness test three factors were considered: percentage v/v of acetonitrile, flow rate and temperature; an increase in the flow rate results in a decrease of the drug found concentration, while the percentage of organic modifier and temperature have no important effect on the response. For intermediate precision measure the considered variables were: analyst, equipment and days. The RSD value (2.27%, n=24) indicated a good precision of the analytical method. PMID- 12110394 TI - Validation of a RP-LC method for the simultaneous determination of isoniazid, pyrazinamide and rifampicin in a pharmaceutical formulation. AB - A simple and accurate liquid chromatographic method was developed and validated for estimation of isoniazid (ISN), pyrazinamide (PYR) and rifampicin (RIF) in combined dosage forms. Drugs were chromatographed on a reverse phase C18 column using a mobile phase gradient and monitored at the corresponding maximum of each compounds. Peaks were identified with retention time as compared with standards and confirmed with characteristic spectra using diode-array detector. Solution concentrations were measured on a weight basis to avoid the use of an internal standard. The method does not require any specific sample preparation except the use of a guard column. The method is linear (r(2)>0.999), precise (RSD%: 0.50% for ISN, 0.12% for PYR and 0.98% for RIF), accurate (overall average recovery yields: 98.55% for ISN, 98.51 for PYR and 98.56% for RIF) and selective. Due to its simplicity and accuracy the method is suitable for routine quality control analysis of antitubercolosis combination dosage form. PMID- 12110396 TI - Analysis of aliphatic amines in air samples by HPLC with electrochemical detection. AB - A method was developed for the analysis of primary aliphatic amines by high performance liquid chromatography coupled with electrochemical detector. The electrochemical oxidation of aliphatic amines derivatized with 2,5 dihydroxybenzaldehyde was investigated at porous graphite electrodes. The derivatization reactions were performed off-line, before the chromatographic separation. The compounds were separated on a reversed phase column with a methanol-acetonitrile-phosphate buffer and detected setting at an oxidation potential of +0.5 V. The influence of the mobile phase buffer concentration and pH on the detector response was also studied. The derivatization was shown to be quantitative and the response linear between 50 and 200 ng/ml. The method is sensitive, selective and could be applicable for the assay of volatile amines in the field of environmental toxicology and also for biological monitoring after occupational exposure. PMID- 12110397 TI - Determination of adrenergic agonists from extracts and herbal products of Citrus aurantium L. var. amara by LC. AB - The purpose of this study was to set up a HPLC method to separate adrenergic amines (dl-octopamine, dl-synephrine and tyramine) and to determine their content in fruits, extracts and herbal products of Citrus aurantium L. var. amara. A rapid method for the quantitative analysis of these amines is described, based on their separation by RP-HPLC technique with UV detection. The analysis were conducted on a Lichrospher RP-18 column at room temperature, using a mobile phase consisting of 0.02 M citric acid-0.02 M NaH2PO4 (7:3 v/v) and adjusted to a final pH of 3. The detection was at 220 nm. Since some of these amines are chiral compounds and their enantiomers showed different pharmacological activity, the direct separation of synephrine enantiomers was carried out with HPLC on a beta cyclodextrin stationary phase. The mobile phase consisted of methanol-NaH2PO4 25 mM pH 3.5 (20:80 v/v) and tetrabutylammonium hydrogen sulfate 10 mM in ratio of 30:70 v/v in isocratic condition and the detection was at 220 nm. The two proposed methods were applied to the analysis of fruits, extracts and herbal products of C. aurantium L. var. amara. Taking into account that some authors have reported that l-synephrine may be converted into its d-form by high temperature, this optical isomerization was monitored by the same HPLC method used for the separation of enantiomers. PMID- 12110398 TI - Development and validation of a reversed-phase liquid chromatographic method for the assay of lidocaine in aqueous humour samples. AB - A simple, fast and reliable reversed-phase liquid chromatographic method was developed for the assay of lidocaine in human aqueous humour samples. The samples were analysed without any preliminary treatment on a C8 column with UV detection at 225 nm. The mobile phase consisted of methanol/sodium dihydrogen phosphate (30 mM) containing sodium pentansulphonate (10 mM) adjusted to pH 2.5 with phosphoric acid (50:50 v/v). Validation of the method showed it to be precise, accurate and linear over the concentration range of analysis with a limit of detection of 0.2 microgml(-1). The limit of quantitation was 2.5 microgml(-1) with a relative standard deviation of 2.5%. Linear regression analysis in the range 2.5-60 microgml(-1) gave correlation coefficients higher than 0.999. No interference from three commonly co-administered drugs was observed. The method developed was applied to the analysis of lidocaine in aqueous humour samples in order to evaluate and compare the efficacy of two different forms of administration of lidocaine for topical anaesthesia in cataract surgery. PMID- 12110399 TI - Rapid methods for determination of fluoxetine in pharmaceutical formulations. AB - Two different analytical methods for the quality control of fluoxetine in commercial formulations have been developed and compared: a spectrofluorimetric method and a capillary zone electrophoretic (CZE) method. The fluorescence emission values were measured at lambda=293 nm when exciting at lambda=230 nm. The CZE method used an uncoated fused-silica capillary and pH 2.5 phosphate buffer as the background electrolyte. The extraction of fluoxetine from the capsules consisted of a simple one-step dissolution with methanol/water, filtration and dilution. Both methods gave satisfactory results in terms of precision; the best results were obtained for the electrophoretic method, with RSD% values always lower than 2.0%. The accuracy was assessed by means of recovery studies, which gave very good results, between 97.5 and 102.6%. Furthermore, both methods also have the advantage of being very rapid. PMID- 12110401 TI - FT-IR spectroscopy: a powerful tool in pharmacology. AB - In the present work we report a Fourier Transform Infrared (FT-IR) analysis performed on rat encephalon samples in the CH-OH vibrational stretching region (2400-3800) cm(-1), in order to reveal the presence of a very diffuse commercial benzodiazepine: VALIUM. The comparison between the spectral features of normal brain and the ones of samples with administrated substance has unambiguously showed that the CH stretching region seems not to suffer from any change for the pharmacological treatment, instead the OH band is strongly modified probably due to the presence of a new spectral contribution characteristic of diazepam molecule. PMID- 12110400 TI - Determination of lecithin drug specialties and diet integrators; by means of first or second derivative enzymatic-spectrophotometric analysis. AB - The possibility has been investigated of applying derivative analysis to a classical enzymatic-spectrophotometric method for lecithin determination for the purpose of developing an analytical direct method that does not require long pretreatment of the test sample even in the case of turbid samples. Several samples of drugs and food integrators containing lecithin were thus tested using both the standard and the derivative method and then comparing the results obtained. The RSD% values of measurements on real (food or drugs) samples were always <2.5. Using a first derivative spectrophotometric method, average recovery was always between 102 and 105%. PMID- 12110402 TI - Estimation of impurity profiles of drugs and related materials: part XXI. HPLC/UV/MS study of the impurity profile of ethynodiol diacetate. AB - The impurity profile of ethynodiol diacetate was investigated using the HPLC/UV/MS method. Using the slightly modified HPLC method of USP 24 two impurities, earlier isolated by preparative HPLC and investigated by NMR spectroscopy were separated and characterised. The mass spectra amended by the diode-array UV spectra supported the earlier found structures (E and Z isomers of 17alpha-ethinyl-estr-4-ene-3beta,17-diol-3-acetate-17-(3'-acetoxy-2'-butenoate). Another, hitherto not described impurity, 17alpha-ethinyl-estr-4-ene-3beta,17 diol-3-acetate-17-(3-oxo-butanoate) has also been separated and characterised by means of its mass spectrum, NMR and UV spectra. PMID- 12110403 TI - Validation of a spectrophotometric method for the determination of iron (III) impurities in dosage forms. AB - A spectrophotometric method (lambda=535 nm) for the iron (III) impurities determination in iron protein-succinylate complex syrup using thioglycolic acid in basic ambient was proposed and validated. Assay samples were treated with 0.1 N hydrochloric acid and centrifuged to remove the interfering active drug. Linear response (r=0.9999) was observed over the range of 0.005-0.2% of the iron (III) with respect to the complex nominal concentration. The accuracy could be considered very satisfactory (recovery=97-99%). The intra-day precision (RSD) of impurity amongst six independent sample preparations, was 1.4%, and there was no significant difference between intra- and inter-day studies. Intermediate precision indicated that the assay possessed high degrees of ruggedness. The limit of quantitation was 0.005% of impurity with respect to the active drug. The results obtained for iron (III) were compared statistically with those obtained with the standard addition method by means of the Student's t-test and the variance ratio F-test; no significative difference was found. PMID- 12110404 TI - Age-related changes in the collagen network and toughness of bone. AB - The hypothesis of this study is that the mechanical integrity of the collagen network in bone deteriorates with age, and such adverse changes correlate with the decreased toughness of aged bone. To test the hypothesis, 30 human cadaveric femurs from donors ranging from 19 to 89 years of age were tested to determine the age-related changes in the mechanical properties of demineralized bone and fresh bone samples. Along with bone porosity, bone density, and weight fractions of the mineral and organic phases, collagen denaturation and concentrations of collagen cross-links (HP, hydroxylysylpyridinoline; LP, lysylpyridinoline; PE, pentosidine) were determined for these bone specimens as a function age. Analysis of variance (ANOVA) showed that age-dependent changes were reflected in the decreased strength, work to fracture, and fracture toughness of bone; in the decreased strength, elastic modulus, and work to fracture of the collagen network; as well as in the increased concentration of pentosidine (a marker of nonenzymatic glycation) and increased bone porosity. Regression analyses of the measured parameters showed that the age-related decrease in work to fracture of bone (especially its postyield portion) correlated significantly with deterioration in the mechanical integrity of the collagen network. The results of this study indicate that the adverse changes in the collagen network occur as people age and such changes may lead to the decreased toughness of bone. Also, the results suggest that nonenzymatic glycation may be an important contributing factor causing changes in collagen and, consequently, leading to the age-related deterioration of bone quality. PMID- 12110405 TI - The contribution of the organic matrix to bone's material properties. AB - Bone is a two-phase porous composite material comprised primarily of collagen and mineral, which together provide its mechanical properties. The contribution of the mineral phase to bone's mechanical properties has dominated scientific thinking. Collagen's role has been underappreciated and not very well studied. However, there is evidence that changes in collagen content, or changes to inter- and intrafibrillar collagen cross-linking, can reduce the energy required to cause bone failure (toughness), and increase fracture risk. Although collagen may have less effect on bone's strength and stiffness than does mineral, it may have a profound effect on bone fragility. Collagen changes that occur with age and reduce bone's toughness may be an important factor in the risk of fracture in older women with low bone mass. PMID- 12110406 TI - Osteogenesis imperfecta type VII: an autosomal recessive form of brittle bone disease. AB - Osteogenesis imperfecta (OI) is a heritable disease of bone with low bone mass and bone fragility. The disease is generally classified into four types based on clinical features and disease severity, although recently fifth and sixth forms have also been reported. Most forms of OI are autosomal dominant. Rarely, autosomal recessive disease has been described. We report the clinical, radiological, and histological features of four children (age 3.9-8.6 years at last follow-up; all girls) and four adults (age 28-33 years; two women) with a novel form of autosomal recessive OI living in an isolated First Nations community in northern Quebec. In keeping with the established numeric classification for OI forms, we have called this form of the disease OI type VII. The phenotype is moderate to severe, characterized by fractures at birth, bluish sclerae, early deformity of the lower extremities, coxa vara, and osteopenia. Rhizomelia is a prominent clinical feature. Histomorphometric analyses of iliac crest bone samples revealed findings similar to OI type I, with decreased cortical width and trabecular number, increased bone turnover, and preservation of the birefringent pattern of lamellar bone. The disease has subsequently been localized to chromosome 3p22-24.1, which is outside the loci for type I collagen genes. The underlying genetic basis for the disease remains to be determined. PMID- 12110407 TI - Osteogenesis imperfecta type VII maps to the short arm of chromosome 3. AB - We have identified a novel form of autosomal recessive osteogenesis imperfecta (OI) in a small First Nations community from northern Quebec. Mutation screening of the COL1A1/COL1A2 genes revealed no detectable mutations, and type I collagen protein analyses were also normal. By linkage analysis, we mapped this unique autosomal recessive variant of osteogenesis imperfecta to chromosome 3p22-24.1. Based on the assumption of a founder effect, genome-wide screening was performed on a DNA sample pooled from seven affected individuals. Familial as well as historical recombinations identified within an extended haplotype of 19 markers localized the disease between markers D3S2324 and D3S1561, separated by <5 cM. Based on chromosomal localization to 3p22-24.1, the transforming growth factor beta receptor 2 gene and the parathyroid hormone/parathyroid hormone-related peptide receptor were tested, but were excluded as being associated with the phenotype. This study excludes type I collagen mutations in the pathogenesis of the disease and assigns this form of OI to a locus other than the ones containing the type I collagen genes. PMID- 12110408 TI - Intervention thresholds for osteoporosis. AB - The aim of this study was to determine the threshold of fracture probability at which interventions become cost-effective. We modeled the effects of a treatment costing $500/year, given for 5 years, that decreased the risk of all osteoporotic fractures by 35%, followed by a waning of effect for 5 years. Sensitivity analyses included a range of effectiveness (10%-50%) and a range of intervention costs (200-500 dollars/year). Data on costs and risks were from Sweden. Costs included direct costs and costs in added years of life, but excluded indirect costs due to morbidity. A threshold for cost-effectiveness of 60,000 dollars per quality-adjusted life-year (QALY) gained was used. Costs of added years were excluded in a sensitivity analysis for which a threshold value of 30,000 dollars per QALY was used. In the base case, intervention was cost-effective when treatment was targeted to women at average risk at age of >or=65 years. Irrespective of the efficacy modeled (10%-50%) or of cost of intervention (200 500 dollars/year) segments of the population at average risk could be targeted cost-effectively: The lower the intervention cost and the higher the effectiveness, the lower the age at which intervention was cost-effective. With the base case (500 dollars/year; 35% efficacy) treatment in women was cost effective with a 10 year hip fracture probability that ranged from 1.4% at the age of 50 years to 4.4% at the age of 65 years. The exclusion of osteoporotic fractures other than hip fracture would increase the threshold to a 9%-11% 10 year probability because of the substantial morbidity from fractures other than hip fracture, particularly at younger ages. We conclude that the inclusion of all osteoporotic fractures has a marked effect on intervention thresholds, that these vary with age, and that available treatments can be cost-effectively targeted to individuals at moderately increased risk. PMID- 12110409 TI - The impact of osteoporosis on quality-of-life: the OFELY cohort. AB - Although the long-term outcomes of osteoporosis (Op) such as fracture, kyphosis, and pain are well known, the physical, psychological, and social consequences, beyond fracture and pain, are less clear. The Osteoporosis-targeted Quality-of life (OPTQoL) questionnaire aimed at assessing the physical difficulty, fears, and adaptations to one's daily life was developed as a cross-sectional instrument to characterize the burden of Op within a community. The purpose of this study was to assess the impact of Op and related factors on community women participating in the OFELY study in France. Femoral neck bone mineral density (BMD) and OPTQoL questionnaire data were collected from women randomly selected from a large insurance company. Data were obtained for 756 women (mean age 59 years, range 36-92), most of whom were white. Women were classified into five groups based on the extent of physical manifestations and family history of Op. Women who had prior fractures, height loss, and/or kyphosis or both reported greater physical difficulty, more adaptations to their lives, and greater fears than women reporting no such changes. Scores on the Physical Difficulty domain, however, did not differ significantly based on BMD alone (BMD T score 20 years after menopause (n = 102; 204 alleles, p = 0.024). The type 7 allele was associated with high bone density in women more than 20 years after menopause (p = 0.042). The association study with spondylosis of postmenopausal women (n = 221) revealed that another distinct allele, type 8, was significantly associated with low spondylosis score at L-4/5 (p = 0.019) and L-5/S-1 (p = 0.048) levels in the subpopulation equal to or younger than the average age ( 0.65 and p < 0.01) as well as Fe and star marrow space (eta(2) > 0.45 and p < 0.05). The cylinders from subjects with high pyrrole or low HP in their bone collagen had a relatively thick and simple structure. Those with low pyrrole and high HP had relatively thin trabeculae that were more numerous and spread over a complex network. The relative concentrations of the pyrrole and pyridinoline cross-links appear to reflect the structural organization of the trabeculae. PMID- 12110416 TI - Mechanical strength of the thoracolumbar spine in the elderly: prediction from in situ dual-energy X-ray absorptiometry, quantitative computed tomography (QCT), upper and lower limb peripheral QCT, and quantitative ultrasound. AB - The objective of this study was to compare the ability of clinically available densitometric measurement techniques for evaluating vertebral strength in elderly individuals. Measurements were related to experimentally determined failure strength in the thoracic and lumbar spine. In 127 specimens (82 women and 45 men, age 80 +/- 10 years), dual-energy X-ray absorptiometry (DXA) was performed at the lumbar spine, femur, radius, and total body, and peripheral-quantitative computed tomography (pQCT) at the distal radius, tibia, and femur under in situ conditions with intact soft tissues. Spinal QCT and calcaneal ultrasound parameters were performed ex situ in degassed specimens. Mechanical failure loads of thoracic vertebrae 6 and 10 (T-6 and -10), and lumbar vertebra 3 (L-3) were determined in axial compression on functional three-segment units. In situ anteroposterior DXA and QCT of the lumbar spine explained approximately 65% of the variability of thoracolumbar failure. A combination of cortical and trabecular density (QCT) provided the best prediction in the lumbar spine. However, this was not the case in the thoracic spine, for which lumbar cortical density (QCT) and DXA provided significantly better estimates than trabecular density (QCT). pQCT was significantly less correlated with the strength of lumbar and thoracic vertebrae (r(2) = 40%), but was equivalent to femoral or radial DXA. pQCT measurements in the lower limb showed no advantage over those at the distal radius. Ultrasound explained approximately 25% of the variability of vertebral failure strength and added independent information to spinal QCT, but not to spinal DXA. These experimental results advocate site-specific assessment of vertebral strength by either spinal DXA or QCT. PMID- 12110417 TI - The bone formation defect in idiopathic juvenile osteoporosis is surface specific. AB - We have previously shown that idiopathic juvenile osteoporosis (IJO) is characterized by a decreased cancellous bone volume and a very low bone formation rate on cancellous surfaces. Whether IJO similarly affects cortical bone is unknown. We therefore compared tetracycline double-labeled transfixing iliac crest bone biopsies from eight children with typical clinical features of IJO (six girls; age 10-12 years) and from nine children (four girls; age 9-12 years) without metabolic bone disease. No differences in intracortical remodeling activity were detected. Both structural parameters reflecting intracortical remodeling (cortical porosity, active canal diameter, and quiescent canal diameter) and bone surface-based metabolic parameters (osteoid, osteoblast, mineralizing, osteoclast and eroded surfaces, and bone formation rate) were similar in IJO patients and controls (p > 0.2 each, t-test). Although the internal cortex of the biopsy was thinner in IJO patients than in controls (660 +/- 170 microm vs. 980 +/- 320 microm; p = 0.02), there was no difference in the width of the external cortex (p = 0.36). In growing children, both cortices exhibit an external modeling drift. Therefore, the difference in internal cortical width point to a decreased modeling activity on the endocortical surface of the internal cortex. In fact, bone formation rate on this surface was 48% lower in IJO patients than in controls (82 +/- 45 microm(3)/microm(2) per year vs. 159 +/- 162 microm(3)/microm(2) per year). However, this difference did not achieve statistical significance (p = 0.21) due to the high variability of bone formation rate on modeling surfaces. The disturbance of bone remodeling in IJO is limited to cancellous bone, but there may be a modeling defect affecting the internal cortex. Thus, the process causing IJO appears to mainly affect bone surfaces that are in contact with the bone marrow cavity. PMID- 12110418 TI - Correlation of bone mineral density with strength and microstructural parameters of cortical bone in vitro. AB - The aim of this study was to evaluate the influence of microstructural parameters, such as porosity and osteon dimensions, on strength. Therefore, the predictive value of bone mineral density (BMD) measured by quantitative computed tomography (QCT) for intracortical porosity and other microstructural parameters, as well as for strength of cortical bone biopsies, was investigated. Femoral cortical bone specimens from the middiaphysis of 23 patients were harvested during total hip replacement while drilling a hole (dia. 4.5 mm) for the relief of the intramedullary pressure. In vitro structural parameters assessed in histological sections as well as BMD determined by quantitative computed tomography were correlated with yield stress, and elastic modulus assessed by a compression test of the same specimens. Significant correlations were found between BMD and all mechanical parameters (elastic modulus: r = 0.69, p < 0.005; yield stress: r = 0.64, p < 0.005). Significant correlations between most structural parameters assessed by histology and yield stress were discovered. Structural parameters related to pore dimensions revealed higher correlation coefficients with yield stress (r = -0.69 for average pore diameter and r = -0.62 for fraction of porous structures, p < 0.005) than parameters related to osteons (r = 0.60 for osteon density and average osteonal area, p < 0.005), whereas elastic modulus was predicted equally well by both types of parameters. Significant correlations were found between BMD and parameters related to porous structures (r = 0.85 for porosity, 0.80 for average pore area, and r = 0.79 for average pore diameter in polynomial regression, p < 0.005). Histologically assessed porosity correlated significantly with parameters describing porous structures and haversian canal dimensions. Our results indicate a relevance of osteon density and fraction of osteonal structures for the mechanical parameters of cortical bone. We consider the measurement of BMD by quantitative computed tomography to be helpful for the estimation of bone strength as well as for the prediction of intracortical porosity and parameters related to porous structures of cortical bone. PMID- 12110419 TI - Intravenous pamidronate in the treatment of transient osteoporosis of the hip. AB - The aim of this study was to evaluate the efficacy of intravenous pamidronate in patients with transient osteoporosis of the hip (TOH). Thirteen men and three women (mean age 38.3 years, range 30-49) were recruited. The diagnosis was made by means of radiographs, bone scintigraphy, and magnetic resonance imaging (MRI). Pamidronate (45 mg) was intravenously administered three times, once every third day. The outcome measures included a clinical assessment using a pain visual analog scale (VAS; range 0-100), and the WOMAC functional impairment score (FUI; range 0-100). The bone mineral density (BMD) of the total hip and femoral neck was measured using dual-energy X-ray absorptiometry (DXA). Clinical assessments were made before treatment (T(0)) and 1 month later (T(1)), and the densitometric measurements at T(0), and then after 2 (T(2)) and 4 months (T(4)). A further MRI scan was made 3 months after treatment. In comparison to the unaffected side, there was a significant decrease at T(0) in the BMD of both the total hip (median 16.6%, range 8.5%-29.1%, p < 0.00001) and femoral neck (median 22.5%, range 12.0% 34.2%, p < 0.00001). By T(1), both VAS and FUI had decreased significantly (p < 0.00001). By T(2), the total hip and femoral neck BMD had increased by 10.9% (range 2.7%-23.6%, p < 0.00001) and 12.3% (range 7.8%-26.9%, p < 0.00001), respectively, and all patients were asymptomatic. By T(3), the MRI findings had normalized in all patients and, at T(4), there was a further increase in BMD. None of the patients experienced symptom relapse during the follow-up of 39.5 +/- 17.7 months. These results suggest that a short course of pamidronate is effective in treating TOH, and leads to a prompt and long-lasting recovery. PMID- 12110420 TI - Reduced bone mineral density in postmenopausal women self-reporting premenopausal wrist fractures. AB - Postmenopausal fractures are associated with low bone mass; however, the role of low peak bone mass in young adults in determining subsequent osteoporosis suggests that premenopausal fractures may also be relevant. We therefore sought to determine whether a self-reported previous history of premenopausal wrist and nonwrist fractures could also be associated with bone density and therefore be used to predict osteoporosis. We recruited 453 volunteer women with a median age of 64 years (range 50-83 years), with no metabolic bone disease, previous femoral neck fracture, or prevalent vertebral fracture. Bone density at the femoral neck (FN) and lumbar spine (LS) was measured using a Lunar DPX-L. As expected, the 319 women who did not report any fracture had a higher T score at LS (-0.93 +/- 1.44) than the 134 women who reported a previous fracture at any site and at any age (T score -1.60 +/- 1.21, p < 0.001). The findings for the FN were similar. Compared with fracture-free women, the women who reported a first wrist fracture before menopause now had a lower LS T score (-1.77 +/- 1.20, n = 15, p < 0.05), whereas those who reported a nonwrist fracture showed no significant decrease in their LS T score (-1.26 +/- 1.00, n = 36). When both wrist and nonwrist fractures had occurred after menopause, the T score was significantly lower. Twenty percent of the fracture-free women were osteoporosis patients. After adjusting for body weight, age, hormonal replacement therapy (HRT), and hip fracture in the family, the relative risk (RR) of osteoporosis for premenopausal wrist fractures was 2.7 (95% confidence interval 1.4-4.3) vs. 1.2 (0.7-2.4) for women with premenopausal nonwrist fractures. We conclude that self-reported premenopausal wrist fractures, but no other fractures occurring before menopause, are likely to be associated with osteoporosis at 65 years of age, and therefore constitute strong grounds for screening. PMID- 12110421 TI - The influence of microcomputed tomography threshold variations on the assessment of structural and mechanical trabecular bone properties. AB - In this study, we investigate how morphological parameters and mechanical properties derived from microcomputed tomography (microCT) are affected by small errors in threshold value when variable bone structures and different bone volume fractions are involved. For this purpose, biopsies of vertebrae of 6-, 23-, and 230-week-old female pigs were scanned using microCT. For each specimen, five threshold values were determined within the range of thresholds that an observer could select realistically, in steps of 0.5%. The scans were converted to microfinite-element (microFE) models, used to determine the elastic moduli. A variation of 0.5% in threshold resulted in a 5% difference in bone volume fraction and 9% difference in maximal stiffness for bone cubes with a volume fraction of <0.15. When the volume fraction was >0.2, these differences were only 2% and 3%, respectively. For all bone cubes, the differences for trabecular thickness and bone surface density were <3%. The effects on morphological anisotropy and trabecular number were negligible for threshold variations of 0.5%. These findings suggest that threshold selection is important for the accurate determination of volume fraction and mechanical properties, especially for low bone volume fractions; the architectural directionality is less sensitive to changes in threshold. PMID- 12110422 TI - Gender-specific pubertal changes in volumetric cortical bone mineral density at the proximal radius. AB - It is well established that puberty affects the geometry of cortical bone differently in females and males. In the present study we investigated whether there are also gender differences in the volumetric bone mineral density of the cortical compartment (BMDcort). BMDcort was determined at the proximal radial diaphysis in 362 healthy children and adolescents (age 6-23 years; 185 females, 177 males) and in 107 adults (age 29-40 years; 88 women, 19 men) using peripheral quantitative computed tomography (pQCT). The densitometric result for BMDcort was similar in prepubertal girls and boys, but was significantly higher in females after pubertal stage 3. pQCT results for BMDcort are influenced by cortical thickness due to the partial volume effect. Therefore, these gender differences were reanalyzed in groups of subjects of the same developmental stage who were matched for cortical thickness. Thus calculated, no gender difference in BMDcort was detected in prepubertal children. However, adolescent females after pubertal stage 3 and adult women had a 3%-4% higher BMDcort than males at the same developmental stage. BMDcort is an integrated measure of both cortical porosity and mean material density of cortical bone. The metabolic activity of cortical bone (intracortical remodeling) increases cortical porosity and decreases the mean material density of cortical bone. Our results therefore suggest that intracortical remodeling is lower in postpubertal females than in males. PMID- 12110423 TI - Amelioration of osteoporosis by menatetrenone in elderly female Parkinson's disease patients with vitamin D deficiency. AB - Significant reduction in bone mineral density (BMD) occurs in patients with Parkinson's disease (PD), correlating with immobilization and with vitamin D deficiency, and increasing the risk of hip fracture, especially in elderly women. As a biological indicator of compromised vitamin K status, an increased serum concentration of undercarboxylated osteocalcin (Oc) has been associated with reduced BMD in the hip and an increased risk of fracture in otherwise healthy elderly women. We evaluated treatment with vitamin K(2) (menatetrenone; MK-4) in maintaining BMD and reducing the incidence of nonvertebral fractures in elderly female patients with PD. In a random and prospective study of PD patients, 60 received 45 mg of MK-4 daily for 12 months, and the remaining 60 (untreated group) did not. At baseline, patients of both groups showed vitamin D and K(1) deficiencies, high serum levels of ionized calcium, and glutaminic residue (Glu) Oc, and low levels of parathyroid hormone (PTH) and 1,25-dihydroxyvitamin D [1,25 (OH)(2)D], indicating that immobilization-induced hypercalcemia inhibits renal synthesis of 1,25-(OH)(2)D and compensatory PTH secretion. BMD in the second metacarpals increased by 0.9% in the treated group and decreased by 4.3% in the untreated group (p < 0.0001). Vitamin K(2) level increased by 259.8% in the treated group. Correspondingly, significant decreases in Glu Oc and calcium were observed in the treated group, in association with an increase in both PTH and 1,25-(OH)(2)D. Ten patients sustained fractures (eight at the hip and two at other sites) in the untreated group, and one hip fracture occurred among treated patients (p = 0.0082; odds ratio = 11.5). The treatment with MK-4 can increase the BMD of vitamin D- and K-deficient bone by increasing vitamin K concentration, and it can also decrease calcium levels through inhibition of bone resorption, resulting in an increase in 1,25-(OH)(2)D concentration. PMID- 12110424 TI - Secondary osteoporosis and the risk of distal forearm fractures in men and women. AB - Secondary osteoporosis plays an important role in the pathogenesis of hip and spine fractures, but relatively little is known about the potential impact of secondary osteoporosis and fall-related disorders on the risk of distal forearm fractures. To address this issue, we conducted a population-based, nested case control study comparing 496 Rochester, Minnesota, residents with an initial distal forearm fracture to an equal number of age- and gender-matched controls. Potential risk factors were assessed by review of each subject's complete (inpatient and outpatient) medical records in the community (median duration >30 years) and analyzed using multiple logistic regression. Although history of diabetes mellitus in women (odds ratio [OR] 0.34, 95% confidence interval [CI] 0.15-0.75) and long-term anticonvulsant use in both genders (OR 3.58, 95% CI 1.26 10) were independently associated with fracture risk in a multivariate analysis, the conditions linked with secondary osteoporosis had, in aggregate, no statistically significant association with distal forearm fractures. Fall-related conditions altogether were associated with a borderline increase in risk (OR 1.36, 95% CI 0.98-1.91) and might have accounted for 19% of forearm fracture occurrence in the community. Among women (OR 2.72, 95% CI 1.20-6.19), but not men, a history of prior osteoporotic fracture was also associated with an increase in distal forearm fractures. These factors do not appear to account for the discrepancy in forearm fracture incidence in women when compared with men. PMID- 12110426 TI - Age-related differences between thinning of horizontal and vertical trabeculae in human lumbar bone as assessed by a new computerized method. AB - To investigate whether vertical trabeculae undergo compensatory thickening with age in the human vertebral body, a new computerized method was developed that is able to distinguish between horizontal and vertical trabeculae on normal histological sections. Study subjects included 48 individuals (24 women aged 19 97 years, and 24 men aged 23-95 years). From each L-2, thick frontal sections of half of the vertebra were embedded undecalcified in methylmetacrylate and cut into 10-microm-thick sections. A simple method able to classify image pixels as belonging to either vertical or horizontal trabeculae was developed and implemented in a computer program. The parallel-plate model was modified so that it was able to determine trabecular thickness, number, and separation (Tb.Th, Tb.N, and Tb.Sp) for horizontal and vertical trabeculae separately. The histomorphometric parameters were measured in three different regions: whole section, mid-third, and sub-endplate, for both horizontal and vertical trabeculae. It was found that the trabecular thickness of vertical trabeculae was independent of age in all investigated regions. The thickness of the horizontal trabeculae, in contrast, decreased significantly with age in all these regions. Tb.N decreased significantly with age for both horizontal and vertical trabeculae in all regions. However, the relative loss of trabeculae per unit length was largest for the horizontal trabeculae, whereas the absolute loss of trabeculae per unit length was largest for the vertical trabeculae. Tb.Sp was found to increase significantly with age for both vertical and horizontal trabeculae in all regions. No significant gender-related differences were found. In conclusion, in this study we describe a new, simple method for separation of horizontal and vertical bone tissue. This method has been applied on frontal vertebral sections. Trabecular bone thickness measured with the parallel-plate model showed that the thickness of horizontal trabeculae decreases significantly with age, whereas the thickness of vertical trabeculae did not decrease significantly with age. Furthermore, although the relative loss of trabeculae was larger for horizontal trabeculae than for vertical trabeculae, the absolute loss of vertical trabeculae was higher than that of horizontal trabeculae. PMID- 12110425 TI - Change in bone mass distribution induced by hormone replacement therapy and high impact physical exercise in post-menopausal women. AB - The purpose of this intervention trial was to determine whether changes in bone mass distribution could be observed in postmenopausal women following hormone replacement therapy (HRT) and/or high-impact physical exercise. Eighty healthy women, aged 50-57 years, at <5 years after the onset of menopause and with no previous use of HRT, were randomly assigned to one of four groups: HRT; exercise (Ex); HRT + Ex (ExHRT); and control (Co). HRT administration was conducted in a double-blind manner for 1 year using estradiol plus noretisterone acetate (Kliogest). The exercise groups participated in a 1 year progressive training program consisting of jumping and bounding activities. Subjects participated in two supervised sessions per week and were asked to perform a series of exercises at home 4 days/week. Bone measurements using a quantitative computed tomography scanner (Somatom DR, Siemens) were obtained from the proximal femur, midfemur, proximal tibia, and tibial shaft. Data were analyzed with a software program (BONALYSE 1.3) calculating density (g/cm(3)), cross-sectional area (CSA; mm(2)), and moments of inertia (I(max), I(min), I(polar)). In addition, the bone mass spectrum was determined as a function of the angular distribution around the bone mass center (polar distribution) and the distance from the bone mass center through the diaphyseal wall (radial distribution). After the 1 year period, there was an overall interaction of group x time in bone mineral density (BMD) at the proximal femur (p = 0.05) and tibial shaft (p = 0.035). Women in the ExHRT and HRT groups had increased proximal femur and tibial shaft BMD when compared with the change observed in the Co group (p = 0.024-0.011). The change was more pronounced in the cortical tibia, wherein the ExHRT group also differed from the Ex group (p = 0.038). No significant changes were found in bone CSA at any of the measured sites. The radial distribution indicated an increase of BMD in the endocortical part of the measured sites in the HRT and ExHRT groups and in the proximal tibia in the Ex group. The polar distribution showed that bone mass was redistributed in the anteroposterior direction. The changes in I(max), I(min), and I(polar) in the HRT and ExHRT groups differed from those in the Co group at the proximal femur, midfemur, and proximal tibia (p = 0.047-0.001). The Ex group also differed from the Co group in I(max) and I(polar) at the proximal tibia (p = 0.018 and 0.039, respectively). These results support the idea that HRT acts primarily at the bone-marrow interface. The exercise intervention chosen for this study contributed to the maintenance of bone mass. Our results suggest that both HRT and exercise have local effects on bone mass. The change in bone mass distribution induced by HRT and exercise may play an important role in the alteration of bone strength. PMID- 12110427 TI - Changes in gene expression associated with the bone anabolic effects of basic fibroblast growth factor in aged ovariectomized rats. AB - Basic fibroblast growth factor (bFGF) stimulates bone formation in vitro and in vivo. The purpose of this study was to determine changes in gene expression for bone matrix proteins, growth factors, and cytokines associated with the stimulatory effects of bFGF on bone formation in aged ovariectomized (ovx) rats. At 3 months of age, female Sprague-Dawley rats were sham-operated (sham) or ovariectomized (ovx), then maintained untreated for 1 year. At 15 months of age, baseline (BSL) sham and ovx rats were killed. All other rats received daily intravenous injections of bFGF (200 microg/kg) or vehicle (veh) for 14 days. Lumbar vertebrae were processed for quantitative bone histomorphometry or molecular analyses. Ovariectomy decreased vertebral cancellous bone volume by approximately 33% and increased most indices of bone turnover. Treatment of aged ovx rats with bFGF for only 14 days significantly increased cancellous bone volume compared with vehicle treatment of ovx rats, but this variable remained lower than in sham + veh rats. Osteoid volume, osteoblast surface, and osteoid surface were markedly increased, and osteoclast surface was significantly decreased in ovx + bFGF rats compared with sham + veh and ovx + veh rats. Northern analyses revealed that mRNA levels for osteocalcin and type I collagen, relative to 18S RNA, were significantly higher in ovx + bFGF rats than in ovx + veh rats by a factor of >10. RNase protection assays revealed that insulin-like growth factor (IGF-I) mRNA levels, relative to L32 housekeeping gene, were also significantly higher, by nearly a factor of 3, in ovx + bFGF rats than in ovx + veh rats. Treatment of ovx rats with bFGF did not appear to affect message levels for transforming growth factor-beta (TGF-beta), interleukin-6 (IL-6), and interferon-gamma (IFN-gamma). These in vivo results suggest that bFGF treatment upregulates gene expression for IGF-I, which may mediate, at least in part, the increased gene expression for bone matrix proteins and the bone anabolic effects of bFGF in aged ovx rats. PMID- 12110428 TI - Site- and compartment-specific changes in bone with hindlimb unloading in mature adult rats. AB - The purpose of this study was to examine site- and compartment-specific changes in bone induced by hindlimb unloading (HU) in the mature adult male rat (6 months old). Tibiae, femora, and humeri were removed after 14, 21, and 28 days of HU for determination of bone mineral density (BMD) and geometry by peripheral quantitative computed tomography (pQCT), mechanical properties, and bone formation rate (BFR), and compared with baseline (0 day) and aging (28 day) controls. HU resulted in 20%-21% declines in cancellous BMD at the proximal tibia and femoral neck after 28 day HU vs. 0 day controls (CON). Cortical shell BMD at these sites was greater (by 4%-6%) in both 28 day HU and 28 day CON vs. 0 day CON animals, and nearly identical to that gain seen in the weight-bearing humerus. Mechanical properties at the proximal tibia exhibited a nonsignificant decline after HU vs. those of 0 day CON rats. At the femoral neck, a 10% decrement was noted in ultimate load in 28 day HU rats vs. 28 day CON animals. Middiaphyseal tibial bone increased slightly in density and area during HU; no differences in structural and material properties between 28 day HU and 28 day CON rats were noted. BFR at the tibial midshaft was significantly lower (by 90%) after 21 day HU vs. 0 day CON; this decline was maintained throughout 28 day HU. These results suggest there are compartment-specific differences in the mature adult skeletal response to hindlimb unloading, and that the major impact over 28 days of unloading is on cancellous bone sites. Given the sharp decline in BFR for midshaft cortical bone, it appears likely that deficits in BMD, area, or mechanical properties would develop with longer duration unloading. PMID- 12110429 TI - Effect of recombinant human osteogenic protein-1 on the healing of a freshly closed diaphyseal fracture. AB - Osteogenic protein-1 (OP-1), or bone morphogenetic protein-7, is an osteoinductive morphogen that is involved in embryonic skeletogenesis and in bone repair. In bone defect models without spontaneous healing, local administration of recombinant human OP-1 (rhOP-1) induces complete healing. To investigate the ability of rhOP-1 to accelerate normal physiologic fracture healing, an experimental study was performed. In 40 adult female goats a closed tibial fracture was made, stabilized with an external fixator, and treated as follows: (1) no injection; (2) injection of 1 mg rhOP-1 dissolved in aqueous buffer; (3) injection of collagen matrix; and (4) injection of 1 mg rhOP-1 bound to collagen matrix. The test substances were injected in the fracture gap under fluoroscopic control. At 2 and 4 weeks, fracture healing was evaluated with radiographs, three dimensional computed tomography (CT), dual-energy X-ray absorptiometry, biomechanical tests, and histology. At 2 weeks, callus diameter, callus volume, and bone mineral content at the fracture site were significantly increased in both rhOP-1 groups compared with the no-injection group. As signs of accelerated callus maturation, bending and torsional stiffness were higher and bony bridging of the fracture gap was observed more often in the group with rhOP-1 dissolved in aqueous buffer than in uninjected fractures. Treatment with rhOP-1 plus collagen matrix did not result in improved biomechanical properties or bony bridging of the fracture gap at 2 weeks. At 4 weeks there were no differences between groups, except for a larger callus volume in the rhOP-1 plus collagen matrix group compared with the control groups. All fractures showed an advanced stage of healing at 4 weeks. In conclusion, the healing of a closed fracture in a goat model can be accelerated by a single local administration of rhOP-1. The use of a carrier material does not seem to be crucial in this application of rhOP-1. PMID- 12110430 TI - Improvement of fracture healing by systemic administration of growth hormone and local application of insulin-like growth factor-1 and transforming growth factor beta1. AB - Fracture healing is influenced by numerous hormones, growth factors, and cytokines. The systemic administration of growth hormone (GH) has shown to accelerate bone regeneration. Local application of growth factors, such as insulin-like growth factor-1 (IGF-1) and transforming growth factor-beta-1 (TGF beta1), are known to stimulate bone metabolism. Until now, the exact local and systemic mechanisms that lead to improved bone regeneration remain unclear. In addition, the effect of systemic administration of GH as compared with locally delivered growth factors on fracture healing in rats is not known. A midshaft fracture of the right tibia of 5-month-old female Sprague-Dawley rats (n = 80) was intramedullary stabilized with IGF-1 and TGF-beta1 coated vs. uncoated titanium K-wires. The growth factors were incorporated in a poly(D,L-lactide) (PDLLA) coating and released continuously throughout the experiment. Recombinant species-specific (rat) GH was applied systemically (2 mg/kg body weight) by daily subcutaneous injection and compared with a placebo group. The healing process was radiologically monitored. Twenty-eight days after fracture biomechanical torsional testing was performed. The consolidation and callus composition, including quantification of cartilage and mineralized tissue, was traced in histomorphometrical investigations using an image analysis system. Both methods, the systemic administration of GH and the local application of growth factors, showed significant biomechanical and histological effects on fracture healing. The local growth factor application showed a stronger effect on fracture healing than the systemic GH injection. The combined application of both methods did not accelerate the effect on bone healing compared with the single application. It is therefore concluded that combining local and systemic stimulating methods does not provide further additive effects with regard to fracture healing. PMID- 12110431 TI - Effects of cerivastatin and parathyroid hormone on the lumbar vertebra of aging male Sprague-Dawley rats. AB - Both men and women lose bone at a late age (aging bone loss). The aim of this study was to determine whether cerivastatin and parathyroid hormone (PTH) can prevent aging bone loss in men. Bone loss in aged male Sprague-Dawley (SD) rats was used as a model for age-related bone loss in men. Nine-month-old male SD rats were divided into six groups: (1) baseline controls (killed at the beginning of the study); (2) age-matched controls; (3) parathyroid hormone (PTH; 80 microg/kg body weight per day for 5 days/week) treated; (4) low-dose cerivastatin (0.2 mg/kg body weight per day) treated; and (5) medium-dose cerivastatin (0.4 mg/kg body weight per day) treated; and (6) high-dose cerivastatin (0.8 mg/kg body weight per day) treated. Groups 2-6 were treated for 23 weeks between the ages of 9 and 15 months and killed at the end of 23 weeks. The fourth lumbar vertebra was analyzed using peripheral quantitative computed tomography (pQCT). It is shown that age-matched controls had decreased cancellous bone mineral content (Cn. BMC) by 19% (p < 0.05) and cancellous bone mineral density Cn. BMD) by 22% (p < 0.01) when compared with baseline controls. All three doses of cerivastatin resulted in lower Cn. BMC and Cn. BMD when compared with age-matched controls, but this decrease was not statistically significant. In the PTH-treated group, Cn. BMC increased by 5% (p < 0.0001) and Cn. BMD increased by 37% (p < 0.0001) when compared with age-matched controls. In age-matched controls, cortical bone mineral content (Ct. BMC) and cortical bone mineral density (Ct. BMD) decreased slightly, but not significantly, when compared with baseline controls. Ct. BMD did not change significantly at any of the three doses in the cerivastatin treated groups. In the PTH-treated group, Ct. BMC increased by 23% (p < 0.0001) when compared with age-matched controls. We confirmed that male SD rats lose bone with aging in the lumbar vertebra, and it is concluded that cerivastatin, at all doses administered, did not prevent this age-related bone loss. In contrast, PTH prevented age-related bone loss in the vertebra of male SD rats. PMID- 12110432 TI - Genistein, a soybean isoflavone, affects bone marrow lymphopoiesis and prevents bone loss in castrated male mice. AB - Soybean isoflavones exhibit selective effects on bone metabolism in postmenopausal women as well as in ovariectomized animals. Recently, the role of estrogen in bone metabolism in men has also received attention, because a man with a mutated estrogen receptor-alpha (ER(alpha)) gene will exhibit osteoporotic phenotypes. To examine the possible role of genistein, a soybean isoflavone, in bone marrow hemopoiesis and bone metabolism in men, male mice were orchidectomized (orx) and treated with genistein (0.4-0.8 mg/day) or 17beta estradiol (E(2); 0.03 microg/day) subcutaneously for 3 weeks. In orx mice, seminal vesicle weight decreased markedly, and it was not affected by the administration of genistein or E(2). The number of bone marrow cells was markedly increased after orx, and the majority was B-220 weakly positive pre-B cells. Increased B-lymphopoiesis was restored completely by E(2) or genistein administration. In orx mice, bone mineral density of the femur decreased markedly, and this bone loss was prevented to a significant extent by treatment with genistein as well as E(2). Histomorphometry showed that the trabecular bone volume in the femoral distal metaphysis decreased markedly after orx, and genistein and E(2) prevented this bone loss. These results suggest that soybean isoflavones prevent bone loss due to androgen deficiency in males. PMID- 12110433 TI - Mechanical strain and fluid movement both activate extracellular regulated kinase (ERK) in osteoblast-like cells but via different signaling pathways. AB - Extracellular regulated kinases (ERKs)-1 and -2 are members of the MAPK family of protein kinases involved in the proliferation, differentiation, and apoptosis of bone cells. We have shown previously that ROS 17/2.8 cells show increased activation of ERK-1 or -2, which is sustained for 24 h, when the strips onto which they are seeded are subjected to a 10 min period of cyclic four point bending that produces physiological levels of mechanical strain along with associated fluid movement of the medium. Movement of the strips through the medium without bending causes fluid movement without strain. This also increases ERK-1/2 activation, but in a biphasic manner over the same time period. Our present study investigates the role of components of signaling pathways in the activation of ERK-1/2 in ROS 17/2.8 cells in response to these stimuli. Using a range of inhibitors we show specific differences by which ERK-1 and ERK-2 are activated in response to fluid movement alone, compared with those induced in response to strain plus its associated fluid movement. ERK-1 activation induced by fluid movement was markedly reduced by nifedipine, and therefore appears to involve L-type calcium channels, but was unaffected by either L-NAME or indomethacin. This suggests independence from prostacyclin (PGI(2)) and nitric oxide (NO) production. In contrast, ERK-1 activation induced by application of strain (and its associated fluid disturbance) was abrogated by TMB-8 hydrochloride, L-NAME, and indomethacin. This suggests that strain-induced ERK-1 activation is dependent upon calcium mobilization from intracellular stores and production of NO and PGI(2). ERK-2 activation appears to be mediated by a separate mechanism in these cells. Its activation by fluid movement alone involved both PGI(2) and NO production, but its activation by strain was not affected by any of the inhibitors used. The G protein inhibitor, pertussis toxin, did not cause a reduction in the activation of ERK-1 or -2 in response to either stimulus. These results are consistent with earlier observations of ERK activation in bone cells in response to both strain (with fluid movement) and fluid movement alone, and further demonstrate that these phenomena stimulate distinct signaling pathways. PMID- 12110434 TI - Correcting calf girth discriminates the incidence of falling but not bone mass by broadband ultrasound attenuation in elderly female subjects. AB - Calf circumference has been cited as an independent risk factor for hip fracture. Correcting this measured girth for subcutaneous adipose tissue or fluid accumulation provides a more valid estimate of lean tissue, but has not been reported in elderly populations. Two hundred eighty-eight randomly selected female volunteers, aged >or=70 years, were assessed by quantitative ultrasound (QUS) and clinical risk factors as part of a larger screening study for hip fracture risk. This involved measuring broadband ultrasound attenuation (BUA) and administering a structured risk factor questionnaire that included estimated daily skeletal loading (time standing or walking). Body mass index (BMI) was estimated using current body mass and height at age 25 years. Calf girth was measured using a standard anthropometric tape, the medial calf skinfold (a vertical fold at the point of maximum calf girth) was measured, and corrected calf girth (CCG) was calculated by subtracting the skinfold (in centimeters) multiplied by pi from calf girth. Subjects were aged 76.9 +/- 5.0 years, had BMIs of 24.3 +/- 3.9 kg/m(2), and spent an average of 5.5 +/- 2.0 h on their feet each day. Age, current body mass, BMI calf girth, and CCG all correlated with BUA (p < 0.01). CCG correlated with hours on the feet (p < 0.05), whereas calf girth did not (p > 0.05). Dividing the sample into tertiles by these correlates of BUA and predicting BUA using stepwise regression revealed different predictors for each tertile. Of the total sample, 93 had fallen in the last 12 months, whereas 195 had not. Independent t-tests showed these groups to be similar in age, BMI, and calf girth (p > 0.05), but fallers spent less time on their feet each day, and had smaller CCG (p < 0.05). This suggests that larger calf muscles may be protective against falling-possibly as a result of enhanced stability or greater neuromuscular control. PMID- 12110435 TI - Heparan sulfate abnormalities in exostosis growth plates. AB - Hereditary multiple exostoses (HME), a condition associated with development and growth of bony exostoses at the ends of the long bones, is caused by germline mutations in the EXT genes. EXT1 and EXT2 function as glycosyltransferases that participate in the biosynthesis of heparan sulfate (HS) to modify proteoglycans. HS proteoglycans, synthesized by chondrocytes and secreted to the extracellular matrix of the growth plate, play critical roles in growth plate signaling and remodeling. As part of studies to delineate the mechanism(s) by which an exostosis develops, we have systematically evaluated four growth plates from two HME and two solitary exostoses. Mutational events were correlated with the presence/absence and distribution of HS and the normally abundant proteoglycan, perlecan (PLN). DNA from the HME exostoses demonstrated heterozygous germline EXT1 or EXT2 mutations, and DNA from one solitary exostosis demonstrated a somatic EXT1 mutation. No loss of heterozygosity was observed in any of these samples. The chondrocyte zones of four exostosis growth plates showed absence of HS, as well as diminished and abnormal distribution of PLN. These results indicate that, although multiple mutational events do not occur in the EXT1 or EXT2 genes, a complete loss of HS was found in the exostosis growth plates. This functional knockout of the exostosis chondrocytes' ability to synthesize HS chains further supports the observations of cytoskeletal abnormalities and chondrocyte disorganization associated with abnormal cell signaling. PMID- 12110436 TI - Behavior of osteoblast, adipocyte, and myoblast markers in genome-wide expression analysis of mouse calvaria primary osteoblasts in vitro. AB - Several genes, such as alkaline phosphatase, osteocalcin, and Cbfa1/Osf2, are known to be regulated during osteoblastic differentiation and are commonly used as "osteoblast markers" for in vitro or in vivo studies. The number of these genes is very limited, however, and it is of major interest to identify new genes that are activated or repressed during the process of osteoblast differentiation and bone formation as well as to extend the available information on gene families relevant to this particular differentiation pathway. To identify such genes, we have implemented a genome-wide analysis by determining changes in expression levels of 27,000 genes during in vitro differentiation of primary osteoblasts isolated from mouse calvaria. This study focuses on the description of the analytical and filtering process applied; on the transcriptional analysis of well-established "bone," "adipocyte," and "muscle" pathway markers; and on a description of the regulation profiles for genes recently described in the Skeletal Gene Database. We also demonstrate that new array technologies constitute reliable and powerful tools to monitor the transcription of genes involved in osteoblastic differentiation, allowing a more integrated vision of the biological pathways regulated during osteoblast commitment, differentiation, and function. PMID- 12110437 TI - Expression of leukemia inhibitory factor (LIF)/interleukin-6 family cytokines and receptors during in vitro osteogenesis: differential regulation by dexamethasone and LIF. AB - The leukemia inhibitory factor/interleukin-6 (LIF/IL-6) family of cytokines is known to play a major role in bone physiology. Although much work has focused on the regulation of bone resorption by IL-6 and related cytokines, their effects on osteoblast development and bone formation have not been as well studied. Previously, we reported that LIF inhibits, in a non-IL-6-dependent manner, osteoblast differentiation and bone nodule formation in the rat calvaria (RC) model, an effect that is antagonized by dexamethasone (Dex). The culture time sensitive window suggested that LIF targets late preosteoblasts or early osteoblasts, and that this stage-specific effect coincided with a period of low endogenous production of LIF and IL-6. To detect potential crosstalk between members of this family, we have extended these observations by assessing the expression levels of other LIF/IL-6 cytokines (CNTF, OSM, IL-11, CT-1) and their receptors in the same RC cell model treated with or without LIF or Dex. Semiquantitative reverse transcription-polymerase chain reaction (RT-PCR) analysis showed that IL-11 and its receptor, CNTF and its receptor, LIFR, and gp130 were constitutively expressed throughout the culture period. Expression of CT-1 and OSM increased with culture time - that is, with osteoblast differentiation - whereas the specific receptor for OSM (OSMR) was highly expressed at early timepoints and either plateaued or decreased thereafter. Continuous treatment with Dex (10(-8) mol/L) inhibited the endogenous production of IL-6, LIF, OSM, IL-11R, and OSMR, but had no detectable effect on the expression of IL-11, CT-1, CNTF, CNTFR, LIFR, or gp130. Finally, treatment with exogenously added LIF stimulated IL-6, LIF, LIFR, and OSMR, but had no other detectable effects. These data indicate that multiple members of the LIF/IL-6 family and their receptors are expressed in RC cell cultures, and are differentially regulated by Dex and LIF, suggesting that these cytokines play a complex and interdependent role, further modulated by glucocorticoid levels, in osteoprogenitor differentiation and bone nodule formation. PMID- 12110438 TI - Glucocorticoid-induced osteoporosis in the rat is prevented by the tyrosine phosphatase inhibitor, sodium orthovanadate. AB - Glucocorticoid-induced osteoporosis is characterized by decreased osteoblast numbers and a marked impairment of new bone formation. We found that, in vitro, dexamethasone inhibits both preosteoblast proliferation and mitogenic kinase activity in response to mitogens, and that inhibition of protein tyrosine phosphatases (PTPs) using sodium orthovanadate prevents this. Therefore, dexamethasone may act by either upregulating antiproliferative PTPs or downregulating promitogenic tyrosine-phosphorylated substrates. In this study, osteoporosis was induced in 3.5-month-old rats by subcutaneous injection with methylprednisolone 3.5 mg/kg per day for 9 weeks. Rats were treated with steroid alone or in combination with 0.5 mg/mL sodium orthovanadate, administered continuously in drinking water. Steroid-treated bones were significantly (p < 0.005) osteopenic (according to dual-energy X-ray absorptiometry) and physically weaker (p < 0.05) than controls. Quantitative bone histology confirmed a significant decrease in osteoid surfaces (p < 0.001), osteoblast numbers (p < 0.05), and rate of bone formation (p < 0.001). Concomitant treatment with vanadate largely prevented the densitometric, histologic, and physical abnormalities induced by prednisolone. This study supports our finding that PTPs are central to the negative regulation of osteoblast proliferation by glucocorticoids and, furthermore, suggests that PTP inhibitors such as sodium orthovanadate should be considered as novel anabolic agents for the treatment of steroid-induced osteoporosis. PMID- 12110439 TI - Adaptive adjustment of the adolescent porcine mandibular condyle. AB - The effect of occlusal support during primary dentition on the mandibular condyle remains controversial. We sought to determine whether unilateral loss of occlusal support leads to quantifiable adaptive changes of the condyle. Quantitative analysis of condylar growth and spongy bone volume after unilateral removal of teeth on the left side in adolescent minipigs was examined over a period of 4 months. Serial sagittal sections of the temporomandibular joint were examined using microradiography, fluorescence microscopy, and light microscopy. The condyles on the nonextracted side showed a higher growth rate than those on the extracted side, with a 1.56-fold thicker (p = 0.003) additional vertical bone layer. This factor was greater ventrally than dorsally (p = 0.0311), increasing from dorsomedial (1.33) to ventrolateral (2.38). There was therefore a reciprocal change of the condylar surface curve between the left and right condyles. Increased condylar growth correlated with a lower subchondral spongy bone volume (7.38% difference, p = 0.002). The amount of mineralized bone matrix generated was estimated to be about 1.33-fold higher in the nonextraction side condyles compared with those on the extraction side. Thus, unilateral loss of occlusal support was shown to lead to quantifiable alterations of condylar vertical growth and spongy bone volume in minipigs. PMID- 12110440 TI - Ultrasound stimulates nitric oxide and prostaglandin E2 production by human osteoblasts. AB - We have previously shown that the therapeutic range of ultrasound heals osteoradionecrotic bone and induces bone formation in vitro. It is well established that nitric oxide (NO) and prostaglandins are crucial early mediators in mechanically induced bone formation. The therapeutic range of ultrasound may act in the same way; therefore, we have investigated the effect of the therapeutic range of ultrasound on NO induction and prostaglandin E(2) (PGE(2)) production in vitro. Two ultrasound machines were evaluated, "traditional" (1 MHz, pulsed 1:4, tested at four intensities) and a "long-wave" (45 kHz, continuous, also tested at four intensities) devices. Ultrasound was applied to human mandibular osteoblasts for 5 min, and incubated at 37 degrees C for up to 24 h. The control group (sham insonated) was treated in the same way. NO was determined by measuring the nitrite concentration in the culture media colorimetrically, and PGE(2) was assayed by radioimmunoassay. Ultrasound produced a significant increase in both induced nitrite and PGE(2) production. The NO synthesis appeared to be via inducible NO synthase (iNOS) on the basis of the time course and levels of nitrite obtained, although the inhibition of other NOS isoforms by aminoguanidine cannot be excluded. PGE(2) synthesis appeared to be via COX-2. With the 45 kHz machine, a significant increase in NO was achieved at three intensities, 5, 30, and 50 mW/cm(2). The 1 MHz machine stimulated the synthesis of both NO and PGE(2), but was significant at only one dose (0.1 W/cm(2(SAPA))). There was no difference between the two machines with regard to PGE(2) synthesis. The time-course experiment revealed peak production to be 12-18 h for both NO and PGE(2). The therapeutic range of ultrasound stimulates both NO and PGE(2) synthesis by human osteoblasts, and the 45 kHz machine appeared to be more effective than the traditional short-wave length. These results may reflect the healing effect of ultrasound on fractures and osteoradionecrosis. PMID- 12110441 TI - Characterization of the bone-resorptive effect of interleukin-11 in cultured mouse calvarial bones. AB - Interleukin-11 (IL-11) is a stromal cell-derived cytokine that can enhance osteoclast formation and stimulate bone resorption. In the present study, the characteristics of the resorptive effect of IL-11 in mouse calvarial bones were investigated. Both recombinant mouse IL-11 and human IL-11 caused concentration- and time-dependent stimulations of (45)Ca release from prelabeled mouse calvariae. Half-maximal responses were obtained at 0.7 ng/mL (approximately 40 pmol/L). Mouse and human IL-11 also stimulated release of (3)H from [(3)H] proline-labeled bones. The magnitude of the (45)Ca and (3)H release (1.4-1.6 fold) caused by a maximally effective concentration of IL-11 was less than the stimulation (2.5-4.0-fold) elicited by a maximum concentration of parathyroid hormone (PTH). Release of (45)Ca by IL-11 was unaffected by the mitotic inhibitors, hydroxyurea and aphidicolin. In addition to resorption of bone, IL-11 caused a small (1.5-2.0-fold) enhancement of prostaglandin E(2) (PGE(2)) biosynthesis in calvariae, but had no effect on the mRNA expression of cyclooxygenase-1 and -2, or cytosolic phospholipase A(2). Indomethacin and flurbiprofen abolished the formation of PGE(2) and partially reduced (45)Ca release stimulated by IL-11. When either mouse interleukin-4 (IL-4) or interleukin-13 (IL-13) was added to calvariae treated with IL-11, (45)Ca release was inhibited. Resorption caused by IL-11 was also inhibited by both anti-mouse glycoprotein 130 (gp130) and an antibody neutralizing IL-11, but these agents had no effect on (45)Ca release caused by PTH or 1,25(OH)(2)vitamin D(3) (D(3)). Real time, quantitative polymerase chain reaction (PCR) analysis (TaqMan PCR) and semiquantitative reverse transcription-polymerase chain reaction (RT-PCR) demonstrated that IL-11 caused concentration-dependent enhancements of receptor activator of nuclear factor-kappaB ligand (RANKL) and osteoprotegerin (OPG) mRNA, without affecting the mRNA expression of RANK. Mouse RANKL stimulated (45)Ca release in the calvarial bones. The stimulatory effects of RANKL and IL-11 were inhibited by mouse OPG. These data demonstrate that IL-11 stimulates osteoclastic resorption in mouse calvariae by mechanisms that are independent of cell proliferation; partially dependent on prostaglandin biosynthesis; sensitive to inhibition by IL-4, IL-13, and OPG; and associated with enhanced expression of RANKL and OPG. In addition, IL-11 was not found to play an essential role in resorption stimulated by other calciotropic agents in calvariae. PMID- 12110442 TI - Regulation of receptor activator of nuclear factor-kappa B ligand and osteoprotegerin mRNA expression by parathyroid hormone is predominantly mediated by the protein kinase a pathway in murine bone marrow cultures. AB - Parathyroid hormone (PTH) stimulates receptor activator of nuclear factor-kappaB ligand (RANKL) mRNA and inhibits osteoprotegerin (OPG) mRNA expression in murine bone marrow cultures. To understand the mechanisms influencing these responses, we investigated the role of the protein kinase A (PKA) and protein kinase C (PKC) pathways in the regulation of RANKL and OPG mRNA expression in murine bone marrow cultures. Murine bone marrow cells were stimulated with bovine PTH(1-34) and (1 34) amide, which activate both pathways; PTH(3-34), which more selectively activates the PKC and calcium pathways; and human PTH (1-31), which stimulates adenylyl cyclase, but not protein kinase C. We also examined agents that more directly activate either the PKA pathway (forskolin [FSK] and 8-bromo cAMP [8-Br cAMP]) or the PKC pathway (phorbol 12-myristate 13-acetate [PMA]) in murine bone marrow cultures. After 1 h, RANKL mRNA expression was stimulated to a similar degree by agents that activate either or both the PKA and PKC pathways. However, this effect was sustained for 24 h only with agents that stimulated PKA. OPG mRNA expression was inhibited by all agents that stimulated PKA at 6 h. In contrast, PKC-specific stimulators [PMA and bPTH(3-34)] had no effect on OPG regulation in this culture system. To determine the involvement of the PKC signaling pathway in responses of RANKL, bone marrow cells were pretreated with PMA for 24 h and then treated with PTH(1-34) or FSK for 2 h. PMA pretreatment did not alter the ability of PTH or FSK to stimulate RANKL or inhibit OPG mRNA expression. Treatment of cells with H-89, a PKA inhibitor, significantly reduced the ability of PTH and FSK to induce RANKL and inhibit OPG mRNA expression. Calphostin C, a PKC inhibitor, significantly reduced PMA-stimulated RANKL mRNA expression without altering PTH- or FSK-mediated effects on RANKL or OPG mRNA. Cycloheximide, an inhibitor for protein synthesis, inhibited PTH-stimulated RANKL mRNA expression by 60% without altering the effect of PTH on OPG mRNA expression. To examine the involvement of prostaglandin in PMA-mediated responses, cells were treated with indomethacin, a nonspecific prostaglandin G/H synthase (PGHS) inhibitor, or NS 398, a selective inhibitor of PGHS-2. Neither PGHS inhibitor altered PMA-induced effects on RANKL and OPG mRNA expression. These results demonstrate that the PKA pathway is predominantly involved in the effects of PTH on RANKL mRNA expression in murine bone marrow cultures, but there is also a PKC-mediated response, which is not sustained. Inhibition of OPG by PTH appears to be a selective PKA response. PMID- 12110443 TI - Role of polyunsaturated fatty acids in the inhibitory effect of human adipocytes on osteoblastic proliferation. AB - As previously reported, the association of bone loss with an increase in bone marrow adipose volume may be related to the inhibition of human osteoblastic cell proliferation in the presence of human adipocytes. In the osteoblastic supernatant, fatty acid composition varied after coculture with mature adipocytes, with a marked increase in the proportion of docosahexaenoic acid (22:6 n-3; DHA) (+90 +/- 8%). This suggests that polyunsaturated fatty acids (PUFA) may contribute to the inhibitory effect of adipocytes on osteoblastic cell proliferation. The purpose of the present study was to evaluate the effects of two PUFA, DHA and arachidonic acid (20:4 n-6; AA), on the proliferation of primary human osteoblastic (hOB) cells and human osteosarcoma cell line, MG-63. The effects of cholesterol and oleic acid, a monounsaturated FA (18:1 n-9; OA), both being present in adipocyte lipidic vacuoles, were also investigated. At between 10 and 50 micromol/L, DHA and AA induced a significant dose-dependent decrease in hOB cell proliferation (p < 0.0001 and p < 0.006 for DHA and AA, respectively) when compared with control hOB cells exposed to the vehicle (bovine serum albumin). This inhibition reached -50% with 50 micromol/L of DHA or 20 micromol/L of AA. This effect was not related to cell apoptosis, as shown by terminal deoxynucleotidyltransferase-mediated dUTP-fluorescein nick end labeling (TUNEL) and Hoechst dye staining. In contrast, OA and cholesterol had no effect on hOB cell proliferation, even at a high concentration (200 micromol/L). Similar results were observed with regard to MG-63 cell proliferation. In addition, flow cytometric analysis showed that the number of hOB cells in the S phase of the cycle was twofold lower when treated with 50 micromol/L of DHA or AA. In vitro results indicate that mature adipocytes may contribute to age-related bone loss through the release of polyunsaturated fatty acids, which impair osteoblastic proliferation. PMID- 12110444 TI - Effects of predictable and unpredictable intermittent noise on spatial learning in rats. AB - The effects of predictable (periodic) and unpredictable (aperiodic) intermittent noise of moderate intensity (68 dB) on the learning of a complex T-maze by genetically defined rats were investigated. In Experiment 1, three groups (n=8) of rats learned a multiple T-maze, one group under control conditions, one group with predictable intermittent noise and one group with unpredictable intermittent noise. Results showed a profound effect of noise on learning and behavioural scores. Noise-exposed animals made less errors, finished their trials sooner and explored less. There was no difference between predictable and unpredictable noise. Further tests, during which formerly noise-exposed groups learned a new route under control conditions (Experiment 2) or the former controls learned a new route with noise (Experiment 3), suggest that the effects of noise on learning were caused by an effect of noise on memory formation and/or retrieval, rather than by long-term shifts in behavioural strategies. PMID- 12110445 TI - Sensory preconditioning in rats with lesions of the anterior thalamic nuclei: evidence for intact nonspatial 'relational' processing. AB - Rats with neurotoxic lesions centered in the anterior thalamic nuclei were trained in two versions of a nonspatial, sensory preconditioning procedure. In both versions, two stimulus compounds (AX and BY) were first presented and then X, but not Y, was paired with an aversive unconditioned stimulus. This procedure resulted in greater conditioned responding to A than B. Anterior thalamic lesions had no apparent effect on these two examples of sensory preconditioning, nor did they affect fear conditioning or conditioned taste aversion. In contrast, the same lesions led to a severe deficit on a test of spatial memory. These results help to refine our understanding of the contribution of the anterior thalamic nuclei to spatial memory. PMID- 12110446 TI - CREB phosphorylation as a molecular marker of memory processing in the hippocampus for spatial learning. AB - Regulation of gene transcription via the cyclic adenosine 3',5'-monophosphate (cAMP)-mediated second messenger pathway has been implicated in learning and memory. Although the cAMP response element-binding protein (CREB) is an important transcription factor involved in long-term memory, it remains to be determined whether the CREB-dependent events are attributed to spatial learning and memory in a radial arm maze. Here we demonstrate that cAMP-dependent protein kinase A (PKA) and CREB are activated in the course of spatial learning. The radial maze training in rats resulted in a significant increase in PKA and CREB phosphorylation in the hippocampus in the course of spatial learning, which was followed by spatial memory formation. On the other hand, neither the phosphorylation of Ca2+/calmodulin-dependent protein kinase II (CaMKII) and extracellular signal-regulated kinase (ERK) nor the mRNA level of brain-derived neurotrophic factor was significantly affected. These results suggest that activation of the PKA/CREB signaling pathway in the hippocampus plays an important role in spatial memory formation. PMID- 12110447 TI - Differential responses in central dopaminergic activity induced by apomorphine in IPL nude rat. AB - The IPL nude rat, derived by spontaneous mutation from the Sprague-Dawley strain, presents alterations in the prolactin synthesis and secretion due to an increased dopaminergic inhibition. However, there are no reports concerned to central dopamine activity. The corpus striatum is a brain area involved in the development of stereotyped behavior after the activation of mesolimbic and/or nigro-striatal dopamine pathways. In order to identify possible mesolimbic and/or nigro-striatal dysfunctions in the IPL nude rat, we study the spontaneous oral behaviors and the effects of apomorphine-induced dopaminergic activation on stereotyped behavior and neurochemical changes. Males from both strains were injected with saline or apomorphine (2 and 5 mg/kgs.c.) and evaluated during 30 min in a stereotypes oral tests. The corpus striatum and nucleus accumbens were used to measure dopamine (DA), 3,4-dihydroxyphenylalanine (DOPA) and 3,4 dihydroxyphenylacetic acid (DOPAC) by HPLC. The concentrations were expressed as synthesis rate (DA/DOPA) and turnover rate (DOPAC/DA). We observed that the spontaneous gnaw movements were significantly different between the untreated IPL nude and Sprague-Dawley (SD) rats. Apomophine injection decreased the amount of stereotyped gnawing in IPL nude rats at the two doses used, but it induced an increase in SD rats. Apomorphine also caused an enhancement in the number of biting and sniffing without modifying the licking behavior. In addition, modifications of the dopaminergic activity were also observed. Synthesis rate in the striatum of IPL nude rats was higher than in SD rats after the injection of saline. Apomorphine caused a reduction of the synthesis rate in both strains. Turnover rate was significantly lower in the striatum of IPL nude rats than in the SD rats injected with saline. Apomorphine caused an increase in the turnover rate in both strains. Contrary to observed in the striatum, the 2 mg/kg dose of apomorphine caused a significant increase in the DA synthesis rate in nucleus accumbens, while 5 mg/kg decrease it in both strains. The DA turnover rate in the same area was lower in IPL nude than in SD rats after saline injection. Apomorphine enhances the DA turnover rate in both strains. We conclude that the modifications of the oral spontaneous and induced stereotypical patterns observed in the IPL nude rats could be related to the differential responses in dopaminergic activity in the two brain areas examined. PMID- 12110449 TI - Time-dependent involvement of PKA/PKC in contextual memory consolidation. AB - Rats were implanted bilaterally with cannulae into the dorsal hippocampus and trained in a Pavlovian fear-conditioning paradigm. Four groups of rats were infused intra-cranially with 1-(5'-isoquinolinesulfonyl)-2-methylpiperazine (H7 dihydrochloride), a potent inhibitor of both protein kinase C (PKC) and cAMP dependent protein kinase (PKA), at different time intervals in order to examine their involvement in the acquisition and consolidation of contextual fear memory. We demonstrate a significant consolidation deficit of long-term contextual fear conditioning memory that is maximal when PKA and PKC are inhibited at 90 min post training. These results suggest the existence of a critical time window, during which these enzymes must be activated for the consolidation of long-term memories. PMID- 12110448 TI - Postnatally induced differences in adult pain sensitivity depend on genetics, gender and specific experiences: reversal of maternal deprivation effects by additional postnatal tactile stimulation or chronic imipramine treatment. AB - Postnatal endotoxin exposure, handling or maternal deprivation produce long lasting individual differences in various neuroendocrine and behavioural responses. However, the impact of postnatal experiences on adult pain sensitivity and its reversibility by postnatal additional tactile stimulation or antidepressants in adulthood is not well understood. Therefore, postnatal endotoxin application as a model for infection, maternal deprivation as a model for depression, and postnatal handling as a model for stimulation were compared with respect to the effects on pain sensitivity in adult Fischer 344 (F344) and Lewis (LEW) rats. Handling increased hot plate latencies in adult F344 and LEW rats, while maternal deprivation shortened hot plate latencies only in LEW rats. Prophylactic treatment strategies, such as tactile stimulation of the dorsal neck region of pups directly after maternal deprivation, or chronic treatment of adult maternally deprived rats using imipramine, successfully provide protection against the maternal deprivation-induced shortening of hot plate latencies. Thus, there is considerable specificity of certain postnatal experiences in modulating adult pain sensitivity and the maternal deprivation-induced hyperalgesia is reversible by different interventional regimes. These findings may explain some of the individual differences in pain sensitivity of humans and the differential efficacy of antidepressants in pain syndromes. PMID- 12110450 TI - Impairment of pronation, supination, and body co-ordination in reach-to-grasp tasks in human Parkinson's disease (PD) reveals homology to deficits in animal models. AB - Animal (monkey, rat, mouse) models are widely used to investigate degenerative processes and potential therapeutic treatments for human Parkinson's disease (PD). One task that has proved useful in these investigations is a reach-to-grasp task (skilled reaching) in which an animal reaches for a piece of food that it then consumes. Rats with extensive unilateral Dopamine depletions are impaired in using the contralateral limb. The qualitative features of posture, lifting and advancing the limb, pronating the paw to grasp food, and in withdrawing and supinating the paw to place the food in the mouth are impaired, as is reaching success. Humans with PD are often described as having poor manual dexterity that worsens as the disease progresses. As there have been no detailed comparisons of reaching movements in the animal models and in PD subjects, the following descriptive analysis was performed. Ten subjects with PD, eight age matched controls and 14 young normal subjects were studied as they used a natural movement of reaching for a small piece of food that they then placed in the mouth to eat. The reaching movements were described using Eshkol-Wachman Movement Notation (EWMN), supplemented with kinematic analyses. From this description, a 21-point rating scale was devised to describe the component movements of the reach. Movements included: orienting the head and eyes to the target, adjusting posture, lifting the hand, shaping and aiming the digits to the target, pronating the hand to grasping the food with a pincer grip, lifting and supinating the hand to transporting the food to the mouth, and further supinating the hand and opening the digits to place food in the mouth, and finally returning the hand to the starting position. Analysis indicated that most aspects of the reaching movements of the PD subjects were significantly different relative to both young control subjects and old control subjects. As compared to the control groups, postural and reaching components of the movements were fragmented, movements were achieved using more proximal segments of the body, and rotatory movements of the hand were limited. The PD subjects did use a pincer grasp to obtain the food, but the grasp was less independent of other digit movements than was observed in the control subjects. These results are discussed in terms of a homology to impairments displayed animal models of PD. PMID- 12110451 TI - Arginine vasotocin effects on courtship behavior in male white perch (Morone americana). AB - Arginine vasotocin (AVT) and its mammalian homologue, arginine vasopressin (AVP), have been shown to have widespread behavioral effects in vertebrates. AVT was evaluated for its effectiveness in stimulating an important courtship behavior termed 'attending' in male white perch, Morone americana. Attending consists of close and continuous following of the female with occasional contact in the abdominal area. We tested the behavioral effectiveness of AVT in stimulating attending when administered either intraperitoneally (IP) or intracerebroventricularly (ICV). We also tested IP injections of AVT alone and in combination with an AVP V(1) receptor antagonist (Manning compound). None of the IP injections of either AVT or Manning compound produced consistent effects on attending behavior. In contrast, ICV injections of AVT did significantly increase attending behavior and at low dosages. Circulating levels of testosterone and 11 ketotestosterone were not affected approximately 80 min following injection by any of the treatments. The strong behavioral effects observed with ICV administration support a central site of action for AVT in stimulating attending behavior. This is a complex behavior that shows similarities to behaviors mediated by AVT and AVP in other vertebrates, providing further evidence of a conserved behavioral role for these peptides. PMID- 12110453 TI - Changes in the septohippocampal cholinergic system following removal of molar teeth in the aged SAMP8 mouse. AB - We investigated the effect of dysfunctional teeth on age-related changes in the septohippocampal cholinergic system by assessing acetylcholine (ACh) release and choline acetyltransferase (ChAT) activity in the hippocampus and ChAT immunohistochemistry in the medial septal nucleus and the vertical limb of the diagonal band in young-adult and aged SAMP8 mice after removal of their upper molar teeth (molarless condition). Aged molarless mice showed decreased ACh release and ChAT activity in the hippocampus and a reduced number of ChAT immunopositive neurons in the medial septal nucleus compared to age-matched control mice, whereas these effects were not seen in young-adult mice. The results suggest that the molarless condition in aged SAMP8 mice may enhance an age-related decline in the septohippocampal cholinergic system. PMID- 12110452 TI - Prenatal cocaine exposure alters sensitivity to the effects of idazoxan in a distraction task. AB - The present study was designed to test whether prenatal cocaine (COC) exposure alters sensitivity to the attentional effects of idazoxan (IDZ), an alpha-2 adrenergic antagonist that increases coeruleocortical NE activity. The task assessed subjects' ability to selectively attend to an unpredictable light cue and disregard olfactory distractors. IDZ increased commission errors specifically under conditions of distraction, an effect that was similar in the COC and control groups. In contrast, COC animals were significantly more sensitive than controls to the effects of IDZ on omission errors and nontrials. The pattern of effects suggests that the differential treatment response to IDZ on these latter measures resulted from an alteration in norepinephrine (NE)-modulated dopamine release in the COC animals, reflecting lasting changes in dopaminergic and/or noradrenergic systems as a result of the early cocaine exposure. Based on the behavioral measures that showed a differential response to IDZ in the COC animals, it seems likely that these changes may contribute to the alterations in sustained attention and arousal regulation that have been reported in both animals and humans exposed to cocaine in utero. PMID- 12110454 TI - Lateralization in human nasal chemoreception: differences in bilateral electrodermal responses related to olfactory and trigeminal stimuli. AB - The study of olfactory lateralization in humans has given rise to many publications, but the findings have often been contradictory. There is growing evidence to suggest that the nature of the olfactory stimulus influences the processes of lateralization. An important factor could be the trigeminal component. Indeed, most odorants simultaneously stimulate both olfactory (CN I) and trigeminal (CN V) systems which differ in terms of their central projections, ipsilaterally for CN I and contralaterally for CN V. The aim of this study was to investigate variations in psychophysiological measurements between a nasal input with low (phenyl ethyl alcohol (PEA)) and high (allyl isothiocyanate (AIC)) intranal trigeminal stimulation. In a first experiment (20 subjects), the intensity, hedonicity and irritation levels of stimulus were tested with a psychophysical evaluation to study the possible influences of perceptual characteristics. A second experiment (37 subjects) used bilateral electrodermal recordings and compared the skin conductance responses (SCRs) for both nasal inputs on either monorhinal and birhinal stimulations. Firstly, the electrodermal activity (EDA) results showed no differences between the two nostrils for PEA as well as AIC, but differences in relation to the type of stimulus, e.g. higher amplitude in response to AIC versus PEA. Secondly, the results indicated bilateral differences in EDA recordings related to the nature of the stimulus and are discussed in terms of hemispheric asymmetric activation. PMID- 12110455 TI - Adolescent cocaine exposure and offensive aggression: involvement of serotonin neural signaling and innervation in male Syrian hamsters. AB - Repeated low-dose cocaine treatment (0.5 mg/kg/day) during adolescence facilitates offensive aggression in male Syrian hamsters (Mesocricetus auratus). The current study assessed whether adolescent cocaine-facilitated offensive aggression was inhibited by increased serotonin activity and if cocaine exposure during this developmental period influenced serotonin development in the primary aggression areas of hamster brain. In a first experiment, hamsters were treated with low doses of cocaine throughout adolescence and then scored for offensive aggression following the systemic administration of vehicle or fluoxetine, a selective serotonin reuptake inhibitor. Vehicle-treated hamsters showed high levels of offensive aggression, while treatment with fluoxetine inhibited the cocaine-facilitated aggressive response. Only one out of ten fluoxetine-treated animals both attacked and bit intruders, compared to nine out of ten saline treated animals. In a second experiment, hamsters were administered low doses of cocaine or saline throughout adolescence, tested for offensive aggression, and then examined for differences in serotonin afferent innervation to regions of the hamster brain implicated in aggressive responding. Aggressive cocaine-treated hamsters showed significant reductions (35-50%) in the number of serotonin immunoreactive varicosities and fibers in several aggression areas, including the anterior hypothalamus, lateral septum, medial amygdala, and bed nucleus of the stria terminalis. Together, these results support a role for serotonin innervation and function in adolescent cocaine-facilitated offensive aggression. PMID- 12110456 TI - Noradrenergic control of auditory information processing in female canaries. AB - An ethological procedure, based on the study of the sexual responsiveness of female canaries (Serinus canaria) to song playbacks was used to investigate the function of central noradrenergic inputs in the processing of auditory information. The effects of a noradrenergic denervation on sexual responses was analyzed in females exposed to playbacks of biological relevant auditory stimuli, i.e. sexually stimulating songs, presented alone or masked by auditory distractors. A decrease in behavioral responsiveness was observed as a function of the amount of masking distractors indicating that female canaries have the perceptual ability to discriminate and selectively attend to biologically relevant songs. After the systemic administration of DSP-4, a specific noradrenergic neurotoxin, females exhibited an overall decrease in sexual responsiveness to songs masked or not by distractors. No effect of DSP-4 were detected on the motor activity nor on reproductive behaviors. These results indicate that central noradrenergic inputs modulate the sexual behavior of female canaries by affecting the auditory processing of relevant information contained in sexually stimulating songs. PMID- 12110457 TI - The behavioral and dendritic growth effects of focal sensorimotor cortical damage depend on the method of lesion induction. AB - Using different models of focal cortical injury in adult rats, the neural structural and behavioral outcomes of unilateral lesions of the forelimb representation of the sensorimotor cortex (SMC) were assessed. Lesions were produced using either electrolytic, aspiration, or combined ('electroaspiration') techniques. Measurements of dendritic arborization in layer V of the motor cortex opposite the lesion revealed a growth of pyramidal neuron dendritic processes following electrolytic lesions in comparison to shams. This effect was not found in either the aspiration or electroaspiration lesion groups. Behaviorally, animals in all lesion groups developed a hyper-reliance on the forelimb ipsilateral to the lesion and proportionate disuse of the contralateral (impaired) forelimb for postural support behaviors. In comparison to sham operated animals, the initial asymmetries in behaviors expressed during movement were similar between lesion groups, but were less enduring following electrolytic lesions than following aspiration and electroaspiration lesions. Furthermore, both aspiration lesion groups had more prevalent adduction of the impaired forelimb than the electrolytic-only lesion rats. Thus, cortical aspiration resulted in more severe and enduring forelimb impairments than the electrolytic lesions, despite similar lesion sizes, as assessed using cortical volume measures. These findings suggest that the aspiration lesion procedures, at least as performed in the present study, exacerbate the behavioral effects of focal cortical injury and limit compensatory plasticity in the contralateral cortex. PMID- 12110459 TI - Differential learning strategies in spatial and nonspatial versions of the Morris water maze in the C57BL/6J inbred mouse strain. AB - We recently developed a new nonspatial version of the Morris water maze that requires the use of four visually distinct intra-maze patterns to efficiently locate a hidden platform. The nonspatial version was designed to match the spatial version on complexity of cue usage, and differs only on spatiality of cues, thereby allowing more meaningful comparisons between the two versions. Following a previous experiment that demonstrated nonspatial learning with the BXSB inbred mouse strain, C57 inbred mice were tested in this study. They received spatial and nonspatial training in a counter-balanced order so that Test Order and information transfer could be assessed. Subjects that received spatial training first had superior performance in both the spatial and the nonspatial tasks when compared to mice that received nonspatial training first. The mice that received spatial training first used extra-maze cues as a spatial strategy. However, during nonspatial testing they did not use the intra-maze cues to locate the platform; instead, the mice used an egocentric strategy of circling through the platform annulus. Subjects that received spatial testing first were superior on the nonspatial task to those subjects that received nonspatial training first. Moreover, subjects that received nonspatial testing first were unable to learn the spatial version. Overall, C57 mice can learn both the spatial and nonspatial versions of the Morris maze presented here; however, the nonspatial version is more difficult and is solved using an egocentric strategy. PMID- 12110458 TI - Spatial and nonspatial Morris maze learning: impaired behavioral flexibility in mice with ectopias located in the prefrontal cortex. AB - About half of BXSB/MpJ-Yaa (BXSB) mice have neocortical ectopias (misplaced clusters of neurons located in layer I of cortex). Previous behavioral studies have suggested that ectopic mice have superior spatial, but equivalent nonspatial, reference memory learning. However, since spatial and nonspatial learning were not assessed in the same apparatus and with the same testing procedure, it is unclear if this conclusion is accurate. We have created a new nonspatial Morris maze for mice that differs from the spatial task only in the type of cues that must be utilized to efficiently locate the platform (intra-maze black/white patterns vs. extra-maze room cues) and does not differ in the level of task complexity or the presence of objects within the maze. Ectopic mice were very good in utilizing extra-maze cues when learning the spatial version and in utilizing intra-maze cues when learning the nonspatial version of the Morris maze, while non-ectopics were not, suggesting that ectopics have superior spatial and nonspatial reference memory. Ectopias in BXSB mice are usually located in prefrontal and/or motor cortex. The prefrontal cortex is involved in behavioral flexibility (e.g. being able to easily switch from using spatial to nonspatial cues). Only ectopic mice with ectopias specifically located in the prefrontal region of cortex demonstrated difficulty switching from using extra-maze to intra maze cues and vice versa. Thus, the presence of one or more ectopias in the prefrontal region of cortex disrupted one of the normal functions of the prefrontal cortex. PMID- 12110460 TI - Early postnatal protein malnutrition affects learning and memory in the distal but not in the proximal cue version of the Morris water maze. AB - Learning and memory of early postnatal protein malnourished rats were investigated in the Morris water maze. During the lactation period (21 days) each litter (mother plus six male and two female pups) was provided with 16% (well nourished) or 6% (malnourished) protein diets. After weaning, rats remained on the same diet until 49 days of age. From day 50 on all animals were fed a commercial lab chow. Experiments started on day 70. In experiment I (proximal cue version) the animals were trained to escape from water to a visible platform (3 cm above the water level) in six trials daily for four consecutive days, completing 24 trials. In experiment II (distal cue version) the animals were trained to escape from water to a submerged platform using the same procedure as in experiment II. After the 24th trial, the platform was removed and the animals were submitted to a 60-s trial (probe trial). Seven and twenty-eight days after training, the retention test was conducted in one 180-s trial. The results showed no impairment of the learning or memory of malnourished animals tested in the proximal cue version but an increased latency and distance traveled to find the submerged platform in the distal cue version of the procedure. In the distal cue version the malnourished animals also showed increased latency to find the platform 7 and 28 days after the test training. No difference due to diet was found in the probe trial test indicating that, once the task is acquired, malnourished rats can manage extra-maze cues as easily as well-nourished rats. It is suggested that the present results can be due to alterations produced by protein malnutrition in the hippocampal formation or also to reflect the higher emotionality of rats following early malnutrition, specially considering the fact that postnatally malnourished animals are more reactive to unpleasant or aversive stimuli as cold water. PMID- 12110461 TI - Hippocampal lesioned rats are able to learn a spatial position using non-spatial strategies. AB - In the last two decades, many experiments have demonstrated that the hippocampus plays a role in the learning and processing of spatial and contextual information. Despite these demonstrations, some recent publications have indicated that the hippocampus is not the only structure involved in spatial learning and that even after hippocampal lesions, rats can perform spatial tasks. However, it is not well established whether animals with hippocampal dysfunction still have some spatial learning capacities or develop non-spatial solutions; these may require lengthier acquisition training. We now report the effects of conventional, dorsal hippocampal ablation on rats' performance on the water maze. We tested rats using a short (4 days) versus a long (16 days) acquisition period. We demonstrated that animals with dorsal hippocampal lesions have some residual capacity for learning the localization of a hidden escape platform in a pool during both a reference memory task and a working memory task. The animals with dorsal hippocampal lesions learned to escape at a fixed location, but only with extended training. It is suggested that these animals used non-spatial strategies to compensate for a spatial memory impairment. The results are discussed with respect to the experimental procedure and the strategy applied by the lesioned rats. PMID- 12110462 TI - Chemosensory input and lateralization of brain function in the domestic chick. AB - One-day old domestic chicks (Gallus gallus domesticus) show concentration dependent behavioural responses to olfactory cues. In the present study we investigated the lateralized olfactory responses of 1-day-old chicks to the odours of eugenol and iso-amyl acetate. In experiment 1 different concentrations of each odour were presented in repeated trials to chicks housed individually. The odours were presented together with a small coloured bead at which the chick pecked. When tested with the highest concentration of eugenol (100% v/v), the chicks demonstrated more head shaking when their left nostril was occluded (RN; right nostril in use) than when their right nostril was occluded (LN; left nostril in use). No such lateralization occurred in response to iso-amyl acetate. This result was confirmed in a second experiment in which the chicks were tested with unscented stimuli, 100% eugenol and 100% isoamyl acetate. In experiment 3 we found that occluding both the chicks' nostrils abolished the head shaking response to eugenol and to iso-amyl acetate. Thus, the chicks' head shaking responses to the odorants eugenol and iso-amyl acetate are mediated primarily by inputs from within the nasal cavity, and not by oral or occular inputs. The present results are consistent with the hypothesis that there is lateralization to olfactory cues and that it is dependent on the involvement of receptors inside the nasal cavity. We suggest that differences in lateralized olfactory responses to different odours are affected by the relative involvement of intranasal olfactory and trigeminal chemoreceptors. PMID- 12110463 TI - Running increases ethanol preference. AB - Wheel running performed by rats is reinforcing, rewarding and possibly addictive. In this study we analyzed if wheel running could affect ethanol preference. Lewis rats, known to be both addiction-prone and to develop an excessive wheel running behavior, were given access to ethanol in a two-bottle free-choice paradigm. The animals reached a high and stable ethanol intake after 5 weeks. In the next phase, rats were subjected to ethanol withdrawal for 1, 2 or 4 weeks with or without access to running wheels. Finally animals were again given access to ethanol in the same two-bottle free-choice paradigm, combined with access to running wheels. The rats that ran in running wheels during 1 or 2, but not 4, weeks of ethanol withdrawal increased both ethanol intake and preference as compared with the control group that did not have access to the wheels. Previous studies have demonstrated that low doses of morphine increases ethanol preference. Here we show that also running potentiates ethanol intake and preference. Thus, running which shares many of the reinforcing properties with addictive drugs appears to potentiate rats to an increased preference for ethanol. Our results describe a behavioral interaction where running increases ethanol consumption. PMID- 12110464 TI - Unihemispheric memory in pigeons-knowledge, the left hemisphere is reluctant to share. AB - In the present study, pigeons were trained under binocular conditions in a conditional visual discrimination in which they were faced with two identical patterns arranged one above the other. In half of these stimulus pairs the animals had to peck the upper pattern, in the other half the lower one. Although only six pairs of stimuli were used, only four out of eight birds reached learning criterion. These animals needed up to 6 months of training with 3050 to 6650 trails. Then, the experiment proceeded under identical conditions using eye caps restricting vision alternatively to the left or the right eye. These monocular tests revealed that three out of four birds virtually had no knowledge of the task contingencies using their left eye (right hemisphere). Again, several thousand trials were needed to train the birds to criterion with their left eye, while they were simultaneously discriminating at a very high level with their right. These results show that memories on task contingencies are stored unihemispherically in the visually dominant left side despite extensive training with both eyes open. Additionally, it can be concluded that the subsequent read out by the 'naive' hemisphere can be largely restricted, resulting in a 'natural split-brain' like situation in birds. It is speculated that the absence of a corpus callosum in birds restricts interhemispheric transfer of information. PMID- 12110465 TI - Limitations of the sensitive period for sexual imprinting: neuroanatomical and behavioral experiments in the zebra finch (Taeniopygia guttata). AB - In the course of developmental sensitive periods, an animal receives input from the environment which shapes its behaviour and neuronal connectivity. While normally restricted to early development, it has been shown frequently that an extension of the sensitive period to later ages is possible by sensory deprivation or inadequate stimulation. This raises the question whether sensitive periods can be shifted to any age, or whether a time window exists within which sensitive period shifts are possible. We show here for sexual imprinting that such a time window exists, and we also show that the spine density changes during development in brain areas involved in imprinting predict the limits for sensitive period shifts. Based on these results, we speculate about the mechanisms which may underlay the regulation of spine density and thus the imprinting process. PMID- 12110466 TI - Emotional changes related to age in rats--a behavioral analysis. AB - The present study investigated age-related differences in the emotional behavior of rats using factor analysis to identify motivational factors influencing spontaneous behavior in open field with illuminated center (OF), plus maze (EPM) and social interactions test. Animals of the same strain, bred under the same conditions, formed two experimental groups: young adults (YA, N=20) tested at the age of 4 months and old rats (OA, N=16) tested at the age of 24 months. The computer video based tracking system EthoVision was used for automated acquisition and analysis of data. The results of each test were analyzed separately for YA and OA by factor analysis. Two main independent factors emerged from the analysis of OF measures-factor 1, which appeared to reflect motor activity, and factor 2, reflecting anxiety. The measures best reflecting motor activity (distance moved in the peripheral zone) and anxiety (time spent in central zone) decreased significantly with age. Factor analysis for EPM measures revealed, in both groups, three independent factors. In YA, factor 1 reflected motor activity, factor 2-anxiety, in OA measures of anxiety loaded on factor 1, measures of activity on factor 2. Factor 3 in both groups appeared to represent a decision making process. The number of entries to the closed arms declined significantly in OA, showing an age related decrease of motor activity. Also, the ratio of open arms entries in relation to the total number of entries decreased in OA, indicating a higher anxiety level. Three independent factors emerged from the analysis of social interaction measures. The pattern of factor loading was different in young and old animals, although the number and time of social interactions did not show age-related differences. In addition to a decrease of motor activity we conclude that old rats also differ from young animals in emotional and social behavior. PMID- 12110467 TI - The partial reinforcement extinction effect in humans: effects of schizophrenia, schizotypy and low doses of amphetamine. AB - The partial reinforcement extinction effect (PREE) was studied in human subjects. It has been suggested that the PREE depends on neural mechanisms critical to the cognitive dysfunction which underlines acute schizophrenia. We therefore predicted that the PREE should be reduced, through decreased resistance to extinction in the partial reinforcement (PR) condition, in various types of individual: (a) healthy volunteers given low doses of oral amphetamine; (b) those in the acute (but not chronic) phase of a schizophrenic illness and; (c) healthy volunteers with high scores on personality measures of schizotypy. Despite obtaining robust demonstrations of PREE in all experiments, none of these predictions were confirmed. A single, low dose, of amphetamine had no effect on either continuous reinforcement (CR) or partial reinforcement (PR). Acute and chronic schizophrenic patients showed a reduced PREE compared to controls. However this was due to increased resistance to extinction in the CR groups. Finally, high schizotypy scores were associated with greater PREE, attributable to both decreased extinction in the CR condition and increased extinction in the PR condition. The results of these experiments on human PREE provide no support that PREE is a valid paradigm with which to explore the cognitive dysfunction underlying schizophrenia. PMID- 12110468 TI - Entrainment of locomotor activity rhythm in pinealectomized adult Syrian hamsters by daily melatonin infusion. AB - Melatonin entrains circadian rhythms in several species of rodents, but a role for melatonin as a Zeitgeber in the adult Syrian hamster is debated. The aim of this study was to define the conditions of daily programmed melatonin infusion in which an entrainment of the locomotor activity rhythm is obtained in adult male Syrian hamsters. The animals were pinealectomized, cannulated with a subcutaneous infusion system and submitted to dim red light conditions. They were initially daily infused with vehicle until free-running was established. Then, the animals were divided into three experimental groups, each group corresponding to a specific melatonin dose and infusion duration: (1) 10 microg melatonin/h for 5 h; (2) 30 microg melatonin/h for 5 h; and (3) 50 microg melatonin/h for 1 h. Of the total 64 hamsters, 37 hamsters fully entrained to the melatonin infusion regardless of whether the animals expressed during pre-treatment a free-running period (tau)< or >24 h, 20 animals presented a transient entrainment and seven did not entrain. Of the 37 animals entrained, withdrawal of melatonin re established free-running rhythms, although often with a different tau compared with that observed during pre-treatment. These results indicate that after a long time of daily infusion, melatonin is able to entrain the free-running rhythm in adult Syrian hamster. The mechanism involved is not known, but the change in tau observed after melatonin treatment in some animals suggests that melatonin, directly or indirectly, affects the functioning of the clock. PMID- 12110469 TI - Altered performance characteristics in cognitive tasks: comparison of the albino ICR and CD1 mouse strains. AB - With the advent of recombinant DNA technology the mouse has become a favored model organism in brain research. Numerous mouse strains are available to use as a host for carrying genetic alteration induced by targeted or random mutagenesis. Most strains differ in their genetic makeup and phenotypical characteristics. The choice of the host strain thus can be crucial for the analysis of functional effects of the induced mutation. In the present paper we analyze the behavior of two related outbred albino strains of mice, ICR and CD1, that are often used in transgenic research. Using two frequently employed learning tasks, the Morris water maze (MWM) and the context-dependent fear conditioning (CFC) as well as other behavioral tests, we demonstrate significant performance differences between the strains. ICR suffers from a severe visual impairment making this strain difficult to use in several behavioral paradigms that require good visual perception, e.g. the MWM. CD1 does not suffer from grossly impaired vision but, similarly to the ICR strain, CD1 mice exhibit decreased freezing in all phases of CFC. Although the strains are able to learn, such deficits can render them significantly impaired dependent on the performance demands of the cognitive test employed. Our findings underscore the need for careful examination of the characteristics of the host strain, the choice of which must be made in accordance with the expected functional alterations induced by the mutation. PMID- 12110470 TI - Anxiolytic effect of NKP608, a NK1-receptor antagonist, in the social investigation test in gerbils. AB - NKP608 is a potent, selective and orally active non-peptidic neurokinin-1 (NK1) receptor antagonist for which anxiolytic- and antidepressant-like effects have been described in various animal models in rats. Since species differences have been reported for some NK1-receptor antagonists, NKP608 was tested here in the social investigation test in gerbils, in a species in which the NK1-receptor is close to the human receptor. NKP608 (0.01 to> or =0.3 mg/kg p.o.) increased the time investigating the partner comparable to that seen following treatment with chlordiazepoxid (7 mg/kg p.o.), thus clearly indicating that NKP608 also has a robust anxiolytic effect in the social investigation test in gerbils. Such findings are in line with previous data obtained in rats, extend them to gerbils and corroborate the potential of NKP608 (and other representatives of the class of NK1-receptor antagonists) as new therapeutic agents beneficial in psychiatric disorders such as anxiety and/or depression. PMID- 12110472 TI - The hardness of food plays an important role in food selection behavior in rats. AB - To examine the importance of the hardness of foods, we conducted behavioral and electromyographical experiments in Wistar male rats using three kinds of pellets, a hard commercially made pellet (MF), a hard privately produced pellet (H) and a soft privately produced pellet (S). MF and H had the same hardness but contained different ingredients, and S and H had the same ingredients but different degrees of hardness (S/=1 x 10(9) cells/l, except for day 29 when the WBC had to be >/=3 x 10(9) cells/l. Six patients were treated at each dose level. The dose of paclitaxel was increased by 10 mg/m(2) per level. Of the 24 patients enrolled, 13 had adenocarcinoma, 10 had squamous cell carcinoma and one had an undifferentiated carcinoma. All patients were evaluable for toxicity and 22 of 24 patients were evaluable for response. The paclitaxel dose could be escalated to 110 mg/m(2). At this dose, 3 out of 6 patients developed dose-limiting toxicity (DLT) including neutropenic enterocolitis with sepsis, vomiting and diarrhoea. Diarrhoea grades 3 and 4 was seen in 4 (17%) patients. Two of these patients died of neutropenic enterocolitis. Neutropenia grades 3 or 4 was seen in 20 (83%) patients, but apart from the two patients with neutropenic enterocolitis no other infectious complications were seen. Mild to moderate sensory neurotoxicity was seen in 11 (46%) patients (grade 1 in 8 patients and grade 2 in 3 patients). Other toxicities were mild and easily manageable. Of the 22 evaluable patients, 11 (50%) patients achieved a partial or complete response with a median duration of 13 months. Ten patients with either locally advanced disease or supraclavicular or celiac lymph nodes received additional local treatment after response to chemotherapy, seven patients are still without evidence of disease after a median follow-up of 32 months. Paclitaxel at a dose 100 mg/m(2) infused over 3 h followed by a 3-h infusion of 70 mg/m(2) cisplatin can be recommended for further studies in patients with metastatic or unresectable oesophageal cancer. Occurring diarrhoea should be handled with caution because it may be a sign of neutropenic enterocolitis. The response rate of this dose-dense schedule seems encouraging. PMID- 12110497 TI - Prognostic factors for survival and factors associated with long-term remission in patients with advanced melanoma receiving cytokine-based treatments: second analysis of a randomised EORTC Melanoma Group trial comparing interferon-alpha2a (IFNalpha) and interleukin 2 (IL-2) with or without cisplatin. AB - The aim of this study was to define prognostic factors for survival, and especially for long-term survival in a mature data-set of patients with stage IV melanoma treated within a randomised trial of cytokine-based protocols. Long-term follow-up data on patients enrolled into a European Organization for Research and Treatment of Cancer (EORTC) trial comparing interferon-alpha (IFNalpha) plus interleukin-2 (IL-2) with or without cisplatin were collected. Univariate and multivariate Cox regression analyses were performed to define prognostic factors for survival. The characteristics of patients alive at 2 and 5 years after randomisation were compared with the entire cohort using the chi(2) test. The minimum potential follow-up of the 131 evaluable patients was 5 years. 18 patients (14%) were alive 2 years after randomisation, and 11 (8%) 5 years after randomisation. Pretreatment performance status (PS), serum lactate dehydrogenase (LDH) and tumour mass were significant predictors for survival, whereas site of metastases and number of sites were non-significant. PS and LDH were the only independent prognostic factors. All except 1 patient alive at 2 and 5 years had a pretreatment PS of 100%, and only three long-term survivors had elevated pretreatment LDH. There was no association between the site of metastases and long-term survival. Response to treatment was a major predictor for long-term survival, whereas addition of cisplatin did not impact upon overall survival probability or on long-term survival. The probability of long-term survival in stage IV melanoma patients after IL-2-based treatments is governed by pretreatment PS, serum LDH and response to treatment. Site of metastases, the basis for the M-subcategories of the new AJCC staging system, was not informative in this study. PMID- 12110498 TI - Lack of interstitial chromosome 1p deletions in clinically-detected neuroblastoma. AB - Loss of heterozygosity (LOH) of the distal part of the short arm of chromosome 1 in neuroblastoma is a well characterised phenomenon. In addition, previous reports have described interstitial deletions outside the common region of loss on chromosome 1p36, suggesting additional tumour suppressor loci. In this study, we have searched extensively for interstitial 1p deletions in a panel of 67 neuroblastoma samples from clinically-detected cases. We used three VNTR probes and 10 dinucleotide markers from the 1p32-36 regions reported to show interstitial deletions. Fifteen (22%) tumours showed telomeric LOH without evidence for more proximal interstitial deletions. Forty-five tumours showed no LOH or allelic imbalance. Seven (10%) tumours demonstrated allelic imbalance for one or more markers. These tumours were subsequently analysed by fluorescent in situ hybridisation (FISH) and flow cytometry. The patterns found in all seven tumours were consistent with copy number changes of the entire chromosome 1, without evidence for interstitial deletions. This study indicates that interstitial deletions of chromosome 1p are rare in clinically-detected neuroblastoma when analysed by a combination of molecular and cytogenetic techniques. PMID- 12110499 TI - Lung cancer risks in women with previous breast cancer. AB - Evaluation of the adverse effects of breast cancer treatment is becoming increasingly important in light of the earlier detection and prolonged survival of the patients. The beneficial effect of post-surgical radiotherapy has lately been challenged. The Swedish Cancer Registry (SCR) was used to identify approximately 141000 women with breast cancer, diagnosed between 1958 and 1997, followed-up for the occurrence of lung cancer. Standardised incidence ratios and expected number of lung cancers were calculated using incidence rates from the SCR. There were 613 subsequent lung cancers and a statistically significant increased risk of lung cancer was seen >5 years after breast cancer diagnosis, in contrast to a significantly decreased risk the first five years after the breast cancer diagnosis. The latter finding was confined to those >60 years of age when diagnosed with breast cancer. When restricting the analyses to those cases with information on the laterality of breast and lung cancer, an increased risk of a lung cancer on the same side as the breast cancer was seen >10 years after the breast cancer diagnosis. Birth cohorts with a higher smoking prevalence, i.e. 1930-1949, revealed a higher risk of lung cancer, than previous birth cohorts. Women with breast cancer have a significantly increased risk of developing a subsequent lung cancer possibly related to an interaction between radiotherapy and smoking. PMID- 12110500 TI - Antiproliferative activity induced by the somatostatin analogue, TT-232, in human pancreatic cancer cells. AB - Somatostatin analogues have been developed as antiproliferative agents, but their administration as general antitumour agents is limited, mainly because of the wide distribution of somatostatin receptors throughout the human body. TT-232, a new somatostatin structural analogue, was reported to have tumour-selective antiproliferative activity without an antisecretory action. We examined whether TT-232 had antiproliferative activity in human pancreatic cancer cell lines, and compared its antiproliferative activity with that of RC-160 and other TT-232 derivatives. TT-232 inhibited the growth of all of the cell lines used in this study and induced apoptotic cell death. RC-160 showed no such growth inhibition. TT-232 also inhibited tumour formation in a xenograft model. A competitive binding assay was performed using the cell membrane fraction and 111In-DTPA-TT 232 in order to show the existence of a specific binding site on the cells. A specific binding site was detected in MIAPaCa-2 cells. It has been shown that the activation of protein tyrosine phosphatase (PTPase) is one of the main intracellular pathways responsible for somatostatinergic inhibition of cell growth. We found a significant PTPase stimulation after TT-232 administration using an immunoblot analysis assessing the level of protein tyrosine phosphorylation, and also a direct measurement of the PTPase activity. We also demonstrated that PTPase stimulation by TT-232 was involved in its antiproliferative activity as this activity was reversed by the addition of sodium orthovanadate, a PTPase inhibitor. Our results indicate that TT-232 could be a potentially useful therapeutic agent if these data are translated into clinical practice. PMID- 12110501 TI - Expression of the vascular endothelial cell protein C receptor in epithelial tumour cells. AB - The rat monoclonal antibody LMR-42 has previously been shown to react with an external epitope of a plasma membrane protein with a M(r) of approximately 55,000 that was upregulated in multidrug-resistant (MDR) tumour cells. Here, we report the isolation of the cDNA encoding the LMR-42 antigen from the MDR human fibrosarcoma cell line HT1080/DR4 and the lung cancer cell line GLC4/ADR by expression cloning. Sequence analysis showed that the LMR-42 antigen is identical to the endothelial cell protein C receptor (EPCR). Using the LMR-42 Mab for cytochemical analyses of a disease-oriented panel of 45 non-drug selected tumour cell lines of the National Cancer Institute (NCI), we found high EPCR expression in 47% of the primary tumour cell lines, including melanomas, renal- and colon carcinomas. In a small panel of human tumours, occasionally very high EPCR expression was detected in endothelial vessels, but expression in the tumour cells was a rare event. The functional significance of overexpression of EPCR on both primary and drug-selected tumour cells is still unclear. As the protein is related to MHC class I molecules and shares no characteristics with any of the currently known transporter proteins, EPCR is not expected to play a causal role in the resistant phenotype of the MDR tumour cells. Nevertheless, exposure of tumour cells to cytostatic drugs may frequently lead to EPCR overexpression. Since EPCR is known to play a pivotal role in preventing blood coagulation through binding of (activated) protein C, it might endow tumour cells, both of mesenchymal and epithelial derivations, with increased growth potential by local anti-coagulant activity. PMID- 12110502 TI - DNA interstrand cross-linking and TP53 status as determinants of tumour cell sensitivity in vitro to the antibody-directed enzyme prodrug therapy ZD2767. AB - Cellular determinants of sensitivity to the bifunctional alkylating agent 4-[N,N bis(2-iodoethyl)amino]phenol (ZD2767D), the active drug produced by ZD2767 antibody-directed enzyme prodrug therapy (ADEPT), were studied. The prodrug 4 [N,N-bis(2-iodoethyl)amino]phenoxycarbonyl L-glutamic acid (ZD2767P)+activating enzyme carboxypeptidase G2 (CPG2) displayed growth inhibitory activity (IC(50) 0.04-2.2 microM) in colorectal tumour and non-small cell lung cancer (NSCLC) cell lines, and was more potent than a monofunctional ZD2767D analogue (colorectal cell lines-IC(50) 18-38 microM), synthesized for the first time. ZD2767P + CPG2 rapidly formed DNA-DNA interstrand cross-links (maximal at 10 min), and semi quantitative analyses indicate that levels were similar in 3 of 4 cell lines studied (25-75 rad equivalents) at equitoxic (10 x IC(50)/LC(50)) concentrations. In matched HCT116 TP53 functional/non-functional cell lines, there was no significant difference in the sensitivity to ZD2767P+CPG2. Together, these results suggest that cellular sensitivity to ZD2767P+CPG2 is, in part, related to the levels of interstrand crosslinks, but that TP53 status does not markedly effect chemosensitivity. PMID- 12110503 TI - The promise of translational physiology. PMID- 12110504 TI - Plasminogen activator inhibitor-1 and the kidney. AB - Plasminogen activator inhibitor-1 (PAI-1) is a serine protease inhibitor that was isolated 20 years ago. First recognized as an inhibitor of intravascular fibrinolysis, it is now evident that PAI-1 is a multifunctional protein with actions that may be dependent on or independent of its protease inhibitory effects. The latter often involve interactions between PAI-1 and vitronectin or the urokinase receptor. The protease-inhibitory actions of PAI-1 extend beyond fibrinolysis and include extracellular matrix turnover and activation of several proenzymes and latent growth factors. PAI-1 has been implicated in several renal pathogenetic processes, including thrombotic microangiopathies and proliferative and/or crescentic glomerulopathies. Most recently, it has become clear that PAI-1 also plays a pivotal role in progressive renal disease, both glomerulosclerosis and tubulointerstitial fibrosis. An active area of present research interest, untold stories are likely to be uncovered soon. PMID- 12110505 TI - Abnormal regulation of ENaC: syndromes of salt retention and salt wasting by the collecting duct. AB - Although the aldosterone-responsive segments of the nephron together reabsorb <10% of the filtered Na+, certain single-gene defects that affect the epithelial Na+ channel (ENaC) in the luminal membrane of the collecting duct (CD) or its regulation by aldosterone cause severe hypertension, whereas others cause salt wasting and hypotension. These rare defects illustrate the key role of the distal nephron in maintaining normal extracellular volume and blood pressure. Genetic defects that increase the Cl- conductance of the junctional complexes may also lead to salt retention and hypertension. Less dramatic alterations in regulatory actions of other hormones such as vasopressin (VP), either alone or with other genetic variations, diet, or environmental factors, may also produce Na+ retention or loss. Although VP acts primarily to regulate water balance, it is also an antinatriuretic hormone. Elevated basal plasma VP levels, and/or augmented VP release with increased Na+ intake, have been linked to essential hypertension in humans and in animal models of congestive heart failure and cirrhosis. Norepinephrine, dopamine, and prostaglandin E2 can inhibit the antinatriuretic effects of VP, and changes in the actions of these autocrine and paracrine regulators may also be involved in abnormal regulation of Na+ reabsorption. PMID- 12110506 TI - Rapid microplate method for PAH estimation. AB - Evaluation of renal hemodynamics requires estimation of effective renal plasma flow, which is commonly measured by the renal clearance of p-aminohippuric acid (PAH). There are many existing methods for PAH assay that are complicated, expensive, or time consuming. We describe a rapid, precise, and accurate microplate-based assay of PAH using p-dimethylaminocinnamaldehyde, which produces a red color on reaction with PAH, and compare it with a reference HPLC method. Renal PAH clearances were measured in 10 volunteers, and clearances were calculated by using the new and HPLC methods. There was excellent agreement between the HPLC and the microplate method of PAH assay. The average ratio of microplate to HPLC method was nearly 1.0, and the limits of agreement (2 SD) for plasma, urine, and clearances were 17.2, 19.3, and 25.5%, respectively. Intraday coefficient of variation for urine and plasma was <7%; interday coefficient of variation was <6% for urine and plasma samples. The microplate method is a reliable alternative to a reference HPLC method and can be performed for a fraction of the cost, time, and reagents. PMID- 12110507 TI - Bladder permeability barrier: recovery from selective injury of surface epithelial cells. AB - The mammalian bladder maintains high electrochemical gradients between urine and blood, permitting the kidney to modify body chemistries through urinary excretion. To perform this function, the urothelium maintains a tight permeability barrier. When this barrier is damaged, leakage of urine components into the underlying bladder layers results, with symptoms of cystitis. In these studies, we develop a model of selective urothelial injury using protamine sulfate (PS) and define the process by which this epithelium recovers from damage. Exposure to PS (10 mg/ml), but not vehicle, caused a rapid fall in transepithelial resistance as well as striking increases in water and urea permeabilities. These changes were accompanied by necrosis and sloughing of sheets of umbrella cells, as seen by scanning and transmission electron microscopy. Over the 72 h after PS exposure, barrier function recovered, with transepithelial resistance and water and urea permeabilities returning to normal values. After loss of umbrella cells, the underlying intermediate cells underwent rapid maturation, as evidenced by increased expression of uroplakins and gradual formation of well-defined tight junctions. At day 5 after PS exposure, barrier function was restored and the surface cells exhibited normal-appearing tight junctions and normal labeling for uroplakins and zonula occludens 1. However, the cells remained smaller than umbrella cells until day 10 after exposure, when normal size was restored. These studies develop for the first time a controlled model of selective urothelial damage and demonstrate a characteristic process by which barrier function is restored and underlying intermediate cells develop into mature umbrella cells. This model will be useful in defining the mechanisms that regulate repair of urothelial damage. PMID- 12110508 TI - Effect of immunosuppressive agents on glucocorticoid receptor function in A6 cells. AB - Immunosuppressive agents such as FK-506 and rapamycin inhibit aldosterone- stimulated Na+ transport in A6 cells. Concentration dependence is consistent with the known affinities of these agents for immunophilins. The inhibition was also dependent on time, requiring preincubation with FK-506 or rapamycin before inhibition was seen. The present studies were designed to determine whether this inhibition was pretranscriptional and whether it was due to an effect on either receptor translocation or nuclear accumulation. Because transport effects of steroids in A6 cells are mediated by glucocorticoid receptors (GRs), we examined the transcriptional response of GR-regulated reporters transfected into these cells. Preincubation of cells with FK-506 and rapamycin completely blocked reporter gene activation, whereas preincubation with cyclosporin A partially inhibited this activation. A minimum of 8 h of preincubation was required before the effect was seen. Using a transiently transfected green fluorescent protein-GR construct, we examined the effect of FK-506 and rapamycin on GR translocation. GR translocation induced by dexamethasone was extremely rapid (<5 min) and was largely unaffected by FK-506 or rapamycin but was completely blocked by geldanamycin. Digital deconvolutions revealed a punctate nuclear accumulation of GR, which was still seen after preincubation with immunosuppressive agents. These agents clearly inhibit steroid action by blocking GR-stimulated gene transcription, but this effect is not mediated by altered translocation or nuclear accumulation of receptors. Inhibition of steroid-regulated gene transcription by immunosuppressive agents may explain the electrolyte abnormalities seen in patients receiving these drugs. PMID- 12110509 TI - Cell proliferation is insufficient, but loss of tuberin is necessary, for chemically induced nephrocarcinogenicity. AB - Although 2,3,5-tris-(glutathion-S-yl)hydroquinone (TGHQ; 2.5 micromol/kg ip) markedly increased cell proliferation within the outer stripe of the outer medulla (OSOM) of the kidney in both wild-type (Tsc2(+/+)) and mutant Eker rats (Tsc2(EK/+)), only TGHQ-treated Tsc2(EK/+) rats developed renal tumors, indicating that cell proliferation per se was not sufficient for tumor development. Tuberin expression was initially induced within the OSOM after TGHQ treatment but was lost within TGHQ-induced renal tumors. High extracellular signal-regulated kinase (ERK) activity occurred in the OSOM of Tsc2(EK/+) rats at 4 mo and in TGHQ-induced renal tumors. Cyclin D1 was also highly expressed in TGHQ-induced renal tumors. Reexpression of Tsc2 in tuberin-negative cells decreased ERK activity, consistent with the growth-suppressive effects of this tumor suppressor gene. Thus 1) stimulation of cell proliferation after toxicant insult is insufficient for tumor formation; 2) tuberin induction after acute tissue injury suggests that Tsc2 is an acute-phase response gene, limiting the proliferative response after injury; and 3) loss of Tsc2 gene function is associated with cell cycle deregulation. PMID- 12110510 TI - LPA is a novel lipid regulator of mesangial cell hexokinase activity and HKII isoform expression. AB - The prototypical extracellular phospholipid mediator, lysophosphatidic acid (LPA), exhibits growth factor-like properties and represents an important survival factor in serum. This potent mesangial cell mitogen is increased in conditions associated with glomerular injury. It is also a known activator of the classic mitogen-activated protein kinase (MAPK) pathway, which plays an important role in the regulation of mesangial cell hexokinase (HK) activity. To better understand the mechanisms coupling metabolism to injury, we examined the ability of LPA to regulate HK activity and expression in cultured murine mesangial cells. LPA increased total HK activity in a concentration- and time-dependent manner, with maximal increases of >50% observed within 12 h of exposure to LPA concentrations > or =25 microM (apparent ED(50) 2 microM). These effects were associated with increased extracellular signal-regulated kinase (ERK) activity and were prevented by the pharmacological inhibition of either MAPK/ERK kinase or protein kinase C (PKC). Increased HK activity was also associated with increased glucose (Glc) utilization and lactate accumulation, as well as selectively increased HKII isoform abundance. The ability of exogenous LPA to increase HK activity was both Ca2+ independent and pertussis toxin insensitive and was mimicked by LPA-generating phospholipase A2. We conclude that LPA constitutes a novel lipid regulator of mesangial cell HK activity and Glc metabolism. This regulation requires sequential activation of both Ca2+-independent PKC and the classic MAPK pathway and culminates in increased HKII abundance. These previously unrecognized metabolic consequences of LPA stimulation have both physiological and pathophysiological implications. They also suggest a novel mechanism whereby metabolism may be coupled to cellular injury via extracellular lipid mediators. PMID- 12110511 TI - Water transport in neonatal and adult rabbit proximal tubules. AB - We have recently demonstrated that although the osmotic water permeability (P(f)) of neonatal proximal tubules is higher than that of adult tubules, the P(f) of brush-border and basolateral membrane vesicles from neonatal rabbits is lower than that of adults. The present study examined developmental changes in the water transport characteristics of proximal convoluted tubules (PCTs) in neonatal (9-16 days old) and adult rabbits to determine whether the intracellular compartment or paracellular pathway is responsible for the maturational difference in transepithelial water transport. The permeability of n-butanol was higher in the neonatal PCT than the adult PCT at all temperatures examined, whereas the diffusional water permeability was identical. Increasing the osmotic gradient increased volume absorption in both the neonatal and the adult PCT to the same degree. The P(f) was not different between the neonatal and the adult PCT at any osmotic gradient studied. To assess solvent drag as a measure of the paracellular transport of water, the effect of the osmotic gradient on mannitol and chloride transport were measured. There was no change in chloride or mannitol transport with the increased osmotic gradient in either group, indicating that there was no detectable paracellular water movement. In addition, the mannitol permeability of the neonatal PCT was found to be lower than that of the adult PCT with the isotonic bath (8.97 +/- 4.01 vs. 40.49 +/- 13.89 microm/s, P < 0.05). Thus the intracellular compartment of the neonatal PCT has a lower resistance for water transport than the adult PCT and is responsible for the higher than expected P(f) in the neonatal PCT. PMID- 12110512 TI - Glucose stimulates O2 consumption, NOS, and Na/H exchange in diabetic rat proximal tubules. AB - Endothelial nitric oxide synthase (NOS) and neuronal NOS protein increased in proximal tubules of acidotic diabetic rats 3-5 wk after streptozotocin injection. NOS activity (citrulline production) was similar in nondiabetic and diabetic tubules incubated with low glucose (5 mM glucose + 20 mM mannitol); but after 30 min with high glucose (25 mM), Ca-sensitive citrulline production had increased 23% in diabetic tubules. Glucose concentration did not influence citrulline production in nondiabetic tubules. High glucose increased carboxy-2-phenyl 4,4,5,5,-tetramethylimidazoline 1-oxyl-3-oxide (cpt10)-scavenged NO sevenfold in a suspension of diabetic tubules but did not alter NO in nondiabetic tubules. Diabetes increased ouabain-sensitive 86Rb uptake (141 +/- 9 vs. 122 +/- 6 nmol x min(-1) x mg(-1)) and oligomycin-sensitive O2 consumption (QO2; 16.0 +/- 1.7 vs. 11.3 +/- 0.7 nmol x min(-1) x mg(-1)). Ethylisopropyl amiloride-inhibitable QO2 (6.5 +/- 0.6 vs. 2.4 +/- 0.3 nmol x min(-1) x mg(-1)) accounted for increased oligomycin-sensitive QO2 in diabetic tubules. N(G)-monomethyl-L-arginine methyl ester (L-NAME) inhibited most of the increase in 86Rb uptake and QO2 in diabetic tubules. L-NAME had little effect on nondiabetic tubules. Inhibition of QO2 by ethylisopropyl amiloride and L-NAME was only 5-8% additive. Uncontrolled diabetes for 3-5 wk increases NOS protein in proximal tubules and makes NOS activity sensitive to glucose concentration. Under these conditions, NO stimulates Na-K ATPase and QO2 in proximal tubules. PMID- 12110513 TI - Role of prostanoids in regulation of the renin-angiotensin-aldosterone system by salt intake. AB - We investigated the effect of cyclooxygenase (COX) activity on the regulation of the renin-angiotensin-aldosterone system by salt intake. Therefore, Sprague Dawley rats were subjected to different salt diets [0.02, 0.6, and 8% NaCl (wt/wt)] and treated with the selective COX-2 inhibitor rofecoxib (10 mg x kg body wt(-1) x day(-1)) or with ketorolac at a dose selective for COX-1 inhibition (2 mg x kg body wt(-1) x day(-1)) for 3, 7, 14, and 21 days. Rofecoxib and ketorolac caused a similar reduction of renocortical PGE2 formation with a low salt diet. Rofecoxib did not change plasma renin activity or renocortical renin mRNA abundance with any of the diets but clearly lowered plasma aldosterone concentration. In contrast, ketorolac delayed the increase in plasma renin activity and of renin mRNA in response to low salt intake but did not change plasma aldosterone concentration. Prolonged treatment with rofecoxib but not with ketorolac caused an upregulation of COX-2 expression while COX-1 mRNA abundance remained unchanged. These findings suggest that COX-1-derived, but not COX-2 derived, prostanoids are of relevance for the regulation of the renin system by salt intake. PMID- 12110514 TI - Proliferation and osmotic tolerance of renal inner medullary epithelial cells in vivo and in cell culture. AB - Renal inner medullary (IM) cells survive interstitial osmolality that ranges from 600 to 1,700 mosmol/kgH2O or more. In contrast, much smaller acute changes killed the cells previously studied in tissue culture, such as mouse IM collecting duct 3 (mIMCD3) cells, that are immortalized with SV40 and proliferate rapidly. Proliferation and DNA replication sensitize mIMCD3 cells to hypertonicity. In the present studies, we observed that proliferating cells were scarce in rat IM. Then, we prepared passage 2 mouse IM epithelial (p2mIME) cells. They have a much lower incidence of DNA replication than do mIMCD3 cells. p2mIME cells survive much greater acute increases in NaCl than do mIMCD3 cells and also tolerate significantly greater acute increases of urea and of NaCl plus urea, but still not to levels as high as occur in vivo. We conclude that immortalization and continued DNA replication account for part of the previously observed difference in osmotic tolerance between IM cells in vivo and in cell culture but that other factors must also be involved. PMID- 12110515 TI - Water permeability of aquaporin-4 is decreased by protein kinase C and dopamine. AB - Aquaporin-4 (AQP4) plays an important role in the basolateral movement of water in the collecting duct. Here we show that this water channel can be dynamically regulated. Water permeability (P(f)) was measured in individual LLC-PK1 cells that were transiently transfected with AQP4. To identify which cells were transfected, AQP4 was tagged at the NH2 terminus with green fluorescent protein. Transfected cells showed a strong fluorescent signal in basolateral membrane and a low-to-negligible signal in the cytosol and apical membrane. Activation of protein kinase C (PKC) with phorbol 12,13-dibutyrate (PDBu) significantly decreased P(f) of cells expressing AQP4 but had no effect on neighboring untransfected cells. No redistribution of AQP4 in response to PDBu was detected. Dopamine also decreased the P(f) in transfected cells. The effect was abolished by the PKC inhibitor Ro 31-8220. Reduction of AQP4 water permeability by PDBu and dopamine was abolished by point mutation of Ser(180), a consensus site for PKC phosphorylation. We conclude that PKC and dopamine decrease AQP4 water permeability via phosphorylation at Ser180 and that the effect is likely mediated by gating of the channel. PMID- 12110516 TI - The electrical resistance breakdown assay determines the role of proteinases in tumor cell invasion. AB - The electrical resistance breakdown of the Madin-Darby canine kidney (MDCK) cell monolayer provides a continuous assay system for cancer invasion that detects functional changes before morphological alterations. In this study, we address the question of whether physical contact between tumor cell and epithelial monolayer is a prerequisite for tumor cell invasion. When human melanoma cells were seeded directly (i.e., physical contact) on top of an electrically tight epithelial cell layer (5,800 +/- 106 Omega x cm2), electrical monolayer leakage led to an 18 +/- 3% reduction of transepithelial electrical resistance within 24 h. However, when melanoma cells were seeded close to the basolateral surface of the epithelial cell monolayer but separated by a filter membrane (i.e., no physical contact), electrical leakage occurred even more quickly (42 +/- 3% reduction in 24 h). Atomic force microscopy detected discrete structural changes between cells. Electrical leakage was effectively blocked by alpha2-macroglobulin or ilomastat, inhibitors of matrix metalloproteinases. We conclude that exocytosis of soluble proteases causes electrical breakdown of the MDCK monolayer, independently of physical contact between tumor cells and the monolayer. PMID- 12110517 TI - Effects of fatty acids on mitochondrial beta-oxidation enzyme gene expression in renal cell lines. AB - Regulatory effects of fatty acids on gene expression of medium-chain acyl-CoA dehydrogenase (MCAD), a mitochondrial beta-oxidation enzyme, were investigated in rabbit kidney cell lines derived from proximal tubule (RC.SV1), thick ascending limb of Henle's loop (RC.SV2), or collecting duct (RC.SV3). Exposure to long chain fatty acids led to significant increases (2-fold) in MCAD mRNA abundance in RC.SV1 and RC.SV2 cells; kinetics and dose-response studies established that maximal MCAD gene stimulation was reached 4 h after addition of 50 microM oleate (C18:1) in the culture medium. These effects of fatty acids were totally abolished in the presence of 1 microg/ml actinomycin D, a transcription inhibitor. Staining of cellular lipids revealed that fatty acid-induced gene stimulation could occur in the absence of cellular fatty acid accumulation. Altogether, these data indicate that small changes in cellular fatty acid flux can have direct short-term effects on fatty acid oxidation enzyme gene expression in renal cells, and this might take part in the regulation of cellular fatty acid homeostasis in response to changes in tubular fluid composition. PMID- 12110518 TI - Stimulation of protein kinase C pathway mediates endocytosis of human nongastric H+-K+-ATPase, ATP1AL1. AB - The human nongastric H+-K+-ATPase, ATP1AL1, shown to reabsorb K+ in exchange for H+ or Na+, is localized in the luminal plasma membrane of renal epithelial cells. It is presumed that renal H+-K+-ATPases can be regulated by endocytosis. However, little is known about the molecular mechanisms that control plasma membrane expression of renal H+-K+-ATPases. In our study, activation of protein kinase C (PKC) using phorbol esters (phorbol 12-myristate 13-acetate) leads to clathrin dependent internalization and intracellular accumulation of the ion pump in stably transfected Madin-Darby canine kidney cells. Functional inactivation of the H+-K+-ATPase by PKC activation is shown by intracellular pH measurements. Proton extrusion capacity of ATP1AL1-transfected cells is drastically reduced after phorbol 12-myristate 13-acetate incubation and can be prevented with the PKC blocker bisindolylmaleimide. Ion pump internalization and inactivation are specifically mediated by the PKC pathway, whereas activation of the protein kinase A pathway has no influence. Our results show that the nongastric H+-K+ ATPase is a specific target for the PKC pathway. Therefore, PKC-mediated phosphorylation is a potential regulatory mechanism for apical nongastric H+-K+ ATPase plasma membrane expression. PMID- 12110519 TI - Specificity of human and rat orthologs of the concentrative nucleoside transporter, SPNT. AB - To understand the roles that nucleoside transporters play in the in vivo distribution of clinically important nucleoside analogs, the substrate specificity of each transporter isoform should be determined. In the present work, we studied the substrate specificities of the human and rat orthologs of the Na+-dependent purine-selective nucleoside transporter (SPNT; concentrative nucleoside transporter 2), for nucleosides, nucleobases, and base- and ribose modified nucleoside analogs. The two-electrode voltage-clamp technique in Xenopus laevis oocytes expressing these transporters was used. Purine nucleosides and uridine induced currents in oocytes expressing rat SPNT (rSPNT) or human SPNT1 (hSPNT1). The rank order of magnitude of nucleoside-induced currents was guanosine > uridine > adenosine > inosine and guanosine > uridine > inosine > adenosine for rSPNT- and hSPNT1-expressing oocytes, respectively. Uridine analogs (modified at the 5-position of the base) induced little or no current, suggesting that these compounds are only poorly transported by either transporter. Cladribine induced currents in oocytes expressing rSPNT (K(0.5) = 57 +/- 12 microM) but not hSPNT1. The ribose-modified nucleoside analogs, adenine arabinoside, and 2',3'-dideoxyadenosine induced currents in rSPNT-expressing, but not in hSPNT1-expressing, oocytes. These data suggest that there are notable species differences in the specificity of SPNT for synthetic nucleoside analogs. PMID- 12110520 TI - Higher basal serine phosphorylation of D1A receptors in proximal tubules of old Fischer 344 rats. AB - Dopamine (DA) and D1-like receptor agonists promote an increase in Na excretion by means of activation of the D1-like receptor signaling cascade and subsequent inhibition of the Na/H exchanger and Na-K-ATPase in renal proximal tubules. Recently, our laboratory reported that DA and the D1-like receptor agonist failed to inhibit Na-K-ATPase activity in old Fischer 344 rats because of uncoupling of D1A receptors from G proteins and that this abnormality led to a diminished natriuretic response to DA in old Fischer 344 rats. In this study, we have tested the hypothesis that the mechanism of this uncoupling may be an altered phosphorylation of D1A receptors in old rats. In experiments performed in renal cortical slices, both DA and SKF-38393, a D1-like receptor agonist, increased the serine phosphorylation of D1A receptors in adult (6 mo) but not old (24 mo) rats. Interestingly, the basal serine phosphorylation of D1A receptors was higher in old than in adult rats. Competition ligand binding ([3H]SCH-23390) experiments on the D1-like receptor in adult and old rats with fenoldopam, a D1-like receptor agonist, revealed the presence of two affinity states of the receptors. There was a rightward shift in the agonist displacement of the ligand in old compared with adult rats, as reflected in the IC50 values (adult vs. old, 7.46 x 10(-9) +/- 2.26 vs. 7.93 x 10(-7) +/- 1.33 M). Also, there was a reduction in agonist affinity in the low-affinity receptors in old compared with adult rats (IC50, adult vs. old, 5.67 x 10(-5) +/- 1.33 vs. 12.60 x 10(-5) +/- 6.50 M). Moreover, the abundance of D1A receptor proteins was approximately 47% lower in the membranes of old compared with adult rats. We speculate that higher basal serine phosphorylation of D1A receptors may have rendered the D1A receptor uncoupled from G protein, leading to a reduced agonist affinity state and thus diminished natriuretic response to DA in old rats. PMID- 12110522 TI - The promise of translational physiology. PMID- 12110523 TI - Glandular regulation of interstitial diffusion: a model and simulation of a novel physiological mechanism. AB - In endocrine glands, vigorous and coordinated responses are often elicited by modest changes in the concentration of the agonist molecule. The mammalian parathyroid gland is a representative case. Small (5%) changes in serum calcium result in 10-fold (1,000%) changes in glandular parathyroid hormone (PTH) release. In vitro, single isolated cells are observed to secrete fewer hormones than cells residing within a connected group, suggesting that a network has emergent regulatory properties. In PTH-secreting tumors, however, the ability to respond quickly to changes in calcium is strongly damped. A unifying hypothesis that accounts for these phenomena is realized by extracellular modulation of calcium diffusivity. A theoretical model and computational experiments demonstrate qualitative agreement with published experimental results. Our results suggest that, in addition to the cellular mechanisms, endocrine glandular networks may have regulatory prowess at the level of interstitial transport. PMID- 12110521 TI - Glucocorticoid regulation of the murine PHEX gene. AB - The phosphate-regulating gene with homologies to endopeptidases on the X chromosome (PHEX) is a member of the neutral endopeptidase family, which is expressed predominantly on the plasma membranes of mature osteoblasts and osteocytes. Although it is known that the loss of PHEX function results in X linked hypophosphatemic rickets, characterized by abnormal bone matrix mineralization and renal phosphate wasting, little is known about how PHEX is regulated. We therefore sought to determine whether the murine PHEX gene is regulated by glucocorticoids (GCs), which are known to influence phosphate homeostasis and bone metabolism. Northern blot analysis revealed increased PHEX mRNA expression in GC-treated suckling mice (1.5-fold) and in rat osteogenic sarcoma (UMR-106) cells (2.5-fold). An increase was also seen in PHEX promoter activity in transiently transfected UMR-106 cells with GC treatment. Analysis of nested promoter deletions revealed that an atypical GC response element was located between -337 and -315 bp. Mutational analysis and electrophoretic mobility shift assays further identified -326 to -321 bp as a site involved in GC regulation. Supershift analyses and electrophoretic mobility shift assay competition studies indicated that the core binding factor alpha1-subunit transcription factor is able to bind to this region and may therefore play a role in the GC response of the murine PHEX gene. PMID- 12110524 TI - K(ATP) channels and insulin secretion disorders. AB - ATP-sensitive potassium (K(ATP)) channels are inhibited by intracellular ATP and activated by ADP. Nutrient oxidation in beta-cells leads to a rise in [ATP]-to [ADP] ratios, which in turn leads to reduced K(ATP) channel activity, depolarization, voltage-dependent Ca(2+) channel activation, Ca(2+) entry, and exocytosis. Persistent hyperinsulinemic hypoglycemia of infancy (HI) is a genetic disorder characterized by dysregulated insulin secretion and, although rare, causes severe mental retardation and epilepsy if left untreated. The last five or six years have seen rapid advance in understanding the molecular basis of K(ATP) channel activity and the molecular genetics of HI. In the majority of cases for which a genotype has been uncovered, causal HI mutations are found in one or the other of the two genes, SUR1 and Kir6.2, that encode the K(ATP) channel. This article will review studies that have defined the link between channel activity and defective insulin release and will consider implications for future understanding of the mechanisms of control of insulin secretion in normal and diseased states. PMID- 12110525 TI - Genetic approaches to the molecular understanding of type 2 diabetes. AB - The appreciation that individual susceptibility to type 2 diabetes (T2D) and related components of the dysmetabolic syndrome has a strong inherited component provides a coherent framework within which to develop a molecular understanding of the pathogenesis of T2D. This review focuses on the main approaches currently adopted by researchers seeking to identify the inherited basis of T2D and the present state of our knowledge. One central theme that emerges is that progress in defining the genetic basis of the common, multifactorial forms of T2D is hindered by etiological heterogeneity: T2D is likely to represent the final common pathway of diverse interacting primary disturbances. Such heterogeneity equally compromises efforts to understand the basis for T2D by use of other approaches, such as cellular biochemistry and classical physiology. Analyses that seek to ally sophisticated physiological characterization with measures of genomic variation are likely to provide powerful tools for redressing the loss of power associated with such heterogeneity. PMID- 12110526 TI - Metabolic changes in the glucose-induced apoptotic blastocyst suggest alterations in mitochondrial physiology. AB - Mammalian preimplantation embryos experience a critical switch from an oxidative to a predominantly glycolytic metabolism. In this study, the change in nutrient metabolism between the 2-cell and blastocyst stages was followed by measuring single embryo concentrations of tricarboxylic acid (TCA) cycle and glycolytic metabolites with microfluorometric enzymatic cycling assays. When the normal values were established, further changes that occur as a result of the induction of apoptosis by exposure to high-glucose conditions were examined. From the 2 cell to the blastocyst stage, the embryos experienced an increase in TCA metabolites and a dramatic increase in fructose 1,6-bisphosphate (FBP). The high TCA metabolites may result from accumulation of substrate due to a slowing of TCA cycle metabolism as glycolysis predominates. Embryos exposed to elevated glucose conditions experienced significantly lower FBP, suggesting decreased glycolysis, significantly higher pyruvate, suggesting increased pyruvate uptake by the embryos in response to decreased glycolysis, and increased TCA metabolites, suggesting an inability to oxidize the pyruvate and a slowing of the TCA cycle. We speculate that the glycolytic changes lead to dysfunction of the outer mitochondrial membrane that results in the abnormal TCA metabolite pattern and triggers the apoptotic event. PMID- 12110527 TI - Lactate induces insulin resistance in skeletal muscle by suppressing glycolysis and impairing insulin signaling. AB - Elevation of plasma lactate levels induces peripheral insulin resistance, but the underlying mechanisms are unclear. We examined whether lactate infusion in rats suppresses glycolysis preceding insulin resistance and whether lactate-induced insulin resistance is accompanied by altered insulin signaling and/or insulin stimulated glucose transport in skeletal muscle. Hyperinsulinemic euglycemic clamps were conducted for 6 h in conscious, overnight-fasted rats with or without lactate infusion (120 micromol x kg(-1) x min(-1)) during the final 3.5 h. Lactate infusion increased plasma lactate levels about fourfold. The elevation of plasma lactate had rapid effects to suppress insulin-stimulated glycolysis, which clearly preceded its effect to decrease insulin-stimulated glucose uptake. Both submaximal and maximal insulin-stimulated glucose transport decreased 25-30% (P < 0.05) in soleus but not in epitrochlearis muscles of lactate-infused rats. Lactate infusion did not alter insulin's ability to phosphorylate the insulin receptor, the insulin receptor substrate (IRS)-1, or IRS-2 but decreased insulin's ability to stimulate IRS-1- and IRS-2-associated phosphatidylinositol 3 kinase activities and Akt/protein kinase B activity by 47, 75, and 55%, respectively (P < 0.05 for all). In conclusion, elevation of plasma lactate suppressed glycolysis before its effect on insulin-stimulated glucose uptake, consistent with the hypothesis that suppression of glucose metabolism could precede and cause insulin resistance. In addition, lactate-induced insulin resistance was associated with impaired insulin signaling and decreased insulin stimulated glucose transport in skeletal muscle. PMID- 12110528 TI - Enhanced O-GlcNAc protein modification is associated with insulin resistance in GLUT1-overexpressing muscles. AB - O-linked glycosylation on Ser/Thr with single N-acetylglucosamine (O GlcNAcylation) is a reversible modification of many cytosolic/nuclear proteins, regulated in part by UDP-GlcNAc levels. Transgenic (T) mice that overexpress GLUT1 in muscle show increased basal muscle glucose transport that is resistant to insulin stimulation. Muscle UDP-GlcNAc levels are increased. To assess whether GLUT4 is a substrate for O-GlcNAcylation, we translated GLUT4 mRNA (mutated at the N-glycosylation site) in rabbit reticulocyte lysates supplemented with [(35)S]methionine. O-GlcNAcylated proteins were galactosylated and separated by lectin affinity chromatography; >20% of the translated GLUT4 appeared to be O GlcNAcylated. To assess whether GLUT4 or GLUT4-associated proteins were O GlcNAcylated in muscles, muscle membranes were prepared from T and control (C) mice labeled with UDP-[(3)H]galactose and immunoprecipitated with anti-GLUT4 IgG (or nonimmune serum), and N-glycosyl side chains were removed enzymatically. Upon SDS-PAGE, several bands showed consistently two- to threefold increased labeling in T vs. C. Separating galactosylated products by lectin chromatography similarly revealed approximately threefold more O-GlcNAc-modified proteins in T vs. C muscle membranes. RL-2 immunoblots confirmed these results. In conclusion, chronically increased glucose flux, which raises UDP-GlcNAc in muscle, results in enhanced O-GlcNAcylation of membrane proteins in vivo. These may include GLUT4 and/or GLUT4-associated proteins and may contribute to insulin resistance in this model. PMID- 12110529 TI - Regulation of IGF-I mRNA by GH: putative functions for class 1 and 2 message. AB - This study investigated mechanisms regulating hepatic insulin-like growth factor (IGF)-I class 1 and 2 mRNA levels. Lambs were treated with growth hormone (GH) either as an acute, single dose or over a longer term. Total hepatic unspliced, pre-mRNA levels increased after the single dose of GH but were attenuated after 8 days of GH, with exon 1- and 2-derived pre-mRNA levels displaying coordinate responses. Surprisingly, changes in total spliced, mature mRNA levels did not reflect those for pre-mRNA, instead being augmented after 8 days of GH. GH also induced a differential increase in the ratio of mature class 2-to-class 1 IGF-I mRNA; therefore, this must be predominantly via posttranscriptional mechanisms. Increases in the ratio of class 2-to-class 1 mRNA were observed in polysomal vs. total RNA preparations derived from GH-treated but not control lambs, indicating an increased proportion of class 2 transcripts engaged in translation. Our findings indicate that GH may stabilize mature class 2 transcripts or destabilize mature class 1 transcripts and that class 2 mRNA may have a greater translational potential. The following two main functions of hepatic class 2 IGF-I mRNA are suggested: an efficient "monitor" of GH status via providing a rapid negative feedback mechanism and a coordinator of endocrine-regulated tissue growth. PMID- 12110530 TI - Effect of enteral vs. parenteral glucose delivery on initial splanchnic glucose uptake in nondiabetic humans. AB - To determine if enteral delivery of glucose influences splanchnic glucose metabolism, 10 subjects were studied when glucose was either infused into the duodenum at a rate of 22 micromol x kg(-1) x min(-1) and supplemental glucose given intravenously or when all glucose was infused intravenously while saline was infused intraduodenally. Hormone secretion was inhibited with somatostatin, and glucose (approximately 8.5 mmol/l) and insulin (approximately 450 pmol/l) were maintained at constant but elevated levels. Intravenously infused [6,6 (2)H(2)]glucose was used to trace the systemic appearance of intraduodenally infused [3-(3)H]glucose, whereas UDP-glucose flux (an index of hepatic glycogen synthesis) was measured using the acetaminophen glucuronide method. Despite differences in the route of glucose delivery, glucose production (3.5 +/- 1.0 vs. 3.3 +/- 1.0 micromol x kg(-1) x min(-1)) and glucose disappearance (78.9 +/- 5.7 vs. 85.0 +/- 7.2 micromol x kg(-1) x min(-1)) were comparable on intraduodenal and intravenous study days. Initial splanchnic glucose extraction (17.5 +/- 4.4 vs. 14.5 +/- 2.9%) and hepatic UDP-glucose flux (9.0 +/- 2.0 vs. 10.3 +/- 1.5 micromol x kg(-1) x min(-1)) also did not differ on the intraduodenal and intravenous study days. These data argue against the existence of an "enteric" factor that directly (i.e., independently of circulating hormone concentrations) enhances splanchnic glucose uptake or hepatic glycogen synthesis in nondiabetic humans. PMID- 12110531 TI - Changes in LPLa and reverse cholesterol transport variables during 24-h postexercise period. AB - We investigated the time course of exercise-induced lipoprotein lipase activity (LPLa) and reverse cholesterol transport (RCT) during the 24-h postexercise period. Subjects were 10 sedentary normolipidemic males [NTG; fasting triglyceride (TG) = 89.1 +/- 8.6 mg/dl] and 6 hyperlipidemic males (HTG; fasting TG = 296.8 +/- 64.0 mg/dl). Each subject performed a control trial (no exercise) and 4 exercise trials. In the exercise trials, a subject jogged on a treadmill at 60% of his maximal O(2) consumption for 1 h. Pre- and postheparin blood samples were taken before exercise (baseline) and at 4, 8, 12, and 24 h after exercise. There was no group difference in LPLa (P > 0.05) over the time points. When the LPLa data from the two groups were combined, LPLa at 24 h after exercise was higher than baseline or at 4, 8, 12 h after exercise (P < 0.05). Plasma TG and lecithin-cholesterol acyltransferase activity (LCATa) were higher in HTG than in NTG, and the total high-density lipoprotein-cholesterol (HDL(tot)-Chol) was lower in HTG than in NTG (P < 0.05). HDL(2)-Chol, LCATa, and cholesterol ester transfer protein activity did not differ during the 24-h postexercise period (P > 0.05). These results suggest that LPLa is still increasing 24 h after an acute aerobic exercise and that the magnitude of the increase in exercise-induced LPLa in HTG was similar to that in NTG. Furthermore, in the sedentary population with or without HTG, the variables related to RCT do not change during the 24-h period after exercise. PMID- 12110532 TI - Regulation of endogenous glucose production after a mixed meal in type 2 diabetes. AB - The extent and time course of suppression of endogenous glucose production (EGP) in type 2 diabetes after a mixed meal have been determined using a new tracer methodology. Groups of age-, sex-, and weight-matched normal controls (n = 8) and diet-controlled type 2 diabetic subjects (n = 8) were studied after ingesting a standard mixed meal (550 kcal; 67% carbohydrate, 19% fat, 14% protein). There was an early insulin increment in both groups such that, by 20 min, plasma insulin levels were 266 +/- 54 and 190 +/- 53 pmol/l, respectively. EGP was similar basally [2.55 +/- 0.12 mg x kg(-1) x min(-1) in control subjects vs. 2.92 +/- 0.16 mg x kg(-1) x min(-1) in the patients (P = 0.09)]. After glucose ingestion, EGP declined rapidly in both groups to approximately 50% of basal within 30 min of the meal. Despite the initial rapid decrease, the EGP was significantly greater in the diabetic group at 60 min (1.75 +/- 0.12 vs. 1.05 +/- 0.14 mg x kg( 1) x min(-1); P < 0.01) and did not reach nadir until 210 min (0.96 +/- 0.17 mg x kg(-1) x min(-1)). Between 60 and 240 min, EGP was 47% higher in the diabetic group (0.89 +/- 0.09 vs. 1.31 +/- 0.13 mg x kg(-1) x min(-1), P < 0.02). These data quantitate the initial rapid suppression of EGP after a mixed meal in type 2 diabetes and the contribution of continuing excess glucose production to subsequent hyperglycemia. PMID- 12110533 TI - Interrelationships of serum testosterone and free testosterone index with FFM and strength in aging men. AB - Muscle mass and strength losses during aging may be associated with declining levels of serum testosterone (T) in men. Few studies have shown a direct relationship between T and muscle mass and strength. Subjects were 262 men, aged 24-90 yr, from the Baltimore Longitudinal Study of Aging, who had T and sex hormone-binding globulin sex hormone-binding globulin (SHBG) measurements, from which the free T index (FTI) was calculated (T/SHBG) from serum samples collected longitudinally since 1963, total body fat mass and arm and leg fat-free mass (FFM) by dual-energy X-ray absorptiometry and arm and leg strength by dynanomometry. Mixed-effects models estimated T and FTI at the time of mass and strength measurements. Age, total body fat, arm and leg FFM, T, and FTI were significantly associated with concentric and eccentric strength. FTI, not T, was modestly, but directly, related to arm and leg strength after fat, arm and leg FFM, height, and age were accounted for and indirectly through body mass. FTI is a better predictor of arm and leg strength than T in aging men. PMID- 12110534 TI - No apparent suppression by insulin of in vivo skeletal muscle lipolysis in nonobese women. AB - To investigate the antilipolytic effect of insulin in skeletal muscle and adipose tissue in vivo, the rates of glycerol release from the two tissues were compared in 10 nonobese women during a two-step euglycemic hyperinsulinemic clamp. Tissue interstitial glycerol levels were determined by microdialysis, and tissue blood flow was assessed with the (133)Xe clearance technique. Absolute rates of glycerol release were estimated according to Fick's principle. In both adipose tissue and muscle, glycerol levels decreased significantly already during the low insulin infusion rate. The fractional release of glycerol (difference between interstitial glycerol and arterialized venous plasma glycerol) was reduced by more than one-half in adipose tissue (P < 0.0001) in response to insulin, whereas it remained unaltered in skeletal muscle. Muscle blood flow rates increased by 60% (P < 0.02) during insulin infusion; in adipose tissue, blood flow rates did not change significantly in response to insulin. The basal rate of glycerol release from skeletal muscle amounted to approximately 15% of that from adipose tissue. After insulin infusion, the rate of adipose tissue glycerol release was markedly suppressed, whereas in skeletal muscle the rate of glycerol mobilization did not change significantly in response to insulin. It is concluded that insulin does not inhibit the rate of lipolysis in skeletal muscle of nonobese women. PMID- 12110535 TI - Differences between mouse and rat pancreatic islets: succinate responsiveness, malic enzyme, and anaplerosis. AB - Succinic acid methyl esters are potent insulin secretagogues in rat pancreatic islets, but they do not stimulate insulin release in mouse islets. Unlike rat and human islets, mouse islets lack malic enzyme and, therefore, are unable to form pyruvate from succinate-derived malate for net synthesis of acetyl-CoA. Dimethyl [2,3-(14)C]succinate is metabolized in the citric acid cycle in mouse islets to the same extent as in rat islets, indicating that endogenous acetyl-CoA condenses with oxaloacetate derived from succinate. However, without malic enzyme, the net synthesis from succinate of the citric acid cycle intermediates citrate, isocitrate, and alpha-ketoglutarate cannot occur. Glucose and other nutrients that augment alpha-ketoglutarate formation are secretagogues in mouse islets with potencies similar to those in rat islets. All cycle intermediates can be net synthesized from alpha-ketoglutarate. Rotenone, an inhibitor of site I of the electron transport chain, inhibits methyl succinate-induced insulin release in rat islets even though succinate oxidation forms ATP at sites II and III of the respiratory chain. Thus generating ATP, NADH, and anaplerosis of succinyl-CoA plus the four-carbon dicarboxylic acids of the cycle and its metabolism in the citric acid cycle is insufficient for a fuel to be insulinotropic; it must additionally promote anaplerosis of alpha-ketoglutarate or two intermediates interconvertible with alpha-ketoglutarate, citrate, and isocitrate. PMID- 12110536 TI - Somatostatin-28 regulates GLP-1 secretion via somatostatin receptor subtype 5 in rat intestinal cultures. AB - Five somatostatin receptors (SSTRs) bind somatostatin-14 (S-14) and somatostatin 28 (S-28), but SSTR5 has the highest affinity for S-28. To determine whether S-28 acting through SSTR5 mediates inhibition of glucagon-like peptide-1 (GLP-1), fetal rat intestinal cell cultures were treated with somatostatin analogs with relatively high specificity for SSTRs 2-5. S-28 dose-dependently inhibited GLP-1 secretion stimulated by gastrin-releasing peptide more potently than S-14 (EC(50) 0.01 vs. 5.8 nM). GLP-1 secretion was inhibited by an SSTR5 analog, BIM-23268, more potently than S-14 and nearly as effectively as S-28. The SSTR5 analog L 372,588 also suppressed GLP-1 secretion equivalent to S-28, but a structurally similar peptide, L-362,855 (Tyr to Phe at position 7), was ineffective. An SSTR2 selective analog was less effective than S-28, and an SSTR3 analog was inactive. Separate treatment with GLP-1-(7-36)-NH(2) increased S-28 and S-14 secretion by three- and fivefold; BIM-23268 abolished S-28 without altering S-14, whereas the SSTR2 analog was inactive. The results indicate that somatostatin regulation of GLP-1 secretion occurs via S-28 through activation of SSTR5. GLP-1-stimulated S 28 secretion is also autoregulated by SSTR5 activation, suggesting a feedback loop between GLP-1 and S-28 modulated by SSTR5. PMID- 12110537 TI - Alterations of nPKC distribution, but normal Akt/PKB activation in denervated rat soleus muscle. AB - Denervation has been shown to impair the ability of insulin to stimulate glycogen synthesis and, to a lesser extent, glucose transport in rat skeletal muscle. Insulin binding to its receptor, activation of the receptor tyrosine kinase and phosphatidylinositol 3'-kinase do not appear to be involved. On the other hand, it has been shown that denervation causes an increase in the total diacylglycerol (DAG) content and membrane-associated protein kinase C (PKC) activity. In this study, we further characterize these changes in PKC and assess other possible signaling abnormalities that might be related to the decrease of glycogen synthesis. The results reveal that PKC-epsilon and -theta;, but not -alpha or zeta, are increased in the membrane fraction 24 h after denervation and that the timing of these changes parallels the impaired ability of insulin to stimulate glycogen synthesis. At 24 h, these changes were associated with a 65% decrease in glycogen synthase (GS) activity ratio and decreased electrophoretic mobility, indicative of phosphorylation in GS in muscles incubated in the absence of insulin. Incubation of the denervated soleus with insulin for 30 min minimally increased glucose incorporation into glycogen; however, it increased GS activity threefold, to a value still less than that of control muscle, and it eliminated the gel shift. In addition, insulin increased the apparent abundance of GS kinase (GSK)-3 and protein phosphatase (PP)1 alpha in the supernatant fraction of muscle homogenate to control values, and it caused the same increases in GSK-3 and Akt/protein kinase B (PKB) phosphorylation and Akt/PKB activity that it did in nondenervated muscle. No alterations in hexokinase I or II activity were observed after denervation; however, in agreement with a previous report, glucose 6 phosphate levels were diminished in 24-h-denervated soleus, and they did not increase after insulin stimulation. These results indicate that alterations in the distribution of PKC-epsilon and -theta; accompany the impairment of glycogen synthesis in the 24-h-denervated soleus. They also indicate that the basal rate of glycogen synthesis and its stimulation by insulin in these muscles are diminished despite a normal activation of Akt/PKB and phosphorylation of GSK-3. The significance of the observed alterations to GSK-3 and PP1 alpha distribution remain to be determined. PMID- 12110538 TI - RXR receptor agonist suppression of thyroid function: central effects in the absence of thyroid hormone receptor. AB - High-affinity agonists for the retinoic acid X receptors (RXR) have pleotropic effects when administered to humans. These include induction of hypertriglyceridemia and hypothyroidism. We determined the effect of a novel high affinity RXR agonist with potent antihyperglycemic effects on thyroid function of female Zucker diabetic rats and nondiabetic littermates and in db/db mice. In both nondiabetic and ZFF rats, AGN194204 causes a 70-80% decrease in thyrotropin (TSH), 3,3',5-triiodothyronine, and thyroxine (T(4)) concentrations. In the db/db mouse, AGN194204 causes a time-dependent decrease in thyroid hormone levels with the fall in TSH that was significant after 1 day of treatment preceding the fall in T(4) levels that was significant at 3 days of treatment. Treatment with AGN194204 caused an initial increase in hepatic 5'-deiodinase mRNA levels which then fell to undetectable levels by 3 days of treatment and continued to be low at 7 days of treatment. After treatment for 5 days with AGN194204, both wild-type and thyroid hormone receptor beta (TR beta(-/-))-deficient mice demonstrated a nearly 50% decrease in serum TSH and T(4) concentrations. The results suggest that a high-affinity RXR agonist with antihyperglycemic activity can cause central hypothyroidism independently of TR beta, the main mediator of hormone induced TSH suppression. PMID- 12110539 TI - Metabolic basis of HIV-lipodystrophy syndrome. AB - Human immunodeficiency virus (HIV)-lipodystrophy syndrome (HLS) is characterized by hypertriglyceridemia, low high-density lipoprotein-cholesterol, lipoatrophy, and central adiposity. We investigated fasting lipid metabolism in six men with HLS and six non-HIV-infected controls. Compared with controls, HLS patients had lower fat mass (15.9 +/- 1.3 vs. 22.3 +/- 1.7 kg, P < 0.05) but higher plasma glycerol rate of appearance (R(a)), an index of total lipolysis (964.71 +/- 103.33 vs. 611.08 +/- 63.38 micromol x kg fat(-1) x h(-1), P < 0.05), R(a) palmitate, an index of net lipolysis (731.49 +/- 72.36 vs. 419.72 +/- 33.78 micromol x kg fat(-1) x h(-1), P < 0.01), R(a) free fatty acids (2,094.74 +/- 182.18 vs. 1,470.87 +/- 202.80 micromol x kg fat(-1) x h(-1), P < 0.05), and rates of intra-adipocyte (799.40 +/- 157.69 vs. 362.36 +/- 74.87 micromol x kg fat(-1) x h(-1), P < 0.01) and intrahepatic fatty acid reesterification (1,352.08 +/- 123.90 vs. 955.56 +/- 124.09 micromol x kg fat(-1) x h(-1), P < 0.05). Resting energy expenditure was increased in HLS patients (30.51 +/- 2.53 vs. 25.34 +/- 1.04 kcal x kg lean body mass(-1) x day(-1), P < 0.05), associated with increased non-plasma-derived fatty acid oxidation (139.04 +/- 24.17 vs. 47.87 +/- 18.81 micromol x kg lean body mass(-1) x min(-1), P < 0.02). The lipoatrophy observed in HIV lipodystrophy is associated with accelerated lipolysis. Increased hepatic reesterification promotes the hypertriglyceridemia observed in this syndrome. PMID- 12110540 TI - Role of PKC isoforms in glucose transport in 3T3-L1 adipocytes: insignificance of atypical PKC. AB - To elucidate the involvement of protein kinase C (PKC) isoforms in insulin induced and phorbol ester-induced glucose transport, we expressed several PKC isoforms, conventional PKC-alpha, novel PKC-delta, and atypical PKC isoforms of PKC-lambda and PKC-zeta, and their mutants in 3T3-L1 adipocytes using an adenovirus-mediated gene transduction system. Endogenous expression and the activities of PKC-alpha and PKC-lambda/zeta, but not of PKC-delta, were detected in 3T3-L1 adipocytes. Overexpression of each wild-type PKC isoform induced a large amount of PKC activity in 3T3-L1 adipocytes. Phorbol 12-myristrate 13 acetate (PMA) activated PKC-alpha and exogenous PKC-delta but not atypical PKC lambda/zeta. Insulin also activated the overexpressed PKC-delta but not PKC alpha. Expression of the wild-type PKC-alpha or PKC-delta resulted in significant increases in glucose transport activity in the basal and PMA-stimulated states. Dominant-negative PKC-alpha expression, which inhibited the PMA activation of PKC alpha, decreased in PMA-stimulated glucose transport. Glucose transport activity in the insulin-stimulated state was increased by the expression of PKC-delta but not of PKC-alpha. These findings demonstrate that both conventional and novel PKC isoforms are involved in PMA-stimulated glucose transport and that other novel PKC isoforms could participate in PMA-stimulated and insulin-stimulated glucose transport. Atypical PKC-lambda/zeta was not significantly activated by insulin, and expression of the wild-type, constitutively active, and dominant-negative mutants of atypical PKC did not affect either basal or insulin-stimulated glucose transport. Thus atypical PKC enzymes do not play a major role in insulin stimulated glucose transport in 3T3-L1 adipocytes. PMID- 12110541 TI - Free fatty acid-induced peripheral insulin resistance augments splanchnic glucose uptake in healthy humans. AB - To investigate the effect of elevated plasma free fatty acid (FFA) concentrations on splanchnic glucose uptake (SGU), we measured SGU in nine healthy subjects (age, 44 +/- 4 yr; body mass index, 27.4 +/- 1.2 kg/m(2); fasting plasma glucose, 5.2 +/- 0.1 mmol/l) during an Intralipid-heparin (LIP) infusion and during a saline (Sal) infusion. SGU was estimated by the oral glucose load (OGL)-insulin clamp method: subjects received a 7-h euglycemic insulin (100 mU x m(-2) x min( 1)) clamp, and a 75-g OGL was ingested 3 h after the insulin clamp was started. After glucose ingestion, the steady-state glucose infusion rate (GIR) during the insulin clamp was decreased to maintain euglycemia. SGU was calculated by subtracting the integrated decrease in GIR during the period after glucose ingestion from the ingested glucose load. [3-(3)H]glucose was infused during the initial 3 h of the insulin clamp to determine rates of endogenous glucose production (EGP) and glucose disappearance (R(d)). During the 3-h euglycemic insulin clamp before glucose ingestion, R(d) was decreased (8.8 +/- 0.5 vs. 7.6 +/- 0.5 mg x kg(-1) x min(-1), P < 0.01), and suppression of EGP was impaired (0.2 +/- 0.04 vs. 0.07 +/- 0.03 mg x kg(-1) x min(-1), P < 0.01). During the 4-h period after glucose ingestion, SGU was significantly increased during the LIP vs. Sal infusion study (30 +/- 2 vs. 20 +/- 2%, P < 0.005). In conclusion, an elevation in plasma FFA concentration impairs whole body glucose R(d) and insulin mediated suppression of EGP in healthy subjects but augments SGU. PMID- 12110542 TI - Salmon cardiac natriuretic peptide is a volume-regulating hormone. AB - The present study tested the hypothesis that salmon cardiac peptide (sCP), a new member of the family of natriuretic peptides, has an important role in the regulation of fluid balance and cardiovascular function. Intra-arterial administration of sCP increased urine output in salmon. It had a diuretic effect in rat as well, but the potency was lower. sCP increased the sodium excretion in proportion to the increased urine flow. Blood pressure was not affected by sCP in either species. Acute volume expansion elevated the plasma level of sCP in salmon, and an acute transfer of salmon from fresh to sea water decreased the circulating sCP level. Cardiac immunoreactive sCP or sCP mRNA levels were not affected by transfer to sea water. These results indicate that sCP has an important physiological role in defending salmon against volume overload but that it does not appear to contribute to the short-term regulation of blood pressure. sCP provides an excellent model of the general mechanisms of regulation of the A type (atrial) natriuretic peptide system. PMID- 12110543 TI - Compensatory glomerular growth after unilateral nephrectomy is VEGF dependent. AB - Various growth factors and cytokines have been implicated in different forms of kidney enlargement. Vascular endothelial growth factor (VEGF) is essential for normal renal development and plays a role in diabetic glomerular enlargement. To explore a possible role for VEGF in compensatory renal changes after uninephrectomy, we examined the effect of a neutralizing VEGF-antibody (VEGF-Ab) on glomerular volume and kidney weight in mice treated for 7 days. Serum and kidney insulin-like growth factor I (IGF-I) levels were measured, since IGF-I has been implicated in the pathogenesis of compensatory renal growth, and VEGF has been suggested to be a downstream mediator of IGF-I. Placebo-treated uninephrectomized mice displayed an early transient increase in kidney IGF-I concentration and an increase in glomerular volume and kidney weight. In VEGF-Ab treated uninephrectomized animals, increased glomerular volume was abolished, whereas renal hypertrophy was partially blocked. Furthermore, the renal effects of VEGF-Ab administration were seen without affecting the renal IGF-I levels. In conclusion, these results demonstrate that compensatory glomerular growth after uninephrectomy is VEGF dependent. PMID- 12110544 TI - Low-dose dexamethasone in the rat: a model to study insulin resistance. AB - The main aim of this study was to set up a new animal model to study insulin resistance. Wistar rats (6 or 7 per group) received the following for 4 wk in experiment 1: 1) vehicle, 2) 2 microg/day subcutaneous dexamethasone, 3) metformin (400 mg x kg(-1) x day(-1) os), and 4) dexamethasone plus metformin. In experiment 2 the rats received the following: 1) vehicle, 2) dexamethasone, 3) dexamethasone plus arginine (2%; as substrate of the nitric oxide synthase for nitric oxide production) in tap water, and 4) dexamethasone plus isosorbide dinitrate (70 mg/kg; as direct nitric oxide donor) in tap water. Insulin sensitivity was significantly reduced by dexamethasone already at week 1, before the increase in blood pressure (day 15) and without significant changes in body weight compared with vehicle. Dexamethasone-treated rats had significantly higher triglycerides, hematocrit, and insulin, whereas serum total nitrates/ nitrites were lower compared with vehicle. The concomitant treatment with metformin minimized all the described effects of dexamethasone. In experiment 2, only isosorbide dinitrate was able to prevent the observed dexamethasone-induced metabolic, hemodynamic, and insulin sensitivity changes. Chronic low-dose subcutaneous dexamethasone (2 microg/day) is a useful model to study the relationships between insulin resistance and blood pressure in the rat, and dexamethasone might decrease insulin sensitivity and increase blood pressure through an endothelium-mediated mechanism. PMID- 12110545 TI - GLUT4-containing vesicles are released from membranes by phospholipase D cleavage of a GPI anchor. AB - We have previously developed a cell-free assay from rat skeletal muscle that displayed in vitro glucose transporter 4 (GLUT4) transfer from large to small membrane structures by the addition of a cytosolic protein fraction. By combining protein fractionation and the in vitro GLUT4 transfer assay, we have purified a glycosylphosphatidylinositol (GPI) phospholipase D (PLD) that induces transfer of GLUT4 from small to large membranes. The in vitro GLUT4 transfer was activated and inhibited by suramin and 1,10-phenanthroline (an activator and an inhibitor of GPI-PLD activity, respectively). Furthermore, upon purification of the GLUT4 transporter protein, the protein displayed an elution profile in which the molecular mass was related to the charge, suggesting the presence or absence of phosphate. Second, by photoaffinity labeling of the purified GLUT4 with 3 (trifluoromethyl)-3-(m-[(125)I]iodopenyl)diazirine, both labeled phosphatidylethanolamine and fatty acids (constituents of a GPI link) were recovered. Third, by using phase transition of Triton X-114, the purified GLUT4 was found to be partly detergent resistant, which is a known characteristic of GPI-linked proteins. Fourth, the purified GLUT4 protein was recognized by an antibody raised specifically against GPI links. In conclusion, GLUT4-containing vesicles may be released from a membrane compartment by action of a GPI-PLD. PMID- 12110547 TI - New creatine transporter assay and identification of distinct creatine transporter isoforms in muscle. AB - Despite the pivotal role of creatine (Cr) and phosphocreatine (PCr) in muscle metabolism, relatively little is known about sarcolemmal creatine transport, creatine transporter (CRT) isoforms, and subcellular localization of the CRT proteins. To be able to quantify creatine transport across the sarcolemma, we have developed a new in vitro assay using rat sarcolemmal giant vesicles. The rat giant sarcolemmal vesicle assay reveals the presence of a specific high-affinity and saturable transport system for Cr in the sarcolemma (Michaelis-Menten constant 52.4 +/- 9.4 microM and maximal velocity value 17.3 +/- 3.1 pmol x min( 1) x mg vesicle protein(-1)), which cotransports Cr into skeletal muscle together with Na(+) and Cl(-) ions. The regulation of Cr transport in giant vesicles by substrates, analogs, and inhibitors, as well as by phorbol 12-myristate 13 acetate and insulin, was studied. Two antibodies raised against COOH- and NH(2) terminal synthetic peptides of CRT sequences both recognize two major polypeptides on Western blots with apparent molecular masses of 70 and 55 kDa, respectively. The highest CRT expression occurs in heart, brain, and kidney, and although creatine kinase is absent in liver cells, CRT is also found in this tissue. Surprisingly, immunofluorescence staining of cultured adult rat heart cardiomyocytes with specific anti-CRT antibodies, as well as cell fractionation and cell surface biotinylation studies, revealed that only a minor CRT species with an intermediate molecular mass of approximately 58 kDa is present in the sarcolemma, whereas the previously identified major CRT-related protein species of 70 and 55 kDa are specifically located in mitochondria. Our studies indicate that mitochondria may represent a major compartment of CRT localization, thus providing a new aspect to the current debate about the existence and whereabouts of intracellular Cr and PCr compartments that have been inferred from [(14)C]PCr/Cr measurements in vivo as well as from recent in vivo NMR studies. PMID- 12110546 TI - Temporal gradients in shear stimulate osteoblastic proliferation via ERK1/2 and retinoblastoma protein. AB - Bone cells are subject to interstitial fluid flow (IFF) driven by venous pressure and mechanical loading. Rapid dynamic changes in mechanical loading cause transient gradients in IFF. The effects of pulsatile flow (temporal gradients in fluid shear) on rat UMR106 cells and rat primary osteoblastic cells were studied. Pulsatile flow induced a 95% increase in S-phase UMR106 cells compared with static controls. In contrast, ramped steady flow stimulated only a 3% increase. Similar patterns of S-phase induction were also observed in rat primary osteoblastic cells. Pulsatile flow significantly increased relative UMR106 cell number by 37 and 62% at 1.5 and 24 h, respectively. Pulsatile flow also significantly increased extracellular signal-regulated kinase (ERK1/2) phosphorylation by 418%, whereas ramped steady flow reduced ERK1/2 activation to 17% of control. Correspondingly, retinoblastoma protein was significantly phosphorylated by pulsatile fluid flow. Inhibition of mitogen-activated protein (MAP)/ERK kinase (MEK)1/2 by U0126 (a specific MEK1/2 inhibitor) reduced shear induced ERK1/2 phosphorylation and cell proliferation. These findings suggest that temporal gradients in fluid shear stress are potent stimuli of bone cell proliferation. PMID- 12110548 TI - Reasons for poor understanding of when and how to access GP care for childhood asthma in Auckland, New Zealand. AB - BACKGROUND: Attempts to explain why some patients lack the understanding needed to access GP care for childhood asthma are uncommon and have tended to be based on reported statistical associations. OBJECTIVES: The aims of this study were to describe and account for poor patient understanding of when and how to access GP care for childhood asthma in Auckland, New Zealand. METHODS: A general inductive approach was used to analyse 29 semi-structured, personal interviews, during March-May 2001, with Auckland key informants selected through maximum variation sampling. Informant checking and the literature supported the text analysis by two independent researchers. RESULTS: Key informants reported wide variations in the extent to which guardians and asthmatic children understand when and how to access GP services. Two sets of barriers to patient understanding were identified. The first limits the willingness of people to seek understanding and the second limits their ability to understand, even if they want to understand. CONCLUSIONS: Use of qualitative methodology was able to reveal barriers to patient understanding. Strategies operating at the GP and system levels were identified to help overcome these barriers. PMID- 12110549 TI - Illness behaviour and patient satisfaction as correlates of self-referral in Japan. AB - BACKGROUND: Patients who frequently change physicians without letters of referral are common, and this has become a source of concern among primary care doctors in Japan. Previous studies have shown a correlation between psychiatric disorders and patient dissatisfaction and the utilization of medical resources. Abnormal illness behaviours such as hypochondria and inappropriate treatment seeking have been associated with various psychiatric disorders. The relationship between illness behaviour and self-referral in Japan has yet to be fully explored. OBJECTIVES: Our aim was to describe the characteristic illness behaviour and satisfaction level of self-referred patients in the general medicine clinic of Saga Medical School Hospital. METHODS: Using the Japanese version of the Illness Behaviour Questionnaire (J-IBQ), we examined the illness behaviour of 277 self referred patients visiting the clinic. Patient satisfaction with previous medical care was examined with the use of our original Patient Satisfaction Questionnaire. The results were compared with those for physician-referred patients. RESULTS: Self-referred patients differed significantly from original visit patients on the GH (general hypochondriasis), DC (disease conviction), AD (affect disturbance) and I (irritability) scales and from physician-referred patients on the GH and DC scales. In comparison with physician-referred patients, self-referred patients showed significant dissatisfaction with their most recent medical visit elsewhere. Dissatisfaction toward the medical staff, especially the doctors, was stronger than that toward the medical environment, waiting time or the on-site medical equipment. CONCLUSIONS: It is important to give patients appropriate overall support, not only physical but also emotional, when they first visit a general physician for medical advice. The J-IBQ may be a useful instrument for primary identification of self-referral patients with probable somatization syndromes. Open doctor-doctor and patient-doctor communication is necessary to increase patient satisfaction, which may be helpful to minimize the self-referral phenomenon in Japan. PMID- 12110550 TI - Meeting patient expectations of care: the major determinant of satisfaction with out-of-hours primary medical care? AB - BACKGROUND: Client or consumer expectation is considered to influence their satisfaction with the service provided, but its importance has not been quantified in health care. OBJECTIVE: The aim of this study was to determine the effect of "patient expectations of care" on satisfaction with care provided by out-of-hours services. METHODS: We surveyed 3457 patients who requested out-of hours care from five practices, two general practice out-of-hours co-operatives and a deputizing service in an English health authority during late 1997. The independent variables were: the service providing the care (service type), where out-of-hours care was given (location of care) and whether the care met the patient's expectations. The independent variable was overall patient satisfaction with out-of-hours care. RESULTS: Patients who received the care they hoped for (their idealized expectation was met) were more satisfied than those who did not. Patients who attended centres were more satisfied with the care received than those who had had home visits. Patients were more satisfied if they received care from the co-operative which did not employ assistants than from the deputizing service. Idealized expectation (care which was hoped for) match, location of care and service type explained 34, 2 and 4% of the variance, respectively. Age, sex, ethnicity, access to a car, normative/comparative expectation (care which was expected) and whether patients expected and received telephone advice, a home visit or domiciliary care, and the delay between request for care and care provision were not independently associated with satisfaction. CONCLUSIONS: Meeting or failing to meet the care patients hoped for is an important predictor of patient satisfaction with out-of-hours care. Purchasers and providers of out of-hours care should consider whether and how patient expectation of service can be managed. This may reduce patient dissatisfaction with the service they provide. These findings also have important implications for the design of studies which use patient satisfaction as an outcome variable. PMID- 12110552 TI - The role of patient, physician and systemic factors in the management of type 2 diabetes mellitus. AB - BACKGROUND: Few studies have explored the contextual dimensions and subsequent interactions that contribute to a lack of adherence in the application of guidelines for diabetes management. OBJECTIVE: The purpose of this qualitative study was to explore family physicians' issues and perceptions regarding the barriers to and facilitators of the management of patients with type 2 diabetes mellitus (DM). METHODS: Four focus groups composed of family physicians (n= 30) explored the participants' experiences in the management of patients with type 2 DM. A semi-structured interview guide began with questions on family physicians' experience of providing care and included specific probes to stimulate discussion about the various barriers to and facilitators of the management of type 2 DM in family practice. RESULTS: Participants clearly identified type 2 DM as a chronic disease most often managed by family physicians. The findings revealed distinct barriers and facilitators in managing patients with type 2 DM which fell into three domains: patient factors; physician factors; and systemic factors. There was a dynamic interplay among the three factors. The important role of education was common to each. CONCLUSIONS: The interactions of patient, physician and systemic factors have implications for the implementation of a diabetes management model. The care of patients with type 2 DM exemplifies the ongoing challenges of caring for patients with a chronic disease in family practice. The findings, while specific to the management of type 2 DM, have potential transferability to other chronic illnesses managed by family physicians. PMID- 12110551 TI - Patient and GP agreement on aspects of general practice care. AB - OBJECTIVE: The aim of the present study was to compare patient and GP priorities for general practice care. METHODS: A questionnaire survey was carried out in general practice in Denmark which included 900 consecutive patients aged over 18 years from 15 practices collected in 1995, and 919 randomly sampled GPs in 1999. The postal questionnaire, developed by the EUROPEP group, contained 40 questions about eight aspects of primary care. Participants were asked to state their priorities for each question ranging from "not at all important" to "most important". A reminder questionnaire was sent to non-responders after 2 weeks. Top priority percentages ("very/most important") were calculated for each question as were differences between participant groups. RESULTS: Questionnaires were answered by 771 (85.7%) patients and 584 (64.2%) GPs. Their priorities were highly correlated (r = 0.754, P < 0.001). Patients gave higher priority than GPs to availability and accessibility of the practice and seeing the same GP. The GP should be capable of providing information on illness, investigations and treatments and patient associations, and should know the patient's history and be regularly updated through courses. CONCLUSIONS: Patient and GP priorities for primary care were highly correlated. The higher priority awarded by patients than by GPs to specific aspects of primary care should be acknowledged when organizing and developing general practice. PMID- 12110553 TI - Severity of episodes of care assessed by family physicians and patients: the DUSOI/WONCA as an extension of the International Classification of Primary Care (ICPC). AB - OBJECTIVES: The DUSOI/WONCA is included in the second edition of the International Classification of Primary Care (ICPC-2), as an extension to assess the severity of episodes of care. We studied (i) family physician's (FPs') assessment of three DUSOI/WONCA parameters per episode of care; (ii) how these relate to patient and episode of care characteristics, and to the interventions that occur; and (iii) how FPs' and patients' assessment of severity compare. METHODS: Twelve FPs participated and coded patient and encounter data with ICPC. Also, they answered three DUSOI/WONCA questions, that were also answered (after the consultation) by 300 patients. Odds ratios were calculated for the relationships of the severity elements to patient and episode characteristics, and interventions. The relative agreement between FPs' and patients' ratings of severity was assessed. RESULTS: In 2033 consultations, 2860 episodes of care were documented, with a subset of 411 with a paired assessment by patient and FP. Patients appeared to be less hindered by symptoms/ complaints than the FPs thought, and less optimistic about the prognosis without care than the FP. Clear relationships existed between the FPs' assessment of severity and the patient, encounter and episode of care characteristics. Substantial agreement existed between FPs' and patients' assessment of severity. CONCLUSIONS: This study confirms the feasibility for FPs routinely to code the separate elements of severity for episodes of care, simultaneously using ICPC and DUSOI/WONCA. The studied elements of severity all provide relevant information: the interventions that occurred all related to them in a logical fashion. The FP-patient agreement on severity is satisfactory, also in the sense that it seems realistic to include these elements of severity as a topic in the communication with the patient. PMID- 12110554 TI - Predictors of differences in symptom perception of older patients and their doctors. AB - BACKGROUND: Doctors and their older patients do not necessarily agree on what health problem ought to be treated. Discordance influences diagnostic procedures, patient satisfaction and treatment outcome negatively. OBJECTIVE: The purpose of the present study was to determine the psychosocial factors influencing differences in symptom reports of patients and physicians. METHODS: A cohort study was carried out in a medical out-patient clinic. A total 141 women and 213 men agreed to give symptom reports while waiting for their doctor's appointment and allowed their treating physician to evaluate symptoms afterwards. RESULTS: Disagreement between patients and physicians on which symptoms triggered the visit appeared in one-third of the cases. This was more likely the longer symptoms existed, the less intensely patients experienced their symptom and the more restricted they felt because of the symptom. Psychosocial factors did not have a significant influence. CONCLUSIONS: Independently of psychosocial variables, the different illness concepts of patients and their treating physician influence the subject of the consultation. Doctors and patients were most likely to agree when patients reported their symptoms as being of recent onset and being intense. Symptom intensity and the associated degree of restriction seem to reflect two different conceptual dimensions of symptom evaluation. PMID- 12110555 TI - Genital infection by Chlamydia trachomatis in Lisbon: prevalence and risk markers. AB - BACKGROUND: There is little information about the prevalence and risk markers for Chlamydia trachomatis infections in Portugal. OBJECTIVES: Our aim was to assess the prevalence of C. trachomatis genital infection and to study variables associated with this infection in a group of sexually active women aged < or =30 years living in the Lisbon area and to estimate the prevalence of C. trachomatis infection among partners of infected patients. METHODS: A systematic sample of women observed in general practice family planning and teenager clinics was collected. A questionnaire was administered, followed by a pelvic examination. A first-catch urine sample was taken for polymerase chain reaction (PCR) Amplicor assay. When a sample tested positive, the woman was invited to obtain a urine sample from her partner. Socio-demograhic, behavioural and clinical variables were studied and their association with the PCR Amplicor result was assessed. RESULTS: A total of 1108 women, aged between 14 and 30 years, were studied. Fifty one women (4.6% of total sample) tested positive for C. trachomatis. The prevalence of infection was slightly higher in patients aged < or =19 years (5.3%) than in age groups 20-25 (4.8%) and 26-30 years (3.9%). African ethnicity was related to a higher percentage of infection than European ethnicity: 9.8% versus 3.8%, P= 0.0067. Use of condoms "sometimes/never" was associated with a higher prevalence of infection: 5.2% versus 2.3% in those responding "always/almost always" (P= 0.0447). An altered cervix was associated with a higher prevalence of infection: 7.3% versus 3.7% with a normal cervix (P= 0.0106). Urine samples were obtained from 16 partners of infected patients. Six partners (37.5%) tested positive for C. trachomatis. CONCLUSIONS: A 4.6% prevalence of C. trachomatis genital infection was found. African ethnicity, using condoms "sometimes/never" and an altered cervix were associated with C. trachomatis infection, but showed low positive predictive value for C. trachomatis infection. Younger age may be associated with a slight increase in risk. Contact tracing for diagnosis and treatment remains a difficult issue to approach effectively. PMID- 12110556 TI - Accuracy of a first diagnosis of asthma in primary health care. AB - BACKGROUND: In a postal questionnaire study, the prevalence of asthma in southern Sweden has been found to be 5.5%. However, the register prevalence of asthma obtained from the medical records in the same municipality and age groups was found to be only 2.1%. OBJECTIVES: The aims of the study were to investigate whether the low register prevalence of asthma was caused by an underdiagnosis of asthma in primary health care and to validate a first diagnosis of asthma set by GPs in primary health care. METHODS: During a period of 3 months in 1997, all patients seeking care in the primary health care units of the municipality of Lund (population 171 877) with upper or lower airway infections, prolonged cough, allergic rhinitis, fatigue or a first positive diagnosis of asthma were recorded ( n = 3025). RESULTS: In the whole group of 3025 patients, 99 patients were found to have received a diagnosis of asthma for the first time during the study period. The diagnosis was verified in 52 of those 68 patients who attended a follow-up and examination by a respiratory physician. Among the remaining 2926 patients, 221 patients were selected randomly to constitute a control group. In this group, three patients were found to have asthma. Thus, the specificity of an asthma diagnosis set in primary health care was 0.99 [95% confidence interval (CI) 0.99-1.00] and the sensitivity was 0.59 (95% CI 0.31-0.81). CONCLUSIONS: The GPs in this study were good at excluding those who did not have asthma (specificity 99%) but less good in correctly diagnosing those who actually had current asthma (sensitivity 59%), which suggests an underdiagnosis of asthma. PMID- 12110558 TI - Labelling of acute respiratory illness: evidence of between-practitioner variation in the UK. AB - BACKGROUND: It is unclear which symptoms and signs GPs use when attributing diagnostic labels to patients with acute respiratory illness (ARI). OBJECTIVE: We sought to ascertain GPs' self-reported definitions of ARI. METHODS: A postal questionnaire concerned with the diagnosis of ARI was sent to all registered GPs in Avon Health Authority. GPs were asked to choose a clinical term that would describe the clinical presentation in four hypothetical patients, and the next three questions asked them to define acute bronchitis, upper respiratory tract infection (URTI) and any other term they used for ARI (excluding pneumonia). We measured proportions and compared responses across the three diagnostic categories. RESULTS: The majority (88%) of GPs agreed that cough associated with fever should be labelled as a URTI. When sputum and chest signs were also present, opinion was more divided, with 62% diagnosing acute bronchitis in young patients and 72% lower respiratory tract infection in old patients. CONCLUSIONS: This study demonstrates that there is more consistent use of diagnostic labels for URTI than for acute bronchitis or other terms used to label ARI. In the future, researchers should quantify the prognostic significance of symptoms and signs in ARI and provide GPs with a more rational approach to the diagnosis and management of ARI. PMID- 12110557 TI - Striking the right balance in colorectal cancer care--a qualitative study of rural and urban patients. AB - BACKGROUND: Colorectal cancer is the second most common cause of cancer-related death in Scotland. For patients, the journey from diagnosis through treatment is complex and there are inequalities in survival rates. OBJECTIVES: The aim of the present study was to explore how patients with colorectal cancer perceive their care. METHODS: This was a qualitative study involving 95 patients and relatives of patients with colorectal cancer in the North, Northeast and Northern Isles, Scotland. Focus groups (32 participants) were conducted in hospital, and interviews (63 participants) in patients' own homes in order to explore their experiences of health services for cancer-what was good, what was bad and what was needed. Analysis was inductive, with exploration of similar and divergent perspectives within themes. RESULTS: Patients wanted rapid diagnosis, specialist treatment and good communication, but their experiences of and perspectives on these areas were often divergent. Delays in diagnosis could stem from late presentation by patients, but also from early presentation when the cancer could go undetected. GP continuity was desirable, but sometimes implicated in delays. Patients preferred their GPs to be advocates, not gatekeepers. The context, however, was one where some patients pursued their care tenaciously while others did not. For some, speed of progress through the system was everything, but others found this could be impersonal. Outlying patients had to balance transport difficulties with the benefits of distant specialist treatment. Some patients wanted full information to be provided directly, but others could not cope with this. CONCLUSIONS: From patients' perspectives, ideal cancer care cannot be achieved in a uniform way. For some of the key goals of cancer treatment to be met, including rapid access to specialist treatment for all and good communication of bad news and test results, a balance must be struck which tailors care to individuals. PMID- 12110559 TI - Health care follow-up after stroke: opportunities for secondary prevention. AB - BACKGROUND: Stroke patients have a 15-fold increased risk of a recurrence, but management of risk factors following stroke has been found to be unsatisfactory. Little is known about health service follow-up of patients after stroke or, consequently, the opportunities for providing secondary prevention to patients. OBJECTIVE: The aim of the present study was to investigate the relationship between health service follow-up and management of risk factors after stroke. METHODS: The study used data from the population-based South London Stroke Register, collected prospectively between 1995 and 1998. Main measures included risk factor change and follow-up by hospital physicians, GPs and district nurses. Logistic regression was used to determine relationships between these measures. RESULTS: Seven hundred and seventeen stroke survivors were registered with first stroke between 1995 and 1998. Most patients were followed-up on at least one occasion by at least one service within the first 3 months after stroke: 51% saw a hospital specialist; 72% saw a GP; and 14% saw a community nurse. However, 14% of patients did not see a doctor at all. Disabled patients were less likely to see a doctor, only 17% of severely disabled patients seeing a hospital specialist [odds ratio (OR) 0.17; 95% confidence interval (CI) 0.07-0.41]. Doctor-led follow up was related to treatment of physiological risk factors (e.g. 73% of hypertensive patients who had seen a GP were treated compared with 59% who had seen only a hospital specialist and 47% who had seen neither). Contact with health services was not associated with behavioural risk factor change. CONCLUSIONS: Opportunities for delivering secondary prevention existed through a range of services, but problems of continuity and effectiveness of care are evident. Further investigation is needed to determine how best to intervene to address these issues. In other words, whether interventions should concentrate on improving access and availability of current services, or whether the focus should be on making current strategies more effective. PMID- 12110560 TI - Promoting influenza vaccination of elderly patients in primary care. AB - BACKGROUND: Though influenza is a serious health problem for elderly people, their influenza vaccination rate remains low in Switzerland. OBJECTIVE: Our aim was to assess the impact of an intervention combining multiple strategies to promote influenza vaccination of elderly patients in primary care. METHODS: We conducted a pre-/post-intervention study in a university-based primary care clinic in Geneva, Switzerland, where an annual community-wide campaign promotes influenza vaccination of people at high risk. We included 318 and 346 patients aged over 64 years attending the clinic during the last trimesters of 1995 and 1996, respectively. The intervention included: patient information by leaflets and posters, a walk-in vaccination clinic, a training workshop for physicians, record reminders and peer comparison feedback on vaccination performance. Using the computerized database, medical records and the vaccination register, we measured influenza immunization rates and relative benefits (RBs) of the intervention. RESULTS: Influenza vaccine uptake globally increased from 21.7% before the intervention to 51.7% thereafter. Among 144 patients attending in both phases, the immunization rate rose from 29.2 to 69.4% [matched RB estimate () = 2.4; 95% confidence interval (CI) 1.9-3.0]; vaccine uptake increased particularly among all chronic patients ( = 3.2; 95% CI 2.2-4.6), cardiac patients ( = 3.4; 95% CI 2.1-5.4) and diabetics ( = 3.3; 95% CI 1.9-5.9). For 376 patients attending in a single phase, the vaccination rate rose from 15.5 to 39.1% (adjusted RB = 2.8; 95% CI 1.8-4.4), particularly among the elderly aged 65-75 years (adjusted RB = 5.7; 95% CI 2.7-12.4). CONCLUSION: An intervention combining strategies targeting patients, physicians and care delivery significantly increased influenza vaccine uptake of elderly patients in primary care, particularly those at high risk. PMID- 12110561 TI - Traumatic events in a general practice population: the patient's perspective. AB - OBJECTIVES: The aim of the present study was to describe the patient's perspective on the GP's care after violent events: which role is the GP assigned; and how is the care appreciated. Events studied were serious accidents, burglary, robbery, physical and sexual abuse, disasters and war. METHOD: A postal questionnaire was sent to a random sample of 2997 patients (> or =20 years) from the practice population of 32 GPs (67 500 patients). RESULTS: The response was 50%. Forty-two per cent of the respondents had experienced one or more events. Twenty-eight per cent of the victims desired some kind of professional help; more than half of them desired that care from their GP, three-quarters actually seeking it. Most frequently sought care was sympathy, "a number of good talks", and care for physical complaints. Overall, contentment with the GP's contribution was high; patients especially appreciate sympathy and support, as well as initiative on the GP's part in commencing and pursuing care. Of those who felt no need for professional help, 88% found that they could cope with the traumatic event well enough, with or without the help of family and friends. For those who did not seek help, although they did desire it, the main reasons were that they considered their problems insufficiently medical or felt that their GP lacked the time. In the case of physical and sexual abuse, feelings of guilt and issues of patient confidentiality played a role for some patients. CONCLUSIONS: The number of events experienced by our respondents is lower than in previous studies for burglary, robbery, physical and sexual abuse (adults and children); the occurrence of accidents is similar. The majority of the people who experience traumatic events cope with them well enough without professional help. For those seeking help, the GP plays an important role. Care could be improved as follows: the GP should make it clear to patients that he/she can play a role in caring for them in the aftermath of a traumatic event and stress the confidential nature of the consultation. On the whole, GPs should be more supportive and attentive when being consulted about this topic; also patients would like their doctors to be more active in raising the subject, as well as in initiating follow-up. PMID- 12110562 TI - Depression in primary care. A nationwide epidemiological survey. AB - BACKGROUND: Most epidemiological studies on depression in primary care are conducted at single sites, and variations in reported prevalence may depend on characteristics of health care services and other local factors. OBJECTIVES: Our aim was to investigate the prevalence of depression in primary care in Italy and its association with physical illness, disability and health care utilization. METHODS: This nationwide epidemiological study involved 191 primary care physicians (PCPs) who assessed during one index week 1896 patients aged 14 and over attending their clinics. Screening was conducted by using the General Health Questionnaire-12. Probable cases were assessed by PCPs with the WHO ICD-10 Checklist for Depression and rated for severity of physical illness. RESULTS: The prevalence of current depression ranged between 7.8 and 9.0% in the three main Italian areas, with no significant variations. A linear increase from North to South was observed for psychological distress, disability and frequency of medical consultation. Depression was associated with severe, but not with mild or moderate physical illness. Depression was also associated with disability and accounted for an increased rate of consultation. CONCLUSION: Because of the disability associated with depression and of its impact on health care utilization, guidelines and intervention strategies are needed. PMID- 12110563 TI - Tackling teenage turmoil: primary care recognition and management of mental ill health during adolescence. AB - This paper examines how primary care can improve for teenagers who are experiencing mental or emotional turmoil. This is an important health issue because at least 15% of teenagers experience mental health problems at any one time, there are indications that this proportion is rising, and there is evidence that suicide rates are rising in young people. The paper discusses how troubled teenagers can be identified, cared for and managed by primary care providers within the UK, although some of the information presented is from other countries. It identifies inter-relationships with other health behaviours and risk factors. The GP's role in assessing a troubled teenager is discussed, as well as a consideration of individual and contextual issues to frame a "triple" diagnosis, i.e. a diagnosis simultaneously in biomedical, individual and contextual terms. A review of present knowledge of management is presented. The paper concludes that there are several deficiencies at present, namely a lack of identification of teenage distress, a lack of training for GPs in teenage health, a lack of a research base, a lack of resources and finally a lack of information provided by any teenagers who have experienced turmoil and could give useful insights into their experience. PMID- 12110564 TI - Five years of family health care in Sao Jose. AB - In 1994, the Federal Government of Brazil enacted legislation to share the costs with municipalities of establishing or remodelling up to 20 000 health clinics, covering a population of 69 million people. Sao Jose clinic was established with family physicians in 1993 in a community of 3000 in the City of Curitiba. The clinic was functioning by 1995 when the Canadian four principles of Family Medicine were introduced to clinic staff. The impact of the clinic's work has measured improvements in perinatal mortality and child nutrition, reduced hepatitis A infection and produced dramatic improvements in delivery of preventive services. The presence of the clinic has empowered a poor community to demand improved municipal services that have helped to improve overall health. The introduction of Family Health Clinics in Brazil, and assistance provided by Canada, has achieved the objectives of the national Government in one sample site. PMID- 12110565 TI - Graduates' evaluation of a postgraduate diploma course in family medicine. AB - BACKGROUND: Postgraduate programmes offer an opportunity to learn family medicine for physicians in practice who were unable to obtain formal training in the immediate postgraduate phase of their career. Since 1985, the Chinese University of Hong Kong has provided a part-time 1-year diploma course at hours convenient for private practitioners. The curriculum has evolved, reducing public health components and increasing family medicine concepts. Between six and 16 students took the course each year until 1999, when formal recognition led to increased popularity. OBJECTIVE: The aim of this study was to evaluate the components and outcomes of the course as a prelude to further development. METHODS: Evaluation comprised a structured telephone interview conducted with two enrollees from each year of the course (total 28), selected randomly from class lists. RESULTS: Participants were mostly young doctors, with an average of 5 years in general practice. Many graduates are now prominent in training and development of family medicine in Hong Kong. Graduates rated most components favourably, but found the original research components too demanding, and not useful subsequently for most. Counselling, family dynamics, consultation and practice organization skills were valued. Conventional continuing education components, such as lectures by specialists, were evaluated poorly. CONCLUSIONS: This course has proved useful in the Hong Kong context, being practical for physicians, and allowing them to study ideas they would not otherwise encounter. Critical appraisal and evidence-based medicine exercises now replace the former research components. PMID- 12110566 TI - Selections from current literature. Horton hears a Who but no murmurs--does it matter? PMID- 12110569 TI - Projection structure at 8 A resolution of the melibiose permease, an Na-sugar co transporter from Escherichia coli. AB - The ion-coupled sugar membrane symporter or co-transporter melibiose permease (MelB), responsible for alpha-galactoside accumulation in Escherichia coli, is a representative member of the glycoside-pentoside- hexuronide family of the vast class of electrochemical potential-driven porters. Pure solubilized preparations of a MelB recombinant protein were subjected to two-dimensional crystallization trials and several crystal forms were observed. Two of these appeared as large wide tubes suitable for analysis by electron crystallography. Flattened tubes on carbon support film, embedded in amorphous ice prior to electron cryomicroscopy, showed two-sided plane group symmetries P12(1) or P222(1), with unit cell dimensions a = 89.9 A, b = 51.6 A, gamma = 91.9 degrees and a = 188.9 A, b = 48.8 A, gamma = 90 degrees, respectively. The projection map from the P222(1 )crystals at 8 A resolution displayed an asymmetric protein unit consisting of two domains lining a central and curve-shaped cleft. Together, the MelB monomer could host the 12 predicted transmembrane alpha-helices. Overall, the MelB helix packing arrangement compared more favorably with that of the Na(+)/H(+) antiporter NhaA than that of the oxalate antiporter. PMID- 12110570 TI - Three-dimensional structure of the type 1 inositol 1,4,5-trisphosphate receptor at 24 A resolution. AB - We report here the first three-dimensional structure of the type 1 inositol 1,4,5 trisphosphate receptor (IP(3)R). From cryo-electron microscopic images of purified receptors embedded in vitreous ice, a three-dimensional structure was determined by use of standard single particle reconstruction techniques. The structure is strikingly different from that of the ryanodine receptor at similar resolution despite molecular similarities between these two calcium release channels. The 24 A resolution structure of the IP(3)R takes the shape of an uneven dumbbell, and is approximately 170 A tall. Its larger end is bulky, with four arms protruding laterally by approximately 50 A and, in comparison with the receptor topology, probably corresponds to the cytoplasmic domain of the receptor. The lateral dimension at the height of the protruding arms is approximately 155 A. The smaller end, whose lateral dimension is approximately 100 A, has structural features indicative of the membrane-spanning domain. A central opening in this domain, which is occluded on the cytoplasmic half, outlines a pathway for calcium flow in the open state of the channel. PMID- 12110571 TI - Domain movements of plasma membrane H(+)-ATPase: 3D structures of two states by electron cryo-microscopy. AB - H(+)-ATPase is a P-type ATPase that transports protons across membranes using the energy from ATP hydrolysis. This hydrolysis is coupled to a conformational change between states of the protein, in which the proton-binding site is alternately accessible to the two sides of the membrane with an altered affinity. When isolated from Neurospora crassa, H(+)-ATPase is a 600 kDa hexamer of identical 100 kDa polypeptides. We have obtained the three-dimensional structures of both ligand-free and Mg(2+)/ADP-bound states of this complex using single-particle electron cryo- microscopy. Structural comparisons, together with the difference map, indicate that there is a rearrangement of the cytoplasmic domain on Mg(2+)/ADP binding, which consists of a movement of mass towards the 6-fold axis causing the structure to become more compact, accompanied by a modest conformational change in the transmembrane domain. PMID- 12110572 TI - Alternative splicing modulates the frequency-dependent response of CaMKII to Ca(2+) oscillations. AB - Ca(2+) oscillations are required in various signal trans duction pathways, and contain information both in their amplitude and frequency. Remarkably, the Ca(2+)/calmodulin(CaM)-dependent protein kinase II (CaMKII) can decode such frequencies. A Ca(2+)/CaM-stimulated autophosphorylation leads to Ca(2+)/CaM independent (autonomous) activity of the kinase that outlasts the initial stimulation. This autonomous activity increases exponentially with the frequency of Ca(2+) oscillations. Here we show that three beta-CaMKII splice variants (beta(M), beta and beta(e)') have very similar specific activity and maximal autonomy. However, their autonomy generated by Ca(2+) oscillations differs significantly. A mechanistic basis was found in alterations of the CaM activation constant and of the initial rate of autophosphorylation. Structurally, the splice variants differ only in a variable 'linker' region between the kinase and association domains. Therefore, we propose that differences in relative positioning of kinase domains within multimeric holoenzymes are responsible for the observed effects. Notably, the beta-CaMKII splice variants are differentially expressed, even among individual hippocampal neurons. Taken together, our results suggest that alternative splicing provides cells with a mechanism to modulate their sensitivity to Ca(2+) oscillations. PMID- 12110573 TI - Conformational changes in surface structures of isolated connexin 26 gap junctions. AB - Gap junction channels mediate communication between adjacent cells. Using atomic force microscopy (AFM), we have imaged conformational changes of the cytoplasmic and extracellular surfaces of native connexin 26 gap junction plaques. The cytoplasmic domains of the gap junction surface, imaged at submolecular resolution, form a hexameric pore protruding from the membrane bilayer. Exhibiting an intrinsic flexibility, these cytoplasmic domains, comprising the C terminal connexin end, reversibly collapse by increasing the forces applied to the AFM stylus. The extracellular connexon surface was imaged after dissection of the gap junction with the AFM stylus. Upon injection of Ca(2+) into the buffer solution, the extracellular channel entrance reduced its diameter from 1.5 to 0.6 nm, a conformational change that is fully reversible and specific among the divalent cations tested. Ca(2+) had a profound effect on the cytoplasmic surface also, inducing the formation of microdomains. Consequently, the plaque height increased by 0.6 nm to 18 nm. This suggests that calcium ions induce conformational changes affecting the structure of both the hemichannels and the intact channels forming cell-cell contacts. PMID- 12110574 TI - PKC epsilon controls the traffic of beta1 integrins in motile cells. AB - Protein kinase C (PKC) has been implicated in beta 1 integrin-mediated cell migration. Expression of the novel PKC isoform, PKC epsilon, in PKC epsilon(-/-) cells is shown here to stimulate directional migration of cells towards beta 1 integrin substrates in a manner dependent on PKC catalytic activity. On PKC inhibition, integrin beta 1 and PKC epsilon become reversibly trapped in a tetraspanin (CD81)-positive intracellular compartment, correlating with reduced haptotaxis. Immunofluorescence and pulse labelling studies indicate that this is a previously uncharacterized recycling compartment trapped by inhibition of PKC. Electron microscopy demonstrated the co-localization of PKC epsilon and integrin beta 1 on the vesicular membranes. Finally, using a reconstituted in vitro system, the dissociation of PKC epsilon from these vesicles is shown to be dependent on both the presence of cytosolic components and energy, and on PKC catalytic activity. The evidence presented indicates that PKC epsilon controls an internal traffic step that under uninhibited conditions permits the recycling of beta 1 integrin, contributing to cell motility. PMID- 12110575 TI - How Tlg2p/syntaxin 16 'snares' Vps45. AB - Soluble N-ethylmaleimide sensitive factor-attachment protein receptors (SNAREs) and Sec1p/Munc18-homologs (SM proteins) play key roles in intracellular membrane fusion. The SNAREs form tight four-helix bundles (core complexes) that bring the membranes together, but it is unclear how this activity is coupled to SM protein function. Studies of the yeast trans-Golgi network (TGN)/endosomal SNARE complex, which includes the syntaxin-like SNARE Tlg2p, have suggested that its assembly requires activation by binding of the SM protein Vps45p to the cytoplasmic region of Tlg2p folded into a closed conformation. Nuclear magnetic resonance and biochemical experiments now show that Tlg2p and Pep12p, a late- endosomal syntaxin that interacts functionally but not directly with Vps45p, have a domain structure characteristic of syntaxins but do not adopt a closed conformation. Tlg2p binds tightly to Vps45p via a short N-terminal peptide motif that is absent in Pep12p. The Tlg2p/Vps45p binding mode is shared by the mammalian syntaxin 16, confirming that it is a Tlg2p homolog, and resembles the mode of interaction between the SM protein Sly1p and the syntaxins Ufe1p and Sed5p. Thus, this mechanism represents the most widespread mode of coupling between syntaxins and SM proteins. PMID- 12110576 TI - Ferrichrome induces endosome to plasma membrane cycling of the ferrichrome transporter, Arn1p, in Saccharomyces cerevisiae. AB - Siderophores are small iron-binding molecules that are synthesized and secreted in the iron-free form by microorganisms. Saccharomyces cerevisiae takes up iron bound to siderophores by two separate systems, one of which requires the ARN family of sidero phore-iron transporters. Arn1p and Arn3p are expressed in endosome-like intracellular vesicles. Here we present evidence that, in the absence of its specific substrate, ferrichrome, Arn1p is sorted directly from the Golgi to the endosomal compartment and does not cycle to the plasma membrane. When cells are exposed to ferrichrome at low concentrations, Arn1p stably relocalizes to the plasma membrane. At higher concentrations of ferrichrome, Arn1p relocalizes to the plasma membrane and rapidly undergoes endocytosis. Plasma membrane localization of Arn1p occurs only in the presence of its specific substrate, and not in the presence of other siderophores. Despite expression of Arn1p on the plasma membrane, mutant strains with defects in endocytosis exhibit reduced uptake of ferrichrome-iron. Thus, siderophores influence the trafficking of the Arn transporters within the cell and this trafficking is important for transporter function. PMID- 12110577 TI - Light reception and circadian behavior in 'blind' and 'clock-less' mutants of Neurospora crassa. AB - The filamentous fungus Neurospora crassa is a model organism for the genetic dissection of blue light photoreception and circadian rhythms. WHITE COLLAR-1 (WC 1) and WC-2 are considered necessary for all light responses, while FREQUENCY (FRQ) is required for light-regulated asexual development (conidia formation); without any of the three, self-sustained (circadian) rhythmicity in constant conditions fails. Here we show that light-regulated and self-sustained development occur in the individual or mutant white collar strains. These strains resemble wild type in their organization of the daily bout of light-regulated conidiation. Molecular profiles of light- induced genes indicate that the individual white collar-1 and white collar-2 mutants utilize distinct pathways, despite their similar appearance in all aspects. Titration of fluence rate also demonstrates different light sensitivities between the two strains. The data require the existence of an as-yet-unidentified photoreceptor. Furthermore, the extant circadian clock machinery in these mutant strains supports the notion that the circadian system in Neurospora involves components outside the WC-FRQ loop. PMID- 12110578 TI - Absence of the prion protein homologue Doppel causes male sterility. AB - The agent that causes prion diseases is thought to be identical with PrP(Sc), a conformer of the normal prion protein PrP(C). PrP(C)-deficient mice do not exhibit major pathologies, perhaps because they express a protein termed Dpl, which shares significant biochemical and structural homology with PrP(C). To investigate the physiological function of Dpl, we generated mice harbouring a homozygous disruption of the Prnd gene that encodes Dpl. Dpl deficiency did not interfere with embryonic and postnatal development, but resulted in male sterility. Dpl protein was expressed at late stages of spermiogenesis, and spermatids of Dpl mutants were reduced in numbers, immobile, malformed and unable to fertilize oocytes in vitro. Mechanical dissection of the zona pellucida partially restored in vitro fertilization. We conclude that Dpl regulates male fertility by controlling several aspects of male gametogenesis and sperm-egg interaction. PMID- 12110579 TI - The protein import motor of mitochondria: a targeted molecular ratchet driving unfolding and translocation. AB - Unfolding and import of preproteins into mitochondria are facilitated by a molecular motor in which heat shock protein 70 (Hsp70) in the matrix plays an essential role. Here we present two different experimental approaches to analyze mechanisms underlying this function of Hsp70. First, preproteins containing stretches of glutamic acid (polyE) or glycine (polyG) repeats in front of folded domains were imported into mitochondria. This occurred although Hsp70 cannot pull on these stretches to unfold the folded domains, since it does not bind to polyE and polyG. Secondly, preproteins containing titin immunoglobulin (Ig)-like domains were imported into mitochondria, despite the fact that forces of >200 pN are required to mechanically unfold these domains. Since molecular motors generate forces of approximately 5 pN, Hsp70 could not promote unfolding of the Ig-like domains by mechanical pulling. Our observations suggest that Hsp70 acts as an element of a Brownian ratchet, which mediates unfolding and translocation of preproteins across the mitochondrial membranes. PMID- 12110580 TI - A GAF-domain-regulated adenylyl cyclase from Anabaena is a self-activating cAMP switch. AB - The gene cyaB1 from the cyanobacterium Anabaena sp. PCC 7120 codes for a protein consisting of two N-terminal GAF domains (GAF-A and GAF-B), a PAS domain and a class III adenylyl cyclase catalytic domain. The catalytic domain is active as a homodimer, as demonstrated by reconstitution from complementary inactive point mutants. The specific activity of the holoenyzme increased exponentially with time because the product cAMP activated dose dependently and nucleotide specifically (half-maximally at 1 microM), identifying cAMP as a novel GAF domain ligand. Using point mutants of either the GAF-A or GAF-B domain revealed that cAMP activated via the GAF-B domain. We replaced the cyanobacterial GAF domain ensemble in cyaB1 with the tandem GAF-A/GAF-B assemblage from the rat cGMP stimulated phosphodiesterase type 2, and converted cyaB1 to a cGMP-stimulated adenylyl cyclase. This demonstrated the functional conservation of the GAF domain ensemble since the divergence of bacterial and eukaryotic lineages >2 billion years ago. In cyanobacteria, cyaB1 may act as a cAMP switch to stabilize committed developmental decisions. PMID- 12110581 TI - Selenoproteins and selenocysteine insertion system in the model plant cell system, Chlamydomonas reinhardtii. AB - Known eukaryotic selenocysteine (Sec)-containing proteins are animal proteins, whereas selenoproteins have not been found in yeast and plants. Surprisingly, we detected selenoproteins in a member of the plant kingdom, Chlamydomonas reinhardtii, and directly identified two of them as phospholipid hydroperoxide glutathione peroxidase and selenoprotein W homologs. Moreover, a selenocysteyl tRNA was isolated that recognized specifically the Sec codon UGA. Subsequent gene cloning and bioinformatics analyses identified eight additional selenoproteins, including methionine-S-sulfoxide reductase, a selenoprotein specific to Chlamydomonas: Chlamydomonas selenoprotein genes contained selenocysteine insertion sequence (SECIS) elements that were similar, but not identical, to those of animals. These SECIS elements could direct selenoprotein synthesis in mammalian cells, indicating a common origin of plant and animal Sec insertion systems. We found that selenium is required for optimal growth of Chlamydomonas: Finally, evolutionary analyses suggested that selenoproteins present in Chlamydomonas and animals evolved early, and were independently lost in land plants, yeast and some animals. PMID- 12110582 TI - Chk1 is activated transiently and targets Cdc25A for degradation at the Xenopus midblastula transition. AB - In Xenopus embryos, cell cycle elongation and degradation of Cdc25A (a Cdk2 Tyr15 phosphatase) occur naturally at the midblastula transition (MBT), at which time a physiological DNA replication checkpoint is thought to be activated by the exponentially increased nucleo-cytoplasmic ratio. Here we show that the checkpoint kinase Chk1, but not Cds1 (Chk2), is activated transiently at the MBT in a maternal/zygotic gene product-regulated manner and is essential for cell cycle elongation and Cdc25A degradation at this transition. A constitutively active form of Chk1 can phosphorylate Cdc25A in vitro and can target it rapidly for degradation in pre-MBT embryos. Intriguingly, for this degradation, however, Cdc25A also requires a prior Chk1-independent phosphorylation at Ser73. Ectopically expressed human Cdc25A can be degraded in the same way as Xenopus Cdc25A. Finally, Cdc25A degradation at the MBT is a prerequisite for cell viability at later stages. Thus, the physiological replication checkpoint is activated transiently at the MBT by developmental cues, and activated Chk1, only together with an unknown kinase, targets Cdc25A for degradation to ensure later development. PMID- 12110583 TI - c-FLIP(L) is a dual function regulator for caspase-8 activation and CD95-mediated apoptosis. AB - Activation of the caspase cascade is a pivotal step in apoptosis and can occur via death adaptor-mediated homo-oligomerization of initiator procaspases. Here we show that c-FLIP(L), a protease-deficient caspase homolog widely regarded as an apoptosis inhibitor, is enriched in the CD95 death-inducing signaling complex (DISC) and potently promotes procaspase-8 activation through hetero-dimerization. c-FLIP(L) exerts its effect through its protease-like domain, which associates efficiently with the procaspase-8 protease domain and induces the enzymatic activity of the zymogen. Ectopic expression of c-FLIP(L) at physiologically relevant levels enhances procaspase-8 processing in the CD95 DISC and promotes apoptosis, while a decrease of c-FLIP(L) expression results in inhibition of apoptosis. c-FLIP(L) acts as an apoptosis inhibitor only at high ectopic expression levels. Thus, c-FLIP(L) defines a novel type of caspase regulator, distinct from the death adaptors, that can either promote or inhibit apoptosis. PMID- 12110585 TI - Essential role of PDK1 in regulating cell size and development in mice. AB - PDK1 functions as a master kinase, phosphorylating and activating PKB/Akt, S6K and RSK. To learn more about the roles of PDK1, we generated mice that either lack PDK1 or possess PDK1 hypomorphic alleles, expressing only approximately 10% of the normal level of PDK1. PDK1(-/-) embryos die at embryonic day 9.5, displaying multiple abnormalities including lack of somites, forebrain and neural crest derived tissues; however, development of hind- and midbrain proceed relatively normally. In contrast, hypomorphic PDK1 mice are viable and fertile, and insulin injection induces the normal activation of PKB, S6K and RSK. Nevertheless, these mice are 40-50% smaller than control animals. The organ volumes from the PDK1 hypomorphic mice are reduced proportionately. We also establish that the volume of a number of PDK1-deficient cells is reduced by 35 60%, and show that PDK1 deficiency does not affect cell number, nuclear size or proliferation. We provide genetic evidence that PDK1 is essential for mouse embryonic development, and regulates cell size independently of cell number or proliferation, as well as insulin's ability to activate PKB, S6K and RSK. PMID- 12110584 TI - Tyrosine phosphorylation of Mdm2 by c-Abl: implications for p53 regulation. AB - The p53 tumor suppressor is inhibited and destabilized by Mdm2. However, under stress conditions, this downregulation is relieved, allowing the accumulation of biologically active p53. Recently we showed that c-Abl is important for p53 activation under stress conditions. In response to DNA damage, c-Abl protects p53 by neutralizing the inhibitory effects of Mdm2. In this study we ask whether this neutralization involves a direct interplay between c-Abl and Mdm2, and what is the contribution of the c-Abl kinase activity? We demonstrate that the kinase activity of c-Abl is required for maintaining the basal levels of p53 expression and for achieving maximal accumulation of p53 in response to DNA damage. Importantly, c-Abl binds and phosphorylates Mdm2 in vivo and in vitro. We characterize Hdm2 (human Mdm2) phosphorylation at Tyr394. Substitution of Tyr394 by Phe394 enhances the ability of Mdm2 to promote p53 degradation and to inhibit its transcriptional and apoptotic activities. Our results suggest that phosphorylation of Mdm2 by c-Abl impairs the inhibition of p53 by Mdm2, hence defining a novel mechanism by which c-Abl activates p53. PMID- 12110587 TI - TGF-beta receptor-activated p38 MAP kinase mediates Smad-independent TGF-beta responses. AB - Through the action of its membrane-bound type I receptors, transforming growth factor-beta (TGF-beta) elicits a wide range of cellular responses that regulate cell proliferation, differentiation and apoptosis. Many of the signaling responses induced by TGF-beta are mediated by Smad proteins, but certain evidence has suggested that TGF-beta can also signal independently of Smads. We found in mouse mammary epithelial (NMuMG) cells, which respond to TGF-beta treatment in multiple ways, that TGF-beta-induced activation of p38 MAP kinase is required for TGF-beta-induced apoptosis, epithelial-to-mesenchymal transition (EMT), but not growth arrest. We further demonstrated that activation of p38 is independent of Smads using a mutant type I receptor, which is incapable of activating Smads but still retains the kinase activity. This mutant receptor is sufficient to activate p38 and cause NMuMG cells to undergo apoptosis. However, it is not sufficient to induce EMT. These results indicate that TGF-beta receptor signals through multiple intracellular pathways and provide first-hand biochemical evidence for the existence of Smad-independent TGF-beta receptor signaling. PMID- 12110586 TI - Loss of p27(Kip1) but not p21(Cip1) decreases survival and synergizes with MYC in murine lymphomagenesis. AB - The cyclin-dependent kinase (CDK) inhibitors p21(Cip1) and p27(Kip1) are induced in response to anti-proliferative stimuli and block G(1)/S-phase progression through the inhibition of CDK2. Although the cyclin E-CDK2 pathway is often deregulated in tumors the relative contribution of p21(Cip1) and p27(Kip1) to tumorigenesis is still unclear. The MYC transcription factor is an important regulator of the G(1)/S transition and its expression is frequently altered in tumors. Previous reports suggested that p27(Kip1) is a crucial G(1) target of MYC. Our study shows that in mice, deficiency for p27(Kip1) but not p21(Cip1) results in decreased survival to retrovirally-induced lymphomagenesis. Importantly, in such p27(Kip1) deficient lymphomas an increased frequency of Myc activation is observed. p27(Kip1) deficiency was also shown to collaborate with MYC overexpression in transgenic lymphoma models. Thus, in vivo, the capacity of MYC to promote tumor growth is fully retained and even enhanced upon p27(Kip1) loss. We show that in lymphocytes, MYC overexpression and p27(Kip1) deficiency independently stimulate CDK2 activity and augment the fraction of cells in S phase, in support of their distinct roles in tumorigenesis. PMID- 12110588 TI - Phosphorylation by p38MAPK and recruitment of SUG-1 are required for RA-induced RAR gamma degradation and transactivation. AB - The nuclear retinoic acid receptor RAR gamma 2 undergoes proteasome-dependent degradation upon ligand binding. Here we provide evidence that the domains that signal proteasome-mediated degradation overlap with those that activate transcription, i.e. the activation domains AF-1 and AF-2. The AF-1 domain signals RAR gamma 2 degradation through its phosphorylation by p38MAPK in response to RA. The AF-2 domain acts via the recruitment of SUG-1, which belongs to the 19S regulatory subunit of the 26S proteasome. Blocking RAR gamma 2 degradation through inhibition of either the p38MAPK pathway or the 26S proteasome function impairs its RA-induced transactivation activity. Thus, the turnover of RAR gamma 2 is linked to transactivation. PMID- 12110589 TI - Transcription factor-mediated lineage switching reveals plasticity in primary committed progenitor cells. AB - The developmental plasticity of transplanted adult stem cells challenges the notion that tissue-restricted stem cells have stringently limited lineage potential and prompts a re-evaluation of the stability of lineage commitment. Transformed cell systems are inappropriate for such studies, since transformation potentially dysregulates the processes governing lineage commitment. We have therefore assessed the stability of normal lineage commitment in primary adult haematopoietic cells. For these studies we have used prospectively isolated primary bipotent progenitors, which normally display only neutrophil and monocyte differentiation in vitro. In response to ectopic transcription factor expression, these neutrophil/monocyte progenitors were reprogrammed to take on erythroid, eosinophil and basophil-like cell fates, with the resultant colonies resembling the mixed lineage colonies normally generated by multipotential progenitors. Clone-marking and daughter cell experiments identified lineage switching rather than differential cell selection as the mechanism of altered lineage output. These results demonstrate that the cell type-specific programming of apparently committed primary progenitors is not irrevocably fixed, but may be radically re specified in response to a single transcriptional regulator. PMID- 12110590 TI - Growth factors can activate ATF2 via a two-step mechanism: phosphorylation of Thr71 through the Ras-MEK-ERK pathway and of Thr69 through RalGDS-Src-p38. AB - Transcription factor ATF2 regulates gene expression in response to environmental changes. Upon exposure to cellular stresses, the mitogen-activated proteinkinase (MAPK) cascades including SAPK/JNK and p38 can enhance ATF2's transactivating function through phosphorylation of Thr69 and Thr71. How ever, the mechanism of ATF2 activation by growth factors that are poor activators of JNK and p38 is still elusive. Here, we show that in fibroblasts, insulin, epidermal growth factor (EGF) and serum activate ATF2 via a so far unknown two-step mechanism involving two distinct Ras effector pathways: the Raf-MEK-ERK pathway induces phosphorylation of ATF2 Thr71, whereas subsequent ATF2 Thr69 phosphorylation requires the Ral-RalGDS-Src-p38 pathway. Cooperation between ERK and p38 was found to be essential for ATF2 activation by these mitogens; the activity of p38 and JNK/SAPK in growth factor-stimulated fibroblasts is insufficient to phosphorylate ATF2 Thr71 or Thr69 + 71 significantly by themselves, while ERK cannot dual phosphorylate ATF2 Thr69 + 71 efficiently. These results reveal a so far unknown mechanism by which distinct MAPK pathways and Ras effector pathways cooperate to activate a transcription factor. PMID- 12110591 TI - Lariat formation and a hydrolytic pathway in plant chloroplast group II intron splicing. AB - Lariat formation has been studied intensively only with a few self-splicing group II introns, and little is known about how the numerous diverse introns in plant organelles are excised. Several of these introns have branch-points that are not a single bulge but are adjoined by A:A, A:C, A:G and G:G pairs. Using a highly sensitive in vivo approach, we demonstrate that all but one of the barley chloroplast introns splice via the common pathway that produces a branched product. RNA editing does not improve domain 5 and 6 structures of these introns. The conserved branch-point in tobacco rpl16 is chosen even if an adjacent unpaired adenosine is available, suggesting that spatial arrangements in domain 6 determine correct branch-point selection. Lariats were not detected for the chloroplast trnV intron, which lacks an unpaired adenosine in domain 6. Instead, this intron is released as linear molecules that undergo further polyadenylation. trnV, which is conserved throughout plant evolution, constitutes the first example of naturally occurring hydrolytic group II intron splicing in vivo. PMID- 12110592 TI - The splicing of U12-type introns can be a rate-limiting step in gene expression. AB - Some protein-coding genes in metazoan genomes contain a minor class of introns that are excised by a distinct, low-abundance spliceosome. We have developed a quantitative RT-PCR assay that allows comparison of the relative rates of intron removal from the transcripts present in a pre-mRNA population. We show that the U12-type introns are more slowly spliced than the major-class (U2-type) introns from three endogenous pre-mRNAs in human tissue culture cells. In Drosophila melanogaster S2 cells, using minigene constructs designed to produce nearly identical mRNAs, we observe increased expression of fluorescent protein and mature mRNA upon mutation of a U12-type to a U2-type intron. These results provide evidence that the level of gene expression in vivo is lowered by the presence of a U12-type intron and implicate the U12-type spliceosome as a target in the post-transcriptional regulation of gene expression. PMID- 12110593 TI - Site-specific cross-linking analyses reveal an asymmetric protein distribution for a box C/D snoRNP. AB - Methylation of the ribose 2'-hydroxyl, the most widespread modification of ribosomal and splicesomal RNAs, is guided by the box C/D class of small nucleolar RNAs (snoRNAs). Box C/D small nucleolar ribonucleoproteins (snoRNPs) contain four core proteins: fibrillarin, Nop56, Nop58 and 15.5 kDa. We constructed U25 snoRNAs containing a single photoactivatable 4-thiouridine at each U position within the conserved box C/D and C'/D' motifs. Proteins assembled on the snoRNA after injection into Xenopus oocyte nuclei were identified by cross-linking, and reconstituted particles characterized by functional rescue and mutational analyses. Our data argue that box C/D snoRNPs are asymmetric, with the C' box contacting Nop56 and fibrillarin, the C box interacting with Nop58, and the D and D' boxes contacting fibrillarin. No cross-link to 15.5 kDa was detected; its binding is disrupted by 4-thiouridine substitution in position 1 of the C box. Repositioning the guide sequence of U25 upstream of box D instead of D' revealed that both C/D motifs have the potential to function as guide centers, but, surprisingly, there was no alteration in protein cross-linking. PMID- 12110594 TI - Class I tyrosyl-tRNA synthetase has a class II mode of cognate tRNA recognition. AB - Bacterial tyrosyl-tRNA synthetases (TyrRS) possess a flexibly linked C-terminal domain of approximately 80 residues, which has hitherto been disordered in crystal structures of the enzyme. We have determined the structure of Thermus thermophilus TyrRS at 2.0 A resolution in a crystal form in which the C-terminal domain is ordered, and confirm that the fold is similar to part of the C-terminal domain of ribosomal protein S4. We have also determined the structure at 2.9 A resolution of the complex of T.thermophilus TyrRS with cognate tRNA(tyr)(G Psi A). In this structure, the C-terminal domain binds between the characteristic long variable arm of the tRNA and the anti-codon stem, thus recognizing the unique shape of the tRNA. The anticodon bases have a novel conformation with A-36 stacked on G-34, and both G-34 and Psi-35 are base-specifically recognized. The tRNA binds across the two subunits of the dimeric enzyme and, remarkably, the mode of recognition of the class I TyrRS for its cognate tRNA resembles that of a class II synthetase in being from the major groove side of the acceptor stem. PMID- 12110595 TI - tadA, an essential tRNA-specific adenosine deaminase from Escherichia coli. AB - We report the characterization of tadA, the first prokaryotic RNA editing enzyme to be identified. Escherichia coli tadA displays sequence similarity to the yeast tRNA deaminase subunit Tad2p. Recombinant tadA protein forms homodimers and is sufficient for site-specific inosine formation at the wobble position (position 34) of tRNA(Arg2), the only tRNA having this modification in prokaryotes. With the exception of yeast tRNA(Arg), no other eukaryotic tRNA substrates were found to be modified by tadA. How ever, an artificial yeast tRNA(Asp), which carries the anticodon loop of yeast tRNA(Arg), is bound and modified by tadA. Moreover, a tRNA(Arg2) minisubstrate containing the anticodon stem and loop is sufficient for specific deamination by tadA. We show that nucleotides at positions 33-36 are sufficient for inosine formation in mutant Arg2 minisubstrates. The anticodon is thus a major determinant for tadA substrate specificity. Finally, we show that tadA is an essential gene in E.coli, underscoring the critical function of inosine at the wobble position in prokaryotes. PMID- 12110596 TI - Dissolution of the maskin-eIF4E complex by cytoplasmic polyadenylation and poly(A)-binding protein controls cyclin B1 mRNA translation and oocyte maturation. AB - Cytoplasmic polyadenylation stimulates the translation of several dormant mRNAs during oocyte maturation in Xenopus. Polyadenylation is regulated by the cytoplasmic polyadenylation element (CPE), a cis-acting element in the 3' untranslated region of responding mRNAs, and its associated factor CPEB. CPEB also binds maskin, a protein that in turn interacts with eIF4E, the cap-binding factor. Here, we report that based on antibody and mRNA reporter injection assays, maskin prevents oocyte maturation and the translation of the CPE containing cyclin B1 mRNA by blocking the association of eIF4G with eIF4E. Dissociation of the maskin-eIF4E complex is essential for cyclin B1 mRNA translational activation, and requires not only cytoplasmic polyadenylation, but also the poly(A)-binding protein. These results suggest a molecular mechanism by which CPE- containing mRNA is activated in early development. PMID- 12110597 TI - Crystal structure of human 53BP1 BRCT domains bound to p53 tumour suppressor. AB - The BRCT (BRCA1 C-terminus) is an evolutionary conserved protein-protein interacting module found as single, tandem or multiple repeats in a diverse range of proteins known to play roles in the DNA-damage response. The BRCT domains of 53BP1 bind to the tumour suppressor p53. To investigate the nature of this interaction, we have determined the crystal structure of the 53BP1 BRCT tandem repeat in complex with the DNA-binding domain of p53. The structure of the 53BP1 p53 complex shows that the BRCT tandem repeats pack together through a conserved interface that also involves the inter-domain linker. A comparison of the structure of the BRCT region of 53BP1 with the BRCA1 BRCT tandem repeat reveals that the interdomain interface and linker regions are remarkably well conserved. 53BP1 binds to p53 through contacts with the N-terminal BRCT repeat and the inter BRCT linker. The p53 residues involved in this binding are mutated in cancer and are also important for DNA binding. We propose that BRCT domains bind to cellular target proteins through a conserved structural element termed the 'BRCT recognition motif'. PMID- 12110598 TI - Structure and mechanism of T4 polynucleotide kinase: an RNA repair enzyme. AB - T4 polynucleotide kinase (Pnk), in addition to being an invaluable research tool, exemplifies a family of bifunctional enzymes with 5'-kinase and 3'-phosphatase activities that play key roles in RNA and DNA repair. T4 Pnk is a homotetramer composed of a C-terminal phosphatase domain and an N-terminal kinase domain. The 2.0 A crystal structure of the isolated kinase domain highlights a tunnel-like active site through the heart of the enzyme, with an entrance on the 5' OH acceptor side that can accommodate a single-stranded polynucleotide. The active site is composed of essential side chains that coordinate the beta phosphate of the NTP donor and the 3' phosphate of the 5' OH acceptor, plus a putative general acid that activates the 5' OH. The structure rationalizes the different specificities of T4 and eukaryotic Pnk and suggests a model for the assembly of the tetramer. PMID- 12110599 TI - Lesion bypass in yeast cells: Pol eta participates in a multi-DNA polymerase process. AB - Replication through (6-4)TT and G-AAF lesions was compared in Saccharomyces cerevisiae strains proficient and deficient for the RAD30-encoded DNA polymerase eta (Pol eta). In the RAD30 strain, the (6-4)TT lesion is replicated both inaccurately and accurately 60 and 40% of the time, respectively. Surprisingly, in a rad30 Delta strain, the level of mutagenic bypass is essentially suppressed, while error-free bypass remains unchanged. Therefore, Pol eta is responsible for mutagenic replication through the (6-4)TT photoproduct, while another polymerase mediates its error-free bypass. Deletion of the RAD30 gene also reduces the levels of both accurate and inaccurate bypass of AAF lesions within two different sequence contexts up to 8-fold. These data show that, in contrast to the accurate bypass by Pol eta of TT cyclobutane dimers, it is responsible for the mutagenic bypass of other lesions. In conclusion, this paper shows that, in yeast, translesion synthesis involves the combined action of several polymerases. PMID- 12110600 TI - Accessory factors determine the order of strand exchange in Xer recombination at psi. AB - Xer site-specific recombination in Escherichia coli converts plasmid multimers to monomers, thereby ensuring their correct segregation at cell division. Xer recombination at the psi site of plasmid pSC101 is preferentially intramolecular, giving products of a single topology. This intramolecular selectivity is imposed by accessory proteins, which bind at psi accessory sequences and activate Xer recombination at the psi core. Strand exchange proceeds sequentially within the psi core; XerC first exchanges top strands to produce Holliday junctions, then XerD exchanges bottom strands to give final products. In this study, recombination was analysed at sites in which the psi core was inverted with respect to the accessory sequences. A plasmid containing two inverted-core psi sites recombined with a reversed order of strand exchange, but with unchanged product topology. Thus the architecture of the synapse, formed by accessory proteins binding to accessory sequences, determines the order of strand exchange at psi. This finding has important implications for the way in which accessory proteins interact with the recombinases. PMID- 12110601 TI - Replication fork collapse at replication terminator sequences. AB - Replication fork arrest is a source of genome re arrangements, and the recombinogenic properties of blocked forks are likely to depend on the cause of blockage. Here we study the fate of replication forks blocked at natural replication arrest sites. For this purpose, Escherichia coli replication terminator sequences Ter were placed at ectopic positions on the bacterial chromosome. The resulting strain requires recombinational repair for viability, but replication forks blocked at Ter are not broken. Linear DNA molecules are formed upon arrival of a second round of replication forks that copy the DNA strands of the first blocked forks to the end. A model that accounts for the requirement for homologous recombination for viability in spite of the lack of chromosome breakage is proposed. This work shows that natural and accidental replication arrests sites are processed differently. PMID- 12110603 TI - Human immunodeficiency virus type 1 Vpr-mediated G(2) cell cycle arrest: Vpr interferes with cell cycle signaling cascades by interacting with the B subunit of serine/threonine protein phosphatase 2A. PMID- 12110602 TI - Promoter competition as a mechanism of transcriptional interference mediated by retrotransposons. AB - Enhancers can function over great distances and interact with almost any kind of promoter, but insulators or promoter competition generally limit their effect to a single gene. We provide in vivo evidence that retroelements may establish promoter competition with their neighboring genes and restrict the range of action of an enhancer. We report that the retroelement Idefix from Drosophila melanogaster inhibits white gene expression in testes by a promoter competition mechanism that does not occur in the eyes. The sequence specificity of the two TATA-less promoters of white and Idefix is a prime determinant in the competition that takes place in tissues where both are transcriptionally active. This study brings to light a novel mechanism whereby transcriptional interference by an active retrotransposon may perturb expression of neighboring genes. This capacity to interfere with the transcriptional regulation of their host, together with the facts that retroelements preferentially move within the germline and do not excise to replicate, suggest that these elements are cis-regulatory sequences able to imprint specific and heritable controls essential for eukaryotic gene regulation. PMID- 12110604 TI - Enhanced inhibition of L-type calcium currents by troglitazone in streptozotocin induced diabetic rat cardiac ventricular myocytes. AB - 1. Troglitazone, an insulin-sensitizing agent shown to improve cardiac function in both experimental animals and patients with diabetes, inhibits voltage dependent L-type Ca(2+) currents (I(Ca,L)) in cardiac myocytes, which may underlie its cardioprotective effects. However, inhibition by troglitazone of I(Ca,L) in diabetic cardiac myocytes has not been characterized. 2. Using whole cell voltage-clamp techniques, I(Ca,L) was measured in ventricular myocytes isolated from 4-6 weeks streptozotocin (STZ)-induced diabetic rats and age matched control rats. 3. Under control conditions with CsCl internal solution, diabetic myocytes did not differ from control myocytes in membrane capacitance, current density or voltage-dependent properties of I(Ca,L). 4. Troglitazone decreased amplitude of I(Ca,L) in both control and diabetic myocytes in a concentration-dependent manner. This inhibition was more potent in diabetic than in control myocytes; half-maximum inhibitory concentrations of troglitazone measured at a holding potential of -50 mV were 4.3 and 9.5 micromol l(-1), respectively. 5. Troglitazone at 5 micromol l(-1) did not significantly influence the voltage dependency of steady-state inactivation or the inactivation time course of I(Ca,L) in either control or diabetic myocytes. 6. Since troglitazone inhibits I(Ca,L) more effectively in STZ-induced diabetic ventricular myocytes, this agent may prevent cardiac dysfunction in diabetes. PMID- 12110605 TI - Effects of oestrogen replacement on steady and pulsatile haemodynamics in ovariectomized rats. AB - 1. The effects of ovariectomy (Ovx), menopause and oestrogen replacement on the haemodynamics remain controversial. The present study employed the technique of arterial impedance analysis to measure and calculate the steady and pulsatile haemodynamics. The purpose was to determine the haemodynamic consequence of ovariectomy and oestrogen replacement. 2. Ovariectomy was carried out under anaesthesia on female Sprague Dawley rats aged 9 weeks. Oestrogen (17 beta estradiol or E(2)) replacement started 1 week after ovariectomy for 4 weeks. Ovx increased the body weight (BW), while it greatly reduced the uterus weight. Left ventricular weight (LVW) was slightly increased, but LVW/BW ratio was slightly reduced. These changes were reversed after E(2) replacement. 3. Compared to sham group, Ovx with or without E(2) replacement did not significantly affect the systolic, mean and diastolic pressure. In Ovx, pulse pressure (PP) and heart rate were significantly increased, while stroke volume and cardiac output were slightly decreased. Total peripheral resistance (TPR) was largely elevated, indicating Ovx induced systemic vasoconstriction. These changes all returned to close normal values (sham group) after E(2) replacement, except PP. 4. Ovx increased the characteristic input impedance (Zc) and pulse wave reflection, while it decreased arterial compliance. E(2) treatment reversed these changes, except Zc. 5. These results demonstrate that Ovx influences both the resistance and Windkessel functions of the artery. E(2) treatment effectively reverses most the effects of Ovx both on the steady and pulsatile haemodynamics. PMID- 12110606 TI - Pressor and pulmonary responses to ET-1(1-31) in guinea-pigs. AB - 1. Endothelin-1(1-31) (ET-1(1-31); 0.25 to 4 nmol kg(-1); i.v.) induced, in the guinea-pig, graded increases in MAP and an indomethacin-sensitive enhancement of pulmonary insufflation pressure (PIP). At all doses, ET-1(1-31) induced a monophasic pressor response, except at 4 nmol kg(-1), which caused a rapid and transient response (first phase: over first 10 min after injection) followed by a more slowly-developing and sustained (second phase: between 10 and 45 min after injection) increase in MAP. ET-1(1-31) was 4 to 10 fold less potent than ET-1 on PIP responses. 2. Phosphoramidon (5 and 10 mg kg(-1)) reduced both pressor and PIP effects of ET-1(1-31). Thiorphan (0.25 and 2.5 mg kg(-1)) did not affect the pressor responses to ET-1(1-31) although its PIP effects were markedly reduced by the NEP inhibitor. A selective endothelin-converting enzyme (ECE) inhibitor, CGS 35066 (1 mg kg(-1)), significantly reduced the second phase pressor response and increase in PIP triggered by ET-1(1-31). 3. The second (but not the first) pressor phase of ET-1(1-31) (4 nmol kg(-1)) was markedly reduced by BQ-123 (selective ET(A) antagonist), whereas the increase of PIP was significantly reduced by BQ-788 (selective ET(B) antagonist). Co-administration of BQ-123 plus BQ-788 abolished ET-1(1-31)-induced increase in PIP, but blockade of the second pressor phase afforded by BQ-123 was now reversed. 4. In guinea-pig isolated perfused lungs, ET-1(1-31) (50 nM) induced the release of prostacyclin and thromboxane A(2), which was inhibited by BQ-788 (5 nM) or thiorphan (25 microM), but not BQ-123 (1 microM). 5. These results suggest that ET-1(1-31) enhances MAP. Its sustained, but not transient, pressor effects are mediated via ET(A) receptor activation. Furthermore, ET-1(1-31) increases airway resistance in vivo and triggers prostacyclin and thromboxane A(2) release from perfused lungs predominantly via ET(B) receptor activation. ET-1(1-31) failed to display any selectivity of action towards either ET(A) or ET(B) receptors in these models. 6. We suggest that, in order to raise MAP, ET-1(1-31) requires conversion to ET-1, predominantly by ECE and to a lesser extent neutral endopeptidase 24.11, whereas the reverse holds true regarding its pharmacological effects in airways. PMID- 12110607 TI - Expression and pharmacological profile of the human organic cation transporters hOCT1, hOCT2 and hOCT3. AB - 1. Organic cation transporters (OCTs) are involved in the elimination of monoamines and cationic xenobiotics. To examine whether some cell lines express several different OCTs, we investigated seven human cell lines for the mRNA expression pattern of the human (h) transporters hOCT1, hOCT2 and hOCT3. hOCT1 mRNA was found in all cell lines, six additionally expressed hOCT3 and only two cell lines contained all three hOCTs. 2. Among the three OCTs only for the OCT3 (also designated as 'uptake(2)' or 'extraneuronal monoamine transporter') 'selective' inhibitors are described in the literature. The affinities of the OCT3 inhibitors for the other two OCTs are largely unknown. Therefore, we compared the potencies of eight compounds as inhibitors of hOCT-mediated uptake of the organic cation [(3)H]-1-methyl-4-phenylpyridinium ([(3)H]-MPP(+)) in human embryonic kidney 293 (HEK293) cells stably expressing hOCT1, hOCT2 or hOCT3. Decynium-22 inhibited hOCT3 with 10 fold higher potency than hOCT1 and hOCT2. Corticosterone was about 100 fold more potent as inhibitor of hOCT3 than of hOCT1 or hOCT2, and O-methylisoprenaline (OMI) inhibited almost exclusively hOCT3. Progesterone and beta-Oestradiol preferentially inhibited hOCT3 and hOCT1, whereas prazosin was a potent inhibitor of hOCT1 and hOCT3. Phenoxybenzamine (PbA) inhibited with about equal apparent potency all three hOCTs, whereas the PbA derivative SKF550 ((9-fluorenyl)-N-methyl-beta-chloroethylamine) preferentially inhibited hOCT3 and hOCT2. 3. PbA reversibly inhibited hOCT1 and irreversibly hOCT2 and hOCT3; SKF550 also irreversibly inhibited hOCT3 but hOCT2 in a reversible manner. 4. These compounds enable a functional discrimination of the three hOCTs: hOCT1 is selectively inhibited by prazosin, reversibly inhibited by PbA and it is not sensitive to inhibition by SKF550 and OMI; hOCT2 is reversibly inhibited by SKF550, irreversibly by PbA and not by prazosin, beta oestradiol and OMI, whereas hOCT3 is selectively inhibited by corticosterone, OMI and decynium22. PMID- 12110608 TI - FcepsilonRI cross-linking-induced actin assembly mediates calcium signalling in RBL-2H3 mast cells. AB - 1. To determine the role of actin assembly in the Ca(2+) signalling of mast cells activated by cross-linking of FcepsilonRI, we examined the effects of cytochalasin D, an inhibitor of actin polymerization. 2. In the RBL-2H3 cells, F actin content was increased by sensitization with anti-dinitrophenol (DNP) IgE. In these cells, cytochalasin D induced oscillatory increases in cytosolic Ca(2+) ([Ca(2+)](i)); these increase were inhibited by jasplakinolide, a stabilizer of actin filaments. 3. In the IgE-sensitized RBL-2H3 cells, DNP-human serum albumin (DNP-HSA) augmented actin assembly. DNP-HSA also increased the production of IP(3), [Ca(2+)](i) and degranulation. Cytochalasin D enhanced all of these DNP HSA-induced effects. 4. In a Ca(2+)-free solution, DNP-HSA induced a transient increase in [Ca(2+)](i), and this increase was accelerated by cytochalasin D. After cessation of the DNP-HSA-induced Ca(2+) release, the re-addition of Ca(2+) induced a sustained increase in [Ca(2+)](i) through capacitative Ca(2+) entry (CCE), and this increase was enhanced by cytochalasin D. 5 The effect of cytochalasin D in enhancing the CCE activity was prevented by xestospongin C. 6. In contrast, neither the Ca(2+) release nor the CCE activation that was induced by thapsigargin was affected by cytochalasin D. 7. These results suggest that actin de-polymerization stimulates the FcepsilonRI-mediated signalling to augment the release of Ca(2+) from the endoplasmic reticulum in RBL-2H3 cells. PMID- 12110610 TI - Investigation of neurotransmission in vas deferens from alpha(2A/D)-adrenoceptor knockout mice. AB - 1. We have investigated pre- and post-junctional responsiveness in vas deferens from wild-type and alpha(2A/D)-adrenoceptor knockout mice. The response to a single stimulus was not significantly different between wild-type and knock-out mice. The isometric contraction to 10 Hz stimulation for 4 s was significantly larger in vas deferens from knockout as compared with wild-type. 2. The maximum potentiation of 10 Hz stimulation-evoked contractions by yohimbine was to 206.2+/ 38.0% of control in wild-type but to 135.8+/-13.6% of control in knockout. The alpha(2A/D)-adrenoceptor selective antagonist BRL 44408 significantly increased the 10 Hz stimulation-evoked contraction in wild-type but not knockout, and the reverse was true for the alpha(2C)-adrenoceptor selective antagonist spiroxatrine. The alpha(2B)-adrenoceptor antagonist imiloxan had no effect on the evoked contraction except at high concentrations, and only in wild-type. Following cocaine (3 microM) and BRL 44408 (1 microM), 10 Hz responses were similar in shape and maximum between wild-type and knock-out. 3. The alpha(2) adrenoceptor agonist xylazine virtually abolished the early component of the contraction to 10 Hz stimulation in the presence of nifedipine (10 microM) in vas deferens from knockout mice in a way consistent with a change of receptor subtype but without clear evidence for a reduced receptor number. However, the late component of the contraction to 10 Hz stimulation was significantly potentiated by xylazine in tissues from knock-out mice. 4. It is concluded that, although non alpha(2A/D)-adrenoceptors replace alpha(2D)-adrenoceptors in this knockout, the alpha(2)-adrenoceptor agonist and antagonist data are contradictory. The antagonist data suggest a major loss of prejunctional alpha(2)-adrenoceptors, but this is not necessarily supported by the agonist data. PMID- 12110609 TI - P2Y(1) and P2Y(2) receptors are coupled to the NO/cGMP pathway to vasodilate the rat arterial mesenteric bed. AB - 1. To assess the role of nucleotide receptors in endothelial-smooth muscle signalling, changes in perfusion pressure of the rat arterial mesenteric bed, the luminal output of nitric oxide (NO) and guanosine 3',5' cyclic monophosphate (cGMP) accumulation were measured after the perfusion of nucleotides. 2. The rank order of potency of ATP and analogues in causing relaxation of precontracted mesenteries was: 2-MeSADP=2-MeSATP>ADP>ATP=UDP=UTP>adenosine. The vasodilatation was coupled to a concentration-dependent rise in NO and cGMP production. MRS 2179 selectively blocked the 2-MeSATP-induced vasodilatation, the NO surge and the cGMP accumulation, but not the UTP or ATP vasorelaxation. 3. mRNA encoding for P2Y(1), P2Y(2) and P2Y(6) receptors, but not the P2Y(4) receptor, was detected in intact mesenteries by RT-PCR. After endothelium removal, only P2Y(6) mRNA was found. 4. Endothelium removal or blockade of NO synthase obliterated the nucleotides-induced dilatation, the NO rise and cGMP accumulation. Furthermore, 2 MeSATP, ATP, UTP and UDP contracted endothelium-denuded mesenteries, revealing additional muscular P2Y and P2X receptors. 5. Blockade of soluble guanylyl cyclase reduced the 2-MeSATP and UTP-induced vasodilatation and the accumulation of cGMP without interfering with NO production. 6. Blockade of phosphodiesterases with IBMX increased 15-20 fold the 2-MeSATP and UTP-induced rise in cGMP; sildenafil only doubled the cGMP accumulation. A linear correlation between the rise in NO and cGMP was found. 7. Endothelial P2Y(1) and P2Y(2) receptors coupled to the NO/cGMP cascade suggest that extracellular nucleotides are involved in endothelial-smooth muscle signalling. Additional muscular P2Y and P2X receptors highlight the physiology of nucleotides in vascular regulation. PMID- 12110612 TI - Pharmacological evidence for putative CCK(1) receptor heterogeneity in human colon smooth muscle. AB - 1. The pharmacology of the cholecystokinin CCK(1) receptors endogenously expressed in human gallbladder and human ascending colon smooth muscle tissue was compared using radioligand binding assays. 2. Saturation analysis of the interaction between the radiolabelled, selective CCK(1)-receptor antagonist, [(3)H]-L-364,718, and enriched gastrointestinal tissue membranes suggested the presence of multiple binding sites in human colon but not human gallbladder. 3. Competition studies, using a range of structurally diverse, CCK-receptor selective ligands provided further evidence for CCK(1) receptor heterogeneity in human colon tissue (n(H) values significantly less than unity for SR27897=0.77+/ 0.07, 2-NAP=0.73+/-0.03, YM220=0.70+/-0.09 and PD-134,308=0.83+/-0.01). Moreover, the competition data for SR27897, 2-NAP and YM220 were consistent with the interaction of these compounds at two binding sites. In contrast, in the human gallbladder assay, a single binding site model provided a good fit of the competition curve data obtained with all the CCK receptor selective compounds. 4. The data obtained are consistent with the presence of a single CCK(1) receptor binding site in the gallbladder but not in the colon. A two-site analysis of the colon data, indicated that one of the two sites was indistinguishable from that characterized in the gallbladder. The molecular basis of the apparent receptor heterogeneity in the colon remains to be established. PMID- 12110611 TI - Attenuation of morphine tolerance after antisense oligonucleotide knock-down of spinal mGluR1. AB - 1. Chronic systemic treatment of rats with morphine leads to the development of opioid tolerance. This study was designed to examine the effects of intrathecal (i.t.) infusion of a metabotropic glutamate receptor 1 (mGluR1) antisense oligonucleotide, concomitant with chronic morphine treatment, on the development of tolerance to morphine's antinociceptive effects. 2. All rats received chronic (6 day) s.c. administration of morphine to induce opioid tolerance. Additionally, rats were treated with either mGluR1 antisense (AS), missense (MIS) or artificial cerebrospinal fluid (ACSF) by i.t. infusion via chronically implanted i.t. catheters connected to osmotic mini-pumps. The effects of acute i.t. or s.c. morphine on tail-flick latencies were assessed prior to and following chronic s.c. morphine treatment for all chronic i.t. infusion groups. mGluR1 protein level in the spinal cord was determined by Western blot analysis for all treatments, assessing the efficiency of knock-down with AS treatment. 3. Acute i.t. morphine dose-dependently produced antinociception in the tail-flick test in naive rats. Systemic morphine-treated rats administered i.t. ACSF or MIS developed tolerance to i.t. morphine. Chronic i.t. infusion with mGluR1 AS significantly reduced the development of tolerance to i.t. morphine. 4. In contrast to i.t. morphine, tolerance developed to the antinociceptive effects of s.c. morphine, in all i.t. infusion groups, including the mGluR1 AS group. 5. The spinal mGluR1 protein level was dramatically decreased after mGluR1 AS infusion when compared to control animals (naive and ACSF-treated animals). 6. These findings suggest that the spinal mGluR1 is involved in the development of tolerance to the antinociceptive effects of morphine. Selective blockade of mGluR1 may be beneficial in preventing the development of opioid analgesic tolerance. PMID- 12110613 TI - Molecular modelling of the interactions of carbamazepine and a nicotinic receptor involved in the autosomal dominant nocturnal frontal lobe epilepsy. AB - 1. The normal and a mutant (S248F) human neuronal alpha4beta2 nicotinic receptors, and their interaction with the channel blocker carbamazepine (CBZ) have been modelled. The mutant, responsible for the autosomal dominant nocturnal frontal lobe epilepsy (ADNFLE), has an enhanced sensitivity to and a slower recovery from desensitization, a lower conductance, short open times, reduced calcium permeability, and is 3 fold more sensitive to CBZ, a drug used in the treatment of partial epilepsies. 2. Mutant channel properties are explained by the physicochemical properties of the two Phe248 side chains, including size and cation-pi interaction, and their dynamic behaviour. A defective mechanism of dehydration might be responsible for the reduced calcium influx. 3. Phe248 residues are the main component of CBZ binding sites in the mutant, while this is not true for Ser248 in the normal receptor. 4. A higher number of blocking binding sites and a predicted higher affinity found for CBZ in the mutant account for its differential sensitivity to CBZ. 5. Aromatic-aromatic interactions between CBZ and the two Phe248 account for the difference in affinity, which is at least 12 times higher for the mutant, depending on the method used for calculating K(i). 6. Normal vs mutant differences in K(i), enhanced by the higher number of blocking binding sites in the mutant, seem excessive compared to the differential sensitivities to CBZ experimentally found. The negative cooperativity suggested by a predicted overlapping of blocking and non-blocking binding sites gives an explanation, as overlapping is higher in the mutant. 7. For both types of receptors we found that the carbamyl group of the best blocking conformers of CBZ forms hydrogen bonds with serine residues, which may explain the fundamental role of that moiety for this molecule to act as antiepileptic drug. PMID- 12110614 TI - The highly selective CRF(2) receptor antagonist K41498 binds to presynaptic CRF(2) receptors in rat brain. AB - 1. Novel analogues of antisauvagine-30 (aSvg-30), a selective antagonist for CRF(2) receptors, have been synthesized and characterized in vitro and in vivo. 2. The analogues were tested for their ability to compete for [(125)I-Tyr(0)]Svg binding and to inhibit Svg-stimulated adenylate cyclase activity in human embryonic kidney (HEK) 293 cells, permanently transfected with cDNA coding for the human CRF(1) (hCRF(1)), hCRF(2alpha) and hCRF(2beta) receptor. One analogue [D-Phe(11), His(12), Nle(17)]Svg(11-40), named K41498, showed high affinity binding to hCRF(2alpha) (K(i)=0.66+/-0.03 nM) and hCRF(2beta) (K(i)=0.62+/-0.01 nM) but not the hCRF(1) receptor (k(i)=425+50 nM) and decreased Svg-stimulated cAMP accumulation in hCRF(2) expressing cells. In conscious Wistar-Kyoto rats, K41498 (1.84 microg, i.v.) antagonized the hypotensive response to systemic urocortin (1.4 microg, i.v.), but did not block the pressor response to centrally administered urocortin (2.35 microg, i.c.v.). 3. K41498 was subsequently radio iodinated, and in autoradiographic studies, specific (sensitive to rat urocortin, astressin and aSvg30, but insensitive to antalarmin) binding of (125)I-K41498 (100 pM) was detected in the heart and in selected brain regions including the nucleus tractus solitarius (NTS), spinal trigeminal nucleus, lateral septum and around the anterior and middle cerebral arteries. 4. Following unilateral nodose ganglionectomy, binding of (125)I-K41498 was reduced by 65% in the ipsilateral NTS, indicative of presynaptic CRF(2) receptors on vagal afferent terminals. 5. These data demonstrate that K41498 is a useful tool to study native CRF(2) receptors in the brain and periphery. PMID- 12110617 TI - Prevention of experimental carotid and coronary artery thrombosis by the glycoprotein IIb/IIIa receptor antagonist CRL42796. AB - 1. The antithrombotic effect of the glycoprotein IIb/IIIa receptor antagonist, CRL42796, was examined in canine models of carotid and coronary artery thrombosis. 2. In the carotid artery thrombosis model, occlusion occurred in all control vessels (time to thrombosis 47.6+/-8.9 min). After treatment with low dose CRL42796 (15 microg kg(-1) loading dose +0.31 microg kg(-1) min(-1) i.v.), two of five vessels occluded. Time to thrombosis increased significantly to 155.2+/-23.1 min. When the drug infusion was increased (0.69 microg kg(-1) min( 1)), each of five vessels remained patent. 3. Ex vivo platelet aggregation in response to arachidonic acid (AA) and ADP was examined in platelet rich plasma (PRP) prepared from citrate or heparin anticoagulated blood. CRL42796 reduced platelet reactivity at low and high doses in PRP from citrate anticoagulated blood. However, in PRP from heparin anticoagulated blood, only the higher infusion dose produced a significant reduction in ex vivo platelet responses. 4. A combination of oral aspirin (4.6 mg kg(-1) -41, -17 h) and the low infusion dose of CRL42796 did not produce an additional benefit beyond that provided by CRL42796 alone. 5. Coronary artery thrombosis was inhibited in four of five vessels treated with the lower infusion dose of CRL42796 and in five of five vessels treated with the higher infusion. Time to thrombosis increased with both doses (Control, 90.8+/-10.4 min; low dose, 165.8+/-14.2 min; high dose, >180.0+/ 0 min). 6. The results indicate that CRL42796 is an effective in vivo antithrombotic agent against experimentally-induced carotid and coronary artery thrombosis. PMID- 12110615 TI - Protective effects of M40403, a selective superoxide dismutase mimetic, in myocardial ischaemia and reperfusion injury in vivo. AB - 1. Myocardial injury caused by ischaemia and reperfusion comes from multiple pathogenic events, including endothelial damage, neutrophil extravasation into tissue, mast cell activation, and peroxidation of cell membrane lipids. These events are followed by myocardial cell alterations resulting eventually in cell necrosis. An enhanced formation of reactive oxygen species is widely accepted as a stimulus for tissue destruction and cardiac failure. 2. In this study, we have investigated the cardioprotective effects of M40403 in myocardial ischaemia reperfusion injury. M40403 is a low molecular weight, synthetic manganese containing superoxide dismutase mimetic (SODm) that selectively removes superoxide anion. Ischaemia was induced in rat hearts in vivo by ligating the left anterior descending coronary artery. Thirty minutes after the induction of ischaemia, the ligature was removed and reperfusion allowed to occur for at least 60 min. M40403 (0.1-1 mg kg(-1)) was given intravenously 15 min before ischaemia. 3. The results obtained in this study showed that M40403 significantly reduced the extent of myocardial damage, mast cell degranulation and the incidence of ventricular arrhythmias. Furthermore, M40403 significantly attenuated, in a dose dependent manner, neutrophil infiltration in the myocardium as well as the associated induction of lipid peroxidation. Calcium overload seen post reperfusion of the ischaemic myocardium was also reduced by M40403. 4. Immunohistochemical analysis for nitrotyrosine revealed a positive staining in cardiac tissue taken after reperfusion: this was attenuated by M40403. Moreover reperfused cardiac tissue sections showed positive staining for P-selectin and for anti-intercellular adhesion molecule (ICAM-1) in the vascular endothelial cells. M40403 treatment markedly reduced the intensity and degree of P-selectin and ICAM-1 in these tissues. No staining for nitrotyrosine, P-selectin or ICAM-1 was found in cardiac tissue taken at the end of the ischaemic period. 5. Overall, M40403 treatment reduced the morphological signs of myocardial cell injury and significantly improved survival. 6. Taken together, these results clearly indicate that M40403 treatment exerts a protective effect against ischaemia reperfusion-induced myocardial injury, supporting a key role for superoxide anion in reperfusion injuries. This suggests that synthetic enzymes of SOD such as M40403, offer a novel therapeutic approach for the treatment of ischaemic heart disease where superoxide anion plays a dominant role. PMID- 12110616 TI - K+-induced hyperpolarization in rat mesenteric artery: identification, localization and role of Na+/K+-ATPases. AB - 1. Mechanisms underlying K(+)-induced hyperpolarizations in the presence and absence of phenylephrine were investigated in endothelium-denuded rat mesenteric arteries (for all mean values, n=4). 2. Myocyte resting membrane potential (m.p.) was -58.8+/-0.8 mV. Application of 5 mM KCl produced similar hyperpolarizations in the absence (17.6+/-0.7 mV) or presence (15.8+/-1.0 mV) of 500 nM ouabain. In the presence of ouabain +30 microM barium, hyperpolarization to 5 mM KCl was essentially abolished. 3. In the presence of 10 microM phenylephrine (m.p. 33.7+/-3 mV), repolarization to 5 mM KCl did not occur in the presence or absence of 4-aminopyridine but was restored (-26.9+/-1.8 mV) on addition of iberiotoxin (100 nM). Under these conditions the K+-induced repolarization was insensitive to barium (30 microM) but abolished by 500 nM ouabain alone. 4. In the presence of phenylephrine + iberiotoxin the hyperpolarization to 5 mM K(+) was inhibited in the additional presence of 300 nM levcromakalim, an action which was reversed by 10 microM glibenclamide. 5. RT-PCR, Western blotting and immunohistochemical techniques collectively showed the presence of alpha(1)-, alpha(2)- and alpha(3) subunits of Na(+)/K(+)-ATPase in the myocytes. 6. In K(+)-free solution, re introduction of K(+) (to 4.6 mM) hyperpolarized myocytes by 20.9+/-0.5 mV, an effect unchanged by 500 nM ouabain but abolished by 500 microM ouabain. 7. We conclude that under basal conditions, Na(+)/K(+)-ATPases containing alpha(2)- and/or alpha(3)-subunits are partially responsible for the observed K(+)-induced effects. The opening of myocyte K(+) channels (by levcromakalim or phenylephrine) creates a 'K(+) cloud' around the cells which fully activates Na(+)/K(+)-ATPase and thereby abolishes further responses to [K(+)](o) elevation. PMID- 12110618 TI - Different actions of protein kinase C isoforms alpha and epsilon on gastric acid secretion. AB - 1. The phorbol ester TPA, an activator of protein kinase C (PKC), inhibits cholinergic stimulation of gastric acid secretion but increases basal H(+) secretion. 2. Since these contradictory findings suggest the action of different PKC isozymes we analysed the role of calcium-dependent PKC-alpha, and calcium independent PKC-epsilon in gastric acid secretion. 3. Inhibition of PKC-alpha by the indolocarbazole Go 6976 revealed that about 28% of carbachol-induced acid secretion was inhibited by PKC-alpha. In the presence of Go 6976 approximately 64% of the carbachol-induced signal transduction is mediated by Ca(2+)/calmodulin dependent protein kinase II (CaMKII), and 14% is conveyed by PKC-epsilon as deduced from the inhibition with the bisindolylmaleimide Ro 31-8220. 4. Inhibition of carbachol-induced acid secretion by TPA was accompanied by a decrease in CaMKII activity. 5. The stimulation of basal acid secretion by TPA was biphasic with a peak at a very low concentration (10 pM), resulting in an activation of the calcium-sensor CaMKII. The activation was determined with a phosphospecific polyclonal antibody against active CaMKII. The TPA-induced increase of H(+) secretion was sensitive to the cell-permeable Ca(2+)-chelator BAPTA/AM, Ro 31-8220, and the CaMKII-inhibitor KN-62, but not to Go 6976. 6. Since TPA induced the translocation of PKC-epsilon but not of PKC-alpha in resting parietal cells, PKC-epsilon seems to be at least responsible for an initial elevation of free intracellular calcium to initiate TPA-induced acid secretion. 7. Our data indicate the different roles of two PKC isoforms: PKC epsilon activation appears to facilitate cholinergic stimulation of H(+) secretion likely by increasing intracellular calcium. In contrast, PKC-alpha activation attenuates acid secretion accompanied by a down-regulation of CaMKII activity. PMID- 12110620 TI - Aristolochic acid as a probable human cancer hazard in herbal remedies: a review. AB - The old herbal drug aristolochic acid (AA), derived from Aristolochia spp., has been associated with the development of a novel nephropathy, designated aristolochic acid nephropathy (AAN), and urothelial cancer in AAN patients. There is clear evidence that the major components of the plant extract AA, aristolochic acid I (AAI) and aristolochic acid II (AAII), both nitrophenanthrene carboxylic acids, are genotoxic mutagens forming DNA adducts after metabolic activation through simple reduction of the nitro group. Several mammalian enzymes have been shown to be capable of activating both AAI and AAII in vitro and in cells. The activating metabolism has been elucidated and is consistent with the formation of a cyclic nitrenium ion with delocalized charge leading to the preferential formation of purine adducts bound to the exocyclic amino groups of deoxyadenosine and deoxyguanosine. The predominant DNA adduct in vivo, 7-(deoxyadenosin-N(6) yl)aristolactam I (dA-AAI), which is the most persistent of the adducts in target tissue, is a mutagenic lesion leading to AT-->TA transversions in vitro. This transversion mutation is found at high frequency in codon 61 of the H-ras oncogene in tumours of rodents induced by AAI, suggesting that dA-AAI might be the critical lesion in the carcinogenic process in rodents. DNA-binding studies confirmed that both AAs bind to the adenines of codon 61 in the H-ras mouse gene and preferentially to purines in the human p53 gene. In contrast, the molecular mechanism of renal interstitial fibrosis in humans after chronic administration of AA remains to be explored. However, preliminary findings suggest that DNA damage by AA is not only responsible for the tumour development but also for the destructive fibrotic process in the kidney. It is concluded that there is significant evidence that AA is a powerful nephrotoxic and carcinogenic substance with an extremely short latency period, not only in animals but also in humans. In particular, the highly similar metabolic pathway of activation and resultant DNA adducts of AA allows the extrapolation of carcinogenesis data from laboratory animals to the human situation. Therefore, all products containing botanicals known to or suspected of containing AA should be banned from the market world wide. PMID- 12110621 TI - Smoking, lung cancers and their TP53 mutations. PMID- 12110619 TI - Potent analgesic and anti-inflammatory actions of a novel thymulin-related peptide in the rat. AB - 1. The present study examines the effect of PAT (peptide analogue of thymulin) in two rat models of inflammatory hyperalgesia induced by either i.pl. (1.25 microg in 50 microl saline) or i.p. (50 microg in 100 microl) injections of endotoxin ET. 2. Pretreatment with PAT (1, 5 or 25 microg in 100 microl saline, i.p.) decreased, in a dose dependent manner, both mechanical hyperalgesia, determined by the paw pressure (PP) test and thermal hyperalgesia determined by the hot plate (HP), the paw immersion (PI) and the tail flick (TF) tests. 3. Compared to the tripeptides K(D)PT and K(D)PV, known to antagonize interleukin (IL)-1beta or IL-1beta and PGE(2) mechanisms, PAT, at lower dosages, exerted stronger anti hyperalgesic effects. 4. When compared with the effect of a steroidal (dexamethasone) and a non-steroidal (indomethacin) anti-inflammatory drugs (NSAID), PAT demonstrated equal analgesic actions. 5. Pretreatment with PAT, reduced significantly the increased concentration of IL-1beta, IL-6, TNF-alpha and NGF due to i.pl. injection of ET. 6. Injection of i.p. ET produced sickness behaviour characterized by hyperalgesia and fever. Pretreatment with PAT prevented the hyperalgesia and maintained the body temperature within the normal range and was accompanied by a down-regulation of the levels of pro-inflammatory cytokines and PGE(2) in the liver. 7. PAT, in all doses used, did not result in any evident changes in the physiological parameters or in the normal behaviour of the rats. 8. The anti-hyperalgesic and anti-inflammatory effects of PAT can be attributed, at least partially, to the down-regulation of pro-inflammatory mediators. PMID- 12110622 TI - Hunting for electrophiles that harm human DNA: Frits Sobels Award Lecture. AB - This lecture is dedicated to Frits Sobels and his farsighted vision on research directions in genetic toxicology. Some accomplishments by the author's research group in the area of cancer etiology research and pre-clinical drug safety evaluation are presented. Praziquantel, an antischistosomal drug, was found to be devoid of any genetic effects which determined the drug companies to proceed with further safety evaluation and marketing. This highly efficient life-saving drug is now in use world wide. Biomonitoring methods have been developed to quantitate carcinogens, their metabolites or DNA adducts in humans exposed environmentally and endogenously to genotoxic agents. The methods were applied in ecological and case-control studies aimed at establishing causal relationships between exposure and disease. Results from both field studies in Iran and laboratory investigations supported the hypothesis that opium use, in particular ingestion of its pyrolysates, may be a risk factor for esophageal cancer in this region, probably acting together with nutritional deficiencies and thermal injury. By applying the nitrosoproline (NPRO) test in ecological studies on esophageal cancer causation in China some support was obtained for the involvement of N nitroso compounds. In inhabitants of high risk areas endogenous nitrosamine synthesis could be markedly reduced by ingestion of vitamin C. Ultrasensitive detection methods for etheno-DNA adducts, which are formed by lipid peroxidation products resulting from increased oxidative stress, have been developed. Known cancer risk factors such as metal storage, chronic inflammatory processes and a high omega-6 PUFA fat diet increased the background level of these miscoding DNA adducts many times. They were found to increase progressively in premalignant lesions of cancer-prone tissues of humans and rodents, probably contributing to the genetic instability that drives cells to malignancy. Etheno-DNA adducts are thus promising markers to verify the efficiency of chemopreventive measures in humans. PMID- 12110623 TI - The effect of dietary restriction during development in utero on the frequency of spontaneous somatic mutations. AB - Caloric or dietary restriction is known to be protective against cancer in humans and in mice but the mechanism is uncertain. Given that somatic mutations are important in carcinogenesis, dietary restriction may act by changing mutation rates. Indeed, previous studies have shown that reductions in caloric intake during development or in adult life make mice less susceptible to high doses of mutagens. In these studies there have been hints that the spontaneous mutant frequency may also be reduced, but no significant decrease has been observed save in one study of very old mice. Since the spontaneous mutant frequency is already low, reductions from this level require the use of much larger sample sizes than usual and larger than those used in the previous studies. As pre-existing mutations cannot be eliminated, it is necessary to reduce the dietary intake over a period of time when a substantial proportion of spontaneous mutations arise in order to see an effect. To overcome such problems, the dietary restriction in this study was applied during the time of the highest mutation rate, early development, and many more than the usual number of animals were studied. SWR female mice were crossed with Muta(TM)Mouse males to obtain F(1) progeny for analysis of mutant frequency. At conception, the dams were put into two groups, one that was fed ad libitum and another which was fed 80% of the ad libitum diet. Pups were killed at birth, DNA was extracted from the whole animal and used to measure the mutant frequencies of the mice at the cII locus. Although the weights of the pups from dams whose diet was restricted were significantly less than those of the ad libitum mice (P = 0.003), the litter sizes in the two groups were approximately the same and did not differ significantly (P = 0.13). There was no significant difference in the mutant frequencies in the dietarily restricted and ad libitum groups (P = 0.43). In addition, there was no significant correlation between the weights of the pups and their mutant frequency in either the ad libitum or dietarily restricted groups (r(2) = 0.14 and r(2) = 0.024). No difference was observed in mutant frequency between the ad libitum and dietarily restricted mice from litters of the same size (P = 0.61). These results indicate that the protective effect of dietary restriction on cancer rates is not mediated by an alteration in the spontaneous rate of mutation but rather by another mechanism, such as its effect on induced mutation. PMID- 12110624 TI - Mutagenicity of two 2-phenylbenzotriazole derivatives, 2-[2-(acetylamino)-4 (diethylamino)-5-methoxyphenyl]-5-amino- 7-bromo-4-chloro-2H-benzotriazole and 2 [2-(acetylamino)-4-(diallylamino)-5-methoxyphenyl]-5-amino-7-bromo-4-chloro-2H benzotriazole and their detection in river water in Japan. AB - We recently detected five 2-phenylbenzotriazole (PBTA)-type mutagens (PBTA-1, PBTA-2, PBTA-3, PBTA-4 and PBTA-6) in concentrates from several rivers that flow in geographically different areas in Japan containing textile-related industries. On the basis of synthesis studies, these five PBTA derivatives were deduced to have originated from the corresponding dinitrophenylazo dyes, which are industrial chemicals used in textile dyeing, via reduction and chlorination. 2 [(2-Bromo-4,6-dinitrophenyl)azo]-5-(diethylamino)-4-methoxyacetanilide (Color Index name Disperse Blue 291, CAS registry no. 56548-64-2) and 2-[(2-bromo-4,6 dinitrophenyl)azo]-5-(diallylamino)-4-methoxyacetanilide (Color Index name Disperse Blue 373, CAS registry no. 51868-46-3) are used in textile dyeing and have 2-[(2-bromo-4,6-dinitrophenyl)azo]-4-methoxyacetanilide moieties in their structures, which are thought to be essential for their conversion to mutagenic PBTA derivatives. In the present study we have synthesized 2-[2-(acetyl-amino)-4 (diethylamino)-5-methoxyphenyl]-5-amino-7-bromo-4-chloro-2H-benzotriazole (PBTA 7) and 2-[2-(acetylamino)-4-(diallylamino)-5-methoxyphenyl]-5-amino-7-bromo-4 chloro-2H-benzotriazole (PBTA-8) from Disperse Blue 291 and Disperse Blue 373, respectively, by reduction with iron powder and subsequent chlorination with sodium hypochlorite. Both PBTA-7 and PBTA-8 exerted strong mutagenicity in Salmonella typhimurium TA98 and YG1024 in the presence of S9 mix (43 000 and 1 430 000 revertants/nmol for PBTA-7 and 40 700 and 2 213 000 revertants/nmol for PBTA-8 in TA98 and YG1024). To clarify whether PBTA-7 and PBTA-8 exist in the environment, water samples were collected at seven sites in six rivers flowing through two different regions where textile dyeing industries are located. All water samples were mutagenic in Salmonella typhimurium YG1024 with S9 mix and their potencies ranged from 108 000 to 1 990 000 revertants/g blue rayon. PBTA-7 and PBTA-8 were detected in water samples from both regions at levels of <0.1 101.4 ng/g blue rayon and <0.1-48.9 ng/g blue rayon, respectively. In some samples PBTA-7 and PBTA-8 could contribute up to 15% of the water mutagenicity. PMID- 12110625 TI - Mutation spectrum of 1,2-dibromo-3-chloropropane, an endocrine disruptor, in the lacI transgenic Big Blue Rat2 fibroblast cell line. AB - 1,2-Dibromo-3-chloropropane (DBCP), a soil fumigant against nematodes, has been extensively studied for genotoxicity, carcinogenicity and damage to male reproduction-related organs, as a possible endocrine disruptor. However, the precise mechanisms involved in DBCP-induced mutagenesis and carcinogenesis are as yet unknown. Thus, in this study the mutagenicity and mutation spectrum of DBCP was determined using the lacI transgenic Big Blue Rat2 fibroblast cell line. In determining the optimal concentration of DBCP in Big Blue Rat2 fibroblast cells, the 50% inhibition concentration was calculated to be 0.75 mM. When cells were exposed to DBCP concentrations of 0.21, 0.39 and 0.75 mM, the respective relative survival rates were approximately 80, 70 and 50%. The mean mutant frequencies (MFs) (x 10(-5), +/- SEM) of the medium and 1% DMSO solvent controls were determined as 6.43 +/- 0.616 and 5.28 +/- 1.086, respectively. The MFs (x 10(-5), +/- SEM) of cells exposed to 0.21, 0.39 and 0.75 mM DBCP were 8.09 +/- 1.02, 10.86 +/- 2.17 and 12.26 +/- 0.79, respectively, with a dose-dependent effect (ANOVA, P = 0.007). Moreover, MF values for the 0.75 and 0.39 mM DBCP-treated groups were statistically significant (ANOVA, P < 0.05). The majority of recovered mutations (31/40, 77.5%) were single base pair substitutions in the DBCP-induced groups. Among 31 single base pair substitutions, 25 (62.5%) occurred at G:C base pairs, while six (15%) were at A:T base pairs. The predominant mutation was G:C-->A:T transitions (16/40, 40%), followed by G:C-->T:A transversions (9/40, 22.5%). We conclude that DBCP is a possible base substitution mutagen, especially at guanine bases. PMID- 12110626 TI - Chromosomal aberration, micronucleus and Comet assays on peripheral blood lymphocytes of leprosy patients undergoing multidrug treatment. AB - To evaluate the genetic damage in leprosy patients, we carried out the alkaline Comet assay and chromosomal aberration (CA) and micronucleus (MN) tests in peripheral blood lymphocytes of 50 leprosy patients receiving multidrug treatment (MDT) and 50 healthy individuals. The Comet assay showed statistically higher mean values for length to width ratios of DNA mass (P < 0.01) and for mean frequencies of tailed cells (P < 0.001) in cells of leprosy patients than in those of controls. Similarly, the mean frequencies of micronucleated cells (per 1000 cytochalasin B-induced binucleated cells) were significantly greater (P < 0.001) in leprosy patients (19.92 +/- 2.564) than in controls (1.6 +/- 0.231). A statistically significant 10-fold increase in the frequency of CAs (11.16 +/- 0.411) was observed in leprosy patients compared with controls (1.28 +/- 0.242). In multiple regression analyses, when patients and controls were considered together, disease factor alone significantly influenced the genotoxicity markers. In the control group, age and alcohol consumption significantly influenced MN and length to width ratios and CA frequency, respectively. However, in MDT-treated leprosy patients none of the other confounding factors (sex, age, smoking and alcohol drinking) significantly affected the extent of genetic damage. PMID- 12110627 TI - Characterization of Trp(+) reversions in Escherichia coli strain WP2uvrA. AB - The Escherichia coli strain WP2uvrA is widely used in general mutagenicity screening tests because of its high sensitivity to many kinds of mutagens and it serves as a supplement to the standard Salmonella typhimurium tester strains. In contrast to Salmonella His(+) revertants, E.coli Trp(+) revertants have not been characterized at the molecular level. In this study we found that in the trpE65 allele of WP2uvrA the triplet that codes for the fourth amino acid from the N terminus of anthranilate synthetase was an ochre stop codon (TAA) instead of a glutamine codon (CAA). In spontaneous Trp(+) revertants the ochre codon had been changed to glutamine (CAA), lysine (AAA), glutamic acid (GAA), leucine (TTA), serine (TCA) or tyrosine (TAC, TAT). Since tryptophan prototrophy could also be restored by ochre suppressor mutations at the anticodon sites in the genes for tRNA(Glu) (glnU), tRNA(Lys) (lysT) and tRNA(Tyr) (tyrT, tyrU), the Trp(+) reversion system with E.coli WP2uvrA detected five types of base substitutions, A.T-->T.A, A.T-->C.G, A.T-->G.C, G.C-->A.T and G.C-->T.A. About 30-50% of Trp(+) revertants induced by N-ethyl-N'-nitro-N-nitrosoguanidine, captan and angelicin plus UVA irradiation were attributable to reversion at the trpE65 ochre locus; the others were attributable to suppressor mutations. In contrast, almost all revertants induced by N-methyl-N'-nitro-N-nitrosoguanidine, 3-chloro-4 (dichloromethyl)-5-hydroxy-2(5H)-furanone and furylfuramide were caused by suppressor mutations. Thus, the high mutagen sensitivity of WP2uvrA is due to several target sites consisting of A.T base pairs (trpE65, lysT) and G.C base pairs (glnU, tyrT, tyrU). PMID- 12110629 TI - Comparison of the computer programs DEREK and TOPKAT to predict bacterial mutagenicity. Deductive Estimate of Risk from Existing Knowledge. Toxicity Prediction by Komputer Assisted Technology. AB - The performance of two computer programs, DEREK and TOPKAT, was examined with regard to predicting the outcome of the Ames bacterial mutagenicity assay. The results of over 400 Ames tests conducted at Glaxo Wellcome (now GlaxoSmithKline) during the last 15 years on a wide variety of chemical classes were compared with the mutagenicity predictions of both computer programs. DEREK was considered concordant with the Ames assay if (i) the Ames assay was negative (not mutagenic) and no structural alerts for mutagenicity were identified or (ii) the Ames assay was positive (mutagenic) and at least one structural alert was identified. Conversely, the DEREK output was considered discordant if (i) the Ames assay was negative and any structural alert was identified or (ii) the Ames assay was positive and no structural alert was identified. The overall concordance of the DEREK program with the Ames results was 65% and the overall discordance was 35%, based on over 400 compounds. About 23% of the test molecules were outside the permissible limits of the optimum prediction space of TOPKAT. Another 4% of the compounds were either not processable or had indeterminate mutagenicity predictions; these molecules were excluded from the TOPKAT analysis. If the TOPKAT probability was (i) > or =0.7 the molecule was predicted to be mutagenic, (ii) < or =0.3 the compound was predicted to be non-mutagenic and (iii) between 0.3 and 0.7 the prediction was considered indeterminate. From over 300 acceptable predictions, the overall TOPKAT concordance was 73% and the overall discordance was 27%. While the overall concordance of the TOPKAT program was higher than DEREK, TOPKAT fared more poorly than DEREK in the critical Ames-positive category, where 60% of the compounds were incorrectly predicted by TOPKAT as negative but were mutagenic in the Ames test. For DEREK, 54% of the Ames-positive molecules had no structural alerts and were predicted to be non-mutagenic. Alternative methods of analyzing the output of the programs to increase the accuracy with Ames-positive compounds are discussed. PMID- 12110628 TI - Cell studies of the DNA bis-intercalator Delta-Delta [mu-C4(cpdppz)(2) (phen)(4)Ru(2)](4+): toxic effects and properties as a light emitting DNA probe in V79 Chinese hamster cells. AB - Coordination complexes of type [Ru(L)(3)](2+), where L is a nitrogen-containing aromatic bidentate ligand, can often be photolytically reduced, making them useful in studies of DNA- or protein-mediated electron transfer and in artifical photosynthesis model systems. Upon binding to DNA some Ru(L) complexes have been found to display strongly increased fluorescence compared with when free in solution, making those compounds interesting to test as DNA probes. Thus, they are becoming widely used in the chemistry community. Here, asynchronous cultures of V79 Chinese hamster cells were exposed to the DNA bis-intercalator Delta-Delta [mu-C4(cpdppz)(2)-(phen)(4)Ru(2)](4+) at 10(-10)-10(-4) M. The extraordinarily strong binding of the compound to DNA was the reason for testing its possible interference with DNA metabolism in intact mammalian cells. Exposure for 1 h to 10(-10)-10(-4) M did not significantly decrease DNA synthesis. Cells exposed to 10(-5) M for 27 h showed no staining of the nucleus, while DNA was stained in cells electroporated in the presence of the compound. However, the Ru dimer was probably taken up by pinocytosis, because numerous minute precipitates could be observed in the cytoplasm. Treatment for 24 h at concentrations of 10(-10)-10(-5) M did not inhibit growth, as indicated by cell density and mitotic activity. Neither did it affect chromosomal arrangements during mitosis. However, at 10(-4) M the density of cultures was reduced by approximately 45% and apoptotic cells were frequent, as opposed to mitoses. We also investigated the properties of the Ru dimer as a fluorescent DNA stain. The compound appears attractive as a red DNA stain when broad excitation in the visible range is desirable and extremely low background staining is essential. The low toxicity of the compound is a favourable trait in this context. PMID- 12110630 TI - Combined treatment with 4-(N-methyl-N-nitrosamino)-1- (3-pyridyl)-1-butanone and dibutyl phthalate enhances ozone-induced genotoxicity in B6C3F1 mice. AB - Potential toxicological interactions of 4-(N-methyl-N-nitrosamino)-1-(3-pyridyl) 1-butanone (NNK) and/or dibutyl phthalate (DBP) with ozone were investigated. Male and female B6C3F1 mice were exposed to ozone (0.5 p.p.m.), NNK (1.0 mg/kg), DBP (5000 p.p.m.) and different combinations of these toxicants 6 h/day for 16, 32 and 52 weeks. Two cytogenetic end-points, determined by the chromosomal aberration (CA) and supravital micronucleus (SMN) assays, were investigated in vivo. Our results show that all treated groups of both sexes showed genotoxic effects when compared with the control group. Additive and/or synergistic responses were observed in the CA assay for all test periods when mice of both sexes were exposed to ozone and NNK, ozone and DBP and the combination of ozone, NNK and DBP. In the SMN assay, additive interactions were noted for both sexes in the 16 and 32 week studies, similar to the results with the CA assay. All combination groups of both sexes showed synergistic interactions in the 52 week study. The results indicate that combined exposure to ozone, NNK and DBP in both sexes of mice has enhanced genotoxic effects compared with exposure to ozone alone. PMID- 12110631 TI - Elimination of micronucleated cells by apoptosis after treatment with inhibitors of microtubules. AB - Two major mechanisms responsible for chromosome segregation errors are non disjunction and chromosome loss, both leading to aneuploidy. Previous studies in our laboratory showed the existence of thresholds for the induction of chromosome non-disjunction and chromosome loss and the induction of apoptosis by microtubule inhibitors. From a mechanistic point of view one can expect that apoptosis contributes to the elimination of cells with premutagenic/mutagenic lesions. If aneuploid cells were eliminated by the induction of apoptosis below the threshold concentrations for chromosome loss and non-disjunction, the defined thresholds would not be applicable to cells unable to undergo apoptosis. The aim of this study was to investigate whether apoptosis was induced directly or indirectly as a response to aberrant chromosome segregation below the thresholds for the induction of chromosome loss and non-disjunction, as previously defined by us. Therefore, human lymphocytes were exposed in vitro to five concentrations of nocodazole and five concentrations of carbendazim representing the threshold concentrations for chromosome non-disjunction and chromosome loss, two concentrations below the lowest threshold and one concentration between the two threshold values. After 48 h exposure to the aneugens, induction of apoptosis was analysed by the annexin-V test. The frequencies of chromosome non-disjunction and chromosome loss were estimated in cytokinesis-blocked human lymphocytes in combination with FISH; this methodology was applied to whole cell cultures as well as to apoptotic and viable cell fractions obtained using magnetic annexin microbead cell sorting. Our results suggest that elimination of aneuploid cells does occur. However, the efficiency of disappearance of micronucleated cells is higher than for cells presenting chromosome non-disjunction. The correlation found between early apoptotic events and micronucleus formation could account, at least in part, for the specific elimination of aneuploid cells. PMID- 12110632 TI - Genotoxic effects of oestrogens in breast cells detected by the micronucleus assay and the Comet assay. AB - Cumulative exposure to oestrogen has been linked to increased risk of breast cancer. Whilst oestrogens induce cancers in rodent bioassays it is unclear whether the mechanisms involved are genotoxic and/or epigenetic. The cytokinesis block micronucleus (CBMN) and the alkaline single cell-gel electrophoresis 'Comet' assays were used to examine MCF-7 cells for chromosomal damage and DNA single-strand breaks (SSBs), respectively. The comet-forming activities of oestrogens were also tested in a 72 h primary culture of cells isolated from freshly expressed breast milk. Micronuclei (MN) were scored in 500 binucleate cells per treatment and SSBs were quantified by comet tail length (CTL) (microm). Effects on mitotic rate (per cent binucleate cells) and cell viability (per cent plating efficiency) were also assessed. beta-Oestradiol, oestrone and oestriol were tested for genotoxicity in the 10(-10)-10(-4) M and 10(-10)-10(-2) M concentration ranges in the CBMN and Comet assays, respectively. Beta-Oestradiol, following 24 h treatment but not 120 h treatment, induced increases (up to 3 fold) in MN at a concentration of 10(-9) M. Oestrone induced dose-related increases in MN (up to 5-fold) following both 24 and 120 h treatment, whereas oestriol appeared not to induce MN. All three oestrogens induced dose-related increases in per cent binucleate cells suggesting that they enhance mitotic rate. In the Comet assay both beta-oestradiol and oestrone induced dose-related increases in SSBs (up to 7-fold over control CTL) and were significantly comet forming (P < 0.0001) at concentrations as low as 10(-9) and 10(-8) M, respectively, whereas oestriol was less genotoxic. All three oestrogens were significantly comet-forming (P < 0.0001) in a primary culture of breast milk cells, suggesting that they can damage the target cells from which breast cancers may eventually arise. PMID- 12110633 TI - Comparison of kinetics of induction of DNA adducts and gene mutations by a nitrofuran compound, 7-methoxy-2-nitronaphtho[2,1-b]furan (R7000), in the caecum and small intestine of Big Blue mice. AB - In previous experiments, i.p. injection of the 5 nitronaphthofuran derivative 7 methoxy-2-nitronaphtho[2,1-b]furan (R7000) to lacI transgenic Big Blue mice led to an increase in the mutant frequency (MF), especially in the caecum and the small intestine. In the present work, the in vivo genotoxicity of R7000 in these two target organs was further investigated. Big Blue mice were treated with a single daily i.p. injection of R7000 of 0.05-0.5 mg/day for five consecutive days and killed 28 days later. These treatments led to significant increases in MF of 1.8-, 3- and 5.4-fold at 0.1, 0.2 and 0.5 mg/day R7000, respectively, in the small intestine. In the caecum, a mutagenic effect, of 4.5-fold, was only observed at the highest dose. DNA adduct formation and MFs resulting from R7000 were also analysed in parallel at various times after the last injection. R7000 led to 14 and seven different nucleotide modifications in the caecum and small intestine, respectively. Three hours after the final injection the level of induced DNA adducts was 10 times higher in the caecum than in the small intestine. From 3 h to 5 days after the final injection, 93 and 58% of DNA adducts disappeared in the caecum and small intestine, respectively. The resulting MF values were similar when comparing the two organs. Analysis of the R7000-induced mutation spectrum in the caecum showed that single G:C and large, > or =3 bp deletions and GC-->CG transversions were the first induced mutations at the end of the treatment. Fifteen days later, the R7000 mutation specificity characteristics already reported in Escherichia coli and in the small intestine of Big Blue mice were evident in the caecum, with the two major events being GC- >TA transversions and deletions of one G:C base pair. In both organs, a relationship between the decrease in R7000-DNA adducts and induction of MF was evident. However, the efficiency of this compound in damaging DNA was not correlated with the capacity of DNA lesions to lead to mutations. Some discrepancies in the R7000 genotoxic effects between the two organs were observed, which may be attributable to differences in the metabolic activation pathway of the compound, as well as to DNA repair proficiency in each tissue. PMID- 12110634 TI - Sensitivity of superselective arteriography for small hepatocellular carcinoma compared with proximal arteriography and computed tomography during superselective arteriography. AB - PURPOSE: To obtain successful arterial chemoembolization for hepatocellular carcinoma (HCC), we evaluated the sensitivity of proximal arteriography, superselective (subsegmental or more distal branch) arteriography and computed tomography (CT) during superselective arteriography and assessed the method for the injection of contrast medium. METHODS: Thirty-two patients with 38 HCCs (< or = 5 cm) with a mean diameter of 2.2 cm underwent digital subtraction arteriography of proximal and superselective arteriography. In addition, they also had helical CT during superselective arteriography. The contrast medium was injected with a mechanical injector (n = 6 lesions) or by hand (n = 32) for superselective arteriography and CT during superselective arteriography. The amount of contrast medium used for superselective arteriography and CT during superselective arteriography with the mechanical injector was 3.5 times and 9 times that with manual injection, respectively. RESULTS: Overall, 31 lesions (81.6%) were detected by proximal arteriography, 25 (65.8%) by superselective arteriography and 35 (92.1%) by CT during superselective arteriography. CT during superselective arteriography was significantly superior to superselective arteriography (P = 0.005). In both studies, manual injection of contrast medium had a significantly higher sensitivity than mechanical injection (P = 0.013). CONCLUSION: To detect small HCC, CT during superselective arteriography showed significantly higher sensitivity than superselective arteriography. Manual injection of contrast medium was significantly superior to mechanical injection. Therefore, manual injection CT during superselective arteriography is recommended for accurately targeted, transarterial chemoembolization therapy. PMID- 12110635 TI - Increased frequency of p53 mutation in sporadic colorectal cancer from cigarette smokers. AB - BACKGROUND: Cigarette smoking has been shown to increase the risk of colorectal cancer. However, the relation between smoking and genetic alterations has not been clarified in this type of cancer. METHODS: Mutations of p53, APC, beta catenin and K-ras-2 genes were analyzed in colorectal carcinomas from 28 smokers and 33 non-smokers. Frequencies and types of mutations were compared between smokers and non-smokers. RESULTS: The frequency of carcinomas with p53 mutation was higher in smokers (20/28, 71%) than in non-smokers (15/33, 45%) (P = 0.037). The common type of mutation was single-base substitution including G:C to A:T transition in both groups (68% in smokers and 67% in non-smokers). With respect to G:C to A:T transitions, mutation at CpG sites was less frequent in smokers (9/15, 60%) than in non-smokers (10/10, 100%), whereas mutation at non-CpG sites was more frequent in smokers (6/16, 40%) than in non-smokers (0/10, 0%) (P = 0.028). The frequency of APC mutation was not significantly different between smokers (14/28, 50%) and non-smokers (15/33, 45%). No beta-catenin mutation was detected in carcinomas from smokers. K-ras-2 mutation occurred in smokers at a similar frequency (9/28, 32%) to that in non-smokers (13/33, 39%). Concerning pathological aspects, Dukes' A carcinomas were less frequent in smokers (11%) than in non-smokers (33%), whereas Dukes' D carcinomas were more frequent in smokers (25%) than in non-smokers (15%). CONCLUSION: The present results suggest that an increased frequency of p53 gene mutation, including G:C to A:T transitions at non-CpG sites, is associated with an increased risk of colorectal carcinogenesis in cigarette smokers. PMID- 12110636 TI - Disposition of radioiodine (131)I therapy for thyroid carcinoma in a patient with severely impaired renal function on chronic dialysis: a case report. AB - The aim of this study was to analyze the disposition of radioiodine used for the ablation of thyroid remnants after radical surgery for a differentiated thyroid carcinoma in a patient on chronic hemodialysis in order to deliver the optimal (131)I dose to improve the healing rate in these rare cases and to serve as a useful reference to other health care professionals who might face a similar dilemma. A 50 mCi dose of (131)I was administered orally and dialysis sessions were performed 24, 72 and 144 h after therapy. Patient effluent dialyzate waste samples were collected and blood radioactivity analyses were performed at each dialysis session. The (131)I disposition half-life was 2.7 +/- 0.8 h. The amounts of remnant radioactivity in total body patient were 58.7, 38.9 and 27.1%, respectively, after each of the three dialysis sessions and the effective period calculated was 1.4 days. The extents of water purification in blood were 69.7, 47.9 and 22.7% at the beginning of each dialysis and 37.7, 42.8 and 18.1% at the end of each dialysis. Effective periods of radioiodine for thyroid cancer in a patient on hemodialysis resulted in rapid iodine clearance, thereby reducing the effective radiation dose and promoting the need to use larger treatment doses. Hemodialysis was safe and effective during treatment with radioiodine. PMID- 12110637 TI - A long-term survivor of intrahepatic cholangiocarcinoma with lymph node metastasis: a case report. AB - We describe a case of intrahepatic cholangiocarcinoma (ICC) in a 50-year-old man. A well-defined, hypoechoic tumor, 3.5 cm in greatest diameter, was detected in the left medial segment of the liver with ultrasonography. Celiac angiography showed staining at the same location. Computed tomography revealed lymph node swelling around the head of the pancreas. On October 10, 1993, the patient underwent partial hepatectomy with pancreatoduodenectomy and lymph node dissection around the hepatoduodenal ligament and along the common hepatic artery. Postoperative histopathological examination showed a moderately differentiated tubular adenocarcinoma which had metastasized to the dissected lymph nodes at the posterior surface of the head of the pancreas and at the root of the middle colic artery. Eight years after surgery, the patient is alive and well with no sign of recurrence. Immunohistochemical staining showed ductal-type mucin core protein-1 expression in the tumor, which indicates more favorable survival after surgery. Patients with ICC and lymph node metastasis are considered to have poor prognosis; however, further study of the characteristics of ICC with lymph node metastasis is needed. PMID- 12110638 TI - Aggressive undifferentiated carcinoma of unknown primary site complicated by lactic acidosis after bleeding: a case report. AB - Undifferentiated carcinoma of unknown primary site complicated by lactic acidosis has not been documented. We describe a young female with undifferentiated carcinoma of unknown primary site manifested by widespread lymph node and hepatic infiltration, hyperuricemia and very high levels of lactate dehydrogenase. She developed lactic acidosis suddenly after an episode of bleeding following nasal biopsy. The bleeding episode is likely to have caused subclinical hepatic hypoperfusion and hypoxemia, thereby aggravating lactate overproduction by tumor cells and clearance impairment due to diffuse hepatic infiltration to result in rapidly fatal acidosis before cytotoxic agents could be instituted. Although uncommon, when a critical event occurs in aggressive malignancies with massive hepatic involvement, the clinician should be alert for the development of lactic acidosis because the life-threatening metabolic complication is best avoided by prompt and effective cytoreduction therapy. PMID- 12110640 TI - Concurrent mutations of K-ras oncogene at codons 12 and 22 in colon cancer. AB - K-ras mutation is the most common oncogenic alteration in various human cancers including colorectal carcinomas. Point mutations have the potential to activate the K-ras gene if they occur in the critical coding sequences. Almost all of these mutations have been localized in codons 12, 13 and 61. We report a case of colon cancer presenting point mutations at both codons 12 and 22 of the K-ras gene. PCR-SSCP and subsequent sequencing revealed that GGT (glycine, wild-type) to AGT (serine) substitution at codon 12 and CAG (glutamine, wild-type) to CGG (arginine) substitution at codon 22 occurred in the same allele. PMID- 12110639 TI - A novel germline mutation of hMLH1 in a patient with hereditary non-polyposis colorectal cancer. AB - DNA mismatch repair genes, hMLH1 and hMSH2, assigned on chromosome 3p21-23 and 2p21-22 are involved in hereditary non-polyposis colorectal cancer (HNPCC). The heterozygous carrier of the mutated allele results in a mutator phenotype and accelerating tumorigenesis, which especially causes carcinomas in the gastrointestinal and genitourinary tracts. We screened germline mutations of mismatch repair genes hMLH1 and hMSH2 in a patient with multiple primary neoplasms (multiple stomach cancers, colon cancer and brain tumor) in a cancer clustered HNPCC family. Screening by long RT-PCR from the RNA extracted from puromycin-treated heparinized blood showed skipping of the exon 2 in hMLH1. The analysis of the genomic DNA showed a GT deletion in the splice-donor site of the exon 2, which is compatible with the splicing variant detected by long RT-PCR analysis. This is a novel germline mutation that has not been reported previously. PMID- 12110641 TI - Accuracy in estimating tumor extension according to mammographic subtypes in patients with ductal carcinoma in situ. AB - BACKGROUND: The association between subtypes of mammographic findings and histopathological tumor extension in patients with ductal carcinoma in situ has remained unclear. The purpose of this study was to investigate the relationship between tumor extension on mammography, by stratifying four subtypes, and histopathological tumor size in patients with ductal carcinoma in situ. METHODS: This study was performed on 109 breasts with ductal carcinoma in situ. They were treated by mastectomy at our Hospital between January 1990 and December 1999. Findings on mammography were categorized as microcalcification type, spiculated type, circumscribed type or fibrocystic-change type. The microcalcification type consisted of breasts with malignant microcalcifications, regardless of the presence or absence of tumor shadow. We analyzed the relationship between tumor size on mammography in each category and histopathological tumor size. In the breasts with palpable tumors, we compared palpated tumor size and histopathological tumor size according to the mammographic subtypes. RESULTS: There was no statistical difference between mammographic tumor size and histopathological tumor size for each mammographic subtype (microcalcification type, P = 0.60; spiculated type, P = 0.72; circumscribed type, P = 0.055). The size of the ductal carcinoma in situ in microcalcification and spiculated type was estimated approximately by mammography. However, mammography tended to overestimate the circumscribed type. In the cases of palpable tumor, we statistically underestimated the size of ductal carcinoma in situ by palpation in microcalcification and fibrocystic-change type (microcalcification type, P = 0.0001; fibrocystic-change type, P = 0.040). CONCLUSION: Mammographic categorization is useful for surgical planning of ductal carcinoma in situ, particularly when considering breast-conserving surgery. PMID- 12110642 TI - Nitric oxide levels and lipid peroxidation in plasma of patients with gastric cancer. AB - BACKGROUND: The aim was to investigate the levels of malondialdehyde and total NO(2)(-) plus NO(3)(-) marker for NO(*) generation in gastric carcinoma and to correlate their levels with the cancer stage. METHOD: The pretreatment plasma samples were obtained from 38 patients with gastric cancer (seven patients at stage II, 19 at stage III and 12 at stage IV). Nitrite (NO(2)(-)) and nitrate (NO(3)(-)) levels, the end products of nitric oxide (NO(*)), were determined in these samples. NO(2)(-) was measured by using the Griess reaction and after enzymatic conversion of NO(3)(-) into NO(2)(-) by nitrate reductase, the resultant NO(2)(-) was also measured by the same method. Malondialdehyde (MDA), a lipid peroxidation marker, was measured by the thiobarbituric acid method. RESULTS: The levels of plasma MDA, NO(*) and NO(3)(-) were significantly higher in patients with gastric cancer compared with the healthy control group. Higher levels of MDA, NO(*) and NO(3)(-) were observed as the stage of the disease increased. CONCLUSION: We found that increased NO(*) production and MDA levels were present in plasma of patients with gastric cancer. These increases can be associated with the oxidant-antioxidant status in these patients. PMID- 12110643 TI - Cyclooxygenase-2 overexpression in colorectal cancer is associated with non polypoid growth. AB - BACKGROUND: Cyclooxygenase-2 is considered to play an important role in colorectal tumorigenesis. The purpose of this study was to clarify the relationship between cyclooxygenase-2 expression and morphology in colorectal cancer. METHODS: We investigated cyclooxygenase-2 mRNA expression and type of growth (polypoid or non-polypoid according to Shimoda's classification) in 69 colorectal cancers by reverse transcription-polymerase chain reaction and histologic examination. Cyclooxygenase-2 mRNA expression was assessed as a ratio relative to cyclooxygenase-1 mRNA expression (cyclooxygenase-2 index). RESULTS: Cyclooxygenase-2 indices in normal colorectal mucosa, polypoid cancer and non polypoid cancer were 0.7 +/- 0.1, 1.0 +/- 0.1 and 1.9 +/- 0.1, respectively. The cyclooxygenase-2 index in non-polypoid cancer was significantly higher than in polypoid cancer (P = 0.0002) or normal mucosa (P < 0.0001). No difference in cyclooxygenase-2 index was found between polypoid cancer and normal mucosa (P = 0.67). CONCLUSIONS: These results suggest that cyclooxygenase-2 overexpression in colorectal cancer is associated with non-polypoid growth. PMID- 12110645 TI - Endoscope-assisted minilaparotomy (endoscopic minilaparotomy) for retroperitoneal Schwannoma: experience with three cases. AB - We have been applying endoscope-assisted minilaparotomy (endoscopic minilaparotomy) to retroperitoneal operations with favorable outcomes. Here, endoscopic minilaparotomy through a single flank incision (4-7 cm) was performed in three cases of incidentally discovered retroperitoneal Schwannoma. Resection of the tumor was successfully completed. The postoperative course was uneventful. Wound pain was mild and full oral feeding and walk were resumed the day following operation. It is concluded that endoscopic minilaparotomy is applicable to retroperitoneal Schwannoma with excellent postoperative recovery. PMID- 12110644 TI - Preoperative prediction of extracapsular tumor extension at radical retropubic prostatectomy in Taiwanese patients with T1c prostate cancer. AB - BACKGROUND: To find a predictor for extracapsular tumor extension at radical retropubic prostatectomy (RRP) in Taiwanese patients with stage T1c prostate cancer (PC), preoperative transrectal sonoguiding prostate biopsy outcomes and clinicopathological data obtained from these patients were reviewed. METHODS: Fifty-five consecutive men who underwent radical retropubic prostatectomy for stage T1c PC were included. Preoperative sextant needle biopsies of the prostate were performed and whole-mount prostatectomy specimens were processed. The pathological end point was tumor capsular perforation extending entirely through the prostate capsule. Preoperative prostate-specific antigen (PSA), free-to-total PSA ratio, prostate volume, PSA density, Gleason score, number of positive biopsy cores, percentage cancer of sextant biopsies, percentage cancer of one lobe and percentage cancer of one core were analyzed for their ability to predict extracapsular tumor extension at RRP. RESULTS: Eighteen of the 55 specimens showed evidence of tumor capsular perforation. Those with extracapsular tumor extension (ECE) had higher PSA than organ-confined disease (OCD) (18.4 vs 8.3 ng/ml, P < 0.01). The ECE had a higher PSA density than OCD (0.556 vs 0.226, P < 0.01). The percentage of cancer in biopsies, percentage cancer of one lobe and percentage cancer of one core were all higher in ECE than OCD (P < 0.05). The ECE had a higher biopsy Gleason score than OCD (5.6 vs 4.5, P < 0.01). CONCLUSIONS: The four strongest predictors for extracapsular tumor extension of patients with T1c PC were PSA density >or=0.35, biopsy Gleason score >or=6, >or=20% cancer in biopsies and PSA >or=10 ng/ml. PMID- 12110646 TI - Survival trends of patients treated at the National Cancer Center Hospital, Japan, from 1962 to 1994. PMID- 12110647 TI - Normal mammalian skeletal muscle and its phenotypic plasticity. AB - Since muscle mass makes up such a high proportion of total body mass, there must have been considerable selective pressure to minimize the cost of maintenance and to maximize the functionality of muscle tissue for all species. Phenotypic plasticity of muscle tissue allows the species blueprint of muscle tissue to be modified to accommodate specific demands experienced by animals over their lifetime. In this review, we report the scaling of muscle structural compartments in a set of mammals spanning five orders of magnitude (17 g woodmice to 450 kg horses and steers). Muscle mass, muscle myofibrillar volume and sarcoplasmic space were found to represent similar relative quantities in all species studies (scaling factor close to unity). Mitochondrial volumes were found to be systematically smaller in larger animals (scaling factor 0.91) and closely related to the scaling of (O(2)max) (0.92) and were tracked by the scaling of total capillary length (0.95). In this set of species, we therefore found that maximal metabolic rate and supporting structures did not scale to the 0.75 power of body mass as generally suggested. Muscle phenotypic plasticity is reasonably well characterized on a structural and functional basis, but we still know little about the signals that cause the changes in gene expression necessary for phenotypic changes in muscle. The molecular responses of human m. vastus lateralis to endurance exercise indicate that a single bout of exercise causes specific transient transcriptional adaptations that may gradually accumulate after their translation into the (structural) modifications seen with phenotypic plasticity. Metabolic and mechanical factors are recognized candidate factors for the control of exercise-induced gene transcription in muscle. Distinct protein kinases and transcription factors emerge as possible interfaces that integrate the mechanical (MAPKs and jun/fos) and metabolic (AMPK, HIF-1alpha and PPARalpha) stimuli into enhanced gene transcription in skeletal muscle. PMID- 12110648 TI - Muscle designed for maximum short-term power output: quail flight muscle. AB - Take-off in birds at high speeds and steep angles of elevation requires a high burst power output. The mean power output of the pectoralis muscle of blue breasted quail (Coturnix chinensis) during take-off is approximately 400 W kg(-1) muscle, as determined using two independent methods. This burst power output is much higher than has been measured in any other cyclically contracting muscle. The power output of muscle is determined by the interactions between the physiological properties of the muscle, the stimulation regime imposed by the central nervous system and the details of the strain cycle, which are determined by the reciprocal interaction between the muscle properties and the environmental load. The physiological adaptations that enable a high power output to be achieved are those that allow the muscle to develop high stresses whilst shortening rapidly. These characteristics include a high myofibrillar density, rapid twitch contraction kinetics and a high maximum intrinsic velocity of shortening. In addition, several features of the strain cycle increase the power output of the quail pectoralis muscle. First, the muscle operates at a mean length shorter than the plateau of the length/force relationship. Second, the muscle length trajectory is asymmetrical, with 70 % of the cycle spent shortening. The asymmetrical cycle is expected to increase the power output substantially. Third, subtle deviations in the velocity profile improve power output compared with a simple asymmetrical cycle with constant lengthening and shortening rates. The high burst power outputs found in the flight muscles of quail and similar birds are limited to very brief efforts before fatigue occurs. This strong but short flight performance is well-suited to the rapid-response anti-predation strategy of these birds that involves a short flight coupled with a subsequent sustained escape by running. These considerations serve as a reminder that the maximum power-producing capacities of muscles need to be considered in the context of the in vivo situation within which the muscles operate. PMID- 12110649 TI - Etruscan shrew muscle: the consequences of being small. AB - The skeletal muscles of the smallest mammal, the Etruscan shrew Suncus etruscus, are functionally and structurally adapted to the requirements of an enormously high energy turnover. Isometric twitch contractions of the extensor digitorum longus (EDL) and soleus muscles are shorter than in any other mammal, allowing these muscles to contract at outstandingly high frequencies. The skeletal muscles of S. etruscus contract at up to 900 min(-1) for respiration, up to 780 min(-1) for running and up to 3500 min(-1) for shivering. All skeletal muscles investigated lack slow-twitch type I fibres and consist only of fast-twitch type IID fibres. These fibres are optimally equipped with properties enabling a high rate of almost purely oxidative metabolism: they have a small diameter, their citrate synthase activity is higher and their lactate dehydrogenase activity is lower than in the muscles of any other mammal and they have a rapid shortening velocity. Differences in isometric twitch contraction times between different muscles are, at least in part, probably due to differences in cytosolic creatine kinase activities. PMID- 12110650 TI - Supercontracting muscle: producing tension over extreme muscle lengths. AB - Muscle mechanics dictates a trade-off between the ability of a muscle to generate isometric force and its length. This intrinsic trade-off is the result of the need for overlap between thick and thin filaments upon extension of the sarcomere and of the limitations imposed by the physical interference between the thin filaments and the thick filaments with the Z-disk upon contraction. However, previously published data indicate that chameleons are able to produce a nearly constant tongue retraction force over a wide range of tongue extension lengths, made possible by the presence of supercontracting muscle in the tongue retractors. Investigation of the length/tension properties and ultrastructure of the tongue retractor in a closely related agamid lizard (Pogona vitticeps) indicates that the ability to generate tension at extreme elongation is probably a derived feature for chameleons. Whereas chameleons are unique among vertebrates in possessing supercontracting muscle, this seems to be a common phenomenon in invertebrates. However, the presence of supercontracting muscle in chameleons and in several invertebrate groups seems to be coupled to the need to generate tension over large changes in muscle length and might be a more general solution for this problem. PMID- 12110651 TI - Energy-saving mechanisms in muscle: the minimization strategy. AB - Many mechanisms reduce the cost of muscle activity. Here, we describe a set of specializations that reduce the cost of contraction in the high-frequency twitches that are used by a wide variety of animals for either sound production or flight. Minimizing the cost of these contractions means that cellular ATP production can meet ATP demand and sustain the high contractile rate. Two classes of specialization are found that minimize the contractile cost. The first class reduces the muscle work required per contraction. Light appendages such as rattles, insect limbs and membranous wings that require little work for movement are used in high-frequency contractions. The second set of specializations involves processes that minimize energy use. High-frequency muscles tend to have a lower cross-bridge content, fewer attached cross-bridges and shorter length changes per contraction. The result is low muscle-specific forces (stress), small length changes (strain) and rapid contraction times that suggest that these muscles push the lower limit of contractile function. The consequence of function at this lower extreme of contraction is to minimize the contractile cost of high frequency muscles. Thus, specializations that permit rapid contractions at a low rate of ATP use per twitch are the basis of a minimization strategy for energy saving in muscles contracting at high frequency. PMID- 12110652 TI - Weakfish sonic muscle: influence of size, temperature and season. AB - The influence of temperature, size and season on the sounds produced by the sonic muscles of the weakfish Cynoscion regalis are categorized and used to formulate a hypothesis about the mechanism of sound generation by the sonic muscle and swimbladder. Sounds produced by male weakfish occur at the time and location of spawning and have been observed in courtship in captivity. Each call includes a series of 6-10 sound pulses, and each pulse expresses a damped, 2-3 cycle acoustic waveform generated by single simultaneous twitches of the bilateral sonic muscles. The sonic muscles triple in mass during the spawning season, and this hypertrophy is initiated by rising testosterone levels that trigger increases in myofibrillar and sarcoplasmic cross-sectional area of sonic muscle fibers. In response to increasing temperature, sound pressure level (SPL), dominant frequency and repetition rate increase, and pulse duration decreases. Likewise, SPL and pulse duration increase and dominant frequency decreases with fish size. Changes in acoustic parameters with fish size suggest the possibility that drumming sounds act as an 'honest' signal of male fitness during courtship. These parameters also correlate with seasonally increasing sonic muscle mass. We hypothesize that sonic muscle twitch duration rather than the resonant frequency of the swimbladder determines dominant frequency. The brief (3.5 ms), rapidly decaying acoustic pulses reflect a low-Q, broadly tuned resonator, suggesting that dominant frequency is determined by the forced response of the swimbladder to sonic muscle contractions. The changing dominant frequency with temperature in fish of the same size further suggests that frequency is not determined by the natural frequency of the bladder because temperature is unlikely to affect resonance. Finally, dominant frequency correlates with pulse duration (reflecting muscle twitch duration), and the inverse of the period of the second cycle of acoustic energy approximates the recorded frequency. This paper demonstrates for the first time that the dominant frequency of a fish sound produced by a single muscle twitch is apparently determined by the velocity of the muscle twitch rather than the natural frequency of the swimbladder. PMID- 12110653 TI - Phylogenetic implications of the superfast myosin in extraocular muscles. AB - Extraocular muscle exhibits higher-velocity and lower-tension contractions than other vertebrate striated muscles. These distinctive physiological properties are associated with the expression of a novel extraocular myosin heavy chain (MYH). Encoded by the MYH13 gene, the extraocular myosin heavy chain is a member of the fast/developmental MYH gene cluster on human chromosome 17 and the syntenic MYH cluster on mouse chromosome 11. Comparison of cDNA sequences reveals that MYH13 also encodes the atypical MYH identified in laryngeal muscles, which have similar fast contractile properties. Comparing the MYH13 sequence with the other members of the fast/developmental cluster, the slow/cardiac MYH genes and two orphan skeletal MYH genes in the human genome provides insights into the origins of specialization in striated muscle myosins. Specifically, these studies indicate (i) that the extraocular myosin is not derived from the adult fast skeletal muscle myosins, but was the first member of the fast/developmental MYH gene cluster to diverge and specialize, (ii) that the motor and rod domains of the MYH13 have evolved under different selective pressures and (iii) that the MYH13 gene has been largely insulated from genomic events that have shaped other members of the fast/developmental cluster. In addition, phylogenetic footprinting suggests that regulation of the extraocular MYH gene is not governed primarily by myogenic factors, but by a hierarchical network of regulatory factors that relate its expression to the development of extraocular muscles. PMID- 12110654 TI - 'Superfast' or masticatory myosin and the evolution of jaw-closing muscles of vertebrates. AB - There are four fibre types in mammalian limb muscles, each expressing a different myosin isoform that finely tunes fibre mechanics and energetics for locomotion. Functional demands on jaw-closer muscles are complex and varied, and jaw muscles show considerable phylogenetic plasticity, with a repertoire for myosin expression that includes limb, developmental, alpha-cardiac and masticatory myosins. Masticatory myosin is a phylogenetically ancient motor with distinct light chains and heavy chains. It confers high maximal muscle force and power. It is highly jaw-specific in expression and is found in several orders of eutherian and marsupial mammals including carnivores, chiropterans, primates, dasyurids and diprotodonts. In exceptional species among these orders, masticatory myosin is replaced by some other isoform. Masticatory myosin is also found in reptiles and fish. It is postulated that masticatory myosin diverged early during gnathostome evolution and is expressed in primitive mammals. During mammalian evolution, mastication of food became important, and in some taxa jaw closers replaced masticatory myosin with alpha-cardiac, developmental, slow or fast limb myosins to adapt to the variety of diets and eating habits. This occurred early in some taxa (rodents, ungulates) and later in others (macropods, lesser panda, humans). The cellular basis for the uniqueness of jaw-closing muscles lies in their developmental origin. PMID- 12110655 TI - Do muscles function as adaptable locomotor springs? AB - During normal animal movements, the forces produced by the locomotor muscles may be greater than, equal to or less than the forces acting on those muscles, the consequences of which significantly affect both the maximum force produced and the energy consumed by the muscles. Lengthening (eccentric) contractions result in the greatest muscle forces at the lowest relative energetic costs. Eccentric contractions play a key role in storing elastic strain energy which, when recovered in subsequent contractions, has been shown to result in enhanced force, work or power outputs. We present data that support the concept that this ability of muscle to store and recover elastic strain energy is an adaptable property of skeletal muscle. Further, we speculate that a crucial element in that muscle spring may be the protein titin. It too seems to adapt to muscle use, and its stiffness seems to be 'tuned' to the frequency of normal muscle use. PMID- 12110656 TI - Physiological basis of temperature-dependent biogeography: trade-offs in muscle design and performance in polar ectotherms. AB - Polar, especially Antarctic, oceans host ectothermic fish and invertebrates characterized by low-to-moderate levels of motor activity; maximum performance is reduced compared with that in warmer habitats. The present review attempts to identify the trade-offs involved in adaptation to cold in the light of progress in the physiology of thermal tolerance. Recent evidence suggests that oxygen limitations and a decrease in aerobic scope are the first indications of tolerance limits at both low and high temperature extremes. The cold-induced reduction in aerobic capacity is compensated for at the cellular level by elevated mitochondrial densities, accompanied by molecular and membrane adjustments for the maintenance of muscle function. Particularly in the muscle of pelagic Antarctic fish, among notothenioids, the mitochondrial volume densities are among the highest known for vertebrates and are associated with cold compensation of aerobic metabolic pathways, a reduction in anaerobic scope, rapid recovery from exhaustive exercise and enhanced lipid stores as well as a preference for lipid catabolism characterized by high energy efficiency at high levels of ambient oxygen supply. Significant anaerobic capacity is still found at the very low end of the activity spectrum, e.g. among benthic eelpout (Zoarcideae). In contrast to the cold-adapted eurytherms of the Arctic, polar (especially Antarctic) stenotherms minimize standard metabolic rate and, as a precondition, the aerobic capacity per milligram of mitochondrial protein, thereby minimizing oxygen demand. Cost reductions are supported by the downregulation of the cost and flexibility of acid-base regulation. At maintained factorial scopes, the reduction in standard metabolic rate will cause net aerobic scope to be lower than in temperate species. Loss of contractile myofilaments and, thereby, force results from space constraints due to excessive mitochondrial proliferation. On a continuum between low and moderately high levels of muscular activity, polar fish have developed characteristics of aerobic metabolism equivalent to those of high-performance swimmers in warmer waters. However, they only reach low performance levels despite taking aerobic design to an extreme. PMID- 12110657 TI - Temperature plasticity of contractile proteins in fish muscle. AB - Three myosin heavy chain isoforms with different actin-activated Mg(2+)-ATPase activities were found in the fast skeletal muscle from carp (Cyprinus carpio) acclimated to 10 and 30 degrees C. The composition of three types of myosin heavy chain was dependent on acclimation temperature, demonstrating the presence of temperature-specific myosin isoforms in carp. Subsequently, the temperature dependence of the sliding velocity of fluorescent F-actin in myosins isolated from 10 degrees C- and 30 degrees C-acclimated carp was measured. At 8 degrees C, the filament velocity was three times higher for myosin from 10 degrees C- than from 30 degrees C-acclimated fish. Activation energies (E(a)) for the sliding velocity of F-actin were 63 and 111 kJ mol(-1) for myosins from 10 degrees C- and 30 degrees C-acclimated fish, respectively. Activation energy for actin-activated Mg(2+)-ATPase activity was 0.46 kJ mol(-1) in myosin from 10 degrees C-acclimated fish and 0.54 kJ mol(-1) in myosin from 30 degrees C-acclimated fish. The inactivation rate constant (K(D)) of Ca(2+)-ATPase was 7.5x10(-4)s(-1) at 30 degrees C for myosin from 10 degrees C-acclimated fish, which was approximately twice that for myosin from 30 degrees C-acclimated fish. It is suggested that these differences in thermostability reflect a more flexible structure of the myosin molecule in cold-acclimated carp, which results in a reduced activation enthalpy for contraction and, hence, a higher sliding velocity at low temperatures. Structural analysis of cDNAs encoding the carp myosin heavy chain demonstrated striking differences in two surface loops of myosin subfragment-1 (S1), loops 1 and 2, between the 10 degrees C and 30 degrees C types, which were predominantly expressed in carp acclimated to 10 degrees C and 30 degrees C, respectively. Chimeric myosins composed of Dictyostelium discoideum myosin backbones with loop sequences of carp S1 heavy chain isoforms demonstrated that the diversity of the loop 2 sequence of carp S1 affected the V(max) of actin activated Mg(2+)-ATPase activity. PMID- 12110658 TI - Going with the flow or life in the fast lane: contrasting mitochondrial responses to thermal change. AB - Temperature is one of the most important environmental factors affecting the physiology of animals. Seasonal fluctuations in temperature are of particular importance in aquatic ectotherms since their body temperature is in equilibrium with their environment. When an organism faces adverse environmental conditions, it can either remain active or enter into metabolic depression, adopting the strategy that maximises its fitness. Physiological responses to environmental stress occur at many different levels of organisation in an animal. Here, we focus on mitochondria, given their central importance in cellular energy metabolism. We contrast the thermal biology of skeletal muscle mitochondria from cold-active species with that of species that spend their winters in a metabolically depressed state. Specifically, we examine the modifications of mitochondrial properties during thermal/seasonal acclimation and examine mechanisms by which these modifications can arise. While compensatory responses to cold acclimation include increases in mitochondrial abundance, in the oxidative capacities of individual mitochondria and adjustments of ADP affinities, metabolic depression can reduce tissue levels of mitochondrial enzymes and mitochondrial proton leak rates. PMID- 12110659 TI - Design of heterothermic muscle in fish. AB - Among the tremendous diversity of fish, there are a small number that are considered elite in their swimming performance. These include representatives from the tunas, billfish and sharks. In addition to being elite swimmers, these fish share numerous specialized anatomical features including the structure of their swimming muscles and some form of regional endothermy, termed heterothermy. These heterothermies fall into two classes: those that maintain elevated temperatures in swimming muscles and those that have muscle-derived tissues specialized for delivering warm blood to the brain. Because these versions of heterothermy are manifest in fish whose swimming performance is considered elite, it has been parsimonious to hypothesize that heterothermy is part of an integrated high-performance design. Recognizing that the design of skeletal muscle is hierarchical, the design of heterothermic muscle in fish will be examined within a hierarchical framework. This paper will examine, in order, the specific anatomical specializations, the performance of muscle as a biomaterial and then as a dynamic mechanical structure or device - in each case looking at the extent to which heterothermy is part of an integrated high-performance design or is perhaps just a happy accident. This examination will reveal how difficult it is to make a case for the central importance of heterothermy in the design of these swimming muscle systems. PMID- 12110660 TI - Adaptive thermogenesis in hummingbirds. AB - The occurrence of non-shivering thermogenesis in birds has long been a controversial issue. Although birds are endothermic vertebrates, sharing with mammals (placental mammals and marsupials) a common ancestor, they do not possess brown adipose tissue or a similar type of tissue, unlike their mammalian counterparts. Some bird species are, however, able to withstand very low ambient temperatures (-70 degrees C) or undergo periods of heterothermia, and there is now good experimental evidence showing that non-shivering thermogenesis may indeed occur in birds under such conditions. The skeletal muscles of birds, particularly the flight muscles, occupy a significant fraction (approximately 30 %) of the total body mass, and recent results have shown that they are likely to be the main sites for non-shivering thermogenesis. The precise mechanisms involved in adaptive thermogenesis in birds are still not fully understood. The translocation of Ca(2+) between intracellular compartments and the cystosol mediated by the sarcoplasmic reticulum Ca(2+)-ATPase, uncoupled from ATP synthesis, is one mechanism whereby chemi-osmotic energy can be converted into heat, and it has been proposed as one of the possible mechanisms underlying non shivering thermogenesis in birds on the basis of data obtained mainly from ducklings acclimatized to cold conditions. The recent characterization of an uncoupling protein homolog in avian skeletal muscle and the expression of its mRNA at different stages of the torpor/rewarming cycle of hummingbirds indicate that it has the potential to function as an uncoupling protein and could play a thermogenic role during rewarming in these birds. PMID- 12110662 TI - Adaptive plasticity of skeletal muscle energetics in hibernating frogs: mitochondrial proton leak during metabolic depression. AB - The common frog (Rana temporaria) spends the coldest months of each year overwintering in ice-covered ponds where temperatures can vary from 0.5 to 4.0 degrees C. Over the course of a winter season, the animals enter progressively into a state of metabolic depression that relies almost exclusively on aerobic production of ATP. However, if aerobic metabolism is threatened, for example by increasingly hypoxic conditions, decreases in the animal's metabolic rate can reach upwards of 75% compared with the 50% decrease seen during normoxia. Under these conditions, the major proportion of the overall reduction in whole-animal metabolic rate can be accounted for by metabolic suppression of the skeletal muscle (which makes up approximately 40% of body mass). Little is known about the properties of mitochondria during prolonged periods of metabolic depression, so we have examined several aspects of mitochondrial metabolism in the skeletal muscle of frogs over periods of hibernation of up to 4 months. Mitochondria isolated from the skeletal muscle of frogs hibernating in hypoxic water show a considerable reorganisation of function compared with those isolated from normoxic submerged animals at the same temperature (3 degrees C). Both the active (state 3) and resting (state 4) respiration rates of mitochondria decrease during hypoxic, but not normoxic, hibernation. In addition, the affinity of mitochondria for oxygen increases during periods of acute hypoxic stress during normoxic hibernation as well as during long-term hibernation in hypoxic water. The decrease in mitochondrial state 4 respiration rates during hypoxic hibernation evidently occurs through a reduction in electron-transport chain activity, not through a lowered proton conductance of the mitochondrial inner membrane. The reduced aerobic capacity of frog skeletal muscle during hypoxic hibernation is accompanied by lowered activities of key enzymes of mitochondrial metabolism caused by changes in the intrinsic properties of the mitochondria. In the absence of oxygen, the mitochondrial F(1)F(o)-ATPase (the ATP synthase) begins to run backwards as it actively pumps protons from the matrix in an attempt to maintain the mitochondrial membrane potential. At this time, the ATP synthase functions as an ATPase to preserve a certain proton-motive force. Frogs limit ATP wastage during anoxia by a profound inhibition of the ATP synthase. Taken together, our studies show that protonmotive force is lowered aerobically by restricting electron supply and during anoxia by restricting mitochondrial ATPase activity. PMID- 12110661 TI - UCP2 and UCP3 in muscle controlling body metabolism. AB - The uncoupling protein-1 (UCP1) homologues UCP2 and UCP3 are able to uncouple ATP production from mitochondrial respiration, thereby dissipating energy as heat and affecting energy metabolism efficiency. In contrast to UCP1, which plays an important role in adaptive thermogenesis, UCP2 and UCP3 do not have a primary role in the regulation of energy metabolism. UCP2, which is expressed in a wide variety of tissues, including white adipose tissue, skeletal muscle and tissues of the immune system, has been suggested to affect the production of reactive oxygen species. UCP2 has also been suggested to regulate the [ATP]/[ADP] ratio and was recently shown to influence insulin secretion in the beta-cells of the pancreas. UCP3, in contrast, is expressed predominantly in skeletal muscle and has been associated with whole-body energy metabolism. However, the primary function of UCP3 is not the regulation of energy metabolism. For example, fasting, a condition attenuating energy expenditure, upregulates UCP3 expression. Moreover, UCP3-knockout mice have a normal metabolic rate. The exact function of UCP3 therefore remains to be elucidated, but putative roles for UCP3 include involvement in the regulation of ROS, in mitochondrial fatty acid transport and in the regulation of glucose metabolism in skeletal muscle. Whatever the primary function of these novel uncoupling proteins, a secondary effect via uncoupling might allow them to influence (but not to regulate) energy metabolism, which would be consistent with the observations from linkage and association studies. Therefore, UCP2 and UCP3 remain interesting targets for pharmacological upregulation in the treatment of obesity and diabetes. PMID- 12110663 TI - Maintaining muscle mass during extended disuse: aestivating frogs as a model species. AB - Prolonged muscle disuse in vertebrates can lead to a pathological change resulting in muscle wasting and a loss of muscle strength. In this paper, we review muscle disuse atrophy in the vertebrates and examine the factors that influence the magnitude of the atrophic response during extended periods of inactivity, both artificially imposed (e.g. limb immobilisation) and naturally occurring, such as the quiescence associated with dormancy (e.g. hibernation and aestivation). The severity of muscle atrophy is positively correlated with mass specific metabolic rate, and the metabolic depression that occurs during dormancy would appear to have a protective role, reducing or preventing muscle atrophy despite periods of inactivity lasting 6-9 months. In the light of these findings, the role of reactive oxygen species and antioxidants during muscle disuse is emphasised. PMID- 12110664 TI - Thermal plasticity of skeletal muscle phenotype in ectothermic vertebrates and its significance for locomotory behaviour. AB - Seasonal cooling can modify the thermal preferenda of ectothermic vertebrates and elicit a variety of physiological responses ranging from winter dormancy to an acclimation response that partially compensates for the effects of low temperature on activity. Partial compensation of activity levels is particularly common in aquatic species for which seasonal temperature changes provide a stable cue for initiating the response. Thermal plasticity of locomotory performance has evolved independently on numerous occasions, and there is considerable phylogenetic diversity with respect to the mechanisms at the physiological and molecular levels. In teleosts, neuromuscular variables that can be modified include the duration of motor nerve stimulation, muscle activation and relaxation times, maximum force and unloaded shortening velocity (V(max)), although not all are modified in every species. Thermal plasticity in V(max) has been associated with changes in myosin ATPase activity and myosin heavy chain (MyHC) composition and/or with a change in the ratio of myosin light chain isoforms. In common carp (Cyprinus carpio), there are continuous changes in phenotype with acclimation temperature at lower levels of organisation, such as MyHC composition and V(max), but a distinct threshold for an effect in terms of locomotory performance. Thus, there is no simple relationship between whole-animal performance and muscle phenotype. The nature and magnitude of temperature acclimation responses also vary during ontogeny. For example, common carp acquire the ability to modify MyHC composition with changes in acclimation temperature during the juvenile stage. In contrast, the thermal plasticity of swimming performance observed in tadpoles of the frog Limnodynastes peronii is lost in the terrestrial adult stage. Although it is often assumed that the adjustments in locomotory performance associated with temperature acclimation enhance fitness, this has rarely been tested experimentally. Truly integrative studies of temperature acclimation are scarce, and few studies have considered both sensory and motor function in evaluating behavioural responses. Developmental plasticity is a special case of a temperature acclimation response that can lead to temporary or permanent changes in morphology and/or physiological characteristics that affect locomotory performance. PMID- 12110665 TI - A role for cyclic AMP response element-binding protein (CREB) but not the highly similar ATF-2 protein in sterol regulation of the promoter for 3-hydroxy-3 methylglutaryl coenzyme A reductase. AB - Sterol regulatory element-binding proteins (SREBPs) activate promoters for key genes of metabolism to keep pace with the cellular demand for lipids. In each SREBP-regulated promoter, at least one ubiquitous co-regulatory factor that binds to a neighboring recognition site is also required for efficient gene induction. Some of these putative co-regulatory proteins are members of transcription factor families that all bind to the same DNA sequence elements in vitro and are often expressed in the same cells. These two observations have made it difficult to assign specific and redundant functions to the unique members of a specific gene family. We have used the chromatin immunoprecipitation (ChIP) technique coupled with a transient complementation assay in Drosophila SL2 cells to directly compare the ability of two members of the CREB/ATF family to function as co regulatory proteins for SREBP-dependent activation of the HMG-CoA reductase promoter. Results from both of these experimental systems demonstrate that CREB is an efficient SREBP co-regulator but ATF-2 is not. PMID- 12110666 TI - Site-directed mutagenesis of the basic N-terminal cluster of pancreatic bile salt dependent lipase. Functional significance. AB - Previous studies have postulated the presence of a heparin-binding site on the bile salt-dependent lipase (BSDL), whereas two bile salt-binding sites regulate the enzyme activity. One of these sites may overlap with the tentative heparin binding site at the level of an N-terminal basic cluster consisting of positive residues Lys(32), Lys(56), Lys(61), Lys(62), and Arg(63). The present study uses specific site-directed mutagenesis to determine the functional significance of this basic cluster. Mutations in this sequence resulted in recombinant enzymes that were able to bind to immobilized and to cell-associated heparin before moving throughout intestinal cells. Recombinant BSDL was fully active on soluble substrate, but mutants were less active on micellar cholesteryl oleate in comparison with the wild-type enzyme. Activation studies by primary (sodium taurocholate) and by secondary (sodium taurodeoxycholate) bile salts revealed that the activation of BSDL by sodium taurocholate at concentrations below the critical micellar concentration, and not that evoked by micellar bile salts, was affected by substitutions, suggesting that this N-terminal basic cluster likely represents the specific bile salt-binding site of BSDL. Substitutions also affected the activation of the enzyme promoted by anionic phospholipids, extending the function of this site to that of a cationic regulatory site susceptible to accommodate anionic ligands. PMID- 12110668 TI - A unique developmental pattern of Oct-3/4 DNA methylation is controlled by a cis demodification element. AB - Oct-3/4 is the earliest expressed transcription factor that is known to be crucial in murine pre-implantation development. In this report we asked whether methylation participates in controlling changes in Oct-3/4 expression and thus may play an important role in controlling normal embryogenesis. We show that the Oct-3/4 gene is unmethylated from the blastula stage but undergoes de novo methylation at 6.5 days post-coitum and remains modified in all adult somatic tissues analyzed. Oct-3/4 remains unmethylated in 6.25 days post-coitum epiblast cells when other genes, such as apoAI, undergo de novo methylation. We show that methylation of the Oct-3/4 promoter sequence strongly compromises its ability to direct efficient transcription. Moreover, DNA methylation inhibits basal transcription of the endogenous Oct-3/4 gene in vivo. We found that the Oct-3/4 gene harbors a cis-specific demodification element that includes the proximal enhancer sequence. This element leads to demethylation in embryonal carcinoma cells when the sequence is initially methylated and protects the local region from de novo methylation in post-implantation embryos. These results indicate that in the embryo protection from de novo methylation is not a unique feature of imprinted or housekeeping genes that carry a CpG island, but is also applicable to tissue-specific genes expressed during early stages of embryogenesis. Methylation of Oct-3/4 may be analogous to methylation of CpG islands on the inactive X chromosome that also occurs at later stages of development. PMID- 12110667 TI - Vascular smooth muscle alpha-actin gene transcription during myofibroblast differentiation requires Sp1/3 protein binding proximal to the MCAT enhancer. AB - The conversion of stromal fibroblasts into contractile myofibroblasts is an essential feature of the wound-healing response that is mediated by transforming growth factor beta1 (TGF-beta1) and accompanied by transient activation of the vascular smooth muscle alpha-actin (SmalphaA) gene. Multiple positive-regulatory elements were identified as essential mediators of basal SmalphaA enhancer activity in mouse AKR-2B stromal fibroblasts. Three of these elements bind transcriptional activating proteins of known identity in fibroblasts. A fourth site, shown previously to be susceptible to single-strand modifying agents in myofibroblasts, was additionally required for enhancer response to TGF-beta1. However, TGF-beta1 activation was not accompanied by a stoichiometric increase in protein binding to any known positive element in the SmalphaA enhancer. By using oligonucleotide affinity isolation, DNA-binding site competition, gel mobility shift assays, and protein overexpression in SL2 and COS7 cells, we demonstrate that the transcription factors Sp1 and Sp3 can stimulate SmalphaA enhancer activity. One of the sites that bind Sp1/3 corresponds to the region of the SmalphaA enhancer required for TGF-beta1 amplification. Additionally, the TGF beta1 receptor-regulated Smad proteins, in particular Smad3, are rate-limiting for SmalphaA enhancer activation. Whereas Smad proteins collaborate with Sp1 in activating several stromal cell-associated promoters, they appear to operate independently from the Sp1/3 proteins in activating the SmalphaA enhancer. The identification of Sp and Smad proteins as essential, independent activators of the SmalphaA enhancer provides new insight into the poorly understood process of myofibroblast differentiation. PMID- 12110669 TI - The immunoglobulin D-binding part of the outer membrane protein MID from Moraxella catarrhalis comprises 238 amino acids and a tetrameric structure. AB - Moraxella catarrhalis IgD-binding protein (MID), a 200-kDa outer membrane protein comprising 2,139 amino acids, has recently been isolated and shown to display a unique and specific affinity for human IgD. To identify the IgD-binding region, MID was digested with proteases. In addition, a series of truncated fragments of MID were manufactured and expressed in Escherichia coli followed by analysis for IgD binding in Western and dot blots. The smallest fragment with essentially preserved IgD binding was comprised of 238 amino acid residues (MID(962-1200)). Shorter recombinant proteins gradually lost IgD-binding capacity, and the shortest IgD-binding fragment comprising 157 amino acids (MID(985-1142)) displayed a 1,000-fold reduced IgD binding compared with the full-length molecule. The truncated MID(962-1200) was efficiently attracted to a standard IgD serum and to purified myeloma IgD(kappa) and IgD(lambda) sera but not to IgG, IgM, or IgA myeloma sera. Furthermore, the fragment specifically bound to peripheral blood B lymphocytes, and the binding was inhibited by preincubation with anti-IgD-Fab polyclonal antibodies. Results obtained by introducing five amino acids randomly into MID(962-1200) using transposons suggested that alpha helix structures were important for IgD binding. Ultracentrifugation experiments and gel electrophoresis revealed that native MID(962-1200) was a tetramer. Interestingly, tetrameric MID(962-1200) attracted IgD more than 20-fold more efficiently than the monomeric form. Thus, a tetrameric structure of MID(962 1200) is crucial for optimal IgD-binding capacity. PMID- 12110670 TI - Folding of the striated muscle myosin motor domain. AB - We have investigated the folding of the myosin motor domain using a chimera of an embryonic striated muscle myosin II motor domain fused on its COOH terminus to a thermal stable, fast folding variant of green fluorescent protein (GFP). In in vitro expression assays, the GFP domain of the chimeric protein, S1(795)GFP, folds rapidly enabling us to monitor the folding of the motor domain using fluorescence. The myosin motor domain folds very slowly and transits through multiple intermediates that are detectable by gel filtration chromatography. The distribution of the nascent protein among these intermediates is strongly dependent upon temperature. At 25 degrees C and above the predominant product is an aggregate of S1(795)GFP or a complex with other lysate proteins. At 0 degrees C, the motor domain folds slowly via an energy independent pathway. The unusual temperature dependence and slow rate suggests that folding of the myosin motor is highly susceptible to off-pathway interactions and aggregation. Expression of the S1(795)GFP in the C2C12 muscle cell line yields a folded and functionally active protein that exhibits Mg(2+)ATP-sensitive actin-binding and myosin motor activity. In contrast, expression of S1(795)GFP in kidney epithelial cell lines (human 293 and COS 7 cells) results in an inactive and aggregated protein. The results of the in vitro folding assay suggest that the myosin motor domain does not fold spontaneously under physiological conditions and probably requires cytosolic chaperones. The expression studies support this conclusion and demonstrate that these factors are optimized in muscle cells. PMID- 12110671 TI - c-Myc represses and Miz-1 activates the murine natural resistance-associated protein 1 promoter. AB - Iron is essential for growth, and impaired iron homoeostasis through a non conserved mutation within murine Nramp1, also termed Slc11a1, contributes to susceptibility to infection. Nramp1 depletes the macrophage cytosol of iron, with effects on iron-regulated gene expression and iron-dependent processes. Wu and colleagues (Wu, K.-J., Polack, A., and Dalla-Favera, R. (1999) Science 283, 676 679) showed converse control of iron regulatory protein expression (IRP2) and H ferritin by c-Myc, suggesting a role for c-Myc in enhancing cytoplasmic iron levels for growth. We investigated if c-Myc also regulates Nramp1 expression. We show an inverse correlation with cell growth, and in co-transfection experiments c-Myc represses the Nramp1 promoter. Within the Nramp1 promoter we identified six non-canonical E boxes, which are not important for c-Myc repression. By deletion analysis the repressor site maps to one or more initiator elements flanking the transcriptional initiation site. Co-transfections with the c-Myc interacting zinc finger protein (Miz-1) show that Miz-1 can overcome c-Myc repression of Nramp1, and, from a deletion construct lacking E box sites, Miz-1 activates the Nramp1 promoter. These studies reinforce the link between c-Myc and iron regulation and provide further evidence that c-Myc negatively regulates genes that decrease the iron content of the cytosol. The results provide further support for a divalent cation antiporter function for Nramp1. PMID- 12110673 TI - A functionally active human F1F0 ATPase can be purified by immunocapture from heart tissue and fibroblast cell lines. Subunit structure and activity studies. AB - Human mitochondrial F(1)F(0) ATP synthase was isolated with a one-step immunological approach, using a monoclonal antibody against F(1) in a 96-well microplate activity assay system, to establish a method for fast high throughput screening of inhibitors, toxins, and drugs with very small amounts of enzyme. For preparative purification, mitochondria from human heart tissue as well as cultured fibroblasts were solubilized with dodecyl-beta-d-maltoside, and the F(1)F(0) was isolated with anti-F(1) monoclonal antibody coupled to protein G agarose beads. The immunoprecipitated F(1)F(0) contained a full complement of subunits that were identified with specific antibodies against five of the subunits (alpha, beta, OSCP, d, and IF(1)) and by MALDI-TOF and/or LC/MS/MS for all subunits except subunit c, which could not be resolved by these methods because of the limits of detection. Microscale immunocapture of F(1)F(0) from detergent-solubilized mitochondria or whole cell fibroblast extracts was performed using anti-F(1) monoclonal antibody immobilized on 96-well microplates. The captured complex V displayed ATP hydrolysis activity that was fully oligomycin and inhibitor protein IF(1)-sensitive. Moreover, IF(1) could be co isolated with F(1)F(0) when the immunocapture procedure was carried out at pH 6.5 but was absent when the ATP synthase was isolated at pH 8.0. Immunocaptured F(1)F(0) lacking IF(1) could be inhibited by more than 90% by addition of recombinant inhibitor protein, and conversely, F(1)F(0) containing IF(1) could be activated more than 10-fold by brief exposure to pH 8.0, inducing the release of inhibitor protein. With this microplate system an ATP hydrolysis assay of complex V could be carried out with as little as 10 ng of heart mitochondria/well and as few as 3 x 10(4) cells/well from fibroblast cultures. The system is therefore suitable to screen patient-derived samples for alterations in amount or functionality of both the F(1)F(0) ATPase and IF(1). PMID- 12110672 TI - Highly purified scavenger receptor class B, type I reconstituted into phosphatidylcholine/cholesterol liposomes mediates high affinity high density lipoprotein binding and selective lipid uptake. AB - The murine class B, type I scavenger receptor mSR-BI is a high and low density lipoprotein (HDL and LDL) receptor that mediates selective uptake of cholesteryl esters. Here we describe a reconstituted phospholipid/cholesterol liposome assay of the binding and selective uptake activities of SR-BI derived from detergent solubilized cells. The assay, employing lysates from epitope-tagged receptor (mSR BI-t1)-expressing mammalian and insect cells, recapitulated many features of SR BI activity in intact cells, including high affinity and saturable (125)I-HDL binding, selective lipid uptake from [(3)H]cholesteryl ether-labeled HDL, and poor inhibition of HDL receptor activity by LDL. The novel properties of a mutated receptor (Q402R/Q418R, normal LDL binding but loss of most HDL binding) were reproduced in the assay, as was the ability of the SR-BI homologue CD36 to bind HDL but not mediate efficient lipid uptake. In this assay, essentially homogeneously pure mSR-BI-t1, prepared by single-step immunoaffinity chromatography, mediated high affinity HDL binding and efficient selective lipid uptake from HDL. Thus, SR-BI-mediated HDL binding and selective lipid uptake are intrinsic properties of the receptor that do not require the intervention of other proteins or specific cellular structures or compartments. PMID- 12110674 TI - A conserved motif common to the histone acetyltransferase Esa1 and the histone deacetylase Rpd3. AB - Post-translational modification of histones enables dynamic regulation of chromatin structure in eukaryotes. Histone acetyltransferase (HAT) and histone deacetylase (HDAC) modify the N-terminal tails of histones by adding or removing acetyl groups to specific lysine residues. A particular pair of HAT (Esa1) and HDAC (Rpd3) is proposed to modify the same lysine residue in vitro and in vivo. Thus, HAT and HDAC might have similar structural and functional motifs. Here we show that HAT (Esa1 family) and HDAC (Rpd3 family) have similar amino acid stretches in the primary structures through evolution. We refer to this region as the "ER (Esa1-Rpd3) motif." In the tertiary structure of Esa1, the ER motif is located near the active center. In Rpd3, for which the tertiary structure remains unclear, we demonstrate that the ER motif contains the same secondary structure as found in Esa1 by circular dichroism analysis. We did alanine-scanning mutagenesis and found that the ER motif regions of Esa1 or Rpd3 are required for HAT activity of Esa1 or HDAC activity of Rpd3, respectively. Our discovery of the ER motif present in the pair of enzymes (HAT and HDAC) indicates that HAT and HDAC have common structural bases, although they catalyze the reaction with opposite functions. PMID- 12110675 TI - Characterization of the conserved interaction between GATA and FOG family proteins. AB - The N-terminal zinc finger (ZnF) from GATA transcription factors mediates interactions with FOG family proteins. In FOG proteins, the interacting domains are also ZnFs; these domains are related to classical CCHH fingers but have an His --> Cys substitution at the final zinc-ligating position. Here we demonstrate that different CCHC fingers in the FOG family protein U-shaped contact the N terminal ZnF of GATA-1 in the same fashion although with different affinities. We also show that these interactions are of moderate affinity, which is interesting given the presumed low concentrations of these proteins in the nucleus. Furthermore, we demonstrate that the variant CCHC topology enhances binding affinity, although the His --> Cys change is not essential for the formation of a stably folded domain. To ascertain the structural basis for the contribution of the CCHC arrangement, we have determined the structure of a CCHH mutant of finger nine from U-shaped. The structure is very similar overall to the wild-type domain, with subtle differences at the C terminus that result in loss of the interaction in vivo. Taken together, these results suggest that the CCHC zinc binding topology is required for the integrity of GATA-FOG interactions and that weak interactions can play important roles in vivo. PMID- 12110676 TI - Molecular and global time-resolved analysis of a psbS gene dosage effect on pH- and xanthophyll cycle-dependent nonphotochemical quenching in photosystem II. AB - Photosynthetic light harvesting in plants is regulated by a pH- and xanthophyll dependent nonphotochemical quenching process (qE) that dissipates excess absorbed light energy and requires the psbS gene product. An Arabidopsis thaliana mutant, npq4-1, lacks qE because of a deletion of the psbS gene, yet it exhibits a semidominant phenotype. Here it is shown that the semidominance is due to a psbS gene dosage effect. Diploid Arabidopsis plants containing two psbS gene copies (wild-type), one psbS gene (npq4-1/NPQ4 heterozygote), and no psbS gene (npq4 1/npq4-1 homozygote) were compared. Heterozygous plants had 56% of the wild-type psbS mRNA level, 58% of the wild-type PsbS protein level, and 60% of the wild type level of qE. Global analysis of the chlorophyll a fluorescence lifetime distributions revealed three components in wild-type and heterozygous plants, but only a single long lifetime component in npq4-1. The short lifetime distribution associated with qE was inhibited by more than 40% in heterozygous plants compared with the wild type. Thus, the extent of qE measured as either the fractional intensities of the PSII chlorophyll a fluorescence lifetime distributions or steady state intensities was stoichiometrically related to the amount of PsbS protein. PMID- 12110677 TI - The respiratory complex I (NDH I) from Klebsiella pneumoniae, a sodium pump. AB - The electrogenic NADH:Q oxidoreductase from the enterobacterium Klebsiella pneumoniae transports Na(+) ions. The complex was purified with an increase of the specific Na(+) transport activity from 0.2 micromol min(-1) mg(-1) in native membrane vesicles to 4.7 micromol min(-1) mg(-1) in reconstituted enzyme specimens. The subunit pattern resembled that of complex I from Escherichia coli, and two prominent polypeptides were identified as the NuoF and NuoG subunits of complex I. During purification the typical cofactors of complex I were enriched to yield approximately 17 nmol mg(-1) iron, 24 nmol mg(-1) acid-labile sulfide, and 0.79 nmol mg(-1) FMN in the purified sample. The enzyme contained approximately 1.2 nmol mg(-1) Q6 and 1.5 nmol mg(-1) Q8. The reduction of ubiquinone by NADH was Na(+)-dependent, which indicates coupling of the chemical and the vectorial reaction of the pump. The Na(+) activation profile corresponded to the Hill equation with a Hill coefficient K(H)(Na(+)) = 1.96 and with a half maximal saturation at 0.33 mm Na(+). The reconstituted complex I from Klebsiella pneumoniae catalyzed deamino-NADH oxidation, Q1 reduction, and Na(+) translocation with specific activities of 2.6 units mg(-1), 2.4 units mg(-1), and 4.7 units mg(-1), respectively, which indicate a Na(+)/electron stoichiometry of one. PMID- 12110678 TI - The consequence of sequence alteration of an amphipathic alpha-helical antimicrobial peptide and its diastereomers. AB - The search for antibiotics with a new mode of action led to numerous studies on antibacterial peptides. Most of the studies were carried out with l-amino acid peptides possessing amphipathic alpha-helix or beta-sheet structures, which are known to be important for biological activities. Here we compared the effect of significantly altering the sequence of an amphipathic alpha-helical peptide (15 amino acids long) and its diastereomer (composed of both l- and d-amino acids) regarding their structure, function, and interaction with model membranes and intact bacteria. Interestingly, the effect of sequence alteration on biological function was similar for the l-amino acid peptides and the diastereomers, despite some differences in their structure in the membrane as revealed by attenuated total reflectance Fourier-transform infrared spectroscopy. However, whereas the all l-amino acid peptides were highly hemolytic, had low solubility, lost their activity in serum, and were fully cleaved by trypsin and proteinase K, the diastereomers were nonhemolytic and maintained full activity in serum. Furthermore, sequence alteration allowed making the diastereomers either fully, partially, or totally protected from degradation by the enzymes. Transmembrane potential depolarization experiments in model membranes and intact bacteria indicate that although the killing mechanism of the diastereomers is via membrane perturbation, it is also dependent on their ability to diffuse into the inner bacterial membrane. These data demonstrate the advantage of the diastereomers over their all l-amino acid counterparts as candidates for developing a repertoire of new target antibiotics with a potential for systemic use. PMID- 12110680 TI - Focal adhesion kinase activated by beta(4) integrin ligation to mCLCA1 mediates early metastatic growth. AB - Early metastatic growth occurs at sites of vascular arrest of blood-borne cancer cells and is entirely intravascular. Here we show that lung colonization by B16 F10 cells is licensed by beta(4) integrin adhesion to the mouse lung endothelial Ca(2+)-activated chloride channel protein mCLCA1. In a manner independent of Met, beta(4) integrin-mCLCA1-ligation leads to complexing with and activation of focal adhesion kinase (FAK) and downstream signaling to extracellular signal-regulated kinase (ERK). FAK/ERK signaling is Src-dependent and is interrupted by adhesion blocking antibodies and by dominant-negative (dn)-FAK mutants. Levels of ERK activation in B16-F10 cells transfected with wild-type or mutant FAK are closely associated with rates of proliferation and bromodeoxyuridine (BrdUrd) incorporation of tumor cells grown in mCLCA1-coated dishes, the ability to form tumor cell colonies on CLCA-expressing endothelial cell monolayers, and the extent of pulmonary metastatic growth. Parallel with the transfection rates, B16 F10 cells transfected with dn-FAK mutants and injected intravenously into syngeneic mice generate approximately half the number and size of lung colonies that vector-transfected B16-F10 cells produce. For the first time, beta(4) integrin ligation to its novel CLCA-adhesion partner is shown to be associated with FAK complexing, activation, and signaling to promote early, intravascular, metastatic growth. PMID- 12110679 TI - Expression of the palmitoylation-deficient CD151 weakens the association of alpha 3 beta 1 integrin with the tetraspanin-enriched microdomains and affects integrin dependent signaling. AB - Transmembrane proteins of the tetraspanin superfamily are assembled in multimeric complexes on the cell surface. Spatial orientation of tetraspanins within these complexes may affect signaling functions of the associated transmembrane receptors (e.g. integrins, receptor-type tyrosine kinases). The structural determinants that control assembly of the tetraspanin complexes are unknown. We have found that various tetraspanins and the alpha(3) integrin subunit are palmitoylated. The stability and molecular composition of the palmitoylated alpha(3)beta(1)-tetraspanin complexes are not affected by adhesion. To assess the significance of palmitoylation in the function of the alpha(3)beta(1)-tetraspanin complexes we mapped the sites of palmitoylation for CD151. Mutation of six cysteines, Cys(11), Cys(15), Cys(79), Cys(80), Cys(242), and Cys(243) was necessary to completely abolish palmitoylation of CD151. The association of the palmitoylation-deficient mutant of CD151 (CD151Cys8) with CD81 and CD63 was markedly decreased, but the interaction of the alpha(3)beta(1)-CD151Cys8 complex with phosphatidylinositol 4-kinase was not affected. Ectopic expression of CD151Cys8 in Rat-1 cells impaired the interactions of the endogenous CD63 and CD81 with the alpha(3)beta(1) integrin. Although the expression of the palmitoylation-deficient CD151 does not change cell spreading on the extracellular matrix, the number of focal adhesions increased. Adhesion-induced phosphorylation of PKB/c-Akt is markedly increased in cells expressing a palmitoylation-deficient mutant, thereby providing direct evidence for the role of the tetraspanin microdomains in regulation of the integrin-dependent phosphatidylinositol 3-kinase signaling pathway. In contrast, activation of FAK and ERK1/2 were not affected by the expression of CD151Cys8. Our results demonstrate that palmitoylation of tetraspanins is critical not only for the organization of the integrin-tetraspanin microdomains but also has a specific role in modulation of adhesion-dependent signaling. PMID- 12110681 TI - Cloning, characterization, and functional studies of a 47-kDa platelet receptor for type III collagen. AB - A 1.2-kb cDNA fragment encoding a platelet 47-kDa protein has been isolated from a human bone marrow cDNA library by using a degenerate oligonucleotide of the sequenced amino terminus of the purified platelet protein with a poly(dT)(12).(dG) by polymerase chain reaction. A computer search revealed that the cDNA represents the coding sequence of a protein with a fragmentary homology to several proteins. Using a prokaryotic expression system, pBad TOPO-47 cDNA, a 47-kDa recombinant protein was obtained and purified to apparent homogeneity by nickel-nitrilotriacetic acid resin and collagen affinity column. The recombinant protein binds to type III but not type I collagen-Sepharose 2B affinity columns. Anti-47-kDa but not anti-65-kDa antibody inhibits the binding of the recombinant protein to the type III collagen-coated micro titer wells in a dose-dependent manner. Like the receptor protein purified from platelet membranes, the recombinant protein inhibits type III collagen-induced platelet aggregation also in a dose-dependent manner. We have defined two active peptides from the cloned deduced amino acid sequence. Both peptides inhibit type III but not type I collagen-induced platelet aggregation in a dose-dependent fashion. These results suggest that the active binding site of the platelet receptor to type III collagen resides in these portions of the protein. PMID- 12110682 TI - Elf1p, a member of the ABC class of ATPases, functions as a mRNA export factor in Schizosacchromyces pombe. AB - Rae1p and Mex67p/Tap are conserved mRNA export factors. We have used synthetic lethal genetic screens in Schizosaccharomyces pombe to identify mutations in genes that are functionally linked to rae1 and mex67 in mRNA export. From these screens, we have isolated mutations in a putative S. pombe homologue of the Candida albicans elf1 gene. The elf1 of S. pombe is not an essential gene. When elf1 mutations are combined with rae1-167 mutation, growth and mRNA export is inhibited in the double mutants. This inhibition can be suppressed by the multicopy expression of mex67 suggesting that Mex67p can substitute for the loss of Elf1p function. Elf1p is a non-membrane member of the ATP-binding cassette (ABC) class of ATPase and the GFP-Elf1p fusion localizes to the cytoplasm. Elf1p, expressed and purified from Escherichia coli, binds and hydrolyzes ATP. A mutant Elf1p that carries a glycine to aspartic acid (G731D) mutation within the Walker A domain of the second ATP site retains the ATP binding but loses its ATPase activity in vitro. This mutant protein no longer functions in mRNA export. Taken together, our results show that Elf1p functions as a mRNA export factor along with Rae1p and Mex67p in S. pombe. PMID- 12110683 TI - Protein-tyrosine phosphatase-sigma is a novel member of the functional family of alpha-latrotoxin receptors. AB - Receptor-like protein-tyrosine phosphatase sigma (PTPvarsigma) is essential for neuronal development and function. Here we report that PTPvarsigma is a target of alpha-latrotoxin, a strong stimulator of neuronal exocytosis. alpha-Latrotoxin binds to the cell adhesion-like extracellular region of PTPvarsigma. This binding results in the stimulation of exocytosis. The toxin-binding site is located in the C-terminal part of the PTPvarsigma ectodomain and includes two fibronectin type III repeats. The intracellular catalytic domains of PTPvarsigma are not required for the alpha-latrotoxin binding and secretory response triggered by the toxin in chromaffin cells. These features of PTPvarsigma resemble two other previously described alpha-latrotoxin receptors, neurexin and CIRL. Thus, alpha latrotoxin represents an unusual example of the neurotoxin that has three independent, equally potent, and yet structurally distinct targets. The known structural and functional characteristics of PTPvarsigma, neurexin, and CIRL suggest that they define a functional family of neuronal membrane receptors with complementary or converging roles in presynaptic function via a mechanism that involves cell-to-cell and cell-to-matrix interaction. PMID- 12110684 TI - Mutations in the nucleotide binding domain 1 signature motif region rescue processing and functional defects of cystic fibrosis transmembrane conductance regulator delta f508. AB - The gene encoding the cystic fibrosis transmembrane conductance regulator (CFTR), an ATP binding cassette (ABC) transporter that functions as a phosphorylation- and nucleotide-regulated chloride channel, is mutated in cystic fibrosis (CF) patients. Deletion of a phenylalanine at amino acid position 508 (DeltaF508) in the first nucleotide binding domain (NBD1) is the most prevalent CF-causing mutation and results in defective protein processing and reduced CFTR function, leading to chloride impermeability in CF epithelia and heterologous systems. Using a STE6/CFTRDeltaF508 chimera system in yeast, we isolated two novel DeltaF508 revertant mutations, I539T and G550E, proximal to and within the conserved ABC signature motif of NBD1, respectively. Western blot and functional analysis in mammalian cells indicate that mutations I539T and G550E each partially rescue the CFTRDeltaF508 defect. Furthermore, a combination of both revertant mutations resulted in a 38-fold increase in CFTRDeltaF508-mediated chloride current, representing 29% of wild type channel activity. The G550E mutation increased the sensitivity of CFTRDeltaF508 and wild type CFTR to activation by cAMP agonists and blocked the enhancement of CFTRDeltaF508 channel activity by 2 mm 3-isobutyl-1-methylxanthine. The data show that the DeltaF508 defect can be significantly rescued by second-site mutations in the nucleotide binding domain 1 region, that includes the LSGGQ consensus motif. PMID- 12110685 TI - Salt bridges at the inter-ring interface regulate the thermostat of GroEL. AB - The chaperonin GroEL consists of a double-ring structure made of identical subunits and displays unusual allosteric properties caused by the interaction between its constituent subunits. Cooperative binding of ATP to a protein ring allows binding of GroES to that ring, and at the same time negative inter-ring cooperativity discharges the ligands from the opposite ring, thus driving the protein-folding cycle. Biochemical and electron microscopy analysis of wild type GroEL, a single-ring mutant (SR1), and two mutants with one inter-ring salt bridge of the chaperonin disrupted (E461K and E434K) indicate that these ion pairs form part of the interactions that allow the inter-ring allosteric signal to be transmitted. The wild type-like activities of the ion pair mutants at 25 degrees C are in contrast with their lack of inter-ring communication and folding activity at physiological temperatures. These salt bridges stabilize the inter ring interface and maintain the inter-ring spacing so that functional communication between protein heptamers takes place. The characterization of GroEL hybrids containing different amounts of wild type and mutant subunits also indicates that as the number of inter-ring salt bridges increases the functional properties of the hybrids recover. Taken together, these results strongly suggest that inter-ring salt bridges form a stabilizing ring-shaped, ionic zipper that ensures inter-ring communication at the contact sites and therefore a functional protein-folding cycle. Furthermore, they regulate the chaperonin thermostat, allowing GroEL to distinguish physiological (37 degrees C) from stress temperatures (42 degrees C). PMID- 12110687 TI - Coordinated folding and association of the LIN-2, -7 (L27) domain. An obligate heterodimerization involved in assembly of signaling and cell polarity complexes. AB - LIN-2, -7 (L27) homology domains are putative protein-protein interaction modules found in several scaffold proteins involved in the assembly of polarized cell signaling structures. These specific interaction pairs are well conserved across metazoan species, from worms to man. We have expressed and purified L27 domains from multiple species and find that certain domains from proteins such as Caenorhabditis elegans LIN-2 and LIN-7 can specifically heterodimerize. Biophysical analysis of interacting L27 domains demonstrates that the domains interact with a 1:1 stoichiometry. Circular dichroism studies reveal that the domains appear to function as an obligate heterodimer; individually the domains are largely unfolded, but when associated they show a significant increase in helicity, as well as a cooperative unfolding transition. These novel obligate interacting pairs are likely to play a key role in regulating the organization of signaling proteins at polarized cell structures. PMID- 12110686 TI - Cyclosporine A enhances leukocyte binding by human intestinal microvascular endothelial cells through inhibition of p38 MAPK and iNOS. Paradoxical proinflammatory effect on the microvascular endothelium. AB - The calcineurin inhibitor cyclosporine A (CsA) modulates leukocyte cytokine production but may also effect nonimmune cells, including microvascular endothelial cells, which regulate the inflammatory process through leukocyte recruitment. We hypothesized that CsA would promote a proinflammatory phenotype in human intestinal microvascular endothelial cells (HIMEC), by inhibiting inducible nitric-oxide synthase (iNOS, NOS2)-derived NO, normally an important mechanism in limiting endothelial activation and leukocyte adhesion. Primary cultures of HIMEC were used to assess CsA effects on endothelial activation, leukocyte interaction, and the expression of iNOS as well as cell adhesion molecules. CsA significantly increased leukocyte binding to activated HIMEC, but paradoxically decreased endothelial expression of cell adhesion molecules (E selectin, intercellular adhesion molecule 1, and vascular cell adhesion molecule 1). In contrast, CsA completely inhibited the expression of iNOS in tumor necrosis factor-alpha/lipopolysaccharide-activated HIMEC. CsA blocked p38 MAPK phosphorylation in activated HIMEC, a key pathway in iNOS expression, but failed to inhibit NFkappaB activation. These studies demonstrate that CsA exerts a proinflammatory effect on HIMEC by blocking iNOS expression. CsA exerts a proinflammatory effect on the microvascular endothelium, and this drug-induced endothelial dysfunction may help explain its lack of efficacy in the long-term treatment of chronically active inflammatory bowel disease. PMID- 12110688 TI - Structures of the complexes of a potent anti-HIV protein cyanovirin-N and high mannose oligosaccharides. AB - The development of anti-human immunodeficiency virus (HIV) microbicides for either topical or ex vivo use is of considerable interest, mainly due to the difficulties in creating a vaccine that would be active against multiple clades of HIV. Cyanovirin-N (CV-N), an 11-kDa protein from the cyanobacterium (blue green algae) Nostoc ellipsosporum with potent virucidal activity, was identified in the search for such antiviral agents. The binding of CV-N to the heavily glycosylated HIV envelope protein gp120 is carbohydrate-dependent. Since previous CV-N-dimannose structures could not fully explain CV-N-oligomannose binding, we determined the crystal structures of recombinant CV-N complexed to Man-9 and a synthetic hexamannoside, at 2.5- and 2.4-A resolution, respectively. CV-N is a three-dimensional domain-swapped dimer in the crystal structures with two primary sites near the hinge region and two secondary sites on the opposite ends of the dimer. The binding interface is constituted of three stacked alpha1-->2-linked mannose rings for Man-9 and two stacked mannose rings for hexamannoside with the rest of the saccharide molecules pointing to the solution. These structures show unequivocally the binding geometry of high mannose sugars to CV-N, permitting a better understanding of carbohydrate binding to this potential new lead for the design of drugs against AIDS. PMID- 12110689 TI - Fibroblast growth factor 2 induction of the osteocalcin gene requires MAPK activity and phosphorylation of the osteoblast transcription factor, Cbfa1/Runx2. AB - Fibroblast growth factor 2 (FGF-2) is an important regulator of bone formation and osteoblast activity. However, its mechanism of action on bone cells is largely unknown. A major route for FGF signaling is through the mitogen-activated protein kinase (MAPK) pathway. We showed recently that this pathway is important for activation and phosphorylation of Cbfa1/Runx2, an osteoblast-related transcription factor (Xiao, G., Jiang, D., Thomas, P., Benson, M. D., Guan, K., Karsenty, G., and Franceschi, R. T. (2000) J. Biol. Chem. 275, 4453-4459). The present study examined the mechanism of FGF-2 regulation of the mouse osteocalcin gene in MC3T3-E1 preosteoblastic cells. FGF-2 stimulated osteocalcin mRNA and promoter activity in a dose- and time-dependent manner in MC3T3-E1 preosteoblastic cells. Similar results were obtained in mouse bone marrow stromal cells. This stimulation required Runx2 and its DNA binding site in the osteocalcin promoter. FGF-2 also dramatically increased phosphorylation of extracellular signal-regulated kinase 1 and 2 (ERK1/2) followed by phosphorylation of Runx2. Furthermore, a specific ERK1/2 phosphorylation inhibitor, U0126, completely blocked both FGF-2-stimulated Runx2 phosphorylation and osteocalcin promoter activity, indicating that this regulation requires the MAPK pathway. Deletion studies showed that the C-terminal PST domain of Runx2 is required for the FGF-2 response. This study is the first demonstration that Runx2 is phosphorylated and activated by FGF-2 via the MAPK pathway and suggests that FGF-2 plays an important role in regulation of Runx2 function and bone formation. PMID- 12110690 TI - New insights into the role of the N terminus in conformational transitions of the Na,K-ATPase. AB - The deletion of 32 residues from the N terminus of the alpha1 catalytic subunit of the rat Na,K-ATPase (mutant alpha1M32) shifts the E(1)/E(2) conformational equilibrium toward E(1), and the combination of this deletion with mutation E233K in the M2-M3 loop acts synergistically to shift the conformation further toward E(1) (Boxenbaum, N., Daly, S. E., Javaid, Z. Z., Lane, L. K., and Blostein, R. (1998) J. Biol. Chem. 273, 23086-23092). To delimit the region of the cytoplasmic N terminus involved in these interactions, the consequences of a series of N terminal deletions of alpha1 beyond Delta32 were evaluated. Criteria to assess shifts in conformational equilibrium were based on effects of perturbation of the entire catalytic cycle ((i) sensitivity to vanadate inhibition, (ii) K(+) sensitivity of Na-ATPase measured at micromolar ATP, (iii) changes in K'(ATP), and (iv) catalytic turnover), as well as estimates of the rates of the conformational transitions of phospho- and dephosphoenzyme (E(1)P --> E(2)P and E(2)(K(+)) --> E(1) + K(+)). The results show that, compared with alpha1M32, the deletion of up to 40 residues (alpha1M40) further shifts the poise toward E(1). Remarkably, further deletions (mutants alpha1M46, alpha1M49, and alpha1M56) reverse the effect, such that these mutants increasingly resemble the wild type alpha1. These results suggest novel intramolecular interactions involving domains within the N terminus that impact the manner in which the N terminus/M2-M3 loop regulatory domain interacts with the M4-M5 catalytic loop to effect E(1) <--> E(2) transitions. PMID- 12110691 TI - Functional mapping of Bas2. Identification of activation and Bas1-interaction domains. AB - The transcriptional activator protein Bas2 is required to express more than 20 genes in pathways for purine nucleotide and histidine biosynthesis, phosphate utilization, and the HO endonuclease by acting with co-regulator proteins Bas1, Pho4, and Swi5. The role that Bas2 plays in transcriptional activation may be to unmask latent activation domains in the co-regulator and to promote ternary complex formation between Bas2, the co-regulator, and DNA. We show that Bas2 also contributes to transcriptional activation by providing an activation domain. We localize this domain in Bas2 to the C-terminal 156 amino acids using deletion analysis and fusion to a heterologous DNA binding domain. Additionally, we show that Bas2 makes direct contacts with Bas1. This interaction is detected by co immunoprecipitation and by two-hybrid analysis. We localize the interaction region to the central portion of Bas2, from amino acids 112 to 404. PMID- 12110692 TI - Beta-glucoside kinase (BglK) from Klebsiella pneumoniae. Purification, properties, and preparative synthesis of 6-phospho-beta-D-glucosides. AB - ATP-dependent beta-glucoside kinase (BglK) has been purified from cellobiose grown cells of Klebsiella pneumoniae. In solution, the enzyme (EC ) exists as a homotetramer composed of non-covalently linked subunits of M(r) approximately 33,000. Determination of the first 28 residues from the N terminus of the protein allowed the identification and cloning of bglK from genomic DNA of K. pneumoniae. The open reading frame (ORF) of bglK encodes a 297-residue polypeptide of calculated M(r) 32,697. A motif of 7 amino acids (AFD(7)IG(9)GT) near the N terminus may comprise the ATP-binding site, and residue changes D7G and G9A yielded catalytically inactive proteins. BglK was progressively inactivated (t(12) approximately 19 min) by N-ethylmaleimide, but ATP afforded considerable protection against the inhibitor. By the presence of a centrally located signature sequence, BglK can be assigned to the ROK (Repressor, ORF, Kinase) family of proteins. Preparation of (His6)BglK by nickel-nitrilotriacetic acid agarose chromatography provided high purity enzyme in quantity sufficient for the preparative synthesis (200-500 mg) of ten 6-phospho-beta-d-glucosides, including cellobiose-6'-P, gentiobiose-6'-P, cellobiitol-6-P, salicin-6-P, and arbutin-6-P. These (and other) derivatives are substrates for phospho-beta-glucosidase(s) belonging to Families 1 and 4 of the glycosylhydrolase superfamily. The structures, physicochemical properties, and phosphorylation site(s) of the 6 phospho-beta-d-glucosides have been determined by fast atom bombardment-negative ion spectrometry, thin-layer chromatography, and (1)H and (13)C NMR spectroscopy. The recently sequenced genomes of two Listeria species, L. monocytogenes EGD-e and L. innocua CLIP 11262, contain homologous genes (lmo2764 and lin2907, respectively) that encode a 294-residue polypeptide (M(r) approximately 32,200) that exhibits approximately 58% amino acid identity with BglK. The protein encoded by the two genes exhibits beta-glucoside kinase activity and cross-reacts with polyclonal antibody to (His6)BglK from K. pneumoniae. The location of lmo2764 and lin2907 within a beta-glucoside (cellobiose):phosphotransferase system operon, may presage both enzymatic (kinase) and regulatory functions for the BglK homolog in Listeria species. PMID- 12110693 TI - Identification, characterization, and functional study of the two novel human members of the semaphorin gene family. AB - We cloned two novel human transmembrane semaphorins, (HSA)SEMA6C and (HSA)SEMA6D, that belong to the class VI subgroup of the semaphorin family. The genes for SEMA6C and SEMA6D are mapped on chromosome 1q12-21.1 and 15q21.1, respectively. Among the adult tissues, SEMA6C is expressed only in skeletal muscle, whereas SEMA6D is expressed abundantly in kidney, brain, and placenta and moderately in the heart and skeletal muscles. During murine development, neither SEMA6C nor SEMA6D was expressed in embryonic day 10.5 (E10.5) embryos, but both were highly expressed in the areas of the lateral ventricle, the striatum, the wall of the midbrain, the pons/midbrain junction, and the choroid plexus of E13 embryos. Were neurons, neither axons nor astrocytes, highly expressed both semaphorins. Three isoforms of SEMA6C and five isoforms of SEMA6D derived from alternative splicing were identified, and their expression was regulated in a tissue- and development dependent manner. Deletion analysis indicated that a sema domain and a PSI domain are integrally necessary for correct post-translation modification and subcellular localization. The extracellular domain of SEMA6C inhibited axonal extension of nerve growth factor-differentiated PC12 cells and induced the growth cone collapse of chicken dorsal root ganglion, rat hippocampal neurons, and rat cortical neurons in a dose-responsive manner. SEMA6D acted like SEMA6C except it had no significant effect on the growth cones of rat cortical neurons. PMID- 12110694 TI - Properties of long myosin light chain kinase binding to F-actin in vitro and in vivo. AB - Short and long myosin light chain kinases (MLCKs) are Ca(2+)/calmodulin-dependent enzymes that phosphorylate the regulatory light chain of myosin II in thick filaments but bind with high affinity to actin thin filaments. Three repeats of a motif made up of the sequence DFRXXL at the N terminus of short MLCK are necessary for actin binding (Smith, L., Su, X., Lin, P., Zhi, G., and Stull, J. T. (1999) J. Biol. Chem. 274, 29433-29438). The long MLCK has two additional DFRXXL motifs and six Ig-like modules in an N-terminal extension, which may confer unique binding properties for cellular localization. Two peptides containing either five or three DFRXXL motifs bound to F-actin and smooth muscle myofilaments with maximal binding stoichiometries consistent with each motif binding to an actin monomer in the filaments. Both peptides cross-linked F-actin and bound to stress fibers in cells. Long MLCK with an internal deletion of the five DFRXXL motifs and the unique NH(2)-terminal fragment containing six Ig-like motifs showed weak binding. Cell fractionation and extractions with MgCl(2) indicate that the long MLCK has a greater affinity for actin-containing filaments than short MLCK in vitro and in vivo. Whereas DFRXXL motifs are necessary and sufficient for short MLCK binding to actin-containing filaments, the DFRXXL motifs and the N-terminal extension of long MLCK confer high affinity binding to stress fibers in cells. PMID- 12110695 TI - Of mice and men: the tale of two therapies. PMID- 12110696 TI - Bone marrow-derived stem cells can differentiate into retinal cells in injured rat retina. AB - It has recently been shown that bone marrow cells can differentiate into various lineage cells including neural cells in vitro and in vivo. We therefore examined whether bone marrow stem cells can differentiate into retinal neural cells in adult rats. PKH-67-labeled stem cell-enriched bone marrow cells (BMCs) were injected into the vitreous space of eyes in which the retinas had been mechanically injured using a hooked needle. Two weeks after the injection of these cells, immunohistochemical examinations were carried out. The stem cell enriched BMCs had been incorporated and had differentiated into retinal neural cells in the injured retina. The stem cell-enriched BMCs had accumulated mainly in the outer nuclear layer around the injured sites. The incorporated cells expressed glial fibrillary acidic protein, calbindin, rhodopsin, and vimentin. These results raise the possibility that stem cell-enriched BMCs have the ability to differentiate into retinal neural cells, and that the injection of stem cell enriched BMCs into the retina would help repair damaged retinal cells. PMID- 12110697 TI - Identification of insulin-producing cells derived from embryonic stem cells by zinc-chelating dithizone. AB - BACKGROUND AND AIMS: Embryonic stem (ES) cells have a pluripotent ability to differentiate into a variety of cell lineages in vitro. We have recently identified the emergence of cellular clusters within differentiated ES cell cultures by staining with dithizone (DTZ). DTZ is a zinc-chelating agent known to selectively stain pancreatic beta cells because of their high zinc content. The aim of the present study was to investigate the characteristics of DTZ-stained cellular clusters originating from ES cells. METHODS: Embryoid bodies (EBs), formed by a 5-day hanging drop culture of ES cells, were allowed to form outgrowths in the culture. The outgrowths were incubated in DTZ solution (final concentration, 100 microg/ml ) for 15 minutes before being examined microscopically. The gene expression of endocrine pancreatic markers was also analyzed by reverse transcriptase-polymerase chain reaction. In addition, insulin production was examined immunohistochemically, and its secretion was examined using enzyme-linked immunosorbent assay. RESULTS: DTZ-stained cellular clusters appeared after approximately 16 days in the EB culture and became more apparent by day 23. They were found to be immunoreactive to insulin and expressed pancreatic-duodenal homeobox 1 (PDX1), proinsulin 1, proinsulin 2, glucagon, pancreatic polypeptide, glucose transporter-2 (GLUT2), and islet-specific glucose 6-phosphatase catalytic subunit-related protein (IGRP) mRNA. They were also able to secrete detectable amounts of insulin. CONCLUSIONS: ES cell-derived DTZ positive cellular clusters possess characteristics of the endocrine pancreas, including insulin secretion. Further, DTZ staining is a useful method for the identification of differentiated pancreatic islets developed from EBs in vitro. PMID- 12110698 TI - Age-related alterations in the lymphohematopoietic and B-lineage precursor populations in NZB mice. AB - Significant disturbances in B lineage populations of New Zealand Black (NZB) mice have been reported, both with respect to their phenotypes as well as to their function. Notably, there is a profound age-dependent decrease in B-cell precursors in this strain of lupus prone mice. In efforts to characterize the impact of this disturbance in disease, we performed an intensive phenotypic and B cell population analysis in young and old NZB mice. Our results revealed that there was a significant age-dependent decrease in B cell precursors at all levels of the B-cell-lineage developmental pathway. Analysis of the proliferative capacity of these cell populations showed a comparative decrease in cycling activity in the B-cell-lineage populations of old NZB mice. Furthermore, these cell subsets were much more susceptible to spontaneous apoptosis when compared with similar populations from age-matched BALB/c or young NZB mice. Since the frequency of cells that express the interleukin-7 receptor (IL-7R) declines as NZB mice age, we hypothesize that impairment of IL-7R signal transduction pathways could contribute to severe perturbations of B-cell function in aged NZB mice. PMID- 12110699 TI - Transplanted hematopoietic cells seed in clusters in recipient bone marrow in vivo. AB - The process of hematopoietic stem and progenitor cell (HSPC) seeding in recipient bone marrow (BM) early after transplantation is not fully characterized. In vivo tracking of HSPCs, labeled with PKH dyes, through an optical window surgically implanted on the mouse femur revealed that transplanted cells cluster in the recipient BM. Within the first day after intravenous injection, 86 +/- 6% of the cells seeded in clusters (p < 0.001 versus scattered cells) in the endosteal surfaces of the epiphyses. The primary clusters were formed by concomitant seeding of 6-10 cells over an area of approximately 70 microm, and secondarily injected cells did not join the already existing clusters but formed new clusters. Major antigen-disparate HSPCs participated in formation of the primary clusters, and T lymphocytes were also incorporated. After 4 to 5 days, some cellular clusters were observed in the more central regions of the BM, where the brightness of PKH fluorescence decreased, indicating cellular division. These later clusters were classified as secondary, assuming that the mechanisms of migration in the BM might be different from those of primary seeding. Some clusters remained in the periphery of the BM and retained bright fluorescence, indicating cellular quiescence. The number of brightly fluorescent cells in the clusters decreased exponentially to two to three cells after 24 days (p < 0.001). The data suggest that the hematopoietic niche is a functional unit of the BM stromal microenvironment that hosts seeding of a number of transplanted cells, which form a cluster. This may be the site where auxiliary non-HSPC cells, such as T lymphocytes, act in support of HSPC engraftment. PMID- 12110700 TI - Treatment of Shwachman syndrome by Japanese herbal medicine (Juzen-taiho-to): stimulatory effects of its fatty acids on hemopoiesis in patients. AB - Juzen-taiho-to (a Japanese herbal medicine) has been traditionally administered to patients with anemia, neutropenia, or wasting syndrome. We previously attempted to isolate and purify the hemopoiesis-stimulatory components in Juzen taiho-to extracts using an in vitro hemopoietic stem cell (HSC) assay method in which mouse HSCs can proliferate on a stromal cell line (MS-5). We have found that fatty acids (particularly oleic acid and linolenic acid) actively promote the proliferation of HSCs, and that the effect is mediated by stromal cells, rather than by any direct action on the HSCs. In the present study, we show, using human normal bone marrow cells (BMCs) and umbilical cord blood cells, that similar stimulatory effects are due to the presence of oleic acid and linolenic acid, which stimulate the proliferation of HSCs in stroma-based culture systems. Furthermore, a marked stimulatory effect was noted on BMCs from patients with Shwachman syndrome, which shows pancreatic and bone marrow dysfunctions. We also show the data on hemopoietic recovery after the administration of Juzen-taiho-to to a patient with Shwachman syndrome. These findings suggest that decreased fatty acid levels in the blood, caused by exocrine pancreatic insufficiency, induce bone marrow dysfunction in Shwachman syndrome. PMID- 12110701 TI - Higher pH promotes megakaryocytic maturation and apoptosis. AB - Megakaryocytic (Mk) cells mature adjacent to bone marrow (BM) sinus walls and subsequently release platelets within the sinusoidal space or in lung capillaries. In contrast, primitive stem and Mk progenitor cells reside the furthest away from the BM sinus walls. The existence of pH gradients in the BM raises the question of whether pH affects Mk maturation and differentiation. We generated Mk cells from peripheral blood CD34(+) cells in a serum-free medium at different pH levels (7.2, 7.4, and 7.6) and found that higher pH resulted in an earlier and higher polyploidization of CD41(+) Mk cells and an earlier onset of Mk-cell apoptosis. The peak day of high ploidy was correlated well with the onset day of Mk apoptosis, thus suggesting that a decline in the fraction of high ploidy Mk cells at the late culture stage is caused by Mk-cell apoptosis. We further explored the relationship between Mk-cell maturation and apoptosis by employing an antiapoptotic agent Z-Val-Ala-Asp(Ome)-FMK (zVAD). Addition of zVAD led to an average 30% higher and 2.8-day delayed polyploidization, while apoptosis was delayed by 2.4 days. Faster depletion of CD34(+) cells and an earlier peak in the fraction of larger colony-forming Mk cells (BFU-Mks) were also observed at higher pH. Taken together, these data suggest that higher pH promotes Mk-cell differentiation, maturation, and apoptosis. PMID- 12110702 TI - Preimplantation human embryos and embryonic stem cells show comparable expression of stage-specific embryonic antigens. AB - Cell-surface antigens provide invaluable tools for the identification of cells and for the analysis of cell differentiation. In particular, stage-specific embryonic antigens that are developmentally regulated during early embryogenesis are widely used as markers to monitor the differentiation of both mouse and human embryonic stem (ES) cells and their malignant counterparts, embryonic carcinoma (EC) cells. However, there are notable differences in the expression patterns of some such markers between human and mouse ES/EC cells, and hitherto it has been unclear whether this indicates significant differences between human and mouse embryos, or whether ES/EC cells correspond to distinct cell types within the early embryos of each species. We now show that human ES cells are characterized by the expression of the cell-surface antigens, SSEA3, SSEA4, TRA-1-60, and TRA-1 81, and by the lack of SSEA1, and that inner cell mass cells of the human blastocyst express a similar antigen profile, in contrast to the corresponding cells of the mouse embryo. PMID- 12110703 TI - AFP(+), ESC-derived cells engraft and differentiate into hepatocytes in vivo. AB - A major problem in gene therapy and tissue replacement is accessibility of tissue specific stem cells. One solution is to isolate tissue-specific stem cells from differentiating embryonic stem (ES) cells. Here, we show that liver progenitor cells can be purified from differentiated ES cells using alpha-fetoprotein (AFP) as a marker. By knocking the green fluorescent protein (GFP) gene into the AFP locus of ES cells and differentiating the modified ES cells in vitro, a subpopulation of GFP(+) and AFP-expressing cells was generated. When transplanted into partially hepatectomized lacZ-positive ROSA26 mice, GFP(+) cells engrafted and differentiated into lacZ-negative and albumin-positive hepatocytes. Differentiation into hepatocytes also occurred after transplantation of GFP(+) cells in apolipoprotein-E- (ApoE) or haptoglobin-deficient mice as demonstrated by the presence of ApoE-positive hepatocytes and ApoE mRNA in the liver of ApoE deficient mice or by haptoglobin in the serum and haptoglobin mRNA in the liver of haptoglobin-deficient mice. This study describes the first isolation of ES cell-derived liver progenitor cells that are viable mediators of liver-specific functions in vivo. PMID- 12110704 TI - Hypoxia modifies proliferation and differentiation of CD34(+) CML cells. AB - We previously showed that hypoxia (1% O(2)) favors the self-renewal of murine and human normal hematopoietic stem cells. This study represents the first attempt to characterize the effects of hypoxia on the maintenance of chronic myeloid leukemia (CML) progenitors. CD34(+) cells isolated from apheresis products of CML patients were incubated in hypoxia (1% O(2)) and normoxia (20% O(2)). After 8 days of culture, their proliferation, capacity for colony-forming-cell (CFC) generation in secondary cultures (pre-CFC), and phenotype (CD34 and platelet activating factor receptor [PAF-R]) were compared with those of normal cells, and tyrosine phosphorylation in CML cells was measured. Hypoxia inhibits the proliferation of CD34(+) cells and preserves the pre-CFC capacity and cell surface CD34 expression of CML cells better than normoxia. The PAF-R expression, which was absent on freshly isolated cells, was detected at the cell surface in both populations after 8 days of culture, but with a lower percentage of positive cells in CML cell cultures. Incubation in hypoxia suppressed the PAF-R expression of normal cells and increased it in CML cells, resulting in a similar expression in the two populations. These effects could be linked to inhibition by hypoxia of the tyrosine hyperphosphorylation of cellular proteins, a major hallmark of CML cells. PMID- 12110705 TI - The molecular perspective: Bcl-2 and apoptosis. PMID- 12110707 TI - High-resolution CT and CT angiography of peripheral pulmonary vascular disorders. AB - Peripheral pulmonary vascular disorders that can be evaluated with computed tomography (CT) include various disease entities with overlapping imaging features and a wide range of clinical manifestations. The overall accuracy of CT in the diagnosis of pulmonary vascular disorders increases with improved spatial resolution, administration of a high-flow contrast material bolus, and the use of cardiac gating. The integration of high-resolution CT and CT angiographic techniques into one scanning protocol has important clinical implications for multisection CT and makes it the imaging modality of choice in the evaluation of this complex group of disorders. PMID- 12110708 TI - MR imaging of the postoperative knee: a pictorial essay. AB - Magnetic resonance (MR) imaging of the postoperative knee has become more common because more arthroscopic repair procedures are being performed. The most common procedures include partial meniscectomy and meniscal repair, anterior cruciate ligament (ACL) reconstruction, and cartilage repair procedures. Specific findings of a retorn meniscus following meniscal repair or partial meniscectomy are increased signal intensity extending through the site of repair on T2-weighted images, displaced meniscal fragments, and abnormal signal intensity at a site distant from the repair. Findings of ACL graft disruption on T2-weighted MR images include absence of intact graft fibers and increased signal intensity similar to that of fluid within the expected region of the graft. Partial tears of the graft appear as areas of increased signal intensity affecting a portion of the graft with some intact fibers still present. An impinged ACL graft may appear to be draped over the anterior inferior edge of the intercondylar roof or be posteriorly bowed. Localized anterior arthrofibrosis appears on T1-weighted MR images as a focal nodular lesion of low signal intensity that is anterior to the ACL graft in the intercondylar notch and is indistinguishable from adjacent joint fluid. On T2-weighted images, the nodule is well differentiated from high-signal intensity joint fluid. Finally, MR imaging has been shown to be accurate in the evaluation of cartilage repair tissue. Knowledge of the normal MR imaging appearance of the knee after the more common repair procedures will allow radiologists to recognize complications associated with such procedures. PMID- 12110709 TI - Anatomy and MR imaging appearances of synovial plicae of the knee. AB - Synovial plicae are normal anatomic structures of the knee that sometimes become symptomatic. Magnetic resonance (MR) imaging and MR arthrography are useful tools in the evaluation of synovial plicae and allow differentiation of these entities from other causes of knee pain. At MR imaging, synovial plicae appear as bands of low signal intensity within the high-signal-intensity joint fluid. Gradient-echo T2-weighted and fat-suppressed T2-weighted or proton density-weighted MR images are optimal for the evaluation of plicae. Plica syndrome, the painful impairment of knee function in which the only finding that helps explain the symptoms is the presence of a thickened and fibrotic plica, should be included in the differential diagnosis of internal derangement of the knee. A diffusely thickened synovial plica, perhaps associated with synovitis or erosion of the articular cartilage of the patella or femoral condyle, in a patient with no other significant MR imaging findings suggests the diagnosis of plica syndrome. Once the diagnosis has been made, nonsurgical treatment is preferable initially. Failure of the patient to improve with conservative treatment leaves arthroscopic excision of the pathologic plica as the treatment of choice. PMID- 12110710 TI - Gynecologic causes of acute pelvic pain: spectrum of CT findings. AB - Although ultrasound (US) is the primary imaging modality of choice in the radiologic evaluation of the female patient with acute pelvic pain, the role of computed tomography (CT) in the evaluation of abdominal and pelvic pain continues to expand. CT may be performed if a gynecologic disorder is not initially suspected, if US findings are equivocal, or if the abnormality extends beyond the field of view achievable with the endovaginal probe and further characterization of pelvic disease is required. Many gynecologic disorders that cause acute pelvic pain (eg, uterine disorders, ovarian disorders, endometriosis, pelvic inflammatory disease, postoperative or postpartum complications) demonstrate characteristic CT findings. Familiarity with these CT appearances is important: It will allow the radiologist to guide appropriate treatment of affected patients and may eliminate the need for further imaging evaluation. PMID- 12110711 TI - Sonohysterographic findings of endometrial and subendometrial conditions. AB - Sonohysterography has become the standard test in the evaluation of dysfunctional uterine and postmenopausal bleeding because it allows reliable differentiation between focal and diffuse endometrial and subendometrial lesions, with the most common being polyps and submucosal fibroids. An endometrial polyp usually appears as a well-defined, homogeneous, polypoid lesion that is isoechoic to the endometrium with preservation of the endometrial-myometrial interface. Atypical polyps have cystic components, multiplicity, a broad base, and hypoechogenicity or heterogeneity. Submucosal fibroids are usually broad-based, hypoechoic, well defined, solid masses with shadowing and an overlying layer of echogenic endometrium that distorts the endometrial-myometrial interface. Atypical fibroids are pedunculated or have a multilobulated surface. The major advantage of sonohysterography is that it can accurately depict the percentage of the fibroid that projects into the endometrial cavity. Endometrial hyperplasia usually appears as diffuse thickening of the echogenic endometrial stripe without focal abnormality, but occasionally focal hyperplasia can be seen. Endometrial cancer is typically a diffuse process, but early cases can appear as a polypoid mass. Adhesions usually appear as mobile, thin, echogenic bands that bridge a normally distensible endometrial cavity, but occasionally thick, broad-based bands or complete obliteration of the endometrial cavity is seen. Although endometrial lesions have characteristic features, a wide range of appearances is possible, with significant overlap between entities. Radiologists should be familiar with the broad spectrum of findings that may be seen at sonohysterography in both benign and malignant processes to raise the appropriate level of concern and to direct the clinician toward the appropriate means of diagnostic biopsy or surgery. PMID- 12110712 TI - Experience of 4 years with open MR defecography: pictorial review of anorectal anatomy and disease. AB - Functional disorders of the pelvic floor are a common clinical problem. Diagnosis and treatment of these disorders, which frequently manifest with nonspecific symptoms such as constipation or incontinence, remain difficult. Fluoroscopic x ray defecography has been shown to aid in detection of functional and morphologic abnormalities of the anorectal region. With the advent of open-configuration magnetic resonance (MR) imaging systems, MR defecography with the patient in a vertical position became possible. MR defecography permits analysis of the anorectal angle, the opening of the anal canal, the function of the puborectal muscle, and the descent of the pelvic floor during defecation. Good demonstration of the rectal wall permits visualization of intussusceptions and rectoceles. Excellent demonstration of the perirectal soft tissues allows assessment of spastic pelvic floor syndrome and descending perineum syndrome and visualization of enteroceles. MR defecography with an open-configuration magnet allows accurate assessment of anorectal morphology and function in relation to surrounding structures without exposing the patient to harmful ionizing radiation. PMID- 12110713 TI - Atypical low-signal-intensity renal parenchyma: causes and patterns. AB - Certain renal diseases manifest as low signal intensity of the renal parenchyma on magnetic resonance images. Sometimes, the appearance is sufficiently characteristic to allow a specific radiologic diagnosis to be made. The causes of this finding can be classified into three main categories on the basis of the pathophysiology: hemolysis, infection, and vascular disease. The first category includes paroxysmal nocturnal hemoglobinuria (PNH), hemosiderin deposition in the renal cortex from mechanical hemolysis, and sickle cell disease. The second category includes hemorrhagic fever with renal syndrome (HFRS). The third category includes acute renal vein thrombosis, renal cortical necrosis, renal arterial infarction, rejection of a transplanted kidney, and acute nonmyoglobinuric renal failure with severe loin pain and patchy renal vasoconstriction. These disease processes have different patterns of low signal intensity. PNH, hemosiderin deposition from mechanical hemolysis, and sickle cell disease involve the entire cortex including the columns of Bertin. HFRS involves the medulla, especially the outer medulla, whereas cortical necrosis involves the inner cortex including the columns of Bertin. In renal vein thrombosis, low signal-intensity lesions involve the outer medulla, an appearance resembling that of HFRS. Wedge-shaped low-signal-intensity regions involving both the cortex and the medulla are seen in arterial infarction. PMID- 12110714 TI - Benign regenerative nodules in Budd-Chiari syndrome and other vascular disorders of the liver: radiologic-pathologic and clinical correlation. AB - Large regenerative nodules are benign liver lesions that are frequently seen in Budd-Chiari syndrome and less commonly in other vascular disorders of the liver or systemic conditions such as autoimmune disease, myeloproliferative disorders, and lymphoproliferative disorders. They are usually multiple, with a typical diameter of 0.5-4 cm. At pathologic analysis, large regenerative nodules are well circumscribed, round lesions that may distort the contour of the liver. Only a minority of these nodules are detected at cross-sectional imaging. At multiphasic helical computed tomography, large regenerative nodules are markedly and homogeneously hyperattenuating on arterial dominant phase images and remain slightly hyperattenuating on portal venous phase images. Large regenerative nodules are bright on T1-weighted magnetic resonance images and show the same enhancement characteristics after intravenous bolus administration of gadolinium contrast material. They are predominantly isointense or hypointense relative to the liver on T2-weighted images. There is no evidence that large regenerative nodules degenerate into malignancy. If these nodules are misdiagnosed as multifocal hepatocellular carcinoma, patients might be denied transplantation or offered inappropriately aggressive therapy such as transcatheter arterial chemoembolization. Understanding the clinical setting and imaging appearance of large regenerative nodules can help avoid misdiagnosis as other hypervascular masses. PMID- 12110715 TI - Chronic mesenteric ischemia: imaging and percutaneous treatment. AB - Chronic mesenteric ischemia (CMI) is rare and is often diagnosed late. Fatal malabsorption-related complications or acute ischemic events occur in the absence of treatment. Diagnosis depends on careful acquisition of a medical history and elimination of other conditions. No sensitive and specific tests are available for functional diagnosis of CMI. If other causes of abdominal pain and weight loss have been confidently ruled out, evidence of visceral artery occlusion at noninvasive imaging (Doppler ultrasonography, computed tomographic angiography, and magnetic resonance angiography) suggests CMI. Until the 1990s, open surgery was considered the treatment of choice; percutaneous transluminal angioplasty (PTA) was reserved for patients for whom surgery carried a high risk. However, open surgery carries a nonnegligible risk of morbidity and mortality. In recent years, PTA with stent placement has been recognized as a minimally invasive means of obtaining good long-term results with an acceptable recurrence rate and consequently has been suggested for primary treatment of CMI. New treatments including administration of fibrinolytic agents before PTA of chronic occlusions, routine revascularization of one or more arteries, and stent placement will probably be validated in the near future. Similarly, new data on selection of the best approach will become available soon. PMID- 12110717 TI - Collateral pathways in patients with celiac axis stenosis: angiographic-spiral CT correlation. AB - Although celiac axis stenosis is a frequently encountered occlusive vascular disease, clinically significant ischemic bowel disease caused by celiac axis stenosis is rarely reported due to rich collateral circulation from the superior mesenteric artery (SMA). The most important and frequently encountered collateral vessels from the SMA in patients with celiac axis stenosis are the pancreaticoduodenal arcades and the dorsal pancreatic artery. Subtypes of collateral pathways via the dorsal pancreatic artery include a longitudinal pathway between the celiac branches and the SMA or its branches and a transverse pathway to either the splenic or gastroduodenal artery. A communicating channel between the right hepatic artery and the SMA can be a route for collateral circulation. Hepatic artery variants cause the development of unique collateral pathways that have different characteristics depending on the type of variant. These collateral pathways include intrahepatic interlobar collateral vessels, right gastric to left gastric arterial anastomoses, left hepatic to left gastric arterial anastomoses, and peribiliary arterial plexuses. Major collateral pathways in patients with celiac axis stenosis can be identified with spiral CT, and knowledge concerning this collateral circulation may be important for certain medical procedures such as interventional procedures for the management of hepatic tumors, pancreaticobiliary surgery, and liver transplantation. PMID- 12110718 TI - Practice corner: what did the CT scan show? PMID- 12110721 TI - Reviewing RSNA education exhibits for RadioGraphics: the keystone to journal success. PMID- 12110719 TI - Quantification of flow dynamics in congenital heart disease: applications of velocity-encoded cine MR imaging. AB - Velocity-encoded cine (VEC) magnetic resonance (MR) imaging is a valuable technique for quantitative assessment of flow dynamics in congenital heart disease (CHD). VEC MR imaging has a variety of clinical applications, including the measurement of collateral flow and pressure gradients in coarctation of the aorta, differentiation of blood flow in the left and right pulmonary arteries, quantification of shunts, and evaluation of valvular regurgitation and stenosis. After surgical repair of CHD, VEC MR imaging can be used to monitor conduit blood flow, stenosis, and flow dynamics. There are some pitfalls that can occur in VEC MR imaging. These include potential underestimation of velocity and flow, aliasing, inadequate depiction of very small vessels, and possible errors in pressure gradient measurements. Nevertheless, VEC MR imaging is a valuable tool for preoperative planning and postoperative monitoring in patients with CHD. PMID- 12110722 TI - Neuroblastoma. PMID- 12110723 TI - Neuroblastoma, ganglioneuroblastoma, and ganglioneuroma: radiologic-pathologic correlation. AB - Neuroblastoma, ganglioneuroblastoma, and ganglioneuroma are tumors of the sympathetic nervous system that arise from primitive sympathogonia and are referred to collectively as neuroblastic tumors. They arise wherever sympathetic tissue exists and may be seen in the neck, posterior mediastinum, adrenal gland, retroperitoneum, and pelvis. The three tumors differ in their degree of cellular and extracellular maturation; immature tumors tend to be aggressive and occur in younger patients (median age, just under 2 years), whereas mature tumors occur in older children (median age, approximately 7 years) and tend to behave in a benign fashion. The most benign tumor is the ganglioneuroma, which is composed of gangliocytes and mature stroma. Ganglioneuroblastoma is composed of both mature gangliocytes and immature neuroblasts and has intermediate malignant potential. Neuroblastoma is the most immature, undifferentiated, and malignant tumor of the three. Neuroblastoma, however, may have a relatively benign course, even when metastatic. Thus, these neuroblastic tumors vary widely in their biologic behavior. Features such as DNA content, tumor proto-oncogenes, and catecholamine synthesis influence prognosis, and their presence or absence aids in categorizing patients as high, intermediate, or low risk. Treatment consists of surgery and, usually, chemotherapy. Despite recent advances in treatment, including bone marrow transplantation, neuroblastoma remains a relatively lethal tumor, accounting for 10% of pediatric cancers but 15% of cancer deaths in children. PMID- 12110724 TI - Cystic pheochromocytoma. PMID- 12110725 TI - Search for isotropic resolution in CT from conventional through multiple-row detector. AB - Computed tomography (CT) is a method of acquiring and reconstructing the image of a thin cross section on the basis of measurements of attenuation. In comparison with conventional radiographs, CT images are free of superimposing tissues and are capable of much higher contrast due to elimination of scatter. Most of the developments in CT since its introduction can be considered as attempts to provide faster acquisition times, better spatial resolution, and shorter computer reconstruction times. From the early designs, the technology progressed with faster scanning times and higher scanning plane resolution, but true three dimensional (3D) imaging became practical only with helical scanning capabilities. The recent advent of multiple-row detector helical scanners has the capability to produce 3D images that approach the ideal of a true "3D radiograph." Current multiple-row detector scanners can scan 40-cm volume lengths in less than 30 seconds with near-isotropic resolution and image quality that could not be envisioned at the time of Hounsfield's invention. PMID- 12110727 TI - A five-step approach to digital image manipulation for the radiologist. AB - Digital manipulation of images plays a key role in development of multimedia presentations. Five basic steps to digitizing images and preparing them for publication and computer presentation are scanning, correction, editing and labeling, saving files, and producing final output. These steps can be completed with commercially available hardware and image manipulation software (eg, Photoshop). The higher the quality of the original scanned image, the more image data there will be to edit: A good image cannot be created from an inferior scan. The most important functions for properly scanning images are size, resolution, and color. Resolutions of 300 ppi and 72 ppi should be used for print publication and computer presentations, respectively. The higher resolution image has the larger file size. The scanned image should be saved as a TIFF (tagged image file format), which is an uncompressed file type used for printed images. The Joint Photographic Experts Group (JPEG) format compresses the size of the image file but also reduces image quality. The JPEG format is a good choice if a small file size is needed, such as in Web and PowerPoint presentations. If the user needs to save an image as a JPEG file, the image should be edited first and then saved once in JPEG format. With Photoshop, the user can rotate and crop an image; adjust its brightness, contrast, and color; remove unwanted patient information, dust, and scratches; and add text and symbol labels to enhance images for teaching purposes. Digital manipulation can be fast and effective if the user has some basic knowledge and tools. PMID- 12110726 TI - Computer-aided diagnosis scheme for detection of polyps at CT colonography. AB - Colon cancer is one of the leading causes of cancer deaths in the United States. However, most colon cancers can be prevented if precursor colonic polyps are detected and removed. An advanced computer-aided diagnosis (CAD) scheme was developed for the automated detection of polyps at computed tomographic (CT) colonography. A region encompassing the colonic wall is extracted from an isotropic volume data set obtained by interpolating CT colonographic scans along the axial direction. Polyp candidates are detected with computation of three dimensional (3D) geometric features that characterize polyps, followed by extraction of polyps with hysteresis thresholding and fuzzy clustering using these geometric features. The number of false-positive findings is reduced by extracting 3D texture features from polyp candidates and applying quadratic discriminant analysis to the candidates. This CAD scheme was applied in 71 patients who underwent CT colonography, 14 of whom had colonoscopically confirmed polyps (n = 21). At by-patient analysis, sensitivity was 100%, with an average false-positive rate of 2.0 per patient. At by-polyp analysis, the scheme detected 90% of the polyps at the same false-positive rate. This CAD scheme permits accurate detection of suspicious lesions and thus has the potential to reduce radiologists' interpretation time and improve their diagnostic accuracy in the detection of polyps at CT colonography. PMID- 12110728 TI - Image editing with Adobe Photoshop 6.0. AB - The authors introduce Photoshop 6.0 for radiologists and demonstrate basic techniques of editing gray-scale cross-sectional images intended for publication and for incorporation into computerized presentations. For basic editing of gray scale cross-sectional images, the Tools palette and the History/Actions palette pair should be displayed. The History palette may be used to undo a step or series of steps. The Actions palette is a menu of user-defined macros that save time by automating an action or series of actions. Converting an image to 8-bit gray scale is the first editing function. Cropping is the next action. Both decrease file size. Use of the smallest file size necessary for the purpose at hand is recommended. Final file size for gray-scale cross-sectional neuroradiologic images (8-bit, single-layer TIFF [tagged image file format] at 300 pixels per inch) intended for publication varies from about 700 Kbytes to 3 Mbytes. Final file size for incorporation into computerized presentations is about 10-100 Kbytes (8-bit, single-layer, gray-scale, high-quality JPEG [Joint Photographic Experts Group]), depending on source and intended use. Editing and annotating images before they are inserted into presentation software is highly recommended, both for convenience and flexibility. Radiologists should find that image editing can be carried out very rapidly once the basic steps are learned and automated. PMID- 12110729 TI - Herbivory, plant resistance, and climate in the tree ring record: interactions distort climatic reconstructions. AB - To understand climate change, dendrochronologists have used tree ring analyses to reconstruct past climates, as well as ecological processes such as herbivore population dynamics. Such reconstructions, however, have been hindered by a lack of experiments that separate the influences of confounding impacts on tree rings, such as herbivores and the interactions of multiple factors. Our long-term experiments with scale insects on resistant and susceptible pines demonstrate three major points that are important to the application of this commonly used tool. (i) Herbivory reduced tree ring growth by 25-35%. (ii) The impact on ring growth distorted climate reconstruction, resulting in the overestimation of past moisture levels by more than 2-fold. Our data suggest that, if distortion because of herbivory has been a problem in previous reconstructions, estimates of the magnitude of recent climate changes are likely to be conservative. (iii) Our studies support a detectible plant resistance x herbivore x climate interaction in the tree ring record. Because resistance and susceptibility to herbivory are known to be genetically based in many systems, the potential exists to incorporate plant genetics into the field of dendrochronology, where it may be used to screen distortions from the tree ring record. PMID- 12110730 TI - The Epstein-Barr virus oncogene product, latent membrane protein 1, induces the downregulation of E-cadherin gene expression via activation of DNA methyltransferases. AB - The latent membrane protein (LMP1) of Epstein-Barr virus (EBV) is expressed in EBV-associated nasopharyngeal carcinoma, which is notoriously metastatic. Although it is established that LMP1 represses E-cadherin expression and enhances the invasive ability of carcinoma cells, the mechanism underlying this repression remains to be elucidated. In this study, we demonstrate that LMP1 induces the expression and activity of the DNA methyltransferases 1, 3a, and 3b, using real time reverse transcription-PCR and enzyme activity assay. This results in hypermethylation of the E-cadherin promoter and down-regulation of E-cadherin gene expression, as revealed by methylation-specific PCR, real-time reverse transcription-PCR and Western blotting data. The DNA methyltransferase inhibitor, 5'-Aza-2'dC, restores E-cadherin promoter activity and protein expression in LMP1 expressing cells, which in turn blocks cell migration ability, as demonstrated by the Transwell cell migration assay. Our findings suggest that LMP1 down-regulates E-cadherin gene expression and induces cell migration activity by using cellular DNA methylation machinery. PMID- 12110731 TI - Alternative splicing of RGS8 gene determines inhibitory function of receptor type specific Gq signaling. AB - The regulators of G protein signaling (RGS) proteins modulate heterotrimeric G protein signaling. RGS8 is a brain-specific RGS protein of 180 aa. Here we identified a short isoform of RGS8, RGS8S, that arises by alternative splicing. RGS8S cDNA encodes a N terminus of 7 aa instead of amino acids 1-9 of RGS8 and 10 180 of RGS8. The subcellular distribution of RGS8 and RGS8S did not differ significantly in transfected cells. RGS8S accelerated, not as efficiently as RGS8, the turning on and off of Gi/o-mediated modulation of G protein-gated inwardly rectifying K(+) channels in Xenopus oocytes. We next examined the effects of RGS8 and RGS8S on Gq-mediated signaling. RGS8 decreased the amplitude of the response upon activation of m1 muscarinic or substance P receptors, but did not remarkably inhibit signaling from m3 muscarinic receptors. In contrast, RGS8S showed much less inhibition of the response of either of these Gq-coupled receptors. By quantitative analysis of the inhibitory effect and the protein expression level, we confirmed that the difference of inhibitory effect is caused by both the qualitative difference between RGS8 and RGS8S and the quantitative difference of the protein expression level. We also confirmed that the receptor type specificity of inhibition is not caused by the difference of the expression level of the receptors. In summary, we showed that 9 aa in the N terminus of RGS8 contribute to the function to inhibit Gq-coupled signaling in a receptor type specific manner and that the regulatory function of RGS8S is especially diminished on Gq-coupled responses. PMID- 12110732 TI - Overexpression of a splice variant of DNA methyltransferase 3b, DNMT3b4, associated with DNA hypomethylation on pericentromeric satellite regions during human hepatocarcinogenesis. AB - DNA hypomethylation on pericentromeric satellite regions is an early and frequent event associated with heterochromatin instability during human hepatocarcinogenesis. A DNA methyltransferase, DNMT3b, is required for methylation on pericentromeric satellite regions during mouse development. To clarify the molecular mechanism underlying DNA hypomethylation on pericentromeric satellite regions during human hepatocarcinogenesis, we examined mutations of the DNMT3b gene and mRNA expression levels of splice variants of DNMT3b in noncancerous liver tissues showing chronic hepatitis and cirrhosis, which are considered to be precancerous conditions, and in hepatocellular carcinomas (HCCs). Mutation of the DNMT3b gene was not found in HCCs. Overexpression of DNMT3b4, a splice variant of DNMT3b lacking conserved methyltransferase motifs IX and X, significantly correlated with DNA hypomethylation on pericentromeric satellite regions in precancerous conditions and HCCs (P = 0.0001). In particular, the ratio of expression of DNMT3b4 to that of DNMT3b3, which is the major splice variant in normal liver tissues and retains conserved methyltransferase motifs I, IV, VI, IX, and X, showed significant correlation with DNA hypomethylation (P = 0.009). Transfection of human epithelial 293 cells with DNMT3b4 cDNA induced DNA demethylation on satellite 2 in pericentromeric heterochromatin DNA. These results suggest that overexpression of DNMT3b4, which may lack DNA methyltransferase activity and compete with DNMT3b3 for targeting to pericentromeric satellite regions, results in DNA hypomethylation on these regions, even in precancerous stages, and plays a critical role in human hepatocarcinogenesis by inducing chromosomal instability. PMID- 12110733 TI - Global repression of exotoxin synthesis by staphylococcal superantigens. AB - Virulent Staphylococcus aureus strains typically produce and secrete large quantities of many extracellular proteins involved in pathogenesis. Such strains cause the classical staphylococcal lesion--local tissue destruction and aggressive inflammation accompanied by the massive influx of polymorphonuclear leukocytes, leading to the formation of pus. Most strains causing toxic shock syndrome, however, produce and secrete very small quantities of most exoproteins although they elaborate high levels of toxic shock syndrome toxin-1 (TSST-1). These strains cause local infections that are remarkably apurulent although potentially fatal owing to the superantigen. We have analyzed this disparity and have found that TSST-1 itself is a negative global regulator of exoprotein gene transcription. TSST-1 not only represses most exoprotein genes but determines its own high expression level by autorepression. We report also that a second superantigen, enterotoxin B, has similar regulatory properties. PMID- 12110734 TI - Viruses and lymphomas. PMID- 12110735 TI - A controlled trial of arthroscopic surgery for osteoarthritis of the knee. AB - BACKGROUND: Many patients report symptomatic relief after undergoing arthroscopy of the knee for osteoarthritis, but it is unclear how the procedure achieves this result. We conducted a randomized, placebo-controlled trial to evaluate the efficacy of arthroscopy for osteoarthritis of the knee. METHODS: A total of 180 patients with osteoarthritis of the knee were randomly assigned to receive arthroscopic debridement, arthroscopic lavage, or placebo surgery. Patients in the placebo group received skin incisions and underwent a simulated debridement without insertion of the arthroscope. Patients and assessors of outcome were blinded to the treatment-group assignment. Outcomes were assessed at multiple points over a 24-month period with the use of five self-reported scores--three on scales for pain and two on scales for function--and one objective test of walking and stair climbing. A total of 165 patients completed the trial. RESULTS: At no point did either of the intervention groups report less pain or better function than the placebo group. For example, mean (+/-SD) scores on the Knee-Specific Pain Scale (range, 0 to 100, with higher scores indicating more severe pain) were similar in the placebo, lavage, and debridement groups: 48.9+/-21.9, 54.8+/-19.8, and 51.7+/-22.4, respectively, at one year (P=0.14 for the comparison between placebo and lavage; P=0.51 for the comparison between placebo and debridement) and 51.6+/-23.7, 53.7+/-23.7, and 51.4+/-23.2, respectively, at two years (P=0.64 and P=0.96, respectively). Furthermore, the 95 percent confidence intervals for the differences between the placebo group and the intervention groups exclude any clinically meaningful difference. CONCLUSIONS: In this controlled trial involving patients with osteoarthritis of the knee, the outcomes after arthroscopic lavage or arthroscopic debridement were no better than those after a placebo procedure. PMID- 12110736 TI - Regression of splenic lymphoma with villous lymphocytes after treatment of hepatitis C virus infection. AB - BACKGROUND: Some epidemiologic studies suggest a link between hepatitis C virus (HCV) infection and some B-cell non-Hodgkin's lymphomas. We undertook this study after a patient with splenic lymphoma with villous lymphocytes had a hematologic response after antiviral treatment of HCV infection. METHODS: Nine patients who had splenic lymphoma with villous lymphocytes and HCV infection were treated with interferon alfa-2b (3 million IU three times per week) alone or in combination with ribavirin (1000 to 1200 mg per day). The outcomes were compared with those of six similarly treated patients with splenic lymphoma with villous lymphocytes who tested negative for HCV infection. RESULTS: Of the nine patients with HCV infection who received interferon alfa, seven had a complete remission after the loss of detectable HCV RNA. The other two patients had a partial and a complete remission after the addition of ribavirin and the loss of detectable HCV RNA. One patient had a relapse when the HCV RNA load again became detectable in blood. In contrast, none of the six HCV-negative patients had a response to interferon therapy. CONCLUSIONS: In patients with splenic lymphoma with villous lymphocytes who are infected with HCV, treatment with interferon can lead to regression of the lymphoma. PMID- 12110737 TI - Inactivating mutations in the gene for thyroid oxidase 2 (THOX2) and congenital hypothyroidism. AB - BACKGROUND: Several genetic defects are associated with permanent congenital hypothyroidism. Immunologic, environmental, and iatrogenic (but not genetic) factors are known to induce transient congenital hypothyroidism, which spontaneously resolves within the first months of life. We hypothesized that molecular defects in the thyroid oxidase system, which is composed of at least two proteins, might be involved in the pathogenesis of permanent or transient congenital hypothyroidism in babies with defects in iodide organification, for which the oxidase system is required. METHODS: Nine patients were recruited who had idiopathic congenital hypothyroidism (one with permanent and eight with transient hypothyroidism) and an iodide-organification defect and who had been identified by the screening program for congenital hypothyroidism. The DNA of the patients and their relatives was analyzed for mutations in the genes for thyroid oxidase 1 (THOX1 ) and 2 (THOX2 ). RESULTS: The one patient with permanent and severe thyroid hormone deficiency and a complete iodide-organification defect had a homozygous nonsense mutation in the THOX2 gene that eliminates all functional domains of the protein. Three of the eight patients with mild transient congenital hypothyroidism and a partial iodide-organification defect had heterozygous mutations in the THOX2 gene that prematurely truncate the protein, thus abolishing its functional domains. CONCLUSIONS: Biallelic inactivating mutations in the THOX2 gene result in complete disruption of thyroid-hormone synthesis and are associated with severe and permanent congenital hypothyroidism. Monoallelic mutations are associated with milder, transient hypothyroidism caused by insufficient thyroidal production of hydrogen peroxide, which prevents the synthesis of sufficient quantities of thyroid hormones to meet the large requirement for thyroid hormones at the beginning of life. PMID- 12110738 TI - Risk of renal allograft loss from recurrent glomerulonephritis. AB - BACKGROUND: Recurrent glomerulonephritis is a known cause of renal allograft loss; however, the incidence of this complication is poorly defined. We determined the incidence, timing, and relative importance of allograft loss due to the recurrence of glomerulonephritis. METHODS: A total of 1505 patients with biopsy-proved glomerulonephritis received a primary renal transplant in Australia from 1988 through 1997. Recurrence was confirmed by renal biopsy. The Kaplan Meier method was used to estimate the 10-year incidence of allograft failure due to recurrent glomerulonephritis, and this incidence was compared with the incidence of acute rejection, chronic rejection, and death with a functioning allograft. Characteristics of the recipients and donors were examined as potential predictors of recurrence. RESULTS: Allograft loss due to the recurrence of glomerulonephritis occurred in 52 recipients, with a 10-year incidence of 8.4 percent (95 percent confidence interval, 5.9 to 12.0). The type of glomerulonephritis, the sex of the recipient, and the peak level of panel reactive antibodies were independent predictors of the risk of recurrence. Recurrence was the third most frequent cause of allograft loss at 10 years, after chronic rejection and death with a functioning allograft. Despite the effect of recurrence, the overall 10-year incidence of allograft loss was similar among transplant recipients with biopsy-proved glomerulonephritis and among those with other causes of renal failure (45.4 percent [95 percent confidence interval, 40.9 to 50.2] vs. 45.8 percent [95 percent confidence interval, 42.3 to 49.3], P=0.09). CONCLUSIONS: Recurrence is an important cause of allograft loss for those with renal failure due to glomerulonephritis. No risk factors for recurrence were identified that warrant altering the approach to transplantation. However, accurate estimates of risk can now be provided to potential recipients of renal allografts. PMID- 12110739 TI - Images in clinical medicine. Chondrodysplasia punctata. PMID- 12110740 TI - Cystinosis. PMID- 12110742 TI - Debridement and lavage for osteoarthritis of the knee. PMID- 12110741 TI - Case records of the Massachusetts General Hospital. Weekly clinicopathological exercises. Case 21-2002. A 21-year-old man with arthritis during treatment for hyperthyroidism. PMID- 12110743 TI - The reciprocal dance between cancer and development. PMID- 12110745 TI - Retraction: Barbaro et Al. Incidence of dilated cardiomyopathy and detection of HIV in myocardial cells of HIV-positive patients. N Engl J Med 1998;339:1093-9. PMID- 12110744 TI - Is placebo surgery unethical? PMID- 12110746 TI - Outcomes in young adulthood for very-low-birth-weight infants. PMID- 12110747 TI - Oral and topical corticosteroids in bullous pemphigoid. PMID- 12110748 TI - Antibiotic-associated diarrhea. PMID- 12110749 TI - The antiphospholipid syndrome. PMID- 12110750 TI - Pharmacy compliance with a prescription-drug discount program. PMID- 12110751 TI - Case 8-2002: pleural effusion. PMID- 12110752 TI - Ooplasmic transfer. PMID- 12110753 TI - Interferon alfa-induced adverse effects in patients with a psychiatric diagnosis. PMID- 12110754 TI - Absolute fracture risk varies with bone densitometry technique used. A theoretical and in vivo study of fracture cases. AB - The lack of consensus of how the results of peripheral bone mineral density (BMD) measurements should be interpreted is proving a barrier to the wider use of these devices. One approach is to interpret peripheral measurements using thresholds (so-called equivalent T-scores) defined to have the same absolute fracture risk as a femoral neck T-score of -2.5. For this concept to be valid, the estimates of fracture risk for a population should be the same irrespective of the measurement technique used. We tested this prediction both theoretically and in vivo using data for 63 postmenopausal women with Colles fracture and 191 control subjects. The theoretical analysis showed that if the normal population has a Gaussian BMD distribution and fracture risk varies exponentially with Z-score as exp(-beta Z) then patients who experience a low-trauma fracture have a fracture risk that is larger by a factor exp(beta(2)) compared with the fracture risk of the whole population. Using data from the in vivo study, fracture risk predictions were compared for seven different types of measurement (lumbar spine; femoral neck; total hip BMD; and speed of sound [SOS] at the radius, tibia, phalanx, and metatarsal). When quantitative estimates of fracture risk were made for individual subjects, the average risk of fracture for the Colles group varied between 1.03 times larger (for tibial SOS) and 2.77 times larger (for total hip BMD) than the average fracture risk for the whole population. As predicted by the theoretical study, fracture risk varied according to the odds ratio determined by logistic regression analysis. Therefore, estimates of fracture risk derived for the same group of patients varied almost threefold according to the type of measurement. It was concluded that equating estimates of absolute fracture risk for different types of scan should not be used as the basis of deriving equivalent T-scores for interpreting peripheral measurements. PMID- 12110755 TI - Bone mineral density testing in healthy postmenopausal women. The role of clinical risk factor assessment in determining fracture risk. AB - The ease of measurement and the quantitative nature of bone mineral densitometry (BMD) is clinically appealing. Despite BMD's proven capability to stratify fracture risk, data indicate that clinical risk factors provide complementary information on fracture susceptibility that is independent of BMD. Methods to quantify fracture risk using both clinical and BMD variables would have great appeal for clinical decision-making. We describe a procedure for quantifying hip fracture risk (5-yr and remaining lifetime) based on (1) the individual's age alone (base model, assuming average clinical risk factors and bone density), (2) incorporation of multiple patient-specific clinical risk factor data in the base model, and (3) incorporation of both patient-specific clinical risk factor data and BMD results. PMID- 12110756 TI - Comparison of heel ultrasound and finger DXA to central DXA in the detection of osteoporosis. Implications for patient management. AB - The goal of the study was to investigate the potential discordance in patient management when a clinician assumes that a peripheral device is a diagnostic surrogate for central DXA in the detection and treatment of osteoporosis. Over a period of 2 mo, asymptomatic women seeking conventional central DXA evaluation for osteoporosis at a diagnostic imaging center were also evaluated with heel ultrasound and finger DXA peripheral imaging devices. T-Scores of -2.5 or less in screening examinations were used to evaluate the discordance between the two peripheral devices and central DXA in the identification of patients with osteoporosis. Higher T-score cutoffs (>-2.5) were also evaluated. Using central DXA as the standard for comparison, the sensitivity of heel ultrasound for screening cases was 0.34 and specificity was 0.92. For finger DXA, sensitivity was 0.23 and specificity was 0.92. Overall discordance between the peripheral devices and central DXA was 21% (heel) and 23% (finger). Heel ultrasound identified 7 out of every 22 osteoporotic patients diagnosed with central DXA. Finger DXA identified 5 out of every 22 osteoporotic patients. Using lower T scores for the peripheral devices increased sensitivity but markedly increased discordance with DXA. The peripheral devices we studied cannot be considered equivalent surrogates for central DXA in the screening of asymptomatic women for osteoporosis. PMID- 12110757 TI - Efficacy and tolerability of cyclical intravenous pamidronate in patients with low bone mass. AB - Oral bisphosphonates are an established mode of therapy for the prevention and treatment of osteoporosis. However, many patients are unable to take them either because of poor tolerability or some established contraindications. This retrospective study describes our clinical experience with the efficacy and tolerability of cyclical intravenous pamidronate in patients with osteopenia or osteoporosis at the American University of Beirut Medical Center. Twenty patients received intravenous pamidronate, as a 30-mg infusion administered intravenously over 2 h every 3 mo. All patients were maintained on calcium and vitamin D supplementation: 1000 mg of calcium and 400-800 IU of vitamin D, respectively. Bone mineral densities of the spine and/or hip were measured at baseline and within an average of 14 mo of study entry while on cyclical pamidronate. Two thirds of patients had a significant increase in bone mineral density either at the lumbar spine or hip. Five patients (25%) developed the expected acute-phase reaction symptoms. Pamidronate constitutes an attractive alternative therapy in patients who cannot tolerate oral bisphosphonates. PMID- 12110758 TI - Effect of HMG-CoA reductase inhibitors (statins) on bone mineral density. AB - Recent studies have suggested that 3-hydroxy-3-methylglutaryl - coenzyme A (HMG CoA) reductase inhibitors (statins) can increase the bone mineral density (BMD). Our objective was to determine if patients on statin drugs were more likely to have a greater bone mineral density and lower risk of osteoporosis than patients not taking these drugs. A computerized pharmacy system provided complete medication dispensing records for the 983 patients (697 men and 286 women) referred for bone mineral density testing at a single Veterans Affairs Medical Center. In an analysis of covariance model that adjusted for age, body mass index, race, and vitamin use, men using statin drugs were more likely to have a greater BMD of the spine (p < 0.005). The mean difference (effect size) was 0.05 g/cm2 (95% confidence interval of [CL] 0.02-0.09), about 5.3% greater BMD. In women, the association was not significant. The risk of osteoporosis (defined as a T-score < or = -2.5) was determined using logistic regression analysis after adjustment for potential confounding variables. Although not statistically significant, men who received statin drugs for more than 2 yr were approximately half as likely to develop osteoporosis (odds ratio [OR] =.55, 95% CI = 0.28 1.08). A similar effect was observed in women taking statins for any length of time (OR = 0.36, 95% CI = 0.12-1.07). This study suggests that statin drugs may decrease osteoporosis risk, warranting a randomized controlled trial. PMID- 12110759 TI - Ultrasound of the phalanges is not related to a previous fracture. A comparison between ultrasound of the phalanges, calcaneus, and DXA of the spine and hip in 75-year-old women. AB - Recently, an ultrasound (US) device for measurement of amplitude-dependent speed of sound in four proximal phalanges of the hand (DBM Sonic 1200, IGEA, Carpi, Mo, Italy) has been introduced but has not been thoroughly investigated in populations at most risk for fragility fractures (i.e., elderly women). As part of the Malmo Osteoporosis Prospective Risk Assessment study (OPRA), we investigated 1044 randomly selected women, all 75 yr of age, with US of the phalanges and, for comparison, also with two more established methods for bone mass measurement: US of the calcaneus and dual-energy X-ray absorptiometry (DXA) of the hip and spine, both methods having an ability to predict fracture. A self assessment questionnaire was used to obtain information on previous fracture and age at fracture event. We found a low correlation between US of the phalanges and US of the calcaneus speed of sound (SoS) (r = 0.11, p < 0.01), US of the calcaneus (stiffness) (r = 0.09, p < 0.05), DXA of the femoral neck (r = 0.09, p < 0.05), and DXA of the spine (r = 0.10, p < 0.01) and no significant correlation between US of the phalanges and US of the calcaneus broadband ultrasound attenuation (BUA) and DXA trochanter. Also, no differences in US of the phalanges were found when comparing women without any fracture with women with at least one fracture, whereas US of the calcaneus (SoS, BUA, and stiffness) and DXA of the femoral neck, trochanter, and spine were all lower in the women with a fracture history (p < 0.0001). In addition, the precision of the US of the phalanges method was evaluated and found to be lower in these elderly women, compared to the precision reported by others and the manufacturer. In summary, the present data indicate that US of the phalanges is not a usable tool for estimating fracture risk in an elderly female population. PMID- 12110760 TI - Bone mineral assessment with tibial ultrasonometry and dual-energy X-ray absorptiometry in long-term survivors of acute lymphoblastic leukemia in childhood. AB - Acute lymphoblastic leukemia (ALL) in childhood is a serious disease that can affect growth and the attainment of maximal peak bone mass. The latter has recently been recognized as a risk factor for the development of osteoporosis later in life. To determine long-term effects of the disease itself and its treatment, we assessed the bone status of a group of long-term survivors of childhood ALL, all treated with high doses of steroids (dexamethasone) and methotrexate and without cranial irradiation. All 21 subjects enrolled in this cross-sectional study were diagnosed to have non-high-risk precursors acute lymphoblastic leukemia (12 boys and 9 girls, mean age 16.5 yr, range 12.2-25.4 yr). Standard deviation (SD) scores were calculated using a tibial ultrasound device and spinal dual-energy X-ray absorptiometry (DXA) device as bone assessment techniques. SD scores of those two different bone assessment techniques were compared. The mean SOS (speed of sound) SD scores (SDS) of the tibia (mean 0.26, standard deviation [sd] 1.00) were not significantly different from our reference value of 0. There was no significant difference between the SOS SDS in boys and girls. With DXA, no significant difference was seen between the mean BMD SDS and the reference data and no significant difference in BMD between boys and girls was found. The individual mean SDS for bone mineral density (BMD) of lumbar spine are 0.24 (sd 1.02), total body 0.17 (sd 1.00), and apparent BMD (BMAD) 0.07 (sd 1.09). Spearman's correlation between mean SOS SDS and mean BMD of lumbar spine was 0.47, mean SOS SDS and mean BMAD SDS was 0.43, and mean SOS SDS and mean BMD of total body was 0.49. These correlations were significant at the 0.05 level (two tailed). Despite high-dose dexamethasone and methotrexate used for treatment of these children with ALL, no long-term side effects on the bone mineral status of the subjects, measured with DXA or tibial ultrasonometry, could be determined. PMID- 12110761 TI - Frame size, ethnicity, lifestyle, and biologic contributors to areal and volumetric lumbar spine bone mineral density in Indian/Pakistani and American Caucasian premenopausal women. AB - Published data on the spinal bone mineral density (BMD) of premenopausal women originating from the Indian subcontinent (Indian/Pakistani) are few. We compared anteroposterior (AP) and lateral areal BMD (aBMD) using dual X-ray absorptiometry and calculated volumetric BMD (vBMD) in Indian/Pakistani (n = 47) vs American (n = 47) women with dissimilar statures and skeletal sizes. To account for differences, we "adjusted" lumbar aBMD separately for vertebral size (aBMD/the square root of the projected area), height (aBMD/height), and hip skeletal width (aBMD/hip width). We "corrected" bone mineral content (BMC), aBMD, and vBMD for frame size, collectively using height, hip width, and vertebral size. Unadjusted mean aBMD values for AP lumbar (L1-L4, p = 0.0086; L3-L4, p = 0.044) spine were higher in Americans than Indians/Pakistanis,whereas lateral vBMD (p = 0.56) or aBMD (p = 0.060) values were not different. After adjusting for height, hip width, or vertebral size, or correcting for frame size, differences in aBMD disappeared. Regression analyses indicated that the best measures to correct for frame size were: vertebral area for BMC, hip width for aBMD, and vertebral width for lateral vBMD. Height was not significant in any model. In correcting for frame size, we accounted for 73-85% of the variability in BMC, 22-28% in aBMD, and 27% in lateral vBMD. After frame size was corrected, we accounted for 34% of the variability in AP BMC and aBMD, in contrast with 6-9% in the lateral models. Five significant biologic and lifestyle factors remained in AP models; only body weight remained for lateral spine. Upon accounting for frame size using regression, much variability in BMD, aBMD, and vBMD was explained by lifestyle and biologic factors, not by ethnicity. PMID- 12110762 TI - Influence of boron supplementation on vertebral and femoral bone mass in rats on strenuous treadmill exercise. A morphometric, densitometric, and histomorphometric study. AB - We studied the effect of boron supplement on experimental osteopenia caused by strenuous exercise in 93- d-old female Wistar rats. A control group of 15 rats was not manipulated. The exercise group of 30 rats was divided into 2 groups of 15 rats each, one that was fed a diet supplemented with 50 mg/kg of boron in the form of Na(2)B(4)O(7), and other that, did not receive a boron supplement. The length and weight were determined in the femur and fifth lumbar vertebra and the bone mineral content and density were assessed through densitometry, and trabecular bone volume, trabecular number, trabecular thickness, and trabecular separation with histomorphometry. The femur length and weight, and vertebra weight, and femur and vertebra bone mineral content and density were significantly lower and the trabecular separation was higher in the exercise group than in the others (p < 0.005 in all). The femur weight, bone mineral content and density, trabecular bone volume and trabecular thickness, were significantly higher in the exercise plus boron group (p < 0.005 to 0.0001). It was concluded that boron preserves bone mass in rats that have been exposed to intense exercise. PMID- 12110763 TI - A study on the influence of calcified intervertebral disk and aorta in determining bone mineral density. AB - This study utilized dual-energy X-ray absorptiometry (DXA) to determine the association that age-related calcinosis of the aorta and intervertebral disks have in determining bone mineral density (BMD). Eight cadavers were chosen at random and were scanned with DXA before and after the removal of the aorta and intervertebral disks. Our results showed that the removal of sclerotic aortas decreased the vertebral BMD an average of 4.64% and the removal of two lumbar intervertebral disks further decreased BMD an average of 11.93%. These results were deemed significant at the 0.01 level using a Friedman two-way analysis of variance by ranks. It can be concluded that the presence of aortic arteriosclerotic lesions and intervertebral disk chondrocalcinosis add a significant contribution to BMD. PMID- 12110764 TI - Comparative evaluation of the DPX and DPX-IQ lunar densitometer systems following software upgrade. AB - Bone densitometry research departments perform system and software upgrades infrequently in order to maintain high precision. This study compares the results obtained on a Lunar densitometer with DPX, and DPX-IQ installed to achieve year 2000 compliance. The DPX-IQ provides an improved femur edge detection algorithm with an expanded reference database. Two hundred data files for each measurement site acquired on DPX were reanalyzed on DPX-IQ. There was no change to the bone mineral density (BMD), bone mineral content (BMC), T-scores or Z-scores for the L2-L4 spine, radius (ultradistal and 33%), and total body. There was a significant high correlation for the femoral neck BMD (r = 0.98; p < 0.05). The mean differences in BMD, BMC, T-scores, and Z-scores at the femoral neck and Ward's and trochanteric regions were not significant (p > 0.05). The limits of agreement within the 95% confidence interval for the femoral neck BMD using the Bland and Altman method was between -0.057 and 0.063 g/cm(2). This order of magnitude magnifies the long-term precision error and alters the usual confidence limits for interpretation of true change in densitometry practice. Therefore, it is important for reanalysis of DPX data files with the DPX-IQ to be performed so that longitudinal changes in BMD can be accurately assessed. PMID- 12110765 TI - Bone adaptation response to sham and bending stimuli in mice. AB - This study presents inbred-strain-related differences in tibial bone adaptation response to low-force loading in four-point bending and sham (pad pressure) arrangements in mice. Our previous work in mice has shown that at relatively high but equal bending forces (9 N or a bending moment of 16.88 N-mm), C57BL/6J mice respond with significantly greater bone formation than C3H/HeJ mice. Because of high tibial strains, the majority of the bone response in our previous study was woven bone. In this, study, we reduced the loading forces to 5 N or a bending moment of 9.38 N-mm (to decrease the woven-bone formation response) and investigated inbred-strain-related bone adaptation differences resulting from bending and sham loading (reported here for the first time in C57BL/6J) in these mice. Twenty-four female mice within each inbred mouse strain (C3H/HeJ [C3H] and C57BL/6J [B6]) were randomly divided into the two loading groups (12 per group sham and bending, total of 48 mice). All of the external loading was done for 36 cycles at 2 Hz, 3 d/wk for 3 wk. The bone adaptation response at lower forces exhibited a pattern similar to that seen for the higher forces in the previous study, suggesting that the patterns of bone adaptation were inbred strain related and independent of bending force magnitude. The bending-related periosteal mineral apposition surface (pMS) and mineral apposition rate (MAR) were respectively 40% and 45% greater in B6 than in C3H. The cortical bone adaptation response to bending was greater when compared to sham or pad pressure for each inbred strain of mice, suggesting that the majority of the bone adaptation response was the result of bending stimulus and not local pressure from pad contact. In addition, regardless of loading arrangement (sham or bending), the bone adaptation response in C57BL/6J mice was greater than C3H/HeJ. PMID- 12110766 TI - Sphincter of Oddi and acute pancreatitis: a new treatment option. PMID- 12110767 TI - Islet redox stress: the manifold toxicities of insulin resistance, metabolic syndrome and amylin derived islet amyloid in type 2 diabetes mellitus. AB - CONTEXT: Redox stress, reactive oxygen species, reactive nitrogen species, and oxygen free radicals ("toxic oxygen") are increasingly being reported as important cellular signaling mechanisms. It has been known for over a hundred years that type 2 diabetes mellitus is a manifold disease, not only in its etiology, but also in its associated manifold toxicities and multiple complications of the diabetic opathies. The presence of islet amyloid has also been described in association with type 2 diabetes mellitus for a century. OBJECTIVE: This review will attempt to remain focused on the relationship between redox stress, the reactive oxygen species and the reactive nitrogen species in the islet, and how these interact with the multiplicative effect of the toxicities of insulin resistance, metabolic syndrome, amylin (hyperamylinemia), amylin derived islet amyloid and type 2 diabetes mellitus. CONCLUSIONS: Redox sensitive cellular signaling systems play an important role in the development, progressive nature (remodeling) and damaging effects on the beta cell within the islet of the pancreas. Furthermore, redox stress may play an important role in the remodeling and development of islet amyloid creating a space-occupying lesion with a resultant secretory and absorptive defect within the islet. The presence of manifold toxicities necessitates an approach of global risk reduction in the prevention and treatment of type 2 diabetes mellitus. An improved understanding of the dynamic relationship between these toxicities and redox stress within the islet will aid both the researcher and the clinician. PMID- 12110768 TI - Overexpression of the Sm-like proto-oncogene in primary and metastatic pancreatic endocrine tumors. AB - CONTEXT: The cancer associated Sm-like proto-oncogene mRNA has been found to be overexpressed in the majority of pancreatic adenocarcinomas and is necessary for the transformed phenotype in pancreatic cancer cell lines. However, expression levels have not been examined in other types of pancreatic neoplasms, such as pancreatic endocrine tumors. SETTING: Fifteen primary pancreatic endocrine tumors, including five insulinomas and 10 non-functioning tumors, along with seven hepatic metastatic pancreatic endocrine tumors. MAIN OUTCOME MEASURES: Quantitative expression levels of cancer associated Sm-like mRNA were measured by real-time PCR. Overexpression was defined as a two-fold or greater value when compared to the expression levels found in normal pancreatic islet cells obtained from healthy donors. RESULTS: In primary tumors, four of the 10 non-functioning pancreatic endocrine tumors were found to overexpress cancer associated Sm-like mRNA (40%). Three of the five (60%) insulinomas also overexpressed cancer associated Sm-like mRNA. In total, cancer associated Sm-like mRNA was overexpressed in seven of 15 primary tumors (47%) and in the majority (71%, 5 of 7) of the hepatic metastases. CONCLUSIONS: Our results indicate that the cancer associated Sm-like mRNA gene may also play a role in the tumorigenesis of pancreatic endocrine tumors. PMID- 12110769 TI - Endothelial dysfunction in the pathogenesis of atherosclerosis. AB - A healthy endothelium plays a central role in cardiovascular control. Therefore, endothelial dysfunction (ED), which is characterized by an imbalance between relaxing and contracting factors, procoagulant and anticoagulant substances, and between pro-inflammatory and anti-inflammatory mediators, may play a particularly significant role in the pathogenesis of atherosclerosis. ED is closely related to different risk factors of atherosclerosis, to their intensity and their duration. The involvement of risk factors in ED is also supported by results of intervention studies that showed regression of ED with treatment of risk factors. The common denominator whereby different risk factors cause ED is most probably increased oxidative stress and/or inflammation. ED promotes atherosclerosis and probably plays an important role in the development of thrombotic complications in the late stages of the disease. As ED is a key underlying factor in the atherosclerotic process, markers of endothelial abnormalities have been sought. Detection of ED is based on tests of endothelium-dependent vasomotion (dilation capability of peripheral and coronary arteries) and on circulating markers of endothelial function (endothelin-1, von Willebrand factor, tissue plasminogen activator, plasminogen activator inhibitor, adhesion molecules). Using these tests it is possible to follow the dose-response of harmful effects or risk factors, and the effects of preventive procedures on vessel wall function. PMID- 12110770 TI - Carotid atherosclerosis and ischemic stroke in young patients. AB - BACKGROUND: Epidemiological studies indicate a high prevalence of carotid atherosclerosis in elderly patients with ischemic stroke. The aim of this study was to investigate the presence of early carotid atherosclerotic lesions in young subjects with ischemic stroke, in the absence of the common atherosclerotic risk factors. METHODS: We studied 98 young patients with first ischemic stroke (54 males and 44 females; mean age 41.2 years; range 32-50) and 96 healthy controls. All subjects underwent ultrasonographic scanning of the carotid arteries according to a standardized protocol. RESULTS: The carotid intima-media thickness was significantly increased in the patient group (p<0.001) compared with controls. In addition, the prevalence of carotid atherosclerotic plaques was greater in the patients than in the controls (p<0.001). In particular, we detected 18 non-occlusive carotid plaques and 16 thrombotic occlusions. In 8 patients, the lesions were bilateral. The echographic pattern of the plaques was hard in 8 cases, soft in 5 cases, and mixed in the remaining 5 cases. CONCLUSIONS: We detected an increased wall thickness of the carotid arteries and an increased prevalence of carotid atherosclerotic lesions and carotid thrombotic occlusions in young patients with ischemic stroke, with a relative low incidence of cardiovascular risk factors. This finding suggests that arterial intima-media thickness per se is an important determinant of vascular disease in young patients. The data also provide indirect support for the potential role of genetic factors in the genesis of atherosclerosis in young patients. PMID- 12110771 TI - The proximal form of mural thrombus in aortoiliac occlusive disease using computed tomography. AB - BACKGROUND: It is important to know the nature of proximal thrombus in patients with aortoiliac occlusive disease (AIOD) when juxtarenal aortic clamping is scheduled. This study was aimed to evaluate the shape and nature of thrombi at several sites of the abdominal aorta in patients with AIOD using enhanced computed tomography (CT). Final judgment was made according to the operative findings. METHODS: Between the years 1999 and 2001, 22 patients, who underwent aortobifemoral bypass, were enrolled. The shape and nature of their thrombi were examined at 4 points (superior mesenteric, suprarenal, juxtarenal and infrarenal arteries at the level of the 2 cm before the renal artery) and 88 slices of CT were examined retrospectively. RESULTS: There was mural thrombus in 31 slices, which could be classified into 4 shapes (crescent-shaped: 10 cases; magatama: 2; wavy: 12; circular: 6). The wavy and circular shaped thrombi were found to be atheromatous. Nine cases (40.9%) on operative findings were atheroma (wavy: 4; circular shaped: 5). The crescent shape might correspond to fibrin thrombus. Atheromatous thrombus clamping near the renal artery was thought to cause microthromboembolism to surrounding organs. CONCLUSIONS: It is recommended that the more proximal aorta or splanchnic arteries should be temporarily clamped during proximal procedures in patients with wavy or circular shaped thrombi at the juxtarenal aorta to prevent kidney or bowel infarction. PMID- 12110772 TI - Coagulation in diabetic and non-diabetic claudicants. AB - BACKGROUND: In order to compare hemostasis in diabetic and non-diabetic claudicants we evaluated endothelial (von Willebrand factor, vWF), rheologic (fibrinogen, hematocrit), coagulation system (thrombin-antithrombin complex, TAT) and platelet (platelet factor 4, PF4, aggregation on thrombin, collagen and ADP stimulation) parameters in both groups and healthy controls. METHODS: Twenty-five diabetic, 34 non-diabetic patients with claudication and 26 healthy individuals were enrolled into the study. RESULTS: The severity of lower limbs ischemia was similar in two groups of claudicants but coronary heart disease and cerebral ischemia were significantly more common in diabetic than in non-diabetic claudicants. vWF level was significantly higher in diabetic than non-diabetic claudicants and healthy controls (184+/-43%, 147+/-43%, and 103+/-42%, respectively). Fibrinogen was significantly higher in diabetic and non-diabetic claudicants compared to controls (4.2+/-1.7, and 3.9+/-1.1, versus 2.9+/-0.5 g/l) and TAT plasma concentration was much higher in diabetic compare to non-diabetic patients and controls (9.8+/-4.4, 1.7+/-1.1, and 1.3+/-0.6 microg, respectively). PF4 concentration was significantly higher in non-diabetic patients with PAOD (34+/-29 UI/ml) when compare to healthy controls (14+/-9 UI/ml), but diabetic PAOD patients with the disease showed lower PF4 concentration (26+/-30 UI/ml). Platelet aggregation with all used activators was similar in all groups likewise hematocrit values, and platelet count. CONCLUSIONS: Complicated DM is linked with significant endothelial perturbation when compared with healthy, but also with PAOD individuals; rheologic parameters are not different from those found in PAOD patients; coagulation system activation but not platelet hyperactivity is associated with DM complicated by PAOD when compared to both control groups. PMID- 12110773 TI - Role of cytoplasmic calcium in platelet aggregation. AB - BACKGROUND: Although adherence and aggregation of platelets on an active surface such as exposed subendothelial matrix or foreign surfaces is integral to the occlusion of blood vessels, its mode of action is not fully understood. METHODS: The role of cytoplasmic ionized Ca(2+) concentration ([Ca(2+)](i)) in platelet activation induced by contact with a glass surface under shear-stress was studied by employing confocal laser scanning microscopy (CLSM) in conjunction with a parallel plate flow chamber. Changes in [Ca(2+)](i) and morphology of aggregating platelets on glass surface was simultaneously examined. RESULTS: Under static condition, contact with glass caused platelet adhesion to the surface, which was associated with [Ca(2+)](i) rise and morphological change; however, platelets did not develop a large aggregation on the surface. Under lower shear-stress, the number of the single platelets adsorbed on the surface was less than that under static condition. Although shear-stress increased the number of single platelets involved and enhanced morphological change in aggregating platelets in a shear stress related manner, the peak [Ca(2+)](i) value in individual platelets were not increased. CONCLUSIONS: These observations may suggest the crucial roles of shear-stress in platelet aggregate formation at the site of arterial stenosis. Shear-stress might enhances platelet aggregate growth not through the enhancement of [Ca(2+)](i) rise. PMID- 12110774 TI - Subclavian carotid transposition for symptomatic subclavian artery stenosis or occlusion. A comparison with the endovascular procedure. AB - BACKGROUND: Although subclavian-carotid transposition (SCT), among all extrathoracic revascularization procedures, has emerged as the treatment of choice for symptomatic subclavian artery (SA) stenosis or occlusion, some authors advocate percutaneous transluminal angioplasty with stenting as the optimum primary therapy. AIM: to assess safety, efficacy and durability of SCT in the treatment of symptomatic SA stenosis or occlusion. METHODS: DESIGN: review of a prospectively maintained vascular surgical registry. SETTING: University vascular surgical service. PATIENTS: 39 patients requiring surgery for symptomatic stenosis or occlusion of the proximal SA from September 1985 to August 1999. INTERVENTION: SCT. MEASURES: data were collected prospectively from hospital charts and office medical records to determine demographics, risk factors, presenting clinical manifestation, blood pressure differentials, location of the SA lesion and early postoperative outcome. Long-term follow-up was available in all patients. Patency of the revascularization was determined by physical examination and non-invasive laboratory studies. RESULTS: The perioperative mortality and morbidity rates were 2.5% (1 of 39) and 2.5% (1 of 39), respectively. Immediate relief of symptoms was achieved in 100% of cases. Mean follow-up was 6.8 years. Revascularization neither failed during the follow-up period, nor did patients have recurrent symptoms. The overall survival rates at 1, 3, 5 and 10 years were 100%, 100%, 86% and 68%. Overall late mortality rate was 18.4%: no death was stroke related. CONCLUSIONS: SCT is a very safe and effective surgical procedure for the treatment of symptomatic SA atherosclerotic disease, ensuring an excellent long-term patency. PMID- 12110775 TI - Prosthetic grafts for above-knee femoropopliteal bypass. A multicenter retrospective study of 564 grafts. AB - BACKGROUND: Many prosthetic grafts including expanded polytetrafluoroethylene (ePTFE) and polyethylene terephthalate (Dacron) have recently been used for above knee femoropopliteal bypass. The purpose of this study was to identify the factors affecting patency performance and patient survival. METHODS: A multicenter retrospective analysis of 496 patients who received 564 grafts between 1990 and 1999 (325 ePTFE and 239 Dacron). Follow-up extended to 114.5 months, with a mean of 30.8 months (+/-25.9 months). RESULTS: The overall primary patency rate for all grafts was 71.4% at 5 years, 73.7% for ePTFE, and 68.9% for Dacron grafts. The secondary patency rates at 5 years were 84.1% for ePTFE, and 83.8% for Dacron. No significant differences were found. The logistic regression analysis revealed that younger age at operation and smoking history were correlated with decreased primary patency rate. The patency rates were unaffected by postoperative administration of oral anticoagulants or antiplatelet agents, although pharmacotherapy contributed to the improvement of survival rates. Renal failure, cerebral infarction and Dacron decreased survival rate. CONCLUSIONS: We conclude that the patency performances of prosthetic grafts are satisfying. However, the choice of prosthetic grafts for younger patients or patients with a smoking history need to be carefully considered. Cerebral infarction, chronic renal failure and Dacron grafts may decrease the survival rate. The operative indications should be determined carefully in these cases. The administration of beraprost sodium is recommended for postoperative pharmacotherapy. PMID- 12110776 TI - The stiffness of the common carotid artery in patients with Graves' disease. AB - BACKGROUND: Thyroid hormone excess is accompanied by many cardiovascular symptoms. We hypothesised that mechanical properties of the large arteries are also involved in hyperthyroidism-induced cardiovascular changes and set out to investigate this further. METHODS: We compared 2 groups: one composed of hospitalized patients with newly diagnosed, non-treated Graves' disease (Graves' group) and the other (control group) composed of healthy persons or patients with euthyroid goiter (n=25 and n=23, respectively). We determined values of cross sectional compliance (CC(100)), distensibility (DC(100)) and stiffness coefficients (alpha(100)) estimated for blood pressure P(n)=100 mmHg by measuring the blood pressure with the oscillometric method and measuring differences in the diameter of the common carotid artery with ultrasonographic vascular echo Doppler system. RESULTS: We found no differences in CC(100) (10.6 vs 10.3 10( 4)m(2)/Mpa), DC(100) (29.7 vs 29.6 1/Mpa) or alpha(100) (2.67 vs 2.72) between the 2 groups. However, there were strong correlations between all stiffness parameters and plasma thyroid hormone concentrations in the Graves' group (r(s)=0.4698, p<0.001 for DC(100), r(s)=0.4342, p<0.003 for CC(100), r(s)= 0.4698, p<0.001 for alpha(100) and fT3, respectively). This relationships remained significant after statistical corrections for age and lipid levels (r(p)=0.22, p<0.03 for DC(100) and fT3 or r(p)=-0.457, p<0.02 for alpha(100) and fT3, respectively). CONCLUSIONS: These results indicate that in the Graves' group the stiffness of the common carotid artery might be affected by the thyroid hormone level and the lower the level, the more severe the activity of the hyperthyroidism reflected by the plasma concentration of fT3 or fT4. PMID- 12110777 TI - Decreasing plasma endothelin-1 and unchanged plasma neopterin during folate supplementation in hyperhomocysteinemia. AB - BACKGROUND: Hyperhomocysteinemia is a risk factor for atherosclerosis and venous thrombosis, probably exerting its effects through endothelial function. Homocysteine levels are lowered by folate supplementation, and such treatment improves endothelial function. However, whether folate supplementation decreases vascular risk and improves survival is unknown. The aim of this study was to evaluate endothelial function and mononuclear leukocyte inflammatory activity during homocysteine lowering in patients with hyperhomocysteinemia and vascular disease. METHODS: Endothelial function assessed as plasma (p-)endothelin(ET)-1 and intraplatelet cGMP and cAMP, and mononuclear leukocyte inflammatory activity, assessed as p-neopterin were studied during homocysteine lowering in 50 patients with hyperhomocysteinemia and vascular disease, randomized to folate supplementation or no treatment for 3 months. RESULTS: P-homocysteine decreased during the 3 months not only in patients on folate supplementation (from 27 [21 52] to 14 [8-41] micromol/l; p<0.001), but also in the untreated group (from 23 [20-35] to 19 [4-31] micromol/l; p<0.001). P-ET-1 decreased during folate supplementation (from 5.7 [2.7-11.6] to 4.1 [1.8-9.0] pg/ml; p<0.01), but was unchanged in the untreated group 4.1 [2.0-9.5] pg/ml and 4.5 [2.7-7.1] pg/ml). P neopterin was unchanged in patients on folate supplementation (9.7 [5.1-54.4] and 7.6 [5.7-73.0] nmol/l), but increased in the untreated group (from 8.2 [4.7-19.5] to 8.6 [4.6-24.6] nmol/l; p<0.05). Intraplatelet cGMP decreased in patients on folate supplementation (from 0.86 [0.21-2.00] platelets to 0.56 [0.17-1.42] pmol/10(9) platelets; p<0.05), but was unchanged in the untreated group. No significant differences concerning intraplatelet cAMP occurred in either group. CONCLUSIONS: Folate supplementation in hyperhomocysteinemia is associated with decreasing levels of both ET-1 and intraplatelet cGMP, and the absence of an increase in the levels of the inflammatory mediator neopterin. PMID- 12110778 TI - Vascular endothelial growth factor in patients with critical limb ischemia before and after amputation. AB - BACKGROUND: It is known that levels of vascular endothelial growth factor (VEGF) in biological fluids increase with inflammation and vascular proliferation. Theraputic angiogenesis by injection of VEGF or genes encoding for it may be a promising strategy for treatment of critical limb ischemia. However growth factors are also implicated in the development of vascular disease by smooth muscle cell proliferation. METHODS: We have previously shown VEGF levels to be increased in juvenile diabetic subjects with no clinical evidence of vascular disease. We have measured serum VEGF using an enzyme linked immunosorbent assay and cellular VEGF expression using flow cytometry in patients with critical limb ischemia prior to and 6 months postamputation to determine whether removal of the ischemic limb leads to changes in systemic endothelial cell function. RESULTS: Baseline VEGF levels were significantly increased in the patient group compared to controls with levels returning to control levels at 6 months postsurgery. Monocyte and neutrophil VEGF expression was significantly reduced in the patient group. Platelet expression of VEGF was also reduced but this failed to reach statistical significance. CONCLUSIONS: The results suggest that it may be useful to determine the balance between VEGF production and cellular receptor expression prior to treatment. PMID- 12110779 TI - Genetic evaluation of the common variant of the endothelial nitric oxide synthase (Glu(298)->Asp) can be found in a higher degree in patients with TAO compared to healthy controls. METHODS: We enrolled 42 patients with TAO and 149 healthy subjects. RESULTS: Nineteen patients (45.2%) showed homozygosity for Glu(298) versus 76 (51%) in the control group. The heterozygous status for Glu(298)-->Asp was found among 18 patients (42.9%) compared to 61 (40.9%) members of the control group, which was a nearly equal distribution. Homozygosity for the mutant Asp(298) was slightly elevated in the patient group with 11.9 versus 8.1% in the control group. Allele frequency for Asp(298) was 0.333. CONCLUSIONS: Our data do not show elevated homozygosity for the common variant Glu(298)-->Asp in patients with TAO compared to healthy controls. The limitation of our evaluation is the small number of patients, which is a general problem when evaluating patients with TAO, as this is not a common disease. PMID- 12110780 TI - Clinical manifestations of atherosclerosis in an elderly population are related to plasma neopterin, NGAL and endothelin-1, but not to Chlamydia pneumoniae serology. AB - BACKGROUND: Inflammatory mediators secreted by leukocytes are implicated in atherogenesis. Chlamydia (C.) pneumoniae infection has been suggested as a trigger of this process. We investigated relationships between C. Pneumoniae serology, inflammatory mediators and symptomatic cardiovascular disease in old age. METHODS: In a cross-sectional study at baseline with a prospective 4 year follow-up, intraplatelet cyclic 3'-5'adenosine monophosphate (cAMP) and cyclic 3' 5'guanosine monophosphate (cGMP), plasma neutrophil gelatinase associated lipocalin (NGAL), plasma soluble tumor necrosis factor receptor-1 (TNFR-1) plasma neopterin and plasma endothelin-1 (ET-1) were analysed together with IgG and IgA antibodies for C. Pneumoniae in an elderly reference population (n=140, median age 71 years, 71 females). Twenty-one subjects had clinical manifestations of cardiovascular disease at baseline and another 21 were diagnosed with cardiovascular disease during follow-up. RESULTS: In age adjusted logistic regression, subjects with cardiovascular disease showed higher plasma levels of neopterin (p=0.02), NGAL (p=0.04), and ET-1 (p<0.01). If subjects with cardiovascular disease at baseline were excluded from the analysis, higher plasma neopterin (p=0.01) and lower serum HDL cholesterol (p=0.03) predicted cardiovascular disease during follow-up. The presence of IgG or IgA against C. pneumoniae was not associated with cardiovascular disease. Neither were there any associations between inflammatory or endothelial parameters and C. pneumoniae serology. CONCLUSIONS: The inflammatory mediators neopterin and NGAL and endothelial derived vasoconstrictive ET-1 were increased in elderly subjects with symptomatic cardiovascular disease. Increased plasma neopterin predicted cardiovascular disease during follow-up. No relationships were found between C. Pneumoniae serology and cardiovascular disease. PMID- 12110781 TI - Increased plasma total nitric oxide among patients with severe chronic venous disease. AB - BACKGROUND: Skin damage in chronic venous disease (CVD) may be partially attributable to free radical injury including that of nitric oxide (NO). The aim of this study was to measure total plasma NO in patients with CVD compared to control subjects. METHODS: Forty-four patients with CVD and 13 control subjects with no arterial or venous disease were included in this study. The patients underwent duplex ultrasonography to confirm the extent of the venous disease and were assigned to the appropriate CEAP clinical stage. Exhaustive exclusion criteria were applied to prevent the influence of dietary intake and other diseases in total NO production. Patients were studied after resting supine for 10 min in room temperature. Blood samples were taken from the long saphenous or a dorsal foot vein. Plasma was separated and frozen at -80 degrees C within 1 hour of venesection. Total NO was assayed with a colorimetric test using the Griess reaction (R&D systems, UK). RESULTS: The median total NO levels among patients with the C5 was 55 micromol/L (interquartile range 51 to 64); with C4 was 53 micromol/L (interquartile range 44 to 58); C2 and C3 was 44 micromol/L (interquartile range 36 to 50) and with control subjects it was 43 micromol/L (interquartile range 41 to 50). Those with healed ulcers (C5) showed statistically raised NO levels compared to controls (median difference 12 micromol/L [95% confidence interval: 5-22]). Combining the data in the C4 and C5 patients results in a set of data significantly different from control subjects (median difference 9 [95% C.I. 2-15]). CONCLUSIONS: Raised total NO is strongly associated with the more severe stages (lipodermatosclerosis and healed ulceration), in patients with venous disease. PMID- 12110783 TI - Etiology of leg ulcers, healing and recurrence rates in octo- and nonagenarians. AB - BACKGROUND: A high proportion of leg ulcers refractory to ambulatory compression therapy have a mixed etiology. This study evaluates this mixed etiology and the healing and recurrence rate in octo- and nonagenarians. METHODS: The study group comprised 101 patients with 119 legs affected by ulcers. Concomitant diseases, ulcer size, healing time and time for the ulcer to recur were documented. RESULTS: Sixty-four ulcers were of venous origin [healing rate (HR): 45.3%, recurrence rate [(RR): 10.3%], 23 ulcers were complicated by 1 additional disease process (HR: 47.8%, RR: 45.5%), 13 ulcers were complicated by 2 or more concomitant diseases (HR: 46.2%, RR: 16.6%) and 19 ulcers (HR: 26.3%, RR: 20%) were of non-venous origin. CONCLUSIONS: This study showed that venous reflux in combination with local or systemic disease in our elderly patient group increases the chance of recurrence. Non-venous ulcers appear to have a reduced healing rate possibly due to the underlying pathology. PMID- 12110782 TI - Clinical presentation and anatomic distribution of chronic venous insufficiency of the lower limb in a typical Mediterranean population. AB - BACKGROUND: The aim of this study was to demonstrate the characteristics of lower limb chronic venous insufficiency (CVI) in a homogeneous Mediterranean population. METHODS: Investigation of 694 patients with uni- or bilateral symptoms and signs of lower limb CVI using colour duplex scanning. Limbs with previous venous surgery were excluded. The limbs were classified according to history and ultrasonic findings into those with post-thrombotic and those with primary CVI. The clinical presentation according to the CEAP classification was correlated to the anatomic distribution of venous reflux. RESULTS: Most of the symptomatic limbs (537/656, 81.5%) with primary CVI belonged to classes 1 to 3. In these limbs reflux confined to superficial veins was very common (64.5%, 424/656) whereas the prevalence of deep and perforator vein reflux was 18.5 and 25.5%, respectively. In contrast most of the limbs (69.5%) with post-thrombotic CVI belonged to classes 4 to 6, had a complex pattern of reflux, and involvement of deep and perforator veins was common (86.5 and 48%, respectively). In about a quarter (24%) of patients with suspected primary CVI no reflux was found in either limb on duplex scanning. Most of them (48%) had telangiectasis. Bilateral reflux was found in 71% of the patients with primary CVI. CONCLUSIONS: The clinical presentation was worse in limbs with post-thrombotic CVI than in those with a primary disease. Post-thrombotic CVI was associated with a complex pattern of reflux, affecting mostly the deep and perforator veins, whereas superficial reflux was the most common pattern in limbs with primary CVI. Therefore, surgery aiming to eliminate superficial reflux would confer only a minimal benefit in limbs with post-thrombotic CVI but would treat the majority of the limbs with the primary CVI. The high prevalence of bilateral reflux found in patients with primary CVI suggests a bilateral predisposition, which supports the hypothesis of the existence of a generalised venous disease. PMID- 12110784 TI - Changes in the use of health resources by patients with chronic phlebopathies after thermal hydrotherapy. Report from the Naiade project, a nation-wide survey on thermal therapies in Italy. AB - BACKGROUND: Chronic venous disorders carry lifelong medical and social burdens. Within conservative approaches, spa hydrotherapy is popular among patients with venous disorders in Europe, but whether the practice is associated with health or social benefits remains controversial. METHODS: The present work is a substudy of the nation-wide Italian Naiade Project, a large multicenter observational exercise on spa treatments in different disease groups. The "Chronic Phlebopathies" substudy included 2504 patients with primary or secondary varicosis or non-varicose venous insufficiency. After a first visit and administration of a detailed questionnaire, patients underwent a "thermal cycle" of 15-20 days consisting of underwater active and passive physical therapy with mineral waters. The same procedures were repeated after 1 year on the 1352 patients (54%) who spontaneously returned to the same spa. Primary endpoints of the study were some indicators of the use of health resources related to the year after the first thermal cycle, compared with the same indicators recorded at first visit using appropriate statistical methods. RESULTS: The occurrence of acute venous episodes, working days missed, number and duration of hospital admissions, consumption of drugs and physical therapies were all significantly reduced in the year after thermal therapy, thus indicating lesser use of health resources. CONCLUSIONS: The study suggests that thermal hydrotherapy in patients with chronic venous disorders is associated with health and social benefits. PMID- 12110785 TI - Intermittent claudication unmasking underlying Fabry's disease. AB - In a 53-year-old woman, admitted to our Department with leg pain, peripheral arterial occlusive disease (PAOD) was diagnosed. The absence of cardiovascular risk factors in this middle-aged woman, the unexplained burning pain during both effort and rest of the lower extremities mimicking severe ischemia, decreased sweating and cold induced Raynaud's phenomenon raised the suspicion of an underlying predisposing disease. The coexistence of painful acroparesthesias, angiokeratomas, left ventricular hypertrophy (LVH), corneal opacities and lenticular lesions suggested the diagnosis of Fabry's disease, which was confirmed by low serum levels of a-galactosidase-A activity. This case, presented with intermittent claudication due to generalized atherosclerosis, is quite unusual, since Fabry's disease rarely produces symptoms in female carriers. PMID- 12110786 TI - Biological role and clinical implications of mast cells in surgery. PMID- 12110787 TI - Surgeons' tone of voice: a clue to malpractice history. AB - BACKGROUND: Interpersonal aspects of care, such as the communication behaviors of physicians, are often cited as central to patients' decisions to initiate malpractice litigation. Relatively little is known, however, about the impact of the communication behaviors of surgeons. In the current study, we investigated the relationship between judgments of surgeons' voice tone and their malpractice claims history. METHODS: We examined the relationship between surgeons' voice tone during routine office visits and their history of malpractice claims. Surgeons were audiotaped while speaking to their patients during office visits, and very brief samples of the conversations were rated by coders blind to surgeons' claims status. Two 10-second clips were extracted for each surgeon from the first and last minute of their interactions with 2 different patients. Several variables were rated that assessed warmth, hostility, dominance, and anxiety from 10-second voice clips with content and 10-second voice clips with just voice tone. RESULTS: Controlling for content, ratings of higher dominance and lower concern/anxiety in their voice tones significantly identified surgeons with previous claims compared with those who had no claims (odds ratio [OR] 2.74, 95% confidence interval [CI] 1.16 to 6.43 for dominance; OR 0.46, 95% CI 0.21 to 1.01 for concern/anxiety). CONCLUSIONS: Surgeons' tone of voice in routine visits is associated with malpractice claims history. This is the first study to show clear associations between communication and malpractice in surgeons. Specific types of affect associated with claims can be judged from brief audio clips, suggesting that this method might be useful in training surgeons. PMID- 12110789 TI - Invited commentary: financial implications of early hospital discharge and readmission. PMID- 12110788 TI - The effect of decreasing length of stay on discharge destination and readmission after coronary bypass operation. AB - BACKGROUND: Over the decade of the 1990s, hospital stay after operation declined in response to prospective payment and managed care. As a result, complications previously detected and treated in the hospital may have begun to occur after discharge. In addition, discharge to nursing homes and rehabilitation hospitals may have increased. To address these questions, we used a statewide database to look at the use of postacute care and the 30-day readmission and mortality after coronary bypass operation. METHODS: A modification of the Commonwealth of Massachusetts Division of Health Care Finance and Policy discharge data to include a unique patient identifier allowed us to retrospectively track patient destination at discharge and study 30-day readmission to all hospitals in the state. RESULTS: Over the 3-year period after the institution of the unique patient identifier (1993 to 1996), postoperative length of stay after coronary bypass operation decreased from 7.4 to 6 days (19%, P <.0005), but the 30-day readmission rate (17.7%) did not increase. Discharge to rehabilitation hospitals and skilled nursing facilities rose significantly (11.7% to 23.8%), especially in the Medicare population (17.2% to 38.5%). Mortality in the 30 days after discharge remained constant at 0.3%. CONCLUSIONS: A shorter postoperative length of stay did not appear to disadvantage coronary artery bypass patients by increasing their likelihood of readmission or death. Cost savings from reduced length of stay were offset by increased use of postacute services. PMID- 12110790 TI - Radioisotope-navigated video-assisted thoracoscopic operation for ectopic mediastinal parathyroid. PMID- 12110791 TI - The expression of proliferating cell nuclear antigen, p53, p21, and apoptosis in primary gastric lymphoma. AB - BACKGROUND: We have investigated the correlations among the expression of proliferating cell nuclear antigen (PCNA), p53, p21, and the prognosis of primary gastric lymphoma, and we have investigated apoptosis by using transferase mediated deoxyuridine triphosphate nick-end labeling staining. METHODS: A retrospective study was performed on 33 cases of primary gastric lymphoma that were surgically resected. Histopathologic examination was undertaken according to the Working Formulation classification. Immunohistochemical staining was performed by using the avidin-biotin-peroxidase complex method, with anti-PCNA antibody, anti-p53 antibody, and anti-p21 antibody. Apoptosis was quantified in situ from paraffin-embedded specimens by using an in situ apoptosis detection kit. RESULTS: On histologic grade, the survival rate of high-grade type was significantly lower than that of low-grade type and than that of intermediate grade type. The PCNA index of low-grade type was significantly lower than that of intermediate-grade type and than that of high-grade type. Those patients who experienced recurrence were both p53(+) and p21(-). Two of these patients were receiving postoperative chemotherapy but nevertheless experienced recurrence at 18 months and at 5 months after surgery. The PCNA index of p53(+) p21(-) cases was significantly higher than that of p53(-) p21(+) cases. The apoptotic count of p53(-) p21(+) cases was significantly higher than that of p53(+) p21(-) cases. CONCLUSIONS: The Working Formulation classification and the PCNA index were each significant factors predicting the prognosis of primary gastric lymphoma. The prognosis of p53(+) p21(-) cases was poor, and the apoptotic count of p53(+) p21( ) cases was low. Accordingly, the effect of combined chemotherapy for those cases was thought to be poor. PMID- 12110792 TI - Extended lymphadenectomy and preoperative radiotherapy for lower rectal cancers. AB - BACKGROUND: Extended lymphadenectomy including lateral node dissection (EXT-L) contributes to a low incidence of local recurrence of lower rectal cancer. However, EXT-L is frequently associated with impairment of sexual and urinary function. We therefore compared the effectiveness of preoperative radiotherapy with that of EXT-L. METHODS: One hundred fifteen patients were studied. Seventy eight patients underwent preoperative radiotherapy with a total dose of 50 Gy (Rad[+] group), and 37 did not (Rad[-] group). Seventy-five patients received EXT L (EXT-L[+] group), and 40 did not (EXT-L[-] group). Patients were further divided into 4 subgroups (Rad[+]&EXT-L[-], Rad(+)&EXT-L[+], Rad[-]&EXT-L(+), and Rad[-]&EXT-L[-]), and clinicopathologic features were examined. In the Rad(+) group, the relation between the p53 gene and survival was also examined. RESULTS: There was a significant difference in disease-free survival between the Rad(+) and Rad(-) groups (5-year disease-free survival rate, 74.6% vs 45.9%; P =.006). However, there was no significant difference between the Rad(+)&EXT-L[-] and Rad[ ]&EXT-L(+) groups. The p53 gene status did not affect survival in the Rad(+) group. CONCLUSIONS: This study suggests that in terms of curative effect, preoperative radiotherapy can be one alternative therapy in place of EXT-L for patients with lower rectal cancer. PMID- 12110793 TI - Real-time rapid reverse transcriptase-polymerase chain reaction for intraoperative diagnosis of lymph node micrometastasis: clinical application for cervical lymph node dissection in esophageal cancers. AB - BACKGROUND: New molecular techniques have been designed to detect cancer micrometastases that are otherwise missed by conventional histologic examination. The aim of this study was to establish a sensitive and rapid genetic assay to detect lymph node micrometastasis and to assess its usefulness clinically for cervical lymphadenectomy in esophageal cancer. We have recently shown that metastasis in the lymph node chain along the recurrent laryngeal nerves (rec LNs) is a predictor of cervical node metastasis in esophageal cancer. In our retrospective study, the positive rate of cervical lymph node metastasis with rec LNs metastasis was 51.6%, and the rate without rec LNs metastasis was 11.6%. There was a significant difference in both positive rates (P =.0002). METHODS: Rec LNs obtained from 50 patients with esophageal cancer were assessed prospectively by intraoperative histopathologic examination (HE) and genetic analysis. The latter involved a real-time quantitative reverse transcriptase polymerase chain reaction (RT-PCR) system with multiple markers, carcinoembryonic antigen, squamous cell carcinoma, and melanoma antigen-3, whose messenger RNAs are highly and frequently expressed in esophageal cancers. Cervical lymphadenectomy was subsequently performed in a subset of these patients. RESULTS: Ten of 50 patients (20%) were scored as node positive by HE, and 24 patients (48%) were scored positive by genetic diagnosis, including 9 HE-positive cases. Genetic diagnosis of rec LNs accurately predicted all 9 cases with cervical lymph node metastasis and 2 cases with cervical lymph node recurrence, whereas HE missed 2 cases with cervical lymph node metastasis and 2 cases with cervical lymph node recurrence. CONCLUSIONS: Our real-time rapid RT-PCR assay can improve the sensitivity of HE for detection of lymph node metastasis and might be potentially useful for intraoperative genetic diagnosis for subsequent cervical lymphadenectomy in esophageal cancer surgery. PMID- 12110794 TI - Prognostic significance of serum p53 antibody in patients with esophageal squamous cell carcinoma. AB - BACKGROUND: The p53 protein overexpression that usually results from genetic alterations has been reported to induce serum antibodies against p53. There is little information about the clinicopathologic and prognostic significance of preoperative serum p53 antibody in patients with esophageal cancer. METHODS: A highly specific enzyme-linked immunosorbent assay was used to analyze serum p53 antibodies in 105 patients with esophageal squamous cell carcinoma. The cutoff level of 1.3 U/mL was used to indicate seropositive patients, and the cutoff level of 10 U/mL was used to identify high titer patients. At 3 months after surgery, seropositive patients were examined again. RESULTS: A total of 28 patients (26.7%) were positive for serum p53 antibodies. The patients who remained seropositive were more likely to develop tumor recurrence (P =.025). Seropositive patients had worse outcome than seronegative patients. The high titer group had significant association with advanced tumor stages and worse outcomes than the low titer group. High serum p53 antibody titer was an independent prognostic factor (P <.001). CONCLUSIONS: We found that serum p53 antibody was useful to detect esophageal cancer and to identify those with a high risk of tumor recurrence and a poor prognosis. PMID- 12110795 TI - Biliary complications in adult living donor liver transplantation with duct-to duct hepaticocholedochostomy or Roux-en-Y hepaticojejunostomy biliary reconstruction. AB - BACKGROUND: The aim of this study was to compare the incidence of biliary complications after adult living donor liver transplantation (ALDLT) with Roux-en Y hepaticojejunostomy (R-Y HJ) or duct-to-duct hepaticocholedochostomy (D-D HC). METHODS: Biliary complications were reviewed in 20 consecutive ALDLT recipients surviving more than 1 month, including 10 patients who underwent R-Y HJ and 10 patients who underwent D-D HC reconstructions. RESULTS: Ten biliary complications were seen in 8 patients (40%) from the study group. Specifically, 1 case of biliary leakage and 1 case of biliary hemorrhage were observed in the R-Y HJ group (20%), and 2 biliary leakages, 4 biliary strictures, and 2 C-tube related biliary leakages were seen in 6 patients from the D-D HC group (60%). Three of the 5 patients (60%) who underwent right lobe graft ALDLTs experienced biliary stricture. All cases of biliary leakage and biliary hemorrhage were stopped spontaneously by continuous drainage. Three patients in the D-D HC group with anastomotic strictures were successfully treated with percutaneous interventions. Only 1 patient with anastomotic stricture in the D-D HC group with left lobe graft required intrahepatic R-Y HJ reanastomosis. Two cases of C-tube related biliary leakages were treated with endoscopic management. CONCLUSIONS: Biliary complications such as anastomotic strictures were common in the D-D HC group rather than in the R-Y HJ group. D-D HC reconstruction should be applied cautiously, especially in the right lobe graft ALDLT cases. PMID- 12110796 TI - Off-pump coronary artery bypass grafting for patients with three-vessel disease. AB - BACKGROUND: Off-pump coronary artery bypass grafting (CABG) has been performed since 1996 in our institute, and its application has recently been expanded to patients with three-vessel disease. A study was performed to clarify the benefit of off-pump CABG for patients with three-vessel disease. METHODS: Between June 1, 1991 and September 30, 2001, a total of 1089 patients with three-vessel disease (832 men and 257 women; mean age, 64.0 +/- 9.1 years) underwent on-pump CABG. After 1997, a total of 310 patients with three-vessel disease (223 men and 87 women; mean age, 68.8 +/- 8.6 years) underwent off-pump CABG. Data of the historical cohort of on-pump CABG and the concurrent cohort of off-pump CABG were analyzed retrospectively. RESULTS: The 2 groups were age and gender matched. Significant comorbidities were more often observed in the off-pump group than in the on-pump group. The mean number of bypass grafts in the off-pump versus on pump CABG was 3.3 vs 3.7, P <.0001. The mean intubation period, intensive care unit stay, and postoperative hospital stay were 7.9 vs 11.7 hours, 2.2 vs 3.0 days, and 14.5 vs 17.5 days, respectively (P <.0001). In-hospital mortality rate (0.6% vs 1.2%, respectively) and morbidity rates (10.3% vs 12.9%, respectively) were not significantly different. After surgery, calculated event-free rates at 2 years were 93.3% vs 91.9%, respectively; P = not significant. CONCLUSIONS: In patients with multivessel disease, off-pump CABG provided early recovery, and its complication rates and early follow-up results were equivalent to on-pump CABG. PMID- 12110797 TI - Single nucleotide polymorphism (G994-->T) in the plasma platelet-activating factor-acetylhydrolase gene is associated with graft patency of femoropopliteal bypass. AB - BACKGROUND: Plasma platelet-activating factor-acetylhydrolase (PAF-AH) is known to catalyze platelet-activating factor (PAF). The single nucleotide polymorphism (SNP) of plasma PAF-AH gene (G994 -->T in exon 9) is associated with a decreased level of plasma PAF-AH activity. This study analyzed the risk of the SNP on graft occlusion of femoropopliteal bypass in patients with atherosclerotic occlusive disease. METHODS: We retrospectively assessed the patency of 50 above-knee femoropopliteal bypass grafting in 50 patients. Genomic DNA was analyzed for the mutant allele. Plasma PAF-AH activity was measured by radioimmunoassay. RESULTS: The 10-year cumulative primary patency of the bypass was 78.5% in GG (normal genotype) and 50.0% in GT (heterozygous) or TT (homozygous deficient) (P <.05, Kaplan-Meier method). The relative risk of graft failure in GT or TT genotypes was 1.68 (P =.08, Cox proportional hazards model). PAF-AH activity (nmol/min/50 microL) was 1.92 +/- 0.82 in patients with patent grafts and 1.42 +/- 0.47 in those with occluded grafts (mean +/- standard deviation; P <.05, unpaired t test). CONCLUSIONS: The SNP of plasma PAF-AH was associated with a decreased primary graft patency of above-knee femoropopliteal bypass. The risk of graft failure may increase when patients have the SNP. To confirm the independent risk of graft failure by the SNP, further study is necessary and prospective study should be performed. PMID- 12110798 TI - A novel leflunomide derivative, FK778, for immunosuppression after kidney transplantation in dogs. AB - BACKGROUND: Leflunomide and its metabolite, A77 1726, interfere with pyrimidine metabolism and exert potent immunosuppression in experimental organ transplantation. However, clinical use of the agents has been restrained because of an extended half-life. FK778, one synthetic malononitrilamide derived from A77 1726, is synthesized to overcome this problem while maintaining similar therapeutic efficacy. METHODS: Immunosuppressive effect, pharmacokinetics, and adverse events of FK778, as a single drug treatment or combination with tacrolimus or cyclosporine, were determined in a canine kidney transplantation model. The agents were daily administered orally to the animals for 90 days after surgery. Animal survival, pharmacokinetics, biochemistry, hematology, and histopathology were evaluated. RESULTS: FK778 at 4 mg/kg prolonged median survival of the control animals from 10 days to 30.5 days. Administration of 4 mg/kg FK778 with 0.3 mg/kg tacrolimus or 10 mg/kg cyclosporine increased median survival to 75.5 days and 50.5 days, respectively. In combined treatments, trough levels of FK778 at 4 mg/kg ranged between 40 microg/mL and 100 microg/mL after 1 month. Vomiting and diarrhea were common in animals given FK778. Bone marrow suppression was seen at higher doses. CONCLUSIONS: FK778 is a promising new immunosuppressant that may be used in combination with current standard drugs in organ transplantation. PMID- 12110799 TI - Surgical management of intraductal papillary mucinous tumor of the pancreas. AB - BACKGROUND: Intraductal papillary mucinous tumor (IPMT) is a type of pancreatic cystic neoplasm. IPMT consists of intraductal papillary mucinous adenoma (benign IPMT) and intraductal papillary mucinous carcinoma (malignant IPMT). Preoperative diagnosis of malignancy is difficult; the invasiveness and metastatic character are not well known. The purpose of the study was to evaluate the optimal diagnostic and therapeutic strategy of IPMT. METHODS: Medical charts of 38 patients with final diagnosis of IPMT in Kyoto University Hospital were retrospectively reviewed. Preoperative imaging, mode of operation, and clinical and histopathologic findings were analyzed. RESULTS: In 38 IPMTs, imaging of localization was correct in 82% by computed tomography, 90% by ultrasonography, 70% by endoscopic retrograde cholangiopancreatography, 100% by magnetic resonance cholangiopancreatography, and 100% by endoscopic ultrasonography. Evaluation of malignancy by endoscopic ultrasonography resulted in sensitivity and specificity of 81% and 78%, respectively. Pylorus-preserving pancreaticoduodenectomy was preferably performed in 20 of 38 patients with IPMT. Twenty-two patients had histologically malignant disease. Half of them had an invasive component in the adjacent stroma. One case of malignant IPMT showed lymph node metastasis, and the patient had no recurrence after pancreaticoduodenectomy with regional lymphadenectomy. No case was diagnosed as margin positive; however, 27% showed a dysplasia with atypia in the epithelial cells of the cut edge of the pancreas. One patient with negative atypia at the cut edge of the pancreas developed a recurrent tumor in the remnant pancreas. CONCLUSIONS: The preoperative diagnosis of malignancy is difficult, and 50% of malignant IPMT showed an invasive component. Thus, radical resection of the pancreas with regional lymph node dissection should be the choice of treatment. Lymph node metastasis and intraductal distant invasion should be carefully managed in the surgical treatment of these lesions. PMID- 12110800 TI - Peritoneal inflammatory cells in acute pancreatitis: Relationship of infiltration dynamics and cytokine production with severity of illness. AB - BACKGROUND: The purpose of this study was to clarify the still poorly understood dynamics of peritoneal inflammatory cells (PICs) in acute pancreatitis. METHODS: Acute pancreatitis of 3 different degrees of severity was induced in male Wistar rats. Peritoneal lavage was performed at 1, 6, 12, and 24 hours after the induction, and the fluids collected were analyzed for the number and subpopulation of PICs. The levels of apoptosis and necrosis, cytokines, and bacterial infection were also investigated. RESULTS: The number of PICs was increased in mild and moderate pancreatitis, and the infiltration of inflammatory cells had occurred. In severe pancreatitis, the number of PICs increased until 6 hours after the induction, but thereafter the number decreased. Infiltration of neutrophils occurred 6 hours after the induction, but it was not sustained thereafter and infiltration of peritoneal macrophages did not occur. Cytokines in the lavage fluid increased in all 3 models during the first 6 hours after the induction. Subsequently, cytokines were reduced in mild and moderate pancreatitis but significantly increased in severe pancreatitis. The level of bacterial infection increased according to the severity. CONCLUSIONS: The relationship between the PIC dynamics and cytokine levels in severe pancreatitis is very different from that observed in mild or moderate pancreatitis. PMID- 12110801 TI - Indications for abdominal para-aortic lymph node dissection in patients with esophageal squamous cell carcinoma. AB - BACKGROUND: Abdominal para-aortic lymph node (APAL) dissection of esophageal cancer is not widely accepted. The aim of this article is to propose the indications for APAL dissection in esophageal cancer patients from the viewpoint of micrometastases. METHODS: To evaluate the value of APAL dissection in patients with esophageal cancer, the status of APAL metastases and recurrence in 230 patients with esophageal squamous cell carcinoma (1989 to 1998) was examined retrospectively. On the basis of our findings, 16 patients received a prophylactic APAL dissection from January 1999 to March 2001. Micrometastases in the dissected lymph nodes were examined using cytokeratin staining and reverse transcription-polymerase chain reaction of squamous cell carcinoma antigen messenger RNA. RESULTS: Among the 230 patients who had esophageal squamous cell carcinoma, 21 had APAL metastases (including micrometastases) or APAL recurrence. Among the 21 patients with APAL metastases and recurrence, 20 (95.2%) had metastases (including micrometastases) in perigastric lymph nodes (paracardial and lesser curvature nodes). Among 51 patients with lower thoracic esophageal carcinoma, 13 (25.5%) had APAL metastases or recurrence. On the basis of these results, prophylactic APAL dissection was performed in patients with lower thoracic esophageal cancer who were suspected of perigastric lymph node metastases during operations. APAL metastases (including micrometastases) were detected in 6 (38%) of these patients, and 2 patients with APAL micrometastases survived without recurrence. However, 7 patients had hematogenic recurrence after the operation. CONCLUSIONS: Our results suggested that the indications for APAL dissection were limited. Patients with lower thoracic esophageal cancer who are suspected to have perigastric lymph node metastasis and APAL micrometastases may be considered for APAL dissection. PMID- 12110802 TI - Forces for change in surgical training: Unintended consequences. PMID- 12110803 TI - O. H. W. and the gift of total recall. PMID- 12110804 TI - Bouveret's syndrome. PMID- 12110805 TI - Juvenile fibroadenoma of the breast. PMID- 12110806 TI - Leiomyosarcoma arising from the superior mesenteric vein. PMID- 12110808 TI - Communicating esophageal duplication with Barrett's esophagus and high-grade dysplasia. PMID- 12110807 TI - Necrotic infected liver metastasis from colon cancer. PMID- 12110811 TI - The immune tolerance network: a new paradigm for developing tolerance-inducing therapies. AB - Immune tolerance therapies are designed to reprogram immune cells in a highly specific fashion to eliminate pathogenic responses while preserving protective immunity. A concept that has tantalized immunologists for decades, the development of tolerance-inducing therapies, would revolutionize the management of a wide range of chronic and often debilitating diseases by obviating the need for lifelong immunosuppressive regimens. The advances of the past decade have provided a more detailed understanding of the molecular events associated with T cell recognition and activation. Building on these advances, immunologists have demonstrated the feasibility of various tolerance-inducing approaches in small- and large-animal models of autoimmunity, allergy, and transplant graft rejection. Unprecedented opportunities to test these approaches in a variety of human diseases have now emerged. To capitalize on these advances, the National Institutes of Health recently established the Immune Tolerance Network (ITN), an international consortium of more than 70 basic and clinical immunologists dedicated to the evaluation of novel tolerance-inducing therapies and associated studies of immunologic mechanisms. By using a unique interactive approach to accelerate the development of clinical tolerance therapies, the ITN is partnering with the biotechnology and pharmaceutical industries to examine innovative tolerogenic approaches in a range of allergic and autoimmune diseases and to prevent graft rejection after transplantation. Two years since its inception, the ITN now has approximately 2 dozen clinical trials or tolerance assays studies ongoing or in later stages of protocol development. This report summarizes the rationale for emphasizing clinical research on immune tolerance and highlights the progress of the ITN. PMID- 12110810 TI - HIV: clinical manifestations. AB - HIV infection in the United States appeared early in the 1980s, when previously healthy homosexual men manifested opportunistic infections attributable to apparent underlying immunodeficiency. After these initial isolated reports, there appeared many other groups of patients at risk for development of this devastating disease. From these meager beginnings, the problem has escalated exponentially. HIV infection can affect every system in the human body. Since the era of highly active antiretroviral therapy, however, the prevalence of opportunistic infections and HIV-AIDS clinical manifestations has declined dramatically. In addition to antiretroviral therapy, management of HIV-infected persons requires knowledge of the extent of system involvement, as well as highly active antiretroviral therapy-related adverse effects, so as to recognize complications and initiate appropriate intervention. In the following review we will attempt to comprehensively summarize the clinical manifestations of HIV infection for both pediatric and adult populations. PMID- 12110812 TI - Rethinking doctrine: bronchitis, eosinophils, and bronchoscopy in pediatric asthma. PMID- 12110813 TI - Vicilins: a case study in allergen pedigrees. PMID- 12110814 TI - Exhaled breath condensate: an evolving tool for noninvasive evaluation of lung disease. AB - Exhaled breath condensate (EBC) contains aerosolized airway lining fluid and volatile compounds that provide noninvasive indications of ongoing biochemical and inflammatory activities in the lung. Rapid increase in interest in EBC has resulted from the recognition that in lung disease this easily sampled fluid has measurable characteristics that differ prominently from health. These assays have provided evidence of airway and lung redox deviation, acid-base status, and degree and type of inflammation in acute and chronic asthma, chronic obstructive pulmonary disease, adult respiratory distress syndrome, occupational diseases, and cystic fibrosis. Characterized by uncertain and variable degrees of dilution, EBC does not provide precise assessment of individual solute concentrations within native airway lining fluid. However, it can provide useful information when concentrations differ substantially between health and disease or are based on ratios of solutes found in the sample. Because they can be used to measure the targets of modern therapy, EBC assays are likely to become integral components of future clinical studies, and after further technical work is accomplished, they might be used to diagnose and monitor therapy in individual patients. PMID- 12110815 TI - Material safety data sheets: are they reliable in identifying human hazards? AB - The material safety data sheet (MSDS) is an integral part of a worker's evaluation for suspected occupational asthma and dermatitis. However, established US federal guidelines for creating an MSDS do not require that certain key information relevant to the diagnosis of these disorders be included. This rostrum is intended to highlight the limitations of MSDSs as they pertain to the diagnosis of occupational asthma and occupational dermatitis so that future consideration can be given to modification of the existing MSDS guidelines. This article summarizes the origins of MSDS documents, provides an overview of their format, and discusses some of their inherent limitations, which at times impede proper medical evaluation by physicians and other health care professionals. MSDSs are an essential part of making the workplace a safer environment. More complete disclosure about both irritation and sensitization risks in these documents would facilitate the evaluation of workers for OA and OD. Their current ambiguity often delays the diagnosis of these occupational diseases and places the worker at further risk for development of occupational-related long-term disorders. Health care professionals have an obligation to better educate themselves regarding the interpretation of MSDSs and to recognize that they sometimes provide incomplete data. PMID- 12110816 TI - The risk of hospitalization in patients with asthma switched from an inhaled corticosteroid to a leukotriene receptor antagonist. AB - Asthma-related hospitalization rates were compared over a 2-year period between a cohort of patients with asthma who switched from an inhaled corticosteroid in year 1 to a leukotriene modifier in year 2 (n = 285) and a matched cohort continuously treated with an inhaled corticosteroid (n = 570). During year 1, patients were well maintained, with a hospitalization rate of 1.1% to 1.4%. During year 2, 2.5% of the patients switched to a leukotriene modifier had one or more asthma-related hospitalizations compared with 0.6% of the patients continuously receiving an inhaled corticosteroid. Patients treated with a leukotriene modifier were at 7 times greater risk for an asthma-related hospitalization compared with patients who continued to receive an inhaled corticosteroid (risk-adjusted odds ratio, 7.1; 95% CI, 2.79-17.95). These data are consistent with the results of well-controlled clinical trials showing that leukotriene modifiers may be associated with deterioration of asthma control relative to inhaled corticosteroids. Considered in aggregate, the data support the conclusion that leukotriene modifiers should not be substituted for inhaled corticosteroids as a single-controller therapy for asthma. PMID- 12110817 TI - Clinical significance of bronchoalveolar eosinophils in childhood asthma. AB - To evaluate the relationship between clinical parameters and differential cell counts, bronchoalveolar lavage was performed in 79 asthmatic infants and children with unusual asthma. Multivariate analysis showed significant associations between (1) allergic asthma and the presence of alveolar eosinophils and (2) persistent and longer asthma and an increased number of alveolar neutrophils. Our results provide the first evidence that in asthmatic infants and children eosinophilic inflammation is related to allergic sensitization. PMID- 12110818 TI - Switching from conventional to extrafine aerosol beclomethasone dipropionate therapy in children: a 6-month, open- label, randomized trial. AB - BACKGROUND: In adults with asthma, hydrofluoralkane-134a beclomethasone dipropionate (HFA-BDP) extrafine aerosol provides equivalent asthma control at half the daily dose of conventional chlorofluorocarbon (CFC)-BDP. OBJECTIVE: We sought to compare the efficacy and tolerability of switching from CFC-BDP to HFA BDP at half the daily dose in children with stable asthma. METHODS: This 6-month, open-label, randomized, multicenter study enrolled 520 children aged 5 to 11 years with well-controlled asthma receiving inhaled CFC-BDP or budesonide 200 to 800 microg/d x. (Four hundred fifty-two patients were using doses within the recommended range of 200-400 microg and were analyzed separately.) During a 4 week run-in period, patients used CFC-BDP plus a spacer (CFC-BDP+S) at approximately the same dose as they were using before study entry. Patients were then randomized in a 1:3 ratio to continue on CFC-BDP+S or switch to HFA-BDP Autohaler at half the daily dose. RESULTS: The change from baseline in morning peak expiratory flow was significantly greater in patients receiving 100-200 microg of HFA-BDP compared with those receiving 200-400 microg of CFC-BDP+S at weeks 7 to 8 (8.5 and 0.4 L/min, respectively; P =.014), with continuing improvement in both groups over 6 months (12.2 and 12.4 L/min, respectively, at month 6). There were no significant differences between treatments in mean change from baseline in FEV(1), percentage of days or nights without asthma symptoms, and daily beta-agonist use over the 6-month treatment period. The proportion of patients who had one or more asthma exacerbations, the incidence of adverse events, and the percentage change from baseline in 24-hour urinary free cortisol levels were similar in the 2 treatment groups. CONCLUSIONS: This study confirms that asthma control can be well maintained in children when switching from CFC BDP+S to an HFA-BDP Autohaler at doses as low as 100 to 200 microg/d. PMID- 12110819 TI - Use of inhalant medications with and without chlorofluorocarbon propellants in the United States, 1996-2000. PMID- 12110820 TI - Prevalence of pachycondyla chinensis venom allergy in an ant-infested area in Korea. AB - Recently, immediate allergic reactions, including anaphylaxis, after Pachycondyla chinensis ant stings have been frequently reported in Korea. To estimate the prevalence of these reactions and the sensitization rate to P chinensis, we undertook a visit-questionnaire survey of the 327 adult residents living in a town in an ant-infested area in Korea. Skin prick tests with 6 common inhalant allergens, 3 bee venom allergens, and P chinensis whole body extract were performed on all ant-allergic subjects, on 86 asymptomatic residents, and on 37 controls outside the area. The serum-specific IgE to P chinensis extract was determined by ELISA. Seven subjects (2.1%) reported that they had experienced systemic allergic reactions to P chinensis stings; 4 of them had anaphylactic reactions. Large local reactions occurred in an additional 5 subjects (1.6%). All subjects with systemic allergic reactions had positive skin prick test results to P chinensis extract, whereas 23.3% of asymptomatic residents and 2.7% of the controls showed positive skin prick test results. The serum-specific IgE level was significantly higher in the subjects with systemic reactions than in the subjects with local reactions, the asymptomatic sensitizers, and the nonatopic controls. Sensitization to bee venom was found in 25% of the P chinensis allergic subjects; this was significantly higher than the 3% rate seen in nonsensitized subjects. In conclusion, we report a 2.1% prevalence of systemic allergic reactions after P chinensis stings, based on self-reported symptoms, in an ant-infested area in Korea. PMID- 12110821 TI - Inadequate use of asthma medication in the United States: results of the asthma in America national population survey. AB - BACKGROUND: Studies of the use of anti-inflammatory asthma therapy have been limited to selected populations or have been unable to assess the appropriateness of therapy for individuals. OBJECTIVE: We sought to describe the current use of asthma medication in the United States population and to examine the influence of symptoms and sociodemographics on medication use. METHODS: This study was based on a cross-sectional, national, random-digit-dial household telephone survey in 1998 designed to identify adult patients and parents of children with current asthma. Respondents were classified as having current asthma if they had a physician's diagnosis of asthma and were either taking medication for asthma or had asthma symptoms during the past year. RESULTS: One or more persons met the study criteria for current asthma in 3273 (7.8%) households in which a screening questionnaire was completed. Of these, 2509 persons (721 children <16 years) with current asthma were interviewed. Current use of anti-inflammatory medication was reported by 507 (20.1%). Of these, most were using inhaled corticosteroids (72.5%), with use of antileukotrienes reported by 11.4% and use of cromolyn nedocromil reported by 18.6%. Of persons with persistent asthma symptoms in the past month, 26.2% reported current use of some form of anti-inflammatory medication. In bivariate analysis persons reporting lower income, less education, and present unemployment, as well as smokers, were significantly (P <.001) less likely to report current anti-inflammatory use than were other populations. In a multiple regression model nonsmokers and those of white, non-Hispanic ethnicity, as well as persons reporting less asthma control, were more likely to report current anti-inflammatory medication use. CONCLUSION: In the United States use of appropriate asthma therapy remains inadequate. Strategies to increase use of anti inflammatory therapy among patients with asthma are needed. These might include methods to increase access to asthma care for minorities and the socioeconomically disadvantaged. PMID- 12110822 TI - Maternal asthma, infant feeding, and the risk of asthma in childhood. AB - Controversy surrounds the issue of whether children with asthmatic mothers should be breast-fed. The aim of this study was to investigate whether maternal asthma status alters the association between asthma and breast-feeding. In a cohort study of 2602 West Australian children enrolled before birth and followed prospectively, we collected data on method of infant feeding, maternal asthma (as reported by parental questionnaire), atopy (as measured by skin prick test), and current asthma (defined as a physician's diagnosis of asthma and wheeze in the last year) at 6 years of age. The risk of childhood asthma increased if exclusive breast-feeding was stopped (other milk was introduced) before 4 months (odds ratio, 1.28; 95% CI, 1.01-1.62; P =.038), and this risk was not altered by atopy or maternal asthma status. After adjusting for covariates, exclusive breast feeding for less than 4 months was a significant risk factor for current asthma (odds ratio, 1.35; 95% CI, 1.00-1.82; P =.049). There was no formal statistical interaction between breast-feeding and maternal asthma status (P =.970). In this study maternal asthma status did not modify the association between asthma and breast-feeding duration. We recommend that infants with or without a maternal history of asthma be exclusively breast-fed for 4 months and beyond. PMID- 12110823 TI - The effect of omalizumab on nasal allergic inflammation. AB - BACKGROUND: In sensitized patients, coupling between IgE and FcepsilonRI receptors on mast cells leads to release of proinflammatory mediators and a subsequent influx of inflammatory cells to the affected organ. Omalizumab (Xolair; formerly rhuMAb-E25) binds to circulating IgE, thus preventing induction of the allergic process. OBJECTIVE: We investigated the effect of treatment with omalizumab on seasonal allergic rhinitis and related changes in inflammatory cell numbers in nasal biopsy specimens. METHODS: Patients were randomized to treatment with omalizumab or placebo before the pollen season; the treatment was started and continued during season. Symptoms and use of medication were recorded, and blood samples and nasal biopsy specimens were obtained before and during season. Immunocytochemistry was performed on biopsy sections through use of the following antibodies: anti-CD4, CD8 (T lymphocytes), EG2, and anti-eosinophil peroxidase (eosinophils), anti-tryptase (mast cells), human neutrophil lipocalin (neutrophils), and antibodies against IgE and FcepsilonRI. RESULTS: During the season, blood eosinophils increased in placebo-treated patients but not in omalizumab-treated patients (P =.01); the difference between the treatment groups was significant (P =.04). Free IgE in serum decreased significantly (P =.0002) in omalizumab-treated patients but not in placebo-treated patients; the difference between the groups was significant (P =.0001). In nasal biopsy specimens, the number of eosinophil peroxidase-positive staining cells increased in the placebo treated patients (P =.003) but not in the actively treated patients during the season; the difference between the groups was significant (P =.0001). The number of IgE(+) staining cells decreased significantly in the omalizumab group during the season in comparison with the placebo group (P =.04). CONCLUSION: The clinical benefit of treatment with omalizumab is associated with an anti inflammatory effect on cellular markers in blood and nasal tissue. PMID- 12110824 TI - Maternal vaginal microflora during pregnancy and the risk of asthma hospitalization and use of antiasthma medication in early childhood. AB - BACKGROUND: Infants with wheezing and allergic diseases have a microflora that differs from that of healthy infants. The fetus acquires microorganisms during birth when exposed to the maternal vaginal microflora. It is therefore conceivable that the maternal vaginal microflora might influence the establishment of the infant flora and, as a consequence, the development of wheezing and allergic diseases. OBJECTIVE: We sought to study the associations between the composition of the maternal vaginal microflora and the development of wheezing and asthma in childhood. METHODS: We performed a population-based cohort study in Denmark. Vaginal samples for bacterial analysis were obtained during pregnancy. A total of 2927 women (80% of the invited women) completed the study and had 3003 live infants. Infant wheezing was assessed as one or more hospitalizations for asthma between 0 and 3 years of age. Asthma was assessed as use of 3 or more packages of antiasthma medication between 4 and 5 years of age. RESULTS: Maternal vaginal colonization with Ureaplasma urealyticum during pregnancy was associated with infant wheezing (odds ratio [OR], 2.0; 95% CI, 1.2 3.6), but not with asthma, during the fifth year of life. Maternal colonization with staphylococci (OR, 2.2; 95% CI, 1.4-3.4) and use of antibiotics in pregnancy (OR, 1.7; 95% CI, 1.1-2.6) were associated with asthma during the fifth year of life. CONCLUSION: The composition of the maternal vaginal micro-flora might be associated with wheezing and asthma in the offspring up to 5 years of age. PMID- 12110825 TI - Exposure to birch pollen in infancy and development of atopic disease in childhood. AB - BACKGROUND: The relationship between early allergen exposure, sensitization, and development of atopic disease remains controversial. In 1993, extremely high levels of birch pollen were recorded in Stockholm, Sweden, creating the unique opportunity to study children with different exposures during infancy. OBJECTIVE: We sought to assess the influence of early high-dose exposure to an inhalant allergen (birch pollen) on sensitization and development of atopic disease in children. METHODS: A total of 583 children with atopic heredity born in Stockholm in February through April 1992, 1993, or 1994 were investigated at age 4.5 to 5 years. The children were examined and underwent skin prick testing with inhalant and food allergens. IgE antibodies (RAST) against birch pollen and recombinant birch pollen allergen (rBet v 1) were analyzed in serum. RESULTS: The children born in 1993 (high-dose exposure at 0-3 months) were more often sensitized (ie, positive skin prick test response) to birch pollen than the children born in 1994 (low-dose exposure; 17.8% and 8.8%, respectively; odds ratio [OR], 2.4; 95% CI, 1.2-4.6). A tendency in the same direction was seen for children born in 1992 (high-dose exposure at 12-15 months; OR, 1.7; 95% CI, 0.9-3.2). The results were supported by the RAST analyses. The prevalence of bronchial asthma, allergic rhinoconjunctivitis, and atopic dermatitis did not differ between the birth-year groups. However, the prevalence of pollen- and animal dander-induced allergic asthma was increased in the children born in 1993 (OR, 2.6; 95% CI, 1.2-5.6). An interaction between early high-dose exposure to birch pollen and cat in the household was suggested for sensitization to cat (P =.06). CONCLUSION: Exposure to high levels of birch pollen in infancy increases the risk of sensitization to the same allergen, as well as the risk of allergic asthma. PMID- 12110826 TI - Fexofenadine modulates T-cell function, preventing allergen-induced airway inflammation and hyperresponsiveness. AB - BACKGROUND: Antihistamines have been evaluated for usefulness in the treatment of asthma for more than 50 years. Interest was limited until the introduction of newer compounds that were free of much of the dose-limiting sedation associated with the earlier drugs. OBJECTIVE: In a murine model of allergen-induced airway inflammation and hyperresponsiveness, the efficacy of an H1 receptor antagonist to prevent allergic inflammation and altered airway function was evaluated. METHODS: Mice were sensitized and challenged to an allergen, ovalbumin, which elicited marked airway and tissue eosino-philia and airway hyperresponsiveness. Fexofenadine was administered before challenge, and airway responsiveness to inhaled methacholine, airway and tissue eosinophilia, bronchoalveolar lavage fluid cytokine levels, and serum IgE levels were assayed. In a second group of experiments, sensitized and challenged mice were treated or not treated with fexofenadine before challenge. T cells were isolated from the lungs and adoptively transferred into naive recipients before exposure to limited airway allergen challenge, and lung function and inflammation were evaluated. RESULTS: Fexofenadine treatment of sensitized mice prevented the development of airway hyperresponsiveness in both the primary sensitization and challenge, as well as in the adoptive transfer experiments. These changes were accompanied by decreases in bronchoalveolar lavage and tissue eosinophilia, lymphocyte numbers, and T(H)2 cytokine production. CONCLUSION: The results demonstrate the efficacy of an H1 receptor antagonist in preventing allergen-induced alterations in pulmonary inflammation and airway function. The data support the evaluation of drugs such as fexofenadine in the treatment of allergic asthma. PMID- 12110827 TI - The effect of cysteinyl leukotrienes on growth of eosinophil progenitors from peripheral blood and bone marrow of atopic subjects. AB - BACKGROUND: The accumulation of eosinophils into the peripheral blood and airways of asthmatic subjects is, in part, dependent on cysteinyl leukotrienes (cysLTs). However, the effect of cysLTs on peripheral blood and bone marrow eosinophil pro genitor cells in allergic subjects is not known. OBJECTIVE: The purpose of this study was to evaluate the effects of leukotriene (LT) D(4) and LTE(4) and the cysLT(1) receptor antagonist montelukast on peripheral blood and bone marrow eosinophil-basophil progenitor growth and development in atopic subjects. METHODS: Semisolid methylcellulose cultures for peripheral blood and bone marrow eosinophil-basophil colonies were counted after incubation with or without addition of LTD(4), LTE(4), and montelukast in the presence of suboptimal concentrations of GM-CSF, IL-3, and IL-5. RESULTS: Peripheral blood eosinophil basophil colony-forming unit cultures grown in the presence of GM-CSF and bone marrow eosinophil-basophil colony-forming units grown in the presence of IL-5 were significantly increased by the addition of LTD(4) (0.1 micromol/L). This increase was suppressed by montelukast (1 micromol/L). CONCLUSION: This study has demonstrated that the cysLT LTD(4) can stimulate proliferation of eosinophil hematopoietic progenitor cells in the presence of eosinophilopoietic cytokines. The suppressive effect by montelukast demonstrates that this is a cysLT(1) receptor-mediated effect. PMID- 12110828 TI - Recombinant allergens promote expression of CD203c on basophils in sensitized individuals. AB - BACKGROUND: Traditionally, the diagnosis of type I allergies is based on clinical data, skin test results, and laboratory test results with allergen extracts. During the past few years, several attempts have been made to refine diagnostic assays in clinical allergy by introducing recombinant allergens and novel markers of IgE-dependent cell activation. OBJECTIVES: We have identified the ectoenzyme CD203c as a novel basophil antigen that is upregulated on IgE receptor cross linkage. In this study we applied CD203c and a panel of recombinant allergens to establish a novel basophil test that allows for a reliable quantification of IgE dependent responses at the effector cell level. METHODS: Patients allergic to birch (Bet v 1, n = 15; Bet v 2, n = 8) and grass (Phl p 1, n = 15; Phl p 2, n = 10; Phl p 5, n = 14) pollen allergens, as well as 10 nonallergic donors, were examined. Basophils were exposed to various concentrations of recombinant allergens for 15 minutes and then examined for expression of CD203c by means of flow cytometry. CD203c upregulation was correlated with the increase in CD63. RESULTS: Exposure to recombinant allergens resulted in a dose-dependent increase in expression of CD203c on peripheral blood basophils in sensitized individuals, whereas no increase was seen in healthy control subjects. The effects of the recombinant allergens on CD203c expression were also time dependent. There was a good correlation between allergen-induced upregulation of CD203c and upregulation of CD63 (R = 0.76). CONCLUSION: Flow cytometric quantitation of CD203c on blood basophils exposed to recombinant allergens is a useful approach to determine the allergic state in sensitized individuals and represents a basis for a sensitive novel allergy test. PMID- 12110829 TI - Endotoxins prevent murine IgE production, T(H)2 immune responses, and development of airway eosinophilia but not airway hyperreactivity. AB - BACKGROUND: Contact with immunomodulatory factors, such as LPS, in early infancy is associated with decreased allergen sensitization. OBJECTIVE: We sought to study the effects of systemic or airway exposure with LPS on the development of allergen sensitization, eosinophilic airway inflammation, and increased in vivo airway reactivity (AR) in a mouse model. METHODS: BALB/c mice were systemically sensitized with ovalbumin (OVA) plus adjuvant on days 1 and 14 and challenged through the airways with allergen on days 34 to 36. We performed measurement of OVA-specific IgE serum levels, in vitro T(H)2 cytokine production, differential cell counts in bronchoalveolar lavage fluids, and assessment of in vivo AR to inhaled methacholine by means of barometric whole-body plethysmography. RESULTS: Systemic LPS administration before OVA sensitization reduced OVA-specific IgE serum levels (426 +/- 76 vs 880 +/- 104 U/mL, P <.01), T(H)2 cytokine production by splenic mononuclear cells (IL-4: 0.08 +/- 0.01 vs 0.17 +/- 0.01 ng/mL; IL-5: 1.98 +/- 0.52 vs 4.11 +/- 0.54 ng/mL; P <.01), and extent of airway eosinophilia (total cell counts: 93 vs 376 x 10(3)/mL; eosinophils: 23% vs 51%; P <.01) compared with that in OVA-sensitized mice. Local LPS administration to sensitized mice before airway allergen challenges particularly induced IFN-gamma production by peribronchial lymph node cells in vitro (1718 +/- 315 vs 483 +/- 103 ng/mL, P <.01) associated with reduced airway eosinophilia compared with that seen in OVA sensitized mice. Development of increased AR was not affected by systemic or local LPS exposure. Inhibitory effects of LPS on allergen sensitization and eosinophilic airway inflammation were inhibited by administration of anti-IL-12 antibodies before LPS exposure. CONCLUSION: These data indicate that local and systemic application of LPS modulates systemic and local T(H)1/T(H)2 immune responses in a distinct but similarly IL-12-dependent mode. PMID- 12110830 TI - IgE(+), Kit(-), I-A/I-E(-) myeloid cells are the initial source of Il-4 after antigen challenge in a mouse model of allergic pulmonary inflammation. AB - BACKGROUND: IL-4 is generated within hours after antigen lung challenge and influences events that take place early in the induction of pulmonary inflammation. However, the cells responsible for this early IL-4 production in the lung are unknown. OBJECTIVES: We sought to characterize the initial inflammatory events in the lung after antigen challenge and to identify cells responsible for producing IL-4 at early time points. METHODS: Mice were sensitized with ovalbumin or passive IgE and challenged intranasally. Histologic measures of inflammation were used, and lung tissue cytokine production was analyzed by means of RT-PCR. Cells producing IL-4 were characterized by means of in situ hybridization and flow cytometry. RESULTS: IL-4 mRNA was detectable 100 minutes after challenge in sensitized animals. Blockade of this early IL-4 downregulated vascular cell adhesion molecule 1 mRNA expression and attenuated the early recruitment of eosinophils to the lung. CD4-depleted and mast cell deficient mice both expressed early IL-4. Cellular analysis revealed the presence of IL-4 protein at 100 minutes exclusively in IgE(+) myeloid cells that did not express CD3, Kit, or I-A/I-E. Moreover, IL-4 production induced by means of passive IgE sensitization and abrogated in FcR gamma-chain-deficient mice supports the conclusion that this IL-4 production is dependent on IgE/gamma-chain interaction. CONCLUSION: IL-4 production by an IgE/gamma-chain-dependent mechanism occurs rapidly after allergen challenge. At these early time points, IL 4 is produced by a myeloid cell with the characteristics of a mouse basophil (IgE(+), Kit(-), I-A/I-E(-)). These data thus suggest that strategies targeting basophils should be considered in the treatment of early lung inflammation. PMID- 12110831 TI - T(H)2 cytokine expression in atopic children with otitis media with effusion. AB - BACKGROUND: Otitis media with effusion (OME) is more common in atopic children. Few studies have looked for the presence of inflammatory mediators in the middle ear effusions of this population. OBJECTIVE: We hypothesize that atopic children with OME have a different inflammatory cell and cytokine profile than nonatopic patients with the disease. METHODS: Twenty-six patients with OME undergoing myringotomy and ventilation tube placement were recruited at the McGill University Hospital Center. The atopic status was determined for each patient by using standard skin testing. By means of immunocytochemistry, fluid specimens were assessed for T lymphocytes (CD3), eosinophils (major basic protein), neutrophils (elastase), mast cells (tryptase), and basophils (BB1). By using in situ hybridization, the expression of IL-4, IL-5, and IFN-gamma was assessed. RESULTS: There is a higher percentage of eosinophils and T lymphocytes in atopic patients with OME (n = 8) compared with that seen in nonatopic patients (n = 18, P <.01). There is a higher percentage of neutrophils in nonatopic patients with OME compared with that seen in atopic patients (P <.01). In examining cytokine profiles, there is a higher percentage of cells expressing IL-4 and IL-5 in atopic patients with OME compared with that seen in nonatopic patients (P <.01). CONCLUSION: The predominance of eosinophils, T lymphocytes, and T(H)2 mediators in the middle-ear effusions of atopic children provides evidence that allergy might play a role in the pathogenesis of OME. PMID- 12110832 TI - Quadrupole time-of-flight mass spectrometry: a method to study the actual expression of allergen isoforms identified by PCR cloning. AB - BACKGROUND: Over the past 2 decades, molecular biology has shown that most major allergens exist in multiple isoforms. Very little is known about the relevance of allergen isoforms at the level of expressed protein (ie, actual allergen exposure). OBJECTIVE: The aim of this study was to evaluate the applicability of state-of-the-art quadrupole time-of-flight mass spectrometry (Q-TOF MSMS) to the identification and quantification of allergen isoforms at the protein level. METHODS: Q-TOF MSMS is a mass spectrometric approach for sequencing peptides and proteins. In our study it was applied to recombinant (r)Mal d 1, rBet v 1a and rBet v 1d, and natural (n)Mal d 1 from fruits of Malus domestica, cultivar Granny Smith. RESULTS: Q-TOF MSMS allowed sequencing of about 70% of all amino acids in Mal d 1 and about 60% of those in Bet v 1. Mixing experiments with rBet v 1 isoforms and with rMal d 1 and nMal d 1 revealed that the technique allows identification of isoforms in mixtures down to a level of at least 5%. Recombinant Mal d 1 was identified as a Mal d 1a representative, whereas Granny Smith apples were shown to contain Mal d 1b-like allergen isoforms. In this apple cultivar hitherto unreported modifications of Mal d 1b were identified. Q-TOF MSMS allowed semiquantitative measurement of allergen at the femtomole to picomole level. CONCLUSION: Q-TOF MSMS is a powerful tool to find out whether an allergen isoform, detected at the cDNA level, is really expressed in quantities relevant for the allergic patient. PMID- 12110833 TI - Presence of high contents of thymus and activation-regulated chemokine in platelets and elevated plasma levels of thymus and activation-regulated chemokine and macrophage-derived chemokine in patients with atopic dermatitis. AB - BACKGROUND: T(H)2 cells and eosinophils selectively express CC chemokine receptor 4 and CCR3, respectively, and their chemokine ligands are likely to play important roles in the pathogenesis of atopic dermatitis (AD). OBJECTIVE: The purpose of this study was to demonstrate the presence of thymus and activation regulated chemokine (TARC) in platelets and its release during clotting and to evaluate the circulating levels of TARC, macrophage-derived chemokine (MDC), and eotaxin in control subjects and patients with AD. METHODS: We compared plasma and serum contents of TARC, MDC, and eotaxin. We measured TARC contents in platelet lysates. We analyzed the correlation of plasma levels of TARC, MDC, and eotaxin with various clinicolaboratory parameters in patients with AD. RESULTS: Serum contents of TARC rapidly increased during clotting, whereas those of MDC and eotaxin increased only slightly. We demonstrated that platelets contained TARC, and its levels were dramatically elevated in patients with AD. Platelets also released TARC on stimulation with thrombin. We therefore evaluated circulating levels of these chemokines in control subjects and patients with AD by using plasma samples. Plasma TARC levels were significantly increased in patients with AD (P <.0001) and showed significant correlations with severity scoring of atopic dermatitis (SCORAD) index (r = 0.665, P <.00001), serum lactate dehydrogenese levels (r = 0.696, P =.00001), eosinophil counts (r = 0.381, P =.007), and platelet counts (r = 0.562, P <.0001). Similarly, plasma MDC levels were significantly increased in patients with AD (P <.0001) and showed significant correlations with SCORAD index (r = 0.727, P <.0001), serum lactate dehydrogenese levels (r = 0.861, P <.0001), eosinophil counts (r = 0.505, P =.005), and platelet counts (r = 0.370, P =.01). On treatment, plasma TARC and MDC levels were dramatically decreased in accordance with improved SCORAD scores (P =.0012 and P =.0007, respectively). On the other hand, plasma eotaxin levels did not show any significant increase or correlation with any of the clinical parameters in patients with AD. CONCLUSION: Platelets from patients with AD contain high levels of TARC. Thus platelets might play an important role in AD pathogenesis by releasing T(H)2-attracting TARC on activation. Furthermore, circulating levels of TARC and MDC, but not those of eotaxin, correlate well with the disease activity of AD. PMID- 12110834 TI - Environmental tobacco smoke exposure and nocturnal symptoms among inner-city children with asthma. AB - BACKGROUND: Environmental tobacco smoke (ETS) is a frequent exposure and is linked to asthma among inner-city children. OBJECTIVE: We sought to examine the relationship among ETS exposure, select asthma symptoms, and consequences among inner-city children with asthma. METHODS: Data from interviews with primary caregivers of inner-city elementary school children with asthma were evaluated (n = 590). Caregiver reports of child asthma symptoms, exercise limitations, asthma management, health care use, and ETS exposure were examined. RESULTS: Smoking in the home was reported by 29.4% of primary caregivers. ETS exposure (yes/no) was not related to frequency of child nocturnal symptoms or other select asthma morbidity markers. However, among children exposed to ETS, the frequency and severity of child nocturnal symptoms were highest among children exposed to moderate-to-heavy levels of ETS. After controlling for child age, anti inflammatory medication use, asthma primary care, and caregiver's education, exposure to higher levels of ETS was associated with nearly a 3-fold increase in nocturnal symptoms in children (odds ratio, 2.83; 95% CI, 1.22-6.55). CONCLUSION: Among elementary school inner-city children with asthma, exposure to higher levels of ETS was associated with increased frequency of nocturnal symptoms. Reducing the exposure of children with asthma to ETS should be a clear priority in developing effective asthma management plans for inner-city families. PMID- 12110835 TI - Identification of sesame seed allergens by 2-dimensional proteomics and Edman sequencing: seed storage proteins as common food allergens. AB - BACKGROUND: Sesame seed allergy is becoming increasingly prevalent, probably because of its use in international fast-food and bakery products. Despite this fact, few studies have focused on the identification of its major allergenic proteins. OBJECTIVE: The aim of this study was to identify allergenic proteins of sesame seeds (Sesamum indicum). METHODS: Extracted sesame seed proteins were separated by means of SDS-PAGE and 2-dimensional (2-D) PAGE. Immunolabeling was performed with individual patient sera from 20 patients with sesame seed allergy. Selected proteins were further analyzed by means of Edman sequencing. RESULTS: IgE-binding proteins were identified at 78, 52, 45, 34, 32, 29, 25, 20, 9, and 7 kd. Analyzing internal sequences, the protein at 45 kd, which was recognized by 75% of the patients, was found to be a 7S vicilin-type globulin, a seed storage protein of sesame and named Ses i 3. The protein at 7 kd was found to be a 2S albumin, another seed storage protein of sesame and named Ses i 2. Seed storage proteins are known food allergens in peanut, walnut, Brazil nut, and soybean. Interestingly, one known IgE-binding epitope of the peanut allergen Ara h 1 has 80% homology with the corresponding area of Ses i 3. The different amino acids were previously shown not to be critical for IgE binding in Ara h 1. In addition, the proteins at 78 and 34 kd were found to be homologous to the embryonic abundant protein and the seed maturation protein of soybeans, respectively. CONCLUSION: The identification of 4 sesame seed allergens is the first step toward generating recombinant allergens for use in future immunotherapeutic approaches. In addition, the detection of conserved IgE binding epitopes in common food allergens might be a useful tool for predicting cross-reactivity to certain foods. PMID- 12110836 TI - Ana o 1, a cashew (Anacardium occidental) allergen of the vicilin seed storage protein family. AB - BACKGROUND: The allergens responsible for cashew food allergy have not been well characterized. OBJECTIVES: We initiated a study to clone cDNAs encoding cashew food allergens. METHODS: A cashew cDNA library was screened with human serum for IgE-reactive clones and rabbit IgG anti-cashew extract antisera. Reactive clones were sequenced and expressed, and linear epitopes were identified by means of solid-phase overlapping peptide analysis. Immunoblot inhibition was used to identify the native peptide in cashew extract. RESULTS: Four closely related clones reactive with both human and rabbit antisera were sequenced. Sequence analysis showed that these encode members of the vicilin/sucrose-binding protein family of plant seed storage proteins. Screening of the recombinant protein with sera from 20 patients with cashew allergy and 8 cashew-tolerant patients with allergies to other tree nuts showed that 50% and 25% of sera from patients with cashew allergy and cashew-tolerant subjects, respectively, bound the recombinant protein. The corresponding native allergen protein, designated Ana o 1, was located at approximately 50 kd. Epitope mapping revealed 11 linear IgE-binding epitopes, of which 3 appear to be immunodominant. None of the epitopes were shared in common with those of the peanut vicilin allergen Ara h 1. CONCLUSION: Ana o 1, a vicilin-like protein, is a major food allergen in cashews. Cashew and peanut vicilins do not share linear epitopes. PMID- 12110837 TI - Component-resolved diagnosis with recombinant allergens in patients with cherry allergy. AB - BACKGROUND: In pollen-related food allergy, extracts for skin prick tests (SPTs) are often not standardized, and the test reliability is affected by false negative reactions. OBJECTIVE: We sought to evaluate a panel of recombinant allergens (RAs) derived from one allergenic food for use in component-resolved in vivo diagnosis, taking cherry as a model food. METHODS: Seventy-nine subjects were included in the study: 24 Swiss patients (group 1) with a positive double blind placebo-controlled food challenge result to cherries, 23 patients with birch pollen allergy but without cherry allergy (group 2), 23 nonatopic subjects (group 3), and 9 Spanish patients with a history of a cherry allergy (group 4). SPTs were performed in duplicate by using recombinant cherry allergens (Bet v 1 related allergen: recombinant (r) Pru av 1; profilin: rPru av 4; and lipid transfer protein: rPru av 3) in concentrations of 10, 50, and 100 microg/mL. Furthermore, IgE reactivity to rPru av 1, rPru av 4, and rPru av 3 was assessed by means of immunoblot analysis. RESULTS: SPT responses with rPru av 1, rPru av 4, and rPru av 3 were positive in 92%, 17%, and 4% of the patients in group 1; in 74%, 30%, and 0% of the patients in group 2; in 0%, 22%, and 89% of the patients in group 4; and negative for all nonatopic subjects (group 3). Thus the sensitivity of a positive SPT response to at least one of the 3 RAs was 96%. The specificities, negative predictive values, and positive predictive values with the 3 RAs were 100%, 96%, and 100% if calculated in relation to the nonatopic control group but 17%, 79%, and 60% when calculated in relation to the control group with birch pollen allergy. The correlation between SPT and immunoblotting results was excellent. Sensitization to rPru av 3 was associated with more severe symptoms than sensitization to rPru av 1. CONCLUSIONS: SPTs with RAs proved to be highly sensitive for diagnosis of cherry allergy. Component-resolved in vivo diagnosis with standardized amounts of stable RAs allows us to determine sensitization patterns directly, to correlate them with severity of clinical symptoms, and to analyze geographic differences. PMID- 12110838 TI - Venom immunotherapy improves health-related quality of life in patients allergic to yellow jacket venom. AB - BACKGROUND: Venom immunotherapy (VIT) is effective in preventing anaphylactic reactions after insect stings. The effect of VIT on health-related quality of life (HRQL) was studied to evaluate whether this treatment is of importance to patients. OBJECTIVE: We compared HRQL outcomes measured with a disease-specific instrument (Vespid Allergy Quality-of-Life Questionnaire [VQLQ]) in patients allergic to yellow jacket venom treated with VIT or with an adrenalin self administration device (EpiPen) in an open-label, randomized, controlled trial. METHODS: Consenting patients were block randomized to either VIT or EpiPen. Patients received uniform, standardized information, which specified the risk of their condition and the risks and benefits of both treatment options. HRQL measures took place before and after 1 year of treatment with VIT or EpiPen. RESULTS: Seventy-four patients agreed to be randomized, of whom 36 received VIT and 38 an EpiPen. The mean change in VQLQ score in the group randomized to VIT was 1.07 (95% CI, 0.68-1.46), and this improvement was statistically significant (P <.0001) compared with that seen in the group randomized to the EpiPen, in which this change was -0.43 (95% CI, -0.71 to -0.16). These differences were seen in both men and women, persons with more or less general anxiety, and those stung recently and those stung more than a year before their outpatient department visit. The overall proportion of patients receiving benefit from VIT is 0.72, generating a number needed to treat of 1.4. CONCLUSIONS: VIT results in a clinically important improvement in HRQL in patients allergic to yellow jacket venom in all subgroups studied. Of every 3 patients treated with VIT, 2 patients experience an important improvement in their quality of life. PMID- 12110839 TI - Respiratory infections and the prevalence of atopy in children. PMID- 12110840 TI - Wheezing and dog exposure. PMID- 12110842 TI - The C(2)B Ca(2+)-binding motif of synaptotagmin is required for synaptic transmission in vivo. AB - Synaptotagmin is a synaptic vesicle protein that is postulated to be the Ca(2+) sensor for fast, evoked neurotransmitter release. Deleting the gene for synaptotagmin (syt(null)) strongly suppresses synaptic transmission in every species examined, showing that synaptotagmin is central in the synaptic vesicle cycle. The cytoplasmic region of synaptotagmin contains two C(2) domains, C(2)A and C(2)B. Five, highly conserved, acidic residues in both the C(2)A and C(2)B domains of synaptotagmin coordinate the binding of Ca(2+) ions, and biochemical studies have characterized several in vitro Ca(2+)-dependent interactions between synaptotagmin and other nerve terminal molecules. But there has been no direct evidence that any of the Ca(2+)-binding sites within synaptotagmin are required in vivo. Here we show that mutating two of the Ca(2+)-binding aspartate residues in the C(2)B domain (D(416,418)N in Drosophila) decreased evoked transmitter release by >95%, and decreased the apparent Ca(2+) affinity of evoked transmitter release. These studies show that the Ca(2+)-binding motif of the C(2)B domain of synaptotagmin is essential for synaptic transmission. PMID- 12110843 TI - A first-generation linkage disequilibrium map of human chromosome 22. AB - DNA sequence variants in specific genes or regions of the human genome are responsible for a variety of phenotypes such as disease risk or variable drug response. These variants can be investigated directly, or through their non random associations with neighbouring markers (called linkage disequilibrium (LD)). Here we report measurement of LD along the complete sequence of human chromosome 22. Duplicate genotyping and analysis of 1,504 markers in Centre d'Etude du Polymorphisme Humain (CEPH) reference families at a median spacing of 15 kilobases (kb) reveals a highly variable pattern of LD along the chromosome, in which extensive regions of nearly complete LD up to 804 kb in length are interspersed with regions of little or no detectable LD. The LD patterns are replicated in a panel of unrelated UK Caucasians. There is a strong correlation between high LD and low recombination frequency in the extant genetic map, suggesting that historical and contemporary recombination rates are similar. This study demonstrates the feasibility of developing genome-wide maps of LD. PMID- 12110845 TI - Synaptotagmins I and IV promote transmitter release independently of Ca(2+) binding in the C(2)A domain. AB - At nerve terminals, a focal and transient increase in intracellular Ca(2+) triggers the fusion of neurotransmitter-filled vesicles with the plasma membrane. The most extensively studied candidate for the Ca(2+)-sensing trigger is synaptotagmin I, whose Ca(2+)-dependent interactions with acidic phospholipids and syntaxin have largely been ascribed to its C(2)A domain, although the C(2)B domain also binds Ca(2+) (refs 7, 8). Genetic tests of synaptotagmin I have been equivocal as to whether it is the Ca(2+)-sensing trigger of fusion. Synaptotagmin IV, a related isoform that does not bind Ca(2+) in the C(2)A domain, might be an inhibitor of release. We mutated an essential aspartate of the Ca(2+)-binding site of the synaptotagmin I C(2)A domain and expressed it in Drosophila lacking synaptotagmin I. Here we show that, despite the disruption of the binding site, the Ca(2+)-dependent properties of transmission were not altered. Similarly, we found that synaptotagmin IV could substitute for synaptotagmin I. We conclude that the C(2)A domain of synaptotagmin is not required for Ca(2+)-dependent synaptic transmission, and that synaptotagmin IV promotes rather than inhibits transmission. PMID- 12110846 TI - A chemical imbalance. PMID- 12110844 TI - Association of the ADAM33 gene with asthma and bronchial hyperresponsiveness. AB - Asthma is a common respiratory disorder characterized by recurrent episodes of coughing, wheezing and breathlessness. Although environmental factors such as allergen exposure are risk factors in the development of asthma, both twin and family studies point to a strong genetic component. To date, linkage studies have identified more than a dozen genomic regions linked to asthma. In this study, we performed a genome-wide scan on 460 Caucasian families and identified a locus on chromosome 20p13 that was linked to asthma (log(10) of the likelihood ratio (LOD), 2.94) and bronchial hyperresponsiveness (LOD, 3.93). A survey of 135 polymorphisms in 23 genes identified the ADAM33 gene as being significantly associated with asthma using case-control, transmission disequilibrium and haplotype analyses (P = 0.04 0.000003). ADAM proteins are membrane-anchored metalloproteases with diverse functions, which include the shedding of cell surface proteins such as cytokines and cytokine receptors. The identification and characterization of ADAM33, a putative asthma susceptibility gene identified by positional cloning in an outbred population, should provide insights into the pathogenesis and natural history of this common disease. PMID- 12110847 TI - Gaining a lot from translation. PMID- 12110848 TI - After the gold-rush. PMID- 12110849 TI - Dubious data remain in print two years after misconduct inquiry. PMID- 12110850 TI - Botswana's AIDS laboratory squares up to HIV pandemic. PMID- 12110851 TI - Europe gets tough on labelling genetically modified foodstuffs. PMID- 12110853 TI - AIDS meeting demands more than lip service. PMID- 12110854 TI - Race is on to find alternative to animal tests. PMID- 12110856 TI - Physicists plot mass production of neutrinos. PMID- 12110857 TI - Public body appointed to clean up UK's nuclear legacy. PMID- 12110855 TI - Japan lays out 55-point patent plan. PMID- 12110858 TI - Supercomputer to aid African HIV research. PMID- 12110859 TI - Misconduct in physics: time to wise up? PMID- 12110860 TI - The brave new world of RNA. PMID- 12110861 TI - Jury out on Jefferson's alleged descendants. PMID- 12110863 TI - Rebel army need not be a barrier to conservation. PMID- 12110864 TI - Interoperability calls for an unusual mix of skills. PMID- 12110869 TI - The bigger picture. PMID- 12110871 TI - Out on the edge. PMID- 12110870 TI - Hominid revelations from Chad. PMID- 12110872 TI - Do the locomotion. PMID- 12110873 TI - Bacterial game dynamics. PMID- 12110874 TI - Cleaning up catalysts. PMID- 12110875 TI - Apoptosis: repulsive encounters. PMID- 12110876 TI - Theoretical biology: ants on a Turing trail. PMID- 12110878 TI - Snake circumvents constraints on prey size. AB - For animals who are unable to take bites out of their food, the size of the food item that can be consumed is constrained by the maximal size of the mouth opening (gape)--snakes are an example of gape-limited predators and they usually swallow their prey whole. Here we describe unique feeding behaviours in two closely related species of snake, which circumvent their gape limitation by removing and consuming pieces from newly moulted crabs that are too large to be swallowed intact. This evolutionary innovation is surprising, as the needle-like teeth and highly mobile bones that facilitate the capture and engulfment of large, whole prey by snakes are ill-suited both to cutting and to generating large bite forces. PMID- 12110879 TI - Recombination: Multiply infected spleen cells in HIV patients. AB - The genome of the human immunodeficiency virus is highly prone to recombination, although it is not obvious whether recombinants arise infrequently or whether they are constantly being spawned but escape identification because of the massive and rapid turnover of virus particles. Here we use fluorescence in situ hybridization to estimate the number of proviruses harboured by individual splenocytes from two HIV patients, and determine the extent of recombination by sequencing amplified DNA from these cells. We find an average of three or four proviruses per cell and evidence for huge numbers of recombinants and extensive genetic variation. Although this creates problems for phylogenetic analyses, which ignore recombination effects, the intracellular variation may help to broaden immune recognition. PMID- 12110880 TI - A new hominid from the Upper Miocene of Chad, Central Africa. AB - The search for the earliest fossil evidence of the human lineage has been concentrated in East Africa. Here we report the discovery of six hominid specimens from Chad, central Africa, 2,500 km from the East African Rift Valley. The fossils include a nearly complete cranium and fragmentary lower jaws. The associated fauna suggest the fossils are between 6 and 7 million years old. The fossils display a unique mosaic of primitive and derived characters, and constitute a new genus and species of hominid. The distance from the Rift Valley, and the great antiquity of the fossils, suggest that the earliest members of the hominid clade were more widely distributed than has been thought, and that the divergence between the human and chimpanzee lineages was earlier than indicated by most molecular studies. PMID- 12110881 TI - Geology and palaeontology of the Upper Miocene Toros-Menalla hominid locality, Chad. AB - All six known specimens of the early hominid Sahelanthropus tchadensis come from Toros-Menalla site 266 (TM 266), a single locality in the Djurab Desert, northern Chad, central Africa. Here we present a preliminary analysis of the palaeontological and palaeoecological context of these finds. The rich fauna from TM 266 includes a significant aquatic component such as fish, crocodiles and amphibious mammals, alongside animals associated with gallery forest and savannah, such as primates, rodents, elephants, equids and bovids. The fauna suggests a biochronological age between 6 and 7 million years. Taken together with the sedimentological evidence, the fauna suggests that S. tchadensis lived close to a lake, but not far from a sandy desert, perhaps the oldest record of desert conditions in the Neogene of northern central Africa. PMID- 12110882 TI - Possible in situ formation of meteoritic nanodiamonds in the early Solar System. AB - Grains of dust that pre-date the Sun provide insights into their formation around other stars and into the early evolution of the Solar System. Nanodiamonds recovered from meteorites, which originate in asteroids, have been thought to be the most abundant type of presolar grain. If that is true, then nanodiamonds should be at least as abundant in comets, because they are thought to have formed further out in the early Solar System than the asteroid parent bodies, and because they should be more pristine. Here we report that nanodiamonds are absent or very depleted in fragile, carbon-rich interplanetary dust particles, some of which enter the atmosphere at speeds within the range of cometary meteors. One interpretation of the results is that some (perhaps most) nanodiamonds formed within the inner Solar System and are not presolar at all, consistent with the recent detection of nanodiamonds within the accretion discs of other young stars. An alternative explanation is that all meteoritic nanodiamonds are indeed presolar, but that their abundance decreases with heliocentric distance, in which case our understanding of large-scale transport and circulation within the early Solar System is incomplete. PMID- 12110883 TI - Phonon-enhanced light matter interaction at the nanometre scale. AB - Optical near fields exist close to any illuminated object. They account for interesting effects such as enhanced pinhole transmission or enhanced Raman scattering enabling single-molecule spectroscopy. Also, they enable high resolution (below 10 nm) optical microscopy. The plasmon-enhanced near-field coupling between metallic nanostructures opens new ways of designing optical properties and of controlling light on the nanometre scale. Here we study the strong enhancement of optical near-field coupling in the infrared by lattice vibrations (phonons) of polar dielectrics. We combine infrared spectroscopy with a near-field microscope that provides a confined field to probe the local interaction with a SiC sample. The phonon resonance occurs at 920 cm(-1). Within 20 cm(-1) of the resonance, the near-field signal increases 200-fold; on resonance, the signal exceeds by 20 times the value obtained with a gold sample. We find that phonon-enhanced near-field coupling is extremely sensitive to chemical and structural composition of polar samples, permitting nanometre-scale analysis of semiconductors and minerals. The excellent physical and chemical stability of SiC in particular may allow the design of nanometre-scale optical circuits for high-temperature and high-power operation. PMID- 12110884 TI - Pressure-induced insulator conductor transition in a photoconducting organic liquid-crystal film. AB - Intermolecular separation determines the extent of orbital overlap and thus the rate of electron transfer between neighbouring molecules in an organic crystal. If such a crystal is compressed, the resistivity decreases owing to a diminishing intermolecular distance. Metal insulator transitions have been observed by applying hydrostatic pressure to, for example, Langmuir films of metal nanoparticles. But previous attempts to observe a clear transition point in organic crystals, such as anthracene and tetracene, were not successful owing to difficulties with electrically insulating the high-pressure cell. Here we report a different approach by using a sample that is photoconductive and forms an organized film. A cylindrical tip (approximately 100 microm in diameter) was used to compress the sample instead of a piston/cylinder structure, entirely eliminating the problem of electrical insulation. Furthermore, by illuminating the sample with a laser, the conductivity of the sample is increased by several orders of magnitude. By monitoring the photocurrent with sensitivity at the 10( 13) A level, changes in resistivity at very low pressure could be monitored. We observe a sharp increase in current that could indicate a transition from hopping to delocalized conduction. PMID- 12110885 TI - Self-regeneration of a Pd-perovskite catalyst for automotive emissions control. AB - Catalytic converters are widely used to reduce the amounts of nitrogen oxides, carbon monoxide and unburned hydrocarbons in automotive emissions. The catalysts are finely divided precious-metal particles dispersed on a solid support. During vehicle use, the converter is exposed to heat, which causes the metal particles to agglomerate and grow, and their overall surface area to decrease. As a result, catalyst activity deteriorates. The problem has been exacerbated in recent years by the trend to install catalytic converters closer to the engine, which ensures immediate activation of the catalyst on engine start-up, but also places demanding requirements on the catalyst's heat resistance. Conventional catalyst systems thus incorporate a sufficient excess of precious metal to guarantee continuous catalytic activity for vehicle use over 50,000 miles (80,000 km). Here we use X-ray diffraction and absorption to show that LaFe(0.57)Co(0.38)Pd(0.05)O(3), one of the perovskite-based catalysts investigated for catalytic converter applications since the early 1970s, retains its high metal dispersion owing to structural responses to the fluctuations in exhaust-gas composition that occur in state-of-the-art petrol engines. We find that as the catalyst is cycled between oxidative and reductive atmospheres typically encountered in exhaust gas, palladium (Pd) reversibly moves into and out of the perovskite lattice. This movement appears to suppress the growth of metallic Pd particles, and hence explains the retention of high catalyst activity during long-term use and ageing. PMID- 12110886 TI - A satellite geodetic survey of large-scale deformation of volcanic centres in the central Andes. AB - Surface deformation in volcanic areas usually indicates movement of magma or hydrothermal fluids at depth. Stratovolcanoes tend to exhibit a complex relationship between deformation and eruptive behaviour. The characteristically long time spans between such eruptions requires a long time series of observations to determine whether deformation without an eruption is common at a given edifice. Such studies, however, are logistically difficult to carry out in most volcanic arcs, as these tend to be remote regions with large numbers of volcanoes (hundreds to even thousands). Here we present a satellite-based interferometric synthetic aperture radar (InSAR) survey of the remote central Andes volcanic arc, a region formed by subduction of the Nazca oceanic plate beneath continental South America. Spanning the years 1992 to 2000, our survey reveals the background level of activity of about 900 volcanoes, 50 of which have been classified as potentially active. We find four centres of broad (tens of kilometres wide), roughly axisymmetric surface deformation. None of these centres are at volcanoes currently classified as potentially active, although two lie within about 10 km of volcanoes with known activity. Source depths inferred from the patterns of deformation lie between 5 and 17 km. In contrast to the four new sources found, we do not observe any deformation associated with recent eruptions of Lascar, Chile. PMID- 12110887 TI - Local dispersal promotes biodiversity in a real-life game of rock-paper-scissors. AB - One of the central aims of ecology is to identify mechanisms that maintain biodiversity. Numerous theoretical models have shown that competing species can coexist if ecological processes such as dispersal, movement, and interaction occur over small spatial scales. In particular, this may be the case for non transitive communities, that is, those without strict competitive hierarchies. The classic non-transitive system involves a community of three competing species satisfying a relationship similar to the children's game rock-paper-scissors, where rock crushes scissors, scissors cuts paper, and paper covers rock. Such relationships have been demonstrated in several natural systems. Some models predict that local interaction and dispersal are sufficient to ensure coexistence of all three species in such a community, whereas diversity is lost when ecological processes occur over larger scales. Here, we test these predictions empirically using a non-transitive model community containing three populations of Escherichia coli. We find that diversity is rapidly lost in our experimental community when dispersal and interaction occur over relatively large spatial scales, whereas all populations coexist when ecological processes are localized. PMID- 12110888 TI - Exceptional sperm cooperation in the wood mouse. AB - Spermatozoa from a single male will compete for fertilization of ova with spermatozoa from another male when present in the female reproductive tract at the same time. Close genetic relatedness predisposes individuals towards altruism, and as haploid germ cells of an ejaculate will have genotypic similarity of 50%, it is predicted that spermatozoa may display cooperation and altruism to gain an advantage when inter-male sperm competition is intense. We report here the probable altruistic behaviour of spermatozoa in an eutherian mammal. Spermatozoa of the common wood mouse, Apodemus sylvaticus, displayed a unique morphological transformation resulting in cooperation in distinctive aggregations or 'trains' of hundreds or thousands of cells, which significantly increased sperm progressive motility. Eventual dispersal of sperm trains was associated with most of the spermatozoa undergoing a premature acrosome reaction. Cells undergoing an acrosome reaction in aggregations remote from the egg are altruistic in that they help sperm transport to the egg but compromise their own fertilizing ability. PMID- 12110889 TI - Transmitter-evoked local calcium release stabilizes developing dendrites. AB - In the central nervous system, dendritic arborizations of neurons undergo dynamic structural remodelling during development. Processes are elaborated, maintained or eliminated to attain the adult pattern of synaptic connections. Although neuronal activity influences this remodelling, it is not known how activity exerts its effects. Here we show that neurotransmission-evoked calcium (Ca(2+)) release from intracellular stores stabilizes dendrites during the period of synapse formation. Using a ballistic labelling method to load cells with Ca(2+) indicator dyes, we simultaneously monitored dendritic activity and structure in the intact retina. Two distinct patterns of spontaneous Ca(2+) increases occurred in developing retinal ganglion cells--global increases throughout the arborization, and local 'flashes' of activity restricted to small dendritic segments. Blockade of local, but not global, activity caused rapid retraction of dendrites. This retraction was prevented locally by focal uncaging of caged Ca(2+) that triggered Ca(2+) release from internal stores. Thus, local Ca(2+) release is a mechanism by which afferent activity can selectively and differentially regulate dendritic structure across the developing arborization. PMID- 12110890 TI - Release of chromatin protein HMGB1 by necrotic cells triggers inflammation. AB - High mobility group 1 (HMGB1) protein is both a nuclear factor and a secreted protein. In the cell nucleus it acts as an architectural chromatin-binding factor that bends DNA and promotes protein assembly on specific DNA targets. Outside the cell, it binds with high affinity to RAGE (the receptor for advanced glycation end products) and is a potent mediator of inflammation. HMGB1 is secreted by activated monocytes and macrophages, and is passively released by necrotic or damaged cells. Here we report that Hmgb1(-/-) necrotic cells have a greatly reduced ability to promote inflammation, which proves that the release of HMGB1 can signal the demise of a cell to its neighbours. Apoptotic cells do not release HMGB1 even after undergoing secondary necrosis and partial autolysis, and thus fail to promote inflammation even if not cleared promptly by phagocytic cells. In apoptotic cells, HMGB1 is bound firmly to chromatin because of generalized underacetylation of histone and is released in the extracellular medium (promoting inflammation) if chromatin deacetylation is prevented. Thus, cells undergoing apoptosis are programmed to withhold the signal that is broadcast by cells that have been damaged or killed by trauma. PMID- 12110891 TI - Reciprocal regulation of CD4/CD8 expression by SWI/SNF-like BAF complexes. AB - Thymic development produces two sub-lineages of T cells expressing either CD4 or CD8 co-receptors that assist antibody production and mediate cell killing, respectively. The mechanisms for mutually exclusive co-receptor expression remain poorly defined. We find that mutations in the high mobility group (HMG) domain of BAF57--a DNA-binding subunit of the mammalian SWI/SNF-like chromatin-remodelling BAF complexes--or in the BAF complex ATPase subunit Brg, impair both CD4 silencing and CD8 activation. Brg is haploinsufficient for CD8 activation, but not for CD4 silencing, whereas BAF57 mutations preferentially impair CD4 silencing, pointing to target- and subunit-specific mechanisms of chromatin remodelling. BAF complexes directly bind the CD4 silencer, but the BAF57 HMG domain is dispensable for tethering BAF complexes to the CD4 silencer or other chromatin loci in vivo, or for remodelling reconstituted templates in vitro, suggesting that chromatin remodelling in vivo requires HMG-dependent DNA bending. These results indicate that BAF complexes contribute to lineage bifurcation by reciprocally regulating lineage-specific genes, reminiscent of the role of the yeast SWI/SNF complex in mediating mating-type switching. PMID- 12110892 TI - Apoptosis disables CD31-mediated cell detachment from phagocytes promoting binding and engulfment. AB - Macrophage recognition and ingestion of 'self' cells undergoing apoptosis in vivo protects tissues from the toxic contents of dying cells and modulates macrophage regulation of inflammatory and immune responses. However, the complex molecular mechanisms mediating macrophage discrimination between viable and apoptotic cells are poorly understood. In particular, little is known of why viable nucleated cells are not engulfed by macrophages. To reveal active repulsion of viable cells and to seek specific capture or 'tethering' of apoptotic cells, we studied macrophage binding of viable and apoptotic leukocytes under conditions of flow. We found that homophilic ligation of CD31 (ref. 4) on viable leukocytes promoted their active, temperature-dependent detachment under low shear, whereas such CD31 mediated detachment was disabled in apoptotic leukocytes, promoting tight binding and macrophage ingestion of dying cells. Here we propose that CD31 (also known as platelet-endothelial cell adhesion molecule-1, PECAM-1) is an example of a cell surface molecule that prevents phagocyte ingestion of closely apposed viable cells by transmitting 'detachment' signals, and which changes function on apoptosis, promoting tethering of dying cells to phagocytes. PMID- 12110893 TI - Overexpression of beta-carotene hydroxylase enhances stress tolerance in Arabidopsis. AB - Plant stress caused by extreme environmental conditions is already a principal reason for yield reduction in crops. The threat of global environment change makes it increasingly important to generate crop plants that will withstand such conditions. Stress, particularly stress caused by increased sunlight, leads to the production of reactive oxygen species that cause photo-oxidative cell damage. Carotenoids, which are present in the membranes of all photosynthetic organisms, help protect against such light-dependent oxidative damage. In plants, the xanthophyll cycle (the reversible interconversion of two carotenoids, violaxanthin and zeaxanthin) has a key photoprotective role and is therefore a promising target for genetic engineering to enhance stress tolerance. Here we show that in Arabidopsis thaliana overexpression of the chyB gene that encodes beta-carotene hydroxylase--an enzyme in the zeaxanthin biosynthetic pathway- causes a specific twofold increase in the size of the xanthophyll cycle pool. The plants are more tolerant to conditions of high light and high temperature, as shown by reduced leaf necrosis, reduced production of the stress indicator anthocyanin and reduced lipid peroxidation. Stress protection is probably due to the function of zeaxanthin in preventing oxidative damage of membranes. PMID- 12110894 TI - NMR analysis of a 900K GroEL GroES complex. AB - Biomacromolecular structures with a relative molecular mass (M(r)) of 50,000 to 100,000 (50K 100K) have been generally considered to be inaccessible to analysis by solution NMR spectroscopy. Here we report spectra recorded from bacterial chaperonin complexes ten times this size limit (up to M(r) 900K) using the techniques of transverse relaxation-optimized spectroscopy and cross-correlated relaxation-enhanced polarization transfer. These techniques prevent deterioration of the NMR spectra by the rapid transverse relaxation of the magnetization to which large, slowly tumbling molecules are otherwise subject. We tested the resolving power of these techniques by examining the isotope-labelled homoheptameric co-chaperonin GroES (M(r) 72K), either free in solution or in complex with the homotetradecameric chaperonin GroEL (M(r) 800K) or with the single-ring GroEL variant SR1 (M(r) 400K). Most amino acids of GroES show the same resonances whether free in solution or in complex with chaperonin; however, residues 17 32 show large chemical shift changes on binding. These amino acids belong to a mobile loop region of GroES that forms contacts with GroEL. This establishes the utility of these techniques for solution NMR studies that should permit the exploration of structure, dynamics and interactions in large macromolecular complexes. PMID- 12110897 TI - The antiquity of RNA-based evolution. AB - All life that is known to exist on Earth today and all life for which there is evidence in the geological record seems to be of the same form--one based on DNA genomes and protein enzymes. Yet there are strong reasons to conclude that DNA- and protein-based life was preceded by a simpler life form based primarily on RNA. This earlier era is referred to as the 'RNA world', during which the genetic information resided in the sequence of RNA molecules and the phenotype derived from the catalytic properties of RNA. PMID- 12110898 TI - The chemical repertoire of natural ribozymes. AB - Although RNA is generally thought to be a passive genetic blueprint, some RNA molecules, called ribozymes, have intrinsic enzyme-like activity--they can catalyse chemical reactions in the complete absence of protein cofactors. In addition to the well-known small ribozymes that cleave phosphodiester bonds, we now know that RNA catalysts probably effect a number of key cellular reactions. This versatility has lent credence to the idea that RNA molecules may have been central to the early stages of life on Earth. PMID- 12110899 TI - The involvement of RNA in ribosome function. AB - The ribosome is a particle made of RNA and protein that is found in abundance in all cells that are actively making protein. It catalyses the messenger RNA directed synthesis of proteins. Recent structural work has demonstrated a profound involvement of the ribosome's RNA component in all aspects of its function, supporting the hypothesis that proteins were added to the ribosome late in its evolution. PMID- 12110900 TI - Alternative pre-mRNA splicing and proteome expansion in metazoans. AB - The protein coding sequences of most eukaryotic messenger RNA precursors (pre mRNAs) are interrupted by non-coding sequences called introns. Pre-mRNA splicing is the process by which introns are removed and the protein coding elements assembled into mature mRNAs. Alternative pre-mRNA splicing selectively joins different protein coding elements to form mRNAs that encode proteins with distinct functions, and is therefore an important source of protein diversity. The elaboration of this mechanism may have had a significant role in the expansion of metazoan proteomes during evolution. PMID- 12110901 TI - RNA interference. AB - A conserved biological response to double-stranded RNA, known variously as RNA interference (RNAi) or post-transcriptional gene silencing, mediates resistance to both endogenous parasitic and exogenous pathogenic nucleic acids, and regulates the expression of protein-coding genes. RNAi has been cultivated as a means to manipulate gene expression experimentally and to probe gene function on a whole-genome scale. PMID- 12110903 TI - Molecular Recognition through Isosteric Interaction--Implication from Studies on Insulin. PMID- 12110902 TI - Emerging clinical applications of RNA. AB - RNA is a versatile biological macromolecule that is crucial in mobilizing and interpreting our genetic information. It is not surprising then that researchers have sought to exploit the inherent properties of RNAs so as to interfere with or repair dysfunctional nucleic acids or proteins and to stimulate the production of therapeutic gene products in a variety of pathological situations. The first generation of the resulting RNA therapeutics are now being evaluated in clinical trials, raising significant interest in this emerging area of medical research. PMID- 12110904 TI - Interaction between Jak3 and Nucleosome Assembly Protein 1. AB - Tyrosine kinase Jak3 plays a critical role in the interleukin 2 IL-2 signaling because it not only participates the Jak-Stat pathway, but also interacts with unidentified signal transducers and regulates expression of some oncogenes such as c-fos and c-myc. Abundant evidence demonstrated that phosphorylated tyrosine was necessary for the interaction between two proteins. Therefore, in order to clarify the role of Jak3 in IL-2 signal transduction, the tyrosine phosphorylation-involved yeast two-hybrid system was constructed and the N terminal region JH3-JH7 of Jak3 was used as a bait to screen a peripheral blood cDNA library. About 50 double-positive colonies were obtained. Sequence analysis indicated that one of them was from nucleosome assembly protein 1 gene (Nap1), and encoded a protein of 392 amino acid residues. Two-hybrid system results demonstrated that interaction between Jak3 and Nap1 depended on the level of tyrosine phosphorylation. Furthermore, immunoprecipitation and Western blot experiments confirmed that Jak3 really interacted with Nap1 in murine pro-B lymphocyte BAF/BO3beta cells. PMID- 12110905 TI - Cloning and Overexpression of the Mitomycin C Resistance Gene (mcr) from Streptoverticillium caespitosum. AB - Streptoverticillium caespitosum ATCC27422 is a major producer of an anti-cancer drug, mitomycin C. A 6.6 kb DNA fragment containing the mitomycin C resistance gene (mcr) was isolated from ATCC27422 by shotgun method in order to learn the molecular mechanism of mitomycin C resistance. By constructing a series of subclones from this 6.6 kb DNA fragment, the mitomycin C resistance gene was localized on a 3.1 kb DNA fragment. Sequence analysis revealed that the open reading frame of mcr gene was 1 347 bp in size, encoding 448 amino acids with ATG as initiation codon and TGA as termination codon. The mcr gene was specifically expressed under the control of T7 promoter in E.coli, and the resistance to mitomycin C in the transformant was over 100-fold higher than that in wild-type strain. The overexpression of mcr gene in E.coli is very helpful for the further research about the molecular mechanism of drug resistance. PMID- 12110906 TI - Fusion Expression and Antibody Preparation of the Human Transcription Factor hB1F. AB - Human B1F was cloned from a human liver cDNA library by yeast one-hybrid screening and characterized. It is a transcription factor which belongs to nuclear receptor superfamily. The cDNA segment 450-930 of human liver transcription factor hB1F was cloned into the pGEX-3X expression vector and was expressed in E.coli. The purified GST-fused hB1F, named GST-CS, were used to immunize mice to get polyclonal antibodies to hB1F. The antiserum showed specificity to hB1F after the purification by GST-Sepharose 4B. It can be used in further investigations of the structure and function of hB1F. PMID- 12110907 TI - Cloning, High Level Expression and Purification of Rat betaB2-crystallin. AB - beta-crystallins are the largest group of lens structural proteins, which are necessary for both the high refractive index and the transparency of the eye lens, and have been implicated in various kinds of cataracts. To obtain abundant betaB2-crystallin for the study of the mechanism of their oligomerization, a bacterial expression system for betaB2-crystallin and a rapid method for its purification were developed. cDNA encoding rat betaB2-crystallin was cloned using RT-PCR. After the induction of recA promoter with nalidixic acid, abundant protein was produced in E. coli. betaB2-crystallin comprised about 30% of the total bacterial proteins and it is water-soluble. After anion-exchange chromatography on DEAE cellulose and gel filtration on Sephadex G-100, the protein was obtained pure as judged by SDS-PAGE. The high level expression and rapid purification of recombinant betaB2-crystallin will facilitate the further study of structure-function relationship of betaB2-crystallin. PMID- 12110908 TI - Construction and Characterization of a Mutant of Single-chain Urokinase-type Plasminogen Activator Ser(175)-His(187)-mscu-PA AB - Single-chain urokinase-type plasminogen activator scu-PA is the precursor of double-chain urokinase tcu-PA , which has a much higher intrinsic catalytic activity than other zymogens of the serine protease family. To restore the "zymogen triad" of Asp-His-Ser in the serine protease family, the mutant gene of scu-PA mscu-PA, Ala(175)right curved arrow Ser(175), Tyr(187)right curved arrow His(187) was constructed by the method of oligonucleotide-directed, site-specific mutagenesis in order to reduce its intrinsic catalytic activity. mscu-PA was expressed in E. coli BL21. After denaturation and renaturation in vitro, the mscu PA was purified to homogeneity by SP-Sepharose ion-exchange chromatography, Sephacryl S-200 chromatography and Benzamidine-Sepharose affinity adsorption. mscu-PA had the same activity to plasmin as scu-PA. The catalytic efficiency measured by k(cat)/K(m) to synthetic substrate S(2444) was 2.5-fold lower than that of scu-PA, and the activity against Glu-plasminogen was also reduced. After activation by plasmin, mtcu-PA and tcu-PA had similar catalytic efficiency against S(2444) and Glu-plasminogen. The results suggest that the intrinsic catalytic activity of mscu-PA be really reduced after restoring the "zymogen triad". PMID- 12110909 TI - Preparation of Inositol Tetrakisphosphate and Its Application in Modification of Porcine Hemoglobin. AB - Modification of hemoglobin using inositol tetrakisphosphate (IP(4)) can improve the oxygen affinity of hemoglubin. Phytase was extracted from wheat bran and purified by ammonium sulphate fractionation, followed by Sephadex G-50 gel filtration and Mono Q chromatography. The purified phytase was used for hydrolysis of phytic acid under controled conditions. IP(4), as a major composition of the hydrolysate, was further purified on resin 714 column. The purified IP(4) was oxidized by periodate to obtain dialdehyde-IP(4). By the reaction between the aldehyde group of IP(4) derivative and the amino group of porcine hemoglobin (pHb), pHb-IP(4) conjugate was formed and was found to have better ability of O(2) binding and releasing than the native hemoglobin. PMID- 12110910 TI - Exogenous p21(WAF1) Transfection Affects Apoptosis in Human Fibroblasts. AB - pDOR-p21 sense and pDOR-p21 antisense retroviral expression vectors were constructed and successfully transfected by Lipofectin into normal human diploid fibroblasts(2BS line), resulting in 2BS-p21s and 2BS-p21a cell lines, respectively. Southern blot analysis verified that the exogenous p21(WAF1) cDNA were integrated into genomic DNA. Compared with the cells transfected with pDOR neo empty vector, p21(WAF1) mRNA expression increased in 2BS-p21s cells, which were less sensitive to apoptosis induced by NaBu, and showed higher cell viability, delayed appearance of DNA ladder, and less area of apoptosis peak. On the other hand, p21(WAF1) mRNA expression decreased in 2BS-p21a cells, which were more sensitive to apoptosis induced by NaBu. These results indicated that the expression amount of p21(WAF1) in 2BS cells was negatively related with its susceptibility to apoptosis induced by NaBu. PMID- 12110911 TI - An ESR Study on the Effect of Hydration on the Dynamic Property of RNase A. AB - A method is described for the measurement of dynamic property of RNase A by ESR under xeric conditions. The relationship between relative humidity and hydration degree of RNase A was determined by hydration isotherm. A solution of RNase A was allowed to react with a solution containing maleimide nitroxide label at 25 degrees, then was dialysed and lyophilized. The stable powder of RNase A - maleimide nitroxide label compound was put into the tubules, then was hydrated under different relative humidity for 11 days. After hydration, the tubules were closed and measured by ESR. The relationship between hydration value and A(max) was detected. The results showed that the lowest water content that could induce motion of RNase A by water is about 0.20 g of water per g of RNase A. That means the motion of RNase A molecule becomes detectable when there are 152 water molecules around one RNase A molecule. PMID- 12110912 TI - Purification and Functional Characterization of a Shark Cartilage Factor Inhibitory to Angiogenesis. AB - SCAIF-I, an inhibitor of angiogenesis from shark cartilage was purified to homogeneity. The 4 mol/L guanidinium chloride extract of shark cartilage was fractionally precipitated with 35%-65% acetone, then purified by Resource Q ion exchange chromatography, Sephacryl S-300 gel filtration, and reverse-phase high performance liquid chromatography. The pure inhibitor was homogeneous as a single band on a silver-stained 15% sodium dodecyl sulfate-polyacrylamide gel. SCAIF-I had an molecular weight of 18 kD. It specifically inhibited proliferation of endothelial cells, and strongly blocked endothelial cell movement and angiogenesis in the chorioallantoic membrane of chick embryos. Systemic administration of SCAIF-I at the dose of 5 mg/kg.d suppressed 87.93% of the growth of Lewis lung carcinoma implanted in C57BL/6 mice. PMID- 12110913 TI - An NF-IL6 3'UTR RNA-specific Binding Protein in E. coli Purification and Partial Protein Sequencing. AB - protein was found in E.coli which can specifically bind to NF-IL6 mRNA 3'UTR. After a series of purification steps, the RNA-specific binding protein was directly sequenced on the Porton LF3200 Protein Sequencer. A sequence of 10 amino acids of the purified NF-IL6 3'UTR binding protein was obtained, namely, Ala-Thr Arg-Ile-Glu-Phe-His-Gly-Cyss(?)-Gly. A BLAST search with the 10 amino acids against NCBI database failed to identify any protein with identical sequence. The significance of identifying an eukaryotic mRNA binding protein in E.coli is discussed. PMID- 12110914 TI - Fusion and Expression of the Genes Encoding Murine-Human Chimeric Heavy Chain to Carcinoembryonic Antigen with Core-streptavidin Gene. AB - In order to reduce the human anti-murine antibody (HAMA) in radioimmunotherapy, the gene encoding the heavy chain variable region (V(H)) of the murine monoclonal antibody with high specificity and affinity to carcinoembryonic antigen was fused to the human C(gamma3) gene to construct the murine-human chimeric heavy chain antibody gene, then was linked to core-streptavidin which can specifically bind to biotin, facilitating its purification and radioisotope labeling. The fusion gene was expressed in E.coli at high level, accounting for 24% of the total bacteria protein, and was characterized by SDS-PAGE and Western-blots. When using RIA the content of inclusion bodies was denatured and followed by renaturation, it was shown by using RIA to possess ability to bind to its specific antigen of CEA. Using horseradish peroxidase (HRP) labeled biotin as antibody in Western blots, one band of 70 kD only was detected, demonstrating the fusion protein not only had the ability to bind CEA, but also could bind biotin specifically. PMID- 12110915 TI - High Expression and Purification of Recombinant Human Serum Albumin from Pichia pastoris. AB - Recombinant human serum albumin (rHSA) was produced in methylotrophic yeast Pichia pastoris. By optimization of expression, about 150 mg/L of rHSA was obtained from broth of Pichia pastoris GS115/HAS (his+Mut(S)) supernatant. The rHSA was isolated and purified by hollow-fiber ultrafiltration, Phenyl-Sepharose hydrophobic chromatography and antibody-immunoadsorbent chromatography. Finally, rHSA was purified to electrophoretic purity. PMID- 12110916 TI - Ubiquitin Conjugating Enzyme Ubc9 is Involved in Protein Degradation of Redox Factor-1 (Ref-1). AB - Redox factor-1 (Ref-1) is a bifunctional protein playing an important role in both cellular redox regulation and DNA apurinic/apyrimidinic sites' repair. To find Ref-1interacting proteins (Rips), a yeast two-hybrid screening was performed by using Ref-1 redox domain as the 'bait', and five positive clones were obtained. One of them (Rip3) was identified to be the ubiquitin-conjugating enzyme Ubc9. Simultaneous overexpression of Ubc9 in Hela cells dramatically inhibited the enhancement of AP-1 reporter gene by Ref-1. Western blot indicated that the protein level of Ref-1 dropped down as the result of simultaneous overexpression of Ubc9. These results suggest that Ubc9 is involved in the protein degradation of Ref-1, resulting in the downregulation of Ref-1 physiological function. PMID- 12110917 TI - Thiophosphorylation and Fluorescent Labeling of Substrate Peptide of Protein Kinase. AB - Study on the conditions of thiophosphorylation reaction and fluorescent labeling reaction of the substrate of protein kinase A was carried out by using Kemptide LRRASLG as a model. The suitable concentration of the fluorescent regent 5-{ ((2 iodoacetyl)amino)ethyl amino} naphthelene-1-sulfonic acid (1,5-IAEDANS) and the suitable pH of labeling reaction buffer were 1.6 mmol/L and 8, respectively. Stability of the labeled peptide under the conditions of automatic N-terminal protein sequencing, electrospray mass spectrometry and in the presence of 0.1% trifluoroacetic acid was investigated, respectively. The specific differences in UV spectra between the labeled and unlabeled peptides were observed. Therefore, the possibility to detect the thiophosphorylated and fluorophore labeled peptide during high performance liquid chromatography peptide mapping was primarily shown. PMID- 12110918 TI - Immobilized Tryptophanyl-tRNA Synthetase from Bacillus subtilis. AB - In order to investigate the recognition mechanism and the relationship between structure and function of tRNA(Trp) with tryptophanyl-tRNA synthetase (TrpRS), TrpRS from Bacillus subtilis was purified and immobilized on CNBr-activated Sepharose 4B. Protein recovery and activity recovery of the immobilization were 95.5% and 31.3%, respectively. Properties of immobilized TrpRS were studied in detail. The thermal stability and the shelf stability of immobilized TrpRS were much higher than those of the native TrpRS. Besides these, the immobilized TrpRS, with good operation stability, had increased optimum temperature and optimum pH. A 56-base single-stranded RNA library containing 20 consecutive completely randomized bases was subjected to 3 successive rounds of immobilized TrpRS column selection with SELEX method, resulting in a sharp increase of the percentage of the RNA pool that could bind immobilized TrpRS from 4.3% for the first round pool to 14.7% for the third-round pool. After sequencing the third-round RNA pool, a RNA secondary structure resembling the structure of the acceptor stem in tRNA(Trp) was obtained though the selection. All the results indicated that immobilized TrpRS could be used as an affinity chromatography matrix and was qualified for the SELEX of a RNA pool simulating tRNA(Trp) molecule. PMID- 12110919 TI - A Novel Type of Apoptosis Induced by Cadmium in BA/F3beta Cells. AB - Apoptosis is usually accompanied by DNA fragmentation and up-regulation of reactive oxygen species, and it can be inhibited by overexpression of Bcl-2. Here, cadmium was found to induce apoptosis in BA/F3beta cells. MTT assay, Hochest 33258 staining, and transmission electron microscopy analysis were used to detect the apoptosis, however, neither DNA fragmentation nor up-regulation of reactive oxygen species were observed in this type of apoptosis. Furthermore, Bcl 2 overexpression had no effect on this type of apoptosis. In conclusion, these data suggested that cadmium induced a novel type of apoptosis in BA/F3beta cells. PMID- 12110920 TI - cDNA Cloning of the Guinea Pig Growth Hormone Receptor. AB - cDNA cloning and 1 899 bp sequence of growth hormone receptor (GHR) from guinea pig liver are described. The guinea pig GHR consists of 610 amino acids. The structural feature and homology comparison of guinea pig GHR are also reported. PMID- 12110921 TI - Purification and Characterization of a Novel Anticoagulation Factor (ACF II) from the Venom of Agkistrodon acutus. AB - A novel anticoagulation factor (ACF II) was first isolated from the venom of Agkistrodon acutus by DEAE-Sephadex A-50, Sephadex G-75 and CM-Sephadex C-50 column chromatography. The purified ACF II showed a single band in PAGE, S DS PAGE and IEF-PAGE, respectively. ACF II consisted of two peptide chains lin ke d together by disulfide bond(s) and the molecular weights of the two peptide cha ins were about 14.6 kD as determined by SDS-PAGE. The isoelectric point of ACF II was 7.0. ACF II possessed distinct anticoagulant activity and its minimum conc entration to prolong plasma prothrombin time in vitro was 0.4 mg/L. No throm bin- like activity, fibrinolytic activity, phospholipase A(2) activity, haemorrhagic activity, o r lethal activity was detected in ACF II, which may be useful as an effective ve nom-based anticoagulant. PMID- 12110922 TI - Establishment of Cell Line with the FEN-1 Gene Blocked by Its Antisense. AB - FEN-1 is a structure-specific endo/exonuclease, which is involved in the process of both DNA duplication and DNA repair. In this work a mammalian expression vector expressing antisense FEN-1 gene fragment pMAMneoAmp(-)FNB(-) was constructed, after cloning the NcoI-BamHI fragment of FEN-1 gene into the mammalian expression vector pMAMneoAmp(-) in antisense orientation. After FL cell was transfected with pMAMneoAmp(-)FNB(-) and selected by G418, the FL-FEN-1(-) cell line, in which the FEN-1 gene expression was blocked, was established. It was found that the growth of FL-FEN-1(-) was decreased upon the induction with dexamethasone and its T(D) was 3.03 d, while the T(D) of controls FL and FL-M induced with dexamethasone was 2.03 and 2.22 d, respectively, and the T(D) of the FL-FEN-1(-) cell without dexamethasone was 2.38 d. PMID- 12110923 TI - Cloning and Functional Expression of Neurotoxin cDNA from Naja naja atra. AB - Three new cDNAs named NL1, NL2 and NL3 were cloned from the total RNA of Naja naja atra by RT-PCR. The protein sequences encoded by them showed 77%, 72% and 98% structure identity to cobrotoxin which is a postsynaptic neurotoxin from Taiwan cobra (Naja naja atra), respectively. The five conservative residues, Tyr(25), Lys(27), Trp(29), Arg(33) and Lys(47), essential for the function of cobrotoxin were also found in the three cDNAs. The NL3, the most homologous to cobrotoxin, was expressed in E.coli BL21(DE3) by cloning into pET28b+. The expressed product was insoluble inclusion bodies and could be purified up to 90% purity in the range of 6 mg per liter culture cells by a single affinity step. The purified protein was refolded in vitro and the toxicity was assessed to be less than the native cobrotoxin in mice. PMID- 12110924 TI - An Emerging Field in Bioscience High Hydrostatic Pressure Study. AB - The goal of this mini review is to explain how hydrostatic pressure acts on bio macromolecules(mainly on protein), giving some basic knowledge. Moreover, it will be shown that high pressure is a powerful tool both for the study of bio macromolecules such as protein conformation, protein-protein (or protein-nucleic acid, or protein-ligands) interactions and the modulation of enzymatic activity etc. and for study of biotechnological application. PMID- 12110925 TI - Sequence Analysis of 47 Low-abundance ESTs Expressed in Early Human Embryo. AB - The availability of high quality cDNA libraries has proven essential to positional gene cloning efforts differential gene expression studies and EST sequencing/mapping projects.In order to isolate and identify new genes expressed during early human development a 3-week-old human embryo cDNA library was constructed and a pre-screening procedure was used to select cDNAs corresponding to low abundance mRNAs. 6 508 clones were hybridized with a mixture of cDNA probes. Approximately 1 677 clones (26%) did not hybridize with the cDNA probes and represent low abundance mRNAs as well as empty vectors.Partial sequences were generated from one or both end of 47 low abundance cDNA clones and the sequences comparisons with genetic databases revealed that 38.3% of them was annotated human genes 10.6% was highly similar to those from either human or other species 40.4% was partial sequence matched with ESTs that had already been detected and 8.5% of the cDNAs appeared to be unknown in the genetic databases. PMID- 12110926 TI - Purification and Characterization of the Antenna Protein CP29 from Spinach Photosystem II. AB - A Chla/b-binding protein, CP29, was purified from PS II core complex of spinach by DEAE-Toyopearl-650S anion-exchange chromatography, after treatment with the mild nonionic detergent beta-dodecyl maltoside and high concentration of LiClO(4). At room temperature, purified CP29 had a maximum absorption at 677 nm and a fluorescence maximum at 681 nm and doublet CD signals which indicated the presence of excitonic interactions between chlorophylls. The pigment content of the CP29 was 5 D7 Chla molecules and 2 D3 Chlb molecules per CP29 polypeptide, which was determined by spectroscopic method. These results suggested that the purified CP29 was in a native state. The conformational contents of CP29 were analyzed with the help of the room temperature CD spectra of CP29. The secondary structure of CP29 was predicted by using Chou-Fasman method. PMID- 12110927 TI - Overexpressed Human TNF Receptor-75 Can Independently Mediate hTNFalpha-induced Cytotoxicity in BHK-21 Cells. AB - A recombinant bicistronic expression vector was constructed containing human TNF receptor-75 gene and encephalomyocarditis virus(EMCV)internal ribosome entry site(IRES), followed by a neomycin phosphotransferase gene as selectable marker. BHK-21 cells were transfected with this vector using electroporation method and positive clones overexpressing the genes of interest were obtained after selection with G-418. The expression of two hTNFRs in those cells at RNA transcriptional and the protein translation levels had been proved by RT-PCR and indirect ELISA. It was found that the hTR75 could mediate cytotoxicity independently in BHK-21 cells after those cells were treated by hTNFalpha. Furthermore, hTR55 did not display the synergistic activity on this function of hTR75. These results put forward a new way for further studies on structure and function relationships of hTR75 and the interactions between two hTNFRs. PMID- 12110929 TI - Effects of Deleting the Amino-terminal Domain of GRK-2 on Its Function. AB - To reveal the possible role of the amino-terminal domain of G protein-coupled receptor kinases(GRKs)in receptor phosphorylation and/or modulation of its kinase activity, a truncated mutant of GRK-2 lacking the amino-terminal domain(deltaN GRK2)was made. deltaN-GRK2 was expressed effectively in E.coli as a GST fusion protein and was purified by affinity chromatography on a GSH-Sepharose column. deltaN-GRK2 was then separated from GST tag by thrombin cleavage and recovered. Although deltaN-GRK2 had nearly identical activity with wild-type GRK-2 in phosphorylation of peptide substrate, it completely lost the ability to phosphorylate the light-activated receptor rhodopsin. Furthermore, deletion of the amino-terminal domain rendered GRK-2 unresponsive to the regulation of kinase activity by a truncated form of rhodopsin, (329)G-Rho(*) and beta gamma subunits of G protein. These results demonstrated that the amino-terminal domain was necessary to GRK2 for both the phosphorylation of receptor and the regulation of its kinase activity by the receptor. It was reasonable to postulate that this domain has little, if any effect on the catalytic domain of natural form of GRK2. PMID- 12110928 TI - Construction of Early Human Embryo cDNA Libraries and Screening Objective Genes. AB - The construction, evaluation, and application of cDNA libraries from 3-, 4-, and 5-week-old human embryos are described. Total RNAs were extracted from whole embryos using a modified single-step method. mRNA purified by two passes through oligo (dT) columns was reverse-transcripted into single-stranded cDNA. Alkaline agarose electrophoresis showed that the double-strand cDNA fragments ranged from 0.4 9.0 kb and most of them were in the range of 1.0 2.0 kb. After separation on SizeSep 400 Spun columns to eliminate excess adaptors and small cDNA fragments(less than 400 bp), the cDNAs were ligated into pSPORT1 plasmid and lambdaZipLox phage. The plasmid libraries have complexities of 2.6x10(5), 1.7x10(5) and 2.1x10(5) clones and the phage cDNA libraries have complexities of 3.4x10(6), 3.7x10(6) and 2.3x10(6) clones, respectively. Three whole length cDNAs encoding human CD59, MCP and DAF were amplified by PCR using 3-week-old phage library as templates, and human tPA gene with whole length cDNA was screened from 4-week-old plasmid library by hybridization. It was shown that these libraries are of high quality and are suitable to screen rarely expressed genes. The libraries are a valuable source for the study of novel gene expression during human development. PMID- 12110930 TI - Expression of E-selectin in Endothelial Cells Effects of Wild Type p53 Gene. AB - To investigate expression of E-selectin in endothelial cells(EC)and effect of wild type p53 gene transduction on its expression, flow cytometry and RT-PCR were employed to measure the E-selectin protein and mRNA, respectively. The effects of TNFalpha stimulation and p53 gene transduction on the expression of E-selectin was studied. Flow cytometric analysis revealed that there was no E-selectin expression on the surface of resting EC, although its mRNA was detectable by RT PCR. TNFalpha (10-1 000 ku/L)induced E-selectin expression on the surface of EC in a concentration-dependent manner. The level of E-selectin mRNA was also increased with the increasing concentrations of TNFalpha. The expression of E selectin protein was notable after 2 h of TNFalpha stimulation and reached a peak at 4-6 h, then rapidly declined to around the basic level by 12 h. Transduction of p53 gene inhibited TNFalpha-induced EC expression of E-selectin. The E selectin positive cell proportion was reduced from(21.31+/-1.06)% to(11.83+/ 0.98)%, (n = 3, P < 0.001). The E-selectin mRNA level was also reduced correspondingly. In resting EC, neither E-selectin nor its mRNA level was affected by the transduction of p53 gene. The results indicated that p53 gene inhibited TNFalpha-induced expression of E-selectin on the surface of EC, at least partially, by reducing its mRNA level. PMID- 12110931 TI - Epstein-Barr Virus Encoded Latent Membrane Protein 1 Increases Expression of Immunoglobulin kappa Light Chain through NFkappaB in a Nasopharyngeal Carcinoma Cell Line. AB - NFkappaB activity analysis was performed with pNFkappaB-luc reporter plasmid. Expression of NFkappaB in nuclear and expression of Igkappa in intact cells were analyzed by Western Blot to identify whether Epstein-Barr virus encoded latent membrane protein 1 (LMP1) increases the expression of immunoglobulin kappa light chain through NFkappaB signal transduction pathway in nasopharyngeal carcinoma. Results showed that LMP1 could increase NFkappaB activity and promote the accumulation of NFkappaB p65 in nucleus, and LMP1 also increased Igkappaexpression. In addition, phosphorothioate oligonucleotides to antisense NFkappaB p65 and p50 inhibited the Igkappaexpression. These imply that LMP1 can regulate Igkappa expression via NFkappaB signal transduction pathway in nasopharyngeal carcinoma. PMID- 12110932 TI - Purification and Characterization of an Alkaline Lipase from Penicillium cyclopium PG37. AB - An extracellular, novel alkaline lipase produced by Penicillium cyclopium PG37 was purified by centrifugation, ammonium sulfate precipitation, and phenyl Sepharose CL-4B, DEAE Sepharose fast flow and Sephadex G-75 column chromatographies. A 16.5-fold purification of the enzyme was achieved which had a specific activity of 5 200 u/mg protein, and the recovery of the activity was 33.2%. The purified enzyme exhibited a single band on SDS-polyacrylamide gel electrophoresis (SDS-PAGE) and polyacrylamide gel electrophoresis (PAGE). The molecular weight of the native lipase was estimated to be about 29 kD by gel filtration using Sephadex G-150, and that of the denatured lipase was determined to be about 27.5 kD by its mobility on SDS-PAGE, indicating that the lipase was a monomer. The N-terminal amino acid sequence was determined with automatic protein sequencer to be ATADAAAFPD, which has no homology with other sequences of known lipases. The optimum temperature of the action of this enzyme was 25 degrees and the lipase was stable below 30 degrees, but only 30% of its activity remained after 20 min incubation at 40 degrees. The enzyme was stable at pH from 6.5 to 10.5, and its optimal pH for activity is 10.0. Low concentration of alkaline proteinase has little effect on the lipase PG37, therefore these two enzymes can be used as ingredients that are added to commercial detergents simultaneously. PMID- 12110933 TI - Analysis of 5' Upstream Regulatory Sequences of Lignin Peroxidase(LIP) Genes of Phanerochaete chrysosporium. AB - Phanerochaete chrysosporium yields exocellular isozymes of lignin peroxidases and Mn peroxidases which are able to degrade lignin. In order to investigate the regulatory mechanism of LIP genes at transcriptional level, 6 DNA fragments subcloned from 5' upstream sequences of LIP genes (GLG3 and GLG6) were obtained, then gel mobility shift assay was carried out to screen DNA fragment(s) with ability to be bound specifically by proteins isolated from P.chrysosporium mycelia. The results showed that a 670 bp DNA fragment localized upstream of a LIP gene GLG6 was capable of being bound specifically by protein component, and analysis of its nucleotide sequence indicated that the DNA fragment contains the putative sequence characteristic of protein binding site. This result suggests that protein(s) in this fungus interact with the putative cis-element(s) in the 5' upstream sequence of LIP genes, regulating the expression of LIP genes. PMID- 12110934 TI - Expression of a Gene for Single-chain Antibody to Carcinoembryonic Antigen in Bombyx mori cells and Its Larvae. AB - Recombinant Bm-BacScFv virus which contains a cDNA encoding anti-CEA single-chain antibody was generated by cotransfection into Bm N cells with the transfer plasmid pBacPAK-(His)(6)-ScFv and the modified Bombyx mori nuclear polyhedrosis virus Bm-BacPAK genomic DNA. Bombyx mori cells and larvae were infected by the recombinant virus, respectively. The anti-CEA single-chain antibody were expressed both in Bombyx mori cells and larvae, the former accounted for 6% of the total cell protein, the later 0.3 mg per larvae. The histidine-tagged ScFvs were respectively purified with nickel-iminodiacetic acid affinity chromatography. The purity of the former reached over 90% and that of the latter was lower. Purified protein products was able to bind to CEA with an affinity constant of 5.4x10(8)/mol.L(-1) and 2.3x10(8)/mol.L(-1), slightly lower than its parental monoclonal antibody E(7)B(10) which had 2.7x10(9)/mol.L(-1). PMID- 12110935 TI - Cloning and Expression of a Novel Mutated Osteoprogerin/Osteoclastogenesis Inhibitory Factor Gene. AB - Total RNA was isolated from normal Chinese human liver cell line L02. A mutated osteoprotegerin/osteoclastogenesis Inhibitory Factor(OPG/OCIF) cDNA was amplified by RT-PCR using the total RNA as template and was inserted into pBS-sk plasmid. Sequence analysis showed that the OPG/OCIF cDNA from L02 cells was a mutated OPG/OCIF gene which had a nonsense mutation(Ochre) at the codon of Gln(394). The OPG/OCIF isoform was 8 amino acid residues less at the C terminal than the OPG/OCIF reported. The 3'fragment of OPG/OCIF gene from genomic DNA of cell line 293(ATCC CRL 1573) was also cloned. Sequence analysis indicated that the sequence of genomic DNA was the same as that of cDNA. The mutated OPG/OCIF gene was inserted into yeast expression plasmid pPIC3.5K, and the recombinant plasmid was used to transform Pichia pastoris GS115. SDS-PAGE analysis revealed that the human OPG/OCIF isoform was highly expressed and accumulated up to over 30% of soluble protein of yeast after the induction by methanol for 3 to 5 days. The longer the transformant was induced, the higher the ratio of glycosylated protein was. PMID- 12110936 TI - Properties of Recombinant Aeromonas punctata Prolyl Endopeptidase. AB - The properties of recombinant Aeromonas punctata prolyl endopeptidase(apPEP) were studied using specific substrate and peptides. Results show that the optimum catalytic temperature and pH was 34 degrees and 8.4, the stability of the apPEP was in the range of 4-32 degrees and pH 6.0-10.0, and its K(m) was 0.03 mmol/L based on the Z-Gly-Pro-betaNA. The apPEP was not sensitive to PMSF, TLCK, TPCK, Trypsion inhibitor, EDTA, tetrathionate and some metal ions, but was sensitive to SDS and Zn(2 ), and was completely inhibited by DFP. Oxytocin and calcitonin could be specifically hydrolyzed by apPEP at the carboxyl site of proline residue, but the hydrolysis efficiency of calcitonin by the enzyme was less than for oxytocin and for Z-Gly-Pro-betaNA. PMID- 12110937 TI - Molecular Cloning and Expression of a New A Chain of beta-Bungarotoxin. AB - The cDNA encoding a new A chain of beta-bungarotoxin was cloned from Bungarus multicinctus. It encoded a 27-amino acid signal peptide and a mature protein of 120 amino acids. The mature protein had the same 13 cysteines as the other A chains and shared high homology with them. The cDNA encoding the PLA(2) from Naja naja atra was also cloned by the use of the same degenerate primers. The cDNAs encoding the new A chain and the PLA(2) were highly expressed in E. coli as soluble fusion proteins and purified by affinity chromatography. The two recombinant proteins achieved after digestion by X(a) factor showed weak and strong PLA(2) activity, respectively. PMID- 12110938 TI - Analysis of Peptide Mapping Glycosylated Site and Glycan Structure in Ribonuclease B by Liquid Chromatography-Electrospray Mass Spectrometry. AB - Liquid chromatography-electrospray mass spectrometry was utilized to analyze peptide mapping of a glycoprotein ribonuclease B to obtain its primary structure. The glycosylated site was determined by comparison of peptide mapping before and after glycanase treatment.Tandem MS(MS/MS)was performed to analyze the structure of N-linked glycan and deglycosylated peptide. The nature of glycan was determined to be of highmannose type by mass spectrometry after the treatment with alpha-mannosidase. In addition the relative abundance of heterogeneous glycopeptides was quantified. This method is rapid and sensitive for the characterization of glycoproteins with N-linked glycan. PMID- 12110939 TI - Immunomodulated Signaling in Macrophages:Regulation of the MAPK Signaling Pathways by PKA and PKC. AB - Previous experimental investigation indicated that immuno-suppressor activities of suppressor macrophages on T B lymphocytes and NK cells could be prevented by treatment with LPS but the tumoricidal activities of those macrophages could be kept or even enhanced after the same treatment. During this complicated course LPS-mediated immuno-modulation was accompanied by activation of PKC and MAPK signal pathways. In order to explore the effect of another signal on MAPK pathway this model of immuno-modulated macrophage was utilized to study the regulatory effect on the activation of three family members of MAPK (ERK1/2 JNK and p38) by cAMP/PKA and PMA/PKC. The results showed that 1) LPS-mediated immuno-modulation was accompanied by dynamic changes of intracellular cAMP amount and PKA activity. 2) A specific PKC activator PMA induced strongly the activation of ERK1/2 JNK and p38 MAPK. 3) In contrast the activation of cAMP/PKA mediated a significant inhibiton of the phosphorylation of ERK1/2 JNK and p38 MAPK and ATF-2 but it enhanced the phosphorylation of CREB. These results suggest that a complicated "cross-talk" may exist among PKC and PKA and MAPK signaling pathways in the regulation of murine peritoneal suppressor macrophages by LPS. PMID- 12110940 TI - Cloning and Expression of Human Manganese-superoxide Dismutase cDNA. AB - The cDNA encoding human manganese-superoxide dismutase was amplified from the human liver cell (L02) total RNA by RT-PCR and sequenced. The human Mn-SOD cDNA was ligated into expression vector pET-24a(+) under T7 promoter. After 5 h induction with 1 mmol/L IPTG 800 mol/L Mn(2+) human Mn-SOD was highly expressed in E.coli BL21(DE3). The protein product was up to 50% of the bacteria total protein in soluble form and had specific SOD activity. PMID- 12110941 TI - Gene Expression Profiles of Human Fetal Nasopharyngeal Tissue. AB - To study differentially expressed genes in nasopharynx tissues of embryo during development, and to observe the gene changes, total RNAs were respectively extracted from the nasopharynx tissue which came from fetuses of 5, 6, 7, and 8 month, Probes were yielded by reverse transcription and were used to hybridize with the Atlas(TM) human cDNA expression arrays. The results showed that the genes expression profile were distinctly different and different expression levels were found in the same gene during development. Results indicated that gene expression pattern had a character of time-dependence, such as early growth response protein 1 and the cDNA expression array provided a powerful method for studying gene expression profile in a large range of genes. PMID- 12110942 TI - Adsorption on Cellulose of the Schizophyllum commune Cellobiose Dehydrogenase. AB - Equilibrium of adsorption of cellobiose dehydrogenase (CDH) of Schizophyllum commune on cotton cellulose, microcrystalline cellulose, and phosphate-swollen cellulose was reached within 4 min. The binding isotherms of CDH adsorbed on the above substrates were a straight line in Scatchard plot analysis, indicating that the binding fitted to the one-binding-site model. It was found that CDH adsorption onto microcrystalline cellulose was strongly affected by temperature, pH, NaCl, and ethylene glycol, respectively. PMID- 12110943 TI - The Relationship between Yeast Coexpressed Gene Clusters and Their Upstream cis acting Elements. AB - DNA microarrays bring biology a new approach to study functions of genes and genomes from their expression pattern on a genomic scale. With its fully sequenced genome and newly published expression patterns available, budding yeast (Saccharomyces cerevisiae) was chosen to carry out an investigation of the relationship between gene 5' upstream cis-acting elements and expression patterns using bioinformatic tools. Results show that genes in the same cluster share common cis-acting element candidates and can be regulated by same transfactors. In the sites found by this study, some sites are corresponding to known cis acting elements, while others may indicate new ones that can be tested by experiments. The results are helpful to understand more about gene functions, metabolic pathways and genetic networks. PMID- 12110944 TI - Depletion of the other genome-mitochondrial DNA depletion syndromes in humans. AB - We present the current knowledge on the genetic and phenotypic aspects of mitochondrial DNA depletion syndromes. The human mitochondrial DNA encodes 13 of the 82 structural proteins of the mitochondrial electron transport chain. The replication and maintenance of the mtDNA require a large number of nuclear encoded enzymes and balanced nucleotide pools. Mitochondrial nucleotide synthesis is of major importance because of the constant need for nucleotides for mtDNA maintenance even in quiescent cells. As de novo enzymes are not present in the mitochondria, synthesis is accomplished via the salvage pathway. Defective mtDNA synthesis and maintenance manifest by multiple deletions or by depletion of the mitochondrial genome. Patients with multiple deletions typically present with progressive external ophthalmoplegia, ptosis and, exercise intolerance after the first decade of life. mtDNA depletion is usually an infantile disease characterized by severe muscle weakness, hepatic failure, or renal tubulopathy with fatal outcome. Linkage analysis in families with multiple mtDNA deletions reveal mutations in proteins that participate in mtDNA replication, the mitochondrial DNA polymerase gene, and the Twinkle gene, a putative mitochondrial helicase and in factors which play a role in mitochondrial nucleotide metabolism, the adenine nucleotide translocator, and the thymidine phosphorylase gene. We have recently identified mutations in an additional two essential proteins in the nucleotide salvage pathway, the mitochondrial deoxyribonucleoside kinases. The phenotype was distinctive for each gene, with hepatic failure and encephalopathy associated with mutations in the deoxyguanosine kinase gene and isolated devastating myopathy as the sole manifestation of thymidine kinase 2 deficiency. The tissue selectivity of these disorders and especially the exclusive muscle involvement in thymidine kinase 2 mutations is puzzling. The normal sequence of the remaining mtDNA copies in spite of a serious mitochondrial nucleotide imbalance is also unexpected. We propose several tissue-specific protective mechanisms and a time window, likely encompassing fetal life and even early infancy, during which nuclear nucleotide synthesis provides mitochondrial needs in all organs. We also speculate on future genes to be discovered in other phenotypes of mtDNA depletion. PMID- 12110945 TI - CD4(+) T helper cells and the role they play in viral control. AB - The natural history of untreated HIV-1 infection is characterized by progressive erosion of the immune system with eventual loss of viral control. T helper cell responses to HIV-1 are typically weak or absent in chronically HIV-1 infected individuals. CD4(+) T helper cell responses have been shown to be important in a number of experimental viral infection systems, and here we explore the ways in which T helper cells might aid in the control of viruses in general and HIV-1 in particular. We first review the relationship between T helper cells and other arms of the immune system. We then focus on the role T helper cells play in viral control in murine and nonhuman primate models, and finally review what is known of CD4(+) T helper cell function in HIV-1 infection. PMID- 12110946 TI - Functional relationships between three novel homozygous mutations in the ACTH receptor gene and familial glucocorticoid deficiency. AB - Familial glucocorticoid deficiency (FGD) is an autosomal recessive disorder characterized by a glucocorticoid adrenal insufficiency without mineralocorticoid deficiency. Mutations of the ACTH receptor (MC2-R) gene have been reported in some FGD cases, but only a few of them have been functionally studied. We reported clinical features and MC2-R gene analysis in three families. For each proband, an homozygous mutation was identified after amplification and sequencing of the whole intronless MC2-R gene. One mutation converted Val-142 located in the second intracellular loop to Leu. Another mutation in the sixth transmembrane domain converted Ala-233 to Pro. The last mutation converted the negatively charged Asp-103 in the first extracellular loop to an uncharged Asn. Functional studies of these mutations as well as the S120R mutation were performed after stable transfection of M3 cells and measurement of ACTH-induced cAMP production. For the S120R, V142L, and A233P mutated MC2-R, cAMP production curves were similar to that obtained with M3 parental cells, confirming that these mutations are responsible for the FGD in the affected patients. The D103N-mutated MC2-R had an impaired cAMP response to physiological doses of ACTH, but the maximal response at very high concentrations of ACTH was similar to that obtained for the wild-type MC2-R. All these results demonstrated clear relationships based on functional studies between MC2-R homozygous mutations and FGD phenotype. PMID- 12110947 TI - Mutations in the cardiac myosin-binding protein C gene are the predominant cause of familial hypertrophic cardiomyopathy in eastern Finland. AB - Hypertrophic cardiomyopathy (HCM) is a genetic disorder characterized by cardiac hypertrophy caused by mutations in genes encoding sarcomere proteins. This study screened all patients with HCM from the Kuopio University Hospital region in eastern Finland for variants in the cardiac myosin-binding protein C gene ( MYBPC3). All 35 exons of MYBPC3 were screened by the single-strand conformation polymorphism method in 37 unrelated patients with HCM. In MYBPC3 we identified seven novel (Gln1061X, IVS5-2A-->C, IVS14-13G-->A, Ex25DeltaLys, Pro147Leu, Ser236Gly, and Arg1138His) and two previously reported (Arg326Gln, Val896Met) variants, all of which are predicted to affect the structure of the encoded protein. Four of the nine variants, a nonsense mutation Gln1061X, a splice acceptor mutation (IVS5-2A-->C), a novel substitution in intron 14 (IVS14-13G- >A), and a novel 3-bp deletion in exon 25 (Ex25DeltaLys) were concluded to be disease-causing mutations because they cosegregated with the HCM phenotype or were absent in more than 200 normal chromosomes, or both. The mutation Gln1061X was found most frequently, being present in 6 families (23 subjects) while the other three mutations were found in single families each. Haplotype analysis indicated a likely founder effect among the families carrying the Gln1061X mutation. We found four novel mutations in MYBPC3, accounting for approx. 38% of familial and 24% of all cases of HCM. In our previous and unpublished studies no more frequent cause of HCM has been found in genetic analyses of other eight sarcomeric proteins. Consequently MYBPC3 is the predominant gene for HCM in eastern Finland. In addition, several amino acid substitutions in MYBPC3 suspected to be not associated with HCM were identified, indicating that some of the missense variants found in MYBPC3 are possibly not disease-causing mutations. PMID- 12110948 TI - Mutations in hepatocyte nuclear factor 4alpha (HNF4alpha) gene associated with diabetes result in greater loss of HNF4alpha function in pancreatic beta-cells than in nonpancreatic beta-cells and in reduced activation of the apolipoprotein CIII promoter in hepatic cells. AB - Mutations in the HNF4alpha gene have been correlated with maturity-onset diabetes of the young, which is characterized mainly by pancreatic beta-cell dysfunction and is also associated with mild liver abnormalities. HNF4alpha D126Y and D126H mutations were found in a patient with early-onset type 2 diabetes, and the R324H mutation was found in a common type 2 diabetic nephropathic patient. We investigated whether these mutations, which have not yet been functionally characterized, impair HNF4alpha function in three cell models: HEK 293 embryonal kidney cells, HepG2 hepatoma cells, and betaTC3 pancreatic beta-cells. The R324H mutation had no effect on HNF4alpha function with either the HNF1alpha and L-type pyruvate kinase (LPK) promoters, but the D126Y and D126H mutations impaired HNF4alpha transcriptional activities in all tested cell lines. These impairments by D126Y and D126H mutations, which are located in the T box, are not due to a loss of dimerization but to a loss of DNA binding. Interestingly, the strongest functional consequences of these mutations were observed on the HNF1alpha promoter in betaTC3 cells. Given the key role of the transcription factor HNF1alpha in pancreatic beta-cell function, it can be inferred that impairment of HNF4alpha function by these mutations affects metabolic pathways in pancreatic beta-cells and contributes to development of diabetes. Moreover, the HNF4alpha mediated activation of the apolipoprotein CIII promoter in HepG2 cells was significantly impaired by D126Y and D126H mutations. These results support clinical findings that liver function can also be impaired in diabetic patients having HNF4alpha mutations. PMID- 12110949 TI - Expression and mutation analysis of BRUNOL3, a candidate gene for heart and thymus developmental defects associated with partial monosomy 10p. AB - Partial monosomy 10p is a rare chromosomal aberration. Patients often show symptoms of the DiGeorge/velocardiofacial syndrome spectrum. The phenotype is the result of haploinsufficiency of at least two regions on 10p, the HDR1 region associated with hypoparathyroidism, sensorineural deafness, and renal defects (HDR syndrome) and the more proximal region DGCR2 responsible for heart defects and thymus hypoplasia/aplasia. While GATA3 was identified as the disease causing gene for HDR syndrome, no genes have been identified thus far for the symptoms associated with DGCR2 haploinsufficiency. We constructed a deletion map of partial monosomy 10p patients and narrowed the critical region DGCR2 to about 300 kb. The genomic draft sequence of this region contains only one known gene, BRUNOL3 ( NAPOR, CUGBP2, ETR3). In situ hybridization of human embryos and fetuses revealed as well as in other tissues a strong expression of BRUNOL3 in thymus during different developmental stages. BRUNOL3 appears to be an important factor for thymus development and is therefore a candidate gene for the thymus hypoplasia/aplasia seen in partial monosomy 10p patients. We did not find BRUNOL3 mutations in 92 DiGeorge syndrome-like patients without chromosomal deletions and in 8 parents with congenital heart defect children. PMID- 12110950 TI - Stimulation of human natural interferon-alpha response via paramyxovirus hemagglutinin lectin-cell interaction. AB - A lectin-carbohydrate recognition event without enzymatic function is proposed as molecular basis for an important innate immune response to enveloped viruses. It involves the hemagglutinin-neuraminidase (HN) glycoprotein of Newcastle disease virus (NDV) and sialic acid expressing cellular receptors on human natural interferon (IFN) alpha producing cells. This conclusion is based on two types of experimental evidence: (a) strong UV irradiation of NDV, which destroyed the cell binding and hemadsorption (HAd) but not the neuraminidase (NA) activity of HN, also destroyed its IFN-alpha inducing activity; (b) DNA transfectants expressing HN mutant molecules with greatly reduced NA but not HAd activity induced IFN alpha while transfectants expressing HN mutant molecules with greatly reduced NA and HAd activity were incapabable of inducing IFN-alpha in human peripheral blood mononuclear cells. The results clarify molecular mechanisms involved in pattern recognition during innate immune responses. PMID- 12110951 TI - Association of polymorphisms of the estrogen receptor alpha gene with bone mineral density of the femoral neck in elderly Japanese women. AB - The estrogen receptor alpha gene is a candidate locus for genetic influence on bone mass. The possible association between two polymorphisms in the first intron of this gene, alone or in combination, and bone mineral density at various sites was examined in participants in the National Institute for Longevity Sciences Longitudinal Study of Aging, a population-based prospective cohort study of aging and age-related diseases. The relationship of the TC ( PvuII) and AG ( XbaI) polymorphisms in the first intron of the estrogen receptor alpha gene to bone mineral density was determined in 2230 subjects (1120 men, 1110 women) and in 2238 subjects (1128 men, 1110 women), respectively, all of whom were community dwelling individuals aged 40-79 years. Bone mineral density at the radius was measured by peripheral quantitative computed tomography and that for the lumbar spine, right femoral neck, right trochanter, right Ward's triangle, and total body was measured by dual-energy X-ray absorptiometry. Estrogen receptor alpha genotypes were determined with an automated fluorescent allele-specific DNA primer assay system. Analysis of the TC ( PvuII) polymorphism revealed that bone mineral density for the total body, femoral neck, and trochanter was significantly lower in women aged 60 years or over with the CC genotype than in those with the TT genotype, but statistical significance was not achieved after adjustment for age, body mass index, and smoking status. Analysis of the AG ( XbaI) polymorphism revealed that bone mineral density for the femoral neck was significantly lower in women aged 60 years or over with the GG genotype than in those with the AA genotype. After adjustment for age, body mass index, and smoking status, bone mineral density for the femoral neck was significantly lower in women aged 60 years or over with the GG genotype than in those with the AA or AG genotypes. Analysis of combined genotypes in women aged 60 years or over revealed that bone mineral density for the femoral neck was significantly lower in women with the CC/ GG genotype than in those with the TT/ AA or TC/ AA genotypes. After adjustment for age, body mass index, and smoking status, bone mineral density for the femoral neck was significantly lower in women aged 60 years or over with the CC/ GG genotype than in those with other genotypes. No differences in bone mineral density at the various sites were detected among TC ( PvuII), AG ( XbaI), or combined genotypes in women aged under 60 years or in men. These results suggest that the estrogen receptor alpha gene is a susceptibility locus for bone mass, especially for the femoral neck, in elderly Japanese women. PMID- 12110952 TI - Differential changes in glutamate concentration in the primate prefrontal cortex during spatial delayed alternation and sensory-guided tasks. AB - Glutamate is a major neurotransmitter in the mammalian brain and glutamatergic neurotransmission in the frontal cortex is indicated to play important roles in cognitive operations. We previously examined changes in extracellular dopamine in the primate frontal cortex in cognitive tasks, and in this paper we extend this to glutamate. We employed, as cognitive tasks, a delayed alternation task where the animal must retain information in working memory, and a sensory-guided task in which there is no working memory requirement but there may be more sensory processing requirements. Using the in vivo microdialysis method, we examined changes in extracellular glutamate concentration in the dorsolateral, arcuate, orbitofrontal, and premotor areas of the primate frontal cortex. Compared to basal rest levels, we observed significant increases in glutamate concentration in dorsolateral and arcuate areas of the prefrontal cortex during the sensory guided task, but did not find significant changes in any of the frontal areas examined during the delayed alternation task. When glutamate concentration was compared between the delayed alternation and sensory-guided tasks, difference was observed only in the dorsolateral prefrontal cortex, especially in the ventral lip area of the principal sulcus. The results indicate the importance of glutamate in processing sensory information but not in retaining information in working memory in the primate dorsolateral and arcuate prefrontal cortex. We also compared the concentration of glutamate and dopamine in the tasks. We found a double dissociation in the concentration of glutamate and dopamine in the dorsolateral area: there was an increase in glutamate but no change in dopamine during the sensory-guided task, whereas there was an increase in dopamine but no change in glutamate during the delayed alternation task. It is thus suggested that in the primate dorsolateral prefrontal cortex, increased glutamate tone without dopamine increase facilitates sensory-guided task performance, while increased dopamine tone without glutamate increase is beneficial for working memory task performance. PMID- 12110954 TI - Effect of GABAB receptor blockade on receptive fields of raccoon somatosensory cortical neurons during reorganization. AB - Single unit recordings were made in the fourth digit representation of raccoon somatosensory cortex after amputation of the fourth digit. The receptive fields of neurons in this "reorganized" cortex were studied before and after microiontophoretic application of a highly specific GABA(B) receptor antagonist, CGP 55845. When the recordings were performed early in the reorganization process, 3-5 weeks after amputation, CGP 55845 produced a greater expansion of the receptive field (184%) than at longer post-amputation intervals (17-34 weeks, 137% increase) or previously reported in intact animals (105%). Most of the receptive fields in these animals were restricted to either an adjacent digit or the palm and the expansion was adjacent to the predrug receptive field. However, in some cases block of GABA(B) receptors unmasked new responsive areas that were separated from the predrug field. In addition, GABA(B) receptor block often revealed higher threshold fields on an adjacent digit or palm. These results reinforce previous studies that used a GABA(A) receptor antagonist, and together they reveal the importance of GABAergic synapses in regulating the various inputs to cortical somatosensory neurons during reorganization. PMID- 12110953 TI - Effect of aging on the human initial interaural linear vestibulo-ocular reflex. AB - To determine age-related changes, the initial linear vestibulo-ocular reflex (LVOR) of eight older subjects of mean age 65+/-7 years (mean +/- SD, range 56-75 years) was compared with that of nine younger subjects of mean age 24+/-5 years (range 18-31 years) in response to random transients of whole-body heave (interaural) translation at peak acceleration of 0.5 g delivered by a pneumatic actuator. Binocular eye rotations were measured with magnetic search coils, while linear head position and acceleration were measured with a potentiometer and piezoelectric accelerometer. Subjects viewed targets 200 cm, 50 cm, or 15 cm distant immediately before the unpredictable onset of randomly directed translation in darkness (LVOR) and in light (LVVOR). All subjects maintained ideal vergence of 1.5-2 degrees for the 200-cm target, 6-8 degrees for the 50-cm target, and 21-26 degrees for the 15-cm target, with actual vergences depending on individual interpupillary distances. Search coil recording of angular position of the upper teeth showed head rotation to be negligible (less than 0.5 degrees ) for the first 250 ms after onset of head translation, excluding a role for the angular VOR in the responses studied. The LVOR response to heave translation was an oppositely directed eye rotation occurring after a mean latency of 62+/-3 ms for older and 42+/-3 ms (mean +/- SD) for younger subjects ( P<0.0001). The peak of the latency distribution was 60-100 ms for older and 20-60 ms for younger subjects. During the early interval, 70-80 ms from head motion onset prior to a pursuit contribution or saccades, all subjects had significantly enhanced LVOR with decreasing target distance. In this interval, the LVOR position amplitude of younger subjects was 0.17+/-0.01 degrees, 0.40+/-0.01 degrees, 0.57+/-0.01 degrees (mean +/- SE), respectively, in descending order of target distance. Early sensitivities were significantly reduced for older subjects to 0.07+/-0.01 degrees, 0.23+/-0.01 degrees, 0.40+/-0.01 degrees ( P<0.0001). There was no significant effect of target visibility in either group during the first 110 ms ( P>0.05). Visual-otolith interaction was mainly reflected not by the vestibular slow phase, but by vestibular catch-up saccades (VCUS) in the compensatory direction. The effect of aging on the initial human LVOR is thus to: prolong latency, reduce early sensitivity, and reduce occurrence of vestibular catch-up saccades. PMID- 12110955 TI - Two-dimensional analysis of the spacing of ocular dominance columns in normally raised and strabismic kittens. AB - In the primary visual cortex of cats, ferrets and macaque monkeys, the thalamocortical afferents conveying signals from the two eyes terminate in alternating regions of layer IV known as ocular dominance columns. Previous experiments have indicated that the periodicity of these columns can be influenced by visual experience: compared to normally raised animals both strabismic cats and cats raised with alternating monocular exposure displayed an increased spacing of adjacent ocular dominance columns in the primary visual cortex (area 17). However, recently it was shown that the formation of ocular dominance columns begins much earlier than previously supposed, indicating that early visual experience might only have a limited influence on the development of the spatial pattern of ocular dominance columns. We therefore visualized the complete pattern of ocular dominance columns in area 17 of normally raised and strabismic kittens during early postnatal development (age 3-6 weeks), particularly focussing on littermates. In addition, we used a previously developed spatial analysis (period statistics) to quantify columnar spacing two dimensionally. We observed a pronounced interindividual variability in both normally raised and strabismic animals, with column spacings ranging from 783 to 1362 microm. In contrast to previous reports, there were no significant differences in columnar periodicity between normally raised and strabismic cats. These data indicate that rearing has less influence on column spacing while the interindividual variability is much greater than previously supposed, suggesting that genetic differences have an influence on column spacing. PMID- 12110956 TI - Non-invasive regime for language lateralization in right- and left-handers by means of functional MRI and dichotic listening. AB - Language lateralization was assessed by two independent functional techniques, fMRI and a dichotic listening test (DLT), in an attempt to establish a reliable and non-invasive protocol of dominance determination. This should particularly address the high intraindividual variability of language lateralization and allow decision-making in individual cases. Functional MRI of word classification tasks showed robust language lateralization in 17 right-handers and 17 left-handers in terms of activation in the inferior frontal gyrus. The DLT was introduced as a complementary tool to MR mapping for language dominance assessment, providing information on perceptual language processing located in superior temporal cortices. The overall agreement of lateralization assessment between the two techniques was 97.1%. Conflicting results were found in one subject, and diverging indices in ten further subjects. Increasing age, non-familial sinistrality, and a non-dominant writing hand were identified as the main factors explaining the observed mismatch between the two techniques. This finding stresses the concept of an intrahemispheric distribution of language function that is obviously associated with certain behavioral characteristics. PMID- 12110957 TI - Modulation of visual cortical excitability in migraine with aura: effects of 1 Hz repetitive transcranial magnetic stimulation. AB - Recent studies showed hyperexcitability of the occipital cortex in subjects affected by migraine with aura. It has been shown that 1 Hz repetitive transcranial magnetic stimulation (rTMS) reduces excitability of visual cortex in normal subjects. The aim of the study was to investigate the effects of low frequency (1 Hz) rTMS on visual cortical excitability by measuring changes in phosphene threshold (PT) in subjects with migraine with aura. Thirteen patients with migraine with aura and 15 healthy controls were examined. Using a standardized transcranial magnetic stimulation protocol of the occipital cortex, we assessed the PT (the lowest magnetic stimulation intensity at which subjects just perceived phosphenes) before and after a 1-Hz rTMS train delivered at PT intensity for 15 min. The difference in the proportion of subjects reporting phosphenes in migrainer and control groups was significant (migrainers: 100% vs controls 47%; P<0.05), and 1 Hz rTMS over the occipital cortex led to a significantly increased visual cortex excitability expressed as a decrease in PT in subjects affected by migraine with aura. Conversely, after a 1-Hz TMS train normal subjects showed increased PT values, which suggests a decreased visual cortex excitability. Our findings confirm that the visual cortex is hyperexcitable in migrainers and suggest a failure of inhibitory circuits, which are unable to be upregulated by low frequency rTMS. PMID- 12110958 TI - Systemic acetyl-L-carnitine eliminates sensory neuronal loss after peripheral axotomy: a new clinical approach in the management of peripheral nerve trauma. AB - Several hundred thousand peripheral nerve injuries occur each year in Europe alone. Largely due to the death of around 40% of primary sensory neurons, sensory outcome remains disappointingly poor despite considerable advances in surgical technique; yet no clinical therapies currently exist to prevent this neuronal death. Acetyl- L-carnitine (ALCAR) is a physiological peptide with roles in mitochondrial bioenergetic function, which may also increase binding of nerve growth factor by sensory neurons. Following unilateral sciatic nerve transection, adult rats received either one of two doses of ALCAR or sham, or no treatment. Either 2 weeks or 2 months later, L4 and L5 dorsal root ganglia were harvested bilaterally, in accordance with the Animal (Scientific Procedures) Act 1986. Neuronal death was quantified with a combination of TUNEL [TdT (terminal deoxyribonucleotidyl transferase) uptake nick end labelling] and neuron counts obtained using the optical disector technique. Sham treatment had no effect upon neuronal death. ALCAR treatment caused a large reduction in the number of TUNEL positive neurons 2 weeks after axotomy (sham treatment 33/group; low-dose ALCAR 6/group, P=0.132; high-dose ALCAR 3/group, P<0.05), and almost eliminated neuron loss (sham treatment 21%; low-dose ALCAR 0%, P=0.007; high-dose ALCAR 2%, P<0.013). Two months after axotomy the neuroprotective effect of high-dose ALCAR treatment was preserved for both TUNEL counts (no treatment five/group; high-dose ALCAR one/group) and neuron loss (no treatment 35%; high-dose ALCAR -4%, P<0.001). These results provide further evidence for the role of mitochondrial bioenergetic dysfunction in post-traumatic sensory neuronal death, and also suggest that acetyl- L-carnitine may be the first agent suitable for clinical use in the prevention of neuronal death after peripheral nerve trauma. PMID- 12110959 TI - Differential distribution of interneurons in the neural networks that control walking in the mudpuppy (Necturus maculatus) spinal cord. AB - Locomotor behavior is believed to be produced by interneuronal networks that are intrinsically organized to generate the underlying complex spatiotemporal patterns. In order to study the temporal correlation between the firing of individual interneurons and the pattern of locomotion, we utilized the spinal cord-forelimb preparation from the mudpuppy, in which electrophysiological recordings of neuronal activity were achieved during walking-like movement of the forelimb induced by bath application of N-methyl- D-aspartate (NMDA). Intra- and extracellular recordings were made in the C2 and C3 segments of the spinal cord. These segments contain independent flexor and extensor centers for the forelimb movement about the elbow joint during walking. Among the 289 cells recorded in the intermediate gray matter (an area between the ventral and dorsal horns) of the C2 and C3 segments, approximately 40% of the cells fired rhythmically during "walking." The firing rates were 6.4+/-0.4 impulses/s (mean +/- SE). These rhythmically active cells were classified into four types based on their phase of activity during a normalized step cycle. About half the rhythmic cells fired in phase with either the flexor (F) or extensor (E) motoneurons. The rest fired in the transitions between the two phases (F-->E and E-->F). Longitudinal distributions of the four types of interneurons along the spinal cord were in agreement with observations that revealed distinct but overlapping flexor and extensor centers for walking. Some cells triggered short-latency responses in the elbow flexor or extensor muscles and may be last-order interneurons. These observations suggest that there is a differential distribution of phase-specific interneurons in the central pattern generator of the mudpuppy spinal cord for walking. PMID- 12110960 TI - Neurons with radial receptive fields in monkey area V4A: evidence of a subdivision of prelunate gyrus based on neuronal response properties. AB - In recordings from two awake, behaving macaque monkeys we found that neurons in the crown of the prelunate gyrus differed in their responsiveness to simple visual stimuli. Neurons in the posterior part of the gyrus (area V4) responded strongly to stationary or moving bars, while neurons in the anterior part (area V4A) responded only weakly to such stimuli. Most receptive fields in area V4A were elongated with long axes oriented radially towards the fovea. These neurons were sensitive to radial movements, especially to sudden shifts of real 3D objects. The border between areas V4 and V4A coincided with the representation of the horizontal meridian. Area V4A extended into the posterior bank of the superior temporal sulcus, where its border corresponded to the representation of the vertical meridian. The sequence of the representations of the horizontal and vertical meridians over the prelunate gyrus suggests the existence of another area between V4A and V4t. PMID- 12110961 TI - Digital nerve anaesthesia decreases EMG-EMG coherence in a human precision grip task. AB - There is increasing evidence that the primary motor cortex is involved in the generation of electromyographic (EMG) oscillations at frequencies in the range of 15-30 Hz that are observed during performance of a precision grip task. Since the level of the corticomuscular coherence varies according to the nature of the object that is gripped, it seemed possible that somatosensory inputs from the hand might affect this coherence. The aim of this study was to investigate whether interrupting cutaneous inputs from the digits would affect the coherence between hand muscles during precision grip of a compliant object. Subjects performed a precision grip hold-ramp-hold task before, during and after digital nerve anaesthesia of the index finger and thumb. There were marked deficits in the performance of the task, particularly during the initial formation of the grip and first hold period. Local digital nerve anaesthesia reduced but did not abolish 14-31 Hz coherence between EMG activity recorded from different hand and forearm muscles. Coherence was measured during the second hold phase of the task. Digital nerve anaesthesia did not affect the predominant frequencies in the EMG power spectra compiled from the same phase of the task. We conclude that during a precision grip task, cutaneous input enhances oscillatory synchrony between pairs of hand muscles. PMID- 12110962 TI - Percept-related changes in horizontal optokinetic nystagmus at different body orientations in space. AB - Large-field motion of the visual environment is a powerful stimulus to induce the perception of contra-directional self-motion in a stationary observer. We investigated the interrelations between horizontal optokinetic nystagmus and subjective states of motion perception under variation of subjects' orientation with respect to gravity. Subjects were tested sitting upright and lying supine, and signalled transitions between object- and self-motion perception whilst viewing an optokinetic stimulus rotating about the subjects' longitudinal axis at a range of angular velocities. Optokinetic stimulation in the supine condition resulted in subjects perceiving a graviceptive conflict and the illusory perception of whole body tilt in a direction opposite to optokinetic stimulus rotation, whereas during upright viewing the axis of stimulus rotation was aligned with the direction of gravity and thus did not result in a conflict or perception of tilt. In both postures, self-motion perception coincided with an increased deviation of mean horizontal gaze position in the perceived direction of heading with a concurrent reduction in optokinetic nystagmus slow-phase gain. Slow-phase gain was also significantly reduced in the supine position as well as at increasing stimulus velocities. The results demonstrate that spontaneous transitions between the perception of object-motion and that of self-motion consistently coincide with spatial attentional and orientational strategies, shifting from passive monitoring to active oculomotor exploration and anticipation. PMID- 12110963 TI - The breakdown of Fitts' law in rapid, reciprocal aiming movements. AB - According to Fitts' law, there is speed-accuracy trade-off in a wide variety of discrete aiming movements. However, it is unknown whether the same law applies to cyclic aiming movements. In the present study, a comparison is made between discrete versus cyclic aiming movements. A group of 24 healthy participants made graphical pen movements in 12 different aiming tasks in which successive finger and wrist movements were emphasized, consecutively executed as discrete and cyclic movements and varying in three target widths. In the cyclic condition, aiming movements consisted of back-and-forth movements that were performed in immediate succession for 20 s. In the discrete condition, back-and-forth aiming movements were drawn as 20 single strokes, starting after a go signal and stopping after reaching the target area. The targets had various levels of spatial accuracy and the movements had different directions (from lower left to upper right; from lower right to upper left) elicit either predominantly wrist or finger movements. The amount of information processed per unit of time (bits per second; index of performance, IP), tangential velocity, the pen pressure, and the ratio of peak-over-mean velocity were studied to gain understanding about the differences in control between discrete and cyclic movements. It was found that the IP and movement velocity were almost twice as large in cyclic versus discrete movements. In contrast, the axial pen pressure and the ratios of peak-over-mean velocity were much lower in cyclic movements (1.24 N versus 0.94 N; 2.26 N versus 1.81 N). The results of our study indicate that the predicted constant IP does not hold for rapid cyclic aiming movements and that speed-accuracy trade-off is different. It is concluded that cyclic movements exploit the energetic and physiological properties of the neuromotor system. Expected differences in brain activity related to discrete and cyclic aiming movements are discussed as well as several neurophysiological mechanisms, which predict more economic force recruitment and information processing in cyclic than in discrete movements. PMID- 12110964 TI - Conflicting sources of spatial information in a distance-reproduction task. AB - Previous research has shown that the reproduction of a criterion distance is biased towards previously coded endpoints. The purpose of this research was to illustrate that, in addition to the retention of endpoint information, the presence of conflicting sources of spatial information within a trial causes systematic response biases in distance reproduction. Three experiments were conducted in which participants performed rapid aiming movements on a digitising tablet that translated to movement of a cursor on a computer monitor. The required movement amplitude in all three experiments was 20 cm. In experiment 1, the location of the home and target positions on the monitor was fixed, but the initial position of the hand was varied randomly from trial to trial. In experiment 2, the change in position of the limb was matched by a corresponding change in the location of the monitor display. In experiment 3, the initial position of the limb was fixed, but the location of the display on the monitor varied from trial to trial. The results of experiments 1 and 2 showed that error varied as a function of the initial position of the limb. However, this effect was greater in experiment 1, where the mapping between the location of the monitor display and limb position varied from trial to trial. There was also an effect of varying the location of the monitor display in experiment 3, but this was smaller than varying initial limb position in experiment 1. These findings suggest that both the retrieval of previously specified endpoints and conflicts in the coding of spatial information contributed to the observed response biases in distance reproduction. PMID- 12110965 TI - Haptic discrimination of object shape in humans: two-dimensional angle discrimination. AB - The human ability to recognize objects on the basis of their shape, as defined by active exploratory movements, is dependent on sensory feedback from mechanoreceptors located both in the skin and in deep structures ( haptic feedback). Surprisingly, we have little information about the mechanisms for integrating these different signals into a single sensory percept. With the eventual aim of studying the underlying central neural mechanisms, we developed a shape discrimination test that required active exploration of objects, but was restricted to one component of shape, two-dimensional (2D) angles. The angles were machined from 1-cm-thick Plexiglas, and consisted of two 8-cm-long arms that met to form an angle of 90 degrees (standard) or 91 degrees to 103 degrees (comparison angles). Subjects scanned pairs of angles with the index finger of the outstretched arm and identified the larger angle of each pair explored. Discrimination threshold (75% correct) was 4.7 degrees (range 0.7 degrees to 12.1 degrees), giving a precision of 5.2% (0.8-13.4%: difference/standard). Repeated blocks of trials, either in the same session or on different days, had no effect on discrimination threshold. In contrast, the motor strategy was partly modified: scanning speed increased but dwell-time at the intersection did not change. Finally, 2D angle discrimination was not significantly modified by rotating the orientation of one of the angles in the pair (0 degrees, 4 degrees or 8 degrees rotation towards the midline, in the vertical plane), providing evidence that subjects evaluated each angle independently in each trial. Subject reports indicated that they relied on cutaneous feedback from the exploring digit (amount of compression of the finger at the angle) and mental images of the angles, most likely arising from proprioceptive information (from the shoulder) generated during the to-and-fro scans over the angle. In terms of shoulder angles, the mean discrimination threshold here was 0.54 degrees (range 0.08 degrees to 1.36 degrees). These values are lower than previous estimates of position sense at the shoulder. In light of the subjects' strategies, it therefore seems likely that both cutaneous and proprioceptive (including both dynamic and static position related signals) feedback contributed to the haptic discrimination of 2D angles. PMID- 12110966 TI - Haptic discrimination of object shape in humans: contribution of cutaneous and proprioceptive inputs. AB - Using two-dimensional (2D) angles composed of two straight, 8-cm-long arms that formed an angle, we investigated the importance of cutaneous feedback from the exploring index finger, and proprioceptive feedback from the shoulder (scanning movements made with the outstretched arm), to the human ability to discriminate small differences in the angles. Using a two-alternative forced-choice paradigm, subjects identified the larger angle in each pair explored (standard angle, 90 degrees; comparison angles, 91 degrees to 103 degrees). Subjects were tested under four experimental conditions: (1) active touch (reference condition); (2) active touch with digital anaesthesia; (3) passive touch (a computer-controlled device displaced the angle under the subject's immobile digit); and (4) passive touch with digital anaesthesia. When only proprioceptive feedback from the shoulder was available (condition 2), there was a significant increase in discrimination threshold, from 4.0 degrees in the reference condition (condition 1) to 7.2 degrees, indicating that cutaneous feedback from the exploring digit contributed to task performance. When only cutaneous feedback from the finger was available (condition 3), there was also a significant increase in threshold from 4.2 degrees in the active condition to 8.7 degrees. This suggested that proprioceptive feedback from the shoulder, potentially from a variety of deep (muscle and joint) but also cutaneous receptors, contributed to the ability to discriminate small changes in 2D angles. When both sources of feedback were eliminated (condition 4), subjects were unable to discriminate even the largest difference presented (13 degrees). The results suggest that this sensory task is truly an integrative task drawing on sensory information from two different submodalities and so, following the definition of Gibson, is haptic in nature. The results are discussed in relation to the potential neural mechanisms that might underlie a task that requires integration across two anatomically separate body parts and two distinct modalities. PMID- 12110967 TI - C-fos induction in rat superficial dorsal horn following cutaneous application of noxious chemical or mechanical stimuli. AB - The method of c-fos immunodetection was used to map the distributions of neurons in the lumbar spinal dorsal horn that were activated following intracutaneous (i.c.) microinjection, or iontophoretic application, of different irritant chemicals to the lateral hindpaw of rats. Microinjections (1 microl) of histamine, serotonin (5-HT), nicotine, capsaicin, or formalin each elicited similar distributions of Fos-like immunoreactivity (FLI) in laminae I-II of the ipsilateral superficial dorsal horn, with little or no FLI in deeper laminae or contralaterally. In laminae I and II, FLI cell counts were significantly higher following i.c. histamine, 5-HT, capsaicin, formalin, and noxious pinch, compared to i.c. saline controls. Capsaicin-evoked FLI was dose-dependent. Multivariate analysis of variance revealed no significant difference in spatial distributions of FLI induced by any of the chemical or pinch stimuli. Iontophoretic application of histamine, 5-HT, or nicotine also elicited similar distributions of FLI in the superficial dorsal horn, and cell counts of FLI were significantly higher compared to controls receiving iontophoretic vehicle (methyl cellulose). These results indicate either that individual laminae I-II neurons are activated by each of the irritant chemicals, or that neurons selectively responsive to a given irritant are comingled without any apparent laminar segregation. PMID- 12110968 TI - Event-related fMRI responses in the human frontal eye fields in a randomized pro- and antisaccade task. AB - We examined whether the frontal eye fields (FEF) are involved in the suppression of reflexive saccades. Simultaneous recording of horizontal eye movements and functional magnetic resonance imaging enabled us to perform a randomized pro- and antisaccade task and to sort blood oxygenation level dependent (BOLD) time series on the basis of task performance. Saccadic reaction time distributions were comparable across tasks indicating a similar effort in preprocessing of the saccades. Furthermore, we found similar BOLD activation in FEF during both correctly performed pro- and antisaccades. Frontal eye field activation started prior to target presentation and saccade generation. While we observed only few erroneous antisaccades, these were associated with a decrease in BOLD activity prior to target presentation, and increased BOLD activity after target presentation relative to correctly performed antisaccades. These findings are consistent with a role of the FEF in the suppression of reflexive saccades. The increase in activity after target presentation for antisaccade errors can only be indirectly linked to such a role but may also reflect activity related to the generation of a correction saccade. Frontal eye field BOLD activity may further represent general arousal, preparatory set, short-term memory, or salience-map related activity. PMID- 12110969 TI - Development of a flow-injection analysis (FIA) enzyme sensor for fructosyl amine monitoring. AB - An enzyme-sensor system with flow-injection analysis (FIA) has been developed for the detection of fructosyl amine compounds; the sensor utilizes fructosyl amine oxidase isolated from the marine yeast Pichia sp. N1-1 strain. With this FIA system 0.2 to 10 mmol L(-1) fructosyl valine can be determined. The sensor is approximately five times more sensitive to fructosyl valine, a model compound for glycated hemoglobin HbA1c, than to N(epsilon)-fructosyl lysine, a model compound for glycated albumin. This FIA system can also be used to detect fructosyl dipeptides. The operational stability of the sensor enabled more than 120 consecutive sample injections over a period of approximately 20 h. PMID- 12110970 TI - Monitoring, and estimation of kinetic data, by piezoelectric impedance analysis, of the binding of the anticancer drug mitoxantrone to surface-immobilized DNA. AB - A new method of monitoring drug binding to DNA, in real-time, by means of the piezoelectric quartz-crystal-impedance (PQCI) technique, is proposed. The method was used to monitor the binding of an anticancer drug, mitoxantrone (MX), to double-stranded calf-thymus DNA covalently immobilized on the thioglycollic acid modified gold electrode surface of a quartz crystal. Optimum experimental conditions for the immobilization were established. The DNA-anchored piezoelectric sensor was in contact with MX solution. The time courses of the resonant frequency and the equivalent circuit characteristics of the sensor were also obtained during the study of DNA-drug binding. On the basis of the analysis of the multidimensional information provided by the PQCI technique the observed frequency decrease was mainly ascribed to the increase in the mass of the sensor surface resulting from the binding. The kinetics of the binding process were studied quantitatively by monitoring the frequency change with time and a piezoelectric response model for the binding was derived theoretically. The experimental data were fitted to the model and the binding and dissociation rate constants and the binding equilibrium constant were estimated to be 66.0+/-0.1 mol(-1) L s(-1), 1.4+/-0.1x10(-4) s(-1), and 4.71+/-0.07x10(5) mol(-1) L, respectively, at 37 degrees C. PMID- 12110971 TI - Preparation of refractive index matching polymer film alternative to oil for use in a portable surface-plasmon resonance phenomenon-based chemical sensor method. AB - In order to simplify the procedure for assembling a surface-plasmon resonance (SPR) sensor, a refractive index matching polymer film was prepared as an alternative to the conventionally used matching oil. The refractive index matching polymer film, the refractive index of which was nearly equal to the prism and sensor chip material (a cover glass) of the SPR sensor, was prepared by casting a tetrahydrofuran solution of poly (vinyl chloride) (PVC) containing equal weights of dioctyl phthalate and tricresyl phosphate. The refractive index matching polymer film was found to have a refractive index of 1.516, which is identical to that of the prism and the cover glass used for the present SPR sensor. The utility of the matching polymer film for the SPR sensor was confirmed by the detection of anti-human albumin, based on an antigen-antibody reaction. PMID- 12110972 TI - A flow-through solid-phase spectroscopic sensing device implemented with FIA solution measurements in the same flow cell: determination of binary mixtures of thiamine with ascorbic acid or acetylsalicylic acid. AB - A continuous and simple UV-photometric flow-through optosensor has been developed for the simultaneous determination of a binary mixture of two species with different electric charges present at very different concentrations - ascorbic acid (or acetylsalicylic acid) and thiamine. The sensing device is based on the selective sorption and determination of a cationic analyte on a cation-exchange gel (Sephadex SP-C25) while the other, anionic, analyte is determined in the solution among the interstices of the cation-exchange gel in the same flow cell. The analytes arrive in sequence at the sensing zone, which detects their intrinsic absorbance at 253 nm, as a result of on-line separation by use of a minicolumn filled with the same cation-exchange gel as in the cell, and placed before the flow cell. Thiamine is retained in the minicolumn whereas ascorbic acid or acetylsalicylic acid pass through it and produce their signal as a result of absorbance in the interstitial solution among the resin beads. Thiamine is then eluted from the precolumn, transported to the flow cell, and temporarily retained in the sensing zone from this eluted solution. Calibration graphs were linear over the range 3-50, 25-400, and 300-3000 microg mL(-1) (600 microL sample volume) and the relative standard deviations were 2.56, 1.85, and 1.25 % for thiamine, ascorbic acid, and acetylsalicylic acid, respectively. The proposed method was satisfactorily applied to the determination of binary mixtures of thiamine with ascorbic acid or acetylsalicylic acid in pharmaceutical preparations and semi-synthetic samples. PMID- 12110973 TI - Determination of total platinum in plasma and plasma ultrafiltrate, from subjects dosed with the platinum-containing N-(2-hydroxypropyl)methacrylamide copolymer AP5280, by use of graphite-furnace Zeeman atomic-absorption spectrometry. AB - AP5280 is an N-(2-hydroxypropyl)methacrylamide (HPMA) copolymer to which are linked tetrapeptide side chains containing bioactive platinum complexes at their C-terminal sides. We have developed and validated a rapid and sensitive analytical assay for the determination of total platinum concentrations in plasma, and free platinum of an AP5280 origin in plasma ultrafiltrate (PUF), of subjects dosed with AP5280. The total platinum levels were determined by use of graphite-furnace atomic-absorption spectrometry (GF-AAS) with Zeeman correction after appropriate dilution of the plasma sample with plasma-hydrochloric acid 0.2 mol L(-1) (1:5) as diluent. The limit of quantitation of this assay is 0.25 micromol L(-1) platinum in plasma. Linear calibration curves were obtained over the concentration range 0.25-5.0 micromol L(-1). Accuracy was between 87.7% and 104.2% and precision was 15.3% at the lowest concentration and less than 14% for all other levels tested. Accuracy and precision were thus in accordance with generally accepted criteria for analytical methods. Analysis of samples obtained from patients receiving AP5280 demonstrated the applicability of the described assay. Analysis of free platinum in PUF was performed by use of a previously validated and reported assay from our institute in which the same instrumental method is used. PMID- 12110974 TI - Speciation of Fe(II) and Fe(III) by the modified ferrozine method, FIA spectrophotometry, and flame AAS after cloud-point extraction. AB - A method has been developed for the simultaneous determination of traces of Fe(III) and Fe(II) in water by on-line coupling of spectrophotometry with flame atomic absorption spectrometry (FAAS). The method involves cloud-point extraction (CPE) of both species with ammonium pyrrolidinecarbodithioate (APDC) under standard conditions, which facilitates the in situ complexation and extraction of both species. Differentiation of the oxidation states of iron is achieved by using mathematical equations to overcome the interference of Fe(III) in the spectrophotometric determination of Fe(II) when they are both present in the same solution. In this manner the time-consuming and labor-intensive steps of preoxidation of Fe(II) or reduction of Fe(III) are eliminated. By preconcentrating a 10-mL sample solution detection limits as low as 7 microg L( 1), were obtained after a single-step extraction procedure. The relative standard deviation (n=4, 30 microg L(-1)) was 2.6 % and 1.8 % for spectrophotometry and FAAS, respectively. Recoveries in the range of 96-105 % were obtained by analysis of spiked real samples. The method was further verified by analyzing a certified reference material (IMEP-9); for this the recovery was 98.5 %. PMID- 12110975 TI - Capability of diatomaceous earth to preconcentrate and store Pb and Cr: on-line determination by FI-FAAS. AB - The diatomaceous earth (DE) has an important ability to retain metals such as Cd, Cr, Mn and Pb, which can be used for their stabilization in the environment and for analytical purposes. In this paper a fast on-line preconcentration method for the determination of Cr and Pb in waters by flow injection flame atomic absorption spectrometry is described. Preconcentration was based on the retention of Cr and Pb on a DE immobilized in silica gel at pH 3.0 and subsequent elution with 200 microL of 3 mol L(-1) HCl. The preconcentration factors were 100 and 150 for Pb and Cr respectively, for 16 mL water sample volume. The detection limits under these conditions were 3 ng mL(-1) and 1 ng mL(-1) for Pb and Cr, respectively. The stability of Cr and Pb retained on silica gel-DE columns was established. Silica gel-DE microcolumns with the retained analytes were stored for 2 months at two different temperatures: 4 degrees C and room temperature. At regular time intervals, both metals were eluted and quantified. The results showed the potential of the procedure for sampling and storing water samples for subsequent metal determination, avoiding the problems associated with maintaining species integrity in aqueous solution, and the possibility to of decontaminating polluted spaces. PMID- 12110976 TI - Comparison of UV derivative-spectrophotometry and partial least-squares (PLS-1) calibration for determination of methotrexate and leucovorin in biological fluids. AB - Binary mixtures of methotrexate (MTX) and leucovorin (LV) have been resolved by application of first-derivative spectrophotometry and partial least squares calibration (PLS-1). By measuring the first-derivative signals of MTX and LV at 354 and 300 nm, respectively, simultaneous determination was possible. The mean recoveries for urine samples were 91 and 96% for MTX and LV, respectively. Partial least squares (PLS-1) multivariate calibration has been applied to the determination of these compounds in serum and in urine without pretreatment of the samples. The absorption spectra of serum or urine samples spiked with methotrexate and/or leucovorin, were used to optimize the calibration matrixes by the PLS-1 method. The sensitivity and selectivity of the proposed procedures were calculated. Mean recoveries were 101 and 97% for MTX and LV, respectively, for serum samples, and 101 and 98% for MTX and LV, respectively, for urine samples. PMID- 12110977 TI - Estimation of the composition of heparin mixtures from various origins using proton nuclear magnetic resonance and multivariate calibration methods. AB - A multivariate calibration method for the characterization of heparin samples based on the analysis of (1)H nuclear magnetic resonance (NMR) spectral data is proposed. Heparin samples under study consisted of two-component or four component mixtures of heparins from porcine, ovine and bovine mucosae and bovine lung. Although the (1)H NMR spectra of all heparin types were highly overlapping, each origin showed some particular features that could be advantageously used for the quantification of the components. These features mainly concerned the anomeric H, which appeared in the range 5.0-5.7 ppm and the peaks of acetamidomethyl protons at 2.0-2.1 ppm. The determination of the percentage of each heparin class depended on these differences and was carried out using partial least squares regression (PLS) as a calibration method. Prior to the PLS analysis, the spectral data were standardized using the internal standard peak (sodium 4,4-dimethyl-4-silapentanoate- 2,2,3,3- d (4), TSP) as the reference. The quantification of each heparin type in the samples using PLS models built with 4 or 5 components was satisfactory, with an overall prediction error ranging from 3% to 10%. PMID- 12110978 TI - Quick measurement of protein sulfhydryls with Ellman's reagent and with 4,4' dithiodipyridine. AB - Since its introduction in 1959, Ellman's reagent (5,5'-dithio-bis(2-nitrobenzoic acid)) has been the favorite reagent for spectrophotometric measurement of protein sulfhydryls. Meanwhile however, evidence has accumulated that many protein sulfhydryls give an incomplete reaction with Ellman's reagent, even during prolonged assay times. In the present study, the kinetic problem was solved by including cystamine as a "mediator" between the protein sulfhydryl and Ellman's reagent, as previously applied in an enzymatic thiol assay [9]. As an alternative, 4,4'-dithiodipyridine (DTDP) was used in place of Ellman's reagent. Due to its small size, amphiphilic nature, and lack of charge, DTDP quickly reacts with poorly accessible protein sulfhydryls, without any catalysis by cystamine. The DTDP method and the Ellman/cystamine method were both optimized for maximal sensitivity, minimal sample consumption (detection limit 0.2 nmol mL( 1), determination limit 0.6 nmol mL(-1)), and minimal assay time (5 min). In validation experiments, both methods gave identical results and the measured sulfhydryls/protein matched the expected values. Electronic supplementary material to this paper can be obtained by using the Springer Link server located at http://dx.doi.org/10.1007/s00216-002-1347-2. PMID- 12110979 TI - Electrocatalytic amperometric determination of amitrole using a cobalt phthalocyanine-modified carbon paste electrode. AB - Cobalt-phthalocyanine-modified carbon paste electrodes are shown to be excellent indicators for electrocatalytic amperometric measurements of triazolic herbicides such as amitrole, at low oxidation potentials (+0.40 V). The detection and determination of amitrole in flow injection analysis with a modified carbon paste electrode with Co-phthalocyanine is described. The concentrations of amitrole in 0.1 M NaOH solutions were determined using the electrocatalytic oxidation signal corresponding to the Co(II)/Co(III) redox process. A detection limit of 0.04 microg mL(-1) (4 ng amitrole) was obtained for a sample loop of 100 microL at a fixed potential of +0.55 V (vs. Ag/AgCl) in 0.1 M NaOH and a flow rate of 4.0 mL min(-1). Furthermore, the modified carbon paste electrodes offers reproducible responses in such a system, and the relative standard deviation was 3.3% using the same surface, 5.1% using different surface, and 6.9% using different pastes. The performance of the cobalt-phthalocyanine-modified carbon paste electrodes is illustrated here for the determination of amitrole in commercial formulations. The response of the electrodes is stable, with more than 80% of the initial retained activity after 50 min of continuous use. PMID- 12110980 TI - Electrochemical detection of parathion at a glassy-carbon electrode modified with hexadecane. AB - A modified electrode, based on a hexadecane (C(16))-coated glassy-carbon electrode (GCE) has been developed for the determination of parathion. The electrochemical behavior of parathion was investigated by cyclic voltammetry (CV). The peak potential and peak current were found to depend on the supporting electrolyte and the pH of the buffer solution. The reduction currents for parathion were proportional to parathion concentration over the range 8x10(-8) 2x10(-5) mol L(-1). The detection limit was 2x10(-8) mol L(-1) parathion for an accumulation time of 30 s. The effects of organic and inorganic species on the determination of parathion were also studied. A procedure was developed to extract parathion from spiked soil samples using a mixture of dichloromethane and acetone as the extraction solvent. The complete extraction and analytical procedure are simple, inexpensive and rapid. Parathion was determined in a soil sample by use of linear scan voltammetry (LSV). PMID- 12110981 TI - Test strip for determination of nitrite in water. AB - A disposable test strip is proposed for the determination of nitrite in waters. The strip is an inert rectangular strip of polyester with a 6 mm o.d. circular, transparent and colorless film attached to its surface. This film contains the chemicals required for reaction and fixation of the dye formed, sulfanilamide, N (1-naphthyl)ethylenediamine on Nafion. When the test strip is placed in an acidified (pH 2.0) sample solution containing nitrite a red-violet color develops; the absorbance of this is measured at 536 nm. The linear range of the method depends on the time of equilibration of the test strip with the sample solution. When the equilibration time was 45 min, the linear range was 8.9-500 microg L(-1) whereas for an equilibration time of 60 min it was 4.7-200 microg L( 1). The detection limit was 1.4 microg L(-1) for an equilibration time of 60 min. The precision of the method, expressed as RSD, was 8.8 % at 100.0 microg L(-1). The method was applied, and validated chemometrically, for the determination of nitrite in different types of water (spring, mineral, tap, well, and sea). PMID- 12110982 TI - Development of an FIA system with amperometric detection for determination of bentazone in estuarine waters. AB - On the basis of its electrochemical behaviour a new flow-injection analysis (FIA) method with amperometric detection has been developed for quantification of the herbicide bentazone (BTZ) in estuarine waters. Standard solutions and samples (200 microL) were injected into a water carrier stream and both pH and ionic strength were automatically adjusted inside the manifold. Optimization of critical FIA conditions indicated that the best analytical results were obtained at an oxidation potential of 1.10 V, pH 4.5, and an overall flow-rate of 2.4 mL min(-1). Analysis of real samples was performed by means of calibration curves over the concentration range 2.5x10(-6) to 5.0x10(-5) mol L(-1), and results were compared with those obtained by use of an independent method (HPLC). The accuracy of the amperometric determinations was ascertained; errors relative to the comparison method were below 4% and sampling rates were approximately 100 samples h(-1). The repeatability of the proposed method was calculated by assessing the relative standard deviation (%) of ten consecutive determinations of one sample; the value obtained was 2.1%. PMID- 12110983 TI - Diformylhydrazine as analytical reagent for spectrophotometric determination of iron(II) and iron(III). AB - The bidentate ligand diformylhydrazine (OHC-HN-NH-CHO), DFH, combines with iron(II) and iron(III) in alkaline media in the pH range 7.3-9.3 to form an intensely colored red-purple iron(III) complex with an absorption maximum at 470 nm. Beer's law is obeyed for iron concentrations from 0.25 to 13 microg mL(-1). The molar absorptivity was in the range 0.3258x10(4)-0.3351x10(4) L mol(-1) cm( 1) and Sandell's sensitivity was found to be 0.0168 microg cm(-2). The method has been applied to the determination of iron in industrial waste, ground water, and pharmaceutical samples. PMID- 12110984 TI - Total synchronous fluorescence scan spectra of petroleum products. AB - Extending the two-dimensional synchronous fluorescence scan to a three dimensional total synchronous fluorescence scan (TSFS) spectral measurement gives the total synchronous fluorescence characteristics of a multifluorophoric sample at various possible wavelength intervals (Deltalambda), which could help to characterize multifluorophoric systems better. TSFS spectra of petroleum products such as diesel, kerosene, petrol, engine oil etc., available in the Indian market, are reported. Fluorescence in these samples is due to the presence of polycyclic aromatic hydrocarbons (PAHs) of various ring sizes. The TSFS contour plot profiles of the neat samples measured at right-angle geometry is a result of various energy-degrading photophysical processes such as inner filter effect, light attenuation, resonance energy transfer, collisional quenching etc. TSFS plots make it easy to obtain the optimized Deltalambda of an unknown sample of analytical interest. TSFS and the excitation-emission matrix (EEM) techniques are similar, but the contour profiles generated are different. The response of the TSFS contour profiles to dilution is different from that in the EEM contour profiles. Thus, TSFS can provide an alternative way of presenting the fluorescence response of concentrated multifluorophoric samples. PMID- 12110985 TI - ETAAS determination of nickel in serum and urine. AB - Methods for the direct determination of Ni in human blood serum and urine by electrothermal atomic absorption spectrometry (ETAAS) are described. Hydrogen peroxide was proposed as matrix modifier, assisting thermal decomposition of proteins during the ashing step. A pyrolysis temperature of 1,200 degrees C was found to be optimal while 2,100 degrees C and 2,200 degrees C were found to be optimal atomizing temperatures for Ni in serum and urine respectively. Calibration was performed by using a calibration curve prepared with aqueous standard solutions of Ni (glycine must be used as modifier for Ni in aqueous solutions). The limits of detection, defined as the blank values plus 3 times the standard deviation of the blank values, were 0.2 microg/L for both serum and urine samples. Relative standard deviations for serum samples with concentrations of Ni in the range 0.5-2 microg/L were 10-15% and for urine samples with Ni concentrations in the range 0.5-2.5 microg/L were 8-10%. PMID- 12110986 TI - Influence of soluble polymer polyvinylpyrrolidone on separation of small peptides and amino acids by microchip-based capillary electrophoresis. AB - In microchip-based capillary electrophoresis, the resolution and separation efficiency of small peptides and amino acids can be noticeably improved by adding a low molecular weight (30,000) soluble polymer additive, polyvinypyrrolidone in the separation medium. Several separation conditions such as injection time and electrophoretic buffer have been investigated and optimized. Using an electro stacking scheme, the resolution and separation efficiency of small peptides and amino acids can be enhanced significantly. Under the optimal conditions, the separation of fluorescein isothiocyanate Isomer I-labeled small peptides and amino acids was successfully achieved within 100 s. PMID- 12110987 TI - Effects of typical and atypical antipsychotics on prepulse inhibition in schizophrenia: a critical evaluation of current evidence and directions for future research. AB - Prepulse inhibition (PPI) of the startle response refers to an attenuation in response to a strong stimulus (pulse) if this is preceded shortly by a weak non startling stimulus (prepulse). PPI provides a simple operational measure of sensorimotor gating, serving to prevent the interruption of ongoing perceptual and early sensory analysis. In accord with postulated deficits in early stages of information processing, there is ample evidence that PPI is disrupted in individuals with schizophrenia. PPI in animals is thought to represent a well validated model for evaluating potential new treatments for schizophrenia. Currently, available data on the differential effects of typical and atypical antipsychotics suggest that atypical antipsychotics, in particular clozapine and risperidone, may be more effective than typical antipsychotics in improving PPI deficits in schizophrenia. However, studies have so far used small samples and/or between-subjects designs, and not examined the effects of other concomitant medications that may also influence PPI. The directions are identified for further applications of this model using within-subjects longitudinal designs and reasonable sample sizes to establish superiority of particular atypical antipsychotics over typical antipsychotics in improving PPI in schizophrenic populations. PMID- 12110988 TI - Nicotine addiction: insights from recent animal studies. AB - RATIONALE: Recent preclinical behavioral and neurobiological research has characterized important behavioral features and has identified neurobiological substrates that may underlie nicotine reinforcement and addiction. OBJECTIVE: To examine recent advances on nicotine exposure in preclinical models, from three perspectives: (a) the chronopharmacokinetics of nicotine, (b) behavioral studies on nicotine reinforcement, withdrawal, and reinstatement/relapse, and (c) effects of nicotine on neurobiological substrates after repeated exposure. RESULTS: Preclinical studies can be used to operationally model selected aspects of nicotine reinforcement, withdrawal, and reinstatement or relapse. These may be used to investigate the functional in vivo consequences of acute and long-term changes in neuronal acetylcholine receptor populations that follow nicotine exposure. Behavioral studies focusing on distinct stages of nicotine exposure (e.g., active reinforcement vs. cessation or reinstatement) may also be used in parallel with studies on dopaminergic function, a proposed substrate for the reinforcing effects of nicotine, and of opioid receptor function, a possible site of neuroadaptations secondary to nicotine exposure. CONCLUSIONS: While no single current animal model may capture the experience of human smoking or nicotine addiction, increasingly, separate animal models are capturing the full spectrum of behavioral and neurobiological dimensions of this complex condition. PMID- 12110989 TI - Oxygen saturation and cognitive performance. AB - The purpose of the experiments was to investigate how inhalation of 100% oxygen affected cognitive performance. A test battery was developed that was designed to capture different aspects of cognitive processes, i.e., perception, attention, working memory, long-term memory and prospective memory. All tests were verbally based, thus reducing cognitive spatial processes to a minimum. In experiment 1, 48 participants volunteered in a complete factorial within-participant design. Two different conditions for type of gas were used, inhalation of 100% oxygen and inhalation of breathing air (approximately 21% oxygen balanced with nitrogen). The inhalation was performed during the 1 min prior to starting each separate test. The instructions for each test were given during the inhalation period. All participants inhaled oxygen or breathing air through a Swedish military pilot mask. Physiological (heartbeats per minute and blood oxygen saturation level) reactions were recorded continuously throughout the session. Participants also completed a mood-state questionnaire before and after the test battery. The results revealed that cognitive performance were not affected by inhalation. Hence, this experiment does not replicate previous findings that suggest that inhalation of 100% oxygen could increase cognitive performance. Another experiment was performed to control for methodological issues. Experiment 2 revealed exactly the same pattern, i.e., inhalation of oxygen did not affect cognitive functioning. PMID- 12110990 TI - Nicotine-induced enhancement of attention in the five-choice serial reaction time task: the influence of task demands. AB - RATIONALE: Beneficial effects of nicotine on cognitive processes including attention have potential therapeutic uses and have been proposed as incentives for tobacco smoking. OBJECTIVES: To establish task conditions under which the effects of nicotine on attention are obtained reliably and to characterise such effects further. METHODS: Rats were trained in a modified version of the five choice serial reaction time task (5-CSRTT) to detect 1-s light stimuli with greater than 70% accuracy and fewer than 20% omission errors. Nicotine was tested under different task requirements by varying signal event rate, stimulus duration and stimulus predictability, and by introducing white-noise distractors. RESULTS: Nicotine (0.05-0.2 mg/kg, s.c.) repeatedly improved accuracy and reduced omission errors and reaction times, leading to increases in numbers of reinforcers earned. Anticipatory responding was increased. Parametric modifications intended to increase demands on sustained attention did not affect performance in a manner suggesting that this subtype of attention was being taxed, and the effects of nicotine were not more marked under such conditions. Shorter stimulus durations impaired performance, but this manipulation weakened the effect of nicotine on accuracy. In contrast, the presence of noise distractors facilitated the effects of nicotine to the extent that distractor-induced impairments were abolished by the drug. CONCLUSIONS: The 5-CSRTT can provide a sensitive rodent model for the attention-enhancing effects of nicotine. Changes made to the procedure may have increased its sensitivity to nicotine, particularly with respect to accuracy. There were indications that the effects of nicotine were largest on processes of selective attention or on disengaging attention from irrelevant events and shifting it to behaviourally significant stimuli. PMID- 12110991 TI - Sensorimotor gating effects produced by repeated dopamine agonists in a paradigm favoring environmental conditioning. AB - RATIONALE: It is not known whether dopamine agonist-induced disruption of prepulse inhibition (PPI) can be conditioned to the environment, a phenomenon established for dopamine agonist-induced locomotor activation and other behaviors. Furthermore, the literature is contradictory regarding whether PPI disruption, like locomotor activity, can become sensitized after repeated dopamine agonist administration. Differences in methodology (e.g. drug environment pairing) may have contributed to these contradictory findings. OBJECTIVES: In a series of studies, we investigated whether dopamine agonist induced disruption of PPI could be conditioned and whether repeated administration of dopamine agonists, in a paradigm favorable to conditioning, would produce sensitization to dopamine agonist-induced disruption of PPI. METHODS: One group of rats were administered subcutaneous apomorphine (0.5 mg/kg) daily for 7 (experiment 1) or 5 (experiment 3) consecutive days contingent with startle testing (in testing rooms, immediately before test sessions). A second group received the same apomorphine dose daily in a manner non-contingent with behavioral testing (in home cages after test sessions). The following day, all rats received vehicle injections contingent with the test environment to assess for environmental conditioning (vehicle challenge day). The next day, all animals received a challenge of apomorphine (0.5 mg/kg) contingent with the test environment to assess the contribution of drug-environment pairing on changes observed in apomorphine-induced disruption of PPI (apmorphine challenge day). PPI was measured immediately after drug injections in the test environment. A separate study (experiment 2) tested amphetamine (3.0 mg/kg) using a similar methodology. In a fourth study, rats were pretreated with haloperidol (1.0 mg/kg) or saline prior to receiving daily apomorphine to see if haloperidol could modify the changes in PPI produced by repeated apomorphine administration. RESULTS: On the vehicle challenge day, PPI exhibited by the rats that received daily apomorphine contingent with the testing environment did not differ from the group that received vehicle contingent with the testing environment. However, animals receiving apomorphine contingent with testing exhibited partial tolerance to its PPI effects during the conditioning period. The PPI exhibited by both groups did not differ significantly on the apomorphine-challenge day. Amphetamine produced a complete tolerance to its PPI effects by day 3. Haloperidol pretreatment blocked the PPI tolerance produced by repeated apomorphine injections. CONCLUSIONS: These results suggest: 1) unlike locomotion, PPI disruption induced by apomorphine cannot be conditioned to the environment; 2) unlike locomotion, repeated adminstration of dopamine agonists produce tolerance, rather than sensitization, to PPI; 3) environmental factors do not seem to be critical for PPI tolerance; and 4) dopamine receptors mediate PPI tolerance to apomorphine. PMID- 12110993 TI - Differential modulation of efficiency in a food-rewarded "differential reinforcement of low-rate" 72-s schedule in rats by norepinephrine and serotonin reuptake inhibitors. AB - RATIONALE: The differential reinforcement of low-rate 72-s (DRL 72-s) schedule, in which rats must withhold a response for at least 72 s to obtain a reward (generally water), is an attractive procedure for the characterisation of potential antidepressant agents. Indeed, several antidepressants have been shown to improve efficiency (ratio of reinforcement rate to response rate) in this model, either by decreasing response rates and/or by increasing reinforcement rates. OBJECTIVE: Herein, we compared the actions of antidepressants known to inhibit serotonin (5-HT), norepinephrine (NE) and/or dopamine (DA) reuptake in a food-rewarded DRL 72-s schedule. METHODS: Rats trained in a food-rewarded DRL 72 s schedule and showing stable baseline performance were administered with drugs i.p. once a week. In independent experiments, the influence of drugs on food intake, spontaneous locomotor activity and extracellular levels of monoamines in the frontal cortex was evaluated. RESULTS: In confirmation of previous studies, the tricyclic agent imipramine (10.0 mg/kg) and the "atypical" agent mianserin (40.0 mg/kg) significantly increased efficiency. In analogy, the selective NE reuptake inhibitors (NARIs) desipramine (20.0 mg/kg), nortriptyline (2.5 mg/kg) and reboxetine (0.63 mg/kg) all displayed marked enhancements in efficiency. In contrast, the selective 5-HT reuptake inhibitors (SSRIs) citalopram (10.0 mg/kg), fluvoxamine (10.0 mg/kg) and paroxetine (10.0 mg/kg) all significantly decreased efficiency. The mixed 5-HT/NE reuptake inhibitors (SNRIs) venlafaxine (2.5 mg/kg, 10.0 mg/kg) and S33005 (0.16-10.0 mg/kg), likewise, did not increase efficiency. Further, the DA reuptake inhibitors (DARIs) bupropion (0.16-10.0 mg/kg) and GBR12935 (0.63-10.0 mg/kg) had no effect on DRL 72-s performance. All drug classes exerted a similar, mild inhibitory influence on food intake and locomotor behaviour. Imipramine, mianserin and NARIs markedly increased extracellular levels of NE, and SSRIs elevated levels of 5-HT, while SSRIs augmented levels of both. CONCLUSIONS: The present experimental procedure demonstrates, in analogy to imipramine and mianserin, robust and consistent increases in efficiency with NARIs. Their effects may be distinguished from a decrease in efficiency elicited by SSRIs, and a lack of activity of SNRIs and DARIs. While the reasons underlying the ineffectiveness of SSRIs (in contrast to previous studies) remain to be clarified, these data underline the importance of adrenergic mechanisms in the control of behaviour under conditions of delayed responding. Further, they support the interest of DRL 72-s procedures for the characterisation of diverse classes of antidepressant agent. PMID- 12110992 TI - Influence of forced swimming-induced stress on the anxiolytic-like effect of 5HT(1A) agents in mice. AB - RATIONALE: Stress has been related to both anxiety and mood disorders. Forced swimming (FS) is a type of stress that is able to modify the activity of serotonin (5-HT) and GABA in the central nervous system. 5-HT(1A) compounds have been shown to be anxiolytic in a variety of behavioral models and in clinical studies. OBJECTIVE: The main purpose of the present study was to analyze the effect of FS on the anxiolytic-like actions of three 5-HT(1A) compounds. METHODS: Stressed (ST) and unstressed (UST) mice were evaluated in the exploratory behavior test (EBT) or burying behavior test (BBT). In addition, the action of increasing doses of the 5-HT(1A) compounds buspirone, 8-OH-DPAT and indorenate in ST and UST mice was analyzed using the EBT. A spontaneous ambulatory behavior test was carried out immediately after the anxiety tests. RESULTS: One session of FS induced anxiolytic-like behavior in mice tested in both the EBT and the BBT. This effect of FS was blocked by a previous administration of either picrotoxin or WAY 100635. The 5-HT(1A) compounds produced a clear anxiolytic-like effect in UST animals. By contrast, with low doses of either 8-OH-DPAT (0.01 mg/kg), buspirone (0.03 mg/kg) or indorenate (0.3, 0.6 mg/kg) ST mice showed a decrease in the anti-anxiety-like effect observed after FS. No change in ambulation that could mask the results of the anxiety test was registered. CONCLUSIONS: The present data provide evidence that FS induces changes in the effect of 5-HT(1A) agents. The participation of the 5-HT and/or GABA systems in these stress-induced effects is discussed. PMID- 12110994 TI - Stimulation by antipsychotic agents of mitogen-activated protein kinase (MAPK) coupled to cloned, human (h)serotonin (5-HT)(1A) receptors. AB - RATIONALE: There is evidence that serotonergic mechanisms contribute to the functional profiles of antipsychotic drugs, several of which display affinity for human (h)5-HT(1A) receptors. OBJECTIVE: Here, we compared the interaction of several antipsychotic agents at h5-HT(1A) receptors employing mitogen-activated protein kinase (MAPK), an intracellular marker. METHODS: The influence of antipsychotics on MAPK phosphorylation was quantified in Chinese hamster ovary (CHO) cells stably transfected with h5-HT(1A) receptors by use of a highly selective antibody. RESULTS: The novel antipsychotic agent, S16924, concentration dependently (pEC(50), 8.10) stimulated the phosphorylation of MAPK. Its maximal effect (96%) was similar to that of the prototypical 5-HT(1A) agonist, (+)8-OH DPAT (pEC(50), 8.54) (defined as 100%). The selective 5-HT(1A) receptor antagonist WAY100,635, which was inactive alone, abolished stimulation of MAPK by S16924 with a pK(b) of 9.66. This stimulatory influence of S16924 on MAPK was potently mimicked by the benzoisoxazole, antipsychotic ziprasidone (pEC(50), 7.25; 93%). The atypical antipsychotic clozapine also activated MAPK, albeit with lower potency and efficacy (pEC(50), 5.43 and 43%). These actions of ziprasidone and clozapine were also blocked by WAY100,635. Evaluated at a single, high concentration, several other antipsychotics stimulated MAPK phosphorylation with variable efficacy: quetiapine (75%), ocaperidone (74%), tiospirone (57%), olanzapine (54%) and risperidone (21%). In all cases, their actions were abolished by WAY100,635. In contrast, haloperidol, thioridazine and sertindole did not stimulate MAPK. CONCLUSIONS: Antipsychotics display contrasting efficacies in modulating MAPK phosphorylation at h5-HT(1A) receptors, ranging from high (e.g. S16924 and ziprasidone), via intermediate (e.g. clozapine) to low (e.g. haloperidol). Differential modulation of 5-HT(1A) receptor-coupled MAPK may contribute to their contrasting functional profiles. PMID- 12110996 TI - Physiological and performance effects of pyridostigmine bromide in healthy volunteers: a dose-response study. AB - RATIONALE: Questions have been raised about the role pyridostigmine bromide (PB) plays in the etiology of Gulf War veterans' illnesses. There is a need to understand better the physiological and behavioral effects of this drug, particularly at the 30-mg/8-h regimen recommended by the US Military. OBJECTIVE. To perform a double-blind, cross-over, dose-response study of PB in 67 healthy, young volunteers (31 women, 36 men). METHODS: Volunteers were initially trained on a standardized test battery. Supervised administration of placebo (PL) and PB (every 8 h/5 days) occurred in each of two dosing weeks, separated by a non dosing week. One group received 30 mg PB and PL, and the other 60 mg PB and PL. In each dosing week, the battery was performed after the first pill and again when steady-state plasma PB levels were achieved. RESULTS: PB was associated with an overall improvement in reaction time on tests of memory and attention, and with a reduction in RMS error on a tracking task. PB slowed heart rate and decreased the high frequency component of heart rate variability (HF HRV). Dose response effects were found only for HF HRV, and RMS error. The extent of cholinesterase inhibition was directly related to the magnitude of the HF HRV decrease, and was predicted by the weight-normalized PB dose. Cholinesterase inhibition was not related to the extent or severity of reported drug side effects. CONCLUSIONS: PB does not appear to have detrimental physiological or performance consequences at the recommended 30-mg dose, or at twice that dose, when evaluated under non-stressful laboratory conditions. PMID- 12110995 TI - Enhancement of cocaine-seeking behavior by repeated nicotine exposure in rats. AB - RATIONALE: Drugs with addictive liability have a high probability of co-abuse in many addicts. For example, cocaine users are several times more likely to smoke cigarettes than non-cocaine users, and smoking increases during cocaine use. Previous work has provided evidence that nicotine and cocaine have interactive neurochemical effects, particularly with regard to dopamine (DA) transmission. OBJECTIVES: The present study examined the impact of nicotine treatment on the reinforcement efficacy of self-administered cocaine and non-reinforced responding for cocaine in rats. METHODS: Rats were trained to self-administer cocaine (i.v.) on a progressive ratio (PR) schedule of reinforcement. Self-administration training continued until stable responding was obtained. Acute nicotine pretreatment consisted of a subcutaneous injection (0.15, 0.3 and 0.6 mg/kg) 3 min prior to cocaine access. In the repeated treatment condition, a separate group of animals was given nicotine (0.6 mg/kg, s.c.) 3 min prior to cocaine access for 14 consecutive days. During extinction trials, these animals were injected with nicotine (0.6 mg/kg, s.c.) after 45 min of non-reinforced responding. RESULTS: Acute nicotine treatment produced an inverted U-shaped dose response function with lower doses increasing and the highest dose decreasing the number of cocaine infusions obtained during a session. Animals treated repeatedly with the highest dose of nicotine showed a significant increase in the number of cocaine infusions by day 8 of nicotine treatment. During extinction sessions when cocaine was not available, injections of nicotine in these animals caused a reinstatement of the previously rewarded lever-press behavior. CONCLUSIONS: These findings indicate that nicotine can facilitate cocaine reinforcement, may contribute to the transition from moderate drug-taking to an escalation of drug intake which is characteristic of addiction, and may trigger relapse. PMID- 12110997 TI - In vitro receptor screening of pure constituents of St. John's wort reveals novel interactions with a number of GPCRs. AB - RATIONALE: Hypericum perforatum L. (St. John's wort; SJW) is one of the leading psychotherapeutic phytomedicines and great effort has been devoted to clarifying its mechanism of action. OBJECTIVE: We have undertaken a comprehensive analysis of several pure compounds isolated from the crude extract to gain further insight into the molecular actions of various substituents of SJW. METHODS: We characterized the in vitro pharmacology of the naphthodianthrones hypericin and pseudohypericin, the phloroglucinol derivative hyperforin, and several flavonoids at 42 biogenic amine receptors and transporters using the resources of the National Institute of Mental Health Psychoactive Drug Screening Program. RESULTS: The biflavonoid amentoflavone significantly inhibited binding at serotonin (5 HT(1D), 5-HT(2C)), D(3)-dopamine, delta-opiate, and benzodiazepine receptors. The naphthodianthrone hypericin had significant activity at D(3)- and D(4)-dopamine receptors and beta-adrenergic receptors. With the exception of the D(1)-dopamine receptor, the phloroglucinol derivative hyperforin was less active than other SJW constituents tested on all screened receptors. CONCLUSION: Our present in vitro data clearly show that several pure substances in SJW are potential CNS psychoactive agents and may contribute to the antidepressant efficacy of the plant in a complex manner. Our data also reveal novel and heretofore unexpected interactions of pure compounds in SJW at a number of GPCRs, transporters, and ion channels. We hypothesize that additive or synergistic actions of different single compounds may be responsible for the antidepressant efficacy of SJW. These results and this general approach may impact our understanding of phytomedicines in general and H. perforatum specifically. PMID- 12110998 TI - Neuropsychological evidence of a relatively selective profile of temporal dysfunction in drug-free MDMA ("ecstasy") polydrug users. AB - RATIONALE: Experimental evidence has shown that 3,4-methylenedioxymethamphetamine (MDMA, "ecstasy") can act as a serotonergic neurotoxin in laboratory animals. The serotonin system predominantly innervates frontal and limbic regions of the brain and has been associated with consolidatory learning and mnemonic processes in humans. OBJECTIVES: The aim of the present study was to investigate the cognitive neuropsychological profile of drug-free ecstasy users by employing a selection of tasks previously associated with lesion or neurodegenerative damage to the temporal lobe or fronto-striatal regions. METHODS: The study comprised 40 participants: 20 ecstasy polydrug users and 20 polydrug users who had never taken ecstasy. RESULTS: Ecstasy users were significantly impaired on a recognition task for complex visual patterns and spatial working memory, as a function of task difficulty rather than systematic search strategy. They also showed a trend towards impairment on several learning paradigms. Ecstasy users remained relatively unimpaired on most measures associated with prefrontal functioning, with the exception of verbal fluency "letter" generation. CONCLUSIONS: Initial cognitive deficits in ecstasy polydrug users may be more apparent in tasks known to be sensitive to temporal functioning. PMID- 12110999 TI - Methodological problems with ecstasy and the SCL-90. PMID- 12111001 TI - Difference between kidney and liver in decreased manganese superoxide dismutase activity caused by exposure of mice to mercuric chloride. AB - Dysfunction of antioxidant enzymes caused by mercuric compounds is partially associated with substantial induction of oxidative stress. In the present study, changes in renal and hepatic enzyme activity of an antioxidant protein manganese superoxide dismutase (Mn-SOD) after exposure to mercuric chloride (HgCl(2)) were examined in ICR mice. Subcutaneous administration of HgCl(2) (0.25-3 mg/kg) resulted in a decrease in renal Mn-SOD activity in a dose-dependent manner, whereas the hepatic enzyme activity was unaffected following injection of HgCl(2). Mercury accumulation in the kidney was drastically higher (34-75 times) than that in the liver after HgCl(2) administration. Examining interactions of purified Mn-SOD with HgCl(2) indicated that mercury ions suppressed Mn-SOD activity by reduction of the native form. These results suggest that inorganic mercury can directly interact with murine Mn-SOD, resulting in decrease of the enzyme activity and that the HgCl(2)-mediated significant reduction of renal, but not hepatic, Mn-SOD activity in vivo appears to be associated with the tissue specificity for mercury accumulation. PMID- 12111002 TI - Sharp dose- and time-dependent toxicity of mercuric chloride at the cellular level in sea urchin embryos. AB - Sea urchin embryos ( Sphaerechinus granularis) offer the opportunity to analyse toxicity towards cell division and stages of early development. Mercuric chloride (HgCl(2)) arrested early development at the level of the first cell cycle. The toxic effect occurred in a very sharp concentration range around 7 microM HgCl(2). At sub-toxic concentrations of HgCl(2), the morphology and kinetics of early development were comparable to control embryos. The time-dependence of toxicity was short; a 5-min exposure to the toxic concentration of 10 microM HgCl(2) was sufficient to provoke developmental dysfunction whereas continuous exposure to 5 microM HgCl(2) allowed development to occur normally. The effects on early development over this range of concentrations were specific to HgCl(2) toxicity since other heavy metal chlorides had no effect at 30 microM. Thus, the sea urchin model may provide new clues to the molecular mechanisms of HgCl(2) toxicity. PMID- 12111003 TI - Neonatal exposure of newborn mice to pyrethroid (permethrin) represses activity dependent c-fos mRNA expression in cerebellum. AB - In a previous report, we demonstrated that the exposure of cultured mouse cerebellar granule cells to permethrin, a type I pyrethroid insecticide, repressed the induction of activity-dependent c- fos and brain-derived neurotrophic factor (BDNF) gene expression, accompanying a decrease in Ca(2+) influx into neurons. In addition, it has been suggested that some pyrethroids, including permethrin, are endocrine-modulating chemicals and accumulate in human breast milk. In this study, therefore, we investigated whether lactational exposure of newborn mice to permethrin influenced c- fos, BDNF and beta-actin gene expression in the developing neonatal cerebellum. In the cerebella of control neonates, c- fos mRNA expression was characterized by a significant increase in postnatal weeks 2 and 3, followed by a marked decrease. In the cerebella of permethrin-treated neonates, the expression of c- fos mRNA was dose dependently repressed by cis-permethrin more effectively than by trans-permethrin at postnatal week 3, without alterations in the body or cerebellum weights of neonates. In the fourth and fifth week, however, c- fos mRNA expression had decreased to the same level as that in the control and permethrin-treated neonates. A decrease in BDNF mRNA expression tended to be observed in the cerebella of newborn mice on exposure to permethrin. Thus, our results indicate that the activity-dependent gene expressions in cerebellar neuronal cells can be repressed by permethrin both in vitro and in vivo, and suggest that lactational exposure to pyrethroids might affect the postnatal development of the mammalian brain. PMID- 12111004 TI - Altered gene expression of hepatic lanosterol 14alpha-demethylase (CYP51) in lead nitrate-treated rats. AB - Effects of lead nitrate (LN), a hepatic mitogen, on hepatic gene expressions of lanosterol 14alpha-demethylase (CYP51) and the sterol regulatory element binding proteins (SREBP-1a, SREBP-1c and SREBP-2), which are thought to be transcription factors for hepatic CYP51 gene, were examined by the methods of Northern blot and/or real time reverse transcriptase-polymerase chain reaction (RT-PCR). In both immature (4-week-old) and mature (7-week-old) rats, LN treatment resulted in definite increases in hepatic gene expression of CYP51 at 12 h and in the liver weight at 48 h. As for transcription factors for the CYP51 gene, enhanced gene expression of SREBP-2 was observed 6-12 h after LN treatment, whereas no enhanced gene expression of other SREBPs, SREBP-1a and SREBP-1c, was observed at any time after the treatment; for SREBP-1a, there was no significant change; for SREPB-1c, there was a drastic decrease. In addition, the serum total cholesterol level was increased 12 h after LN treatment to 7-week-old rats, and the increased level was maintained at least up to 48 h later. In the present study, we demonstrate for the first time that LN, a heavy-metal ion, activates the expression of the SREBP 2 and CYP51 genes without decreasing the serum total cholesterol level and further suggest that only SREBP-2 among SREBPs might play an important role in the LN-enhanced CYP51 gene expression. PMID- 12111005 TI - 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) induces plasminogen activator inhibitor-1 through an aryl hydrocarbon receptor-mediated pathway in mouse hepatoma cell lines. AB - 2,3,7,8-Tetrachlorodibenzo- p-dioxin (TCDD), a ubiquitous environmental pollutant, elicits a variety of toxicities and is a well-known carcinogen. TCDD alters the expression of many genes including CYP1A1/2, CYP1B1, glutathione S transferase Ya, aldehyde-3-dehydrogenase, NAD(P)H:quinone oxidoreductase, transforming growth factor (TGF)-alpha and TGF-beta. The present study was aimed at characterization of TCDD to induce plasminogen activator inhibitor-1 (PAI-1) in mouse hepatoma cell lines. A Hepa1c1c7 wild-type cell [H1(wt)], an aryl hydrocarbon receptor (AhR)-deficient mutant [H1(AhR(-))] and an AhR nuclear translocator (Arnt)-deficient mutant [H1(Arnt(-))] were used for this study. TCDD induced PAI-1 in H1(wt) cells, but not in H1(AhR(-)) and H1(Arnt(-)) mutants, indicating a functional role of the AhR-Arnt complex in this effect. Cycloheximide (CHX) treatment resulted in increased PAI-1 mRNA induction, indicating that this response to TCDD is a direct effect on transcription and not a secondary effect mediated by other TCDD-induced proteins. Transfection with PAI 1 promoter led to increased PAI-1 promoter activity in H1(wt) cells treated with TCDD, but no such effect occurred in H1(AhR(-)) or H1(Arnt(-)) cells, implying involvement of the AhR and Arnt. In addition, alpha-naphthoflavone and phenanthroline, two AhR antagonists, each blocked the enhancing effect of TCDD on PAI-1 promoter-coupled luciferase activity in H1(wt) cells. PAI-1 promoter deletion analysis indicated that TCDD-induced PAI-1 transcription was distinctly different from TGF-beta-dependent PAI-1 transcription, particularly in the region between -161 to +73. In summary, TCDD induced the PAI-1 gene directly via an AhR- and Arnt-dependent mechanism, which was distinctly different from TGF-beta-driven PAI-1 transcription. PMID- 12111007 TI - Effects of butyl paraben on the male reproductive system in mice. AB - Parabens are alkyl ester compounds of p-hydroxybenzoic acid widely used as preservatives in foodstuffs, cosmetics, toiletries and pharmaceuticals. These compounds are known to exert a weak estrogenic activity in estrogen receptor assays in vitro and in uterotrophic assays in vivo. In this paper, we have shown that butyl paraben had an adverse effect on the male mouse reproductive system and that it damaged the late steps of spermatogenesis in the testis. Butyl paraben was administered to 4-week-old Crj:CD-1 mice assigned to groups of eight animals, at doses of 0.01%, 0.10%, and 1.00% in the diet for 10 weeks. The average butyl paraben intake from the calculated food consumption was 14.4+/ 3.60, 146+/-35.9, and 1504+/-357 mg/kg per day for the 0.01%, 0.10%, and 1.00% dietary butyl paraben groups, respectively. There were no treatment-related effects of butyl paraben on the liver, ventral prostates, seminal vesicles, and preputial glands (both in terms of absolute weight and relative to body weight) in any of the study groups. Both the absolute and relative weights of the epididymides were significantly higher in 1.00% group when compared with controls. A dose-dependent decrease of both round and elongated spermatid counts in stages VII-VIII seminiferous tubules was observed, and the elongated spermatid counts were significantly lower in all of the treated groups. The numbers of spermatogonia and spermatocytes did not differ from control values. The serum testosterone concentration decreased in a dose-dependent fashion and was significant at 1.00%. These data demonstrated that butyl paraben can exert an adverse effect on the male reproductive system at doses that are well below those of the accepted daily intake (ADI) in Japan. PMID- 12111006 TI - Effects of the dithiocarbamate fungicide propineb in primary neuronal cell cultures and skeletal muscle cells of the rat. AB - After repeated-dose toxicity studies with the fungicide propineb, reversible effects on muscle functions were found. Therefore, mechanistic investigations should contribute to clarification of its mode of action in relation to disulfiram and diethyldithiocarbamate neurotoxicity or direct effects on muscle cells. In principle, besides the dithiocarbamate effects, two different mechanisms have been discussed for this fungicide. One mechanism is the degradation to carbon disulfide (CS(2)) and propylenthiourea (PTU) and the other are direct effects of zinc. Primary neuronal cell cultures of the rat are a well established model to identify neurotoxic compounds like n-hexane or acrylamide. In this cell culture model, endpoints such as viability, energy supply, glucose consumption and cytoskeleton elements were determined. Additionally, skeletal muscle cells were used for comparison. Propineb and its metabolite PTU were investigated in comparison to CS(2), disulfiram and diethyldithiocarbamate. The toxicity of zinc was tested using zinc chloride (ZnCl(2)). It was clearly shown that propineb exerted strong effects on the cytoskeleton of neuronal and non neuronal cell cultures (astrocytes, muscle cells). This was similar to ZnCl(2,) but not to CS(2). With CS(2) and disulfiram effects on the energy supply were more prominent. In conclusion, the toxicity of propineb is not comparable to disulfiram, diethyldithiocarbamate or CS(2) neurotoxicity. In regard to these findings, a direct reversible effect of propineb on skeletal muscle cells seems to be more likely. PMID- 12111008 TI - Assessment of DNA damage in workers occupationally exposed to pesticide mixtures by the alkaline comet assay. AB - The potential genetic hazard of pesticides to human beings is of great concern in occupational and environmental settings because of the widespread use of these chemicals for domestic and industrial applications. Various studies have revealed a significantly elevated risk for particular tumours in humans exposed to some pesticides. Results from the biological monitoring or cytogenetic methods for the detection of health risks to pesticides have given both positive and negative results of mutagenicity. In this study DNA damage in peripheral lymphocytes of 33 pesticide-exposed workers employed in the municipality of Ankara (Turkey) for at least 1 year was examined by alkaline single-cell gel electrophoresis, the 'comet' technique. Results were compared with those from 33 controls of comparable age, sex and smoking habits, which were not occupationally exposed to pesticides. Work characteristics of the exposed workers and the use of personnel protective measures were also investigated. The DNA damage observed in lymphocytes of the workers was significantly higher than that in the controls ( P<0.001). The observed DNA damage was found to be significantly lower ( P<0.001) in workers applying some of the necessary individual safety protections during their work. Cigarette smoking was not related to increases in DNA damage; also, no significant association was found between the duration of occupational exposure to pesticides and the degree of DNA damage. PMID- 12111011 TI - Biochemical indices of bone turnover and the assessment of fracture probability. PMID- 12111012 TI - A new approach to the development of assessment guidelines for osteoporosis. PMID- 12111013 TI - Relation of statin use and bone loss: a prospective population-based cohort study in early postmenopausal women. AB - Recent experimental and epidemiologic studies have suggested that the lipid lowering drugs, statins, may have bone-protective effects. We studied the effects of statin use on the change in bone mineral density (BMD) in a prospective 4.5 year cohort study based on subjects from the Kuopio Osteoporosis Risk Factor and Prevention (OSTPRE) Study, Finland. Six hundred and twenty women aged 53-64 years were divided into four groups: 55 women reported continuous and 63 women occasional statin use during the follow-up; 142 non-users of statins reported hypercholesterolemia whereas 360 non-users did not. Spinal and femoral BMDs were measured by dual-energy X-ray densitometry in 1995-1996 and 1999-2000 and the BMD changes of the four groups were compared. Characteristics of the study population were obtained with postal inquiries. The mean annual spinal and femoral BMD changes of the study groups were 0.29% and -0.50% for the continuous statin users, 0.19% and -0.57% for the occasional statin users, 0.52% and -0.29% for the hypercholesterolemic non-users of statins, and 0.39% and -0.33% for the non-users of statins without hypercholesterolemia, ( p = 0.398 and p = 0.404) respectively. The corresponding BMD changes adjusted for age, years since menopause, body mass index, BMD at baseline, calcium intake, estrogen and cortisone therapy, duration of follow-up and statin use before the baseline were -0.20% and -0.47%, 0.19% and -0.54%, 0.54% and -0.32%, 0.47% and -0.33% ( p = 0.134 and p = 0.628), respectively. Our results suggest that statins do not protect from early postmenopausal bone loss. Randomized trials are needed to confirm these results. PMID- 12111014 TI - Hip fracture risk and proximal femur geometry from DXA scans. AB - In this retrospective study of hip fracture risk evaluation from hip dual-energy X-ray absorptiometry (DXA) scans, our objectives were to determine which part of the femoral neck length contributes most to the fracture risk and to define a geometric parameter better than hip axis length (HAL) for discriminating hip fracture patients. Forty-nine Caucasian women with a nontraumatic femoral neck fracture were matched on age to 49 normal women and on both age and femoral neck bone mineral density (BMD) to 49 unfractured women. In addition to BMD, geometric parameters including neck-shaft angle, neck width and several HAL segments were evaluated by discriminant analysis to determine which was the best hip fracture discriminator. Neck-shaft angle had a limited influence on the hip fracture risk. Age-related bone loss was associated with a neck width increase in unfractured and fractured patients. HAL was significantly longer in fractured patients and was a significant discriminator between fractured patients and normal controls. HAL was not significant as a discriminator between fractured and low-BMD unfractured patients. The intertrochanter-head center distance (from the intertrochanteric line to the femoral head center) coincides with the femoral lever arm and includes no segments that adapt to BMD changes, such as the greater trochanter-intertrochanter distance. Among all tested lengths, this segment was the part of HAL that discriminated best between fractured and low-BMD unfractured patients. A longer intertrochanter-head center distance increased the risk of femoral neck fracture among low-BMD patients. Including automatic measurement of this segment in standard DXA protocols may prove useful in identifying patients at high risk for hip fracture. At present, HAL remains the easier neck length to measure, but automatic evaluation of the intertrochanter-head center distance must be a goal for future image analysis development. PMID- 12111015 TI - Variations in bone density among persons of African heritage. AB - The epidemiology of bone loss in populations of African heritage is still poorly known. We compared a convenience sample of 47 African-American (AA) residents of Rochester, Minnesota (32 women, 15 men) and 66 recent immigrants from Somalia (all women) with 684 white subjects (349 women, 335 men) previously recruited from an age-stratified random sample of community residents. Areal bone mineral density (BMD, g/cm(2)) and volumetric bone mineral apparent density (BMAD, g/cm(3)) were determined for lumbar spine and proximal femur using the Hologic QDR 2000 for white subjects and the QDR 4500 for the others; the instruments were cross-calibrated from data on 20 volunteers. Lumbar spine BMD was 18% higher in AA ( p<0.001) and 4% lower in Somali ( p = 0.147) than white women. Femoral neck BMD was 27% higher in AA women but also 11% greater in Somali women (both p<0.001) compared with whites. Lumbar spine BMD was 6% higher ( p = 0.132) and femoral neck BMD 21% higher ( p<0.001) in AA than white men. No Somali men were studied. After correcting for bone size differences, both lumbar spine ( p<0.01) and femoral neck BMAD ( p<0.001) were greater for Somali than white women, but the difference between Somali and AA women persisted. Lumbar spine and femoral neck BMAD values also remained significantly greater for AA women (both p<0.001) and men ( p<0.05; p<0.001) compared with whites. Weight was associated with BMAD at both skeletal sites in all groups, but adjustment for differences in weight did not reduce the discrepancy in BMAD values between Somali and AA women or between the latter group and whites. This heterogeneity among different ethnic groups of African heritage may provide an opportunity for research to better explain race-specific differences in bone metabolism. PMID- 12111016 TI - The decision to accept treatment for osteoporosis following hip fracture: exploring the woman's perspective using a stage-of-change model. AB - Despite the availability of medications that reduce fracture risk, most women who sustain a hip fracture are not evaluated or treated for osteoporosis. While a number of studies have attributed this to a lack of physician awareness, no studies have evaluated this problem from the patient's perspective. To explore the process a woman negotiates when deciding to accept pharmacologic treatment for osteoporosis after hip fracture, we used a stage-of-change model to characterize a consecutive series of 70 postmenopausal women (mean age 85 years) admitted to a tertiary care hospital with an acute low-impact hip fracture between May 2000 and August 2000. We measured stage-of-change using a modified form of the Weinstein Precaution Adoption Process Model (PAPM). The majority of patients (65%) were ineligible because of dementia or delirium; only 29 were eligible and 21 were enrolled. Most women (62%) were in stages 1 or 2 of the PAPM, indicating that they were unaware of osteoporosis or had never considered pharmacologic treatment for it. The only factors associated with a more advanced PAPM stage (indicating active consideration or currently taking treatment) were a previous bone mineral density (BMD) evaluation ( p = 0.007) and a diagnosis of osteoporosis ( p = 0.001). Although 48% of women had a previous fragility fracture and osteoporosis knowledge was poor overall (mean score 52% correct), neither was associated with a more advanced PAPM stage in this sample. In conclusion, women evaluated after hip fracture were not ready to accept pharmacologic treatment for osteoporosis; they were unaware that they had osteoporosis or had never considered treatment for it. For a woman to advance through the behavior change process, she must first be made aware of the problem that requires a change in behavior. Physicians play a crucial role in promoting awareness of the diagnosis of osteoporosis after fracture, which in turn is associated with patient advancement through the behavior change process and the decision to accept pharmacologic intervention. The large number of cognitively impaired patients in this population, however, will certainly make efforts to improve osteoporosis awareness, diagnosis and intervention more challenging. PMID- 12111018 TI - Differential effects of primary hyperparathyroidism on ultrasound properties of bone. AB - Primary hyperparathyroidism (PHPT) may result in greater cortical than trabecular bone loss. Ultrasound is able to predict osteoporotic fracture risk independent of densitometric measurements, but little is known about the changes in ultrasound variables with PHPT. The aim of our study was to examine the effect of PHPT on ultrasound variables and bone density measurements at cortical (hand) and trabecular (lumbar spine and heel) sites, and to evaluate their reversibility following surgical treatment. We recruited 25 postmenopausal women diagnosed with PHPT ages 51-76 years (mean 62 years) and 95 postmenopausal controls ages 57-80 years (mean 67 years). Measurements were made at baseline and 1 year. Speed of sound (SOS) and broadband ultrasound attenuation (BUA) of the heel were measured using the Lunar Achilles (LA+) and McCue CUBA Clinical (CC). Amplitude-dependent speed of sound (AD-SoS) and ultrasound bone profile index (UBPI) of the fingers were measured using the IGEA DBM Sonic. Bone mineral density (BMD) of the hand and lumbar spine (LS) were measured by dual-energy X-ray absorptiometry (DXA). At baseline, hand BMD, LS BMD and heel BUA were significantly lower and finger UBPI significantly higher in the PHPT patients compared with controls ( p<0.001). There were no differences in Stiffness Index, heel SOS or finger AD-SoS between control and PHPT subjects. At 1 year postoperatively, there was a mean (+/-SD) increase in LS and hand BMD of 3+/-1% ( p<0.01). BUA at the heel increased (11+/ 5%, p<0.001), and UBPI of the fingers decreased (17+/-7%, p<0.001) probably reflecting different modes of attenuation in trabecular (scattering) and cortical (absorption) bone. Stiffness Index, SOS of the heel and AD-SoS of the fingers did not change. BUA, UBPI and BMD returned towards normal postmenopausal values following surgery. There were no changes in BMD or QUS variables at 1 year in the control group. Quantitative ultrasound (QUS) measurements provide different information about bone structure than densitometric measurements and cannot be regarded as simply reflecting bone density. With further research the combined use of BMD and QUS could improve the assessment of skeletal status in patients with PHPT before and after surgery. PMID- 12111017 TI - Incidence of limb fracture across Europe: results from the European Prospective Osteoporosis Study (EPOS). AB - The aim of this population-based prospective study was to determine the incidence of limb fracture by site and gender in different regions of Europe. Men and women aged 50-79 years were recruited from population registers in 31 European centers. Subjects were invited to attend for an interviewer-administered questionnaire and lateral spinal radiographs. Subjects were subsequently followed up using an annual postal questionnaire which included questions concerning the occurrence of new fractures. Self-reported fractures were confirmed where possible by radiograph, attending physician or subject interview. There were 6451 men and 6936 women followed for a median of 3.0 years. During this time there were 140 incident limb fractures in men and 391 in women. The age-adjusted incidence of any limb fracture was 7.3/1000 person-years [pyrs] in men and 19 per 1000 pyrs in women, equivalent to a 2.5 times excess in women. Among women, the incidence of hip, humerus and distal forearm fracture, though not 'other' limb fracture, increased with age, while in men only the incidence of hip and humerus fracture increased with age. Among women, there was evidence of significant variation in the occurrence of hip, distal forearm and humerus fractures across Europe, with incidence rates higher in Scandinavia than in other European regions, though for distal forearm fracture the incidence in east Europe was similar to that observed in Scandinavia. Among men, there was no evidence of significant geographic variation in the occurrence of these fractures. This is the first large population-based study to characterize the incidence of limb fracture in men and women over 50 years of age across Europe. There are substantial differences in the descriptive epidemiology of limb fracture by region and gender. PMID- 12111019 TI - Independent living after fractures in the elderly. AB - Although fractures are an important source of disability among the growing elderly populations of industrialized societies, patient-centered multidimensional outcome information is scarce. The purpose of this study was to quantify the natural history of recovery from fractures of the upper and lower extremities. From the 1994/95 WHO MONICA survey in Augsburg, Germany, we selected all persons aged 58-78 years who had experienced a fracture during the preceeding 10 years, along with a control population twice as large. The Health Assessment Questionnaire (HAQ) and the Medical Outcomes Study Short Form 36 (SF-36) were administered to these subjects in 1998. Patients' recollection of fracture type and location were validated against medical records. The most recent fracture was in the upper extremity in 45 cases, lower extremity in 55 cases and elsewhere in 46 cases. Extremity fractures resulted in persistent and measurable impairment of the activities of daily living or general quality of life in patients 65 years or older, especially if the femur was involved. More than 40% of the interindividual variation of functional disability in the study group could be explained by age, sex, history of a fracture within 12 years and perceived difficulties walking. Existing generic and specific musculoskeletal outcome measurement instruments thus allow the assessment of functional recovery and health status after fractures in an elderly population. Geriatric assessment following fractures at higher age may improve ability to live independently. Difficulty walking deserves special attention, as it is associated with more general functional disability among the elderly. PMID- 12111020 TI - Quantitative ultrasound assessment of acute bone loss following spinal cord injury: a longitudinal pilot study. AB - Spinal cord injury (SCI) results in substantial and rapid osteoporosis. Given its rapid onset, assessment of bone changes in the early stages (first 6 months) following SCI is important. This is particularly pertinent if intervention is to be implemented. Quantitative ultrasound (QUS) represents a potential assessment tool for the evaluation of skeletal changes in the early stages following SCI. This longitudinal pilot study assessed changes in QUS measures of calcaneal broadband ultrasound attenuation (BUA) and speed of sound (SOS) in 15 male subjects (age 23.9+/-7.3 years) over a 6-week period. Their mean time since SCI was 110.3+/-34.5 days. Also assessed were bone mineral density of the calcaneus (BMDc) and proximal tibia (BMDt) using dual-energy X-ray absorptiometry (DXA). Confirming the rapid onset of bone loss following SCI, BMDc and BMDt decreased by 7.5+/-3.0% ( p<0.001) and 5.3+/-4.2% ( p<0.001), respectively. QUS was sensitive to these changes. BUA decreased by 8.5+/-6.9% ( p<0.001), whilst SOS decreased by 1.5+/- 1.3% ( p<0.001). Suggesting an influence of the material properties of bone on BUA, BUA was correlated with BMDc at both the initial ( r = 0.68, p<0.01) and final ( r = 0.62, p<0.01) assessments. There were no significant correlations in the magnitude of change over the 6-week assessment period between any of the skeletal measures (all p>0.05). This suggests that skeletal qualities other than material properties also influence QUS measures. Overall, this study confirmed the rapid onset of bone loss following SCI and showed QUS to be a useful portable measure of acute bone changes. This may allow assessment of bone loss and the efficacy of intervention on this loss in the early stages following injury, a period where traditional axial DXA assessment is limited by practical constraints. PMID- 12111021 TI - Use of clinical risk factors in elderly women with low bone mineral density to identify women at higher risk of hip fracture: The EPIDOS prospective study. AB - Elderly women with very low bone mineral density (BMD) ( T-score or = 75 %). RESULTS: The 118 patients were dissatisfied with items that indicated curtailment of their freedom, while the 35 staff were dissatisfied with the physical facilities for care. Highest satisfaction for patients and staff were for items on staff-patient relationship. Barely satisfactory items for patients included the time spent with doctors. Patients had a higher positive appraisal of the adequacy of physical facilities than staff, while staff had a more positive appraisal of their relationship with patients. There were no significant differences in satisfaction among diagnostic groups. CONCLUSION: The logical and discriminating manner in which patients assessed satisfaction supports the impression that they can be relied upon to make objective appraisal of the process of care, and that patient satisfaction is a valid index of the quality of care. PMID- 12111035 TI - An empirical comparison of substance and alcohol dependence patterns in the homeless in Madrid (Spain) and Los Angeles (CA, USA). AB - BACKGROUND: Alcohol and drug use continue to figure heavily in the experience of the contemporary homeless population. The comparison among pattern of use plays a central role in the cross-cultural view of this topic. This article shows the results of comparing the data concerning alcohol and other drug abuse and dependence among the homeless population of Madrid (Spain) and Los Angeles (USA). METHODS: Data come from two studies carried out independently in each city. Both studies used a comparable methodology which included the same inclusion and diagnostics criteria, representative sampling methods and similar diagnostic structured interviews. In the present study, the data from these two studies are combined in a unique database which allows global and item-to-item comparison between the two studies. RESULTS: The results show different sociodemographic profiles for each city. Once controlled for the sociodemographic differences (age, education, current employment status and marital status), the life and 12 month prevalence rates of alcohol and other drug disorders (DSM-III-R) are also different. There are also significant differences in social, emotional and health problems associated with the consumption of alcohol and other drugs. The Madrid and LA samples also present differences in the time patterns of the beginning of the homelessness situation and the onset of alcohol- and drug-related disorders. CONCLUSIONS: The pattern of results is discussed in the light of the differences in both socioeconomic and cultural among Madrid and Los Angeles which might explain, in turn, differences in the homelessness situation as well as in the alcohol and other drug use patterns. PMID- 12111036 TI - The neutrophil as a mediator of myocardial ischemia-reperfusion injury: time to move on. AB - Granulocytes, especially neutrophils, are recruited in myocardium during the evolution of acute myocardial infarction. Because the neutrophil reaction is most intense during reperfusion and because these cells are a rich source of toxic oxidant species and proteolytic enzymes, it has become a widely held view that neutrophils are an important mechanism of myocardial injury extension during reperfusion. However, on close examination the evidence underlying this contention is equivocal. The basic experimental situation can be summarised thus. (1) All forms of reperfusion injury (i.e., cytotoxic or lethal cell injury, myocardial stunning, endothelial dysfunction, and reperfusion-induced arrhythmias) can be observed in neutrophil-free conditions. (2) "Anti-neutrophil" interventions (e.g., anti-inflammatory drugs, adenosine, anti-neutrophil antisera, leukocyte filters and inhibitors of the various pathways of neutrophil adhesion) do not consistently prevent reperfusion injury and they certainly do not consistently limit infarct size. (3) The time course of neutrophil accumulation in post-ischaemic myocardium may be different to the time course of injury. (4) Despite more than two decades of research, no double-blind, randomised controlled clinical trial assessing an anti-neutrophil therapy in myocardial infarction has yet reported a positive benefit that is attributable to inhibition of neutrophil recruitment. The evidence weighs against a pivotal role of neutrophils as a causal factor in most forms of ischemia-reperfusion injury. An exception may be microvascular injury and capillary plugging leading to the "no-reflow" phenomenon but even here the evidence suggests that the extent of neutrophil accumulation and microvascular injury is determined by, rather than a cause of, myocyte necrosis. PMID- 12111037 TI - The role of p38 mitogen-activated protein kinase in myocardial ischemia/reperfusion injury; relationship to ischemic preconditioning. AB - Myocardial mitogen-activated protein kinases can be activated by ischemia and reperfusion, and they may play important roles in the evolution of ischemic injury. Considerable work has been performed to evaluate the role of different MAPK signaling pathways in ischemia/reperfusion injury. The focus of this review is the p38 MAPK pathway, specifically whether activation of the p38 MAPK signaling pathway is beneficial or detrimental. Different studies have come to conflicting conclusions. This review will examine the literature on the role of p38 MAPK in myocardial ischemia/reperfusion injury, highlight areas of controversy and areas of general agreement, examine possible downstream targets of p38 during acute ischemia, and attempt to draw some conclusions. PMID- 12111038 TI - Contribution of I(K,ADO) to the negative dromotropic effect of adenosine. AB - OBJECTIVE: Despite the pathophysiological and therapeutic significance of the negative dromotropic effect of adenosine, its underlying ionic mechanism, and specifically the role of the adenosine-activated K(+) current (I(K,ADO)) is not experimentally defined. Therefore, we studied the contribution of I(K,ADO) to the negative dromotropic effect of adenosine. METHODS: Effects of adenosine on single atrioventricular nodal and left atrial myocytes from rabbits were studied using the whole cell configuration of the patch clamp technique. Complementary experiments were done in rabbit and guinea pig isolated hearts instrumented to measure the atrium-to-His bundle interval. RESULTS: In contrast to its effect in atrial myocytes, Ba(2+) selectively and completely blocked I(K,ADO) at membrane potentials from -70 to 0 mV in atrioventricular nodal myocytes and abolished the adenosine-induced leftward shift of the reversal membrane potential. Ba(2+) alone did not significantly prolong the A-H interval, but markedly attenuated the A-H interval prolongation caused by adenosine. In guinea pig heart, EC(50) values ( pD(2) +/- SEM) for adenosine-induced atrium-to-His bundle interval prolongation were 3.3 micromol/L (5.48 +/- 0.04) and 13.2 micromol/L (4.88 +/- 0.05, P < 0.001) in the absence and presence of Ba(2+), respectively. Despite species dependent differences in sensitivities to adenosine (guinea pig > rabbit), the relative contribution of adenosine-activated K(+) current to the atrium-to-His bundle interval prolongation was nearly identical. In guinea pig hearts it ranged from 37.8 % (P = 0.013) to 72.5 % (P < 0.001) at 2 to 6 micromol/L adenosine, respectively. CONCLUSION: I(K,ADO) contributes significantly to the negative dromotropic effect of adenosine, but predominantly at relatively high concentrations of the nucleoside. PMID- 12111039 TI - Potentiation of the negative dromotropic effect of adenosine by rapid heart rates: possible ionic mechanism. AB - Adenosine-induced slowing of atrioventricular nodal conduction is a rate dependent process that is potentiated by the A(1)-adenosine receptor allosteric enhancer, PD 81,723. The ionic mechanisms underlying these phenomena were investigated in guinea pig isolated hearts and single atrial myocytes by measuring the atrium-to-His bundle (A-H) interval and using patch-clamp recordings, respectively.A decrease in atrial cycle length from 300 to 190 ms decreased the concentration of adenosine needed to cause atrioventricular nodal block from 7.8 +/- 1.0 to 2.6 +/- 0.7 micromol/L (P < 0.001). Ba(2+) (100 micromol/L), a selective blocker of the adenosine-activated inward rectifier K(+) current I(K,ADO) in the atrioventricular node, failed to abolish this rate dependent effect of adenosine. PD 81,723 (5 micromol/L) potentiated the negative dromotropic effect of adenosine even after I(K,ADO) was blocked by Ba(2+) and after attenuation of I(Ca,L) by adenosine was prevented by 8-Br-cAMP (1.5 mmol/L). In atrial myocytes, adenosine augmented a time- and voltage-dependent K(+) current (Ado-I(K)). Ado-I(K) was more sensitive to adenosine than I(K,ADO) (EC(50) values, 0.8 versus 1.4 micromol/L, P < 0.01). PD 81,723 blocked I(K,ADO), but potentiated Ado-I(K). Ado-I(K) was insensitive to Ba(2+) (P = 0.98), whereas it was blocked by chromanol 293B (5 micromol/L, P < 0.001). Unlike I(K,ADO), Ado I(K) increased during rapid stimulation of myocytes (P < 0.001). Adenosine augments a time- and voltage-dependent K(+) current, Ado-I(K). The pharmacological and kinetic properties of Ado-I(K) are consistent with it playing an important role in the negative dromotropic effect of adenosine at lower concentrations of the nucleoside, at fast heart rates and in the presence of PD 81,723. PMID- 12111040 TI - Influence of perfusate calcium concentration on the inotropic insulin effect in isolated guinea pig and rat hearts. AB - Conflicting data on the inotropic effect of insulin are present in the literature suggesting a positive inotropic property or no inotropic effect or even a negative influence. To clarify the reason for these diverging findings, dose response curves of insulin have been performed in isolated working rat and guinea pig hearts perfused with Krebs-Henseleit buffer containing 0.9, 1.25, 2.5, and 5 mM Ca(2+) at 37 degrees C. At 1.25 mM [Ca(2+)], insulin (8 to 16 IU) regularly improved the inotropic state. LVdP/dt(max) increased significantly from 1,900 to 2,300 mm Hg/s (+21 %) in guinea pig hearts and from 3,197 to 4,345 mm Hg/s (+36 %) in rat hearts. LVEDP did not change significantly. Myocardial oxygen consumption increased parallel with contractility. Heart rate was not influenced in either species. Coronary flow increased by 16.5 % in guinea pig hearts, but decreased in rat hearts by 13.6 % (p < 0.05 each). With 0.9 mM [Ca(2+)] the positive inotropic effect of insulin did not further augment. At 2.5 mM [Ca(2+)] insulin exhibited in both species no significant change of LVdP/dt(max) but very high insulin doses depressed the heart. At 5 mM [Ca(2+)] insulin depressed the heart significantly already at lower concentrations. At 31 degrees C and 1.25 mM [Ca(2+)] the positive inotropic insulin effect was preserved. We conclude that the positive inotropic insulin effect in rat and guinea pig hearts depends on the extracellular [Ca(2+)], i.e., is maximal around 1.25 mM [Ca(2+)] and is reduced or absent at higher [Ca(2+)] or may even become negative. PMID- 12111041 TI - Myocardial ischemia and reperfusion reduce the levels of cyclic ADP-ribose in rat myocardium. AB - Cyclic ADP-ribose (cADPR) is a novel Ca(2+)-mobilizing second messenger in mammalian cells including cardiomyocytes. It is unknown whether myocardial ischemia and reperfusion affect the metabolism of cADPR in the myocardium. The present study therefore examined the effects of myocardial ischemia and reperfusion on the concentrations of myocardial cADPR using high-performance liquid chromatography. Basal levels of cADPR in rat myocardium were 5.3 +/- 1.8 nmol x mg(-1) protein. Myocardial ischemia for 30 min significantly decreased cADPR concentrations to 2.1 +/- 0.4 nmol x mg(-1) protein. During reperfusion, cADPR was maintained at ischemic levels. The activity of ADP-ribosyl cyclase was expressed as the conversion rate of nicotinamide guanine dinucleotide (NGD(+)) to cyclic GDP-ribose. Myocardial ischemia and reperfusion did not alter the activity of ADP-ribosyl cyclase. However, cADPR hydrolase activity, as measured by the conversion rate of cADPR to ADP-ribose, was significantly elevated by ischemia and reperfusion. To determine the mechanism resulting in the enhancement of cADPR hydrolase activity, we examined the effects of changes in ADP, ATP, pH, and PO(2) on the conversion rate of cADPR to ADPR. Alterations of ADP, ATP, or pH in myocardial tissue had no effect on the degradation of cADPR, whereas a decrease in tissue PO(2) markedly increased the hydrolysis of cADPR. These results suggest that myocardial ischemia and reperfusion decrease cADPR in the myocardium by increasing its hydrolysis. Tissue hypoxia may be one of the important mechanisms to activate cADPR hydrolase. PMID- 12111042 TI - Central AT1 receptors are involved in the enhanced cardiac sympathetic afferent reflex in rats with chronic heart failure. AB - The purpose of this study was to determine if the cardiac sympathetic afferent reflex (CSAR) was augmented in rats with coronary ligation-induced chronic heart failure (CHF), and if central angiotensin II (ANG II) was involved in this enhancement. Under alpha-chloralose and urethane anesthesia, mean arterial pressure (MAP), heart rate (HR) and renal sympathetic nerve activity (RSNA) were recorded in sino-aortic denervated and cervical vagotomized rats. An intracerebroventricular cannula was implanted. The CSAR was examined by epicardial application of bradykinin (BK, 0.04 and 0.4 microg in 2.0 microl) or capsaicin (0.04 and 0.4 microg in 2.0 microl) to the anterior and posterior wall of the left ventricle. The CSAR evoked by BK or capsaicin was augmented in rats with CHF. In sham rats, there was no significant difference of the CSAR induced by BK or capsaicin between anterior and posterior epicardial application. However, in rats with CHF, the CSAR induced by BK (0.04 microg) to anterior epicardial application was blunted compared with posterior application. Intracerebroventricular injection of losartan (500 nmol) normalized the enhanced CSAR in rats with CHF, but had no significant effects on the CSAR in sham rats. However, intravenous application of the same dose of losartan only decreased the baseline MAP, but did not alter baseline RSNA or the CSAR in sham or CHF rats. Pre-treatment with epicardial application of lidocaine to the anterior wall abolished the CSAR evoked by application of BK or capsaicin but had no effects on the CSAR evoked by epicardial application of BK or capsaicin to the posterior wall. These results suggest that the CSAR induced by epicardial application of BK and capsaicin is enhanced in the rats with CHF, and the enhanced CSAR is mediated by central AT(1) receptors. PMID- 12111043 TI - Angiotensin II type 1 receptor expression in human coronary arteries with variable degrees of atherosclerosis. AB - Autopsy specimens of human coronary arteries were collected from 65 men and women ranging in age from 40-76 years of age. We made 209 coronary artery sections, which were graded in terms of severity of atherosclerosis by means of the Stary classification. Sections were stained using an antibody directed against the angiotensin II type 1 (AT1) receptor. We found that in non-atherosclerotic sections, staining was confined to vascular smooth muscle cells in the media. However, with the advent of atherosclerosis, AT1 receptor expression was also present in atherosclerotic plaque and involved other cell types including inflammatory cells and myofibroblasts. We identified a remarkable correlation between AT1 receptor staining and the severity of coronary atherosclerosis as well as intima-media thickness. These human data correspond well to animal models of atherosclerosis indicating an upregulation of AT1 receptor expression in atherosclerotic tissue. PMID- 12111044 TI - Angiotensin II-augmented migration of VSMCs towards PDGF-BB involves Pyk2 and ERK 1/2 activation. AB - Activation of the local and systemic renin-angiotensin system is directly and indirectly involved in mechanisms of vascular remodeling during chronic hypertension. This study investigated the effect of angiotensin II (AII) on rat vascular smooth muscle cell (VSMC) migration towards platelet-derived growth factor-BB (PDGF-BB) in vitro. Pre-treatment with AII (1 microM) for 48 or 72 h induced a significant increase in PDGF-BB-directed migration by 77 +/- 21 % and 58 +/- 24 %, respectively (both p < 0.01). This effect was concentration dependent and inhibited by the selective angiotensin receptor type I (AT(1)) blocker DUP 753. PDGF-directed migration of VSMCs was significantly inhibited by antibodies against beta(3)-and beta(5)-integrins, indicating an important role of these integrins in VSMC migration. However, AII augmented migration was not accompanied by an increased expression of beta(3)- and beta(5)-integrin mRNA and protein levels in VSMCs. Inhibition of the mitogen-activated protein kinase ERK 1/2 with PD 98059 (30 microM) completely abolished the effect of AII on PDGF-BB directed VSMC migration (p < 0.01). The proline-rich tyrosine kinase 2 (Pyk2) and focal adhesion kinase (FAK) are cytoskeleton-associated protein kinases participating in integrin-dependent signaling. Therefore, expression and phosphorylation of these kinases was determined 48 h after AII treatment, revealing a significant increase in Pyk2 and FAK protein levels (up to 2-fold, both p < 0.05) and increased phosphorylation of Pyk2 (2-fold, p < 0.05) and ERK 1/2 (4-fold, p < 0.05) as compared to controls. Furthermore, immunofluorescence and Western blot analysis demonstrated a translocation of Pyk2 from the plasma membrane to the cytosol, as well as a perinuclear enrichment of ERK 1/2 protein 48 h after AII treatment. In conclusion, our data suggest that changes in the levels of Pyk2 and ERK 1/2 phosphorylation, responsible for integrin-dependent signaling, as well as their subcellular translocation are important for the enhanced chemotactic response of VSMCs after AII pre-treatment. PMID- 12111045 TI - Spinach and tomato consumption increases lymphocyte DNA resistance to oxidative stress but this is not related to cell carotenoid concentrations. AB - BACKGROUND: The increased consumption of fruit and vegetables has been linked to protection against different chronic diseases, but the dietary constituents responsible for this association have not been clearly identified. AIM OF THE STUDY: We evaluated the effect of spinach and spinach+tomato puree consumption on cell DNA resistance to an oxidative stress. METHODS: To this aim, in a dietary controlled intervention study, 9 healthy female volunteers consumed a basal diet low in carotenoids (< 600 microg/day) enriched with daily portions (150 g) of spinach (providing about 9 mg lutein, 0.6 mg zeaxanthin, 4 mg beta-carotene) for 3 weeks (from day 0 to day 21) followed by a 2 week wash-out period (basal diet) and finally another 3 weeks (from day 35 to day 56) of diet enriched with daily portions of spinach (150 g) + tomato puree (25 g, providing about 7 mg lycopene, 0.3 mg beta-carotene). At the beginning and the end of each period of vegetable intake, blood samples were collected for lymphocyte separation. Carotenoid concentrations of lymphocytes were determined by HPLC and DNA damage was evaluated by the comet assay following an ex vivo treatment with H(2)O(2). RESULTS: During the first period of spinach consumption, lymphocyte lutein concentration did not increase significantly (from 1.6 to 2.2 micromol/10(12) cells) while lycopene and beta-carotene concentrations decreased significantly (from 1.0 to 0.1 micromol/10(12) cells, P < 0.001, and from 2.2 to 1.2 micromol/10(12) cells, P < 0.05, respectively). Lutein and lycopene concentrations increased after spinach+tomato puree consumption (from 1.2 to 3.5 micromol/10(12) cells, P < 0.01, and from 0.1 to 0.7 micromol/10(12) cells, P < 0.05, respectively). The increase may be attributed to the addition of tomato puree to spinach; however, the different concentrations of carotenoids in lymphocytes registered at the beginning of the two intervention periods may have affected the results. DNA resistance to H(2)O(2) insult increased significantly after both the enriched diets (P < 0.01); however, no "additive effect" was seen after spinach + tomato puree consumption. In the spinach + tomato intervention period an inverse correlation was observed between lymphocyte lycopene concentration and DNA damage, but this seems not able to explain the protection observed. CONCLUSIONS: The consumption of carotenoid-rich foods even for a short period of time gives protection against oxidative stress. The results obtained seem to suggest that this protective role is not specifically related to carotenoids. However they may contribute together with other substances present in vegetables to lymphocyte resistance to oxidative damage. PMID- 12111046 TI - The efficacy of dietary intervention alone or combined with hormone replacement therapy in postmenopausal women with hypercholesterolemia in Seoul, Korea. AB - BACKGROUND: Women have an increased incidence of cardiovascular disease (CVD) due to hormone imbalance-induced changes in blood lipid profiles after menopause. AIM OF STUDY: This study was done to compare the effects of dietary intervention and hormone replacement therapy, alone or in combination, on blood lipids and body weight in Korean postmenopausal women with hypercholesterolemia. METHOD: The subjects were treated by one of three different treatments for 12 weeks: hormone replacement therapy (HRT group, n = 8), dietary intervention (DIET group, n = 8) and hormone replacement therapy combined with dietary intervention (HRT+DIET group, n = 8). RESULTS: Serum TC and LDL-C levels decreased by 13-16 % and 24-28 % in the HRT group, by 17-19 % and 21-23 % in the DIET group and by 19-26 % and 32-39 % in the HRT+DIET group, respectively ( P < 0.05). Serum HDL-C levels decreased in the DIET group (-6.4 %, P < 0.05) but not in the HRT and HRT+DIET groups. Serum TG levels increased in the HRT group (18 %, P < 0.05) but decreased in the DIET group (-24.4 %, P < 0.05). In the HRT+ DIET group, serum TG levels did not change. Body weight decreased only in the DIET group. CONCLUSIONS: We can conclude that dietary intervention produces a considerable improvement in blood lipid profiles and body weight, even though our study is limited by the sample size. Thus, the treatment to reduce risk of CVD should be individualized on the basis of the patient's dietary intake status, and at least, HRT should not be substituted for dietary intervention. PMID- 12111047 TI - A new equation especially developed for predicting resting metabolic rate in the elderly for easy use in practice. AB - BACKGROUND: Equations published in the literature for predicting resting metabolic rate (RMR) in older individuals were exclusively derived from studies with small samples of this age group. AIM: of the present investigation was therefore to compare the measured RMR of a relatively large group of older females and males with values for RMR calculated from the most commonly used WHO [1] equations. Furthermore, on the basis of the data collected by our study group a new equation for calculating RMR in the elderly was to be developed. Variables used in this equation should be easily and exactly determinable in practice. SUBJECTS AND METHODS: RMR was measured by indirect calorimetry after an overnight fast in a sample of 179 female (age 67.8 +/- 5.7 y, BMI 26.4 +/- 3.7 kg/m(2)) and 107 male (age 66.9 +/- 5.1 y, BMI 26.3 +/- 3.1 kg/m(2)) participants in the longitudinal study on nutrition and health status in an aging population of Giessen, Germany. The subjects were at least 60 years old, did not suffer from thyroid dysfunction, and were not taking thyroid hormones. Stepwise multiple linear regression analysis was used to estimate the best predictors of RMR. RESULTS: In females there was no significant difference between our measured RMR (5504 +/- 653 kJ/d) and RMR predicted with the WHO [1] equation (5458 +/- 440 kJ/d), whereas in males measured RMR (6831 +/- 779 kJ/d) was significantly higher than calculated RMR (6490 +/- 550 kJ/d). Results of regression analysis, considering body weight, body height, age, and sex, showed that RMR is best calculated by the following equation: RMR [kJ/d]= 3169 + 50.0 x body weight [kg] 15.3 x age [y] + 746 x sex [female = 0, male = 1]. The variables of this equation accounted for 74 % (R(2)) of the variance in RMR and predicted RMR within +/- 486 kJ/d (SEE). CONCLUSION: On the basis of the data determined in a large group of older individuals, we offer a new equation for calculating RMR in the elderly that is both easy and accurate for use in practice. PMID- 12111048 TI - Beta(2)-adrenergic receptor mutation and abdominal obesity risk: effect modification by gender and HDL-cholesterol. AB - OBJECTIVE AND DESIGN: A case-control study was conducted to examine the association between the 27Glu polymorphism of the beta(2)-adrenergic receptor gene (ADRB2) and the risk of abdominal obesity (defined by a waist/hip ratio: WHR higher than 0.85). METHODS: The case series encompassed 112 obese subjects with body mass index (BMI) > 30 kg/m(2) and WHR > 0.85 and no other major disease except for type 2 diabetes, while the controls were 127 healthy subjects, BMI < 25 kg/m(2) and WHR < 0.85. RESULTS: The association between the risk of abdominal obesity and the 27Glu polymorphism was estimated using multivariate logistic regression. A higher crude odds ratio (OR) of 4.08 (95 % confidence interval: 0.98-16.3) for the 27Glu allele was found among men, while no increased risk was apparent among female participants. Moreover, when the model was adjusted for age, male subjects carriers of the 27Glu allele had a significant ten-fold higher risk of abdominal obesity (OR = 10.31; 95 % CI: 1.4-76.8) and the product-term for the interaction (effect modification) between gender and the ADRB2 mutation was near to the limits of statistical significance (Likelihood ratio test p = 0.056). Interestingly, we also found an effect modification with higher OR among individuals with low HDL-cholesterol (< 1.5 mmol/l) after adjustment for age and gender (OR = 2.87 95 % CI 1.09-7.50) and the product-term for interaction between the 27Glu allele and HDL-cholesterol was statistically significant (Likelihood ratio test p = 0.003). CONCLUSIONS: Our results showed that the 27Glu allele of the ADRB2 gene appears to be a risk factor for abdominal obesity among male subjects, specially among those with lower HDL-cholesterol levels. PMID- 12111049 TI - Tocopherol metabolites 2, 5, 7, 8-tetramethyl-2-(2'-carboxyethyl)-6 hydroxychroman (alpha-CEHC) and 2, 7, 8-trimethyl-2-(2'-carboxyethyl)-6 hydroxychroman (gamma-CEHC) in human serum after a single dose of natural vitamin E. AB - BACKGROUND: alpha- and gamma-Tocopherol are vitamin E compounds in human blood and tissues. alpha-CEHC (2,5,7,8-tetramethyl-2-(2'-carboxyethyl)-6 hydroxychroman) and gamma-CEHC (2,7,8-trimethyl-2-(2'-carboxyethyl)-6 hydroxychroman) have been identified as water-soluble metabolites which are excreted with the urine in humans. AIM OF THE STUDY: To assess over-time changes of serum levels of alpha- and gamma-CEHC in humans after a single dose of vitamin E from a natural source. METHODS: Twenty-one healthy subjects ingested a single dose of vitamin E (306 mg of RRR-alpha-tocopherol and 1.77 mg of gamma tocopherol). Blood was collected before (baseline) and 2, 6, 12, 24, 35, 50, and 74 h after ingestion. Serum was separated and levels of alpha- and gamma tocopherol and alpha- and gamma-CEHC were determined by HPLC. RESULTS: After vitamin E ingestion, a statistically significant increase was observed for alpha tocopherol and alpha-CEHC. Maximum serum levels for both compounds were measured 12 h after application (33.3 +/- 11.1 micromol alpha-toco-pherol /L and 42.4 +/- 18.3 nmol alpha-CEHC /L); baseline values were reached again after 72 h. While gamma-tocopherol levels decreased during the study period, an increase in the metabolite gamma-CEHC was observed. The optical isomer formed in the metabolism of RRR-alpha-tocopherol was assigned as S-alpha-CEHC. CONCLUSIONS: alpha-CEHC levels increase after administration of a single dose of natural vitamin E in humans. The appearance of the metabolite in blood parallels that of the parent compound. The gamma-tocopherol analog appears to be metabolized more efficiently than alpha-tocopherol. PMID- 12111050 TI - Systematic review of fatty acid composition of plasma phospholipids of venous cord blood in full-term infants. AB - The purpose of this review was to systematically evaluate the variability of the fatty acid composition of venous cord blood phospholipids in different populations. In an attempt to review published evidence systematically, we found 19 data sets describing fatty acid composition of venous cord blood phospholipids in 11 European and 2 American countries. The amount of saturated-, monounsaturated- and parent essential polyunsaturated fatty acids exhibited relatively moderate variability among the data sets reviewed. Values of arachidonic acid and docosahexaenoic acid showed two-fold variability among the data sets. The highest values of docosahexaenoic acid were observed in countries with apparently higher consumption of dietary fat from sea fish. Considering the differences in blood sampling, laboratory methods and data presentation, we conclude that fatty acid composition of venous cord blood phospholipids in healthy, full-term infants shows relatively modest variability; hence, it is suitable for the estimation of in utero fatty acid supply. PMID- 12111051 TI - Impact of parental BMI on the manifestation of overweight 5-7 year old children. AB - BACKGROUND: There is an increase in the prevalence of overweight and obese children. Genetic and environmental factors are contributing factors but the influence of parental nutritional state on early manifestation of overweight is not well characterised. AIM OF THE STUDY: To systematically investigate the impact of parental BMI on the manifestation of overweight in 5 to 7 year old children. METHODS: Cross-sectional study (as a part of the Kiel Obesity Prevention Study [KOPS]) of 3306 children aged 5-7 years and their parents. The nutritional state of the children (BMI, triceps skinfold, fat mass, prevalence of overweight) was investigated in subgroups differing with respect to parental BMI. RESULTS: BMI of the children was significantly correlated with parental BMI (r = 0.272, p < 0.01). Children's BMI showed closer associations with maternal than with paternal BMI (r = 0.254 vs. 0.159, p < 0.01). A multivariate regression analysis showed that parental BMI explained 7.6 % of the variance in children's BMI. OR for overweight was elevated in children with at least one overweight parent (overweight mother: OR 2.9 (boys)/3.1 (girls); overweight father: OR 1.8 (boys)/2.4 (girls). OR was highest for children with two obese parents (OR 7.6 (boys)/6.3 (girls). Children with one obese parent were more frequently overweight than children with one overweight parent. CONCLUSIONS: Parental BMI showed only a weak correlation with the BMI of their children. However, children's risk of becoming overweight increased with parental overweight and obesity. Thus, familial disposition has to be taken into account to identify risk groups for preventive measures. PMID- 12111052 TI - Traumatic injuries: imaging of peripheral musculoskeletal injuries. AB - The current dominant role of conventional radiography must be reassessed at increasingly shorter intervals in view of the continuing emergence of new imaging modalities that are available to diagnose peripheral musculoskeletal injuries. In comparison with conventional radiography, digital radiographic techniques offer advantages for optimization of image quality and dose, such as a wider dynamic range and post-processing of images. Currently, digital luminescence radiography (storage phosphor radiography) is the most commonly used digital method for obtaining radiographs, using the established positioning projections and routines of the film-screen technique. A new process, radiography with flat-panel amorphous silicon detectors, is still under development. Computed tomography is a valuable tool for diagnosing injuries of the peripheral musculoskeletal system, especially when three-dimensional data sets are acquired; these allow reformating images in all planes desired (2D technique) or in a volumetric format (3D technique). Established indications for CT in the peripheral skeleton are hip fractures, wrist injuries and calcaneal fractures; however, CT may be used as a supplement to radiography in every region of the body. Sonography is beginning to play an increasingly important role in trauma. Muscle and tendon injuries are the most common indications, but worthwhile information can be gained of the shoulder, elbow, hip, and knee joints, supplementing conventional or digital radiography. Magnetic resonance imaging effectively visualizes traumatic changes of the skeleton and the peripheral soft tissues. It is the method of choice to detect occult fractures. It can be used to diagnose muscle and tendon injuries. Joint injuries, especially in the knee and the shoulder joint, are common indications for MRI in the posttraumatic setting. PMID- 12111053 TI - Traumatic injuries: imaging and intervention of large arterial trauma. AB - Traumatic vessel injury can cause bleeding, thrombosis, embolization, or malperfusion due to external compression and spasm. Non-traumatic causes of acute large arterial emergencies include rupture of an aneurysm and pseudoaneurysm, dissection, embolization, and thrombosis in hypercoagulability syndromes. Ultrasonography is, of course, the imaging modality of choice in emergency cases; however, in central vascular injuries, spiral CT with contrast enhancement is the imaging modality that provides the most information. Angiography may be necessary for detailed information and before intervention. Stent-grafts are used to close large vascular lacerations, ruptured aortic aneurysms, and the entry tear of dissections. Interventional radiology methods play a major role in managing vascular emergencies. PMID- 12111054 TI - Non-traumatic neurological emergencies: imaging of cerebral ischemia. AB - Cardiovascular disease is the leading cause of death worldwide with almost one third of all cardiovascular deaths ascribed to stroke. Imaging modalities, such as CT, MRI, positron emission tomography (PET), and single photon emission CT (SPECT) provide tremendous insight into the pathophysiology of acute stroke. Computed tomography is considered the most important initial diagnostic study in patients with acute stroke, because underlying structural lesions, such as tumor, vascular malformation, or subdural hematoma, can mimic stroke clinically. Diffusion-weighted imaging (DWI) has the ability to visualize changes in diffusion within minutes after the onset of ischemia and has become a powerful tool in the evaluation of patients with stroke syndrome. Territories with diffusion and perfusion mismatch may define tissues at risk, but with potential recovery. An alternative strategy with CT technology uses rapid CT for dynamic perfusion imaging, with similar goals in mind. Angiography can be performed in the hyperacute stage if thrombolytic therapy is being considered. Indications for diagnostic angiography include transient ischemic attacks in a carotid distribution, amaurosis fugax, prior stroke in a carotid distribution, a high grade stenotic lesion in a carotid artery, acquiring an angiographic correlation of magnetic resonance angiography (MRA) or computed tomographic angiography (CTA) concerning stenotic findings. In 50% of all angiograms performed in the hyperacute stage, occlusion of a vessel is observed; however, the need for angiography has been made less necessary due to the improvements of MRA, duplex ultrasound, and CTA. Numerous etiologies can lead to infarction. In children, pediatric stroke is very uncommon. The most common cause is an embolus from congenital heart disease with right-to-left shunts. Also a dissection of large extracranial vessels may result in cerebral infarction, and although the brain is equipped with numerous venous drainage routes, the occlusion of a large sinus or a widespread vein obstruction will eventually lead to venous infarction. Thus, optimal stroke care requires not only early and exact identification of ischemia, but also a close collaboration between the clinician and radiologist. PMID- 12111055 TI - Non-traumatic neurological emergencies: emergency neuroradiological interventions. AB - This article summarizes recent developments in the growing field of interventional neuroradiology for the treatment of acute cerebrovascular disease. We describe the possibilities in endovascular therapy of acute cerebral aneurysms using electrolytically detachable coils combined with trispan neck bridging devices and stent implantation to occlude acute wide neck aneurysms. Techniques and results of local intra-arterial thrombolytic therapy in acute stroke and central retinal artery occlusion are described and we discuss the potential for rapid, large-burden thrombus removal in cases of internal carotid artery thrombosis by rheolytic thrombectomy, percutaneous transluminal angioplasty and stent implantation. Emergency endovascular therapy using the transvenous approach to treat severe intracranial or intraocular hypertension and multifocal haemorrhagic venous infarction due to cerebral sinus thrombosis or dural fistulas is also described. In cases of acute bleeding of head and neck lesions following trauma, tumours after radiotherapy, arteriovenous malformations, epistaxis or from iatrogenic origin, angiography plays a major role in localizing the source of bleeding and occluding the damaged vessel during the same session using the same endovascular approach. PMID- 12111056 TI - Intraindividual comparison of contrast-enhanced electron-beam computed tomography and navigator-echo-based magnetic resonance imaging for noninvasive coronary artery angiography. AB - The aim of this study was to compare contrast-enhanced electron-beam computed tomography (EBCT) and navigator-echo-based MRI of the coronary arteries in the same patient population. Both methods were assessed for visualization of the coronary arteries and their diagnostic accuracy in identifying significant coronary artery stenoses compared with conventional coronary angiography. Twenty patients with known coronary artery disease were examined with both contrast enhanced EBCT and a respiratory-gated MRI sequence. A grading system was used to evaluate the image quality. Sensitivity and specificity for the detection of significant coronary artery stenoses was evaluated compared with conventional coronary angiography. With EBCT, 89% of the main coronary arteries could be completely visualised in the proximal and middle segments; with MRI, 83% were visualised. With EBCT the sensitivities for identifying significant (>/=50%) stenoses in proximal and middle vessel segments were 75% in the main stem, 88% in the left anterior descending coronary artery, 75% in the left circumflex coronary artery, and 90% in the right coronary artery. Respective sensitivities for MRI angiograms were 75, 82, 75 and 80%. With both modalities a sufficient image quality of the main coronary arteries can be obtained in most cases. The diagnostic capability for detecting significant stenoses is comparable for both methods. PMID- 12111057 TI - Current concepts in imaging of laryngeal and hypopharyngeal cancer. AB - In adjunct to direct laryngoscopy, CT and/or MRI are needed for an accurate staging of laryngeal and hypopharyngeal carcinomas because both cross-sectional imaging modalities are known to reliably evaluate deep tumor infiltration. Except for the clinical background, this article reviews technical aspects of CT and MRI, the pathologic appearance of laryngeal and hypopharyngeal carcinomas, and therapeutically relevant diagnostic aspects. PMID- 12111058 TI - Virtual endoscopy of the middle ear: experimental and clinical results of a standardised approach using multi-slice helical computed tomography. AB - Virtual endoscopy (VE) enables non-invasive 3D endoluminal imaging of the middle ear by post-processing of CT data. To optimise the clinical application a standardised approach was evaluated in normal and pathologic cases. Data acquisition was performed using multi-slice helical CT in 20 normal patients and 15 patients with malformation or trauma. Virtual endoscopy of the tympanic cavity and 3D images of the ossicles were generated using surface and volume rendering. Qualitative assessment of the representation of anatomical structures was performed in normal patients. In 15 pathological cases the diagnostic benefit was evaluated by comparing the 3D images to the 2D images and intra-operative findings. In all 35 cases 3D imaging was possible using the standardised approach. The ossicular chain as well as the bony and soft tissue structures of the tympanic cavity were visualised in 20 normal patients. In 7 of 8 patients with malformation and 1 of 7 patients with trauma the original diagnosis was changed by 3D imaging. Standardisation and evaluation of the method in normal patients is essential as it enhances the diagnostic reliability. Virtual endoscopy facilitates understanding of the complex anatomy of the middle ear. In cases of suspected malformation and confirmed trauma it is helpful for diagnosis and surgical planning. PMID- 12111059 TI - Low-dose CT of the paranasal sinuses with eye lens protection: effect on image quality and radiation dose. AB - The purpose of the study was to assess the effect of lens protection on image quality and radiation dose to the eye lenses in CT of the paranasal sinuses. In 127 patients referred to rule out sinusitis, an axial spiral CT with a lens protection placed on the patients eyes was obtained (1.5/2/1, 50 mAs, 120 kV). Coronal views were reconstructed at 5-mm interval. To quantify a subjective impression of image quality, three regions of interest within the eyeball were plotted along a line perpendicular to the protection at 2, 5, and 9 mm beneath skin level on the axial images. Additionally, dose reduction of a bismuth containing latex shield was measured using a film-dosimetry technique. The average eyeball density was 17.97 HU (SD 3.7 HU). The relative increase in CT density was 180.6 (17.7), 103.3 (11.7), and 53.6 HU (9.2), respectively. There was no diagnostic information loss on axial and coronal views observed. Artifacts were practically invisible on images viewed in a bone window/level setting. The use of the shield reduced skin radiation from 7.5 to 4.5 mGy. The utilization of a radioprotection to the eye lenses in paranasal CT is a suitable and effective means of reducing skin radiation by 40%. PMID- 12111060 TI - Screen film vs full-field digital mammography: image quality, detectability and characterization of lesions. AB - The objective of this study was to compare screen-film mammography (SFM) to full field digital mammography (FFDM) regarding image quality as well as detectability and characterization of lesions using equivalent images of the same patient acquired with both systems. Two mammography units were used, one with a screen film system (Senographe DMR) and the other with a digital detector (Senographe 2000D, both GEMS). Screen-film and digital mammograms were performed on 55 patients with cytologically or histologically proven tumors on the same day. Together with these, 75 digital mammograms of patients without tumor and the corresponding previous screen-film mammograms not older than 1.5 years were reviewed by three observers in a random order. Contrast, exposure, and the presence of artifacts were evaluated. Different details, such as the skin, the retromamillary region, and the parenchymal structures, were judged according to a three-point ranking scale. Finally, the detectability of microcalcifications and lesions were compared and correlated to histology. Image contrast was judged to be good in 76%, satisfactory in 20%, and unsatisfactory in 4% of screen-film mammograms. Digital mammograms were judged to be good in 99% and unsatisfactory in 1% of cases. Improper exposure of screen-film system occurred in 18% (10% overexposed and 8% underexposed). Digital mammograms were improperly exposed in 4% of all cases but were of acceptable quality after post-processing. Artifacts, most of them of no significance, were found in 78% of screen-film and in none of the digital mammograms. Different anatomical regions, such as the skin, the retromamillary region, and dense parenchymal areas, were better visualized in digital than in screen-film mammography. All malignant tumors were seen by the three radiologists; however, digital mammograms allowed a better characterization of these lesions to the Breast Imaging Reporting and Data System (BI-RADS;) [corrected] categories (FFDM better than SFM in 23 of 165 vs 9 of 165 judged cases in SFM). In conclusion, digital mammography offers a consistent, high image quality in combination with a better contrast and without artifacts. Lesion detection in digital images was equal to that in screen-film images; however, categorization of the lesions to the BI-RADS classification was slightly better. PMID- 12111061 TI - An experience with the Advanced Breast Biopsy Instrumentation (ABBI) system in the management of non-palpable breast lesions. AB - Our objective was to evaluate our experience with the Advanced Breast Biopsy Instrumentation system (ABBI) in non-palpable breast lesions in a prospective study from July 1998 to November 2000. The ABBI system was included in a protocol for BIRADS 4 non-palpable, small (<15 mm) breast lesions. Digital radiographs of both specimen and biopsy cavity were obtained to validate the procedure. A total of 255 ABBI biopsies were performed in 254 patients. In 251 cases the lesions were successfully removed (98.4%). Mammographic lesions consisted of 176 cases of microcalcifications (69%), 51 cases of architectural distortions (20%) and 28 cases of nodules (11%). Seventy-two carcinomas were diagnosed (28.2%). Affected margins were found in 41 cases (56.9%). Residual tumour was seen in 31 patients (43%). Seventeen borderline results and 33 benign architectural distortions obviated further procedures. The complication rate in 10 cases was as follows: 3 wound infections; 4 haematomas; and 3 vasovagal reactions. The main utility of the ABBI system is to allow a reliable diagnosis in complex lesions, such as small clusters of microcalcifications and especially architectural distortions. Surgery can be avoided for borderline cases if the lesion is completely removed and free margins are obtained in the pathology study. Therapeutic use is controversial and can be applied only in selected cases. PMID- 12111062 TI - Breast MRI for monitoring response of primary breast cancer to neo-adjuvant chemotherapy. AB - The objective of the present study was to monitor response to preoperative chemotherapy with breast MRI in patients with large breast cancer. Fifty-eight women in whom core biopsy had confirmed the presence of breast carcinoma underwent breast MRI prior to beginning chemotherapy and before surgical excision. In 24 cases patients underwent one or two additional examinations during chemotherapy to monitor their progress. Breast MRI included both T2 weighted spin-echo sequences and T1-weighted gradient-echo sequences before and 1, 2, 3, and 8 min after bolus injection of gadolinium-DTPA. Tumor size and the dynamic contrast medium uptake patterns of the respective carcinomas were evaluated and compared with the final histology findings. Based on their MR tomographic findings (change in tumor size and intensity of contrast media uptake), patients were assigned to groups with non-response (NR), partial response (PR), and complete response (CR). Based on MR tomographic findings, there were 12 patients in the NR group, 34 in the PR group, and 12 in the CR group. In NR group contrast medium uptake tended to increase or show no more than minimal decrease. Diagnostic accuracy for assigning patients to the NR group was 83.3% and to the PR group 82.4%. In patients whose tumors showed only slight response to chemotherapy, breast MRI proved very reliable in determining the size of the lesions. In patients whose tumors displayed significant response and in the CR group, the size of the residual tumor was underestimated in 8 of 12 cases. In 66.7% of patients in the CR group histology revealed residual tumor masses in areas up to 5 cm in diameter. During chemotherapy, intensity of contrast medium uptake decreased in 88.2% of patients with PR and in all patients with CR. Reliable determination of response was possible within 6 weeks following the initiation of chemotherapy. Breast MRI is suitable as a monitoring method. The determination of residual tumor size is unreliable in carcinomas exhibiting significant response to chemotherapy which may lead to false-negative results. The method may be employed for monitoring response to chemotherapy after 6 weeks. PMID- 12111063 TI - European quadricentric evaluation of a breast MR biopsy and localization device: technical improvements based on phase-I evaluation. AB - Our purpose was to report about technical success, problems and solutions, as experienced in a first multicentre study on MR-guided localisation or vacuum biopsy of breast lesions. The study was carried out at four European sites using a dedicated prototype breast biopsy device. Experiences with 49 scheduled localisation procedures and 188 vacuum biopsies are reported. Apart from 35 dropped indications, one localisation procedure and 9 vacuum biopsies were not possible (3 times space problems due to obesity, 2 times too strong compression, 3 times impaired access from medially, 2 times impaired access due to a metal bar). Problems due to too strong compression were recognised by repeat MR without compression. During the procedure problems leading to an uncertain result occurred in eight vacuum biopsies, two related to the procedure: one limited access, and one strong post-biopsy enhancement. Improvements after phase-I study concerned removal of the metal bar, development of an improved medial access, of a profile imitating the biopsy gun, optimisation of compression plates and improved software support. The partners agreed that the improvements answered all important technical problems. PMID- 12111064 TI - Whole-body computed tomography in polytrauma: techniques and management. AB - An interdisciplinary team should be involved in the diagnosis and management of severely injured patients. The adoption of criteria for starting treatment for multiple trauma avoids underestimation of seriousness of injury. These criteria are established by the circumstances of the accident, the patterns of trauma, and the vital findings. Basic diagnosis comprises a limited number of plain films in the trauma room, including supine chest, lateral cervical spine, and pelvis, and ultrasound of abdomen, pleura, and pericardium. Organ diagnosis using CT is complementary and depends on the clinical findings and findings from the basic investigations. We recommend spiral CT (skull base 2/2/4 mm, cerebrum 8/8/8 mm native) and after intravenous contrast medium thoracic (5/7.5/5 mm) and abdominal CT (8/12/8 mm). Image reconstruction of bony structures can be added. The CT and the trauma center should be in close proximity; time-consuming transfers must be avoided. If this is not possible, a CT can be integrated in the trauma room. Our hospital trauma registry contains over 2200 entries. A quality committee has been established and external quality control is implemented. PMID- 12111066 TI - Transabdominal sonography of the gastrointestinal tract. AB - Like other cross-sectional imaging methods, transabdominal sonography is increasingly used for evaluation of gastrointestinal diseases. The potentials and limitations of sonography in evaluation of the gastrointestinal tract are discussed. Transabdominal sonography proved to be of clinical value in assessment of appendicitis, diverticulitis, bowel obstruction, chronic inflammatory bowel diseases, intussusception and infantile hypertrophic pyloric stenosis. The sonographic morphology of the most common gastrointestinal diseases is discussed. In experienced hands sonography can be used as primary imaging in several gastrointestinal diseases. The gastrointestinal tract should be included in the sonographic examination of the abdomen, especially if symptoms could be related to the intestine. PMID- 12111065 TI - Clinical evaluation of digital radiography based on a large-area cesium iodide amorphous silicon flat-panel detector compared with screen-film radiography for skeletal system and abdomen. AB - The aim of this clinical study was to compare the image quality of digital radiography using the new digital Bucky system based on a flat-panel detector with that of a conventional screen-film system for the skeletal structure and the abdomen. Fifty patients were examined using digital radiography with a flat-panel detector and screen-film systems, 25 for the skeletal structures and 25 for the abdomen. Six radiologists judged each paired image acquired under the same exposure parameters concerning three observation items for the bone and six items for the abdomen. Digital radiographic images for the bone were evaluated to be similar to screen-film images at the mean of 42.2%, to be superior at 50.2%, and to be inferior at 7.6%. Digital radiographic images for the abdomen were judged to be similar to screen-film images at the mean of 43.4%, superior at 52.4%, and inferior at 4.2%; thus, digital radiographic images were estimated to be either similar as or superior to screen-film images at over 92% for the bone and abdomen. On the statistical analysis, digital radiographic images were also judged to be preferred significantly in the most items for the bone and abdomen. In conclusion, the image quality of digital radiography with a flat-panel detector was superior to that of a screen-film system under the same exposure parameters, suggesting that dose reduction is possible with digital radiography. PMID- 12111067 TI - CT findings in isolated ischemic proctosigmoiditis. AB - The purpose of our study was to describe the CT features of ischemic proctosigmoiditis in correlation with clinical, laboratory, endoscopic, and histopathologic findings. Our study included seven patients with isolated ischemic proctosigmoiditis. Patients were identified by a retrospective review of all histopathologic records of colonoscopic biopsies performed during a time period of 4 years. All patients presented with left lower abdominal quadrant pain, bloody stools, and leukocytosis, and four patients had fever at the time of presentation. Four of seven patients suffered from diarrhea, one of seven was constipated and two of seven had normal stool consistency. The CT examinations were reviewed by two authors by consensus and compared with clinical and histopathologic results as well as with the initial CT diagnosis. The CT showed a wall thickening confined to the rectum and sigmoid colon in seven of seven patients, stranding of the pararectal fat in four of seven, and stranding of the perisigmoidal fat in one of seven patients. There were no enlarged lymph nodes, but five of seven patients showed coexistent diverticulosis and in three of these patients CT findings were initially misinterpreted as sigmoid diverticulitis. Endoscopies and histopathologic analyses of endoscopic biopsies confirmed non transmural ischemic proctosigmoiditis in all patients. Isolated ischemic proctosigmoiditis often presents with unspecific CT features and potentially misleading clinical and laboratory findings. In an elderly patient or a patient with known cardiovascular risk factors the diagnosis of ischemic proctosigmoiditis should be considered when wall thickening confined to the rectum and sigmoid colon is seen that is associated with perirectal fat stranding. PMID- 12111068 TI - Clinical evaluation of further-developed MRCP sequences in comparison with standard MRCP sequences. AB - The purpose of this study was the comparison of technically improved single-shot magnetic resonance cholangiopancreatography (MRCP) sequences with standard single shot rapid acquisition with relaxation enhancement (RARE) and half-Fourier acquired single-shot turbo spin-echo (HASTE) sequences in evaluating the normal and abnormal biliary duct system. The bile duct system of 45 patients was prospectively investigated on a 1.5-T MRI system. The investigation was performed with RARE and HASTE MR cholangiography sequences with standard and high spatial resolutions, and with a delayed-echo half-Fourier RARE (HASTE) sequence. Findings of the improved MRCP sequences were compared with the standard MRCP sequences. The level of confidence in assessing the diagnosis was divided into five groups. The Wilcoxon signed-rank test at a level of p<0.05 was applied. In 15 patients no pathology was found. The MRCP showed stenoses of the bile duct system in 10 patients and choledocholithiasis and cholecystolithiasis in 16 patients. In 12 patients a dilatation of the bile duct system was found. Comparison of the low- and high spatial resolution sequences and the short and long TE times of the half Fourier RARE (HASTE) sequence revealed no statistically significant differences regarding accuracy of the examination. The diagnostic confidence level in assessing normal or pathological findings for the high-resolution RARE and half Fourier RARE (HASTE) was significantly better than for the standard sequences. For the delayed-echo half-Fourier RARE (HASTE) sequence no statistically significant difference was seen. The high-resolution RARE and half-Fourier RARE (HASTE) sequences had a higher confidence level, but there was no significant difference in diagnosis in terms of detection and assessment of pathological changes in the biliary duct system compared with standard sequences. PMID- 12111069 TI - Acute cholecystitis in high-risk patients: percutaneous cholecystostomy vs conservative treatment. AB - Our objective was to compare the effectiveness of percutaneous cholecystostomy (PC) vs conservative treatment (CO) in high-risk patients with acute cholecystitis. The study was randomized and comprised 123 high-risk patients with acute cholecystitis. All patients fulfilled the ultrasonographic criteria of acute inflammation and had an APACHE II score > or =12. Percutaneous cholecystostomy guided by US or CT was successful in 60 of 63 patients (95.2%) who comprised the PC group. Sixty patients were conservatively treated (CO group). One patient died after unsuccessful PC (1.6%). Resolution of symptoms occurred in 54 of 63 patients (86%). Eleven patients (17.5%) died either of ongoing sepsis (n=6) or severe underlying disease (n=5) within 30 days. Seven patients (11%) were operated on because of persisting symptoms (n=3), catheter dislodgment (n=3), or unsuccessful PC (n=1). Cholecystolithotripsy was performed in 5 patients (8%). Elective surgery was performed in 9 cases (14%). No further treatment was needed in 32 patients (51%). In the CO group, 52 patients (87%) fully recovered and 8 patients (13%) died of ongoing sepsis within 30 days. All successfully treated patients showed clinical improvement during the first 3 days of treatment. Percutaneous cholecystostomy in high-risk patients with acute cholecystitis did not decrease mortality in relation to conservative treatment. Percutaneous cholecystostomy might be suggested to patients not presenting clinical improvement following 3 days of conservative treatment, to critically ill intensive care unit patients, or to candidates for percutaneous cholecystolithotripsy. PMID- 12111070 TI - Current status of power Doppler and color Doppler sonography in the differential diagnosis of lymph node lesions. AB - The use of Power Doppler sonography in nodal diseases provides an improvement of early and noninvasive diagnosis of regional metastatic involvement. By using Power Doppler sonography it is possible to characterize lymph nodes as reactively enlarged, metastases, malignant lymphoma, tuberculosis and to study cervical cysts. The high diagnostic accuracy is based on perfusion-characteristics of these lymph nodes. Reactive lymph nodes show increased central perfusion of the hilum, whereas metastases tend to show increased peripheral perfusion. Affected lymph nodes in patients with by malignant lymphoma are highly perfused in the center but also peripheral. Power Doppler sonography is still not able to discriminate small (<8 mm) nonnecrotic metastasis or micrometastases from reactive lymph nodes. The purpose of this paper is to provide a summary of the current status of power Doppler and Doppler sonography in the differential diagnosis of lymph nodes. PMID- 12111071 TI - Differential diagnosis of lymphadenopathy: power Doppler vs color Doppler sonography. AB - Our objective was to compare color and power Doppler sonography of superficial lymph nodes. One hundred ninety-three lymph nodes in 161 patients were assessed by color and power Doppler sonography using standardized settings. We tested which modality displayed more intranodal vessels and checked if these differences would have altered the diagnosis. Additional vessels were seen by color Doppler sonography in 18 nodes and by power Doppler sonography in 58 nodes. Amongst those nodes were 15 nodes which showed no vascularization in color Doppler sonography and 23 nodes with only few intranodal flow signals; however, the better sensitivity of power Doppler sonography had no impact on the diagnosis in 42 of 58 nodes. Diagnostic confidence was increased in 7 nodes which showed normal vessels only in power Doppler sonography, although missing flow signals were defined as a benign finding. Pathological vessels were displayed only by power Doppler sonography in 9 nodes, but 6 of these 9 results proved to be false positive. Power Doppler sonography displays more intranodal flow signals than color Doppler sonography, but the diagnostic impact is low because of an increased risk of false-positive results. PMID- 12111072 TI - Spiral CT arthrography of the knee: technique and value in the assessment of internal derangement of the knee. AB - Computed tomography imaging has achieved excellent multiplanar capability and submillimeter spatial resolution due to the development of the spiral acquisition mode and multidetector row technology. Multidetector spiral CT arthrography (CTA) yields valuable information for the assessment of internal derangement of the joints. This article focuses on the value of spiral CTA of the knee in the assessment of the meniscus, anterior cruciate ligament, and hyaline cartilage lesions. Advantages and disadvantages of spiral CTA with respect to MR imaging are presented. PMID- 12111073 TI - Value of opposed-phase gradient-echo technique in distinguishing between benign and malignant vertebral lesions. AB - Our objective was to evaluate the possible role of opposed-phase gradient-echo (GRE) sequence in predicting the nature of vertebral lesions supposing that in the case of malignancy fat is completely replaced while in the case of benign lesion fat is still present. Eighty-six patients with vertebral lesions underwent MR examination at 0.5 T. The MR protocol included a T1-weighted spin-echo (SE) and an opposed-phase GRE using the same parameters (TR=280-320 ms, flip angle=90 degrees, slice thickness=3.5-4 mm, matrix=256x160-192, field of view=34-36 cm, no. of excitations=2-4) except for TE (10 ms in SE vs 7 ms in GRE) to obtain opposed-phased images. Qualitative (nature of lesion, detectability, degree of signal intensity (SI), marrow pattern) and quantitative (SI on opposed-phase GRE minus SI on T1-weighted SE minus SI ratio=SI on out-of-phase GRE images divided by SI on T1-weighted SE images) analysis were performed. The SI ratio values were analysed using Mann-Whitney rank-sum test and receiver operating characteristics (ROC) curve. Lesions resulted to be malignant in 45 and benign in 41 patients (23 biopsies, 20 MR follow-ups, 43 clinical and other imaging follow-ups). Based on visual inspection of opposed-phased images, visual SI was evaluated high in 38 (34 malignant, 34 benign), mild in 28 (9 malignant, 19 benign) and low in 20 (2 malignant, 18 benign) patients. Based on region-of-interest measurements, SI ratio values range was 0.36-6.2 (mean value=1.68+/-0.82) for malignant and 0.07 1.54 (mean value=0.77+/-0.44) for benign lesions. A cut-off value of 1.2 gave a sensitivity, specificity, accuracy, negative predictive value and positive predictive value, respectively, of 88.8, 80.49, 84.88, 86.4 and 83.33%. The ROC analysis of the SI ratio showed an area under ROC curve of 0.92 and a statistically significant difference between the two groups of lesions was observed ( p<0.01). The GRE opposed-phase sequence can help to predict the nature of a vertebral lesion. This fast and widely available technique together with morphological criteria can improve the accuracy of MRI. PMID- 12111074 TI - MR imaging assessment of clinical problems in rheumatoid arthritis. AB - Although MR imaging has been increasingly recognized as a useful tool in the diagnosis of early rheumatoid arthritis (RA) and in the assessment of disease activity, these applications have not yet been usually included in the routine management of this condition. Our goal is to review the current role of MRI in the everyday clinical management of patients with RA. The usefulness of MRI in the evaluation of articular and para-articular changes in specific locations, mainly the craniocervical region and the temporomandibular joint, are reviewed. Clinical problems derived from local extra-articular involvement, such as tenosynovitis, "rice-bodies" bursitis, and Baker's cyst rupture, are also described. Finally, we also review the value of MRI in evaluation of some complications of RA such as tendinous rupture, osteonecrosis, stress fracture, and septic arthritis/osteomyelitis. PMID- 12111075 TI - Craniopharyngiomas: identification of different semiological patterns with MRI. AB - Our objectives were to analyze different semiological patterns in craniopharyngiomas studied with CT and MR sequences. Retrospective study of 26 patients with confirmed craniopharyngiomas. All cases were examined with CT and MR imaging using a variety of pulse sequences (spin echo, inversion recovery, gradient echo in-phase and opposed-phase). The analyzed component patterns were classified as solid, calcium, proteic-like, cerebrospinal fluid (CSF)-like, hematic-like, and fatty patterns. The different patterns were related by means of contingency tables and the Fisher exact test and also to epidemiological findings and tumor size. A solid pole was detected in all patients, whereas a cystic component was present in 92.3% of the cases. Calcification was visualized in 65.3%, proteic-like in 53.8%, CSF-like in 23%, hematic-like in 19.2%, and fatty component in 15.3%. There were no statistical associations between patterns, with the exception that in no case did CSF-like and proteic-like patterns coexist ( P=0.004). Tumor size was related to components. Hematic-like (17.0+/-18.9 vs 3.9+/-2.6 mm, non-present vs present) and CSF-like (16.9+/-19.6 vs 6.5+/-4.0 mm) patterns were observed most frequently in smaller tumors, whereas larger tumors usually had proteic-like (5.9+/-5.4 vs 21.1+/-21.0 mm) and calcified (4.6+/-1.9 vs 19.1+/-19.9 mm) patterns. Computed tomography and a combination of different MR images frequently allow the detection of different semiological patterns in these tumors. Semiological patterns were correlated only to tumor size. PMID- 12111076 TI - Thalamic involvement in a patient with kernicterus. AB - We report the MR imaging findings of a 16-month-old boy with dyskinetic cerebral palsy resulting from kernicterus. T2-weighted images showed symmetric bilateral hyperintensity in the thalamus in addition to the globus pallidus. PMID- 12111077 TI - Cavernous angioma associated with ipsilateral hippocampal sclerosis. AB - We report two cases with extratemporal cavernous angioma (CA) and coexisting ipsilateral hippocampal sclerosis. Classically dual pathology is defined as the association of hippocampal sclerosis with an extrahippocampal lesion. Subtle changes in hippocampus might be overlooked in the presence of an unequivocal extrahippocampal abnormality. Seizure outcome after epilepsy surgery in cases with dual pathology is less favourable if only one of the lesions is removed. Dual pathology must always be considered in diagnostic imaging of patients with intractable epilepsy and CA. PMID- 12111078 TI - Hyperechogenicity of medial gastrocnemial veins during ultrasound scanning of the calf in sitting patients: a normal variant. AB - The goal of this study was to evaluate the prevalence and significance of the high echogenicity of blood frequently encountered in medial gastrocnemial veins (MGVs) during venous ultrasound scanning of the calf in sitting patients. Using a commercially available multi-frequency linear probe an experienced radiologist prospectively analysed a series of 97 legs from 59 patients referred to the ultrasound department for various clinical reasons; echogenic ambulatory outpatients being free of calf venous thrombosis were only considered. Hyperechogenicity of blood in MGVs was visible in 67 of 97 calfs (69%); it was discrete or moderate in 23 calfs (23.7%) and intense in 44 calfs (45,36%); 97% of these 67 calfs contained large MGVs whose diameter was at least twice that of the other muscular veins. The "washing out" manoeuvre applied to these veins by direct compression with the probe was effective to prove their normal permeability; after releasing of compression, hyperechogenicity was spontaneously restored in a mean time of 47.45+/-16.85 s. Because of a very low flow and a diameter commonly larger than other muscular veins, MGVs probably frequently reach a sufficiently low shear rate to promote aggregation causing echogenicity of blood when patients are examined in a sitting position. For this reason, hyperechogenicity of MGVs, instead of being a manifestation of local stasis, represent a reversible physiological normal variant. Nevertheless, the careful dynamic compression of these veins during ultrasound scanning is necessary to distinguish this normal variant from deep venous thrombosis. PMID- 12111079 TI - Double bubble sign. PMID- 12111080 TI - The Trier mummy Pai-es-tjau-em-aui-nu: radiological and histological findings. AB - The ancient Egyptian mummy Pai-es-tjau-em-aui-nu, now on exhibition in the Rheinische Landesmuseum in Trier, Germany, was examined by conventional X-ray radiography, computed tomography, and digital fluoroscopy. In addition, some tissues were biopsied for further histologic identification. Along with some representative images, the peculiarities of the Trier mummy concerning the mummification process are presented. PMID- 12111081 TI - Nonspecific interstitial pneumonia as pulmonary involvement of primary Sjogren's syndrome. AB - The pathologic patterns of lung involvement in nine patients with Sjogren's syndrome (SjS) are evaluated. The patients consisted of three males and six females, with a median age of 59 years. The SjS was diagnosed according to the criteria of the First International Seminar on SjS. In all patients, high resolution computed radiographic scanning (HRCT) of the lungs was performed, and apparent honeycomb or microhoneycomb formation was observed in six patients. Pathologically, six patients were diagnosed with usual interstitial pneumonia (UIP), and three were diagnosed with nonspecific interstitial pneumonia/fibrosis (NSIP) (group II). There were no apparent honeycomb formations on HRCT in patients diagnosed with NSIP. In conclusion, NSIP is also a possible histologic classification of interstitial pneumonia associated with SjS. PMID- 12111082 TI - Trace elements and antioxidant enzymes in Behcet's disease. AB - Free oxygen radicals and insufficiency of antioxidant enzymes have been implicated in the pathogenesis of Behcet's disease (BD). Trace elements function as cofactors to antioxidant enzymes. The antioxidant system and trace elements were investigated in many different studies, including BD, but these subjects have not been investigated as a whole in these patients. The aim of the present study was to investigate the antioxidative system and trace elements in BD to contribute to the knowledge of pathogenesis and treatment of this disease. We examined glutathione peroxidase (GSH-Px) and superoxide dismutase (SOD) activities together with selenium (Se), copper (Cu), zinc (Zn), manganese (Mn), and iron (Fe) levels in plasma and erythrocytes of 50 patients with BD and 30 healthy controls. It was found that in patients with BD, erythrocyte GSH-Px and SOD activities and erythrocyte Se, plasma Fe, Mn, and Zn levels were significantly lower than those of controls and that plasma Cu, erythrocyte Zn, and Mn levels were significantly higher in patients with BD. Insufficient antioxidant enzyme activities were observed in patients with BD. The mechanism(s) of this phenomenon is not clear. Therefore, supplementation with trace elements involved in the antioxidative processes may increase scavenger enzyme activities, and consequently, an improvement in clinical symptoms may be expected. PMID- 12111084 TI - Effectiveness of a home-based exercise therapy and walking program on osteoarthritis of the knee. AB - Osteoarthritis (OA) of the knee is a very common rheumatological disease, and there are various treatment modalities for it. The aim of this study was to investigate the effects of home-based exercise and walking programs in the treatment of OA. A total of 90 patients with knee osteoarthritis were included. Their ages ranged between 48 and 71 years. The patients were separated into three groups. None of them had practiced a daily simple exercise program during the previous year. Group 1 ( n=30) was given a home-based exercise program. Group 2 ( n=30) had regular a walking program three times per week, starting with 10-min duration. Group 3 ( n=30) was accepted as the control group. Patients were assessed according to pain, functional capacity, and quality of life parameters. Pain was evaluated by the Western Ontario McMaster osteoarthritis index (WOMAC) of pain score and visual analogue scale (VAS). Functional capacity was measured by WOMAC physical function index. Quality of life was assessed by the Nottingham Health Profile questionnaire (NHP). All groups continued the program for 3 months. At the end of the therapy, the patients were called and 81 were accepted to come to the hospital. Although WOMAC pain and physical functional scores and VAS scores were statistically lower in both groups than in the control group ( P<0.001), the difference between groups 1 and 2 was not statistically significant ( P>0.05). But the result of the NHP showed a statistically significant improvement in the walking group when compared to the home-based exercise and control groups ( P<0.001). As a result, we conclude that a simple home-based exercise therapy and a regular walking program are effective in treating the symptoms of OA. PMID- 12111083 TI - Expression of caspase-1 in synovial membrane-like interface tissue around loosened hip prostheses. AB - Caspase-1 expression in synovial membrane-like interface tissue (SMLIT) around loosened hip prostheses and osteoarthritic synovial samples was studied. Caspase 1 mRNA was found in SMLIT and synovial tissue. There is no difference in the copy numbers of caspase-1 mRNA between these samples. Both precursor and active forms of caspase-1 proteins appeared in these samples, but the number of positive cells was higher in SMLIT than in synovial tissue. Double labeling revealed that most caspase-1-positive cells were macrophages and fibroblasts. In the lining-like layers and deep stroma of SMLIT, many cells were double positive for active caspase-1 and interleukin-1 beta (IL-1beta). In contrast, the number of active caspase-1/IL-18 double-positive cells was very low. We conclude that caspase-1 synthesis is increased in SMLIT. Caspase-1 can be involved in implant loosening by processing IL-1beta precursor into its mature form, which is a potent osteoclast-activating factor and a major proinflammatory mediator. PMID- 12111086 TI - Does fibromyalgia affect the outcomes of local steroid treatment in patients with carpal tunnel syndrome? AB - Carpal tunnel syndrome (CTS) and fibromyalgia (FM) are common diseases in adult women. The aim of this study was to investigate the effect of FM on the outcome of local steroid treatment in patients with CTS. Forty-eight female patients with CTS and 26 female patients with CTS who also met the 1990 American College of Rheumatology (ACR) criteria for FM were enrolled the study. All patients underwent single steroid injections into the carpal tunnel. Response to injection was measured using Boston Questionnaire (BQ) and electrophysiological tests. The BQ scores and electrophysiological findings showed significant improvement 3 months after the treatment in both groups ( P<0.001). However, less improvement in BQ symptom severity scores was observed in the FM group than the other group ( P<0.05). Bilateral CTS was also more common in the FM group ( P<0.05). The present data suggest that FM must be kept in mind in patients with CTS giving poor response to local steroid treatment. PMID- 12111085 TI - The chemoattraction of lymphocytes by rheumatoid arthritis - synovial fluid is not dependent on the chemokine receptor CCR5. AB - OBJECTIVE: The objective was to study the potential role of the chemokine receptor CCR5 in the chemoattraction of lymphocytes by rheumatoid arthritis synovial fluid (RA-SF). METHODS: The expression of the CCR5 receptor was studied by flow cytometry. Chemotaxis of peripheral blood lymphocytes in response to RA SF was analyzed on transmigration chambers. Chemotaxis of immortalized lymphocytes from individuals homozygous for the Delta32 deletion of the CCR5 gene (CCR5-/-) was analyzed. The effect of a neutralizing anti-CCR5 antibody on the migration of CCR5+/+ cells was also studied. RESULTS: We confirmed an increase in the proportion of CCR5-expressing lymphocytes in RA-SF and a preferential migration of CCR5+ lymphocytes toward RA-SF in vitro. CCR5-/- lymphocytes showed decreased chemotactic responses to the chemokine MIP-1beta but not to RA-SF. The chemotactic responses of CCR5+/+ lymphocytes to RA-SF were not modified by anti CCR5 neutralizing antibody. CONCLUSIONS: We confirm a preferential accumulation of CCR5-expressing lymphocytes into RA-SF. However, the chemotactic responses of lymphocytes to RA-SF were not dependent on a functional CCR5 receptor, suggesting that CCR5 is a marker of a lymphocyte subset rather than a specific mediator of chemotactic responses to chemokines in RA-SF. PMID- 12111087 TI - Type II autosomal dominant osteopetrosis. AB - Two principal types of osteopetrosis have been distinguished. One is the dominantly inherited, relatively benign condition which is often detected radiologically in asymptomatic adults. A second type is the recessive, lethal, malignant form. Autosomal dominant osteopetrosis (ADO) has two distinct radiological subtypes known as types I and II. We report here a 23-year-old patient with ADO type II. Radiographic investigations of a skeletal survey showed generalised osteosclerosis with thickened cortex. Magnetic resonance imaging (MRI) scan disclosed osteosclerosis in superior and inferior portions of the vertebral bodies which produced a 'sandwich' appearance. The 'bone-within-bone' appearance was seen in the ileum of the patient. The vertebral bone density was found markedly elevated. The carbonic anhydrase II level was found to be normal. We discuss here the genetic etiology of this disorder. PMID- 12111088 TI - Primary antiphospholipid syndrome presenting with abdominal angina and splenic infarction. AB - The antiphospholipid syndrome is an autoimmune hypercoagulability syndrome in which a wide variety of thromboembolic diseases may occur. Gastrointestinal manifestations associated with vascular occlusion include Budd-Chiari syndrome, hepatic and splenic infarction, pancreatitis, omental and intestinal infarction, and esophageal variceal bleeding due to portal vein thrombosis, but chronic mesenteric ischemia associated with mesenteric arterial thrombosis is very rare in this syndrome. We experienced a female patient with primary antiphospholipid syndrome with abdominal angina and splenic infarction associated with celiac trunk and mesenteric arterial thromboses. This is the first report describing chronic mesenteric ischemia and splenic infarction in a patient with primary antiphospholipid syndrome. PMID- 12111089 TI - Thrombotic thrombocytopenic purpura associated with mixed connective tissue disease. AB - We report a male case of thrombotic microangiopathy with mixed connective tissue disease (MCTD). Thrombocytopenia and hemolytic anemia developed despite steroid treatment for acute interstitial pneumonitis. The patient became confused and eventually developed coma. Diffusion-weighted MRI revealed high intensity at the brainstem, frontal lobes, basal nuclei, and insula, findings compatible with multiple brain edema due to microthrombosis. Despite the treatment of successive plasma exchange and steroid pulse therapy, he eventually died by multiple organ failure. This rare case suggested that diffusion-weighted MRI is very sensitive at detecting early alterations of thrombotic thrombocytopenic purpura (TTP) induced ischemic lesions in the brain. Neuropsychiatric symptoms due to thrombotic microangiopathy could be some of the fatal complications in MCTD patients. PMID- 12111090 TI - C4 deficiency state in antiphospholipid antibody-related recurrent preeclampsia evolving into systemic lupus erythematosus. AB - The case of an apparent healthy woman who developed recurrent preeclampsia with antiphospholipid antibodies and evolved towards systemic lupus erythematosus during her last pregnancy is presented. The diagnostic dilemma between lupus renal flare and toxemia is discussed. The potential role of immunological alterations, such as complement genetic deficiencies, in women with primary antiphospholipid syndrome who develop systemic lupus erythematosus is also discussed. PMID- 12111091 TI - A case of Waldenstroem's disease with a monoclonal IgM antiphospholipid antibody. AB - The antiphospholipid syndrome (APS) was described in 1983 as a clinical entity characterized by venous and arterial thrombosis, thrombocytopenia, and recurrent fetal loss. The serological markers of APS are antiphospholipid antibodies (APLA) directed mainly against anionic phospholipids, usually cardiolipin but also phosphatidylserine. Some APLA exhibit lupus anticoagulant activity. Monoclonal gammopathy sometimes occurs with the presence of autoantibodies. In this paper, we describe a patient with the diagnosis of immunocytoma with an IgM, kappa paraprotein with apparent specificity against anionic phospholipids, and lupus anticoagulant activity, but no clinical signs of APS. We describe in this patient the presence of a high titer of monoclonal APLA, which does apparently not induce the clinical symptoms of APS. This might be indicative for the presence of pathogenic and nonpathogenic antiphospholipid antibodies. PMID- 12111093 TI - Septins: a ring to part mother and daughter. AB - The septins are well conserved GTPases found in animals and fungi. In yeast, they are required for the formation of 10-nm filaments, with which they co-localize at the bud neck. Therefore, septins have been proposed to be components of the neck filaments and to have polymerization properties. In support of this hypothesis, septin complexes purified from yeast and flies form filaments in vitro. However, recent studies have questioned the relevance of septin filament formation for septin function. Particularly, septin polymerization may not be required for their function in cytokinesis. New septin functions have also been recently uncovered: in budding yeast, the septin ring is required for the maintenance of cell polarity. It forms a cortical barrier that prevents lateral diffusion of membrane-associated proteins through the bud neck. Here, we review the most recent functional and biochemical data, to discuss whether there is a link between septin polymerization properties and septin function. PMID- 12111094 TI - The URH1 uridine ribohydrolase of Saccharomyces cerevisiae. AB - In the yeast Saccharomyces cerevisiae, uridine ribohydrolase activity is important for recycling, via the salvage pathway, pyrimidine deoxy- and ribonucleosides into uracil required for the growth of strains lacking the de novo pyrimidine synthesis pathway. We have shown that not only uridine and cytidine, but also 5-fluorouridine, 5-fluorocytidine and deoxycytidine are substrates for this enzyme. We identified, cloned and characterized the corresponding URH1 gene and its physiological function was determined by the measurement of metabolic fluxes in several mutants impaired in the pyrimidine salvage pathway. Sequence comparison revealed strong homology between Urh1p and the inosine/uridine-preferring nucleosidase and inosine/adenosine/guanosine nucleoside hydrolase proteins from the parasitic organisms Crithidia fasciculata and Trypanosoma brucei brucei. Moreover, the Asp and His residues in the putative active site were conserved. Site-directed mutagenesis demonstrated that the conserved His residue is involved in catalysis. These results allow us to speculate that the structure and catalytic mechanism of Urh1p are similar to the inosine/uridine nucleoside hydrolase from C. fasciculata. PMID- 12111095 TI - Immunosuppressant-like effects of phenylbutyrate on growth inhibition of Saccharomyces cerevisiae. AB - Phenylbutyrate (4-phenylbutyric acid; PB) and its metabolite, phenylacetate, are effective anti-neoplastic agents in tissue culture and have shown promise in clinical trials for a variety of neoplasms. PB is a drug of remarkably low toxicity that acts in vitro as a differentiating agent, causing reversion of the transformed phenotype by an unknown mechanism. We attempted to identify the cellular target(s) for PB using Saccharomyces as a model. PB inhibits growth of yeast on rich medium at concentrations of 0.1-1.0 mM, concentrations similar to plasma concentrations observed in human trials. Yeast cells treated with 1 mM PB remain over 90% viable for 24 h. PB inhibits tryptophan uptake, and resistance to PB can be conferred by tryptophan prototrophy, by supplementing tryptophan auxotrophs with the high levels of tryptophan, by overexpression of the aromatic amino acid permeases Tat1p or Tat2p, and by disruption of TAT1. Since tryptophan auxotrophy and transport influences resistance to PB, phytosphingosine, and the immunosuppressant FK506, these drugs might affect the same pathway. We isolated and characterized a mutant resistant to 1 mM PB and identified the mutant as bul1. A chromosomal BUL1 deletion displayed all phenotypes shown by the PB resistant mutant. PMID- 12111096 TI - Interaction between mating-type proteins from the homothallic fungus Sordaria macrospora. AB - Mating-type genes control sexual development in ascomycetes. Little is known about their function in homothallic species, which are self-fertile and do not require a mating partner for sexual reproduction. The function of mating-type genes in the homothallic fungus Sordaria macrospora was assayed using a yeast system in order to find properties typical of eukaryotic transcription factors. We were able to demonstrate that the mating-type proteins SMTA-1 and SMTa-1 have domains capable of activating transcription of yeast reporter genes. Two-hybrid analysis for heterodimerization and homodimerization revealed the ability of SMTA 1 to interact with SMTa-1 and vice versa. These two proteins are encoded by different mating types in the related heterothallic species Neurospora crassa. The interaction between SMTA-1 and SMTa-1 was defined by experiments with truncated versions of SMTA-1 and in vitro by means of protein cross-linking. Moreover, we gained evidence for homodimerization of SMTA-1. Possible functions of mating-type proteins in the homothallic ascomycete S. macrospora are discussed. PMID- 12111097 TI - Functional expression and cellular localization of cercosporin-resistance proteins fused with the GFP in Cercospora nicotianae. AB - The Cercospora nicotianae pdx1 and crg1 genes were previously identified as genes required for resistance to the singlet oxygen ((1)O(2))-generating toxin cercosporin. The pdx1 gene has subsequently been shown to be required for pyridoxine biosynthesis, but both the precise biochemical function of the PDX1 protein and the function of the CRG1 protein remain undefined, as both sequences lack defined enzymatic domains or cofactor-binding sites. The gfp gene encoding green fluorescent protein was translationally fused with pdx1 and crg1. Transformation of these constructs into strains mutant in these respective genes resulted in green-fluorescent transformants complemented for the mutant phenotype. Microscopic studies revealed that in transformants transformed with gfp alone, fluorescence was distributed evenly throughout the cytoplasm and excluded from the vacuoles. Expression of PDX1::GFP either under the constitutive Aspergillus nidulans gpdA promoter or its own native promoter was visualized as distinct fluorescent circular structures in the cytoplasm, suggesting that PDX1::GFP was probably localized in the intracellular vesicles. Expression of CRG1 fused with GFP at either its N- or C-terminus resulted in low green fluorescence, compared with that of GFP alone or PDX1::GFP. The green fluorescence of either of the CRG1::GFP fusion proteins was barely observable in transformants and was generally seen as a few scattered regions of fluorescence in the hyphae. Southern blot analysis indicated multiple copies of the constructs were integrated into the fungal genome. Northern analysis revealed that pdx1:: gfp and crg1:: gfp were each expressed as an intact transcriptional unit. Cell fractionation followed by immunoblotting against a GFP antibody showed that GFP alone and PDX1::GFP were detected exclusively in the cytoplasmic fraction. The two CRG1::GFP proteins were barely detected in the cytoplasmic fraction and not at all from the membrane fraction, a result inconsistent with microscopic observation and computer sequence analysis, which suggests that CRG1 is a membrane protein. PMID- 12111098 TI - Cloning and genetic analysis of subtilases in sapstaining fungi. AB - In order to assess the genetic variance of a group of homologous subtilases in sapstaining fungi, molecular techniques were employed. First, PCR screening with degenerate primers and dot-blot analyses were used to screen 31 different isolates, representing nine species, of sapstaining fungi for the presence of subtilase-like sequences. Restriction fragment length polymorphism PCR and sequence analysis techniques were then used to determine the inter- and intraspecies variation of these genes. A labelled chemiluminescent probe was then used to screen an Ophiostoma floccosum 387N genomic library for subtilase genes. Over ten positive clones were found and one was subcloned and sequenced. Randomly amplified cDNA ends PCR techniques were then employed to obtain full-length subitilase gene sequence information from isolates of four different species. The obtained sequences were found to be homologous with other fungal subtilases and common structural features of the inferred proteins with proteinase K were apparent. Southern blot analysis was used to verify and determine the copy number of the subtilase genes in these fungi. In the work presented here, it was found that all of the isolates tested seemed to contain some sort of subtilase gene sequence and that there was inter- and intraspecies variation in the number and type of subtilase genes present. The data indicated that three distinct groups of subtilase genes are present in sapstaining fungi. However, individual isolates were found to contain only one or two of these gene types. PMID- 12111099 TI - Sequences upstream of the YRTA core region are essential for transcription of the tobacco atpB NEP promoter in chloroplasts in vivo. AB - Transcription of the plastid atpB gene is accomplished by plastid-encoded (PEP) and nuclear-encoded (NEP) RNA polymerases. In contrast to NEP promoters of many other plastid genes, the tobacco atpB NEP promoter exhibits robust activity in chloroplasts in vivo. Previously, in vitro transcription assays using extracts from non-photosynthetic cells identified two elements required for full atpB NEP promoter activity, a core sequence and an upstream GAA box. Of these, only the core sequence containing the motif YRTA is conserved in the majority of NEP promoters. We used plastid transformation to examine the requirements for atpB NEP promoter activity in chloroplasts. Our results demonstrate that sequences upstream of the core element are essential for promoter activity in vivo, and that transcription of the NEP promoter in chloroplasts is not dependent on activity from an overlapping PEP promoter. PMID- 12111100 TI - Mathematical modelling of oxygen transport to tissue. AB - The equations governing oxygen transport from blood to tissue are presented for a cylindrical tissue compartment, with blood flowing along a co-axial cylindrical capillary inside the tissue. These governing equations take account of: (i) the non-linear reactions between oxygen and haemoglobin in blood and between oxygen and myoglobin in tissue; (ii) diffusion of oxygen in both the axial and radial directions; and (iii) convection of haemoglobin and plasma in the capillary. A non-dimensional analysis is carried out to assess some assumptions made in previous studies. It is predicted that: (i) there is a boundary layer for oxygen partial pressure but not for haemoglobin or myoglobin oxygen saturation close to the inflow boundary in the capillary; (ii) axial diffusion may not be neglected everywhere in the model; (iii) the reaction between oxygen and both haemoglobin and myoglobin may be assumed to be instantaneous in nearly all cases; and (iv) the effect of myoglobin is only significant for tissue with a low oxygen partial pressure. These predictions are validated by solving the full equations numerically and are then interpreted physically. PMID- 12111101 TI - Minimal number of generations out of polymorphism in the one-locus two-allele model with unpredictable fertilities. AB - Fertilities varying in time in an unpredictable manner raise the question of the maintenance of polymorphism, and the subsequent question of the minimal total number of generations spent by the system in impoverished polymorphism. In the one-locus two-allele model, level surfaces of this minimal time out of a given set K of constraints defining polymorphism are delineated. The surface of null minimal time is also the largest set of genotype frequencies from which there exists at least one route remaining in K forever. The influence of the range of admissible fertilities clarifies the trade-off between homozygotes and heterozygotes. Notably, ranges of fertility forbidding a system leaving K to ever return to it are determined. Generally, keeping a chance to regain polymorphism demands a sufficiently relatively high fertility for matings involving heterozygotes. PMID- 12111102 TI - Invasion dynamics and attractor inheritance. AB - We study the dynamics of a population of residents that is being invaded by an initially rare mutant. We show that under relatively mild conditions the sum of the mutant and resident population sizes stays arbitrarily close to the initial attractor of the monomorphic resident population whenever the mutant has a strategy sufficiently similar to that of the resident. For stochastic systems we show that the probability density of the sum of the mutant and resident population sizes stays arbitrarily close to the stationary probability density of the monomorphic resident population. Attractor switching, evolutionary suicide as well as most cases of "the resident strikes back" in systems with multiple attractors are possible only near a bifurcation point in the strategy space where the resident attractor undergoes a discontinuous change. Away from such points, when the mutant takes over the population from the resident and hence becomes the new resident itself, the population stays on the same attractor. In other words, the new resident "inherits" the attractor from its predecessor, the former resident. PMID- 12111103 TI - The general mixing of addicts and needles in a variable-infectivity needle sharing environment. AB - In this paper we develop and analyse a model for the spread of HIV/AIDS amongst a population of injecting drug users. The model we discuss focuses on the transmission of HIV through the sharing of contaminated drug injection equipment and in particular we examine the mixing of addicts and needles when the AIDS incubation period is divided into three distinct infectious stages. The impact of this assumption is to greatly increase the complexity of the HIV transmission mechanism. We begin the paper with a brief literature review followed by the derivation of a model which incorporates three classes of infectious addicts and three classes of infectious needles and where a general probability structure is used to represent the interaction of addicts and needles of varying levels of infectivity. We find that if the basic reproductive number is less than or equal to unity then there exists a globally stable disease free equilibrium. The model possesses an endemic equilibrium solution if the basic reproductive number exceeds unity. We then conduct a brief simulation study of our model. We find that the spread of disease is heavily influenced by the way addicts and needles of different levels of infectivity interact. PMID- 12111104 TI - A phase I study of suramin with once- or twice-monthly dosing in patients with advanced cancer. AB - PURPOSE: The optimal schedule of administration of suramin has not been well defined. The purpose of this study was to determine the maximum tolerated dose and toxicities of suramin when administered using a fixed dosing scheme on a once or twice-monthly schedule. METHODS: A total of 40 patients were treated on this phase I dose-escalation study employing a once-monthly (day 1 of each 28-day cycle) and a twice-monthly (days 1 and 8 of each 28-day cycle) schedule. RESULTS: The most common dose-limiting events included fatigue, neuropathy, and anorexia. We identified the 1440 mg/m(2) dose level to be the maximal tolerated dose for both schedules, with 83% of patients developing dose-limiting toxicity (DLT) on the twice-monthly schedule, and 67% developing DLT on the monthly schedule. At the 1200 mg/m(2) dose level, only 25% developed DLT on the twice-monthly schedule and 33% developed DLT on the monthly schedule. Trough suramin levels gradually increased with higher dose levels but fell well below the putative toxic concentration of 350 microg/ml. CONCLUSION: Suramin can be safely administered using a monthly schedule without using plasma concentrations to guide dosing. PMID- 12111105 TI - Phase II study of liposomal annamycin in the treatment of doxorubicin-resistant breast cancer. AB - Liposomal annamycin (L-AN) has shown antitumor activity in preclinical studies. It may selectively target tumors and bypass MDR-1 resistance. A total of 13 women with doxorubicin-resistant breast cancer were treated on this phase II study. The median number of prior chemotherapy regimens was two, and six patients had two or more organ sites of involvement. L-AN was administered at 190-250 mg/m(2) as an i.v. infusion over 1-2 h every 3 weeks. No responses were observed. Of the 13 patients, 12 had clear deterioration and new tumor growth after one or two courses. The 13th patient had prolonged grade 2 thrombocytopenia after one course, and was taken off study when the lung metastases increased 62 days after treatment. L-AN at this dose and on this schedule had no detectable antitumor activity in patients with doxorubicin-resistant metastatic breast cancer. PMID- 12111106 TI - Rapid biotransformation of satraplatin by human red blood cells in vitro. AB - PURPOSE: Satraplatin is an orally administered platinum complex that has demonstrated clinical activity and manageable toxicity in phase II trials. The presence of several different platinum-containing species and very little intact parent drug in the systemic circulation indicates extensive biotransformation of satraplatin in vivo. To investigate the basis for the biotransformation of satraplatin, studies were carried out into the stability of the drug in whole blood and various other biological fluids in vitro. METHODS: Concentrations of satraplatin and platinum-containing biotransformation products in incubation fluids were measured using high-performance liquid chromatography-inductively coupled plasma mass spectrometry (HPLC-ICPMS). The fate of satraplatin-derived platinum in whole blood in vitro was determined by analysis of blood fractions for platinum by ICPMS. RESULTS: In fresh human whole blood in vitro, satraplatin concentrations fell very rapidly, resulting in a half-life for the disappearance of the drug of only 6.3 min (95% CI, 5.9 to 6.7 min). After the addition of drug to red blood cells that had been prepared from whole blood and suspended in 0.9% NaCl, satraplatin also disappeared very rapidly. Satraplatin was much more stable in fresh human plasma (t(1/2) 5.3 h) and fully supplemented cell culture medium (t(1/2) 22 h). Two new platinum-containing species appeared on HPLC-ICPMS platinum chromatograms of methanol extracts of plasma after the addition of the drug to whole blood. Their identities were assigned as the platinum(II) complex known as JM118 and a platinated protein with similar electrophoretic mobility to that of serum albumin. During the incubation of satraplatin in blood, platinum associated with red blood cells at an accumulation half-life of 9.5 min (95% CI, 7.1 to 14.2 min). At equilibrium, 62% of the added platinum was associated with red blood cells in a form that was not exchangeable in methanol or 0.9% NaCl. CONCLUSIONS: The rapid disappearance of satraplatin from human blood in vitro depends upon the presence of red blood cells. Generation of JM118 and irreversibly bound membrane- and protein-associated platinum indicates that satraplatin undergoes rapid biotransformation in whole blood. PMID- 12111107 TI - A population analysis of the pharmacokinetics of Cremophor EL using nonlinear mixed-effect modelling. AB - PURPOSE: The purpose of this study was to develop a population pharmacokinetic model for Cremophor EL used as a formulation vehicle for paclitaxel. METHODS: Plasma concentration-time data from 70 patients (85 courses) treated with paclitaxel dissolved in Cremophor EL were used. The nonlinear mixed-effect modelling (NONMEM) program was used for the population pharmacokinetic analysis. The influence of patient characteristics on the pharmacokinetics of Cremophor EL was determined. The stability of the final model was evaluated using bootstrapping. RESULTS: The data were optimally fitted to a three-compartment model with Michaelis-Menten elimination from the central compartment. The following pharmacokinetic parameters were estimated: volume of the central compartment (V1=2.59 l), volumes of two peripheral compartments (V2=1.81 l, V3=1.61 l), intercompartmental clearance between central and peripheral compartments (Q12=1.44 l/h, Q13=0.155 l/h), maximal elimination rate (Vmax=0.193 ml/h), and concentration at half Vmax (Km=0.122 ml/l). Interindividual variability of the pharmacokinetic parameters was quantified for V1 (25%), V2 (36%) and Vmax (31%). Residual variability consisted of a combined additional (0.095 ml/l) and proportional error (7%). Gender, body surface area and performance status according to the World Health Organization were significantly correlated with V1, V2 and Vmax, respectively ( P<0.0001). The median parameter estimates of 1000 bootstrap samples were in accordance with those obtained with the original data set, indicating the validity of the population model. CONCLUSIONS: The population model was able to adequately describe the pharmacokinetic parameters and influence of covariates on the pharmacokinetics of Cremophor EL. This model can be used when studying the relationship between the pharmacokinetics and toxicity of Cremophor EL, and the drug's influence on the pharmacokinetics of paclitaxel. PMID- 12111108 TI - Pharmacokinetic study of S-1, a novel oral fluorouracil antitumor agent in animal model and in patients with impaired renal function. AB - PURPOSE: S-1 is a novel oral fluorouracil antitumor drug that combines tegafur (FT), 5-chloro-2,4-dihydroxypyridine (CDHP), which inhibits dihydropyrimidine dehydrogenase (DPD), and potassium oxonate (Oxo). As 50% of CDHP is excreted in the urine, renal dysfunction may directly affect the DPD inhibitory effect and lead to increased 5-fluorouracil (5-FU) concentrations. We sought to determine the influence of impaired renal function on the pharmacokinetics of S-1 in an animal model and in patients with gastric cancer. METHODS: An experimental renal failure model induced by cisplatin was developed in rabbits, and plasma concentrations of FT, 5-FU, CDHP and Oxo were determined after S-1 injection. Four patients with various degrees of renal impairment with unresectable gastric cancer were recruited to the study, and the pharmacokinetics in these four patients were analyzed following single and consecutive S-1 administrations. RESULTS: In experimental renal failure, plasma clearance of CDHP and 5-FU was retarded corresponding to the degree of renal impairment and there was a close correlation between creatinine clearance (CLcr) and plasma CDHP and 5-FU clearance. In the single administration study, half standard dose was used in three patients (CLcr > or = 50 ml/min) and one-third in the other (CLcr <50 ml/min). In patients with CLcr more than 75 ml/min, C(max), T(max), AUC((0 infinity)), and T(1/2) of 5-FU and CDHP were not different between single and consecutive administrations. In contrast, in patients with mild and moderate renal dysfunction (CLcr 55 and 36 ml/min, respectively), the T(1/2) values of CDHP with consecutive administrations (7.6 and 15.3 h, respectively) were longer than the values with single administration (4.6 and 8.2 h, respectively). The T(1/2) of 5-FU was 5.7 h with single administration and 8.5 h with consecutive administration in patients with moderate renal impairment. The AUC((0-infinity)) of 5-FU with consecutive administrations (3089.7 ng.h/ml) was far greater than with single administration (430.4 ng.h/ml). There was also a strong correlation between CLcr and plasma CDHP clearance. Based on the pharmacokinetics following multiple consecutive administrations, S-1 administration resulted in no severe adverse reactions in any of the four patients. CONCLUSIONS: CDHP clearance was prolonged in the presence of renal impairment, leading to a delayed T(1/2), and high AUC of 5-FU. These findings demonstrate that administration of S-1 to patients with impaired renal function may need individualized dosing and pharmacokinetic monitoring. PMID- 12111109 TI - Leucocyte versus erythrocyte thioguanine nucleotide concentrations in children taking thiopurines for acute lymphoblastic leukaemia. AB - PURPOSE: The aim of this study was to compare leucocyte and erythrocyte thioguanine nucleotide (TGN) cytotoxic metabolite concentrations in children with lymphoblastic leukaemia taking mercaptopurine (MP) or thioguanine (TG) as part of their long-term remission maintenance chemotherapy. METHODS: Ten consecutive children treated on the MRC ALL97 protocol were studied. Six were randomized to TG and four to MP. Leucocyte and erythrocyte thiopurine nucleotide metabolites were measured after the children had been titrated to the standard thiopurine protocol dose, or higher. RESULTS: Children taking TG accumulated significantly higher erythrocyte TGN concentrations than those taking MP (median difference 1171 pmol/8 x 10(8) erythrocytes, 95% CI 766 to 2169, P<0.02), but there was no significant difference in the concentration range of leucocyte TGNs generated from TG or MP. In those children taking TG, median TGN concentrations were 5142 pmol/8 x 10(8) leucocytes and 1472 pmol/8 x 10(8) erythrocytes (3.5-fold difference, median difference 3390 pmol/8 x 10(8) cells, 95% CI 1559 to 7695, P=0.005), compared to 5422 pmol/8 x 10(8) leucocytes and 261 pmol/8 x 10(8) erythrocytes (20-fold difference, median difference 5054 pmol/8 x 10(8) cells, 95% CI 2281 to 6328, P=0.03) in those taking MP. CONCLUSIONS: Despite the accumulation of significantly higher erythrocyte TGN concentrations for TG compared with MP, the accumulation of leucocyte TGNs in children taking TG was similar to the range of leucocyte TGNs in children taking MP. Therefore, when correlating intracellular TGNs to clinical effect, the range of erythrocyte TGN metabolites will be higher for those children taking TG than in those taking MP. PMID- 12111110 TI - Analysis of age, estimated creatinine clearance and pretreatment hematologic parameters as predictors of fludarabine toxicity in patients treated for chronic lymphocytic leukemia: a CALGB (9011) coordinated intergroup study. AB - PURPOSE: Fludarabine is a renally excreted agent that is an effective treatment for chronic lymphocytic leukemia (CLL), a disease predominantly of the elderly. We sought to determine whether age, renal function or pretreatment hematologic status predicted toxicity of fludarabine treatment for CLL. METHODS: We evaluated 192 patients with previously untreated B-cell CLL who were entered onto the fludarabine treatment arm (25 mg/m(2) daily for 5 days every 28 days) of CALGB study 9011, an intergroup study with participation from SWOG, CTG/NCI-C and ECOG. Patients were required to have serum creatinine within 1.5 times normal. Hematologic indices and infections were recorded during the first 28-day cycle of treatment. A time-to-toxicity endpoint was evaluated over the entire course of fludarabine treatment. Creatinine clearance (CrCl(est)) was estimated using serum creatinine, age and body mass index. RESULTS: The median age was 64 years (range 37-87 years) and median CrCl(est) was 62 ml/min (range 27-162 ml/min, interquartile range 52-79 ml/min). We found no association between age and incidence of hematologic toxicity or infection during the first cycle of treatment. There was a strong association between CrCl(est) and the time-to toxicity endpoint. Patients with CrCl(est) below 80 ml/min had increased incidence of toxicity during their treatment course ( P<0.0001). Pretreatment anemia, thrombocytopenia and Rai stage were highly associated with the incidence of neutrophil toxicity and grade III/IV hematologic toxicities during the first cycle of treatment ( P<0.0001). CONCLUSIONS: Patient age was not an independent risk factor for fludarabine-related toxicity, but CrCl(est) was associated with time to toxicity. PMID- 12111112 TI - The effect of keratinocyte growth factor on tumour growth and small intestinal mucositis after chemotherapy in the rat with breast cancer. AB - PURPOSE: Mucositis from cancer chemotherapy is a common problem for which there is no definitive treatment. It produces significant morbidity and occasional mortality. Prevention and successful treatment could significantly enhance the quality of life of patients, and improve survival; however any potential preventative agent must not enhance tumour growth. The aims of this study were to assess the effect of keratinocyte growth factor (KGF) on breast tumour growth, and in preventing small intestinal mucositis induced by methotrexate (MTX). METHODS: Tumour-bearing rats received KGF or saline for 5 days prior to either MTX or saline treatment, and were killed 24 h after the last MTX injection. The weights of the tumour, small and large intestines, and liver were recorded. Apoptosis was assessed by TUNEL assay in the tumour and jejunum. Intestinal morphometry was used to assess villus area, crypt length and mitotic crypt count. Tumour proliferation was assessed by mitotic count. RESULTS: KGF increased the weight of the small intestine prior to chemotherapy but the weight was not maintained after chemotherapy. KGF synergized with MTX to increase apoptosis in both intestinal crypts and the breast cancer. KGF also reduced tumour size. CONCLUSIONS: We conclude that KGF had a modest effect on intestinal growth prior to chemotherapy. It did not protect the gut from mucositis, nor did it worsen morphometry. It reduced tumour size. PMID- 12111111 TI - Chemotherapeutic agents enhance TRAIL-induced apoptosis in prostate cancer cells. AB - PURPOSE: Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a member of the tumor necrosis factor (TNF) family that preferentially kills tumor cells. In this study, we sought to determine whether chemotherapeutic agents augment TRAIL-induced cytotoxicity in human prostate cancer cells, and whether this sensitivity can be blocked by overexpression of bcl-2. METHODS: Prostate cancer cells, PC3 and LNCaP, were treated with TRAIL alone, drug alone or a combination of both for 24 h. Cytotoxicity was determined by DNA fragmentation and clonogenic survival assay. RESULTS: Treatment with the conventional chemotherapeutic agents cisplatin (2 and 5 microg/ml), etoposide (10 microM and 20 microM) and doxorubicin (30 and 60 n M) dramatically augmented TRAIL-induced apoptosis in LNCaP and PC3 cells. TRAIL-induced apoptosis was partially abrogated by overexpression of bcl-2 in these two cell lines when it was used in combination with the above agents. Similar results were obtained using clonogenic survival assays where bcl-2 overexpression was also found to marginally protect against TRAIL- and chemotherapy-induced cell killing. CONCLUSIONS: This study demonstrates that combination treatment of prostate cancer cells with TRAIL and chemotherapeutic agents overcomes their resistance by triggering caspase activation. This greater than additive effect of cotreatment with TRAIL and chemotherapy may provide the basis for a new therapeutic approach to induce apoptosis in otherwise resistant cancer cells. PMID- 12111114 TI - Suicide gene therapy using E. coli beta-galactosidase. AB - PURPOSE: Suicide gene therapy offers the potential to increase the selective toxicity of antitumor agents by intratumoral expression of exogenous enzymes that convert nontoxic prodrugs to toxic products. The use of herpes simplex virus thymidine kinase with ganciclovir, and E. coli cytosine deaminase with 5 fluorocytosine are well-known examples of this approach. The purpose of this study was to investigate a novel suicide gene therapy using E. coli beta galactosidase (beta-gal) as the prodrug-activating enzyme. Advantages of this approach include: (1) the ability to use prodrugs that are cleaved by beta-gal to agents that are known to possess activity against human solid tumors, and (2) the extensive experience gained with targeting beta-gal to specific tumors in experimental animals and in humans. METHODS: Two different structural types of anthracycline prodrugs, N-[4"-(beta- D-galactopyranosyl)-3" nitrobenzyloxycarbonyl]daunomycin (Daun02) and N-[(4" R,S)-4"-ethoxy-4"-(1"'- O beta- D-galactopyranosyl)butyl]daunorubicin (gal-DNC4) were investigated. The prodrugs were evaluated as substrates for beta-gal. Cytotoxicity studies of Daun02 were conducted against a murine tumor (Panc02), two human breast tumors (MCF-7 and T47D), and three human prostate tumors (PC3, DU145 and LNCAP) that had been transduced to express beta-gal. Antitumor studies of Daun02 were conducted against mouse tumor Panc02 xenografts implanted subcutaneously. RESULTS: Daun02 was a good substrate for beta-gal. By comparison, gal-DCN4 was a poor substrate. Except for PC3, the beta-gal-transduced tumors showed 3- to 60-fold increased sensitivity to Daun02 compared with mock-transduced control cells. Daunomycin was formed from Daun02 in tissue culture medium containing beta-gal-transduced cell lines but was not observed in the medium from mock-transduced controls. In vivo therapeutic studies of Daun02 against the Panc02 tumor in athymic mice showed no significant inhibition of tumor growth. Pharmacokinetic studies showed limited distribution of the prodrug beyond the vascular space. CONCLUSIONS: E. coli beta gal may be useful as a prodrug-activating enzyme in suicide gene therapy and has the potential to increase the selective toxicity of conventional antitumor agents. Although this approach worked well against tumor cells in vitro, it was not effective against a xenograft model in vivo, apparently because of poor drug tissue distribution. PMID- 12111113 TI - Urinary and fecal excretion of topotecan in patients with malignant solid tumours. AB - PURPOSE: The objectives of the study were to determine the pharmacokinetics and routes of excretion of topotecan following intravenous or oral administration to patients with refractory solid tumours. METHODS: Patients were randomized to receive either oral (2.3 mg/m(2)) or intravenous (1.5 mg/m(2)) topotecan once daily for 5 days in course 1. Patients who received in course 1 oral topotecan received in course 2 intravenous topotecan on day 1 followed by oral topotecan on days 2 to 5. Patients who received in course 1 intravenous topotecan received in course 2 oral topotecan once daily for 5 days. Plasma pharmacokinetics were performed on day 1 of course 1 (all patients) and course 2 (only patients receiving intravenous topotecan on that day). In course 1, urine and feces were collected for up to 9 days after the first dosage. The amounts of topotecan and N desmethyl topotecan in plasma, urine and feces were determined by validated high performance liquid chromatographic assays. RESULTS: A total of 11 patients were enrolled in the study. Nine patients were evaluable for pharmacokinetics. Plasma pharmacokinetics were similar to those previously reported. The principal route of excretion was the urine, with approximately 49% of the intravenously administered topotecan dose and 20% of the oral dose collected in the urine as parent drug. Approximately 18% and 33% of the intravenous and oral dose, respectively, were recovered unchanged in the feces. Only small amounts of N desmethyl topotecan were found in the excreta. CONCLUSIONS: Fecal and urinary excretion of unchanged topotecan were the major routes of topotecan elimination. Approximately 28% of the intravenous dose and 43% of the oral dose of topotecan were unaccounted for and eliminated through other routes. PMID- 12111115 TI - Structure-activity relationships of alkylphosphocholine derivatives: antineoplastic action on brain tumor cell lines in vitro. AB - Erucylphosphocholine (ErPC) is a promising candidate for the treatment of human brain tumors. The aim of the present study was to investigate whether structural modifications of ErPC would improve its antineoplastic activity in vitro. The novel alkylphosphocholine (APC) derivatives docosenyl-( cis-10,11) phosphocholine, tricosenyl-( cis-12,13)-phosphocholine, heneicosenyl-( cis-12,13) phosphocholine and erucyl- N, N, N-trimethylpropanolaminophosphate all reduced cell growth and viability of rat and human astrocytoma/glioblastoma (AC/GBM) cell lines (C6, T98G, U87MG, A172) and had improved antineoplastic activity when compared to the prototypical APC hexadecylphosphocholine (HePC). However, the four cell lines differed in their sensitivity to the APC derivatives. A172 cells were most sensitive to their cytostatic action and T98G cells to their cytotoxic action. The LC(50) values for T98G cells after a 72-h exposure to the novel derivatives varied between 25 and 54 microM compared to 45+/-8.1 microM for ErPC. Complete killing of T98G cells was obtained with all derivatives at 90 microM. Structural modifications of the chain length of the alcohol moiety as well as changing the position of the double bond within the alkyl chain improved cytotoxicity of the APC in C6 and A172 cells and to a lesser extent in T98G cells, whereas U87MG cells showed almost similar sensitivities to the novel drugs and ErPC. Increasing the distance between the phosphorus and nitrogen atoms within the polar phosphocholine group did not alter antineoplastic activity but modified physicochemical characteristics, e.g. increased the solubility in water. In a similar manner to ErPC, all derivatives induced growth arrest in the G(2)/M phase of the cell cycle and apoptotic cell death. Importantly, none of the derivatives showed hemolytic activity. As there was no clear superiority of any of the novel derivatives, ErPC remains the leading APC derivative for future clinical trials in brain tumor chemotherapy. PMID- 12111116 TI - Pilot study of gemcitabine (10 mg/m(2) per min) and cisplatin. AB - PURPOSE: Gemcitabine and cisplatin are routinely used in combination. In this combination, gemcitabine at conventional doses of 1000-1500 mg/m(2) is delivered weekly as a 30-minute bolus. Laboratory data suggest that the synthesis of gemcitabine triphosphate is saturable, and that gemcitabine infused at a rate of 10 mg/m(2) per min optimizes accumulation of the triphosphate. Early clinical experience suggests that gemcitabine delivered by a more prolonged infusion is more myelosuppressive. In this pilot study, we wished to assess if full-dose gemcitabine when given with cisplatin could be delivered by this more prolonged infusion. METHODS: In this study, all patients received cisplatin 75 mg/m(2). All gemcitabine doses were delivered at 10 mg/m(2) per min. For the initial cohort (level 1) the gemcitabine dose was 800 mg/m(2) per min. Subsequent escalations were 1000 mg/m(2) per min (level 2), and 1250 mg/m(2) per min (levels 3 and 4). For the first three cohorts, patients received gemcitabine on days 1, 8, and 15 and cisplatin on day 15 on a 28-day cycle. Patients on level 4 received gemcitabine on days 1 and 8 and cisplatin on day 8 on a 21-day cycle. Dose omissions or delays (holds) were mandated for NCI CTC grade 3 or 4 granulocytopenia or grade 2-4 thrombocytopenia. RESULTS: Entered onto this dose finding study were 23 patients (12 male, 11 female) with advanced solid tumors. Seven patients were treatment-naive. Six patients were treated on level 1, five each on levels 2 and 3 and seven on level 4. Patients received one to seven cycles of treatment. Myelosuppression-related dose holds occurred at all levels. First-cycle dose holds occurred in three of six, four of five, three of five and two of seven patients on successive levels. First-cycle grade 3 or 4 granulocytopenia/thrombocytopenia occurred in three patients on level 1, one patient on level 2, two patients on level 3 and three patients on level 4. There were no partial or complete responses. Four patients were removed for toxicity (three myelosuppression, one nephrotoxicity), one at physician discretion, and 15 with disease progression. Three patients stopped therapy with stable disease after 5-6 months. On level 3, three of five patients remained on therapy for 4 months or more, compared to only one patient on each of the other three levels. CONCLUSIONS Weekly gemcitabine 1250 mg/m(2), utilizing a 10 mg/m(2) per min infusion rate, can be delivered with cisplatin 75 mg/m(2) with tolerable toxicity. When used in combination with cisplatin, however, the benefit of this fixed dose rate infusion gemcitabine compared to standard bolus gemcitabine remains to be determined. PMID- 12111117 TI - Immature myeloid cells and cancer-associated immune suppression. AB - Impaired balance between mature and immature myeloid cells is one of the hallmarks of cancer. In cancer patients as well as in mouse models there is increasing evidence that progressive tumor growth is associated with an accumulation of immature myeloid cells, monocytes/macrophages, and with a decreased number and function of dendritic cells (DC). This review examines recent findings on the contribution of immature myeloid cells (ImC) to cancer induced immune suppression. PMID- 12111118 TI - Eliciting cytotoxic T lymphocytes against acute myeloid leukemia-derived antigens: evaluation of dendritic cell-leukemia cell hybrids and other antigen loading strategies for dendritic cell-based vaccination. AB - Dendritic cells (DC) have been successfully used in clinical pilot studies to induce tumor-specific immunity as well as clinical response in selected patients. However, DC-based immunotherapy remains a challenge and several parameters need to be examined in order to optimize the induction of anti-tumor immune responses. This study focuses on DC vaccination for leukemia and evaluates the in vitro efficacy of three different strategies for generating antigen-loaded DC-based vaccines for the induction of major histocompatibility complex (MHC) class I restricted anti-leukemia cytotoxic T lymphocyte (CTL) responses. These included direct fusion of DC with leukemia cells to generate DC-leukemia cell hybrids, and DC pulsed with either apoptotic leukemia cell fragments or whole tumor cell lysates. Using either the U937 cell line or primary human acute myeloid leukemia blasts (AML), DC-leukemia cell hybrids were found to be the most potent in vitro inducers of CTL activity. DC pulsed with apoptotic tumor cell fragments were less efficient, but induced a more potent CTL response compared to tumor lysate-pulsed DC. The CTL responses were both MHC class I-restricted and antigen-specific, as shown by the inability of the CTL to lyse other control targets. The data presented here suggest that the method of antigen loading onto DC may be critical in the design of tumor vaccines. PMID- 12111119 TI - Hsp110 over-expression increases the immunogenicity of the murine CT26 colon tumor. AB - Several studies have suggested a positive correlation between heat shock protein (hsp) expression and tumor immunogenicity. Independently, many studies have shown that hsp purified from tumors can be used as a tumor-specific vaccine. In this study, we have explored the connection between hsp expression and anti-tumor immunity by transducing murine CT26 colon carcinoma cells with the cDNA of a major hsp, i.e. hsp110. We have shown that over-expression of hsp110 has no effect on CT26 tumor cell growth in vitro, and does not inhibit their anchorage independent growth capacity. However, in situ, hsp110 over-expressing CT26 tumor (CT26-hsp110) grew at a significantly reduced rate as compared to the wild-type CT26 tumor in immunocompetent mice. Moreover, immunization of mice with inactivated CT26-hsp110 cells significantly inhibited the growth of wild-type CT26 tumor. This immunity was associated with an increased frequency of tumor specific T cells after vaccination. An in vivo antibody depletion assay demonstrated that inactivated CT26-hsp110 cells elicited anti-tumor responses involving CD8(+) T cells and natural killer (NK) cells, but not CD4(+) T cells. Lastly, the effect of the addition of granulocyte-macrophage colony stimulating factor (GM-CSF) to these vaccine formulations was determined. Mice immunized with irradiated CT26-hsp110 cells combined with GM-CSF-producing bystander cells revealed a complete inhibition of CT26 tumor growth, indicating a synergy between inactivated CT26-hsp110 vaccine activity and GM-CSF. These observations demonstrate that manipulation of hsp110 expression in tumors, specifically when combined with GM-CSF, represents a potentially powerful approach to cancer vaccine formulation. PMID- 12111120 TI - Effect of the dose and composition of an autologous hapten-modified melanoma vaccine on the development of delayed-type hypersensitivity responses. AB - We have reported that treatment of melanoma patients with a vaccine consisting of autologous tumor cells modified with the hapten, dinitrophenyl (DNP) and preceded by low-dose cyclophosphamide induces delayed-type hypersensitivity (DTH) to autologous, unmodified tumor cells and that this response is a significant predictor of survival. We analyzed the vaccines prepared for 284 patients who were treated following resection of regional or distant metastases to find out whether the dose and composition determined the immunological response. A positive DTH response (> or =5 mm induration) to unmodified autologous tumor cells was induced in 57% of the patients (median: 5 mm; range: 0-22 mm). Regression analysis showed no significant association between the magnitude of DTH and the number of live (trypan blue exclusion) melanoma cells per dose over a dosage range of 0.5-25.0 x 10(6). Surprisingly, there was a small but significant positive relationship between the mean number of dead cells in the vaccines of a given patient and that patient's maximum DTH to unmodified melanoma cells. Only 37% of patients whose vaccines contained >50% live cells developed DTH, as compared with 69% and 65% of patients whose vaccines contained 26% to 50% or < or =25% live cells, respectively. Thus, it appears that dead tumor cells contribute to the immunogenicity of the DNP vaccine, but other factors such as the administration schedule may be more important determinants of immunological and clinical outcome. PMID- 12111122 TI - Linkage of CD40L to a self-tumor antigen enhances the antitumor immune responses of dendritic cell-based treatment. AB - CD40-CD40 ligand (CD40L) interaction is an important costimulatory signal in the interaction between T cells and antigen-presenting cells (APC). In the present study, we determined whether the linkage of CD40L to the tumor-specific idiotype (Id) derived from a murine B-cell lymphoma, 38C13, could enhance its immunogenicity when presented by dendritic cells (DC). We showed that bone marrow derived DC pulsed with Id-CD40L upregulated the expression of CD40, CD80, CD86, and major histocompatibility complex (MHC) class II molecules with the increased production of interleukin-12 (IL-12). Mice immunized with DC loaded with Id-CD40L showed high levels of anti-Id antibody response of both IgG2a and IgG1 isotypes. In addition, nylon wool-enriched T cells from these immunized mice showed a tumor specific T-cell proliferative response upon stimulation with Id protein. Mice immunized with DC pulsed with Id alone failed to show any of these immune responses. Immunization with DC pulsed with Id-CD40L showed increased resistance to the challenge by 38C13 tumor, and tumor growth was significantly retarded. Together, these results show that linkage of CD40L to a self-tumor antigen enhances the anti-tumor immune response in DC-based treatment. PMID- 12111121 TI - Molecular requirements for CD8-mediated rejection of a MUC1-expressing pancreatic carcinoma: implications for tumor vaccines. AB - Previous studies have indicated that different effector cells are required to eliminate MUC1-expressing tumors derived from different organ sites and that different vaccine strategies may be necessary to generate these two different MUC1-specific immune responses. In this study, we characterized molecular components that are required to produce immune responses that eliminate Panc02.MUC1 tumors in vivo by utilizing mice genetically deficient in molecules related to immunity. A parallel study has been reported for a B16.MUC1 tumor model. We confirmed that a CD8(+) effector cell was required to eliminate MUC1 expressing Panc02 tumors, and demonstrated that T cells expressing TCR-alpha/beta and co-stimulation through CD28 and CD40:CD40L interactions played critical roles during the initiation of the anti-Panc02.MUC1 immune response. TCR-alpha/beta(+) cells were required to eliminate Panc02.MUC1 tumors, while TCR-gamma/delta(+) cells played a suppressive non-MUC1-specific role in anti-Panc02 tumor immunity. Type 1 cytokine interferon-gamma (IFN-gamma), but not interleukin-12 (IL-12), was essential for eliminating MUC1-expressing tumors, while neither IL-4 nor IL-10 (type 2 cytokines) were required for tumor rejection. In vitro studies demonstrated that IFN-gamma upregulated MHC class I, but not MHC class II, on Panc02.MUC1 tumor cells. Surprisingly, both perforin and FasL played unique roles during the effector phase of immunity to Panc02.MUC1, while lymphotoxin-alpha, but not TNFR-1, was required for immunity against Panc02.MUC1 tumors. The findings presented here and in parallel studies of B16.MUC1 immunity clearly demonstrate that different effector cells and cytolytic mechanisms are required to eliminate MUC1-expressing tumors derived from different organ sites, and provide insight into the immune components required to eliminate tumors expressing the same antigen but derived from different tissues. PMID- 12111123 TI - Immunogenicity of WT-1 peptides. PMID- 12111124 TI - Establishment of radioactive astatine and iodine uptake in cancer cell lines expressing the human sodium/iodide symporter. AB - The sodium/iodide symporter (NIS) has been recognized as an attractive target for radioiodine-mediated cancer gene therapy. In this study we investigated the role of human NIS for cellular uptake of the high LET alpha-emitter astatine-211 ((211)At) in comparison with radioiodine as a potential radionuclide for future applications. A mammalian NIS expression vector was constructed and used to generate six stable NIS-expressing cancer cell lines (three derived from thyroid carcinoma, two from colon carcinoma, one from glioblastoma). Compared with the respective control cell lines, steady state radionuclide uptake of NIS-expressing cell lines increased up to 350-fold for iodine-123 ((123)I), 340-fold for technetium-99m pertechnetate ((99m)TcO(4)(-)) and 60-fold for (211)At. Cellular (211)At accumulation was found to be dependent on extracellular Na(+) ions and displayed a similar sensitivity towards sodium perchlorate inhibition as radioiodide and (99m)TcO(4)(-) uptake. Heterologous competition with unlabelled NaI decreased NIS-mediated (211)At uptake to levels of NIS-negative control cells. Following uptake both radioiodide and (211)At were rapidly (apparent t(1/2) 3-15 min) released by the cells as determined by wash-out experiments. Data of scintigraphic tumour imaging in a xenograft nude mice model of transplanted NIS-modified thyroid cells indicated that radionuclide uptake in NIS expressing tumours was up to 70 times ((123)I), 25 times ((99m)TcO(4)(-)) and 10 times ((211)At) higher than in control tumours or normal tissues except stomach (3-5 times) and thyroid gland (5-10 times). Thirty-four percent and 14% of the administered activity of (123)I and (211)At, respectively, was found in NIS tumours by region of interest analysis ( n=2). Compared with cell culture experiments, the effective half-life in vivo was greatly prolonged (6.5 h for (123)I, 5.2 h for (211)At) and preliminary dosimetric calculations indicate high tumour absorbed doses (3.5 Gy/MBq(tumour) for (131)I and 50.3 Gy/MBq(tumour) for (211)At). In conclusion, NIS-expressing tumour cell lines of different origin displayed specific radionuclide uptake in vitro and in vivo. We provide first direct evidence that the high-energy alpha-emitter (211)At is efficiently transported by NIS. Application of (211)At may direct higher radiation doses to experimental tumours than those calculated for (131)I. Thus, (211)At may represent a promising alternative radionuclide for future NIS-based tumour therapy. PMID- 12111125 TI - Co-expressed peptide receptors in breast cancer as a molecular basis for in vivo multireceptor tumour targeting. AB - Breast cancers can express different types of peptide receptors such as somatostatin, vasoactive intestinal peptide (VIP), gastrin-releasing peptide (GRP) and NPY(Y(1)) receptors. The aim of this in vitro study was to evaluate which is the most appropriate peptide receptor or peptide receptor combination for in vivo diagnostic and therapeutic targeting of breast cancers. Seventy-seven primary breast cancers and 15 breast cancer lymph node metastases were investigated in vitro for their expression of somatostatin, VPAC(1), GRP and NPY(Y(1)) receptors using in vitro receptor autoradiography on successive tissue sections with (125)I-[Tyr(3)]-octreotide, (125)I-VIP, (125)I-[Tyr(4)]-bombesin and (125)I-[Leu(31),Pro(34)]-PYY respectively. This study identified two groups of tumours: a group of 68 tumours (88%) with at least one receptor expressed at high density (>2,000 dpm/mg tissue) that may provide a strong predictive value for successful in vivo targeting, and a group of nine tumours (12%) with no receptors or only a low density of them (<2,000 dpm/mg tissue). In the group with high receptor density, 50 of the 68 tumours (74%) expressed GRP receptors, 45 (66%) expressed NPY(Y(1)) receptors, 25 (37%) expressed VPAC(1) receptors and 14 (21%) expressed somatostatin receptors. Mean density was 9,819+/-530 dpm/mg tissue for GRP receptors, 9,135+/-579 dpm/mg for NPY(Y(1)) receptors, 4,337+/-528 dpm/mg for somatostatin receptors and 3,437+/-306 dpm/mg for VPAC(1) receptors. It is of note that tumours expressing NPY(Y(1)) or GRP receptors, or both, were found in 63/68 (93%) cases. Lymph node metastases showed a similar receptor profile to the corresponding primary tumour. This in vitro study strongly suggests that the combination of radiolabelled GRP and Y(1) analogues should allow targeting of breast carcinomas and their lymph node metastases for in vivo peptide receptor scintigraphy and radiotherapy. PMID- 12111126 TI - A novel quantitative dual-isotope method for simultaneous ventilation and perfusion lung SPET. AB - A quantitative dual-isotope single-photon emission tomography (SPET) technique for the assessment of lung ventilation (V) and perfusion (Q) using, respectively, technetium-99m labelled Technegas (140 keV) and indium-113m labelled macro aggregated albumin (392 keV), is presented, validated and clinically tested in a healthy volunteer. In order to assess V, Q and V/Q distributions in quantitative terms, algorithms which correct for down scattering, photon scattering and attenuation, as well as an organ outline algorithm, were implemented. Scatter and down-scatter correction were made in the spatial domain by pixel-wise image subtraction of projection-dependent global scattering factors obtained from the energy domain. The attenuation correction was based on an iterative projection/back-projection method. All studies were made on a three-headed SPET system (Trionix) with medium-energy parallel-hole collimators. The set of input data for quantification was based on SPET acquisition of emission data in four separate energy windows, the associated cumulative energy spectra and transmission data. The attenuation correction routine as well as the edge detection algorithm utilized data from (99m)Tc transmission tomography. Attenuation data for (113m)In were obtained by linear scaling of the (99m)Tc attenuation maps. The correction algorithms were experimentally validated with a stack phantom system and applied on a healthy volunteer. The mean difference between the corrected SPET data of the dense stack lung phantom and those obtained from the corresponding scatter- and attenuation-"free" version was only 1.9% for (99m)Tc and 0.9% for (113m)In. The estimated fractional V/Q distribution in the 3-D lung phantom volume had its peak at V/Q=1, with a width (FWHM) of 0.31 due to noise, particularly in the (113m)In images, and to partial volume effects. For a healthy volunteer, the corresponding values were 0.9 and 0.35, respectively. This method allows accurate assessment of radionuclide distribution on a regional basis. For basic lung physiology and clinical practice, the method allows assessment of the global frequency functions of the V, Q and V/Q distributions. PMID- 12111127 TI - 99mTc-MIBI SPET in non-small cell lung cancer in relationship with Pgp and prognosis. AB - Higher technetium-99m methoxyisobutylisonitrile (MIBI) uptake in non-small cell lung cancer (NSCLC) has been reported to be associated with a positive response to chemotherapy. It has previously been found that in tumour cells, P glycoprotein (Pgp) expression is of importance for tracer uptake. However, some studies have indicated that Pgp expression does not play an important role in (99m)Tc-MIBI uptake in NSCLC; indeed, a negative correlation between (99m)Tc-MIBI uptake and Pgp expression has been reported. Against the background of conflicting results, our aim was to evaluate the relationship between (99m)Tc MIBI uptake, prognosis and Pgp expression in NSCLC. A total of 37 patients with NSCLC underwent (99m)Tc-MIBI single-photon emission tomography (SPET) before chemotherapy. In 19 patients both Pgp and p53 expression, and in two patients only p53 expression (due to the limited biopsy material), were measured with immunohistochemical staining. (99m)Tc-MIBI uptake was significantly higher in responders than in non-responders: 3.09+/-1.14 vs 2.24+/-0.88 ( P<0.03) and 3.09+/-1.08 vs 2.37+/-1.06 ( P<0.05) for the early ratio (ER) and the delayed ratio (DR), respectively. The wash-out rate (WR) of responders was not significantly different from that of non-responders. We found no significant differences in ER, DR and WR among the groups positive or negative for Pgp and p53 status. There was a significant positive correlation between the survival rate and both ER and DR: r=0.49 ( P=0.003) and r=0.40 ( P=0.018), respectively. Patients with ER and DR values above 3 showed significantly longer survival than those with values below 3: 14.7+/-8.5 months vs 7.3+/-5.1 months ( P<0.009) and 13.2+/-8.4 months vs 7.4+/-5.3 months ( P<0.04) for ER and DR, respectively. However, interestingly, and in contrast to expectations, patients with a Pgp score of +2 showed significantly longer survival (12.9+/-6.7 months) than those with Pgp scores of +1 (4.4+/-3.0 months) or - (negative) (3.8+/-2.2 months) ( P<0.009 and P<0.02, respectively). Our results suggest that in NSCLC, patients with higher (99m)Tc-MIBI uptake tend to show a positive response to chemotherapy, and patients with ER and DR values above 3 have a significantly better prognosis. We also found that Pgp expression seems to play only a minor role in (99m)Tc-MIBI uptake. Our finding that patients with ER and DR values above 3 have a better prognosis needs to be confirmed in larger series of patients. PMID- 12111128 TI - Low-dose dobutamine stress gated SPET for identification of viable myocardium: comparison with stress-rest perfusion SPET and PET. AB - The detection of viable myocardium is important for the prediction of functional recovery after revascularisation. However, a fixed perfusion defect often includes viable myocardium, and perfusion imaging then underestimates myocardial viability. We previously reported that low-dose dobutamine stress gated single photon emission tomography (SPET) provides similar findings to dobutamine stress echocardiography in the assessment of myocardial viability. The present study investigated whether low-dose dobutamine stress gated SPET is of additional value as compared with stress-rest technetium-99m tetrofosmin SPET for the detection of myocardial viability. Standard stress-rest perfusion SPET, low-dose dobutamine stress gated SPET and fluorine-18 fluorodeoxyglucose positron emission tomography (FDG PET) were studied in 23 patients (mean age 67+/-7.6 years) with previous myocardial infarction. Twenty-one of them were successfully studied with each technique. FDG PET viability (FDG uptake >/=50%) was employed as the gold standard. One-day stress-rest (99m)Tc-tetrofosmin myocardial SPET was performed. After the resting study, gated SPET was acquired following infusion of 7.5 microg kg(-1) min(-1) of dobutamine. Left ventricular wall motion in 16 segments was assessed by cine mode display using a four-point scale. Myocardial viability was considered present when there was improvement by one point. Of a total of 336 segments analysed, 53 had persistent defects on stress-rest perfusion SPET. FDG viability was seen in 16 of 17 dobutamine-responsive segments, but in only 11 of 36 dobutamine non-responsive segments ( P<0.01). Thus, in the segments with persistent defects, viability findings on low-dose dobutamine stress gated SPET were concordant with those on FDG PET in 77% of segments (kappa value =0.55). For the detection of FDG-viable myocardium, the combination of stress-rest perfusion SPET and low-dose dobutamine stress gated SPET achieved a better sensitivity than stress-rest perfusion SPET alone (35/46, 76% vs 19/46, 41.3%, P<0.001), with a similar specificity (25/29, 86% vs 26/29, 90%, P=NS). We conclude that in the identification of viable myocardium, low-dose dobutamine stress gated SPET may provide additional information missed on a routine stress-rest perfusion scan. Dobutamine stress gated SPET may provide new insights into myocardial viability on the basis of ischaemia and contractile reserve. PMID- 12111129 TI - Deferoxamine improves coronary vascular responses to sympathetic stimulation in patients with type 1 diabetes mellitus. AB - Effects of oxygen-derived free radicals are suggested to be a potential pathogenic factor for endothelial dysfunction. In this study we sought to evaluate the effect of hydroxyl radicals on the human coronary vascular bed in type I diabetes mellitus using positron emission tomography (PET). Thirteen patients with type 1 diabetes underwent PET using nitrogen-13 ammonia at rest and during sympathetic stimulation with the cold pressor test (CPT). The rest-stress study protocol was repeated twice (on different days) using pre-stress infusion of either saline as placebo or deferoxamine, an iron chelator which inhibits generation of hydroxyl radicals. At rest, global MBF was higher in diabetics than in normal controls (78.1+/-17.5 vs 63.2+/-14.9 mg 100 g(-1) min(-1), P<0.05) and myocardial vascular resistance (MVR) showed a trend towards lower values (patients, 1.28+/-0.35; controls, 1.55+/-0.32, P=NS). CPT increased MBF in all controls while 7/13 diabetics responded normally. CPT decreased MVR in 10/13 controls but in only 4/13 diabetics. There was no significant difference in the duration of diabetes, HbA1c, daily insulin dose, body mass index, or lipid profiles between patients with and patients without abnormal MBF or MVR responses. Pre-stress infusion of deferoxamine normalized MBF response in all six patients, and MVR response in six of the nine patients. Another group consisting of seven patients underwent a rest-rest protocol after infusion of deferoxamine and saline to investigate the effect of deferoxamine on resting MBF. Deferoxamine did not change the resting MBF (deferoxamine, 81+/-17 ml 100 g(-1) min(-1); saline, 75+/-19 ml 100 g(-1) min(-1), P=NS) or MVR (deferoxamine, 1.0+/-0.5 mmHg ml(-1) 100 g(-1) min(-1); saline, 1.2+/-0.6 mmHg ml(-1) 100 g(-1) min(-1), P=NS). In conclusion, inhibition of hydroxyl radical formation using deferoxamine significantly improved the responses of coronary microvasculature to sympathetic stimulation. Hydroxyl radicals may play a role in the pathogenesis of flow abnormalities in type 1 diabetes. PMID- 12111130 TI - Limbic system perfusion in Alzheimer's disease measured by MRI-coregistered HMPAO SPET. AB - The goal of this study was to perform a systematic, semi-quantitative analysis of limbic perfusion in patients with Alzheimer's disease (AD) using coregistered single-photon emission tomography (SPET) images aligned to magnetic resonance (MR) images. Limbic perfusion in 40 patients with mild to moderate AD was compared with that of 17 age-, sex-, and education-matched normal controls (NC). HMPAO SPET scans and 3D T1-weighted MR images were acquired for each subject. Structures of the limbic system (i.e. hippocampus, amygdala, anterior thalamus, hypothalamus, mamillary bodies, basal forebrain, septal area and cingulate, orbitofrontal and parahippocampal cortices) were traced on the MR images and transferred to the coregistered SPET scans. Perfusion ratios for all limbic regions were calculated relative to cerebellar perfusion. General linear model multivariate analysis revealed that, overall, limbic structures showed significant hypoperfusion (F=7.802, P<0.00001, eta (2)=0.695 ) in AD patients compared with NC. Greatest differences (d > or = 0.8) were found in the hippocampus, as well as all areas of the cingulate cortex. Significant relative hypoperfusion was also apparent in the parahippocampal cortex, amygdala/entorhinal cortex, septal area and anterior thalamus, all of which showed medium to large effect sizes (d=0.6-0.8). No significant relative perfusion differences were detected in the basal forebrain, hypothalamus, mamillary bodies or orbitofrontal cortex. Logistic regression indicated that posterior cingulate cortex perfusion was able to discriminate AD patients from NC with 93% accuracy (95% sensitivity, 88% specificity). The current results suggest that most, but not all, limbic structures show significant relative hypoperfusion in AD. These findings validate previous post-mortem studies and could be useful in improving diagnostic accuracy, monitoring disease progression and evaluating potential treatment strategies in AD. PMID- 12111131 TI - Coupling of porcine bone blood flow and metabolism in high-turnover bone disease measured by [(15)O]H(2)O and [(18)F]fluoride ion positron emission tomography. AB - Previously, we identified a parathyroid hormone-related high-turnover bone disease after gastrectomy in mini pigs. Dynamic [(18)F]fluoride ion positron emission tomography (PET) revealed that bone metabolism was significantly increased, but that bone blood flow derived from permeability-surface area product (PS product)-corrected K(1) values was not. Since bone blood flow and metabolism are coupled in normal bone tissues, we hypothesised that the capillary permeability and/or surface area might be altered in high-turnover bone disease. The "true" bone blood flow ( f(H2O)) was measured in vertebral bodies by dynamic [(15)O]H(2)O PET, followed by a 120-min dynamic [(18)F]fluoride ion PET study, 6 months after total gastrectomy (n=5) and compared with results in sham-operated animals (n=5). Estimates for bone blood flow based on PS-corrected K(1) values (f) and the net uptake of fluoride in bone tissue (K(i)), representing the bone metabolic activity, were calculated using standard compartmental modelling and non-linear fitting. Gastrectomy was followed by a significant elevation of K(i) and k(3) ( P<0.05), which was mainly caused by an increase of the fraction of bound tracer in tissue (P<0.01). In contrast, f(H2O), f, the single-pass extraction fraction of [(18)F]fluoride (E) and the volume of distribution (DV) of [(18)F]fluoride were not significantly different between groups. In both groups, a coupling of the mean f(H2O) and K(i) values was found, but the intercept with the y-axis was higher in high-turnover bone disease. It is concluded that in high turnover bone disease following gastrectomy, the PS product for [(18)F]fluoride remains unchanged. Therefore, even in high-turnover bone diseases, [(18)F]fluoride ion PET can provide reliable blood flow estimates (f), as long as a proper PS product correction is applied. The increased bone metabolism in high turnover bone disease after gastrectomy is mainly related to an up-regulation of the amount of ionic exchange of [(18)F]fluoride with the bone matrix, while tracer delivery remains unchanged. PMID- 12111132 TI - FDG-PET improves the staging and selection of patients with recurrent colorectal cancer. AB - Whole-body fluorine-18 fluorodeoxyglucose positron emission tomography (FDG-PET) has proved effective in the diagnosis and staging of recurrent colorectal cancer. In this study, we analysed how PET affects the management of patients with recurrent colorectal cancer by permitting more accurate selection of candidates for curative resection. The data of 79 patients with known or suspected recurrent colorectal cancer were analysed. Conventional imaging modalities (CIM) and PET results were compared with regard to their accuracy in determining the extent and the resectability of tumour recurrence. Recurrence was demonstrated in 68 of the 79 patients. The data indicate that PET was superior to CIM for detection of recurrence at all sites except the liver. Based on the CIM+PET staging, surgery with curative intent was proposed in 39 patients and was indeed achieved in 31 of them (80%). PET was more accurate than CIM alone in predicting the resectability or non-resectability of the recurrence (82% vs 68%, P=0.02). It is concluded that whole-body FDG-PET is highly sensitive for both the diagnosis and the staging of patients with recurrent colorectal cancer. Its use in conjunction with conventional imaging procedures results in a more accurate selection of patients for surgical treatment with curative intent. PMID- 12111133 TI - PET attenuation coefficients from CT images: experimental evaluation of the transformation of CT into PET 511-keV attenuation coefficients. AB - The CT data acquired in combined PET/CT studies provide a fast and essentially noiseless source for the correction of photon attenuation in PET emission data. To this end, the CT values relating to attenuation of photons in the range of 40 140 keV must be transformed into linear attenuation coefficients at the PET energy of 511 keV. As attenuation depends on photon energy and the absorbing material, an accurate theoretical relation cannot be devised. The transformation implemented in the Discovery LS PET/CT scanner (GE Medical Systems, Milwaukee, Wis.) uses a bilinear function based on the attenuation of water and cortical bone at the CT and PET energies. The purpose of this study was to compare this transformation with experimental CT values and corresponding PET attenuation coefficients. In 14 patients, quantitative PET attenuation maps were calculated from germanium-68 transmission scans, and resolution-matched CT images were generated. A total of 114 volumes of interest were defined and the average PET attenuation coefficients and CT values measured. From the CT values the predicted PET attenuation coefficients were calculated using the bilinear transformation. When the transformation was based on the narrow-beam attenuation coefficient of water at 511 keV (0.096 cm(-1)), the predicted attenuation coefficients were higher in soft tissue than the measured values. This bias was reduced by replacing 0.096 cm(-1) in the transformation by the linear attenuation coefficient of 0.093 cm(-1) obtained from germanium-68 transmission scans. An analysis of the corrected emission activities shows that the resulting transformation is essentially equivalent to the transmission-based attenuation correction for human tissue. For non-human material, however, it may assign inaccurate attenuation coefficients which will also affect the correction in neighbouring tissue. PMID- 12111135 TI - Towards in vivo nuclear microscopy: iodine-125 imaging in mice using micro pinholes. AB - Position-sensitive gamma-radiation detectors equipped with collimators have been used for in vivo imaging of the distribution of radiolabelled molecules in laboratory animals and humans for several decades. To date, the best image resolution achieved in a rodent is on the order of 1 mm. Here we demonstrate how a basic and compact gamma camera can be constructed for in vivo radionuclide imaging in small animals, at much higher spatial resolution. Resolution improvements were obtained by combining dense, shaped, micro-pinhole apertures with iodine-125, an isotope with low energy emissions, ease of incorporation into a wide range of molecules, and straightforward translation into the clinic via other isotopes of iodine that are suitable for nuclear medicine imaging. (125)I images of test distributions and a mouse thyroid have been obtained at a resolution of as high as 200 microm using this simple bench-top camera. Possible future applications and extension to ultra-high-resolution emission tomography are discussed. PMID- 12111134 TI - Combining iodine-131 Lipiodol therapy with low-dose cisplatin as a radiosensitiser: preliminary results in hepatocellular carcinoma. AB - A prospective pilot trial was performed in 20 patients randomised to receive either (131)I-Lipiodol therapy alone (n=10) or (131)I-Lipiodol combined with a short low-dose cisplatin infusion (n=10), the aim being to evaluate the possible positive influence of a radiosensitiser on toxicity and tumour response. An activity of 1,354-2,128 MBq (mean 1,824 MBq) [36.6-57.5 mCi (mean 49.3 mCi)] (131)I-labelled Lipiodol was administered by selective instillation in the hepatic artery. Cisplatin was given in a dose of 30 mg/m(2) at day -1 and day +6 (day 0: (131)I-Lipiodol). The primary endpoint of this trial was toxicity of therapy; points of secondary interest were tumour response and survival at 6 months. With the use of cisplatin we found a higher percentage of stable or diminished tumour size (90%, vs 40% without). A benefit in group survival at 6 months was not evident. Low-grade stomatitis in one patient and minor changes in peripheral blood count were probably directly related to cisplatin, but its administration is unlikely to be associated with an excess of serious side effects. The use of low-dose cisplatin infusion as a radiosensitising agent in (131)I-Lipiodol therapy for hepatocellular carcinoma seems safe and may be beneficial for tumour control. Larger patient groups are necessary for confirmation and to establish the future role of (131)I-Lipiodol in hepatocellular carcinoma. PMID- 12111136 TI - Clinical relevance of left ventricular volume assessment by gated myocardial SPET in patients with coronary artery disease. AB - Coronary artery disease (CAD) is the leading cause of mortality in the Western world. Multiple parameters have been investigated to predict prognosis in CAD patients. The prognostic value of the left ventricular ejection fraction (LVEF) in patients with CAD is well established. More recently, left ventricular (LV) volumes have also shown prognostic value. Due to the favourable imaging characteristics of technetium-99m (high count density), the development of (99m)Tc-labelled myocardial perfusion tracers has made it possible to perform an electrocardiogram-gated acquisition during routine myocardial perfusion imaging. This enables assessment of LVEF and LV volumes during myocardial perfusion scintigraphy. This review considers the possible prognostic abilities of LV volume assessment by gated cardiac SPET. PMID- 12111137 TI - Drugs from the seas - current status and microbiological implications. AB - The oceans are the source of a large group of structurally unique natural products that are mainly accumulated in invertebrates such as sponges, tunicates, bryozoans, and molluscs. Several of these compounds (especially the tunicate metabolite ET-743) show pronounced pharmacological activities and are interesting candidates for new drugs primarily in the area of cancer treatment. Other compounds are currently being developed as an analgesic (ziconotide from the mollusc Conus magus) or to treat inflammation. Numerous natural products from marine invertebrates show striking structural similarities to known metabolites of microbial origin, suggesting that microorganisms (bacteria, microalgae) are at least involved in their biosynthesis or are in fact the true sources of these respective metabolites. This assumption is corroborated by several studies on natural products from sponges that proved these compounds to be localized in symbiotic bacteria or cyanobacteria. Recently, molecular methods have successfully been applied to study the microbial diversity in marine sponges and to gain evidence for an involvement of bacteria in the biosynthesis of the bryostatins in the bryozoan Bugula neritina. PMID- 12111138 TI - Industrial processes with animal cells. AB - Industrial processes involving animal cells for the production of useful products still seem to be rather uncommon. Nevertheless, during the last four decades of the last century the number of relevant processes has increased from production of virus vaccines to monoclonal antibodies and finally complex structured glycoproteins. As soon as cell lines became permanent and culture medium changed from purely biological fluids to more or less defined chemical media, large-scale cultivation could begin. The developments of the 1970s - fusion of cells to form hybridomas, and genetic engineering - triggered a second wave of products. Monoclonal antibodies and recombinant proteins for diagnosis and therapy set new challenges for the inventors. Historically, there has been no straightforward process development since the product dictates the process operation. Therefore, the scale of production covers the whole range from small multiple-unit reactors (flasks or roller bottles) up to 10,000-l single-unit batch reactors. Products with high value and small demand can be produced in multiple-unit systems whereas "bulk" products for vaccination and therapy may need large-scale bioreactors to be cost effective. All the different systems have their advantages and disadvantages and significant challenges that curb the development of effective perfusion cultures still remain. PMID- 12111139 TI - Biotechnology and bioremediation: successes and limitations. AB - With advances in biotechnology, bioremediation has become one of the most rapidly developing fields of environmental restoration, utilizing microorganisms to reduce the concentration and toxicity of various chemical pollutants, such as petroleum hydrocarbons, polycyclic aromatic hydrocarbons, polychlorinated biphenyls, phthalate esters, nitroaromatic compounds, industrial solvents, pesticides and metals. A number of bioremediation strategies have been developed to treat contaminated wastes and sites. Selecting the most appropriate strategy to treat a specific site can be guided by considering three basic principles: the amenability of the pollutant to biological transformation to less toxic products (biochemistry), the accessibility of the contaminant to microorganisms (bioavailability) and the opportunity for optimization of biological activity (bioactivity). Recent advances in the molecular genetics of biodegradation and studies on enzyme-tailoring and DNA-shuffling are discussed in this paper. PMID- 12111141 TI - Thermophilic production of lactic acid using integrated membrane bioreactor systems coupled with monopolar electrodialysis. AB - The thermophilic Bacillus strain BS119 was selected for this study to demonstrate the long term performance of lactic acid production and simultaneous pre purification. Integrated continuous cell recycle cultivation using ultra filtration membrane bioreactor (MBR) systems was investigated. The permeate from the MBR was routed to an on-line electrodialysis (ED) to recover, pre-purify and concentrate lactate. The cultivation and ED was operated at 60 degrees C for more than 1,000 h at a pH of 6.5. At lower dilution rate (0.02 h(-1)), lactate concentration reached a maximum of 55 g l(-1) with clearly lower residual glucose levels. At 0.04 h(-1), lactate concentration was significantly lower at 35 g l( 1). Maximal volumetric productivities of 1.38 g l(-1) h(-1) were achieved. Under stable conditions, lactic acid yield on consumed glucose appeared stable at around 80%. It could be demonstrated that the addition of supplements like yeast extract and peptone severely influences product formation. Integration of mono polar ED with the MBR systems yields lactate solutions with concentrations of up to 115 g l(-1). Because of the low substrate feed concentrations (less than 50 g l(-1)), lactate flux was rather poor, reaching a low maximum of 140 g m(-2) h( 1); nevertheless, stack energy consumption was positive with an average of 0.49 kWh kg(-1) lactate. PMID- 12111140 TI - A new photobioreactor for continuous marennin production with a marine diatom: influence of the light intensity and the immobilised-cell matrix (alginate beads or agar layer). AB - In oyster ponds, the marine diatom Haslea ostrearia synthesises and excretes a hydrosoluble pigment of commercial interest called marennin. During the benthic stage, when algal cells are naturally immobilised in their own polysaccharides, marennin production is higher. To optimise this production, axenic cultures of H. ostrearia were immobilised in a polysaccharidic matrix (alginate or agar) and introduced into a new photobioreactor device for continuous marennin production. Solute diffusion was improved using an alginate beads monolayer, leading to higher levels of cell growth (a 2-fold higher cell concentration) and marennin productivity (7.57-8.80 mg day(-1) l(-1)). An increase in the light intensity (from 3.0 to 8.5x10(16) quanta cm(-2) s(-1)) led to an earlier and 1.3-fold higher production of marennin. However, the higher light intensity led to a higher rate of cell death [0.29 instead of 0.40 ng chlorophyll a (10(6) cells)( 1)]. Due to the secondary nature of marennin metabolism, it would be necessary to alternate between culture conditions favouring cell growth (moderate light intensity and no limiting nitrate supply) and those promoting marennin production (high light intensity and limiting nitrate supply). PMID- 12111142 TI - Coupled lactic acid fermentation and adsorption. AB - Polyvinylpyridine (PVP) and activated carbon were evaluated for coupled lactic acid fermentation and adsorption, to prevent the product concentration from reaching inhibitory levels. The lactic acid production doubled as a result of periodical circulation of the fermentation broth through a PVP adsorption column. The adsorbent was then regenerated and the adsorbed lactate harvested, by passing 0.1 N NaOH through the column. However, each adsorption-regeneration cycle caused about 14% loss of the adsorption capacity, thus limiting the practical use of this rather expensive adsorbent. Activated carbon was found much more effective than PVP in lactic acid and lactate adsorption. The cells of Lactobacillus delbrueckii subsp. delbrueckii (LDD) also had strong tendency to adsorb on the carbon. A study was therefore conducted using an activated carbon column for simultaneous cell immobilization and lactate adsorption, in a semi-batch process with periodical medium replacement. The process produced lactate steadily at about 1.3 g l(-1)h(-1) when the replacement medium contained at least 2 g l(-1) of yeast extract. The production, however, stopped after switching to a medium without yeast extract. Active lactic acid production by LDD appeared to require yeast extract above a certain critical level (<2 g l(-1)). PMID- 12111143 TI - Inoculum production of the ectomycorrhizal fungus Pisolithus microcarpus in an airlift bioreactor. AB - Many important tree species in reforestation programs are dependent on ectomycorrhizal symbiosis in order to survive and grow, mainly in poor soils. The exploitation of this symbiosis to increase plant productivity demands the establishment of inoculum production methods. This study aims to propose an inoculum production method of the ectomycorrhizal fungus Pisolithus microcarpus (isolate UFSC-Pt116) using liquid fermentation in an airlift bioreactor with external circulation. The fungus grew as dark dense pellets during a batch fermentation at 25.5 degrees C and air inlet of 0.26-0.43 vvm. The maximum biomass (dry weight) achieved in the airlift bioreactor was approximately 5 g.l( 1) after 10-11 days. The specific growth rate (micro(x)) in the exponential phase was 0.576 day(-1), the yield factor (Y(X/S)) 0.418, and the productivity (P(X)) 0.480 g.l(-1).day(-1). This specific growth rate was higher than that observed by other authors during fermentation processes with other Pisolithus isolates. The method seems to be very suitable for biomass production of this fungus. However, new studies on the fungus growth morphology in this system, as well as on the efficiency of the process for the cultivation of other ectomycorrhizal fungi, are necessary. It is also necessary to test the infectivity and efficiency of the inoculum towards the hosts. PMID- 12111144 TI - Dietary-fiber-degrading enzymes from a human intestinal Clostridium and their application to oligosaccharide production from nonstarchy polysaccharides using immobilized cells. AB - The secretion of nonstarchy polysaccharide-degrading enzymes from an anaerobic human intestinal bacterium, Clostridium butyricum- beijerinckii (isolated from human feces), was investigated. Growth of the bacterium was found when laminarin, konjac glucomannan, and pectic acid were added separately to the culture media as sole carbon source. The corresponding degrading enzymes for these dietary fibers, laminarinase (endo-1,3- beta-glucanase), endo-1,4-beta-mannanase, endo- and exo pectate lyases, and pectin methylesterase, were then purified and characterized. These extracelluar enzymes, which were secreted by the bacterium in the human large intestine, were considered to contribute to digestion of the ingested dietary fibers to their oligosaccharides, following by short-chain fatty acid fermentation by the bacterium. We have developed cell immobilization techniques of the bacterium on cellulose-foam carriers that are effective for continuous production of the oligosaccharides from the dietary fibers in a fed-batch reactor system. From 9 g of pectic acid, a total of 3.96 g of 4,5-unsaturated digalacturonic acid was produced over 40 h in four 500-ml batchcultures. In the same manner, the corresponding oligosaccharides were obtained from konjac glucomannan and laminarin with average conversion rates of around 30-40%. PMID- 12111145 TI - Molecular cloning and the biochemical characterization of two novel phytases from B. subtilis 168 and B. licheniformis. AB - A novel phytase gene ( phyL) was cloned from Bacillus licheniformis by multiple steps of degenerate and inverse PCR. The coding region of the phyL gene was 1,146 bp in size and a promoter region of approximately 300 bp was identified at the upstream sequence. This gene, together with a phytase gene ( 168phyA) identified in the B. subtilis strain 168 genome by a homology search, was cloned and over expressed in B. subtilis using a phi105MU331 prophage vector system. Up to 35 units of phytase/ml were secreted into the culture media; and mature enzymes of around 44-47 kDa were purified for characterization. Both phytases exhibited broad temperature and pH optima and showed high thermostability. Of the two, the phytase encoded by phyL exhibited higher thermostability, even at a lower calcium concentration, as it was able to recover 80% of its original activity after denaturation at 95 degrees C for 10 min. With their neutral pH optima and good temperature stabilities, these Bacillus phytases are good candidates for animal feed applications and transgenic studies. PMID- 12111146 TI - Purification and characterisation of a novel laccase from the ascomycete Melanocarpus albomyces. AB - A novel laccase from the ascomycete Melanocarpus albomyces was purified and characterised. The enzyme was purified using anion exchange chromatography, hydrophobic interaction chromatography and gel filtration, and the purified laccase was biochemically characterised. It had activity towards typical substrates of laccases including 2,2'-azinobis-(3-ethylbenzthiazoline-6 sulphonate), dimethoxyphenol, guaiacol, and syringaldazine. The laccase showed good thermostability and it had a pH optimum at neutral pH, both unusual properties for most known fungal laccases. The activity of the laccase from M. albomyces was highest at 60-70 degrees C. With guaiacol and syringaldazine the pH optima were rather broad: 5-7.5 and 6-7, respectively. It retained 50% of its activity after 5 h incubation at 60 degrees C. The molecular weight of the laccase was about 80 kDa and the isoelectric point 4.0. The ultraviolet-visible absorption and electron paramagnetic resonance spectra of the purified laccase indicated that the typical three types of copper were present. PMID- 12111147 TI - Influence of threonine exporters on threonine production in Escherichia coli. AB - Threonine production in Escherichia coli threonine producer strains is enhanced by overexpression of the E. coli rhtB and rhtC genes or by heterologous overexpression of the gene encoding the Corynebacterium glutamicum threonine excretion carrier, thrE. Both E. coli genes give rise to a threonine-resistant phenotype when overexpressed, and they decrease the accumulation of radioactive metabolites derived from [(14)C] L-threonine. The evidence presented supports the conclusion that both RhtB and RhtC catalyze efflux of L-threonine and other structurally related neutral amino acids, but that the specificities of these two carriers differ substantially. PMID- 12111148 TI - Construction of self-disruptive Bacillus megaterium in response to substrate exhaustion for polyhydroxybutyrate production. AB - In order to establish a novel recovery system for polyhydroxyalkanoates, a self disruptive strain of Bacillus megaterium that responds to substrate exhaustion was constructed. A gene cassette carrying the lysis system of Bacillus amyloliquefaciens phage - holin and endolysin - was inserted into the Escherichia coli- Bacillus subtilis shuttle vector pX under the control of a xylose-inducible expression system, xylR-xylA '. In this system, the expression of a target gene is induced by xylose but inhibited by glucose, which acts as an anti-inducer. B. megaterium was transformed with pX conveying the phage lysis system, which was integrated into the amyE locus of chromosomal DNA of B. megaterium by homologous recombination. The lysis system caused self-disruption of the transformant cells effectively even when expression of the lysis genes was induced during stationary phase. For the production of polyhydroxybutyrate (PHB), the transformant was grown in a medium containing glucose as a substrate in the presence of xylose. When the glucose concentration approached zero, self-disruption was spontaneously induced, releasing intracellularly accumulated PHB into the culture broth. This system realizes timely cell disruption immediately after the PHB content in the cell reaches a maximum level. PMID- 12111149 TI - Bioengineered emulsans from Acinetobacter calcoaceticusRAG-1 transposon mutants. AB - Transposon mutants of Acinetobacter calcoaceticus strain RAG-1 were studied in an effort to control fatty acid (FA) substitution patterns of emulsan, a bioemulsifier secreted by the organism. The disrupted genes, involved in the biosynthetic pathways of biotin, histidine, cysteine or purines, influenced the level and types of FAs incorporated into emulsan. The structural variants of emulsan generated by the transposon mutants were characterized for yield, FA content, molecular weight, and emulsification behavior when grown on a series of FAs of different chain lengths from C11 to C18. Yields of emulsan from the transposon mutants were found to be lower than the parent strain and depended on the type of FA used to supplement the growth medium. Mutants 13D (His-) and 52D (Cys-) grown on LB plus C16 or C14, respectively, exhibited enhanced emulsifying activity compared to A. calcoaceticus RAG-1. The presence and composition of long chain FAs on the polysaccharide backbone influenced emulsification behavior: particularly a high mole percentage of C16 (48%) and C18 (42%). The results provide important insight into the bioengineering of bioemulsifier-producing microorganisms and provide a path towards highly tailored novel amphipathic structures to utilize as biodegradable in environmental, biomedical, and personal care applications. PMID- 12111150 TI - Metabolic engineering of Saccharomyces cerevisiae for production of novel lipid compounds. AB - The yeast Saccharomyces cerevisiae has been modified successfully for production of numerous metabolites and therapeutic proteins through metabolic engineering, but has not been utilized to date for the production of lipid-derived compounds. We developed a lipid metabolic engineering strategy in S. cerevisiae based upon culturing techniques that are typically employed for studies of peroxisomal biogenesis; cells were grown in media containing fatty acids as a sole carbon source, which promotes peroxisomal proliferation and induction of enzymes associated with fatty acid beta-oxidation. Our results indicate that growth of yeast on fatty acids such as oleate results in extensive uptake of these fatty acids from the media and a subsequent increase in total cellular lipid content from 2% to 15% dry cell weight. We also show that co-expression of plant fatty acid desaturases 2 and 3 ( FAD2 and FAD3), using a fatty acid-inducible peroxisomal gene promoter, coupled the processes of fatty acid uptake with the induction of a new metabolic pathway leading from oleic acid (18:1) to linolenic acid (18:3). Finally, we show that cultivation of yeast cells in the presence of triacylglycerols and exogenously supplied lipase promotes extensive incorporation of triglyceride fatty acids into yeast cells. Collectively, these results provide a framework for bioconversion of low-cost oils into value-added lipid products. PMID- 12111151 TI - Introduction of the carbohydrate-activated promoter P(malK) for recombinant protein production. AB - A production protocol for the use of the malK promoter was established. The protocol includes two phases: an initial fed-batch phase on glucose to reach a high cell density and a fed-batch phase on maltose for production of the desired recombinant protein. It is suggested that this cultivation scheme could be used for all promoters that are catabolite repressed by glucose and where growth and production need to be separated. The specific feature of this system is shown by its ability to control the rate of synthesis of the product protein, ss galactosidase. In the production phase with a constant feed or an exponential feeding of 0.1 h(-1) it took 4 h longer to reach the maximum specific production rate than with the higher dilution rates of 0.25 h(-1) and 0.4 h(-1), respectively. In the above experiments a dilution rate of 0.3 h(-1) in the growth phase was used. The volumetric production of this system could furthermore be extended to 40 h. All protocol procedures so far tested resulted in the same maximum production rate, but reached in different lengths of time. It is argued that this system is particularly well suited for the production of proteins that have a complex structure and/or need to be produced in a soluble form or to be exported to the periplasm. PMID- 12111152 TI - High-level production of TaqI restriction endonuclease by three different expression systems in Escherichia coli cells using the T7 phage promoter. AB - Three different expression systems were constructed for the high-level production of TaqI restriction endonuclease in recombinant Escherichia colicells. In system [R], the TaqI endonuclease gene was cloned and expressed under the control of the strong T7 RNA polymerase promoter. To protect cellular DNA, methylase protection was provided by constitutive co-expression of TaqI methylase activity either by cloning the TaqI methylase gene on a second plasmid (system [R,M]) or by constructing a recombinant plasmid harboring both the endonuclease and methylase genes (system [R+M]). In batch shake flasks containing complex media, co expression of the methylase gene in systems [R,M] and [R+M] resulted in a 2- and 3-fold increase in volumetric productivity over system [R], yielding activities of 250x10(6) U l(-1) and 350x10(6) U l(-1), which were 28 and 39 times higher than the data in the literature, respectively. Under controlled bioreactor conditions in chemically defined medium, co-expression of methylase activity greatly improved the yield and specific TaqI endonuclease productivity of the recombinant cells, and reduced acetic acid excretion levels. System [R,M] is preferable for high expression levels at longer operation periods, while system [R+M] is well-suited for high expression levels in short-term bioreactor operation. PMID- 12111153 TI - Expression of a gene for Mn-peroxidase from Coriolus versicolor in transgenic tobacco generates potential tools for phytoremediation. AB - In efforts aimed at the detoxification of contaminated areas, plants have many advantages over bacteria and fungi. We are attempting to enhance the environmental decontamination functions of plants by transferring relevant genes from microorganisms. When the gene for Mn-peroxidase (MnP) from Coriolus versicolor was expressed in transgenic tobacco plants, one line (designated fMnP21) expressed MnP activity at levels 54-fold higher than in control lines. When undamaged roots of transgenic plants were applied to liquid medium supplemented with 250 microM pentachlorophenol (PCP), the decrease in the level of PCP in fMnP21 (86% reduction) was about 2-fold higher than that in control lines (38% reduction). Expression of the gene for MnP in the transgenic plants had no obvious negative effects on their vegetative and sexual growth. Our system should contribute to the development of novel methods for the removal of hazardous chemicals from contaminated environments using transgenic plants. PMID- 12111155 TI - Cell surface-engineered yeast with ability to bind, and self-aggregate in response to, copper ion. AB - In order to construct a cell surface-engineered yeast Saccharomyces cerevisiae that facilitates adsorption and recovery of heavy metal ions, we endowed it with the ability to self-aggregate in response to binding and accumulation of copper ion. A fusion gene for the expression of GTS1, which encodes a putative zinc finger transcription factor related to occurrence of cell-aggregation, was constructed under the control of the copper ion-inducible CUP1 promoter from the yeast metallothionein gene. The multicopy plasmid carrying the fusion gene was introduced into a cell surface-engineered yeast displaying histidine hexa peptide, which can chelate copper ion. This transformant strain aggregated in medium only in the presence of copper ion, with aggregation induced by as little as 1 mM copper ion. The copper ion-induced aggregation did not interfere with the copper ion-adsorbing function of the cell surface-engineered yeast, indicating that this transformant strain has the twin features of enhanced cell surface adsorption of copper ion and self-aggregation in response to environmental copper ion. PMID- 12111154 TI - Production of active bovine cathepsin C (dipeptidyl aminopeptidase I) in the methylotrophic yeast Candida boidinii. AB - The heterologous production of active bovine cathepsin C (CTC; dipeptidyl aminopeptidase I) was investigated. Attempts to express CTC in Escherichia coli were hampered by formation of inclusion bodies that were partially degraded. To overcome this impediment, secretion of recombinant CTC was attempted in the methylotrophic yeast Candida boidinii. A DNA fragment encoding bovine procathepsin C was synthesized based on preferred codon usage in C. boidinii and placed downstream of the C. boidinii proteinase A signal sequence resulting in secretion of active CTC into the culture medium. The gene was expressed under the control of the methanol-inducible formate dehydrogenase gene promoter. Production levels were significantly improved by using a protease-deficient strain, changing medium composition, and by lowering the temperature of induction. When the recombinant C. boidinii was grown for 90 h in a jar-fermenter, active CTC was secreted with a yield of up to approximately 12 mg/l. PMID- 12111156 TI - Sinorhizobium fredii isolates can be specifically identified by a 260 bp fragment from the nolXWBTUV locus. AB - A pair of primers homologous to the nolXWBTUV locus generated a 260 bp fragment by PCR only in the presence of Sinorhizobium fredii template DNA of different quality. This resulted in a fast and accurate method for the identification of S. fredii either from pure DNA, whole bacterial cells or nodule extracts. By means of two PCR fragments, one specific for S. fredii (260-bp) and the other specific for Bradyrhizobium japonicum (RSalpha), we found that S. fredii strain SMH12 and B. japonicum E109 were equally efficient at developing nodules on soybean plants grown under controlled environmental conditions. PMID- 12111157 TI - Nitrate regulation of alpha-aminoadipate reductase formation and lysine inhibition of its activity in Penicillium chrysogenum and Acremonium chrysogenum. AB - alpha-Aminoadipate reductase (alpha-AAR) is a key enzyme in the branched pathway for lysine and beta-lactam biosynthesis of filamentous fungi since it competes with alpha-aminoadipyl-cysteinyl-valine synthetase for their common substrate L alpha-aminoadipic acid. The alpha-AAR activity in two penicillin-producing Penicillium chrysogenum strains and two cephalosporin-producing Acremonium chrysogenum strains has been studied. The alpha-AAR activity peaked during the growth-phase preceding the onset of antibiotic production, which coincides with a decrease in alpha-AAR activity, and was lower in high penicillin- or cephalosporin-producing strains. The alpha-AAR required NADPH for enzyme activity and could not use NADH as electron donor for reduction of the alpha-aminoadipate substrate. The alpha-AAR protein of P. chrysogenum was detected by Western blotting using anti-alpha-AAR antibodies. The mechanism of lysine feedback regulation in these two filamentous fungi involves inhibition of the alpha-AAR activity but not repression of its synthesis by lysine. This is different from the situation in yeasts where lysine feedback inhibits and represses alpha-AAR. Nitrate has a strong negative effect on alpha-AAR formation as shown by immunoblotting studies of alpha-AAR. The nitrate effect was reversed by lysine. PMID- 12111158 TI - Effect of medium composition, flow rate, and signaling compounds on the formation of soluble extracellular materials by biofilms of Chromobacterium violaceum. AB - Biofilms of the homoserine-lactone-defective strain Chromobacterium violaceum CV026 were grown on silicone surfaces in a flow chamber. The effect of medium flow rate, different levels of N-butanoyl-homoserine lactone, and nutrients on biofilm activity was studied by quantifying the proteins and exopolysaccharides attached to the cells. To compare the effect of each of the three variables within a wide range, the experiments were designed as a full factorial search with three levels for each variable. Calculated contour plots demonstrated that N acyl homoserine lactone is an important determinant of both the amount and the composition of the compounds excreted into the medium. PMID- 12111159 TI - Degradation of chlorophenols by Phanerochaete chrysosporium: effect of 3,4 dichlorophenol on extracellular peroxidase activities. AB - Extracellular peroxidases play an important role in the degradation of chlorophenols by Phanerochaete chrysosporium. Depending on the moment of 3,4 dichlorophenol addition, the production of lignin peroxidase and manganese peroxidase in C-limited agitated cultures was affected in opposite ways. In cultures that received 3,4-dichlorophenol at the time of inoculation, fungal growth was reduced and peroxidases were not produced, whereas peroxidase activities were stabilized after 3,4-dichlorophenol addition to pregrown cultures. Further investigation revealed that mRNA encoding lignin peroxidase was not produced in cultures started with 3,4-dichlorophenol, suggesting that the onset of secondary metabolism was affected. In addition, the stabilization of lignin peroxidase activity was not the result of an activation of lignin peroxidase gene transcription, as shown by Northern blot experiments, but likely due to the inhibition of peroxidase degradation by extracellular proteases. PMID- 12111161 TI - Quantitative determination of the spatial distribution of pure- and mixed-strain immobilized cells in gel beads by immunofluorescence. AB - A new method was developed to detect and quantify two strains, Lactococcus lactis subsp. lactis biovar. diacetylactis MD and Bifidobacterium longum ATCC 15707, immobilized separately and co-immobilized in gel beads, using specific polyclonal antibodies and confocal laser-scanning microscopy. The establishment of biomass concentration profiles for each strain was measured during colonization of beads using successive pH-controlled batch fermentations. Growth occurred preferentially in 200- and 300-microm peripheral layers of the beads for L. diacetylactis and B. longum, respectively. Repeated-batch cultures with immobilized cells permitted the production of a mixed culture containing a non competitive strain of bifidobacteria, as a result of immobilized-cell growth and high cell-release activity from the beads. During co-immobilized fermentations, there were no apparent interactions between the strains. PMID- 12111160 TI - Effect of glucose on glycerol bioconversion by Lactobacillus reuteri. AB - The impact of glucose on glycerol metabolism, especially on 3 hydroxypropionaldehyde (3-HPA) accumulation by resting cells of Lactobacillus reuteri has been investigated. Two systems were used in the study: MRS(-) (modified MRS - omitting glucose, acetate and Tween 80) and distilled water (H(2)O). In MRS(-), addition of glucose enhanced glycerol metabolism in resting cells of L. reuteri, consequently increasing the accumulation of 3-HPA by regulating the NAD/NADH ratio. Enhanced glycerol metabolism correlated positively with the concentration of glucose. NADH produced during glucose metabolism was preferentially reoxidized to NAD by the reduction of 3-HPA to 1,3-propanediol; an adequate supply of glycerol therefore outweighed the repression of glucose on the accumulation of 3-HPA. At a molar ratio of glucose to glycerol no greater than 0.33, accumulation of 3-HPA was favored. In non-growing medium (H(2)O), addition of glucose seemed to be counter-productive with respect to 3-HPA accumulation. Lactate had a positive impact on glycerol metabolism, presumably by altering the redox flux, resulting in enhanced 3-HPA accumulation in both MRS(-) and H(2)O systems. PMID- 12111162 TI - Use of carbon and energy balances in the study of the anaerobic metabolism of Enterobacter aerogenes at variable starting glucose concentrations. AB - The anaerobic metabolism of Enterobacter aerogenes was studied in batch culture at increasing initial glucose levels (9.0< S(o) <72 g l(-1)). The ultimate concentrations of fermentation products were utilized to check a metabolic flux analysis based on simple carbon mass and energy balances that promise to be suitable for the study of different fermentation processes, either under aerobic or anaerobic conditions. The stoichiometric coefficients of products collected at increasing starting glucose concentrations under anaerobic conditions suggest: (a) little influence of starting glucose level on the formation of the main fermentation products (2,3-butanediol and ethanol); (b) possible inhibition of 2,3-butanediol and lactate formations by increased ethanol concentration; (c) consequent increase in carbon flux through the remaining metabolic pathways with increased molar productions of succinate, acetate and hydrogen; (d) relative constancy of the molar production of ATP and CO(2). PMID- 12111163 TI - Fed-batch cultivation of baker's yeast followed by nitrogen or carbon starvation: effects on fermentative capacity and content of trehalose and glycogen. AB - An industrial strain of Saccharomyces cerevisiae (DGI 342) was cultivated in fed batch cultivations at a specific growth rate of 0.2 h(-1). The yeast was then exposed to carbon or nitrogen starvation for up to 8 h, to study the effect of starvation on fermentative capacity and content of protein, trehalose and glycogen. Nitrogen starvation triggered the accumulation of trehalose and glycogen. After 8 h of starvation, the content of trehalose and glycogen was increased 4-fold and 2-fold, respectively. Carbon starvation resulted in a partial conversion of glycogen into trehalose. The trehalose content increased from 45 to 64 mg (g dry-weight)(-1), whereas the glycogen content in the same period was reduced from 55 to 5 mg (g dry-weight)(-1). Glycogen was consumed faster than trehalose during storage of the starved yeast for 1 month. Nitrogen starvation resulted in a decrease in the protein content of the yeast cells, and the fermentative capacity per gram dry-weight decreased by 40%. The protein content in the carbon-starved yeast increased as a result of starvation due to the fact that the content of glycogen was reduced. The fermentative capacity per gram dry-weight was, however, unaltered. PMID- 12111164 TI - Alkanotrophic Rhodococcus ruber as a biosurfactant producer. AB - In this report we examined the structure and properties of surface-active lipids of Rhodococcus ruber. Most historical interest has been in the glycolipids of Rhodococcus erythropolis, which have been extensively characterised. R. erythropolis has been of interest due to its great metabolic diversity. Only recently has the metabolic potential of R. ruber begun to be explored. One major difference in the two species is that most R. ruber strains are able to oxidise the gaseous alkanes propane and butane. In preparation for investigation of the effects of gas metabolism on biosurfactant production, we set out to characterise the biosurfactants produced during growth on liquid n-alkanes and to compare these with R. erythropolis glycolipids. PMID- 12111165 TI - Biodesulfurization of benzothiophene and dibenzothiophene by a newly isolated Rhodococcus strain. AB - Rhodococcus sp. KT462, which can grow on either benzothiophene (BT) or dibenzothiophene (DBT) as the sole source of sulfur, was newly isolated and characterized. GC and GC-MS analyses revealed that strain KT462 has the same BT desulfurization pathway as that reported for Paenibacillus sp. A11-2 and Sinorhizobium sp. KT55. The desulfurized product of DBT produced by this strain, as well as other DBT-desulfurizing bacteria such as R. erythropolis KA2-5-1 and R. erythropolis IGTS8, was 2-hydroxybiphenyl. A resting cells study indicated that this strain was also able to degrade various alkyl derivatives of BT and DBT. PMID- 12111166 TI - An arming yeast with the ability to entrap fluorescent 17beta-estradiol on the cell surface. AB - We constructed a novel surface-engineered yeast displaying the ligand-binding domain of the rat estrogen receptor (ERLBD). ERLBD, display of which on the yeast cell surface was confirmed by immunofluorescence, possessed strong binding activity to fluorescent 17beta-estradiol - an analogue of the natural ligand of the estrogen receptor - that was comparable to the activity of the native receptor. Environmental homeostasis has recently been disturbed by endocrine disruptors, which cause confusion in the hormone secretion system. It is therefore very important to identify chemical compounds with hormone-like activity and remove them from the environment. The present results demonstrate that the new arming yeast displaying ERLBD on its cell surface will be capable of screening, entrapping, and removing estradiol-like compounds from the environment. PMID- 12111167 TI - Hydraulic selection pressure-induced nitrifying granulation in sequencing batch reactors. AB - The effect of hydraulic selection pressure on the development of nitrifying granules was investigated in four column-type sequencing batch reactors (SBR). The nature of SBR is cycle operation, thus SBR cycle time can serve as a main hydraulic selection pressure imposed on the microbial community in the system. No nitrifying granulation was observed in the SBR operated at the longest cycle time of 24 h, due to a very weak hydraulic selection pressure, while the washout of nitrifying sludge was found in the SBR run at the shortest cycle time of 3 h, and led to a failure of nitrifying granulation. Excellent nitrifying granules with a mean diameter of 0.25 mm and specific gravity of 1.014 were developed in a SBR operated at cycle times of 6 h and 12 h, respectively. The results further showed that a short cycle time would stimulate microbial activity, production of cell polysaccharides and also improve the cell hydrophobicity. These hydraulic selection pressure-induced microbial changes favour the formation of nitrifying granules. This work, probably for the first time, shows that nitrifying granules can be developed at a proper hydraulic selection pressure in terms of SBR cycle time. Nitrifying granulation is a novel biotechnology which has a great potential for wastewater nitrification. PMID- 12111168 TI - Effects of nitrite and ammonium on methane-dependent denitrification. AB - For effective application of methane-dependent denitrification (MDD) in the treatment of wastewater containing NO(2)(-) or NH(4)(+), the effect of these inorganic nitrogen compounds on MDD activity needs to be clarified. The MDD activity of sludge acclimatized with CH(4) and O(2) was determined with mineral media of different nitrogen-compound compositions in the presence of 0.21 atm CH(4) and 0.20 atm O(2). Incubations with media containing only NO(2)(-) or two of the three inorganic nitrogen compounds (NO(3)(-)+NO(2)(-), NO(2)(-)+NH(4)(+) or NH(4)(+)+NO(3)(-)) resulted in MDD activity equal to or higher than that with media containing only NO(3)(-). However, there was no MDD activity in media containing NO(2)(-) at 10 degrees C, probably because of serious inhibition of NO(2)(-) on methane oxidation. MDD occurred in media containing only NH(4)(+), although the total nitrogen removal efficiency was very low. These results show that NO(2)(-) and NH(4)(+), in the presence of NO(x)(-), do not inhibit but rather promote MDD. Consequently, NH(4)(+) does not need to be completely oxidized to NO(3)(-) in the nitrification reactor before MDD. However, under psychrophilic conditions, NO(2)(-) seriously inhibited MDD. Therefore, the nitrification reactor must not discharge effluent containing NO(2)(-) under psychrophilic conditions. PMID- 12111169 TI - Adaptation of anaerobic ammonium-oxidising consortium to synthetic coke-ovens wastewater. AB - A consortium with autotrophic anaerobic ammonium oxidising (AAAO) activity was developed from municipal sludge, and its ability to remove high ammonium concentrations in a toxic wastewater such as coke ovens wastewater is presented here. The enriched AAAO consortium was acclimatised to a synthetic coke ovens wastewater to establish anaerobic ammonium oxidation (AAO) activity. Phenol was the main carbon component of the synthetic wastewater whereby it was added stepwise from 50+/-10 to 550+/-10 mg l(-1) into an anammox enrichment medium. Ammonium-N removal was initially impaired; however, it gradually recovered. After 15 months of further selection and enrichment, the ammonium removal rate reached 62+/-2 mg NH(4)(+)-N l(-1) day(-1), i.e. 1.5 times the rate in the original AAAO reactor. The new consortium demonstrated higher ammonium and nitrite removal rates, even under phenol perturbation (up to 330+/-10 mg l(-1)). It is therefore concluded that the AAO activity in the consortium was resistant to high phenol and has potential for treating coke-ovens wastewater. PMID- 12111170 TI - Evaluation of white-rot fungi for detoxification and decolorization of effluents from the green olive debittering process. AB - Wastewater produced by the debittering process of green olives (GOW) is rich in polyphenolics and presents high chemical oxygen demand and alkalinity values. Eight white-rot fungi ( Abortiporus biennis, Dichomitus squalens, Inonotus hispidus, Irpex lacteus, Lentinus tigrinus, Panellus stipticus, Pleurotus ostreatus and Trametes hirsuta) were grown in GOW for 1 month and the reduction in total phenolics, the decolorization activity and the related enzyme activities were compared. Phenolics were efficiently reduced by P. ostreatus (52%) and A. biennis (55%), followed by P. stipticus (42%) and D. squalens (36%), but only P. ostreatus had high decolorization efficiency (49%). Laccase activity was the highest in all of the fungi, followed by manganese-independent peroxidase (MnIP). Substantial manganese peroxidase (MnP) activity was observed only in GOW treated with P. ostreatus and A. biennis, whereas lignin peroxidase (LiP) and veratryl alcohol oxidase (VAOx) activities were not detected. Early measurements of laccase activity were highly correlated ( r(2)=0.91) with the final reduction of total phenolics and could serve as an early indicator of the potential of white rot fungi to efficiently reduce the amount of total phenolics in GOW. The presence of MnP was, however, required to achieve efficient decolorization. Phytotoxicity of GOW treated with a selected P. ostreatus strain did not decline despite large reductions of the phenolic content (76%). Similarly, in GOW treated with purified laccase from Polyporus pensitius, a reduction in total phenolics which exceeded 50% was achieved; however, it was not accompanied by a decline in phytotoxicity. These results are probably related to the formation of phenoxy radicals and quinonoids, which re-polymerize in the absence of VAOx but do not lead to polymer precipitation in the treated GOW. PMID- 12111171 TI - Effects of 2,4-dichlorophenol on activated sludge. AB - The effects of 2,4-dichlorophenol (2,4-DCP) on both acclimated and unacclimated activated sludge were investigated in batch reactors. The IC(50) values on the basis of maximum specific growth rate ( micro(m)), percent chemical oxygen demand (COD) removal efficiency and sludge activity were found to be 72, 60 and 47 mg l( 1), respectively, for unacclimated culture. The percent COD removal efficiencies of unacclimated culture were affected adversely, even at low concentrations, whereas culture acclimated to 75 mg 2,4-DCP l(-1) could tolerate about 200 mg 2,4 DCP l(-1)on the basis of COD removal efficiency. Although yield coefficient values of unacclimated culture increased surprisingly to very high values with the addition of 2,4-DCP, a linear decrease with respect to 2,4-DCP concentrations was observed for acclimated culture. Although no removal was observed with unacclimated culture, almost complete removal of 2,4-DCP up to a concentration of 148.7 mg l(-1) was observed with acclimated culture. It was showed that the culture could use 2,4-DCP as sole organic carbon source, although higher removal efficiencies in the presence of a readily degradable substrate were observed. Culture acclimated to 4-chlorophenol used 2,4-DCP as sole organic carbon source better than those acclimated to 2,4-DCP. PMID- 12111172 TI - Use of a two-phase partitioning bioreactor for degrading polycyclic aromatic hydrocarbons by a Sphingomonas sp. AB - A two-phase partitioning bioreactor (TPPB) utilizing the bacterium Sphingomonas aromaticivorans B0695 was used to degrade four low molecular weight (LMW) polycyclic aromatic hydrocarbons (PAHs). The TPPB concept is based on the use of a biocompatible, immiscible organic solvent in which high concentrations of recalcitrant substrates are dissolved. These substances partition into the cell containing aqueous phase at rates determined by the metabolic activity of the cells. Experiments showed that the selected solvent, dodecane, could be successfully used in both solvent extraction experiments (to remove PAHs from soil) and in a TPPB application. Further testing demonstrated that solvent extraction from spiked soil was enhanced when a solvent combination (dodecane and ethanol) was used, and it was shown that the co-solvent did not significantly affect TPPB performance. The TPPB achieved complete biodegradation of naphthalene, phenanthrene, acenaphthene and anthracene at a volumetric consumption rate of 90 mg l(-1) h(-1) in approximately 30 h. Additionally, a total of 20.0 g of LMW PAHs (naphthalene and phenanthrene) were biodegraded at an overall volumetric rate of 98 mg l(-1) h(-1) in less than 75 h. Degradation rates achieved using the TPPB and S. aromaticivorans B0695 are much greater than any others previously reported for an ex situ PAH biodegradation system operating with a single species. PMID- 12111173 TI - Detection and characterization of erythromycin-resistant methylase genes in Gram positive bacteria isolated from poultry litter. AB - The epidemiology of four erythromycin-resistant methylase ( erm) genes, ermA, ermB, ermC and msrA, was determined in erythromycin-resistant staphylococci, enterococci and streptococci isolated from poultry litter. All isolates were resistant to multiple antibiotics. Southern hybridization indicated that 4 of the 20 staphylococci contained the ermC gene on plasmids: on a 2.2 kb plasmid in Staphylococcus hominis and S. sciuri, on a 6.0 kb plasmid in S. xylosus, and on a 7.0 kb plasmid in S. lentus. In 16 of the 20 staphylococci, the ermA gene was harbored exclusively on the chromosome, as a double chromosomal insert on 8.0 and 6.2 kb EcoRI fragments. None of the staphylococci harbored the msrA gene. Dot blot analysis indicated that all enterococci and streptococci hybridized with a biotinylated ermB gene probe. Southern hybridization indicated that only 2 of the 19 erythromycin-resistant enterococci contained the ermB gene on plasmids. The gene was localized on 4.0 kb and 5.9 kb plasmids, respectively, in two Enterococcus faecium isolates. Results from our studies indicate that the patterns of occurrence of erm genes, the sizes of the plasmids and the copy numbers of the inserts were different from the existing information on the presence of erm genes in clinical strains of Staphylococcus spp. PMID- 12111174 TI - Nickel accumulation and nickel oxalate precipitation by Aspergillus niger. AB - A strain of Aspergillus niger isolated from a metal-contaminated soil was able to grow in the presence of cadmium, chromium, cobalt, copper, and unusually high levels of nickel on solid (8.0 mM) and in liquid (6.5 mM) media. This fungus removed >98% of the nickel from liquid medium after 100 h of growth but did not remove the other metals, as determined by inductively coupled plasma spectroscopy. Experiments with non-growing, live fungal biomass showed that nickel removal was not due to biosorption alone, as little nickel was bound to the biomass at the pH values tested. Furthermore, when the protonophore carbonyl cyanide p-(trifluoremetoxy) phenyl hydrazone (FCCP) was added to the actively growing fungus nickel removal was inhibited, supporting the hypothesis that energy metabolism is essential for metal removal. Analytical electron microscopy of thin-sectioned fungal biomass revealed that metal removed from the broth was localized in the form of small rectangular crystals associated with the cell walls and also inside the cell. X-ray and electron diffraction analysis showed that these crystals were nickel oxalate dihydrate. PMID- 12111175 TI - The role of antisense oligonucleotides in the treatment of bladder cancer. AB - Both intravesical and systemic chemotherapy are limited in their efficacy in the treatment of bladder cancer patients. These limitations are centred around an inability to induce apoptosis in bladder tumour cells. This resistance to apoptosis induction is commonly associated with the overexpression of antiapoptotic proteins such as Bcl-2. Strategies to decrease the cellular expression of such proteins would enhance chemotherapy effectiveness. One such strategy is to use antisense oligonucleotides which are short sequence specific single stranded DNA or RNA molecules designed to bind to the RNA of the target protein. By binding to the target RNA, protein production is interrupted and target protein levels decease. When used to target antiapoptotic proteins, antisense oligonucleotides can therefore be used as a pre-treatment before chemotherapy to help chemosensitise the tumour cell. This review outlines the rationale for this strategy and the work done to date with antisense oligonucleotides in bladder cancer. PMID- 12111176 TI - New method: the intravital videomicroscopic characteristics of the microcirculation of the urinary bladder in rats. AB - Intravital fluorescence microscopy (IVM) is a widely used method to study the microcirculation in several organs. Our aim was to develop a standard rat model to evaluate the microcirculatory characteristics of the urinary bladder under physiological pressure conditions using the most advanced fluorescence videomicroscopic techniques. Spraque-Dawley rats were used after filling their bladders with a constant volume of saline solution. The intravesical pressure was continuously monitored. The bladder was positioned on a specially designed stage and IVM measurements were made at the beginning, the 90th, 120th and 180th min. Arteriolar and venular diameters, functional capillary density, venular red blood cell velocity, arteriolar and venular macromolecular leakage and leukocyte endothelial cell interactions (observation of rolling and firmly adherent leukocytes) were quantitatively assessed by a computer assisted analysis system. Neither microcirculatory parameters nor the intravesical pressure changed significantly during the observation period of 180 min using constant filling volume. We successfully established a new, well functioning and reproducible model to study the microcirculation of the rat bladder using intravital fluorescent microscopy. PMID- 12111177 TI - Diverse patterns of acid-base abnormalities associated with a modified sigmoid neobladder. AB - In this study, we analyzed the pattern of metabolic acidosis in patients following the construction of a sigmoid neobladder and then search for the risk factors which affecting this. In 23 men aged 43-73 years and nine women aged 49 74 years who underwent sigmoid neobladder surgery, we performed physical examinations and blood tests every 3 months for 13-75 months (38.7+/-16.6: mean+/ SD). We monitored acid-base balance, serum electrolytes, creatinine, lipid and liver function in patients for up to 6 years postoperatively. Creatinine clearance over 24 h was determined preoperatively. According to pH and base excess measured during follow-up, patients were classified into three groups (normal, 17 patients; temporary acidosis, eight patients; persistent acidosis, seven patients). Patients with temporary acidosis could compensate spontaneously by 1 year without being given sodium bicarbonate; those with persistent acidosis could not compensate spontaneously and five of them required medication with sodium bicarbonate after 1 year. Serum creatinine in patients with persistent acidosis was consistently higher during follow-up than in the other two groups. Preoperative creatinine clearances in the normal, temporary, and persistent groups were 94.25+/-27.47, 95.19+/-18.63, and 69.18+/-16.18 ml/min/1.73 m(2), respectively, being significantly lower in the persistent group ( P<0.05). In this group, patients with creatinine clearances less than 70 ml/min/1.73 m(2) could not compensate for metabolic acidosis. Normal and temporary groups showed different changes of serum chloride and bicarbonate during follow-up (respectively higher and lower) although the renal functions of the two groups were similar. Chloride and bicarbonate varied reciprocally with pH and base excess. Temporal hyperchloremic metabolic acidosis was observed until a year after surgery. In conclusion, temporary acidosis can be caused in some patients in spite of normal renal function, although it is difficult to predict it. In addition, careful follow-up is required, especially in patients with a creatinine clearance <70 ml/min/1.73 m(2) who can encounter persistent acidosis. PMID- 12111178 TI - Elevation of urinary IL-1beta levels during transplant associated nephropathy in rat model of the nephrotic syndrome. AB - A recurrence of nephrotic syndrome is a well-known phenomenon in patients who receive kidney transplantation. In this study, we attempt to establish a rat model of recurrent nephrotic change after renal transplantation using puromycin aminonucleoside (PA) induced nephrotic rats. We then examine the mechanism leading to recurrence. Female Sprague-Dawley rats (8 weeks) were divided into four groups: group A, allogenic renal transplantation of a normal kidney to PA induced nephrotic rats; group B, PA injection only as a nephrotic control; group C, allogenic transplantation of a normal kidney to rats receiving a saline injection; group D, rats receiving only a saline injection. The serum and urinary levels of protein, tumor necrosis factor (TNF)-alpha, interleukin (IL)-1beta, IL 2 and interferon (IFN)-gamma were measured. The cytokine levels were assessed by performing enzyme-linked immunosorbent assays. Six days after renal transplantation, the kidneys were excised, tissue sections were stained with hematoxylin-eosin and the specimens were studied for pathological alterations. The urinary protein levels and the histological results of protein casts in renal tubules of the autologous kidneys of the nephrotic rats and the transplanted kidneys confirmed the recurrence of nephrotic syndrome in the transplanted kidney. After renal transplantation, urinary protein and IL-1beta levels were significantly elevated in recurrent transplanted kidney groups compared with those in the control groups, while the TNF-alpha, IL-2, and IFN-gamma levels were not elevated. In addition, serum levels of TNFalpha and IL-1beta were not elevated. These results suggested that IL-1beta may relate to the recurrent nephropathy in the transplanted kidney in the nephrotic rat. PMID- 12111179 TI - Effects of amrinone on bilateral renal ischemia/reperfusion injury. AB - Renal ischemia/reperfusion injury could arise as a consequence of clinical conditions such as renal transplantation, shock, cardiac arrest, hemorrhage and renal artery surgery. In this experimental study, we aimed to determine the preventive effects of amrinone on bilateral renal ischemia/reperfusion injury in rats. A total of 60 Wistar-albino rats were divided into six groups ( n=10). Midline laparotomies were made under ketamine anesthesia. In the sham, amrinone1 and amrinone2 without ischemia (AWI1 and AWI2) groups saline, 5 and 10 mg/kg of amrinone was infused, respectively. In the ischemia, ischemia plus amrinone1 (IPA1) and ischemia plus amrinone2 (IPA2) groups, saline and 5 and 10 mg/kg of amrinone was infused, respectively, at the beginning of reperfusion, subsequent to 45 min of bilateral renal artery occlusion. Following 6 h of reperfusion, blood was drawn to study serum BUN and creatinine and a bilateral nephrectomy was done to determine tissue malonyldialdehyde ( MDA) and myeloperoxidase (MPO) levels. The results were analysed by Mann-Whitney U-test. The parameters studied were statistically higher in the ischemia group compared with the other groups ( P<0.05 for each comparison), indicating renal I/R injury. These parameters were lower in the amrinone without ischemia groups (AWI1 and AWI2) than in the sham group, however there were no significant differences between the groups ( P>0.05, for each comparison). The treatment groups IPA1 and IPA2 had statistically similar results compared with the sham group, showing the preventive effect of amrinone on renal I/R injury at the given doses. We conclude that amrinone prevented experimental renal ischemia/reperfusion injury in rats, independently of the administered doses. This preventive effect of the agent could depend on its effect of regulating the microcirculation, in decreasing intracellular calcium and in preventing neutrophil activation. We propose that this preventive effect of amrinone - which has gained clinical application especially in cases of cardiac insufficiency - could also be exploited in clinical conditions related with renal ischemia/reperfusion. PMID- 12111180 TI - Losartan attenuates renal vasoconstriction in response to acute unilateral ureteral occlusion in pigs. AB - Unilateral ureteral obstruction in pigs is associated with an enhanced, de novo generation of angiotensin II from the ipsilateral kidney. In order to further investigate the role of this system during unilateral ureter obstruction, the renal hemodynamic response to the non-peptide angiotensin II antagonist losartan was investigated. Danish land race pigs were operated on under general anesthesia. Catheters were placed in both renal veins by x-ray and ultrasonic flow probes were mounted on the renal arteries. Losartan (2 mg/kg/h) was administered intravenously to an experimental group ( n=9) continuously over 8 h of unilateral ureteral occlusion. This group was then compared to a matched control group which received only saline ( n=6). Ipsilateral pelvic pressure, renal blood flow using ultrasound transit time, glomerular filtration rate, mean arterial pressure and heart rate were measured. Renal handling of angiotensin II was examined by determining the renal extraction and secretion rates of immunoreactive angiotensin II. The anticipated reduction in ipsilateral renal blood flow after the onset of obstruction was attenuated in the losartan treated pigs, but the ipsilateral glomerular filtration rate was unaffected as compared with the controls. In the losartan group, the increase in renal vascular resistance was significantly reduced compared with un-treated controls (141+/-25% vs 217+/-24%, P<0.05). Plasma immunoreactive angiotensin II increased significantly from all three sample locations in both groups after the onset of obstruction, being more pronounced in the losartan treated group in which immunoreactive angiotensin II from the ipsilateral renal vein increased from 5.1+/-0.5 pmol/l to 41.6+/-19.6 pmol/l, P=0.027. In the controls immunoreactive angiotensin II increased from 2.7+/-0.3 pmol/l to 24.8+/-10.2 pmol/l. Furthermore, plasma aldosterone was significantly reduced after losartan administration (from 80.4 pmol/l to 36.0 pmol/l, P=0.005), indicating effective blockade of the angiotensin II type-1 receptor. The results from the present study suggest that continuous intravenous administration of losartan blocks the angiotensin II receptor mediated effects in the pig. Losartan is able to reduce ipsilateral vasoconstriction in the obstructed kidney during unilateral ureter obstruction supporting the view that angiotensin II is an important mediator of vasoconstriction during unilateral ureter obstruction in the pig model with acute unilateral occlusion of the ureter. PMID- 12111181 TI - The early effects of partial outflow obstruction on contractile properties of diabetic and non-diabetic rat bladder. AB - The aim of this study is to determine the early effects of partial outflow obstruction (POO) on the detrusor contractility of diabetic (DM) and non-diabetic rats. A total of 67 adult female Wistar rats with average weight of 214+/-3.1 g were randomized into five groups as control ( n=6), sham operated ( n=6), obstructed ( n=18), DM ( n=19), and DM with obstruction ( n=18). Intraperitoneal injection of 60 mg/kg streptozotocin was performed to achieve DM. Partial bladder neck obstruction was created surgically by ligating the urethra around a 3F feeding tube. Bladder strips were obtained and inspected on days 3, 7, and 14 of both the diabetic period and POO. Mean detrusor weights were measured and the maximal contractile responses to carbachol (Car), adenosine 5'-triphosphate (ATP), substance P (SP) and electrical field stimulation (EFS) of detrusor strips in all groups were studied in vitro. After 14 days of obstruction, no remarkable difference was observed between the maximal contractile responses to Car and SP of strips from obstructed-only (POO) and diabetic-obstructed (DM-POO) rats compared to the control group. The responses to EFS and ATP in the POO rats were significantly lower than the controls ( P<0.01, P<0.01, respectively). In the DM POO group however, the responses were significantly better than the POO group, reaching almost similar levels with the controls. The contractile responses of DM POO rats were higher than the POO group but lower than the DM group. Better contractile responses of the rats with DM-POO than POO group can be explained by the early enhancing effects of DM on detrusor contractility. In early DM+POO period, the negative effects of POO on detrusor muscle contractility is masked by diabetes mellitus. PMID- 12111182 TI - Characteristics of ureteral bolus in diabetic rats. AB - Changes in the dynamics of the ureteral bolus in diabetics was investigated by using videomicroscopic imaging. Diabetes mellitus (DM) was induced in 8-week-old male rats by the administration of streptozotocin. The pressure of the renal pelvis (via nephrostomy) and visualization of the dynamics of the ureteral bolus were recorded in three groups (DM, sucrose-fed, and control) 8 weeks after administration. Peristaltic velocity, frequency and bolus length were analyzed on the basis of image processing using indigo carmine dissolved in a saline solution. This was perfused via the nephrostomy. The dilated ureters were macroscopically observed in all diabetic rats. There were significant decreases in the velocity of the bolus in diabetics (4.3+/-0.8 mm/s) compared to sucrose diuretic (11.0+/-3.2 mm/s) and control (8.3+/-1.4 mm/s) rats. In addition, the length of the bolus in diabetics (9.3+/-1.4 mm) was about twofold longer than those of control rats (4.2+/-0.6 mm). There were also significant decreases in the frequency of renal pelvis contraction in diabetics (19.5+/-1.9 min(-1)) compared to sucrose-diuretic (25.3+/-1.0 min(-1)) and control (29.0+/-2.3 min( 1)) rats. These results indicate that the decrease in the velocity of the bolus and the frequency of renal pelvis contraction result in the disability of urine transport in the upper urinary tract in DM although the volume per bolus increases. Besides hyperglycemia, these changes may predispose diabetics to upper urinary tract infections such as pyelonephritis. PMID- 12111184 TI - Therapeutic strategy after microsurgical varicocelectomy in the modern assisted reproductive technology era. AB - In this study, we searched for prognostic factors at preoperative examination for the improvement in spermatogenesis of patients undergoing varicocelectomy. Eighty patients with varicocele testis underwent microsurgical varicocelectomy. Before surgery, the seminogram, testicular volume, varicocele grade, and serum FSH, LH, testosterone, prolactin, and estradiol were evaluated. Postoperatively, semen analysis was performed every 3 months. We assessed the associations between the preoperative variables and postoperative seminogram improvement. 0f 80 patients, 37 showed improvement, usually by 6 months. Patient age, duration of sterility, testicular volume, sperm motility, morphology, semen volume, serum LH, testosterone, prolactin, and estradiol showed little difference between responders and non-responders. A small left testis, or a grade III varicocele decreased the likelihood of improvement. Patients with a sperm count of 10-20 x 10(6)/ml were significantly more likely to respond to varicocelectomy than those with sperm counts <5 x 10(6)/ml. Patients with elevated FSH were less likely to respond, as were those with a Johnsen score below 6. Varicocelectomy alone is unlikely to improve sperm counts of patients with a sperm count below 5 x 10(6)/ml, high FSH, small left testes, or Johnsen scores below 6. In conclusion, for couples in this situation, assisted reproductive technology coupled with varicocelectomy should be proposed. PMID- 12111183 TI - Antioxidative effects of exogenous nitric oxide versus antioxidant vitamins on renal ischemia reperfusion injury. AB - The objectives of this study were to compare the protective influence of exogenous nitric oxide on renal ischemia reperfusion (I/R) injury with that of the antioxidant vitamins C and E. Sprague-Dawley rats were divided into three groups ( n=12 per group). Normal saline solution was given in group 1, a vitamin C (200 mg/kg/d) plus vitamin E (100 mg/kg/d) combination in group 2 for 3 days before operating and Na-nitroprusside (5 mg/kg/d) in group 3 before reperfusion. The left kidneys were exposed to warm ischemia for 40 min followed by reperfusion for 90 min. The right kidneys were used as internal controls. After both kidneys were removed, histopathological examinations were performed, and oxidative and antioxidative parameters were measured. In the postischemic reperfused rat kidneys, the renal lipid peroxidation level was significantly lower, and the renal GSH level higher in the group given Na-nitroprusside compared with groups 1 and 2. Renal specific xanthine oxidase activity was also significantly lower in the group treated with Na-nitroprusside than in the groups given vitamins or saline. There was a significant, negative correlation between lipid peroxidation and reduced glutathione levels. Our results suggest that the exogenous nitric oxide (Na-nitroprusside) inhibits xanthine oxidase, and has more apparent preventive features for renal I/R injury than the antioxidant vitamins C+E. PMID- 12111185 TI - Influence of pertussis toxin on superficial bladder carcinoma in rats. AB - The proliferation and migration of cells is a fundamental process for the metastasis of malignant tumour cells. In several in vitro studies, pertussis toxin (PTX) inhibited cell proliferation and cell motility in the human transitional cell carcinoma cell line J82. The present study investigated the effect of the intravesical application of PTX on the development of superficial bladder cancer in rats. We used the model of N-butyl-N-(4 hydroxybutyl)nitrosamine (BBN, 0.05% via drinking water x10 weeks) to induce superficial bladder carcinomas in 40 female rats. After 16 weeks the rats were treated in two groups with 0.4 ml PTX (1 microg/ml) or 0.4 ml phosphate buffered saline (PBS) by intravesical application (once a week for 10 weeks). In the 25th week urine cytology was determined and all rats were killed at week 26 followed by histological evaluation. In the control group, the urine cytology was positive for G2/G3 cells in ten of 17 rats. In the PTX group G2/G3 cells were determined in five of 20 rats ( P two tailed <0.05). Histopathologically 12 rats (71%) of the control group and 11 rats (55%) of the PTX group developed T1-T2 transitional cell carcinomas. No local or systemic side effects were disclosed. PTX treatment reduces the development of G3 transitional cell carcinomas in rats and may represent a new approach for local therapy in superficial bladder cancer. PMID- 12111186 TI - Variables influencing bacteriological outcome in patients with streptococcal tonsillopharyngitis treated with penicillin V. AB - Despite the fact that group A streptococci (GAS) remain susceptible to penicillin V (pen V), an increasing rate of bacteriological treatment failures has occurred. A recent study has suggested that the major variables associated with pen V treatment failures were the number of days ill prior to initiation of treatment (<2 days) and age <6 years. In order to study the link between pen V treatment failures and individual variables, we reviewed the files of all children enrolled in four randomised multicentre trials of oral antibiotic therapy, carried out from 1993 to 1999. A standard protocol and follow-up examination were used in these four studies: cultures were obtained 4 days and 1 month after completion of treatment. Total DNA restriction fragment length polymorphism was used to compare pre- and post-treatment GAS isolates. We enrolled 1560 children aged 3 to 12 years, 685 received a 10 day pen V regimen (45 mg/kg per day divided into three doses/day), among them 536 were assessable for bacteriological efficacy at the first and second follow-up visit. We found the only variable associated with penicillin treatment failure was the age of the child when infected. The rate of failure was statistically more important for children younger than 6 years (35.5%, 95% CI 29.9--41.1) than for older children (21.9%, 95% CI 16.9-26.9). CONCLUSION: in this study only young age (<6 years) increases penicillin V treatment failures for group A streptococcal tonsillopharyngitis. This may lead to different antibiotic regimens and follow-up modalities for these targeted patients. PMID- 12111187 TI - Familial isolated congenital asplenia: a rare, frequently hereditary dominant condition, often detected too late as a cause of overwhelming pneumococcal sepsis. Report of a new case and review of 31 others. AB - Congenital isolated asplenia may arise as a minor form of situs abnormalities or result from an unrelated specific defect of spleen development. It is a rare life threatening condition and pneumococcal sepsis is often the first sign of the disease. We report on the case of a deceased 11-month-old girl and her father who developed recurrent pneumococcal meningitis. The fatal evolution in the girl was due to Streptococcus pneumoniae serotype 23 with intermediate penicillin sensitivity 4 h after amoxicillin (100 mg/kg i.v.) administration. Establishing the diagnosis of congenital isolated asplenia in the case of pneumococcal sepsis can be achieved by performing two easy and non-invasive investigations: searching for Howell-Jolly bodies on blood smears and performing ultrasound examination of the abdomen to look for the spleen. In the case of congenital isolated asplenia, use of appropriate prophylaxis could save the lives of affected children. Our review of the literature yielded 31 cases of congenital isolated asplenia. Thirteen were sporadic and 18 were familial cases involving eight families. CONCLUSION: in the case of Streptococcus pneumoniae sepsis, a systematic search for Howell-Jolly bodies on blood smears and ultrasound examination of the abdomen for the presence of asplenia should be mandatory to detect isolated congenital asplenia. If asplenia is found, potentially life-saving antibiotic prophylaxis and pneumococcal vaccination should be initiated. PMID- 12111188 TI - Circulating anti-prolactin auto-antibodies must be considered in the differential diagnosis of hyperprolactinaemia in adolescents. AB - In four adolescents (two girls and two boys), who were investigated for galactorrhoea or short stature, we found moderately elevated serum prolactin (PRL) levels, but without a decreased gonadotropin and sex hormone production. Serum PRL levels were not responding to the intravenous injection of thyrotropin releasing hormone. Magnetic resonance imaging of the hypothalamic-pituitary region revealed no abnormality. Initial diagnoses were idiopathic and drug induced hyperprolactinaemia. The presence of anti-prolactin auto-antibodies was suspected because of low recovery of PRL after precipitation with polyethylene glycol and confirmed by immunoprecipitation with anti-human IgG-agarose. CONCLUSION: anti-prolactin auto-antibodies causing high prolactin values in some immunoassays can cause hyperprolactinaemia in adolescents without other signs of hormonal disturbances or auto-immune disease. PMID- 12111189 TI - Progressive neurologic disability in methylmalonic acidemia despite transplantation of the liver. AB - Methylmalonic acidemia unresponsive to cobalamin is often fatal in infancy. Patients have been considered candidates for hepatic transplantation and experience has been that the procedure eliminates the life-threatening episodes of ketoacidosis that characterize this disease. CONCLUSION: experience with a 24 year-old patient treated with hepatic transplantation indicates that this procedure does not prevent progressive renal failure and neurologic dysfunction. PMID- 12111190 TI - The accuracy of voiding urosonography in detecting vesico-ureteral reflux: a summary of existing data. AB - The primary objective of this review was to assess the diagnostic accuracy of voiding urosonography (VUS) in detecting reflux (VUR). As a secondary objective, the reported technical suggestions and diagnostic mistakes were shown to improve the examination protocol and provide the most accurate results. Using a Medline Database search, the published articles comparing the grey-scale (GS) or colour Doppler (CD) VUS with voiding cystourethrography (VCUG) as the gold standard were selected. Articles were excluded when data were not sufficient to construct 2x2 tables or when the gold standard was different from VCUG. For the analyses of diagnostic accuracy values, 95% confidence intervals were given. Agreements in the results of GSVUS and VCUG and in those of CDVUS and VCUG were determined by Kappa statistics. GSVUS and CDVUS were compared for diagnostic accuracy by the McNemar test. Results showed that the range of GSVUS sensitivity and specificity in detecting VUR was 69%-100% and 86%-97%, respectively. The agreement between GSVUS and VCUG diagnoses ranged from 90% to 97% (K score range 0.61-0.92; P<0.001). The range of CDVUS sensitivity and specificity in detecting VUR was 93% 100% and 86%-93%, respectively. The agreement between CDVUS and VCUG diagnoses ranged from 89% to 96% (K score range 0.77-0.91; P<0.001). One study comparing both VUS modalities with VCUG in the same group of patients, showed that the diagnostic accuracy of CDVUS was significantly higher than that of GSVUS (96% versus 90% of cases correctly classified; McNemar chi squared =4; P<0.05). CONCLUSION: the existing data indicate that false-negative voiding urosonographic diagnoses (8%-31%) and underestimated reflux grading cases using the same technique are related to anatomical conditions, patient cooperation and contrast medium administration. False-positive (3%-14%) and overestimated reflux grading cases using voiding urosonography could be correctly assessed cases. The intermittent nature of vesico-ureteral reflux is better detected by a technique employing a prolonged observation time, such as voiding urosonography. This might question the current role of voiding cystourethrography in the investigation of reflux. PMID- 12111191 TI - Low-volume peritoneal dialysis in 116 neonatal and paediatric critical care patients. AB - Acute renal insufficiency accounts for high mortality in paediatric intensive care patients, particularly in infants. Peritoneal dialysis, usually carried out with dialysate volumes of >20 ml/kg body weight, increases pulmonary artery pressure, which may compromise myocardial function in critical illness. In this paper we report our experiences with the use of lower dialysate volumes in the treatment of critically ill children with renal impairments. We suggest that low volume peritoneal dialysis is able to achieve adequate ultrafiltration, which relieves overhydration in ventilated and haemodynamically compromised children. A total of 116 paediatric intensive care patients treated between 1992 and 2000 was the subject of this investigation. Diagnosis, indication for dialysis, arterial and central venous pressure, blood gases, creatinine, blood urea nitrogen, urinary output at installation, ultrafiltration, fluid balance, duration and complications during dialysis as well as survival were investigated. The overall mortality was 53%. The respective diagnoses and mortality rates were as follows: 65% of the patients suffered from cardiac diseases (54% mortality), 7% from renal diseases (13%) and 28% from multi-organ system failure (62%). Low-volume peritoneal dialysis was started at evidence of total body fluid overload with inadequate urinary output and resulted in a mean ultrafiltration of 2.8 ml/kg body weight per h. A negative fluid balance was achieved in 53% of patients, mainly in those suffering from hypervolaemia and minor oliguria. None of the complications resulted in death. CONCLUSION: early installation of low-volume peritoneal dialysis offers a safe and adequate ultrafiltration procedure for paediatric critical care patients suffering from minor oliguria and fluid overload. PMID- 12111192 TI - Auto-immune pancytopenia in a child with DiGeorge syndrome. AB - We report on the development of auto-immune pancytopenia in a child with DiGeorge syndrome carrying the 22q11 microdeletion. She had congenital heart disease, dysmorphic facies, thymic hypoplasia, immunodeficiency, velopharyngeal insufficiency, scoliosis, and a hearing deficit. She had a low T-cell count with a normal CD4/CD8 ratio, IgA deficiency and a normal lymphoblastic response to mitogens. She has presented with pancytopenia since 10 years of age (leucocytes 3,300/mm(3), haemoglobin 107 g/l, platelets 80,000/mm(3)). Platelet-associated antibodies, anti-neutrophil antibodies and Coombs' positive red cells were present. At 14 years of age, she presented with a severe episode of haemolysis with pancytopenia. Steroids were effective in treating the pancytopenia at a dose of 2 mg/kg per day for 6 weeks. Since 15 years of age, she has had episodes of acrocyanosis. At 16 years of age, she still had mild pancytopenia without any treatment. CONCLUSION: the clinical spectrum of the 22q11 microdeletion syndrome is very broad. This case suggests that auto-immune disease such as pancytopenia is part of the 22q11 microdeletion syndrome. PMID- 12111193 TI - TSC1 and TSC2 mutations in tuberous sclerosis, the associated phenotypes and a model to explain observed TSC1/ TSC2 frequency ratios. AB - Tuberous sclerosis (TSC) is a multisystem disease with manifestations in the central nervous system, skin, kidneys, heart, and other visceral organs. The development of TSC is associated with alterations within a gene on chromosome 9q34 ( TSC1) and a gene on chromosome 16p13 ( TSC2). Most de-novo patients show a mutation in TSC2, whereas only 50% of all familial cases can be related to TSC2 mutations. In the present study, 68 unrelated patients with confirmed clinical manifestations of TSC were tested for mutations in the TSC1 and TSC2 genes. In total, we studied 59 sporadic cases and 9 familial cases, including one large family with TSC2 linkage. Two pathogenic mutations were found in TSC1. The TSC2 gene analysis revealed 29 mutations, including 3 large deletions and 26 small mutations, 15 of them truncating. CONCLUSION: the TSC1-TSC2 mutation ratio in our group of patients differs significantly from the 1:1 ratio previously predicted on the basis of linkage studies. There is an obvious paradox between the observed frequency of TSC1 mutations in familial cases and sporadic cases. An interestingly mild phenotype, observed in one of our TSC1 mutation carriers, led to the elaboration of a model that provides a plausible explanation for this paradox. We propose the presence of a very mildly affected patient group with TSC1-related disease who are not regularly detected by clinical diagnosis. PMID- 12111194 TI - Beta-catenin and the morphogenesis of colorectal cancer. AB - Tumor growth and progression are morphogenetic processes that are characterized by ongoing changes in tumor structure and differentiation. These processes show similarities to morphogenesis in embryonic development, as supported by the fact that the main pathways regulating morphogenesis in early embryogenesis and organogenesis are directly or indirectly altered in most neoplasms. In colorectal adenocarcinomas, different morphogenetic areas can be clearly defined. The focus of this review is on combining morphology-based aspects and recent molecular and genetic data on the progression of colorectal carcinomas. The decisive genetic alteration in most colorectal cancers is the loss of function mutation in the adenomatous polyposis coli tumor suppressor gene, leading to an accumulation of the oncoprotein beta-catenin, the main effector of the embryonic Wnt/wingless pathway. The possibility is discussed that, on the basis of this genetic alteration, the tumor microenvironment is an additional driving force of tumor progression. PMID- 12111195 TI - Pancreatic diseases past and present: a historical examination of exhibition specimens from the Collectio Rokitansky in Vienna. AB - The Viennese collection of pathological specimens (Collectio Rokitansky) comprises a large number of objects from all fields of pathological anatomy and is one of the largest historical collections in the entire world. We reviewed the original diagnoses in a series of pancreatic specimens using modern histopathological techniques. It was found that the histological structure of eleven pancreatic specimens was surprisingly well preserved. In three cases of extrapancreatic pseudocysts, we identified chronic pancreatitis as the underlying disease. Two specimens contained tumours that proved to be ductal adenocarcinomas. A third, rather large tumour was identified as a solid pseudopapillary carcinoma and a fourth one as a neuroendocrine carcinoma. The remaining cases were classified as fibrotic atrophy, congenital cysts, microcystic serous cystadenoma, and necrotic sequestration of the pancreas. The application of immunohistochemical methods failed. In conclusion, the surprisingly well-preserved exhibits from the Collectio Rokitansky, which have been on display for more than 100 years, are accessible to modern histopathological investigation without the use of immunohistochemical techniques. Such examinations allow us to assess the occurrence of diseases and tumours in the sociocultural environment of the 19th century. PMID- 12111196 TI - Ampulla of vater cancers: T-stage and histological subtype but not Dpc4 expression predict prognosis. AB - Loss of immunohistochemical expression of Dpc4 occurs in about 50% of pancreatic ductal cancers and its loss correlates with DPC4/Smad4 gene inactivation. Dpc4 expression was also lost in 6 of 16 (37%) ampulla of Vater cancers (AVCs) previously analyzed. Furthermore, chromosomal losses involving 18q, where DPC4 is located, have been observed in 34% of AVCs and are associated with decreased survival. To evaluate the possibility that expression of Dpc4 may be correlated with survival, we analyzed 89 AVCs for inactivation of DPC4 by immunohistochemical staining. Thirty-seven cases showed no expression of Dpc4 (41%). Multivariate survival analysis was performed including age, sex, tumor size, histological subtype (intestinal or pancreatobiliary), grade of differentiation, T-stage, lymph-node metastases and Dpc4 status. T-stage and histological subtype were selected as independent prognostic factors, while Dpc4 immunostaining was not significantly associated with any clinicopathological variable, including histological subtype. Although Dpc4 expression is of no clinical relevance, its involvement in AVC gives additional weight to the hypothesis that, among all pancreatic exocrine and endocrine tumors, only AVC and common ductal adenocarcinomas have similar molecular fingerprints. Moreover, comparison of the frequencies of allelic loss on chromosomal arm 18q and the loss of Dpc4 expression (34% and 41%, respectively) is highly suggestive that DPC4 is the major target of these losses. PMID- 12111197 TI - Detection of the Epstein-Barr virus in primary adenocarcinoma of the lung with Signet-ring cells. AB - The Epstein-Barr virus (EBV) is directly implicated in the pathogenesis of a variety of undifferentiated carcinomas and has also been identified in conventional adenocarcinomas of the stomach. To date, the association of EBV with non-small cell lung carcinoma is restricted to Asian patients. To evaluate the presence of EBV in lung cancers from Europeans, we investigated primary lung adenocarcinomas with a similar morphological tumour pattern to those of the stomach, specifically rare tumours with components of signet-ring cells. Three tumours of signet-ring cell type were examined by means of polymerase chain reaction (PCR). To localise the virus to the neoplastic cells, in situ hybridisation (ISH) was performed using an antisense Epstein-Barr virus encoded RNA probe. Immunohistochemistry was performed to evaluate the expression of latent membrane protein-1 (LMP-1) and EBV nuclear antigen 2 (EBNA-2). PCR investigation confirmed the presence of EBV in one case. Positive signals confined to tumour cells were present on ISH. None of the tumours showed expression of LMP-1 and EBNA-2. To our knowledge, this is the first report on the presence of EBV in primary adenocarcinoma of the lung in a Caucasian patient. The present study indicates that EBV may infect some lung cancers with a specific tumour pattern. PMID- 12111198 TI - High levels of cellular retinol binding protein-1 expression in leiomyosarcoma: possible implications for diagnostic evaluation. AB - Retinoid bioavailability is regulated by the activity of specific cytoplasmic receptors. High levels of cellular retinol binding protein-1 (CRBP-1) have been documented during experimental arterial and wound-healing processes, but data concerning neoplastic smooth muscle tissues are scarce and/or controversial. This study reports that the expression of CRBP-1 is markedly higher in uterine and gastrointestinal leiomyosarcomas than in leiomyomas and normal myometrium; CRBP-1 was practically absent in normal gastrointestinal smooth muscle tissue. CRBP-1 positivity was particularly elevated in the epithelioid variant of leiomyosarcoma; it was associated with increased proliferative and apoptotic rates and inversely related to smooth muscle differentiation evaluated by alpha- and gamma-smooth muscle actin and desmin expression. Western blotting and reverse transcription polymerase chain reaction confirmed the observations concerning CRBP-1 and actin isoform expression and revealed higher NF-kappa-Bp65 and RAR alpha and lower Bax protein levels in leiomyosarcoma than in the other conditions. These findings document that a high CRBP-1 expression is associated with smooth muscle malignancy and suggest that CRBP-1 expression represents a new useful marker for the classification of unusual variants of smooth muscle tumors. PMID- 12111199 TI - E-cadherin, E-selectin and vascular cell adhesion molecule: immunohistochemical markers for differentiation between mesothelioma and metastatic pulmonary adenocarcinoma? AB - Pleural mesotheliomas, especially pure epithelioid mesotheliomas, may histologically be easily confused with peripheral pulmonary adenocarcinomas or pleural carcinosis. As there is no specific antibody for mesotheliomas, today a panel of immunohistochemical markers is used for the differential diagnosis of these two tumour entities. In search of further significant antibodies for application onto formalin-fixed, paraffin-embedded tissue, we immunohistochemically investigated the expression pattern of three adhesion molecules: vascular cell adhesion molecule (VCAM), E-selectin and E-cadherin. A comparatively large number of 44 mesotheliomas (15 epithelioid, 15 biphasic, 14 sarcomatoid) and 18 peripheral pulmonary adenocarcinomas were analysed. While for these two tumour entities there were no significant differences of the staining patterns for VCAM and E-selectin, there were significant differences in the expression of E-cadherin: while nearly all adenocarcinomas stained positively, there was almost no staining reaction of the mesotheliomas. Therefore, E-cadherin -- in contrast to E-selectin and VCAM -- appears to be a further relevant immunohistochemical marker for the distinction between adenocarcinomas and mesotheliomas. PMID- 12111200 TI - Three-dimensional analysis of alveolar structure in usual interstitial pneumonia. AB - The etiology of usual interstitial pneumonia (UIP), a progressive lung disease, remains unclear. We examined alveolar structure in UIP three-dimensionally. Lung biopsy specimens from five patients with idiopathic pulmonary fibrosis were used. Sections 150-microm thick were stained with elastica solution for elastic fibers, with alpha-smooth muscle actin antibody for myofibroblasts, with anti-Thomsen Friedenreich antibody for type-II pneumocytes and with anti-CD34 antibody for blood vessels. We examined them three-dimensionally using a laser confocal microscope or light microscope. In the fibrotic lesions, the thick elastic fibers forming the alveolar framework were not particularly dense considering the reduction in alveolar volume. Near the fibrotic lesions, some of the thin elastic fibers in the alveolar wall were slightly sinuous and ended with rounded tips. Type-II pneumocytes had proliferated and were distributed uniformly over the alveolar surface. Smooth muscle actin filaments were detected only around the alveolar orifice. These findings show that in UIP destruction of the elastic fiber framework of the alveoli may lead to irreversible focal alveolar collapse after damage to the alveolar epithelial cells, and proliferation of type-II pneumocytes may be involved with this elastolysis. PMID- 12111202 TI - Isolated amoebic appendicitis. AB - Amoebiasis, a disease of worldwide distribution, is endemic in tropical countries with suboptimal sanitation facilities. Isolated amoebic appendicitis (IAA) is regarded as a rare manifestation of the disease globally. Because there are no defined clinical features that distinguish IAA from bacterial appendicitis, diagnosis is usually dependent on histopathological examination. A 9-year retrospective study was undertaken to investigate the clinicopathological aspects of IAA. The main complaints were fever and abdominal pain. None of the patients had dysentery. The pre-operative clinical diagnosis was acute appendicitis and acute abdomen in 13 and 8 patients, respectively. In all cases the intra operative diagnosis was acute appendicitis. Gross pathological appraisal revealed peritonitis and perforation in 19 and 17 cases, respectively. Histopathological examination of these appendices demonstrated appendiceal ulceration, transmural mixed inflammation, haematophagous amoebic trophozoites and necrosis in all cases. Vascular pathology comprised venous and capillary luminal plugging (11 cases), necrotising small vessel vasculitis (11 cases), thrombophlebitis of medium sized veins (9 cases) and arteritis with associated thrombosis (1 case). Organising fibrinopurulent peritonitis was present in 19 cases. Two appendices that appeared normal macroscopically demonstrated ulceration and inflammation that were confined to the mucosa and submucosa. All of 18 patients who were treated with metronidazole survived without further surgery, while three patients who were untreated succumbed to the disease. Appendicectomy, accurate histopathological appraisal thereof and optimal, timely management of IAA were critical to the favourable outcome in the present study. PMID- 12111201 TI - Cell proliferation and apoptosis in human uterine leiomyomas and myometria. AB - To determine the role of cell proliferation and apoptosis in uterine leiomyoma growth, we studied protein expression of two major regulatory proteins of apoptosis -- Bcl-2 (anti-apoptotic) and Bax (pro-apoptotic) -- and two endogenous markers of cell replication - proliferating cell nuclear antigen (PCNA) and Ki-67 - in tumors and matched myometrium from premenopausal women. Conventional mitotic indices also were determined, and all proliferation data were correlated to tumor size. In situ end-labeling of fragmented DNA and routine histology were used to assess apoptosis. Our results showed that the apoptosis-regulating proteins (Bcl 2 and Bax) were expressed in the cytoplasm of the leiomyoma and myometrial smooth muscle cells throughout the menstrual cycle. Bax expression differed from Bcl-2 in that it also was found in the cytoplasm of vascular smooth muscle cells of the myometria and tumors. Both tumors and myometrial samples expressed 26-kDa and 21 kDa proteins that reacted with antibodies directed towards Bcl-2 and Bax, respectively. Apoptosis was not a prominent feature of uterine leiomyomas or myometrium. PCNA- and Ki-67-labeling and mitotic counts were significantly ( P<0.05) higher in leiomyomas than in matched myometrial samples. Proliferative activity was variable for individual tumors of the same patient and independent of tumor size. Our results suggest that altered apoptosis by overexpression of Bcl-2 or by decreased expression of Bax does not appear to be a major factor in uterine leiomyoma growth. We conclude that increased cell proliferation is the most significant contributor to growth and that the proliferative state is autonomous for each tumor in a given patient and is independent of tumor size. PMID- 12111203 TI - Immunohistochemical studies in acute and chronic canine chagasic cardiomyopathy. AB - A major characteristic of Chagas' disease is a myocarditis constituted primarily of mononuclear cells, both during the acute and chronic phases of the disease. Using monoclonal antibodies and image analyses we have quantified canine CD8(+) T cells (caCD8(+) T cells), canine CD4(+) T cells (caCD4(+) T cells) and neutrophils in canine chagasic myocardiopathy induced by two strains isolated from the first human clinical case of Chagas' disease. We also evaluated the influence of tissue parasitism in the genesis of chronic myocarditis through immunohistochemistry. As in human myocarditis, there was a predominance of T lymphocytes in the inflammatory infiltrate in all animals studied. In the dogs inoculated with strain Berenice 78 (Be78) and necropsied during the acute phase of infection, we found 58% caCD8(+) and 42% caCD4(+) T cells. In chronically infected animals, 53% of T cells were represented by caCD8(+) and 47% were caCD4(+) T cells. Since normal canine lymphoid organs are constituted by 70-80% caCD4(+) T cells and 20-30% caCD8(+) T cells our results indicate a higher proliferation of caCD8(+) T cells in dogs inoculated with the Be78 strain. In chronic myocarditis induced by the Berenice 62 (Be62) strain, caCD8(+) cells constituted 33% of the T cells and 67% were caCD4(+) T cells, a proportion similar to that found in normal canine lymphoid organs. Since the Be78 strain induces greater loss of myocardiocytes than strain Be62, we believe that the caCD8(+) T cells, among other factors, can be important in the genesis of these lesions. Amastigote nests and immunohistochemically labelled Trypanosoma cruzi antigen were not found in dogs necropsied during the chronic phase. The absence of the parasite in the myocardium suggests the involvement of other mechanisms in the genesis of the inflammatory process. PMID- 12111205 TI - Gynecomastia with pseudoangiomatous stromal hyperplasia and multinucleated giant cells. Association with neurofibromatosis type 1. PMID- 12111204 TI - Cyclooxygenase-2 and Bcl-2 expression in the stomach mucosa of Wistar rats exposed to Helicobacter pylori, N'-methyl- N'-nitro- N-nitrosoguanidine and bile. AB - Cyclooxygenase-2 (COX-2) and Bcl-2 have been implicated in upper gastrointestinal tract carcinomas, but the underlying mechanisms are not known. In the present study we assessed the correlation of COX-2 and Bcl-2 to known cell kinetics in the glandular stomach mucosa of 104 Wistar rats given combinations of Helicobacter pylori, MNNG ( N'-methyl- N'-nitro- N-nitrosoguanidine) and bile. COX-2 expression, Bcl-2 and cell proliferation (Ki-67) in antral and corpus mucosa were determined immunohistochemically. Apoptotic cells were detected using terminal uridine deoxynucleotidyl nick end labelling technique. Expression of COX 2 was found in the lower portion of glandular corpus epithelium, and Bcl-2 positivity was mainly seen in the proliferative zone of both antrum and corpus mucosa. COX-2 expression in histologically normal-appearing corpus mucosa was associated with cell proliferation, atrophy and intestinal metaplasia in antrum and with Bcl-2 expression in corpus mucosa. No correlation was found between apoptosis and Bcl-2 expression. MNNG but not H. pylori significantly increased COX-2 in corpus mucosa. H. pylori, however, promoted the COX-2 expression in corpus when bile was added and Bcl-2 expression in antrum. Abnormal expression of both COX-2 and Bcl-2 seem to be involved in H. pylori-induced gastric carcinogenesis by altering the gastric cell kinetics. PMID- 12111206 TI - Vibrio vulnificus infection in patients with liver disease: report of five autopsy cases. AB - Five autopsy cases of Vibrio vulnificus infection with liver disease are reported. All five patients ate raw seafood 24 h before the onset of illness. The clinical presentation was of primary septicemia, with positive cultures in both the blood and cutaneous lesions. Stool cultures were positive for the organism in one patient with gastrointestinal symptoms. Autopsy examination revealed liver cirrhosis in three cases and alcoholic liver disease in two; all showed portal hypertension. Gastrointestinal mucosal changes were seen in four patients: edema, hemorrhagic necrosis, and lymphocyte infiltration. One case was of an human immunodeficiency virus infected patient in which histology showed a rare intestinal disease, phlegmonous colitis. We believe this is the first description of a case of concomitant phlegmonous enterocolitis and V. vulnificus infection. Patients with liver disease should be warned about the possibility of life threatening infections and complications associated with the consumption of raw seafood. PMID- 12111207 TI - Sporadic, syndromic, and Zollinger-Ellison syndrome associated fundic gland polyps consistently express cytokeratin 7. PMID- 12111210 TI - Developmental potential of fused Caenorhabditis elegans oocytes: generation of giant and twin embryos. AB - With their first cleavage blastomeres in Caenorhabditis elegans are fixed to very different developmental programs going along with differential segregation of maternal gene products. To investigate whether indications for a prelocalization of cytoplasmic components can already be found in unfertilized egg cells, we fused mature C. elegans oocytes with the help of a laser microbeam. Fertilization of two fused oocytes resulting in triploid zygotes showed an essentially normal early cleavage pattern with the establishment of five somatic cell lineages and a germline and also a normal spatial arrangement of blastomeres. A considerable fraction of such embryos hatched and developed into fertile giant nematodes. The numbers of cell nuclei in freshly hatched and adult giant animals were found to be essentially the same as in untreated controls. When three fused oocytes were fertilized, two alternative patterns of early embryogenesis were observed. Half of the embryos followed the normal cleavage mode. The other half, however, developed in a twin-like fashion with all cells present in two copies, apparently due to fertilization by two sperm. In such embryos, two areas of gastrulation were established, resulting in the generation of two separate gut primordia. In summary, our results suggest that (1) in contrast to the uncleaved zygote in the mature oocyte of C. elegans no cytoplasmic regionalization exists, (2) the invariable cell numbers typical for the C. elegans embryo are not controlled via cell size, and (3) the entry of a second sperm can induce a cascade of events in the egg leading to the formation of two complete embryo anlagen. PMID- 12111212 TI - The jing gene is required for embryonic brain development in Drosophila [corrected]. AB - Loss- and gain-of-function studies have demonstrated a crucial role for the jing zinc finger transcription factor in neuronal and glial differentiation and survival in the embryonic central nervous system midline of Drosophila. Here, we have studied the role of jing during embryonic brain development. Proper jing function is required for the formation of the primary brain axon scaffold. In homozygous jing (3) mutant embryos the preoral commissure is not pioneered and never forms. Other axon pathways are pioneered but subsequently do not form properly, including the postoral tritocerebral commissure, the circumesophageal connectives and the pathways that connect the brain with the ventral nerve cord. To understand the cellular basis of the axon phenotype the jing expression pattern in the brain was characterized using a jing-lacZ enhancer trap. jing-lacZ enhancer trap expression occurs in glia and neurons in the brain midline and lateral clusters as determined by co-localization of the lacZ gene product with Repo and Castor, respectively. In addition, the jing-lacZ enhancer trap and the basic helix-loop-helix-PAS gene, single-minded ( sim), are expressed in the only glial midline cluster present in stage-14 wild-type embryos. jing function is required for the differentiation of Repo-, Castor- and Sim-positive cells in the embryonic brain as each of these populations contain a reduced number of cells in homozygous jing (3) mutant embryos. We further find that jing is required for neuronal and glial survival as repo- and castor-expressing cells undergo cell death in homozygous jing (3) mutant embryos, as revealed by double labeling with Tunel. Expression of jing in sim-expressing cells in the brain disrupts the entire axon scaffold but most significantly results in loss of the preoral and postoral tritocerebral commissures. In addition, circumesophageal connectives are repelled after expression of two copies of UAS- jing in sim-expressing cells, suggesting the activation of axon repellent molecules. Over-expression of sim in the brain is also associated with loss of preoral and postoral tritocerebral commissures. Therefore, the proper dosage of jing and sim in the brain is critical for the formation of the primary axon scaffold. These results show that an important role for jing in the developing brain is the regulation of neuronal and glial differentiation and survival. PMID- 12111211 TI - Yan regulates Lozenge during Drosophila eye development. AB - The Drosophila eye offers an excellent opportunity to understand how general developmental processes are subtly altered to result in specific cell fates. Numerous transcription factors have been characterized in the developing eye; most of these are active in overlapping subsets of cells. Mechanisms used to regulate transcription factors act at many levels, and include competition for cognate binding sites, post translational modification, transcriptional regulation and cofactor availability. In undifferentiated cells of the larval eye imaginal disc, the transcriptional repressor Yan outcompetes the transcriptional activator Pointed for ETS binding sites on the prosperoenhancer. During differentiation, the Ras signaling cascade alters the Yan/Pointed dynamic through protein phosphorylation, effecting a developmental switch. In this way, Yan and Pointed are essential for prospero regulation. Hyperstable Yan (ACT) cannot be phosphorylated and blocks prospero expression. Lozenge is expressed in undifferentiated cells, and is required for prospero regulation. We sequenced the eye-specific enhancer of lozenge in three Drosophila species spanning 17 million years of evolution and found complete conservation of three ETS consensus binding sites. We show that lozengeexpression increases as cells differentiate, and that Yan (ACT) blocks this upregulation at the level of transcription. We find that expression of Lozenge via an alternate enhancer alters the temporal expression of Prospero, and is sufficient to rescue Prospero expression in the presence of Yan (ACT). These results suggest that Lozenge is involved in the Yan/Pointed dynamic in a Ras-dependent manner. We propose that upregulated Lozenge acts as a cofactor to alter Pointed affinity, by a mechanism that is recapitulated in mammalian development. PMID- 12111213 TI - Retinoic acid differently affects the formation of palps and surrounding neurons in the ascidian tadpole. AB - The anterior-most surface of the ascidian tadpole larvae is composed of specialized complex structures, including adhesive organs (palps) and the surrounding sensory neurons (RTENs) connected to neurons inside the palps. These are derived from a-line blastomeres by inductive effects from A-line blastomeres. The induction is reported to coordinate the expression of homeobox genes in the anterior epidermis, which can be affected by all- trans retinoic acid (RA). RA treatment also results in failure of the morphological formation of palps. Here we first isolated a gene intensely expressed in the cells of the anterior structure from the time of their lineage restriction, and then found that the RA treatment did not affect the specific gene expression in the presumptive palp cells but did affect that in the RTENs. These results suggest that the palp formation involves at least two different processes, a RA-insensitive cell-type specification process and a RA-sensitive morphogenetic process. RA treatment also affects the morphogenetic process of the palp formation and also disturbs the precise patterning of the surrounding epidermis, which may contribute to the regulation of RTEN development. PMID- 12111214 TI - An orphan PRD class homeobox gene expressed in mouse brain and limb development. AB - We report the cDNA sequence and expression of a mouse homeobox gene, Dmbx1, from the PRD class and comparison to its human orthologue. The gene defines a new homeobox gene family, Dmbx, phylogenetically distinct from the Ptx, Alx, Prx Otx, Gsc, Otp and Pax gene families. The Dmbx1 gene is expressed in the developing mouse diencephalon, midbrain and hindbrain, and has dynamic expression during forelimb and hindlimb development. Unusually for homeobox genes, there is no orthologue in the Drosophila or Caenorhabditis genomes; we argue this reflects secondary loss. PMID- 12111215 TI - Insights into the G1/S transition in plants. AB - The G1/S transition generally represents the principal point of commitment to cell division. Many of the components of the cell cycle core machinery regulating the G1/S transition in plants have been recently identified. Although plant regulators of the G1/S transition display structural and biochemical homologies with their animal counterparts, their functions in integrating environmental stimuli and the developmental program within cell cycle progression are often plant-specific. In this review, recent progress in understanding the role of plant G1/S transition regulators is presented. Emerging evidence concerning the mechanisms of G1/S control in response to factors triggering the cell cycle and the integration of these mechanisms with plant development is also discussed. PMID- 12111216 TI - Immunolocalization of beta-D-glucans, pectins, and arabinogalactan-proteins during intrusive growth and elongation of nonarticulated laticifers in Asclepias speciosa Torr. AB - Nonarticulated laticifers are latex-containing cells that elongate indefinitely and grow intrusively between the walls of meristematic cells. To identify biochemical mechanisms involved in the growth of nonarticulated laticifers, we have analyzed the distribution of various polysaccharides and proteoglycans in walls of meristematic cells in contact with laticifers, nonadjacent to laticifers, and in laticifer walls. In the shoot apex of Asclepias speciosa, the levels of callose and a (1-->4)-beta-galactan epitope are lower in meristematic walls in contact with laticifers than in nonadjacent walls. In contrast, we did not detect a decline in xyloglucan, homogalacturonan, and arabinogalactan-protein epitopes upon contact of meristematic cells with laticifers. Laticifer elongation is also associated with the development of a homogalacturonan-rich middle lamella between laticifers and their neighboring cells. Furthermore, laticifers lay down walls that differ from those of their surrounding cells. This is particularly evident for epitopes in rhamnogalacturonan I. A (1-->5)-alpha-arabinan epitope in this pectin is more abundant in laticifers than meristematic cells, while the opposite is observed for a (1-->4)-beta-galactan epitope. Also, different cell wall components exhibit distinct distribution patterns within laticifer walls. The (1-->5)-alpha-arabinan epitope is distributed throughout the laticifer walls while certain homogalacturonan and arabinogalactan-protein epitopes are preferentially located in particular regions of laticifer walls. Taken together, our results indicate that laticifer penetration causes changes in the walls of meristematic cells and that there are differences in wall composition within laticifer walls and between laticifers and their surrounding cells. PMID- 12111217 TI - The "drought-inducible" histone H1s of tobacco play no role in male sterility linked to alterations in H1 variants. AB - Tobacco ( Nicotiana tabacum L.) has two major H1 variants (H1A and H1B), which account for over 80% of chromatin linker histones, and four minor variants: H1C, H1D, H1E and H1F. We have shown previously [M. Prymakowska-Bosak et al. (1999) Plant Cell 11:2317-2329] that reversal of the natural proportion of major to minor H1 variants in transgenic tobacco plants results in a characteristic male sterility phenotype identical to that occurring in many plant species subjected to water deficit at the time of male meiosis. It has been proposed by others that the drought-induced arrest of male gametophyte development is linked to decreased sugar delivery to reproductive tissues. Within the family of angiosperm H1s there is a well-defined class of minor H1 variants named "drought inducible" because some of its members have been shown to be induced by water deficit. We have identified and cloned the tobacco H1C gene, which, based on sequence similarity, represents a "drought-inducible" minor H1 variant. Analysis of the un-translated mRNA and promoter regions of H1C suggests a regulation by sucrose concentration. Antisense silencing of H1C and its close homologue H1D in plants that do not express H1A and H1B does not affect the characteristic H1A(-)/ H1B(-) male sterility phenotype. Silencing of H1C and H1D also has no effect on growth and development of plants. Our findings demonstrate that H1C and H1D are dispensable for normal growth and development of tobacco, and that the compensatory up regulation of "drought-inducible" H1s observed in H1A(-)/ H1B(-) plants is not the direct cause of male sterility linked to alterations in H1 variants. PMID- 12111218 TI - Calcium oxalate crystal morphology mutants from Medicago truncatula. AB - Plants accumulate crystals of calcium oxalate in a variety of shapes and sizes. The mechanism(s) through which a plant defines the morphology of its crystals remains unknown. To gain insight into the mechanisms regulating crystal shapes, we conducted a mutant screen to identify the genetic determinants. This is the first reported mutant screen dedicated to the identification of crystal morphology mutants. A single leaf was harvested from individual Medicago truncatula L. plants that had been chemically mutagenized. Each leaf was visually inspected, using crossed-polarized light microscopy, for alterations in crystal shape and size. Seven different crystal morphology defective ( cmd) mutants were identified. Six cmd mutants were recessive and one dominant. Genetic analysis of the six recessive mutants suggested that each mutant was affected at a different locus. Each cmd mutant represents a new locus different than any previously identified. The plant phenotype of the cmd mutants appeared similar to that of the wild type in overall growth and development. This observation, coupled with the finding that several of the mutants had drastically altered the amount of calcium they partition into the oxalate crystal, questions current hypotheses regarding crystal function. Comparisons between the mutant crystals and those present in other legumes indicated the likelihood that simple point mutations contributed to the evolution of the variations in prismatic crystal shapes commonly observed in these plants today. The availability of cmd mutants provides the opportunity to investigate aspects of crystal shape and size that have been recalcitrant to previous approaches. PMID- 12111219 TI - The host guides morphogenesis and stomatal targeting in the grapevine pathogen Plasmopara viticola. AB - The oomycete grape downy mildew (Plasmopara viticola Berk. & Curt. Ex de Bary) is a serious pathogen of grapevine and spreads by extremely efficient cycles of asexual propagation. The high efficiency must involve efficient sensing of the host. We therefore analyzed the time course and morphology of the early development of this pathogen in a host system, by infection of leaf discs of grapevine (Vitis vinifera L. cv. Muller-Thurgau), and in a host-free system. Host factors were demonstrated to influence pathogen development in the following ways: (i) the release of zoospores from mature sporangia was accelerated, (ii) the morphogenesis of the germ tube was coordinated, and (iii) the zoospores were targeted to the stomata by factors that depended on stomata closure. The findings show that the early development of P. viticola is regulated, specifically and coordinately, by factors originating from the host plant. PMID- 12111220 TI - Compartmentation of aluminium in leaves of an Al-accumulator, Fagopyrum esculentum Moench. AB - Buckwheat (Fagopyrum esculentum Moench.) is an Al-accumulating plant, but the internal mechanism(s) of detoxification of Al is not fully understood. We investigated the subcellular localization of Al in the leaves of this plant (cv. Jianxi) by directly isolating protoplasts and vacuoles. Pure protoplasts and vacuoles from the leaves of buckwheat, grown hydroponically in Al solution, were obtained based on light-microscopic observation and the activities of marker enzymes of cytosol and vacuoles. More than 80% of total Al in the leaves was present in the protoplasts, and was identified as an Al-oxalate complex (1:3 ratio) by (27)Al-nuclear magnetic resonance. Oxalate and Al in the protoplasts was localized in the vacuoles. These results suggest that internal detoxification of Al in the buckwheat leaves is achieved by both complexation with oxalate and sequestration into vacuoles. PMID- 12111221 TI - Novel iron-storage particles may play a role in aluminum tolerance of Cyanidium caldarium. AB - Cyanidium caldarium (Tilden) Geitler, a unicellular red alga, has extraordinarily high aluminum (Al) tolerance. Algal cells cultured in the presence or absence of Al were subjected to transmission electron microscopy and energy dispersive X-ray analysis. Substantial changes to the thylakoid lumens were observed for the algal cells cultured in medium containing 200 mM Al, while other organelles were largely unaffected. Several spherical electron-dense bodies were found in the cytoplasm near the nucleus of both of the control and Al-treated cells. Although high levels of Fe and P were found in the bodies of control cells, immunocytochemical and morphological analysis data did not match the criteria established for Fe-accumulating substances like ferritin and phytate. In addition to these elements, Al was found in the bodies of the Al-treated cells. These results suggest that the electron-dense bodies function as an Fe-storage site under normal culture conditions, and that sequestration of Al in these bodies contributes to the high Al tolerance exhibited by C. caldarium. PMID- 12111222 TI - Aluminium triggers malate-independent potassium release via ion channels from the root apex in wheat. AB - The regulatory mechanisms for the aluminium (Al)-induced efflux of K(+) and malate from the root apex of Al-resistant wheat ( Triticum aestivum L. cv. Atlas) were characterized. Treatment with 20 mM tetraethylammonium (TEA) chloride, a K(+)-channel inhibitor, blocked the Al-induced K(+) efflux by 65%, but blocked the Al-induced malate efflux only slightly. Lanthanum (La(3+)) or ytterbium (Yb(3+)) strongly inhibited the K(+) efflux, but slightly increased malate efflux. These lanthanides applied together with Al did not affect the Al-induced malate efflux, but reduced the Al-induced K(+) efflux by 57% for La(3+) and by 35% for Yb(3+). By contrast, pretreatment with 50 microM niflumic acid, an anion channel inhibitor, strongly suppressed the Al-induced malate efflux, but did not affect the Al-induced K(+) efflux. The efflux of K(+) uncoupled with that of malate resulted in the alkalization of intracellular pH in the root apex, suggesting that the release of K(+) coupled with malate plays an important role in stabilizing intracellular pH. Copper (Cu(2+)) induced the release of K(+) via a TEA-insensitive pathway without the release of malate in both Al-resistant and Al-sensitive (cultivar Scout) wheat. Simultaneous application of Al and Cu(2+) to the root apices resulted in TEA-sensitive K(+) efflux in Atlas but not in Scout, suggesting that Al competes with Cu(2+) for K(+) efflux. Taken together, these results suggest that Al-induced K(+) efflux is mediated by both TEA- and lanthanide-sensitive K(+) channels, although this induction is not a prerequisite for the induction of the release of malate. PMID- 12111223 TI - Prevention of stomatal closure by immunomodulation of endogenous abscisic acid and its reversion by abscisic acid treatment: physiological behaviour and morphological features of tobacco stomata. AB - Transgenic tobacco ( Nicotiana tabacum L.) plants ubiquitously accumulating a single-chain variable-fragment (scFv) antibody against abscisic acid (ABA) to high concentrations in the endoplasmic reticulum (RA plants) show a wilty phenotype. High stomatal conductance and loss of CO(2) and light dependence of stomatal conductance are typical features of these plants. ABA was applied to these plants either via the petioles or by daily spraying over several weeks in order to normalise the phenotype. During the long-term experiments, scFv protein concentrations, total and (calculated) free ABA contents, and stomatal conductance and its dependence on CO(2) concentration and light intensity were monitored. The wilty phenotype of transgenic plants could not be normalised by short-term treatment with ABA via the petioles. Only a daily long-term treatment during plant development normalised the physiological behaviour completely. Scanning electron microscopy of stomata showed morphological changes in RA plants compared with wild-type plants that, for structural reasons, prevented regular stomatal movements. After long-term treatment with ABA this defect could be completely eliminated. Guard-cell-specific expression of the anti-ABA scFv did not cause any changes in physiological behaviour compared to the wild type. In addition, mesophyll-specific expression starting in leaves that were already fully differentiated resulted in normal phenotypes, too. We conclude that changes in distribution and availability of ABA in the cells of developing leaves of RA plants cause the development of structural features in stomata that prevent normal function. PMID- 12111224 TI - Expression of the NH(+)(4)-transporter gene LEAMT1;2 is induced in tomato roots upon association with N(2)-fixing bacteria. AB - Plants growing in close association with N(2)-fixing bacteria are able to overcome growth limitations in N-depleted soils. The molecular mechanism by which free-living, N(2)-fixing bacteria promote plant growth is still a matter of debate. By inoculating N-depleted tomato (Lycopersicon esculentum Mill.) plants with Azospirillum brasilense or Azoarcus sp. we could demonstrate the induction of the root NH(+)(4)-transporter gene, LEAMT1;2 (L. esculentum ammonium transporter 1;2), indicating that bacterial NH(+)(4) might be used as an N source under these conditions. Azospirillum brasilense (nif(-)) mutants, which lack the structural nifDK genes, failed to induce LEAMT1;2 expression. This suggests that root-associated N(2)-fixing bacteria do excrete NH(+)(4) to levels that can be sensed by tomato roots and is in agreement with the induction of expression of LEAMT1;2 with as low as > or = 1 microM external NH(+)(4). While peak expression was obtained with 2-5 microM NH(+)(4), a further increase in NH(+)(4) reduced LEAMT1;2-mRNA levels in a concentration-dependent manner. The inhibition of LEAMT1;2 expression by glutamine and the glutamine synthetase blocker L methionine sulfoximine (MSX) provided evidence for the control of LEAMT1;2 expression by cytoplasmic NH(+)(4) concentration or the plant N status. Since micromolar concentrations of NH(+)(4) strongly increased the LEAMT1;2-mRNA levels, the transported NH(+)(4) ion itself could represent a key signal in the associative interaction between higher plants and N(2)-fixing micro-organisms. PMID- 12111226 TI - Altered cell wall disassembly during ripening of Cnr tomato fruit: implications for cell adhesion and fruit softening. AB - The Cnr ( C olourless n on- r ipening) tomato ( Lycopersicon esculentum Mill.) mutant has an aberrant fruit-ripening phenotype in which fruit do not soften and have reduced cell adhesion between pericarp cells. Cell walls from Cnr fruit were analysed in order to assess the possible contribution of pectic polysaccharides to the non-softening and altered cell adhesion phenotype. Cell wall material (CWM) and solubilised fractions of mature green and red ripe fruit were analysed by chemical, enzymatic and immunochemical techniques. No major differences in CWM sugar composition were detected although differences were found in the solubility and composition of the pectic polysaccharides extracted from the CWM at both stages of development. In comparison with the wild type, the ripening-associated solubilisation of homogalacturonan-rich pectic polysaccharides was reduced in Cnr. The proportion of carbohydrate that was chelator-soluble was 50% less in Cnr cell walls at both the mature green and red ripe stages. Chelator-soluble material from ripe-stage Cnr was more susceptible to endo-polygalacturonase degradation than the corresponding material from wild-type fruit. In addition, cell walls from Cnr fruit contained larger amounts of galactosyl- and arabinosyl containing polysaccharides that were tightly bound in the cell wall and could only be extracted with 4 M KOH, or remained in the insoluble residue. The complexity of the cell wall alterations that occur during fruit ripening and the significance of different extractable polymer pools from cell walls are discussed in relation to the Cnr phenotype. PMID- 12111225 TI - Generation, characterization, and heterologous expression of wild-type and up regulated forms of Arabidopsis thaliana leaf ADP-glucose pyrophosphorylase. AB - ADP-glucose pyrophosphorylase (AGPase), a key enzyme in starch biosynthesis of higher plants, consists of a pair of regulatory large (LS) and catalytically small (SS) subunits. In plants, these subunits are coded by multiple genes resulting in the formation of tissue-specific enzyme forms, which are differentially regulated during plant growth and development. Some AGPase isoforms differ in catalytic and regulatory properties as well as intracellular location. In an effort to gain a better understanding of the role of the leaf AGPase in carbon partitioning and its effect on plant productivity, the Arabidopsis leaf AGPase containing the mature forms of the SS and LS was expressed in a heterologous expression system and characterized enzymatically. The Arabidopsis recombinant AGPase had kinetic values for 3-phosphoglyceric acid, glucose-1-phosphate and Mg(2+) similar to those of the native enzyme. As the N terminus of the LS has been suggested to be involved in enzyme function, the length of the N-terminal region was extended or shortened. Of the five modified LSs analyzed, only the T5 form lacking six residues of the mature N-terminus was able to form detectable levels of enzyme activity, indicating that the N-terminal region is critical for enzyme function. Two up-regulatory LS mutations that allosterically activate the potato enzyme, a stem isoform, were introduced into the corresponding Arabidopsis LS sequences and co-expressed with wild-type SS. Both modified enzymes showed up-regulatory properties, indicating that these specific residue changes were also operational in the leaf isoform. PMID- 12111227 TI - The non-photosynthetic phosphoenolpyruvate carboxylases of the C4 dicot Flaveria trinervia -- implications for the evolution of C4 photosynthesis. AB - C4 phospho enolpyruvate carboxylases (PEPCase; EC 4.1.1.3) have evolved from ancestral non-photosynthetic (C3) isoforms during the evolution of angiosperms and thereby gained distinct kinetic and regulatory properties. In order to obtain insight into this evolutionary process we have studied the C3 isoforms, ppcB and ppcC, of the C4 dicot Flaveria trinervia (Spreng.) C. Mohr and compared them with the C4 enzyme of this species, ppcA, and its orthologue in the C3 species F. pringlei Gandoger. Phylogenetic analyses indicate that the ppcB PEPCase is the closest relative of the ppcA enzyme. In addition, the presence of ppcB also in the closely related C3 species F. pringlei suggests that this gene was present already in the ancestral C3 species and consequently that ppcA has evolved by gene duplication of ppcB. Investigation of the enzymatic properties of the ppcB and ppcC enzymes showed low and similar K(0.5)-PEP values and limited activation by glucose-6-phosphate, typical of non-photosynthetic PEPCases, at pH 8.0. However, at the more physiological pH of 7.6, the ppcC enzyme displayed a substantially higher K(0.5)-PEP than the ppcB counterpart, indicating their involvement in different metabolic pathways. This indication was strengthened by malate inhibition studies in which the ppcC enzyme showed 10 times higher tolerance to the inhibitor. The ppcA enzyme was, however, by far the most tolerant enzyme towards malate. Interestingly, the increased malate tolerance was correlated with a decrease in enzyme efficiency displayed by the turnover constant k(cat). We therefore suggest that the increased malate tolerance, which is imperative for an efficient C4 cycle, is connected with a decreased enzyme efficiency that in turn is compensated by increased enzyme expression. PMID- 12111228 TI - Seasonal responses of photosynthetic electron transport in Scots pine (Pinus sylvestris L.) studied by thermoluminescence. AB - The potential of photosynthesis to recover from winter stress was studied by following the thermoluminescence (TL) and chlorophyll fluorescence changes of winter pine needles during the exposure to room temperature (20 degrees C) and an irradiance of 100 micromol m(-2) s(-1). TL measurements of photosystem II (PSII) revealed that the S(2)Q(B)(-) charge recombinations (the B-band) were shifted to lower temperatures in winter pine needles, while the S(2)Q(A)(-) recombinations (the Q-band) remained close to 0 degrees C. This was accompanied by a drastically reduced (65%) PSII photochemical efficiency measured as F(v)/ F(m,) and a 20-fold faster rate of the fluorescence transient from F(o) to F(m) as compared to summer pine. A strong positive correlation between the increase in the photochemical efficiency of PSII and the increase in the relative contribution of the B-band was found during the time course of the recovery process. The seasonal dynamics of TL in Scots pine needles studied under field conditions revealed that between November and April, the contribution of the Q- and B-bands to the overall TL emission was very low (less than 5%). During spring, the relative contribution of the Q- and B-bands, corresponding to charge recombination events between the acceptor and donor sides of PSII, rapidly increased, reaching maximal values in late July. A sharp decline of the B-band was observed in late summer, followed by a gradual decrease, reaching minimal values in November. Possible mechanisms of the seasonally induced changes in the redox properties of S(2)/S(3)Q(B)(-) recombinations are discussed. It is proposed that the lowered redox potential of Q(B) in winter needles increases the population of Q(A)(-), thus enhancing the probability for non-radiative P680(+)Q(A)(-) recombination. This is suggested to enhance the radiationless dissipation of excess light within the PSII reaction center during cold acclimation and during cold winter periods. PMID- 12111229 TI - Xylem embolism and drought-induced stomatal closure in maize. AB - Water relations during drought and xylem vulnerability to embolism were studied on four maize ( Zea mays L.) genotypes having contrasting grain yields under drought conditions. Drought provoked a drop in xylem pressure, leaf water potential and whole-plant transpiration. Transpiration was reduced to a minimum value when xylem pressures reached ca. -1.6 MPa. This value corresponded to the threshold xylem pressure below which xylem embolism developed to a substantial degree in leaf midribs. Therefore, xylem embolism always remained low in leaf veins, even when plants exhibited clear water-stress symptoms. This suggests that stomatal closure during drought contains xylem embolism to a minimum value. Cavitation resistance was not related to grain yield under drought conditions for the four genotypes evaluated. However, it can be speculated that an increase in cavitation resistance by cultural practices or genetic selection may increase drought survival in maize. PMID- 12111230 TI - Degradation of a peptide in pitcher fluid of the carnivorous plant Nepenthes alata Blanco. AB - Carnivorous plants acquire substantial amounts of nitrogen from insects. The tropical carnivorous plant Nepenthes produces trapping organs called pitchers at the tips of tendrils elongated from leaf ends. Acidic fluid is secreted at the bottoms of the pitchers. The pitcher fluid includes several hydrolytic enzymes, and some, such as aspartic proteinase, are thought to be involved in nitrogen acquisition from insect proteins. To understand the nitrogen-acquisition process, it is essential to identify the protein-degradation products in the pitcher fluid. To gain insight into protein degradation in pitcher fluid, we used the oxidized B-chain of bovine insulin as a model substrate, and its degradation by the pitcher fluid of N. alata was investigated using liquid chromatography-mass spectrometry (LC-MS). LC-MS analysis of the degradation products revealed that the oxidized B-chain of bovine insulin was initially cleaved at aromatic amino acids such as phenylalanine and tyrosine. These cleavage sites are similar to those of aspartic proteinases from other plants and animals. The presence of a series of peptide fragments as degradation products suggests that exopeptidase(s) is also present in the pitcher fluid. Amino acid analysis and peptide fragment analysis of the degradation products demonstrated that three amino acids plus small peptides were released from the oxidized B-chain of bovine insulin, suggesting that insect proteins are readily degraded to small peptides and amino acids in the pitcher fluid of N. alata. PMID- 12111231 TI - Anthocyanins protect light-sensitive thiarubrine phototoxins. AB - Thiarubrines are phototoxic plant pigments that decompose to thiophenes when exposed to sunlight. We investigated the mechanism of thiarubrine photoprotection in Ambrosia chamissonis (Less.) Greene (Asteraceae), which contains high amounts of these chemicals in its stems and leaf petioles. Thiarubrines are compartmentalized in laticifers that are surrounded by anthocyanin-containing cells. When this light-screening sheath was removed and laticifers exposed to light, rapid bleaching of the thiarubrine contents occurred. The leaves and stems of A. chamissonis seedlings were found to contain 10.5+/-6.8 microg/g total anthocyanins, predominantly cyanidin 3-O-(6'-O-malonylglucoside) and cyanidin 3-O glucoside, while none was detected in roots. To correlate anthocyanin distribution with thiarubrine photoprotection, changes in thiarubrine A and thiophene A levels were measured in seedlings exposed to light. In roots, thiarubrine A levels decreased by 94% after 30 min of irradiation, and thiarubrines were completely absent after 4 h. A concomitant 3-fold increase in thiophene A levels in roots occurred during light exposure. In leaves and stems, thiarubrine A levels did not change appreciably during light exposure, with a nominal increase from 102.8+/-33.1 microg/g FW to 108.4+/-20.7 microg/g FW after 4 h. To confirm their photoprotective function, solutions of cyanidin 3-O glucoside were used to filter visible light incident on a solution of thiarubrine A. Anthocyanin solutions of greater than 0.1 mM completely prevented thiarubrine photoconversion. This is the first report that anthocyanins function to photoprotect light-sensitive defensive chemicals in plants. PMID- 12111232 TI - Characterization of a fertilization-induced and developmentally regulated plasma membrane aquaporin expressed in reproductive tissues, in the wild potato Solanum chacoense Bitt. AB - Fertilization triggers a unique and complex developmental program leading to embryogenesis and seed set. Growth and differentiation are accompanied by rapid metabolic changes as well as massive cellular reorganization inside the fertilized ovule. Using differential display to isolate genes involved in pollen pistil interactions and early fertilization events, we isolated from Solanum chacoense Bitt. a fertilization-induced plasma-membrane intrinsic protein of the PIP2 family that is predominantly expressed in pistil and anthers tissues. Major intrinsic proteins (MIPs) consist of a large family of highly conserved membrane spanning proteins that are mainly represented in plants by the aquaporins. Aquaporins, mainly of the PIP and TIP type, have been shown in many species to serve as water channels. In S. chacoense the ScPIP2a mRNA is developmentally regulated during anthesis, with mRNA levels gradually decreasing as the pistil reaches maturity. In flowers, strongest expression was observed in elongating styles, in stamens, and transiently in ovules following fertilization. In styles, maximal expression levels correlated with phases of rapid style elongation and with the formation of epidermal papillae. ScPIP2a mRNA was also strongly expressed in developing fruit, consistent with a role in cell expansion during maturation and development. PMID- 12111233 TI - Oligogalacturonides inhibit the induction of late but not of early auxin responsive genes in tobacco. AB - Oligogalacturonides (OGs) released from the plant cell wall regulate several defense responses, as well as various aspects of plant growth and development. In these latter effects, OGs exhibit auxin-antagonist activity. To shed light on the mechanism by which OGs antagonise auxin, we analysed the ability of these oligosaccharides to inhibit the activity of four auxin-up-regulated promoters [pGm-GH3 of soybean (Glycine max L. Merr.), pNt114 of tobacco (Nicotiana tabacum L.), and prolB and prolD of Agrobacterium rhizogenes] driving the expression of the beta-glucuronidase reporter gene (GUS) in transgenic tobacco seedlings. Our results indicate that OGs at submicromolar concentrations inhibit the activation by auxin of pNt114, prolB and prolD, but not that of pGm-GH3. Comparative analysis of the kinetics of activation of the four promoters in response to the hormone shows that, while pGm-GH3 is rapidly activated, the other three promoters exhibit a delayed activation, with a lag of at least 4 h before the appearance of GUS activity. The lack of effect of the OGs on early auxin-responsive genes was confirmed by RNA gel blot analysis of the tobacco genes Nt-GH3 and Nt-iaa2.3/2.5. Our results suggest that the auxin-antagonist action of OGs affects the expression of late but not of early auxin-responsive genes. PMID- 12111234 TI - Solar ultraviolet radiation affects the activity of ribulose-1,5-bisphosphate carboxylase-oxygenase and the composition of photosynthetic and xanthophyll cycle pigments in the intertidal green alga Ulva lactuca L. AB - The effect of solar UV radiation on the physiology of the intertidal green macroalga Ulva lactuca L. was investigated. A natural Ulva community at the shore of Helgoland was covered with screening foils, excluding UV-B or UV-B + UV-A from the solar spectrum. In the sampled material, changes in the activity and concentration of ribulose-1,5-bisphosphate carboxylase-oxygenase (Rubisco), and the concentration of photosynthetic and xanthophyll cycle pigments were determined. Exclusion of UV radiation from the natural solar spectrum resulted in an elevated overall activity of Rubisco, related to an increase in its cellular concentration. Among the photosynthetic pigments, lutein concentration was substantially elevated under UV exclusion. In addition, marked UV effects on the xanthophyll cycle were found: exclusion of solar UV radiation (and particularly UV-B) resulted in an increased ratio of zeaxanthin concentration to the total xanthophyll content, indicating adverse effects of UV-B on the efficiency of photoprotection under high irradiances of photosynthetically active radiation. The results confirm a marked impact of present UV-B levels on macroalgal physiology under field conditions. PMID- 12111235 TI - Molecular characterization of a gene encoding a cystatin expressed in the stems of American chestnut (Castanea dentata). AB - A cDNA clone with similarity to genes encoding cystatin was recently isolated from a cDNA library created using mRNA extracted from stem tissues of Castanea dentata (Marsh.) Borkh. (CASde:Pic1). All of the requisite motifs for inhibitory activity were found upon examination of the deduced amino acid sequence. Reverse transcription-polymerase chain reaction was used to detect the cystatin transcript in healthy stem, leaf and seed tissues, as well as in diseased tissues. Gene fragments encoding this putative cystatin were cloned from American and Chinese (Castanea mollissima Blume) chestnuts and a comparison of these sequences revealed significant differences within the intron, including deletions and alterations in restriction-enzyme sites. The long-term goal of this study is to determine whether the cystatin allele in Chinese chestnut correlates to a resistance gene and, if so, if this allele could be used to enhance resistance in American chestnut. PMID- 12111236 TI - Export of acyl chains from plastids isolated from embryos of Brassica napus (L.). AB - We report the first measurements of the kinetics of transmembrane transport of acyl chains in plants. This was achieved by separating the period of in vitro synthesis of fatty acids from their export and by making use of acyl-CoA-binding protein (ACBP), which specifically binds long-chain acyl-CoAs. In the absence of added CoA but in the presence of ACBP, newly synthesised acyl chains accumulated as free fatty acids (FFAs) in plastids isolated from embryos of oilseed rape (Brassica napus L.). When CoA was added to plastids that had accumulated FFAs, the acyl chains were converted to acyl-CoAs that, in the presence of ACBP, were exported to the incubation medium. The rate of export was dependent on the CoA concentration and, at a saturating CoA concentration, was similar to the rate at which the fatty acids had been synthesised prior to CoA addition. PMID- 12111237 TI - A WEE1 homologue from Arabidopsis thaliana. AB - Little is known about the genes that regulate cyclinB-Cdc2 complexes at the G2/M transition of the plant cell cycle although in yeast and animals cdc25 and wee1 are central regulators of cdc2. Here we describe the isolation, by reverse transcription polymerase chain reaction (RT-PCR), of a WEE1 cDNA (AtWEE1) in Arabidopsis thaliana (L.) Heynh. Semi-quantitative RT-PCR showed that AtWEE1 expression was confined to actively dividing regions of the plant. The overexpression of AtWEE1 in fission yeast (Schizosaccharomyces pombe) caused cells to arrest, and to grow but not divide, resulting in very elongated cells. Our data provide evidence for a functional WEE1 in A. thaliana. PMID- 12111238 TI - What can we learn about cell signalling by combining optical imaging and patch clamp techniques? AB - Optical imaging is a powerful technique with which to investigate the activity, distribution and movement of biomolecules. The increased resolution of images obtained with confocal microscopy now allows us to visualize the signalling events in individual intracellular organelles. Local photobleaching and uncaging of caged compounds enable investigators to control the activity of many biologically important molecules in small localized regions of both cytosol and internal spaces of cellular organelles. Uncaging and photobleaching conveniently complement laser scanning confocal microscopy. The whole-cell recording configuration of the patch-clamp technique has been widely used not only to measure ionic currents, but also to control the concentration of important molecules in the cytosol. The cell-attached configuration of patch clamp was utilized for local stimulation of the cell and local delivery of the second messengers. This paper describes the advantages of combining patch-clamp and optical imaging methods as well as some of the recent achievements using this approach. PMID- 12111239 TI - Novel phosphate-regulating genes in the pathogenesis of renal phosphate wasting disorders. AB - Over the past decade, three classes of Na/Pi cotransporters have been identified in mammalian kidney. The type IIa Na/Pi cotransporter, Npt2, is the most abundant and is expressed in the brush-border membrane of renal proximal tubular cells where the bulk of filtered inorganic phosphate (Pi) is reabsorbed. Disruption of the Npt2 gene in mice underscored the importance of Npt2 in the overall maintenance of Pi homeostasis and demonstrated that Npt2 is the target for regulation of proximal tubular Pi reabsorption by parathyroid hormone and dietary Pi. The regulation is post-transcriptional and largely occurs by brush-border membrane retrieval and insertion of Npt2 protein. Of great interest is the recent identification of novel Pi regulating genes, PHEX and FGF23, that play a role in the pathophysiology of inherited (X-linked hypophosphatemia and autosomal dominant hypophosphatemic rickets) and acquired (oncogenic hypophosphatemic rickets) disorders characterized by renal Pi wasting and associated skeletal abnormalities. Studies are currently underway to elucidate the molecular basis for impaired renal Pi reabsorption in these disorders and to determine the precise physiological role of PHEX and FGF-23 in the regulation of Pi homeostasis. PMID- 12111240 TI - Regulation of cardiac CFTR Cl(-) channel activity by a Mg(2+)-dependent protein phosphatase. AB - Dephosphorylation of the CFTR Cl(-) channel is known to be induced by both okadaic-acid- (OA-) sensitive and -insensitive protein phosphatases (PPs). In the present study, the effects of cytosolic free Mg(2+) on the cardiac CFTR Cl(-) current were examined in relation to the latter PP activity in guinea pig ventricular myocytes. Even when maintaining intracellular Mg-ATP at millimolar concentrations under whole-cell patch-clamp mode, cAMP-activated Cl(-) conductance was reversibly suppressed by cytosolic free Mg(2+), with an IC(50) of around 2.5 mmol/l. In contrast, changes in the cytosolic concentration of free Mg(2+) ([Mg(2+)](i)) had no effect on genistein-activated CFTR Cl(-) currents. The Mg(2+) effect on cAMP-activated CFTR Cl(-) conductance was completely reversed by application of anthracene-9-carboxylic acid (9-AC), which was previously shown to inhibit an OA-insensitive PP in cardiac myocytes. A 9-AC sensitive fraction of endogenous PP activity in the extract of guinea pig ventricle was found to be activated by free Mg(2+) at millimolar concentrations but to be inactive at micromolar concentrations. The intracellular application of OA failed to activate basal Cl(-) conductance at millimolar [Mg(2+)](i). In the presence of OA, however, basal Cl(-) conductance became activated either by reducing [Mg(2+)](i) to micromolar concentrations or by applying 9-AC. Thus, we conclude that a Mg(2+)-dependent PP sensitive to 9-AC plays a key role in the cAMP-mediated regulation of cardiac CFTR Cl(-) channel at physiological [Mg(2+)](i)under both basal and cAMP-activated conditions. Also, it appears that the genistein-activated conformation of the cardiac CFTR channel is not sensitive to the Mg(2+)-dependent PP. PMID- 12111241 TI - Cyclic AMP stimulates renin gene transcription in juxtaglomerular cells. AB - Although the cyclic AMP signalling cascade is considered to be the main activator of renin gene expression in renal juxtaglomerular (JG) cells, the molecular pathways along which cAMP exerts this effect remain a matter of controversy. Here in this study we used the mouse JG cell line As4.1, which shares a number of functional similarities with native JG cells. We found that forskolin, an activator of adenylate cyclase, in the presents of IBMX time-dependently increased renin mRNA levels and prorenin secretion up to threefold. The stimulation of renin gene expression by forskolin/IBMX was markedly attenuated by an inhibitor of protein kinase A (H-89, 10 microM). Forskolin/IBMX had no effect on the decline of renin mRNA after general inhibition of transcription by actinomycin D (2 microM). Conversely, forskolin/IBMX increased the activity of a 2.8-kb fragment of the renin promoter threefold. The promoter region responsible for the stimulatory effect of forskolin/IBMX was narrowed down to three 4 bp of the mouse Ren1(C) gene, which are known as putative CRE-sites. The CRE-binding protein was found to be phosphorylated under forskolin/IBMX stimulation. It appears likely therefore that cAMP stimulates renin gene expression in JG cells by activating protein kinase A and subsequent phosphorylation of the CRE-binding protein. PMID- 12111242 TI - Expression and functional implications of troponin T isoforms in soleus muscle fibers of rat after unloading. AB - The expression pattern of troponin T (TnT) isoforms was studied in rat soleus muscle fibers in control and after hindlimb unloading (HU) conditions. To determine the functional consequence of TnT expression, the fibers were also examined for their calcium activation characteristics. With regard to TnT expression, four populations of fibers were distinguished in control muscle. Slow fibers expressing only slow isoforms of TnT (TnT1s, 2s, 3s ) were predominant (54%). Hybrid slow fibers (16%) differed from slow fibers by the additional expression of two TnTf isoforms. Hybrid fast fibers (22%) expressed slow and fast isoforms of TnT while fast fibers (8%) expressed only fast TnT isoforms. The expression of the other regulatory protein isoforms was checked for each population. The contractile experiments revealed steeper slopes of the tension/pCa relationship from hybrid slow fibers expressing fast TnT in a completely slow molecular environment. The expression of TnTs in hybrid fast fibers did not modulate the intrinsic co-operativity. After HU, the fast population was increased and reached 55%. The slow population decreased to 41% and a very small amount of hybrid slow fibers remained (4%). These data demonstrated the implication of TnT isoforms in the calcium activation properties and, more particularly, in the modulation of co-operativity within the myofibrillar lattice. Regulation of TnT expression appeared as a very fast and complete process compared to moderate changes of TnC and TnI. PMID- 12111243 TI - Regulation of the mouse Nas1 promoter by vitamin D and thyroid hormone. AB - The renal sodium-sulfate cotransporter, NaS(i)-1, a protein implicated to control serum sulfate levels, has been shown to be regulated in vivo by 1,25 dihydroxyvitamin D(3) (1,25-(OH)(2)D(3)) and tri-iodothyronine (T(3)). Recently, we cloned the mouse NaS(i)-1 gene ( Nas1) and in the present study identified a 1,25-(OH)(2)D(3)- and T(3)-responsive element located within the Nas1 promoter. Mutational analysis of the Nas1 promoter resulted in identification of a direct repeat 6-type vitamin-D-responsive element (DR6 VDRE) at -525 to -508 and an imperfect inverted repeat 0-type T(3)-responsive element (IR0 T(3)RE) at -436 to 425 which conferred 1,25-(OH)(2)D(3) and T(3) responsiveness, respectively. In summary, we have identified responsive elements that mediate the enhanced transcription of Nas1 by 1,25-(OH)(2)D(3) and T(3), and these mechanisms may provide important clues to the physiological control of sulfate homeostasis. PMID- 12111244 TI - Assessment of sarcoplasmic reticulum Ca(2+) flux pathways in cardiomyocytes from rabbits with infarct-induced left-ventricular dysfunction. AB - The aim of the study was to correlate intracellular Ca(2+) transients with Ca(2+) uptake and efflux characteristics of the sarcoplasmic reticulum (SR) in ventricular myocytes isolated from rabbits with left-ventricular dysfunction (LVD). Chronic (8 weeks) ligation of a coronary artery caused marked LVD in rabbits. Measurements of intracellular [Ca(2+)] were made using Fura-2 on intact, single, left-ventricular myocytes. SR Ca(2+) flux rates associated with sarco endoplasmic reticulum Ca(2+) ATPase type 2 (SERCA2)-mediated uptake, ryanodine receptor type 2 (RyR2)-mediated Ca(2+) efflux and background SR Ca(2+) leak were measured in suspensions of permeabilised myocytes. Measurements on single, permeabilised myocytes were used to assess the steady-state Ca(2+) content of the SR and the characteristics of spontaneous SR Ca(2+) release. Peak systolic [Ca(2+)] was significantly lower; time-to-peak and Ca(2+) transient duration were significantly longer in LVD myocytes. SERCA2-mediated Ca(2+) uptake was reduced to approximately 50% in myocytes from the LVD group. Ruthenium red (RuR) sensitive Ca(2+) efflux (mediated by the RyR2) was also reduced in the LVD group by approximately 50%, as was the remaining (RuR-insensitive) background Ca(2+) leak. Measurements from single, permeabilised myocytes showed a lower steady state SR Ca(2+) content. The frequency and amplitude of spontaneous SR Ca(2+) release from LVD hearts was also reduced. Partial inhibition of SERCA2 by thapsigargin depressed both the amplitude and the frequency of spontaneous release. Partial inhibition of RyR2-mediated-Ca(2+) efflux with tetracaine enhanced spontaneous Ca(2+) release amplitude and decreased frequency. Increased background Ca(2+) leak with ionomycin decreased the frequency of spontaneous release. It is concluded that partial inhibition of SERCA2 mimics some aspects of altered SR function in LVD, but reduced RyR2 function cannot explain the other functional alterations observed. Reduced background Ca(2+) leak from the SR may compensate partly for the reduced Ca(2+) uptake capacity of the SR in the LVD group. PMID- 12111245 TI - Hypothalamic tryptophan hydroxylase and the hypothalamic-pituitary-adrenocortical response to water deprivation and hydration in vasopressin-deficient and normal rats. AB - The effects of water deprivation and hydration on plasma corticosterone concentration and the activity of tryptophan hydroxylase (TPH), the rate-limiting enzyme in serotonin (5-HT) biosynthesis, in the hypothalamus of vasopressin- (AVP ) deficient homozygous Brattleboro and normal Wistar rats were studied. In the Wistar rats, water deprivation caused an increase in the TPH activity in the anterior and middle (infundibular) parts of the hypothalamus, while hydration did not affect the activity of the enzyme in the anterior hypothalamus but produced an increase in its middle part. In contrast, in the Brattleboro rats, water deprivation had no effect on TPH activity in the anterior and middle parts of the hypothalamus but hydration produced a decrease in TPH activity in the anterior hypothalamus. After 48 h of water deprivation, the plasma corticosterone concentration significantly increased in water-deprived and decreased in hydrated Wistar rats. Under water deprivation, the rise in corticosterone concentration in the homozygous Brattleboro rats was significantly greater than that in the Wistar rats. The data provide evidence that the CRH-like activity of AVP is not necessary for activation of the hypothalamic-pituitary-adrenal system induced by water deprivation. The observations show that AVP is involved in the activation of TPH induced by water deprivation. This suggests that AVP modulates the metabolism of 5-HT and the response of the 5-HT-ergic system to water deprivation. PMID- 12111246 TI - Disturbances of colonic ion secretion in inflammation: role of the enteric nervous system and cAMP. AB - We used the trinitrobenzenesulphonic acid (TNBS) rat model of experimental colitis to study the alterations in electrogenic ion transport in the inflamed distal colon. The distal colon exhibited decreased basal transport and reduced short-circuit current responses to carbachol and isobutylmethylxanthine (IBMX). The concentration/response curve for IBMX was also shifted to the right. Ion substitution experiments indicated that electrogenic transport was attributable chiefly to Cl(-) secretion. The mucosal layer of the inflamed distal colon (devoid of the submucosa) exhibited normal maximal responses to carbachol and IBMX, although the concentration/response curve for the latter was again shifted to the right. Tetrodotoxin markedly increased the response of the normal distal colon to both secretagogues and nullified the inhibition of the response to carbachol, but not that to IBMX, in the inflamed colon. The response of the mucosal preparation to 8-bromoadenosine 3',5'-cyclic monophosphate was similar in the normal and inflamed intestine, while the G protein activator NaF had a greater effect in the latter. The expression of the cystic fibrosis transmembrane conductance regulator (CFTR), as assessed by Northern blotting, was unchanged. cAMP levels in isolated colonocytes were markedly reduced by inflammation. We conclude that colonic inflammation produces disturbances of the enteric nervous system resulting in defective mucosal cAMP production and inhibition of ionic secretion. PMID- 12111247 TI - Inwardly rectifying potassium currents in rat basophilic leukaemia (RBL-1) cells: regulation by spermine and implications for store-operated calcium influx. AB - In non-excitable cells, the major Ca(2+) influx pathway is the store-operated one. Store-operated Ca(2+) entry is intimately related to the prevalent membrane potential, in that hyperpolarisation enhances Ca(2+) influx and depolarisation reduces it. Inwardly rectifying potassium channels are important determinants of the membrane potential and hence will regulate, indirectly, the rate and extent of Ca(2+) entry through store-operated channels. Here we investigated inwardly rectifying potassium currents ( I(RK)) in rat basophilic leukaemia (RBL-1) cells, a model system for studying store-operated Ca(2+) influx. I(RK) was voltage dependent in that the current decays during strong hyperpolarisations. Recovery from this decay was both time and voltage dependent. Close to the resting potential of RBL-1 cells, however, I(RK) was stable. Neither store depletion per se nor the subsequent rise in intracellular [Ca(2+)] appeared to alter I(RK) activity. Receptor stimulation reduced the current only weakly. Unexpectedly, intracellular spermine inhibited I(RK) quite strongly and via a mechanism that seemed distinct from that responsible for current rectification. The relevance of these findings to store-operated Ca(2+) influx is discussed. PMID- 12111248 TI - Exhaled nitric oxide level during and after heavy exercise in athletes with exercise-induced hypoxaemia. AB - Endogenous nitric oxide (NO) is an important mediator of vasodilatation, bronchodilatation and lung inflammation. We hypothesised that the exhaled NO level may be modified in some endurance-trained athletes during and after intense exercise. Nine athletes with exercise-induced hypoxaemia (EIH), 12 athletes without EIH and 10 untrained subjects exercised for 15 min at 90% maximal oxygen consumption (VO(2)max). Exhaled NO was measured during exercise, and after 1 h and 22 h of recovery. Exhaled NO concentration ( C(NO)) decreased significantly during exercise in all subjects and returned to basal values after 1 h of recovery with no further modification. Exhaled NO output (V(NO)) rose significantly during exercise, rapidly dropped down following exercise and was similar to resting values after 1 h and 22 h of recovery. The results also showed that C(NO) and V(NO) were significantly lower in the athletes with EIH in comparison with the untrained subjects (V(NO) was 5.32 +/- 0.77 nmol/min versus 3.61 +/- 0.72 nmol/min at rest, 18.52 +/- 1.50 nmol/min versus 15.00 +/- 2.06 nmol/min during heavy exercise, and 5.52 +/- 1.04 nmol/min versus 3.79 +/- 0.76 nmol/min after 22 h recovery, in untrained subjects and EIH athletes, respectively). These findings do not confirm the hypothesis of pulmonary inflammation associated with EIH. However, potential NO epithelial down regulation may occur and contribute to the development of gas exchange abnormality in some endurance-trained athletes. PMID- 12111249 TI - Caudal expiratory neurones in the rat. AB - The main source of expiratory drive to respiratory muscles in the rat is thought to be the caudal expiratory neurones. However, their projections to the spinal cord and the coordination of the population activity with the respiratory cycle are largely unknown. We examined their bulbospinal projections using antidromic activation, and the coordination of their activity using cross-correlation. Of 76 expiratory neurones examined, 40% projected to the C2 segment of the spinal cord, all unilaterally and all but 4 to the contralateral side. Thirteen contralateral axons were located in the ventromedial funiculus, when activation thresholds were less than 25 microA. For 29 neurones, the mean (+/- SEM) conduction velocity, calculated from the single-point activation and a measure of the direct distance between recording and stimulating electrodes, was 7.4 +/- 0.4 m/s. We calculated 88 cross-correlograms from ipsilateral pairs of expiratory neurones recorded on the same side of the medulla, and 176 contralateral pairs recorded from opposite sides of the medulla. All of the latter were featureless, but 23% of cross correlograms for ipsilateral pairs displayed broad peaks at time zero, which we interpreted as due to activation of both neurones from a common source. We conclude that in the adult rat, approximately half of the caudal expiratory neurones project unilaterally and contralaterally to the spinal cord and, although common activation serves to coordinate some of the ipsilateral population, we suggest that neither common activation nor excitatory cross connections exist as sufficient means for coordinating left and right populations to the same respiratory rhythm. PMID- 12111250 TI - Barttin increases surface expression and changes current properties of ClC-K channels. AB - The term Bartter syndrome encompasses a heterogeneous group of autosomal recessive salt-losing nephropathies that are caused by disturbed transepithelial sodium chloride reabsorption in the distal nephron. Mutations have been identified in the NKCC2 (Na(+)-K(+)-2Cl(-)) cotransporter and ROMK potassium channel, which cooperate in the process of apical sodium chloride uptake, and ClC Kb chloride channels, which mediate basolateral chloride release. Recently, mutations in barttin, a protein not related to any known ion transporter or channel, were described in BSND, a variant of Bartter syndrome associated with sensorineural deafness. Here we show that barttin functions as an activator of ClC-K chloride channels. Expression of barttin together with ClC-K in Xenopus oocytes increased ClC-K current amplitude, changed ClC-K biophysical properties, and enhanced ClC-K abundance in the cell membrane. Co-immunoprecipitation revealed a direct interaction of barttin with ClC-K. We performed in situ hybridization on rat kidney slices and RT-PCR analysis on microdissected nephron segments to prove co-expression of barttin, ClC-K1 and ClC-K2 along the distal nephron. Functional analysis of BSND-associated point mutations revealed impaired ClC-K activation by barttin. The results demonstrate regulation of a CLC chloride channel by an accessory protein and indicate that ClC-K activation by barttin is required for adequate tubular salt reabsorption. PMID- 12111251 TI - Alterations in single muscle fiber calcium sensitivity with resistance training in older women. AB - The purpose of this investigation was to determine the effects of a 12-week progressive resistance-training program (PRT) on single muscle fiber calcium sensitivity in six older women (73 +/- 2 years). Muscle biopsy samples of the vastus lateralis were obtained pre- and post-PRT. Chemically skinned single muscle fibers ( n=274) were dissected and studied. The experimental sequence for each fiber was the determination of peak maximal isometric tension ( P(o)) at pCa 4.5 (pCa=-log[Ca(2+)]), and then subsequent submaximal activations of the fiber at nine Ca(2+) concentrations (pCa 6.8 to 4.7). Myosin heavy chain (MHC) I fiber (slow-twitch) diameter increased 16% ( P<0.05) with no change in MHC IIa fibers (fast-twitch) pre- to post-PRT, respectively. P(o) in MHC I fibers increased 34% ( P<0.05) as a result of the training with no change in MHC IIa fibers. The mean MHC I Ca(2+) activation threshold (minimal amount of Ca(2+) necessary to induce tension) increased from 6.83 +/- 0.02 to 6.91 +/- 0.01 ( P<0.05), as did the mean half-maximal activation (pCa(50)), 5.51 +/- 0.02 to 5.71 +/- 0.03 ( P<0.05) with PRT. The slope of the Hill plot above ( n(1)) the pCa(50) for MHC I did not change significantly with the PRT. However, the slope of the Hill plot below ( n(2)) the pCa(50) for MHC I demonstrated an increase ( P<0.05) with training. There were no differences with MHC IIa fibers with PRT for any of the variables measured. In conclusion, the results of this investigation indicate that myofibril Ca(2+) sensitivity and activation properties are altered in MHC I, but not MHC IIa fibers with PRT in older women. The alterations in the MHC I Ca(2+) properties appear to have an effect on the mechanisms involved with skeletal muscle adaptability in older women following PRT. PMID- 12111252 TI - Stimulation of Xenopus oocyte Na(+),K(+)ATPase by the serum and glucocorticoid dependent kinase sgk1. AB - The serum and glucocorticoid-dependent kinase-1 (sgk1) is expressed in a wide variety of tissues including renal epithelial cells. As it is up-regulated by aldosterone, it is considered to participate in the regulation of renal Na(+) reabsorption. Indeed, co-expression of sgk1 with the renal epithelial Na(+) channel (ENaC) augments Na(+) channel activity. The aim of the present study was to examine possible effects of sgk1 on Na(+)/K(+)-ATPase activity. To this end dual-electrode voltage-clamp experiments were performed in Xenopus oocytes expressing the active kinase (S422D)sgk1 or the inactive mutant (K127N)sgk1. Na(+)/K(+)-ATPase activity was estimated from the hyperpolarization (delta V(m)) and the outwardly-directed current ( I(P)) created by addition of extracellular K(+) in the presence of K(+) channel blocker Ba(2+). Both delta V(m) and I(P) were significantly larger in oocytes expressing (S422D)sgk1 than in those expressing (K127N)sgk1 or having been injected with water. I(P) was fully inhibited by ouabain. Ion-selective microelectrodes showed that the stimulation of pump current was not the result of altered cytosolic Na(+) activity or pH. The present results thus point to an additional action of sgk1 that may participate in the regulation of renal tubular Na(+) transport. Moreover, sgk1 may be involved in the regulation of Na(+)/K(+)-ATPase in extrarenal tissues. PMID- 12111253 TI - Cardiac microtubules are more resistant to chemical depolymerisation in streptozotocin-induced diabetes in the rat. AB - Evidence exists for a specific diabetic cardiomyopathy independent of concurrent vascular disease. Our aim was to test the hypothesis that a change in the microtubular cytoskeleton may contribute to cardiac dysfunction in type-1 diabetes. Resting sarcomere length and characteristics of unloaded shortening were measured in ventricular myocytes from rats 2 months after injection of streptozotocin (STZ). Microtubular density and organisation were assessed using immunofluorescence confocal microscopy and the effects of microtubule disruption by colchicine on shortening and microtubules were examined. Diabetic myocytes showed a significant reduction in resting sarcomere length and a 30% increase in time to peak shortening. The microtubule disruptor colchicine (10 micromol/l) had no effect on the amplitude or kinetics of shortening in myocytes from control or diabetic rats. Cardiac microtubular density and organisation were similar in control and diabetic animals, yet although colchicine significantly reduced microtubule density in control myocytes, microtubules in diabetic myocytes were resistant to its effects. These observations of an increase in microtubular stability in STZ-diabetes of 2 months duration imply a disruption to the normal balance between populations of dynamic and drug-stable microtubules. Such disruption has been observed in other pathological conditions and may contribute to diabetic cardiomyopathy. PMID- 12111254 TI - ATP and nitric oxide modulate a Ca(2+)-activated non-selective cation current in macrovascular endothelial cells. AB - We have studied the properties of a non-selective cation current (NSC(Ca)) in macrovascular endothelial cells derived from human umbilical vein (EA cells) that is activated by an increase of intracellular Ca(2+) concentration, [Ca(2+)](i). Current-voltage relationships are linear and the kinetics of the current is time independent. Current-[Ca(2+)](i) relationships were fitted to a Ca(2+) binding site model with a concentration for half-maximal activation of 417 +/- 76 nM, a Hill coefficient of 2.3 +/- 0.8 and a maximum current of -23.9 +/- 2.7 pA/pF at 50 mV. The Ca(2+)-activated channel is more permeable to Na(+) than for Cs(+) ( P(Cs)/ P(Na)=0.58, n=7), but virtually impermeable to Ca(2+). Current activation was transient if ATP was omitted from the pipette solution. The maximal currents at 300 and 500 nM [Ca(2+)](i) were smaller than in the absence of ATP, but were not significantly different at 2 microM. The intracellular Ca(2+) concentration for half-maximal activation of the Ca(2+)-activated current was shifted to 811 +/ 12 nM in the absence of ATP. Substitution of ATP by the non-hydrolysable ATP analogue adenylylimidodiphosphate (AMP-PNP) did not affect current activation. Sodium nitroprusside (SNP) decreased NSC(Ca) in a concentration-dependent manner. The nitric oxide (NO) donors S-nitroso- N-acetylpenicillamine (SNAP) and 3 morpholinosydnonimine (SIN-1) also inhibited NSC(Ca). In contrast, nitro- L arginine (NLA), which inhibits all NO-synthases, potentiated NSC(Ca), whereas superoxide dismutase (SOD), which inhibits the breakdown of NO, inhibited NSC(Ca). It is concluded that the Ca(2+)-activated non-selective action channel in EA cells is modulated by the metabolic state of the cell and by NO. PMID- 12111255 TI - Effects of hyperosmotic shrinking on ventricular myocyte shortening and intracellular Ca(2+) in streptozotocin-induced diabetic rats. AB - Evidence exists for a specific diabetic cardiomyopathy independent of concurrent vascular disease. We tested the hypothesis that chronic hyperglycaemia found in streptozotocin- (STZ) induced diabetic rats leads to an altered response to and contractile effects of hyperosmotic shrinkage in ventricular myocytes. Analysis confirmed significant hyperglycaemia and revealed significant blood hyperosmolarity in STZ-treated rats. Myocyte volume changes, shortening and intracellular Ca(2+) ([Ca(2+)](i)) transients were measured in cells superfused with normal Tyrode (NT, 300 mmol/kg) and then hyperosmotic Tyrode (HT, 440 mmol/kg) at 35-36 degrees C. Shrinking significantly reduced the amplitude of shortening, whilst the [Ca(2+)](i) transient was significantly increased. The time course of both shortening and the [Ca(2+)](i) transient were prolonged in myocytes from STZ-treated compared to control rats. Time to peak shortening was 130.3 ms in STZ compared to 100.2 ms in control myocytes. Time to peak [Ca(2+)](i) transient was 70.8 ms in STZ compared to 44.6 ms in control myocytes and the time from peak to half recovery was 191.0 ms in STZ compared to 169.1 ms in control myocytes. Fractional SR Ca(2+) release, assessed by the application of caffeine, was increased by shrinking. However, the effects of raised extracellular osmolarity on volume changes, contractility and [Ca(2+)](i) were not altered by the chronic hyperglycaemia found in STZ-treated rats. PMID- 12111256 TI - Separation of neonatal rat ventricular myocytes and non-myocytes by centrifugal elutriation. AB - The preparation of pure cardiac myocyte cultures from neonatal rats is hampered by the presence of non-myocytes, which can proliferate during culturing, thereby causing a progressive decrease in the proportion of myocytes. In order to obtain myocyte cell suspensions of high purity, a method based on centrifugal elutriation was developed. Cardiac cells, isolated from neonatal rat heart ventricles, were subjected to elutriation using flow rates that increased step wise from 20 to 80 ml/min. The cell fraction obtained at 80 ml/min consisted of 68-90% myocytes. Still, upon culturing, the remaining non-myocytes proliferate, causing the proportion of myocytes to decrease to 60 +/- 2% at day 5. A second elutriation protocol was developed in which myocytes and non-myocytes were separated after a period of co-culturing for 4-5 days. By this approach a fibroblast-rich cell fraction (87 +/- 5%) and a myocyte-rich cell fraction (82 +/ 6%) were obtained. In conclusion, centrifugal elutriation creates the opportunity to separate neonatal rat myocytes from non-myocytes, either freshly isolated or after a period of culturing. Particularly, cell separation after a period of culturing ventricular cells offers an advantage to analyse the experimental effects on myocytes and non-myocytes separately. PMID- 12111257 TI - Subtotal colectomy with primary ileorectostomy is effective for unlocalized, diverticular hemorrhage. AB - BACKGROUND: A large proportion of patients with acute colonic diverticular bleeding undergo emergency surgery without successful prior localization of the bleeding site. This study sought to determine the surgical techniques of choice for unlocalized, diverticular hemorrhage. METHODS: We reviewed the data on 42 consecutive patients (median age 76 years, range 44-91) with acute colonic diverticular bleeding operated on between November 1993 and December 2000. Mean follow-up was 4.1 years. RESULTS: Preoperative localization of the bleeding site was possible in six patients (14%), by colonoscopy in two and by angiography in four. Ten patients underwent segmental colectomy with primary anastomosis (5 "directed", 5 "blind") and 32 subtotal colectomy with primary ileorectostomy (1 "directed", 31 "blind"). Subtotal colectomy is the more extensive surgical procedure (longer resected bowel, greater blood loss), and although it was performed in older patients, there were no significant differences between segmental and subtotal colectomy with respect to operation time, morbidity, mortality, hospital stay, number of bowel movements, continence scores, rebleeding rate, or patient satisfaction. CONCLUSIONS: Subtotal colectomy with primary ileorectostomy for unlocalized colonic diverticular bleeding is a safe and effective surgical procedure providing complete bleeding control and preserving continence. PMID- 12111258 TI - The impact of comorbidity on the overall survival and the cause of death in patients after colorectal cancer resection. AB - BACKGROUND: Retrospective investigation to identify associations between certain patient characteristics and survival in 531 patients with resected colorectal cancer (CRC). Special reference is given to a standardized comorbidity. METHODS: To compare different levels of exposure we determined hazard ratios (HR) in Cox proportional hazards models for survival times and odds ratios (OR) in logistic regression models. RESULTS: Overall survival was associated with tumor stages (III+IV vs. I+II; HR 7.48), tumor differentiation (low vs. high; HR 1.84), blood transfusions (>2 vs. < or =2; HR 1.88), and comorbidity (Charlson Comorbidity Index >2 vs. < or =2; HR 1.77). Low tumor stage (I+II vs. III+IV; OR 11.1), elevated Charlson Comorbidity Index (>2 vs. < or =2; OR 3.83), and longer ICU stay (>2 days vs. < or =2 days; OR 3.40) more frequently lead to non-cancer related death than to cancer-related death. CONCLUSION: Standardized comorbidity should be considered as a factor in survival studies of CRC. PMID- 12111259 TI - Esophageal cancer patients surviving 6 years after esophagectomy. AB - BACKGROUND: Esophageal cancer is one of the most malignant tumors, with a dismal prognosis in spite of recent advances in early diagnosis and extended lymphadenectomy. These patients need to be stratified according to prognostic variables for precise identification of high-risk group. MATERIAL AND METHODS: Seventy-six patients with esophageal carcinoma were uniformly treated with curative intent between 1980 and 1992 with at least 6 years follow-up. Results and prognostic factors of long-term survival were analyzed by univariate and multivariate analyses. RESULTS: Thirty patients (39.5%) survived 6 years, and the remaining 46 patients died within 6 years: recurrent esophageal cancer in 27 and causes unrelated to esophageal cancer in 19. The 1-, 2-, 3-, and 6-year overall survival rates in all 76 patients were 77.6%, 57.9%, 53.9%, and 39.5%, respectively. The factors influencing survival rate verified by univariate analysis were Borrmann classification (0, 1 vs. 2, 3), size of tumor (< or =3.0 vs. >3.0 cm), depth of invasion (T1, 2 vs. T3, 4), pN category (pN0 vs. pN1), number of lymph node metastasis (< or =4vs. >4), metastatic lymph node ratio (< or =0.1 vs. >0.1), time of operation (< or =480 vs. >480 min), and amount of perioperative blood transfusion given (< or =2 vs. >2 U). Among the significant variables independent prognostic factors for survival determined by multivariate analysis were metastatic lymph node ratio and amount of blood transfusion. CONCLUSIONS: A significant number of patients can thus apparently be cured of esophageal carcinoma by extensive resection. Patients with many metastatic lymph nodes and much blood transfusion, on the other hand, should receive appropriate treatment against such esophageal carcinoma. PMID- 12111260 TI - Efficacy of ukrain in the treatment of pancreatic cancer. AB - BACKGROUND: This monocentric study evaluated the effect of ukrain in the treatment of pancreatic cancer. MATERIAL AND METHODS: Between January 1996 and December 1999 we treated 21 patients with 10 mg ukrain every second day x10. The control group received supportive treatment only. RESULTS: Ukrain treatment was well tolerated. Mean values on pain measure and Karnofsky index were significantly better in the ukrain group than in controls ( P<0.05). One-year survival was 76% in the ukrain group, compared to 9.5% in the control group. Median survival after treatment with ukrain was 574 days, compared to 197 days in the control group. CONCLUSIONS: Our data demonstrate that ukrain improves quality of life in patients suffering from advanced pancreatic cancer and significantly prolongs survival time in these patients. PMID- 12111261 TI - Conversion of locally inoperable primary rectal cancer with multiple liver metastases to an option for cure after local down-staging and hepatic arterial infusion chemotherapy. AB - AIMS: This case report presents a patient with local unresectable primary rectal cancer and multiple synchronous liver metastases. METHODS: The treatment consisted of primary tumor resection after down-staging by local irradiation followed by hepatic arterial infusion chemotherapy. RESULTS: The patient is now without any signs of tumor growth 44 months after beginning of treatment. PMID- 12111262 TI - Early postoperative results of surgery for rectal carcinoma as a function of the distance of the tumor from the anal verge: results of a multicenter prospective evaluation. AB - BACKGROUND: The problems associated with rectal surgery are frequently discussed with no reference being made to the distance of the tumor from the anal verge. This study examined the effect of the location of the tumor on early postoperative results. PATIENTS AND METHODS: This was a multicenter study involving 75 German hospitals and 3756 patients, of whom 1463 had rectal carcinoma. On the basis of the location of the tumor (distance from the anal verge), four groups were distinguished: <4, 4-7.9, 8-11.9, and 12-16 cm. RESULTS: Resection and abdominoperineal resection rates and the incidence of postoperative complications depended on the location of the tumor. Significantly higher resection rates and fewer specific complications, and a significant reduction in overall postoperative morbidity were found with tumor locations more than 8 cm from the anal verge. The highest anastomotic leak rate was observed with anastomoses less than 7 cm from the anal verge. The logistic regression showed that the distance of the tumor from the anal verge is an independent variable for the development of an anastomotic leak. CONCLUSIONS: Early results are greatly affected by the location of the rectal carcinoma. This applies to both abdominoperineal resection rates and specific postoperative complications, such as anastomotic leak rate and operation morbidity in general. PMID- 12111263 TI - Childbirth and incontinence: a prospective study on anal sphincter morphology and function before and early after vaginal delivery. AB - PURPOSE: Disturbance of anal continence is a well-known problem after vaginal delivery. However, only few and incongruent data on the incidence and pathogenesis of postpartum incontinence are available. This study examined the effects of vaginal delivery on anal continence prospectively. METHODS: In 42 unselected women anal vector manometry and endoanal ultrasonography were performed, and pudendal nerve terminal motor latency (PNTML) and rectal sensibility were measured in the 32th week of pregnancy and 6 weeks after delivery. Continence was evaluated according to the Kelly-Holschneider score. Patients with occult sphincter defects were additionally followed-up 12 weeks after vaginal delivery. To exclude any effect of pregnancy alone ten patients with elective cesarian section served as controls. RESULTS: Overall continence after vaginal delivery did not differ significantly from that before delivery, there was a significant reduction in postpartum anal squeeze and resting pressures in all patients. Obstetric tears of grade III or IV occurred in 9% of the patients. Endosonography revealed occult lesions of the internal and external anal sphincter in an additional 19% of women who clinically seemed to have an intact sphincter. Manometric results and continence in these women did not differ significantly from those with intact sphincter and remained unchanged after 12 weeks. PNTML and rectal sensibility were not affected by vaginal delivery. After cesarian section there were no changes in continence, anal pressures, rectal sensibility, or PNTML. CONCLUSIONS: Vaginal delivery leads to direct mechanical trauma to the anal sphincters, while stretch and distension of the pudendal nerve seem to be of minor importance. Only endoanal ultrasonography is suitable for detection of occult sphincter lesions. PMID- 12111264 TI - Drifting towards a diffuse control model of exploratory motor learning: A comparison of global and within-trial performance measures. AB - Accurate measurement is crucial for understanding the processes that underlie exploratory patterns in motor learning. Accordingly, measures of learning should be sensitive to the changes that take place during skill acquisition. Most studies, however, use trial-based global measures that assess performance but do not actually measure gradual changes taking place within trials. The present study attempted to remedy this shortcoming by analysing a visual adaptation task, and comparing traditional global measures of learning with new, within-trial measures. Movement time was the only global measure sensitive to changes in the movement trajectory during learning. Three new measures were expected to reveal changes to the movement trajectory that are associated with learning: (i) the length of runs, (ii) change of trajectory angle in relation to the target, and (iii) drift (change in distance from the target). All three measures were sensitive to learning and indicated a gradual straightening of the movement trajectories over trials. Furthermore, three different methods to partition trajectories into segments were examined. The new within-trial measures, irrespective of partitioning method, prove promising for the development of a diffuse control model of exploratory learning. PMID- 12111265 TI - Adaptation of reflexive feedback during arm posture to different environments. AB - In this study we have examined the ability of the central nervous system (CNS) to use spinal reflexes to minimize displacements during postural control while continuous force perturbations were applied at the hand. The subjects were instructed to minimize the displacements of the hand from a reference position that resulted from the force perturbations. The perturbations were imposed in one direction by means of a hydraulic manipulator of which the virtual mass and damping were varied. Resistance to the perturbations came from intrinsic and reflexive stiffness, and from the virtual environment. It is hypothesized that reflexive feedback during posture maintenance is optimally adjusted such that position deviations are minimal for a given virtual environment. Frequency response functions were estimated, capturing all mechanical properties of the arm at the end point (hand) level. Intrinsic and reflexive parameters were quantified by fitting a linear neuromuscular model to the frequency responses. The reflexive length feedback gain increased strongly with damping and little with the eigenfrequency of the total combined system (i.e. arm plus environment). The reflexive velocity feedback gain decreased slightly with relative damping at the largest eigenfrequency and more markedly at smaller eigenfrequencies. In the case of highest reflex gains, the total system remained stable and sufficiently damped while the responses of only the arm were severely underdamped and sometimes even unstable. To further analyse these results, a model optimization was performed. Intrinsic and reflexive parameters were optimized such that two criterion functions were minimized. The first concerns performance and penalized hand displacements from a reference point. The second one weights afferent control effort to avoid inefficient feedback. The simulations showed good similarities with the estimated values. Length feedback was adequately predicted by the model for all conditions. The predicted velocity feedback gains were larger in all cases, probably indicating a mutual gain limiting relation between length and velocity afferent signals. The results suggest that both reflex gains seem to be adjusted by the CNS, where in particular the length feedback gain was optimal so as to maximize performance at minimum control effort. PMID- 12111266 TI - Self-selected modular recurrent neural networks with postural and inertial subnetworks applied to complex movements. AB - It has been shown that dynamic recurrent neural networks are successful in identifying the complex mapping relationship between full-wave-rectified electromyographic (EMG) signals and limb trajectories during complex movements. These connectionist models include two types of adaptive parameters: the interconnection weights between the units and the time constants associated to each neuron-like unit; they are governed by continuous-time equations. Due to their internal structure, these models are particularly appropriate to solve dynamical tasks (with time-varying input and output signals). We show in this paper that the introduction of a modular organization dedicated to different aspects of the dynamical mapping including privileged communication channels can refine the architecture of these recurrent networks. We first divide the initial individual network into two communicating subnetworks. These two modules receive the same EMG signals as input but are involved in different identification tasks related to position and acceleration. We then show that the introduction of an artificial distance in the model (using a Gaussian modulation factor of weights) induces a reduced modular architecture based on a self-elimination of null synaptic weights. Moreover, this self-selected reduced model based on two subnetworks performs the identification task better than the original single network while using fewer free parameters (better learning curve and better identification quality). We also show that this modular network exhibits several features that can be considered as biologically plausible after the learning process: self-selection of a specific inhibitory communicating path between both subnetworks after the learning process, appearance of tonic and phasic neurons, and coherent distribution of the values of the time constants within each subnetwork. PMID- 12111268 TI - Force and torque production in static multifinger prehension: biomechanics and control. I. Biomechanics. AB - We studied the coordinated action of fingers during static tasks involving exertion of force and torque on a handheld object. Subjects were asked to keep a handle with an attachment that allowed for independent change of the suspended load (0.5-2.0 kg) and external torque (0.375-1.5 N m) in a vertical position while applying minimal effort. Normal and shear forces were measured from the thumb; normal forces only were measured from the four fingers. EXPERIMENTAL RESULTS: (1) the thumb shear force increased during supination efforts and decreased during pronation efforts; (2) the total moment of the normal finger forces only counterbalanced approximately 50% of the external torque, hence shear forces accounted for approximately one-half of the total torque exerted on the object; (3) the total normal force increased with external torque, and the total force magnitude did not depend on the torque direction; (4) the forces of the 'peripheral' (index and little) fingers depended mainly on the torque while the forces exerted by the 'central' (middle and ring) fingers depended both on the load and torque; (5) there was a monotonic relationship between the mechanical advantage of a finger (i.e., its moment arm during torque production) and the force produced by that finger; and (6) antagonist finger moments acting opposite to the intended direction of the total moment were always observed - at low torques the antagonist moments were as high as 40-60% of the agonist moments. MODELING: A three-zone model of coordinated finger action is suggested. In the first zone of load/torque combinations, activation of antagonist fingers (i.e., fingers that generate antagonist moments) is necessary to prevent slipping. In the second zone, the activity of agonist fingers is sufficient for preventing slips. In the third zone, the performer has freedom to choose between either activating the antagonist fingers or redistributing activities amongst the agonist fingers. The findings of this study provide the foundation for neural network and optimization modeling described in the companion paper [Zatsiorsky et al. (2002) Biol Cybern DOI 10.1007/s00422-002-0320-7]. PMID- 12111267 TI - Force and torque production in static multifinger prehension: biomechanics and control. II. Control. AB - The coordination of digits during combined force/torque production tasks was further studied using the data presented in the companion paper [Zatsiorsky et al. Biol Cybern this issue, Part I]. Optimization was performed using as criteria the cubic norms of (a) finger forces, (b) finger forces normalized with respect to the maximal forces measured in single-finger tasks, (c) finger forces normalized with respect to the maximal forces measured in a four-finger task, and (d) finger forces normalized with respect to the maximal moments that can be generated by the fingers. All four criteria failed to predict antagonist finger moments when these moments were not imposed by the task mechanics. Reconstruction of neural commands: The vector of neural commands c was reconstructed from the equation c=W(-1)F, where W is the finger interconnection weight matrix and F is the vector of finger forces. The neural commands ranged from zero (no voluntary force production) to one (maximal voluntary contraction). For fingers producing moments counteracting the external torque ('agonist' fingers), the intensity of the neural commands was well correlated with the relative finger forces normalized to the maximal forces in a four-finger task. When fingers produced moments in the direction of the external torque ('antagonist' fingers), the relative finger forces were always larger than those expected from the intensity of the corresponding neural commands. The individual finger forces were decomposed into forces due to 'direct' commands and forces induced by enslaving effects. Optimization of the neural commands resulted in the best correspondence between actual and predicted finger forces. The antagonist moments are, at least in part, due to enslaving effects: strong commands to agonist fingers also activated antagonist fingers. PMID- 12111269 TI - Temporal-to-spatial dynamic mapping, flexible recognition, and temporal correlations in an olfactory cortex model. AB - This paper proposes temporal-to-spatial dynamic mapping inspired by neural dynamics of the olfactory cortex. In our model the temporal structure of olfactory-bulb patterns is mapped to the spatial dynamics of the ensemble of cortical neurons. This mapping is based on the following biological mechanism: while anterior part of piriform cortex can be excited by the afferent input alone, the posterior areas require both afferent and association signals, which are temporally correlated in a specific way. One of the functional types of the neurons in our model corresponds to the cortical spatial dynamics and encodes odor components, and another represents temporal activity of association-fiber signals, which, we suggest, may be relevant to the encoding of odor concentrations. The temporal-to-spatial mapping and distributed representation of the model enable simultaneous rough cluster classification and fine recognition of patterns within a cluster as parts of the same dynamic process. The model is able to extract and segment the components of complex odor patterns which are spatiotemporal sequences of neural activity. PMID- 12111270 TI - On the distribution of fast-phase intervals in optokinetic and vestibular nystagmus. AB - Histograms of fast-phase intervals in human optokinetic and vestibular nystagmus were generated, and fitted to statistical distributions used in previous studies. The distributions did not depend on stimulation type (optokinetic or vestibular). An inverse Gaussian or a gamma distribution fitted the data better than did a reciprocal Gaussian distribution, but none fitted the data especially well. In some cases, however, the interpretation of these distributions is more physiologically satisfactory than in others. Recommendations are made on which class of models is preferred, and the experiments needed to support the particular models. Our results call into question the validity of previous studies that fit statistical distributions to data sets of a size comparable to ours. PMID- 12111271 TI - A feedforward model of suppressive and facilitatory habituation effects. AB - Simple exposure to repetitive stimulation is known to induce short-term learning effects across a wide range of species. These effects can be both suppressive and facilitatory depending on stimulus conditions: repetitive presentation of a weak stimulus decreases the strength of the response (habituation), whereas presentation of a tonic stimulus following a series of weak stimuli transiently increases the response strength (dishabituation). Although these phenomena have been comprehensively characterized at both behavioral and cellular levels, most existing models of nonassociative learning focus exclusively on the suppressive or facilitatory changes in response, and do not attempt to relate cellular events to behavior. I propose here a feedforward model of habituation effects that explains both suppressive and facilitatory changes in response relying on the interaction between excitatory and inhibitory processes that develop in parallel on two different timescales. The model's properties are used to explain the rate sensitivity property of habituation and recovery and stimulus dishabituation. PMID- 12111272 TI - The discontinuous nature of motor execution II. Merging discrete and rhythmic movements in a single-joint system -- the phase entrainment effect. AB - Initiation of rapid discrete flexion movements is significantly altered when a secondary rhythmic movement is performed simultaneously with the same limb; the onset of a stimulus-evoked discrete movement tends to occur time-locked to the oscillation: i.e., the rhythmic movement entrains the discrete response. This nonlinear interaction may reflect a specific principle of coordination of motor tasks which are simultaneously executed with the same effector. This part II of a tripartite research report on such single-muscle multiple-task coordination investigates the contribution of the dynamic properties of the muscle and its reflex circuitry to phase entrainment. Assuming a simple threshold-linear relationship between the control signals generated by the central nervous system and the observable kinematic and electromyographic signals, a secondary rhythmic movement will cause an additional phase-dependent delay between the central "go" command and the first observable change in actual kinematics of the compound movement. Several indicators for such threshold-linear interaction are derived and tested on real data obtained in psychophysical experiments. Four healthy subjects performed rapid lateral abductions of the index finger in response to a visual "go" signal. During a portion of the experiments, subjects produced additional low-amplitude oscillatory movements before stimulus presentation with either the same finger (one-handed task), or with the index finger of the other hand (two-handed task). Results showed phase entrainment and modulation of reaction times when the cyclic and the discrete movements were simultaneously executed by the same finger. But there was no entrainment in the bimanual execution of the tasks. The model was capable of reproducing the observed effects. It is concluded that coordination of voluntary movements which are concurrently performed by the same effector involves specific discontinuous operations, which represents an essential part of the mechanism of motor coordination. Phase entrainment reflects this characteristic discontinuous behavior of the lower stages of motor execution and does not necessarily require nonlinear interaction of motor commands at higher levels of motor processing. PMID- 12111273 TI - Influence of anatomical, physical, and detection-system parameters on surface EMG. AB - Many previous studies were focused on the influence of anatomical, physical, and detection-system parameters on recorded surface EMG signals. Most of them were conducted by simulations. Previous EMG models have been limited by simplifications which did not allow simulation of several aspects of the EMG generation and detection systems. We recently proposed a model for fast and accurate simulation of the surface EMG. It characterizes the volume conductor as a non-homogeneous and anisotropic medium, and allows simulation of EMG signals generated by finite-length fibers without approximation of the current-density source. The influence of thickness of the subcutaneous tissue layers, fiber inclination, fiber depth, electrode size and shape, spatial filter transfer function, interelectrode distance, length of the fibers on surface, single-fiber action-potential amplitude, frequency content, and estimated conduction velocity are investigated in this paper. Implications of the results on electrode positioning procedures, spatial filter design, and EMG signal interpretation are discussed. PMID- 12111274 TI - Unified neurophysical model of EEG spectra and evoked potentials. AB - Evoked potentials -- the brain's transient electrical responses to discrete stimuli -- are modeled as impulse responses using a continuum model of brain electrical activity. Previous models of ongoing brain activity are refined by adding an improved model of thalamic connectivity and modulation, and by allowing for two populations of excitatory cortical neurons distinguished by their axonal ranges. Evoked potentials are shown to be modelable as an impulse response that is a sum of component responses. The component occurring about 100 ms poststimulus is attributed to sensory activation, and this, together with positive and negative feedback pathways between the cortex and thalamus, results in subsequent peaks and troughs that semiquantitatively reproduce those of observed evoked potentials. Modulation of the strengths of positive and negative feedback, in ways consistent with psychological theories of attentional focus, results in distinct responses resembling those seen in experiments involving attentional changes. The modeled impulse responses reproduce key features of typical experimental evoked response potentials: timing, relative amplitude, and number of peaks. The same model, with further modulation of feedback, also reproduces experimental spectra. Together, these results mean that a broad range of ongoing and transient electrocortical activity can be understood within a common framework, which is parameterized by values that are directly related to physiological and anatomical quantities. PMID- 12111275 TI - A spin-glass-like Lyapunov function for a neurotrophic model of neuronal development. AB - We derive a spin-glass-like energy or Lyapunov function for our previously studied neurotrophic model of anatomical synaptic plasticity and neuronal development. This function is then used in Monte-Carlo simulations of the model applied to the development of ocular dominance columns. We discuss the relationship between our model and other models, and speculate on the implications of underlying spin glass structures in many models of neuronal development, learning and plasticity. PMID- 12111276 TI - MEG responses during rhythmic finger tapping in humans to phasic stimulation and their interpretation based on neural mechanisms. AB - The phase-resetting experiment was applied to human periodic finger tapping to understand how its rhythm is controlled by the internal neural clock that is assumed to exist. In the experiment, the right periodic tapping movement was disturbed transiently by a series of left finger taps in response to impulsive auditory cues presented randomly at various phases within the tapping cycle. After each left finger tap, the original periodic tapping was reestablished within several tapping cycles. Influences of the disturbance on the periodic right finger tapping varied depending on the phase of the periodic right finger tapping at which each left finger tap was made. It was confirmed that the periodic tapping was disturbed not by the auditory cues but by the left finger taps. Based on this fact, in this paper each single left tap was considered as the stimulus, and the phase of the periodic tapping of the right index finger when the left tap was executed as the phase of the stimulus. Responses of the neural activities (magnetoencephalography, MEG), the tapping movement, and the corresponding muscle activities (electromyography) were simultaneously measured. Phase-resetting curves (PRCs) representing the degree of phase reset as a function of the phase of the stimulus were obtained both for the left sensorimotor cortex MEG response and for the right index finger tapping response. The shapes of both PRCs were similar, suggesting that the phase reset of the left sensorimotor cortex activities and that of the finger tapping rhythm were the same. Four out of eight subjects showed type-0 reset in Winfree's definition, and the others showed type-1 reset. For general limit-cycle oscillators, type-0 reset is obtained for relatively strong perturbations and type 1 for weak perturbations. It was shown that the transient response of MEG to the single left tap stimuli in type-0 subjects, where the phase was progressively reset, were different from those in type-1 subjects. Based on detailed analysis of the differences, a neural network model for the phase reset of the tapping rhythm is proposed. PMID- 12111277 TI - Modeling complex tone perception: grouping harmonics with combination-sensitive neurons. AB - Perception of complex communication sounds is a major function of the auditory system. To create a coherent precept of these sounds the auditory system may instantaneously group or bind multiple harmonics within complex sounds. This perception strategy simplifies further processing of complex sounds and facilitates their meaningful integration with other sensory inputs. Based on experimental data and a realistic model, we propose that associative learning of combinations of harmonic frequencies and nonlinear facilitation of responses to those combinations, also referred to as "combination-sensitivity," are important for spectral grouping. For our model, we simulated combination sensitivity using Hebbian and associative types of synaptic plasticity in auditory neurons. We also provided a parallel tonotopic input that converges and diverges within the network. Neurons in higher-order layers of the network exhibited an emergent property of multifrequency tuning that is consistent with experimental findings. Furthermore, this network had the capacity to "recognize" the pitch or fundamental frequency of a harmonic tone complex even when the fundamental frequency itself was missing. PMID- 12111279 TI - A metabolic cart for measurement of oxygen uptake during human exercise using inspiratory flow rate. AB - This study evaluated an ergo-spirometry system based on mixed expired gas for gas analyses and an inspiratory based determination of flow. There were 74 paired samples of oxygen uptake (VO(2)) and related variables including pulmonary ventilation (V(E)), fractional concentrations of expired CO(2) and O(2) ( F(E)CO(2) and F(E)O(2), respectively), as well as CO(2) output (VCO(2)) which were obtained from the metabolic cart and a Douglas bag system during 22 min submaximal and 5-8 min maximal running on a treadmill. For F(E)CO(2) and VCO(2) the metabolic cart gave readings that were 2.6% and 1.8% higher and for F(E)O(2) 0.2% lower than the Douglas bag method ( P<0.05). For the metabolic cart and the Douglas bag method the coefficient of variation (CV) for repeated determinations of VO(2) was 1.9% and 1.8%, respectively. For VO(2) and V(E), there were no significant differences between the two methods and the 95% confidence interval of the difference in VO(2) was within -30 and +20 ml min(-1). The CV of the differences in VO(2) between the two systems was 2.4% and it is concluded that a metabolic cart method based on inspiratory flow rate is suitable for measurement of VO(2) and V(E) during both submaximal and maximal exercise. PMID- 12111278 TI - Effect of endurance training and/or fish oil supplemented diet on cytoplasmic fatty acid binding protein in rat skeletal muscles and heart. AB - Endurance training and/or a fish oil supplemented diet affect cytoplasmic fatty acid binding protein (FABP(c)) content in rat skeletal muscles and heart. After 8 weeks of swimming, trained rats exhibited higher FABP(c) content in the extensor digitorum longus (EDL) and in the gastrocnemius than did control rats (30%). The FABP(c) increase was associated with an increase of citrate synthase activity (85% and 93%, respectively, in the two muscles), whereas lactate dehydrogenase activity decreased significantly. In contrast, in the soleus and in the heart we did not observe any effect of exercise either on FABP(c) or on the metabolic profile. Therefore, increasing oxidative capacities of muscle by exercise resulted in a concomitant increase of the FABP(c) content. Giving a polyunsaturated fatty acid (omega-3) supplemented diet for eight weeks induced a large rise of the FABP(c) in EDL (300%), gastrocnemius (250%), soleus (50%) and heart (15%) without a concurrent accumulation of intramuscular triglycerides or modification of the citrate synthase activity, suggesting that polyunsaturated fatty acids may increase FABP(c) content by up-regulating fatty acid metabolism genes via peroxisome proliferator-activated receptor alpha activation. Endurance trained rats fed with an omega-3 diet had similar FABP(c) content in the gastrocnemius muscle when compared to sedentary omega-3 fed rats, whereas an additive effect of exercise and diet was observed in the EDL. The FABP(c) in the soleus and in the heart of rats fed with omega-3 supplements remained constant whether rats performed exercise or not. As a result, both exercise and omega-3 enriched diet influenced FABP(c) content in muscle. These two physiological treatments presumably acted on FABP(c) content by increasing fatty acid flux within the cell. PMID- 12111280 TI - Theoretical and practical considerations in the application of whole body plethysmography to sleep research. AB - The sleep-wake state has a profound influence on many, perhaps most, aspects of normal physiology and is strongly implicated in the mediation (or remediation) of impaired health and performance. Many sleep disorders stem from abnormal respiratory anatomy or sleep-induced changes in respiratory control, underscoring the need for research into the effects of the sleep-wake state on respiratory control processes. Whole body plethysmography is being increasingly used to study respiration in freely behaving animals, and is especially well suited to studies of sleeping animals and human subjects. The method is simple in principle, but care is required in its application to ensure reliable results, and there are circumstances in which it is an inappropriate technique. This review describes the main advantages, pitfalls, and limitations inherent in the use of whole body plethysmography for non-invasive measurement of lung ventilation and metabolic rate in sleeping animals. Sources of potential error, and ways of avoiding such errors, are discussed, with reference to studies involving animal models of sleep related breathing disorders. PMID- 12111281 TI - Relationship between passive properties of the calf muscles and plantarflexion concentric isokinetic torque characteristics. AB - The purpose of this study was to use a model of aging to examine the relationships between passive properties of the calf muscles and plantarflexion concentric isokinetic torque characteristics. Eighty-one active women 20-84 years of age were tested using a Kin-Com isokinetic dynamometer interfaced with electromyography (EMG). The passive properties were tested by stretching the muscles from relaxed plantarflexion to a maximal dorsiflexion (DF) angle at a rate of 5 degrees.s(-1) (0.087 rad.s(-1)) with minimal raw EMG activity (<0.05 mV). The maximal concentric torque was tested from maximal passive DF into plantarflexion at four randomly ordered velocities of 30, 60, 120, and 180 degrees.s(-1). Pearson correlation coefficients (Bonferroni adjusted) indicated a hierarchical order of high to moderate positive correlations between four passive properties and the peak and mean concentric torque for all test velocities. Correlation coefficients for the four passive properties ranged from 0.50 to 0.78 ( P<0.001), and the coefficients of determination ( r(2)) from higher to lower were: (1) maximal DF passive resistive torque ( r(2): 0.50-0.62), (2) length extensibility ( r(2): 0.40-0.49), (3) maximal muscle length ( r(2): 0.28-0.41), and (4) passive elastic stiffness in the last half of the full-stretch range of motion ( r(2): 0.25-0.31). The maximal DF passive resistive torque and the length extensibility accounted for 50-62% and 40-49% of the variability in the concentric torque, respectively. The results indicate that the concentrically stronger calf muscles of active women were positively correlated with passively stronger, longer, and stiffer calf muscles, which are characteristics of the calf muscles of younger women. Further studies are needed to examine whether therapeutic interventions, such as stretching and strengthening, can promote adaptations in the calf muscles of older women to attain these more youthful characteristics. PMID- 12111283 TI - In vitro interleukin-6 release in whole blood cultures in samples taken at rest from triathletes and professional rugby players. AB - The aim of this study was to investigate the endotoxin-induced interleukin-6 (IL 6) release in whole blood cultures from samples taken at rest, 24 h post exercise, from a control group of recreationally trained individuals (C), a group of highly trained triathletes (TA) and a group of highly trained professional rugby players (RP). Fifteen RP [mean (SD): age 26 (3) years, height 1.90 (0.2) m, body mass 104.5 (12.2 kg)], 13 male TA [age 33 (5) years, height 1.78 (0.1) m, body mass 76.3 (12.6) kg] and eight recreationally active male volunteers [age 28 (6) years, height 1.80 (0.1) m, body mass 72.3 (7.3) kg] participated in the study. Plasma IL-6 concentration and in vitro IL-6 synthesis by whole blood cultures were measured in samples taken at rest. Plasma IL-6 concentration was significantly higher ( P<0.01) for the RP and TA groups than for C, as were the in vitro basal and endotoxin activated concentrations. However, after endotoxin stimulation, newly induced IL-6 concentration was significantly lower ( P<0.01) in the RP and TA than in the C group. Therefore, professional rugby players have a similar IL-6 release of whole blood cultures in vitro to that of triathletes. Specifically, mononuclear cells appear to be chronically activated to spontaneously release IL-6, but have a decreased capacity to respond to a further stimulus. Amongst possible explanations for this, the most likely is counter regulation due to already elevated IL-6 release. PMID- 12111282 TI - Serum lipoprotein cholesterols in older oarsmen. AB - We evaluated effects of age and rowing on concentrations of lipids and lipoprotein cholesterols in the blood. Maximal oxygen uptake (VO(2max)), and concentrations of total cholesterol (TC), triglyceride (TG), low-density lipoprotein cholesterol (LDL-C), and high-density lipoprotein cholesterol (HDL-C) were measured in 17 oarsmen [mean (SD)] [age 64 (4) years, body mass 69 (6) kg] and in sedentary men [age 65 (3) years, body mass 70 (7) kg] who were matched on the basis of body size. Also the variables were obtained from young oarsmen [age 22 (2) years, body mass 70 (4) kg] and young sedentary men [age 22 (3) years, body mass 69 (7) kg]. The percentage body fat of the older oarsmen was lower than that of the older sedentary men [18 (4)% compared to 23 (4)%, P<0.05], but it was similar to that of the young sedentary men [17 (4)%]. Although older oarsmen possessed a lower VO(2max) than the young oarsmen [3.0 (0.4) l.min(-1) compared to 4.1 (0.3) l.min(-1), P<0.01], they showed a VO(2max) similar to that of the young sedentary men [3.1 (0.5) l.min(-1)] but a higher value than obtained from the older sedentary men [2.2 (0.3) l.min(-1), P<0.05]. Although the indices of risk factors for coronary artery disease in the older oarsmen were higher than those in the young oarsmen [LDL-C/HDL-C 1.7 (0.2) compared to 1.3 (0.4), TC/HDL-C 3.1 (0.2) compared to 2.6(0.4), P<0.05], they were lower than those in both the older [2.1 (0.3), 3.6 (0.3), P<0.05] and the young sedentary men [2.1 (0.4), 3.5 (0.4), P<0.05]. The results suggest that rowing is an appropriate type of exercise for the promotion of health. PMID- 12111284 TI - Relationship between the curvature constant parameter of the power-duration curve and muscle cross-sectional area of the thigh for cycle ergometry in humans. AB - For high-intensity cycle ergometer exercise, the relationship between power output ( P) and its tolerable duration ( t) has been well characterized by the hyperbolic relationship: ( P- theta;(F)). t=W', where theta;(F) has been termed the "critical power" or "fatigue threshold". The curvature constant (W') reflects a constant amount of work which can be performed above theta;(F), and it may be regarded as a muscle energy store. The relationship of this energy store to muscle mass is not known. Therefore, the purpose of this study was to determine the relationships among W', accumulated peak oxygen deficit (accumulated peak O(2)-deficit), and muscle cross-sectional area (CSA) of the thigh for high intensity cycle ergometry in humans. A group of 17 healthy male subjects (aged 21 41 years) participated in this study. The theta;(F) and W' of the P- t hyperbolic relationship and the accumulated peak O(2)-deficit was calculated by standard procedures. The CSA of muscle, fat and bone in the right thigh were measured using ultrasonography. The mean (SD) of theta;(F), W', accumulated peak O(2) deficit, and muscle CSA of the thigh were 200.0 (17.8) W, 12.60 (2.94) kJ, 2.29 (0.41) l, and 185.3 (22.6) cm(2), respectively. The muscle CSA of the thigh was positively correlated with W' ( r=0.59, P<0.01) and with accumulated peak O(2) deficit ( r=0.54, P<0.05). The relationship between W' and accumulated peak O(2) deficit also showed a positive correlation ( r=0.63, P<0.005). Our results indicated that W' derived from the P- t hyperbolic curve as anaerobic working capacity is related to the CSA of muscle. PMID- 12111285 TI - Temperature-related overestimation of energy expenditure, based on heart-rate monitoring in obese boys. AB - An equation has been developed to reduce the error in predicting energy expenditure (EE) from heart rate (HR) monitoring, by correcting for climate related HR increases. To evaluate the effects of such a correction in a "real life" situation, a group of 9- to 14-year-old obese boys [mean (SD) 36.6 (3.3)% body fat; n=14] was monitored for 6 days during the summertime. An activity interview was taken daily. During times of outdoor activities, measurements of HR were corrected for the influence of ambient temperature ( T) using the aforementioned equation. Time spent outdoors was 196.7 (21.1) min/day at a T of 17-34 degrees C. Temperature correction reduced the time spent at HR>140 beats/min and >160 beats/min by 20-25% during outdoor activity. Estimated outdoor EE corrected for the influence of temperature was 2.46 (2.37) MJ; the uncorrected value was 2.68 (2.45) MJ. The respective values for the waking hours and over 24 h were 7.34 (2.77) MJ versus 7.55 (2.81) MJ and 10.83 (2.72) MJ versus 11.05 (2.77) MJ, respectively. The excessive HR measured in a warm summer climate could explain 8.8 (3.5)% of EE during outdoor activities, which resulted in a 2.9 (2.7)% overestimate of EE during daily waking hours and a 1.9 (1.8)% overestimate of the total 24-h EE. Thus, the influence of temperature on HR may, at least in part, explain the previously reported overestimate of EE calculated using the HR/oxygen consumption relationship versus that calculated using the doubly labelled water method. We recommend correcting HR for the influence of climate in studies using HR profiles to determine outdoor physical activity levels and/or EE during summertime. However, when total daily EE or waking hours EE is assessed, even in subjects who are engaging in long intervals of outdoor play, the temperature correction required is minimal. PMID- 12111286 TI - Onset of mild lower body negative pressure induces transient change in mean arterial pressure in humans. AB - Mild (0 to -20 mmHg) lower body negative pressure (LBNP) has traditionally been considered to elicit reflex responses mediated by cardiopulmonary baroreceptors only, without any arterial baroreflex involvement. Mild LBNP has therefore frequently been used to study the influence of cardiopulmonary baroreceptors on the human circulatory system. In a previous study we found that mean arterial pressure (MAP) was transiently but strongly affected by rapid (0.3 s) onset and release of -20 mmHg LBNP. In the present study we tested whether MAP is also transiently affected by slow onset and release of -20 mmHg LBNP. A group of 12 subjects participated in this study, which was approved by the local Ethics Committee. Heart rate, stroke volume, cardiac output, MAP, total peripheral resistance, acral and non-acral skin blood flow, and blood flow velocity in the brachial artery were continuously recorded during the pre-LBNP period, during LBNP and during the post-LBNP period. The LBNP was gradually applied and released over a 15 s period. The main finding was that MAP was transiently but strongly affected by the gradual onset of LBNP as mild as -20 mmHg. During onset of LBNP MAP was significantly ( P=0.003) lower than MAP in the pre-LBNP period. This shows that not only the cardiopulmonary baroreceptors but also the arterial baroreceptors must be activated during mild LBNP. PMID- 12111287 TI - Influence of time pressure and verbal provocation on physiological and psychological reactions during work with a computer mouse. AB - The overall aim of this study was to investigate whether time pressure and verbal provocation has any effect on physiological and psychological reactions during work with a computer mouse. It was hypothesised that physiological reactions other than muscle activity (i.e. wrist movements, forces applied to the computer mouse) would not be affected when working under stressful conditions. Fifteen subjects (8 men and 7 women) participated, performing a standardised text-editing task under stress and control conditions. Blood pressure, heart rate, heart rate variability, electromyography, a force-sensing computer mouse and electrogoniometry were used to assess the physiological reactions of the subjects. Mood ratings and ratings of perceived exertion were used to assess their psychological reactions. The time pressure and verbal provocation (stress situation) resulted in increased physiological and psychological reactions compared with the two control situations. Heart rate, blood pressure and muscle activity in the first dorsal interosseus, right extensor digitorum and right trapezius muscles were greater in the stress situation. The peak forces applied to the button of the computer mouse and wrist movements were also affected by condition. Whether the increases in the physiological reactions were due to stress or increased speed/productivity during the stress situation is discussed. In conclusion, work with a computer mouse under time pressure and verbal provocation (stress conditions) led to increased physiological and psychological reactions compared to control conditions. PMID- 12111288 TI - Effects of long-term training specificity on maximal strength and power of the upper and lower extremities in athletes from different sports. AB - Maximal concentric one repetition maximum half-squat (1RM(HS)), bench-press (1RM(BP)), power-load curves during concentric actions with loads ranging from 30% to 100% of 1RM(HS) and 1RM(BP)were examined in 70 male subjects divided into five groups: weightlifters (WL, n=11), handball players (HP, n=19), amateur road cyclists (RC, n=18), middle-distance runners (MDR, n=10) and age-matched control subjects (C, n=12). The 1RM(HS)values in WL, HP and RC were 50%, 29% and 28% greater, respectively, ( P<0.001-0.01) than those recorded for MDR and C. The half-squat average power outputs at all loads examined (from 30% to 100%) in WL and HP ( P<0.001 at 45% and 60% with HP) were higher ( P<0.05-0.001) than those in MDR, RC and C. Average power output at the load of 30% of 1RM(HS) in RC was higher ( P<0.05) than that recorded in MDR and C. Maximal power output was produced at the load of 60% for HP, MDR and C, and at the load of 45% for WL and RC. The 1RM(BP) in WL was larger ( P<0.05) than those recorded in HP, RC, MDR and C. In the bench press, average muscle power outputs in WL and HP were higher ( P<0.05-0.001) than those in MDR, RC and C, and were maximized at a load of 30% of 1RM for WL and HP, and at 45% for RC, MDR and C. In addition, the velocities that elicited the maximal power in the lower extremities were lower ( approximately 0.75 m.s(-1)) than those occurring in the upper extremities ( approximately 1 m.s(-1)). The data suggest that the magnitude of the sport-related differences in strength and/or muscle power output may be explained in part by differences in muscle cross-sectional area, fibre type distribution and in the muscle mechanics of the upper and lower limbs as well as by training background. PMID- 12111290 TI - Voice monitoring to measure emotional load during short-term stress. AB - It is well known that there is a relationship between the voice the human emotional status. Previous studies have demonstrated that changes of fundamental frequency ( f(0)), in particular, have a significant relationship with emotional load. The aim of the present study was to investigate how f(0) changes in response to an unknown emotionally stressful task under real-life conditions. A further question was whether repetitions of this task lead to an adaptation of f(0), indicating a lower emotional load. The participants of this study included 26 healthy males. f(0) and heart rate ( f(c)) were recorded for baseline testing (BLT) under relaxed laboratory conditions. Then the participants were asked to negotiate a natural obstacle by way of sliding down a rope hanging from a handlebar without any safety provisions, thus being exposed to the danger of a fall from a height of up to 12 m into shallow water (guerrilla slide I, GSI). The task was repeated after 30 min (GSII) and after 3 days of physical strain (GSIII). Immediately before starting the task the participants were asked to give a standardised speech sample, during which f(c) was recorded. The mode value of f(0) ( f(0,mode)) of the speech samples was used for further analysis. The mean (SD) value of f(0,mode) at BLT was 114.9 (14.8) Hz; this increased to 138.8 (19.6) Hz at GSI ( P<0.000), decreased to 135.9 (19.6) Hz at GSII and to 130.0 (21.5) Hz at GSIII ( P=0.012). The increase in f(c) was significantly different from BLT to GSI ( P<0.000). The repetitions of the task did not produce significant changes in f(c). It was shown that f(0,mode) is a sensitive parameter to describe changes in emotional load, at least in response to short-term psychoemotional stress, and seems to throw light upon the amount of adaptation caused by increased experience. PMID- 12111289 TI - Effects of vitamin E deficiency on fatigue and muscle contractile properties. AB - Radical-mediated oxidative damage of skeletal muscle membranes has been implicated in the fatigue process. Vitamin E (VE) is a major chain breaking antioxidant that has been shown to reduce contraction-mediated oxidative damage. We hypothesized that VE deficiency would adversely affect muscle contractile function, resulting in a more rapid development of muscular fatigue during exercise. To test this postulate, rats were fed either a VE-deficient (EDEF) diet or a control (CON) diet containing VE. Following a 12-week feeding period, animals were anesthetized and mechanically ventilated. Muscle endurance (fatigue) and contractile properties were evaluated using an in situ preparation of the tibialis anterior (TA) muscle. Contractile properties of the TA muscle were determined before and after a fatigue protocol. The muscle fatigue protocol consisted of 60 min of repetitive contractions (250 ms trains at 15 Hz; duty cycle=11%) of the TA muscle. Prior to the fatigue protocol, no significant differences existed in the force-frequency curves between EDEF and CON animals. At the completion of the fatigue protocol, muscular force production was significantly ( P<0.05) lower in the EDEF group (reduced by 69%) compared to CON group (reduced by 38%). Following the fatigue protocol, a right shift existed in the force-frequency curve at low stimulation frequencies (G (E360G). The other was a splicing mutation, 2027-2028+2delAAGT, which caused aberrant splicing transcripts, whereas 1876-2028del, 2027-2028insGTCTCTTCC ACTTCTTCC, and 2026 2028del were three aberrant transcripts that kept reading in-frame. In this report, an expression assay using SV40 transformed fibroblasts was performed to investigate the consequences of these two mutations on enzyme activity and protein levels. Molecular analysis in this study revealed that the two mutations 1079A>G and 2028+2delAAGT were the disease-causing mutations. PMID- 12111369 TI - Molecular analysis of the transferrin gene in a patient with hereditary hypotransferrinemia. AB - We previously reported a patient with hereditary hypotransferrinemia who suffered from severe anemia and growth retardation and was diagnosed on the basis of an extremely low level of serum transferrin (TF). By an isoelectric focusing analysis, we found that the patient and his father shared a variant TF protein with an abnormal isoelectric point. The study suggested that the patient was a compound heterozygote with a variant allele, encoding the mutant TF, of paternal origin and a null allele of maternal origin. In the present study, we investigated the TF gene of the patient and his family. We showed that the patient and his father shared a variant TF gene bearing a GAA to AAA transition at codon 394. This nucleotide substitution causes a nonconservative amino acid change from glutamate to lysine in amino acid residue 375 of the TF protein. This single amino acid mutation is predicted to cause a conformational change in the coiled region of the carboxyl-terminal iron-binding lobe. As for the maternal null allele, no mutation was found in either the coding region or the exon-intron boundaries, suggesting an abnormality in the transcription or stability of mRNA of maternal allele origin. PMID- 12111370 TI - Strategies for genome-wide association studies: optimization of study designs by the stepwise focusing method. AB - Recently, the use of genome-wide linkage disequilibrium (LD) analysis to localize traits has attracted much attention because of the introduction of high throughput genotyping systems. However, a limitation of such studies is often the total cost of genotyping in addition to sample size. Therefore, it is important to estimate optimal conditions for such a study given the total cost of genotyping. In the present study, we have introduced the "stepwise focusing method," in which candidate markers are selected in a stepwise fashion. In the first focusing step, samples from both case and control groups are genotyped at a certain number of single-nucleotide polymorphisms (SNPs) (for example, 50000), and the markers that exhibit significant intergroup differences by a chi(2) test are selected. In the first step, the risk of type I error is set rather high (for example, 0.1), and, therefore, most of the selected markers are false positives. In the second step, the markers selected in the first step are tested by using samples obtained from a different set of case-control samples. We performed extensive simulation studies to estimate both the type I error and the power of the test by changing parameters such as genotype relative risk, disease allele frequency, and sample size. If the total number of genotypings was limited, the stepwise focusing method yielded optimal conditions and was more powerful than conventional methods. PMID- 12111371 TI - Clinical variant of Tangier disease in Japan: mutation of the ABCA1 gene in hypoalphalipoproteinemia with corneal lipidosis. AB - Despite progress in molecular characterization, specific diagnoses of disorders belonging to a group of inherited hypoalphalipoproteinemias, i.e., apolipoprotein AI deficiency, lecithin-cholesterol acyltransferase deficiency, Tangier disease (TD), and familial high-density lipoprotein (HDL) deficiency, remain difficult on a purely clinical basis. Several TD patients were recently found to be homozygous for mutations in the ABCA1 gene. We have documented here a clinical variant of TD in a Japanese patient who manifested corneal lipidosis and premature coronary artery disease as well as an almost complete absence of HDL-cholesterol, by identifying a novel homozygous ABCA1 mutation (R1680W). We propose that patients with apparently isolated HDL deficiency who are found to carry ABCA1 mutations may in fact belong to a category of TD patients whose phenotypic features are only partially expressed, and that a number of hidden clinical variants of TD might exist among other HDL deficiency patients who have escaped correct clinical diagnosis. PMID- 12111373 TI - Molecular cloning and characterization of a novel human cAMP response element binding ( CREB) gene ( CREB4). AB - Cyclic adenosine monophosphate (cAMP) response element-binding (CREB) proteins are a family of mammalian transcription activators. We identified a novel human CREB gene ( CREB4) that was 1592 bp long and encoded a protein of 395 amino acid residues. The protein shared high homology to mouse CREB3 (identity 62%, similarity 72%). The expression pattern of the human CREB4 gene showed transcripts in prostate, brain, pancreas, skeletal muscle, small intestine, testis, leukocyte, and thymus, whereas in heart, lung, liver, kidney, placenta, spleen, ovary, and colon, specific bands of the transcript could not be detected. The CREB4 gene consisted of ten exons and nine introns and was mapped to chromosome 1q21.3 by means of a bioinformatics analysis. PMID- 12111372 TI - Identification of single-nucleotide polymorphisms in the human LPIN1 gene. AB - Because mutations in the murine analog of human LPIN1 cause lipodystrophy in mice, LPIN1 is a candidate gene for human lipodystrophy syndromes. To identify possible disease mutations and/or common single-nucleotide polymorphisms (SNPs), we developed primer pairs to amplify the 21 exons of LPIN1. We used these primer pairs to sequence LPIN1 in lipodystrophy patients who had no mutations in known lipodystrophy genes, and also in normal control subjects. We found no rare LPIN1 coding sequence variants that were exclusive to patients with lipodystrophy. However, we found four silent SNPs, namely, +17C>T in exon 3, 935C>T in exon 5, and 1040G>A and 1079G>C in exon 6, and one nonsynonymous SNP, namely, 2211C>T (P616S) in exon 15. The findings suggest that LPIN1mutations are not commonly seen in patients with lipodystrophy who had no mutations in known disease genes. However, the identification of amplification primers and SNPs provides tools to further investigate LPIN1for association with other phenotypes. PMID- 12111374 TI - Web-based detection of genotype errors in pedigree data. AB - For linkage analysis using pedigree data, it is important to eliminate contradictions concerning Mendelian inheritance from genotypic data. Such contradictions may derive from either genotyping errors or pedigree errors. We implemented an error-checking algorithm in a World Wide Web-based program that can be used through the Internet even by computer nonspecialists. This program, named Checkfam, uses two error-checking algorithms to detect and report contradictions concerning Mendelian inheritance. With this program, users can guess the causes of genotype errors (technical problems in genotyping, pedigree misrecording, or errors in input data concerning familial relationships). The present program will be useful to researchers for checking genotypic data and for preparing correct input files for linkage analysis. PMID- 12111376 TI - Identification of CAG repeat-containing genes expressed in human brain as candidate genes for autosomal dominant spinocerebellar ataxias and other neurodegenerative diseases. AB - To obtain novel candidate genes for autosomal dominant spinocerebellar ataxia and other neurodegenerative disorders in which gene mutations remain unidentified, we screened a human fetal brain cDNA library using (CAG)(10) repeat probes. Sixteen cDNAs were isolated and mapped to chromosomes 1, 2, 3, 6, 9, 13, 15, 16, 22, and X. Although we failed to detect abnormal CAG repeat expansion within these genes in Japanese patients with inherited neurodegenerative diseases, these genes remain potential candidate genes for neurodegenerative diseases that feature anticipation. PMID- 12111375 TI - Lack of evidence for a significant association between nonsyndromic cleft lip with or without cleft palate and the retinoic acid receptor alpha gene in the Japanese population. AB - Nonsyndromic cleft lip with or without cleft palate (NSCLP) is one of the most common craniofacial malformations. Both genetic and environmental factors are thought to be involved in the pathogenesis. The retinoic acid receptor alpha ( RARA) is one of the candidate genes for the pathogenesis of NSCLP. An association between a PstI restriction fragment length polymorphism or D17S579 microsatellite marker polymorphism of the RARA gene and NSCLP was previously suggested, but no nucleotide change that may influence the gene expression or the protein sequence has been reported to date. To investigate the possible association between the RARA gene and NSCLP in Japanese patients, we performed a transmission disequilibrium test (TDT) using three microsatellite markers at the RARA locus in 48 parent-offspring trios. The allele-wise TDT did not show evidence of an association between the RARA gene and NSCLP. We also screened nucleotide changes in eight patients with family histories using exon-by-exon direct sequencing. We found a novel nucleotide change (1161C>T) that is located in exon 1 of the RARA gene in one patient whose father also had the disease. Because the 1161C>T was inherited from the patient's healthy mother, the mutation was not considered to be responsible for NSCLP. We then screened all probands for the nucleotide changes in the promoter region of the RARA gene using denaturing high-performance liquid chromatography. A novel nucleotide length polymorphism of a thymidine tract was identified in three patients. No association between this polymorphism and NSCLP was observed. Our findings suggest that the RARA gene variations do not contribute to the development of NSCLP in the Japanese population. PMID- 12111377 TI - Complete XY gonadal dysgenesis and aspects of the SRYgenotype and gonadal tumor formation. AB - XY gonadal dysgenesis can be classified as either complete or incomplete according to gonadal morphology. The disease is a sex-reversal disorder resulting from embryonic testicular regression sequences and is induced by mutations in the sex-determining region Y ( SRY) gene. The incidence of SRY mutations is thought to be approximately 20%. As the disease is characterized by a frequent complication of gonadal tumors, patients are usually advised to undergo prophylactic gonadectomy. In this study, we searched for mutations in SRY open reading frames from three patients with the complete form of XY gonadal dysgenesis, and detected missense mutations in two patients. Combined with the results of our previous study, in which SRY abnormalities were also detected in two out of three complete-type patients, the final incidence of SRY abnormalities was 67% (four of six patients), which is much higher than previously thought. The incidence of gonadal tumor formation in patients with SRY abnormalities was 50% (two of four patients), which is similar to the result of a metanalysis of patients with SRY abnormalities that revealed an incidence of 52.5%. Therefore, it is possible that the lower incidences of SRY abnormalities previously reported were caused by the inclusion of patients with the incomplete form or other sex reversal disorders. Moreover, our results suggest that clinicians should carefully examine patients with SRY abnormalities. PMID- 12111378 TI - Catalog of 605 single-nucleotide polymorphisms (SNPs) among 13 genes encoding human ATP-binding cassette transporters: ABCA4, ABCA7, ABCA8, ABCD1, ABCD3, ABCD4, ABCE1, ABCF1, ABCG1, ABCG2, ABCG4, ABCG5, and ABCG8. AB - Single-nucleotide polymorphisms (SNPs) at some gene loci are useful as markers of individual risk for adverse drug reactions or susceptibility to complex diseases. We have been focusing on identifying SNPs in and around genes encoding drug metabolizing enzymes and transporters, and have constructed several high-density SNP maps of such regions. Here we report SNPs at additional loci, specifically 13 genes belonging to the superfamily of ATP-binding cassette transporters ( ABCA4, ABCA7, ABCA8, ABCD1, ABCD3, ABCD4, ABCE1, ABCF1, ABCG1, ABCG2, ABCG4, ABCG5, and ABCG8). Sequencing a total of 416 kb of genomic DNA from 48 Japanese volunteers identified 605 SNPs among these 13 loci: 14 in 5' flanking regions, 5 in 5' untranslated regions, 37 within coding elements, 529 in introns, 8 in 3' untranslated regions, and 12 in 3' flanking regions. By comparing our data with SNPs deposited in the dbSNP database of the National Center for Biotechnology Information (US) and with published reports, we determined that 491 (81%) of the SNPs reported here were novel. We also detected 107 genetic variations of other types among the loci examined (insertion-deletions or mono- di-, or trinucleotide polymorphisms). The high-density SNP maps we constructed on the basis of these data should provide useful information for investigating associations between genetic variations and common diseases or responsiveness to drug therapy. PMID- 12111379 TI - Phylogeographic analysis of mtDNA variation in four ethnic populations from Yunnan Province: new data and a reappraisal. AB - Two sets of mitochondrial DNA (mtDNA) hypervariable segment I (HVS-I) data from four ethnic populations (Tibetan, Va, Dai, and Lahu) from Yunnan Province, China, were analyzed here by using phylogeographic methods. The results suggest that more attention should be paid to sampling methodology when addressing the genetic relationship and affinity among ethnic populations. Comparison of related data from different labs may serve as a check for the credibility of the data and will help discern the origin of the ethnic populations. Generally, Tibetan populations have more north-prevalent haplogroups (clades of the mtDNA phylogeny), while Dai and Lahu populations have high frequencies of south-prevalent haplogroups. The Vas, although autochthonous according to historical records, show signs of gene admixtures from northern and southern populations, for they harbor high frequencies of the south-prevalent haplogroup F and the north-prevalent haplogroup D as well as other northern mtDNA lineages such as M9 and G2a. The consanguineous marriage customs of the Lahu, together with possible genetic drift during this group's historical migration, left a conspicuous genetic imprint on its current gene pool. PMID- 12111380 TI - Study of MTHFR and MS polymorphisms as risk factors for NTD in the Italian population. AB - Homozygosity for the C677T mutation in the methylenetetrahydrofolate reductase ( MTHFR) gene is a risk factor for neural tube defects (NTDs) in many populations, including Italians. Another common mutation on the MTHFR gene, A1298C, has also been described as a risk mutation. Furthermore, several studies have suggested that a defective methionine synthase ( MS) enzyme could be a critical defect in folate-related NTDs. An A-to-G transition at bp 2756 on the MS gene has also been reported. In this case-control study, we studied the frequencies of these two polymorphisms in 203 Italian probands with non-syndromic NTDs: 98 mothers, 67 fathers, and 210 control individuals. Although the A1298C polymorphism is common in the Italian population (0.25), the allelic frequency was significantly higher among NTD cases and their parents. Heterozygous patients and mothers have an odds ratio (OR) of 1.98 and 2.11, respectively. The risk associated with the 1298CC genotype was higher for cases (OR = 3.67), for fathers (OR = 3.28), and, above all, for mothers (OR = 6.23). The prevalence of the A2756G polymorphism of the MS gene was determined (0.15). No increased prevalence of the mutated G allele was found in NTD families. This study shows that the MTHFRA1298C polymorphism is a genetic determinant for NTD risk in Italy. No association between the MSA2756G and NTD susceptibility was found. PMID- 12111381 TI - Double deletions and missense mutations in the first nucleotide-binding fold of the ATP-binding cassette transporter A1 ( ABCA1) gene in Japanese patients with Tangier disease. AB - Tangier disease (TD) is a rare autosomal recessive disease characterized by plasma high-density lipoprotein deficiency caused by an ATP-binding cassette transporter A1 ( ABCA1) gene mutation. We describe three different mutations in Japanese patients with TD. The first patient was homozygous for double deletions of 1221 bp between intron 12 and 14 and 19.9 kb between intron 16 and 31. The breakpoint sequence analyses suggest that it is a simultaneous event caused by double-loop formation through multiple Alu. The second patient was homozygous for a novel mutation of A3198C in exon 19, resulting in Asn935His. The third patient was homozygous for A3199G of exon 19 that leads to Asn935Ser, which is the same mutation found in German and Spanish families. Both Asn mutations involved Walker A motif of the first nucleotide-binding fold. PMID- 12111383 TI - Molecular dynamics and IR spectroscopy in investigation of phase transitions in molecular crystal 4,4'-bis(11-hydroxy-1-undecyloxy)biphenyl. AB - Molecular dynamics (MD) simulations combined with temperature-dependent IR spectroscopic measurements were used to study phase transitions in molecular crystals of the mesogenic diol 4,4'-bis(11-hydroxy-1-undecyloxy)biphenyl. DSC measurements revealed four phase transitions in this molecular crystal at approximately 327.1 K, 389.8 K, 419.1 K and 431.9 K. Analysis of the dynamic trajectories at temperatures of 300 K, 360 K, 400 K and 480 K revealed changes in conformation of the mesogenic diol molecules and consequently changes in crystal packing and crystal structure in the temperature range 300-480 K and enabled us to understand the mechanism of the phase transitions. PMID- 12111382 TI - Genomic structure and eight novel exonic polymorphisms of the human N-cadherin gene. AB - Analysis of the detailed genomic structure of human N-cadherin revealed that the 16-exon gene is more than 72 kb in length and that it consists of a mosaic of exons. Five repeated cadherin domains, a transmembrane domain, and a cytoplasmic domain are encoded by exons 4 to 13, 13 and 14, and 14 to 16, respectively. A search for molecular variants in the entire coding region in 96 Japanese individuals resulted in the identification of eight sequence polymorphisms including three CCT- or GCC-type trinucleotide repeat polymorphisms adjacent to the initiation codon and five other novel single-nucleoticle polymorphisms (SNPs) in the coding region. Three of the five SNPs accompanied an amino acid substitution: Ala118Thr, Ala826Thr, and Asn845Ser. Knowlege of the fine gene structure and eight novel polymorphisms will be useful for the genetic study of the role of N-cadherin in diseases involving cell adhesion in the brain and in cardiomyocytes. PMID- 12111384 TI - A study on the structures of the substituted (aminomethyl)lithium and (thiomethyl)lithium compounds. AB - KEYWORDS: The structures of substituted (aminomethyl)lithium and (thiomethyl)lithium compounds have been examined. Geometric parameters, charge densities, bond orders, dipole moments and heats of formation for all the members of the two series of monomers and dimers of the units LiCN(R)2 and LiCSR where R=H, CH3(Me), C6H5(Ph) have been calculated. The structures of the three complex compounds containing the same units; [[Li(CH2SMe)(THF)]X], [Li2(CH2SPh)2(THF)4] and [Li2(CH2NPh2)2(THF)3] have also been modeled. Geometry optimizations have been performed with the semiempirical PM3 method. The molecular orbital calculations have been carried out by a self-consistent field method using the restricted Hartree-Fock formalism. Comparisons have been made with the corresponding properties of methyl lithium monomer and dimer. The results show that in all of the nitrogen-containing monomers, the C-Li bonds weaken and the Li C-H(N) angles decrease due to the coordination of lithium with nitrogen. Substitution of hydrogen atoms by methyl or phenyl groups decreases the Li-N coordination. In the sulfur-containing compounds, sulfur behaves similarly to nitrogen but the changes are smaller because the 3p lone-pair orbital of sulfur is higher in energy than the 2p lone-pair of nitrogen. All the dimers of nitrogen/sulfur-containing methyl lithium derivatives form six-membered rings in which the Li-N(S) coordination is greater than the one in the corresponding monomers. Dimerization reactions have been found to be exothermic and the formation of all the dimers is favored. The results obtained for the three complex structures are comparable to the experimental results reported in the literature. PMID- 12111386 TI - Supramol--a program for structure analysis of intercalates using molecular simulations: the structure of VOPO4*C6H4O2. AB - A method of structure analysis of intercalates has been developed that uses a combination of molecular simulations with powder diffraction. The program Supramol for the determination of intercalated structures uses crystal energy minimization in conjunction with powder diffraction data. The program solves the multiple minima problem in molecular mechanics, generating initial models systematically and searching for the global energy minimum by comparing the experimental and calculated diffraction patterns. The program is compatible with the Cerius2 modeling environment. Two intercalated crystal structures solved by Supramol are presented in the present paper: vanadyl phosphate intercalated with p-benzoquinone and the high temperature phase of vanadyl phosphate intercalated with dioxane. The structure of vanadyl phosphate intercalated with p-benzochinone is tetragonal, space group I4/ m, the unit cell parameters a=6.21 A, b=6.21 A, c=20.18 A and the density is rho=2.30 g x cm(-3), Z=4. The crystal structure of vanadyl phosphate intercalated with dioxane (high temperature phase) is monoclinic, space group C2/ m, unit cell parameters are: a= b=8.94 A, c=8.22 A, alpha=gamma=90 degrees, beta=106.30 degrees, Z=4, density 2.248 g x cm(-3). PMID- 12111387 TI - Analysis of distributions of amino acids in the primary structure of tumor suppressor p53 family according to the random mechanism. AB - It is well known that the evolutionary process leads to the majority of amino acids clustering in some regions rather than being homogenously distributed along a protein. Among numerous factors affecting the evolutionary process is chance, whose impact therefore should be present in a protein primary structure. The issue of how to measure the random distribution of amino acids in a primary structure is of importance for the understanding of protein structure and functions. In this study, we use the random principle as a tool to analyze and compare the distributions of amino acids in the primary structure of the p53 protein family. The results, for example, show that the amino acids are distributed more randomly in mouse p53 and less randomly in common tree shrew p53, the distribution ranks of amino acids are relatively lower in the functional regions (about 0.5 on average) than in the whole sequences (about 1.2 on average) except for mouse p53. From the probabilistic distribution view, the composition of human p53 is relatively stable in the functional regions rather than in the whole sequence, which may suggest one of the potential effects on the mutations inducing human cancers. In general, we can use the distribution probability to present quantitatively a type of distribution of amino acids in a protein, to compare quantitatively the magnitude of clusters between different proteins and to track the effect of chance on the evolutionary process. PMID- 12111388 TI - A QSPR-approach to the estimation of the pK(HB) of six-membered nitrogen heterocycles using quantum mechanically derived descriptors. AB - Descriptors derived from semiempirical (AM1) molecular orbital calculations have been used to construct a quantitative structure-property relationship (QSPR) for the thermodynamic hydrogen-bond basicity, p K(HB), of a series of six-membered aromatic nitrogen-heterocycles. The resulting model uses four-descriptors (the Coulson charge on the nitrogen atom, the energy of the localized nitrogen lone pair orbital, the p-orbital contribution to this MO and an accessibility angle). The model gives r2(cv)=0.95 for 51 compounds with a standard deviation between calculation and experiment of 0.13 log units. PMID- 12111385 TI - Docking and molecular dynamics studies at trypanothione reductase and glutathione reductase active sites. AB - A theoretical docking study on the active sites of trypanothione reductase (TR) and glutathione reductase (GR) with the corresponding natural substrates, trypanothione disulfide (T[S]2) and glutathione disulfide (GSSG), is reported. Molecular dynamics simulations were carried out in order to check the robustness of the docking results. The energetic results are in agreement with previous experimental findings and show the crossed complexes have lower stabilization energies than the natural ones. To test DOCK3.5, four nitro furanic compounds, previously designed as potentially active anti-chagasic molecules, were docked at the GR and TR active sites with the DOCK3.5 procedure. A good correlation was found between differential inhibitory activity and relative interaction energy (affinity). The results provide a validation test for the use of DOCK3.5 in connection with the design of anti-chagasic drugs. PMID- 12111389 TI - Structural characterization of the catalytic domain of the human 5-lipoxygenase enzyme. AB - Leukotrienes are inflammatory mediators involved in several diseases. The enzyme 5-lipoxygenase initiates the synthesis of leukotrienes from arachidonic acid. Little structural information is available regarding 5-lipoxygenase. In this study, we found that the primary structure of the catalytic domain of human 5 lipoxygenase is similar to that of the rabbit 15-lipoxygenase. This similarity allowed the development of a theoretical model of the tertiary structure of the 5 lipoxygenase catalytic domain, using the resolved structure of rabbit 15 lipoxygenase as a template. This model was used in conjunction with primary and secondary structural information to investigate putative nucleotide binding sites, a MAPKAP kinase 2 phosphorylation site, and a Src homology 3 binding site on the 5-lipoxygenase protein, further. Results indicate that the putative nucleotide binding sites are spatially distinct, with one on the beta-barrel domain and the other(s) on the catalytic domain. The MAPKAP kinase 2 phosphorylation site involves a four amino acid insertion in mammalian 5 lipoxygenases that significantly alters molecular structure. This target for post translational modification is both common and unique to 5-lipoxygenases. The Src homology 3 binding site, found in all lipoxygenases, appears to lack the characteristic left-handed type II helix structure of known Src homology 3 binding sites. These results, which highlight the unique nature of the MAPKAP kinase site, underscore the utility of structural information in the analysis of protein function. PMID- 12111390 TI - Theoretical studies of biliverdin: energetics of the reduction pathways to bilirubin. AB - Geometries and energies of formation of bilirubin formed by reduction of biliverdin via three meso carbon sites, the beta, gamma and delta positions, have been calculated using semiempirical methods. It has been shown that gamma bilirubin with a ridge-tile conformation forms six intramolecular hydrogen bonds and is the most stable of the three above mentioned positions by at least 22 kcal mol(-1). Reduction pathways for beta-, gamma- and delta-bilirubin formations from biliverdin are studied in detail. The roles of loss of conjugation and hydrogen bond formations in stability of different conformers have been discussed. Gamma bilirubin was fully optimized by using ab initio methods. Fine refinements of calculated results show excellent agreement with experimental results. PMID- 12111391 TI - Density functional models of the mechanism for decarboxylation in orotidine decarboxylase. AB - The mechanism of orotidine 5-monophosphate decarboxylase (ODCase) has been modeled using hybrid Density Functional Theory (B3LYP functional). The main goal of the present study was to investigate if much larger quantum chemical models of the active site than previously used could shed new light on the mechanism. The models used include the five conserved amino acids expected to be the most important ones for catalysis. One result of this model is that a mechanism involving a direct cleavage of the C-C bond followed by a protonation of C6 by Lys93 appears unlikely, with a barrier for decarboxylation 20 kcal mol(-1) too high. Additional effects like electrostatic stress and ground-state destabilization have been estimated to have only a minor influence on the reaction barrier. The conclusion from the calculations is that the negative charge developing on the substrate during decarboxylation must be stabilized by a protonation of the carbonyl O2 of the substrate. For this mechanism, the addition of the catalytic amino acids decreases the reaction barrier by 25 kcal mol(-1), but full agreement with experimental results has still not been reached. Further modifications of this mechanism are discussed. PMID- 12111392 TI - Human betacellulin structure modeled from other members of EGF family. AB - We have modeled betacellulin (BTC) to gain insight into the structural elements that can explain its properties. The epidermal growth factor (EGF) signal transduction pathway, a significant mediator of several cell functions, is based on four closely related tyrosine kinase receptors. The ErbB receptors are transmembrane glycoproteins and signal transduction is initiated by ligand binding that induces receptor homo- or heterodimerization to form a complex containing two molecules of ligand and two molecules of receptor. The EGF family of ligands can be divided into three groups based on their ability to bind and activate distinct ErbB receptor homo- and heterodimers. Each member of the group formed by BTC, heparin binding EGF (HB-EGF) and epiregulin (EP) can interact with both the EGF receptor (EGFR) and heregulin receptors (ErbB-3 and ErbB-4), and are hence called "bispecific" ligands. BTC and EP also present the distinctive feature that they activate all possible heterodimeric ErbB receptors. BTC has been modeled with the program MODELLER, using human EGF, human transforming growth factor alpha (hTGFalpha), human HB-EGF and human heregulin one alpha (hHRG 1alpha) as templates. The structure of the model as well as that of the templates were optimized and a simulation of 100 ps was run for all. The main structural properties of the model and the templates were compared and in conclusion the hBTC conformation was closely similar to that of hTGFalpha. PMID- 12111393 TI - Calculation of molecular integrals over Slater-type orbitals using recurrence relations for overlap integrals and basic one-center Coulomb integrals. AB - The recurrence relations are established for the basic one-center Coulomb integrals over Slater-type orbitals (STOs). These formulae and the recurrence relations for basic overlap integrals are utilized for the calculation of multicenter electron-repulsion integrals. The calculations of multicenter electron-repulsion integrals are performed by the use of translation formulae for STOs obtained from the Lambda and Coulomb Sturmian exponential-type functions (ETFs). It is shown that these integrals show a faster convergence rate in the case of Coulomb Sturmian ETFs. The accuracy of the results is quite high for the quantum numbers of STOs and for the arbitrary values of internuclear distances and screening constants of atomic orbitals. PMID- 12111394 TI - Molecular modeling of poly(ethylene oxide) model cofactors; 1,3,6-tri-O-galloyl beta- d-glucose and corilagin. AB - The most stable structures of two poly(ethylene oxide) (PEO) model cofactors, beta-1-O-galloyl-3,6-( R)-hexahydroxydiphenoyl- d-glucose (corilagin) and 1,3,6 tri-O-galloyl-beta- d-glucose (TGG), are calculated using molecular modeling and PM3 semiempirical molecular orbital theories. The theoretical PM3 structures agree with interpreted structures from experimental NMR; the glucopyranose ring of corilagin has a boat and TGG a chair conformation, for which the heats of formation, torsion angles, distances, van der Waals surface, and the infrared spectra are calculated. PMID- 12111395 TI - Molecular modeling analysis: "Why is 2-hydroxypyridine soluble in water but not 3 hydroxypyridine?". AB - Molecular mechanics and semiempirical calculations using HyperChem 5 were carried out to investigate whether the results obtained can explain why 2-hydroxypyridine is far more soluble in water than 3-hydroxypyridine. The results of molecular mechanics calculations show that in solution in water the total energy of 2 hydroxypyridine in the oxo form is less than that of 3-hydroxypyridine in the zwitterionic form by 2.14 kcal x mol(-1). The difference is much greater for the AM1 optimized H-bonded molecules. The greater amount of energy released in dissolution and H-bond formation by 2-hydroxypyridine than by 3-hydroxypyridine together with a higher crystal lattice energy for the latter provide an explanation as to why 3-hydroxypyridine is much less soluble in water than 2 hydroxypyridine. When the predicted electronic spectral lines of the compounds were compared with the observed lambda(max) values, it is found that generally the results obtained using AM1 agree more closely with the experimentally observed values. PMID- 12111396 TI - Excited state conformational dynamics of semiflexibly bridged electron donor acceptor systems: a semiempirical CI-study including solvent effects. AB - Semiempirical (AM1) molecular orbital theory with configuration interaction has been used to investigate the harpooning mechanism proposed from experimental studies on semiflexibly bridged electron donor-acceptor systems. Our calculations on the charge-transfer state of N-phenyl-4-(4-cyano-naphth-1-ylmethyl) piperidine, 1, confirm the proposed harpooning mechanism including an intermediate loosely folded charge-transfer state and reproduce the thermodynamics obtained from our spectroscopic studies closely. The structural details of the extended (ECT), intermediate (ICT) and compact (CCT) charge transfer states are discussed, as are the transition states connecting them. Solvent effects have been modeled using self-consistent reaction field (SCRF) calculations within the polarized continuum model. The effect of solvent polarity on the stabilities of the three charge-transfer states is discussed. PMID- 12111397 TI - Structure-cytotoxicity relationships for a series of HEPT derivatives. AB - Structure-cytotoxicity relationships were studied for a series of 90 HEPT derivatives by means of multiple linear regression (MLR) and artificial neural network (ANN) techniques. The values of log(1/CC50) (CC50=cytotoxic dose of compound required to reduce the proliferation of normal uninfected MT-4 cells by 50%) of the studied compounds were correlated with the descriptors encoding the chemical structures. Using the pertinent descriptors revealed by the regression analysis, a correlation coefficient of 0.935 ( s=0.149) for the training set ( n=81) was obtained for the ANN model with a 5-6-1 configuration. The results obtained from this study indicate that the cytotoxicity of HEPT derivatives is strongly dependent on hydrophobic factors, mainly log P(R1), and dependent on the steric factors, especially SigmaMW(R3+R4). Comparison of the descriptors' contribution obtained in MLR and ANN analysis shows that the contribution of some of the descriptors to cytotoxicity may be non-linear. PMID- 12111398 TI - Comparative molecular field analysis (CoMFA) for sulfoxidation reactions in Mortierella isabellina ATCC 42613 and Helminthosporium sp. NRRL 4671. AB - Previous models for mechanisms of enzymatic sulfoxidation have been somewhat limited by a lack of knowledge of the essential features of substrate-enzyme versus product-enzyme relationships. Computerized methods for modeling ligand protein (substrate-enzyme) interactions can overcome some of these limitations. Specifically, CoMFA (comparative molecular field analysis) provided a useful general approach in which to evaluate substrate-enzyme and product-enzyme relationships. The present investigation examined the relationship between substrate and product structure in predicting enantioselective sulfoxidation reactions using CoMFA for two species of microorganisms that have been used as models for mammalian metabolism, Mortierella isabellina and Helminthosporium sp. The overall enantioselectivity observed was based on the composite stereoselectivity of sulfoxide formation, sulfone formation (from the sulfoxide), and sulfoxide reduction back to the achiral substrate (sulfide). PMID- 12111399 TI - Binding free energy of selected anticancer compounds to DNA--theoretical calculations. AB - Many studies have elucidated structures and thermodynamics of complexes formed by different ligands with DNA. However, in most cases structural and free energy binding studies were not correlated with each other because of the problem of identifying which experimental free energy of binding corresponds to which experimental DNA-ligand structure. In the present work, Poisson-Boltzmann and solvent-accessible surface area methods were used to predict unknown modes of interaction between DNA and three different ligands: mitoxantrone and two pyrimidoacridine derivatives. In parallel, experimental measurements of binding free energy for the studied complexes were performed to compare experimental and calculated values. Our studies showed that the calculated values of free energy are only close to experimental data for some models of interaction between ligands and DNA. Based on this correlation, the most likely models of DNA-ligand complexes were postulated: (i) mitoxantrone and one derivative of pyrimidoacridine, both with two charged side chains, intercalate from the minor groove of DNA and bind with both chains in this groove; (ii) pyrimidoacridine, with only one side chain, very likely does not intercalate into DNA at all. Additionally, the non-electrostatic and electrostatic parts of the calculated binding free energy for the DNA-ligands studied are discussed. PMID- 12111400 TI - Mesoscopic dynamics of colloids simulated with dissipative particle dynamics and fluid particle model. AB - We report results of numerical simulations of complex fluids, using a combination of discrete-particle methods. Our molecular modeling repertoire comprises three simulation techniques: molecular dynamics (MD), dissipative particle dynamics (DPD), and the fluid particle model (FPM). This type of model can depict multi resolution molecular structures found in complex fluids ranging from single micelle, colloidal crystals, large-scale colloidal aggregates up to the mesoscale processes of hydrodynamical instabilities in the bulk of colloidal suspensions. We can simulate different colloidal structures in which the colloidal beds are of comparable size to the solvent particles. This undertaking is accomplished with a two-level discrete particle model consisting of the MD paradigm with a Lennard Jones (L-J) type potential for defining the colloidal particle system and DPD or FPM for modeling the solvent. We observe the spontaneous emergence of spherical or rod-like micelles and their crystallization in stable hexagonal or worm-like structures, respectively. The ordered arrays obtained by using the particle model are similar to the 2D colloidal crystals observed in laboratory experiments. The micelle shape and its hydrophobic or hydrophilic character depend on the ratio between the scaling factors of the interactions between colloid-colloid to colloid-solvent. Unlike the miscellar arrays, the colloidal aggregates involve the colloid-solvent interactions prescribed by the DPD forces. Different from the assumption of equilibrium growth, the two-level particle model can display much more realistic molecular physics, which allows for the simulation of aggregation for various types of colloids and solvent liquids over a very broad range of conditions. We discuss the potential prospects of combining MD, DPD, and FPM techniques in a single three-level model. Finally, we present results from large scale simulation of the Rayleigh-Taylor instability and dispersion of colloidal slab in 2D and 3D. Electronic supplementary material to this paper can be obtained by using the Springer LINK server located at http://dx.doi.org/10.1007/s00894-001-0068-3. PMID- 12111401 TI - Involvement of apoptotic cell death in autoimmune diseases. AB - A low rate of as well as a high rate of apoptotic cell death is involved in the development of various human autoimmune diseases. In rheumatoid arthritis (RA), impaired apoptosis of rheumatoid synovial cells appears to induce hyperplasia of the synovial tissues, whereas the acceleration of apoptotic cell death of osteoblasts may contribute to periarticular bone loss in patients with RA. Humoral factors including cytokines and growth factors present in the rheumatoid synovial tissues modulate the expression of apoptosis-related molecules in the cells, which inhibits or stimulates the apoptotic process of synovial cells and osteoblasts. In addition, investigations of animal arthritis models suggest that an enforced induction of apoptotic cell death of synovial cells ameliorates synovial tissue hyperplasia. The increase of salivary gland cell apoptosis and the resistance of apoptotic cell death in salivary infiltrating mononuclear cells have been observed in patients with Sjogren's syndrome (SS). Immunohistochemical studies indicate that X chromosome linked inhibitor of apoptosis protein in salivary gland cells as well as Bcl-2/Bcl-xL in salivary infiltrating mononuclear cells may be critical anti-apoptogenic molecules in each cell type. Human T lymphotropic virus type I (HTL V-I) is one of the pathogenic organisms for RA and SS, and we demonstrated that HTL V-I tax stimulates NF-kappa B nuclear translocation, inhibiting apoptotic cell death of human host cells, which may accelerate the autoimmune process. The association between the apoptosis of thyrocytes and the process of autoimmune thyroid diseases has also been examined, and our data suggest that Fas-mediated apoptosis of human thyrocytes is modulated by thyroid-stimulating antibodies, thyroid stimulation blocking antibodies, and cytokines. These data indicate that the correction of apoptotic cell death in each cell type will become a new therapeutic strategy for treatment of human autoimmune diseases. PMID- 12111402 TI - Mutation of beta-catenin gene in endometrial cancer but not in associated hyperplasia. AB - Endometrial hyperplasia is a recognized effect of excessive or unopposed estrogen stimulation and is considered to be a precancerous condition of endometrial adenocarcinoma. We have previously shown that the subcellular localization of beta-catenin in the human endometrium is changed according to cell proliferation, suggesting a role of intercellular transduction in cell-growth control in human endometrium, not only in the physiological condition but also in the carcinogenic endometrium. In the present study, to clarify at which stage of endometrial carcinogenesis molecular alteration of the beta-catenin gene occurs, we analyzed the subcellular localization of beta-catenin by immunohistochemistry, and we analyzed exon 3 of the beta-catenin gene, in 25 patients - with endometrial hyperplasia and 20 patients with endometrial cancers associated with endometrial hyperplasia, digesting DNA from the cancer and hyperplasia parts, separately. Fourteen of the 25 (56.0%) endometrial hyperplasia samples, 12 (60.0%) endometrial cancers, and 11 (55.0%) associated hyperplasias of the 20 endometrial cancers associated with hyperplasia showed nuclear localization of beta-catenin. Mutation in exon 3 of the beta-catenin gene was found in 2 of the 20 endometrial cancer samples; however, it was not found in the 25 endometrial hyperplasias or the 20 associated hyperplasias. The results suggest that molecular alteration of the beta-catenin gene occurs in atypical hyperplasia or cancer, rather than in simple or complex hyperplasia without atypia, during endometrial carcinogenesis. PMID- 12111403 TI - Expression of matrix metalloproteinases, tissue inhibitors of metalloproteinase, collagens, and Ki67 antigen in pleural malignant mesothelioma: an immunohistochemical and electron microscopic study. AB - Because matrix metalloproteinases (MMPs) degrade extracellular matrix, including basement membrane, and because tissue inhibitors of MMP (TIMPs) suppress MMP activities, MMPs and TIMPs are considered to play important roles in invasion and metastasis in many malignancies. We examined immunohistochemically the expression of MMPs (MMP-1, -2, -3, -7, and -9), TIMPs (TIMP-1 and -2), and collagens (types I, III, and IV) in 16 patients with pleural malignant mesothelioma (PMM; 8 with the epithelial, 4 with the sarcomatous, and 4 with the biphasic type). Electron microscopy revealed that the tumor cells in all types possessed the characteristics of malignant mesotheliomas, including numerous microvilli and moderate amounts of intermediate filaments. Basement lamina was present only focally. The proliferative Ki67 index was at a high level, compared with values reported in various other malignancies. Positive staining for MMP-1 was observed in most tumor cells in all 16 patients (100%). MMP-2 was expressed in most tumor cells in 2 patients (13%). In contrast, MMP-3, -7, and -9 were not detected in any PMM. TIMP-1 and TIMP-2 were expressed in 3 patients (19%) and 2 patients (13%), respectively. The stromal cells were simultaneously positive for MMPs or TIMPs in the patients whose tumor parenchymal cells were positive for each enzyme. These results indicate that the expression of MMP-1 and MMP-2 may be related to PMM invasion and spread. In particular, as MMP-1 was overexpressed in contrast to the lower expression of TIMP-1, MMP-1 is strongly suggested to play an important role in PMM invasion by degrading the tumor stroma. In spite of general agreement that epithelial-type PMM has a better prognosis than other types, there was no significant difference in the Ki67 index among the histological types of PMM. PMID- 12111404 TI - Abnormality of vascular elastic fibers in the macular mouse and a patient with Menkes' disease: ultrastructural and immunohistochemical study. AB - The macular mouse is a mutant mouse with the same gene abnormality as that of Menkes' disease, and it exhibits symptoms and abnormalities similar to those of Menkes' disease. In an electron microscopic study, we examined morphological changes in the internal elastic lamina (IEL) of the elastic arteries (EA) and the muscular arteries (MA) in a patient with Menkes' disease and in the macular mouse, an animal model of this disease. The IEL of the EA was significantly thinner in the macular mouse than that in controls, but the IEL of the MA in the macular mouse was significantly thicker than that of the controls. These contrary results for the thickness of the IEL in the MA and the EA in this animal model of Menkes' disease may reflect differences in the anatomical and pathophysiological properties of the two types of vessels. PMID- 12111405 TI - Morphological analysis of skin in senescence-accelerated mouse P10. AB - Senescence-accelerated mice (SAM) were established as a kind of group of related inbred strains that have been used as animal models for accelerated senescence and age-associated disorders. To analyze the characteristics of skin in SAM, the present study examined its morphology at the histological and ultrastructural levels. Histologic comparison of skin from senescence-accelerated-prone (SAM P10) and -resistant (SAM R1) mice revealed that the most characteristic features of SAM P10 were remarkable increases in the number of mast cells and in the density of collagen fibers in the dermis. Therefore, cutaneous allergic responsiveness and the proliferative activity of fibroblasts were also examined. Ultrastructurally, mast cells in the skin of SAM P10 possessed specific granules which exhibited considerable heterogeneity in electron density and various degrees of degranulation. In contrast, mast cells in the skin of control SAM R1 possessed a population of stable granules. Mast cell granules were frequently in contact with fibroblasts and were in close apposition to collagen fibers in the dermis of SAM P10. The collagen bundles were disorganized, and various diameters of collagen fibers were observed. SAM P10 demonstrated a significantly reduced wheal-and-flare reaction to histamine and tachykinins such as substance P, which suggests that skin aging may cause reduced sensitivity of mast cells and/or blood vessels to extrinsic stimuli. An in-vitro study using organ and monolayer culture demonstrated that the proliferative capacity of fibroblasts in the skin of SAM P10 was reduced in comparison with SAM R1. This is the first report that demonstrates the detailed morphological characteristics of skin in SAM P10. The findings obtained suggest that SAM P10 is a useful animal model of aged human skin, because of its many similar morphological features, including the reduction of the cutaneous allergic response, represented by neurogenic inflammation via the axon reflex, and its decreased fibroblast proliferation. PMID- 12111406 TI - Electron microscopic study of monkey retina after photodynamic treatment. AB - The purpose of this histological study was to determine the effects of photodynamic treatment, using a hematoporphyrin derivative and argon laser, on normal retinas of monkeys. Ten cynomolgus monkeys were treated with a hematoporphyrin derivative, given intravenously at a dose of 2.5 mg/kg. Forty minutes or 1 or 3 days after the injection, argon laser photoradiation was given over a 2.0-mm-diameter with a 10-min exposure and at an intensity of 40, 100, or 200 mW. The eyes were enucleated 1, 3, 4, 15, 18, 21, 35, or 38 days after the photoradiation and tissue samples were observed under a transmission electron microscope. The most fragile regions in the retina were the retinal nerve fibers, the outer segments of the visual cells, and the retinal pigment epithelium. Vascular endothelial cells were also fragile. The retinal capillary was easily obstructed, and the choriocapillaris was also occluded in an animal with severe retinal damage. The Mueller cells had the highest tolerance to the photodynamic treatment. Thus, exposing the normal part of the retina to light during photodynamic therapy should be avoided. PMID- 12111407 TI - Ultrastructure of human pheochromocytoma cells cultured for long periods. AB - We conducted ultrastructural analysis of human pheochromocytoma (PC) cells maintained in primary culture for about 10 months. The cells were first isolated by the enzymatic treatment of a surgically resected tissue specimen obtained from a 37-year-old man with PC, a condition which is characterized by elevated blood levels of adrenaline and noradrenaline. It was found that noradrenaline production in the medium continued until the 90th day of culture (1330 pg/ml). The production level decreased to 20 pg/ml on the 180th day, and to 18 pg/ml on the 300th day. Examination under a transmission electron microscope (TEM) at 4 weeks of culture revealed electron-dense granules (about 200 nm in size and, presumably, rich in catecholamines), which were also observed in the tumor cells from the original PC tissue. Neurite-like processes grew at around 1 week of culture, and were still maintained at 6 months of culture. But, after 6 months of culture, the neurite-like processes contained a rosary-like elevated structure, which was suggestive of cell degeneration, as determined by a plasma polymerization replica method and observed with a scanning electron microscope. When cells were examined under the TEM, fewer electron-dense granules were observed in the cell bodies, with more numerous lipofuscin-like granules and filaments. Thus, electron-dense granules, which, presumably, contain catecholamines, were seen in a long-term culture of human PC cells. These granules decreased in number in parallel with the decrease in catecholamine levels in the culture. PMID- 12111408 TI - Remarkable variability of apple mosaic virus capsid protein gene after nucleotide position 141. AB - Eight new sequences of European isolates from almond, apple, hop, prune and pear of the Apple mosaic ilarvirus (ApMV) capsid protein gene are presented. A consensus sequence was established as having 654 nucleotides (nt) and two American and two European isolates were identified to have insertions 6 to 15 nucleotides after nt position 141. The insertion resulted in the American isolate A inframeshift repaired with two point insertions 17 and 68 nt downstream. The RNA around the insertion point can potentially form a stable secondary structure with three hairpins. The insertions could stabilise this structure or could be neutral. The predicted folding of the translated protein is not influenced by the insertions or frameshift, and we speculate that the region after nt position 141 is without reasonable selection pressure and represents a hot spot for the accumulation of insertion mutations in ApMV. PMID- 12111409 TI - Avian paramyxoviruses and influenza viruses isolated from mallard ducks (Anas platyrhynchos) in New Zealand. AB - A comprehensive study using virological and serological approaches was carried out to determine the status of live healthy mallard ducks (Anas platyrhynchos) in New Zealand for infections with avian paramyxoviruses (APMV) and influenza viruses (AIV). Thirty-three viruses isolated from 321 tracheal and cloacal swabs were characterized as: 6 AIV (two H5N2 and four H4N6), 10 APMV-1 and 17 APMV-4. Of 335 sera samples tested for AIV antibodies, 109 (32.5%) sera were positive by nucleoprotein-blocking ELISA (NP-B-ELISA). Serum samples (315) were examined for antibody to APMV-1, -2, -3, -4, -6, -7, -8, -9 by the haemagglutination inhibition test. The largest number of reactions, with titres up to > or =1/64, was to APMV-1 (93.1%), followed by APMV-6 (85.1%), APMV-8 (56%), APMV-4 (51.7%), APMV-7 (47%), APMV-9 (15.9%), APMV-2 (13.3%) and APMV-3 (6.0%). All of the H5N2 isolates of AIV and the APMV-1 isolates from this and earlier New Zealand studies had low pathogenicity indices assessed by the Intravenous Pathogenicity Index (IVPI) with the result 0.00 and Intracerebral Pathogenicity Index (ICPI) with results 0.00-0.16. Partial genomic and antigenic analyses were also consistent with the isolates being non-pathogenic. Phylogenetic analysis of the 10 APMV-1 isolates showed 9 to be most similar to the reference APMV-1 strain D26/76 originally isolated in Japan and also to the Que/66 strain, which was isolated in Australia. The other isolate was very similar to a virus (MC 110/77) obtained from a shelduck in France. PMID- 12111411 TI - Full length genome sequence of Tioman virus, a novel paramyxovirus in the genus Rubulavirus isolated from fruit bats in Malaysia. AB - A novel paramyxovirus in the genus Rubulavirus, named Tioman virus (TiV), was isolated in 1999 from a number of pooled urine samples of Island Flying Foxes (Pteropus hypomelanus) during the search for the reservoir host of Nipah virus. TiV is antigenically related to Menangle virus (MenV) that was isolated in Australia in 1997 during disease outbreak in pigs. Sequence analysis of the full length genome indicated that TiV is a novel member of the genus Rubulavirus within the subfamily Paramyxovirinae, family Paramyxoviridae. However, there are several features of TiV which make it unique among known paramyxoviruses and rubulaviruses in particular: (1) TiV, like MenV, uses the nucleotide G as a transcriptional initiation site, rather than the A residue used by all other known paramyxoviruses; (2) TiV uses C as the +1 residue for all intergenic regions, a feature not seen for rubulaviruses but common for all other members within the subfamily Paramyxovirinae; (3) Although the attachment protein of TiV has structural features that are conserved in other rubulaviruses, it manifests no overall sequence homology with members of the genus, lacks the sialic acid binding motif N-R-K-S-C-S and has only two out of the six highly conserved residues known to be important for the catalytic activity of neuraminidase. PMID- 12111410 TI - Characterisation of an Indonesian very virulent strain of infectious bursal disease virus. AB - An Indonesian very virulent (vv) strain of infectious bursal disease virus (IBDV), designated Tasik94, was characterised both in vivo and at the molecular level. Inoculation of Tasik94 into 5-week-old specific-pathogen-free (SPF) chickens resulted in 100% morbidity and 45% mortality. The complete nucleotide and predicted amino acid sequences of genomic segments A and B were determined. Across each of the three deduced open reading frames (ORFs), Tasik94 shared the greatest nucleotide homology to Dutch vv strain D6948. Phylogenetic analyses were performed using 15 full-length polyprotein sequences and a total of 105 VP2 hypervariable region sequences from geographically and pathogenically diverse strains. In each case, Tasik94 grouped closely with vv strains, particularly those from Europe. The deduced VP1, VP2, VP3, VP4 and VP5 protein sequences of Tasik94 were aligned with those from published strains and putative virulence determinants were identified in VP2, VP3 and VP4. Alignment of additional protein sequences across the VP2 hypervariable region confirmed that residues Ile[242], Ile[256] and Ile[294] were highly-conserved amongst vv strains, and may account for their enhanced virulence. PMID- 12111412 TI - Characterization of gamma2-human herpesvirus-8 glycoproteins gH and gL. AB - The gamma2 human herpesvirus-8 (HHV-8) or Kaposi's sarcoma associated herpesvirus (KSHV) ORFs 22 and 47 are counterparts to glycoproteins gH and gL, respectively, that are conserved among the members of herpesviruses. To define HHV-8 gH and gL, rabbit polyclonal antibodies were raised against GST-gH and GST-gL fusion proteins. Anti-gL and anti-gH antibodies reacted with the surface of virus carrying BCBL-1 cells. Both antibodies immunoprecipitated the HHV-8 envelope associated 120 kDa and 41-42 kDa proteins. In transfected COS-1 cells, gH was expressed as an endo-H sensitive 110 kDa glycoprotein, which was absent on the surface of the cells. However, after co-transfection with gL, gH was detected as an endo-H resistant 120 kDa glycoprotein, and was expressed on the surface of the cells. Non-covalent complex formation between gH and gL was detected in the transfected COS-1 cells. Anti-gH and anti-gL antibodies neutralized HHV-8 infectivity in the absence of complement, individually and more efficiently together. However, virus binding to the target cells was not inhibited. These studies suggest that HHV-8 gL is required for gH processing and expression on the cell surface membranes, and gH/gL complex plays an important role in the post binding step of HHV-8 infection. PMID- 12111413 TI - Isolate-specific synergy in disease symptoms between cauliflower mosaic and turnip vein-clearing viruses. AB - Simultaneous infection of a plant by two viruses can cause more severe disease than is caused by infection with either virus alone. Such synergy may be due to effects on the replication of one virus by the second virus or to other causes. The tobamovirus turnip vein-clearing virus (TVCV), itself causing almost imperceptible symptoms in infected turnips, exacerbated symptoms of infection of turnip by the Cabbage S isolate of the caulimovirus cauliflower mosaic virus (CaMV). The synergy in symptom production was most evident in a reduced size of leaves, providing an objective measure of synergy. In contrast, synergy did not occur when the CM4-184 isolate of CaMV was used in combination with TVCV. Both isolates of CaMV increased the level of TVCV accumulated in leaves. TVCV did not increase the level of the Cabbage S CaMV isolate. The use of Cabbage S-CM4-184 chimeras revealed that a region critical for isolate synergy in stunting was within the coat protein gene and/or the 5' one third of the reverse transcriptase gene. We conclude that the disease symptom synergy between TVCV and Cabbage S CaMV is not caused by altered levels of accumulation of the viruses, but instead reflects subtle genetic interactions mapping to the ORF IV-ORF V region of CaMV DNA. PMID- 12111414 TI - Genetic diversity and response to IFN of the NS3 protease gene from clinical strains of the hepatitis C virus. AB - The N-terminal one-third of the hepatitis C virus nonstructural gene 3 (NS3) codes for a serine protease. To investigate natural genetic diversity of this enzyme a nested PCR reaction was developed to obtain NS3 protease sequence data directly from patient strains. This data was used to determine genetic diversity, phylogenetic and evolutionary rates, and selection of variants by interferon therapy. The potential effect of genetic diversity on enzyme structure using molecular modeling was also attempted. Results show significant variability in clinical HCV strains at both the nucleotide (30.2% for 1a and 25.8% for 1b) and amino acid sequences (11.0% for 1a and 9.9% for 1b). Phylogenic analysis shows two distinct clades with two HCV isolates grouping as a sister clade to 1b. Structural analysis reveals that most mutations lie in the N-terminus of the enzyme. When strains were sorted as to whether or not the patient had received antiviral therapy, no difference was found in the number or locations of mutations in 1a strains. However, 1b strains demonstrated an overall drop in the number of positions that were mutated. This study demonstrates significant differences among natural strains that may pose a problem for structure based drug development. PMID- 12111415 TI - Inhibition of vaccinia virus replication by adenosine in BSC-40 cells: involvement of A(2) receptor-mediated PKA activation. AB - In the present study, we show that adenosine (Ado) affects vaccinia virus (VV) replication in BSC-40 cells. In order to identify its effects on VV replicative cycle we analyzed the synthesis of virus macromolecules in cells incubated with 0.5 mM Ado. A 50% reduction in the steady-state level of virus DNA synthesis was observed. Consequently, virus post-replicative gene expression was also affected. A prolonged synthesis of the F11L early virus protein was also observed and it is likely related to a slow decline in the steady-state level of early mRNAs, as suggested by northern blot analysis of the VGF early transcript. The involvement of cAMP-signaling pathway as mediator of Ado response was also evaluated. Ado stimulated a three-fold increase in cAMP levels in BSC-40 cells and cAMP-mimetic agents reduced virus yield in a dose-dependent manner. Co-incubation of infected cells with H-89 reduced the inhibitory effects of 8-Br-cAMP and Ado on VV yields suggesting PKA involvement. A(2) receptor-mediated activation of PKA was indicated by antagonism of Ado response by theophylline and DMPX. Together, these results indicate that virus DNA replication is the main target of Ado. The mechanism involved is not related to reduction of the pyrimidine nucleotide synthesis. Furthermore, Ado-induced PKA activation modulates negatively an unidentified step of the virus replicative cycle. PMID- 12111417 TI - The mouse is not permissive for equine herpesvirus 2 (EHV-2), however viral DNA persisted in lung and spleen depending on the inoculation route. AB - BALB/c mice were inoculated with 3 EHV-2 low passage isolates. After intranasal inoculation, viral DNA was detected by virus-specific nested PCR in the lung up to day 30 post inoculation and in nasal turbinates till day 7. In trigeminal ganglia, olfactory bulb, brain and lymph nodes viral DNA was randomly shown by PCR. After intraperitoneal inoculation viral DNA was present in lymphoid tissues. The spleen was PCR positive up to day 30 and showed a splenomegaly. Clinical signs, virus replication and viraemia, were not observed and no virus strain specific differences were obvious. Control mice inoculated with equine herpesvirus 4 were PCR negative in all tissues. PMID- 12111416 TI - The invasion routes of neurovirulent A/Hong Kong/483/97 (H5N1) influenza virus into the central nervous system after respiratory infection in mice. AB - A/Hong Kong/483/97 (H5N1) influenza virus (HK483) isolated from the third patient during the outbreak of chicken and human influenza in Hong Kong in 1997 was shown to be neurovirulent in mice. HK483 was inoculated intranasally to mice, and the invasion routes of the virus in the central nervous system (CNS) were investigated by immunohistochemical and in situ hybridization. The pathological changes consisted of bronchopneumonia, ganglionitis, and nonpurulent encephalomyelitis of the brain stem and the anterior part of the thoracic cord. Viral antigens and viral nucleic acids (RNA and mRNA) were demonstrated in the pterygopalatine, trigeminal and superior ganglions prior to or simultaneously with their detection in the CNS. The antigens and nucleic acids were also observed in the olfactory bulb from an early stage of the infection. In the spinal cord, virus-infected cells were first demonstrated in the grey matter of the thoracic cord. The virus, which primarily replicated in the lungs, was considered to invade the thoracic cord via cardiopulmonary splanchnic nerves and sympathetic nerves. These findings indicate that the virus reached the CNS through afferent fibers of the olfactory, vagal, trigeminal, and sympathetic nerves following replication in the respiratory mucosa. PMID- 12111418 TI - Molecular epidemiology and phylogenetic analysis of Sapporo-like viruses. AB - Sapporo-like viruses (SLV) are a causative agent of gastroenteritis in humans. SLV-specific primers were newly designed in capsid protein-coding region and 529 fecal samples collected from gastroenteritis patients were tested. Thirty-five samples (6.6%) were found to be positive by reverse transcriptase-polymerase chain reaction (RT-PCR). Out of 35 positive samples, 25 were classified into a genogroup SG-I, typified by the Sapporo virus, and 9 were classified as a genogroup SG-II, typified by the London virus. Interestingly, one sample could not be classified into any genogroup, which suggests that it may be part of a new SLV genogroup. The RT-PCR used in this study appeared to be capable of widely detecting SLV genogroups, and it was seen to be powerful enough for molecular epidemiological as well as phylogenetic studies on SLV. PMID- 12111419 TI - Plaque assay for African swine fever virus on swine macrophages. AB - A plaque assay developed to detect the infection of African Swine Fever Virus on swine macrophages is described. Plaques were generated by all of the virus isolates tested. The method is suitable not only for virus titration but also for the selection of clones in protocols for isolation/purification of recombinant viruses. PMID- 12111420 TI - The nucleolus--a gateway to viral infection? AB - A number of viruses and viral proteins interact with a dynamic sub-nuclear structure called the nucleolus. The nucleolus is present during interphase in mammalian cells and is the site of ribosome biogenesis, and has been implicated in controlling regulatory processes such as the cell cycle. Viruses interact with the nucleolus and its antigens; viral proteins co-localise with factors such as nucleolin, B23 and fibrillarin, and can cause their redistribution during infection. Viruses can use these components as part of their replication process, and also use the nucleolus as a site of replication itself. Many of these properties are not restricted to any particular type of virus or replication mechanism, and examples of these processes can be found in DNA, RNA and retroviruses. Evidence suggests that viruses may target the nucleolus and its components to favour viral transcription, translation and perhaps alter the cell cycle in order to promote virus replication. Autoimmunity to nucleolin and fibrillarin have been associated with a number of diseases, and by targeting the nucleolus and displacing nucleolar antigens, virus infection might play a role in the initiation of these conditions. PMID- 12111421 TI - Characterization of simian and human immunodeficiency chimeric viruses re isolated from vaccinated macaque monkeys after challenge infection. AB - Monkeys that have been vaccinated with nef-deleted SHIVs were either fully or partially protected against challenge with acute pathogenic SHIV-89.6 P. Viruses isolated from these vaccinated monkeys were all found to be the 89.6 P challenge virus using PCR amplification and restriction enzyme analysis of the env region of the viruses. Analysis of the 3'-end of the env region and 5'-half of the nef region using a heteroduplex mobility assay revealed that the parental 89.6 P and re-isolated viruses from unvaccinated 89.6 P-infected monkeys had quite an abundant and similar heterogeneous quasispecies population. In contrast, the viruses isolated from the vaccinated monkeys had different and fewer quasispecies indicating a selective immune pressure in the vaccinated monkeys. The in vitro replication of the viruses isolated from the vaccinated monkeys in human and macaque peripheral blood mononucular cells (PBMCs) as well as in established cell lines such as M8166 and HSC-F cells, were slow and delayed when compared to the parental 89.6 P and re-isolated viruses from unvaccinated 89.6 P-infected monkeys. Further comparison revealed that in HSC-F cells the viruses from vaccinated monkeys again showed delayed and weak CD4(+) cell down-modulation as well as having little or no effect on cell growth or cell viability on HSC-F cells and monkey PBMC. Thus we noticed that these re-isolated 89.6 P viruses from the vaccinated monkeys had changed or had been selected for low pathogenic viruses in the monkeys. This suggests that though the vaccination did not completely prevent the replication of the challenge virus in the monkeys it did contain the challenge virus by suppressing the pathogenic variants. This further enhances the prospects of this nef-deleted SHIV as the bases for effective anti HIV vaccine candidates. PMID- 12111422 TI - St. Louis encephalitis virus induced pathology in cultured cells. AB - Apoptosis is a highly regulated process of cellular self-destruction with diverse functions in multicellular organisms. It is known to be one of the mechanisms of viral pathogenesis. St. Louis encephalitis virus (SLEV), an arthropod-borne flavivirus, causes encephalitis disease of varying severity mostly in North America and in some regions of South America. This virus induces cytopathic effects in vertebrate cell lines, however, the mechanism by which this occurs is yet to be elucidated. SLEV induced cytopathic effects in K562 cells, a human mononuclear cell line, and in Neuro 2a cells, a mouse neuroblastoma cell line. SLEV-infected K562 and Neuro 2a cells underwent apoptotic cell death, whereas neither the cells inoculated with UV-inactivated virus nor the mock-infected cells developed cytopathic effects. The gene expression of regulators of apoptosis was investigated in K562 cells. A rise in the expression of the pro apoptotic bax gene was detected specifically in the SLEV-infected K562 cells. These findings suggest that up-regulation of bax mRNA is correlated with cytopathic effects in SLEV-infected K562 cells. PMID- 12111424 TI - Characterization of sealpox virus, a separate member of the parapoxviruses. AB - A disease outbreak characterized by lesions of the skin and mucosa of the oral cavity was recognized in harbor seals ( Phoca vitulina) from the German North Sea. Using electron microscopy typical parapoxvirus particles were observed. The presence of parapoxvirus was confirmed by PCR and nucleotide sequencing of part of the putative major envelope protein coding gene. Comparative sequence analysis revealed that the virus from seal is significantly different from the established parapoxvirus species Orf virus, Bovine papular stomatitis virus, Pseudocowpox virus, and Parapoxvirus of red deer in New Zealand. The results of our analysis provide evidence for inclusion of the seal parapoxvirus as member of a separate species within the genus Parapoxvirus. PMID- 12111423 TI - Infection of macaques with chimeric simian and human immunodeficiency viruses containing Env from subtype F. AB - Chimeric simian and human immunodeficiency viruses (SHIVs) are useful for investigating the pathogenicity of human immunodeficiency virus (HIV-1) and to develop an anti-HIV-1 vaccine. We attempted to construct SHIVs containing Env from various subtypes, because almost all SHIVs which have been reported so far have Env from HIV-1 that belongs to subtype B. Two infectious SHIVs containing Env from two strains of HIV-1, CMR304 and CMR306, which belong to subtype F and A, respectively, were newly obtained. These SHIVs essentially showed a coreceptor usage and a neutralization pattern that were similar to those of the parental HIV 1s. In macaque PBMC, SHIVcmr304 replicated with kinetics similar to that of prototypic SHIV-NM-3rN with HIV-1 NL432 Env, but SHIVcmr306 replicated poorly. Inoculation of four rhesus macaques with SHIVcmr304 resulted in an increase of plasma viral load in all the macaques, though viral RNA copies were 100-fold lower than that in the infection with NM-3rN. This SHIV containing Env from HIV-1 subtype F will be a valuable source for the analysis of HIV-1 subtype F and the evaluation of vaccine candidates as a genetically divergent challenge virus. PMID- 12111425 TI - Reassortment between genetically distinct Japanese and US strains of Soil-borne wheat mosaic virus: RNA1 from a Japanese strain and RNA2 from a US strain make a pseudorecombinant virus. AB - Soil-borne wheat mosaic virus (SBWMV), the type species of the genus Furovirus, has a plus-sense bipartite RNA genome. Japanese and US strains of SBWMV are genetically distantly related, despite their biologically identical properties. Here we report formation of a pseudorecombinant virus consisting of RNA1 from a Japanese strain and RNA2 from a US strain, using infectious in vitro transcripts for both strains. Full-length infectious cDNA clones for a Japanese strain were previously constructed (Yamamiya and Shirako [38]). For RNA1 of a US strain, due to instability of full-length cDNA clones in Escherichia coli cells, it was necessary to prepare a full-length template DNA for in vitro transcription by combining overlapping 5'-terminal and 3'-terminal cDNAs individually cloned in two plasmids, whereas for RNA2 a full-length cDNA clone was the template. For infectivity assays, Chenopodium quinoa, a local lesion host, and wheat, a systemic host, were used. A mixture of Japanese RNA1 transcripts and US RNA2 transcripts caused formation of local lesions on C. quinoa leaves and systemic infection to wheat plants. The nucleotide sequence of the progeny viral RNA2 was identical to that of the US RNA2. The reciprocal combination was not infectious to either host. These results confirm that the Japanese and US SBWMV are genetically distantly related strains belonging to a single species. PMID- 12111426 TI - Murray Valley encephalitis virus recombinant subviral particles protect mice from lethal challenge with virulent wild-type virus. AB - We report on the development and characterisation of a recombinant Murray Valley encephalitis virus (MVE) envelope glycoprotein expression system that results in the secretion of subviral particles (SVPs) upon transfection of the murine fibroblast (COS-7) cell line. Initially, aspects of the physical and antigenic structure of cell-associated and secreted forms of the MVE envelope glycoproteins (prM and E) are presented. We then show that BALB/c mice inoculated with SVPs purified from pcDNA(3)-prM/E-transfected COS-7 cell supernatants are protected from lethal challenge with the virulent prototype strain MVE-1-51 and that this protection correlates with the development of a neutralising humoral immune response by the host. By contrast, prior immunisation with cell-associated, recombinant MVE envelope glycoproteins did not protect mice from challenge with MVE-1-51 and this was associated with the development of antibody that was unable to neutralise virus infectivity in vitro. These studies demonstrate that SVPs derived from the in vitro expression of recombinant MVE prM and E genes are an effective candidate vaccine for the prevention of encephalitis in the mouse model. PMID- 12111427 TI - Molecular analysis of two complete rice tungro bacilliform virus genomic sequences from India. AB - The complete genomic sequences of two geographically distinct isolates of rice tungro bacilliform virus (RTBV) from India were determined. Both the sequences showed equal divergence from previously reported Southeast Asian isolates. Numerous insertions, deletions and substitutions, mostly in the intergenic regions, were found. The genome sizes were 7907 and 7934 bp respectively, 95 and 68 residues short of an infectious clone reported earlier. Between them, both the isolates showed high homology all along the genome, except for a 30-nucleotide insertion/deletion close to the 3' end of ORF III in one of them. Both the isolates indicated an unconventional start codon in ORF I, similar to the type isolate. In addition, as novel features, both the Indian isolates showed an unconventional start codon for ORF IV. Considering the low amounts of genome variability noticed in other RTBV isolates, the Indian isolates show that they have diverged sufficiently from the rest and should be considered belonging to a distinct strain. PMID- 12111428 TI - Building a mouse model hallmarking the congenital human cytomegalovirus infection in central nervous system. AB - To investigate the mechanisms that human cytomegalovirus (HCMV) can vertically transmit from the placenta of mice to infect their offspring in the central nervous system (CNS) and cause congenital anomalies, and in order to provide basic research for preparing HCMV vaccine, we have developed a new type of mouse model of HCMV congenital CNS infection. Pure strain mice were propagated after being infected with HCMV. Then the degree of infection by HCMV to offspring was determined. The experiment shows that in the infection groups the mortality of fetal mice and the fatality of neonatal mice in one week are higher than that of the control groups (P < or = 0.05). At the same time we investigated the CNS of fetus's mice whose mothers were infected by HCMV. Our results showed: 1. The virus was successfully isolated from their cerebral cortex. 2. The signal of HCMV hybridization print was found in their nervous cell through in situ hybridization. 3. Especially human herpes virus-like particles and inclusion bodies in the plasm of nerve cell were found in the tissue of their brain under the electron microscope. This new type of mouse model of HCMV inherent CNS infection will help prepare HCMV vaccine and research HCMV congenital infection in CNS. PMID- 12111429 TI - Differences between influenza virus receptors on target cells of duck and chicken. AB - H5, H7, and H9 subtype influenza viruses in land-based poultry often differ from viruses of wild aquatic birds by deletions in the stalk of the neuraminidase, by the presence of additional carbohydrates on the hemagglutinin, and by occasional changes in the receptor specificity. To test whether these differences could reflect distinctions between the virus receptors in different avian species, we compared the binding of duck, chicken and human influenza viruses to cell membranes and gangliosides from epithelial tissues of duck, chicken and African green monkey. Human viruses bound to cell membranes of monkey and chicken but not to those of duck, suggesting that chicken cells unlike duck cells contain Sia(alpha2-6)Gal-terminated receptors recognized by human viruses. Duck virus bound to gangliosides with short sugar chains that were abundant in duck intestine. Human and chicken viruses did not bind to these gangliosides and bound more strongly than duck virus to gangliosides with long sugar chains that were found in chicken intestinal and monkey lung tissues. Our data suggest that the spectrum of sialylglycoconjugates which can serve as influenza virus receptors in chicken is more similar to the spectrum of receptors in the respiratory epithelia of monkey than to that in the epithelial tissues of duck. This notion could explain the recent emergence of avian H9N2 virus lineage with human virus-like receptor specificity and emphasizes the role of the chicken as a potential intermediate host for the transmission of viruses from aquatic birds to humans. PMID- 12111430 TI - Prevalence of swine influenza virus subtypes on swine farms in the United States. AB - Serologic and virologic prevalence of infection with different swine influenza virus (SIV) subtypes was investigated using swine sera, nasal swabs and lung samples that had been submitted for a diagnosis to the Minnesota Veterinary Diagnostic Laboratory. A total of 111,418 pig sera were tested for SIV antibody between 1998 and 2000, and 25,348 sera (22.8%) were found to be positive by the hemagglutination inhibition (HI) test. Of the positive samples, 16,807 (66.7%) and 8,541 (33.7%) had antibody to H1 and H3 subtypes, respectively. Between January 1998 and May of 2001, a total of 3,561 nasal swabs or lung samples were examined for the presence of SIV, and SIV was isolated from 1,124 samples (31.7%). Of these isolates, 869 (77.3%) and 255 (22.7%) were subtyped as H1 and H3, respectively, by the HI method. For further characterization, 120 SIV isolates each from 1998 to 2001 were randomly selected from a culture collection and their hemagglutinin (HA) and neuraminidase genes examined by reverse transcription-PCR and sequencing. Of the 480 isolates, 322 (67.1%), 22 (4.6%) and 129 (26.9%) were subtyped as H1N1, H1N2 and H3N2, respectively. The remaining 7 samples (1.5%) were found to contain both H1N1 and H3N2 viruses. The SIV H1N2 subtype was isolated from 1, 8, and 13 samples in 1999, 2000, and 2001, respectively. The 22 H1N2 isolates originated from 9 different states of the United States. Genetic screening of the HA genes of 12 selected H1N2 isolates showed that 8 of them had a close phylogenetic relationship with the Indiana isolate of H1N2 (A/Swine/Indiana/9K035/99), while 4 isolates were closely related to classical SIV H1N1. PMID- 12111431 TI - Expression of the Cydia pomonella granulovirus iap3 gene. AB - The IAP3 protein of Cydia pomonella granulovirus (CpGV) was the first identified member of the baculovirus IAP family of proteins, which have been shown to block apoptosis in diverse systems. However, little is known of the expression and subcellular localisation of CpGV IAP3 during a viral infection. This study examined IAP3 in cells infected by CpGV and in cells infected by an Autographa californica nucleopolyhedrovirus (AcMNPV) recombinant that carried the CpGV iap3 gene. The levels of iap3 specific transcripts were monitored and production of the protein was assessed using an IAP3-specific antiserum. The data showed that iap3 is expressed during both early and late phases of infection, with a switch occurring from distal early transcription start sites to proximal late start sites. Protein levels are highest after DNA replication. IAP3 is localised exclusively in the cytoplasm. Subcellular fractionation experiments demonstrated that the protein is present in both soluble and membrane-bound cytosolic fractions. The membrane-bound fraction includes IAP3 that is associated with the mitochondria. However, the data do not support the hypothesis that release of cytochrome C from the mitochondria is involved in baculovirus-induced apoptosis. PMID- 12111432 TI - Characterisation of potyviruses from sugarcane and maize in China. AB - Sugarcane or maize leaves with mosaic virus symptoms were collected from 13 sites in China. Sequence data showed that all 8 samples from maize contained Sugarcane mosaic virus (SCMV); complete sequences were determined from 2 samples and partial sequences (the CI coding region and the 3'-part of the genome) from the others. The 5 sugarcane samples all contained a virus tentatively described as Sorghum mosaic virus (SrMV) and in three of them SCMV was also detected; 2 SrMV sequences and the 3 SCMV ones were completely determined and partial SrMV sequences were obtained from the remaining 3 samples. The features of the complete sequences of SCMV and SrMV are described for the first time. Sequence comparisons and phylogenetic analysis showed three distinct groups of Chinese SCMV sequences (sugarcane isolates from Zhejiang province, a maize isolate from Guangdong and maize isolates from other provinces). The SrMV sequences were similar to one another (> 93% identical nucleotides); they resembled published sequences in the coat protein but were less similar in the 3'-UTR. The complete sequences of SCMV and SrMV had about 70% nucleotides identical to one another and to Maize dwarf mosaic virus (MDMV). MDMV was not detected in any of the samples. PMID- 12111433 TI - Immunoelectron microscopy analysis of HCMV gpUL73 (gN) localization. AB - Human Cytomegalovirus (HCMV) UL73 encodes for a polymorphic structural glycoprotein, gpUL73(gN), conserved among herpesviruses. This study analyzed the intracellular and intraviral localization of gpUL73 by immunoelectron-microscopy comparing the reactivity of two different antibodies. We found that gN is an envelope component of the mature viral particle with at least a portion exposed at the virus surface and another at the internal side of the envelope. Furthermore, gpUL73 is also present in the matrix of dense bodies and "black holes". These results, as well as immunoblotting analysis, suggest that the two antibodies recognize different forms, fully processed or unprocessed, of gpUL73 gN. PMID- 12111434 TI - Bean common mosaic virus isolates causing different symptoms in asparagus bean in China differ greatly in the 5'-parts of their genomes. AB - Potyvirus isolates from asparagus bean ( Vigna sesquipedalis) plants in Zhejiang province, China, caused either rugose and vein banding mosaic symptoms (isolate R) or severe yellowing (isolate Y) in this host, but were otherwise similar in host range. Both isolates were completely sequenced and shown to be isolates of Bean common mosaic virus (BCMV). The complete sequences were 9992 (R) or 10062 (Y) nucleotides long and shared 91.7% identical nucleotides (93.2% identical amino acids) in their genomes and were more distantly related to the BCMV-Peanut stripe virus sequence (PStV). The isolates were much less similar to one another in the 5'-UTR and the N-terminal region of the P1 protein. In the P1, isolate Y was closer to PStV (76.1% identical amino acids) than to isolate R (64.8%). Phylogenetic analyses of the coat protein region showed that the new isolates grouped with other isolates from Vigna spp., forming the blackeye cowpea mosaic strain subgroup of BCMV with 94-98% nucleotides (96-99% amino acids) identical to one another and about 90% identity to other BCMV isolates. Other significant subgroupings amongst published BCMV isolates were detected. PMID- 12111435 TI - Production of immunogenic VP6 protein of bovine group A rotavirus in transgenic potato plants. AB - We report here the production of transgenic potato plants expressing the major capsid protein VP6 of bovine group A rotavirus (GAR). Transgenic plants under the control of a cauliflower mosaic virus 35S promoter, or a modified promoter linked to the tobacco mosaic virus 5'-untranslated sequence were positive for GAR antigens by ELISA. The expressed protein was consistent in size with VP6 of GAR by Western blot assay. The presence of the VP6 gene and its transcript was detected by PCR and RT-PCR. Adult BALB/c mice were immunized intraperitoneally with concentrated transgenic potato extracts emulsified in Freund's adjuvant. Sera collected after immunization showed the anti-VP6 response in ELISA and Western blot assay. These results suggest that the immunogenic VP6 protein expressed in plants could be useful for the preparation of diagnostic reagents. PMID- 12111436 TI - The aging brain. Changes in the neuronal insulin/insulin receptor signal transduction cascade trigger late-onset sporadic Alzheimer disease (SAD). A mini review. AB - Aging of the brain has been demonstrated to be the main risk factor for late onset sporadic AD what is in contrast to early-onset familial AD in which mutations predominate the pathology. Aging of the brain was found to be associated with a multitude of aberrancies from normal in morphological, cellular and molecular terms. Recent findings provide clear evidence that the function of the neuronal insulin/insulin receptor signal transduction cascade is of pivotal significance to maintain normal cerebral blood flow and oxidative energy metabolism, work of the endoplasmatic reticulum/Golgi apparatus and the cell cycle in terminally differentiated neurons no longer in the cell cycle. It has become evident that normal metabolism of both amyloid precursor protein and tau protein is part of interactive processes controlled by the neuronal I/IR signal transduction cascade. In normal brain aging, the function of this cascade starts to fail compared to normal resulting in adverse effects in CBF/oxidative energy metabolism, work of the endoplasmatic reticulum/Golgi apparatus and cell cycle. The aberrancies may not be drastic, but multifold and permanently existing, inclusive the metabolism of APP and tau-protein. The amount of intraneuronally formed betaA4 may increase, and tau-protein may become hyperphosphorylated. These processes as a whole may increase the vulnerability of the aging brain and may facilitate the generation of late-onset sporadic AD. PMID- 12111437 TI - The cholinergic pathology in Alzheimer's disease--discrepancies between clinical experience and pathophysiological findings. AB - The cholinergic hypothesis of Alzheimer's disease (AD) states 1. that cholinergic neurons in the basal forebrain are severely affected in the course of disease, detectable both histopathologically by a loss of neurons and neurochemically, by a loss of marker enzymes for acetylcholine synthesis and degradation, and 2. that the resulting cerebral cholinergic deficit leads to memory loss and other cognitive and non-cognitive symptoms, which are characteristic for the illness. This hypothesis was mainly based on studies, which had been conducted on brains of patients with advanced dementia. Nevertheless, it has served as the rationale for the development of drugs, i.e. acetylcholine-esterase inhibitors (AChE-I), which have shown consistent, but modest clinical efficacy against cognitive decline and behavioural symptoms of dementia for a limited period of time. These drugs are presently regarded the standard treatment of dementia in Alzheimer's disease. Now, due to a more sensitive and reliable clinical diagnosis, neurobiological investigations can be performed on early stages of disease, when the changes detected presumably are more relevant for the pathogenesis. New studies on the pathophysiology of the cholinergic system in AD suggest 1. that the cholinergic deficit occurs only late in the disease, 2. that at the earliest stages there even is an upregulation of cholinergic activity in the brain, and 3. that an increased activity of AChE may develop under therapy with AChE-I's. These data show that there is a plasticity of the central cholinergic system in AD and that the positive clinical effects of AChE-I are to be weighted against possible detrimental effects on a pathophysiological level. These data challenge the cholinergic hypothesis in its present form, e.g. should stimulate studies on the underlying process, which leads the cholinergic system to increase its activity in patients in the early stage of AD and may have clinical consequences regarding cholinergic drug therapy. PMID- 12111438 TI - Increase of transcriptional levels of egr-1 and nur77 genes due to both nicotine treatment and withdrawal in pheochromocytoma cells. AB - The influence of nicotine on the expression of egr-1 and nur77 genes by nicotine treatment and withdrawal was assessed using PC12 cells. Nicotine treatment significantly increased the amount of mRNA for egr-1 and nur77 genes at 0.5 h post-nicotine treatment in the PC12 cells. In addition, nicotine withdrawal also elevated transcriptional levels of egr-1 and nur77 genes in Northern blot analyses. Nicotine treatment (200 microM) was also found to significantly increase expressional levels of Egr-1 and Nur77 proteins at 0.5 h post-nicotine treatment. In contrast, Egr-1 and Nur77 protein levels were dramatically decreased by nicotine withdrawal. These results suggest that expressional levels of Egr-1 and Nur77 proteins in neural cells may affect the transcriptional activity of late-response genes after nicotine withdrawal. PMID- 12111439 TI - No association of the -48CT polymorphism of the presenilin 1 gene with Alzheimer disease in a late-onset sporadic population. AB - Recently, a polymorphism located in the promoter of the presenilin 1 gene was associated with early-onset Alzheimer disease (EOAD). To determine if this polymorphism is also a risk factor for late-onset Alzheimer's disease (LOAD), we analysed its potential impact in a French population of LOAD patients only. Genotype and allelic distributions of the -48CT polymorphism were similar for controls and AD patients. Our result suggests that this polymorphism may not influence the development of LOAD. Other studies need to be undertaken to confirm this association restricting the impact of this polymorphism to EOAD patients. PMID- 12111441 TI - Cerebrospinal fluid levels of thiamine in patients with Alzheimer's disease. AB - Thiamine is an essential cofactor for several important enzymes involved in brain oxidative metabolism, such as the alpha-ketoglutarate dehydrogenase complex (KGDHC), pyruvate-dehydrogenase complex (PDHC), and transketolase. Some investigators reported decreased thiamine-diphosphate levels and decreased activities of KGDHC, pyruvate-dehydrogenase complex and transketolase in the brain tissue of Alzheimer's disease (AD) patients. We measured cerebrospinal (CSF) levels of thiamine-diphosphate, thiamine-monophosphate, free thiamine, and total thiamine, using ion-pair reversed phase high performance liquid chromatography, in 33 patients with sporadic AD and 32 matched controls. The mean CSF levels of thiamine-derivatives did not differ significantly from those of controls, while the mean plasma levels of thiamine-diphosphate, free and total thiamine were significantly lower in the AD-patient group. CSF and plasma thiamine levels were not correlated with age, age at onset, duration of the disease, and scores of the MiniMental State Examination, with the exception of plasma thiamine-diphosphate with MiniMental State Examination (r = 0.41, p < 0.05) in the AD-patients group. CSF and plasma values did not predict dementia progression, assessed with the MiniMental State Examination scores. These results suggest that CSF thiamine levels are not related with the risk for and the progression of AD. PMID- 12111440 TI - The insulin gene VNTR polymorphism in Alzheimer's disease: results of a pilot study. AB - Insulin (INS) and insulin-like growth factors include different polypeptides involved in growth and development. Possibly they play a role in the pathogenesis of neurodegenerative disorders such as Alzheimer's disease (AD). A variable number of tandem repeats (VNTR) polymorphism at the human INS 5'-flanking region consisting of three distinct allele classes has been shown to influence the tissue-specific expression of INS and the insulin-like growth factor 2 (IGF-2). Since alterations in the expression of INS or IGF-2 might be relevant in AD, we investigated the association between the INS VNTR polymorphism and the risk for AD. We found no association between the INS VNTR genotype and the risk for AD (p = 0.873). However, survival analysis revealed that class III homozygotes of the INS VNTR polymorphism had an earlier initial onset in patients suffering from early AD (p = 0.002). Our preliminary results suggest, that genetically determined alterations of the INS/IGF-2 metabolism might modify the course of AD. Further studies are warranted to confirm these data in larger study samples. PMID- 12111442 TI - Magnetic resonance in the differential diagnosis of dementia. AB - Magnetic resonance became an important tool for the differential diagnosis of dementia. Magnetic resonance imaging is the preferred method to exclude treatable entities accompanied by dementing symptoms. New techniques including diffusion and perfusion magnetic resonance imaging are helpful for the differentiation between vascular dementia and degenerative disorders. Magnetic Resonance spectroscopy evolves as a tool for the diagnosis of different forms of degenerative dementia. Multimodal magnetic resonance holds promise to diagnose Alzheimer's disease at early clinical stages and to monitor the progression of the disease. PMID- 12111443 TI - Inhibition of acetyl- and butyryl-cholinesterase in the cerebrospinal fluid of patients with Alzheimer's disease by rivastigmine: correlation with cognitive benefit. AB - Cholinesterase (ChE) inhibition represents the most efficacious treatment approach for Alzheimer's disease (AD) to date. This multiple-dose study has examined the relationship between inhibition of acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) activities in the cerebrospinal fluid (CSF) and cognitive change (measured by the Computerised Neuropsychological Test Battery [CNTB]) following administration of the ChE inhibitor, rivastigmine (Exelon). In 18 patients with mild to moderate AD, CNTB scores, activities of AChE and BuChE in the CSF, and plasma BuChE activity were determined prior to treatment with rivastigmine. Doses of rivastigmine were then titrated (1 mg b.i.d./week) to final doses of 1, 2, 3, 4, 5 or 6 mg b.i.d. (n = 3 per dose). Following treatment with the target dose of rivastigmine for at least 3 days, CNTB scores were re determined. CSF samples were continuously collected together with plasma samples prior to and for 12 hours after the final dose of rivastigmine, and AChE and BuChE activities determined.AChE in CSF and BuChE in plasma were dose-dependently inhibited by rivastigmine treatment. The inhibition of BuChE in CSF was not clearly dose-dependent. A statistically significant correlation was observed between the change in CNTB summary score and inhibition of AChE activity (r = 0.56, p < 0.05) and BuChE activity (r = -0.65, p < 0.01) in CSF. Improvement in speed-, attention- and memory-related subtests of the CNTB correlated significantly with inhibition of BuChE but not AChE activity in CSF. Weak or absent correlation with change in cognitive performance was noted for inhibition of plasma BuChE. These results indicate that cognitive improvement with rivastigmine in AD is associated with central inhibition of ChEs and support a role for central BuChE in addition to AChE inhibition in modulating cholinergic function in AD. PMID- 12111445 TI - Potential neurotoxic inflammatory responses to Abeta vaccination in humans. AB - Studies in transgenic mouse models of Alzheimer's disease suggested the development of a vaccine that would induce the production of antibodies against amyloid-beta (Abeta) peptide, which in turn would stimulate microglia to phagocytose and remove senile plaques. However, some patients in the human clinical trials developed symptoms of brain inflammation, demonstrated by lymphocyte infiltration and elevated protein levels. These parameters are indicative of a breakdown of the blood-brain-barrier and entry of T-cells into the brain. Abeta-specific activated T-helper cells have the potential to amplify the existing pro-inflammatory conditions that are present in the brains of Alzheimer's disease patients. Cytotoxic T-cells might even attack the amyloid precursor protein which is present on the surface of many cells, including neurons. Before undertaking further vaccination trials there is a need to re assess the risks associated with Abeta vaccination and with the therapeutic containment of a neuroinflammatory response. These risks may not be justified in the light of recent studies which have shown the efficacy of conventional, low risk treatments in slowing the progress of AD. PMID- 12111444 TI - Effect of subchronic administration of metrifonate, rivastigmine and donepezil on brain acetylcholine in aged F344 rats. AB - The changes in extracellular acetylcholine levels were investigated by microdialysis in the cortex and hippocampus of aging rats after administration of metrifonate (80 mg/kg), rivastigmine (0.75 mg/kg), donepezil (1.5 mg/kg) or vehicle for 21 days (twice daily p.o.). Eighteen h after the last administration, cholinesterase inhibition was 85, 52 and 39% after metrifonate, rivastigmine and donepezil, respectively, and was accompanied by 988, 590 and 75% increase in cortical acetylcholine level. In the hippocampus, metrifonate and rivastigmine brought about a 169 and 108% increase in acetylcholine levels. A challenge dose of metrifonate, rivastigmine and donepezil was followed by a further increase in cortical and hippocampal acetylcholine levels. The retrograde perfusion of the M(2)-M(4) receptor antagonist AFDX-384 (10 microM) induced a 500 and 300% increase in cortical and hippocampal acetylcholine release, in control and rivastigmine-treated rats, respectively, no increase in metrifonate-treated rats, and a 210% increase in donepezil-treated rats. In conclusion, chronic treatment of aging rats with metrifonate, rivastigmine and donepezil induces a long-lasting increase in acetylcholine levels, and reveals marked differences between the three drugs. PMID- 12111446 TI - Cerebrolysin in Alzheimer's disease: a randomized, double-blind, placebo controlled trial with a neurotrophic agent. AB - Cerebrolysin (Cere) is a compound with neurotrophic activity. It has been shown to be effective in the treatment of Alzheimer's disease (AD) in earlier trials. In this multicenter, randomized, double-blind, placebo-controlled, parallel-group study, patients were injected intravenously with placebo or 30 mL Cere five days per week for four weeks. Effects on cognition and global function were evaluated with the Alzheimer Disease Assessment Scale-Cognitive Subscale (ADAS-Cog) and the Clinicians Interview-based Impression of Change with Caregiver Input scale (CIBIC+) 4, 12, 24 weeks after the beginning of the injections. 192 patients were enrolled, 95 were randomized to placebo, and 97 to Cere. At baseline, there was a significant difference between groups for age, age of onset of dementia, and the number of patients with hallucinations. At week 12 there was a significant difference on the CIBIC+ (p = 0.033) in favor of Cere. The number of CIBIC+ responders (score < or = 4), was significantly higher (p = 0.007), with 68 (76%) in the Cere group and 51 (57%) in the placebo group. Trends were noted in the Disability Assessment in Dementia scale and the Cornell Depression Scale. Adverse events were recorded in 73% of placebo and 64% of Cere patients. Most common adverse events were headaches, dizziness, weight loss and anxiety. CONCLUSIONS: Cere treatment was well tolerated and resulted in significant improvements in the global score two months after the end of active treatment. PMID- 12111447 TI - Beta-CIT SPECT demonstrates reduced availability of serotonin transporters in patients with Fatal Familial Insomnia. AB - Fatal familial insomnia (FFI) is a rare hereditary human prion disease with unique clinical features including progressive sleep impairment and autonomic dysfunction. The serotonergic system is considered to be involved in the regulation of the sleep-wake cycle. In this study we demonstrate a reduced availability of serotonin transporters of 57% and 73% respectively in a thalamus hypothalamus region of two FFI patients examined with beta-CIT SPECT as compared to age-expected control values. PMID- 12111448 TI - Initial serotonin transport into viable platelets and imipramine binding to platelet membranes. AB - Transport of serotonin into human platelets is a paradigm for neuronal reuptake to investigate putatively low neurotransmitter availability in certain psychiatric diseases. However, inconsistent results have been obtained on serotonin binding to platelet membranes at equilibrium and transport during initial phase into isolated platelets. In the present study we applied a rapid oil-centrifugation technique to study (14)C-serotonin transport for 15s into viable human platelets during the initial phase, compared to the binding of (3)H imipramine at equilibrium (60 min) to membranes isolated from platelets of the same individuals and to their blood serotonin levels. Platelets were viable for two h after the isolation procedure; concomitant with the decrease in viability transport also decreased. Initial transport into viable cells was observed for two min. Across 19 healthy individuals blood serotonin levels correlated with the halfmaximal saturation constants of binding, K(D), for imipramine but not with any other transport or binding parameters, such as V(max) or B(max). Inhibition studies with psychoactive drugs showed good correlation between transport during the initial phase and binding at equilibrium (r = 0.83). It is speculated that changes in the V(max) of transport reflect problems with isolation, pretreatment with drugs, the energy load of the cell, and polymorphism of the serotonin transporter. The latter shows a polymorphism in the 5'regulatory region with a 44 bp insertion (l, long form) or deletion (s, short form). Results by Greenberg et al. indicate that in platelets from healthy men the l-variant was associated with increased initial serotonin uptake. Thus for genotyping, we suggest to subdivide patient and control groups in addition to psychopathology also according to their peripheral biochemical lesions. PMID- 12111449 TI - Increased hippocampal DNA oxidation in serotonin transporter deficient mice. AB - The serotonin transporter (5HTT) is the molecule responsible for the high affinity reuptake of 5HT from the synaptic cleft. Mice lacking the 5HTT exhibit highly elevated extracellular concentrations of 5HT. We assessed whether the glutathione detoxification system is altered in 5HTT-deficient mice. While levels of reduced and oxidized glutathione were unchanged, glutathione metabolising enzymes showed a differential pattern of modulation. Glutathione peroxidase was reduced in frontal cortex, brainstem, and cerebellum of 5HTT-deficient mice, though not to a statistically significant extent, while a putative isoform of the detoxifying enzyme glutathione-S-transferase pi was decreased in a number of brain regions, especially in brainstem. At the level of the DNA, we found an increase of oxidative DNA adducts in the hippocampus of 5HTT-deficient mice. Given the importance of the hippocampus in learning and memory, this may be the most important neurochemical consequence of the absence of the 5HTT. PMID- 12111450 TI - Untangling the human estrogen receptor gene structure. AB - Awareness of estrogen's neuroprotective and behavioral effects is broadening rapidly and has served as an incentive to investigate estrogen signaling in central nervous system disorders. The present analysis focuses on two human nuclear estrogen receptors, ER alpha and ER beta, which have been shown to play key roles in the complex integration of estrogen's genomic and non-genomic modes of action. The corresponding genes are estimated to have diverged from an ancestral ER gene over 450 million years ago and are candidate genes for a variety of brain disorders. Recent progress in the Human Genome Project has greatly aided our understanding of the molecular blueprint and provides the means for reassessing both genes' genomic organization. Analyses of multiple alternatively spliced transcripts, large untranslated sequences and neighbouring genes reveal several novel features which suggest an increasingly versatile transcriptional machinery. We outline additional exons in the genes' 5'- and 3' untranslated regions, a new polymorphic ER alpha microsatellite and a nested gene which lend themselves to further evolutionary and functional studies. PMID- 12111451 TI - Inhibition of the neuronal isoform of nitric oxide synthase significantly attenuates 1-methyl-4-phenylpyridinium (MPP(+)) toxicity in vitro. AB - The possible protection against the toxicity of 1-methyl-4-phenylpyridinium (MPP(+)) afforded by inhibitors of nitric oxide synthase (NOS) and the antagonist of N-methyl-D-aspartate receptor function, MK-801, was studied in a brain-slice superfusion system. Significant decreases in levels of dopamine and its metabolites 3,4-dihyroxyphenylacetic acid (DOPAC) and homovanillic acid were observed following incubation of slices with 25 microM MPP(+). The activity of intracellular lactate dehydrogenase (LDH), a marker of cell viability, was also significantly decreased. These effects were attenuated by preincubation with I mM 7-nitroindazole (7NI), a selective inhibitor of the neuronal isoform of nitric oxide synthase (NOS). In contrast, the nonspecific NOS inhibitor N(omega)-nitro-L arginine, also at 1 mM, had no effect on levels of dopamine metabolites but did show a small attenuation of the levels of dopamine. 7NI alone caused some increase in levels of dopamine and a decrease in the metabolite DOPAC, which is consistent with it also acting as an inhibitor of monoamine oxidase-B. MK-801 afforded no significant protection of aminergic cells, although changes in LDH activity suggested that there may have been some protection of non-aminergic neurons affected by this, relatively high concentration of MPP(+). PMID- 12111452 TI - Methylmercury induces neurite degeneration in primary culture of mouse dopaminergic mesencephalic cells. AB - Methylmercury cation (MeHg) is an hazardous environmental pollutant with neurotoxic action. Little is known about the effects of MeHg on catecholaminergic neurons. In the present study we have used epifluorescence microscopy and confocal microscopy to investigate the alterations induced by MeHg in primary DA (dopaminergic) cells isolated from the ventral mesencephalon of CD-1 embryonic mice and cultured for six days in vitro. DA cells were identified in the multi culture by immunocytochemistry using a tyrosine-hydroxylase antibody. The morphometric analysis of DA neurons exposed to 1 microM MeHg demonstrated a striking decrease in the number of neurites, indicative of cytoskeletal alteration. In addition, DA neurons displayed cell shrinkage and a significant increase of nuclei with chromatin condensation. Based on these results it is concluded that MeHg is highly toxic to primary DA neurons. PMID- 12111453 TI - Mitochondria determine the survival and death in apoptosis by an endogenous neurotoxin, N-methyl(R)salsolinol, and neuroprotection by propargylamines. AB - In neurodegenerative disorders, such as Parkinson's disease, selective neuronal death characterizes clinical signs and symptoms. Recently apoptosis was reported to be a common type of cell death in some disorders, and well-controlled apoptotic cascade is proposed to be a target of neuroprotective therapy. In our studies to find endogenous neurotoxins as a pathogenic factor in Parkinson's disease, dopamine-derived N-methyl(R)salsolinol was found to induce apoptosis in dopamine neurons of rat models of Parkinson's disease. In human dopaminergic SH SY5Y cells, apoptosis was initiated by decline in mitochondrial membrane potential, and anti-apoptotic Bcl-2 family protein regulated apoptotic signal transduction. In addition, a series of propargylamines were found to prevent apoptosis through stabilization of mitochondrial membrane potential, which also involved Bcl-2. The role of mitochondria and the involvement of Bcl-2 in apoptosis and neuroprotection were clearly demonstrated using isolated mitochondria. These results indicate that mitochondria are the site to determine the cell death induced by neurotoxins and also the neuroprotection by anti apoptotic propargylamines. PMID- 12111454 TI - Phospholipase A2 activity in rat embryonic brain and in primary cultures of cortical neurons. AB - We investigated, with the aid of a radio-enzymatic method, the developmental changes in the activity of 85 kDa calcium-dependent cytosolic phospholipase A(2) (cPLA(2)) in pre-term embryonic rat brain and in primary cultures of cortical neurons. PLA(2) activity was highest in the brains of 18 day embryos (E18) and gradually decreased toward birth (E18 through E21). No significant differences were found regarding cPLA(2) activity in different topographies, namely hippocampus, cortex, and total brain. In primary cultures of E18/E19 cortical neurons, cPLA(2) activity was higher than the respective embryonic tissue activity, straight from early development. In vitro cPLA(2) activity peaked on the 4th day in culture, and decreased after the six days of incubation, when the cultures became mature. This result reinforces previous evidence that cPLA(2) plays an important role in the early development of the nervous system, and further suggests that it may be implicated in neurodevelopmental processes and in in vitro neuronal survival. We believe that we have produced a useful model for the tissue culture study of PLA(2) metabolism and biochemistry, which can be further addressed to the investigation of the biology of neurodegenerative and neuropsychiatric disorders. PMID- 12111455 TI - Pergolide protects dopaminergic neurons in primary culture under stress conditions. AB - Dopamine agonists are an important therapeutic strategy in the treatment of Parkinson's disease. They postpone the necessity for and reduce the required dose of L-3,4-dihydroxyphenylalanine (L-DOPA) medication thus protecting against the development of motor complications and potential oxidative stress due to L-DOPA metabolism. In primary cultures from mouse mesencephalon we show that pergolide, a preferential D(2) agonist enhanced the survival of healthy dopaminergic neurons at low concentrations of 0.001 microM. About 100 fold higher concentrations (0.1 microM) were necessary to partially reverse the toxic effects of 10 microM 1 methyl-4-phenylpyridinium (MPP(+)). Pergolide was equally effective in preventing the reduction of dopamine uptake induced by 200 microM L-DOPA. Furthermore, between 0.001-0.1 microM it also reduced lactate production thus promoting aerobic metabolism. The present findings suggest that pergolide protects dopaminergic neurons under conditions of elevated oxidative stress. PMID- 12111456 TI - Intrastriatal iron perfusion releases dopamine: an in-vivo microdialysis study. AB - We perfused iron as FeCl(3) directly into the striatum of normal rats and used the in vivo microdialysis technique to monitor striatal levels of dopamine (DA). KCl was perfused to assess the functional integrity of the DA receptors at the end of each dialysis experiment. Cu(+2) (as CuSO(4)) and Cl(-) (as NaCl) were perfused to compare the effects of Fe(+3) to that of other heavy metal and donors of Cl(-) anion. Perfusion of FeCl(3) (1 mM for 15 min) produced a 250% increase in striatal levels of DA. Perfusion of CuSO(4) (1 mM for 15 min) or NaCl (10 mM for 15 min) did not affect striatal DA levels. There was a significant increase in DA levels with KCl stimulation (56 mM for 15 min) after perfusion with FeCl(3). We conclude that iron releases DA from striatal nerve endings without the immediate destruction of the DA terminals. The implications of chronic release of dopamine as a cause of dopaminergic cell death are discussed. PMID- 12111457 TI - Synthetic neuromelanin is toxic to dopaminergic cell cultures. AB - In the present study, primary cultures of mesencephalic dopaminergic cells were exposed to synthetic dopamine neuromelanin (NM) for 48 hrs at concentrations of 0, 1, 10, 20, 50 and 100 microg NM/ml medium. Differently prepared synthetic NM with or without incorporated iron and NM oxidatively damaged by hydrogen peroxide were used. All NMs affected cellular structures e.g. as swelling of neural processes, rounding of cells, and occasional inclusion of neuromelanin particles. Cell numbers were uniformly and dose dependently reduced. Exposure to MPP(+) and ferric iron led to cytotoxic changes which could be further aggravated by oxidatively damaged NM, suggesting cytotoxicity of soluble compounds of NM in predamaged neurons. PMID- 12111458 TI - The neuromelanin of human substantia nigra and its interaction with metals. AB - Neuromelanin (NM) is a peculiar biochemical component of several neurons in the Substantia Nigra (SN), the target area of the degenerative process in Parkinson Disease (PD). SN NM has peculiarities as to its composition and an impressive capacity of chelating metals, iron in particular, but not exclusively. Gaining insights into the structural and functional characteristics of NM should help understanding the reasons of selective vulnerability of nigral neurons in many parkinsonian conditions. From the present data a protective role of NM can be postulated until the buffering capability toward heavy metals are exhausted. The overloading of NM with iron and other metals in neurons may trigger inflammatory and degenerative processes aggravating the underlying pathological condition. PMID- 12111459 TI - Effect of lazaroid pretreatment on dopamine-induced impairment of the rat nigrostriatal system. AB - Oxidative stress induced by enhanced catecholamine metabolism may subsequently cause damages to the nervous system. We used in vivo-pulse voltammetry to study an enhanced brain dopamine (metabolism) induced either by intranigral dopamine (DA) injection or reduction of cerebral blood flow. One week after intranigral injection of 10 microg DA or unilateral occlusion of one carotid the DA activity in the ipsilateral striatum was decreased as compared to the contralateral side. Three weeks after DA application and carotid clamping the DA activity was restored to normal. The significant reduction of 3,4-dihydroxyphenylacetic acid (DOPAC) after one week was attenuated by pretreatment with the lazaroid U-74389G, injected 20 min before surgery. The results are in accordance with the view that radical mechanisms play a crucial role in the impairment of the nigrostriatal system induced by oligemia. PMID- 12111460 TI - 5-Hydroxytryptophan (5-HTP) uptake and decarboxylation in the kitten brain. AB - This study reports the presence of noradrenergic (NA) neurons which are capable to take up 5-hydroxytryptophan (5-HTP) and decarboxylate it to 5 hydroxytryptamine (5-HT serotonin) in the kitten brain. After loading of 5-HTP and monoamine oxidase inhibitor (MAOI), we could demonstrate 5-HT immunoreactivity (IR) not only in hypothalamic and midbrain dopaminergic (DA) cell bodies, but also in NA ones located in the pons and medulla oblongata of the new born kitten aged from 1 to 7 days. NA cell bodies could no longer show 5-HT IR after this treatment in the kitten older than 1 month. On the other hand, 5-HT IR in the ventrolateral posterior hypothalamic (VLPH) cells was very weak at birth and became more and more intense after 15 days of age. Finally, after loading of tryptophan (TP) and MAOI, 5-HTP uptake cells mentioned above did not express 5-HT-IR in the kitten brain. PMID- 12111461 TI - Partial loss of dopaminergic neurons in the substantia nigra, ventrotegmental area and the retrorubral area -- model of the early beginning of Parkinson's symptomatology? AB - To test substances which might have protective effects on the dopaminergic system it is necessary to use models with a pathological symptomatology of the early beginning, i.e. models in which the chance exists to arrest the otherwise progressive pathological processes (see Heim et al., 2001). 6-hydroxydopamine (6 OHDA) injected unilaterally into the ventrolateral striatum of rats (6 microg dissolved in 2 microl 0.2% ascorbic acid) leads to specific stereotyped movements after subcutaneous injection of apomorphine both 3 and 13 weeks after surgery. Ten weeks after surgery decreased spontaneous motor activity could be observed. Twelve weeks after 6-hydroxydopamine injection, the animals had difficulties in performing a spatial navigation task when the submerged escape platform was moved to another position. The switching of motor programs was less pronounced. The application of tyrosine-hydroxylase-staining showed a loss of ipsilateral neurones of the substantia nigra compacta as well as of dendrites in the pars reticulata, neurones in the ventral tegmental area and in the retrorubral area ipsilaterally as well as a loss of dopaminergic fibres both ipsilaterally and contralaterally in the striatum which should belong to the contralateral acting substantia nigra afferents. The loss of the neurones and the afferents was induced by the retrograde denervation following the 6-OHDA injection within the ventrolateral striatum. The question arises whether the model used here with the partially loss of dopaminergic neurons and fibres reflects some of pathological symptoms of Parkinson's disease in the early states. PMID- 12111462 TI - Serum antioxidant capacity in neurological, psychiatric, renal diseases and cardiomyopathy. AB - The role of free radicals (FR) in the pathogenesis and in the progression of many diseases has been often discussed, but not widely investigated. However, the total antioxidant capacity in the serum seems to be of great evidence. Total antioxidant capacity was determined using oxygen absorbance capacity assay (ORAC) in serum of patients suffering from depression, schizophrenia, Alzheimer's disease (AD), anorexia nervosa, Parkinson's disease (PD), amyotrophic lateral sclerosis (ALS), Aids-encephalopathy, diabetic polyneuropathy (PNP), cardiomyopathy (CM), renal disease, and healthy individuals as controls (C). The results showed that the total antioxidant capacity in serum decreased significantly (p < 0.01) by 24, 20, 13, and 17% for anorexia nervosa, Aids encephalopathy, PNP and CM respectively. In serum of patients with renal disease significantly elevated antioxidant capacity was found. The data indicated that increased oxidative stress can be involved in the pathogenesis or in the progression of PNP and CM. Decrease of serum antioxidant capacity in patients with anorexia nervosa and Aids-encephalopathy are probably due primarily to malnutrition and secondly to insufficient antioxidant and immune system. In renal disease, the accumulation of urea in serum seems to be responsible for high antioxidant capacity. In contrast, there were no changes in PD, AD, depression syndrome and schizophrenia. PMID- 12111463 TI - Novel mitochondrial DNA mutations in Parkinson's disease. AB - Despite the recent discovery of several chromosomal gene mutations in familial Parkinson's disease (PD) the genetic background for idiopathic PD remains to be elusive. Since the discovery of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) action on dopaminergic neuronal cells and the specific decrease of mitochondrial complex I activity in substantia nigra of PD patients mitochondrial biochemistry and genetics emerged to become Pandora's box in the pathogenesis of PD. One approach was to establish the potential role of defective mitochondrial DNA (mtDNA). As complex I genes are the most vulnerable part of mtDNA we analyzed the mitochondrial MTND1 and MTND2 genes of 10 substantia nigra and 85 platelet samples from PD patients. We were uneventful to detect heteroplasmic base changes even applying techniques able to visualize mutations with low percentage of heteroplasmy but here we report novel homoplasmic base changes. These results add further evidence that there are no inherited disease specific mtDNA mutations, hence individual homoplasmic mutations or very low grade heteroplasmic mutations in the vicinity of mitochondrial metabolism and oxidative stress may contribute to selective neuronal vulnerability in PD. PMID- 12111464 TI - Parkinson's disease: changes in apoptosis-related factors suggesting possible gene therapy. AB - Specific degeneration of the nigrostriatal dopamine (DA) neurons of the substantia nigra pars compacta and the resulting loss of nerve terminals accompanied by DA deficiency in the striatum are responsible for most of the movement disturbances called parkinsonism, i.e., muscle rigidity, akinesia, and resting tremor, observed in Parkinson's disease (PD). We and other workers have found changes in the levels of cytokines, neurotrophins, and other apoptosis related factors in the nigro-striatal region of postmortem brain and/or in the cerebrospinal fluid (CSF) from PD patients, or from animal models of PD such as 1 methyl-4-phenyl-1,2,3,4-tetrahydropyridine (MPTP)-induced PD in mice or 6 hydroxydopamine (6-OHDA)-induced PD in rats. The most remarkable changes observed specifically in the nigrostriatal region were decreased levels of neurotrophins supporting DA neurons. These results indicate that the process of cell death in the nigrostriatal DA neurons in PD may be the so-called programmed cell death, i.e., apoptosis. Thus gene therapy for PD should aim both at supplementing the decreased striatal DA level by introducing the genes of DA-synthesizing enzymes into non-DA cells in the striatum and at supporting or restoring DA neurons by preventing apoptosis by introducing genes that block the process of apoptosis. PMID- 12111465 TI - Macaque animal model for HIV-induced neurological disease. AB - The pathogenesis of HIV-induced neurological disorders is still incompletely understood. Since many aspects of this disease are difficult to explore in humans, animal models are necessary to fill the gaps in our knowledge. Based on the high concordance with the human system, the SIV-infection of macaques currently provides the best animal model to study pathogenesis, therapy and prevention of HIV-infection. In this review, important features of the CNS infection in this model are outlined. Recent virological, immunological, neurophysiological and neurochemical findings obtained with this animal model are presented and key factors in the development of neurological disease are identified. PMID- 12111466 TI - Parkinsonism in HIV dementia. AB - A great number of human immunodeficiency virus (HIV)-infected patients develop a central nervous system disorder, commonly called HIV dementia or AIDS dementia complex (ADC). HIV dementia is independent of opportunistic infections and is due to the virus itself. Symptoms include psychomotor slowing, apathy and motor disorders similar to the bradykinesia and postural and gait abnormalities observed in late Parkinson's disease. Consequently, HIV has been discussed during the last few years as an additional cause for parkinsonism, and parkinsonian syndromes as manifestations of HIV dementia. Moreover, the early phase of HIV infection gains increasing interest because of studies which report subtle neurological symptoms at this stage. Accordingly, we found in SIV-infected monkeys that dopamine is reduced by 44% within as few as two months of infection, indicating that changes during early infection must be thoroughly evaluated. In this short review, we discuss alterations in the nigrostriatal dopaminergic system during early and late immunodeficiency virus infection and the common clinical and biochemical features shared by HIV dementia and Parkinson's disease. PMID- 12111467 TI - Treatment of Parkinson's with L-DOPA. The early discovery phase, and a comment on current problems. PMID- 12111468 TI - Different modes of action of catecholamine-O-methyltransferase inhibitors entacapone and tolcapone on adenylyl cyclase activity in vitro. AB - Catechol-O-methyltransferase (COMT) inhibitors such as entacapone and tolcapone are used as adjuncts to L-DOPA ( l-3,4-dihydroxyphenylalanine, levodopa) in the treatment of Parkinson's disease. Tolcapone has been reported to associate with diarrhoea, a common reason for study withdrawal. The mechanism of this adverse effect is not yet understood. Cholera toxin causes diarrhoea by permanent activation of G(s) proteins, resulting in increased adenylyl cyclase (AC) activity. The aim of this study was to examine the effects of the COMT inhibitors entacapone and tolcapone on AC activity in membranes isolated from rat striatum, a brain structure enriched with dopaminergic G-protein-coupled receptors and AC activity. This study demonstrates differential effects of tolcapone and entacapone on Gpp(NH)p/dopamine-stimulated AC activity. Entacapone enhanced the stimulatory effect of Gpp(NH)p/dopamine, whereas tolcapone attentuated this effect, suggesting that diarrhoea associated with tolcapone treatment is not caused by permanent activation of G(s) proteins. PMID- 12111469 TI - The effect of deprenyl washout in patients with long-standing Parkinson's disease. AB - Deprenyl, an irreversible MAO-B inhibitor, is known to have a symptomatic effect in de novo patients with Parkinson's disease (PD). It has, however, not been studied thoroughly in patients with advanced PD and response fluctuations. This study evaluated the effect of washout of deprenyl in patients with long-standing PD. Eleven PD patients who were on chronic treatment with deprenyl (mean age 57 +/- 8 years, mean disease duration 8.4 +/- 2.9 years), seven with response fluctuations, were enrolled in a double-blind study of a novel MAO-B inhibitor. A deprenyl washout period of one month was required prior to initiation of treatment with the trial drug. Patients were evaluated by Unified Parkinson's Disease Rating scale (UPDRS) before and one month after the washout period. Motor function was quantified by computerized tapping speed and movement time test. Results showed that neither total UPDRS scores (22 +/- 16 vs. 18 +/- 16, respectively) nor tapping speed and movement time changed significantly (4.4 +/- 0.5 vs. 4.2 +/- 0.3 Hz and 159 +/- 45 vs. 161 +/- 40 seconds; p > 0.1, respectively). However, eight patients reported various degrees of subjective deterioration, among them were the seven fluctuating patients. Two patients first began to experience response fluctuations during the washout period. It seems that deprenyl has a symptomatic effect, especially in patients with response fluctuations, and it may postpone the appearance of fluctuations in patients with PD. Attempts to discontinue treatment with deprenyl may aggravate disease symptoms. PMID- 12111470 TI - Olfactory function in idiopathic Parkinson's disease (IPD): results from cross sectional studies in IPD patients and long-term follow-up of de-novo IPD patients. AB - Olfactory loss is a prominent symptom in idiopathic Parkinson's disease (IPD). Experiment 1 re-investigated the diagnostic value of psychophysical testing in the differentiation between idiopathic Parkinson disease (IPD) from non-IPD; 50 consecutive PS patients participated. In Experiment 2 five de-novo patients received 3 olfactory tests spread over a period of approximately one year. Nineteen IPD patients were anosmic, and 18 were hyposmic. All but one patient with MSA and PSP had mild/moderate hyposmia. Normosmia was found in CBD/misdiagnosed PS/psychogenic movement disorder. In Experiment 2, one of the de novo patients was normosmic, 3 hyposmic, and 1 anosmic. Follow up investigations indicated decreased olfactory function in 3 patients while it improved in one. The normosmic patient retained olfactory abilities. This patient failed to respond to pharmacological treatment. In summary, olfactory tests differentiate IPD from non-IPD. Furthermore, tests of olfactory function may also be of interest in investigations related to treatment of PS. PMID- 12111471 TI - Alzheimer disease and cerebrovascular pathology: an update. AB - Recent epidemiological and clinico-pathologic data suggest overlaps between Alzheimer disease (AD) and cerebrovascular lesions that may magnify the effect of mild AD pathology and promote progression of cognitive decline or even may precede neuronal damage and dementia. Vascular pathology in the aging brain and in AD includes: 1. cerebral amyloid angiopathy (CAA) with an incidence of 82-98% often associated with ApoE epsilon 2 and causing a) cerebral mass hemorrhages (around 70%, mainly in the frontal and parietal lobes), b) multiple or recurrent microhemorrhages (15%), and c) ischemic (micro-)infarcts or lacunes (around 20%). The frequency of these lesions increases with the severity of CAA and shows no correlation with that of senile amyloid plaques. CAA, significantly more frequent in patients with cerebral hemorrhages or infarcts than in aged controls, is an important risk factor for cerebrovascular lesions in AD. 2. Microvascular changes with decreased density and structural abnormalities causing regional metabolic and blood-brain barrier dysfunctions with ensuing neuronal damage. In large autopsy series of demented aged subjects, around 80% show Alzheimer type pathology, 20-40% with additional, often minor vascular lesions, 7-10% "pure" vascular dementia, and 3-5% "mixed" dementia (combination of AD and vascular encephalopathy). AD cases with additional minor cerebrovascular lesions have significantly more frequent histories of hypertension or infarcts than "pure" AD patients. Vascular lesions in AD include cortical microinfarcts, subcortical lacunes, white matter lesions / leukoencephalopathy, small hemorrhages and corticosubcortical infarcts, while in mixed type dementia multiple larger or hemispheral infarcts are more frequent. Small infarcts in AD patients have no essential impact on global cognitive decline which mainly depends on the severity of Alzheimer pathology, but in early stage of AD they may influence and promote the development of dementia. Recent studies showed lower density of plaques and tangles in brains with cerebrovascular lesions, and similar severity of dementia was related to fewer AD lesions in brains with than in those without small vascular lesions. Further studies will help to elucidate the risk factors and impact of cerebrovascular lesions on the development and progression of dementia in AD. PMID- 12111472 TI - In vivo imaging of region and cell type specific neocortical neurodegeneration in Alzheimer's disease. Perspectives of MRI derived corpus callosum measurement for mapping disease progression and effects of therapy. Evidence from studies with MRI, EEG and PET. AB - Neuropathological studies in Alzheimer's disease (AD) indicate specific loss of layer III and V large pyramidal neurons in association cortex. These neurons give rise to long cortico-cortical connections, projecting through the corpus callosum, in an anterior-posterior topology. Based on these findings we hypothesized that regional corpus callosum atrophy may be a potential in vivo marker for neocortical neuronal loss in AD. To evaluate this hypothesis, we developed a method to measure cross-sectional area of the corpus callosum and of five corpus callosum subregions on midsagittal magnetic resonance imaging scans (MRI). In a subsequent series of six experimental studies using MRI, (18)FDG-PET and EEG, we investigated the relation of white matter hyperintensities (WMH) to corpus callosum size and correlated regional pattern of corpus callosum atrophy with regional cortical metabolic decline as well as intracortical coherencies. Mean total corpus callosum area was reduced significantly in AD patients compared to healthy age-matched controls, with the greatest changes in the rostrum and the splenium and relative sparing of the truncus. The regional pattern of corpus callosum atrophy was independent of WMH load and correlated significantly with pattern of regional metabolic decline measured with (18)FDG-PET, the degree of cognitive impairment and regional decline of bilateral intracortical-coherency in EEG in AD patients. We further found that hippocampus atrophy, as a marker of early allocortical degeneration, was more pronounced than total corpus callosum atrophy in mild stages of AD. Regional corpus callosum atrophy in mild disease, however, suggested early neocortical degeneration in AD. In a longitudinal study, AD patients showed significantly greater rates of corpus callosum atrophy than controls. Rates of atrophy correlated with progression of clinical dementia severity in AD. Our results indicate that regional corpus callosum atrophy in AD patients represents the loss of callosal efferent neurons in corresponding regions of the neocortex. As these neurons are a subset of cortico-cortical projecting neurons, region-specific corpus callosum atrophy may serve as a marker of progressive neocortical disconnection in AD. In combination with measurement of hippocampal atrophy, assessment of corpus callosum atrophy over time in individual patients is useful to evaluate effects on brain structure of currently developed drugs, thought to slow or modify AD progression. PMID- 12111473 TI - A short review of cognitive and functional neuroimaging studies of cholinergic drugs: implications for therapeutic potentials. AB - In the last 20 years a cholinergic dysfunction has been the major working hypothesis for the pharmacology of memory disorders. Cholinergic antagonists and lesions impair and different classes of cholinomimetics (i.e. acetylcholine precursors, cholinergic agonists and acetylcholinesterase inhibitors) enhance attention and memory in experiment animals, healthy human subjects and Alzheimer disease patients. In addition, acetylcholinesterase inhibitors improve different cognitive (i.e. visuospatial and verbal) functions in a variety of unrelated disorders such as dementia with Lewy bodies, Parkinson disease, multiple sclerosis, schizoaffective disorders, iatrogenic memory loss, traumatic brain injury, hyperactivity attention disorder and, as we recently reported, vascular dementia and mild cognitive impairment. In animals, different cholinomimetics dose-dependently increased regional cerebral metabolic rates for glucose (rCMRglc) and regional blood flow (rCBF), two indices of neuronal function, more markedly in subcortical regions (i.e. thalamus, hippocampus and visual system nuclei). In both healthy human subjects and Alzheimer disease patients acetylcholinesterase inhibitors increased rCMRglc and rCBF in subcortical and cortical brain regions at rest but attenuated rCBF increases during cognitive performances. Hence, acetylcholinesterase inhibitors may enhance cognition and rCMRglc by acting primarily on subcortical regions that are involved in attentional (i.e. thalamus) and memory (i.e. hippocampus) processes; such an effect probably is not specific for Alzheimer disease and can be beneficial in patients suffering from a wide array of neuropsychiatric disorders. PMID- 12111474 TI - Anxiolytic-like effects of acute and chronic GABA transporter inhibition in rats. AB - Acute GABA transporter inhibition can induce anxiolytic-like behaviors. The present analysis addressed whether chronic treatment (23 days via drinking water) with a GABA transporter inhibitor affects rat behavior similar to acute treatment and interferes with additional benzodiazepine-receptor agonistic treatment. Seventy-one rats divided into seven groups were acutely treated with either vehicle, diazepam (2 mg/kg), zolpidem (0.05 mg/kg), tiagabine (19 mg/kg) or chronically with tiagabine with or without acute diazepam or zolpidem. Animals were behaviorally characterized in an elevated plus-maze. None of the treatments induced changes in the activity of the animals. Acute and chronic treatment with tiagabine induced anxiolytic-like effects, similar to acute doses of diazepam. Acute diazepam did not enhance chronic tiagabine effects, whereas acute zolpidem attenuated the anxiolytic-like effects of chronic tiagabine. It is concluded that anxiolytic effects of acute GABA-uptake inhibition by tiagabine persist under chronic treatment and are sensitive to concomitant use of benzodiazepine receptor ligands. PMID- 12111475 TI - Understanding the neurotransmitter pathology of schizophrenia: selective deficits of subtypes of cortical GABAergic neurons. AB - Research aimed at understanding the neurotransmitter pathology of schizophrenia has been underway for half a century, with much emphasis on the dopamine system. Although this approach has advanced our understanding of treatment mechanisms, identification of primary dopaminergic abnormalities in the disease has been elusive. The increasing emphasis on a neuronal pathology of schizophrenia has led to the identification of abnormalities in GABAergic and glutamatergic systems; and we have identified selective deficits in GABAergic interneurons containing the calcium binding proteins parvalbumin and calbindin. Here we report further evidence for a loss of parvalbumin-immunoreactive neurons in both dorsolateral prefrontal and medial temporal cortex, indicating that these deficits are consistent with a subtle neurodevelopmental pathogenesis and hypothesizing that they may contribute to a further degenerative process in schizophrenia. PMID- 12111477 TI - Cycloid psychoses -- from clinical concepts to biological foundations. AB - The modern concept of cycloid psychoses is primarily based upon the clinical delineation of their phenotypes according to Leonhard. By settling the dilemma of Kraepelinean "atypical psychoses", their description may be considered one of the major achievements of clinical psychiatry in the last century. In particular, this had been facilitated by the work of Wernicke and Kleist. Albeit not yet generally recognized, cycloid psychoses have already stimulated great efforts of research yielding remarkable results. In this article, we elucidate the concept of cycloid psychoses and present recent findings pertaining to their putative biological foundations. Finally, future perspectives for the field of biological psychiatry are proposed fostering the heuristics of Leonhard's nosology. PMID- 12111478 TI - Exposure to prenatal infections, genetics and the risk of systematic and periodic catatonia. AB - The meaning of heterogeneity in schizophrenia and the impact of genetic and environmental factors on etiology are a matter of continuous debate in psychiatric research. Different clinical and birth history variables were investigated in a sample of 68 patients with chronic catatonic schizophrenia according to DSM III-R, classified into Leonhard's systematic schizophrenia (n = 32) and periodic catatonia (n = 36). Parental transmission of the disease was evident in 44% of the periodic catatonia cases compared to one case in systematic catatonia (3%; p = 0.0003). In systematic catatonia, 34% of the index cases were exposed to prenatal infections compared to 8% in periodic catatonia (p = 0.008). Using logistic regression analysis exposure to gestational maternal infections predicted diagnosis of systematic catatonia at p = 0.008, and parental psychosis predicted diagnosis of periodic catatonia in the index cases at p = 0.0001. The latter finding is substantiated by the recent mapping of a periodic catatonia susceptibility locus on chromosome 15q15 with evidence for autosomal dominant transmission. These findings support the hypothesis that distinct schizophrenia phenotypes are based on different etiological mechanisms. PMID- 12111476 TI - Hippocampal neurons in schizophrenia. AB - The hippocampus is crucial for normal brain function, especially for the encoding and retrieval of multimodal sensory information. Neuropsychiatric disorders such as temporal lobe epilepsy, amnesia, and the dementias are associated with structural and functional abnormalities of specific hippocampal neurons. More recently we have also found evidence for a role of the hippocampus in the pathophysiology of schizophrenia. The most consistent finding is a subtle, yet significant volume difference in schizophrenia. Here we review the cellular and molecular basis of smaller hippocampal volume in schizophrenia. In contrast to neurodegenerative disorders, total hippocampal cell number is not markedly decreased in schizophrenia. However, the intriguing finding of a selective loss of hippocampal interneurons deserves further study. Two neurotransmitter receptors, the GABAA and AMPA/kainate glutamate receptors, appear to be abnormal, whereas changes of the NMDA glutamate receptor are less robust. The expression of several genes, including those related to the GABAergic system, neurodevelopment, and synaptic function, is decreased in schizophrenia. Taken together, recent studies of hippocampal cell number, protein expression, and gene regulation point towards an abnormality of hippocampal architecture in schizophrenia. PMID- 12111479 TI - Multidimensional analysis of the concentrations of 17 substances in the CSF of schizophrenics and controls. AB - The concentrations of 17 substances were determined in the cerebrospinal fluid (CSF) of 28 paranoid schizophrenic patients and 16 controls. Results were standardized and simultaneously evaluated through Multidimensional Scaling (MDS). The full data set can be considered as a cloud of points consisting of the 44 subjects in the 17-dimensional parameter space. MDS seeks a two-dimensional representation of this 17-dimensional cloud of points, while retaining as much as possible the distances between the subjects. The two-dimensional reduction of the 17 CSF parameters correctly separated 15 of 16 controls from the schizophrenic subjects. This indicates that a biological heterogeneity between schizophrenic and nonschizophrenic subjects can be detected by the simultaneous analysis of the CSF concentrations of substances related directly or indirectly to the neuronal activity in the brain. PMID- 12111480 TI - Evolutionary conserved microsatellites in the promoter region of the 5 hydroxytryptamine receptor 2C gene (HTR2C) are not associated with bipolar disorder in females. AB - Two polymorphic dinucleotide repeats separated by a short spacer are localized in the promoter region of the serotonin receptor 2C gene ( HTR2C). One of the repeats was found to be evolutionary conserved between humans and rhesus monkeys. Although promoter-associated microsatellites have previously been shown to regulate expression of different genes, we did not find any significant influence of distinct HTR2C promoter microsatellite alleles on transcriptional efficiency as measured by luciferase activity and receptor availability as assayed by [(3)H] mesulergine binding. Furthermore, no association of specific alleles with bipolar disorder was found. These results indicate that the HTR2C promoter polymorphism does not contribute significantly to the etiopathogenesis of bipolar disorder in females. PMID- 12111481 TI - When cells become depressed: focus on neural stem cells in novel treatment strategies against depression. AB - Clinical neuroscience enters a new era in understanding the pathophysiology of depressive illness and the mode of action of antidepressant therapy. While elucidation of factors that lead to depression is still in its infancy, biochemical malfunctions appear to have well defined morphological correlations, especially in the hippocampus. Hippocampus is one of the main sites in the brain habouring neural stem cells. Cytokines and neurotrophic factors like brain derived neurotrophic factor (BDNF) play a pivotal role in neural plasticity and potentially influence growth and migration of these progenitors. Not surprisingly, antidepressant drugs interfering with neurotransmitters such as serotonin (5-HT) influence neurotrophins like BDNF, since 5-HT homeostasis is essential for brain development, neurogenesis, and neuroplasticity as well as complex behavior. In this review, the new area of neural stem cell research and the avenues of ongoing and future research sustaining the development of novel treatments for depression will be explored. PMID- 12111482 TI - Smoking only explains part of the associations between platelet monoamine oxidase activity and personality. AB - Platelet monoamine oxidase (MAO) activity has been shown to be inversely associated with personality traits such as sensation seeking, impulsiveness and extraversion. Those personality traits have also been linked to vulnerability for substance abuse, e.g. tobacco smoking and early onset or "type 2" alcoholism. Compounds in cigarette smoke have been shown to be inhibitors of MAO, which has led several authors to claim that there is no association between alcoholism, which is the most studied psychiatric condition, and platelet MAO if the effect of smoking is removed. With regard to the association between personality and platelet MAO, authors have in general been cautious. In the present paper we describe a number of results which show that there is such an association, both in clinical series if the effect of smoking is removed and in series where smoking have never taken place. A cornerstone in this regard is the significant association between platelet MAO activity and both behaviour/personality, voluntary alcohol intake and biochemical measures of CNS serotonergic activity in non-human primates. Strong evidence that the regulation of platelet MAO activity takes place on a transcriptional level with an involvement of transcription factors, likely to also regulate central monoaminergic activity, are presented. PMID- 12111483 TI - Electrophysiological measurements of anterior cingulate function. AB - Based on recent findings from various areas of brain research the anterior cingulate cortex (ACC) within the prefrontal cortex is increasingly considered as a brain region activated during tasks requiring conflict-monitoring and allocation of attention. In the present study with event-related potentials (ERPs) the question has been addressed, whether the NoGo-condition of the Continuous Performance Test is associated with enough conflict-monitoring and allocation of attention in order to activate the ACC in healthy controls. Low Resolution Electromagnetic Tomography (LORETA), a new three-dimensional source localization method, revealed significantly increased brain electrical activity during the NoGo-ERP as compared to the Go-ERP with its maximum located exactly within the ACC in four independent samples of healthy subjects. These results relate the conflict-monitoring requirements associated with inhibition of a prepared motor response (NoGo-condition) to a powerful brain electrical ACC activity. This non-invasive, easy to perform and inexpensive electrophysiological measurement, therefore, provides a new method for the assessment of ACC-function in healthy subjects. PMID- 12111484 TI - Five-year outcome of normal pressure hydrocephalus with or without a shunt: predictive value of the clinical signs, neuropsychological evaluation and infusion test. AB - BACKGROUND: Between 1993-1995, 51 patients under 75 years of age with clinical symptoms and CT-based diagnosis of normal pressure hydrocephalus were investigated prospectively in order to clarify the value of neuropsychological tests, clinical symptoms and signs and infusion test in the differential diagnosis and prediction of outcome in normal pressure hydrocephalus. METHODS: Patients had a thorough neurological examination, and neuropsychological evaluation. A 24-hour intraventricular ICP-measurement, infusion test, neurophysiological investigations and MRI study were performed, and a cortical biopsy was obtained. The ICP measurement defined the need for a shunt. All 51 patients were re-examined three and twelve months later. The final follow-up was accomplished five years postoperatively. FINDINGS: 25 of the patients needed a shunt operation. One year after a shunt placement 72% of these patients had a good recovery concerning activities of daily living, 58% benefited in their urinary incontinence and 57% walked better. During the 5 years of follow-up 8 patients with shunt and 9 without shunt had died. Positive effect of shunting remained. Only one neuropsychological test, recognition of words test, distinguishes the patients with the need for a shunt. Simple mini mental examination test was not different in those who improved. In the postoperative follow-up patients with shunt showed no change in neuropsychological tests even if they were subjectively better. The infusion test was of no value in diagnosing NPH. The 16 patients with Alzheimer's disease did worse after one year than those without pathological changes, but the mortality was not increased. INTERPRETATION: Specific neuropsychological tests are of little value in diagnosing NPH. Mini-Mental status examination was neither of value in diagnosing NPH nor in prediction of the outcome. In this study the infusion test did not improve diagnostic accuracy of NPH, but shunt placement relieves urinary incontinence and walking disability in patients with increased ICP. The patients with positive Alzheimer diagnosis on biopsy did not improve. PMID- 12111485 TI - Laboratory testing of hydrocephalus shunts -- conclusion of the U.K. Shunt evaluation programme. AB - 16 models of valves, currently in use in the U.K., have been tested long-term in the U.K. Shunt Evaluation Laboratory according to the protocol based on the new ISO 7197 standard. Valves tested were: Medtronic PS Medical: Delta Valve, Flow Control and Lumbo-Peritoneal Shunt, Heyer-Schulte Nero-Care: In-line, Low Profile and Pudenz Flushing Valve, Codman: Codman-Hakim Programmable, Hakim Precision, Accu-Flo, Holter, Uni-Shunt, and siphon-preventing device -- SiphonGuard, NMT: Orbis-Sigma Valve, Omni-Shunt and Hakim Valve, Sophysa: Sophy Programmable Valve, Radionics: Contour-Flex Valve. The majority of the valves had a non physiologically low hydrodynamic resistance (with the exception of Orbis-Sigma, PS Lumbo-Peritoneal and Heyer-Schulte In-Line). This may result in overdrainage both related to posture and during nocturnal cerebral vasogenic waves. A long distal catheter increases the resistance of these valves by 100-200%. Drainage through valves without siphon-preventing mechanism is very sensitive to body posture. This may produce grossly negative intracranial pressure after implantation. A few shunts (Delta, Low Profile and Pudenz-Flushing with Anti Siphon Devices) offer a reasonable resistance to negative outlet pressure, and hence potentially might prevent complications related to overdrainage. On the other hand, valves with siphon-preventing devices may be blocked by raised subcutaneous pressure (exception: SiphonGuard, but this device may block the drainage because of its faulty design). In most of the silicone-diaphragm valves, closing pressure varied and reached values lower than that specified by the manufacturer (exception: Heyer-Schulte Pudenz Flushing Valve). All programmable valves are susceptible to overdrainage in the upright body position. Programmed settings may be changed by external magnetic fields. Most shunts are very sensitive to the presence of small particles in the drained fluid. The behavior of a valve revealed during such testing is of immediate relevance to the surgeon and may not be adequately described in the manufacturer's product information. These results are also relevant to the assessment of shunt function in-vivo using an infusion test. PMID- 12111487 TI - Analysis of fluid in craniopharyngioma-related cysts in children: proteins, lactate and pH. AB - The purpose of this study was to evaluate the possible role of blood-brain barrier disruption in cyst formation in craniopharyngioma. Fifteen samples of cyst fluid and 14 samples of blood serum were collected from 14 patients with cystic forms of craniopharyngiomas and studied biochemically regarding total protein, albumin, immunoglobulins G and M contents, lactate and pH. Analysis of the data obtained for cyst fluids according to Felgenhauer and comparing them to those obtained for the corresponding blood sera led us to prove the hypothesis of blood-brain barrier impairment in patients with cyst formations in craniopharyngioma. We have also revealed an elevated lactate content and decreased pH in cyst fluids compared with blood sera. Thus the pathogenesis of craniopharyngiomal cyst appears to be much more akin to those described for cysts accompanying other brain tumours than it was believed earlier. PMID- 12111486 TI - Cervical cage fusion with 5 different implants: 250 cases. AB - BACKGROUND: Anterior decompression with interbody fusion is the surgical procedure of choice in cervical spondylosis. Graft harvesting complications occurring from classical fusion procedures favoured ongoing development of cage technology. To evaluate efficiency of cage fusion for surgical treatment of discogenic cervical disorders, this six-year retrospective study analyses 250 consecutive cases treated by interbody cage fusion with 5 different implants. METHODS: Indications for fusion concerned degenerative discopathies, disc herniations and selected cases of failed surgery presenting with radiculopathy (228 cases) or myelopathy (22 cases). Screwed threaded titanium cages (149 cases), impacted squared or anatomically shaped Peek cages (59 cases), and impacted titanium cages (42 cases) were used together with local graft or bone substitute. Additional plating was indicated in 16 unstable cases. FINDINGS: Excellent outcome for neck pain (96%) and radiculopathy (97%) was noted, but a less favourable one for myelopathy (60%). All cases were stabilised at 1 year. Complications leading to reoperation included cage migration and subsidence, adjacent level degeneration and stenotic myelopathy. INTERPRETATION: Cage technology simplified anterior cervical interbody fusion and proved efficient. The fact there was no graft harvesting saved operating time and hospital stay. STATEMENT: It is not the intention of the author to indicate material preference in this article. PMID- 12111488 TI - Endocrinological outcome following first time transsphenoidal surgery for GH-, ACTH-, and PRL-secreting pituitary adenomas. AB - BACKGROUND: To study remission rates and pituitary functions following transsphenoidal surgery of newly diagnosed GH-, ACTH-, and PRL-secreting pituitary adenomas. METHODS: Out of a series of 329 newly diagnosed pituitary adenomas, 131 (39.8%) were hormone (67 GH-, 27 ACTH-, 37 PRL-) secreting. PRL secreting adenomas were subjected to surgery because they failed to respond to previous medical treatment therapy. The data on secreting adenomas, regarding the results of standardised endocrinological testing, MRI findings and water metabolism disturbances, were extracted retrospectively from the pituitary data base of the hospital. The mean follow-up was 3.7 years. RESULTS: The overall remission rate for PRL-secreting adenomas (27%) was significantly lower than for GH- (71.6%) and ACTH-secreting (81.5%) ones. Remission rates correlated negatively with the magnitude of preoperative hormone excess (not in Cushing's disease), tumour size (not in prolactinoma) and invasiveness. Generally, the improvement of the adenopituitary functions was statistically significant during the first three postoperative months, and thereafter remained unchanged. Diabetes insipidus persisting for more than three months occurred with similar frequency in the three patient groups (in 9.4% of GH-, in 6.7% of ACTH-, and in 10% of PRL secreting adenomas). Tumour regrowth occurred more often in PRL-(20%) than in ACTH- (9.1%) and GH- (0%) secreting tumours. CONCLUSIONS: In GH- and ACTH secreting pituitary adenomas, remission rates were significantly higher and recurrence rates lower than in PRL-secreting adenomas, which had failed to respond to previous medical therapy. The overall postoperative adenopituitary function was improved in all patient groups. Diabetes insipidus occurred with similar frequency in all patient groups. When reporting on results of surgery for secreting pituitary adenomas, not only remission and recurrence rates, but also the results of the pituitary function should be included. PMID- 12111489 TI - Medically intractable epilepsy from insular low-grade gliomas: improvement after an extended lesionectomy. AB - OBJECTIVE: With the aim to better evaluate the role of the insula in seizures and the therapeutic implications, we analyzed the rate of insular low-grade gliomas (LGG) presenting with medically refractory epilepsy, detailed their ictal symptoms, and described the epileptological results following their surgical resection. METHODS: Eleven patients (8 men, 3 women, mean age: 35 years) harboring an insular LGG generating intractable seizures, underwent tumor removal and perilesional opercular cortex resection (lesionectomy "plus") using intraoperative functional electrical mapping, combined with ultrasonography and/or neuronavigation. RESULTS: Despite the occurrence of five postoperative deficits, the patients recovered in all cases. The epileptological results showed improvement in all cases: 9 Engel's Class I (82%), 1 Class II and 1 Class III. Ten lesionectomies were total [3] or subtotal [7], while one resection was partial (the patient in Engel's III). CONCLUSION: The high rate of pharmacologically intractable seizures associated with insular LGG, and the favorable epilepsy outcome following surgical resection of these tumors seemingly indicate that the insular cortex itself may induce chronic seizures when injured. These results suggest, taking account of the technical surgical progress allowing now to minimize the morbidity after surgery in this region, that there is another indication than the sole oncological reason for surgery in patients with diffuse insular glioma -- even if total tumor removal is not systematically possible. PMID- 12111490 TI - The complexity of trauma to the cranio-cervical junction: correlation of clinical presentation with Doppler flow velocities in the V3-segment of the vertebral arteries. AB - BACKGROUND: Trauma to the craniocervical junction (CCJ) encompasses a wide and complex spectrum of injury with an even broader range of clinical presentation, and since trauma to this area borderlines neurosurgical main areas of interest, these relatively rare injuries can be overlooked. In fact, there has been an abundance of documentation in the literature since Sir Charles Bell first reported on a case of occipital condyle fracture in 1817 in which a patient suddenly dropped dead as he turned his head to thank physicians and nurses while leaving the hospital. The death was attributed to compression of the medulla oblongata by a fractured occipital condyle. METHOD: At our department, we have been applying specialized diagnostic procedures in our clinical work-up in the acute and chronic situation of isolated injury to the CCJ as well as polytrauma with CCJ involvement in a total of 33 patients throughout a 3-year period (1997 to 2000). In this article, we present some of our experiences with these diagnostic procedures, introducing the application of V3-segment Doppler of the vertebral artery (VA), which we consider to be an effective supplementary method in a precise work-up in CCJ injury. The V3-segment of the vertebral artery, especially where it courses through its' groove behind the superior facet of the atlas is particularly vulnerable to injury. The authors measured flow velocity of the V3-segment from a point just below and roughly 2 to 3 centimeters dorsal to the mastoid process. FINDINGS: All patients were investigated for the occurrence of changes in the blood flow in the V3-segment of the VA after suffering a CCJ injury. There was an increase or decrease in the flow velocity in trauma patients with whiplash injuries as well as in patients with occipital condyle fractures and associated ligament injury. In severe injuries, some with fatal outcome, a dissection or thrombosis of the VA was found with a loss of the V3-doppler flow signal, or rather flow void. CONCLUSION: CCJ injuries in the polytrauma patient as well as in patients presenting with delayed onset of symptoms often remain unrecognised, despite the advances in medicine. CCJ injuries are potentially lethal and can be treated if detected early. The V3 method enables rapid, effective, feasible and inexpensive diagnosis in the initial work-up of CCJ injury. PMID- 12111491 TI - Surgical treatment of chronic subdural hygromas in infants and children. AB - BACKGROUND: Extra-axial chronic fluid collections are pathological conditions occurring more frequently in the paediatric age, particularly in children less than two-year-old. Although recognised for many years and repeatedly reported in the literature, some aspects of their treatment are still under debate. This review of the management of chronic subdural collections is based on our experience of 72 cases treated between January 1984 and December 2000. METHOD: Patients were divided into four groups according to aetiology. Thirty-four cases (47%) occurred following various surgical procedures; 14 cases (20%) occurred as complications of leptomeningeal infections; 13 cases (18%) were post-traumatic, whereas no obvious aetiology could be detected in the last 11 cases (15%). The collections were unilateral in 34 patients (47%) and bilateral in the other 38 (53%). FINDINGS: Thirty-five patients (48%) were treated by means of a temporary subdural external drainage (SED) (for a total of 38 SED procedures), which was maintained for a mean of 5.8 (+/-3.4) days; it was effective in 26 cases, whereas in the other 9 it was necessary to perform a subdural-peritoneostomy (SPS). Three of these 38 SEDs were complicated by infection. In one more child the external drainage was complicated by a chronic subdural haematoma. A SPS was performed in 44 cases (61%), 9 being failed external drainages. Only in 16 (38%) of these 44 patients the SPS was removed after an average of 27.0 (+/-16.6) months. Three patients (4%) were treated by craniotomy and resection of the neomembranes lining the subdural fluid collection. Obstruction of the SPS occurred in 6 children, infection in 4. Good results were obtained with either external or internal drainage. In fact post-operative neuroradiological investigation disclosed in all the cases complete or almost complete cerebral re-expansion; also the clinical outcome was generally very good, although largely dependent upon the basic clinical condition. Unilateral subdural-peritoneal and/or external drainage was effective also in case of bilateral collections. Only 2 patients required temporary bilateral external drainage. PMID- 12111492 TI - Endoscopic transthoracic sympathicotomy and peripheral microcirculation: effects of electric sympathetic chain stimulation, thermocoagulation and anaesthetic agents. AB - BACKGROUND: During endoscopic transthoracic sympathicotomy (ETS) in patients with hyperhidrosis it is useful to assess the effect of surgery peroperatively. However, the autonomic system is affected in various ways by different anesthetic agents. In the present study, the effect of ETS during either isoflurane or propofol anesthesia was evaluated by measuring changes in the finger pulp microcirculation using laser Doppler flowmetry (LDF). Electric stimulation of the sympathetic chain was used for identifying the sympathetic chain and to explore whether the anesthetic agents differentially influenced the LDF response to stimulation. METHODS: From a group of 12 patients with incapacitating palmar hyperhidrosis, six were randomly assigned to isoflurane and six to propofol anesthesia. LDF probes were attached to the ipsilateral finger pulp for continuous recording of peripheral blood flow during the ETS procedure. Electric stimulation (1 Hz, 10 Hz, and 100 Hz) was applied to the sympathetic paravertebral chain at the levels of the 2nd and 3rd sympathetic ganglia via a custom designed bipolar electrode. In eight of the patients LDF recordings were also performed in the awake state and compared with records obtained from eight healthy subjects. FINDINGS: In patients anesthetized with isoflurane, the base line finger pulp blood flow did not significantly differ from that of awake normal subjects, while those anesthetized with propofol had a lower base line flow, similar to that of awake subjects with hyperhidrosis. Stimulation of the sympathetic chain induced marked reduction of finger pulp microcirculation in both anesthetic groups, and this effect was frequency dependent during isoflurane anesthesia. After ETS a significant increase in flow was recorded only in the propofol group. Interpretation. The study demonstrates that the completeness of the sympathicotomy can only be peroperatively evaluated if an anesthetic agent with relatively low vasodilatory capacity, as e.g. propofol, is utilized. PMID- 12111493 TI - CT-guided percutaneous selective cordotomy for treatment of intractable pain in patients with malignant pleural mesothelioma. AB - Malignant mesotheliomas are neoplasms that arise from mesothelial cells and cause intractable pain in the chest wall, usually located unilaterally. This local pain can be well controlled by computerized tomography (CT)-guided percutaneous cordotomy (PC). One hundred and fifty-three patients suffering from intractable pain due to malignancy were treated with CT-guided cordotomy between 1988 and 2001. Seventy of the 153 patients had pulmonary malignancy. Among these, 40 had bronchogenic carcinoma, 11 had Pancoast tumors and the remaining 19 had mesothelioma. The latter 19 cases with malignant mesothelioma suffering from unilateral pain were treated with CT-guided PC. In 18 cases, pain was controlled totally and, in one, partial pain control was obtained. Selective pain control was obtained in 15 cases, in whom narcotic drugs were discontinued postoperatively. Post-cordotomy dysesthesia was noted in only one case, and no complication or mortality was observed. In the treatment of intractable pain, CT guided cordotomy is a perfect method in selected cases with malignancy. This is the most effective and suitable treatment modality for local pain due to malignant mesothelioma. PMID- 12111494 TI - Recurrent intracranial germinoma outside the initial radiation field with progressive malignant transformation. AB - A 19-year-old man with a pure germinoma in the pineal region was successfully treated with chemotherapy followed by 24 Gy local irradiation. Eight months later, magnetic resonance (MR) imaging detected ventricular wall dissemination outside the radiation field. Near complete response was achieved again after 28.8 Gy whole brain and 24 Gy whole spine irradiation. Two months later, MR imaging demonstrated recurrence of a mass at the corpus callosum. Gamma knife radiosurgery did not control this mass, so tumour resection was performed. Histological examination revealed immature teratoma. Enlargement of the recurrent mass at the trigone of the left lateral ventricle was found in spite of additional chemotherapy. Tumour extirpation was performed and histological examination revealed embryonal carcinoma. The patient died of tumour progression 34 months after the initial treatment. By a combination of chemotherapy regiments in use today, the initial radiation field to treat intracranial germinomas should not be confined to the tumour bed. PMID- 12111497 TI - Regrowth of residual ruptured aneurysms treated by Guglielmi's Detachable Coils which demanded further treatment by surgical clipping: report of 7 cases and review of the literature. AB - OBJECT: The management of intracranial aneurysms has truly evolved after the introduction of endovascular treatment by Guglielmi Detachable Coils (GDC). In our department, for every case (ruptured or unruptured aneurysm) we discuss in the first place endovascular treatment. When coiling is feasible, it is done as a first choice. If not (intracranial compressive haematoma, coiling unfeasible or dangerous), the patient is operated upon. Failure of the endovascular technique, like incomplete treatment and regrowth of the residual sac, becomes a subject of discussion. Some cases need complementary treatment for large or unstable residual aneurysm. METHODS: Thus, between 1997 and 2000, 59 ruptured aneurysms were treated using an endovascular method by means of GDC. In 15 of this cases complementary treatment was needed, due to the size or instability of the residual aneurysm. In 8 cases a new embolization was possible and in 7 cases a complementary surgical procedure was needed, due to the impossibility of further endovascular treatment. RESULTS: Out of these 7 cases who were operated upon after coiling, clipping of the residual neck was possible in 4 cases; in 3 cases clipping was impossible due to the partial filling of the aneurysm neck by the coils. In these 3 cases, a ligation of the residual neck, associated with coagulation of the sac was performed. DISCUSSION: The difficulty of the treatment of an residual aneurysm after coiling is discussed as well as those surgical techniques alternative to clipping (wrapping or coagulation of the residual sac). PMID- 12111498 TI - Effect of carotid endarterectomy on the ophthalmic artery. AB - BACKGROUND: To evaluate the effect of carotid endarterectomy on ophthalmic artery flow direction and peak systolic flow velocity, the authors examined the ophthalmic artery on 32 patients who had undergone carotid endarterectomy. METHODS: The 32 patients had more than 70% stenosis of the internal carotid artery at its origin on angiography. The ophthalmic artery ipsilateral to the carotid endarterectomy was evaluated by the ophthalmic artery color Doppler flow imaging before surgery and then at one week, one month, and three months after surgery. FINDINGS: (1) Before carotid endarterectomy: eight patients showed reversed ophthalmic artery direction. In the other 24 patients with antegrade ophthalmic artery flow direction, the average peak systolic flow velocity was 0.17+/-0.10 m/sec. (2) At one week after carotid endarterectomy: The reversed ophthalmic artery flow direction was resolved in each patient. The average peak systolic flow velocity in the patients with preoperative antegrade flow rose significantly to 0.28+/-0.10 m/sec (p<0.05). (3) At one month and three months after carotid endarterectomy: All patients showed the antegrade ophthalmic artery flow direction. The average peak systolic flow velocities showed no significant change compared to the value at one week after carotid endarterectomy. (4) During the followed up period, there was no patient showing worsening or recurrence of clinical symptoms including the visual symptoms. INTERPRETATION: Carotid endarterectomy brought about the correction of the reversed flow and an increase in the peak systolic flow velocity of the ipsilateral ophthalmic artery immediately after surgery. PMID- 12111499 TI - Magnetic resonance imaging after transsphenoidal surgery of clinically non functional pituitary macroadenomas and its impact on detecting residual adenoma. AB - BACKGROUND AND PURPOSE: In clinically non-functional pituitary macroadenomas, prospective follow-up magnetic resonance imaging (MRI) was conducted after transsphenoidal surgery both to study the changes of the sellar contents at the post-operative site over time and to assess the amount of residual adenoma tissue. METHODS: A total of 50 patients with clinically non-functional pituitary macroadenomas were treated by transsphenoidal tumour resection and were examined by MRI before and directly after surgery (early MR) and 3 months (intermediate MR) and 1 year after surgery (late MR). Changes in the sellar contents over time and the degree of tumour excision were studied on T1-weighted enhanced and unenhanced scans. All patients underwent complete neuro-ophthalmological and endocrinological assessments before and 3 months after surgery. For the interpretation of the post-operative images the results of the endocrinological examinations after surgery were also taken into account. RESULTS: The maximum size of tumour extension on coronal T1-weighted images ranged from 1.2 cm to 5.0 cm (mean 2.3 cm). Despite tumour resection, early post-operative images still showed a persistent mass in the sella in 83% that was usually caused by post operative haemorrhage, fluid collection and implanted fat material. However, rapid improvement in visual symptoms was noted in 89%. Changes in the sellar region at the early post-operative site markedly hindered the interpretation of MR images for detecting residual tumour tissue, which was suspected in half of the patients (1 intrasellar, 13 suprasellar, and 11 parasellar). Regression of the post-operative mass in the sella was present 3 months after surgery, resulting in a 50% change in the volume of the coronal sellar extension, which also improved the reliability in interpreting the post-operative MR images. On the intermediate MR images residual tumour tissue was detected in 30% of the patients (4 intrasellar, 2 suprasellar and 9 parasellar). Because the suprasellar mass descended over time, an increasing rate of tumour remnant within the sella was seen 3 months following surgery. Before surgery the pituitary gland was visible superiorly or posterosuperiorly to the macroadenomas in 35 patients. However, at the early post-operative site the remaining gland was only visible in 12 patients. Under the condition that endocrinological function tests confirmed adequate hormonal function, the remaining gland was detectable by MRI in 36 patients 3 months after surgery. CONCLUSION: Delayed regression of the sellar contents after transsphenoidal surgery of pituitary macroadenomas was demonstrated by this prospective MR study. Owing to the changes at the post operative site, it was difficult to interpret early post-operative images and detect residual adenoma tissue. With respect to the delayed regression of the sellar contents, the interpretation of post-operative images for detection of residual adenoma was improved 3 months after surgery. At this time, residual adenoma tissue was found in 30% of clinically non-functional macroadenomas, mostly at the parasellar and, after descent from the suprasellar space, at the intrasellar site. PMID- 12111500 TI - Relevance of image fusion for target point determination in functional neurosurgery. AB - BACKGROUND: For the treatment of medically refractory movement disorders such as Parkinson's disease, essential tremor and primary dystonia, deep brain stimulation (DBS) has become one of the main treatment options. The targets for implantation of the stimulation electrodes are various nuclei within the basal ganglia or the thalamic and subthalamic area. Accurate target localisation is of major importance for outcome and patient safety. The goal of this study was to evaluate the role of image fusion in the determination of target co-ordinates. METHOD: We conducted a retrospective study on 10 patients in whom 17 DBS electrodes had been implanted. Coordinates of the anterior and posterior commissures and of the DBS targets were compared on pre- and postoperative computerised tomography (CT) and fused CT/magnetic resonance scans. The targets as defined on the images were further compared with the targets derived intra operatively with microelectrode recordings (MER) and macrostimulation. FINDINGS: The achievable mean target accuracy was of the order of the diameter of the DBS electrode and of the accuracy of the image fusion algorithm, i.e. about 1 mm. However, the maximal differences were between 1.8 mm and 3.2 mm. INTERPRETATION: Image fusion is a helpful tool for accurate determination of target point co ordinates in DBS. In combination with intraoperative, electrophysiological recordings and stimulation which are still considered to be the most reliable localisation methods, image fusion may help to discern the anatomical and functional three-dimensionality of the target nuclei. Image fusion may reduce the number of trajectories needed for intraoperative electrophysiological determination of the optimal electrode localisation and thus lower the risk of complications. PMID- 12111501 TI - Microsurgical DREZotomy for pain of spinal cord and Cauda equina injury origin: clinical characteristics of pain and implications for surgery in a series of 26 patients. AB - The result of the DREZotomy procedure used for the treatment of chronic intractable neuropathic pain caused by injuries at the T9-L4 spine level in 26 patients has been reported. For the purpose of identifying the most favorable pain pattern for DREZ surgery we retrospectively analyzed the effectiveness of surgical treatment on different forms of pain in the follow-up period of 13-50 months, 37 months on average. All pain forms were classified according to subjective sensory pain expression including the rhythm and topography of the pain. Three groups of pain were formed according to subjective sensory equivalents: pain of thermal quality (burning, boiling, baking, warm etc.), pain of mechanical-nonthermal quality (shooting, cutting, stabbing, sharp, incisive, cramping, constriction, distraction, throbbing etc.). The third group was the combination of the previous two. Success in pain relief has been defined as a 50% or greater reduction in pain after surgery such that pain no longer interferes with patient activities of daily living and sleeping pattern and no longer requires routine analgesic pain medication. Our results revealed that the pain of mechanical-nonthermal nature and intermittent rhythm, confined to segmental topography was the most responsive to the DREZ surgical treatment so that 90% patients suffering from this pain pattern experienced a good long-term pain relief (70% had complete long term pain relief). Neuropathic pain of thermal quality with the diffuse infralesional distribution and steady rhythm was the most resistant to the DREZ surgical treatment: neither patient had long-term relief of this pain pattern. In the group of patients suffering from pain consisting of combined mechanical and thermal sensory components with confined pain territory, 75% experienced a good long-term pain relief (50% had complete long-term pain relief). Immediate pain relief was obtained in 88% of patients and was long lasting in 69% of the total series. Our results pointed to confined territory, intermittent rhythm and mechanical nature of the pain as the most relevant predictors of the expected pain relief achieved by the DREZ surgery. PMID- 12111502 TI - The evaluation and comparison of cerebellar mutism in children and adults after posterior fossa surgery: report of two adult cases and review of the literature. AB - BACKGROUND: Although there are some cases of cerebellar mutism in adults after posterior fossa surgery for cerebellar tumour it generally occurs in children. Reversible pathophsiology and the anatomical substrate of this syndrome still remain unclear. The predominance of cerebellar mutism in children is suggested to be related to the higher incidence of posterior fossa tumours in children. However, the question regarding the reason for the obvious difference in the incidence of this syndrome between the paediatric and adult population still remaining unanswered. The aim of this study was to evaluate and compare children and adult groups separately to understand the incidence and the clinical characteristics better and to elucidate the pathophysiological basis and predictive factors for this syndrome. METHOD: We reviewed, analysed, and compared the cases of cerebellar mutism individually in children and in adults reported in the English literature. We found 106 reported cases in children and 11 cases in adults which were suitable for analysis. We added two adult cases to these. FINDINGS: The ages of the patients ranged from 2 to 16 (mean, 6.4 year) in children and from 17 to 74 (mean, 38.7 year) in adults. Although vermis was the main location in both groups, the incidence of vermis lesions was considered higher in the paediatric population (%91.5 versus %69.2). The rate of brain stem invasion was prominent in children (%31.1) when compared with adults (%7.6). The latency for the development of mutism and the duration of the mutism were similar in children and adults (mean, 1.4 d versus 2 d and mean, 5.07 wk versus 4.2 wk respectively). Mutism was transient in all the cases of both groups. INTERPRETATION: Recent concepts of cerebellar physiology disclose the importance of the cerebellum in learning, language, and mental and social functions. Pontine nuclei, the thalamus, motor and sensory areas and supplementary motor areas have been proven necessary for the initiation of speech. It can be hypothesized that uncompleted maturation of the reciprocal links in childhood connecting the cerebellum to these structure makes the children more vulnerable to have postoperative cerebellar mutism in comparison to the adult population. PMID- 12111503 TI - Complications of closed continuous lumbar drainage of cerebrospinal fluid. AB - Sixty three patients who had a lumbar subarachnoid catheter placed for closed continuous cerebrospinal fluid drainage and the complications are presented. The drain was successful in achieving the desired goal in 59 patients (93,6%). The complications are mainly divided into 3 groups; A - complications related to alterations in CSF drainage rate, B - complications due to mechanical failure of the catheter, C - infection. The overall complication rate is found to be 44,4%. Overdrainage, pneumocephalus and meningitis are found to be the most severe complications, but most of these complications are reversible with early recognition. Unfortunately one patient died following meningitis and hepatic failure. Lumbar subarachnoid drainage is a safe method unless the development of any neurological findings should prompt rapid discontinuation of lumbar drainage and immediate radiographic evaluation. PMID- 12111504 TI - Expression of cyclin kinase inhibitor p21/WAF1 protein in pituitary adenomas: correlations with endocrine activity, but not cell proliferation. AB - p21/WAF1 blocks cell cycle progression through inhibition of cyclin/cyclin dependent kinases (cdks) complexes and, simultaneously, has been associated with cell cycle exit and the process of differentiation. In this series, the expression of p21/WAF1 was assessed immunohistochemically in 47 cases of pituitary adenomas in relation to endocrine activity and cell proliferation. To evaluate cell proliferation, a monoclonal antibody, MIB-1, against Ki-67 antigen was used in all of the available cases. The study revealed positive p21/WAF1 staining in 24 cases of 26 functioning parenchymas, whereas 14 cases of 21 non functioning parenchymas stained negative. The MIB-1 index ranged from less than 1% to 5.1% (mean: less than 1.7%) in functioning adenomas, and from less than 1% to 3.6% (mean: less than 1.6%) in non-functioning adenomas. Regardless of endocrine activity, p21/WAF1 positivity did not correlate with the MIB-1 index. Double staining techniques revealed the co-expression of p21/WAF1/GH or p21/WAF1/PRL in functioning adenomas. In 22 cases of p21/WAF1-positive functioning adenomas, p21/WAF1 immunoreactivity was seen in the cytoplasma as well as nuclei. These results indicate that in pituitary adenomas, p21/WAF1 expression is associated with endocrine activity, but not with cell proliferation. Taken together with recent findings demonstrating that cytoplasmic p21/WAF1 acts as an inhibitor of apoptosis, it is possible that pituitary adenomas expressing cytoplasmic p21/WAF1 have resistance against DNA-damaging agents such as ionizing radiation. PMID- 12111505 TI - Indirect carotid cavernous fistula appeared after balloon embolization of direct CCF. AB - The authors describe a case of indirect carotid cavernous fistula (CCF) appearing five months after embolization for traumatic direct CCF, which was treated six months after the trauma. Long-term (six months) venous hypertension to the affected cavernous sinus due to direct CCF and cavernous sinus thrombosis following a balloon embolization were considered as an etiology of the de novo dural arteriovenous fistula. The recurrent symptoms of CCF are usually related to detached balloon disorder, but delayed recurrence may be caused by the de novo dural AVF, if the direct CCF was treated in the chronic state. PMID- 12111506 TI - Vertebral epidural arteriovenous fistula and radicular pain in neurofibromatosis type I. AB - A minor trauma caused opening of an arteriovenous fistula between the right vertebral artery and cervical spinal epidural venous plexus in a patient with neurofibromatosis Type I. Subsequent dilation of the plexus caused compression of the spinal cord and radicular symptomology of the right upper extremity. The single-hole fistula and its arterial feeder were filled with electrodetachable coils via an intra-arterial approach. This lead into shrinkage of the plexus, reformation of the cord caliber and full and stable clinical recovery. The achieved endovascular occlusion of the fistula proved to be permanent on follow up. PMID- 12111508 TI - Complex developmental venous anomaly of the brain. PMID- 12111507 TI - Spontaneous bilateral chronic subdural haematoma of the posterior fossa. Case report and review of the literature. AB - INTRODUCTION: Chronic subdural haematomas of the posterior fossa in adults without a history of trauma are very rare. To our knowledge, only 15 cases have so far been reported in the literature, including those with anticoagulation therapy. A case of spontaneous bilateral infratentorial chronic subdural haematoma associated with anticoagulation therapy in an alive adult is presented and the relevant literature is reviewed. CASE REPORT: A 70 year old female presented with progressive dizziness, vertigo and gait ataxia. She was on anticoagulation therapy for heart disease. Neuro-imaging revealed bilateral infratentorial subdural masses. The subdural masses were suspects for chronic subdural haematomas by neuroradiological criteria. Because of the progressive symptomatology, the haematomas were emptied through burrhole trepanations. Chocolate-colored fluid, not containing clotted components, gushed out under great pressure. The source of bleeding could not be identified. The patient recovered well from surgery, but died 4 months later shortly after admission to another hospital from heart failure. DISCUSSION: The chronic subdural haematomas in this patient may have been due to rupture of bridging veins caused by a very mild trauma not noticed by the patient and possibly aggravated by the anticoagulation therapy. Infratentorial chronic subdural haematoma should at least be a part of the differential diagnosis in elderly patients with cerebellar and vestibular symptomatology even without a history of trauma. PMID- 12111509 TI - Candida albicans granuloma imitate clivus chordoma. PMID- 12111510 TI - Spinal cord injury with a stingray spine. PMID- 12111511 TI - Cost-effective and time-saving surgical treatment of pulmonary hydatid cysts with multiple localization. AB - PURPOSE: The most frequent anatomic locations of hydatid cysts are the liver and lungs. Because there is no effective medical therapy against this parasitic disease, surgery is the treatment of choice. The aim of this retrospective study was to compare the cost and effect of a one-stage operation with those of two- or three-stage operations in the treatment of lung hydatid cysts with multiple localizations. METHODS: We evaluated 364 patients who underwent surgical treatment for hydatid cysts, all of whom had multiple localizations. To avoid two or three-staged operations, we performed median sternotomy, simultaneous bilateral thoracotomy and unilateral thoracotomy with a transdiaphragmatic approach. RESULTS: For the treatment of 460 hydatid cyst localizations in 364 patients, a collective 381 operations were performed. The number of operations and periods of hospitalization were reduced. CONCLUSION: A one-stage surgical procedure for bilateral lung and liver hydatid cysts is superior to the traditional two- and three-stage operations because it reduces morbidity, hospital stay, and cost. PMID- 12111512 TI - Left retroperitoneal approach using a retractor to repair abdominal aortic aneurysms: a comparison with the transperitoneal approach. AB - PURPOSE: The purpose of this study was to compare the effectiveness of the retroperitoneal approach (RP) using a Thompson retractor with the conventional transperitoneal approach (TP), to repair infrarenal abdominal aortic aneurysms (AAA). METHODS: A total of 91 consecutive patients were divided into two groups; group A ( n = 21) underwent surgery using the TP, and group B ( n = 70) underwent surgery using the RP with a Thompson retractor. RESULTS: There were no significant differences in the operation time, aortic cross-clamp time, incidence of postoperative cardiac events, or the development of wound complications; however, a significantly higher rate of postoperative respiratory complications and ileus was observed in group A. Moreover, oral feeding was commenced later and the hospital stay was prolonged in group A ( P < 0.01). CONCLUSION: These findings clearly demonstrate that our RP method, especially when using a Thompson retractor, is a preferable alternative to TP for AAA surgery. PMID- 12111513 TI - Changes in the intestinal flora after the administration of prophylactic antibiotics to patients undergoing a gastrectomy. AB - PURPOSE: Changes in the intestinal flora were investigated in patients administered antibiotics for 4 days after a gastrectomy. METHODS: Twenty-four patients were divided into each of the following groups: cefazolin, 1 g every 8 h; cefozopran, 0.5 g every 8 h; flomoxef, 1 g every 8 h. All drugs were administered intravenously for a 4-day period. RESULTS: Antibiotic prophylaxis caused no major change in the total number of anaerobes because of the preservation of the Bacteroides fragilis group. However, the number of Bifidobacterium, Lactobacillus, Eubacterium, and Veillonella spp. decreased by an average of 100-fold. These changes were accompanied by an overgrowth of Enterococcus spp. and Pseudomonas aeruginosa. CONCLUSIONS: Four days of antibiotic prophylaxis after surgery caused a suppression of colonization resistance in the intestinal flora. Certain genera of anaerobes may thus play an important role in preventing overgrowth of antibiotic-resistant organisms after surgery. Surgeons should decide the duration of antibiotic use after carefully considering its influence on the intestinal flora. PMID- 12111514 TI - Evaluation of treatment for synchronous hepatic metastases from gastric cancer with special reference to long-term survivors. AB - PURPOSE: The purpose of this study was to determine the most effective means of treating liver metastases from gastric cancer. METHODS: We retrospectively examined 43 patients with liver metastases, but without peritoneal dissemination, who had received different forms of treatment. RESULTS: The crude 5-year survival rates of patients who underwent gastrectomy with complete hepatectomy (curative gastrectomy), noncurative gastrectomy, and no gastrectomy were 33.3%, 3.7%, and 0%, respectively. The curative gastrectomy group showed the highest survival rate (not significant). The median survival of patients given hepatic artery infusion (HAI), systemic chemotherapy, and no chemotherapy were 353, 189, and 61 days, respectively. The patients given chemotherapy survived significantly longer than those not given chemotherapy. Three patients survived for more than 5 years without any signs of recurrence. The long-term survivors all had primary lesions without serosal invasion (T2) and no other noncurative factors. Two patients underwent curative gastrectomy and one underwent noncurative gastrectomy. All were given postoperative chemotherapy; as HAI in two cases, and as systemic chemotherapy in one case. CONCLUSION: These findings suggest that curative gastrectomy combined with HAI or systemic chemotherapy should be attempted for patients with primary tumors without serosal invasion or any other noncurative factors. PMID- 12111515 TI - Surgical management of spontaneous intrabiliary rupture of hydatid liver cysts. AB - PURPOSE: The most common complication of hydatid liver cysts is spontaneous rupture into the biliary tract. This study was conducted to evaluate the surgical management of spontaneous intrabiliary rupture of a hydatid liver cyst in 41 patients. METHODS: The preoperative diagnosis was confirmed by ultrasound in all 41 patients, 37 of whom were jaundiced. RESULTS: According to Gharbi's classification, 39% of the cysts were type III and they ranged from 3 to 18 cm in diameter, with a mean diameter of 9 cm. The mean total bilirubin and alkaline phosphatase values were 6.3 mg/dl and 450 IU, respectively. Partial cystectomy, cholecystectomy, and common bile duct exploration were performed in all patients. In seven patients, the visible biliary duct within the cyst cavity was sutured with 2/0 silk. Intraoperative cholangiography was performed in all patients, and choledochoscopy was performed in 11 patients. A T-tube was inserted after the biliary tract content was thought to have been totally cleaned out in 38 patients (93%), and a choledochoduodenostomy was performed in 3 patients (7%). An external biliary fistula developed in five patients, persisting for 11-25 days. The fistulae healed within a mean period of 5 days after endoscopic sphincterotomy (EST). For patients without a fistula the mean hospitalization time was 8 days and there was no mortality. CONCLUSION: These results suggest that when a hydatid liver cyst ruptures into the biliary tract, common bile duct exploration should be conducted using intraoperative cholangiography and choledochoscopy. If the biliary tract is cleaned of all cystic content, T-tube drainage should be sufficient, but EST is an effective technique for treating persistent extended external biliary fistulae. PMID- 12111516 TI - Reevaluation of the indications for radical pancreatectomy to treat pancreatic carcinoma: is portal vein infiltration a contraindication? AB - PURPOSE: Portal vein resection (PVR) has become more widely performed owing to improvements in the perioperative mortality rate. The present study was performed to determine whether portal vein infiltration is a contraindication against radical pancreatectomy for patients with pancreatic carcinoma. METHODS: Between 1990 and 1997, a total of 66 patients with invasive ductal carcinoma of the pancreas underwent surgical resection at the Department of Surgery II, Hokkaido University Hospital. After the exclusion of those who underwent distal pancreatectomy, the remaining 43 patients were divided into a PVR(+) group ( n = 28) and a PVR(-) group ( n = 15). The clinicopathological characteristics, morbidity, and mortality were statistically compared between the two groups. RESULTS: The overall survival rate of the patients who required PVR was not significantly different from that of those who underwent pancreatic resection without PVR. CONCLUSION: These findings suggest that combined PVR should not be a contraindication to radical pancreatectomy for pancreatic carcinoma with positive vascular invasion. PMID- 12111517 TI - Significance of serum endotoxin and antiendotoxin antibody levels in predicting the severity of acute pancreatitis. AB - PURPOSE: A close association between endotoxemia and acute pancreatitis has been reported, and attempts have been made to predict the severity of pancreatitis by estimating the levels of endotoxin. The present study was designed to correlate endotoxemia with the severity and complications of acute pancreatitis as graded by contrast-enhanced computed tomography and Blamey's criteria. METHODS: We examined 20 patients with acute pancreatitis, using Blamey's criteria to assess the severity of pancreatitis. The endotoxin level was estimated by the Limulus Amoebocyte Lysate method and the antiendotoxin antibody level was assayed by the enzyme-linked immunoassay technique measuring combined levels of IgG and IgM. RESULTS: Severe pancreatitis was confirmed in 9 of the 20 patients, 17 (85%) of whom were found to have endotoxemia. There was no correlation between the presence and level of endotoxemia and the severity of pancreatitis; however, antiendotoxin antibody titers were significantly lower in patients with severe disease ( P < 0.05), those who suffered of major complications ( P < 0.01), and those who died of the disease ( P < 0.01). CONCLUSION: The findings of this study demonstrated that the presence of endotoxemia accompanied by a fall in antiendotoxin antibody titer predicts poor outcome in patients with acute pancreatitis. PMID- 12111518 TI - c-myc amplification and enhancement of sensitivity to cytosine arabinoside: an in vitro and in vivo study on four sublines established from a pulmonary adenocarcinoma. AB - PURPOSE: Meticulous treatment strategies taking tumor heterogeneity into account are considered essential to achieve breakthroughs in current cancer therapy. We analyzed tumor heterogeneity in the primary tumor of a patient with pulmonary adenocarcinoma characterized by a poor prognosis. METHODS: Four sublines with different growth characteristics in vitro were established from the tumor using a method for short-term selective cultivation. We examined the differences in morphological, biochemical, and genetic findings of these sublines. RESULTS: Differences in the histological features of the transplanted tumors were seen in the four sublines. The 88-2T and 88-2 tumors revealed a well-differentiated adenocarcinoma; the 88-2F tumor revealed a large cell-like carcinoma resembling the metastatic tumor in the lymph nodes; and the 88-2FA tumor was composed of signet-ring cells. There were differences in oncogenes, with the 88-2F line alone exhibiting 12-fold amplification of c-myc. Sensitivity to cytosine arabinoside (Ara C) was specifically increased in the 88-2F cell line, alone. CONCLUSIONS: These sublines demonstrate that human pulmonary adenocarcinoma has various types of heterogeneity within the primary tumor. Furthermore, c-myc amplification may play an important role in altering phenotype and growth characteristics in vitro and in vivo, and for increasing sensitivity to Ara C and the potential of cancer cells to metastasize to lymph nodes. PMID- 12111519 TI - Can nonpenetrating vascular closure staples and hepatocyte growth factor prevent intimal hyperplasia following ePTFE grafting of the carotid artery in rabbits? AB - PURPOSE: To evaluate whether nonpenetrating vascular closure staples (VCS) and hepatocyte growth factor (HGF) can effectively prevent anastomotic intimal hyperplasia. METHODS: An expanded polytetrafluoroethylene graft, 2 mm in diameter, was implanted in the common carotid artery of rabbits divided into three experimental groups. In the control group, distal anastomosis was performed with interrupted suturing; in the VCS group, clips were applied along the lateral suture line after the placement of stay sutures; and in the VCS + HGF group, the same anastomotic technique was performed as in the VCS group, followed by the administration of the HGF for 4 days. RESULTS: The time taken to complete the anastomosis was significantly less in both the VCS groups than in the control group ( P < 0.0001). On postoperative day (POD) 28, the patency rate was significantly lower ( P < 0.05) in the VCS group (42.9%) than in the control group (100%), but the rate in the VCS + HGF group (100%) was the same as that in the control group. Intimal thickness was significantly less in the control group than in either the VCS or VCS + HGF groups ( P< 0.05). The percentage of area stenosis was significantly less ( P< 0.01) in the control group than in the VCS group. CONCLUSION: The VCS clip failed to suppress intimal thickness or reduce the percentage of stenosis at the anastomotic site. PMID- 12111520 TI - Simultaneous occurrence of papillary intrafollicular and microcarcinomas with bilateral medullary microcarcinoma of the thyroid in a patient with multiple endocrine neoplasia type 2A: report of a case. AB - We report the case of a simultaneous occurrence of papillary intrafollicular and microcarcinomas with bilateral medullary microcarcinoma of the thyroid in a patient with multiple endocrine neoplasia type 2A. The concurrent presence of two thyroid carcinomas is rare. The simultaneous occurrence of two different tumors in the same thyroid each being multifocal and smaller than 1 cm in diameter has not been previously reported in the literature. Furthermore, we define the first case of intrafollicular papillary thyroid carcinoma (carcinoma in situ). PMID- 12111521 TI - Primary thyroid carcinoma found with esophageal stenosis: report of a case. AB - The typical symptoms of primary thyroid cancer are a cervical mass and cervical lymphadenopathy, while dyspnea, hoarseness, or dysphagia can occur in the presence of extrathyroidal involvement. Esophageal involvement or stenosis causing dysphagia without any influence on the trachea is rare because of the anatomical location of the esophagus and the trachea. We report herein a case of primary thyroid carcinoma advancing behind the esophagus with the trachea intact, which was difficult to diagnose by esophagoscopy or roentgenogram. Thus, thyroid tumors need to be differentiated from other possible causes of esophageal stenosis. Computed tomography, magnetic resonance imaging, and ultrasonography are useful diagnostic modalities to detect thyroid tumors causing esophageal stenosis. PMID- 12111523 TI - The use of a Greenfield filter to treat a pregnant woman for internal jugular venous thrombosis: report of a case. AB - Internal jugular venous thrombosis is an unusual entity with the potential to develop into pulmonary embolism (PE). A 28-year-old woman at 15 weeks gestational age of pregnancy was referred to our hospital for pain and swelling on the left side of her neck. Magnetic resonance imaging and computed tomography of her neck revealed an occlusion of the left internal jugular vein. Left internal jugular venous thrombosis was thus diagnosed. She was successfully treated by placement of a Greenfield filter in the superior vena cava and delivered a full-term healthy infant. This procedure could be an effective and safe method to prevent PE in patients of internal jugular venous thrombosis in whom anticoagulation therapy has either failed or is contraindicated. PMID- 12111522 TI - An atypical lung carcinoid tumor resected after induction therapy with involvement of the superior sulcus region: report of a case. AB - This report presents a case of lung carcinoid tumor that showed a growth pattern similar to that of a superior sulcus tumor (SST). A 63-year-old man was referred to our hospital and was diagnosed to have a stage IV (T3N2M1) SST on his right side. After three cycles of induction therapy with MVP (methotrexate/vinblastine/prednisolone) and a total dose of 45 Gy radiation given to the chest lesion, the clinical stage was restaged down to IIB (T3N0M0). A salvage operation was performed (upper lobectomy plus chest wall combined resection including the first to fourth ribs) followed by targeting adjuvant brachytherapy. The results of a histologic examination of the resected specimen revealed the tumor to be an atypical carcinoid tumor. PMID- 12111524 TI - Successful local excision of ileostomy adenocarcinoma after colectomy for familial adenomatous polyposis: report of a case. AB - Primary adenocarcinoma rarely develops at the site of an ileostomy performed for ulcerative colitis (UC), familial adenomatous polyposis (FAP), or Crohn's disease. We describe a case of ileostomy cancer found 14 years after proctocolectomy for FAP with cancer of both the sigmoid colon and rectum. Resection of the ileal mucosa around the stoma was performed three times. To our knowledge, only 35 other such cases have ever been reported. Our review of these cases indicates that routine examination of the stoma by a physician, or even by the patient, may lead to earlier detection of this rare complication, and a better chance of cure through minimal surgery. PMID- 12111525 TI - Diffuse small bowel Crohn's disease treated with side-to-side isoperistaltic strictureplasty: report of two cases and description of a variation of the original technique. AB - Diffuse small bowel Crohn's disease is unusual and it is characterized by multiple diseased segments involving the jejunum and ileum. The most frequent indication for surgery is an intestinal obstruction, often complicated by a high grade of malnutrition. The natural history of this clinical form is not well defined and the optimal surgical approach remains controversial. We herein present our surgical policy in two cases of diffuse small bowel Crohn's disease, who were particularly at risk of developing short bowel syndrome. We focused our attention on the use of side-to-side isoperistaltic strictureplasty as described by Michelassi for the treatment of stenoses longer than 20 cm. We also propose the application of this technique for the treatment of shorter stenosis cases. PMID- 12111527 TI - Mesenteric Castleman's disease: report of a case. AB - A 77-year-old woman was admitted with intermittent abdominal dull pain. Hypochromic anemia, hypergammaglobulinemia, and elevated C-reactive protein were found in this case. Ultrasonography, computed tomography, magnetic resonance imaging, and angiography indicated either mesenteric leiomyoma or leiomyosarcoma, but no definitive preoperative diagnosis could be established. A surgical resection of the tumor revealed a mesenteric Castleman's tumor with small daughter lymphoid tumors of plasma cell type. PMID- 12111526 TI - Renal focal tubulointerstitial fibrosis with short bowel syndrome: report of a case. AB - We report a male patient with short bowel syndrome (SBS) and renal focal tubulointerstitial fibrosis (FTIF). Seven years after surgery, he was introduced to us due to severe undernutrition, an impairment of growth hormone (GH) secretion, and abnormally low levels of plasma citrulline and arginine at 11 years 7 months of age, just before nutritional support using total parenteral nutrition (TPN) was begun. Thereafter, the support was changed to home TPN with GH supplementation. After an improvement of the disorders, GH was stopped at 17 years 3 months of age. However, hyperuricemia appeared and a renal biopsy revealed FTIF at 20 years of age. Home TPN was continued twice a week because the plasma arginine level was still low. His follow-up biopsy at 23 years of age showed morphometric amelioration. Arginine deficiency following SBS may be associated with FTIF. The cause of hyperuricemia after SBS therefore needs to be investigated in detail. PMID- 12111529 TI - Laparoscopic treatment for peripheral pancreatic duct injury after blunt abdominal trauma: report of a case. AB - We report the successful use of laparoscopy to treat a blunt pancreatic trauma (BPT) in a 33-year-old woman involved in a traffic accident. Computed tomography showed peripancreatic effusion and indicated an injury to the main pancreatic duct (MPD). Urgent endoscopic retrograde pancreatography (ERP) was performed. The ERP revealed a leakage of contrast medium from the peripheral pancreatic duct. The patient underwent drainage of pancreatic effusion using laparoscopic techniques. The patient had an uneventful course, and no complications have been detected a year after the operation. These results indicate that emergency ERP and laparoscopic drainage are appropriate for patients with peripancreatic effusion due to peripheral pancreatic duct injury. PMID- 12111528 TI - Hepatic portal venous gas caused by blunt abdominal trauma: is it a true ominous sign of bowel necrosis? Report of a case. AB - A case of transient portal venous gas in the liver following blunt abdominal trauma is described. Computed tomography (CT) demonstrated hepatic portal venous gas 4 h after the injury. An exploratory laparotomy revealed segmental necrosis of the small intestine with a rupture of the bladder. Pneumatosis intestinalis was evident on the resected bowel. A histopathologic study revealed congestion and bleeding in the bowel wall and a great deal of the mucosa had been lost because of necrosis. However, neither thrombus nor atherosclerotic changes were observed in the vessels. A bacteriological examination demonstrated anaerobic bacteria from the bowel mucosa, which was most likely to produce portal venous gas. Although the present case was associated with bowel necrosis, a review of literature demonstrated that portal venous gas does not necessarily indicate bowel necrosis in trauma patients. There is another possibility that the portal venous gas was caused by a sudden increase in the intra-abdominal pressure with concomitant mucosal disruption, which thus forced intraluminal gas into the portal circulation in the blunt trauma patients. PMID- 12111530 TI - Hand-assisted laparoscopic distal pancreatectomy in pigs using our original hand device (Lap Disc). AB - We analyzed the safety and feasibility of a hand-assisted laparoscopic distal pancreatectomy (HALDP) in pigs. We performed HALDP in five cases using our original hand device (Lap Disc). The procedures in all cases were successfully performed. The Lap Disc was very useful for performing HALDP because of its compact design and simple usage. This preliminary experience suggests that this device could be used to perform HALDP as well as other procedures in humans. PMID- 12111532 TI - "What watch?...such much!" Complexity and evolution of circadian clocks. AB - This review uses three examples to summarise our knowledge about the complexity and the evolution of circadian systems. The first example describes the ecology of unicellular algae, which use their circadian system to optimise the daily exploitation of resources that are spatially separated. The second example looks at the role of clocks in tissues and cells within a complex organism, and the third speculates on how the circadian system may have evolved. PMID- 12111533 TI - Clock mechanisms in Drosophila. AB - Mechanisms underlying circadian clock function in Drosophila melanogaster have been revealed by genetic and molecular approaches. Two interlocked transcriptional feedback loops involving at least the period, timeless, Clock,and cycle genes generate molecular oscillations that are believed to control behavioral rhythmicity and other clock outputs. These oscillations are further enhanced and fine-tuned to match the duration of the solar day by post transcriptional and post-translational mechanisms depending on the PERIOD and TIMELESS proteins and on the protein kinases DOUBLE-TIME and SHAGGY. Light is the principal zeitgeber for synchronizing molecular and behavioral rhythmicity via the blue-light photoreceptor CRYPTOCHROME and the TIMELESS protein. In addition, light seems required for maintaining robust molecular oscillations at least in peripheral clock-gene-expressing tissues like the eyes, antennae, or Malpighian tubules. Relaying temporal information to cells and tissues expressing overt biological rhythms involves regulation of "output genes" at multiple levels. Although their regulation depends on the major clock genes, the majority of the clock-controlled genes are not direct targets of clock factors. PMID- 12111534 TI - Clock mechanisms in zebrafish. AB - Recent research on the circadian system of the zebrafish is reviewed. This teleost has become an attractive model system because of its advantages for genetic analyses. Circadian rhythms of zebrafish behavior, visual system function, and pineal melatonin synthesis have been described, and behavioral and pineal rhythms are being used to identify and characterize clock mutants. Zebrafish heart, kidney, and embryonic cell lines contain circadian oscillators and phototransduction mechanisms for entrainment, suggesting that circadian pacemaking functions may be distributed throughout the animal. Studies of circadian system development in zebrafish have found that a molecular circadian oscillation in unfertilized oocytes persists through embryonic development with its phase intact, but that the pacemakers that drive rhythms of melatonin synthesis and behavior require environmental entraining signals late in development for initial synchronization. Zebrafish homologs of several of the core clock genes identified in other animals have been cloned. Transcripts for most of these are rhythmically expressed in multiple tissues. The interactions of clock gene products are for the most part similar to their interactions in mammals, although there are some potentially interesting differences. PMID- 12111535 TI - Rhythm and soul in the avian pineal. AB - The avian pineal gland, like that of mammals, displays a striking circadian rhythm in the synthesis and release of the hormone melatonin. However, the pineal gland plays a more prominent role in avian circadian organization and differs from that in mammals in several ways. One important difference is that the pineal gland in birds is relatively autonomous. In addition to making melatonin, the avian pineal contains photoreceptors and a circadian clock (thus, an entire circadian system) within itself. Furthermore, avian pineals retain their circadian properties in organ or dispersed cell culture, making biochemical components of regulatory pathways accessible. Avian pinealocytes are directly photosensitive, and novel candidates for the unidentified photopigments involved in the regulation of clock function and melatonin production, including melanopsin, pinopsin, iodopsin, and the cryptochromes, are being evaluated. Transduction pathways and second messengers that may be involved in acute and entraining effects, including cyclic nucleotides, calcium fluxes, and protein kinases, have been, and continue to be, examined. Moreover, several clock genes similar to those found in Drosophila and mouse are expressed, and their dynamics and interactions are being studied. Finally, the bases for acute and clock regulation of the key enzyme in melatonin synthesis, arylalkylamine N acetyltransferase (AA-NAT), are described. The ability to study entrainment, the oscillator itself, and a physiological output in the same tissue at the same time makes the avian pineal gland an excellent model to study the bases and regulation of circadian rhythms. PMID- 12111536 TI - Molecular machinery of the circadian clock in mammals. AB - The discovery of clock genes and the general principle of their oscillation has stimulated research on biological clocks and this research has had a major impact on the field of life sciences. The mammalian circadian core oscillator is thought to be composed of an autoregulatory transcription-(post)translation-based feedback loop involving a set of clock genes. The real time monitoring of clock gene oscillation at the levels of genes, cells, tissues, and systems will clarify the issue of how the time signal is born and how it is integrated into the organismic level. Investigations of circadian systems in various organisms employ multiple methods including ethology, physiology, neuroscience, molecular biology, cell biology and genetics. The circadian system has thus become a unique example in the elucidation of the general principles of how genes control cellular, systemic and behavioral functions. PMID- 12111537 TI - Opsins and mammalian photoentrainment. AB - Research over the past decade has provided overwhelming evidence that photoreception in the vertebrate eye is not confined to the rod and cone photoreceptors. It appears that photoreceptor cells within the inner retina provide irradiance information to a wide variety of different photosensory tasks including photoentrainment, pupillary constriction and masking behaviour. Action spectra in mice lacking all rod and cone photoreceptors ( rd/rd cl) have demonstrated the existence of a previously uncharacterised, opsin/vitamin-A-based photopigment with peak sensitivity at 479 nm (opsin photopigment/OP(479)). The review addresses the question: has the gene encoding OP(479) already been isolated, and if not, what type of gene should we be seeking and where in the eye might this gene be expressed? On the basis of available data, the gene that encodes OP(479) remains unidentified, and two broad possibilities exist. On the assumption that OP(479) will be like all of the other vertebrate photopigments (ocular and extraocular) and share a close phylogenetic relationship based upon amino acid identity and a conserved genomic structure, then the gene encoding OP(479) has yet to be isolated. Alternatively, there may have been a separate line of photopigment evolution in the vertebrates that has given rise to the melanopsin family. If true then the mammalian melanopsin gene may encode OP(479). Only when melanopsin and other candidates for OP(479) have been functionally expressed, and shown to encode a photopigment that matches the action spectrum of OP(479), can firm conclusions about the identity of the non-rod, non-cone ocular photoreceptor of mammals be made. PMID- 12111538 TI - Neurotransmitters of the retino-hypothalamic tract. AB - The brain's biological clock, which, in mammals, is located in the suprachiasmatic nucleus (SCN), generates circadian rhythms in behaviour and physiology. These biological rhythms are adjusted daily (entrained) to the environmental light/dark cycle via a monosynaptic retinofugal pathway, the retinohypothalamic tract (RHT). In this review, the anatomical and physiological evidence for glutamate and pituitary adenylate cyclase-activating polypeptide (PACAP) as principal transmitters of the RHT will be considered. A combination of immunohistochemistry at both the light- and electron-microscopic levels and tract tracing studies have revealed that these two transmitters are co-stored in a subpopulation of retinal ganglion cells projecting to the retino-recipient zone of the ventral SCN. The PACAP/glutamate-containing cells, which constitute the RHT, also contain a recently identified photoreceptor protein, melanopsin, which may function as a "circadian photopigment". In vivo and in vitro studies have shown that glutamate and glutamate agonists such as N-methyl- D-aspartate mimic light-induced phase shifts and that application of glutamate antagonists blocks light-induced phase shifts at subjective night indicating that glutamate mediates light signalling to the clock. PACAP in nanomolar concentrations has similar phase-shifting capacity as light and glutamate, whereas PACAP in micromolar concentrations modulates glutamate-induced phase shifts. Possible targets for PACAP and glutamate are the recently identified clock genes Per1 and Per2, which are induced in the SCN by light, glutamate and PACAP at night. PMID- 12111539 TI - Suprachiasmatic nucleus organization. AB - The suprachiasmatic nucleus (SCN) of the hypothalamus is a dominant circadian pacemaker in the mammalian brain controlling the rest-activity cycle and a series of physiological and endocrine functions to provide a foundation for the successful elaboration of adaptive sleep and waking behavior. The SCN is anatomically and functionally organized into two subdivisions: (1) a core that lies adjacent to the optic chiasm, comprises predominantly neurons producing vasoactive intestinal polypeptide (VIP) or gastrin-releasing peptide (GRP) colocalized with GABA and receives dense visual and midbrain raphe afferents, and (2) a shell that surrounds the core, contains a large population of arginine vasopressin (AVP)-producing neurons in its dorsomedial portion, and a smaller population of calretinin (CAR)-producing neurons dorsally and laterally, colocalized with GABA, and receives input from non-visual cortical and subcortical regions. In this paper, we present a detailed quantitative analysis of the organization of the SCN core and shell in the rat and place this in the context of the functional significance of the subdivisions in the circadian control of regulatory systems. PMID- 12111540 TI - Signaling in the suprachiasmatic nucleus: selectively responsive and integrative. AB - The suprachiasmatic nucleus (SCN) contains a biological clock that generates timing signals that drive daily rhythms in behaviors and homeostatic functions. In addition to this pacemaker function, the SCN gates its own sensitivity to incoming signals, which permits appropriate temporal adjustment to achieve synchrony with environmental and organismic states. A series of time-domains, in which the SCN restricts its own sensitivity to a limited set of stimuli that adjust clock phase, can be distinguished. Pituitary adenylyl cyclase-activating peptide (PACAP) and cAMP directly reset clock phase during the daytime domain; both cause phase advances only during the clock's day-time domain, but are without effect at night. In contrast, acetylcholine and cGMP analogs phase advance the clock only when applied during the night. Sensitivity to light and glutamate arises concomitant with sensitivity to acetylcholine and cGMP. Light and glutamate cause phase delays in the early night, by elevating intracellular Ca(2+) via neuronal ryanodine receptors. In late night, light and glutamate utilize a cGMP-mediated mechanism to induce phase advances. Nocturnal responses of SCN primed by light or glutamate can be modulated by effectors of phase resetting in daytime, namely, PACAP and cAMP. Finally, the dusk and dawn domains are characterized by sensitivity to the pineal hormone, melatonin, acting through protein kinase C. These changing patterns of sensitivities demonstrate that the circadian clock controls multiple intracellular gates, which ensures that they can be opened selectively only at specific points in the circadian cycle. Discerning the molecular bases of these changes is fundamental to understanding integrative and regulatory mechanisms in the circadian system. PMID- 12111541 TI - Output pathways of the mammalian suprachiasmatic nucleus: coding circadian time by transmitter selection and specific targeting. AB - Every day, we experience profound changes in our mental and physical condition as body and brain alternate between states of high activity during the waking day and rest during night-time sleep. The fundamental evolutionary adaptation to these profound daily changes in our physiological state is an endogenous 24-h clock. This biological clock enables us to prepare ourselves to these daily changes, instead of only being able to show a passive and delayed response. During the past decade, enormous progress has been made in determining possible molecular components of the biological clock. An important question remains, however, regarding how the rhythmic signal from the biological clock is spread throughout the body to control its physiology and behavior. Indeed, ultimately, the only raison d'etre for the biological clock is its output (Green 1998). In the present review, we propose that the main mechanism for the spreading time-of day information throughout the body consists of different circadian waves of suprachiasmatic nucleus (SCN) transmitter release, directed to a restricted number of specific SCN target areas, and affecting both neuroendocrine mechanisms and the peripheral autonomic nervous system. PMID- 12111542 TI - The mammalian retina as a clock. AB - Many physiological, cellular, and biochemical parameters in the retina of vertebrates show daily rhythms that, in many cases, also persist under constant conditions. This demonstrates that they are driven by a circadian pacemaker. The presence of an autonomous circadian clock in the retina of vertebrates was first demonstrated in Xenopus laevis and then, several years later, in mammals. In X. laevis and in chicken, the retinal circadian pacemaker has been localized in the photoreceptor layer, whereas in mammals, such information is not yet available. Recent advances in molecular techniques have led to the identification of a group of genes that are believed to constitute the molecular core of the circadian clock. These genes are expressed in the retina, although with a slightly different 24-h profile from that observed in the central circadian pacemaker. This result suggests that some difference (at the molecular level) may exist between the retinal clock and the clock located in the suprachiasmatic nuclei of hypothalamus. The present review will focus on the current knowledge of the retinal rhythmicity and the mechanisms responsible for its control. PMID- 12111543 TI - Control of melatonin synthesis in the mammalian pineal gland: the critical role of serotonin acetylation. AB - The large daily rhythm in circulating melatonin levels is a highly conserved feature of vertebrate physiology: high values always occur at night. The dynamics of the rhythm are controlled by the next-to-last enzyme in melatonin synthesis (serotonin --> N-acetylserotonin --> melatonin), arylalkylamine N acetyltransferase (AANAT), the "melatonin rhythm enzyme". In vertebrate biology, AANAT plays a unique time-keeping role as the molecular interface between the environment and the hormonal signal of time, melatonin. This chapter describes the mammalian AANAT regulatory system, which includes the retina, neural structures, transsynaptic processes, and molecular events. In addition, special attention is paid to the functional characteristics of the systems which insure that the nocturnal increase in melatonin is an accurate and reliable indicator of the duration of the night, and why the melatonin rhythm is the most reliable output signal of the Mind's Clock. PMID- 12111544 TI - The anatomy and innervation of the mammalian pineal gland. AB - The parenchymal cells of the mammalian pineal gland are the hormone-producing pinealocytes and the interstitial cells. In addition, perivascular phagocytes are present. The phagocytes share antigenic properties with microglial and antigen presenting cells. In certain species, the pineal gland also contains neurons and/or neuron-like peptidergic cells. The peptidergic cells might influence the pinealocyte by a paracrine secretion of the peptide. Nerve fibers innervating the mammalian pineal gland originate from perikarya located in the sympathetic superior cervical ganglion and the parasympathetic sphenopalatine and otic ganglia. The sympathetic nerve fibers contain norepinephrine and neuropeptide Y as neurotransmitters. The parasympathetic nerve fibers contain vasoactive intestinal peptide and peptide histidine isoleucine. Recently, neurons in the trigeminal ganglion, containing substance P, calcitonin gene-related peptide, and pituitary adenylate cyclase-activating peptide, have been shown to project to the mammalian pineal gland. Finally, nerve fibers originating from perikarya located in the brain containing, for example, GABA, orexin, serotonin, histamine, oxytocin, and vasopressin innervate the pineal gland directly via the pineal stalk. Biochemical studies have demonstrated numerous receptors on the pinealocyte cell membrane, which are able to bind the neurotransmitters located in the pinealopetal nerve fibers. These findings indicate that the mammalian pinealocyte can be influenced by a plethora of neurotransmitters. PMID- 12111545 TI - Mammalian melatonin receptors: molecular biology and signal transduction. AB - The pineal hormone, melatonin, is an important regulator of seasonal reproduction and circadian rhythms. Its effects are mediated via high-affinity melatonin receptors, located on cells of the pituitary pars tuberalis (PT) and suprachiasmatic nucleus (SCN), respectively. Two subtypes of mammalian melatonin receptors have been cloned and characterized, the MT1 (Mel(1a)) and the MT2 (Mel(1b)) melatonin receptor subtypes. Both subtypes are members of the seven transmembrane G protein-coupled receptor family. By using recombinant melatonin receptors it has been shown that the MT1 melatonin receptor is coupled to different G proteins that mediate adenylyl cyclase inhibition and phospholipase C beta activation. The MT2 receptor is also coupled to inhibition of adenylyl cyclase and additionally it inhibits the soluble guanylyl cyclase pathway. In mice with a targeted deletion of the MT1 receptor, the acute inhibitory effects of melatonin on SCN multiunit activity are completely abolished, while the phase shifting responses to melatonin (given in physiological concentrations) appear normal. Furthermore, melatonin inhibits the phosphorylation of the transcription factor cyclic AMP response element binding protein, induced by the pituitary adenylate cyclase-activating polypeptide in SCN cells predominantly via the MT1 receptor. However, a functional MT2 receptor in the rodent SCN is partially able to compensate for the absence of the MT1 receptor in MT1 receptor-deficient mice. These findings indicate redundant and non-redundant roles of the receptor subtypes in regulating SCN function. In the PT, a functional MT1 receptor is essential for the rhythmic synthesis of the clock gene product mPER1. Melatonin produces a long-lasting sensitization of adenylyl cyclase and thus amplifies cyclic AMP signaling when melatonin levels decline at dawn. This action of melatonin amplifies gene expression rhythms in the PT and provides a mechanism for reinforcing rhythmicity in peripheral tissues which themselves lack the capacity for self-sustained oscillation. Mice with targeted deletion of melatonin receptor subtypes provide an excellent model to understand cellular mechanisms through which melatonin modulates circadian and photoperiodic rhythmicity. PMID- 12111546 TI - The pars tuberalis as a target of the central clock. AB - The pars tuberalis (PT) of the pituitary has emerged from being a gland of obscure and unknown function to a tissue of central importance to our understanding of how photoperiod regulates seasonal responses. The discovery of melatonin receptors on this gland first pointed to its involvement in seasonal physiology. However, the more recent demonstration of the expression of clock genes in the PT, such as Per1, has heightened interest in the gland. Recent work shows how photoperiod, through the hormone melatonin, affects the timing and amplitude of expression of the Per1 gene, as well as other genes such as Icer. The effect of photoperiod and melatonin on the expression of Per1 in the PT is distinct to its effects on the SCN, and this probably reflects distinct functions of the clock genes in the two tissues - acting as part of the biological clock in the SCN, but as an interval timing system within the PT. The changes in amplitude of Per1 gene expression in response to altered length of photoperiod have provided the first clues as to how the durational melatonin signal is decoded within the neuroendocrine system. PMID- 12111547 TI - Organisation of the circadian system in melatonin-proficient C3H and melatonin deficient C57BL mice: a comparative investigation. AB - In all vertebrates melatonin is rhythmically synthesised in the pineal gland and functions as a hormonal message encoding for the duration of darkness. This review focuses on the role of melatonin in the circadian organisation of mammals by comparing signal transduction mechanisms in the pineal organ, the suprachiasmatic nucleus and the hypophyseal pars tuberalis in melatonin proficient (C3H) and melatonin-deficient (C57BL) mice strains. Surprisingly, the major signal transduction cascades in the pineal organ did not differ between the two mouse strains. With regard to the suprachiasmatic nucleus, the site of the endogenous clock, it was found that melatonin at most sets the gain for clock error signals mediated via the retinohypothalamic tract, but has no effect on the rhythm generation itself or on the maintenance of the oscillation. In contrast, melatonin plays an essential role in the control of the hypophyseal pars tuberalis. Here it acts in concert with adenosine to elicit rhythms in clock gene expression. Melatonin opens a temporally restricted gate for adenosine to induce cyclic AMP (cAMP)-sensitive genes by sensitising the adenylyl cyclase. This interaction, which grants a temporally precise regulation of gene expression, may reflect the central role of melatonin, i.e. in synchronising peripheral clock cells that require unique phasing of output signals with the master clock in the brain. PMID- 12111548 TI - The chronobiotic properties of melatonin. AB - In mammals, the exact role of melatonin (Mel) in the circadian timing system remains to be determined. However, exogenously administered Mel, as reported in the present mini-review, has been shown to affect the circadian clock. The sites and mechanisms of action involved in this "chronobiotic" effect of Mel have begun to be characterized. The suprachiasmatic nuclei (SCN) appear to be an important site for the entrainment effect of Mel and the presence of Mel receptors appears to be a prerequisite. However, the pharmacological dose of Mel needed to entrain circadian rhythms means that very probably other sites and mechanisms also play a role. PMID- 12111549 TI - Clock genes in mammalian peripheral tissues. AB - For many years, neurons of the suprachiasmatic nucleus (SCN) in the hypothalamus were thought to contain the unique mammalian clock controlling circadian rhythmicity of peripheral tissues via neural and humoral signals. Surprisingly, the cloning and characterisation of mammalian clock genes have revealed that they are expressed in a circadian manner throughout the body. It is generally accepted now that peripheral cells contain a circadian clock which is similar to the one present in SCN neurons, although only the latter seems to be self-sustained. It is still unclear how these peripheral clocks are synchronised by the central SCN clock, albeit humoral signals appear to be crucial. Interestingly, peripheral clocks can be uncoupled from the central clock in particular conditions such as restricted-feeding, allowing peripheral tissues to adapt themselves to cues incompatible to other cues perceived by the SCN (mainly the photoperiod). Whereas circadian clocks have been intensively dissected, little is known about the mechanisms by which these clocks regulate the expression of clock-controlled genes. Direct regulation for some of them by the products of clock genes was recently documented, but this probably represents the exception rather than the rule. We should soon be able to describe complete circadian transcriptional cascades from clock genes to enzymes and structural proteins. In addition to circadian humoral and neural signals, these cascades should help us to understand how gene expression, physiology and behaviour are influenced by the rotation of the Earth around its axis. PMID- 12111550 TI - The novel Streptomyces olivaceoviridis ABC transporter Ngc mediates uptake of N acetylglucosamine and N,N'-diacetylchitobiose. AB - During cultivation in the presence of N-acetylglucosamine or chitin, Streptomyces olivaceoviridis mycelium efficiently takes up [(14)C]-labelled N acetylglucosamine. Uptake of the labelled compound can be completely inhibited by unlabelled N-acetylglucosamine and partially by chitobiose. After extraction of the membrane with Triton X-100, two forms of a protein that binds to N acetylglucosamine and N, N'-diacetylchitobiose (chitobiose) were purified to homogeneity by two consecutive rounds of anionic exchange chromatography. The protein was named NgcE. Using surface plasmon resonance, its binding parameters were determined. It showed highest affinity for N-acetylglucosamine (K(D)=8.28 x 10(-9) M) and for chitobiose (K(D)=2.87 x 10(-8) M). Varying equilibrium dissociation constants in the micromolecular range were ascertained for chitotetraose (K(D)=4.5 x 10(-6) M), chitopentaose (K(D)=1.03 x 10(-6) M) and chitohexaose (K(D)=3.02 x 10(-6) M); the lowest value was measured for chitotriose (K(D)=19.4 x 10(-6) M). After having determined the sequences of several internal peptides from the binding protein by Edman degradation, the corresponding ngcE gene, which encodes a predicted lipid-anchored protein, was identified by reverse genetics. Using a genomic phage library of S. olivaceoviridis genes encoding two other membrane proteins (named NgcF and NgcG) were identified adjacent to ngcE. Each of these is predicted to have six membrane spanning helices and a consensus motif for integral membrane proteins characteristic of ABC transporters. In addition, the gene for a predicted regulator protein (NgcR) was detected. The ngcEFG operon lacks a gene for an ATP hydrolysing protein. NgcE is a new member of the CUT-1 family of ABC transporters for carbohydrates. Comparative studies of the wild-type and a mutant strain carrying an insertion within the ngc operon clearly demonstrate that the Ngc system mediates the uptake of N-acetylglucosamine and chitobiose in vivo. PMID- 12111552 TI - Physical interaction of Cdc28 with Cdc37 in Saccharomyces cerevisiae. AB - The Cdc37 protein in Saccharomyces cerevisiae is thought to be a kinase-targeting subunit of the chaperone Hsp90. In a genetic screen, four protein kinases were identified as interacting with Cdc37 - Cdc5, Cdc7, Cdc15 and Cak1. This result underlines the importance of Cdc37 for the folding of protein kinases. In addition, we showed that Ydj1, a yeast DnaJ homolog belonging to the Hsp40 family of chaperones, genetically interacts with Cdc37. No physical interaction has so far been detected between Cdc37 and Cdc28, although genetic interactions (synthetic lethality and mutation suppression), and biochemical studies have suggested that these two proteins functionally interact. We found that, when separately expressed, the N-terminal lobe of Cdc28 interacted strongly with the C terminal moiety of Cdc37 in a two-hybrid system. This was not the case for the full-length Cdc28 protein. We present models to explain these results. PMID- 12111551 TI - Direct interaction of the extracellular matrix protein DANCE with apolipoprotein(a) mediated by the kringle IV-type 2 domain. AB - Lipoprotein(a) [Lp(a)] consists of LDL and apolipoprotein(a), and has been shown to be a major, independent, risk factor for arteriosclerosis and thrombosis in humans. To further elucidate the (patho)physiological function(s) of Lp(a)/apo(a), we searched for new protein ligands, using the yeast two-hybrid system and employing the highly repetitive kringle IV type 2 (KIV-2) sequence from apo(a) as bait. The extracellular matrix protein DANCE [developmental arteries and neural crest epidermal growth factor (EGF)-like] recently implicated in atherogenesis was identified as an interactor. A direct physical interaction between DANCE and apo(a) was confirmed by co-purification of both recombinant proteins derived from culture supernatants of transiently transfected COS-1 cells. Furthermore, binding of human plasma-derived Lp(a) to recombinant DANCE was also observed. Finally, when monoclonal anti-apo(a) and polyclonal anti-DANCE antibodies were applied to tissue slices of atherosclerotic carotid artery, the two proteins were found to be co-localized in endothelial and smooth muscle cells, suggesting that they occur together in the arterial wall. However, as yet, the in vivo relevance and the possible functional role of this newly identified DANCE:Lp(a)/apo(a) interaction remains speculative. PMID- 12111553 TI - Genome-wide distribution and potential regulatory functions of AtATE, a novel family of miniature inverted-repeat transposable elements in Arabidopsis thaliana. AB - A study of transgenic promoter::beta-glucuronidase lines showed that the promoters of the two Arabidopsis ARGININE DECARBOXYLASE paralogues, ADC1 and ADC2, exhibited extremely different patterns of activity. One major feature of the promoter of ADC1 was the presence of a novel transposable element, which was shown to possess all of the characteristics of Miniature Inverted-repeat Transposable Elements (MITEs), and to be present in 26 full-length copies and 1617 partial copies and fragments distributed throughout the Arabidopsis genome. TRANSFAC analysis showed that this transposable element possesses a significant number of transcription-factor binding motifs. A bioinformatics approach based on a suffix-tree compilation was used to obtain an exhaustive description of exact copy numbers and positions of the element in the Arabidopsis genome. The distribution among the chromosomes was non-random, and a significant number of copies were found in regions flanking genes. Full-length copies of the transposable element were detected in the immediate vicinity of 22 genes, either upstream or downstream. PMID- 12111554 TI - Two broad-spectrum blast resistance genes, Pi9( t) and Pi2( t), are physically linked on rice chromosome 6. AB - To understand the molecular basis of broad-spectrum resistance to rice blast, fine-scale mapping of the two blast resistance (R) genes, Pi9( t) and Pi2( t), was conducted. These two genes were introgressed from different resistance donors, previously reported to confer resistance to many blast isolates in the Philippines, and were mapped to an approximately 10-cM interval on chromosome 6. To further test their resistance spectrum, 43 blast isolates collected from 13 countries were used to inoculate the Pi2( t) and Pi9( t) plants. Pi9( t)-bearing lines were highly resistant to all isolates tested, and lines carrying Pi2( t) were resistant to 36 isolates, confirming the broad-spectrum resistance of these two genes to diverse blast isolates. Three RAPD markers tightly linked to Pi9( t) were identified using the bulk segregant analysis technique. Twelve positive bacterial artificial chromosome (BAC) clones were identified and a BAC contig covering about 100 kb was constructed when the Pi9( t) BAC library was screened with one of the markers. A high-resolution map of Pi9( t) was constructed using BAC ends. The Pi2( t) gene was tightly linked to all of the Pi9( t) markers in 450 F(2) plants. These data suggest that Pi9( t) and Pi2( t) are either allelic or tightly linked in an approximately 100-kb region. The mapping results for Pi9( t) and Pi2( t) provide essential information for the positional cloning of these two important blast resistance genes in rice. PMID- 12111555 TI - A group of deuterostome Ty3/ gypsy-like retrotransposons with Ty1/ copia-like pol domain orders. AB - Here we report the existence of an unusual group of LTR retrotransposons, termed Gmr1-like elements. The members of this group are most similar in sequence to elements of the Ty3/ gypsy group, yet, unlike typical Ty3/ gypsy elements, they have pol genes in which the integrase domain lies upstream, rather than downstream, of the reverse transcriptase domain. Such an arrangement was formerly believed to be a characteristic peculiar to Ty1/ copia elements. The group includes the previously described retrotransposon Gmr1 from the Atlantic cod Gadus morhua, together with elements from a variety of other vertebrates, such as the zebrafish Danio rerio, the pufferfish Fugu rubripes and the clawed toad Xenopus laevis, as well as elements from non-vertebrate deuterostomes such as the sea squirt Ciona intestinalis. No Gmr1-like elements were found outside of the deuterostomes, nor were any other groups of elements with atypical pol-domain orders identified. Phylogenetic analyses show that the Gmr1-like elements form a monophyletic group within the larger group of Ty3/ gypsy elements. Some of the newly identified elements appear to be structurally intact and may still be active. The identification of this group challenges some previously held beliefs concerning the structure and evolution of LTR retrotransposons. PMID- 12111556 TI - The genes encoding subunits of ATP synthase are conserved in the reduced plastid genome of the heterotrophic alga Prototheca wickerhamii. AB - The heterotrophic unicellular alga Prototheca wickerhamii is closely related to the photoautotrophic Chlorella vulgaris but has a 54,100-bp plastid DNA (ptDNA) that is much smaller than the chloroplast DNA of C. vulgaris (150,613 bp). The nucleotide sequence of 28,093 bp of the Prototheca ptDNA has been determined. No genes for photosynthetic functions have been found, except for sequences encoding six subunits of the ATP synthase ( atpA, atpB, atpE, atpF, atpH, and atpI). Transcripts of these atp genes have also been detected. Whether the leucoplasts of Prototheca contain a functional ATP synthase has still to be elucidated. Identified genes further include tufA, minD, cysT, and genes coding for three rRNAs, 22 tRNAs, and 12 ribosomal proteins. The results support the idea that, in the reduced plastid genome of Prototheca, genes coding for components of the plastid translational apparatus have been preferentially retained, and might be needed for the expression of the atp genes and some unassigned ORFs. PMID- 12111557 TI - A regulator gene for acetate utilisation from Neurospora crassa. AB - The Neurospora crassa homologue of the Aspergillus nidulans regulatory gene facB has been cloned. The gene encodes a putative transcriptional activator of 865 amino acids that contains a DNA-binding domain with a Zn(II)(2)Cys(6) binuclear cluster, a linker region and a leucine zipper-like heptad repeat. Two internal amino acid sequences are identical to peptide sequences determined from proteolytic fragments of a DNA-binding protein complex specific for genes involved in acetate utilisation and expressed in acetate-induced mycelia of N. crassa. Recombinant expression of the predicted DNA-binding domain demonstrates that it is capable of independent recognition of a subset of the promoter sequences that bind the protein complex from N. crassa. A duplication-induced mutation in the corresponding gene results in an acetate non-utilising phenotype that is characterised by inefficient induction of the enzymes required for acetate utilisation. The new gene does not fall into any existing complementation group and has been designated acu-15. PMID- 12111558 TI - A receptor-like protein kinase with a lectin-like domain from lombardy poplar: gene expression in response to wounding and characterization of phosphorylation activity. AB - Plant receptor-like protein kinases (RLKs) are thought to be involved in various cellular processes mediated via signal transduction pathways. To clarify the initial step in such a signal transduction pathway in woody plants, we cloned a cDNA encoding PnLPK (a P opulus n igra var. italica lectin-like protein kinase) from lombardy poplar. The C-terminal region of the predicted PnLPK protein includes a protein kinase catalytic domain consisting of the 12 subdomains typical of the eukaryotic protein kinase superfamily. Following the signal peptide at the N-terminus, a domain that shows homology to legume lectins retains the putative Mn(2+)- and Ca(2+)-binding amino acids, which are highly conserved among lectin-related proteins. Because a putative hydrophobic transmembrane domain was localized between the lectin-like domain and the protein kinase domain, PnLPK was determined to be a member of the plant RLK subfamily with a lectin-like domain. Transcripts of the PnLPK gene accumulate in roots, mature leaves and calli of lombardy poplar, whereas only trace amounts of the transcripts are detectable in stems, young leaves and apical buds. Wounding of the young leaves increased the amount of PnLPK mRNA, but none of several phytohormones tested had any effect on the transcription of PnLPK. When incubated in the presence of divalent metal cations such as Mn(2+), the C-terminal catalytic domain of PnLPK showed significantly higher autophosphorylation activity than the full-length PnLPK protein. The phosphorylation activity of PnLPK was also detected using beta-casein as substrate. Phosphoamino acid analysis indicated that PnLPK is a serine/threonine kinase. PMID- 12111559 TI - Expression analysis of RNA14, a gene involved in mRNA 3' end maturation in yeast: characterization of the rna14-5 mutant strain. AB - In Saccharomyces cerevisiae, Rna14 protein is involved in both cleavage and polyadenylation of mRNA in the nucleus. Previous work has demonstrated that this protein is also localized in mitochondria. Moreover, all known rna14 mutants can be separated into two distinct classes: the poly(A)-negative class, which contains mutants that are deficient in mRNA 3'-end processing, and the poly(A) positive class, which includes those mutants that are not impaired in any of the steps in mRNA metabolism investigated. This suggests that in addition to its involvement in mRNA polyadenylation, Rna14p could have a second function related to mitochondrial metabolism. Here we investigated the regulation of RNA14 by characterizing the rna14-5 mutant, which is the only poly(A)-positive allele that also overproduces the RNA14 mRNA. We showed that both deregulation of RNA14 transcription and modification of RNA14 mRNA stability contribute to the strong accumulation of the transcripts in this mutant. Surprisingly, the RNA14 promoter itself is not essential for this phenotype of the rna14-5 mutant. However, the 3' UTR of the mRNA is necessary for overproduction of the transcripts, although it is not sufficient to deregulate a reporter gene by itself. Site-directed mutagenesis experiments provided additional data suggesting that the rna14-5 mutation acts at the protein level rather than modifying the properties of the RNA14 transcripts themselves. A tentative model accounting for the data is discussed, in light of the proposed extranuclear function of the Rna14p and its mitochondrial localization. PMID- 12111560 TI - Identification and genomic organization of gene loci negatively controlled by the virulence regulatory BvgAS two-component system in Bordetella bronchiseptica. AB - The two-component system BvgAS positively controls transcription of the virulence genes of Bordetella pertussis and B. bronchiseptica, which include several genes for toxins and adhesins. On the other hand, the BvgAS system negatively controls the expression of a poorly characterized set of genes, the so-called virulence repressed ( vrg) genes. To investigate the function of this group of genes and their relationship to virulence, we identified several novel vrg genes of B. bronchiseptica via the generation of transcriptional fusions with gfp (the ORF encoding Green Fluorescent Protein) by transposon mutagenesis. Expression of all of the vrg genes was enhanced under phenotype-modulating growth conditions and in phase variants, demonstrating that their transcription is indeed controlled by the BvgAS system. In addition, transcription of most of these new vrg genes was found to be affected by the growth phase of the bacteria, with maximal expression being observed in the late logarithmic or stationary phase. The majority of these genes encode putative metabolic functions involved in redox reactions and amino acid transport. Interestingly, several vrg genes of B. bronchiseptica are not expressed or have been lost in B. pertussis, indicating that, possibly as a consequence of its adaptation to a single host organism, many vrg genes of B. pertussis are gradually decaying. PMID- 12111561 TI - The stress-response sigma factor sigma(H) controls the expression of ssgB, a homologue of the sporulation-specific cell division gene ssgA, in Streptomyces coelicolor A3(2). AB - By using a previously established method for the identification of promoters recognized by a particular sigma factor of RNA polymerase, we identified a promoter in Streptomyces coelicolor A3(2) that is recognized by a heterologous RNA polymerase containing the late sporulation-specific sigma factor sigma(F). The promoter directed the expression of a gene named ssgB, which is related to the sporulation-specific cell division gene ssgA. These genes, together with three others, constitute a new family of paralogous genes specific for Streptomyces. S1-nuclease mapping using RNA prepared from an Escherichia coli strain that expresses ssgB under the control of sigma(F), and from S. coelicolor A3(2) at various developmental stages, identified identical transcription start points in both strains, corresponding to the promoter ssgBp. The promoter is developmentally regulated in S. coelicolor: it is induced at the time of aerial mycelium formation and is most active during sporulation. However, the level of the ssgB transcript was unaffected in a sigF mutant of S. coelicolor A3(2). Interestingly, the level of the transcript was substantially reduced in an S. coelicolor strain that was mutant for the sigH gene, which encodes a stress response sigma factor (sigma(H)) that is essential for sporulation in S. coelicolor A3(2). This dependence of ssgB upon sigH was confirmed by an in vitro transcription assay, in which sigma(H), in the presence of the S. coelicolor core RNA polymerase, was able to recognize ssgBp. These results suggest that the S. coelicolor ssgB gene is under the control of the stress-response sigma(H). Transcription of ssgB was investigated in S. coelicolor A3(2) mutants with lesions in each of six known early whi genes required for sporulation septation. Expression of ssgB was deregulated in three of the mutants ( whiA, whiI, and whiJ). Based on these data, it is proposed that the ssgB gene product plays a role in the developmental process in S. coelicolor A3(2). PMID- 12111562 TI - Regeneration of transgenic citrus plants under non selective conditions results in high-frequency recovery of plants with silenced transgenes. AB - Insertion of foreign DNA into plant genomes frequently results in the recovery of transgenic plants with silenced transgenes. To investigate to what extent regeneration under selective conditions limits the recovery of transgenic plants showing gene silencing in woody species, Mexican lime [ Citrus aurantifolia (Christm.) Swing.] plants were transformed with the p25 coat protein gene of Citrus tristeza virus (CTV) with or without selection for nptII and uidA. Strikingly, more than 30% of the transgenic limes regenerated under non-selective conditions had silenced transgenes, and in all cases silencing affected all the three transgenes incorporated. These results indicate that the frequency of transgene silencing may be greatly underestimated when the rate of silencing is estimated from the number of regenerants obtained under selective conditions. To our knowledge, this is the first report in which the frequency of gene silencing after transformation has been quantified. When the integration pattern of T-DNA was analyzed in silenced and non-silenced lines, it was observed that inverted repeats as well as direct repeats and even single integrations were able to trigger gene silencing. Gene silencing has often been associated with the insertion of DNA sequences as inverted repeats. Interestingly, here, direct repeats and single-copy insertions were found in both silenced and non-silenced lines, suggesting that the presence of inverted-repeat T-DNAs and the subsequent formation of dsRNAs triggering gene silencing cannot account for all silencing events. PMID- 12111563 TI - The magic bullet in the new millennium -- cause for optimism. PMID- 12111564 TI - The history of antibiotics: the Japanese story. AB - It is well-known that Japan was the third country, after the United States and the United Kingdom, to become self-sufficient in penicillin manufacture, as early as 1948. Besides the production of penicillin, much effort was made nationwide in exploratory research on anti-infective, anti-cancer, and agricultural antibiotics, and also other bioactive microbial products. One can list upwards of 117 useful antibiotics and related agents of Japanese origin, and among them, 41 agents have been licensed out around the world. The first antibiotic from Japan was colistin (discovered in 1950) followed by well-known agents such as mitomycin C (1955), kanamycin (1957), bleomycin (1965), cefazolin (1969), amikacin (1972), piperacillin (1976), norfloxacin (1977), cefoperazone (1978), ofloxacin (1980), clarithromycin (1984), meropenem (1987), and others. The major group is the beta lactam antibiotics (up to 15), followed by 7 fluoroquinolones and anti-cancer and agricultural antibiotics (4 of each). Noteworthy was the expansion of antibiotics research into bioactive microbial products, which led to the discovery of extremely beneficial agents for human quality of life (QOL), such as pravastatin for hyperlipidemia and tacrolimus for atopic diseases. It is to be expected that Japan will continue a high level of activity in exploratory research on beneficial chemotherapeutic agents in the twenty-first century. The Japanese Society of Chemotherapy (JSC), established in 1953, has played a central role for about a half century in the evaluation of novel agents discovered in Japan and those introduced from abroad. Besides the evaluation of chemotherapeutic agents, both fundamentally and clinically, the JSC has taken an active part in the establishment of a variety of guidelines for chemotherapy and in the education of clinical scientists who have evaluated all agents of Japanese origin appropriately. PMID- 12111565 TI - Trends in antimicrobial susceptibility of the Streptococcus milleri group. AB - A collection of 114 clinical Streptococcus milleri group (SMG) strains at the Ryukyu University Hospital obtained in 1999 and 2000, was identified and tested for susceptibility to 12 antibiotics. The percentage of strains with intermediate susceptibility to penicillin G was relatively high (14%). Cefaclor and cefotiam, with a MIC 90 of 2 microg/ml, were less active than cefotaxime. Strains nonsusceptible to erythromycin, clindamycin, and azithromycin were found with a frequency of 8%, 5%, and 4%, respectively. Almost all of the SMG strains were susceptible to fluoroquinolones (except for 1% of the strains nonsusceptible to levofloxacin), and sitafloxacin (DU-6859a) was the most active agent among the 12 tested antibiotics. PMID- 12111567 TI - Rapid detection of quinolone resistance-associated gyrA mutations in Neisseria gonorrhoeae with a LightCycler. AB - Afluorometric real-time polymerase chain-reaction (PCR)-hybridization system, the LightCycler was developed for the detection of Neisseria gonorrhoeae in clinical samples and the analysis of point mutations associated with quinolone resistance in the gyrAgene. This system allowed us to amplify the N. gonorrhoeae-specific gyrA gene from an amount of DNA corresponding to five genome copies per reaction. We were able to detect N. gonorrhoeae in either 55 control strains or 36 nonisolated clinical urethral swab specimens, and to analyze the presence of mutations in codons Ser-91 and Asp-95 of the gyrA gene within 1 h. The mutation status in the gyrA gene assessed by the LightCycler assay was completely in agreement with the results of our previous conventional sequencing analysis, and was associated with the susceptibility to ciprofloxacin. PMID- 12111566 TI - CP6679, a new injectable cephalosporin with broad spectrum and potent activities against methicillin-resistant Staphylococcus aureus and Pseudomonas aeruginosa. AB - The antibacterial activities of CP6679, a new injectable cephalosporin with a broad antibacterial spectrum, were compared with those of other cephalosporins. CP6679 had stronger in-vitro activity than ceftazidime and cefpirome against methicillin-resistant Staphylococcus aureus(MRSA), methicillin-resistant coagulase-negative staphylococci, and Pseudomonas aeruginosa. Its activity against MRSA was eight times stronger than that of cefpirome, and it showed high binding affinity for penicillin-binding protein 2' of MRSA. Furthermore, the antibacterial activity of CP6679 against ceftazidime-resistant and imipenem resistant P. aeruginosawas eight times stronger than that of ceftazidime and four times stronger than that of imipenem. In addition to its in-vitro activities, CP6679 showed the highest efficacy among all cephalosporins tested in murine models of systemic infection induced by MRSA or P. aeruginosa. It was more effective than vancomycin and cefpirome against respiratory tract infections induced by MRSA in mice. PMID- 12111568 TI - Evaluation of panipenem/betamipron (PAPM/BP) in pneumonia in elderly patients. AB - The present multicenter study evaluated the clinical and bacteriological efficacy and safety of panipenem/betamipron (PAPM/BP) for treating pneumonia in elderly patients. Forty-three episodes of pneumonia in 43 patients were treated with PAPM/BP as the sole antibiotic agent. All patients were 65 years of age or older, and were given PAPM/BP at a total daily dosage range of 0.5-2.0 g. The clinical efficacy rate, expressed as a percentage of the total number of excellent and good responses, was 56.4%. Of the 43 patients, 13 were evaluated bacteriologically. In these 13 patients, the eradication rate, expressed as a percentage of the total number of "eradicated" and "replaced" efficacies, was 30.8%. Adverse effects and abnormal laboratory findings occurred in 2 patients, which was 4.6% of the total number of patients evaluated. No serious adverse effects were observed. We concluded that PAPM/BP was well tolerated overall, and was effective and safe for most of the elderly patients. PMID- 12111569 TI - Characterization of cefotaxime-resistant Escherichia coli isolates from a nosocomial outbreak at three geriatric hospitals. AB - Over a 22-month period, there was an unusual upsurge in the incidence of cefotaxime-resistant Escherichia coli among hospitalized patients in three geriatric hospitals in the same district. Sixteen highly cefotaxime-resistant strains were obtained from clinical specimens during the period January 1996 through October 1997. All strains were characterized by antibiotic resistance pattern analysis, the detection of the TEM- and Toho-type beta-lactamase or CTX-M type beta-lactamase gene by polymerase chain reaction (PCR), plasmid profiling, Southern hybridization analysis, and pulsed-field gel electrophoresis (PFGE). Antibiotic resistance analysis showed that all strains were highly resistant to ampicillin, piperacillin, carbenicillin, cephaloridine, and cefotaxime; intermediately resistant to cefoxitin; moderately susceptible to moxalactam and ceftazidime; and susceptible to imipenem. Detailed analysis of beta-lactamase content revealed that all cefotaxime-resistant strains harbored a plasmid that mediated an extended-spectrum beta-lactamase of the Toho-type or CTX-M-type by PCR and Southern hybridization analysis. PCR detection showed that all the E. colistrains, except for strains TUM1023, TUM1101, TUM1227, and TUM1229, also possessed bla(TEM) genes. Furthermore, Southern hybridization analysis showed that all strains, except for TUM1102, gave a similar signal with the Toho probe. The PFGE profiles of the E. colistrains obtained with XbaI showed four patterns that correlated well with the plasmid profiles. The Dice value of 15 strains, including Toho-2 producer (TUM1083), for their PFGE patterns indicated a similarity of 80% or more. Our results suggest that 15 of the 17 Toho type beta lactamase-producing E. coli strains (including strain TUM1083) studied belong to a single epidemic strain, while the other two strains are different from them, and the Toho-type or CTX-M-type beta-lactamase encoding gene may be acquired by plasmid conjugation or a mobile element. PMID- 12111570 TI - Survey of dental hygienists and healthcare workers for microorganisms in the oral cavity. AB - Over 350 bacterial species are known to form communities on tooth surfaces in the oral cavity, and opportunistic pathogens are frequently isolated from samples taken from the oral cavities of immunocompromised hosts, as well as elderly people with poor oral hygiene. To confirm the correlation between the prevalence of opportunistic pathogens in the oral cavity and oral hygiene, dental plaque and saliva samples from 15 dental hygienists and 15 healthcare workers were investigated. No opportunistic pathogens were found in the samples from dental hygienists, however, they were isolated from 10 of the 15 healthcare workers (66.7%). The total amounts of Streptococcus sp. and Lactobacillus sp. were also significantly higher in the healthcare workers. The amount of Streptococcus mutans was also higher, although the difference was not significant. From our results, we concluded that the prevalence of opportunistic pathogens and cariogenic bacteria in the oral cavity may be reduced by proper oral hygiene measures. PMID- 12111571 TI - Molecular epidemiological studies of Staphylococcus aureus in urinary tract infection. AB - In recent years, the increasing incidence of urinary tract infection (UTIs) caused by Staphylococcus aureus has been noted at the urology ward, Okayama University Hospital. We investigated the molecular epidemiological characteristics of 139 UTI isolates, including 45 methicillin-sensitive S. aureus (MSSA) and 94 methicillin-resistant S. aureus (MRSA), collected over a 10-year period from 1990 to 1999. The antibiotic resistance genes ( mecA, aph(3')-III, aac(6')-aph(2"), ant(4')-I) and the toxin genes (tst, sea, seb, and sec) were detected by using multiplex polymerase chain reaction (PCR). Since 1996, the prevalence of the ant(4')-I, tstand secgenes has increased markedly in coagulase type II S. aureus possessing the mecA gene (MRSA). The presence of toxin genes in MRSA was higher than that in MSSA; 66.0% and 26.7% for tst, 7.4% and 4.4% for sea, 24.5% and 8.9% for seb, and 66.0% and 28.9% for sec, respectively. In the review of medical records, it was found that febrile episodes occurred in 12 of 72 patients with monomicrobial UTI caused by S. aureus. For the febrile patients, S. aureus isolates with both the tst and sec genes were found significantly more often (11 of 12; 91.7%) than those without the tst and sec genes ( P = 0.0484). Molecular typing of MRSA isolates, by using random amplified polymorphic DNA analysis and pulsed-field gel electrophoresis analysis, revealed no apparent clonality of these isolates over the 10 years, suggesting that most of the recent MRSA infections are not due to cross-infection in the urology ward. PMID- 12111572 TI - Acute disseminated encephalomyelitis following dengue fever. AB - A58-year-old man suffered from acute disseminated encephalomyelitis (ADEM) after dengue fever. ADEM has not been described as the cause of neurological complications in dengue fever. However, the increasing use of magnetic resonance imaging in endemic areas may help to identify ADEM as being responsible for neurological complications in dengue fever. PMID- 12111573 TI - Disseminated coccidioidomycosis with intra- and paravertebral abscesses. AB - We report a case of paravertebral and intravertebral abscesses caused by Coccidioides immitis in a Japanese man. The patient had lived in Arizona, United States, for 5 years, and suffered from overt disease after coming back to Japan. Culture of pus from the paravertebral abscess revealed Coccidioides immitis, and a diagnosis of disseminated coccidioidomycosis was made. Fluconazole (600 mg/day), taken orally, was started, and the abscesses surrounding the vertebral bodies disappeared after 2 years of treatment. The abscess in the vertebral bodies also responded to treatment, but a small lesion was still left in the 10th vertebral body after 2 years of treatment. Coccidioidomycosis is a fungal infection that is endemic in the southwestern United States and in Central and South America. Although coccidioidomycosis causes self-limiting flu-like illness or pneumonia, a small proportion of the infections progress to disseminated diseases. Because the incidence of coccidioidomycosis is increasing year by year, physicians not only in endemic but also in nonendemic areas have to consider coccidioidomycosis as one of the differential diagnoses when they examine patients from endemic areas. PMID- 12111574 TI - Pyogenic vertebral osteomyelitis caused by Prevotella intermedia. AB - We describe a case of vertebral osteomyelitis caused by Prevotella intermedia, which is an extremely unusual cause of vertebral osteomyelitis. The organism was isolated from vertebral biopsies and the patient was treated successfully with intravenous ampicillin-sulbactam and clindamycin. Diagnosis and management of this condition are described, and the importance of anaerobic bacteria in the pathogenesis of vertebral osteomyelitis is discussed. PMID- 12111576 TI - Susceptibility change to antibiotics of Staphylococcus aureus strains isolated from skin infections between July 1994 and November 2000. AB - We measured the in-vitro susceptibility of 833 Staphylococcus aureus strains, isolated from skin infections at our hospital between July 1994 and November 2000, to 13 antimicrobial agents (ampicillin, methicillin, cephalexin, cefaclor, cefpodoxime proxetil, gentamicin, erythromycin, clindamycin, minocycline, vancomycin, fusidic acid, ofloxacin, and nadifloxacin). The concentrations required to inhibit 50% of the strains (MIC(50)) of all these antimicrobial agents was extremely stable and ranged at levels below 3.13 microg/ml, except for gentamicin; in contrast, the MIC(90) was not uniform. The MIC(90) of vancomycin, fusidic acid, and nadifloxacin was very low and stable. No strain was resistant to vancomycin. The incidence of MRSA was 10%-20%. PMID- 12111577 TI - In-vitro activity of ciprofloxacin combined with flomoxef against Bacteroides fragilis, compared with that of ciprofloxacin combined with clindamycin. AB - Using checkerboard and time-kill assays, the in-vitro activity of ciprofloxacin alone and in combination with flomoxef against clinical Bacteroides fragilis strains was evaluated. In addition, the microbiological efficacy of this combination was compared with that of ciprofloxacin plus clindamycin. In 88% of the 25 strains tested, the combination of ciprofloxacin plus flomoxef exhibited a synergistic or an additive effect, whereas only 56% of the 25 strains ( P< 0.01, chi(2) test) tested with the combination of ciprofloxacin plus clindamycin exhibited similar effects. In a time-kill study using 7 clinical strains, a synergistic or additive effect of the combination of ciprofloxacin plus flomoxef was observed in all 7 strains. In conclusion, the combination of ciprofloxacin plus flomoxef is very active against B. fragilis, suggesting that this combination may be very useful in the treatment of aerobic and B. fragilis mixed infections, because ciprofloxacin has an expanded spectrum against aerobes. PMID- 12111575 TI - A rare case of Salmonella soft-tissue abscess. AB - A healthy 6-year-old boy had complained of fever and chest pain for 3 days. On admission, he had a mass on the sternum, 3.7 x 2.5 cm in size. Abnormal laboratory findings included a white blood cell count of 12,900/microl, erythrocyte sedimentation rate (ESR), 74 mm/h, and C-reactive protein (CRP), 9.7 mg/dl. Ultrasound examination of the chest revealed a hypoechoic lesion on the sternum that was 30 x 15 mm in size. Chest computed tomography (CT) scan showed no bone fracture or bone erosion. The patient received cefpirome, given intravenously at 60 mg/kg per day for 10 days. Incision and drainage was performed on the seventh day in the hospital, and we collected 0.5 ml of pus. Salmonella enteritidis was detected from the drainage. However, the patient had no gastrointestinal symptoms. He was discharged on the fourteenth hospital day, as he was asymptomatic. Results of all physical and laboratory examinations including blood and stool cultures and ultrasound examinations, were within the normal limits upon discharge. PMID- 12111578 TI - Effects of antimicrobial agents on spontaneous and endotoxin-induced cytokine release of human peripheral blood mononuclear cells. AB - Because the immunomodulatory effects of antibiotics could possibly influence the degree of the systemic and local response to infection, knowledge of their intrinsic influence on the host's inflammatory response appears to be essential. Therefore, this study investigated the effects of frequently used antimicrobial agents (beta-lactams, quinolones gentamicin, vancomycin and metronidazole) on the in-vitro tumor necrosis factor (TNF)-alpha and interleukin (IL)-6 production of isolated human peripheral blood mononuclear cells (PBMNC), cultured with or without endotoxin, in comparison with those effects obtained in a whole-blood assay system. In the presence of ciprofloxacin, ofloxacin, gentamicin, vancomycin, and metronidazole, a significant inhibition of the endotoxin stimulated TNF-alpha production of human peripheral blood mononuclear cells (PBMNC) was found at therapeutic levels. Only ofloxacin showed a significant inhibitory influence on the endotoxin-induced IL-6 production of PBMNC. In the whole-blood assay, significant effects were not detectable. None of the antibiotics showed cytotoxicity. It is concluded that, at present, the direct immunological effects of antibiotics should be interpreted carefully with regard to the experimental conditions, and regardless of the therapeutic implications. To assess the potential direct immunomodulatory effect of antimicrobial agents, different cell culture procedures should be used. PMID- 12111579 TI - Disseminated coccidioidomycosis with intra- and paravertebral abscesses. PMID- 12111580 TI - The prognostic significance of overexpression of the decoy receptor for Fas ligand (DcR3) in patients with gastric carcinomas. AB - BACKGROUND: The FasL-Fas system has an important role in mediating immune cytotoxic killing of cells such as virus-infected or tumor cells. It was recently reported that there is a soluble decoy receptor (DcR3), which binds to FasL and inhibits FasL-induced apoptosis, and certain tumors may escape FasL-dependent immune-cytotoxic attack by expressing a decoy receptor that blocks FasL. We evaluated whether DcR3 has clinical relevance in actual human gastric cancers. METHODS: : The expression of DcR3 was investigated by Northern blot analysis in a series of 84 primary gastric carcinomas and compared with clinicopathological features and prognosis. The DcR3 expression level was analyzed and quantified densitometrically. The location of DcR3 mRNA in gastric carcinoma tissue was detected by in situ hybridization. RESULTS: The frequency of DcR3 overexpression was 26% (22 of 84 surgical specimens). The DcR3 expression level was significantly associated with lymph node metastasis and pathological stage, but did not correlate with tumor size, metastatic status, or histological type. In situ hybridization demonstrated that DcR3 mRNA was expressed in tumor cells. When the patients were followed up for 63 months, DcR3 overexpression was found to be associated with a significantly shortened duration of overall survival compared with findings in patients having normal DcR3 expression. CONCLUSION: The DcR3 decoy receptor for FasL may be involved in the progression of gastric cancer. Further evaluation of these possible roles of DcR3 and the regulation of DcR3 expression in malignant cells will be critically important for the development of new strategies for controlling the growth of malignant cells that escape host immune surveillance. PMID- 12111581 TI - Quantitative detection of disseminated cancer cells in the greater omentum of gastric carcinoma patients with real-time RT-PCR: a comparison with peritoneal lavage cytology. AB - BACKGROUND: Peritoneal lavage cytology is an excellent prognostic determinant but lacks sensitivity. Quantification of carcinoembryonic antigen (CEA) mRNA in peritoneal washes by real-time reverse-transcriptase polymerase chain reaction (RT-PCR) was found to be a more sensitive method to detect free cancer cells. It may be beneficial to explore, by the same method, a sample of the omentum, which is known to harbor cancer cells before the establishment of gross peritoneal metastasis. METHODS: Greater omentum and peritoneal washes were obtained from 90 gastric carcinoma patients during laparotomy. The CEA mRNA levels in these materials were quantified using a real-time RT-PCR system with hybridization probes. The significance of CEA mRNA levels in both materials, as a predictive factor of peritoneal metastasis, was explored by univariate and multivariate analyses. RESULTS: After a median follow-up of 718 days, 13 patients had clinical evidence of peritoneal metastasis. Under the assumption that these patients had free cancer cells in the peritoneal cavity, the sensitivity and specificity of conventional cytology for the detection of these cells were 31% and 100%, respectively. The sensitivity and specificity of the CEA mRNA levels extracted from the peritoneal washing samples were 77% and 94%, while those of the omentum were 46% and 90%. Multivariate analysis, with the diagnosis of peritoneal metastasis as an endpoint, revealed that CEA mRNA level in the peritoneal washes was the only significant risk factor. CONCLUSION: Quantitative RT-PCR of peritoneal washes remains the first choice as a tool to sensitively predict intraperitoneal recurrence in gastric carcinoma patients. PMID- 12111582 TI - Reduced thiamine (vitamin B1) levels following gastrectomy for gastric cancer. AB - BACKGROUND: Vitamin B1 deficiency is well known as a possible complication following gastric restrictive surgery for morbid obesity; however, reduced vitamin B1 levels in patients who have undergone gastrectomy for gastric cancer have not been discussed previously. METHODS: Serum vitamin B1 levels were determined after the return to normal daily activity in 54 patients with distal gastrectomy for gastric cancer, 32 patients with total gastrectomy for gastric cancer, and 30 patients with radical surgery for colorectal cancer. Changes from serum vitamin B1 levels before operation to those after return to normal daily activity, without nutritional support, were investigated in 25 patients with gastrectomy for gastric cancer and 26 patients with radical surgery for colorectal cancer. RESULTS: Decreased serum vitamin B1 levels, below the normal range, were recognized in 7 of the 54 distally gastrectomized patients and in 5 of the 32 totally gastrectomized patients, whereas no such decrease was recognized in any patient after colorectal surgery. Decreased serum vitamin B1 level was recognized within 6 months after the operation in 6 of the 7 distally gastrectomized patients showing a decreased vitamin B1 level and in 3 of the 5 totally gastrectomized patients showing a decreased vitamin B1 level. Postoperative serum vitamin B1 levels were significantly lower than those before operation in patients with gastrectomies, whereas there was no significant difference in serum vitamin B1 levels before and after the surgeries in patients with surgery for colorectal cancer. CONCLUSION: Vitamin B1 levels may be reduced in gastrectomized patients, especially within 6 months after operation, even after their return to normal daily activity without nutritional support. PMID- 12111583 TI - Endoscopic evaluation of the remnant stomach after gastrectomy: proposal for a new classification. AB - BACKGROUND: In Japan, approximately half of gastric cancers are diagnosed in the early stage. Longer survival has been provided for patients with early gastric cancer (EGC). Several new surgical procedures have been employed for some EGCs. To compare the functional results of these techniques with those of classic distal gastrectomy, it is important to evaluate the remnant stomach in relation to quality of life (QOL) and secondary cancers. We propose a new endoscopic classification, regarding several aspects of the remnant stomach, which enables common understanding and description of the condition. METHODS: Of 651 patients who underwent a distal gastrectomy or pylorus-preserving gastrectomy (PPG), 324 had at least one upper gastrointestinal (GI) endoscopy during the follow-up period. Ninety-three of the 324 patients underwent a Roux-en-Y reconstruction (RY); 175, Billroth type 1 (B1); and 56, PPG. Endoscopic findings regarding residual food, gastritis, and bile reflux in the gastric stump were investigated for these patients. We classified the amount of residual food into five grades, the degree and the extent of gastritis into five grades, and bile reflux into two grades. First, we evaluated the consistency of diagnosis between two endoscopists, in the first 200 patients, and then we applied the classification to all 324 patients to examine the usefulness of this classification. RESULTS: Consistency of diagnosis was obtained between two endoscopists who classified the patients independently. The agreement rate was 98.5% for residual food, 93% for gastritis, and 100% for bile reflux. Residual food was observed in 14.0% of the RY group, 22.3% of the B1 group, and 37.5% of the PPG group. These differences were significant (RY versus B1; P < 0.05 and RY versus PPG; P < 0.01). The remnant stomach after B1 showed significantly more severe and extensive gastritis than that after RY and PPG ( P < 0.01). As for bile reflux, there was no significant difference among the three groups. CONCLUSION: The classification (RGB classification: Residue, Gastritis, Bile) can be used easily and is practical. The results suggest some important differences among methods of reconstruction. This classification seems to be useful to describe these findings and to further evaluate these reconstructive methods. PMID- 12111584 TI - Phase II study of paclitaxel with 3-h infusion in patients with advanced gastric cancer. AB - BACKGROUND: To increase the options for agents for gastric cancer chemotherapy, we performed a phase II clinical trial on the use of a 3-h infusion of paclitaxel to confirm its efficacy and the feasibility of its use in patients with advanced gastric cancer. METHODS: Thirty-two (32) patients with measurable metastatic gastric cancer were enrolled in this study. Seventeen patients (53%) had received prior chemotherapy for metastatic disease, 4 patients (13%) had adjuvant chemotherapy alone, and 11 patients (34%) were chemotherapy-naive. Paclitaxel was intravenously infused for 3 h, at a dose of 210 mg/m(2), once every 3 weeks. To prevent hypersensitivity reactions, standard premedication was administered to all patients. RESULTS: Nine (28%; 9/32 ) objective partial responses (PRs) were observed (95% confidence interval [CI], 14%-47%), and the remaining 23 patients showed stable (12 patients; 37.5%) and progressive disease (11 patients; 34.4%). The median time to response was 20 days (range, 14-38 days). The median response duration was 87 days (range, 50-103 days). The median survival of all patients was 234 days (range, 13-646+ days). The major adverse reactions were myelosuppression (grade 3/4 leukopenia and neutropenia were observed in 59% and 88% of the patients, respectively), alopecia, and peripheral neuropathy. Peripheral neuropathy was observed in 19 patients, however, most of the patients recovered after the completion of treatment. CONCLUSION: A 3-h infusion of paclitaxel is an effective therapy for advanced gastric cancer and is clinically well tolerated by the patients. PMID- 12111585 TI - An oral anticancer drug, TS-1, enabled a patient with advanced gastric cancer with Virchow's metastasis to receive curative resection. AB - We encountered a patient with advanced gastric cancer, with Virchow's lymph node metastasis, who subsequently underwent curative resection after neoadjuvant chemotherapy with the newly developed oral anticancer drug, TS-1. The patient was a 67-year-old woman who had a type 2 tumor in the middle third of the stomach, and Virchow's lymph node metastasis, which was diagnosed by fine-needle aspiration cytology; she also had swollen paraaortic lymph nodes. Curative resection was considered impossible, and TS-1 (100 mg/day) was administered for 28 days in one course, mainly in the outpatient clinic. Although grade 2 stomatitis interrupted the therapy on day 21 of the second course and on day 7 of the third course, the type 2 tumor showed marked remission (partial response; PR) and the metastasis in the Virchow's and paraaortic lymph nodes had completely disappeared after the third course (complete response; CR). Eleven weeks after the completion of the TS-1 treatment, total gastric resection with D3 lymph node dissection was performed. Histopathological examination revealed tumor involvement only in the mucosal and submucosal layers of the stomach and the no. 4d lymph node. Most of the tumor was replaced with fibrosis with granulomatous change in the muscularis propria of the stomach and in the no. 3, no. 6, and no. 7 lymph nodes. This may be the first report of a patient with advanced gastric cancer with Virchow's lymph node metastasis who successfully received curative resection following neoadjuvant chemotherapy with a single oral anticancer drug. PMID- 12111587 TI - Intramuscular metastasis from gastric cancer. AB - Skeletal muscle is an uncommon site of hematogenous metastasis of gastric carcinoma. We report here a rare case of gastric carcinoma with multiple intramuscular metastases. Our patient had advanced gastric carcinoma and complained of left gluteal induration with tenderness. Because magnetic resonance imaging (MRI) revealed that the gluteal tumor showed iso-signal intensity on T1 weighted images and high signal intensity on T2-weighted images, with reticulated texture around the tumor, and the patient had advanced gastric carcinoma, we speculated that the tumor was an intramuscular metastatic tumor from primary gastric carcinoma. There were also multiple intramuscular metastatic lesions in both gluteal muscles on the MRI findings that were not detected by physical examination. Therefore, the patient underwent total gastrectomy with combined resection of spleen, with subsequent chemotherapy. Three months after the operation, we excised the gluteal tumor to alleviate the gluteal pain. Histological examinations confirmed that the gluteal tumor was a metastasis from primary gastric carcinoma. PMID- 12111586 TI - Rare case of early mucosal gastric cancer presenting with metastasis to the bulbar conjunctiva. AB - Early gastric cancer has an excellent outcome following surgical treatment. In particular, mucosal gastric cancer (m-cancer) very rarely results in metastatic dissemination and may be successfully treated by local surgical resection. We report a 64-year-old Japanese woman who presented with a recurrent cystic lesion on the left bulbar conjunctiva, with a biopsy specimen revealing metastatic signet-ring cell carcinoma. Gastrointestinal investigations revealed an early gastric cancer in the lesser curvature of the stomach. Biopsy specimen of the gastric lesion indicated poorly differentiated adenocarcinoma, which was identical to findings in the lesion in the bulbar conjunctiva. She underwent a distal gastrectomy and made an uneventful recovery. Histopathological analysis indicated a gastric signet-ring cell carcinoma that was limited to the mucosal layer, without evidence of lymphatic spread. Although the exact mechanism of metastatic dissemination to the bulbar conjunctiva is unclear, this case is very unusual, because ocular metastases almost invariably occur in the context of documented and established malignant disease. This is, to our knowledge, the first reported case of a patient with gastric mucosal cancer who presented with a conjunctival metastatic deposit and who subsequently received curative surgical treatment for both conditions. PMID- 12111588 TI - A patient with primary gastric choriocarcinoma who received a correct preoperative diagnosis and achieved prolonged survival. AB - Primary choriocarcinoma of the stomach is an extremely rare and highly malignant tumor. A 68-year-old Japanese man was admitted to our department with symptoms of epigastric pain, abdominal fullness, and dizziness. Primary gastric choriocarcinoma with regional lymph node metastases was diagnosed preoperatively by the detection of elevated serum levels of a tumor marker, radiography, and immunohistochemical staining of biopsy specimens. The patient underwent total gastrectomy and jejunal reconstruction (Roux-en-Y method), followed by chemotherapy. Histological examination of the resected stomach revealed typical choriocarcinoma accompanied by common adenocarcinomatous elements. After postoperative chemotherapy the patient survived for 4 years and 6 months, and died with no evidence of recurrence of carcinoma. This is the first known patient with primary gastric choriocarcinoma to have survived for such a long period without recurrent elevation of the serum level of human chorionic gonadotropin (HCG), which was a useful marker when reevaluating the patient. PMID- 12111589 TI - Correlations between light penetration into skin and the therapeutic outcome following laser therapy of port-wine stains. AB - For several years the flashlamp-pumped pulsed dye laser (FPDL) has been the favoured method for the treatment of port-wine stains (PWS). The therapeutic outcome of FPDL laser therapy depends on the anatomical location of the PWS and is mainly attributed to morphological parameters such as size and depth of the PWS blood vessels. The aim of this study was to show a correlation between the therapeutic outcome following FPDL therapy and the optical properties of the skin overlying the PWS vessels. For this purpose the therapeutic outcome following FPDL treatment (585 nm; 0.45 ms) of 884 PWS situated on different body sites was evaluated by judging the grade of fading of PWS colour. On the other hand the light penetration into 123 skin samples (thickness 0.10-1.35 mm) was determined between 450 nm and 1030 nm and compared with the PWS laser therapy outcome for equal locations by statistical analysis. PWS on the neck, trunk, arms or legs yielded a higher mean grade of fading as compared to PWS on the head. Within the face, a wide range of fading was evident. The light penetration into skin increased linearly with increasing wavelength and location-dependent differences were found. The attenuation coefficient was 22.8+/-5.3 mm(-1) at 585 nm. No significant or strong correlation was observed between the therapeutic outcome of PWS laser therapy and the light penetration into skin. However, a correlation was obvious by plotting the respective profile plots. Therefore, among other effects, in particular morphological parameters of PWS vessels, the optical properties of the skin contribute to a small extent to the clinical outcome of PWS laser therapy. PMID- 12111590 TI - Tissue levels, histologic changes and plasma pharmacokinetics of meta-Tetra (hydroxyphenyl) chlorin (mTHPC) in the cat. AB - The biodistribution and pharmacokinetics of meta-tetra(hydroxyphenyl)chlorin (mTHPC)(1) have been documented in humans, rats, dogs and rabbits. It has been demonstrated to be an effective photodynamic therapy agent for treatment of squamous cell carcinoma. Squamous cell carcinoma is a common feline neoplasm, causing significant morbidity and mortality in the feline population. The association between ultraviolet radiation exposure and occurrence of this neoplasm in the cat provides a useful model for the study of human cutaneous squamous cell carcinoma. In this study, we document the biodistribution, pharmacokinetics and toxicity of mTHPC in a group of normal cats. Four groups of cats were given the drug intravenously at dosages of 0, 0.15, 0.30 and 0.60 mg/kg. mTHPC levels were measured in plasma and tissues at 0, 24, 48, 72, 96 and 336 h after drug administration. Additionally, plasma samples were collected at 1 and 6 h post-injection and analysed. Biodistribution and pharmacokinetics of mTHPC in cats mirrors that in other animal species. There were no clinical or pathological changes associated with administration of the drug. The biodistribution and pharmacokinetics of mTHPC in cats mirrors that in other species studied. There were no clinical or pathological changes attributable to administration of the drug at the doses administered. mTHPC may be a useful photodynamic therapy drug in cats. PMID- 12111591 TI - Fluorescence biodistribution and photosensitising activity of toluidine blue o on rat buccal mucosa. AB - The antimicrobial activity of toluidine blue O (TBO) in the presence of red light has been demonstrated for a wide range of microorganisms. The response of tissues to TBO-induced photosensitisation is an important factor in assessing the clinical usefulness of this technique for the treatment of infectious diseases. The aims of this study were to determine the effect of TBO-mediated photosensitisation on rat buccal mucosa and the biodistribution of the photosensitiser in this tissue. An aqueous solution of TBO was applied to one side of the buccal mucosa of the animals. A 6 mm diameter area was then exposed to light (633 nm) from a copper vapour pumped-dye laser. The opposite, untreated, side of the buccal mucosa served as a control. TBO concentrations of 25, 50 and 200 microg/ml, laser light doses of 110, 170 and 340 J/cm(2) were assessed. Control groups of animals were subjected to 340 J/cm(2) laser light alone or to 200 microg/ml TBO alone. Serial sacrifices were performed after 72 h to obtain mucosal tissue samples for histological examination. For the determination of TBO biodistribution, additional groups received the same TBO doses and were sacrificed after 1 min or 10 min. Specimens were removed and frozen immediately for digital fluorescence imaging. No necrotic or inflammatory changes were found in the buccal mucosa of the animals with any of the treatments (using up to 200 microg/ml TBO and 340 J/cm(2) laser light). A high TBO fluorescence in the epithelium, particularly in the keratinised layer, with almost no fluorescence in the underlying connective tissue was demonstrated by the digital imaging. The results of this study suggest that TBO-mediated PDT (within the concentrations and light doses tested) could be a safe antimicrobial approach for the oral infections without damaging the adjacent normal tissue. PMID- 12111592 TI - Microleakage of class V composite restorations following Er:YAG and Nd:YAP laser irradiation compared to acid-etch: an In vitro study. AB - This study compared microleakage at enamel/composite and dentine/composite interfaces following Er:YAG laser, Nd:YAP laser, or acid-etch preparation. Class V cavities produced on the lingual and buccal surfaces of 20 extracted carie- and restoration-free human teeth were randomly assigned to four groups of 10 cavities. Teeth were treated with 37% phosphoric acid and primer (group 1), irradiated with an Er:YAG laser (group 2) or an Nd:YAP laser (group 3), or served as controls (group 4). The specimens were restored with Scotchbond Multipurpose/Z100 (3M), stored in physiological solution at 37 degrees C for 7 days, thermocycled 500 times between 5 degrees C and 55 degrees C, placed in a 0.5% solution of basic fuchsin for 48 h, embedded in resin, and sectioned. Microleakage was assessed according to the depth of dye penetration along the restorative composite. The results showed that irradiation with Er:YAG and Nd:YAP lasers did not produce a good seal. Mean microleakage was greater than with acid etch and statistically comparable to that of control cavities (Kruskal-Wallis test). PMID- 12111593 TI - Optimisation of illumination for photodynamic therapy with mTHPC on normal colon and a transplantable tumour in rats. AB - Recent reports suggest that the effect of photodynamic therapy (PDT) can be enhanced by fractionating the light dose or reducing the light fluence rate. We assessed these options on two tissues in rats (normal colon and a transplanted fibrosarcoma) using the photosensitiser meta-tetrahydroxyphenylchlorin (mTHPC). Animals were sensitised with 0.3 mg/kg mTHPC, 3 days prior to illumination with red light (652 nm) using a single fibre touching the target tissue and killed 1-3 days later for quantitative measurement of the extent of PDT necrosis. Results were similar for both tissues, although the differences between illumination regimens were less marked in tumour tissue. Using continuous illumination and a fixed low energy in colon, the extent of necrosis was up to almost three times larger with 5 mW than with 100 mW, although the maximum attainable necrosis was independent of power. The long treatment time using 5 mW could be halved without loss of effect by increasing the power during treatment. Dividing the light into two equal fractions at 100 mW increased the lesion size by up to 20% in colon (independent of the timing of the dark interval), but by only 10% in tumour and had no effect at 20 mW. Previous studies using 5-aminolaevulinic acid (ALA) showed a much larger effect of fractionation that was critically dependent on the timing of the dark interval. We postulate that enhancement of PDT by fractionation is due to improved oxygen supply to the treated area which may be due to reversal of temporary vascular occlusion (more likely with ALA) or less rapid photochemical consumption of oxygen (more likely with mTHPC). At lower fluence rates, the oxygen consumption rate is not fast enough to be improved by fractionation. We conclude that fractionated or low power light delivery can enhance PDT with mTHPC. Although the effects are not large, this may be of value for interstitial treatment of solid tumours when multiple sites are treated simultaneously. PMID- 12111596 TI - Retrospective analysis of clinical and microbiological aspects of Klebsiella oxytoca bacteremia over a 10-year period. AB - From 1991 to 2000, 125 sporadic cases of Klebsiella oxytoca bacteremia were analyzed retrospectively to review clinical features and to identify the risk factors associated with resistance to extended-spectrum cephalosporins and fatal outcome. Bacteremia was acquired nosocomially in 52% of the patients. Almost all patients (97%) had an underlying disease, with biliary and pancreatic disease occurring most frequently (55%). The biliary tract was the most common site of infection (44%). Resistance to extended-spectrum cephalosporins was identified in 22 of the 125 (18%) Klebsiella oxytoca blood isolates and resistance to ciprofloxacin in 9 (7%). Only previous antibiotic therapy was strongly associated with resistance to extended-spectrum cephalosporins in patients with Klebsiella oxytoca bacteremia ( P=0.009). The mortality rate was 24% and was higher in patients infected with isolates resistant to extended-spectrum cephalosporins (41% vs. 20%; P=0.04). In multivariate analysis, fatal outcome was independently associated with septic shock, deteriorated mental status, polymicrobial bacteremia, and solid tumor. Surgical therapy had a protective effect (OR, 0.06; 95% CI, 0.005-0.7; P=0.03). In conclusion, Klebsiella oxytoca bacteremia was most commonly associated with biliary tract infection. Previous antibiotic therapy was strongly associated with resistance to extended-spectrum cephalosporins in patients with Klebsiella oxytoca bacteremia. PMID- 12111597 TI - Double-blind, placebo-controlled study comparing the effect of azithromycin with clarithromycin on oropharyngeal and bowel microflora in volunteers. AB - The purpose of this double-blind study was to assess the effect of azithromycin and clarithromycin on oral and fecal microflora. Bacterial species from fecal samples and throat washes from healthy volunteers were identified and quantified before, during and after receipt of either placebo ( n=6), azithromycin (500 mg once daily for 3 days; n=6) or clarithromycin (500 mg twice daily for 7 days; n=6). In both antibiotic groups, the changes in oropharyngeal aerobic microflora following antibiotic administration were minor. Antibiotics neither changed the bacterial load of Streptococcus spp. compared with placebo, nor did macrolide resistant streptococci emerge. In the fecal aerobic microflora, the number of organisms of the family Enterobacteriaceae decreased slightly after antibiotic administration in both the clarithromycin and the azithromycin groups, but levels normalized by day 21 after therapy. No colonization with nonfermenters or Clostridium difficile was seen, and the total number of anaerobic bacteria was not affected in any study group. In conclusion, there were no significant differences between azithromycin and clarithromycin in their effect on human oropharyngeal and intestinal microflora, nor was the use of these antibiotics associated with colonization by resistant, gram-positive organisms or overgrowth of opportunistic microorganisms. PMID- 12111598 TI - Changing epidemiology of infective endocarditis: a retrospective survey of 108 cases, 1990-1999. AB - The aim of this study was to report the experience with infective endocarditis over the past decade, describe the changing clinical and epidemiological features of the disease, and attempt to determine the optimal number of blood culture sets required for diagnosis. All cases diagnosed during a 10-year period were reviewed clinically and microbiologically. In addition, a retrospective assessment of blood culture data was performed. From the period 1990-1999, 108 cases that met the von Reyn or Duke's criteria were recorded. The major underlying cardiac condition was the presence of a prosthetic valve ( n=33 patients, 31%). Among patients with native valves, nonrheumatic valvular heart disease of the elderly was the most common underlying factor ( n=19 patients, 25%). Overall, 13 patients (11%) died. Predictors on admission for increased mortality were shortness of breath, age >60 years, time to defervescence, erythrocyturia, hemoglobin level <10 g/dl, and leukocytosis >15,000 (all P<0.05). Analysis of blood culture data showed that the diagnostic yield among groups from whom either only one or more than six blood culture sets were drawn was reduced compared with that among groups from whom between two and five culture sets had been taken. The outcome of endocarditis in this series from a community hospital was much more favorable compared with that reported in surveys from large tertiary centers. Moreover, obtaining more than two or three blood cultures is neither helpful nor cost effective in the initial assessment of patients with suspected endocarditis. PMID- 12111599 TI - Prevalence of GB virus C (hepatitis G virus) and risk factors for infection in Sao Paulo, Brazil. AB - The prevalence of GB virus C (GBV-C) varies widely throughout the world. A cross sectional study was conducted in the city of Sao Paulo, Brazil, to estimate the prevalence of GBV-C infection and to identify associated risk factors, using a large sampling of the general population rather than blood donors or an illness related group of subjects. GBV-C RNA was detected by reverse-transcriptase polymerase chain reaction using primers directed to the 5' noncoding region (NCR) and nonstructural 5A region (NS5A) in serum samples from 1,039 healthy individuals 2 years of age or more. Fifty-two individuals were positive for both sets of primers and one was positive for NS5A only (prevalence of GBV-C infection, 5.1%; 95%CI, 3.9-6.7%). No child under 5 years of age was found positive. Among subjects aged 5 years or more, the prevalence of infection increased consistently with age, up to 30-39 years (8.3%), and decreased from then on. The number of sexual partners in the last 3 years (2 or more: OR, 2.6; 95%CI, 1.3-5.5) and history of contact with blood-sucking insects (OR, 2.5; 95%CI 1.2-5.4) were independently associated with GBV-C infection. In conclusion, the prevalence of GBV-C infection is high in Sao Paulo. In addition to parenteral transmission, another route, e.g. sexual or vertical, may be involved. PMID- 12111600 TI - Comparison of two surveillance methods for detecting nosocomial infections in surgical patients. AB - Nosocomial infections play a role in quality and cost control in health care. Surveillance of these infections is the only way to gain more insight into their frequency and causes. Since the results of surveillance may lead to changes in both patient and hospital management, which are sometimes major, it is necessary that all healthcare workers involved agree on the criteria used for the diagnosis and surveillance of these complications. In order to compare the efficacy of two surveillance methods, nosocomial infections in surgical patients were registered by both the Department of Surgery (complication surveillance [CS]) and the Department of Infection Control (nosocomial infection surveillance [NIS]) at the University Medical Center Utrecht, The Netherlands, over a 2-month period. The CS team used the national criteria of the Association of Surgeons of the Netherlands and the NIS team used the international criteria of the Centers for Disease Control and Prevention, USA, to define cases of nosocomial infection. A total of 515 patients were included in both arms of the study. The CS team diagnosed 69 infections in 49 patients, and the NIS team diagnosed 64 infections in 45 patients. Of 104 total infections, 39 were diagnosed by the CS team exclusively, 35 by the NIS team exclusively and only 30 by both. The main reasons for the inconsistent results were as follows: (i) the lack of follow-up after discharge in the NIS arm, (ii) the use of clinical criteria for the definition of a nosocomial infection in the CS arm, and (iii) the use of positive cultures as part of the criteria in the NIS arm. From the perspective of infection control, the CS system cannot be recommended for the surveillance of nosocomial infections. PMID- 12111601 TI - Differences between reference laboratories of the European community in their ability to detect Salmonella species. AB - The ability of national reference laboratories for Salmonella of the European Union member states to detect Salmonella bacteria was tested in four collaborative studies during the period 1995 through 1999. Three different methods were prescribed in the four studies. Capsules containing various numbers of Salmonella Typhimurium or Salmonella Enteritidis were tested. In studies II, III and IV, Salmonella bacteria were isolated in the presence of competitive microorganisms. Significant differences were found between the four studies due to varying levels of difficulty with regard to the level of contamination, the use of serotypes and the presence of competitive organisms. There were also significant differences between the laboratories in the results obtained. Possible reasons for these differences will be further investigated by the European Union Community Reference Laboratory for Salmonella. PMID- 12111602 TI - Evaluation of a new commercial assay for diagnosis of pulmonary and nonpulmonary tuberculosis. AB - A new commercial assay for the diagnosis of tuberculosis, the BDProbeTec ET Direct Detection assay (Becton Dickinson, USA), was evaluated using 351 respiratory and 372 nonrespiratory specimens. The results were compared to detection of Mycobacterium tuberculosis complex (MTC) by conventional culture. Among the 351 respiratory specimens, MTC bacteria were identified in 150, of which 85 were positive by both microscopy and the assay. Sixty-five specimens culture positive for MTC were microscopy negative; of these, 39 were positive in the assay. All 26 specimens culture positive for nontuberculous mycobacteria (NTM) were negative by the assay. Of 175 specimens culture negative for MTC, 3 were falsely positive by the assay and 1 yielded inhibition. The overall sensitivity and specificity values were 82.7% and 98.5%, respectively. The sensitivity for microscopy-positive and -negative respiratory specimens was 100% and 60%, respectively. After correction for discrepancies, the specificity was 99% compared with notification data. The BDProbeTec ET assay detected 66 of 67 microscopy-positive and 50 of 125 microscopy-negative nonrespiratory specimens. The result for one specimen was inconclusive. All nine specimens containing NTM were negative by the assay. Of 171 specimens culture negative for MTC, 6 were falsely positive by the assay. The overall sensitivity and specificity values obtained with nonrespiratory specimens were 60.7% and 96.7%, respectively. After examining discrepancies by reviewing the patients' histories, the specificity was 98.9%. The sensitivity was 98.5% in microscopy-positive specimens and 40.3% in microscopy-negative specimens. The overall inhibition rate was 0.3%. The BDProbeTec ET assay is a fast, effective, and user-friendly system that can be used for rapid detection of MTC bacteria in respiratory and microscopy-positive nonrespiratory specimens as an important supplement to smear and culture. PMID- 12111603 TI - Evaluation of a rapid immunochromatographic test for serodiagnosis of visceral leishmaniasis. AB - The purpose of this study was to compare the performance of a rapid immunochromatographic dipstick test for the qualitative detection of circulating antibodies to the leishmanial recombinant antigen K39 with that of a classical immunofluorescent antibody test for serodiagnosis of visceral leishmaniasis. Sera from 143 Italian subjects, including 69 patients with clinically suspected visceral leishmaniasis, 23 patients with hypergammaglobulinemia and 51 healthy controls, were tested. The immunochromatographic test was performed according to the manufacturer's instructions, using antigen-impregnated nitrocellulose paper strips. The immunofluorescent antibody test was performed according to an established method, using promastigotes of Leishmania infantum zymodeme Montpellier 1 as antigen. In 11 patients, diagnosis of active Leishmania infection was established by microscopic examination of biopsy samples and/or clinical response to meglumine antimoniate. Results of the two tests correlated for all but two sera examined. In two patients, one with proven infectious mononucleosis and one with bacterial pneumonia, the immunofluorescent antibody test was positive and the dipstick test was negative. In the restricted sample of patients in whom a definitive diagnosis was established, the immunochromatographic test was positive in 11 of 11 patients with confirmed Leishmania infection and negative in 103 of 103 subjects who either had other documented diseases or were healthy controls, showing 100% sensitivity and 100% specificity. PMID- 12111604 TI - Occurrence and clinical presentation of systemic pneumococcal infections in an unselected population in Oslo, Norway, between 1993 and 1997. AB - In order to describe the clinical and microbiological manifestations of systemic pneumococcal infection in an unselected urban population, 147 cases that occurred in the period 1993-1997 were retrospectively reviewed. An unexpected finding was that gastrointestinal symptoms were remarkably common. All pneumococcal isolates were fully susceptible to penicillin. The 7-valent conjugated vaccine covered 71% of those under 2 years of age, but only 21% of those 15-65 years of age were covered. Although the case fatality rate was 17%, the rate of early fatality due to systemic pneumococcal infection was unchanged compared with data published in the era before antibiotics. This study emphasizes the importance of continuing efforts to prevent systemic pneumococcal infections. PMID- 12111605 TI - Mycobacterium tuberculosis and Mycobacterium fortuitum osteomyelitis of the foot and septic arthritis of the ankle in an immunocompetent patient. AB - Mycobacteria, both tuberculous and nontuberculous, are recognized as a cause of chronic bone and joint infection. However, the diagnosis of mycobacterial infection is easily missed because of the absence of systemic involvement. Moreover, specific microbiologic techniques are required to detect mycobacteria in clinical specimens. Infections due to uncommon pathogens such as mycobacteria are more likely to occur in the immunocompromised host. A case of septic arthritis of the ankle and osteomyelitis of the foot due to both tuberculous and nontuberculous mycobacteria in an immunocompetent host is reported here. PMID- 12111606 TI - Clinical impact of nosocomial Klebsiella bacteremia in critically ill patients. AB - In order to determine the clinical impact of Klebsiella bacteremia on critically ill patients, a matched cohort study was conducted between January 1992 and December 2000. During the study period, all intensive care unit (ICU) patients with nosocomial Klebsiella bacteremia were defined as cases (n=52), but two of these patients were excluded from the matched cohort due to incomplete medical records. The remaining 50 patients were matched at a ratio of 1:2 with control patients (n=100) on the basis of the APACHE II severity of disease classification system. Patients with Klebsiella bacteremia experienced acute renal failure and hemodynamic instability more often than controls. They also had a longer ICU stay and longer ventilator dependence. In-hospital mortality rates for cases and controls were nearly equal (36% vs. 37%, respectively; P=0.905). In conclusion, after adjusting accurately for severity of underlying disease and acute illness, no difference in mortality was found between ICU patients with Klebsiella bacteremia and their matched control subjects. PMID- 12111607 TI - Spectrum of abdominal and pelvic infections caused by pneumococci in previously healthy adult women. AB - Eleven cases of pneumococcal infection of abdominal and pelvic origin that occurred in previously healthy adults are described. All cases occurred in women who were admitted to a county hospital in Norway with acute abdominal symptoms such as pain, nausea, vomiting and diarrhea. Explorative laparotomy was performed in eight patients. Three patients had diffuse peritonitis and seven cases were classified as pelvic inflammatory disease; five of these seven patients had pelvic peritonitis. All patients but one were treated with antibiotics, and all survived without long-term sequelae. Although abdominal and genital infections caused by pneumococci are rare, their potential occurrence should not be neglected. PMID- 12111608 TI - Primary HIV type-1 infection misdiagnosed as Mediterranean spotted fever. AB - The case studies of four patients, two men and two women between the ages of 42 and 54 years, are described. They presented to a hospital emergency department during the summer months with acute fever and exanthema. These are the primary symptoms of Mediterranean spotted fever (MSF), an endemic rickettsial disease in the Mediterranean basin that is seen particularly during the summer. The patients were clinically diagnosed as having MSF, but their diagnoses were not confirmed by serological testing. One patient was diagnosed with primary human immunodeficiency virus type 1 infection (HIV-1) 10 days later. The remaining three patients were diagnosed with HIV infection years later, but it is very likely that they also had primary HIV infection when MSF was presumed. When a patient develops sudden onset of fever and a maculopapular rash that is characteristic of MSF, the possibility of primary HIV-1 infection should be considered. PMID- 12111609 TI - First definite case of aortic valve endocarditis due to Moraxella phenylpyruvica. AB - Described here is the first definite case of endocarditis due to Moraxella phenylpyruvica, which occurred in a 50-year-old male with a bicuspid aortic valve. The diagnosis was delayed because of the confounding positivity of the Widal and Wright tests. The patient was cured with surgical valve replacement and antibiotic treatment. PMID- 12111610 TI - Serogroups, serotypes, serosubtypes and antimicrobial susceptibility of Neisseria meningitidis isolates in Casablanca, Morocco. AB - Since antigenic characterization and antibiotic susceptibility testing are useful for generating prophylactic recommendations and treatment guidelines, a total of 163 Neisseria meningitidis isolates obtained between January 1992 and September 2000 at the microbiology laboratory of the IbnRochd University Hospital of Casablanca, Morocco, were serogrouped, serotyped, serosubtyped and tested for their susceptibility to five antibiotics. Serogroup B was detected most frequently (75.5%), followed by serogroup A (13.5%). The phenotype B:4:P1.15 represented 74.8% of all serogroup B isolates. Seven (4.3%) isolates demonstrated decreased susceptibility to penicillin G. All isolates tested were susceptible to cefotaxime, chloramphenicol and rifampin. All isolates were inhibited by spiramycin at a concentration of 0.4 mg/l. PMID- 12111611 TI - Chronic osteomyelitis due to Actinomyces neuii subspecies neuii and Dermabacter hominis. PMID- 12111613 TI - Is the CACNA1A gene involved in familial migraine with aura? AB - The discovery of mutations in the neural calcium channel (CACNA1A) gene in familial hemiplegic migraine (FHM), variant of migraine with aura, led to the suggestion that this gene might be involved in familial migraine with aura (FMA). We investigated whether the mutations in FHM are present in FMA patients, analyzing genomic DNA by PCR, single stranded conformation polymorphism, sequencing and restriction enzyme. No mutations were found. A known polymorphism (5682-14C>T) was found in exon 36. These findings suggest that the mutations found in FHM and the other known mutations of the CACNA1A gene are not the genetic basis of FMA. Genetic alterations in FMA patients may be localized on chromosome 19 but not in the CACNA1A exons we investigated. PMID- 12111612 TI - Comparative trial of the effect of pneumococcal vaccine on viral load and CD4+ lymphocytes in asymptomatic and antiretrovirally naive HIV-infected patients. PMID- 12111614 TI - Familial hemiplegic migraine: clinical features and probable linkage to chromosome 1 in an Italian family. AB - We describe an Italian family with familial hemiplegic migraine (FHM), subtle cerebellar signs and probable linkage to chromosome 1. FHM is genetically heterogeneous; in about 50% of families it is caused by mutations within the CACNA1A gene on chromosome 19. Linkage to 1q31 and 1g21-23 has also been established. Other families do not link either to chromosome 19 or 1. Chromosome 19-linked FHM may display nystagmus and cerebellar ataxia. Affected family members were neurologically examined; linkage analysis was performed with markers for chromosomes 19p13, 1q21-23, and 1q32. Five family members had hemiplegic migraine, and 3 displayed additional cerebellar signs (scanning speech and nystagmus). In 1 patient, episodes of hemiplegic migraine triggered by mild head trauma. Epilepsy and mental retardation were also found in 1 affected relative each. Lod scores for linkage to 19p13 were negative, while the maximum two-point lod score was 1.81 to 1q21-23. This family with FHM and associated subtle cerebellar signs, epilepsy and mental retardation showed probable linkage to 1q21 23. PMID- 12111615 TI - A case-control study on Alzheimer's disease and exposure to anesthesia. AB - Aretrospective hospital-based case-control study was performed with the aim to evaluate the association between exposure to anesthesia and Alzheimer's disease (AD). A total of 115 AD patients, 230 Parkinson's disease (PD) patients and 230 patients with non-degenerative neurological disease were studied. Each AD case was matched for sex, age (+/-3 years) and geographic area of residence with four controls (2 PD patients and 2 with other neurological disease). Information about exposure to general anesthesia and other variables was gathered through hospital records. No associations were found between the risk of AD and the exposure to anesthesia in the 1 and 5 years preceding disease onset, nor between the risk of AD and the number of surgical operations. A significant difference was observed between the mean age of AD patients and controls undergoing surgical procedures. The present study reveals a lack of association between exposure to general anesthesia and AD. Prospective epidemiological studies are needed in order to investigate levels of exposure to anesthesia, as well as any possible relationships between anesthetic exposure and genetic factors (e. g. APOEepsilon4 genotype). PMID- 12111616 TI - Incidence, risk factors and short-term mortality of stroke in Vittoria, southern Italy. AB - We determined the incidence, risk factors, and short-term mortality of stroke in a well-defined area of southern Italy, i. e. the city of Vittoria, Sicily (58 833 inhabitants). The medical recores of the local hospitals and the outpatient files of the local neurologist referring to the calendar year 1991 were retrospectively investigated. Stroke was defined according to standard criteria and classified as first-ever (FE) and recurrent (R). Risk factors for stroke were diagnosed from medical history, laboratory and instrumental findings, and in the presence of specific treatments. Short-term mortality was assessed as 30-day case-fatality rate. The sample included 120 cases (61 men) aged 34-94 years, 89 of whom (48 men) had a FE stroke. The overall annual crude incidence rate of FE stroke was 165.3 per 100 000 (men, 178.4; women 152.2); for FE and R stroke together it was 222.9 (men, 226.8; women 219.1). The standardized rates were 245.3 (FE stroke) and 321.9 (FE and R stroke). The age-specific rates for FE stroke were 9.4 (<55 years), 262.2 (55-64 years), 645.2 (65-74 years), 2019.7 (75-84 years), and 3246.8 (> or =85 years). The corresponding values for FE and R stroke were 11.7, 412.0, 887.1, 2565.5, and 4220.8. In patients with FE stroke, cerebral infarction was the main type. Hypertension, diabetes and cardiac disorders were the commonest risk factors, with similar distribution among FE and R stroke. The 30 day case-fatality rate was 28% for FE and R stroke and 38% for FE stroke. Compared to other reports, the incidence of stroke in Vittoria was lower in the youngest but higher in the oldest age groups. Although the small sample size and possibility of misdiagnoses may partly explain our findings, the roles of different dietary, social, and genetic factors in the local population warrant investigation. PMID- 12111617 TI - SPASE-I: a multicenter observational study on pharmacological treatment of acute stroke in the elderly. The Italian Study of Pharmacological Treatment of Acute Stroke in the Elderly Group. AB - We investigated the clinical outcome and current therapeutic management of ischemic stroke in elderly patients in different neurological wards throughout Italy. Twelve centers in 10 Italian regions were involved. The clinical and instrumental data were prospectively collected for 335 patients 65 years or older, presenting in a 12-months period. At the multivariate analysis, death was associated with the presence of total anterior circulation infarct and the use of antiedema agents. Stroke-related disability was related to female sex, unified stroke scale (USS) scores at entry and mini-mental state examination (MMSE) scores at the 1-months follow-up. The majority of patients (70%) received antiplatelets to prevent stroke recurrence. Very elderly (> or =80 years) patients had worse presentation and prognosis than less elderly stroke victims (<80 years). Age played an important role in the therapeutic approach to stroke: while antiplatelets were widely used, oral anticoagulants were underused even when specifically indicated. PMID- 12111618 TI - External ventricular drainage for acute obstructive hydrocephalus developing following spontaneous intracerebral haemorrhages. AB - There is no consensus in the literature on the effects of the development of hydrocephalus on survival and disability after intracerebral haemorrhage (ICH) and the benefits of external ventricular drainage (EVD). In this open, prospective study, we investigated the clinical courses, radiological findings and outcome scores of 47 consecutive patients who were admitted to our clinic with spontaneous ICH. Hydrocephalus developed in 6 (12.8%) of the 47 patients, and EVD was applied in these 6 cases. In one of the 6 patients, the lesion was additionally excised due to the large cerebellar haematoma. Intraventricular haemorrhage was more common in patients developing hydrocephalus (83.3% vs. 29.3% in patients without hydrocephalus; p<0.05) and the lesions of all the patients were in the proximity of the ventricular system. Hospital mortality and functional outcome were not significantly different between patients with and without hydrocephalus. Our results shown that acute obstructive hydrocephalus should be anticipated if haematoma is near the ventricle or if it is opening to the ventricle. EVD is a life-saving and effective procedure that should be performed in patients who develop hydrocephalus following spontaneous intracerebral haemorrhage. PMID- 12111619 TI - Sleep disturbances associated with minor psychiatric disorders in medical students. AB - We performed a cross-sectional study with 342 medical students (age range, 18-35 years) to identify, among a group of sleep disturbances, those which are related to minor psychiatric disorders in this population. The instruments employed for data collection were the self-reporting questionnaire (SRQ-20), the morningness/eveningness questionnaire, the Epworth sleepiness scale, and a general questionnaire regarding demographic characteristics, use of drugs, history of psychopathology, usual fall-asleep time, usual wake-up time, amount of sleep, arousal during the night, and insomnia. We used a logistic regression model to determine independent factors associated with minor psychiatry disorders. Daytime sleepiness [odds ratio (OR), 2.12; 95% CI, 1.21-3.71], arousal [OR, 4.54; 95% CI, 1.97-10.47], insomnia [OR 2.45; 95% CI, 1.32-4.56], and sleeping less than 7 hours per night [OR, 2.02; 95% CI, 1.11-3.67] were associated with minor psychiatric disorders. This group of variables determined a cumulative risk ratio of 5.47 [95% CI, 2.87-10.41] for the main outcome. PMID- 12111620 TI - Donepezil in the treatment of hallucinations and delusions in Parkinson's disease. AB - As cholinergic mechanisms may be at least partially responsible for hallucinations and delusions in Parkinson's disease (PD), we conducted an open study in 8 PD patients to assess the efficacy and tolerability of the cholinesterase inhibitor donepezil, 5 mg at bedtime for two months, in the treatment of these complications. Hallucinations and delusions improved significantly in all patients. Donezepil was overall well tolerated, but a deterioration in motor disability was noted in 2 out of 8 patients. PMID- 12111621 TI - Leukocyte count in the synovial fluid of children with culture-proven brucellar arthritis. AB - Brucellosis is an important cause of paediatric septic arthritis in endemic areas. Because the Gram stain is frequently negative and culture results are unavailable at the time of the patient's admission, the diagnosis of brucellar arthritis is usually entertained on the bases of epidemiological considerations and cytological examination of the synovial fluid aspirate. The aim of this study was to assess the sensitivity of a synovial fluid leukocyte count >50 000 WBC/mm(3) for detecting culture-proven brucellar arthritis in children. The medical records of all children with brucellar arthritis diagnosed since 1994 in a hospital serving an endemic area for brucellosis in southern Israel were reviewed. Nine patients (six males and three females), aged 3-14 years, were identified. A single joint was affected in all patients. The median leukocyte count in the synovial fluid was 9500 WBC/mm(3) (range 300-61 500 WBC/mm(3)), and in eight of the nine patients it was less than 50 000 WBC/mm(3). Brucella melitensis was recovered from the synovial fluid culture in all patients. The diagnosis of brucellar septic arthritis cannot be excluded on the basis of a low leukocyte count in the joint aspirate. A high index of suspicion and use of modern culture techniques are recommended to improve the diagnosis of brucellar arthritis. PMID- 12111622 TI - Absence of association of the serotonin transporter gene polymorphism with the mentally healthy subset of fibromyalgia patients. AB - The serotonin transporter (5-HTT) gene is considered to be a promising candidate for genetic involvement in some mood disorders owing to its role in the regulation of serotoninergic neurotransmission. In this study, we aimed to assess the significance of the 5-HTT gene in fibromyalgia syndrome (FS) as well as to find out whether the 5-HTT gene polymorphism is associated with this disease. Fifty-three mentally healthy fibromyalgia patients and 60 unrelated healthy volunteer controls were included in the study. Symptom Checklist-90-Revised (SCL 90-R), Beck Depression Inventory (BDI), and State and Trait Anxiety Inventory tests (STAI-I and II) were applied to both patients and controls. A PCR analysis of 5-HTT gene polymorphism was performed, and the results of the patients with FS and healthy controls were compared. In both FS patients and healthy controls the S/S, S/L and L/L alleles of the 5-HTTLPR genotype were represented in 24.5 % and 33%, 56.6% and 38.3%, and 18.9% and 28.3%, respectively. Additionally, in FS patients and healthy controls the 10/10, 10/12 and 12/12 alleles of the VNTR variant were represented in 5.9% and 11.7, 51% and 36.7%, and 43.1% and 51.7%, respectively. The 5-HTTLPR and VNTR results of the patients and controls were not significantly different ( P>0.05). We concluded that neither 5-HTT nor its polymorphism is associated with FS. Our results also address the frequencies of 5 HTT gene alleles in our population. Further studies are required to better understand the genetic basis of FS. PMID- 12111623 TI - Alkaline phosphatase in rheumatoid arthritis patients: possible contribution of bone-type ALP to the raised activities of ALP in rheumatoid arthritis patients. AB - Raised serum alkaline phosphatase (ALP) activity in rheumatoid arthritis (RA) has been reported, although its aetiology is not clear. In this paper we investigate whether synovial tissue is a possible source of raised ALP activity in RA. The activities and isozymes of ALP were determined in sera and synovial fluids from 22 RA and seven osteoarthritis (OA) patients. The expression of both protein and ALP mRNA in synovial tissue was investigated immunohistochemically and by reverse transcription (RT) PCR. ALP activity was higher in serum and synovial fluid from RA patients than in those from OA patients. In addition, the ratio of levels of bone-type ALP to those of liver-type ALP was significantly higher in synovial fluid than in serum from RA patients. Bone-type ALP was positive around the perivascular area and the subepithelial cells in the synovial tissue from RA patients. In contrast, the synovial tissue from OA patients exhibited no staining. The mRNA of bone-type ALP was detected in RA synoviocytes. In conclusion, ALP levels were elevated in both serum and synovial fluid from RA patients. Bone-type ALP derived from the synovial tissue may contribute to the raised activities of ALP in RA patients. PMID- 12111624 TI - Ultrasound and clinical evaluation of quadricipital tendon enthesitis in patients with psoriatic arthritis and rheumatoid arthritis. AB - Enthesitis is an inflammatory lesion of the tendon, ligament and capsular insertions into the bone, and it is a fundamental element in the diagnosis of spondyloarthropathies. Sonography is the method of choice for studying periarticular soft tissues because it is capable of detecting both the early (oedema, thickening) and the late alterations (erosions and enthesophytes); it is also an inexpensive, biologically harmless and easily repeatable technique. The aim of this study was to compare the prevalence of quadricipital enthesitis in psoriatic arthritis (PsA) and rheumatoid arthritis (RA) patients, and to document any clinical and echostructural differences in this lesion between the two diseases. The results show that enthesitis is more frequent in PsA patients, more than half of whom are asymptomatic. Knee inflammation was found in the PsA patients with enthesitis regardless of the concomitant presence of joint effusion; none of the RA patients suffered from enthesitis alone. Quadricipital enthesitis is more frequent in male patients. There was no significant correlation between the presence of peripatellar psoriatic lesions and enthesitis. Sonographic examinations of patients with enthesitis revealed that those with RA had dominantly inflammatory lesions, whereas PsA patients also showed major new bone deposition. PMID- 12111626 TI - Effect of interferon-alpha(2a) on neutrophil adhesion and phagocytosis in chronic myeloid leukemia and Behcet's disease. AB - Alpha-interferon (alpha-IFN) is implicated in a Behcet's disease (BD)-like syndrome observed in a small number of chronic myeloid leukemia (CML) patients. The effect of alpha-IFN on neutrophil adhesion and phagocytosis in CML patients, BD patients and healthy volunteers was investigated to clarify the reason for this observation. Ten subjects were studied for each group by incubating neutrophils with various doses of alpha-IFN. Basal neutrophil adhesions for CML patients, BD patients and healthy volunteers were similar. However, BD patients had greater basal phagocytosis than CML patients, and both groups had greater basal phagocytosis than healthy volunteers. Neutrophil adhesion and phagocytosis of CML patients increased following incubation with higher doses of alpha-IFN, and phagocytosis approached the high levels observed with BD neutrophils. This study provides evidence that alpha-IFN activates neutrophils in CML patients in a dose-dependent manner, and leads to a neutrophil function profile that resembles BD. PMID- 12111625 TI - Transverse myelitis in patients with antiphospholipid antibodies--the importance of early diagnosis and treatment. AB - Transverse myelitis (TM) is a rare manifestation of systemic lupus erythematosus (SLE) and the antiphospholipid syndrome (APS). No uniform therapeutic protocol exists for its treatment, and the prognosis is usually poor. Here we describe four patients having TM associated with antiphospholipid antibodies. Treatment measures and delay in diagnosis between symptom onset and the initiation of treatment varied between patients, but the earlier the diagnosis and the more aggressive the treatment the better was the patient's outcome. Based on these cases and on a literature review we suggest that early aggressive treatment (usually with pulses of methylprednisolone and cyclophosphamide) might improve the prognosis of patients with TM associated with antiphospholipid antibodies. PMID- 12111627 TI - A prospective study on silicone breast implants and the silicone-related symptom complex. AB - This cohort study prospectively evaluated the prevalence of the silicone-related symptom complex (SRSC) in relation to antinuclear antibodies (ANA) and magnetic resonance imaging (MRI) of silicone breast implants (SBI) 1 year after implantation. A total of 57 women undergoing mastectomy followed by immediate breast reconstruction (IBR) and SBI between March 1995 and March 1997 at the University Hospital Rotterdam/Daniel den Hoed Cancer Centre, were prospectively evaluated. Just before and 1 year after IBR the sera of these women were tested for the presence of ANA and they were screened for the prevalence of SRSC-related symptoms by questionnaire. All prostheses were evaluated by MRI 1 month and 1 year after IBR. Just before operation 11% of the women had a Sjogren score of more than 2, whereas 30% had such a score 1 year after IBR ( P = 0.01). One year postoperatively women had significantly more RA/Raynaud-related complaints: 21% preoperatively versus 40% 1 year after IBR ( P = 0.03). Within the undefined complaints-related group 19% had a score of 2 or more preoperatively and 33% 1 year after IBR ( P = 0.09). There were no new cases of ANA positivity 1 year after IBR. The linguine sign was seen by MRI in three implants: one 1 month after IBR and two 1 year after IBR. There was no relation to changes in SRSC expression and these MRI findings. In conclusion, 1 year after SBI implantation women had more SRSC-related complaints, especially Sjogren's and RA/Raynaud's. Moreover there was no correlation between elevated SRSC expression and changes in the presence of ANA or changes in MRI of the SBI 1 year after IBR. PMID- 12111628 TI - Analysis of the reasons for DMARD therapy discontinuation in patients with rheumatoid arthritis in the Czech and Slovak republics. AB - The aim of the study was to evaluate the efficacy and safety of disease-modifying drugs (DMARDs) in everyday clinical practice in Central European States (the Czech and Slovak republics). This was a retrospective, multicentre study. With the help of a special questionnaire, the medical files of 760 patients in 15 centres were analysed looking for reasons for DMARD discontinuation (e.g. insufficient efficacy, toxicity). The secondary endpoints were duration of therapy with individual DMARDs and the influence of other factors (demographic, disease specific, concomitant therapy) on duration of therapy. In 47.1 % of patients therapy was interrupted because of lack of efficacy, in 43.2 % because of adverse events, and in 9 % for undefined reasons. Toxic reactions leading to withdrawal were most common with gold (62.6 %) and methotrexate (62.5 %). Because of insufficient effect, treatment was most frequently interrupted with antimalarials (62.3 %) and penicillamine (53.2 %), but in only 22% treated with methotrexate. The mean duration of one treatment episode with DMARDs was 28.1 +/- 48.9 months. Surprisingly, it was longest for cyclophosphamide (53.5 + 55.1 months) and shortest for cyclosporin (7.0 +/- 6.7 months). The mean duration of treatment with methotrexate was only 14.9; +/- 16.2 months. The mean duration of treatment with one DMARD was statistically longer in patients with positive rheumatoid factor, extra-articular disease and age lower than 50 years. There was no impact of sex, concomitant steroid treatment and high or low sedimentation rate on treatment duration. Considerable differences in everyday clinical practice with DMARDs between Central European states and published data from the US and western Europe have been found. More education about modern strategies in the treatment of RA is probably necessary for practising rheumatologists. PMID- 12111629 TI - Cervical spine disorders in patients with rheumatoid arthritis and amyloidosis. AB - The aim of this radiographic study was to ascertain the extent of inflammatory cervical spine disorders in patients with rheumatoid arthritis (RA) complicated by secondary amyloidosis (SA). The study involved 147 patients with RA and SA, whose cervical spine radiographs were available. They were treated at the Rheumatism Foundation Hospital, Heinola, during the period 1989-2000 and had had RA for a mean of 24 years. The inflammatory abnormalities of the cervical spine were studied from radiographs taken at or after the diagnosis of SA during flexion and extension. One-hundred and eleven (76%) patients had subluxations, impaction or apophyseal joint ankylosis. Atlantoaxial impaction (AAI) was seen in 76 (52%) patients and anterior atlantoaxial subluxation (AAS) in 59 (40%). Apophyseal joint ankylosis was the third most frequent finding, seen in 34 (23%) cases. A combination of AAI and apophyseal joint ankylosis was noted in 26 (18%) patients. Eight (5%) patients had undergone surgery on the cervical spine. In conclusion, inflammatory and destructive changes are frequent in the cervical spine of patients with RA and SA. Characteristic changes are AAI and AAS. RA patients with SA have more severe disease than those in epidemiological studies when cervical spine disorders are concerned. PMID- 12111630 TI - Systems to assess the progression of finger joint osteoarthritis and the effects of disease modifying osteoarthritis drugs. AB - Our objective was to assess the progression of osteoarthritis (OA) using scoring systems based on the anatomical changes recorded in the finger joints on standard radiographs and to test how far these scoring systems could be used to evaluate the effects of candidate "disease modifying osteoarthritis drugs" (DMOAD). The appearance and growth of osteophytes, narrowing of the joint space and subchondral bone changes allowed the classic OA-associated anatomical lesions to be used to score the progression of finger joint OA. Progression of OA in the finger joints was also assessed by the their evolution through previously described and predictable anatomical phases on standard X-rays. These phases were characterised by complete loss of the joint space preceding or coinciding with the appearance of subchondral cysts eroding the entire subchondral plate, and have been described in "inflammatory" or "erosive" OA. The erosive episodes were followed by processes of remodelling. In order to interfere with the progression of osteoarthritis, two chondroitin sulphates with possible DMOAD effects were used in two series of patients with OA of the finger joints. The patients were included in two separate randomised, double-blind placebo-controlled trials: 46 of them received chondroitin polysulphate and 34 received chondroitin sulphate. Eighty-five patients were kept on placebo medication and were used as controls. All 165 patients were followed for 3 years. Posteroanterior X-rays of the metacarpophalangeal and interphalangeal (IP) finger joints were obtained at the start of this prospective study and at yearly intervals thereafter. Almost 80% of the distal IP and 50% of the proximal IP were affected at study entry. In approximately 40% of the patients the classic picture of OA of the IP joints was complicated by manifest erosive OA changes. The two systems to score the progression of OA (Anatomical Lesion and Anatomical Phase Progression Score System) showed definite progression within 3 years of follow-up, especially in the IP joints. When compared with the placebo controls, none of the chondroitin sulphates prevented OA from occurring in previously normal finger joints. However, when the classic OA-associated anatomical lesions were considered, OA was less progressive in both active treatment groups. Furthermore, fewer patients from both chondroitin sulphate- and chondroitin polysulphate-treated groups developed "erosive" osteoarthritis. In conclusion, conventional radiographs can be used to assess the morbidity and progression of hand OA. The systems used to score the progression of finger joint OA allowed the DMOAD effects of both chondroitin sulphates to be evaluated. The data recorded during these pilot studies should help investigators to design future long-term clinical experiments. PMID- 12111631 TI - Systemic sclerosis therapy with iloprost: a prospective observational study of 30 patients treated for a median of 3 years. AB - Iloprost is useful in the short-term treatment of severe Raynaud's phenomenon and ischaemic ulcers in patients with systemic sclerosis (SSc), but its long-term effects are largely unknown. The aim of this study was to report long-term outcome (median follow-up 36 months) in a prospective observational study of a cohort of 30 consecutive patients with SSc who received iloprost therapy with maintenance infusions every 3 weeks after an initial cycle of 5 consecutive days. At the end of the observation, compared to the pretreatment point, we observed complete healing of digital ulcers in 19/21 patients (90%), a decrease of the Raynaud's phenomenon visual analogue score from 10/10 (25th-75th percentile 7-10) to 5/10 (4-6.75) ( P <0.001) and, in patients with diffuse cutaneous involvement, of the modified Rodnan skin thickness score from 25.5 (16.5-31.5) to 16 (13.5-20) ( P = 0.02), minimal improvement of the Health Assessment Questionnaire from 0.87 (0.68-1.37) to 0.75 (0.62-1.25), which was neither statistically nor clinically significant. The forced vital capacity was not significantly changed, but the diffusion capacity corrected for the alveolar volume decreased from 71% (54-76.7) of the expected value to 62% (51.5-71) ( P = 0.02). In one patient with limited SSc a positive effect on pulmonary hypertension was observed. Six patients, after a median of 25 months of treatment and healing of digital ulcers, discontinued the therapy; after a median of 10 months ulcers did not recur in five of these six. Other reasons for discontinuation were: tolerability (1), disease progression (normotensive renal crisis: 1), and death due to intracranial haemorrhage (1). This same patient had previously suffered a central retinal vein thrombosis. In conclusion, long-term therapy with iloprost in patients with SSc has a durable effectiveness on ischaemic ulcers and Raynaud's phenomenon, but it is not possible to conclude that the natural history of the disease was modified. PMID- 12111632 TI - Elevated homocysteine levels in patients with Raynaud's phenomenon secondary to systemic lupus erythematosus. AB - The objective of this study was to analyse the association between serum homocysteine (Hcy) levels and the presence of Raynaud's phenomenon (RP) in a cohort of systemic lupus erythematosus (SLE) patients. We enrolled premenopausal, disease-inactive SLE patients ( n = 34) with RP (group I, n = 11) or without RP (group II, n = 23), and age-matched healthy premenopausal women as controls (group III, n = 20). Fasting Hcy levels were determined for all these subjects. The results reveal that group I patients exhibited significantly greater serum Hcy levels (11.68+/-2.98 mmol/l) than group II patients (8.29+/-2.89 mmol/l) ( P = 0.003), and group III healthy subjects (8.00+/-1.50 mmol/l) ( P = 0.001); however, in this regard there was no significant difference between group II and group III patients ( P = 0.927). In conclusion, elevated serum Hcy levels were noted for this cohort of SLE patients with RP compared to those not evidencing RP and healthy controls. Although the pathogenesis of RP still remains obscure, this study suggests that Hcy may play a role in the aetiopathogenesis of SLE patients with RP. PMID- 12111634 TI - Coexistence of diffuse idiopathic skeletal hyperostosis and ankylosing spondylitis: a case report. AB - Diffuse idiopathic skeletal hyperostosis and ankylosing spondylitis are two diseases which are listed in the differential diagnosis of each other. There have been limited numbers of case reports regarding the coexistence of both diseases in the literature. We describe a patient who demonstrated the features of diffuse idiopathic skeletal hyperostosis with coexisting features resembling ankylosing spondylitis in order to discuss the association of the two diseases. PMID- 12111633 TI - Interleukin-1 receptor antagonist gene polymorphism in chinese patients with systemic lupus erythematosus. AB - The purpose of this study was to determine whether IL-1 receptor antagonist (IL 1Ra) gene polymorphism is a marker of susceptibility to or severity of systemic lupus erythematosus (SLE) in Chinese patients. The study included 52 Chinese patients with SLE. One hundred and three unrelated, healthy individuals living in central Taiwan served as controls. From genomic DNA, the polymorphism of the gene for IL-1Ra was typed. Allelic frequencies and carriage rates were compared between SLE patients and controls. The relationship between allelic frequencies and clinical manifestations of SLE was evaluated. We found an increased frequency of IL1RN*2 in the SLE patients compared to normal controls (chi(2) = 4.15, P<0.05), with an odds ratio (of allele frequency) of 2.63 (95% confidence interval 1.00-6.96). The carriage rate of IL1RN*2 was also higher in the SLE patients (6.8% in the controls vs. 17.3% in the SLE patients). We observed increased frequencies of malar rash and photosensitivity among patients with IL1RN*2 (77.8%) compared to patients without the allele (48.8%). However, this difference did not reach statistical significance (chi(2) = 2.51, P = 0.11). This study indicated that the frequency of IL1RN*2 is higher in Chinese SLE patients than in Chinese normal controls in Taiwan. However, there was no association between the frequency of IL1RN*2 and clinical manifestations. PMID- 12111635 TI - Churg-Strauss syndrome presenting as spontaneous subarachnoid haemorrhage. AB - Churg-Strauss syndrome (CSS) is a systemic small-vessel vasculitis characterised by the presence of asthma and eosinophilia. Central nervous system involvement (cerebral infarctions or intracerebral haemorrhage) is rare in CSS. Spontaneous subarachnoid hemorrhage (SAH) has been described in other systemic vasculitides. SAH is exceptional in CSS. We present a 47-year-old woman with CSS presenting as a spontaneous SAH with cerebral angiography findings consistent with vasculitis of the basilar artery and without aneurysms or arteriovenous malformations. She received treatment with prednisone and cyclophosphamide, and 2 months later the basilar artery was normal on magnetic resonance angiography. PMID- 12111636 TI - Pancreatic pseudocyst in paediatric systemic lupus erythematosus. AB - Pancreatitis is a rare complication of paediatric systemic lupus erythematosus (SLE). We describe a child with severe form of SLE who initially developed acute pancreatitis, subsequently complicated by extensive pancreatic pseudocyst. The treatment and outcome are discussed. PMID- 12111637 TI - Idiopathic intracranial hypertension with elevated cerebrospinal fluid level of interleukin-6 in a patient with systemic lupus erythematosus. PMID- 12111638 TI - Thirty distinct CACNA1F mutations in 33 families with incomplete type of XLCSNB and Cacna1f expression profiling in mouse retina. AB - X-linked CSNB patients may exhibit myopia, nystagmus, strabismus and ERG abnormalities of the Schubert-Bornschein type. We recently identified the retina specific L-type calcium channel alpha1 subunit gene CACNA1F localised to the Xp11.23 region, which is mutated in families showing the incomplete type (CSNB2). Here, we report comprehensive mutation analyses in the 48 CACNA1F exons in 36 families, most of them from Germany. All families were initially diagnosed as having the incomplete type of CSNB, except for two which have been designated as Aland Island eye disease (AIED)-like. Out of 33 families, a total of 30 different mutations were identified, of which 24 appear to be unique for the German population. The mutations, 20 of which are published here for the first time, were found to be equally distributed over the entire gene sequence. No mutation could be found in a classic AIED family previously shown to map to the CSNB2 interval. Cacna1f expression in photoreceptor-negative mice strains indicate that the gene is expressed in the outer nuclear, the inner nuclear, and the ganglion cell layer. Such a distribution points to the central role of calcium regulation in the interaction of retinal cells that mediate signal transmission. PMID- 12111639 TI - Identification of novel SDHD mutations in patients with phaeochromocytoma and/or paraganglioma. AB - Familial paraganglioma is a dominantly inherited disorder characterised by the development of highly vascular tumours in the head and neck. Recently, a relationship between hereditary tumours derived from the autonomic nervous system and germline mutations in the gene encoding succinate dehydrogenase complex subunit D (SDHD) is increasingly a subject of study. Familial paraganglioma syndrome is embryologically related to phaeochromocytoma, another neuroendocrine tumour that shows great aetiological and genetic heterogeneity. Some hereditary phaeochromocytomas may be associated with germline mutations in VHL, RET and NF1 genes in genetic disorders such as von Hippel-Lindau disease (VHL), multiple endocrine neoplasia type 2 (MEN 2) and neurofibromatosis type 1 (NF 1), respectively. However, there are many cases that cannot be explained by mutations in these genes. In this report, we describe two previously unreported mutations in two patients from 25 unrelated kindreds with phaeochromocytoma and/or paraganglioma disorders and with or without familial antecedents: a mutation featuring the change of tryptophan to a termination codon in exon 2, and a 4-bp deletion in exon 4 that results in a truncated protein. We also describe one missense substitution of uncertain significance. The patients had previously tested negative for germline mutations in VHL and RET genes and had not been previously selected. The involvement of SDHD mutations in familial phaeochromocytoma and/or paraganglioma predisposition is of considerable interest since other studies have shown these alterations to be associated with highly expressed angiogenic factors. PMID- 12111640 TI - Rapid detection of common autosomal aneuploidies by quantitative fluorescent PCR on uncultured amniocytes. AB - Prenatal diagnosis of chromosomal abnormalities by cytogenetic analysis is time consuming, expensive, and requires highly qualified technicians. Rapid diagnosis of aneuploidies followed by reassurance for women with normal results can be performed by molecular analysis of uncultured foetal cells in less than 24 h. Today, all molecular techniques developed for a fast diagnosis of aneuploidies rely on the semi-quantification of fluorescent PCR products from short tandem repeat (STR) polymorphic markers. Our objective was to test a chromosome quantification method based on the analysis of fluorescent PCR products derived from non-polymorphic target genes. An easy to set up co-amplification of portions of DSCR1 (Down Syndrome Critical Region 1), DCC (Deleted in Colorectal Carcinoma), and RB1 (Retinoblastoma 1) allowed the molecular detection of aneuploidies for chromosomes 21, 18 and 13 respectively. Quantitative analysis was performed in a blind prospective study of 400 amniotic fluids. Four samples (1%) could not be analysed by PCR probably because of a low concentration of foetal DNA. Follow up karyotype analysis was done on all samples and molecular results were in agreement with the cytogenetic data with no false-positive or false-negative results. Our gene based fluorescent PCR approach is an alternative molecular method for a rapid and reliable detection of aneuploidies which can be helpful for the clinical management of high-risk pregnancies. PMID- 12111641 TI - Unique (Y;13) translocation in a male with oligozoospermia: cytogenetic and molecular studies. AB - The incidence of Y/autosome translocations is low. Whereas involvement of non acrocentric chromosomes often leads to infertility, cases related with acrocentric chromosomes are usually familial with no or minimal effect on fertility. A de novo (Yp/13p) translocation was found in a 32-year-old male referred for severe oligozoospermia. Conventional cytogenetic procedures (GTG, CBG and NOR banding) and molecular cytogenetic techniques (Fluorescence In Situ Hybridization, FISH) were performed on high-resolution chromosomes obtained after peripheral blood lymphocyte culture as also on interphase nuclei of spermatogenic cells from semen samples. Screening of AZF microdeletions in the Yq11.2 region known to be involved with spermatogenesis defects was also performed. GTG banding showed a (Yp/13p) translocation in all scored metaphases. CBG and NOR staining of the derivative chromosome revealed the maintenance of Yq heterochromatin and of the 13p NOR region. FISH with centromeric Y and 13/21 probes, SRY specific probe and X/Y (p and q arms) sub-telomeric probes gave the expected number/location of fluorescent signals. Hybridisation with a pan-telomeric repeat (TTAGGG) probe showed an absence of the telomeric sequences at the fusion point of the rearranged chromosome. FISH analysis with probes to chromosomes X, Y, 13 and 18 showed an abnormal segregation of the translocated chromosome during meiosis I, which explains that only 13.6% of the secondary spermatocytes were normal. Most of these became arrested, as after meiosis II the large majority of the round spermatids were normal (70%), as were in consequence most of the sperm (85.1%). Multiplex-PCR confirmed the intactness of the SRY region and showed absence of AZF microdeletions. We report a novel de novo (Yp;13p) translocation characterised by loss of the 13p and Yp telomeres. Meiotic studies using FISH demonstrated meiosis I chromosome unpairing and mal segregation that justifies the severe oligozoospermia. Although most sperm have a normal chromosomal constitution, preimplantation genetic diagnosis should be considered an option for this patient. PMID- 12111642 TI - Both common and unique susceptibility genes in different rat strains with pristane-induced arthritis. AB - Pristane-induced arthritis (PIA) in rats is an animal model for rheumatoid arthritis (RA). We have previously identified seven quantitative trait loci (QTLs), which regulate arthritis development using a cross between the susceptible DA strain and the resistant E3 strain of rats (Pia2-8). In the present study the inbred rat strain LEW.1F was used as the susceptible strain in a cross with the E3 strain. The results confirmed the locus Pia4 on chromosome 12, which previously was shown to be associated with PIA, and also with experimental allergic encephalomyelitis, in crosses between the rat strains E3 and DA. On chromosome 1, linked to the albino locus, we identified a novel QTL, Pia9 in the LEW.F1 cross. This locus was associated with arthritis severity in the early phase of disease. A locus on chromosome 16, denoted Pia11, was also associated with arthritis severity in the early phase of the disease. A suggestive locus was detected on chromosome 14, which was associated with arthritis severity at the time when PIA progresses into a chronic phase. Using a congenic LEW.1F strain, which carries E3 alleles at the Pia9 locus, we confirmed that the E3 allele significantly suppresses arthritis severity during the early phase of the disease. The results revealed synergistic effects between different susceptibility loci using ANOVA analysis. These interactions were influenced by gender. Rats with Pia9 alleles from LEW.1F and Pia11 alleles from E3, were shown to suffer from much more severe arthritis in the early stage of the disease. On the other hand, the Pia9 and the suggestive locus on chromosome 14 affected only males during the chronic phase of the disease. These findings provide clues to how genetic factors by themselves, and in interaction with each other, regulate the development of a disease, which displays many similarities to RA. PMID- 12111643 TI - MECP2 gene mutation analysis in Chinese patients with Rett syndrome. AB - Rett syndrome (RTT) is a progressive neurodevelopmental disorder that affects almost exclusively girls. Mutations in the X-linked methyl-CpG-binding protein 2 gene (MECP2) have been found to be a cause. In order to study the spectrum of MECP2 mutations in Chinese patients, we employed PCR and sequencing of the coding region of MECP2 gene in 31 Chinese cases of classical sporadic RTT. Mutations in MECP2 were found in about 55%. Twelve different mutations in exon 3 were identified in 17 of these 31 patients; two of these are novel. A novel missense variant was detected in the C-terminal region in a patient and her father who was normal. In addition, there was a single nucleotide variant in the 3'UTR. PMID- 12111644 TI - Low frequency of MECP2 mutations in mentally retarded males. AB - A high frequency of mutations in the methyl CpG-binding protein 2 (MECP2) gene has recently been reported in males with nonspecific X-linked mental retardation. The results of this previous study suggested that the frequency of MECP2 mutations in the mentally retarded population was comparable to that of CGG expansions in FMR1. In view of these data, we performed MECP2 mutation analysis in a cohort of 475 mentally retarded males who were negative for FMR1 CGG repeat expansion. Five novel changes, detected in seven patients, were predicted to change the MECP2 coding sequence. Except for one, these changes were not found in a control population. While this result appeared to suggest a high mutation rate, this conclusion was not supported by segregation studies. Indeed, three of the five changes could be traced in unaffected male family members. For another change, segregation analysis in the family was not possible. Only one mutation, a frameshift created by a deletion of two bases, was found to be de novo. This study clearly shows the importance of segregation analysis for low frequency mutations, in order to distinguish them from rare polymorphisms. The true frequency of MECP2 mutations in the mentally retarded has probably been overestimated. Based on our data, the frequency of MECP2 mutations in mentally retarded males is 0.2% (1/475). PMID- 12111645 TI - A novel polymorphism in exon 11 of the WKL1 gene, shows no association with schizophrenia. AB - A missense mutation in exon 11 of the WKL1 gene on chromosome 22 was found to be associated with cases of catatonic schizophrenia in a single large pedigree. We have screened exon 11 of the WKL1 gene in 174 cases of schizophrenia, including cases of 22 cases of catatonic schizophrenia, but could not detect the previously reported mis-sense mutation. However in exon 11, we observed an insertion/deletion polymorphism, one-missense substitution and two synonymous substitutions. In addition, we also identified a nucleotide substitution in intron 11. All these polymorphisms appeared to be in complete linkage disequilibrium with one another. The polymorphisms were also identified in a UK pedigree with schizophrenia, however the polymorphisms did not segregate with the disease. To test for potential association between these polymorphisms and schizophrenia we sequenced an equal number of UK control individuals who were free of all psychiatric symptoms and had negative family histories for mental illness; the frequency of the insertion/deletion polymorphism was not significantly different in schizophrenia cases (42 out of 348 chromosomes, allele frequency 12%) compared to normal controls (40 out of 356 chromosomes, allele frequency 11%). The insertion/deletion was found to be in Hardy Weinberg equilibrium in both the schizophrenic and control groups. The insertion/deletion is composed of repeated sequence from exon 11 and intron 11 and is predicted to affect WKL1 protein structure. PMID- 12111646 TI - Mutations of Cx26 gene (GJB2) for prelingual deafness in Taiwan. AB - Mutations in the Cx26 (GJB2) gene have been shown to be responsible for a major part of autosomal recessive non-syndromic inherited prelingual deafness. We have sequenced the coding region of GJB2 gene from 169 Taiwanese patients with prelingual deafness and 100 unrelated normal individuals. In the deaf patients, three mutations were found: two novel mutations, 551G-->A, and 299-300delAT, and one previously described mutation, 235delC. Four previously reported polymorphisms, 79G-->A, 109G-->A, 341A-->G, and 608T-->C, were also found in both deaf patients and normal individuals and one new possible polymorphism, 558G-->A, which was only found in a patient. Interestingly, we did not find the 35delG allele, which is commonly found in the Caucasian population, either in the patients or in normal individuals we examined. Our data also showed 235delC to be the most common type of mutation found in Cx26 mutants (approximately 57%). Therefore, based on our findings, we have developed a simple molecular test for the 235delC mutation and it should be of considerable help to those families to understand the cause of their children having the prelingual deafness. PMID- 12111647 TI - Management of disease-related anemia in patients with multiple myeloma or chronic lymphocytic leukemia: epoetin treatment recommendations. AB - Multiple myeloma (MM) and chronic lymphocytic leukemia (CLL) patients often develop anemia due to the disease process and effects from disease therapy. Blood transfusion, the established treatment, has an immediate effect in improving patients' hemoglobin levels. However, this effect is transient and transfusion is associated with several risks, including infections and mild to life-threatening immunologic reactions. A newer option is recombinant human erythropoietin (epoetin); a biological treatment that leads to increased hemoglobin levels over an extended time without the risks of blood transfusion. Extensive evidence has shown that epoetin is effective in the treatment of cancer-associated anemia. An international expert panel met to develop treatment recommendations for the use of epoetin in MM and CLL patients. Based on the available data, it is recommended that treatment be initiated only after other possible causes of anemia are eliminated. Epoetin should be administered to any patient with hemoglobin < or=10 g/dl. Patients with hemoglobin 10-12 g/dl should receive epoetin if they suffer from significant symptoms of anemia and/or have progressively decreasing hemoglobin values. Dosage should be initiated at 10 000 IU three times/week or 40 000 IU once/week and be titrated to maintain hemoglobin at 12 g/dl. Nonresponsive patients (<1 g/dl increase over four weeks) may have their dose increased to 20 000 IU three times/week or 60 000 IU once/week, respectively. Epoetin treatment should be discontinued if there is no response to the increased dosage, or hemoglobin >14 g/dl. Treatment should resume for patients who exceed 14 g/dl, at a reduced dosage, if their hemoglobin falls below 12 g/dl. PMID- 12111648 TI - A novel treatment approach for low grade lymphoproliferative disorders using PKC412 (CGP41251), an inhibitor of protein kinase C. AB - INTRODUCTION: PKC412 (formally CGP41251) selectively inhibits protein kinase C (PKC) isoforms and has been shown to be cytotoxic to malignant cells in vitro. We have undertaken a single centre, open-label, multi-dose, exploratory Phase II clinical trial of PKC412 in patients with CLL and low grade NHL. METHODS: Thirteen CLL patients and eight stage IV NHL patients were treated at three oral dose levels of 25, 150 and 225 mg/day for 14 days. RESULTS: There was a median decrease of 29.4% in the lymphocyte count in 11 out of 18 patients with circulating disease following treatment. Two NHL patients without circulating disease showed loss of immunophenotypic evidence of marrow disease and a third showed an improvement in blood counts and transfusion requirements. Adverse events were mostly gastrointestinal (16 patients) requiring little or no intervention. In nine patients there was an asymptomatic rise in serum amylase and/or transaminases. Asymptomatic hyperglycemia was also observed in eight patients. All returned to normal following cessation of treatment. In 14 out of 20 patients total PKC activity measured in peripheral blood and/or bone marrow lymphocytes was reduced during treatment to a mean of 54% of pre-treatment level. CONCLUSION: PKC412 is safe, well tolerated and reduces the tumor load in chronic B-cell malignancies. Inhibition of PKC offers a novel approach to the chemotherapy of B-cell malignancies. PMID- 12111649 TI - The myelodysplastic syndromes: analysis of prognostic factors and comparison of prognostic systems in 128 Chinese patients from a single institution. AB - INTRODUCTION: Although retrospective analysis were frequently undertaken, and many prognostic systems for myelodysplastic syndromes (MDS) have been proposed worldwide, few such studies have been performed and the effectiveness of different scoring systems have not yet been verified in independent patient populations in China. The aim of this single center study was to evaluate the prognostic factors and compare the prognostic scoring systems in Chinese patients with MDS. MATERIALS AND METHODS: One hundred and twenty-eight patients diagnosed as primary MDS in our Institution were studied retrospectively to identify significant prognostic factors and to assess the predictive value of 11 previously described prognostic systems, including French-American-British (FAB) classification, World Health Organization (WHO) classification, Mufti, Sanz, Morra, Aul, Oguma, Toyama, Morel and international prognostic scoring system (IPSS). RESULTS: The median age of the patients was 50 years (range 13-82). The 2 and 5-year survival rate of the patients were 55.22+/-4.90% and 26.09+/-6.36% respectively, with a median survival of 31 months (range 1-127 months). Fifty patients (39.1%) had progressed to acute leukemia (AL) with a median time of 8 months (range 1-43 months). Major independent variables indicated by multivariate analysis were the percentage of bone marrow (BM) blast cells and complex karyotype aberrations for survival (P=0.042 and 0.042, respectively) and only the percentage of BM blast cells for AL transformation (P=0.023). All the systems except Mufti scores successfully discriminated risk groups concerning both survival and AL evolution, especially in the high risk group, ranging from 10 to 20 months and from 4 to 7 months, respectively. The FAB and WHO classification, as well as Sanz, Oguma, Morel and IPSS possessed lower P value (P<0.0001) than that of the rest scoring systems. CONCLUSION: The patients in our study were younger than these of the Western population, whereas the survival and AL transformation ratio were comparable to these previous studies. The BM blast proportion and complex chromosomal defects were highly significant for predicting outcome in MDS patients. Most investigated systems effectively stratified patients into groups with different life expectancies and identified a subset of patients with poor clinical outcome. The FAB, WHO classification, as well as Sanz, Oguma, Morel and IPSS scoring systems were more applicable for predicting survival and leukemia progression. PMID- 12111650 TI - Epidemiology of hairy cell leukemia in Iceland. AB - INTRODUCTION: Hairy cell leukemia (HCL) is a rare B-cell lymphoproliferative disorder. Previous epidemiological studies have mainly focused on cases derived from single institutions or from localized cancer registries. This is the first study in which all cases diagnosed nationwide over a long period of time in a well defined population are analysed. We report the epidemiology of all HCL patients in Iceland, their clinical characteristics, treatment and follow-up. PATIENTS AND METHODS: : All patients diagnosed with HCL in Iceland over a 20 year period, were included in this study. Data was collected retrospectively. RESULTS: Sixteen patients, 13 males and three females were diagnosed with HCL in Iceland from 1981-2000, giving a mean incidence of 4.7/million/year (95% CI: 2.7-7.6) in the population 20 years and older. Eleven patients were treated with a purine analogue, 10 of whom achieved CR. One other patient obtained CR following splenectomy and IFN, giving a total CR rate of 69%. Three other patients (19%) obtained PR, giving a total response rate of 88%. One patient had a variant of HCL and did not respond to any therapy and one patient died of sepsis before any chemotherapy could be given. Six patients with HCL have died, one from complications of HCL. Three patients developed a second malignancy (19%). CONCLUSIONS: The mean incidence of HCL in Iceland is 4.7/million/year. This is slighty higher than the reported incidence in England and Wales, although not significantly higher. The incidence is based on a nationwide information from a well defined stable and racially homogenous island population. Other results are in accordance with previously published studies. PMID- 12111651 TI - Platelet kinetic study in patients with idiopathic thrombocytopenic purpura (ITP) refractory or relapsing after corticosteroid treatment. AB - BACKGROUND: A platelet kinetic study (PKS) is not indicated in the evaluation of adult patients with idiopathic thrombocytopenic purpura (ITP) at presentation. However, in ITP patients refractory to or relapsing after corticosteroid therapy, its appropriateness is considered uncertain. METHODS: We prospectively performed a PKS with (111)In oxine-labeled autologous platelets in 93 consecutive adult ITP patients failing steroid treatment. RESULTS: In 22 patients (24%) a primary condition accounting for thrombocytopenia was identified (17 with myelodysplastic syndrome and three aplastic anemia). Non-ITP patients had significantly longer platelet circulating life span (P=0.0001), lower splenic platelet uptake (P=0.008) and higher liver platelet uptake (P=0.05) compared to 71 patients with confirmed ITP. Among ITP patients with platelets persistently <50 x 10(9)/L, splenectomy was considered in 48 cases. In 23 (48%) it was prospectively excluded because of platelet life span > or = 7 days (11 cases), no splenic platelet uptake together with high liver uptake (10 cases), or both conditions (two cases). Splenectomy was successfully carried out in the other 25 patients, obtaining a response rate of 100% (22 complete responses; three partial responses). Persistent relapse occurred in six of 25 (24%) splenectomized patients after a median of three months (range 1-8). PKS parameters were not able to predict post-splenectomy relapse, although relapsed patients had lower splenic/hepatic platelet uptake ratio (2.6 in relapsed vs 4.9 in persistently responsive patients; P=0.08). CONCLUSIONS: It was concluded that in patients with chronic ITP failing steroid therapy, some PKS parameters may be prospectively used to increase the short term success rate of splenectomy. PMID- 12111652 TI - E2A/HLF fusion gene in an acute lymphoblastic leukemia patient with disseminated intravascular coagulation and a normal karyotype. AB - INTRODUCTION: Disseminated intravascular coagulation (DIC) is a rare event in acute lymphoblastic leukemia (ALL). However, it has been described in a few cases of pre-B ALL with translocation t(17;19)(q22;p13) which results in the fusion of E2A gene with sequences of HLF gene. Here, we report a case of pre-B ALL with DIC and an apparently normal karyotype by R banding. MATERIALS AND METHODS: Fluorescent in situ hybridization (FISH) studies were performed on bone marrow cells from the patient at presentation and after three months of therapy. RT-PCR was used to detect the E2A-HLF transcript. The type of rearrangement was characterized by sequencing. RESULTS: The t(17;19)(q22;p13) was detected by FISH analysis. The fusion E2A-HLF was amplified by RT-PCR and sequenced, giving a type I rearrangement with a long insertion (146 nucleotides) between E2A exon 13 and HLF exon 4. CONCLUSION: While translocation t(17;19) is undetectable by R-banding technique, it can be detected with FISH and amplified with RT-PCR. Therefore, systematic molecular investigations should be conducted for all patients with pre B ALL associated with DIC, in order to appreciate the incidence and the prognostic value of this rare abnormality. PMID- 12111653 TI - Amplification of a novel c-Kit activating mutation Asn(822)-Lys in the Kasumi-1 cell line: a t(8;21)-Kit mutant model for acute myeloid leukemia. AB - INTRODUCTION: A subset of AML-M2/M4Eo patients has been shown to carry c-kit mutations suggesting that myelomonoblastic leukemia cells, disrupting core binding factor through t(8;21) or inv(16) chromosomal rearrangements, have a common differentiation stage suitable to c-kit mutation. In rare core binding factor leukemia patients an increased dosage of a mutated Asp816(Tyr/Val) kit allele is achieved through nonrandom duplication of chromosome 4 where the c-kit gene is located. MATERIALS AND METHODS: The c-kit gene was studied in the core binding factor leukemia cell line Kasumi-1 with t(8;21) by fluorescence in situ hybridization and mutation analysis. The dosage of Asn822(Lys) mutated allele was evaluated by fluorescence semiquantitative PCR. The correct membrane homing of KIT receptor and its activating status was analysed by immunofluorescence and Western blotting respectively. RESULTS: We identified in the Kasumi-1 cell line a novel Asn822(Lys) ligand-independent c-kit activating mutation and demonstrated by semiquantitative PCR that the mutated allele is about fivefold amplified compared to the normal allele. Fluorescence In Situ Hybridization analysis revealed that c-kit amplification maps to minute 4cen-q11 derived marker chromosome, often carrying duplicated signals, which are unequally distributed in the cell population. The Asn822(Lys) mutation affects a highly conserved codon within the tyrosine kinase activation loop leading, likewise the Asp(816) mutants, to constitutive ligand-independent activation of the KIT receptor. DISCUSSION: Results obtained point to the Kasumi-1 cell line as powerful in-vitro model for further investigation of altered KIT signal transduction pathways in acute myeloid leukemia with core binding factor rearrangements and a useful tool for pharmacological therapeutic targeting. PMID- 12111654 TI - Hodgkin's disease primarily involving the oropharynx: case report and review of the literature. AB - We report a rare case of Hodgkin's disease primarily involving the oropharyngeal region. The patient presented with stage IIEA disease with a favorable response to treatment. Our literature search revealed that this disease affected rarely the lymphoid tissues of Waldeyer's ring. In this atypical location, tonsil and nasopharynx represent the most frequently involved sites. The involvement of the posterior wall of the oropharynx by Hodgkin's disease has been reported only once so far. When Waldeyer's ring is involved, the disease is usually localized, with occasional involvement of cervical lymph nodes. In this location, mixed cellularity is the most frequent histologic subtype and the Epstein-Barr virus is found in the majority of cases. This report is followed by a discussion on the clinical and pathological features, the implication of Epstein-Barrs virus and a literature review for Hodgkin's disease involving Waldeyer's ring. PMID- 12111656 TI - [Prevalence of asymptommatic coeliac disease in children and adults in the Dresden region of Germany]. AB - BACKGROUND AND OBJECTIVE: Coeliac disease (CD) can be present without any, only a few, or many symptoms. Since asymptomatic CD can have the same complications and also carries the same risk for malignant disease as clinically typical CD inadequately treated by diet, early diagnosis is essential. The prevalence of asymptomatic CD in the Dresden region was determined by antibody screening. At the same time the sensitivity and specificity of the different antibodies were calculated. MATERIAL AND METHODS: Anti-gliadin and endomysium antibodies and total IgA content were measured in the serum of 3004 children (group A), aged 5 12 years, and of 4313 blood donors (group B), aged 17-64 years. Small-intestine biopsies were recommended if either (1) endomysium antibodies (EmA) or (2) anti gliadin antibodies (ACA) and clinical symptoms or (3) AGA-IgG in the presence of total IgA deficiency and clinical symptoms had been demonstrated. RESULTS: EmA were demonstrated in 0.17% of group A and in 0.28% of group B. But AGA were found much more frequently (group A: 3.89%, group B: 3.76%). The number of cases of CD confirmed by biopsy indicated a prevalence of asymptomatic CD of 1 in 500 children and 1 in 540 adults. Sensitivity and specificity of EmA were significantly higher than those of AGA. CONCLUSION: Compared with a previous study on the prevalence of clinically typical CD in the same region, the present investigation indicates a four-fold higher prevalence of asymptomatic CD. Coeliac specific antibodies should, therefore, be measured much more widely in the presence of certain symptoms and risk factors. While in adults the measurement of EmA is sufficient to provide the indication for a small-intestine biopsy, both EmA and AGA should be determined before a biopsy is undertaken in children. PMID- 12111655 TI - [Prevention of asthma in childhood]. AB - SUMMARY: Environmental factors during early childhood seem to influence the onset of asthma, a disease that affects 10% of all German children, respectively. Prevention strategies are needed to reduce the burden of the disease. Breast feeding and the avoidance of environmental tobacco smoke exposure are recommended for primary prevention. Furthermore, secondary prevention measurements are suggested to reduce the onset of symptoms, once the disease is present. PMID- 12111657 TI - [Poisoning with a podophyllin-containing wart-treating tincture]. AB - HISTORY AND ADMISSION FINDINGS: A 57-year-old depressive and alcohol-dependent man was admitted because of frequent nausea and vomiting and abdominal complaints after he had ingested 40 ml of a tincture for treating warts. He was under the influence of alcohol, but normally oriented and without contributory findings other than his gastrointestinal complaints and tachypnea. INVESTIGATIONS: Transaminases were raised (GOT 1197 U/l, GPT 170 U/l, gammaGT 150 U/l, LDH 2047 U/l), as were creatine phosphokinase (426 U/l) and ferritin (12 200 ng/ml). Platelet count was 36000 mm3, Leucocytes count 11 700/mm (3). Gastroscopy showed marked mucosal necrosis along the entire esophagus and the pulled-up small intestine (state after gastrectomy). DIAGNOSIS, TREATMENT AND COURSE: The patient became comatose within 5 hours, acidotic, oliguric, required ventilation and went into severe shock. The symptoms and the fact that podophyllin (pod.) was the main agent in the wart preparation confirmed the suspicion of pod. poisoning. Symptomatic and intensive care measures stabilized his critically grave condition. He was extubated on the 7th day after ingestion and on the 10th day was discharged at his own request in a relatively good general state. When he was re-admitted after 4 weeks he was without psychiatric symptoms but deeply depressed, and he had signs of a polyneuropathy in all limbs, typical of pod. toxin poisoning. CONCLUSION: Pod. toxin, a spindle poison, is the toxic agent of pod., the resin from the roots and rhizomes of various Berberis plants. While the potential toxicity of the resin, taken either orally or applied externally, has been long known, the poorly definied raw product is still being added to anti wart tinctures. PMID- 12111658 TI - [Frequency adaptive pacing in patients with chronotropic incompetence--case report]. PMID- 12111659 TI - [Frequency adaptive pacing in patients with chronotropic incompetence- diagnosis]. PMID- 12111660 TI - [Frequency adaptive pacing in patients with chronotropic incompetence--therapy]. PMID- 12111663 TI - [Respiratory effects of leptin]. PMID- 12111664 TI - [Previous history in chronic reflux esophagitis]. PMID- 12111665 TI - [Which antihypertensive therapy is indicated in type 2 diabetic patients?]. PMID- 12111667 TI - Powerful regression-based quantitative-trait linkage analysis of general pedigrees. AB - We present a new method of quantitative-trait linkage analysis that combines the simplicity and robustness of regression-based methods and the generality and greater power of variance-components models. The new method is based on a regression of estimated identity-by-descent (IBD) sharing between relative pairs on the squared sums and squared differences of trait values of the relative pairs. The method is applicable to pedigrees of arbitrary structure and to pedigrees selected on the basis of trait value, provided that population parameters of the trait distribution can be correctly specified. Ambiguous IBD sharing (due to incomplete marker information) can be accommodated in the method by appropriate specification of the variance-covariance matrix of IBD sharing between relative pairs. We have implemented this regression-based method and have performed simulation studies to assess, under a range of conditions, estimation accuracy, type I error rate, and power. For normally distributed traits and in large samples, the method is found to give the correct type I error rate and an unbiased estimate of the proportion of trait variance accounted for by the additive effects of the locus-although, in cases where asymptotic theory is doubtful, significance levels should be checked by simulations. In large sibships, the new method is slightly more powerful than variance-components models. The proposed method provides a practical and powerful tool for the linkage analysis of quantitative traits. PMID- 12111669 TI - A note on the calculation of empirical P values from Monte Carlo procedures. PMID- 12111668 TI - Comprehensive detection of genomic duplications and deletions in the DMD gene, by use of multiplex amplifiable probe hybridization. AB - Duplications and deletions are known to cause a number of genetic disorders, yet technical difficulties and financial considerations mean that screening for these mutations, especially duplications, is often not performed. We have adapted multiplex amplifiable probe hybridization (MAPH) for the screening of the DMD gene, mutations in which cause Duchenne muscular dystrophy (DMD) and Becker muscular dystrophy. MAPH involves the quantitative recovery of specifically designed probes following hybridization to immobilized genomic DNA. We have engineered probes for each of the 79 exons of the DMD gene, and we analyzed them by using a 96-capillary sequencer. We screened 24 control individuals, 102 patients, and 23 potential carriers and detected a large number of novel rearrangements, especially small, one- and two-exon duplications. A duplication of exon 2 alone was the most frequently occurring mutation identified. Our analysis indicates that duplications occur in 6% of patients with DMD. The MAPH technique as modified here is simple, quick, and accurate; furthermore, it is based on existing technology (i.e., hybridization, PCR, and electrophoresis) and should not require new equipment. Together, these features should allow easy implementation in routine diagnostic laboratories. Furthermore, the methodology should be applicable to any genetic disease, it should be easily expandable to cover >200 probes, and its characteristics should facilitate high-throughput screening. PMID- 12111671 TI - Differentiating neurological disorders from psychiatric and the psychological impact of neurological diseases. PMID- 12111672 TI - Depression, dementia, and pseudodementia. PMID- 12111673 TI - Dealing with patients who have medically unexplained symptoms. PMID- 12111674 TI - Neurological effects of psychopharmacological agents. PMID- 12111675 TI - Psychiatric effects of neuropharmacological agents. PMID- 12111676 TI - Neurological disorders and depression. PMID- 12111677 TI - Hand and body position during locomotor behavior in the aye-aye (Daubentonia madagascariensis). AB - Aye-ayes (Daubentonia madagascariensis) have unique hands among primates, with extraordinarily long fingers in relation to body size. These long digits may be vulnerable to damage from forces during locomotion, particularly during head first descent-a locomotor mode that the aye-aye utilizes frequently. Previous behavioral studies of aye-aye locomotion reported that Daubentonia must curl its fingers during horizontal quadrupedalism and/or descent to reduce potential stresses on its long fingers. To test this hypothesis, we examined hand and body position in three captive adult aye-ayes while they walked quadrupedally on horizontal and oblique branches. Substantial variation in hand position was observed among individuals for each substrate orientation. While hand postures with curled fingers were preferred by one individual during descent, they were not preferred by the other two individuals, contrary to our expectations. Differences in body position were more consistent among all three individuals. The angle of the body relative to the substrate was significantly reduced during descent (8.4 degrees ) compared to horizontal locomotion (16.9 degrees ). These results suggest that changes in body position, rather than hand position, may help reduce stresses on the digits. A biomechanical model is proposed that demonstrates how a reduction in the body angle in relation to substrate may act to move the center of mass more caudally. This mechanism of moderating loads by altering body position, rather than hand position, may represent an important functional aspect of arboreal locomotion in aye-ayes and other primates. PMID- 12111678 TI - Squirrel monkey chuck call: vocal response to playback chucks based on acoustic structure and affiliative relationship with the caller. AB - Adult female squirrel monkeys (genus Saimiri) that are socially familiar often exchange the chuck vocalization, which differs acoustically across individuals. We used behavioral observations, vocalization playback experiments, and analysis of the acoustic properties of vocalizations to investigate the effect of caller identity and acoustic structure on vocal response to playback chucks in two all female social groups (n=10 females). Females were most likely to respond with a chuck to the playback chucks of their closely affiliated partners compared to those of nonaffiliated group members. This shows for the first time that the chuck stimulus alone is sufficient to elicit a chuck response from a female's affiliated partner. Additionally, females responded with a chuck mostly to familiar playback chucks from their own group and least to playbacks of silent controls. Unfamiliar playback chucks from the same species and a different squirrel monkey species elicited chuck responses intermediate between familiar chucks and silent controls. Post-hoc discriminant function analyses provide preliminary evidence that females are most likely to respond to unfamiliar chucks when those chucks are close in acoustic structure to familiar chucks from their own social group. These results provide a provisional explanation for error in the squirrel monkey signal processing system, in which unfamiliar chucks never heard before nevertheless elicit a chuck response if they are similar in acoustic structure to familiar group chucks. PMID- 12111679 TI - Attachment and social preferences in cooperatively-reared cotton-top tamarins. AB - In many primate species a close attachment between mother and infant provides a secure base for the infant when the infant is frightened or under stress. In cooperatively breeding primates infant carrying is divided among several individuals in the group, with the mother often doing little more than nursing. In these species it is not clear which individual would best serve as a secure base for the infant. We studied eight infant cotton-top tamarins from birth through 20 weeks of age, noting who carried the infant during the first 100 days, who transferred food with the infants, and, as infants became independent, with whom they associated during social play and affiliative behavior. From week 9 to week 20, when infants were independent of carriers most of the time, we presented families with six trials (once every 2 weeks) with a threatening stimulus (a human dressed in a lab coat and wearing an animal mask). Infants played primarily with their twin or youngest sibling and had affiliative interactions with many family members. However, in fearful situations, infants ran to those who had carried them and transferred food with them the most-their father or oldest brother (never to the mother). Although adults increased rates of mobbing calls in response to the threat, infants significantly reduced their vocalization rate. For these cooperatively breeding monkeys, the attachment object for infants is the family member that invested the most effort in carrying the infant and transferring food with the infant. These results parallel and extend results from bi-parental infant care species in which the father serves as the primary attachment figure. PMID- 12111680 TI - Female reproductive parameters and fruit availability: factors determining onset of estrus in Japanese macaques. AB - This study examined the onset of estrus, nutritional conditions during the mating season, and female reproductive parameters in a troop of unprovisioned Japanese macaques (Macaca fuscata fuscata). The mean interbirth interval (IBI) was 25.1 mo, suggesting a 2-yr birth cycle. The mean IBI following the death of an infant before the mating season was only 19.8 mo, whereas the mean IBI was 27.8 mo when the infant survived. Estrus occurred during the time of the greatest food quality, in the autumn (i.e., mating season). A female whose infant did not survive the mating season tended to come into estrus more readily than one with a surviving infant. Only one of the 19 females whose infants survived came into estrus, whereas 43 of the 58 (74.1%) females whose infants did not survive came into estrus. Fruit conditions during the mating season each year were evaluated by the seed crop of the two species (Fagus crenata and Zelkova serrata) that constituted the main food source for the macaques; fruiting levels were classified as "high-fruit" or "low-fruit." Females came into estrus more often during high-fruit mating seasons than during low-fruit mating seasons: 36 (97.3%) of 37 females came into estrus during high-fruit mating seasons, whereas eight (20%) of 40 females came into estrus during low-fruit mating seasons. Although females came into estrus under both high- and low-fruit conditions during the mating season, estrus seldom occurred when females had a surviving infant, and it appears that the absence of a surviving infant is an almost essential variable for the onset of estrus. When females with a surviving infant were excluded from the analysis because they were unlikely to come into estrus, all 35 remaining females (100%) came into estrus during high-fruit mating seasons, whereas only eight (34.8%) of 23 females came into estrus during low-fruit mating seasons. Therefore, the onset of estrus is strongly affected not only by the survival of infants through the mating season, but also by fruiting conditions, especially the availability of high-quality food. PMID- 12111681 TI - Genetic assessment of a white-collared x red-fronted lemur hybrid zone at Andringitra, Madagascar. AB - We examined a purported lemur (Eulemur fulvus rufusxE. albocollaris) hybrid zone at Andringitra, Madagascar, using sequences from five genes (one mitochondrial gene (d-loop) and four nuclear introns (hemopexin, malic enzyme, ceruloplasmin, and microsatellite 26 flanking region)), from 60 individuals (E. albocollaris (n = 16), E.f. rufus (n = 14), E. collaris (n = 9), and purported hybrids from Andringitra (n = 21)). Diagnostic (d-loop and microsatellite 26) and private sites (all other genes) were found in all gene regions for E. albocollaris and E.f. rufus. Also, private sites were found for the purported hybrid population in two gene regions (d-loop and ceruloplasmin). When the putative hybrids were examined for diagnostic and private markers, 18 of 21 were found to contain markers from both E. albocollaris and E.f. rufus populations. The remaining three individuals were found to contain only markers for E. albocollaris. These results indicate that the population at Andringitra is a hybrid population between E. albocollaris and E.f. rufus. PMID- 12111683 TI - Excretion and measurement of estradiol and progesterone metabolites in the feces and urine of female squirrel monkeys (Saimiri sciureus). AB - The first objective of the present study was to determine the metabolic form and rate of excretion of ovarian hormone metabolites in the urine and feces of female squirrel monkeys injected with radiolabeled progesterone (Po) and estradiol. The major portion of the urinary metabolites of both hormones was excreted within 16 24 hr post-injection. Estrogen and Po isotopes in feces exhibited an excretion peak at 16 hr post-injection. The majority of recovered radiolabel of both hormones was excreted in feces. Chromatographic separation of fecal extractions indicated that the major estrogen metabolites in feces are in the free as opposed to the conjugated form. The radioactivity and immunoreactivity for estrone and estradiol (E(1) and E(2), respectively) in eluates of fecal samples subjected to celite co-chromatography indicated that both free E(1) and E(2) exist as excretion products in the feces of female squirrel monkeys. The major radioactive peaks for Po metabolites showed peaks in the elution profile at or very near the Po standard, and corresponded with the celite co-chromatography elution profile of Po standard when subjected to enzyme immunoassay (EIA). The second objective was to validate the application of EIA systems to measure fecal metabolites. Reproductive events of one female squirrel monkey across one annual reproductive cycle are described using the endocrine profile generated from fecal steroid assays. Examination of this profile confirmed that longitudinal fecal sampling and steroid hormone metabolite measurement in feces was not only feasible and practical, but accurately detected known reproductive events as well. PMID- 12111682 TI - Mineral content as a basis for food selection by western lowland gorillas in a forest clearing. AB - The forests in northwest Republic of Congo contain a number of herbaceous swamp clearings that provide foraging sites for lowland gorillas (G.g. gorilla). A 10 month study at the Maya Nord clearing (Parc National d'Odzala) showed that feeding activities occupied 72% of the time visiting gorillas spent on the clearing. They fed on four plant species: Enydra fluctuans (Asteraceae), Cyperus sp., Pycreus mundtii, and Rhynchospora corymbosa (Cyperaceae) among the 45 species recorded on the clearing. These clearing food species have higher mineral contents (especially Na and Ca) than the dominant Marantaceae species (Haumania liebrechtsiana) that constituted a staple food plant for gorillas in this forest. They also have higher potassium contents and contain less lignin than non-eaten clearing items/species. Finally, the most actively searched for clearing food (Enydra fluctuans) was characterized by the highest amount of Na and Ca. These results suggest that the mineral content (especially in Na, Ca, and/or K) could determine the feeding selectivity of gorillas at the clearing. They also tend to confirm that the amount of fiber plays a deterrent role in food selectivity, as has been found by many authors. The high density of gorillas in that region could result from the combination of the large areas of Marantaceae forests that provide abundant though monotonous food, and the number of clearings that provide sufficient mineral supplies. Clearings should thus be considered as key habitats for the conservation of gorillas. PMID- 12111684 TI - Distribution, population structure, and conservation of lion-tailed macaques (Macaca silenus) in the Anaimalai Hills, Western Ghats, India. AB - The lion-tailed macaque is an endangered species, and hence it is necessary that the remaining populations in the rainforests of the Western Ghats, India, be located and their habitats assessed for effective conservation. The Anaimalai Hills in the state of Tamil Nadu harbor 31 groups of lion-tailed macaques. However, the rainforest in these hills is highly fragmented. Since lion-tailed macaques are typically arboreal, the groups have become isolated. Two large rain forest complexes in these hills harbor 12 and seven groups, respectively, and the remaining 12 groups inhabit small, isolated forest fragments. Group size ranges from six to 53 individuals, with a mean size of 16.3. In the small forest fragments, the standard deviation (SD) of group size was considerably higher than it was in the larger forest complexes. The disturbed fragments also had a higher variability in group size than the relatively undisturbed habitats. It is believed that fragmentation may impede male migration. We suggest that the fragments be managed in such a way that male migration among groups can be facilitated to overcome the potential effects of isolation. PMID- 12111685 TI - An alternate characterization of hazard in occupational epidemiology: years of life lost per years worked. AB - BACKGROUND: Standardized mortality ratios (SMRs) and other measures of relative risk by themselves may not suffice as descriptors of occupational hazards for many audiences including decision-makers and those at direct risk from hazardous work. To explore other approaches, we calculated excess years of potential life lost and excess lifetime risk for both lung diseases and fatal injuries in a cohort of uranium miners with historical records of exposure to radon gas. METHODS: We used relatively simple life table (SMR) methods and also analyzed lung cancer mortality with Poisson regression methods permitting control for smoking. RESULTS: Among uranium miners hired after 1950, whose all-cause SMR was 1.5, 28 percent would experience premature death from lung diseases or injury in a lifetime of uranium mining. On average, each miner lost 1.5 yr of potential life due to mining-related lung cancer, or almost 3 months of life for each year employed in uranium mining. As a consequence of all excess lung disease and injury risks combined, a year of mining was associated with 5.9 months loss of potential life. For each year actually working underground, miners lost more than 8 months of potential life. When controlled for smoking (and healthy worker effect) with Poisson regression, the estimates for radon-related lung cancer effects were slightly larger. Although chronic disease deaths dominated in excess years of life lost (due to radon, silica and possibly other exposures), more years were lost on average per individual injury death (38 yr), than per excess lung cancer (20 yr) or other lung disease death (18 yr). Fatal-injury dominated the potential years of life lost up to about age 40. CONCLUSIONS: Years of life lost per years employed provides another, more intuitive summary of occupational mortality risk. PMID- 12111686 TI - Occupation and risk of esophageal and gastric cardia adenocarcinoma. AB - BACKGROUND: Adenocarcinomas of the esophagus and gastric cardia have risen dramatically in incidence over the past few decades, however, little research has been conducted on the occupational risk factors for these cancers. METHODS: In this population-based case-control study, lifetime job histories were compared between cases of esophageal adenocarcinoma (n = 283), gastric cardia adenocarcinoma (n = 259), and population controls (n = 689). Odds ratios (OR) and 95% confidence intervals (CI) for ever employment and by duration in various occupational and industrial categories were calculated using unconditional logistic regression. RESULTS: The risk of esophageal adenocarcinoma was elevated for persons ever employed in administrative support (OR = 1.5; 95%CI = 1.0-2.1); financial, insurance, and real estate (OR = 1.6; 95%CI = 1.0-2.4); and health services (OR = 2.2; 95%CI = 1.2-3.9). The risk of gastric cardia adenocarcinoma was increased among transportation workers (OR = 1.7; 95%CI = 1.1-2.6), as well as among carpenters (OR = 1.8; 95%CI = 0.9-3.9) and workers in the furniture manufacturing industry (OR = 2.4; 95%CI = 0.9-6.3). However, we observed few duration-response relations between length of employment in any category and cancer risk. CONCLUSIONS: This study revealed associations of esophageal adenocarcinoma with employment in administrative support, health services, and a category of financial, insurance, and real estate industries, and of gastric cardia adenocarcinoma with transportation and certain woodworking occupations. Some of these findings may be due to the play of chance associated with the multiple comparisons made in this study. Our results suggest that, overall, workplace exposures play a minor role in the etiology and upward trend of esophageal and gastric cardia adenocarcinomas. PMID- 12111687 TI - Population-based study of non-infectious rhinitis in relation to occupational exposure, age, sex, and smoking. AB - BACKGROUND: Many types of occupational exposure are associated with the risk of non-infectious rhinitis. We investigate the risk factors for this association. METHODS: A random population of 2,044 subjects (aged 21-51) answered a questionnaire that was comprised of detailed questions on occupational exposure, nasal complaints, and smoking. NIR was defined as the sensation of nasal blockage and/or attacks of sneezing without having a cold. The incidence rates for NIR among exposed and unexposed were calculated. In the different exposed groups, only NIR with onset after the start of exposure was regarded as exposed. If a subject reported NIR before the relevant exposure started, he/she was excluded from that analysis. Relative risks (RR) were calculated as incidence rate ratios. Odds ratios controlling for smoking, age, and atopy were also calculated. RESULTS: The incidence rate for NIR was 13.5/1,000 person-years. Males exposed to fire fumes (RR 2.5, 95% confidence interval (CI) 1.5-4.1), women exposed to paper dust (RR 2.0, 95% CI 1.5-2.9), and male cleaners (RR 3.1, 95% CI 1.9-5.1) displayed an increased risk of developing NIR. Smoking was associated with an increased risk of NIR for both sexes. CONCLUSIONS: Exposure to several occupational irritants is associated with a higher risk of developing NIR. PMID- 12111688 TI - Association between asbestos exposure, cigarette smoking, myeloperoxidase (MPO) genotypes, and lung cancer risk. AB - BACKGROUND: As observed in tobacco-associated carcinogenesis, genetic factors such as the polymorphic metabolic/oxidative enzyme myeloperoxidase (MPO) could modulate individual susceptibility to asbestos-associated carcinogenesis. METHODS: RFLP-PCR analysis identified the MPO genotypes in 375 Caucasian lung cancer cases and 378 matched controls. An epidemiological interview elicited detailed information regarding smoking history and occupational history and exposures. RESULTS: Asbestos exposure was associated with a significantly elevated risk estimate (OR = 1.45; 95% CI 1.04-2.02). On stratified analysis, we found the MPO genotypes modified the effect of asbestos exposure on lung cancer risk. Specifically, G/G carriers who were exposed to asbestos had an odds ratio (OR) of 1.72 (95% CI; 1.09-2.66), while A-allele carriers (G/A + A/A) exposed to asbestos exhibited a reduced OR of 0.89 (95% CI; 0.56-1.44). The OR was further reduced to 0.73 (0.49-1.06) for A-allele carriers not exposed to asbestos. A similar trend was observed for the joint effects between the MPO genotypes and pack-years smoking. Next, all three risk factors (MPO genotypes, asbestos exposure, and smoking) were analyzed simultaneously for joint effects. Heavy smokers with the G/G genotype and a history of asbestos exposure demonstrated a statistically significant elevated risk estimate (OR = 2.19; 95% CI 1.16-4.11), while the A-allele carriers with the same exposure profile were at a lower risk for lung cancer (OR = 1.18; 95% CI 0.58-2.38). The A-allele genotypes demonstrated similar protective effects for the other three exposure profiles. CONCLUSIONS: For a similar level of exposure to established carcinogens, individuals with the MPO A-allele genotypes appear to have a reduced risk of lung cancer. PMID- 12111689 TI - Occupational determinants of bone and blood lead levels in middle aged and elderly men from the general community: the Normative Aging Study. AB - BACKGROUND: Few studies of the general population have investigated risk factors for elevated levels of lead in bone in relation to occupation. METHODS: Six hundred and fifty six community-exposed men had their bone and blood lead levels measured (by K-X-ray fluorescence). Based on their occupational histories, participants were categorized into those who worked in white-collar (WC) occupations (59%) or blue-collar (BC) occupations (41%). No subjects had worked in a primary lead industry (e.g., smelting). RESULTS: In multivariate regression models that adjusted for age, race, education, smoking, alcohol ingestion and retirement status, BC subjects had tibia and patella lead concentrations that were 5.5 (95% CI: 3.2-7.8) and 6.5 (95% CI: 3.1-9.8) microg/g higher than WC subjects, respectively. Interaction terms pairing race with occupational status indicated that in non-white BC subjects, tibia and patella lead levels were higher still by 11.3 (95% CI: -2 to 24.5) and 20.5 (95% CI: 1.2-39.8) microg/g, respectively. Blood lead levels were low for these mostly retired men (mean [SD]: 6.1 [3.9] microg/g) and in multivariate regression models, occupational status was not a significant predictor of blood lead levels; however, an interaction between race and occupational status was also suggested, with non-white BC subjects having blood lead levels that were predicted to be higher by 4.5 (95% CI: 0.3-8.7) microg/dl. CONCLUSIONS: Bone lead levels are higher in the men who worked in BC occupations even if they have not worked in primary lead-exposed occupations. This effect is markedly stronger in non-white BC workers and suggests an interaction between occupational exposures and race/ethnicity with respect to cumulative exposure to lead. A similar interaction was suggested by models of blood lead levels. PMID- 12111690 TI - Fatal asthma from powdering shark cartilage and review of fatal occupational asthma literature. AB - BACKGROUND: Work-related asthma (WRA) is the most common work-associated respiratory disease in developed countries. METHOD: We report shark cartilage dust as a new potential cause of occupational asthma (OA) in the context of other fatal OA case reports. RESULTS: A 38-year-old white male worked for 8 years in a facility which primarily granulated and powdered various plastics. Sixteen months prior to his death, the plant began grinding shark cartilage. After 10 months of exposure, he reported chest symptoms at work in association with exposure to shark cartilage dust and a physician diagnosed asthma. Six months later, he complained of shortness of breath at work and died from autopsy-confirmed asthma. The latency from onset of exposure to symptoms and from symptom onset to death was shorter than 10 previously reported OA fatalities. CONCLUSION: Recognition of occupational causes and triggers of asthma and removal of affected individuals from these exposures is critical and can prevent progression to irreversible or even fatal asthma. PMID- 12111691 TI - Respiratory protection: associated factors and effectiveness of respirator use among underground coal miners. AB - BACKGROUND: We investigated factors associated with the use of respiratory protection and explored the effectiveness of respirators among coal miners. METHODS: Between 1987 and 1992, respiratory symptoms, smoking, lung function, and dust exposures were assessed longitudinally among 185 underground bituminous coal miners. Self-reported use of respiratory protection was expressed as mean percent time wearing a respirator. RESULTS: Miners' respirator use increased with mean dust concentration, but decreased with tobacco consumption. Increasing age was associated with greater respirator use. Miners who had respiratory symptoms at the initial survey subsequently reported greater use of respirators. A significant protective association was found between the miners' respirator use and FEV(1) levels at both the initial and follow-up surveys. CONCLUSIONS: These results provide additional evidence that respirator use is protective of lung health. When respiratory protection programs are developed, factors that may affect respirator use behavior, such as age, smoking, and respiratory symptoms, should be considered. Future studies of respiratory health will need to consider workers' use of respiratory protection. PMID- 12111692 TI - Occupational burns from oxygen resuscitator fires: the hazard of aluminum regulators. AB - BACKGROUND: There have been over 30 incidents of oxygen resuscitator fires over the last 6 years, causing severe burns to a number of fire fighters, emergency medical service personnel, health care workers, and patients. The National Institute for Occupational Safety and Health (NIOSH) was requested to investigate three such incidents. METHODS: NIOSH conducted site investigations of the incidents, and the requesters also sent the involved oxygen resuscitators to a forensic engineering company for a causal analysis. RESULTS: The investigated fires were associated with aluminum regulators, all from one manufacturer, on compressed pure oxygen cylinders. The investigations indicated that the cause of the fires was an initial small ignition in the high-pressure area of the aluminum regulator, which then consumed itself in a massive burnout. CONCLUSIONS: Aluminum regulators used with high-pressure oxygen systems are subject to rare, but potentially catastrophic combustion in normal use. Replacement of such regulators with those made of more fire-resistant materials or designs, as well as education and improved safety practices are needed to reduce this hazard. PMID- 12111693 TI - Expression of urocortin mRNA and peptide in the human prostate and in prostatic adenocarcinoma. AB - BACKGROUND: Urocortin (UCN) is a recently described neuropeptide member of the CRF family, responsible for the secretion of the proopiomelanocortin-derived peptides from the pituitary gland. Although previous results have demonstrated the synthesis of several neuroendocrine factors in the prostate, no studies have been carried out on the expression of UCN in the human gland. METHODS: UCN expression was evaluated in benign prostatic hyperplasia and prostatic tumor tissues by RT-PCR and immunohistochemistry. RESULTS: UCN mRNA and peptide were demonstrated in all specimens tested. In nonneoplastic tissues, UCN was localized in the secretory luminal epithelial and basal layer cells, in the smooth muscle component of the stroma, and in lymphoid infiltrates. An intense immunostaining was evident in prostate adenocarcinoma cells. CONCLUSIONS: The results of the present study demonstrate for the first time UCN expression in the human prostate and in prostate cancer, and suggest a potential involvement of UCN in prostate physiopathology. PMID- 12111694 TI - Inhibition of proliferation of PC-3 human prostate cancer by antagonists of growth hormone-releasing hormone: lack of correlation with the levels of serum IGF-I and expression of tumoral IGF-II and vascular endothelial growth factor. AB - BACKGROUND: Antagonists of growth hormone-releasing hormone (GHRH) such as JV-1 38 can inhibit androgen-independent prostate cancer directly by several mechanisms and/or indirectly by suppressing growth hormone/insulin-like growth factor-I (GH/IGF-I) axis. To shed more light on the mechanisms involved, the effects of JV-1-38 on PC-3 human prostate cancer were compared with those of somatostatin analog RC-160 in vivo and in vitro. METHODS: Nude mice bearing PC-3 tumors received JV-1-38 (20 microg), RC-160 (50 microg) or a combination of JV-1 38 and RC-160. The concentration of IGF-I in serum and the expression of mRNA for IGF-II and vascular endothelial growth factor (VEGF) in tumor tissue were investigated. RESULTS: In vivo, the final volume of PC-3 tumors treated with JV-1 38 was significantly lowered by 49% (P < 0.01), whereas RC-160 exerted only 30% inhibition (NS), compared with controls. Combined use of both compounds augmented tumor inhibition to 63% (P < 0.001). Serum IGF-I levels were decreased only in mice treated with RC-160. JV-1-38 suppressed mRNA for IGF-II in PC-3 tumors by 42%, whereas RC-160 alone or in combination with JV-1-38 caused a 65% reduction. JV-1-38 and RC-160 used as single drugs decreased the expression of VEGF by 50%, and their combination caused a 63% reduction. In vitro, JV-1-38 inhibited the proliferation of PC-3 cells by 39%. This effect could be partially reversed by addition of IGF-I to the serum-free medium. RC-160 alone did not affect the PC-3 cell growth in vitro, but in combination with JV-1-38 it augmented the antiproliferative effect of the GH-RH antagonist to 72%. Exposure to JV-1-38 in vitro reduced the expression of mRNA for IGF-II in PC-3 cells by 55% but did not change VEGF mRNA levels, whereas RC-160 had no effect. CONCLUSIONS: The antiproliferative effect of JV-1-38 was not associated with the suppression of serum IGF-I and was only partially correlated with the expression of IGF-II and VEGF in PC-3 tumors, suggesting that other mechanisms play a role in the antitumor action of GHRH antagonists. Nevertheless, the stronger inhibition of tumor growth after combined treatment with JV-1-38 and RC-160 indicates that the interference with multiple local stimulatory factors leads to an enhanced inhibition of prostate cancer. PMID- 12111695 TI - Mechanisms of constitutive NF-kappaB activation in human prostate cancer cells. AB - BACKGROUND: Activation of the NF-kappaB transcription factor has been previously demonstrated in two androgen receptor negative prostate cancer cell lines. We wished to extend this work to additional prostate cancer cells and to characterize the mechanisms responsible for constitutive NF-kappaB activation. METHODS: Electrophoretic mobility shift assays were performed to measure NF kappaB DNA-binding activity in prostate cancer cell lines, and immunohistochemistry was performed to detect nuclear localization of NF-kappaB in prostate cancer tissues. Western blot analysis was used to study the status of IkappaBalpha. Transient transfection assays were employed to characterize the contributions of IkappaB kinase (IKK), MAPK kinase kinases (MAPKKKs), androgen receptor (AR), and tyrosine phosphorylation to the constitutive activation of NF kappaB in the prostate cancer cell lines. RESULTS: Constitutive NF-kappaB activity was observed in AR-negative cell lines as well as in the prostate cancer patient samples, but was not present in AR positive cells. A "super-repressor" IkappaBalpha, as well as dominant negative forms of IKKbeta and NF-kappaB inducing kinase (NIK), and tyrosine kinase inhibition were able to suppress NF kappaB activity in the cells with constitutive activation. CONCLUSIONS: The constitutive activation of NF-kappaB observed in prostate cancer cells is likely due to a signal transduction pathway involving tyrosine kinases, NIK, and IKK activation. PMID- 12111696 TI - Mechanisms of the growth inhibitory effects of the isoflavonoid biochanin A on LNCaP cells and xenografts. AB - BACKGROUND: Isoflavones inhibit the growth of some types of tumor cells, including prostate adenocarcinoma. This study used LNCaP cells and xenografts to investigate the mechanisms of the antiproliferative effects of biochanin A, a major isoflavone present in red clover but not soy-derived products. METHODS: LNCaP cells were exposed to varying doses of biochanin A to evaluate viability, DNA synthesis, and DNA fragmentation (TUNEL) analysis. Regulation of gene expression was determined by using Western immunoblotting and cDNA microarrays. Anti-tumorigenic effects were evaluated by using athymic mice with LNCaP flank tumors. RESULTS: Biochanin A induced a dose-dependent inhibition of proliferation and [(3)H]thymidine incorporation that correlated with increased DNA fragmentation, indicative of apoptosis. Western blot analyses of cell cycle regulatory proteins revealed that biochanin A significantly decreased expression of cyclin B and p21, whereas flow cytometry showed that cells were accumulating in the G(0)/G(1) phase. cDNA microarray analyses identified 29 down-regulated genes with six reduced below assay detection limits. Eleven genes were up regulated, including 9 that were undetectable in controls. In mice with LNCaP xenografts, biochanin A significantly reduced tumor size and incidence. CONCLUSION: These results indicate that biochanin A inhibits prostate cancer cell growth through induction of cell cycle arrest and apoptosis. Biochanin A regulated genes suggest multiple pathways of action. Biochanin A inhibits the incidence and growth of LNCaP xenograft tumors in athymic mice. PMID- 12111697 TI - Hormones and prostate cancer: current perspectives and future directions. AB - Prostate cancer is the most commonly diagnosed non-skin cancer in men in most western countries. Despite the high morbidity and mortality from prostate cancer, its etiology remains obscure. Although compelling laboratory data suggest a role for androgens in prostate carcinogenesis, most epidemiologic data on humans are inconclusive. To provide insights and directions for future epidemiologic research on hormones and prostate cancer, this review focuses on current perspectives of serum-based studies and polymorphisms in relevant hormone-related genes. We highlight the importance of methodologic studies and investigations of hormone levels in the prostatic tissue to help clarify the often-contradictory data on serologic studies. We recommend careful analysis and cautious interpretation of studies of genetic markers, including repeats and single nucleotide polymorphisms (SNPs), as false positive and negative results may arise in many current and future studies with limited statistical power and non representative samples from the population. The review also highlights the reasons to perform functional analyses of SNPs, a critical and often under appreciated component of molecular epidemiologic investigations. The time is ripe for large-scale multidisciplinary investigations that incorporate molecular genetics, biochemistry, histopathology, and endocrinology into traditional epidemiologic studies. Such collaboration will lead to a deeper understanding of the etiologic pathways of prostate cancer, ultimately yielding better preventive, diagnostic, and therapeutic strategies. PMID- 12111698 TI - Altered apoptotic gene expression and acquired apoptotic resistance in cadmium transformed human prostate epithelial cells. AB - BACKGROUND: Cadmium is a suspected prostatic carcinogen, although the underlying mechanisms are unclear. To investigate these mechanisms, we performed molecular comparisons between the cadmium-transformed prostate epithelial cell line CTPE and the nontumorigenic parental line RWPE-1. METHODS: Gene expression patterns were compared by using cDNA arrays, RNase protection assays, and Western blots. Apoptosis was analyzed by using flow cytometry to quantify apoptotic nuclei and an enzyme-linked immunosorbent assay method to measure DNA fragmentation. Caspase 3 activity was measured colorimetrically. RESULTS: Among the genes down-regulated in CTPE cells were those encoding several members of the caspase family of apoptotic proteases as well as the apoptotic regulator Bax. Ribonuclease protection assays confirmed global down-regulation of caspase gene expression in CTPE. Decreased Bax expression in CTPE was confirmed by Western blots, which also revealed increased expression of anti-apoptotic Bcl-2. Consistent with these changes, CTPE cells exhibited increased resistance to apoptosis induced by cadmium, cisplatin, and etoposide. CTPE cells also exhibited lower caspase-3 activity vs. RWPE-1 after etoposide treatment. CONCLUSIONS: CTPE cells exhibited altered expression of important apoptotic regulators as well as resistance to several apoptotic stimuli. We hypothesize that acquired apoptotic resistance may be a key aspect of cadmium-induced malignant transformation of prostate epithelial cells and that this may contribute to both tumor initiation and the acquisition of aggressive characteristics subsequent to tumor formation. PMID- 12111699 TI - Down-regulation of (IIIb) and (IIIc) isoforms of fibroblast growth factor receptor 2 (FGFR2) is associated with malignant progression in human prostate. AB - BACKGROUND: Fibroblast growth factors (FGFs) and their receptors (FGFRs) have a critical function in the cellular stroma/epithelium interaction for the development and homeostasis of human prostate. Imbalance in expression of these factors is associated with malignancy in several cancers. METHODS: To quantify the expression of fibroblast growth factor receptor isoforms FGFR2(IIIb), FGFR2(IIIc), FGFR1(IIIc), and fibroblast growth factors FGF2 and FGF7 in normal and tumoral human prostate tissues, and human prostatic epithelial cell lines, we used quantitative real-time polymerase chain reaction. RESULTS: The expression of FGFR2(IIIb) mRNA is down-regulated in 60% of the tumors studied (P < 0.0001). Furthermore, FGFR2(IIIb) is significantly reduced in androgen-independent tumors (AI) compared with androgen-responsive tumors (AD) (P = 0.02). A significant reduction in FGFR2(IIIc) expression is also observed in 80% of tumors (P = 0.001). However, unlike FGFR2(IIIb), the down-regulation of FGFR2(IIIc) is not related to the androgen-independent status (P = 0.09). On the other hand, neither FGFR1(IIIc) nor FGF2 and FGF7 have shown any significant variation in expression between normal and cancerous specimens. CONCLUSIONS: These findings propose that decreased expression of not only FGFR2(IIIb) but also FGFR2(IIIc) isoforms may be a critical step in prostate cancer progression and furthermore suggest that FGFR2(IIIb) expression could be used as a marker for prostate cancer evolution from androgen-dependent to androgen-independent status. PMID- 12111700 TI - Metallothionein isoform 3 expression inhibits cell growth and increases drug resistance of PC-3 prostate cancer cells. AB - BACKGROUND: The third isoform of metallothionein (MT-3) is overexpressed in prostate cancers and PIN lesions. The expression of MT-3 is highly variable but appears to correlate to Gleason score. The goal of the present study was to determine the effect of MT-3 overexpression on the growth of the PC-3 prostate cancer cell line. METHODS: PC-3 cells were stably transfected with either the MT 3 or MT-1E gene. Cell growth was determined by counting DAPI-stained nuclei, drug resistance by the colony formation assay, MT mRNA expression by reverse transcriptase-polymerase chain reaction, and MT protein by immunoblot. RESULTS: PC-3 cells that overexpress the MT-3 gene are growth inhibited compared with either untransfected cells, cells with blank vector, or cells with similar overexpression of the MT-1E gene. Furthermore, increased chemotherapeutic drug resistance occurred in PC-3 clones derived from MT-3- and MT-1E-transfected cells. CONCLUSION: The overexpression of MT-3 can influence the growth and chemotherapeutic drug resistance of the PC-3 prostate cancer cell line. PMID- 12111701 TI - Insulin-like growth factors and risk of benign prostatic hyperplasia. AB - BACKGROUND: Insulin-like growth factors (IGFs) have potent growth mitogenic and anti-apoptotic effects on prostate tissue, whereas IGF binding proteins (IGFBPs) inhibit growth of prostatic tissue. The IGF axis has been implicated in prostate cancer risk, but its role in benign prostatic hyperplasia (BPH) is unclear. METHODS: Plasma levels of IGF-I, IGF-II, IGFBP-1, and IGFBP-3 were determined from the fasting bloods of 206 BPH cases admitted for treatment and 306 randomly selected population controls in Shanghai, China. RESULTS: Relative to the lowest tertile, men in the highest tertile of IGF-I levels had a significantly elevated risk of BPH (odds ratio [OR] = 2.80, 95% confidence interval [95% CI] = 1.60 4.92; P(trend) < 0.001). Results for IGF-I were more pronounced after adjustment for serum androgens. In contrast, men in the highest IGFBP-3 tertile had a significantly reduced risk (OR = 0.40; 95% CI = 0.23-0.69; P(trend) < 0.001). No associations of BPH with IGF-II and IGFBP-1 were observed. CONCLUSION: As has been previously observed for prostate cancer, we found that IGF-I and IGFBP-3 are associated with BPH risk in China. Further investigation is needed to elucidate the role of the IGF axis in BPH etiology. PMID- 12111702 TI - Melatonin and prostate cancer cell proliferation: interplay with castration, epidermal growth factor, and androgen sensitivity. AB - BACKGROUND: Potential modulatory effects of melatonin on the proliferation of androgen-sensitive LNCaP and androgen-insensitive PC-3 and DU 145 prostate cancer cells were reported recently. In this study, we investigated the effects of combined melatonin and castration on LNCaP tumor growth in vivo, the interactions between melatonin and epidermal growth factor (EGF) on LNCaP cell proliferation, and melatonin actions on the proliferation of PC-3 and DU 145 cells. METHODS: Tumor development and growth in castrated nude mice inoculated with LNCaP cells or in intact animals inoculated with DU 145 cells, with or without daily melatonin treatment, were monitored by observation and caliper measurement. MT(1) receptor expression in native or transfected prostate cancer cell lines was examined by immunocytochemistry or 2-[(125)I]iodomelatonin binding. Cyclin D1 expression in LNCaP cells was assessed by Western blotting, and cell proliferation was measured by thymidine incorporation and/or cell count. RESULTS: Melatonin treatment was associated with further decreases in LNCaP tumor incidence and growth rate in castrated nude mice. Melatonin and 2-iodomelatonin (a melatonin receptor agonist) attenuated EGF-stimulated increases in LNCaP cell proliferation and cyclin D1 levels. Melatonin had no effect on the proliferation or growth of MT(1) receptor-expressing DU 145 cells, and of PC-3 cells in which MT(1) receptor protein was undetectable. The proliferation of transfected PC-3 cells expressing MT(1) receptor was unaffected by 2-iodomelatonin. CONCLUSION: Together with previous data, the present results indicate synergistic action of melatonin and castration in inhibiting the growth of androgen-sensitive LNCaP tumor. Androgen-sensitive prostate cancer cell proliferation may be modulated by opposite changes in cyclin D1 levels induced by activated MT(1) and EGF receptors. In androgen-insensitive prostate cancer cells, MT(1) receptor-mediated signal transduction may become defective not only through changes in membrane receptor protein expression and/or functions, but also by means of alterations in downstream postreceptor signaling events. PMID- 12111704 TI - Association of polymorphisms within androgen receptor, 5alpha-reductase, and PSA genes with prostate volume, clinical parameters, and endocrine status in elderly men. AB - BACKGROUND: The aim of this study was to assess the impact of polymorphisms of three genes within the androgen pathway on prostate volume, clinical parameters, and endocrine status. METHODS: Elderly men with lower urinary tract symptoms underwent clinical and endocrine work-up. In parallel, polymorphisms within the 5alpha-reductase gene (SRD5A2 V89L and A49T), the androgen receptor gene (AR; number of CAG repeats), and the prostate specific antigen (PSA) gene (A --> G substitution at position-158) were determined by polymerase chain reaction and restriction-length polymorphism analysis by using DNA from peripheral blood. RESULTS: A total of 190 men (66.5 +/- 9.2 yr) were analyzed. The number of CAG repeats within the AR and the PSA polymorphism revealed no associations to clinical and endocrine parameters. Individuals carrying the mutated SRD5A2 A49T allele (5.3% of the total population) had larger prostates (54.1 vs. 39.3 ml), higher PSA levels (12.2 vs. 4.3 ng/ml), and a 35% reduction in prostatic stroma/epithelial cell ratio. Men with the mutated SRD5A2 V89L gene had lower testosterone levels. CONCLUSIONS: In contrast to prostate cancer, polymorphisms within AR and PSA genes do not seem to be of importance for benign prostatic hyperplasia. Polymorphisms within the 5alpha-reductase gene are interesting biomarkers for the development of benign prostatic hyperplasia and benign prostatic enlargement. PMID- 12111703 TI - Stat3 enhances the growth of LNCaP human prostate cancer cells in intact and castrated male nude mice. AB - BACKGROUND: Prostate cancer frequently progresses from an initial androgen dependence to androgen independence, rendering the only effective androgen ablation therapy useless. The mechanism underlying the androgen-independent progression is unknown. Stat3, a member of the family of signal transducers and activators of transcription, is activated in numerous cancers, including prostate. This study is to investigate the role of Stat3 activation in the growth of prostate cancer cells. METHODS: A constitutively active Stat3 was ectopically expressed in androgen-sensitive LNCaP prostate cancer cells and resulting stable clones expressing activated Stat3 were isolated. The effect of Stat3 activation on LNCaP cell growth in response to androgen in vitro and in vivo was examined. RESULTS: We show that the levels of activated Stat3 are associated with the progression of androgen-independent prostate cancer. Activation of Stat3 in androgen-sensitive LNCaP prostate cancer cells results in enhancement of tumor growth in both intact and castrated male nude mice and enhances androgen receptor mediated prostate specific antigen expression. CONCLUSIONS: These findings demonstrate that intracellular signaling mediated by Stat3 can enhance the growth of androgen-sensitive human LNCaP prostate cancer cells in both intact and castrated male nude mice. PMID- 12111705 TI - Molecular characterization of prostate hyperplasia in prolactin-transgenic mice by using cDNA representational difference analysis. AB - BACKGROUND: Transgenic mice overexpressing the rat prolactin (PRL) gene develop a dramatic enlargement of the prostate gland. The objective of this study was to characterize the molecular mechanisms in the prostate of importance for the prostate hyperplasia seen in these transgenic mice. METHODS: cDNA representational difference analysis (cDNA RDA) was used to isolate differentially expressed transcripts in the prostate hyperplasia of the transgenic mice compared with wildtype littermates. Furthermore, cDNA microarray analysis was used to verify the RDA output. RESULTS: Here we report 10 transcripts, some of them described to be involved in proliferation and apoptosis, which are differentially expressed in the enlarged transgenic prostates compared with controls. CONCLUSION: The identified differentially expressed transcripts presented herein supports molecular similarities between the prostate hyperplasia of PRL-transgenic mice and human BPH that may contribute to explain the molecular basis of prostate hyperplasia. PMID- 12111706 TI - Using multiple cutpoints for the free-to-total prostate specific antigen ratio improves the accuracy of prostate cancer detection. AB - BACKGROUND: Using a single cutpoint for the free-to-total (F/T) prostate specific antigen (PSA) ratio loses important diagnostic information. We evaluated the performance of multiple F/T PSA cutpoints in detecting prostate cancer in men with nonspecific PSA values. METHODS: We extracted sensitivity and specificity data from 12 studies reporting on >or=30 cancer patients with PSA values between 2.0 and 10.0 ng/mL. We calculated stratum-specific likelihood ratios (LR) and areas under the receiver operating characteristic (ROC) curves. RESULTS: Multiple cutpoints for the F/T PSA ratio significantly increased the area under the ROC (0.70) compared with the single investigator-selected cutpoint (0.62), P < 0.004. The LR for the most positive cutpoint stratum (2.62) was significantly higher than the LR for a positive test from the single cutpoint (1.36), P < 0.004. CONCLUSIONS: Using multiple cutpoints increased the discriminating power of the F/T PSA ratio and led to greater probability revisions in the most positive test result strata. PMID- 12111707 TI - Identification of a plasma membrane Ca2+-ATPase in epithelial cells and aposomes of the rat coagulating gland. AB - BACKGROUND: Epithelial cells of the rat coagulating gland secrete a minor fraction of proteins by means of an alternative export mode named apocrine secretion. Thereby, proteins are released by means of membrane bounded blebs or "aposomes" arising from the apical plasma membrane. Little is known about the composition of the aposomal membrane and whether or not proteins located in the apical plasma membrane are integrated into the aposomes. METHODS: To show expression and localization of Ca(2+)-ATPase in rat coagulating gland, reverse transcriptase-polymerase chain reaction, Western blotting experiments, and Ca(2+) ATPase activity assays, as well as immunofluorescence studies were performed. RESULTS: Ca(2+)-ATPase is located in the apical plasma membrane of the epithelial cells of the rat coagulating gland and is also included in the aposomes. Mg(2+) dependent and Mg(2+)-independent Ca(2+)-ATPase activity was observed in coagulating gland primary epithelial cells and tissue. Gene expression of plasma membrane Ca(2+)-ATPase isoforms 1 and 4 was detected in cultured primary epithelial cells of the rat coagulating gland and coagulating gland tissue. CONCLUSIONS: We show for the first time that Ca(2+)-ATPase as an important, functionally active membrane protein of the apical plasma membrane is incorporated into the aposomal membranes and is released from the cells during apocrine secretion process. PMID- 12111708 TI - Embryotoxicity of the pesticide mirex in vitro. AB - Mirex is a pesticide that is environmentally stable, accumulates in body tissues, and is embryo- and feto-toxic at high concentrations in vivo. This study is the first to evaluate the effects of mirex on organogenesis-stage embryos in vitro. Mouse embryos were exposed on gestation day 8.5 for 24 h in whole-embryo culture to mirex at 100, 200, or 400 microg/ml dissolved in xylene and compared with xylene-treated controls (1, 2, or 4 microl/ml, respectively) and untreated controls. Embryos were evaluated for malformations, somite number, total protein content, and visceral yolk sac circulation. Potential embryotoxic mechanisms were evaluated by using PCNA stain for cell proliferation and the TUNEL assay for apoptotic cell death. Mirex-exposed embryos demonstrated increased malformation rates and decreased total embryonic protein contents at > or =200 microg/ml mirex, and decreased somite numbers and VYS circulation at > or =100 microg/ml mirex, compared with xylene-treated controls. There was no difference in PCNA levels or TUNEL staining in mirex-treated embryos compared with xylene-treated controls or untreated controls. Thus, mirex is embryotoxic in vitro to early organogenesis stage mouse embryos at concentrations > or =100 microg/ml, but the effects do not appear to be mediated by changes in cell proliferation or apoptotic cell death. PMID- 12111709 TI - Incision of AP sites in lung cancer patients: a pilot study. AB - DNA base excision repair (BER) removes frequent DNA lesions of either endogenous or exogenous origin. Some indications point to BER defects in lung cancer patients. We have investigated the ability of ten lung cancer patients to repair natural AP sites, the most frequent genetic lesion, using an in vitro assay in which peripheral blood lymphocytes (PBL) extracts incise randomly depurinated plasmid DNA. The median value of repair activity in patients was lower than that of matched controls but the difference did not reach significance. Unlike other BER enzymatic steps, marked defects in incision of AP sites may not be associated with lung carcinogenesis. PMID- 12111711 TI - Monitoring urban air particulate matter (fractions PM 2.5 and PM 10) genotoxicity by plant systems and human cells in vitro: a comparative analysis. AB - Increased incidence of mortality and sickness due to cardiopulmonary complications has been associated with elevated levels of urban air particles (UAP), with an aerodynamic diameter of 10 microm (PM 10) and 2.5 microm (PM 2.5). In the present report alternative plant systems and human cells in vitro are associated with human hazard and genotoxic risk assessment of UAP. The genotoxic activities associated with the coarse (PM 10) and the fine fraction (PM 2.5) of airborne particulates have been analyzed by evaluating micronuclei induction and/or sister-chromatid exchange (SCE) using in vitro models of Daucus carota and HS 27 human fibroblast cell suspensions and Zea mays root meristems. Results show variability in the response of the test systems and indicate that the mutagenicity trend in both plant and human cell cultures was directly correlated to the concentration of carbon-rich particles in the fraction of the PM 2.5 airborne particulates. Moreover, in plant tissues, the frequency of micronuclei and SCE was related to an enhancement of the specific activity of the stress related enzyme peroxidase. PMID- 12111710 TI - Study of chromosome damage in patients with breast cancer treated by two antineoplastic treatments. AB - The frequencies of chromosomal aberrations (CAs) and sister chromatid exchanges (SCEs) were determined in peripheral blood lymphocyte cultures from women with breast cancer treated by chemotherapy (CT) with FEC (5-fluorouracil, epirubicin, and cyclophosphamide) or CMF (cyclophosphamide, methotrexate, and 5-fluorouracil) cocktail in six CT cycles. The number of patients in each CT cycle were from 1 to 3 for SCE and 2 to 5 for CA. Samples were collected before and 48 h after CT. Although the size of the sample was limited and interindividual variability was wide, it appears that a 21-day interval between CT sessions is sufficient for cell recovery. This fact was demonstrated by the reduction in CA and SCE frequencies between cycles in parallel with the unchanged mitotic index and proliferative index values. PMID- 12111712 TI - Anti-mutagenic activity of Phyllanthus amarus Schum & Thonn in vitro as well as in vivo. AB - Methanolic extract of Phyllanthus amarus was tested for its anti-mutagenic activity in Salmonella typhimurium strains TA1535, TA100, and TA102 (Ames test). P. amarus extract was able to inhibit the activation and mutagenicity of 2 acetaminofluorene (2-AAF) and aflatoxinB(1) at concentrations of 0.25-2 mg/plate. It was also found to inhibit mutagenicity induced by direct acting mutagens sodium azide (NaN(3)), N-methyl-N-nitro-N-nitrosoguanidine (MNNG), and 4-nitro-0 phenylenediamine (NPD), at concentrations of 1 mg to 0.25 mg/plate. Urinary mutagenicity produced in rats by benzo[a] pyrene was found to be significantly inhibited by the oral administration of Phyllanthus extract. These results indicate significant anti-mutagenicity of the extract in vitro as well as in vivo. PMID- 12111714 TI - Effects of methionine on selenium embryotoxicity in cultured rat embryos. AB - Effects of methionine, an essential amino acid, on the embryotoxicity of selenium (Se) were examined using the rat embryo culture. Rat embryos at day 9.5 of gestation were cultured for 48 h in the presence of sodium selenite at 10 and 20 microM or sodium selenate at 30 and 100 microM with or without the addition of 1 mM DL-methionine. Selenite at 20 microM or selenate at 100 microM alone increased the incidence of embryonic malformation and inhibited the embryonic growth. The addition of methionine increased the incidence of embryonic malformation at 10 microM of selenite but decreased the incidence of embryonic malformation at 100 microM of selenate. On the other hand, the addition of methionine partially restored the inhibited embryonic growth at 20 microM of selenite or at 100 microM of selenate. It was considered from these results that the methionine availability in the embryonic environment and the oxidation state of Se are critical in Se embryotoxicity. PMID- 12111713 TI - Lack of modification of rat hepatocarcinogenesis by fernane-type triterpenoids, isolated from a Euphorbia genus. AB - Inhibitors of topoisomerases, enzymes that produce an unusual type of DNA damage, are considered as antitumor agents. Recently it has been reported that the fernane-type triterpenoid EC-2 and its hydroxyl derivative, isolated from Euphorbia, are potent topoisomerase II inhibitors. In this study, the modifying effects of EC-2 and EC-4 on the development of putative preneoplastic lesions, glutathione S-transferase placental form (GST-P)-positive foci, in the liver of rats were investigated using a medium-term bioassay system. Fisher 344 male, 6 week-old rats were given a single intraperitoneal injection (200 mg/kg b.w.) of diethylnitrosamine or saline at the beginning of the experiment and subjected to 2/3 partial hepatectomy at the 3rd week. The test compounds were administered five times/week by i.g. gavage at a dose of 1 mg/kg b.w. from 2 to 8 weeks. Quantitation of the numbers and areas per cm(2) of induced GST-P positive foci did not demonstrated any significant differences among the groups and no variation in cell proliferation as indicated by 5-bromo- 2'-deoxyuridine (BrdU) labeling. Our results suggest that EC-2 and EC-4 have no modifying effects on rat hepatocarcinogenesis. PMID- 12111715 TI - Protective effect of tenuazonic acid against dimethyl benz(a)antracene-induced skin carcinogenesis in mice. AB - Anticarcinogenic potential of tenuazonic acid (TA), a mycotoxin isolated from a fungus, Alternaria alternata, on skin tumorigenesis induced by 7,12-dimethyl benz(a) antracene (DMBA) was investigated. Female Swiss albino mice were exposed topically to 100 nmole of DMBA twice weekly for 20 weeks. Another group of animals was treated with 250 microg TA in acetone daily for a period of 1 week, followed by the same dose of TA prior to every application of DMBA. At the end of 14 weeks, all the animals in the group that was exposed to DMBA alone developed tumors, while 40% of the animals in TA-treated group were found to be tumor free. After 20 weeks, there was no further increase in the number of tumor-bearing animals. Results indicated that prior application of TA significantly delayed the onset of tumorigenesis and also reduced the cumulative number of tumors per tumor bearing animals. The present studies reveal the antitumor and protective potential of TA against polycyclic aromatic hydrocarbon induced skin carcinogenesis. PMID- 12111717 TI - Identification of the mouse Loop-tail gene: a model for human craniorachischisis? AB - Neural tube defects are one of the commonest human birth defects, with more than 0.5% of some populations affected. Mouse models are being used in an attempt to identify genes that could be involved in these malformations. Only two mouse mutations are known to lead to craniorachischisis, failure of closure of almost the entire neural tube. Two recent papers report that the gene for one of these, Loop-tail, has now been identified and sequenced.1, 2 It has been given the designation Ltap/Lpp1 and appears to function in floor plate formation. It will be of great interest to investigate the role of this gene in human neural tube defects. PMID- 12111716 TI - p53 and ageing: too much of a good thing? AB - A recent report by Tyner et al.(1) suggests that p53 is bad for longevity. Heterozygotic mice carrying a p53 mutation that apparently enhances the stability of the wild-type protein showed shorter lifespans and faster ageing while also developing fewer tumours. This fits with the idea that cellular ageing is the price paid for better protection against unlimited proliferation of cancer cells. But other work shows that there is a strong positive association between DNA repair-mediated protection against cancer and ageing. So what are we to make of the new data with regard to overall understanding of the mechanisms of ageing? PMID- 12111719 TI - Transient existence of life without a cell membrane: a novel strategy of siphonous seaweed for survival and propagation. AB - Siphonous seaweeds, which constitute a vital component of coral reefs, are structurally simple, single-celled coenocytic macroscopic green algae. Kim et al.1 have recently shown the extraordinary wound-repair and propagation mechanism of one such siphonous green alga--Bryopsis plumosa. Nucleocytoplasmic aggregates, which are released after injury to this plant, are membraneless structures that can survive in seawater for 10-20 minutes, before they are surrounded by a gelatinous envelope. Subsequently, a cell membrane and cell wall are synthesized around each of these aggregates and the resulting individual cells, so formed, develop into new plants. This report represents a significant advance in our understanding of wound response and, more significantly, is probably the first example of transient survival of life without a cell membrane! PMID- 12111718 TI - Inheriting a structural scaffold for Golgi biosynthesis. AB - In animal cells, the Golgi complex undergoes reversible disassembly during mitosis. The disassembly/reassembly process has been intensively studied in order to understand the mechanisms that govern organelle assembly and inheritance during cell division. A long-standing controversy in the field has been whether formation of Golgi structure is template-mediated or self-organizes from components of the endoplasmic reticulum. A recent study1 however, has demonstrated that a subset of proteins that form a putative Golgi matrix can be inherited during cell division in the absence of membrane input from the endoplasmic reticulum. The outcome of this study suggests that a templating mechanism for the formation of Golgi structure may exist. This study has important implications for understanding mechanisms that govern Golgi biogenesis. PMID- 12111720 TI - Myoblast fusion in Drosophila. AB - Somatic muscle formation is an unusual process as it requires the cells involved, the myoblasts, to relinquish their individual state and fuse with one another to form a syncitial muscle fiber. The potential use of myoblast fusion therapies to rebuild damaged muscles has generated continuing interest in elucidating the molecular basis of the fusion process. Yet, until recently, few of the molecular players involved in this process had been identified. Now, however, it has been possible to couple a detailed understanding of the cellular basis of the fusion process with powerful classical and molecular genetic strategies in the Drosophila embryo. We review the cellular studies, and the recent genetic and biochemical analyses that uncovered interacting extracellular molecules present on fusing myoblasts and the intracellular effectors that facilitate fusion. With the conservation of proteins and protein functions across species, it is likely that these findings in Drosophila will benefit understanding of the myoblast fusion process in higher organisms. PMID- 12111721 TI - Understanding Rho/Rac biology in T-cells using animal models. AB - Experiments with cell lines have unveiled the implication of the Rho/Rac family of GTPases in cytoskeletal organization, mitogenesis, and cell migration. However, there have not been adequate animal models to investigate the role of these proteins in more physiological settings. This scenario has changed recently in the case of the T-cell lineage after the generation of animal models for Rho/Rac family members, their regulators, and effectors. These studies have revealed the implication of these GTPases on multiple regulatory layers of T cells, including the coordination of cytoskeletal change, activation of kinase cascades, stimulation of calcium fluxes, and the induction of gene expression. These pathways affect the transition of different T-cell maturation stages, the positive/negative selection of thymocytes, T-cell responses to antigens, and the homeostasis of peripheral T-lymphocytes. Moreover, these animals have revealed interesting cross-talks between Rho/Rac pathways and other signal transduction routes that participate in lymphocyte responses. PMID- 12111722 TI - Patterning the marginal zone of early ascidian embryos: localized maternal mRNA and inductive interactions. AB - Early animal embryos are patterned by localized egg cytoplasmic factors and cell interactions. In invertebrate chordate ascidians, larval tail muscle originates from the posterior marginal zone of the early embryo. It has recently been demonstrated that maternal macho-1 mRNA encoding transcription factor acts as a localized muscle determinant. Other mesodermal tissues such as notochord and mesenchyme are also derived from the vegetal marginal zone. In contrast, formation of these tissues requires induction from endoderm precursors at the 32 cell stage. FGF-Ras-MAPK signaling is involved in the induction of both tissues. The responsiveness for induction to notochord or mesenchyme depends on the inheritance of localized egg cytoplasmic factors. Previous studies also point to critical roles of directed signaling in polarization of induced cells and in subsequent asymmetric divisions resulting in the formation of two daughter cells with distinct fates. One cell adopts an induced fate, while the other assumes a default fate. A simple model of mesoderm patterning in ascidian embryos is proposed in comparison with that of vertebrates. PMID- 12111723 TI - The role of calcium-binding proteins in the control of transcription: structure to function. AB - Transcriptional regulation is coupled with numerous intracellular signaling processes often mediated by second messengers. Now, growing evidence points to the importance of Ca(2+), one of the most versatile second messengers, in activating or inhibiting gene transcription through actions frequently mediated by members of the EF-hand superfamily of Ca(2+)-binding proteins. Calmodulin and calcineurin, representative members of this EF-hand superfamily, indirectly regulate transcription through phosphorylation/dephosphorylation of transcription factors in response to a Ca(2+) increase in the cell. Recently, a novel EF-hand Ca(2+)-binding protein called DREAM has been found to interact with regulatory sequences of DNA, thereby acting as a direct regulator of transcription. Finally, S100B, a dimeric EF-hand Ca(2+)-binding protein, interacts with the tumor suppressor p53 and controls its transcriptional activity. In light of the structural studies reported to date, this review provides an overview of the structural basis of EF-hand Ca(2+)-binding proteins linked with transcriptional regulation. PMID- 12111725 TI - My favorite cell--Paramecium. AB - A Paramecium cell has a stereotypically patterned surface, with regularly arranged cilia, dense-core secretory vesicles and subplasmalemmal calcium stores. Less strikingly, there is also a patterning of molecules; for instance, some ion channels are restricted to certain regions of the cell surface. This design may explain very effective and selective responses, such as that to Ca(2+) upon stimulation. It enables the cell to respond to a Ca(2+) signal precisely secretion (exocytosis) or by changing its ciliary activity. These responses depend on the location and/or type of signal, even though these two target structures co-exist side-by-side, and normally only limited overlap occurs between the different functions. Furthermore, the patterning of exocytotic sites and the possibility of synchronous exocytosis induction in the sub-second time range have considerably facilitated analyses, and thus led to new concepts of exocytotic membrane fusion. It has been possible to dissect complicated events like overlapping Ca(2+) fluxes produced from external sources and from internal stores. Since molecular genetic approaches have become available for Paramecium, many different gene products have been identified only some of which are known from "higher" eukaryotes. Although a variety of basic cellular functions are briefly addressed to demonstrate the uniqueness of this unicellular organism, this article focuses on exocytosis regulation. PMID- 12111724 TI - The folic acid biosynthesis pathway in bacteria: evaluation of potential for antibacterial drug discovery. AB - The potential of the folic acid biosynthesis pathway as a target for the development of antibiotics has been acknowledged for many years and validated by the clinical use of several drugs. Recently, the crystal structures of all but one of the enzymes in the pathway from GTP to dihydrofolate have been determined. Given that structure-based drug design strategies are now widely employed, these recent developments have prompted a re-evaluation of the potential of each of the enzymes in the pathway as a target for development of specific inhibitors. Here, we review the current knowledge of the structure and mechanism of each enzyme in the bacterial folic acid biosynthesis pathway from GTP to dihydrofolate and draw conclusions regarding the potential of each enzyme as a target for therapeutic intervention. PMID- 12111726 TI - MEN, destruction and separation: mechanistic links between mitotic exit and cytokinesis in budding yeast. AB - Cellular events must be executed in a certain sequence during the cell division in order to maintain genome integrity and hence ensure a cell's survival. In M phase, for instance, chromosome segregation always precedes mitotic exit (characterized by mitotic kinase inactivation via cyclin destruction); this is then followed by cytokinesis. How do cells impose this strict order? Recent findings in budding yeast have suggested a mechanism whereby partitioning of chromosomes into the daughter cell is a prerequisite for the activation of mitotic exit network (MEN). So far, however, a regulatory scheme that would temporally link the initiation of cytokinesis to the execution of mitotic exit has not been determined. We propose that the requirement of MEN components for cytokinesis, their translocation to the mother-daughter neck and triggering of this translocation by inactivation of the mitotic kinase may be the three crucial elements that render initiation of cytokinesis dependent on mitotic exit. PMID- 12111727 TI - Is human aging still mysterious enough to be left only to scientists? AB - The feasibility of reversing human aging within a matter of decades has traditionally been dismissed by all professional biogerontologists, on the grounds that not only is aging still poorly understood, but also many of those aspects that we do understand are not reversible by any current or foreseeable therapeutic regimen. This broad consensus has recently been challenged by the publication, by five respected experimentalists in diverse subfields of biogerontology together with three of the present authors, of an article (Ann NY Acad Sci 959, 452-462) whose conclusion was that all the key components of mammalian aging are indeed amenable to substantial reversal (not merely retardation) in mice, with technology that has a reasonable prospect of being developed within about a decade. Translation of that panel of interventions to humans who are already alive, within a few decades thereafter, was deemed potentially feasible (though it was not claimed to be likely). If the prospect of controlling human aging within the foreseeable future cannot be categorically rejected, then it becomes a matter of personal significance to most people presently alive. Consequently, we suggest that serious public debate on this subject is now warranted, and we survey here several of the biological, social and political issues relating to it. PMID- 12111728 TI - Evo-devo comes into focus. PMID- 12111729 TI - A genetic attack on the defense complex. AB - An increasing number of "non-model" organisms are becoming accessible to genetic analysis in the field, as evolutionary biologists develop dense molecular genetic maps. Peichel et al.'s recent study(1) provides a microsatellite-based map for threespine stickleback fish (Gasterosteus aculeatus), and the first evidence for QTL affecting feeding morphology and defensive armor. This species has undergone rapid and parallel morphological and behavioral evolution, and there is now hope that some of the genes responsible for the divergence may soon be identified. PMID- 12111730 TI - A map of the common chimpanzee genome. AB - The completion of the chimpanzee genome will greatly help us determine which genetic changes are unique to humanity. Chimpanzees are our closest living relative, and a recent study has made considerable progress towards decoding the genome of our sister taxon.1 Over 75,000 common chimpanzee (Pan troglodytes) bacterial artificial chromosome end sequences were aligned and mapped to the human genome. This study shows the remarkable genetic similarity (98.77%) between humans and chimpanzees, while highlighting intriguing areas of potential difference. If we wish to understand the genetic basis of humankind, the completion of the chimpanzee genome deserves high priority. PMID- 12111731 TI - The ups and downs of daily life: profiling circadian gene expression in Drosophila. AB - Circadian rhythms are responsible for 24-hour oscillations in diverse biological processes. While the central genes governing circadian pacemaker rhythmicity have largely been identified, clock-controlled output molecules responsible for regulating rhythmic behaviors remain largely unknown. Two recent reports from McDonald and Rosbash(1) and Claridge-Chang et al.2 address this issue. By identifying a large number of genes whose mRNA levels show circadian oscillations, the reports provide important new information on the biology of circadian rhythm. In addition, the reports illustrate both the power and limitations of microarray-based methods for profiling mRNA expression on a genomic scale. PMID- 12111732 TI - Minimally disturbed, multicycle, and reproducible synchrony using a eukaryotic "baby machine". AB - A eukaryotic "baby machine" has been developed that produces synchronized cultures that display up to four synchronous cell cycles. That such cells can be produced implies that methods unable to produce successive synchronized cell cycles may not actually synchronize cells. But most important, the baby machine method now opens the way for the study of the cell cycle of minimally disturbed, artifact-free, well-synchronized, mammalian cells. PMID- 12111733 TI - Checking in on Cds1 (Chk2): A checkpoint kinase and tumor suppressor. AB - Together, DNA repair and checkpoint responses ensure the integrity of the genome. Coordination of cell cycle checkpoints and DNA repair are especially important following genotoxic radiation or chemotherapy, during which unusually high loads of DNA damage are sustained. In mammalian cells, the checkpoint kinase, Cds1 (also known as Chk2) is activated by ATM in response to DNA damage. The role of Cds1 as a checkpoint kinase depends on its ability to phosphorylate cell cycle regulators such p53, Cdc25 and Brca1. A role for Cds1 in repair is suggested by the finding that it interacts with the Holliday junction resolving activity Mus81. This review focuses on the many questions generated by recent progress in understanding the function and regulation of human Cds1. PMID- 12111734 TI - Memory in bacteria and phage. AB - Whenever the state of a biological system is not determined solely by present conditions but depends on its past history, we can say that the system has memory. Bacteria and bacteriophage use a variety of memory mechanisms, some of which seem to convey adaptive value. A genetic type of heritable memory is the programmed inversion of specific DNA sequences, which causes switching between alternative patterns of gene expression. Heritable memory can also be based on epigenetic circuits, in which a system with two possible steady states is locked in one or the other state by a positive feedback loop. Epigenetic states have been observed in a variety of cellular processes, and are maintained by diverse mechanisms. Some of these involve alternative DNA methylation patterns that are stably transmitted to daughter molecules and can affect DNA-protein interactions (e.g., gene transcription). Other mechanisms exploit autocatalytic loops whereby proteins establish the proper conditions for their continued synthesis. Template polymers other than nucleic acids (e.g., components of the cell wall) may also propagate epigenetic states. Non-heritable memory is exemplified by parasitic organisms that bear a signature of their previous host, such as host-controlled modification of phage DNA or porin hitchhiking in predatory bacteria. The heterogeneous nature of the examples known may be indicative of widespread occurrence of memory mechanisms in bacteria and phage. However, the actual extent, variety and potential selective value of prokaryotic memory devices remain open questions, still to be addressed experimentally. PMID- 12111735 TI - Embryonic origin of the eyes in teleost fish. AB - The developmental history of the vertebrate eye begins at an early embryonic stage, with the formation of the body axes and induction of neural tissue. Several recent experimental embryological and genetic studies in teleost fish have produced new insights into the morphogenetic and molecular regulation of eye formation. Molecular signaling pathways and patterned expression of transcription factors implicated in eye determination are discussed, and the importance of morphogenetic cell movements is emphasized. PMID- 12111736 TI - The corticostriatal junction: a crucial region for forebrain development and evolution. AB - Most parts of the brain are conserved across reptiles and birds (sauropsids) and mammals. Two major qualitative differences occur in the upper part, or pallium, of the telencephalon, the most rostral part of the brain. Mammals have a six layered neocortex and also exhibit a different morphological organization in the lateral half, or sector, of their pallium than do sauropsids. These differences of lateral pallial construction may derive from small but crucial differences in migration patterns of neuronal precursors generated at or above the corner of the lateral ventricle, the corticostriatal junction (CS). Sauropsids have a large structure, the dorsal ventricular ridge, that is proliferated from this region, and its anterior part (ADVR) receives ascending projections from the dorsal thalamus. Mammals have multiple structures in this same region-the lateral part of neocortex, amygdala, and claustrum-endopiriform formation. We propose here that, as the degree of development of structures that form the deeper tier of the pallium varies across the stages of embryology and across phylogeny, mutations may have occurred during evolution at the origin of mammals that had profound consequences for the fate of neural populations generated in the region of the CS and its neighboring pallial germinal zone. PMID- 12111737 TI - Muscular dystrophies, the cytoskeleton and cell adhesion. AB - Muscular dystrophies are associated with mutations in genes encoding several classes of proteins. These range from extracellular matrix and integral membrane proteins to cytoskeletal proteins, but also include a heterogeneous group of proteins including proteases, nuclear proteins, and signalling molecules. Muscular dystrophy phenotypes have also become evident in studies on various knockout mice defective in proteins not previously considered or known to be mutated in muscular dystrophies. Some unifying themes are beginning to emerge from all of these data. This review will consider recent advances in our understanding of the molecules involved and bring together data that suggest a role for the cytoskeleton and cell adhesion in muscular dystrophies. PMID- 12111738 TI - The nature of robustness in development. AB - A trait is robust to a genetic or environmental variable if its variation is weakly correlated with variation in that variable. The source of robustness lies in the fact that the developmental processes that give rise to complex traits are nonlinear. A consequence of this nonlinearity is that not all genes are equally correlated with the trait whose ontogeny they control. Here we explore how developmental mechanisms determine and alter the correlation structure between genes and the traits that they control. A formula is developed by which the correlation of a gene or environmental variable with a trait can be calculated if the mechanism that gives rise to the trait is known. The nature of robustness and the ways in which robustness can evolve are discussed in the context of the problems that arise in the analysis of inherently nonlinear systems. PMID- 12111739 TI - Search for enhancers: teleost models in comparative genomic and transgenic analysis of cis regulatory elements. AB - Homology searches between DNA sequences of evolutionary distant species (phylogenetic footprinting) offer a fast detection method for regulatory sequences. Because of the small size of their genomes, tetraodontid species such as the Japanese pufferfish and green spotted pufferfish have become attractive models for comparative genomics. A disadvantage of the tetraodontid species is, however, that they cannot be bred and manipulated routinely under laboratory conditions, so these species are less attractive for developmental and genetic analysis. In contrast, an increasing arsenal of transgene techniques with the developmental model species zebrafish and medaka are being used for functional analysis of cis regulatory sequences. The main disadvantage is the much larger genome. While comparison between many loci proved the suitability of phylogenetic footprinting using fish and mammalian sequences, fast rate of change in enhancer structure and gene duplication within teleosts may obscure detection of homologies. Here we discuss the contribution and potentials provided by different teleost models for the detection and functional analysis of conserved cis regulatory elements. PMID- 12111741 TI - Vegetative incompatibility in the ascomycete Rosellinia necatrix studied by fluorescence microscopy. AB - We describe our examination of the cytological characteristics of the vegetative incompatibility reaction in a filamentous ascomycetes, Rosellinia necatrix, by analyzing the fluorescence emitted by ethidium bromide and acridine orange stained nuclei. Hyphal anastomosis between incompatible strains, which were field and single ascospore isolates, were observed with cell death showing fused hyphae, and nuclei debris which were intensified by staining with ethidium bromide. In contrast, the nuclei in a living cell were not intensified by staining with ethidium bromide but were intensified by staining with acridine orange. A strain was found which did not form a barrier reaction, but which could be shown to undergo cell death and therefore showed a positive vegetative incompatibility reaction. We also examined the vegetative incompatibility among five single ascospore isolates and the putative parent strain from the same perithecium; all strains were incompatible. These results strongly suggest that vegetative incompatibility in R. necatrix is regulated by many loci. PMID- 12111742 TI - Molecular characterization of local Bacillus thuringiensis strains recovered from Northern Jordan. AB - A total of 80 isolates of Bacillus thuringiensis were recovered from different habitats of Northern Jordan. These isolates were grouped into three classes based on crystal morphology (spherical, bipyramidal, and both bipyramidal and cuboidal). The isolates that produced spherical crystals were the most common and the most toxic to diptera. SDS-PAGE analysis of the isolates and some reference strains with similar crystal morphology showed similar protein profiles with heterogeneous multiple protein components. The plasmid DNA content of the isolates in comparison with B. thuringiensis serovar israelensis gave a similar single intense DNA band. PMID- 12111743 TI - Alcohol production from starch by mixed cultures of Aspergillus awamori and immobilized Saccharomyces cerevisiae at different agitation speeds. AB - The influence of different agitation speeds on alcoholic fermentation by free and immobilized cells of Saccharomyces cerevisiae in a co-culture with free cells of Aspergillus awamori was investigated. Starch hydrolysis and glucose, glucoamylase and alpha-amylase accumulation in the fermentation medium was affected by agitation speed. Maximum amounts of the enzymes were obtained at 150-200 rpm after 72 h where, maximum growth and glucose accumulations were noticed at 200 300 rpm. Alcohol production was stimulated by low agitation speed (50 rpm) and decreased at higher speeds. The results also showed that the amount of produced alcohol was affected by the time of yeast inoculation. When the inoculation of yeast was carried out after the growth of fungi for 72 h, the amounts of produced alcohol increased by 84, 75, 89 and 68% at 50, 100, 150 and 200 rpm, respectively than that produced when the two organisms were inoculated together at the beginning of the fermentation process. A batch culture of the two organisms produced about 2% (v/v) alcohol from 12% (w/v) corn starch in 72 h at 50 rpm. On the other hand, the immobilized yeast and suspended A. awamori produced more alcohol reaching 3.7% (v/v) at 200 rpm under the same cultivation conditions. The fermentation process was less affected by alginate and cell concentrations of the immobilized yeast. Repeated batch fermentation by co-culture of A. awamori and immobilized yeast cells were successfully used for 12 times without a significant loss in alcohol production. PMID- 12111744 TI - A mutation in the folA promoter delays adaptation to minimal medium by Escherichia coli K-12. AB - In its natural environment, Escherichia coli is subjected to frequent changes in nutrients and physical parameters such as temperature. Adapting to new conditions requires an orderly progression of events. We show that a folA mutant that overexpressed dihydrofolate reductase because of a C to T promoter mutation, adapted very slowly when transferred from rich medium at 37 degrees C to glucose minimal medium at 42 degrees C. It adapted more rapidly when serine and threonine were added. At 37 degrees C, it was more sensitive than its parent to serine, pyruvate and valine. We propose that the mutant does not increase synthesis of serine and threonine normally upon transfer to minimal medium. This results in a limited availability of serine, threonine and isoleucine during adaptation. Phenotypically, this is manifested as a growth delay at 42 degrees C and an increased sensitivity to isoleucine restriction at 37 degrees C. PMID- 12111745 TI - Independently expressed N-terminal pro-domain of aqualysin I precursor complements the folding of its mature domain to active form in Escherichia coli. AB - Aqualysin I is a subtilisin-type serine protease secreted into the culture medium by Thermus aquaticus YT-1. It is first produced as a large precursor that consists of a signal peptide, an N-terminal pro-domain, the mature protease domain and a C-terminal pro-domain. To investigate whether the N-terminal pro domain supplied in trans as an independent peptide plays an important role in the folding and secretion of the protease, the N-terminal pro-domain in E. coli has been expressed independent of the mature domain with or without the C-terminal pro-domain using an expression system with separate promoters and signal peptides. Protease assay and SDS-PAGE clearly showed that the N-terminal pro domain plays an essential role in guiding the proper folding in trans of the enzymatically active conformation of aqualysin I. The N-terminal amino acid sequences of the purified enzymes were identical and had no signal peptides. These results indicate that independently expressed domains are secreted into the periplasmic space before the N-terminal pro-domain-assisted folding of the mature domain. PMID- 12111746 TI - Integration analysis of pSK41 in the chromosome of a methicillin-resistant Staphylococcus aureus K-1. AB - An integrate of pUB110 carrying aadD and blmS genes conferring resistance to 4' OH containing aminoglycosides and bleomycin, respectively, is found in the Staphylococcal conjugative plasmid pSK41 and in the mec chromosomal region. In this study, we found an integrated copy of pSK41 in the chromosome of an arbekacin-resistant MRSA strain, K-1, clinically isolated in Japan. This is the first report of the chromosomal integration of this large plasmid in MRSA. Analysis of the nucleotide sequence and restriction map revealed a complete homology of the blmS-containing region of the integrate to wild type pSK41, except for the deletion of aadD and a part of the repU gene. In addition, we have discovered an insertion of IS1182 within the pSK41 integrate. Pulse-field gel electrophoresis-Southern analysis showed that the integration occurred outside the mec region. We also found that deletion of aadD resulted in an aberration of blmS transcription. PMID- 12111747 TI - Influence of pH on the xylose reductase activity of Candida guilliermondii during fed-batch xylitol bioproduction. AB - The influence of pH on the xylose reductase (XR) activity was studied. The fed batch fermentation using exponential feeding rate was employed to produce xylitol. The feeding started when the xylose concentration in the fermenter reached 50 g/l and the pH was adjusted to 2.5, 4.0 or 6.0. The best results for XR activity (0.567 U/mg(protein)) and xylitol volumetric productivity (1.06 g/l.h) were achieved with pH 6.0. PMID- 12111748 TI - Candida dubliniensis, a new fungal pathogen. AB - There is a high interest in Candida species other than Candida albicans because of the rise and the epidemiological shifts in candidiasis. These emerging Candida species are favored by the increase of immunocompromised patients and the use of new medical practices, and m. Most oropharyngeal candidiasis can be foundare observed in those HIV-infected patients infected with human immunodeficiency virus (HIV). Candida dubliniensis is a recently described opportunistic pathogen that is closely related to C. albicans but differs from it with respect to epidemiology, certain virulence characteristics, and the ability to develop fluconazole resistance in vitro. C. dubliniensis has been linked to oral candidiasis in AIDS patients, although it has recently been associated to invasive disease. C. dubliniensis shares diagnostic characteristics with C. albicans, as germ tube- and chlamydospore-production, and it is generally misclassified as C. albicans by standard diagnostic procedures. Several recent studies have attempted to elucidate useful phenotypic and genotypic characteristics for separating both species. A large variety of methods have been developed with the aim of facilitating rapid and, accurate identification of this species. These have included differential chromogenic isolation platesculture media, direct immunological tests, and enhanced manual and automated biochemical and enzymatic panels. Chromogenic isolation media, as CHROMagar Candida, demonstrate better detection rates than traditional media, and allow the presumptive identification of C. dubliniensis by means of colony color (dark green colonies). API 20 C AUX system is considered a reference method, but ID 32 C strip, the VITEK Yeast Biochemical Card and the VITEK 2 ID-YST system correctly identify most C. dubliniensis isolates, being the latter the most accurate. Spectroscopic methods, such as Fourier transformed-infrared spectroscopy, offer potential advantages. However, many authors consider that standard methods for differentiation of Candida species are time-consuming, often insensitive and can fail to distinguish C. dubliniensis. To overcome these low sensitivity, poor specificity and intolerable delay,drawbacks, molecular tools have been developed to discriminate C. dubliniensis, and particularly those based on the polymerase chain reaction. But, molecular tools prove difficult and too complex for routine use in the clinical laboratory setting and new developments are necessary. Moreover, an increased resistance to antifungal drugs has been described. Although preliminary studies indicate that most strains of C. dubliniensis are susceptible to antifungal agents, fluconazole-resistant strains have been detected. Furthermore, fluconazole-resistant strains are easily derived in vitro, showing an increased expression of multidrug resistance transporters, as MDR1. PMID- 12111749 TI - Bacterial protease inhibitors. AB - Serine-, cysteine-, and metalloproteases are widely spread in many pathogenic bacteria, where they play critical functions related to colonization and evasion of host immune defenses, acquisition of nutrients for growth and proliferation, facilitation of dissemination, or tissue damage during infection. Since all the antibiotics used clinically at the moment share a common mechanism of action, acting as inhibitors of the bacterial cell wall biosynthesis or affecting protein synthesis on ribosomes, resistance to these pharmacological agents represents a serious medical problem, which might be resolved by using new generation of antibiotics, possessing a different mechanism of action. Bacterial protease inhibitors constitute an interesting such possibility, due to the fact that many specific as well as ubiquitous proteases have recently been characterized in some detail in both gram-positive as well as gram-negative pathogens. Few potent, specific inhibitors for such bacterial proteases have been reported at this moment except for some signal peptidase, clostripain, Clostridium histolyticum collagenase, botulinum neurotoxin, and tetanus neurotoxin inhibitors. No inhibitors of the critically important and ubiquitous AAA proteases, degP or sortase have been reported, although such compounds would presumably constitute a new class of highly effective antibiotics. This review presents the state of the art in the design of such enzyme inhibitors with potential therapeutic applications, as well as recent advances in the use of some of these proteases in therapy. PMID- 12111750 TI - Glycogen synthase kinase 3 (GSK-3) inhibitors as new promising drugs for diabetes, neurodegeneration, cancer, and inflammation. AB - Glycogen synthase kinase 3 (GSK-3) was initially described as a key enzyme involved in glycogen metabolism, but is now known to regulate a diverse array of cell functions. Two forms of the enzyme, GSK-3alpha and GSK-3beta, have been previously identified. Small molecules inhibitors of GSK-3 may, therefore, have several therapeutic uses, including the treatment of neurodegenerative diseases, diabetes type II, bipolar disorders, stroke, cancer, and chronic inflammatory disease. As there is lot of recent literature dealing with the involvement of GSK 3 in the molecular pathways of different diseases, this review is mainly focused on the new GSK-3 inhibitors discovered or specifically developed for this enzyme, their chemical structure, synthesis, and structure-activity relationships, with the aim to provide some clues for the future optimization of these promising drugs. PMID- 12111751 TI - Current trends in QSAR on NO donors and inhibitors of nitric oxide synthase (NOS)*. AB - This article evaluates the quantitative structure-activity relationships (QSAR) of nitric oxide (NO) radical donors and nitric oxide synthases (NOS) inhibitors, using the C-QSAR program of Biobyte. Furoxans, triazines, amidoximes, tetrazoles, imidazoles and N(omega)-2-nitroarylamino acid analogues were included in this survey. In nine out of seventeen cases, the clog P plays a significant part in the QSAR of the NO radical donors and of the NOS inhibition. Many of the compounds must be interacting with a hydrophobic space in a non-specific way. In some cases molecular refractivity CMR/MR as well as sterimol parameters (B(1) and L) are important. Electronic effects, with the exception of the Hammett's constant sigma and the Swain-Lupton parameter F, are not found to govern the biological activity. Stereochemical and electronic features are also found to be important. Indicator variables were used after the best model was found to account for the usual structural features. PMID- 12111752 TI - Diet and arterial hypertension: is the sodium ion alone important? AB - Hypertension is a widespread phenomenon whose ultimate cause is still unknown. Many factors contribute to this disease, and partially for this reason, hypertension responds to different treatments in different individuals. It is difficult to generalize about therapies for general populations. In particular, the role of electrolytes in hypertension varies widely across individuals. This review focuses its attention on sodium, potassium, calcium, and magnesium ions in order to investigate whether these electrolytes play a role in the pathogenesis of arterial hypertension and its treatment. Some individuals are especially sensitive to sodium, and changing their intake of dietary sodium may lead to variations in the levels of the other electrolytes. These changes in electrolyte levels can complicate treatments for arterial hypertension in some patients. PMID- 12111753 TI - Artificial static and geomagnetic field interrelated impact on cardiovascular regulation. AB - Spreading evidence suggests that environmental and artificial magnetic fields have a significant impact on cardiovascular system. The modulation of cardiovascular regulatory mechanisms may play a key role in observed effects. The objective was to study interrelated impacts of artificial static magnetic field (SMF) and natural geomagnetic field (GMF) on arterial baroreceptors. We studied baroreflex sensitivity (BRS) in conscious rabbits before and after 40 min of sham (n = 20) or application of Nd2-Fe14-B alloy magnets (n = 26) to the sinocarotid baroreceptor region in conjunction with GMF disturbance during the actual experiment, determined by K- and A(k)-indexes from a local geomagnetic observatory. SMF at the position of baroreceptors was 0.35 T. BRS was estimated from peak responses of mean arterial pressure (MAP) and heart rate expressed as percentages of the resting values preceding each pair of pressure (phenylephrine) and depressor drug (nitroprusside) injections. We observed a significant increase in BRS for the nitroprusside depressor test (0.78 +/- 0.1 vs. 1.15 +/- 0.14 bpm/mmHg%, initial value vs. SMF exposure, P <.0002) and a tendency for phenylephrine pressor test to increase in BRS. Prior to SMF exposure, a significant positive correlation was found between actual K index values and MAP (t = 2.33, P =.025, n = 46) and a negative correlation of the K index with BRS (t = -3.6, P =.001, n = 46). After SMF exposure we observed attenuation of the geomagnetic disturbance induced a decrease in BRS. Clinical trials should be performed to support these results, but there is a strong expectation that 0.35 T SMF local exposure to sinocarotid baroreceptors will be effective in cardiovascular conditions with arterial hypertension and decreased BRS, due to a favorable SMF effect on the arterial baroreflex. Magnets to the sinocarotid triangle along with modification of the pharmacotherapy for hypertension should be especially effective on days with intense geomagnetic disturbance, in moderating sympathetic activation and baroreceptor dysfunction. PMID- 12111754 TI - Gene expression of cytokine receptors in HL60 cells exposed to a 50 Hz magnetic field. AB - The effects of a 50 Hz extremely low frequency (ELF) sinusoidal magnetic field (MF) on the expression of genes relating to cytokine receptors were studied in HL60 cells. Transcription levels of tumor necrosis factor receptor (TNFR) p55 and p75, interleukin-6 receptor-alpha (IL-6Ralpha) and transforming growth factor beta receptor 1 (TGFbetaR1) were quantified in cells exposed to an intensity of 0.1 or 0.8 mT for periods ranging from 30 min to 72 h. Cells treated with 10 nM of phorbol 12-myristate 13-acetate (PMA) for 8 h served as a positive control. Gene expression values were assessed by the ribonuclease protection assay (RPA) and normalized to those of the noninducible gene GAPDH. The results showed that MF exposure at 0.1 and 0.8 mT for 72 h increased TNFR p75 and IL-6Ralpha mRNA expression in HL60 cells. No significant change in gene expression levels of TNFR p55 and TGFbetaR1 was observed under any of the exposure conditions. In addition, we report here for the first time that IL-6Ralpha mRNA expression can be suppressed by PMA in HL60 cells. PMID- 12111755 TI - ELF magnetic fields increase amino acid uptake into Vicia faba L. roots and alter ion movement across the plasma membrane. AB - Vicia faba seedlings, subjected to a 10 microT 50 Hz square wave magnetic field for 40 min together with a radioactive pulse, showed a marked increase in amino acid uptake into intact roots. A more modest increase was observed with a 100 microT 50 Hz square wave. An increase in media conductivity at low field intensities from 10 microT 50 Hz square wave, 100 microT 50 Hz sine wave, and 100 microT 60 Hz square wave fields, indicated an alteration in the movement of ions across the plasma membrane, most likely due to an increase in net outflow of ions from the root cells. Similarly, marked elevation in media pH, indicating increased alkalinity, was observed at 10 and 100 microT for both square and sine waves at both 50 and 60 Hz. Our data would indicate that low magnetic field intensities of 10 and 100 microT at 50 or 60 Hz can alter membrane transport processes in root tips. PMID- 12111756 TI - No effect of extremely low-frequency magnetic field observed on cell growth or initial response of cell proliferation in human cancer cell lines. AB - An effect on the tumor promotion process, as represented by accelerated cell growth, has been indicated as one example of areas that demonstrate the possibility of biological effects of extremely-low frequency magnetic fields. We, therefore, exposed the five cell lines (HL-60, K-562, MCF-7, A-375, and H4) derived from human tumors to a magnetic field for 3 days to investigate the effects on cell growth. Prior to exposure or sham exposure, the cells were precultured for 2 days in low serum conditions. The number of growing cells was counted in a blind manner. To investigate the effect on the initial response of cell proliferation, two cell lines were synchronized in G1 phase by serum starvation and then exposed to a magnetic field for 18 h (H4 cells) or 24 h (MCF 7 cells), both with and without serum stimulation. The rate of DNA synthesis, taken as a measure of the cell proliferation, was determined by following the incorporation of [(3)H]-thymidine into the DNA. Three different magnetic field polarizations at both 50 and 60 Hz were used: linearly polarized (vertical); circularly polarized; and an elliptically polarized field. Magnetic field flux densities were set at 500, 100, 20 and 2 microT (rms) for the vertical field and at 500 microT (rms) for the rotating fields. No effect of magnetic field exposure was observed on either cell growth or the initial response of cell proliferation. PMID- 12111757 TI - Studies of 50 Hz circularly polarized magnetic fields of up to 350 microT on reproduction and embryo-fetal development in rats: exposure during organogenesis or during preimplantation. AB - Groups of mated female Sprague-Dawley rats were simultaneously exposed to 0 (sham exposed), 7, 70, or 350 microT (rms) circularly polarized 50 Hz magnetic fields (MF) for 22 h/day on gestational day 8-15, the period of rat fetal organogenesis (organogenesis study) or from day 0 to day 7 of gestation, the rat preimplantation period (preimplantation study). Developmental toxicity was assessed on gestational day 20. Identical experiments were repeated to confirm reproducibility of both studies. In both studies, statistically significant differences between exposed and sham exposed animals were observed in several measured parameters; however, these differences only appeared in one, but not both replicate experiments and generally at only an isolated exposure level. Because these differences were not reproducible and did not show a dose response relationship, they were not considered related to MF exposure. In the organogenesis study, lower kidney weights of dams were seen at 70 and 350 microT in Experiment 1. Lower dam liver weights and lower mean body weights of viable female and male fetuses were seen at 70 microT in Experiment 2. Otherwise, there were no differences in these parameters or in group means for fetal loss after implantation, number of viable fetuses, fetal body weight and sex ratio, incidences of external, visceral, and skeletal abnormalities or variations, or tissue abnormalities after histopathological examination. In the preimplantation study, dam health and indices for reproduction and embryo-fetal development, including pre or postimplantation loss, number and body weight of live fetuses, and sex ratio, external, skeletal abnormalities and variations, and skeletal ossification did not differ. Dam inorganic phosphorous concentration at 350 microT was elevated in one experiment and depressed in another. In one experiment, visceral abnormalities, primarily thymic remnant in neck and accessory liver lobe, were increased in the 7 microT group. Based on these results from two studies, we conclude that circularly polarized 50 Hz MF exposure of up to 350 microT during the fetal organogenesis or during the preimplantation period does not affect reproduction and embryo-fetal development in Sprague Dawley rats. PMID- 12111758 TI - Potential motion related bias in the worn dosimeter measurements of two childhood leukemia studies. AB - Time averaged field measurements produced by a Positron dosimeter worn by study subjects was the primary method of exposure evaluation in two Canadian studies of childhood leukemia and AC magnetic field exposure. Statistically significant but mutually contradictory results obtained in the two studies, done in different locales but under similar study conditions, have not been explained. This report examines operational features of the Positron meter, including an unanticipated sensitivity to wearer motion. If the convalescent cases studied were less active than their healthy controls, as one might expect, then the meter's characteristic responses to motion, particularly as they would affect case-control distributions above and below the different referent group cutpoints used in the two studies, could help to explain both the unexpected inverse risks reported in the larger study and the unusually high risks reported in the smaller study. PMID- 12111759 TI - Effects of pulsed electromagnetic field (PEMF) stimulation on bone tissue like formation are dependent on the maturation stages of the osteoblasts. AB - The effects of pulsed electromagnetic field (PEMF, 15 Hz pulse burst, 7 mT peak) stimulation on bone tissue-like formation on osteoblasts (MC3T3-E1 cell line) in different stages of maturation were assessed to determine whether the PEMF stimulatory effect on bone tissue-like formation was associated with the increase in the number of cells and/or with the enhancement of the cellular differentiation. The cellular proliferation (DNA content), differentiation (alkaline phosphatase activity), and bone tissue-like formation (area of mineralized matrix) were determined at different time points. PEMF treatment of osteoblasts in the active proliferation stage accelerated cellular proliferation, enhanced cellular differentiation, and increased bone tissue-like formation. PEMF treatment of osteoblasts in the differentiation stage enhanced cellular differentiation and increased bone tissue-like formation. PEMF treatment of osteoblasts in the mineralization stage decreased bone tissue-like formation. In conclusion, PEMF had a stimulatory effect on the osteoblasts in the early stages of culture, which increased bone tissue-like formation. This stimulatory effect was most likely associated with enhancement of the cellular differentiation, but not with the increase in the number of cells. PMID- 12111760 TI - Comment on "Extremely low frequency magnetic fields can either increase or decrease analgesia in the land snail depending on field and light conditions" by Frank S. Prato, M. Kavaliers, and A.W. Thomas. PMID- 12111763 TI - Independent effect of vitamin B12 deficiency on hematological status in older Chinese vegetarian women. AB - We have examined the independent effect of vitamin B(12) deficiency on hematological indices in older Chinese vegetarian women using a cross-sectional study design: 119 women older than 55 years who had been vegetarian for more than 3 years were studied. Fasting blood samples were taken for complete blood count, serum iron, total serum iron binding capacity, serum iron saturation, serum vitamin B(12), serum folate, serum methylmalonic acid levels (MMA), and renal function test. Subjects with iron deficiency (iron saturation <15%) and those with serum creatinine >150 mmol/L were excluded. The prevalence of definite vitamin B(12) deficiency (vitamin B(12) level < 150 pmol/L and MMA >or= 0.4 micromol/L) was 42%. Another 32.8% had possible vitamin B(12) deficiency (either criterion). The prevalence of iron deficiency was 10%. After exclusions, 96 subjects were further analyzed. Vitamin B(12) deficiency defined by serum vitamin B(12) and MMA was associated with a decrease in hemoglobin concentrations by up to 0.9 g/dL, but it was not associated with an increase in mean corpuscular volume (MCV). Serum MMA but not vitamin B(12) levels correlated inversely with hemoglobin and platelet counts and positively with MCV, after adjustment of confounding factors. However, the percentage of subjects with anemia did not increase significantly until serum MMA became >1.0 micromol/L. In conclusion, vitamin B(12) deficiency was associated with a significant decrease in hemoglobin concentration. However, anemia associated with vitamin B(12) deficiency was seldom macrocytic. We recommend that older vegetarians should be given vitamin B(12) supplements routinely. PMID- 12111762 TI - Phase II trial of sequential therapy with fludarabine followed by cyclophosphamide, mitoxantrone, vincristine, and prednisone for low-grade follicular lymphomas. AB - Advanced follicular lymphomas, grades I and II, are indolent tumors but are not considered curable with standard therapy. Fludarabine has the highest single agent response rates in this disease. However, fludarabine-based combination chemotherapy regimens have been associated with significant myelotoxicity. Data exist suggesting that the best way to combine partially non-cross-resistant agents may be to use them sequentially. Patients with bulky stage II, stage III, or stage IV follicular lymphoma (grade I or II) were entered on this protocol. Patients were treated with 3 cycles of fludarabine followed by 6-8 cycles of cyclophosphamide, mitoxantrone, vincristine, and prednisone (CNOP). Response was assessed after the 3(rd) cycle of fludarabine and after the 4(th), 6(th), and 8(th) cycles of CNOP. Twenty-seven patients were entered on the protocol. Median follow-up was 50 months. Eighteen patients (67%) attained a complete response (CR), and eight patients (30%) attained a partial response (PR), for an overall response rate of 97%. Median relapse-free survival was 34 months, and median overall survival was not reached for the entire cohort. While all patients who achieved only PR progressed, more than half of those in CR remain free of progression at 39-84 months of follow-up. The regimen was well tolerated. The sequential combination of fludarabine and CNOP appears to be active and well tolerated in patients with grade I and II follicular lymphoma. Patients who achieve CR fare best, and many remain disease-free long term. While these results are encouraging, the addition of other active agents such as rituximab to this regimen may further enhance efficacy and is under investigation. PMID- 12111765 TI - Testing strategies for diagnosing lupus anticoagulant: decision analysis. AB - Clinicians commonly evaluate patients with thrombosis or a prolonged activated partial thromboplastin time (aPTT) for the presence of a lupus anticoagulant (LA). We evaluated strategies for detecting LA, in three clinical settings, with decision-modeling techniques. A decision tree was constructed with 12 strategies, using a combination of aPTT and dilute Russell viper venom times (dRVVT) with confirmatory tests, tissue thromboplastin time (TTI), platelet neutralization procedures, and mixing studies. Probabilities and costs of adverse events and test costs were obtained from the literature. Patient preference for each strategy was evaluated by assigning utilities to each outcome. On the basis of assay results in 90 healthy people and 77 patients, we calculated sensitivities and specificities for each strategy, with true positives defined as suggested by the International Society on Thrombosis and Haemostasis. The least costly strategy for evaluation of patients with a prolonged aPTT, or with thrombosis, is not to test and to assume that LA is absent. For patients with systemic lupus erythematosus (SLE), it is least expensive not to test, although testing with TTI alone can also be considered an efficient strategy. The strategy of highest utility to patients with SLE is testing with TTI, followed by dRVVT. On the basis of these cost and utility results, clinicians' strategies for detecting LA may need modification. These strategies would then optimally be tested in clinical trials. PMID- 12111764 TI - Acetaminophen and diphenhydramine as premedication for platelet transfusions: a prospective randomized double-blind placebo-controlled trial. AB - Non-hemolytic transfusion reactions (NHTR) occur in up to 30% of patients receiving platelet transfusions. Premedication with acetaminophen and diphenhydramine is a common strategy to prevent NHTR, but its efficacy has not been studied. In this prospective trial, transfusions in patients receiving pre storage leukocyte-reduced single-donor apheresis platelets (SDP) were randomized to premedication with either acetaminophen 650 mg PO and diphenhydramine 25 mg IV, or placebo. Fifty-one patients received 98 transfusions. Thirteen patients had 15 NHTR: 15.4% (8/52) in the treatment arm and 15.2% (7/46) in the placebo arm. Premedication prior to transfusion of pre-storage leukocyte reduced SDP does not significantly lower the incidence of NHTR as compared to placebo. PMID- 12111766 TI - Outcome in hemoglobin SC disease: a four-decade observational study of clinical, hematologic, and genetic factors. AB - Over the past 40 years, we observed 284 subjects with hemoglobin SC disease (Hb SC) for 2,837 person-years. We examined the association of the course of clinical events with hematologic and genetic factors. The mean entry age was 21 years, although 15% entered before one year of age. The mean Hb concentration was 11.3 g/dL, the mean fetal hemoglobin was 2.5%, and the mean MCV was 84.4 fL. Twenty five subjects died at a median age of 37 years. Chronic organ-specific complications occurred in 112 subjects (39.4%), with advanced retinopathy in 65 subjects (22.9%) and osteonecrosis (avascular necrosis) in 42 subjects (14.8%). We identified the beta-globin haplotypes in 82 subjects and the alpha-gene status in 79. Twenty-nine percent had alpha-thalassemia-2. The beta(CI) haplotype was present in 85.4%. We found a decreased incidence of retinopathy in the beta(CI) subjects compared to the non-beta(CI) subjects (33% vs. 67%; P = 0.049) with a later mean onset age (29 years vs. 21 years; log-rank test, P= 0.026). We also found a consistent pattern of decreased morbidity in subjects who had alpha thalassemia-2 in comparison to those who did not. We found a reduced risk of chronic organ-specific complications (log-rank test, P= 0.003), lower incidence of sickle crisis (48% vs. 80%, P= 0.001), later onset of gallbladder disease (age of onset: 55 years vs. 34 years; P= 0.055), and lower risk of osteonecrosis (log rank test, P= 0.024). Our findings suggest that Hb SC subjects who have not inherited alpha-thalassemia-2 might benefit from erythrocyte rehydration therapy. PMID- 12111767 TI - Sickle erythrocyte adherence to endothelium at low shear: role of shear stress in propagation of vaso-occlusion. AB - Under venular flow conditions, sickle cell adherence to endothelium is mediated by cell adhesion molecules and adhesive proteins associated with inflammation, coagulation, and endothelial perturbation. Periodic and reduced blood flow are observed in sickle microcirculation during hematologic steady state, suggesting that blood flow is compromised in sickle microcirculation. We tested the hypothesis that low blood flow enhances adherence by quantifying sickle cell adhesion to endothelium under venular flow (1.0 dyne/cm(2) shear stress) and low flow (0.1 dyne/cm(2) shear stress), with and without addition of adhesion promoting agonists. Under low flow, sickle cell adherence to endothelium increases with contact time in the absence of endothelial activation or adhesive protein addition. In contrast, at venular shear stress, sickle cell adherence only occurs following endothelial activation with TNF-alpha or addition of thrombospondin. Analysis of these data with a mathematical model reveals that at low flow adherence is "transport-controlled," meaning that contact time between sickle cells and endothelium is a more important determinant of adherence than high-affinity receptor-ligand interactions. Low-affinity interactions are sufficient for adhesion at low flow. In contrast, at venular flow (1 dyne/cm(2) shear stress) adherence is "affinity-controlled," meaning that adherence requires induction of specific high-affinity receptor-ligand interactions. These findings demonstrate that in addition to activating factors and adherence proteins, microvascular shear stress is an important determinant of sickle cell adhesion to endothelium. This suggests that in vivo, erythrostasis is an important determinant of adhesion that can act either independently or concurrently with ongoing acute events to induce adhesive interactions and vaso-occlusion. PMID- 12111768 TI - Silent brain infarcts are rare in Kuwaiti children with sickle cell disease and high Hb F. AB - Overt stroke is rare among sickle cell disease (SCD) patients in Kuwait. However, there are no previous studies of silent cerebral infarcts, which have been described in up to 20% of American children with Hb SS. We have carried out a prospective brain MRI study among otherwise normal SCD patients, who were consecutive patients seen in a 1-year period to document the prevalence of silent cerebral infarcts in children with sickle cell disease in Kuwait. Any patient with a previous seizure or other neurological abnormality was excluded. MRI was done with a 1.5 Tesla unit with super-conducting magnet. T1- and T2-weighted sagittal and axial sections and proton density axial images were obtained in 5-mm thick sections. The study group consisted of 30 (23 SS and 7 Sbeta(0)Thal) patients-19 males and 11 females-whose ages ranged from 6 to 17 (mean of 9.8 +/- 3.5) years. Hb F ranged from 11% to 35% with a mean of 22.8% +/- 5.7%. Only one patient, a 10-and-a-half-year-old boy with Hb SS, showed hyperintense signals in the parietal white matter, consistent with small infarcts, thus giving a prevalence of 3.3%. Silent brain infarcts are uncommon in our patients, and the protective factors remain to be fully elucidated. PMID- 12111769 TI - Asymptomatic homozygous deletional beta(0)-thalassemia in an African individual. AB - Homozygosity or compound heterozygosity for beta(0)-thalassemia mutations most commonly results in a transfusion-dependent thalassemia major phenotype. In this report, we describe a 55-year-old male, from Guinea-Bissau, that had been asymptomatic and never transfused until being admitted to hospital with anemia, fever, splenomegaly, and asthenia. Following hospital admission, HIV-2 and Mycobacterium tuberculosis infections were diagnosed, and biochemical and molecular studies revealed homozygosity for beta(0)-thalassemia. At the molecular level, this is the first description of homozygosity for the beta(0)-Black 1,393 bp deletion. In this case, the complete absence of beta-globin gene expression seems to be compensated by an unusually high fetal globin gene expression (Hb F 96%). Beta-globin haplotyping results were compatible with the propositus being homozygous for the Black 2 haplotype and for the absence of the XmnI polymorphism at -158 of (G)gamma-globin gene (-/-). Co-inheritance of genetic factors usually associated with high Hb F levels was not detected. Otherwise, the propositus is a heterozygote for the alpha-globin gene 3.7-kb deletion that is a beneficial modulating factor but not sufficient to explain this extremely mild phenotype. This unusual genotype/phenotype association is discussed in terms of the mechanisms underlying hemoglobin switching during development. PMID- 12111770 TI - Chylothorax in chronic lymphocytic leukemia patient. AB - Chronic lymphocytic leukemia (CLL) is rarely complicated by chylothorax: we present a 93-year-old woman with CLL who developed recurrent pleural effusions that were ultimately found to be chylous in nature. Despite eight repeated thoracenteses, she continued to experience re-accumulation of fluid, and therefore, video-assisted thoracotomy with mass ligation of the thoracic duct region plus pleurodesis was performed to resolve the chylothorax. Despite her age and underlying disease, she did well during follow-up. The etiology and management of chylothorax are also reviewed. PMID- 12111771 TI - Multiple pulmonary nodules: an unusual presentation of fludarabine pulmonary toxicity: case report and review of literature. AB - Fludarabine monophosphate, a purine analogue is used in the treatment of lymphoid malignancies. A 73-year-old woman presented with fever and cough 2 weeks after completing the third cycle of fludarabine for chronic lymphocytic leukemia (CLL). Chest roentgenogram showed multiple pulmonary nodules. Pulmonary histopathology demonstrated a mononuclear interstitial infiltrate without evidence of malignant, infectious, granulomatous, or vascular causes. Her symptoms and pulmonary nodules resolved following treatment with corticosteroids. To our knowledge, four cases of interstitial pneumonitis associated with fludarabine have been reported in medical literature. Fludarabine induced lung toxicity must be considered in all patients who develop unexplained lung disease while receiving fludarabine. It is reversible with discontinuation of drug and administration of corticosteroids. This case extends the spectrum of fludarabine pulmonary toxicity to include pulmonary nodules. PMID- 12111772 TI - Platelet satellitism, spurious neutropenia, and cutaneous vasculitis: casual or causal association? AB - About 90 cases of platelet satellitism (PS) have been published, most of them involving neutrophils, a few comprising monocytes, and one case involving basophils. The case of a 30-year-old female patient with cutaneous vasculitis who developed asymptomatic severe neutropenia is reported. All blood smears (BS) prepared from peripheral blood samples collected with either ethylenediaminetetraacetic acid, trisodium citrate, and heparin showed PS involving neutrophils, eosinophils, and monocytes. Immunohistochemistry analysis of her skin biopsy and BS, employing peroxidase stain for the detection of antibodies directed against IgG, IgA, IgM, and C3, revealed an intense positive reaction only for IgG in the endothelium and leukocyte clumps within the microvasculature, as well as in peripheral blood neutrophils displaying the PS phenomenon. Transfer of the PS phenomenon was demonstrated by incubating the patient's plasma with leukocytes from an ABO-compatible healthy donor. In our patient, PS did not disappear after incubation at 37 degrees C, suggesting that this might be a different or "atypical" PS phenomenon probably mediated by an autoimmune process involving an IgG-class antibody responsible for both conditions, PS, and cutaneous vasculitis. PMID- 12111774 TI - Cholecystitis as the presenting manifestation of acute myeloid leukemia: report of a case. AB - We report a case of acute leukemia, which presented as cholecystitis in a previously healthy middle-aged adult. Leukemic involvement of the gastrointestinal tract is a well-known clinicopathological entity. However, leukemic infiltration of the gall bladder wall is a rare occurrence. It has only been previously described in the setting of relapsed disease and, to our knowledge, has never been reported as the primary manifestation of de novo acute leukemia. The patient in question was treated with standard induction and consolidation chemotherapy and remains in complete remission 19 months after his presentation with symptoms consistent with cholecystitis. PMID- 12111773 TI - Bone marrow necrosis with dyspnea in a patient with malignant lymphoma and plasma levels of thrombomodulin, tumor necrosis factor-alpha, and D-dimer. AB - Non-Hodgkin's lymphoma (peripheral T cell, unspecified, clinical stage IIIEA) was diagnosed in a 54-year-old male. He was treated weekly with chemotherapy consisting of pirarubicin hydrochloride, cyclophosphamide, methotrexate, vincristine sulfate, etoposide, and prednisolone. After 6 weeks of treatment in a state of partial remission, he exhibited sudden dyspnea, chest pain, fever, and drowsiness. The patient had received 100 microg/day of granulocyte colony stimulating factor (G-CSF) for 6 days before the onset. Laboratory data showed an elevated lactate dehydrogenase (LDH) level, leukoerythroblastosis in the peripheral blood, and no decrease in the serum haptoglobin level. There were no findings of acute myocardial infarction or pulmonary thromboembolism. Bone marrow specimen showed the characteristic features of necrosis without any signs of the involvement of lymphoma cells. No indications of infections were found. This patient was diagnosed as having bone marrow necrosis (BMN) during the recovery phase of bone marrow with G-CSF treatment after chemotherapy for malignant lymphoma. After conservative therapy, inhalation of oxygen and stopping the administration of G-CSF, all clinical symptoms subsided except that the elevation of LDH continued for 1 month. The plasma level of tumor necrosis factor-alpha (TNF-alpha) was high just after the onset of BMN. The thrombomodulin (TM) level was high just before the onset of BMN and continued to be high for 2 weeks. Cross linked fibrin degradation products (D-dimer) were also high just after the onset of BMN and continued to be high for 3 weeks after the onset. Although dyspnea is a rare symptom of BMN, it is important to rule out in BMN during the recovery phase of bone marrow with G-CSF treatment after chemotherapy for malignant lymphoma. Activated neutrophils in the small vessels of the lung by G-CSF and microthrombi, suggested by the elevation of D-dimer, may participate in the onset of dyspnea, which is a rare symptom of the onset of BMN. PMID- 12111775 TI - Herbs for HIV infection. PMID- 12111776 TI - Successful treatment of non-Hodgkin's lymphoma complicated with autoimmune neutropenia. PMID- 12111777 TI - Thalidomide-associated gynecomasty in a patient with multiple myeloma. PMID- 12111779 TI - Autosomal non-dominant hereditary spherocytosis: does it occur in India? PMID- 12111778 TI - First case of hemoglobin San Diego in Italy. PMID- 12111780 TI - Variation of PDGF, TGFbeta, and bFGF levels in essential thrombocythemia patients treated with anagrelide. AB - We studied 15 patients with essential thrombocythemia (ET) before treatment and after normalization of platelet count by anagrelide. Significantly increased plasma levels of PDGF, TGFbeta, and bFGF were found. Patients with mild reticulin fibrosis in bone marrow had higher PDGF levels. During treatment, plasma TGFbeta and bFGF levels remained elevated in most patients (P < 0.0001 and P < 0.01, respectively). Intraplatelet PDGF levels were low before treatment (P < 0.006) and normal on hematological remission, without relation with the presence or absence of reticulin fibrosis in bone marrow. Intraplatelet TGFbeta levels were normal regardless of the platelet count. Intraplatelet bFGF levels were raised before (P < 0.001) and during treatment (P < 0.01). By immunostaining, TGFbeta and bFGF were seen in megakaryocytes and lymphocytes with a similar pattern of intensity in patients and controls, suggesting that other cells might also contribute to the raised plasma values. We believe that the plasma increment of these cytokines suggests that they play a role in the pathogenesis of ET. The normal PDGF plasma level found during treatment may be in relation with the platelet count. However, the persistent increase of TGF-beta in plasma and bFGF both in plasma and platelets may indicate dysregulation of cytokine synthesis in TE. PMID- 12111781 TI - Successful treatment of anemia in idiopathic myelofibrosis with recombinant human erythropoietin. AB - Thirteen patients with idiopathic myelofibrosis (5 osteomyelosclerosis) were treated with recombinant human erythropoietin (rHuEpo) for transfusion-dependent anemia. All but 7 patients were concomitantly treated with alpha interferon, and 5 patients also received a interferon before the start of erythropoietin (EPO) treatment. All but two patients became transfusion independent. The highly positive results of the present study of transfusion-dependent patients with idiopathic myelofibrosis calls for further studies to delineate more precisely in larger series those patients who are likely to respond to rHuEpo. PMID- 12111782 TI - Increased circulating activated endothelial cells, vascular endothelial growth factor, and tumor necrosis factor in thalassemia. AB - An increased number of circulating endothelial cells (CECs) was demonstrated in alpha- and beta-thalassemic patients, beta-thalassemia/hemoglobin E (BE), both splenectomized (BE[S]) and non-splenectomized (BE[NS]), had higher numbers of CECs than alpha-thalassemia, both HbH (alpha-thal l/alpha-thal 2; H) and HbH with hemoglobin Constant Spring (alpha-thal 1/CS; H/CS). CECs were also increased in heterozygous HbE (EA) and homozygous HbE (EE). The highest level of tumor necrosis factor-alpha (TNF-alpha) was found in HbH/CS patients, whereas the highest levels of vascular endothelial growth factor (VEGF) was observed in BE[S] patients. Significant decreases, in protein C and protein S levels were found in both alpha- and beta-thalassemia compared with normal. Good correlations between the numbers of CEC and TNF-alpha, VEGF, protein C, and protein S levels were demonstrated in this study. In addition, markers for endothelial cell activation and injury (intercellular adhesion molecule-1, ICAM-1/CD54; vascular cell adhesion molecule-1, VCAM-1/CD106; and E-selectin, ELAM-1/CD62E) were detected on the surface of isolated CECs using immunofluorescence technique. Appearance of CECs with markers for endothelial cell activation, together with increased levels of TNF-alpha and VEGF and decreased levels of protein C and protein S in the circulation, may account for the propensity of vascular perturbation in thalassemic subjects. PMID- 12111784 TI - Expression and prognostic significance of Bcl-2 family proteins in myelodysplastic syndromes. AB - Excessive apoptosis is implicated in the pathogenesis of myelodysplastic syndromes (MDS). We assessed by flow cytometry the expression of several members of the Bcl-2 family in bone marrow mononuclear cells (BMMNC) of 168 MDS samples at diagnosis. The proteins studied were Bcl-2, Bcl-xL (anti-apoptotic), Bax, Bad, Bak, and Bcl-xS (pro-apoptotic). The percentage of BMMNC expressing Bcl-2 and Bcl xL was higher in refractory anemia with excess of blasts (RAEB), RAEB in transformation (RAEB-T), and chronic myelomonocytic leukemia (CMML) than in refractory anemia (RA) and RA with ringed sideroblasts (RAS). Conversely pro apoptotic proteins Bad, Bak, and Bcl-xS were detected in a higher percentage of cells in RA and RAS. RA and RAS were associated with an increased Bcl-xS/Bcl-xL ratio. The expression of anti-apoptotic proteins was also correlated with that of CD34 and P170 and with the percentage of blast cells. Two-color analyses demonstrated that CD34 and Bcl-2 were usually expressed in the same cells. No significant correlation was found with cytogenetic abnormalities. Higher expression of pro-apoptotic Bcl-2-family proteins (Bak, Bad, Bcl-xS) and higher Bcl-xS/Bcl-xL ratio were associated with longer survival and decreased risk of leukemic transformation in univariate analysis, whereas expression of anti apoptotic proteins was associated with decreased survival. Consequently Bcl-2 proteins expression was well correlated with the International Prognostic Scoring System (IPSS). Our data confirm that the control of apoptosis is deregulated in MDS cells. Moreover, the study of markers such as CD34 (or Bcl-2), Bcl-xL, and Bcl-xS provides additional prognostic information. PMID- 12111783 TI - Hyperhomocysteinemia and cobalamin deficiency in young Asian Indians in the United States. AB - Hyperhomocysteinemia, a risk factor for vascular disease, may be a particular problem in Asian Indians, but information is limited, especially in the U.S., despite its growing Asian population. Moreover, suggestions have been made that folate deficiency is responsible for the hyperhomocysteinemia in Indians. Therefore, we studied homocysteine status in healthy Asian Indians in the U.S. prospectively, determined the frequency of cobalamin and folate deficiency as contributors to it, and examined whether food-cobalamin absorption contributed to cobalamin deficiency. Homocysteine levels were higher in Asian Indian men than in 4 other ethnic groups (P < 0.0001); 10/39 Indian men (25.6%) were hyperhomocysteinemic. Cobalamin levels were lower in Indian men (P = 0.000005) and women (P = 0.03) than in non-Indians; low levels were found more frequently in both Indian men (23/39; 59.0%) and women (5/21; 23.8%) than in others. Measuring methylmalonic acid in 10 selected subjects showed that the low cobalamin levels reflected cobalamin deficiency, and high methylmalonic acid levels were found in some subjects without hyperhomocysteinemia. Evidence of folate deficiency was not found in any subjects. Food-cobalamin absorption was normal in all 13 Indian subjects tested, including those with Helicobacter pylori infection. The results show that hyperhomocysteinemia is strikingly common in apparently healthy, young Asian Indian men. The cause appears to be cobalamin deficiency, which affected more than half of the Indian men, may be largely subclinical, is underestimated by homocysteine levels alone which were not always abnormal, and is probably largely dietary in origin. Folate deficiency is rare. This public health problem is amenable to prevention and treatment in this growing segment of the U.S. population. It was, parenthetically, noteworthy that many of the affected subjects were young physician trainees. PMID- 12111785 TI - Prevalence of hereditary thrombophilia in patients with thrombosis in different venous systems. AB - The prevalence of hereditary thrombophilia is well known in patients with lower extremity thrombosis but only poorly studied in patients with thrombosis at unusual sites. Consequently, it is still unclear whether such patients should generally be screened for hereditary thrombophilia. We retrospectively analyzed 260 patients with thrombosis at unusual sites including thrombosis in portal, cerebral, retinal, and upper-extremity veins with respect to the prevalence of FV Leiden, prothrombin G20210A, protein C, protein S, and antithrombin deficiency. In addition, all thrombotic episodes were analyzed for circumstantial risk factors. Used as controls, healthy volunteers (120) and patients with lower extremity thrombosis (292) showed overall prevalence of hereditary thrombophilia of 9.1% and 39.0%, respectively. The corresponding numbers were 33.3%, 34.3%, and 39.0% in patients with portal vein, upper-extremity, and lower-extremity thrombosis, respectively. In patients with cerebral vein thrombosis, however, the prevalence was significantly lower (23.5%). Patients with retinal vein occlusion did not show an increased frequency of thrombophilia at all (5.9%). In all five groups FV Leiden was by far the most frequent defect (4.4-27.1%), while prothrombin G20210A occurred rarer (2.5-7.6%). Protein C, protein S, and antithrombin deficiency were much less prevalent (0-3.1%) except for patients with portal vein thrombosis (4.8-7.1%). Compared to healthy individuals, the relative risk of thrombosis was 4.3 (2.2-8.1), 3.8 (1.8-7.7), 2.5 (1.0-6.1), 3.7 (1.5-8.6), and 0.6 (0.2-2.1) for patients with lower-extremity, upper-extremity, cerebral vein, portal vein, and retinal vein thrombosis, respectively. Circumstantial risk factors were more frequent in patients without than with hereditary thrombophilia and were found most often in patients with upper extremity thrombosis. In each group the most frequent circumstantial risk factor was different. However, oral contraceptives and cancer were found in all five groups. In conclusion, independent upon the presence of circumstantial risk factors, screening for hereditary thrombophilia is warranted in all patients with thrombosis at unusual sites except in those with retinal vein occlusion. PMID- 12111786 TI - Cytogenetic findings in reactive lymphoid hyperplasia: significance of non-clonal t(3;14) and t(3;22). AB - Despite several reports of the molecular detection of recurrent lymphoma translocations in reactive lymph nodes (LN), cytogenetic analysis is seldom performed on such cases. We report the clinical and cytogenetic analysis results on 30 reactive LN. Of these, 17 cases yielded either no growth (n = 9) or normal metaphases only (n = 8), and seven of the 17 patients subsequently developed lymphoma. Lymphoma developed in all 10 patients with a clonal karyotype (median of 2.6 months). Three patients (1 HIV-positive) had non-clonal t(3;14) or t(3;22). Their lymphadenopathy resolved spontaneously, and none progressed to lymphoma at 4-6 years of follow-up. Molecular methods detected a small B-cell clone in one case and an oligoclonal B-cell population in the other. Cytogenetic analysis may be useful for interpreting cases of lymphoid hyperplasia. A clonal abnormality is highly predictive of concurrent and/or subsequent lymphoma. A lymphoma-specific but non-clonal abnormality does not necessarily herald the development of subsequent lymphoma. PMID- 12111787 TI - A reliable plasma marker of platelet activation: does it exist? AB - Despite the clear importance of the platelet in various conditions and diseases, there are few opportunities to assess the physiological and pathological functions of this cell. Those without access to a flow cytometer or platelet aggregometer rely on secreted or release products of the platelet. Principle among these molecules are alpha granule components beta thromboglobulin and platelet factor four, and membrane constituents such as P selectin, gpV and glycocalicin. However, notwithstanding the ease of measurement of these markers (i.e., by ELISA) each one has its own particular disadvantage, mostly of methodology and specificity. Nevertheless, if our goal is to improve our ability to recognize and treat subjects with thrombotic disorders, then additional studies on these molecules may prove to be a sound investment. PMID- 12111788 TI - Spontaneous gas gangrene in malignant lymphoma: an underreported complication? AB - We report a case of spontaneous gas gangrene (SGG), the most rapidly progressive form of clostridial infection, in a patient with non-Hodgkin's lymphoma (NHL). We review the literature and examine the association between these two entities. A 43-year-old man with NHL developed fatal C. perfringens-associated SGG and massive hemolysis during induction chemotherapy. Although patients with NHL usually have several risk factors of SGG, such as bowel involvement or neutropenia, only two cases have been described previously in detail. Common features of all reports are a delayed diagnosis and a fatal outcome. Awareness of this condition should result in prompt antibiotic therapy at the onset of typical presenting symptoms in any lymphoma patient, especially if risk factors are present. PMID- 12111789 TI - Multiple sclerosis associated with interferon-alpha therapy for chronic myelogenous leukemia. AB - We report a chronic myelogenous leukemia (CML) patient in chronic phase (CP) who developed multiple sclerosis (MS) in association with interferon-alpha (IFN alpha) administration. In our patient, recombinant IFN-alpha2b therapy induced hematologically complete and cytogenetically major partial response for CML first, and sequential central nervous system dysfunction evolved, which subsided shortly after the cessation of its administration. Restarting IFN-alpha therapy by changing to a natural type of IFN-alpha resulted in rapid exacerbation of MS. The patient's neurological symptoms progressed gradually, but partial hematologic response persisted without any IFN-alpha derivatives or anti-cancer agents until a matched unrelated donor transplant procedure was performed. Myeloablative therapy led to lasting stable state of MS and finally to complete cytogenetic remission of CML. This patient's presenting clinical course and laboratory data suggest that both exertion of anti-leukemic activity and autoimmune process of MS might be mediated by mutual mechanisms, such as enhancement of specific cellular immunity induced by IFN-alpha. PMID- 12111790 TI - Acquired pulmonary alveolar proteinosis after umbilical cord blood transplantation for acute myeloid leukemia. AB - Pulmonary alveolar proteinosis (PAP) is a heterogeneous disease that occasionally develops with hematological malignancy. However, PAP in association with hematopoietic stem cell transplantation is quite rare. Here we present the first report of a patient who developed PAP after cord blood transplantation (CBT). A 45-year-old female with AML underwent unrelated CBT. On day +2 after CBT she developed congestive heart failure with diffuse alveolar infiltrates in the bilateral lungs. Despite treatment, the alveolar infiltrates further increased with progression of multiple organ failure (MOF). She died from MOF before hematopoietic recovery on day +27. Post-mortem study revealed that massive amorphous materials positive for periodic acid-Schiff stain filled in the pulmonary alveoli. These findings led to a diagnosis of PAP. The bone marrow was hypocellular without the leukemic cells. The impaired immunity during the period of leukopenia as well as the impaired clearance of surfactant proteins might contribute to the development of PAP. PMID- 12111791 TI - Platelet spherocytosis: a new bleeding disorder. AB - The present study evaluated a child with thrombocytopenia and a rare congenital bleeding disorder associated with platelet spherocytosis. Differential interference phase contrast microscopy (DIC) revealed that his platelets were spherical in form. Examination of thin sections in the electron microscope showed that his platelets were nearly devoid of microtubules (MT) and microtubule coils (MTC) that support the discoid shape in 100% of normal platelets. Immunofluorescence with a monoclonal antibody to tubulin, the precursor protein of MT, revealed bright rings in normal cells and diffuse fluorescence of patient platelets. The brightness of the fluorescence emitted by patient platelets was comparable to the diffuse fluorescence of normal platelets after chilling to dissolve intact MT, suggesting normal and patient cells contained comparable amounts of tubulin. Exposure of patient platelets to Taxol, an agent that stabilizes and induces MT, caused MT formation in 82% of patient platelets and MTC development in 11%, resulting in their conversion to discs. Glycoproteins GPIIb/IIIa and GPIb were present on patient platelets, and the cells contained normal numbers of dense bodies, ruling out storage pool disease. The patient's platelets adhered to and spread normally on glass but failed to undergo rapid, irreversible aggregation when stirred with agents that produced a complete response in normal discoid platelets, even when the patient's platelets were concentrated. The poor response of spherical platelets was associated with failure to become irregular and extend long filopodia. Thus, the spherical shape of patient platelets may contribute to the thrombocytopenia and to the clinical bleeding symptoms. PMID- 12111792 TI - Monoclonal gammopathy after low-grade MALT lymphoma: evidence for a second neoplasm. AB - We report the case of a patient with lymphoma of the salivary gland, at first diagnosed as lymphoma of mucosa-associated lymphoid tissue (MALT lymphoma) but later found to infiltrate the bone marrow. At diagnosis, the patient had a polyclonal increase of gamma-globulins. Five years after initial diagnosis, the patient presented with monoclonal gammopathy and infiltration of the bone marrow with neoplastic cells. Initially, the patient had received chemotherapy with different protocols (including etoposide, cyclophosphamide, fludarabin, methotrexate, and vincristine), none of which induced a lasting response. Therapy with rituximab (chimeric anti-CD20 monoclonal antibody) finally led to partial remission. Eighteen months after rituximab, progressive lymphoma in the abdomen and a monoclonal gammopathy developed. The bone marrow showed infiltration by lymphoplasmacytoid cells (monoclonal expression of the light-chain type lambda, positive for CD20, heterogeneous expression of CD45). The patient achieved another short clinical response with 4 cycles of the CHOP-protocol, but soon the lymphoma progressed again. Five years and 8 months after the initial diagnosis, the patient died from septicemia and progressive lymphoma. By polymerase chain reaction (PCR) for the IgH gene it was shown that lymphoma cells were initially oligoclonal in the salivary gland and, later, biclonal in the bone marrow. Sequencing of two bands of apparently same length showed that these manifestations of lymphoma were not identical. Taken together, our data show that the initial low-grade oligoclonal MALT lymphoma was no longer present and a more aggressive biclonal lymphoma with plasmacytoid differentiation had developed. The new lymphoma was clonally distinct and produced high amounts of monoclonal IgG lambda by immunoelectrophoresis. The relationship of the second lymphoma to the initial MALT lymphoma is discussed. PMID- 12111793 TI - Amyloidoma of lymph node. PMID- 12111794 TI - Lichenoid eruption to STI 571. PMID- 12111795 TI - Caspases on the brain. AB - The basic mechanisms that underlie neurodegenerative diseases are unknown. Loss of function of specific regions of the brain is due to incapacitation of cells that constitute those regions. Cells can simply stop functioning normally (neurons may cease to transmit signals), or they may die. There is now evidence that the pathology of several neurodegenerative diseases is due to inappropriate apoptosis. This being the case, an understanding of the mediators of apoptosis, their identities, and their role in orchestrating death would be a vital step toward remedying the diseases. The central components of apoptotic pathways, proteases of the caspase family, are present in latent forms in all nucleated cells. Their activity is balanced by specific activation and inactivation events, and the molecular and biochemical controls have been well established in vitro and in model transformed cell lines. In this Mini-Review, we consider the current status of the basic control mechanisms and how these may be subverted during neurodegeneration. PMID- 12111796 TI - BDNF activated TrkB/IRR receptor chimera promotes survival of sympathetic neurons through Ras and PI-3 kinase signaling. AB - Insulin receptor-related receptor (IRR) expression is tightly coupled to the nerve growth factor (NGF) receptor, TrkA, throughout development. Expression of both receptors is primarily localized to neural crest derived sensory and sympathetic neurons. In contrast to TrkA, however, the physiological ligand for IRR is unknown. To analyze the intracellular signaling and potential function of the orphan IRR in neurons, an adenovirus expressing a TrkB/IRR chimeric receptor was used to infect cultured mouse superior cervical ganglion neurons that normally require NGF for survival. Brain derived neurotrophic factor (BDNF) activated TrkB/IRR induced neuronal survival. We utilized numerous receptor mutants in order to identify the intracellular domains of IRR necessary for signaling and neuron survival. Finally, we employed adenovirus encoding dominant negative forms of the extracellular signal-regulated kinase (ERK) signaling cascade to demonstrate that IRR, like TrkA, requires ras activation to promote neuron survival. Therefore, by use of the chimeric TrkB/IRR receptor, we have demonstrated the ability of IRR to elicit activation of signaling cascades resulting in a biological response in superior cervical ganglion (SCG) neurons. PMID- 12111797 TI - Brain-derived neurotrophic factor applied to the motor cortex promotes sprouting of corticospinal fibers but not regeneration into a peripheral nerve transplant. AB - Previous experiments from our laboratory have shown that application of brain derived neurotrophic factor (BDNF) to the red nucleus or the motor cortex stimulates an increase in the expression of regeneration-associated genes in rubrospinal and corticospinal neurons. Furthermore, we have previously shown that BDNF application stimulates regeneration of rubrospinal axons into a peripheral graft after a thoracic injury. The current study investigates whether application of BDNF to the motor cortex will facilitate regeneration of corticospinal neurons into a peripheral nerve graft placed into the thoracic spinal cord. In adult Sprague Dawley rats, the dorsal columns and the corticospinal tract between T9 and T10 were ablated by suction, and a 5-mm-long segment of predegenerated tibial nerve was autograft implanted into the lesion. With an osmotic pump, BDNF was infused directly into the parenchyma of the motor cortex for 14 days. Growth of the corticospinal tract into the nerve graft was then evaluated by transport of an anterograde tracer. Anterogradely labeled corticospinal fibers were not observed in the peripheral nerve graft in animals treated with saline or BDNF. Serotinergic and noradrenergic fibers, as well as peripheral sensory afferents, were observed to penetrate the graft, indicating the viability of the peripheral nerve graft as a permissive growth substrate for these specific fiber types. Although treatment of the corticospinal fibers with BDNF failed to produce regeneration into the graft, there was a distinct increase in the number of axonal sprouts rostral to the injury site. This indicates that treatment of corticospinal neurons with neurotrophins, e.g., BDNF, can be used to enhance sprouting of corticospinal axons within the spinal cord. Whether such sprouting leads to functional recovery after spinal cord injury is currently under investigation. PMID- 12111798 TI - Secretory phospholipase A(2) induces delayed neuronal COX-2 expression compared with glutamate. AB - Agonists of the binding site for secretory phospholipase A(2) (sPLA(2)) potentiate glutamate-induced neuronal cell death in primary cell cultures and in vivo (Kolko et al. [1996] J. Biol. Chem. 271:32722; Kolko et al. [1999] Neurosci. Lett. 274:167]. Here, we tested the hypothesis that COX-2 expression participates in the brain response to sPLA(2). sPLA(2)-OS(2), a selective ligand of a neuronal sPLA(2)-binding site, was injected into the rat striatum, and early-response gene expression was monitored by in situ hybridization using (35)S-radiolabeled oligonucleotide probes and immunohistochemistry. An up-regulation of COX-2, c fos, and c-jun, but not COX-1, was observed around the lesion as well as in the neocortex 4 hr after the injection. Hippocampal up-regulation of COX-2 was seen in dentate gyrus 8 hr after injection. When glutamate was injected, up-regulation of the early-response genes peaked after 2 hr. Our studies showed 1) that sPLA(2) selectively induced neuronal COX-2; 2) that this induction was delayed (4 hr after injection of sPLA(2)) compared with that elicited by glutamate (2 hr after injection), suggesting different signaling; and 3) that c-fos and c-jun were induced around the infarct area as soon as 2 hr after injection, but in other aspects followed a time course similar to that of COX-2. We conclude that sPLA(2) may modulate neuronal COX-2 expression through mechanisms that differ from those of glutamate-induced COX-2 expression. PMID- 12111799 TI - H(2)O(2) induces upregulation of Fas and Fas ligand expression in NGF differentiated PC12 cells: modulation by cAMP. AB - Fas, (APO-1/CD95), a transmembrane glycoprotein belonging to the tumor necrosis (TNF) receptor superfamily, transduces apoptotic death upon crosslinking by its cognate ligand (FasL). As upregulation of Fas/FasL expression occurs in neuropathological conditions (e.g., stroke, central nervous system [CNS] trauma and seizures) associated with oxidative damage, we questioned whether reactive oxygen species (ROS) can directly affect Fas and FasL expression in neuronal cells. Utilizing rat PC12 cells neuronally differentiated with nerve growth factor (NGF), we observed that concentrations of H(2)O(2) inducing apoptotic cell death rapidly trigger the expression of Fas mRNA and protein as well as FasL mRNA. Although NGF-addition to naive PC12 downregulated constitutive Fas and FasL transcription, the H(2)O(2)-induced Fas and FasL mRNA upregulation invariably occurred either in the presence or in the absence of NGF. Similarly, phorbol 1,2 myristate 1, 3-acetate (PMA), a potent protein kinase C (PKC) activator, did not modify Fas and FasL mRNA upregulation subsequent to H(2)O(2) exposure. On the contrary, forskolin and dibutyryl cAMP, which elevate intracellular cAMP by independent mechanisms, both counteracted H(2)O(2)-induced Fas, but not FasL, mRNA upregulation and increased constitutive expression of FasL mRNA. Altogether, our data show that oxidative stress is a major stimulus in eliciting Fas and FasL expression in NGF-differentiated PC12 cells. Moreover, we describe here for the first time the existence of cAMP-dependent mechanism(s) modulating Fas and FasL expression. PMID- 12111800 TI - Apoptotic death following Fas activation in human oligodendrocyte hybrid cultures. AB - The purpose of this study was to determine how oligodendrocytes die following Fas receptor activation. An immortalized human oligodendrocyte hybrid line (MO3.13) was challenged with Fas ligand (FasL), and cell death was assessed by flow cytometry and DNA gel electrophoresis. Caspase activation was determined by either Western immunoblotting on cell extracts or by whole-cell flow cytometry. FasL challenge clearly induced substantial apoptotic cell death in the oligodendrocyte hybrid cell line, as judged by flow cytometry and by the presence of prominent low molecular weight DNA banding patterns after gel electrophoresis. Western immunoblots showed marked increases in cleaved caspase-1, 8, and 3, indicating that the extrinsic Fas death receptor-induced pathway was activated. The intrinsic mitochondrial pathway was also activated, but only at a minimal level. These findings demonstrate that there are several independent molecular sites within the extrinsic caspase cascade in oligodendrocytes where inhibitory compounds may be capable of blocking cell death in vivo. PMID- 12111802 TI - Insulin-like growth factor binding proteins-1 and -2 differentially inhibit rat oligodendrocyte precursor cell survival and differentiation in vitro. AB - Insulin-like growth factor-1 (IGF-1) is a growth and survival factor for oligodendrocyte lineage cells and induces myelination. Its actions are modulated by IGF binding proteins (IGFBPs) that are present in the extracellular fluids or on the cell surface. Additionally, IGFBPs are also known to exert actions that are independent of IGF-1. We studied whether IGF-binding proteins (IGFBPs)-1 and 2 modulate rat oligodendrocyte precursor (O2A) cell survival and differentiation in vitro both in the absence and presence of exogenously added IGF-1. The data reveal that IGFBP-1 and -2 reduced O2A cell survival in the absence and presence of exogenously added IGF-1. The effects of IGFBP-1 on cell survival in the presence of exogenously added IGF-1 were IGF-1-dependent, whereas IGFBP-2 displayed both IGF-1-dependent and IGF-1-independent effects. Furthermore, IGFBP 1 and -2 inhibited O2A cell differentiation in the presence of IGF-1 as reflected by decreased expression levels of two myelin proteins, CNPase (2',3'-cyclic nucleotide 3'-phosphohydrolase) and MAG (myelin associated glycoprotein). Analysis of medium samples revealed that O2A cells do not secrete proteases that degrade these IGFBPs. Taken together the data show that IGFBP-1 and -2 are negative effectors of oligodendrocyte survival and differentiation. Accordingly, the role of IGFBPs should be explicitly taken into account when investigating IGF 1 effects on oligodendrocytes, especially in the context of therapeutic purposes. PMID- 12111801 TI - Molecular evidence of apoptotic death in malignant brain tumors including glioblastoma multiforme: upregulation of calpain and caspase-3. AB - Cell death in the core of human brain tumors is triggered by hypoxia and lack of nutrients, but the mode of cell death whether necrosis or apoptosis is not clearly defined. To identify the role of apoptosis in brain tumor cell death, we investigated macromolecular (RNA and protein) synthesis and activity in the central to peripheral region of benign [desmoplastic infantile ganglioglioma (DIG) and transitional meningioma (TMG)] and malignant [ependymoma (END), anaplastic astrocytoma (APA), and glioblastoma multiforme (GBM)] brain tumors derived from five patients who had not received previously radiotherapy or chemotherapy. Normal brain tissue (NBT) served as control. RT-PCR analysis of tumor tissues covering central to peripheral regions detected mRNA overexpression of pro-apoptotic gene bax in malignant tumors, indicating a commitment to apoptosis. The mRNA expression of calpain (a Ca(2+)-dependent cysteine protease) and calpastatin (endogenous calpain inhibitor) was altered resulting in an elevated calpain/calpastatin ratio. Calpain content and activity were increased, suggesting a role for calpain in cell death. In the mitochondria-dependent death pathway, caspase-9 and caspase-3 were also overexpressed in tumors. The increased caspase-3 activity cleaved poly(ADP-ribose) polymerase (PARP). Agarose gel electrophoresis detected a mixture of random and internucleosomal DNA fragmentation in malignant brain tumors. Overexpression of pro-apoptotic bax, upregulation of calpain and caspase-3, and occurrence of internucleosomal DNA fragmentation are now presented indicating that one mechanism of cell death in malignant brain tumors is apoptosis, and that enhancement of this process therapeutically may promote decreased tumor growth. PMID- 12111803 TI - Subcellular localization and detergent solubility of MVP17/rMAL, a lipid raft associated protein in oligodendrocytes and myelin. AB - Detergent-insoluble, glycosphingolipid-cholesterol-enriched microdomains (lipid rafts) have been implicated in both protein trafficking and signal transduction. Previously we identified in oligodendrocytes and myelin the lipid raft associated, integral membrane protein myelin vesicular protein of 17 kDa (MVP17)/rMAL. Here we have examined the subcellular localization and/or detergent insolubility of native and recombinant MVP17/rMAL in transfected oligodendrocytes and COS-7 cells and purified myelin. Consistent with our previous report regarding the insolubility of MVP17/rMAL in the zwitterionic detergent 3-[(3 chloramidopropyl)-dimethylammonio]-1-propane-sulfonate (CHAPS), MVP17/rMAL from purified myelin and oligodendrocytes in culture was mostly insoluble upon extraction at 4 degrees C with the non-ionic detergent Triton X-100 and floated to a low density in sucrose gradient ultracentrifugation, but became detergent soluble at 37 degrees C. Data obtained by immunofluorescence microscopy of the expression of epitope-tagged MVP17/rMAL transfected into oligodendrocytes and COS 7 cells were consistent with a model in which both the N- and C-termini of this protein face the cytoplasm. Mutational analysis identified domains of MVP17/rMAL important for its subcellular localization and for its detergent solubility profile. In particular, insertional mutagenesis of loop II prevented the insertion of the mutant protein into the plasma membrane of COS-7 cells and rendered it insoluble in TX-100. Expression of full-length constructs of MVP17/rMAL in COS-7 cells resulted in an enlargement of transfected COS-7 cells, consistent with a proposed role of rMAL in vesicular trafficking. PMID- 12111804 TI - Proteins of peripheral myelin are associated with glycosphingolipid/cholesterol enriched membranes. AB - A characteristic feature of the vertebrate nervous system is the ensheathment of axons by myelin, a multilamellar membrane specialization produced by polarized glial cells. Although the main protein and lipid components of the myelin sheath are well characterized, relatively little is known about the mechanisms of their intracellular distribution to the respective sites of assembly within the myelin sheath. To analyze whether peripheral myelin protein trafficking is mediated by glycosphingolipid/cholesterol-enriched membranes (GEMs), we studied the association of established myelin proteins, peripheral myelin protein 22 (PMP22), protein zero (P0), plasmolipin, and myelin basic protein (MBP), with these membrane microdomains. To examine the association of the selected peripheral myelin proteins with detergent-insoluble GEMs, purified myelin from sciatic nerve of adult rat was extracted with Triton X-100 at 4 degrees C and 37 degrees C and, in additional experiments, was pretreated with the cholesterol chelator methyl beta-cyclodextrin. The material was then centrifuged to equilibrium in sucrose gradients, and fractions were analyzed by Western blotting. Here we demonstrate for the first time that PMP22, P0, and plasmolipin prepared from purified peripheral myelin are associated with GEMs. To characterize whether the association of these proteins is a specialized feature of myelinating Schwann cells, we studied the distribution of PMP22, P0, and plasmolipin in transiently transfected HeLa cells. These experiments confirm the specific association of these proteins with GEMs in both neural and nonneural cell types. PMID- 12111805 TI - Alterations in myelination in the central nervous system of dystonia musculorum mice. AB - Dystonia musculorum (dt) is an autosomal recessive sensory neuropathy in mice resulting from a mutation in the gene encoding the cytoskeletal linker protein Bpag1. In addition to neurodegeneration, dt mice display myelination abnormalities in the peripheral nervous system. In this report we investigated whether myelination abnormalities are also present in the central nervous system of dt(Tg4) mice. Transcripts for both neural isoforms of Bpag1 (a1 and a2) were detected in optic nerves and spinal cords of wild-type mice. Light microscopy of resin-embedded thin sections revealed a reduction in myelinated axons in both optic nerves and spinal cords in dt(Tg4) mice. As well, hypermyelinated axons were detected in these tissues. Ultrastructural analysis of optic nerves and spinal cords from dt(Tg4) mice revealed an increase in the number of amyelinated axons, the presence of hypo- and hypermyelinated axons, and redundant myelin that course away from axons. Changes in the level of myelin proteins accompanied the morphological alterations. Myelin-associated glycoprotein levels were reduced in optic nerves of dt(Tg4) mice, and myelin basic protein levels were altered in optic nerves, sciatic nerves, and spinal cords of affected mice. Short-term cultures of oligodendrocytes derived from dt(Tg4) mice did not show morphological alterations. PMID- 12111806 TI - Differential distribution of laminins in Alzheimer disease and normal human brain tissue. AB - Immunocytochemistry, Western blotting, and RT-PCR were used to identify the isoforms of laminin expressed in the Alzheimer disease, but not in normal human brain tissue. We found that alpha 1 laminin was heavily over-expressed in Alzheimer disease frontal cortex, and localized in reactive astrocytes of the grey and white matter, and as punctate deposits in the senile placques of the Alzheimer brain tissue. Antibodies against the C-terminal neurite outgrowth domain of the gamma 1 laminin demonstrated expression of the gamma 1 laminin in GFAP-immunoreactive reactive astrocytes of the Alzheimer disease frontal cortex. The gamma 1 laminin was also heavily over-expressed in reactive astrocytes of both grey and white matter. Although antibodies against the C-terminal neurite outgrowth domain failed to localize gamma 1 laminin in senile plaques, antibodies against the N-terminal domains of the gamma 1 laminin demonstrated gamma 1 laminin as punctate deposits in the senile plaques. The present results indicate that enhanced and specialized expression patterns of alpha 1 and gamma 1 laminins distinctly associate these two laminins with the Alzheimer disease. The fact that domain specific antibodies localize both alpha1 and gamma 1 laminins in the senile plaques as punctate deposits and in astrocytes of both the gray and white matter indicate that these laminins and their specific domains may have distinct functions in the pathophysiology of the Alzheimer disease. PMID- 12111807 TI - Effect of amyloid beta-peptide on permeability transition pore: a comparative study. AB - A potentially central factor in neurodegeneration is the permeability transition pore (PTP). Because of the tissue-specific differences in pore properties, we directly compared isolated brain and liver mitochondria responses to the neurotoxic A beta peptides. For this purpose, the following parameters were examined: mitochondrial membrane potential (Delta Psi m), respiration, swelling, ultrastructural morphology, and content of cytochrome c. Both peptides, A beta(25 35) (50 microM) and A beta(1-40) (2 microM), had a similar toxicity, exacerbating the effects of Ca(2+), although, per se, they did not induce (PTP). In liver mitochondria, A beta led to a drop in Delta Psi m and potentiated matrix swelling and disruption induced by Ca(2+). In contrast, brain mitochondria, exposed to the same conditions, demonstrated a higher capacity to accumulate Ca(2+) before the Delta Psi m drop and a slight increase of mitochondrial swelling compared with liver mitochondria. Furthermore, mitochondrial respiratory state 3 was depressed in the presence of A beta, whereas state 4 was unaltered, resulting in an uncoupling of respiration. In both types of mitochondria, A beta did not affect the content of cytochrome c. The Delta Psi m drop was reversed when Ca(2+) was removed by EGTA or when ADP plus oligomycin was present. Pretreatment with cyclosporin A or ADP plus oligomycin prevented the deleterious effects promoted by A beta and/or Ca(2+). It can be concluded that brain and liver mitochondria show a different susceptibility to the deleterious effect of A beta peptide, brain mitochondria being more resistant to the potentiation by A beta of Ca(2+) induced PTP. PMID- 12111809 TI - Protective effects of atypical antipsychotic drugs on PC12 cells after serum withdrawal. AB - Atypical antipsychotic drugs are widely used in the treatment of schizophrenia, and clinical evidence has shown that early and prolonged intervention with these drugs will improve the long-term outcome. It is still unclear, however, whether the atypical antipsychotic drugs are also neuroprotective. To clarify this matter, we used PC12 cell cultures and the MTT assay for cell viability to determine whether various concentrations of the atypical antipsychotics clozapine, quetiapine, and risperidone are neuroprotective after serum withdrawal. In addition, to explore the drugs' actions, Northern blot was used to examine the gene expression of SOD1 (Cu/Zn superoxide dismutase) and p75NTR (p75 neurotrophin receptor). The results demonstrated that 1) the antipsychotic drugs can protect PC12 cells from death after serum withdrawal; cell viability in these drug treatment groups is significantly different from that in the groups without serum in the medium (P < 0.01); and 2) these drugs up-regulated the SOD1 gene expression to more than 120% (P < 0.05) and also down-regulated p75NTR mRNA levels to less than 65% of their respective control values (P < 0.05). These findings suggest that the atypical antipsychotics clozapine, quetiapine, and risperidone may exert a neuroprotective function through the modulation of SOD1 and p75NTR expression. PMID- 12111810 TI - Extracellular proteolysis in brain injury and inflammation: role for plasminogen activators and matrix metalloproteinases. AB - The role of intracellular proteases (e.g., calpains and caspases) in the pathophysiology of neuronal cell death has been extensively investigated. More recently, accumulating data have suggested that extracellular proteolysis also plays a critical role. The two major systems that modify the extracellular matrix in brain are the plasminogen activator (PA) and matrix metalloproteinase (MMP) axes. This Mini-Review delineates major pathways of PA and MMP action after stroke, brain trauma, and chronic inflammation. Deleterious effects include the disruption of blood-brain barrier integrity, amplification of inflammatory infiltrates, demyelination, and possibly interruption of cell-cell and cell matrix interactions that may trigger cell death. In contrast, PA-MMP actions may contribute to extracellular proteolysis that mediates parenchymal and angiogenic recovery after brain injury. As the mechanisms of deleterious vs. potentially beneficial PA and MMP actions become better defined, it is hoped that new therapeutic targets will emerge for ameliorating the sequelae of brain injury and inflammation. PMID- 12111808 TI - Exposure to lead elevates induction of zif268 and Arc mRNA in rats after electroconvulsive shock: the involvement of protein kinase C. AB - Exposure to lead is well known to impair cognitive function in young children. Because of the importance of gene regulation for neurodevelopment, we examined the effect of lead on the induction of the mRNA of the immediate early genes zif268 and Arc. The time course for the induction of zif268 mRNA and Arc mRNA by electroconvulsant shock (ECS) was altered in the area of the dentate gyrus of the hippocampus in rats exposed to lead from postnatal days (PND) 1 to 28. Other areas of the hippocampus were not affected by lead. The effects on the induction of zif268 mRNA were observed at blood lead levels as low as 12 microg/dl. No change in the induction of zif268 mRNA was observed in the hippocampus of rats exposed to lead from PND 28 to PND 56. Because of the possible involvement of protein kinase C (PKC) in the effect of lead, activation of different isoforms of PKC was investigated. An increase in the amount of PKC epsilon and PKC gamma was observed at 60 min after ECS in the membrane fraction from hippocampus, indicating activation of these isoforms. The amount of PKC epsilon in membranes was higher in rats exposed to lead than in rats not exposed to lead after ECS. Taken together, the data suggest that lead may disturb regulation of specific immediate early genes by activating PKC epsilon. PMID- 12111811 TI - Reversal of amyloid beta toxicity in Alzheimer's disease model Tg2576 by intraventricular antiamyloid beta antibody. AB - There are considerable data on synaptic dysfunction in Alzheimer's disease (AD). However, the precise molecular basis for synaptotoxicity in AD is not known. We tested the hypothesis that amyloid beta (Abeta), as produced in Tg2576 mice overexpressing a mutant form of amyloid precursor protein, leads to changes in SNAP-25, a molecule required for Ca-sensitive neurotransmitter vesicle exocytosis. Anti-Abeta antibody was injected into the third ventricle (icv) of 10 month-old Tg2576 mice, preceding formation of plaques. Immunodensity of glial fibrillary acidic protein (GFAP) and SNAP-25 were quantitated in the hippocampus 1 month later. SNAP-25 was reduced by 96% in the inner molecular layer (SMi) of dentate gyrus, by 95% in the hilum, and by 75-76% in stratum lucidum (SL), stratum oriens (SO), and stratum radiatum (SR) of CA1-CA3 of the Tg2576 mice. GFAP was increased by more than 50-fold, specifically within the neuropil of CA1 CA3, and by twofold in portions of fimbria. One injection of 10 microg of anti Abeta antibody into the third ventricle at 10 months completely prevented or restored changes in GFAP at 11 months of age. The restoration of SNAP-25 by anti Abeta antibody compared with wild type was 69% in CA1-SO, 93% in CA1-SR, 85% in CA3-SL, 77% in SMi, and 60-73% in hilum. In addition, whereas control injections of saline or IgG produced greatly increased GFAP diffusely in the hippocampus of Tg2576 animals, there was no increase in GFAP after anti-Abeta injection, suggesting a synergistic interaction of nonspecific trauma with Abeta in the transgenic mice. This is the first report of depleted SNAP-25 immunoreactivity in Tg models and the first report of icv injection of anti-Abeta antibody in this model of AD. The largest reductions of the SNAP-25 are in hilum and SMi, so either reduction in the septal-hilum-SMi path is primary or reduction in this path begins at an earlier age than in CA3-CA1 fields. A single icv injection of anti-Abeta antibody is potent in reversing Abeta effects and, therefore, represents a suitable model for investigating early Abeta toxicity. In addition, intrathecal or icv antibody may be an efficient means of treating or preventing toxicity in AD, particularly under conditions of immune hyporesponsivity. PMID- 12111812 TI - Molecular mechanisms of enhanced susceptibility to apoptosis in differentiating oligodendrocytes. AB - Several studies have shown that the progression of oligodendrocyte progenitors along the lineage correlates with increased susceptibility to death stimuli. The molecular basis of this phenomenon remains unclear. This study demonstrates that the protein levels of several proapoptotic molecules, including Bax, Bad (nonphosphorylated form), and certain caspase proforms, increase during oligodendrocyte development. In contrast, the steady-state levels of antiapoptotic molecules, such as Bcl2 and Bcl(XL), remain constant. This altered equilibrium between proapoptotic and antiapoptotic molecules correlates with increased cytochrome C in the cytosol. We conclude that, as oligodendrocytes mature, their susceptibility to apoptosis increases because of a change in the balance between protective mechanisms and proapoptotic pathways. This suggests the possible existence of a death susceptibility program, which is intrinsic to differentiating oligodendrocyte progenitors. PMID- 12111814 TI - Platelet-activating factor induces permeability transition and cytochrome c release in isolated brain mitochondria. AB - Platelet-activating factor (PAF), a potent bioactive phospholipid implicated in neuronal excitotoxic death, was assessed as a mediator of brain mitochondrial dysfunction. Carbamyl PAF, a non-hydrolyzable PAF analog, added to neurons in culture resulted in decreased mitochondrial membrane potential (DeltaPsi(M)) as measured by the DeltaPsi(M)-sensitive fluorophore 5,5', 6,6'-tetrachloro-1, 1', 3,3'-tetraethylethylbenzimidazolo-carbocyanide iodide (JC-1). To investigate whether PAF has a direct effect on the mitochondria, the mediator was added to rat brain mitochondria preparations and an increase in the permeability of the mitochondrial membrane, termed permeability transition (PT), and cytochrome c release were measured. We report that PAF causes both dose-dependent PT and cytochrome c release from isolated mitochondria. Furthermore, the selective PAF antagonist tetrahydro-4,7,8,10 methyl-1 (chloro-2 phenyl)-6 (methoxy-4 phenyl carbamoyl)-9 pyrido [4',3'-4,5] thieno [3,2-f] triazolo-1,2,4 [4,3-a] diazepine 1,4 (BN50730), which has affinity for intracellular binding sites, and the peripheral benzodiazepine receptor ligands 7-chloro-5- [4'-chlorophenyl]-1,3 dihydro-1-methyl-2H-1,4-benzodiazepin-2-one (Ro5-4864) and 1-(-2-chlorophenyl)-N methyl-N-(1-methylpropyl)-3-isoquinolinecarboxamide (PK11195), inhibit PAF induction of PT and cytochrome c release. These results suggest that PAF excitotoxicity involves, at least in part, alterations of the mitochondrial membrane. PMID- 12111815 TI - Estrogen treatment suppresses forebrain p75 neurotrophin receptor expression in aged, noncycling female rats. AB - There is increasing evidence that estrogen has beneficial effects on cognition, both in humans and in rodents, and may delay Alzheimer's disease onset in postmenopausal women. Several rodent studies have utilised the ovariectomy model to show estrogen regulation of the p75 neurotrophin receptor, TrkA, and markers of acetylcholine synthesis in the cholinergic basal forebrain. We studied estrogenic effects in aged (16-17-month-old), noncycling rats. Estrogen treatment for 10 days drastically reduced p75(NTR) immunoreactivity in the rostral parts of the basal forebrain. The number of p75(NTR)-immunoreactive neurons was decreased, and those neurons remaining positive for p75(NTR) showed reduced p75(NTR) staining intensity. In vehicle-treated rats, almost all choline acetyltransferase immunoreactive neurons were p75(NTR) positive (and vice versa), but, in estrogen treated rats, large numbers of choline acetyltransferase-immunoreactive cells were negative for p75(NTR). Similar levels of p75(NTR) down-regulation in the rostral basal forebrain were found when estrogen treatment was extended to 6 weeks. There was no reduction in the number of p75(NTR)-immunoreactive neurons in the caudal basal forebrain after 10 days of treatment. After 6 weeks of treatment, however, there was evidence of p75(NTR) down-regulation in the caudal basal forebrain. There was no evidence of hypertrophy or atrophy of cholinergic neurons even after 6 weeks of estrogen treatment. Considering the evidence for the role of p75(NTR) in regulating survival, growth and nerve growth factor responsiveness of cholinergic basal forebrain neurons, the results indicate an important aspect of estrogen's effects on the nervous system. PMID- 12111813 TI - Attenuated plasticity in neurons and astrocytes in the mouse model of Sanfilippo syndrome type B. AB - Sanfilippo syndrome type B (MPS III B) is a neurodegenerative disorder characterized by profound mental retardation and early death. It is caused by deficiency of a lysosomal enzyme involved in heparan sulfate (HS) degradation. Because HS accumulation can be a major feature of this disease, we have examined crucial molecular systems associated with HS function. Using a knockout mouse with disruption of the gene responsible for HS degradation, we evaluated the effects of possible HS accumulation on neuroplasticity that are within the spectrum of action of fibroblast growth factors (FGFs) and their receptor (FGFR). We found that levels of mRNA for the FGFR-1 were attenuated in the mutant mice by the age of 6 months, whereas the mRNAs for FGF-1 and FGF-2 were reduced or unchanged in the brain regions tested. Neurogenesis, in which FGF-2 is involved, was inhibited in the MPS III B mouse brain at both young and adult ages. We also examined the expression of the glial fibrillary acidic protein (GFAP) gene and GFAP-positive cell density in both normal and injured conditions to study the functional response of astrocytes to insult. We found that, although the mutation alone caused drastic induction of reactive astrocytes, acute injury to the mutant brains failed to induce additional reactive astrocytes. Our results showed important alterations in the expression of several genes involved in the maintenance of neuroplasticity in the MPS III B. This in turn may result in reduction of neuronal health and brain function. PMID- 12111817 TI - Atypical neuroleptics stimulate neurogenesis in adult rat brain. AB - Schizophrenia has been treated effectively with atypical neuroleptics without serious side effects. We have shown previously that long-term treatment with atypical neuroleptics is correlated with an improvement of cognition in adult rats. We report here that atypical neuroleptics stimulate a 2- to 3-fold increase in newly divided cells in the subventricular zone in the rat and that some of these new cells in the subventricular zone and hippocampus also express a neuronal marker. We used bromodeoxyuridine (BrdU) to identify newly divided cells and confirmed the observation with antibody to a cell-cycle-specific, endogenous proliferating cell nuclear antigen (PCNA). Identification of BrdU-positive cells in the anterior subventricular zone (SVZa) particularly in rats treated with the atypical neuroleptics but not in those in the haloperidol-treated and control rats, suggests increased rostral migratory stream (RMS) cell traffic to replenish neurons in the olfactory bulb. Expression of a neuronal marker, NeuN, in BrdU positive cells in rats treated with atypical neuroleptics, also suggests that these compounds may modulate in vivo differentiation of neuronal progenitor cells even within a day of BrdU injection. Our results indicate that atypical neuroleptics have a mechanism of action other than the previously proposed mechanisms, which might explain their role in improved cognition in animal and in schizophrenic patients. If substantiated by future studies, our findings may lead to an expanded use of atypical neuroleptics in other neurodegenerative diseases to stimulate neuronal replacement and repair. PMID- 12111816 TI - Influence of cell density and thyroid hormone on glial cell development in primary cultures of embryonic rat cerebral hemisphere. AB - The influence of cell density and thyroid hormone (TH) on the development of astrocytes and oligodendrocytes was investigated in primary cultures prepared from rat cerebral hemisphere on embryonic day (E)18. At the beginning of the culture, most of the cells were microtubule-associated protein 2 (MAP2)-positive neurons, whereas O1-positive oligodendrocytes and glial fibrillary acidic protein (GFAP)-positive astrocytes were rarely observed. After the cells were maintained in serum-free defined medium, astrocytes developed at high cell density but rarely at a low one. When leukemia inhibitory factor (LIF) was supplemented in low-density cultures, the levels of GFAP expression markedly increased to almost the same extent as in high-density culture without TH. This suggests that, in low density cultures, astrocyte progenitors could not differentiate because of insufficient astrocyte-inducing factors. Interestingly, the addition of TH increased GFAP expression levels only at high density. The number of oligodendrocytes increased with TH addition at both cell densities, although the effects were more remarkable at high density. These results suggest that cell density and TH are pivotal factors in the development of both astrocytes and oligodendrocytes. It is also suggested that the effects of TH on glial cell development could be accelerated via cell-cell communications. PMID- 12111818 TI - Ethanol-induced increase of sphingosine recycling for ganglioside biosynthesis: a study performed on cerebellar granule cells in culture. AB - The effect of ethanol on ganglioside metabolism was assessed in cultured rat cerebellar granule cells. Cells were incubated in the presence of tritiated serine or galactose, and the synthesis of radioactive gangliosides was followed. The rate of de novo biosynthesis of gangliosides labeled in the oligosaccharide moiety (deriving from tritiated galactose) was not affected by the presence of ethanol. On the contrary, the biosynthesis of gangliosides labeled in the ceramide long chain base moiety (deriving from tritiated serine), dramatically decreased in the presence of alcohol. These results suggest that the gap between the extent of the biosynthesis of lipid and polar portions observed in the presence of ethanol, is filled by an increased recycling of sphingosine produced from ganglioside degradation. This hypothesis was confirmed by pulse-chase experiments with GM1 ganglioside, tritiated in the sphingosine moiety, and following radiolabeled gangliosides deriving from its metabolic processing. In fact, the radioactivity carried by gangliosides whose labeling could derive exclusively (GD1b + GT1b) or partially (GD1a) from the recycling of catabolic radiolabeled sphingosine, dramatically increased in ethanol-treated cells during the chase period. Taken together, these results suggest that ethanol increases ceramide sphingosine recycling for ganglioside biosynthesis. PMID- 12111819 TI - Expression of the myelin and oligodendrocyte progenitor marker sulfatide in neurons and astrocytes of adult rat brain. AB - Sulfatide is a myelin component of the central (CNS) and peripheral nervous system (PNS) and is used extensively to identify oligodendrocyte progenitor cells. We have explored sulfatide expression in CNS gray matter (cerebellum, cerebral cortex, and hippocampus) and the PNS in adult rats using an anti sulfatide antibody (Sulph I) and confocal microscopy. Biochemical analyses revealed two Sulph I antigens, sulfatide and seminolipid; sulfatide was present at about five times higher concentration, and the affinity of Sulph I for sulfatide was 2.5 times higher than that for seminolipid. Thus sulfatide was considered the dominant antigen. We found Sulph I immunostaining, in addition to that in myelinated areas in subpopulations of astrocytes and neurons. Astrocyte Sulph I staining was localized to the cell bodies and in some cases also to the processes. In the cerebellum, some Sulph I-positive astrocytes corresponded to Golgi epithelial cell bodies. We also found Sulph I staining in neuronal cell bodies, which in some neurons was clearly localized to the cytoplasm and in others to the nuclear membrane. Sulph I immunostaining in the PNS was located in the myelin sheath and paranodal end segments. These results demonstrate the expression of sulfatide in cell types other than oligodendrocytes and Schwann cells, showing that sulfatide is not a selective marker for adult oligodendrocyte progenitor cells. Moreover, these findings show that sulfatide is localized also to intracellular compartments and indicate that other roles of sulfatide in astrocytes and neurons, compared to myelin, might be considered. PMID- 12111820 TI - Cytokines, chemokines, and cytokine receptors in human microglia. AB - Enriched populations of human microglial cells were isolated from mixed cell cultures prepared from embryonic human telencephalon tissues. Human microglial cells exhibited cell type-specific antigens for macrophage-microglia lineage cells including CD11b (Mac-1), CD68, B7-2 (CD86), HLA-ABC, HLA-DR and ricinus communis aggulutinin lectin-1 (RCA-1), and actively phagocytosed latex beads. Gene expression and protein production of cytokines, chemokines and cytokine/chemokine receptors were investigated in the purified populations of human microglia. Normal unstimulated human microglia expressed constitutively mRNA transcripts for interleukin- 1beta (IL-1beta) -6, -8, -10, -12, -15, tumor necrosis factor-alpha (TNF-alpha), macrophage inflammatory protein-1alpha (MIP 1alpha), MIP-1beta, and monocyte chemoattractant protein-1 (MCP-1), while treatment with lipopolysaccharide (LPS) or amyloid beta peptides (Abeta) led to increased expression of mRNA levels of IL-8, IL-10, IL-12, TNF-alpha, MIP-1alpha, MIP-1beta, and MCP-1. Human microglia, in addition, expressed mRNA transcripts for IL-1RI, IL-1RII, IL-5R, IL-6R, IL-8R, IL-9R, IL-10R, IL-12R, IL-13R, and IL 15R. Enzyme-linked immunosorbent assays (ELISA) showed increased protein levels in culture media of IL-1beta, IL-8, TNF-alpha, and MIP-1alpha in human microglia following treatment with LPS or Abeta. Increased TNF-alpha release from human microglia following LPS treatment was completely inhibited with IL-10 pretreatment, but not with IL-6, IL-9, IL-12, IL-13, or transforming growth factor-beta (TGF-beta). Present results should help in understanding the basic microglial biology, but also the pathophysiology of activated microglia in neurological diseases such as Alzheimer disease, Parkinson disease, Huntington disease, amyotrophic lateral sclerosis, stroke, and neurotrauma. PMID- 12111821 TI - Variant PC12 cell line that spontaneously differentiates and extends neuritic processes. AB - The rat pheochromocytoma PC12 cells differentiate into neuronal-like cells in response to treatment with neurotrophins. The cells have been extensively used for investigating neuronal differentiation and axonal growth. Here we report the isolation of a variant PC12 cell line, named PC12-N1, which spontaneously differentiates and extends neuritic processes. The PC12-N1 cells expressed many neuronal specific proteins, including the synaptosomal associated protein of 25 kDa (SNAP-25), synaptotagmin, and synaptobrevin (also known as VAMP). The cells also expressed neurofilament protein of 68 kDa, a marker for differentiated neurons. In addition to the spontaneous neurite outgrowth, the PC12-N1 cells showed a marked increase in neurite outgrowth upon treatment with nerve growth factor (NGF), basic fibroblast growth factor (bFGF), and cyclic AMP (cAMP). The activation of mitogen-activated protein (MAP) kinases was examined by immunoblot analysis using phospho-specific antibodies. No overactivation was observed with ERK1/2 or p38. However, the c-Jun N-terminal kinase JNK/SAPK was activated approximately 10-fold over the parental PC12 cells. These results suggest that activation of JNK/SAPK may be involved in the spontaneous neurite extension in the PC12-N1 cells. Moreover, the PC12-N1 cells may be used as a model for investigating molecular signaling mechanisms underlying neuronal differentiation and axonal outgrowth. PMID- 12111823 TI - Regulation of the ferritin H subunit by vitamin B12 (cobalamin) in rat spinal cord. AB - Cobalamin-deficient (Cbl-D) central neuropathy is a pure myelinolytic disease, in which gliosis is also observed. Iron is abundant in the mammalian central nervous system, where it is required for various essential functions including myelinogenesis. It is predominantly located in the white matter and oligodendrocytes, which also actively synthesize the major iron proteins (e.g., ferritin, transferrin). We investigated the expression of the main proteins of iron metabolism in the spinal cord (SC) of totally gastrectomized Cbl-D rats 2 months after surgery (i.e., when the Cbl-D status has become severe). There were no significant changes in iron content, the activity of iron regulatory proteins, or the expression of transferrin or its receptor in the SC. We observed a significant decrease in the levels of both H and L ferritin subunits, with a more marked reduction in the latter. Post-operative cobalamin replacement therapy normalized only the H-ferritin subunits, and only in the SC. Our results therefore suggest that permanent cobalamin deficiency affects iron metabolism in the rat SC preferentially from a functional point of view, because H-ferritin is known to be involved in the uptake and release of iron. PMID- 12111822 TI - Immunoglobulin Fc gamma receptor promotes immunoglobulin uptake, immunoglobulin mediated calcium increase, and neurotransmitter release in motor neurons. AB - Receptors for the Fc portion of immunoglobulin G (IgG; FcgammaRs) facilitate IgG uptake by effector cells as well as cellular responses initiated by IgG binding. In earlier studies, we demonstrated that amyotrophic lateral sclerosis (ALS) patient IgG can be taken up by motor neuron terminals and transported retrogradely to the cell body and can alter the function of neuromuscular synapses, such as increasing intracellular calcium and spontaneous transmitter release from motor axon terminals after passive transfer. In the present study, we examined whether FcgammaR-mediated processes can contribute to these effects of ALS patient immunoglobulins. F(ab')(2) fragments (which lack the Fc portion) of ALS patient IgG were not taken up by motor axon terminals and were not retrogradely transported. Furthermore, in a genetically modified mouse lacking the gamma subunit of the FcR, the uptake of whole ALS IgG and its ability to enhance intracellular calcium and acetylcholine release were markedly attenuated. These data suggest that FcgammaRs appear to participate in IgG uptake into motor neurons as well as IgG-mediated increases in intracellular calcium and acetylcholine release from motor axon terminals. PMID- 12111824 TI - Gamma-aminobutyric acid transporter (BGT-1) expressed in human astrocytoma U373 MG cells: pharmacological and molecular characterization and phorbol ester induced inhibition. AB - The properties of a transport system specific for gamma-aminobutyric acid (GABA) expressed in human U373 MG astrocytoma cells were examined. The uptake of [(3)H]GABA was dependent on both extracellular Na(+) and Cl(-) ions and was inhibited by (+/-)-nipecotic acid, guvacine, and beta-alanine, with a pharmacological profile corresponding to that reported for the human homologue of the GABA/betaine transporter (BGT-1). Accordingly, [(3)H]GABA uptake was also inhibited by betaine, and reverse transcriptase-polymerase chain reaction (RT PCR) analysis of total RNA from U373 MG cells with specific BGT-1 primers resulted in the amplification of a 440 bp fragment that was further characterized by restriction analysis and sequencing. In addition, Western blot analysis with anti-BGT-1 antiserum revealed the presence of a characteristic 60 kDa band. The primary structure of the human BGT-1 protein predicts two putative phosphorylation sites for the Ca(2+)/diacylglicerol-dependent protein kinase (PKC), and treatment of U373 MG cells with the PKC activator phorbol 12-myristate 13-acetate (TPA) led to a concentration- and time-dependent decrease in [(3)H]GABA uptake. The maximal effect was detected at 2 hr of incubation, to disappear after 4 hr. TPA-induced reduction in [(3)H]GABA uptake was reversed by preincubation with staurosporine. Taken together, these results indicate that U373 MG cells express a GABA transporter of the BGT-1 subtype whose function is regulated by phosphorylation events through PKC. PMID- 12111825 TI - d-Amphetamine-induced increase in neurotensin and neuropeptide Y outflow in the ventral striatum is mediated via stimulation of dopamine D1 and D2/3 receptors. AB - The neuroanatomical and functional relationships between dopamine (DA) and neurotensin (NT) and DA and neuropeptide Y (NPY) suggest a role for these neuropeptides in DA-related neuropsychiatric disorders. By employing a microdialysis technique in conjunction with radioimmunoassay (RIA), the effects of d-amphetamine per se or after pretreatment with DA receptor antagonists on NT and NPY outflow were determined in the ventral striatum (VSTR) of the rat. One hour after a subcutaneous (s.c.) injection of saline, the DA-D(1) receptor antagonist SCH 23390 (0.3 mg/kg), or the DA-D(2/3) receptor antagonist raclopride (1.0 mg/kg), animals were injected s.c. with either saline or d-amphetamine (1.5 mg/kg). d-Amphetamine significantly increased extracellular NT- and NPY-like immunoreactivity (LI) concentrations compared with control animals. Administration of SCH 23390 or raclopride did not significantly affect NT-LI or NPY-LI concentrations. However, pretreatment with either SCH 23390 or raclopride abolished the stimulatory effect of d-amphetamine on NT-LI and NPY-LI. These findings demonstrate that d-amphetamine increases extracellular concentrations of NT-LI and NPY-LI in the VSTR through a mechanism that initially involves stimulation of either DA-D(1) or DA-D(2/3) receptors but appears to require both. In conclusion, changes in dopaminergic neurotransmission via DA-D(1) and DA D(2/3) receptors affect the outflow of both NT and NPY in the VSTR. PMID- 12111826 TI - Oligodendrocytes as providers of growth factors. AB - Glial cells recently are being appreciated as supporters of brain neurons. This review addresses their role as growth factor providers. While the function of astrocytes in this capacity is known, new data indicate that oligodendrocytes, the myelinating cells of the brain, exhibit similar abilities. Oligodendrocytes provide trophic signals to nearby neurons and synthesize defined growth factors. Expression of growth factors is influenced by neural signals. The review summarizes these roles and their implications in brain function. PMID- 12111827 TI - A secreted Delta1-Fc fusion protein functions both as an activator and inhibitor of Notch1 signaling. AB - Signaling induced through interactions between DSL (Delta, serrate, LAG-2) ligand signaling cells and Notch-responding cells influences the developmental fate of a wide variety of invertebrate and vertebrate cell types. Consistently with a requirement for direct cell-cell interactions, secreted DSL ligands expressed in flies do not appear to activate Notch signaling but rather produce phenotypes reminiscent of losses in Notch signaling. In contrast, secreted DSL ligands expressed in Caenorhabditis elegans or supplied to mammalian cells in culture produce effects indicative of Notch activation. In fact, engineered secreted DSL ligands have been used to study Notch signaling in neurogenesis, gliogenesis, hematopoeisis, neurite morphogenesis and ligand-induced nuclear translocation of the Notch intracellular domain. Using a recombinant, secreted form of the DSL ligand Delta1, we found that antibody-induced oligomerization (termed "clustering") was required for this soluble ligand to bind specifically to Notch1 expressing cells, undergo internalization, and activate downstream signaling. Interestingly, clustering with either limiting or excess antibody led to ligand binding in the absence of Notch signaling, indicating that ligand binding is necessary but not sufficient for activation of Notch signaling. Moreover, such antibody clustering conditions blocked Notch1 signaling induced by membrane-bound DSL ligands. We propose that multimerization influences whether ligand binding to Notch results in activation or inhibition of downstream signaling and suggest that differences in ligand presentation might account for why secreted forms of DSL ligands have been reported to function as agonists and antagonists of Notch signal transduction. PMID- 12111828 TI - Comparative expression profiles of Trk receptors and Shc-related phosphotyrosine adapters during retinal development: potential roles of N-Shc/ShcC in brain derived neurotrophic factor signal transduction and modulation. AB - Neurotrophins (NTs) have multiple roles in retinal development and survival, which are mediated through their specific receptors and signaling molecules. An emerging family of adapter protein, Shc (Src homology and collagen)-related molecules, i.e., Shc/ShcA, Sck/ShcB, and N-Shc/ShcC, has been implicated in various phosphotyrosine signal transduction mechanisms, including that for NTs. To explore the potential role(s) of Shc-related adapters in NT signaling in the retina, we compared the developmental changes of the mRNA expression of TrkA -B, and -C in the rat retina, on one hand and, on the other hand, studied which members of the Shc family were activated after brain-derived neurotrophic factor (BDNF) application in axotomized rat retinas. Early in development, both TrkA and ShcA were highly expressed, whereas, in late development to adulthood, TrkB/C and ShcB/C were highly expressed. In the mature retinal ganglion cell layer, the expression of ShcB/C and TrkB/C was evident. Immunoreactivity of ShcC was located in the retinal ganglion cells, amacrine cells, and inner plexiform layer. The response of ShcC following retinal axotomy was most profound with the administration of BDNF, and there was some response with neurotrophin-3. These results indicate that ShcC could be a potential phosphotyrosine adapter among the Shc family members for BDNF signaling and function during retinal development and regeneration in vivo. PMID- 12111829 TI - Role of long-range repulsive forces in organizing axonal neurofilament distributions: evidence from mice deficient in myelin-associated glycoprotein. AB - When the axon of a motor neuron is sectioned and visualized by electron microscopy, a two-dimensional distribution of neurofilaments (NFs) with nonrandom spacing is revealed; this ordered arrangement implies the presence of physical interactions between the NFs. To gain insight into the molecular basis of this organization, we characterized NF distributions from mouse sciatic nerve cross sections using two statistical mechanical measures: radial distribution functions and occupancy probability distributions. Our analysis shows that NF organization may be described in terms of effective pairwise interactions. In addition, we show that these statistical mechanical measures can detect differences in NF architecture between wild-type and myelin-associated glycoprotein null mutant mice. These differences are age dependent, with marked contrast between the NF distributions by 9 months of age. Finally, using Monte Carlo simulations, we compare the experimental results with predictions for models in which adjacent NFs interact through rigid cross bridges, deformable cross bridges, and long range repulsive forces. Among the models tested, a model in which the filaments interact through a long-range repulsive force is most consistent with the results of our analysis. PMID- 12111830 TI - Monocyte recruitment and myelin removal are delayed following spinal cord injury in mice with CCR2 chemokine receptor deletion. AB - The inflammatory response initiated after spinal cord injury (SCI) is characterized by the accumulation of macrophages at the impact site. Monocyte chemoattractant protein-1 (MCP-1) is a strong candidate for mediating chemotaxis of monocytes to the injured nervous system. To help in defining the role of MCP-1 in inflammation after SCI, we evaluated the time course of macrophage accumulation for 2 weeks following a midthoracic spinal cord contusion injury in mice lacking CCR2, a principal receptor for MCP-1. Mice with a deletion of CCR2 resulted in significantly reduced Mac-1 immunoreactivity restricted to the lesion epicenter at 7 days postinjury. The regions devoid of Mac-1 immunoreactivity corresponded to areas of reduced myelin degradation at this time. By 14 days postinjury, however, there were no differences in Mac-1 staining between CCR2 (+/+) and CCR2 (-/-) mice. Analyses of mRNA levels by RNase protection assay (RPA) revealed increases in MCP-1 as well as MCP-3 and MIP-2 mRNA at 1 day postinjury compared with 7 day postinjury. There were no differences in chemokine expression between CCR2-deficient mice and wild-type littermate controls. The CCR2-deficient mice also exhibited reduced expression of mRNA for chemokine receptors CCR1 and CCR5. Together, these results indicate that chemokines acting through CCR2 contribute to the early recruitment of monocytes to the lesion epicenter following SCI. PMID- 12111831 TI - SHP-1 expression in avian mixed neural/glial cultures. AB - The central nervous system response to injury includes astrocyte proliferation and hypertrophy as well as microglial activation and proliferation. However, not all glial cells enter the cell cycle following damage, and the mechanism that determines which glial cells will proliferate and which will remain quiescent has yet to be elucidated. Protein tyrosine phosphorylation has been shown to play an important role in the regulation of the cell cycle in a number of different systems and has been implicated in both astrocyte proliferation and differentiation. Of particular interest is the protein tyrosine phosphatase SHP-1 (Src homology 2-containing protein tyrosine phosphatase 1), which: (1) modulates cellular proliferation in the hematopoietic system, (2) is involved in various growth factor second messenger signaling cascades, and (3) has been demonstrated by our laboratory to increase in immunoreactivity within a subpopulation of astrocytes following deafferentation of the chicken auditory brainstem. These SHP 1+ cells appear to be those which fail to enter the cell cycle following deafferentation. The present study examines whether manipulation of cellular proliferation in vitro modifies the expression of SHP-1 immunoreactivity in mixed neural/glial cultures of the avian auditory brainstem. In addition, the effect of the protein tyrosine phosphatase inhibitor sodium orthovanadate on cellular proliferation was assessed in these cultures. Our results demonstrate that SHP-1 expression can be modulated by changes in proliferation and that inhibiting tyrosine phosphatase activity results in increased proliferation. Taken together, these results indicate that SHP-1 may play central role in negatively regulating glial proliferation following injury. PMID- 12111832 TI - Striatal neurochemical changes in transgenic models of Huntington's disease. AB - Transgenic mouse models of Huntington's disease (HD) were examined following the onset of overt behavioral symptoms. The HD transgenic mice demonstrated profound striatal losses in D1, D2, and D3 dopamine (DA) receptor proteins in comparison with their nonsymptomatic, age-matched littermate controls. In parallel, a robust increase in the striatal D5 DA receptor subtype occurred in the transgenic compared with the wild-type control mice. This receptor elevation was accompanied by heightened cyclic AMP levels, which may be induced by the adenylyl cyclase linked D5 receptor. This is a unique result; normal striatal D5 protein levels are modest and not thought to contribute substantially to cyclic AMP-mediated DA signaling mechanisms. Simple compensatory up-regulation of D5 DA receptors in response to D1 receptor subtype loss does not explain our findings, because genetic inactivation of the D1 DA receptor does not alter levels of D5 DA receptor expression. Immunofluorescent detection of tyrosine hydroxylase showed that nigrostriatal DA containing terminals were reduced, further supporting that disturbances in DA signaling occurred in HD transgenic models. The substance P containing striatal efferent pathway was more resistant to the HD mutation than met-enkephalin-producing striatal projection neurons in the transgenics, based on neuropeptide immunofluorescent staining. Analogous findings in multiple transgenic models suggest that these changes are due to the presence of the transgene and are not dependent on its composition, promotor elements, or mouse strain background. These findings suggest modifications in the striatal DA system and that its downstream signaling through cyclic AMP mechanisms is disrupted severely in HD following onset of motor symptoms. PMID- 12111834 TI - Studies on neuronal death in the mouse model of Niemann-Pick C disease. AB - A mouse model of Niemann-Pick disease type C (NPC) carries a genetic defect that causes biochemical changes in lipid levels and a progressive neuropathology that parallels the effects of NPC disease in humans. It is a moot point whether or not the loss of Purkinje and other neuronal cells proceeds by apoptotic death. Therefore, we have introduced into these mice a transgene expressing human Bcl-2 protein which has previously been demonstrated to prevent developmental neuronal death and death induced by a variety of stimuli. The human Bcl-2 transgene was driven by the neuron-specific enolase promoter and was abundantly expressed in Purkinje and other neuronal cells. npc1(-/-)/bcl-2 transgenic mice did not show a significant delay in the onset of neurological disorders. Neuropathological examination of the npc1(-/-)/bcl-2 transgenic mice did not disclose significant differences in numbers of surviving Purkinje cells between the npc1(-/-), tg(+) and npc1(-/-), tg(-) mice. When the npc1(-/-) mice were treated with minocycline, a drug which was shown to inhibit apparent apoptotic death in other mouse models of neurological disease, no delay in onset of neurological disorders were observed in either npc1(-/-), or npc1(-/-) /mdrla(-/-) mice (mdr1a deficiency was used to enhance brain availability of minocycline). Caspase-1 levels were not altered in npc1(-/-) mice, with or without minocycline treatment. These results suggest that Purkinje cell loss in npc1(-/-) mice does not proceed by an apoptotic pathway that can be inhibited by Bcl-2 or minocycline. PMID- 12111833 TI - GeneChip analysis shows altered mRNA expression of transcripts of neurotransmitter and signal transduction pathways in the cerebral cortex of portacaval shunted rats. AB - Identifying the gene expression changes induced by hepatic encephalopathy (HE) leads to a better understanding of the molecular mechanisms of HE-induced neurological dysfunction. Using GeneChip and real-time PCR, the present study evaluated the gene expression profile of rat cerebral cortex at 4 weeks after portacaval shunting. Among 1,263 transcripts represented on the chip, mRNA levels of 31 transcripts were altered (greater than twofold; 16 increased and 15 decreased) in the portacaval shunted (PCS) rat compared to sham control. Changes observed by GeneChip analysis were confirmed for 20 transcripts (8 increased, 7 decreased, and 5 unchanged in PCS rat brain) by real-time PCR. Neurotransmitter receptors, transporters, and members of the second messenger signal transduction are the major groups of genes altered in PCS rat brain. Of importance was that the increased heme oxygenase-1 and decreased Cu,Zn-superoxide dismutase expression observed raise the possibility of oxidative stress playing a pathogenic role in chronic HE. PMID- 12111835 TI - Expression of inducible nitric oxide synthase and cyclooxygenase-2 after excitotoxic damage to the immature rat brain. AB - It is well established that after adult brain damage the enzymes cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) play an important role in the inflammatory processes and oxidative stress, which are considered to be the leading factors contributing to delayed cell death. The contribution of these enzymes to postnatal brain damage, however, is poorly understood. In our study, excitotoxic lesions were induced by the injection of N-methyl-D-aspartate in the cortex of postnatal day 9 rats. After different survival times ranging from 4 hr to 7 days post-lesion, brain sections were processed for the immunocytochemical demonstration of COX-2 and iNOS and double labeling with neuronal, glial and neutrophil markers. First and maximal de novo induction of iNOS and COX-2 expression was found at 10 hr post-lesion. Expression of both enzymes started to diminish at 24 hr, reaching basal levels at day 3. iNOS-expressing cells were mainly identified as infiltrated neutrophils as well as highly ramified protoplasmic astrocytes closely associated with blood vessels. Moreover, scattered iNOS-positive neurons were found at the lesion borders. In contrast, COX-2 was mainly observed in reactive microglial cells and neuronal cells. COX-2 positive neurons were found within the degenerating area at 10 hr and at the borders of the lesion later on. This study shows that maximal iNOS and COX-2 expression precedes the period of massive neuronal death observed at 24 hr post lesion, and may therefore contribute to the evolution of the inflammatory response and the neurodegenerative process after an excitotoxic lesion to the postnatal brain. PMID- 12111836 TI - Specific up-regulation of GADD153/CHOP in 1-methyl-4-phenyl-pyridinium-treated SH SY5Y cells. AB - Growth arrest DNA damage-inducible 153 (GADD153) expression was increased in 1 methyl-4-phenyl-pyridinium (MPP(+))-treated human SH-SY5Y neuroblastoma cells as determined by gene microarray analysis. GADD153 expression increased after 24 hr of MPP(+) (1 mM) exposure and preceded activation of caspase 3. Comparison of GADD153 expression among cultures treated with other toxins whose primary mode of action is either via mitochondrial impairment (rotenone) or via oxidative stress (6-hydroxydopamine or hydrogen peroxide) showed that GADD153 was uniquely up regulated by MPP(+). Together these data suggest that a cellular mechanism distinct from mitochondrial impairment or oxidative stress contributes significantly to the up-regulation of GADD153 by MPP(+) and that GADD153 may function as an inducer of apoptosis following MPP(+) exposure. Published 2002 Wiley-Liss, Inc. PMID- 12111837 TI - Developmental changes in the expression of iron regulatory proteins and iron transport proteins in the perinatal rat brain. AB - The perinatal brain requires a tightly regulated iron transport system. Iron regulatory proteins (IRPs) 1 and 2 are cytosolic proteins that regulate the stability of mRNA for the two major cellular iron transporters, transferrin receptor (TfR) and divalent metal transporter-1 (DMT-1). We studied the localization of IRPs, their change in expression during perinatal development, and their relationship to TfR and DMT-1 in rat brain between postnatal days (PND) 5 and 15. Twelve-micron frozen coronal sections of fixed brain tissue were obtained from iron-sufficient Sprague-Dawley rat pups on PND 5, 10, and 15, and were visualized at 20 to 1,000x light microscopy for diaminobenzidine activity after incubation with specific primary IRP-1, IRP-2, DMT-1, and TfR antibodies and a universal biotinylated secondary and tertiary antibody system. IRP and transport protein expression increased in parallel over time. IRP1, IRP2, and DMT 1 were partially expressed in the choroid plexus epithelial cells at PND 5 and 10, and fully expressed at PND 15. The cerebral blood vessels and ependymal cells strongly expressed IRP1, IRP2, and DMT-1 as early as PND 5. Substantive TfR staining was not seen in the choroid plexus or ependyma until PND 15. Glial and neuronal expression of IRP1, IRP2, DMT-1, and TfR in cortex, hippocampal subareas and striatum increased over time, but showed variability in cell number and intensity of expression based on brain region, cell type, and age. These developmental changes in IRP and transporter expression suggest potentially different time periods of brain structure vulnerability to iron deficiency or iron overload. PMID- 12111838 TI - Elevation of brain glutathione by gamma-glutamylcysteine ethyl ester protects against peroxynitrite-induced oxidative stress. AB - Elevation of glutathione (GSH) has been recognized as an important method for modulating levels of reactive oxygen species (ROS) in the brain. We investigated the antioxidant properties of gamma-glu-cys-ethyl ester (GCEE) in vitro and its ability to increase GSH levels upon in vivo i.p. injection. GCEE displays antioxidant activity similar to GSH as assessed by various in vitro indices such as hydroxyl radical scavenging, dichlorofluorescein fluorescence (DCF), protein specific spin labeling, glutamine synthetase (GS) activity, and protein carbonyls. Intraperitoneal injection of GCEE to gerbils resulted in a 41% increase in brain total GSH levels in vivo as determined by the DTNB-GSH reductase recycling method. Gerbils injected with buthionine sulfoximine (BSO), an inhibitor of gamma-glutamylcysteine synthetase, had 40% less total brain glutathione. Gerbils injected with BSO followed by a GCEE injection had GSH levels similar to vehicle-injected controls, suggesting that GCEE upregulates GSH biosynthesis by providing gamma-glutamylcysteine and not cysteine. Cortical synaptosomes from GCEE-injected animals were less susceptible to peroxynitrite induced oxidative damage as assessed by DCF fluorescence, protein-specific spin labeling, and GS activity. These experiments suggest that GCEE is effective in increasing brain GSH levels and may potentially play an important therapeutic role in attenuating oxidative stress in neurodegenerative diseases associated with oxidative stress such as Alzheimer disease. PMID- 12111839 TI - Chronological changes in pyridoxine-5'-phosphate oxidase immunoreactivity in the seizure-sensitive gerbil hippocampus. AB - To identify the roles of pyridoxine-5'-phosphate (PNP) oxidase in epileptogenesis and the recovery mechanisms in spontaneous seizure, a chronological and comparative analysis of PNP oxidase expression was conducted. PNP oxidase immunoreactivity in a preseizure group of seizure-sensitive (SS) gerbils was detected more strongly than that in a seizure-resistant (SR) group. The density of PNP oxidase immunoreactivity in a 30 min postictal group was significantly lower than that in a preseizure group. In a 12 hr postictal group, PNP oxidase immunodensity had recovered to a preseizure level. The overexpression of PNP oxidase in the hippocampus of preseizure SS gerbils suggests that PNP or pyridoxal 5'-phosphate plays an important role in the modulation of glutamic acid decarboxylase activity and gamma-aminobutyric acid reuptake as mediated by membrane transporter via neurons. In addition, this change in the PNP oxidase immunoreactivity following seizure may be a compensatory response designed to reduce epileptic activity in this animal. PMID- 12111840 TI - Stem cell repair of central nervous system injury. AB - Neural stem cells (NSCs) have great potential as a therapeutic tool for the repair of a number of CNS disorders. NSCs can either be isolated from embryonic and adult brain tissue or be induced from both mouse and human ES cells. These cells proliferate in vitro through many passages without losing their multipotentiality. Following engraftment into the adult CNS, NSCs differentiate mainly into glia, except in neurogenic areas. After engraftment into the injured and diseased CNS, their differentiation is further retarded. In vitro manipulation of NSC fate prior to transplantation and/or modification of the host environment may be necessary to control the terminal lineage of the transplanted cells to obtain functionally significant numbers of neurons. NSCs and a few types of glial precursors have shown the capability to differentiate into oligodendrocytes and to remyeliate the demyelinated axons in the CNS, but the functional extent of remyelination achieved by these transplants is limited. Manipulation of endogenous neural precursors may be an alternative therapy or a complimentary therapy to stem cell transplantation for neurodegenerative disease and CNS injury. However, this at present is challenging and so far has been unsuccessful. Understanding mechanisms of NSC differentiation in the context of the injured CNS will be critical to achieving these therapeutic strategies. PMID- 12111841 TI - Hippocampal brain-derived neurotrophic factor gene regulation by exercise and the medial septum. AB - Brain-derived neurotrophic factor (BDNF) enhances synaptic plasticity and neuron function. We have reported that voluntary exercise increases BDNF mRNA levels in the hippocampus; however, mechanisms underlying this regulation have not been defined. We hypothesized that medial septal cholinergic and/or gamma amino butyric acid (GABA)ergic neurons, which provide a major input to the hippocampus, may regulate the baseline gene expression and exercise-dependent gene upregulation of this neurotrophin. Focal lesions were produced by medial septal infusion of the saporin-linked immunotoxins 192-IgG-saporin or OX7-saporin. 192 IgG-saporin produced a selective and complete loss of medial septal cholinergic neurons with no accompanying GABA loss. Baseline BDNF mRNA was reduced in the hippocampus of sedentary animals, but exercise-induced gene upregulation was not impaired, despite complete loss of septo-hippocampal cholinergic afferents. OX7 saporin produced a graded lesion of the medial septum characterized by predominant GABA neuron loss with less reduction in the number of cholinergic cells. OX7-saporin lesion reduced baseline hippocampal BDNF mRNA and attenuated exercise-induced gene upregulation, in a dose-dependent manner. These results suggest that combined loss of septal GABAergic and cholinergic input to the hippocampus may be important for exercise-dependent BDNF gene regulation, while cholinergic activity on its own is not sufficient. These results are discussed in relation to their implications for aging and Alzheimer's disease. PMID- 12111842 TI - Cellular mechanisms of connexin32 mutations associated with CNS manifestations. AB - Both oligodendrocytes and myelinating Schwann cells express the gap junction protein connexin32 (Cx32). Mutations in the gene encoding Cx32 (GJB1) cause the X linked form of Charcot-Marie-Tooth disease (CMTX). Although most CMTX patients do not have clinical central nervous system (CNS) manifestations, subclinical evidence of CNS dysfunction is common. We investigated the cellular effects of a subgroup of GJB1/Cx32 mutations that have been reported to cause clinical CNS dysfunction. We hypothesized that these mutants have dominant-negative effects on other connexins expressed by oligodendrocytes, specifically Cx45. We expressed these and other Cx32 mutants in communication-incompetent as well as Cx45 expressing HeLa cells, and analyzed the transfected cells by immunocytochemistry and immunoblotting. In communication-incompetent cells, the mutants associated with CNS phenotypes failed to reach the cell membrane and were instead retained in the endoplasmic reticulum (A39V, T55I) or Golgi apparatus (M93V, R164Q, R183H), although rare gap junction plaques were found in cells expressing M93V or R183H. In HeLa cells stably expressing Cx45, these Cx32 mutants showed a similar expression pattern, and did not alter the pattern of Cx45 expression. These results indicate that Cx32 mutants that are associated with a CNS phenotype do not interact with Cx45, but may instead have other toxic effects in oligodendrocytes. PMID- 12111843 TI - Regulation and subcellular localization of the microtubule-destabilizing stathmin family phosphoproteins in cortical neurons. AB - Stathmin is a ubiquitous cytosolic phosphoprotein, preferentially expressed in the nervous system, and the generic element of a protein family that includes the neural-specific proteins SCG10, SCLIP, and RB3 and its splice variants, RB3' and RB3". All phosphoproteins of the family share with stathmin its tubulin binding and microtubule (MT)-destabilizing activities. To understand better the specific roles of these proteins in neuronal cells, we performed a comparative study of their expression, regulation, and intracellular distribution in embryonic cortical neurons in culture. We found that stathmin is highly expressed ( approximately 0.25% of total proteins) and uniformly present in the various neuronal compartments (cell body, dendrites, axon, growth cones). It appeared mainly unphosphorylated or weakly phosphorylated on one site, and antisera to specific phosphorylated sites (serines 16, 25, or 38) did not reveal a differential regulation of its phosphorylation among neuronal cell compartments. However, they revealed a subpopulation of cells in which stathmin was highly phosphorylated on serine 16, possibly by CaM kinase II also active in a similar subpopulation. The other proteins of the stathmin family are expressed about 100 fold less than stathmin in partially distinct neuronal populations, RB3 being detected in only about 20% of neurons in culture. In contrast to stathmin, they are each mostly concentrated at the Golgi apparatus and are also present along dendrites and axons, including growth cones. Altogether, our results suggest that the different members of the stathmin family have complementary, at least partially distinct functions in neuronal cell regulation, in particular in relation to MT dynamics. PMID- 12111844 TI - Rev-erbalpha gene expression in the mouse brain with special emphasis on its circadian profiles in the suprachiasmatic nucleus. AB - Rev-erbalpha is an orphan nuclear receptor that constitutively suppresses gene transcription. In the present study, the expression of Rev-erbalpha was investigated in the mouse brain by in situ hybridization using antisense cRNA probe. Positive Rev-erbalpha mRNA signals were detected widely in the brain with the highest expression in the suprachiasmatic nucleus (SCN). In the constant dark condition, the circadian expression profiles of Rev-erbalpha m RNA in the SCN showed a peak at early daytime (CT4) and a trough at early night time (CT16). The environmental lighting condition (light-dark environmental condition and exposure in the subjective night) did not alter the expression profiles. These findings indicate that Rev-erbalpha gene is a transcription factor intimately related to the circadian clock in the SCN. PMID- 12111845 TI - Glutamate signalling and secretory phospholipase A2 modulate the release of arachidonic acid from neuronal membranes. AB - The lipid mediators generated by phospholipases A(2) (PLA(2)), free arachidonic acid (AA), eicosanoids, and platelet-activating factor, modulate neuronal activity; when overproduced, some of them become potent neurotoxins. We have shown, using primary cortical neuron cultures, that glutamate and secretory PLA(2) (sPLA(2)) from bee venom (bv sPLA(2)) and Taipan snake venom (OS2) elicit synergy in inducing neuronal cell death. Low concentrations of sPLA(2) are selective ligands of cell-surface sPLA(2) receptors. We investigated which neuronal arachidonoyl phospholipids are targeted by glutamate-activated cytosolic calcium-dependent PLA(2) (cPLA(2)) and by sPLA(2). Treatment of (3)H-AA-labeled cortical neurons with mildly toxic concentrations of sPLA(2) (25 ng/ml, 1.78 nM) for 45 min resulted in a two- to threefold higher loss of (3)H-AA from phosphatidylcholine (PC) than from phosphatidylethanolamine (PE) and in minor changes in other phospholipids. A similar profile, although of greater magnitude, was observed 20 hr posttreatment. Glutamate (80 microM) induced much less mobilization of (3)H-AA than did sPLA(2) and resulted in a threefold greater degradation of (3)H-AA PE than of (3)H-AA PC by 20 hr posttreatment. Combining sPLA(2) and glutamate resulted in a greater degradation of PC and PE, and the N methyl-D-aspartate receptor antagonist MK-801 only blocked glutamate effects. Thus, activation of the arachidonate cascade induced by glutamate and sPLA(2) under experimental conditions that lead to neuronal cell death involves the hydrolysis of different (perhaps partially overlapping) cellular phospholipid pools. PMID- 12111846 TI - Differential neuropathological alterations in transgenic mice expressing alpha synuclein from the platelet-derived growth factor and Thy-1 promoters. AB - Accumulation of alpha-synuclein has been associated with neurodegenerative disorders, such as Lewy body disease and multiple system atrophy. We previously showed that expression of wild-type human alpha-synuclein in transgenic mice results in motor and dopaminergic deficits associated with inclusion formation. To determine whether different levels of human alpha-synuclein expression from distinct promoters might result in neuropathology mimicking other synucleopathies, we compared patterns of human alpha-synuclein accumulation in the brains of transgenic mice expressing this molecule from the murine Thy-1 and platelet-derived growth factor (PDGF) promoters. In murine Thy-1-human alpha synuclein transgenic mice, this protein accumulated in synapses and neurons throughout the brain, including the thalamus, basal ganglia, substantia nigra, and brainstem. Expression of human alpha-synuclein from the PDGF promoter resulted in accumulation in synapses of the neocortex, limbic system, and olfactory regions as well as formation of inclusion bodies in neurons in deeper layers of the neocortex. Furthermore, one of the intermediate expressor lines (line M) displayed human alpha-synuclein expression in glial cells mimicking some features of multiple system atrophy. These results show a more widespread accumulation of human alpha-synuclein in transgenic mouse brains. Taken together, these studies support the contention that human alpha-synuclein expression in transgenic mice might mimic some neuropathological alterations observed in Lewy body disease and other synucleopathies, such as multiple system atrophy. PMID- 12111847 TI - Interleukin-4, interleukin-10, and interleukin-1-receptor antagonist but not transforming growth factor-beta induce ramification and reduce adhesion molecule expression of rat microglial cells. AB - The activity of microglial cells is strictly controlled in order to maintain central nervous system (CNS) immune privilege. We hypothesized that several immunomodulatory factors present in the CNS parenchyma, i.e., the Th2-derived cytokines interleukin (IL)-4 and IL-10, interleukin-1-receptor-antagonist (IL-1 ra), or transforming growth factor (TGF)-beta can modulate microglial morphology and functions. Microglial cells were incubated with IL-4, IL-10, IL-1-ra, TGF beta, or with astrocyte conditioned media (ACM) and were analyzed for morphological changes, expression of intercellular adhesion molecule (ICAM)-1, and secretion of IL-1beta or tumor necrosis factor (TNF)-alpha. Whereas untreated controls showed an amoeboid morphology both Th2-derived cytokines, IL-1-ra, and ACM induced a morphological transformation to the ramified phenotype. In contrast, TGF-beta-treated microglial cells showed an amoeboid morphology. Even combined with the neutralizing antibodies against IL-4, IL-10, or TGF-beta ACM induced microglial ramification. Furthermore, ACM did not contain relevant amounts of IL-4 and IL-10, as measured by enzyme-linked immunosorbent assay (ELISA). Flow cytometry showed that lipopolysaccharide (LPS)-induced ICAM-1 expression on microglial cells was strongly suppressed by ACM, significantly modulated by IL-4, IL-10, or IL-1-ra, but not influenced by TGF-beta. The LPS induced secretion of IL-1beta and TNF-alpha was only reduced after application of ACM, whereas IL-4 or IL-10 did not inhibit IL-1beta- or TNF-alpha secretion. TGF beta enhanced IL-1beta- but not TNF-alpha secretion. In summary, we demonstrate that IL-4, IL-10, and IL-1-ra induce microglial ramification and reduce ICAM-1 expression, whereas the secretion of proinflammatory cytokines is not prevented. TGF-beta has no modulating effects. Importantly, unidentified astrocytic factors that are not identical with IL-4, IL-10, or TGF-beta possess strong immunomodulatory properties. PMID- 12111848 TI - Establishment and characterization of spontaneously immortalized Schwann cells from murine model of globoid cell leukodystrophy (twitcher). AB - The twitcher mouse is a murine model of human globoid cell leukodystrophy (GLD; Krabbe disease) caused by a genetic defect in the activity of galactosylceramidase (GALC). An accumulation of cytotoxic metabolite, galactosylsphingosine (psychosine), in myelin forming cells (oligodendrocytes and Schwann cells) of the twitcher mouse as well as patients with GLD has been suggested to cause dysfunction of these cells and subsequent demyelination in the central and peripheral nervous system. To investigate further the cellular pathomechanism of GLD, we established spontaneously immortalized Schwann cell lines from the twitcher mouse. Long-term cultures of Schwann cells derived from dorsal root ganglia and consecutive peripheral nerves of 3-week-old twitcher mice were maintained for 6 months, and spontaneously developed colonies were expanded further and characterized. One of the cell lines, designated TwS1, showed distinct Schwann cell phenotypes, was passaged twice a week and maintained for over 10 months without phenotypic alterations. The TwS1 cells had a nonsense mutation in the GALC genome, and showed markedly reduced GALC activity and elevated psychosine levels. Ultrastructurally, varieties of cytoplasmic inclusions were demonstrated in TwS1 cells. When TwS1 cells were infected with a retrovirus vector encoding GALC, GALC activity was markedly increased and psychosine levels were significantly decreased. These immortalized Schwann cells can be useful in studies on the nervous system lesions in GLD. PMID- 12111849 TI - Effects of locally administered pentylenetetrazole on nigral single unit activity and severity of dystonia in a genetic model of paroxysmal dystonia. AB - The dt(sz) hamster is a well-established animal model of idiopathic paroxysmal dystonia. Previous investigations of this mutant have indicated dysfunctions of the gamma-aminobutyric acid (GABA)-ergic system within the basal ganglia. Systemic administration of the central stimulant pentylenetetrazole (PTZ) aggravated dystonia at subconvulsant doses, whereas GABA-mimetic drugs have beneficial effects in dt(sz) hamsters. GABA mimetics also provide clinical benefit in humans with idiopathic paroxysmal dystonia. The spontaneous discharge rates of substantia nigra pars reticulata (SNr) neurons was unaltered in anesthetized dt(sz) hamsters, but systemic application of subconvulsant doses of PTZ caused significantly greater increases of discharge rates in dystonic hamsters compared with nondystonic controls. The present study tested the hypothesis that SNr neurons are more sensitive to local application of PTZ in dt(sz) hamsters than in nondystonic hamsters. PTZ applied locally by pressure injection at 2, 3, and 5 mM to the SNr during in vivo single unit recordings revealed a dose-dependent increase of SNr discharge rates in mutants and controls relative to predrug rates, with a significantly greater increase in mutants at 3 mM PTZ. To examine the functional relevance of the increased susceptibility of SNr neurons to PTZ in mutants, the effects of PTZ on severity of dystonia were investigated after microinjections into the SNr of freely moving dt(sz) hamsters. Bilateral nigral microinjection of 40 ng PTZ did not aggravate dystonia but exerted moderate antidystonic effects. Therefore, the previous findings of prodystonic effects of systemic administration of PTZ in dt(sz) hamsters are related to extranigral effects rather than to the elevation of nigral discharge rates in response to systemic, or locally applied, PTZ. The greater susceptibility of neurons within the SNr to PTZ suggests dysfunctions of the GABA(A) receptor in dt(sz) mutants. PMID- 12111850 TI - Characterization of astrocytes derived from human NTera-2/D1 embryonal carcinoma cells. AB - Astrocytes are the predominant cell type in the vicinity of glutamatergic synapses, where they monitor and maintain low levels of glutamate. Synaptic homeostasis of glutamate involves its removal from the synaptic cleft via high affinity glutamate transporters, glutamate transporter-1 (GLT-1)/excitatory amino acid transporters (EAAT)2 and glutamate and aspartate transporter (GLAST)/EAAT1, and glutamate-catabolizing enzyme, glutamine synthase. Glutamate transporters have been mostly characterized in rodent astrocytes, due to the lack of a convenient human cell system. We report here that NTera-2 (NT2/D1, a cell line derived from a human teratocarcinoma and known to differentiate into neurons) can also be differentiated by a 4-week treatment with retinoic acid into functional astrocytes (NT2/A). Differentiation was accompanied by decreased cell proliferation and cell-cycle arrest, as measured by flow cytometry, immunostaining for Ki67 and incorporation of 5-bromo-2'deoxyuridine (BrdU). Immunocytochemistry and Western blot analysis showed that NT2/A expressed glial fibrillary acidic protein, vimentin and S100beta. Reverse transcription polymerase chain reaction (PCR) detected mRNA encoding glutamate transporters GLT 1/EAAT2 and GLAST/EAAT1. The expression level of GLAST/EAAT1 was higher than that of GLT-1/EAAT2, which is a typical expression pattern for primary astrocytes. Functionality of the transporters was demonstrated by the uptake of (3)H glutamate. NT2/A also expressed active glutamine synthase, and treatment with glutamate (up to 1 mM for 24 hr) was non-toxic, suggesting that these cells were capable of converting it to non-toxic metabolites. NT2/A and NT2-derived neurons could be grown as mixed cultures and this may prove to be a useful experimental model to study molecular mechanisms underlying glutamate excitotoxicity. PMID- 12111851 TI - Recovery of deficient cholinergic calcium signaling by adenosine in cultured rat cortical astrocytes. AB - The regulation of the cholinergic calcium signaling in astroglial cells is thought to play a crucial role in the pathogenesis of Alzheimer's disease. We investigated the action of the cell modulator adenosine on acetylcholine (Ach) mediated intracellular calcium ([Ca(2+)](i)) transients in cultured rat cortical astrocytes using the Ca(2+) imaging technique. The stable adenosine analog 2 chloroadenosine (2ClA) potentiated the [Ca(2+)](i) rise induced by activation of muscarinic Ach receptors by shifting approximately 30-fold the half-effective Ach concentration. This 2ClA effect was maintained upon removal of extracellular Ca(2+), indicating that Ach-induced [Ca(2+)](i) elevation was due mainly to Ca(2+) mobilization from intracellular stores. Pharmacological studies demonstrated that the 2ClA action was mediated by A1 receptors. Incubation with pertussis toxin abrogated the 2ClA effect but left unchanged the [Ca(2+)](i) rise produced by Ach alone. The [Ca(2+)](i) response elicited by Ach alone was abolished upon blockade of muscarinic receptor subtypes that stimulate phospholipase C, whereas the [Ca(2+)](i) elevation generated by the combined action of subthreshold Ach and 2ClA was not affected. Collectively, these results suggest that the impaired cholinergic signaling, the cardinal symptom of Alzheimer's disease, can be reinforced at the second messenger level by an alternative intracellular Ca(2+) mobilizing path, which can be brought into play by the concomitant activation of A1 purinoceptors and muscarinic receptors negatively coupled to adenylyl cyclase. PMID- 12111852 TI - Regulation of kynurenic acid synthesis in C6 glioma cells. AB - Studies with brain slices have provided evidence that synthesis of kynurenic acid (KYNA) from kynurenine (KYN), which occurs in astrocytes, is modulated by changes in the ionic composition of the medium and the presence of depolarizing agents or the excitatory amino acid glutamate (Glu). The present study analyzed the effects of changes in incubation medium on KYNA synthesis in cultured C6 glioma cells. The synthesis was not affected by omission of Na(+) and raising K(+) concentration to 50 mM, conditions that in brain slices stimulate or inhibit KYNA formation, respectively. KYNA synthesis in C6 cells was inhibited by the absence of Ca(2+), which contrasts with its Ca(2+) independence in brain slices. Also, lack of Mg(2+) and addition of a chloride channel blocker, 4-acetamido-4' isothiocyanatostilbene-2,2'-disulfonate (SITS), did not affect the synthesis. KYNA synthesis in C6 cells was dose dependently inhibited by Glu. The inhibitory effect of Glu was not affected by GDPbetaS, an antagonist of metabotropic Glu receptors, the receptor class prevailing in C6 cells, suggesting that Glu acted intracellularly. NH(4)Cl and veratridine decreased KYNA production, mirroring the effects noted in brain slices. KYNA synthesis was strongly reduced in the presence of leucine (Leu), and the uptake of [(14)C]Leu was inhibited by the KYNA precursor KYN, which points to Leu as a potential endogenous modulator of KYNA formation in CNS cells. PMID- 12111854 TI - Acute administration of Ginkgo biloba extract (EGb 761) affords neuroprotection against permanent and transient focal cerebral ischemia in Sprague-Dawley rats. AB - We examined the neuroprotective action of a standardized extract of Ginkgo biloba leaves (EGb 761) in permanent and transient middle cerebral artery (MCA) occlusion models in Sprague-Dawley rats. Forty-four animals were given either EGb 761 (50-200 mg/kg) or vehicle intraperitoneally, 1 hr before permanent MCA occlusion, to evaluate the dose-response effects. An additional 58 animals received EGb 761 (200 mg/kg) or vehicle, 0.5- 4 hr after permanent MCA occlusion, for establishing the therapeutic window. Delayed treatment was also employed in 110 animals treated with either EGb 761 (100-200 mg/kg) or vehicle at 2-3 hr following transient focal cerebral ischemia induced by MCA occlusion for 2 hr. Neurobehavioral scores were determined 22-24 hr after permanent MCA occlusion and either 3 or 7 days after transient MCA occlusion, and brain infarction volumes were measured upon sacrifice. Local cortical blood flow (LCBF) was serially measured in a subset of animals receiving EGb 761 (100-200 mg/kg) or vehicle, 0.5 hr and 2 hr after permanent and transient MCA occlusion, respectively. Relative to vehicle-treated controls, rats pretreated with EGb761 (100 and 200 mg/kg) had significantly reduced infarct volumes, by 36% and 49%, respectively, and improved sensory behavior (P < 0.05). Delayed treatment with EGb 761 also significantly reduced brain infarction, by 20-29% and 31%, when given up to 2 and 3 hr following transient and permanent MCA occlusion, respectively, whereas improved neurobehavioral scores were noted up to 2 hr after the onset of MCA occlusion (P < 0.05). LCBF was significantly improved in the ipsilateral cortex following the EGb 761 treatment, whereas a higher dose showed a more sustained effect. In conclusion, EGb 761 protected against transient and permanent focal cerebral ischemia and was effective after a prolonged reperfusion period even when therapy is delayed up to 2 hr. This neuroprotection may be at least partially attributed to the beneficial effects of selectively improved LCBF in the area at risk of infarction. PMID- 12111853 TI - Mevastatin induces degeneration and decreases viability of cAMP-induced differentiated neuroblastoma cells in culture by inhibiting proteasome activity, and mevalonic acid lactone prevents these effects. AB - Statins with a closed-ring structure (mevastatin, lovastatin, and simvastatin) and with an open-ring structure (pravastatin and fluvastatin) are widely used in the human population to manage hypercholesterolemia. These statins may have neuroprotective or neurotoxic effects, but these effects remain controversial. We have utilized adenosine 3',5'-cyclic monophosphate-induced terminally differentiated murine neuroblastoma (NB) cells in culture as an experimental model to study the effect of statins. Results showed that mevastatin induced degenerative changes and reduced the viability of differentiated NB cells by inhibiting proteasome activity. Lactacystin, an established inhibitor of proteasome, also produced similar degenerative changes in these cells. In contrast, pravastatin neither affected the degeneration and viability of differentiated NB cells nor the proteasome activity. High-performance liquid chromatography (HPLC) analysis of the extract obtained from mevastatin-treated growth medium and differentiated cells revealed that about 50% of mevastatin is converted to an open-ring structure in the growth medium; however, differentiated cells did not convert any portion of mevastatin into an open-ring structure and accumulated only mevastatin with a closed-ring structure. Mevalonic acid lactone by itself did not affect the viability of differentiated NB cells or the proteasome activity, but it completely prevented mevastatin-induced degeneration and decreased viability by reducing the uptake of mevastatin and by blocking its action on proteasome activity. Mevalonic acid failed to prevent lactacystin induced degeneration and inhibition of proteasome activity. Our results suggest that mevastatin could act as a neurotoxic agent or neuroprotective agent, depending upon the extent of its hydrolysis to an open-ring structure and the level of mevalonic acid. PMID- 12111855 TI - A population of oligodendrocytes derived from multipotent neural precursor cells expresses a cholinergic phenotype in culture and responds to ciliary neurotrophic factor. AB - Because oligodendrocytes and their precursors possess receptors for classical transmitters, and neurotransmitters such as glutamate and noradrenaline can mediate oligodendroglial proliferation and differentiation, it is possible that other neurotransmitters can also exert regulatory roles in oligodendrocyte function. We used mitogen-proliferated multipotent neuroepithelial precursors (neurospheres) and identified oligodendroglia that expressed markers traditionally found in cholinergic neurons. Regardless of culture conditions, there existed a large population of cells that resembled oligodendrocytes morphologically and coexpressed the oligodendrocyte-specific marker galactocerebroside (GalC) and the acetylcholine (ACh)-synthesizing enzyme choline acetyltransferase (ChAT). These cells did not express neuronal markers, and whole cell recordings from cells with similar morphology displayed only outward currents in response to depolarizing voltage steps, further supporting their oligodendroglial identity. Another cholinergic marker, the vesicular ACh transporter, was also detected in GalC(+) oligodendrocytes. Furthermore, neurospheres cultured in the presence of the cholinergic receptor antagonist atropine showed a decrease in the number of GalC(+) spheres, implicating the muscarinic ACh receptor in oligodendrocyte development. The actions of neurotrophins and ciliary neurotrophic factor (CNTF) on these ChAT(+) oligodendrocytes were examined. Among these, CNTF treatment significantly increased oligodendrocytic process outgrowth. These results demonstrate classical cholinergic neuronal markers in oligodendrocytes as well as an effect of muscarinic receptor blockade on oligodendrocyte differentiation. PMID- 12111856 TI - Deletion of the C-terminal domain of the NR2B subunit alters channel properties and synaptic targeting of N-methyl-D-aspartate receptors in nascent neocortical synapses. AB - Channel properties and synaptic targeting of N-methyl-D-aspartate (NMDA) receptors determine their importance in synaptic transmission, long-term synaptic plasticity, and developmental reorganization of synaptic circuits. To investigate the involvement of the C-terminal domain of the NR2B subunit in regulating channel properties and synaptic localization, we analyzed gene-targeted mice expressing C-terminally truncated NR2B subunits (NR2B(DeltaC/DeltaC) mice; Sprengel et al. [1998] Cell 92:279-89). Because homozygous NR2B(DeltaC/DeltaC) mice die perinatally, we studied embryonic neocortical neurons differentiating in culture. At early stages in vitro, neurons predominantly expressed NR1/NR2B receptors, as shown by the NR2B subunit-specific antagonist ifenprodil. At these nascent synapses, NMDA excitatory postsynaptic currents (EPSCs) in neurons from NR2B(DeltaC/DeltaC) mice showed a strong-amplitude reduction to 20% of control, but AMPA EPSCs were unaltered. Analysis of the MK-801 block of NMDA receptor mediated whole-cell currents revealed a decreased peak open probability of NMDA receptor channels (to about 60%) in neurons from NR2B(DeltaC/DeltaC) mice, although their single channel conductance was unchanged. To study effects on synaptic targeting, we determined the fraction of synaptically localized NMDA receptors relative to the whole-cell NMDA receptor population. In neurons from NR2B(DeltaC/DeltaC) mice, the synaptic NMDA receptor fraction was drastically reduced, suggesting that the C-terminal domain of the NR2B subunit plays a major role in synaptic targeting of NMDA receptors at nascent synapses. With increasing time in culture, the reduction in NMDA EPSCs in neurons from NR2B(DeltaC/DeltaC) mice diminished. This is explained by the expression of additional NMDA receptor subtypes containing NR2A subunits at more mature synapses. PMID- 12111857 TI - Long-term fate of human telencephalic progenitor cells grafted into the adult mouse brain: effects of previous amplification in vitro. AB - We assessed the developmental potential of human telencephalic progenitor cells, with and without previous amplification in vitro, following grafting into the nonlesioned adult mouse CNS. Cell suspensions, shown to contain neuroepithelium like and neuroblast-like cells, were injected into the subventricular zone (SVZ) and the striatum. These two regions were selected for comparative studies because one, the SVZ, is mitotically active, whereas the other, the striatum, is mitotically inactive. In situ hybridization with a human-specific Alu probe showed that the cells survived for up to 30 weeks in both targets and migrated away from the injection site. Fresh cells continued to proliferate and gave rise to very extended grafts before differentiating into neurons and glia. We further show that, when grown in vitro prior to grafting, human cells acquired new properties: Their proliferation was very limited, and they differentiated more rapidly. This study therefore provides new information about the use of these cells, which are a potential tool for both cellular and gene therapy. PMID- 12111858 TI - Temporal analysis of cytokine gene expression during infiltration of porcine neuronal grafts implanted into the rat brain. AB - A large array of evidence supports the involvement of infiltrating T lymphocytes in the rejection process of intracerebral neuronal xenografts. Little is known, however, about the molecular mechanisms that drive the recruitment of this cell type. In the present work, we used real-time RT-PCR methodology to investigate the kinetics of cytokine gene expression during the infiltration of fetal porcine neurons (PNEU) implanted into the striatum of LEW.1A rats. T lymphocyte infiltration was followed by measuring the intracerebral levels of transcripts encoding the beta chain of the T cell receptor. These transcripts remained barely detectable until the fourth week (28 days) postimplantation, when a sudden accumulation occurred. Their kinetics, which support previous immunohistochemical observations, indicate that alphabetaT lymphocyte recruitment occurs rapidly after a delay of several weeks in this experimental model. Infiltration of PNEU grafts by T lymphocytes was accompanied by a concomitant, dramatic augmentation of transcripts coding for monocyte chemotactic protein-1 and RANTES (for regulated on activation, normal T cell expressed and secreted), two chemokines targeting this cell type, among others. Likewise, a sudden accumulation of transcripts of proinflammatory lymphokines [interleukin (IL)-1alpha, tumor necrosis factor-alpha, IL-6] as well as Th1 cytokines (IL-2, interferon-gamma) was also detected. In contrast, IL-4, -10, and -13 mRNA remained barely detectable at the different time points. No significant changes were noticed for IL-12 or transforming growth factor-beta transcripts. These data support the concept that T lymphocyte infiltration of PNEU grafts is actively promoted by a local production of chemokines and proinflammatory lymphokines and is based on a Th1 polarization. PMID- 12111859 TI - Regeneration of primary sensory axons into the adult rat spinal cord via a peripheral nerve graft bridging the lumbar dorsal roots to the dorsal column. AB - This study investigated the feasibility of using a peripheral nerve autograft (NAG) to promote and guide regeneration of sensory axons from the caudal lumbar dorsal roots to the rostral dorsal column following a lower thoracic cordotomy in adult rats. After a left hemicordotomy at the T13 vertebra level and ipsilateral L3 and L4 rhizotomies, a peripheral NAG (peroneal nerve) was connected to the distal roots stumps, then implanted into the left dorsal column 10 mm rostral to hemicordotomy site (n = 12). After surgery, all animals of the experimental group experienced complete anesthesia in their left hindlimb. Three months later, a slight response to nociceptive stimulation reappeared in L3 and/or L4 dermatomes in 6 of the 12 experimental animals. None of these animals exhibited self mutilation. Nine months after surgery, we performed retrograde tracing studies by injecting horseradish peroxidase (HRP) into the left dorsal column 30 mm rostral to the NAG implantation site. In eight animals, we found HRP-stained neurons in the left L3 and/or L4 dorsal root ganglia (DRG). The mean number of HRP-stained neurons per DRG was 71 +/- 92 (range 2-259). In control groups, no HRP-stained neurons were found in L3 or L4 DRG. Histological analysis of the NAG showed evidence of axonal regeneration in all 8 animals with positive retrograde labeling of DRG neurons. However, we did not find a statistical correlation between the number of HRP-stained neurons and the degree of sensory recovery. This study demonstrates that an NAG joining dorsal roots to the dorsal column, thus shunting the original CNS-PNS junction, can support regeneration of central axons from DRG primary sensory neurons into the dorsal column over distances of at least 30 mm despite the inhibitory influence of the CNS white matter. PMID- 12111860 TI - Re-evaluation of phencyclidine low-affinity or "non-NMDA" binding sites. AB - TCP and its derivative gacyclidine (+/- GK11) are high-affinity non-competitive antagonists of N-methyl-D-aspartate (NMDA) receptors (NMDARs) and as such exhibit significant neuroprotective properties. These compounds also bind with a low affinity to binding sites whose pharmacological profiles are different from that of NMDARs. With the intention to develop new strategies of neuroprotection, we found it mandatory to investigate whether 1-[1-(2-thienyl)cyclohexyl]piperidine (TCP) and gacyclidine low-affinity sites are similar. The effects of several drugs selective for either NMDARs or the [(3)H]TCP low-affinity site (or PCP(3) site) on (+), (-)[(3)H]GK11 and [(3)H]TCP specific binding were investigated. Competition experiments on cerebellum homogenates revealed substantial differences between the pharmacological profiles of the PCP(3) site and that of gacyclidine's enantiomers low-affinity sites. Under experimental conditions preventing the interaction of the radioligands with NMDARs, the autoradiographic study showed, however, that the distributions of both [(3)H]TCP and (-)[(3)H]GK11 specific binding were similar. The specific labelling was low and uniform in telencephalic structures, whereas in the cerebellum it was higher in the molecular than in the granular layer. Finally, the analysis of competition experiments performed on tissues slices demonstrated that PCP(3) selective ligands were unable to prevent [(3)H]TCP or (-)[(3)H]GK11 binding to "non-NMDA" binding sites. As a whole, our data suggest that: (1) the different pharmacological profiles of [(3)H]TCP and [(3)H]gacyclidine enantiomers on low affinity sites are due to their selectivity for specific NMDARs subpopulations; (2) the pharmacological isolation of TCP and gacyclidine "non-NMDA" binding sites is the most appropriate way to further study the low-affinity component of their specific binding. Obtaining reliable and specific pharmacological tools for those binding sites is of particular interest, since it is likely that they play a substantial role in the low neurotoxicity, and therefore tolerability, of gacyclidine, a new neuroprotective drug currently evaluated in clinical trials for the treatment of brain and spinal cord injuries. PMID- 12111861 TI - Cytokine activity contributes to induction of inflammatory cytokine mRNAs in spinal cord following contusion. AB - Injury of the spinal cord leads to an inflammatory tissue response, probably mediated in part by cytokines. Because a common therapy for acute spinal cord injury is the use of an antiinflammatory synthetic glucocorticoid (methylprednisolone), we sought to determine mechanisms contributing to inflammation shortly after acute injury. Cytokine mRNAs [interleukin (IL)-1alpha, IL-1beta, tumor necrosis factor (TNF)-alpha, and IL-6] were increased during the first 2 hr following weight-drop compression injury by RNase protection assay, prior to the reported appearance of circulating lymphocytes. This immediate pattern of cytokine mRNA induction could be replicated in cultured, explanted spinal cord slices but not in whole blood of injured animals, which is consistent with a tissue source of cytokine mRNAs. Western blotting detected IL-1beta-like immunoreactivity released into culture medium following explantation and pro-IL 1beta-like immunoreactivity in freshly dissected spinal cord tissue. Pharmacologically blocking IL-1 and TNF-alpha receptors significantly reduced expression of IL-1alpha, IL-1beta, and TNF-alpha mRNAs. Finally, mice lacking both IL-1 and TNF-alpha receptors exhibited diminished induction of TNF-alpha, IL 6, and IL-1ra mRNAs following injury. Therefore, we conclude that contusion injury induces an immediate release of cytokines, which then contributes to the induction of cytokine mRNAs. PMID- 12111862 TI - Ceramide-induced cell death in primary neuronal cultures: upregulation of ceramide levels during neuronal apoptosis. AB - Ceramide is a sphingolipid that has been implicated both in apoptosis and protection from cell death. We show that in both rat cerebellar granule cells and cortical neuronal cultures application of C(2)-ceramide causes cell death in a dose- and time-dependent manner. Similar effects were observed with the exogenous application of bacterial sphingomyelinase, which hydrolyzes sphingomyelin located on the outer leaflet of the plasma membrane and leads to endogenous ceramide accumulation. Furthermore, endogenous ceramide levels were increased during apoptosis induced by nutrient deprivation or etoposide treatment. These findings suggest that upregulation of ceramide levels, which may be generated through activation of sphingomyelinase, contributes to neuronal apoptosis. PMID- 12111863 TI - Defective neuronal sprouting by human apolipoprotein E4 is a gain-of-negative function. AB - The apolipoprotein E (apoE) epsilon 4 allele (apoE4) is a major risk factor for neurodegenerative conditions, including Alzheimer's disease (AD). A role for apoE in regeneration of synaptic circuitry after neural injury has been shown in several in vitro studies in which apoE3 supports neuronal sprouting better than apoE4. We evaluated sprouting in an in vitro mouse organotypic hippocampal slice culture system derived from transgenic mice expressing apoE3 or apoE4, in which apoE-dependent granule cell mossy fiber sprouting in the presence of apoE4 is only 51% of the level of apoE3. Sprouting supported by apoE4 had a dose response opposite that by supported by apoE3: although increasing E3 expression increased sprouting, increasing E4 expression decreased sprouting, suggesting that the defect in E4 in supporting neuronal sprouting is a gain-of-negative activity. These results may have important pharmacogenomic implications for AD therapies that modulate apoE expression levels. PMID- 12111865 TI - Down-regulation of gamma-glutamylcysteine synthetase regulatory subunit gene expression in rat brain tissue during aging. AB - The mechanism underlying age-related neurodegenerative diseases is still an area of significant controversy. Increased evidence suggests that oxidative stress contributes importantly to neuronal damage observed in the brains of aged animals and in neurodegenerative diseases. Glutathione (GSH), the most abundant intracellular nonprotein thiol, plays an important role in antioxidant defense. The concentration of this important antioxidant decreases with age in the brain, which is accompanied by an increase in oxidative damage to macromolecules. The mechanism underlying the age-associated decline in GSH content in the brain, however, is not clear. In this study, we demonstrate for the first time that the expression of the regulatory subunit of gamma-glutamylcysteine synthetase (GCS), the rate-limiting enzyme in de novo GSH synthesis, decreases with age in cerebellum, cerebral cortex, and hippocampus of Fisher 344 rats. This was accompanied by a decline in GCS activity and GSH content. There were no significant differences in either the concentrations of cysteine and glutathione disulfide (GSSG) or the activities of glutathione synthetase (GS), gamma-glutamyl traspeptidase (GGT), and glutathione reductase (GR) in the brains from different age groups. Our results suggest that the age-associated decrease in GSH in the brain may result from the down-regulation of GCS regulatory subunit and consequently a decrease in the activity of GCS. PMID- 12111864 TI - Inhibition of COX-2 reduces the age-dependent increase of hippocampal inflammatory markers, corticosterone secretion, and behavioral impairments in the rat. AB - Brain aging as well as brain degenerative processes with accompanying cognitive impairments are generally associated with hyperactivity of the hypothalamus pituitary-adrenal axis, the end product of which, the glucocorticoid hormone, has been warranted the role of cell damage primum movens ("cascade hypothesis"). However, chronic inflammatory activity occurs in the hippocampus of aged rats as well as in the brain of Alzheimer's disease patients. The concomitant increase in the secretion of the glucocorticoid hormone, the endogenous anti-inflammatory and pro-inflammatory markers, has prompted us to investigate the two phenomena in the aging rat, and to work out its meaning. This study shows that: (I) interleukin 1beta (IL-1beta), tumor necrosis factor alpha (TNFalpha), and prostaglandin E(2) (PGE(2)) increase with age in the rats hippocampus, and (II) chronic oral treatment with celecoxib, a selective cycloxygenase-2 (COX-2) inhibitor, is able to contrast the age-dependent increase in hippocampal levels of pro-inflammatory markers and circulating anti-inflammatory corticosterone, provided that it is started at an early stage of aging. Under these conditions, age-related impairments in cognitive ability may be ameliorated. Taken together, these results indicate that there is a natural tendency to offset the age-dependent increase in brain inflammatory processes via the homeostatic increase of the circulating glucocorticoid hormone. PMID- 12111866 TI - Hypothermia and thiopentone sodium: individual and combined neuroprotective effects on cortical cultures exposed to prolonged hypoxic episodes. AB - Because there are many conflicting reports on cerebroprotective effects of hypothermia and barbiturates, we examined the degree of neuroprotection at defined temperatures (normothermia, 37 degrees C; mild hypothermia, 32 degrees C; deep hypothermia, 22 degrees C; and profound hypothermia, 17 degrees C) and various concentrations (low, 4 microM; moderate, 40 microM; and high, 400 & microM) of thiopentone sodium (TPS), alone and in combination in cortical cultures exposed to prolonged hypoxia (24-48 hr). The survival rate of embryonic day (E)16 Wistar rat cortical neurons was evaluated on photomicrographs before and after experiments. During the 24-hr hypoxic period, the survival rate of neurons was maximal with combinations of mild hypothermia with 40 microM (91.6 +/ 0.7%) and 400 microM TPS (90.8 +/- 0.7%) or deep hypothermia combined with all concentrations of TPS (4 microM, 90.6 +/- 1.0%; 40 microM, 91.4 +/- 0.8%; 400 microM, 91.8 +/- 1.2%). During 48 hr hypoxia, the highest survival rate was seen with the combination of deep hypothermia and either 40 microM (90.9 +/- 0.6%) or 400 microM (91.1 +/- 1.4%) TPS. In the presence of profound hypothermia in combination with all concentrations of TPS, the survival rate was significantly reduced (P< 0.01) compared to combined application of either mild or deep hypothermia with TPS. In summary, maximal neuroprotection was attained with hypothermia and TPS in combination rather than applied individually, during prolonged hypoxic episodes (24- 48 hr). During a 24-hr hypoxic period, both mild and deep hypothermia combined with a clinically relevant concentration of TPS (40 microM) offered the highest neuroprotection. Only deep hypothermia provided maximal neuroprotection when combined with 40 microM TPS, during 48-hr hypoxia. Combination of profound hypothermia and TPS did not confer considerable neuroprotection during long lasting hypoxia. PMID- 12111867 TI - Impairment of adenylyl cyclase and of spatial memory function after microsphere embolism in rats. AB - The purpose of the present study was to characterize alterations in the adenylyl cyclase (AC), cyclic adenosine 3',5'-monophosphate (cAMP), and spatial memory function after sustained cerebral ischemia. Sustained cerebral ischemia was induced by injection of 900 microspheres (48 microm in diameter) into the right (ipsilateral) hemisphere of rats. Alterations in the AC and cAMP in the cerebral cortex and hippocampus were examined up to 7 days after the embolism. A decrease in the cAMP content was seen in the ipsilateral hemisphere throughout the experiment. Microsphere embolism (ME) decreased the activity of Ca(2+)/calmodulin (CaM)-sensitive AC in the ipsilateral hemisphere throughout the experiment, whereas the basal and 5'-guanylyl imidodiphosphate (Gpp(NH)p)-sensitive AC activities were not altered. Immunoblotting analysis of AC subtypes with specific antibodies showed a decrease in the immunoreactivity of AC-I in the ipsilateral hemisphere during these periods. No significant differences in the immunoreactivity of AC-V/VI and AC-VIII were observed after ME. The levels of GTP binding proteins Galpha(s), Galpha(i), and Gbetawere unchanged. Furthermore, microsphere-embolized rats showed prolongation of the escape latency in the water maze task determined on the seventh to ninth day after the operation. These results suggest that sustained cerebral ischemia may induce the impairment of the AC, particularly a selective reduction in the AC-I level and activity, coupled with the decrease in cAMP content. This reduction may play an appreciable role in the disturbance in cAMP-mediated signal transduction system, possibly leading to learning and memory dysfunction. PMID- 12111869 TI - Role of Bcl-2 family of proteins in malignancy. AB - B cell lymphoma gene-2 (Bcl-2) is the prototypic member of a growing family of proteins that play evolutionarily conserved, key regulatory roles in apoptosis. The Bcl-2 family members are characterized by the presence of one or more Bcl-2 homology domains and are comprised of both the prosurvival and proapoptotic proteins. Bcl-2 itself is a prosurvival member of the family and its aberrant expression has been linked to a variety of different cancers, including several hematological malignancies. Although the exact mechanism of action of Bcl-2 family of proteins in regulating apoptosis is still a matter of some debate, these proteins appear to act upstream of caspase activation. Many recent studies have shown the therapeutic potential of targeting Bcl-2 family members for the treatment of cancer. This article summarizes what is currently known about Bcl-2 like proteins and how the evolving understanding of the biology of these proteins is paving way for the development of novel cancer therapeutics. PMID- 12111868 TI - Tuberculosis in blood and marrow transplant recipients. AB - Although one third of the world's population is infected with tuberculosis (TB), TB in blood and marrow transplant (BMT) recipients is relatively less well studied, as the incidence of TB is relatively low in developed countries with BMT units. Since the report of the first two cases in 1983, 52 cases of TB complicating BMT have been reported in the English literature from BMT centers in ten different countries. Not unexpectedly, the two largest series were reported from areas with a high incidence of TB in the general population, with about 45 cases per 10(5) inhabitants per year in Spain and about 100 cases per 10(5) inhabitants per year in Hong Kong respectively. The overall frequency of occurrence of TB in BMT recipients was 0.4% (52 cases among 13 881 BMT recipients), with a male:female ratio of 11:9 and median age of 33 (range 7-57). The incidence of TB in the general population is a major predictor of a higher frequency of occurrence in BMT recipients. Moreover, allogeneic transplantation, graft-versus-host disease, and total body irradiation were found to be risk factors associated with TB. Among the 48 cases in whom the time of manifestation were reported, only one case manifested during the neutropenic period (day 11). On the other hand, 11 cases (23%) manifest after engraftment but before day 100, and 36 (75%) manifest after day 100. The most important aspect towards making the diagnosis is a high index of suspicion, as TB occurred in relatively low frequencies especially in developed countries, and the clinical patterns usually mimic other more common infectious and non-infectious complications after BMT. As the incidence of drug resistant TB is increasing, we prefer to treat our patients for at least one year (as compared with six months in immunocompetent hosts) with four drugs in the first six months and two or three drugs for another six months. In those patients who could not tolerate oral medication, we used an intravenous regimen of rifampicin, ciprofloxacin, and amikacin until oral therapy could be instituted. The absence of relapse after termination of treatment in our patients suggested that secondary prophylaxis would not be necessary as long as immune function has been restored. With the rising incidence of TB in countries that previously enjoyed a very low prevalence of TB, attributed to the growing population of HIV-infected subjects with TB, and the changing patterns of population migration, it is important to bear a high index of suspicion of Mycobacterium tuberculosis as a pathogen in BMT recipients. PMID- 12111872 TI - Recent publications in hematological oncology. PMID- 12111871 TI - Cd8(+)/V beta 5.1(+) large granular lymphocyte leukemia associated with autoimmune cytopenias, rheumatoid arthritis and vascular mammary skin lesions: successful response to 2-deoxycoformycin. AB - We report a case of CD8(+)/V beta 5.1(+) T-cell large granular lymphocyte leukemia (T-LGL leukemia) presenting with mild lymphocytosis, severe autoimmune neutropenia, thrombocytopenia, polyarthritis and recurrent infections with a chronic disease course. Immunophenotyping showed an expansion of CD3(+)/TCR alpha beta(+)/CD8(+bright)/CD11c(+)/CD57(-)/CD56(-) large granular lymphocytes with expression of the TCR-V beta 5.1 family. Southern blot analysis revealed a clonal rearrangement of the TCR beta-chain gene. Hematopoietic growth factors, high dose intravenous immunoglobulin and corticosteroids were of limited therapeutic benefit to correct the cytopenias. During the disease course, the patient developed a severe cutaneous leg ulcer and bilateral vascular mammary skin lesions. Treatment with 2-deoxycoformycin resulted in both clinical and hematological complete responses, including the resolution of vascular skin lesions. Combined immuno-staining with relevant T-cell associated and anti-TCR-V beta monoclonal antibodies proved to be a sensitive method to assess the therapeutic effect of 2-deoxycoformicin and to evaluate the residual disease. PMID- 12111870 TI - Isotype class switching and the pathogenesis of multiple myeloma. AB - Translocations at the immunoglobulin heavy chain locus (14q32) are now considered the commonest karyotypic change in multiple myeloma. These translocations are thought to be intimately involved in the pathogenesis of this disease. The heavy chain locus is strongly transcriptionally active in B and plasma cells and transfer of a potential oncogene to 14q32 would result in its dysregulation. Molecular characterization suggests that the majority of these breakpoints cluster in switch regions within the heavy chain locus. Switch regions are normally involved in the regulated process of isotype switching so that in myeloma the rearrangements are believed to be a result of so-called illegitimate (aberrant) switch recombination and are likely to be an early event in myeloma development. A legitimate switch recombination event occurs between two switch regions producing a hybrid switch; this is necessary for class switching to proceed on a productive allele. In this review we describe the process of isotype switching and how illegitimate class switching may be related to the pathogenesis of multiple myeloma. PMID- 12111873 TI - Baseline information in structural failure time estimators for the effect of observed treatment compliance. AB - Structural accelerated failure time models allow expression of the effect of treatment actually received in placebo-controlled randomized trials with non compliance. Without further assumptions, the structural parameter is typically estimated via a series of auxiliary logrank tests, searching for the structural parameter that back-transforms treated survival times to latent treatment-free survival times which are equally distributed between randomized arms. In this paper we investigate to what extent score tests involving baseline covariates provide more powerful auxiliary tests and lead to more precise estimates of the structural parameter without compromising the alpha-level. We propose a set of estimating equations which combines score components for covariate effects based on the control arm only, with a log-likelihood score for treatment effect based on both arms. Analytic results for exponential models as well as simulation studies for the semi-parametric approach indicate that in many practical situations this incorporation of baseline covariates leads to more precise estimators of the structural effect. Relative efficiency is shown to depend on the selective nature of compliance. In a leukaemia trial we find the length of the 95 per cent confidence interval for the structural parameter is reduced to two-thirds of the original length by incorporating baseline covariates in this way. PMID- 12111874 TI - A Bayesian analysis of a proportion under non-ignorable non-response. AB - The National Health Interview Survey (NHIS) is one of the surveys used to assess one aspect of the health status of the U.S. population. One indicator of the nation's health is the total number of doctor visits made by the household members in the past year. We study the binary variable of at least one doctor visit versus no doctor visit by all household members to each of the 50 states and the District of Columbia. The proportion of households with at least one doctor visit is an indicator of the status of health of the U.S. population. There is a substantial number of non-respondents among the sampled households. The main issue we address here is that the non-response mechanism should not be ignored because respondents and non-respondents differ. The purpose of this work is to estimate the proportion of households with at least one doctor visit, and to investigate what adjustment needs to be made for non-ignorable non-response. We consider a non-ignorable non-response model that expresses uncertainty about ignorability through the ratio of odds of a household doctor visit among respondents to the odds of doctor visit among all households, and this ratio varies from state to state. We use a hierarchical Bayesian selection model to accommodate this non-response mechanism. Because of the weak identifiability of the parameters, it is necessary to 'borrow strength' across states as in small area estimation. We also perform a simulation study to compare the expansion model with an alternative expansion model, an ignorable model and a non-ignorable model. Inference for the probability of a doctor visit is generally similar across the models. Our main result is that for some of the states the non response mechanism can be considered non-ignorable, and that 95 per cent credible intervals of the probability for a household doctor visit and the probability that a household responds shed important light on the NHIS data. PMID- 12111875 TI - A new parametric method based on S-distributions for computing receiver operating characteristic curves for continuous diagnostic tests. AB - Receiver operating characteristic (ROC) curves provides a method for evaluating the performance of a diagnostic test. These curves represent the true positive ratio, that is, the true positives among those affected by the disease, as a function of the false positive ratio, that is, the false positives among the healthy, corresponding to each possible value of the diagnostic variable. When the diagnostic variable is continuous, the corresponding ROC curve is also continuous. However, estimation of such curve through the analysis of sample data yields a step-line, unless some assumption is made on the underlying distribution of the considered variable. Since the actual distribution of the diagnostic test is seldom known, it is difficult to select an appropriate distribution for practical use. Data transformation may offer a solution but also may introduce a distortion on the evaluation of the diagnostic test. In this paper we show that the distribution family known as the S-distribution can be used to solve this problem. The S-distribution is defined as a differential equation in which the dependent variable is the cumulative. This special form provides a highly flexible family of distributions that can be used as models for unknown distributions. It has been shown that classical statistical distributions can be represented accurately as S-distributions and that they occur in a definite subspace of the parameter space corresponding to the whole S-distribution family. Consequently, many other distributional forms that do not correspond to known distributions are provided by the S-distribution. This property can be used to model observed data for unknown distributions and is very useful in constructing parametric ROC curves in those cases. After fitting an S-distribution to the observed samples of diseased and healthy populations, ROC curve computation is straightforward. A ROC curve can be considered as the solution of a differential equation in which the dependent variable is the ratio of true positives and the independent variable is the ratio of false positives. This equation can be easily obtained from the S-distributions fitted to observed data. Using these results, we can compute pointwise confidence bands for the ROC curve and the corresponding area under the curve. We shall compare this approach with the empirical and the binormal methods for estimating a ROC curve to show that the S-distribution based method is a useful parametric procedure. PMID- 12111876 TI - Properties of the summary receiver operating characteristic (SROC) curve for diagnostic test data. AB - The summary receiver operating characteristic (SROC) curve has been recommended to represent the performance of a diagnostic test, based on data from a meta analysis. However, little is known about the basic properties of the SROC curve or its estimate. In this paper, the position of the SROC curve is characterized in terms of the overall diagnostic odds ratio and the magnitude of inter-study heterogeneity in the odds ratio. The area under the curve (AUC) and an index Q(*) are discussed as potentially useful summaries of the curve. It is shown that AUC is maximized when the study odds ratios are homogeneous, and that it is quite robust to heterogeneity. An upper bound is derived for AUC based on an exact analytic expression for the homogeneous situation, and a lower bound based on the limit case Q(*), defined by the point where sensitivity equals specificity: Q(*) is invariant to heterogeneity. The standard error of AUC is derived for homogeneous studies, and shown to be a reasonable approximation with heterogeneous studies. The expressions for AUC and its standard error are easily computed in the homogeneous case, and avoid the need for numerical integration in the more general case. SE(AUC) and SE(Q(*)) are found to be numerically close, with SE(Q(*)) being larger if the odds ratio is very large. The methods are illustrated using data for the Pap smear screening test for cervical cancer, and for three tests for the diagnosis of metastases in cervical cancer patients. PMID- 12111877 TI - Estimating cancer prevalence using mixture models for cancer survival. AB - Knowledge of cancer prevalence is useful for estimating the ongoing level of resources utilized in the treatment of disease and is of some public health interest. Cancer prevalence is estimated first as the proportion of persons previously diagnosed (PD) with cancer that are still alive; and second as the proportion of individuals in the population who were previously diagnosed with cancer and who have not been cured (NC). The proportion of cases that are cured is estimated by assuming that the cured and uncured cases have distinct survival patterns. The hazard for cured cases is assumed to be a multiple of the hazard from causes other than cancer in the general population. The hazard for uncured cases is assumed to have two independent components: one corresponding to the disease-specific hazard, and the other a multiple of the population hazard from 'other causes'. Future prevalence estimates are obtained by projecting the survival of current prevalent cases as well as the survival of future incident cases. PMID- 12111878 TI - Jointly modelling the relationship between survival and pulmonary function in cystic fibrosis patients. AB - Modelling the relationship between pulmonary function and survival in cystic fibrosis (CF) is complicated by the fact that measures of pulmonary function commonly used such as the forced expiratory volume in one second (FEV(1)) are measured with error, and patients with the poorest lung function are increasingly censored by death, that is, data are available only for the patients who have survived to the current age. We assume a linear random effects model for FEV1 per cent predicted, where the random intercept and slope of FEV(1) per cent predicted, along with a specified transformation of the age at death follow a trivariate normal distribution. We illustrate how this model can be used to describe the relationship between age at death and parameters of the individual patient's regression of FEV(1) per cent predicted versus age, such as the slope and the intercept or true value of FEV(1) per cent predicted at a given age. We also illustrate how the model provides empirical Bayes estimates of these individual parameters. In particular, we explore how the predicted value of the age at death might be used as a prognostic or severity index. The model and methods are illustrated on a cohort of 188 cystic fibrosis patients with a common genotype (homozygous for the DeltaF508 mutation), born on or after 1965 and followed at the CF Center at the Rainbow Babies and Children's Hospital, Cleveland, OH, U.S.A. PMID- 12111879 TI - Methods for evaluating the performance of diagnostic tests in the absence of a gold standard: a latent class model approach. AB - In many areas of medical research, 'gold standard' diagnostic tests do not exist and so evaluating the performance of standardized diagnostic criteria or algorithms is problematic. In this paper we propose an approach to evaluating the operating characteristics of diagnoses using a latent class model. By defining 'true disease' as our latent variable, we are able to estimate sensitivity, specificity and negative and positive predictive values of the diagnostic test. These methods are applied to diagnostic criteria for depression using Baltimore's Epidemiologic Catchment Area Study Wave 3 data. PMID- 12111880 TI - A new approach to training back-propagation artificial neural networks: empirical evaluation on ten data sets from clinical studies. AB - We present a new approach to training back-propagation artificial neural nets (BP ANN) based on regularization and cross-validation and on initialization by a logistic regression (LR) model. The new approach is expected to produce a BP-ANN predictor at least as good as the LR-based one. We have applied the approach to ten data sets of biomedical interest and systematically compared BP-ANN and LR. In all data sets, taking deviance as criterion, the BP-ANN predictor outperforms the LR predictor used in the initialization, and in six cases the improvement is statistically significant. The other evaluation criteria used (C-index, MSE and error rate) yield variable results, but, on the whole, confirm that, in practical situations of clinical interest, proper training may significantly improve the predictive performance of a BP-ANN. PMID- 12111881 TI - Sample size requirements for estimating intraclass correlations with desired precision. AB - A method is developed to calculate the approximate number of subjects required to obtain an exact confidence interval of desired width for certain types of intraclass correlations in one-way and two-way ANOVA models. The sample size approximation is shown to be very accurate. PMID- 12111882 TI - Designing complex group sequential survival trials. AB - This paper presents methodology for designing complex group sequential survival trials when the survival curves will be compared using the logrank statistic. The method can be applied to any treatment and control survival curves as long as each hazard function can be approximated by a piecewise linear function. The approach allows arbitrary accrual patterns and permits adjustment for varying rates of non-compliance, drop-in and loss to follow-up. The calendar-time information-time transformation is derived under these complex assumptions. This permits the exploration of the operating characteristics of various interim analysis plans, including sample size and power. By using the calendar-time information-time transformation, information fractions corresponding to desired calendar times can be determined. In this way, the interim analyses can be scheduled in information time, assuring the desired power and realization of the spending function, while the interim analyses will take place according to the desired calendar schedule. PMID- 12111883 TI - Bayesian two-stage designs for phase II clinical trials. AB - Many different statistical designs have been used in phase II clinical trials. The majority of these are based on frequentist statistical approaches. Bayesian methods provide a good alternative to frequentist approaches as they allow for the incorporation of relevant prior information and the presentation of the trial results in a manner which, some feel, is more intuitive and helpful. In this paper, we propose two new Bayesian designs for phase II clinical trials. These designs have been developed specifically to make them as user friendly and as familiar as possible to those who have had experience working with two-stage frequentist phase II designs. Thus, unlike many of the Bayesian designs already proposed in the literature, our designs do not require a distribution for the response rate of the currently used drug or the explicit specification of utility or loss functions. We study the properties of our designs and compare them with the Simon two-stage optimal and minimax designs. We also apply them to an example of two recently concluded phase II trials conducted at the National Cancer Centre in Singapore. Sample size tables for the designs are given. PMID- 12111884 TI - Meta-analysis of method comparison studies. AB - Methods for the meta-analysis of results from randomized controlled trials are well established. However, there are currently no methods for the meta-analysis of method comparison studies. Here the combination of results from studies comparing two methods of measurement on the same unit of observation is required. We compare standard methods for the pooling of k samples from the same Normal population to those for pooling parameter estimates, in order to estimate the pooled mean difference and 95 per cent limits of agreement. Methods for investigating heterogeneity across studies and for calculating random effects estimates are proposed. We postulate that for published studies either the estimated mean or variance of the difference between measurements will tend to be smaller than for unpublished studies and investigate the evidence for the existence of such publication bias. The methods are illustrated with an example evaluating the accuracy of temperature measured at the axilla compared to the rectum in children. PMID- 12111885 TI - On the assessment of adverse drug reactions from spontaneous reporting systems: the influence of under-reporting on odds ratios. AB - A well-known problem in spontaneous reporting systems (SRSs) for adverse drug reactions (ADRs) is under-reporting, that is, the problem that not all occurrences of ADRs are reported to the SRS. We look at the question of how to draw statistical conclusions from analyses of SRS data using reporting odds ratios. We will show that certain under-reporting problems play no role in assessing ADRs from SRSs: the results from the analyses turn out to be biased by some specific under-reporting problems, but not by others. SRS data can be particularly useful for the assessment of drug-drug interactions. If the assumption holds that there is an under-reporting problem for a first drug, and an under-reporting problem for a second drug, but that these two under-reporting problems do not influence each other, then reporting odds ratios estimated from SRSs are useful for signalling drug-drug interactions in the ADR-experiencing population. Similar results hold for covariate-drug interactions. We illustrate our results using two examples. PMID- 12111886 TI - Comparison of bandwidth selection methods for kernel smoothing of ROC curves. AB - In this paper we compared four non-parametric kernel smoothing methods for estimating an ROC curve based on a continuous-scale test. All four methods produced a smooth ROC curve of the test. The difference in these four methods lay with the way they chose their bandwidth parameters. To assess the relative performance of the four bandwidth selection methods, we conducted a simulation study using different underlying distributions, along with varied sample sizes. The results from our simulation study suggested that the kernel smoothing method originally proposed by Altman and Leger for estimation of the distribution function was the best choice for estimation of an ROC curve. We illustrated these methods with a real example. PMID- 12111887 TI - A Bayesian space varying parameter model applied to estimating fertility schedules. AB - We propose a spatial generalized linear model (GLM) to analyse the vital rates for small areas. In each small area, we have a response vector and covariates to explain its variability. The statistical methodology is based on a spatial Bayesian approach and it allows the covariates' parameters of the generalized linear model to vary smoothly on space. Hence, the effect of a covariate on the response varies depending on the random variables measurement location. Our model is an extension of disease mapping models allowing the space-covariate interaction to be modelled in a natural way and giving space a position of intrinsic interest. We introduce the model in the context of fertility curve estimation. In each small area, we have a curve describing the variation of fertility rates by age modelled by Coale's fertility model, which implies a GLM in each area. A simulation shows the advantages of our approach. In addition, the paper applies the procedure to census data used to study the diffusion of low fertility behaviour in Brazil. PMID- 12111888 TI - Time-varying exposure and the impact of stressful life events on onset of affective disorder. AB - Stressful life events are now established as risk factors for the onset of affective disorder but few studies have investigated time-varying exposure effects. Discrete (grouped) time survival methods provide a flexible framework for evaluating multiple time-dependent covariates and time-varying covariate effects. Here, we use these methods to investigate the time-varying influence of life events on the onset of affective disorder. Various straightforward time varying exposure models are compared, involving one or more (stepped) time dependent covariates and time-dependent covariates constructed or estimated according to exponential decay. These models are applied to data from two quite different studies. The first, a small scale interviewer-based longitudinal study (n = 180) concerned with affective disorder onset following loss (or threat of loss) event experiences. The second, a questionnaire assessment as part of an ongoing population study (n = 3353), provides a history of marital loss events and of depressive disorder onset. From the first study the initial impact of loss events was found to decay with a half-life of 5 weeks. Psychological coping strategy was found to modify vulnerability to the adverse effects of these events. The second study revealed that while men had a lower immediate risk of disorder onset following loss event experience their risk period was greater than for women. Time-varying exposure effects were well described by the appropriate use of simple time-dependent covariates. PMID- 12111889 TI - Modelling HIV viral rebound using non-linear mixed effects models. AB - Individuals infected with the human immunodeficiency virus type 1 (HIV-1) who initiate antiretroviral therapy typically experience a marked decline in concentrations of HIV-1 RNA in plasma. Often, however, viral rebound occurs within the first year of treatment and this rebound may be associated with resistance to antiretroviral therapy. For this reason, it is important to study the patterns of virological response of HIV-1 RNA to treatment. In particular, there is interest in the relationship between the lowest level of plasma HIV-1 RNA attained after initiation of therapy (nadir value) and the time until rebound. To investigate this question, we implement a simple and flexible non linear mixed effects model for the trajectory of the HIV-1 RNA until rebound. This model is also consistent with biological insights into the effects of treatment. We also show how the problem of censoring of HIV-1 RNA values at the lower limit of assay quantification can be addressed using a multiple imputation scheme. The algorithm is simple to implement and is based on accessible software. Our application makes use of data from clinical trial 315 conducted by the AIDS Clinical Trials Group (ACTG 315). We find a strong relationship between HIV-1 RNA nadir and time to rebound, with potentially important consequences for the management of HIV-infected individuals. PMID- 12111890 TI - Kappa coefficients in medical research. AB - Kappa coefficients are measures of correlation between categorical variables often used as reliability or validity coefficients. We recapitulate development and definitions of the K (categories) by M (ratings) kappas (K x M), discuss what they are well- or ill-designed to do, and summarize where kappas now stand with regard to their application in medical research. The 2 x M(M>/=2) intraclass kappa seems the ideal measure of binary reliability; a 2 x 2 weighted kappa is an excellent choice, though not a unique one, as a validity measure. For both the intraclass and weighted kappas, we address continuing problems with kappas. There are serious problems with using the K x M intraclass (K>2) or the various K x M weighted kappas for K>2 or M>2 in any context, either because they convey incomplete and possibly misleading information, or because other approaches are preferable to their use. We illustrate the use of the recommended kappas with applications in medical research. PMID- 12111892 TI - Alternative confidence intervals for the assessment of bioequivalence in four period cross-over designs. AB - Bioequivalence studies are conducted to demonstrate equivalence in the bioavailability of the active ingredient in different formulations. The U.S. Food and Drug Administration (FDA) requires pharmaceutical companies to show bioequivalence between different formulations or generic companies to show bioequivalence between generic drugs and brand drugs before approval. A recent FDA guidance on bioequivalence proposes criteria for assessment of population bioequivalence (PBE) and individual bioequivalence (IBE) in a four-period cross over design. In this paper, computer simulation is used to compare modified large sample (MLS) upper bounds with those proposed by the FDA to test for both PBE and IBE. The comparison criteria are the ability to maintain the stated test size and the simulated power of tests based on these bounds. PMID- 12111891 TI - Adaptive dose finding for phase I clinical trials of drugs used for chemotherapy of cancer. AB - Phase I clinical trials of cancer chemotherapy drugs are intended to determine the maximum tolerable dose (MTD). Thestandard method employed is a rule-based dose-escalation scheme in which escalation depends on the number of patients at a dose level that have dose-limiting toxicity (DLT). The MTD is thus defined in terms of the rules and a series of dose levels selected for sampling. For some trials it is desirable to have a more precise definition of the MTD, and to determine the MTD more accurately than possible with the rule-based schemes. Continuous reassessment methods (CRMs) define the MTD to be the dose at which a fixed fraction of patients experience DLT, and thus appear suited to these trials. It is shown, however, that these methods can have failure modes that in fact make them unattractive. An alternative data-driven dose-finding method is described that combines the robustness of the rule-based methods and with features of CRMs. The method has two stages. In the first stage, doses are escalated by a factor of 1.5. In the second stage, which begins at the first instance of DLT, a two-parameter logistic dose-response model estimates the MTD from the DLT experience of all patients. The model is initialized by setting the dose (d10) at which 10 per cent of patients would experience DLT to half the dose at which the first DLT was observed, and the dose (d90) at which 90 per cent would experience DLT to ten times d10. Weights are assigned such that the information at d10 and d90 is equivalent to that of one patient at each of the two doses. Cohorts of three patients are treated in both stages, and the dose for a new cohort in the second stage is the estimated MTD. The only prior information required to specify the design completely is the dose which will be given to the first cohort. Two stopping rules are investigated; among the requirements for these are that at least three (or four) DLTs be observed and at least nine patients be treated in the second stage. Simulations show that a coefficient of variation of approximately 0.4 on a target DLT probability of 0.3 is obtained over a wide variation in dose-response characteristics of the study drug. The performance of the new method is compared to that of rule-based methods. PMID- 12111893 TI - Receiver operating characteristic (ROC) analysis for diagnostic examinations with uninterpretable cases. AB - Receiver operating characteristic (ROC) analysis plots true positive rates over false positive rates to describe the discriminatory power of a test to differentiate between two specifiable populations (that is 'normal' from 'abnormal') using a continuous dichotomous threshold. This assumes that the test separates cases into observed 'normal' or 'abnormal'. However, in practice many tests have some uninterpretable results as an inherent feature of the test itself, whether independent or dependent on the sample populations. This paper defines a method to describe the ability of tests to discriminate between specifiable populations when uninterpretable results occur non-informatively about disease status. A mixed model modified ROC curve is developed. Formulae to estimate the area under the modified ROC curve are given. Comparisons of conventional with the mixed model ROC analysis are shown in mathematical formulae as well as simulated experiments. An example of diagnosis of spinal fracture in renal transplant patients is presented. PMID- 12111894 TI - Analysis of mutational spectra: locating hotspots and clusters of mutations using recursive segmentation. AB - Mutations within different regions of disease-causing genes can vary in their impact on disease initiation and progression. Determining how individual mutations within such genes affect disease risk and progression can improve the accuracy of prognoses and help guide treatment selection. Estimates of mutation specific risks can be poor, however, when genes have a large number of distinct mutations, and data for any given mutation is sparse. To address this problem, we present here a method of analysing the spectrum of mutations observed across a gene that pools together mutations that appear to have similar effects on disease. One of the assumptions underlying the analysis of mutational spectra created in this manner is that the frequency of the mutation in the sample reflects the degree of its effect on disease development. Additionally, mutations that disrupt the same functionally important region of the gene are expected to have a similar impact on disease development. These mutations tend to form a cluster within the spectrum. Therefore, we developed an algorithm that segments a spectrum into regions containing sites with similar mutational frequencies, and have derived by simulation equations that allow one to evaluate whether segmentation is needed. We used this approach to investigate the spectrum of mutations observed in the p53 tumour suppressor gene in colorectal cancer tumours. Here, recursive segmentation identified the boundaries of apparent clusters better than did other methods, and this approach could identify clusters of mutations which corresponded to biologically important regions of the p53 protein. PMID- 12111895 TI - Concordance of disease form in kindreds ascertained through affected individuals. AB - When designing or conducting genetic epidemiological studies of a disease with several distinct forms, it is useful to know whether susceptibilities to the different forms are conferred by different genes or whether there are genes that confer susceptibility to multiple forms. A natural approach to exploring these issues is to examine how the disease forms cluster in kindreds. When inclusion in the study is based on the affection status of multiple relatives, however, distorted patterns of familial clustering of disease form can be evident. The purpose here is to present statistical methods for adjusting for this distortion. In particular, approaches to testing two null hypotheses are presented: a null hypothesis that corresponds to all genes acting in the same way on the relative risk of the different disease forms, and a null hypothesis that corresponds to each gene conferring susceptibility to distinct disease forms. The approaches are illustrated through an application to the generalized and localization-related forms of epilepsy. PMID- 12111896 TI - A Pearson-type goodness-of-fit test for stationary and time-continuous Markov regression models. AB - Markov regression models describe the way in which a categorical response variable changes over time for subjects with different explanatory variables. Frequently it is difficult to measure the response variable on equally spaced discrete time intervals. Here we propose a Pearson-type goodness-of-fit test for stationary Markov regression models fitted to panel data. A parametric bootstrap algorithm is used to study the distribution of the test statistic. The proposed technique is applied to examine the fit of a Markov regression model used to identify markers for disease progression in psoriatic arthritis. PMID- 12111897 TI - Diagnostics for conformity of paired quantitative measurements. AB - Matched pairs data arise in many contexts - in case-control clinical trials, for example, and from cross-over designs. They also arise in experiments to verify the equivalence of quantitative assays. This latter use (which is the main focus of this paper) raises difficulties not always seen in other matched pairs applications. Since the designs deliberately vary the analyte levels over a wide range, issues of variance dependent on mean, calibrations of differing slopes, and curvature all need to be added to the usual model assumptions such as normality. Violations in any of these assumptions invalidate the conventional matched pairs analysis. A graphical method, due to Bland and Altman, of looking at the relationship between the average and the difference of the members of the pairs is shown to correspond to a formal testable regression model. Using standard regression diagnostics, one may detect and diagnose departures from the model assumptions and remedy them - for example using variable transformations. Examples of different common scenarios and possible approaches to handling them are shown. PMID- 12111898 TI - Permutation tests for detecting and estimating mixtures in task performance within groups. AB - We propose a two-sample permutation test incorporating mixture models as a general tool for detecting and quantifying effects on task performance. We illustrate the proposed method with examples where the dependent measures under investigation are recorded for normal controls and relatives of patients with schizophrenia on a delayed response, spatial and object working memory task. Our mixture modelling in relatives allows the component distributions to arise from different continuous parametric families. We also investigate the effects of the within-family correlation and the prior distribution of the mixing proportion on the test results. The power of the test depends on sample sizes, the mixing proportion, the difference in component means and the ratio of component variances. PMID- 12111899 TI - Power comparison of robust approximate and non-parametric tests for the analysis of cross-over trials by J.A. Correa and F. Bellavance, Statistics in Medicine 2001; 20: 1185-1196. PMID- 12111901 TI - Test statistics and sample size formulae for comparative binomial trials with null hypothesis of non-zero risk difference or non-unity relative risk by C. P. Farrington and G. Manning, Statistics in Medicine 1990; 9: 1447-1454. PMID- 12111903 TI - Using inverse weighting and predictive inference to estimate the effects of time varying treatments on the discrete-time hazard. AB - We estimate the effects of non-randomized time-varying treatments on the discrete time hazard, using inverse weighting. We consider the special monotone pattern of treatment that develops over time as subjects permanently discontinue an initial treatment, and assume that treatment selection is sequentially ignorable. We use a propensity score in the hazard model to reduce the potential for finite-sample bias due to inverse weighting. When the number of subjects who discontinue treatment at any given time is small, we impose scientific restrictions on the potentially observable discontinuation hazards to improve efficiency. We use predictive inference to account for the correlation of the potential hazards, when comparing outcomes under different durations of initial treatment. PMID- 12111906 TI - Estimating the causal effect of zidovudine on CD4 count with a marginal structural model for repeated measures. AB - Even in the absence of unmeasured confounding factors or model misspecification, standard methods for estimating the causal effect of a time-varying treatment on the mean of a repeated measures outcome (for example, GEE regression) may be biased when there are time-dependent variables that are simultaneously confounders of the effect of interest and are predicted by previous treatment. In contrast, the recently developed marginal structural models (MSMs) can provide consistent estimates of causal effects when unmeasured confounding and model misspecification are absent. We describe an MSM for repeated measures that parameterizes the marginal means of counterfactual outcomes corresponding to prespecified treatment regimes. The parameters of MSMs are estimated using a new class of estimators - inverse-probability of treatment weighted estimators. We used an MSM to estimate the effect of zidovudine therapy on mean CD4 count among HIV-infected men in the Multicenter AIDS Cohort Study. We estimated a potential expected increase of 5.4 (95 per cent confidence interval -1.8,12.7) CD4 lymphocytes/l per additional study visit while on zidovudine therapy. We also explain the theory and implementation of MSMs for repeated measures data and draw upon a simple example to illustrate the basic ideas. PMID- 12111907 TI - Early selection in a randomized phase II clinical trial. AB - A randomized phase II clinical trial design can be employed when one wishes to be able to select one of several similar therapies or variants of a therapy for inclusion in a subsequent, definitive, phase III trial. It is not necessary in this type of trial to formally identify a superior arm using the usual parameters and stringent criteria employed for hypothesis testing. Simon, Wittes and Ellenberg have described methods and sample sizes for selecting the superior arm from among k possible arms. In this paper, we describe an approach based on statistical selection theory which allows one to potentially make a decision to end accrual to a randomized phase II trial at an interim point of the trial, and to select one of two arms as being worthy of further evaluation in a subsequent study. This method requires that an adequate gap in the number of responses between the two arms be observed at the interim point in order to limit the probability that the selected arm is actually inferior by more than an indifference amount. Published in 2002 by John Wiley & Sons, Ltd. PMID- 12111908 TI - Reproducibility probability in clinical trials. AB - For marketing approval of a new drug product, the United States Food and Drug Administration (FDA) requires that substantial evidence of the effectiveness of the drug product be provided through the conduct of at least two adequate and well-controlled clinical trials. The purpose of conducting the second clinical trial is to study whether the clinical result from the first trial is reproducible in the second trial with the same study protocol. Under certain circumstance, the FDA Modernization Act of 1997 includes a provision to allow data from one adequate and well-controlled clinical trial investigation and confirmatory evidence to establish effectiveness for risk/benefit assessment of drug and biological candidates for approval. In this paper, we introduce the concept of reproducibility probability for a given clinical trial, which is useful in providing important information for regulatory agencies in deciding whether a single clinical trial is sufficient and for pharmaceutical companies in adjusting the sample size in a future clinical trial. Three approaches, the estimated power approach, the method of confidence bounds and the Bayesian approach, are studied in evaluating reproducibility probabilities under several study designs commonly used in clinical trials. PMID- 12111909 TI - Assessing rater agreement using marginal association models. AB - New models that are useful in the assessment of rater agreement, particularly when the rating scale is ordered or partially ordered, are presented. The models are parameterized to address two important aspects of rater agreement: (i) agreement in terms of the overall frequency in which raters assign categories; and (ii) the extent to which raters agree on the category assigned to individual subjects or items. We present methodology for the simultaneous modelling of univariate marginal responses and bivariate marginal associations in the K-way contingency table representing the joint distribution of K rater responses. The univariate marginal responses provide information for evaluating agreement in terms of the overall frequency of responses, and the bivariate marginal associations provide information on category-wise agreement among pairs of raters. In addition, estimated scores within a generalized log non-linear model for bivariate associations facilitate the assessment of category distinguishability. PMID- 12111910 TI - Simultaneous estimation of intrarater and interrater agreement for multiple raters under order restrictions for a binary trait. AB - It is valuable in many studies to assess both intrarater and interrater agreement. Most measures of intrarater agreement do not adjust for unequal estimates of prevalence between the separate rating occasions for a given rater and measures of interrater agreement typically ignore data from the second set of assessments when raters make duplicate assessments. In the event when both measures are assessed there are instances where interrater agreement is larger than at least one of the corresponding intrarater agreements. This implies that a rater agrees less with him/herself and more with another rater. In the situation of multiple raters making duplicate assessments on all subjects, the authors propose properties for an agreement measure based on the odds ratio for a dichotomous trait: (i) estimate a single prevalence across two reading occasions for each rater; (ii) estimate pairwise interrater agreement from all available data; (iii) bound the pairwise interrater agreement above by the corresponding intrarater agreements. Estimation of odds ratios under these properties is done by maximizing the multinomial likelihood with constraints using generalized log linear models in combination with a generalization of the Lemke-Dykstra iterative incremental algorithm. An example from a mammography examination reliability study is used to demonstrate the new method. PMID- 12111911 TI - The predictive value of microbiologic diagnostic tests if asymptomatic carriers are present. AB - If a proper gold standard is not available, then the predictive value of a test cannot be estimated. In this paper the concept of etiologic predictive value (EPV) is introduced. It is a quantity that will yield the predictive value of a test to predict presence of a specified disease in situations for which no proper gold standard is available. This is achieved by using information obtained from a healthy control population. This quantity requires that the marker in our test is present in all individuals having the specified disease, as in the case where the marker is the aetiologic factor for the specified disease. Furthermore this quantity requires that asymptomatic carriers are present. This means that not all individuals with the marker has the specified disease. EPV is developed with special reference to the evaluation of bacterial cultures, or rapid tests to detect a bacterium, but the quantity might be used in other circumstances as well. EPV is applied to an example in which conventional throat culture is evaluated. Further information concerning EPV can be found at http://www.infovice.se/fou/epv. PMID- 12111912 TI - Sample size and power determination for clustered repeated measurements. AB - It is common in epidemiological and clinical studies that each subject has repeated measurements on a single common variable, while the subjects are also 'clustered'. To compute sample size or power of a test, we have to consider two types of correlation: correlation among repeated measurements within the same subject, and correlation among subjects in the same cluster. We develop, based on generalized estimating equations, procedures for computing sample size and power with clustered repeated measurements. Explicit formulae are derived for comparing two means, two slopes and two proportions, under several simple correlation structures. PMID- 12111913 TI - A comparison of methods to detect publication bias in meta-analysis by P. Macaskill, S. D. Walter and L. Irwig, Statistics in Medicine, 2001; 20:641-654. PMID- 12111915 TI - Proceedings of the 3rd Symposium on Systemic Review Methodology. Oxford, England. July 3-5, 2000. PMID- 12111916 TI - Investigating causes of heterogeneity in systematic reviews. AB - What causes heterogeneity in systematic reviews of controlled trials? First, it may be an artefact of the summary measures used, of study design features such as duration of follow-up or the reliability of outcome measures. Second, it may be due to real variation in the treatment effect and hence provides the opportunity to identify factors that may modify the impact of treatment. These factors may include features of the population such as: severity of illness, age and gender; intervention factors such as dose, timing or duration of treatment; and comparator factors such as the control group treatment or the co-interventions in both groups. The ideal way to study causes of true variation is within rather than between studies. In most situations however, we will have to make do with a study level investigation and hence need to be careful about adjusting for potential confounding by artefactual factors such as study design features. Such investigation of artefactual and true causes of heterogeneity form essential steps in moving from a combined effect estimate to application to particular populations and individuals. PMID- 12111917 TI - Statistical methods for assessing the influence of study characteristics on treatment effects in 'meta-epidemiological' research. AB - Biases in systematic reviews and meta-analyses may be examined in 'meta epidemiological' studies, in which the influence of trial characteristics such as measures of study quality on treatment effect estimates is explored. Published studies to date have analysed data from collections of meta-analyses with binary outcomes, using logistic regression models that assume that there is no between- or within-meta-analysis heterogeneity. Using data from a study of publication bias (39 meta-analyses, 394 published and 88 unpublished trials) and language bias (29 meta-analyses, 297 English language trials and 52 non-English language trials), we compare results from logistic regression models, with and without robust standard errors to allow for clustering on meta-analysis, with results using a 'meta-meta-analytic' approach that can allow for between- and within-meta analysis heterogeneity. We also consider how to allow for the confounding effects of different trial characteristics. We show that both within- and between meta analysis heterogeneity may be of importance in the analysis of meta epidemiological studies, and that confounding exists between the effects of publication status and trial quality. PMID- 12111918 TI - Exploring sources of heterogeneity in systematic reviews of diagnostic tests. AB - It is indispensable for any meta-analysis that potential sources of heterogeneity are examined, before one considers pooling the results of primary studies into summary estimates with enhanced precision. In reviews of studies on the diagnostic accuracy of tests, variability beyond chance can be attributed to between-study differences in the selected cutpoint for positivity, in patient selection and clinical setting, in the type of test used, in the type of reference standard, or any combination of these factors. In addition, heterogeneity in study results can also be caused by flaws in study design. This paper critically examines some of the potential reasons for heterogeneity and the methods to explore them. Empirical support for the existence of different sources of variation is reviewed. Incorporation of sources of variability explicitly into systematic reviews on diagnostic accuracy is demonstrated with data from a recent review. Application of regression techniques in meta-analysis of diagnostic tests can provide relevant additional information. Results of such analyses will help understand problems with the transferability of diagnostic tests and to point out flaws in primary studies. As such, they can guide the design of future studies. PMID- 12111919 TI - Quantifying heterogeneity in a meta-analysis. AB - The extent of heterogeneity in a meta-analysis partly determines the difficulty in drawing overall conclusions. This extent may be measured by estimating a between-study variance, but interpretation is then specific to a particular treatment effect metric. A test for the existence of heterogeneity exists, but depends on the number of studies in the meta-analysis. We develop measures of the impact of heterogeneity on a meta-analysis, from mathematical criteria, that are independent of the number of studies and the treatment effect metric. We derive and propose three suitable statistics: H is the square root of the chi2 heterogeneity statistic divided by its degrees of freedom; R is the ratio of the standard error of the underlying mean from a random effects meta-analysis to the standard error of a fixed effect meta-analytic estimate, and I2 is a transformation of (H) that describes the proportion of total variation in study estimates that is due to heterogeneity. We discuss interpretation, interval estimates and other properties of these measures and examine them in five example data sets showing different amounts of heterogeneity. We conclude that H and I2, which can usually be calculated for published meta-analyses, are particularly useful summaries of the impact of heterogeneity. One or both should be presented in published meta-analyses in preference to the test for heterogeneity. PMID- 12111920 TI - How should meta-regression analyses be undertaken and interpreted? AB - Appropriate methods for meta-regression applied to a set of clinical trials, and the limitations and pitfalls in interpretation, are insufficiently recognized. Here we summarize recent research focusing on these issues, and consider three published examples of meta-regression in the light of this work. One principal methodological issue is that meta-regression should be weighted to take account of both within-trial variances of treatment effects and the residual between trial heterogeneity (that is, heterogeneity not explained by the covariates in the regression). This corresponds to random effects meta-regression. The associations derived from meta-regressions are observational, and have a weaker interpretation than the causal relationships derived from randomized comparisons. This applies particularly when averages of patient characteristics in each trial are used as covariates in the regression. Data dredging is the main pitfall in reaching reliable conclusions from meta-regression. It can only be avoided by prespecification of covariates that will be investigated as potential sources of heterogeneity. However, in practice this is not always easy to achieve. The examples considered in this paper show the tension between the scientific rationale for using meta-regression and the difficult interpretative problems to which such analyses are prone. PMID- 12111921 TI - Issues in the selection of a summary statistic for meta-analysis of clinical trials with binary outcomes. AB - Meta-analysis of binary data involves the computation of a weighted average of summary statistics calculated for each trial. The selection of the appropriate summary statistic is a subject of debate due to conflicts in the relative importance of mathematical properties and the ability to intuitively interpret results. This paper explores the process of identifying a summary statistic most likely to be consistent across trials when there is variation in control group event rates. Four summary statistics are considered: odds ratios (OR); risk differences (RD) and risk ratios of beneficial (RR(B)); and harmful outcomes (RR(H)). Each summary statistic corresponds to a different pattern of predicted absolute benefit of treatment with variation in baseline risk, the greatest difference in patterns of prediction being between RR(B) and RR(H). Selection of a summary statistic solely based on identification of the best-fitting model by comparing tests of heterogeneity is problematic, principally due to low numbers of trials. It is proposed that choice of a summary statistic should be guided by both empirical evidence and clinically informed debate as to which model is likely to be closest to the expected pattern of treatment benefit across baseline risks. Empirical investigations comparing the four summary statistics on a sample of 551 systematic reviews provide evidence that the RR and OR models are on average more consistent than RD, there being no difference on average between RR and OR. From a second sample of 114 meta-analyses evidence indicates that for interventions aimed at preventing an undesirable event, greatest absolute benefits are observed in trials with the highest baseline event rates, corresponding to the model of constant RR(H). The appropriate selection for a particular meta-analysis may depend on understanding reasons for variation in control group event rates; in some situations uncertainty about the choice of summary statistic will remain. PMID- 12111922 TI - Bayesian random effects meta-analysis of trials with binary outcomes: methods for the absolute risk difference and relative risk scales. AB - When conducting a meta-analysis of clinical trials with binary outcomes, a normal approximation for the summary treatment effect measure in each trial is inappropriate in the common situation where some of the trials in the meta analysis are small, or the observed risks are close to 0 or 1. This problem can be avoided by making direct use of the binomial distribution within trials. A fully Bayesian method has already been developed for random effects meta-analysis on the log-odds scale using the BUGS implementation of Gibbs sampling. In this paper we demonstrate how this method can be extended to perform analyses on both the absolute and relative risk scales. Within each approach we exemplify how trial-level covariates, including underlying risk, can be considered. Data from 46 trials of the effect of single-dose ibuprofen on post-operative pain are analysed and the results contrasted with those derived from classical and Bayesian summary statistic methods. The clinical interpretation of the odds ratio scale is not straightforward. The advantages and flexibility of a fully Bayesian approach to meta-analysis of binary outcome data, considered on an absolute risk or relative risk scale, are now available. PMID- 12111923 TI - A comparison of handsearching versus MEDLINE searching to identify reports of randomized controlled trials. AB - This study aims to compare handsearching to a basic MEDLINE search for the identification of reports of randomized trials in specialized health care journals. Twenty-two specialized health care journals, published in the U.K., were handsearched for all reports of controlled trials (as defined by the Cochrane Collaboration). The reports of trials, which were judged to be definitely randomized, were identified from a random sample of three years per journal and form one element of this study. A MEDLINE search using the publication type terms 'randomized controlled trial' and 'controlled clinical trial' was also performed for the same journal years. The reports of trials retrieved by handsearching were then compared against those retrieved from the MEDLINE search, to identify differences in retrieval between the two techniques. Reports of randomized trials identified by the MEDLINE search but not found by handsearching were individually assessed to see if they met the Cochrane eligibility criteria for a report of a randomized trial. A total of 714 reports of randomized trials were found by using a combination of both handsearching and MEDLINE searching. Of these, 369 (52 per cent) were identified only by handsearching and 32 (4 per cent) were identified only by MEDLINE searching. Of the reports identified only by handsearching, 252 had no MEDLINE record, of which 232 (92 per cent) were meeting abstracts or published in supplements; 117 (25 per cent) of the 462 reports of randomized trials which had a MEDLINE record were missed by the electronic search because they did not have either of the publication type terms 'randomized controlled trial' or 'controlled clinical trial'. This proportion varied depending on when the reports of randomized trials were published (that is, before or after the introduction of the MEDLINE publication type terms above). The highest additional yield from handsearching compared to MEDLINE searching was for reports of randomized trials published prior to 1991 and from handsearching the non-MEDLINE indexed parts of a journal. The results of this study suggest that a combination of MEDLINE and handsearching is required to identify adequately reports of randomized trials. PMID- 12111924 TI - Identification of randomized controlled trials in systematic reviews: accuracy and reliability of screening records. AB - A study was conducted to estimate the accuracy and reliability of reviewers when screening records for relevant trials for a systematic review. A sensitive search of ten electronic bibliographic databases yielded 22 571 records of potentially relevant trials. Records were allocated to four reviewers such that two reviewers examined each record and so that identification of trials by each reviewer could be compared with those identified by each of the other reviewers. Agreement between reviewers was assessed using Cohen's kappa statistic. Ascertainment intersection methods were used to estimate the likely number of trials missed by reviewers. Full copies of reports were obtained and assessed independently by two researchers for eligibility for the review. Eligible reports formed the 'gold standard' against which an assessment was made about the accuracy of screening by reviewers. After screening, 301 of 22 571 records were identified by at least one reviewer as potentially relevant. Agreement was 'almost perfect' (kappa>0.8) within two pairs, 'substantial' (kappa>0.6) within three pairs and 'moderate' (kappa>0.4) within one pair. Of the 301 records selected, 273 complete reports were available. When pairs of reviewers agreed on the potential relevance of records, 81 per cent were eligible (range 69 to 91 per cent). If reviewers disagreed, 22 per cent were eligible (range 12 to 45 per cent). Single reviewers missed on average 8 per cent of eligible reports (range 0 to 24 per cent), whereas pairs of reviewers did not miss any (range 0 to 1 per cent). The use of two reviewers to screen records increased the number of randomized trials identified by an average of 9 per cent (range 0 to 32 per cent). Reviewers can reliably identify potentially relevant records when screening thousands of records for eligibility. Two reviewers should screen records for eligibility, whenever possible, in order to maximize ascertainment of relevant trials. PMID- 12111925 TI - Simultaneous echo refocusing in EPI. AB - A method to encode multiple two-dimensional Fourier transform (2D FT) images within a single echo train is presented. This new method, simultaneous echo refocusing (SER), is a departure from prior echo planar image (EPI) sequences which use repeated single-shot echo trains for multislice imaging. SER simultaneously acquires multiple slices in a single-shot echo train utilizing a shared refocusing process. The SER technique acquires data faster than conventional multislice EPI since it uses fewer gradient switchings and fewer preparation pulses such as diffusion gradients. SER introduces a new capability to simultaneously record multiple spatially separated sources of physiologic information in subsecond image acquisitions, which enables several applications that are dependent on temporal coherence in MRI data including velocity vector field mapping and brain activation mapping. PMID- 12111926 TI - On the application of a non-CPMG single-shot fast spin-echo sequence to diffusion tensor MRI of the human brain. AB - The strong sensitivity of Carr-Purcell-Meiboom-Gill (CPMG) fast spin-echo (FSE) sequences, such as rapid acquisition with relaxation enhancement (RARE), to the phase of the prepared transverse magnetization means that artifact-free single shot diffusion-weighted images can currently only be obtained with a 30-50% reduction in the signal-to-noise ratio (SNR). However, this phase sensitivity and signal loss can be addressed in FSE sequences that use quadratic phase modulation of the radiofrequency (RF) refocusing pulses to generate a sustained train of stable echoes. Here the first application of such a non-CPMG single-shot FSE (ssFSE) sequence to diffusion tensor MR imaging (DT-MRI) of the human brain is described. This approach provides high SNR diffusion-weighted images that have little or no susceptibility to poor B(0) magnetic field homogeneity and the strong eddy currents typically present in DT-MRI experiments. PMID- 12111927 TI - Effect of adrenergic stimulation on Rb(+) uptake in normal and ischemic areas of isolated pig hearts: (87)Rb MRI study. AB - The role of adrenergic stimulation in Rb(+) uptake in normal and ischemic areas of pig hearts was investigated using an agonist (dobutamine) and an antagonist (S propranolol). The left anterior descending artery (LAD) in isolated pig hearts was cannulated to maintain adequate perfusion of the LAD bed (2.2-2.8 ml/min/g). Rb(+) loading was initiated by switching perfusion to Rb(+)-containing Krebs Henseleit (KH) buffer in the presence of 0.1 microM dobutamine, 0.5 microM propranolol, or no drug (control), and the LAD flow was reduced to 10% of the baseline for 120 min. The flow through the LAD was then restored to normal and the drugs were removed. In all groups the rate of Rb(+) uptake obtained from serial (87)Rb images was severely depressed in the ischemic anterior wall relative to that in the normal posterior wall. Dobutamine increased the rate of Rb(+) uptake in the posterior wall (6.3% +/- 1.4% vs. 2.5% +/- 0.25% per minute in control) and did not affect the rate in the anterior wall (0.86% +/- 0.40% vs. 1.14% +/- 0.19% per minute in control). Propranolol did not affect Rb(+) uptake in either of these areas. We conclude that the stimulation of Rb(+) uptake by dobutamine is related to activation of Na(+)/K(+) ATPase and passive Na(+) influx in normal tissue. The lack of any effect of propranolol implies a low level of endogenous norepinephrine during low-flow (LF) ischemia. PMID- 12111928 TI - Magnetic field dependence of proton spin-lattice relaxation times. AB - The magnetic field dependence of the water-proton spin-lattice relaxation rate (1/T(1)) in tissues results from magnetic coupling to the protons of the rotationally immobilized components of the tissue. As a consequence, the magnetic field dependence of the water-proton (1/T(1)) is a scaled report of the field dependence of the (1/T(1)) rate of the solid components of the tissue. The proton spin-lattice relaxation rate may be represented generally as a power law: 1/T(1)omega = A omega(-b), where b is usually found to be in the range of 0.5 0.8. We have shown that this power law may arise naturally from localized structural fluctuations along the backbone in biopolymers that modulate the proton dipole-dipole couplings. The protons in a protein form a spin communication network described by a fractal dimension that is less than the Euclidean dimension. The model proposed accounts quantitatively for the proton spin-lattice relaxation rates measured in immobilized protein systems at different water contents, and provides a fundamental basis for understanding the parametric dependence of proton spin-lattice relaxation rates in dynamically heterogeneous systems, such as tissues. PMID- 12111929 TI - Improved accuracy of human cerebral blood perfusion measurements using arterial spin labeling: accounting for capillary water permeability. AB - A two-compartment exchange model for perfusion quantification using arterial spin labeling (ASL) is presented, which corrects for the assumption that the capillary wall has infinite permeability to water. The model incorporates an extravascular and a blood compartment with the permeability surface area product (PS) of the capillary wall characterizing the passage of water between the compartments. The new model predicts that labeled spins spend longer in the blood compartment before exchange. This makes an accurate blood T(1) measurement crucial for perfusion quantification; conversely, the tissue T(1) measurement is less important and may be unnecessary for pulsed ASL experiments. The model gives up to 62% reduction in perfusion estimate for human imaging at 1.5T compared to the single compartment model. For typical human perfusion rates at 1.5T it can be assumed that the venous outflow signal is negligible. This simplifies the solution, introducing only one more parameter than the single compartment model, PS/v(bw), where v(bw) is the fractional blood water volume per unit volume of tissue. The simplified model produces an improved fit to continuous ASL data collected at varying delay time. The fitting yields reasonable values for perfusion and PS/v(bw). PMID- 12111930 TI - Apparent diffusion of water, ions, and small molecules in the Xenopus oocyte is consistent with Brownian displacement. AB - The incoherent displacement of water in living tissues is of considerable interest because of the widespread use of diffusion-weighted MRI, for which image contrast is based on the water apparent diffusion coefficient (ADC). It has been hypothesized that the decrease in water ADC associated with brain injury is primarily due to a reduction in the ADC of water in the intracellular space. Xenopus oocytes permit direct measurement of ADC values for intracellular molecules, thereby providing insight into the nature of intracellular motion. In this study, the measured ADC values of small molecules and ions are shown to be primarily size-dependent, indicating that intracellular water motion in the oocyte is mainly Brownian displacement with little or no role for cytoplasmic streaming. Further, intracellular water ADC values show no dependence on diffusion time over a broad range (3.4-100 ms), suggesting that barriers to displacement are finely spaced (< or = 2-3 microm). The water diffusion shows some small anisotropy, suggesting that the cell has structure, giving water displacement a directional preference. The calculated intracellular apparent viscosity, which reflects the combined effects of barriers to motion, intermolecular binding, and fluid phase viscosity was 2.07 +/- 0.09 cP. PMID- 12111931 TI - Application of the static dephasing regime theory to superparamagnetic iron-oxide loaded cells. AB - The relaxation rates of iron-oxide nanoparticles compartmentalized within cells were studied and found to satisfy predictions of the static dephasing (SD) regime theory. THP-1 cells in cell culture were loaded using two different iron-oxide nanoparticles (superparamagnetic iron-oxide (SPIO) and ultrasmall SPIO (USPIO)) with four different iron concentrations (0.05, 0.1, 0.2, and 0.3 mg/ml) and for five different incubation times (6, 12, 24, 36, and 48 hr). Cellular iron-oxide uptake was assessed using a newly developed imaging version of MR susceptometry, and was found to be linear with both dose and incubation time. R(2)* sensitivity to iron-oxide loaded cells was found to be 70 times greater than for R(2), and 3100 times greater than for R(1). This differs greatly from uniformly distributed nanoparticles and is consistent with a cellular bulk magnetic susceptibility (BMS) relaxation mechanism. The cellular magnetic moment was large enough that R(2)' relaxivity agreed closely with SD regime theory predictions for all cell samples tested [R(2)'=2 pi/(9 x the square root of 3) x gamma LMD] where the local magnetic dose (LMD) is the sample magnetization due to the presence of iron oxide particles). Uniform suspensions of SPIO and USPIO produced R(2)' relaxivities that were a factor of 3 and 8 less, respectively, than SD regime theory predictions. These results are consistent with theoretical estimates of the required mass of iron per compartment needed to guarantee SD-regime-dominant relaxivity. For cellular samples, R(2) was shown to be dependent on both the concentration and distribution of iron-oxide particles, while R(2)' was sensitive to iron-oxide concentration alone. This work is an important first step in quantifying cellular iron content and ultimately mapping the density of a targeted cell population. PMID- 12111932 TI - Test-retest reliability estimation of functional MRI data. AB - Functional magnetic resonance imaging (fMRI) data are commonly used to construct activation maps for the human brain. It is important to quantify the reliability of such maps. We have developed statistical models to provide precise estimates for reliability from several runs of the same paradigm over time. Specifically, our method extends the premise of maximum likelihood (ML) developed by Genovese et al. (Magn Reson Med 1997;38:497-507) by incorporating spatial context into the estimation process. Experiments indicate that our methodology provides more conservative estimates of true positives compared to those obtained by Genovese et al. The reliability estimates can be used to obtain voxel-specific reliability measures for activated as well as inactivated regions in future experiments. We derive statistical methodology to determine optimal thresholds for region- and context-specific activations. Empirical guidelines are also provided on the number of repeat scans to acquire in order to arrive at accurate reliability estimates. We report the results from experiments involving a motor paradigm performed on a single subject several times over a period of 2 months. PMID- 12111933 TI - Changes in axonal morphology in experimental autoimmune neuritis as studied by high b-value q-space (1)H and (2)H DQF diffusion magnetic resonance spectroscopy. AB - Experimental autoimmune neuritis (EAN) has been studied in rat sciatic nerves by a combination of high b-value (1)H and (2)H double quantum filtered (DQF) diffusion MRS. The signal decays of water in the (1)H and (2)H DQF diffusion MRS were found to be not monoexponential and were analyzed using the q-space approach. The q-space analysis of the (1)H diffusion data detected two diffusing components, one having broad and the other having narrow displacement profiles. These components were shown to be very sensitive to the progression of EAN disease. The q-space parameters were found to be abnormal at day 9 postimmunization before the appearance of clinical signs. The assignment of the component with the narrow displacement profile to axonal water has been corroborated by the (2)H DQF diffusion MRS results. The displacement and the relative population of this slow and restricted diffusing component followed the processes of demyelination, axonal loss, and remyelination that occur in EAN. The displacements extracted from the slow-diffusing component with the narrow displacement correlated well with the average size of the axons as deduced from electron microscopy (EM). The component with the broad displacement showed significant changes which were attributed to the formation of endoneurial edema. This observation was also corroborated by the (2)H DQF diffusion MRS experiments. It seems, therefore, that q-space analysis of high b-values diffusion MRS is a promising new approach for early detection and better characterization of the different pathologies associated with EAN. This study demonstrates the utility of high-b-value q-space diffusion MRS for studying white matter-associated disorders in general. PMID- 12111935 TI - Imaging of intracellular sodium with shift reagent aided (23)Na CSI in isolated rat hearts. AB - 23Na chemical shift imaging (CSI) in conjunction with shift reagents was used to obtain images of intracellular (Na(i)) and extracellular sodium (Na(e)) in isolated rat hearts. It was demonstrated that the increase of Na(i) concentration in ischemic myocardium can be detected with this technique. 3D acquisition weighted (23)Na CSI datasets with a nominal spatial resolution of 1.7 x 1.7 x 2.9 mm were acquired in 30 min in normoxic hearts and in globally or locally ischemic hearts. The shift reagent Tm(DOTP)(5-) was used to discriminate Na(i) and Na(e) signals. Na(i) maps could be generated in ischemic hearts, but not in normoxic hearts as the signal-to-noise ratio is too low. The Na(i) signal increased by more than 100% and the Na(e) signal decreased by more than 50% in myocardium of globally ischemic hearts (n = 3) compared to normoxic hearts (n = 3). In hearts with an acute occlusion of the left anterior descending coronary artery (n = 3), there was a local Na(i) signal increase in the anterior wall in the range of 60 110% compared to remote, normoxic tissue. PMID- 12111934 TI - High-resolution MRI of cardiac function with projection reconstruction and steady state free precession. AB - The purpose of this study was to investigate the trabecular structure of the endocardial wall of the living human heart, and the effect of that structure on the measurement of myocardial function using MRI. High-resolution MR images (0.8 x 0.8 x 8 mm voxels) of cardiac function were obtained in five volunteers using a combination of undersampled projection reconstruction (PR) and steady-state free precession (SSFP) contrast in ECG-gated breath-held scans. These images provide movies of cardiac function with new levels of endocardial detail. The trabecular papillary muscle complex, consisting of a mixture of blood and endocardial structures, is measured to constitute as much as 50% of the myocardial wall in some sectors. Myocardial wall strain measurements derived from tagged MR images show correlation between regions of trabeculae and papillary muscles and regions of high strain, leading to an overestimation of function in the lateral wall. PMID- 12111936 TI - Validation of diffusion tensor MRI-based muscle fiber tracking. AB - Diffusion-tensor (DT) MRI fiber tracking may potentially be used for in vivo structural analysis. The purpose of this study was to assess quantitatively the ability of a DT-MRI fiber-tracking algorithm to measure the fiber orientation (pennation) in skeletal muscle in vivo. In five adult Sprague-Dawley rats, the pennation angle (theta) was measured in the rat lateral gastrocnemius with DT-MRI (theta(DT-MRI)) and by direct anatomical inspection (DAI) (theta(DAI)). The mean theta(DT-MRI) was not significantly different from the mean theta(DAI). In addition, the two methods were highly correlated (r = 0.89) and the regression of theta(DT-MRI) on theta(DAI) resulted in a slope not significantly different from 1 and an intercept not significantly different from zero. These data indicate that DT-MRI-based fiber tracking as implemented here is a valid tool for in vivo structural analysis of small-animal skeletal muscle. PMID- 12111937 TI - Cardiac diffusion MRI without motion effects. AB - We present a method for diffusion tensor MRI in the beating heart that is insensitive to cardiac motion and strain. Using a stimulated echo pulse sequence with two electrocardiogram (ECG) triggers, diffusion-encoding bipolar gradient pulses are applied at identical phases in consecutive cardiac cycles. In this experiment, diffusion is encoded at a single phase in the cardiac cycle of less than 30 ms in duration. This encoding produces no phase shifts for periodic motion and is independent of intervening strains. Studies in a gel phantom with cyclic deformation confirm that by using this sequence we can map the diffusion tensor free of effects of cyclic motion. In normal human subjects, myocardial diffusion eigenvalues measured with the present method showed no significant change between acquisitions encoded at maximum contractile velocity (peak) vs. at myocardial standstill (end-systole), demonstrating motion independence of in vivo diffusion measurements. Diffusion tensor images acquired with the present method agree with registered data acquired with a previous cardiac diffusion MRI method that was shown to be valid in the normal heart, strongly supporting the validity of MRI diffusion measurement in the beating heart. Myocardial sheet and fiber dynamics measured during systole showed that normal human myocardial sheet orientations tilt toward the radial during systole, and fiber orientations tilt toward the longitudinal, in qualitative agreement with previous invasive studies in canines. These results demonstrate the technique's ability to measure myocardial diffusion accurately at any point in the cardiac cycle free of measurable motion effect, as if the heart were frozen at the point of acquisition. PMID- 12111938 TI - Pulse-wave velocity measured in one heartbeat using MR tagging. AB - A noninvasive method for measuring the aortic pulse-wave velocity (PWV) in a single heartbeat is introduced. The method sinusoidally tags a column of blood within the vessel, and rapidly acquires a series of 1D projections of the tags as they move (in practice, 64 projections at 4-ms intervals). From these projections, the relative motion of blood at different positions along the vessel is measured. The PWV is obtained by fitting a mathematical model of blood flow to the tag trajectories. Tests of this method in a pulsatile flow phantom are presented using latex and polyurethane tubes. The PWV measured in these tubes was (mean +/- standard deviation) 4.4 +/- 0.5 m/s and 2.3 +/- 0.2 m/s, respectively. The distensibility of each tube was calculated from the PWV (latex = (7 +/- 2) 10(-3) mm Hg(-1), poly. = (25 +/- 4) 10(-3)mmHg(-1)) and found to agree within error with distensibility measurements based on the change of tube area with pressure (latex = (6.3 +/- 0.3) 10(-3)mmHg(-1), poly. = (27 +/- 1) 10(-3) mmHg( 1)). To test its feasibility, the PWV measurement was applied to four normal volunteers. The measured PWV values were 3.9 +/- 0.8 m/s, 3.6 +/- 0.9 m/s, 3.9 +/ 0.5 m/s, and 5.3 +/- 0.8 m/s. By acquiring an independent PWV measurement each heartbeat, errors introduced by arrhythmia and trigger variability appear to be avoided with this method. PMID- 12111939 TI - Extravascular proton-density changes as a non-BOLD component of contrast in fMRI of the human spinal cord. AB - The fractional signal intensity change (Delta S/S) observed during activation in T(2)-weighted fMRI of the spinal cord has previously been shown to depend linearly on the echo time (TE) but to have a positive value of roughly 2.5% extrapolated to zero TE. In this study we investigated the origin of this finding by measuring the Delta S/S in spinal fMRI with very short TEs. Our results demonstrate that the Delta S/S does not approach zero, but has a value as high as 3.3% at TE = 11 ms. At TEs > 33 ms we observed the linear relationship between Delta S/S and TE as in previous studies. These data demonstrate that there is a non-BOLD contribution to signal changes observed in spinal fMRI. We hypothesize that this contribution is a local proton density increase due to increased water exudation from capillaries with increased blood flow during neuronal activation, and term this effect "signal enhancement by extravascular protons" (SEEP). PMID- 12111940 TI - Diffusion tensor imaging using single-shot SENSE-EPI. AB - SENSitivity Encoding (SENSE) greatly enhances the quality of diffusion-weighted echo-planar imaging (EPI) by reducing blurring and off-resonance artifacts. Such improvement would also be desirable for diffusion tensor imaging (DTI), but measures derived from the diffusion tensor can be extremely sensitive to any kind of image distortion. Whether DTI is feasible in combination with SENSE has not yet been explored, and is the focus of this study. Using a SENSE-reduction factor of 2, DTI scans in eight healthy volunteers were carried out with regular- and high-resolution acquisition matrices. To further improve the stability of the SENSE reconstruction, a new coil-sensitivity estimation technique based on variational calculus and the principles of matrix regularization was applied. With SENSE, maps of the trace of the diffusion tensor and of fractional anisotropy (FA) had improved spatial resolution and less geometric distortion. Overall, the geometric distortions were substantially removed and a significant resolution enhancement was achieved with almost the same scan time as regular EPI. DTI was even possible without the use of quadrature body coil (QBC) reference scans. Geometry-factor-related noise enhancement was only discernible in maps generated with higher-resolution matrices. Error boundaries for residual fluctuations in SENSE reconstructions are discussed. Our results suggest that SENSE can be combined with DTI and may present an important adjunct for future neuroimaging applications of this technique. PMID- 12111941 TI - Image distortion correction in EPI: comparison of field mapping with point spread function mapping. AB - Echo-planar imaging (EPI) can provide rapid imaging by acquiring a complete k space data set in a single acquisition. However, this approach suffers from distortion effects in geometry and intensity, resulting in poor image quality. The distortions, caused primarily by field inhomogeneities, lead to intensity loss and voxel shifts, the latter of which are particularly severe in the phase encode direction. Two promising approaches to correct the distortion in EPI are field mapping and point spread function (PSF) mapping. The field mapping method measures the field distortions and translates these into voxel shifts, which can be used to assign image intensities to the correct voxel locations. The PSF approach uses acquisitions with additional phase-encoding gradients applied in the x, y, and/or z directions to map the 1D, 2D, or 3D PSF of each voxel. These PSFs encode the spatial information about the distortion and the overall distribution of intensities from a single voxel. The measured image is the convolution of the undistorted density and the PSF. Measuring the PSF allows the distortion in geometry and intensity to be corrected. This work compares the efficacy of these methods with equal time allowed for field mapping and PSF mapping. PMID- 12111942 TI - Motion correction using the k-space phase difference of orthogonal acquisitions. AB - Rigid body translations of an object in MRI create image artifacts along the phase-encode (PE) direction in standard 2DFT imaging. If two images are acquired with swapped PE direction, it is possible to determine and correct for arbitrary in-plane translational interview motions in both images directly from phase differences in the k-space acquisitions by solving a large system of linear equations. For example, if one assumes two N x N 2D acquisitions with in-plane translational interview motion, 4N unknown motions may corrupt the two images, but the phase difference at each point in k-space yields a system of N(2) equations in these 4N unknowns. If the acquisitions have orthogonal PE directions, this highly overdetermined system of equations can be solved to provide the motion records, which in turn can be used to correct the motion artifacts in each image. The theory of this orthogonal k-space phase difference (ORKPHAD) technique is described, and results are presented for synthetic and in vivo motion-corrupted data sets. In all cases, the data showed clear improvement of translation-induced artifacts. These methods do not require special pulse sequences and are theoretically generalizable to partial Fourier imaging and 3D acquisitions. PMID- 12111943 TI - Multishot 3D slice-select tailored RF pulses for MRI. AB - A multishot 3D slice-select tailored RF pulse method is presented for the excitation of slice profiles with arbitrary resolution. This method is derived from the linearity of the small tip angle approximation, allowing for the decomposition of small tip angle tailored RF pulses into separate excitations. The final image is created by complex summation of the images acquired from the individual excitations. This technique overcomes the limitation of requiring a long pulse to excite thin slices with adequate resolution. This has implications in applications including T*(2)-weighted functional MRI in brain regions corrupted by intravoxel dephasing artifacts due to susceptibility variations. Simulations, phantom experiments, and human brain images are presented. It is demonstrated that at most four shots of 40 ms pulse length are needed to excite a 5 mm-thick slice in the brain with reduced susceptibility artifacts at 3T. PMID- 12111944 TI - Mapping myocardial perfusion with an intravascular MR contrast agent: robustness of deconvolution methods at various blood flows. AB - Evaluation of quantitative parameters such as regional myocardial blood flow (rMBF), blood volume (rMBV), and mean transit time (rMTT) by MRI is gaining acceptance for clinical applications, but still lacks robust postprocessing methods for map generation. Moreover, robustness should be preserved over the full range of myocardial flows and volumes. Using experimental data from an isolated pig heart preparation, synthetic MR kinetics were generated and four deconvolution approaches were evaluated. These methods were then applied to the first-pass T(1) images of the isolated pig heart using an intravascular contrast agent and rMBF, rMBV and rMTT maps were generated. In both synthetic and experimental data, the fit between calculated and original data reached equally good results with the four techniques. rMBV was the only parameter estimated correctly in numerical experiments. Moreover, using the algebraic method ARMA, abnormal regions were well delineated on rMBV maps. At high flows, rMBF was underestimated at the experimental noise level. Finally, rMTT maps appeared noisy and highly unreliable, especially at high flows. In conclusion, over the myocardial flow range, i.e., 0-400 ml/min/100g, rMBF identification was biased in presence of noise, whereas rMBV was correctly identified. Thus, rMBV mapping could be a fast and robust way to detect abnormal myocardial regions. PMID- 12111945 TI - Independent component analysis of fMRI data in the complex domain. AB - In BOLD fMRI a series of MR images is acquired and examined for task-related amplitude changes. These functional changes are small, so it is important to maximize detection efficiency. Virtually all fMRI processing strategies utilize magnitude information and ignore the phase, resulting in an unnecessary loss of efficiency. As the optimum way to model the phase information is not clear, a flexible modeling technique is useful. To analyze complex data sets, independent component analysis (ICA), a data-driven approach, is proposed. In ICA, the data are modeled as spatially independent components multiplied by their respective time-courses. There are thus three possible approaches: 1) the time-courses can be complex-valued, 2) the images can be complex-valued, or 3) both the time courses and the images can be complex-valued. These analytic approaches are applied to data from a visual stimulation paradigm, and results from three complex analysis models are presented and compared with magnitude-only results. Using the criterion of the number of contiguous activated voxels at a given threshold, an average of 12-23% more voxels are detected by complex-valued ICA estimation at a threshold of /Z/ > 2.5. Additionally, preliminary results from the complex models reveal a phase modulation similar to the magnitude time-course in some voxels, and oppositely modulated in other voxels. PMID- 12111946 TI - New approach to gridding using regularization and estimation theory. AB - When sampling under time-varying gradients, data is acquired over a non-equally spaced grid in k-space. The most computationally efficient method of reconstruction is first to interpolate the data onto a Cartesian grid, enabling the subsequent use of the inverse fast Fourier transform (IFFT). The most commonly used interpolation technique is called gridding, and is comprised of four steps: precompensation, convolution with a Kaiser-Bessel window, IFFT, and postcompensation. Recently, the author introduced a new gridding method called Block Uniform ReSampling (BURS), which is both optimal and efficient. The interpolation coefficients are computed by solving a set of linear equations using singular value decomposition (SVD). BURS requires neither the pre- nor the postcompensation steps, and resamples onto an n x n grid rather than the 2n x 2n matrix required by conventional gridding. This significantly decreases the computational complexity. Several authors have reported that although the BURS algorithm is very accurate, it is also sensitive to noise. As a consequence, even in the presence of a low level of measurement noise, the resulting image is often highly contaminated with noise. In this work, the origin of the noise sensitivity is traced back to the potentially ill-posed matrix inversion performed by BURS. Two approaches to the solution are presented. The first uses regularization theory to stabilize the inversion process. The second formulates the interpolation as an estimation problem, and employs estimation theory for the solution. The new algorithm, called rBURS, contains a regularization parameter, which is used to trade off the accuracy of the result against the signal-to-noise ratio (SNR). The results of the new method are compared with those obtained using conventional gridding via simulations. For the SNR performance of conventional gridding, it is shown that the rBURS algorithm exhibits equal or better accuracy. This is achieved at a decreased computational cost compared to conventional gridding. PMID- 12111947 TI - Coupling and decoupling theory and its application to the MRI phased array. AB - In classical MRI phased-array design, optimal coil overlapping is used to minimize coupling between nearest-neighbor coils, and low input impedance preamplifiers are used to isolate the relatively weak coupling between non nearest neighbors. However, to make the complex sensitivities of phased-array coils sufficiently distinct in parallel spatially-encoded MRI, it is desirable to have no overlapping between coils. Also, if phased arrays are used as transmit coils in MRI, one can no longer rely on the low input impedance of the preamplifiers for decoupling. Here a coupling and decoupling theory is introduced to provide a better understanding of the relations between coupled and uncoupled signals in the MRI phased array, and to offer a new method for decoupling phased array coils without overlapping the nearest coil pairs. The new decoupling method is based on the assumption that any n-element phased array can be decoupled by a 2n-port interface system between phased array and preamplifiers. The detailed analysis and the experimental results show that a four-port interface can be used to decouple a two-element phased array. Furthermore, the four-port interfaces can serve as building blocks to construct a 2n-port decoupling interface. This new method allows one to place the coil elements anywhere that could optimize parallel spatial encoding without concern for coupling between the coils. The method can also be used for phased-array transmit coils. PMID- 12111949 TI - MRI of the lungs using hyperpolarized noble gases. AB - The nuclear spin polarization of the noble gas isotopes (3)He and (129)Xe can be increased using optical pumping methods by four to five orders of magnitude. This extraordinary gain in polarization translates directly into a gain in signal strength for MRI. The new technology of hyperpolarized (HP) gas MRI holds enormous potential for enhancing sensitivity and contrast in pulmonary imaging. This review outlines the physics underlying the optical pumping process, imaging strategies coping with the nonequilibrium polarization, and effects of the alveolar microstructure on relaxation and diffusion of the noble gases. It presents recent progress in HP gas MRI and applications ranging from MR microscopy of airspaces to imaging pulmonary function in patients and suggests potential directions for future developments. PMID- 12111950 TI - Toward direct mapping of neuronal activity: MRI detection of ultraweak, transient magnetic field changes. AB - A novel method based on selective detection of rapidly changing DeltaB(0) magnetic fields and suppression of slowly changing DeltaB(0) fields is presented. The ultimate goal of this work is to present a method that may allow detection of transient and subtle changes in B(0) in cortical tissue associated with electrical currents produced by neuronal activity. The method involves the detection of NMR phase changes that occur during a single-shot spin-echo (SE) echo-planar sequence (EPI) echo time. SE EPI effectively rephases all changes in B(0) that occur on a time scale longer than the echo time (TE) and amplifies all DeltaB(0) changes that occur during TE/2. The method was tested on a phantom that contains wires in which current can be modulated. The sensitivity and flexibility of the technique was demonstrated by modulation of the temporal position and duration of the stimuli-evoked transient magnetic field relative to the 180 RF pulse in the imaging sequence-requiring precise stimulus timing. Currently, with this method magnetic field changes as small as 2 x 10(-10) T (200 pT) and lasting for 40 msec can be detected. Implications for direct mapping of brain neuronal activity with MRI are discussed. PMID- 12111951 TI - 2D JPRESS of human prostates using an endorectal receiver coil. AB - A localized 2D J-resolved (JPRESS) MR spectroscopic sequence was evaluated in human prostates in vivo. Voxels of typically 2 ml were placed in the peripheral zone of the prostate. Eight healthy volunteers, three subjects with benign prostatic hyperplasia, and three patients with prostatic cancer were scanned on a 1.5T MR scanner, using a body coil for RF transmission and a pelvic phased-array coil combined with a disposable endorectal coil for signal reception. The total acquisition time for a 2D JPRESS spectrum was approximately 17 min. A major advantage of the endorectal 2D JPRESS was the ability to resolve the peaks of choline-containing compounds and those of spermine unequivocally. Spectral results clearly showed the biochemical changes in cancer and benign compared to healthy prostates, in conformity with ex vivo biochemical findings. The preliminary results suggest that the endorectal 2D JPRESS could be successfully implemented for the diagnostic examination of human prostates. . PMID- 12111952 TI - Automatic control of hyperthermic therapy based on real-time Fourier analysis of MR temperature maps. AB - Local hyperthermia is increasingly being used for therapeutic purposes, such as tumor ablation. Heat conduction and energy absorption in vivo during the hyperthermic procedure are largely unknown, thus making feedback temperature control highly desirable. Here, a general method for temperature control based on Fourier transformation (FT) of the bio-heat equation is presented, taking into account heat diffusion (D) and energy absorption (alpha) together with temperature distribution derived from rapid, continuous MR temperature mapping. The main advantages of the new method are: 1) the spatial distribution of heat deposition and conduction over the full region of interest (ROI) is taken into account, and 2) the high speed resulting from the use of fast FT (FFT) of temperature maps allows rapid feedback coupling. Initial tests based on MRI guided focused ultrasound (FUS) demonstrated that high-quality temperature regulation can be obtained even for erroneous values of D and alpha, so long as their relative error remained in the same range. Performance of the automated control procedure was validated ex vivo and in vivo on rabbit thigh using moderate FUS heating. During the procedure, the standard deviation (SD) of the temperature remained in the range of temperature noise obtained by MRI, indicative of the performance of the regulation algorithm. PMID- 12111953 TI - Pulmonary ventilation and perfusion scanning using hyperpolarized helium-3 MRI and arterial spin tagging in healthy normal subjects and in pulmonary embolism and orthotopic lung transplant patients. AB - Conventional nuclear ventilation/perfusion (V/Q) scanning is limited in spatial resolution and requires exposure to radioactivity. The acquisition of pulmonary V/Q images using MRI overcomes these difficulties. When inhaled, hyperpolarized helium-3 ((3)He) permits MRI of gas distribution. Magnetic labeling of blood (arterial spin-tagging (AST)) provides images of pulmonary perfusion. Three normal subjects, two patients who had undergone single lung transplantation for emphysema, and one subject with pulmonary embolism (PE), were imaged. (3)He distribution and blood perfusion appeared uniform in the normal subjects and throughout the lung allografts. Gas distribution and perfusion in the emphysematous lungs were non-uniform and paralleled radiographic abnormalities. AST imaging alone revealed a lower-lobe wedge-shaped perfusion defect in the patient with PE that corresponded to computed tomography (CT) imaging. Hyperpolarized (3)He gas is demonstrated to provide ventilation images of the lung. Blood perfusion information may be obtained during the same examination using the AST technique. The sequential application of these imaging methods provides a novel tool for studying V/Q relationships. PMID- 12111954 TI - Phase coherent averaging in magnetic resonance spectroscopy using interleaved navigator scans: compensation of motion artifacts and magnetic field instabilities. AB - The quality of spectra in (1)H magnetic resonance spectroscopy (MRS) is strongly affected by temporal signal instabilities during the acquisition. One reason for these instabilities are hardware imperfections, e.g., drifts of the main magnetic field in superconducting magnets. This is of special concern in high-field systems where the specification of the field stability is close to the spectral linewidth. A second major potential source of artifacts, particularly in clinical MRS, is patient motion. Using standard acquisition schemes of phase-cycled averaging of the individual acquisitions, long-term effects (field drifts) as well as changes on a shorter time scale (motion) can severely reduce spectral quality. The new technique for volume-selective MRS presented here is based on the additional interleaved acquisition of a navigator signal during the recovery time of the metabolite acquisition. It corrects for temporal signal instabilities by means of a deconvolution of the metabolite and the navigator signal. This leads to phase-corrected individual metabolite scans and upon summation to a phase-coherent averaging scheme. The interleaved navigator acquisition does not require any user interaction or supervision, while sequence efficiency is maintained. PMID- 12111955 TI - Characterization of anisotropy in high angular resolution diffusion-weighted MRI. AB - The methods of group theory are applied to the problem of characterizing the diffusion measured in high angular resolution MR experiments. This leads to a natural representation of the local diffusion in terms of spherical harmonics. In this representation, it is shown that isotropic diffusion, anisotropic diffusion from a single fiber, and anisotropic diffusion from multiple fiber directions fall into distinct and separable channels. This decomposition can be determined for any voxel without any prior information by a spherical harmonic transform, and for special cases the magnitude and orientation of the local diffusion may be determined. Moreover, non-diffusion-related asymmetries produced by experimental artifacts fall into channels distinct from the fiber channels, thereby allowing their separation and a subsequent reduction in noise from the reconstructed fibers. In the case of a single fiber, the method reduces identically to the standard diffusion tensor method. The method is applied to normal volunteer brain data collected with a stimulated echo spiral high angular resolution diffusion weighted (HARD) acquisition. PMID- 12111956 TI - A model of blood-brain barrier permeability to water: accounting for blood inflow and longitudinal relaxation effects. AB - A noninvasive technique for measuring the permeability of the blood-brain barrier (BBB) to water could help to evaluate changes in the functional integrity of the BBB that occur in different pathologies, such as multiple sclerosis or growth of brain tumor. Recently, Schwarzbauer et al. (Magn Reson Med 1997;37:769-777) proposed an MR method to measure this permeability based on the T(1) reductions induced by injecting various doses of paramagnetic contrast agent. However, this method may be difficult to implement in a clinical environment. Described here is a two-point technique, in which a spatially selective inversion is used to measure T(1) prior to and after injection of an intravascular contrast agent. Measurements made in the rat brain are compared to numerical simulations generated with a physiological model that accounts for blood flow and includes two different blood volumes: nonexchanging and exchanging blood volumes. Our results suggest that BBB permeability could be evaluated from the change in T(1) caused by the vascular contrast agent. This technique might provide an approach for monitoring changes in BBB permeability to water in clinical studies. PMID- 12111957 TI - Water exchange and inflow affect the accuracy of T1-GRE blood volume measurements: implications for the evaluation of tumor angiogenesis. AB - The goal of this study was to determine the degree to which vascular water exchange and blood flowing into an imaging slice affect the accuracy of blood volume measurements of brain and tumor tissue when using intravascular T(1) contrast agents. The study was performed using 2D and 3D gradient-echo imaging sequences, since these are two of the most commonly used MRI methods used to evaluate tissue blood volume fraction. Computer simulations were performed and measurements made in a rat 9L gliosarcoma brain tumor model. The computer simulations demonstrate that, with either water exchange or inflow effects alone, the dependence on the physiologic and imaging parameters can be well characterized and therefore potentially offset. In the exchange only case, the parametric dependence of 3D simulations suggest that the best accuracy is achieved with high flip angles, short TR, and low blood contrast agent concentrations. However, for a 2D GRE sequence which is influenced by both water exchange and inflow, the simulations predict that the error trend as a function of the imaging and physiologic parameters is unpredictable and therefore difficult to compensate. With both 2D and 3D GRE the measured blood volume data in rat brain and tumor tissue demonstrate tissue-specific trends, which reflect differences in the considered physiologic parameters. The experimental data strongly support the computer simulations and also indicate that minimization of the physiological effects by proper selection of imaging parameters, contrast concentration, and volume calculation methods is crucial for accurate assessment of absolute blood volume fraction. PMID- 12111958 TI - Dy-DTPA derivatives as relaxation agents for very high field MRI: the beneficial effect of slow water exchange on the transverse relaxivities. AB - Proton longitudinal and transverse relaxivities of Dy(DTPA)(2-) and Dy-DTPA bisamide derivatives (Dy(DTPA-BA): Dy-DTPA bisamide, Dy(DTPA-BEA): Dy-DTPA bisethylamide, Dy(DTPA-BnBA): Dy-DTPA bis-n-butylamide, and Dy(DTPA-BBMA): Dy DTPA bisbismethylamide) were analyzed between 0.47 T and 18.8 T. Curie longitudinal relaxation was clearly observed at magnetic fields larger than 2.4 T, but the longitudinal relaxivities are limited by the fast rotation of the complexes. Rotational correlation times were separately assessed by deuterium relaxometry of the diamagnetic deuterated lanthanum analogs. Transverse relaxivity, which depends on the square of the magnetic field and on the residence time of the coordinated water molecule (tau(M)), was more than 7.5 times larger at 18.8 T and 310 K for Dy(DTPA-BA) and Dy(DTPA-BEA) as compared to Dy(DTPA)(2-). This difference is mainly related to the slower water exchange of the bisamide complexes, as confirmed by the values of tau(M) measured by oxygen 17 relaxometry. Such Dy-complexes, characterized by relatively long tau(M) values (tauM310 larger than 100 ns but smaller than 1 micros), thus appear to be useful as negative T(2) (or transverse) contrast agents for high-field imaging. This was demonstrated by the spin-echo images of phantoms obtained at 4.7 T on samples containing Dy(DTPA)(2-) and Dy(DTPA-BEA). PMID- 12111959 TI - Theory of nonexponential NMR signal decay in liver with iron overload or superparamagnetic iron oxide particles. AB - A quantitative theory is proposed for the nonexponential NMR proton signal decay observed in liver with iron overload or superparamagnetic iron oxide particles. This effect occurs for Carr-Purcell-Meiboom-Gill (CPMG) sequences and is argued to be a direct consequence of the strong magnetic field inhomogeneities generated by the iron, rather than being due to tissue compartments. An approximate mathematical form is given for the signal decay, which is fit to experimental data for samples of rat liver with iron oxide particles, for samples of marmoset liver with hemosiderosis, and for in vivo human liver with hereditary hemochromatosis. The fitting parameters obtained are consistent with the pattern of iron deposition determined from histology. For the case of hereditary hemochromatosis, a good correlation is found between a parameter characterizing the nonexponential decay and the iron concentration. Implications for practical MR quantification of hepatic iron are discussed. PMID- 12111960 TI - Dynamic oxygen-enhanced MRI reflects diffusing capacity of the lung. AB - The purpose of this study was to demonstrate the feasibility of dynamic oxygen enhanced MRI in a clinical setting. We hypothesized that dynamic oxygen enhancement can reflect the regional diffusing capacity of the lung. Ten patients with pulmonary emphysema and seven healthy volunteers were examined with a respiratory-synchronized inversion recovery single-shot turbo spin-echo sequence (TR = 3200-5000 ms, TE = 16 ms, TI = 720 ms, ETS = 4 ms) following 100% oxygen inhalation, using a 1.5 T whole-body scanner. Maximum mean relative enhancement ratios calculated by averaging six defined regions of interest (ROIs) in both lungs were statistically compared between healthy volunteers and patients, and were correlated with diffusing lung capacity (%DL(CO)). In patients with pulmonary emphysema, maximum mean relative enhancement ratios were significantly decreased compared to those in healthy volunteers (P = 0.0008). Maximum mean relative enhancement ratio had excellent correlation with % DL(C0) (r(2) = 0.83). Dynamic oxygen-enhanced MRI may reflect the diffusing capacity of the lung; therefore, imaging of oxygen enhancement with MRI may provide maps of the diffusing capacity. PMID- 12111961 TI - Cerebral blood volume measurements by rapid contrast infusion and T2*-weighted echo planar MRI. AB - Cerebral blood volume (CBV) provides information complementary to that of cerebral blood flow in cerebral ischemia, tumors, and other conditions. We have developed an alternative theory and method for measuring CBV based on dynamic imaging by MRI or CT during a short contrast infusion. This method avoids several limitations of traditional approaches that involve waiting for steady state or measuring the area under the curve (AUC) during bolus contrast injection. Anesthetized dogs were studied by T2*-weighted echo planar imaging during gadolinium-DTPA infusions lasting 30-60 sec. CBV was calculated from the ratio of the signal changes in tissue and artery. Method responsiveness was compared to AUC measurements using the vasodilator acepromazine. The ratio of signal change in tissue to that in artery rapidly approached an asymptotic value even while the amount of contrast in artery continued to increase. Using 30-sec infusions, the mean (+/- SD) of CBV for control animals was 3.6 +/- 0.9 ml blood/100 g tissue in gray matter and 2.3 +/- 0.8 ml blood/100 g tissue in white matter (ratio = 1.6). Acepromazine increased CBV to 5.7 +/- 1.5 ml blood/100 g tissue in gray matter and 3.1 +/- 0.8 ml blood/100 g tissue in white matter (ratio = 2.0). AUC measurements after bolus injection yielded similar values for control animals but failed to demonstrate any change after acepromazine. It is possible to measure CBV using dynamic MRI or CT during 30-60-sec contrast infusions. This method may be more sensitive to changes in CBV than traditional AUC methods. PMID- 12111962 TI - Serial MRI evaluation of cardiac structure and function in mice after reperfused myocardial infarction. AB - This study evaluated the utility of cardiac MRI for assessing the impact of myocardial infarction (MI) on cardiac structure and function in mice following reperfused 1- or 2-hr occlusions of the left anterior descending coronary artery (LAD). When assessed 1 day after MI, the left ventricular ejection fraction (LVEF) had declined by more than half, and remained depressed for the duration of the study. Furthermore, MI initiated dramatic increases in both LV end-systolic volume (LVESV) and end-diastolic volume (LVEDV), with a greater than threefold increase in LVESV and a twofold increase in LVEDV by 4 weeks post-MI. Transmural LV wall thickening (WTh) analysis revealed that noninfarcted myocardium in the remote septal region exhibited an early deficit in contractile function after MI that transiently resolved by day 7, only to be followed by a late phase of dysfunction that became fully manifest by day 28 post-MI. In conclusion, MRI allows the serial assessment of cardiac structure and function after MI in mice, with a resolution adequate to document both regional and temporal changes. The application of these imaging techniques in transgenic and knock-out mice will greatly expedite research aimed at defining the functional roles of individual genes in the pathophysiology of LV remodeling (LVR) after reperfused MI. PMID- 12111963 TI - 3D MRA coronary axis determination using a minimum cost path approach. AB - A method is introduced to automatically find the coronary axis based on two or more user-defined points, even in the presence of a severe stenosis. The coronary axis is determined by finding a minimum cost path (MCP) in a feature image in which the tubular-like structures are enhanced. The results of the proposed method were compared with manually drawn central axes to estimate the accuracy. In 32 3D TFE-EPI acquisitions of patients and volunteers, 14 right coronary arteries (RCAs), 15 left anterior descending arteries (LADs), and eight left circumflex arteries (LCXs) were manually tracked twice by two operators to determine a reference axis and to assess the inter- and intra-user variability. On average, the maximum distance to the reference axis, based on only two user defined points, is less than 1.5 mm; the average distance is around 0.65 mm, which is less than the average in-plane resolution. The results of the method are comparable to those of the manual operators. PMID- 12111964 TI - Quantification of spinal cord atrophy from magnetic resonance images via a B spline active surface model. AB - A method is presented that aims at segmenting and measuring the surface of the spinal cord from MR images in order to detect and quantify atrophy. A semiautomatic segmentation with very little intervention from an operator is proposed. It is based on the optimization of a B-spline active surface. The method allows for the computation of orthogonal cross-sections at any level along the cord, from which measurements are derived, such as cross-sectional area or curvature. An evaluation of the accuracy and reproducibility of the method is presented. PMID- 12111965 TI - Two-dimensional spatially-selective RF excitation pulses in echo-planar imaging. AB - Two-dimensional spatially-selective RF (2DRF) excitation pulses were developed for single-shot echo-planar imaging (EPI) with reduced field of view (FOV) in the phase-encoding direction. The decreased number of k-space lines significantly shortens the length of the EPI echo train. Thus, both gradient-echo and spin-echo 2DRF-EPI images of the human brain at 2.0 T exhibit markedly reduced susceptibility artifacts in regions close to major air cavities. Based on a blipped-planar trajectory, implementation of a typical 2DRF pulse resulted in a 26-ms pulse duration, a 5-mm section thickness, a 40-mm FOV along the phase encoding direction, and a 200-mm distance of the unavoidable side excitations from the center of the FOV. For the above conditions and at 2 x 2 mm(2) resolution, 2DRF-EPI yielded an echo train length of only 21 ms, as opposed to 102 ms for conventional EPI. This gain in time may be used to achieve higher spatial resolution. For example, spin-echo 2DRF-EPI of a 40-mm FOV at 1 x 1 mm(2) resolution led to an echo train of 66 ms. Although the current implementation still lacks user-friendliness, 2DRF pulses are likely to become a useful addition to the arsenal of advanced MRI tools. . PMID- 12111966 TI - T1 relaxation time at 0.2 Tesla for monitoring regional hyperthermia: feasibility study in muscle and adipose tissue. AB - The purpose of this study was to characterize T(1), particularly in the hyperthermia temperature range (ca. 37-44 degrees C), in order to control regional hyperthermia with MR monitoring using 0.2 Tesla, and to improve T(1) mapping. A single-slice and a new multislice "T One by Multiple Read-Out Pulses" (TOMROP) pulse sequence were used for fast T(1) mapping in a clinical MRI hyperthermia hybrid system. Temporal stability, temperature sensitivity, and reversibility of T(1) were investigated in a polyamidacryl gel phantom and in samples of muscle and adipose tissues from turkey and pig, and verified in patients. In the gel phantom a high linear correlation between T(1) and temperature (R(2) = 0.97) was observed. In muscle and adipose tissue, T(1) and temperature had a linear relationship below a breakpoint of 43 degrees C. Above this breakpoint muscle tissue showed irreversible tissue changes; these effects were not visible in adipose tissue. The ex vivo results were confirmed in vivo under clinical conditions. T(1) mapping allows the characterization of hyperthermia-related tissue response in healthy tissue. T(1), in combination with fast mapping, is suitable for controlling regional hyperthermia at 0.2 T within the hybrid system. PMID- 12111967 TI - Generalized autocalibrating partially parallel acquisitions (GRAPPA). AB - In this study, a novel partially parallel acquisition (PPA) method is presented which can be used to accelerate image acquisition using an RF coil array for spatial encoding. This technique, GeneRalized Autocalibrating Partially Parallel Acquisitions (GRAPPA) is an extension of both the PILS and VD-AUTO-SMASH reconstruction techniques. As in those previous methods, a detailed, highly accurate RF field map is not needed prior to reconstruction in GRAPPA. This information is obtained from several k-space lines which are acquired in addition to the normal image acquisition. As in PILS, the GRAPPA reconstruction algorithm provides unaliased images from each component coil prior to image combination. This results in even higher SNR and better image quality since the steps of image reconstruction and image combination are performed in separate steps. After introducing the GRAPPA technique, primary focus is given to issues related to the practical implementation of GRAPPA, including the reconstruction algorithm as well as analysis of SNR in the resulting images. Finally, in vivo GRAPPA images are shown which demonstrate the utility of the technique. PMID- 12111968 TI - Noise and motion correction in dynamic contrast-enhanced MRI for analysis of atherosclerotic lesions. AB - Dynamic contrast-enhanced MRI of atherosclerotic vessels after contrast agent injection may provide unique information regarding lesion structure and vulnerability. The high-resolution images necessary for viewing lesion substructures, however, are often corrupted by patient motion and low signal-to noise ratios, making pixel-level analyses difficult. This article presents a postprocessing method that enables pixel-level analysis of dynamic images by eliminating motion and enhancing image quality. Noise and motion correction are performed using optimal statistical methods under the assumption that noise and contrast agent dynamics are random processes. The method is demonstrated and validated on dynamic images of atherosclerotic plaques in human carotid arteries. PMID- 12111969 TI - Design of a SENSE-optimized high-sensitivity MRI receive coil for brain imaging. AB - An 8-channel receive-only detector array was developed for SENSE MRI of human brain. The coil geometry was based on a gapped element design and used ultra-high impedance preamplifiers for mutual decoupling of the elements. Computer simulations of the electric and magnetic fields showed that excellent signal-to noise ratio (SNR) and SENSE performance could be achieved by placing the coil elements close to the head and maintaining a substantial gap between the elements. Measurements with a 1.5 T prototype coil showed a 2.7-fold improvement of the SNR averaged over the brain compared to a conventional quadrature birdcage receive coil and an average geometrical noise amplification factor (g-value) of 1.06 and 1.38 for rate-2 and rate-3 SENSE, respectively. PMID- 12111970 TI - Direct reconstruction of MR images from data acquired on a non-Cartesian grid using an equal-phase-line algorithm. AB - The equal-phase line (EPL) algorithm is proposed as a means of allowing rapid Fourier transform (FT) reconstruction of MR image data acquired on a non Cartesian grid. The pixels on the image are grouped according to their positions. The pixels in a group have the same phase in the complex exponential function exp[j2pi(xu + yv)] and receive the same contribution from a data point. Each group is related to an EPL in the image space. The contribution of a data point can then be distributed to the pixels along the EPLs. The described EPL algorithm enables a decrease of the reconstruction time to about 40% of the direct FT (DrFT) for the non-Cartesian data. A numerical phantom and two sets of in vivo spiral data were used to investigate an optimal number of the EPLs and to measure the reconstruction time. The EPL algorithm runs nearly as fast as the look-up table (LUT) method (Dale et al. IEEE Trans Med Imaging 2001;20:207-217), but it does not require a large memory to store the coefficients in advance, as is required in the LUT method. Thus, the EPL algorithm can be used to reconstruct images up to 512 x 512 pixels in size in a PC of limited memory, and may be more conveniently applied to a multiprocessor system. PMID- 12111971 TI - Alterations in carbohydrate and fatty acid levels of Lymantria dispar larvae caused by a microsporidian infection and potential adverse effects on a co occurring endoparasitoid, Glyptapanteles liparidis. AB - Infection of Lymantria dispar host larvae by the entomopathogenic microsporidium Vairimorpha sp. has a negative impact on the performance of the endoparasitic braconid Glyptapanteles liparidis. To investigate possible causes for this effect, we studied to what extent nutritional host suitability is altered by the microsporidium. Therefore, we analyzed carbohydrates and fatty acids in host larvae after Vairimorpha infection and/or parasitism by G. liparidis. Trehalose levels were significantly reduced in the hemolymph of infected hosts. After day five post infection, it was detected only in traces. Four to six days later, the glycogen resources were depleted in infected larvae. Parasitism by G. liparidis, on the other hand, led to increased hemolymph trehalose levels during the early endoparasitic phase but to a significant decrease at the end of its larval development. No effect of parasitism on the glycogen content was ascertained. Hemolymph levels of the fatty acids analyzed, such as palmitic, stearic, oleic, linoleic, and linolenic acid, were significantly reduced in microsporidia infected L. dispar. Vairimorpha sp. develops as an intracellular parasite in the fat body of the host larva and synthesis of trehalose and fatty acids may be disturbed. Moreover, microsporidia may also harness metabolites or energy produced by host cells. We conclude that the microsporidia-induced decrease in hemolymph carbohydrates and fatty acids adversely affects growth and development of parasitoid larvae. PMID- 12111972 TI - Content and composition of phosphoglycerols and neutral lipids at different developmental stages of the eggs of the codling moth, Cydia pomonella (Lepidoptera: Tortricidae). AB - Phosphoglycerol, triacylglycerol, diacylglycerol, and free fatty acid content was studied in eggs of the codling moth Cydia pomonella at the white, red ring, and black head developmental stages. The composition of total phosphoglycerols and of the three classes of neutral lipids was also analyzed. The highest total lipid content was found in eggs at the white stage, the amount decreasing during development mainly as a result of a diminution in the quantity of phosphoglycerols, which account for approximately 50% of total content at all stages of egg development. The amount of triacylglycerols and free fatty acids changes significantly during development, whereas only minor changes were found in diacyglycerol levels. The total phosphoglycerol acyl composition of eggs at the white and red ring stages is similar, whereas differences are evident at the black head stage of development. Triacylglycerols and free fatty acids are enriched in saturated fatty acids in all analyzed stages. The acyl profile of diacylglycerols is different at each stage. The unsaturation index decreases in diacylglycerols and free fatty acids as a function of egg development. The results of the present paper suggest that triacylglycerols may constitute an important source of energy during the final period of egg development while phosphoglycerols may function as fuel during the beginning. Phosphoglycerols could be precursors for the triacylglycerol biosynthesis that takes place between white and red ring stages. PMID- 12111973 TI - Activation of the Sarcophaga lectin gene promoter in transgenic Drosophila. AB - We established transgenic Drosophila strains in which the lacZ gene was expressed under the control of the 5'-upstream regulatory region of the Sarcophaga lectin gene promoter (3.1 kbp). The reporter gene was expressed in the fat bodies of the transgenic larvae when they were immunized by body pricking or treatment with Escherichia coli, which was the same as the Sarcophaga lectin gene expression in Sarcophaga larvae. However, the same reporter gene was found to be expressed constitutively in the digestive tracts of the transgenic larvae even without immunization. PMID- 12111974 TI - Juvenile hormone stimulated tyrosine kinase-mediated protein phosphorylation in the CNS of the silk worm, Bombyx mori. AB - In vitro studies with the larval CNS of the silkworm, Bombyx mori revealed the phosphorylation of a 48-kDa protein, which was not dependent on cyclic nucleotides. Studies also revealed modest phosphorylation of this protein by a calcium-dependent but calmodulin-independent mechanism. However, phosphorylation of this protein was greatly enhanced in the presence of juvenile hormone (JH) I by a calcium-independent mechanism. This stimulatory effect of JH was seen in both homogenates as well as in intact CNS of Bombyx. Immunoblotting studies revealed the cross-reaction of this 48-kDa protein with phosphotyrosine monoclonal antibody and the phosphorylation of this protein was inhibited by genistein. This study suggests that the 48-kDa protein is a substrate for tyrosine kinase. The phosphorylation of this protein was also observed in other larval tissues such as salivary gland, fat body, and epidermis of Bombyx. PMID- 12111975 TI - Purification and characterization of nucleoside diphosphate kinase from the brain of Bombyx mori. AB - Nucleoside diphosphate kinase in the brain of Bombyx mori was purified by ammonium sulfate fractionation, and a sequence of chromatographies on DEAE Cellulofine, hydroxyapatite, Mono-S, and Mono-Q column. The purified enzyme preparation was found to be electrophoretically homogeneous on SDS-PAGE, and its molecular mass was determined to be 18 kDa. The purified protein was digested and the amino acid sequences of resulting peptides were determined. The enzyme showed high similarity to the amino acid sequences of the Drosophila NDP kinase. The enzyme showed NDP kinase activity and mediated the phosphorylation of myelin basic protein. Gel filtration and Hill plot analysis indicate that the purified NDP kinase forms a tetramer and shows little interaction among substrates. Dephosphorylation of NDP kinase by bacterial alkaline phosphatase increased NDP kinase activity. This result indicates that phosphorylation of NDP kinase represses NDP kinase activity. PMID- 12111976 TI - Surgery:...an enabling profession? PMID- 12111977 TI - Fabricating autologous tissue to engineer artificial nerve. AB - This study reports on the successful fabrication of artificial nerves with tissue engineering methods. Schwann cells were cultured for 2 weeks, seeded on polyglactin 910 scaffolds, and biomembrane-coated with rat-tail glue and laminin. Observation of the scaffolds' adsorptivity to Schwann cells, and of the growth and migration of Schwann cells, was made using a light microscope, and by scanning and transmission electron microscopy. The Schwann cells were able to migrate and proliferate on the polyglactin 910 fiber. Schwann cells were well distributed, and formed a Bungner band on which the Schwann cells produced more matrices. Schwann cells on the biomembrane also grew well. We investigated the role of the tissue engineering conduit guide in vivo, using an established rabbit peripheral nerve regeneration model. At 8 weeks, axonal regeneration was observed in the distal nerve stump. Adult Schwann cells can be produced on the coated fiber and the biomembrane. Three-dimensional scaffolds with Schwann cells had the basic characteristics of the artificial nerve. These findings will provide a practical method for fabricating tissue-engineered artificial nerve to repair long nerve defects. PMID- 12111978 TI - Nerve regeneration through a healthy peripheral nerve trunk as a nerve conduit: a preliminary study of a new concept in peripheral nerve surgery. AB - The popularity of nerve conduits has increased recently due to the need for alternative nerve reconstruction techniques, obviating the harvest of nerve grafts. Based on ideas suggesting nerve tissue itself, which was the most physiologic environment for nerve regeneration, a study using 40 sciatic nerves of 20 rats was performed. The proximal stumps of transected peroneal nerves were sutured to the lateral wall of healthy tibial nerve trunks after removal of the epineurium. Twelve weeks later, tissue samples were taken from the anastomosis sites and from the levels above and below these sites. Configurations of fascicles were evaluated, and numbers of fibers were estimated. It was observed that the fibers of peroneal nerves regenerated into and through the tibial nerve trunk distally. This suggested that active regenerating fibers of a proximal stump of a nerve could regenerate and progress as a fascicular unit in optimum condition at the trunk of another healthy nerve. This preliminary study should stimulate further studies based on this new concept: that a nerve trunk can serve as the host for the regenerating fibers of another nerve. PMID- 12111979 TI - Insulin growth factor-1 decreases muscle atrophy following denervation. AB - Despite modern microsurgical techniques for nerve repair, functional outcome following proximal injury is often unsatisfactory because irreversible muscle atrophy may develop before reinnervation occurs. Because insulin growth factor-1 (IGF-1) has been shown to improve muscle regeneration after injury, and may have a role in muscle preservation following denervation, the purpose of this investigation was to evaluate the histological, immunohistochemical, and electrophysiological differences between normal, denervated, and IGF-1-injected denervated muscle over an 8-week period. Denervated mice gastrocnemius muscles demonstrated a decrease in muscle weight, a decrease in myofiber diameter, an absence of muscle regeneration, an early increase in the number of neuromuscular junctions (NMJs), and a decrease in fast-twitch and maximum tetanic strength as compared to normal muscle up to 8 weeks following denervation. IGF-1-injected denervated muscle, on the other hand, sustained muscle diameter and muscle weight, maintained a smaller number of NMJs, and relatively sustained fast-twitch and maximum tetanic strength as compared to normal muscle over 8 weeks. These data suggest that IGF-1 may help prevent muscle atrophy and secondary functional compromise after denervation. PMID- 12111980 TI - Effect of isovolemic hemodilution with 3% albumin on thrombus formation at venous microanastomosis in rats. AB - The present investigation was undertaken to evaluate the effect of isovolemic hemodilution with 3% albumin per se on the prevention of microvascular venous thrombosis, using an experimental model in rats mimicking the thrombotic mechanisms similar to those in clinical situations. Male Wistar rats were allocated into two groups of 15 rats each: group 1, the nonhemodiluted group, consisted of animals submitted to microsurgical procedure to induce thrombosis without previous hemodilution; group 2, the hemodiluted group, consisted of animals submitted to microsurgical procedure to induce thrombosis with previous hemodilution. The histopathological examination clearly showed that the nonhemodiluted group presented well-defined venous thrombosis in the anastomoses in 9 animals, totally occlusive in 8, and mural in 1. In contrast, in the hemodiluted group none of the anastomoses presented venous thrombosis, except for the presence of nonadherent blood clots. The clinical analysis of the anastomoses' patency, as evaluated by the empty-full test, showed a significant reduction in rate of venous occlusion in the previously hemodiluted animals as compared with the nonhemodiluted animals 20 min after microsurgical anastomosis. Forty-eight hours after anastomosis, the rate of venous occlusion was 66.7% in the nonhemodiluted group and 33.3% in the hemodiluted group; even though no significant difference could be detected during this time, the patency of anastomoses tended to be greater in the hemodiluted group than that of controls. The rate of spontaneous recanalization after 48 h was identical in both groups (16.6%) in comparison with the rates observed 20 min after the surgical procedure. In summary, moderate isovolemic hemodilution with 3% albumin was effective in reducing the rate of venous microanastomosis occlusion in rats 20 min after the surgical procedure, tending to maintain the anastomoses' patency after 48 h. In addition, 48 h after the microsurgical procedure, the rate of venous thrombosis as evaluated microscopically was significantly lower in the hemodiluted animals as compared with nonhemodiluted control animals. This points out that hemodilution with albumin could be a safe and adequate procedure to prevent thrombus formation at venous microanastomosis. PMID- 12111981 TI - Improved energetic recovery of skeletal muscle in response to ischemia and reperfusion injury followed by in vivo 31P-magnetic resonance spectroscopy. AB - It is of great clinical interest to improve postischemic tissue recovery during microsurgical transfers. The effect of singlet oxygen energy (SOE) as photon illumination at 634 nm on rat skeletal muscle during ischemia and postischemic reperfusion was investigated noninvasively and continuously by in vivo (31)P magnetic resonance spectroscopy ((31)P-MRS). A model of pedicled rat rectus femoris muscle was used, where phosphorous metabolites were followed before onset of ischemia (control), after 4 h of ischemia, and after 1 h of reperfusion. Two groups were studied: one control group (n = 10), and one SOE-treated group (n = 10). Blood perfusion was measured by laser Doppler flowmetry (LDF) during the study. After 4 h of ischemia, ATP levels were 72% and 51% of normal control values in the illuminated group and the control group, respectively (P < 0.05). After 1 h of postischemic reperfusion, phosphocreatine (PCr) recovered to 79% and adenosine triphosphate (ATP) to 71% in the illuminated group, whereas in the control group, the recovery was 57% and 51%, respectively (P < 0.05). It is concluded that singlet oxygen energy has a beneficial effect on the energy state of skeletal muscle during ischemia and postischemic reperfusion. PMID- 12111982 TI - Skeletal fixation in digital replantation. AB - Although the primary objective of replantation is revascularization and ultimately viability of the amputated digit(s), skeletal stabilization is an important cornerstone of the composite repair and reconstructive process. If performed rapidly and securely, anatomic (or near anatomic) fracture reduction and fixation may contribute profoundly to the protection of the revascularization and the repair or reconstruction of nerves, tendons, and integument; reliable fracture healing; functional restoration; and final outcome. Conversely, less than anatomic (or near anatomic) reduction or unreliable and insecure fixation may deter successful early revascularization and, later, good function. This article reviews the various methods of fracture stabilization that may be employed, and their advantages and disadvantages. We believe that anatomic (or near anatomic) fracture reduction, reliable and stable fracture fixation, minimal additional dissection, and early active range-of-motion exercises will have a substantial effect on both viability and functional outcome in digital replantation. PMID- 12111985 TI - Disability versus futility in rationing health care services: defining medical futility based on permanent unconsciousness--PVS, coma, and anencephaly. AB - All societies must ration health care services in the face of unlimited demand. The concept of medical futility may appear to be an uncontroversial means by which to ration services. Depending upon how it is applied, however, limiting services based on alleged medical futility may violate prohibitions against disability-based discrimination. In particular, use of medical futility to require removal of life-sustaining interventions has been held to violate the Americans with Disabilities Act. The ADA protects both people with disabilities who are conscious and people in unconscious states, such as permanent vegetative state (PVS), coma, and anencephaly. Ultimately, as the number of people in permanently unconscious states increases, our society will have to recognize that consciousness is an essential characteristic defining human beings and determining whether a legal right to unlimited life-sustaining intervention should apply. This article proposes to define medical futility to preclude life sustaining interventions after a stated period of permanent unconsciousness and to redefine the end of life consistently as neocortical death. PMID- 12111987 TI - Legal implications for failure to comply with advance directives: an examination of the incompetent individual's right to refuse life-sustaining medical treatment. AB - Life-sustaining medical technology in the past century has created a growing body of case law and legislation recognizing the incompetent individual's right to make his or her own end-of-life decisions. This article focuses on California's leadership in the area of these specific end-of-life issues: specifically, exploring the right of an incompetent individual to refuse life-sustaining medical treatment. The article examines advance directives along with various judicial decision-making standards for incompetent individuals and explores the sociobehavioral and legal rationale for compliance with incompetent individual's rights to make end-of-life decisions. Finally this article concludes (i) that advance directives allow competent individuals to state the medical treatment they would prefer in the event they should later become incompetent and (ii) that when advance directives are properly executed in a detailed manner, under laws currently in effect in some jurisdictions, the preferences stated in the directive bind health care providers. PMID- 12111986 TI - The role of proxies in treatment decisions: evaluating functional capacity to consent to end-of-life treatments within a family context. AB - Psychology as a profession has entered the arena of palliative and hospice care later in the process than other health care professions. Through the use of Familial Advance Planning Evaluations (FAPEs), however, psychologists can assist individuals and families in facing end-of-life transitions in important ways. Hospice and palliative care philosophy treats the patient and family as the unit of care. End-of-life decision-making is therefore a family matter as well as a normative developmental transition. Yet, little is known about the decision making process. This paper reviews the literature regarding informed consent, advance care planning, and proxy decision-making and outlines a theoretical model for familial decision-making. Previous models of end-of-life capacity evaluations and family assessments are presented and serve as the basis for a comprehensive assessment of familial decision-making at the end of life. Functional capacity evaluations of individuals at the end of life regarding decisions about life sustaining medical treatments enable both the individual patient and one identified proxy from his or her family to discuss important issues families may face during medical crises at the end of life. The information gleaned from such evaluations has the potential to assist psychologists and other professionals in designing family-specific interventions to reduce caregiving distress, improve quality of life for dying patients, and ease the transition to bereavement for caregivers. PMID- 12111988 TI - End-of-life care in nursing homes: the interface of policy, research, and practice. AB - Much attention has begun to focus on the quality of care for persons near the end of life. Palliative care, especially through hospice, has generated much discussion as possibly the most holistic care available. Consideration of how chronically ill older adults receive such care as a result of public policy can benefit from adopting a multidimensional perspective. This paper adopts Bronfenbrenner's ecological model to understand current end-of-life care for nursing home residents, followed by consideration of how each of these dimensions or levels of influence can be used to foster both research agendas and policy reforms to improve end-of-life care of nursing home residents. Specifically, the benefits of considering the influence of such policy initiatives as the Medicare hospice benefit and the Patient Self-Determination Act (PSDA) on end-of-life care in nursing homes and the means through which policy can be informed by clinical research is emphasized. PMID- 12111989 TI - Reinterpreting the controlled substances act: predictions for the effect on pain relief. AB - On November 6, 2001 U.S. Attorney General Ashcroft reinterpreted the Controlled Substances Act in a manner consistent with the Pain Relief Promotion Act, a bill introduced in the last Congress. This reinterpretation criminalizes actions related to the provision of controlled substances (e.g. opioids, barbiturates) in the event that a patient dies and external observers believe the intent of providing the medication was to hasten the patient's death. This paper analyzes the arguments regarding the need for reinterpretation and the potential consequences of such an action on pain relief. The conclusion is that the weight of the data clearly leads to the prediction that this reinterpretation of the Controlled Substances Act will lead physicians to be less likely to provide adequate types and doses of pain medication to patients who may die. PMID- 12111990 TI - Characterization of the nuclear matrix proteins in a transgenic mouse model for prostate cancer. AB - The nuclear matrix (NM) contains a number of proteins that have been found to be associated with transformation. We have previously identified changes in the NM associated with prostate cancer. In this study, we examine the molecular changes that are associated with prostate cancer development in transgenic adenocarcinoma of mouse prostate (TRAMP) model by studying the differences in the NM proteins (NMPs). We collected prostates from the TRAMP males at six critical time points: 6 weeks (puberty), 11 and 19 weeks (development of mild hyperplasia), 25 weeks (development of severe hyperplasia), 31 and 37 weeks (development of neoplasia). The nuclear matrices from the prostates collected at these time points were then isolated and the NMPs were characterized by high-resolution two-dimensional gel electrophoresis. We found three NMPs (E1A, E1B, and E1C) that were present in the 6-week-old prostate and two NMPs (E2A and E2B) that were present in the 11-week old prostate. These NMPs were absent in the 31- and 37-week-old prostate. We also found five NMPs (E3A-E3E) that were present in the 31-week-old prostate, but absent in the earlier time points. In addition, three NMPs (Le1, Le2, Le3) were present at higher expression in the 6-, 11-, 19-, and 25-weeks old TRAMP prostates, but they were expressed lower during the development of neoplasia at 31- and 37-weeks old. Identification of these NMPs permits the development of novel markers that can characterize various stages of prostate cancer development as well as potentially therapeutic targets. PMID- 12111991 TI - Nuclear matrix proteins as biomarkers in prostate cancer. AB - The nuclear matrix (NM) is the structural framework of the nucleus that consists of the peripheral lamins and pore complexes, an internal ribonucleic protein network, and residual nucleoli. The NM contains proteins that contribute to the preservation of nuclear shape and its organization. These protein components better known as the NM proteins have been demonstrated to be tissue specific, and are altered in many cancers, including prostate cancer. Alterations in nuclear morphology are hallmarks of cancer and are believed to be associated with changes in NM protein composition. Prostate cancer is the most frequently diagnosed cancer in American men and many investigators have identified unique NM proteins that appear to be specific for this disease. These NM protein changes are associated with the development of prostate cancer, as well as in some cases being indicative of cancer stage. Identification of these NM proteins specific for prostate cancer provides an insight to understanding the molecular changes associated with this disease. This article reviews the role of NM proteins as tumor biomarkers in prostate cancer and the potential application of these proteins as therapeutic targets in the treatment of this disease. PMID- 12111992 TI - Helix 12 in the human estrogen receptor (hER) is essential for the hER function by overcoming nucleosome repression in yeast. AB - When exogenous human estrogen receptor (hER) binds with estrogen, it can activate transcription of target genes in yeast cells. The estrogen dose-response expression patterns in yeast are very similar to those in human cells. This implies that hER may function in yeast cells via mechanisms similar to those in human cells. In this study, Saccharomyces cerevisiae was used to dissect mechanisms of hER-activated transcription in yeast. The hER contains two transcription activation domains: ER-AF-1 and ER-AF-2 (LBD or HBD). In both human and wild-type yeast cells, hER must bind with estrogen in order to activate transcription. In those cells, ER-AF-2 is independently active upon hormone binding, but ER-AF-1 by itself is inactive. In a mutagenesis screen, we found a mutant strain in which the ER-AF-1 was independently active. It was determined that this mutant strain carried a Tup1 mutation. More interestingly, a small hER fragment ER-AF-0, containing neither ER-AF-1 nor ER-AF-2, was also fully active in the DeltaTup1 cells. This suggests that in this strain, hormone binding is not required for transcription activation by hER. It is known that the Tup1/Ssn6 complex plays an important role in general transcription repression by protecting histone acetylation sites thus stabilizing nucleosomes. In the DeltaTup1 cells, nucleosomes are known to be unstable because histones can be easily accessed by acetylase and cause nucleosome disassociation. Two point mutations in helix 12 (H12) in ER-AF-2, which abolished hER function in human cells, also completely abolished hER function in the wild-type yeast cells. This suggested that H12 is essential for hER transcription activation function. However, hER with the H12 mutation is able to activate transcription in DeltaTup1 cells. This indicates that the normal function of H12 is required for transcription activation by hER only if nucleosomes are not acetylated and are therefore stable. The results of this work suggest that there is a close relationship between hER function and nucleosome remodeling. It also provides insight about H12 activity and its functional relationship with other domains in hER. We propose here that H12 is essential for hER function by recruiting strong nucleosome remodeling proteins to the promoter region thus overcoming nucleosome repression. PMID- 12111994 TI - Effects of estrogen on collagen synthesis by cultured human osteoblasts depend on the rate of cellular differentiation. AB - Estrogen is known to act on osteoblasts according to their stage of differentiation and estrogen receptor (ER) isoform expression. The aim of this study was to determine when type I collagen (COL1) synthesis by cultured low passage, human bone-derived osteoblasts (hOBs) is upregulated in response to estrogen. Cell lines from female donors aged 1 and 66 years were cultured for 11 days on collagen in growth medium supplemented with human serum, hydrocortisone, and beta-glycerophosphate. Young-donor hOBs grew more quickly than old-donor hOBs and did not mineralize. Old-donor hOBs formed mineralized nodules 5 days after reaching confluence. Changes in mRNA levels with time for ERs, type I collagen, and alkaline phosphatase reflected the faster differentiation of the old-donor cells. The ERbeta/ERalpha ratio fell threefold in young-donor hOBs but rose 300 fold in old-donor hOBs. Increased ERbeta/ERalpha ratios prevented ligand dependent downregulation of ERalpha transcription, resulting in reduced proliferation in old-donor hOBs. Upregulation of COL1 mRNA expression in response to estrogen was confined to intermediate stages of differentiation, resulting in significant increases in COL1 mRNA by estradiol only in young-donor cells. Since the young and old-donor hOBs were cultured under identical conditions, our results indicate that the response of hOBs to estrogen is largely dependent on intracellular mechanisms that control the timing of cellular differentiation. PMID- 12111993 TI - Chronic pulsatile shear stress impacts synthesis of proteoglycans by endothelial cells: effect on platelet aggregation and coagulation. AB - Endothelial-derived proteoglycans are important regulators of the coagulation pathway in vivo and our primary objective of this study was to determine whether chronic shear stress affected the synthesis, release, and activity of proteoglycans from bovine aortic endothelial cells (BAEC). BAEC were cultured under shear and proteoglycans were purified from BAEC conditioned media and analyzed using both anionic exchange and size exclusion chromatography. The overall amount of proteoglycans produced per cell was significantly greater for the high shear-treated samples compared to the low shear-treated samples indicating that the shear magnitude did impact cell responsiveness. While overall size and composition of the proteoglycans and glycosaminoglycan (GAG) side chains were not altered by shear, the relative proportion of the high and low molecular weight species was inversely related to shear and differed significantly from that found under static tissue culture conditions. Moreover, a unique proteoglycan peak was identified from low shear stress (5 +/- 2 dynes/cm(2)) conditioned media when compared to high shear conditions (23 +/- 8 dynes/cm(2)) via anionic exchange chromatography, suggesting that subtle changes in the GAG structures may impact activity of these molecules. In order to characterize whether these changes impacted proteoglycan function, we studied the effects of shear specific proteoglycans on the inhibition of thrombin-induced human platelet aggregation as well as on platelet-fibrin clot dynamics. Proteoglycans from high shear-treated samples were less effective inhibitors of both platelet aggregation and blood coagulation inhibition than proteoglycans from low shear-treated samples and both were less effective than proteoglycans isolated from static tissue culture samples. However, due to changes in the overall proteoglycan synthesis and release rate, the high and low shear-treated sample had essentially identical effects on these activities, suggesting that the cells were able to compensate for stress-induced proteoglycan changes. Our data suggests that shear stress, by altering proteoglycan synthesis and fine structure, may play a role in maintaining vascular hemodynamics and hemostasis. PMID- 12111995 TI - Contribution of HDL-apolipoproteins to the inhibition of low density lipoprotein oxidation and lipid accumulation in macrophages. AB - High-density lipoprotein (HDL) is known as a protective factor against atherosclerosis. However, whether HDL-apolipoproteins (apo-HDL) contribute to the protection in arterial cells remains unclear. The localization patterns of human apolipoproteins in atherosclerotic arteries were determined using immunohistochemical examination. The results indicate that several apolipoproteins are retained in component cells of the coronary artery walls. To elucidate the possible roles of apo-HDL in the protection of atherosclerotic lesion formation, we investigated the effects of apo-HDL on the formation of conjugated diene (CD) in a cell-free system and thiobarbituric acid-reactive substances (TBARS) in the medium of a macrophage-mediated LDL oxidation system. The results showed that apo-HDL significantly exerted an inhibitory effect on LDL lipid oxidation in vitro. In addition, apo-HDL decreased cholesterol influx but enhanced cholesterol efflux from J774 macrophages in a dose-dependent manner. These results are consistent with the notion that there is reduced intracellular lipid accumulation in apo-HDL treated macrophages. These data provide a direct evidence for apo-HDL in protecting LDL from oxidative modification and in reducing the accumulation of cholesterol and lipid droplets by J774 macrophages. PMID- 12111996 TI - Differential involvement of protein kinase C in basal versus acetylcholine regulated prolactin secretion in rat anterior pituitary cells during aging. AB - Although it is well known that plasma concentration of prolactin (PRL) increases during aging in rats, how the anterior pituitary (AP) aging per se affects PRL secretion remains obscure. The objectives of this study were to determine if changes in the pituitary PRL responsiveness to acetylcholine (ACh; a paracrine factor in the AP), as compared with that to other PRL stimulators or inhibitors, contribute to the known age-related increase in PRL secretion, and if protein kinase C (PKC) is involved. We also determined if replenishment with aging declined hormones such as estrogen/thyroid hormone influences the aging-caused effects on pituitary PRL responses. AP cells were prepared from old (23-24-month old) as well as young (2-3-month-old) ovariectomized rats. Cells were pretreated for 5 days with diluent or 17beta-estradiol (E(2); 0.6 nM) in combination with or without triiodothyronine (T(3); 10 nM). Then, cells were incubated for 20 min with thyrotropin-releasing hormone (TRH; 100 nM), angiotensin II (AII; 0.2-20 nM), vasoactive intestinal peptide (VIP; 10(-9)-10(-5) M), dopamine (DA; 10(-9) 10(-5) M), or ACh (10(-7)-10(-3) M). Cells were also challenged with ACh, TRH, or phorbol 12-myristate 13-acetate (PMA; 10(-6) M) following PKC depletion by prolonged PMA (10(-6) M for 24 h) pretreatment. We found that estrogen priming of AP cells could reverse the aging-caused effects on pituitary PRL responses to AII and DA. In hormone-replenished cells aging enhanced the stimulation of PRL secretion by TRH and PMA, but not by AII and VIP. Aging also reduced the responsiveness of cells to ACh and DA in suppressing basal PRL secretion, and attenuated ACh inhibition of TRH-induced PRL secretion. Furthermore, ACh suppressed TRH-induced PRL secretion mainly via the PMA-sensitive PKC in the old AP cells, but via additional mechanisms in young AP cells. On the contrary, basal PRL secretion was PKC (PMA-sensitive)-independent in the old AP cells, but dependent in the young AP cells. Taken together, these results suggest differential roles of PMA-sensitive PKC in regulating basal and ACh-regulated PRL responses in old versus young AP cells. The persistent aging-induced differences in AP cell responsiveness to ACh, DA, TRH, and PMA following hormone (E(2)/T(3)) replenishment suggest an intrinsic pituitary change that may contribute, in part, to the elevated in vivo PRL secretion observed in aged rats. PMID- 12111997 TI - Sox9 transactivation and testicular expression of a novel human gene, KIAA0800. AB - The Sry and Sox9 sex-determination factors initiate and promote testis differentiation by gene transactivation through similar promoter elements. However, knowledge is limited concerning what genes are regulated by Sry/Sox9 in the testis. Identification and characterization of Sry/Sox9-regulated genes are critical for understanding sexual differentiation. We now demonstrate that a novel human gene, KIAA0800, is preferentially expressed in the testis and is transactivated by Sox9. The KIAA0800 promoter is repressed by an upstream element involving a polyT track and two Alu repeats. Two specific Sox9-bindings sites have been identified in the KIAA0800 promoter by using DNaseI footprinting assays and gel electrophoretic mobility shift assays. Sox9 transactivation of the KIAA0800 promoter appears to be exerted mainly through the relief of promoter repression. Genes homologous to the human KIAA0800 exist in organisms with differentiated sex tissues including mouse, Drosophila, and C. elegans, but not in unicellular organisms, including yeast and bacteria. Further, our recent sequence analysis shows that KIAA0800 protein is 97% identical between human and mouse. Thus, KIAA0800 is a novel Sox9-activated gene that is evolutionarily conserved and potentially involved in sexual differentiation. PMID- 12111998 TI - Effects of cell swelling on intracellular calcium and membrane currents in bovine articular chondrocytes. AB - Chondrocytes experience a dynamic extracellular osmotic environment during normal joint loading when fluid is forced from the matrix, increasing the local proteoglycan concentration and therefore the ionic strength and osmolarity. To exist in such a challenging environment, chondrocytes must possess mechanisms by which cell volume can be regulated. In this study, we investigated the ability of bovine articular chondrocytes (BAC) to regulate cell volume during a hypo-osmotic challenge. We also examined the effect of hypo-osmotic stress on early signaling events including [Ca2+](i) and membrane currents. Changes in cell volume were measured by monitoring the fluorescence of calcein-loaded cells. [Ca2+](i) was quantified using fura-2, and membrane currents were recorded using patch clamp. BAC exhibited regulated volume decrease (RVD) when exposed to hypo-osmotic saline which was inhibited by Gd3+. Swelling stimulated [Ca2+](i) transients in BAC which were dependent on swelling magnitude. Gd3+, zero [Ca2+](o), and thapsigargin all attenuated the [Ca2+](i) response, suggesting roles for Ca2+ influx through stretch activated channels, and Ca2+ release from intracellular stores. Inward and outward membrane currents significantly increased during cell swelling and were inhibited by Gd3+. These results indicate that RVD in BAC may involve [Ca2+](i) and ion channel activation, both of which play pivotal roles in RVD in other cell types. These signaling pathways are also similar to those activated in chondrocytes subjected to other biophysical signals. It is possible, then, that these signaling events may also be involved in a mechanism by which mechanical loads are transduced into appropriate cellular responses by chondrocytes. PMID- 12111999 TI - Effect of exogenous stress on the tissue-cultured mouse lens epithelial cells. AB - The effects of heat, oxidative and osmotic stress on heat shock proteins (HSP 70(I), HSC-70, and HSP-40 of tissue cultured mouse lens epithelial cells (alphaTN 4) were investigated. The effect of stress on the heat shock transcription factor (HSF-1), and a nuclear matrix protein (NM-60) of alphaTN-4 cells was also examined. Cells were exposed to heat (45 degrees C), oxidative stress (50 mM H(2)0(2)) and osmotic (600 mM medium) shock for 30 min, and then allowed to recover for 18 h in physiological medium. Control cells were maintained at 37 degrees C in an isosmolar medium. Cellular proteins were isolated and fractionated by SDS-PAGE. Western blot was used to determine the levels of HSP and nuclear proteins. Heat stressed cells were also examined, by immunofluorescence, for the specific localization of NM-60 and HSF-1. The results revealed that both NM-60 and HSF-I were present in specific locations in normal and heat-stressed cell nuclei. Nuclei isolated immediately after stress showed localization of fluorescence near the nuclear membrane. When heat stressed cells were allowed to recuperate at 37 degrees C, most of the fluorescence were relocated in discrete areas of the nucleus. These experiments showed that alphaTN 4 cells responded to stress by overexpression of HSP-70(I) and HSP-40. This increase was not present immediately after the end of the stress period, but clearly evident at 18 h of recovery in physiological medium. Immunofluorescent data suggest that heat stress induced the localization of NM-60 and HSF-1 near the nuclear membrane. PMID- 12112001 TI - Molecular characterization of protein kinase C-alpha binding to lamin A. AB - Previous results from our laboratory have identified lamin A as a protein kinase C (PKC)-binding protein. Here, we have identified the regions of PKC-alpha that are crucial for this binding. By means of overlay assays and fusion proteins made of glutathione-S-transferase (GST) fused to elements of rat PKC-alpha, we have established that binding occurs through both the V5 region and a portion of the C2 region (i.e., the calcium-dependent lipid binding (CaLB) domain) of the kinase. In particular, we have found that amino acid 200-217 of the CaLB domain are essential for binding lamin A, as a synthetic peptide corresponding to this stretch of amino acids prevented the interaction between the CaLB domain and lamin A. We also show that the presence of four lysine residues of the CaLB domain (K205, K209, K211, and K213) was essential for the binding. We have determined that binding of elements of PKC-alpha to lamin A does not require the presence of cofactors such as phosphatidylserine (PS) and Ca(2+). We have also found that the binding site of lamin A for the CaLB domain of PKC-alpha is localized in the carboxyl-terminus of the lamin, downstream of amino acid 499. Our findings may prove to be important to clarify the mechanisms regulating PKC function within the nucleus and may also lead to the synthesis of isozyme specific drugs to attenuate or reverse PKC-dependent nuclear signaling pathways important for the pathogenesis of cancer. PMID- 12112002 TI - Nitric oxide and prostaglandin E2 participate in lipopolysaccharide/interferon gamma-induced heme oxygenase 1 and prevent RAW264.7 macrophages from UV irradiation-induced cell death. AB - Induction of heme oxygenase (HO)-1 during inflammation has been demonstrated in many cell types, but the contribution of inflammatory molecules nitric oxide (NO) and prostaglandin E(2) (PGE(2)) has remained unresolved. Here we show that NO donors including sodium nitroprusside (SNP) and spermine nonoate (SP-NO), and PGE(2) significantly stimulate HO-1 expression in RAW264.7 macrophages, associated with alternative induction on NO and PGE(2) in medium, respectively. NO donors also show the inductive effect on cyclo-oxygenase 2 protein and PGE(2) production. In the presence of lipopolysaccharide and interferon-gamma (LPS/IFN gamma), HO-1 protein was induced slightly but significantly, and SNP, SP-NO, and PGE(2) enhanced HO-1 protein induced by LPS/IFN-gamma. L-Arginine analogs N-nitro L-arginine methyl ester (L-NAME) and N-nitro-L-arginine (NLA) significantly block HO-1 protein induced by LPS/IFN-gamma associated with a decrease in NO (not PGE(2)) production. And, NSAIDs aspirin and diclofenase dose dependently inhibited LPS/IFN-gamma-induced HO-1 protein accompanied by suppression of PGE(2) (not NO) production. PD98059 (a specific inhibitor of MEKK), but not SB203580 (a specific inhibitor of p38 kinase), attenuated PGE(2) (not SP-NO) induced HO-1 protein. Under UVC (100 J/m(2)) and UVB (50 J/m(2)) irradiation, PGE(2) or SP-NO treatment prevents cells from UVC or UVB-induced cell death, and HO-1 inhibitor tin protoporphyrin (SnPP) reverses the preventive effects of PGE(2) and SP-NO. The protective activity induced by PGE(2) on UVC or UVB irradiation-induced cell death was blocked by MAPK inhibitor PD98059 (not SB203580). These results demonstrated that inflammatory molecules NO and PGE(2) were potent inducers of HO 1 gene, and protected cells from UV-irradiation-induced cell death through HO-1 induction. PMID- 12112000 TI - High levels of MMP-1 expression in the absence of the 2G single nucleotide polymorphism is mediated by p38 and ERK1/2 mitogen-activated protein kinases in VMM5 melanoma cells. AB - Matrix metalloproteinase-1 (MMP-1) is one of only a few enzymes with the ability to degrade the stromal collagens (types I and III) at neutral pH, and high expression of MMP-1 has been associated with aggressive and invasive cancers. We recently reported a single nucleotide insertion/deletion polymorphism (SNP) in the collagenase-1 (MMP-1) promoter (Rutter et al. [1998] Can. Res. 58:5321-5325), where the insertion of an extra guanine (G) at -1607 bp creates the sequence, 5' GGAA-3 (2G allele), compared to the sequence 5'-GAA-3' (1G allele). The presence of 2G constitutes a binding site for the ETS family of transcription factors, and increases MMP-1 transcription in fibroblasts and A2058 melanoma cells cultured in vitro. In addition, the presence of the 2G allele has been linked to several aggressive malignancies as well as to enhanced expression of MMP-1. In this study, we describe a melanoma cell line, VMM5, that is 1G homozygous, but that is invasive and expresses high levels of MMP-1 constitutively. The high level of MMP 1 expression in VMM5 cells is due to the utilization of both the p38 and ERK1/2 transduction pathways. In contrast, in the A2058 cell line, which also expresses MMP-1 constitutively and which is 2G homozygous, only the ERK pathway is activated. Thus, our data suggest that in the absence of 2G allele and in the presence of the appropriate transcription factors, tumor cells may use alternative signal/transduction pathways and cis-acting sequences to achieve high levels of MMP-1 expression, which contribute to the ability of tumor cells to invade, regardless of their genotype. PMID- 12112003 TI - Role of GST P1-1 in mediating the effect of etoposide on human neuroblastoma cell line Sh-Sy5y. AB - The oxidative stress could have a dual action on glutathione S-transferase (GST) P1-1 metabolism: transcriptional induction and/or polymerization. The former should represent a form of adaptation to oxidative stress and contribute to protect the cell, the latter one should activate apoptosis via c-Jun N-terminal kinase (JNK). We studied the effect of etoposide on human neuroblastoma cell line SH-SY5Y and on an etoposide-resistant clone to investigate whether a pleiotropic effect of etoposide on the redox status of the cell exists which is able to interfere with apoptosis through the GST P1-1 system. Etoposide treatment was able to induce GST P1-1 polymerization and activation of apoptosis. The data obtained from our etoposide-resistant clone and the possibility to reverse the sensitive phenotype to a resistant one by means of hexyl-glutathione preincubation, seem to suggest that cellular levels of glutathione have a key role in protecting GST P1-1 by oxidation and consequently the cell's decision between life and death. PMID- 12112004 TI - Differential regulation of Cbfa1/Runx2 and osteocalcin gene expression by vitamin D3, dexamethasone, and local growth factors in primary human osteoblasts. AB - Core binding factor alpha 1 (Cbfa1) is an osteoblast-specific transcription factor essential to develop a mature osteoblast phenotype. However, its exact role in the signaling of various osteotropic-differentiating agents is still unclear. In this study, we assessed the effects of 1,25-(OH)(2)-D3 (D3), ascorbic acid, bone morphogenetic protein-2 (BMP-2), dexamethasone (Dex), and transforming growth factor-beta (TGF-beta) on Cbfa1 and osteocalcin (OCN) mRNA steady state levels (by semiquantitative RT-PCR) in an in vitro model of osteoblast differentiation. TGF-beta increased Cbfa1 mRNA levels in normal primary human osteoblasts (pHOB) by 2.6-fold in a time-dependent fashion with maximum effect on day 28 (P < 0.001). Similarly, the glucocorticoid Dex enhanced Cbfa1 gene expression by pHOB in a time-dependent fashion by up to 4.6-fold (P < 0.001). In contrast, Dex inhibited OCN gene expression levels by 68% (P < 0.01). Treatment with BMP-2 resulted in an earlier enhancement of Cbfa1 and led to a 4.2-fold increase with a maximum on day 21 (P < 0.001). Ascorbic acid did not modulate Cbfa1 and OCN gene expression. The effect of vitamin D (D3) on Cbfa1 mRNA expression was influenced by the duration of treatment, being inhibitory after 1 h and having a stimulatory effect after 48 h. Time course experiments indicated a stimulatory effect of D3 on Cbfa1 mRNA levels (by 2.5-fold after 48 h; P < 0.01). Analysis of the late cellular differentiation marker osteocalcin revealed that D3 increased OCN gene expression by 14-fold (P < 0.001). In conclusion, in normal primary human osteoblasts, the rapid and pronounced increase of OCN after treatment with D3 seems not to be mediated by Cbfa1. These data imply that Cbfa1 gene expression is differentially regulated by various osteoblastic differentiating agents and is dependent on the stage of maturation. PMID- 12112005 TI - Specific induction of heat shock protein 27 by glucocorticoid in osteoblasts. AB - It is generally recognized that osteoporosis is a common complication of patients with glucocorticoid excess and that glucocorticoid receptor is associated with heat shock protein (HSP) 70 and HSP90 in a heterocomplex. In the present study, we investigated whether glucocorticoid induces HSP27, HSP70, and HSP90 in osteoblast-like MC3T3-E1 cells. Dexamethasone time-dependently increased the levels of HSP27, while having no effect on the levels of HSP70 or HSP90. The effect of dexamethasone was dose-dependent in the range between 0.1 nM and 0.1 microM. Dexamethasone induced an increase of the levels of mRNA for HSP27. Dexamethasone induced the phosphorylation of p38 mitogen-activated protein (MAP) kinase. SB203580 and PD169316, inhibitors of p38 MAP kinase, suppressed the HSP27 accumulation by dexamethasone. In addition, SB203580 reduced the dexamethasone stimulated increase of the mRNA levels for HSP27. The dexamethasone-induced phosphorylation of p38 MAP kinase was reduced by SB203580. These results strongly suggest that glucocorticoid stimulates the induction of neither HSP70 nor HSP90, but HSP27 in osteoblasts, and that p38 MAP kinase is involved in the induction of HSP27. PMID- 12112006 TI - Characterization of stably transfected fusion protein GFP-estrogen receptor-alpha in MCF-7 human breast cancer cells. AB - Tagging hormone receptors with the green fluorescent protein (GFP) has increased our knowledge of ligand dependent sub-cellular trafficking of hormone receptors. However, the effect of the tagged hormone receptor expression on the corresponding wild type hormone receptor and endogenous gene expression has not been investigated. In this study, we constructed a MCF-7 cell line stably expressing GFP-tagged human estrogen receptor-alpha (ER) under control of the tetracycline-on system to determine the effect of GFP-ER expression on cell proliferation and expression of endogenous ER and hormone-responsive genes. Further, the inducible system was applied to determine the ligand dependent turnover rates of GFP-ER protein and mRNA. Our results demonstrate that GFP-ER expression did not affect cell cycling. Independent of ligand, GFP-ER markedly reduced the level of endogenous ER mRNA and protein, suggesting that ER negatively autoregulates its expression. Cisplatin cross-linking studies showed that GFP-ER is associated with nuclear DNA in situ, suggesting that GFP-ER is partially replacing ER at estrogen response elements. Furthermore, GFP-ER expression did not affect the estradiol induced temporal expression of hormone responsive genes c-myc and pS2. PMID- 12112007 TI - Targeted disruption of hsf1 leads to lack of thermotolerance and defines tissue specific regulation for stress-inducible Hsp molecular chaperones. AB - The rapid synthesis of heat shock proteins (Hsps) in cells subjected to environmental challenge is controlled by heat shock transcription factor-1 (Hsf1). Regulation of Hsps by Hsf1 is highly complex and, in the whole organism, remains largely unexplored. In this study, we have used mouse embryo fibroblasts and bone marrow progenitor cells from hsf1-/- mice as well as hsp70.3-lacZ knock in mice bred on the hsf1deficient genetic background (hsf1-/--hsp70.3+/--lacZ), to further elucidate the function of Hsf1 and its participation as a transcriptional activator of Hsp70 synthesis under normal or heat-induced stress conditions in vitro and in vivo. The results revealed that heat-induced Hsp70 expression in mouse tissue is entirely controlled by Hsf1, whereas its activity is not required for tissue-specific constitutive Hsp70 expression. We further demonstrate that Hsf1 is critical for maintaining cellular integrity after heat stress and that cells from hsf1-/- mice lack the ability to develop thermotolerance. This deficiency is explained by the elimination of stress inducible Hsp70 and Hsp25 response in the absence of Hsf1 activity, leading to a lack of Hsp-mediated inhibition of apoptotic cell death via both caspase dependent and caspase-independent pathways. The pivotal role of the Hsf1 transactivator in regulating rapid synthesis of Hsps as a critical cellular defense mechanism against environmental stress-induced damage is underlined. PMID- 12112008 TI - Identification of PSF as a protein kinase Calpha-binding protein in the cell nucleus. AB - Protein kinase C (PKC) isoforms are present in the cell nucleus in diverse cell lines and tissues. Since little is known about proteins interacting with PKC inside the cell nucleus, we used Neuro-2a neuroblastoma cells, in which PKCalpha is present in the nucleus, to screen for nuclear binding partners for PKC. Applying overlay assays, we detected several nuclear proteins which bind to PKCalpha. Specificity of binding was shown by its dependence on PKC activation by phorbol ester, calcium, and phosphatidylserine. The PKC-binding proteins were partially purified and analyzed by microsequencing and mass spectrometry. Four proteins could be identified: PTB-associated splicing factor (PSF), p68 RNA helicase, and the heterogeneous nuclear ribonucleoprotein (hnRNP) proteins A3 and L. In the case of PSF, binding to PKC could also be demonstrated in a GST-pull down assay using GST-PKCalpha, expressed in insect cells. Phosphorylation experiments revealed that PSF is a weak in vitro substrate for PKCalpha. PMID- 12112009 TI - Identification of novel protein/DNA interactions within the promoter of the bone related transcription factor Runx2/Cbfa1. AB - The runt homology transcription factor Runx2/Cbfa1 is essential for bone development and osteoblast differentiation. Regulatory mechanisms that govern Runx2 transcription in osteoblasts define the osteogenic pathways that control skeletal development. In this study, we systematically examined transcription factor binding within the upstream Runx2 P1 promoter, which regulates expression of the bone-related Runx2 factor. We identified two novel protein/DNA interactions that are mediated by sequence specific factors, based on cross competition experiments, point mutations, and gel-shift immunoassays. One complex recognizes a non-canonical Runx2 site, whereas the other factor binds to a palindromic sequence. Site-directed mutagenesis of the novel Runx2 motif (5'TCCCAC3') within the 0.6 kb rat Runx2 promoter reduces transcription by 2 fold, indicating that this site supports enhancement of Runx2 promoter activity. Mutation of the palindromic motif (5'AGTACT3') results in a 2-3-fold activation of the Runx2 promoter, demonstrating that the wild type sequence contributes to transcriptional repression. These studies, together with our previous findings of auto-suppression of the Runx2 promoter and negative regulation by 1,25(OH)(2) Vitamin D3, suggest that physiological control of Runx2 gene expression is mediated by a series of intricate regulatory mechanisms. PMID- 12112010 TI - Inhibition of the p38 pathway upregulates macrophage JNK and ERK activities, and the ERK, JNK, and p38 MAP kinase pathways are reprogrammed during differentiation of the murine myeloid M1 cell line. AB - Mitogen-activated protein (MAP) kinases have been implicated as important mediators of the inflammatory response. Here we report that c-Jun NH(2)-terminal kinase (JNK), extracellular signal-regulated kinase (ERK), and p38 MAP kinase activities are reprogrammed during the IL-6 induced macrophage-like differentiation of the murine myeloid M1 cell line. Moreover, p38 inhibition upregulates JNK and ERK activity in M1 cells and in thioglycollate-elicited peritoneal exudate macrophages. IL-6-induced M1 differentiation also induces expression of the anti-inflammatory cytokine IL-10, and p38 inhibition potentiates this increase in IL-10 expression in an ERK-dependent manner. Thus, we speculate that during inflammatory conditions in vivo macrophage p38 may regulate JNK and ERK activity and inhibit IL-10 expression. These data highlight the importance of p38 in the molecular mechanisms of macrophage function. PMID- 12112011 TI - Differential regulation of P-glycoprotein genes in primary rat hepatocytes by collagen sandwich and drugs. AB - P-glycoprotein (Pgp) is a small family of plasma membrane proteins, which are capable of transporting substrates across cell membranes. Class I and II Pgp are able to transport drugs and have been shown to mediate multidrug resistance (MDR). Class III Pgp is a long chain phospholipid transporter and does not mediate MDR. The regulation of all three Pgp genes is still poorly understood. For instance, it is not clear if the three Pgp genes are co-regulated or differentially regulated by external stimuli. This study examined the effect of drugs and collagen sandwich system on expression and transcription of all the three Pgp genes in primary rat hepatocytes. Consistent with previous findings, dramatic overexpression (25-fold) of Class II Pgp mRNA was seen, upon culturing of hepatocytes onto a single layered collagen gel. Hepatocytes sandwiched between two layers of collagen gel exhibited decreased (4.5-fold) Class II Pgp mRNA expression as compared to the single layer system. Treatment of hepatocytes cultured on the single layer collagen system with cytoskeletal disrupting (cytochalasin D, colchicine) but not cytoskeletal stabilizing (phalloidin, taxol) drugs, suppressed Class II Pgp expression. In all cases, no change in Class II Pgp transcription was observed as demonstrated by nuclear run-on studies. This suggests that collagen configuration and drugs affect Class II Pgp mRNA expression predominantly through post-transcriptional mechanisms. In contrast, parallel increases in mRNA expression and transcription of Class I Pgp gene were observed upon culturing of hepatocytes, in the collagen sandwich system, and treatment with some drugs (cytochalasin D, colchicine, and phalloidin). This suggests that Class I Pgp gene is regulated primarily via transcriptional mechanisms by these stimuli. On the other hand, Class III Pgp gene appears to be post-transcriptionally co-regulated with Class II Pgp gene by treatment with the drugs, while collagen configuration affected both transcription and post transcription of Class III Pgp gene. Finally, dose-dependent studies using cycloheximide provided further evidence that the two MDR-associated genes are not co-regulated. This study has implications for future studies on the molecular mechanisms of Pgp gene regulation. PMID- 12112013 TI - Does PTH have a direct effect on intestine? AB - Dogma for the past three decades has dictated that parathyroid hormone (PTH) has no direct effect on intestine with regard to calcium or phosphate absorption, but rather that PTH acts to promote the synthesis of a hormonally active form of vitamin D, namely 1,25-dihydroxyvitamin D(3) [1,25(OH)(2)D(3)]. However, diverse laboratories have each provided some evidence to suggest PTH does indeed have a direct effect on intestine. We will briefly review the evidence for biological effects, biochemical effects, and the presence of intestinal receptors for PTH, and conclude with the implications for biomedical research. PMID- 12112014 TI - Activation of bone sialoprotein gene transcription by flavonoids is mediated through an inverted CCAAT box in ROS 17/2.8 cells. AB - Bone sialoprotein (BSP) is a major noncollagenous protein of the mineralized bone extracellular matrix that has been implicated in the nucleation of hydroxyapatite. Recent studies have shown that BSP is also expressed by osteotropic cancers suggesting BSP might play a role in the pathogenesis of bone metastases. The present study investigates regulation of BSP transcription in rat osteosarcoma ROS 17/2.8 cells by flavonoids: genistein (an inhibitor of protein tyrosine kinases), daidzein (an inactive compound of genistein), flavone, and flavanone. Genistein, daidzein, and flavone (50 microM) increased steady state levels of BSP mRNA about 1.7-fold at 12 h. From transient transfection assays using various sized BSP promoter-luciferase constructs, genistein increased luciferase activities within 12 h. Constructs including the promoter sequence nucleotides (nts) -116 to -43 (pLUC3) were found to enhance transcriptional activity approximately 2.6-fold in ROS 17/2.8 cells treated with genistein (50 microM). Daidzein, flavone, and flavanone (50 microM) also increased luciferase activities. In contrast, the tyrosine kinase inhibitors, herbimycin A and lavendustin A, which do not have a flavonoid structure, did not stimulate BSP transcription. Transcriptional stimulation by genistein was almost completely abrogated in a construct that included 2 bp mutations in the inverted CCAAT box. A monoclonal antibody against NF-YA, a CCAAT box-binding transcription factor, inhibited formation of DNA-NF-Y protein complex in gel shift assays formed by nuclear extracts of ROS 17/2.8 cells. These data suggest that the inverted CCAAT box is required for flavonoid-induced BSP expression and that the stimulatory action is dependent on the flavone structure and does not involve an inhibitory action on protein tyrosine kinase. PMID- 12112012 TI - Development-dependent disappearance of caspase-3 in skeletal muscle is post transcriptionally regulated. AB - Caspase-3, a major player in apoptosis, engages apoptosis-activated cells into an irreversible pathway leading to cell death. In this article, we report that caspase-3 protein is absent from rat and mouse adult skeletal muscles, despite the abundant presence of its mRNA. During skeletal muscle development, caspase-3 protein is present in neonatal animals, but its expression gradually decreases, and disappears completely by 1 month of age, when there is still abundant caspase 3 mRNA. This discordance between caspase-3 message and protein expression is unique to skeletal muscle, as in all other analyzed tissues the protein presence correlates with the presence of the mRNA. The only circumstance in which caspase 3 protein appears in adults is in regenerating muscles; once regeneration is complete, however, it again becomes undetectable in repaired muscles. We conclude that caspase-3 protein in skeletal muscle is uniquely regulated at the post transcriptional level, unseen in other tissues such as brain, heart, lung, kidney, thymus, spleen, liver, or testis. The post-transcriptional regulation of caspase-3 might serve as a fail-safe mechanism to avoid accidental cell death. PMID- 12112015 TI - Alternative splicing during chondrogenesis: modulation of fibronectin exon EIIIA splicing by SR proteins. AB - The alternative exon EIIIA of the fibronectin gene is included in mRNAs produced in undifferentiated mesenchymal cells but excluded from differentiated chondrocytes. As members of the SR protein family of splicing factors have been demonstrated to be involved in the alternative splicing of other mRNAs, the role of SR proteins in chondrogenesis-associated EIIIA splicing was investigated. SR proteins interacted with chick exon EIIIA sequences that are required for exon inclusion in a gel mobility shift assay. Addition of SR proteins to in vitro splicing reactions increased the rate and extent of exon EIIIA inclusion. Co transfection studies employing cDNAs encoding individual SR proteins revealed that SRp20 decreased mRNA accumulation in HeLa cells, which make A+ mRNA, apparently by interfering with pre-mRNA splicing. Co-transfection studies also demonstrated that SRp40 increased exon EIIIA inclusion in chondrocytes, but not in HeLa cells, suggesting the importance of cellular context for SR protein activity. Immunoblot analysis did not reveal a relative depletion of SRp40 in chondrocytic cells. Possible mechanisms for regulation of EIIIA splicing in particular, and chondrogenesis associated splicing in general, are discussed. PMID- 12112017 TI - Retinoid X receptors and retinoid response in neuroblastoma cells. AB - Retinoic acid (RA) modulates differentiation and apoptosis of neural cells via RA receptors (RARs) and retinoid X receptors (RXRs). Neuroblastoma cells are potentially useful models for elucidating the molecular mechanisms of RA in neural cells, and responses to different isomers of RA have been interpreted in terms of differential homo- and heterodimerization of RXRs. The aim of this study was to identify the RXR types expressed in neuroblast and substrate-adherent neuroblastoma cells, and to study the participation of these RXRs in RAR heterodimers. RXRbeta was the predominant RXR type in N-type SH SY 5Y cells and S type SH EP cells. Gel shift and supershift assays demonstrated that RARbeta and RARgamma predominantly heterodimerize with RXRbeta. In SH SY 5Y cells, RARgamma/RXRbeta was the predominant heterodimer binding to the DR5 RARE in the absence of 9-cis RA (9C), whereas the balance shifted in favor of RARbeta/RXRbeta in the presence of ligand. There was a marked difference between the N- and S type neuroblastoma cells in retinoid receptor-DNA interactions, and this may underlie the differential effects of retinoids in these neuroblastoma cell types. There was no evidence to indicate that 9C functions via RXR homodimers in either SH SY 5Y or SH EP neuroblastoma cells. The results of this study suggest that interactions between retinoid receptors and other nuclear proteins may be critical determinants of retinoid responses in neural cells. PMID- 12112018 TI - Purification and characterization of protein tyrosine phosphatase PTP-MEG2. AB - PTP-MEG2 is an intracellular protein tyrosine phosphatase with a putative lipid binding domain at the N-terminus. The present study reports expression, purification, and characterization of the full-length form of the enzyme plus a truncated form containing the catalytic domain alone. Full-length PTP-MEG2 was expressed with an adenovirus system and purified from cytosolic extracts of human 293 cells infected with the recombinant adenovirus. The purification scheme included chromatographic separation of cytosolic extracts on fast flow Q Sepharose, heparin-agarose, l-histidyldiazobenzylphosphonic acid agarose, and hydroxylapatite. The enrichment of PTP-MEG2 from the cytosol was about 120-fold. The truncated form of PTP-MEG2 was expressed in E. coli cells as a non-fusion protein and purified by using a chromatographic procedure similar to that used for the full-length enzyme. The purified full-length and truncated enzymes showed single polypeptide bands on SDS-polyacrylamide gel electrophoresis under reducing conditions and behaved as monomers on gel exclusion chromatography. With para nitrophenylphosphate and phosphotyrosine as substrates, both forms of the enzyme exhibited classical Michaelis-Menten kinetics. Their responses to pH, ionic strength, metal ions, and protein phosphatase inhibitors are similar to those observed with other characterized tyrosine phosphatases. Compared with full length PTP-MEG2, the truncated DeltaPTP-MEG2 displayed significantly higher V(max) and lower K(m) values, suggesting that the N-terminal putative lipid binding domain may have an inhibitory role. The full-length and truncated forms of PTP-MEG2 were also expressed as GST fusion proteins in E. coli cells and purified to near homogeneity through affinity columns. However, the specific phosphatase activities of the GST fusion proteins were 10-25-fold below those obtained with the correspondent non-fusion proteins. PMID- 12112016 TI - Retroviral mediated expression of the human myeloid nuclear antigen in a null cell line upregulates Dlk1 expression. AB - The human myeloid nuclear differentiation antigen (MNDA) is a hematopoietic cell specific nuclear protein. MNDA and other related gene products interact with and alter the activity of a large number of proteins involved in regulating specific gene transcription. MNDA and related genes exhibit expression characteristics, which suggest functions unique to specific lineages of cells, in addition to mediating the effects of interferons. Cells of the human K562 myeloid line do not express MNDA and are relatively immature compared to lines that express MNDA (HL 60, U937, and THP1). The hypothesis that MNDA influences the expression of specific genes was tested by creating MNDA expressing K562 cells using stable retroviral mediated gene transfer followed by evaluation of transcription profiles. Two macroarrays containing a total of 2,350 cDNAs of known genes showed a specific up-regulation of Dlk1 expression in MNDA expressing K562 cell clones. Real time quantitative RT-PCR analysis confirmed an average of over 3- and 7-fold upregulation of Dlk1 in two clones of MNDA expressing K562 cells. The effects on Dlk1 were also confirmed by Northern blotting. Dlk1 is essential for normal hematopoiesis and abnormal expression is a proposed marker of myelodysplastic syndrome. Additional screening of transcription profiles after induced erythroid and megakaryoblastic differentiation showed no additional gene transcripts altered by the presence of MNDA. These results indicate that MNDA alters expression of a gene essential for normal hematopoiesis. PMID- 12112019 TI - Effects of bone morphogenetic protein-7 stimulation on osteoblasts cultured on different biomaterials. AB - The objective of the present study was to investigate the effects of an in vitro stimulation of human osteoblasts by recombinant human bone morphogenetic protein 7 (rhBMP-7) on the collagen types and the quantity of the collagen cross-links synthesized in a three-dimensional culture on various biomaterials for bone replacement. Trabecular bone chips were harvested from human iliac crests, and cell cultures were established at standard conditions. One hundred and fifty nanograms per milliliter of rhBMP-7 was added. For the second passage a cell scraper was used to bring the cells into suspension, and 100 microl osteoblasts (at a density of 3.3 x 10(5)) were transferred onto nine blocks of either Bio Oss, Tutoplast, or PepGen p-15. Blocks incubated with cells that were not treated with rhBMP-7 served as controls. Cell colonization of the biomaterials was observed by scanning electron microscopy (SEM) and transmission electron microscopy (TEM) after a period of 2, 4, and 6 weeks. Throughout the experiment medium, supernatants were collected and collagen was characterized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Finally, the collagen cross-link residues hydroxylysylpyridinoline (HP) and lysylpyridinoline (LP) were quantified by HPLC. Within 4 weeks the cells became confluent on all of the studied biomaterials. All samples synthesized bone specific LP and collagen type I. However, in rhBMP-7-stimulated samples, the amount of HP and LP found was increased by 45% compared to non-stimulated samples. Cell proliferation and collagen synthesis was similar on the different biomaterials, but was consistently reduced in specimen not stimulated with rhBMP-7. In vitro stimulation of osteoblasts on Bio-Oss, Tutoplast, or PepGen p-15 with rhBMP-7 and subsequent transplantation of the constructs might lead to an enhanced osseointegration of the biomaterials in vivo. PMID- 12112020 TI - Sam68 associates with the SH3 domains of Grb2 recruiting GAP to the Grb2-SOS complex in insulin receptor signaling. AB - The 68 kDa Src substrate associated during mitosis (Sam68) is an RNA binding protein with Src homology (SH) 2 and 3 domain binding sites. We have recently found that Sam68 is a substrate of the insulin receptor (IR) that translocates from the nucleus to the cytoplasm and that Tyr-phosphorylated Sam68 associates with the SH2 domains of p85 PI3K and GAP, in vivo and in vitro. In the present work, we have further demonstrated the cytoplasmic localization of Sam68, which is increased in cells overexpressing IR. Besides, we sought to further study the association of Sam68 with the Ras-GAP pathway by assessing the interactions with SH3 domains of Grb2. We employed GST-fusion proteins containing the SH3 domains of Grb2 (N or C), and recombinant Sam68 for in vitro studies. In vivo studies of protein-protein interaction were assessed by co-immunoprecipitation experiments with specific antibodies against Sam68, GAP, Grb2, SOS, and phosphotyrosine; and by affinity precipitation with the fusion proteins (SH3-Grb2). Insulin stimulation of HTC-IR cells promotes phosphorylation of Sam68 and its association with the SH2 domains of GAP. Sam68 is constitutively associated with the SH3 domains of Grb2 and it does not change upon insulin stimulation, but Sam68 is Tyr phosphorylated and promotes the association of GAP with the Grb2-SOS complex. In vitro studies with fusion proteins showed that Sam68 association with Grb2 is preferentially mediated by the C-terminal SH3 domains of Grb2. In conclusion, Sam68 is a substrate of the IR and may have a role as a docking protein in IR signaling, recruiting GAP to the Grb2-SOS complex, and in this way it may modulate Ras activity. PMID- 12112021 TI - Progesterone attenuates the inhibitory effects of cardiotonic digitalis on pregnenolone production in rat luteal cells. AB - Previous studies have shown that digoxin decreases testosterone secretion in testicular interstitial cells. However, the effect of digoxin on progesterone secretion in luteal cells is unclear. Progesterone is known as an endogenous digoxin-like hormone (EDLH). This study investigates how digitalis affected progesterone production and whether progesterone antagonized the effects of digitalis. Digoxin or digitoxin, but not ouabain, decreased the basal and human chorionic gonadotropin (hCG)-stimulated progesterone secretion as well as the activity of cytochrome P450 side chain cleavage enzyme (P450scc) in luteal cells. 8-Br-cAMP and forskolin did not affect the reduction. Neither the amount of P450scc, the amount of steroidogenic acute regulatory (StAR) protein, nor the activity of 3beta-hydroxysteroid dehydrogenase (3beta-HSD) was affected by digoxin or digitoxin. Moreover, in testicular interstitial and luteal cells, progesterone partially attenuated the reduction of pregnenolone by digoxin or digitoxin and the progesterone antagonist, RU486, blocked this attenuation. These new findings indicated that (1) digoxin or digitoxin inhibited pregnenolone production by decreasing the activity of P450scc enzyme, but not Na(+)-K(+) ATPase, resulting in a decrease on progesterone secretion in rat luteal cells, and (2) the inhibitory effect on pregnenolone production by digoxin or digitoxin was reversed partially by progesterone. In conclusion, digoxin or digitoxin decreased progesterone production via the inhibition of pregnenolone by decreasing P450scc activity. Progesterone, an EDLH, could antagonize the effects of digoxin or digitoxin in luteal cells. PMID- 12112022 TI - Different cellular localization, translocation, and insulin-induced phosphorylation of PKBalpha in HepG2 cells and hepatocytes. AB - Protein kinase B (PKB), a serine/threonine protein kinase, prevents apoptosis and promotes cellular transformation. PKB activity is stimulated by insulin. In this report, we examined the relative amounts of expression, location, and translocation upon insulin stimulation of PKBalpha in normal primary hepatocytes and carcinoma cells, HepG2 cells. Non-phosphorylated PKBalpha was present in both types of unstimulated cells. The phosphorylated form of the enzyme was present in the nucleus of unstimulated HepG2 cells but not in normal hepatocytes. In the cytoplasm, PKBalpha was found in greater abundance in the hepatocytes as compared in HepG2 cells. Insulin induced the translocation of phosphorylated PKBalpha from the nucleus to the nuclear membrane in HepG2 cells. In contrast, insulin caused translocation and phosphorylation of PKBalpha from the cytosol to the plasma membrane in normal hepatocytes. In addition, there is a higher expression of PKBalpha in the HepG2 cells as compared to normal primary hepatocytes. These findings provide an important distinction between hepatocellular HepG2 cells and normal liver cells and suggest that the presence of constitutively active nuclear PKB in the transformed cells might be an important contributor in cell transformation and immortality of hepatoma cells. PMID- 12112023 TI - 1,25(OH)2-vitamin D3 induces translocation of the vitamin D receptor (VDR) to the plasma membrane in skeletal muscle cells. AB - 1,25-dihydroxy-vitamin D(3) (1,25(OH)(2)D(3)), the hormonally active form of vitamin D(3), acts through two different mechanisms. In addition to regulating gene expression via the specific intracellular vitamin D receptor (VDR), 1,25(OH)(2)D(3) induces rapid, non-transcriptional responses involving stimulation of transmembrane signal transduction pathways. The activation of second messengers supports the hypothesis that a membrane-bound steroid receptor similar to those that mediate peptide hormone biology exists. Skeletal muscle is a target tissue for 1,25(OH)(2)D(3). Avian embryonic skeletal muscle cells (myoblasts/myotubes) have been shown to respond both genomically and non genomically to the hormone. The present study provides evidence indicating that short-term treatment (1-10 min) with 1,25(OH)(2)D(3) induces translocation of the VDR from the nuclear to the microsomal fraction in chick myoblasts. This translocation is blocked by colchicine, genistein, or herbimycin, suggesting the involvement of microtubular transport and tyrosine kinase/s in the relocation of the receptor. By isolation of plasma membranes, it was demonstrated that the hormone increases the amounts of VDR specifically in this fraction. These results suggest that the nuclear VDR may be the receptor that mediates the non-genomic effects of 1,25(OH)(2)D(3) in chick myoblasts. PMID- 12112025 TI - Metallothionein is required for zinc-induced expression of the macrophage colony stimulating factor gene. AB - Macrophage colony stimulating factor (M-CSF) plays an important role in the proliferation and differentiation of mononuclear phagocytes. The present study investigates the effect of zinc on M-CSF expression in MC3T3-E1 and L929 cells. Zinc dose-dependently increased M-CSF mRNA levels. The time-course of zinc induced M-CSF mRNA expression peaked at 6 h. Stability studies of mRNA using actinomycin D revealed that zinc does not affect M-CSF mRNA stability. We examined the function of the M-CSF gene promoter using a luciferase reporter assay. A construct containing the -467/+39 region of the promoter was upregulated by zinc. In the presence of cycloheximide, zinc did not induce a greater increase in the M-CSF mRNA than cycloheximide alone. To confirm the effect of MT on M-CSF mRNA expression, mouse lung fibroblasts (MLFs) were prepared from MT+/+ and MT-/- mice. Zinc induced an increase in the expression of M-CSF in MT+/+ MLFs, but this response was not evident in MT-/- MLFs. Moreover, overexpression of MT upregulated M-CSF mRNA expression as well as M-CSF secretion. Our findings suggest that MT expression mediates zinc regulation of M-CSF gene expression at the transcriptional level. PMID- 12112024 TI - Expression and cellular localization of naturally occurring beta estrogen receptors in uterine and mammary cell lines. AB - The protein ER-alpha has been exhaustively characterized in estrogen-sensitive tissues and cell lines. However, little is known regarding the expression and cellular distribution of the newly identified ER-beta protein. We first quantified the specific estradiol binding site content in the estrogen-responsive cell lines MCF-7 (mammary) and SHM (myometrial). In the two cell types, these sites were associated to the expression of both ER-alpha and -beta isoforms. Native ER-beta was visualized to reside inside the nucleus by means of conventional indirect immunofluorescence. The cells expressed ER-beta as a tight approximately 50 kDa triplet when resolved by sodium dodecyl sulfate polyacrylamide gels (SDS-PAGE) and blotted using antibodies mapping different domains of the cloned ER-beta version. When the cells were subjected to homogenization and differential centrifugation, a substantial proportion of ER beta immunolabeling was localized at membrane subfractions. ER-beta expression and partitioning was confirmed by Ligand blotting assays using estrogen derivatives coupled to different macromolecular tags. However, ER-alpha was expressed as the major estrogen binding protein in both cell lines. Similar localization experiments were performed on HeLa cells (cervix). Though usually considered ER-negative, this cell line displayed basal significant estrogen binding capacity and co-expression of both ER isoforms. Taken as a whole, the results indicate that ER-beta could be expressed as functional estrogen binding proteins among a dominant population of ER-alpha sites in the cell lines under study. PMID- 12112026 TI - Activation of stress response by ionomycin in rat hepatoma cells. AB - All living systems respond to a variety of stress conditions by inducing the synthesis of stress or heat shock proteins (HSPs), which transiently protect cells. HSP synthesis was preceded by an increase in intracellular free calcium concentration [(Ca(2+))i]. In this study, we show that Ca(2+) ionophore, ionomycin, induced an immediate increase in intracellular free Ca(2+) and examined how this increase affects heat shock response in rat hepatoma cell line H4II-E-C3. Results indicate that incubating H4II-E-C3 cells with 0.3 microM ionomycin at 37 degrees C for 15 min results in the induction of HSP 70 in both Ca(2+)-containing and Ca(2+)-free medium. Associated with this increase in free Ca(2+) is an in vivo change in membrane organization and activation of signaling molecules like ERKS and SAPKs/JNK. In Ca(2+) containing medium HSP 70 induction mediated by HSF-HSE interaction was faster upon ionomycin treatment as compared to heat shock. Our results show that ionomycin, at sub lethal concentration, increases intracellular free Ca(2+) concentration, activates SAPK/JNK and HSF-HSE interaction, and induces HSP 70 synthesis. PMID- 12112027 TI - High levels of exogenous C2-ceramide promote morphological and biochemical evidences of necrotic features in thyroid follicular cells. AB - CD95 and ceramide are known to be involved in the apoptotic mechanism. The triggering of CD95 induces a cascade of metabolic events that progressively and dramatically modifies the cell shape by intense membrane blebbing, leading to apoptotic bodies production. Although the CD95 pathway has been abundantly described in normal thyrocytes, the effects of cell permeable synthetic ceramide at morphological and biochemical levels are not fully known. In the present study, we show that thyroid follicular cells (TFC) exposed to 20 microM of C(2) ceramide for 4 h are characterized by morphological features of necrosis, such as electron-lucent cytoplasm, mitochondrial swelling, and loss of plasma membrane integrity without drastic morphological changes in the nuclei. By contrast, TFC treated with 2 microM of C(2)-ceramide for 4 h are able to accumulate GD3, activate caspases cascade, and induce apoptosis. Furthermore, we provide evidence that 20 microM of C(2)-ceramide determine the destruction of mitochondria and are not able to induce PARP cleavage and internucleosomal DNA fragmentation, suggesting that the apoptotic program is not activated during the death process and nuclear DNA is randomly cleaved as the consequence of cellular degeneration. Pretreatment with 30 microM of zVAD-fmk rescued TFC from 2 microM of C(2) ceramide-induced apoptosis, whereas, 20 microM of C(2)-ceramide exposure induced necrotic features. Deltapsi(m) was obviously altered in cells treated with 20 microM of C(2)-ceramide for 4 h (75% +/- 3.5%) compared with the low percentage (12.5% +/- 0.4%) of cells with altered Deltapsi(m) exposed to 2 microM of C(2) ceramide. Whereas, only 20% +/- 1.1% of cells treated with anti-CD95 for 1 h showed altered Deltapsi(m). Additionally, Bax and Bak, two pro-apoptotic members, seem to be not oligomerized in the mitochondrial membrane following ceramide exposure. These results imply that high levels of exogenous ceramide contribute to the necrotic process in TFC, and may provide key molecular basis to the understanding of thyroid signaling pathways that might promote the apoptotic mechanism in thyroid tumoral cells. PMID- 12112028 TI - Regulation of Her2/neu promoter activity by the ETS transcription factor, ER81. AB - Overexpression of the HER2/Neu receptor is correlated to a poor prognosis in tumor patients and leads to stimulation of mitogen-activated protein kinase (MAPK) signaling pathways, which in turn activate transcription factors, such as the ETS protein ER81. Here, we have analyzed whether, on the other hand, ER81 may regulate the Her2/neu gene. Indeed, ER81, together with its co-activators, p300 and CBP, activates the Her2/neu promoter, and this activation is enhanced upon stimulation of MAPK pathways as well as by oncogenic HER2/Neu protein. Furthermore, ER81 interacts with one ETS binding site in the Her2/neu promoter, whose mutation decreases ER81-mediated transcription. Activation of the Her2/neu promoter is also diminished upon mutation of MAPK-dependent phosphorylation sites in ER81 or upon deletion of ER81 transactivation domains. In addition, the ER81 DNA-binding domain on its own functions as a dominant-negative molecule, effectively repressing any stimulation of the Her2/neu promoter. Altogether, our results show that ER81 is a component of a positive regulatory feedback loop, in which the HER2/Neu protein activates ER81, as well as p300/CBP via MAPKs causing the upregulation of the Her2/neu gene. PMID- 12112029 TI - Role of regucalcin as an activator of sarcoplasmic reticulum Ca2+-ATPase activity in rat heart muscle. AB - The expression of regucalcin, a regulatory protein of Ca(2+) signaling, and its effect on Ca(2+) pump activity in the microsomes (sarcoplasmic reticulum) of rat heart muscle was investigated. The expression of regucalcin mRNA was demonstrated by reverse transcription-polymerase chain reaction (RT-PCR) analysis in heart muscle using rat regucalcin-specific primers. Results with Western blot analysis showed that regucalcin protein was present in the cytoplasm, although it was not detected in the microsomes. Microsomal Ca(2+)-ATPase activity was significantly increased in the presence of regucalcin (10(-10)-10(-8) M) in the enzyme reaction mixture. This increase was not seen in the presence of thapsigargin (TP) (10(-5) M), a specific inhibitor of the microsomal Ca(2+) pump enzyme. Regucalcin (10( 10)-10(-8) M) significantly stimulated ATP-dependent (45)Ca(2+) uptake by the microsomes. The effect of regucalcin (10(-8) M) in increasing microsomal Ca(2+) ATPase activity was completely prevented in the presence of digitonin (10(-3) or 10(-2)%), which has a solubilizing effect on membranous lipid, or N ethylmaleimide (NEM), a modifying reagent of sulfhydryl (SH) groups. Dithiothreitol (DTT; 5 mM), a protecting reagent of SH groups, increased markedly Ca(2+)-ATPase activity. In the presence of DTT (5 mM), regucalcin could not significantly enhance the enzyme activity. Also, the effect of regucalcin in increasing Ca(2+)-ATPase activity was completely inhibited by the addition of vanadate (1 mM), an inhibitor of phosphorylation of enzyme. In addition, the effect of regucalcin on Ca(2+)-ATPase activity was not significantly modulated in the presence of dibutyryl cyclic AMP (10(-4) M), inositol 1,4,5-trisphosphate (10(-3) M), or calmodulin (5 microg/ml) which is an intracellular signaling factor. The present study demonstrates that regucalcin can activate Ca(2+) pump activity in rat heart microsomes, and that the protein may act the SH groups of Ca(2+)-ATPase by binding to microsomal membranes. PMID- 12112030 TI - Differential expression of a WD protein during squamous differentiation of tracheal epithelial cells. AB - The lining of the trachea consists of a pseudostratified, mucociliary epithelium that under a variety of conditions, such as vitamin A deficiency, toxic and mechanical injury, becomes a stratified squamous epithelium. Several in vitro cell culture models have been established to study the process of differentiation of airway epithelium. Such studies have indicated that mucosecretory differentiation of tracheal epithelial cells can be modulated by substratum. This study was undertaken to understand molecular mechanisms of squamous differentiation in tracheal epithelia. Primary cultured tracheal cells grown on uncoated filters were differentiated to single layer of squamous cells, whereas cells were grown as stratified columnar cells on collagen-I coated filters. The responses to secretagogues were altered according to culture conditions. DD-PCR revealed that FAK and a WD protein expression was increased in squamous tracheal epithelia. Expression of a WD protein was changed by the treatment of retinoic acid in various epithelial cells. These results indicated that squamous differentiation of tracheal cells changes the expression of a variety of genes, and that the experimental model for this study can be employed to study molecular mechanisms of squamous differentiation in airway epithelial cells. PMID- 12112031 TI - Inhibition of CYP 17, a new strategy for the treatment of prostate cancer. AB - Androgens are growth factors for approximately 80 percent of all prostate cancers. Suppressing androgen biosynthesis is therefore an important therapeutic strategy in order to inhibit tumor growth. Unfortunately, the drugs currently applied to lower androgen levels only affect testicular androgen production. Since androgens are also synthesized in the adrenal glands, tumor stimulation cannot be blocked completely. A new therapeutic target, CYP 17 (P450 17, 17alpha hydroxylase-C17, C20 lyase), is likely to improve this situation. CYP 17 is a P450 enzyme and catalyzes the last step of androgen biosynthesis in both testes and adrenals. Inhibition of this enzyme will therefore result in a complete block of androgen production. This paper gives an overview of the current situation in this novel field of drug research and focuses on the development of steroidal and non-steroidal inhibitors of CYP 17. PMID- 12112033 TI - Non-thiol farnesyltransferase inhibitors: evaluation of different AA(X) peptidomimetic substructures in combination with arylic cysteine replacements. AB - In the course of our studies on non-thiol farnesyltransferase inhibitors based on the 2, 5-diaminobenzophenone AAX-peptidomimetic substructure, we have developed the (4-nitrophenyl)butyryl (R(1)), the (2-naphthyl)acryloyl (R(2)), the 4 nitrocinnamoyl (R(3)), and the 5-(4-nitrophenyl)furylacryloyl (R(4)) groups as useful cysteine replacements. In this study, we combined these four groups with other AA(X)-peptidomimetic substructures (5-10: R = H) reported in the literature. The 5-(4-nitrophenyl)furylacryloyl moiety (R(4)) turned out to be the most useful non-thiol cysteine replacement yielding in all cases the most active inhibitors. By combination of this 5-(4-nitrophenyl) furylacryloyl moiety (R(4)) with the structurally simple AAX-peptidomimetics 4-aminobenzophenone (5) and 4 aminodiphenylsulfone (6) potent, readily accessible non-thiol farnesyltransferase inhibitors were obtained (IC(50) = 12 nMand 10 nM). PMID- 12112032 TI - Anticonvulsant and sedative-hypnotic activities of N-substituted isatin semicarbazones. AB - A series of N-methyl/acetyl, 5-(un)-substituted isatin-3-semicarbazones were screened for anticonvulsant and sedative-hypnotic activities. The results revealed that protection was obtained in all the screens i.e., MES, scPTZ, and scSTY. Compounds 2, 4, 6, 10 but not 1 and 3 showed low neurotoxicity when compared to clinically used drugs. Compounds 5, 7, 8 and 9 were completely non toxic. Compound 6 showed good activity in the rat oral MES screen. Among all the compounds, 3 and 6 emerged as the most active compounds as indicated by the protection they exhibit in MES, scSTY, and scPTZ screens. All the compounds showed significant sedativehypnotic activity. PMID- 12112034 TI - Heterologous expression of a bacterial homospermidine synthase gene in transgenic tobacco: effects on the polyamine pathway. AB - Homospermidine synthase (HSS) is a branch-point enzyme that links the secondary pathway (pyrrolizidine alkaloids) to primary metabolism (polyamines). Since the diamine putrescine is a precursor of homospermidine and nicotine in tobacco, we performed heterologous expression of a bacterial homospermidine synthase gene (hss)in Nicotiana tabacum and determined the effect on free and conjugated polyamine levels. The hss gene from Rhodopseudomonas viridis was placed under the control of the cauliflower mosaic virus (CaMV) 35S promoter in Agrobacterium tumefaciens Ti-plasmid in sense and antisense orientation and both hss constructs were transformed into tobacco plants. Expression of the hss gene was verified by "Northern" and "Southern Blot" analysis. 2 transgenic sense lines were generated from 1000 calli which showed weak expression of homospermidine synthase, i.e. 50 pktal/mg protein and 45 pktal/mg protein. These transgenic sense plants showed a significantly decreased content of free spermidine while the pool of conjugated spermidine was not affected. The 2 sense plants exhibited a range of abnormal phenotypes such as dwarfness and stunted growth. Homospermidine was sporadically detectable in wild type tobacco. To our knowledge, this is the first biotechnological approach to express a prokaryotic homospermidine synthase gene in tobacco plants. PMID- 12112035 TI - Synthesis and antibacterial activity of 1beta-methyl-2-(5-substituted thiazolidinopyrrolidin-3-ylthio)carbapenems and related compounds. AB - The synthesis of a new series of 1beta-methylcarbapenems containing the substituted thiazolidinopyrrolidine moiety is described. Their in vitro antibacterial activities against both Gram-positive and Gram-negative bacteria were tested and the effect of substituent on the thiazolidine ring was investigated.A particular compound (18 c) having a 2-amide substituted thiazolidine moiety showed the most potent antibacterial activity. PMID- 12112036 TI - Tritylamino aromatic heterocycles and related carbinols as blockers of ca 2+ activated potassium ion channels underlying neuronal hyperpolarization. AB - A series of novel aromatic tritylamino heterocycles has been synthesized and the compounds have been tested in comparison with clotrimazole for their ability to inhibit the slow afterhyperpolarization current (sI (AHP)) in cultured rat hippocampal pyramidal neurones. Some analogues of the clotrimazole metabolite, 2 chlorophenyl-diphenyl methanol, having different chlorination substitution in the triphenyl group have also been examined. Two compounds in particular, 3-[(2 chlorophenyl)-diphenylmethylamino] pyridine (3a, UCL 1880) and 2 tritylaminothiazole (6, UCL 2027), are of special interest; they are effective blockers of the sI (AHP) (IC (50) = 1.1-1.2 microM) and are much more selective than clotrimazole since they have less effect on the high voltage-activated Ca2+ current. PMID- 12112037 TI - Oxidation and degradation products of papaverine. Part I: Gadamer and Schulemann's papaverinol synthesis revisited. AB - In 1915 Gadamer published in this journal [1] a procedure for the synthesis of papaverinol 2 from papaverine 1 in excellent yield. However, he did not investigate the formation of a violet fluorescence produced upon crystallization of papaverinol 2 from ethanol. The compound responsible for this fluorescence was isolated and identified as the yet unknown quaternary ammonium ion 4, a 6a, 12a diazadibenzo-[a, g]fluorenylium derivative. The isolation of 4 and its structure determination by spectroscopic methods are described. However, its formation mechanism is unknown. PMID- 12112038 TI - Message from the editors. PMID- 12112040 TI - Hyperglycemia and stroke outcome: vindication of the ischemic penumbra. PMID- 12112041 TI - Bourneville and Taylor: a developing story? PMID- 12112042 TI - Cortical control of spatial memory in humans: the visuooculomotor model. AB - We review current knowledge of the cortical control of spatial memory, studied using visuooculomotor paradigms. Spatial memory is an essential cognitive process that can be involved in preparing motor responses. Our knowledge of spatial memory in humans recently has progressed thanks to the use of ocular saccades as a convenient model of motor behavior. Accuracy of memory-guided saccades, made to the remembered locations of visual targets, is a reflection of spatial memory. For the performance of memory-guided saccades with brief delays (up to 15-20 seconds), that is, involving short-term spatial memory, lesion studies have shown that the posterior parietal cortex, the dorsolateral prefrontal cortex, and the frontal eye field play significant roles. Studies of memory-guided saccades using transcranial magnetic stimulation have suggested that the right posterior parietal cortex is involved at the initial stage (<300 milliseconds) of visuospatial integration, whereas the dorsolateral prefrontal cortex in both hemispheres controls the following phase of short-term memorization, the frontal eye field mainly serving to trigger saccades. The new concept of a medium-term spatial memory has emerged from a behavioral study of memory-guided saccades in normal subjects, showing a paradoxical spontaneous improvement of spatial memory after delays of approximately 20 seconds. Lesion studies have shown that the parahippocampal cortex could specifically control this medium-term spatial memory. Last, different experimental and clinical arguments suggest that, after a few minutes, the hippocampal formation finally takes over the control of spatial memory for long-term spatial memorization. Therefore, spatial memory involved in the memorization of visual items could be successively controlled by the dorsolateral prefrontal cortex (short-term spatial memory), the parahippocampal cortex (medium-term spatial memory), and the hippocampal formation (long-term spatial memory), depending on specific periods of times. The applicability of this simple visuooculomotor model of spatial memory to other types of stimuli and general motoricity has yet to be confirmed. PMID- 12112043 TI - Acute hyperglycemia adversely affects stroke outcome: a magnetic resonance imaging and spectroscopy study. AB - Controversy exists whether acute hyperglycemia is causally associated with worse stroke outcome or simply reflects a more severe stroke. In reversible ischemia models, hyperglycemia is associated with lactic acidosis and conversion of penumbral tissue to infarction. However, the relationship between hyperglycemia, lactic acidosis, and stroke outcome has not been explored in humans. Sixty-three acute stroke patients were prospectively evaluated with serial diffusion-weighted and perfusion-weighted magnetic resonance imaging and acute blood glucose measurements. Patients with hypoperfused at-risk tissue were identified by acute perfusion-diffusion lesion mismatch. As a substudy, acute and subacute magnetic resonance spectroscopy was performed in the 33 most recent patients to assess the relationship between acute blood glucose and lactate production in the ischemic region. In 40 of 63 patients with acute perfusion-diffusion mismatch, acute hyperglycemia was correlated with reduced salvage of mismatch tissue from infarction, greater final infarct size, and worse functional outcome. These correlations were independent of baseline stroke severity, lesion size, and diabetic status. Furthermore, higher acute blood glucose in patients with perfusion-diffusion mismatch was associated with greater acute-subacute lactate production, which, in turn, was independently associated with reduced salvage of mismatch tissue. In contrast, acute blood glucose levels in nonmismatch patients did not independently correlate with outcome measures, nor was there any acute subacute increase in lactate in this group. Acute hyperglycemia increases brain lactate production and facilitates conversion of hypoperfused at-risk tissue into infarction, which may adversely affect stroke outcome. These findings support the need for randomized controlled trials of aggressive glycemic control in acute stroke. PMID- 12112044 TI - Focal cortical dysplasia of Taylor's balloon cell type: mutational analysis of the TSC1 gene indicates a pathogenic relationship to tuberous sclerosis. AB - Focal cortical dysplasia (FCD) is characterized by a localized malformation of the neocortex and underlying white matter. Balloon cells, similar to those observed in tuberous sclerosis, are present in many cases (FCD(bc)). In these patients, a hyperintense funnel-shaped subcortical lesion tapering toward the lateral ventricle was the characteristic finding on fluid-attenuated inversion recovery magnetic resonance imaging scans. Surgical lesionectomy results in complete seizure relief. Although the pathogenesis of FCD(bc) remains uncertain, histopathological similarities indicate that FCD(bc) may be related pathogenetically to tuberous sclerosis. Here, we studied alterations of the TSC1 and TSC2 genes in a cohort of patients with chronic, focal epilepsy and histologically documented FCD(bc) (n = 48). DNA was obtained after microdissection and laser-assisted isolation of balloon cells, dysplastic neurons, and nonlesional cells from adjacent normal brain tissue. Sequence alterations resulting in amino acid exchange of the TSC1 gene product affecting exons 5 and 17 and silent base exchanges in exons 14 and 22 were increased in patients with FCD(bc) compared with 200 control individuals (exon 5, 2.3% FCD(bc) vs 0% C; exon 17, 35% FCD(bc) vs 1.0% C; exon 14, 37.8% FCD(bc) vs 15% C; exon 22, 45% FCD(bc) vs 23.8% C). Sequence alterations could be detected in FCD(bc) and in adjacent normal cells. In 24 patients, DNA was suitable to study loss of heterozygosity at the TSC1 gene locus in microdissected FCD(bc) samples compared with control tissue. Eleven FCD(bc) cases exhibited loss of heterozygosity. In the TSC2 gene, only silent polymorphisms were detected at similar frequencies as in controls. Our findings indicate that FCD(bc) constitutes a clinicopathological entity with distinct neuroradiological, neuropathological, and molecular genetic features. These data also suggest a role of the TSC1 gene in the development of FCD(bc) and point toward a pathogenic relationship between FCD(bc) and the tuberous sclerosis complex. PMID- 12112045 TI - Succinate in dystrophic white matter: a proton magnetic resonance spectroscopy finding characteristic for complex II deficiency. AB - A deficiency of succinate dehydrogenase is a rare cause of mitochondrial encephalomyopathy. Three patients, 2 sisters and 1 boy from an unrelated family, presented with symptoms and magnetic resonance imaging signs of leukoencephalopathy. Localized proton magnetic resonance spectroscopy indicated a prominent singlet at 2.40ppm in cerebral and cerebellar white matter not present in gray matter or basal ganglia. The signal was also elevated in cerebrospinal fluid and could be identified as originating from the two equivalent methylene groups of succinate. Subsequently, an isolated deficiency of complex II (succinate:ubiquinone oxidoreductase) was demonstrated in 2 patients in muscle and fibroblasts. One of the sisters died at the age of 18 months. Postmortem examination showed the neuropathological characteristics of Leigh syndrome. Her younger sister, now 12 months old, is also severely affected; the boy, now 6 years old, follows a milder, fluctuating clinical course. Magnetic resonance spectroscopy provides a characteristic pattern in succinate dehydrogenase deficiency. PMID- 12112046 TI - Application of the new McDonald criteria to patients with clinically isolated syndromes suggestive of multiple sclerosis. AB - Traditionally, multiple sclerosis (MS) has been diagnosed on the basis of clinical evidence of dissemination in time and space. Previously, it could not be diagnosed in patients with single clinical episodes of demyelination known as clinically isolated syndromes. New diagnostic criteria from the International Panel of McDonald and colleagues incorporate MRI evidence of dissemination in time and space to allow a diagnosis of MS in patients with clinically isolated syndromes. From clinical and MRI examinations performed prospectively at baseline, 3 months, 1 year, and 3 years of follow-up, the frequency of developing MS was ascertained by the application of both the new McDonald criteria and the Poser criteria for clinically definite MS. The specificity, sensitivity, positive and negative predictive value, and accuracy of the new criteria for the development of clinically definite MS were assessed. At 3 months, 20 of 95 (21%) patients had MS with the McDonald criteria, whereas only 7 of 95 (7%) had developed clinically definite MS. After 1 year, the corresponding figures were 38 of 79 (48%) and 16 of 79 (20%), and after 3 years, they were 29 of 50 (58%) and 19 of 50 (38%). The development of MS with the new MRI criteria after 1 year had a high sensitivity (83%), specificity (83%), positive predicative value (75%), negative predictive value (89%), and accuracy (83%) for clinically definite MS at 3 years. Use of the new McDonald criteria more than doubled the rate of diagnosis of MS within a year of presentation with a clinically isolated syndrome. The high specificity, positive predictive value, and accuracy of the new criteria for clinically definite MS support their clinical relevance. PMID- 12112047 TI - Minocycline markedly protects the neonatal brain against hypoxic-ischemic injury. AB - Hypoxic-ischemic brain injury in the perinatal period is a major cause of morbidity and mortality. Presently, there are no proven effective therapies with which to safeguard the human neonatal brain against this type of injury. Minocycline, a semisynthetic tetracycline, has been shown to be neuroprotective in certain adult ischemic injury/stroke and neurodegenerative disease models. However, minocycline's neuroprotective effects have not been assessed after insults to the neonatal brain. We now report that minocycline administered either immediately before or immediately after a hypoxic-ischemic insult substantially blocks tissue damage in a rodent model of neonatal hypoxic-ischemic brain injury. Minocycline treatment prevents the formation of activated caspase-3, a known effector of apoptosis, as well as the appearance of a calpain cleaved substrate, a marker of excitotoxic/necrotic cell death. To our knowledge, this is the first report of a systemic treatment that can be administered after a hypoxic-ischemic insult, which provides robust, nearly complete neuroprotection to the developing brain. Our data suggest that minocycline or a related neuroprotective tetracycline may be a candidate to consider in human clinical trials to protect the developing brain against hypoxic-ischemic-induced damage. PMID- 12112048 TI - Magnetization transfer imaging in normal aging, mild cognitive impairment, and Alzheimer's disease. AB - The purpose of this study was to assess whether structural brain damage as detected by volumetric magnetization transfer imaging (MTI) is present in mild cognitive impairment (MCI) and Alzheimer's disease (AD) and, if so, whether these abnormalities are global in character or restricted to the temporal lobe. Volumetric MTI analysis of the whole brain and temporal and frontal lobes was performed in 25 patients with probable AD, in 13 patients with MCI, and in 28 controls. Magnetization transfer ratio (MTR) histograms were produced, from which we derived measures for structural brain damage and atrophy. The peak heights of the MTR histograms of MCI and AD patients were lower than those of controls for the whole brain and temporal and frontal lobes, reflecting structural brain damage. AD patients had more atrophy than controls in all regions that were studied. MCI patients differed from controls for temporal lobe atrophy only. Volumetric MTI demonstrates structural changes that are related to cognitive decline in large parts of the brain of AD patients. Moreover, structural changes also were observed in MCI patients, indicating that widespread brain damage can be demonstrated before patients are clinically demented. PMID- 12112050 TI - Mechanisms of acute cerebral infarctions in patients with middle cerebral artery stenosis: a diffusion-weighted imaging and microemboli monitoring study. AB - Although most therapeutic efforts and experimental stroke models focus on the concept of complete occlusion of the middle cerebral artery as a result of embolism from the carotid artery or cardiac chamber, relatively little is known about the stroke mechanism of intrinsic middle cerebral artery stenosis. Differences in stroke pathophysiology may require different strategies for prevention and treatment. We prospectively studied 30 consecutive acute ischemic stroke patients with middle cerebral artery stenosis detected by transcranial Doppler and magnetic resonance angiography. Patients underwent microembolic signal monitoring by transcranial Doppler and diffusion-weighted magnetic resonance imaging. Characteristics of acute infarct on diffusion-weighted magnetic resonance imaging were categorized according to the number (single or multiple infarcts) and the pattern of cerebral infarcts (cortical, border zone, or perforating artery territory infarcts). The data of microembolic signals and diffusion-weighted magnetic resonance imaging were assessed blindly and independently by separate observers. Diffusion-weighted magnetic resonance imaging showed that 15 patients (50%) had single acute cerebral infarcts and 15 patients had multiple acute cerebral infarcts. Among patients with multiple acute infarcts, unilateral, deep, chainlike border zone infarcts were the most common pattern (11 patients, 73%), and for single infarcts, penetrating artery infarcts were the most common (10 patients, 67%). Microembolic signals were detected in 10 patients (33%). The median number of microembolic signals per 30 minutes was 15 (range, 3-102). Microembolic signals were found in 9 patients with multiple infarcts and in 1 patient with a single infarct (p = 0.002, chi(2)). The number of microembolic signals predicted the number of acute infarcts on diffusion weighted magnetic resonance imaging (linear regression, adjusted R(2) =0.475, p < 0.001). Common stroke mechanisms in patients with middle cerebral artery stenosis are the occlusion of a single penetrating artery to produce a small subcortical lacuna-like infarct and an artery-to-artery embolism with impaired clearance of emboli that produces multiple small cerebral infarcts, especially along the border zone region. PMID- 12112049 TI - Sudomotor testing predicts the presence of neutralizing botulinum A toxin antibodies. AB - The increasing number of patients being treated with botulinum toxin A complex (BoNT/A) has led to a higher incidence of neutralizing anti-BoNT/A antibodies (ABAs). Because BoNT/A is known to inhibit sweating, here we report sudometry as a possibility for predicting the presence of ABA. Sixteen patients suffering from spasmodic torticollis were selected: in 2 patients, BoNT/A treatment continued to be effective, in 9 patients, the treatment effect was impaired, and in 5 patients, secondary treatment failure developed. BoNT/A (100 mouse units, Dysport; Ipsen Pharma, Berkshire, United Kingdom) was injected subcutaneously into the lateral calves. Sweating was visualized with iodine starch staining. In addition, quantitative sudomotor axon reflex testing was performed at the injection site. Individual ABA titers were determined with a mouse bioassay. Results of sudometry significantly correlated with the BoNT/A treatment success. The quantitative sudomotor axon reflex testing was 0.58 +/- 0.63 fraction of the normal mean in patients with treatment failure, 0.18 +/- 0.13 fraction of the normal mean in those who responded partially, and 0 in responders (p < 0.01). Accordingly, the areas of the anhidrotic skin after subcutaneous injections were 4.5 +/- 10.3 cm(2), 32.7 +/- 16.5 cm(2), and 62 cm(2) (p < 0.01). Discrimination analysis indicated that the presence of ABA (6 ABA-positive and 10 ABA-negative) could be predicted correctly in all patients from the results of sudometry. Therefore, sudometry is a useful tool for identifying patients with neutralizing ABAs and might be helpful for identifying reasons for BoNT/A treatment failure. PMID- 12112051 TI - Working memory after traumatic brain injury in children. AB - To investigate the effects of traumatic brain injury on working memory in children, we administered semantic (letter identity) and phonological (letter rhyme) N-back tasks to children who were on average 5 years post-mild (n = 54) or -severe (n = 26) traumatic brain injury and 44 typically developing children who were comparable in age. The correct detection of targets and false alarms were measured for each task. Memory load (which varied from 0 to 3 letters back) and age significantly affected the detection of targets and false alarms in both tasks. The severity of traumatic brain injury affected the correct detection of letters on the identity task and false alarms on the rhyme task. Traumatic brain injury severity also interacted with memory load in its effect on false alarms on the rhyme task. Traumatic brain injury results in impaired working memory and diminished inhibition in children. The N-back working memory task is feasible for administration to brain-injured children and potentially could be useful for studying brain activation associated with working memory and effects of drug therapy in this group of patients. PMID- 12112053 TI - Adenine nucleotide translocator 1 deficiency associated with Sengers syndrome. AB - Sengers syndrome is characterized by congenital cataracts, hypertrophic cardiomyopathy, mitochondrial myopathy, and lactic acidosis, but no abnormalities have been found with routine mitochondrial biochemical diagnostics (the determination of pyruvate oxidation rates and enzyme measurements). In immunoblot analysis, the protein content of the mitochondrial adenine nucleotide translocator 1 (ANT1) was found to be strongly reduced in the muscle tissues of two unrelated patients with Sengers syndrome. In addition, low residual adenine nucleotide translocator activity was detected upon the reconstitution of detergent-solubilized mitochondrial extracts from the patients' skeletal or heart muscle into liposomes. Sequence analysis and linkage analysis showed that ANT1 was not the primary genetic cause of Sengers syndrome. We propose that transcriptional, translational, or posttranslational events are responsible for the ANT1 deficiency associated with the syndrome. PMID- 12112052 TI - Interaction of cognitive aging and memory deficits related to epilepsy surgery. AB - Temporal lobe epilepsy surgery can cause significant memory impairment. This study was intended to examine whether surgery also could affect prognosis of memory in older age. Age regression of verbal memory was examined in 187 patients (before and 1 year after left temporal lobe surgery) and 264 healthy controls. Eighty patients underwent selective amygdalohippocampectomy, and 107 patients underwent anterior two-thirds temporal lobectomy. Amygdalohippocampectomy patients had mesiotemporal epilepsy; anterior two-thirds temporal lobectomy patients had more extramesial or diffuse seizure onset zones. Memory was assessed by word list learning for its more mesial (consolidation/retrieval) and more neocortical (learning) aspects. Patients showed significant preoperative memory impairment. Independent of seizure outcome and surgical approach, surgery had significant negative effects on learning and consolidation/retrieval. In the amygdalohippocampectomy group, preoperative and postoperative age regressions of learning and consolidation/retrieval were not different from those of controls. In the anterior two-thirds temporal lobectomy group, age regression of verbal learning became steeper after surgery, and consolidation/retrieval was negatively correlated with older age and later onset of epilepsy even before surgery. The data confirm that age regression of verbal memory in left temporal lobe epilepsy is similar to that in healthy controls. Both left anterior two-thirds temporal lobectomy and amygdalohippocampectomy worsen verbal learning and memory and bring patients closer to cognitive disability. Particularly in anterior two-thirds temporal lobectomy patients, surgery and reduced capacities for compensation cause acceleration of lifetime memory decline. The results support earlier and tailored epilepsy surgery and suggest that memory prognosis in older age should be considered if more extensive temporal resections would be inevitable. PMID- 12112054 TI - Angiostrongylus cantonensis infection mimicking a spinal cord tumor. AB - Angiostrongylus cantonensis is the most common cause of eosinophilic meningitis and meningoencephalitis. Almost all cases are self-limiting and are diagnosed by cerebrospinal fluid eosinophilia and enzyme-linked immunosorbent assay; pathology reports are restricted to postmortem samples from lethal cases. We report on what we believe is the first case of A. cantonensis infection diagnosed by biopsy in a living patient. The spinal cord was biopsied because of the unusual clinical presentation of a myelopathy without meningeal symptoms, together with a mass lesion that was clinically and radiologically diagnosed as a spinal cord tumor. PMID- 12112055 TI - Microscopic R2* mapping of reduced brain iron in the Belgrade rat. AB - R2* mapping has recently been used to detect iron overload in patients with movement disorders. We demonstrate here that this technique can also be used to detect reduced brain iron, as in the case of a missense mutation in the iron transporting protein divalent metal transporter 1. Surprisingly, we found that the same brain regions are affected (ie, the globus pallidus, substantia nigra, and cerebellar dentate nucleus); this suggests a much more extensive role for these structures in regulating overall brain iron homeostasis. Therefore, for the clinical monitoring of movement disorders for which normal brain iron homeostasis (either overload or reduction) may be implicated, R2* mapping appears to be well suited. PMID- 12112056 TI - Preferential loss of paternal 19q, but not 1p, alleles in oligodendrogliomas. AB - Portions of chromosomes 1p and 19q, which are frequently deleted in oligodendrogliomas, are subject to genomic imprinting, suggesting that the putative tumor suppressor genes could be monoallelically expressed. The parental origins of 1p and 19q allele losses were determined in 6 cases of pure oligodendroglioma. An equilibrated parental loss (3 maternal and 3 paternal) was found for 1p deletions. In contrast, 19q deletions always occurred on the paternal copy (p = 0.015). In this setting, a cloning strategy based on a search for homozygous deletion or mutation of the remaining allele would be appropriate for identifying the tumor suppressor gene located on 1p but inappropriate for identifying the presumably monoallelically expressed tumor suppressor gene located on 19q. PMID- 12112057 TI - Minicolumnar pathology in dyslexia. AB - The minicolumn is an anatomical and functional unit of the brain whose genesis accrues from germinal cell divisions in the ventricular zone of the brain. Disturbances in the morphometry of minicolumns have been demonstrated recently for both autism and Down's syndrome. We report minicolumnar abnormalities in the brain of a dyslexic patient. The corresponding developmental disturbance (ie, large minicolumns) could account for the perceptual errors observed in dyslexia. PMID- 12112058 TI - Anti-Hu antibodies in Merkel cell carcinoma. AB - Anti-Hu antibody is an antineuronal autoantibody found in a subset of patients with paraneoplastic neurological disease. The antibody was first associated with small cell carcinoma of the lung and is most often used as a marker for this neoplasm in patients presenting with suspected paraneoplastic syndromes. Here we report a patient with a multifaceted neurological disorder in the setting of Merkel cell carcinoma. The patient's serum contained antibodies against the Hu antigen, and the expression of the Hu antigen was demonstrated in the patient's tumor. PMID- 12112059 TI - Creutzfeldt-Jakob disease cluster in an Australian rural city. AB - Through the Australian National Creutzfeldt-Jakob Disease Registry, 6 pathologically confirmed sporadic cases were recognized over a 13-year period in persons who had been long-term residents of a moderate-sized rural city, whereas the expected number was 0.923. An extensive investigation could not find any point-source or case-to-case transmission links. This occurrence is highly statistically significant (p = 0.0027) when viewed in isolation and remains significant (p < 0.02) when only the cases that arose after the cluster was recognized were taken into account. However, a more conservative statistical analysis suggests that such a grouping could have arisen by chance in at least one population group of this size when the whole country is taken into consideration. PMID- 12112060 TI - In vivo study indicating loss of intracortical inhibition in tumor-associated epilepsy. AB - Transcranial magnetic stimulation was performed in 2 patients with focal motor seizures in the right hand caused by a circumscribed tumor process affecting the left precentral gyrus. In both cases, paired-pulse transcranial magnetic stimulation showed a loss of intracortical inhibition for interstimulus intervals of 2 to 4msec that was replaced by an enormous facilitation in the lesioned hand motor cortex. The uniform impairment of inhibitory mechanisms in epileptogenic tumors with different histologies suggests a common, nonspecific cause of tumor related epileptogenesis. PMID- 12112061 TI - Germline mosaicism of a novel mutation in lysosome-associated membrane protein-2 deficiency (Danon disease). AB - We identified a family with lysosome-associated membrane protein-2 deficiency (Danon disease) associated with a novel 883 ins-T mutation in the lysosome associated membrane protein-2 gene located at Xq24. Although the affected son and daughter carried the same mutation, it was not detected in their mother's peripheral blood or buccal cells; this indicated germline mosaicism. This is the first molecular evidence for germline mosaicism in Danon disease and has important implications for genetic counseling. PMID- 12112063 TI - Creatine supplementation results in elevated phosphocreatine/adenosine triphosphate (ATP) ratios in the calf muscle of athletes but not in patients with myopathies. PMID- 12112064 TI - Rabbit model of Guillain-Barre syndrome. PMID- 12112066 TI - Mistaken treatment of anterior ischemic optic neuropathy with interferon beta-1a. PMID- 12112069 TI - Blockade of blocking antibodies in Guillain-Barre syndromes: "unblocking" the mystery of action of intravenous immunoglobulin. PMID- 12112070 TI - Hereditary spastic paraplegia: the pace quickens. PMID- 12112072 TI - Novel locus for autosomal dominant pure hereditary spastic paraplegia (SPG19) maps to chromosome 9q33-q34. AB - Hereditary spastic paraplegia is a clinically and genetically heterogeneous disorder characterized by progressive spasticity of the lower limbs. Seven loci for autosomal dominant pure hereditary spastic paraplegia (ADPHSP) have already been mapped on chromosomes 14q, 2p, 15q, 8p, 12q, 19q, and 2q. We report on an Italian family affected by ADPHSP for which we excluded linkage with the known loci and performed a genome-wide search. Linkage analysis and haplotype construction permitted the identification of a novel ADPHSP locus on the long arm of chromosome 9, designated SPG19. The phenotype was characterized by late onset (range, 36-55 years) and mild disability, with only 1 patient bound to a wheelchair after 31 years of disease. Urinary disturbances (urgency and/or incontinence) were always present, even in young patients with a short disease history. PMID- 12112071 TI - Intravenous immunoglobulins neutralize blocking antibodies in Guillain-Barre syndrome. AB - Intravenous immunoglobulin (IVIg) treatment ameliorates the course of Guillain Barre syndrome (GBS), but its specific mode of action is unknown. We attempted to delineate the effect of IVIg on neuromuscular blocking antibodies in GBS. A total of seven GBS serum samples were examined for blocking antibodies and the effect of IVIg with a macro-patch-clamp technique in mouse hemidiaphragms. First, serum was tested before and after treatment with IVIg. Second, we investigated with coincubation experiments whether the IVIg was capable of neutralizing neuromuscular blocking antibodies in GBS serum or affinity-purified immunoglobulin G (IgG) fractions. Finally, the mechanism of the neutralizing effect was studied by the coincubation of active blocking GBS IgG with Fab and Fc fragments prepared from IVIg. All GBS sera (two adults and two children) and GBS IgG fractions (three adults) taken before treatment with IVIg blocked evoked quantal release by approximately 90%. Blocking activity was markedly reduced in sera obtained after treatment with IVIg. Coincubation of the pretreatment blocking serum with the posttreatment serum, or with the IVIg preparation used for treatment, reduced the blocking activity of the pretreatment GBS serum. When GBS IgG was coincubated with IVIg, the blocking activity of GBS IgG was diminished dose-dependently. Monovalent and divalent Fab fragments prepared from the IVIg were as effective as whole IVIg, but Fc fragments were ineffective. Therapeutic IVIg is capable of neutralizing neuromuscular blocking antibodies in GBS by a dose-dependent, antibody-mediated mechanism. This may, in part, explain its therapeutic efficacy. PMID- 12112073 TI - Evolution of the response to levodopa during the first 4 years of therapy. AB - The short-duration response, long-duration response, and dyskinetic response to levodopa change during long-term levodopa therapy. How these responses evolve, and which changes contribute to the emergence of motor fluctuations, remain unclear. We studied 18 subjects with Parkinson's disease before they began levodopa therapy and after 6, 12, 24, and 48 months of long-term levodopa therapy. The responses to 2-hour levodopa infusions after overnight and after 3 days of levodopa withdrawal were studied from 6 months onward. The mean magnitude of the short-duration response and the long-duration response measured after overnight without antiparkinsonian medications did not change during the 4 years. However, after 3 days without levodopa, it was apparent that the short-duration response magnitude was progressively increasing (p < 0.0001) and that the long duration response was decaying more rapidly (p = 0.0004). The short-duration response magnitude at 4 years was inversely related to the long-duration-response magnitude (p = 0.022), suggesting that the long-duration response was one determinant of the short-duration-response magnitude. Dyskinesia increased progressively in severity during the study (p = 0.013). The duration of the short duration response and dyskinesia did not change during the 4 years. Subject reports of motor fluctuations tended to be associated with a large short-duration response (p = 0.054). We suggest that a larger long-duration response, rather than a shortened one, is more important to the development of fluctuations. Improving the baseline or practical-off motor function to reduce the magnitude of the short-duration response may be a strategy to treat fluctuations. PMID- 12112075 TI - Relapsed and late-onset Nipah encephalitis. AB - An outbreak of infection with the Nipah virus, a novel paramyxovirus, occurred among pig farmers between September 1998 and June 1999 in Malaysia, involving 265 patients with 105 fatalities. This is a follow-up study 24 months after the outbreak. Twelve survivors (7.5%) of acute encephalitis had recurrent neurological disease (relapsed encephalitis). Of those who initially had acute nonencephalitic or asymptomatic infection, 10 patients (3.4%) had late-onset encephalitis. The mean interval between the first neurological episode and the time of initial infection was 8.4 months. Three patients had a second neurological episode. The onset of the relapsed or late-onset encephalitis was usually acute. Common clinical features were fever, headache, seizures, and focal neurological signs. Four of the 22 relapsed and late-onset encephalitis patients (18%) died. Magnetic resonance imaging typically showed patchy areas of confluent cortical lesions. Serial single-photon emission computed tomography showed the evolution of focal hyperperfusion to hypoperfusion in the corresponding areas. Necropsy of 2 patients showed changes of focal encephalitis with positive immunolocalization for Nipah virus antigens but no evidence of perivenous demyelination. We concluded that a unique relapsing and remitting encephalitis or late-onset encephalitis may result as a complication of persistent Nipah virus infection in the central nervous system. PMID- 12112074 TI - Peroxisome proliferator-activated receptor-gamma agonists prevent experimental autoimmune encephalomyelitis. AB - The development of clinical symptoms in multiple sclerosis and its animal model experimental autoimmune encephalomyelitis (EAE) involves T-cell activation and migration into the central nervous system, production of glial-derived inflammatory molecules, and demyelination and axonal damage. Ligands of the peroxisome proliferator-activated receptor (PPAR) exert anti-inflammatory effects on glial cells, reduce proliferation and activation of T cells, and induce myelin gene expression. We demonstrate in two models of EAE that orally administered PPARgamma ligand pioglitazone reduced the incidence and severity of monophasic, chronic disease in C57BL/6 mice immunized with myelin oligodendrocyte glycoprotein peptide and of relapsing disease in B10.Pl mice immunized with myelin basic protein. Pioglitazone also reduced clinical signs when it was provided after disease onset. Clinical symptoms were reduced by two other PPARgamma agonists, suggesting a role for PPARgamma activation in protective effects. The suppression of clinical signs was paralleled by decreased lymphocyte infiltration, lessened demyelination, reduced chemokine and cytokine expression, and increased inhibitor of kappa B (IkB) expression in the brain. Pioglitazone also reduced the antigen-dependent interferon-gamma production from EAE-derived T cells. These results suggest that orally administered PPARgamma agonists could provide therapeutic benefit in demyelinating disease. PMID- 12112076 TI - Periaxin mutations cause a broad spectrum of demyelinating neuropathies. AB - Previous studies have demonstrated that apparent loss-of-function mutations in the periaxin gene cause autosomal recessive Dejerine-Sottas neuropathy or severe demyelinating Charcot-Marie-Tooth disease. In this report, we extend the associated phenotypes with the identification of two additional families with novel periaxin gene mutations (C715X and R82fsX96) and provide detailed neuropathology. Each patient had marked sensory involvement; two siblings with a homozygous C715X mutation had much worse sensory impairment than motor impairment. Despite early disease onset, these siblings with the C715X mutation had relatively slow disease progression and adult motor impairment typical of classic demyelinating Charcot-Marie-Tooth neuropathy. In contrast, a patient with the homozygous R82fsX96 mutation had a disease course consistent with Dejerine Sottas neuropathy. The neuropathology of patients in both families was remarkable for demyelination, onion bulb and occasional tomacula formation with focal myelin thickening, abnormalities of the paranodal myelin loops, and focal absence of paranodal septate-like junctions between the terminal loops and axon. Our study indicates a prominent sensory neuropathy resulting from periaxin gene mutations and suggests a role for the carboxyl terminal domain of the periaxin protein. PMID- 12112077 TI - Association between high-density lipoprotein and cognitive impairment in the oldest old. AB - Low high-density lipoprotein cholesterol is associated with an increased risk for cardiovascular disease and stroke. At the same time, cardiovascular disease and stroke are important risk factors for dementia. We assessed the association between total and fractionated cholesterol and cognitive impairment and explored whether observed associations were dependent on or independent of atherosclerotic disease. In a population-based study, total cholesterol, triglycerides, low density lipoprotein cholesterol, and high-density lipoprotein cholesterol were measured in 561 subjects 85 years old and grouped in three equal strata representing decreasing serum concentrations. History of cardiovascular disease and stroke was determined. All subjects completed the Mini-Mental State Examination (MMSE), and the presence of dementia was determined. Median MMSE scores were significantly lower in subjects with low high-density lipoprotein cholesterol (25 points vs 27 points, p < 0.001). No differences in MMSE scores were found for other lipids and lipoproteins. MMSE scores in subjects with and without cardiovascular disease were 26 and 27 points (p = 0.007), respectively, and in subjects with and without stroke were 21 and 26 points (p < 0.001), respectively. The associations between low MMSE scores and low high-density lipoprotein cholesterol remained significant after subjects with cardiovascular disease or stroke were excluded. In a comparison of subjects with low high density lipoprotein cholesterol with subjects with high high-density lipoprotein cholesterol, the odds ratio for dementia was 2.3 (95% confidence interval, 1.2 4.3), and in subjects without cardiovascular disease or stroke, it was 3.7 (95% confidence interval, 1.3-10.1). All odds ratios were unaffected by education, low density lipoprotein cholesterol, triglycerides, and survival. Low high-density lipoprotein cholesterol is associated with cognitive impairment and dementia. At least part of the association between high-density lipoprotein cholesterol and cognitive function is independent of atherosclerotic disease. PMID- 12112078 TI - Combined effect of age and severity on the risk of dementia in Parkinson's disease. AB - Age and severity of extrapyramidal signs have been consistently associated with incident dementia in Parkinson's disease. We evaluated the separate and combined effects of age and severity of extrapyramidal signs on the risk of incident dementia in Parkinson's disease in the setting of a population-based prospective cohort study. Age and the total Unified Parkinson's Disease Rating Scale motor score at baseline evaluation were dichotomized at the median. Four groups of Parkinson's disease patients were defined: younger age/low severity (reference), younger age/high severity, older age/low severity, and older age/high severity. Risk ratios for incident dementia were calculated with Cox proportional hazards models controlling for gender, education, ethnicity, and duration of Parkinson's disease. Of 180 patients, 52 (28.9%) became demented during a mean follow-up period of 3.6 +/- 2.2 years. The median age at baseline of the Parkinson's disease patients was 71.8 years (range, 38.5-95.9 years), and the median total Unified Parkinson's Disease Rating Scale motor score was 24 (range, 2-65). The group with older age/high severity had a significantly increased risk of incident dementia (relative risk, 9.7; 95% confidence interval, 3.9-24.4) compared with the group with younger age/low severity (reference), whereas the groups with older age/low severity (relative risk, 1.6; 95% confidence interval, 0.5-4.8) and younger age/high severity (relative risk, 1.2; 95% confidence interval, 0.5-3.2) did not. These findings suggest that the increased risk of incident dementia in Parkinson's disease associated with age and severity of extrapyramidal signs is related primarily to their combined effect rather than separate effects. PMID- 12112079 TI - Sporadic Pick's disease: a tauopathy characterized by a spectrum of pathological tau isoforms in gray and white matter. AB - Pick's disease is characterized neuropathologically by distinct tau immunoreactive intraneuronal inclusions known as Pick bodies and by insoluble tau proteins with predominantly three microtubule-binding repeat tau isoforms. However, recent immunohistochemical studies showed that the antibody specific for exon 10, which encodes the fourth microtubule-binding repeat, detected other tau lesions in Pick's disease. To better define the spectrum of tau pathology in Pick's disease, we used biochemical, immunohistochemical, and ultrastructural techniques to analyze the tau isoform composition in 14 Pick's disease brains. Western blot analysis showed that both three and four microtubule-binding repeat pathological tau isoforms are present in gray and white matter of various brain regions. Using phosphorylation-dependent anti-tau antibodies, we show that major tau phosphoepitopes are present in sarcosyl-insoluble gray and white matter regions of Pick's disease brains. Also, for the first time to our knowledge, we demonstrated that isoforms with four microtubule-binding repeat tau isoforms are present in Pick bodies from selected brains. Isolated tau filaments were straight or twisted and formed by three microtubule-binding repeat or four microtubule binding repeat tau isoforms. Major tau phosphorylation-dependent and exon 10 specific epitopes were present in filaments. Therefore, Pick's disease is characterized by an accumulations of Pick bodies in the hippocampal region and cortex as well as the presence of three and four microtubule-binding repeat tau pathology in both cortical gray and white matter that distinguish this tauopathy from other neurodegenerative disorders. PMID- 12112080 TI - Partial epilepsy with pericentral spikes: a new familial epilepsy syndrome with evidence for linkage to chromosome 4p15. AB - The genetic analysis of simple Mendelian epilepsies remains a key strategy in advancing our understanding of epilepsy. In this article, we describe a new family epilepsy syndrome, partial epilepsy with pericentral spikes, which we map to chromosome 4p15. We distinguish it clinically, electrophysiologically, and genetically from previously described Mendelian epilepsies. The family described is a large Brazilian kindred of Portuguese extraction in which affected family members manifest a variety of seizure types, including hemiclonic, hemitonic, generalized tonic-clonic, simple partial (stereotyped episodes of epigastric pain), and complex partial seizures consistent with temporal lobe epilepsy. The syndrome is benign, either requiring no treatment or responding to a single antiepileptic medication. Seizure onset is in the first or second decades of life, with seizures in individuals up to the age of 71 years and documented encephalogram changes up to the age of 30 years. A key feature of partial epilepsy with pericentral spikes is a characteristic encephalogram abnormality of spikes or sharp waves in the pericentral region (centroparietal, centrofrontal, or centrotemporal). This distinctive encephalogram abnormality of pericentral spikes unites these several seizure types into a discrete family epilepsy syndrome. As with other familial epilepsies, the inherited nature of this new syndrome may be overlooked because of the variability in penetrance and seizure types among affected family members. PMID- 12112082 TI - Platelet-activating factor acetylhydrolase activity in cerebrospinal fluid of children with acute systemic or neurological illness. AB - Platelet-activating factor acetylhydrolase was analyzed in cerebrospinal fluid samples taken from children with a variety of neurological conditions (85 patients; mean age, 3.8 years) to determine it is involved in the defense mechanism against the toxic effect of inflammatory mediators in the central nervous system. A significant increase in cerebrospinal fluid activity was seen in the patients with meningitis and acute febrile illness in comparison with the control subjects. The activity was also significantly higher in the patients with meningitis than in the patients with inflammatory neurological diseases. In addition, the biochemical profile of cerebrospinal fluid platelet-activating factor acetylhydrolase was different from other known acetylhydrolases. These findings suggest that cerebrospinal fluid platelet-activating factor acetylhydrolase activity may be a sensitive marker of the host response to central nervous system infections. PMID- 12112081 TI - A recessive form of central core disease, transiently presenting as multi minicore disease, is associated with a homozygous mutation in the ryanodine receptor type 1 gene. AB - Multi-minicore disease is an autosomal recessive congenital myopathy characterized by the presence of multiple, short-length core lesions (minicores) in both muscle fiber types. These lesions being nonspecific and the clinical phenotype being heterogeneous, multi-minicore disease boundaries remain unclear. To identify its genetic basis, we performed a genome-wide screening in a consanguineous Algerian family in which three children presented in infancy with moderate weakness predominant in axial muscles, pelvic girdle and hands, joint hyperlaxity (hand involvement phenotype), and multiple minicores. We mapped the disease to chromosome 19q13 in this family and, subsequently, in three additional families showing a similar phenotype, with a maximum LOD score of 5.19 for D19S570. This locus was excluded in 16 other multi-minicore disease families with predominantly axial weakness, scoliosis, and respiratory insufficiency ("classical" phenotype). In the Algerian family, we identified a novel homozygous missense mutation (P3527S) in the ryanodine receptor type 1 gene, a positional candidate gene responsible for the autosomal dominant congenital myopathy central core disease. New muscle biopsies from the three patients at adulthood demonstrated typical central core disease with rods; no cores were found in the healthy parents. This subgroup of families linked to 19q13 represents the first variant of central core disease with genetically proven recessive inheritance and transient presentation as multi-minicore disease. PMID- 12112083 TI - Activity of the hypothalamic-pituitary-adrenal axis in multiple sclerosis: correlations with gadolinium-enhancing lesions and ventricular volume. AB - The known interaction between the immune system and the hypothalamic-pituitary adrenal axis led us to explore the interrelation between magnetic resonance imaging findings and the hypothalamic-pituitary-adrenal axis activity in 53 multiple sclerosis patients. The cortisol release induced by the dexamethasone corticotropin-releasing hormone test was negatively associated with the presence and number of gadolinium-enhancing lesions and positively associated with ventricular size. This finding suggests a protective effect of the hypothalamic- pituitary-adrenal drive on acute lesional inflammation in multiple sclerosis, probably by limiting immune overshoot. In contrast, the nature of the correlation between hypothalamic-pituitary-adrenal hyperdrive and brain atrophy remains to be determined. PMID- 12112084 TI - Striatal monoaminergic terminals in Lewy body and Alzheimer's dementias. AB - Vesicular monoamine transporter type 2 and benzodiazepine binding site expressions were examined with quantitative autoradiography in postmortem striata from 19 patients with dementia with Lewy bodies, seven patients with dementia with Lewy bodies and Alzheimer's disease, 12 patients with Alzheimer's disease, and eight neurologically normal subjects. Striatal vesicular monoamine transporter type 2 expression in dementia with Lewy bodies and in dementia with Lewy bodies plus Alzheimer's disease was reduced significantly, indicating degeneration of nigrostriatal projections. Striatal benzodiazepine binding site expression was unchanged, indicating preserved intrinsic striatal neuropil. Vesicular monoamine transporter type 2 and benzodiazepine binding site expressions were each preserved in Alzheimer's disease striatum. We conclude that dementia with Lewy bodies may be distinguished from Alzheimer's disease by postmortem examination or by future in vivo measurements of the striatal vesicular monoamine transporter type 2. PMID- 12112085 TI - Human herpesvirus 6 encephalitis associated with hypersensitivity syndrome. AB - Hypersensitivity syndrome, a serious systematic reaction to a limited number of drugs, is associated with the reactivation of human herpesvirus 6. A 56-year-old man developed acute limbic encephalitis followed by multiple organ failure during the course of toxic dermatitis induced by aromatic anticonvulsants. The clinical features of skin eruptions, high fever, eosinophilia, and atypical lymphocytosis were compatible with drug hypersensitivity syndrome. The patient showed seroconversion for human herpesvirus 6, and polymerase chain reaction detected human herpesvirus 6 DNA in the cerebrospinal fluid. To our knowledge, this is the first report of human herpesvirus 6 encephalitis associated with hypersensitivity syndrome. PMID- 12112086 TI - Mitochondrial DNA nucleotide changes C14482G and C14482A in the ND6 gene are pathogenic for Leber's hereditary optic neuropathy. AB - A novel mitochondrial DNA nucleotide transversion, C14482A (M64I), different from the previously reported C14482G (M64I), was found to cause Leber's hereditary optic neuropathy with visual recovery in an Italian family. These equivalent changes are the fifth pathogenic mutation for pure Leber's hereditary optic neuropathy. This confirms that the ND6 gene of complex I is a mutational hot spot and suggests that different amino acid substitutions at residue 64, as induced by C14482G or C14482A (M64I) and the common T14484C (M64V) mutations, are associated with visual recovery. PMID- 12112087 TI - Brain proteasomal function in sporadic Parkinson's disease and related disorders. AB - Because genetic defects relating to the ubiquitin-proteasome system were reported in familial parkinsonism, we evaluated proteasomal function in autopsied brains with sporadic Parkinson's disease. We found that proteasome peptidase activities in a fraction specific to the proteasome were preserved in five brain areas (including the striatum) of Parkinson's disease where neuronal loss is not observed. Striatal protein levels of two proteasome subunits were normal in Parkinson's disease but reduced mildly in disease controls (multiple system atrophy). Our brain data suggest that a systemic, global disturbance in the catalytic activity and degradation ability of the proteasome itself is unlikely to explain the cause of Parkinson's disease. PMID- 12112088 TI - Increased expression of the amyloid precursor beta-secretase in Alzheimer's disease. AB - Beta-secretase cleavage represents the first step in the generation of Abeta polypeptides and initiates the amyloid cascade that leads to neurodegeneration in Alzheimer's disease. By comparative Western blot analysis, we show a 2.7-fold increase in protein expression of the beta-secretase enzyme BACE in the brain cortex of Alzheimer's disease patients as compared to age-matched controls. Similarly, we found the levels of the amyloid precursor protein C-terminal fragment produced by beta-secretase to be increased by nearly twofold in Alzheimer's disease cortex. PMID- 12112090 TI - Thrombolysis induces cerebral hemorrhage in a mouse model of cerebral amyloid angiopathy. AB - We studied the impact of cerebral amyloid angiopathy on tissue plasminogen activator-induced cerebral hemorrhages in APP23 transgenic mice. Results show that the intravenous administration of tissue plasminogen activator in APP23 mice leads to an increase in cerebral amyloid angiopathy-associated microhemorrhages and can provoke parenchymal and subarachnoidal hematomas. We conclude that cerebral amyloid angiopathy is a risk factor for cerebral hemorrhage caused by tissue plasminogen activator administration in mice and stress the need for more comprehensive studies of the relation between cerebral amyloid angiopathy and tissue plasminogen activator-induced cerebral hemorrhages in elderly and Alzheimer's disease patients. PMID- 12112089 TI - High headache prevalence among women with hemochromatosis: the Nord-Trondelag health study. AB - In this large cross-sectional population-based study, 51,272 persons responded to a headache questionnaire and were screened for hemochromatosis. Phenotypic hemochromatosis and the C282Y/C282Y genotype were both associated with an 80% increase in headache prevalence evident only among women. The reason for this association is unclear, but one may speculate that iron overload alters the threshold for triggering a headache by disturbing neuronal function. PMID- 12112091 TI - Toxicity of beta-amyloid vaccination in patients with Alzheimer's disease. PMID- 12112092 TI - Further evidence that SPG3A gene mutations cause autosomal dominant hereditary spastic paraplegia. PMID- 12112093 TI - Association of interleukin-1 polymorphisms with Alzheimer's disease in Australia. PMID- 12112094 TI - Prion protein deposits match magnetic resonance imaging signal abnormalities in Creutzfeldt-Jakob disease. PMID- 12112097 TI - Does the right make it right? Questions about recovery of language after stroke. PMID- 12112098 TI - Altered reproductive endocrine regulation in men with epilepsy: implications for reproductive function and seizures. PMID- 12112099 TI - Sickle cell disease: the neurological complications. AB - The genetic cause of sickle cell disease has been known for decades, yet the reasons for its clinical variability are not fully understood. The neurological complications result from one point mutation that causes vasculopathy of both large and small vessels. Anemia and the resultant cerebral hyperemia produce conditions of hemodynamic insufficiency. Sickled cells adhere to the endothelium, contributing to a cascade of activated inflammatory cells and clotting factors, which result in a nidus for thrombus formation. Because the cerebrovascular reserve becomes exhausted, the capacity for compensatory cerebral mechanisms is severely limited. There is evidence of small-vessel sludging, and a relative deficiency of nitric oxide in these vessels further reduces compensatory vasodilatation. Both clinical strokes and silent infarcts occur, affecting motor and cognitive function. New data suggest that, in addition to sickle cell disease, other factors, both environmental (eg, hypoxia and inflammation) and genetic (eg, mutations resulting in thrombogenesis), may contribute to a patient's stroke risk. The stroke risk is polygenic, and sickle cell disease can be considered a model for all cerebrovascular disease. This complex disease underscores the potential intellectual and practical distance between the determination of molecular genetics and effective clinical application and therapeutics. PMID- 12112100 TI - A physiological change in the homotopic cortex following left posterior temporal lobe infarction. AB - We investigated the relationship between cerebral activity (measured with positron emission tomography) and word rate in normal subjects and aphasic patients listening to monosyllabic words at rates up to those encountered in normal speech. By measuring the slope of the regression of the individual activity-word rate responses in the temporal cortex in normal subjects, we identified a functional asymmetry in the posterior superior temporal sulcus. The response was greater (steeper) on the left. Using the same study design, we identified a steeper activity-word rate response in the right posterior superior temporal sulcus of patients who had recovered single-word auditory comprehension after infarction of the left posterior temporal cortex. This response was significantly different from that observed in both normal subjects and a group of patients with left hemisphere infarcts that spared the posterior temporal cortex. The results demonstrated a change in physiological responsiveness in the contralateral homotopic cortex after posterior temporal infarction. PMID- 12112101 TI - Interictal and postictal alterations of pulsatile secretions of luteinizing hormone in temporal lobe epilepsy in men. AB - Mesial temporal lobe epilepsy has been associated with abnormalities of reproductive physiology, but the mechanisms of hormonal dysregulation are not clear. Chronic effects of the epileptic state and the acute impact of seizures could alter hypothalamic function, which is represented by the downstream pulsatile secretion of luteinizing hormone. This study evaluates the interictal and postictal secretion of luteinizing hormone in mesial temporal lobe epilepsy. We characterized luteinizing hormone secretion in patients with mesial temporal lobe epilepsy during two 24-hour epochs: an interictal baseline and a postictal interval initiated by an electrographically confirmed spontaneous seizure. Males, rather than females, were studied so that menstrual cycles could not account for differences between epochs. Blood luteinizing hormone and prolactin (as a positive control) were measured every 10 minutes. Deconvolution analysis defined luteinizing hormone secretion in terms of interpulse interval, amplitude, and mass. Approximate entropy quantitated relative degradation in the orderliness of serial luteinizing hormone release. Interictal baseline epochs were compared to those of healthy controls with unpaired Student's t tests and between interictal and postictal epochs within epileptic patients with paired t tests. Ten epileptic men completed both interictal and postictal epochs. Interictally, seizure patients had lower mean concentrations, slower pulse rates, and higher peak amplitudes than healthy male controls. Within epileptic patients, mean interpulse interval, pulse amplitude, and pulse mass were not affected by the occurrence of seizures, whereas the orderliness of pulse mass decreased postictally. Acute seizures induced timing irregularity in luteinizing hormone secretion, whereas chronic epilepsy was associated with changes in luteinizing hormone pulse frequency, amplitude, and mass. Altered timing and regularity of neuroendocrine pulse patterns may underlie other disorders of homeostasis in mesial temporal lobe epilepsy. PMID- 12112103 TI - The prevalence of narcolepsy among Chinese in Hong Kong. AB - Narcolepsy is a lifelong, crippling sleep disorder. Although the discovery of the hypocretin system has been a breakthough in genetics, the epidemiological aspects of narcolepsy remain elusive. Ethnic predisposition was suggested to partially account for the 2,500-fold difference in the reported prevalence rates of narcolepsy between Japanese (0.59%) and Israeli Jews (0.00023%). We carried out a general population study, conducting a random telephone survey with a structured questionnaire, which included a validated screening instrument (a Chinese version of the Ullanlinna Narcolepsy Scale). It was followed by clinical-polysomnographic HLA confirmation of the subjects determined to be positive for narcolepsy based on the questionnaire. Of 9,851 subjects interviewed, 28 subjects (0.28%, 58% female) were screened positive. Ninety percent had a second detailed interview, 64% had HLA typing, and over half of them had a sleep assessment. Only three subjects were found to have genuine narcolepsy. The most common nonnarcolepsy diagnoses were sleep apnea syndrome and sleep-wake schedule disorder. The prevalence rate of narcolepsy in Southern (Hong Kong) Chinese was found to be 0.034% (95% confidence interval = 0.010-0.117%). All available narcoleptic subjects were HLA DRB1-1501 positive and 50% were DQB1-0602 positive. The prevalence rate of narcolepsy among Chinese is comparable to the rates for other populations in studies with stringent epidemiological designs, suggesting that major cross-ethnic differences in the prevalence rates of narcolepsy previously reported likely resulted from methodological limitations. PMID- 12112102 TI - Alzheimer's neurofibrillary pathology and the spectrum of cognitive function: findings from the Nun Study. AB - The development of interventions designed to delay the onset of dementia highlights the need to determine the neuropathologic characteristics of individuals whose cognitive function ranges from intact to demented, including those with mild cognitive impairments. We used the Braak method of staging Alzheimer's disease pathology in 130 women ages 76-102 years who were participants in the Nun Study, a longitudinal study of aging and Alzheimer's disease. All participants had complete autopsy data and were free from neuropathologic conditions other than Alzheimer's disease lesions that could affect cognitive function. Findings showed a strong relationship between Braak stage and cognitive state. The presence of memory impairment was associated with more severe Alzheimer's disease pathology and higher incidence of conversion to dementia in the groups classified as having mild or global cognitive impairments. In addition to Braak stage, atrophy of the neocortex was significantly related to the presence of dementia. Our data indicate that Alzheimer's neurofibrillary pathology is one of the neuropathologic substrates of mild cognitive impairments. Additional studies are needed to help explain the variability in neuropathologic findings seen in individuals whose cognitive performance falls between intact function and dementia. PMID- 12112105 TI - Sensory training for patients with focal hand dystonia. AB - Some patients with focal hand dystonia have impaired sensory perception. Abnormal sensory processing may lead to problems with fine motor control. For patients with focal hand dystonia who demonstrate sensory dysfunction, sensory training may reverse sensory impairment and dystonic symptoms. We studied the efficacy of learning to read braille as a method of sensory training for patients with focal hand dystonia. Sensory spatial discrimination was evaluated in 10 patients who had focal hand dystonia and 10 age- and gender-matched controls with a spatial acuity test (JVP domes were used in this test). Clinical dystonia evaluation included the Fahn dystonia scale and time needed to write a standard paragraph. Each individual was trained in braille reading at the grade 1 level for 8 weeks, between 30 and 60 minutes daily, and was monitored closely to ensure that reading was done regularly. Both controls and patients demonstrated improvement on the spatial acuity test. Patients showed a significant mean difference from baseline to 8 weeks on the Fahn dystonia scale. Sixty percent of the patients shortened the time they needed to write a standard paragraph. Improved sensory perception correlated positively with improvement on the Fahn dystonia scale. We conclude that training in braille reading improves deficits in spatial discrimination and decreases disability in patients with focal hand dystonia. PMID- 12112104 TI - Mapping of autosomal recessive chronic distal spinal muscular atrophy to chromosome 11q13. AB - Distal spinal muscular atrophy is a heterogeneous group of neuromuscular disorders caused by progressive anterior horn cell degeneration and characterized by progressive motor weakness and muscular atrophy, predominantly in the distal parts of the limbs. Here we report on chronic autosomal recessive distal spinal muscular atrophy in a large, inbred family with onset at various ages. Because this condition had some of the same clinical features as spinal muscular atrophy with respiratory distress, we tested the disease gene for linkage to chromosome 11q and mapped the disease locus to chromosome 11q13 in the genetic interval that included the spinal muscular atrophy with respiratory distress gene (D11S1889 D11S1321, Z(max) = 4.59 at theta = 0 at locus D11S4136). The sequencing of IGHMBP2, the human homologue of the mouse neuromuscular degeneration gene (nmd) that accounts for spinal muscular atrophy with respiratory distress, failed to detect any mutation in our chronic distal spinal muscular atrophy patients, suggesting that spinal muscular atrophy with respiratory distress and chronic distal spinal muscular atrophy are caused by distinct genes located in the same chromosomal region. In addition, the high intrafamilial variability in age at onset raises the question of whether nonallelic modifying genes could be involved in chronic distal spinal muscular atrophy. PMID- 12112106 TI - Effects of ovarian hormones on human cortical excitability. AB - Ovarian steroids appear to alter neuronal function in women, but direct physiological evidence is lacking. In animals, estradiol enhances excitatory neurotransmission. Progesterone-derived neurosteroids increase GABAergic inhibition. The effect of weak transcranial magnetic stimulation of the motor cortex on the motor evoked potential (MEP) from transcranial magnetic stimulation given milliseconds later is changed by GABAergic and glutamatergic agents. Using this technique previously, we showed more inhibition in the luteal phase relative to the midfollicular menstrual phase, which is consistent with a progesterone effect. To detect the effects of estradiol, we have now divided the follicular phase. We tested 14 healthy women during the early follicular (low estradiol, low progesterone), late follicular (high estradiol, low progesterone), and luteal (high estradiol, high progesterone) phases, with interstimulus intervals from 2 to 10msec (10 trials at each interval and 40 unconditioned trials). We calculated the ratio of the conditioned MEP at each interval to the mean unconditioned MEP: the higher the ratio, the less inhibition and the more facilitation caused by the first stimulus. The combined ratios increased significantly from the early follicular phase to the late follicular phase and then decreased again in the luteal phase. These findings demonstrate an excitatory neuronal effect associated with estradiol and confirm our earlier finding of inhibition associated with progesterone. PMID- 12112107 TI - Impact of sustained deprenyl (selegiline) in levodopa-treated Parkinson's disease: a randomized placebo-controlled extension of the deprenyl and tocopherol antioxidative therapy of parkinsonism trial. AB - Deprenyl (selegiline) delays the need for levodopa therapy in patients with early Parkinson's disease, but the long-term benefits of this treatment remain unclear. During 1987 to 1988, 800 patients with early Parkinson's disease were randomized in the Deprenyl and Tocopherol Antioxidative Therapy of Parkinsonism trial to receive deprenyl, tocopherol, combined treatments, or a placebo and were then placed on active deprenyl (10mg/day). A second, independent randomization was carried out in early 1993 for 368 subjects who by that time had required levodopa and who had consented to continuing the deprenyl treatment (D subjects) or changing to a matching placebo (P subjects) under double-blind conditions. The first development of wearing off, dyskinesias, or on-off motor fluctuations was the prespecified primary outcome measure. During the average 2-year follow-up, there were no differences between the treatment groups with respect to the primary outcome measure (hazard ratio, 0.87; 95% confidence interval, 0.63, 1.19; p = 0.38), withdrawal from the study, death, or adverse events. Although 34% of D subjects developed dyskinesias and only 19% of P subjects did (p = 0.006), only 16% of D subjects developed freezing of gait but 29% of P subjects did (p = 0.0003). Decline in motor performance was less in D subjects than P subjects. Levodopa-treated Parkinson's disease patients who had been treated with deprenyl for up to 7 years, compared with patients who were changed to a placebo after about 5 years, experienced slower motor decline and were more likely to develop dyskinesias but less likely to develop freezing of gait. PMID- 12112108 TI - Effects of oxcarbazepine on sodium concentration and water handling. AB - Oxcarbazepine, a keto-analogue of carbamazepine, was recently approved in the United States for the treatment of seizures of partial onset. Some patients treated with oxcarbazepine showed the development of hyponatremia, which in most instances was asymptomatic. Understanding the mechanisms by which oxcarbazepine can lead to a reduction of serum sodium levels could have therapeutic implications for the few patients in whom symptomatic hyponatremia develops. In this study, we evaluated sodium and water handling in patients with epilepsy and in healthy subjects titrated over 3 weeks to a maximum daily oxcarbazepine dose of 2,400mg. All subjects were evaluated in a hospital setting after an overnight fast and after an acute water-load test performed before oxcarbazepine exposure and after maintenance on the medication for 3 weeks. Before oxcarbazepine exposure, the percentage of water load excreted was normal as both groups excreted more than 80% of the administered water load. After the intake of oxcarbazepine, the water load resulted in a reduction of the serum sodium and free water clearance without a concomitant increase in the arginine vasopressin serum levels. Most subjects in both groups failed to excrete 80% or more of the water load, suggesting that the effect of oxcarbazepine is physiological. We found that, after the water load, serum sodium and free water clearance were diminished in both groups without a concomitant increase in the arginine vasopressin serum levels. These findings indicate that oxcarbazepine-induced hyponatremia is not attributable to the syndrome of inappropriate secretion of antidiuretic hormone. Possible mechanisms include a direct effect of oxcarbazepine on the renal collecting tubules or an enhancement of their responsiveness to circulating antidiuretic hormone. PMID- 12112109 TI - Role of parkin mutations in 111 community-based patients with early-onset parkinsonism. AB - Early-onset parkinsonism is frequently reported in connection with mutations in the parkin gene. In this study, we present the results of extensive genetic screening for parkin mutations in 111 community-derived early-onset parkinsonism patients (age of onset <50 years) from Germany with an overall mutation rate of 9.0%. Gene dosage alterations represented 67% of the mutations found, underlining the importance of quantitative analyses of parkin. In summary, parkin mutations accounted for a low but significant percentage of early-onset parkinsonism patients in a community-derived sample. PMID- 12112110 TI - Paraneoplastic chorea associated with CRMP-5 neuronal antibody and lung carcinoma. AB - Paraneoplastic chorea is described in 16 patients: 11 with limited small-cell carcinoma, 2 with lung cancer revealed by imaging, 1 with renal cell carcinoma, and 1 with lymphoma. All had CRMP-5-IgG; 6 also had ANNA-1 (anti-Hu), including 1 without evident cancer. Chorea was the initial and most prominent symptom in 11 patients, asymmetric or unilateral in 5 patients, and part of a multifocal syndrome in 14 patients. Basal ganglia abnormalities were revealed by magnetic resonance imaging and at autopsy (as perivascular inflammation and microglial activation). Four patients improved with chemotherapy, and 2 improved with intravenous methylprednisolone. PMID- 12112111 TI - Mitochondrial DNA C4171A/ND1 is a novel primary causative mutation of Leber's hereditary optic neuropathy with a good prognosis. AB - A novel mitochondrial DNA C4171A mutation in the ND1 gene in two Korean families with Leber's hereditary optic neuropathy is described. All affected patients recovered spontaneously after suffering months to years of initial visual loss. This mutation replaces leucine with methionine in a conserved extramembrane loop of the ND1 gene and was absent in 514 normal controls and in 63 Leber's hereditary optic neuropathy lineages harboring the primary mutations. We consider mitochondrial DNA C4171A/ND1 a primary causative mutation of Leber's hereditary optic neuropathy with a good prognosis. PMID- 12112112 TI - Deficiency of tetralinoleoyl-cardiolipin in Barth syndrome. AB - Barth syndrome is an X-linked cardiac and skeletal mitochondrial myopathy. Barth syndrome may be due to lipid alterations because the product of the mutated gene is homologous to phospholipid acyltransferases. Here we document that a single mitochondrial phospholipid species, tetralinoleoyl-cardiolipin, was lacking in the skeletal muscle (n = 2), right ventricle (n = 2), left ventricle (n = 2), and platelets (n = 6) of 8 children with Barth syndrome. Tetralinoleoyl-cardiolipin is specifically enriched in normal skeletal muscle and the normal heart. These findings support the notion that Barth syndrome is caused by alterations of mitochondrial lipids. PMID- 12112113 TI - Brain biopterin and tyrosine hydroxylase in asymptomatic dopa-responsive dystonia. AB - It is assumed that brain biopterin and dopamine loss should not be as severe in asymptomatic dopa-responsive dystonia caused by GCH1 mutations as it is in symptomatic dopa-responsive dystonia. However, the actual status of dopaminergic systems in asymptomatic cases is unknown. In the autopsied putamen of an asymptomatic GCH1 mutation carrier, we found that brain biopterin loss (-82%) paralleled that reported in dopa-responsive dystonia patients (-84%). However, tyrosine hydroxylase protein and dopamine levels (-52 and -44%, respectively) were not as severely affected as in symptomatic patients (exceeding -97 and -88%, respectively). Our data suggest that the extent of striatal tyrosine hydroxylase protein loss may be critical in determining dopa-responsive dystonia symptomatology. PMID- 12112114 TI - Seizure-associated hippocampal volume loss: a longitudinal magnetic resonance study of temporal lobe epilepsy. AB - This longitudinal quantitative magnetic resonance imaging study of 24 patients with mild temporal lobe epilepsy shows an ipsilateral hippocampal volume decrease of 9% (range, -30 to +0.5%; p = 0.002, paired t test) over a period of 3.5 +/- 0.7 years. The hippocampal volume loss was correlated to the number of generalized seizures between the scans (p = 0.0007, r = 0.6), suggesting seizure associated hippocampal damage. PMID- 12112115 TI - A novel D104G mutation in the adenine nucleotide translocator 1 gene in autosomal dominant progressive external ophthalmoplegia patients with mitochondrial DNA with multiple deletions. AB - Autosomal dominant progressive external ophthalmoplegia is a mitochondrial disorder characterized by multiple large deletions of mitochondrial DNA. A recent study showed pathogenic heterozygous missense mutations in the heart/skeletal muscle isoform of the adenine nucleotide translocator 1 gene in autosomal dominant progressive external ophthalmoplegia patients. In one Japanese autosomal dominant progressive external ophthalmoplegia family, we found a novel A-to-G heterozygous mutation at nucleotide 311 of the adenine nucleotide translocator 1 gene, which segregated with affected individuals and could not be detected in the genomic DNA sequence of 120 normal controls. This mutation converted a highly conserved aspartic acid into a glycine at codon 104. Polymerase chain reaction analysis of single muscle fibers showed the presence of one type of deletion in each fiber, suggesting clonal expansion of mitochondrial DNA with deletions. These findings support the pathogenesis of the adenine nucleotide translocator 1 gene mutation in human disease. PMID- 12112116 TI - Electrophysiological findings in X-linked myopathy with excessive autophagy. AB - We report electrophysiological features and magnetic resonance imaging muscle findings in 4 patients and 1 female carrier of X-linked myopathy with excessive autophagy. Motor units were polyphasic with high mean amplitude and normal duration. The thigh muscles were most severely involved, but myotonic discharges were abundant in both clinically affected and unaffected muscles. Along with the clinicopathological features, these electrophysiological findings distinguish X linked myopathy with excessive autophagy from other limb-girdle myopathies. PMID- 12112117 TI - Diffuse signal abnormalities in the spinal cord in multiple sclerosis: direct postmortem in situ magnetic resonance imaging correlated with in vitro high resolution magnetic resonance imaging and histopathology. AB - In this study, we compared direct postmortem in situ (whole-corpse) sagittal spinal cord magnetic resonance imaging (1.5T) of 7 multiple sclerosis cases with targeted high-resolution in vitro axial magnetic resonance imaging (4.7T) and histopathology. On sagittal in situ magnetic resonance imaging, 1 case had a normal spinal cord, 2 had only focal lesions, 3 had a combination of focal and diffuse abnormalities, and 1 had only diffuse abnormalities. All spinal cords showed abnormalities on high-resolution magnetic resonance imaging and histopathology, confirming the existence of diffuse cord changes as genuine multiple sclerosis-related abnormalities while highlighting the limited resolution of in vivo magnetic resonance imaging. PMID- 12112118 TI - Clinical variability in 3-hydroxy-2-methylbutyryl-CoA dehydrogenase deficiency. AB - We report the identification of two new 7-year-old patients with 3-hydroxy-2 methylbutyryl-CoA dehydrogenase deficiency, a recently described inborn error of isoleucine metabolism. The defect is localized one step above 3-ketothiolase, resulting in a urinary metabolite pattern similar to that seen for deficiency of the latter. One patient has progressive neurodegenerative symptoms, whereas the clinical phenotype of the other patient is characterized by psychomotor retardation without loss of developmental milestones. A short-term biochemical response to an isoleucine-restricted diet was observed in both children. PMID- 12112121 TI - Studies of mitochondrial DNA in Devic's disease revealed no pathogenic mutations, but polymorphisms also found in association with multiple sclerosis. PMID- 12112120 TI - Low cerebrospinal fluid hypocretin (orexin) and altered energy homeostasis in human narcolepsy. PMID- 12112122 TI - Trinucleotide repeat expansions in the junctophilin-3 gene are not found in Caucasian patients with a Huntington's disease-like phenotype. PMID- 12112123 TI - Leonard T. Kurland: 1921-2001. PMID- 12112126 TI - Neurogenesis in the mossy chiton, Mopalia muscosa (Gould) (Polyplacophora): evidence against molluscan metamerism. AB - Neurogenesis in the chiton Mopalia muscosa (Gould, 1846) was investigated by applying differential interference contrast microscopy, semithin serial sectioning combined with reconstruction techniques, as well as confocal laser scanning microscopy for the detection of fluorescence-conjugated antibodies against serotonin and FMRFamide. The ontogeny of serotonergic nervous structures starts with cells of the apical organ followed by those of the cerebral commissure, whereas the serotonergic prototroch innervation, pedal system, and the lateral cords develop later. In addition, there are eight symmetrically arranged serotonergic sensory cells in the dorsal pretrochal area of the larva. FMRFamide-positive neural elements include the cerebral commissure, specific "ampullary" sensory cells in the pretrochal region, as well as the larval lateral and pedal system. In the early juvenile the cerebral system no longer stains with either of the two antibodies and the pedal system lacks anti-FMRFamide immunoreactivity. Outgroup comparison with all other molluscan classes and related phyla suggests that the cord-like, nonganglionized cerebral system in the Polyplacophora is a reduced condition rather than a primitive molluscan condition. The immunosensitivity of the pedal commissures develops from posterior to anterior, suggesting independent serial repetition rather than annelid-like conditions and there is no trace of true segmentation during nervous system development. Polyplacophoran neurogenesis and all other available data on the subject contradict the idea of a segmented molluscan stem species. PMID- 12112127 TI - Comparative studies of the development and differentiation of chloride cells in tilapine fish with different reproductive styles. AB - Using light and electron microscopy and fluorescent probes, we followed the ontogenesis of selected organs in embryos of several species of tilapia (Cichlidae, Pisces) with emphasis on chloride cell differentiation in species with two different reproductive styles: we compared the substrate-brooder Tilapia zillii and the mouth-brooders Oreochromis niloticus, O. aureus, Sarotherodon galilaeus, and Tristramella sacra. In all species a transitory blood network system nurtured by the vena caudalis inferiores supplied the yolk sac and preanal finfold during the advanced stages of embryonic and initial stages of larval development. During these stages chloride cells occurred on the yolk sac, as a part of the abdominal epithelium. The cells and their associated blood plexus remained active here until the gill-lamellae, operculum, and mouth became functional. The chloride cells of their epithelium and blood system then took over, concomitant with a gradual degradation of the transitory blood system on the yolk sac. Ontogenesis of these systems (transitory and permanent) progressed at a higher rate in substrate-brooders than in mouth-brooders and was correlated with the earlier functioning of the gill-operculum system. Thus, at a constant temperature of 26 degrees C, the more exposed T. zillii progeny completed metamorphosis at 7-8 days after fertilization, calculated around 5,000 +/- 80 h/temp, whereas juveniles of more protected mouth-brooders attained a similar stage only 15 +/- 1 days after fertilization and around 9,000 +/- 200 h/temp. This earlier development of chloride cells and other pivotal organs in environmentally exposed progeny of substrate-brooders, as compared to the protected progeny of mouth-brooders, shows that their ontogeny was selected for the optimal survival style under specific etho-ecological conditions. PMID- 12112128 TI - In the absence of Rauber's sickle material, no blood islands are formed in the avian blastoderm. AB - Using the quail-chick chimera technique, we followed the fate of Rauber's sickle cells in older whole blastoderms (cultured for approximately 2 days): after removal of the autochthonous Rauber's sickle from an unincubated chicken blastoderm, a quail Rauber's sickle was grafted isotopically and isochronically in its place. In transverse sections through these chimeras, the grafted quail Rauber's sickle cells were seen to have transformed into a broad row or ridge of quail junctional endoblast cells extending at the inner border of the area containing blood islands. After unilateral removal of the junctional endoblast from an intermediate streak chicken blastoderm (Stage 3; Hamburger and Hamilton [1951] J Morphol 88:49-92), we observed during further in vitro culture that at the operated side, in the area previously occupied by this junctional endoblast, blood islands no longer developed. If after such a unilateral removal of the chicken junctional endoblast quail junctional endoblast was apposed in its place, then blood islands reappeared in the operated area. The intimate contact between the apposed quail junctional endoblast and the recently formed blood islands, derived from peripherally migrating mesoderm, was very obvious on sections through such chimeras. We further demonstrate that Rauber's sickle vs. junctional endoblast is indispensable for the anlage of blood islands in avian blastoderms. Indeed, in the absence of Rauber's sickle material no blood islands develop (even when mesoderm is present after ingression of the upper layer via a primitive streak) in the isolated central region of the area centralis of unincubated chicken blastoderms after culture in vitro. Also, no junctional endoblast and no sickle canal appear in these explants. By contrast, if a Rauber's sickle fragment is placed on such an isolated central blastoderm region, then blood islands develop. These blood islands start to develop from peripherally migrating mesoderm in the neighborhood of the Rauber's sickle-derived junctional endoblast. PMID- 12112129 TI - Ephemeroptera egg chorion characters: a test of their importance in assessing phylogenetic relationships. AB - The egg chorion ultrastructures of the Hermanella-Traverella (Insecta: Ephemeroptera) species complex were studied from a comparative point of view and used for the first time in a cladistic analysis. Egg characters, along with other nymphal and adult morphological characters, were used to assess the phylogenetic relationships of the species complex. In order to test the value of egg characters, analyses were performed on three matrices: 1) egg characters alone, 2) adult and nymphal characters, and 3) adult, nymphal, and egg characters. The computer program Pee-Wee was used to carry out the analysis. The cladistic analysis confirmed the value and potential of egg chorionic characters in assessing the phylogenetic relationships among ephemeropteran species. Egg characters, when added to the nymphal and adult character matrix, provided extra support to the monophyletic nature of the Hermanella-Traverella complex. Previously weakly defined clades were also resolved based on the new evidence. In the species studied the egg chorionic structures as well as their shape did not change after oviposition or water immersion, remaining constant through the different maturation stages of each species (mature nymph, subimago, and imago). For this reason, the eggs are a valuable source of information to unambiguously identify and associate a nymph to its correspondent adult stage when rearing is not possible. PMID- 12112130 TI - Spermatophore transfer in the hermit crab Clibanarius vittatus (Crustacea, Anomura, Diogenidae). AB - Although mating has been described in several hermit crab species, the mechanics of spermatophore transfer have not previously been demonstrated. Evidence from pleopod and gonopore morphology, video observations, and inseminated females indicates that in Clibanarius vittatus the male applies a spermatophoric mass directly onto the female via the gonopores rather than with modified pleopods 1-2 (gonopods) and/or genital papillae as in many other decapods. The single second pleopod of males of C. vittatus has a simple endopod with no apparent modifications for sperm transfer. There are no genital papillae extending from the male gonopores. The globular spermatophores are aligned in rows surrounded by a seminal secretion in the male ducts (vasa deferentia that terminate in ejaculatory ducts opening to the exterior via the gonopores). During copulation, described from time-lapse video recordings, the ventral surface of the last thoracic segment of the male, bearing the gonopores, was apposed to the ventral cephalothorax of the female. A massive amount of seminal secretion containing spermatophore ribbons, termed here the spermatophoric mass and described for the first time in a hermit crab species, was observed covering the sternites and coxae of pereopods 1-5 of a recently copulated female. It is suggested that during copulation the male emits the contents of the ejaculatory ducts directly onto the female without the aid of gonopods or genital papillae. Although spermatophore transfer is simple in C. vittatus, the presence of modified anterior pleopods or elongate genital papillae (sexual tubes) in other paguroidean species suggests the possibility of a more complex insemination process in these other hermit crabs. PMID- 12112131 TI - Serous cutaneous glands in new world hylid frogs: an ultrastructural study on skin poisons confirms phylogenetic relationships between Osteopilus septentrionalis and Phrynohyas venulosa. AB - Transmission electron microscope investigations of the serous (poison) skin glands in the New World tree frogs Osteopilus septentrionalis and Phrynohyas venulosa revealed that they produce granules with closely similar substructures, namely, a dense cortex and pale medulla. In both species these features, that contrast the complex, sometimes repeating patterns described in other hylid frogs, derive from similar secretory and maturational processes starting from the Golgi phase of poison biosynthesis. Observations on secretory discharge showed that the two species share common release mechanisms, based on bulk discharge (holocrine) processes. Our data provide novel evidence of the extensive ultrastructural polymorphism of serous skin products in Hylidae and agree with phylogenies that regard this family as polyphyletic in origin. Assuming that ultrastructural features of cutaneous poison biosynthesis and maturation are adequate clues for tracking anuran phylogeny, the present findings also support a close relationship between Osteopilus and Phrynohyas taxa as previously suggested by osteological evidence. PMID- 12112132 TI - Gill-derived glands in glandulocaudine fishes (teleostei: Characidae: Glandulocaudinae). AB - The Glandulocaudinae is a subfamily of neotropical characid fishes from Central and South America. A unifying feature of the subfamily is the caudal gland, found almost exclusively in males. The gland consists of tissue on the base of the caudal fin covered in part by hypertrophied scales. Scale movement as the caudal fin is flexed appears to facilitate the release of chemical compounds from the glandular tissue. We describe here a different structure, found in the gill cavity of mature males in 12 of 17 glandulocaudine genera examined. Termed a gill gland, it develops as a male secondary sex character and appears morphologically suited to release chemical signals. The gland forms by the growth of tissue over and around 4-13 anterior gill filaments on the first gill arch, forming chambers with ventral openings. Within the gland chambers, gill secondary lamellae usually shorten and may disappear. When secondary lamellae persist, simple columnar epithelial cells develop between them. In the absence of secondary lamellae, the gland chambers are lined with a simple cuboidal or columnar epithelium. Gland size and the degree of gill modification vary among species. Gill glands appear absent in five glandulocaudine genera, suggesting character reversals based on current phylogenetic hypotheses and systematic classification. Gill gland morphology suggests that this structure releases chemical compounds into the gill current. The presence of gill glands only in mature males suggests a function in reproduction and/or male aggression. Together with studies of the caudal gland, this research suggests that chemical signals may play important roles in glandulocaudine reproduction. PMID- 12112133 TI - Bite performance in clariid fishes with hypertrophied jaw adductors as deduced by bite modeling. AB - Within clariid fishes several cranial morphologies can be discerned. Especially within anguilliform representatives an increase in the degree of hypertrophy of the jaw adductors occurs. The hypertrophy of the jaw adductors and skeletal modifications in the cranial elements have been linked to increased bite force. The functional significance of this supposed increase in bite force remains obscure. In this study, biomechanical modeling of the cranial apparatus in four clariid representatives showing a gradual increase in the hypertrophy of the jaw adductors (Clarias gariepinus, Clariallabes melas, Channallabes apus, and Gymnallabes typus) is used to investigate whether bite force actually increased. Static bite modeling shows that the apparent hypertrophy results in an increase in bite force. For a given head size, the largest bite forces are predicted for C. apus, the lowest ones for C. gariepinus, and intermediate values are calculated for the other species. In addition, also in absolute measures differences in bite force remain, with C. apus biting distinctly harder than C. gariepinus despite its smaller head size. This indicates that the hypertrophy of the jaw adductors is more than just a correlated response to the decrease in absolute head size. Further studies investigating the ecological relevance of this performance difference are needed. PMID- 12112134 TI - Kinesin-II is not essential for mitosis and cell growth in Chlamydomonas. AB - The FLA10 gene product (Fla10p) in Chlamydomonas, a heterotrimeric kinesin-II, plays a crucial role in flagellar assembly as a motor protein driving intraflagellar transport. This protein has also been suggested to play a role in mitosis based on its localization to mitotic spindle. A role for Fla10p in mitosis has been difficult to test because to date only conditional (temperature sensitive) mutant alleles were available, and it is not known whether these retain residual function for mitosis at the non-permissive temperature. In this report, we describe a null allele of fla10 produced by insertional mutagenesis. This mutant does not assemble flagella, but proliferates at a rate identical to that of wild type cells. Observation of microtubule organization in the cell body revealed that normal mitotic spindles are formed in dividing mutant cells. Thus, we conclude that FLA10 kinesin plays no significant roles in mitosis. PMID- 12112135 TI - Myosins of Babesia bovis: molecular characterisation, erythrocyte invasion, and phylogeny. AB - Using degenerate primers, three putative myosin sequences were amplified from Australian isolates of Babesa bovis and confirmed as myosins (termed Bbmyo-A, Bbmyo-B, and Bbmyo-C) from in vitro cultures of the W strain of B. bovis. Comprehensive analysis of 15 apicomplexan myosins suggests that members of Class XIV be defined as those with greater than 35% myosin head sequence identity and that these be further subclassed into groups bearing above 50-60% identity. Bbmyo A protein bears a strong similarity with other apicomplexan myosin-A type proteins (subclass XIVa), the Bbmyo-B myosin head protein sequence exhibits low identity (35-39%) with all members of Class XIV, and 5'-sequence of Bbmyo-C shows strong identity (60%) with P. falciparum myosin-C protein. Domain analysis revealed five divergent IQ domains within the neck of Pfmyo-C, and a myosin-N terminal domain as well as a classical IQ sequence unusually located within the head converter domain of Bbmyo-B. A cross-reacting antibody directed against P. falciparum myosin-A (Pfmyo-A) revealed a zone of approximately 85 kDa in immunoblots prepared with B. bovis total protein, and immunofluorescence inferred stage-specific myosin-A expression since only 25% of infected erythrocytes with mostly paired B. bovis were immuno-positive. Multiplication of B. bovis in in vitro culture was inhibited by myosin- and actin-binding drugs at concentrations lower than those that inhibit P. falciparum. This study identifies and classifies three myosin genes and an actin gene in B. bovis, and provides the first evidence for the participation of an actomyosin-based motor in erythrocyte invasion in this species of apicomplexan parasite. PMID- 12112136 TI - Simultaneous quantification of cell motility and protein-membrane-association using active contours. AB - We present a new method for the quantification of dynamic changes in fluorescence intensities at the cell membrane of moving cells. It is based on an active contour method for cell-edge detection, which allows tracking of changes in cell shape and position. Fluorescence intensities at specific cortical subregions can be followed in space and time and correlated with cell motility. The translocation of two GFP tagged proteins (CRAC and GRP1) from the cytosol to the membrane in response to stimulation with the chemoattractant cAMP during chemotaxis of Dictyostelium cells and studies of the spatio-temporal dynamics of this process exemplify the method: We show that the translocation can be correlated with motility parameters and that quantitative differences in the rate of association and dissociation from the membrane can be observed for the two PH domain containing proteins. The analysis of periodic CRAC translocation to the leading edge of a cell responding to natural cAMP waves in a mound demonstrates the power of this approach. It is not only capable of tracking the outline of cells within aggregates in front of a noisy background, but furthermore allows the construction of spatio-temporal polar plots, capturing the dynamics of the protein distribution at the cell membrane within the cells' moving co-ordinate system. Compilation of data by means of normalised polar plots is suggested as a future tool, which promises the so-far impossible practicability of extensive statistical studies and automated comparison of complex spatio-temporal protein distribution patterns. PMID- 12112138 TI - Geometric Clutch model version 3: the role of the inner and outer arm dyneins in the ciliary beat. AB - The Geometric Clutch model of ciliary and flagellar beating uses the transverse force (t-force) that develops between the outer doublets of the axoneme as the regulator for activating and deactivating the dynein motors and organizing the flagellar beat. The version of the model described here adds detail to the formulations used in the two previous versions as follows: (1) In place of two opposing sets of dyneins, the new model has four sets of dyneins, corresponding to two sets on each side of the axoneme acting in series. (2) The four sets of dyneins are each subdivided into two ranks representing inner and outer arm dyneins. (3) The force produced by each dynein is governed by a force-velocity relationship that is independently specified for the inner and outer arms. Consistent with the original model, the new version of the Geometric Clutch model can simulate both the effective and recovery stroke phases of the ciliary beat using a single uniform algorithm. In addition, the new version can operate with the outer arms disabled. Under this condition, the simulation exhibits a beat pattern similar to the original but the beat frequency is reduced to approximately one third. These results are contingent on using force-velocity relationships for the inner and outer arms similar to those described by Brokaw [1999: Cell Motil. Cytoskeleton 42:134-148], where the inner arms contribute most of the driving force at low shear velocities. This constitutes the first examination of the effects of the force-velocity characteristics of dynein on a cilia-like beat in a theoretical framework. PMID- 12112137 TI - Contraction-mediated pinocytosis of RGD-peptide by dermal fibroblasts: inhibition of matrix attachment blocks contraction and disrupts microfilament organisation. AB - Force generation in collagen and matrix contraction are basic functions of fibroblasts and important elements of tissue repair. Cell-matrix attachment is critical to this contraction, involving RGD-binding integrins. We have investigated how this process operates, in terms of force generation (in the Culture Force Monitor) and cytoskeletal structure, using a synthetic RGD decapeptide. The RGD-peptide blocked force generation over the first 6 h, followed by near complete recovery by 20 h. However, dose response was complex indicating multiple processes were operating. Analysis of cytoskeletal structure after treatment with RGD-peptide indicated major disruption with condensed aggregates of actin and microtubular fragmentation. Fluorescent labeling and tracking of the RGD-peptide demonstrated intracellular uptake into discrete cytoplasmic aggregates. Critically, these RGD-peptide pools co-localised with the condensed actin microfilament aggregates. It is concluded that RGD-peptide uptake was by a form of contraction-mediated pinocytosis, resulting from mechanical tension applied to the untethered RGD-peptide-integrin, as contractile microfilament were assembled. These findings emphasize the importance of sound mechanical attachment of ligand-occupied integrins (e.g., to extracellular matrix) for normal cytoskeletal function. Conversely, this aspect of unrestrained cytoskeletal contraction may have important pathogenic and therapeutic applications. PMID- 12112139 TI - Host focal adhesion protein domains that bind to the translocated intimin receptor (Tir) of enteropathogenic Escherichia coli (EPEC). AB - Enteropathogenic Escherichia coli (EPEC) attach to the plasma membrane of infected host cells and induce diarrhea in a variety of farm animals as well in humans. These bacteria inject a three-domain protein receptor, Tir (translocated intimin receptor), that is subsequently inserted into the plasma membrane. EPEC induce the host cell to form membrane-covered actin-rich columns called pedestals. Focal adhesion constituents, alpha-actinin, talin, and vinculin, are localized along the length of the pedestals and we have previously reported they bind the two cytoplasmic domains of Tir, (Tir I and Tir III) [Freeman et al., 2000: Cell Motil. Cytoskeleton 47:307-318]. In the present study, various constructs were made expressing different regions of these three focal adhesion proteins to determine which domains of the proteins bound Tir I. Three different assays were used to detect Tir I/host protein domain interactions. In co precipitation assays, His-Tir I bound to the 27-kDa region of alpha-actinin; to four different domains of talin; and to the N-terminal domain of the vinculin head and the vinculin tail domain. A yeast two-hybrid analysis of Tir I and the various focal adhesion fusion proteins revealed a region near the C-terminus of talin was the only domain to interact with Tir I. Finally, to assess direct binding interactions, biotinylated Tir I was used in overlay assays and confirmed the binding of Tir I with the 27-kDa region of alpha-actinin, the four regions of talin, and the vinculin tail. These binding interactions between hostfocal adhesion proteins and EPEC Tir may facilitate the adhesion of EPEC to the host cell surface. PMID- 12112140 TI - Buckling of actin stress fibers: a new wrinkle in the cytoskeletal tapestry. AB - Intracellular tension is considered an important determinant of cytoskeletal architecture and cell function. However, many details about cytoskeletal tension remain poorly understood because these forces cannot be directly measured in living cells. Therefore, we have developed a method to characterize the magnitude and distribution of pre-extension of actin stress fibers (SFs) due to resting tension in the cytoskeleton. Using a custom apparatus, human aortic endothelial cells (HAECs) were cultured on a pre-stretched silicone substrate coated with a fibronectin-like polymer. Release of the substrate caused SFs aligned in the shortening direction in adhered cells to buckle when compressed rapidly (5% shortening per second or greater) beyond their unloaded slack length. Subsequently, the actin cytoskeleton completely disassembled in 5 sec and reassembled within 60 sec. Quantification of buckling in digital fluorescent micrographs of cells fixed and stained with rhodamine phalloidin indicated a nonuniform distribution of 0-26% pre-extension of SFs in non-locomoting HAECs. Local variability suggests heterogeneity of cytoskeletal tension and/or stiffness within individual cells. These findings provide new information about the magnitude and distribution of cytoskeletal tension and the dynamics of actin stress fibers, and the approach offers a novel method to elucidate the role of specific cytoskeletal elements and crosslinking proteins in the force generating apparatus of non-muscle cells. PMID- 12112141 TI - Redox-based control of the gamma heavy chain ATPase from Chlamydomonas outer arm dynein. AB - The outer dynein arm from Chlamydomonas flagella contains two redox-active thioredoxin-related light chains associated with the alpha and beta heavy chains; these proteins belong to a distinct subgroup within the thioredoxin family. This observation suggested that some aspect of dynein activity might be modulated through redox poise. To test this, we have examined the effect of sulfhydryl oxidation on the ATPase activity of isolated dynein and axonemes from wildtype and mutant strains lacking various heavy chain combinations. The outer, but not inner, dynein arm ATPase was stimulated significantly following treatment with low concentrations of dithionitrobenzoic acid; this effect was readily reversible by dithiol, and to a lesser extent, monothiol reductants. Mutational and biochemical dissection of the outer arm revealed that ATPase activation in response to DTNB was an exclusive property of the gamma heavy chain, and that enzymatic enhancement was modulated by the presence of other dynein components. Furthermore, we demonstrate that the LC5 thioredoxin-like light chain binds to the N-terminal stem domain of the alpha heavy chain and that the beta heavy chain associated LC3 protein also interacts with the gamma heavy chain. These data suggest the possibility of a dynein-associated redox cascade and further support the idea that the gamma heavy chain plays a key regulatory role within the outer arm. PMID- 12112142 TI - Arabidopsis thaliana protein, ATK1, is a minus-end directed kinesin that exhibits non-processive movement. AB - The microtubule cytoskeleton forms the scaffolding of the meiotic spindle. Kinesins, which bind to microtubules and generate force via ATP hydrolysis, are also thought to play a critical role in spindle assembly, maintenance, and function. The A. thaliana protein, ATK1 (formerly known as KATA), is a member of the kinesin family based on sequence similarity and is implicated in spindle assembly and/or maintenance. Thus, we want to determine if ATK1 behaves as a kinesin in vitro, and if so, determine the directionality of the motor activity and processivity character (the relationship between molecular "steps" and microtubule association). The results show that ATK1 supports microtubule movement in an ATP-dependent manner and has a minus-end directed polarity. Furthermore, ATK1 exhibits non-processive movement along the microtubule and likely requires at least four ATK1 motors bound to the microtubule to support movement. Based on these results and previous data, we conclude that ATK1 is a non-processive, minus-end directed kinesin that likely plays a role in generating forces in the spindle during meiosis. PMID- 12112143 TI - Microtubule displacements at the tips of living flagella. AB - We have observed that the flagellar axoneme of the Chinese hamster spermatozoon undergoes periodic changes in length at the same frequency as the flagellar beat. The amplitude of the length oscillation recorded at the tip is maximally about 0.5 microm or 0.2% of the total length. In some favourable cells, it was possible to see the opposing "halves" of the axoneme moving at the tip in a reciprocating manner and 180 degrees out-of-phase. This behaviour, when analysed quantitatively, is broadly consistent with predictions made from the sliding doublet theory of ciliary and flagellar motility and thus it constitutes an additional verification of the theory, for the first time in a living cell. However, on close examination, there is a partial mismatch between the timing of the length oscillation and the phase of the beat cycle. We deduce from this that there is some sliding at the base of the flagellum, sliding that is accommodated by elastic compression of the connecting piece. Micrographic evidence for such compression is presented. PMID- 12112144 TI - Yeast polypeptide chain release factors eRF1 and eRF3 are involved in cytoskeleton organization and cell cycle regulation. AB - Termination of translation in eukaryotes is controlled by two interacting polypeptide chain release factors, eRF1 and eRF3. eRF1 recognizes nonsense codons UAA, UAG, and UGA, while eRF3 stimulates polypeptide release from the ribosome in a GTP- and eRF1-dependent manner. In the yeast Saccharomyces cerevisiae, eRF1 and eRF3 are encoded by the SUP45 and SUP35 genes, respectively. Here we show that in yeast shortage of any one of the release factors was accompanied by a reduction in the levels of the other release factor and resulted in a substantial increase of nonsense codon readthrough. Besides, repression of the genes encoding these factors caused different effects on cell morphology. Repression of the SUP35 gene caused accumulation of cells of increased size with large buds. This was accompanied by the disappearance of actin cytoskeletal structures, impairment of the mitotic spindle structure, and defects in nuclei division and segregation in mitosis. The evolutionary conserved C-terminal domain of eRF3 similar to the elongation factor EF-1alpha was responsible for these effects. Repression of the SUP45 gene caused accumulation of unbudded cells with 2C and higher DNA content, indicating that DNA replication is uncoupled from budding. The data obtained suggest that eRF1 and eRF3 play additional, nontranslational roles in the yeast cell. PMID- 12112145 TI - Distribution of the class II beta-tubulin in developmental and adult rat tissues. AB - During a screen of monoclonal antibodies raised against a cytoskeletal preparation of neonatal rat cerebrum, we have identified a monoclonal antibody, MAb58A, that is specific for the class II beta-tubulin isotype. Immunoscreening of a rat brain cDNA library using MAb58A yielded the cDNA retaining a class II specific nucleotide sequence. The specificity of MAb58A to the class II beta tubulin isotype was confirmed by immunoreactivity to synthetic peptides corresponding to isotype-specific sequence of class I, II, III, IVa, or IVb. Further, the results of an immunoassay against a series of overlapping octapeptides derived from a class II-specific region revealed that the antibody epitope was a heptapeptide that consists of Glu-Glu-Glu-Glu-Gly-Glu-Asp (EEEEGED). Immunoblot analysis revealed that the class II isotype represented a significant portion of beta-tubulin present in the adrenal gland, brain, and testis of adult rats. In fetal tissues, this isotype was detected in skeletal muscle, as well as in the brain. Immunohistochemically, MAb58A reacted predominantly with components of the developing rat nervous system, such as migrating neuroblasts, peripheral nerves and ganglion cells, and sensory organs. MAb58A-immunoreactivity was also found in developing skeletal and smooth muscle cells, chondrocytes, and vascular endothelia. In adults, MAb58A-immunoreactivity was remarkably diminished, but persisted in peripheral nerves and ganglion cells, chondrocytes, and capillary components. Together, our results demonstrate that MAb58A is specific for the class II beta-tubulin isotype, which may retain an embryonic nature in both neuronal and non-neuronal tissues. PMID- 12112146 TI - Arrest of cell cycle progression during first interphase in murine zygotes microinjected with anti-PCM-1 antibodies. AB - To investigate the function of the centrosome protein PCM-1, antibodies against PCM-1 were microinjected into either germinal vesicle stage meiotic oocytes or fertilized mouse eggs, and cell cycle progression events (i.e., microtubule assembly, chromosome and centrosome organization, meiotic maturation) were assayed. These studies determined that microinjected PCM-1 antibodies arrested cell cycle progression, with anti-PCM-1 arresting fertilized eggs at the pronucleate stage when injected during G1. Analysis of the injected eggs determined that centrosome disruption and microtubule cytaster disorganization accompanied the cell cycle arrest. Anti-PCM-1 blocked neither pronuclear centration, completion of mitosis when microinjected into zygotes at G2, nor meiotic maturation when microinjected into immature oocytes. These results identify a novel role for PCM- 1 in cell cycle regulation, and indicate that PCM 1 must fulfill an essential function for cells to complete interphase. PMID- 12112148 TI - Inducing precocious anaphase in cultured mammalian cells. AB - The spindle checkpoint, which prevents anaphase onset upon spindle damage or incorrect chromosome alignment, presents a problem for experimental analysis of protein function in anaphase and cytokinesis. This is because the functional disruption of many proteins before anaphase onset can activate this checkpoint, preventing anaphase and subsequent cell cycle events. This paper compares new and old methods of overriding the spindle checkpoint in prometaphase mammalian tissue culture cells. PMID- 12112149 TI - Transient concentration of a gamma-tubulin-related protein with a pericentrin related protein in the formation of basal bodies and flagella during the differentiation of Naegleria gruberi. AB - The distribution of two proteins in Naegleria gruberi, N-gammaTRP (Naegleria gamma-tubulin-related protein) and N-PRP (Naegleria pericentrin-related protein), was examined during the de novo formation of basal bodies and flagella that occurs during the differentiation of N. gruberi. After the initiation of differentiation, N-gammaTRP and N-PRP began to concentrate at the same site within cells. The percentage of cells with a concentrated region of N-gammaTRP and N-PRP was maximal (68%) at 40 min when the synthesis of tubulin had just started but no assembled microtubules were visible. When concentrated tubulin became visible (60 min), the region of concentrated N-gammaTRP and N-PRP was co localized with the tubulin spot and then flagella began to elongate from the region of concentrated tubulin. When cells had elongated flagella, the concentrated N-gammaTRP and N-PRP were translocated to the opposite end of the flagellated cells and disappeared. The transient concentration of N-gammaTRP coincided with the transient formation of an F-actin spot at which N-gammaTRP and alpha-tubulin mRNA were co-localized. The concentration of N-gammaTRP and formation of the F-actin spot occurred without the formation of microtubules but were inhibited by cytochalasin D. These observations suggest that the regional concentration of N-gammaTRP and N-PRP is mediated by actin filaments and might provide a site of microtubule nucleation for the assembly of newly synthesized tubulins into basal bodies and flagella. PMID- 12112150 TI - Contractility of single human dermal myofibroblasts and fibroblasts. AB - Human dermal myofibroblasts, characterised by the expression of alpha-smooth muscle actin, are part of the granulation tissue and implicated in the generation of contractile forces during normal wound healing and pathological contractures. We have compared the contractile properties of single human dermal fibroblasts and human dermal myofibroblasts by culturing them on flexible silicone elastomers. The flexibility of the silicone substratum permits the contractile forces exerted by the cells to be measured [Fray et al., 1998: Tissue Eng. 4:273 283], without changing their expression of alpha-smooth muscle actin. The mean contractile force produced by myofibroblasts (2.2 microN per cell) was not significantly different from that generated by fibroblasts (2.0 microN per cell) when cultured on a substrata with a low elastomer stiffness. Forces produced by fibroblasts were unaffected by increases in elastomer stiffness, but forces measured for myofibroblasts increased to a mean value of 4.1 microN/cell. This was associated with a higher proportion of myofibroblasts being able to produce wrinkles on elastomers of high stiffness compared to fibroblasts. We discuss the force measurements at the single cell level, for both fibroblast and myofibroblasts, in relation to the proposed role of myofibroblasts in wound healing and pathological contractures. PMID- 12112151 TI - Growth cones contain myosin II bipolar filament arrays. AB - Nonmuscle myosin II is among the most abundant forms of myosin in nerve growth cones. At least two isoforms of myosin II (A and B) that have overlapping but distinct distributions are found in growth cones. It appears that both myosin IIA and IIB may be necessary for normal nerve outgrowth and motility, but the molecular interactions responsible for their activity remain unclear. For instance, it is unknown if these myosin II isoforms produce bipolar "minifilaments" in growth cones similar to those observed in other nonmuscle cells. To determine if minifilaments are present in growth cones, we modified the electron microscopy preparative procedures used to detect minifilaments in other cell types. We found structures that appeared very similar to bipolar minifilaments found in noneuronal cells. They also labeled with antibodies to either myosin IIA or IIB. Thus, the activity of myosin II in growth cones is likely to be similar to that in other nonmuscle cells. Bipolar filaments interacting with oppositely oriented actin filaments will produce localized contractions or exert tension on actin networks. This activity will be responsible for the myosin II dependent motility in growth cones. PMID- 12112152 TI - Micropatterning tractional forces in living cells. AB - Here we describe a method for quantifying traction in cells that are physically constrained within micron-sized adhesive islands of defined shape and size on the surface of flexible polyacrylamide gels that contain fluorescent microbeads (0.2 microm diameter). Smooth muscle cells were plated onto square (50 x 50 microm) or circular (25- or 50-microm diameter) adhesive islands that were created on the surface of the gels by applying a collagen coating through microengineered holes in an elastomeric membrane that was later removed. Adherent cells spread to take on the size and shape of the islands and cell tractions were quantitated by mapping displacement fields of the fluorescent microbeads within the gel. Cells on round islands did not exhibit any preferential direction of force application, but they exerted their strongest traction at sites where they formed protrusions. When cells were confined to squares, traction was highest in the corners both in the absence and presence of the contractile agonist, histamine, and cell protrusions were also observed in these regions. Quantitation of the mean traction exerted by cells cultured on the different islands revealed that cell tension increased as cell spreading was promoted. These results provide a mechanical basis for past studies that demonstrated a similar correlation between spreading and growth within various anchorage-dependent cells. This new approach for analyzing the spatial distribution of mechanical forces beneath individual cells that are experimentally constrained to defined sizes and shapes may provide additional insight into the biophysical basis of cell regulation. PMID- 12112153 TI - Separating centrosomes interact in the absence of associated chromosomes during mitosis in cultured vertebrate cells. AB - We detail here how "free" centrosomes, lacking associated chromosomes, behave during mitosis in PtK(2) homokaryons stably expressing GFP-alpha-tubulin. As free centrosomes separate during prometaphase, their associated astral microtubules (Mts) interact to form a spindle-shaped array that is enriched for cytoplasmic dynein and Eg5. Over the next 30 min, these arrays become progressively depleted of Mts until the two centrosomes are linked by a single bundle, containing 10-20 Mts, that persists for > 60 min. The overlapping astral Mts within this bundle are loosely organized, and their plus ends terminate near its midzone, which is enriched for an ill-defined matrix material. At this time, the distance between the centrosomes is not defined by external forces because these organelles remain stationary when the bundle connecting them is severed by laser microsurgery. However, since the centrosomes move towards one another in response to monastrol treatment, the kinesin-like motor protein Eg5 is involved. From these results, we conclude that separating asters interact during prometaphase of mitosis to form a spindle-shaped Mt array, but that in the absence of chromosomes this array is unstable. An analysis of the existing data suggests that the stabilization of spindle Mts during mitosis in vertebrates does not involve the chromatin (i.e., the RCC1/RanGTP pathway), but instead some other chromosomal component, e.g., kinetochores. PMID- 12112155 TI - Primary hyperparathyroidism in an elderly woman: surgical reversibility of profound mental state problems due to mild hypercalcaemia. AB - BACKGROUND: Clinical case of primary hyperparathyroidism in an 89 year old woman causing profound neuropsychiatric symptoms associated with relatively mild increases in serum calcium levels. OBJECTIVES: To demonstrate that lowering of serum calcium levels by medical treatment (biphosphonate), provided a scientific basis from which to treat a very elderly woman with neuropsychiatric symptoms with definitive surgery. METHOD: Case report. RESULTS: The use of biphosphonate therapy led to marked but temporary improvements in her mental state. CONCLUSION: A trial of reducing serum calcium levels with biphosphonate appears to be indicated in elderly patients with mild primary hyperparathyroidism and neuropsychiatric disorders. PMID- 12112154 TI - An unexplained sequestration of latrunculin A is required in neutrophils for inhibition of actin polymerization. AB - Latrunculin A (LatA) is a toxic natural product that causes disruption of the actin cytoskeleton in many eukaryotic cells at submicromolar concentrations. LatA has been found to bind G-actin with a dissociation constant of 0.2 microM, and more recently to bind profilin-G-actin and, weakly, thymosin beta4-G-actin. A number of investigators have used LatA as a G-actin sequestering agent. Thus, we studied neutrophil chemotaxis and its requisite conversion of G-actin to F-actin, supported by an extensive pool of G-actin, mainly bound to thymosin beta4. Calculations suggest that the affinity of LatA is insufficient to cause significant sequestration of this pool, and the pool's buffering action should protect neutrophils from depletion of productive G-actin species by submicromolar LatA. Nonetheless, we found that both chemoattractant stimulated migration and F actin polymerization in neutrophils were inhibited by LatA at these concentrations. The latter effect was accompanied by sequestration of LatA and showed a cell density dependence that was consistent with G-actin sequestration. The apparent contradiction between the calculations and the experimental observations could be reconciled by assuming the presence of an accessory species, of unknown normal function, which forms a high affinity ternary complex with LatA and G-actin, thus causing the cells to concentrate LatA. Other models that could not be ruled out also invoke new actions of LatA, suggesting caution in the interpretation of its effects on cells. PMID- 12112156 TI - Sleep disturbances among nursing home residents. AB - STUDY OBJECTIVES: This study assesses the prevalence and characteristics of sleep disturbances among an entire nursing home population, consisting of 29, mainly demented, long-term patients. DESIGN AND SETTING: Sleep was evaluated for 14 consecutive days using actigraphic measurements and nursing staff observations. No alterations were made in every-day routines or medications during the observation period. MEASUREMENTS AND RESULTS: Actigraphy showed a mean sleep onset latency of one hour and a mean wake after sleep onset of more than two hours, while there was no findings of early morning awakening. Mean sleep efficiency was 75%, and more than 13 hours were spent in bed. 72% of the subjects had sleep efficiency below 85%. Nursing staff reported sleep onset latency of more than 30 minutes in 158 of the 203 analysed days, while early morning awakening was reported in only 12 of 203 days. Actigraphical measurements and nursing staff observations gave similar results. The validity of actigraphy in this population is discussed. CONCLUSION: Sleep disturbances were common among the residents in this nursing home. Sleep onset latency was prolonged, and the patients experienced frequent wake bouts after sleep onset. The diminished ability of sustained sleep may have been influenced by the prolonged time in bed. PMID- 12112157 TI - Estrogen replacement therapy is associated with less progression of subclinical structural brain disease in normal elderly women: a pilot study. AB - BACKGROUND: Cortical atrophy, central atrophy, deep white-matter hyperintensities, and periventricular hyperintensities are reported in normal aging. OBJECTIVES: We examined the effects of estrogen replacement therapy (ERT) on these forms of 'subclinical structural brain disease' (SSBD) in normal, postmenopausal women in a pilot, naturalistic, longitudinal study of 15 subjects. METHODS: Two assessments were performed at least two years apart, with volumetric magnetic resonance imaging (MRI) and neuropsychological testing. RESULTS: Women receiving open-label ERT showed significantly less progression of SSBD than those who did not. CONCLUSIONS: The association between reduced SSBD progression and ERT suggests this intervention could help preserve normal brain structure in healthy elderly women. PMID- 12112158 TI - Prophylactic therapy with lithium in elderly patients with unipolar major depression. AB - OBJECTIVES: To compare the relapse rate of elderly depressed patients taking low dose lithium as an additional therapy with antidepressant medication to those receiving antidepressant medication alone. METHODS: Fifty elderly subjects recovering from a major depressive illness taking continuation antidepressants were randomised, in a double blind study, to receive additional lithium carbonate or placebo and followed up over a two year period for evidence of relapse. RESULTS: Relapse rate was significantly greater in those subjects taking antidepressant medication alone compared to subjects taking additional lithium therapy. After six months four (17%) subjects taking antidepressant medication alone had relapsed, whereas none of the subjects taking additional lithium had relapsed. After two years eight (33%) subjects taking antidepressant medication alone had relapsed, whereas only one (4%) of the subjects taking additional lithium had relapsed. CONCLUSION: This preliminary study suggests that long-term low dose lithium therapy is well tolerated and protects elderly patients from a relapse of depressive illness. PMID- 12112159 TI - Access to a community aged psychiatry service by elderly from non-English speaking backgrounds. AB - OBJECTIVE: With the ageing of Australia's ethnic communities, aged mental health services need to examine issues pertaining to accessibility and appropriateness in the context of this sociodemographic change. The aim of this review of referrals to a community aged-psychiatry service was to compare for differences between patients from non-English-speaking backgrounds (NESB) and English speaking backgrounds (ESB). METHOD: Sociodemographic and clinical variables were retrospectively collated for a 12-month period and analysed according to NESB and ESB status. The 1996 Australian Census data were used for comparison of catchment area representation of different ethnic groups. RESULTS: 40.8% of patients referred to the service were from NESB, and 78.8% of these were assessed with an interpreter. Taken as broad ethnic groups, the referral of elderly European migrants was similar to their representation in the local population, while elderly migrants from Asia and other non-European backgrounds were under represented. Sociodemographic differences were found in that the elderly from NESB were more likely to be poorly educated, have a low proficiency in English, and to have been employed in unskilled occupations. On the other hand, patterns of referral, diagnosis, past psychiatric history and outcome did not differ significantly between the two groups. Clinical variables relating to referral source, past psychiatric history, and outcome did not differ significantly across the two groups. CONCLUSIONS: This 12-month review of referrals to an aged psychiatry community service found that nearly half were of elderly patients from NESB. The lower utilisation of the service by certain ethnic groups may reflect obstacles in their pathway to care. Alternatively, strong family networks, or a lower prevalence of mental illness in these elderly, may explain the findings in this report. PMID- 12112160 TI - Serum cholesterol and depressive symptoms in elderly Finnish men. AB - OBJECTIVE: Evidence from previous studies suggests that alterations in lipid levels may be associated with depression in old age. The objective of this study was to investigate the association between serum lipids and depressive symptoms in a population of elderly men. SUBJECTS AND METHODS: Altogether 470 men born between 1900 and 1919 were examined in the 30-year follow-up of the Seven Countries Study in 1989. Zung Self-Rating Depression Scale was used to determine the depressive status of the subjects. The depressive status was dichotomised and used as the dependent variable in the present study. RESULTS: The depressive status was available for 421 men aged 70 to 89 years in 1989. The prevalence of depression, defined as the Zung sum score equal to or greater than 48, was 15.2% (n = 64). A low serum total cholesterol (odds ratio (OR) 0.67, 95% confidence intervals (CI) 0.48-0.94, p = 0.022) and low low density lipoprotein cholesterol (OR 0.67, 95% CI: 0.46-0.98, p = 0.041) were independently associated with depression. No association with depression was found for high density lipoprotein (HDL) concentration or HDL/total cholesterol ratio after the adjustment for other putative correlates for depression. CONCLUSIONS: Our study of a well-documented population of elderly Finnish men confirms that low total serum cholesterol is associated with a high amount of depressive symptoms independently of weight change or chronic disease. Our study is the first to show an independent association of low LDL-cholesterol concentration with a high amount of depressive symptoms in the old-old. PMID- 12112161 TI - Apolipoprotein E epsilon 4 allele is not associated with the cognitive impairment in community-dwelling normal elderly individuals. AB - OBJECTIVES: The aim of this study was to examine whether the APOE epsilon 4 allele also confers a risk for the cognitive impairment in normal aging. METHODS: We administered all the eight neuropsychological tests from the CERAD neuropsychological battery to the CVD-free, community-dwelling normal elderly individuals, and compared their performance by the occurrence of the APOE epsilon 4 allele. RESULTS: Either the impact of APOE epsilon 4 allele itself or its interaction terms with age and gender of the subjects did not influence the performance of the eight neuropsychological tests (epsilon p > 0.1 by ANCOVA). CONCLUSIONS: The APOE epsilon 4 allele is not a risk factor for the cognitive decline in normal elderly individuals regardless of age and gender. PMID- 12112163 TI - Very early-onset familial Alzheimer's disease: a novel presenilin 1 mutation. AB - BACKGROUND: Early-onset familial Alzheimer's disease (EOFAD) is linked to mutations in three autosomal dominant genes: PS1, PS2 and APP. The clinical presentation and age of onset of mutations is variable. OBJECTIVES: The aim of this report is to describe a novel PS1 mutation believed to be causal for a very early onset of AD. METHODS: This is a case history using information from medical records, relative interviews and genetic testing results to describe the pre clinical prodrome and clinical course of a patient with EOFAD. RESULTS: A previously undescribed G206V mutation in PS1 was found in the proband. CONCLUSION: The G206V mutation in PS1 is probably causal of a case of EOFAD with significant premorbid features. PMID- 12112164 TI - The meaning of acute confusional state from the perspective of elderly patients. AB - OBJECTIVES: The aim of this study was to illuminate lived experience of having been in an acute confusional state (ACS) as narrated by elderly patients in orthopaedic care. METHOD: Qualitative study with phenomenological hermeneutic method for analysing the data based on narrative interviews. Fifty patients (67 96 years of age) who developed ACS during hospitalisation and in all cases the ACS ceased during their stay on the ward were interviewed once lucid again regarding the course of the event, their experiences, memories and interpretation of what had happened during the ACS. RESULTS: The meaning of the patients' lived experiences of being and having been confused was interpreted as 'Being trapped in incomprehensible experiences and a turmoil of past and present and here and there', comprising the themes trying to get a grip on the experience of the confusion, encountering past, present and the realm of the imagination as reality during the period of confusion and confronting the idea of having been confused. Contradictory to earlier research the patients remembered and could tell in great detail about their ACS. While confused, the confusional state means that impressions of all kinds invade the mind of the person and are experienced as reality, making him/her a victim of these impressions rather than the one who controls what comes into his/her mind. While in the middle of these experiences the person simultaneously senses that the impressions are unreal, thus indicating that he/she is in some sort of borderland between understanding and not understanding. The things that come into the mind of the person can either be frightening or neutral or enjoyable scenarios that seem to be mainly familiar but can also be unknown. These scenarios seem to be a mixture of past and present, of events and people while they seem to float from location to location. CONCLUSIONS: The findings indicates that what takes place during the ACS is not nonsense but probably a mix of the patient's life history, their present situation and above all a form of communication concerning their emotional state and inner experiences in this new situation. The findings also indicated that one possible approach to the patients is to confirm and support the patients in narrating their experiences both during the confusion and also after the ACS had ceased. PMID- 12112162 TI - Do visuospatial and constructional disturbances differentiate frontal variant of frontotemporal dementia and Alzheimer's disease? an experimental study of a clinical belief. AB - BACKGROUND: In recent years several attempts have been made to distinguish frontotemporal dementia (FTD) from Alzheimer's disease (AD) on neuropsychological grounds; in particular, it has been suggested that FTD patients show spared spatial abilities with respect to AD patients. OBJECTIVE: We aimed at verifying whether patients with the frontal variant of frontotemporal dementia (fv-FTD) and AD patients perform differently on visuospatial and constructional tasks. METHODS: We assessed a wide range of visuospatial abilities and provided a qualitative analysis of constructional performances in 14 fv-FTD patients and 11 AD patients, matched for general cognitive abilities. RESULTS: The two groups of patients achieved similar scores on two copying tasks, presented similar drawing procedures in copying Rey complex figure and made a similar quantitative and qualitative pattern of errors in copying simple geometrical drawings. Moreover, no significant difference was found between fv-FTD and AD patients on a specific battery for visuospatial abilities. CONCLUSIONS: Our data and a review of the literature suggest that basic visuospatial and constructional skills cannot be taken as a reliable diagnostic criterion for distinguishing fv-FTD and AD at a mild to moderate disease stage and that the clinical belief of spared spatial abilities in fv-FTD has to be referred to the lack of topographic disorientation in comparison to AD. PMID- 12112165 TI - The relation of White Matter Hyperintensities to implicit learning in healthy older adults. AB - OBJECTIVE: This study examined whether MRI evidence of cerebrovascular disease in the form of white matter hyperintensities (WMH) was associated with decreased implicit sequence learning performance in a high-functioning group of normal elderly volunteers. METHOD: One hundred and eight community-dwelling elderly individuals received an MRI and performed an implicit sequence learning task, the serial reaction time (SRT) task. RESULTS: Hyperintensities present in the white matter were associated with a decreased learning effect. This association was found with both deep white matter and periventricular changes. Other factors affecting SRT performance (i.e., baseline reaction time and switch-cost) were not significantly related to the presence of WMH. CONCLUSIONS: The results indicate that in addition to previously identified generalized cognitive deficits, WMH are also associated with a specific decrease in the implicit learning of sequences. PMID- 12112166 TI - Types of nursing home residents with self-destructive behaviours: analysis of the Harmful Behaviours Scale. AB - OBJECTIVE: To investigate the types of self-destructive behaviours identified by the Harmful Behaviours Scale (HBS) and the variables associated with them. METHOD: A cross sectional survey involving 647 residents in 11 nursing homes in the eastern suburbs of Sydney, Australia. The following instruments were used: Harmful Behaviours Scale (HBS); Behavioural Pathology in Alzheimer's Disease Rating Scale; Functional Assessment Staging Scale; Resident Classification Index; Health of the Nation Outcome Scale; Even Briefer Assessment Scales for Depression; Abbreviated Mental Test Scale; and the suicide item from the Structured Hamilton Depression Rating Scale. Diagnosis of dementia was obtained from nursing home records. RESULTS: Latent class analysis of the HBS identified four groups of residents, described as 'aggressive resistant' (34.9%), 'food refusal' (26.8%), 'behaviourally disturbed' (5.4%) and a 'non-symptomatic' group (33.0%) with little self-destructive behaviour. The behaviourally disturbed group engages in a widespread combination of direct and indirect self-destructive behaviours and displays other behavioural and psychological symptoms of dementia. In contrast, the food refusal group whose only behavioural symptom was refusal to eat and drink had the most cognitive impairment and did not show higher levels depression or suicidal ideation. CONCLUSIONS: We have found three groups of residents with self-destructive behaviours and each group is associated with a different pattern of variables. PMID- 12112167 TI - Policies, principles and pragmatism: old age psychiatrists' attitudes and practice regarding influenza immunisation for long stay patients. AB - This study aimed to examine the attitudes and practice of old age psychiatrists with respect to influenza immunisation for their patients in long stay care. A questionnaire was mailed out with a copy of the government immunisation policy. There was considerable disagreement among responders regarding the government policy, quality of life issues and the appropriateness of immunisation. There was a consensus in favour of immunising those who could not consent and for seeking relatives' views in this scenario. Staff immunisation status and patients' prior wishes were highlighted, amongst other factors, as affecting immunisation decisions. The government policy might be more acceptable to psychiatrists if there was more emphasis on the individual nature of clinical decisions and the policy will have to change in the light of new legislation. PMID- 12112168 TI - Influenza vaccination of people with dementia in long-stay wards in the United Kingdom: what is going on? PMID- 12112169 TI - Unrecognised and untreated depression in geriatric patients with hip fractures. PMID- 12112171 TI - The relationship between biological and environmental determinants of delusions in mild Alzheimer's disease patients. PMID- 12112170 TI - The agreement of the Mattis Dementia Rating Scale with the Mini-Mental State Examination. PMID- 12112173 TI - The Mini-Mental State exam may help in the differentiation of dementia with Lewy bodies and Alzheimer's disease. AB - OBJECTIVE: Since patients with dementia with Lewy bodies (DLB) tend to have greater impairment of attention and construction and better memory ability on neuropsychological tests than patients with Alzheimer's disease (AD), we determined if the items that measure attention, memory, and construction in the Mini-Mental State Examination (MMSE) help to distinguish DLB from AD early in the course of the dementia. DESIGN: We retrospectively studied the first available MMSE exam for each of our patients with DLB or AD and compared their MMSE subscores for attention, memory, and construction. SETTING: A university dementia brain bank in central Illinois, USA. PATIENTS: All patients with neuropathologically-proven DLB or AD with MMSE scores > or =13. RESULTS: We identified 17 DLB and 27 AD patients for whom we had MMSE exams. The attention and construction subtest scores of the DLB group were worse (p=0.0071 and p=0.0038, respectively) than those of the AD group. The memory subscores of the DLB group were better, although the difference did not reach statistical significance (p=0.22). When a mathematical equation was used to combine the three subscores with equal weighting (Attention-5/3Memory+5.Construction), the scores of the DLB group were worse (p=0.00007). Using this equation, a score less than 5 points was associated with DLB with a sensitivity of 0.82 (95% Confidence Interval (CI)=0.57-0.96) and a specificity of 0.81 (95% CI=0.62-0.94). CONCLUSIONS: Our findings support the work of others regarding the relative neuropsychological impairments of DLB and AD and indicate that the MMSE may be helpful in the differentiation of DLB and AD. PMID- 12112174 TI - Data management and quality assurance for an International project: the Indo-US Cross-National Dementia Epidemiology Study. AB - BACKGROUND: Data management and quality assurance play a vital but often neglected role in ensuring high quality research, particularly in collaborative and international studies. OBJECTIVE: A data management and quality assurance program was set up for a cross-national epidemiological study of Alzheimer's disease, with centers in India and the United States. METHODS: The study involved (a) the development of instruments for the assessment of elderly illiterate Hindi speaking individuals; and (b) the use of those instruments to carry out an epidemiological study in a population-based cohort of over 5000 persons. Responsibility for data management and quality assurance was shared between the two sites. A cooperative system was instituted for forms and edit development, data entry, checking, transmission, and further checking to ensure that quality data were available for timely analysis. A quality control software program (CHECKS) was written expressly for this project to ensure the highest possible level of data integrity. CONCLUSIONS: This report addresses issues particularly relevant to data management and quality assurance at developing country sites, and to collaborations between sites in developed and developing countries. PMID- 12112175 TI - Hippocampal volume and antidepressant response in geriatric depression. AB - BACKGROUND: Biological markers of treatment response may include structural brain changes seen on neuroimaging. While most imaging studies have focused on cerebrovascular disease, evidence is growing that the hippocampus may play a role in depression, particularly geriatric depression. METHOD: We studied 60 depressed elderly patients enrolled in a longitudinal study who were treated with antidepressant medications using a treatment guideline-based approach. Baseline and 12-week Montgomery-Asberg Depression Rating Scale (MADRS) scores were obtained via interview with a geriatric psychiatrist. All subjects had a baseline magnetic resonance imaging (MRI) brain scan. MRI scans were processed using standard protocols to determine total cerebral volume and right and left hippocampal volumes. Hippocampal volumes were standardized for total cerebral volume. MADRS scores less than 10 were used to define remission. RESULTS: When the group with the lowest quartile of standardized hippocampal volumes was compared to those above the first quartile, those with small right and total hippocampal volumes were less likely to achieve remission. In a subsequent logistic regression model controlling for age small standardized right hippocampal volumes remained significantly associated with remission. CONCLUSION: Further studies with larger sample are needed to determine if left-right hippocampal volume differences do exist in depression, and basic neuroscience studies will need to elucidate the role of the hippocampus in geriatric depression. PMID- 12112176 TI - Longitudinal increase in the volume of white matter hyperintensities in late onset depression. AB - BACKGROUND: Cerebrovascular disease is thought to play a role in the pathogenesis of geriatric major depression. One finding supporting such a "vascular depression" is the increased neuropathology in the form of white matter hyperintensities (WMH) found in patients diagnosed with a late-onset depression. However, at present there is little evidence that a longitudinal increase in WMH burden within an individual is associated with the onset of a late-life depression. METHODS: This study examined three-year longitudinal change in WMH volume and in cognition in: (a) an older man who developed his first episode of major depression during the study period, and (b) a comparison group of twelve older individuals who remained depression free. All subjects received at baseline and three years later a structural magnetic resonance imaging (MRI) using fast FLAIR technology. The images were analyzed with semi-automated computerized software to obtain WMH volumes. Subjects also received at both time points the Mini Mental State Exam (MMSE) as well a series of cognitive tasks assessing executive abilities (verbal fluency, Trail Making Test and Stroop test) since executive dysfunction is thought to be characteristic of a vascular depression. RESULTS: The individual who became depressed during the followup showed an increase in WMH volume that exceeded the 95% Confidence Intervals (CI) for change in the comparison group. This individual also showed a similar decline on the measures of executive function but not on the MMSE. CONCLUSIONS: These results are consistent with cerebrovascular disease being a factor in the pathogenesis of late-onset depression (i.e. "vascular depression"). PMID- 12112177 TI - Gender, aggression and serotonergic function are associated with response to sertraline for behavioral disturbances in Alzheimer's disease. AB - BACKGROUND: Indications for serotonergic medications in the treatment of behavioral disorders associated with Alzheimer's disease (AD) remain to be established. METHOD: Sertraline (100 mg OD) was evaluated in a double-blind, randomized, placebo-controlled cross-over study in 22 nondepressed patients with severe probable AD and significant behavioral disturbance. Each subject was given a fenfluramine challenge to evaluate central serotonergic tone. RESULTS: Eight of 21 (38%) completers responded to sertraline. Drug responsive behaviors included aggression/agitation, irritability and aberrant motor behavior. Low aggression, female gender and large prolactin increase were associated with a better response. There was a trend for decreased aggression during sertraline versus placebo (p = 0.08). CONCLUSION: Aggression, gender and serotonergic function were associated with sertraline response. Larger randomized controlled trials are needed to clarify the profile of responders. PMID- 12112178 TI - Gender related differences in clinical characteristics and hospital based resource utilization among older adults with schizophrenia. AB - OBJECTIVE: This report is an analysis of gender related differences in clinical characteristics and hospital based health resource utilization among older adults with schizophrenia and schizoaffective disorder in an acute care, state hospital over a one-year period. METHODS: This retrospective record review is an analysis of age of illness onset, psychiatric and medical comorbidity, hospital utilization, and psychotropic medication use. RESULTS: There were a total of 66 individuals with either schizophrenia or schizoaffective disorder. Mean age of this group was 55.2 +/- 4.62 years. Women were significantly over-represented among individuals with late onset schizophrenia and schizoaffective disorder. Men with schizophrenia had more comorbid substance abuse compared to women with schizophrenia (p < 0.05). Women and men did not differ significantly in hospital length of stay, amount or type of antipsychotic medication prescribed, or in utilization of seclusion/restraint in hospital. Both genders had substantial utilization of antipsychotic medication. Use of conventional antipsychotic medication monotherapy was always associated with use of anti-extrapyramidal symptom (anti-EPS) medication, while use of atypical antipsychotic medication monotherapy was more rarely associated with use of anti-EPS medication. CONCLUSIONS: In later life, women and men may have some areas of differing health care needs. Women in particular may benefit from psychoeducational approaches that address the experience of psychiatric illness of relatively recent onset (for example, symptom identification and acceptance of illness). Men may benefit from particular emphasis on treatment of comorbid substance abuse disorders. PMID- 12112179 TI - The relationship between delusions and depression in Alzheimer's disease. AB - OBJECTIVES: The aim of this investigation was to study the relationship between delusions and depression in Alzheimer's disease (AD). DESIGN: This was a cross sectional, case control study. SETTING: Neuropsychiatry Service, the Johns Hopkins School of Medicine, USA. PARTICIPANTS: 303 community-residing patients with probable AD according to NINCDS/ADRDA criteria were included in the study. Seventy-five patients with delusions only were compared to a control group of 228 patients who had neither delusions nor hallucinations. Patients with only hallucinations or both delusions and hallucinations were excluded. MEASURES: Patients were assessed clinically for the presence of delusions using the DSM-IV glossary definitions. They were also rated on standardized measures of depression, cognitive impairment, staging of dementia, general medical health, and functional impairment. RESULTS: There was an association between delusions and depression among patients with AD. Before adjustment for other variables, the presence of depression conferred a 1.8-fold (95% confidence intervals (CI) = 1.0 3.1; p = 0.04) higher risk of delusions. After adjustment for multiple other variables, this risk increased further to 6.8-fold (95% CI = 2.1-21.6; p = 0.001). CONCLUSIONS; Delusions in AD are strongly associated with depression after statistical adjustment for all confounding variables, which might distort this association. This finding has implications for our understanding of the etio pathogenesis and management of delusions and depression in AD. PMID- 12112180 TI - The prevalence of major depression or dysthymia among aged Medicare Fee-for Service beneficiaries. AB - RESEARCH OBJECTIVE: Estimates of the prevalence of major depression vary widely. Current estimates range from 2 to 14 % depending upon the definition and procedure for diagnosis. Further, most estimates are for special populations, either living in selected geographic areas or receiving specific types of medical care. A national survey of Medicare Fee-for-Service (FFS) beneficiaries provides an opportunity to assess the current level of major depression or dysthymia among a diverse population of older Americans. STUDY DESIGN: The Health Outcomes Survey (HOS) was administered to a national random sample of 1,000 Medicare FFS beneficiaries. We used the Mental Component Summary (MCS) measure of the SF-36 to estimate the prevalence of major depression or dysthymia. Logistic regression was used to examine associated factors. RESULTS: The response rate was 61.7%. Using an MCS score of 42 or lower, prevalence of major depression or dysthymia was estimated to be 25% for respondents age 65 years and older. Logistic regression analysis revealed that the likelihood of major depression or dysthymia was associated with years of education (Odds Ratio (OR) = 0.87), difficulties performing activities of daily living (OR = 1.72), and Medicaid enrollment (OR = 2.67). CONCLUSIONS: The results revealed that one-quarter of the respondents reported mental health problems consistent with major depression or dysthymia. This is higher than previously reported. Like previous studies, years of education, physical impairment, and poverty are strong predictors of major depression or dysthymia. The high rate of major depression or dysthymia implies there may be considerable unmet need among elderly Medicare FFS beneficiaries for diagnosing and treating mental illness. PMID- 12112181 TI - Risk of mortality for dementia in a developing country: the Yoruba in Nigeria. AB - BACKGROUND: Limited data exist on the impact of dementia in developing nations, including its association with mortality. OBJECTIVE: The purpose of this paper is to assess the relationship between dementia and five-year mortality on a community dwelling elderly Yoruba population in the developing country of Nigeria and to compare those results with those from an elderly African-American community in Indianapolis. METHODS: A two-phase design was used to ascertain dementia status in two sites. In the first phase, the Community Screening Instrument for Dementia (CSI-D) was administered. In the second phase, subjects were sampled for the clinical assessment according to their CSI-D performance category. Proportional hazards regression was used to assess the relationship between mortality and cognitive status at both sites after adjusting for demographics and chronic disease conditions. RESULTS: For the entire screened population, poor and intermediate performance on the CSI-D is associated with increased mortality at both sites; however the effect of CSI-D performance did not significantly differ between the two sites. For the clinically assessed sample, dementia was significantly associated with increased mortality at both sites (Ibadan RR = 2.83, Indianapolis RR = 2.05), but the effect was not significantly different across the two sites. CONCLUSION: Dementia resulted in an increased risk of mortality for Yoruba of a magnitude similar to African Americans suggesting that the impact of dementia on mortality risk may be similar for developing and developed countries. PMID- 12112182 TI - The clinical profile of older patients' response to antidepressants--an open trial of sertraline. AB - BACKGROUND: A number of studies have examined the predictive utility and time to response of rating scales and demographic variables. Very few community samples have been examined in this way, and no studies examining the prognostic validity of early symptomatic response have been found in the literature. OBJECTIVES: This study aims to describe how treatment response is reflected in rating scales in older community residents treated with sertraline and to explore the utility of these instruments in predicting response. METHODS: The study examines the open label therapeutic and continuation phases of a maintenance trial. RESULTS: 225 older depressed community residents were treated (openly) with sertraline. Fifty three percent had a good outcome, 13% did not respond to sertraline and had a poor long-term prognosis. Increased age was associated with poor outcome and increased anxiety symptoms with a good outcome. In the compliant sub-sample, GMS/AGECAT schizophrenia symptoms were associated with poor response to treatment. Baseline HDRS items and related symptom clusters were not of predictive utility, however early changes in HDRS score (improvement from baseline of four or more by four weeks) was associated with good outcome. All symptom clusters improved within two weeks of treatment with sleep symptoms improving by six weeks. Optimum symptomatic improvement was achieved by eight weeks. CONCLUSIONS: Clinicians in primary care can expect 53% response to treatment. In the absence of symptomatic improvement by one month (HDRS score of four or more) treatment should be reviewed. Optimum treatment response is usually achieved within eight weeks. PMID- 12112183 TI - Sodium valproate in the treatment of aggressive behavior in patients with dementia--a randomized placebo controlled clinical trial. AB - OBJECTIVES: The efficacy and tolerability of sodium valproate 2 x 240 mg compared to placebo were investigated in aggressive behavior in dementia. DESIGN: A randomized, placebo controlled, double-blind cross-over design. The trial included a baseline period (one week); a placebo period (three weeks); a wash-out period with placebo (one week); and a treatment period with sodium valproate (three weeks). SETTING: A psychogeriatric short-stay ward at a psychiatric teaching hospital. PARTICIPANTS: Demented patients who met Patel's criteria for aggressive behavior and had a score of > or =3 on at least one of the items of the Social Dysfunction and Aggression scale-9 (SDAS-9). INTERVENTION: A fixed dose of sodium valproate 2 x 6 ml of a 40 mg/ml suspension (daily defined dose of 480 mg) was compared to placebo. MEASUREMENTS: Primary outcome variables were changes of the score of SDAS-9 and Clinical Global Impression scale (CGI) performed at the last week of each treatment period. RESULTS: Data of 42 patients (F=25 and M=17; age 80.4+/-6.8 years) were analyzed. Treatment with sodium valproate showed no differences compared to placebo on aggressive behavior. The mean plasma level of sodium valproate was 40.9+/-10.8 microg/ml. Regression analysis showed a trend for improvement between the plasma levels of sodium valproate and the SDAS-9 and the CGI scores. Adverse events were not related to the plasma levels of sodium valproate. Secondary outcome measurements showed significant improvement on restless, melancholic and anxious behavior; a trend for improvement was found on suspicious and dependent behavior. Possible limitations of this study are the low dose of sodium valproate, the relatively short treatment period (three weeks), and the absence of statistical corrections for multiple comparisons. CONCLUSION: This study showed no effect of sodium valproate 2 x 240 mg over placebo on aggressive behavior in dementia. PMID- 12112184 TI - Feasibility of treating mild cognitive impairment with cholinesterase inhibitors. PMID- 12112185 TI - Voting practices of the residents of nursing and residential homes. PMID- 12112187 TI - Current awareness in geriatric psychiatry. PMID- 12112188 TI - Rasmussen Syndrome and movement disorder. PMID- 12112189 TI - Fetal neural grafts for Huntington's disease: a prospective view. AB - Intrastriatal transplantation of striatal neuroblasts from human fetuses is a promising approach for treatment of Huntington's disease, on the basis of many experimental animal studies and, most recently, pilot clinical trials. Technically, several issues remain to be resolved (e.g., the precise site of dissection of the fetal tissue; the number and location of the fetal striatal implants; or the use of immunosuppressive therapy), and await larger-scale trials and purposely designed protocols. Further clinical data must also be obtained, and preliminary promising results must be replicated in a patient group large enough to provide conclusive results. It is important to establish (1) the amount of clinical benefit provided to the patient by the grafted cells; (2) the anticipated duration of clinical benefits; and (3) the secondary rate of decline after the benefit of the graft has been overbalanced. Evaluation of these parameters will require very long-term follow-up of the patients involved, over several years after grafting, before the technique can eventually be proposed widely to patients. PMID- 12112190 TI - Beyond the iron mask: towards better recognition and treatment of depression associated with Parkinson's disease. AB - This review examines the frequency of depression complicating Parkinson's disease (PD), its aetiology and clinical features, and also how it may be recognised and treated. Studies investigating the frequency of depression in PD have yielded figures ranging between 2.7% and 70%. Methodological differences account for much of the disparity. The aetiology of depression in PD is complex, and probably relates to both biological and exogenous factors. Dysfunction of multiple neurotransmitter systems, including the serotonergic system, may be involved. Mood disturbances resulting from deep brain stimulation of the subthalamic nucleus may provide a fruitful area for future research, and assist our understanding of the neural networks involved in mediating depression. Several recent studies have confirmed that depression in the PD patient is a major determinant of quality of life and that this is closely related to dysfunction in other clinically important health areas. The validity for many existing scales in the screening, diagnosis, and monitoring of depression in the PD patient has not been established. The Montgomery-Asberg Depression Rating Scale and the Hamilton Rating Scale for Depression appear to have good diagnostic sensitivity and specificity when compared with DSM-IV criteria. Recommendations for the optimal drug treatment of depression in PD are difficult to give, due to an inexplicable dearth of sizeable, placebo-controlled studies. A majority of physicians would probably now opt for a selective serotonin reuptake inhibitor in the depressed PD patient. There is no good evidence that these drugs are associated with a worsening of motor features, but they should probably not be coprescribed with selegiline, because of the risk of causing a potentially serious serotonin syndrome. Several studies have suggested that depression in the PD patient is associated with a more rapid deterioration in cognitive and motor functions, perhaps as a surrogate marker for more extensive brainstem cell loss. PMID- 12112191 TI - Characterisation of striatal NMDA receptors involved in the generation of parkinsonian symptoms: intrastriatal microinjection studies in the 6-OHDA lesioned rat. AB - Treatments for Parkinson's disease based on replacement of lost dopamine have several problems. Following loss of dopamine, enhanced N-methyl-D-aspartate (NMDA) receptor-mediated transmission in the striatum is thought to be part of the cascade of events leading to the generation of parkinsonian symptoms. We determined the localisation and pharmacological characteristics of NMDA receptors that play a role in generating parkinsonian symptoms within the striatum. Rats were lesioned unilaterally with 6-hydroxydopamine (6-OHDA), and cannulae implanted bilaterally to allow injection of a range of NMDA receptor antagonists at different striatal sites. When injected rostrally into the dopamine-depleted striatum, the glycine site partial agonist, (+)-HA-966 (44-400 nmol) caused a dose-dependent contraversive rotational response consistent with an antiparkinsonian action. (+)-HA-966 (400 nmol) had no effect when infused into more caudal regions of the dopamine-depleted striatum, or following injection into any striatal region on the dopamine-intact side. To determine the pharmacological profile of NMDA receptors involved in inducing parkinsonism in 6 OHDA-lesioned rats, a range of NMDA receptor antagonists was infused directly into the rostral striatum. Ifenprodil (100 nmol) and 7-chlorokynurenate (37 nmol), but not MK-801 (15 nmol) or D-APV (25 nmol) elicited a dramatic rotational response when injected into the dopamine-depleted striatum. This pharmacological profile is not consistent with an effect mediated via blocking NR2B-containing NMDA receptors. The effect of intrastriatal injection of ifenprodil was increased in animals previously treated with levodopa (L-dopa) methyl ester. This was seen as an increase in on-time and in peak rotational response. We propose that stimulation of NR2B-containing NMDA receptors in the rostral striatum underlies the generation of parkinsonian symptoms. These studies are in line with previous findings suggesting that administration of NR2B-selective NMDA receptor antagonists may be therapeutically beneficial for parkinsonian patients, when given de novo and following L-dopa treatment. PMID- 12112192 TI - Repetitive magnetic stimulation of cortical motor areas in Parkinson's disease: implications for the pathophysiology of cortical function. AB - We investigated the neurophysiological and clinical effects of repetitive magnetic stimulation (rTMS) delivered to the cortical motor areas in healthy subjects and patients with Parkinson's disease. rTMS was delivered with a high speed magnetic stimulator (Cadwell, Kennewick, WA) through a figure-eight coil centred on the primary motor area at a stimulus intensity of 120% motor threshold. Trains of 10 stimuli were delivered at frequencies of 5 Hz while subjects were at rest and during a voluntary contraction of the contralateral first dorsal interosseous muscle. In normal subjects at rest, the muscle evoked responses (MEPs) to each stimulus in a train of magnetic stimuli progressively increased in size during the train. rTMS left the MEPs unchanged in patients off therapy and had a small facilitatory effect in those on therapy. In normal subjects and patients, 5-Hz rTMS trains delivered during a voluntary contraction of the target muscle left the MEP unchanged in size. MEPs were followed by a silent period that increased in duration during the course of the train. The silent period duration increased to a similar extent in patients and controls. The reduced rTMS-induced facilitation of MEPs in patients with Parkinson's disease reflects a decreased facilitation of the excitatory cells in the cortical motor areas. PMID- 12112193 TI - Evaluation of sleep and driving performance in six patients with Parkinson's disease reporting sudden onset of sleep under dopaminergic medication: a pilot study. AB - Six patients with Parkinson's disease (PD) reporting unusually fast or sudden onset of sleep under the addition of dopamine agonists to a previous levodopa containing therapy were examined using a sleep-wake diary, the Epworth sleepiness scale (ESS), polysomnography, multiple sleep latency tests (MSLT), a standardized vigilance test, and driving simulation. In all patients, ESS scores were increased and polysomnography showed disruption of the sleep pattern, a tendency towards poor sleep efficiency, and reduced proportions of slow- wave and rapid eye movement sleep. Pathological results in the MSLT or the vigilance test were obtained in five cases. For evaluation of driving performance, the standard deviation from the mean lane position during driving simulation was calculated. Three of five patients had clearly increased mean SDLP values. With respect to the measurement of daytime sleepiness (ESS, MSLT, vigilance test, and driving simulation), each patient had pathological results in at least two of these examinations. However, only a limited transfer of the routine vigilance assessment to driving performance was possible. In summary, this pilot study indicates that unusually fast or sudden onset of sleep in PD patients is a phenomenon of daytime sleepiness. PMID- 12112194 TI - Autosomal dominant adult neuronal ceroid lipofuscinosis: parkinsonism due to both striatal and nigral dysfunction. AB - We describe a family with adult neuronal ceroid lipofuscinosis, with apparent autosomal dominant inheritance, observed in six affected individuals in three generations. Disease onset was usually in the fifth decade, but was earlier in the youngest generation. Early symptoms consisted of myoclonus in face and arms, epilepsy, auditory symptoms, cognitive decline, or depression. Parkinsonism occurred a few years after disease onset, with stooped posture, shuffling gait, bradykinesia, and mask face. Four subjects deteriorated to a state of severe handicap, with severe dementia, contractures, dysphagia, and dysarthria. Leg weakness evolved to flaccid paraparesis in two patients. Diagnosis was confirmed by brain biopsy in one patient and full autopsy in two patients. Abundant intraneuronal storage of autofluorescent material was found throughout the brain. Electron microscopy showed granular osmiophilic deposits and scarce fingerprint profiles. Striking loss of neurons in the substantia nigra pars compacta and reticulata was found. (123)I-IBZM Single photon emission computed tomography in two patients showed loss of postsynaptic D2 receptor binding in the striatum. We conclude that parkinsonism in ANCL is likely to be caused by both presynaptic nigral cell loss and postsynaptic striatal degeneration. PMID- 12112195 TI - Reaction time is not impaired by stimulation of the ventral-intermediate nucleus of the thalamus (Vim) in patients with tremor. AB - We studied the effect of high-frequency electrical stimulation of the ventral intermediate nucleus of the thalamus (Vim) in four patients implanted with chronic stimulators to determine whether this procedure adversely affects reaction time to a proprioceptive stimulus. Two patients had undergone this surgery for treatment of tremor resulting from Parkinson's disease insufficiently responsive to levodopa therapy and two patients for treatment of essential tremor. Reaction times to auditory, visual, cutaneous, and proprioceptive stimuli were tested in a simple motor task requiring flexion of the elbow joint to a visual target in response to each stimulus. Reaction times were tested postoperatively with and without the stimulator turned on. We found that reaction time for all stimulus modalities was not increased when the stimulator was turned on; in fact, reaction times were, on average, slightly shorter during stimulation, but this difference was not statistically significant. We conclude that transmission of somatosensory inputs, necessary for initiating voluntary movement, from the periphery to the cortex is not significantly impaired by stimulation of the ventral-intermediate nucleus of the thalamus in patients with pathological tremor. PMID- 12112196 TI - Descending control of muscles in patients with cervical dystonia. AB - It was reported recently that specific features in the frequency analysis of electromyographic (EMG) activity in the sternocleidomastoid (SCM) and splenius (SPL) muscles were able to distinguish between rotational idiopathic cervical dystonia (CD) and voluntary torticollis in individual subjects. Those with CD showed an abnormal drive to muscles at 5 to 7 Hz and an absence of the normal 10 to 12 Hz peak in the autospectrum of SPL. We sought to determine whether the same abnormalities in the frequency domain are found in complex CD, in which the head is displaced in more than two planes. EMG activity was recorded in the SCM, SPL, trapezius, and levator scapulae muscles bilaterally in 10 patients with complex CD. Frequency analysis of EMG was compared with conventional clinical and polymyographic assessment. The autospectrum of SPL during free dystonic contraction showed an absence of a significant peak at 10 to 12 Hz in 8 of the 10 patients. The presence of a 5 to 7 Hz frequency drive showed a significant association with muscle pairs determined as dystonic by means of polymyography (P < 0.005). The neck posture predicted blindly, based on the low-frequency drive, correlated significantly with the clinical assessment of posture (P < 0.01). Conventional assessment and the results of frequency analysis correlated, suggesting that a low-frequency drive to neck muscle may be a general feature of simple rotational and more complex cervical dystonia. The pattern of coherence between the EMG in different neck muscles may provide a means of identifying leading dystonic muscles, especially in patients with complex cervical dystonia. PMID- 12112197 TI - Dopamine miracle: from brain homogenate to dopamine replacement. PMID- 12112198 TI - Case-control study of estrogen receptor gene polymorphisms in Parkinson's disease. AB - We investigated the association of Parkinson's disease (PD) with two estrogen receptor gene polymorphisms. In a sample of 319 unrelated PD cases and 196 control subjects including both men and women, we observed no association of PD with the estrogen receptor genotypes derived from XbaI and PvuII digests. Analyses restricted to women or to cases and controls of European origin yielded similar findings. Further analyses stratified by age at examination or by family history did not show associations. While exogenous and endogenous estrogen may modify the risk of PD in women, the two estrogen receptor gene polymorphisms considered here do not seem to contribute to PD susceptibility. PMID- 12112199 TI - Diagnosing Parkinson's disease using videotaped neurological examinations: validity and factors that contribute to incorrect diagnoses. AB - Field work is commonly required in movement disorders research. Sending neurologists into the field can be logistically challenging and costly. Alternatively, neurological examinations may be videotaped and reviewed later. There is little knowledge of the validity of the videotaped neurological examination in the diagnosis of Parkinson's disease (PD). We examined the validity of the videotaped Unified Parkinson's Disease Rating Scale (UPDRS) motor examination in the diagnosis of PD, and sought to determine which factors are associated with incorrect diagnoses. PD patients and controls were enrolled in a familial aggregation study between August of 1998 and June of 2000, and as part of that study each was examined by a physician who performed an in-person UPDRS motor examination. Each also underwent a second, videotaped UPDRS motor examination. Based on the review of this videotape, a neurologist, who was blinded to the previous clinical diagnosis, assigned a diagnosis of PD or normal. A total of 211 of 231 PD patients (sensitivity = 91.3%), and 170 of 172 controls (specificity = 98.8%) were correctly identified based on the videotape. True positives had a higher mean rest tremor score (1.7 vs. 0.3; P < 0.001), action tremor score (0.9 vs. 0.3; P < 0.001), bradykinesia score (11.2 vs. 7.4; P = 0.02), and disease of longer mean duration (8.9 vs. 5.8 years; P = 0.001) than false negatives. False negatives did not differ from true positives in terms of age, total dose of levodopa, Hoehn and Yahr score, or rigidity, gait and posture, or facial masking scores (each assessed during the in-person examination). The videotaped UPDRS motor examination is a useful means of diagnosing PD and provides an alternative approach for the diagnosis of PD in field studies. A limitation is that patients with milder PD of shorter duration may not be recognized as PD. PMID- 12112201 TI - Disordered respiration as a levodopa-induced dyskinesia in Parkinson's disease. AB - Symptomatic respiratory disturbance as a consequence of levodopa (L-dopa) therapy for Parkinson's disease (PD) has been described only rarely and may be underrecognized in clinical practice. We report on two patients with PD in whom the introduction or augmentation of L-dopa therapy was associated with the development of irregular and rapid breathing. Analysis of breathing patterns before and after L-dopa demonstrated a striking change in respiratory rate after administration of L-dopa, with the emergence of irregular tachypnea alternating with brief periods of apnea, in a pattern consistent with a central origin. In both cases, the temporal relationship of the respiratory disturbance to the administration of L-dopa suggested a peak-dose drug effect. Previous reports of L dopa-induced respiratory dyskinesia are reviewed, and the potential mechanisms whereby L-dopa might influence the central control of respiration to produce irregular breathing patterns are discussed. PMID- 12112200 TI - [123I]beta-CIT SPECT distinguishes vascular parkinsonism from Parkinson's disease. AB - We investigated whether [(123)I]-beta-CIT and single-photon emission computed tomography (SPECT) imaging distinguishes patients with clinically suspected vascular parkinsonism (VP) from patients with idiopathic Parkinson's disease (PD). [(123)I]beta-CIT SPECT is a sensitive marker of dopaminergic degeneration, and the degree of striatal binding reduction in PD correlates with disease severity. Thirteen patients who fulfilled rigid clinical criteria for VP (mean +/ S.D.: age, 76.5 +/- 5.3 years; disease duration, 3.6 +/- 2.8 years), 20 PD patients (age, 66.2 +/- 9.5 years; disease duration, 4.3 +/- 2.7 years), and 30 healthy persons (age, 44.6 +/- 19.2 years) underwent [(123)I]beta-CIT SPECT imaging. Age-corrected striatal beta-CIT binding was reduced on average by 40.8% in PD but was near normal in the VP group (mean reduction, 1.2%). This difference was statistically significant (Z = 4.68; P < 0.001). The left-right asymmetry of striatal beta-CIT binding was significantly increased in the PD group compared with normal controls and the VP group (F(2) = 17.4, P <0.001). Moreover, putamen caudate nucleus ratios were significantly reduced in PD compared with both VP patients and healthy controls (F(2) = 65.5, P < 0.001). Whole striatal beta-CIT binding was more than one standard deviation above the mean PD values in all but one of the individual VP patients. Our findings suggest that the presynaptic dopaminergic deficits seen in PD are absent in most patients with VP. [(123)I]beta-CIT SPECT imaging may be useful to help distinguish between PD and VP patients during life. PMID- 12112202 TI - Therapeutic efficacy of bilateral prefrontal slow repetitive transcranial magnetic stimulation in depressed patients with Parkinson's disease: an open study. AB - Recent studies have suggested that both high- and low-frequency repetitive transcranial magnetic stimulation (rTMS) have antidepressant effects in patients with major depression. We conducted an open study to assess the effects of slow rTMS on mood changes in patients with depression associated with Parkinson's disease (PD). Ten depressed patients with PD (four with major depression and six with dysthymia) received daily sessions of rTMS (frequency, 0.5 Hz; pulse duration, 0.1 msec; field intensity, 10% above the motor threshold) over both prefrontal regions (a total of 100 stimuli per prefrontal region daily) over 10 consecutive days. This treatment resulted in a moderate but significant decrease in scores of the Hamilton Depression Rating Scale (33-37%) and the Beck Depression Inventory (24-34%), which persisted 20 days after finishing the stimulation. In parallel, we observed mild improvement (18-20%) of motor symptoms. No significant adverse effects were reported. These preliminary results suggest the therapeutic potential of daily prefrontal low-frequency rTMS (0.5 Hz) in depression associated with PD. PMID- 12112203 TI - Bilateral pallidotomy in Parkinson's disease: a retrospective study. AB - We evaluated the effects of bilateral pallidotomy in patients with advanced Parkinson's disease. Thirteen patients with Parkinson's disease had a staged bilateral pallidotomy if they had severe response fluctuations, dyskinesias, painful dystonia, or bradykinesia despite optimum pharmacological treatment. Assessment scales were the Unified Parkinson's Disease Rating scale (UPDRS), the Schwab and England scale, and a questionnaire on the effects of disability in activities of daily living and adverse effects. Postoperative magnetic resonance imaging was evaluated for lesion location and extension. The median off-phase UPDRS motor score was reduced from 43.5 to 29 after the first pallidotomy, and it was further reduced to 23.5 after the second pallidotomy (n = 8). The UPDRS activities of daily living off-phase score improved from 28.5 to 20.5 after the first pallidotomy and to 19 after the second pallidotomy (n = 6). The Schwab and England scale off-phase score showed an improvement after both procedures, first from 40 to 60, and thereafter to 90 (n = 8). On-phase dyskinesias were reduced substantially. Eight patients had adverse effects, of whom five had problems with speech. One patient became hemiplegic due to a delayed infarction. Ten patients experienced further benefit from the second procedure. Bilateral pallidotomy reduces dyskinesias. A second contralateral pallidotomy may reduce parkinsonism, although to a lesser degree compared with the first pallidotomy and with an increased risk for adverse effects. PMID- 12112204 TI - Health-related quality of life in patients with advanced Parkinson's disease treated with deep brain stimulation of the subthalamic nuclei. AB - Parkinson's disease (PD) is a progressive neurodegenerative disorder for which there is as yet no cure. It affects many aspects of patients' lives, only some of which can be monitored by available clinical rating scales. In the past decade, there has been a new emphasis on the use of health-related quality of life (HRQOL) measures to describe patient response to treatment. We describe patient reported HRQOL in subjects who underwent bilateral deep brain stimulation (DBS) of the subthalamic nuclei (STN) for the treatment of PD, compared with a similar group of subjects who did not receive surgical treatment. A consecutive series of patients (n = 11) with advanced idiopathic PD were treated with DBS of the STN. This surgically treated group was compared prospectively with a similar group of patients (n =13) awaiting surgery. Self-reported HRQOL, measured by the Parkinson's Disease Questionnaire (PDQ-39) was evaluated at three time periods T(0), T(3), and T(6). The surgery group was evaluated according to the Unified Parkinson's Disease Rating Sale (UPDRS) before (T(0)), 3 (T(3)), and 6 months (T(6)) after surgery. HRQOL, UPDRS part II and III, duration of off periods, and dyskinesias improved significantly from T(0) to T(3) and from T(0) to T(6) for the surgery group but not for the nonsurgery group. Ten of the 11 patients treated with DBS of the STN reported a lower summary score (indicating better HRQOL) 6 months after surgery. The results of this prospective controlled study suggest that patients with advanced idiopathic PD treated with DBS of the STN obtain significant improvements in patient reported HRQOL and in clinical outcomes 3 and 6 months after surgery. PMID- 12112205 TI - Effects of a startle on heart rate in patients with multiple system atrophy. AB - The patient cooperation usually required for neurophysiological assessment of autonomic cardioregulatory function is difficult to obtain from patients with bradykinesia. A particularly interesting condition occurs in multiple system atrophy (MSA), which features both bradykinesia and autonomic dysfunction. Another characteristic of patients with MSA is their normal motor reaction to a startling stimulus. We used startle as a stimulus for testing autonomic cardioregulatory function in patients with MSA, thus avoiding the need for patient cooperation. In 10 healthy volunteers and 8 MSA patients, we recorded the electrocardiographic QRS complex with surface electrodes attached over the chest and delivered an acoustic startle stimulus after 8 seconds of baseline recording. We calculated the ratio between the pre-stimulus and the post-stimulus heart beat intervals (R-R ratio) by dividing the mean prestimulus R-R interval by the shortest R-R interval obtained within 10 seconds poststimulus. Healthy volunteers had a significant shortening of the R-R interval. The peak of the effect occurred after 2 to 5 seconds, with a mean R-R ratio of 1.14 (S.D. = 0.09). In contrast, R R shortening was markedly reduced in patients, even though they had a normal motor response. The mean R-R ratio in patients was 1.03 (S.D. = 0.03), significantly lower than in healthy volunteers (P < 0.01). Our results demonstrate an abnormally reduced modulation of the heart beat frequency in patients with MSA, compatible with a dysfunction on pathways responsible for autonomic regulation. The method described here may be useful in the assessment of cardioregulatory function in poorly cooperative patients with normal startle responses. PMID- 12112206 TI - Further extension of the H1 haplotype associated with progressive supranuclear palsy. AB - The recent finding of disequilibrium among several polymorphisms along the tau gene and the strong association of one of the two haplotypes formed by these polymorphisms (H1) with progressive supranuclear palsy (PSP) suggests that a single allele in or near the tau gene at 17q21 is responsible for increased risk in most of the PSP cases. We sought to determine whether mutations in the tau gene are responsible for the disease in 45 sporadic PSP patients. Furthermore, we analyzed some markers located in the common region of linkage (D17S800-D17S791), associated with some cases of familial frontotemporal dementia (FTDP-17), and the SNPs rs1816 and rs937 close to the tau gene, to determine their possible association with sporadic PSP. We did not find pathogenic mutations in exons 9, 10, 12, or 13 of the tau gene, indicating that tau mutations in both the splice site region of the exon 10 and in the microtubule-binding region of tau gene are not a cause of PSP in this study group. We found significant overrepresentation of the haplotypes H1, extended up to the promoter of the tau gene (H1P), in PSP patients as compared with controls. In addition, a significant overrepresentation of the D17S810 2/2 and 3/2 genotypes, of the SNP rs1816 A/A, and of the SNP rs937 delG/delG genotypes was detected in PSP, further extending the haplotype described previously. These results are consistent with the hypothesis that a change either in the 5' or in the 3' flanking regions of the tau gene, or even other genes contained in the H1E haplotype, could increase the genetic susceptibility to PSP. PMID- 12112208 TI - Corticobasal degeneration syndrome with basal ganglia calcification: Fahr's disease as a corticobasal look-alike? AB - A 57-year-old man with a 5-year history of progressive left-sided rigidity and apraxia had extensive bilateral calcification of basal ganglia, centrum semiovale, dentate nuclei, and cerebellar white matter on brain imaging. The case is an example of radiological Fahr's disease accompanying a clinical syndrome of corticobasal degeneration. Possible pathogenetic and nosological implications of this association are discussed. PMID- 12112207 TI - Steele-Richardson-Olszewski-syndrome: reduction of dopamine D2 receptor binding relates to the severity of midbrain atrophy in vivo: (123)IBZM SPECT and MRI study. AB - Patients with the clinical diagnosis of progressive supranuclear palsy (PSP) show heterogeneous neuropathological findings. In neuropathologically proven cases with numerous neurofibrillary tangles of neuropil threads, the brainstem and striatum are always affected. We compared (123)I-iodobenzamide single photon emission computed tomography (IBZM-SPECT) for imaging of striatal dopamine D(2) receptors in vivo with high-resolution magnetic resonance imaging (MRI) in 13 patients with possible or probable PSP. Clinically, all patients exhibited similar signs including supranuclear vertical-down-gaze palsy, axial rigidity especially involving the neck, bradykinesia, instability of balance with easy falls, and a poor response to dopaminergic drugs. Specific striatal dopamine D(2) receptor binding in IBZM-SPECT was reduced in 10 patients but was normal in three patients. Mean midbrain diameter was 23.7 mm. The reduction of IBZM-binding was statistically significantly correlated to midbrain atrophy (P = 0.010) either in all 13 patients or in those without striatal or white matter lesions (P = 0.015). We suggest that technical investigations, mainly MRI, are able to corroborate the in vivo diagnosis, if PSP is clinically suspected. PMID- 12112210 TI - Severe generalized dystonia due to primary putaminal degeneration: case report and review of the literature. AB - Putaminal lesions of a variety of etiologies may cause secondary dystonia. We report on a case of primary putaminal degeneration as a cause of severe childhood onset generalized dystonia and review the literature of the pathology of dystonia. A 44-year-old patient with severe generalized childhood-onset dystonia and macrocephaly underwent neurological evaluation and neuropathological examination. Neurological examination was normal apart from dystonia and signs referable to prior cryothalamotomy. Workup for metabolic and genetic causes of dystonia was negative. Neuroimaging showed severe bilateral putaminal degeneration, which subsequently correlated with the neuropathological findings of gliosis, spongiform degeneration, and cavitation. The substantia nigra pars compacta contained a normal number of neurons but decreased tyrosine hydroxylase immunoreactivity. There were no histopathological markers of other metabolic or degenerative diseases. PMID- 12112209 TI - Dopamine transporter density is decreased in parkinsonian patients with a history of manganese exposure: what does it mean? AB - Manganese (Mn) exposure can cause parkinsonism. Pathological changes occur mostly in the pallidum and striatum. Two patients with a long history of occupational Mn exposure presented with Mn-induced parkinsonism. In one patient, magnetic resonance imaging (MRI) showed findings consistent with Mn exposure, and Mn concentration was increased in the blood and urine. However, this patient's clinical features were typical of idiopathic Parkinson disease (PD). Previous pathological and positron emission tomography studies indicate that striatal dopamine transporter density is normal in Mn-induced parkinsonism, whereas it is decreased in PD. Therefore, we performed [(123)I]-(1r)-2 beta-carboxymethoxy 3beta-(4-iodophenyl)tropane ([(123)I]-beta-CIT) single-photon emission computed tomography. Severe reduction of striatal beta-CIT binding was indicated, which is consistent with PD. We propose three interpretations: (1) the patients have PD, and Mn exposure is incidental; (2) Mn induces selective degeneration of presynaptic dopaminergic nerve terminals, thereby causing parkinsonism; or (3) Mn exposure acts as a risk of PD in these patients. Our results and careful review of previous studies indicate that the axiom that Mn causes parkinsonism by pallidal lesion may be over-simplified; Mn exposure and parkinsonism may be more complex than previously thought. Further studies are required to elucidate the relationship between Mn and various forms of parkinsonism. PMID- 12112212 TI - Effect of treatment with intravenous immunoglobulin on quality of life in patients with stiff-person syndrome. AB - The therapeutic effects of intravenous immunoglobulin (IVIG) on the stiff-person syndrome (SPS) have been described exclusively in case reports or open-label studies in terms of clinical outcomes. We investigate whether IVIG improves quality of life (QoL) in the SPS. Six patients with the classic form of SPS completed a generic QoL instrument, the SF-36, and a Visual Analogue Scale (VAS) before treatment as well as 2 weeks after completion of a course of IVIG. There was significant improvement in the SF-36 subscores for pain, social functioning, general mental health, and energy-vitality with treatment. The VAS also improved significantly. We conclude that treatment with IVIG improves QoL in the SPS. PMID- 12112211 TI - Friedreich's ataxia with chorea and myoclonus caused by a compound heterozygosity for a novel deletion and the trinucleotide GAA expansion. AB - Friedreich's ataxia (FRDA) is the most common hereditary ataxia, affecting about 1 in 50,000 individuals. It is caused by mutations in the frataxin gene; 98% of cases have homozygous expansions of a GAA trinucleotide in intron 1 of the frataxin gene. The remaining 2% of patients are compound heterozygotes, who have a GAA repeat expansion in one allele and a point mutation in the other allele. FRDA patients with point mutation have been suggested to have atypical clinical features. We present a case of compound heterozygotes in a FRDA patient who has a deletion of one T in the start codon (ATG) of the frataxin gene and a GAA repeat expansion in the other allele. The patient presented with chorea and subsequently developed FRDA symptoms. The disease in this case is the result of both a failure of initiation of translation and the effect of the expansion. This novel mutation extends the range of point mutations seen in FRDA patients, and also broadens the spectrum of FRDA genotype associated with chorea. PMID- 12112213 TI - Effects of apomorphine on flexor reflex and periodic limb movement. AB - It has been suggested that periodic leg movements (PLM) and spinal flexor reflex (FR) share common mechanisms. Although dopaminergic agents improve PLM in humans and strongly influence spinal FR circuitry in animal studies, its effects on FR have not been documented in humans. We describe a 65-year-old man with PLM after overnight withdrawal of dopaminergic agents. The electromyographic pattern of spontaneous PLM closely resembled that of the FR elicited by medial plantar nerve stimulation. Thirty minutes after subcutaneous injection of apomorphine, both PLM and FR were completely abolished. These findings demonstrate that dopaminergic agents can suppress exaggerated FR in humans, and support the hypothesis of common mechanisms for PLM and FR. PMID- 12112214 TI - Camptocormia in Parkinson's disease mimicked by focal myositis of the paraspinal muscles. AB - We report on a 63-year-old man with idiopathic Parkinson's disease who developed kyphosis and a severe forward flexion of the thoracolumbar spine. A typical feature was an increase during walking or standing and it completely disappeared in the supine position, mimicking the clinical phenomenon of camptocormia (bent spine). In addition to the abnormal posture, a weakness of the erector spinal muscles, local pain, reddening, and elevated temperature of the paraspinal muscles were evident. Creatine kinase was initially elevated, electromyography showed spontaneous activity and a myopathic pattern. Magnetic resonance imaging and bioptic examinations revealed a focal myositis of the paraspinal muscles. This case indicates that camptocormia can be mimicked by focal myositis of paraspinal muscles and must be included in the differential diagnosis, especially when additional symptoms as inflammatory signs or weakness are present. PMID- 12112215 TI - Reversible parkinsonian syndrome in systemic and brain vasculitis. AB - A young female patient with chronic renal failure due to a systemic vasculitis and a parkinsonian syndrome secondary to brain vasculitis, most likely systemic lupus erythematosus, is described. The patient had a dramatic response to a pulse of methylprednisolone, with remission of her parkinsonian symptoms. PMID- 12112217 TI - Spongiform encephalopathy mimicking corticobasal degeneration. AB - The presentation of subacute spongiform encephalopathies (SSE) is varied. The following case of SSE presented clinically similar to corticobasal degeneration. The SSE diagnosis was suspected because of magnetic resonance imaging (MRI) findings and confirmed pathologically. PMID- 12112216 TI - Paradoxical response to apomorphine in a case of atypical parkinsonism. AB - We describe a patient with clinical signs of parkinsonism showing a paradoxical response to apomorphine injection. We discuss possible pathogenetic mechanisms with regard to the literature and suggest the diagnosis of a striatonigral degeneration at an early stage. PMID- 12112218 TI - Possible sporadic rapid-onset dystonia-parkinsonism. AB - Rapid-onset dystonia-parkinsonism is a hereditary disease characterized by a combination of dystonic and parkinsonian symptoms. Bulbar musculature is predominantly affected by dystonia. The onset is usually abrupt and the progression of the disease over years is minimal or absent. Homovanillic acid levels in cerebrospinal fluid can be diminished, suggesting that the pathogenesis of the disease is related to some dysfunction in dopaminergic neurotransmission. However, no abnormality has been found in positron emission tomography studies and levodopa does not improve symptoms. The genetic abnormality is not known, but evidence for linkage to markers on chromosome 19q13 has been reported. We describe the case of a woman with a clinical picture highly suggestive of rapid onset dystonia-parkinsonism (RDP) and no family history of the disease. PMID- 12112219 TI - Dystonia, athetosis, and epilepsia partialis continua in a patient with late onset Rasmussen's encephalitis. AB - Rasmussen's encephalitis is a rare autoimmune disorder characterized by intractable epilepsy and progressive hemispheric dysfunction. The disorder usually affects children, although cases have been reported with symptom onset in late adolescence or adulthood. Myoclonus is common in Rasmussen patients, usually occurring as part of epilepsia partialis continua (EPC); however, other hyperkinetic movements are rare. This report documents a 19-year-old woman with Rasmussen's encephalitis whose clinical presentation was dominated by foot dystonia, arm athetosis, and EPC. Intravenous immunoglobulin improved both hyperkinetic movements and EPC, but benefit was transient. The clinical significance and implications of these findings are discussed. PMID- 12112220 TI - Myoclonic dystonia as unique presentation of isolated vitamin E deficiency in a young patient. AB - We describe a young patient affected by vitamin E deficiency with mutation in the tocopherol transfer protein alleles and the unique presentation as myoclonic dystonia, which was practically the only symptom for 6 years before ataxia became evident. Vitamin E supplementation markedly improved both symptoms. This unusual clinical phenotype must be considered, because isolated vitamin E deficiency is eminently treatable. PMID- 12112221 TI - Levetiracetam in the treatment of paroxysmal kinesiogenic choreoathetosis. AB - Anticonvulsants are frequently used in the treatment of paroxysmal kinesiogenic choreoathetosis (PKC). Although they are often extremely effective in eliminating paroxysmal movements, short- and long-term side-effects may limit their use in young patients. Levetiracetam (Keppra), a novel antiepileptic drug approved for the treatment of partial seizures is well tolerated in patients with epilepsy. We report on the use of levetiracetam in the treatment of PKC. Levetiracetam was effective in eliminating paroxysmal events and should be considered as an alternative to standard antiepileptic medications in this disorder. PMID- 12112222 TI - Painful arm and moving fingers: clinical features of four new cases. AB - The syndrome of painful arm and moving fingers associates pain in one arm or hand with involuntary movement of one or several fingers. In the four cases described, an association between a central and a peripheral nervous system lesion is demonstrated or suspected. Treatment of the condition is disappointing. PMID- 12112223 TI - Hallervorden-Spatz syndrome resembling a typical Tourette syndrome. AB - A young man presenting with a Tourette syndrome-like disorder that was the main clinical manifestation of Hallervorden-Spatz syndrome is described. It is recommended that, even in the case of slow progression, HSS should be considered in the differential diagnosis of TS-like disorders. PMID- 12112224 TI - Tardive tremor due to metoclopramide. AB - Tardive tremor is a very uncommon neuroleptic-induced tardive syndrome which was initially described in five patients by Stacy and Jankovic (Stacy and Jankovic, Mov Disord 1992;7:53-57). Since then, there have been only three additional case reports attesting to the apparent rarity of this condition, although it is unknown whether other unreported cases have been observed. We describe a patient with persistent tardive tremor that was associated with tardive dyskinesia, who closely resembles previously reported cases. PMID- 12112226 TI - Subthalamotomy for end-stage severe Parkinson's disease. PMID- 12112225 TI - Treatment of pseudobulbar laughter after gamma knife thalamotomy. AB - We describe a case of pathological laughter after gamma knife thalamotomy which resolved after treatment with sertraline. It is important to identify this potentially treatable complication of surgical therapy. PMID- 12112228 TI - Stop codon decoding in Candida albicans: from non-standard back to standard. AB - The human pathogen Candida albicans translates the standard leucine-CUG codon as serine. This genetic code change is mediated by a novel ser-tRNA(CAG), which induces aberrant mRNA decoding in vitro, resulting in retardation of the electrophoretic mobility of the polypeptides synthesized in its presence. These non-standard decoding events have been attributed to readthrough of the UAG and UGA stop codons encoded by the Brome Mosaic Virus RNA 4, which codes for the virion coat protein, and the rabbit globin mRNAs, respectively. In order to fully elucidate the behaviour of the C. albicans ser-tRNA(CAG) towards stop codons, we have used other cell-free translation systems and reporter genes. However, the reporter systems used encode several CUG codons, making it impossible to distinguish whether the slow migration of the polypeptides is caused by the replacement of leucines by serines at the CUG codons, readthrough, or a combination of both. Therefore, we have constructed new reporter systems lacking CUG codons and have used them to demonstrate that aberrant mRNA decoding in vitro is not a result from stop codon readthrough or any other non-standard translational event. Our data show that a single leucine to serine replacement at only one of the four CUG codons encoded by the BMV RNA-4 gene is responsible for the aberrant migration of the BMV coat protein on SDS-PAGE, suggesting that this amino acid substitution (ser for leu) significantly alters the structure of the virion coat protein. The data therefore show that the only aberrant event mediated by the ser-tRNA(CAG) is decoding of the leu-CUG codon as serine. PMID- 12112229 TI - The SCR1 gene from Schwanniomyces occidentalis encodes a highly hydrophobic polypeptide, which confers ribosomal resistance to cycloheximide. AB - In Saccharomyces cerevisiae, the SCR1 gene from Schwanniomyces occidentalis is known to induce ribosomal resistance to cycloheximide (cyh). A 2.8 kb DNA fragment encoding this gene was sequenced. Its EMBL Accession No. is AJ419770. It disclosed a putative tRNA(Asn) (GUU) sequence located downstream of an open reading frame (ORF) of 1641 nucleotides. This ORF was shown to correspond to SCR1. It would encode a highly hydrophobic polypeptide (SCR1) with 12 transmembrane domains. SCR1 is highly similar to a variety of yeast proteins of the multidrug-resistance (MDR) family. However, SCR1 only conferred resistance to cyh but not to benomyl or methotrexate. The cyh-resistance phenotype induced by SCR1 was confirmed in several S. cerevisiae strains that expressed this gene to reside at the ribosomal level. In contrast, a beta-galacosidase-tagged SCR1 was found to be integrated in the endoplasmic reticulum (ER). It is proposed that the ribosomes of yeast cells expressing SCR1 undergo a conformational change during their interaction with the ER, which lowers their affinity for cyh-binding. If so, these findings would disclose a novel ribosomal resistance mechanism. PMID- 12112230 TI - A history of research on yeasts 4: cytology part II, 1950-1990. PMID- 12112231 TI - Homing at an extragenic locus mediated by VDE (PI-SceI) in Saccharomyces cerevisiae. AB - PI-SceI (VDE), a homing endonuclease with protein splicing activity, is a genomic parasite in the VMA1 gene of Saccharomyces cerevisiae. In a heterozygous diploid of the VDE-less VMA1 allele and a VDE-containing VMA1 allele, VDE specifically cleaves its recognition sequence (VRS) in the VDE-less VMA1 allele at meiosis, followed by 'homing', i.e. a conversion to a VDE-containing allele. We found that upon VDE expression, homing of a marker gene at an extragenic locus occurs only when a 45 bp element containing the VRS is inserted at its allelic site, while mutants of VDE with no endonuclease activity lack authentic extragenic homing activity. Thus, both the VRS and VDE are required for homing. Insertion of the VRS in a homozygous diploid significantly lowered the spore germination ability, indicating that a template for gene repair at its allelic locus is essential for efficient homing and survival of yeast cells. PMID- 12112232 TI - Saccharomyces cerevisiae Big1p, a putative endoplasmic reticulum membrane protein required for normal levels of cell wall beta-1,6-glucan. AB - Deletion of Saccharomyces cerevisiae BIG1 causes an approximately 95% reduction in cell wall beta-1,6-glucan, an essential polymer involved in the cell wall attachment of many surface mannoproteins. The big1 deletion mutant grows very slowly, but growth can be enhanced if cells are given osmotic support. We have begun a cell biological and genetic analysis of its product. We demonstrate, using a Big1p-GFP fusion construct, that Big1p is an N-glycosylated integral membrane protein with a Type I topology that is located in the endoplasmic reticulum (ER). Some phenotypes of a big1Delta mutant resemble those of strains disrupted for KRE5, which encodes another ER protein affecting beta-l,6-glucan levels to a similar extent. In a big1Deltakre5Delta double mutant, both the growth and alkali-soluble beta-l,6-glucan levels were reduced as compared to either single mutant. Thus, while Big1p and Kre5p may have similar effects on beta-l,6-glucan synthesis, these effects are at least partially distinct. Residual beta-l,6-glucan levels in the big1Deltakre5Delta double mutant indicate that these gene products are unlikely to be beta-l,6-glucan synthase subunits, but rather may play some ancillary roles in beta-l,6-glucan synthase assembly or function, or in modifying proteins for attachment of beta-l,6-glucan. PMID- 12112233 TI - Schizosaccharomyces pombe spPABP, a homologue of Saccharomyces cerevisiae Pab1p, is a non-essential, shuttling protein that facilitates mRNA export. AB - Poly(A)-binding proteins play important roles in mRNA metabolism in eukaryotic cells. We examined the role of the Schizosaccharomyces pombe homologue of the Saccharomyces cerevisiae poly(A)-binding protein, Pab1p, in cellular growth and mRNA export. In contrast to PAB1, the sppabp gene is not essential for cellular viability. Like the human hPABP1 protein, spPABP is cytoplasmically localized and can shuttle between the nucleus and the cytoplasm. We found that a spPABP-GFP fusion protein expressed from a multicopy plasmid could suppress the growth and mRNA export defect of rae1-16 7 nup184-1 synthetic lethal mutations. However, about 20-25% of cells in the population exhibited a pronounced nuclear accumulation of poly(A)(+) RNA. The same cells also localized the spPABP-GFP fusion to the nucleus, suggesting that the shuttling ability of spPABP is related to its function in mRNA export. When a heterologous nuclear export activity from spMex67p was fused to spPABP-GFP fusion protein, it overcame the nuclear retention but did not increase nuclear mRNA export. We discuss the implications of these observations in relation to how spPABP could function in mRNA export. Published in 2002 by John Wiley & Sons, Ltd. PMID- 12112235 TI - Current awareness on yeast. PMID- 12112236 TI - Two mechanisms for oxidation of cytosolic NADPH by Kluyveromyces lactis mitochondria. AB - Null mutations in the structural gene encoding phosphoglucose isomerase completely abolish activity of this glycolytic enzyme in Kluyveromyces lactis and Saccharomyces cerevisiae. In S. cerevisiae, the pgi1 null mutation abolishes growth on glucose, whereas K.lactis rag2 null mutants still grow on glucose. It has been proposed that, in the latter case, growth on glucose is made possible by an ability of K. lactis mitochondria to oxidize cytosolic NADPH. This would allow for a re-routing of glucose dissimilation via the pentose-phosphate pathway. Consistent with this hypothesis, mitochondria of S. cerevisiae cannot oxidize NADPH. In the present study, the ability of K. lactis mitochondria to oxidize cytosolic NADPH was experimentally investigated. Respiration-competent mitochondria were isolated from aerobic, glucose-limited chemostat cultures of the wild-type K. lactis strain CBS 2359 and from an isogenic rag2Delta strain. Oxygen-uptake experiments confirmed the presence of a mitochondrial NADPH dehydrogenase in K.lactis. This activity was ca. 2.5-fold higher in the rag2Delta mutant than in the wild-type strain. In contrast to mitochondria from wild-type K. lactis, mitochondria from the rag2Delta mutant exhibited high rates of ethanol dependent oxygen uptake. Subcellular fractionation studies demonstrated that, in the rag2Delta mutant, a mitochondrial alcohol dehydrogenase was present and that activity of a cytosolic NADPH-dependent 'acetaldehyde reductase' was also increased. These observations indicate that two mechanisms may participate in mitochondrial oxidation of cytosolic NADPH by K. lactis mitochondria: (a) direct oxidation of cytosolic NADPH by a mitochondrial NADPH dehydrogenase; and (b) a two-compartment transhydrogenase cycle involving NADP(+)- and NAD(+)-dependent alcohol dehydrogenases. PMID- 12112237 TI - A microarray-assisted screen for potential Hap1 and Rox1 target genes in Saccharomyces cerevisiae. AB - Saccharomyces cerevisiae adapts to altered oxygen availability by differentially expressing a number of genes. Under aerobic conditions oxygen control of gene expression is exerted through the activator Hap1 and the repressor Rox1. The Hap1 transcription factor senses cellular heme status and increases expression of aerobic genes in response to oxygen. The repression of hypoxic genes under normoxic conditions results from Hap1-mediated activation of ROX1 transcription. To allow the identification of additional Hap1 and Rox1 target genes, genome-wide expression was analysed in aerobically, chemostat-cultivated hap1 and rox1 null mutants. The microarray results show that deletion of HAP1 causes a lower transcript level of 51 genes. Transcription of 40 genes was increased in rox1 mutant cells compared to wild-type cells. Combining these results with our previously described transcriptome data of aerobically and anaerobically grown cells and with computational analysis of the promoters identified 24 genes that are potentially regulated by Hap1, and 38 genes satisfied the criteria of being direct targets of Rox1. In addition, this work provides further evidence that Rox1 controls transcription of anaerobic genes through repression under normoxic conditions. PMID- 12112238 TI - Two non-complementing genes encoding enzymatically active methylenetetrahydrofolate reductases control methionine requirement in fission yeast Schizosaccharomyces pombe. AB - By transforming two methionine auxotrophic mutants from fission yeast Schizosaccharomyces pombe with a wild-type gene library, we defined two genes, met9 and met11, which both encode a methylenetetrahydrofolate reductase. The genes cannot complement each other. We detected single transcripts for both. In vitro measurements of enzymatic activities showed that the met11-encoded enzyme was responsible for only 15-20% of the total methylenetetrahydrofolate reductase activity. A strain in which gene met9 was disrupted required significantly more methionine for full growth and efficient mating and sporulation than the strain disrupted for gene met11. The in vitro and in vivo data thus indicated that met9 was the major expressed gene. Our results are in accordance with the assumption that the two methylenetetrahydrofolate reductases generate the methyl groups necessary for methionine synthetase to convert homocysteine to methionine, and suggest that expression of the two genes is an important parameter in the control of methionine biosynthesis. PMID- 12112239 TI - Post-translational modifications of the AFET3 gene product: a component of the iron transport system in budding cells and mycelia of the yeast Arxula adeninivorans. AB - The yeast Arxula adeninivorans is characterized by a temperature-dependent dimorphism. A. adeninivorans grows as budding cells at temperatures up to 42 degrees C, but forms mycelia at higher temperatures. A strong correlation exists between morphological status and iron uptake, achieved by two transport systems that differ in iron affinity. In the presence of high Fe(II) concentrations (>2 microm), budding cells accumulate iron concentrations up to seven-fold higher than those observed in mycelia, while at low Fe(II) concentrations (<2 microm), both cell types accumulate similar amounts of iron. The copper-dependent Fe(II) oxidase Afet3p, composed of 615 amino acids, is a component of the high-affinity iron transport system. This protein shares a high degree of homology with other yeast iron transport proteins, namely Fet3p of Saccharomyces cerevisiae, Cafet3p of Candida albicans and Pfet3p of Pichia pastoris. Expression of the AFET3 gene is found to be strongly dependent on iron concentration but independent of the morphological stage; however, cell morphology was found to influence post translational modifications of the gene product. O-glycosylation was observed in budding cells only, whereas N-glycosylation occurred in both cell types. The N glycosylated 103 kDa glycoprotein matures into the 108.5 kDa form, further characterized by serine phosphorylation. Both N-glycosylation and phosphorylation occur at low iron concentrations (< or =5 microm). The mature Afet3p of 108.5 kDa is uniformly distributed within the plasma membrane in cells of both morphological stages. PMID- 12112240 TI - Sequencing and functional analysis of the Hansenula polymorpha genomic fragment containing the YPT1 and PMI40 genes. AB - A 6.0 kb genomic DNA segment was isolated by its ability to rescue the temperature-sensitive growth defect and the hypersensitivity to sodium deoxycholate of a spontaneous vanadate-resistant mutant derived from Hansenula polymorpha DL-1. The genomic fragment contains four open reading frames homologous to the Saccharomyces cerevisiae genes YPT1 (which codes for a GTP binding protein; 75% amino acid identity), PMI40 (encoding phosphomannose isomerase; 61% identity), YLR065c (30% identity) and CST13 (28% identity). The H. polymorpha YPT1 homologue (HpYPT1) was found to be responsible for the complementation of the temperature-sensitive phenotype and the sodium deoxycholate sensitivity of the mutant strain. Disruption of the H. polymorpha PMI40 homologue (HpPMI40) resulted in the auxotrophic requirement for D-mannose. The heterologous expressions of HpYPT1 and HpPMI40 were able to complement the temperature-sensitive phenotype of S. cerevisiae ypt1-1 mutant and the mannose auxotrophy of S. cerevisiae pmi40 null mutant, respectively, indicating that the H. polymorpha genes encode the functional homologues of S. cerevisiae YPT1 and PMI40 proteins. The nucleotide sequence has been submitted to GenBank under Accession No. AF454544. PMID- 12112241 TI - Dehydroepiandrosterone (DHEA) metabolism in Saccharomyces cerevisiae expressing mammalian steroid hydroxylase CYP7B: Ayr1p and Fox2p display 17beta hydroxysteroid dehydrogenase activity. AB - We have engineered recombinant yeast to perform stereospecific hydroxylation of dehydroepiandrosterone (DHEA). This mammalian pro-hormone promotes brain and immune function; hydroxylation at the 7alpha position by P450 CYP7B is the major pathway of metabolic activation. We have sought to activate DHEA via yeast expression of rat CYP7B enzyme. Saccharomyces cerevisiae was found to metabolize DHEA by 3beta-acetylation; this was abolished by mutation at atf2. DHEA was also toxic, blocking tryptophan (trp) uptake: prototrophic strains were DHEA resistant. In TRP(+) atf2 strains DHEA was then converted to androstene 3beta,17beta-diol (A/enediol) by an endogenous 17beta-hydroxysteroid dehydrogenase (17betaHSD). Seven yeast polypeptides similar to human 17betaHSDs were identified: when expressed in yeast, only AYR1 (1-acyl dihydroxyacetone phosphate reductase) increased A/enediol accumulation, while the hydroxyacyl-CoA dehydrogenase Fox2p, highly homologous to human 17betaHSD4, oxidized A/enediol to DHEA. The presence of endogenous yeast enzymes metabolizing steroids may relate to fungal pathogenesis. Disruption of AYR1 eliminated reductive 17betaHSD activity, and expression of CYP7B on the combination background (atf2, ayr1, TRP(+)) permitted efficient (>98%) bioconversion of DHEA to 7alpha-hydroxyDHEA, a product of potential medical utility. PMID- 12112243 TI - Current awareness. PMID- 12112242 TI - The role of the 5' untranslated region (UTR) in glucose-dependent mRNA decay. AB - When S. cerevisiae are grown with glucose, SDH2 mRNA encoding the iron protein of the succinate dehydrogenase complex is unstable and present at low level. In yeast grown without glucose, SDH2 mRNA is stable and its level rises. Addition of glucose to a glucose-limited culture causes the SDH2 mRNA level to fall rapidly with a half-life of approximately 5-7 min. Previously the 5'UTR of the mRNA of SDH2 was shown to be necessary and sufficient to destabilize it in glucose (Lombardo et al., 1992). We now show that the SDH1 and SUC2 5'UTRs are capable of conferring glucose-sensitive mRNA instability. We also examine how changes in the SDH2 5'UTR affect glucose-triggered degradation. Finally, we show that changes in mRNA stability are correlated with changes in translational efficiency for these transcripts. PMID- 12112244 TI - Genetic epidemiology and microarrays. PMID- 12112245 TI - Microarrays and genetic epidemiology: a multipurpose tool for a multifaceted field. AB - The advent of molecular technologies that allow the collection and analysis of large amounts of genetic data is rapidly transforming the field of genetic epidemiology. Whether monitoring infectious outbreaks or identifying genotypic variations that underlie disease susceptibility, genetic epidemiology relies heavily on the analysis of multiple, independently derived results. By allowing the simultaneous monitoring of thousands of genetic or expression data points, microarrays are emerging as particularly powerful tools. Several recent reviews have described array manufacturing and the types of scientific questions that can exploit this technology, but few have addressed how the intended use of an array can dictate its design. This review will focus on this latter issue, with particular emphasis on the genetic epidemiology of infectious disease. The design of arrays for genotyping, expression profiling, and fingerprinting are presented, and examples of recent epidemiological studies are used to illustrate the applications' strong points and limitations. In addition to discussing arrays' ability to provide global views of gene identity or function, the review will describe design options for creating arrays that detect multiple genetic variations. It will also examine the reliability of array-generated fingerprints, assay accessibility, and possibilities for sharing and comparing data across studies. Although many challenges lie ahead, microarrays' multiple abilities appear uniquely poised to accelerate the advance of genetic epidemiology's multiple fronts. PMID- 12112246 TI - Design of studies using DNA microarrays. AB - DNA microarrays are assays that simultaneously provide information about expression levels of thousands of genes and are consequently finding wide use in biomedical research. In order to control the many sources of variation and the many opportunities for misanalysis, DNA microarray studies require careful planning. Different studies have different objectives, and important aspects of design and analysis strategy differ for different types of studies. We review several types of objectives of studies using DNA microarrays and address issues such as selection of samples, levels of replication needed, allocation of samples to dyes and arrays, sample size considerations, and analysis strategies. PMID- 12112247 TI - Role of gene expression microarray analysis in finding complex disease genes. AB - The promise of gene expression studies using microarray technology has inspired much new hope for finding complex diseases genes. It has become clear that complex diseases result from collective actions of many genetic and nongenetic factors. Therefore, genetic dissection of complex diseases should be carried out in a global context. The technology of gene expression microarray analysis (GEMA) can provide such global information on transcription activities of essentially all genes simultaneously. It is hoped that this promising technology can be applied to samples drawn from large-scale, well-defined genetic epidemiological studies and help us untangle the web of pathways leading to complex diseases. However, extremely noisy GEMA data pose serious challenges in terms of the statistical methodologies needed. Extensive work is needed in order to respond to the challenges before one can fully utilize the potential power provided by GEMA. We begin in this paper by identifying several statistical problems related to the application of GEMA to genetic epidemiological analysis, and consider study designs that might benefit from this promising new technology. While it is still too early to tell how much of the enormous potential of GEMA will be realized ultimately, its success will probably depend most critically on the ability of statistical genetics to rise to the challenge of mining information from a sea of noise. PMID- 12112249 TI - Empirical bayes methods and false discovery rates for microarrays. AB - In a classic two-sample problem, one might use Wilcoxon's statistic to test for a difference between treatment and control subjects. The analogous microarray experiment yields thousands of Wilcoxon statistics, one for each gene on the array, and confronts the statistician with a difficult simultaneous inference situation. We will discuss two inferential approaches to this problem: an empirical Bayes method that requires very little a priori Bayesian modeling, and the frequentist method of "false discovery rates" proposed by Benjamini and Hochberg in 1995. It turns out that the two methods are closely related and can be used together to produce sensible simultaneous inferences. PMID- 12112248 TI - Symbolic discriminant analysis of microarray data in autoimmune disease. AB - New laboratory technologies such as DNA microarrays have made it possible to measure the expression levels of thousands of genes simultaneously in a particular cell or tissue. The challenge for genetic epidemiologists will be to develop statistical and computational methods that are able to identify subsets of gene expression variables that classify and predict clinical endpoints. Linear discriminant analysis is a popular multivariate statistical approach for classification of observations into groups. This is because the theory is well described and the method is easy to implement and interpret. However, an important limitation is that linear discriminant functions need to be prespecified. To address this limitation and the limitation of linearity, we have developed symbolic discriminant analysis (SDA) for the automatic selection of gene expression variables and discriminant functions that can take any form. In the present study, we demonstrate that SDA is capable of identifying combinations of gene expression variables that are able to classify and predict autoimmune diseases. PMID- 12112250 TI - Mantel statistics to correlate gene expression levels from microarrays with clinical covariates. AB - Mantel statistics provide an additional step to standard approaches in the analysis of gene expression and covariate data, allow the calculation of standard statistics such as correlation, partial correlation, and regression coefficients, and, with permutation tests, provide P values for these statistics to relate the sample covariates to the expression levels. In this article we describe the Mantel statistics and illustrate their use and interpretation with data from a study of seven human oligodendrogliomas (brain tumors) where expression levels of 1013 genes and five covariates were previously analyzed using standard approaches. In the previous analysis of these data, qualitative relationships were found between gene expressions and two of the clinical covariates. We show in this article how the Mantel statistics are able to formally quantify and provide P values to determine statistical significance of these relationships. We also show how the Mantel statistics can be used to rank subsets of genes, found using standard clustering methods, in terms of differential expression across samples. PMID- 12112251 TI - Rapid 18O analysis of small water and CO2 samples using a continuous-flow isotope ratio mass spectrometer. AB - High-frequency throughput is often needed in isotopic studies in biological and medical fields. Here we report that high-precision oxygen isotope ratio measurements of water (+/-0.13 per thousand) were rapidly and routinely made on small samples (40-100 microL) using an isotope ratio mass spectrometer operated in continuous-flow mode. Simple modifications to existing instrumentation allow for rapid manual analyses of dilute CO2 (10% CO2/90% N2), including the addition of a septum port and water trap prior to the gas chromatography (GC) column (elemental analyzer column in this study) and the extension of fused-silica capillary tubing between the mass spectrometer source and the effluent tubing from the GC column (located within the CONFLO unit on Finnigan mass spectrometers). We routinely analyzed 20 small-volume samples per hour using this technique, without sacrificing precision of the oxygen isotope ratio measurement. PMID- 12112252 TI - Liquid chromatography/electrospray tandem mass spectrometric analysis of incurred ractopamine residues in livestock tissues. AB - Ractopamine HCl is a beta-adrenergic agonist (beta-agonist) recently approved by the U.S. Food and Drug Administration, but not other governmental agencies, for use in finishing swine. For these reasons, it was important to develop and validate mass spectrometric methods for the detection and confirmation of ractopamine residues in livestock marker tissues. Incurred tissues in cattle, sheep, turkeys, and ducks were generated during 7-day ractopamine feeding (20 ppm in diets) periods. Disposition of ractopamine residues in liver and pigmented retinal epithelium was determined in animals slaughtered with withdrawal periods of 0, 3, and 7 days. Ractopamine residues, purified using solid-phase extraction, were measured using liquid chromatography (LC) and electrospray with detection by tandem mass spectrometry (MS/MS) in the multiple reaction-monitoring (MRM) mode. Total ractopamine residues (parent ractopamine + hydrolyzed conjugates) in liver were detected in all species on withdrawal day 0 (2-97 ppb) and were greatly diminished in all species by withdrawal day 7 (<1 ppb). Bovine and ovine retina had lower levels of ractopamine (0.5-3 ppb) than liver, and occular residues increased with withdrawal time, suggesting redistribution into this tissue. Lower limits of quantification were found to be approximately 0.1 ppb in liver and retina. Incurred ractopamine residues were confirmed by the precise and accurate agreement of MRM intensity ratios of diagnostic fragment ions (m/z 284, 164, and 121) from the protonated molecule between ractopamine residues in incurred samples and an authentic ractopamine standard. The limits of confirmation in liver and retina using recognized acceptance criteria were below 1 ppb. The high sensitivity and specificity for measurement and confirmation of ractopamine residues suggests this method will be applicable for regulatory residue surveillance programs. PMID- 12112253 TI - Direct-injection high performance liquid chromatography ion trap mass spectrometry for the quantitative determination of olanzapine, clozapine and N desmethylclozapine in human plasma. AB - A specific and sensitive direct-injection high performance liquid chromatography electrospray ionization tandem mass spectrometry (HPLC/ESI-MS/MS) method has been developed for the rapid identification and quantitative determination of olanzapine, clozapine, and N-desmethylclozapine in human plasma. After the addition of the internal standard dibenzepin and dilution with 0.1% formic acid, plasma samples were injected into the LC/MS/MS system. Proteins and other large biomolecules were removed during an online sample cleanup using an extraction column (1 x 50 mm i.d., 30 microm) with a 100% aqueous mobile phase at a flow rate of 4 mL/min. The extraction column was subsequently brought inline with the analytical column by automatic valve switching. Analytes were separated on a 5 microm Symmetry C18 (Waters) analytical column (3.0 x 150 mm) with a mobile phase of acetonitrile/0.1% formic acid (20:80, v/v) at a flow rate of 0.5 mL/min. The total analysis time was 6 min per sample. The inter- and intra-assay coefficients of variation for all compounds were <11%. By eliminating the need for extensive sample preparation, the proposed method offers very large savings in total analysis time. PMID- 12112254 TI - Detection of norbolethone, an anabolic steroid never marketed, in athletes' urine. AB - Norbolethone (13-ethyl-17-hydroxy-18,19-dinor-17alpha-pregn-4-en-3-one) is a 19 nor anabolic steroid first synthesized in 1966. During the 1960s it was administered to humans in efficacy studies concerned with short stature and underweight conditions. It has never been reported by doping control laboratories. Norbolethone was identified in two urine samples from one athlete by matching the mass spectra and chromatographic retention times with those of a reference standard. The samples also contained at least one likely metabolite. The samples were also unusual because the concentrations of endogenous steroids were exceptionally low. Since norbolethone is not known to be marketed by any pharmaceutical company, a clandestine source of norbolethone may exist. PMID- 12112255 TI - Sample preparation in matrix-assisted laser desorption/ionization mass spectrometry of whole bacterial cells and the detection of high mass (>20 kDa) proteins. AB - Three sample preparation strategies commonly employed in matrix-assisted laser desorption/ionization mass spectrometry (MALDI-TOFMS) of whole bacterial cells were investigated for the detection of high mass signals; these included the dried droplet, the seed-layer/two-layer, and the bottom-layer methods. Different sample preparation approaches favoured the detection of high- or low-mass proteins. The low-mass peaks were best detected using the bottom-layer method. By contrast, the dried droplet method using a solvent with higher water content, and hence effecting a slower crystallization process, gave the best results for the detection of high-mass signals. Signals up to m/z 158 000 could be detected with this methodology for Bacillus sphaericus. Sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) analysis of the same extracts used for MALDI-TOFMS showed bands in the molecular weight range in which high-mass peaks were observed in MALDI-MS, suggesting that the high-mass signals are not polymeric adducts of low-mass protein monomers. In addition, one of the high molecular weight proteins (approximately 126 kDa) was putatively identified as an S-layer protein by an in gel tryptic digest. The bacterial samples spotted on the target wells for MALDI TOFMS, using the different sample preparation strategies, were examined under a scanning electron microscope and differences were observed between the different strategies, suggesting that the nature of the crystals and the distribution of the analytes amidst the crystals could influence the spectral pattern observed in MALDI-TOFMS of whole bacterial cells. Finally, evidence is presented to indicate that, although the determinants are intact cells, cell lysis occurs both before and during the MALDI process. PMID- 12112256 TI - Singlet-singlet annihilation in ultraviolet matrix-assisted laser desorption/ionization studied by fluorescence spectroscopy. AB - Laser-induced fluorescence spectroscopy was carried out on microcrystalline samples of three typical matrices under conditions of matrix-assisted laser desorption/ionization (MALDI). The emitted fluorescence intensity was determined as a function of incident laser fluence and a sublinear increase of the fluorescence intensity with laser fluence was found. A very good fit was obtained when the experimental fluorescence vs. fluence data were compared with a numerical model assuming that under typical MALDI fluence conditions a large fraction of molecules in the excited singlet state undergoes singlet-singlet annihilation. Throughout the fluence range relevant for MALDI, however, the experimental data could not be fit well to a model assuming resonant two-photon absorption as the process depopulating the singlet state. In a separate set of experiments, the singlet lifetimes of several typical crystalline MALDI matrices were determined and found to be considerably shorter than previously reported. While both singlet-singlet annihilation and resonant two-photon absorption have been discussed in the literature as candidates for pathways to primary matrix ion generation in MALDI, the data presented here suggest that singlet-singlet annihilation is the dominant mechanism for depopulating the singlet state in a matrix crystal excited at typical MALDI fluences. PMID- 12112257 TI - Quantitative determination of polar and ionic compounds in petroleum fractions by atmospheric pressure chemical ionization and electrospray ionization mass spectrometry. AB - The capabilities of atmospheric pressure chemical ionization (APCI) and electrospray ionization (ESI) methods for quantitative analysis of polar and ionic compounds in petroleum fractions have been examined. The requirements of the analysis for sensitivity, linear dynamic range, and structural characterization have been discussed. ESI was found to be approximately two orders of magnitude more sensitive than APCI and is most suitable for the detection of analytes in weak concentrations. Equivalent relative linear dynamic ranges were observed by the two methods (at least three orders of magnitude). For the relatively high analyte concentrations examined here (e.g., 1-100 ppm or higher), the absolute area counts increased linearly with the analyte amount only in APCI, making this method more attractive for quantitative liquid chromatography/mass spectrometry (LC/MS) applications. Nevertheless, a wider range of ionic compounds can be detected by ESI than by APCI. PMID- 12112258 TI - A simple algorithm improves mass accuracy to 50-100 ppm for delayed extraction linear matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. AB - A simple mathematical technique for improving mass calibration accuracy of linear delayed extraction matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (DE MALDI-TOFMS) spectra is presented. The method involves fitting a parabola to a plot of Delta(m) vs. mass data where Delta(m) is the difference between the theoretical mass of calibrants and the mass obtained from a linear relationship between the square root of m/z and ion time of flight. The quadratic equation that describes the parabola is then used to correct the mass of unknowns by subtracting the deviation predicted by the quadratic equation from measured data. By subtracting the value of the parabola at each mass from the calibrated data, the accuracy of mass data points can be improved by factors of 10 or more. This method produces highly similar results whether or not initial ion velocity is accounted for in the calibration equation; consequently, there is no need to depend on that uncertain parameter when using the quadratic correction. This method can be used to correct the internally calibrated masses of protein digest peaks. The effect of nitrocellulose as a matrix additive is also briefly discussed, and it is shown that using nitrocellulose as an additive to the alpha-cyano-4-hydroxycinnamic acid (alphaCHCA) matrix does not significantly change initial ion velocity but does change the average position of ions relative to the sample electrode at the instant the extraction voltage is applied. PMID- 12112259 TI - Inductively coupled plasma mass spectrometric determination of molybdenum in urine. AB - A method was developed for the determination of molybdenum (Mo) in human urine by direct dilution of the sample in doubly distilled water with 1% HNO3 (v/v) and inductively coupled mass spectrometry (ICP-MS). In and Y were used as internal standards. Since (98)Mo provides a higher sensitivity, it was chosen as the reference isotope. The influence of different factors, such as sample dilution, HNO3 concentration and the stability of the analyte were evaluated. The detection limit (LOD) was assessed at 0.2 microg/L Mo, while the lower limit of quantification (LOQ) was 0.6 microg/L. Recoveries ranged between 97.2 and 100.7% from solutions containing from 10 to 50 microg/L Mo. Linear calibration curves were generated from 2.1 and 52.1 microg/L with coefficients of variation (CV ) ranging from 1.62 to 3.56%. In order to establish reference values (RV) for molybdenum, the procedure presented here was used to determine Mo in the urine of a population group living in Tuscany, Italy. PMID- 12112260 TI - On-line high-performance liquid chromatography/mass spectrometric characterization of native oligosaccharides from glycoproteins. AB - An on-line high-performance liquid chromatography/mass spectrometry (HPLC/MS) method is described for the rapid characterization of any type of oligosaccharide released from glycoproteins. The procedure can be applied without further manipulation to fractions collected from a high-performance anion-exchange chromatography-pulse amperometric detection (HPAEC-PAD) system commonly used for glycosylation mapping of glycoproteins, or to a pool of oligosaccharides directly released from glycoproteins. The system consists of a porous graphitized high performance chromatography column (Hypercarb) coupled to a quadrupole time-of flight (TOF) mass spectrometer. Oligosaccharides are eluted from the column with a gradient of ammonium acetate/acetonitrile and directly identified following in source fragmentation. Some applications of the method are presented, as well as information about the spectra and fragmentation behavior observed for N- and O linked oligosaccharides released from some recombinant glycoproteins. Low femtomole limits of detection are achieved using proper miniaturization. PMID- 12112261 TI - Use of a liquid chromatography/ion trap mass spectrometry/triple quadrupole mass spectrometry system for metabolite identification. PMID- 12112263 TI - Forensic identification of explosive oxidizers by electrospray ionization mass spectrometry. AB - The mass spectrometry of a group of inorganic oxidizers was studied using the electrospray ionization technique. It was found that a series of cluster ions were predominant in both positive- and negative-ion mode, allowing for the characterization of the investigated oxidizers. The identity of the recorded cluster ions was further confirmed by using some isotopically labeled compounds and tandem mass spectrometry with collision-induced dissociation. The use of electrospray ionization mass spectrometry for positive identification of major oxidizer components in explosive formulations was demonstrated by three samples of forensic interest. PMID- 12112264 TI - Identification of hemes and related cyclic tetrapyrroles by matrix-assisted laser desorption/ionization and liquid secondary ion mass spectrometry. AB - Mass spectrometry has proven to be a powerful technique applicable on trace amounts for the identification of known hemes and cyclic tetrapyrroles, and for providing critical information for the structure of new and novel versions. This report describes investigations of the practical limits of detection for such bioinorganic prosthetic groups, primarily by liquid secondary ion mass spectrometry (LSIMS) and matrix-assisted laser desorption/ionization time-of flight mass spectrometry (MALDI-TOFMS), including a survey of the utility of common matrices. The lower limit of detection under favorable conditions extends to low picomole amounts. Certain derivatization techniques, such as methyl esterification and chelation to zinc, both increase the sensitivity of analyses and provide spectroscopic signatures that enable heme/cyclic tetrapyrrole ions to be identified in the presence of contaminants. PMID- 12112265 TI - Analysis of protein ions in the range 3000-12000 Th under partial (no discharge) atmospheric pressure chemical ionization conditions using ion trap mass spectrometry. AB - A new approach, based on the use of atmospheric pressure chemical ionization ion trap mass spectrometry (APCI-ITMS), but without a corona discharge, was investigated for application to creating and monitoring protein ions. It must be emphasized that APCI is not usually used in protein analysis. In order to verify the applicability of the proposed method to the analysis of proteins, two standard proteins (horse cytochrome c and horse myoglobin) were analyzed. A mixture of the two proteins was also analyzed showing that this novel approach, based on the use of APCI, can be used in the analysis of protein mixtures. PMID- 12112266 TI - Considerations for design of a Fourier transform mass spectrometer in the 4.2 K cold bore of a superconducting magnet. AB - An external source Fourier transform mass spectrometer (FTMS) constructed inside the vertical cold bore of a superconducting magnet will have dramatic advantages in effective magnetic field, noise figures, and base pressure over current commercially available external source FTMS systems. There are substantial, but solvable, difficulties in the design, primarily with regard to control of the helium boiloff rate to an acceptable level, as well as relatively minor design challenges with heat sinks, contraction of metallic ion optic elements in the extreme temperature, and tandem mass spectrometry experiments. However, the ability to construct the FTMS inside the narrow bore tube of existing, commercially available vertical bore NMR magnets will allow access to the upper magnetic field limit currently used by 900 MHz (21 Tesla) - 1 GHz (23.3 Tesla) NMR experiments. The vacuum system, simply by being held inside the cold bore at 4.2 K, will cryopump itself dropping base pressures substantially, and heat sinking the input resistor of the preamplifier to this cryogenically cooled vacuum chamber will allow reduction of the input Johnson noise by a factor of 8.4 with associated 8.4-fold improvement in signal/noise, sensitivity, and dynamic range. The simultaneous improvement of three fundamental limiting factors in the FTMS (field strength, base pressure, and Johnson noise figure) will clearly outweigh the concomitant increased helium boiloff rate particularly if this rate can be dropped to the estimated <5 L/day range. The additional use of modern cryorefrigerators will further reduce helium boiloff to zero except during MS(n) experiments and system cooldown. PMID- 12112267 TI - Structural characterization of steroidal saponins by electrospray ionization and fast-atom bombardment tandem mass spectrometry. AB - The structural characterization of four steroidal saponin compounds involving two and three sugar groups, namely spirostanol saponins and furostanol saponins, were investigated by positive ion fast-atom bombardment (FAB), electrospray ionization mass spectrometry (ESI-MS) and tandem mass spectrometry (MS/MS) techniques. Important structural information was obtained from collision-induced dissociation (CID) and FAB-MS spectra with different liquid matrices. It was found that a characteristic fragmentation involving the loss of 144 Da arising from the cleavage of the E-ring was observed when there was no sugar chain at the C-26 position. When a glucoside group was substituted at the C-26 position, this C-26 sugar moiety was preferentially eliminated. All of these compounds produced a major product ion with a stable skeleton structure at m/z 255. The results of this paper can assist structural analysis of mixtures of steroidal saponins. PMID- 12112269 TI - Electron ionisation mass spectral analysis and fragmentation pathways for some thiophosphorylic p-carboxybenzene sulfonamides. AB - The work presents the 70 eV electron impact mass spectra of some thiophosphorylic p-carboxybenzene sulfonamides, and proposes rationalisation of their fragmentation pathways. Accurate mass measurements of the fragment ions, and metastable ion analyses performed in the MIKES mode, were used to elucidate the most abundant ion compositions and to elucidate the fragmentation patterns of these pharmacologically interesting compounds. PMID- 12112268 TI - Electron impact mass spectral fragmentation of chiral bisbenzoxazole, bisbenzothiazole and biscarbamide derivatives of 2,2-dimethyl-1,3-dioxolane. AB - The mass spectrometric fragmentation behaviour of five pairs of (R,R)- and (S,S) 4,5-bis(benzoxazol-2-yl)-2,2-dimethyl-1,3-dioxolane derivatives, one pair of (R,R)- and (S,S)-4,5-bis(benzothiazol-2-yl)-2,2-dimethyl-1,3-dioxolanes, and three pairs of (R,R)- and (S,S)-N,N'-bis(2-hydroxyaryl)-2,2-dimethyl-1,3 dioxolane-4,5-dicarbamides, all important compounds for asymmetric catalysis (P. Jiao et al., Tetrahedron Asymmetry 2001; 12: 3081), has been studied with the aid of mass-analyzed ion kinetic energy spectrometry and accurate mass measurements under electron impact ionization conditions. The spectral observations have been rationalized in terms of fragment ion structures and fragmentation mechanisms that will provide an aid to spectral interpretation for new compounds of this type. PMID- 12112270 TI - Determination of glutathione in spruce needles by liquid chromatography/tandem mass spectrometry. AB - For the determination of glutathione (GSH) and its oxidized form (GSSG) in spruce needles their electrospray mass and MS/MS spectra were recorded with an ion trap mass spectrometer (ITMS, LCQ, Finnigan) and a triple stage quadrupole mass spectrometer (TSQ, Quattro II, Micromass). A study of the stability of GSH in aqueous solutions shows the presence of dimeric and trimeric forms of GSH, as well as GSSG, GSH-sulfonate and GSH-sulfinic acid. The same components were also found in extracts of spruce needles. We developed an assay which is suitable for monitoring low concentrations of GSH and similar compounds in plant tissues, employing the sensitivity and specificity of LC/MS/MS. Preliminary results on the mass spectrometric determination of GSH in spruce needles are given. PMID- 12112271 TI - Tri-alpha-naphthylbenzene as a crystalline or glassy matrix for matrix-assisted laser desorption/ionization: a model system for the study of effects of dispersion of polymer samples at a molecular level. AB - Tri-alpha-naphthylbenzene (TalphaNB) can exist as either a crystalline or glassy solid at ambient temperatures, making it a unique matrix in matrix-assisted laser desorption/ionization (MALDI) spectroscopy. Electrosprayed TalphaNB is crystalline and has a melting point of 180 +/- 2 degrees C, as measured by differential scanning calorimetry (DSC). A glass of TalphaNB is obtained upon heating above the crystalline melting point with a glass transition temperature of 68 +/- 2 degrees C having no remaining crystallinity. MALDI samples containing mass fraction 1% polystyrene (PS) are run in both the crystalline and amorphous states. In the crystalline state, there is a strong spectrum typical of PS, but upon melting and quenching to the glassy state, the MALDI signal disappears. If the transparent, amorphous sample is treated with 1-butanol, it becomes white, and the MALDI signal returns. DSC shows that the 1-butanol treatment leads to the return of some of the crystallinity. Small angle neutron scattering (SANS) shows that the crystalline state has large aggregations of PS while the amorphous state has molecularly dispersed PS molecules. MALDI gives strong signals only when there are large aggregations of polymer molecules, with individually dispersed molecules producing no signal. PMID- 12112273 TI - Short capillary ion-pair high-performance liquid chromatography coupled to electrospray (tandem) mass spectrometry for the simultaneous analysis of nucleoside mono-, di- and triphosphates. AB - A liquid chromatography/mass spectrometry (LC/MS) method for the analysis of complex mixtures of nucleoside mono-, di- and triphosphates has been developed. A short capillary column (35mm x 0.3mm i.d.) was operated under ion-pair high performance liquid chromatography conditions and hyphenated to (negative) electrospray (tandem) mass spectrometry. As such, the separation of 12 nucleotides was performed by a binary gradient elution using CH(3)OH/H(2)O and N,N-dimethylhexylamine (N,N-DMHA) as ion-pairing agent. The influence of different N,N-DMHA concentrations on the chromatographic and mass spectrometric performance was evaluated to achieve optimal LC/MS conditions. In addition it was demonstrated that a controlled admission of ammonium dihydrogen phosphate (NH(4)H(2)PO(4)) improved both chromatographic performance and mass spectrometric detection. Because the system was hyphenated to an orthogonal designed electrospray interface (Z-spraytrade mark), long acquisition times were possible without loss of sensitivity. PMID- 12112272 TI - Matrix-assisted laser desorption/ionization mass spectrometry compatible beta elimination of O-linked oligosaccharides. AB - A new beta-elimination procedure has been introduced to cleave O-linked oligosaccharides from low- to sub-microgram amounts of glycoproteins prior to analysis by mass spectrometry. Borane-ammonia complex in aqueous ammonia is used as a cleaving solution alternative to the sodium borohydride/sodium hydroxide medium conventionally used in beta-elimination. The procedure results in minimum sample purification, leading to minimal sample loss and consequently an overall enhancement in sensitivity. It was applied successfully in the analysis of bovine fetuin and submaxillary mucin, as well as to a complex bile-salt-stimulated lipase glycoprotein isolated from human milk. PMID- 12112274 TI - Evaluation of low-pressure gas chromatography linked to ion-trap tandem mass spectrometry for the fast trace analysis of multiclass pesticide residues. AB - A rapid multiresidue method for the analysis of 72 pesticides has been developed using a single injection with low-pressure gas chromatography/tandem mass spectrometry (LP-GC/MS/MS). The LP-GC/MS/MS method used a short capillary column of 10 m x 0.53 mm i.d. x 0.25 microm film thickness coupled with a 0.6 m x 0.10 mm i.d. restriction at the inlet end. Optimal LP-GC conditions were determined which achieved the fastest separation in MS/MS detection mode. Also MS/MS conditions were optimized in order to increase sensitivity and selectivity. The analytical parameters of the LP-GC/MS/MS method were compared with those obtained by GC/MS/MS using a conventional capillary column (30 m x 0.25 mm i.d. x 0.25 microm film thickness). Better precision and sensitivity values were obtained with the LP-GC/MS/MS approach. The limits of detection (LOD) of the compounds ranged from 0.1 to 14.1 microg L(-1) for LP-GC/MS/MS, lower than those obtained for conventional GC/MS/MS that ranged from 0.1 to 17.5 microg L(-1). The peak widths obtained with the short column in LP-GC are similar to those obtained using conventional capillary GC columns, and the peaks can be successfully identified by MS/MS detection with the conventional scan speed of ion-trap instruments. In addition, the analysis time was significantly reduced with LP GC/MS/MS (32 min) versus GC/MS/MS (72 min), allowing the number of samples analyzed per day in a routine laboratory to be doubled. PMID- 12112275 TI - Liquid chromatography/tandem mass spectrometric quantification with metabolite screening as a strategy to enhance the early drug discovery process. AB - Throughput for early discovery drug metabolism studies can be increased with the concomitant acquisition of metabolite screening information and quantitative analysis using ultra-fast gradient chromatographic methods. Typical ultra-fast high-performance liquid chromatography (HPLC) parameters used during early discovery pharmacokinetic (PK) studies, for example, employ full-linear gradients over 1-2 min at very high flow rates (1.5-2 mL/min) on very short HPLC columns (2 x 20 mm). These conditions increase sample throughput by reducing analytical run time without sacrificing chromatographic integrity and may be used to analyze samples generated from a variety of in vitro and in vivo studies. This approach allows acquisition of more information about a lead candidate while maintaining rapid analytical turn-around time. Some examples of this approach are discussed in further detail. PMID- 12112276 TI - Variety identification of wheat using mass spectrometry with neural networks and the influence of mass spectra processing prior to neural network analysis. AB - The performance of matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry with neural networks in wheat variety classification is further evaluated.1 Two principal issues were studied: (a) the number of varieties that could be classified correctly; and (b) various means of pre-processing mass spectrometric data. The number of wheat varieties tested was increased from 10 to 30. The main pre-processing method investigated was based on Gaussian smoothing of the spectra, but other methods based on normalisation procedures and multiplicative scatter correction of data were also used. With the final method, it was possible to classify 30 wheat varieties with 87% correctly classified mass spectra and a correlation coefficient of 0.90. PMID- 12112277 TI - Determination of 4-hydroxynonenal in rat plasma by gas chromatography/mass spectrometry. PMID- 12112278 TI - Qualitative determination of trace quantities of nonyl phenyl polyethylene glycol ether in water based on solid-phase microextraction combined with surface assisted laser desorption/ionization mass spectrometry. PMID- 12112280 TI - Enantiomeric quantification of the bioactive peptide seryl-histidine methyl ester by electrospray ionization mass spectrometry and the kinetic method. PMID- 12112279 TI - A mixed liquid matrix for infrared matrix-assisted laser desorption/ionization of oligonucleotides. PMID- 12112281 TI - Simple two-point calibration of hybrid quadrupole time-of-flight instruments using a synthetic lipid standard. PMID- 12112282 TI - Biological activities of lavender essential oil. AB - Essential oils distilled from members of the genus Lavandula have been used both cosmetically and therapeutically for centuries with the most commonly used species being L. angustifolia, L. latifolia, L. stoechas and L. x intermedia. Although there is considerable anecdotal information about the biological activity of these oils much of this has not been substantiated by scientific or clinical evidence. Among the claims made for lavender oil are that is it antibacterial, antifungal, carminative (smooth muscle relaxing), sedative, antidepressive and effective for burns and insect bites. In this review we detail the current state of knowledge about the effect of lavender oils on psychological and physiological parameters and its use as an antimicrobial agent. Although the data are still inconclusive and often controversial, there does seem to be both scientific and clinical data that support the traditional uses of lavender. However, methodological and oil identification problems have severely hampered the evaluation of the therapeutic significance of much of the research on Lavandula spp. These issues need to be resolved before we have a true picture of the biological activities of lavender essential oil. PMID- 12112283 TI - Inhibitory effects of echinoisoflavanone and sophoraisoflavanone D in Sophora chrysophylla SEEM on copper ion-induced protein oxidative modification of mouse brain homogenate in vitro. AB - We present the results of an in vitro investigation of the inhibitory effects of echinoisoflavanone and sophoraisoflavanone D isolated from Sophora chrysophylla SEEM on copper-induced protein oxidative modification of mouse brain homogenate in vitro. They inhibited copper-induced protein oxidative modification. The order of inhibitory effects of these isoflavanones and mannitol as a hydroxyl radical scavenger was sophoraisoflavanone D > mannitol > echinoisoflavanone. The results suggest that these natural products may be of use in cases where oxidative stress is present. PMID- 12112284 TI - Enhanced conditioned inhibitory avoidance by a combined extract of Zingiber officinale and Ginkgo biloba. AB - Previous work has shown that intragastric administration of Zingicomb, a preparation consisting of Zingiber officinale and Ginkgo biloba extracts, has anxiolytic-like properties. The aim of the present study was to assess the effects of acute treatment with this preparation on inhibitory avoidance learning. The influence of pre-trial administered Zingicomb (ZC) on inhibitory avoidance conditioning was investigated in adult male Wistar rats, with a one trial step-through avoidance task. The animals were treated intragastrically with either vehicle, 0.5, 1, 10 or 100 mg/kg ZC 60 min prior to the acquisition trial. When tested 24 h after training, rats which had received 10 mg/kg ZC exhibited significantly longer step-through latencies than vehicle treated animals. This result, thus, demonstrates the beneficial effects of Zingicomb on conditioned inhibitory avoidance. Unlike conventional anxiolytic drugs, such as the benzodiazepines, which tend to have amnesic properties, this phytopharmacon is a potent anxiolytic agent which, additionally, can facilitate performance on a learning task, indicating promising clinical applications. PMID- 12112285 TI - Effects of Melilotus officinalis on acute inflammation. AB - Our study investigated the effects of Melilotus officinalis L. extract, containing 0.25% coumarin, on acute inflammation induced with oil of turpentine in male rabbits. The results were compared with those from a group treated with hydrocortisone sodium hemisuccinate and one injected with coumarin before inflammation was induced. The effects were evaluated by measuring serum citrulline, a test of in vitro phagocytosis, total leukocyte count and differential leukocyte count expressed as a percentage. M. officinalis had antiinflammatory effects because it reduced the activation of circulating phagocytes and lowered citrulline production. These properties were similar to those of hydrocortisone sodium hemisuccinate and coumarin. In the bone marrow acute phase response, M. officinalis had an inhibitory action that was lower than that of hydrocortisone sodium hemisuccinate and similar to coumarin. PMID- 12112286 TI - Cytotoxic flavonoids from the stem bark of Lonchocarpus aff. fluvialis. AB - Activity-guided fractionation of a chloroform-soluble extract of Lonchocarpus aff. fluvialis stem bark using a human epidermoid (KB) tumour cell line as a monitor afforded five rotenoids, one pterocarpan, one chalcone, three flavanones, one flavone and one triterpenoid. All of the compounds isolated proved to be of previously known structure. Among them, the rotenoids (-)-sumatrol and (+/-) villosinol, the dibenzoylmethane derivative (+)-3,4-methylenedioxy-2'-methoxy [2",3":4',3']-furanodibenzoylmethane, and the flavanones (-)-isoglabrachromene and (-)-candidone have been shown to exhibit significant cytotoxic activity against human cancer cells for the first time. This is the first report of the chemical constituents of this species, and the profile of compounds obtained was in accordance with the established chemosystematic patterns of species in the tribe Tephrosieae (Leguminosae, Papilionoideae). PMID- 12112287 TI - Psychopharmacological studies on Tragia involucrata root extract. AB - The methanol fraction of the root extract of Tragia involucrata was investigated for psychopharmacological activity in rodents. It produced an alteration of behavioural pattern and a reduction in spontaneous motility. A significant potentiation of pentobarbitone-induced sleep, a decrease of body temperature, suppression of the aggressive behaviour pattern and of the conditioned avoidance response (CAR) were observed with the fraction. These suggest that the methanol fraction of the T. involucrata root extract possesses significant central nervous system depressant action. PMID- 12112288 TI - Antioxidant activity of the methanol fraction of Pluchea indica root extract. AB - Studies were carried out to evaluate the influence of the methanol fraction of Pluchea indica Less root extract (PIRE), the dual inhibitors (BW 755C and phenidone) and vitamin on both in vivo and in vitro free radical-scavenging activities, CCl(4)-induced lipid peroxidation and the metabolism of arachidonic acid by lipoxygenase. PIRE produced significant antiinflammatory activity against glucose oxidase-induced paw oedema (in vivo), inhibited hydroxyl radical and superoxide generation, lysis of erythrocytes induced by hydrogen peroxide, CCl(4) induced lipid peroxidation and also dioxygenase activity of lipoxygenase (both in the presence and absence of hydrogen peroxide). Significantly higher free radical scavenging activity was observed with BW 755C and phenidone compared with PIRE. However, both BW 755C and phenidone stimulated hydroxyl radical generation compared with the observed inhibitory effects of PIRE and vitamin E. PMID- 12112289 TI - Activity of essential oils of three Micromeria species (Lamiaceae) against micromycetes and bacteria. AB - The chemical composition of essential oils from three Micromeria species: M. dalmatica Benth., M. albanica (Griceb. ex K. Mal) Silic and M. thymifolia (Scop.) Fritsch were investigated by GC and GC-MS and their antibacterial and antifungal activities against seven fungal and six bacterial species were evaluated. Biological assays showed strong fungitoxicity of oils from all three Micromeria spp., particularly M. albanica, against all fungi tested. Essential oils of these species also exerted antibacterial effect against Streptococcus aureus, Escherichia coli, Bacillus subtilis and Micrococcus luteus at low concentrations. Higher concentrations of essential oil of M. albanica and M. dalmatica were active against Gram-negative Pseudomonas aeruginosa, which could be due to the high content of piperitenone oxide. PMID- 12112290 TI - Evaluation of antilipid peroxidative action of propolis ethanol extract. AB - The antilipid peroxidative action of the ethanol extract of Brazilian propolis at a concentration of 47% (w/v) was evaluated by examining the inhibitory effect of the extract on the formation of hydroperoxide- and endoperoxide-type lipid peroxides during heating of authentic polyunsaturated fatty acids and on Fe(3+) ADP/ascorbic acid- and Fe(3+)-ADP/NADPH-dependent lipid peroxidation reactions in rat liver microsomes. Hydroperoxide-type lipid peroxides were measured by the haemoglobin-methylene blue method and endoperoxide-type lipid peroxides by the thiobarbituric acid (TBA), Fe(3+)-TBA and LPO-586 methods. Propolis ethanol extract inhibited dose-dependently the formation of hydroperoxide- and endoperoxide-type lipid peroxides during heating of linoleic acid, linolenic acid or arachidonic acid and the amount of the extract causing a half inhibition of these lipid peroxide formations ranged between 20 and 75 microg. Propolis ethanol extract inhibited dose-dependently both Fe(3+)-ADP/ascorbic acid- and Fe(3+) ADP/NADPH-dependent lipid peroxidation reactions in rat liver microsomes when lipid peroxides produced in both reactions were measured by the TBA method. The amount of propolis extract causing a half inhibition of the Fe(3+)-ADP/ascorbic acid-dependent lipid peroxidation was about 5 microg, while that of the extract causing a half inhibition of the Fe(3+)-ADP/NADPH-dependent lipid peroxidation was about 0.15 microg. These results indicate that the propolis ethanol extract exerts an antilipid peroxidative action at very low doses. PMID- 12112291 TI - An antiviral principle present in a purified fraction from Melia azedarach L. leaf aqueous extract restrains herpes simplex virus type 1 propagation. AB - Meliacine (MA), an antiviral principle isolated from leaves of Melia azedarach L., exhibits potent antiviral activity against herpes simplex virus type 1 (HSV 1) by inhibiting specific infected-cell polypeptides (ICPs) produced late in infection. Some of these are involved in DNA synthesis and in the assembly of nucleocapsids. The present report provides additional evidence to elucidate the mode of action of MA against HSV-1. Time-of-addition experiments confirmed that MA affects a late event in the multiplication cycle of HSV-1. We showed that MA diminished the synthesis of viral DNA and inhibited the spread of infectious viral particles when HSV-1 that expresses beta-galactosidase activity was used. In addition, the lack of a protein with an apparent MW of 55 KD was detected in MA-treated cell extracts. Ultrastructural analysis of infected cells showed that, in the case of MA treatment, a large number of unenveloped nucleocapsids accumulated in the cytoplasm and a minor proportion of mature virus was found in cytoplasmic vesicles.These findings suggest that MA exerts an antiviral action on both the synthesis of viral DNA and the maturation and egress of HSV-1 during the infection of Vero cells. PMID- 12112292 TI - Flow cytometric estimation on cytotoxic activity of leaf extracts from seashore plants in subtropical Japan: isolation, quantification and cytotoxic action of ( )-deoxypodophyllotoxin. AB - The cytotoxic activity of methanol extracts of leaves collected from 39 seashore plants in Iriomote Island, subtropical Japan was examined on human leukaemia cells (K562 cells) using a flow cytometer with two fluorescent probes, ethidium bromide and annexin V-FITC. Five extracts (10 microg/mL) from Hernandia nymphaeaefolia, Cerbera manghas, Pongamia pinnata, Morus australis var. glabra and Thespesia populnea greatly inhibited the growth of K562 cells. When the concentration was decreased to 1 microg/mL, only one extract from H. nymphaeaefolia still inhibited the cell growth. A cytotoxic compound was isolated from the leaves by bioassay-guided fractionation and was identified as (-) deoxypodophyllotoxin (DPT). The fresh leaves of H. nymphaeaefolia contained a remarkably high amount of DPT (0.21 +/- 0.07% of fresh leaf weight), being clarified by a quantitative HPLC analysis. DPT at 70-80 pM started to inhibit the growth of K562 cells in an all-or-none fashion and at 100 pM or more it produced complete inhibition in all cases. Therefore, the slope of the dose-response curve was very steep. DPT at 100 pM or more decreased the cell viability to 50%-60% and increased the number of cells undergoing apoptosis (annexin V-positive cells). The results indicate that DPT contributes to the cytotoxic action of the extract from the leaves of H. nymphaeaefolia on K562 cells. PMID- 12112293 TI - Effect of saikosaponin-A, a triterpenoid glycoside, isolated from Bupleurum falcatum on experimental allergic asthma. AB - The separation and isolation of an extract of Bupleurum falcatum were performed based on antiallergic activities in preliminary studies with higher plants. The final active compound was identified as saikosaponin-A (SSA), a triterpenoid glycoside. SSA at more than 1 mg/kg (i.v.) significantly inhibited the passive cutaneous anaphylaxis reaction in rats in a dose-dependent manner, attaining a maximum inhibition of approximately 60% with 10 mg/kg. SSA at 3 and 10 mg/kg also suppressed asthmatic bronchoconstriction in sensitized guinea-pigs. SSA possesses a weak inhibitory activity on histamine-induced tracheal contraction in guinea pigs and on histamine release induced by A-23187 in rat mast cells. These results indicate that SSA has an inhibitory activity against allergic asthma. This activity seems to derive from both antagonism of the histamine action and inhibition of allergic mediators. Additional mechanisms may also be involved. PMID- 12112294 TI - Peroxynitrite scavenging activity of herb extracts. AB - Peroxynitrite (ONOO(-)) is a cytotoxicant with strong oxidizing properties toward various cellular constituents, including sulphydryls, lipids, amino acids and nucleotides and can cause cell death, lipid peroxidation, carcinogenesis and aging. The aim of this study was to characterize ONOO(-) scavenging constituents from herbs. Twenty-eight herbs were screened for their ONOO(-) scavenging activities with the use of a fluorometric method. The potency of scavenging activity following the addition of authentic ONOO(-) was in the following order: witch hazel bark > rosemary > jasmine tea > sage > slippery elm > black walnut leaf > Queen Anne's lace > Linden flower. The extracts exhibited dose-dependent ONOO(-) scavenging activities. We found that witch hazel (Hamamelis virginiana L.) bark showed the strongest effect for scavenging ONOO(-) of the 28 herbs. Hamamelitannin, the major active component of witch hazel bark, was shown to have a strong ability to scavenge ONOO(-). It is suggested that hamamelitannin might be developed as an effective peroxynitrite scavenger for the prevention of ONOO( ) involved diseases. PMID- 12112295 TI - Inhibition of cholesterol biosynthesis in HepG2 cells by artichoke extracts is reinforced by glucosidase pretreatment. AB - High-dose aqueous extracts from artichoke leaves were found to inhibit cholesterol biosynthesis from (14)C-acetate rather moderately in HepG2 cells in contrast to primary cultured rat hepatocytes in which the inhibition was stronger. Preincubation of the extracts with several glycohydrolases revealed that pretreatment with beta-glucosidase considerably reinforced the inhibition. A significant reduction of acetate incorporation was found above extract concentrations of 0.01 mg/mL and at 0.2 mg/mL almost 60% inhibition was observed. Cytotoxic effects detected by the MTT-assay were restricted to higher concentrations of the extracts with and without beta-glucosidase pretreatment. Since cynaroside represents a major glucoside in artichoke extracts, both cynaroside and its aglycone luteolin were tested. It could be demonstrated that cynaroside is indeed one of the targets of beta-glucosidase and that the liberated luteolin is responsible for the inhibitory effect. Direct measurements of beta-glucosidase activity in rat hepatocytes and HepG2 cells revealed that endogenous enzyme activity in hepatocytes may be sufficient to convert cynaroside to its aglycone, while in HepG2 cells this may not be the case. These findings emphasize the importance of beta-glucosidase-dependent liberation of luteolin for the ability of artichoke extracts to inhibit hepatic cholesterol biosynthesis. PMID- 12112296 TI - Inhibitory effects of Keishi-bukuryo-gan on free radical induced lysis of rat red blood cells. AB - Keishi-bukuryo-gan (KBG) prevents the progression of atherosclerosis in cholesterol-fed rabbits by its antioxidative effect. The present study was to test our hypothesis that ingestion of KBG would protect red blood cell (RBC) membranes from free radical induced oxidation if polyphenolic antioxidants in KBG could be absorbed and circulated in the blood. When incubated with a RBC suspension, KBG and four of five herb medicines constituting KBG provided strong protection for RBC membranes against haemolysis induced by 2,2-azo-bis (2 amidinopropane) dihydrochloride (AAPH), an azo free radical initiator. The inhibitory effect was dose dependent at concentrations of 100-1000 microg/mL. Furthermore, the ingestion of 200 mg of KBG was associated with a significant decrease in susceptibility of RBC to haemolysis in rats. PMID- 12112297 TI - Preventative effects of the flowers of Inula britannica on autoimmune diabetes in C57BL/KsJ mice induced by multiple low doses of streptozotocin. AB - We have reported that an aqueous extract from the flowers of Inula britannica L. subsp. japonica Kitam. (IB) prevented immunologically induced experimental hepatitis in mice and suggested that the antihepatitic effect of IB is due to inhibition of IFN-gamma production. We then investigated the effects of IB on diabetes in mice induced by multiple low doses of streptozotocin (MLDSTZ), which is a mouse model for IFN-gamma-dependent autoimmune diabetes. C57BL/KsJ mice (male, 7 weeks) were provided with IB extract (500 mg/ kg/ day) in drinking water ad libitum, starting 7 days before the first STZ injection. Autoimmune diabetes was induced by MLDSTZ (40 mg/kg/day for 5 daily doses, i.p.). The IB treatment significantly suppressed the increase of blood glucose levels. Histological analysis of the pancreas showed that the degree of insulitis and destruction of beta-cells were reduced by IB treatment. The IFN-gamma production from stimulated splenic T lymphocytes was inhibited by the IB treatment. Moreover, the proportion of IFN-gamma-producing cells in the CD4(+) population, which was increased by MLDSTZ, was significantly decreased by the IB treatment. These results suggest that IB has a preventative effect on autoimmune diabetes by regulating cytokine production. PMID- 12112298 TI - Investigation on the hypoglycaemic effects of extracts of four Mexican medicinal plants in normal and alloxan-diabetic mice. AB - The hypoglycaemic activities of four water ethanol extracts (WEE) prepared from Bidens pilosa L., Salvia officinalis L., Psacalium peltatum H.B.K. (Cass) and Turnera diffusa Willd. were investigated in healthy and alloxan-diabetic mice. The WEE of S. officinalis significantly reduced the blood glucose of fasting normal mice 120 (15.7%) and 240 min (30.2%) after intraperitoneal administration (p < 0.05). The WEE of P. peltatum and B. pilosa also significantly diminished glycaemia in healthy mice at 240 min (19.6% and 13.8%, respectively). In mildly diabetic mice, the WEE of P. peltatum lowered the basal blood glucose level 120 (16%) and 240 min (54%) after intraperitoneal administration (p < 0.05 and p < 0.01, respectively). The WEE of B. pilosa and S. officinalis also significantly diminished the hyperglycaemia in mildly diabetic mice at 240 mins (32.6% and 22.7%, respectively). The administration of these three extracts to animals with severe hyperglycaemia did not cause a significant decrease. The WEE of T. diffusa did not show any hypoglycaemic activity. Thus, three of the WEE studied conserved the hypoglycaemic activity originally detected in the traditional preparations of the studied antidiabetic plants. It appears that these extracts require the presence of insulin to show hypoglycaemic activity. PMID- 12112299 TI - ent-Kaur-16-en-19-oic acid, a KB cells cytotoxic diterpenoid from Elaeoselinum foetidum. AB - Toxic and cytotoxic activities of the toxic plant Elaeoselinum foetidum (Apiaceae) were evaluated using the brine shrimp toxicity (BST) and KB cell cytotoxicity assays. The active chloroform extract was subjected to a bioactivity directed fractionation, monitored by the BST assay, that led to the isolation of the diterpenoid ent-kaur-16-en-19-oic acid. This compound was potent against the brine shrimp (LC(50) = 4.8 microg/mL) and KB cells (IC(50) = 1.6 microg/mL). PMID- 12112301 TI - Plumieride from Allamanda cathartica as an antidermatophytic agent. AB - Plumieride has been isolated as an active principle of the leaves of Allamanda cathartica. It showed strong fungitoxicity against some dermatophytes causing dermatomycosis to animals and human beings. It exhibited a noncytotoxic nature against a P(388) mouse leukaemia cell line. PMID- 12112300 TI - Globularia alypum L. extracts reduced histamine and serotonin contraction in vitro. AB - The present study analyses in in vitro models the pharmacological activity of methanol and dichloromethane extracts (1, 10 and 100 microg/mL) obtained from the leaves and stems of Globularia alypum L. Preincubation of the guinea-pig ileum and rat uterus with both extracts produced a dose dependent abolition of the contractile effects of histamine and serotonin, respectively. At the same doses, neither methanol nor dichloromethane extract reduced the contractile effects of acetylcholine on rat duodenum or noradrenaline on rat vas deferens. PMID- 12112302 TI - Anticonvulsant and sedative effects of Salvadora persica L. stem extracts. AB - The effect of Salvadora persica L. stem extracts on the potentiation of sodium pentobarbital activity and on generalized tonic-clonic seizure produced by pentylenetetrazol (PTZ) on the rat is reported. The extracts of Salvadora persica L. extended sleeping-time and decreased induction-time induced by sodium pentobarbital; in addition it showed protection against pentylenetetrazol-induced convulsion by increasing the latency period and diminishing the death rate. PMID- 12112303 TI - Human platelet aggregation inhibitors from thyme (Thymus vulgaris L.). AB - Two antiaggregant compounds, thymol (compound 1) and 3,4,3',4'-tetrahydroxy-5,5' diisopropyl-2,2'-dimethylbiphenyl (compound 2) were isolated from the leaves of thyme (Thymus vulgaris L.). The structures were determined by (1)H-, (13)C-NMR and mass spectra (MS) studies. These compounds inhibited platelet aggregation induced by collagen, ADP, arachidonic acid (AA) and thrombin except that compound 2 did not inhibit platelet aggregation induced by thrombin. PMID- 12112305 TI - Induction of caspase-mediated apoptosis and cell-cycle G1 arrest by selenium metabolite methylselenol. AB - Previous work based on mono-methyl selenium compounds that are putative precursors of methylselenol has strongly implicated this metabolite in the induction of caspase-mediated apoptosis of human prostate carcinoma and leukemia cells and G1 arrest in human vascular endothelial and cancer epithelial cells. To test the hypothesis that methylselenol itself is responsible for exerting these cellular effects, we examined the apoptotic action on DU145 human prostate cancer cells and the G1 arrest effect on the human umbilical vein endothelial cells (HUVECs) of methylselenol generated with seleno-L-methionine as a substrate for L methionine-alpha-deamino-gamma-mercaptomethane lyase (EC4.4.1.11, also known as methioninase). Exposure of DU145 cells to methylselenol so generated in the sub micromolar range led to caspase-mediated cleavage of poly(ADP-ribose) polymerase, nucleosomal DNA fragmentation, and morphologic apoptosis and resulted in a profile of biochemical effects similar to that of methylseleninic acid (MSeA) exposure as exemplified by the inhibition of phosphorylation of protein kinase AKT and extracellularly regulated kinases 1/2. In HUVEC, methylselenol exposure recapitulated the G1 arrest action of MSeA in mitogen-stimulated G1 progression during mid-G1 to late G1. This stage specificity was mimicked by inhibitors of phosphatidylinositol 3-kinase. The results support methylselenol as an active selenium metabolite for inducing caspase-mediated apoptosis and cell-cycle G1 arrest. This cell-free methylselenol-generation system is expected to have significant usefulness for studying the biochemical and molecular targeting mechanisms of this critical metabolite and may constitute the basis of a novel therapeutic approach for cancer, using seleno-L-methionine as a prodrug. PMID- 12112306 TI - Protective effect of vitamin E on ultraviolet B light-induced damage in keratinocytes. AB - Ultraviolet (UV) B radiation is the most common environmental factor in the pathogenesis of skin cancer. Exposure of human skin to UVB radiation leads to the depletion of cutaneous antioxidants, the activation of nuclear factor kappa B (NF kappaB), and programmed cell death (apoptosis). Although antioxidant supplementation has been shown to prevent UVB-induced photooxidative damage, its effect on components of cell signaling pathways leading to gene expression has not been clearly established. In the present study, the effect of the antioxidant vitamin, alpha-tocopherol (alpha-T), and its acetate analog, alpha-tocopherol acetate (alpha-TAc), on UVB-induced damage in primary and neoplastic mouse keratinocytes was investigated. The ability of both vitamins to modulate UVB induced apoptosis and activation of the transcription factor NF-kappaB were studied. Treatment of normal and neoplastic mouse epidermal keratinocytes (308 cells) with 30-60 mJ/cm(2) UVB markedly decreased viable cell number and was accompanied by DNA fragmentation. When both vitamins were applied to cells at times before and after UVB radiation, a significant increase in the percentage of viable cells and concomitant decrease in the number of apoptotic cells was noted, with vitamin pretreatment providing a better protection than posttreatment. Simultaneous posttreatment of irradiated cells with alpha-TAc abolished the cytotoxic effects of UVB and restored cell viability to control levels. In addition, simultaneous posttreatment of irradiated cells with alpha-T reduced the number of apoptotic cells by half, indicating a synergistic effect of two such treatments compared with any single one. Flow cytometry analysis indicated that vitamin treatment suppressed both an increase in pre-G0 cells and a decrease in cycling cells by UVB exposure. In addition, NF-kappaB activation was detected 2 h after UV exposure and was maintained for up to 8 h. Pretreatment with vitamins significantly inhibited NF-kappaB activation at 4 and 8 h. These results indicate that vitamin E and its acetate analog can modulate the cellular response to UVB partly through their action on NF-kappaB activation. Thus, these antioxidant vitamins are potential drugs for the protection from or the reduction of UVB associated epidermal damage. PMID- 12112307 TI - Overexpression of cyclin D2 is associated with increased in vivo invasiveness of human squamous carcinoma cells. AB - Overexpression of cyclin D2 was studied in 10 human squamous cell carcinoma lines, to establish whether this gene plays a role in tumor progression. We found that those cell lines that overexpressed cyclin D2 (CCND2) had the most invasive in vivo behavior. The invasive ability of the cell lines was determined by evaluating the penetration of carcinoma cells into the tracheal wall in an in vivo assay with de-epithelialized tracheas transplanted into the subcutaneous tissue of nude mice. From five cell lines that exhibited low invasive ability, we selected two that had very little CCND2 expression (SCC9 and SCC15), to evaluate whether CCND2 gene transfer would increase the invasive behavior. After confirming the successful transfer of CCND2 by Northern, Western, and kinase activity assays, we assessed the in vivo invasive behavior of the CCND2 transfected cells and their respective vector alone-transfected controls. The cell lines containing the transferred CCND2 gene had a significantly higher invasive ability than respective controls. This was accompanied by a moderate increase in gelatinase activity. In addition, the in vitro proliferative abilities, under normal culture conditions, of the parental CCND2-transfected and vector alone-transfected cells were found to be similar, as was the in vivo labeling index of Ki-67 in the tracheal transplants. These results indicated that the overexpression of CCND2 in squamous cell carcinoma lines modulates cell proliferation after induced quiescence and also has a powerful enhancing effect on in vivo aggressive growth behavior. PMID- 12112308 TI - Mutation rate and specificity analysis of tetranucleotide microsatellite DNA alleles in somatic human cells. AB - We have systematically varied microsatellite sequence composition to determine the effects of repeat unit size, G+C content, and DNA secondary structure on microsatellite stability in human cells. The microsatellites were inserted in frame within the 5' region of the herpes simplex virus thymidine kinase (HSV-tk) gene. The polypyrimidine/polypurine microsatellites displayed enhanced S1 nuclease sensitivity in vitro, consistent with the formation of non-B-form DNA structures. Microsatellite mutagenesis studies were performed with a shuttle vector system in which inactivating HSV-tk mutations are measured after replication in a nontumorigenic cell line. A significant increase in the HSV-tk mutation frequency per cell generation was observed after insertion of [TTCC/AAGG]9, [TTTC/AAAG]9, or [TCTA/AGAT]9 sequences (P 50%). The results indicated an apparent switch from focal expression of matrilysin in UC-related low-grade dysplasia to widespread expression in high grade dysplasia and invasive cancer, mimicking the pattern of expression in sporadic colorectal cancer. Although the sample size is small and further investigation therefore is required, the results suggest the possible role of anti-matrix metalloproteinase therapy in reducing the risk of progression from LGD to cancer in patients with ulcerative colitis. Published 2002 Wiley-Liss, Inc. PMID- 12112312 TI - A major breakpoint cluster domain in murine radiation-induced acute myeloid leukemia. AB - Cytogenetic and molecular studies have provided evidence of the clustering of chromosome 2 deletion breakpoints in radiation-induced murine acute myeloid leukemia (AML). Moreover, clustering occurs in at least two fragile domains rich in telomere-like arrays. Here we describe a physical map of the distal breakpoint cluster and confirm the presence of inverted head-to-head telomeric sequence arrays. These potentially recombinogenic sequences were not, however, the direct focus for post-irradiation chromosome breakage in AML. Instead, the two arrays bordered a 2.5-kb sequence with properties expected of a nuclear matrix attachment region (MAR). The putative MAR co-localized in the fragile domain with genes important to the hemopoietic system (leukocyte tyrosine kinase, zinc finger protein 106, erythrocyte protein band 4.2, and beta(2)-microglobulin (beta2m)); the beta2m subdomain was a particular focus of breakage. On the basis of these and other data, we suggest that AML-associated chromosome 2 fragility in the mouse is a consequence of domain-specific fragility in genomic domains containing numerous genes critical to the hemopoietic system. Recorded with the permission of the controller of Her Majesty's Stationery Office. Published by Wiley-Liss, Inc. PMID- 12112313 TI - Protection against human papillomavirus type 16-E7 oncogene-induced tumorigenesis by in vivo expression of dominant-negative c-jun. AB - Expression of the human papillomavirus (HPV) type 16 E6 and E7 gene products is a risk factor for human cervical carcinogenesis as well as skin and oral carcinogenesis. Expression of the HPV-16 E7 gene in mouse skin induces hyperplasia and enhances tumor promotion. Expression of dominant-negative c-jun (TAM67) in the mouse skin protects mice from 7,12-dimethylbenz[a]anthracene (DMBA)/12-O-tetradecanoylphorbol-13-acetate (TPA)-induced papillomagenesis without blocking mitogen-induced hyperproliferation. To determine the role of activator protein-1 (AP-1) in HPV-induced cancer, we crossed HPV-16 E7 mice with TAM67 mice and analyzed the effects of DMBA/TPA on tumor promotion. We showed that expression of TAM67 protected mice from HPV-16 E7-enhanced tumorigenesis, suggesting AP-1 as a target for prevention of HPV-induced cancer. PMID- 12112314 TI - Comparative analysis of loss of heterozygosity of specific chromosome 3, 13, 17, and X loci and TP53 mutations in human epithelial ovarian cancer. AB - We previously reported the identification of three minimal regions of deletion on the short arm of chromosome 3 (3p) in epithelial ovarian tumor specimens, suggesting that the inactivation of tumor-suppressor genes in these regions may be important in terms of ovarian tumorigenesis. Another previous study of ovarian cancer observed that allele loss of chromosome 179 was frequently found in ovarian tumors that also showed loss of heterozygosity (LOH) of chromosomes 3p, 13q, 17p, and Xp. In an independent study, we also reported a high frequency of LOH for selected chromosome 17 loci in high-grade and late-stage ovarian tumors. We have extended our LOH analysis of chromosome 3p to include 102 ovarian tumor specimens (29 and 73 samples were previously examined for LOH of chromosome 3p and 17 markers, respectively), using additional polymorphic markers, to assess the coordinate LOH of loci representing the three chromosome 3p minimal regions of deletions [von Hippel-Lindau syndrome (VHL), thyroid hormone receptor beta, and fragile histidine triad (FHIT)] and LOH of other important loci [tumor protein 53 (TP53), breast cancer 1 early onset (BRCA1), breast cancer 2 early onset, retinoblastoma 1, ornithine carbamoyltransferase, and androgen receptor] or somatic mutations in TP53. There was a significant association between LOH of any chromosome 3p marker and LOH of any chromosome 17 marker (P = 0.026). The frequency of LOH at the TP53 locus was higher in the group of samples that displayed LOH of a 3p marker (P = 0.019), as was the frequency of LOH at the BRCA1 locus (P = 0.014). LOH of chromosome 3p was noted in four specimens that did not display LOH of either the BRCA1 or the TP53 locus, indicating that LOH of these loci need not precede LOH of the chromosome 3p loci. We found a significant association between LOH of the VHL (3p25) locus and LOH of any chromosome 17 marker (P = 0.005), suggesting that there may be an important relationship, in the tumorigenesis of epithelial ovarian cancer, between a gene at 3p25 and a gene located on chromosome 17. Our results indicate that inactivation of p53 by somatic mutation is unlikely to be a prerequisite to chromosome 3p LOH, because we found no significant association between mutations in TP53 and LOH of the three chromosome 3p loci. The frequency of LOH at the FHIT locus at 3p14 increased significantly with advancing age at diagnosis (P = 0.018), as did the frequency of somatic TP53 mutations (P = 0.008). PMID- 12112315 TI - Expression of prostate-specific antigen is transcriptionally regulated by genistein in prostate cancer cells. AB - Prostate cancer is the second-leading cause of cancer-related deaths in men in the United States. Unfortunately, there is no effective therapy when prostate cancer becomes metastatic and refractory to conventional treatments. For this reason, the identification and exploration of new agents that reduce prostate cancer cell growth are of paramount importance. High consumption of plant-derived phytoestrogens is inversely associated with the incidence and mortality rate of prostate cancer. Previous studies, including our own, have shown that the phytoestrogen genistein inhibits prostate cancer cell growth in vitro and in vivo and decreases secreted and intracellular levels of the androgen-regulated protein prostate-specific antigen (PSA), but the role of genistein as an agonist/antagonist for hormone receptors remains unclear. To elucidate the mechanism by which genistein modulates PSA protein expression in prostate cancer cells, we investigated the effects of genistein on androgen-mediated and estrogen mediated transcriptional regulation of PSA, androgen receptor (AR) mRNA and protein expression, and the ability of nuclear proteins to bind to androgen response elements (AREs) in LNCaP cells. We showed that genistein decreased the transcriptional activation of PSA by both androgen-dependent and androgen independent methods in LNCaP cells. The reduction of androgen-mediated transcriptional activation of PSA was correlated with decreased AR protein and mRNA levels and decreased binding to AREs. In contrast, genistein had differential effects on 17beta-estradiol-mediated PSA expressions. Low concentrations of genistein enhanced 17beta-estradiol-mediated PSA expressions, whereas high concentrations of genistein inhibited estrogen-mediated PSA expression in LNCaP cells. Genistein did not inhibit AR protein expression in the presence of 17beta-estradiol. These results suggest that ligand-dependent differences in the ability to activate PSA expression may contribute to the agonistic/antagonistic responses observed with genistein in prostate cancer cells. PMID- 12112316 TI - Nordihydroguaiaretic acid-mediated inhibition of ultraviolet B-induced activator protein-1 activation in human keratinocytes. AB - Nordihydroguaiaretic acid (NDGA) is a polyphenolic compound from the Larrea tridentata bush that has been identified as a chemopreventive drug in animal studies. Topically applied NDGA has been shown to prevent phorbol ester promotion of tumors in mouse skin, suggesting that NDGA may be a candidate drug for the chemoprevention of skin cancer. Ultraviolet (UV) B irradiation from sunlight exposure is the major cause of human skin cancer. UVB irradiation causes epigenetic alterations in target keratinocytes, such as the upregulation of signal transduction pathways that induce the expression of transcription factors. Specifically, UVB induces activator protein-1 (AP-1), a transcription factor complex that alters normal cellular gene expression. A component of the UVB induced AP-1 complex, c-fos, also was identified as a mediator of the signaling pathway that leads to AP-1 activation. Thus, NDGA was investigated as a potential inhibitory agent for UVB-induced signaling pathways in the human keratinocyte cell line HaCaT. NDGA significantly inhibited UVB-induced c-fos and AP-1 transactivation. In addition, NDGA was found to inhibit activity of phosphatidylinositol 3-kinase (PI 3-kinase), a UVB-inducible enzyme that contributes to the induced expression of c-fos and AP-1. Therefore, NDGA prevents UVB-induced c-fos expression and AP-1 transactivation by inhibiting the PI 3 kinase signaling pathway. Effective skin chemoprevention strategies may incorporate NDGA to inhibit components of the UVB-induced cell signaling pathways that increase AP-1 activity. PMID- 12112317 TI - Transgene stability and features of rasH2 mice as an animal model for short-term carcinogenicity testing. AB - The transgenic mouse rasH2 line, in which the mouse carries the human c-Ha-ras gene under the control of its own enhancer and promoter, has been proposed as one of the alternative short-term models for carcinogenicity testing. To apply this purpose, we have produced a genetically homogeneous population as C57BL/6JJic TgN(RASH2) (Tg-rasH2) by continuous backcrossing. In this study, we examined the transgene stability between different generations and the detailed transgene architecture of the integrated human c-Ha-ras gene. Fluorescence in situ hybridization analysis showed that the integrated human c-Ha-ras gene was stably located on chromosome 15E3 in Tg-rasH2 mice at generation number (N) 15 and 20. Southern and Northern blot analysis did not show any differences in the hybridized band pattern in each generation. Southern blot analyses showed that the Tg-rasH2 mouse contained three copies of the human c-Ha-ras gene arrayed in a head-to-tail configuration. We also determined the nucleotide sequence of the transgene in the Tg-rasH2 mouse at N20 and confirmed that the sequence of the coding region was perfectly matched with human c-Ha-ras cDNA. Cloning and sequencing of genome/transgene junctions revealed that integration of the microinjected human c-Ha-ras gene into mouse host genome resulted in a 1820-bp deletion in the rasH2 line. The deleted sequence did not have any sequence homologies with known functional genes. We assumed that either the deletion or the transgene insertion, or both, would not cause insertional mutation. In short term carcinogenicity testing with a genetically engineered mouse model, confirmation of the transgene or modified gene stability at each generation is one of the important factors that affect the sensitivity to carcinogenic compounds in the same way as the genetic background, age and route of administration. PMID- 12112318 TI - APC-dependent regulation of ornithine decarboxylase in human colon tumor cells. AB - Mutation/deletion of the adenomatous polyposis coli (APC) tumor suppressor gene in germline cells of rodents and humans is associated with increased intestinal activity of ornithine decarboxylase (ODC), the first enzyme in polyamine synthesis, and intestinal neoplasia. To study the role of APC in signaling ODC expression, we used the human colon tumor cell line HT29 (wtAPC-/-), which has been stably transfected with a zinc-inducible wild-type APC gene. The addition of ZnCl(2) to HT29-APC cells increased wild-type APC protein and Mad1 RNA and protein and decreased levels of c-myc and ODC RNA and protein, relative to these parameters in HT29 cells transfected with the same plasmid containing the beta galactosidase gene in place of APC. Upon induction of APC expression, ODC promoter activity and RNA levels were suppressed. When the e-box domain in the 5' flanking region of the ODC gene was mutated, ODC promoter activity was unaffected by wild-type APC expression. Antisense, but not missense, c-myc oligonucleotides decreased ODC activity in HT29 cells expressing mutant APC. These results demonstrated that wild-type APC suppressed c-myc and activated Mad1 expression in HT29 colon-derived cells. These proteins, in turn, regulated the transcription of target genes, including ODC. The data presented indicate that ODC is a modifier of APC-dependent signaling in intestinal cells and tissues. PMID- 12112319 TI - Alterations of the Fhit gene in hepatocellular carcinomas induced by N nitrosodiethylamine in rats. AB - The present study was conducted to assess whether Fhit gene alterations are a feature of hepatocellular carcinomas (HCCs) induced by N-nitrosodiethylamine (DEN) in male Fischer 344 rats. Animals, 6 wk old, received a single intraperitoneal injection of DEN at a dose of 10 mg/kg body weight, followed by combined treatment with partial hepatectomy and colchicine to induce cell-cycle disturbance and a selection procedure, consisting of 2-acetylaminofluorene and carbon tetrachloride. Fourteen HCCs were obtained 42 wk after the beginning of the experiment; total RNA was extracted for the assessment of aberrant transcription of the Fhit gene by reverse transcriptase-polymerase chain reaction analysis. Aberrant transcripts were detected in nine of the 14 HCCs (64.3%). Sequence analysis showed that these resulted from the absence of nt -9 to 279, nt -9 to 348, nt -98 to 279, nt -26 to 365, or nt -98 to 348. Western blot analysis demonstrated reduced expression of Fhit protein in six of 10 HCCs (60.0%), with a perfect correlation with Fhit gene alterations. These results indicated that changes in the Fhit gene occur frequently and may thus play some role in the development of HCCs induced by DEN in rats. PMID- 12112320 TI - Inhibition of proliferation and expression of T1 and cyclin D1 genes by thyroid hormone in mammary epithelial cells. AB - The relationship between thyroid hormone (triiodothyronine, T(3)) and breast cancer is unclear. We studied the effect of the c-erbA/TR alpha proto-oncogene encoding a functional T(3) receptor (TR alpha 1), of its ligand T(3), and of its retroviral, mutated counterpart, the v-erbA oncogene, on the proliferation capacity of nontumorigenic mammary epithelial cells (EpH4). We found that EpH4 cells expressing ectopically TR (EpH4 + TR alpha 1) or v-erbA (EpH4 + v-erbA) proliferated faster than parental EpH4 cells that contained low levels of endogenous TR. T(3) inhibited DNA synthesis and proliferation in EpH4 + TR alpha 1 cells but not EpH4 or EpH4 + v-erbA cells. The study of cell-cycle genes showed that T(3) decreased cyclin D1 RNA and protein levels in EpH4 + TR alpha 1 cells. In addition, T(3) downregulated the expression of T1, a gene that is overexpressed in human breast adenocarcinomas and is induced by mitogens, serum, and several oncogenes and cytokines. Inhibition of the T1 gene by T(3) required both de novo mRNA and protein synthesis. Furthermore, T(3) abolished the induction of T1 by the tumor promoter 12-O-tetradecanoylphorbol-13-acetate and inhibited the activity of an activation protein 1-dependent promoter (-73-Col CAT) in EpH4 + TR alpha 1 cells, suggesting that interference with activation protein 1 transcription factor plays a part in the inhibition of the T1 gene. Our results showed that T(3) reduced the proliferation of mammary epithelial cells and inhibited the expression of cyclin D1 and T1 genes. PMID- 12112321 TI - Loss of heterozygosity, microsatellite instability, and mismatch repair protein alterations in the radial growth phase of cutaneous malignant melanomas. AB - Little is known about genomic alterations during development of the radial growth phase (RGP) of cutaneous malignant melanomas (CMMs). In this investigation polymerase chain reaction-based microsatellite assays were applied to analyze 13 RGP-CMMs with 18 microsatellite markers at six chromosomal regions: 1p, 3p, 4q, 6q, 9p, and 10q. Loss of heterozygosity (LOH) was found in eight cases (62%), at 9p22, 1p36, and 10q11, suggesting the presence of tumor-suppressor genes at these regions. LOH was encountered frequently at the interferon-alpha (31%) and D10S249 loci (15%). Low-level microsatellite instability (MSI) (11-16% of investigated loci unstable) was noted in three cases (23%). Two MSI banding patterns were seen: band shift and the presence of additional bands. To investigate the underlying mechanisms of the low-level MSI pattern, we analyzed the lesions for expression of mismatch repair (MMR) proteins with immunoperoxidase methods and mouse monoclonal antibodies. The average percentages of positively stained cells for human MutL homolog 1 (hMLH1), human MutS homolog 2 (hMSH2), and human MutS homolog 6 (hMSH6) in RGP-CMM (75.6 +/- 3.4%, 67.20 +/- 7.71%, and 76.6 +/- 2.1%, respectively) were reduced compared with benign nevi. No statistically significant differences in MMR protein expression were found between microsatellite-stable and low-level MSI lesions (P = 0.173, P = 0.458, and P = 0.385 for hMLH1, hMSH2, and hMSH6, respectively). There was a direct correlation between values for percentages of positively stained cells for hMSH2 and hMSH6 (r = +0.9, P = 0.03), suggesting that common mechanisms regulate their expression. In conclusion, LOH, MSI, and reduced MMR protein expression appear to be present in at least some RGP-CMMs and may play a role in their pathogenesis. Further studies are necessary to support these finding and to determine their diagnostic and prognostic significance. PMID- 12112322 TI - Removal of Cdk inhibitors through both sequestration and downregulation in zearalenone-treated MCF-7 breast cancer cells. AB - Treatment of MCF 7 cells with the fungal estrogen zearalenone induced cyclin E associated kinase activity transiently within 9-12 h; total cyclin-dependent kinase (Cdk) 2 activity was elevated for 24 h and beyond. This increased cyclin E/Cdk2 activity was associated with sequestration of the Cdk inhibitor p27 Cdk inhibitor 1B (p27(KIP1)) by newly formed cyclin D1/Cdk4 complexes and with downregulation of p27(KIP1) expression. The activation of cyclin A/Cdk2 activity corresponded with virtual elimination of p27(KIP1). The activity of cyclin E/Cdk2 complexes from zearalenone-treated lysates was inhibited in vitro by recombinant p27(KIP1), and this inhibition was relieved by the addition of recombinant cyclin D1/Cdk4 complexes. Thus, sequestration of p27(KIP1) by cyclin D1/Cdk4 resulted in activation of Cdk2 in vitro. Cdk inhibitory activity in lysates of zearalenone treated cells was depleted by anti-p27(KIP1) and anti-Cdc2 interacting protein (p21(CIP1)) antibodies. Overexpression of the Cdk4/6-specific Cdk inhibitor of Cdk4 p16(INK4A) was associated with increased association of p27(KIP1) with Cdk2, concomitant with disruption of D cyclin/Cdk4 complexes. The proteasome inhibitor 2-leu-leu-leu-H aldehyde (MG-132) was relatively ineffective in inhibiting the initial, sequestration-dependent activation of cyclin E/Cdk2 yet was as effective as p16(INK4A) in inhibiting activation of cyclin A/Cdk2 later in G(1). Downregulation of p27(KIP1) proceeded in p16(INK4A)-expressing cells after zearalenone treatment, and G(1) arrest afforded by p16(INK4A) expression was reversible upon prolonged treatment with zearalenone. Zearalenone treatment of MCF-7 cells elicited expression of F-box protein S phase kinase-associated protein 2 (p45(SKP2)), a substrate-specific component of the ubiquitin-ligase complex that targets p27(KIP1) for degradation in the proteasome. These studies suggest that both sequestration of Cdk inhibitors by cyclin D1/Cdk4 complexes and downregulation of p27(KIP1) play major roles in the induction of Cdk2 activity and S phase entry elicited by estrogens in MCF-7 cells. PMID- 12112324 TI - Emerging methods for the rapid determination of enantiomeric excess. AB - Methods for enantiomeric excess determination using a variety of spectroscopic techniques are summarized. Particular attention is paid to techniques that have promise for application to problems of combinatorial catalyst discovery but have not yet been so employed. PMID- 12112325 TI - Preferential enrichment: full crystallographic analysis of the unusual phenomenon in the mixed crystals' version. AB - Full characterization of the crystal structures of the racemate, nonracemate (20% ee), and pure enantiomer of [2-[4-(3-ethoxy-2 hydroxypropoxy)phenylcarbamoyl]ethyl]trimethylammonium p-bromobenzenesulfonate (NBMe3), which has successfully shown an unusual enantiomeric resolution phenomenon, "preferential enrichment," was achieved by means of X-ray crystallographic analysis and construction of the binary melting point phase diagram. The crystalline nature of the racemate is not a racemic compound but a fairly ordered mixed crystal composed of the two enantiomers. The crystal structure of the nonracemate (20% ee) is virtually identical with that of the racemate and similar to that of the pure enantiomer. The binary melting point phase diagram of NBMe3 is consistent with the nature of a mixed crystal composed of the two enantiomers. PMID- 12112326 TI - Asymmetric autocatalysis and its application to chiral discrimination. AB - Chiral pyrimidyl, quinolyl, and pyridyl alkanols act as asymmetric autocatalysts with significant amplification of enantiomeric excess (ee) in the enantioselective addition of diisopropylzinc to pyrimidine-5-, quinoline-3-, and pyridine-3-carbaldehydes, respectively. 2-Alkynyl-5-pyrimidyl alkanol with as low as 0.6% ee automultiplies during the consecutive asymmetric autocatalysis with increasing ee to as high as >99.5%. Asymmetric autocatalysis is applied to chiral discrimination of organic compounds. In the presence of methyl mandelate or 2 butanol with very low ee's (0.05-0.1%) as chiral initiators, the reaction between pyrimidine-5-carbaldehyde and diisopropylzinc affords pyrimidyl alkanol with higher ee's with the correlated absolute configurations to those of the chiral initiators. Chirality of amino acids (such as leucine) and helicenes with very low ee's are also discriminated by asymmetric autocatalysis, affording pyrimidyl alkanol with very high ee's. Asymmetric autocatalysis also discriminates the chirality of primary alcohols-alpha-d, monosubstituted [2.2]paracyclophanes and octahedral cobalt complex with achiral ligands of which the chirality is due to the topology of coordination of the achiral ligand. Even the chirality of inorganic crystals such as quartz and sodium chlorate is discriminated by asymmetric autocatalysis of pyrimidyl alkanol. Thus, asymmetric autocatalysis provides a unique method for the discrimination of chiral compounds and crystals. PMID- 12112327 TI - Chirality induction in supramolecular aggregate: chiral recognition between armed cyclen-Na+ complexes having quadruplicated helical geometry. AB - A water-soluble self-aggregate of octadentate Na+ complex with chiral cholesterol armed cyclen provides unique microenvironments for chirality induction of guest cyclen-Na+ complex via supramolecular interaction. PMID- 12112328 TI - Enzyme immobilization utilizing a porous ceramics support for chiral synthesis. AB - A novel porous ceramics support, named "Toyonite," for the immobilization of enzymes was prepared from the minerals of kaolinite under acidic conditions. Modification of the porous surface of Toyonite with two different organic coating agents gave Toyonite 200-M (TN-M), and Toyonite 200-A (TN-A), possessing methacryloyloxy and amino groups on the respective surfaces. Compared with other solid supports, TN-M and TN-A supports exhibited high selectivity for lipase PS (Pseudomonas cepacia, Amano) and glucoamylase (Gluczyme AF 6, Amano) proteins, respectively. The activities of both the transesterification of rac-1 with TN-M PS lipase and the hydrolysis of starch with TN-A glucoamylase were greater than those of similar reactions with these two enzymes immobilized on other solid supports. Further, TN-M PS lipase showed higher reactivity toward synthetic substrates, including aromatic and aliphatic secondary alcohols, than the free enzyme powder. PMID- 12112329 TI - Survey of factors determining the circularly polarised luminescence of macrocyclic lanthanide complexes in solution. AB - The development of emissive lanthanide complexes as structural or reactive probes to signal changes in their local chiral or ionic environment has been inhibited by the lack of understanding of correlating structural and electronic spectral information. The definition of relatively rigid enantiopure macrocyclic lanthanide complexes, whose inter- and intramolecular exchange dynamics have been defined, offers scope for remedying this situation. Chiral axially symmetric lanthanide complexes in solution give rise to large emission dissymmetry values (g(em)) in CPL spectra. The sign and magnitude of g(em) are determined by the degree of twist about the principal axis, which is predicted to be a maximum at +/-22.5 degrees, and by the site symmetry and local ligand field. In particular, the polarisability of the ligand donor atoms, especially for any axial donor, is very important. Examples of each case are discussed for structurally related cationic Eu(III) complexes. PMID- 12112330 TI - Enantioselective photoreaction of achiral tropolone ether in inclusion crystals with optically active host compounds. AB - Achiral 4-isopropyltropolone methyl ether (3) included in an optically active host compound (-)-1 yielded optically active photocyclization products (-)-5 and (-)-6 in 96 and 90% ee, respectively, upon photoirradiation in the solid state. PMID- 12112331 TI - Mechanism of asymmetric hydrogenation by rhodium complexes with unsymmetrical P chirogenic bisphosphine ligands. AB - Using a series of the rhodium complexes with (1S,2S)-1-(R(1))methylphosphino-2 (R(2))(R(3))phosphinoethane (R(1), R(2) and R(3) = 1-adamantyl, t-butyl, cyclohexyl, cyclopentyl, methyl; abbreviated as unsymmetrical BisP*), very high enantioselectivities were observed when the di- or tri- substituted and tetra substituted dehydro-alpha-amino acid derivatives were used as the substrates. The main factor to give high enantioselectivity is the repulsive interaction between the functional groups of the substrate and the bulky substituents of the unsymmetrical BisP*. Since the unsymmetrical BisP* has two independent chiral phosphorous atoms in the vicinity of the active site, the higher enantioselectivity than those by the C2 symmetric BisP* complexes can be obtained. Moreover, the fine-tuning to obtain extremely high enantioselectivity may be possible by changing the combination of the substituents on the two phosphorous atoms of the unsymmetrical BisP*. PMID- 12112332 TI - Asymmetric total synthesis of highly symmetric squalene-derived cytotoxic polyethers. AB - The efficient total synthesis of the cytotoxic meso polyether teurilene (1), rarely occurring in nature, was achieved through the effective combination of the concept of two-directional synthesis and rhenium(VII) chemistry. In the key rhenium(VII)-promoted syn oxidative cyclization reaction of the two-directional substrate 16, trans-syn diastereoselectivity (steric control) has been observed in contrast to our early observation of cis-syn diastereoselectivities (chelation control) for bishomoallylic tertiary alcohols 24-27 possessing the neighboring tetrahydrofuran (THF) ring. This synthesis in only 10 steps from commercially available methyl tiglate is significantly shorter than the previous one requiring 25 steps. The total syntheses of four possible C(s) symmetric (3 and 47-49) and one C2 symmetric (4) compounds for biogenetically squalene-derived pentaTHF polyether, glabrescol, was also achieved in relatively few steps by taking advantage of its intrinsic symmetry and one- and two-directional modes of double THF ring formations with vanadium catalyst, tert-butyl hydroperoxide (TBHP), and trifluoroacetic acid (TFA). Thus, the meso structure 3 originally proposed by Jacobs et al. has been revised to the optically pure C2 symmetric structure 4 through its enantioselective total synthesis. PMID- 12112333 TI - Synthesis and structure of poly(binaphthyl salen manganese complex) and its application to asymmetric epoxidation. AB - Poly(binaphthyl salen manganese complex)es 3-Mn were synthesized from a 3,3' diformylbinaphthol derivative, alpha,omega-diamines, and Mn(OAc)2. Their helical structures were well-supported by their IR, UV, and CD spectra. The catalysis of 3-Mn in an asymmetric epoxidation was investigated. PMID- 12112334 TI - Role of chirality and optical purity in nucleic acid recognition by PNA and PNA analogs. AB - Peptide nucleic acids are DNA mimics able to form duplexes with complementary DNA or RNA strands of remarkable affinity and selectivity. Oligopyrimidine PNA can displace one strand of dsDNA by forming PNA(2):DNA triplexes of very high stability. Many PNA analogs have been described in recent years, in particular, chiral PNA analogs. In the present article the results obtained recently using PNA derived from N-aminoethylamino acids 7 are illustrated. In particular, the dependence of optical purity on synthetic methodologies and a rationale for the observed effects of chirality on DNA binding ability is proposed. Chirality as a tool for improving sequence selectivity is also described. PNA analogs derived from D- or L-ornithine 8 were also found to be subjected to epimerization during solid phase synthesis. Modification of the coupling conditions or the use of a submonomeric strategy greatly reduced epimerization. The optically pure oligothymine PNAs 8 were found to bind to RNA by forming triplexes of unusual CD spectra. The melting curves of these adducts presented two transitions, suggesting a conformational change followed by melting at high temperature. PMID- 12112335 TI - Stereoselective protein binding of alprenolol in the renal diseased state. AB - The investigation was undertaken to study the stereoselective protein binding of alprenolol in renal disease patient sera, compared to that in the sera of healthy volunteers. The in vitro stereoselective protein binding of beta-blockers was determined in undiluted serum and in isolated alpha(1)-acid glycoprotein (AGP) solutions by ultrafiltration. The stereoselctive serum protein binding of alprenolol, a beta-adrenergic blocking agent, in healthy volunteers was significantly altered in renal disease patients. We investigated the effects of AGP concentration and endogenous substances, including uremic toxins, on the stereoselective protein binding of alprenolol in renal disease patients. A good correlation between the unbound (R)/(S) ratio (F(R)/F(S) ratio), an apparent index of stereoselectivity in alprenolol serum binding and AGP concentration in serum, was found. However, stereoselective protein binding was not influenced by endogenous substances. This result can be explained by the difference in binding affinities of (R) and (S)-isomers of alprenolol to AGP. We conclude that the stereoselective protein binding of alprenolol in healthy volunteers and renal disease patients varies as a result of changes in AGP concentration. Accordingly, these findings might be useful in alprenolol therapy in renal disease patients. PMID- 12112336 TI - Molecular recognition in solid-state crystallization: colored chiral adduct formations of 1,1'-bi-2-naphthol derivatives and benzoquinone with a third component. AB - Molecular recognition in solid-state crystallization involving derivatives of 1,1'-bi-2-naphthol and benzoquinone was studied. No adduct crystal was formed when crystals of biphenyl were further added as a third component to a grinding mixture of crystals of chiral 1,1'-bi-2-naphthol and benzoquinone, which by itself did not form an adduct. This contrasts with the case in which further addition of naphthalene crystals to the same mixture produced a new red crystal. Adduct formations using chiral 6,6'-dibromo-1,1'-bi-2-naphthol in place of 1,1' bi-2-naphthol were also studied. In this case, adducts were produced either with or without biphenyl as a third compound, but the colors of the adducts differed significantly in the two cases: red and bluish-black. The same three-component adduct crystals were produced from solid-grinding and solution crystallization and the structure was determined by X-ray diffractometry. Based on the crystal structures, theoretical calculations were carried out to compare the mechanism of colorations in the binary and the ternary complexes. PMID- 12112338 TI - Chiral recognition phenomena probed by steady-state luminescence measurements: Stern-Volmer analysis of chiral discriminatory behavior. AB - The chiral recognition phenomenon observed in enantioselective excited-state energy transfer processes currently requires the use of chiroptical spectroscopic techniques to probe the magnitude and sense of the discriminatory interactions. The use of chiroptical spectroscopic techniques limits the study of chiral recognition to those molecular species with strong absorption or emission dissymmetry factors. This study presents the theoretical and experimental methodology to determine the magnitude of chiral discriminatory interactions with unpolarized, steady-state luminescence measurements. Based on bimolecular luminescence quenching kinetics for a system containing chiral molecules, the Stern-Volmer equation is derived and contains a chiral discriminatory term for a system containing a chiral but racemic luminophore and an enantiomerically resolved quencher species. The utility of this methodology is confirmed by examining the enantioselective excited-state quenching between several Ln(dpa)(3)(3-) complexes (where Ln = Eu(3+), Tb(3+), or Dy(3+) and dpa = pyridine 2,6-dicarboxylate) acting as the energy donor and either racemic or enantiomerically resolved [Co(dach)(3)](3+) (where dach = trans-1R,2R (or 1S,2S) diaminocyclohexane) acting as the energy acceptor in an aqueous solution. The results of this study confirm the utility of unpolarized, steady-state luminescence measurements as a probe of chiral discriminatory behavior. PMID- 12112339 TI - Novel direct access to enantiomerization barriers from peak profiles in enantioselective dynamic chromatography: enantiomerization of dialkyl-1,3 allenedicarboxylates. AB - The axially chiral allenes dimethyl-1,3-allenedicarboxylate 1 and diethyl-1,3 allenedicarboxylate 2 show characteristic plateau formation during enantioselective GC separation on the chiral stationary liquid phase Chirasil beta-Dex. The elution profiles, obtained from temperature-dependent dynamic GC (DGC) experiments (1: 100-140 degrees C; 2: 110-150 degrees C) were evaluated with the recently derived approximation function (AF) k1(approx) = f(t(R)(A),t(R)(B),w(h)(A),h(plateau), N) to yield the enantiomerization rate constant directly k(1). These values were compared with those obtained by computer-aided simulation with ChromWin. The Eyring activation parameters of the experimental interconversion profiles were determined to be: DeltaG(#)(298.15 K) = 103.6 +/- 0.9 kJ mol(-1), DeltaH(#) = 44.7 +/- 0.4 kJ mol(-1), DeltaS(#) = -198 +/- 7 J K(1) mol(-1) for dimethyl-1,3-allenedicarboxylate 1, and DeltaG(#)(298.15 K) = 103.5 +/- 1.1 kJ mol(-1), DeltaH(#) = 44.7 +/- 0.5 kJ mol(-1), DeltaS(#) = 197 +/- 9 J K(-1) mol(-1) for diethyl-1,3-allenedicarboxylate 2. The approximation function (AF) presented here allows the fast determination of rate constants k(1) and activation barriers of enantiomerization DeltaG(#) from chromatographic parameters without extensive computer simulation. PMID- 12112340 TI - Synthesis and circular dichroism studies of N,N-bis(2-quinolylmethyl)amino acid Cu(II) complexes: determination of absolute configuration and enantiomeric excess by the exciton coupling method. AB - We report a method to determine the absolute configuration of alpha-amino acids by exciton coupled circular dichroism (ECCD). Naturally occurring amino acids were successfully derivatized with 2-bromomethylquinoline. Complexation of these conformationally flexible ligands with Cu(II) salts yielded defined propeller like structures. The direction of the twist (i.e., the relative orientation of the chromophores to each other) is governed by the asymmetric amino acid carbon center. The transition moments of the chromophores couple and yield a bisignate circular dichroism spectrum, the sign of which corresponds to the absolute configuration of the chiral center of the amino acid. Enantiomeric excess (e.e.) of amino acid derivatives is linearly related to the differential extinction coefficient Delta(epsilon) and can be assessed easily utilizing a standard curve. This efficient, sensitive technique requires low analyte concentrations, offers several advantages over established methods, and could be applied in medicinal, pharmaceutical, or chemical retail and manufacturing industry. PMID- 12112341 TI - A search for circular dichroism in the VUV photofragmentation mass spectra of 2 amino-1-butanol. AB - The dissociative photoionization of a single-enantiomer chiral molecule by circularly polarized synchrotron radiation was investigated, for the first time, in the gas phase. Photoion mass spectra were produced by the interaction of (+) (S)-, (-)-(R)- and rac-2-amino-l-butanol with circularly polarized light. Comparison of these spectra places an upper bound of approximately 2% on circular dichroism in the dissociative photoionization of 2-amino-l-butanol at 21 eV, which may have consequences for the theory that the origin of biological homochirality was predominantly enantioselective photofragmentation by circularly polarized light. We have also identified and elucidated many of the difficulties of performing gas phase CD measurements in crossed beam experiments. PMID- 12112343 TI - Gram-scale synthesis of (+)- and (-)-trans-10-amino-9-hydroxy-9,10 dihydrophenanthrene by enzymatic hydrolysis. AB - The enzymatic resolution of (+/-)-trans-10-azido-9-acetoxy-9,10 dihydrophenanthrene by Candida cylindracea lipase-catalyzed hydrolysis was achieved on the gram-scale with excellent yield and enantiomeric excess. The resulting (+)- and (-)-amino alcohols were derivatized to alpha-hydroxy-N benzenesulphonamides. PMID- 12112342 TI - Overall view of the use of chiral platinum(II) complexes as chiral derivatizing agents (CDAs) for the enantiodiscrimination of unsaturated compounds by 195Pt NMR. AB - The use of organometallic CDAs based on ethene-platinum(II) complexes, covalent cis- and trans-dichloro(Am)(ethylene)platinum(II), and ionic [PtCl(3)(C(2)H(4))]( )[AmH](+), containing primary and secondary amines, for the (195)Pt NMR enantiodiscrimination of chiral unsaturated compounds is reviewed. PMID- 12112345 TI - Enantioseparation properties of (1-->6)-alpha-D-glucopyranan and (1-->6)-alpha-D mannopyranan tris(phenylcarbamate)s as chiral stationary phases in HPLC. AB - Synthetic polysaccharides, (1-->6)-alpha-D-glucopyranan (3a) and (1-->6)-alpha-D mannopyranan (3b), were prepared by the cationic ring-opening polymerization of 1,6-anhydro-2,3,4-tri-O-allyl-beta-D-glucopyranose (1a) and 1,6-anhydro-2,3,4-O-6 allyl-beta-D-mannopyranose (1b), followed by the cleavage of the allyl ether linkage of 2,3,4-tri-O-allyl-(1-->6)-alpha-D-glucopyranan (2a) and 2,3,4-tri-O allyl-(1-->6)-alpha-D-mannopyranan (2b), respectively. 2,3,4-Tris-O-(3,5 dimethylphenylcarbamoyl)- and 2,3,4-tris-O-(3,5-dichlorophenylcarbamoyl)-(1-->6) alpha-D-glucopyranan (CSP-1 and CSP-2, respectively) and 2,3,4-tris-O-(3,5 dimethylphenylcarbamoyl)- and 2,3,4-tris-O-(3,5-dichlorophenylcarbamoyl)-(1-->6) alpha-D-mannopyranan (CSP-3 and CSP-4, respectively) were prepared by the reaction of 3 with the corresponding 3,5-disubstituted phenylisocyanates and the chiral recognition abilities of CSP-1-4 as chiral stationary phases (CSPs) in high-performance liquid chromatography (HPLC) were evaluated. Racemic compounds such as trans-cyclopropanedicarboxylic acid dianilide (9), 1,2,2,2 tetraphenylethanol (10), flavanone (11), Troger's base (12), benzoin (13), and cobalt(III) tris(acetylacetonate) (14) were efficiently resolved using CSP-1-4. For comparison among CSPs, the chiral recognition properties of the (1-->6)-alpha D-glucopyranan CSPs were different from that of the (1-->6)-alpha-D-mannopyranan CSPs, and CSP-4 exhibited the highest chiral recognition ability among the CSPs. The resolution factors of 12 and 14 were 0.42 and 0.56 for CSP-1, 0.32 and 2.16 for CSP-2, 1.80 and 0.84 for CSP-3, and 2.31 and 8.26 for CSP-4, respectively. PMID- 12112344 TI - Regio- and stereoselective hydroxylation of taxoids by filamentous fungi. AB - Paclitaxel (Taxol), is one of the most promising chemotherapeutic agents developed for cancer treatment in past two decades. Microorganisms such as filamentous fungi are known to perform regio- and stereoselective hydroxylation of taxoids. Herein, we describe highly regio- and stereoselective hydroxylation at the 1beta and 9alpha positions of the taxane skeleton. Such hydroxylation reactions proceed readily for the taxadienes as substrates rather than taxoids having an oxetane ring. The presence of different oxygen substituents on the taxane nucleus, such as 5-acetoxy, has a significant effect on the selectivity and yield of the hydroxylation catalyzed by the microbial oxidases. PMID- 12112346 TI - Liquid chromatographic resolution of N-acyl-alpha-amino acids as their anilide derivatives on a chiral stationary phase based on (S)-leucine. AB - A chiral stationary phase (CSP 1) derived from N-(3,5-dinitrobenzoyl)leucine N phenyl N-alkylamide was used for the liquid chromatographic resolution of anilide derivatives of N-acyl-alpha-amino acids and the chromatographic resolution results were compared with those from four other commercial CSPs. The chromatographic resolution results showed that CSP 1 was most effective among five CSPs used in this study. The chiral recognition mechanism exerted by CSP 1 for the resolution of anilide derivatives of N-acyl-alpha-amino acids is proposed to involve a face-to-face pi-pi interaction and two hydrogen bonding interactions between the CSP and the analytes from the chromatographic resolution behaviors of slightly modified anilide derivatives of N-acyl-alpha-amino acids. The chiral recognition mechanism proposed is quite similar to that advanced previously for the resolution of N-(3,5-methoxybenzoyl)-alpha-amino acids on CSP 1, even though the interaction sites of the two types of analytes were totally different from each other. The apparent similarity of the two chiral recognition mechanisms was assumed to stem from the identical interaction modes of the two types of analytes with the CSP. In addition, the dependence of the enantioselectivity of anilide derivatives of N-acyl-alpha-amino acids on the length of the alkyl tail of the N acyl group of analytes was rationalized to stem from the intercalation of the N acyl group of the (R)-enantiomer of analytes between the tethers of the CSP. PMID- 12112347 TI - Quantitative analysis of chiral compounds from unresolved peaks in capillary electrophoresis using multivariate calibration with experimental design and artificial neural networks. AB - Quantitation of optical isomers can be achieved even from incompletely resolved peaks with a multivariate calibration applying a combination of experimental design and artificial neural networks (ANN). Using the proposed approach, method development can be more efficient and analysis time shortened while quantitation of optical isomers with acceptable precision (+/-1-3%) can be achieved. PMID- 12112348 TI - Antibodies directed against L and D isovaline using a chemical derivatizing reagent for the measurement of their enantiomeric ratio in extraterrestrial samples: first step-production and characterization of antibodies. AB - Determining the enantiomeric ratio of amino acids in meteorites requires very sensitive and precise measurements. In this study, an immunochemical approach, combined with new chemical derivatizing agents, was investigated for the measurement of the enantiomeric ratio of isovaline. In the initial step, L and D isovaline were derivatized with epsilon-benzyloxycarbonyl-L-lysine-(t-butyl ester)-chloroethylnitrosourea (Z-L-Lys-(OtBu)-CENU). The Z group was hydrolyzed and the resulting isovaline derivatives (L-Lys(OtBu)-L-isovaline and L-Lys(OtBu) D-isovaline) were conjugated with protein using glutaraldehyde and reduced with sodium borohydride. Rabbits were immunized with the immunogenic conjugates thus obtained. Antibodies were characterized using many compounds, both derivatized and underivatized, in competitive ELISA tests. These competition experiments performed enabled us to establish the following results: 1) unconjugated L Lys(OtBu)-L-isovaline and L-Lys(OtBu)-D-isovaline were poorly recognized; 2) all related L-Lys(OtBu)-alpha-hydrogenated amino acids (L and D) were not recognized at all, which eliminates the possibility of the measurements being distorted by contamination; 3) only conjugated L-Lys(OtBu)-alpha-amino-isobutyric acid (AIB) was recognized by the antibody, 4) the enantiomeric discrimination of L and D isovaline through their derivatives (diastereoisomeric L-Lys(OtBu)-L-isovaline and L-Lys(OtBu)-D-isovaline) was in accordance with the measurement of their enantiomeric ratio. Immunopurification was shown to enhance antibody specificity. The strategy employed shows potential for the quantification of meteoritic amino acids. PMID- 12112350 TI - Individual variations in aging of the male urethral rhabdosphincter in Japanese. AB - Although the degenerative changes with aging of the male urethral rhabdosphincter (URS) have been investigated, its individual morphological variations are still unclear. To provide an anatomical basis for clinical evaluation of the individual URS function in the aged, we investigated the structural differences in the URS of 25 elderly Japanese men using semiserial sections stained immunohistochemically and by hematoxylin-eosin. Before removal of the histological specimens, we dissected the ischioanal fossa and labeled several structures by carbon particles to allow proper orientation during the histological observations. In addition, macroscopic slices (10 mm thickness) made from five other male pelves were examined and, when necessary, followed by routine histological procedure to confirm the gross observations. An extended circular URS (over (1/2) circumferential configuration) was found in 15/25 cadavers, but showed very limited height (proximal-distal length) and thickness. A more restricted URS, including even a thin, arc-like pattern, was observed in the remaining cadavers. The attachment of the URS to the smooth muscle layer was loose and usually clearly separated. Continuation between the URS and deep transverse perineal muscle was sometimes observed. The thick fascia of the levetor ani, with high content of smooth muscles, usually provided the lateral or dorsal insertions of the URS. Our results in elderly Japanese subjects suggest that the sphincteric action is weak or incomplete. We suggest that the elderly URS maintains continence by retracting the urethra backward and upward with the aid of the levator sling, rather than the real sphincteric action expected in younger men. PMID- 12112349 TI - Chirality and the origin of life: in situ enantiomeric separation for future space missions. AB - Two different methods of derivatization were studied in order to select and optimize one for the in situ enantiomeric separation of amino acids present in Martian samples. The method, using DMF-DMA [N,N-dimethylformamide dimethyl acetal], is simple and easily automated. However, byproducts of the reaction interfere in the gas chromatograms and mass spectrometry detection is needed for in situ analysis. The chloroformate derivatization has several advantages, including the use of achiral robust capillary column, room temperature reaction, and short analysis. The choice of the definitive derivatization method will depend on the energy and time devoted to the analysis of amino acids in the next Mars exploration missions. PMID- 12112351 TI - William Hunter's Gravid Uterus: the specimens and plates. AB - William Hunter's collection of anatomical specimens of the pregnant uterus forms one of the finest displays in the Anatomy Museum at the University of Glasgow. We were interested to know which specimens in the Museum matched the plates in Hunter's The Anatomy of the Human Gravid Uterus Exhibited in Figures (1774). In our investigation we were greatly assisted by Teacher's Catalogue of the Anatomical and Pathological Preparations of Dr William Hunter (1900). Thirteen specimens in the Museum and one from the pathological collection at the Royal Infirmary are represented in Hunter's book. The specimens can be recognized in 25 of its illustrations. A further three specimens may correspond to figures but we could not prove this. With one possible exception, all the specimens matching plates noted in Teacher's catalogue remain in the Museum and one believed missing in Marshall's (1970) revision of the catalogue has been found. PMID- 12112352 TI - Anatomical variations of the sural nerve. AB - An anatomical study of the formation of the sural nerve (SN) was carried out on 76 Thai cadavers. The results revealed that 67.1% of the SNs were formed by the union of the medial sural cutaneous nerve (MSCN) and the lateral sural cutaneous nerve (LSCN); the MSCN and LSCN are branches of the tibial and the common fibular (peroneal) nerves, respectively. The site of union was variable: 5.9% in the popliteal fossa, 1.9% in the middle third of the leg, 66.7% in the lower third of the leg, and 25.5% at or just below the ankle. One SN (0.7%) was formed by the union of the MSCN and a different branch of the common fibular nerve, running parallel and medial to but not connecting with the LSCN, which joined the MSCN in the lower third of the leg. The remaining 32.2% of the SNs were a direct continuation of the MSCN. The SNs ranged from 6-30 cm (mean = 14.41 cm) in length with a range in diameter of 3.5-3.8 mm (mean = 3.61 mm), and were easily located 1-1.5 cm posterior to the posterior border of the lateral malleolus. The LSCNs were 15-32 cm long (mean = 22.48 cm) with a diameter between 2.7-3.4 mm (mean = 3.22 mm); the MSCNs were 17-31 cm long (mean = 20.42 cm) with a diameter between 2.3-2.5 mm (mean = 2.41 mm). Clinically, the SN is widely used for both diagnostic (biopsy and nerve conduction velocity studies) and therapeutic purposes (nerve grafting) and the LSCN is used for a sensate free flap; thus, a detailed knowledge of the anatomy of the SN and its contributing nerves are important in carrying out these and other procedures. PMID- 12112353 TI - Morphologic study of the connections between the accessory nerve and the posterior root of the first cervical nerve. AB - The purpose of this study was to evaluate the anastomotic relationships between the accessory nerve and the posterior root of the first cervical nerve and to determine the course of the posterior root nerve fibers after anastomosis. The relationships between these two nerves were studied in 100 sides of the spinal cord and then classified into four types. In the most common type of anastomosis (38% of cases), either a branch from the posterior C1 root was seen to course cranially and join the accessory nerve, or the posterior root and accessory nerve fused as they coursed orthogonal to one other. In the second most common type (36% of cases), the accessory nerve anastomosed with a posterior C1 root that had no direct connection with the spinal cord; the nerve fibers of the posterior root were observed in stained samples to course caudally along the accessory nerve and join the posterior C2 root. In the least common type (6% of cases), no connection was observed between the accessory nerve and the posterior C1 root. In the next least common type (20% of cases), the posterior C1 root was absent and a connecting branch was sometimes observed between the accessory nerve and the anterior C1 root. PMID- 12112354 TI - Sympathetic fiber origin of the superior laryngeal nerve and its branches: an anatomic study. AB - An anatomic study of 50 fresh adult human cadavers was carried out to classify the nervous anastomoses between the ipsilateral cervical sympathetic chain (CSC)/superior cervical sympathetic ganglion (SCSG) and the superior laryngeal nerve with its branches (SLNwb). Following the pattern of a prospective randomized study, the necks were dissected bilaterally to identify the SLNwb, CSC and SCSG. The nervous connections between the CSC/SCSG and the SLNwb were grouped according to Sun and Chang's (1991, Surg. Radiol. Anat. 13:175-180) types as well as according to a new classification, tie, based on the location and number of connections between the CSC/SCSG and the SLNwb. Connections between the CSC/SCSG and the SLNwb were found in 95 of 100 dissections. According to our tie classification, the results demonstrated that the e(n) was the most frequent tie group in the sample (59%), followed by t(n) (15%), i(n)e(n) (11%), t(n)e(n) (8%), no nervous anastomosis (5%), and i(n) (2%), where t, i, e represent nervous connections between the CSC/SCSG and the trunk, external branch, and internal branch of the SLN, respectively, and n represents the number of connections. Therefore, the nervous communications between the CSC/SCSG and the SLNwb were frequent and could easily be classified by the origin and number of nervous connections. Although their physiologic role remains unclear, the SLNwb may be an important pathway for nerve fibers from the CSC/SCSG to reach the larynx. PMID- 12112355 TI - Obturator canal fat plug: a pre-hernial condition? AB - Although obturator hernias are rare, they are associated with a high mortality, as diagnosis is often delayed and the condition tends to occur in the elderly. Previous researchers have postulated that obturator hernias begin with invagination of pre-peritoneal fat through the pelvic orifice of the obturator canal, forming a fat plug. With chronically raised intra-abdominal pressure, or sudden weight loss, this can progress to a clinical obturator hernia. We dissected 95 Caucasian cadaveric hemi-pelvises (47 males; 48 females) in order to investigate the frequency of fat plugs and examine the validity of the current hypothesis regarding obturator hernia pathogenesis. The mean age (SD) of the specimens was 83 (9) years, with 48 (24 male; 24 female) from the left side and 47 (23 male; 24 female) from the right side. A fat plug was found in 21 canals (22%); the majority were female (71%, P = 0.03), and right sided (62%, P = 0.20). The mean (SD) length was 20.0 mm (6.3 mm), with mean diameter 5.9 mm (1.8 mm). A shallow peritoneal dimple was also found overlying fat plugs in two specimens. No visceral herniations were found. The sex and side distribution of the obturator canal fat plugs we found are similar to those of obturator hernias reported in the literature, supporting the hypothesis that fat plugs are pre-hernial. It is unlikely that fat plugs are a high-risk condition, but dimples over these plugs may be a marker of potential hernia formation. PMID- 12112356 TI - Inguinal canal "lipoma". AB - Groin dissection was performed in adult male post-mortem subjects to establish the prevalence of inguinal canal "lipoma." Thirty-six body halves (age range 24 92 years) were studied. Of these, 27 (75%) contained a discrete mass of fat within the inguinal canal. This mass was always continuous with the preperitoneal fat through the deep inguinal ring. Nineteen of these 27 masses (70%), displayed a characteristic pedunculated form with a bulbous distal end. Eighteen of the 36 dissections (50%), revealed a mass more than 4 cm in length. Six dissections showed extension of the mass beyond the superficial inguinal ring and three of these six (8% of the 36 groins studied) showed distortion of the proximal spermatic cord with a mass at the superficial inguinal ring. The masses submitted for histology comprised mature adipose tissue and all but two of these were reported as having an adherent capsule. No significant correlation was found between mass length and either subject age or body mass index (BMI) but a statistically significant correlation between the length of the fat mass on the left and right sides was shown. This study demonstrates that the inguinal canal "lipoma" is a common feature in an adult male population and may be of sufficient size to cause clinical misdiagnosis. The high prevalence, characteristic location and appearance of the "lipoma" suggest a developmental etiology. PMID- 12112357 TI - Extensor digitorum brevis manus: anatomical, radiological and clinical relevance. A review. AB - The extensor digitorum brevis manus (EDBM) is a supernumerary muscle in the dorsum of the hand frequently misdiagnosed as a dorsal wrist ganglion, exostosis, tendon sheath cyst or synovitis. Its presence in a living subject, confirmed by magnetic resonance imaging (MRI), is presented together with a review of the hitherto reported cases and the results of an anatomical study on 128 adult human cadavers (59 males and 69 females). The EDBM was found in three (2.3%) of the 128 cadavers. It occurred in two (3.4%) of the 59 male cadavers (one bilateral and one unilateral on the right side) and in one (1.5%) of the 69 female cadavers (unilateral on the left side). Consequently, the EDBM was found in four (1.6%) of the 256 upper limbs. It originated from the dorsal wrist capsule within the compartment deep to the extensor retinaculum for the extensor digitorum and inserted into the extensor hood of the index finger in one case and into that of the middle finger in three cases. In both hands of the living subject, the origin was similar but the insertion was into the index and middle fingers. In all cases, it was innervated by the posterior interosseous nerve and its blood supply was provided by the posterior interosseous artery. PMID- 12112358 TI - Tibiofemoral angle in Malawians. AB - We determined the tibiofemoral angle from 323 unilateral radiographs of the knee joint in adult black Malawians, comprising 219 males and 104 females, aged 18-55 years. The mean tibiofemoral angle in males was 174.14 with a standard deviation (SD) of 3.47 degrees and, in females, 174.46 SD 4.30 degrees. The range of the angle for both genders was 164-185 degrees. There was no significant difference in the angle between male and female Malawian subjects (P > 0.1). The results showed ethnic differences from Caucasian values (often quoted in standard textbooks of anatomy and orthopedics) and did not show the sexual dimorphism seen in Caucasian subjects. Comparison of the results with those in Nigerians and East Africans demonstrated regional and country-to-country variations. The clinical importance of the angle in the diagnosis and management of valgus and varus deformities is stressed for practicing orthopedic surgeons. PMID- 12112359 TI - Application of osteology to forensic medicine. AB - The four main features of biological identity are sex, age, stature, and ethnic background. The forensic osteologist aims to establish these attributes for an individual from their skeletal remains. Many techniques are available for the osteological determination of sex in the adult but it is one of the most difficult biological factors to ascribe to juvenile remains. Conversely, there are a multitude of markers to estimate age in the young skeleton but ageing becomes less accurate with increasing years. Stature is usually a relatively straightforward parameter to establish in the adult. In the juvenile, it is naturally correlated with age but is complicated by differences in rates of growth both between the sexes and between individuals. Determination of ethnic identity is the least reliable and is hampered by lack of data on many populations. This paper reviews the principal methods used to establish identity and comments on their reliability and accuracy in the forensic context. PMID- 12112360 TI - Hepaticocystic duct: a rare anomaly of the extrahepatic biliary system. PMID- 12112361 TI - Glial induction of blood-brain barrier-like L-system amino acid transport in the ECV304 cell line. AB - The blood-brain barrier (BBB) is formed by the presence of tight junction complexes between brain endothelial cells that restrict paracellular permeability. As a consequence, a number of transport proteins are expressed on cerebral endothelial cells to facilitate the transport of nutrients into the brain. Although the modulation of barrier tight junction properties by glial conditioned medium and by second messengers is well established, little is known about the effects of these factors on carrier-mediated BBB transport processes. The ECV304 cell line shows an endothelial phenotype and can be induced to upregulate certain BBB features in the presence of glial factors. In the present study, we have examined the effect of conditioned medium derived from rat C6 glioma cells (C6CM) on the function of the L-system amino acid transporter in ECV304 cells, using L-leucine as the model substrate, and have determined whether the changes observed can be mimicked by modulating intracellular cAMP levels. ECV304 cells exposed to C6CM exhibited a significant increase in both the affinity of leucine transport and the diffusional constant (Michaelis-Menten), while the maximal transport capacity remained unchanged. Conversely, acute exposure to modulators of the PKA and PKC second messenger pathways was found to reduce significantly the maximal transport capacity and diffusion constants, while transport affinity remained unchanged. In both cases, the maximal flux of leucine was increased, indicating transport of greater efficiency. This study indicates that exposure of ECV304 cells to C6CM provides an influence inducing L system transport properties characteristic of brain endothelial cells. Furthermore, it appears that L-system-mediated transport of amino acids can be modulated by several distinct pathways. PMID- 12112362 TI - Ceramide levels are inversely associated with malignant progression of human glial tumors. AB - Ceramide represents an important sphingoid mediator involved in the signaling pathways that control cell proliferation, differentiation, and death. To determine whether ceramide levels correlate with the malignant progression of human astrocytomas, we investigated these levels in surgical specimens of glial tumors of low-grade and high-grade malignancy. Tumor samples obtained from 52 patients who underwent therapeutic removal of primary brain tumors were used. The tumors were classified according to standard morphologic criteria and were grouped into tumors of low-grade and high-grade malignancy. Sections of normal brain tissue adjacent to the tumor were also analyzed in 11 of the 52 patients. After extraction and partial purification, ceramide was measured by quantitative derivatization to ceramide-1-phosphate using diacylglycerol kinase and [gamma (32)P]ATP. Ceramide levels were significantly lower in the combined high-grade tumors compared with low-grade tumors and in both tumor groups compared with peritumoral tissue. The results indicate an inverse correlation between the amount of ceramide and tumor malignancy as assessed by both the histological grading and ganglioside pattern. Moreover, overall survival analysis of 38 patients indicates that ceramide levels are significantly associated with patient survival. The present findings indicate that ceramide is inversely associated with malignant progression of human astrocytomas and poor prognosis. The downregulation of ceramide levels in human astrocytomas emerges as a novel alteration that may contribute to glial neoplastic transformation. The low ceramide levels in high-grade tumors may provide an advantage for their rapid growth and apoptotic resistant features. This study appears to support the rationale for the potential benefits of a ceramide-based chemotherapy. PMID- 12112363 TI - Stage-specific gene expression in early differentiating oligodendrocytes. AB - The screening of a differential library from precursor and differentiated oligodendrocytes, obtained through the representational difference analysis (RDA) technique, has generated a number of cDNA recombinants corresponding to mRNA coding for known and unknown proteins: (1) mRNA coding for proteins involved in protein synthesis, (2) mRNA coding for proteins involved in the organization of the cytoskeleton, and (3) mRNA coding for proteins of unknown function. The expression profile of the mRNA was studied by Northern blot hybridization to the poly-A(+) mRNA from primary rat progenitor and differentiated oligodendrocytes. In most cases, hybridization to the precursor was higher than hybridization to the differentiated mRNA, supporting the validity of the differential screening. Hybridization of the cDNA to rat cerebral hemisphere and brain stem poly-A(+) mRNA, isolated from 1- to 90-day-old rats, confirms the results obtained with the mRNA from differentiating oligodendrocytes. The intensity of the hybridization bands decreases as differentiation proceeds. The pattern of expression observed in oligodendrocytes is different from that found in the brain only in the case of the nexin-1 mRNA, the level of which remains essentially constant throughout differentiation both in the brain stem and in the cerebral hemispheres, in agreement with the published data. In contrast, the intensity of hybridization to the oligodendrocyte mRNA is dramatically lower in the differentiated cells compared with the progenitor oligodendrocyte cells. Some of the recombinant cDNA represent mRNA sequences present at high frequency distribution in the cells, while others belong to the rare sequences group. Six recombinants code for proteins of the ribosomal family, suggesting that of approximately 70 known ribosomal proteins, only a few are upregulated during oligodendrocyte differentiation. The third category of open reading frame (ORF) is represented by rare messengers coding for proteins of unknown functions and includes six clones: RDA 279, 11, 95, 96, 254, and 288. PMID- 12112364 TI - Accumulation of intracellular ascorbate from dehydroascorbic acid by astrocytes is decreased after oxidative stress and restored by propofol. AB - Primary rat astrocyte cultures absorbed dehydroascorbic acid from the medium and reduced it to intracellular ascorbate. Uptake of dehydroascorbic acid (5-200 microM) was inhibited only partially by glucose (10 mM). The remaining glucose insensitive component of dehydroascorbic acid uptake was inhibited reversibly by sulfinpyrazone (IC(50) = 80 microM). Dehydroascorbic acid uptake was not mediated by Na(+)-ascorbate cotransporters or volume-sensitive anion channels because it was neither Na(+)-dependent nor blocked by the channel antagonist, 4,4' diisothiocyanatostilbene-2,2'-disulfonic acid. Oxidative stress, induced in astrocytes by the lipophilic radical generator tert-butyl hydroperoxide, decreased intracellular glutathione concentration and inhibited accumulation of intracellular ascorbate from dehydroascorbic acid. Subsequent administration of either the native antioxidant alpha-tocopherol (200 microM) or anesthetic concentrations of the antioxidant sedative propofol (1-8 microM, administered 30 min after tert-butyl hydroperoxide), did not change glutathione concentration but restored the ability of astrocytes to accumulate intracellular ascorbate from dehydroascorbic acid. These results are consistent with a novel mechanism of astrocytic ascorbate accumulation that is inhibited by lipophilic radicals and protected by lipophilic antioxidants such as propofol. PMID- 12112365 TI - A new neuromodulatory pathway with a glial contribution mediated via A(2a) adenosine receptors. AB - A low concentration (10 nM) of adenosine potentiated hippocampal neuronal activity via A(2a) adenosine receptors without affecting presynaptic glutamate release or postsynaptic glutamatergic conductance. Adenosine inhibited glutamate uptake through the glial glutamate transporter, GLT-1, via A(2a) adenosine receptors. In addition, adenosine stimulated GLT-1-independent glutamate release from astrocytes, possibly in response to a rise in intracellular Ca(2+), via A(2a) adenosine receptors involving PKA activation. Those adenosine actions could lead to an increase in synaptic glutamate concentrations responsible for the potentiation of hippocampal neuronal activity. The results of the present study thus represent a novel neuromodulatory pathway with a glial contribution, bearing both inhibition of GLT-1 function and stimulation of glial glutamate release, as mediated via A(2a) adenosine receptors. PMID- 12112366 TI - Vasoactive intestinal peptide and pituitary adenylate cyclase-activating polypeptide inhibit chemokine production in activated microglia. AB - Microglia react to even minor disturbances in CNS homeostasis and function as critical regulators of CNS inflammation. Activated microglia secrete inflammatory mediators such as cytokines and chemokines, which contribute to the pathophysiological changes associated with several neuroimmunologic disorders. Microglia-derived inflammatory chemokines recruit various populations of immune cells, which initiate and maintain the inflammatory response against foreign antigens. Entry and retention of activated immune cells in the CNS is a common denominator in a variety of traumatic, ischemic, and degenerative diseases. Vasoactive intestinal peptide (VIP) and pituitary adenylate cyclase-activating polypeptide (PACAP) are two structurally related neuropeptides that function as potent anti-inflammatory factors in the periphery. Here we investigated the effects of VIP and PACAP on chemokine production by activated microglia. VIP and PACAP inhibit the expression of the microglia-derived CXC chemokines MIP-2 and KC, and of the CC chemokines MIP-1alpha, -1beta, MCP-1, and RANTES. The inhibition of chemokine gene expression correlates with an inhibitory effect of VIP/PACAP on NFkB binding. The VIP/PACAP inhibition of both chemokine production and of NFkB binding is mediated through the specific receptor VPAC1 and involves a cAMP-dependent intracellular pathway. Of biological significance is the fact that the inhibition of chemokine production by VIP/PACAP leads to a significant reduction in the chemotactic activity generated by activated microglia for peripheral leukocytes, i.e., neutrophils, macrophages, and lymphocytes. Because reduction in the number and activation of infiltrating leukocytes represents an important factor in the control of inflammation in the CNS, VIP and/or PACAP released by neurons during an inflammatory response could serve as neuronal survival factors by limiting the inflammatory process. PMID- 12112367 TI - Chlorotoxin, a scorpion-derived peptide, specifically binds to gliomas and tumors of neuroectodermal origin. AB - Highly migratory neuroectodermal cells share a common embryonic origin with cells of the central nervous system (CNS). They include enteric, parasympathetic, sympathoadrenal, and sensory neurons of the peripheral nervous system, Schwann cells, melanocytes, endocrine cells, and cells forming connective tissue of the face and neck. Because of their common embryologic origin, these cells and the tumors that derive from them can share genetic and antigenic phenotypes with gliomas, tumors derived from CNS glia. We recently discovered that chlorotoxin (ClTx), a 4-kD peptide purified from Leiurus quinquestriatus scorpion, is a highly specific marker for glioma cells in biopsy tissues (Soroceanu et al. Cancer Res 58:4871-4879, 1998) that can target tumors in animal models. We report on the specificity of ClTx as a marker for tumors of neuroectodermal origin that include peripheral neuroectodermal tumors (PNET) and gliomas. Specifically, we histochemically stained frozen and paraffin tissue sections of human biopsy tissues from 262 patients with a synthetically manufactured and biologically active ClTx bearing an N-terminal biotin. The vast majority (74 of 79) of primary human brain tumors investigated showed abundant binding of ClTx with greater than 90% ClTx-positive cells in each section. By comparison, 32 biopsies of uninvolved brain used for comparison were largely ClTx-negative, with only a few isolated reactive astrocytes showing some ClTx binding. However, as with gliomas, the vast majority of PNETs examined showed specific ClTx binding (31 of 34). These include medulloblastomas (4 of 4), neuroblastomas (6 of 7), ganglioneuromas (4 of 4), melanomas (7 of 7), adrenal pheochromocytomas (5 of 6), primitive PNET (1), small cell lung carcinoma (2 of 3), and Ewing's sarcoma (2 of 2). Under identical staining conditions, normal tissues from brain, skin, kidney, and lung were consistently negative for ClTx. These results suggest that chlorotoxin is a reliable and specific histopathological marker for tumors of neuroectodermal origin and that chlorotoxin derivatives with cytolytic activity may have therapeutic potential for these cancers. PMID- 12112368 TI - GM-CSF and M-CSF modulate beta-chemokine and HIV-1 expression in microglia. AB - Significant numbers of patients with acquired immunodeficiency syndrome (AIDS) develop CNS infection primarily in macrophages and microglial cells. Therefore, the regulation of human immunodeficiency virus type 1 (HIV-1) infection and activation of the brain mononuclear phagocytes subsequent to infection are important areas of investigation. In the current report, we studied the role of granulocyte-macrophage colony-stimulating factor (GM-CSF) and macrophage-CSF (M CSF) in the expression of antiviral beta-chemokines and HIV-1 p24 in cultures of primary human fetal microglia. We found that stimulation with GM-CSF or M-CSF induced macrophage inflammatory proteins (MIP-1alpha and MIP-1beta) and augmented RANTES expression, after HIV-1 infection of microglia. This was not due to the effect of GM-CSF on viral expression because GM-CSF was neither necessary nor stimulatory for viral infection, nor did GM-CSF enhance the expression of env pseudotyped reporter viruses. Blocking GM-CSF-induced microglial proliferation by nocodazole had no effect on beta-chemokine or p24 expression. The functional significance of the GM-CSF-induced beta-chemokines was suggested by the finding that, in the presence of GM-CSF, exogenous beta-chemokines lost their anti-HIV-1 effects. We further show that although HIV-1-infected microglia produced M-CSF, they failed to produce GM-CSF. In vivo, GM-CSF expression was localized to activated astrocytes and some inflammatory cells in HIV-1 encephalitis, suggesting paracrine activation of microglia through GM-CSF. Our results demonstrate a complex interplay between CSFs, chemokines, and virus in microglial cells and may have bearing on the interpretation of data derived in vivo and in vitro. PMID- 12112369 TI - MHC class II-positive perivascular microglial cells mediate resistance to Cryptococcus neoformans brain infection. AB - Acquired resistance to the CNS pathogen Cryptococcus neoformans is mediated by CD4(+) T lymphocytes primed by exposure to antigen in the context of major histocompatibility class II (MHC II) molecules. In mouse brain, parenchymal and perivascular microglial cells may express interferon-gamma (IFN-gamma)-inducible MHC class II marker and thus interact with CD4(+) T cells. Primed effector T cells are retained in the infected CNS if antigen is encountered in proper MHC context and may deliver signals that potentiate microglia to enhanced fungistasis. Vaccinated C57BL6/J mice resist an ordinarily lethal C. neoformans rechallenge, but identically treated congenic Abeta(o/o) mice (MHC class II deficient; CD4(+) T-cell-deficient) do not. Nor can Abeta(o/o) mice be adoptively immunized by infusion of lymphocytes from vaccinated C57BL6/J donors, as are severe combined immunodeficient (SCID) mice (MHC class II-intact, lymphocyte deficient). Chimeric (C57BL/6J:Abeta(o/o)) mice with class II expression likely on perivascular microglia only were, like SCID mice, capable of adoptive immunization against C. neoformans brain infection. To the contrary, chimeric mice with class II expression likely only on parenchymal microglia were not capable of effective adoptive immunization against C. neoformans brain infection. Therefore, in order to mediate resistance to infection, primed CD4(+) T cells must interact with the replenishable perivascular microglial subset that lies in close proximity to cerebral vasculature. Although T cells may supply help in the form of inflammatory cytokines to parenchymal microglia, expression of class II on these cells appears unnecessary for antifungal activity. PMID- 12112370 TI - Increased syndecan expression by pleiotrophin and FGF receptor-expressing astrocytes in injured brain tissue. AB - Syndecan-1, -2, -3, and -4 are heparan sulfate proteoglycans that are differentially expressed during development and wound repair. To determine whether syndecans are also involved in brain injury, we examined the expression of syndecan core proteins genes in cryo-injured mouse brain, using in situ hybridization. All syndecan mRNA transcripts were similarly expressed in the region surrounding the necrotic tissue, exhibiting peak levels at day 7 after injury. Comparison with cellular markers showed that reactive astrocytes were the primary source of syndecans. Syndecans serve as co-receptors for fibroblast growth factor (FGF) and as a reservoir for another heparin-binding growth factor, pleiotrophin (PTN, or heparin-binding growth-associated molecule. In our model, FGF receptor1 (FGFR1) and PTN mRNA levels were upregulated in reactive astrocytes. The distribution patterns of FGFR1 and PTN overlapped considerably with those of syndecan-1 and -3 mRNAs, respectively. These results suggest that syndecans are expressed primarily in reactive astrocytes, and may provide a supportive environment for regenerating axons in concert with heparin-binding growth factors (e.g., FGF and PTN) in the injured brain. PMID- 12112371 TI - Long-term modulation of glucose utilization by IL-1 alpha and TNF-alpha in astrocytes: Na+ pump activity as a potential target via distinct signaling mechanisms. AB - Interleukin-1alpha (IL-1alpha) and tumor necrosis factor-alpha (TNF-alpha) markedly stimulate glucose utilization in primary cultures of mouse cortical astrocytes. The mechanism that gives rise to this effect, which takes place several hours after application of cytokine, has remained unclear. Experiments were conducted to identify the major signaling cascades involved in the metabolic action of cytokine. First, the selective IL-1 receptor antagonist (IL-1ra) prevents the effect of IL-1alpha on glucose utilization in a concentration dependent manner, whereas it has no effect on the action of TNF-alpha. Then, using inhibitors of three classical signaling cascades known to be activated by cytokines, it appears that the PI3 kinase is essential for the effect of both IL 1alpha and TNF-alpha, whereas the action of IL-1alpha also requires activation of the MAP kinase pathway. Participation of a phospholipase C-dependent pathway does not appear critical for both IL-1alpha and TNF-alpha. Inhibition of NO synthase by L-NAME did not prevent the metabolic response to both IL-1alpha and TNF-alpha, indicating that nitric oxide is probably not involved. In contrast, the Na(+)/K(+) ATPase inhibitor ouabain prevents the IL-1alpha- and TNF-alpha stimulated 2-deoxyglucose (2DG) uptake. When treatment of astrocytes with a cytokine was followed 24 h later by an acute application of glutamate, a synergistic enhancement in glucose utilization was observed. This effect was greatly reduced by ouabain. These data suggest that Na(+) pump activity is a common target for both the long-term metabolic action of cytokines promoted by the activation of distinct signaling pathways and the enhanced metabolic response to glutamate. PMID- 12112372 TI - RANTES stimulates inflammatory cascades and receptor modulation in murine astrocytes. AB - Cultured mouse astrocytes respond to the CC chemokine RANTES by production of chemokine and cytokine transcripts. Stimulation of astrocytes with 1 nM RANTES or 3-10 nM of the structurally related chemokines (eotaxin, macrophage inflammatory protein-1alpha and -beta [MIP-1alpha, MIP-1beta]) induced transcripts for KC, monocyte chemoattractant protein-1 (MCP-1), tumor necrosis factor-alpha (TNF alpha), MIP-1alpha, MIP-2, and RANTES in a chemokine and cell-specific fashion. Synthesis of chemokine (KC and MCP-1) and cytokine (TNF-alpha) proteins was also demonstrated. RANTES-mediated chemokine synthesis was specifically inhibited by pertussis toxin, indicating that G-protein-coupled chemokine receptors participated in astrocyte signaling. Astrocytes expressed CCR1 and CCR5 (the redundant RANTES receptors). Astrocytes derived from mice with targeted mutations of either CCR1 or CCR5 respond after RANTES stimulation, suggesting multiple chemokine receptors may separately mediate RANTES responsiveness in astrocytes. Preliminary data suggest activation of the MAP kinase pathway is also critical for RANTES-mediated signaling in astrocytes. Treatment with RANTES specifically modulated astrocyte receptors upregulating intercellular adhesion molecule 1 (ICAM-1) and downregulating CX3CR1 expression. Thus, after chemokine treatment, astrocytes release proinflammatory mediators and reprogram their surface molecules. The combined effects of RANTES may serve to amplify inflammatory responses within the central nervous system. PMID- 12112373 TI - Evidence for possible interactions between PLP and DM20 within the myelin sheath. AB - PLP and its smaller DM20 isoform constitute the major proteins of CNS myelin. Previous studies indicated a role for the proteins in maintaining the intraperiod line of the myelin sheath and the integrity of axons and suggested that both isoforms were necessary to provide these functions. The present study shows that each isoform is capable individually of inserting into compact myelin. Employing chromatographic extraction procedures designed to maintain the natural conformation of the proteins we found that most PLP and DM20 remained associated. Using an antibody specific to the PLP isoform, we were able to co immunoprecipitate DM20 from the major fraction of the extracted equine myelin and from mouse native whole myelin. We suggest that PLP and DM20 may form a hetero oligomeric complex within the myelin sheath, probably in association with specific lipids and that this arrangement is essential for the normal structure of myelin and axons. PMID- 12112374 TI - Mimosine prevents the death of glucose-deprived immunostimulated astrocytes by scavenging peroxynitrite. AB - Immunostimulated astrocytes become highly vulnerable to glucose deprivation (Choi and Kim: J Neurosci Res 54:870-875, 1998a). The increased vulnerability is caused by the enhanced level of peroxynitrite endogenously produced in glucose-deprived immunostimulated astrocytes. In the present study, we report that the plant amino acid mimosine can attenuate the increased death by scavenging peroxynitrite. Treatment with mimosine blocked the increase of nitrotyrosine immunoreactivity, a marker of peroxynitrite, in glucose-deprived immunostimulated astrocytes. Furthermore, mimosine directly inhibited the nitration of tyrosine residues of bovine serum albumin and the oxidation of dihydrorhodamine-123 to rhodamine-123 by peroxynitrite. Mimosine has been used experimentally as a cell cycle G1/S phase transition blocker (Lalande: Exp Cell Res 186:332-339, 1990; Hoffman et al.: Cytometry 12:26-32, 1991). Flow cytometry analysis, however, showed that the cytoprotective effect of mimosine was not attributed to its inhibition of cell cycle progression. Furthermore, under our experimental conditions, mimosine did not alter the levels of cell cycle regulatory proteins, including p21(WAF1/CIP1), cyclins D1 and E, and proliferating cell nuclear antigen. In addition, cyclin dependent kinase inhibitors olomoucine and roscovitine did not block the increased death. These results indicate that mimosine inhibits the augmented death of glucose-deprived immunostimulated astrocytes by scavenging peroxynitrite rather than suppressing the cell cycle progression. PMID- 12112375 TI - Characterization of myelination in the developing zebrafish. AB - Myelination, the process by which glial cells ensheath and electrically insulate axons, has been investigated intensely. Nevertheless, knowledge of how myelination is regulated or how myelinating cells communicate with neurons is still incomplete. As a prelude to genetic analyses of these processes, we have identified zebrafish orthologues of genes encoding major myelin proteins and have characterized myelination in the larval zebrafish. Expression of genes corresponding to proteolipid protein (PLP/DM20), myelin protein zero (P0), and myelin basic protein (MBP) is detected at 2 days postfertilization (dpf), first in the ventral hindbrain, close to the midline. During the next 8 days, expression spreads rostrally to the midbrain and optic nerve, and caudally to the spinal cord. DM20 is expressed in the CNS only, while MBP transcripts are detected both in the CNS and in Schwann cells of the lateral line, cranial nerves, and spinal motor nerves. Unlike its closest homologue, trout IP1, zebrafish P0 transcripts were restricted to the CNS. Ultrastructurally, the expression of myelin genes correlated well with myelination, although myelination showed a temporal lag. Myelinated axons were first detected at 4 dpf in the ventral hindbrain, where they were loosely wrapped by processes of glia cells. By 7 dpf, bundles of heavily myelinated axons were observed in the same region. Axons in the lateral line and optic nerves were also surrounded by compact myelin. Conservation in gene expression patterns and the early appearance of myelinated axons, support using the zebrafish to dissect the process of myelination by a genetic approach. PMID- 12112376 TI - Glutamate transporters and retinal excitotoxicity. AB - Glutamate appears to play a major role in several degenerative retinal disorders. However, exogenous glutamate is only weakly toxic to the retina when glutamate transporters on Muller glial cells are operational. In an ex vivo rat retinal preparation, we previously found that exogenous glutamate causes Muller cell swelling but does not trigger excitotoxic neurodegeneration unless very high concentrations that overwhelm the capacity of glutamate transporters are administered. To determine the role of glutamate transporters in Muller cell swelling and glutamate-mediated retinal degeneration, we examined the effects of DL-threo-beta-benzyloxyaspartate (TBOA), an agent that blocks glutamate transport but that unlike most available transport inhibitors is neither a substrate for transport nor a glutamate receptor agonist. We found that TBOA triggered severe retinal neurodegeneration attenuated by ionotropic glutamate receptor antagonists. TBOA-induced neuronal damage was also diminished by riluzole, an agent that inhibits endogenous glutamate release. In the presence of riluzole, to inhibit glutamate release plus TBOA to block glutamate uptake, the addition of low concentrations of exogenous glutamate triggered severe excitotoxic neuronal damage without inducing Muller cell swelling. We conclude that TBOA-sensitive glutamate transporters play an important role in regulating the neurodegenerative effects of glutamate in the rat retina. PMID- 12112377 TI - Astrocytes in adult rat brain express type 2 inositol 1,4,5-trisphosphate receptors. AB - Astrocytes respond to neuronal activity by propagating Ca(2+) waves elicited through the inositol 1,4,5-trisphosphate pathway. We have previously shown that wave propagation is supported by specialized Ca(2+) release sites, where a number of proteins, including inositol 1,4,5-trisphosphate receptors (IP(3)R), occur together in patches. The specific IP(3)R isoform expressed by astrocytes in situ in rat brain is unknown. In the present report, we use isoform-specific antibodies to localize immunohistochemically the IP(3)R subtype expressed in astrocytes in rat brain sections. Astrocytes were identified using antibodies against the astrocyte-specific markers, S-100 beta, or GFAP. Dual indirect immunohistochemistry showed that astrocytes in all regions of adult rat brain express only IP(3)R2. High-resolution analysis showed that hippocampal astrocytes are endowed with a highly branched network of processes that bear fine hair-like extensions containing punctate patches of IP(3)R2 staining in intimate contact with synapses. Such an organization is reminiscent of signaling microdomains found in cultured glial cells. Similarly, Bergmann glial cell processes in the cerebellum also contained fine hair-like processes containing IP(3)R2 staining. The IP(3)R2-containing fine terminal branches of astrocyte processes in both brain regions were found juxtaposed to presynaptic terminals containing synaptophysin as well as PSD 95-containing postsynaptic densities. Corpus callosum astrocytes had an elongated morphology with IP(3)R2 studded processes extending along fiber tracts. Our data suggest that PLC-mediated Ca(2+) signaling in astrocytes in rat brain occurs predominantly through IP(3)R2 ion channels. Furthermore, the anatomical arrangement of the terminal astrocytic branches containing IP(3)R2 ensheathing synapses is ideal for supporting glial monitoring of neuronal activity. PMID- 12112378 TI - IGF-I and microglia/macrophage proliferation in the ischemic mouse brain. AB - We have used a model of hypoxic-ischemic brain injury in adult male C57BL/6 mice to study insulin-like growth factor-I (IGF-I) and IGF-binding protein (IGFBP) expression in response to cerebral hypoxia-ischemia (H/I) in the adult mouse. A period of 20 min of H/I that resulted in histopathology in cortex, striatum, and thalamus was correlated with induction of mRNA for IGF-I, IGFBP-2, IGFBP-3, IGFBP 5, and glial fibrillary acidic protein (GFAP) by 4 days of recovery. Increased IGF-I mRNA was located within damaged regions and was surrounded by IGFBP-2 mRNA expression. The results of combined immunostaining/in situ hybridzation showed that the cells expressing IGFBP-2 mRNA were also GFAP-positive and comprised a subset of activated astrocytes immediately surrounding areas of damage. In contrast, staining within damaged regions showed high numbers of cells immunopositive for F4/80 and lectin B(4) indicative of microglia and macrophages but no cells immunopositive for the astrocytic proteins GFAP or S-100beta. Microglia/macrophages within the damaged areas expressed IGF-I mRNA and were also immunopositive for the proliferating cell nuclear antigen. To determine whether expression of IGF-I could contribute to proliferation of microglia, we treated purified cultures of adult brain microglia with IGF-I in the presence of (3)H thymidine. IGF-I stimulated a twofold increase in DNA synthesis in cultures of adult brain microglia. Taken together with previous data demonstrating that IGF-I promotes proliferation of peripheral macrophages, these data support the hypothesis that IGF-I is an autocrine/paracrine mitogen for microglia/macrophages after H/I. PMID- 12112380 TI - Molecular analysis of mutations at the HPRT and TK loci of human lymphoblastoid cells after combined treatments with 3'-azido-3'-deoxythymidine and 2',3' dideoxyinosinedagger. AB - Combinations of antiretroviral drugs that include nucleoside reverse transcriptase inhibitors (NRTIs) are superior to single-agent regimens in treating or preventing HIV infection, but the potential long-term health hazards of these treatments in humans are uncertain. In earlier studies, our group found that coexposure of TK6 human lymphoblastoid cells to 3'-azido-2',3' dideoxythymidine (AZT) and 2',3'-dideoxyinosine (ddI), the first two NRTIs approved by the FDA as antiretroviral drugs, produced multiplicative synergistic enhancement of DNA incorporation of AZT and mutagenic responses in both the HPRT and TK reporter genes, as compared with single-drug exposures (Meng Q et al. [2000a]: Proc Natl Acad Sci USA 97:12667-12671). The purpose of the current study was to characterize the mutational specificity of equimolar mixtures of 100 microM or 300 microM AZT + ddI at the HPRT and TK loci of exposed cells vs. unexposed control cells, and to compare the resulting mutational spectra data to those previously found in cells exposed to AZT alone (Sussman H et al. [1999]: Mutat Res 429:249-259; Meng Q et al. [2000b]: Toxicol Sci 54:322-329). Molecular analyses of HPRT mutant clones were performed by reverse transcription-mediated production of cDNA, PCR amplification, and cDNA sequencing to define small DNA alterations, followed by multiplex PCR amplification of genomic DNA to define the fractions of deletion events. TK mutants with complete gene deletions were distinguished by Southern blot analysis. The observed HPRT mutational categories included point mutations, microinsertions/microdeletions, splicing-error mutations, and macrodeletions including partial and complete gene deletions. The only significant difference or shift in the mutational spectra for NRTI-treated cells vs. control cells was the increase in the frequency of complete TK gene deletions following exposures (for 3 days) to 300 microM AZT-ddI (P = 0.034, chi square test of homogeneity); however, statistical analyses comparing the observed mutant fraction values (measured mutant frequency x percent of a class of mutation) between control and NRTI-treated cells for each class of mutation showed that the occurrences of complete gene deletions of both HPRT and TK were significantly elevated over background values (0.34 x 10(-6) in HPRT and 6.0 x 10(-6) in TK) at exposure levels of 100 microM AZT-ddI (i.e., 1.94 x 10(-6) in HPRT and 18.6 x 10(-6) in TK) and 300 microM AZT-ddI (i.e., 5.6 x 10(-6) in HPRT and 34.6 x 10(-6) in TK) (P < 0.05, Mann-Whitney U-statistic). These treatment related increases in complete gene deletions were consistent with the spectra data for AZT alone (ibid.) and with the known mode of action of AZT and ddI as DNA chain terminators. In addition, cotreatments of ddI with AZT led to substantial absolute increases in the mutant fraction of other classes of mutations, unlike cells exposed solely to AZT [e.g., the frequency of point mutations among HPRT mutants was significantly increased by 130 and 323% over the background value (4.25 x 10(-6)) in cells exposed to 100 and 300 microM AZT-ddI, respectively]. These results indicate that, at the same time that AZT-ddI potentiates therapeutic or prophylactic efficacy, the use of a second NRTI with AZT may confer a greater cancer risk, characterized by a spectrum of mutations that deviates from that produced solely by AZT. PMID- 12112381 TI - Mutant frequency and mutational spectra in the Tk and Hprt genes of N-ethyl-N nitrosourea-treated mouse lymphoma cellsdagger. AB - The mouse lymphoma assay (MLA) utilizing the Tk gene is widely used to identify chemical mutagens. The autosomal location of the Tk gene allows for the detection of a wide range of mutational events, from point mutations to chromosome alterations. However, chemically induced point mutation spectra in the Tk gene of mouse lymphoma cells have not been characterized. In this study, we determined and compared the mutagenicity and mutational spectra of N-ethyl-N-nitrosourea (ENU) in the Tk and Hprt genes of mouse lymphoma cells. Treatment of L5178Y mouse lymphoma cells with 100 microg/ml ENU induced a Tk mutant frequency of 756 x 10( 6) and an Hprt mutant frequency of 311 x 10(-6). Sequence analysis of Tk and Hprt mutant cDNAs showed a similar overall mutation pattern in the two genes with base pair substitutions accounting for 83% of non-loss of heterozygosity mutations in the Tk gene and 75% of all mutations in the Hprt gene. The most common point mutation induced by ENU was G:C --> A:T transition (36 and 28% of independent mutations detected in the Tk and Hprt genes, respectively). The mutation spectra induced by ENU in both the Tk and Hprt genes were different from the respective patterns produced in mutants from untreated cells. About 9% of Tk and 7% of Hprt mutations from control cells were in-frame deletions, whereas no such mutations were found among the ENU-induced Tk and Hprt mutations. Our results indicate that ENU produces a chemical-specific point mutational profile in the Tk gene of mouse lymphoma cells that is remarkably similar to that found in the X-linked Hprt gene. This study provides evidence that the MLA can be used not only to detect point mutagens but also for analysis of mutational spectra. PMID- 12112382 TI - Urothelial cell DNA adducts in rubber workers. AB - Workers employed in the rubber industry appear to have a significant excess cancer risk in a variety of sites, including cancer of the urinary bladder. In this cross-sectional study, we investigated the occurrence of DNA adducts in exfoliated bladder cells of currently exposed, nonsmoking rubber workers (n = 52) and their relationship with occupational exposure estimates and acetylation phenotype (NAT2). Four DNA adducts were identified, with the proportion of positive samples (e.g., DNA samples with quantifiable levels of a specific DNA adduct) ranging from 3.8 to 79%. The highest proportion of positive samples and the highest relative adduct labeling levels were in workers involved in the production functions "mixing" and "curing," areas with potential for substantial exposure to a wide range of chemical compounds used in rubber manufacturing (P < 0.05 for adducts 2 and/or 3, compared to all other departments). No statistically significant relationships were found between identified DNA adducts and urinary mutagenicity or personal inhalable and dermal exposure estimates. Interestingly, subjects with a fast NAT2 acetylation phenotype tended to have higher levels of DNA adducts. This study suggests that rubber workers engaged in mixing and curing may be exposed to compounds that can form DNA adducts in urothelial cells. Larger studies among rubber workers should be conducted to study in more detail the potential carcinogenicity of exposures encountered in these work areas. PMID- 12112383 TI - Identification of N-(deoxyguanosin-8-yl)-4-azobiphenyl by (32)P-postlabeling analyses of DNA in human uroepithelial cells exposed to proximate metabolites of the environmental carcinogen 4-aminobiphenyl. AB - DNA adducts formed in human uroepithelial cells (HUC) following exposure to N hydroxy-4-aminobiphenyl (N-OH-ABP), the proximate metabolite of the human bladder carcinogen 4-aminobiphenyl (ABP), were analyzed by the (32)P-postlabeling method. Two adducts detected by (32)P-postlabeling were previously identified as the 3',5'-bisphospho derivatives of N-(deoxyguanosin-8-yl)-4-aminobiphenyl (dG-C8 ABP) and N-(deoxyadenosin-8-yl)-4-aminobiphenyl (dA-C8-ABP) (Frederickson S et al. [1992] Carcinogenesis 13: 955-961; Hatcher and Swaminathan [1995b] Carcinogenesis 16: 295-301). In contrast to the dG-C8-ABP adduct, which was 3' dephosphorylated by nuclease P1, dA-C8-ABP was resistant to nuclease P1, thus providing an enrichment step before postlabeling. Autoradiography of the two dimensional thin-layer chromatogram of the postlabeled products obtained following nuclease P1 digestion revealed several minor adducts, one of which has been identified in the present study. Postlabeling analyses following nuclease P1 digestion of the products obtained from the reaction of N-acetoxy-4-aminobiphenyl with deoxyguanosine-3'-monophosphate (dGp) demonstrated the presence of this minor adduct. The 3'-monophosphate derivative of the adduct was subsequently chromatographically purified and subjected to spectroscopic analyses. Based on proton NMR and mass spectroscopic analyses of the synthetic product, the chemical structure of the adduct has been identified as N-(deoxyguanosin-N(2)-yl)-4 azobiphenyl (dG-N==N-ABP). (32)P-Postlabeling analysis of the nuclease P1 enriched DNA hydrolysate of HUCs treated with N-OH-ABP or N-hydroxy-4 acetylaminobiphenyl (N-OH-AABP) showed the presence of the dG-N==N-ABP adduct. It was also detected in calf thymus DNA incubated with HUC cytosol and N-OH-ABP in the presence of acetyl-CoA, or incubated with HUC microsomes and N-OH-AABP. These results demonstrate that in the target cells for ABP carcinogenesis in vivo, N-OH ABP and N-OH-AABP are bioactivated by acyltransferases to reactive arylnitrenium ions that covalently interact at the N2 position of deoxyguanosine in DNA. PMID- 12112384 TI - Induction of genotoxic damage is not correlated with the ability to methylate arsenite in vitro in the leukocytes of four mammalian species. AB - Arsenic is a natural drinking water contaminant that impacts the health of large populations of people throughout the world; however, the mode or mechanism by which arsenic induces cancer is unclear. In a series of in vitro studies, we exposed leukocytes from humans, mice, rats, and guinea pigs to a range of sodium arsenite concentrations to determine whether the lymphocytes from these species showed differential sensitivity to the induction of micronuclei (MN) assessed in cytochalasin B-induced binucleate cells. We also determined the capacity of the leukocytes to methylate arsenic by measuring the production of MMA [monomethylarsinic acid (MMA(V)) and monomethylarsonous acid (MMA(III))] and DMA [dimethylarsinic acid (DMA(V)) and dimethylarsonous acid (DMA(III))]. The results indicate that cells treated for 2 hr at the G(0) stage of the cell cycle with sodium arsenite showed only very small to negligible increases in MN after mitogenic stimulation. Treatment of actively cycling cells produced induction of MN with increasing arsenite concentration, with the human, rat, and mouse lymphocytes being much more sensitive to MN induction than those of the guinea pig. These data gave an excellent fit to a linear model. The leukocytes of all four species, including the guinea pig (a species previously thought not to methylate arsenic), were able to methylate arsenic, but there was no clear correlation between the ability to methylate arsenic and the induction of MN. PMID- 12112385 TI - Estimation of mutation rate at human glycophorin A locus in hematopoietic stem cell progenitors. AB - Surveys of human mutant cells exhibit a few individuals with relatively high "outlying" values, which might be explained by rare mutations occurring during development. To estimate how commonly this occurs, mutant red cell frequencies at the glycophorin A locus in 135 neonates and 109 children and adolescents from three research centers are compared with simulations in which mutations arise from successive cycles of binary fission. The simulations predict the data most accurately when the mutation rate in stem cell precursors is about 2-4 x 10(-7) per division cycle, which is similar to previous estimates from adult stem cell divisions. If these mutation rates are accurate, and the number of stem cell divisions during adult life is as low as previously estimated, it is predicted that up to one-sixth of mutant stem cells over a lifetime arose in early life. However, these mutant stem cells would be difficult to detect in surveys because their distribution within the general population is so skewed. PMID- 12112387 TI - Catalytic inhibitors of topoisomerase II are DNA-damaging agents: induction of chromosomal damage by merbarone and ICRF-187. AB - Merbarone is a catalytic inhibitor of topoisomerase II (topo II) that has been proposed to act primarily by blocking topo II-mediated DNA cleavage without stabilizing DNA-topo II-cleavable complexes. In this study merbarone was used as a model compound to investigate the genotoxic effects of catalytic inhibitors of topo II. The clastogenic properties of merbarone were evaluated using in vitro and in vivo micronucleus (MN) assays combined with CREST staining. For the in vitro MN assay, ICRF-187, a different type of catalytic inhibitor, and etoposide, a topo II poison, were used for comparison. Treatment of TK6 cells with all three of these drugs resulted in highly significant dose-related increases in kinetochore-lacking MN and, to a lesser extent, kinetochore-containing MN. In addition, a good correlation between p53 accumulation and MN formation was seen in the drug-treated cells. A mouse MN assay was performed to confirm that similar DNA-damaging effects would occur in vivo. Bone marrow smears from merbarone treated B6C3F1 mice showed a dose-related increase in micronucleated polychromatic erythrocytes with a mean of 26 MN per 1000 cells being seen at the 60 mg/kg dose. Almost all MN lacked a kinetochore signal, indicating that merbarone was predominantly clastogenic under these conditions in vivo. The present study clearly shows that merbarone is genotoxic both in vitro and in vivo, and demonstrates the inaccuracy of earlier statements that merbarone and other catalytic inhibitors block the enzymatic activity of topo II without damaging DNA. PMID- 12112386 TI - Mutagenicity of gamma-radiation, mitomycin C, and etoposide in the Hprt and Tk genes of Tk(+/-) mice. AB - The recently developed Tk(+/-) mouse detects in vivo somatic cell mutation in the endogenous, autosomal Tk gene. To evaluate the sensitivity of this model, we have treated Tk(+/-) mice with three agents that induce DNA damage by different mechanisms, and determined spleen lymphocyte mutant frequencies (MFs) in the autosomal Tk gene and in the X-linked Hprt gene. gamma-Radiation, which produces single- and double-strand breaks by nonspecific oxidative stress, efficiently increased Hprt MF, but not Tk MF. Mitomycin C, which produces bulky DNA monoadducts and crosslinks, was mutagenic in both the Hprt and Tk genes, but the response was greater in the Tk gene. An inhibitor of the ligase function of DNA topoisomerase II, etoposide, did not increase Hprt MF, and induced a small, but nonsignificant increase in Tk MF. Combined with previous data, the results indicate that the two genes are differentially sensitive to many agents, and that the Tk gene is more sensitive than the Hprt gene to some, but not all types of DNA damage. PMID- 12112388 TI - Activities of erythrocyte antioxidant enzymes in cancer patients. AB - We measured the activities of erythrocyte superoxide dismutase (SOD) and glutathione peroxidase (GPX) before therapy in 97 patients with cancer in various sites (gastrointestinal tract (GIT) (n=40), breast (n=30), and others (n=27)), and in 60 matched controls to assess antioxidant enzyme protection. Hemolysate hemoglobin (Hb) was measured spectrophotometrically. The activity of SOD (U/g Hb) was significantly lower in all sites (when all the cancer sites were considered as a group), GIT, breast, and other sites compared to the controls (P<0.0001, P<0.0001, P<0.0001 and P<0.0001, respectively). The activity of GPX (U/g Hb) was significantly decreased in all sites, GIT, and breast cancer sites than in the controls (P=0.024, P=0.033, and P=0.043, respectively). Age showed a weak negative correlation with enzyme activities in controls and patients. There was no significant association between SOD and GPX activities in either the controls or the patients. These results suggest that there may be a greater antioxidant burden for SOD than GPX in cancer, and that a weak association exists between the activities of the two enzymes in antioxidant protection. PMID- 12112389 TI - Bidirectional (positive/negative) interference of spironolactone, canrenone, and potassium canrenoate on serum digoxin measurement: elimination of interference by measuring free digoxin or using a chemiluminescent assay for digoxin. AB - Spironolactone and potassium canrenoate (aldosterone antagonist diuretics) are often used with digoxin in clinical practice. Spironolactone, potassium canrenoate, and their common metabolite canrenone cross-react with the fluorescence polarization immunoassay (FPIA) for digoxin, and can falsely elevate serum digoxin concentrations. Serum digoxin concentrations were falsely lowered when the microparticle enzyme immunoassay (MEIA) was used. Aliquots of drug-free serum were supplemented with therapeutic and above-therapeutic concentrations of spironolactone, canrenone, and potassium canrenoate, and apparent digoxin activities were measured. We observed digoxin-like activities in the FPIA, but observed no activity with the MEIA or the chemiluminescent assay (CLIA). However, when serum digoxin pools prepared from patients receiving digoxin were supplemented with these compounds, we observed suppression of total digoxin levels with the MEIA. In contrast, no interference was observed in the presence of these compounds when CLIA was used for digoxin measurement. These compounds are strongly protein-bound, and no apparent digoxin activity was observed in the protein-free ultrafiltrate when drug-free sera were spiked with high levels of these compounds. Taking advantage of strong protein binding of these compounds and weak protein binding of digoxin (25%), interference of spironolactone, canrenone, and potassium canrenoate in FPIA and MEIA digoxin assays can be mostly eliminated by monitoring free digoxin concentration. Another approach to avoid this interference is to use the CLIA digoxin assay. PMID- 12112390 TI - Decorin transfection in human mesangial cells downregulates genes playing a role in the progression of fibrosis. AB - The proteoglycan decorin inhibits TGF-beta; therefore, it could antagonize progression of fibrotic diseases associated with activation of TGF-beta(1). The effect of decorin transfection in human mesangial cells (HMCs) on the expression of genes related to kidney fibrosis was investigated. HMCs, isolated from glomeruli of healthy portions of human kidneys removed due to carcinoma, were histochemically typed. Decorin cDNA cloned in a eukaryotic expression vector was transfected into HMCs. Gene expression of fibrogenetic cytokines and fibrotic proteins TGF-beta(1), PDGF-beta, alpha(1) collagen type IV, alpha(1) collagen type I, fibronectin, and tenascin was analyzed, by reverse transcription polymerase chain reaction (RT-PCR), 24 hr after transfection. Immunoblotting analysis of protein extracts using anti-decorin IgG, revealed a positive signal of about 52 MDa, corresponding to the molecular weight of decorin, in cultures transfected with the decorin gene. Decorin mRNA increased about 12 times in cultures transfected with the construct pCR3.1-Deco. Cells with increased decorin synthesis showed a 61% decrease of TGF-beta(1) mRNA, a 71% reduction of alpha1 collagen type IV mRNA, and a 29% reduction of fibronectin mRNA. This study is the first to investigate decorin transfection into human mesangial cells, and supports the use of the decorin gene to control the progression of glomerular and interstitial fibrosis in kidney diseases. PMID- 12112391 TI - Superiority of a functional leukocyte adhesiveness/aggregation test over the white blood cell count to discriminate between mild and significant inflammatory response in patients with acute bacterial infections. AB - Electronic cell counters may underestimate the white blood cell count (WBCC) in the presence of aggregated leukocytes. In the present study we focused on the possibility of using a functional, as opposed to an anatomic, count to circumvent this eventual underestimation. A model of bacterial infection was used because of the importance of leukocytosis in the physician's clinical decision-making process. There were 35 patients with low C-reactive protein (CRP) concentrations (0.5-4.9 mg/dL), 45 with intermediate (5-9.9 mg/dL), and 120 with relatively high (>10 mg/dL) CRP concentrations. A significant (P=0.008) difference was noted between the state of leukocyte adhesiveness/aggregation in the peripheral blood of individuals with low CRP concentrations (3.5%+/-4.3%) and those with high CRP concentrations (7.4%+/-8%), while there was no significant difference in the respective number of WBCs per cubic millimeter (cmm) (11,600 +/- 5,500 and 14,000 +/- 7,200, respectively). We raise the possibility that a functional test might be superior over an anatomic count in patients with acute bacterial infection and a significant acute phase response. PMID- 12112393 TI - Human immunodeficiency virus genotype and hypertriglyceridemia. AB - Many HIV patients develop a progressive syndrome of abnormal body fat distribution accompanied by hypertriglyceridemia. Antiretroviral agents are thought to be etiologic in the syndrome, often termed "highly active antiretroviral therapy (HAART)-associated lipodystrophy." In the course of clinical HIV genotype testing, we observed that our HIV patients with hypertriglyceridemia had viral genotypes that were more highly mutated than those of our therapy-matched control patients. Hypertriglyceridemia was statistically associated with predicted resistance for three nucleoside reverse transcriptase inhibitors: zidovudine, abacavir, and stavudine. Statistical analysis of 51 patients in retrospect revealed a strong association of mutations at reverse transcriptase codons M41 and T215 with hypertriglyceridemia (chi-square (chi(2)) = 8.375, P=.0038; and chi(2)=7.445, P=.0064, respectively). This was in contrast to silent mutations, which occurred at equivalent rates in retroviral genotypes of patients with and without hypertriglyceridemia. The findings imply that the HIV genotype itself may be a significant etiologic factor in antiretroviral associated lipodystrophy. PMID- 12112392 TI - Anti-smooth muscle antibodies (ASMAs) and anti-cytoskeleton antibodies (ACTAs) in liver diseases: a comparison of classical indirect immunofluorescence with ELISA. AB - In the diagnosis of autoimmune hepatitis type I (AIH-I), the routine assay of indirect immunofluorescence (IFL), used for the detection of anti-smooth muscle antibodies (ASMAs), has a low predictive value. On the other hand, the enzyme linked immunosorbent assay (ELISA), which detects anti-cytoskeleton antibodies (ACTAs), presents contradictory results concerning their specific antigenic target. In this study, we first looked for the immunological properties (isotypes and antigenic targets) of autoantibodies in AIH-I and two other control liver diseases: primary biliary cirrhosis (PBC) and viral hepatitis (VH), using ELISA based on cytoskeleton proteins: F-actin, G-actin, myosin, tropomyosin, troponin, desmin, vimentin, keratin, and an extract of HEp-2 carcinoma cells. We also compared the diagnostic value of IFL and ELISA. In contrast to previous studies, we found that actin was not specific for AIH-I. No autoantigen and no antibody class or subclass discriminated AIH-I from the control diseases. IFL is more suitable for AIH-I diagnosis, as 97% of AIH-I sera but only 22% of PBC sera were ASMA-positive. Additionally, 96% of ASMA-positive, and all ASMA-negative sera from all three liver diseases were ACTA-positive. ASMA were mainly IgG, while >50% of ACTA also contained IgA and IgM. These data suggest that ACTAs recognize additional epitopes as compared to ASMAs, and they frequently occur in all liver diseases. PMID- 12112394 TI - Chronic cocaine increases kappa-opioid receptor density: lack of effect by selective dopamine uptake inhibitors. AB - Continuous infusion of cocaine or the selective dopamine uptake inhibitors GBR 12909 or RTI-117 increases locomotor stimulation, to which partial tolerance occurs. In addition, all three drugs produce significant decreases in tyrosine hydroxylase immunoreactivity in caudate putamen and nucleus accumbens core, suggesting a decreased dopaminergic tone. An interaction between cocaine and opioids has long been documented. Chronic cocaine significantly increases mu and kappa-opioid receptors and treatment with a kappa-opioid agonist markedly reduces the behavioral effects of cocaine. In addition, chronic cocaine, but not GBR 12909, increases prodynorphin gene expression in caudate putamen. To further understand the interaction between cocaine and the kappa-opioid system, the effects of a chronic continuous infusion for 14 days of cocaine or one of the selective dopamine uptake inhibitors GBR 12909 or RTI-117 via osmotic minipump were examined on kappa-opioid receptors using the selective kappa-opioid ligand [3H] U-69593. [3H] U-69593 binding density was significantly increased in caudate putamen, nucleus accumbens shell, claustrum, and endopiriform nucleus after cocaine, while neither GBR 12909 nor RTI-117 had any effect. The increased kappa opioid receptor densities observed following cocaine are likely not related to dopamine uptake inhibition, since they were not produced by selective dopamine uptake inhibitors. These findings suggest that regulation of kappa-opioid receptors by cocaine may be via inhibition of serotonin or norepinephrine uptake, by a combination of effects on two or three monoamine transporters, or by a mechanism unrelated to transporter inhibition. PMID- 12112395 TI - Repeated ventral tegmental area amphetamine administration alters dopamine D1 receptor signaling in the nucleus accumbens. AB - Neuroadaptations of the mesoaccumbens dopamine (DA) system likely underlie the emergence of locomotor sensitization following the repeated intermittent systemic administration of amphetamine (AMPH). In the nucleus accumbens (NAc), such neuroadaptations include enhanced DA overflow in response to a subsequent AMPH challenge as well as increased sensitivity to the inhibitory effects of D1 DA receptor (D1R) activation and an altered profile of D1R-dependent induction of immediate early genes (IEGs). Previous results indicate that AMPH acts in the ventral tegmental area (VTA) to initiate those changes leading to sensitization of the locomotor activity and NAc DA overflow produced by systemic administration of this drug. These observations are intriguing, given that acute infusion of AMPH into the VTA does not stimulate locomotor activity or, as we report presently, increase extracellular NAc DA concentrations. Two experiments, therefore, assessed the ability of repeated VTA AMPH to produce adaptations in D1R signaling in the NAc. Rats were administered three bilateral VTA infusions of saline or AMPH (2.5 microg/0.5 microl/side, one every third day). In the first experiment, in vivo extracellular electrophysiological recordings revealed that previous exposure to VTA AMPH enhanced the sensitivity of NAc neurons to the inhibitory effects of iontophoretic application of the D1R agonist SKF 38393. This effect was observed early (2-3 days) and at 1 month of withdrawal, but not after 2 months. Similarly, in the second experiment it was found that the D1R dependent induction by AMPH of Fos, FosB, and JunB, but not NGFI-A, in the NAc was enhanced in rats exposed 1 week earlier to repeated VTA AMPH. These findings indicate that repeated VTA AMPH administration initiates relatively long-lasting adaptations in D1R signaling in the NAc that may, together with presynaptic adaptations affecting DA overflow, contribute to the expression of locomotor sensitization by this drug. PMID- 12112396 TI - Preferential action of gabapentin and pregabalin at P/Q-type voltage-sensitive calcium channels: inhibition of K+-evoked [3H]-norepinephrine release from rat neocortical slices. AB - Gabapentin (GBP; Neurontin) and pregabalin (PGB; CI-1008), efficacious drugs in several neurological and psychiatric disorders, inhibit neurotransmitter release from mammalian brain slices at therapeutically relevant concentrations. A detailed investigation, exploring the basis for this in vitro phenomenon, focused on norepinephrine (NE) and rat neocortical tissue in complementary assays of neurotransmitter release and radioligand binding. The results are consistent with the hypothesis that GBP, PGB, and related substances decrease neocortical NE release by acting at the alpha2delta subunit of presynaptic P/Q-type voltage sensitive Ca2+ channels (VSCC) subserving Ca2+ influx in noradrenergic terminals. The inhibitory action appears competitive with [Ca2+]o and preferential to those neurons undergoing prolonged depolarization. Other results indicate that the reduction of exocytotic NE release is independent of L- and N-type VSCC, classical drug/peptide binding sites on VSCC, Na+ channels, alpha2-adrenoceptors, NE transporter, and system L amino acid transporter. These findings suggest a selective modulation of P/Q-type VSCC that are implicated in neurotransmission and several GBP-responsive pathologies. PMID- 12112397 TI - 5-HT6 receptor binding sites in schizophrenia and following antipsychotic drug administration: autoradiographic studies with [125I]SB-258585. AB - The 5-hydroxytryptamine (5-HT; serotonin)-6 receptor (5-HT6R) is a putative target of atypical antipsychotic drugs and its mRNA expression is altered in schizophrenia. [125I]SB-258585 is a selective 5-HT6R antagonist which has been well characterized for use in the rat brain. The present study evaluated its suitability for receptor autoradiography in the human brain and its application to quantitative studies. The affinity (K(d) approximately 1.2 nM) and relative distribution of binding sites (striatum >> cortex approximately hippocampus) were similar to the rat. The distribution of [125I]SB-258585 binding in these regions was also consistent with that of 5-HT6R mRNA, determined in parallel using in situ hybridization. [125I]SB-258585 binding site densities were measured in dorsolateral prefrontal cortex of 20 patients with chronic schizophrenia and compared with 17 normal subjects. No differences were seen between groups. Neither were [125I]SB-258585 binding site densities affected in the frontal cortex or striatum of rats following 2 weeks' administration of the antipsychotic drugs haloperidol, chlorpromazine, olanzapine, risperidone, or clozapine. In summary, [125I]SB-258585 is a suitable radioligand for studies of human brain 5 HT6R binding sites and shows that their distribution is broadly similar to that of the rodent. The lack of effect of schizophrenia or antipsychotic drug administration on [125I]SB-258585 binding suggests that an altered receptor density does not contribute to any involvement which the 5-HT6R may have in the disease or its treatment. PMID- 12112398 TI - Inhibition of autonomic nerve-mediated inotropic responses in guinea pig atrium by bafilomycin A. AB - Neurosecretory vesicles actively accumulate neurotransmitter by consuming proton motive force generated by vacuolar H+-ATPase (V-ATPase). The effects of bafilomycin A, a macrolide antibiotic that inactivates V-ATPase, on nerve stimulation-mediated inotropic responses of the left atrium were studied to explore the role of the enzyme in the cholinergic and adrenergic neurotransmissions. On field stimulation, the contractility of paced atrium exhibited initial atropine-sensitive depression followed by propranolol-sensitive facilitation. Both the negative and positive inotropic effects were abolished by bafilomycin A. The inhibitions were irreversible and followed a similar time course and the inhibitory effects were accelerated by intense nerve stimulation. In contrast, bafilomycin A had no effect on the inotropic responses produced by muscarinic acetylcholine or alpha-adrenergic receptor agonist. Stimulation of neuronal nicotinic acetylcholine receptor also elicited biphasic changes of contractile force, which were depressed by bafilomycin A. Compared with the inhibitory effects on field stimulation, the depressions progressed slowly and incompletely. The results suggest that inhibition of V-ATPase depressed the synaptic transmissions at autonomic nerve-muscle junctions. Furthermore, bafilomycin A preferentially inhibited neurotransmitter release emanating from the immediately releasable pool. PMID- 12112399 TI - Cocaine increases serotonergic activity in the hippocampus and nucleus accumbens in vivo: 5-HT1a-receptor antagonism blocks behavioral but potentiates serotonergic activation. AB - The hippocampus is an important mediator of learning and reinforcement, but its role in cocaine effects has received little attention. Neuronal activity in the hippocampus and the nucleus accumbens (Nac) depend on serotonergic (5-HT) transmission. Here we describe for the first time a cocaine-induced increase in 5 HT concentration in the hippocampus and the Nac parallel to behavioral activation. In addition, pretreatment with the 5-HT(1A)-receptor antagonist WAY 100635 blocked the behavioral activation after cocaine while potentiating the 5 HT increase in the hippocampus and the Nac. In vivo microdialysis was used in behaving rats to measure extracellular concentration of 5-HT in the hippocampus and the Nac. Four groups of animals received one of the following drug combinations: WAY 100635 (0.4 mg/kg) and cocaine (10 mg/kg), saline and cocaine (10 mg/kg), WAY 100635 (0.4 mg/kg) and saline, or saline and saline. The injections were administered i.p. and spaced 30 min apart. It was found that 1.) cocaine, at a dose that activates behavior, increases 5-HT levels in the hippocampus and in the Nac, and 2.) 5-HT(1A)-receptor antagonism can cause a dissociation of the hippocampal and Nac 5-HT activity from behavioral activation after cocaine. These results are discussed within the framework of the hippocampal-accumbens projection and its contribution to behavioral activity. They suggest that the hippocampus may have a role in mediating the behavioral and neurochemical effects of cocaine. PMID- 12112400 TI - Dopaminergic synapses in the matrix of the ventrolateral striatum after chronic haloperidol treatment. AB - Antipsychotic drugs (APD) are used in the treatment of schizophrenia and other psychotic disorders and exert their effects, in part, through dopamine receptor blockade. APD treatment causes many changes in the brains of humans and experimental animals including therapeutic, pathologic, or changes associated with motor side effects. Typical APD given chronically to animals induce behavioral sequelae that mimic tardive dyskinesia in several ways. Our previous work has shown that chronic treatment with haloperidol decreases striatal synaptic density but that symmetric synapses are lost only in rats that develop oral dyskinesias. The goals of this study were to determine if the density of dopaminergic terminals was affected by chronic haloperidol treatment and/or correlated with dyskinesias. Rats were given haloperidol (1.5 mg/kg/rat) or water, as a control. After 6 months of treatment, rats were divided into nondyskinetic or dyskinetic groups according to the behavior scores determined in the last month. Striatal volume was similar between controls and drug-treated rats. Synaptic density, calculated using stereological methods, was obtained from the matrix of the ventrolateral striatum. The density of symmetric synapses (mean +/- SD, per 100/microm(3)) formed by tyrosine hydroxylase (TH) containing terminals in haloperidol treated rats (3.58 +/- 1.64) was not significantly different from that of controls (3.06 +/- 1.00). The density of TH-labeled terminals forming symmetric synapses in the nondyskinetic group (3.65 +/- 1.67) vs. the dyskinetic group (3.54 +/- 1.73) was similar and neither was different from that of the controls. These data indicate that terminals other than dopaminergic ones form fewer symmetric synapses in dyskinetic rats. Moreover, these data have implications for interpreting results obtained in humans treated with typical antipsychotic drugs. PMID- 12112401 TI - Gabaergic modulation of the stress response in frontal cortex and amygdala. AB - GABAergic neurotransmission is thought to play an important role in the modulation of the central response to stress. In the present study we evaluate the influence of a brief restraint exposure on GABA-stimulated chloride influx in diverse brain areas presumed to have a major role in the mediation of emotional behaviors following aversive stimulation. A reduced chloride uptake after stress exposure was only observed in frontal cortex and amygdala. Moreover, rats subjected to such stressor performed an anxiogenic behavior when exposed later to the elevated plus-maze. A comparable behavior in the elevated plus-maze was observed between animals that were allowed to chew during the restraint experience and those without any stressful manipulation, suggesting that chewing served as an efficient coping behavioral strategy during such threatening situations. In order to explore if chewing during the restraint experience could suppress the reduction in GABA-stimulated chloride uptake induced by this stressor, rats were allowed or not to chew during restraint and in both cases GABA-stimulated chloride influx was assayed in frontal cortex and amygdala. The finding of this experiment showed that restrained rats that have the possibility to chew exhibited a similar GABA-stimulated chloride uptake in cortical tissue to that shown by control, unstressed rats. Moreover, chewing in response to restraint attenuated the reduction of GABA-stimulated chloride uptake in amygdala, supporting the notion that chewing is an effective coping response to restraint. These experiments suggest that a reduced GABAergic inhibitory control in these areas could be implicated in the emotional sequelae generated by this uncontrollable stressor and that the suppression of this reduction seems to be associated with the occurrence of coping behavioral response to such fear inducing stimulus. PMID- 12112402 TI - Pharmacological evaluation of a Br-76 analog of epibatidine: a potent ligand for studying brain nicotinic acetylcholine receptors. AB - [(76)Br]-Norchlorobromoepibatidine ([(76)Br]BrPH) is a specific and high affinity radioligand for the nicotinic acetylcholine receptors (nAChRs). In vitro, on rat thalamus membranes [(76)Br]BrPH bound to two sites with apparent affinities of 8 pM and 3 nM. The density of binding sites were 1.9 and 70 fmol/mg protein, respectively. In vivo, in biodistribution and autoradiographic studies in rats the regional distribution of [(76)Br]BrPH paralleled the neuroanatomical localization of nAChRs. Two hours postinjection, the highest concentration in the brain was found in thalamus and colliculi (4% ID/g). Competition experiments with specific nicotinic, muscarinic, dopaminergic, and serotoninergic drugs confirmed that the in vivo binding of [(76)Br]BrPH was consistent with neuronal nicotinic receptors. PET imaging of [(76)Br]BrPH in baboon demonstrated a rapid and high uptake in the brain. Peak uptake occurred at 30-40 min for the thalamus. Due to the constant washout in the cerebellum, the thalamus to cerebellum ratio was 5 at 2 h postinjection. Subcutaneous injection of cytisine (1 mg/kg), 3 h postinjection of [(76)Br]BrPH reduced the radioactivity concentration in thalamus and cortex by 58 and 50%, respectively, as observed 1 h later. Cytisine pretreatment (5 mg/kg s.c.) inhibited completely the radioligand accumulation in the thalamus. Chronic MPTP pretreatment resulted in reduction of [(76)Br]BrPH uptake in all brain regions except in cerebellum. These preliminary results suggest that [(76)Br]BrPH has the potential to be a useful radioligand for studying the pharmacology of nicotinic acetylcholine receptors in preclinical experiments. PMID- 12112403 TI - Serotonin modulation of cerebral glucose metabolism measured with positron emission tomography (PET) in human subjects. AB - To develop a method to measure the dynamic response of the serotonin system in vivo, the effects of intravenously administered citalopram (the most selective of the serotonin reuptake inhibitors) on cerebral glucose metabolism were evaluated. Cerebral glucose metabolism was measured with positron emission tomography (PET) in 14 normal subjects scanned after administration of saline placebo and citalopram administered on 2 separate days. Citalopram administration resulted in a decrease in metabolism in the right anterior cingulate gyrus (BA 24/32), right superior (BA 9) and right middle frontal gyrus (BA 6), right parietal cortex (precuneus), right superior occipital gyrus, left thalamus, and right cerebellum. Increased metabolism was observed in the left superior temporal gyrus and left occipital cortex. Alterations in metabolism by acute citalopram administration involved the heteromodal association cortices that also show metabolic alterations in patients with geriatric depression and overlap with the regions affected by antidepressant treatment. Future studies will evaluate how the acute metabolic response to citalopram relates to the metabolic response after chronic treatment in patients with geriatric depression. PMID- 12112404 TI - Region-specific targeting of dopamine D2-receptors and somatodendritic vesicular monoamine transporter 2 (VMAT2) within ventral tegmental area subdivisions. AB - Throughout the ventral tegmental area (VTA), dopamine is packaged within subcellular organelles by the vesicular monoamine transporter-2 (VMAT2). Somatodendritically released dopamine in this region binds to the D2 receptor (D2R) to modulate ongoing neurotransmission. Although autoregulation of mesocortical dopaminergic neurons in the parabrachial VTA (PB-VTA) is known to be less efficacious than that of mesolimbic dopaminergic neurons in the paranigral (PN-VTA), the cellular basis for this regional heterogeneity is not known. For this reason, we used electron microscopic immunocytochemistry to determine the subcellular localization of the dopamine storage vesicles (identified by the presence of VMAT2) in relation to the D2R in these VTA subdivisions. In both regions, D2R immunoreactivity was principally located on extrasynaptic dendritic plasma membranes near excitatory-type synapses. Equivalent percentages (72 and 74%) of the D2R-labeled dendrites in each region contained VMAT2-immunoreactive tubulovesicles. Of the total VMAT2-labeled dendrites, however, a significantly lower percentage in the PB-VTA (26%) than in the PN-VTA (38%) contained D2R labeling. In contrast, a significantly higher number of VMAT2 immunogold-silver deposits was seen within individual dendrites in the PB-VTA than in PN-VTA. In both regions, D2R immunoreactivity was also detected in VMAT2-negative axon terminals that formed synapses on dendrites containing VMAT2. Our results are the first to demonstrate that within VTA neurons and their afferents the D2R is strategically positioned for activation by dopamine released from dendritic storage vesicles. These findings also suggest that the potential for D2R activation may affect the expression levels of VMAT2 in VTA dendrites. PMID- 12112405 TI - Parametric mapping of [18F]FPCIT binding in early stage Parkinson's disease: a PET study. AB - We have shown that fluorinated N-3-fluoropropyl-2-beta-carboxymethoxy-3-beta-(4 iodophenyl) nortropane ([(18)F]FPCIT) and PET offer a valuable means of quantifying regional abnormality in dopamine transporter (DAT) imaging associated with Parkinson's disease (PD). The objective of this study was to delineate the topographic distribution of DAT binding in early stage idiopathic PD using statistical parametric analysis of [(18)F]FPCIT PET data. We performed dynamic PET studies in 15 hemi-parkinsonian (Hoehn & Yahr I) patients and 10 age-matched normal volunteers over 100 min and calculated images of [(18)F]FPCIT binding ratios on a pixel-by-pixel basis. Statistical parametric mapping (SPM) was then used to localize binding reductions in PD and to compute the absolute change relative to normal. [(18)F]FPCIT binding decreased significantly in the contralateral posterior putamen of the PD group (P < 0.001, corrected). A significant reduction was also seen in the ipsilateral putamen, which was smaller in extent but localized more posteriorly. A quantitative comparison of DAT binding in the two clusters showed that the onset of motor symptoms in PD was associated with an approximate 70% loss relative to the normal mean in the contralateral posterior putamen. These results suggest that SPM analysis of [(18)F]FPCIT PET data can be used to quantify and map abnormalities in DAT activity within the human striatum. This method provides a useful tool to track the onset and progression of PD at its earliest stages. PMID- 12112406 TI - Dose-dependent effects of Delta9-tetrahydrocannabinol on rates of local cerebral glucose utilization in rat. AB - Recent reports have demonstrated that Delta(9)-tetrahydrocannabinol (Delta(9) THC) stimulates locomotor activity at low doses (<2.5 mg/kg), while higher doses (>2.5 mg/kg) produce decreases in spontaneous activity. Using quantitative 2 [(14)C]deoxyglucose (2-DG) autoradiography, we systematically studied the effects of acute Delta(9)-THC on rates of local cerebral glucose utilization. The first series of experiments was designed to determine if Delta(9)-THC-mediated changes in cerebral metabolism followed a clear dose-response relationship. Adult male Sprague-Dawley rats were treated with either vehicle or Delta(9)-THC (0.25-2.5 mg/kg) and the 2-DG procedure was initiated 15 min following exposure. Administration of 2.5 mg/kg Delta(9)-THC produced significant decreases in cerebral metabolism in most brain regions studied. In contrast, administration of 0.25 mg/kg Delta(9)-THC produced no significant alterations in any brain region studied, while 1.0 mg/kg of Delta(9)-THC produced a restricted pattern of metabolic decreases. Significant decreases in metabolism following 1.0 mg/kg were concentrated in structures subserving limbic and sensory functions. In a second series of experiments, the effects of pretreatment with the cannabinoid receptor antagonist SR141716A (1.0 mg/kg) on Delta(9)-THC-induced changes in functional activity were measured. Pretreatment with SR141716A attenuated the majority of functional changes produced by Delta(9)-THC, suggesting that these effects are primarily mediated by central cannabinoid receptors. Moreover, these findings indicate that the effects of Delta(9)-THC on cerebral metabolism are dose dependent and that there are regional differences in the metabolic response to acute cannabinoid exposure. PMID- 12112407 TI - Serotonin mediates CA1 spine density but is not crucial for ovarian steroid regulation of synaptic plasticity in the adult rat dorsal hippocampus. AB - The activity of the serotonin (5-hydroxytryptamine, 5-HT) system is sensitive to estradiol and progesterone. During the ovarian cycle, dendritic spines on CA1 pyramidal neurons of the dorsal hippocampus are increased by estradiol and later decreased by progesterone. We sought to determine whether 5-HT is involved in maintaining CA1 spine density and/or in steroid regulation of synaptic plasticity in dorsal hippocampus. Ovariectomized rats were treated (sc) over 10 days with the tryptophan hydroxylase inhibitor parachlorophenylalanine (pCPA) to deplete 5 HT, followed by estradiol benzoate on days 10 and 11. A subset of animals received progesterone on day 12. The day after the last treatment, rats were perfused and brains were processed for Golgi impregnation. Separate groups were processed for radioimmunocytochemistry (RICC) for the spine-associated protein, spinophilin, or high-performance liquid chromatography (HPLC) for monoamine analysis. Golgi and RICC data indicate that CA1 apical spine density was significantly decreased by pCPA (17-20%). Estradiol increased spine density in both saline- and pCPA-treated rats compared to respective controls (30%); however, pCPA animals maintained significantly fewer spines. No differences in spine densities were observed between saline- and pCPA-treated rats given estradiol and progesterone. Depletion of 5-HT by pCPA was confirmed in the CA1 ( 90%) and dorsal raphe (-80%) by HPLC analysis. While 5-HT depletion was associated with a 57% decrease in CA1 norepinephrine (NE), there was no difference in dorsal raphe NE. Thus, whereas 5-HT is involved in maintaining spine density in the adult female rat CA1, it is not crucial for steroid-mediated plasticity. 5-HT-regulated spines/synapses may represent distinct populations from those modulated by estradiol and progesterone in dorsal hippocampus. PMID- 12112408 TI - PET imaging of brain acetylcholinesterase using [11C]CP-126,998, a brain selective enzyme inhibitor. AB - PET and [(11)C]CP-126,998, an N-benzylpiperidinebenzisoxazole, were used to image brain acetylcholinesterase (AChE) distribution in healthy controls before and after administration of 5 mg donepezil p.o., a reversible AChE inhibitor. Logan plots were used to compute distribution volumes (V(T)). The V(T) of [(11)C]CP 126,998 was highest in the basal ganglia and cerebellum and lowest in the cerebral cortex, thalamus, amygdala, and hippocampus. The regional V(T) values correlated well with AChE concentration measured in vitro. Donepezil, given 4 h before PET scanning, induced a substantial inhibition of [(11)C]CP-126,998 binding (43-62%) in all brain regions when compared to the baseline PET study. The results of this study indicate that PET imaging of [(11)C]CP-126,998 may be useful in quantifying the distribution of regional brain AChE. This new PET radiotracer may potentially be employed in the diagnosis and treatment of patients with disorders of cholinergic neurotransmission, such as Alzheimer's disease. PMID- 12112409 TI - Differentiation of spinous synapses in the mouse organ of corti. AB - The inner hair cells, the primary auditory receptors, are perceived only as a means for transfer of sound signals via the auditory nerve to the central nervous system. During initial synaptogenesis, they receive relatively few and mainly somatic synapses. However, around the onset of hearing (10-14 postnatal days in the mouse), a complex network of local spinous synapses differentiates, involving inner hair cells, their afferent dendrites, and lateral olivocochlear terminals. Inner hair cell spines participate in triadic synapses between olivocochlear terminals and afferent dendrites. Triadic synapses have not yet been confirmed in the adult. Synaptic spines of afferent dendrites form axodendritic synapses with olivocochlear terminals and somatodendritic synapses with inner hair cells. The latter are of two types: ribbon-dendritic spines and stout dendritic spines surrounded only by a crown of synaptic vesicles. Formation of spinous afferent synapses results from sprouting of dendritic filopodia that intussuscept inner hair cell cytoplasm. This process continues in the adult, indicating ongoing synaptogenesis. Spinous processes of olivocochlear synaptic terminals contact adjacent afferent dendrites, thus integrating their connectivity. They develop about 14 postnatal days, but their presence in the adult has yet to be confirmed. Differentiation of spinous synapses in the organ of Corti results in a total increase of synaptic contacts and in a complexity of synaptic arrangements and connectivity. We propose that spinous synapses provide the morphological substrate for local processing of initial auditory signals within the cochlea. PMID- 12112410 TI - Mu-receptor agonism with alfentanil increases striatal dopamine D2 receptor binding in man. AB - Animal studies indicate that mu-opioids indirectly modulate neurotransmission in the nigrostriatal dopaminergic pathway. We used positron emission tomography (PET) to study the effects of alfentanil (a mu-opioid receptor agonist) on striatal dopamine D2 receptor binding in eight healthy male volunteers. D2 receptor binding was determined by using [(11)C]raclopride as radioligand. Each subject underwent two PET sessions on the same day, the first without the drug (control) and the second during alfentanil infusion. Alfentanil was administered as target-controlled infusion to maintain pseudo steady-state plasma concentration of 80 ng/ml throughout the PET session. A freeze lesion model was used for pain testing at the end of both PET sessions. A mechanical pain stimulus of 5 N was rated by the subjects using a visual analog scale. Regions of interest for the putamen, caudate nucleus, and cerebellum were drawn on MRI images and transferred to PET images. Alfentanil increased the binding potential of [(11)C]raclopride in the putamen by 6.0% (P = 0.04) and in the caudate nucleus by 7.4% (P = 0.008). Alfentanil caused a small reduction in respiratory rate (P = 0.046) and oxygen saturation (P < 0.001), and a moderate consistent increase in end-tidal CO(2) (P < 0.001). Pain scores were significantly smaller after alfentanil PET scan (median VAS 9 (0-42) vs. 23.5 (15-52), P = 0.008). These results indicate that pharmacologically relevant concentrations of alfentanil increase D2 dopamine receptor binding in the striatum in man. This increase is assumed to reflect reduced dopamine release. PMID- 12112411 TI - (R)-N-[11C]methyl-3-pyrrolidyl benzilate, a high-affinity reversible radioligand for PET studies of the muscarinic acetylcholine receptor. AB - We recently reported the synthesis and binding affinity of ligands for the muscarinic acetylcholine receptor (mAChR) based on both the pyrrolidyl and piperidyl benzilate scaffold. One of these, (R)-3-pyrrolidyl benzilate, was successfully radiolabeled with [(11)C]methyl triflate and the resulting compound, (R)-N-[(11)C]methyl-3-pyrrolidyl benzilate (3-[(11)C]NMPYB), was evaluated as a reversible, acetylcholine-sensitive tracer for the mAChR (K(i) of unlabeled 3 NMPYB is 0.72 nM). This compound displayed high, receptor-mediated retention in regions of the mouse and rat brain known to have high concentrations of mAChRs. Moreover, bolus studies in a pigtail monkey showed that this compound had superior clearance from the brain when compared to muscarinic radiotracers previously employed in human PET studies. Infusion studies in the same monkey revealed that it was possible to achieve equilibrium of radiotracer distribution for 3-[(11)C]NMPYB in both the striatum and cortex. Sensitivity to endogenous acetylcholine levels was evaluated by injecting phenserine (5 mg/kg) into rats prior to administration of 3-[(11)C]NMPYB in an equilibrium infusion protocol. This pretreatment produced a modest, statistically significant decrease (9-11%) in the distribution volume ratios for muscarinic receptor rich regions of the rat brain as compared to controls. PMID- 12112412 TI - Age-related changes in the striatal dopaminergic system in the living brain: a multiparametric PET study in conscious monkeys. AB - In the present study, age-related changes in the striatal dopaminergic system were examined in the living brains of conscious young (6.2 +/- 1.5 years old) and aged (20.2 +/- 2.6 years old) monkeys (Macaca mulatta) using positron emission tomography (PET). L-[beta-(11)C]DOPA and [(11)C]beta-CFT were applied to determine dopamine presynaptic functions such as synthesis rate and transporter (DAT) availability, respectively. Striatal dopamine D(1)- (D(1)R) and D(2)-like receptor (D(2)R) binding were measured with [(11)C]SCH23390 and [(11)C]raclopride, respectively. Although the markers of presynaptic terminals showed parallel age-related declines, the reduction of dopamine synthesis rate measured with L-[beta-(11)C]DOPA was slightly smaller than that of DAT determined with [(11)C]beta-CFT. The binding of [(11)C]raclopride to D(2)R in vivo was significantly reduced with aging, while that of [(11)C]SCH23390 to D(1)R showed no such marked age-related reduction. When the DAT inhibitor GBR12909 (0.5 and 5 mg/kg) was administered, DAT availability, dopamine synthesis, and D(2)R binding were significantly decreased in a dose-dependent manner in both age groups; however, the degrees of the decreases in these parameters were significantly higher in young rather than in aged animals. Dopamine concentration in the striatal extracellular fluid (ECF), as measured by microdialysis, was increased by administration of GBR12909 in a dose-dependent manner and the degree of the increase in dopamine level decreased with age. These results demonstrate that age related changes of dopamine neuronal functions were not limited to the resting condition but were also seen in the functional responses to the neurotransmitter modulation. PMID- 12112413 TI - Conditioned-fear stress increases Fos expression in monoaminergic and GABAergic neurons of the locus coeruleus and dorsal raphe nuclei. AB - Many studies have demonstrated that physical or psychological stress can increase Fos expression in brainstem monoaminergic nuclei. Little is known, however, about the extent to which stress increases the expression of Fos in monoaminergic and nonmonoaminergic neurons in the brainstem. We examined the effects of conditioned fear (CF) stress following mild footshock (FS) as unconditioned stress on Fos expression in the monoaminergic and GABAergic neurons of the ventral tegmental area (VTA), locus coeruleus (LC), and dorsal raphe nucleus (DR) in rats. The CF stress significantly increased the number of Fos-positive (Fos+) cells in both the LC and DR, whereas it did not increase the number in the VTA. Using a double labeling technique, we combined Fos immunostaining with that for tyrosine hydroxylase (TH), serotonin (5-HT), or GABA for histochemical identification of the CF stress-induced Fos+ neurons. The percentage of TH/Fos double-labeled cells resulting from CF stress was 63% of the Fos+ cells in the LC, whereas 52% of the Fos+ cells contained 5-HT in the DR. We also found that approximately 60% of the CF stress-induced Fos+ cells were GABAergic neurons in these brain regions. These results indicate that CF stress induces intense Fos expression in the noradrenergic LC and serotonergic DR neurons, but not in the dopaminergic VTA neurons. They also indicate that not only monoaminergic neurons but also GABAergic neurons within the LC and DR are activated by the stress. PMID- 12112414 TI - Dopamine transporter-dependent induction of C-Fos in HEK cells. AB - The psychostimulants cocaine and amphetamine increase expression of the immediate early gene (IEG) c-fos indirectly, via D1 dopamine receptor activation. To determine whether dopamine transporter substrates and inhibitors can affect c-Fos expression directly, we investigated their effects on c-Fos protein and c-fos mRNA in HEK-293 (HEK) cells transfected with the human dopamine transporter (hDAT). In untransfected HEK cells, methylphenidate and cocaine produced a small but statistically significant increase in c-Fos, whereas dopamine and amphetamine did not. In hDAT cells, DAT substrates (dopamine, amphetamine) increased c-Fos immunoreactivity 6- and 3-fold (respectively). The DAT inhibitors cocaine, methylphenidate, and bupropion also increased c-Fos approximately 3-fold in hDAT cells. If coincubated with dopamine, the inhibitors attenuated dopamine-induced c Fos in hDAT cells. The magnitude of c-fos mRNA induction by substrates and inhibitors paralleled induction of c-Fos protein immunoreactivity. The results indicate that substrates or inhibitors of the DAT can trigger induction of IEG expression in the absence of D1 dopamine receptor. For substrates, IEG induction is DAT-dependent, but for certain DAT inhibitors the cellular response can be elicited in the absence of the DAT in HEK cells. Oxidative stress may partly, but not fully, account for the DA-induced c-Fos induction as an inhibitor of oxidative stress Trolox C, attenuated DA-induced c-Fos induction. Protein kinase C (PKC) may also partially account for c-Fos induction as a specific inhibitor of PKC Bisindolylmaleimide I (BIS) attenuated DA-induced c-Fos by 50%. DAT substrate and inhibitor effects on IEGs, other fos-related antigens, and possible mechanisms that contribute to c-Fos induction warrant investigation in presynaptic neurons as a potential contribution to the long-term effects of psychostimulants. PMID- 12112416 TI - Immune response by host after allogeneic chondrocyte transplant to the cartilage. AB - Chondrocytes constitutively express class I and, in some species, class II major histocompatibility complex (MHC). It is also possible that they possess specific differentiation antigen(s). Furthermore, lymphocytic cells, corresponding to NK cells, display spontaneous cytotoxic activity against chondrocytes. Studies on articular cartilage repair by transplants of allogeneic chondrocytes were mainly done on non-inbred animals, such as rabbits and hens. Surprisingly, only in single instances these transplants were rejected. In inbred rats, allogeneic chondrocytes transplanted into full-thickness defects in articular cartilage immediately after isolation evoked systemic immunological reaction and produced cartilage was rejected. Combined immunosuppression with cyclosporin A and cladribine did not prevent rejection of such transplants. Mechanical separation of transplants from bone marrow prevented sensitization of recipients and rejection of the produced cartilage. Successful allogeneic chondrocyte transplants in rabbits and hens could be tentatively explained by a certain degree of inbreeding among experimental animals, by the use of chondrocytes cultivated before grafting in artificial scaffolds and thus protected by matrix produced in vitro, and also by creation of a temporary mechanical barrier between transplant and bone marrow by tissues damaged during preparation of the defect. PMID- 12112415 TI - Introduction for "donor transplant vs. recipient cells in musculoskeletal system". PMID- 12112417 TI - Response of the donor and recipient cells in mesenchymal cell transplantation to cartilage defect. AB - To facilitate the repair of articular cartilage defects, autologous mesenchymal cells from bone marrow or periosteum were transplanted in a rabbit model. Two weeks after the transplantation of the mesenchymal cells, the whole area of the original defect was occupied by cartilage. From the deep area of the reparative cartilage, which contacted with host bone, chondrocytes became hypertrophic and the invasion of bone with vasculature started, until the replacement reached the natural junction of the host cartilage and the subchondral bone about 4 weeks after transplantation. Twelve weeks after the transplantation, the repair cartilage in the defect became a little thinner than the adjacent normal cartilage, which became a little thinner 24 weeks after the transplantation (the longest observation period in the study). Large, full-thickness defects of the weight-bearing region of the articular cartilage were repaired with hyaline-like cartilage after implantation of autologous mesenchymal cells. The repair process by mesenchymal cell transplantation was explained as follows: The donor transplanted cell differentiated into cartilage and the defects were completely filled with cartilage. Then, mesenchymal cells that entered the chondrogenic lineage rapidly progressed through this lineage to the hypertrophic state, which was then the target for erosion and vascular invasion. Although this vasculature and the newly formed bone were considered to be host-derived, there was no evidence to that effect. To prove this, suitable experimental marking of these donor cells is needed. In the case of chondrocyte transplantation, the repair cartilage maintained its thickness to the full depth of the original defect; the tissue derived from the implanted chondrocytes was not invaded by vessels or replaced by subchondral bone. PMID- 12112418 TI - Behavior of graft and host cells in underlying subchondral bone after transplantation of osteochondral autograft. AB - Transplantation of osteochondral autograft is widely used as a therapeutic strategy for the defect of articular cartilage. In the repair process, although underlying subchondral bone becomes necrotic and then is followed by bone reconstruction, the fate of graft and host cells during remodeling of underlying subchondral bone has not been elucidated. The objectives of this study were to establish a method to follow graft and host cells after transplantation of osteochondral autograft, and to elucidate the fate of both graft and host cells during remodeling of underlying subchondral bone. For these purposes, autologous transplantation models employing transgenic rats and wild-type rats, which were genetically identical to each other except for transgenes, were used. Two transplantation models were designed so that either the graft or the host cells had transgenes. Model I: transgenic rats were the donor, and wild-type rats were the recipient; model II: conversely, wild-type rats were the donor, and transgenic rats were the recipient. The grafted bone marrow cells and osteocytes in the trabeculae survived in the graft at 3 weeks after transplantation. Invasion of the host bone marrow cells into the graft was also found. Thus, bone marrow cells in the host as well as both bone marrow cells and osteocytes in the graft could potentially participate in the remodeling of underlying subchondral bone. Furthermore, the interface between graft and host was consisted with both graft and host derived cells. Since new bone formation was found in this space, both graft and host cells could have the potential to contribute to remodeling of underlying subchondral bone. The two models of the transplantations using the transgenic rats were found to be beneficial in following graft cells as well as host cells and in understanding their function on healing after autologous transplantation. PMID- 12112419 TI - Donor cell survival and repopulation after intraarticular transplantation of tendon and ligament allografts. AB - The specific cells within ligaments and tendons are important to maintain the unique structural and material properties of these tissues. The use of tendon and ligament allografts with living cells for ligament reconstruction would be desirable assuming that these cells would survive after transplantation and continue to function. We assessed the fate of donor cells in fresh allografts of the patellar and anterior cruciate ligaments after transplantation. The cells in these allografts used to reconstruct the anterior cruciate ligament did not survive. This was demonstrated using a DNA probe technique that clearly distinguished donor cells from host cells in the Spanish goat model. The donor cells were replaced by host cells in a rapid manner. The host cells that repopulated the allografts assumed the histologic similarity to the fibroblasts they replace. Simultaneous full-thickness skin transplants in the same animals were not rejected during the interval of rapid loss of donor DNA from the allografts. The absence of rejection of the skin grafts at the one-week interval suggests that no pre-existing antibody associated with an immune reaction was responsible for the rapid loss of DNA in the allografts. The clinical basis for utilizing intra-articular allografts with living donor cells needs further justification to account for their increased expense, more complicated surgical logistics, and higher potential risk of disease transmission. PMID- 12112420 TI - Response of donor and recipient cells after transplantation of cells to the ligament and tendon. AB - The mechanical properties of healing ligaments and tendons are not comparable to those of normal tissue. To improve the quality of the ligament healing, therapeutic strategies include gene transfer or placement of mesenchymal stem cells at the healing site. Studies show that marker genes, growth factors, and antisense oligonucleotides can be delivered to both normal and healing ligaments and tendons by gene transfer. Cells with and without genetic modification have been successfully transplanted to ligaments and tendons and remain viable. Tendon healing can be improved using collagen gel implants seeded with autologous mesenchymal stem cells. Even though these early results are encouraging, more work is required regarding the response of the recipient site to donor cells or vectors. PMID- 12112421 TI - Fate of donor bone marrow cells in medial collateral ligament after simulated autologous transplantation. AB - A potential strategy to enhance ligament healing by transplantation of mesenchymal stem cells (MSCs), which are demonstrated to differentiate into fibroblast-like cells in vitro, is presented. The objective of this study was to follow transplanted nucleated cells from bone marrow, which contain MSCs, in the healing medial collateral ligament (MCL) over time, and to examine their phenotype and survivability. It was hypothesized that MSCs in nucleated cells from bone marrow would differentiate into fibroblast-like cells in the healing ligament following adaptation to the environment. The transplantation model employed in this study eliminates the immune response to a donor by the recipient using a transgenic rat (donor), which does not produce foreign protein from transgenes, and its wild-type rat (recipient) in order to simulate autologous transplantation. The MCL of the wild-type rat was ruptured, where 1 x 10(6) nucleated cells of bone marrow from the transgenic rat were injected. The transgenes in transplanted nucleated cells were detected throughout the healing MCL for 28 days by in situ hybridization. At 3 days, many donor cells were evident in the injury site and fascial pocket, and some were found in the midsubstance. Morphologically, transplanted cells with elongated nuclei were found at the ruptured edge of the midsubstance and surface of the unruptured site after 3 days. At 28 days, these cells continued to survive in the healing MCL. Their shapes were similar to those of surrounding recipient MCL fibroblasts. Thus, transplanted cells might differentiate into fibroblasts. Therefore, it was demonstrated that there is a potential for nucleated cells from bone marrow to serve as a vehicle for therapeutic molecules as well as to be a source in enhancing healing of ligaments. PMID- 12112422 TI - Adenovirus mediated gene transfer to skeletal muscle. AB - Transfer of therapeutic genes into muscle tissue has promise for the treatment of a variety of muscular dystrophies. Various vectors have been used to deliver genes to skeletal muscle but their application has faced several major limitations including: (1) the lack of transgene persistence caused by the immune rejection of transduced myofibers and/or vector toxicity, and (2) the maturation dependence of viral transduction. While the immunorejection and/or cytotoxic problems are being overcome with the development of new vectors, maturation dependent viral transduction is still a major hurdle in gene transfer to skeletal muscle. Poor adenoviral transduction in mature myofibers has been attributed to: (1) the extracellular matrix of mature myofibers may form a physical barrier and prevent the passage of large viral particles; (2) viral receptors are down regulated with muscle maturation; and (3) loss of myoblasts with muscle maturation, which serve as intermediaries in the viral transduction. In this review, we will focus on recent developments in overcoming those hurdles of gene therapy in skeletal muscle, especially to adenovirus (Ad), including: (1) new mutant vectors lacking all viral genes to decrease immunogenicity, and hence, improve persistence of transgene expression in muscle in vivo; (2) using tissue specific promoters to evade immunorejection; (3) permeabilization of the extracellular matrix; (4) modifying the viral receptors in mature myofibers; and (5) myoblast or muscle stem cell mediated ex vivo gene transfer. PMID- 12112424 TI - Introduction to the biology of phagocytosis. PMID- 12112423 TI - Histological and electrophysiological analysis of the peripheral nerve allografts using an immunosuppressive agent. AB - In peripheral nerve allografts, use of an immunosuppressive agent is one of the ways of reducing nerve rejection. FK506 is a newly discovered substance, extracted from Streptomyces tsukubaensis, and has strong immunosuppressive effects. In the present study, immunosuppressive effects of FK 506 were examined using allografts of rat sciatic nerves. Good nerve regeneration was demonstrated through 12 weeks in this model. The immunosuppressed group gained weight over the course of the experiment. Another study was performed to observe the histological changes caused by ceasing the administration of FK506. Administration of FK506 was terminated 12 weeks after grafting. At 8 weeks after cessation, cellular infiltration and large unmyelinated axons were observed in the extended subperineurial space of grafts. At 12 weeks, histological characteristics of rejection were not observed. In the electrophysiological study, the temporal dispersions were recorded at 4 and 8 weeks. However, the normal electrophysiological waves were recorded at 12 weeks after cessation. It was concluded that FK506 is effective for preventing rejection of nerve allografts without any serious side effects on rats, and findings of total rejection of grafts were not recognized after ceasing the administration of FK 506. In peripheral nerve allografts, short-term administration of an immunosuppressive agent is sufficient to lead to good nerve regeneration. PMID- 12112425 TI - Phagocytosis--the mighty weapon of the silent warriors. AB - Professional phagocytes, comprising polymorphonuclear neutrophils and monocyte/macrophage cells, play an important role in the host defense. Any defect in their function exposes the organism to microbial intruders terminating in fatal diseases. The functional responses of the phagocytes to bacterial and fungal infections include chemotaxis, actin assembly, migration, adhesion, aggregation, phagocytosis, degranulation, and reactive oxygen species production. Superoxide generation by phagocytic NADPH oxidase is an imperative step toward bacterial killing. Phagocytes participate in inflammatory reactions and exert tumoricidal activity. They are supported by serum factors such as immunoglobulins, cytokines, complement, the acute phase reactant C-reactive protein, production of antibacterial proteins, and others. In addition to their principal task to eliminate bacteria, they are engaged in removing damaged, senescent, and apoptotic cells. Engulfed cell debris, large particles such as latex beads, fat, and oil droplets, are examples of phagocytic activity illustrated in the present review with transmission and scanning electron microscope micrographs. Numerous factors, such as diseases and stressful conditions, affect the engulfing activity of the professional phagocytes. Our experience regarding the impaired phagocytic capacity of cells in patients with diabetes and chronic renal failure is discussed. The results obtained in our laboratory from experiments detecting the effect of strenuous physical exercise, hypothermia, fasting, and abdominal photon irradiation on the phagocytic capacity of human polymorphonuclear neutrophils and rat peritoneal macrophages are hereby summarized and the reports on those subjects in the recent literature are reviewed. A variety of assays are applied for quantifying phagocytosis. Flow cytometry based on incubation of phagocytic cells with fluorescent conjugated particles and measuring the amount of fluorescence as an indicator of the engulfing capacity of the cells is a useful method. A direct visualization of the ingested particles using light or electron microscopy is a valuable tool for estimation of phagocytic function. In our hands, the use of semithin sections of embedded phagocytes following their incubation with latex particles provided satisfactory results for measuring the total number of phagocytic cells, as well as the internalizing capacity of each individual cell. Microbiological assays, the nitroblue tetrazolium test, quantitation of antibody- and antigen-mediated phagocytosis, as well as methods reviewed in detail in other reports are additional applications for determination of this intricate process. PMID- 12112426 TI - Actin machinery of phagocytic cells: universal target for bacterial attack. AB - Uptake of microorganisms by eukaryotic cells depends on proper functioning of the actin machinery. It creates a driving force for the cell membrane deformations necessary for ingestion and killing of microbes by phagocytes. Therefore, specific alterations in the activity of the actin apparatus could be favorable for pathogenic bacteria, representing an efficient mechanism in their virulence. Such alterations are supposed to be achieved in two principle ways. One is accomplished via binding of bacterial ligands to certain surface receptors, which initiate subsequent actin cytoskeleton rearrangements. Another is to introduce cytoskeleton-targeted products directly into eukaryotic cells and in this way modulate the activity of the actin apparatus. Indeed, Legionella and some other intracellular parasites possess ligands able to stimulate certain receptors on the surface of phagocytes and possess devices suitable for translocation of effector molecules into eukaryotic cytoplasm. The results of such events could be increased uptake of these microbes and their subsequent transportation to permit multiplication in their intracellular niche. On the contrary, representatives of Clostridium and a number of other extracellular pathogens create products which penetrate eukaryotic cells and disorganize the actin cytoskeleton network, thus making uptake of these pathogens by phagocytes impossible. PMID- 12112427 TI - Oxidative stress in phagocytes--"the enemy within". AB - Phagocytes represent a powerful defense system against invading microorganisms that threaten the life or functional integrity of the host. The capacity to generate and release substantial amounts of reactive oxygen species is a unique property of activated polymorphonuclear and mononuclear phagocytes. The crucial role of these molecules in killing microorganisms and their consecutive contribution to tissue damage during injury and inflammation is widely known. Although much research has been done to explore the molecular events involved in the interaction of oxygen intermediates with microbes or host tissue, surprisingly little attention has been paid to the effect of reactive metabolites on the phagocyte itself. This fact is especially surprising, since it is apparent that the activated phagocyte is directly exposed to its own toxic metabolites. The potential damage occurring during excessive radical formation might notably alter the vital functions of these primarily immunocompetent cells. Moreover, the critical role of oxygen radicals in apoptosis of leukocytes has been recently revealed. Apoptosis is now supposed to represent a key mechanism in neutrophil deactivation and resolution of inflammation. Therefore, this review will focus on the delicate balance between released oxidants and antioxidative protection within the phagocytes themselves. General and phagocyte-specific antioxidative mechanisms, which have co-evolved with the radical generating machinery of phagocytes, are discussed, since the outcome of local inflammation can directly depend on this antioxidative capacity and might range from adequate elimination of the pathogen with minimal acute tissue damage to progression towards a systemic inflammatory response syndrome. PMID- 12112428 TI - Phagocytosis by invertebrate hemocytes: causes of individual variation in Panorpa vulgaris scorpionflies. AB - An in vitro phagocytosis assay, adjusted to as little as 1 microL of insect hemolymph, enables the microscopic determination of phagocytosis for single individuals of small insects. Even repeated determination over the lifetime of individuals is possible. This method makes it feasible to study individual variation in invertebrate phagocytic capacity. Possible sources of such variation are reviewed in this article: genetic differences, development, aging, reproduction, presence of parasites, and diverse environmental influences are natural sources of individual variation in phagocytosis. However, the methods used for phagocytosis and microscopic evaluation are also (unwelcome) sources of variation. To optimize incubation time for in vitro phagocytosis, time courses were determined. Furthermore, the reliability of visual counting and image analysis for the microscopic quantification of phagocytosis are compared. The influences of larval development and adult aging on phagocytosis by Panorpa vulgaris hemocytes are subsequently demonstrated. During development, a decrease in hemocyte numbers but a simultaneous increase in the proportion of phagocytosing hemocytes was observed when larvae reached pupation. On the other hand, adults showed a dramatic decrease in phagocytic capacity with age, while cell numbers remained fairly constant. The results show that individual variation in phagocytosis can be determined accurately in small invertebrates and related to its causes. This might be especially interesting in the context of studies relating individual immunocompetence to ecology, life history variation, and behavior. PMID- 12112429 TI - Bacteria-hemocyte interactions and phagocytosis in marine bivalves. AB - Marine bivalves (such as mussels, oysters, and clams) are widespread mollusks in coastal waters at different latitudes; due to their filter-feeding habits, they accumulate large numbers of bacteria from the harvesting waters and may act as passive carriers of human pathogens. To cope with this challenge, bivalves possess both humoral and cellular defense mechanisms with remarkably effective capabilities. The circulating cells, or hemocytes, are primarily responsible for defense against parasites and pathogens; microbial killing results from the combined action of the phagocytic process with humoral defense factors such as agglutinins (e.g., lectins), lysosomal enzymes (e.g., acid phosphatase, lysozyme), toxic oxygen intermediates, and various antimicrobial peptides. In this work, current knowledge of the mechanisms underlying the interactions between bacteria and the hemolymph components of marine bivalves is summarized. Bacterial susceptibility to hemolymph killing in different bivalve species may be a consequence of the different ability of bacterial products to attract phagocytes, the presence or absence of specific opsonizing molecules, the hemocyte capability to bind and engulf different bacteria, and the different bacterial sensitivity to intracellular killing. The role of soluble (e.g., agglutinins and opsonins) and surface-bound factors in bacterial phagocytosis by hemocytes of the most common marine bivalve species is described and the possibility that environmental temperatures and other seasonal factors may influence this process is considered. Moreover, the potential strategies used by bacteria to evade phagocytic killing by hemocytes are discussed. From the available data it is clear that several questions need further investigation; the elucidation of the factors influencing phagocytosis in bivalves and the fundamental strategies used by bacteria to escape hemolymph killing are important not only to understand bivalve immune defenses but also to explain the persistence of pathogenic bacteria in bivalve tissues and to predict the consequent impact on human health. PMID- 12112430 TI - Amphibia Kupffer cells. AB - Amphibia Kupffer cells (i.e., liver resident macrophages) show many common characteristics when compared with Mammalia Kupffer cells: filopodia, microvillous-like structures, lamellipodia, fuzzy coat, coated vesicles, bristled vacuoles, nonspecific esterase activity, and pinocytotic and phagocytic activity are present both in Amphibia and Mammalia Kupffer cells. On the other hand, some differences are present between Kupffer cells of both zoological classes: phagocytosed red cells and their derivatives, iron-protein complexes, and lipofuscin bodies are normally present in Amphibia Kupffer cells, but absent in the same cells of healthy mammals. Worm-like structures are not seen in Amphibia and endogenous peroxidase activity is very weak in these animals compared with Mammalia. The most important difference lies in the ability of Amphibia Kupffer cells to produce melanins: in fact the tyrosinase gene is expressed, "melanosome centers" are present, and dopa oxidase activity is demonstrable. PMID- 12112431 TI - Are thrombocytes and platelets true phagocytes? AB - Thrombocytes and platelets, beyond their primary function in hemostasis, seem to play an active role in inflammation. As regards their phagocytic ability, the results to date are confusing, incomplete, and somewhat contradictory. Whereas the interaction of avian thrombocytes or mammalian platelets with bacteria both in vitro and in vivo has received wide attention, almost no information exists on the topic in "lower" vertebrates. The aim of this work is to review the available information on the phagocytic properties of thrombocytes and platelets. Particular attention is payed to the ontogeny of these cells, the soluble factors involved in the inflammatory process derived from them, and their interaction with particulate material, mainly with bacteria. PMID- 12112432 TI - Role of the phagocytes on embryos: some morphological aspects. AB - Phagocytosis in embryos was studied by Elie Metchnikoff more than a century ago and is a pillar of the Phagocytic Theory. Throughout the last three decades phagocytosis in embryos has been studied from different perspectives, which this review describes and analyzes. The following branches were identified: 1) the search for the origin and first identification of well-known adult phagocytes in embryos, including their role after induced injuries; 2) the search for the occurrence of phagocytosis in embryos and its role during their physiological development; and 3) the search for phagocytosis in embryos, as a tool to study identity and self-recognition. It is possible to verify that different cell types are able to undertake phagocytosis, under a variety of different stimuli, and that the nature of what is phagocytosed also varies widely. Although the overwhelming majority of species described among metazoarians are invertebrates, most published articles in this field relate to mammals (particularly mice and humans) and birds (particularly chicks). In order to enrich this field of knowledge, research using a wider variety of vertebrate and invertebrate species should be undertaken. Furthermore, the present knowledge of phagocytosis in embryos needs a revised paradigm capable of embracing all the above-mentioned research trends under a single, more general, biological theory. In this sense, Metchnikoff's Phagocytic Theory, which is based on a broad biological paradigm and is thus capable of dealing with all research trends mentioned herein, should be revisited in order to contribute to this edification. PMID- 12112433 TI - Studies on glycogen autophagy: effects of phorbol myristate acetate, ionophore A23187, or phentolamine. AB - The effects of agents that could manipulate the lysosomal calcium such as phorbol myristate acetate, ionophore A23187, and phentolamine on the lysosomal glycogen degradation were studied by electron microscopy, morphometric analysis, and biochemical assays in newborn rat hepatocytes. Phorbol myristate acetate, which promotes the input of calcium to lysosomes, increased the total volume of autophagic vacuoles and the activity of lysosomal glycogen-hydrolyzing acid alpha 1,4 glucosidase and decreased the fractional volume of undigested glycogen inside the autophagic vacuoles and also decreased the activity of acid mannose 6 phosphatase. Ionophore A23187, which releases lysosomal calcium, produced opposite results in these enzyme activities. Phentolamine, an alpha-adrenergic blocking agent which interferes with the generation of phosphoinositides and may activate the lysosomal calcium uptake pump, increased the total volume of autophagic vacuoles and the activity of lysosomal glycogen-hydrolyzing acid glucosidase and decreased the fractional volume of undigested glycogen inside the autophagic vacuoles. The results of this study constitute evidence that changes in lysosomal calcium may influence certain aspects of autophagy, including the degradation of glycogen inside the autophagic vacuoles. They also support our previous postulate [Kalamidas and Kotoulas (2000a,b) Histol Histopathol 15:29-35, 1011-1018] that stimulation of autophagic mechanisms in newborn rat hepatocytes may be associated with acid mannose 6-phosphatase activity-deficient lysosomes. PMID- 12112434 TI - Morphological aspects of particle uptake by lung phagocytes. AB - Macrophages residing on the inner epithelial surfaces of airways and alveoli are the only lung phagocytes exposed directly to the environment. Their phagocytic and microbicidal activities are essential for maintaining this organ in a clean and sterile state. The morphology of these phagocytes can be investigated in situ only after implementing special techniques, which involve intravascular triple perfusion of aqueous fixatives or instillation of nonpolar ones. Such studies have revealed the engulfment of particles by these cells to be rapid, the process being essentially complete within a day. Particles are entrapped within phagosomes and the host cells eventually transported out of the lungs by mucociliary action, macrophages with higher loads being more rapidly eliminated than those with lower ones. Very small particles or those persisting on the epithelial surfaces may be taken up by the eponymous cells. Translocation of particles into the underlying connective tissue and their subsequent phagocytosis by interstitial macrophages prolongs their retention time in the lungs. The still poorly studied pleural macrophages might be involved in cell-mediated immune responses within the pleural space. Intravascular pulmonary macrophages figure largely in the phagocytosis of circulating particles. The role played by dendritic cells in particle uptake by the lungs is not well understood. Airway and alveolar macrophages are the primary phagocytes of the lung. In nonoverload situations and for particles >1 microm, a small proportion of those recruited suffices to remove material from the epithelial surface before other phagocytes, with an apparently greater immunological potential, gain access to it. PMID- 12112435 TI - Changes in macrophage morphology and prolonged cell viability following exposure to polyethylene particulate in vitro. AB - The interaction of macrophages and ultra-high molecular weight polyethylene (PE) wear plays an important role in perpetuating chronic inflammation at the bone implant interface, leading to peri-implant osteolysis and mechanical failure of the implant. A model to study the interaction of human mature macrophages with orthopaedic biomaterial wear has been previously developed. With the use of the model, in this study, the mature human monocyte-derived macrophages (MDMs) were observed with light, fluorescent, and scanning electron microscopy (SEM), as well as transmission electron microscopy (TEM). The cell viability was investigated using calcein and ethidium staining. Following exposure to PE particulate, the morphology of the human MDMs was heterogeneous: rounded, flattened, and elongated. There was no morphological evidence of cytotoxicity or apoptosis. The MDM viability was not influenced by phagocytosis of PE particulate in a negative fashion. In fact, more prolonged cell viability was observed in the human MDMs exposed to PE particulate when compared to controls. PMID- 12112436 TI - Phagocytosis of apoptotic cells by liver: a morphological study. AB - The present review deals with the morphological features of the removal of apoptotic cells by liver. The engulfment of cells undergoing apoptosis can be considered a specialized form of phagocytosis, playing a major role in the general tissue homeostasis in physiological and pathological conditions. In fact, defects of phagocytosis of apoptotic cells might have deleterious consequences for neighboring healthy cells, i.e., pathogenesis of inflammatory disease or dysregulation of the immune system. Phagocytosis of apoptotic cells by liver is a complex phenomenon, involving multiple molecular mechanisms of recognition (i.e., lectin-like receptors and receptors for externalized phosphatydilserine) of both parenchymal (hepatocytes) and nonparenchymal (Kupffer and endothelial cells) liver cells, often operating in cooperation. The data discussed in the present review are drawn from studies of phagocytosis of apoptotic cells in the liver, carried out with in vivo and in situ adhesion experiments as well as in vitro assays. Our results indicate that the three main liver cell types (hepatocytes, Kupffer, and endothelial cells) are able to recognize and internalize apoptotic cells by means of specific receptors (galactose and mannose-specific receptor; receptor for phosphatydilserine) and by cytoskeletal reorganization that favors the engulfment of the apoptotic cells. The "flags" for the identification of apoptotic cells by the liver are modifications of the surface of dead cells, i.e., sugar residues and phosphatydilserine exposition. Vitronectin receptor is not involved in such a recognition. The adhesions between modified cell surfaces of apoptotic cells and phagocytes generate cytoplasmatic signaling pathways that drive apoptotic cells to their final fate within the phagocytes (i.e., lysosomal digestion). PMID- 12112437 TI - Myelin phagocytosis by macrophages and nonmacrophages during Wallerian degeneration. AB - The literature concerning Schwann cells (SCs) and macrophages in myelin phagocytosis during Wallerian degeneration is reviewed. SCs carry out the first step in the removal of myelin by segmenting myelin and then incorporating the degraded myelin. The recruited macrophages then join in the myelin-phagocytosis event, appearing to make full use of their original phagocyte abilities until the end of myelin clearance. The molecular mechanisms of the two cells underlying myelin phagocytosis are thought to be different; myelin phagocytosis by SCs being lectin-mediated, i.e., opsonin-independent, whereas that of macrophages is mainly opsonin-dependent. It is important to note that SCs and macrophages cooperatively accomplish myelin phagocytosis. PMID- 12112438 TI - Vascular permeability in blood capillary. PMID- 12112439 TI - Transcytosis of plasma macromolecules in endothelial cells: a cell biological survey. AB - The modern exploration of endothelial cell biology is a largely interdisciplinary exercise. Cell biological, physiological, and more recently molecular biology approaches were used to study the pathways and the organelles involved in transcytosis of macromolecules in endothelial cell (EC). Here we discuss mainly the cell biological findings that revealed that EC have the attributes to fulfill the transport function. They are polarized cells, heterogeneous, and, thus, structurally and functionally adapted to the vascular bed in which they reside. The structural heterogeneity involves the number and distribution of plasmalemmal vesicles (caveolae), their generated channels, and the organization of intercellular junctions. The closely related functional heterogeneity comprises the degree of permeability for plasma molecules that vary as a function of organ. The EC are endowed with the cellular machinery to perform (1) endocytosis, that is to take up plasma proteins and the molecules they carry to be used for themselves (cholesterol-carrying low density lipoproteins, fatty acid carrying albumin, iron carrying transferrin, etc.), and (2) transcytosis, which implies to transport plasma proteins to the subjacent cells and tissues. The possible pathways for transport of molecules are transcellular, via caveolae and channels, and paracellular via intercellular junctions. Most of the results obtained, so far, indicate that transcytosis of albumin, low-density lipoproteins, metaloproteases, and insulin, is performed by cargo-vesicles and their generated channels. The paracellular pathway can be used for water and ions; in postcapillary venules, at the level of which approximately 30% of junctions are open to a space of 6 nm, small molecules may take this route. Recent data obtained by molecular biology techniques revealed that caveolae are endowed with the molecular machinery for fusion/fission, docking, and movement across cells. Moreover, the various and numerous molecules that have been detected in the caveolae membrane and the different functions assumed by this differentiated microdomain strengthen the postulate that there are at least two or more types of vesicles molecularly tailored for the local physiological requirements. PMID- 12112440 TI - Ultrastructural studies define soluble macromolecular, particulate, and cellular transendothelial cell pathways in venules, lymphatic vessels, and tumor associated microvessels in man and animals. AB - We present de novo studies and review published efforts from our laboratory, spanning 12 years (from 1988 to 2000), where we have used ultrastructural approaches to study the functional anatomy of the microvasculature in man and animals in health and disease. These efforts have defined a new endothelial cell organelle, termed the vesiculo-vacuolar organelle (VVO), which participates in the regulated transendothelial cell passage of soluble macromolecules. The studies defining this organelle utilized ultrathin serial sections, three dimensional computer-assisted reconstructions, and ultrastructural electron-dense tracers to establish luminal to abluminal transendothelial cell continuity of VVOs. Commonality of VVOs and caveolae is suggested by the ultrastructural anatomy of individual units of VVOs and caveolae, the presence of caveolin in both structures, and a mathematical analysis of morphometric data, all of which suggest that VVOs form from fusions of individual size units equivalent to vesicles of caveolar size. Ultrastructural studies have localized potent permeability factors and their specific receptors to VVOs in in vivo tumor and allergic inflammation models. Regulation of permeability through VVOs has been quantified and shown to be increased in tumor microvessels and in control vessels exposed to potent permeability-inducing mediators. The transendothelial cell passage of particulate macromolecules occurs by vacuolar transport in tumor vessels; in permeability factor-exposed control vessels, colloidal carbon traversed endothelial cells via the development of pores that did not communicate with or disrupt intercellular junctions by gap formation. Serial section and computer-assisted reconstructions established these findings and suggested the possible development of transendothelial cell pores from VVOs. Serial sectioning and computer-assisted three-dimensional reconstructions of ultrastructural samples of an acute inflammation model revealed a transendothelial cell traffic route for motile neutrophils and platelets in the absence of classical ultrastructural criteria for regulated secretion from either cell. PMID- 12112441 TI - Morphological and cytochemical aspects of capillary permeability. AB - Transport of plasma soluble constituents across the capillary wall is of primordial importance in cardiovascular physiology. While physiological experiments have concluded with the existence of two sets of pores, a large one responsible for the transport of proteins and a small one designed for the diffusion of small solutes, the morphological counterparts have yet to get general agreement. In this review, we present the different proposed paths within and between the endothelial cells that do allow passage of plasma constituents and may respond to the definitions established by physiological means. The vesicular system existing in endothelial cells has been the first transendothelial path to be proposed. Several data have demonstrated the involvement of this system in transport, although others have systematically brought controversy. One alternative to the vesicles has been the demonstration of membrane-bound tubules creating, in certain cases, transendothelial channels that would allow diffusion of plasma proteins and other constituents across the capillary wall. Access to this tubulo-vesicular system could be restrained by the stomatal diaphragm and facilitated by specific membrane receptors. Further, we have demonstrated for the first time with morpho-cytochemical tools, that the intercellular clefts are the site of diffusion for small molecules such as peptides having a molecular weight inferior to 3,000. For the fenestrated capillary bed, we have shown that fenestrae are the site through which plasma constituents cross the capillary wall. However, and in spite of the existence of these large open pores, the endothelial cells still display the tubulo-vesicular system involved in transport of large molecules and their intercellular clefts are also the site of diffusion of small molecules. Making consensus on the existence of an intracellular tubulo-vesicular system in non-fenestrated capillaries, responsible for the transport of large molecules by the endothelial cells, and understanding the rational for the fenestrated capillary to have three paths for transport--the fenestrae, the tubulo-vesicular system, and the inter endothelial clefts--require further investigation. PMID- 12112442 TI - Structure and function of endothelial caveolae. AB - Caveolae are spherical invaginations of the plasma membrane and associated vesicles that are found at high surface densities in most cells, endothelia included. Their structural framework has been shown to consist of oligomerized caveolin molecules interacting with cholesterol and sphingolipids. Caveolae have been involved in many cellular functions such as endocytosis, signal transduction, mechano-transduction, potocytosis, and cholesterol trafficking. Some confusion still persists in the field with respect to the relationship between caveolae and the lipid rafts, which have been involved in many of the above functions. In addition to all these, endothelial caveolae have been involved in capillary permeability by their participation in the process of transcytosis. This short review will focus on their structure and components, methods used to determine these components, and the role of caveolae in the transendothelial exchanges between blood plasma and the interstitial fluid. PMID- 12112443 TI - Functional expression and localization of P-glycoprotein at the blood brain barrier. AB - Until recently, the blood-brain barrier was viewed as a static lipid membrane barrier. Physical attributes of the cerebral endothelial cells such as the presence of tight junctions, paucity of vesicles or caveolae, and high electrical resistance were believed to be the primary components that provide the membrane selectivity of the blood-brain barrier to a variety of circulating compounds from the periphery. However, results from molecular biology, immunocytochemistry, biochemistry, and transport studies show that the cerebral endothelial cells possess an asymmetrical array of metabolic enzymes (i.e., alkaline phosphatase, cytochrome P450 enzymes, glutathione transferases) and energy-dependent efflux transport proteins (i.e., P-glycoprotein and Multidrug-resistance proteins) that are instrumental to the barrier function. P-glycoprotein, a membrane-associated, energy-dependent, efflux transporter, is expressed in brain parenchyma (i.e., astrocytes and microglia) as well as in blood-brain and blood-cerebrospinal fluid barriers. Its function along the blood-brain barrier is believed to prevent the accumulation of potentially harmful compounds in the brain by actively removing them from the brain into the peripheral circulation. This is a brief review on the expression and activity of P-glycoprotein at the blood-brain barrier, which reports on the localization of the protein in rat brain capillaries in situ as well as in a well-characterized in vitro model of the blood-brain barrier, an immortalized rat brain endothelial cell line, the RBE4. Immunocytochemical analysis employing various P-glycoprotein monoclonal antibodies, demonstrated the presence of the protein along the plasma membrane, in plasmalemmal vesicles and nuclear envelope of rat cerebral endothelial cells, both in situ and in vitro. Western blot analysis revealed a single band with a molecular weight of 170-180 kDa, a size previously reported for P-glycoprotein, in RBE4 cells. In addition, results from functional studies show that the accumulation of the P-glycoprotein substrate digoxin by RBE4 monolayer cells is significantly enhanced in the presence of standard P-glycoprotein inhibitors (verapamil, cyclosporin A, PSC 833), protease inhibitors (saquinavir, ritonavir, indinavir), and the metabolic inhibitor, sodium azide. These results demonstrate the functional expression of P glycoprotein in the immortalized rat brain endothelial cell line, RBE4. Novel in situ and in vitro intracellular locations of P-glycoprotein in cerebral endothelial cells have been identified suggesting that this transporter may play a significant role in the subcellular distribution of substrates in the brain. PMID- 12112444 TI - Diversity in unity: the biochemical composition of the endothelial cell surface varies between the vascular beds. AB - The vascular endothelium represents a population of squamous epithelial cells characterized by a particular histological localization (intima of blood vessels) and by several physiological functions such as the transport of substances between blood and tissues, the modulation of the vascular tone, the control of blood coagulation and that of the leukocyte extravasation. In spite of all these elements in common and of an identical embryonic origin, endothelial cells show definite morphological and physiological variations that divide them into types and subtypes, each specifically associated to various categories of organs. Even within the vasculature of the same organ, there are clear segmental (arterial/capillary/venous) differentiations of the endothelial cells. While the morphological and physiological differences between endothelial cells are well documented, there are very few data on the biochemistry underlying this heterogeneity. This work presents several data suggesting that, at present, the domain is ripe for a comprehensive analysis of this biochemical diversity, at least in what concerns the luminal aspect of the endothelial plasmalemma, a compartment of crucial importance in the biology and pathology of the cardiovascular system. PMID- 12112445 TI - Diabetic vascular dysfunction: links to glucose-induced reductive stress and VEGF. AB - A complete biochemical understanding of the mechanisms by which hyperglycemia causes vascular functional and structural changes associated with the diabetic milieu still eludes us. In recent years, the numerous biochemical and metabolic pathways postulated to have a causal role in the pathogenesis of diabetic vascular disease have been distilled into several unifying hypotheses. These involve either increased reductive or oxidative stress to the cell, or the activation of numerous protein kinase pathways, particularly protein kinase C and mitogen-activated protein kinases. As detailed below, there is tremendous crosstalk between these competing hypotheses. We propose that increased tissue glucose levels alter cytosolic coenzyme balance by increased flux of glucose through the sorbitol pathway increasing free cytosolic NADH levels. Increased NADH levels can generate reactive oxygen species via numerous mechanisms, lead to the formation of intracellular advanced glycation end products, and induce growth factor expression via mechanisms involving protein kinase C activation. The elevation in growth factors, particularly vascular endothelial growth factor (VEGF), is responsible for the vascular dysfunction via numerous mechanisms reported here in detail. PMID- 12112446 TI - Sequential morphological and permeability changes in the rete capillaries during hyperglycaemia. AB - With the rete model of the eel swimbladder, we have studied the appearance and development of a microangiopathy during a 2-year period of hyperglycaemia. Hyperglycaemia was induced in the eel by chronic exposure to cold water. At 3-5 months, basement membrane thickness was twice the normal value and increased only slightly thereafter. Diffusion coefficients of permeability were measured in counter-current perfusion experiments for a variety of tracers that are believed to use different pathways of transcapillary transport. The permeability to sucrose was the first to significantly increase, at 6-8 months, followed by that of albumin, insulin, and inulin, at 9-11 months and that of sodium, at 18-24 months. The permeability to water and antipyrine remained stable throughout the study. The results indicate that in the rete model, chronic hyperglycaemia induces a rapid thickening of the capillary basement membrane and selective permeability increments in the various paths of transcapillary transport. PMID- 12112447 TI - Neurobiology of Fragile X syndrome: from molecular genetics to neurobehavioral phenotype. PMID- 12112448 TI - Molecular phenotype of Fragile X syndrome: FMRP, FXRPs, and protein targets. AB - Fragile X syndrome (FraX) is one of the most prevalent genetic causes of mental retardation. FraX is associated with an unstable expansion of a polymorphism within the 5' untranslated region of the FMR1 gene. The main consequence of this mutation is a reduction in the levels of the gene product (FMRP). FMRP is an RNA binding protein with multiple spliced variants (isoforms) and high levels of expression in a variety of tissues, including neurons. In the latter cells, it is localized not only to the perikaryon but also to dendrites and dendritic spines. FMRP belongs to a family of proteins that includes the Fragile X Related Proteins or FXRPs. FXRPs share high homology in their functional domains with FMRP, and also associate with mRNA and components of the protein synthesis apparatus. However, FXRPs do not have the same temporo-spatial pattern of distribution (and other properties) of FMRP. Immunochemical assays have confirmed that a functionally uncompensated FMRP deficit is the essence of the FraX molecular phenotype. Here, we report our preliminary study on FXRPs levels in leukocytes from FraX males. By immunoblotting, we found that a marked reduction in FMRP levels is associated with a modest increase in FXR1P and no changes in FXR2P levels. The consequences of this reduced FMRP expression on protein synthesis, in other words, the identification of FMRP targets, can be studied by different molecular approaches including protein interaction and proteomics methods. By two dimensional gel electrophoresis, we showed that in FraX leukocytes there is a defect in acetylation that involves prominently the regulatory protein annexin-1. Extension of current studies of the molecular phenotype to more brain-relevant tissue samples, a wider range of proteomics-based methods, and correlative analyses of FMRP homologues and FMRP targets with multiple behavioral measures, will greatly expand our understanding of FraX pathogenesis and it will help to develop and monitor new therapeutic strategies. PMID- 12112449 TI - Fragile X mental retardation: misregulation of protein synthesis in the developing brain? AB - Fragile X mental retardation results from the absence of a selective RNA-binding protein, FMRP. Previous studies demonstrated that FMRP forms messenger ribonucleoprotein (mRNP) complexes to associate with translating polyribosomes, suggesting that FMRP is involved in regulating protein synthesis. We are now facing the changing questions: How does FMRP influence protein synthesis in the brain? What is the target for FMRP in learning and memory? How does the absence of FMRP cause misregulation of protein synthesis, which in turn leads to mental impairment in fragile X syndrome? Models for abnormal neuronal function as a result of misregulated translation due to the absence of FMRP are discussed. PMID- 12112451 TI - Effects of Fragile X syndrome and an FMR1 knockout mouse model on forebrain neuronal cell biology. AB - The neurological deficits exhibited by patients with Fragile X syndrome (FraX) have been attributed to the absence of the Fragile X Mental Retardation Protein (FMRP), the product of the FMR1 gene, which is nonfunctional in these individuals. While a great deal has been learned about FraX using non-invasive techniques and autopsy tissue from humans, the limited availability of subjects and specimens severely restricts the rate at which such data can be collected and the types of experimental questions posed. In view of these limitations, a transgenic mouse model of FraX has been constructed in which the FMR1 gene is selectively knocked out (KO) [Bakker et al. (1994) Cell 78:23-33]. These mice show molecular, morphological, and behavioral alterations consistent with phenotypes observed in FraX patients, making them good models to study the absence of FMRP expression. PMID- 12112450 TI - Fragile X syndrome, the Fragile X related proteins, and animal models. AB - The Fragile X syndrome (FraX), which is characterized among other physical and neurologic impairments by mental retardation, is caused by the absence of the product of the FMR1 gene. The Fragile X Mental Retardation Protein (FMRP) is a member of a novel family of RNA-binding proteins. The latter includes two other proteins highly homologous with FMRP: the fragile X related proteins 1 and 2 (FXRP1 and FXRP2). Characterization of FXRPs, including their interaction with FMRP, will provide critical information about the mechanisms of action of FMRP and the role of this group of proteins in FMRP-deficient conditions such as FraX. Genetic manipulations of FMRP and the FXRPs should also provide valuable tools for investigating pathophysiology and gene therapies in FraX. The present review summarizes the strategies used for identifying the FXRPs, their chromosomal localization, molecular structure, and tissue distribution. It also reviews interactions between different members of this family of RNA-binding proteins. Animal models, both knockout and transgenic, of FMRP and the FXRPs are discussed. Phenotypic features of the FMR1 knockout mouse, the FMR1 transgenic rescue mouse, and other novel strategies for manipulating and delivering FMRP and FXRPs to the brain and other tissues are described. PMID- 12112452 TI - Cerebral growth in Fragile X syndrome: review and comparison with Down syndrome. AB - Neuroimaging studies have shown selective changes in brain size in Fragile X syndrome (FraX), which include reductions in the posterior cerebellar vermis, age dependent increases in hippocampal volume, and enlarged caudate nucleus and thalamus. Contrasting with these limbic and subcortical anomalies, much less is known about the neocortex in FraX. The present study attempted to examine cerebral and lobar-level volumetric changes in young males with FraX (2-7 years), by comparing groups of subjects with full mutation (FM) and mosaicism (Mos) with both age-matched controls and subjects with developmental language delay (DLD) and Down syndrome (DS). For this purpose, we used high resolution (i.e, SPGR) MRI scans and semi-automated methods for segmenting (tissue class) and parcellating (i.e., Talairach) the brain. In agreement with previous studies, we found no changes in overall brain or cerebrum size in FraX. Nevertheless, boys with FM FraX had relative reductions in temporal lobe volume (primarily gray matter) and relative preservation/enlargement of parietal white matter volume. While temporal lobe reductions were not specific, since they were also observed in DLD and DS subjects, parietal preservation/enlargement was only seen in FraX. The relevance of these preliminary findings was emphasized by comparisons between FraX groups, which revealed more marked changes in FM FraX than in Mos FraX (i.e., gene dosage). While cross-sectional analyses revealed marked age-dependent decreases in DS, a group showing marked global and lobar volumetric reductions, there were no changes over time in FraX. These neuroimaging data are discussed in the context of FraX neurobiology and other developmental disorders. PMID- 12112453 TI - Influence of stimulants on electrodermal studies in Fragile X syndrome. AB - Attention Deficit Hyperactivity Disorder (ADHD) is seen in the majority of children with Fragile X Syndrome (FraX). Previous work has documented an enhanced sweat response to stimuli in children with FraX compared to controls utilizing electrodermal response (EDR) measures. The present study assesses the EDRs both on and off stimulants in 19 children with ADHD and FraX compared to 17 age- and IQ-matched control patients with ADHD and developmental delays. Although the baseline EDRs were comparable between FraX patients and controls, the patients with FraX had a significant decrease in EDR amplitude and number of peaks when treated with stimulants compared to controls. This suggests that patients with FraX are more responsive to the enhancement of inhibitory systems that occur with stimulant use for ADHD. The use of a quantifiable measure, such as EDR, is recommended in future studies of treatment efficacy. PMID- 12112454 TI - Early development in males with Fragile X syndrome: a review of the literature. AB - This article reviews the current bibliographic knowledge on early neurobehavioral development and milestones in Fragile X syndrome (FraX), with emphasis on males affected by the condition. Three broad areas of early development were examined: (1) gross and fine motor, (2) speech and language, and (3) social. The result of the current review indicates very limited information on the developmental milestones in all three areas. The scarce literature on motor development shows that in FraX there is an early developmental delay. Research on speech and language demonstrates pervasive deficits in conversational skills and severe developmental delay, with increasing discrepancy between language level and chronological age in young males with FraX. Finally, deficits in social development in FraX include abnormal gaze, approach and avoidance conflict, and high incidence of autistic spectrum disorders. PMID- 12112455 TI - A developmental approach to understanding Fragile X syndrome in females. AB - The psychological phenotype of females with fragile X syndrome (FraX) is discussed, focusing primarily on empirical findings over the past decade and on studies of probands with the full mutation (FM). A developmental approach is used to help characterize specific patterns of cognitive, neuropsychological, social, emotional, and behavioral functioning across the lifespan of females with FraX. Approximately half of females with the syndrome present with cognitive abilities that fall in the borderline to mentally retarded range, and the remaining females with average intellectual functioning may experience relative deficits in math achievement and problems with attention and executive functioning. Reports of socio-emotional functioning are somewhat inconsistent, due in part, perhaps, to methodological differences in study design. To date, much of what we understand about the psychological phenotype of FraX is based on cross-sectional studies of girls and women with the disorder. Symptoms associated with shyness, and social anxiety and avoidance have been reported in some school-age, adolescent, and adult females with FraX. Only recently have efforts begun to identify the developmental trajectory of FraX in infants and toddlers. There is a void of information specific to these developmental periods. Identifying key deficits in cognitive and socio-emotional functioning has important implications for early detection and intervention for girls with FraX. Directions for future research are discussed. PMID- 12112456 TI - Embryonic and fetal rat myoblasts form different muscle fiber types in an ectopic in vivo environment. AB - Limb muscle development is characterized by the migration of muscle precursor cells from the somite followed by myoblast differentiation and the maturation of myotubes into distinct muscle fiber types. Previous in vitro experiments have suggested that rat limb myoblasts are composed of at least two distinct myoblast subpopulations that appear in the developing hindlimb at different developmental stages. These embryonic and fetal myoblast subpopulations are believed to generate primary and secondary myotubes, respectively. To test this hypothesis, cells obtained from embryonic day 14 (ED 14) and ED 20 rat hindlimbs were analyzed for myosin heavy chain expression after long-term differentiation in adult rat brains. Fetal myoblasts from ED 20 hindlimbs produced muscle fibers with a phenotype similar to that seen in tissue culture--predominantly fast myosin with a small proportion also coexpressing slow myosin. However, injection sites populated by embryonic myoblasts from ED 14 hindlimbs produced a different phenotype from that previously reported in culture, with fibers expressing an entire array of myosin isoforms. In addition, a subpopulation of fibers expressing exclusively slow myosin was found only in the embryonic injection sites. Our results support the existence of at least three myogenic subpopulations in early rat limb buds with only one exhibiting the capability to differentiate in vitro. These findings are consistent with a model of muscle fiber type development in which the fiber type potential of myoblast populations is established before differentiation into myotubes. This process establishes myogenic subpopulations that have restricted adaptive ranges regulated by both intrinsic and extrinsic factors. PMID- 12112457 TI - Expression, gene regulation, and roles of Fisp12/CTGF in developing tooth germs. AB - Odontogenesis involves multiple events, including tissue-tissue interactions, cell proliferation, and cell differentiation, but the underlying mechanisms of regulation are far from clear. Because Fisp12/CTGF is a signaling protein involved in similar events in other systems, we asked whether it is expressed in developing tooth germs and what roles it may have. Indeed, Fisp12/CTGF transcripts were first expressed by dental laminas, invaginating epithelium, and condensing mesenchyme at the bud stage, and then became abundant in enamel knot and preameloblasts. Fisp12/CTGF was present not only in inner dental epithelium but also in stratum intermedium and underlying dental mesenchyme. Fisp12/CTGF expression decreased markedly in secreting ameloblasts. Tissue reconstitution experiments showed that Fisp12/CTGF expression in dental epithelium required interaction with mesenchyme but was maintained by treatment of epithelium with transforming growth factor-1, a factor regulating Fisp12/CTGF expression in other systems, or with bone morphogenetic protein-2. Loss-of-function studies using CTGF neutralizing antibodies revealed that interference with endogenous factor action in tooth germ explants led to a severe inhibition of proliferation in both epithelium and mesenchyme and a marked delay in cytodifferentiation of ameloblasts and odontoblasts. Treatment of dental epithelial and mesenchymal cells in culture with recombinant CTGF stimulated cell proliferation, whereas treatment with neutralizing antibodies inhibited it. The data demonstrate for the first time that Fisp12/CTGF is expressed during odontogenesis. Expression is confined to specific sites and times, is regulated by epithelial-mesenchymal interactions and critical soluble factors, and appears to be needed for proliferation and differentiation along both ameloblast and odontoblast cell lineages. PMID- 12112458 TI - Gene-dosage-sensitive genetic interactions between inversus viscerum (iv), nodal, and activin type IIB receptor (ActRIIB) genes in asymmetrical patterning of the visceral organs along the left-right axis. AB - We have shown previously that mice deficient in the activin type IIB receptor (ActRIIB) exhibit right isomerism, which is characterized by mirror-image symmetrical right lungs, complex cardiac malformations, and hypoplasia of the spleen. These observations led us to hypothesize that the signaling of a TGF-beta family member by means of ActRIIB is necessary for the determination of the left sidedness of the visceral organs. To test this hypothesis, we examined laterality defects in mice carrying mutations in both ActRIIB and inversus viscerum (iv) genes, because iv(-/-) mice display a spectrum of laterality defects, including situs inversus, right isomerism, and left isomerism. We found that all mice homozygous for both iv and ActRIIB mutations displayed the right isomerism. The phenotype of right isomerism in the double mutants was also more severe than that in ActRIIB(-/-) mice as shown by persistent left inferior vena cava, right atrial isomerism, and hypoplasia of spleen. Interestingly, the incidence of right isomerism also increased significantly in iv(-/-);ActRIIB(+/-) and iv(+/ );ActRIIB(-/-) mice compared with homozygous mice carrying either of single gene mutations. A mechanism of the genetic modulation between ActRIIB and iv genes may be that iv modulates the asymmetric expression of a TGF-beta family member that signals through activin type II receptors, ActRIIA and ActRIIB, to specify the "left-sidedness." Nodal is the most likely candidate. We show here that the penetrance and severity of the right isomerism is significantly elevated in nodal(+/-); ActRIIB(-/-) mice, compared with ActRIIB(-/-) mice. Furthermore, the chimeric mice derived from nodal(-/-) ES cells displayed right isomerism, indistinguishable from that in (iv(-/-);ActRIIB(-/-)) mice. We propose that iv functions to establish asymmetric expression of nodal in a gene-dosage-sensitive manner and that nodal signals through the activin type II receptors to specify the left-sidedness by means of a threshold mechanism. PMID- 12112459 TI - Efficient ectopic gene expression targeting chick mesoderm. AB - The chick model has been instrumental in illuminating genes that regulate early vertebrate development and pattern formation. Targeted ectopic gene expression is critical to dissect further the complicated gene interactions that are involved. In an effort to develop a consistent method to ectopically introduce and focally express genes in chick mesoderm, we evaluated and optimized several gene delivery methods, including implantation of 293 cells laden with viral vectors, direct adenoviral injection, and electroporation (EP). We targeted the mesoderm of chick wing buds between stages 19 and 21 (Hamburger and Hamilton stages) and used beta galactosidase and green fluorescent protein (GFP) to document gene transfer. Expression constructs using the cytomegalovirus (CMV) promoter, the beta-actin promoter, and vectors with an internal ribosomal entry sequence linked to GFP (IRES-GFP) were also compared. After gene transfer, we monitored expression for up to 3 days. The functionality of ectopic expression was demonstrated with constructs containing the coding sequences for Shh, a secreted signaling protein, or Hoxb-8, a transcription factor, both of which can induce digit duplication when ectopically expressed in anterior limb mesoderm. We identified several factors that enhance mesodermal gene transfer. First, the use of a vector with the beta-actin promoter coupled to the 69% fragment of the bovine papilloma virus yielded superior mesodermal expression both by markers and functional results when compared with several CMV-driven vectors. Second, we found the use of mineral oil to be an important adjuvant for EP and direct viral injection to localize and contain vector within the mesoderm at the injection site. Lastly, although ectopic expression could be achieved with all three methods, we favored EP confined to the mesoderm with insulated microelectrodes (confined microelectroporation- CMEP), because vector construction is rapid, the method is efficient, and results were consistent and reproducible. PMID- 12112460 TI - Cell reorganisation during epithelial fusion and perforation: the case of ascidian branchial fissures. AB - In this study, we have analysed ultrastructurally the mechanism of epithelial fusion and perforation during the development of branchial fissures in the larva and bud of the colonial urochordate Botryllus schlosseri. Perforation of membranes represents an important process during embryogenesis, occurring to create communication between two separate compartments. For example, all chordate embryos share the formation of pharyngeal plates, which are constituted of apposed endodermal and ectodermal epithelia, which have the capacity to fuse and perforate. Although the process of perforation is extremely common, its cellular mechanism remains little understood in detail, because of the complexity of the structures involved. In B. schlosseri, two epithelial monolayers, the peribranchial and the branchial ones, with no interposed mesenchymal cells, participate in pharyngeal perforation. Blood flows in the interspace between the two cellular leaflets. Apico-lateral zonulae occludentes seal the cells of each epithelium, so that the blood compartment is separated from the environment of the peribranchial and branchial chambers; here, sea water will flow when the zooid siphons open. Stigmata primordia appear as contiguous thickened discs of palisading cells of branchial and peribranchial epithelia. The peribranchial component invaginates to contact the branchial one. Here, the basal laminae intermingle, compact, and are degraded, while the intercellular space between the two epithelia is reduced to achieve the same width as that found between the lateral membranes of adjacent cells. Cells involved in this fusion rapidly change their polarity: they acquire a new epithelial axis, because part of the adhering basal membrane becomes a new lateral surface, whereas the original lateral membranes become new apical surfaces. Before disassembling the old tight junctions and establishing communication between branchial and peribranchial chambers, cells of the stigmata rudiments form new tight junctions organised as distinct entities, so that the structural continuum of the epithelial layers is maintained throughout the time of fusion and perforation. PMID- 12112461 TI - Comparative analysis of the expression and regulation of Wnt5a, Fz4, and Frzb1 during digit formation and in micromass cultures. AB - Previous studies have shown that three members of the Wnt signaling pathway, the ligand WNT5A, the receptor FZ4, and the Wnt antagonist FRZB1, are implicated in the formation and differentiation of the digits. In this study, we have attempted to establish a functional correlation between them by comparing their expression patterns and their regulation by the signals controlling proliferation and differentiation of the limb mesoderm during formation of the avian digits in vivo and in micromass cultures. In vivo Wnt5a and Fz4 are expressed in the undifferentiated mesoderm of the autopod and in the differentiating digit cartilages. In the undifferentiated mesoderm, the expression of both genes is regulated positively by FGFs and negatively by bone morphogenetic proteins (BMPs). As chondrogenic differentiation starts, Fz4 becomes intensely up regulated in the aggregate and in the developing perichondrium, whereas transcripts of Wnt5a are excluded from the core of the aggregate but maintained in the surrounding mesenchyme and perichondrium. In addition, at this stage, the expression of both genes become positively regulated by BMPs. These changes in expression and regulation are coincident with the induction of Frzb1 in the chondrogenic aggregate, which is expressed under the positive control of BMPs. Our findings fit with a role of Wnt5a/Fz4 negatively regulating in vivo the initiation and progression of cartilage differentiation. In vitro, only Frzb1 is expressed and regulated in a manner resembling that observed in vivo. Wnt5a and Fz4 are both expressed in the differentiating mesenchyme of micromass cultures, but their expression is not significantly regulated by the addition of FGF-2 or BMP-7 to the culture medium. PMID- 12112462 TI - Regulation of cadherin junctions during mouse submandibular gland development. AB - Submandibular gland (SMG) development involves branching morphogenesis of the salivary epithelium into the surrounding mesenchyme, accompanied by proliferation and differentiation of immature salivary cells along acinar and ductal cell lineages. During development, salivary cell sorting and cell-cell adhesion are likely to be directed by cadherin adhesion receptors. We show that two classic cadherins, N- and E-cadherin, participate in SMG development. Early in embryonic morphogenesis, both cadherins displayed diffuse staining with regionalized localization to cell-cell borders. At this stage, significant pools of N- and E cadherins were Triton-soluble, suggesting that fractions of these molecules were not localized to stable junctional complexes associated with the actin cytoskeleton. With cytodifferentiation, cadherins became progressively Triton insoluble, and this correlated with their organization at cell-cell interfaces. In the cytodifferentiated SMG, N-cadherin was absent, whereas E-cadherin remained at cell-cell interfaces. Early in morphogenesis, beta-catenin was also primarily Triton-soluble, and its association with the actin cytoskeleton and localization to the adherens junctions increased with cytodifferentiation. Greater recruitment of cadherins and beta-catenin to cell-cell borders was paralleled by changes in membrane association of two Rho GTPases, Cdc42 and RhoA. N-cadherin was detected only at early stages of postnatal development, whereas E-cadherin and beta catenin became progressively Triton-insoluble during differentiation. Our results indicate that N-cadherin functions transiently in SMG development. On the other hand, E-cadherin and beta-catenin appear to play different roles during tissue organization and cytodifferentiation. In early morphogenesis, E-cadherin and beta catenin are likely to participate in SMG remodeling, whereas during cytodifferentiation, they form stable cell-cell contacts, and may collaborate with Rho GTPases in the establishment and maintenance of salivary cell polarity. PMID- 12112463 TI - Zebrafish Smad7 is regulated by Smad3 and BMP signals. AB - Growth factors of the TGF-beta superfamily such as BMPs and Nodals are important signaling factors during all stages of animal development. Smad proteins, the cytoplasmic mediators of most TGF-beta signals in vertebrates, play central roles not only for transmission but also in controlling inductive TGF-beta signals by feedback regulation. Here, we describe cloning, expression pattern, transcriptional regulation, and functional properties of two novel zebrafish Smad proteins: the TGF-beta agonist Smad3b, and the anti-Smad Smad7. We show that zebrafish Smad3b, in contrast to the related zebrafish Smad2, can induce mesoderm independently of TGF-beta signaling. Although mammalian Smad3 was shown to inhibit expression of the organizer-specific genes goosecoid, zebrafish smad3b activates organizer genes such as goosecoid. Furthermore, we show that Smad3 and BMP signals activate smad7. Because Smad7 blocks distinct TGF-beta signals in early zebrafish development, our data provide hints for new roles of smad3 genes in the regulation and modulation of TGF-beta signals. In summary, our analyses point out differences of Smad3b and Smad2 functions in zebrafish and provide the first link of smad3 and smad7 function in context of vertebrate development. PMID- 12112464 TI - Pax3 and Dach2 positive regulation in the developing somite. AB - In vertebrates, skeletal muscles of the body arise from cells of somitic origin. Recently, somite culture experiments have identified a set of genes, including Pax3, Six1, Eya2, and Dach2, that appear to play an important role in early myogenesis during somite development (Heanue et al. [1999] Genes Dev. 13:3231 3243). In somite culture Pax3, Six1, Eya2, and Dach2 not only function to activate myogenesis, but they form a complex network regulating each other's transcription. We sought to examine whether this putative Pax3/Six1/Eya2/Dach2 network of regulation actually functions in vivo. In particular, we tested whether Pax3 and Dach2 participate in a positive regulatory feedback loop in vivo as they do in culture. To test in vivo Pax3/Dach2 interregulation, we took advantage of the known dependence of both factors on ectodermal signals. Somites isolated from the overlying ectoderm lose expression of Pax3 and Dach2. Therefore, we attempted to rescue Pax3 or Dach2 expression in somites isolated from the ectoderm by retroviral misexpression of the complementary factor. Indeed misexpression of Pax3 or Dach2 resulted in rescue of Dach2 or Pax3, respectively. These rescue experiments demonstrate that Pax3 and Dach2 positively regulate each other's expression in vivo and support the validity of the Pax3/Six1/Eya2/Dach 2 network in regulating myogenesis. PMID- 12112465 TI - Expression of ErbB3, ErbB4, and neuregulin-1 mRNA during tooth development. AB - The receptor tyrosine kinases ErbB3 and ErbB4, which bind to various variants of neuregulin (NRG), play fundamental roles in neural development and in organs, which form through epithelial-mesenchymal interactions. Here, we demonstrate that NRG-1 and the receptors ErbB3 and ErbB4 are expressed locally during rodent tooth development. However, the mRNA expression patterns of ErbB3 and ErbB4 were distinctly different during odontogenesis. Examinations of teeth in genetically heart-rescued ErbB4-/- mice did not reveal any obvious deviation from the normal phenotype. The results suggest that ErbB3 and ErbB4 may participate in tooth morphogenesis. The specific interactions between NRG isoforms and ErbB receptors during this process remain to be determined. PMID- 12112466 TI - Proliferation and growth factor expression in abnormally enlarged placentas of mouse interspecific hybrids. AB - It has been shown previously that abnormal placental growth occurs in crosses and backcrosses between different mouse (Mus) species. In such crosses, late gestation placentas may weigh between 13 and 848 mg compared with a mean placental weight of approximately 100 mg in late gestation M. musculus intraspecific crosses. A locus on the X-chromosome was shown to segregate with placental dysplasia. Thus in the (M. musculus x M. spretus)F1 x M. musculus backcross, placental hyperplasia cosegregates with a M. spretus derived X chromosome. Here we have investigated whether increased cell proliferation and aberrant expression of two genes that are involved in placental growth control, Igf2 and Esx1, may cause, or contribute to placental hyperplasia. Increased bromodeoxyuridine labeling of nuclei, reflecting enhanced proliferation, was indeed observed in hyperplastic placentas when compared with normal littermate placentas. Also, increased expression of Igf2 was seen in giant cells and spongiotrophoblast. However, when M. musculus x M. spretus F1 females were backcrossed with males that were heterozygous for a targeted mutation of the Igf2 gene, placentas that carried a M. spretus derived X-chromosome and were negative for a functional Igf2 allele exhibited an intermediate placental phenotype. Furthermore, in early developmental stages of placental hyperplasia, we observed a decreased expression of the X-chromosomal Esx1 gene. This finding suggests that abnormal expression of both Igf2 and Esx1 contributes to abnormal placental development in mouse interspecific hybrids. However, Esx1 is not regulated by IGF2. PMID- 12112467 TI - Characterization of transgene expression and Cre recombinase activity in a panel of Thy-1 promoter-Cre transgenic mice. AB - The regulatory elements of the murine Thy1.2 gene were used to drive Cre recombinase expression in the nervous system (NS) of transgenic mice. Eleven Thy1 Cre lines exhibited transgene expression in several regions of the central and peripheral nervous systems, including the cerebral cortex, cerebellum, spinal cord, retina, and dorsal root ganglion. Thy1-Cre expression also resulted in region-specific activation of Cre reporter transgenes. Although Thy-1 expression is normally initiated in postmitotic neurons in the perinatal period, we show that the Thy-1.2 expression cassette drives Cre expression in immature proliferative zones in the NS as early as embryonic day 11 and in non-neural tissue. The Thy1.2 transgene cassette, therefore, does not impart transgene expression that is restricted to the NS or to postmitotic neurons within the NS. This panel of Thy1-Cre transgenic mice, however, will be useful reagents for the ablation of genes whose transcripts are spatially or temporally restricted in the developing NS. PMID- 12112468 TI - Mesenchymal expression of vascular endothelial growth factors D and A defines vascular patterning in developing lung. AB - The lung has specific vascular patterning requirements for effective gas exchange at birth, including alignment of airways and blood vessels and lymphatic vessels. Vascular endothelial growth factors (VEGF) are potent effectors of vascular development. We examined the temporal and spatial expression of VEGF-D and specific VEGF-A isoforms at each stage of lung development. VEGF-D, expressed only by cadherin-11-positive cells of the mesenchyme, is first detected at embryonic day (E) 13.5, a period of active vasculogenesis. VEGFR-3, its cognate receptor, is detected earlier on days E11.5 to E14.5, in both blood vessels and lymphatic vessels and later, on day E17.5, in only lymphatic vessels. VEGF-A is expressed in the mesenchyme throughout lung development and also by the epithelium midway through organogenesis. Before E14, the predominant forms of VEGF-A are the soluble isoforms, VEGF-A120 and 164. Not until E14.5 do epithelial cells at the tips of expanding airways express VEGF-A, including VEGF-A188, an isoform with high affinity for extracellular matrix. Our results demonstrate unique temporal and spatial expression of VEGF-D and specific VEGF-A isoforms during lung development and suggest these related factors have distinct functions in vascular and lymphatic patterning of the lung. PMID- 12112470 TI - Characterizing gene expression during lens formation in Xenopus laevis: evaluating the model for embryonic lens induction. AB - Few directed searches have been undertaken to identify the genes involved in vertebrate lens formation. In the frog Xenopus, the larval cornea can undergo a process of transdifferentiation to form a new lens once the original lens is removed. Based on preliminary evidence, we have shown that this process shares many elements of a common molecular/genetic pathway to that involved in embryonic lens development. A subtracted cDNA library, enriched for genes expressed during cornea-lens transdifferentiation, was prepared. The similarities/identities of specific clones isolated from the subtracted cDNA library define an expression profile of cells undergoing cornea-lens transdifferentiation ("lens regeneration") and corneal wound healing (the latter representing a consequence of the surgery required to trigger transdifferentiation). Screens were undertaken to search for genes expressed during both transdifferentiation and embryonic lens development. Significantly, new genes were recovered that are also expressed during embryonic lens development. The expression of these genes, as well as others known to be expressed during embryonic development in Xenopus, can be correlated with different periods of embryonic lens induction and development, in an attempt to define these events in a molecular context. This information is considered in light of our current working model of embryonic lens induction, in which specific tissue properties and phases of induction have been previously defined in an experimental context. Expression data reveal the existence of further levels of complexity in this process and suggests that individual phases of lens induction and specific tissue properties are not strictly characterized or defined by expression of individual genes. PMID- 12112469 TI - Mouse PeP: a novel peroxisomal protein linked to myoblast differentiation and development. AB - The identification of several peroxisomal proteins in the past decade has deepened our understanding of the biology of peroxisomes and their involvement in human disorders. We report the cloning and expression pattern during the mouse development of a cDNA encoding a novel protein, named PeP, and show that its product is imported specifically to the peroxisome matrix in a variety of cell types. We also demonstrate that PeP is imported to the organelle through the PEX5 receptor pathway, which indicates that the C-terminal tripeptide SKI behaves as a type 1 peroxisomal targeting signal (PTS1). PeP expression is tightly regulated, as shown by Northern and in situ hybridization experiments. Thus during embryonic development in the mouse, PeP mRNA is detected almost exclusively in the skeletal muscle, whereas in adult mice, strong expression is also found in the heart and brain. In addition, PeP mRNA accumulation is induced after myoblast differentiation in vitro, when myotube formation is promoted. Sequence analysis reveals that PeP has no significant homology to any known protein, except for a short stretch of amino acids containing the fingerprint of the fibronectin type III superfamily, a domain present in proteins often related to molecular and cellular recognition and binding processes. Thus our data suggest a connection between the function of PeP and murine cell differentiation and development. PMID- 12112471 TI - Expression of differentially expressed nucleolar transforming growth factor-beta1 target (DENTT) in adult mouse tissues. AB - Differentially expressed nucleolar TGF-beta1 target (DENTT) is a novel member of the TSPY/TSPY-L/SET/NAP-1 (TTSN) superfamily that we have previously identified in human lung cancer cells. Here, we have investigated the expression of this protein in the adult mouse. By Western analysis, DENTT is highly expressed in the pituitary gland and moderately in the adrenals, brain, testis, and ovary. Immunohistochemical staining analysis for DENTT showed differential cytoplasmic and nuclear staining patterns in several cell types. The pituitary gland showed the highest level of immunostaining for DENTT, with strong cytoplasmic immunoreactivity in the anterior lobe, moderate levels in the posterior lobe, and a few cells showing nuclear staining in the intermediate lobe. In contrast, the intermediate lobe of the pituitary showed intense cytoplasmic staining for TGF beta1. Nuclear and cytoplasmic staining for DENTT was present in the islets of Langerhans in the pancreas. Cytoplasmic staining for DENTT was particularly intense in the cortex of the adrenal gland, whereas the medulla showed weak nuclear staining. In the nervous system, the choroid plexus showed the highest immunoreactivity, with cortical motoneurons and Purkinje cells having relatively high levels of staining for DENTT as well. DENTT immunoreactivity was found in Leydig interstitial cells, Sertoli cells, and primary spermatocytes in the testis. In the female reproductive system, DENTT immunoreactivity was present in oocytes, thecal cells, and corpora lutea. The bronchial epithelium of the lung showed moderate levels of staining for DENTT localized to the cell nucleus. Additionally, three rodent pituitary cell lines (AtT20, GH3, and alphaT3-1, representing corticotropes, lactotropes, and gonadotropes, respectively) showed expression of DENTT. Addition of TGF-beta1 or serum to AtT20 cells increased DENTT protein production by 4 hr and, after reaching maximal levels at 2.4-fold above basal level by 8 hr, decreased, whereas no more than a 1.5-fold increase in DENTT protein occurred in GH3 or alphaT3-1 cells. Transient transfection studies showed that ectopic DENTT expression significantly increased the level of p3TP Lux reporter transcription in AtT20 cells, but not in GH3 or alphaT3-1 cells. Interestingly, addition of TGF-beta1 had no significant effect on the ability of DENTT expression to influence p3TP-Lux reporter transcription in AtT20 cells. This report is the first detailed immunohistochemical examination of a member of the TTSN superfamily in the adult mouse. Expression of DENTT in endocrine tissues, nervous system, lung, oocytes, and thecal cells, in addition to the testis, suggests new roles for the TTSN superfamily. The differential patterns of expression of DENTT and TGF-beta1 in some tissues, including the pituitary, suggest that other factors are likely to be regulators of DENTT besides TGF beta1. PMID- 12112472 TI - Molecular characterization of Calymmin, a novel notochord sheath-associated extracellular matrix protein in the zebrafish embryo. AB - During the screening of a zebrafish postsomitogenesis embryo cDNA library, we have identified a cDNA corresponding to a novel type of protein localized to the notochordal sheath-associated extracellular matrix (ECM) of the embryo. The 4.049 kb mRNA encodes a predicted polypeptide of 1,207 amino acids (122 kDa, pI 10.50) with a potential signal peptide of 20 amino acids. After the signal peptide, the mature protein consists of 1,187 amino acids (119 kDa, pI 10.46), for which the name "Calymmin" (from Greek chialphalambdanumumualpha, to envelop, to cover) is proposed. The Calymmin mRNA is highly and transiently expressed by the notochord cells of the embryo from the 10- to 12-somite stage to the pharyngula period (13 and 24 hours postfertilization, respectively), and light and electron microscopical immunolocalization analysis revealed that the protein was specifically localized within a granular and filamentous layer of the ECM compartment surrounding the notochord. In zebrafish no tail mutants (ntl(tc41)), in which the notochord precursor cells are present but fail to differentiate, the Calymmin protein was not detected, confirming the notochord origin of Calymmin. These results indicate that Calymmin is a novel constitutive protein of the ECM compartment associated to the perinotochordal sheath in the zebrafish embryo, which is specifically expressed by the differentiating notochord cells. PMID- 12112473 TI - Induction of chondrogenesis in neural crest cells by mutant fibroblast growth factor receptors. AB - Activating mutations in human fibroblast growth factor receptors (FGFR) result in a range of skeletal disorders, including craniosynostosis. Because the cranial bones are largely neural crest derived, the possibility arises that increased FGF signalling may predispose to premature/excessive skeletogenic differentiation in neural crest cells. To test this hypothesis, we expressed wild-type and mutant FGFRs in quail embryonic neural crest cells. Chondrogenesis was consistently induced when mutant FGFR1-K656E or FGFR2-C278F were electroporated in ovo into stage 8 quail premigratory neural crest, followed by in vitro culture without FGF2. Neural crest cells electroporated with wild-type FGFR1 or FGFR2 cDNAs exhibited no chondrogenic differentiation in culture. Cartilage differentiation was accompanied by expression of Sox9, Col2a1, and osteopontin. This closely resembled the response of nonelectroporated neural crest cells to FGF2 in vitro: 10 ng/ml induces chondrogenesis, Sox9, Col2a1, and osteopontin expression, whereas 1 ng/ml FGF2 enhances cell survival and Sox9 and Col2a1 expression, but never induces chondrogenesis or osteopontin expression. Transfection of neural crest cells with mutant FGFRs in vitro, after their emergence from the neural tube, in contrast, produced chondrogenesis at a very low frequency. Hence, mutant FGFRs can induce cartilage differentiation when electroporated into premigratory neural crest cells but this effect is drastically reduced if transfection is carried out after the onset of neural crest migration. PMID- 12112474 TI - Effect of elevated homocysteine on cardiac neural crest migration in vitro. AB - A positive correlation between elevated maternal homocysteine (Hcys) and an increased risk of neural tube, craniofacial, and cardiac defects is well known. Studies suggest Hcys perturbs neural crest (NC) development and may involve N methyl-D-aspartate (NMDA) receptors (Rosenquist et al., 1999). However, there is no direct evidence that Hcys alters NC cell behavior. Here, we evaluated the effect of Hcys on cardiac NC cell migratory behavior in vitro. Neural tube segments from chick embryos treated in ovo with or without Hcys were placed in culture and the migratory behavior of emigrating NC cells was monitored. Hcys significantly increased in vitro NC cell motility at all embryonic stages examined. NC cell surface area and perimeter were also increased. However, the relative distance NC cells migrated from their original starting point only increased in NC cells treated in ovo at stage 6 or at the time neural tube segments were cultured. Cysteine had no effect. NMDA mimicked Hcys' effect on NC motility and migration distance but had no effect on cell area or perimeter. The noncompetitive inhibitor of NMDA receptors, MK801+, significantly inhibited NC cell motility, reduced migration distance, and also blocked the effects of NMDA and Hcys on NC motility and migratory distance in vitro. A monoclonal antibody directed against the NMDA receptor immunostained NC cells in vitro and, in western blots, bound a single protein with the appropriate molecular weight for the NMDA receptor in NC cell lysates. These data are consistent with the hypothesis that a Hcys-sensitive NMDA-like receptor is expressed by early emigrating NC cells or their precursors, which is important in mediating their migratory behavior. Perturbation of this receptor may be related to some of the teratogenic effects observed with elevated Hcys. PMID- 12112475 TI - Coexpression of SCL and GATA3 in the V2 interneurons of the developing mouse spinal cord. AB - The differentiation of neural progenitors into the many classes of neurons that exist in the mature spinal cord is a process that relies heavily on the activation of precise combinations of transcription factors. Defining these transcription factor combinations is an important aspect of research in developmental neurobiology that promises to provide incredible insights into the structure, function, and pathology of the central nervous system. The present study aimed to investigate a possible role for the Stem Cell Leukemia (SCL) gene, a basic helix-loop-helix (bHLH) transcription factor gene, in the specification of a population of neural cells in the ventral neural tube. Section RNA in situ hybridisation revealed that SCL is transiently expressed within the V2 postmitotic domain of the developing mouse spinal cord between 10.5 and 13.5 days post coitum. Double-immunofluorescence experiments were subsequently carried out to directly compare the expression of SCL with other V2-specific markers at the cellular level. These experiments revealed that SCL is expressed in a medially restricted subpopulation of GATA-3 producing cells, suggesting a possible role for this factor in the differentiation of the GATA population of V2 interneurons. PMID- 12112476 TI - Cell death along the embryo midline regulates left-right sidedness. AB - During embryogenesis, left-right sidedness is established by asymmetric expression of laterality genes. A recent model predicts the presence of a functional midline that divides the left side of the embryonic disc from the right side, separating left- and right-inducing signals. We show evidence that this midline is formed from a distinct population of cells within the primitive streak. Cells in the dorsal midline of the chick primitive streak display unique expression of the gastrulation markers fgf-8 and brachyury. These midline cells are fated to die, and dead cells remain in the midline during gastrulation. Inhibition of midline cell death compromises the early expression of laterality genes, such as shh and nodal and randomizes the direction of heart looping. We suggest that cell death along the primitive streak midline is a novel mechanism involved in the regulation of left-right asymmetry during early embryogenesis. PMID- 12112477 TI - Mouse alpha1(I)-collagen promoter is the best known promoter to drive efficient Cre recombinase expression in osteoblast. AB - Cell- and time-specific gene inactivation should enhance our knowledge of bone biology. Implementation of this technique requires construction of transgenic mouse lines expressing Cre recombinase in osteoblasts, the bone forming cell. We tested several promoter fragments for their ability to drive efficient Cre expression in osteoblasts. In the first mouse transgenic line, the Cre gene was placed under the control of the 2.3-kb proximal fragment of the alpha1(I) collagen promoter, which is expressed at high levels in osteoblasts throughout their differentiation. Transgenic mice expressing this transgene in bone were bred with the ROSA26 reporter (R26R) strain in which the ROSA26 locus is targeted with a conditional LacZ reporter cassette. In R26R mice, Cre expression and subsequent Cre-mediated recombination lead to expression of the LacZ reporter gene, an event that can be monitored by LacZ staining. LacZ staining was detected in virtually all osteoblasts of alpha1(I)-Cre;R26R mice indicating that homologous recombination occurred in these cells. No other cell type stained blue. In the second line studied, the 1.3-kb fragment of osteocalcin gene 2 (OG2) promoter, which is active in differentiated osteoblasts, was used to drive Cre expression. OG2-Cre mice expressed Cre specifically in bone. However, cross of OG2-Cre mice with R26R mice did not lead to any detectable LacZ staining in osteoblasts. Lastly, we tested a more active artificial promoter derived from the OG2 promoter. The artificial OG2-Cre transgene was expressed by reverse transcriptase-polymerase chain reaction in cartilage and bone samples. After cross of the artificial OG2-Cre mice with R26R mice, we detected a LacZ staining in articular chondrocytes but not in osteoblasts. Our data suggest that the only promoter able to drive Cre expression at a level sufficient to induce recombination in osteoblasts is the alpha1(I)-collagen promoter. PMID- 12112478 TI - Cancer patients' perceptions of do not resuscitate orders. AB - Patients' perceptions of do not resuscitate (DNR) orders and how and when to present the information were sought to aid in framing DNR policy. Semi-structured interviews of 23 patients being treated for cancer, were conducted by a clinical psychologist. The interviews were transcribed and analysed with the aid of a qualitative software package. Discourse analysis enabled hypotheses to be formed based on consistencies and variations of the language used. Most patients understood what DNR meant and preferred DNR orders to 'good palliative care' orders. They saw it as their autonomous right and responsibility to make such decisions. They would seek information on the likely medical outcomes of resuscitation but also would use non-rational criteria based on emotional and social factors to make their decisions. Family considerations suggest that personal autonomy is not the overriding basis of the decision. Patients were unsure of the best timing of a DNR discussion and were prepared to defer to doctors' intuition. Most advocated written DNR orders but few had them. Families were construed as advocates but also seen as constraining individual autonomy. When considering DNR orders, patients recognise the diversity of preferences likely to exist that belie a one policy fits all approach. PMID- 12112479 TI - Employment patterns of long-term cancer survivors. AB - As more people are diagnosed at earlier stages and surviving cancer, they are increasingly likely to be at working ages, where issues regarding productivity and employment continuation must be addressed by patients and employers alike. To this end, we studied the employment patterns of 253 long-term cancer survivors in the Detroit Metropolitan Area. Of those working at the time of their initial diagnosis, 67% were employed 5-7 years later. Patients who stopped working did so because they retired (54%), were in poor health/disabled (24%), quit (4%), cited other reasons (9%), or their business closed (9%). Many employed patients worked in excess of 40 h per week although some reported various degrees of disability that interfered with job performance. Overall, the ability of cancer patients to continue employment appears optimistic. PMID- 12112480 TI - Health status and life satisfaction among breast cancer survivor peer support volunteers. AB - Two measures of health-related quality of life (HRQOL), the Medical Outcomes Survey Short Form 36 (SF-36) and the Satisfaction with Life Domains Scale for Cancer (SLDS-C), were compared to examine the relationship between health status and life satisfaction among breast cancer survivors (BCSs). A total of 586 BCSs, all of whom were volunteers in peer support programs, met inclusion criteria and completed the self-report measures. Significant correlation coefficients were shown between life satisfaction and measures of health status. SF-36 scores were significantly higher for physical functioning, emotional well-being, and vitality subscales compared to population norms. BCSs expressed greatest dissatisfaction with their sexual ability, physical strength, and bodies in general. Small age differences were found. Results suggest that incorporating multiple measures of HRQOL contribute to the understanding and measurement of the effects of cancer on perceived health status and life satisfaction. PMID- 12112481 TI - Communication between physicians and cancer patients about complementary and alternative medicine: exploring patients' perspectives. AB - The aim of this paper is to identify barriers to communication between physicians and cancer patients regarding complementary and alternative medicine (CAM) by exploring the perspectives of patients. In face of the recent popularity of CAM use among cancer patients, the lack of communication is a serious problem. A number of CAM therapies may interfere with conventional treatments and thus impact patients' well-being and chances of survival. In addition, lack of communication is problematic in the health care context because the development of openness and trust between health care providers and clients is contingent upon effective interpersonal communication. We conducted semi-structured interviews with 143 cancer patients to explore their experiences with CAM use. Using a qualitative research method, we examined interview data from 93 CAM users who provided sufficient information about communication issues. As a result, three themes emerged describing barriers to unsuccessful communication as perceived from the patient's point of view: physicians' indifference or opposition toward CAM use, physicians' emphasis on scientific evidence, and patients' anticipation of a negative response from their physician. Increasing education about CAM and regular assessment of CAM use may help physicians to be more aware of their patients' CAM use. As a result, physicians may provide patients with information on risks and benefits of CAM use and refer patients to other services that may address unmet needs. Given a difference in epistemiologic beliefs about cancer and its treatment, the challenge is to find a common ground for an open discussion in which physicians consider that scientific evidence is not all that counts in the life of an individual facing a serious disease. PMID- 12112482 TI - A structural model of the relationships among self-efficacy, psychological adjustment, and physical condition in Japanese advanced cancer patients. AB - We made detailed research for relationships among physical condition, self efficacy and psychological adjustment of patients with advanced cancer in Japan. The sample consisted of 85 (42 males and 43 females) advanced cancer patients. Interviews were conducted with some measurement scales including the Self efficacy scale for Advanced Cancer (SEAC), and the Hospital Anxiety and Depression Scale (HADS). Karnofsky Performance Status (KPS) and medication status were also recorded from the evaluation by physicians. We used structural equation modeling (SEM) for statistical analysis. The analysis revealed that the model, including three self-efficacy subscales, depression, anxiety, KPS, meal-, liquid intake, prognosis and three latent variables: 'Self-efficacy', 'Emotional Distress', and 'Physical Condition,' fit the data (chi-square(24)=28.67, p=0.23; GFI=0.93; CFI=0.98; RMSEA=0.05). In this model, self-efficacy accounted for 71% of the variance in emotional distress and physical condition accounted for 8% of the variance in self-efficacy. Overall, our findings suggest clearly that close relationships existed among physical condition, self-efficacy and emotional distress. That is, patients in good physical condition had a high self-efficacy, and patients with high self-efficacy were less emotionally distressed. These results imply that psychological intervention which emphasizes self-efficacy would be effective for advanced cancer patients. PMID- 12112483 TI - Predicting major depression in brain tumor patients. AB - Very few studies have been performed utilizing DSM criteria to diagnose major depressive disorder (MDD) in adult brain tumor patients. This study aimed to diagnose MDD in this population using DSM-IV criteria.Eighty-nine adult brain tumor patients were examined in an ambulatory neuro-oncology clinic setting using a structured psychiatric interview which followed current DSM-IV diagnostic criteria for MDD. This sample was interviewed and evaluated on a one-time basis. The patients were referred for evaluation on a consecutive basis. Multiple regression was used to model critical independent variables to predict MDD.Twenty eight percent of the sample (N=89) were found to have major depressive disorder using DSM-IV criteria. Key predictors of MDD included frontal region of tumor location (p=0.001), combined sadness and lack of motivation symptoms (p=0.0001), and family psychiatric history (p=0.006). The multiple regression models account for 37% of variance in predicting MDD (R(2)=0.37).A substantially higher incidence of MDD was found in this sample of adult brain tumor patients compared with other adult, ambulatory cancer patients previously evaluated with DSM criteria. The incidence of MDD was about triple that found in other published studies using DSM criteria. PMID- 12112484 TI - Individual quality of life in patients with leukaemia and lymphoma. AB - With increasingly sophisticated chemotherapy regimes being prescribed the quality of life of cancer patients has become a key outcome measure. Little has been reported concerning the experience of patients with haematological malignancy receiving chemotherapy. The objective of this study was to evaluate the clinical usefulness of a novel quality of life measure-the Schedule for the Evaluation of Individual Quality of Life-Direct Weighting (SEIQoL-DW) in a sample of patients with either leukaemia or lymphoma. Fifty-one patients from the haematology clinic and in-patient unit at The Royal Devon and Exeter Hospital completed the SEIQoL DW; in addition, each patient completed the Hospital Anxiety and Depression Scale (HADS) and a ten item questionnaire covering aspects of their treatment and satisfaction with information provided. The practical application of the SEIQoL DW is described and two patients quality of life profiles are illustrated for comparison. The relationship between quality of life, satisfaction with information provided and psychological distress as measured by the HADS is discussed. PMID- 12112485 TI - Pulsed electrostatic fields (ETG) to reduce hair loss in women undergoing chemotherapy for breast carcinoma: a pilot study. AB - AIMS: To determine whether specific pulsed electrostatic fields, or electrotrichogenesis (ETG), could potentially prevent or reduce hair loss in patients undergoing adjuvant cyclophosphamide, methotrexate and 5-fluorouracil (CMF) chemotherapy for breast cancer. METHODS: Thirteen women were followed during their adjuvant ETG and chemotherapy treatment to determine the efficacy of ETG. All patients were treated for 12 min, twice weekly with a pulsed electrostatic field. Quantitative hair loss was measured by photographic assessment, and manual hair count. Quality of life assessment was conducted at the end of the study. RESULTS: Twelve out of 13 participants had good hair retention throughout the chemotherapy period and afterwards. There were no reported side effects attributable to ETG. CONCLUSIONS: This study shows encouraging results in an area where no other appropriate treatment is available Reducing alopecia, secondary to chemotherapy has the potential to increase CMF treatment compliance, enhance patient self-esteem, and improve overall quality of life during this stressful period. PMID- 12112486 TI - The EORTC QLQ-C30 (version 3.0) Quality of Life questionnaire: validation study for Spain with head and neck cancer patients. AB - The EORTC Quality of Life Study Group has developed a questionnaire for evaluating Quality of Life in international clinical trials: QLQ-C30. The purpose of the present work is to validate the third version of this questionnaire (3.0) for use in Spain. Two hundred and one head and neck cancer patients completed the QLQ-C30 at one or two time points during the treatment and follow-up periods, and a subsample completed the questionnaire on three occasions. Psychometric evaluation of the structure, reliability and validity of the questionnaire was undertaken. The data support the structure and the reliability of the scales. Validity was confirmed in three ways: the interscale correlations are statistically significant and moderate, several scales and items discriminate among groups of patients with different scores on the clinical variables, and some scales reflect significant changes during treatment and the follow-up period. The EORTC QLQ-C30 (version 3.0) is a reliable and valid questionnaire when applied to a sample of Spanish head and neck cancer patients. These results are in line with previous studies. PMID- 12112488 TI - Does prospective payment reduce inpatient length of stay? AB - A change in payment mechanism for inpatient care from per diem to per episode creates two incentives - a marginal and an average price effect - to change length of stay. The decrease in marginal price per day to zero should reduce the length of stay, while an increase in average price per inpatient stay should increase the length of stay. This study uses data from a natural experiment to estimate both marginal and average price elasticities, and to test whether the length of stay falls after the introduction of prospective payment in a sample of 8509 severely mentally ill patients. We estimate that the marginal price elasticity is zero, but the average price elasticity is between 0.16 and 0.20. The results were generally robust for short- and long stayers, and for persons admitted early and late after the change in payment mechanism. The model controlled for hospital fixed effects and individual random effects. PMID- 12112489 TI - The benefits of switching smoking cessation drugs to over-the-counter status. AB - This paper provides an analysis of the benefits to society from the conversion of nicotine replacement drugs (nicotine patches and gum) in 1996 from sale by prescription only in the United States to over-the-counter (OTC) sales. To estimate these benefits, we first estimate statistical demand functions for nicotine patches and gum. Second, we calculate the effects of OTC conversion on sales of each type of nicotine replacement drug. Third, we survey the literature on the effects of nicotine replacement drugs on total quits of cigarette smoking. Fourth, we survey the literature on the effects of quits achieved on expected lifespan, and on the estimated monetary value of longer lives from smoking cessation. Finally, we use all this evidence to calculate the value of the social benefits of the OTC conversion to the US. As a result of the OTC conversion, consumption of nicotine replacement drugs has increased substantially, by 78-92% for nicotine patches and 180% for nicotine gum. We estimate that the resulting increase in smoking cessation generated annual net social benefits of the order of magnitude of 1.8-2 billion dollars, based on conservative estimates both of the number of quits achieved and the value of added quality-adjusted life years from the reduced smoking. PMID- 12112490 TI - The impact of diabetes on adult employment and earnings of Mexican Americans: findings from a community based study. AB - Epidemiological studies indicate that minority populations in the US - including African Americans, Native Americans and Mexican Americans - are particularly at risk for diabetes and that their complications are more frequent and severe. Using microdata from a 1994-1999 population based study of middle aged and older Mexican Americans in the Southwest, this study analyzes the impact of diabetes on the employment and earnings outcomes of adults 45 years of age and older. The empirical results from estimating maximum likelihood employment and earnings models suggest that diabetes leads to lower productivity and earnings for women but has no statistically significant impact on their employment probability. In the case of men, however, diabetes leads to a lower employment propensity but has no effect on earnings. Thus, the problems associated with this condition could lead to potential future financial difficulties particularly for high-risk populations in their later years. PMID- 12112491 TI - Something old, something new, something borrowed, something blue: a framework for the marriage of health econometrics and cost-effectiveness analysis. AB - Economic evaluation is often seen as a branch of health economics divorced from mainstream econometric techniques. Instead, it is perceived as relying on statistical methods for clinical trials. Furthermore, the statistic of interest in cost-effectiveness analysis, the incremental cost-effectiveness ratio is not amenable to regression-based methods, hence the traditional reliance on comparing aggregate measures across the arms of a clinical trial. In this paper, we explore the potential for health economists undertaking cost-effectiveness analysis to exploit the plethora of established econometric techniques through the use of the net-benefit framework - a recently suggested reformulation of the cost effectiveness problem that avoids the reliance on cost-effectiveness ratios and their associated statistical problems. This allows the formulation of the cost effectiveness problem within a standard regression type framework. We provide an example with empirical data to illustrate how a regression type framework can enhance the net-benefit method. We go on to suggest that practical advantages of the net-benefit regression approach include being able to use established econometric techniques, adjust for imperfect randomisation, and identify important subgroups in order to estimate the marginal cost-effectiveness of an intervention. PMID- 12112492 TI - Social risk management options for medical care in Indonesia. AB - This paper investigates the extent to which price subsidies for medical care are a suitable instrument to reduce household's exposure to catastrophic financial risks associated with ill-health in Indonesia. Using the 1995 SUSENAS household survey, the observed distribution of user fees and health expenditures is used to derive a distribution of 'needed' medical expenditures. The trade-off between the tax burden and effectiveness in reducing the exposure to catastrophic risk is analyzed for two existing price regimes along with a number of hypothetical regimes. We find that the existing regimes significantly reduce the exposure to catastrophic shocks but do not eliminate them. Simulations suggest that further reductions could be achieved if a larger proportion of government subsidies were directed to inpatient care. Subsidizing outpatient treatment is a cost effective policy to reduce exposure to catastrophic risks only for the very poor. PMID- 12112493 TI - A new explanation for the difference between time trade-off utilities and standard gamble utilities. AB - This paper gives a new explanation for the systematic disparity between standard gamble (SG) utilities and time trade-off (TTO) utilities. The common explanation, which is based on expected utility, is that the disparity is caused by curvature of the utility function for duration. This explanation is, however, incomplete. People violate expected utility and these violations lead to biases in SG and TTO utilities. The paper analyzes the impact on SG and TTO utilities of three main reasons why people violate expected utility: probability weighting, loss aversion, and scale compatibility. In the SG, the combined effect of utility curvature, probability weighting, loss aversion, and scale compatibility is an upward bias. In the TTO these factors lead both to upward and to downward biases. This analysis can also explain the tentative empirical finding that the TTO better describes people's preferences for health than the SG. PMID- 12112494 TI - Using stated preference discrete choice modelling to evaluate the introduction of varicella vaccination. AB - Applications of stated preference discrete choice modelling (SPDCM) in health economics have been used to estimate consumer willingness to pay and to broaden the range of consequences considered in economic evaluation. This paper demonstrates how SPDCM can be used to predict participation rates, using the case of varicella (chickenpox) vaccination. Varicella vaccination may be cost effective compared to other public health programs, but this conclusion is sensitive to the proportion of the target population immunised. A choice experiment was conducted on a sample of Australian parents to predict uptake across a range of hypothetical programs. Immunisation rates would be increased by providing immunisation at no cost, by requiring it for school entry, by increasing immunisation rates in the community and decreasing the incidence of mild and severe side effects. There were two significant interactions; price modified the effect of both support from authorities and severe side effects. Country of birth was the only significant demographic characteristic. Depending on aspects of the immunisation program, the immunisation rates of children with Australian-born parents varied from 9% to 99% while for the children with parents born outside Australia they varied from 40% to 99%. This demonstrates how SPDCM can be used to understand the levels of attributes that will induce a change in the decision to immunise, the modification of the effect of one attribute by another, and subgroups in the population. Such insights can contribute to the optimal design and targeting of health programs. PMID- 12112495 TI - Health economics resources - an update. PMID- 12112496 TI - Reproducibility of total cerebral blood flow measurements using phase contrast magnetic resonance imaging. AB - PURPOSE: To evaluate reproducibility of total cerebral blood flow (CBF) measurements with phase contrast magnetic resonance imaging (pcMRI). MATERIALS AND METHODS: We repeated total CBF measurements in 15 healthy volunteers with and without cardiac triggering, and with and without repositioning. In eight volunteers measurements were performed at two different occasions. In addition, measurement of flow in a phantom was performed to validate MR measurements. RESULTS: A difference of 40.4 ml/minute was found between CBF measurements performed with and without triggering (P < 0.05). For repeated triggered measurements, the coefficient of variation (CV) was 7.1%, and for nontriggered measurements 10.3%. For repeated measurements with repositioning, the CV was 7.1% with and 11.2% without triggering. Repeated measurements at different occasions showed a CV of 8.8%. Comparing measured with real flow in the phantom, the triggered differed 4.9% and the nontriggered 8.3%. CONCLUSION: The findings of this study demonstrate that pcMRI is a reliable method to measure total CBF in terms of both accuracy and reproducibility. PMID- 12112497 TI - A method for the direct electrical stimulation of the auditory system in deaf subjects: a functional magnetic resonance imaging study. AB - PURPOSE: To develop a safe functional magnetic resonance imaging (fMRI) procedure for auditory assessment of deaf subjects. MATERIALS AND METHODS: A gold-plated tungsten electrode has been developed which has zero magnetic susceptibility. Used with carbon leads and a carbon reference pad, it enables safe, distortion free fMRI studies of deaf subjects following direct electrical stimulation of the acoustic nerve. Minor pickup of the radio frequency (RF) pulses by the electrode assembly is difficult to eliminate, and a SPARSE acquisition sequence is used to avoid any effects of unintentional auditory nerve stimulation. RESULTS: The procedure is demonstrated in a deaf volunteer. Activation is observed in the contralateral but not the ipsilateral primary auditory cortex. This is in sharp contrast to studies of auditory processing in hearing subjects, but consistent with the small number of previous positron emission tomography (PET) and MR studies on adult deaf subjects. CONCLUSION: The fMRI procedure is able to demonstrate whether the auditory pathway is fully intact, and may provide a useful method for preoperative assessment of candidates for cochlear implantation. PMID- 12112498 TI - Applications of neural network analyses to in vivo 1H magnetic resonance spectroscopy of Parkinson disease patients. AB - PURPOSE: To apply neural network analyses to in vivo magnetic resonance spectra of controls and Parkinson disease (PD) patients for the purpose of classification. MATERIALS AND METHODS: Ninety-seven in vivo proton magnetic resonance spectra of the basal ganglia were recorded from 31 patients with (PD) and 14 age-matched healthy volunteers on a 1.5-T imager. The PD patients were grouped as follows: probable PD (N = 15), possible PD (N = 11), and atypical PD (N = 5). Total acquisition times of approximately five minutes were achieved with a TE (echo time) of 135 msec, a TR (repetition time) of 2000 msec, and 128 scan averages. Neural network (back propagation, Kohonen, probabilistic, and radial basis function) and related (generative topographic mapping) data analyses were performed. RESULTS: Conventional data analysis showed no statistically significant differences in metabolite ratios based on measuring signal intensities. The trained networks could distinguish control from PD with considerable accuracy (true positive fraction 0.971, true negative fraction 0.933). When four classes were defined, approximately 88% of the predictions were correct. The multivariate analysis indicated metabolic changes in the basal ganglia in PD. CONCLUSION: A variety of neural network and related approaches can be successfully applied to both qualitative visualization and classification of in vivo spectra of PD patients. PMID- 12112499 TI - In vivo magnetic resonance imaging of the human cervical spinal cord at 3 Tesla. AB - PURPOSE: To demonstrate the feasibility of obtaining high-quality magnetic resonance (MR) images of the human cervical spinal cord in vivo at a magnetic field strength of 3 T and to optimize the signal contrast between gray matter, white matter, and cerebrospinal fluid (CSF) on 2D gradient recalled echo (GRE) images of the cervical spinal cord. MATERIALS AND METHODS: Using a custom-built, anatomically molded radio frequency (RF) surface coil, the repetition time and flip angle of a 2D GRE sequence were systematically varied in five volunteers to assess tissue contrast in the cervical spinal cord. RESULTS: The 2D GRE parameters for an optimal balance between gray-white matter and CSF-white matter contrast at 3 T were determined to be a time-to-repetition (TR) of 2000 msec and a flip angle of 45 degrees, with the constant short time-to-echo (TE) of 12 msec used in this study. Excellent tissue contrast and visualization of the internal anatomy of the spinal cord was demonstrated reproducibly in eight subjects using these optimal parameters. CONCLUSION: This study demonstrates that imaging the cervical spinal cord and delineating internal spinal cord structures such as gray and white matter is feasible at 3 T. PMID- 12112501 TI - Influence of oxygen flow rate on signal and T(1) changes in oxygen-enhanced ventilation imaging. AB - PURPOSE: To investigate the optimal oxygen flow rate for oxygen-enhanced MR ventilation imaging. MATERIALS AND METHODS: Using a cardiac-triggered nonselective inversion recovery (IR) half Fourier single-shot fast spin echo sequence, series of images were acquired with the subject alternately inhaling room air and 100% oxygen. Oxygen flow rates of 5 L/min, 10 L/min, 15 L/min, 20 L/min, and 25 L/min were studied, and signal intensity from the oxygen-enhanced ventilation images and T(1) of the lung were measured. RESULTS: The average signal intensity was 63.0 +/- 21.0 for 5 L/min, 98.7 +/- 26.8 for 10 L/min, 133.8 +/- 20.0 for 15 L/min, 138.7 +/- 19.7 for 20 L/min, and 139.2 +/- 37.9 for 25 L/min. The average T(1)'s of the lung were 1399 msec +/- 130 msec for room air, 1314 msec +/- 101 msec for 5 L/min, 1276 msec +/- 105 msec for 10 L/min, 1207 msec +/- 71 msec for 15 L/min, 1206 msec +/- 90 msec for 20 L/min, and 1207 msec +/- 42 msec for 25 L/min. CONCLUSION: The optimal flow rate is 15 L/min for oxygen-enhanced ventilation imaging. PMID- 12112500 TI - Imaging inflammation: direct visualization of perivascular cuffing in EAE by magnetic resonance microscopy. AB - PURPOSE: To determine if the architectural features revealed by magnetic resonance microscopy (MRM) allow one to detect microscopic abnormalities associated with neuroinflammation in fixed brain sections from animals with experimental allergic encephalomyelitis (EAE), an animal model for multiple sclerosis (MS). MATERIALS AND METHODS: Imaging was performed at the Center for In Vivo Microscopy (CIVM) using a 9.4-Tesla, 89-mm bore, superconducting magnet with actively shielded gradients capable of 850 mT/m. A number of MR contrasts and spatial resolutions were explored. RESULTS: The assessment of EAE brain showed that it is possible to visualize perivascular cuffing in vitro by MRM on three dimensional T1 proton stains. CONCLUSION: Inflammatory cell infiltration is a prerequisite for the development of lesions in EAE and MS. Thus, the ability to directly detect individual perivascular cuffs of inflammation may provide a useful means of monitoring the time course of inflammatory events, as conventional histopathological scoring of perivascular cuffs is utilized, but in the absence of sectioning and staining. PMID- 12112502 TI - Optimal post-contrast timing of breast MR image acquisition for architectural feature analysis. AB - PURPOSE: To define a post-contrast imaging time span during which diagnostic accuracy of breast magnetic resonance (MR) architectural feature analysis is maintained. MATERIALS AND METHODS: Seventy-five patients with mammographically visible or palpable findings underwent MR examination. Three sequential post contrast, fat-saturated, three-dimensional gradient-echo imaging runs were acquired spanning 0-90, 90-180, and 180-270 seconds after contrast injection. Five readers independently predicted the malignant potential of the MR abnormalities. RESULTS: Receiver-operator characteristics (ROC) curves were our primary measure of diagnostic accuracy. The accuracy of four readers was unchanged over the three post-contrast runs. One reader was slightly more accurate using the second and third runs than using the first. CONCLUSION: For most readers, a single post-contrast run performed at any point during the first four minutes and 30 seconds following injection should yield an equivalent diagnostic accuracy. If any time period is less optimal, it is that of our first run, performed between 0-90 seconds after contrast injection. PMID- 12112503 TI - Spoiled gradient-echo as an arterial spin tagging technique for quick evaluation of local perfusion. AB - PURPOSE: To introduce a simple gradient-echo arterial spin tagging (GREAST) technique available for most commercial magnetic resonance (MR) systems, for a quick evaluation of tissue perfusion. MATERIALS AND METHODS: The GREAST technique uses a combination of a short TR spoiled gradient-echo (SPGR) sequence with a selective presaturation radio frequency (RF) pulse that allows acquiring each tagged and control image within 10-20 seconds. The phantom and human studies were performed for evaluating the feasibility in measurement of local perfusion and the efficacy in alleviation of the asymmetric magnetization transfer (MT) and slice profile effects. RESULTS: Results show a good linear relationship between the signal attenuation caused by the presaturation pulse and flow rates in the phantom experiment and effective alleviation of the asymmetric MT and slice profile effects for various orientations of imaging slices. Human studies showed good perfusion results in brain imaging. Perfusion imaging on the liver and kidney were also conducted. The results could be significantly improved by effectively lessening motion-related artifacts. CONCLUSION: The GREAST technique is simple, easy to use, and applicable to examine local perfusion of the brain and other organs in the trunk. PMID- 12112504 TI - Axillary lymph node metastases: a statistical analysis of various parameters in MRI with USPIO. AB - PURPOSE: To assess the value of plain vs. iron oxide-enhanced MRI vs. the combined study (plain + postcontrast) based on qualitative and quantitative parameters of three pulse sequences. MATERIALS AND METHODS: Data from two sites were acquired using the same technique; therefore, this data could be pooled. T1W SE, T2W-FSE, and 3D-PSIF were used before and 24-36 hours after MRI with ultra small particles of iron oxide (USPIO) was performed. A total of 52 lymph nodes (LNs) in nine patients (25 benign, 27 malignant) were evaluated by two readers who were visually and quantitatively blinded to the histology. Combinations of the following diagnostic parameters were compared using logistic regression analysis: the short-axis diameter of the LN, the signal distribution of the LN on postcontrast agent MRI (homogeneous or heterogeneous), and qualitatively and quantitatively determined signal changes of the LN following administration of contrast agent in the three evaluated sequences. RESULTS: Using pre- and postcontrast data, the optimized accuracy based on the statistically most significant parameters (LN diameter > 6 mm, visual assessment of signal change on T2W-SE) was 87% (81% sensitivity, 92% specificity). Precontrast data alone yielded 75% accuracy (63% sensitivity, 86% specificity). Postcontrast data alone yielded 75% accuracy (56% sensitivity, 96% specificity). CONCLUSION: Based on our results, USPIO-MRI improved the diagnosis of metastatic axillary LNs compared with precontrast MRI alone. Both pre- and postcontrast studies are needed. T1W-SE and T2W-PSIF did not yield significant additional information. This study may help to further improve the technique of USPIO imaging. PMID- 12112505 TI - Assessment of patellofemoral relationships using kinematic MRI: comparison between qualitative and quantitative methods. AB - PURPOSE: To compare the level of agreement between quantitative and qualitative methods in determining patellofemoral relationships, since controversy exists regarding the use of quantitative vs. qualitative criteria to interpret images of the patellofemoral joint (PFJ) obtained using kinematic magnetic resonance (MR) imaging. MATERIALS AND METHODS: One hundred twenty mid-patellar axial plane images obtained using kinematic MR imaging from fifteen subjects were randomly selected for analysis. MR images represented various knee flexion angles ranging from 0 to 60 degrees. Quantitative analysis (bisect offset and patellar tilt angle) was performed by two examiners using a computer-assisted software program. Based on data from previously published literature, MR images were characterized as demonstrating normal, medial, or lateral patellar subluxation, and/or normal, medial, or lateral tilt. Using similar categories, two different examiners experienced in reading MR images of the PFJ then applied qualitative criteria to the same images. RESULTS: The average agreement between the quantitative and qualitative assessments of horizontal patellar displacement and patellar tilt ranged from poor to moderate (Kappa coefficient values of 0.27 and 0.45, respectively). Quantitative and qualitative techniques demonstrated acceptable intra- and inter-observer reliability. CONCLUSION: These findings indicate that the use of quantitative criteria does not compare well to qualitative criteria in the analysis of kinematic MR images of the PFJ. One explanation for this discrepancy relates to the fundamental difference between the techniques. That is, quantitative measurements are based on the use of osseous landmarks, while the qualitative assessments tend to rely on a description of patellofemoral relationships based on joint surfaces. PMID- 12112506 TI - MR imaging of the forefoot under weight-bearing conditions: position-related changes of the neurovascular bundles and the metatarsal heads in asymptomatic volunteers. AB - PURPOSE: To assess practicability of weight-bearing magnetic resonance (MR) imaging of the forefoot, and to demonstrate position-related changes of the neurovascular bundles and the metatarsal heads in asymptomatic volunteers. MATERIALS AND METHODS: With an open-configuration MR system, 32 feet of 32 asymptomatic individuals aged 20-60 years were studied in supine and weight bearing body positions. Transverse T1-weighted spin-echo MR images were performed. MR images were evaluated qualitatively with regard to image quality, visibility, and position-dependent changes of the neurovascular bundle. In addition, the position of the metatarsal heads was analyzed quantitatively. RESULTS: Weight-bearing MR imaging was feasible in all 32 feet. Quality of MR images obtained in the weight-bearing position was superior to that obtained in the supine position (P < 0.05). A change in the position of the neurovascular bundle from below a virtual line paralleling the plantar cortical line to above this line was present in 50 of 61 instances (82%) between the supine and the weight-bearing positions. When changing from the supine to the weight-bearing position, there was a significant decrease in the distance between the plantar skin and the metatarsal heads for the second (mean 4.5 mm), third (mean 4.4 mm), and fourth metatarsal heads (mean 3.7 mm) (P < 0.0001). However, the difference for the first (mean 0.5 mm) and fifth (mean 0.9 mm) metatarsal heads was not significant. CONCLUSION: Weight-bearing MR imaging of the forefoot is feasible using an open-configuration MR system and demonstrates position-related changes of the neurovascular bundles relative to the metatarsal heads, as well as position-related changes of the metatarsal heads themselves. PMID- 12112508 TI - Intrahepatic arterioportal fistula: demonstration by dynamic 3D magnetic resonance angiography in under four seconds. AB - We report a case of a 35-year-old patient with clinical symptoms of portal hypertension that underwent dynamic contrast-enhanced 3D magnetic resonance angiography (MRA) of the abdomen. On breath-hold dynamic contrast-enhanced MRA in less than 4 seconds, contrast passage from the arterial to the portal system was successfully demonstrated. Patient was managed according to MRA findings. PMID- 12112507 TI - Interactive body magnetic resonance fluoroscopy using modified single-shot half Fourier rapid acquisition with relaxation enhancement (RARE) with multiparameter control. AB - PURPOSE: To develop a technique for interactive fluoroscopic abdominal magnetic resonance imaging (MRI) based on a single-shot half-Fourier rapid acquisition and relaxation-enhanced sequence. MATERIALS AND METHODS: The sequence was modified by incorporating inner-volume excitation, driven-equilibrium signal enhancement, and reduced flip angle refocusing pulses. Contrast control was provided by integrating "on-the-fly" selection of phase encoding view order, fat suppression, and section thickness. The resulting sequence was evaluated with phantom experiments to quantify the signal-to-noise ratio (SNR) effects of the modifications and in healthy volunteers for imaging the bile ducts, stomach, and duodenum using water and gaseous contrast media. RESULTS: Observed SNR relating to driven-equilibrium and flip angle scaling matched theoretical predictions. Volunteer examinations demonstrated the ability of the modified sequence to provide interactive, artifact-free imaging of the abdomen, including switching between conventional proton density-weighted, T2-weighted imaging and "hydrographic" projection imaging. CONCLUSION: Refinement of this technique may provide an abdomino-pelvic imaging capability similar in concept to real-time ultrasound, but with the advantages of MRI. PMID- 12112509 TI - Magnetic resonance safety testing of a newly-developed fiber-optic cardiac pacing lead. AB - PURPOSE: To assess magnetic resonance (MR) safety for a newly developed, fiber optic cardiac pacing lead. MATERIALS AND METHODS: MR safety was assessed for the fiber-optic cardiac pacing lead by evaluating magnetic field interactions and heating. Translational attraction and torque were evaluated using a 1.5-Tesla MR system and previously described, standardized techniques. MR imaging-related heating was assessed using a 1.5-Tesla MR system and a transmit/receive, body radiofrequency (RF) coil with the fiber-optic lead positioned to simulate an in vivo condition in a saline-filled phantom. The phantom had dimensions similar to a human subject's torso and head. A fluoroptic thermometry system was used to record temperatures on and near the electrodes of the fiber-optic pacing lead at five-second intervals immediately before and during 20 minutes of MR imaging performed at a whole-body-averaged specific absorption rate (SAR) of 1.5 W/kg. Temperatures were also recorded from a reference site during this experiment. RESULTS: Magnetic field interactions for the fiber-optic lead were minimal (deflection angle, 23 degrees; torque, +2). The highest temperature change recorded for the fiber-optic cardiac pacing lead and reference site was +0.8 degrees C. CONCLUSION: The minor magnetic field interactions and relative lack of heating for the fiber-optic pacing lead indicate that it should be safe for patients with this device to undergo MR imaging procedures using MR systems operating at 1.5-T or less and at a whole-body-averaged SARs up to 1.5 W/kg. PMID- 12112511 TI - fMRI of the auditory cortex in patients with unilateral carotid artery steno occlusive disease. AB - PURPOSE: To examine whether an internal carotid artery (ICA) steno-occlusive disease leads to a reduced blood oxygenation level dependent (BOLD)-signal change due to a decreased vasodilatory reserve capacity. MATERIALS AND METHODS: Patients suffering from unilateral ICA stenosis or occlusion were examined using functional magnetic resonance imaging (fMRI) of the auditory cortex with a bilateral stimulation applying a pulsed 1000 Hz sine-tone. RESULTS: Compared to control subjects, who showed symmetric bilateral BOLD-responses within the auditory cortex, the ICA patients revealed either a normal bilateral cortical activation pattern or a reduced cortical activation on the steno-occluded side. This latter decrease of BOLD-signal change might indicate a depressed vasomotor reserve capacity. The observed coincidence between this asymmetric reduction in BOLD-signal and ischemic borderzone lesions on the affected side in this subgroup of patients strongly supports this assumption. CONCLUSION: This study shows that fMRI of the auditory cortex appears to have clinical merit in the workup of cerebrovascular conditions. PMID- 12112510 TI - Passive catheter tracking using MRI: comparison of conventional and magnetization prepared FLASH. AB - PURPOSE: To compare a magnetization-prepared gradient-echo (GRE) sequence with a conventional GRE sequence for visualizing contrast agent-filled catheters. MATERIALS AND METHODS: Passive visualization of endovascular catheters using MRI was compared between two imaging sequences: 1) inversion recovery (IR)-fast low angle shot (FLASH), and 2) conventional FLASH. Two-dimensional projection images of the catheters filled with 4% diluted contrast agent in a phantom and the aorta of swine were obtained with each sequence with a temporal resolution of two frames per second. We compared background suppression and catheter visibility using the catheter-to-background signal ratio and the ratings of two radiologists. RESULTS: In the phantom, IR-FLASH allowed for a 200% increase in catheter-to-background ratio (p < 0.01) and improved depiction of catheters over conventional FLASH. In swine, the IR-FLASH images showed a statistically significant improvement of 80% (p < 0.001) over conventional FLASH in all comparisons of the catheter-to-background signal ratio, and an improvement of 160% (p < 0.05) in comparison with the radiologists' observations. CONCLUSION: This study shows that IR-FLASH is a better technique for passive tracking of contrast agent-filled catheters than conventional FLASH. PMID- 12112512 TI - Effects of indomethacin on cerebral blood flow at rest and during hypercapnia: an arterial spin tagging study in humans. AB - PURPOSE: To investigate using an arterial spin tagging (AST) approach the effect of indomethacin on the cerebral blood flow (CBF) response to hypercapnia. MATERIALS AND METHODS: Subjects inhaled a gas mixture containing 6% CO(2) for two 5-minute periods, which were separated by a 10-minute interval, in which subjects inhaled room air. In six subjects, indomethacin (i.v., 0.2 mg/kg) was infused in the normocapnic interval between the two hypercapnic periods. RESULTS: Indomethacin reduced normocapnic gray matter CBF by 36 +/- 5% and reduced the CBF increase during hypercapnia from 43 +/- 9% to 16 +/- 5% in gray matter (P < 0.001) and from 48 +/- 11% to 35 +/- 9% in white matter (P < 0.025). CONCLUSION: The results demonstrate that an AST approach can measure the effects of indomethacin on global CBF increases during hypercapnia and suggest that an AST approach could be used to investigate pharmacological effects on focal CBF increases during functional activation. PMID- 12112513 TI - Diffusion-weighted MRI of cholesteatomas of the petrous bone. AB - PURPOSE: To investigate if primary cholesteatomas of the petrous bone show high signal in diffusion-weighted imaging (DWI). MATERIALS AND METHODS: In this blinded study, we compared 15 patients with clinically certain cases and later surgically proven cholesteatomas vs. 12 patients with clinically acute otitis of the middle ear and 20 volunteers without petrous bone disease. Two blinded readers without knowledge of the clinical data decided in consensus agreement whether there was a pathologic signal increase in the petrous bone in an anisotropic single-shot echo-planar imaging (EPI) DWI sequence, an artifact, or no signal increase. RESULTS: Thirteen of 15 patients with cholesteatomas showed bright signal in EPI DWI, whereas 10 of 12 patients with acute otitis media and all volunteers presented the usual low signal of petrous bone. CONCLUSION: EPI DWI is a fast diagnostic method that may be an additional valuable tool in the workup of suspected cholesteatomas. The ability of this technique to differentiate between cholesteatomas and granulomas or chronic otitis is not yet available. PMID- 12112514 TI - Fast phase contrast cardiac magnetic resonance imaging: improved assessment and analysis of left ventricular wall motion. AB - PURPOSE: To evaluate the use of CINE phase contrast magnetic resonance imaging (MRI) to assess and characterize left ventricular wall motion by two- or three directional velocity vector fields that reflect the temporal evolution of myocardial velocities over the whole cardiac cycle. MATERIAL AND METHODS: A fast imaging protocol is presented that permits the assessment of the pixel-wise full in-plane velocity information of the beating heart within a single breath-hold measurement. Temporal resolution of the acquired images is improved by the use of high-speed gradients and application of view sharing to black blood k-space segmented gradient echo imaging. A novel tool for data analysis is presented based on correlating locally different myocardial motion patterns to averaged left ventricular velocities reflecting nonpathological myocardial function. RESULTS: Measurement protocol and postprocessing options were evaluated in a study with 16 normal volunteers. Simulations showed that correlation analysis can be used to differentiate regions with altered velocity waveforms from global radial velocities. Results of patient examinations are presented on an exemplary basis and demonstrate that correlation analysis provides an effective method for identification and classification of myocardial dynamics. CONCLUSION: Within the framework of our volunteer and patient examinations, fast phase contrast cardiac MRI has proven to be a reliable method to assess and analyze myocardial performance on the basis of two-directional velocity vector fields. PMID- 12112516 TI - Respiratory motion in coronary magnetic resonance angiography: a comparison of different motion models. AB - PURPOSE: To assess respiratory motion models for coronary magnetic resonance angiography (CMRA). In this study various motion models that describe the respiration-induced 3D displacements and deformations of the main coronary arteries were compared. MATERIALS AND METHODS: Multiple high-resolution 3D coronary MR images were acquired in healthy volunteers using navigator-based respiratory gating, each depicting the coronary vessels at different respiratory motion states. In the images representing the different inspiratory states the displacements and deformations of the main coronary vessels with respect to the end-expiratory state were determined, by means of elastic registration. Several correction models (superior-inferior (SI) translation, 3D translation, and 3D affine transformation) were tested and compared with respect to their ability to map a selected inspiratory to the end-expiratory motion state. RESULTS: 3D translation was found to be superior over SI translation, which is commonly used for prospective motion correction in CMRA. The 3D affine transformation was found to be the best correction model considered in this study. Furthermore, a large intersubject variability of the model parameters was observed. CONCLUSION: The results of this study indicate that a patient-adapted 3D correction model (3D translation or better 3D affine) will considerably improve prospective motion correction in CMRA. PMID- 12112515 TI - Reproducibility of left ventricular mass measurement using a half-Fourier black blood single-shot fast spin-echo sequence within a single breath hold: comparison with a conventional multiple breath-hold segmented gradient echo technique in patients. AB - PURPOSE: To compare the reproducibility of left ventricular (LV) mass measurements using a black-blood half-Fourier single-shot fast spin-echo (SSFSE) and a segmented gradient echo magnetic resonance (MR) pulse sequence. MATERIAL AND METHODS: Breath-hold SSFSE and segmented gradient echo cardiac MR examinations were performed twice in 32 patients and manual detection of the LV endocardium and epicardium was applied by two blinded reviewers. The SSFSE pulse sequence allowed whole-heart coverage in a single breath hold, while multiple breath holds were required using the segmented gradient echo sequence. Spatial presaturation slabs were used with the SSFSE pulse sequence to reduce the field of view (FOV) and thereby achieve higher spatial resolution. RESULTS: Intraclass correlation coefficients were higher with the SSFSE pulse sequence than with the segmented gradient echo pulse sequence: intraobserver reproducibility reached 0.999 vs. 0.991; interobserver reproducibility: 0.997 vs. 0.981; and interstudy reproducibility: 0.998 vs. 0.936. These higher levels of reproducibility were confirmed on Bland and Altman plots. CONCLUSION: LV mass measurements can be assessed more reproducibly with the single breath-hold SSFSE technique than with the standard multiple breath-hold segmented gradient echo method. PMID- 12112517 TI - Nonuniform flow dynamics in the aorta of normal children: a simplified approach to measurement using magnetic resonance velocity mapping. AB - PURPOSE: To determine regional flow dynamics in the normal aorta (Ao) in children. Understanding flow dynamics in children is important in cardiovascular energetics, in designing improved aortic reconstructions by crafting the surgery to mimic normal aortic flow, and in Doppler flow calculations. The objective of this study was to determine regional flow dynamics in the normal Ao in children. MATERIALS AND METHODS: We performed magnetic resonance velocity mapping on 13 subjects (ages 7.2 +/- 6.2 years) with normal Aos to determine flow dynamics in four equal quadrants in the ascending (AAo) and descending aorta (DAo) aligned along the long axis of the Ao. Statistical significance was set at P <.05. RESULTS: In the AAo, the left posterior quadrant displayed significantly less blood flow (16% +/- 5%) than the other quadrants (26-29%) over the cardiac cycle. In the DAo, both anterior quadrants carried significantly less blood flow (20% and 21%) than the posterior quadrants (27% and 32%). At maximum flow (15% +/- 5% into the cardiac cycle for the AAo; 27% +/- 15% for the DAo), there was significantly more flow in the right posterior quadrant (28% +/- 2%) than other quadrants (22-23%) in the AAo. In the DAo, both posterior quadrants had significantly higher flow rates (27% and 30%) than the anterior quadrants (21% and 22%). Maximum velocity in both the AAo and the DAo occurred in the left posterior quadrant in 10/13 at 16-24% into the cardiac cycle. At end-systole, a short flow reversal was noted in the posterior quadrants in the AAo in 11/13; in the DAo, this occurred in the anterior quadrants in 10/13. CONCLUSION: Flow dynamics in the normal Ao in children are not symmetrical; the flow distributions are asymmetric in both the AAo and the DAo throughout systole, including flow reversal related to the dicrotic notch. These results may help improve Ao surgery. PMID- 12112518 TI - Short echo time MR spectroscopic imaging of the lung parenchyma. AB - PURPOSE: To perform short echo time MR spectroscopic imaging of the lung parenchyma on normal volunteers. MATERIALS AND METHODS: A short echo time projection-reconstruction spectroscopic imaging sequence was implemented on a commercial 1.5T whole body MRI scanner. Images and spectra of the lung parenchyma were obtained from five normal volunteers. Breath-held spectroscopic imaging was also performed. RESULTS: Spectroscopic imaging of short-T2* species allows visualization of different anatomic structures based upon their frequency shifts. A characteristic peak from the parenchyma was seen at three ppm from water frequency. CONCLUSION: Short echo time MR spectroscopic imaging of the lung parenchyma was demonstrated in normal volunteers. This method may improve proton imaging of the lungs and add specificity to the diagnosis of pulmonary disease. PMID- 12112519 TI - Prediction of postoperative pulmonary function using perfusion magnetic resonance imaging of the lung. AB - PURPOSE: To assess semiquantitatively the regional distribution of lung perfusion using magnetic resonance (MR) perfusion imaging. MATERIALS AND METHODS: Subjects were 20 consecutive patients with bronchogenic carcinoma, who underwent MR imaging (MRI) and radionuclide (RN) perfusion scans for preoperative evaluation. Three-dimensional (3D) images of whole lungs were obtained before and 7 seconds after bolus injection of contrast material (5 ml of Gd-DTPA). Subtraction images were constructed from these dynamic images. Lung areas enhanced with the contrast material were measured and multiplied by changes in signal intensity, summed for the whole lung, and the right-to-left lung ratios were calculated. The predicted postoperative forced expiratory volume in 1 second (FEV1) was estimated using MR and RN perfusion ratios. RESULTS: The correlation between perfusion ratios derived from the MR and RN studies was excellent (r = 0.92). Sixteen of 20 patients underwent surgery, and 12 patients had postoperative pulmonary function tests. The predicted FEV1 derived from the MR perfusion ratio correlated well with the postoperative FEV1 in the 12 patients (r = 0.68). CONCLUSION: Perfusion MRI is suitable for semiquantitative evaluation of regional pulmonary perfusion. PMID- 12112520 TI - In vivo diffusion-weighted MRI of the breast: potential for lesion characterization. AB - PURPOSE: To investigate the potential of apparent diffusion coefficients (ADCs) in characterizing breast lesions in vivo. MATERIALS AND METHODS: Two diffusion weighted (DW) sequences were implemented on a 1.5 Tesla scanner, with low b-value orthogonal and high b-value tetrahedral sensitized sequences. The orthogonal sequence was evaluated on 16 normal volunteers and 23 patients with known lesion types (six benign and 17 malignant). The tetrahedral sequence was evaluated on a smaller number of subjects: two normal, two malignant, and two benign. RESULTS: The mean value of the ADC of the malignant tumors was reduced compared to that of the benign lesions and normal tissue. This finding was related to the increased cellularity of the malignant lesions. The ADC values were elevated for all tissue types with the low b-value sequence as compared to the high b-value sequence, indicating contributions from perfusion effects at the low b-values. CONCLUSION: The study clearly shows that DW-MRI can help characterize breast lesions in vivo. PMID- 12112521 TI - MR pattern of hyperacute cerebral hemorrhage. AB - Magnetic resonance (MR) pattern of cerebral hemorrhage relates mainly to the relaxation and susceptibility effects of iron-containing hemoglobin degradation products, as well as to their intra- or extracellular location. The purpose of this article is to report two acute stroke patients who underwent thrombolytic therapy and developed hyperacute cerebral hemorrhage during their admission cerebral MR survey. They constitute the earliest MR appearance of hyperacute intracerebral bleeding reported in the literature, featuring increased diffusion properties and persistent susceptibility effect on perfusion-weighted imaging (PWI)-series. PMID- 12112522 TI - Real-time MRI of joint movement with trueFISP. AB - PURPOSE: To develop a technique for dynamic magnetic resonance imaging (MRI) of joint motion based on a combination of real-time TrueFISP (fast imaging with steady state precession) imaging with surface radiofrequency (RF) coils. MATERIALS AND METHODS: The metacarpal, elbow, tarsal, and knee joint of five volunteers and the knees of four patients were examined with a real-time TrueFISP sequence during movement of the joints. RESULTS: All examined joints could be assessed under dynamic conditions with high image contrast and high temporal resolution. CONCLUSION: Dynamic MRI of joints with TrueFISP is feasible and can provide information supplemental to static joint examinations. PMID- 12112523 TI - Loss of heterozygosity analysis: practically and conceptually flawed? AB - The Knudson "two-hit" hypothesis has provided the rationale for studies that aim to identify tumor-suppressor genes by mapping regions of allelic loss (loss of heterozygosity, LOH). Although LOH has been found in practically all types of tumors, very few such projects have been successful in identifying their tumor suppressor targets. The prime explanation for this failure is probably that researchers have, in general, been too credulous about the two-hit hypothesis, and too willing to ignore factors such as intratumor heterogeneity, contamination by normal cells, karyotypic complexity, homozygous deletions, gene dosage changes, and polymerase chain reaction artifacts. We suggest ways of minimizing these problems. Unfortunately, there is no guarantee that existing or newer methods, such as genomic microarrays and in situ single-nucleotide polymorphism analysis, will solve the difficulties of LOH analysis. The future prospects for LOH studies are, as ever, uncertain. PMID- 12112524 TI - Identification of novel fusion partners of ALK, the anaplastic lymphoma kinase, in anaplastic large-cell lymphoma and inflammatory myofibroblastic tumor. AB - ALK-positive anaplastic large-cell lymphoma (ALCL) has been recognized as a distinct type of lymphoma in the heterogeneous group of T/Null-ALCL. While most of the ALK-positive ALCL (ALKomas) are characterized by the presence of the NPM ALK fusion protein, the product of the t(2;5)(p23;q35), 10-20% of ALKomas contain variant ALK fusions, including ATIC-ALK, TFG-ALK, CLTC-ALK (previously designated CLTCL-ALK), TMP3-ALK, and MSN-ALK. TMP3-ALK and TMP4-ALK fusions also have been detected in inflammatory myofibroblastic tumors (IMTs), making clear that aberrations of the ALK gene are not associated exclusively with the pathogenesis of ALK-positive ALCL. Here we report results of molecular studies on two lymphoma cases and one IMT case with variant rearrangements of ALK. Our study led to the detection of the CLTC-ALK fusion in an ALCL case and to the identification of two novel fusion partners of ALK: ALO17 (KIAA1618), a gene with unknown function, which was fused to ALK in an ALCL case with a t(2;17)(p23;q25), and CARS, encoding the cysteinyl-tRNA synthetase, which was fused to ALK in an IMT case with a t(2;11;2)(p23;p15;q31). These results confirm the recurrent involvement of ALK in IMT and further demonstrate the diversity of ALK fusion partners, with the ability to homodimerize as a common characteristic. PMID- 12112525 TI - Clinical significance of alterations of chromosome 8 detected by fluorescence in situ hybridization analysis in pathologic organ-confined prostate cancer. AB - Loss of 8p22 and gain of 8q24 are known to be common chromosomal alterations in prostate cancer. We have previously demonstrated that concurrent 8q24 overrepresentation and 8p22 loss were associated with a poor prognosis in patients with high-grade, locally advanced prostate cancer. We evaluated the alteration of 8p22 and 8q24 in a large cohort of pathologic organ-confined prostate cancer using fluorescence in situ hybridization (FISH) analysis. All 195 patients with Gleason scores > 5, pathologic stage T(2)N(0)M(0) (pT(2)N(0)M(0)) prostate cancer, who underwent a radical prostatectomy at the Mayo Clinic between 1987 and 1991, and for whom blocks were available, were selected for this study. The median follow-up period was 9.5 years, and endpoints of this study were biochemical and clinical disease progression. The latter includes local as well as systemic disease progression. FISH analysis using paraffin-embedded tissues was performed for 8p22 (LPL), centromere 8 (8cen), and 8q24 (MYC) and was successful for 156 tumors (80.0%). Of these tumors, 104 (66.6%) had one or more numeric alterations of the 3 loci evaluated. An increased copy number of 8q24 was observed in 66 (42.3%) tumors, of which 20 (12.8%) had an additional increase (AI) of 8q24, and 46 (29.5%) had a gain of 8q24 with an equivalent gain of 8cen. Losses and gains of 8p22 were detected in 81 (51.9%) and 20 (12.8%) tumors, respectively. An AI of 8q24 was significantly associated with the tumor Gleason score (P = 0.042). Univariate analysis indicated that loss of 8p22 was a significant predictor of biochemical and clinical disease progression (P = 0.025 and P = 0.011, respectively). Furthermore, the group with loss of 8p22 concurrent with an AI of 8q24 (Loss 8p22-any 8cen-AI 8q24) had an increased rate of biochemical disease progression (P = 0.052). Multivariate analysis demonstrated that neither individual nor the Loss-any-AI combination of alterations was a significant independent predictor of disease progression when adjusting for Gleason score, preoperative PSA levels, and DNA ploidy. These data suggest that loss of 8p22 is associated with a poor prognosis, specifically when it is accompanied by AI of 8q24 in pT(2)N(0)M(0) prostate cancer. PMID- 12112526 TI - Preferential expression of a mutant allele of the amplified MDR1 (ABCB1) gene in drug-resistant variants of a human sarcoma. AB - Activation of the MDR1 (ABCB1) gene is a common event conferring multidrug resistance (MDR) in human cancers. We investigated MDR1 activation in MDR variants of a human sarcoma line, some of which express a mutant MDR1, which facilitated the study of allelic gene expression. Structural alterations of MDR1, gene copy numbers, and allelic expression were analyzed by cytogenetic karyotyping, oligonucleotide hybridization, Southern blotting, polymerase chain reaction, and DNA heteroduplex assays. Both chromosome 7 alterations and several cytogenetic changes involving the 7q21 locus are associated with the development of MDR in these sarcoma cells. Multistep-selected cells and their revertants contain three- to six-fold MDR1 gene amplification compared with that of the drug sensitive parental cell line MES-SA and single-step doxorubicin-selected mutants. MDR1 gene amplification precedes the emergence of a mutant allele in cells that were coselected with doxorubicin and a cyclosporin inhibitor of P-glycoprotein (P gp). Allele-specific oligonucleotide hybridization showed that the endogenous mutant allele was present as a single copy, with multiple copies of the normal allele. Reselection of revertant cells with doxorubicin in either the presence or the absence of the P-gp inhibitor resulted in exclusive reexpression of the mutant MDR1 allele, regardless of the presence of multiple wild-type MDR1 alleles. These data provide new insights into how multiple alleles are regulated in the amplicon of drug-resistant cancer cells and indicate that increased expression of an amplified gene can result from selective transcription of a single mutant allele of the gene. PMID- 12112527 TI - Analysis of 11q21-24 loss of heterozygosity candidate target genes in breast cancer: indications of TSLC1 promoter hypermethylation. AB - Loss of heterozygosity (LOH) at the distal half of chromosome arm 11q is frequent in a variety of human tumors, including breast cancer, and is often associated with poor prognosis. In an ongoing attempt to locate and characterize the main target genes within this chromosome region, we first looked for aberrations in known genes either suggested to be involved in tumorigenesis or shown to suppress tumor formation. We examined 31 primary breast tumors showing LOH in 11q21-24 for mutations in the MRE11A, CHK1, PPP2R1B, and TSLC1 genes. The absence of intragenic alterations related to cancer led us next to evaluate possible gene silencing resulting from promoter region CpG hypermethylation, using the bisulfite sequencing technique. In addition to the four genes mentioned above, we also analyzed the ATM gene, which had been investigated for certain germline mutations in an earlier study. Only the TSLC1 promoter region exhibited aberrant methylation patterns, and altogether 33% (10/30) of the successfully analyzed tumors showed evidence of elevated levels of TSLC1 CpG methylation. Ten percent (3/30) of the tumors showed significantly increased methylation. Thus, as has been shown in lung and some other forms of cancer, hypermethylation of the TSLC1 promoter region is also frequently a second hit along with LOH in breast cancer. PMID- 12112528 TI - Functional evidence from microcell-mediated chromosome transfer of myeloid leukemia suppressor genes on human chromosomes 7 and 11. AB - The long arm of human chromosome 7 between 7q22 and 7q36 has been identified as a region harboring one or more tumor-suppressor genes (TSGs) inactivated in acute myeloid leukemia (AML). Additional TSGs mapping to other chromosomes may well be involved in the etiology of this disease. For example, experiments using a mouse model system have indicated the possible presence of an AML TSG at 11p11-12. Microcell-mediated chromosome transfer (MMCT) has been used to introduce human chromosomes 7 and 11 into a murine myeloid leukemia cell line. A proportion of MMCT hybrid clones containing either whole chromosome 7 or fragments of chromosome 11 showed a significant delay in leukemogenic onset when injected into syngeneic mice. Screening of hybrid clones did not associate any human microsatellite markers with decreased leukemogenic potential in vivo. However, preliminary evidence was obtained of allelic loss at chromosomal regions homologous with human 7q22 in murine F1 hybrid AMLs. Our data provide functional evidence of AML-associated TSGs localized to human chromosomes 7 and 11 in support of previously published studies on cytogenetic and allelic losses associated with AML development. PMID- 12112529 TI - No evidence for hypermethylation of the hSNF5/INI1 promoter in pediatric rhabdoid tumors. AB - The hSNF5/INI1 gene on chromosome 22 has been implicated as a tumor suppressor gene in pediatric rhabdoid tumor, an aggressive malignancy that generally occurs in the first two years of life. The most common sites for tumor development are the brain and kidney. We and other investigators have identified deletions and mutations of the INI1 gene in the majority of rhabdoid tumors of the central nervous system, kidney, and extrarenal tissues. At least 20% of cases do not have genomic alterations of INI1, although expression at the RNA or protein level may be decreased. The aim of this study was to determine whether hypermethylation or mutation of the 5' promoter region of INI1, or hypermethylation of CpG dinucleotides in a GC-rich repeat region within the first intron, could account for the decreased expression of INI1 observed in these tumors. We employed bisulfite modification, polymerase chain reaction, and sequence analysis to determine the methylation status of the cytosine nucleotides in the predicted promoter region of the INI1 gene, and two GC repeat regions in intron 1. DNA from 24 tumors with or without coding-sequence mutations was analyzed. None of the tumors demonstrated methylation of the promoter or intron 1 regions. This mechanism is unlikely to account for the inactivation of INI1 in rhabdoid tumors without coding-sequence mutations. One tumor demonstrated a potential mutation in the promoter region, but further studies are required for determining its functional significance. PMID- 12112530 TI - Transcriptional profiling reveals that several common fragile-site genes are downregulated in ovarian cancer. AB - Previous transcriptional profiling analysis of 14 primary ovarian tumors identified approximately 12,000 genes as decreased in expression by at least twofold in one or more of the tumors sampled. Among those genes were several known to be mapped to common fragile sites (CFSs), some of which had previously been shown to exhibit a loss of expression in ovarian carcinoma. Therefore, we selected a subset of genes to determine whether they localized within CFSs. Of the 262 genes that were downregulated at least twofold in 13 of 14 tumors, 10 genes were selected based on the following criteria: localization to a CFS band; documented aberrations in at least one malignancy; and feasibility of scoring breakage at the specific CFS. Fluorescence in situ hybridization analysis was performed using bacterial artificial chromosome clones encompassing portions of the genes to determine the position of the genes relative to their corresponding CFSs. Nine genes were determined to localize within seven previously uncloned CFSs. Semiquantitative reverse-transcription/polymerase chain reaction analysis of the cell lines and primary ovarian tumors validated the downregulation of seven of the 10 genes. We identified portions of seven uncloned CFSs and provide data to suggest that several of the genes mapping within CFSs may be inactivated in ovarian cancer. PMID- 12112531 TI - Comprehensive analysis of genomic alterations in gliosarcoma and its two tissue components. AB - Gliosarcoma is a variant of glioblastoma multiforme characterized by two components displaying gliomatous or sarcomatous differentiation. We investigated 38 gliosarcomas for aberrations of tumor-suppressor genes and proto-oncogenes that are commonly altered in glioblastomas. Amplification of CDK4, MDM2, EGFR, and PDGFRA were found in 11% (4/35), 8% (3/38), 8% (3/38), and 3% (1/35) of the tumors, respectively. Nine of 38 gliosarcomas (24%) carried TP53 mutations. PTEN mutations were identified in 45% (9/20) of the investigated tumors. Twenty gliosarcomas were analyzed by comparative genomic hybridization (CGH). Chromosomal imbalances commonly detected were gains on chromosomes 7 (15/20; 75%), X (4/20; 20%), 9q, and 20q (3/20, 15% each); and losses on chromosomes 10 and 9p (7/20, 35% each), and 13q (3/20, 15%). Five different high-level amplifications were mapped to 4q12-q21 (1 case), 6p21 (1 case), 7p12 (2 cases), proximal 12q (4 cases), and 14q32 (1 case) by CGH. Southern blot and/or differential PCR analyses identified amplification of PDGFRA (4q12), CCND3 (6p21), EGFR (7p12), CDK4 (12q14) and/or MDM2 (12q14.3-q15), and AKT1 (14q32.3) in the respective tumors. Separate analysis of the gliomatous and sarcomatous components of eight gliosarcomas by CGH after microdissection and universal DNA amplification revealed that both components shared 57% of the chromosomal imbalances detected. Taken together, our data indicate that the genomic changes in gliosarcomas closely resemble those found in glioblastomas. However, the number of chromosomes involved in imbalances in gliosarcomas was significantly lower than that in glioblastomas, indicating a higher genomic stability in gliosarcomas. In addition, we provide further support for the hypothesis that the gliomatous and sarcomatous components are derived from a single precursor cell clone, which progressed into subclones with distinct morphological features during tumor evolution. According to our data, gain/amplification of genes on proximal 12q may facilitate the development of a sarcomatous phenotype. PMID- 12112532 TI - Breakpoint position on 17q identifies the most aggressive neuroblastoma tumors. AB - Gain of chromosome arm 17q is a powerful prognostic factor in neuroblastoma, and the distribution of 17q breakpoints suggests that the dosage of one or more genes in 17q22-23 to 17qter is critical for tumor progression. To identify the smallest region of 17q gain, we used eight probes to map translocation breakpoints in 48 primary neuroblastoma tumors. We identified at least five different breakpoints, all localized within the proximal part of 17q (from D17Z1 to MPO). The shortest region of gain identified by these probes extends from MPO (17q23.1) to 17qter. Surprisingly, we found that breakpoints localized proximal to ERBB2 (17q12) were associated with significantly better patient survival than breakpoints localized distal to ERBB2. Breakpoints localized distal to ERBB2 identified patients with a particularly poor prognosis, higher mitotic karyorrhectic index, and stage 4 disease. This implies that breakpoint position on 17q is a discriminative factor within this prognostically poor group of patients. This result also suggests that the biological effect of 17q gain during neuroblastoma progression has a complex basis. We propose that this involves dosage alterations of genes localized on both sides of the 17q breakpoints, with a gene or genes mapping between 17cen and 17q12 acting to suppress progression, and a gene or genes mapping between 17q23.1 and 17qter acting to promote tumor progression. PMID- 12112533 TI - The chromosome translocation t(7;11)(p15;p15) in acute myeloid leukemia results in fusion of the NUP98 gene with a HOXA cluster gene, HOXA13, but not HOXA9. AB - The nucleoporin gene NUP98 has been reported to be fused to 9 partner genes in hematologic malignancies with 11p15 translocations. The NUP98-HOXA9 fusion gene has been identified in acute myeloid leukemia (AML) and chronic myelogenous leukemia with t(7;11)(p15;p15). We report here a novel NUP98 partner gene, HOXA13, in a patient with de novo AML having t(7;11)(p15;p15). The HOXA13 gene is part of the HOXA cluster genes and contains 2 exons, encoding a protein of 338 amino acids with a homeodomain. The NUP98-HOXA13 fusion protein consists of the N terminal phenylalanine-glycine repeat motif of NUP98 and the C-terminal homeodomain of HOXA13, similar to the NUP98-HOXA9 fusion protein. Reverse transcriptase-polymerase chain reaction (RT-PCR) analysis in various leukemic cell lines showed that the HOXA13 gene was expressed significantly more frequently in acute monocytic leukemic cell lines than in other leukemic cell lines (P = 0.039). HOXA13 and three HOXA cluster genes (A9, A10, A11) located at the 5' end of the HOXA9 gene were frequently expressed in myeloid leukemic cell lines. Our results revealed that t(7;11)(p15;p15) was not a single chromosomal abnormality at the molecular level. The protein encoded by the NUP98-HOXA13 fusion gene is similar to that encoded by NUP98-HOXA9, and the expression pattern of the HOXA13 gene in leukemic cell lines is similar to that of the HOXA9 gene, suggesting that the NUP98-HOXA13 fusion protein may play a role in leukemogenesis through a mechanism similar to that of the NUP98-HOXA9 fusion protein. PMID- 12112534 TI - Percutaneous tracheostomy with the guide wire dilating forceps technique: presentation of 171 consecutive patients. AB - BACKGROUND: Evaluation of percutaneous tracheostomy (PT) with the guide wire dilating forceps (GWDF) technique. METHODS: Prospective study of perioperative complications, retrospective analysis of early and late complications in an ICU in a teaching university hospital. RESULTS: The success rate of the procedure was 96.5%. The average procedure time in 171 consecutive patients was 5.0 min. Perioperative complications requiring surgical or medical intervention occurred in 6.4% of 171 patients. This included conversion to surgical tracheostomy, which was necessary in six patients (3.5%). Major complications while being cannulated occurred in 2.4% of 164 patients but seemed mostly unrelated with the GWDF technique itself. Late complications (after decannulation) were mostly minor and occurred in 22.6% of 106 patients. Only one patient (0.9%) had a symptomatic tracheal stenosis developed. CONCLUSION: Percutaneous tracheostomy with the guide wire dilating forceps technique is easy to perform at the bedside with few late complications. However, in our study, perioperative and immediate postoperative bleeding complications (minor and major) occur quite often. PMID- 12112535 TI - Immunohistochemical study of epidermal growth factor receptor in adenoid cystic carcinoma of salivary gland origin. AB - BACKGROUND: Epidermal growth factor (EGF) and its receptor (EGFR) are involved in the development of salivary gland tumors. Recently, treatment modalities for EGFR inhibition have shown an enhanced clinical response in carcinomas of different locations. Adenoid cystic carcinoma (ACC) of salivary gland origin is a malignant tumor with a poor long-term outcome. If salivary gland ACC does exhibit EGFR, then immunotherapy could have a major impact on improving its prognosis. METHODS: The study consisted of 34 samples of formalin-fixed, paraffin-embedded specimens of salivary gland ACC. Specimens were stained with a mouse antihuman monoclonal antibody for immunohistochemical detection of EGFR. Overlying oral mucosa and adjacent normal salivary ducts served as internal controls. Both membrane and cytoplasmic staining were evaluated. Staining score was calculated by multiplying the percentage of positively stained tumor cells by the intensity of the staining. The highest score for a given tumor was equal to 2. RESULTS: In the final analysis, 27 of the 34 specimens were included; 7 were excluded, because the internal control did not reveal any staining. Of these 27 specimens, 23 (85%) stained positively for EGFR with a staining score of 0.05 to 1.8. Three palatal tumors attained the highest scores (one tumor, 1.2, and the remaining two, 1.8). CONCLUSIONS: Most salivary gland ACC stained positively for EGFR, and in some the staining was quite intense. On the basis of the already proven antitumoral effect of agents acting as EGFR inhibitors, it is suggested that patients with ACC might benefit from these agents, especially when surgery has failed or in those with recurrent or metastatic disease. PMID- 12112536 TI - "FAR" chemoradiotherapy improves laryngeal preservation rates in patients with T2N0 glottic carcinoma. AB - BACKGROUND: The appropriate treatment approach for patients with T2N0 laryngeal cancer remains highly controversial. Because radiotherapy alone is associated with a high risk of local recurrence, we have developed a triple combination treatment approach consisting of 5-fluorouracil (250 mg/day, i.v.), vitamin A (50,000 unit/day, i.m.) and external radiation (2.0 Gy/day), which we have termed "FAR therapy." METHODS: Patients with T2N0 glottic carcinoma were initially treated with 15 days of FAR therapy, which included a cumulative radiation dose of 30Gy (i.e., "30 Gy of FAR therapy"). Those patients who demonstrated a complete response either clinically or pathologically continued to receive further FAR therapy, with up to 60-70 Gy. All other patients received laryngectomy without any additional treatment. RESULTS: Ninety-five patients were treated according to this program, and most of the patients (98%) were able to complete this treatment course. Eighty-eight patients (93%) were treated with FAR therapy alone. The local control and ultimate local control rates were 91% (85 of 93), and 99% (92 of 93), respectively. The cumulative 5-year voice preservation and complete laryngeal preservation rates were 91% and 87%, respectively. The cumulative 5-year disease-specific survival rate was 97%. CONCLUSIONS: Because a high rate of laryngeal preservation was achieved without compromising disease specific survival, our treatment approach based on FAR therapy may be promising for the treatment of patients with T2N0 glottic carcinoma. PMID- 12112537 TI - Computer assisted analysis of the microvasculature in metastasized and nonmetastasized squamous cell carcinomas of the tongue. AB - BACKGROUND: Quantification of microvessels in solid malignancies is regarded as a potential test to predict their clinicobiologic behavior. However, discordant results have been reported for head and neck cancer that may be explained by varying methods. METHODS: In this retrospective study, we therefore quantified the microvasculature in 20 nonmetastasized and 20 metastasized squamous cell carcinomas of the tongue, using recently developed methods. For immunohistochemical visualization of the vessels, we used anti-CD34 with a signal amplification step based on the catalyzed deposition of biotinylated tyramine. This protocol results in enhanced staining quality compared with standard protocols. For each tumor, a representative tissue section was systematically sampled with 40 to 60 standardized test fields. True color image analysis system was used to measure microvessel density (MVD) and to obtain additional information with regard to size categories of vessels. RESULTS: Remarkably, in the group of nonmetastasized tumors, the MVD was greater than in the metastasized tumors (p =.007). However, the microvessels with a diameter in the range of 10 to 15 microm predominated in the group of metastasized tongue carcinomas (p =.03). A logistic regression model based on the percentage of vessels smaller than 5 microm, classified 85% of patients with a metastasized tumor correctly and 75% of patients with a nonmetastasized tumor, independently of the clinical stage of the tumor. CONCLUSIONS: These results suggest that only vessels with a diameter larger than 10 microm, consistent with functional vessels, play a role in the process of metastasis. Further research more specifically into structural and functional characterization of blood and lymphatic vessels might help provide more insight into the relationship between microvasculature and the pathogenesis of metastasis in tongue carcinomas. PMID- 12112538 TI - Thyroid calcification and its association with thyroid carcinoma. AB - AIM: Calcification within the thyroid gland may occur in both benign and malignant thyroid disease, and its detection on ultrasonography is frequently dismissed by many clinicians as an incidental finding of little significance. As a tertiary referral center, most of our thyroid patients will have had thyroid ultrasonography before being referred to us, and in our experience, the incidence of malignancy in a thyroid nodule containing calcification seems to be higher than that in the average thyroid nodule. To assess this risk, we conducted this retrospective review. MATERIALS AND METHODS: Our analysis included 462 consecutive patients who underwent thyroid surgery at our institution between 1995 and 1999. We reviewed all the patients' charts for data regarding clinical findings, preoperative diagnostic investigations, and histopathologic diagnosis. Of the 462 patients, 361 (78.1%) had thyroid ultrasonography before surgery, and 49 (13.6%) of these ultrasounds showed intrathyroidal calcification. RESULTS: Of the 49 patients whose ultrasounds showed intrathyroidal calcification, 29 (59.2%) were found on histopathologic examination to have thyroid carcinoma. Twelve of the remaining 20 patients had multinodular goiters. Of the 29 patients with malignancy, seven (24.1%) had preoperative fine-needle aspirates that were reported as benign. After excluding patients who were initially seen with multinodular disease, in the subset of 37 patients who presented with a solitary thyroid lesion with calcification, 28 (75.7%) were found to have carcinoma. CONCLUSIONS: When calcification is noted within a solitary thyroid nodule, the risk of malignancy is very high. Surgery should be recommended regardless of the result of fine-needle aspiration cytologic findings. PMID- 12112539 TI - Histopathologic parameters in the evaluation of T1 squamous cell carcinomas of the oral cavity. AB - BACKGROUND: Prognostic indicators that could assist in a more precise selection of patients with small oral carcinomas for differentiated therapy would be valuable. A significant fraction of patients with stage I disease have a relatively poor prognosis despite the small size of the tumor, but in general stage I tumors of the oral cavity have a favorable prognosis. METHODS: Seventy eight patients with stage I (T1N0M0) oral squamous cell carcinoma from two different ENT departments were included in the study. The pretreatment biopsy specimens were graded according to the modified classification of Jakobsson et al. Eight individual parameters were recorded, four parameters describing the tumor cell population and four parameters describing the tumor/host interaction. RESULTS: The only significant prognostic parameter for disease-specific survival was "mode of invasion." The histologic mean score was not significantly correlated to disease-specific or crude survival. CONCLUSIONS: Mode of invasion is the most important histologic parameter when evaluating the prognosis. Histologic evaluation of small squamous cell carcinomas of the oral cavity may assist the design of a differentiated treatment strategy (eg, monotherapy vs combined treatment). PMID- 12112540 TI - A multicenter phase II study of tgDCC-E1A for the intratumoral treatment of patients with recurrent head and neck squamous cell carcinoma. AB - BACKGROUND: The anti-cancer gene, E1A, can be complexed to a lipid carrier, DC Cholesterol:DOPE, to form tgDCC-E1A, which can be injected directly into tumors. METHODS: Twenty-four patients with recurrent, unresectable, head and neck cancer were treated with intratumoral injections of tgDCC-E1A over 8 weeks. Tumor response was assessed using CT scans. Time to progression and overall survival were calculated. RESULTS: Intratumoral tgDCC-E1A was well tolerated in all patients. No significant toxicities related to tgDCC-E1A were reported. One patient (4.2%) had a complete response, two patients (8.3%) had minor response, and seven patients (29.2%) had stable disease by two-dimensional cross-products on blinded CT scans. The median time to progression was 8.6 weeks (range, 3.3 43.7 weeks), and median survival was 4.6 months (range, 1.3-15.6 months). CONCLUSIONS: Intratumoral injections of tgDCC-E1A were safe and well tolerated. Modest tumor response was observed. Further development of tgDCC-E1A is warranted in combination with other treatment modalities. PMID- 12112541 TI - Differentiated thyroid carcinoma: comparison between papillary and follicular carcinoma in a single institute. AB - PURPOSE: To compare and contrast the clinical presentation and treatment outcome of patients with papillary and follicular thyroid carcinoma and to study the pattern of practice of treatment of differentiated thyroid carcinoma in Hong Kong. METHOD: The clinical presentation and treatment outcomes were reviewed for 1057 patients with differentiated thyroid cancers who were treated at the Queen Elizabeth Hospital, Hong Kong, from 1960 to 1997. Eight hundred forty-two patients had papillary thyroid carcinomas (PTC), and 215 had follicular thyroid carcinomas (FTC). The mean follow-up was 9.2 years. RESULTS: The differences in the clinical factors of PTC to FTC were as follows: PTC had a higher incidence (3.9:1); these patients were younger at presentation (median age, 44 vs 49), showed a higher female-male ratio (4.5 vs 2.9) and smaller primary tumor size (median 2 cm vs 3.5 cm), and a higher incidence of multifocal disease (28.3% vs 18.1%), extrathyroidal extension (39.4% vs 14%), and more lymph node metastases (33.3% vs 12.1%). The incidence of distant metastases was higher for patients with FTC (28.8% vs 8.9%), and cause-specific survival rates were lower (p =.001). The locoregional control rates were not significantly different (p =.2). The 10 year cause-specific survival, freedom from distant metastasis, and locoregional failure figures for PTC compared with FTC were 92.1% vs 81%, 90.8% vs 72.3%, and 78.5% vs 83%. CONCLUSIONS: Although patients with PTC tend to have more advanced locoregional disease compared with those with FTC, the likelihood of locoregional control is similar, and the probability of cure is better. PMID- 12112542 TI - Supraomohyoid neck dissection in the management of cervical lymph node metastases of squamous cell carcinoma of the lower lip. AB - BACKGROUND: Supraomohyoid neck dissection (SOHND) is generally considered an adequate staging procedure in selected patients with squamous cell carcinoma (SCC) of the lip and oral cavity, with clinically negative nodes in the neck that are at increased risk for occult metastatic disease. The potential role of SOHND as a therapeutic surgical procedure for cervical metastasis limited to level I is controversial. METHODS: A series of 44 patients with clinical cervical lymph node metastases at level I from SCC of the lower lip is reported to evaluate the results of a treatment protocol consisting of therapeutic SOHND on indication followed by radiotherapy. RESULTS: Regional recurrences were observed in four (9%) patients. All recurrences developed within the SOHND dissected area only. CONCLUSIONS: A therapeutic SOHND, on indication followed by radiotherapy, can be an oncologically sound and effective procedure in the management of regional lymph node metastases at level I from SCC of the lower lip. PMID- 12112543 TI - The impact of second opinion surgical pathology on the practice of head and neck surgery: a decade experience at a large referral hospital. AB - BACKGROUND: A second review of histopathologic diagnoses is a quality assurance practice that helps expose diagnostic errors and guide management of patients being referred from outside hospitals. Identification of anatomic regions and specimen types that are prone to diagnostic error will be helpful in guiding policy decisions regarding mandatory second opinion surgical pathology. METHODS: All available outside pathology reports were retrieved for patients referred to The Johns Hopkins Hospital Department of Otolaryngology-Head and Neck Surgery between January 1, 1990, and January 1, 2000. The outside diagnosis was compared with diagnosis rendered at the referral hospital. A discrepant diagnosis was regarded as any change resulting in a significant modification in therapy or prognosis. RESULTS: Of the 814 cases reviewed, the second opinion surgical pathology diagnosis resulted in 54 (7%) changed diagnoses. Of the changed diagnosis, 13 (24%) involved a change from a benign to a malignant diagnosis; 8 (15%) involved a change from a malignant to a benign diagnosis; and 33 (61%) involved a change in tumor classification. Follow-up information supported the second opinion diagnosis in 41 of 43 cases (95%). CONCLUSIONS: In a consequential number of cases, second opinion surgical pathology results in major therapeutic and prognostic modifications for patients sent to large referral hospitals for head and neck oncologic surgery. PMID- 12112544 TI - Defects in G1-S cell cycle control in head and neck cancer: a review. AB - Tumors gradually develop as a result of a multistep acquisition of genetic alterations and ultimately emerge as selfish, intruding and metastatic cells. The genetic defects associated with the process of tumor progression affect control of proliferation, programmed cell death, cell aging, angiogenesis, escape from immune control and metastasis. Fundamental cancer research over the last thirty years has revealed a multitude of genetic alterations which specify more or less separate steps in tumor development and which are collectively responsible for the process of tumor progression. The genes affected play in normal cells a crucial role in control over cell duplication and the interaction between cells, and between cells and their direct surrounding. This is illustrated on control during the G1/S phase of the cell cycle by its ultimate regulators: cyclins and cyclin dependent kinases. These proteins not only control the transition through the G1/S phase of the cell cycle, but also serve as mediators of the interaction between cells, and between cells and their surrounding. Defaults in the regulation of these proteins are associated with tumor progression, and, therefore, serve as targets for therapy. Defaults in those genes are found in various tumor types, although some of those prevail in particular tumor types. In this review emphasis is given to the defaults that occur in head and neck cancer. PMID- 12112545 TI - "Tongue sandwich" bolster for skin graft immobilization. AB - BACKGROUND: Because of surface irregularities and continuous movement of the tongue, predictable immobilization of split-thickness skin grafts (STSGs) for tongue defects is difficult to achieve. METHODS: A novel composite, bilayer foam bolster was used to immobilize a STSG after reconstruction of more than 80% of the tongue mucosa after resection of a squamous cell carcinoma and extensive leukoplakia. Dorsal and ventral bolster components were placed over the STSG and affixed using transglossal, through-and-through sutures. RESULTS: The composite foam bolster provided uniform compression along the highly irregular and mobile skin-grafted surface. Graft survival was excellent, and there were no complications. CONCLUSIONS: The "tongue sandwich" bolster is quickly and easily fabricated, immobilizes the tongue in a fully expanded position, and provides excellent apposition of STSGs to highly irregular and vascular surfaces. PMID- 12112546 TI - Pendred's syndrome with goiter and enlarged vestibular aqueducts diagnosed by PDS gene mutation. AB - BACKGROUND: Pendred's syndrome (PS) is an autosomal recessive disorder characterized by goiter and congenital sensorineural hearing loss. Recent advances in molecular biology revealed the gene responsible for PS (PDS) and provided an important aid for the diagnosis of this condition. METHODS: A case of PS with huge goiter and congenital hearing impairment was diagnosed by mutational analysis of the PDS gene. RESULTS: Physical examination and computer tomography CT revealed a diffuse swelling of the thyroid gland. Thyroid function tests were normal, and the perchlorate discharge test was negative. Audiologic examination confirmed sensorineural hearing loss, and temporal bone CT revealed bilateral enlarged vestibular aqueducts. The mutational analysis revealed that the patient was homozygous for His 723 Arg (2168A-->G) in exon 19, a missense mutation. CONCLUSIONS: The results of thyroid function tests in PS patients are usually normal, and the positive perchlorate discharge test has been used for the diagnosis. However, this is a nonspecific test and is not sensitive enough for PS. In our case, despite a negative perchlorate test, the patient was diagnosed by mutational analysis and received total thyroidectomy to relieve respiratory distress caused by thyroid enlargement. This is the first report of a mutation detected in the thyroid tissue and clearly shows that the mutation caused histopathologic change in that gland. PMID- 12112547 TI - Prognostic factors of clinically stage I and II oral tongue carcinoma-A comparative study of stage, thickness, shape, growth pattern, invasive front malignancy grading, Martinez-Gimeno score, and pathologic features. AB - PURPOSE: This study aims at evaluation of the different prognostic models, including stage, tumor thickness, shape, malignancy grading of tumor invasive front, Martinez-Gimeno score, and pathologic features in the prediction of subclinical nodal metastasis, local recurrence, and survival of early T1 and T2 oral tongue squamous cell carcinoma. The results will have important implication for the management of patients. PATIENTS AND METHODS: Seventy-two clinically T1 and T2 glossectomy specimens of oral tongue carcinoma were serially sectioned in 3-mm thickness for the evaluation of various pathologic features. The prognostic value in the prediction of subclinical nodal metastasis, local recurrence, and survival of different models were compared. RESULTS: Among all the tumor parameters and predictive models being evaluated, tumor thickness was the only significant factor that had significant predictive value for subclinical nodal metastasis, local recurrence, and survival. With the use of 3-mm and 9-mm division, tumor of up to 3-mm thickness has 8% subclinical nodal metastasis, 0% local recurrence, and 100% 5-year actuarial disease-free survival; tumor thickness of more than 3 mm and up to 9 mm had 44% subclinical nodal metastasis, 7% local recurrence, and 76% 5-year actuarial disease-free survival; tumor of more than 9 mm had 53% subclinical nodal metastasis, 24% local recurrence, and 66% 5-year actuarial disease-free survival. CONCLUSIONS: Tumor thickness should be considered in the management planning of patients with early oral tongue carcinoma. PMID- 12112548 TI - The addition of mood and anxiety domains to the University of Washington quality of life scale. AB - BACKGROUND: There are numerous head and neck specific quality of life questionnaires, each having its own merits and disadvantages. The University of Washington questionnaire has been widely used and is notable by the inclusion of a shoulder dysfunction domain, domain importance ratings, and patient free text. It is short, simple to process, and provides clinically relevant information. However, it has lacked any psychological dimension of quality of life. The aim of this study was to report the inclusion of two psychological domains (mood, anxiety) to the most recent refinement of the questionnaire (version 3). METHOD: A cross-sectional survey was performed in April 2000. Questionnaires were sent to 183 patients alive and disease free after surgery for oral and oro-pharyngeal malignancy. Replies were received from 145 patients (79% response rate). RESULTS: The new domains (mood and anxiety) correlated significantly with the emotional functioning domains from the EORTC C30 and with the pain and appearance domains of UW-QOL. There were also significant correlations between the "global quality of life" item and the two new domains. Mood (p =.005) and anxiety (p <.001) scores were associated with patient age but with no other clinicodemographic variable. CONCLUSION: The addition of mood and anxiety domains makes the UW-QOL version 4 a single broad measure suitable for effective health-related quality of life evaluation in the routine clinical setting. PMID- 12112549 TI - Impact of palatal prosthodontic intervention on communication performance of patients' maxillectomy defects: a multilevel outcome study. AB - BACKGROUND: Obturators have been developed for surgical defects caused by cancer of the maxillary sinus and alveolar ridge. Outcome research is necessary to develop evidence-based practice guidelines. METHODS: Thirty-two consecutively treated maxillectomy patients seen in the Facial Prosthetics Clinic at UNMC from 1994 through 1996 had their defects obturated for 1 month when speech intelligibility, speaking rate, nasality, and communication effectiveness were measured. RESULTS: With the obturator removed, mean speech intelligibility was 61%, speaking rate was 138 words per minute, and nasality was rated as 5.8 on a 0 7 point scale. With the obturator inserted, mean speech intelligibility was 94%, speaking rate was 164 words per minute, and nasality was rated as 1.6. The mean self-perception of communication effectiveness was 75% of what it was before the diagnosis of cancer. CONCLUSIONS: Obturation is an effective intervention for defects of the maxillary sinus and alveolar ridge on speech performance. Variations in effectiveness were noted based on site of defect and patient satisfaction with the intervention. PMID- 12112550 TI - Interval pathologic assessments in patients treated with concurrent hyperfractionated radiation and intraarterial cisplatin (HYPERRADPLAT). AB - BACKGROUND: To minimize surgical morbidity, surgery should be performed within 2 to 3 months of completion of radiation therapy with or without chemotherapy. Pathologic demonstration of cancer at this interval is commonly used to justify early surgical salvage of residual primary head and neck cancer. These assumptions regarding head and neck cancer in patients treated with concurrent hyperfractionated radiation therapy and intraarterial supradose cisplatin (HYPERRADPLAT) have never been evaluated. METHODS: Post-HYPERRADPLAT clinical and pathologic findings in 42 patients with stage III and IV head and neck cancer were compared with their disease outcomes. All patients underwent an interval analysis of response at 6 to 10 weeks after completion of therapy, 28 of these patients had biopsies of the primary tumor site performed. RESULTS: Clinical findings of cancer with pathologic confirmation up to 4 months after therapy can be associated with eventual complete response (CR). Pathologic CR's from deep incisional biopsies can be associated with recurrent disease within 2 months. Six HYPERRADPLAT-treated patients underwent interval surgical resection of primary disease, and only the four patients with cancer identified in the resection specimen died of recurrent disease. CONCLUSION: In patients treated with HYPERRADPLAT, interval clinical and pathologic assessments may be misleading. Only observation of progressive disease is an accurate predictor of local failure. New evaluation techniques such as metabolic imaging and molecular analysis warrant exploration as tools for interval cancer evaluations. PMID- 12112551 TI - Impact of age on clinical care pathway length of stay after complex head and neck resection. AB - OBJECTIVE: This article investigates the effect of patient age on postoperative pathway length of stay (LOS) for head and neck surgery. Aggregate clinical results for 43 patients, enrolled in the CCP from June 1996-July 1997, are described. Patient age, comorbid status, and postoperative complications are analyzed with respect to impact on LOS. SETTING: Tertiary level academic medical center with an operative otorhinolaryngology volume of approximately 1200 cases per year. PATIENTS: Forty-three patients undergoing head and neck resection with primary closure, local flap, or free flap closure were enrolled on CCP from June 1996-July 1997. Length of stay, frequency of selected aggregated comorbidities, and frequencies of complications are analyzed with nonparametric statistics. A pre-pathway group of 87 consecutive patients is used for comparison. MAIN OUTCOME MEASURES: Length of stay and age. RESULTS: Median actual LOS post-pathway for the patients enrolled in the first year of the pathway was 8 days. This met the CCP target and improved on pre-pathway LOS by 5 days (p <.001). The average LOS increased 25% from 8 days to 10 days for patients older than 65 years of age (p =.036, Mann-Whitney U test). Presence of a comorbidity and a complication concomitantly was statistically associated with increased LOS though not with advancing age (p =.003). CONCLUSIONS: The CCP-reported performance improvement achieved by this pathway suggests improved resource use, and improved patient outcomes are achieved for postoperative care of head and neck surgery patients. Our experience suggests that advancing age creates a clinically significant increase in resource use represented by our finding of increasing LOS. This finding warrants further investigation. PMID- 12112552 TI - Chemotherapy with or without radiotherapy in patients with locoregionally recurrent nasopharyngeal carcinoma. AB - BACKGROUND: Treatment of locoregionally recurrent nasopharyngeal carcinoma (NPC) is challenging because of prior radiotherapy, morbidities from disease recurrence, and limited therapeutic options available. METHODS: A retrospective study of patients with locoregionally recurrent NPC. RESULT: Between March 1994 and December 1999, there were 42 patients; most were Chinese (98%) men (81%) with undifferentiated NPC (86%). A repeat course of radiotherapy was feasible in 20 patients and given concurrently with cisplatin followed by adjuvant cisplatin/5 fluorouracil (PF) (group 1). The remaining 22 (group 2) received palliative chemotherapy (PF) with a response rate of 50%. Significant morbidities resulted from cranial nerve palsies. The 2-year progression-free survival of patients in group 1 was expectedly better (58% vs 38%). Six (14%) developed systemic metastases at 12 months (median) from first recurrence. CONCLUSION: Concurrent chemoradiotherapy for locoregional recurrent NPC seems promising. The morbidity experienced resulted from locoregional disease with few progressing to develop systemic involvement. PMID- 12112554 TI - A scanning electron microscopic study on thrombogenicity of intraarterial catheters for chemotherapeutic treatment in head and neck cancer. AB - BACKGROUND: The purpose of this study was to assess the impact of standard anticoagulation and intermittent catheter irrigation on clot formation on intraarterial chemotherapeutical catheters. METHODS: Sixteen nonheparinized catheters were placed in the carotid vessels of 10 patients. Ten catheters were perfused with chemotherapeutic drugs; six catheters were not perfused. Patients received LMWH anticoagulation; catheters were irrigated with boluses of heparinized saline daily. Catheters were retrieved for SEM evaluation after a mean intravessel placement interval of 21 days. RESULTS: All samples demonstrated accumulation of thrombotic debris on luminal sides and outsides of catheters. Obliteration was seen in three samples. Detachment of thrombus fragments was present in several specimens. No significant dependence of clot formation on placement interval and chemotherapeutic perfusion was calculated. CONCLUSIONS: Standard anticoagulation was ineffective in clot prevention. Heparinized catheters might potentially reduce the risk of clot formation. A delivery system should be engaged for continuous irrigation of catheters with heparinized saline. PMID- 12112553 TI - Swallow function and perception of dysphagia in patients with head and neck cancer. AB - BACKGROUND: The relationship between subjective complaints of dysphagia and objective measures of swallow function in patients with cancers of the oral cavity, pharynx, or larynx, treated with radiotherapy +/- chemotherapy has not been well documented in the literature. METHODS: Swallowing function in 132 patients with various lesions was evaluated using videofluoroscopy and analyzed by patient complaint of dysphagia grouping. RESULTS: Patients with complaints of dysphagia demonstrated significantly worse swallow function as indicated by lower oropharyngeal swallow efficiency (OPSE), longer transit times, larger residues, and more swallows with aspiration. Patients with complaints of dysphagia also tended to take less of their nutrition by mouth and less variety of food consistencies in their diet compared with those without complaint. CONCLUSIONS: Patients were able to perceive decrements in their swallowing function as dysphagia and may have limited their oral intake in response to that perception. The ability to accurately perceive swallowing function may be useful for self monitoring changes in dysphagia status during a course of swallow therapy. PMID- 12112555 TI - Expression of glutathione s-transferase pi in benign mucosa, Barrett's metaplasia, and adenocarcinoma of the esophagus. AB - BACKGROUND: Glutathione s-transferase pi (GSTpi) is an enzyme that provides cellular protection against redox-mediated damage by free radicals, which have been implicated in carcinogenesis. METHODS: Forty-three consecutive specimens from 19 patients were reviewed to identify samples of squamous mucosa, Barrett's metaplasia, adenocarcinoma, and peritumoral inflammation. Serial sections were stained with an anti-GSTpi polyclonal antibody, and GSTpi expression was quantified for each histologic group. RESULTS: GSTpi expression was diminished in peritumoral mononuclear inflammatory cells (p <.001) compared with squamous epithelium, Barrett's metaplasia, or adenocarcinoma. Barrett's metaplasia exhibited decreased GSTpi expression compared with squamous mucosa (p =.045). GSTpi expression by >50% of adenocarcinoma cells was associated with an increased risk (2.25x) of disease at last follow-up. CONCLUSIONS: GSTpi is prominently expressed in esophageal squamous mucosa and adenocarcinoma. Mononuclear cells may be susceptible to oxidative damage secondary to weak GSTpi production. GSTpi may protect the tumor cells themselves from the cytotoxic effects of free radicals. The biochemical role of GSTpi expression in malignant transformation deserves further investigation. PMID- 12112556 TI - Primary neck management among patients with cancer of the oral cavity without clinical nodal metastases: A decision and sensitivity analysis. AB - BACKGROUND: A standardized neck management strategy for oral cancer patients without clinical nodal metastases remains to be established. Consequently, a decision and sensitivity analysis of two neck management protocols, involving either prophylactic neck dissection or careful observation, was conducted using the Oral Cancer Registry of Kyushu, Japan. METHODS: We calculated probabilities of subclinical nodal metastases and 5-year survival using the registry data. A two-way sensitive analysis was conducted using the probabilities and parameters of the complete nodal metastasis resection rate (x) and a utility rating that describes the health state induced by dissection (y) compared with the neck condition in a careful-observation group. RESULTS: We solved the threshold curve for y and x for the expected utility between the two groups. The results showed that prophylactic neck dissection must guarantee a complete resection of subclinical nodal metastases with no disadvantage to health state to be evaluated as equally satisfactory as careful observation. CONCLUSIONS: Careful observation involving standardized systematic preoperative and postoperative screening of the neck seems preferable to prophylactic neck dissection for oral cancer patients without subclinical nodal metastases. PMID- 12112557 TI - PCR detection of circulating tumor cells in nasopharyngeal carcinoma patients with distant metastasis: effect of enzyme and sampling. AB - BACKGROUND: Nasopharyngeal carcinoma (NPC) has a high potential to develop distant metastasis after radiotherapy. Cytokeratin 19 (CK-19) mRNA has been frequently used as a marker in the detection of circulating tumor cells of epithelial origin, but has rarely been investigated in NPC. This study was performed to evaluate the effect of blood sampling and different Taq DNA polymerase on the results of nested reverse transcriptase-polymerase chain reaction (RT-PCR) assay. METHODS: Peripheral blood samples from a total of 37 NPC patients with well-documented distant metastasis (M1) were collected before treatment. Eighteen patients had more than one blood sampling. Five different Taq DNA polymerases were used to test the blood from 17 patients. Peripheral blood of 37 nonmetastatic (M0) NPC patients was tested by the same nested RT-PCR system using multiple Taq DNA polymerases to evaluate the impact of multienzyme assay in the prediction of subsequent distant metastasis. RESULTS: Among M1 NPC patients, the accumulative positive rates of CK-19 mRNA were 22.2%, 44.4%, 70.6%, 75.0%, and 80.0% when one, two, three, four, or five blood sampling were taken, respectively. The accumulative positive rates increased as the numbers of different enzymes increased-from 35.5% by one enzyme to 82.4% by five enzymes. Six of 37 M0 patients had distant metastasis develop after a median follow-up time of 20 months. The detection sensitivity for four-enzyme test (5 of 6 = 83.3%) is better than that of one-enzyme test (2 of 6 = 33.3%). CONCLUSIONS: Our data demonstrate that multiple blood sampling or using multiple enzymes for nested RT-PCR assay significantly enhances the sensitivity in the molecular diagnosis of NPC metastasis. PMID- 12112558 TI - Human papillomavirus infection and cyclin D1 gene amplification in laryngeal squamous cell carcinoma: biologic function and clinical significance. AB - BACKGROUND: Human papillomavirus (HPV) infection is suspected to be a risk factor for head and neck, and in particular for laryngeal, carcinogenesis. Cyclin D1 gene (CCND1) overexpression and amplification have been shown to play a role as prognostic factors in many human cancers, among which are head and neck cancers. METHODS: A literature review of the role in head and neck cancers of HPV infection and CCND1 overexpression and amplification was undertaken. We have evaluated the extent of the current knowledge in this field under the light of recent acquisitions, in particular, about a correlation between HPV infection, a suspected risk factor, and CCND1 amplification, a frequent mutation (about 20% of laryngeal cancers) and a prognostic factor in laryngeal SCC. RESULTS AND DISCUSSION: The significant correlation between HPV infection and CCND1 amplification supports the hypothesis of the involvement of HPV infection in laryngeal carcinogenesis and suggests that HPV positive laryngeal cancers may constitute a different subset of tumors with a peculiar molecular pattern and thus with a different clinical behavior. HPV infection may be considered a synergistic risk factor with smoking and/or alcohol consumption to be investigated in heavy smokers and drinkers, thus contributing to the identification of patient at high-risk for the development of laryngeal cancer who should undergo strict follow-up and primary and secondary prevention. PMID- 12112559 TI - Desmoplastic melanoma of the lip. AB - BACKGROUND: This retrospective study looks at the prognosis of desmoplastic melanoma of the lip, correlating it with the clinical course, treatment, and patterns of failure. METHOD: Twenty-two patients with desmoplastic melanoma of the lip were seen at the University of Texas M. D. Anderson Cancer Center from 1965 to 1998. RESULTS: Three disease groups: (I) untreated tumor (3 patients), (II) excisional scar (10 patients), and (III) locoregional recurrence (9 patients). Group I had two cures and one failure. In group II six had no recurrences, and there were four failures. In group III, all patients failed. Ten patients (45%) had no evidence of disease, of which three (30%) had an initial misdiagnosis. Twelve patients (55%) died of disease or were living with disease, of which eight (67%) had an initial misdiagnosis. CONCLUSIONS: Desmoplastic melanoma of the lip is often misdiagnosed and, therefore, inappropriately treated with multiple recurrences and poor prognosis. Accurate diagnosis and combined treatment may improve local control and survival. PMID- 12112560 TI - Management of an acquired tracheoesophageal fistula with a fascial free flap. AB - BACKGROUND: Failure in the primary repair of a benign acquired tracheoesophageal fistula limits the operative options available at revision. Use of a fascial free flap to treat this condition has not been previously reported. METHODS: We review the case of a patient who had a tracheoesophageal fistula develop after percutaneous tracheostomy, who had failed previous primary repair with strap muscle interposition. RESULTS: A radial forearm fascial free flap was used at revision and resulted in resolution of the fistula. CONCLUSION: Use of a fascial free flap to address a persistent acquired tracheoesophageal fistula, when an accompanying stenotic segment is not present, is a viable treatment option. PMID- 12112562 TI - Porotic hyperostosis as a marker of health and nutritional conditions. PMID- 12112561 TI - Kaposiform hemangioendothelioma of the external auditory canal in an adult. AB - BACKGROUND: Kaposiform hemangioendothelioma is an uncommon vascular tumor initially reported to occur exclusively in children. METHODS: The presentation, pathologic evaluation, and management of an unusual case of kaposiform hemangioendothelioma is presented and discussed. RESULTS: A 27-year-old HIV negative man was initially seen with a reddish nodule located in the outer third of the external auditory canal. Histologically, the tumor was composed of spindle shaped cells arranged in short fascicles associated with small endothelial-like vascular spaces, similar in appearance to Kaposi's sarcoma. The lesion was locally excised but recurred 1 month later; then radiation therapy was performed. The patient remains well at 5-year follow-up. CONCLUSIONS: Recognition of kaposiform hemangioendothelioma is important to avoid possible confusion with a variety of vascular neoplasms with different biologic potential. This case presented some diagnostic difficulty because of the age of the patient and the unusual location of the lesion and had to be mainly distinguished from Kaposi's sarcoma. PMID- 12112563 TI - New method to measure minisatellite variant repeat variation in population genetic studies. AB - The classical analysis of minisatellite variant repeat (MVR) variation using modular structures is limited by the lack of knowledge of the mutational process involved in the evolution of most of the minisatellites. In this study a new method to measure MVR variation and to calculate genetic distances using MVR codes is proposed. The method is based on the statistical similarity of MVR patterns and considers the complete variability of the minisatellite, enabling meaningful comparisons of closely related populations. As an example, the method has been applied to analyze variation in MSY1 (DYF155S1) in five sets of data from European and North African populations. PMID- 12112564 TI - Maximum likelihood estimates of admixture in Northeastern Mexico using 13 short tandem repeat loci. AB - Tetrameric short tandem repeat (STR) polymorphisms are widely used in population genetics, molecular evolution, gene mapping and linkage analysis, paternity tests, forensic analysis, and medical applications. This article provides allelic distributions of the STR loci D3S1358, vWA, FGA, D8S1179, D21S11, D18S51, D5S818, D13S317, D7S820, CSF1PO, TPOX, TH01, and D16S539 in 143 Mestizos from Northeastern Mexico, estimates of contributions of genes of European (Spanish), American Indian and African origin in the gene pool of this admixed Mestizo population (using 10 of these loci); and a comparison of the genetic admixture of this population with the previously reported two polymorphic molecular markers, D1S80 and HLA-DQA1 (n = 103). Genotype distributions were in agreement with Hardy Weinberg expectations (HWE) for almost all 13 STR markers. Maximum likelihood estimates of admixture components yield a trihybrid model with Spanish, Amerindian, and African ancestry with the admixture proportions: 54.99% +/- 3.44, 39.99% +/- 2.57, and 5.02% +/- 2.82, respectively. These estimates were not significantly different from those obtained using D1S80 and HLA-DQA1 loci (59.99% +/- 5.94, 36.99% +/- 5.04, and 3.02% +/- 2.76). In conclusion, Mestizos of Northeastern Mexico showed a similar ancestral contribution independent of the markers used for evolutionary purposes. Further validation of this database supports the use of the 13 STR loci along with D1S80 and HLA-DQA1 as a battery of efficient DNA forensic markers in Northeastern Mestizo populations of Mexico. PMID- 12112565 TI - Dissimilarities in the process of formation of Curiau, a semi-isolated Afro Brazilian population of the Amazon region. AB - The genetic consequences of the social policy of the past in relation to the formation of Afro-Brazilian societies are interesting and have been studied at various biological levels (classical polymorphisms and the mitochondrial and nuclear levels. These allow the estimation of the contribution of African genes and the participation of other ethnic groups in the formation of these communities. With this objective, uniparental systems of exclusively maternal (mtDNA) or paternal (Y-DNA) inheritance in the Curiau community were analyzed. The results demonstrate a differential contribution of the maternal and paternal genetic systems. Thirty-three sequences were identified by mtDNA analysis; 53% showing an African and 47% an Amerindian origin. For the paternal system, 57% were of African, 37% of European, and 6% of Amerindian origin. PMID- 12112566 TI - Effects of health information in youth on adult physical activity: 20-year study results from the Amsterdam growth and health longitudinal study. AB - In the Amsterdam Growth and Health Longitudinal Study (AGAHLS), a group of apparently healthy males and females (n = 200) were interviewed about their physical activities on eight separate occasions over a period of 20 years between 13 and 33 years of age (multi-measured group: MM). Information about their health was given based on their personally measured lifestyle (activity, diet, smoking) and biological risk characteristics for chronic diseases (medical check-ups). A comparable group of boys and girls (n = 200) was only measured on two occasions (bi-measured group: BM): at 13 and 33 years. Physical activity was estimated with a structured interview. Total physical activity and sports activity were estimated in three intensity levels (light, moderate, and heavy). It was hypothesized that the eight repeated medical check-ups with health information in the MM group would result in a healthier lifestyle with respect to the determinants and levels of habitual physical activity compared to the BM group. Contrary to the hypothesis, males and females in the BM group showed a significantly higher increase or a lower decrease in physical activities compared to the MM group. This negative effect on the physical activity pattern at 33 years in the MM group may have been caused by more underreporting of physical activities than in the BM group. In conclusion, there does not appear to be a significant effect of long-term (multi-measured) health information with medical check-ups during adolescence and young adulthood on level of physical activity in males and females at 33 years of age. PMID- 12112567 TI - Midsagittal facial soft-tissue growth of French Canadian adolescents. AB - This study examines changes in 12 midline soft-tissue thicknesses from the forehead, nose, lip, and chin regions in girls and boys from 10 to 16 years of age. The soft-tissue changes are compared to changes in two hard-tissue distances (sella-nasion and nasion-menton). The subjects are from a mixed-longitudinal sample studied at the Montreal Human Growth Research Center in the 1960s and 1970s. Total sample size is 242 (from lateral cephalographs of 124 males and 118 females), with numbers varying by age and measurement. For hard-tissues, boys show clearly defined adolescent spurts, while girls display small velocity increases indicative of only very minor spurts. Forehead tissue thicknesses for both sexes change little and show no demonstrable growth spurts. For the nose and philtrum region, which have the greatest absolute soft-tissue growth changes, both boys and girls show adolescent spurts. Peak velocities are attained between 13 and 14 years in boys and between 11.9 and 12.5 years in girls. Boys appear to have small adolescent spurts for upper (13.7 years) and perhaps lower lip thicknesses. Neither sex displays clear evidence for adolescent spurts in the chin region. PMID- 12112568 TI - Body mass index, overweight and obesity in married and never married men and women in Poland. AB - The study examined the relationship between marital status and the body mass index (BMI) and the prevalence of overweight and obesity in the Polish population. The sample included 2,266 men and 4,122 women, 25-60 years of age, who were occupationally active inhabitants of Wroclaw, in southwestern Poland. Marital status was defined by two categories: never married and presently married, and two groups in each category were established on the basis of educational level: well-educated (12 or more years in school) and poorly educated (less than 12 years in school). The subjects were also divided into four age groups: 25-30, 31-40, 41-50, and 51-60 years. Height and weight were measured and the BMI was calculated. Three categories of the BMI were established: normal, BMI < 25.0 kg/m(2), overweight, BMI > or = 25 < 30 kg/m(2), and obese, BMI > or = 30 kg/m(2). In each age and educational group, married individuals had a higher BMI than those who were never married. With the exception of well-educated males 51 60 years, differences in the BMI between married and never married individuals increased with age. In general, married men and women were more likely to be overweight and obese than never married individuals. The results indicated a significant association (P < 0.001) between marital status and the BMI in both sexes. After age, marital status was the most important predictor of overweight/obesity among men (P < 0.001), whereas educational level did not have a significant role. Among women, age, marital status, and education were significantly (P < 0.001) related to the BMI. PMID- 12112569 TI - Somatic comparisons of South Korean children and youths born and reared in a rural area with the descendants of rural to urban migrants. AB - During April-May 2000 somatic data were collected on 236 males and 191 females ages 6, 9, 13, and 15 years residing in rural areas around the communities of Ankye, Euisung, and Kunwi in Kyungsang Puk Do province, South Korea, and 237 males and 219 female of the same ages born and reared at Taegu in families of "rural to urban migrants." Comparisons were made between urban and rural groups for measures of body size and form, skinfold thickness, the body mass index (BMI), and estimated arm muscle area (ARM). Age at menarche was obtained from school records for the 13- and 15-year-olds. The data were analyzed in separate 2 (urban-rural) x 4 (age) analyses of variance with an alpha level of P < 0.01. Age differences were significant for all dimensions. There was a significant main effect for urban-rural differences in arm girth, upper limb index, and ARM. Rural males were larger in these variables. Among females, only arm girth was significantly different. Age at menarche was significantly earlier in the urban sample (12.6 years) than in the rural sample (13.0 years). Compared with both recent and earlier data for South Korea, present-day 6-, 9-, 13-, and 15-year-old males and females are taller and heavier at every age, indicating secular gains. PMID- 12112570 TI - Age-related decrease in energy expenditure at rest parallels reductions in mass of internal organs. AB - This study explores to what extent the mass of internal organs may impact the age related decrease in energy expenditure at rest (EErest). The relationship between direct measurements of EErest in elderly women and predicted EErest based on equations deriving from the metabolic activity in tissue from younger women were also elucidated. Body composition of elderly women was measured by an impedance method. EErest was measured by the Douglas bag method after an overnight fast. These data were compared with predicted values of EErest based on equations derived from studies in younger women. The mass of internal organs was obtained from autopsy material. Young women (mean age 31.7 years, range 14-60, n = 104) and elderly women of 65 years (n = 22), 75 years (n = 26), and 85 years (n = 31) participated in this study. Autopsy data were obtained from women (n = 238) from the same birth cohorts as the elderly women who died at ages 42-87 years. EErest showed a progressive age-related decline, which appeared to parallel a similar reduction in the mass of internal organs derived from autopsy material of women who died at the same ages. In contrast, FFM was identical in the group of 65 and 75-year-old women, but was lower in the 85-year-old women. Predicted and measured EErest revealed a strong correlation in elderly women. Modest reductions in the mass of internal organs with a high metabolic rate appear to contribute markedly to the decline in EErest observed in aging. Further, it is also possible to predict EErest from the body composition of elderly women using equations developed from younger women. PMID- 12112571 TI - Lung function of Han Chinese born and raised near sea level and at high altitude in Western China. AB - Forced vital capacity (FVC), forced expiratory volume at 1 second (FEV(1)), and FEV(1)/FVC ratios were determined for 531 individuals of Han Chinese descent living at low altitude (250 m) near Beijing and for 592 individuals of Han descent who were born and raised at three high altitudes (3,200 m, 3,800 m, 4,300 m) in Qinghai Province, P.R.C. The study included males and females, ages 6-51 years. Thorax widths, depths, and circumferences of Han females and males born and raised at high altitude are similar to those of low-altitude Han. On the other hand, high-altitude children and adolescents have larger relative sitting heights, indicating greater thorax lengths. After adjusting for this variation in morphology, mean FVC values among 6-21 year-old Han at high altitude are only between 136 mL (for females) and 173 ml (for males) greater than those determined at low altitude but the differences are statistically significant and are maintained consistently throughout the growth period. These data indicate that growth at high altitude produces small-to-moderate increases in lung volumes (about 6%) relative to genetically similar groups growing up at low altitude. In addition, there is no evidence that lung volume growth is accelerated relative to morphological growth among Han children born and raised at high altitude. Adults, 22-51 years, also show greater FVC values at high altitude but the size of the increase relative to Han at low altitude is variable (3% in males and 11% in females). Greater lung function at high altitude is unlikely to result from increased activity or lower pollution, and thus appears to be primarily a result of development in a hypoxic environment. Differences in FVC and FEV(1) at 3,200 m, 3,800 m, and 4,300 m are generally not significant, so that living at altitudes between 3,200 m and 4,300 m appears to have little additional effect on volumetric growth. PMID- 12112572 TI - Lipoprotein lipase and APOE/APOC-I/APOC-II gene cluster diversity in native Brazilian populations. AB - Allele and haplotype frequencies for the T-93G, Hind III, and Pvu II variants of the lipoprotein lipase gene (LPL), and Hpa I and Ava II restriction site polymorphisms (RSP) of the APOE/C-I/C-II gene cluster were determined in 143 individuals from five Brazilian Indian tribes. These results were integrated with those previously reported for APOE. Marked interethnic variability occurs in these sites. A strong linkage disequilibrium was observed between the APOE and APOC-I loci (D' = 0.81; P < 0.00001). Linkage disequilibrium between the Hind III and Pvu II RSPs of the LPL gene was also observed (D' = 1; P < 0.001), but none of these RSPs were in linkage disequilibrium with the T-93G mutation. Considering both loci, heterozygosity was estimated as 0.45, but it was lower in the Xavante and Surui populations, in accordance with the historical and biodemographical data of these Amerindians. The results reported here may have implications for understanding interpopulation differences in lipid levels and coronary heart disease prevalences. PMID- 12112573 TI - Seasonal variation in the dietary adequacy of highland Pwo and Sgaw Karen (Thailand). AB - The present report describes seasonal variability (postharvest, preharvest, harvest) in the dietary adequacy of highland Karen farmers (38 Pwo and 51 Sgaw mother-weaned child pairs) in Thailand. Both the Pwo and Sgaw exhibit little functionally significant seasonal variation in either energy or protein intakes, although there is significant seasonal variation in other nutrients. Sgaw Karen mothers and their weaned children had significantly higher intakes of energy, protein, and seven nutrients than their Pwo counterparts during all three study seasons. The cause of this difference appears to be related to the better socioeconomic status of the Sgaw, as well as to differences in belief systems that result in a greater utilization of wild resources by the Sgaw. There were no significant differences in energy intakes between mothers and their children in either ethnic group, while children tended to have significantly greater intakes of protein but significantly lower intakes of other nutrients. These differences in dietary patterns appear to be due to differences in dietary density and dietary preferences, rather than to foods being differentially directed towards adults. PMID- 12112574 TI - Skeletal clues apparently distinguishing Waldenstrom's macroglobulinemia from multiple myeloma and leukemia. AB - This study was conducted to characterize macroscopically and by conventional radiography the bony lesions in a case of Waldenstrom's macroglobulinemia and to compare and contrast it with those of the other major hematologic lymphoproliferative disorders, multiple myeloma and leukemia. Two varieties of lytic skeletal lesions were found in Waldenstrom's macroglobulinemia. One was sharply defined, spheroid lesions with smooth borders and effaced/erased trabeculae. The second was in the form of coalescing pits (holes) with smooth, minimally remodeled edges. The appearance combined features of multiple myeloma and leukemia, but were mutually exclusive in those diseases. Spheroid lesions with effaced edges were absent in leukemia, while pits were absent in multiple myeloma. Fronts of resorption were not noted in Waldenstrom's macroglobulinemia. The combination of some of the features of leukemia and myeloma appear to allow recognition of Waldenstrom's macroglobulinemia. PMID- 12112575 TI - Two novel genes expressed in Xenopus germ line: characteristic features of putative protein structures, their gene expression profiles and their possible roles in gametogenesis and embryogenesis. AB - We compared the secondary spermatogonia and the primary spermatocytes of Xenopus for the proteins in their microsomal fractions and identified a newly synthesized protein (94 kDa) and three other proteins (99, 85, and 72 kDa) which increased their amount after entering the meiotic phase. These four proteins were used as antigens to produce polyclonal antibody which was found to react with the four proteins as well as two other proteins (208 and 60 kDa). Immunoscreening of Xenopus testis cDNA library with this polyclonal antibody yielded two cDNA clones (Xmegs and Xtr) encoding novel proteins. Xmegs mRNA was specifically expressed in the spermatogenic cells from the mid-pachytene stage to completion of two meiotic divisions. The putative Xmegs protein contained 19 tandem repeats of 26 amino acid residues rich in proline as well as potential phosphorylation sites (i.e., serine and threonine residues). Around this repetitive area, we found five PEST sequences known as a proteolytic signal to target protein for degradation. The presence of PEST sequences was believed to allow protein levels to closely parallel mRNA abundance. These results suggested the possible role of this novel protein in the regulation of two meiotic divisions specific to the spermatogenesis in a phosphorylation- and/or dephosphorylation-dependent manner. On the other hand, Xtr mRNA was expressed in both spermatogenic and oogenic cells except for round spermatids and the later stage cells. This mRNA was also expressed in the early stage embryos and its amount was kept constant from the St. I oocyte to the gastrula stage and decreased thereafter. The putative Xtr protein contained four complete and one partial tudor-like domains that were discovered in Drosophila tudor protein which plays an important role in PGC differentiation and abdominal segmentation. The characteristic expression profile of Xtr and the protein structure similar to the Drosophila tudor protein suggested its possible role in the progression of meiosis and PGC differentiation. PMID- 12112576 TI - EST analysis of gene expression in testis of the ascidian Ciona intestinalis. AB - To explore the gene expression underlying spermatogenesis, a large-scale analysis has been done on the cDNAs from testis of the ascidian, Ciona intestinalis. A set of 5,461 expressed sequence tags was analyzed and grouped into 2,806 independent clusters. Approximately 30% of the clusters showed significant sequence matches to the proteins reported in DDBJ/GenBank/EMBL database including a set of proteins closely related to the gene regulation during spermatogenesis, functional and morphological changes of spermatogenic cells during spermiogenesis, and physiological functions of sperm, as well as those with housekeeping functions commonly expressed in other cells. Some clones show similarities to the proteins present in vertebrate lymphocytes, suggesting a primitive immune system in ascidians. We have also found some genes that are known to participate in hormonal regulation of spermatogenesis in vertebrates. The large majority of the genes expressed in Ciona testis show no significant matches to known proteins and the further analysis of these genes may shed new light on the molecular mechanism of spermatogenesis and sperm functions. PMID- 12112577 TI - Regulated expression of heparin-binding EGF-like growth factor (HB-EGF) in the human endometrium: a potential paracrine role during implantation. AB - Heparin-binding epidermal growth factor (HB-EGF) is a recently identified member of the EGF growth factor family found to be expressed in the uterus of both mouse and human at the time of implantation. In the present study, we investigated the expression patterns of HB-EGF in normal cycling endometrium and compared its expression with the fertility-associated endometrial epithelial biomarkers alpha(v)beta(3) integrin, leukemia inhibitory factor (LIF) and homeobox gene, HOXA-10. RNase protection assay (RPA) using RNA made from endometrium collected from different phases of the menstrual cycle demonstrated increased HB-EGF expression during the mid-secretory phase, a pattern similar to, but slightly preceding the expression of alpha(v)beta(3) integrin and HOXA-10. In vitro studies demonstrated stimulation of HB-EGF expression by estradiol-17beta (E(2)) and progesterone (P(4)) alone or in combination in stromal cells. Combined treatment with E(2) + P(4) was, however, required to stimulate epithelial HB-EGF expression. In vitro experiments demonstrated the ability of HB-EGF to stimulate epithelial expression of the key endometrial proteins including LIF, HOXA-10, and the beta(3) integrin subunit. Each has previously been demonstrated to be an important epithelial biomarker expressed during the implantation window. In addition, conditioned media from endometrial stromal cells treated with E(2) + P(4) + relaxin mimicked the stimulatory effect of HB-EGF on epithelial expression of the beta(3) integrin subunit. The stimulatory effect of the stromal conditioned medium was blocked by antibodies that neutralize a known receptor for HB-EGF. These data suggest that uterine receptivity may be regulated in part by the stromal-derived HB-EGF. PMID- 12112578 TI - Molecular cloning and expression during spermatogenesis of a cDNA encoding testicular 11beta-hydroxylase (P45011beta) in rainbow trout (Oncorhynchus mykiss). AB - Cytochrome P450 1beta-hydroxylase (P450(11beta)) is an important steroidogenic enzyme for glucocorticoid and mineralocorticoid production in vertebrates. In teleosts, P450(11beta) also plays a role in the production of 11-ketotestosterone (11-KT), the predominant androgen in male fish. In this study, we cloned a cDNA encoding rainbow trout (Oncorhynchus mykiss) testicular P450(11beta). The cDNA contains 1,740 nucleotides that encode a protein of 551 amino acids which shares 65.2% homology with testicular P450(11beta) from Japanese eel, and 33-45% homology with adrenal P450(11beta) from rat, human, and frog. HEK293 cells transfected with an expression vector containing the rainbow trout P450(11beta) cDNA open reading frame showed 11beta-hydroxylating activity in the presence of exogenous testosterone. Analysis of tissue distribution by RT-PCR showed great abundance of P450(11beta) mRNA in testis and head kidney. In order to clarify the sites of P450(11beta) gene expression, cRNA in situ hybridization analysis was performed. Hybridization signals were detected in Leydig cells and head kidney inter-renal cells. The results of Northern blot analysis indicated a single 1.8 kb transcript encoding P450(11beta) in testis and in head kidney, suggesting that the testicular form of P450(11beta) may also be involved in cortisol production by inter-renal cells. Seasonal changes in total P450(11beta) mRNA levels in testes during spermatogenesis showed a pattern similar to that of plasma androgens. These results suggest that androgen production in male rainbow trout is partially regulated by changes in abundance of P450(11beta) mRNA. PMID- 12112579 TI - Novel double promoter approach for identification of transgenic animals: A tool for in vivo analysis of gene function and development of gene-based therapies. AB - Advances in vertebrate genetics have allowed studies of gene function in developing animals through gene knockout and transgenic analyses. These advances have encouraged the development of gene-based therapies through introduction of exogenous genes to enhance and/or replace dysfunctional or missing genes. However, in vertebrates, such analyses often involve tedious screening for transgenic animals, such as PCR-based genotype determinations. Here, we report the use of double-promoter plasmids carrying the transgene of interest and the crystallin-promotor-driven Green fluorescent protein (GFP) in transgenic Xenopus laevis tadpoles. This strategy allows a simple examination for the presence of GFP in the eyes to identify transgenic animals. PCR-based genotyping and functional characterization confirms that all animals expressing GFP in the eyes indeed carry the desired promoter/transgene units. Thus, the use of this and other similar vectors should dramatically improve current transgenesis protocols and reduce the time and cost for identifying transgenic animals. PMID- 12112581 TI - Bovine embryos contain a higher proportion of polyploid cells in the trophectoderm than in the embryonic disc. AB - The frequency of polyploid cells in the embryonic disc (ED) and in the trophectoderm (TE) was assessed in 50 in vitro produced bovine embryos fixed at days 7-8 post insemination (pi) and in 20 in vitro produced embryos that were transferred to uteri of recipients at day 7 and then recovered and fixed at day 12 pi. Separation of TE and ED cells was obtained by microdissection and the frequency of polyploid cells was determined by interphase cytogenetic analysis using fluorescence in situ hybridization (FISH) with chromosome 6- and chromosome 7-specific probes. The results show that 96% of day 7 embryos contain polyploid cells in the TE, whereas only 58% contain polyploid cells in the ED. In day 12 embryos 85% of TE and 40% of ED preparations contain polyploid cells. Statistical analysis revealed that the frequency of polyploid cells was significantly higher in the TE than in the ED in embryos containing less than 25% polyploid cells (n = 65). The few embryos (n = 5), which contained more than 25% polyploid cells, did not show this difference. Further, it was revealed that the level of polyploidy on day 7-8 was significantly higher than on day 12, both in the TE (two-fold) and in the ED (seven-fold). PMID- 12112580 TI - Transient transmission of a transgene in mouse offspring following in vivo transfection of male germ cells. AB - Sperm-mediated gene transfer in vertebrates has undergone various developments over the last few years, in different laboratories. In the present study, we microinjected a circular plasmid, carrying the lacZ reporter gene mixed with noncommercial cationic lipids, into the seminiferous tubules of anesthetized adult mice. Histochemical analysis was used to estimate the transfection efficiency 48-96 hr and 40 days after injection. As early as 48-96 hr post injection, an efficient transfection was revealed by a beta-galactosidase expression within both immature and differentiated germ cells. By 40 days post injection, the specific LacZ expression was restricted to the most immature germ cells in the basal portion of the seminiferous tubules. At this time, some injected males were mated with wild-type females and the progeny were analyzed by PCR and Southern blot. We showed that the transgene was transmitted to the offspring but remained episomal, as it was found in the tail of the young animals but not at adulthood. Therefore, the plasmid seemed to be lost during the numerous germ cells divisions. This plasmid stayed in some tissues, such as skeletal muscle and cardiac muscle. No integrative forms have yet been found with the use of a circular DNA. PMID- 12112582 TI - Regulation of apoptosis in the bovine blastocyst by insulin and the insulin-like growth factor (IGF) superfamily. AB - Insulin and the insulin-like growth factors, IGF-I and IGF-II, have been reported to exert a mitogenic effect on the preimplantation mammalian embryo. Furthermore, it has been proposed that loss of imprinting of the insulin-like growth factor II receptor gene and the consequent over-production of IGF-II may be involved in the aetiology of the Enlarged Offspring Syndrome, which occurs as an artefact of in vitro embryo production. We have previously shown that apoptosis occurs in the preimplantation bovine embryo and is influenced by in vitro culture conditions. We have therefore sought to establish the effects of insulin, IGF-I and IGF-II on apoptosis and cell proliferation in bovine blastocysts in vitro. Zygotes, obtained by in vitro maturation and fertilization of follicular oocytes, were cultured to blastocysts, with or without exogenous growth factors. Embryos were stained with propidium iodide to label all nuclei and by TUNEL to label apoptotic nuclei and analyzed by epifluorescent and confocal microscopy. IGF-I and IGF-II, but not insulin, were found to increase the proportion of embryos which formed blastocysts. Insulin decreased the incidence of apoptosis without affecting blastocyst cell number. IGF-I acted to decrease apoptosis and increase total cell number and IGF-II increased cell number alone. These data suggest roles for insulin and the IGFs as mitogens and/or apoptotic survival factors during early bovine development. Perturbation of IGF-II regulated growth may be involved in fetal oversize. PMID- 12112583 TI - Two kinase activities are sufficient for sea urchin sperm chromatin decondensation in vitro. AB - Decondensation of compact and inactive sperm chromatin by egg cytoplasm at fertilization is necessary to convert the male germ cell chromatin to an active somatic form. We studied decondensation of sea urchin sperm nuclei in a cell-free extract of sea urchin eggs to define conditions promoting decondensation. We find that egg cytosol specifically phosphorylates two sperm-specific (Sp) histones in vitro in the same regions as in vivo. This activity is blocked by olomoucine, an inhibitor of cdc2-like kinases, but not by chelerythrine, an inhibitor of protein kinase C (PKC). PKC phosphorylates and solubilizes the sperm nuclear lamina, one requirement for decondensation. Olomoucine, which does not inhibit lamina removal, blocks sperm nuclear decondensation in the same concentration range over which it is effective in blocking Sp histone phosphorylation. In a system free of other soluble proteins, neither PKC nor cdc2 alone elicit sperm chromatin decondensation, but the two act synergistically to decondense sperm nuclei. We conclude that two kinases activities are sufficient for sea urchin male pronuclear decondensation in vitro, a lamin kinase (PKC) and a cdc2-like Sp histone kinase. PMID- 12112584 TI - In vitro and in vivo maturation of oocytes from gonadotrophin-treated brushtail possums. AB - The time course of nuclear maturation of oocytes was examined in brushtail possums, Trichosurus vulpecula. Oocytes were recovered from ovarian follicles > 2 mm in diameter after pregnant mares' serum gonadotrophin/porcine luteinizing hormone (PMSG/LH) treatment (in vivo matured) or 72 hr after PMSG treatment (in vitro matured). Oocytes recovered from small (< 2 mm) and large (> 2 mm) follicles were also assessed for their ability to mature in vitro. Staining with the DNA-specific dye Hoechst 33342 was used to assess the stage of nuclear development by fluorescence microscopy. The process of nuclear maturation progressed rapidly in vivo, as oocytes collected at 20-27 hr post-LH all had a GV, but by 28-29.5 hr post-LH approximately a third of eggs were MII. By 30-hr post-LH, more than 70% of oocytes had reached MII stage and all ovulated eggs were MII. In vitro, all oocytes were at germinal vesicle stage at the start of culture. After 24 hr of culture, 67% of oocytes had progressed to metaphase I/anaphase I of meiosis. After 36 hr, 25% of oocytes had completed maturation to metaphase II, increasing to 52% after 48 hr. Maturation of oocytes after 48 hr in culture was unaffected by the presence or absence of granulosa cells, PMSG or LH/porcine follicle stimulating hormone (FSH). More oocytes from large follicles (55%) completed maturation by 48 hr than from small follicles (15%). The potential of oocytes to mature after 48 hr in culture was dependent on the follicle harvested having reaching a critical diameter of 1.5 mm. PMID- 12112585 TI - Embryonic and foetal development of bovine oocytes treated with a combination of butyrolactone I and roscovitine in an enriched medium prior to IVM and IVF. AB - Cattle oocytes were maintained at germinal vesicles (GV) stage for 24 hr using a combination of two specific and potent inhibitors of M-phase promoting factor (MPF) kinase activity, butyrolactone I (BL-I) and roscovitine (ROS). The media used for inhibition were (a) TCM-199 only and (b) TCM-199 supplemented with serum, hormones and growth factors. The effective doses of inhibitors were 6.25 microM BL-I and 12.5 microM ROS in medium (a) and 50 microM BL-I and 12.5 microM ROS in medium (b). After inhibition, about 90% of the oocytes resumed meiosis and reached the metaphase II (MII) stage during 24 hr of maturation. Following fertilisation the percentage of cleavage (D +2), compacted morula (D +6), blastocysts on D +7 and D +8 and the survival to freezing and thawing of grade 1 embryos frozen on D +7 were not different between the experimental treated groups and the control. In order to evaluate early foetal development, two groups of five grade 1 D +7 blastocysts derived from treated oocytes and two groups of five control embryos were transferred nonsurgically in four synchronised recipient heifers. On D +27, the recipients were slaughtered and the foetuses were recovered. In both groups, six foetuses developed out of the 10 embryos transferred. In conclusion, several supplements can be added to the prematuration medium of bovine oocytes without reducing the quality of inhibition but also without improving their subsequent developmental competence versus treated oocytes in TCM-199 only and versus untreated control. Furthermore, the prematuration step used in this study does not interfere with normal foetal development during the first stages of organogenesis. PMID- 12112586 TI - Functional analysis of sperm from c-mos(-/-) mice. AB - The c-mos protooncogene, which is expressed predominantly in male and female germ cells, is crucial for normal oocyte meiosis and female fertility in mice. Inactivation of c-mos results in abnormal oocyte development and leads to ovarian cysts and tumors in vivo. In contrast to the severe effects of c-mos ablation in females, targeted inactivation of c-mos has not been reported to affect spermatogenesis in male mice. However, previously reported studies of male c-mos( /-) mice have been limited to histological analyses of testes and in vivo matings, both of which are relatively insensitive indicators of sperm production and function. Therefore, we assayed sperm function of c-mos(-/-) males under in vitro conditions to determine whether the absence of Mos during development affected sperm production or fertilizing ability. We found no significant differences between the number of sperm collected from c-mos(-/-) and wild type mice. Additionally, sperm from c-mos(-/-) and c-mos(+/+) males performed equally well in assays of in vitro fertilization (IVF) and fertilization-associated events including zona pellucida (ZP) penetration, sperm/egg plasma membrane fusion, and sperm chromatin remodeling. Therefore, we suggest that the function of Mos in spermatogenesis is either not related to the ultimate fertilizing potential of the sperm, or else the absence of Mos is masked by a redundant kinase. PMID- 12112587 TI - Antibodies generated in response to plasmid DNA encoding zona pellucida glycoprotein-B inhibit in vitro human sperm-egg binding. AB - To investigate the immunogenicity of plasmid DNA encoding bonnet monkey (Macaca radiata) zona pellucida (ZP) glycoprotein-B (bmZPB), the cDNA corresponding to bmZPB, excluding the N-terminal signal sequence and C-terminus transmembrane-like domain, was cloned in mammalian expression vector VR1020 downstream of tissue plasminogen activator signal sequence under cytomegalovirus promoter (VRbmZPB). In vitro transfection of COS-1, COS-7, CHO, HEK-293, and UM-449 mammalian cells with VRbmZPB plasmid DNA led to the expression of bmZPB. Expression of bmZPB in transfected cells was cytosolic. Flow cytometry analysis of COS-1 cells transfected with VRbmZPB revealed that approximately 15% cells expressed bmZPB. The expressed bmZPB has an apparent molecular weight of 57 kDa. Immunization of male BALB/cJ mice with VRbmZPB plasmid DNA in saline as compared to VR1020 immunized group, elicited significant antibodies against E. coli expressed recombinant bmZPB as evaluated in ELISA. The antibodies generated by VRbmZPB plasmid DNA recognized bonnet monkey as well as human ZP. The immune sera obtained from mice immunized with VRbmZPB plasmid DNA also inhibited, in vitro, the binding of spermatozoa to the ZP in the hemizona assay. These studies, for the first time, demonstrate the feasibility of DNA vaccine to generate antibodies against ZP that recognize native protein and inhibit human sperm-oocyte binding. PMID- 12112588 TI - Characterization of human membrane cofactor protein (MCP; CD46) on spermatozoa. AB - Membrane cofactor protein (MCP; CD46) is a complement regulator widely expressed as four isoforms that arise via alternative splicing. On human spermatozoa, MCP is expressed on the inner acrosomal membrane and alterations of spermatozoa MCP may be associated with infertility. In rodents, expression of MCP is largely restricted to the testes. MCP on human spermatozoa has a unique M(r) pattern that we have investigated. We also characterized MCP expression in mice transgenic (tg) for human MCP. Human MCP expression in the tg mice mimics the human pattern in that it is located on the inner acrosomal membrane and has a faster M(r) than MCP expressed elsewhere. Sequencing of RT-PCR products from the testis indicates that there is not a unique male reproductive tissue specific cytoplasmic tail. Instead, human spermatozoa express MCP bearing cytoplasmic tail two, which is also utilized in most other tissues and contains several signaling motifs. Further, using N-glycosidases, we demonstrate that the unique lower molecular weight of MCP on spermatozoa is secondary to a modification in the N-linked sugars. Specifically, as the spermatozoa mature, but before they reach the epididymis, the three N-linked sugars of MCP are trimmed to less complex structures. While the purpose of this deglycosylation is unknown, we propose that it is a common feature of proteins expressed on the plasma and inner acrosomal membranes of spermatozoa and hypothesize that it is a spermatozoa specific event critical for facilitating sperm-egg interactions. PMID- 12112590 TI - Identification of the cadherin subtypes present in the human peritoneum and endometriotic lesions: potential role for P-cadherin in the development of endometriosis. AB - Endometriosis is defined as endometrial tissue outside of the uterine cavity. The pathogenesis of this common disease remains poorly understood. However, the implantation and invasion of the viable cells from retrograde menstruation into the peritoneum is a widely accepted theory. To date, the mechanisms by which cell adhesion molecules mediate the development of human endometriosis remain unclear. Cadherins are a family of cell adhesion molecules that mediate cell-cell adhesion in a homophilic manner. In this study, the cadherins present in the peritoneum and endometriotic lesions were identified by RT-PCR using degenerate primers. In addition, differences in the levels of the cadherin mRNA transcripts present in eutopic endometrium and endometriotic lesions of the same patients were then compared by semiquantitative RT-PCR. Multiple cadherins were detected in the peritoneum and endometriotic lesions. Of these, P-cadherin appears to be the predominant cadherin subtype present in the peritoneum. Similarly, P-cadherin mRNA levels in endometriotic lesions were significantly greater than those observed in the corresponding eutopic endometrium. The expression of P-cadherin in both the human peritoneum and endometriotic lesions suggests that this cell adhesion molecule may play a central role in the development of endometriosis by mediating endometrial-peritoneal cell interactions in a homophilic manner. PMID- 12112591 TI - Rescue of infertile transgenic rat lines by intracytoplasmic injection of cryopreserved round spermatids. AB - Transgenic male rats carrying human alpha-lactalbumin with thymidine kinase gene (line name; LAC3) were found to be infertile due to expression of the transgene in the testes. Furthermore, it was not possible to maintain the line even by the use of intracytoplasmic sperm injection (ICSI). Therefore, round spermatids prepared from the LAC3 rats were microinjected into strontium-activated oocytes using a Piezo-driven micromanipulator. Of 263 oocytes microinjected with LAC3 spermatids, 244 (92.8%) survived the injection and 96 (39.3%) developed to the 2 cell stage. Three viable offspring were born after transfer (1.4%, 3/219), and two offspring carried the LAC3 transgene. In the control experiment using spermatids of Wistar rats, similar proportions of post-injection survival (91.3%, 241/264), cleavage (40.2%, 97/241), and development into offspring (0.5%, 1/206) were obtained. Thus, this paper reports not only the first rat offspring derived from round spermatid injection but also the practical application of the microinsemination technique to the rescue of transgenes of infertile transgenic male rats. PMID- 12112589 TI - Cadherins expression during gamete maturation and fertilization in the rat. AB - A role for adhesion molecules in gamete fusion, preceding fertilization, has been previously suggested. We investigated the presence of cadherins, Ca(2+) dependent cell-cell adhesion molecules, in rat oocytes and spermatozoa using an anti-pan cadherin antibody and specific antibodies against the 3 classical cadherins: E- (epithelial), P- (placental), and N- (neural) cadherins. Electrophoretic separation was performed on samples of lysed oocytes of different stages: germinal vesicle oocytes, metaphase II eggs, newly fertilized and 2-cell embryos, as well as spermatozoa from testes, caput and cauda epididymis and ejaculate. Localization of cadherins was determined on intact, gametes by immunocytochemistry, using confocal microscopy. Immunoblotting with the pan cadherin antibody revealed a major band of approximately 120 kD in all oocyte and sperm extracts. Oocytes presented E-cadherin at appropriate molecular weight but N-cadherin only as a specific 40 kD band. In sperm lysate, at all stages, both E- and N-cadherin were demonstrated as major protein bands but a series of lower molecular weight proteins (that may represent protein degradation) were also detected. Immunohistochemical evaluation showed that E- and N-cadherins are already present on the plasma membrane of immature unfertilized oocytes, although their concentration increases after fertilization in early cleavage stage embryos. Cadherin localization on spermatozoa changed during maturation from a dispersed pattern over the entire head plasma membrane of testicular spermatozoa to a restricted equatorial and post-acrosomal plasma membrane staining in ejaculated spermatozoa. These findings suggest a specific cadherin organization at the fusogenic domains of both gametes. PMID- 12112593 TI - Effects of cryopreservation of mouse embryos and in vitro fertilization on genotypic frequencies in colonies. AB - To evaluate the effects of cryopreservation and in vitro fertilization (IVF) on genotypic frequencies in mouse colonies, genotypic frequencies at 15 biochemical, 4 immunological and 20 microsatellite loci were examined in three colonies of MCH (ICR) mice derived from noncryopreserved embryos obtained by natural mating without the induction of superovulation, cryopreserved embryos obtained by natural mating with the induction of superovulation, and cryopreserved embryos obtained by the induction of superovulation and IVF. Three (Pgm-1, Ldr-1 and Hbb) out of the 15 biochemical loci, two (Thy-1 and H2K) out of four immunological loci and five (D5Mit18, D6Mit15, D12Mit5, D13Mit26, and D14Mit7) out of 20 microsatellite loci that showed polymorphisms in every colony were used for detection of genotypic frequencies. The genotypic frequencies of the loci in the three colonies did not differ from the predicted genotypic frequencies (P > 0.05). The results suggested that genetic drift does not occur among colonies established from treated and untreated embryos, and it was clear that the embryo banking by cryopreservation is suitable for preservation of outbred stock without genetic drift. PMID- 12112592 TI - Transgenic pig expressing the enhanced green fluorescent protein produced by nuclear transfer using colchicine-treated fibroblasts as donor cells. AB - Fetal-derived fibroblast cells were transduced with replication defective vectors containing the enhanced green fluorescent protein (EGFP). The transgenic cells were treated with colchicine, which theoretically would synchronize the cells into G2/M stage, and then used as donor nuclei for nuclear transfer. The donor cells were transferred into the perivitalline space of enucleated in vitro matured porcine oocytes, and fused and activated with electrical pulses. A total of 8.3% and 28.6% of reconstructed oocytes showed nuclear envelope breakdown and premature chromosome condensation 0.5 and 2 hr after activation, respectively. Percentage of pronuclear formation was 62.5, 12 hr after activation. Most (91.4%) of the 1-cell embryos with pronuclei did not extrude a polar body. Most (77.2%) embryos on day 5 were diploid. Within 2 hr after fusion, strong fluorescence was detectable in most reconstructed oocytes (92.3%). The fluorescence in all NT embryos became weak 15 hr after fusion and disappeared when culture to 48 hr. But from day 3, cleaved embryos at the 2- to 4-cell stage started to express EGFP again. On day 7, 85.8% of cleaved embryos expressed EGFP. A total of 9.4% of reconstructed embryos developed to blastocyst stage and 71.5% of the blastoctysts expressed EGFP. After 200 reconstructed 1-cell stage embryos were transferred into four surrogate gilts, three recipients were found to be pregnant. One of them maintained to term and delivered a healthy transgenic piglet expressing EGFP. Our data suggest that the combination of transduction of somatic cells by a replication defective vector with the nuclear transfer of colchicine-treated donors is an alternative to produce transgenic pigs. Furthermore, the tissues expressing EGFP from descendents of this pig may be very useful in future studies using pigs that require genetically marked cells. PMID- 12112595 TI - Developmental, qualitative, and ultrastructural differences between ovine and bovine embryos produced in vivo or in vitro. AB - The objective of this study was to compare bovine and ovine oocytes in terms of (1) developmental rates following maturation, fertilization, and culture in vitro, (2) the quality of blastocysts produced in vitro, assessed in terms of their ability to undergo cryopreservation, and (3) the ultrastructural morphology of these blastocysts. In vitro blastocysts were produced following oocyte maturation/fertilization and culture of presumptive zygotes in synthetic oviduct fluid. In vivo blastocysts were used as a control from both species. In Experiment 1, the cleavage rate of bovine oocytes was significantly higher than that of ovine oocytes (78.3% vs. 58.0%, respectively, P < 0.001). The overall blastocyst yield was similar for both species (28.7% vs. 29.0%). However, when corrected for cleavage rate, significantly more ovine oocytes reached the blastocyst stage at all time-points (36.6% vs. 50.0% on day 8, for bovine and ovine, respectively, P < 0.001). Following vitrification, there was no difference in survival between in vivo produced bovine and ovine blastocysts (72 hr: 85.7% vs. 75.0%). However, IVP ovine blastocysts survived at significantly higher rates than IVP bovine blastocysts at all time points (72 hr: 47.4% vs. 18.1%, P < 0.001). At the ultrastructural level, compared with their in vivo counterparts, IVP blastocysts were characterized by a lack of desmosomal junctions, a reduction in the microvilli population, an increase in the average number of lipid droplets and increased debris in the perivitelline space and intercellular cavities. These differences were more marked in bovine IVP blastocysts, which also displayed electron-lucent mitochondria and large intercellular cavities. These observations may in part explain the species differences observed in terms of cryotolerance. In conclusion, the quality of ovine blastocysts was significantly higher than their bovine counterparts produced under identical in vitro conditions suggesting inherent species differences between these two groups affecting embryo quality. PMID- 12112594 TI - Stage of the estrous cycle at the time of pregnant mare's serum gonadotropin injection affects pre-implantation embryo development in vitro in the mouse. AB - The present study aims to analyze in the mouse the effect of the stage of the estrous cycle at the time of pregnant mare's serum gonadotropin (PMSG) injection on fertilization of ovulated cumulus-enclosed oocytes and later embryo development in vitro to the blastocyst stage. Quality of blastocysts was evaluated by staining and counting of total number of nuclei, mitotic index, percentage of apoptotic nuclei, and cell allocation to the inner cell mass (ICM) and trophectoderm (TE) lineage. Superovulation of hybrid (C57Bl/6JIco female x CBA/JIco male) female mice of 4-6 weeks of age was induced by a priming injection of PMSG at different stages of the estrous cycle followed after a 48-hr interval by human chrorionic gonadotropin. Our data indicate that injection of PMSG at the estrus phase gives the best outcome whereas injection of PMSG at the diestrus-1 or diestrus-2 phase provides the worst results. In fact, (1) total number of oocytes ovulated, number of ovulated oocytes enclosed by cumulus cells, and number of TE cells in day-5 blastocysts were significantly lower in diestrus-1 females than in estrus, diestrus-2 and proestrus mice; (2) percentage of day-5 blastocysts and total number of cells in day-5 blastocysts were lower in diestrus 1 and diestrus-2 females than in estrus and proestrus mice; and (3) percentage of apoptotic nuclei in day-5 blastocysts was lower in estrus mice than in diestrus 1, diestrus-2, or proestrus females. These data endorse previous studies suggesting that administration of gonadotropins in mice should be synchronized with the innate estrous cycle of females. PMID- 12112596 TI - Embryonic hatching enzyme strypsin/ISP1 is expressed with ISP2 in endometrial glands during implantation. AB - Embryo hatching and outgrowth are the first critical steps on the way to a successful pregnancy. It is generally held that serine proteases are responsible for this process, although the exact mechanisms of action are not clearly understood. Recently, we described two novel implantation serine proteinase (ISP) genes that are expressed during the implantation period. The ISP1 gene encodes the embryo-derived enzyme strypsin, which is necessary for blastocyst hatching in vitro and the initiation of invasion. The ISP2 gene, which encodes a related tryptase, is expressed in endometrial glands and is regulated by progesterone during the peri-implantation period. Based on similarities between ISP2 gene expression and that of a progesterone-regulated lumenal serine proteinase activity associated with lysis of the zona pellucida, we have suggested that the strypsin related protein, ISP2, may encode a zona lysin proteinase. As tryptases naturally assemble to form tetrameric structures, we have hypothesized that ISP1 and ISP2 tetramerize to form strypsin and lysin, respectively. In this study, we demonstrate that like ISP2, the ISP1 gene is also expressed in endometrial glands and is positively regulated by progesterone during implantation. Using in situ hybridization of adjacent tissue sections, we show that the ISP1 and ISP2 genes are co-expressed within the endometrial gland. Following evidence that ISP1 and 2 can efficiently form homotetramers and heterotetramers in silico, we suggest that ISP heterotetramers may be also be secreted into the uterine lumen during the implantation period. That the embryonic hatching enzyme, may also be secreted into the uterine lumen from uterus, may provide insight into the mechanisms of hatching and implantation initiation. PMID- 12112597 TI - Comparison of glycoprotein hormone alpha-subunits of laboratory animals. AB - The common alpha-subunit of glycoprotein hormones (CGalpha) is a core protein shared by follicle-stimulating hormone (FSH), luteinizing hormone (LH), and thyroid-stimulating hormone (TSH). In order to obtain a molecular basis for an efficient superovulation technique applicable to a wide range of animal species and to discuss the phylogenetic aspect based on molecules related to the reproductive system, we determined cDNA sequences of CGalpha in seven laboratory animals: the guinea pig, Mongolian gerbil, golden hamster, mastomys, Japanese field vole, the JF1 strain of Mus musculus molossinus, and rabbit. Comparison of the inferred CGalpha amino acid sequences of these animals and other mammals (human, mouse, rat, cow, pig, and sheep) showed that the signal peptides and the first ten residues at the N-terminus of the apoprotein were variable, while the rest of the apoproteins were highly conserved. In particular, all rodents had a leucine residue at the apoprotein N-terminus, except the guinea pig, which had a phenylalanine residue, as in the cow, pig, sheep, and rabbit. Phylogenetic trees constructed from amino acid sequences suggest a closer relationship between the guinea pig and artiodactyls than to rodents, confirming the taxonomic peculiarity of the guinea pig. PMID- 12112598 TI - Altered levels of mRNA expression and pharmacological reactivity of alpha1 adrenergic receptor subtypes in the late-pregnant rat myometrium. AB - The adrenergic system plays a major role in the regulation of the uterine contractility during pregnancy. Our previous studies have shown the significance of the alpha1-adrenergic receptors (ARs) in the control of pregnant uterine contractility both in vitro and in vivo. Our present aim was to determine the changes in mRNA expression and pharmacological reactivity of the alpha1-ARs on days 18, 20, and 22 of pregnancy. To demonstrate the expressions of alpha1-AR subtype mRNA, we used a reverse transcription-polymerase chain reaction (RT-PCR); the pharmacological reactivity was tested by electric field stimulation (EFS). The expression of alpha1A-AR mRNA increased from day 18 to 22, while no alpha1B AR mRNA was detectable. We found a small increase in the expression of alpha1D-AR mRNA on day 20, which was not followed by a significant change in pharmacological reactivity. The alpha1D-receptor expression and pharmacological reactivity decreased significantly up to day 22. EFS studies revealed that the alpha1A-AR antagonist 5-methylurapidil had EC50 values (1.9 x 10(-6)-6.3 x 10(-6) M) about one order of magnitude lower than those of the alpha1D-AR antagonist BMY 7378 (4 x 10(-6)-3.6 x 10(-5) M). However, the alpha1B-AR antagonist cyclazosine exerted only a slight effect on the stimulated contractions. Strong correlations were found between the alpha1A-mRNA expression and the EC50 of 5-methylurapidil (r(2) =0.9712), and between the alpha1D-AR mRNA expression and the EC50 of BMY 7378 (r(2) = 0.9937). Our findings suggest that both alpha1A- and alpha1D-ARs are involved in the regulation of the pregnant uterine contractility. The density and pharmacological reactivity indicate that the alpha(1A)-AR seems to play the major role in late-pregnant myometrial contraction. PMID- 12112599 TI - Roles of the Na,K-ATPase alpha4 isoform and the Na+/H+ exchanger in sperm motility. AB - The Na,K-ATPase generates electrochemical gradients that are used to drive the coupled transport of many ions and nutrients across the plasma membrane. The functional enzyme is comprised of an alpha and beta subunit and families of isoforms for both subunits exist. Recent studies in this laboratory have identified a biological role for the Na,K-ATPase alpha4 isoform in sperm motility. Here we further investigate the role of the Na,K-ATPase carrying the alpha4 isoform, showing again that ouabain eliminates sperm motility, and in addition, that nigericin, a H+/K+ ionophore, and monensin, a H+/Na+ ionophore, reinitiate motility. These data, along with the observation that the K+ ionophore valinomycin has no effect on the motility of ouabain-inhibited sperm, suggest that ouabain may change intracellular H+ levels in a manner that is incompatible with sperm motility. We have also localized NHE1 and NHE5, known regulators of intracellular H+ content, to the same region of the sperm as the Na,K-ATPase alpha4 isoform. These data highlight the important role of the Na,K-ATPase alpha4 isoform in regulating intracellular H(+) levels, and provide evidence suggesting the involvement of the Na+/H+ exchanger, which is critical for maintaining normal sperm motility. PMID- 12112600 TI - Evolution of octopod sperm I: comparison of nuclear morphogenesis in Eledone and Octopus. AB - Morphogenesis of the Eledone cirrhosa sperm nucleus, as studied by electron microscopic techniques, is compared with that of Octopus vulgaris. Both species of cephalopods belong to the family Octopodidae. The results indicate that extensive nuclear helicoidization during E. cirrhosa spermiogenesis is brought about by modifications of the function of structural components already present in the late steps of O. vulgaris spermiogenesis. In particular, changes in the regulation of perinuclear microtubule contraction in E. cirrhosa spermatids, as well as a decrease in basicity of protamines, promote nuclear helicoidization. Disulphide bond formation between protamine molecules fixes the completely helicoidal shape of the nucleus in mature sperm of E. cirrhosa. PMID- 12112602 TI - DNA spatial distribution in unfertilized human oocytes by confocal laser scanning microscope. AB - The aim of this study is to investigate the DNA spatial distribution in unfertilized human oocytes by confocal laser scanning microscope (CLSM). One hundred and forty unfertilized oocytes after conventional in vitro fertilization (IVF) and one hundred and three after intracytoplasmic sperm injection (ICSI) were studied. The chromatin was stained with chromomycin A3 (CA3) initially and at a second step with propidium iodide, allowing the observation of the deprotaminated sperm heads in the ooplasm. CLSM offered the opportunity for studying the three-dimensional position of the spermatozoon in the oocyte and the description of the status of the maternal and paternal chromatin. For each of the IVF and ICSI group of unfertilized oocytes, subgroups were performed according to the status of the maternal and paternal chromatin. The maternal chromosomes were either aligned at metaphase II or disarrayed at metaphase II, and the sperm head was either condensed or swollen/decondensed. Few oocytes were activated and formed a pronucleus. The spatial approaching between paternal and maternal chromatin was calculated for all subgroups of unfertilized oocytes and the distances were compared among them. There was no statistical difference in the distance between the spermatozoon and the maternal chromatin among subgroups of the IVF group, while statistical differences occurred among subgroups of the ICSI group. The spermatozoa in the IVF group approached maternal chromatin more than those in the ICSI group. Movement of the spermatozoon into the oocyte and chromatin remodeling for the formation of pronucleus seem to occur independently and are possibly directed by different mechanisms. PMID- 12112601 TI - Evolution of octopod sperm II: comparison of acrosomal morphogenesis in Eledone and Octopus. AB - The first stages of acrosome development during Eledone cirrhosa spermiogenesis are similar to that in Octopus vulgaris, and comprise the initial elongation of both organelles. However, the acrosome in E. cirrhosa does not continue its elongation as it does in O. vulgaris. Instead, its length remains fixed and it undergoes a process of helicoidization that includes the entire organelle. In each spermatid, helicoidization of the E. cirrhosa acrosome occurs simultaneously with helicoidization of the nucleus. The acrosome is associated with special structures that probably are involved in the helical torsion of the organelle. We propose a hypothesis to explain the evolutionary relationship between the acrosomes of O. vulgaris and E. cirrhosa, particularly as it is influenced by nucleomorphogenesis and microtubular contraction. PMID- 12112603 TI - Maintenance of meiotic arrest in bovine oocytes in vitro using butyrolactone I: effects on oocyte ultrastructure and nucleolus function. AB - In this study, we have shown that butyrolactone I (BL-I), a potent inhibitor of cyclin-dependent kinases, affects oocyte cytoplasmic morphology and nuclear function in terms of nucleolar ultrastructure and immunocytochemistry. Bovine oocytes were recovered from three classes of follicle size: 1.5-2, 2-3, and 3-6 mm. The oocytes were incubated for 40 hr with BL-I, and subsequently processed for transmission electron microscopy or immunocytochemistry. A control group of oocytes were processed immediately upon recovery (0 hr). In general, incubation with BL-I for 40 hr disrupted contact between cells of the cumulus oophorous and the oocyte, caused degeneration of the cortical granules and the peripheral migration of all cytoplasmic organelles. At the level of the nucleus, it induced convolution of the nuclear membrane and caused acceleration of nucleolar compaction in oocytes from follicles < 3 mm and fragmentation of nucleoli, particularly evidenced by immunocytochemistry, in oocytes from follicles > 3 mm. Furthermore, the effects appear to be more profound in fully-grown oocytes. PMID- 12112604 TI - Effect of 6-dimethylaminopurine on electrically activated in vitro matured porcine oocytes. AB - The effect of the protein kinase inhibitor, 6-dimethylaminopurine (6-DMAP), on the maturation promoting factor (MPF) activity, pronuclear formation, and parthenogenetic development of electrically activated in vitro matured (IVM) porcine oocytes was investigated. Oocytes were activated by exposure to two DC pulses, each of 1.5 kV/cm field strength and 60 microsec duration, applied 1 sec apart. In the first experiment, subsequent incubation with 2 or 5 mM 6-DMAP for 3 hr increased the incidence of blastocyst formation compared with no treatment, whereas incubation with 2 or 5 mM 6-DMAP for 5 hr did not. In the proceeding experiments, oocytes exposed to 6-DMAP were incubated with 2 mM of the reagent for 3 hr. Assaying histone H1 kinase activity in the second experiment revealed that the levels of active MPF in electrically activated oocytes treated with 6 DMAP were depleted more rapidly and remained depleted for longer compared with electrical activation alone. The kinetics of MPF activity following 6-DMAP treatment were similar to that found in inseminated oocytes in the third experiment. The effect of 6-DMAP was correlated with an increased incidence of parthenogenetic blastocyst formation. A fourth experiment was undertaken to examine the diploidizing effect of 6-DMAP. Electrically activated oocytes treated with 6-DMAP and cytochalasin B, either alone or in combination, displayed a higher incidence of second polar body retention compared with those that were untreated or treated with cycloheximide alone. After 6 days of culture in vitro, parthenotes exposed to 6-DMAP, either alone or in combination with cytochalasin B, formed blastocysts at a greater rate compared with those exposed to cytochalasin B alone, cycloheximide alone or no treatment. The combined 6-DMAP and cytochalasin B treatment induced the highest rate of blastocyst formation (47%), but the numbers of trophectoderm and total cells in these blastocysts were lower compared with those obtained following exposure to 6-DMAP alone. These results suggest that the increased developmental potential of 6-DMAP-treated parthenotes may be attributable to the MPF-inactivating effect of 6-DMAP, rather than the diploidizing effect of 6-DMAP. PMID- 12112605 TI - Fibroin-like substance is a major component of the outer layer of fertilization envelope via which carp egg adheres to the substratum. AB - Seven cDNA encoding silkworm fibroin homologues were cloned from a carp ovarian cDNA library. The encoded proteins are denoted as carp ovarian fibroin-like substances (FLS). FLS contain a repetitive domain consisting of tandem repeats of dipeptide of Gly-X, where X may be any amino acid. Each FLS has its own unique repeating sequence, such as GQGAGQGS, GQGMGQGM, GRGQGEGHGS, and GFGFGQGS, indicating a family of FLS genes exists in carp. FLS is exclusively expressed in oocytes and is stored in cortical granules. During cortical reaction, FLS is exocytosed to perivitelline space and then gradually added to the outer layer of the fertilization envelope (FEo). The FLS of fertilization envelope is conjugated with cystatin and cathepsin-like substance (CLS) and appears in multiple bands of molecular weights ranging from 40 to 205 kDa. After fertilization or artificial activation, carp eggs adhere firmly to the substratum via FEo. FLS is a major component of FEo. The presence of transglutaminase inhibitor, cadaverine or ethylene diaminetetraacetic acid, in the cortical reaction medium can impair or block the recruitment of FLS and other substances to FEo. As a consequence, FEo is not formed or is greatly reduced, resulting in a great reduction of egg adhesion. PMID- 12112606 TI - Nuclear maturation of canine oocytes cultured in protein-free media. AB - The objective of this study was to determine the ability of canine oocytes to complete nuclear maturation in a protein-free medium. Oocytes obtained from ovaries of bitches aged 6 months to 2 years were cultured either in TCM199 or CMRL1066 medium without protein supplementation in 5% or 20% O(2). Sixteen of 121 (13%) oocytes cultured in TCM199 reached metaphase II, but only 1 of 135 oocytes cultured in CMRL1066 did so (P < 0.05). Oxygen concentration did not affect nuclear maturation. An additional 103 oocytes were cultured in TCM199 for 48 hr, inseminated with chilled ejaculated spermatozoa, fixed in 1:3 acetic acid-ethanol and then stained with aceto-orcein; 34% of these oocytes were penetrated by spermatozoa. To determine developmental competence of oocytes cultured in a protein-free medium, 85 oocytes were cultured in TCM 199 for 48 hr, inseminated and then cultured; 7 early stage embryos were produced. The effects of growth hormone, beta-mercaptoethanol (betaME), luteinizing hormone (LH) and energy substrates, alone or in combination, on nuclear maturation of oocytes cultured in a protein-free medium were also determined. Growth hormone enhanced cumulus expansion, but did not improve nuclear maturation. beta-mercaptoethanol had no effect on nuclear maturation. However, percentages of MII oocytes significantly decreased when the oocytes were cultured for 48 hr in the medium containing LH or a high concentration of glucose (P < 0.05). In conclusion, canine oocytes are able to complete nuclear maturation in a protein-free medium. The specific type of medium and other supplements significantly influence the meiotic maturation of canine oocytes. PMID- 12112607 TI - Sperm nuclear halos can transform into normal chromosomes after injection into oocytes. AB - Mouse sperm nuclei extracted with an ionic detergent and 2 M NaCl retain their overall morphology, but upon subsequent reduction of the protamine disulfides they lose all elements of chromatin structure except the organization of DNA into loop that are anchored to the nuclear matrix. These DNA loops appear as a halo surrounding the nuclear matrix, and nuclei extracted in this manner are, therefore, called nuclear halos. Here, we report that sperm nuclear halos injected into oocytes can form pronuclei, then transform into chromosomes with normal morphology. This suggests that sperm nuclear halos retain all the information necessary for normal chromosomal organization, and that micromanipulation of these extracted sperm nuclei can be accomplished without major DNA damage. PMID- 12112608 TI - In-vivo non-invasive study of the thermoregulatory function of the blood vessels in the rat tail using magnetic resonance angiography. AB - In rats, a significant portion of total body heat loss occurs through sympathetically mediated changes in tail blood flow, making the rat tail a convenient model to study vasomotor activity during thermoregulation. Our aim was to perform a non-invasive study of the mechanisms of blood vessel control in the rat tail upon increasing body temperature. In anaesthetized rats, blood vessel temperature was monitored using non-invasive thermistors positioned on the skin surface, covering the ventral artery (Ta) and lateral vein (Tv), and changes in blood vessel size were measured using in-vivo magnetic resonance angiography (MRA). Two important regions of the tail (base and middle) were studied during a gradual rise of rectal temperature (Tr) from 37 to 40 degrees C. MRA data show that increasing Tr causes increased diameter of both arteries and veins of the tail, that venous diameter changes are greater than arterial diameter changes, and that diameter changes of both types of vessel are greater at the base of the tail than in the middle. Temperature data allowed calculation of (Ta - Tv), which we used as an index of flow through arteriovenous anastomoses (AVAs). The data suggest that AVAs near the base of the tail are important in heat exchange, and that they remain open only for Tr values between 38 and 39 degrees C. PMID- 12112609 TI - An additional phase in PCr use during sustained isometric exercise at 30% MVC in the tibialis anterior muscle. AB - The occurrence of an abrupt acceleration in phosphocreatine hydrolysis in the tibial anterior muscle during the last part of a sustained isometric exercise at 30% maximal voluntary contraction until fatigue is demonstrated in seven out of eight healthy subjects by applying in vivo 31P NMR spectroscopy at 1.5 T field strength. This additional third phase in PCr hydrolysis, is preceded by a common biphasic pattern (first fast then slow) in PCr use. The NMR spectra, as localized by a surface coil and improved by proton irradiation, were collected at a time resolution of 16 s. Mean rates of PCr hydrolysis during exercise were -0.44 +/- 0.19% s(-1), -0.07 +/- 0.04% s(-1), and -0.29 +/- 0.10% s(-1) for the three successive phases. The increased rate of PCr hydrolysis, and also the loss of fine force control evident in the force records are consistent with increased involvement of large, fast-fatiguable units later in the contraction. PMID- 12112610 TI - In-vivo visualization of phagocytotic cells in rat brains after transient ischemia by USPIO. AB - Cerebral ischemia provokes tissue damage by two major patho-physiological mechanisms. Direct cell necrosis is induced by diminished access of neurons and glia to essential nutrients such as glucose and oxygen leading to energy failure. A second factor of cellular loss is related to the activation of immune-competent cells within and around the primary infarct. While granulocytes and presumably monocytes are linked to the no-reflow phenomenon, activated microglia cells and monocytes can release cytotoxic substrates, which cause delayed cell death. As a consequence the infarct volume will increase, despite restoration of cerebral perfusion. In the past, visualization of immune competent cells was only possible by histological analysis of post-mortem tissue. However, contrast agents based on small particles of iron oxide are known to accumulate in organs rich in cells with phagocytotic function. These particles can be tracked in vivo by MRI methods based on their relaxation properties. In the present study, the spatio-temporal distribution of USPIO particles was monitored in a rat model of transient cerebral infarction using T1- and T2-weighted MRI sequences. USPIO were detected in vessels at 24 h after administration. At later time points specific accumulation of USPIO was observed within the infarcted hemisphere, with maximal signal enhancement on day 2. Their detectability based on T1-contrast disappeared between day 4 and day 7. Immuno-histochemically (IHC) stains confirmed the presence of macrophages, presumably blood-derived monocytes within areas of T1 signal enhancement. Direct visualization of iron-burdened macrophages by IHC was only possible later than day 3 after occlusion. PMID- 12112611 TI - Serial proton spectroscopy, magnetization transfer ratio and T 2 relaxation in pseudotumoral demyelinating lesions. AB - In some rare cases, demyelinating plaques appear on contrast-enhanced T1-weighted images as pseudotumoral, cyst-like lesions (hypointense, ring enhancing). Serial proton MR spectroscopy, T2 relaxometry and magnetization transfer ratios (MTR) were performed on three pseudotumoral demyelinating lesions to obtain information about their pathological basis. Baseline and 1-month MTR and T2 values were similar to those of cerebrospinal fluid, while spectra showed lactate, lipids and choline. Three-month and 1 year exams showed recovery of MTR, T2 and N acetylaspartate, approaching the contralateral values, while creatine and choline were normal or surpassed contralateral values. Lipids and lactate gradually disappeared. These results suggest that pseudotumoral, cyst-like, ring-enhancing lesions may be characterized by an accumulation of oedema in the extracellular space with an almost complete absence of cells. Reduction of the oedema allows restoration of the tissue to its original location, indicating that cellular destruction was less important than was expected after the first exam. Thus, the evolution of this kind of lesion should be kept in mind when considering lesion volume from T1-weighted images as a marker of disability or irreversible cellular destruction in MS. PMID- 12112612 TI - Sequential proton MRS study of brain metabolite changes monitored during a complete pathological cycle of demyelination and remyelination in a lysophosphatidyl choline (LPC)-induced experimental demyelinating lesion model. AB - Metabolite changes in rat brain internal capsule (ic) area were monitored using volume localized in vivo proton MR spectroscopy (MRS) in a lysophosphatidyl choline (LPC)-induced experimental demyelinating lesion model of multiple sclerosis (MS), during the early phase (pre-acute) as well as during the complete pathological cycle of de- and re-myelination processes. The N-acetyl aspartate (NAA) peak showed reduction during the early phase of the lesion progression (demyelination) until day 10 and increased thereafter during remyelination. However, choline (Cho) and lipid resonances showed increased signal intensity during the early phase and decreased during remyelination. A progressive reduction of the NAA/Cr metabolite ratio in lesioned rats was observed during demyelination (up to day 10) compared with before lesion (control), and the value increased thereafter during remyelination (from day 15). During this period, however, the Cho/Cr ratio was a higher until day 10 and subsequently declined and was close to that calculated before lesion creation. The changes in NAA/Cr and Cho/Cr metabolite ratios correspond to changes in the individual metabolite peaks such as NAA and Cho. The increase in the intensity of the choline resonance during the early phase is indicative of the onset of an inflammatory demyelination process, and its rapid decrease thereafter is due to reduction in the inflammatory process associated with remyelination. Similarly, the increase in the intensity of lipids during the pre-acute stage of the lesion is attributed to active demyelination, which significantly decreased during remyelination. These MR results correlate well with the histology data obtained. PMID- 12112613 TI - Current awareness in NMR in biomedicine. PMID- 12112614 TI - Backbone cleavages of [M - H](-) anions derived from caerin 1 peptides and some synthetic modifications. Molecular recognition initiating internal cyclisation of Glu23. AB - The collision induced spectra of [M - H](-) anions from of caerin 1 peptides and some synthetic modifications show the usual alpha, beta and beta' backbone cleavages together with Ser (epsilon,gamma) and Glu (gamma) cleavages which break the peptide backbone in the vicinity of those residues. All of these cleavages require the peptide backbone to be flexible. There is also a backbone cleavage of a type not observed before. This cleavage involves nucleophilic attack of the carboxylate anion of the Glu23 side chain at the backbone CH of Ile 21. We propose that this cleavage requires the caerin peptide to be in an alpha helical conformation (the 3D structure that this peptide adopts in solution) in order that the interacting groups are held in close proximity. PMID- 12112615 TI - Direct surface analysis of fungal species by matrix-assisted laser desorption/ionization mass spectrometry. AB - In this study various methods of sample preparation and matrices were investigated to determine optimum collection and analysis criteria for fungal analysis by matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS). Intact spores and/or hyphae of Aspergillus niger, Rhizopus oryzae, Trichoderma reesei and Phanerochaete chrysosporium were analyzed by matrix assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI TOFMS). The fungal samples were applied to the MALDI sample target as untreated, sonicated, or acid/heat treated samples, or blotted directly from the fungal culture with double-stick tape. Ferulic acid or sinapinic acid matrix solution was layered over the dried samples and analyzed by MALDI-MS. Statistical analysis showed that simply using double-stick tape to collect and transfer to a MALDI sample plate typically worked as well as the other preparation methods, and required the least sample handling. PMID- 12112616 TI - Analysis of estrogenic contaminants in river water using liquid chromatography coupled to ion trap based mass spectrometry. AB - A precise and reliable method, using liquid chromatography combined with ion trap based mass spectrometry, for the determination of three endogenous estrogens, namely, estrone, estradiol, and estriol, and two synthetic estrogens, ethinyl estradiol and diethylstilbestrol, in environmental water samples was developed. Optimization of the parameter settings of the ion source and mass analyzer as well as evaluation of solvent composition were carried out by continuous introduction of standards through a syringe pump. In negative ion mode the electrospray ionization source gave acceptable results. The optimum solvent used consisted of water/acetonitrile, with no volatile bases or buffers added. A simple, off-line, manual solid-phase extraction method was developed for sample preparation of environmental water samples. Recoveries were over 86% for all compounds. The method was validated and found to be linear, selective, and robust. For analysis of a 50-mL sample, the limit of detection (LOD) ranged from 3.2 to 10.6 ng/L for all compounds, and the limit of quantitation (LOQ) from 10.6 to 35.0 ng/L. Within-day (n = 5) and total (n = 5) reproducibility were investigated at three different concentration levels and ranged from 6.2 to 9.5% and 9.4 to 12.1%, respectively. Finally, the method was applied to real-world samples. PMID- 12112617 TI - An original approach to determining traces of tetracycline antibiotics in milk and eggs by solid-phase extraction and liquid chromatography/mass spectrometry. AB - An original and highly specific method able to identify and quantify traces of five tetracycline antibiotics (TCAs) in milk and eggs is presented. This method uses a single solid-phase extraction (SPE) cartridge for simultaneous extraction and purification of TCAs in the above matrices. After diluting 5 mL of intact whole milk or 2 g egg samples with Na(2)EDTA-containing water, samples are passed through a 0.5-g Carbograph 4 extraction cartridge. After analyte elution from the SPE cartridge, an aliquot of the final extract is injected into a liquid chromatography/mass spectrometry (LC/MS) instrument equipped with an electrospray ion source and a single quadrupole. MS data acquisition is performed in the positive-ion mode and by a time-scheduled multiple-ion selected ion-monitoring program. With methanol as organic modifier, the in-source collision-induced dissociation (CID) process generated fragment ions able to pick up one methanol molecule. In several cases, these methanol-adduct fragment ions have m/z values higher than those of the protonated molecules. This event is rarely encountered in MS, thus making the analysis of TCAs by this method extremely specific. Compared with a conventional published method, the present protocol extracted larger amounts of TCAs from both milk and egg and decreased the analysis time by a factor of 3. Recovery of TCAs in milk at the 25-ppb level ranged between 81 and 96% with relative standard deviation (RSD) no larger than 9%. Recovery of TCAs in egg at the 50-ppb level ranged between 72 and 92% with RSD no larger than 7%. Estimated limits of quantification(S/N = 10) of the method were 2-9 ppb TCAs in whole milk and 2-19 ppb TCAs in eggs. PMID- 12112618 TI - On-line on-chip post-column derivatisation reactions for pre-ionisation of analytes and cluster analysis in gradient micro-liquid chromatography/electrospray mass spectrometry. AB - A system is presented that demonstrates the principle of on-line and on-chip post column derivatisation reactions in micro-high-performance liquid chromatography (micro-HPLC) hyphenated to electrospray time-of-flight mass spectrometry (ESI TOFMS). In this micro-HPLC-chip-MS set-up, the analytes are separated using gradient micro-HPLC and subsequently derivatised on-chip and detected. One of the major limitations of MS detection is its dependency on the degree of ionisation, which is widely variable and compound-specific. Optimising and controlling the degree of ionisation in a simple manner would allow MS detection to be truly generic. One way of achieving this is by pre-ionisation of analytes using simple derivatisation procedures that are both rapid and quantitative. Performing this in situ on the system described here overcomes issues of sample handling and efficiency losses when time-consuming "bench chemistry" is necessary prior to analysis. The power of the system is demonstrated by the separation of primary and secondary amines, which are subsequently derivatised with a positively charged phosphonium complex and detected in an enhanced manner. Typically, molecular cations (M(+)) are detected showing that the ionisation process is dominated by the phosphonium species, leading to more constant ionisation for a variety of compounds. In addition, stable isotopically labelled ((12)C/(13)C) phosphonium reagent is used for the reactions, allowing for inherent signal/noise (S/N) improvement and automated data processing using cluster analysis. A similar reaction scheme is used for the derivatisation of ketones and aldehydes, also demonstrating dramatic increases in sensitivity, especially with increasing temperature. Minimal loss in chromatographic fidelity in terms of retention times is observed by the introduction of the micromixer chip into the system. Optimal flow rates in micro-HPLC and ESI-MS are compatible with flow rates for the chip as well as a multitude of in-line optical detectors including UV and fluorescence. In addition, the micromixer chip can be positioned pre-column if preferred. The system is robust, easily fully automated and applicable to a wide variety of reactions. The system has a major advantage in its simple robust connection to the "normal scale" outside world. PMID- 12112619 TI - Inverse 15N-metabolic labeling/mass spectrometry for comparative proteomics and rapid identification of protein markers/targets. AB - The inverse labeling/mass spectrometry strategy has been applied to protein metabolic (15)N labeling for gel-free proteomics to achieve the rapid identification of protein markers/targets. Inverse labeling involves culturing both the perturbed (by disease or by a drug treatment) and control samples each in two separate pools of normal and (15)N-enriched culture media such that four pools are produced as opposed to two in a conventional labeling approach. The inverse labeling is then achieved by combining the normal (14)N-control with the (15)N-perturbed sample, and the (15)N-control with the (14)N-perturbed sample. Both mixtures are then proteolyzed and analyzed by mass spectrometry (coupled with on-line or off-line separation). Inverse labeling overcomes difficulties associated with protein metabolic labeling with regard to isotopic peak correlation and data interpretation in the single-experiment approach (due to the non-predictable/variable mass difference). When two data sets from inverse labeling are compared, proteins of differential expression are readily recognized by a characteristic inverse labeling pattern or apparent qualitative mass shifts between the two inverse labeling analyses. MS/MS fragmentation data provide further confirmation and are subsequently used to search protein databases for protein identification. The methodology has been applied successfully to two model systems in this study. Utilizing the inverse labeling strategy, one can use any mass spectrometer of standard unit resolution, and acquire only the minimum, essential data to achieve the rapid and unambiguous identification of differentially expressed protein markers/targets. The strategy permits quick focus on the signals of differentially expressed proteins. It eliminates the detection ambiguities caused by the dynamic range of detection. Finally, inverse labeling enables the detection of covalent changes of proteins responding to a perturbation that one might fail to distinguish with a conventional labeling experiment. PMID- 12112620 TI - A rapid and precise technique for measuring delta(13)C-CO(2) and delta(18)O-CO(2) ratios at ambient CO(2) concentrations for biological applications and the influence of container type and storage time on the sample isotope ratios. AB - A simple modification to a commercially available gas chromatograph isotope ratio mass spectrometer (GC/IRMS) allows rapid and precise determination of the stable isotopes ((13)C and (18)O) of CO(2) at ambient CO(2) concentrations. A sample loop was inserted downstream of the GC injection port and used to introduce small volumes of air samples into the GC/IRMS. This procedure does not require a cryofocusing step and significantly reduces the analysis time. The precisions for delta(13)C and delta(18)O of CO(2) at ambient concentration were +/-0.164 and +/ 0.247 per thousand, respectively. This modified GC/IRMS was used to test the effects of storage on the (18)O and (13)C isotopic ratios of CO(2) at ambient concentrations in four container types. On average, the change in the (13)C-CO(2) and (18)O-CO(2) ratios of samples after one week of storage in glass vials equipped with butyl rubber stoppers (Bellco Glass Inc.) were depleted by 0.12 and by 0.20 per thousand, respectively. The (13)C ratios in aluminum canisters (Scotty II and IV, Scott Specialty Gasses) after one month of storage were depleted, on average, by 0.73 and 2.04 per thousand, respectively, while the (18)O ratios were depleted by 0.38 and 1.20 per thousand for the Scotty II and IV, respectively. After a month of storage in electropolished containers (Summa canisters, Biospheric Research Corporation), the (13)C-CO(2) and (18)O-CO(2) ratios were depleted, on average, by 0.26 and enriched by 0.30 per thousand, respectively, close to the precision of measurements. Samples were collected at a mature hardwood forest for CO(2) concentration determination and isotopic analysis. A comparison of CO(2) concentrations determined with an infrared gas analyzer and from sample voltages, determined on the GC/IRMS concurrent with the isotopic analysis, indicated that CO(2) concentrations can be determined reliably with the GC/IRMS technique. The (13)C and (18)O ratios of nighttime ecosystem respired CO(2), determined from the intercept of Keeling plots, were -26.11 per thousand (V-PDB) and -8.81 per thousand (V-PDB-CO(2)), respectively. PMID- 12112621 TI - A method to evaluate tryptic digestion efficiency for high-throughput proteome analyses. PMID- 12112623 TI - Biomarker study of a municipal effluent dispersion plume in two species of freshwater mussels. AB - The toxicological effects of a primary-treated municipal effluent plume were investigated in two species of freshwater mussels, Elliptio complanata and Dreissena polymorpha, exposed for 62 days at sites upstream and downstream of an effluent outfall in the St. Lawrence River (Quebec, Canada). Levels of metallothioneins (MT), cytochrome P4501A1 activity, DNA damage, total lipids, relative levels of vitellins, and phagocytic activity (in E. complanata hemocytes) were determined after the exposure period. A parallel analysis measured heavy metals and coprostanol in mussel tissues. The results show that significant levels of coprostanol and some metals (specifically, Cu, Hg, Sb, Se, and Zn) had accumulated in mussels caged 5 km downstream of the effluent plume. Mixed-function oxidase activity, MT in gills, total lipids, DNA damage (in D. polymorpha only), and total hemolymph bacteria (in E. complanata only) had increased in these mussels, while levels of total cadmium (Cd), MT in digestive glands or whole soft tissues, phagocytic activity, and DNA damage in the digestive gland (in E. complanata only) were diminished. The exposure of mussels to surface waters contaminated by a municipal effluent led to many stress responses, depending on both the tissues and the species being examined. PMID- 12112624 TI - Immunocompetence of bivalve hemocytes as evaluated by a miniaturized phagocytosis assay. AB - Immune function in bivalves can be adversely affected by long-term exposure to environmental contaminants. Investigating alterations in immunity can therefore yield relevant information about the relationship between exposure to environmental contaminants and susceptibility to infectious diseases. We have developed a rapid, cost-effective, and miniaturized immunocompetence assay to evaluate the phagocytic activity, viability, and concentration of hemocytes in freshwater and marine bivalves. Preliminary experiments were performed to optimize various aspects of the assay including 1) the time required for adherence of hemocytes to polystyrene microplate wells, 2) the time required for internalization of fluorescent bacteria, 3) the ratio of hemocytes to fluorescent bacteria in relation to phagocytosis, 4) hemolymph plasma requirements, and 5) the elimination of fluorescence from (noninternalized) bacteria adhering to the external surface of hemocytes. The results of these experiments showed the optimal adherence time for hemocytes in microplate wells to be 1 h, that phagocytosis required at least 2 h of contact with fluorescently labeled E. coli cells, that the number of fluorescent E. coli cells had a positive effect on phagocytic activity, that at least 2.5 million cells/mL were required to measure a significant intake, and that a linear increase in uptake of bacteria (R = 0.91; p < 0.01) could be obtained with concentrations of up to 1.3 x 10(6) hemocytes/mL. Afterward, the assay was used in two field studies to identify sites having the potential to affect the immunocompetence of bivalves. The first study was conducted on Mya arenaria clams collected at selected contaminated sites in the Saguenay River (Quebec, Canada), and the second examined Elliptio complanata freshwater bivalves that had been exposed to a municipal effluent plume in the St. Lawrence River (Quebec, Canada). In the Saguenay River field study a significant increase in phagocytosis was observed at sites closest to polluted areas. Phagocytotic activity varied over time and was highest during the warmest months (June, July, and August), closely paralleling the spawning period of Mya arenaria clams. In contrast, a drop in phagocytic activity was observed in Elliptio complanata mussels exposed to surface water 4 km downstream of a major municipal effluent plume, with a concomitant increase in the number of hemocytes in the hemolymph. It appears that both immunosuppressive and immunostimulative effects are likely to occur in the field and that responses will be influenced by the type and intensity of contaminants at play. PMID- 12112625 TI - Molluscan shellfish biomarker study of the Quebec, Canada, Saguenay Fjord with the soft-shell clam, Mya arenaria. AB - A spatial and temporal survey of six sites in the Saguenay Fjord and of one adjacent site in the St. Lawrence River estuary (Quebec, Canada) was undertaken to study the possible effects of anthropogenic contaminant input on soft-shell clam (Mya arenaria) populations. Bivalve sampling sites were selected because they reflected a range of areas representative of either no known (or apparent) pollution sources or of areas potentially influenced by different gradients and types of contamination sources. The most upstream site selected in the Saguenay Fjord, nearest to a highly populated and industrialized sector, and the most downstream site, near its mouth with the St. Lawrence River estuary, spanned a distance of some 70 km and encompassed the entire intertidal area suitable for Mya arenaria habitat. To measure effects in collected animals, we used a comprehensive battery of biomarkers composed of metallothionein-like proteins (MT), 7-ethoxyresorufin O-deethylase activity (EROD), DNA damage (DD), lipid peroxidation (LPO), vitellinlike proteins (Vn), phagocytosis (PHAG), nonspecific esterase (NspE) activity, and condition factor (weight-to-length ratio of clams). Vn, PHAG, DD, and NspE biomarkers were assayed in hemolymph (or hemocytes), whereas others (MT, EROD, LPO) were determined in the digestive gland. Whole tissue metal content was also quantified in clams collected in the spatial survey. The spatial survey conducted in June 1997 showed significant effects at all sites, and principal component analysis indicated in addition that the more important responses were linked to the MT, LPO, and NspE biomarkers. Clams collected from sites closest to the upstream reaches of the fjord generally displayed higher levels of tissue metals (cadmium, manganese), as well as greater responses of NspE activity, MT, LPO, and PHAG. Animals collected from sites influenced by municipal wastewaters had higher levels of Vn, suggesting the presence of environmental estrogens. The results of the temporal survey (six monthly samplings of clams at three sites from May through October, 1997) showed that the bivalve reproductive cycle (vitellogenesis and spawning) can modulate the expression of several biomarkers. Vn levels, for example, were positively correlated with DD and EROD and negatively correlated with MT, suggesting that reproduction can influence the susceptibility of clams to some contaminants. Discrimination analysis over the 6 months of sampling revealed that the mean value of the discriminant function changed significantly over time, suggesting important changes in the relative contribution of each biomarker. In short, this study has provided evidence that clam populations in the Saguenay Fjord are impacted by multiple sources of contamination whose effects can be modulated by reproduction. PMID- 12112627 TI - A microscale test to measure petroleum oil toxicity to mummichog embryos. AB - A test was developed to compare the toxicity of different petroleum oils to mummichog (Fundulus heteroclitus) embryos. Fertilized eggs were incubated for 11 days at 22.5 degrees C directly on the surface of oil-contaminated sand without a superficial water layer. The mortality rates, the stage of development, and the prevalence of malformations were determined. No effect was found in controls incubated on sand with water and mineral oil as compared with controls on sand with water alone. Two weathered oils, an Alaska North Slope crude oil (ANCO) and a Mesa light crude oil (MLCO), produced similar symptoms of toxicity: retarded growth and development, pericardial edema, hemostasis, hemorrhages, and spinal deformities. These symptoms are consistent with those observed in other fish species exposed to petroleum oils, suggesting that the results of the bioassay would be applicable to other species. MLCO was more embryotoxic than ANCO. The minimal oil concentrations causing a significant reduction in body length were 4.5 microg oil/g sand for MLCO and 12.7 microg oil/g for ANCO, indicating the assay is sensitive. The slopes and the intercepts of the relationships between concentration and growth did not differ in three dose-response experiments conducted with each oil, indicating that the assay is reliable. Finally, the bioassay is less costly than other available options to assess the toxicity of petroleum oils to marine fish embryos. Further work to improve the standardization of the assay will involve comparison of the toxicity of petroleum oils with reference toxicants and selection of a standard substrate. PMID- 12112626 TI - A novel nucleolar biomarker in plant and animal cells for assessment of substance cytotoxicity. AB - The cytotoxicity of three substances (mercury(II), metolachlor, and 4 nitroquinoline-N-oxide) was assessed with a set of nucleolar parameters: the average number of nucleoli, the average volume of a single nucleolus, and the proportion of cells with heteromorphic-paired nucleoli (PNhet). Their toxic impact was studied on cells of animal and plant test organisms: onion (Allium cepa), lettuce (Lactuca sativa), and hydra (Hydra attenuata). In general, at concentrations near IC/LC(50) the three chemicals produced similar cytogenetic effects after 30-360 min of contact. For instance, in plant cells (Allium cepa and Lactuca sativa) the toxicants increased the percentage of cells with PNhet, decreased the volume of single nucleoli, and exerted no significant impact on the nucleolar number. In animal cells (Hydra attenuata), they reduced the size of nucleoli, produced no effect on the number of nucleoli, but decreased the share of cells with PNhet. Also, the chemicals affected the cells of the three test organisms to different degrees. Thus, the effectiveness of our approach of using nucleolar biomarker (use of the proposed set of parameters and time schedule of several determinations in the first hours of toxicant contact, etc.) as a means of assessing cytotoxicity was confirmed. PMID- 12112629 TI - The mycotoxins citrinin, gliotoxin, and patulin affect interferon-gamma rather than interleukin-4 production in human blood cells. AB - Exposure to molds diminishes the numbers of T-helper type 1 (Th1) cells in the peripheral blood of children and is a risk factor for the development of allergic diseases (results of LARS: Leipzig Allergy Risk Children Study, Mueller et al. 2002). We hypothesized that mycotoxins are responsible for this effect and therefore investigated the influence of citrinin, gliotoxin, and patulin on human peripheral blood mononuclear cells (PBMC). CD3/CD28-stimulated PBMC of healthy donors were incubated for 24 h with the mycotoxins in serial dilutions and triplicates. Vitality and proliferation were tested using the MTT assay and T cell function by the expression of cytokines (ELISA, intracellular cytokine staining, and real-time polymerase chain reaction (RT-PCR) for interferon-gamma (IFN-gamma) and interleukin-4 (IL-4). The cytokine secretion was inhibited at concentrations 2-130 times lower compared to vitality (ELISA versus MTT assay). The strongest inhibition of cytokine expression was found for IFN-gamma: 8.3 microg/mL citrinin, 34.2 ng/mL gliotoxin, and 64.8 ng/mL patulin caused a 50% inhibition of the IFN-gamma release (50% inhibitory dose, ID(50)). For IL-4 release the corresponding ID(50) values were 21.6 microg/mL citrinin, 82.8 ng/mL gliotoxin, and 243.2 ng/mL patulin. Furthermore, 3 ng/mL patulin caused a significant increase of IL-4 but a significant suppression of IFN-gamma. On the mRNA level, after 24 h an unaltered or enhanced IL-4 was observed compared to a reduced IFN-gamma expression. Using a method of intracellular cytokine staining, we were able to show that the described effects are caused by a reduction of the number of IFN-gamma-producing T lymphocytes rather than by a reduced functional capacity of the single cell. We suggest that mycotoxins primarily cause stronger inhibition of IFN-gamma-producing Th1 cells, which may lead to T-cell polarization toward the Th2 phenotype and may raise the risk for the development of allergies. PMID- 12112628 TI - The influence of maternal exposure to volatile organic compounds on the cytokine secretion profile of neonatal T cells. AB - Indoor VOC (volatile organic compound) exposure has been shown to be correlated with airway symptoms and allergic manifestations in children. An investigation was conducted within an ongoing birth cohort study (LISA: Lifestyle-Immune System Allergy) of the association between maternal exposure to VOCs and immune status at birth, in particular the cytokine secretion profile of cord-blood T cells. In a randomly selected group of 85 neonates, cytokine-producing cord-blood T cells were analyzed using intracellular cytokine detection. VOC exposure was measured in children's dwellings by passive sampling, while parents were asked to complete questionnaires about possible sources of VOC exposure. Adjusted odds ratios (ORs) were calculated by logistic regression based on categorized quartiles. A positive association was found between elevated percentages of interleukin-4-producing (IL 4) type 2 T cells and exposure to naphthalene (OR = 2.9) and methylcyclopentane (OR = 3.3). Exposure to tetrachloroethylene was associated with reduced percentages of interferon-gamma-producing (IFN-gamma) type 1 T cells (OR = 2.9). In addition, smoking during pregnancy was correlated with a higher indoor air concentration of naphthalene (OR = 3.8), new carpets in infants' bedrooms with elevated methylcyclopentane concentrations (OR = 4.1), and home renovation with a higher trichloroethylene burden (OR = 4.9). Our data suggest that maternal exposure to VOC may have an influence on the immune status of the newborn child. PMID- 12112630 TI - Assessment of mutagenicity and toxicity of different-size fractions of air particulates from La Plata, Argentina, and Leipzig, Germany. AB - Airborne particulates, especially fine particles and bound chemical compounds, are a potential mediator of adverse health effects. In this study an analysis was done of the concentration and size distribution of air particulate matter, the content of bound polycyclic aromatic hydrocarbons (PAHs), and the biological effects of organic extracts from different fractions of dust that had been influenced by urban and industrial emissions from the regions of La Plata, Argentina, and Leipzig, Germany, along with samples from control areas. Air particulate matter was sampled in summer and winter in each region using a high volume sampler with a six-stage cascade impactor, classifying dust in six size fractions from 10-microm particles to those less than 0.49 microm in size. Organic extracts of dust were tested for mutagenicity (Ames test, Salmonella typhimurium TA98 strain, S9+) and cytotoxicity (Tetrahymena pyriformis test system, growth rate, cell respiration). The content of PAHs was analyzed by high performance liquid chromatography (HPLC) with diode array and fluorescence detection. Mutagenic and cytotoxic effects were found to be associated with very fine (<0.49 microm) and fine (<1.5 microm) particle-bound compounds, corresponding to a higher content of PAHs. The mutagenic potency (revertants/m(3)) associated with particles less than 0.49 microm from urban areas of La Plata was about 1 order of magnitude higher than in particles in the range of 3.0 to 0.49 microm. Fine fractions from sites with an industrial burden were also found to exhibit high mutagenic potency. A similar tendency was observed in cytotoxicity tests with T. pyriformis. This cell system proved to be very sensitive to toxicants in tested dust fractions. The observed biological effects were found to be correlated significantly with concentrations of total PAH, carcinogenic PAHs, and benzo[a]pyrene. PMID- 12112631 TI - Overview of results from the WaterTox intercalibration and environmental testing phase II program: Part 1, statistical analysis of blind sample testing. AB - There is an urgent need to evaluate the presence of toxicants in waters used for human consumption and to develop strategies to reduce and prevent their contamination. The International Development Research Centre undertook an intercalibration project to develop and validate a battery of bioassays for toxicity testing of water samples. The project was carried out in two phases by research institutions from eight countries that formed the WaterTox network. Results for the first phase were reported in the special September 2000 issue of Environmental Toxicology. Phase II involved toxicity screening tests of environmental and blind samples (chemical solutions of unknown composition to participating laboratories) using the following battery: Daphnia magna, Hydra attenuata, seed root inhibition with Lactuca sativa, and Selenastrum capricornutum. This battery was also used to assess potential toxicity in concentrated (10x) water samples. Results are presented for a set of six blind samples sent to the participating laboratories over a 1-year period. Analyses were performed for each bioassay to evaluate variations among laboratories of responses to negative controls, violations of test quality control criteria, false positive responses induced by sample concentration, and variability within and between labs of responses to toxic samples. Analyses of the data from all bioassays and labs provided comparisons of false positive rates (based on blind negative samples), test sensitivities to a metal or organic toxicant, and interlaboratory test variability. Results indicate that the battery was reliable in detecting toxicity when present. However, some false positives were identified with a concentrated soft-water sample and with the Lactuca and Hydra (sublethal end-point) tests. Probabilities of detecting false positives for individual and combined toxic responses of the four bioassays are presented. Overall, interlaboratory comparisons indicate a good reliability of the battery. PMID- 12112632 TI - Overview of results from the WaterTox intercalibration and environmental testing phase II program: part 2, ecotoxicological evaluation of drinking water supplies. AB - Because of rapid population growth, industrial development, and intensified agricultural production increasing amounts of chemicals are being released into the environment, polluting receiving water bodies around the world. Given the potential health risk associated with the presence of toxicants in water sources used for drinking yet the scarcity of available data, there is a need to evaluate these waters and develop strategies to reduce and prevent their contamination. The present study examined the applicability of a battery of simple, inexpensive bioassays in environmental management and the relevance of the test results in establishing the toxicological quality of water sources and drinking water within the framework of the eight-country WaterTox Network, sponsored by the International Development Research Centre, Ottawa, Canada. Seventy-six samples were collected from surface and groundwater sources and seven samples from drinking water treatment plants. Each sample was tested with a core battery of bioassays (Daphnia magna, Hydra attenuata, and Lactuca sativa root inhibition tests) and a limited set of physical and chemical parameters. In addition, three labs included the Selenastrum capricornutum test. When no toxic effects were found with the battery, samples were concentrated 10x using a solid-phase extraction (SPE) procedure. Nonconcentrated natural water samples produced a toxic response in 24% of cases with all three core bioassays. When all bioassays are considered, the percentage of raw samples showing toxicity with at least one bioassay increased to 60%. Of seven treated drinkingwater samples, four showed toxicity with at least one bioassay, raising the possibility that treatment processes in these instances were unable to remove toxic contaminants. The Daphnia magna and Hydra attenuata tests indicated a high level of sensitivity overall. Although only three of the eight countries used S. capricornutum, it proved to be an efficient and reliable bioassay for toxicity assessment. PMID- 12112633 TI - Quality of water types in Ukraine evaluated by WaterTox bioassays. AB - The quality of river, ground-, and tap water was analyzed using the basic set of WaterTox bioassays [Daphnia (Daphnia magna), Hydra (Hydra attenuata), and lettuce (Lactuca sativa)] as well as two additional bioassays, onion (Allium cepa) and microalga (Selenastrum gracile). Samples of these waters were also concentrated fivefold using a solid-phase procedure. The results of the Daphnia and Hydra bioassays showed that the winter and spring concentrated and nonconcentrated samples from the Dnieper and Desna rivers, the main water supply sources for Kiev, were nontoxic. In spring, after concentration, the two river samples brought about the same relative decrease in the lettuce root length (by 35%, p < 0.001), where the Desna River sample considerably reduced (by 79.1%, p < 0.001) the number of microalga cells. Samples of groundwater from countryside wells studied in autumn in several villages of the Kiev region were toxic mainly to Hydra (sublethal effects were found in 11%-78%) and lettuce (the root length decreased 15%-56%). Studies of tap water samples from two of the largest cities of Ukraine, Kiev and Kharkiv, were found to be nontoxic to both plants, lettuce and onion, but showed increased sublethal and lethal effects on both animals, Daphnia and Hydra, as well as a reduced number of microalgae. Different bioassays were sensitive to varying degrees to different water types. This reinforces the necessity of using sets of bioassays in toxicity evaluation. In general, all the tested water samples demonstrated some toxicity. These data suggest that drinking water quality in Ukraine needs improvement. PMID- 12112634 TI - Determining toxicity of leachates from Florida municipal solid waste landfills using a battery-of-tests approach. AB - The toxicity and physicochemical parameters of municipal solid-waste (MSW) landfill leachates from six sites in north and north-central Florida, United States, were determined. Landfill leachates are complex mixtures of organic and inorganic constituents. Leachate toxicity was assayed using the acute C. dubia (48-h) and Microtoxtrade mark (15-min) assays and the chronic S. capricornutum (96-h) assay. The landfill leachates were shown to be highly toxic to both C. dubia and S. capricornutum with an EC(50) < 10% and < 15%, respectively. However, Microtoxtrade mark was not as sensitive to the leachates. Based on these results, the assays were ranked for their sensitivities to the landfill leachates; C. dubia > S. capricornutum > Microtoxtrade mark. Chemical analyses showed high concentrations of un-ionized ammonia and salts in the landfill leachates but low concentrations of heavy metals. The toxicity of the landfill leachates varied between the sites sampled and within each of the landfill sites. A significant relationship was found between the total ammonia content of the leachates and toxicity as determined by the C. dubia and S. capricornutum assays. Although the chemical constituents in Florida MSW landfill leachates may be more dilute, the toxicity of the leachates is equivalent to that from industrial waste landfills. PMID- 12112635 TI - Increased metal bioavailability following alteration of freshwater dissolved organic carbon by ultraviolet B radiation exposure. AB - Dissolved organic carbon (DOC) is critically important in the chemistry of freshwater. It complexes heavy metals, making them less bioavailable to aquatic organisms, and absorbs and attenuates UVB radiation, undergoing degradation and alteration in the process. This study examined changes in metal toxicity to the freshwater green alga Pseudokirchneriella subcapitata in high- and low-DOC natural water samples after exposure to UVB radiation. Brown-water and clear water samples were irradiated for 0, 5, and 10 days. The DOC concentrations of the samples were measured, and they were subsequently spiked with Cu, Ni, Zn, Cd, Co, and Pb and used in algal bioassays to measure changes in metal toxicity following irradiation. DOC concentrations declined only 20% with UVB irradiation in both samples, although DOC concentration was much higher in the brown-water sample than in the clear-water sample. In the brown-water sample metal toxicity increased up to 78% after 10 days of UVB irradiation for Cu, Zn, Co and Pb, but not for Ni and Cd. Changes were less evident in the clear-water sample. The differences observed between IC(50) values for relatively fresh, high-DOC water from the headwaters of the Raisin River and much "older," low-DOC water from Lake Simcoe point to the likelihood of the effects observed in this study being many times greater in the natural environment because of very long exposure to solar radiation. Alteration of DOC by UVB irradiation may influence primary productivity and species composition, especially in waters in which metal concentrations are high. PMID- 12112636 TI - Response of the charophyte Nitellopsis obtusa to heavy metals at the cellular, cell membrane, and enzyme levels. AB - The responses of the freshwater macroalga Nitellopsis obtusa to heavy metal (HM) salts of Hg, Cd, Co, Cu, Cr, and Ni were assessed at different levels: whole-cell mortality (96-h LC(50)), in vivo cell membrane (45-min depolarization of resting potential, EC(50)), and enzyme in plasma membrane preparations (K+, Mg2+-specific H+-ATPase inhibition, IC(50)). To measure ATPase activity, a novel procedure for isolation of plasma membrane-enriched vesicles from charophyte cells was developed. The short-term ATPase inhibition assay (IC(50) from 6.0 x 10(-7) to 4.6 x 10(-4) M) was slightly more sensitive than the cell mortality test (LC(50) from 1.1 x 10(-6) to 2.6 x 10(-3) M), and the electrophysiological test with the end point of 45-min depolarization of resting potential was characterized by less sensitivity for HMs (EC(50) from 1.1 x 10(-4) to 2.2 x 10(-2) M). The variability of IC(50) values assessed for HMs in the ATPase assays was close to that of LC(50) values in the mortality tests (CVs from 33.5 to 83.5 and from 12.4% to 57.7%, respectively), whereas the EC(50) values in the electrophysiological tests were characterized by CVs generally below 30%. All three end points identified two separate HM groups according to their toxicity to N. obtusa: Co, Ni, and Cr comprised a group of less toxic metals, whereas Hg, Cu, and Cd comprised a group of more toxic metals. However, the adverse effects within each group were discriminated differently. For example, the maximum difference between the highest and lowest LC(50) for the group of less toxic metals in the long-term mortality test was approximately 60% of the response range, whereas the corresponding difference in IC(50) values in the ATPase assay was 30%. In contrast, the LC(50) values of the more toxic metals occupied only 10% of the response range, whereas the IC(50) values were spread over 70%. Further investigation should be done of the underlying mechanism or mechanisms responsible for the observed differences in the dynamic range of a particular end point of the groups of toxicants of varying strength. PMID- 12112637 TI - Direct toxicity assessment of wastewater: Baroxymeter, a portable rapid toxicity device and the industry perspective. AB - Direct toxicity assessment of wastewater is becoming necessary, and new legislation may render it compulsory for the water industry. At present such assessment is performed at a laboratory away from a site, at considerable cost, and results often come too late, after a toxic event has occurred and the toxin has been released into the environment. Some of the rapid toxicity tests available today require certain conditions to function properly, or their results do not always correlate with other methods. The objective of this study was to assess a portable device, the Baroxymeter, for its suitability as an instrument to test wastewater toxicity. The way the device works is based on monitoring respiration of a bacterial culture by pressure measurements and using respiration inhibition as a toxicity alert. It has been shown that it is possible to detect toxic substances such as 3,5-dichlorophenol and bronopol within 5 min from a 1-mL sample. The benefits and future applications of the Baroxymeter as a high throughput, cost-effective alternative for toxicity screening are discussed in this article. PMID- 12112638 TI - A novel and sensitive test for rapid determination of water toxicity. AB - The performance of a novel, rapid, and sensitive test for detecting chemical toxicants in water is described in this article. The bioassay utilizes a highly sensitive variant of the luminescent bacterium Photobacterium leiognathi that allows the detection in water at levels below milligrams per liter of diverse groups of toxicants, including heavy metals, pesticides, PCBs, polycyclic aromatic hydrocarbons, and fuel traces. For most toxic agents reported in this study, the new assay was markedly more sensitive than the Microtox(trade mark) Vibrio fischeri assay according to the bacterial bioluminescence toxicity data reported in the literature. Additional features of the new bioassay include the ability to discriminate between cationic heavy metals and organic toxicants and the option of being run at ambient temperatures (18 degrees C-27 degrees C), thereby enabling on-site testing with low-cost luminometers. In addition, the stability of the freeze-dried bacterial reagent preparation at ambient temperatures precludes the need for refrigeration or freezing during shipment, which contributes to further reducing overall operational costs. PMID- 12112640 TI - Generation of stable retrovirus packaging cell lines after transduction with herpes simplex virus hybrid amplicon vectors. AB - BACKGROUND: A number of properties have relegated the use of Moloney murine leukemia virus (Mo-MLV)-based retrovirus vectors primarily to ex vivo protocols. Direct implantation of retrovirus producer cells can bypass some of the limitations, and in situ vector production may result in a large number of gene transfer events. However, the fibroblast nature of most retrovirus packaging cells does not provide for an effective distribution of vector producing foci in vivo, especially in the brain. Effective development of new retrovirus producer cells with enhanced biologic properties may require the testing of a large number of different cell types, and a quick and efficient method to generate them is needed. METHODS: Moloney murine leukemia virus (Mo-MLV) gag-pol and env genes and retrovirus vector sequences carrying lacZ were cloned into different minimal HSV/AAV hybrid amplicons. Helper virus-free amplicon vectors were used to co infect glioma cells in culture. Titers and stability of retrovirus vector production were assessed. RESULTS: Simultaneous infection of two glioma lines, Gli-36 (human) and J3T (dog), with both types of amplicon vectors, generated stable packaging populations that produced retrovirus titers of 0.5-1.2 x 10(5) and 3.1-7.1 x 10(3) tu/ml, respectively. Alternatively, when cells were first infected with retrovirus vectors followed by infection with HyRMOVAmpho amplicon vector, stable retrovirus packaging populations were obtained from Gli-36 and J3T cells producing retrovirus titers comparable to those obtained with a traditional retrovirus packaging cell line, Psi CRIPlacZ. CONCLUSIONS: This amplicon vector system should facilitate generation of new types of retrovirus producer cells. Conversion of cells with migratory or tumor/tissue homing properties could result in expansion of the spatial distribution or targeting capacity, respectively, of gene delivery by retrovirus vectors in vivo. PMID- 12112641 TI - The tet-off system is more effective than the tet-on system for regulating transgene expression in a single adenovirus vector. AB - BACKGROUND: Control of transgene expression in mammalian cells is desirable for gene therapy and the study of gene function in basic research. This study evaluates the functionality of single adenovirus (Ad) vectors containing a tetracycline-controllable expression system (tet-off or tet-on system). METHODS: An Ad-mediated binary transgene expression system was generated containing a tet off or a tet-on system, which introduced the gene of interest with a tetracycline regulatable promoter and the tetracycline-responsive transcriptional activator gene into the E1 and E3 deletion regions, respectively. The functionality of the Ad-mediated tet-off and tet-on systems was compared under various conditions in vitro and in vivo. RESULTS: The Ad vector containing the tet-off system provided negative control of gene expression ranging from 20- to 500-fold, depending on the cell type and condition. In contrast, the Ad vector containing the tet-on system increased gene expression by only 2- to 28-fold and required about two-log orders higher concentration of an inducer (doxycycline) to switch on the gene expression, compared with the Ad vector containing the tet-off system. CONCLUSION: Ad vectors containing the tet-off system are a better choice for regulated gene expression than Ad vectors containing the tet-on system. Single Ad vectors containing the tetracycline-controllable expression system will greatly facilitate in vitro and in vivo analyses of gene function and may be useful for gene therapy. PMID- 12112642 TI - Activation of silenced transgene expression in neural precursor cell lines by inhibitors of histone deacetylation. AB - BACKGROUND: Ex vivo gene therapy in the central nervous system (CNS) holds great promise for diseases such as the neurodegenerative disorders. However, achieving stable, long-term transgene expression in grafted cells has proven problematic. This study reports the establishment of an in vitro model of transgene down regulation in cells grafted to the CNS using the immortalized neural progenitor cell lines HiB5 and RN33B. METHODS: Neural cell lines were transduced at 33 degrees C with different GFP constructs, both viral and non-viral, containing either viral or non-viral promoters. Cell differentiation in vitro was obtained by culturing the cells at 37 degrees C in serum-free defined media, which halts cell division, and GFP-expression was analysed by FACS. As early as day 3 of culture at 37 degrees C, the transgene expression decreased markedly in most cell lines. To validate the assay, the same clones were grafted to the adult rat striatum and the down-regulation of GFP-expression was evaluated. RESULTS: The temporal pattern of down-regulation was found to be similar in vitro and in vivo. Using this assay, it was shown that addition of inhibitors of histone deacetylation, but not an inhibitor of DNA methylation, reversed the silencing of GFP in quiescent neural progenitors by up to 308% of control values. CONCLUSION: These results suggest that the same mechanisms controlling gene transcription of the host cell's genome are active in controlling transgene expression and that this should be taken into account when constructing vectors for gene therapy. The assay reported in this study could be used as a screening method to evaluate new vectors. PMID- 12112643 TI - Optimization and direct comparison of the dimerizer and reverse tet transcriptional control systems. AB - BACKGROUND: Exogenously controlled gene expression systems are essential for both the in vivo analysis of gene function and the regulated delivery of therapeutic gene products. However, differences in experimental methods used to characterize the various systems prohibit informative comparisons. The purpose of this study was to identify an optimal system for regulated gene expression studies through a rigorous direct comparison of the dimerizer and the reverse tet transactivator (rtTA) transcriptional switch systems. METHODS: An optimized bicistronic rapamycin-dependent dimerizer construct and an optimized rtTA construct (based on rtTA(s)-M2) were developed that utilize a chimeric mammalian activation domain with a flexible interdomain linker. These constructs were reconstituted in identical eukaryotic expression vectors and compared in transient transfection assays employing target gene reporter constructs that differ only in the relevant DNA binding sites. RESULTS: The optimized rtTA(s)-M2 construct, designated rtTAM2.2, exhibited a twofold increase in the magnitude of doxycycline-dependent reporter gene induction and an eightfold increase in sensitivity as compared to the rtTA(s)-M2 construct. This correlated with a significantly higher level of expression of the rtTAM2.2 protein. In direct comparisons the rtTAM2.2 system mediated inducible expression to a level tenfold greater than the bicistronic dimerizer system. However, while the dimerizer system exhibited no detectable rapamycin-independent expression, a low level of doxycycline-independent target gene expression was detectable. CONCLUSIONS: The improved rtTAM2.2 rtTA system described here may prove optimal when the overall magnitude of target gene induction is critical, while the dimerizer system may be advantageous when the complete absence of ligand-independent target gene expression is essential. PMID- 12112644 TI - Intracellular processing and stability of DNA complexed with histidylated polylysine conjugates. AB - BACKGROUND: Glycosylated polylysines and histidylated polylysines complexed with plasmid DNA (pDNA) were proposed to develop polymer-based gene delivery systems. The present work has been undertaken in two steps to study the uptake and the intracellular processing of pDNA, which are still poorly understood in the polyfection pathway. METHODS AND RESULTS: The kinetics of the uptake and the intracellular processing of pDNA complexed with lactosylated polylysine, histidylated polylysine or histidylated polylysine bearing lactosyl residues (polyplexes) into a CF human airway epithelial cell line were assessed by flow cytometry and confocal microscopy. Complexes formed from histidylated polylysine, even though they were less taken up by cells, show better transfection efficiency with compared with lactosylated complexes. Lactosylated polymers segregated more rapidly when compared with non-lactosylated polymers into compartments different from those containing pDNA on internalization. Intracellular location and pH measurements indicated that polymers ended up in compartments of pH approximately 6.2 while pDNA reached less acidic compartments of pH approximately 6.6. These compartments did not contain the LAMP-1 lysosomal marker. CONCLUSIONS: The present study exhibits that, upon internalization, pDNA and polylysine conjugates underwent segregation with a rate depending on the polylysine substitution and polymer degradation. The better transfection efficiency of polyplexes with histidylated polylysine can be ascribed to their prolonged stability inside the endocytic vesicles that likely favored the pDNA escape in the cytosol. PMID- 12112645 TI - Somatic gene transfer into the lactating ovine mammary gland. AB - BACKGROUND: Somatic gene therapy requires safe and efficient techniques for the gene transfer procedure. The ovine mammary gland is described as a model system for the evaluation of somatic gene transfer methods. METHODS: Different gene delivery formulations were retrogradely injected into the mammary gland of lactating sheep. The efficiency of the gene transfer was subsequently measured by the detection of the secreted transgene products in the milk. To counteract the milk flow in the lactating gland caused by the permanent milk production, a newly developed pretreatment of the mammary gland with hyperosmotic solutions was applied. In addition, in vivo electroporation of DNA into the mammary gland is described. RESULTS: Gene transfer using naked DNA or simple complexes of DNA with polycations did not result in traceable amounts of reporter gene products. However, utilizing the complex cationic lipid DOSPER, a peak expression of about 400 ng/ml was observed 6 days after transfection. Maximum expression rates of more than 1 microg/ml were obtained by combining hyperosmotic pretreatment and receptor-mediated gene transfer. For the in vivo electroporation, the proof of principle for this technique in the mammary gland is reported. CONCLUSIONS: The ovine mammary gland turned out to be a very well suited as a model system for evaluation and optimization of various gene transfer protocols. PMID- 12112646 TI - High efficiency transfection of porcine vascular cells in vitro with a synthetic vector system. AB - BACKGROUND: Gene therapy strategies for the treatment of vascular disease such as the prevention of post-angioplasty restenosis require efficient, non-toxic transfection of vascular cells. In vitro studies in these cells contribute to vector development for in vivo use and for the evaluation of genes with therapeutic potential. The aim of this project was to evaluate a novel synthetic vector consisting of a liposome (L), an integrin targeting peptide (I), and plasmid DNA (D), which combine to form the LID vector complex. METHODS: Cultures of porcine smooth muscle cells and endothelial cells were established and then transfected with the LID vector, using the reporter genes luciferase and green fluorescent protein and the metalloprotease inhibitor TIMP-1. RESULTS: The LID vector system transfected primary porcine vascular smooth muscle cells and porcine aortic endothelial cells with efficiency levels of 40% and 35%, respectively. By increasing the relative DNA concentration four-fold, incubation periods as short as 30 min achieved the same levels of luciferase transgene expression as 4 h incubations at lower DNA concentrations. The transfection did not affect cell viability as measured by their proliferative potential. Serum levels of up to 20% in the transfection medium had no adverse affect on the efficiency of transfer and gene expression in either cell type. Transfections with the cDNA for TIMP-1 produced protein levels that peaked at 130 ng/ml per 24 h and persisted for 14 days at 10 ng/ml per 24 h. CONCLUSION: This novel vector system has potential for studies involving gene transfer to cardiovascular cells in vitro and in vivo. PMID- 12112647 TI - Syngeneic fibroblasts transfected with a plasmid encoding interleukin-4 as non viral vectors for anti-inflammatory gene therapy in collagen-induced arthritis. AB - BACKGROUND: No effective long-term treatment is available for rheumatoid arthritis. Recent advances in gene therapy and cell therapy have demonstrated efficiency in collagen-induced arthritis (CIA). Interleukin-4 (IL-4) is already known to be efficient in CIA in systemic injection or administered by gene therapy. This study was designed to evaluate the effect of a non-viral gene therapy of CIA, involving injection of syngeneic fibroblasts transfected with a plasmid encoding for IL-4. METHODS: Immortalised fibroblasts from DBA/1 mice (DBA/1/0 cells) were transfected with a plasmid expressing IL-4 cDNA (DBA/1/IL-4 cells). Xenogeneic fibroblasts from Chinese hamster ovary (CHO) transfected with a plasmid expressing IL-4 cDNA (CHO/IL-4) were studied also. The cells were engrafted in mice developing CIA by subcutaneous injection of 3 x 10(6) DBA/1/0 or DBA/1/IL-4 or CHO/IL-4 cells. RESULTS: Injection of DBA/1/IL-4 cells, on days 10 and 25 after immunisation, was associated with a significant and lasting improvement in the clinical and histological evidence of joint inflammation and destruction as compared with DBA/1/0 and CHO/IL-4 cells. DBA/1/IL-4 cell treatment decreased also the production of IgG2a antibody to CII and the proliferation of CIIB-specific nodal T cells. Later treatments (engraftments on days 23 and 35 after immunisation) exerted also an anti-inflammatory effect, as evaluated on clinical and histological signs of CIA. CONCLUSIONS: Taken together, these findings indicate that systemic administration of syngeneic cells transfected with an anti-inflammatory cytokine gene, namely IL-4, with a non viral method is effective in CIA and may attenuate the cytokine imbalance seen in this disease. PMID- 12112648 TI - Non-viral vector-mediated uptake, distribution, and stability of chimeraplasts in human airway epithelial cells. AB - BACKGROUND: Chimeraplasty is a novel methodology that uses chimeric RNA/DNA oligonucleotides (chimeraplasts) to stimulate genomic DNA repair. Efficient uptake and nuclear localization of intact chimeraplasts are key parameters to achieve optimal correction of mutation defects into specific cell types. METHODS: A 5'-end FITC-labeled 68-mer RNA/DNA oligonucleotide was complexed with the polycation polyethylenimine (PEI) and the cationic lipids Cytofectin and GenePorter. Flow cytometry was employed to evaluate chimeraplast uptake under different conditions. Intracellular chimeraplast distribution and co-localization with endocytosis markers were assessed by confocal microscopy. Relative quantification of chimeraplast metabolism was performed by denaturing PAGE and GeneScan(trade mark) analysis. RESULTS: In airway epithelial cells, optimized chimeraplast uptake reached near 100% efficiency with the carriers tested. However, chimeraplast nuclear localization could only be achieved using PEI or Cytofectin. Chimeraplast/GenePorter lipoplexes were retained in the cytoplasm. PEI polyplexes and Cytofectin lipoplexes displayed different uptake rates and internalization mechanisms. Chimeraplast/PEI polyplexes were internalized at least partially by fluid-phase endocytosis. In contrast, phagocytosis may have contributed to the internalization process of large-sized chimeraplast/Cytofectin lipoplexes. Moreover, significant chimeraplast degradation was detected 24 h after transfection with both PEI polyplexes and Cytofectin lipoplexes, although the latter seemed to confer a higher degree of protection against nuclease degradation. CONCLUSION: Both Cytofectin and PEI are efficient for chimeraplast nuclear uptake into airway epithelial cells. However, despite the distinct structures and trafficking pathways of the corresponding complexes, none of them could prevent nuclease-mediated metabolism of the chimeric oligonucleotides. These findings should be taken into account for future investigations of chimeraplast-mediated gene repair in airway epithelial cells. PMID- 12112649 TI - DNA transfection of macaque and murine respiratory tissue is greatly enhanced by use of a nuclease inhibitor. AB - BACKGROUND: Nuclease activity present within respiratory tissues contributes to the rapid clearance of injected DNA and therefore may reduce the transfection activity of directly injected transgenes. Most gene transfer technologies transduce or transfect murine tissues more efficiently than corresponding primate tissues. Therefore, it is prudent to assess the utility of novel gene transfer strategies in both rodent and primate models before proceeding with further development. METHODS: This study analyzed the effects of ATA (a nuclease inhibitor) on the direct transfection of macaque and murine lung tissue; compared the levels of DNase activity in murine, primate, and human lung fluids; and tested the inhibitory activity of ATA on the DNase activity present in these samples. Fluorescent microspheres were used to detect areas of transfection in lung. RESULTS: Intratracheal administration of a nuclease inhibitor (ATA) with naked DNA (0.5 microg ATA/g body weight) enhanced direct transfection efficacy in macaque lung by over 86-fold and by over 54-fold in mouse lung. Hematoxylin and eosin staining showed no apparent tissue toxicity. Moreover, macaque, human, and mouse lung fluids were found to possess similar levels of DNase activity and this activity was inhibited by similar concentrations of ATA. The authors also successfully pioneered the use of carboxylate-modified microsphere tracers to identify areas of transfection and/or treatment. CONCLUSION: This work provides evidence that using direct nuclease inhibitors will enhance lung transfection and that nuclease activity is present in all lung fluids tested, which can be inhibited by the use of direct DNase inhibitors. PMID- 12112652 TI - Familial conformational diseases and dementias. AB - Familial conformational diseases occur when a mutation alters the conformation of a protein resulting in abnormal intermolecular interactions, protein aggregation, and consequent tissue damage. The molecular mechanisms of conformational disease are best understood for the serine protease inhibitor (serpin) superfamily of proteins. The serpinopathies include alpha(1)-antitrypsin (SERPINA1) deficiency and the newly characterized familial encephalopathy with neuroserpin inclusion bodies (FENIB) resulting from mutations in the neuroserpin (SERPINI1) gene. This review discusses how insights gained from the study of the serpins may be used to guide our research into other common diseases such as Alzheimer disease, Huntington disease, and Parkinson disease. PMID- 12112650 TI - High-level expression of naked DNA delivered to rat liver via tail vein injection. AB - BACKGROUND: High levels of foreign gene expression in mouse hepatocytes can be achieved by rapid tail vein injection of a large volume of a naked DNA solution, the 'hydrodynamics-based procedure'. Rats are more tolerant of the frequent phlebotomies required for monitoring blood parameters than mice, and thus are better for some biomedical research. METHODS: We tested this technique for the delivery of a therapeutic protein in normal rats, using a rat erythropoietin (Epo) expression plasmid vector, pCAGGS-Epo. RESULTS: We obtained maximal Epo expression when the DNA solution was injected in a volume of 25 ml (approximately 100 ml/kg body weight) within 15 s. We observed a dose-response relationship between serum Epo levels and the amount of injected DNA up to 800 microg. Using quantitative real-time PCR, the vector-derived Epo mRNA expression was mainly detected in the liver. When a lacZ expression plasmid was injected similarly, beta-galactosidase was exclusively detected in the liver, mainly in hepatocytes. Toxicity attributable to the technique was mild and transient, as assessed by histochemical analysis. Epo gene expression and erythropoiesis occurred with Epo gene transfer in a dose-dependent manner, and persisted for at least 12 weeks, the last time point examined. Repeated administration of the plasmid DNA also effectively led to erythropoiesis. CONCLUSIONS: These results demonstrate that gene transfer into the liver via rapid tail vein injection can easily be achieved in the rat, which is more than 10 times larger than the mouse, and has significant value for gene function analysis in rats. PMID- 12112654 TI - Hereditary non-polyposis colorectal cancer (HNPCC): phenotype-genotype correlation between patients with and without identified mutation. AB - Affected members of hereditary non-polyposis colorectal cancer (HNPCC) families develop colorectal cancer at an early age (mean 45 yr) and frequently get extracolonic cancers particularly in the uterus, urinary tract, and small intestine. They have a high risk of developing more than one primary colorectal cancer if not treated with subtotal colectomy at first operation and have more frequent right-sided colon cancers and less frequent rectum cancers, compared to patients with sporadic colorectal cancer. We have screened 31 families fulfilling the Amsterdam criteria and 54 families with a colorectal cancer clustering but not fulfilling the Amsterdam criteria for mutations in MLH1 and MSH2 by direct sequencing, and detected a mutation in 61% of the Amsterdam positive families but only in 15% of the Amsterdam negative families. Genotype-phenotype correlation was compared between 141 affected individuals with an identified mutation and 78 affected individuals from Amsterdam positive families in which a mutation was not identifiable in MLH1 or MSH2. In the affected persons with identified mutations, all expected phenotypic traits were represented, whereas affected persons in whom no mutation was detected fell into two clearly distinguishable subgroups. The minor subgroup, in which no mutation was identified, generally had the same characteristics as found in affected persons with identified mutations. The major subgroup differed significantly in clinical features and exhibited phenotypic traits similar to those found in late-onset families, including abundance of rectal cancer, few HNPCC-related cancers, lower frequency of multiple colorectal cancers, and later age at onset. Finally, for six missense mutations and one single codon deletion, the pathogenic potential was evaluated by domain localization, lod score calculation or segregation analysis when possible, and mutation-induced biochemical change. The results indicate that the majority of missense mutations are pathogenic, although further characterization by functional assays is necessary before implementation in predictive testing programs. PMID- 12112653 TI - A mutational hot spot in the KCNQ4 gene responsible for autosomal dominant hearing impairment. AB - Several different mutations in the KCNQ4 K+ channel gene are responsible for autosomal dominant nonsyndromic hearing impairment (DFNA2). Here we describe two additional families originating from Europe and Japan with a KCNQ4 missense mutation (W276S) that was previously found in one European family. We compared the disease-associated haplotype of the three W276S-bearing families using closely linked microsatellite markers and intragenic single nucleotide polymorphisms. Differences between the haplotypes were found, excluding a single founder mutation for the families. Therefore, the W276S mutation has occurred three times independently, and most likely represents a hot spot for mutation in the KCNQ4 gene. PMID- 12112656 TI - Germline mutation profile of MEN1 in multiple endocrine neoplasia type 1: search for correlation between phenotype and the functional domains of the MEN1 protein. AB - Multiple Endocrine Neoplasia type 1 (MEN1) is an autosomal dominant disease characterized by endocrine tumors of the parathyroids, the pancreatic islets, and the anterior pituitary. The MEN1 gene encodes menin, a nuclear protein interacting with JunD/AP1, Smad3, NFkappaB, and other proteins involved in transcription and cell growth regulation. Here, by exhaustive sequence analysis of 170 probands/families collected through a French clinical network, we identified 165 mutations located in coding parts of the MEN1 gene, which represent 114 distinct MEN1 germline alterations. These mutations have been included in a MEN1-locus specific database available on the world wide web together with approximately 240 germline and somatic MEN1 mutations listed from international published data. Our mutation series included 56 frameshifts, 23 nonsense, 27 missense, and eight deletion or insertion in-frame mutations. Mutations were spread over the entire coding sequence. Taken together, most missense and in-frame MEN1 genomic alterations affect one or all domains of menin interacting with JunD [codons 1-40; 139-242; 323-428], Smad3 [distal to codon 478], and NFkappaB [codons 276-479], three major effectors in transcription and cell growth regulation. No correlation has been observed between genotype and MEN1 phenotype. We suggest that the knowledge of structure and location of a specific mutation has not been useful in clinical practice for the follow-up of affected patients and asymptomatic gene carriers. Our results provide the largest series of MEN1 mutations published to date. They will be a useful tool for further studies focusing on the functional effects of missense mutations and understanding which mechanisms or pathways related to multiple menin interactions might be involved in tumorigenesis of endocrine cells. PMID- 12112655 TI - BRCA1 and BRCA2 mutations in Turkish familial and non-familial ovarian cancer patients: a high incidence of mutations in non-familial cases. AB - Ovarian cancer is a clinically important cancer in Turkey. The contribution of BRCA1 and BRCA2 to ovarian cancer in Turkish patients has not previously been described. In this study we investigated the presence of BRCA1 and BRCA2 mutations in 102 consecutively ascertained, hospital-based, ovarian cancer cases. Four out of 15 (26.7%, 95% confidence interval (CI), 7.8%-55.1%) familial cases were found to carry mutations in BRCA1. Thirteen of the 87 (14.9%, 95% CI, 7.5% 22.4%) non-familial cases had BRCA1 and BRCA2 mutations, six in BRCA1, and seven in BRCA2. We have further studied the non-familial ovarian cancer cases to determine which subgroups have a likelihood of carrying clinically important mutations. Our study shows that those Turkish ovarian cancer patients with serous histopathology harbor a high proportion of mutations (12/58, 20.7%, 95% CI, 10.3% 31.1%) compared to all non-familial cases (14.9%) regardless of pathology. Within the serous sub-group, those that were also diagnosed below age 50 have an even greater percentage of mutations (8/28, 28.6%, 95% CI, 11.8%-45.3%). Our findings demonstrate that a substantial proportion of Turkish ovarian cancer patients, both with and without a family history, carry BRCA1 and BRCA2 mutations, demonstrating the importance of BRCA1 and BRCA2 in the development of ovarian cancer in this population. PMID- 12112657 TI - Intron retention and frameshift mutations result in severe pyruvate carboxylase deficiency in two male siblings. AB - This paper describes the molecular characterization of two male siblings displaying the complex (Type B) form of pyruvate carboxylase (PC) deficiency in which severe neonatal lactic acidosis and redox abnormalities results in death within the first few weeks of life. The two male siblings were found to be compound heterozygous for a TAGG deletion at the exon15/intron15 splice site (IVS15+2-5delTAGG) and a dinucleotide deletion in exon 16 (2491-2492delGT) of the PC gene. We also demonstrate through RT-PCR and sequencing of aberrant transcripts that the IVS15+2-5delTAGG results in the retention of intron 15 during pre-mRNA splicing. In addition, both deletions are predicted to result in a frameshift to generate a premature termination codon such that the encoded mRNA could be subject to nonsense mediated decay. PMID- 12112658 TI - Large-scale determination of SNP allele frequencies in DNA pools using MALDI-TOF mass spectrometry. AB - One of the major challenges in the near future is the identification of genes that contribute to complex disorders. Large scale association studies that utilize a dense map of single nucleotide polymorphisms (SNPs) have been considered as a valuable tool for this purpose. However, genome-wide screens are limited by costs of genotyping thousands of SNPs in a large number of individuals. Here we present a pooling strategy that enables high-throughput SNP validation and determination of allele frequencies in case and control populations. Quantitative analysis of allele frequencies of SNPs in DNA pools is based on matrix-assisted laser desorption/ionization time of flight (MALDI-TOF) mass spectrometry of primer extension assays. We demonstrate the accuracy and reliability of this approach on pools of eight previously genotyped individuals with an allele frequency representation in the range of 0.1 to 0.9. The accuracy of measured allele frequencies was shown in DNA pools of 142 to 186 individuals using additional markers. Allele frequencies determined from the pooled samples deviate from the real frequencies by about 3%. The described method reduces costs and time and enables genotyping of up to thousands of samples by taking advantage of the high-throughput MALDI-TOF technology. PMID- 12112659 TI - Comparison of DNA- and RNA-based methods for detection of truncating BRCA1 mutations. AB - A number of methods are used for mutational analysis of BRCA1, a large multi-exon gene. A comparison was made of five methods to detect mutations generating premature stop codons that are predicted to result in synthesis of a truncated protein in BRCA1. These included four DNA-based methods: two-dimensional gene scanning (TDGS), denaturing high performance liquid chromatography (DHPLC), enzymatic mutation detection (EMD), and single strand conformation polymorphism analysis (SSCP) and an RNA/DNA-based protein truncation test (PTT) with and without complementary 5' sequencing. DNA and RNA samples isolated from 21 coded lymphoblastoid cell line samples were tested. These specimens had previously been analyzed by direct automated DNA sequencing, considered to be the optimum method for mutation detection. The set of 21 cell lines included 14 samples with 13 unique frameshift or nonsense mutations, three samples with two unique splice site mutations, and four samples without deleterious mutations. The present study focused on the detection of protein-truncating mutations, those that have been reported most often to be disease-causing alterations that segregate with cancer in families. PTT with complementary 5' sequencing correctly identified all 15 deleterious mutations. Not surprisingly, the DNA-based techniques did not detect a deletion of exon 22. EMD and DHPLC identified all of the mutations with the exception of the exon 22 deletion. Two mutations were initially missed by TDGS, but could be detected after slight changes in the test design, and five truncating mutations were missed by SSCP. It will continue to be important to use complementary methods for mutational analysis. PMID- 12112660 TI - Ten novel mutations in the human neurofibromatosis type 1 (NF1) gene in Italian patients. AB - The entire NF1 coding region was analyzed for mutations in a panel of 108 unrelated Italian NF1 patients. Using PTT, SSCP, and DNA sequencing, we found 10 mutations which have never been reported before. Clinical diagnosis of NF1 was established according to the NIH consensus criteria in 100 individuals, while 8 were young children with only multiple cafe-au-lait spots. We detected 46 truncated fragments, and 24 of them were fully characterized by SSCP and direct sequencing. Of the 24, 14 were known mutations (R304X, R681X, Q682X, R1306X, R1362X, R1513X, R1748X, Q1794X, R1947X, Y2264X, R2237X, 2674delA, 6789delTTAC, 2027insC). The other 10 mutations represent novel changes that contribute to the germline mutational spectrum of the NF1 gene (K810X, Q2595X, 6772delT, 7190delCT, 7331delA, 1021insTT, 3921insT, 4106insTA, 7149insC, 2033insCG / 2034delA). PTT in a large number of Italian NF1 patients supports the usefulness of this method for characterization of mutations in disorders where the responsible gene is very large and the disease-causing mutations often create a stop codon. In agreement with previous reports, no mutational hotspots within the NF1 gene were detected. PMID- 12112661 TI - Isolated sulfite oxidase deficiency: identification of 12 novel SUOX mutations in 10 patients. AB - We report twelve novel mutations in patients with isolated sulfite oxidase deficiency. The mutations are in SUOX, the gene that encodes the molybdohemoprotein sulfite oxidase. These include two frameshift mutations, a four-basepair deletion (562del4) and a single-basepair insertion (113insC), both resulting in premature termination. Nonsense mutations predicting Y343X and Q364X substitutions were identified in a homozygous state in three patients, the latter in two sibs. The remaining eight are missense mutations generating single amino acid substitutions. From the position of the substituted residues, seven of these mutations are considered to be causative of the enzyme deficiency: I201L, R211Q, G305S, R309H, K322R, Q339R, and W393R. The eighth, a C>T transition, predicts an R319C substitution, which could affect the binding of the molybdenum cofactor and thus severely reduce sulfite oxidase activity. This mutation, however, is downstream of a frameshift mutation and is therefore not the causative mutation in this individual. PMID- 12112662 TI - Biochemical and mutational analyses of the cathepsin c gene (CTSC) in three North American families with Papillon Lefevre syndrome. AB - Papillon Lefevre syndrome (PLS) is an autosomal recessive disorder characterized by palmoplantar hyperkeratosis and severe periodontitis. The disease is caused by mutations in the cathepsin C gene (CTSC) that maps to chromosome 11q14. CTSC gene mutations associated with PLS have been correlated with significantly decreased enzyme activity. Mutational analysis of the CTSC gene in three North American families segregating PLS identified four mutations, including a novel mutation p.G139R. All mutations were associated with dramatically reduced CTSC protease enzyme activity. A homozygous c.96T>G transversion resulting in a p.Y32X change was present in a Mexican PLS proband, while one Caucasian PLS proband was a compound heterozygote for the p.Y32X and p.R272P (c.815G>C) mutations. The other Caucasian PLS proband was a compound heterozygote for c.415G>A transition and c.1141delC mutations that resulted in a p.G139R and a frameshift and premature termination (p.L381fsX393), respectively. The c.415G>A was not present in more than 300 controls, suggesting it is not a CTSC polymorphism. Biochemical analysis demonstrated almost no detectable CTSC activity in leukocytes of all three probands. These mutations altered restriction enzyme sites in the highly conserved CTSC gene. Sequence analysis of CTSC exon 3 confirmed the previously reported p.T153I polymorphism in 4 of the 5 ethnically diverse populations studied. PMID- 12112663 TI - Progressive AV-block and anomalous venous return among cardiac anomalies associated with two novel missense mutations in the CSX/NKX2-5 gene. AB - Non-syndromic cardiac septation defects are common, yet the causative factors remain largely uncharacterised. Septation defects are an integral part of many syndromes, some of which are associated with chromosomal alterations. For the majority, the physiopathogenesis is believed to be multi-factorial, hindering the identification of causative factors. Ten mutations in the gene encoding the transcription factor CSX/NKX2-5 have been described in individuals with ASD and/or atrioventricular conduction defects. In addition, several other cardiac abnormalities were observed, yet the mildest forms are reminiscent of non syndromic septation defects. The CSX/NKX2-5 gene is thus a good candidate for various cardiopathies. We have collected two families with inherited predisposition to cardiac abnormalities. Some members of the families presented ASD and AV block. In both families a novel CSX/NKX2-5 mutation was identified in the homeodomain. Variable expressivity in the phenotype was observed in both families. Importantly, mutation carriers did not present any symptoms at young age. In addition, anomalous venous return, a phenotype not previously associated to CSX/NKX2-5 mutations, was observed in one of the families. We also screened the CSX/NKX2-5 gene in sporadic and familial cases of other cardiopathies. As additional mutations were not found, substitutions in CSX/NKX2-5 gene seem to be a rare cause of cardiopathies without conduction defect. PMID- 12112664 TI - Mutations in myosin VIIA (MYO7A) and usherin (USH2A) in Spanish patients with Usher syndrome types I and II, respectively. AB - Usher syndrome is an autosomal recessive disorder characterized by congenital hearing impairment and retinitis pigmentosa. Three clinical types are known (USH1, USH2 and USH3), and there is an extensive genetic heterogeneity, with at least ten genes implicated. The most frequently mutated genes are MYO7A, which causes USH1B, and usherin, which causes USH2A. We carried out a mutation analysis of these two genes in the Spanish population. Analysis of the MYO7A gene in patients from 30 USH1 families and sporadic cases identified 32% of disease alleles, with mutation Q821X being the most frequent. Most of the remaining variants are private mutations. With regard to USH2, mutation 2299delG was detected in 25% of the Spanish patients. Altogether the mutations detected in USH2A families account for 23% of the disease alleles. PMID- 12112665 TI - Erratum: Identification of five new mutations of PDS/SLC26A4 in Mediterranean families with hearing impairment. AB - Pendred syndrome is an autosomal-recessive disorder characterized by congenital sensorineural hearing loss combined with goiter. This disorder may account for up to 10% of cases of hereditary deafness. The disease gene (PDS/SLC26A4) has been mapped to chromosome 7q22-q31 and encodes a chloride-iodide transport protein. Mutations in this gene are also a cause of non-syndromic autosomal recessive hearing impairment (DFNB4). We have analyzed the PDS/SLC26A4 gene in Spanish and Italian families and we have detected five novel mutations (X781W, T132I, IVS2 2A>G, Y556H and 406del5). PMID- 12112666 TI - Frequencies of GJB2 mutations in German control individuals and patients showing sporadic non-syndromic hearing impairment. AB - Mutations in the GJB2 gene encoding the gap-junction protein connexin 26 have been identified in many patients with childhood hearing impairment (HI). One single mutation, 35delG (30delG), accounts for up to 70% of all analyzed European patients with autosomal recessive inherited HI and 10% of patients with HI of unknown origin, respectively. We screened 188 control individuals and 342 German patients with non-syndromic sporadic HI for the 35delG, compound heterozygosity and other GJB2 mutations by PCR, restriction enzyme based screening, SSCP and sequencing. In all patients, non-progressive hearing impairment varied from moderate to profound involving all frequencies. This study revealed one novel silent mutation (438C/T), three novel gene variants resulting in amino acid substitutions (K112E, T123S, K223R) and two novel HI-related mutations (I82M, 313del14). PMID- 12112667 TI - Identification of fifteen novel mutations in the SLC12A3 gene encoding the Na-Cl Co-transporter in Italian patients with Gitelman syndrome. AB - The SLC12A3 gene encodes the thiazide-sensitive Na-Cl co-transporter (NCCT) expressed in the apical membrane of the distal convoluted tubule of the kidney. Inactivating mutations of this gene are responsible for Gitelman syndrome (GS), a disorder inherited as an autosomal recessive trait. We searched for SLC12A3 gene mutations in 21 Italian patients with the clinical and biochemical features of GS (hypokalemia, hypomagnesemia, metabolic alkalosis, hypocalciuria, and the absence of nephrocalcinosis). All coding regions with their intron-exon boundaries were analyzed using PCR and SSCP techniques followed by sequencing analysis. We identified 21 different mutations evenly distributed throughout the gene without any mutation hot-spot. Fifteen are novel variants, including 12 missense mutations, one deletion, one deletion-insertion and one splice site mutation: R158Q, T163M, W172R, G316V, G374V, G463E, A464T, S615W, V677M, R852S, R958G, C985Y, 2114-2120delACCAAGT, 2144-2158delGCCTTCTACTCGGATinsTG, and 531-2A>G. PMID- 12112668 TI - Two novel mutations and a new STK11/LKB1 gene isoform in Peutz-Jeghers patients. AB - Peutz-Jeghers syndrome (PJS) is a rare autosomal dominantly inherited disorder with variable expression and incomplete penetrance characterized by mucocutaneous pigmentation, predisposition to hamartomatous intestinal polyposis, and various other neoplasms. It occurs in approximately 1 in 8,300 to 29,000 live births. In nearly 50% of patients PJS is caused by germ line mutations in the STK11/LKB1 serine/threonine kinase gene, the only kinase gene currently known to act as a tumor suppressor. We have performed a mutation search in the STK11/LKB1 gene in 8 sporadic cases and 3 PJS families using a combination of different screening techniques. We have identified four mutations, two of which I177N and the IVS2+1A >G, were previously unreported. We have also evaluated the presence of cDNA alterations by means of RT-PCR analysis and direct cDNA sequencing and have found two aberrant transcripts in a single PJS case despite the lack of any apparent genomic alteration. Finally, we report the presence of a novel STK11/LKB1 cDNA isoform observed in all the normal subjects studied as well as in the majority of the PJS patients. PMID- 12112669 TI - Comparison of sequence and structure alignments for protein domains. AB - Profile search methods based on protein domain alignments have proven to be useful tools in comparative sequence analysis. Domain alignments used by currently available search methods have been computed by sequence comparison. With the growth of the protein structure database, however, alignments of many domain pairs have also been computed by structure comparison. Here, we examine the extent to which information from these two sources agrees. We measure agreement with respect to identification of homologous regions in each protein, that is, with respect to the location of domain boundaries. We also measure agreement with respect to identification of homologous residue sites by comparing alignments and assessing the accuracy of the molecular models they predict. We find that domain alignments in publicly available collections based on sequence and structure comparison are largely consistent. However, the homologous regions identified by sequence comparison are often shorter than those identified by 3D structure comparison. In addition, when overall sequence similarity is low alignments from sequence comparison produce less accurate molecular models, suggesting that they less accurately identify homologous sites. These observations suggest that structure comparison results might be used to improve the overall accuracy of domain alignment collections and the performance of profile search methods based on them. PMID- 12112670 TI - Identification of the N-terminal functional domains of Cdk5 by molecular truncation and computer modeling. AB - Cyclin dependent kinase (Cdk) 5, an atypical member of the Cdk family, plays a fundamental role in the development of the nervous system, and may also be involved in the pathogenesis of certain neurodegenerative diseases. Further, Cdk5 is activated by the specific regulatory proteins p39, p35, or p25 rather than cyclins, and in contrast to other members of the Cdk family is not involved in the progression of the cell cycle. A three-dimensional computer model of Cdk5-p25 ATP has been generated previously [Chou et al., Biochem Biophys Res Commun 1999;259:420-428], providing a structural basis for the study of the mechanisms of Cdk5 activation. To assess the predicted ATP and p25 binding domains at the N terminal of Cdk5, two mutants of Cdk5 were prepared in which amino acids 9-15 (Delta9-15) or 9-47 (Delta9-47) were deleted. The results of these studies clearly demonstrate that an N-terminal loop and the PSSALRE helix are indispensable for Cdk5-p25 interactions, and amino acids 9-15 are necessary for ATP binding but are not involved in Cdk5-p25 interactions. Predicted models of Delta9-15 Cdk5 and Delta9-47 Cdk5 were generated, and were used to interpret the experimental data. The experimental and molecular modeling results confirm and extend specific aspects of the original predicted computer model, and may provide useful information for the design of highly selective inhibitors of Cdk5, which could be used in the treatment of certain neurodegenerative conditions. PMID- 12112672 TI - Optimally informative backbone structural propensities in proteins. AB - We use basic ideas from information theory to extract the maximum amount of structural information available in protein sequence data. From a non-redundant set of protein X-ray structures, we construct local-sequence-dependent [phi,psi] distributions that summarize the influence of local sequence on backbone conformation. These distributions, approximations of actual backbone propensities in the folded protein, have the following properties: (1) They compensate for the problem of scarce data by an optimized combination of local-sequence-dependent and single-residue specific distributions; (2) They use multi-residue information; (3) They exploit similarities in the local coding properties of amino acids by collapsing the amino acid alphabet to streamline local sequence description; (4) They are designed to contain the maximum amount of local structural information the data set allows. Our methodology is able to extract around 30 cnats of information from the protein data set out of a total 387 cnats of initial uncertainty or entropy in a finely discretized [phi,psi] dihedral angle space (18 x 18 structural states), or about 7.8%. This was achieved at the hexamer length scale; shorter as well as longer fragments produce reduced information gains. The automatic clustering of amino acids into groups, a component of the optimization procedure, reveals patterns consistent with their local coding properties. While the overall information gain from local sequence is small, there are some local sequences that have significantly narrower structural distributions than others. Distribution width varies from at least 20% less than the average overall entropy to at least 14% above. This spread is an expression of the influence of local sequence on the conformational propensities of the backbone chain. The optimal ensemble of local-sequence-specific backbone distributions produced is useful as a guide to structural predictions from sequence, as well as a tool for further explorations of the nature of the local protein code. PMID- 12112671 TI - Anti-idiotypic protein domains selected from protein A-based affibody libraries. AB - Three pairs of small protein domains showing binding behavior in analogy with anti-idiotypic antibodies have been selected using phage display technology. From an affibody protein library constructed by combinatorial variegation of the Fc binding surface of the 58 residue staphylococcal protein A (SPA)-derived domain Z, affibody variants have been selected to the parental SPA scaffold and to two earlier identified SPA-derived affibodies. One selected affibody (Z(SPA-1)) was shown to recognize each of the five domains of wild-type SPA with dissociation constants (K(D)) in the micromolar range. The binding of the Z(SPA-1) affibody to its parental structure was shown to involve the Fc binding site of SPA, while the Fab-binding site was not involved. Similarly, affibodies showing anti-idiotypic binding characteristics were also obtained when affibodies previously selected for binding to Taq DNA polymerase and human IgA, respectively, were used as targets for selections. The potential applications for these types of affinity pairs were exemplified by one-step protein recovery using affinity chromatography employing the specific interactions between the respective protein pair members. These experiments included the purification of the Z(SPA-1) affibody from a total Escherichia coli cell lysate using protein A-Sepharose, suggesting that this protein A/antiprotein A affinity pair could provide a basis for novel affinity gene fusion systems. The use of this type of small, robust, and easily expressed anti-idiotypic affibody pair for affinity technology applications, including self assembled protein networks, is discussed. PMID- 12112673 TI - Assessing equilibration and convergence in biomolecular simulations. AB - If molecular dynamics simulations are used to characterize the folding of peptides or proteins, a wide range of conformational states needs to be sampled. This study reports an analysis of peptide simulations to identify the best methods for assessing equilibration and sampling in these systems where there is significant conformational disorder. Four trajectories of a beta peptide in methanol and four trajectories of an alpha peptide in water, each of 5 ns in length, have been studied. Comparisons have also been made with two 50-ns trajectories of the beta peptide in methanol. The convergence rates of quantities that probe both the extent of conformational sampling and the local dynamical properties have been characterized. These include the numbers of hydrogen bonds populated, clusters identified, and main chain torsion angle transitions in the trajectories. The relative equilibrium rates of different quantities are found to vary significantly between the two systems studied reflecting both the differences in peptide primary structure and the different solvents used. A cluster analysis of the simulation trajectories is identified as a very effective method for judging the convergence of the simulations. This is particularly the case if the analysis includes a comparison of multiple trajectories calculated for the same system from different starting structures. PMID- 12112674 TI - Protein molecular dynamics with electrostatic force entirely determined by a single Poisson-Boltzmann calculation. AB - The electrostatic force including the intramolecular Coulombic interactions and the electrostatic contribution of solvation effect were entirely calculated by using the finite difference Poisson-Boltzmann method (FDPB), which was incorporated into the GROMOS96 force field to complete a new finite difference stochastic dynamics procedure (FDSD). Simulations were performed on an insulin dimer. Different relative dielectric constants were successively assigned to the protein interior; a value of 17 was selected as optimal for our system. The simulation data were analyzed and compared with those obtained from 500-ps molecular dynamics (MD) simulation with explicit water and a 500-ps conventional stochastic dynamics (SD) simulation without the mean solvent force. The results indicate that the FDSD method with GROMOS96 force field is suitable to study the dynamics and structure of proteins in solution if used with the optimal protein dielectric constant. PMID- 12112675 TI - Physicochemical explanation of peptide binding to HLA-A*0201 major histocompatibility complex: a three-dimensional quantitative structure-activity relationship study. AB - A three-dimensional quantitative structure-activity relationship method for the prediction of peptide binding affinities to the MHC class I molecule HLA-A*0201 was developed by applying the CoMSIA technique on a set of 266 peptides. To increase the self consistency of the initial CoMSIA model, the poorly predicted peptides were excluded from the training set in a stepwise manner and then included in the study as a test set. The final model, based on 236 peptides and considering the steric, electrostatic, hydrophobic, hydrogen bond donor, and hydrogen bond acceptor fields, had q2 = 0.683 and r2 = 0.891. The stability of this model was proven by cross-validations in two and five groups and by a bootstrap analysis of the non-cross-validated model. The residuals between the experimental pIC50 (-logIC50) values and those calculated by "leave-one-out" cross-validation were analyzed. According to the best model, 63.2% of the peptides were predicted with /residuals/ < or = 0.5 log unit; 29.3% with 1.0 < or = /residuals/ < 0.5; and 7.5% with /residuals/ > 1.0 log unit. The mean /residual/ value was 0.489. The coefficient contour maps identify the physicochemical property requirements at each position in the peptide molecule and suggest amino acid sequences for high-affinity binding to the HLA-A*0201 molecule. PMID- 12112676 TI - Atoms-in-molecules study of the genetically encoded amino acids. II. Computational study of molecular geometries. AB - The geometries of the 20 genetically encoded amino acids were optimized at the restricted Hartree-Fock level of theory using the 6-31+G* basis set. A detailed comparison showed the calculated geometries to be in excellent agreement with those determined by X-ray crystallography. The study demonstrated that the geometric parameters for the main-chain group and for the bonds and common functional groups of the side-chains exhibit a high degree of transferability among the members of this set of molecules. This geometric transferability is a necessary prerequisite for the corresponding transferability of their electron density distributions and hence of their bond and atomic properties. The transferability of the electron distributions will be demonstrated and exploited in the following paper of this series, which uses the topology of the electron density to define an atom within the quantum theory of atoms in molecules. Particular features of the geometries of the amino acids are discussed. It has been shown, for example, how the apparent anomaly of the Calpha-N bond length in a peptide being shorter than in the charged species Calpha-NH3+ is resolved when the charge separation is gauged by the differences in the charges of the Calpha and N atoms as opposed to the use of formal charges. A compilation of literature sources on experimental geometries covering each member of the 20 amino acids is presented. A set of rules for labeling the atoms and bonds, complementing the generally accepted IUPAC-IUB rules, is proposed to uniquely identify every atom and bond in the amino acids. PMID- 12112677 TI - Monte Carlo simulations of the peptide recognition at the consensus binding site of the constant fragment of human immunoglobulin G: the energy landscape analysis of a hot spot at the intermolecular interface. AB - Monte Carlo simulations of molecular recognition at the consensus binding site of the constant fragment (Fc) of human immunoglobulin G (Ig) protein have been performed to analyze structural and thermodynamic aspects of binding for the 13 residue cyclic peptide DCAWHLGELVWCT. The energy landscape analysis of a hot spot at the intermolecular interface using alanine scanning and equilibrium-simulated tempering dynamics with the simplified, knowledge-based energy function has enabled the role of the protein hot spot residues in providing the thermodynamic stability of the native structure to be determined. We have found that hydrophobic interactions between the peptide and the Met-252, Ile-253, His-433, and His-435 protein residues are critical to guarantee the thermodynamic stability of the crystallographic binding mode of the complex. Binding free energy calculations, using a molecular mechanics force field and a solvation energy model, combined with alanine scanning have been conducted to determine the energetic contribution of the protein hot spot residues in binding affinity. The conserved Asn-434, Ser-254, and Tyr-436 protein residues contribute significantly to the binding affinity of the peptide-protein complex, serving as an energetic hot spot at the intermolecular interface. The results suggest that evolutionary conserved hot spot protein residues at the intermolecular interface may be partitioned in fulfilling thermodynamic stability of the native binding mode and contributing to the binding affinity of the complex. PMID- 12112678 TI - Prediction of protein secondary structures from conformational biases. AB - We use LINUS (the "Local Independently Nucleated Units of Structure"), a procedure developed by Srinivasan and Rose, to provide a physical interpretation of and predict the secondary structures of proteins. The secondary structure type at a given site is identified by the largest conformational bias during short simulations. We examine the rate of successful prediction as a function of temperature and the interaction window. At high temperatures, there is a large propensity for the establishment of beta-strands whereas alpha-helices appear only when the temperature is lower than a certain threshold value. It is found that there exists an optimal temperature at which the correct secondary structures are predicted most accurately. We find that this temperature is close to the peak temperature of the specific heat. Changing the interaction window or carrying out longer simulations approaching equilibrium lead to little change in the optimal success rate. Our findings are in accord with the observation by Srinivasan and Rose that the secondary structures are mainly determined by local interactions and appear in the early stage of folding. PMID- 12112680 TI - Variants of 3(10)-helices in proteins. AB - An analysis of the shortest 3(10)-helices, containing three helical residues and two flanking capping residues that participate in two consecutive i + 3 --> i hydrogen bonds, shows that not all helices belong to the classic 3(10)-helix, where the three central residues adopt the right-handed helical conformation (alpha(R)). Three variants identified are: 3L10-helix with all residues in the left-handed helical region (alpha(L)), 3EL10-helix where the first residue is in the extended region followed by two residues in the alpha(L) conformation, and its mirror-image, the 3E'R10-helix. In the context of these helices, as well as the equivalent variants of alpha-helices, the length dependence of the handedness of secondary structures in protein structure is discussed. There are considerable differences in the amino acid preferences at different positions in the various types of 3(10)-helices. Each type of 3(10)-helix can be thought to be made up of an extension of a particular type of beta-turn (made up of residues i to i + 3) such that the (i + 3)th residue assumes the same conformation as the preceding residue. Distinct residue preferences at i and i + 3 positions seem to decide whether a particular stretch of four residues will be a beta-turn or a 3(10) helix in the folded structure. PMID- 12112679 TI - Prediction of protein solvent accessibility using support vector machines. AB - A Support Vector Machine learning system has been trained to predict protein solvent accessibility from the primary structure. Different kernel functions and sliding window sizes have been explored to find how they affect the prediction performance. Using a cut-off threshold of 15% that splits the dataset evenly (an equal number of exposed and buried residues), this method was able to achieve a prediction accuracy of 70.1% for single sequence input and 73.9% for multiple alignment sequence input, respectively. The prediction of three and more states of solvent accessibility was also studied and compared with other methods. The prediction accuracies are better than, or comparable to, those obtained by other methods such as neural networks, Bayesian classification, multiple linear regression, and information theory. In addition, our results further suggest that this system may be combined with other prediction methods to achieve more reliable results, and that the Support Vector Machine method is a very useful tool for biological sequence analysis. PMID- 12112681 TI - Susceptibility to proteolysis of triosephosphate isomerase from two pathogenic parasites: characterization of an enzyme with an intact and a nicked monomer. AB - The susceptibility to subtilisin of homodimeric triosephosphate isomerase from Trypanosoma brucei (TbTIM) and Trypanosoma cruzi (TcTIM) was studied. Their amino sequence and 3D structure are markedly similar. In 36 h of incubation at a molar ratio of 4 TIM per subtilisin, TcTIM underwent extensive hydrolysis, loss of activity, and large structural alterations. Under the same conditions, only about 50% of the monomers of TbTIM were cleaved in two sites. The higher sensitivity of TcTIM to subtilisin is probably due to a higher intrinsic flexibility. We isolated and characterized TbTIM that had been exposed to subtilisin. It exhibited the molecular mass of the dimer, albeit it was formed by one intact and one nicked monomer. Its k(cat) with glyceraldehyde 3-phosphate was half that of native TbTIM, with no change in K(m). The intrinsic fluorescence of nicked TbTIM was red-shifted by 5 nm. The association between subunits was not affected. The TbTIM data suggest that there are structural differences in the two monomers or that alterations of one subunit change the characteristics of the other subunit. In comparison to the action of subtilisin on TIMs from other species, the trypanosomal enzymes appear to be unique. PMID- 12112683 TI - Towards atomic resolution with crystals grown in gel: the case of thaumatin seen at room temperature. AB - One reason for introducing a gel in the crystallization medium of proteins is its ability to reduce convection in solution. This can lead to better nucleation and growth conditions, and to crystals having enhanced diffraction properties. We report here the X-ray characterization at room temperature of high-quality crystals of the intensely sweet thaumatin prepared in a sodium tartrate solution gelified with 0.15% (m/v) agarose. Using a synchrotron radiation, these crystals diffracted to a previously unachieved resolution. A diffraction dataset was collected from four crystals at a resolution of 1.2 A with a R(sym) of 3.6% and a completeness of 99%. Refinement was carried out to a final crystallographic R factor of 12.0%. The quality of the electron density map allowed for the observation of fine structural details in the protein and its solvation shell. Crystallization in gel might be used more generally to improve the quality of macromolecular crystals. Advantages provided by the gelified medium in the frame of structural studies are emphasized. PMID- 12112684 TI - Model of the alphaLbeta2 integrin I-domain/ICAM-1 DI interface suggests that subtle changes in loop orientation determine ligand specificity. AB - The interaction of the alphaLbeta2 integrin with its cellular ligand the intercellular adhesion molecule-1 (ICAM-1) is critical for the tight binding interaction between most leukocytes and the vascular endothelium before transendothelial migration to the sites of inflammation. In this article we have modeled the alphaL subunit I-domain in its active form, which was computationally docked with the D1 domain of the ICAM-1 to probe potential protein-protein interactions. The experimentally observed key interaction between the carboxylate of Glu 34 in the ICAM-1 D1 domain and the metal ion-dependent adhesion site (MIDAS) in the open alphaL I-domain was consistently reproduced by our calculations. The calculations reveal the nature of the alphaLbeta2/ICAM-1 interaction and suggest an explanation for the increased ligand-binding affinity in the "open" versus the "closed" conformation of the alphaL I-domain. A mechanism for substrate selectivity among alphaL, alphaM, and alpha2 I-domains is suggested whereby the orientation of the loops within the I-domain is critical in mediating the interaction of the Glu 34 carboxylate of ICAM-1 D1 with the MIDAS. PMID- 12112685 TI - Kinetic analysis of the interaction between the QutA and QutR transcription regulating proteins. AB - The QutR protein is a multidomain repressor protein that interacts with the QutA activator protein. Both proteins are active in the signal transduction pathway that regulates transcription of the quinic acid utilization (qut) gene cluster of the microbial eukaryote Aspergillus nidulans. In the presence of quinate, production of mRNA from the eight genes of the qut pathway is stimulated by the QutA activator protein. The QutR protein plays a key role in signal recognition and transduction, and a deletion analysis has shown that the N-terminal 88 amino acids are sufficient to inactivate QutA function in vivo. Using surface plasmon resonance we show here that the N-terminal 88 amino acids of QutR are able to bind in vitro to a region of QutA that genetic analysis has previously implicated in transcription activation. We further show that increasing the concentration of a full-length (missense) mutant QutR protein in the original mutant strain can restore its repressing function. This is interpreted to mean that the qutR mutation in this strain increases the equilibrium dissociation constant for the interaction between QutA and QutR. We propose a model in which the QutA and QutR proteins are in dynamic equilibrium between bound (transcriptionally inactive) and unbound (transcriptionally active) states. PMID- 12112686 TI - Interaction of Hsp90 with 20S proteasome: thermodynamic and kinetic characterization. AB - The proteasome and heat shock proteins have been found in the centrosome. The evidence of their copurification reported by several studies suggests that they form stable complex. In addition, Hsp90 is involved in the loading of proteasome generated antigenic peptides to the class I major histocompatibility complex. In this article, we report a detailed thermodynamic and kinetic characterization of the Hsp90-20S proteasome interaction, using a surface plasmon resonance technique. The modulation exerted by protons in solution has been investigated, and the results have been discussed, taking into account structural motifs characterizing the binding interface between the two macromolecules. PMID- 12112687 TI - Prediction of strand pairing in antiparallel and parallel beta-sheets using information theory. AB - An information theory approach was developed to predict the alignment of interacting antiparallel and parallel beta-strands. Information scores were derived for the preference of a residue on a beta-strand to be opposite a sequence of residues on an adjacent beta-strand. These scores were used to predict the interstrand register of interacting beta-strands from 10 alternative offset positions either side of the experimentally observed beta-sheet register. The amino acid sequence of an internal beta-strand can be correctly aligned with two beta-strands in a fixed position either side of the strand in 45% of antiparallel and 48% of parallel arrangements. For comparison, when another beta strand from a nonhomologous protein substitutes the internal beta-strand, the same register is predicted for only 24 and 36% of antiparallel and parallel arrangements. As expected, alignment of a single fixed strand with just a second beta-strand sequence was more difficult, and gave a correct register in 31 and 37% of antiparallel and parallel beta-pairs, respectively. These scores are 10% higher than for two randomly selected beta-strand sequences. In general, prediction accuracy was not improved by information tables that distinguished hydrogen-bonding patterns or beta-strand order. These results will contribute to predicting the arrangement of beta-strands in beta-pleated sheets and protein topology. PMID- 12112688 TI - Local energy landscape flattening: parallel hyperbolic Monte Carlo sampling of protein folding. AB - Among the major difficulties in protein structure prediction is the roughness of the energy landscape that must be searched for the global energy minimum. To address this issue, we have developed a novel Monte Carlo algorithm called parallel hyperbolic sampling (PHS) that logarithmically flattens local high energy barriers and, therefore, allows the simulation to tunnel more efficiently through energetically inaccessible regions to low-energy valleys. Here, we show the utility of this approach by applying it to the SICHO (SIde-CHain-Only) protein model. For the same CPU time, the parallel hyperbolic sampling method can identify much lower energy states and explore a larger region phase space than the commonly used replica sampling (RS) Monte Carlo method. By clustering the simulated structures obtained in the PHS implementation of the SICHO model, we can successfully predict, among a representative benchmark 65 proteins set, 50 cases in which one of the top 5 clusters have a root-mean-square deviation (RMSD) from the native structure below 6.5 A. Compared with our previous calculations that used RS as the conformational search procedure, the number of successful predictions increased by four and the CPU cost is reduced. By comparing the structure clusters produced by both PHS and RS, we find a strong correlation between the quality of predicted structures and the minimum relative RMSD (mrRMSD) of structures clusters identified by using different search engines. This mrRMSD correlation may be useful in blind prediction as an indicator of the likelihood of successful folds. PMID- 12112690 TI - Signal transduction in the photoactive yellow protein. II. Proton transfer initiates conformational changes. AB - Molecular dynamics simulation techniques, together with semiempirical PM3 calculations, have been used to investigate the effect of photoisomerization of the 4-hydroxy-cinnamic acid chromophore on the structural properties of the photoactive yellow protein (PYP) from Ectothiorodospira halophila. In this bacteria, exposure to blue light leads to a negative photoactic response. The calculations suggest that the isomerization does not directly destabilize the protein. However, because of the isomerization, a proton transfer from a glutamic acid residue (Glu46) to the phenolate oxygen atom of the chromophore becomes energetically favorable. The proton transfer initiates conformational changes within the protein, which are in turn believed to lead to signaling. PMID- 12112689 TI - Signal transduction in the photoactive yellow protein. I. Photon absorption and the isomerization of the chromophore. AB - Molecular dynamics simulation techniques together with time-dependent density functional theory calculations have been used to investigate the effect of photon absorption by a 4-hydroxy-cinnamic acid chromophore on the structural properties of the photoactive yellow protein (PYP) from Ectothiorodospira halophila. The calculations suggest that the protein not only modifies the absorption spectrum of the chromophore but also regulates the subsequent isomerization of the chromophore by stabilizing the isomerization transition state. Although signaling from PYP is thought to involve partial unfolding of the protein, the mechanical effects accompanying isomerization do not appear to directly destabilize the protein. PMID- 12112691 TI - Crystal structure of the YjeE protein from Haemophilus influenzae: a putative Atpase involved in cell wall synthesis. AB - A hypothetical protein encoded by the gene YjeE of Haemophilus influenzae was selected as part of a structural genomics project for X-ray analysis to assist with the functional assignment. The protein is considered essential to bacteria because the gene is present in virtually all bacterial genomes but not in those of archaea or eukaryotes. The amino acid sequence shows no homology to other proteins except for the presence of the Walker A motif G-X-X-X-X-G-K-T that indicates the possibility of a nucleotide-binding protein. The YjeE protein was cloned, expressed, and the crystal structure determined by the MAD method at 1.7 A resolution. The protein has a nucleotide-binding fold with a four-stranded parallel beta-sheet flanked by antiparallel beta-strands on each side. The topology of the beta-sheet is unique among P-loop proteins and has features of different families of enzymes. Crystallization of YjeE in the presence of ATP and Mg2+ resulted in the structure with ADP bound in the P-loop. The ATPase activity of YjeE was confirmed by kinetic measurements. The distribution of conserved residues suggests that the protein may work as a "molecular switch" triggered by ATP hydrolysis. The phylogenetic pattern of YjeE suggests its involvement in cell wall biosynthesis. PMID- 12112692 TI - Scoring residue conservation. AB - The importance of a residue for maintaining the structure and function of a protein can usually be inferred from how conserved it appears in a multiple sequence alignment of that protein and its homologues. A reliable metric for quantifying residue conservation is desirable. Over the last two decades many such scores have been proposed, but none has emerged as a generally accepted standard. This work surveys the range of scores that biologists, biochemists, and, more recently, bioinformatics workers have developed, and reviews the intrinsic problems associated with developing and evaluating such a score. A general formula is proposed that may be used to compare the properties of different particular conservation scores or as a measure of conservation in its own right. PMID- 12112693 TI - Flexible protein alignment and hinge detection. AB - Here we present a novel technique for the alignment of flexible proteins. The method does not require an a priori knowledge of the flexible hinge regions. The FlexProt algorithm simultaneously detects the hinge regions and aligns the rigid subparts of the molecules. Our technique is not sensitive to insertions and deletions. Numerous methods have been developed to solve rigid structural comparisons. Unlike FlexProt, all previously developed methods designed to solve the protein flexible alignment require an a priori knowledge of the hinge regions. The FlexProt method is based on 3-D pattern-matching algorithms combined with graph theoretic techniques. The algorithm is highly efficient. For example, it performs a structural comparison of a pair of proteins with 300 amino acids in about 7 s on a 400-MHz desktop PC. We provide experimental results obtained with this algorithm. First, we flexibly align pairs of proteins taken from the database of motions. These are extended by taking additional proteins from the same SCOP family. Next, we present some of the results obtained from exhaustive all-against-all flexible structural comparisons of 1329 SCOP family representatives. Our results include relatively high-scoring flexible structural alignments between the C-terminal merozoite surface protein vs. tissue factor; class II aminoacyl-tRNA synthase, histocompatibility antigen vs. neonatal FC receptor; tyrosine-protein kinase C-SRC vs. haematopoetic cell kinase (HCK); tyrosine-protein kinase C-SRC vs. titine protein (autoinhibited serine kinase domain); and tissue factor vs. hormone-binding protein. These are illustrated and discussed, showing the capabilities of this structural alignment algorithm, which allows un-predefined hinge-based motions. PMID- 12112694 TI - Free-energy simulations of the oxidation of c-terminal methionines in calmodulin. AB - In the course of aging or under conditions of oxidative stress, methionine residues of calmodulin undergo oxidation, leading to loss of biological activity of the protein. We have performed free-energy simulations of the effects of C terminal methionine side-chain oxidation on the properties of calmodulin. The simulation results indicate that oxidation should have a destabilizing effect on all three protein functional states: calcium free, calcium loaded, and with both calcium and target peptide bound. Because the different states are destabilized by different amounts, this leads to a more complex pattern in the observable effects on protein thermal stability, calcium affinity, and binding of a target peptide. The influence of oxidation on the free energy of CaM unfolding is estimated by comparing the free-energy cost of oxidizing a Met residue in a Gly Met-Gly peptide and in the protein. The protein thermal stability of the oxidized forms is lowered by a moderate amount 1-3 kcal/mol, in qualitative agreement with experimental results of 0.3 kcal/mol. The calculated changes in affinity for calcium and for the target peptide show opposing trends. Oxidation at position 144 is predicted to enhance peptide binding and weaken calcium binding, whereas oxidation at 145 weakens peptide binding and enhances affinity for calcium. The lower affinity of Met 145-oxidized calmodulin toward the target peptide correlates with experimentally observed lowering of calmodulin-activated Ca ATPase activity when oxidized calmodulin from aged rat brains is used. Thus, our simulations suggest that Met 145 is the oxidation site in the C-terminal fragment of calmodulin. The microscopic mechanism behind the calculated free energy changes appears to be a greater affinity for water of the oxidized Met side-chain relative to normal Met. Structures with Met exposed to solvent had consistently lower free energies than those with buried Met sidechains. PMID- 12112695 TI - Subdomains in the F and G helices of bacteriorhodopsin regulate the conformational transitions of the reprotonation mechanism. AB - We have performed cysteine scanning mutagenesis of the bacteriorhodopsin mutant D85N to explore the role of individual amino acids in the conformational transitions of the reprotonation mechanism. We have used whole-cell reflectance spectroscopy to evaluate the spectral properties of the 59 mutants generated during a scan of the entire F and G helices and the intervening loop region. Cys mutants were grouped into one of six phenotypes based on the spectral changes associated with their M <--> N <--> O intermediate-state transitions. Mutations that produced similar phenotypes were found to cluster in discrete molecular domains and indicate that M, N, and O possess distinct structures and that unique molecular interactions regulate the transitions between them. The distribution of these domains suggests that 1) the extramembranous loop region is involved in the stabilization of the N and M intermediates, 2) lipid-protein interactions play a key role in the accumulation of N, and 3) the amino acid side-chain interactions in the extracellular portion of the interface between helices G and A participate in the accumulation of M. PMID- 12112696 TI - The role of hydrophobic microenvironments in modulating pKa shifts in proteins. AB - The screened Coulomb potential (SCP) method, combined with a quantitative description of the microenvironments around titratable groups, based on the Hydrophobic Fragmental Constants developed by Rekker, has been applied to calculate the pK(a) values of groups embedded in extremely hydrophobic microenvironments in proteins. This type of microenvironment is not common; but constitutes a small class, where the protein's architecture has evolved to lend special properties to the embedded residue. They are of significant interest because they are frequently important in catalysis and in proton and electron transfer reactions. In the SCP treatment these special cases are treated locally and therefore do not affect the accuracy of the pK(a) values calculated for other residues in less hydrophobic environments. Here the calibration of the algorithm is extended with the help of earlier results from lysozyme and of three mutants of staphylococcal nuclease (SNase) that were specially designed to measure the energetics of ionization of titratable groups buried in extremely hydrophobic microenvironments. The calibrated algorithm was subsequently applied to a fourth mutant of SNase and then to a very large dimeric amine oxidase of 1284 residues, where 334 are titratable. The observed pK(a) shifts of the buried residues are large (up to 4.7 pK units), and all cases are well reproduced by the calculations with a root mean square error of 0.22. These results support the hypothesis that protein electrostatics can only be described correctly and self-consistently if the inherent heterogeneity of these systems is properly accounted for. PMID- 12112697 TI - Pathway heterogeneity in protein folding. AB - We generate ab initio folding pathways in two single-domain proteins, hyperthermophile variant of protein G domain (1gb4) and ubiquitin (1ubi), both presumed to be two-state folders. Both proteins are endowed with the same topology but, as shown in this work, rely to a different extent on large-scale context to find their native folds. First, we demonstrate a generic feature of two-state folders: A downsizing of structural fluctuations is achieved only when the protein reaches a stationary plateau maximizing the number of highly protected hydrogen bonds. This enables us to identify the folding nucleus and show that folding does not become expeditious until a topology is generated that is able to protect intramolecular hydrogen bonds from water attack. Pathway heterogeneity is shown to be dependent on the extent to which the protein relies on large-scale context to fold, rather than on contact order: Proteins that can only stabilize native secondary structure by packing it against scaffolding hydrophobic moieties are meant to have a heterogeneous transition-state ensemble if they are to become successful folders (otherwise, successful folding would be too fortuitous an event.) We estimate mutational Phi values as ensemble averages and deconvolute individual-route contributions to the averaged two-state kinetic picture. Our results find experimental corroboration in the well-studied chymotrypsin inhibitor (CI2), while leading to verifiable predictions for the other two study cases. PMID- 12112698 TI - Inhibition of trypsin by cowpea thionin: characterization, molecular modeling, and docking. AB - Higher plants produce several families of proteins with toxic properties, which act as defense compounds against pests and pathogens. The thionin family represents one family and comprises low molecular mass cysteine-rich proteins, usually basic and distributed in different plant tissues. Here, we report the purification and characterization of a new thionin from cowpea (Vigna unguiculata) with proteinase inhibitory activity. Cowpea thionin inhibits trypsin, but not chymotrypsin, binding with a stoichiometry of 1:1 as shown with the use of mass spectrometry. Previous annotations of thionins as proteinase inhibitors were based on their erroneous identification as homologues of Bowman Birk family inhibitors. Molecular modeling experiments were used to propose a mode of docking of cowpea thionin with trypsin. Consideration of the dynamic properties of the cowpea thionin was essential to arrive at a model with favorable interface characteristics comparable with structures of trypsin inhibitor complexes determined by X-ray crystallography. In the final model, Lys11 occupies the S1 specificity pocket of trypsin as part of a canonical style interaction. PMID- 12112699 TI - Role of stabilization centers in 4 helix bundle proteins. AB - Stabilization centers (SCs) were shown to play an important role in preventing decay of three-dimensional protein structures. These residue clusters, stabilized by cooperative long-range interactions, were proposed to serve as anchoring points for arranging secondary structure elements. In all-alpha proteins, SC elements appear less frequently than in all-beta, alpha/beta, and alpha+beta proteins suggesting that tertiary structure formation of all-alpha proteins is governed by different principles than in other protein classes. Here we analyzed the relation between the formation of stabilization centers and the inter-axial angles (Omega) of alpha-helices in 4 helix bundle proteins. In the distance range, where dipoles have dominant effect on the helix pair arrangement, those helix pairs, where residues from both helices participate in SC elements, appear as parallel more frequently than those helices where no SC elements are present. For SC containing helix pairs, the energetic difference between the parallel and anti-parallel states decreases considerably from 1.1 kcal/mol to 0.4 kcal/mol. Although the observed effect is weak for more distant helices, a competition between the SC element formation and the optimal dipole-dipole interaction of alpha-helices is proposed as a mechanism for tertiary structure formation in 4 helix bundle proteins. The SC-forming potential of different arrangements as well as the pitfalls of the SC definition are also discussed. PMID- 12112700 TI - Molecular dynamics simulations of a double unit cell in a protein crystal: volume relaxation at constant pressure and correlation of motions between the two unit cells. AB - Eight molecular dynamics simulations of a double crystal unit cell of ubiquitin were performed to investigate the effects of simulating at constant pressure and of simulating two unit cells compared to a single unit cell. To examine the influence of different simulation conditions, the constant-pressure and constant volume simulations were each performed with and without counterions and using two different treatments of the long-range electrostatic interactions (lattice-sum and reaction-field methods). The constant-pressure simulations were analyzed in terms of unit cell deformation and accompanying protein deformations. Energetic and structural properties of the proteins in the simulations of the double unit cell were compared to the results of previously reported one-unit-cell simulations. Correlation between the two unit cells was also investigated based on relative translational and rotational movements of the proteins and on dipole fluctuations. The box in the constant-pressure simulations is found to deform slowly to reach convergence only after 5-10 ns. This deformation does not result from a distortion in the structure of the proteins but rather from changes in protein packing within the unit cell. The results of the double-unit-cell simulations are closely similar to the results of the single-unit-cell simulations, and little motional correlation is found between the two unit cells. PMID- 12112701 TI - Entropic benefit of a cross-link in protein association. AB - We introduce a method to estimate the loss of configurational entropy upon insertion of a cross-link to a dimeric system. First, a clear distinction is established between the loss of entropy upon tethering and binding, two quantities that are often considered to be equivalent. By comparing the probability distribution of the center-to-center distances for untethered and cross-linked versions, we are able to calculate the loss of translational entropy upon cross-linking. The distribution function for the untethered helices is calculated from the probability that a given helix is closer to its partner than to all other helices, the "Nearest Neighbor" method. This method requires no assumptions about the nature of the solvent, and hence resolves difficulties normally associated with calculations for systems in liquids. Analysis of the restriction of angular freedom upon tethering indicates that the loss of rotational entropy is negligible. The method is applied in the context of the folding of a ten turn helical coiled coil with the tether modeled as a Gaussian chain or a flexible amino acid chain. After correcting for loop closure entropy in the docked state, we estimate the introduction of a six-residue tether in the coiled coil results in an effective concentration of the chain to be about 4 or 100 mM, depending upon whether the helices are denatured or pre-folded prior to their association. Thus, tethering results in significant stabilization for systems with millimolar or stronger dissociation constants. PMID- 12112702 TI - A novel fold recognition method using composite predicted secondary structures. AB - In this work, we introduce a new method for fold recognition using composite secondary structures assembled from different secondary structure prediction servers for a given target sequence. An automatic, complete, and robust way of finding all possible combinations of predicted secondary structure segments (SSS) for the target sequence and clustering them into a few flexible clusters, each containing patterns with the same number of SSS, is developed. This program then takes two steps in choosing plausible homologues: (i) a SSS-based alignment excludes impossible templates whose SSS patterns are very different from any of those of the target; (ii) a residue-based alignment selects good structural templates based on sequence similarity and secondary structure similarity between the target and only those templates left in the first stage. The secondary structure of each residue in the target is selected from one of the predictions to find the best match with the template. Truncation is applied to a target where different predictions vary. In most cases, a target is also divided into N terminal and C-terminal fragments, each of which is used as a separate subsequence. Our program was tested on the fold recognition targets from CASP3 with known PDB codes and some available targets from CASP4. The results are compared with a structural homologue list for each target produced by the CE program (Shindyalov and Bourne, Protein Eng 1998;11:739-747). The program successfully locates homologues with high Z-score and low root-mean-score deviation within the top 30-50 predictions in the overwhelming majority of cases. PMID- 12112703 TI - Performance evaluation of a new algorithm for the detection of remote homologs with sequence comparison. AB - A detailed analysis of the performance of hybrid, a new sequence alignment algorithm developed by Yu and coworkers that combines Smith Waterman local dynamic programming with a local version of the maximum-likelihood approach, was made to access the applicability of this algorithm to the detection of distant homologs by sequence comparison. We analyzed the statistics of hybrid with a set of nonhomologous protein sequences from the SCOP database and found that the statistics of the scores from hybrid algorithm follows an Extreme Value Distribution with lambda approximately 1, as previously shown by Yu et al. for the case of artificially generated sequences. Local dynamic programming was compared to the hybrid algorithm by using two different test data sets of distant homologs from the PFAM and COGs protein sequence databases. The studies were made with several score functions in current use including OPTIMA, a new score function originally developed to detect remote homologs with the Smith Waterman algorithm. We found OPTIMA to be the best score function for both both dynamic programming and the hybrid algorithms. The ability of dynamic programming to discriminate between homologs and nonhomologs in the two sets of distantly related sequences is slightly better than that of hybrid algorithm. The advantage of producing accurate score statistics with only a few simulations may overcome the small differences in performance and make this new algorithm suitable for detection of homologs in conjunction with a wide range of score functions and gap penalties. PMID- 12112704 TI - Swaps in protein sequences. AB - An important question in protein evolution is to what extent proteins may have undergone swaps (switches of domain or fragment order) during evolution. Such events might have occurred in several forms: Swaps of short fragments, swaps of structural and functional motifs, or recombination of domains in multidomain proteins. This question is important for the theoretical understanding of the evolution of proteins, and has practical implications for using swaps as a design tool in protein engineering. In order to analyze the question systematically, we conducted a large scale survey of possible swaps and permutations among all pairs of protein from the Swissport database. A swap is defined as a specific kind of sequence mutation between two proteins in which two fragments that appear in both sequences have different relative order in the two sequences. For example, aXbYc and dYeXf are defined as a swap, where X and Y represent sequence fragments that switched their order. Identifying such swaps is difficult using standard sequence comparison packages. One of the main problems in the analysis stems from the fact that many sequences contain repeats, which may be identified as false-positive swaps. We have used two different approaches to detect pairs of proteins with swaps. The first approach is based on the predefined list of domains in Pfam. We identified all the proteins that share at least two domains and analyzed their relative order, looking for pairs in which the order of these domains was switched. We designed an algorithm to distinguish between real swaps and duplications. In the second approach, we used Blast to detect pairs of proteins that share several fragments. Then, we used an automatic procedure to select pairs that are likely to contain swaps. Those pairs were analyzed visually, using a graphical tool, to eliminate duplications. Combining these approaches, about 140 different cases of swaps in the Swissprot database were found (after eliminating multiple pairs within the same family). Some of the cases have been described in the literature, but many are novel examples. Although each new example identified may be interesting to analyze, our main conclusion is that cases of swaps are rare in protein evolution. This observation is at odds with the common view that proteins are very modular to the point that modules (e.g., domains) can be shuffled between proteins with minimal constraints. Our study suggests that sequential constraints, i.e., the relative order between domains, are highly conserved. PMID- 12112705 TI - Empirical relationships between protein structure and carboxyl pKa values in proteins. AB - Relationships between protein structure and ionization of carboxyl groups were investigated in 24 proteins of known structure and for which 115 aspartate and 97 glutamate pK(a) values are known. Mean pK(a) values for aspartates and glutamates are < or = 3.4 (+/-1.0) and 4.1 (+/-0.8), respectively. For aspartates, mean pK(a) values are 3.9 (+/-1.0) and 3.1 (+/-0.9) in acidic (pI < 5) and basic (pI > 8) proteins, respectively, while mean pK(a) values for glutamates are approximately 4.2 for acidic and basic proteins. Burial of carboxyl groups leads to dispersion in pK(a) values: pK(a) values for solvent-exposed groups show narrow distributions while values for buried groups range from < 2 to 6.7. Calculated electrostatic potentials at the carboxyl groups show modest correlations with experimental pK(a) values and these correlations are not improved by including simple surface-area-based terms to account for the effects of desolvation. Mean aspartate pK(a) values decrease with increasing numbers of hydrogen bonds but this is not observed at glutamates. Only 10 pK(a) values are > 5.5 and most are found in active sites or ligand-binding sites. These carboxyl groups are buried and usually accept no more than one hydrogen bond. Aspartates and glutamates at the N-termini of helices have mean pK(a) values of 2.8 (+/-0.5) and 3.4 (+/-0.6), respectively, about 0.6 units less than the overall mean values. PMID- 12112706 TI - Distinguishing native conformations of proteins from decoys with an effective free energy estimator based on the OPLS all-atom force field and the Surface Generalized Born solvent model. AB - Protein decoy data sets provide a benchmark for testing scoring functions designed for fold recognition and protein homology modeling problems. It is commonly believed that statistical potentials based on reduced atomic models are better able to discriminate native-like from misfolded decoys than scoring functions based on more detailed molecular mechanics models. Recent benchmark tests on small data sets, however, suggest otherwise. In this work, we report the results of extensive decoy detection tests using an effective free energy function based on the OPLS all-atom (OPLS-AA) force field and the Surface Generalized Born (SGB) model for the solvent electrostatic effects. The OPLS AA/SGB effective free energy is used as a scoring function to detect native protein folds among a total of 48,832 decoys for 32 different proteins from Park and Levitt's 4-state-reduced, Levitt's local-minima, Baker's ROSETTA all-atom, and Skolnick's decoy sets. Solvent electrostatic effects are included through the Surface Generalized Born (SGB) model. All structures are locally minimized without restraints. From an analysis of the individual energy components of the OPLS-AA/SGB energy function for the native and the best-ranked decoy, it is determined that a balance of the terms of the potential is responsible for the minimized energies that most successfully distinguish the native from the misfolded conformations. Different combinations of individual energy terms provide less discrimination than the total energy. The results are consistent with observations that all-atom molecular potentials coupled with intermediate level solvent dielectric models are competitive with knowledge-based potentials for decoy detection and protein modeling problems such as fold recognition and homology modeling. PMID- 12112707 TI - Crystal structure of Bacillus subtilis ioli shows endonuclase IV fold with altered Zn binding. PMID- 12112708 TI - Crystal structure of Escherichia coli EC1530, a glyoxylate induced protein YgbM. PMID- 12112709 TI - X-ray structure of Saccharomyces cerevisiae homologous mitochondrial matrix factor 1 (Hmf1). PMID- 12112710 TI - Is N2O3 the main nitrosating intermediate in aerated nitric oxide (NO) solutions in vivo? If so, where, when, and which one? AB - The widespread opinion that N(2)O(3) as a product of NO oxidation is the only nitros(yl)ating agent under aerobic conditions is based on experiments in homogeneous buffered water solutions. In vivo NO is oxidized in heterogeneous media and this opinion is not correct. The equilibrium in the system being dependent on temperature and DeltaG((sol)) for NO, NO(2), isomers of both N(2)O(3), and N(2)O(4). For polar solvents including water, DeltaG((sol)) for N(2)O(3) is high enough, and a stationary concentration of N(2)O(3) in the mixture with other oxides is sufficient to guarantee the hydrolysis of N(2)O(3) to nitrite. In heterogeneous media, the mixture contains solvates NO(2(sol)), N(2)O(3(sol)), and N(2)O(4(sol)) at stationary nonequilibrium concentrations. As far as DeltaG((sol)) is decreased in heterogeneous mixtures with low polar solvents and/or at increased temperatures, the equilibrium in such a system shifts to NO(2). Although NO(2) is a reactive free radical, it almost does not react with water. In contrast, the reaction with most functional protein groups efficiently proceeds by a radical type with the formation of nitrite and new radicals (X) further stabilized in various forms. Therefore, the ratio of the nitrosylated and nitrated products yields depends on actual concentrations of all NO(x). PMID- 12112711 TI - Xenobiotic export pumps, endothelin signaling, and tubular nephrotoxicants--a case of molecular hijacking. AB - This article is a review on recent studies in intact renal proximal tubules that link tubular nephrotoxicants with endothelin (ET) regulation of xenobiotic export pump function. The data show that transport on p-glycoprotein and Mrp2 decreases rapidly when ET signals through an ET(B) receptor, NO synthase (NOS), and protein kinase C (PKC). Surprisingly, nephrotoxicants, such as radiocontrast agents, aminoglycoside antibiotics, and heavy metal salts, "hijack" this signaling pathway, causing ET release from the tubules, hormone binding to its receptor, activation of NOS and PKC, and reduced xenobiotic transport. These findings suggest a new common mechanism by which nephrotoxicants may act to disrupt renal tubular function. PMID- 12112712 TI - Induction of olfactory mucosal and liver metabolism of lidocaine by 2,3,7,8 tetrachlorodibenzo-p-dioxin. AB - Formulation of drugs for administration via the nasal cavity is becoming increasingly common. It is of potential clinical relevance to determine whether intranasal drug administration itself, or exposure to other xenobiotics, can modulate the levels and/or activity of nasal mucosal metabolic enzymes, thereby affecting the metabolism and disposition of the drug. In these studies, we examined changes in several of the major metabolic enzymes in nasal epithelial tissues upon exposure to the environmental contaminant 2,3,7,8-tetrachlorodibenzo p-dioxin (TCDD), as well as the impact of these changes on the metabolism of a model intranasally administered drug, lidocaine. Results of these studies show that TCDD can induce multiple metabolic enzymes in the olfactory mucosa and that the pattern of induction in the olfactory mucosa does not necessarily parallel that which occurs in the liver. Further, increases in enzyme levels noted by Western blot analysis were associated with increased activities of several nasal mucosal enzymes as well as with enhanced conversion of lidocaine to its major metabolite, monoethyl glycine xylidide (MEGX). These results demonstrate that environmental exposures can influence the levels and activity of nasal mucosal enzymes and impact the pharmacology of drugs administered via the nasal route. PMID- 12112713 TI - Formation of nitrosothiols from gaseous nitric oxide at pH 7.4. AB - Nitric oxide (NO) is generated in biological systems and plays important roles as a regulatory molecule. Its ability to bind to haem iron is well known. Moreover, it may lose an electron, forming the nitrosonium ion, involved in the synthesis of S-nitrosothiols (SNOs). It has been suggested that S-nitrosohaemoglobin (-SNO Hb) and low molecular weight SNOs may act as reservoirs of NO. SNOs are formed in vitro, at strongly acidic pH values; however, the mechanism of their formation at neutral pH values is still debated. In this paper we report the anaerobic formation of SNOs (both high- and low-molecular weight) from low concentrations of NO at pH 7.4, provided Hb is also present. We propose a reaction mechanism entailing the participation of Fehaem in the formation of NO(+) and the transfer of NO(+) either to Cysbeta(93) of Hb or to glutathione; we show that this reaction also occurs in human RBCs. PMID- 12112714 TI - Comparison between modifications of lens proteins resulted from glycation with methylglyoxal, glyoxal, ascorbic acid, and fructose. AB - Cataract is generally associated with the breakdown of the lens microarchitecture. Age-dependent chemical modifications and cross-linking of proteins are the major pathways for development of lens opacity. The specific alterations in lens proteins caused by glycation with four carbonyl metabolites, fructose, methylglyoxal, glyoxal, and ascorbic acid, were investigated. Decrease in intensity of tryptophan related fluorescence and level of reduced protein sulfhydryl groups, parameters that are indicative for changes in protein conformation, were observed after reaction with all studied carbonyl compounds. Protein carbonyl content, an index for oxidative damage to proteins, was strongly enhanced in methylglyoxal-treated proteins. Cross-linking of glycated proteins was confirmed by polyacrylamide electrophoresis. alpha-Oxoaldehydes were the most reactive in protein aggregation. They also formed specific chromophores absorbing UV light above 300 nm. Significant loss in lactate dehydrogenase activity resulted from incubation with methylglyoxal, followed by glyoxal and ascorbic acid. The results obtained showed that alterations in lens proteins do not follow the specific reactivity of studied carbonyl compounds. Despite the similarity in chemical structures of alpha-oxoaldehydes and ascorbic acid degradation products, they cause specific alterations in lens protein structure with different biological consequences. PMID- 12112716 TI - Volume cutting for virtual petrous bone surgery. AB - A profound knowledge of anatomy and surgical landmarks of the temporal bone is a basic necessity for any otologic surgeon. Because this knowledge, so far, has been mostly taught by limited temporal bone drilling courses, our objective was to create a system for virtual petrous bone surgery that allows the realistic simulation of specific laterobasal surgical approaches. A major requirement was the development of an interactive drill-like tool, together with a new technique for realistic visualization of simulated cut surfaces. The system is based on a volumetric, high-resolution model of the temporal bone, derived from CT. Interactive volume cutting methods using a new multivolume scheme have been developed. In this scheme, cut regions are modeled independently in additional data volumes using voxelization techniques. The voxelization is adapted to successive cutting operations as needed for the simulation of a drill-like tool. A new visualization technique was developed for artifact-free rendering of sharp edges, as formed by the intersection of a cut and an object surface. The new multivolume visualization technique allows high-quality visualization of interactively generated cut surfaces. This is a necessity for a realistic simulation of petrous bone surgery. Our system therefore facilitates comprehension of the complex morphology, and enables the recognition of surgical landmarks, which is most important if injury to delicate organs (e.g., the facial nerve or auditory ossicles) is to be avoided. The system for virtual petrous bone surgery allows the simulation of specific surgical approaches with high-quality visualization. The user can learn about the complex three-dimensional anatomy of the temporal bone from the viewpoint of a real otosurgical procedure. PMID- 12112715 TI - Deforming a preoperative volume to represent the intraoperative scene. AB - Soft-tissue deformation can be a problem if a preoperative modality is used to help guide a surgical or interventional procedure. We present a method that can warp a preoperative CT image to represent the intraoperative scene shown by an interventional fluoroscopy image. The method is a novel combination of a 2D-3D image registration algorithm and a deformation algorithm that allows rigid bodies to be incorporated into a nonlinear deformation based on radial basis functions. The 2D-3D registration algorithm is used to obtain information on relative vertebral movements between preoperative and intraoperative images. The deformation algorithm uses this information to warp the preoperative image to represent the intraoperative scene more accurately. Images from an aortic stenting procedure were used. The observed deformation in our experiment was 5 degrees flexion and 5 mm lengthening of the lumbar spine over a distance of four vertebrae. The vertebral positions in the warped CT volume represent the intraoperative scene more accurately than in the preoperative CT volume. Although we had no gold standard with which to assess the registration accuracy of soft tissue structures, the position of such structures within the warped CT volume appeared visually realistic. PMID- 12112717 TI - Improved localization of coronary stents using layer decomposition. AB - Accurate placement and expansion of coronary stents is hindered by the fact that most stents are only slightly radiopaque and hence difficult to see in typical coronary X-ray images. We propose a new technique for improved image guidance of multiple coronary stent deployment using layer decomposition of coronary X-ray image sequences. We hypothesize that layer decomposition can improve the accuracy of localization of the end of a deployed stent. Layer decomposition is used to obtain good quality images of a stent in vitro. The resultant background subtracted stent images are embedded into other cine X-ray image sequences to form a database of simulated image sequences. For each simulated sequence, the position of the stent edge is estimated from raw and layer-decomposed images using a small region of the original layer image as a template. Layer decomposition reduced median position errors in 33 of 47 image sequences (70%), including 16 of 18 sequences in which the position errors for raw and layer images differed by 5.0 pixels (0.5 mm) or more. Layer decomposition significantly reduces errors in determination of stent edge location in simulated cine X-ray image sequences. PMID- 12112718 TI - The application accuracy of the NeuroMate robot--A quantitative comparison with frameless and frame-based surgical localization systems. AB - The NeuroMatetrade mark robot system (Integrated Surgical Systems, Davis, CA) is a commercially available, image-guided, robotic-assisted system used for stereotactic procedures in neurosurgery. In this article, we present a quantitative comparison of the application accuracy of the NeuroMate with that of standard frame-based and frameless stereotactic techniques. The article discusses a five-way application accuracy comparison study. The variables of our comparison and their mean errors are as follows: (1) with the robot in a frame-based configuration, the RMS was 0.86 +/- 0.32 mm; (2) with the robot in the frameless configuration, the RMS was 1.95 +/- 0.44 mm; (3) in a standard stereotactic (ZD) frame-based approach, the RMS was 1.17 +/- 0.25 mm; (4) with an infrared tracking system using the frame for fiducial registration, the RMS was 1.47 +/- 0.45 mm; (5) with an infrared tracking system using screw markers for registration, the RMS was 0.68 +/- 0.26 mm. The study was performed with 2-mm sections of CT scans. These results show that the application accuracy of the frame-based NeuroMate robot is comparable to that of standard localizing systems, whether they are frame-based or infrared tracked. PMID- 12112719 TI - Reduction in variability of acetabular cup abduction using computer assisted surgery: a prospective and randomized study. AB - Optimal orientation of the acetabular component of a total hip prosthesis is an important factor in determining the early and long-term result of a total hip arthroplasty (THA). Conventional positioning of the cup component is usually done using a free-hand method, or with the help of a mechanical acetabular alignment guide. However, these methods have proven to be inaccurate, and a great variation in orientation of the cup is found postoperatively. In this study, we wished to determine if the variability of the abduction angle of acetabular cups could be reduced with the use of computer navigation. The abduction angles of the acetabular components of three groups of 50 THAs were assessed. In the first group, a free-hand method was used to position the cup component. This group was operated in the period before we started using computer navigation for hip surgery. In the second group, CT-based computer navigation was used to plan and help position the cup. The third group consisted of 50 THA cases in which a free hand method was used to position the cup, although these procedures were performed in the period after we had begun using the Computer Assisted Surgery (CAS) system. The variability in cup abduction angle was assessed in all three groups and compared. There was a significant reduction in variability in the CAS group compared to the first group. There was also a reduction in variability in the CAS group compared to the third group, although this was not statistically significant. It is concluded that the use of computer navigation helped the surgeon to place the cup component with less variability of the abduction angle, and, more importantly, we found that no cups were placed in the more extreme positions (outliers). PMID- 12112720 TI - Decreased chaos of heart rate time series in children of patients with panic disorder. AB - This study examined the differences of heart rate variability measures between children of parents with panic disorder and children of healthy controls using linear as well as nonlinear techniques. Supine and standing heart rate variability indices were measured in all children using power spectral analysis and a measure of chaos, the largest Lyapunov exponent (LLE) of heart rate time series. No significant differences emerged between the children of panic disorder parents and children of normal controls on any of the spectral heart rate variability measures. However, children of patients with panic disorder had significantly lower LLE of heart rate time series in supine posture, suggesting a relative decrease of cardiac vagal function in this group of children. This suggests a possible heritable effect of certain measures of heart rate variability, as previous studies showed decreased heart rate variability in patients with panic disorder using spectral as well as nonlinear techniques. Recent evidence also suggests that some of these nonlinear measures are superior or of additional value to the traditional time and frequency domain measures of heart rate variability to predict serious ventricular arrhythmias and sudden death. PMID- 12112721 TI - Posttraumatic stress disorder and the structure of common mental disorders. AB - Krueger [1999: Arch Gen Psychiatry 56:921-926] identified a three-factor structure of psychopathology that explained the covariation or grouping of common mental disorders found in the U.S. National Comorbidity Survey (NCS) [Kessler et al., 1994: Arch Gen Psychiatry 51:8-19]. These three fundamental groupings included an externalizing disorders factor and two internalizing disorders factors (anxious-misery and fear). We extended this research through the examination of additional data from a large subsample of the NCS (n=5,877) that contained diagnostic information on posttraumatic stress disorder (PTSD). Factor analytic findings revealed that PTSD showed no affinity with the fear factor defined by panic and phobic disorders, and instead loaded on the anxious-misery factor defined primarily by mood disorders. An identical pattern of results emerged for both lifetime PTSD and 12-month PTSD prevalence figures. Implications of these findings for the classification of PTSD and research on its etiology are briefly discussed. PMID- 12112722 TI - Instrument to assess depersonalization-derealization in panic disorder. AB - There is a long history of scholarly interest on depersonalization-derealization (DD) and its role in clinical anxiety, but there is a paucity of appropriate assessment instruments available. Our objective was to develop and evaluate a self-report measure of DD for use with clinically anxious patients. Panic disorder patients (n=169) were surveyed about DD experiences and provided data on a new item pool for psychometric development. DD episodes were common and a 28 item Depersonalization-Derealization Inventory was found to possess good reliability and validity. DD appears to be prevalent and clinically relevant in panic disorder. Continued study of DD is warranted and may be facilitated by the availability of a suitable instrument with promising psychometric properties. A 12-item version of the instrument may be appropriate as a brief screen. PMID- 12112723 TI - Personality disorder comorbidity in panic disorder patients with or without current major depression. AB - To investigate the relationship between current or past major depressive disorder (MDD) on comorbid personality disorders in patients with panic disorder, we compared the comorbidity of personality disorders using the Structured Clinical Interview for DSM-III-R personality disorders (SCID-II) in 34 panic disorder patients with current MDD (current-MD group), 21 with a history of MDD but not current MDD (past-MD group), and 32 without lifetime MDD comorbidity (non-MD group). With regard to personality disorders, patients in the current-MD group met criteria for at least one personality disorder significantly more often than patients in the past-MD group or the non-MD group (82.4% vs. 52.4% and 56.3%, respectively). The current-MD group showed statistically significantly more borderline, dependent, and obsessive-compulsive personality disorders than the past-MD group or non-MD group. With stepwise regression analyses, number of MDD episodes emerged as an indicator of the comorbidity of cluster C personality disorder and any personality disorders. Future studies should determine whether aggressive treatment of comorbid personality disorders improves the outcome (e.g., lowers the likelihood of comorbid MDD) of patients with panic disorder. PMID- 12112724 TI - Reliability of the self-report version of the panic disorder severity scale. AB - The interview-administered Panic Disorder Severity Scale (PDSS) has been demonstrated to be a reliable and valid measurement of panic disorder severity. The purpose of this paper is to report on the reliability and usefulness of the self-report form (PDSS-SR). The PDSS and PDSS-SR were administered to 108 psychiatric outpatients with and without panic disorder. The internal consistency of the instruments was analyzed with Cronbach's alpha. Test-retest reliability was assessed on 25 of these subjects by using intraclass correlation coefficient (ICC). In addition, decrease in panic symptoms was measured after cognitive behavioral treatment (CBT) treatment on 27 subjects. Cronbach's alpha was.917 for PDSS-SR and.923 for the PDSS. The PDSS-SR had good test-retest reliability (ICC=0.81) and was sensitive to change with treatment, with a mean decrease of 7.3 (S.D.=5.1). The self-report version of the PDSS is a reliable format, which could be useful in clinical and research settings. PMID- 12112725 TI - Social phobia and the recall of autobiographical memories. AB - Although cognitive theories of anxiety suggest that anxious individuals are characterized by the biased tendency to recall negative experiences with perceived threat, few empirical studies have confirmed this notion. To investigate personal memories associated with threatening experiences, individuals with socio phobia (n=16) and nonanxious individuals (n=17) completed an autobiographical memory-cueing procedure. Participants were presented with 15 social threat words and 15 neutral words, and they recorded the first specific memory that came to mind. Autobiographical memories were coded for specificity and affective tone. There were no group differences in the extent to which participants retrieved specific memories. A greater number of memories cued by social threat words were characterized by negative affect in the social phobia group than in the nonanxious group. However, examination of the means suggests that this effect was small and most likely not of practical significance. It is suggested that memory biases toward threat are not a prominent part of cognitive symptoms that characterize individuals with social phobia. PMID- 12112726 TI - Panic attacks in the differential diagnosis and treatment of resistant epilepsy. AB - The authors present four patients displaying panic disorder and a history of epileptic seizures to illustrate difficulties regarding differential diagnosis between epileptic seizures and panic attacks. The cases describe the aversive properties of epileptic seizures, the role of visual seizure-triggering stimuli as phobic cues, and the effectiveness and safety of clomipramine treatment of panic attacks as an adjunct to concurrent antiepileptic medication. PMID- 12112727 TI - Introduction: Rett syndrome. PMID- 12112728 TI - Clinical manifestations and stages of Rett syndrome. AB - The presentation and clinical diagnosis of Rett syndrome at various ages and stages are reviewed. In addition to the classical form, variability in phenotype between different atypical Rett forms is given. Obligatory, supportive, and differential diagnostic criteria are summarized. Long-term follow-up findings in ageing Rett women are addressed. PMID- 12112729 TI - Neurophysiology of Rett syndrome. AB - Neurophysiological evaluations have been widely applied in the study of Rett syndrome (RS) to provide information concerning the developmental aspects of RS; the character and extent of involvement of the central, peripheral, and autonomic nervous system pathways; and evaluation of the clinical symptomatology of RS. The electroencephalogram (EEG) is invariably abnormal and shows characteristic, though not diagnostic, changes: loss of expected developmental features; the appearance of focal, multifocal, and generalized epileptiform abnormalities; and the occurrence of rhythmic slow (theta) activity primarily in the frontal-central regions. Epileptic seizures are reported to occur frequently in RS, and partial and generalized seizures may be experienced by RS girls. However, many events presumed to be seizures have no EEG correlate during video-EEG monitoring, suggesting the possibility of a nonepileptic mechanism. Such monitoring may be necessary to determine appropriate use of antiepileptic drugs. Evoked potentials typically demonstrate intact peripheral auditory and visual pathways and suggest dysfunction of central or "higher" cortical pathways. Somatosensory-evoked potentials may be characterized by "giant" responses, suggesting cortical hyperexcitability. An increased incidence of long QT intervals during electrocardiographic recordings and diminished heart-rate variability, suggesting impairment of the autonomic nervous system, are described in RS. With the discovery of the genetic basis of RS, neurophysiological studies will provide parameters for phenotype-genotype correlations and characterization of animal models. PMID- 12112730 TI - Neuropathology of Rett syndrome. AB - Rett Syndrome is unlike any other pediatric neurologic disease, and its clinical pathologic correlation can not be defined with standard histology techniques. Based on hypotheses suggested by careful clinical observations, the nervous system of the Rett child has been explored utilizing morphometry, golgi preparations, computerized tomography, magnetic resonance imaging, chemistry, immunocytochemistry, autoradiography, and molecular biologic techniques. From these many perspectives we conclude that Rett syndrome is not a typical degenerative disorder, storage disorder, nor the result of gross malformation, infectious or neoplastic processes. There remain regions of the brain that have not been studied in detail but the available data suggest that the neuropathology of Rett syndrome can be summarized as follows: the Rett brain is small for the age and the height of the patient; it does not become progressively smaller over three to four decades; it has small dendritic trees in pyramidal neurons of layers III and V in selected lobes (frontal, motor, and temporal); it has small neurons with an increased neuronal packing density; it has an immature expression of microtubular protein-2 and cyclooxygenase; it exhibits a changing pattern of neurotransmitter receptors with an apparent reduction in many neurotransmitters, possibly contributing to some symptomatology. A mutation in Mecp2 causes this unique disorder of brain development. Neuronal mosaicism for normal and mutated Mecp2 produces a consistent phenotype in the classic female patient and a small brain with some preserved islands of function, but with an inability to support hand use and speech. This paper summarizes our current observations about neuropathology of Rett syndrome. MRDD Research Reviews 2002;8:72-76. PMID- 12112731 TI - Looking from the inside out: a parent's perspective. PMID- 12112732 TI - Genetic basis of Rett syndrome. AB - The origin of Rett syndrome has long been debated, but several observations have suggested an X-linked dominant inheritance pattern. We and others have pursued an exclusion-mapping strategy using DNA from a small number of familial Rett syndrome cases. This work resulted in the narrowing of the region likely to harbor the mutated gene to Xq27.3-Xqter. After systematic exclusion of several candidate genes, we discovered mutations in MECP2, the gene that encodes the transcriptional repressor, methyl-CpG-binding protein 2. Since then, nonsense, missense, or frameshift mutations have been found in at least 80% of girls affected with classic Rett syndrome. Sixty-four percent of mutations are recurrent C > T transitions at eight CpG dinucleotides mutation hotspots, while the C-terminal region of the gene is prone to recurrent multinucleotide deletions (11%). Most mutations are predicted to result in total or partial loss of function of MeCP2. There is no clear correlation between the type and position of the mutation and the phenotypic features of classic and variant Rett syndrome patients, and XCI appears to be a major determinant of phenotypic severity. Further research focuses on the pathogenic consequences of these mutations along the hypothesis of loss of transcriptional repression of a small number of genes that are essential for neuronal function in the maturing brain. PMID- 12112733 TI - MeCP2 and other methyl-CpG binding proteins. AB - DNA methylation is an epigenetic modification that is implicated in transcriptional silencing. Recently, it has become increasingly clear that both correct levels and proper interpretation of methylation are important factors for normal development and function of the human organism. One example is the neurological disorder Rett syndrome (RTT), which affects approximately one in 10,000 girls. RTT is caused by mutations in MeCP2, a protein that was identified by its ability to bind specifically to CpG-methylated DNA. Furthermore, MeCP2 represses transcription in a methylation-dependent manner, and it is the founding member of the family of methyl-CpG binding domain (MBD) proteins. PMID- 12112735 TI - Associations between MeCP2 mutations, X-chromosome inactivation, and phenotype. AB - Rett syndrome is a neurodevelopmental disorder of early postnatal brain growth in girls. Patients show a normal neonatal period with subsequent developmental regression and a loss of acquired skills (communication and motor skills), deceleration of head growth, and development of typical hand stereotypies. Recent studies have shown that mutations in the X-linked methyl CpG binding protein 2 gene (MeCP2) cause most typical cases of Rett syndrome. The MeCP2 gene encodes a protein that binds methylated cytosine residues of CpG dinucleotides and mediates, with histone deacetylases and transcriptional repressors, the transcription "silencing" of other genes. Girls with Rett syndrome exhibit mosaic expression for the MeCP2 defect at the cellular level, with most patients showing random X-inactivation and approximately equal numbers of cells expressing the normal MeCP2 gene and the mutated MeCP2 gene. In rare cases, females with a MeCP2 mutation escape phenotypic expression of the disorder because of nonrandom X inactivation and the preferential inactivation of the mutated MeCP2 allele. Nonrandom patterns of X-inactivation may also contribute to the clinical variability often seen in girls with Rett syndrome. The spectrum of clinical phenotype caused by MeCP2 mutations is wide, including milder "preserved speech" variants, the severe congenital Rett variant, and a subset of X-linked recessive mental retardation in boys. Studies have shown that atypical and classical Rett syndrome can caused by the same MeCP2 mutations, indicating clinical phenotype is variable even among girls with the same MeCP2 mutation. The relationship between type of MeCP2 mutation, X-inactivation status, and clinical phenotype of Rett syndrome is complex and likely involves other environmental and polygenic modifiers. PMID- 12112734 TI - The phenotypic consequences of MECP2 mutations extend beyond Rett syndrome. AB - Although MECP2 was initially identified as the causative gene in classic Rett syndrome (RTT), the gene has now been implicated in several phenotypes that extend well beyond the clinically defined disorder. MECP2 mutations have been found in people with various disorders, including neonatal onset encephalopathy, X-linked recessive mental retardation (MRX), classic and atypical RTT, autism, and Angelman syndrome, as well as mildly affected females and normal carrier females. To make matters more complex, in approximately 20% of classic sporadic RTT cases and more than 50% of affected sister pairs, no mutation in MECP2 has been found. X-chromosome inactivation patterns can clearly affect the phenotypic expression in females, while the effect of the type and position of the mutation is more apparent in the broader phenotype than in RTT. Both males and females are at risk, although an excess of paternally derived mutations are found in most cases of classic RTT. Thus, because of the range of disparate phenotypes, the gene may account for a relatively large portion of mental retardation in the population. PMID- 12112736 TI - Clinical trials and treatment prospects. AB - Prospects for definitive therapeutic intervention for Rett syndrome (RS) have been elevated by the discovery of mutations in the methyl-CpG-binding protein 2 gene (MECP2) in more than 80% of females meeting clinical criteria for this disorder. As such, a review of previous clinical trials, descriptions of the status of clinical management for the prominent medical problems of RS, and a preview of an ongoing clinical trial conducted jointly at the Baylor College of Medicine and the University of Alabama at Birmingham are presented. The conduct of controlled clinical trials requires adherence to diagnostic criteria for RS; stratification by age, stage, and presence of MECP2 mutations; and use of clearly defined outcome measures. Previous clinical trials in RS have been conducted with L-carnitine, the ketogenic diet, and the opiate antagonist, naltrexone. The L carnitine and naltrexone trials were double blind, placebo-controlled and us ed the motor behavioral analysis described in this review. All failed to provide evidence of dramatic improvements in the clinical features of RS. Specific recommendations are presented for clinical management of growth failure, breathing irregularities, seizures, ambulation, scoliosis, gastrointestinal function, self abuse, and habilitation/education although systematic evaluations of each in the context of RS have not been conducted. The only ongoing trial involves dietary supplementation with folate and betaine and is based on the finding that gene expression of some alleles of the agouti gene could be altered by dietary methyl supplementation. The availability of animal models expressing mutations in MECP2 should enhance the evaluation of innovative therapies for RS. PMID- 12112737 TI - Crossing the phase boundary to study protein dynamics and function: combination of amide hydrogen exchange in solution and ion fragmentation in the gas phase. AB - Protein dynamics are the key to understanding their behavior. The static protein structure alone in most cases is insufficient to describe the vast array of complex functions they perform in vivo. Until recently there were relatively few techniques available to investigate the dynamic nature of these proteins. Mass spectrometry has recently emerged as a powerful biophysical method, capable of providing both structural and dynamic information. By utilizing the labile nature of amide hydrogens as a marker of the backbone dynamics in solution, combined with gas-phase dissociation techniques, we now have a high-resolution tool to locate these exchanging hydrogens within the sequence of the protein and to probe the functional importance of its structural elements. In this paper we describe several applications of these methodologies to illustrate the importance of dynamics to the biological functions of proteins. PMID- 12112738 TI - High-performance liquid chromatographic/tandem mass spectrometric identification of the phototransformation products of tebuconazole on titanium dioxide. AB - Tebuconazole is a widely used fungicide. The formation of by-products on irradiated titanium dioxide as a photocatalyst was evaluated. Several species derived from tebuconazole degradation were identified and characterized by HPLC/MS(n). A pattern of reactions accounting for the observed intermediates is proposed. Different parallel pathways are operating (and through these pathways the transformation of the molecule proceeds), leading to a wide range of intermediate compounds. All these molecules are more hydrophylic than tebuconazole. The main steps involved are (1) the hydroxylation of the molecule with the formation of three species having [M + H](+) 324; the hydroxylation occurs on the C-1 carbon and on the aromatic ring in the two ortho-positions; (2) the cleavage of a C--C bond with the release of the tert-butyl moiety and the formation of a species having m/z 250; analogously to step 1, also on this species a further hydroxylation reaction occurs; (3) through the loss of the triazole moiety with the formation of a structure with m/z 257. PMID- 12112739 TI - Identification of methyl salicylate as the principal volatile component in the methanol extract of root bark of Securidaca longepedunculata Fers. AB - Securidaca longepedunculata Fers (Polygalaceae) is commonly used as a medicine in many parts of Africa and shows promise for protecting stored grain against insect pests. Analysis of a methanol extract of the root bark by gas chromatography linked to mass spectrometry (GC/MS) showed a major component accounting for over 90% of the volatile material. This was identified as methyl 2-hydroxybenzoate (methyl salicylate) by comparison of the GC retention times and mass spectrum with those of synthetic standards. This conflicts with an earlier report that the major component is methyl 4-hydroxybenzoate. Two minor components had mass spectra characteristic of 2-hydroxybenzoate esters and were identified as methyl 2-hydroxy-6-methoxybenzoate and its benzyl analogue, again conflicting with an earlier report. PMID- 12112740 TI - Identification of phenylbutyrylglutamine, a new metabolite of phenylbutyrate metabolism in humans. AB - Phenylbutyrate is used in humans for treating inborn errors of ureagenesis, certain forms of cancer, cystic fibrosis and thalassemia. The known metabolism of phenylbutyrate leads to phenylacetylglutamine, which is excreted in urine. We have identified phenylbutyrylglutamine as a new metabolite of phenylbutyrate in human plasma and urine. We describe the synthesis of phenylbutyrylglutamine and its assay by gas chromatography/mass spectrometry as a tert-butyldimethylsilyl or methyl derivative, using standards of [(2)H(5)]phenylbutyrylglutamine and phenylpropionylglutamine. After administration of phenylbutyrate to normal humans, the cumulative urinary excretion of phenylacetate, phenylbutyrate, phenylacetylglutamine and phenylbutyrylglutamine amounts to about half of the dose of phenylbutyrate. Thus, additional metabolites of phenylbutyrate are yet to be identified. PMID- 12112741 TI - Silylation of an OH-terminated self-assembled monolayer surface through low energy collisions of ions: a novel route to synthesis and patterning of surfaces. AB - Using a multi-sector ion-surface scattering mass spectrometer, reagent ions of the general form SiR(3) (+) were mass and energy selected and then made to collide with a hydroxy-terminated self-assembled monolayer (HO-SAM) surface at energies of approximately 15 eV. These ion-surface interactions result in covalent transformation of the terminal hydroxy groups at the surface into the corresponding silyl ethers due to Si--O bond formation. The modified surface was characterized in situ by chemical sputtering, a low-energy ion-surface scattering experiment. These data indicate that the ion-surface reactions have high yields (i.e. surface reactants converted to products). Surface reactions with Si(OCH(3))(3) (+), followed by chemical sputtering using CF(3) (+), yielded the reagent ion, Si(OCH(3))(3) (+), and several of its fragments. Other sputtered ions, namely SiH(OCH(3))(2)OH(2) (+) and SiH(2)(OCH(3))OH(2) (+), contain the newly formed Si--O bond and provide direct evidence for the covalent modification reaction. Chemical sputtering of modified surfaces, performed using CF(3) (+), was evaluated over a range of collision energies. The results showed that the energy transferred to the sputtered ions, as measured by their extent of fragmentation in the scattered ion mass spectra, was essentially independent of the collision energy of the projectile, thus pointing to the occurrence of reactive sputtering.A set of silyl cations, including SiBr(3) (+), Si(C(2)H(3))(3) (+) and Si(CH(3))(2)F(+), were similarly used to modify the HO SAM surface at low collision energies. A reaction mechanism consisting of direct electrophilic attack by the cationic projectiles is supported by evidence of increased reactivity for these reagent ions with increases in the calculated positive charge at the electron-deficient silicon atom of each of these cations. In a sequential set of reactions, 12 eV deuterated trimethylsilyl cations, Si(CD(3))(3) (+), were used first as the reagent ions to modify covalently a HO SAM surface. Subsequently, 70 eV SiCl(3) (+) ions were used to modify the surface further. In addition to yielding sputtered ions of the modified surface, SiCl(3) (+) reacted with both modified and unmodified groups on the surface, giving rise not only to such scattered product ions as SiCl(2)OH(+) and SiCl(2)H(+), but also to SiCl(2)CD(3) (+) and SiCl(2)D(+). This result demonstrates that selective, multi-step reactions can be performed at a surface through low-energy ionic collisions. Such processes are potentially useful for the construction of novel surfaces from a monolayer substrate and for chemical patterning of surfaces with functional groups. PMID- 12112742 TI - Gas-phase ion chemistry in germane-propane and germane-propene mixtures. AB - Germane-propane and germane-propene gaseous mixtures were studied by ion trap mass spectrometry. Variations of ion abundances observed under different partial pressure ratios and mechanisms of ion-molecule reactions elucidated by multiple isolation steps are reported. In addition, the rate constants for the main reactions were experimentally determined and compared with the collisional rate constants to obtain the reaction efficiencies. The yield of ions containing both Ge and C atoms is higher in the germane-propene than in the germane-propane system. In the former mixture, chain propagation takes place starting from germane ions reacting with propene and proceeds with the formation of clusters such as Ge(2)C(4)H(n) (+) and Ge(3)CH(n) (+). PMID- 12112743 TI - Study of non-covalent complexation between catechin derivatives and peptides by electrospray ionization mass spectrometry. AB - The recent development of electrospray ionization mass spectrometry (ESI-MS) has allowed its use to study molecular interactions driven by non-covalent forces. ESI-MS has been used to detect non-covalent complexes between proteins and metals, ligands and peptides and interactions involving DNA, RNA, oligonucleotides and drugs. Surprisingly, the study of the interaction between polyphenolic molecules and peptides/proteins is still an area where ESI-MS has not benefited. With regard to the important influence of these interactions in the biological and food domains, ESI-MS was applied to the detection and the characterization of soluble polyphenol-peptide complexes formed in model solution. The ability to observe and monitor the weak interactions involved in such macromolecular complexation phenomena was demonstrated for monomeric and dimeric flavonoid molecules (catechin-derived compounds) largely encountered in plants and plant derived products. Intact non-covalent polyphenol-peptide complexes were observed by ESI-MS using different experimental conditions. Utilizing mild ESI interface conditions allowed the detection of 1 : 1 polyphenol peptide complexes in all tested solutions and 2 : 1 complexes for the dimers and galloylated polyphenols (flavanols). These results show that there is a preferential interaction between polymerized and/or galloylated polyphenols and peptide compared with that between monomeric polyphenols and peptides. Thus, ESI MS shows potential for the study of small polyphenolic molecule-peptide interactions and determination of stoichiometry. PMID- 12112744 TI - Complexes of bivalent metal cations in electrospray mass spectra of common organic compounds. AB - In the standard electrospray ionization mass spectra of many common, low molecular mass organic compounds dissolved in methanol, peaks corresponding to ions with formula [3M + Met](2+) (M = organic molecule, Met = bivalent metal cation) are observed, sometimes with significant abundances. The most common are ions containing Mg(2+), Ca(2+) and Fe(2+). Their presence can be easily rationalized on the basis of typical organic reaction work-up procedures. The formation of [3M + Met](2+) ions has been studied using N-FMOC-proline methyl ester as a model organic ligand and Mg(2+), Ca(2+), Sr(2+), Ba(2+), Fe(2+), Ni(2+), Mn(2+), Co(2+) and Zn(2+) chlorides or acetates as the sources of bivalent cation. It was found that all ions studied form [3M + Met](2+) complexes with N-FMOC-proline methyl ester, some of them at very low concentrations. Transition metal cations generally show higher complexation activity in comparison with alkaline earth metal cations. They are also more specific in the formation of [3M + Met](2+) complexes. In the case of alkaline earth metal cations [2M + Met](2+) and [4M + Met](2+) complex ions are also observed. It has been found that [3M + Met](2+) complex ions undergo specific fragmentation at relatively low energy, yielding fluorenylmethyl cation as a major product. [M + Na](+) ions are much more stable and their fragmentation is not as specific. PMID- 12112745 TI - Characterization of silicate monomer with sodium, calcium and strontium but not with lithium and magnesium ions by fast atom bombardment mass spectrometry. AB - Fast atom bombardment mass spectrometry (FABMS) was applied to the direct detection of silica species dissolved in LiCl, NaCl, MgCl(2), CaCl(2) and SrCl(2) solutions in order to investigate its dissolution process in solution. Several species of dissolved silicate complexes in the solution were directly detected by FABMS. The peak intensities of [SiO(2)(OH)(2)Na](-), [SiO(3)(OH)Ca](-) and [SiO(3)(OH)Sr](-) increased with increasing concentrations of NaCl, CaCl(2) and SrCl(2), whereas the peak intensities of [SiO(2)(OH)(2)Li](-) and [SiO(3)(OH)Mg]( ) did not increase with increasing concentrations of LiCl and MgCl(2). These results indicte that silicate and cation bind in the solution not after but before ionization. The isotope pattern of Sr(2+) confirmed the existence of the silicate-Sr complex not only with increase of the concentration of silica but also the mass numbers of Sr. The silicate complexes formed with Na(+), Ca(2+) and Sr(2+) showed high stability in chloride solution. This is in good accordance with the fact that Na(+), Ca(2+) and Sr(2+) accelerate the dissolution of silica to form complexes during solution equilibrium. Considering that the stability constant was examined and reported in other papers, this new findings that Mg(2+) does not form a complex with silicic acid (Si(OH)(4)) is very important. PMID- 12112746 TI - Application of 1-alkylamines to a liquid chromatographic/turbo ionspray tandem mass spectrometric method for quantifying metabolites of a new bone anabolic agent, TAK-778, in human serum. AB - We investigated the application of alkylamines, as additives to the mobile phase, to a quantification method for the metabolites, M-III and M-IV, of TAK-778, which is a new bone anabolic agent, in human serum using liquid chromatography/tandem mass spectrometry (LC/MS/MS). Prior to setting up the analytical method, we found that 1-alkylamines co-existing with M-III and M-IV in the turbo ionsprayed solution formed 1-alkylammonium adduct molecules of these metabolites during the ionization process, and the abundance of the adduct ions was considerably higher than that of protonated molecules ([M + H](+)s) of these metabolites. Based on these findings, we investigated a variety of 1-alkylamines and their spiked concentrations in the mobile phase for LC/MS/MS analysis to obtain higher sensitivities for the quantification of these metabolites. After these examinations, we found that 1-hexylamine at a final concentration of 0.05 mmol l( 1) was the most suitable additive for the mobile phase, and set the selected reaction monitoring (SRM) ions for the 1-hexylammonium adduct molecule and [M + H](+), allowing about a fivefold gain in the SRM chromatographic peak compared with that without 1-hexylamine. The adduct ion was considered to be formed by interaction between the amino group of 1-hexylamine and the phosphoryl group of M III and M-IV. The internal standard (I.S.) used was deuterated M-III for each metabolite. The analytes and I.S. were extracted with diethyl ether from serum samples at neutral pH and injected into the LC/MS/MS system with a turbo ionspray interface. The limit of quantification for both analytes was 0.5 ng ml(-1) when 0.1 ml of serum was used, and the calibration curves were linear in the range 0.5 100 ng ml(-1). The method was precise; the intra- and inter-day precisions of the method were not more than 5.6%. The accuracy of the method was good, with deviations between added and calculated concentrations of M-III and M-IV being typically within 16.6%. This method provided reliable pharmacokinetic data for M III and M-IV after the intramuscular administration of TAK-778 sustained-release formulation in humans. PMID- 12112748 TI - Differentiation between isomeric acacetin-6-C-(6"-O-malonyl)glucoside and acacetin-8-C-(6"-O-malonyl)glucoside by using low-energy CID mass spectra. PMID- 12112747 TI - What do matrix-assisted laser desorption/ionization mass spectra reveal about ionization mechanisms? AB - We present ion mass spectra obtained by matrix-assisted laser desorption/ionization for analytes of approximately 1000 Da across the largest range of wavelengths and pulse durations to date. The matrix used in all cases was 2,5-dihydroxybenzoic acid. Based on the data and fundamentals of laser material interactions, we evaluated the plausibility of discriminating among such ion formation mechanisms as multiphoton ionization and excited-state ionization from mass spectra alone. Some previously proposed mechanisms appear unlikely for the matrix-analyte systems that we studied, casting doubt on their general applicability. Moreover, although analysis of mass spectra can apparently exclude certain mechanisms, it cannot establish which of several competing mechanisms is actually operative. This is particularly true with respect to variations in pulse duration and wavelength. PMID- 12112749 TI - Current literature in mass spectrometry. PMID- 12112750 TI - Obituary. Istvan Cornides (1920-1999). PMID- 12112751 TI - Environmental applications of membrane introduction mass spectrometry. AB - The purpose of this review is to highlight the versatility of membrane introduction mass spectrometry (MIMS) in environmental applications, summarize the measurements of environmental volatile organic compounds (VOCs) accomplished using MIMS, present developments in the detection of semi-volatile organic compounds (SVOCs) and forecast possible future directions of MIMS in environmental applications. PMID- 12112752 TI - Quantitative analysis of memantine in human plasma by gas chromatography/negative ion chemical ionization/mass spectrometry. AB - A sensitive and specific method for the determination of memantine in human plasma is presented. Memantine was extracted from plasma and derivatized to the pentafluorobenzoyl derivative in a one-step procedure avoiding any sample concentration steps. Amantadine was used as an internal standard. The compounds were measured by gas chromatography/negative ion chemical ionization mass spectrometry without any further processing. Using this detection mode, the fragment ions at m/z 353 and 325 were obtained at high relative abundance. Calibration graphs were linear over the range 0.117-30 ng ml(-1). At the limit of quantification (LOQ), the inter-assay precision was 2.00% and the intra-assay variability was 3.22%. The accuracy at the LOQ showed deviations of -1.42% (intra assay) and -2.47% (inter-assay). The method is rugged, rapid and robust and was applied to the batch determination of memantine during pharmacokinetic profiling of the drug. PMID- 12112753 TI - Lipid A structure of Pseudoalteromonas haloplanktis TAC 125: use of electrospray ionization tandem mass spectrometry for the determination of fatty acid distribution. AB - The use of electrospray Ionization (ESI) tandem mass spectrometry (MS/MS) for the structural determination of the lipid A components of the psychrophilic bacterium Pseudoalteromonas haloplanktis TAC 125 is reported. The lipid A contains the classical bisphosphorylated beta-(1' --> 6)-linked D-glucosamine disaccharide with 3-hydroxydodecanoyl residues (12 : 0 (3-OH)) linked both as esters and amides to 2', 3' (distal glucosamine) and 2, 3 positions (proximal glucosamine) of the sugar backbone. The hydroxyl of 12 : 0 (3-OH) fatty acid linked at the 3' position is esterified by a dodecanoyl residue (12 : 0). In addition to the pentaacyl component, a minor tetraacyl lipid A, lacking the acyl residue at position 3, was also found in the lipid A fraction. The advantage of this MS technique for the investigation of the intra-ring fragmentation, which is useful for the determination of fatty acyl residue distribution on each glucosamine unit, is emphasized. PMID- 12112754 TI - Low-abundance plasma and urinary [(15)N]urea enrichments analyzed by gas chromatography/combustion/isotope ratio mass spectrometry. AB - We report a method for determining plasma und urinary [(15)N]urea enrichments in an abundance range between 0.37 and 0.52 (15)N atom% (0-0.15 atom% excess (APE) (15)N) using a dimethylaminomethylene derivative. Compared with conventional off line preparation and (15)N analysis of urea, this method requires only small sample volumes (0.5 ml of plasma and 25 microl of urine). The (15)N/(14)N ratio of urea derivatives was measured by gas chromatography/combustion/isotope ratio mass spectrometry (GC/C/IRMS). Two peaks were separated; one was identified by gas chromatography/mass spectrometry (GC/MS) as the complete derivatized urea. Calibration of the complete urea derivative was performed by linear regression of enrichment values of known standard mixtures. Replicate standard (6-465 per thousand delta(15)N) derivatizations showed a relative standard deviation ranging from 0.1 to 7%. In order to test the feasibility of the method, human subjects and rats ingested a single meal containing either 200 mg of [(15)N]glycine (95 AP (15)N) or 0.4 mg of [(15)N]-alpha-lysine (95 AP (15)N), respectively. Urine and plasma were collected at hourly intervals over 7 h after the meal intake. After (15)N glycine intake, maximum urinary urea (15)N enrichments were 330 and 430 per thousand delta(15)N (0.12 and 0.16 APE (15)N) measured by GC/C/IRMS, whereas plasma [(15)N]glycine enrichments were 2.5 and 3.3 APE (15)N in the two human subjects 2 h after the meal. (15)N enrichments of total urine and urine samples devoid of ammonia were higher enriched than urinary [(15)N]urea measured by GC/C/IRMS, reflecting the presence of other urinary N-containing substances (e.g. creatinine). In rats plasma urea (15)N enrichments were 15-20 times higher than those in urinary urea (10-20 per thousand delta(15)N). The different [(15)N]urea enrichments observed after ingestion of [(15)N]-labeled glycine and lysine confirm known differences in the metabolism of these amino acids. PMID- 12112755 TI - Electrospray ionization mass spectrometry of ginsenosides. AB - Ginsenosides R(b1), R(b2), R(c), R(d), R(e), R(f), R(g1), R(g2) and F(11) were studied systematically by electrospray ionization mass spectrometry in positive- and negative-ion modes with a mobile-phase additive, ammonium acetate. In general, ion sensitivities for the ginsenosides were greater in the negative-ion mode, but more structural information on the ginsenosides was obtained in the positive-ion mode. [M + H](+), [M + NH(4)](+), [M + Na](+) and [M + K](+) ions were observed for all of the ginsenosides studied, with the exception of R(f) and F(11), for which [M + NH(4)](+) ions were not observed. The signal intensities of [M + H](+), [M + NH(4)](+), [M + Na](+) and [M + K](+) ions varied with the cone voltage. The highest signal intensities for [M + H](+) and [M + NH(4)](+) ions were obtained at low cone voltage (15-30 V), whereas those for [M + Na](+) and [M + K](+) ions were obtained at relatively high cone voltage (70-90 V). Collision induced dissociation yielded characteristic positively charged fragment ions at m/z 407, 425 and 443 for (20S)-protopanaxadiol, m/z 405, 423 and 441 for (20S) protopanaxatriol and m/z 421, 439, 457 and 475 for (24R)-pseudoginsenoside F(11). Ginsenoside types were identified by these characteristic ions and the charged saccharide groups. Glycosidic bond cleavage and elimination of H(2)O were the two major fragmentation pathways observed in the product ion mass spectra of [M + H](+) and [M + NH(4)](+). In the product ion mass spectra of [M - H](-), the major fragmentation route observed was glycosidic bond cleavage. Adduct ions [M + 2AcO + Na](-), [M + AcO](-), [M - CH(2)O + AcO](-), [M + 2AcO](2-), [M - H + AcO](2-) and [M - 2H](2-) were observed at low cone voltage (15-30 V) only. PMID- 12112756 TI - Monitoring of in vitro deamidation of gliadin peptic fragment by mass spectrometry may reflect one of the molecular mechanisms taking place in celiac disease development. AB - Celiac disease, a chronic disorder of the small intestine, is caused by dietary gluten and is characterized by villous atrophy and local inflammation associated with infiltration of B and T lymphocytes and/or macrophages into the intestinal wall. In genetically predisposed individuals, the infiltrating cells are activated by gluten, gliadin and their proteolytic fragments and produce chemokines, cytokines and reactive radicals. The sequence of one of the macrophage-stimulatory gliadin peptic fragment was determined by mass spectrometry (MS) as VSFQQPQQQYPSSQ. The role of tissue transglutaminase (tTG) in innate immunity stimulation was studied by mass spectrometric monitoring of sequence changes in this active peptide. Two sites of glutamine deamidation in this peptide were localized by high-resolution scanning in MS/MS mode in an ion trap. A single deamidation in the parent peptide led to the complete loss of its stimulatory effect on macrophages. PMID- 12112757 TI - Protein identification with Teflon as matrix-assisted laser desorption/ionization sample support. AB - Protein identification is a critical step in proteomics, and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS) plays an important role in that identification. Polytetrafluoroethylene (Teflon) was tested as a new MALDI sample support to improve protein identification. The tryptic peptides obtained from a model protein were bound to the surface of a modified MALDI sample holder via the hydrophobic interactions that occur between the Teflon surface and the peptide ion-pairs, and the affinity of alpha-cyano-4 hydroxycinnamic acid for the peptides. During that surface-binding step, the peptide mixture was also desalted and concentrated. A greater number of matched peptides and a larger sequence coverage were obtained for the proteins when Teflon was used as the sample support compared with conventional sample preparation methods and a stainless-steel surface. In addition, the characterization of a small amount of protein was improved with Teflon. Nine silver-stained protein spots obtained from 2-D gel of a human cerebrospinal fluid (CSF) proteome were identified by this method. Among the nine protein spots, peptide 6:c3c fragment and procollagen c-proteinase enhancer were not annotated in any published 2-D map of human CSF. A Teflon MALDI sample support is a low cost, simple, and effective method that can be used to improve the quality of the MALDI mass spectrum of a complex tryptic peptide mixture, and to achieve a higher level of reliability and success in protein identification. PMID- 12112758 TI - Identification of anhydroecgonine methyl ester N-oxide, a new metabolite of anhydroecgonine methyl ester, using electrospray mass spectrometry. AB - Cocaine is transformed into hepatotoxic metabolites through oxidative pathways. For anhydroecgonine methyl ester (AEME), the main constituent in crack smoke, the oxidative metabolism has not been studied. Therefore, incubation of AEME with rat liver microsomes was performed and a metabolite of AEME, anhydroecgonine methyl ester N-oxide (AEMENO), was identified. The chemical structure of this new metabolite was confirmed by synthesis and by comparative interpretation of electrospray multiple-stage mass spectra, which were obtained in the positive ion mode. This metabolite was also detected in whole blood, serum and urine samples from crack users. The application of liquid chromatography/electrospray mass spectrometry or nanoelectrospray mass spectrometry was necessary because AEMENO is susceptible to thermal degradation during gas chromatographic/mass spectrometric analysis. This study demonstrated that AEMENO is produced by rat hepatic microsomal metabolism in vitro and is present in body fluids from crack users. PMID- 12112759 TI - Structure, energetics and reactivity of ternary complexes of amino acids with Cu(II) and 2,2'-bipyridine by density functional theory. A combination of radical induced and spin-remote fragmentations. AB - The structures, electron distributions and dissociation energies of gas-phase ternary complexes of Cu(II) with 2,2'-bipyridine and leucine, isoleucine and lysine were addressed by density functional theory using the hybrid B3LYP functional, effective core potentials and the 6-31 + G(d) and LANL2DZ basis sets. The calculations confirm the previously suggested structures, in which amino acid carboxylates coordinate to the Cu atom by the carboxylate and alpha-amino groups in square-planar complexes. The dissociation energies for consecutive eliminations of CO(2) and alkyl radicals from isomeric singly charged complexes of leucine and isoleucine correlate with the ion relative abundances observed in collisionally activated dissociation mass spectra. Doubly charged lysine complexes show extremely low dissociation energies that are consistent with the <1 eV center-of-mass collision energies that were used in previous CAD studies. The calculated charge and spin densities point to radical-induced dissociations of singly charged complexes with an open-electron shell. In contrast, the unpaired electron is virtually inert in doubly charged, open-shell complexes that undergo charge-induced, spin-remote dissociations in the amino acid residues. PMID- 12112760 TI - Negative-ion electrospray mass spectra of carbon dioxide-protected N-heterocyclic anions. PMID- 12112761 TI - Current literature in mass spectrometry. PMID- 12112762 TI - Motor cortex activation is related to force of squeezing. AB - Primate studies have demonstrated that motor cortex neurons show increased activity with increased force of movement. In humans, this relationship has received little study during a power grip such as squeezing, and has previously only been evaluated across a narrow range of forces. Functional MRI was performed in eight healthy subjects who alternated between rest and right hand squeezing at one of three force levels. During scanning, motor performances were recorded using a dynamometer. At each force level, activation volume was measured within left sensorimotor cortex, right sensorimotor cortex, and a midline supplementary motor area. In left sensorimotor cortex, % signal change was also assessed. The range of force generated across the three force levels varied from 4.9 N to 276 N. In left sensorimotor cortex, activation volume increased significantly with greater force. The % signal change also increased with greater force and correlated closely with activation volume. In supplementary motor area, activation volume increased significantly with increasing force, but with greater intersubject variability. In right sensorimotor cortex, a trend for larger activation volumes with greater force did not reach significance. The laterality index, an expression of the relative degree of contralateral vs. ipsilateral sensorimotor cortex activation, did not change across the three force levels. Increased force of squeezing is associated with increased contralateral sensorimotor cortex and supplementary motor area activation. This relationship was found across the full spectrum of forces that the human hand is capable of generating. Use of a valid, reliable method for assessing motor behavior during functional MRI may be important to clinical applications. PMID- 12112763 TI - Functional brain activation during arithmetic processing in females with fragile X Syndrome is related to FMR1 protein expression. AB - Arithmetic processing deficits in persons with fragile X Syndrome (fraX), the most common heritable cause of mental retardation, are well known. In this study, we characterize the neural underpinnings of these performance deficits using functional MRI. Given that a single gene defect (FMR1) is known to be responsible for this disorder, we also assess whether brain activation in arithmetic processing areas is related to amount of FMR1 protein expression (FMRP). Subjects included 16 females with fraX, and 16 female age-matched controls. Subjects viewed arithmetic equations with two (1 + 3 = 4) or three (2 + 3 - 1 = 5) operands, and were asked to judge whether the results were correct or not. Subjects with fraX showed significant impairment in behavioral performance on the 3-operand but not the 2-operand arithmetic equations. Significant brain activation was observed bilaterally in the prefrontal and parietal cortices for unaffected subjects, and bilateral prefrontal and left angular gyrus for subjects with fraX, for both trial types. Subjects with fraX exhibited less overall activation than did unaffected subjects in both types of trials; and, unlike the unaffected group, did not show increased extent of activation in association with greater task difficulty. During the 3-operand trials, activation in bilateral prefrontal and motor/premotor, and left supramarginal and angular gyri were positively correlated with FMRP, suggesting that decreased FMR1 protein expression underlies deficits in math performance in persons with fraX. More broadly, this investigation demonstrates a unique bridging of cognitive and molecular neuroscience and represents a useful approach for the study of brain development and function. PMID- 12112764 TI - Modulation of activity in temporal cortex during generation of inner speech. AB - Monitoring one's thoughts (in the verbal modality) is thought to be critically dependent on the interaction between areas that generate and perceive inner speech in the frontal and temporal cortex, respectively. We used functional magnetic resonance imaging (fMRI) to examine the relationship between activity in these areas while the rate of inner speech generation was varied experimentally. The faster rate was associated with activation in the left inferior frontal gyrus, the right pre- and postcentral gyri and both superior temporal gyri. Thus, temporal cortical activation was associated with increasing the rate of covert articulation, in the absence of external auditory input, suggesting that there is effective fronto-temporal connectivity. Furthermore, this may provide support for the existence of feed forward models, which suggest that activity in regions responsible for verbal perception is modulated by activity in areas that generate inner speech. PMID- 12112766 TI - Modality independence of word comprehension. AB - Functional magnetic resonance imaging (fMRI) was used to examine the functional anatomy of word comprehension in the auditory and visual modalities of presentation. We asked our subjects to determine if word pairs were semantically associated (e.g., table, chair) and compared this to a reference task where they were asked to judge whether word pairs rhymed (e.g., bank, tank). This comparison showed task-specific and modality-independent activation for semantic processing in the heteromodal cortices of the left inferior frontal gyrus (BA 46, 47) and left middle temporal gyrus (BA 21). There were also modality-specific activations in the fusiform gyrus (BA 37) for written words and in the superior temporal gyrus (BA 22) for spoken words. Our findings are consistent with the hypothesis that word form recognition (lexical encoding) occurs in unimodal cortices and that heteromodal brain regions in the anterior as well as posterior components of the language network subserve word comprehension (semantic decoding). PMID- 12112765 TI - Comparison of spatial normalization procedures and their impact on functional maps. AB - The alignment accuracy and impact on functional maps of four spatial normalization procedures have been compared using a set of high resolution brain MRIs and functional PET volumes acquired in 20 subjects. Simple affine (AFF), fifth order polynomial warp (WRP), discrete cosine basis functions (SPM), and a movement model based on full multi grid (FMG) approaches were applied on the same dataset for warping individual volumes onto the Human Brain Atlas (HBA) template. Intersubject averaged structural volumes and tissue probability maps were compared across normalization methods and to the standard brain. Thanks to the large number of degrees of freedom of the technique, FMG was found to provide enhanced alignment accuracy as compared to the other three methods, both for the grey and white matter tissues; WRP and SPM exhibited very similar performances whereas AFF had the lowest registration accuracy. SPM, however, was found to perform better than the other methods for the intra-cerebral cerebrospinal fluid (mainly in the ventricular compartments). Limited differences in terms of activation morphology and detection sensitivity were found between low resolution functional maps (FWHM approximately 10 mm) spatially normalized with the four methods, which overlapped in 42.8% of the total activation volume. These findings suggest that the functional variability is much larger than the anatomical one and that precise alignment of anatomical features has low influence on the resulting intersubject functional maps. When increasing the spatial resolution to approximately 6 mm, however, differences in localization of activated areas appear as a consequence of the different spatial normalization procedure used, restricting the overlap of the normalized activated volumes to only 6.2%. PMID- 12112767 TI - Functional anatomy of syntactic and semantic processing in language comprehension. AB - A functional magnetic resonance imaging (fMRI) study was conducted to map syntactic and semantic processes onto the brain. Chinese-English bilingual subjects performed two experimental tasks: a syntactic plausibility judgment task in which they decided whether a viewed verb phrase was syntactically legal, and a semantic plausibility judgment task in which they decided whether a viewed phrase was semantically acceptable. A font size judgment task was used as baseline. It is found that a large-scale distributed neural network covering the left mid inferior frontal and mid-superior temporal cortices was responsible for the processing of Chinese phrases. The right homologue areas of these left cortical sites were also active, although the brain activity was obviously left lateralized. Unlike previous research with monolingual English speakers that showed that distinct brain regions mediate syntactic and semantic processing of English, the cortical sites contributing to syntactic analysis of Chinese phrases coincided with the cortical sites relevant to semantic analysis. Stronger brain activity, however, was seen in the left middle frontal cortex for syntactic processing (relative to semantic processing), whereas for semantic processing stronger cortical activations were shown in the left inferior prefrontal cortex and the left mid-superior temporal gyri. The overall pattern of results indicates that syntactic processing is less independent in reading Chinese. This is attributable to the linguistic nature of the Chinese language that semantics and syntax are not always clearly demarcated. Equally interesting, we discovered that when our bilingual subjects performed syntactic and semantic acceptability judgments of English phrases, they applied the cerebral systems underlying Chinese reading to the processing of English. PMID- 12112768 TI - Spatial independent component analysis of functional MRI time-series: to what extent do results depend on the algorithm used? AB - Independent component analysis (ICA) has been successfully employed to decompose functional MRI (fMRI) time-series into sets of activation maps and associated time-courses. Several ICA algorithms have been proposed in the neural network literature. Applied to fMRI, these algorithms might lead to different spatial or temporal readouts of brain activation. We compared the two ICA algorithms that have been used so far for spatial ICA (sICA) of fMRI time-series: the Infomax (Bell and Sejnowski [1995]: Neural Comput 7:1004-1034) and the Fixed-Point (Hyvarinen [1999]: Adv Neural Inf Proc Syst 10:273-279) algorithms. We evaluated the Infomax- and Fixed Point-based sICA decompositions of simulated motor, and real motor and visual activation fMRI time-series using an ensemble of measures. Log-likelihood (McKeown et al. [1998]: Hum Brain Mapp 6:160-188) was used as a measure of how significantly the estimated independent sources fit the statistical structure of the data; receiver operating characteristics (ROC) and linear correlation analyses were used to evaluate the algorithms' accuracy of estimating the spatial layout and the temporal dynamics of simulated and real activations; cluster sizing calculations and an estimation of a residual gaussian noise term within the components were used to examine the anatomic structure of ICA components and for the assessment of noise reduction capabilities. Whereas both algorithms produced highly accurate results, the Fixed-Point outperformed the Infomax in terms of spatial and temporal accuracy as long as inferential statistics were employed as benchmarks. Conversely, the Infomax sICA was superior in terms of global estimation of the ICA model and noise reduction capabilities. Because of its adaptive nature, the Infomax approach appears to be better suited to investigate activation phenomena that are not predictable or adequately modelled by inferential techniques. PMID- 12112769 TI - Different activation dynamics in multiple neural systems during simulated driving. AB - Driving is a complex behavior that recruits multiple cognitive elements. We report on an imaging study of simulated driving that reveals multiple neural systems, each of which have different activation dynamics. The neural correlates of driving behavior are identified with fMRI and their modulation with speed is investigated. We decompose the activation into interpretable pieces using a novel, generally applicable approach, based upon independent component analysis. Some regions turn on or off, others exhibit a gradual decay, and yet others turn on transiently when starting or stopping driving. Signal in the anterior cingulate cortex, an area often associated with error monitoring and inhibition, decreases exponentially with a rate proportional to driving speed, whereas decreases in frontoparietal regions, implicated in vigilance, correlate with speed. Increases in cerebellar and occipital areas, presumably related to complex visuomotor integration, are activated during driving but not associated with driving speed. PMID- 12112770 TI - fMRI study comparing names versus pictures of objects. AB - We performed an fMRI one-back recognition study aimed at distinguishing the semantic versus perceptual aspects of how objects and their written forms are processed. There were three types of visually presented items: pictures (schematic drawings of objects); words identifying these objects; and a mixed condition in which pictures were interleaved with words. A semantic decision about object identity was required when pictures were interleaved with words. This condition, contrasted with the other two, invoked a larger signal in multiple areas, including frontal cortex, bilateral occipitotemporal cortex, and the right middle temporal gyrus. We propose that the left occipitotemporal and right temporal activations are indicative of the neural substrate mediating picture-word conversions, whereas the frontal activations reflect the coordinating functions of the central executive. PMID- 12112771 TI - Neural basis of prosopagnosia: an fMRI study. AB - Brain imaging research has identified at least two regions in human extrastriate cortex responding selectively to faces. One of these is located in the mid fusiform gyrus (FFA), the other in the inferior occipital gyrus (IOG). We studied activation of these areas using fMRI in three individuals with severely impaired face recognition (one pure developmental and two childhood prosopagnosics). None of the subjects showed the normal pattern of higher fMRI activity to faces than to objects in the FFA and IOG or elsewhere. Moreover, in two of the patients, faces and objects produced similar activations in the regions corresponding to where the FFA and IOG are found in normal subjects. Our study casts light on the important role of FFA and IOG in the network of areas involved in face recognition, and indicates limits of brain plasticity. PMID- 12112772 TI - Non-invasive assessment of language lateralization by transcranial near infrared optical topography and functional MRI. AB - Near infrared optical topography (OT) is the simultaneous acquisition of hemoglobin absorption from an array of optical fibers on the scalp to construct maps of cortical activity. We demonstrate that OT can be used to determine lateralization of prefrontal areas to a language task that has been validated by functional MRI (fMRI). Studies were performed on six subjects using a visually presented language task. Laterality was quantified by the relative number of activated pixels in each hemisphere for fMRI, and the total hemoglobin responses in each hemisphere for OT. All subjects showed varying degrees of left hemisphere language dominance and the mean laterality indices for subjects who underwent both OT and fMRI were in good agreement. These studies demonstrate that OT gives predictions of hemispheric dominance that are consistent with fMRI. Due to the ease of use and portable nature of OT, it is anticipated that optical topography will be valuable tool for neurological examinations of cognitive function. PMID- 12112773 TI - Mismatch responses to randomized gradient switching noise as reflected by fMRI and whole-head magnetoencephalography. AB - The central auditory system of the human brain uses a variety of mechanisms to analyze auditory scenes, among others, preattentive detection of sudden changes in the sound environment. Electroencephalography (EEG) and magnetoencephalography (MEG) provide a measure to monitor neuronal cortical currents. The mismatch negativity (MMN) or field (MMNm) reflect preattentive activation in response to deviants within a sequence of homogenous auditory stimuli. Functional magnetic resonance imaging (fMRI) allows for a higher spatial resolution as compared to the extracranial electrophysiological techniques. The image encoding gradients of echo planar imaging (EPI) sequences, however, elicit an interfering background noise. To circumvent this shortcoming, the present study applied multi-echo EPI mimicking an auditory oddball design. The gradient trains (SOA = 800 msec, 94.5 dB SPL, stimulus duration = 152 msec) comprised amplitude (-9 dB) and duration (76 msec) deviants in a randomized sequence. Moreover, the scanner noise was recorded and applied in a whole-head MEG device to validate the properties of this specific material. Robust fMRI activation patterns emerged in response to the deviant gradient switching. Changes in amplitude activated the entire auditory cortex, whereas the duration deviants elicited right-lateralized signal increase in secondary areas. The recorded scanner noise evoked reliably right lateralized mismatch MEG responses. Source localization was in accordance with activation of secondary auditory cortex. The presented paradigm provides a robust and feasible tool to study the functional anatomy of early cognitive auditory processing in clinical populations such as schizophrenia. PMID- 12112774 TI - Pseudomonas aeruginosa and other predictors of mortality and morbidity in young children with cystic fibrosis. AB - We conducted a registry-based study to determine prognostic indicators of 8-year mortality and morbidity in young children with cystic fibrosis (CF). Patients ages 1-5 years from the 1990 U.S. Cystic Fibrosis Foundation (CFF) National Patient Registry served as the study cohort (N = 3,323). Registry data provided information on baseline characteristics in 1990, 8-year mortality, and clinical outcomes in 1998.P. aeruginosa respiratory infection was found to be a major predictor of morbidity and mortality. The 8-year risk of death was 2.6 times higher in patients who had respiratory cultures positive for P. aeruginosa in 1990 (95% confidence interval 1.6, 4.1) than in children without P. aeruginosa in their respiratory cultures. Culture-positive patients in 1990 also had a significantly lower percent predicted forced expiratory volume in 1 sec (FEV(1)) and weight percentile at follow-up, and they had an increased risk of continued P. aeruginosa respiratory infection and hospitalization for acute respiratory exacerbation in 1998. Among the other predictors of increased morbidity and mortality were lower baseline weight percentiles and number of CF-related hospitalizations during the baseline year.These findings confirm reports from previous smaller studies of outcomes among young children with CF, and highlight the potential to decrease the morbidity and mortality of young patients with CF through early intervention. PMID- 12112775 TI - Alcaligenes infection in cystic fibrosis. AB - The aim of this study was to investigate the effect of chronic Alcaligenes species infection of the respiratory tract on the clinical status of patients with cystic fibrosis. We conducted a retrospective case-controlled study. The microbiological records of all patients attending the Leeds Regional Pediatric and Adult Cystic Fibrosis Units from 1992-1999 were examined. Chronic Alcaligenes infection was defined as a positive sputum culture on at least three occasions over a 6-month period. These patients were compared with controls matched for age, gender, respiratory function, and Pseudomonas aeruginosa infection status. Respiratory function tests, anthropometric data, Shwachman-Kulczycki score, Northern chest x-ray score, intravenous and nebulized antibiotic treatment, and corticosteroid treatment were compared from 2 years before to 2 years after Alcaligenes infection. From a clinic population of 557, 13 (2.3%) fulfilled the criteria for chronic infection. The median age at acquisition of infection was 17.2 years (range, 6.5-33.6). There was no significant difference in the changes of percentage predicted values for FEV(1), FVC, FEF(25-75), or Shwachman Kulczycki and Northern chest x-ray scores, or in weight, height, and body mass index z-scores between Alcaligenes-infected cases and controls. There was also no significant difference in the use of antibiotics (intravenous and nebulized) or corticosteroids (inhaled and oral). We conclude that in our clinic, chronic infection with Alcaligenes species was uncommon. Chronically infected patients showed no excess deterioration in clinical or pulmonary function status from 2 years before to 2 years after primary acquisition. PMID- 12112776 TI - Prevalence of asthma and asthma-like symptoms in inner-city elementary schoolchildren. AB - American inner-city children are disproportionately affected by asthma. During the 1999-2000 school year, we conducted a survey of 6 Bronx, New York City elementary schools to assess the prevalence of asthma and asthma-like symptoms as reported by parents. Children with probable asthma had symptoms within the last 12 months and parents who indicated that their child had asthma. Children with possible asthma had symptoms within the last 12 months but lacked a diagnosis.Overall, 74% (4,775/6,433) of parents returned completed surveys, identifying 20% (949/4,775) of children as probable asthmatics, and 12% (589/4,775) as possible asthmatics. In multivariate analyses, probable asthma was associated with: Puerto Rican, Black, and white race/ethnicity, male gender, having health insurance, and registration at the poorest school. Possible asthma was associated with lack of health insurance and female gender, but was not associated with any specific race/ethnicity. Our findings support the effectiveness of school-based surveys to identify children at high risk for asthma. The challenge remains to engage children and families in appropriate follow-up care and to manage their illness, either through the use of school based health centers or stronger links to community services. PMID- 12112777 TI - Impact of respiratory illness on expiratory flow rates in normal, asthmatic, and allergic children. AB - We examined the effects of current respiratory illness (RI) on pulmonary function (PF) in 1,103 subjects who underwent spirometry at schools twice within a 4-month period. Before spirometry, subjects were asked if they had a "cold or other chest illness" during the previous month, and if so, whether they had fully recovered. Those who had not recovered were considered to have an RI. We found that children without RI at their first PF test who reported RI on retest had significantly lower forced expiratory volume in 1 sec (FEV(1)) (-0.8%), peak expiratory flow rate (PEFR) (-2.2%), forced expiratory flow between 25-75% of vital capacity (FEF(25-75)) (-3.5%), and forced expiratory flow at 75% of vital capacity (FEF(75)) (-5.1%) than those without RI on both test and retest. Restriction of subjects to those without a history of doctor-diagnosed asthma did not appreciably change these findings. Children with hay fever had significantly larger RI-associated decreases for FEV(1), FEF(25-75), and FEF(75), but not PEFR, than those without hay fever. Among asthmatic subjects, those with active asthma had larger RI-associated decreases in FEF(25-75) and FEF(75), but not PEFR, than those without asthma. There was limited evidence that small airway losses were greater in children less than 12.5 years old. We conclude that RI in children who are well enough to attend school may reduce expiratory flow rates. These effects are greater for children with active asthma or hay fever than in those without, and may be inversely related to age. PMID- 12112778 TI - Cytokine secretion in children with acute Mycoplasma pneumoniae infection and wheeze. AB - The aim of this study was to evaluate cytokine secretion in children with acute Mycoplasma pneumoniae infection and wheeze. We studied 25 patients aged 2-14 years with an acute episode of wheezing (15 with acute M. pneumoniae infection) and 16 healthy controls of similar gender and age (8 with laboratory evidence of asymptomatic acute M. pneumoniae infection). Serum interleukin (IL)-2, interferon (IFN)-gamma, IL-4, and IL-5 concentrations were measured in samples obtained at enrollment, using enzyme-linked immunosorbent assay kits. In the presence of wheezing, IL-5 concentrations were significantly higher in subjects with acute M. pneumoniae infection (33.415 +/- 22.138 pg/mL) than in those without such infection (2.320 +/- 1.846 pg/mL, P < 0.0001). The children with acute M. pneumoniae infection and wheeze had higher IL-5 concentrations (33.415+/-22.138 pg/mL) than those with asymptomatic acute infection and without wheeze (1.740 +/- 2.299 pg/mL, P < 0.0001). No significant between-group differences were observed in terms of IL-2, IFN-gamma, or IL-4 levels, or the prevalence of atopy. Our results show that children with wheezing and acute M. pneumoniae infection have a specific cytokine profile characterized by a significant increase in serum levels of IL-5. This immune response may be important for understanding the pathophysiological mechanisms by which this pathogen contributes to the development of wheeze-related symptoms, and for identifying new treatment strategies. PMID- 12112779 TI - Peripheral blood lymphopenia and neutrophilia in children with severe respiratory syncytial virus disease. AB - It is not known why respiratory syncytial virus (RSV) is associated with prolonged sequelae in many children. Measles virus (also a paramyxovirus), acute stress in sepsis, and cardiac bypass all cause lymphopenia. Using a retrospective analysis of records of children in Bristol with RSV infections over 5 years, we found that children with RSV had lower lymphocyte counts than unstressed, stable children prior to cardiac surgery. Children who required intensive care had the lowest lymphocyte counts. Neutrophil counts were raised in RSV-infected children. These data may offer an insight into pathological mechanisms, and suggest new research avenues. PMID- 12112780 TI - Increased interleukin-8 and monocyte chemoattractant protein-1 concentrations in mechanically ventilated preterm infants with pulmonary hemorrhage. AB - Pulmonary hemorrhage (PH) is a serious complication causing acute respiratory distress in the premature infant, and it is associated with significant mortality and morbidity. The role of inflammatory mediators in this condition is largely undefined. Serial tracheal aspirates (TA) were obtained at intervals from 65 mechanically ventilated infants with birth weights less than 1,250 g during the first 21 days of life. Clinically significant PH developed in 15 infants. TA concentrations of interleukin-8 (IL-8) and monocyte chemoattractant protein-1 (MCP-1) were determined by enzyme-linked immunosorbent assay (ELISA).PH was associated with an increased risk of death, bronchopulmonary dysplasia, intraventricular hemorrhage, and prolonged need for mechanical ventilation and supplemental oxygen. TA aspirate concentrations of IL-8 and MCP-1 (P = 0.001, ANOVA) were significantly increased in infants with PH compared to infants who did not develop this condition. TA cytokine concentrations were also significantly increased in infants who developed bronchopulmonary dysplasia (BPD). Peak TA concentrations of IL-8 and MCP-1 were significantly higher in infants with poor outcome (BPD or death). TA MCP-1 but not IL-8 concentrations were significantly higher in infants who were oxygen-dependent at 36 weeks postconceptional age. These data suggest a pathogenic role for IL-8 and MCP-1 in the development of adverse pulmonary outcome in preterm infants with clinically significant PH. PMID- 12112781 TI - Novel application of biological glue in the management of a complicated pneumothorax in cystic fibrosis. AB - Spontaneous pneumothorax is a frequent complication in advanced lung disease of cystic fibrosis. We describe a case of a complicated pneumothorax in a 21-year old-woman with cystic fibrosis who was effectively treated with the application of biological glue via a minithoracotomy. PMID- 12112783 TI - Unusual cause of chronic cough in a four-year-old cured by uvulectomy. AB - A 4-year-old boy had a history of persistent barking cough unresponsive to medical treatment since infancy. He described a feeling of something in his throat. When investigated by flexible fiberoptic bronchoscopy, the only abnormality was the uvula resting in contact with the epiglottis. The cough was no longer present after uvulectomy. PMID- 12112784 TI - Multiple cavitating pulmonary nodules and clubbing in a 12-year-old girl. AB - We report on a 12-year-old girl with a prolonged history of cough and hemoptysis on three occasions. X-ray and computed tomography of the chest showed several cavitating lesions and mediastinal lymphadenopathy. Lung biopsy revealed nodular sclerosing Hodgkin's disease. Hodgkin's disease should be considered in the differential diagnosis of cavitating pulmonary lesions. PMID- 12112782 TI - Flexible fiberoptic bronchoscopic removal of a fractured synthetic tracheostomy tube in a 3-year-old child. AB - We report on a 3-year-old boy, who was tracheotomized for prolonged ventilatory support in response to respiratory failure due to spinal muscular atrophy. He aspirated a fractured synthetic tracheostomy tube. It was successfully removed by flexible fiberoptic bronchoscopy. PMID- 12112785 TI - Pulmonary lipoblastoma in an 18-month-old child: a unique tumor in children. AB - Lipoblastoma, a rare benign tumor of adipose tissue, typically occurs in the superficial or deep layers of soft tissue on the trunk or extremities. Other sites of occurrence include the head, neck, and retroperitoneum. Lipoblastoma of the chest wall and parietal pleural have been reported, but occurrence within the lung has not been previously described. We report a case of pulmonary lipoblastoma in a young child presenting with complete opacification of the left hemithorax and mediastinal shift on chest radiograph. A lobectomy was performed, and the diagnosis was made by histological examination. PMID- 12112786 TI - Response to "Use of nasal potential differences and sweat chloride as outcome measures in multicenter clinical trials in subjects with cystic fibrosis". PMID- 12112790 TI - Prognostic factors in cystic fibrosis in a single center in Brazil: A survival analysis. AB - The purpose of the present study was to identify prognostic factors related to death in patients with cystic fibrosis (CF). Records of 127 patients with CF submitted to a systematic protocol were retrospectively reviewed. Prognostic factors associated with demographic, nutritional, clinical, and laboratory findings on admission were studied. The median follow-up was 44 months. The analysis was conducted in two steps. First, in a univariate analysis, variables associated with death outcome were identified by the log-rank test. Then the variables that were significantly associated with adverse outcome were included in a multivariate analysis. This analysis, using the Cox proportional hazards model, was performed to identify variables that were independently associated with a worse prognosis. Only variables that remained independently associated with adverse outcome were included in the final model. Three variables were identified as independent predictors of adverse outcome: age at diagnosis under 3 months (relative risk (RR) = 13, 95% CI = 4.5-34, P < 0.001); Shwachman-Kulczycki (S-K score) score below 70; and birth weight under 3,000 g. An interaction effect between S-K score and birth weight was identified. The relative risk of death for the S-K score was 24 (95% CI = 3-195) when birth weight was lower than 3,000 g; on the other hand, when birth weight was 3,000 g or more, the relative risk for the S-K score was 1.4 (95% CI = 0.24-8.83). The combination of three factors (low S-K score, low birth weight, and young age at diagnosis) is indicative of a poor prognosis in CF patients. PMID- 12112791 TI - Effects of salmeterol on arterial oxyhemoglobin saturations in patients with cystic fibrosis. AB - Sleep-related oxygen desaturation has been described in patients with cystic fibrosis (CF). Thus we studied the effects of inhaled Salmeterol xinafoate, a long-acting beta-2 agonist, on transcutaneous oxyhemoglobin saturation in sleeping, stable CF patients. Patients with stable CF (n = 23) were divided into responders and nonresponders to beta-2 agonists, based on an albuterol challenge during daytime testing, and then they received salmeterol before sleep, in a double-blind crossover design. Overnight oxyhemoglobin saturation measurements and spirometry on awakening were performed. Salmeterol administration before sleep resulted in statistically significant increases in mean arterial oxyhemoglobin saturation and in FEV(1) and FEF(25-75) on awakening compared to placebo values, but only in patients responding to daytime albuterol inhalation by showing improvement in lung function. We conclude that salmeterol inhalation at bedtime could prevent or reduce nocturnal hypoxemia in daytime albuterol responsive CF patients, thus improving the long-term clinical outcome of CF lung disease. PMID- 12112792 TI - Spinal muscular atrophy type 1: management and outcomes. AB - Our objectives were to describe survival, hospitalization, speech, and outcomes related to respirator needs for spinal muscular atrophy type 1 (SMA1) patients, using noninvasive or tracheostomy ventilation. From 65 SMA patients referred to our clinic since 1996, we chose 56 SMA1 patients who developed respiratory failure before age 2 years. Patients either had tracheostomy tubes (group A), or used noninvasive ventilation and assisted coughing; a previously reported extubation protocol (group B) was used as needed. Sixteen patients underwent tracheostomy at 10.8 +/- 5.0 months of age, 33 were in group B, and 7 others died without life-support interventions. Compared to group B, group A patients had fewer hospitalizations until age 3 years, but more after age 5, and 15 of 16 lost all spontaneous breathing tolerance posttracheostomy and could not speak. One group A patient died at 16 months of age, and the others were 73.8 +/- 57 months of age (the oldest was 19 years old). Two group B patients died at 6 and 13 months, respectively, whereas the other 31 were 41.8 +/- 26.0 months (and up to 8.3 years) old. Three of 31 in group B required high-span positive inspiratory pressure plus positive end-expiratory pressure (PIP + PEEP) continuously with minimal tolerance for breathing on their own, and 4 could not communicate verbally.In conclusion, SMA type 1 children can survive beyond 2 years of age when offered tracheostomy or noninvasive respiratory support. The latter is associated with fewer hospitalizations after age 5 years, freedom from daytime ventilator use, and the ability to speak. PMID- 12112793 TI - Idiopathic interstitial pneumonitis in children: a national survey in the United Kingdom and Ireland. AB - Interstitial lung disease (ILD) of unknown etiology in immunocompetent patients is rare in children. A national survey was carried out in the United Kingdom and Ireland over a 3-year period in order to identify prevalence, age distribution, histopathology, natural history of the illness, and response to current treatment.Forty-six cases were identified, including 29 males and 17 females. Seventy-six percent presented in the first year of life. Nine (16%) occurred within four families. Conventional treatment with pulsed methylprednisolone, prednisolone, or hydroxychloroquine, singly or in combination, resulted in an excellent response in 65% of cases. Seven children died (15%). The recurrence risk for further children within the same family to develop ILD is estimated to be approximately 10%. The prevalence rate of this condition in the United Kingdom and Ireland during the period of study for children aged 0-16 years is estimated to be 3.6 cases/million. PMID- 12112794 TI - Foreign body aspiration: what is the outcome? AB - Undiagnosed and retained foreign bodies may result in serious complications such as pneumonia, atelectasis, or bronchiectasis. We reviewed a total of 174 children with foreign body aspiration (FBA). Clinical, radiological, and bronchoscopic findings of these patients were evaluated according to the nature of foreign body and elapsed time from aspiration to diagnosis. Significant differences were noted between patients with organic and inorganic FBA in terms of clinical and radiological findings. Cough, recurrent pneumonia, and fever were the most common presenting symptoms in patients with delayed diagnosis. Long-term follow-up was available for 110 patients for a mean duration of 37.8 +/- 23.7 months (range, 1 88 months). We evaluated the course of recovery after bronchoscopic removal. Organic FBA was of comparable duration as for inorganic FBA, and prolonged follow up was associated with increased risk of persistent symptoms and bronchiectasis (P < 0.001). The risk of long-term complications increased with increasing elapsed time from aspiration to diagnosis; complications were as high as 60% in children who were diagnosed 30 days after FBA (P = 0.0035). Bronchiectasis was a major complication, found in 25% of patients whose diagnosis was delayed by more than 30 days (P = 0.0001). Three patients with bronchiectasis underwent lobectomy. Patients with persistent asthma-like symptoms such as cough and wheezing required treatment with inhaled corticosteroids and bronchodilators. The positive response to this treatment was thought to be a confirmation of the development of transient bronchial hyperresponsiveness induced by foreign bodies. We conclude that timely diagnosis and appropriate treatment of FBA is important to prevent long-term complications in affected children. PMID- 12112795 TI - Incidence of exercise-induced arterial hypoxemia in prepubescent females. AB - Due to the recent discovery of exercise-induced arterial hypoxemia (EIAH) in healthy active women with normal levels of peak oxygen uptake (V'(O(2)peak), this study examined the incidence of EIAH in prepubescent females. Nineteen healthy, active, prepubescent females (X +/- SD: age = 11.1 +/- 1.6 years; height = 145.8 +/- 9.1 cm; weight = 35.6 +/- 7.0 kg) performed a progressive maximal exercise test on an electronically braked cycle ergometer starting at 0 W and increasing power by 15 W. min(-1). During this test, expired gases, heart rate (HR), and percent arterial oxyhemoglobin saturation (%SaO(2)) were measured. Results for physiological variables at maximal exercise were as follows: V'(O(2)peak) = 43.7 +/- 7.0 ml x kg(-1) x min(-1); HR(max) = 199 +/- 5 beats x min(-1); %SaO(2) = 96.6 +/- 1.2%. For nearly all subjects, the %SaO(2) at maximal exercise was above levels that would reduce V'(O(2)peak). Therefore, in comparison to previous reports of EIAH in adult women with similar V'(O(2)peak), EIAH does not appear to occur in the prepubescent female population. PMID- 12112796 TI - Normal values for maximal static inspiratory and expiratory pressures in healthy children. AB - Maximal static respiratory pressures are a simple measure of respiratory muscle strength. In order to construct a set of equations describing normal values, we measured maximal inspiratory (P(Imax)) and expiratory (P(Emax)) pressures in 296 children (144 boys and 152 girls), aged 7-14 years, in sitting and standing positions. The boys reached higher values in sitting and standing positions for P(Imax) (-8.29 +/- 2.69 and -8.19 +/- 2.73 kPa, respectively) and P(Emax) (8.02 +/- 2.32 and 7.94 +/- 2.32 kPa, respectively) than girls (-6.53 +/- 1.99 and 6.60 +/- 2.03 kPa for P(Imax) and 6.91 +/- 1.79 and 7.13 +/- 1.81 kPa for P(Emax) for sitting and standing positions, respectively); the differences between boys and girls were highly significant (P < 0.001 in all instances). There were no differences regarding body position during measurements in both genders. Multiple correlation analysis showed significant correlations of pressures to age in boys in all cases, but in girls only for P(Imax) in standing position. Therefore, equations describing reference values were constructed with respect to age as the independent variable. Maximal pressures also correlated with maximal inspiratory and expiratory flows. The measurements of P(Imax) and P(Emax) are useful in assessing respiratory muscle strength despite their relatively large variability. P(Imax) and P(Emax) also correlate with maximum peak expiratory and inspiratory flows. Children generate lower pressures and lower maximal flows than adolescents and adults. PMID- 12112797 TI - Seasonal variation in respiratory syncytial virus chest infection in the tropics. AB - Respiratory syncytial virus (RSV) is the most important cause of lower respiratory tract infection (LRTI) in young children. We determined if there was a seasonal variation in Malaysia in the incidence of RSV infection in young children admitted with LRTI, and possible associations of RSV infection with local meteorological parameters. A total of 5,691 children, aged less than 24 months and hospitalized with LRTI (i.e., bronchiolitis and pneumonia) between 1982-1997, were included in this study. Nasopharyngeal samples were collected and examined for RSV by immunofluorescence, viral culture, or both. Seasonal variations were determined by analyzing the monthly RSV-positive isolation rate via time series analysis. Possible correlations with local meteorological parameters were also evaluated.RSV was isolated in 1,047 (18.4%) children. Seasonal variations in RSV infection rate were evident and peaked during the months of November, December, and January (test statistics [T] = 53.7, P < 0.001). This seasonal variation was evident for both bronchiolitis and pneumonia categories (T = 42.8 and 56.9, respectively, P < 0.001). The rate of RSV infection appeared to correlate with the monthly number of rain days (r = 0.26, P < 0.01), and inversely with the monthly mean temperature (r = -0.38, P < 0.001). In the tropics, seasonal variations in the incidence of RSV infection are evident, with an annual peak in November, December, and January. This information provides a guide for healthcare provisions and implementation of RSV prevention. PMID- 12112798 TI - Beclomethasone dipropionate and salbutamol by metered dose inhaler in infants and small children with recurrent wheezing. AB - The efficacy of beclomethasone dipropionate (BDP) to control respiratory symptoms was evaluated in 31 children under age 2 years with recurrent wheezing. The study was conducted in a double-blind, parallel, and placebo-controlled fashion. The two study groups received either salbutamol plus BDP 200 microg bid by metered dose inhaler (MDI) with a spacer, or salbutamol MDI plus a placebo. Inhaled corticosteroid (IC) and placebo were administered for 8 weeks. Patients were seen every 2 weeks as outpatients, and their progress was evaluated by clinical examination and a daily symptom score card. At the end of the study, patients in both groups had significantly decreased symptoms. No significant difference was found between BDP and placebo groups regarding clinical score, number of salbutamol doses, sleep disturbances, number of symptom-free days, feelings of insecurity of mothers regarding the infants' life due to wheezing, and mothers' perceptions of progress in their infants' respiratory symptoms. We conclude that salbutamol plus 200 microg bid of BDP inhaled from an MDI with a spacer for 8 weeks is no better than salbutamol alone for decreasing recurrent wheezing in small children under age 24 months. PMID- 12112799 TI - Effects of variable concentrations of inhaled nitric oxide and oxygen on the lungs of newborn piglets. AB - We previously demonstrated that inhalation of high concentrations of nitric oxide (iNO) and oxygen for 48 hr causes significant lung injury in newborn piglets. To determine if these effects persist at lower concentrations, groups of newborn piglets were mechanically ventilated for 48 hr with (study 1) constant O(2) (90 100%) and decreasing iNO (100-2 ppm) or (study 2) constant iNO (50 ppm) and decreasing O(2) (95-30%). Bronchoalveolar lavage (BAL) fluid was assayed for surfactant function, and markers of lung inflammation and physiologic parameters were monitored. Neutrophil chemotactic activity (NCA), % neutrophils, and total protein (TP) concentrations decreased significantly in BAL fluid of study 1 piglets as iNO was reduced and inhaled oxygen fraction remained constant, indicating less pulmonary injury at low iNO levels. Low-dose iNO (2 ppm) did not have antiinflammatory effects. However, surfactant function was minimally affected by lowering iNO, and was abnormal in all groups. In contrast, in study 2, pulmonary inflammation and injury were lower when O(2) was decreased to 70% or less, with iNO constant at 50 ppm. Surfactant function normalized and oxygenation improved in study 2 piglets when the inhaled oxygen fraction was decreased and iNO remained constant. These data suggest that iNO- and O(2)-induced lung injury may be minimized by weaning O(2) or iNO, although better physiologic function may be obtained when iNO concentrations are constant and O(2) is reduced. This has important implications in the clinical management of critically ill newborns treated with O(2) and iNO for pulmonary disorders. PMID- 12112800 TI - Factors that influence adherence to exercise and their effectiveness: application to cystic fibrosis. AB - Appropriate, regular exercise is of benefit to patients with cystic fibrosis (CF). As with other segments of the population, it has been difficult to devise exercise programs to which most patients will adhere for long periods of time. In healthy children, factors that are related to positive exercise compliance include social support, perceptions of competency and self-esteem, enjoyment of activity, and availability of a variety of activities. In patients with CF, complications associated with the disease, e.g., time required for other treatment and fatigue, make compliance with recommended exercise activities more difficult. Factors likely to increase compliance in this population include explicit and continued encouragement and support from the family and healthcare team, and the introduction of behavior-changing strategies. PMID- 12112801 TI - Recurrent pneumothoraces associated with nocturnal noninvasive ventilation in a patient with muscular dystrophy. AB - Although a common complication of mechanical ventilation in acute respiratory failure, spontaneous pneumothorax has been rarely reported among patients on chronic, intermittent, noninvasive positive pressure support. We report the first case of recurrent pneumothoraces associated with nocturnal bilevel positive airway pressure ventilation via a nasal mask.A 26-year old man with chronic respiratory failure secondary to an unclassified neuromuscular condition suffered four separate episodes of spontaneous pneumothorax over a 12-month period. Two episodes occurred while he was asleep on bilevel positive airway pressure support. He was found to have numerous subpleural blebs, and we propose a mechanism for their development. Following open pleurodesis and blebectomy, the patient has not had another pneumothorax. Given the increasing utilization of chronic nocturnal bilevel positive airway pressure ventilation, we suggest that healthcare providers and patients be made aware of this potentially life threatening complication. PMID- 12112802 TI - Inflammatory endobronchial polyps in childhood: clinical spectrum and possible link to mechanical ventilation. AB - Inflammatory polyps of the airways are now regarded as histopathologically distinct nonneoplastic endobronchial lesions, which in adults are associated with a variety of chronic inflammatory insults. However, their clinical presentation in the pediatric population is extremely rare, with the etiology of such polyps poorly defined. The clinical and histopathological data from four pediatric patients, identified in the histopathology files of the Royal Brompton Hospital, were retrospectively reviewed. Three out of 4 patients had a history of mechanical ventilation in the neonatal period. In these 3 patients, the polyps were all situated in the proximal airways on the right side. These 3 patients presented at 6 weeks, 7 weeks, and 2 years, respectively, and were successfully treated by polypectomy at rigid bronchoscopy, with subsequent return to normality. One patient, presenting at 12 years of age without history of iatrogenic intervention, underwent a left lower lobectomy for a polyp sited in a segmental bronchus. Presentation in 3 of the 4 patients was with lobar collapse. The fourth patient presented with hyperinflation. We conclude that inflammatory endobronchial polyps may be associated with a history of mechanical ventilation in the neonatal period, polyp formation perhaps being secondary to airway trauma. The small caliber of the main airways in neonates may also be a contributory factor in presentation. PMID- 12112803 TI - Hypoglycemia due to adrenal suppression secondary to high-dose nebulized corticosteroid. AB - High-dose inhaled corticosteroids, greater than 400 mcg per day of beclomethasone dipropionate or equivalent, can cause adrenal insufficiency, but a hypoglycemic crisis has not been reported with the use of nebulized corticosteroids. We describe a 21-month-old asthmatic boy who had a hypoglycemic seizure during a proven acute adrenal crisis secondary to high-dose nebulized budesonide treatment. PMID- 12112804 TI - Risks and complications of nonbronchoscopic bronchoalveolar lavage in a pediatric intensive care unit. PMID- 12112806 TI - Diagnostic value and cost-effectiveness of on-site evaluation of fine-needle aspiration specimens: review of 5,688 cases. AB - Fine-needle aspiration (FNA) has proven to be a safe, economical, accurate, and rapid diagnostic technique. A successful FNA requires a specimen with adequate cellularity, high-quality preparation, an experienced aspirator, and cytopathologist. Up to 32% of FNAs in various organs (thyroid, breast, lung, etc.) may be nondiagnostic due to scant cellularity and poor preparation. On-site immediate evaluation of FNA specimens can be beneficial in determination of adequacy: triage for ancillary studies and provide a preliminary diagnosis of the specimen, which often facilitates rapid clinical decisions. In this study, we compared the on-site FNA interpretation with the final diagnosis and calculated its cost benefit. Reports of 5,688 on-site FNA cases from the files of the University of Pennsylvania Medical Center over a 5-yr period (1/1/96-12/31/00) were reviewed. Data of the immediate on-site interpretation and the final diagnosis in each case were compared to determine the diagnostic accuracy, clinical utility, and cost-effectiveness of on-site FNA evaluation. At our institution the average cost per FNA based on laboratory technical and professional fees (1,743 dollars) and the weighted average cost, based on utilization, of ancillary laboratory studies (328 dollars) and guidance procedures (1,025 dollars) is 3,096 dollars. An additional fee of 231 dollars per case is charged for on-site FNA evaluation by an attending cytopathologist. The average reported rate of nondiagnostic FNAs without on-site evaluation is 20%. Our own nondiagnostic rate for FNAs with on-site evaluation is 0.98%. If one assumes that patients will undergo a repeat FNA for each nondiagnostic specimen, the estimated additional cost in direct institutional charges is 2,022,626 dollars over 5 yr or 404,525 dollars per yr without on-site evaluation. This potential cost savings would be realized by utilizing on-site evaluation despite the additional fee due to a higher rate of specimen adequacy. Based on this study, on-site cytopathologic evaluation of FNA specimens is accurate, cost effective, and has improved patient care at our institution. PMID- 12112807 TI - Rare atypical squamous cells of undetermined significance (ASCUS): a clinically significant diagnosis? AB - To determine the clinical significance of rare atypical squamous cells of undetermined significance (ASCUS) in cervical screening, we studied 748 ASCUS cases prospectively noted to have rare abnormal cells. Comparing the rare ASCUS (RASC) group (defined as five or fewer abnormal cells) statistically to cases diagnosed as within normal limits (WNL), ASCUS unqualified as to number of cells low-grade squamous intraepithelial lesion (LGSIL), and high-grade SIL (HGSIL), we found that the probability of the RASC patients having an abnormal cytology (ASCUS/SIL) or biopsy (dysplasia) result within 1 yr was greater than that of the WNL group, but less than that for ASCUS unqualified, LGSIL, or HGSIL. When only ThinPrep specimens or cases with subsequent definitive SIL/dysplasia were considered, the RASC group was not significantly different from the WNL group. We conclude that RASC increases the risk of a subsequent abnormal cytology/biopsy result in conventional smears, but only when the threshold for abnormality is a subsequent ASCUS. It did not predict dysplasia (SIL/CIN) in those conventional samples. RASC did not have the power to predict any subsequent abnormality and did not appear to be clinically significant in ThinPrep samples. PMID- 12112808 TI - Utility of cytomorphologic criteria and p53 immunolocalization in distinguishing benign from malignant cystic squamous-lined lesions of the neck on fine-needle aspiration. AB - Fine-needle aspiration can effectively distinguish between benign and malignant cystic lesions of the head and neck. However, in some instances it may be difficult to arrive at a definite diagnosis due to limited cellularity, reactive changes, and cellular degeneration. In this study we examined the usefulness of six cytomorphologic features including the presence or prevalence of nuclear atypia, anucleated cells, tissue fragments, necrosis, and background inflammation in distinguishing between benign and malignant cystic lesions of the head and neck. The case cohort comprised 14 benign and 22 malignant cases. P53 immunostain was performed in 19 cases. These features were semiquantitatively measured on a sliding scale of 0-4 in both air-dried Diff-Quik-stained, alcohol-fixed Papanicolaou-stained smears and Millipore filter preparations. Mean and standard errors were calculated and statistical significance was evaluated by unpaired t test (StatView). Increased number of tissue fragments (P < 0.001), greater degree of nuclear atypia (P < 0.001), and background necrosis (P < 0.001) were more frequent in cystically degenerating squamous carcinoma as compared to benign squamous cystic lesions. No significant differences were noted in the number of single cells, anucleated cells, or in the amount of background inflammation found in aspirates of benign vs. malignant cystic squamous lesions. A higher percentage of the malignant cystic squamous lesions FNA cases demonstrated p53 immunolocalization but this difference was not statistically significant. Application of the above-mentioned cytomorphologic criteria and the use of p53 immunostain could effectively distinguish between benign and malignant cystic lesions of the head and neck. PMID- 12112809 TI - Identification of "papillary-like" clusters on endometrial cytologic preparations. AB - The presence of papillary cell clusters is taken as a serious indication in the diagnosis of well-differentiated endometrial adenocarcinoma in Japan. However, papillary-like clusters have been observed in both normal and benign samples. Therefore, overdiagnosis may occur in the observation of cytopreparations. The present study attempts to prepare a histological sample from an Auto Cyto Fix (ACF) sample prepared by the automatic fixation apparatus (ACF1000) using a membrane filter method after cytological observation and to examine its usefulness. In an ACF sample, a papillary-like cluster was observed and the coverslip was then removed, the circumference of the membrane filter was cut off, and the target cell cluster was smeared, embedded, and sliced. In the cases in which a papillary-like cluster was observed, even if differential diagnosis of the derivation was difficult due to a resemblance between the respective morphological findings, it was easily made by histological observation of an ACF sample. PMID- 12112810 TI - Fine-needle aspiration cytology of lobular carcinoma in situ. AB - Fine-needle aspiration cytology (FNAC) plays a key role in the preoperative diagnosis of breast carcinoma but is less reliable in the diagnosis of in situ lesions. The objective of the present study was to investigate the cytological features of lobular carcinoma in situ (LCIS), regarding which little data is available to date. Cytological features of FNAC of the breast from 21 patients with histology-proven LCIS were described and compared with surgical specimens. Aspirates from 8/21 cases had cell groups diagnostic for or compatible with LCIS. Aspirates from an additional two cases demonstrated hypercellular, dissociated, and more pleomorphic tumor cells, which were originally diagnosed as invasive lobular carcinoma (ILC). The remaining 11 aspirates were diagnosed as benign or nondiagnostic. FNAC from the eight diagnostic specimens were characterized by loosely cohesive cell groups composed of uniform cells with occasional intracytoplasmic lumina, slightly irregular and eccentric nuclei. We conclude that the main difficulty in diagnosing LCIS by FNAC is sampling rather than recognition of the lesions. However, one should be aware of the cytological features of LCIS in order to reach a correct diagnosis. There are no reliable cytological criteria that help in differentiating pleomorphic and dissociated LCIS from ILC. PMID- 12112811 TI - Colloid cyst of the third ventricle: cytomorphologic features on stereotactic fine-needle aspiration. AB - Stereotactic brain fine-needle aspiration (FNA) is a valuable diagnostic modality for evaluating space-occupying central nervous system disorders. Colloid cyst (CC) is a rare nonneoplastic lesion thought to arise from misplaced endodermal tissue in the anterosuperior portion of the third ventricle. This study summarizes cytomorphologic features of CC on FNA along with clinical, radiologic, and histopathologic correlation. Ten cases of CC of the third ventricle diagnosed on FNA were retrospectively reviewed for a period of 12 yr (1989-2000). Material was obtained under stereotactic radiologic guidance. Smears were stained with Diff-Quik and Papanicolaou stains and cell block sections with hematoxylin and eosin. The aspirates showed a characteristic sticky and viscous quality on gross examination. Smears showed abundant, amorphous, proteinaceous material with staining qualities similar to colloid aspirated from thyroid. This included a purplish, filmlike coating of the slide with occasional "cracking" artifact; thick, globular, eosinophilic fragments; and granular, ropelike, and somewhat viscous, mucinous material. Pathognomonic radiating hyphae-like structures were not seen. The cellular components varied from isolated cuboidal/columnar cells to large tissue fragments of glandular-type epithelium with focal ciliated border. Goblet cells were frequently identifiable, as were fragments of collagenous cyst wall. Stereotactic FNA of the CC of the third ventricle is an accurate and cost effective diagnostic modality. Cytomorphology coupled with the radiologic features is sufficiently unique for the diagnosis of this rare pathologic entity. PMID- 12112812 TI - Renal and extrarenal congenital rhabdoid tumor: diagnosis by fine-needle aspiration biopsy and FISH. AB - We report on 2 patients with congenital malignant rhabdoid tumor, one located to the kidney and the other to the soft parts of the cheek. Initial diagnosis was performed through percutaneous fine-needle aspiration biopsies, which yielded cytologic smears exhibiting highly characteristic rhabdoid cells, i.e., cells with a large, vesicular nucleus with a prominent nucleolus and cytoplasm exhibiting a large, dense, paranuclear inclusion. Interphase FISH demonstrated only one signal (heterozygous deletion) for the BCR gene in both cases, supporting the diagnosis. Surgical pathology and immunohistochemistry of both cases confirmed the diagnosis. Both patients died within the following 6 mo to 1 yr. PMID- 12112813 TI - Anaplastic T large cell lymphoma diagnosed by exfoliative cytology in a post renal transplant patient. AB - In the last two decades posttransplant lymphoproliferative disorders (PTLDs) have been recognized as a complication of organ transplantation with immunosuppression. The reported incidence of PTLDs in renal transplant patients ranges between 0.3-3% (Birkeland et al., Transplantation 1999;67:876-881). In contrast to the reported incidence of PTLDs in post bone marrow transplant, it is 1% in HLA-matched recipients and up to 20% in HLA mismatched T-cell depleted bone marrow recipients (Curtis et al., Blood 1996;94:2208-2216). In cardiac transplant recipients the reported incidence of PTLDs is between 1.8-9.8 (Mihalov et al., Clin Transplant 1996;10:248-255). PTLDs are predominately extranodal. They have varied morphologic patterns and clonality, but almost all are associated with Epstein-Barr virus (EBV). The vast majority are of B cell lineage; only about 10% are of T-cell origin. We report a T-cell anaplastic large cell lymphoma (ALCL) presenting with bilateral pleural effusion and liver involvement in a renal transplant recipient. PMID- 12112814 TI - Fine-needle aspiration cytology of recurrent granulosa cell tumor: case report with differential diagnosis and immunocytochemistry. AB - We report a case of an adult-type granulosa cell tumor of the ovary which was diagnosed in a 20-yr-old woman. After a 21-yr disease-free interval, she developed a pelvic recurrence, followed by a splenic metastasis and, more recently, omental masses. This report is concerned with the fine-needle aspiration (FNA) diagnosis of the granulosa cell tumor in the latter site and corroboration of the interpretation by immunocytochemistry. Only one previous case is similar to the present one documenting the role of immunocytochemistry in the evaluation of suspected metastatic granulosa cell tumor. The cytopathologic features of metastatic granular cell tumor have been described in a limited number of previous reports. PMID- 12112815 TI - Multiple loss of heterozygosity without K-ras mutation identified by molecular analysis on fine-needle aspiration cytology specimen of acinar cell carcinoma of pancreas. AB - Fine-needle aspiration diagnosis of pancreatic acinar cell carcinoma (PAC) is challenging. Typically, the cytologic findings in PAC are described as a cellular population of loosely cohesive clusters and single neoplastic cells. The individual cells have granular cytoplasm, uniform nuclei, a fine chromatin pattern, and occasional prominent nucleoli. These features are suggestive of PAC but not diagnostic. We illustrate a case in which the combination of cytopathologic findings, clinical information, and molecular analysis enabled us to arrive at the diagnosis of PAC. Although the cytomorphologic features alone were not specific, the presence of a markedly elevated serum lipase level, cutaneous lesions of fat necrosis, and loss of heterozygosity at 1p, 5q25 at the APC locus, 9p21 at the p16 locus, and 17p13 at the p53 locus were essential in excluding the main differential diagnostic entities including pancreatic ductal carcinoma, pancreatic endocrine tumor, and pancreatoblastoma. PMID- 12112816 TI - Peritoneal washing cytology findings of disseminated myxoid leiomyosarcoma of uterus: report of a case with emphasis on possible differential diagnosis. AB - We report the cytology findings of a rare case of myxoid leiomyosarcoma of the uterus with intraabdominal dissemination. The cytology showed uniformly dispersed spindly to polygonal "epithelioid" tumor cells focally linked by background matrix. Spindly tumor cells attaching to and radiating from branching capillary structures were also present. Occasionally, scattered "signet-ring" cells were found, mimicking metastatic poorly differentiated adenocarcinoma. The background mucoid substance was more conspicuous in the cell block sections. Gross and histologic examination of the surgical specimen revealed a large uterine leiomyosarcoma with prominent myxoid change. Ultrastructural study showed that the "signet-ring" appearance was mainly due to degenerative cytoplasmic change with ballooning of mitochondria, dilatation of endoplasmic reticulum, and elevation of outer nuclear membrane. In contrast to other spindle cell malignancies such as sarcomatoid mesothelioma, sarcomatoid carcinoma, or malignant melanoma, true sarcoma cells in general carry a distinctive cytologic appearance in washing/effusion fluid. In a correct clinical setting, the peculiar association with mucoid matrix and absence of classic lipoblasts should also raise the suspicion of metastatic myxoid leiomyosarcoma. PMID- 12112818 TI - Burkitt-type lymphoma of the breast: diagnosis by fine-needle aspiration cytology. PMID- 12112817 TI - Schwannomas: a pitfall in the diagnosis of pleomorphic adenomas on fine-needle aspiration cytology. AB - Fine-needle aspiration cytology (FNAC) is being employed with increasing frequency for the preoperative diagnostic workup of salivary gland lesions. Although most cases show morphologic features very characteristic of specific entities, few lesions, both benign and malignant, can cause problems in interpretation. We report four cases initially diagnosed on FNA as spindle-cell tumors, possibly benign nerve sheath tumors (BNST) in the salivary gland region. These cases were later diagnosed on histologic examination as schwannoma (two cases) and as pleomorphic adenoma (two cases). Review of the cytomorphology of these four cases enabled the correct diagnosis of pleomorphic adenoma in the two cases misinterpreted as BNST. Benign peripheral nerve sheath tumors should always be considered in the differential diagnosis of pleomorphic adenoma. A diligent search for epithelial elements is recommended prior to diagnosing BNST in the head and neck region. PMID- 12112819 TI - Cytohistologic and electron microscopic findings in bronchoalveolar lavage fluid in a case of pulmonary alveolar proteinosis. PMID- 12112820 TI - Solid cell nest in fine-needle aspiration of goiter. PMID- 12112821 TI - Deja vu in pap testing: return of the 5% false-negative fraction and the zero error rate. PMID- 12112822 TI - Litigation cells: their incidence and classification in gynecologic smears. AB - "Litigation cells" are defined as benign cells which may mimic dysplasia or cancer and might be used by plaintiffs' witnesses to imply that the cytotechnologist or pathologist "missed" cells of dysplasia or cancer. We reviewed 180 cervical smears from 166 patients who had hysterectomy for benign leiomyomas. All smears were performed within 12 months prior to hysterectomy. None of the uteri contained dysplasia or cancer on histologic examination. 90.6% of smears reviewed had at least one cell or cell group with atypia mimicking dysplasia or cancer. These "litigation cells" were classified as follows: parabasal cells, metaplastic squamous cells, degenerated endocervical cells, reactive endocervical cells, endometrial cells, neutrophils, histiocytes, and air dried cells. Diseases mimicked by these cells included squamous cell carcinoma, high-grade squamous intraepithelial lesion, low-grade squamous intraepithelial lesion, adenocarcinoma, and glandular dysplasia. These "litigation cells" can be correctly classified by experienced cytotechnologists and cytopathologists and recognized as benign. We recommend that in all cases of alleged malpractice against cytotechnologists and/or pathologists the smears should be reviewed by a panel of individuals trained and experienced in cytopathology. The smears should be reviewed without knowledge of the clinical outcome and in an environment that simulates the normal screening practice. PMID- 12112823 TI - Cytologic findings of malignant vascular neoplasms: a study of twenty-four cases. AB - Cytologic characterization of malignant vascular neoplasms (MVN) is difficult due to the wide range of differential diagnoses and sampling errors. The objective of this study was to identify criteria helpful in the cytologic diagnosis of MVN. The clinical presentation and cytomorphologic features of 22 angiosarcomas and two hemangioendotheliomas were analyzed. The criteria evaluated included cellularity, cellular arrangement, cell shape and size, pleomorphism, cytoplasmic borders, nuclear shape and number, chromatin pattern, nucleoli, background, and presence of angioformative structures. A previous history of MVN was noted in 18 of 24 patients. Specimens with epithelioid morphology were more cellular and pleomorphic and contained multinucleated cells. Specimens with spindle morphology were usually less cellular and less pleomorphic. Angioformative structures were identified in 11 of 24 cases. Awareness of features suggestive of MVN is necessary to raise the possibility of vascular origin, which can be confirmed with appropriate immunocytochemical studies. PMID- 12112824 TI - Cytological grading, apoptosis, and Bcl-2 protein expression in breast cancer. AB - A prospective study was conducted on 27 cases of infiltrating duct carcinoma of breast diagnosed by fine-needle aspirate cytology (FNAC), subjected to mastectomy following the FNA diagnosis. Cytological grading, Bcl-2 score, and quantification of apoptotic cell count were done on FNA material. Next, the carcinomas were graded on the corresponding histopathological sections. The overall concordance between cytological and histological gradings was 77.7%, with maximum concordance in Grade I tumors and minimum in Grade II. The mean apoptotic rates were 0.59 +/- 0.722, 2.11 +/- 0.707, and 2.95 +/- 0.854 in Grades I, II, and III, respectively. Similarly Bcl-2 scores were 1.36 +/- 0.82, 0.22 +/- 0.13, and 0.14 +/- 0.116, in Grades I, II, and III lesions, respectively. When the cytological grade was correlated with histological grade alone, and subsequently along with Bcl-2 scores and apoptotic rates, there was a significant improvement from 0.662-0.713 (P value, 0.001). PMID- 12112825 TI - Fine-needle aspiration cytology of lymphoproliferative disorders in the immunosuppressed patient: the diagnostic utility of in situ hybridization for Epstein-Barr virus. AB - We examined the diagnostic utility of fine-needle aspiration (FNA) in the immunosuppressed patient with particular attention to the value of in situ hybridization for Epstein-Barr virus encoded RNA (EBER), a test frequently performed on tissue sections in this setting. Six patients with adenopathy were identified: three with prior solid organ transplantation, two with prior bone marrow transplantation, and one with human immunodeficiency virus. Cytologic findings in involved tissues were positive for non-Hodgkin's lymphoma in one case, suspicious or atypical in three cases, negative/reactive in one case, and nondiagnostic in one case due to insufficient material. Two aspirates had a polymorphous lymphoid appearance; three showed predominantly monomorphous lymphoid morphology. Retrospectively, 4/4 FNA cell blocks were positive for EBER in a large proportion of atypical lymphoid cells. Together with the cytologic findings, EBER detection allows for more specific classification of these FNA samples as EBV-associated lymphoid proliferations. PMID- 12112826 TI - Diagnostic utility of galectin-3 and CD26/DPPIV as preoperative diagnostic markers for thyroid nodules. AB - The aim of this study was to search for diagnostic markers that could correctly identify thyroid nodular lesions requiring urgent surgical treatment. We investigated whether galectin-3 and dipeptidyl peptidase IV (CD26/DPPIV) could be potential markers for improving the diagnostic accuracy of conventional cytology. Seventy-nine patients with histologically proven thyroid diseases were analyzed. The immunocytochemical staining results showed galectin-3 expression in neoplastic cells of all 37 papillary carcinomas, five of six follicular carcinomas, all three anaplastic carcinomas, one of three medullary carcinomas, and two of 14 follicular adenomas. All 16 adenomatous goiters were negative for galectin-3 immunostaining. On the other hand, all 37 papillary carcinomas, all six follicular carcinomas, and one of three anaplastic carcinomas revealed CD26/DPPIV expression, whereas all three medullary carcinomas were negative. Among benign thyroid lesions, four of 14 follicular adenomas and two of 16 adenomatous goiters exhibited varying degrees of immunoreactivity for CD26/DPPIV. RT-PCR analysis demonstrated overexpression of galectin-3 and CD26/DPPIV mRNAs in all six papillary and all three follicular carcinomas analyzed, whereas the mRNA expressions of these molecules were barely or not detectable in benign thyroid lesions and normal thyroid tissues, except for one case of follicular adenoma. In conclusion, we demonstrate that galectin-3 and CD26/DPPIV were consistently coexpressed at protein and mRNA levels in differentiated thyroid carcinomas. We propose that combined immunostaining for galectin-3 and CD26/DPPIV in the preoperative evaluation of thyroid nodules may play a role in accurate cytodiagnosis. PMID- 12112827 TI - Immunolocalization of glucose transporter 1 and 3 in the placenta: application to cytodiagnosis of Papanicolaou smear. AB - A positive immunostaining for glucose transporter 1 (GLUT1) was exclusively localized in microvilli on the free surface of syncytiotrophoblasts in the placenta. An enhanced immunoreaction for glucose transporter 3 (GLUT3) was elicited in the cell membrane of intermediate trophoblasts and cytotrophoblasts. Neither GLUT1 nor GLUT3 was positive in decidual cells and epithelial components from cervical dysplasia and carcinoma in situ. Cervicovaginal smears from six pregnant women containing atypical cells of unknown origin were subjected to immunocytochemical testing with antibodies against GLUT1 and GLUT3. Atypical cells in smears from two pregnant women were found to be positive for GLUT3 while no specific immunoreaction for GLUT1 was elicited, indicating their origin from either intermediate trophoblasts or cytotrophoblasts. Through the use of antibodies against vimentin and cytokeratin 17, GLUT3-negative atypical cells were further sorted into decidual cells and epithelial components from cervical dysplasia, respectively. PMID- 12112828 TI - Diagnostic role of fine-needle aspiration of bone lesions in patients with a previous history of malignancy. AB - At the present time fine-needle aspiration (FNA) is considered a routine diagnostic procedure in evaluating neoplastic vs. nonneoplastic lesions in many organs, with high sensitivity and specificity. The purpose of this study was to assess the utility of FNA in areas of diagnostic difficulty and its limitations in evaluating bone lesions in patients with a previous history of malignancy. From 1989 to 2000, 249 CT-guided FNAs of bone lesion were performed at our institutions; 187/249 (75.1%) patients had a previous history of malignancy. Aspirated material was air-dried for Diff-Quik stain or fixed in ethanol for Papanicolaou staining. Subsequent surgical tissue was available in 69/187 (36.9%) of the cases. There were 114 males and 73 females, ages 14-86 yr (mean, 64 yr). The primary tumor site was lung 49, genitourinary 46, breast 31, gastrointestinal 28, hematopoietic 26, soft tissue/skin 5, and thyroid 2. There were 125 FNAs of the vertebral spine, 19 from the pelvis, 11 from the ribs, 9 from the sternum, 5 from the femur, and 18 from miscellaneous bone sites. Out of 187, 166 (88.7%) were malignant aspirates confirming the patients' primary malignancies. The most common malignancy encountered was adenocarcinoma, 126/187 (67.4%). Surgical tissue was available for review in 69 patients and the results were in agreement with the FNAs diagnosis in all cases. Nine out of 187 (4.8%) cases were diagnosed as marrow elements on cytological material. These patients have been followed for 1-9 yr and have failed to reveal signs or symptoms of clinical recurrence. Three out of 187 (1.6%) cases showed osteomyelitis. Nine out of 187 (4.8%) were unsatisfactory specimens, with biopsy follow-up available in four cases, showing three metastatic tumors and one case of osteomyelitis. FNA of metastatic bone lesions is a major step in pretreatment diagnosis. On satisfactory specimens, the cytological diagnosis viewed in the clinical-radiological context proves to be similar to surgical diagnosis. FNA is an excellent technique with a high accuracy rate in assessing metastatic bone lesions. PMID- 12112829 TI - Application of adjunct techniques in cytologic material in the diagnosis of rhabdomyosarcoma: case report and review of the literature. AB - A 4(1/2)-yr-old female presented with right-sided pleural effusion and a retroperitoneal mass. Cytologic analysis of the pleural fluid yielded malignant small round blue cells, which were noncohesive, 3-4 times the size of lymphocytes. The malignant cells had hyperchromatic, pleomorphic nuclei with moderate amounts of vacuolated cytoplasm. A few fiber-shaped cells were also seen. Immunostains for desmin, muscle-specific actin were positive; ultrastructural findings of thick and thin actin-myosin filaments confirmed the diagnosis of embryonal rhabdomyosarcoma. This case illustrates the importance of performing appropriate immunohistochemical stains and ultrastructural studies on cytological material to arrive at a definitive diagnosis. PMID- 12112830 TI - Bladder-washing cytology of metastatic ovarian granulosa cell tumor. AB - An 86-year-old Caucasian female presented with two weeks history of discomfort discharging urine, occasional hematuria, and suprapubic pain. The patient had a history of left salpingo-oophorectomy for an ovarian tumor, performed four years earlier. Ultrasound showed a solid mass surrounding the orifice of the left ureter. Bladder washing cytology yielded single, loosely cohesive syncytial aggregates of rather uniform cells. A few discretely grooved nuclei ("coffee bean nuclei") were seen. Histologic examination revealed muscular tissue infiltrated by oval to round cells, arranged in solid and follicular structures. The tumor cells were immunoreactive for estrogen receptor, inhibin, vimentin, and calretinin. The use of antibodies to pancytokeratin, inhibin, estrogen receptor, S-100, calretinin, and chromagranin could help confirm granulosa cell tumor. To my knowledge, there was no previous report on bladder washing cytology of metastatic granulosa cell tumor. PMID- 12112831 TI - Diagnostic intraocular aspiration cytology of choroidal melanoma. AB - Fine-needle aspiration cytology (FNAC) was performed on an intraocular mass in a 32-yr-old Indian woman and a cytologic diagnosis of malignant melanoma was made, supported by immunocytochemistry. The cytologic features included spindle-shaped tumour cells with minimal pleomorphism and the presence of nuclear grooves. No intracellular pigment was identified; however, positivity for HMB-45 allowed a rapid and reliable diagnosis. The utility of FNAC in an atypical presentation of choroidal melanoma is discussed. PMID- 12112832 TI - Cytomorphologic features of papillary cystadenocarcinoma of the parotid. AB - The fine-needle aspiration cytology findings in four cases of recently classified cystadenocarcinoma of the parotid gland are reported. In three cases a recurrent tumor was aspirated. Smear preparations in all four cases were cellular, with numerous papillary projections, single cells, and sheets of cells in varying proportion in a proteinaceous to mucoid background. The background mucin was in varying proportions. The cells were cuboidal to tall columnar with basal nuclei and mild pleomorphism. The cytoplasm was dense in three cases with variable amounts of mucin. In one case (Case 4) the epithelial cells resembled mucin secreting goblet cells, while in another case (Case 1) the cytoplasm showed multiple vacuolations. Mitosis was rare. Lymphoid tissue was seen in one case while macrophages and giant cells were seen in two cases. Epidermoid differentiation was absent in all four cases. Pathologic evaluation of the resected tumor confirmed the cytologic diagnosis. Clinical and radiologic evaluation failed to reveal any other potential primary site. Papillary cystadenocarcinomas of the parotid are rare but can be accurately diagnosed on FNAC. However, they need to be differentiated from mucoepidermoid and papillary acinic cell tumors. PMID- 12112833 TI - Fine-needle aspiration cytology of giant cell fibroblastoma: case report and review of the literature. AB - Giant cell fibroblastoma is an uncommon soft tissue neoplasm occurring in childhood. It appears to be the juvenile form of dermatofibrosarcoma protuberans, with which it shares some histologic, cytogenetic, and immunohistochemical features. We report, to our knowledge, the second description of the cytologic features of giant cell fibroblastoma. The present case represents a recurrent lesion in the soft tissues of the scrotum of a 17-yr-old male. The aspirate produced moderately cellular smears containing mononuclear cells, usually lying singly, but occasionally forming clusters. The majority of the individual cells possessed scanty bipolar cytoplasm or were devoid of cytoplasm. The nuclei were bland, with small nucleoli. Nuclear membranes frequently contained notches, creases, or folds. Small fragments of metachromatic stroma were present in the background and were often associated with small aggregates of cells. Rare multinucleated giant cells containing bland oval or basillary-shaped nuclei were admixed with the spindle-cell component. Necrosis and mitotic figures were not a component of the smears. Surgical resection of the mass confirmed the diagnosis of giant cell fibroblastoma. We review the characteristic cytologic features of giant cell fibroblastoma and compare them with other soft tissue tumors in the differential diagnosis. PMID- 12112834 TI - Pap prior to colposcopy. PMID- 12112836 TI - Nodular fasciitis. PMID- 12112839 TI - Proceedings of the Proteomic Forum. Munich, Germany, 16-19 September 2001. PMID- 12112840 TI - The post-genomic era of interactive proteomics: facts and perspectives. AB - The availability of completed genome sequences of several eukaryotic and prokaryotic species has shifted the focus towards the identification and characterization of all gene products that are expressed in a given organism. In order to cope with the huge amounts of data that have been provided by large scale sequencing projects, high-throughout methodologies also need to be applied in the emerging field of proteomics. In this review, we discuss methods that have been recently developed in order to characterize protein interactions and their functional relevance on a large scale. We then focus on those methodologies that are suitable for the identification and characterization of protein-protein interactions, namely the yeast two-hybrid system and related methods. Several recent studies have demonstrated the power of automated approaches involving the yeast two-hybrid system in building so-called "interaction networks", which hold the promise of identifying the entirety of all interactions that take place between proteins expressed in a certain cell type or organism. We compare the yeast two-hybrid system with several other screening methods that have been developed to investigate interactions between proteins that are not amenable to conventional yeast two-hybrid screenings, such as transcriptional activators and integral membrane proteins. The eventual adaptation of such methods to a high throughput format and their use in combination with automated yeast two-hybrid screenings will help in elucidating protein-protein interactions on a scale that would have been unthinkable just a few years ago. PMID- 12112841 TI - Clonal proteomics: one gene - family of proteins. AB - The work presented here attempts to consolidate our knowledge on cellular transcriptome and proteome. It takes into account that a typical activated cell (lymphocyte) contains 40 000 mRNA molecules at any time, and it represents about 5000 different molecular species of transcripts. Such a cell has about 1 000 000 000 protein molecules, some of them being present at 10 000 000 copies while others at a very low copy number (say 1 to 10 copies per cell). By studying cell free expression of individual cDNA clones (or pools of known complexity) we address to those rare molecular components that will remain undetected by the current analytical means. For our analysis we use cell free translation systems (wheat germ or rabbit reticulocyte origin) and we study polypeptide products originating from intact, or restriction endonuclease-treated cDNA clones. We conclude that in most instances expressed genes yield transcript(s) that translate into several, and often very numerous families of polypeptide species. In our ISODALT two-dimensional gel system we characterize the proteomic profile of the clonal polypeptide families in terms of their molecular mass, charge, multiple products, and appearance. PMID- 12112842 TI - Proteomic analysis of amphiphilic proteins of hexaploid wheat kernels. AB - Wheat proteins and specially gluten proteins have been well studied and are closely associated with baking products. Amphiphilic proteins (proteins that are soluble using nonionic detergent Triton X-114 ) also play an important role in wheat quality. Some of them, like puroindolines, are lipid binding proteins, and are strongly linked to dough foaming properties and to fine crumb texture. However many amphiphilic proteins are still unknown and both their physiological and technological functions remain to be analysed. In order to explore these proteins, proteomic analysis was carried out using 81 F9 lines, progeny obtained from an interspecific cross "W7984"x"Opata", and already used to built a map of more than 2000 molecular markers (International Triticeae Mapping Initiative, ITMImap). Two-dimensional electrophoresis (immobilized pH gradient (pH 6-11)x sodium dodecyl sulfate-polyacrylamide gel electrophoresis) was performed on amphiphilic proteins with three to five replicates for each line. Silver stained gels were analysed using Melanie 3 software. Genetic determinism was carried out on 170 spots segregating between the two parental hexaploid wheats. Many of these spots were mapped on different chromosomes of the ITMImap. Spots of interest were identified using matrix-assisted laser desorption/ionization-time of flight and some of them were partly sequenced using electrospray ionization-tandem mass spectrometry. This proteomic approach provided some very useful information about some proteic components linked to bread wheat quality and particularly to kernel hardness. PMID- 12112843 TI - Identification of the phosphotyrosine proteome from thrombin activated platelets. AB - Signalling cascades are regulated both positively and negatively by tyrosine phosphorylation. Integrin mediated platelet adhesion triggers signal transduction cascades involving translocation of proteins and tyrosine phosphorylation events, ultimately causing large signalling complexes to be assembled. In resting platelets, a small number of phosphorylated proteins are evident with molecular mass of 50-62 kDa and 120-130 kDa. In thrombin activated human platelets, however, there is a large increase in the number of tyrosine phosphorylated signalling proteins detected. These proteins include pCas (130 kDa), FAK (125 kDa), PI(3)k (85 kDa) and src (85 kDa). However, it is unlikely that this list of proteins represents all the dynamic changes that occur after platelet activation and it is understood that more proteins remain unidentified. In this study, we propose a method for the isolation of the phosphotyrosine proteome from both resting and thrombin activated human platelets. All the dynamic phosphotyrosine events that occur in the platelet after thrombin activation were isolated by immunoprecipitation, using the monoclonal antibody 4G10, allowing us to obtain higher concentrations of relatively low abundant proteins. The resulting phosphotyrosine proteomes were separated by two-dimensional gel electrophoresis. Sixty-seven proteins were reproducibly found to be unique in the thrombin activated platelet proteome when compared to resting platelets. We have positively identified ten of these proteins by Western blotting and matrix assisted laser desorption/ionisation-time of flight mass spectrometry and these include FAK, Syk, ALK-4, P2X6 and MAPK kinase kinase. This proteomics approach to understanding the signalling events following platelet activation may elucidate potential drug targets for the treatment of coronary thrombosis. PMID- 12112844 TI - Analysis of N-acetylglucosamine metabolism in the marine bacterium Pirellula sp. strain 1 by a proteomic approach. AB - Pirellula sp. strain 1 is a marine bacterium that can grow with the chitin monomer N-acetylglucosamine as sole source of carbon and nitrogen under aerobic conditions, and that is a member of the bacterial phylum Planctomycetes. As a basis for the proteomic studies we quantified growth of strain 1 with N acetylglucosamine and glucose, revealing doubling times of 14 and 10 h, respectively. Studies with dense cell suspensions indicated that the capacity to degrade N-acetylglucosamine and glucose may not be tightly regulated. Proteins from soluble extracts prepared from exponential cultures grown either with N acetylglucosamine or glucose were separated by two-dimensional gel electrophoresis and visualized by fluorescence staining (Sypro Ruby). Analysis of the protein patterns revealed the presence of several protein spots only detectable in soluble extracts of N-acetylglucosamine grown cells. Determination of amino acid sequences and peptide mass fingerprints from tryptic fragments of the most abundant one of these spots allowed the identification of the coding gene on the genomic sequence of Pirellula sp. strain 1. This gene showed similarities to a dehydrogenase from Bacillus subtilis, and is closely located to a gene similar to glucosamine-6-phosphate isomerase from B. subtilis. Genes of two other proteins expressed during growth on N-acetylglucosamine as well as on glucose were also identified and found to be similar to a glyceraldehyde-3 phosphate-dehydrogenase and a NADH-dehydrogenase, respectively. Thus the coding genes of three proteins expressed during growth of Pirellula sp. strain 1 on carbohydrates were identified and related by sequence similarity to carbohydrate metabolism. PMID- 12112845 TI - Proteomics analysis of the neurodegeneration in the brain of tau transgenic mice. AB - Protein tau, a major microtubule-binding protein in the brain, comprises six isoforms generated through alternative mRNA splicing. A dysfunctional form of mutant and normal tau is associated or implicated in the pathogenesis of several neurodegenerative disorders. The neuropathological hallmark of these tau-opathies are intraneuronal depositions of fibrillary aggregates of which neurofibrillary tangles are most common. Several distinct transgene mouse models confirmed that tau protein can cause neurodegeneration directly. This study was aimed at identifying proteins that might play a role in the cellular disturbances caused by overexpression of the longest isoform of human tau in the brain of transgenic mice. We found 34 proteins which differed in integrated intensity by a factor of at least 1.5. These proteins could be sorted into several categories. Some of the phenotypic characteristics found in the htau transgenic mice could be related to proteins found in this study. Several proteins are linked to processes involving apoptosis and neuronal death and have been discussed in papers describing neurodegenerative disorders. PMID- 12112846 TI - Proteome analysis of rat hippocampal neurons by multiple large gel two dimensional electrophoresis. AB - The goal of the present study was to detect as many protein spots as possible in mammalian cells using two-dimensional gel electrophoresis (2-DE). For proteome analysis, it is of importance to reveal as many proteins as possible. A single standard 2-DE gel (pH 3-10, 18 cm x 20 cm, 13.5% gel) could detect 853 spots from proteins of cultured rat hippocampal neurons when visualized by silver staining. To increase the resolution of the separation and the number of detectable proteins by 2-DE, we utilized seven different narrow pH range immobilized pH gradients in the first dimension. In the second dimension, fourteen long SDS polyacrylamide gels were used: seven 7.5% gels for the separation of high molecular mass proteins (> or = 40 kDa) and seven 13.5% gels for the separation of low molecular mass proteins (< or = 40 kDa). Three hundred and sixty microg of proteins from cultured hippocampal neurons were loaded on to individual gels and visualized by silver staining. All 14 gel images were assembled into a 70 cm x 67 cm cybergel that contained 6677 protein spots, thereby indicating that the utilization of the present strategy led to a 783% increase in the number of detected spots in comparison to the standard procedure. Loading double the amount (720 microg) of proteins on to a 13.5% gel led to a 184% increase in the number of detected spots, thereby indicating that the present strategy has a potential to display more protein spots in the cybergels. PMID- 12112847 TI - Solubilization, two-dimensional separation and detection of the cardiac myofilament protein troponin T. AB - Proteomics strongly relies on the separation of complex protein mixtures, with two-dimensional electrophoresis (2-DE) being a commonly used technique. However efficient separation requires adequate solubilization of the original sample which will determine whether all proteins are accurately represented. Cardiac muscle has presented a particular challenge to solubilization. Here we have optimized the solubilization, separation and detection of the myofilament protein troponin T (TnT). Human left ventricular tissue from a rejected donor transplant heart was homogenized under 19 different conditions and subjected to 2-DE and Western blot analysis for TnT. The optimal conditions for isoelectric focusing of intact TnT requires homogenization in 6 M urea, 2.5 M thiourea, 4% 3-[(3 cholamidopropyl)dimethylamino]-1-propanesulfonate, with the addition of NaCl (2.5 M final concentration). TnT degradation products present in this severely damaged heart however, were differentially extracted from both each other and the intact molecule under the various conditions used. Despite adequate focusing of TnT it was found that a nonglutaraldehyde silver staining protocol, that is compatible with subsequent mass spectrometry, has greatly reduced sensitivity for TnT compared to Coomassie blue. Thus, care is required to avoid misrepresentation of troponin T in proteomic analysis in cardiac tissue. PMID- 12112848 TI - Two-dimensional electrophoresis of recombinant human erythropoietin: a future method for the European Pharmacopoeia? AB - Quality assurance of recombinant protein drugs concerning identity and purity represents a difficult task, in particular, when post-translational modifications lead to a heterogeneous mixture of biomolecules. We chose Neorecormon (rh-EPO, Roche) for our studies to demonstrate the efficiency of two-dimensional electrophoresis (2-DE) to analyse post-translationally modified recombinant drugs. More than 40 protein spots in the range from isoelectric point (pI) 3.5 4.5 and 32-45 kDa could be separated. Enzymatic deglycosylation revealed that the heterogeneity of the protein pattern is mainly caused by variations in glycosylation. In comparison to the separately performed isoelectric focusing and sodium dodecyl sulfate-polyacrylamide gel electrophoresis, as requested by the European Pharmacopoeia, we see a great synergy to use 2-DE for the analysis of rh EPO. A by far higher resolution can be achieved, allowing an improved differentiation of the various rh-EPO glycoforms. Sequential deglycosylation of sialic acids, N-glycosides and the O-glycoside lead to significant shifts both in apparent relative molecular mass and pI. Comparing the 2-DE patterns of rh-EPO before and after deglycosylation allows on the one hand valuable information to be gained on the glycosylation of the recombinant protein and shows on the other hand how significantly the 2-DE protein pattern can be influenced by the glycosylation. As the equipment for the performance of 2-DE has improved significantly over the last decade, we see 2-DE as a reliable method, which should be approved for the routine quality assurance of recombinant drugs and also recommended for the European Pharmacopoeia. PMID- 12112849 TI - Bronchoalveolar lavage protein patterns in children with malignancies, immunosuppression, fever and pulmonary infiltrates. AB - Severe respiratory infections are a major cause of morbidity and mortality in children receiving immunosuppressive therapy for malignancies. The goal of this study was to assess the major changes in the protein patterns in these children. Bronchoalveolar lavage (BAL) fluids of seven control children and of ten children with malignancies and fever not responding to broad spectrum antibiotic treatment was separated by horizontal two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis in the isoelectric point range 3-9. We observed a large increase of alpha(1)-antitrypsin (p = 0.0004) and decreases of the immunoglobulin (Ig) binding factor, transthyretin and cystatin S. Significant changes occurred also in the small acidic proteins. The relative abundance of the IgG heavy and light chains may hinder the separation and identification of many minor protein spots located in the basic area of the gel, suggesting that their removal during sample preparation may be warranted. This study demonstrated significant alterations in BAL fluid proteome in immunosuppressed children with persistent fever and pulmonary infiltrates. Future target regions of interest were identified. Sample prefractionation and the selection of suitable narrow isoelectric point ranges will be necessary for optimized detection and separating conditions. PMID- 12112850 TI - Protein pattern of exhaled breath condensate and saliva. AB - The proteins recovered in exhaled breath condensate (EBC) might be used to non invasively monitor respiratory diseases. However, the range of proteins and their source are still unresolved and contamination by saliva or a similar protein pattern in the nasal and bronchial compartments may make interpretation of the data difficult. We studied nasal EBC (collected through a "free of touch" technique by negative pressure), oral tidal, and oral forced EBC (collected through a rebreathing valve as a saliva trap connected to tubing submerged into ice) and matched saliva samples from five healthy adult subjects. The protein samples were separated by two-dimensional electrophoresis and the silver stained gels were analyzed by Melanie 2 software. In both nasal and oral EBC, three spots (72 kDa/isoelectric point (pI) 6.6-7.0, 66 kDa/pI 5.9-6.7 and 45-48 kDa/pI 8.0 8.6) were consistently present in all subjects. Several other proteins were only sporadically detected. Despite improbable saliva contamination (no phosphorus contamination in the same oral and nasal EBC, no amylase activity in 10 pairs of nasal and oral EBC collected by the same technique), on average 63% and 71% of the spots identified in oral and nasal EBC were also found in the matched individual saliva samples. Compared to saliva, the range and amount of protein in all types of EBC was very small. Even when collected free of saliva contamination the majority of proteins present in EBC was also found in saliva, suggesting that these proteins are present in both compartments, e.g. saliva and secretions of the lower airspaces. The quantification and identification of specific proteins in the various compartments is warranted in future studies to determine the practical value of EBC. PMID- 12112851 TI - Monitoring daunorubicin-induced alterations in protein expression in pancreas carcinoma cells by two-dimensional gel electrophoresis. AB - Tumors of the pancreas are characterized by a high intrinsic potency to develop chemoresistance towards cytotoxic drugs, which is the main cause of ineffective treatment. The phenomenon of multidrug resistance is known to be a multifactorial event in which several mechanisms act simultaneously. We investigated the response of pancreas tumor cells after exposure to the anthracycline daunorubicin (DRC), a well-known antitumor agent in chemotherapy, by two-dimensional gel electrophoresis (2-DE). DRC is known to cause DNA damage and to affect tumor cell growth. Importantly, we aimed at investigating alterations in the protein expression pattern after first contact of the tumor cells with DRC, thus simulating a situation close to clinical chemotherapy and elucidating cell survival strategies following initial drug exposure. A concentration dependent up regulation of a variety of proteins was observed, indicating that cell response to DRC involves multiple signaling events. Since the p53 tumor suppressor is essentially involved in the regulation of cell growth and controlled cell death (apoptosis) after cellular stress (like DNA damage), we investigated the role of p53 in DRC-resistant and -sensitive pancreas carcinoma cells by measuring p53 transcriptional transactivation activities. No differences in p53 activities were observed in response to DRC treatment in both pancreas cell lines, whereas mamma carcinoma cells (MCF-7), possessing wild-type p53, demonstrated the expected increase in p53 transcriptional transactivation activity. Hence, the tested pancreas carcinoma cells harbor a mutant, nonfunctional p53. We additionally analyzed the steady state protein levels of the cyclin dependent kinase inhibitor p21(CIP1), which is known to be involved in cell cycle control. Interestingly, p21(CIP1 )was induced by DRC in sensitive cells in a concentration dependent manner and was highest in resistant cells. In conclusion, our results suggest that the induction of proteins by DRC in pancreas carcinoma cells, as observed by 2-DE, occurs independently from p53 signaling events, but is probably associated with increased levels of p21(CIP1). PMID- 12112852 TI - Detection of cathepsin B up-regulation in neoplastic thyroid tissues by proteomic analysis. AB - Nodular or multinodular goiter is the most common non-neoplastic thyroid disease and may be difficult to distinguish from true neoplastic thyroid diseases using microscopic criteria. We have used two-dimensional gel electrophoresis to study the protein patterns of thyroid tissues including normal thyroid, multinodular goiter, diffuse hyperplasia, follicular adenoma, follicular carcinoma and papillary carcinoma. Specific proteins, in the region of molecular mass 15-30 kDa and isoelectric point 4.5-6.5, were identified by electrospray tandem mass spectrometry and protein sequencing. The most distinctive protein found is cathepsin B, which could be detected as four spots, with differential expression in different thyroid diseases. In particular, two of these cathepsin B spots CB2 and CB3 are strongly up-regulated in neoplastic diseases, compared to non neoplastic diseases. In addition, overexpression of ATP synthase D chain and prohibitin were observed in papillary carcinoma, which should allow it to be differentiated from follicular carcinoma. Changes in expression of other proteins were also observed in disease states compared to normal tissues, namely translationally controlled tumor protein, thioredoxin peroxidase 1, glutathione-S transferase P, DJ-1 protein, superoxide dismutase (Cu, Zn), and heat shock protein 27, but these changes are less characteristic, so they do not allow the differentiation between neoplastic and non-neoplastic tissues. Thus, the proteomic approach is a useful diagnostic tool for studying diseases involving the thyroid nodule. PMID- 12112853 TI - Comparison of one-dimensional and two-dimensional gel electrophoresis as a separation tool for proteomic analysis of rat liver microsomes: cytochromes P450 and other membrane proteins. AB - Gel electrophoresis in combination with peptide mass fingerprinting is the method of choice for proteomic profiling of various in vitro and in vivo biological systems. In the investigation reported here we analyzed the protein composition of hepatic microsomes from untreated and phenobarbital treated rats, using one dimensional (1-DE) and two-dimensional (2-DE) gel electrophoresis, followed by tryptic peptide mapping. To better characterize capabilities of 2-DE 1-DE with regard to microsomal membrane proteins, "ghosts" of microsomal vesicles enriched in membrane proteins were obtained and analyzed. Both 1-DE and 2-DE showed that phenobarbital induces not only cytochromes P450 2B1and 2B2 but such stress related endoplasmic reticulum proteins as protein disulfide isomerase A(3) and A(6) and 78 kDa glucose regulated protein. The analytical performance of 1-DE with regard to endoplasmic reticulum membrane proteins is incomparably greater than that of 2-DE. Twenty-two out of a total of thirty-four known to date microsomal rat membrane proteins were identified by 1-DE in combination with matrix-assisted laser desorption/ionization-mass spectrometry of in-gel digests. At the same time using various types of 2-DE, we were able to identify only three rat microsomal membrane proteins. The data presented in this manuscript clearly demonstrate that 1-DE in combination with peptide mass fingerprinting can be successfully used for cataloging proteins of the endoplasmic reticulum, and that the proteomic analysis of the subcellular organelles containing a considerable number of highly hydrophobic membrane proteins should be performed by combined application of 1-D and 2-D electrophoresis. PMID- 12112854 TI - Identification of caseins in goat milk. AB - The importance of goat milk in infant diet is growing, because it is reported that goat's milk in some cases is less allergenic than cow's milk. This is due probably to the lower presence of caseins associated with a specific type of alpha(s1)-casein. In caprine breeds, four types of alpha(s1)-casein alleles are identified and associated with various amounts of this protein in milk. The contribution of strong alleles to the goat milk is approximately 3.6 g/L of alpha(s1)-casein, while for middle alleles is only 1.6 g/L, weak alleles 0.6 g/L. The contribution of null allele is very low (or non-existent). The quantity of total caseins in caprine milk is positively correlated with the amount of alpha(s1)-casein. Milk from animals possessing strong alleles contain significantly more total caseins than milk from animals without those alleles. This is important because animals with mild alleles can be employed to produce milk for allergic subjects while the other animals can be used to produce milk for the dairy industry. This work shows casein profiles of two types of classified goat milk (B, strong alpha(s1) allele, 0, null alpha(s1) allele) with two-dimensional electrophoresis coupled with matrix-assisted laser desorption/ionization-time of flight mass spectrometry, and it confirms the different polymorphisms at locus alpha(s1) casein. PMID- 12112855 TI - Web-based two-dimensional database of Saccharomyces cerevisiae proteins using immobilized pH gradients from pH 6 to pH 12 and matrix-assisted laser desorption/ionization-time of flight mass spectrometry. AB - An image based two-dimensional (2-D) reference map of very alkaline yeast cell proteins was established by using immobilized pH gradients (IPG) up to pH 12 (IPG 6-12, IPG 9-12 and IPG 10-12) for 2-D electrophoresis and by using matrix assisted laser desorption/ionization-time of flight mass spectrometry peptide mass fingerprinting for spot identification. Up to now 106 proteins with theoretical isoelectric points up to pH 11.15 and molecular mass between 7.5 and 115 kDa were localized and identified. Additionally, due to the improved resolution of steady-state isoelectric focussing with IPGs, even low copy number proteins with codon bias below 0.02 were detected and identified. PMID- 12112856 TI - Initial proteome analysis of mature barley seeds and malt. AB - Several barley (Hordeum vulgare) cultivars are used in the production of malt for brewing. The malt quality depends on the cultivar, its growth and storage conditions, and the industrial process. To enhance studies on malt quality, we embarked on a proteome analysis approach for barley seeds and malt. The proteome analysis includes two-dimensional (2-D) gel electrophoresis, mass spectrometry, and bioinformatics for identification of selected proteins. This project initially focused on proteins in major spots in the neutral isoelectric point range (pI 4-7) including selected spots that differ between four barley cultivars. The excellent malting barley cultivar Barke was used as reference. Cultivar differences in the 2-D gel spot patterns are observed both at the seed and the malt level. In seed extracts one of the proteins causing variations has been identified as an alpha-amylase/trypsin inhibitor. In malt extracts multiple forms of the alpha-amylase isozyme 2 have been identified in varying cultivar characteristic spot patterns. The present identification of proteins in major spots from 2-D gels includes 27 different proteins from 42 spots from mature seed extract, while only three specific proteins were identified by analysing 13 different spots from the corresponding malt extract. It is suggested that post translational processing causes the same protein to occur in different spots. PMID- 12112857 TI - Identification of Staphylococcus aureus exotoxins by combined sodium dodecyl sulfate gel electrophoresis and matrix-assisted laser desorption/ ionization-time of flight mass spectrometry. AB - Staphylococcus aureus is an important human pathogen whose pathogenesis involves the synthesis of cell wall associated virulence factors and secreted toxins with damaging effects on the host cells. Most of these pathogenic factors are synthesized in a growth-phase dependent manner as a response to environmental stress like heat, lack of nutrients or other deleterious conditions. Conventional identification of these pathogenic factors is based on Western blot analysis or enzyme-linked immunosorbent assay (ELISA) and is limited by the commercial availability of antibodies against these toxins. We report here the use of matrix assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry for monitoring the pathogenic factors of S. aureus. For the identification of pathogenic factors, a methicillin sensitive strain of S. aureus, ATCC-29213, was grown at 37 degrees C or 42 degrees C in brain-heart infusion broth and harvested during the early stationary phase of growth. Secreted proteins were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, enzymatically digested with trypsin and analyzed by MALDI-TOF mass spectrometry. When grown at 42 degrees C, alpha- and beta-hemolysins were found to accumulate in S. aureus supernatants while the concentration of protein A was slightly decreased. The identity of some of these toxins was confirmed by Western-blot analysis. MALDI TOF mass spectrometry combined with sodium dodecyl sulfate gel electrophoresis represents a rapid and simple approach to characterize the virulence of S. aureus strains which seems to be particularly valuable for the identification of S. aureus exotoxins for which ELISA is not established. PMID- 12112858 TI - Identification and discrimination of Staphylococcus aureus strains using matrix assisted laser desorption/ionization-time of flight mass spectrometry. AB - Staphylococcus aureus is an important human pathogen frequently resistant to a wide range of antibiotics. Methicillin-resistant S. aureus (MRSA) strains are common nosocomial pathogens that pose a world-wide problem. Rapid and accurate discrimination between methicillin-sensitive S. aureus (MSSA) and methicillin resistant S. aureus is essential for appropriate therapeutic management and timely intervention for infection control. We report here the application of matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) for monitoring the bacterial fingerprints expressed by two well characterized S. aureus strains ATCC 29213 (MSSA) and ATCC 43330 (MRSA). Consistent strain-specific data were obtained from subcultures analyzed over a period of three months as well as after changing the growth media from Mueller Hinton to blood agar indicating the reliability of the method. The bacterial fingerprints of these two strains were compared to independent clinical isolates of S. aureus. A uniform signature profile for MRSA could not be identified. However, the bacterial fingerprints obtained proved to be specific for any given strain. This study demonstrates that MALDI-TOF MS is a powerful method for rapid identification of clonal strains of S. aureus, which might be useful for tracking nosocomial outbreaks of MRSA and for epidemiologic studies of infections diseases in general. PMID- 12112859 TI - The proteome of the bacterium Mycoplasma pneumoniae: comparing predicted open reading frames to identified gene products. AB - An existing proteome map of the bacterium Mycoplasma pneumoniae comprising proteins from 224 genes was extended to 305 genes. This corresponds to about 44% of the 688 proposed genome sequence derived open reading frames (ORFs). The newly assigned gene products were enriched, separated by one-dimensional or two dimensional (2-D) gel electrophoresis and identified by mass spectrometry. The enrichment procedures included differential centrifugation, anion and cation exchange chromatography, affinity chromatography with heparin as a ligand and isolation of biotinylated proteins by binding to immobilized streptavidin. A comparative analysis of the identified proteins from 305 genes with the as yet unverified 383 ORFs concerning isoelectric point, molecular weight and number of transmembrane segments revealed that proteins with more than three predicted transmembrane segments and an isoelectric point above 10.5 are most likely not to be separated by 2-D gel electrophoresis. The mutual benefits of genomics and proteomics were shown by the identification of a todate unannotated 128 amino acid long protein. PMID- 12112860 TI - High pressure effects step-wise altered protein expression in Lactobacillus sanfranciscensis. AB - In this study we investigated the cellular response to the application of high hydrostatic pressure. High pressure is increasingly used for food preservation. With high resolution 2-D electrophoresis we compared the protein patterns of atmospherically grown Lactobacillus sanfranciscensis with those pressure treated up to 200 MPa. We performed the comparative study by using overlapping immobilized pH gradients covering the pH range from 2.5 up to 12 in order to maximize the resolution for the detection of stress relevant proteins. For improved quantitative analysis, staining with SyproRuby was used in addition to silver staining. By computer aided image analysis we detected more than a dozen spots within the pH range from 3.5 to 9 that were more than two-fold increased or 50% decreased in their intensity upon high pressure treatment. Two of them (approx. values: pI 4.0 and 4.2, respectively; M(r) approximately 15 000) have almost identical matrix-assisted laser desorption/ionization-time of flight mass spectrometry spectra and were identified by liquid chromatography-tandem mass spectrometry as putative homologs/paralogs to cold shock proteins of Lactococcus lactis. Their expression is opposed (i.e. the more acidic one is repressed, while the other one is induced); this effect is maximal at 1 h, 150 MPa. It was further remarkable that by monitoring the barosensitivity of the cells within 25 MPa steps, we observed a differential pressure induction or repression of the detected proteins as well. For example one protein (approx. values: pI 4.2, M(r) approximately 15 000) shows a maximum induction after 1 h, 150 MPa while another one (pI 7.5, M(r) approximately 25 000) is maximally induced after 1 h, 50/75 MPa. This indicates a successive cell response and different signalling pathways for these responses. PMID- 12112861 TI - Immunogenic proteins of Helicobacter pullorum, Helicobacter bilis and Helicobacter hepaticus identified by two-dimensional gel electrophoresis and immunoblotting. AB - The ecological niches occupied by various species of Helicobacter are not yet known and the full spectrum of diseases associated with Helicobacter infections are not yet defined. Since these fastidious microaerofilic bacteria require special growth conditions new and improved molecular and serologic diagnostic methods have been developed to increase our understanding of their pathogenesis and virulence characteristics. Immunogenic cell surface proteins of Helicobacter pullorum, Helicobacter bilis, and Helicobacter hepaticus were characterised by proteomic techniques using two-dimensional electrophoresis and immunoblotting with antisera from immunised rabbits. Cross-reactivity between the three Helicobacter species were analysed after a four-step cross-absorption experiment. For H. pullorum, H. bilis and H. hepaticus 21, 13 and 27 specific immunogenic proteins, respectively, were identified. These proteins could be of important sero-diagnostic value for analyses of sera from humans, laboratory animals and for the veterinarian field. PMID- 12112862 TI - Two-dimensional electrophoresis analysis of the extracellular proteome of Bacillus cereus reveals the importance of the PlcR regulon. AB - Many virulence factors are secreted by the gram-positive, spore forming bacterium Bacillus cereus. Most of them are regulated by the transcriptional activator, PlcR, which is maximally expressed at the beginning of the stationary phase. We used a proteomic approach to study the impact of the PlcR regulon on the secreted proteins of B. cereus, by comparing the extracellular proteomes of strains ATCC 14579 and ATCC 14579 Delta plcR, in which plcR has been disrupted. Our study indicated that, quantitatively, most of the proteins secreted at the onset of the stationary phase are putative virulence factors, all of which are regulated, directly or indirectly, by PlcR. The inactivation of plcR abolished the secretion of some of these virulence factors, and strongly decreased that of others. The genes encoding proteins that are not secreted in the DeltaplcR mutant possessed a regulatory sequence, the PlcR box, upstream from their coding sequence. These proteins include collagenase, phospholipases, haemolysins, proteases and enterotoxins. Proteins for which the secretion was strongly decreased, but not abolished, in the DeltaplcR mutant did not display the PlcR box upstream from their genes. These proteins include flagellins and InhA2. InhA2 is a homologue of InhA, a Bacillus thuringiensis metalloprotease that specifically degrades antibacterial peptides. The mechanism by which PlcR affects the production of flagellins and InhA2 is not known. PMID- 12112863 TI - Proteomic approaches to Salmonella Pathogenicity Island 2 encoded proteins and the SsrAB regulon. AB - Type III protein secretion is a common virulence determinant in Gram-negative bacteria and the genetic information is often clustered in pathogenicity islands or on virulence plasmids. We have analyzed the type III secretion system encoded by Salmonella Pathogenicity Island 2 (SPI2) that is indispensable for systemic disease of Salmonella enterica serotype Typhimurium (S. Typhimurium) in mice. Since the low abundance of this secretion system restricted direct analysis by proteomic approaches, several putative proteins were expressed as recombinant products and analyzed by two-dimensional electrophoresis. The map obtained for SPI2 encoded proteins was correlated to the expression pattern of S. Typhimurium. The latter was compared to the proteins induced by SsrAB, the two-component system regulating SPI2 gene expression. Our results exemplify that recombinant expression is a complementary tool for analysis of low abundant proteins or membrane proteins. This approach contributes to the characterization of these proteins by subcellular fractionation. Furthermore, we show that pulse labeling was necessary to analyze growth phase regulated SPI2 proteins that might not be otherwise detectable. PMID- 12112866 TI - Conjugated macrocycles as active materials in nonlinear optical processes: optical limiting effect with phthalocyanines and related compounds. AB - An overview of the optical limiting (OL) processes in phthalocyanines and related compounds is presented, particularly a description of the synthesis and relevant optical properties of a series of axially substituted indium(III), titanium(IV), phthalo- and naphthalocyanines, and octaarylporphyrazines. Several techniques, such as transient absorption, Z-scan, and degenerate four-wave mixing, were used to assess the optical properties and OL performance of the investigated compounds. The versatility of the methods of organic synthesis leads to the achievement of effective systems in terms of OL performance through the appropriate combination and modulation of several structural components. The chemistry of the macrocycles here considered allows the variation of the different chemical features, such as the degree of electronic conjugation of the macrocycle and the nature of the ring substituents, the central atom, and the ligands attached to the central atom. PMID- 12112867 TI - Catalyzing "hot" reactions: enzymes from hyperthermophilic Archaea. AB - We reflect on some of our studies on the hyperthermophilic archaeon, Thermococcus kodakaraensis KOD1 and its enzymes. The strain can grow at temperatures up to the boiling point and also represents one of the simplest forms of life. As expected, all enzymes displayed remarkable thermostability, and we have determined some of the basic principles that govern this feature. To our delight, many of the enzymes exhibited unique biochemical properties and novel structures not found in mesophilic proteins. Here, we focus on a few enzymes that are useful in application, and whose three-dimensional structures are characteristic of thermostable enzymes. PMID- 12112868 TI - Sphericity governs both stereochemistry in a molecule and stereoisomerism among molecules. AB - The concept of sphericity and relevant fundamental concepts that we have proposed have produced a systematized format for comprehending stereochemical phenomena. Permutability of ligands in conventional approaches is discussed from a stereochemical point of view. After the introduction of orbits governed by coset representations, the concepts of subduction and sphericity are proposed to characterize desymmetrization processes, with a tetrahedral skeleton as an example. The stereochemistry and stereoisomerism of the resulting promolecules (molecules formulated abstractly) are discussed in terms of the concept of sphericity as a common mathematical and logical framework. Thus, these promolecules are characterized by point group and permutation group symmetry. Prochirality, stereogenicity, prostereogenicity, and relevant topics are described in terms of the concept of sphericity. PMID- 12112869 TI - Chiral proton donor reagents: tin tetrachloride--coordinated optically active binaphthol derivatives. AB - The Lewis acid-assisted chiral Bronsted acids (chiral LBAs), which are prepared from tin tetrachloride and optically active binaphthol derivatives, are highly effective chiral proton donor reagents for enantioselective protonation and biomimetic polyene cyclization. These chiral LBAs can directly protonate various silyl enol ethers and ketene disilyl acetals to give the corresponding alpha-aryl or alpha-halo ketones and alpha-arylcarboxylic acids, respectively, with high enantiomeric excess (up to 98% ee). A catalytic version of enantioselective protonation was also achieved using stoichiometric amounts of 2,6-dimethylphenol and catalytic amounts of monomethyl ether of optically active binaphthol in the presence of tin tetrachloride. The biomimetic cyclization of simple isoprenoids to polycyclic isoprenoids using chiral LBA is also described. This is the first example of a chiral Bronsted acid-induced enantioselective ene cyclization in synthetic chemistry. Geranyl phenyl ethers, o-geranylphenols, and homogeranylphenol derivatives were directly cyclized in the presence of (R) binaphthol derivatives and tin tetrachloride (up to 90% ee). Compounds bearing a farnesyl group could also be cyclized under the same conditions to give the natural products (-)-ambrox((R)) and (-)-chromazonarol, and (-)-tetracyclic polyprenoids of sedimentary origin. These chiral LBAs recognize the prochiral face of a trisubstituted terminal olefin and site selectively generate carbocations on the substrates. PMID- 12112870 TI - The art of acetylenic scaffolding: rings, rods, and switches. AB - Acetylenic scaffolding with derivatives of tetraethynylethene (TEE, 3,4 diethynylhex-3-ene-1,5-diyne) and (E)-1,2-diethynylethene (DEE, (E)-hex-3-ene-1,5 diyne) provides carbon-rich compounds with interesting physicochemical properties. Thus, these modules are building blocks for monodisperse, linearly pi conjugated oligomers [polytri(acetylene)s, PTAs] extending in length beyond 10nm, and for large, macrocyclic, all-carbon cores (dehydroannulenes and expanded radialenes) exhibiting strong chromophoric properties. The advanced materials' properties were strongly influenced by the presence of electron-donating substituents at the lateral positions, decreasing the decreasing the (HOMO-LUMO) gap in both PTAs and expanded radialenes. Arylated TEEs were found to undergo photochemically induced cis-trans isomerization, paving the way for applications as light-driven molecular switches in optoelectronic devices. Derivatives of 1,3 diethynylallene are new modules that offer the prospect of scaffolding in an orthogonal manner; that is, they represent precursors for helical oligomers. PMID- 12112871 TI - Direct measurements of HOx radicals in the marine boundary layer: testing the current tropospheric chemistry mechanism. AB - OH and HO(2) radicals, atmospheric detergents, and the reservoir thereof, play central roles in tropospheric chemistry. In spite of their importance, we had no choice but to trust their concentrations predicted by modeling studies based on known chemical processes. However, recent direct measurements of these radicals have enabled us to test and revise our knowledge of the processes by comparing the predicted and observed values of the radical concentrations. We developed a laser-induced fluorescence (LIF) instrument and successfully observed OH and HO(2) at three remote islands of Japan (Oki Island, Okinawa Island, and Rishiri Island). At Okinawa Island, the observed daytime level of HO(2) agreed closely with the model estimates, suggesting that the photochemistry at Okinawa is well described by the current chemistry mechanism. At Rishiri Island, in contrast, the observed daytime level of HO(2) was consistently much lower than the calculated values. We proposed that iodine chemistry, usually not incorporated into the mechanism, is at least partly responsible for the discrepancy in the results. At night, HO(2) was detected at levels greater than 1 pptv at all three islands, suggesting the presence of processes in the dark that produce radicals. We showed that ozone reactions with unsaturated hydrocarbons, including monoterpenes, could significantly contribute to radical production. PMID- 12112873 TI - Novel Sxr(a) ES cell line offers hope for Y chromosome gene-targeted mice. AB - A mouse targeted for a Y Chromosome gene has not been reported. Because the Y Chromosome is present in only one copy, and most of its genes are critical for germ cell development, such a mouse would likely be infertile. Thus, we describe a new reproductive strategy to enable transmission of targeted Y Chromosome genes to subsequent generations. The strategy uses two segregating copies of Y Chromosome genes to mimic the autosomal condition. To achieve this, we developed a new embryonic stem cell line from the XYSxr(a) mouse, which carries a duplication of the gene-rich Y Chromosome short arm. Importantly, we demonstrate germ line transmission of the YSxr(a) chromosome and describe this significant new tool as a practical solution to enable reproduction in mice targeted for Y Chromosome genes. PMID- 12112872 TI - A new allele of Gli3 and a new mutation, circletail (Crc), resulting from a single transgenic experiment. AB - While generating transgenic lines, transgene-linked mutations can occur, which are caused by an insertional mutation at a given locus. More rarely, mutations unlinked to the transgene insertion site are observed. In the process of generating a mouse overexpressing the enzyme tyrosinase, we have obtained one transgenic line that appears to carry a semidominant insertional mutation at the Gli3 (extra toes) locus, characterized by polydactyly and skeletal malformations. Additionally, the transgenic line contained a second mutation, Crc (circletail), which appears to be unlinked to the transgene insertion site. Heterozygous Crc mice are incompletely penetrant for a circled-tail phenotype, while all homozygous Crc/Crc mice die at birth of a severe neural tube defect (craniorachischisis). Anatomical evidence from a Crc/Crc; Gli3/+ fetus indicates that these two genes may interact. PMID- 12112874 TI - DEAF-1 function is essential for the early embryonic development of Drosophila. AB - The Drosophila protein DEAF-1 is a sequence-specific DNA binding protein that was isolated as a putative cofactor of the Hox protein Deformed (Dfd). In this study, we analyze the effects of loss or gain of DEAF-1 function on Drosophila development. Maternal/zygotic mutations of DEAF-1 largely result in early embryonic arrest prior to the expression of zygotic segmentation genes, although a few embryos develop into larvae with segmentation defects of variable severity. Overexpression of DEAF-1 protein in embryos can induce defects in migration/closure of the dorsal epidermis, and overexpression in adult primordia can strongly disrupt the development of eye or wing. The DEAF-1 protein associates with many discrete sites on polytene chromosomes, suggesting that DEAF 1 is a rather general regulator of gene expression. PMID- 12112875 TI - Expression of Cre Recombinase in the developing mouse limb bud driven by a Prxl enhancer. AB - We have used a Prx1 limb enhancer to drive expression of Cre Recombinase in transgenic mice. This regulatory element leads to Cre expression throughout the early limb bud mesenchyme and in a subset of craniofacial mesenchyme. Crossing a murine line carrying this transgene to a reporter mouse harboring a floxed Cre reporter cassette revealed that recombinase activity is first observed in the earliest limb bud at 9.5 dpc. By early to mid bud stages at 10.5 dpc recombination is essentially complete in all mesenchymal cells in the limb. Expression of the Cre recombinase was never detected in the limb bud ectoderm. The use of Prx1-Cre mice should facilitate analysis of gene function in the developing limb. PMID- 12112876 TI - Improved mRNA electroporation method for Xenopus neurula embryos. AB - Electroporation has led to new approaches to the analysis of gene regulation of the chick embryonic system. However, application of this method to Xenopus, another model organism of embryology, has left many difficulties to be overcome. The specially devised electrodes, the examination of luciferase activities expressed, and the direct visualization of green fluorescence protein allow us to optimize the conditions of electroporation for Xenopusembryos. The use of mRNA rather than DNA improved the expression efficiency 120 times more than for the case of plasmid DNA, and the effect emerged more immediately after electroporation. The noncontact electroporation adopted here caused less damage to cells and tissues than with the needle type electrode, making it practical for efficient application to early embryos. Furthermore, the mRNA electroporation technique is applicable for other systems in which the DNA electroporation has not had any significant effect because of its low expression efficiency. PMID- 12112877 TI - Axes establishment during eye morphogenesis in Xenopus by coordinate and antagonistic actions of BMP4, Shh, and RA. AB - We have examined the roles of BMP4, Shh, and retinoic acid in establishing the proximal-distal and dorsal-ventral axes in the developing Xenopus eye. Misexpression of BMP4 caused the absence of an optic stalk and the expansion of dorsal and distal markers, tbx2/3/5, and pax6, at the expense of ventral and proximal markers vax2 and pax2. When Shh or Noggin, an antagonist of BMPs, was misexpressed, the reverse expression patterns of these marker genes were observed. These results suggest that BMP4 is involved in the specification of not only dorsal in the optic cup but also distal in the optic vesicle. Because Shh did not suppress bmp4 expression, unlike Noggin, Shh and BMP4 may antagonistically regulate common downstream genes in developing eye. We also found the difference between the effects of Shh and retinoic acid, another possible ventralizing factor, suggesting that Shh could promote ventralization independently of retinoic acid. These findings provide important clues to the coordinate and antagonistic actions of BMP4, Shh, and retinoic acid in axes specifications of Xenopus eyes. PMID- 12112878 TI - Mouse model of split hand/foot malformation type I. AB - Split hand/foot malformation type I (SHFM1) disease locus maps to chromosome 7q21.3-q22, a region that includes the distal-less-related (dll) genes DLX5 and DLX6. However, incomplete penetrance, variable expressivity, segregation distortion, and syndromic association with other anomalies have so far prevented the identification of the SHFM1 gene(s) in man. Here we show that the targeted double inactivation of Dlx5 and Dlx6 in the mouse causes in homozygous mutant animals bilateral ectrodactyly with a severe defect of the central ray of the hindlimbs, a malformation typical of SHFM1. This is the first evidence that the role of dll/Dlx genes in appendage development is conserved from insects to mammals and proves their involvement in SHFM1. PMID- 12112879 TI - Acute and six-month clinical outcome after Helistent stent implantation in coronary arteries: results of the French Helistent multicenter registry. AB - The purpose of our study was to determine the feasibility, safety, and efficacy of the new tubular (Helistent) stent (Hexacath, France) in patients with coronary artery disease. Patients were prospectively included in a multicenter French registry: acute results and 6-month target lesion revascularization rate were assessed. A total of 628 stents were implanted in 527 patients (616 lesions). Mean age was 64 +/- 11 years. Predilatation was performed in 77% of cases. The stent was successfully deployed in 99.4% of attempted lesions. Three patients died during the hospital stay and 10 patients developed myocardial infarction. Angiographic stent thrombosis rate was 1.1%. There was no need for urgent cardiac surgery during hospital stay. Six-month clinical follow-up was obtained in 95% of patients; target lesion revascularization rate was 8.3%. Eight patients (1.6%) (extracardiac death in three patients) died and three patients (0.6%) developed myocardial infarction. In conclusion, the Helistent stent appears safe and effective in minimizing acute complications of elective coronary interventions. Six-month major cardiac events were rare and clinical restenosis was low (< 10%). PMID- 12112880 TI - Long-term clinical outcomes after coronary angioplasty using long stents in small coronary vessels. AB - The role of coronary stenting in challenging situations, such as small vessels and long lesions, remains controversial. The aim of this study was to examine the procedural, in-hospital, and long-term clinical outcomes of patients undergoing angioplasty with long stents in small coronary vessels. We evaluated the procedural success rate and clinical outcomes in 252 consecutive subjects treated by means of the implantation of a single coronary stent in vessels with a mean reference diameter of < 2.5 mm; 128 patients received a short stent (< or = 16 mm) and 124 a long stent (> or = 18 mm). Lesion morphology was more complex in patients treated with long stents (P < 0.05). The mean stent length was 14 +/- 2 mm in the short-stent group and 25 +/- 3 mm in the long-stent group (P < 0.001). The overall procedural success rate (98.4% vs. 97.6%; P = NS) and the rate of major in-hospital adverse events (death, acute myocardial infarction, or target vessel revascularization; 1.6% vs. 2.4%; P = NS) was similar in the two groups. After 11.7 +/- 7 months of follow-up, there was no difference in the incidence of mortality and myocardial infarction (5% vs. 6.6%; P = NS), but revascularization tended to occur more frequently in the patients treated with long stents (21.7% vs. 13.9%; P = NS). In conclusion, the procedural success rate of single short or long stents in small coronary vessels was similar. Although the incidence of target vessel revascularization tended to be higher in the patients treated with longer stents, 2-year event-free survival was equivalent in the two groups (65% vs. 70%; P = NS). PMID- 12112881 TI - Does stent design affect the long-term outcome after coronary stenting? AB - The purpose of our study was to determine the clinical, angiographic, and procedural variables that predict the risk for 2-year target lesion revascularization (TLR) and other clinical events in a large cohort of patients treated with coronary stenting. Between March 1996 and March 1999, 1,340 patients were prospectively enrolled. They underwent at least one stenting procedure with one of the four coronary stent types used during this time period in our institution: Wiktor-I, Nir, Bard XT, or Tenax. Clinical follow-up was obtained for 99.1% of eligible patients (mean, 19.38 +/- 4.97 months). The overall TLR rate was 10.7% at 24 months. Two variables were independently associated with the long-term outcome: MLD post stenting and stent type. Implantation of silicon carbide-coated stents was associated with a twofold decrease in 24-month TLR (P < 0.01). The major adverse cardiac event rate at 2 years was 20.8%. Multivariate analysis showed that three parameters were predictive: diabetes (P < 0.002), recent onset of symptoms (P < 0.03), and high diffusion of coronary atherosclerosis as assessed by Gensini score (P < 0.0069). In conclusion, stent type, and particularly passive silicon carbide coating, appears to affect the 24 month TLR rate but not other major cardiac events. PMID- 12112882 TI - Relation of troponin T release kinetics to long-term clinical outcome in patients with acute ST segment elevation myocardial infarction treated with a percutaneous intervention. AB - The purpose of this study was to determine the relation of troponin T release kinetics to long-term clinical outcome in patients with an acute ST segment elevation myocardial infarction treated with a primary percutaneous intervention. One hundred and four patients with typical ischemic chest pain and > 1.5 mm ST segment elevation in > 2 contiguous leads underwent primary stenting (n = 60) or primary percutaneous transluminal coronary angioplasty (n = 44). Serum troponin T concentrations were obtained prior to and serially postintervention for 72 hr. Mean time to peak serum troponin T concentration was significantly longer in patients with cardiac death (P = 0.02), reinfarction (P = 0.007), target lesion reintervention (P = 0.03), and the composite of these events (13.2 +/- 5.3 vs. 9.3 +/- 4.0 hr; P < 0.0005). Multivariate analysis identified age, Killip class > 2, and time to peak serum troponin T concentration as independent predictors of long-term cardiac event-free survival. Thus, time to peak serum troponin T concentration independently predicts long-term cardiac event-free survival in patients with acute ST segment elevation myocardial infarction treated with a primary percutaneous intervention. PMID- 12112883 TI - Retinal embolization during carotid angioplasty and stenting: mechanisms and role of cerebral protection systems. AB - Carotid stenting has become an accepted alternative to endarterectomy, but fear of embolic stroke has impeded its generalized application. The retina provides a unique observatory for the study of emboli, which may occur either directly or indirectly via collaterals to the ophthalmic artery. Systems under development for cerebral protection differ in their capacity to trap small emboli and in their protection of the collateral circulation. We evaluated 118 sequential patients undergoing carotid stenting using fundoscopy, fluorescein retinal angiography, and visual field examination. The site and size of emboli was assessed, and degree of edema estimated. All patients were treated using distal protection during carotid stent implantation: 38 patients with the Theron system (using routine flushing toward the external carotid) and 80 patients with the Percusurge system (aspiration only). Retinal embolization occurred in 6 of the 118 patients (4%), of whom 2 were symptomatic (1.7%). Using the Theron system, 5 of 38 patients (13.2%) had emboli while 1 of 80 (1.25%) had emboli using the Percusurge system (P = 0.019). Symptoms occurred only with emboli >20 microm. Symptomatic retinal embolization is uncommon during carotid stenting, but is more likely when external to internal carotid collaterals are not protected. Cerebral protection system designs should take into consideration the existence of collaterals and the need to protect against smaller sized emboli, which may cause blindness in the retinal circulation. PMID- 12112884 TI - Usefulness of hydrophilic coating on arterial sheath introducer in transradial coronary intervention. AB - Radial artery spasm is one of the major problems during transradial coronary intervention (TRI). The sheath introducer with hydrophilic coating may reduce the incidence of spasm and reduce the difficulty in removing it from the radial artery under the situation of spasm artery spasm. After we compared the friction resistance between the sheath introducer with hydrophilic coating and that without coating (nine samples each) in vitro, the sheath introducers with and without hydrophilic coating were randomly used in 37 and 36 patients, respectively, who underwent elective TRI with a 6 Fr introducer sheath. Hydrophilic coating of sheath introducer reduced friction resistance by 70% (P < 0.00001) in in vitro model and facilitated sheath removal after finishing TRI (P = 0.0003). Hydrophilic coating of sheath introducer is useful in TRI. PMID- 12112885 TI - One core laboratory at two international sites, is that feasible? An inter-core laboratory and intra-observer variability study. AB - To assess the magnitude of differences in QCA outcomes between two cooperating core laboratories in a single trial, we have carried out an inter-core laboratory variability study. Two QCA experts at the Montreal Heart Institute and Heart Core Leiden both analyzed 32 lesions (pre- and post-intervention) in accordance with previously agreed upon standard operating procedures. One of the experts analyzed the whole image set twice to determine the intraobserver variability. The inter core laboratory differences in the acute gain (n = 31 pairs) are non-significant. The systematic errors of the individual measurements (n = 63 analyses) show an excellent intraclass correlation coefficient of reliability (>75%), except for the stent length (67.7%). The corresponding random errors are small. In general, the intra-observer systematic and random errors are both slightly smaller than those for the inter-core laboratory study. QCA analyses in clinical trials can be carried out in core laboratories at two different locations if and only if highly standardized conditions are maintained. PMID- 12112886 TI - Inflammatory markers and coronary interventions: a potentially useful follow-up modality after stenting. AB - Periprocedural levels of various inflammatory markers have been correlated with prognosis in patients undergoing percutaneous coronary interventions. However, long-term variations of interleukin-1 receptor antagonist (IL-1Ra) or C-reactive protein (CRP) during follow-up after coronary interventions were not previously investigated. The aim of our study was to perform serial evaluations of these markers before and after coronary stenting and to correlate them with clinical status. Plasma levels of IL-1Ra and CRP were measured at baseline and 3 and 6 months after the procedure in 31 patients with symptomatic coronary artery disease undergoing stent implantation, who had no evidence of myocardial ischemia at 6-month follow-up. While at 3 months there were no significant variations of baseline values, 6 months after the procedure a significant decrease from baseline was observed both in IL-1Ra and CRP levels (median -24 pg/ml, P = 0.048, and -0.13 mg/dl, P = 0.017, respectively). Six-month reduction in both IL-1Ra and CRP levels was significant in patients with unstable angina (n = 18; IL-1Ra: from 175 to 119 pg/ml, P = 0.001; CRP: from 0.52 to 0.18 mg/dl, P = 0.002) and nonsignificant in those with stable angina (n = 13) on admission (IL-1Ra: from 123 to 158 pg/ml, P = 0.22; CRP: from 0.19 to 0.10 mg/dl, P = 0.44). In conclusion, a significant reduction of IL-1Ra and CRP levels is observed 6 months after stent implantation in patients with preprocedural unstable angina who remain free of ischemia. This decrease suggests a stabilization of the inflammatory process and may be associated with a favorable prognosis after coronary interventions. PMID- 12112887 TI - Percutaneous interventions in patients with cocaine-associated myocardial infarction: a case series and review. AB - Cocaine-associated myocardial infarction (CAMI) is a well-reported entity. Most previous reports on CAMI have been limited to conservative care utilizing benzodiazepines, aspirin, nitroglycerin, calcium channel blockers, and thrombolytics. Current guidelines on CAMI advocate immediate use of angiography and angioplasty if available rather than routine administration of thrombolytics. However, based on literature search from 1966 to 2001 (using keywords "cocaine," "myocardial infarction," and "angioplasty"), there have been only two case reports of percutaneous coronary intervention (PCI) in patients with cocaine associated myocardial infarction. Both were notable for complications either during or immediately after the procedure. We report a series of 10 patients with cocaine-associated myocardial infarction who were treated with percutaneous interventions, which included angioplasty, stenting, and AngioJet mechanical extraction of thrombus. Despite the different arteriopathic process involved, our findings suggest that PCI can be performed safely and with a high degree of procedural success in patients with CAMI. PMID- 12112888 TI - Results of the Jostent coronary stent graft implantation in various clinical settings: procedural and follow-up results. AB - The Jostent coronary stent graft (CSG) is composed of a PTFE layer sandwiched between two stainless steel stents, initially introduced for the treatment of coronary perforations and aneurysms with excellent results. By providing a mechanical barrier, this stent design also may be beneficial in the treatment of complex ulcerated lesions and in-stent restenosis by preventing debris protrusion and neointimal proliferation through the stent struts. To evaluate the safety and efficacy of this stent graft, we implanted 78 CSGs in 70 patients for a broad range of indications, including coronary perforations, aneurysms, degenerated saphenous vein grafts, complex lesions, and in-stent restenosis. The primary angiographic success rate (95.9%) was high, and using intravascular ultrasound (IVUS) guidance during stent implantation and high inflation pressures (19.3 +/- 3.2 atm), stent expansion with optimal symmetry was achieved in 94.7%. One limitation of the Jostent CSG was the side-branch occlusion rate (18.6%) and the resulting non-Q-wave infarction rate in seven cases (mean CK elevation, 238 U/l), acute Q-wave MI in two cases, and transient ventricular fibrillation in one patient after occlusion of the proximal RCA side branch without further complications. Subacute stent thrombosis occurred in four cases (5.7%) 7 to 70 days after stent implantation, despite using combined antiplatelet therapy with aspirin (ASA), ticlopidine, and/or clopidogrel for 30 days. Angiographic follow up was available in 56 patients (80.0%) after a mean of 159 +/- 49 days, and follow-up IVUS was available in 38 cases. The overall restenosis rate (> 50% diameter stenosis) was 31.6% manifest primarily as edge restenosis (29.8% stent edge vs. 8.8% stent center; P < 0.001). IVUS examinations showed a minimal late lumen loss of 0.4 +/- 2.2 mm(2) within the center of the stent graft vs. 3.2 +/- 2.3 mm(2) at the stent edges (P < 0.001). The restenosis rate in the prespecified subgroups was 33.3% for saphenous vein grafts (2/6 lesions), 30.0% in complex lesions (6/20 lesions), and 38.5% (10/26 lesions) for the treatment of in-stent restenosis. Implantation of the Jostent CSG is feasible and safe, even in complex lesion subsets, and is associated with high primary success rates provided major side branches are avoided. The use of this stent may require an extended time course of antiplatelet therapy. Frequent focal stent edge renarrowing influences the overall restenosis rate. However, in treatment of complex in-stent restenosis and vein graft lesions, stent grafts may offer benefit over conventional therapies. Covered stents such as the JoMed coronary stent graft may become essential for bailout treatment of coronary perforations. PMID- 12112889 TI - Endoluminal stenting of the subclavian artery to treat a surgically created left main equivalent lesion. PMID- 12112890 TI - Laser-facilitated thrombectomy: a new therapeutic option for treatment of thrombus-laden coronary lesions. AB - To overcome the adverse complications of balloon angioplasty in thrombus burden lesions (i.e., distal embolization, platelet activation, no-reflow phenomenon with persistent myocardial hypoxemia), mechanical removal of the thrombus or distal embolization protection devices is required. Pulsed ultraviolet excimer laser light at 308 nm can vaporize thrombus and suppress platelet aggregation. Clinical experience has already shown its efficacy in acute ischemic-thrombotic acute coronary syndromes. Unlike other thrombectomy devices, a 308 nm excimer laser can ablate thrombi as well as the underlying plaque, speed up thrombus clearing, and enhance thrombolytic and GP IIb/IIIa activity. It can also be employed in patients with contraindications for systemic thrombolytic agents or GP IIb/IIIa antagonists. Our report covers clinical data and technical aspects concerning three patients with acute myocardial infarction who presented with a large thrombus burden. After successful laser-transmitted vaporization of the thrombus mass in these patients, the remaining thrombus burden was evacuated, and normal antegrade coronary flow was successfully restored. This approach can be useful for selective patients with acute coronary syndromes. PMID- 12112892 TI - Thou shalt not (look like you're trying to) steal. PMID- 12112891 TI - Functional evaluation of steal by unligated first intercostal branch before transcatheter embolization in recurrent angina after a LIMA-LAD graft. AB - Unligated side branches of the left internal mammary artery (LIMA) have been indicated as a cause of coronary steal resulting in postoperative angina. Although a number of studies have reported successful embolization of the side branches to relieve angina, this phenomenon is still controversial and it has been either emphasized or rejected in studies that attempted to obtain hemodynamic evidence of the steal using angiographic and intravascular Doppler techniques. In this case study, we tried to reproduce physiological decrease in the muscular and coronary beds as it could occur during activity. Our results, using an intracoronary Doppler wire with adenosine combined with forced ventilation, showed that a trial occlusion of the LIMA side branch with a balloon technique can demonstrate whether the flow through the LIMA would increase after embolization of the side branch. PMID- 12112893 TI - Coronary perforation as a potential complication derived from coronary thrombectomy with the X-Sizer device. AB - We report a case of coronary perforation after failed atherectomy with a 2 mm X Sizer catheter in recent totally occluded right coronary artery. The perforation was successfully managed with a polytetrafluoroethylene-covered stent with satisfactory final angiographic results. Possible predictors of this complication with this new device are discussed. PMID- 12112894 TI - Endovascular stent placement for venous obstruction after cardiac transplantation in children and young adults. AB - Survival following cardiac transplantation in children and adults, including the group with complex congenital heart disease, has improved over the last decade secondary to medical and surgical advances in management. There have been rare reports of superior vena cava obstruction at anastomotic sites following transplantation. In patients following heart transplantation, venous stenosis can limit the ability to perform endomyocardial biopsies. We reviewed our experience in three patients who underwent cardiac transplantation and developed significant venous stenosis requiring intervention. All three were successfully treated by transcatheter implantation of endovascular stents. Endovascular stent implantation for venous obstruction in patients following cardiac transplantation was safe and effective, allowing improved ease of catheterization for future posttransplantation monitoring and surveillance. PMID- 12112895 TI - Balloon dilation and stent implant through the side of a previously placed intravascular stent: a new option for the interventionalist. AB - Intravascular stent placement for relief of vascular stenoses often requires that a stent be positioned across the origin of side-branch vessels. Most currently available peripheral vascular stents have a closed-cell design that will not allow catheter manipulation, dilation, or stenting through the stent cells. Therefore, if a side branch arising off a previously stented vessel is stenotic or if its origin has been compromised, transcatheter dilation of the branch vessel has not been possible. We report a series of successful balloon dilation and stent placement through the sides (cells) of previously placed IntraStent DoubleStrut LD stents in the aortic arch and abdominal aorta of pigs. PMID- 12112896 TI - Initial and late results after catheter intervention for neonatal critical pulmonary valve stenosis and atresia with intact ventricular septum: a technique in continual evolution. AB - Critical pulmonary valve stenosis or atresia with intact ventricular septum is a rare congenital cardiac defect that can be technically difficult to alleviate in the catheterization laboratory. Over the past 10 years, several techniques and modifications with variable results have been advocated to facilitate the valvuloplasty procedure. This report describes a single operator's experience using various techniques in 28 neonates with critical pulmonary stenosis or atresia who were considered candidates for transcatheter intervention. The first two patients underwent a gradational balloon valvuloplasty approach that resulted in prolonged fluoroscopy exposure. Thereafter, a "snare assisted" umbilical artery approach was developed which facilitated the valvuloplasty procedure and resulted in significantly fewer balloons used and shorter fluoroscopy times. Early in our experience, stiff guidewire perforation of atretic pulmonary valves was used, whereas in our last two patients, a simplified perforation technique with a new 0.9-mm excimer laser catheter was used. Late echocardiographic and clinical follow-up evaluation in 27 patients demonstrates persistent gradient relief, resolution of tricuspid valve insufficiency, and elimination of right to left shunting at the atrial level. Balloon valvuloplasty is the treatment of choice for critical pulmonary valve stenosis or atresia with intact ventricular septum. When necessary, the use of umbilical artery "snare assistance" facilitates the valvuloplasty technique and shortens procedure time while laser perforation is currently preferable for perforation of the atretic pulmonary valve. PMID- 12112898 TI - Comparing stents performance: is there a need to reinvent the wheel? PMID- 12112897 TI - Open-cell design stents in congenital heart disease: a comparison of IntraStent vs. Palmaz stents. AB - Slotted stainless Palmaz stents (PS) remain the most commonly used in congenital heart disease (CHD). Limitations of PS include rigidity, foreshortening, poor conformability on expansion, balloon rupture, and jailing of side branches. Recently, stents with open-cell design (IntraStent; IS) in appropriate sizes for CHD have been introduced. We reviewed our experience with the IS, comparing performance with the PS in a retrospective nonrandomized, uncontrolled observational study. Between May 1999 and March 2001, 34 IS (10-36 mm) and 34 PS (10-40 mm) were implanted in 57 patients (3 months to 25 years old; median, 3.24 years) in congenital or postoperative lesions. There was no statistically significant difference between the two groups in efficacy, % diameter increase (162% +/- 203% for IS vs. 153% +/- 177% for PS), and % gradient reduction (69% +/ 23% for IS vs. 80% +/- 27% for PS). Other aspects of stent performance differed significantly (P < 0.05): PS forshortened more (mean 18% for PS vs. 0% for IS) and were associated with balloon rupture (9/34 for PS vs. 0/34 for IS; P = 0.002), but had no evidence of intimal protrusion in between struts (0/34 for PS vs. 14/34 for IS; P < 0.001) and no recoil > 15% (0/34 for PS vs. 7/34 for IS; P = 0.006); IS conformed almost twice as well to vessel curvature (P = 0.003). Although these factors did not affect immediate procedural success, balloon rupture in two PS resulted in stent malpositions. Among IS, the origins of three covered side branches were accessed and balloon dilated up to 6 mm through the side of the stent. In conclusion, stents with open-cell design have some characteristics that seem beneficial for their use in CHD: they foreshorten less, are conformable, are less likely to cause balloon rupture, and can allow access to jailed branches. However, they have a higher incidence of significant percent recoil and protrusion of intimal tissue in between struts, which may impact long term stent performance. PMID- 12112899 TI - Radiofrequency-assisted atrial septoplasty for an intact atrial septum in complex congenital heart disease. AB - Septoplasty of the atrial septum was performed with sequential balloon dilation following radiofrequency-assisted perforation of an intact atrial septum in two newborn infants with hypoplastic left heart syndrome and one with double-outlet right ventricle. PMID- 12112900 TI - Late aortic dislocation of a stent following stent angioplasty for ostial renal artery stenosis. AB - A patient with left RAS was treated by stent angioplasty followed by a multivessel percutaneous coronary intervention. Six months later, an aortic dislocation of the stent was diagnosed. The fully expanded stent was caught with a balloon catheter and fixed in the left external iliac artery. Stent migration after initially successful stent angioplasty for RAS is possible. Fully expanded, dislocated balloon-expandable stents can be secured by implanting them into the iliac artery. PMID- 12112902 TI - Pulmonary venous pressure: relationship to pulmonary artery, pulmonary wedge, and left atrial pressure in normal, lightly sedated dogs. AB - Because pulmonary venous pressure has never been measured, it is unclear whether pulmonary wedge pressure measures left atrial pressure, as commonly assumed, or pressure more upstream in the pulmonary venous or capillary beds. Fluid-filled mean pulmonary artery and pulmonary wedge pressure were compared with pulmonary venous and left atrial pressure obtained with high-fidelity micromanometer catheters in eight lightly sedated dogs over a physiologic range of filling pressures. In all conditions, mean pulmonary wedge pressure was virtually identical (r = 0.99) to mean left atrial pressure (slope = 0.99; intercept = 0.46 mm Hg). At the same time, mean pulmonary venous pressure (17.1 +/- 6.5 mm Hg) was intermediate between mean pulmonary artery pressure (20.2 +/- 6.2 mm Hg) and mean pulmonary wedge pressure (13.3 +/- 6.2 mm Hg; P < 0.0001) or mean left atrial pressure (13.4 +/- 6.3 mm Hg; P < 0.0001). These relationships were maintained over normal and increased pressure ranges. As measured by conventional flow-directed pulmonary catheters, mean pulmonary wedge pressure accurately reflects left atrial pressure in lightly sedated, spontaneously breathing normal dogs. PMID- 12112901 TI - Potential emerging therapeutic strategies to prevent restenosis in the peripheral vasculature. AB - Despite the availability of antiplatelet and antithrombotic therapies, recent advances in catheter and stent technology and improved operator skill, restenosis remains the most frequent problem associated with percutaneous and surgical revascularization interventions for both coronary and peripheral arterial disease. Prevention of restenosis in the coronary vasculature has been demonstrated with cilostazol, trapidil, probucol, tranilast, nitric oxide donors, and clopidogrel. Given the similarities in revascularization procedures and in the pathophysiology of restenosis, it is possible that these results may be extrapolated to the setting of restenosis in the peripheral vasculature, making trials with these agents imperative. Several new agents have shown promising preliminary results for the prevention of restenosis in the peripheral vasculature, including cilostazol, low-molecular-weight heparins, and elastase. Several nonpharmacologic treatment modalities are also under study to prevent peripheral and coronary restenosis and have shown favorable initial results. These include endovascular radiation brachytherapy, arterial gene therapy, photoangioplasty, and several novel treatment delivery systems. PMID- 12112903 TI - Anterograde transumbilical venous balloon aortic valvuloplasty. PMID- 12112904 TI - Characterizing PCI outcomes for our patients & ourselves: best case, worst case, real case. PMID- 12112905 TI - Percutaneous therapy of dialysis access failure. PMID- 12112906 TI - Screening for subclavian artery stenosis in patients who are candidates for coronary bypass surgery. AB - We prospectively evaluated 59 patients who were deemed candidates for coronary bypass surgery after coronary artery angiography for subclavian artery narrowing, which could compromise the ipsilateral internal thoracic artery graft. Bilateral arm blood pressure (BP) measurements, auscultation for supraclavicular or cervical bruits, and questioning about cerebrovascular ischemic symptoms were compared to brachiocephalic-subclavian arteriography. One neurologic complication occurred during arteriography. An upper extremity BP difference of > or = 15 mm Hg identified all patients with > or = 50% subclavian artery narrowing. We recommend brachiocephalic-subclavian arteriography only in patients with abnormal noninvasive screening for subclavian stenosis, not routinely. PMID- 12112907 TI - Mechanism of cutting balloon angioplasty for in-stent restenosis: an intravascular ultrasound study. AB - We investigated by intravascular ultrasound (IVUS) the mechanism of action of cutting balloon (CB) angioplasty in patients with in-stent restenosis. Seventy one consecutive restenotic lesions of 66 patients were studied by quantitative coronary angiography (QCA) and IVUS before, immediately after, and, in 20 cases, at 24-hr time interval after CB. CB was selected according to 1:1 CB-to-stent ratio and inflated at 8 atm for 60-90 sec. Both IVUS planar and volumetric (Simpson's rule, 25 patients) analysis were carried out. IVUS measurements included external elastic membrane area (EEMA), stent area (SA), minimal lumen area (MLA), and restenosis area (RA). Following CB, QCA analysis showed increase of minimal lumen diameter (1.17 +/- 0.46 vs. 2.45 +/- 0.51 mm; P < 0.0001) and decrease of diameter stenosis (64% +/- 13% vs. 21% +/- 9%; P < 0.0001). IVUS measurements showed a significant increase of MLA (2.18 +/- 0.80 vs. 7.31 +/- 1.8 mm(2); P < 0.0001), SA (9.62 +/- 2.6 vs. 10.7 +/- 2.75 mm(2); P < 0.0001), and EEMA (17.27 +/- 5 vs. 18.1 +/- 5 mm(2); P < 0.0001) and a decrease of RA (7.43 +/ 2.63 vs. 3.45 +/- 1.39 mm(2); P < 0.0001). No significant change was observed in the original plaque + media area (7.65 +/- 3 vs. 7.38 +/- 2.9 mm(2); P = NS). Thus, of the total lumen enlargement (5.13 +/- 1.85 mm(2)), 23% was the result of increase in mean SA, whereas 77% was the result of a decrease in mean RA. These changes were associated with a 5% increase in EEMA. IVUS volumetric changes paralleled planar variations. Angiographic and IVUS changes were well maintained at 24 hr. CB enlarges coronary lumen mainly by in-stent tissue reduction associated with a moderate degree of additional stent expansion. Favorable QCA and IVUS acute results are maintained at 24 hr. PMID- 12112908 TI - Cardiac catheterization in morbidly obese patients. AB - The safety and findings of cardiac catheterization and coronary angiography in morbidly obese patients with suspected coronary heart disease (CHD) have not been fully examined in the modern era. From a database of 4,978 patients undergoing diagnostic cardiac catheterization, we identified 110 with morbid obesity (body mass > or = 145 kg and body mass index > or = 40 kg/m(2)). Relative to all the other patients in this database, morbidly obese patients had a lower prevalence of CHD (45% vs. 72%; P < 0.05), reflecting a higher prevalence of false positive noninvasive tests. Overall, noninvasive tests were only 75% sensitive and 39% specific for CHD in this group. Use of radial access (66%) and femoral closure devices (24%) was much more frequent in the morbidly obese cohort. Complications were no more frequent in the morbidly obese group, with major (0 vs. 0.9%) and minor (4.7% vs. 3.5%) adverse outcomes being similar to the rest of the database. We conclude that cardiac catheterization using the radial artery or a femoral closure device is a safe and effective method of evaluating CHD in morbidly obese patients. In contrast, noninvasive testing is frequently not definitive and may be misleading. PMID- 12112910 TI - Abciximab administration and clinical outcomes after percutaneous intervention for in-stent restenosis. AB - Abciximab therapy improves clinical outcomes after percutaneous interventions for de novo coronary artery disease. We sought to determine whether clinical outcomes after percutaneous intervention for in-stent restenosis are affected by abciximab administration. Between January 1996 and July 1999, 322 consecutive patients underwent percutaneous intervention for in-stent restenosis; 157 patients received abciximab and 165 patients were treated without abciximab based on operator discretion. Baseline clinical and angiographic variables and type of percutaneous intervention were recorded. Follow-up information was obtained and clinical endpoints were recorded. A multivariate analysis was performed to determine the independent variables associated with adverse clinical outcomes. Baseline clinical and angiographic variables were similar in both groups. Patients who received abciximab were more likely to be treated with rotational atherectomy and less likely to have only balloon angioplasty or repeat stenting. Mean follow-up duration was 19 +/- 12 months. There were no significant differences in the incidence of angina/myocardial infarction (29% vs. 30%; P = 0.9), target vessel revascularization (18% vs. 21%; P = 0.5), death (8% vs. 7%; P = 0.4), or major adverse cardiovascular events (38% vs. 39%; P = 0.9) in both groups. Abciximab administration was not an independent variable associated with adverse outcomes. In this observational study, clinical outcomes after percutaneous intervention for in-stent restenosis did not seem to be affected by abciximab administration. Randomized trials are needed to identify the role of platelet glycoprotein IIb/IIIa inhibitors in the management of in-stent restenosis. PMID- 12112909 TI - Impact of various intravascular ultrasound criteria for stent optimization on the six-month angiographic restenosis. AB - We evaluated the impact of different intravascular ultrasound (IVUS) criteria on 6-month angiographic restenosis in 511 patients with 560 lesions. Seven IVUS criteria were evaluated in this study; stent area at lesion segment 1) > or = 100% of distal reference lumen area, 2) > or = 90% of distal reference lumen area, 3) > or = 80% of average reference lumen area, 4) > or = 90% of average reference lumen area, 5) > or = 55% of average reference vessel area, 6) >/= 7 mm(2), and 7) > or = 2). Using the relative measurement (criteria 1-5), the angiographic restenosis rate was not statistically different. However, absolute measurement of stent area > or = 9 mm(2) (criteria 6 and 7) were associated with significantly lower restenosis rate (14.8% vs. 30.9%, P = 0.001, and 13.5% vs. 24.6%, P = 0.006, respectively). In conclusions, using the relative measurement of IVUS criteria, the occurrence of angiographic restenosis might not be predicted. The absolute measurement of IVUS stent area was the predictor of angiographic restenosis. PMID- 12112911 TI - Feasibility and safety of concomitant left internal mammary arteriography at the setting of the right transradial coronary angiography. AB - We investigated the feasibility and safety of concomitant left internal mammary artery (LIMA) angiography at the setting of the right transradial coronary angiography (TRCA). LIMA angiography was performed using a 5 Fr Simmons-type catheter with a newly modified tip in 184 consecutive patients. The catheter was reformed in the descending (new method) or ascending (traditional method) aorta and manipulated to cannulate the left subclavian artery and LIMA orifice. LIMA angiography was performed selectively in 164 patients (89%) and semiselectively (when the catheter tip reached and was directed to the mammary artery orifice) in 20 patients. There were no procedure-related complications. The image quality of all the semiselective angiograms was also determined satisfactory. Total procedural time was 223 +/- 168 sec. The catheter was reformed using the new method in 160 patients (87%). The catheter reformation time and total procedure time were significantly shorter with the new method than with the traditional method (18 +/- 8 vs. 117 +/- 115 sec, p = 0.000; 204 +/- 191 vs. 309 +/- 139 sec, p = 0.021, respectively). In conclusion, LIMA angiography can be performed readily and safely at the setting of the right TRCA using a Simmons-type catheter. The image quality of the LIMA angiograms is sufficient to obviate the need of the second preoperative angiography via another route. PMID- 12112913 TI - Comparison of outcomes after 8 vs. 9.5 French size intra-aortic balloon counterpulsation catheters based on 9,332 patients in the prospective Benchmark registry. AB - The Benchmark intra-aortic balloon counterpulsation (IABC) registry maintains prospectively gathered clinical information on a large cohort of IABC patients. The purpose of the present report is to compare in-hospital outcomes and complications in patients treated with the newer 8 vs. 9.5 Fr size catheters. Between January 1997 and August 2000, data on 7,078 9.5 Fr and 2,254 8 Fr IABC insertions were submitted to Benchmark. This was not a randomized comparison but rather a posthoc analysis of prospectively gathered data. There was less limb ischemia with the 8 Fr IABC size catheter. There were no significant differences in bleeding or mortality between the two groups. Smaller IABC catheter size is associated with significantly less limb ischemia, especially in higher-risk patients. The large, population-based, ongoing Benchmark registry provides a useful vehicle for outcomes research concerning the evolving practice of IABC. PMID- 12112912 TI - Immediate protamine administration and sheath removal following percutaneous coronary intervention: a prospective study of 429 patients. AB - We tested the approach of reversing anticoagulation following PCI and immediate sheath removal in 429 consecutive patients. On completion of the PCI, protamine was administered, and the vascular sheath was immediately removed. Stents were used in 364 patients (85%) and GP IIb/IIIa inhibitors were used in 52 patients (12%). Time to achieve hemostasis was 30 +/- 17 min. Minor groin bleeding occurred in six patients. One patient required repair of femoral pseudoaneurysm. Mean creatine kinase at 8 and 16 hr post-PCI were 129 +/- 35 and 145 +/- 32 units, respectively. Creatine kinase rose to > 3 times normal in 12 out of 350 patients (3.4%). Prior to 48 hr, eight patients (1.9%) required emergency PCI or coronary bypass surgery. Follow-up at 30 days observed no deaths and only three target vessel revascularizations (0.7%). In conclusion, immediate reversal of anticoagulation and sheath removal after PCI is safe and feasible. PMID- 12112915 TI - Stent deployment within a guide catheter aids removal of a fractured buddy wire. AB - We report a case of fractured buddy wire that was successfully removed by deploying a stent within the guide catheter, trapping the fractured segment of wire between stent and endoluminal surface of the guide catheter. This technique provides an alternative to either percutaneous snare or surgical intervention. PMID- 12112914 TI - Overcoming vascular anatomic challenges to cardiac catheterization by the radial artery approach: specific techniques to improve success. AB - Anatomical variations in the peripheral vasculature can result in decreased procedural success rates for cardiac catheterization performed through the radial artery approach. We describe four categories of vascular challenges encountered in our catheterization laboratory: severe spasm, severe tortuosity, vascular stenosis, and congenital anatomical variations (e.g., accessory radial artery, radioulnar loop). For each situation, we provide a case report illustrating techniques that allowed for successful completion of the case. PMID- 12112916 TI - Coronary stent strut avulsion in aorto-ostial in-stent restenosis: potential complication after cutting balloon angioplasty. AB - We report a case of stent strut avulsion by the cutting balloon during the withdrawal of the deflated balloon catheter in aorto-ostial in-stent restenosis, which was managed successfully by another stent. The proposed mechanisms and recommendations to avoid this rare complication are provided. PMID- 12112917 TI - Aorto-ostial disease and aorto-ostial in-stent restenosis: poorly recognized but very complex lesion subsets. PMID- 12112918 TI - Directional atherectomy of a calcified lesion using a new atherectomy device. AB - We report on a case of directional atherectomy performed on a calcified coronary lesion using a novel device with a hardened titanium cutter. The successful removal of calcified plaque was documented by intravascular ultrasound assessment and confirmed by histopathological analysis of the obtained plaque specimen. PMID- 12112919 TI - Successful management of a resistant renal artery stenosis in a child using a 4 mm cutting balloon catheter. AB - Percutaneous transluminal renal angioplasty (PTRA) is a well-established method to treat renal artery stenosis (RAS) in children and adults. However, a significant number of stenoses might not be treated by interventional techniques due to the inability to dilate the RAS. Conventional balloon angioplasty with a high-pressure coronary angioplasty balloon at 20 atm was unable to dilate a significant RAS in a 12-year-old child with severe renovascular hypertension (RR 195/125 mm Hg). After using a 4 mm cutting balloon, we achieved wide patency of the renal artery and an instant normalization of blood pressure without further need of antihypertensive therapy. PTRA using the cutting balloon technique may offer an additional therapeutic option for selected patients in whom conventional balloon angioplasty was not able to dilate RAS. PMID- 12112920 TI - The Barath cutting balloon: battling the sword of Damocles. PMID- 12112921 TI - A novel use of the Amplatzer muscular ventricular septal defect occluder. AB - We report a case of a 12-month-old-infant with double outlet right ventricle and pulmonary stenosis who presented with signs of superior vena cava syndrome secondary to a dysfunctioning bidirectional Glenn shunt. The patient was successfully treated with transcatheter obstruction of an accessory pulmonary blood flow using the Amplatzer muscular ventricular septal defect occluder. PMID- 12112922 TI - Direct visualization of left ventricular free wall rupture by levocardiography. AB - Two cases of left ventricular free wall rupture and one case of combined left ventricular free wall and ventricular septal rupture are described where ventriculography played a key role in diagnosis. In all three cases of patients with acute myocardial infarction, identification and localization of the defect was made by angiography. This report illustrates the safety and feasibility of ventriculography in patients with suspected cardiac rupture. PMID- 12112923 TI - A case of transradial carotid stenting in a patient with total occlusion of distal abdominal aorta. AB - We report a case with severe carotid stenosis in which carotid stenting was performed via the radial artery due to total occlusion of distal abdominal aorta. The radial approach offers a potential alternative in cases in which the femoral approach is problematic. PMID- 12112924 TI - Potential goals for the dimensions of the pulmonary arteries and aorta with stenting after the Fontan operation. AB - The purpose of this study was to clarify desired stent sizes for stenotic lesions in the post-Fontan circulation. Using angiograms from 22 patients before and at late follow-up (> or = 15 years) after the Fontan operation, we measured the maximum diameters of the proximal pulmonary arteries (PA) and the descending aorta. The diameters of the PA ipsilateral to the inferior vena cava, contralateral to the inferior vena cava, and descending aorta after the Fontan were 10.6-22.6 (15.8 +/- 3.3), 8.0-19.1 (12.9 +/- 3.1), and 12.1-18.9 (15.8 +/- 2.0) mm, respectively, while the percent of normal predicted diameters (% N) were 55%-104% (70% +/- 14%), 38%-99% (66% +/- 17%), and 46%-74% (60% +/- 7%), respectively. Despite somatic growth, the % N of all vessel diameters decreased significantly after the Fontan operation. In conclusion, smaller-sized stents should be acceptable for both the pulmonary artery and descending aorta in the Fontan circulation. PMID- 12112925 TI - IntraStent double-strut LD: collapse and recoil following use in postoperative stenoses. AB - We describe the use of the Intratherapeutics IntraStent in two postoperative patients, its collapse and recoil following deployment, and subsequent transcatheter correction. PMID- 12112926 TI - Management of a large pseudoaneurysm secondary to balloon dilation for native coarctation of the aorta with coil occlusion after stent implantation in a child. AB - We report a case of a 9-year-old girl status post-balloon dilation for native coarctation of the aorta who had a large aortic pseudoaneurysm 1 year after the initial procedure. The aneurysm was occluded with multiple coils after stent implantation to the aorta. The technique and possible advantages of this novel approach are discussed. PMID- 12112928 TI - Interventional catheterization management of perioperative peripheral pulmonary stenosis: balloon angioplasty or endovascular stenting. AB - There is limited reported experience of catheterization therapy for peripheral pulmonary stenosis (PPS) at a surgical site in the early postoperative period. We reviewed the clinical course of patients undergoing interventional catheterization for PPS at a surgical site < 7 weeks after surgery. Successful dilation (SD) was defined as > 50% increase in predilation diameter. From 1984 to 2000, 17 patients had 19 proximal pulmonary arteries dilated 1 to 46 (median 8) days postoperatively. Median age and weight were 3.1 year and 12.7 kg. Three arteries were initially occluded. Seventeen arteries had initial BD with postintervention imaging available in 15; 8 arteries had SD. The arterial diameter increased from 3.9 +/- 2.6 to 5.5 +/- 2.8 mm (P < 0.001). Nine of these arteries had stents placed with diameter increasing to 8.7 +/- 3.7 mm (P < 0.001 compared with post-BD diameter). Stents increased the diameter in all arteries and made four of four failed BD successful. In the two most recent procedures, stents were placed without prior BD with diameter increasing from 1.3 to 9 mm and 8.2 to 14 mm. A stent was placed in 1 of 7 arteries prior to 1993 and in 10 of 12 arteries thereafter (P < 0.004). Three patients prior to 1995 had catheterization related deaths secondary to vessel rupture after BD. BD produces SD in approximately one-half of the procedures but is associated with mortality. Stent placement increases vessel diameter substantially more than BD alone. Stents reduce the acute complication rate and avoid early reoperation in this patient group. PMID- 12112927 TI - Creation of an intra-atrial communication with a new Amplatzer shunt prosthesis: preliminary results in a swine model. AB - A nitinol shunt device was applied in six minipigs to create a precise intra atrial shunt. This self-expanding shunt device consists of two retention disks of 2-8 mm, a 4 mm connecting waist with a 10 mm eccentric hole. It requires a 7 Fr introducer sheath. The device is attached to the delivery cable with a microscrew. Placement technique is identical to that of Amplatzer septal occluder. Balloon dilation was performed immediately and 1 month after placement. One animal died from general anesthesia before device placement. Left atrial angiography showed a patent intra-atrial shunt in 5/5 pigs immediately and 4/4 in 1- to 3-month follow up. Postmortem examination demonstrated patent shunts partially or completely neoendothelialized. The shunt device was found to be an effective and safe way to create a permanent atrial communication. PMID- 12112929 TI - Stent supported angioplasty for coronary arterial stenosis following the arterial switch operation. AB - We present a case of a patient who underwent stent-supported angioplasty for left main bifurcation disease following the arterial switch operation. Prior to this procedure, the patient underwent surgical repair of the left coronary ostium and subsequently coronary artery bypass surgery with an arterial graft to the left anterior descending artery. We could not find a similar report in the literature. PMID- 12112931 TI - Guidelines: minimum standards, gold standards, or ideals for practice? PMID- 12112930 TI - Intracoronary brachytherapy. AB - Patients presenting with in-stent restenosis have an increased risk of need for repeat intervention. Intracoronary brachytherapy is indicated for these patients to prevent recurrent in-stent restenosis. Three intravascular brachytherapy systems are currently FDA-approved for use in patients: one utilizing gamma radiation (Cordis) and two using beta-radiation (Novoste and Guidant). Current evidence and labeling do not support using intracoronary brachytherapy for prevention of restenosis in de novo lesions. Brachytherapy is absolutely contraindicated in patients unable to take prolonged combination antiplatelet drugs. Aspirin and a thienopyridine should be taken for 6 months if no new stent is placed and 12 months if a new stent is placed. If possible, new stent implementation should be avoided. PMID- 12112934 TI - Was Willis right? Thoughts on the interaction of depression and diabetes. AB - Recent studies suggest that depression may be an independent risk factor for developing type 2 diabetes. At a time when this disease is an epidemic, identifying interventions that could prevent or delay the onset of diabetes is a high priority, and treating depression in people predisposed to diabetes could be an effective approach. Future studies should establish a causal relationship between depression and the occurrence of type 2 diabetes, as well as possible mechanisms to explain the relationship, should one be observed. PMID- 12112935 TI - Oxidative stress and diabetic neuropathy: pathophysiological mechanisms and treatment perspectives. AB - Increased oxidative stress is a mechanism that probably plays a major role in the development of diabetic complications, including peripheral neuropathy. This review summarises recent data from in vitro and in vivo studies that have been performed both to understand this aspect of the pathophysiology of diabetic neuropathy and to develop therapeutic modalities for its prevention or treatment. Extensive animal studies have demonstrated that oxidative stress may be a final common pathway in the development of diabetic neuropathy, and that antioxidants can prevent or reverse hyperglycaemia-induced nerve dysfunction. Most probably, the effects of antioxidants are mediated by correction of nutritive blood flow, although direct effects on endoneurial oxidative state are not excluded. In a limited number of clinical studies, antioxidant drugs including alpha-lipoic acid and vitamin E were found to reduce neuropathic symptoms or to correct nerve conduction velocity. These data are promising, and additional larger studies with alpha-lipoic acid are currently being performed. PMID- 12112936 TI - Brain-derived neurotrophic factor (BDNF) regulates glucose and energy metabolism in diabetic mice. AB - Neurotrophins are important regulators in the embryogenesis, development and functioning of nervous systems. In addition to the efficacy of brain-derived neurotrophic factor (BDNF) in neurological disorders, we have found that BDNF demonstrates endocrinological functions and reduces food intake and blood glucose concentration in rodent obese diabetic models, such as C57BL/KsJ-db/db mice. The hypoglycemic effect of BDNF was found to be stronger in younger db/db mice with hyperinsulinemia than in older mice. While BDNF itself did not alter blood glucose in normal mice and streptozotocin (STZ)-treated mice, BDNF enhanced the hypoglycemic effect of insulin in STZ-treated mice. These data indicate that BDNF needs endogenous or exogenous insulin to show hypoglycemic action. In addition, BDNF treatment enhanced energy expenditure in db/db mice. The efficacy of BDNF in regulating glucose and energy metabolism was reproduced through intracerebroventricular administration, suggesting that BDNF acted directly on the hypothalamus, the autonomic center of the brain. PMID- 12112937 TI - Insulin resistance in type 1 diabetes. AB - Insulin resistance plays a larger role in the type 1 diabetes disease process than is commonly recognized. The onset of type 1 diabetes is often heralded by an antecedent illness and/or the onset of puberty, both conditions associated with insulin resistance. In the face of a damaged beta-cell and thus reduced insulin secretion, this change is enough to manifest hyperglycemia. During the first year of clinical disease, considerable evidence suggests that the occurrence of clinical remission or 'honeymoon period' is due to a temporary resolution of the insulin-resistant state present at diagnosis. Intensive diabetes management is associated with both improved insulin sensitivity and beta-cell function. This indicates that the historical data on the changes in insulin secretion post diagnosis may be inappropriate when designing current studies. The known physiological relationship between beta-cell function and insulin sensitivity complicates interpretation of insulin secretion data obtained as part of prevention or intervention trials. While it is recommended that at least a subset of subjects participating in these trials undergo formal measurements of insulin sensitivity to evaluate effects of therapy on this parameter independent of effects on the beta-cell, the sample size must be sufficient to determine an effect if present. Finally, one could speculate that it is possible that subsets of people with mild manifestations of the type 1 autoimmune disease process could benefit from treatments aimed at improving the insulin-resistant state. PMID- 12112938 TI - Diagnosing coronary artery disease in diabetic patients. AB - Although several diagnostic modalities are available to the clinician interested in diagnosing coronary artery disease, very few have been validated in diabetic populations. This review discusses the non-invasive diagnosis of coronary disease in diabetic patients. Evidence regarding the prevalence and prognostic significance of silent ischemia is reviewed and the potential impact of silent ischemia on the diagnostic characteristics of the exercise treadmill test discussed. Other diagnostic tools are considered, and recommendations are made with respect to screening asymptomatic diabetic patients for coronary artery disease. PMID- 12112939 TI - Elevated fasting insulin concentrations associate with impaired insulin signaling in skeletal muscle of healthy subjects independent of obesity. AB - BACKGROUND: Insulin signaling is impaired in the skeletal muscle of obese subjects but whether defects in skeletal muscle insulin signaling also characterize insulin resistance of non-obese individuals is unknown. The detection of insulin signaling defects in muscle biopsies is hampered by the variation of the contaminating non-muscle elements such as blood, connective tissue, fat, and blood vessel structures. Freeze-drying and macroscopic purification of the muscle fibers prior to the analysis might offer a possibility to minimize the analytical variation due to these contaminants. METHODS: In the present study we first determined whether insulin signaling could be reliably assessed in freeze-dried muscle specimens, which are free of non-muscle contaminants, and then applied this method to the study of insulin signaling in weight-matched insulin-sensitive and insulin-resistant non-diabetic men. RESULTS: In rat muscle, increases in tyrosine phosphorylation of insulin receptor (IR) and activity of the insulin receptor substrate-1 (IRS-1)-associated phosphatidylinositol (PI) 3-kinase activity by insulin were similar or higher in freeze-dried and purified muscle than wet muscle. Prior to freeze-drying and purification, biopsies of human vastus lateralis muscle contained between 1% and 40% non-muscle contaminants (11+/-3%, mean+/-SEM, n=19). In freeze-dried biopsies of human vastus lateralis muscle taken before and after 30 min of hyperinsulinemia (serum free insulin 61+/-1 mU/l) in 13 non-diabetic men, insulin increased IR tyrosine phosphorylation 1.4-fold (p<0.05) and IRS-1-associated PI 3 kinase activity 1.7-fold (p<0.005). Insulin-stimulated PI 3-kinase activity was significantly inversely correlated with the fasting serum insulin concentration (r=-0.57, p<0.05). When divided according to the median fasting serum insulin concentration, the men with high fasting insulin [HI, n=7, age 44+/-3 years, body mass index (BMI) 25+/-1 kg/m(2)] as compared to the men with low fasting insulin [LI, n=6, age 45+/-3 years (NS), BMI 24+/-1 kg/m(2) (NS)] had lower rates of whole-body glucose uptake (3.4+/-0.4 vs 5.5+/-0.3 mg/kg min, p<0.005), higher fasting plasma glucose concentrations (5.9+/-0.2 vs 5.2+/-0.1 mmol/l, p<0.05), higher fasting serum triglycerides (1.4+/-0.2 vs 0.9+/-0.1 mmol/l, p<0.05) and lower high-density lipoprotein (HDL) cholesterol concentrations (1.3+/-0.1 vs 1.7+/-0.1 mmol/l, p<0.05). Insulin-stimulated IR tyrosine phosphorylation (p<0.05) and IRS-1-associated PI 3-kinase activity (p<0.05) were significantly lower in the HI than the LI group. CONCLUSIONS: Taken together these data demonstrate that early insulin signaling events can be reliably assessed in freeze-dried human skeletal muscle, and that in vivo insulin resistance and its accompanying features are associated with defects in early insulin signaling events in human skeletal muscle independent of body weight. PMID- 12112940 TI - Regulation of muscle malonyl-CoA levels in the nutritionally insulin-resistant desert gerbil, Psammomys obesus. AB - BACKGROUND: Malonyl-CoA, an allosteric inhibitor of carnitine palmitoyl transferase, controls the oxidation of fatty acids in muscle and other tissues by regulating their entrance into mitochondria. The level of malonyl-CoA in muscle is influenced by the uptake of energy substrates such as glucose, the precursor of its synthesis. The desert gerbil, Psammomys obesus, develops a severe insulin resistance with hyperinsulinemia and hyperglycemia when transferred from its native nutrition to a relative high-energy (HE) rodent chow. In keeping with this it shows a low rate of glucose transport and a failure of insulin to suppress hepatic glucose production during a hyperinsulinemic-euglycemic clamp. METHODS: The concentration of malonyl-CoA has been determined by radio-enzymatic assay together with the levels of citrate and malate in the gastrocnemius muscle of diabetes-prone (DP) and diabetes-resistant (DR) P. obesus during the administration of exogenous insulin, during an hyperinsulinemic-euglycemic clamp and following a 48-h fast. RESULTS: Muscle GLUT4 protein was reduced by 44% in DP Psammomys on a HE diet, compared with normoglycemic-normoinsulinemic animals on a low-energy (LE) diet. Muscle levels of malonyl-CoA, its precursor citrate and the citrate counter-ion malate were not elevated in DP Psammomys on the HE diet despite the hyperinsulinemia. Likewise, the administration of external insulin in subcutaneous (sc) implants to DP Psammomys did not evoke hypoglycemia, decrease glucose production or increase the concentration of malonyl-CoA and citrate in muscle, as it did in both albino rats and a selected line of DR Psammomys. In contrast, fasting significantly reduced muscle malonyl-CoA and citrate levels in the DP and DR Psammomys and promoted the fat oxidative pathway. CONCLUSION: Since non-diabetic Psammomys on a LE diet already show insulin resistance in the fed state, the sustained low malonyl-CoA levels in these animals imply a readiness for the oxidation of fatty acids. In a desert gerbil, adjusted to a food-scarce environment, such a continuing utilization of fatty acids as energy substrate by muscle would preserve the available glucose for glucose-dependent tissues and would diminish the need for gluconeogenesis. PMID- 12112941 TI - Prevention of diabetic nephropathy in mice by a diet low in glycoxidation products. AB - BACKGROUND: Reactive advanced glycation end products (AGEs), known to promote diabetic tissue damage, occur endogenously as well as in heated foods and are orally absorbed. The relative contribution of diet-derived AGEs to diabetic nephropathy (DN) remains unclear. METHODS: We tested a standard mouse food (AIN 93G) found to be rich in AGEs (H-AGE diet) in parallel with a similar diet that contained six-fold lower AGE content (L-AGE), but equal calories, macronutrients, and micronutrients. Non-obese diabetic mice (NOD) with type 1 diabetes (T1D) and db/db mice with type 2 diabetes (T2D) were randomly assigned to each formula for either 4 or 11 months, during which time renal parameters and AGE levels were assessed. RESULTS: Compared to the progressive DN and short survival seen in NOD mice exposed to long-term H-AGE feeding, L-AGE-fed NOD mice developed minimal glomerular pathology and a modest increase in urinary albumin:creatinine ratio (p<0.005), and a significantly extended survival (p<0.0001), consistent with lower serum (p<0.025) and kidney AGEs (p<0.01). Also, in the 4-month study, and in contrast to the H-AGE-fed mice, L-AGE-fed NOD and db/db mice exhibited low levels of renal cortex TGF beta-1 (p<0.05), laminin B1 mRNA (p<0.01) and alpha 1 IV collagen mRNA (p<0.05) and protein, in concert with reduced serum and kidney AGEs (p<0.05, respectively). CONCLUSION: Intake of high-level, food-derived AGEs is a major contributor to DN in T1D and T2D mice. Avoidance of dietary AGEs provides sustained protection against DN in mice; providing the rationale for similar studies in human diabetic patients. PMID- 12112942 TI - The insulin-mediated vascular and blood pressure responses are suppressed in CGRP deficient normal and diabetic rats. AB - BACKGROUND: Calcitonin gene-related peptide (CGRP) is extensively localized in the perivascular or periadventitia nerves throughout the body. CGRP is a potent vasodilator and its release is associated with dilation of these blood vessels. The present study investigated the contribution of the CGRP-mediated vasodilation to the insulin-induced vasodilatory response. METHODS: Male Wistar rats were treated with capsaicin (50 mg/kg) at 1-3 days of age to ablate the CGRP containing neurons. After 8 weeks some animals were made diabetic using streptozotocin. Vehicle-treated animals were used as controls. At 12-13 weeks the animals were fasted, anesthetized with chloralose/urethane and instrumented for recording of cardiovascular dynamics. RESULTS: Body weights and basal, insulin, glucose, mean arterial pressure (MAP), heart rate (HR), and vascular flows were not different in CGRP-deficient rats versus controls. Insulin infusion significantly decreased the MAP in vehicle-treated controls but this response was completely attenuated in CGRP-deficient rats. The decreased response to insulin was associated with a diminished vascular dilatory response in the iliac, renal, and superior mesenteric vessel beds. When insulin was infused in CGRP-deficient diabetic animals there was also a diminished response. Diabetes resulted in an increased renal vascular flow in response to insulin. CONCLUSIONS: From the present studies we conclude that the insulin-mediated vasodilation was due, in part, to the stimulation of perivascular nerves to release CGRP, and the action of CGRP on vascular smooth muscle enhanced directly or indirectly the vasodilatory response to insulin. PMID- 12112944 TI - Practice makes perfect. PMID- 12112943 TI - Current literature in diabetes. PMID- 12112945 TI - Women's health: we are only at the foothills. PMID- 12112946 TI - Bacterial vaginosis and other asymptomatic vaginal infections in pregnancy. AB - Preterm birth is a common cause of neonatal morbidity and mortality. Many asymptomatic genital infections have been associated with preterm birth, but attempts to determine a causal relationship between specific infections and preterm birth have been disappointing. Treatment trials of specific infections have generally failed to show a positive effect, and in some trials have shown a deleterious effect. Although there is a strong association between the presence of bacterial vaginosis and Trichomonas vaginalis in pregnancy and preterm birth, randomized treatment trials have failed to show a benefit of treatment of these organisms. Treatment of asymptomatic bacterial vaginosis or T. vaginalis to prevent preterm birth is not warranted. PMID- 12112947 TI - Antibiotics and preterm labor. AB - Prematurity is a profound obstetric problem and to date no effective treatment or prevention strategies have been found. Many animal and clinical data exist to link infection and preterm labor, yet clinical trials examining the effect of antibiotic treatment in patients with patterns labor and intact membranes have been conflicting and disappointing. Beyond treatment to reduce neonatal group B streptococcal infection, sexually transmitted infections, symptomatic bacterial vaginosis, and bacteriuria, no clinical data exist at this time to support the routine use of antibiotics in patients with preterm labor and intact membranes. PMID- 12112948 TI - Hepatitis C in pregnancy. AB - In epidemiologic studies done primarily in Europe and in the United States, antibody to hepatitis C has been present in approximately 1% to 4% of pregnant women. Although close to 50% of infected women have no known risk factors for infection, routine screening of pregnant women is not currently recommended. Hepatitis C virus (HCV) may be transmitted to the newborn at a rate of approximately 5%; it is not clear whether this occurs in utero, intrapartum, or both. Factors that increase the risk of vertical transmission include concomitant HIV infection and a high maternal HCV viral load. Breast feeding does not appear to significantly increase the risk of neonatal HCV infection. There is currently no treatment for HCV infection that is approved for use in pregnancy or for use in the neonate. PMID- 12112949 TI - Recurrent vulvovaginal candidiasis. AB - Widespread use of over-the-counter antifungal medications has contributed to a large increase in the number of women who experience more than three episodes of candida vulvovaginitis per year. These women are particularly prone to chronic vulvovaginal pain syndromes; as such, the value of aggressive therapy based on detailed diagnosis extends well beyond immediate symptom relief. Diagnosis is complicated by the fact that a larger proportion of cases of are due to non albicans species, which are not readily identifiable at office evaluation, and points to the value of fungal culture in such cases. Although most Candida albicans are sensitive to azole antifungals, non-albicans species are more often resistant, necessitating alternative therapies. In many cases therapy aimed at suppression of recurrence must extend 6 months. Ongoing studies may identify host factors that facilitate recurrence, and thus provide the basis for individually targeted therapy. PMID- 12112950 TI - Outpatient management of pelvic inflammatory disease. AB - Pelvic inflammatory disease (PID) is a spectrum of inflammatory disorders of the female genital tract involving at least the endrometrium and may include the fallopian tubes, ovaries, and pelvic cavity. Over 1 million women each year are treated for PID in the United States, and it is one of the most serious infections diagnosed in women due to its sequelae. Women with PID acutely experience pain and are at risk for sepsis; however, the significant increases in ectopic pregnancy and infertility are the most disturbing long-term complications. It most often is initiated with an infection by a sexually transmitted disease, but can also involve a variety of pathogenic aerobes and anaerobes secondarily. PMID- 12112951 TI - Leak point pressure measurement and stress urinary incontinence. AB - This article reviews literature on leak point pressure (LPP) measurement as a diagnostic test for stress urinary incontinence in women. LPP is not consistently defined and techniques are not standardized, introducing variation in test results. Reproducibility of LPP is poor, both because of biological variation and variation within the test procedure itself (related in part to lack of standardization). Although not well studied, LPP values do correspond to the severity of incontinence symptoms, as a quantitative indication of the level of urethral dysfunction. However, there is no prospective evidence to support the commonly used cutoff of 60 cm H2O as an indication of intrinsic sphincter deficiency. LPP is potentially useful as a clinical and research tool for evaluating stress urinary incontinence in women. However, its use in clinical management is not well supported by evidence and further research is critically important to define its role. PMID- 12112952 TI - Use of synthetic mesh and donor grafts in gynecologic surgery. AB - Traditional surgery for the correction of pelvic organ prolapse continues to result in suboptimal long-term cure rates. In an effort to improve clinical outcomes, various new surgical techniques have been proposed and use of synthetic and donor graft has been advocated. Although the technique of graft placement for the correction of anterior, posterior, and apical vaginal wall reconstruction is easy to perform, controversy exists regarding the optimal choice of material. Synthetic materials have the advantage of being readily available, cost effective, and consistent in quality, but may present with significant complications, including infection and erosion. In contrast, autologous and heterologous donor grafts provide naturally occurring biomaterials that may undergo desired remodelling, but the in vivo tissue response is still not fully understood. The use of graft materials is still in an early period of evaluation and it is expected that its use will steadily rise with increasing experience and new product development. The following review analyzes our current experience with the use of graft materials in reconstructive pelvic surgery. PMID- 12112953 TI - New therapeutic options for urge incontinence. AB - A critical evaluation of the literature published over the past year reveals several therapeutic options for urge incontinence. Basic science advances in understanding the pathophysiology of bladder instability are paramount in the development of new therapeutic options, chief of which is sacral neuromodulation. Epidemiologic studies from around the world impact the therapies and diagnosis. Therapies include hormone delivery systems, pharmaceutical, combined behavioral and drug, botulinum, surgery, magnetic stimulation, and sacral neuromodulation. PMID- 12112954 TI - Recent concepts in fecal incontinence. AB - Fecal incontinence is an inability to defer release of gas or stool from the anus and rectum by mechanisms of voluntary control. It is an important medical disorder affecting the quality of life of more than 2% of the US population. The most common contributing factors include previous vaginal deliveries, pelvic or perineal trauma, previous anorectal surgery, and rectal prolapse. Many physicians lack experience and knowledge related to pelvic floor incontinence disorders, but advancing technology has improved this knowledge. Increased experience with endoanal ultrasound and endoanal magnetic resonance imaging have given us better understanding of the anatomy of the anal canal, and new techniques with muscle translocation and artificial neosphincters and neuromodulation have expanded our armamentarium of options for restoring continence. PMID- 12112955 TI - Prevention of childbirth injuries to the pelvic floor. AB - The majority of childbirth injuries to the pelvic floor occur after the first vaginal delivery. Cesarean sections performed after the onset of labor may not protect the pelvic floor. Elective cesarean section is the only true primary prevention strategy for childbirth injuries to the pelvic floor. Alternative primary prevention strategies include elective cesarean section for women with nonmodifiable risks for childbirth injuries to the pelvic floor, antepartum pelvic floor exercises, or intrapartum pudendal nerve monitoring. Secondary prevention strategies must focus on modifying obstetric practices that predispose women to pelvic floor injury. These factors are best delineated for anal incontinence and include restrictive use of episiotomy, mediolateral episiotomy when necessary, spontaneous over forceps-assisted vaginal delivery, vacuum extraction over forceps delivery, and antepartum perineal massage. Finally, tertiary prevention strategies should address the mode of delivery made for women with childbirth injuries to the pelvic floor who desire future fertility. PMID- 12112956 TI - Adolescent pregnancy trends and demographics. AB - In the United States today, 9% of women aged 15 to 19 years become pregnant each year: 5% give birth, 3% have induced abortions, and 1% have miscarriages or stillbirths--rates much higher than those in other developed countries. Rates are highest among those who are older, from disadvantaged backgrounds, black or Hispanic, married, have much older male partners, and live in southern states. Teen pregnancies are overwhelmingly unintended, reflecting substantial gaps in contraceptive use, and difficulties using reversible methods effectively. Teen pregnancy, birth, and abortion levels have decreased in recent years, primarily because of more effective contraceptive use (responsible for about 75% of the decline), and because of fewer adolescents having sexual intercourse (about 25%). Much work remains to improve the conditions in which young people grow up, provide them with information and education regarding sexuality and relationships, and improve access to sexual and reproductive health services. PMID- 12112957 TI - Techniques of testing for sexually transmitted diseases. AB - Adolescent and young adult women are the highest-risk group for nearly all sexually transmitted infections. This article reviews diagnostic methods for the most common bacterial and viral sexually transmitted infections, with special attention to the use of nucleic acid amplification methods, as well as the utilization of nontraditional clinical specimens. These new modalities should help the care provider identify and manage the large asymptomatic pool of infected patients, and further lower the prevalence and burden of infection. PMID- 12112958 TI - New options in contraception for adolescents. AB - There have been several recent advances in the contraceptive methods available to adolescents in the United States. A new monthly injectable method combines efficacy and ease of compliance with excellent menstrual cycle control. Very low dose oral contraceptive pills containing gonane progestins decrease the incidence of estrogen-related side effects, and are associated with low rates of breakthrough bleeding. Oral contraceptive pills prescribed in continuous cycles can provide relief from menstrual-related symptoms, and may improve contraceptive effectiveness. Noncontraceptive benefits of oral contraceptive pills, such as improvement in dysmenorrhea and acne, may motivate more consistent pill-taking, and should be identified as additional reasons for pill continuation. Maximizing the prescribing time limit of emergency contraception to 120 hours after unprotected intercourse may improve access. Emergency contraception is more effective the sooner it is used, and should be provided in advance to adolescents for immediate use in the event a postcoital method becomes necessary. PMID- 12112960 TI - Preimplantation genetic diagnosis. AB - Preimplantation genetic diagnosis (PGD) has now been used in human fertility centers for a decade. To this end, diagnostic analysis is conducted on polar bodies or single blastomeres from biopsied embryos before the embryos are transferred, allowing the selection of normal embryos before a pregnancy has been established. Advances in technology available for PGD are described, including fluorescent in situ hybridization (FISH), interphase chromosome conversion, comparative genomic hybridization (CGH), fluorescent polymerase chain reaction (PCR), multiplex PCR, and whole genome amplification. These techniques support the diagnosis of a number of diseases at the single-cell level. PMID- 12112959 TI - Early diagnosis, presenting complaints, and management of hyperandrogenism in adolescents. AB - Hyperandrogenism in the adolescent is a common problem that may manifest in several ways, and may be the result of numerous underlying problems. This article reviews hirsutism, acne, adrenal hyperplasia, and polycystic ovarian syndrome. Presenting signs and symptoms, as well as underlying pathophysiology and treatment options, are discussed. PMID- 12112961 TI - In vitro maturation of oocytes. AB - In vitro maturation (IVM) of human oocytes is an emerging assisted reproductive technology with great promise. To be successful, this process must entail both nuclear and cytoplasmic maturation. Endogenous regulation of oocyte maturation is a complex sequence of events regulated by endocrine parameters, oocyte/follicular cross-talk, and intra-oocyte kinase/phosphatase interactions. Although nuclear maturation during human oocyte IVM progresses normally, cytoplasmic maturation is significantly lacking, as exemplified by poor embryonic developmental competence and pregnancy rates. Advances made in immature oocyte isolation and oocyte and embryo culture conditions have increased the clinical feasibility of IVM. However, in order to achieve acceptable birth rates, future studies should focus on characterization and regulation of oocyte cytoplasmic maturation, and how oocyte-derived factors influence zygotic genome activation and embryonic developmental competence. PMID- 12112962 TI - Cryopreservation of ovarian tissue: banking reproductive potential for the future. AB - Cryopreservation of ovarian tissue is a technology that holds promise for banking reproductive potential for the future. It may be apropos for cancer survivors who have undergone treatment with sterility-inducing chemotherapy. Although there is some evidence suggesting cellular and molecular injury with the freezing and thawing process, there are examples in both animals and humans that transplantation of cryopreserved ovarian tissue can effectively bank reproductive potential for the future. This technology may ultimately have applications for in vitro fertilization, and preventing natural or iatrogenic menopause. PMID- 12112963 TI - Recombinant gonadotropins. AB - Recombinant DNA technology makes it possible to produce large amounts of human gene products for pharmacologic applications, supplanting the need for human tissues. The genes for the alpha and beta subunits of follicle-stimulating hormone (FSH), luteinizing hormone (LH), and human chorionic gonadotropin (hCG) have been characterized and cloned. Recombinant FSH (rFSH) has been shown to be safe and effective in the treatment of fertility disorders. In comparison with the urinary gonadotropin products, human menopausal gonadotropins (HMG), and urinary follitropins (uFSH), rFSH is more potent and better tolerated by patients. Recombinant HCG appears to be as efficacious as urinary HCG with the benefit of improved local tolerance. Recombinant LH (rLH) is likely to be recommended as a supplement to rFSH for ovulation induction in hypogonadotropic women. It may also benefit in vitro fertilization patients undergoing controlled ovarian hyperstimulation with rFSH combined with pituitary suppression, with a gonadotropin-releasing hormone agonist or antagonist. PMID- 12112964 TI - Nuclear transfer and cloning. AB - The use of nuclear transfer in human reproductive and therapeutic cloning is reviewed with attention on the origins of this technology from its evolution to the present. The successes and limitations of mammalian reproductive cloning are itemized. A case is made against the use of human reproductive cloning to reproduce an existing person, based on the unacceptable risks to the embryo, fetus, or newborn. However, support is extended for human therapeutic cloning involving the derivation and use of embryonic stem cells to treat human disease. PMID- 12112965 TI - Beyond one's everyday practice: worldwide women's health. PMID- 12112967 TI - Reproductive rights and reproductive wrongs. PMID- 12112966 TI - Violence in the lives of young women: clinical care and management. AB - Millions of female children, adolescents, and young adults from all socioeconomic segments of society will experience physical or sexual violence perpetrated by someone close to them at some point in their lives. Knowledge of risk factors and sequelae associated with interpersonal violence, and of specific screening tools and procedures designed to detect violence, can help clinicians identify potential victims of assault. Careful management and referral of victims is critical. The use of anticipatory guidance to prevent physical and sexual violence is also advised. PMID- 12112968 TI - The mifepristone-misoprostol regimen for early medical abortion. AB - The combination of mifepristone, an antiprogesterone known in the United States as Mifeprex (Danco Laboratories, New York, NY), and misoprostol, a prostaglandin analogue marketed in the United States under the brand name Cytotec (Pharmacia [formerly Searle], Peapack, NJ), provides a nonsurgical abortion in the early stages of a pregnancy. Mifepristone blocks the action of progesterone, a hormone necessary to sustain a pregnancy, whereas misoprostol causes contractions that expel the embryo and other pregnancy tissue. Since September 2000, mifepristone has been approved by the United States Food and Drug Administration for induced abortions of gestations of 49 days or less since the last menstrual period, but is not yet approved for any other uses. Misoprostol has long been approved to prevent and treat stomach ulcers. PMID- 12112969 TI - Release of progestin-only emergency contraception. AB - Progestin-only emergency contraception has been available in a prepackaged product since 1999. In a multicenter randomized trial, the levonorgestrel-only regimen was better tolerated and significantly more effective than the previous standard of care, the Yuzpe regimen. The levonorgestrel-only regimen prevented 85% of unintended pregnancies compared with 57% in the Yuzpe regimen. Emergency contraception is more effective the earlier the treatment begins. With the emergence of specifically prepackaged kits, emergency contraception appears to be more accessible and convenient to providers and to women. However, substantial barriers still exist to women who wish to obtain emergency contraceptive within the recommended initiation of 72 hours after unprotected intercourse. More recent information that emergency contraception is more effective the sooner it is initiated underscores the need for effective educational and distribution strategies. PMID- 12112970 TI - Therapies for the treatment of abnormal uterine bleeding. AB - Abnormal uterine bleeding (AUB) is one of the most common disorders encountered by the gynecologist. Several drugs have been demonstrated to decrease menstrual bleeding in patients with menorrhagia. Non-steroidal anti-inflammatory drugs will decrease bleeding by 30% to 50%. Oral contraceptives may be useful to stop acute bleeding and will decrease menstrual flow by approximately 50%. Tranexamic acid, a plasminogen inhibitor approved for the treatment of hemophilia, will also decrease flow by approximately 50%. Danazol and GnRH analogues both have been used for the treatment of menorrhagia. However, side effects make them unsuitable for long-term use. There are currently two medicated intrauterine devices (IUDs) available in the United States. These IUDs reduce menstrual blood loss by 65% to 85%. Several minimally invasive surgical procedures, including endometrial resection and ablation, may treat menorrhagia in select patients. More recently, various office-based ablation instruments have been developed. These machines conform to the endometrial cavity and may obviate the need for hysteroscopy. PMID- 12112971 TI - Androgen replacement in menopause. AB - Menopause and the years leading to the menopausal transition are associated with significant decline in sex steroid levels. In contrast to the abrupt decline in estrogens at the time of menopause, a fall in the circulating testosterone and the adrenal preandrogens most closely parallel increasing age. Their accelerated decrease occurs in the years preceding menopause. It is now recognized that the decline in androgens plays a significant role in affecting perimenopausal and menopausal symptomatology and quality of life. Loss of circulating levels of androgens affects libido, vasomotor symptoms, mood and well-being, bone structure, muscle mass. Also, it influences cardiovascular profile. In the menopausal female (in whom these symptoms are clearly linked to low levels of bioavailable testosterone levels), hormone replacement therapy may be of benefit. Recently, interest is increasing in the use of androgen replacement not only for women who have undergone premature or surgical menopause but also for those who experience natural menopause and premenopausal loss of libido from diminished free testosterone. PMID- 12112972 TI - Anterior cruciate ligament injuries. AB - Increasingly, epidemiological data has shown differences in the total number of injuries and in the incidence of serious knee injuries among males and females in jumping and pivoting activities. Particularly, strong epidemiological data support increased incidence of non-contact anterior cruciate ligament injuries in females. Despite more than a decade of vigorous debate and intense study on this topic, research has yet to clearly link gender as an individual risk factor for this injury. Risk factors can be divided into intrinsic or extrinsic factors; for this review they are divided into anatomic, hormonal, and neuromuscular categories. The most intriguing of these deals with neuromuscular control of the limb. In particular, neuromuscular issues involving hip and trunk position and its muscular control have been increasingly implicated in this injury etiology. Gender differences have been found in motion patterns, positions, and muscular forces generated with various lower extremity coordinated activities. Thus, many clinicians currently advocate a strengthening program that emphasizes proximal hip control mediated through gluteus and proximal hamstring activation in a close chain fashion. This, combined with skill training in landing and pivoting maneuvers, is thought to help prevent injury. We must encourage continued, structured, and focused research in this area. However, until specific predictive and protective factors are identified, training and prevention programs should continue to be implemented, assessed, and improved. PMID- 12112973 TI - Prevention of anterior cruciate ligament injuries. AB - Numerous studies have found that female athletes who participate in jumping and pivoting sports are four to six times more likely to sustain a knee ligament injury, such as anterior cruciate ligament (ACL) injury, than male athletes participating in the same sports [1-8]. A widening gender gap in the number of serious knee ligament injuries exists due to geometric growth in female athletic participation, coupled with the four- to sixfold higher injury rate. More than 50,000 serious knee injuries are projected to occur in female varsity intercollegiate and high school athletics each year [9, 10]. Most ACL injuries occur by noncontact mechanisms, often during landing from a jump or making a lateral pivot while running [2, 11]. Knee instability, due to ligament dominance (decreased medial-lateral neuromuscular control of the joint), quadriceps dominance (increased quadriceps recruitment and decreased hamstring recruitment and strength), and leg dominance (side-to-side differences in strength, flexibility, and coordination) are possible contributing factors to the increased incidence of knee injury in female athletes [5, 6]. In this review, dynamic neuromuscular analysis (DNA) training is defined, and a rationale is presented for correcting the neuromuscular imbalances that may result in dynamic knee instability during sports play. Dynamic neuromuscular training has been shown to increase knee stability and decrease knee injury rates in female athletes [5, 12.., 13.]. Preliminary research on athlete screening and injury prediction based on the three aforementioned imbalances also is presented with recommendations for developing screening protocols for the identification of high-risk athletes. PMID- 12112974 TI - The name of the game: a review of sexual exploitation of females in sport. AB - Sexual harassment and abuse has been a recognized problem in the workplace, schools, and residential homes for more than three decades. Many professional policies highlight the potential for abusing positions of trust, and therefore forbid intimate relationships between, for example, doctors and patients, and psychologists and clients. Yet, abuse of power in the coach-athlete relationship has only recently been acknowledged. This paper discusses definitions of sexual exploitation, prevalence figures, methods used for entrapping athletes, potential risk factors, signs of abuse and harassment, and the consequences for survivors. PMID- 12112975 TI - Relationship between athletic performance and menstrual cycle. AB - The female sex steroid hormones have multiple actions on body systems other than the reproductive axis. Female athletes, coaches, medical professionals, and researchers have long been concerned about the potential impact of menstrual cycle fluctuations in these hormones on components of athletic performance. Estrogen is known to affect the cardiovascular system, bone, and the brain; progesterone primarily influences thermoregulation and ventilation. Substrate metabolism is likely altered by both hormones. Net physiological effects can be either opposing or synergistic and are determined by the relative proportions of each. Nevertheless, investigations to date have not consistently demonstrated significant differences in aerobic capacity, anaerobic capacity, aerobic endurance, or muscle strength in any specific menstrual cycle phase. The course of some chronic diseases may vary slightly during the menstrual cycle, but the mechanism is currently unknown. Recent research in underlying hormonal causes for anterior cruciate ligament (ACL) injuries also is not convincing. PMID- 12112976 TI - Neuromuscular differences between male and female athletes. AB - Female athletes who participate in jumping and cutting-type sports have a four- to six-fold higher incidence of serious knee injuries compared with male athletes in the same sports. Many of these injuries involve the anterior cruciate ligament, and occur by non-contact mechanisms. The susceptibility of the female athlete's knee to injury is likely multifactorial, although neuromuscular factors seem to have an important role. Recent studies have identified neuromuscular differences between male and female athletes. Specifically, female athletes have decreased stiffness and decreased potential for dynamic stabilization of the knee joint. Proprioception deficits involving the knee and side-to-side strength and coordination imbalances are more frequent in female athletes. Also, female athletes more commonly demonstrate imbalances in strength, timing of activation, and recruitment patterns of the lower extremity muscles. Based on these findings, neuromuscular training programs have been studied in an effort to correct lower extremity neuromuscular imbalances. Preliminary results from these programs have been encouraging in reducing peak landing imact forces in the knee, correcting strength and proprioception deficits, and ultimately in decreasing the incidence of serious knee injuries in female athletes. PMID- 12112977 TI - Commentary review on hypertension. PMID- 12112978 TI - Is less more? Rethinking the extent of surgery for invasive cervical cancer. AB - The surgical treatment of early-stage cancer of the cervix is undergoing renewed scrutiny in an effort to minimize complications, preserve fertility when desired by the patient, and yet avoid any compromise of therapeutic efficacy. With regard to radical hysterectomy, the extent of the parametrial resection and lymphadenectomy are two topics of particular interest. A review of the relevant literature concerning radical hysterectomy and cervical cancer that supports a less radical approach is discussed. For women with stage I cervical cancer, a less radical resection appears to offer more when considered in the context of fewer immediate and long-term complications and comparable long-term survival. PMID- 12112979 TI - Chemotherapy in epithelial ovarian cancer. AB - The management of epithelial ovarian carcinoma can be the most challenging of all the gynecologic malignancies partly because of the paucity of effective screening tools for early-stage disease. Numerous randomized clinical trials identified several cytotoxic agents that are active against epithelial ovarian cancer, but the current standard of care remains optimal surgical cytoreduction followed by platinum and taxane-based chemotherapy. When disease recurs or progresses, alternative chemotherapy regimens often have efficacy, either as a single agent or in combination. While treatment recommendations should be individualized, the disease-free interval after induction chemotherapy is often the most clinically useful parameter to determine selection of second-line therapy. Investigations into novel chemotherapeutic agents and other biologic molecules have led to advances in the therapeutic armamentarium and have improved survival for women with ovarian cancer. PMID- 12112980 TI - Molecular biology of cervical cancer and its precursors. AB - There is strong clinical and experimental evidence that the human papilloma virus (HPV) plays a central role in the development and growth of cervical cancer. However, it is known that the carcinogenesis is a multistep process. Changes in the cytogenetic equilibrium, such as chromosomal imbalances, allelic loss, and structural aberrations, happen during the transformation from normal epithelium to cervical cancer. Numerous studies support the hypothesis that HPV infection is associated with development of malignant or pre-malignant changes of the lower genital tract. However, there is little clinical evidence that HPV detection would allow prediction of development of cervical cancer. Regarding the "human aspect" of carcinogenesis, there are efforts to detect markers that predict the risk of progression. PMID- 12112981 TI - Familial ovarian cancer. AB - Women with mutations in the BRCA1 or BRCA2 genes are at increased risk for the development not only of breast, but of ovarian cancer. The estimated lifetime risk of contracting ovarian cancer for women bearing the mutation is 16% for Ashkenazi Jews and up to 60% for high-risk populations. If a woman is at high familial risk of getting ovarian cancer, an intense screening with transvaginal ultrasound or determination of CA 125 can be done, although these methods have not proved beneficial so far. It is thought by some that the use of the contraceptive pill can prevent up to 50% of family-associated ovarian cancers, but the few existing studies have yielded contradictory results. That prophylactic oophorectomy can prolong the lives of healthy women with a family history or who bear a germline BRCA mutation is quite sure, but it is not helpful in preventing peritoneal carcinoma. The clinicopathologic behavior of mutation associated ovarian cancer differs from the growth pattern of sporadic ovarian cancer. The hope is to develop suitable therapies for both types. PMID- 12112982 TI - Gene therapy of ovarian cancer. AB - For the treatment of ovarian cancer, gene therapy is increasingly viewed as the fourth therapeutic concept (in addition to surgery, chemotherapy, and irradiation). Many approaches that use viral and nonviral delivery systems have been employed to introduce genes into tumor cells, thus changing their malignant phenotype. The development of tissue-specific promoters has enhanced the specificity of adenoviral transduction, the most commonly used transfer method. Phase I clinical trials (targeting p53, BRCA1, Her2/neu, Bcl-2, MDR, EIA, and HSV TK genes) have been performed to test the relative safety of different strategies. Further studies are needed to evaluate the effectiveness of these treatments. New studies must evaluate gene therapy alone and in combination with cytostatic regimens because preclinical studies have shown the chemosensitizing effects of several target genes. The increasing knowledge about the genetic background of ovarian cancer will provide many targets for novel gene therapy approaches. PMID- 12112983 TI - The role of ultrasound in screening for fetal Down syndrome. AB - Antenatal Down syndrome screening begins with maternal age-specific risks. It also may incorporate the maternal serum screen and antenatal ultrasound. These complementary tests improve the efficacy of maternal age in Down syndrome screening. Ultrasound enhances antenatal Down syndrome screening by 1) precise dating of a pregnancy for the interpretation of serum screen results; 2) detecting major structural anomalies that occur in approximately 20% of all second-trimester Down syndrome fetuses; 3) detecting minor markers, such as nuchal fold thickness, pyelectasis, echogenic bowel, short long bones, sandal gap, and hypoplasia of the mid-phalanx of the fifth digit, which are more commonly seen in second-trimester fetuses with Down syndrome; and 4) acting in combination with serum screening in the first trimester using fetal nuchal translucency as the marker. Ultrasound enhances the efficacy of either age or age and the serum screen for the antenatal detection of Down syndrome. PMID- 12112984 TI - Preterm delivery: risks versus benefits of intervention. AB - The objective of this report is to examine the evidence that supports obstetric intervention to prevent preterm birth and to assess the associated neonatal outcomes. The different causes of preterm birth are identified and reviewed. The evidence to support obstetric intervention is examined, and the current protocols for therapy are listed. An example of institutional data review is outlined and includes common neonatal morbidities with comparison to an international data set. This type of review allows clinicians to establish recommendations for obstetric intervention based on neonatal outcome. PMID- 12112985 TI - Recurrent obstetric complications: how placental pathology can contribute to cost effective clinical evaluation and a rational clinical care plan. AB - When a pregnancy is delivered with unexpected outcomes and the potential for newborn risk (growth restriction, preterm birth, preeclampsia) or a pregnancy fails, this fact carries risks for future obstetric complications. There is an ever-expanding range of laboratory tests for patients with "obstetric compromise, not otherwise specified." How can you interpret these test results to give surveillance and treatment only to those patients who will benefit? Four major patterns of placental tissue injury may be distinguished reliably by routine microscopy (acute inflammation, chronic inflammation, maternal/uteroplacental vascular pathology, and clotting problems). For the clinician, this information may serve as a guide to cost-effective and rational patient evaluation and next pregnancy management. PMID- 12112986 TI - Theories in perspective. PMID- 12112987 TI - Developmental trajectories, transitions, and nonlinear dynamical systems: a model of crime deceleration and desistance. AB - Research on crime-related developmental trajectories is reviewed with outcomes revealing the existence of several trajectories rather than a single general pattern. Each trajectory is marked by transitions that define the pattern's path and direction over time. These anticipated transitions differ from the unanticipated transitions known to precipitate crime deceleration and desistance. Borrowing principles from nonlinear dynamical systems theory--sensitive dependence on initial conditions, chaotic attractors, and self-organization in particular--this article offers a model of crime deceleration and desistance in which belief systems congruent with crime are altered in phases--initiation, transition, maintenance--to create belief systems incongruent with crime. The practical implications of this model are discussed and suggestions for future research are outlined. PMID- 12112989 TI - Law, psychology, and the "new sciences": rethinking mental illness and dangerousness. AB - Clinical and legal research on the meaning of mental illness and on definitions (and predictions) of dangerousness continue to offer the mental health law arena mostly disappointing results. In this article, the authors argue that much of this failure is linked to the prevailing Newtonian paradigm of cause-effect relationships, linear logic, and absolute order. In its place, the authors draw attention to the "new sciences"; that is, advances generated from quantum physics and chaos theory. To situate the analysis, the authors explore how psycholegal decision making unfolds. Specifically, the authors examine what contributions the new sciences offer society on the nature and meaning of psychiatric disorder and on the forecasting of violence. Along the way, the authors suggest how the new sciences advance the regard for citizen justice within the domain of mental health law. PMID- 12112988 TI - Social control in China: applications of the labeling theory and the reintegrative shaming theory. AB - This article delineates the underlying philosophy and functions of social control in the Chinese society. This topic is particularly interesting because specific control functions are grounded in a unique macro-control system, which is totally different from that typical of Western countries. The article also scrutinizes the implications of labeling theory and reintegrative shaming theory, as they are elaborated in the West, and tests their sensitivity to cross-cultural differences. Although some caveats are in order, the evidence presented here tends to support the reintegrative shaming theory rather than labeling theory. PMID- 12112990 TI - Stalking, homicide, and catathymic process: a case study. AB - Despite the proliferation of research on various aspects of stalking, there has been relatively little study of stalkers who commit homicide. In this article, a man who stalked and killed a casual acquaintance is reported. He developed an idea to kill her that, at first, seemed ego-alien and unreal but eventually became fixed and was accompanied by a mounting inner pressure to act. The concept of catathymic process--a change in thinking whereby the offender comes to believe that he can resolve his inner conflict by committing an act of extreme violence against someone to whom he feels emotionally bonded--is of particular help in understanding this case, as well as similar cases of stalking that culminate in homicide. PMID- 12112991 TI - The link between recurrent maltreatment and offending behaviour. AB - This article considers recurrent maltreatment and offending behaviour. The sample was 60 males and 19 females (11 to 18 years) resident within a secure institution in England and considered a risk to themselves and/or others. Overall, 20.8% had not experienced maltreatment, 6.5% had experienced a single incident, 11.7% were repeat victims (same perpetrator), 6.5% were revictimised (different perpetrators), and more than half (54.5%) had suffered both repeat and revictimisation. Of those who had committed a violent and/or sexual crime, 74% had experienced some form of revictimisation, compared to 33% of those who committed nonviolent offences. Those young people most likely to have committed violent and/or sexual crimes were those who had been victims of recurrent extrafamilial maltreatment (many of whom had also experienced recurrent intrafamilial maltreatment). Thus, in this sample, revictimisation was associated with serious crimes. However, these findings are preliminary, and prospective research with a larger sample is needed. PMID- 12112992 TI - Inmate family functioning. AB - Two-hundred-nine noninmates and 169 inmates completed questionnaires that assessed retrospective perceptions of 12 dimensions of family life and one overall assessment of quality of family life. Between the inmates and noninmates, means for all dependent variables differed significantly except for self reliance; however, meaningful eta-squares were found only for dimensions of bridging, disengagement, and quality of life. Among the independent-samples t tests for gender, eta-squares were not meaningful. Implications for family life interventions in correctional facilities are suggested. PMID- 12112993 TI - Preventing chronic disease and improving health: broadening the scope of behavioral medicine research and practice. AB - There has been much progress made over the past 50 years in developing and applying the behavioral medicine evidence base to improve the health of individuals and populations. In particular, there has been progress made in applying behavioral and social science research and theories to the challenge of promoting health and disease prevention. These gains notwithstanding, not all sections of the population have benefited equally, either within or between countries. The disparities in social, mental, and physical health between the most advantaged and the most disadvantaged population groups are in the main, as great as, if not greater, than ever. This represents a tremendous challenge to all of us as behavioral medicine practitioners, teachers, and researchers. Although understanding more about the so-called upstream determinants of health that generate such health disparities is clearly very important, this knowledge will only make a difference if it is used to generate appropriate multilevel intervention strategies over a long and sustained period of time. However, such outcomes will also be affected by diverse aspects of the global natural ecological environment, and by striving for what has been described as a health sustaining environment. With the increasing impact of globalization on most aspects of our lives, including health, it is important to consider the implications of this for preventing disease and promoting health across traditional national borders. This should challenge us to think about the importance of appropriate dissemination and diffusion of effective interventions at an institutional or policy level not only within a single country, but also between countries. Given the rapid economic and social changes occurring globally, and the tremendous impact of global environmental change on health, there is a need to broaden the scope and practice of behavioral medicine. PMID- 12112994 TI - Psychosocial vulnerability, hostility, and family history of coronary heart disease among male and female college students. AB - This study evaluated the utility of the psychosocial vulnerability model for understanding the hostility-coronary heart disease (CHD) relationship among college students at risk for CHD. Interrelationships of cognitive, affective, and behavioral hostility with structural and functional social support were examined. College undergraduates with a parental history of CHD (n = 121) and a control group of 125 students with no CHD family history completed measures of hostility and social support. Among women, a significant negative correlation was found between affective-experiential hostility and functional support. Among men, a significant negative correlation was observed between cognitive-experiential hostility and structural support. Path analyses revealed a significant positive effect of expressive hostility on functional support for CHD-negative men and CHD positive women. CHD family history was not associated with hostility or family environment. CHD-positive participants reported less support satisfaction than did CHD-negative participants. Thus, results indicated qualified support for the psychosocial vulnerability model of the hostility-CHD relationship. PMID- 12112995 TI - Living with Huntington's disease: illness perceptions, coping mechanisms, and spouses' quality of life. AB - Chronic illness not only affects the life of those suffering from Huntington's disease but also threatens the quality of life (QOL) of their spouses. In this study, we focus on Huntington's disease (HD). The impact of HD on the QOL of spouses has been hardly studied from a behavioral medicine or health psychology perspective. We hypothesize that spouses' illness perceptions and coping mechanisms will contribute significantly to the prediction of their QOL. Illness perceptions, coping mechanisms, and the QOL of 90 spouses of patients with HD were assessed by means of the Illness Perception Questionnaire, the COPE, and the Medical Outcome Study 36-item Short Form Health Survey, respectively. After controlling for demographic and illness-related variables, coping mechanisms explained a significant amount of variance of spouses' role functioning. Given our results, more empirical and longitudinal research is justified on coping mechanisms and illness perceptions of spouses living with Huntington's disease. PMID- 12112996 TI - Changes in physical symptoms during the menopause transition. AB - This article analyzes physical symptoms experienced by mid-age Australian women in different stages of the menopause transition. A total of 8,623 women, aged 45 to 50 years in 1996, who participated the mid-age cohort of the Australian Longitudinal Study on Women's Health, completed Survey 1 in 1996 and Survey 2 in 1998. Women were assigned to 1 of 6 menopause groups according to their menopausal status at Surveys 1 and 2, and compared on symptoms experienced at Surveys 1 and 2, adjusted for lifestyle, behavioral and demographic factors. At Survey 1, the most commonly reported symptoms were headaches, back pain, stiff joints, tiredness, and difficulty sleeping. Perimenopausal women were more likely than premenopausal or postmenopausal women to report these symptoms. Hot flushes and night sweats were more common among postmenopausal women. Compared with those who remained premenopausal, women who were in the early stages of menopause or perimenopausal were more likely to report tiredness, stiff joints, difficulty sleeping, and hot flushes at Survey 2. Women who remained perimenopausal were also more likely to report back pain and leaking urine. Compared with premenopausal women, odds ratios for night sweats increased for women in consecutive stages of the menopause transition and remained high in the postmenopausal women. PMID- 12112998 TI - Agency in the life course: concepts and processes. PMID- 12112997 TI - Effects of vital exhaustion on cardiac autononomic nervous functions assessed by heart rate variability at rest in middle-aged male workers. AB - We investigated the effects of vital exhaustion (VE) on cardiac autonomic functions in relation to working conditions such as overtime and frequent business trips, and to lifestyles such as smoking on 52 healthy middle-aged male workers. VE was evaluated by an abbreviated Maastricht Vital Exhaustion Questionnaire. Cardiac autonomic function at supine rest was assessed by spectral analysis of heart rate variability in an annual health checkup. The mean amplitude of the high frequency (HF: 0.15-0.4 Hz) component was lower in the high VE group, whereas no significant difference in the ratio of the low frequency (LF: 0.04-0.15 Hz) component power to HF power (the LF/HF ratio) was observed among VE groups. There were significant interactive effects of VE and smoking on HF amplitude, and of VE and frequent business trips on the LF/HF ratio. VE symptoms were related to the suppression of the cardiac para-sympathetic nervous function at rest in middle-aged male workers, but not to the alteration in sympathovagal balance. Smoking and overwork such as frequent business trips may amplify the autonomic dysfunction in relation to VE among workers with a pronounced feeling of VE. PMID- 12112999 TI - History, agency, and the life course. PMID- 12113000 TI - Intentional self-development: exploring the interfaces between development, intentionality, and the self. PMID- 12113001 TI - Parenting, motivation, and the development of children's coping. PMID- 12113002 TI - Are you ready for the genetics revolution? PMID- 12113005 TI - Breast cancer response. PMID- 12113007 TI - When the patient knows best. PMID- 12113008 TI - Hyperbaric oxygen therapy. PMID- 12113009 TI - Emergency. Epiglottitis. PMID- 12113010 TI - Aspergillosis. PMID- 12113011 TI - Narcolepsy: unveiling a mystery. PMID- 12113012 TI - Deceptive placebo administration. PMID- 12113013 TI - When latex allergies limit employment. PMID- 12113014 TI - Living beyond the sword of Damocles: surviving childhood cancer. PMID- 12113015 TI - Carotenoids identify lung cancer risk. PMID- 12113016 TI - Ki-67: potential lung cancer biomarker. PMID- 12113017 TI - Cyberknife image-guided radiosurgery system successful. PMID- 12113018 TI - MedImmune initiates phase I/II clinical study with Vitaxin. PMID- 12113019 TI - New combination promising against colon cancer. PMID- 12113020 TI - Matrix initiates phase II study in hepatocellular carcinoma. PMID- 12113021 TI - BMS and Exelixis target tumor suppression pathways. PMID- 12113022 TI - Anastrozole: pharmacological and clinical profile in postmenopausal women with breast cancer. AB - A significant proportion of breast cancers are estrogen-dependent and are therefore amenable to endocrine therapy. Although tamoxifen has been the mainstream of endocrine treatment for over 20 years, new agents have entered the clinic, which have potentially superior activity and an improved safety profile. The development of orally-active, potent and selective third-line aromatase inhibitors represents a major advantage in the management of hormone-sensitive breast cancer. Anastrozole (Arimidex) was the first of these agents to become available and is currently widely indicated for both first- and second-line treatment for postmenopausal women with breast cancer. This review focuses on the biochemical properties and clinical efficacy of anastrozole, providing an overview of the current clinical status and possible future applications. PMID- 12113023 TI - Rituximab: review and clinical applications focusing on non-Hodgkin's lymphoma. AB - Rituximab (Rituxan) was the first monoclonal antibody approved for cancer therapy and the first single-agent approved for therapy of lymphoma. When combined with CHOP, rituximab is the only drug that has been shown to improve survival of a subpopulation of patients with diffuse large cell lymphoma during the last three decades. It was approved by the FDA for the treatment of patients with relapsed or refractory low-grade or follicular, CD20-positive, B-cell non-Hodgkin's lymphoma in 1997. Rituximab is also being studied in many other B-cell malignancies alone and in combination with other agents. Furthermore, it is currently being evaluated in several nonmalignant diseases, such as autoimmune disorders. This review will focus on the role of rituximab in patients with non Hodgkin's lymphoma. PMID- 12113024 TI - Irinotecan in the treatment of small cell lung cancer. AB - Combination chemotherapy is the cornerstone of treatment of small cell lung cancer, leading to a meaningful survival benefit for these patients. However, there have been no major advances in therapy in the last decade. Therefore, more effective new treatments are necessary to improve the outcome of therapy. Irinotecan is one of the new active agents that provide hope for more effective therapies in the 21st century. A Phase III trial carried out in Japan clearly demonstrated a survival advantage of a combination of cisplatin and irinotecan over the standard regimen of cisplatin and etoposide. This review outlines the treatment results of irinotecan as a single agent as well as in combination with other cytotoxic agents. PMID- 12113025 TI - Management of small cell lung cancer. AB - Small cell lung cancer is a tumor that has a very poor prognosis without treatment. It is however, highly responsive to chemotherapy and radiotherapy. Pretreatment clinical and laboratory parameters--in addition to staging--can prognosticate outcome and help define the aim of treatment. Different schedules of chemotherapy have been developed and varied strategies, such as chemotherapy dose intensification have been tried to improve outcomes. New agents, such as irinotecan, gemcitabine and topotecan have also been tested. Clinical trials have helped to define strategies of integrating thoracic radiotherapy and prophylactic cranial radiotherapy into management of those patients with limited disease to improve survival further. Despite good initial responses to treatment, most patients eventually relapse. Maintenance strategies with ongoing chemotherapy or novel agents, such as interferon, matrix metalloproteinase inhibitors, thalidomide and vaccines are discussed. PMID- 12113026 TI - Advances in systemic therapy of small cell cancer of the lung. AB - Over the last 20 years, progress in the therapy of small cell lung cancer has been painfully slow. Despite dramatic initial responses to chemotherapy, most patients relapse quickly with an overall 5-year survival of about 5%. Recent trials however offer some hope at changing this picture. Combining standard regimens with newer agents has doubled median survival in some cases. The use of novel targeted agents holds the promise of significantly increasing the survival in this disease, with manageable toxicity. This review outlines current treatment strategies, summarizes recent clinical trials and offers a view of what the next 5 years may hold for the treatment of small cell lung cancer. PMID- 12113027 TI - New strategies in the treatment of resectable non-small cell lung cancer. AB - Non-small cell lung cancer is a systemic illness. Given the systemic nature of lung cancer, it seems that chemotherapy should play an essential role. In stage IIIA disease neoadjuvant chemotherapy plus surgical resection improves survival when compared with surgical resection alone. However, randomized trials using postoperative adjuvant chemotherapy with 'older' drugs has shown no substantial improvement in survival. Since new chemotherapeutic agents may provide additional benefits, there are various studies incorporating new agents in the resectable disease treatment setting. One focus for ongoing research is to find better treatment approaches in earlier stages of disease. Some data suggest that induction chemotherapy in stage I-II is feasible and appears not to compromise surgery. Another promising more individual approach is to tailor chemotherapy according to the pattern of genetic variants or abnormalities found in DNA and/or RNA extracted from the bloodstream. Furthermore, at present many types of new agents are available for testing as 'consolidation treatment' following induction treatment, including, angiogenesis inhibitors, antibodies to growth factor receptors, gene therapy and vaccines. PMID- 12113028 TI - Neoadjuvant chemotherapy in early-stage non-small cell lung cancer. AB - Of the patients that undergo complete resection of early-stage non-small cell lung cancer (NSCLC), 30-60% will die. Postoperative adjuvant chemotherapy has yet to demonstrate an unequivocal benefit and there are significant difficulties in administering postoperative chemotherapy to patients with the significant comorbidities found in NSCLC. Currently, several trials are evaluating the role of preoperative chemotherapy in stage I and II NSCLC. This paper reviews the rationale for this approach and potential future developments. PMID- 12113029 TI - Therapeutic advances in the management of metastatic colorectal cancer. AB - Metastatic colorectal cancer has long been considered as a short-term, poor prognosis, chemoresistant disease. Until the early 1990s, the impact of systemic chemotherapy on patient outcome was debated. Recently, the emergence of new therapeutic modalities (5-FU modulations and associations with oxaliplatin and irinotecan) has led to a significant improvement in tumor response rates and patient survival. Thus, the indications of curative surgery of visceral metastases, frequently preceded and followed by chemotherapy, systemic or intra arterial or both, have become more frequent. In this paper we will review and comment on the results of the major clinical trials published in the past 5 years and propose some decision strategies regarding the main clinical situations met in daily practice. PMID- 12113030 TI - Prognostic biomarkers in resected colorectal cancer: implications for adjuvant chemotherapy. AB - Knowledge of the prognostic role of biomarkers in colorectal cancer is limited and the routine determination for clinical practice is not warranted. However, for some of these markers, data are promising enough for further evaluation. This review addresses a comprehensive analysis of prognostic biomarkers in colorectal cancer. Data from published studies were collected and analyzed. A sufficient level of evidence suggests that DNA indexes, angiogenesis indicators and some genetic/biochemical markers identity prognostic differences in patients with early-stage colorectal cancer. High-risk patients could be the target for future adjuvant chemotherapy trials and one or more of these markers may identify prognostic subgroups with the same TNM stage category. PMID- 12113031 TI - Recent advances in the nonsurgical treatment of upper gastrointestinal tract tumors. AB - Cancers of the esophagus, stomach and pancreas constitute a major cause of cancer death worldwide. Despite improvements in both surgical techniques and chemotherapy regimens these tumors remain a great therapeutic challenge and most patients still die of their disease, even after apparent 'curative resection'. There is a clear need for more effective treatments, particularly for patients with advanced disease, since this is the most common presenting stage. In this review we will discuss the clinical strategies in use and identify the best current treatment options based on randomized clinical trial data. Novel strategies under investigation are discussed and speculation made as to which will be standard treatments of the future. PMID- 12113032 TI - Quality of life in patients with cancers of the upper gastrointestinal tract. AB - Accurate assessment of health-related quality of life in patients with upper gastrointestinal cancers is essential to help determine treatment strategies. Questionnaires may be used to screen for physical and psychosocial morbidity, to evaluate new therapies and there is accumulating evidence to suggest that quality of life scores have prognostic value. There are well validated generic measures of quality of life suitable to use in patients with cancers of the upper gastrointestinal tract, but only two systems (EORTC QLQ-C30 and the FACT-G) have site-specific modules that have been constructed for this patient group. The future use of computer-assisted techniques to collect, analyze and interpret quality of life data will enable the implementation of quality of life results in clinical practice. PMID- 12113034 TI - Nonsurgical management of gallbladder cancer: cytotoxic treatment and radiotherapy. AB - Carcinoma of the gallbladder is a rare tumor entity. Apart from surgical intervention, there is no therapeutic measure with curative potential. Therefore, patients with advanced--i.e., unresectable or metastatic-disease present a difficult problem to clinicians, whether to choose a strictly symptomatic treatment or expose the patient to the side effects of potentially ineffective treatment. Despite anecdotal reports about symptomatic palliation and survival advantages, only unrandomized Phase II studies too small to draw meaningful conclusions have been published thus far. Since there is no standard therapy for advanced gallbladder cancer, patients should be offered the opportunity to participate in controlled clinical trials. PMID- 12113033 TI - Management of hepatocellular carcinoma: advances in diagnosis, treatment and prevention. AB - Since the major causes of hepatocellular carcinoma are hepatitis viruses, the difference and similarity of clinical features in relation to the causative virus may indicate that persistent inflammation of the liver is a major role in hepatocellular carcinoma development in both HBV and HCV infection. However, there is a variety of molecular products of virus-inducing mutagenesis, especially in HBV. An advance in the diagnosis of hepatocellular carcinoma is imaging modality to detect hemodynamics of hepatocellular carcinoma with noninvasive methods of ultrasonography and tumor markers. Chemoprevention using synthetic retinoid is another important issue for the prevention of hepatocellular carcinoma development, as well as viral eradication and suppression of inflammation in the liver using interferon and other drugs. PMID- 12113035 TI - Pharmacogenetics of cytotoxic drugs. AB - The main genetic polymorphisms affecting the metabolism and transport of cytotoxic drugs are discussed in this review. Likely future approaches to pharmacogenetics and emerging technologies, such as tumor classification using microarray analysis, are also considered. PMID- 12113036 TI - Clinical applications of radioprotectors. AB - Patients with malignant or benign tumors who receive radiation therapy- frequently in combination with chemotherapy--are likely to experience side effects, either acutely or chronically. After several decades of preclinical and clinical research efforts, a first approved radioprotective drug, amifostine, has been introduced into the clinic. Although thus far it has been demonstrated to be effective only for the prevention of dry mouth following radiotherapy, it has the potential to be applied in many other oncologic situations. PMID- 12113037 TI - New drugs, new drug resistance mechanisms. PMID- 12113038 TI - New studies highlight benefit of Xeloda in new combination treatments for colorectal cancer. PMID- 12113039 TI - ERBITUX as a single agent and in combination in colorectal carcinoma. PMID- 12113040 TI - Docetaxel improves survival and reduces risk of relapse in node-positive early stage breast cancer. PMID- 12113041 TI - Improved survival with combined chemotherapy and radiation in advanced lung cancer patients. PMID- 12113042 TI - Phase II clinical trial of XYOTAX in non-small cell lung cancer to continue. PMID- 12113043 TI - Atrasentan demonstrates survival benefit in hormone-refractory prostate cancer. PMID- 12113044 TI - Cell Genesys reports long-term survival data in Phase II trial of GVAX. PMID- 12113045 TI - Protein design labs begins Phase I trial of Remitogen in cancer patients with solid tumors. PMID- 12113046 TI - GPC Biotech receives Orphan Medicinal Product Designation. PMID- 12113047 TI - Human Genome Sciences announces clearance of IND application for LymphoRad. PMID- 12113048 TI - Letrozole for the management of breast cancer. AB - Letrozole, a third-generation aromatase inhibitor, has been the only aromatase inhibitor to date to show unequivocal superiority to tamoxifen as first-line treatment of metastatic postmenopausal breast cancer. The superiority of letrozole compared with tamoxifen was also reflected in the neoadjuvant setting, in both estrogen receptor-positive and estrogen receptor-unknown patients with differing HER-2 status. Currently, studies are being performed in the adjuvant setting, which will provide important data on the long-term safety of letrozole and help determine its suitability as a chemopreventive agent in healthy women at risk of developing breast cancer. Nevertheless, the superior clinical efficacy and survival data of letrozole suggest that it has the potential to displace tamoxifen as the gold standard in breast cancer treatment in the coming years. PMID- 12113049 TI - Troxacitabine-based therapy of refractory leukemia. AB - Unique among currently approved or in-development nucleoside analogs, troxacitabine (Troxatyl) is an L-nucleoside with significant cytotoxic activity. Its stereochemistry and cellular transport characteristics render it insensitive to some tumor cell mechanisms of resistance to D-nucleosides, such as cytarabine and fludarabine. Troxacitabine's dose-limiting toxicities were mucositis and hand foot syndrome in patients with refractory leukemia. Three complete and one partial remissions were observed in 30 patients with refractory acute myeloid leukemia on a Phase I study. Significant activity in blastic phase of chronic myeloid leukemia was seen on a Phase II study. Combinations of troxacitabine with ara-C, topotecan and idarubicin are active in patients with refractory acute myeloid leukemia (AML). Phase II studies in patients with refractory lymphoproliferative diseases are ongoing. Troxacitabine merits further study in patients with hematological malignancies. PMID- 12113050 TI - Exemestane: a potent irreversible aromatase inactivator and a promising advance in breast cancer treatment. AB - With the introduction of orally-active, potent and selective third-generation aromatase inhibitors and inactivators--anastrozole, letrozole and exemestane- approaches to the treatment of advanced breast cancer are undergoing re evaluation. In advanced breast cancer, aromatase inhibitors and inactivators are likely to become established as the primary choice over tamoxifen in postmenopausal female breast cancer patients when hormonal therapy is indicated in the first-line setting. The current evaluation of exemestane, an oral steroidal irreversible aromatase inactivator, for primary and adjuvant therapy and the potential role of potent estrogen-deprivation therapy in prevention of postmenopausal breast cancer may extend the use of antiaromatase therapy as an increasingly valuable palliative treatment option, conferring survival benefit and possible preventive outcomes across several treatment settings in the management of breast cancer. PMID- 12113051 TI - TNF-alpha targeted therapeutic approaches in patients with hematologic malignancies. AB - Tumor necrosis factor (TNF)-alpha is a major effector and regulatory cytokine with a pleiotropic role in the pathogenesis of several immune-regulated diseases, including graft versus host disease (GVHD) and hematologic malignancies, such as multiple myeloma (MM), myelodysplastic syndrome (MDS) and acute myelogenous leukemia (AML). Curative treatment for the above diseases are not currently available for most patients. Therapeutic approaches inactivating or blocking TNF alpha are being evaluated in clinical trials. This review describes the development of the soluble TNF-alpha receptor (p75 TNF-R: Fc; etanercept) and other agents inactivating or blocking TNF-alpha in the management of patients with hematologic malignancies. The satisfactory safety profile of etanercept--as demonstrated in patients with autoimmune diseases--has been confirmed in patients with hematologic malignancies and GVHD. Studies to assess whether etanercept, either as a single agent or in combination with cytotoxic and/or immune therapy, may increase response rates and/or survival in patients with MM, MDS, AML and other hematologic malignancies are now warranted. PMID- 12113052 TI - Treatment of refractory and relapsed acute myelogenous leukemia. AB - Refractory and relapsed disease occurs in most acute myelogenous leukemia patients. Salvage chemotherapy offers a 30-70% chance of a second complete remission. Unfortunately, this second remission is usually short lived and salvage chemotherapy is rarely curative. Allogeneic bone marrow transplant, either human leukocyte antigen (HLA)-sibling matched or matched unrelated donor, is the only treatment to offer long-term disease-free survival and possible cure. Many patients will be ineligible for allogeneic bone marrow transplant. A conjugated antiCD33 monoclonal antibody, gemtuzumab ozogamicin, has recently been approved for use in relapsed and refractory acute myelogenous leukemia patients. New and novel agents are also undergoing trials in an attempt to improve on the overall poor outcomes. PMID- 12113053 TI - IMRT in the treatment of head and neck cancer: is the present already the future? AB - Disease outcome in locally advanced head and neck cancer patients is far from satisfactory. The main causes of failure remain linked to locoregional recurrences, which are due to incomplete eradication of clonogenic cells. Conventional radiation therapy or 3-dimensional conformal radiation therapy are currently carried out at their extreme possibilities due to their intrinsic limitation--namely the impossibility to generate concave dose distributions without compromising tumor irradiation. Approximately a third of patients treated with radiotherapy and most head and neck cancer cases present concave shapes of the target volumes. With the advent of intensity modulated radiation therapy- clinically available for only few years--head and neck patients can now benefit from strategies based on highly conformal techniques. It is possible to exploit efficiently dose-escalation protocols to increase probabilities to eradicate clonogens, to reduce overall treatment time, to control repopulation problems and to keep as low as reasonably necessary the irradiation of healthy tissues minimizing acute and late complications. Today, both planning and clinical studies demonstrate these advantages but larger controlled trials are necessary to assess the true potentialities of techniques based on intensity modulation for head and neck cancers. In a speculative view, proton therapy, possibly with intensity modulation, or light ion therapy should be considered for selected cases or for reirradiation due to their higher biological efficacy and their degree of dose-conformation to target volumes. PMID- 12113054 TI - Gene therapy for prostate cancer. AB - Prostate cancer is the most common noncutaneous cancer in man. When confined to the prostate it can be cured by radical prostatectomy or irradiation therapy. However, there are no curative therapies for locally advanced, recurrent or metastatic disease. Prostate cancer gene therapy has recently transition from preclinical studies to clinical trials with the goal of developing novel treatments for prostate cancer. The greatest challenge in treating advanced prostate cancer is therapeutic access to and the elimination of metastases. This review details two aspects of prostate cancer gene therapy, the types of delivery systems under development and specific categories of therapeutic genes available with an emphasis on the mechanism of action of specific gene therapy strategies. PMID- 12113056 TI - Polycythemia vera: diagnosis and treatment 2002. AB - Since 1903 when polycythemia vera was designated by Osler as a new identity the clinical manifestations at the time of diagnosis--symptomatology, physical and hematological findings have become well known. Criteria for diagnosis have been established as well as treatment goals. However, agreement on how best to treat this disease has eluded the hematologists particularly as our understanding of the evolution of the hematological findings has become better known. The hemorrhagic and thrombotic complications and acute leukemia in patients managed with myelosuppressive regimens have come to the forefront. Criteria to be used in the comparison of treatment regimens are suggested from which in this author's opinion the use of 32P becomes the treatment of choice, but not all will agree. PMID- 12113055 TI - AntiCD20 mAbs: modifying therapeutic strategies and outcomes in the treatment of lymphoma patients. AB - The first monoclonal antibody for the treatment of cancer, rituximab, was approved 5 years ago and has had a remarkable impact on the treatment of non Hodgkin's lymphomas. Additionally, it sparked a renewed interest in antibodies in general that has revitalized this field of research. Multiple antibodies for a variety of malignant and nonmalignant indications are being evaluated. Some are now approved for anticancer indications (rituximab, ibritumomab, trastuzumab, gemtuzumab, alemtuzumab) and others are completing the review process at US (FDA) and European (EMEA) regulatory agencies. The impact of antiCD20 antibodies, particularly rituximab and ibritumomab, is evident in the way treatment strategies and outcomes have changed for the lymphoma patient. Other unconjugated antibodies (including Hu1D10, epratuzumab and alemtuzumab) and conjugated antibodies (including radiolabeled tositumomab) are under evaluation. PMID- 12113058 TI - The importance of tumor banking: bridging no-mans-land in cancer research. PMID- 12113057 TI - Gold(III) complexes as a new family of cytotoxic and antitumor agents. AB - In recent years, owing to the contributions of a few research groups, some new gold(III) compounds--either simple coordination complexes or organogold compounds -have been prepared that are sufficiently stable under physiological conditions and are promising candidates for pharmacological testing as cytotoxic and antitumor agents. In vitro pharmacological studies point out that some of these novel gold(III) complexes are highly cytotoxic toward cultured human tumor cell lines and are able to overcome resistance to platinum. Significant differences in the spectrum of action were observed compared with cisplatin. Studies are in progress to elucidate the mechanism of action of these compounds. The cellular effects of two representative gold(III) complexes are described. Preliminary results on binding to DNA in vitro are presented, pointing out that the interactions are generally weak. The implications of these results for the development of gold(III) complexes as a new family of cytotoxic and antitumor agents are discussed. PMID- 12113059 TI - Hormonal treatment of endometrial carcinoma. AB - Endometrial cancer has typically been regarded as a relatively benign disease. However, survival rates for patients with advanced-stage or recurrent disease are very poor and more adequate systemic treatment is certainly needed. Being a tumor that arises from a hormone responsive tissue, endometrial cancer is a logical target for endocrine manipulation. This article gives an overview of our current knowledge on hormonal therapy and highlights the large potential for improvement in results of such therapy. Target areas for future research are described. PMID- 12113060 TI - Individualized cancer therapy: molecular approaches to the prediction of tumor response. AB - A major issue with current cancer therapy is how to ensure the maximum benefit with minimum toxicity for individual patients. With the progress of the human genome project have come high-throughput technologies for the analysis of DNA, gene expression or tumor protein in an efficient and quantitative manner. This review will highlight areas of progress with use of molecular approaches to improve the prediction of tumor response, towards the goal of optimized therapy for each individual patient. PMID- 12113061 TI - Anticancer drugs that induce cancer-associated cachectic syndromes. AB - Cachexia--a wasting condition--seriously impairs the quality of life of patients with advanced cancer. Previous studies have shown that several inflammatory cytokines mediate the development of cancer-associated cachexia. Experimentally, cachexia-like symptoms can be induced in tumor-bearing mice and treatment of such mice with chemotherapeutic agents reverses cachexia as a result of its therapeutic action. Nonetheless, cancer chemotherapy occasionally induces anorexia as an adverse reaction. For example, treatment with antitubulin taxanes reduces body weight in tumor-bearing mice more than healthy mice, even when the agents significantly reduce tumor growth. However, the complex relationship between cancer cachexia and the effects of anticancer drugs remains to be elucidated. This review outlines what is known about the development of cachectic reactions, especially in tumor-bearing mice, that occur during treatment with anticancer agents and highlights the clinical relevance of the information. PMID- 12113062 TI - Pemetrexed: a multitargeted antifolate (ALIMTA, LY-231514). AB - Pemetrexed is a new member of the antifolate class of anticancer drugs. Here we describe how it is activated, its mode of action and its pharmacokinetics. Three single-agent Phase I trials are described with particular reference to the observed spectrum of toxicity but also anticancer activity. Several areas of antitumor activity have been explored further in Phase II trials and these are also reported here, as are early studies of pemetrexed in combination with other anticancer agents. Although the results of Phase III studies are not yet available, pemetrexed has promising activity in a number of solid tumors. We conclude by speculating about its future role in the care of patients. PMID- 12113063 TI - CAMPATH (alemtuzumab) for the treatment of chronic lymphocytic leukemia and beyond. AB - CAMPATH (CAMPATH-1H, alemtuzumab, MabCAMPATH), is a lymphocyte-depleting humanized monoclonal antibody that was recently approved in the USA and Europe for the treatment of chronic lymphocytic leukemia (CLL). It targets CD52--a small glycosylphosphatidylinositol-anchored glycoprotein that is highly expressed on normal T- and B-lymphocytes and on a large proportion of lymphoid cell malignancies--but not on hematopoietic progenitor cells. CAMPATH was shown to be effective against CLL refractory to chemotherapy with an acceptable toxicity profile. CAMPATH is also active against T-cell prolymphocytic leukemia and has been extensively used to prevent graft-versus-host disease associated with bone marrow transplantation. CAMPATH is owned by ILEX Pharmaceuticals LP and distributed by Schering AG and its US affiliate Berlex Laboratories. PMID- 12113064 TI - Hormonal therapy for prostate cancer: past, present and future. AB - Prostate cancer is the second leading cause of cancer mortality in men in the USA. For the past six decades, hormonal therapy has been the main treatment of advanced prostate cancer. Hormonal therapy has developed from a surgical procedure to a complex pharmacological treatment. Trials comparing the efficacy of different monotherapies have demonstrated the equivalence of DES, LHRH agonists and orchiectomy. Combined androgen blockade has been compared with monotherapy. However, the results of the different trials have been conflicting. Novel hormonal therapy schedules involving intermittent treatment and peripheral androgen blockade are currently in clinical trials. The role of hormonal therapy in locally advanced disease as part of a multimodality therapy is a new and rapidly developing aspect of hormonal therapy. The mechanism of hormone refractoriness in prostate cancer is an active area of basic science and translational research. PMID- 12113065 TI - Staging in prostate cancer. AB - This article reviews the conventional current techniques that are used for staging prostate cancer. The advantages and limitations of each modality are described. Attention is focused on the areas in which progress is rapidly being made and is likely to be developed in the future. PMID- 12113066 TI - Pharmacia's SU5416 not effective. PMID- 12113067 TI - EMP combination chemotherapy and low-dose monotherapy in advanced prostate cancer. AB - Many chemotherapeutic regimens combined with estramustine phosphate (EMP) have been elaborated for the treatment of hormone-refractory prostate cancer over 30 years. However, older EMP-based combination chemotherapies with vinblastine, vinorelbine, doxorubicin or cyclophosphamide showed relatively low PSA response rate (25-58%) accompanied with high toxicities. On the other hand, newly developed EMP-based combination regimens with etoposide, pacitaxel, carboplatin or docetaxel demonstrated promising PSA response rate (43-77%) with moderate to severe toxicity in the rate of thromboembolic event (5-18%) and of neutropenia (9 41%). Treatment-related death was less in the latter combination group (5/615, 0.8%) than that in the former group (3/234, 1.3%). Of note, in the docetaxel combination with EMP, PSA response rate is as high as 77% with high rate (41%) of neutropenia but no treatment-related death was observed. Docetaxel combination with EMP seems to be the best regimen, though not completely justified by randomized trials, to be selected in the modern era, which will be followed by paclitaxel, carboplatin and EMP combination with PSA response rate of 71%. In addition, an interim report in 83 patients was presented. They were not consecutively enrolled but were treated on low-dose EMP monotherapy for previously untreated advanced prostate cancer in Department of Urology of Tokyo University and our 21 affiliated hospitals. Overall PSA response rate was as high as 93.4% out of 76 assessable patients. However, overall toxicity rate was abnormally high (39.5%) with drug discontinuation rate of 32.1%. The reason of low drug compliance may be attributed to gastrointestinal symptoms. To overcome the low drug compliance, appropriate patients for EMP administration should be selected by using gene analysis on the basis of sophisticated tailor-made medicine. PMID- 12113068 TI - Genta initiates trials with Genasense. PMID- 12113069 TI - British Biotech and MethylGene sign agreement on Phase II antisense drug. PMID- 12113070 TI - Researcher makes breakthrough in cancer research. PMID- 12113071 TI - Current approaches of neoadjuvant chemotherapy in cervical cancer. AB - Despite remarkable improvement in clinical management, the survival of cervical cancer patients has shown only minor progress in the last decade, particularly in patients with advanced and high-risk disease. Multimodal treatment option has been investigated, such as the concurrent use of chemotherapy and radiation, neoadjuvant chemotherapy and radical hysterectomy, or neoadjuvant chemotherapy followed by radiotherapy. Recently, a flow of randomized clinical trials have demonstrated a benefit from the concurrent chemoradiation for the treatment of the cancer of the cervix. This review will summarize the role and benefit of neoadjuvant chemotherapy in combination with sequential or concurrent radiotherapy and radical surgery for treatment of cervical cancer. PMID- 12113073 TI - 10th International Conference on Gene Therapy of Cancer. 13-15 December, 2001, Sidney Kimmel Cancer Center, San Diego, CA, USA. AB - This annual meeting highlighted some incremental advances in the development of gene therapeutics, with genetic prodrug activation, novel tumor suppressor genes and replicating viral vectors at the leading edge for clinical application. Imaging technologies are proving to be very useful for pharmacokinetic and pharmacodynamic studies in preclinical models and should be translated into systems that can help in the development of surrogate markers in clinical trials. PMID- 12113072 TI - Chemoradiation in cervical carcinoma: a must? AB - Cervical carcinoma is one of the most frequent gynecological malignancies. Literature shows that while the rate is exceedingly low in systematically screened populations, the incidence remains high because of large populations of at-risk women--particularly in underserved nations and in medically indigent subpopulations of Western nations--who are not screened. Recently, a series of randomized trials has demonstrated the possibility to dramatically improve the prognosis of these patients by using concurrent chemoradiation. In particular, concurrent chemoradiation represents a major treatment option in patients with bulky IB-IIA disease, IIB-IVA disease and resected IB-IIA disease with poor prognostic factors. However, further studies are necessary to optimize treatment schedules and particularly to define the best drug combination to be used during radiation therapy, improve patients selection by the analysis of anatomical (TNM stage) and non-anatomical (tumor oxygenation, genetic markers, tumor angiogenesis) prognostic factors, explore novel treatment strategies, such as use of neoadjuvant chemoradiation in locally advanced tumors, integration of antiangiogenetic therapies in chemoradiation schedules, use of supportive treatments aimed to overcome tumor hypoxia, to evaluate the possibility of 'cure' of locally recurrent tumors by chemoradiation and finally to define the best 'second-line' treatment for patients failing after chemoradiation with or without surgery. PMID- 12113074 TI - Role of chemotherapy in the management of ovarian cancer. AB - Ovarian cancer is a highly chemotherapy-sensitive malignancy. With current treatment, most women presenting with advanced disease will achieve an objective response and experience major and sustained improvement of cancer-related symptoms. Unfortunately, the majority of patients will ultimately recur, such that second-line treatment options will need to be considered. Further complicating the decision-making process in choosing an 'optimal' second-line regimen is a serious paucity of randomized trials in this clinical setting. PMID- 12113075 TI - Novel target antigens for dendritic cell-based immunotherapy against ovarian cancer. AB - Identification of tumor-specific target antigens has been a major hurdle for the treatment of malignant disease by vaccination or immunotherapy. A second challenge has been the induction of therapeutically effective immune responses to these 'self' antigens. The recent recognition of dendritic cells as powerful antigen-presenting cells capable of inducing primary T-cell responses in vitro and in vivo--in combination with identification of tumor-specific antigens--has generated widespread interest in dendritic cell-based immunotherapy against a wide variety of tumors. In this review, a series of recently identified novel ovarian tumor antigens is discussed, and the potential for therapeutic dendritic cell vaccination targeted against these antigens is assessed. PMID- 12113076 TI - Radioimmunotherapy versus traditional, nontargeted forms of systemic cancer treatment. PMID- 12113077 TI - IDM confirms significant results in bladder cancer program. PMID- 12113078 TI - GlycoDesign expands phase II clinical trials for GD0039. PMID- 12113079 TI - Pentostatin effective as preparative regimen for kidney transplants. PMID- 12113080 TI - Promising results for Arimidex and Femara. PMID- 12113081 TI - Proteasome inhibitor holds promise for patients with refractory multiple myeloma. PMID- 12113082 TI - Encouraging results from phase I trial of GVAX. PMID- 12113083 TI - Therapeutic options and treatment of muscle invasive bladder cancer. AB - Carcinoma of the bladder is the second most common genitourinary malignancy. Although several treatments exist, the gold standard therapy for muscle invasive bladder cancer (> or = stage T2) is cystectomy with urinary diversion. We review various surgical treatments for muscle invasive bladder cancer, focusing on the reported survival rates, complications, advantages and disadvantages of each therapeutic modality. PMID- 12113084 TI - Photodynamic therapy for superficial bladder cancer. AB - In photodynamic therapy, a photosensitizing drug is activated by visible light and in the presence of oxygen, results in local cell death. This evolving modality is now being used to treat and palliate a very wide variety of human solid tumors and carcinoma-in-situ lesions. With regard to bladder cancer, advances in drug development and modern light delivery techniques mean that photodynamic therapy shows promise in the treatment of superficial bladder cancer resistant to conventional treatments. PMID- 12113085 TI - Gene therapy for superficial bladder cancer. AB - In this review, the basics of gene therapy and the strategies to increase the therapeutic effect of gene therapy for superficial bladder cancer are discussed. Strategies considered in detail are modification of the structure of vectors, modification of the promoters of viral vectors and the timing and route of vector administration. Although all of these modifications have shown some degree of improvement for gene transfer, the use of polyamides intravesically in conjunction with an adenoviral system shows the most promise and the greatest potential to supplement or even replace the current treatment modalities for superficial bladder cancer. PMID- 12113086 TI - Retinoids in the prevention of bladder cancer. AB - Superficial bladder cancer has an unpredictable natural history but in many patients it has a tendency for multiple recurrences over many years. Chemoprevention approaches are ideally tested in this type of tumor and may delay or prevent recurrences of superficial bladder cancer or prevent progression to invasive disease. Vitamin A and its derivatives, the retinoids, have been studied in detail in this disease. This article reviews the data on this subject and includes in vitro and in vivo preclinical studies as well as the clinical studies performed in the secondary prevention of this disease. PMID- 12113088 TI - Wilms tumor: progress to date and future considerations. AB - Wilms tumor is the most common tumor of renal origin found in children. In the last 50 years, remarkable progress has been made in the treatment and understanding of children with Wilms tumor. Through the development of multiagent chemotherapy and cooperative pediatric interdisciplinary groups conducting large randomized controlled clinical trials, survival has improved dramatically. In the next century it is expected that 80% of children with Wilms tumor will be long term survivors. Therapy is progressing towards a risk-based management based not only on stage and histology but also incorporated genetic markers. This article reviews progress to date and possible future directions in the treatment of Wilms Tumor. PMID- 12113087 TI - New agents for the treatment of renal cell carcinoma. AB - One-third of patients with renal cell carcinoma present with unresectable or metastatic disease. Immunotherapy, the current standard treatment, induces response in only 10-20% of patients. Chemotherapy with current agents is minimally effective. Other approaches including allogeneic stem cell transplant, vaccine and gene therapy and signal transduction inhibitors, offer promise in early Phase studies. This paper reviews the current treatment options and promising new agents in development. PMID- 12113089 TI - Prognostic factors and molecular markers for renal cell carcinoma. AB - Renal cell carcinoma is the most common cancer in the kidney, affecting nearly 30,000 Americans every year and is associated with over 12,000 deaths annually. If detected early, renal cell carcinomas can be cured surgically. However, once metastatic disease develops the prognosis for long-term survival is poor. Unfortunately, one-third of patients have metastatic disease at the time of diagnosis and approximately 50% of the patients undergoing surgical resection for less advanced disease eventually relapse. This review examines the clinical and molecular prognostic tools currently available or under investigation for kidney cancer. PMID- 12113090 TI - Evolving classification of renal cell neoplasia. AB - The development of consensus classifications for renal epithelial neoplasia in 1996 and 1997 led to the recognition of renal adenoma, renal oncocytoma and metanephric adenoma/adenofibroma as benign tumors and conventional (clear cell) renal cell carcinoma (RCC), papillary RCC, chromophobe RCC and collecting duct carcinoma as malignant morphotypes. While the overwhelming majority of renal adenomas and metanephric adenomas are benign, malignant transformation of both types have been described and genetic predictors of malignant transformation are as yet unknown. The main groups of malignant renal tumors are associated with characteristic genetic changes; conventional RCC (-3p), papillary RCC (+7, +17, Y), chromophobe RCC (hypodiploid). Recent studies have also shown focal loss of heterozygosity of 3p segments in papillary and chromophobe RCC, indicating that 3p mutations are not confined to the conventional RCC morphotype and suggesting the presence of an important tumor suppressor gene at this site. Sarcomatoid metaplasia may occur in any morphotype and this is associated with a poor prognosis. More recently additional varieties of conventional RCC (multilocular cystic RCC), collecting duct carcinoma (medullary renal carcinoma) and papillary RCC (Types 1 and 2), each showing a characteristic morphology, have been recognized. PMID- 12113091 TI - Molecular and pharmacological strategies to overcome multidrug resistance. AB - Multidrug resistance is a major obstacle to the effective treatment of cancer. Despite vast improvements in our understanding of the mechanisms of drug resistance, relatively few significant advances have been made towards effectively circumventing it in a clinical setting. The ability to modulate multidrug resistance has been complicated by the fact that many human tumors simultaneously exhibit multiple resistance mechanisms. In order to effectively overcome multidrug resistance it will be necessary to design new strategies that combine multiple modulating agents and approaches. This review provides an overview of the major causes of multidrug resistance and summarizes many of the current approaches being taken to overcome it. We also describe how liposomal drug delivery systems can be utilized to aid in achieving these goals. PMID- 12113092 TI - Genetic alterations and chemotherapeutic response in human diffuse gliomas. AB - Although for many years adjuvant chemotherapy has been used in the management of human diffuse gliomas, the survival of a large proportion of patients with these tumors has remained unchanged. To date, little is known about the factors that render some of these patients resistant to cytotoxic drugs. Since response to chemotherapy has been heterogeneous irrespective of glioma type, it has been proposed that some genetic alterations associated with glioma formation and progression may be responsible for the variations in the sensitivity of these tumors to adjuvant chemotherapy. In this paper, we review the literature and discuss the usefulness of some of the common genetic alterations in predicting chemotherapeutic response in gliomas. PMID- 12113093 TI - Targeted therapy in oncology: the agony and ecstasy of personalized medicine. PMID- 12113094 TI - Study shows 2-year survival advantage for docetaxel. PMID- 12113095 TI - Genzyme Molecular Oncology receives patent for melanoma immunogens. PMID- 12113096 TI - FDA grants priority review for ZOMETA. PMID- 12113097 TI - Novuspharma announces preliminary results of phase II trials for BBR 3464. PMID- 12113098 TI - GTx begins next phase of clinical trials for prostate cancer. PMID- 12113099 TI - Capecitabine in the management of colorectal cancer. AB - Significant progress has been made in the chemotherapy of colorectal cancer. The author discusses new available options and the development of a new oral fluoropyrimidine, capecitabine (Xeloda). The rational development of this targeted drug with its selective activation in tumor tissue is highlighted. The clinical development of capecitabine and its present and future role in the management of colorectal cancer are reviewed. PMID- 12113101 TI - AE-941 (Neovastat): a novel multifunctional antiangiogenic compound. AB - AE-941 (Neovastat) is a naturally occurring product extracted from cartilage and has antiangiogenic properties. It has reached Phase III clinical trial evaluation for the treatment of solid tumors (non-small cell lung cancer and renal cell carcinoma) and a pivotal Phase II clinical trial in multiple myeloma is ongoing. AE-941 inhibits several steps of the angiogenesis process, including matrix metalloproteinase activities and VEGF signaling pathways. Moreover, AE-941 induces endothelial cell apoptosis and tissue-type plasminogen activator activity, thus suggesting that it is a multifunctional antiangiogenic drug. Results from Phase I/II clinical trials indicate that AE-941, given orally, is well tolerated. Moreover, the median survival time in patients with renal cell carcinoma and non-small cell lung cancer was significantly longer in patients receiving high doses of AE-941 compared to low doses. PMID- 12113100 TI - Raloxifene for the treatment and prevention of breast cancer? AB - Raloxifene is a member of a family of drugs known as selective estrogen receptor modulators (SERMs). Raloxifene is currently approved by the FDA for the prevention and treatment of osteoporosis in postmenopausal women. SERMs hold the potential to treat and prevent breast cancer, osteoporosis and coronary heart disease. Ongoing clinical trials are in place to address the role of raloxifene and SERMs in each of these areas. We review the pharmacology, clinical utility, safety and tolerability of raloxifene and speculate on what the future holds for SERMs and their use in breast cancer. PMID- 12113102 TI - New perspectives for the diagnosis and treatment of oligodendroglioma. AB - Oligodendroglial tumors (OD) constitute a specific type of glioma, with a better prognosis than astrocytic tumors of similar grade. OD are sensitive to chemotherapy, with 60-65% of patients responding to PCV chemotherapy and a median response duration of 1-1.5 years. This holds for both low- and high-grade OD. Despite this high response rate, most if not all patients are ultimately relapsing, requiring improvement of treatment. The presence of chromosomal lesions on the loci 1p36 and 19q13.3 is related to a favorable response to both chemotherapy and radiation therapy. In the near future molecular analysis will become an important tool to identify glial tumors that are likely to respond to treatment and will be used to select patients for clinical trials. Temozolomide appears to be a promising drug in OD. PMID- 12113103 TI - Changing boundaries in the treatment of malignant gliomas. AB - Malignant gliomas are still among the most lethal and difficult tumors to treat; even the most intensive combinations of radio- and chemotherapy are not curative and yield only a modest impact on survival for most of these patients, as long term survivors are less than 10%. There is a major need for new chemotherapeutic drugs and alternative therapeutic modalities. This review aims to define the best standard treatment in the common clinical practice and also summarizes the most promising lines of investigational research in the field of neuro-oncology, which will probably offer new and long-awaited valid therapy options for brain tumor patients. PMID- 12113104 TI - Advances in diagnosis and therapy of neuroendocrine tumors. AB - Neuroendocrine tumors represent a group of neoplasias characterized by significant histopathologic and biological heterogeneity. Diagnosis of neuroendocrine tumors relies upon histological examination augmented by newer techniques such as position emission tomography, meta-iodobenzylguanidine scintigraphy or octreoscan. Surgery represents the definite and curative therapeutic approach in early phase tumors. In metastatic or advanced disease, medical treatment is the best choice. Somatostatin analogs allow adequate control of the carcinoid syndrome, without a significant effect on tumor cell growth. Interferon-alpha may represent an alternative, alone or in association with somatostatin analogs. Chemotherapy is the best choice in the treatment of neuroendocrine tumors characterized by a poor differentiation grade and a high proliferation rate. PMID- 12113105 TI - Therapeutic management of primary CNS lymphoma in immunocompetent patients. AB - The best therapeutic management in primary CNS lymphomas remains to be defined because of the small size and short follow-up of retrospective series, the methodological pitfalls and limited number of prospective studies, and the paucity of randomized trials. The purpose of this article is to analyze, discuss and summarize the current therapeutic approaches, namely chemotherapy or radiotherapy as exclusive treatment, combined treatment, most commonly used drugs, intrathecal chemotherapy and consolidation radiotherapy and to provide recommendations for ordinary clinical practice. Some important therapeutic issues such as the management of intraocular lymphomas, elderly patients and patients without histological diagnosis, as well as the relevance of salvage therapy as a playground for the evaluation of new drugs are also analyzed. Finally, the main open questions as well as current and expected investigation trends are discussed. PMID- 12113106 TI - Current epidemiological trends and surveillance issues in brain tumors. AB - The absence of an overall increase in incidence rates for all primary brain tumors since the 1950s argues against a recently introduced environmental tumorigen impacting these tumors. Historical increases in brain cancer mortality and incidence rates appear to be leveling off following the widespread introduction of CT and MRI scans, indicating that increases in overall rates of malignant tumors are likely to be an artifact of diagnosis and reporting issues. Further studies are needed to understand those tumor types with rates that do appear to be increasing among adults; specifically lymphomas, nerve sheath tumors, pituitary tumors and ependymomas. Patterns of incidence by race, ethnicity, socioeconomic status, and seasonal and regional variation would assist in directing relevant new research questions. Filling in the gap of information on patterns for prevalent, second primaries and metastatic tumors may be useful in understanding the public perception regarding brain tumor rates and would be a valuable addition to healthcare planning tools. PMID- 12113108 TI - Use of adjuvant therapy in cutaneous melanoma. AB - Surgery remains the principal treatment for local, regional and isolated metastatic melanoma. Adjuvant therapy is now available for patients with high risk of recurrence after surgical treatment but is controversial and inconsistently used around the world. In 1995, high-dose interferon-alpha 2b was approved by the FDA, providing clinicians with the first adjuvant therapy for use outside a clinical trial. In this review, we discuss surgical approaches to the management of the primary lesion and regional lymph nodes and the use of high dose interferon. We will also provide guidelines for the use of interferon and discuss current clinical trials evaluating alternate forms of adjuvant therapy. PMID- 12113107 TI - Therapy options in cutaneous T-cell lymphoma. AB - The treatment of cutaneous T-cell lymphoma (CTCL) is continually evolving, as new and emerging drugs are added to the growing arsenal of CTCL therapy. The availability of newly approved investigational therapies, such as bexarotene, denileukin diftitox (DAB389- IL2), monoclonal antibodies and novel chemotherapeutic agents, adds complexity to decisions on the management and treatment of CTCL patients. In formulating a treatment plan, therapeutic options are best approached through consideration of overall clinical staging (stage IA IVB) and skin staging (T1-T4), which affect prognosis and the characteristics of each individual patient's disease. This article will present and discuss the optimal therapeutic agents for all clinical stages of CTCL patients, based on currently available and investigational agents. PMID- 12113109 TI - Melanoma vaccination: state-of-the-art and experimental approaches. AB - Vaccination therapy for human malignancies is an ever-evolving technology. Great strides are being made in our understanding of the various components of the immune system and how best to manipulate these components. PMID- 12113110 TI - Management of Merkel cell carcinoma. AB - Merkel cell carcinoma is an uncommon cutaneous malignancy. Although it is rare, Merkel cell carcinoma has been described as the most malignant primary skin tumor. It is therefore important that once diagnosed, Merkel cell carcinoma is treated appropriately. The aim of this short review is to provide a summary of the available literature to guide clinicians in the future management of such patients. Inevitably in such a rare disease, there are no randomized trials of therapy. The treatment of individual patients will rely on opinion as much as the 'evidence'. PMID- 12113111 TI - Lymphatic mapping and sentinel node biopsy for melanoma. AB - Worldwide experience has now confirmed that the histological status of sentinel nodes (SNs) accurately reflects the status of regional lymph nodes in patients with melanoma. Preoperative lymphoscintigraphy has proved to be invaluable in identifying SNs and in demonstrating unusual lymphatic drainage pathways. Greatest accuracy at the time of surgery is achieved using both blue dye injection and a gamma-probe. Close cooperation between surgeons, nuclear medicine physicians and pathologists is required if the SN biopsy technique is to be reliable. The results of clinical trials will demonstrate whether SN biopsy has any therapeutic value. It is already clear, however, that SN examination provides very accurate staging, and SN biopsy is therefore essential for all melanoma patients entering adjuvant therapy trials. PMID- 12113113 TI - Radiolabeled immunoglobulin therapy: old barriers and new opportunities. AB - The development of cancer-selective therapies, in particular radiolabeled immunoglobulin therapy (RIT), has stalled. RIT limitations/opportunities are identified in translational research in nude mice, beagles and rhesus monkeys, and in patients with Hodgkin's disease, neurological paraneoplastic syndromes or small vessel vasculitis. Intravenous RIT is most successful in patients with hematological malignancies due to high tumor uptake and long tumor retention of radiolabeled immunoglobulins. Patients with solid tumors are only expected to benefit from RIT by the administration of radiolabeled immunoglobulins directly into the tumor. Tumor-reactive IgG is the best vehicle for i.v. RIT. Tumor reactive IgM is the best vehicle for intratumoral RIT. The authors do not intend to review the whole clinical RIT experience, but instead analyze the current limitations to success and how they can be circumvented. When the scientific community can reach a consensus on the development and use of these promising and economical radiopharmaceuticals, increasing numbers of patients with recurrent cancer will start to benefit from RIT. PMID- 12113112 TI - Issues in the epidemiology of melanoma. AB - Cutaneous melanoma is a significant health problem throughout the developing world. Primary and secondary prevention are discussed. The wavelengths of the ultraviolet radiation spectrum and their association with melanoma are discussed. Although excessive sun exposure during childhood is a critical risk factor, excessive sun exposure during adult years is also important. The major risk factors for melanoma--numerous or atypical moles and a sun-sensitive phenotype- are genetic. Their interaction with sun exposure is currently being examined, as well as the interaction of other genetic factors, such as alterations in the melanocortin receptor and the familial melanoma gene, INK4A. Secondary prevention strategies include self-examination and physician examination. New technologies are being developed to supplement visual examination of suspected lesions. These technologies are discussed in detail and include digital photography, digital dermoscopy, confocal scanning laser microscopy and automated diagnosis systems. PMID- 12113114 TI - Treatment of acute leukemia in children: recent advances and future challenges. AB - Recently advances have been made in the treatment of acute leukemia in children, it is now possible to cure more than 70% of children with acute lymphoblastic leukemia. With the introduction of more intensive chemotherapy regimens in patients at higher risk of relapse and the identification of cases that could be less intensely treated to diminish long-term toxicity, it could be possible to improve these excellent results. In contrast, pediatric acute myeloid leukaemia seems to be a more heterogeneous disease and its response to conventional chemotherapy is not as uniform. Introduction of new and more efficacious therapies is necessary to improve the poor outcome, especially among patients with high-risk features. PMID- 12113115 TI - Recent progress in the pharmacotherapy of cancer pain. AB - Cancer pain can be relatively well managed with primary therapies, according to the WHO ladder. However, different conditions may limit the response to the analgesic drug used, which are mainly oploids. Specifically, adverse effects may prevail against the analgesic activity in the clinical setting. New pharmacological strategies may enable a more satisfactory response to be obtained, in terms of balance between analgesia and adverse effects. The change of route of administration or the use of alternative opioids is a first-line option. The use of adjuvant drugs may also improve analgesia with different mechanisms. Recent studies have demonstrated the value of these alternative approaches. However, most of them require definite confirmation in specific well designed studies. PMID- 12113116 TI - Pediatric leukemia in the new millennium. PMID- 12113117 TI - New chemoradiotherapy combinations in head and neck cancer. AB - Head and neck squamous cell cancer is the sixth most common cancer worldwide. Surgical management is effective as a single modality only for early-stage cancers (30% of patients). Making progress in this cancer is of vital interest. The standard treatment for advanced disease is chemoradiotherapy, with the goal of palliation of symptoms and prolongation of life. Response rates to even the best of regimens are approximately 30-40%, with the median survival period of approximately 6 months. Various approaches to treatment of recurrent disease are under investigation, including new drugs or combinations of drugs, with radiotherapy. PMID- 12113118 TI - Molecular biomarkers in HNSCC: prognostic and therapeutic implications. AB - The prognosis of squamous cell carcinoma of head and neck (HNSCC) is influenced by many factors, such as performance status, TNM staging and pathological grading of differentiation. However, these factors are not sufficient for predicting outcome. Therefore, recent research has focused on the identification of molecular biomarkers. These markers help to stage patients in more meaningful prognostic groups and identify high-risk patients who may benefit from a more aggressive treatment approach. They also identify patients who are resistant to radiotherapy or chemotherapy, potentially avoiding the morbidity and cost of ineffective therapies. They can also identify patients with a high risk of recurrence who may benefit from chemoprevention. Finally, these markers may serve as targets for novel therapies, which would eventually change the outcome of HNSCC. PMID- 12113119 TI - Can cure be achieved in patients with head and neck carcinomas? The problem of second neoplasm. AB - The improvement in locoregional control of head and neck carcinomas over the last few decades does not appear to modify the final survival of these patients, mainly due to the appearance of distant metastases and second neoplasms. This article reviews the topic of second neoplasm in patients with an index tumor in the head and neck, making a special point of the incidence and epidemiology of second neoplasms, the influence on prognosis, etiopathogenic theories and the possibility of prevention. PMID- 12113120 TI - Temozolomide: a novel oral alkylating agent. AB - Temozolomide is an imidazotetrazine with a mechanism of action and efficacy similar to dacarbazine (DTIC). However, it differs from DTIC in that it can be taken orally, degrades spontaneously to an active metabolite and penetrates the blood-brain barrier. It is well tolerated, making it a suitable candidate for combination chemotherapy. Trials to date have focussed on its activity in advanced metastatic melanoma and high-grade malignant glioma. Investigations into other indications, in particular solid tumors with central nervous system metastases, are ongoing. Studies of new drug schedules and of drugs to ameliorate temozolomide resistance offer the prospect of increased efficacy. PMID- 12113121 TI - Present status of management of nasopharyngeal carcinoma. AB - Contemporary imaging with endoscopic examination determines tumor extent and this is reflected in the revised staging system. Radiotherapy remains the primary treatment modality. The use of intensity-modulated radiotherapy aims at enhanced tumor control while reducing adverse effects. Concurrent chemotherapy has shown to improve survival, although its efficacy in endemic regions requires confirmation. Determination of circulating cell-free Epstein-Barr virus DNA might detect early recurrence. For patients who developed recurrence in the neck, surgery provides good salvage. For small tumor in the nasopharynx, good results can be obtained with reirradiation therapy, brachytherapy or surgery. The salvage option depends on the size and location of the tumor as well as the expertise available. The long-term effects of these treatment modalities might be significant. PMID- 12113122 TI - Intraperitoneal chemotherapy in the management of malignant disease. AB - The intraperitoneal administration of antineoplastic agents has been proposed as a method to improve the efficacy of therapy of malignant disease principally confined to the peritoneal cavity. Phase I studies have demonstrated the safety and pharmacokinetic advantage for a number of drugs delivered directly into the body compartment, with surgically-defined responses documented in the Phase II trial setting. More recently reported randomized Phase III studies have revealed a survival benefit associated with the use of intraperitoneal cisplatin, compared to i.v. drug delivery when employed as initial treatment of small volume residual advanced ovarian cancer. While there is far less experience with regional therapy of gastrointestinal cancers, limited data suggest intraperitoneal drug administration should be further explored as a potentially important strategy to improve the outcome for this group of malignancies. PMID- 12113123 TI - Mismatch repair defects as a cause of resistance to cytotoxic drugs. AB - In this short review, we aim to bring together the most recent evidence implicating mismatch repair pathway defects as a cause of drug resistance to a spectrum of chemotherapeutic agents in a variety of cancers. Experimental and clinical studies are discussed and possible strategies that may be employed to overcome the multidrug resistant phenotype afforded by such defects. PMID- 12113124 TI - Thalidomide in the treatment of multiple myeloma. AB - Thalidomide--banned from clinical use in the 1960s because of severe teratogenicity--is now back in clinical practice as an effective agent in the treatment of relapsed and refractory multiple myeloma. Several clinical trials have determined that thalidomide is active in 25-35% of patients with relapsed myeloma. The role of thalidomide in early-stage myeloma is being actively investigated. Thalidomide has antiangiogenic and immunomodulatory properties and is an effective inhibitor of TNF-alpha. However, the mechanism of its action in myeloma remains unclear. Major toxicities of thalidomide include constipation, sedation, skin rash, fatigue and peripheral neuropathy. This paper summarizes the current status of thalidomide in multiple myeloma. PMID- 12113126 TI - FDA approves Novartis' Gleevec. PMID- 12113125 TI - Current therapy and future prospects in lymphoma. AB - Today, lymphoma represents one of the most complex and diverse malignancies in routine clinical practice and huge advances have been made in the pathological classification and treatment of this family of diseases. Progress in treatment means that long-term survival is now an achievable goal for many, although a proportion of patients remain incurable. Developments in both conventional chemotherapy as well as biotechnological advances have opened new therapeutic approaches, many of which have already reached the clinic. PMID- 12113127 TI - Berlex Laboratories to market new treatment for B-CLL. PMID- 12113128 TI - AEterna reports on its phase II trial of neovastat. PMID- 12113129 TI - Aronex Pharmaceuticals reports on annamycin phase I trial. PMID- 12113130 TI - Drug used during radiotherapy helps reduce dry mouth problem. PMID- 12113131 TI - OSI Pharmaceuticals, Genentech and Roche announce data from clinical studies of Tarceva. PMID- 12113132 TI - Advances in the management of patients with non-Hodgkin's lymphoma. AB - Non-Hodgkin's lymphoma is the fifth most common cause of death due to cancer and has been rising at a rate of 4% per year for the last four decades. Although 'traditional' chemotherapy and radiotherapy have had important contributions to improving outcomes, new tools in the treatment of non-Hodgkin's lymphoma are needed. This review describes therapeutic modalities that are currently being used or are in the process of being developed and which are based on concepts divergent from 'traditional' approaches to managing non-Hodgkin's lymphoma. PMID- 12113133 TI - Extranodal lymphoma of MALT-type: perspective at the beginning of the 21st century. AB - Extranodal lymphomas arising from mucosa associated lymphoid tissue (MALT) have become a focus of interest in recent years due to their unique pathological and clinical properties. The link between Helicobacter pylori and the development of gastric MALT-type lymphoma has revolutionized treatment options as up to 80% of patients with early gastric MALT-type lymphoma achieve complete remission of the tumor following eradication of H. pylori. As opposed to surgical intervention, which has been the preferred form of treatment in the past, organ conserving approaches are increasingly being applied, as both irradiation and chemotherapy have given excellent results. However, mature data from prospective, randomized studies taking into account the concept of MALT lymphoma as a distinct entity are still lacking in order to define the optimal approach to the management of MALT type lymphoma. PMID- 12113134 TI - 1st annual oncology update: advances and controversies. Steamboat Springs, Colorado, USA, 13-17 February 2001. AB - In addition to the scientific component to the meeting, the conference was successful in many social aspects. The setting permitted the conference to be informal, with casual dress and lively audience participation involvement. In general, the conference was able to provide an excellent multidisciplinary overview of a wide-variety of oncology topics. PMID- 12113135 TI - Role of anti-idiotype vaccines in the modern treatment of human follicular lymphoma. AB - Cancer vaccines are currently conceived as therapeutic tools, in contrast to the prophylactic vaccines that have resolved the problem of a number of infectious diseases. Among the former, anti-idiotype vaccines for human follicular lymphoma have begun to produce tangible clinical results. Just 10 years ago it was not even known whether patients could be immunized against their own tumor antigens and now as many as two independent Phase III clinical trials based on this finding are underway. The rapidity of this development encourages the hope that active immunotherapy may soon become decisive in oncology. For the time being, many important results have already been achieved: the evidence of vaccine induced, tumor-specific humoral/cellular responses and the first documented molecular remissions following vaccination. PMID- 12113136 TI - Immunotherapy of chronic lymphocytic leukemia. AB - Chronic lymphocytic leukemia (CLL) is typically an indolent B-cell malignancy, primarily affecting the aging population. Standard cytotoxic treatment with alkylating agents or purine analogs is very effective at inducing remission. However, curative treatment is not yet available. Immunotherapy is emerging as an exciting modality with significant potential to advance the treatment of this disease. This review discusses the different modalities of immunotherapy under investigation for the treatment of CLL. These modalities include passive immunotherapy with monoclonal antibodies against antigens on CLL B-cells including CD52 and CD20. Active immunotherapy by vaccination with genetically modified autologous leukemia cells is being evaluated in clinical trials. Allogeneic stem cell transplant for adoptive immunotherapy of CLL is yet another modality being investigated. While this modality may have limited application due to morbidity in older patients, it may result in improved survival and possibly cure. The use of immunotherapy in CLL is in the early stages of development. It is likely that this approach will significantly improve the treatment of CLL and possibly contribute to the cure of this disease. PMID- 12113137 TI - Update on prophylaxis and therapy of infection in patients with chronic lymphocytic leukemia. AB - Infections remain a major cause of morbidity and mortality in patients with chronic lymphocytic leukemia. These patients are predisposed to infection due to the immune compromise inherent to the primary disease and to therapy-related immunosuppression. The introduction of purine analogs and agents such as Campath 1H into the therapeutic armamentarium for chronic lymphocytic leukemia has altered the spectrum of infections. Although bacterial infections are most common, opportunistic infections, such as those caused by Candida, Pneumocystis and herpesviruses, may occur, related to T-cell immunosuppression induced by these agents. We will review the pathogenesis of infection, spectrum of infections, risk factors for infection and approaches for infection prophylaxis and therapy. PMID- 12113139 TI - Hairy cell leukemia: treatment prospects. AB - The recent advances in the management of hairy cell leukemia, a chronic and indolent B-cell lymphoproliferative disorder are reviewed. The introduction of alpha-interferon, purine analogs and recombinant monoclonal antibodies/immunotoxins has dramatically improved the outcome in a disease that once had a dismal prognosis. The underlying genetic defect remains unknown. PMID- 12113138 TI - 4th international symposium on leukemia and lymphoma--molecular pharmacology and new treatment modalities. VU Medical Center, Amsterdam, The Netherlands, 7-10 March 2001. PMID- 12113140 TI - Recent advances in the management of squamous cell carcinoma of the head and neck. AB - Therapeutic outcomes of the currently used chemotherapeutic agents for recurrent or advanced head and neck squamous cell carcinoma, such as methotrexate or a combination of 5-fluorouracil and cisplatin, are far beyond satisfaction. New chemotherapeutic agents, such as taxanes, paclitaxel and docetaxel, are among the most active drugs for head and neck cancer and a number of multidrug regimens containing a taxane and cisplatin have produced equivalent or higher response rates than conventional regimens. In addition, early clinical trials of novel molecular-targeted agents, such as epidermal growth factor receptors, tyrosine kinase inhibitors and gene targeted therapy, have shown encouraging results. Further clinical trials will be needed to optimally combine the biologic agents with chemotherapy and assess their effects on long-term control of cancer. PMID- 12113142 TI - Hospital restructuring: identifying the impact on patients and nurses. AB - Health systems throughout the democratic world have been subject to 'reform' in recent years as countries have attempted to contain the rapidly rising costs of health care. Because hospital care accounts for a large proportion of health sector spending, hospital restructuring has been an important part of those changes. In an attempt to make hospitals more efficient and cost-effective, New Zealand, like other countries, has introduced extensive changes to the way in which treatment and care are delivered to patients, and to the way nurses' work is organised and managed. International research has identified links between the way in which nursing is organised in a hospital, and that hospital's patient outcomes. The current authors are part of a team of researchers undertaking research which uses the methodology of the International Hospital Outcomes Study to examine nurse staffing and patient outcomes in New Zealand's secondary and tertiary hospitals across the period 1988-2001. The research involves a large survey of all nurses working in the study hospitals, an examination of the way in which the hospitals have been restructured, and an analysis of patient outcomes. PMID- 12113141 TI - Clinical practice/education exchange: bridging the theory-practice gap. AB - Nursing education is directed toward development of nursing practitioners competent to nurse effectively in the reality of our present society. A major challenge to the nursing profession is to find ways of merging theory and practice in the delivery of nursing education and patient care. One option for achieving this goal is for nurse educators to spend time in clinical practice updating their clinical skills and re-experiencing the realities of practice. Joint appointments with practice, intermittent periods of clinical update in practice and work role exchanges have all been utilized by the profession. However clinical practice/education exchange (CPEE) involving two people--one in clinical practice and the other in education--who exchange jobs for a fixed period of time is a relatively new concept. Central to a CPEE is the aim of enhancing student learning and facilitating meaningful links between theory and practice for them. Hence the exchange occurs between the education institute and the service area where students are placed. This article positions the CPEE within nursing literature and presents narrative accounts from a nurse educator and clinician who exchanged jobs for one year. PMID- 12113144 TI - Randomised controlled trials in nursing and midwifery: an interview with Maralyn Foureur. Interview by Pamela J. Wood. AB - Randomised controlled trials are considered to be one of the best research designs for determining effective care in the clinical setting. Relatively few randomised controlled trials, however, have been carried out in nursing or midwifery practice, so few examples of the practical realities of this research methodology are readily accessible. This is the sixth article in a series based on interviews with nursing and midwifery researchers, designed to offer the beginning researcher a first-hand account of the experience of using particular methodologies. This article focuses on the randomised controlled trial as experienced by Maralyn Foureur (RGON, RM, BA, Grad Dip Clin Epidem, PhD) who used this methodology to demonstrate the effectiveness of a continuity of care model in midwifery practice. PMID- 12113143 TI - Risk assessment of violence to others: time for action. AB - Possible expansion in the scope of practice of mental health nurses, together with the prevalence of nurses being assaulted by patients, accentuates the need for nurses to be more skilled in risk assessment. A literature search was undertaken on the topics of risk assessment, dangerousness, aggression, and violence in the data bases of CINAHL, MEDLINE and PSYCHLIT, in order to determine an evidence based approach to risk assessment of patient violence towards others. In the absence of reliable and valid nursing risk assessment measures, the approach suggested here focuses on the use of observation skills to detect behaviour antecedent to physical assault, and the ability to systematically assess evidence-based risk factors of violent action. Given this rudimentary framework, there is the need to adapt it to specific clinical settings. Failure to proceed rapidly with such developments may jeopardize the safety of both patients and staff. PMID- 12113145 TI - Transcultural concepts in nursing care. PMID- 12113146 TI - The Process of Cultural Competence in the Delivery of Healthcare Services: a model of care. AB - Several models of service care delivery have emerged to meet the challenges of providing health care to our growing multi-ethnic world. This article will present Campinha-Bacote's model of cultural competence in health care delivery: The Process of Cultural Competence in the Delivery of Healthcare Services. This model views cultural competence as the ongoing process in which the health care provider continuously strives to achieve the ability to effectively work within the cultural context of the client (individual, family, community). This ongoing process involves the integration of cultural awareness, cultural knowledge, cultural skill, cultural encounters, and cultural desire. PMID- 12113148 TI - Culture care theory: a major contribution to advance transcultural nursing knowledge and practices. AB - This article is focused on the major features of the Culture Care Diversity and Universality theory as a central contributing theory to advance transcultural nursing knowledge and to use the findings in teaching, research, practice, and consultation. It remains one of the oldest, most holistic, and most comprehensive theories to generate knowledge of diverse and similar cultures worldwide. The theory has been a powerful means to discover largely unknown knowledge in nursing and the health fields. It provides a new mode to assure culturally competent, safe, and congruent transcultural nursing care. The purpose, goal, assumptive premises, ethnonursing research method, criteria, and some findings are highlighted. PMID- 12113147 TI - The Giger and Davidhizar Transcultural Assessment Model. AB - The Giger and Davidhizar Transcultural Assessment Model was developed in 1988 in response to the need for nursing students in an undergraduate program to assess and provide care for patients that were culturally diverse. The model includes six cultural phenomena: communication, time, space, social organization, environmental control, and biological variations. These provide a framework for patient assessment and from which culturally sensitive care can be designed. PMID- 12113149 TI - The Purnell Model for Cultural Competence. AB - This article provides an overview of the Purnell Model for Cultural Competence and the assumptions on which the model is based. The 12 domains comprising the organizing framework are briefly described along with the primary and secondary characteristics of culture, which determine variations in values, beliefs, and practices of an individual's cultural heritage. All health care providers in any practice setting can use the model, which makes it especially desirable in today's team-oriented health care environment. The model has been used by nurses, physicians, and physical and occupational therapists in practice, education, administration, and research in Australia, Belgium, Canada, Central America, Great Britain, Korea, South America, and Sweden. The model has also been translated into Flemish, French, Korean, and Spanish. Although the model is only 4 years old, it shows promise for becoming a major contribution to transcultural nursing and health care. PMID- 12113150 TI - Cultural Diversity in Health and Illness. AB - The purpose of this article is to describe the theoretical models that underlie the book Cultural Diversity in Health and Illness. The book's internal structure; functional structure; conceptual relationships; scope; knowledge antecedents; applications to theory, research, and practice; and areas for further development are discussed. PMID- 12113151 TI - Caregiving between two cultures: an integrative experience. AB - In 1995, more than half of elderly immigrants to the United States were born in Asia or the Pacific region. The purpose of this study was to describe the process of caring for elderly parents by Asian American women. Forty-one women (22 Chinese American and 19 Filipino American) caring for elderly parents were interviewed in a study based on Strauss and Corbin's grounded theory methodology (1990). Although the women were moderately acculturated, indications of being in transition were evident. Analysis of interview data led to development of a substantive theory of caregiving between two cultures, reflecting the paradox of living and caregiving by two sets of standards and worldviews. The primary strategies used to manage the caregiving challenges were connecting and calibrating. Through personal growth and finding meaning, the caregivers integrated the caregiver role into their lives and became more connected with their families and within themselves. PMID- 12113152 TI - Health perceptions of Mexican American women. AB - This article describes the health perceptions of a sample of moderately to highly acculturated Mexican American women. Using an ethnographic design, the author interviewed 13 women to determine their health perceptions. The interviews were guided by the domains of health described in the World Health Organization (WHO) definition of health. Three broad categories of health perceptions were identified: the physical body, the emotional component, and finding balance. With the addition of a spiritual component, the WHO definition was a useful tool for uncovering health perceptions. The process of in-depth ethnographic interviewing provided a contextual view of health in which the complexity of intrafamilial relationships was revealed, as were the importance of spirituality as a coping mechanism and the perception of health as an integrated, holistic experience. PMID- 12113153 TI - A cross-cultural comparison of family resiliency in hemodialysis patients. AB - The purpose of this descriptive comparative survey was to examine differences in family resiliency between hemodialysis patients and caregivers across three ethnically diverse samples that included Anglo-Americans, Mexican Americans, and South Koreans. The study was based on the Family Resiliency Model developed by McCubbin, Thompson, and McCubbin. The patients were in end-stage renal disease and were receiving hemodialysis treatments at either a freestanding hemodialysis unit or a 30-bed dialysis hospital unit. South Korean patients on hemodialysis and their caregivers perceived the stressors imposed by their illness to be significantly greater than either of the other ethnic groups, and they also scored lower on the resiliency measure than the other two groups. Findings of the study support the need to understand the dynamics of family resiliency from a cultural perspective. PMID- 12113154 TI - Context of care for Jordanian women. AB - The purpose of this research study was to document, describe, and analyze diverse and universal care patterns for Jordanian women. The authors used a qualitative design of observation-participation and interview data collected from 15 women in two cities and three villages in Jordan over a 4-month period. Methods used incorporated Leininger's theory of culture care diversity and universality as well as the ethnonursing research method. Themes discovered from the data include (a) culture of caring connectedness, (b) caring for family honor: the agony and the ecstasy, (c) Islam as feminist thought, (d) political care as intertwined with tribal and religious influences, and (e) reviving Rufaida: return to community care. Findings are being used for collaborative curriculum design for new community health nursing roles focusing on women's health. PMID- 12113155 TI - Cultural diversity: students' perspectives. AB - This article is the result of the interest of several faculty members in obtaining information from students about their experiences with cultural diversity. Faculty identified a need for baseline information from the student population to assess the impact of diversity programs on campus. A small, volunteer group of 7 nursing students met for one focus group to share their experiences. The discussion supported the need for ongoing faculty-student interactions to promote cultural sensitivity and knowledge development about diversity within this university population. PMID- 12113156 TI - Building transcultural knowledge through intercollegiate collaboration. AB - Although the importance of providing culturally congruent care is widely recognized among nurses, many nurses lack the skill and knowledge necessary to successfully provide culturally competent care. Nursing programs need to better prepare students in transcultural nursing. This article describes an innovative collaborative learning activity that was implemented between two different nursing programs in different states aimed at improving students' transcultural knowledge. Students worked in small groups and used the Internet to collaborate on case studies that focused on transcultural health care issues. Responses collected by anonymous evaluations at the end of three semesters identified both strengths and weaknesses of the project. Results and suggestions for improvement are reported. PMID- 12113157 TI - Providing nursing care for a Chagga client of Tanzania. AB - A case study is used to present features of a cultural assessment of the Chagga tribe. The purpose is to identify guidelines for provision of culturally congruent nursing care. "Mrs. Chuwa" lives on the foothills of Kilimanjaro. Daily survival is a major concern; the extended family is an important source of support. Traditional beliefs have a major impact on her life; traditional practitioners are alternative resources to dispensaries and clinics. Nurses providing care for Chaggas need to be aware of how important it is to communicate with respect, to obtain an accurate treatment history, and to ensure social and spiritual care is provided. Nurses need to be proactive in issues such as improving health for the community, addressing poverty, and professional development. PMID- 12113158 TI - Everything works. PMID- 12113159 TI - Murderous parents. AB - This article offers observations regarding some of the major manifestations of family violence, neonaticide, infanticide, and filicide with the purpose of aiding in the early identification of parents at risk. They are discussed within the past and present historical and cultural milieu. A brief review of pertinent literature is presented. Pertinent case studies from the forensic psychiatric practice of the author along with psychodynamic reflections are offered. PMID- 12113160 TI - Psychological profiling of offender characteristics from crime behaviors in serial rape offences. AB - Criminal psychological profiling has progressively been incorporated into police procedures despite a dearth of empirical research. Indeed, in the study of serial violent crimes for the purpose of psychological profiling, very few original, quantitative, academically reviewed studies actually exist. This article reports on the analysis of 62 incidents of serial sexual assault. The statistical procedure of multidimensional scaling was employed in the analysis of this data, which in turn produced a five-cluster model of serial rapist behavior. First, a central cluster of behaviors were identified that represent common behaviors to all patterns of serial rape. Second, four distinct outlying patterns were identified as demonstrating distinct offence styles, these being assigned the following descriptive labels brutality, intercourse, chaotic, and ritual. Furthermore, analysis of these patterns also identified distinct offender characteristics that allow for the use of empirically robust offender profiles in future serial rape investigations. PMID- 12113161 TI - Criminality and moral dysfunctions: neurological, biochemical, and genetic dimensions. AB - In this article, the author attempts to demonstrate a relationship between neurobiological dysfunctions and/or genetically determined deviant behavior and personality traits as well as moral abnormalities. Data from neuroscience show that a number of neurological dysfunctions are linked to cognitive and emotional disturbances. Cognitive and emotional abnormalities, in turn, are frequently related to moral dysfunctions. Moreover, neurological disorders can produce dramatic psychological and social problems, personality changes, and behavioral problems in patients. Those mental, emotional, and psychosocial problems and related moral dysfunctions are frequently linked to violence and/or criminal behavior. Genetic research found evidence of inheritability of antisocial traits, which interfere with moral development and activities. This information has consequences for any assessment and disposition within the legal system. More research on the interrelationship between neuro(bio)logical, genetic, emotional, and mental aspects of moral dysfunctions is needed for the development of adequate treatment, prevention, and intervention programs. PMID- 12113162 TI - Causal factors of corporate crime in Taiwan: qualitative and quantitative findings. AB - Street crimes are a primary concern of most criminologists in Taiwan. In recent years, however, crimes committed by corporations have increased greatly in this country. Employing the empirical approach to collect data about causal factors of corporate crime, the research presented in this article is the first systematic empirical study concerning corporate crime in Taiwan. The research sample was selected from a corporation with a criminal record of pollution caused by the release of toxic chemicals into the environment and a corporation with no criminal record. Questionnaire survey and interviews of corporate employees and managers were conducted, and secondary data were collected from official agencies. This research indicated the causal factors of corporate crime as follows: the failure of government regulation, lack of corporate self-regulation, lack of public concern about corporate crime, corporate mechanistic structure, and the low self-control tendency of corporate managers. PMID- 12113163 TI - Violations of out-group and in-group regulations in the eyes of ordinary and protected prisoners: an instance of judgmental modularity. AB - According to the functional theory of cognition, everyday life blaming for wrongdoing reflects individuals' functional morality. The judgments of normative people were found to be modular, that is, changeable as a function of the social role taken at the moment of the judgment. This article presents several empirical indications that prisoners' functional moral judgments are modular as well. These studies were conducted within the experimental framework of functional measurement, which was chosen due to its ability to bypass prisoners' suspicion and resistance to out-group investigations. The studies focus on delinquents' core moral issues such as eyewitness testimony and informing to out-group authorities. As expected, the judgments of ordinary and protected inmates were modular. It is claimed that these findings are applicable to offender rehabilitation and correction and to offender therapy. PMID- 12113164 TI - An evaluation of the effectiveness of substance abuse programming for female offenders. AB - Although a recent meta-analysis reported that substance abuse treatment was associated with moderate reductions in recidivism for female offenders, very few of the tests of treatment (k = 4) focused on adults. The purpose of this study was to contribute to this relatively sparse area of scientific inquiry by exploring the effectiveness of substance abuse programming in reducing recidivism for a sample of 98 federally sentenced female offenders in Canada. Results revealed a significant reduction in general recidivism for treated substance abusers. Moreover, the data indicated that violent reoffending was also reduced for the treated group, although the difference did not reach statistical significance. PMID- 12113165 TI - Maternal cigarette smoking during pregnancy and life-course-persistent offending. AB - Evidence exists documenting the relationship between maternal cigarette smoking and offspring criminal behavior. Although efforts to understand this relationship in a theoretical framework have only recently emerged, attempts made have been grounded in Moffitt's developmental taxonomy of antisocial behavior. Specifically, maternal cigarette smoking is generally viewed as a potential disruption in the offspring's neuropsychological development, which is subsequently associated with life-course-persistent offending. Using a birth cohort of 987 African Americans, the authors extend previous research by empirically assessing, prospectively, the link between maternal cigarette smoking and life-course-persistent offending while using different operationalizations of Moffitt's offending categorization. The authors' findings offer some support for the relationship between maternal cigarette smoking and life-course-persistent offending, which is dependent on how this concept is operationalized. PMID- 12113166 TI - Intimacy training in a forensic psychiatric setting: an experiment. AB - This article is about intimacy training in two forensic psychiatric hospitals. This training is an experiment in which patients are trained in skills relating to intimacy and sexuality through real physical contact with a therapist. It is a way of treatment in those cases in which other, usually verbal methods, have failed to accomplish (sexual) behaviour change, and it can feed or revitalise verbal therapy. The purpose of the training is (a) to make the patient aware of the feelings intimate contact with a woman provokes and (b) to increase the patient's social and sociosexual skills so that he learns how to handle his intimate and sexual wishes, needs, and limits, and those of his partner. The training aims at diminishing the risk of new offences. The experiment is intended to provide answers to questions about the effectiveness of this kind of training in relationship to this aim. PMID- 12113167 TI - Stress-induced Cushing's syndrome in fur-chewing chinchillas. AB - One of the most serious problems in the chinchilla industry is 'fur-chewing', when the chinchilla bites off areas of its own or some other animal's fur. The condition generally develops in both genders at the age of 6-8 months. In chinchilla farms in Croatia an incidence of 15-20% has been observed. A pathomorphological, microbiological and parasitological investigation was conducted on eleven 6- to 11-month-old chinchillas of both sexes with clinical symptoms of 'fur-chewing' and three chinchillas without such signs. Histopathology of the adrenal glands and of the chewed skin revealed changes typical of Cushing's syndrome in 'fur-chewed' chinchillas, such as hyperkeratinisation of the epidermis, epidermal atrophy, pronounced follicular and sebaceous gland atrophy, hyperkeratinisation of the follicles with comedo formations and the presence of calcium salts in subcutis. PMID- 12113168 TI - A possible relationship between bumblefoot responsive to potassium arsenite and micrococci in the blood of three birds of prey. AB - Pododermatitis (bumblefoot) is a major health problem of falcons world-wide because healing processes in the talons are difficult and lengthy. A peregrine (Falco peregrinus), a merlin (Falco columbarius) and a saker falcon (Falco cherrug) with bumblefoot at different stages ranging from III to V, were all found to be carriers of micrococcus-like organisms in the blood and two of them were successfully treated with 0.5% potassium arsenite in low dosage given intravenously. A number of considerations are made on the immune dysfunction aspects of bumblefoot in birds of prey and on the emerging role of arsenic-based medicaments in the treatment of animal and human immune dysfunction syndromes. PMID- 12113169 TI - Apparent cross-infection with a single strain of Malassezia pachydermatis on a pig farm. AB - Twenty-nine isolates of Malassezia pachydermatis were recovered from a single farm of 100 pigs in Croatia. In contrast, 290 farm pigs from other locations (northern parts of Croatia and Slovenia) yielded only two non-lipid dependent isolates of M. pachydermatis using the same swabbing procedure. Ten of the 29 isolates from a single farm had their identity confirmed by karyotyping, and were typed by restriction fragment length polymorphism (RFLP) analysis. All but one of these isolates sub-typed were indistinguishable, one isolate produced a slightly different RFLP profile. Control isolates recovered from dog skin gave RFLP profiles that were easily distinguished from those produced by the pig isolates. These results suggest that a single strain of M. pachydermatis had colonised this pig herd. PMID- 12113170 TI - Tumour suppressor gene p53 mutation in a case of haemangiosarcoma of a dog. AB - Haemangiosarcomas of dogs were analysed by molecular genetic techniques. Regions of the tumour suppressor gene p53, including the well-known tumour hot spots (codons 175, 245, 248, 249, 273 and 282) were screened. A 24 bp deletion was detected in exon 5 of the gene. PMID- 12113172 TI - Comparison of VHS video recording system with apple Macintosh-based image analysis and modified CODA-3 systems in equine motion analysis. AB - A VHS video--computer-based image analysis combination is described as a low sampling rate motion analysis system. Video recordings were taken indoor without any artificial illumination at 25 fps sampling rate. The horse studied was running on a high-speed treadmill and observed at 1.6, 4 and 7 m/s velocities at walk, trot and canter, respectively. Left forelimb and hindlimb were recorded separately from lateral view. For comparison, parallel CODA-3 recordings were taken at the same time from the same position. Joint angles were expressed and compared in angle-time diagrams. Sampling of both systems has been synchronised by a timer device at +/- 1/300 s error level. Results obtained with the two different recording systems were comparable in all joints measured with the exception of the fetlock. Inaccuracies in fetlock recordings are thought to be eliminated by measuring at controlled illumination. As a conclusion, the VHS Macintosh setup appears to be promising as a simplified system for gait analysis. PMID- 12113171 TI - Polymerase chain reaction amplification of 16S-23S spacer region for rapid identification of Salmonella serovars. AB - Polymerase chain reaction (PCR) was used to amplify the spacer regions between the 16S and 23S genes of rRNA genetic loci of Salmonella serovars for their rapid identification. These genetic loci revealed a significant level of polymorphism in length across the species/serovar lines. When the 16S-23S spacer region amplification products were subjected to agarose electrophoresis, the patterns observed could be used to distinguish all the serovars of Salmonella tested. Unique elements obtained in amplification products were mostly clustered at serovar level, although certain genus-specific patterns were also observed. On the basis of the results obtained, the amplification of 16S-23S ribosomal spacer region could suitably be used in a PCR-based identification method for Salmonella serovars. PMID- 12113173 TI - Ochratoxin a contamination of cereal grains and coffee in Hungary in the year 2001. AB - Ochratoxin A (OTA) is a nephrotoxic and carcinogenic mycotoxin, a secondary metabolite produced by mould fungi belonging to several Aspergillus and Penicillium species. It is formed during the storage of cereal grains and other plant-derived products. OTA ingested by humans and animals with the food or feed may exert deleterious effects on health. The purpose of this study was to investigate the ochratoxin contamination of the most important potential sources of OTA. The OTA content of cereal samples for human consumption (36 baking wheat, 16 wheat flour and 6 maize coarse meal samples) and feed grain samples (30 feeding wheat, 32 feeding maize and 20 feeding barley samples) collected in the mid-phase or at the end of the storage period and of 50 commercial coffee samples was determined. The analyses were performed by immunoaffinity column--high performance liquid chromatography (IAC-HPLC). The limit of detection of the method was 0.1 ng/g. Of the wheat samples intended for human consumption, 8.3% contained OTA at 0.29 ng/g on the average (OTA ranges: 0.12-0.5 ng/g; Table 2). The OTA contamination of wheat flour and maize meal samples for human consumption was similar to that of the baking wheat samples. OTA contamination was found in 26.7% of the feeding wheat, 15.6% of the feeding maize and 35% of the feeding barley samples. The average values and the ranges of OTA levels found in the above samples were 12.2 and 0.3-62.8 ng/g, 4.9 and 1.9-8.3 ng/g, and 72 and 0.14 212 ng/g, respectively (Table 3). Sixty-six percent of the coffee samples were contaminated with OA (average level: 0.57 ng/g, ranges: 0.17-1.3 ng/g; Table 4). OTA contamination of baking wheat samples was found to be relatively low, presumably as a result of the favourable weather at harvest and the optimal storage conditions. Calculations made on the basis of the obtained results show that the daily OTA intake of an adult human from edible cereals is only 6.7 ng, while the amount taken up with coffee is 4.1 ng daily. The high prevalence and high levels of OTA contamination in feed grains can be explained by the unfavourable storage conditions, and this finding suggests that OA-related health problems may arise in animals, and that foods of animal origin may be contaminated with this mycotoxin. PMID- 12113174 TI - Development of an antibiotic resistance monitoring system in Hungary. AB - Because of the rapid development and spread of antimicrobial resistance it is important that a system be established to monitor antimicrobial resistance in pathogenic zoonotic and commensal bacteria of animal origin. Susceptibility testing of bacteria from carcasses and different samples of animal origin has been carried out in veterinary institutes for a long time but by an inconsistent methodology. The disc diffusion method proposed by the National Committee for Clinical Laboratory Standards (NCCLS) was introduced in all institutes in 1997. In order to obtain a coherent view of the antimicrobial resistance of bacteria a computer system was consulted, consisting of a central computer to store all data and some local computers attached to it through the network. At these local measuring stations computers are connected to a video camera, which displays the picture of Petri dishes on the monitor, and inhibition zone diameters of bacteria can be drawn with the mouse by the inspector. The software measures the diameters, evaluates whether or not the bacteria are sensitive, and stores the data. The evaluation is based upon the data of the NCCLS. The central computer can be connected to as many local computers with measuring stations as we wish, so it is suitable for an integrated system for monitoring trends in antimicrobial resistance of bacteria from animals, food and humans, facilitating comparison of the occurrence of resistance for each circumstance in the chain. It depends on the examiners which antibiotics they want to examine. Thirty-two different antibiotic panels were compiled, taking into consideration the active ingredients of medicinal products permitted for veterinary use in Hungary, natural resistance and cross-resistance, the mechanism of resistance and the animal species, i.e. which drugs were recommended for treatment in the given animal species, and the recommendations of the OIE Expert Group on Antimicrobial Resistance. The members of the panels can be changed any time, even during the measuring process. In addition to the inhibition zone diameters of bacteria the database also includes information about bacterial and animal species, the age of animals and the sample or organ where the bacteria are from. Since January 2001 the antibiotic susceptibility of E. coli, Salmonella, Campylobacter and Enterococcus strains isolated from the colons of slaughter cows, pigs and broiler chickens has also been examined. Each of the 19 counties of Hungary submits to the laboratory three tied colon samples from a herd of the above-mentioned animals every month. PMID- 12113175 TI - Pulse and continuous oral norfloxacin treatment of experimentally induced Escherichia coli infection in broiler chicks and turkey poults. AB - Experimental colibacillosis was produced in 40 healthy, 7-day-old broiler chickens and turkeys by intratracheal injection of 1 x 10(8) CFU/chick and 1.23 x 10(9) CFU/poult bacteria of an O1:F11 strain of Escherichia coli, respectively. Two days before E. coli challenge all chicks were vaccinated with a live attenuated strain of infectious bronchitis virus (H-52). This model of infection- at least in chicken--proved to be useful for evaluating the efficacy of antimicrobial medication, by recording mortality, body weight gain, pathological alterations and frequency of reisolation of E. coli. Using this model, the efficacy of two different dosing methods of norfloxacin (continuous and pulse dosing) was evaluated. The once-per-day pulse dosing of norfloxacin administered via the drinking water at 15 mg/kg body weight proved to be more efficacious than the continuous dosing method of 100 mg/L for 5 days in chickens, while there were no convincing differences between the two treatment regimens in turkeys. The results confirmed earlier observations on the pharmacokinetic properties of norfloxacin in chicks and turkeys (Laczay et al., 1998). PMID- 12113176 TI - Na+/H+ exchange in primary, secondary and n-butyrate-treated cultures of ruminal epithelial cells: short communication. AB - Rate of amiloride-sensitive Na+ uptake into cultured rumen epithelial cells was studied in order to clarify the influence of culture conditions on Na+/H+ exchange (NHE). Cell cultures were exposed to Na-n-butyrate or not for seven days or subcultured. On the 14th day of culturing, primary cell cultures without butyrate exposure showed both non-stratified and stratified growth. Na-n-butyrate treated 14-day-old cultures and 3-day-old subcultures contained mostly non stratified, i.e. non-keratinised cells. Both n-butyrate treatment and subculturing increased total and amiloride-sensitive Na+ uptake. Our results indicate that Na+ uptake via NHE is determined by the amount and the ratio of non stratified (non-keratinised) cells. PMID- 12113177 TI - Effect of different fat sources on in vitro degradation of nutrients and certain blood parameters in sheep. AB - This study was designed to determine the effects of calcium salt of palm oil fatty acids (CS), hydroxyethylsoyamide (HESA), butylsoyamide (BSA) and soybean oil (SO) on degradation of crude protein and fibre in vitro, and on the blood plasma lipid parameters in vivo. Five mature wethers (body weight 75 kg) were fed five diets in a 5 x 5 Latin square experiment. The control diet consisted of 50% meadow hay and 50% concentrate with no added fat. The control diet was supplemented with CS, HESA, BSA, or SO. Fat was added at 3.5% of dietary dry matter (DM). The final ether extract content of the ration was near 6%. Each period lasted 20 days. Fat supplements, except HESA, consistently decreased the in vitro DM disappearance of soybean meal as compared to control. In contrast to the effect of other treatments, crude protein degradation was greatest in the test tubes with inocula obtained from sheep fed diet with HESA. Fat supplements equally inhibited the DM and fibre breakdown of alfalfa pellet. CS and HESA seemed to be less detrimental to in vitro fermentation of neutral detergent fibre (NDF) than BSA and SO. All fat supplements increased blood plasma triglyceride, cholesterol and total lipid content. Plasma concentration of cholesterol and total lipid was highest with SO. The inclusion of CS in the diet increased 16:0, while all fat supplements increased plasma 18:0 and decreased 16:1 and 18:1 fatty acid content. Plasma 18:2n-6 was not changed by feeding CS and SO. However, compared to the control diet, 18:2n-6 increased with 12 and 41% in plasma fatty acids when sheep were fed HESA and BSA, respectively. The results showed that plasma concentration of linoleic acid was enhanced more when the amide was synthesised from butylamine than when from ethanolamine. PMID- 12113178 TI - Improving reproductive performance in lactating dairy cows by synchronizing ovulation or inducing oestrus. AB - Lactating crossbred Holstein-Friesian dairy cows (n = 331) were started on an Ovsynch regimen 68 +/- 8.2 days after calving; 200 micrograms GnRH intramuscularly (i.m.) on Days 0 and 9, and 35 mg prostaglandin F2 alpha i.m. on Day 7. Thirty-eight and 31 cows (11.5 and 9.4%, respectively) were in oestrus on Days 0 to 6 and 7 to 8, respectively, and inseminated, and the remainder were fixed-time inseminated (on Day 10). For these three groups, pregnancy rates (60 65 days after breeding) were 31.6, 38.7 and 34.0%, respectively (P = 0.82) and calving rates were 100, 100 and 89.9% (P = 0.23). In a preliminary trial, twelve lactating cows (45 to 60 days postpartum) with inactive ovaries were given 1500 IU eCG i.m.; 10 were in oestrus within 10 days after treatment (and inseminated) and eight of these were pregnant (30 days after breeding). The Ovsynch program resulted in acceptable reproductive performance in cyclic cows and eCG treatment has considerable promise for inducing oestrus in anoestrous cows. PMID- 12113180 TI - [Diversity and selectivity in biomolecules]. AB - Nature provides molecular constructions in inexhaustible richness. She combines small building blocksacks, and from the great variety of the possible structures, the really existing molecules are selected by kinetic (actually enzymic) and thermodynamic control. The operation of this fundamental principle is demonstrated by means of the graph analysis, in the large community of the monoterpenoid indole alkaloids isolated from the species of the Apocynaceae, Loganiaceae and Rubiaceae plant families. Strictosidine, an alkaloid glucoside, is formed from a single enantiomer of secologanin and tryptamine by enzymic catalysis and with complete diastereoselectivity. It is the exclusive precursor of more than 2200 alkaloids. The configuration of one or more analogous centers of chirality of them is identical, i.e. homochiral, insofar it will be lost or changed in subsequent reactions. Strictosidine exists in a single, most stable conformer of the possible 324 ones. On simple secologanin derivatives it was proved, that the removal of the glucosyl unit by enzyme results, under kinetic control, in the formation of the epimer pair of the primary aglucone. However, proton catalysed hydrolysis gives, under thermodynamic control, the most stable epimer pair of the possible 48 aglucones formed in 368 elementary steps. In the deglucosylation of the strictosidine type compounds, in 2 series, 1 tricyclic, 8 tetracyclic and 16 pentacyclic aglucone types are formed by further oxa-, carba- and/or azacyclizations. The alkaloids derived from strictosidine can be grouped into one unrearranged (I) and two rearranged (II and III) skeletons, in normal and iso series as well as along a and b lines. The processes are directed partially by proton migrations. The really existing alkaloids are formed in subsequent chemical reactions. The rich molecular library obtained by that way reveals the fundamental unity of Nature in the material multiplicity, i.e. it lets shine up the beauty in the world of the biomolecules, too. PMID- 12113181 TI - [Controlled drug delivery systems. Paradigm change in pharmacology]. AB - This article is to introduce the readers in the colourful world of controlled drug delivery. These structures are to carry the drug into the place of release for let it there out with proper velocity. So the aim of the researches are not to find the best drug, but to get it into the right place in the right time. Some controlled drug delivery processes are already in use. However it comes everyday a physical, chemical or biological idea for making a new style of controlled drug delivery system. This is the reason way we find this part of the pharmacology worth to the attention. We like to introduce it with short presenting of drug delivery systems, they classes, they working and the physical chemistry of drug release. PMID- 12113179 TI - Comparative evaluation of the effect of antioxidants in the conservation of ram semen. AB - The aim of the present study was to develop a treatment supporting the membrane of ram spermatozoa. Semen of different ejaculates collected from breeding rams was mixed and samples of 10(9) sperm cells per ml and Tris-egg yolk extender were completed with the following antioxidants: alpha-tocopherol acetate (E), glutathione peroxidase (GP), Aromex (AR), resveratrol (R), resveratrol + vitamin E (RE), resveratrol + Aromex (RAR), resveratrol + GP (RGP). Peroxidation was evaluated by the analysis of malondialdehyde (MDA) during incubation for 30, 60 and 120 min at 37 degrees C as well as during a 24-h incubation at 5 degrees C. The success of preservation was checked in a 9-day-long period by observing the acrosomal defects and the motility of spermatozoa. Concentration of MDA was 4.06 nmol/10(9) spermatozoa in samples treated with 15 micrograms R while the control sample contained 69.79 nmol MDA per 10(9) spermatozoa after 24-h incubation. Following 30-, 60- and 120-min storage the concentration of MDA in control and R treated samples was 25.89, 36.91, 49.57 and 3.69, 3.74, 3.74 nmol/10(9) spermatozoa, respectively. Moreover, a significantly higher proportion of motile sperm cells was observed in the treated than in the control samples. The frequency of acrosomal defects was lower in the treated groups than in the control. These results indicate that RAR treatment can improve the effects of ram semen preservation. PMID- 12113182 TI - [Introduction of liposomes: examination methods and their importance in research and quality assurance]. AB - After a short introduction, the most important and most commonly used liposome examination methods are described in two parts. At first, we would like to review some of the methods used to examine physical parameters (electronmicroscope, high pressure liquid chromatography, light scattering measurement etc.). In the second part, the methods are reviewed with changes during phase transition of lipids can be described (differential scanning calorimetry, fluorescence). PMID- 12113183 TI - [Therapeutic aspects of HIV/AIDS infected patients and evaluation of therapeutic protocols]. AB - The first HIV-infected patients were treated in 1986, however, at that time medicines inhibiting HIV replication were not available. Solely the complications of AIDS and opportunistic infections could be treated. The HIV replication inhibition capacity of antiretroviral nucleoside and ribavirin were tested in Hungary in 1987, an early date even in international practice. The first nucleoside reverse transcriptase inhibitor (NRTI), azidothymidine, presently called zidovudine (ZDV), was introduced in 1989. By giving patients the appropriate dosage of this, the progression of the disease could be delayed by approximately 6 months to one year. In 1991, the application of two new NRTI was commenced, namely zalcitabine (DDC) and didanosine (DDI). These medicines were applied partly in case of ZDV intolerance and as a part of the sequential monotherapy. In 1994 and 1995 two new NRTI's were introduce, namely stavudine (d4T) and lamivudine (3 TC). At that same time, to obtain a more effective replication inhibition method, a double NRTI combination became part of the therapeutic protocol. The year 1996 resulted in significant changes. At the beginning of the year, two compounds belonging to two new therapeutic procedures. These are saquinavir (SQV) and delavirdine (DLV) belonging to the groups of protease inhibitors and non-nucleoside reverse transcriptase, respectively. The so-called virus cocktails and the effective active antiretroviral therapy (HAART) were applied and, later, to monitor the efficiency of the treatment, the opportunity was provided to measure the copy number of HIV-RNS. The treatment of the HIV disease entails a number of unanswered questions. The maximum result that can be achieved by using today's therapeutic methods is to prolong that particular phase of the HIV disease, which secures the patient a fairly good quality of life. The new combination of the antiretroviral compounds produced by the pharmaceutical industry provides better changes to prolong said period by years, sometimes decades. However, the real solutions to the problem are only theoretically known treatment procedures (gene therapy, cytokins) today. PMID- 12113184 TI - [Formulation of tablets decreasing plasma homocysteine levels]. AB - Clinical studies confirm that intake of folate, vitamin B6 and B12 above the current recommended dietary allowance is of great significance in the primary prevention of coronary heart disease. In order to fulfill the ever increasing requirements concerning the external appearance, reproducible production of medicines and to decrease the time necessary for the preformulation and formulation experiments, I selected methods which enabled rational experimental design and fast objective evaluation. The objective of my thesis was to formulate tablets containing folic acid, vitamin B6 and B12 of optimal therapeutic concentration and of required stability in the presence of novel excipients applied in direct compression. In the course of my preformulation work thermoanalytical (DTG, DSC), chromatographic (HPLC) and spectrometric (NIR) methods were applied. Destabilizing interactions were evaluated with directly compressible excipients, like Ac-Di-Sol, Cellactose, Tablettose, Avicel PH101, Ludipress. Interactions were detected in the 1 + 1 physical mixtures of vitamins and excipients containing lactose (Cellactose, Ludipress, Tablettose) and their destabilizing effect was confirmed. The results of preformulation studies enabled detection of drug-excipients interactions and selection of compatible excipient system to the given drug composition to formulate dosage form of required stability, processability and effectiveness. PMID- 12113186 TI - [University of Florida and Center for Drug Discovery through a visiting scholar's view]. AB - The article introduces University of Florida and the Center for Drug Discovery based on the author's experience as a visiting research fellow. PMID- 12113185 TI - [Synthesis of estrone derivatives containing different halogens and/or heterocyclic moieties]. AB - New compounds derived from the 3-methyl and 3-benzyl ethers of a D-secoestrone aldehyde were synthesized. In the presence of different Lewis acids D homosteroids could form. These reactions which can be explained by an intramolecular Prinstype mechanism follow high stereoselectivity and reactivity and lead to compounds containing 16 beta-oriented halogens in the sterane skeleton. The formation and reaction of the imines derived from the aldehyde and aniline as well as substituted anilines provide a highly efficient access to steroid derivatives. The steroid alkaloids are of pharmacological interest. PMID- 12113187 TI - Breast self-examination. Is it really so dangerous? PMID- 12113188 TI - Reflections on walk-in clinics. PMID- 12113189 TI - Residents as teachers: time for formal training. PMID- 12113190 TI - Taking ACE inhibitors during pregnancy. Is it safe? AB - QUESTION: A pregnant patient is taking enalapril for primary hypertension. How safe are angiotension-converting enzyme inhibitors (ACEI) during pregnancy? ANSWER: Evidence of whether ACEIs cause problems during the first trimester of pregnancy is reassuring. There is evidence that they cause severe renal and other problems during the second and third trimesters, however. These drugs should be avoided during pregnancy. PMID- 12113191 TI - Dermacase. Lip licker's dermatitis. PMID- 12113192 TI - Practice tips. Fine needle aspiration of breast lumps. PMID- 12113193 TI - Radiation treatment for breast cancer. Recent advances. AB - OBJECTIVE: To review recent advances in radiation therapy in treatment of breast cancer. QUALITY OF EVIDENCE: MEDLINE and CANCERLIT were searched using the MeSH words breast cancer, ductal carcinoma in situ, sentinel lymph node biopsy, and postmastectomy radiation. Randomized studies have shown the efficacy of radiation treatment for ductal carcinoma in situ (DCIS) and for invasive breast cancer. MAIN MESSAGE: Lumpectomy followed by radiation is effective treatment for DCIS. In early breast cancer, shorter radiation schedules are as efficacious for local control and short-term cosmetic results as traditional fractionation regimens. Sentinel lymph node biopsy is done in specialized cancer centres; regional radiation is recommended for patients with four or more positive axillary lymph nodes. Postmastectomy radiation has been shown to have survival benefits for high risk premenopausal patients. Systemic metastases from breast cancer usually respond satisfactorily to radiation. CONCLUSION: Radiation therapy continues to have an important role in treatment of breast cancer. There have been great advances in radiation therapy in the last decade, but they have raised controversy. Further studies are needed to address the controversies. PMID- 12113196 TI - Update on Contraception Awareness Project. PMID- 12113194 TI - How, what, and why of sleep apnea. Perspectives for primary care physicians. AB - OBJECTIVE: To review the need for primary care physicians to screen for patients with obstructive sleep apnea (OSA). QUALITY OF EVIDENCE: Literature was reviewed via MEDLINE from 1993 to 2000, inclusive, using the search term "sleep apnea" combined with "epidemiology," "outcome," and "diagnosis and treatment." Citations in this review favour more recent, well controlled and randomized studies, but findings of pilot studies are included where other research is unavailable. MAIN MESSAGE: Obstructive sleep apnea is a disorder with serious medical, socioeconomic, and psychological morbidity, yet most patients with OSA remain undetected. Primary care physicians have a vital role in screening for these patients because diagnosis can be made only through overnight (polysomnographic) studies at sleep clinics. Physicians should consider symptoms of excessive or loud snoring, complaints of daytime sleepiness or fatigue, complaints of unrefreshing sleep, and an excess of weight or body fat distribution in the neck or upper chest area as possible indications of untreated OSA. CONCLUSION: Current research findings indicate that treating OSA patients substantially lowers morbidity and mortality rates and reduces health care costs. Primary care physicians need more information about screening for patients with OSA to ensure proper diagnosis and treatment of those with the condition. PMID- 12113195 TI - Breast cancer screening. First Nations communities in New Brunswick. AB - OBJECTIVE: To determine use of breast cancer screening and barriers to screening among women in First Nations communities (FNCs). DESIGN: Structured, administered survey. SETTING: Five FNCs in New Brunswick. PARTICIPANTS: One hundred thirty three (96%) of 138 eligible women between the ages of 50 and 69 years. INTERVENTIONS: After project objectives, methods, and expected outcomes were discussed with community health representatives, we administered a 32-item questionnaire on many aspects of breast cancer screening. MAIN OUTCOME MEASURES: Rate of use of mammography and other breast cancer screening methods, and barriers to screening. RESULTS: Some 65% of participants had had mammography screening within the previous 2 years. Having mammography at recommended intervals and clinical breast examinations (CBEs) yearly were significantly associated with having had a physician recommend the procedures (P < .001). A family history of breast cancer increased the odds of having a mammogram 2.6-fold (P < .05, 95% confidence interval [CI] 1.03 to 6.54). Rates of screening differed sharply by whether a family physician was physically practising in the community or not (P < .05, odds ratio 2.68, 95% CI 1.14 to 6.29). CONCLUSION: Women in FNCs in one health region in New Brunswick have mammography with the same frequency as off-reserve women. A family physician practising part time in the FNCs was instrumental in encouraging women to participate in breast cancer screening. PMID- 12113197 TI - Strength of teamwork. PMID- 12113198 TI - Hypothesis: the research page. The Cochrane Collaboration: for family physician researchers. PMID- 12113199 TI - Driving-related fear: a review. AB - This article reviews the research on driving-related fear (DRF). Until recently, research has concentrated almost exclusively on the effect of motor vehicle accidents (MVAs) on subsequent levels of DRF. However, recent findings have suggested that MVAs are not solely responsible for this fear reaction, and that non-MVA driving fear can be just as strong. Studies of the broader driving fearful population have encountered difficulty with diagnostic conceptualisation of DRF, although some have investigated a possible typology of DRF. Driving skill has been a neglected issue in the DRF research, and may prove to be a useful part of assessment and remediation of this potentially debilitating problem. Issues of definitional inconsistency are highlighted, and suggestions are made for several directions that future research might profitably take. PMID- 12113200 TI - Anorexia nervosa: obsessive-compulsive disorder, obsessive-compulsive personality disorder, or neither? AB - Anorexia nervosa (AN) is a severe and often chronic disorder with uncertain aetiology and poor prognosis. New approaches to the understanding of the disorder are needed in order to aid the development of more effective treatments. Several authors have suggested that AN has a considerable overlap with obsessive compulsive disorder (OCD) and that this may reflect common neurobiological, genetic, or psychological elements. However, more recent studies have suggested that AN may have a closer relationship with obsessive-compulsive personality traits such as those found in obsessive-compulsive personality disorder (OCPD). In this paper, evidence for links between the three conditions is reviewed, suggestions for further research are outlined and possible implications for the treatment of AN are presented. PMID- 12113201 TI - Distinctive activities of cognitive-behavioral therapy. A review of the comparative psychotherapy process literature. AB - The present review examined the comparative psychotherapy process literature in order to identify the distinctive activities of cognitive-behavioral (CB) treatment. Six techniques and interventions were found to distinguish CB from psychodynamic-interpersonal (PI) therapy: (1) use of homework and outside-of session activities; (2) direction of session activity; (3) teaching of skills used by patients to cope with symptoms; (4) emphasis on patients' future experiences; (5) providing patients with information about their treatment, disorder, or symptoms; and (6) an intrapersonal/cognitive (C) focus. Identifying the distinctive features of CB therapy can improve the measurement of process outcome correlations by more accurately specifying and operationalizing the treatment-specific processes of CB treatment, help researchers differentiate between common and treatment-specific factors, and aid in development of more psychometrically sound instruments assessing adherence and competence in CB therapy. In addition, this review can improve training of CB therapists by providing a guide for clinical practice. PMID- 12113202 TI - Treating antisocial behavior: a context for substance abuse prevention. AB - A large body of literature illustrates an association between antisocial behavior and substance abuse. In the present paper, this association is reviewed from a behavioral-economic standpoint. It is suggested that childhood antisocial behavior is a behavioral complement of substance abuse, and that prosocial behavior is a substitute for substance abuse. Based on this formulation, efforts to reduce or prevent antisocial behavior may be conceptualized as prevention programs for substance abuse. Four empirically supported approaches for the treatment of antisocial behavior are reviewed with respect to their promise for preventing and treating substance abuse. Taken together, they suggest that successful interventions for substance abuse may occur at various points over the course of development, beginning in infancy and extending into adolescence. PMID- 12113203 TI - Psychopathy in juvenile offenders. Can temperament and attachment be considered as robust developmental precursors? AB - Attempts to predict adult psychopathy generally focus on aggressive and antisocial behavior exhibited in childhood and adolescence. Yet, children with conduct problems constitute a heterogeneous group, and many of the unique interpersonal and affective features associated with the construct of psychopathy only apply to a small subset of children displaying antisocial behavior. The current review seeks to derive an understanding of the specific precursors of the apparently amoral, affectionless, and self-centered orientation that psychopathic youngsters display towards other people. The focus is on the notions of temperament and attachment in early childhood, and their links to the emergence of moral emotions later in life. Based on a developmental perspective, the data currently available are examined, highlighting the insights gained from this body of work and outlining the conceptual and methodological challenges that still need to be addressed. PMID- 12113205 TI - Why there is a need for the personality assessment of offenders. PMID- 12113204 TI - Painful procedures in pediatric cancer. A comparison of interventions. AB - Different interventions (i.e., cognitive-behavioral, pharmacological) and their combination were examined and compared to assist pediatric patients with cancer to manage distress during painful procedures. Findings revealed that cognitive protocols are effective in relieving procedural distress for a significant number of children. Pharmacological therapies were found to be relatively safe and effective when carefully administered and monitored by medical personnel. Data from combined cognitive therapies and pharmacological interventions, particularly those more recent pharmacological interventions, reveal generally mixed results, with both types of interventions yielding distinct benefits and disadvantages. Recommendations are made for future studies that match interventions to specific characteristics of the children for whom they are intended, as well as additional studies that combine pharmacological approaches together with cognitive techniques. PMID- 12113206 TI - Introduction to a special series: psychological testing in forensic settings. PMID- 12113207 TI - Treatment planning with the Psychopathy Checklist-Revised (PCL-R). AB - The Psychopathy Checklist-Revised (PCL-R) is an essential component of any assessment protocol within forensic and correctional settings. Both nomothetic and idiographic interpretations aid the clinician in screening and treatment planning. Whereas the PCL-R can be effective in identifying those who are at highest risk of disrupting treatment efforts and jeopardizing the safety of those around them, through item analysis, it also provides clues to the nonpsychopath's unique treatment needs. Specific recommendations are offered regarding the implementation of the PCL-R for screening and treatment planning, and illustrative case examples are provided to enliven essential points. PMID- 12113208 TI - Use of the Rorschach in forensic settings for treatment planning and monitoring. AB - Forensic psychiatric patients exhibit complex clinical issues that are neither readily understood by staff nor necessarily responsive to traditional psychotherapy or treatment milieu approaches. Individualized treatment planning identifies treatment needs and matches them to treatment services, thereby increasing the opportunity for a positive therapeutic outcome. The nature of the Rorschach, particularly that it bypasses volitional resources, enables observation and quantification of personality processes, making the Rorschach uniquely suited for treatment planning in forensic settings. In this article, the authors review relevant Rorschach literature, address the importance of incorporating Rorschach data into the assessment process, and discuss how Rorschach data fit into a thorough assessment that includes historical, clinical, dispositional, and contextual information. The authors offer two case examples to illustrate how Rorschach data are integrated in forensic treatment planning. PMID- 12113209 TI - Use of the MMPI-2 in the treatment of offenders. AB - The MMPI-2 is among the most frequently relied upon inventories for personality assessment. The test is utilized across a variety of nonforensic settings (e.g., psychiatric inpatient and outpatient) as an aid in formulating treatment plans, assessing treatment progress, and measuring treatment outcome. The MMPI-2 can also be utilized in forensic settings in a manner similar to its use in other settings, such as identifying treatment goals and evaluating treatment efficacy. Various MMPI-2 scales can identify an individual's treatment needs, reveal potential obstacles to treatment, and serve as a measure of treatment efficacy. Such information can be very helpful to the clinician in formulating or modifying a course of treatment for offenders. This article provides an overview of the use of the MMPI-2 in treatment planning and describes the relation between scores on the validity, clinical, and various supplementary scales and treatment-related issues. PMID- 12113210 TI - The use of the MCMI-III in the screening and triage of offenders. AB - The Millon Clinical Multiaxial Inventory-III (MCMI-III) is well suited for use in corrections settings. The MCMI-III's scales correlate with the Diagnostic and Statistical Manual of Mental Disorders-Fourth Edition (DSM-IV) personality disorders (frequently found in correctional settings), and the publisher offers a corrections-specific interpretive package. To further elucidate the usefulness of the MCMI-III with offenders and assess its efficacy, the authors administered the test to more than 10,000 inmates of the Colorado Department of Corrections. Scale scores were compared to intake judgment and outcome variables across mental health, substance abuse, and violence variables. A number of scales were found to predict several mental health variables such as subsequent diagnosis, medication prescription, and therapy time. Substance abuse scale elevations corresponded to subsequent intake recommendations. Although more modest, the aggressive personality disorder scales and several of the neurotic scales correlated with future institutional violence. The authors discuss the relevance of these findings to screening, triage, and correctional assessment. PMID- 12113211 TI - The use of the Personality Assessment Inventory (PAI) in assessing offenders. AB - The intersection between the legal and mental health professions is sometimes marked by controversy, and the application of psychological assessments to forensic issues is no exception. However, the field of psychological assessment holds particular promise for clarifying decision making within the forensic arena, as it can bring a particularly well-established body of theory and data to bear upon clinical forensic practice. This article describes one psychometric instrument, the Personality Assessment Inventory, as an example of how particular assessment instruments can help inform this process. PMID- 12113212 TI - Psychological assessment of forensic psychiatric outpatients. AB - The 1960s decrease in long-term residential mental health care resulted in former psychiatric patients being admitted to correctional and forensic psychiatric facilities. Although psychologists face challenges in managing and treating this displaced population, assessment data plays a pivotal role in the determination of appropriate aftercare for the mentally ill parolee. This article discusses the assessment protocol utilized by the Forensic Conditional Release Program (CONREP) in California, summarizes data from these patients, and uses case excerpts to illustrate the potential value of assessment with a forensic psychiatric (outpatient) population. Special emphasis is given to the use of the MMPI-2 and Rorschach. PMID- 12113213 TI - The utility of the State-Trait Anger Expression Inventory with offenders. AB - Self-report instruments can provide useful information as part of a thorough clinical assessment. However, their use in forensic settings can be problematic. The State-Trait Anger Expression Inventory (STAXI) has recently been proposed as an effective instrument for screening and outcome measurement in anger management programs. This study evaluated the effectiveness of this instrument in a sample of both voluntary and court-ordered anger-management clients, all of whom were determined through diagnostic interviews to have significant anger problems. Contrary to findings in nonforensic samples, the STAXI Trait Anger scale identified only about half of the participants as having anger-management problems severe enough to require intervention. Supplemental analysis with two additional scales did not significantly improve sensitivity. In addition to thorough diagnostic interviewing, forensic use of the STAXI (like similar assessment methods) may require additional validity scales to detect denial or socially desirable response patterns. PMID- 12113214 TI - Histamine in health and disease. AB - Histamine is a potent vasoactive agent, bronchial smooth muscle constrictor, and stimulant of nociceptive itch nerves. Activation of H1-receptors plays a central role in the immediate allergic reaction, but has less of an impact in chronic allergic disorders where inflammatory infiltrates, additional mediators such as LTC4/D4/E4 and cytokines, and structural remodeling occur. Histamine, through its H1-receptor-mediated activities, appears to be primarily a proinflammatory agent, yet it does have some homeostatic functions in gastric acid production (H2 receptors) and the central nervous system (predominantly H3-receptors) (97, 98). The realization that first-generation antihistamines often had mixed pharmacological properties (e.g., anticholinergic actions) and crossed the blood brain barrier led to the development of the second-generation drugs, which are more selective for H1-receptors, have less access to the central nervous system, and, therefore, a more favorable benefit-to-risk ratio (therapeutic index). The potential for combined H1-H3-antagonists remains to be fully explored, but offers another exciting opportunity for this ever-expanding family of beneficial drugs. PMID- 12113215 TI - Antiallergic anti-inflammatory effects of H1-antihistamines in humans. AB - Data from in vitro, in vivo, and ex vivo studies suggest that second-generation antihistamines have a number of antiallergic, anti-inflammatory properties that appear to be independent of their H1-blockade activity. First-generation antihistamines also have antiallergic, anti-inflammatory properties, as suggested by the studies with azatadine, chlorpheniramine, mepyramine, and promethazine; most other first-generation antihistamines have not been studied for these properties. In vitro studies have shown that H1-antihistamines reduce the release of proinflammatory mediators from mast cells and basophils, the chemotaxis and activation of inflammatory cells (especially eosinophils), and the expression of adhesion molecules induced by immunological and nonimmunological stimuli in epithelial cell lines. Nasal allergen challenge models have similarly demonstrated that H1-antihistamines inhibit mediator release from mast cells and basophils, and that they decrease inflammatory cell infiltration and the expression of adhesion molecules on epithelial cells. The results of published studies of the effects of H1-antihistamines on nasal allergic inflammation in humans have been summarized in this chapter. Recent investigations indicate that H1-antihistamines may modulate airway inflammation by downregulating the activity of airway epithelial cells, which have an important role in allergic airway inflammation. The modulation of adhesion molecules and of inflammatory cell infiltration by H1-antihistamines may be beneficial during long-term treatment in patients with allergic rhinitis. The rationale for this hypothesis is the persistence of inflammation on the nasal epithelial cells even when patients are symptom-free (16). All of the events affected by H1-antihistamines are important in the allergic inflammation cascade. The underlying mechanisms for such effects remain unclear, but are unrelated to H1-antagonist activity. Several studies have demonstrated that H1-antihistamines can form an ionic association with cell membranes and inhibit calcium ion influx into the mast cell or basophil plasma membrane, or inhibit Ca2+ release within the cells, and may therefore influence the signal transduction pathways. However, these effects appear to occur at concentrations higher than those achieved in therapeutic practice (126-128). It has recently been hypothesized that the anti-inflammatory activity of H1 antihistamines may be a consequence of their ability to influence the activation of genes responsible for the expression and synthesis of proinflammatory mediators (129). The contribution of the antiallergic effects of H1-receptor antagonists to their clinical efficacy is not fully understood. There have been no data suggesting that H1-antihistamines with well-documented antiallergic properties are superior to the others for which such properties have not been as extensively investigated. Additional studies are needed to elucidate the mechanisms(s) by which H1-antihistamines exert anti-inflammatory effects. This knowledge might lead to the development of novel therapies with more potent and specific anti-inflammatory effects. PMID- 12113216 TI - Clinical pharmacology of H1-antihistamines. PMID- 12113217 TI - Antihistamines in rhinoconjunctivitis. AB - In allergic rhinoconjunctivitis, histamine is known to contribute predominantly to nasal itch, sneeze, rhinorrhea, conjunctival itch, and lacrimation and these symptoms benefit most from H1-antihistamine therapy. The discovery in the early 1980s of nonsedating H1-receptor antagonists contributed dramatically to the more widespread acceptance of this mode of therapy. This also led to the undertaking of well-designed clinical trials that have added significantly to our understanding of allergic rhinitis. Oral treatment modifies both nasal and ocular symptoms and provides effective control throughout a 24-h period with once- or twice-daily medication. The advent of topical H1-receptor antagonists offers a wider choice of treatments and provides equal or greater efficacy with lower systemic bioavailability. While having a major impact on rhinoconjunctivitis symptoms, H1-antihistamines do not fully modify disease since histamine is not the only contributor to symptom generation in allergic rhinoconjunctivitis. While the search for oral H1-antihistamines with more widespread "antiallergic" activity continues, the currently available medications modify predominantly histamine-regulated events despite in vitro evidence of greater potential. The development of these new medications may be the next significant advance in this mode of treatment. PMID- 12113218 TI - H1-antihistamines in asthma. AB - Histamine released from mast cells and basophils is an important mediator of airway inflammation in asthma, particularly in the development of the early allergic response. Although histamine has been shown to contribute significantly to the bronchoconstrictor response to allergen or exercise, leukotrienes are likely to play a more prominent role in these responses in asthma. The improved specificity, tolerability, and safety profile of the second-generation H1 antagonists associated with anti-inflammatory activities and bronchodilator activities, may contribute to relieve the symptoms of the upper and lower airways in patients with coexistent mild seasonal asthma and allergic rhinitis. Considering the global rise in the prevalence of allergy and asthma, the suggestion that H1-antagonists may delay the onset of asthma in infants is of considerable interest and merits further assessment. Although it is unlikely that monotherapy with most currently available H1-antagonists will provide significant clinical benefit in asthma, the potential of combined antihistamine and antileukotriene therapy may prove useful, particularly in subjects with poor compliance to inhaled corticosteroid therapy. PMID- 12113219 TI - Antihistamines in urticaria and angioedema. AB - H1-antihistamines are the cornerstone of symptomatic treatment in acute and chronic urticaria, in which they not only relieve itching, but also reduce the number, size, and duration of urticarial lesions. Relief of whealing, flaring, and erythema may be incomplete as the vascular effects of histamine are mediated to its action at H2-receptors as well as at H1-receptors, and other vasoactive substances may also be involved. In randomized, prospective, placebo-controlled, double-blind studies, the new low-sedating H1-antihistamines have been found to be effective and safe in urticaria. Sedating antihistamines, although effective, place patients at risk for adverse effects, including decreased psychomotor performance. The response to H1-antihistamines in some types of urticaria, for example, in urticarial vasculitis, is unsatisfactory. An H2-antihistamine administered concurrently with an H1-antihistamine may modestly enhance relief of itching and wheal formation in some patients with urticaria refractory to treatment with an H1-antihistamine alone. The available evidence does not justify the routine addition of H2-antihistamine treatment to H1-antihistamine treatment. PMID- 12113220 TI - Histamine receptors: specific ligands, receptor biochemistry, and signal transduction. AB - During the past few years, there has been a tremendous increase in our understanding of the histamine receptors. Important progress has been made in the development of H1-receptor agonists and the rationalization of H1-receptor-ligand interaction. The recent observation of constitutive H1- and H2-receptor activity has led to a reclassification of H1- and H2-antagonists. For the H3-receptor, a wide variety of selective and potent ligands are currently available and await clinical application. The recent cloning of the H3-receptor gene and the anticipated generation of transgenic mice will facilitate this development. Within the field of signal transduction, a previously unanticipated complexity has been unravelled. With the cloning of the H3-receptor gene, a similar complexity is to be expected. PMID- 12113221 TI - Histamine and antihistamines in anaphylaxis. AB - Anaphylaxis and anaphylactoid reactions are potentially fatal. These disorders are sometimes iatrogenic, and increase with increased exposure to drugs, synthetic substances, and medical procedures. Non-IgE-mediated anaphylactoid reactions are common in medical settings and are clinically indistinguishable from anaphylaxis. These reactions may be unrecognized if a rigid classic definition of anaphylaxis is used. Histamine is a primary mediator of anaphylaxis and signs and symptoms of anaphylaxis can be reproduced by histamine infusion. Histamine triggers a cascade of inflammatory mediators and modulates its own release. H1-antihistamines are adjunctive treatment therapy for acute anaphylaxis and anaphylactoid reactions, in which many mediators of inflammation are involved. Compared with epinephrine, the first-response medication of choice, antihistamines have a slow onset of action, and they cannot block events that occur subsequent to histamine binding to its receptors. Antihistamines are an important component of regimens for the prevention of anaphylaxis and anaphylactoid reactions in patients at risk, and may eventually have more widespread application in the perioperative setting. In some instances, such as with exercise-induced anaphylaxis and reactions to latex in sensitized individuals, prophylaxis regimens are not always effective. H2-antagonists are not detrimental in the therapy of anaphylaxis and many studies show a favorable outcome when combining H1- and H2-antagonist therapy for prophylaxis. They should be added to therapy at the discretion of the treating physician. Because of decreased antimuscarinic and central nervous system side effects, the newer antihistamines can be given in high doses, allowing more complete blockade of histamine receptors. These agents should lead to a reevaluation of the usefulness of antihistamines in both the treatment of acute anaphylaxis and in prophylactic regimens. The unavailability of parenterally administered second-generation H1 antagonists limits their usefulness in acute anaphylaxis and perioperative prophylaxis. PMID- 12113222 TI - Cost-effectiveness of H1-antihistamines. AB - In the health care arena, assessment of a medication's clinical efficacy is no longer enough. It is important to confirm the medication's cost-effectiveness and the improvement in quality of life it provides in relationship to other options in therapy. H1-antagonists are widely used in the treatment of many atopic disorders, especially allergic rhinitis. This review has pointed out cost effectiveness and quality-of-life studies comparing the different H1-antagonists among themselves, and with other treatments used in allergic rhinitis, such as intranasal corticosteroids and allergen immunotherapy. Cost-effective analyses among H1-antagonists in other allergic diseases, such as atopic dermatitis, urticaria and angioedema, and asthma, are lacking at this time. PMID- 12113223 TI - H1-antihistamines and the central nervous system. AB - An extensive body of research exists on CNS effects of H1-antagonists. There is great interest in this area due to the well-known adverse CNS effects associated with first-generation H1-antagonists, and the many new second-generation agents claiming to have nonsedative properties. Because the CNS effects of H1 antagonists are complex and cannot be reflected in one measurement, a variety of assessments of CNS function are required. These range from the subjective (e.g., self-rating of drowsiness) to the objective (e.g. 24 h EEG sleep latency, P300), and from the simple (e.g., critical flicker fusion) to the complex (e.g., actual driving). When these tests are applied to the evaluation of the H1-antagonists currently available, it is clear that there is a real distinction between the older first-generation H1-antagonists and the newer second-generation ones. At the recommended dosages, all the second-generation H1-antagonists are clearly less sedating in more patients than their predecessors. These newer medications do not cross the blood-brain barrier readily; are highly specific for H1 receptors; have little to no anticholinergic, antiserotoninergic, or anti-alpha adrenergic effects; and do not enhance the adverse CNS effects of alcohol or other CNS-active substances such as the benzodiazepines. Since most second generation H1-antagonists are found to be relatively nonsedating, their benefit/risk ratios will be determined more by their other properties such as non CNS adverse effects (e.g., potential to cause cardiac arrhythmias), potency, onset of action, duration of action, ease of administration, and cost. The future role and usefulness of the older sedating H1-antagonists, given the availability of the safer second-generation agents, are unclear at the present time. When H1 antagonist treatment is indicated, physicians should recommend an effective H1 antagonist with a favorable clinical pharmacology profile and a wide margin of safety in patients of all ages. The common, often subclinical, adverse CNS effects produced by the old H1-antagonists remain a major concern and, therefore, these compounds are no longer medications of choice in the treatment of allergic rhinitis, allergic conjunctivitis, or urticaria. PMID- 12113224 TI - Potential cardiac toxicity of H1-antihistamines. AB - Nonsedating H1-antihistamines are widely prescribed for the treatment of allergic disorders because of their lack of sedative and anticholinergic effects; however, certain nonsedating antihistamines such as terfenadine and astemizole are now known to cause QT prolongation and TdP, particularly in overdosage or with concomitant ingestion of imidazole antifungals or macrolide antibiotics. Mechanistic studies showed that the cardiotoxic effects of some nonsedating antihistamines are due to the inhibition of repolarization potassium channels, particularly IKr, which leads to prolongation of the action potential and QT interval, and the development of early after-depolarization, which triggers TdP. Patients at risk of developing TdP, such as those with congenital long QT syndrome, cardiac disease, liver disease, electrolyte disturbance, or those taking drugs that can prolong QT interval, should avoid nonsedating antihistamines that are also capable of prolonging the QT interval. Many questions still need to be answered, such as the role of other potassium channels (IKs, ITo, and Iped) and the relative expression of various potassium channels in different individuals, which may be important in the pathogenesis of TdP with nonsedating antihistamines. There is also a lack of information on the cardiac actions of newer nonsedating antihistamines. The evidence so far indicates that the potential to cause ventricular arrhythmias is not a class effect and that loratadine, cetirizine, and fexofenadine are not associated with QT prolongation, TdP, or other ventricular arrhythmias. It is hoped that with a better understanding of the arrhythmogenic mechanism of nonsedating antihistamines, we will be able to identify patients at risk and prevent any cardiac toxicity associated with H1-antihistamines, and ultimately, death. PMID- 12113225 TI - H1-antihistamines in pregnancy and lactation. AB - Antihistamines may be used for the treatment of allergic rhinitis, upper respiratory infections, urticaria/angioedema, atopic dermatitis, and, rarely, as adjunctive treatment for anaphylaxis, during pregnancy. Because these illnesses may affect maternal comfort and safety as well as threaten the fetus directly (anaphylaxis) or indirectly, they often require therapy during pregnancy. Based on the information available to date, in this chapter we have attempted to provide rational guidelines for the gestational use of H1-receptor antagonists in a manner that will lead to the optimal well-being of both the mother and her infant. As more information becomes available, the recommendations herein may require modification. Although this chapter has dealt specifically with gestational management, a case can be made for considering this information when making therapeutic decisions in all women of childbearing potential. First, most pregnancies are unplanned, and the peak period of fetal vulnerability to drug induced teratogenesis begins the day a woman's period is due. Second, during gestation, substantial alterations in a previously successful but not optimal-for pregnancy chronic therapeutic regimen may be psychologically threatening to the patient and may lead to either uncontrolled disease or unanticipated side effects. Thus, pregnancy-appropriate regimens should ideally be discussed with all women of childbearing age as part of the informed therapeutic decision-making process. PMID- 12113226 TI - H1-antihistamines in children. AB - In children, as in adults, H1-antagonists are useful in the treatment of allergic rhinoconjunctivitis. Level 1 evidence for their efficacy in this disorder has been obtained in many well-designed pediatric studies. The widespread use of H1 antagonists in upper respiratory tract infections or otitis media in children is not supported by a strong scientific rationale. H1-antagonists are not harmful in children with asthma and, indeed, may have some beneficial effects in children with mild asthma. Their role in delaying or preventing asthma from developing in high-risk infants and toddlers is currently an important area of clinical investigation. The evidence base for their use in children with urticaria or atopic dermatitis still contains large gaps. First-generation H1-antagonists are presumed to be safe for use in infants and children. While they have undoubtedly been administered without apparent harm to millions in this age group, they impair CNS function far more commonly than is generally realized. Their use should be restricted to two uncommon situations: children with urticaria or atopic dermatitis whose pruritus is so severe that the sedation produced by an old H1-antagonist, such as hydroxyzine, is a benefit rather than a risk; and children with anaphylaxis who require intravenous diphenhydramine as adjunctive treatment to epinephrine and other modalities. Apart from these exceptions, in patients of all ages, second-generation H1-antagonists free from CNS adverse effects are clearly the medications of choice. Pediatric formulations of the new H1-antagonists cetirizine, fexofenadine, and loratadine are now available for use. PMID- 12113227 TI - H1-antihistamines in the elderly. AB - In the elderly, H1-antihistamine therapy is commonly prescribed for treatment of rhinitis, conjunctivitis, pruritus, eczema, urticaria, and for prophylaxis of anaphylactoid reactions. Second-generation H1-receptor antagonists provide excellent, safe, and effective alternatives to first-generation antihistamines in this population, as in younger patients. As with all medications, the choice of which agent to use must be tailored to the needs of the individual. Treatment should be planned with consideration of concomitant medications and potential drug-drug interactions and drug-disease interactions. First-generation antihistamines should not be used for treatment of allergic rhinitis or urticaria in the elderly. Age-related physiological changes can enhance or complicate the actions of H1-receptor antagonists, especially when these drugs are taken concurrently with other medications and/or in the presence of comorbid disease. Adjustments in dosages are necessary when some agents are used in patients with renal and/or hepatic disease; however, overall, the use of the newer nonsedating antihistamines is safe, effective, and gratifying in the elderly. PMID- 12113228 TI - Structure and classification of H1-antihistamines and overview of their activities. PMID- 12113229 TI - Group support in breast cancer: realistic hope, realistic benefits. PMID- 12113230 TI - FDA approves ZOMETA for treatment of cancer-related bone complications. PMID- 12113231 TI - Dynavax and Coley Pharmaceutical Group initiate rituximab combination trials. PMID- 12113232 TI - Delcath Systems' phase III trial for inoperable cancer is planned at Sydney Melanoma Unit. PMID- 12113233 TI - TransMolecular receives FDA approval for 131-I-TM-601 IND application. PMID- 12113234 TI - Oncolytics Biotech releases REOLYSIN phase I clinical trial results. PMID- 12113235 TI - Hemosol receives clearance to begin study of HEMOLINK. PMID- 12113236 TI - Caelyx/Doxil for the treatment of metastatic ovarian and breast cancer. AB - Caelyx/Doxil is a novel pegylated liposomal formulation of the first-generation anthracycline, doxorubicin. The pharmacokinetics of this polyethylene-glycol coated liposome are characterized by a reduced volume of distribution, a long intravascular circulating half-life and slow plasma clearance compared with free doxorubicin. This, coupled with a small vesicular size, uniquely promotes the localization of Caelyx/Doxil at tumor sites and explains its altered toxicity profile. The FDA and EMEA have approved its use for the treatment of AIDS-related Kaposi's sarcoma and, more recently, for recurrent epithelial ovarian cancer (EOC). Numerous investigations have focused on its use in the treatment of metastatic breast cancer, as well as recurrent squamous cell cervical carcinoma, soft tissue sarcoma, squamous head and neck cancers, prostate cancers and malignant gliomas. Ongoing clinical studies of combination regimens incorporating Caelyx/Doxil will further clarify its role in the treatment of advanced solid tumors. PMID- 12113237 TI - Fulvestrant (Faslodex): current status in the therapy of breast cancer. AB - Fulvestrant (Faslodex, formerly ICI 182,780) is a potent steroidal antiestrogen that mediates its effects by estrogen receptor downregulation. It appears to act as a pure antiestrogen and exhibits none of the negative side effects associated with the partial agonist activity of tamoxifen. It has been shown to be as effective as the oral aromatase inhibitor anastrozole in postmenopausal women with advanced breast cancer who have progressed on prior endocrine therapy, principally tamoxifen. It therefore provides the clinician with an alternative therapeutic strategy following the development of tamoxifen resistance. Fulvestrant might also have potential as a follow-on therapy after tamoxifen in an adjuvant setting and help alleviate some of the concerns surrounding long-term (up to 5 years) tamoxifen therapy. PMID- 12113238 TI - ZD1839 (IRESSA): a selective EGFR-TK inhibitor. AB - The recent development of highly selective, target-based cancer therapeutics, such as ZD1839 has resulted from a greater understanding of tumor biology. Amongst the most promising of new target-based agents are inhibitors of the epidermal growth factor receptor. ZD 1839 is a potent, selective epidermal growth factor receptor tyrosine kinase inhibitor that has demonstrated promising results in early clinical trials. PMID- 12113239 TI - Combination chemotherapy for metastatic breast cancer. AB - Despite more than four decades of effort, the improvement in survival in metastatic breast cancer has been modest. Recently, however, new drugs such as the taxanes have emerged as pivotal agents in the treatment of metastatic disease and they are now being investigated in the adjuvant setting. In addition, the introduction of molecularly targeted therapies such as trastuzumab provides a new paradigm for the development of biologic treatments. The incorporation of trastuzumab into new combination regimens based on potential molecular synergies is a focus of current research. PMID- 12113241 TI - Management of AIDS-related Kaposi's sarcoma: advances in target discovery and treatment. AB - Kaposi's sarcoma is the most common tumor arising in HIV-infected patients and is an AIDS-defining illness by the Centers for Disease Control guidelines. Recent advances in the elucidation of the pathogenesis of KS are uncovering potential targets for KS therapies. Such targets include the processes of angiogenesis and cellular differentiation and the Kaposi's sarcoma herpesvirus/human herpesvirus 8. With the increasing recognition that effective antiretroviral regimens are associated with both a decreased proportion of new AIDS-defining Kaposi's sarcoma cases and a regression in the size of existing Kaposi's sarcoma lesions, most, if not all, Kaposi's sarcoma patients should be advised to take antiretroviral drugs that will maximally decrease HIV-1 viral load. Five agents are currently approved by the US FDA for the treatment of Kaposi's sarcoma; alitretinoin gel for topical administration; and liposomal daunorubicin, liposomal doxorubicin, paclitaxel and interferon-alpha for systemic administration. Many more agents, particularly angiogenesis inhibitors and other pathogenesis-targeted therapies are in early clinical development. Over the next 5 years, we may see even more of these pathogenesis-targeted therapies in trials and just as important we may identify, develop and validate clinically practical tools for assessing the biological effects of these therapies. The next 5 years may also bring a better understanding of the pharmacokinetic interactions among the many agents in the Kaposi's sarcoma and AIDS armamentariums. PMID- 12113240 TI - Aromatase inhibitors in breast cancer therapy. AB - Estrogens are involved in numerous physiological processes and have crucial roles in certain disease states, such as mammary carcinomas. Estradiol, the most potent endogenous estrogen, is biosynthesized from androgens by the cytochrome P450 enzyme complex called aromatase. Aromatase is found in breast tissue and the importance of intratumoral aromatase and local estrogen production is being unraveled. Inhibition of aromatase is an important approach for reducing growth stimulatory effects of estrogens in estrogen-dependent breast cancer. Steroidal and nonsteroidal aromatase inhibitors have shown clinical efficacy for the treatment of breast cancer. The initial nonselective nature of nonsteroidal inhibitors, such as aminoglutethimide, has been greatly reduced in the later generations of inhibitors, anastrozole and letrozole. Mechanism-based steroidal inhibitors, such as 4-hydroxyandrostenedione and exemestane produce potent aromatase inhibition in patients. The potent and selective third-generation aromatase inhibitors, anastrozole, letrozole and exemestane, are approved for clinical use as first-line endocrine therapy in postmenopausal women with metastatic hormone-dependent breast cancer and as second-line endocrine therapy in postmenopausal patients failing antiestrogen therapy alone or multiple hormonal therapies. PMID- 12113242 TI - Dose-intensive chemotherapy in advanced adult soft tissue sarcoma. AB - The treatment of metastatic soft tissue sarcomas in adults is one of the most challenging areas in oncology. While multidisciplinary management of early-stage, localized disease has led to a number of improved outcomes, therapy of unresectable or advanced disease remains problematic. Virtually every conventional cytotoxic agent has been systematically assessed in this malignancy, yet only a handful have demonstrated significant activity. Adriamycin and ifosfamide are the only chemotherapeutic drugs to have consistently produced response rates of over 20% when given as single agents and these two drugs have been exhaustively studied alone or in combination. Recent efforts to improve response rates and, by inference, disease-free survival and overall survival, have involved exploration of high-dose regimen incorporating growth factors and/or autologous cellular support. In this article, the status of dose-intensive chemotherapy in advanced adult soft tissue sarcomas (excluding pediatric histologies, such as Ewing's sarcoma and rhabdomyosarcoma) will be discussed. Emphasis will be placed on data from randomized Phase III trials but information from Phase I/II studies will also be reviewed and recommendations will be made on a systematic analysis of the data. PMID- 12113243 TI - Telomerase as a diagnostic and therapeutic target for cancer. AB - Telomerase is a ribonucleoprotein enzyme responsible for the elongation of telomeres at the ends of chromosomes. It is widely expressed in most cancers, while absent from most normal somatic cells. Telomerase is partially responsible for the cellular immortalization that allows human cancers to progress indefinitely. Due to its widespread occurrence in cancer and its crucial role in the maintenance of the tumor, telomerase is an attractive target for cancer diagnosis and treatment. PMID- 12113244 TI - Anticoagulants in thrombosis and cancer: the missing link. AB - Many cancer patients reportedly have hypercoaguable state, with recurrent thrombosis due to the impact of cancer cells and chemotherapy on the coagulation cascade. A number of retrospective studies showed that cancer patients are at higher risk of developing venous thromboembolism. In addition to the pathological mechanisms associated with tumor-mediated increase in thrombotic events, cancer therapies including chemotherapy, immobilization, cancer surgery and the use of central venous catheters contribute toward a hypercoaguable state and are therefore independent risk factors of venous thromboembolism in cancer patients. Studies have demonstrated that unfractionated heparin or low molecular weight heparin (LMWH) interferes with various processes involved in tumor growth and metastasis. These processes might include fibrin formation, binding of heparin to angiogenic growth factors--such as basic fibroblast growth factor and VEGF- modulation of tissue factor, release of tissue factor pathway inhibitor and other mechanisms. Clinical trials have suggested an improved efficacy of LMWH, as compared with UFH on the survival of cancer patients with deep vein thrombosis. Similarly, the impact of warfarin on the survival of cancer patients with thromboembolic disorders was demonstrated. Recent studies from our laboratory defined the role of the LMWH (tinzaparin), warfarin, antifactor VIIa and recombinant tissue factor pathway inhibitor in the modulation of angiogenesis, tumor growth and tumor metastasis. PMID- 12113245 TI - Proceedings of the Third Turtle Creek Conference. April 29-May 1, 2001. Dallas, Texas, USA. PMID- 12113246 TI - [Reaction to, "Tetanus prevention in injuries; guidelines for dental practice," Ned Tijdschr Tandheelkd 1995; 102: 266-8]. PMID- 12113247 TI - Abstracts of the 12th Annual Symposium of the London Hypertension Society. London, UK. May 8, 2001. PMID- 12113248 TI - A circle of prayer. Bringing hope to the oncology unit. PMID- 12113249 TI - Vicious cycle. PMID- 12113250 TI - Under control. PMID- 12113251 TI - Switched on. PMID- 12113252 TI - Prayer power. PMID- 12113253 TI - Abstracts of the Fifth Annual Meeting of the Society for Clinical Epidemiology and Health Care Research (SCHR). Atlanta, Georgia, USA. May 3, 2002. PMID- 12113255 TI - Re: Carotid stenting with distal-balloon protection via the transbrachial approach. PMID- 12113254 TI - Re: Analytical modeling and numerical simulation of forces in an endoluminal graft. PMID- 12113256 TI - The actin-severing activity of cofilin is exerted by the interplay of three distinct sites on cofilin and essential for cell viability. AB - Cofilin/actin-depolymerizing factor is an essential and conserved modulator of actin dynamics. Cofilin binds to actin in either monomeric or filamentous form, severs and depolymerizes actin filaments, and speeds up their treadmilling. A high turnover rate of F-actin in actin-based motility seems driven largely by cofilin-mediated acceleration of directional subunit release, but little by fragmentation of the filaments. On the other hand, the filament-severing function of cofilin seems relevant for the healthy growth of cells. In this study, we have characterized three mutants of porcine cofilin to elucidate the molecular mechanism that underlies the filament-severing activity of cofilin. The first mutant could neither associate with actin filaments nor sever them, whereas it effectively accelerated their treadmilling and directional subunit release. The second mutant bound to actin filaments, but failed to sever them and to interfere with phalloidin binding to the filament. The third mutant could associate with actin filaments and sever them, although with a very reduced efficacy. Of these mutant proteins, only the last one was able to rescue Deltacof1 yeast cells and to induce thick actin bundles in mammalian cells upon overexpression. Therefore, the actin-severing activity of cofilin is an essential element in its vital function and suggested to be exerted by co-operation of at least three distinct sites of cofilin. PMID- 12113258 TI - Nonlinear Dynamics and Chaos in Astrophysics: A Festschrift in Honor of George Contopoulos. Proceedings of the thirteenth conference of the series Florida Workshops in Nonlinear Astronomy. February 12-14, 1998. Gainesville, Florida, USA. PMID- 12113259 TI - Scientific Cooperation and Conflict Resolution. Conference of the New York Academy of Sciences. January 28-30, 1998. New York, New York, USA. PMID- 12113260 TI - I'll try to accommodate you. PMID- 12113261 TI - From the Centers for Disease Control and Prevention. Disseminated infection with Simiae-avium group mycobacteria in persons with AIDS--Thailand and Malawi, 1997. PMID- 12113262 TI - From the Centers for Disease Control and Prevention. West Nile Virus activity- United States, 2001. PMID- 12113264 TI - Bibliography. Current world literature. Mineral metabolism. PMID- 12113263 TI - Results from late-breaking clinical trial sessions at the American College of Cardiology 51st Annual Scientific Session. PMID- 12113265 TI - Bibliography. Current world literature. Renal pathophysiology. PMID- 12113266 TI - A comparison of interrupted and continuous sutures for tracheal anastomoses in sheep. AB - OBJECTIVE: To study the tensile strength of tracheal anastomoses. DESIGN: Experimental study. SETTING: University medical school laboratory, Germany. ANIMALS: 15 sheep. INTERVENTIONS: Tracheal anastomoses with three different suturing techniques: a continuous suture and interrupted sutures with either a monofilament or a polyfilament material. Anastomoses were tested to breaking after being in place for 1, 2, 4, 8 or 24 weeks. RESULTS: After one week, with all three materials, the trachea broke at the anastomosis. In animals that survived longer, the trachea broke further away from the anastomosis. There was no significant difference between the mean values of the breaking force for continuous sutures and single interrupted sutures (p = 0.9). CONCLUSION: The suturing technique (continuous or interrupted) has less relevance for the tensile strength of the anastomosis than in vitro experiments suggest. PMID- 12113267 TI - Development and validation of QoL5 for clinical databases. A short, global and generic questionnaire based on an integrated theory of the quality of life. AB - OBJECTIVE: To develop and validate a short, global, and generic quality of life (QoL) questionnaire for clinical databases. The construct validity and item weighting of existing questionnaires are increasingly questioned. DESIGN: Cross sectional population study. SUBJECT: 2460 Danes aged 18-88 years, randomly selected through the Danish Central Person Registry. INTERVENTIONS: Ten questions covering the spectrum of the integrative theory of QoL together with the Nottingham Health Profile (NHP), Sickness Impact Profile (SIP), and self estimated QoL questionnaire were sent by mail. A test-retest study of 50 people was conducted after one month. MAIN OUTCOME MEASURES: Construct and criterion validity, reliability, and sensitivity. RESULTS: QoL5 correlations with SIP, NHP, Self-estimated QoL were 0.37, 0.52, and 0.76, respectively, and increased among those who were unwell. Cronbach's alpha was 0.69. All correlations in Siegel's test were over 0.6, and the test-retest correlation was 0.82. Only 12 respondents in each group will be needed to detect a difference of 10% in the QoL score between two groups. CONCLUSIONS: QoL5 is a valid global and generic QoL measurement. Despite the use of only five questions, internal consistency and sensitivity were acceptable. So a relevant and practical outcome measurement is available for clinical databases. PMID- 12113268 TI - Cholecystokinin stimulation leads to increased oxytocin secretion in women. AB - OBJECTIVE: To find out if cholecystokinin (CCK) stimulates the secretion of oxytocin in humans, and if there are any differences in secretion between healthy women and those with normal-transit constipation. DESIGN: Prospective open study. SETTING: Teaching hospital, Sweden. SUBJECTS: 8 healthy female volunteers and 6 women with chronic refractory normal-transit constipation. INTERVENTIONS: Subjects were fasted before experiments. On one day they were given emulsified corn oil and another an intravenous injection of 1 Ivy dog unit (IDU) CCK/kg body weight. Blood samples were taken before each experiment at 10 minutes and at the time the experiments started. Blood samples were also taken after each experiment at 10, 20, 30, 45, 60, 90 and 120 minutes. MAIN OUTCOME MEASURES: Concentrations of CCK and oxytocin. RESULTS: Ingestion of corn oil significantly increased the plasma concentration of CCK in both groups (healthy women p = 0.03 and constipated women p = 0.008). Injection of CCK also led as expected to hypercholecystokininaemia in both groups (p = 0.008 and p = 0.03, respectively). The corn oil increased oxytocin secretion in both groups (p = 0.02 and 0.03, respectively) and exogenous CCK increased the secretion still further (p = 0.008 and 0.03, respectively). CONCLUSIONS: Both corn oil and injection of CCK led to an increased CCK concentration in plasma. Oxytocin was secreted in response to endogenous as well as exogenous CCK stimulation. There was no difference between healthy and constipated women in either parameter analysed. PMID- 12113270 TI - Simple technique for repair of complete rectal prolapse using a circular stapler with Thiersch procedure. PMID- 12113269 TI - Proteolysis in severe sepsis is related to oxidation of plasma protein. AB - OBJECTIVE: To test the hypothesis that the oxidation of proteins is part of the mechanism of proteolysis in catabolic states. DESIGN: Prospective, observational study. SETTING: Critical care unit at a university teaching hospital, New Zealand. PATIENTS: 13 patients (6 male, 7 female; median age 61, range 26-76 years) who were admitted to the Department of Critical Care Medicine at Auckland Hospital with a diagnosis of severe sepsis. The median APACHE II score during the first 24 hours after admission was 22 (range 15-34). Control values of protein carbonyl in plasma were established in 15 healthy volunteers. INTERVENTIONS: We made serial measurements of total body protein (by neutron activation analysis) and plasma protein carbonyl (by ELISA) concentrations over a period of 10 days. MAIN OUTCOME MEASURE: Plasma protein carbonyl concentration and total body protein. RESULTS: The total amount of body protein decreased significantly over the 10 days (p < 0.001). Plasma protein carbonyl concentrations were significantly higher in the septic patients than in the control group throughout the study period (p < 0.0001). There was a significant reduction in plasma protein carbonyl concentration over the study period (p < 0.008). The early increase in the concentration of protein carbonyl formation was followed by an ongoing loss of body protein. There was a significant positive correlation between total body protein and plasma protein carbonyl (p < 0.03). CONCLUSIONS: Severe sepsis results in oxidation of plasma proteins and this precedes and is related to the loss of body protein. PMID- 12113271 TI - Successful management of lacerated lung by thoracotomy without resection. PMID- 12113272 TI - Resections of the liver, inferior vena cava, and right kidney for recurrences over 10 years after right adrenalectomy for carcinoma. PMID- 12113273 TI - Review of wound healing with reference to an unrepairable abdominal hernia. AB - A 58-year-old man has been under our care with an inguinal hernia that has recurred 8 times. This stimulated us to review the biochemistry of wound repair. We studied the composition of his collagen and tried to find out whether it was intrinsically faulty or whether its fault had been caused by the medication he was taken. PMID- 12113274 TI - Pancreas-sparing duodenectomy: technique and indications. AB - OBJECTIVE: To assess the feasibility, safety, and short-term functional outcome of a pancreas-sparing duodenectomy. DESIGN: Prospective, uncontrolled study. SETTING: University hospital, Sweden. SUBJECTS: Four patients with extensive lesions in the duodenum (2 familial polyposis, 1 villous adenoma, and 1 giant multiple lipoma). RESULTS: All 4 patients had a duodenectomy with sparing of 1 1.5 cm of the duodenal bulb and reinplantation of the biliary and pancreatic ducts into the jejunum. Except for one early postoperative bile leak the operative and postoperative courses were uneventful. The functional results have been promising with unaltered alimentary function in the 3 patients who had no preoperative outlet obstruction and complete resolution of symptoms in the patient with duodenal lipomas who had chronic incomplete obstruction preoperatively. CONCLUSIONS: Although the indications for pancreas-preserving duodenectomy are limited, the procedure can be done safely with gastrointestinal function maintained. PMID- 12113275 TI - Prospective randomised trial of computer-aided diagnosis and contrast radiography in acute small bowel obstruction. AB - OBJECTIVE: to compare the ability of computer-aided diagnosis and contrast radiography for the diagnosis of acute mechanical small bowel obstruction. DESIGN: Prospective randomised trial. SETTING: Kaunas University of Medicine, Lithuania. SUBJECTS: 80 patients with small bowel obstruction with no clinical evidence of strangulation who were randomised into two groups (n = 40 in each) to be investigated by computer-aided diagnosis and contrast radiography. INTENVENTIONS: 37 patients required operation (46%). MAIN OUTCOME MEASURES: specificity, sensitivity, false positive and negative predictive values of the 2 methods; time necessary to make the diagnosis; and morbidity and mortality. RESULTS: The specificity, sensitivity, positive and negative predictive values in the diagnosis of complete acute small bowel obstruction for the computer-aided group were 100%, 87.5%, 100% and 92.3%, and for the contrast radiography group 100%, 76.9%, 100% and 90%, respectively. The mean time period for making the diagnosis was 1 hour in the computer-aided group and 16 hours in the radiography group (p < 0.001). The overall mortality was 3% and morbidity 9%. CONCLUSION: Computer-aided diagnosis had no significant advantage over contrast radiography in the accuracy of diagnosis of the character of small bowel obstruction. However, significantly less time was needed to make the diagnosis in the computer aided group. PMID- 12113276 TI - Prognostic value of various staging and grading systems in proximal colon cancer. AB - OBJECTIVE: To compare the ability of various methods of staging and grading to predict survival in proximal colon cancer. DESIGN: Retrospective study. SETTING: University hospital, Finland. SUBJECTS: 153 patients with primary proximal colon cancer. MAIN OUTCOME MEASURES: Staging by four classification systems, grading by two grading systems, and survival analysis based on Kaplan-Meier survival curves. RESULTS: In all staging systems the survival curves of different stages differed significantly from each other. The modified Dukes' classification was still the best predictor of survival. Grade of tumours had no significant effect on long term survival, but short term survival was affected adversely by the presence of anaplastic tissue. Tumours with no mucin had a worse prognosis than those that produced mucin. All staging methods were superior to either of the histological grading systems tested as prognosticators. Tumour depth correlated with the operator's clinical impression of radicality of operation, and also predicted survival. CONCLUSION: The clinicopathological modified Dukes' staging system was the most powerful prognosticator in proximal colonic cancer and its use in clinical practice should continue. Even a small amount of anaplastic tissue (> or = 5%) had an adverse effect on short term survival. PMID- 12113277 TI - Fournier's gangrene: diagnostic approach and therapeutic challenge. AB - OBJECTIVE: To identify the prognostic variables and to assess the role of aggressive management in patients with Fournier's gangrene. DESIGN: Retrospective study. SETTING: University hospital, Greece. SUBJECTS: 11 patients (9 men and 2 women) with Fournier's gangrene admitted between April 1986 and December 2000. INTERVENTIONS: Early aggressive debridements. MAIN OUTCOME MEASURES: Postoperative course, hospital stay, outcome, morbidity, and mortality. RESULTS: The mean age was 65 years (range 17-90) and the mean (SD) duration of hospital stay was 35 (8) days (range 8-62). The aetiology was identified in 8 patients and idiopathic in 3. Predisposing factors (diabetes, n = 4; heart failure, agranulocytosis, and alcohol misuse, n = 1 each) were identified in 6 patients. All patients except one had raised serum glucose concentrations and low serum albumin values. On admission the white cell count was >15 x 10(9)/L in 10, serum sodium <135 mmol/L, mean (SD) serum creatinine 124 (27) micromol/L, and C reactive protein >150 mg/L was found in all patients. 3/9 male patients required partial excision of the scrotum. Temporary faecal diversion was done for 3 patients. A mean of 3 aggressive repeated debridements (range: 3-6) were required. Nine patients survived and two patients died. CONCLUSION: Rapid and accurate diagnosis remains the key to achieving a successful outcome. Abnormal laboratory variables on admission may suggest the diagnosis. Early, repeated, aggressive debridement is essential for a successful outcome. PMID- 12113279 TI - Premier Initiative improves outcomes in spine surgery. PMID- 12113278 TI - Outcome of treatment of ruptured abdominal aortic aneurysms depending on the type of hospital. AB - OBJECTIVE: To compare the outcome of patients operated on acutely for ruptured abdominal aortic aneurysms (AAA) or otherwise symptomatic aortic aneurysms in a university hospital and in two county hospitals by the same group of vascular surgeons. DESIGN: Retrospective study. SETTING: 1 university and 2 county hospitals, Sweden. SUBJECTS: 108 patients operated on urgently for AAA, 81 at the university hospital, and 27 at the county hospitals between January 1992 and December 1998. INTERVENTION: Repair of the AAA. MAIN OUTCOME MEASURES: Morbidity and mortality. RESULTS: 21 of the 81 patients having urgent repair of an AAA at the university hospital (26%) had been transferred from the county hospitals. Thirteen patients were not operated on, 7 because of their poor general condition and great age (median 84 years), 3 who refused operation, and 3 in whom the diagnosis was incorrect. During the same time period a further 27 haemodynamically unstable patients were operated on by the same vascular surgeons at the county hospitals. The on-table mortality for patients with ruptured AAA and shock was 5/43 (12%) at the university hospital and 4/27 (15%) at the county hospitals. The corresponding in-hospital rates were 11/43 (26%) and 11/27 (41%). Mortality was significantly higher if the operation was delayed by more than 45 minutes. The incidence of postoperative complications was the same in both hospitals. CONCLUSION: If a patient with a ruptured AAA and shock is admitted to the county hospital and operated on by a specialist vascular surgeon the outcome is fully acceptable. The difference seems to be related to the postoperative period. To what extent the delay caused by the surgeon's journey to the county hospital has any influence on the outcome is not possible to evaluate. PMID- 12113280 TI - Software helps reduce adverse events and determine financial costs. PMID- 12113281 TI - COPD care guidelines issued. PMID- 12113282 TI - Guidelines: more patients should get lipid screenings. PMID- 12113283 TI - A method to detect the G894T polymorphism of the NOS3 gene. Clinical validation in familial hypercholesterolemia. AB - An endothelial nitric oxide synthase gene (NOS3) polymorphism in exon 7 (G894T), resulting in Glu298Asp substitution at protein level, has been associated with myocardial infarction, hypertension and coronary atherosclerosis in some populations. This polymorphism is usually identified by polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP). However, the procedures described to date do not eliminate the possibility of misclassification and either require confirmation by DNA sequencing or are time-consuming. In this study, a PCR-RFLP procedure to detect the G894T polymorphism at the NOS3 was optimized by the introduction of a constitutive cleavage site in the amplification product. This cleavage site provides an internal control for enzymatic activity to avoid mistyping. The method was validated by the study of 35 white unrelated individuals with familial hypercholesterolemia and 70 controls. The frequency of the variant allele (T) was similar between both groups (27% vs. 22%, NS), and comparable to the frequency found in other white populations. However, future studies are necessary to confirm these data. In summary, the optimized procedure for detection of the G894T NOS3 polymorphism is rapid, simple, and does not require confirmatory tests. Using this method, we found no association between this polymorphism and familial hypercholesterolemia. PMID- 12113284 TI - Increasing the sensitivity of single-strand conformation polymorphism analysis of the LDLR gene mutations in brazilian patients with familial hypercholesterolemia. AB - Mutations in the low-density lipoprotein receptor (LDLR) gene cause familial hypercholesterolemia (FH), one of the most common single gene disorders. It is thought that FH affects approximately 1 of 500 individuals in most populations. Single-strand conformation polymorphism (SSCP) analysis is widely used to detect mutations in the LDLR gene. However, several factors such as temperature, pH, running time, gel composition and size of the DNA fragments can influence its sensitivity. We have optimized the electrophoretic conditions to screen mutations in the promoter region and exons 1-18 of the LDLR gene by varying temperature (5 degrees C, 8 degrees C, 12 degrees C and 15 degrees C), voltage (300 to 600 V), and running time (1 to 4 hours) in the semi-automated GenePhor system (Amersham Biosciences). The efficiency of the method was evaluated by using 30 positive controls (DNA samples with mutations and polymorphisms in the LDLR gene, previously characterized) and DNA samples from 90 Brazilian patients with FH. Our results show that the use of two temperatures (5 degrees C and 15 degrees C) in combination with other optimized conditions resulted in high mutation detection rate (97%), which was considered appropriate for routine screening. Therefore, this strategy could be useful for the diagnosis of genetic disorders, cancer, and for pharmacogenetic studies. PMID- 12113285 TI - Hormone replacement therapy in postmenopausal women and its effects on plasma lipid levels. AB - Postmenopausal women run the same risks of coronary heart disease as men. The lipid alterations observed at this time reflect increased blood levels of total cholesterol, low-density lipoprotein cholesterol (LDL-C) and lipoprotein (a), and reduced high-density lipoprotein cholesterol (HDL-C) levels. These changes lead to a higher risk of coronary artery disease, and hormonal therapy has a favorable effect on lipid metabolism. In this paper we review the literature on hormone replacement therapy (HRT) in postmenopausal women with the emphasis on the role of lipids in the pathogenesis of coronary heart disease, and on the action of estrogens and their correlation with progestogens, as well as routes of HRT administration. We conclude that the HRT changes the lipid profile in a potentially anti-atherogenic direction, usually reducing LDL-C and increasing HDL C and triglycerides. Otherwise, for postmenopausal women with established coronary disease HRT is not recommended. PMID- 12113286 TI - The effect of N-acetylcysteine supplementation upon viral load, CD4, CD8, total lymphocyte count and hematocrit in individuals undergoing antiretroviral treatment. AB - Individuals infected with the human immunodeficiency virus (HIV-1) present with decreased CD4, a progressive increase in viral load, compromised cell immune defense, and hematologic alterations. The aim of this study was to assess the serum viral load, CD4, CD8, lymphocyte count and hematocrit at the beginning of antiretroviral therapy in individuals who were supplemented with N-acetylcysteine (NAC). Twenty volunteers participated in this double-blind, placebo-controlled 180-day study. Ten participants received 600 mg of NAC per day (NAC group) and the other ten serving as a control group received placebo. The above mentioned parameters were determined before treatment, and after 60, 120 and 180 days. In NAC-treated patients hematocrit remained stable and an increase in CD4 cell count took place earlier than that in the control group. PMID- 12113287 TI - An evaluation of antiretroviral therapy associated with alpha-tocopherol supplementation in HIV-infected patients. AB - In HIV-infected patients, an increase in the production of oxygen-reactive species (ROS) is observed, with a consequent reduction of plasma levels of antioxidants such as alpha-tocopherol. The nuclear transcription factor-kappaB (NF-kappaB) is activated by a prooxidant state in the infected T cells through the release of its inhibitory subunit I-kappaB. The aim of the present work was to evaluate the behavior of hematological parameters and markers of anemia in HIV infected patients who underwent antiretroviral therapy associated with 800 mg/day alpha-tocopherol supplementation. Blood samples were collected from supplemented (n=9) and not-supplemented (n=9) HIV-seropositive patients (n=18). We observed a decreased viral load in the alpha-tocopherol-supplemented group (p<0.05); other changes, such as an increase in the CD4/CD8 ratio, in the hematocrit and in the hemoglobin concentration were also observed, though lacking statistical significance. We conclude that antiretroviral therapy in association with alpha tocopherol (800 mg/day) supplementation is more effective in reducing viral load levels and also, possibly, in recovering other hematological parameters after a 60-day period of use. PMID- 12113288 TI - Effects of re-education in eating habits and physical activity on the lipid profile of obese teenagers. AB - Twenty-five teenagers, 13 males and 12 females, some obese and others overweight, aged between 12 and 18 years, were studied over 8 months, under the supervision of a multidisciplinary team. Effects of re-education in eating habits and physical activity on the lipid profile were evaluated. Dyslipidaemia characterised by increased levels of total cholesterol, low-density lipoprotein cholesterol and triglycerides was obeserved in 64%, 12% and 44% of the teenagers, respectively. Whereas decreased levels of high-density lipoprotein cholesterol were observed in 28%, tendency to hypertension has been observed in 36% of the teenagers. After 8 months, the number of teenagers with total hypercholesterolaemia and hypertriglyceridaemia decreased to 32% and 24%, respectively. Low-density lipoprotein cholesterol levels did not vary significantly. High-density lipoprotein-cholesterol levels increased in 17% of participants. Reduction of blood pressure occurred in most teenagers. These data suggest that re-education programmes in eating habits associated with changes in behaviour and physical activity can benefit obese teenagers and prevent various diseases. PMID- 12113289 TI - Myeloperoxidase-mediated protein oxidation: its possible biological functions. AB - Oxidation of proteins occurs both as a side-effect of aerobic energy metabolism and as an effect of specific metabolism of phagocytic polymorphonuclear granulocytes producing O2- and H2O2. In contrast to other cells, which control their H2O2 level by degrading it to O2 and H2O, polymorphonuclear neutrophilic leukocytes (PMN) use H2O2 as a substrate for oxidizing chloride ions to HOCl which rapidly react with all neighboring thiol, disulfide and amino residues. Chloramines, which are the most abundant HOCl reaction products, react with proteins, modifying only certain exposed methionine and cysteine residues. This may account for selective inactivation of a number of enzymes, carrier proteins and peptide mediators, including the alpha1-proteinase inhibitor, alpha2 macroglobulin and plasminogen activator inhibitor. Inactivaton of plasma proteinase inhibitors protects PMN elastase, collagenase, cathepsin G and other serine proteases in the inflammatory foci. This promotes proteolytic degradation of damaged tissue, removal of bacterial debris and wound healing, as well as tissue remodeling related to the inflammatory processes. Oxidative control of protease-anti-protease balance affects the development of the inflammatory processes. Moreover, inactivation of plasma proteinase inhibitors facilitates primary antigen processing, upregulates lymphocyte proliferative response and activates the local immune response. Oxidation produces a specific protein tagging which attracts and stimulates immune active cells. Therefore, humoral response against oxidatively modified proteins occurs more effectively than that of the native proteins. The effect is dose-dependent with respect to the amount of oxidant employed. Glycol aldehyde, which is the serine chloramine spontaneous decay product, in mice immunized with glycol aldehyde-modified egg-white albumin, yields specific IgG production manifold higher than that in mice immunized with native albumin. Immunopotentiation is produced by proliferation expansion of the same immunocompetent clones. Oxidative tagging of proteins may also affect the autoimmune-type reaction. Thus, a growing body of data suggest that the specific role of protein oxidation by activated PMN is oxidative protein tagging facilitating further development of the immune reaction. PMID- 12113290 TI - Novel haptoglobin insertion/deletion polymorphism is associated with the lipid profile and C-reactive protein (CRP) concentration. AB - We discovered a hitherto undescribed insertion/deletion (I/D) polymorphism of 7 base pairs in intron 4 of the HP1 allele and intron 4 and 6 of the HP2 allele. Genotyping was performed in 311 Belgian subjects. The association between serum haptoglobin (Hp) and lipid concentrations and the Hp I/D genotypes was investigated. Genotype distribution in 103 Hp 1-1 phenotypes (50.5% DD, 39.8% DI, 9.7% II) and D allele frequency (0.70) were in close agreement with the Hardy Weinberg equilibrium. No association between Hp concentrations and the Hp I/D genotypes could be found. Apolipoprotein (apo)A2, apoB and cholesterol concentrations were slightly lower in Hp II compared to DI and DD. Low-density lipoprotein (LDL)-cholesterol and ultrasensitive C-reactive protein (CRP) concentrations and the apoA1/apoA2 ratios differed significantly between Hp D/I genotypes. We added a further component to the molecular heterogeneity of Hp by the detection of an I/D polymorphism. Studies on the Hp I/D polymorphism in various populations open perspectives for further investigation of the distribution pattern of the human Hp gene. The association of the Hp I/D polymorphism with various lipid parameters might add a further component to the complex and multifaceted lipid metabolism. PMID- 12113291 TI - Apolipoprotein E2/E2 genotype in combination with mutations in the LDL receptor gene causes type III hyperlipoproteinemia. AB - The primary genetic cause of type III hyperlipoproteinemia is the homozygous presence of the apolipoprotein E2 allele. However, only approximately 1% of subjects with the apolipoprotein E2/E2 genotype develop type III hyperlipoproteinemia. Other factors are therefore necessary to express type III hyperlipoproteinemia. Two individuals were identified as having type III hyperlipoproteinemia (triglyceride to very low-density lipoprotein (VLDL) cholesterol ratio >0.3). However, in contrast to unchanged or slightly decreased low-density lipoprotein (LDL)-cholesterol levels typically observed in type III patients, elevated LDL-cholesterol levels were observed. The expected apolipoprotein E2/E2 isoform was confirmed by genetic analysis. To explain the elevated LDL-cholesterol level, single strand conformation polymorphism analysis was performed to screen for mutations in the LDL receptor gene. In both individuals, mutations causing an impaired LDL receptor function (2 bp insertion in exon 3 and Glu119 --> Gly mutation in exon 4) were identified. In six more unrelated individuals, these mutations combined with the common apolipoprotein E3/E3 genotype, resulted in an isolated, severe LDL-cholesterol elevation. Our results indicate that the level of LDL receptors plays an important role in remnant clearance, and that the combination of the binding-defective apolipoprotein E2 with a defective LDL receptor precipitate type III hyperlipoproteinemia. PMID- 12113292 TI - High-sensitivity human thyroglobulin (hTG) immunoradiometric assay in the follow up of patients with differentiated thyroid cancer. AB - Circulating human thyroglobulin (hTG) measurement has a pivotal role in the management of patients affected by differentiated thyroid cancer (DTC). Generally, hTG serum concentration less than 1 ng/ml is considered a marker of complete remission after total thyroid ablation. Recently, high-sensitivity immunoradiometric assays (IRMA) have been developed to detect very low hTG serum concentrations. The present study was undertaken to test a newly developed high sensitivity hTG IRMA and to evaluate its diagnostic performance and reproducibility in the follow-up of patients affected by DTC. We retrospectively selected 156 patients without signs of recurrence and 39 patients with DTC recurrence. Serum samples were collected during L-thyroxine (T4) suppressive therapy (ONT4) and 4 weeks after T4 withdrawal (OFFT4), and hTG was measured by a specific high-sensitivity IRMA (DYNOtest Tg-plus, BRAHMS Diagnostica GmbH, Berlin, Germany). Sera showing the presence of antibodies against hTG (AbhTG) or hTG-recovery less than 80% were excluded from the study. The receiver operator characteristic (ROC) curve analysis was performed to select the best cut-off levels, and diagnostic performance of the marker was evaluated. By using ONT4 cut off level of 0.2 ng/ml and OFFT4 cut-off level of 0.5 ng/ml we obtained a sensitivity/specificity/accuracy profile of 0.92/0.98/0.97 and 0.97/0.98/0.98, respectively. We found false-negative results in three (12%) and one (4%) out of 24 patients with cervical recurrence by using 0.2 and 0.5 ng/ml cut-off levels, respectively. However, we found false-negative results in 13 (54%) and six (25%) patients when 1.0 ng/ml cut-off level was used. Finally, DYNOtest Tg-plus showed a very satisfactory intra- and inter-assay reproducibility in the very low hTG concentration range. Based on our data, we conclude that DYNOtest Tg-plus assay is effective and accurate in evaluation of patients with DTC. PMID- 12113293 TI - Tissue transglutaminase-serology markers for coeliac disease. AB - Serology markers of coeliac disease (CD) - antigliadin IgA/IgG antibodies (AGA/AGG) with purified alpha-gliadin, antiendomysium IgA antibodies (EmA) and anti-tissue transglutaminase (atTG) IgA/IgG antibodies--determined in 1451 serum samples, were analysed with respect to different screening algorithms. Determination of atTG using five ELISA methods was compared taking into account the impact of human recombinant antigen and IgG class of atTG. A subgroup of 119 patients undergoing small intestinal biopsy was used to calculate sensitivity and specificity of CD markers. The highest sensitivity (94%) was obtained for AGG, and the highest specificity (93.5%) was obtained for EmA. All coeliac disease patients were detected using the combination of all four CD markers, resulting in 100% sensitivity. CD and type 1 diabetes mellitus autoantigens were determined in 139 diabetic patients. The atTG IgA mean value (16.7 IU/ml) was higher in the antiglutamate dehydrogenase antibody (GAD)-positive subgroup, where at least one CD marker was positive in 83.6% subjects. In the GAD-negative subgroup atTG IgA was 8.73 lU/ml and at least one CD marker was positive in 57.4% subjects. atTG in IgA and IgG classes could be recommended as valuable serological markers of CD in the differential diagnosis of malabsorption as well as in various screening algorithms. ELISA determination of atTG with human antigen could increase the specificity, especially in patients with other autoimmune diseases. PMID- 12113294 TI - Serum adenosine deaminase and cytidine deaminase activities in patients with systemic lupus erythematosus. AB - We evaluated serum total adenosine deaminase, its isoenzymes adenosine deaminase 1 and adenosine deaminase-2, and cytidine deaminase activities in 24 patients with active systemic lupus erythematosus, and in 26 healthy control subjects, and found the means +/- SD values to be 21.38 +/- 5.96 IU/l, 3.74 +/- 2.12 IU/l, 17.72 +/- 5.02 IU/l and 17.89 +/- 4.62 IU/l, respectively in the patients, and 14.97+/- 4.71 IU/l, 4.01 +/- 1.35 IU/l, 10.91 +/- 3.91 IU/l and 7.39 +/- 3.97 IU/l, respectively in the control subjects. When compared to the healthy controls, serum total adenosine deaminase, adenosine deaminase-2 and cytidine deaminase levels were significantly higher (p<0.001) in systemic lupus erythematosus patients, but the decrease of adenosine deaminase-1 level was not statistically significant (p>0.05). The increased adenosine deaminase-2 may be of macrophage origin. It closely correlated with clinical signs of active systemic lupus erythematosus. The membranes of polymorphonuclear neutrophils may be damaged, and cytidine deaminase may be released into serum. In conclusion, serum total adenosine deaminase, adenosine deaminase-2 and cytidine deaminase activities may serve as useful indicators for evaluating disease activity in patients with active systemic lupus erythematosus. PMID- 12113295 TI - Effects of oral N-acetylcysteine on plasma homocysteine and whole blood glutathione levels in healthy, non-pregnant women. AB - Oral N-acetylcysteine supplementation in nine young healthy females induced a quick and highly significant decrease in plasma homocysteine levels and an increase in whole blood concentration of the antioxidant glutathione. N acetylcysteine impresses as an efficient drug in lowering homocysteine concentration and might be beneficial for individuals with hyperhomocysteinemia who are at increased risk of cardiovascular disease. PMID- 12113296 TI - Evaluating sequential values using time-adjusted biological variation. AB - One can compare the difference between two sequential values with the biological variation. Biological variation is a measure of the random disturbances of an analyte's value, measured at different times. When the difference > Z square root 2 SD(BV) then the difference is due to an underlying disease process or physiologic change. A Z value of 1.96 yields a 95% confidence limit. When using multiple sequential values or time periods exceeding that for the empirically derived biological variance, a random walk model allows one to estimate the spread of the variance. For a difference, (delta) to be significant, delta > Z square root 2n SD(BV), where n is the ratio of time reflecting the longer time period divided by the shorter time period. Not all variances grow to this degree over time, because restoring forces diminish the extent of random disturbances. The relationship between a biological variance measured over a longer time period to the one measured over a shorter period can be expressed in terms of this restoring force as SD2 BV,n = SD2 BV,1 sigma(n) j=1 e(-2)(j-1)phi, where n is the ratio of time periods. One can calculate phi using this formula and a spreadsheet. From phi one can calculate the biological variance for any time period, within experimental limits, and compare the difference in sequential values with it. Test intervals can be calculated based on these biological variances. PMID- 12113297 TI - High-dose methylprednisolone therapy in multiple sclerosis increases serum uric acid levels. AB - Uric acid, which is the final product of purine nucleoside metabolism, is a strong peroxynitrite scavenger. Several studies report on lower serum uric acid levels in multiple sclerosis. In this study, we investigated serum uric acid levels before and after high-dose methylprednisolone treatment (intravenous 1 g/day/5 days) in multiple sclerosis patients. Blood samples from 25 definite multiple sclerosis patients (11 male and 14 female) before and after methylprednisolone treatment (days 0, 6 and 30) and from 20 healthy donors (9 male and 11 female) were analyzed. Serum uric acid levels were measured using a quantitative enzymatic assay (Elitech diagnostics, Sees, France) according to the manufacturer's protocol, and the results were standardized using a commercial uric acid standard solution. We observed significantly increased serum uric acid levels 1 day after the termination of the therapy (day 6). These differences were sustained for 30 days after starting treatment (during remission period). Mean serum uric acid levels were significantly higher in the control group. These results suggest that increasing the uric acid concentration may represent one of the possible mechanisms of action of methylprednisolone in multiple sclerosis. PMID- 12113298 TI - Evaluation of accuracy and uncertainty of ELISA assays for the determination of interleukin-4, interleukin-5, interferon-gamma and tumor necrosis factor-alpha. AB - The comparison of analytical results calls for validation of the assays used, i.e. for the documentation of accuracy (trueness and precision), linearity and specificity. In addition, there is a growing demand for evaluation and documentation of traceability and uncertainty of analytical results. However, models for establishing the traceability and uncertainty of immunoassay results are lacking. Sandwich enzyme-linked immunosorbent assays (ELISAs) were developed for determination of the human cytokines interleukin-4 (IL-4), interleukin-5 (IL 5), interferon-y (IFN-gamma) and tumor necrosis factor-alpha (TNF-alpha). The accuracy of each of the assays was evaluated in the ranges of 1-15 microg/l (IL 4), 0.001-1 microg/l (IL-5), 0.5-2.5 microg/l (IFN-T) and 0.14-2.2 microg/l (TNF alpha). Other evaluated performance characteristics were the limit of detection (LOD), immunological specificity, and robustness. Traceability was ensured by the use of World Health Organization International Standards (WHO IS). An uncertainty budget, which combined the contribution from all known uncertainty components, was established for each cytokine ELISA. The between-run relative analytical standard deviation (RSDA) of the assessed ELISAs was found to be in the range of 11-18%, except for IL-5 where RSDA increased at decreasing concentrations. The LOD was 0.12 microg/l, 0.0077 microg/l, 0.0069 microg/l and 0.0063 microg/l for IL-4, IL-5, IFN-gamma and TNF-alpha, respectively. Traceability to the WHO IS was established for each of the cytokines. The combined relative standard uncertainty (U(result)/C(result)) was 28%, 22-62%, 28% and 24% for the IL-4, IL-5, IFN-gamma and TNF-alpha results, respectively. The major contributions to uncertainty came from the relative analytical standard deviation and from the uncertainty of the mass concentration of the WHO IS. The uncertainty of the WHO IS was not stated in the accompanying certificate and was evaluated by other means. The largest sources of uncertainty were located outside our laboratory. This means that the possibilities to improve the reliability of the results produced by the ELISAs are very limited. The task of evaluating measurement uncertainty would be much easier if producers of international reference standards reported the uncertainty of the value of standards. The model for evaluating uncertainty presented in this paper is applicable to other types of assays and to most analytical methods. PMID- 12113299 TI - Development of immunoturbidimetric assays for fourteen human serum proteins on the Hitachi 912. AB - Many laboratories rely on dedicated nephelometers or turbidimeters and commercial reagent kits for the evaluation of serum proteins. However, with growing emphasis on cost containment, laboratories are forced to seek additional operational efficiencies by capitalizing on the use of existing analyzers whenever possible. In the present paper we describe the development of immunoturbidimetric assays for routine analysis of 14 human serum proteins (alpha1-antitrypsin, alpha2 macroglobulin, albumin, apolipoproteins Al and B, complement components 3 and 4, haptoglobin, immunoglobulins A, G, and M, orosomucoid, prealbumin, and transferrin) on the Hitachi 912, a general chemistry analyzer. With this system, we obtained excellent precision at levels corresponding to low, normal, and high physiologic concentrations of each protein (within-run imprecision CVs < or = 3.4%, total imprecision CVs < or = 4.1%). Linearity for each method was within 5% of the expected value throughout the calibration range, and method comparisons with either the Roche turbidimetric or Dade Behring nephelometric assays were in good agreement (r >0.97). We observed no significant interference from bilirubin (up to 718 micromol/l), hemoglobin (up to 8 g/l), triglyceride (up to 14.7 mmol/l) or rheumatoid factor (up to 4,140 IU/ml). Calibration for the 14 protein assays was stable for at least 7 days and onboard refrigerated reagents were stable for at least 3 months. The instrument's automated sample re-run feature minimized sample handling and helped to conserve specimens. In conclusion, the newly developed assays on the Hitachi 912 offer high throughput (>250 tests per hour) without the associated cost of a dedicated instrument for protein assays. PMID- 12113300 TI - Multicenter evaluation of a fully mechanized soluble transferrin receptor assay on the Hitachi and cobas integra analyzers. the determination of reference ranges. AB - Soluble transferrin receptor (sTfR) is reported to be a reliable marker for the diagnosis of iron deficiency, especially when iron metabolism is influenced by inflammatory disorders such as infection, chronic inflammation and cancer-related anemia. In the present multicenter study the analytical performance of a recently introduced, latex-enhanced immunoturbidimetric assay for the determination of soluble transferrin receptor (Tina quant [a] sTfR, Roche Diagnostics) on different fully mechanized analyzers such as Hitachi 917 and 911, and Cobas Integra 400 and 700 was evaluated. Within-run and between-run imprecision showed good results (CV<5% and <7%, respectively). The assay was found to be linear over a wide measuring range (0.4-35 mg/l). Endogenous substances did not interfere with the test results. Comparison of serum sTfR concentrations with those of heparinized plasma revealed good correlation (r>0.976). Method comparison with an existing fully mechanized method as well as with ELISA tests for sTfR showed very good correlation (r>0.987). Because of the lack of international standardization the results differed from each other up to 2.5-fold. The 95% of serum levels in healthy individuals ranged from 1.9 to 4.4 mg/l (n=427). However, the reference ranges should be reported in a sex-dependent manner, as 2.2-5.0 mg/l for men (n=211) and as 1.9-4.4 mg/l for premenopausal (n=216) and postmenopausal (n=45) women. The Tina quant [a] sTfR assay enables the precise, accurate, rapid and convenient determination of sTfR concentrations for routine clinical chemistry purposes. PMID- 12113301 TI - Streptococcus iniae infections in Red Sea cage-cultured and wild fishes. AB - Streptococcus iniae was isolated from 2 moribund wild Red Sea fishes, Pomadasys stridens (Pomadasyidae) and Synodus variegatus (Synodontidae), both collected in shallow waters along the Israeli coast of the Gulf of Eilat. The site is approximately 2 km from a mariculture cage farm in which streptococcal infections were diagnosed in previous years in the red drum Sciaenops ocellatus. This is the first report of S. iniae in Red Sea fishes. Biochemical and molecular similarities between the isolates from cultured fishes and those from the wild specimens suggest that a single strain is involved, and that 'amplification' and dispersal of this pathogen from captive to feral fishes have occurred. At the molecular level, the pathogen is different from the S. iniae isolates that have been afflicting the Israeli freshwater aquaculture in recent years. Although S. iniae prevalence in the wild fish populations of the area remains to be determined, the northernmost region of the Gulf of Eilat, virtually landlocked and with generally calm seas and weak currents, seems to be particularly vulnerable to the impact of diseases that develop in this mariculture system. PMID- 12113302 TI - Response of rainbow trout Oncorhynchus mykiss to exposure to Myxobolus cerebralis above and below a point source of infectivity in the upper Colorado River. AB - We exposed 9 wk old rainbow trout Oncorhynchus mykiss to ambient levels of Myxobolus cerebralis infectious stages at 4 sites of suspected differing infectivity in the Colorado River. Exposure was estimated by periodic filtration of river water at each exposure location. After a 32 d exposure, the fish were held in the Colorado River at a common site for over a year. Resulting infection was evaluated by the presence of clinical signs (whirling behavior, cranial deformity/exophthalmia, and black tail), severity of microscopic lesions, and myxospore counts (8, 10, 12, and 14 mo post-exposure). Two exposure sites that were immediately downstream of Windy Gap Reservoir were much higher in infectivity than the site above the reservoir or the site 26 km downstream of the reservoir. Rainbow trout exposed at those locations showed higher prevalence of clinical signs of whirling disease, more severe histological evidence of infection and higher average myxospore concentrations than those exposed above the reservoir or 26 km below the reservoir. Many more M. cerebralis actinospores were observed from water filtration at the 2 sites immediately below the reservoir compared to the other sites. PMID- 12113303 TI - Myxobolus cycloides on the swimbladder of chub Leuciscus cephalus: a controlled, host-specific localisation. AB - Of 150 wild stock chub, Leuciscus cephalus L. captured in Lower Austrian watercourses, 112 revealed disc like plasmodia of Myxobolus cycloides Gurley, 1893 on the caudal chamber of the swim bladder. Other cyprinid species from the same waters lacked M. cycloides or other myxosporeans in this specific localisation. In chub, the intensity of infection (number of discs on the swim bladder) showed a logarithmic, age-dependent increase. The plasmodia of M. cycloides were situated in the connective tissue--mainly along blood vessels--and exhibited a delicate envelope of host tissue, thus forming a characteristic myxosporean cyst. Occasionally single trophozoites seemed to merge. A general process of fibroblast proliferation leading to encapsulation and degradation of the parasite was observed. This process was initiated by the formation of small multiple encapsulations within the spore containing trophozoid, before thickening of the outer cyst wall occurred. The general non-inflammatory course of the M. cycloides infection, and the obvious good health of the investigated chub suggest that this myxosporean in its host specific localisation cannot be regarded as a serious pathogen--on the contrary: parasite multiplication and degradation seemed to occur in a well-defined equilibrium controlled by the host fish. PMID- 12113305 TI - Antiprotozoals effective in vitro against the scuticociliate fish pathogen Philasterides dicentrarchi. AB - The histophagous ciliate Philasterides dicentrarchi causes fatal scuticociliatosis in farmed turbot Scophthalmus maximus and sea bass Dicentrarchus labrax. The present study screened 52 candidate antiprotozoals for activity against this pathogen in vitro. Of these compounds, 14 were effective (i.e. killed all ciliates within a 24 h assay period). In descending order of efficacy (minimum lethal concentration 100 to 0.8 ppm), these were niclosamide, oxyclozanide, bithionol sulfoxide, toltrazuril, N-(2'-hydroxy-5'-chloro-benzoyl) 2-chloro-4-nitroaniline, furaltadone, doxycycline hyclate, formalin, albendazole, carnidazole, pyrimethamine, quinacrine hydrochloride and quinine sulfate. Administration in filtered seawater rather than phosphate-buffered saline inactivated doxycycline hyclate and albendazole, and markedly reduced that of bithionol sulfoxide and toltrazuril, suggesting that these compounds may not be effective in bath administration. In view of these findings, we discuss the potential utility of chemotherapy as a strategy for the control of scuticociliatosis in farmed turbot and sea bass. PMID- 12113304 TI - Validation of a single round polymerase chain reaction assay for identification of Myxobolus cerebralis myxospores. AB - Validation of a single round PCR-based assay to confirm as Myxobolus cerebralis myxospores obtained from pepsin-trypsin digest preparations is described. The assay is a modification of a PCR assay published previously, based on the amplification of a segment of the gene encoding the 18S ribosomal subunit of M. cerebralis. The sensitivity, specificity and upper and lower detection limits were determined using known M. cerebralis and non-M. cerebralis myxospores and M. cerebralis-free fish. The sensitivity of PCR confirmation was 100% (95% confidence interval of 83.2-100%). The specificity was 100% (95% confidence interval of 87.2-100%). The upper detection limit was approximately 100,000 myxospores per reaction; the lower detection limit was approximately 50 myxospores per reaction. Given the high sensitivity and specificity of the assay, substitution of this assay for histologic confirmation of M. cerebralis infection is encouraged. PMID- 12113306 TI - Complete sequence and structure of ribosomal RNA gene of Heterosporis anguillarum. AB - The ribosomal RNA (rRNA) gene region of the microsporidium Heterosporis anguillarum has been examined. Complete DNA sequence data (4060 bp, GenBank Accession No. AF402839) of the rRNA gene of H. anguillarum are presented for the small subunit gene (SSU rRNA: 1359 bp), the internal transcribed spacer (ITS: 37 bp), and the large subunit gene (LSU rRNA: 2664 bp). The secondary structures of the H. anguillarum SSU and LSU rRNA genes are constructed and described. This is the first complete sequence of an rRNA gene published for a fish-infecting microsporidian species. In the phylogenetic analysis, the sequences, including partial SSU rRNA, ITS, and partial LSU rRNA sequences of the fish-infecting microsporidia, were aligned and analysed. The taxonomic position of H. anguillarum as suggested by Lom et al. (2000; Dis Aquat Org 43:225-231) is confirmed in this paper. PMID- 12113307 TI - Ultrastructure of a haplosporidian containing Rickettsiae, associated with mortalities among cultured paua Haliotis iris. AB - Uninucleate and multinucleate stages of a protozoan parasite are described from cultured abalone Haliotis iris Martyn, 1784 in New Zealand. The parasite is identified as a haplosporidian by the occurrence of multinucleate plasmodia, mitochondria with tubular cristae, lipid droplets, anastomosing endoplasmic reticulum (aER), multivesicular bodies (MVBs), haplosporogenesis by the production of haplosporosome-like bodies from nuclear membrane-bound Golgi, and their maturation to haplosporosomes. Coated pits occurred in the plasma membrane and coated vesicles were scattered in the cytoplasm, particularly in association with the Golgi face away from the nucleus, and aER. It is concluded that the outward face of the Golgi may be the trans face, and that aER is the trans-Golgi network. Coated pits and bristle-coated vesicles are reported from a haplosporidian for the first time. The vesicles in the MVBs resembled the cores and inner membranes of haplosporosomes, without the outer layer. The possible inter-relationships of these features are discussed. The abalone parasite differs from previously described haplosporidians in the apparent absence of a persistent mitotic spindle, and the presence of intracytoplasmic coccoid to rod-shaped bacteria resembling Rickettsiales-like prokaryotes. Phylogenetic analysis of the 16S rRNA gene sequence of the Rickettsiales-like prokaryotes indicated that these organisms belong to the Rickettsia cluster. The prokaryotes have a high (7%) sequence divergence from known Rickettsieae, with Rickettsia sp. and R. massiliae being the closest relatives. The lack of non-molecular evidence prevents us from proposing a new rickettsial genus at this time. PMID- 12113308 TI - Comparative susceptibility of veliger larvae of four bivalve mollusks to a Vibrio alginolyticus strain. AB - The susceptibility of 7 d old veliger larvae of the scallops Argopecten ventricosus and Nodipecten subnodosus, the penshell Atrina maura, and the Pacific oyster Crassostrea gigas to a pathogenic strain of Vibrio alginolyticus was investigated by challenging the larvae with different bacterial concentrations in a semi-static assay. The results indicate that the larvae of the 2 scallop species are more susceptible to the V. alginolyticus strain than those of the oyster and the penshell. Signs of the disease were similar to bacillary necrosis described in previous work. Interspecies differences in susceptibility to pathogens are discussed. PMID- 12113309 TI - Morphological and cytoenzymatic characterization of haemocytes of the venus clam Chamelea gallina. AB - A morphological and enzymatic characterization of Chamelea gallina haemocytes was carried out as a prerequisite for further studies on venus clam immunobiology. Two main types of circulating haemocytes were identified (1) hyalinocytes (79.2%), agranular cells with a central nucleus surrounded by a little cytoplasm, and (2) granulocytes (16.5%), smaller granular cells with smaller nuclei. Small cells with a strongly basophilic nucleus and a thin layer of peripheral cytoplasm, probably undifferentiated blast cells (4.3%), were also observed. Both granulocytes and hyalinocytes can assume a spreading or round morphology. The enzymatic activities of haemocytes were also investigated. Some of the granulocytes and hyalinocytes were positive for hydrolytic enzymes, suggesting a role for these cells in phagocytosis; no oxidative enzymes were detected in C. gallina haemocytes. Granulocytes and hyalinocytes can easily adhere to the substratum and exhibit a low phagocytosis activity towards foreign particles (6.3%), whereas the fraction of cells containing ingested material significantly increased after pre-incubation of test particles with cell-free haemolymph, which suggests the presence of opsonin(s) in the haemolymph. PMID- 12113310 TI - Estimation of prevalence of white spot syndrome virus (WSSV) by polymerase chain reaction in Penaeus monodon postlarvae at time of stocking in shrimp farms of Karnataka, India: a population-based study. AB - White spot disease (WSD) is at present the most serious viral disease affecting cultivated shrimp species globally. The causative agent, white spot syndrome virus (WSSV), is extremely virulent, has a wide host range and can also be transmitted from broodstock to their offspring. The shrimp postlarvae (PL) act as asymptomatic, latent carriers of the virus, and stocking of WSSV-infected PL has been reported as a risk factor for WSD outbreaks in culture ponds. However, there is no population-based study on WSSV prevalence in PL of shrimp. The present manuscript documents the approaches and the results in the estimation of prevalence of WSSV in PL populations of Penaeus monodon at the time of stocking. A maximum of 300 PL from each of the 73 batches of PL stocked at various farms in the west coast of India during September 1999 to January 2000 were tested for the presence of WSSV by 2-step nested PCR. Thirty-six (49%) of the 73 batches tested positive for WSSV either by 1-step alone (3 batches) or after 2-step nested PCR (33 batches). Sub-samples of 5 PL each or 1 PL each tested to quantify the proportion of infected PL within batches showed that WSSV prevalence was very high in 1-step PCR-positive batches and low in 2-step PCR-positive batches. The study also showed that appropriate sampling and sample size were major factors in determining the prevalence of WSSV in PL populations, underlining the need for testing large samples of PL to reduce errors from falsely negative results. PMID- 12113312 TI - Suprapubic aspiration of urine in the diagnosis of urinary tract infection in infants. PMID- 12113311 TI - Unexplained infant crying: an evolutionary perspective. AB - The absence of adverse health outcomes later in development and the similarity of defining features of "colic" across cultures suggest that an evolutionary perspective may give us some insight into the nature of this puzzling condition. Evidence suggests that the larynx evolved first as a protective valve and later as a means to stabilize the upper thorax momentarily for precision upper arm movements. Recently, we found another physiological role for the larynx in regulating respiratory function to promote the recovery of young rats from severe hypothermia. In the process, bursts of calling were emitted by unconscious pups, which were nevertheless effective in eliciting maternal search and retrieval. These unexpected findings reveal how infant calling may have evolved as a communicative signal derived from more primitive physiological functions of the larynx. Repetitive calling in the normal young rat when isolated from its littermates and mother is regulated by multiple sensory cues present in the infant's social interactions, and in the paper it is described how this sensory input projects to central neuromodulatory systems known to be active in the control of anxiety behaviors in adult rats and humans. CONCLUSION: This broad range of functions for infant calling in other mammals suggests several new ways to approach the further study and treatment of unexplained crying in human infants. PMID- 12113313 TI - Coronary artery size in Kawasaki disease: echocardiographic definition. PMID- 12113314 TI - Epinephrine treatment in hypotensive newborns. PMID- 12113315 TI - Beyond education and food: psychosocial well-being of orphans in Africa. PMID- 12113316 TI - Home visitation: from sanitary control to support of the young family. PMID- 12113317 TI - Plasma total homocysteine increases from day 20 to 40 in breastfed but not formula-fed low-birthweight infants. AB - Homocysteine is an intermediate in the folate cycle and methionine metabolism. This study investigated whether formula-fed infants have different plasma total homocysteine to their breastfed counterparts, and during what period any difference developed. Plasma total homocysteine was determined in 53 formula-fed and 15 breastfed healthy low-birthweight babies (< or = 2500 g) around days 10, 20 and 40. Total homocysteine was also measured in human milk. Mean +/- SD plasma total homocysteine levels (micromol l(-1)) at days 10, 20 and 40 were 6.4 +/- 2.6, 6.7 +/- 2.4 and 9.1 +/- 2.4 (breastfed), and 7.5 +/- 3.2, 7.3 +/- 2.1 and 7.4 +/- 1.6 (formula-fed). Homocysteine of breastfed babies at day 40 was higher than that of breastfed babies at day 20 (p < 0.0001), and that of formula-fed counterparts at day 40 (p = 0.002). Homocysteine correlated negatively with formula (day 10) and breast milk (day 40) volume intakes. Median (range) homocysteine in 12 mature human milk samples was 0.30 (not detectable to 0.7) micromol l(-1). CONCLUSION: Increasing plasma total homocysteine in breastfed babies to higher levels compared with formula-fed babies may be caused by a gradually developing suboptimal B-vitamin status in lactating women. PMID- 12113318 TI - Comparing suprapubic urine aspiration under real-time ultrasound guidance with conventional blind aspiration. AB - To determine the optimal method of suprapubic aspiration (SPA), the success rates of real-time ultrasound-guided SPA were compared with those of conventional SPA, and factors associated with success were studied. Thirty infants were randomly allocated to group A (for real-time ultrasound-guided SPA) and 30 infants to group B (for blind SPA with a prehydration protocol). The results showed that the overall success rates for all attempts were similar (26/30 or 87% in group A vs 24/30 or 80% in group B, p > 0.05). The first attempts in both groups were equally successful (both 18/30 or 60%). In comparison with failed attempts, successful ultrasound SPA attempts were associated with a greater bladder depth (mean +/- SD: 28 +/- 11 vs 21 +/- 5 mm, p < 0.01), length (32 +/- 12 vs 23 +/- 9 mm, p < 0.05) and volume (17 +/- 13 vs 8 +/- 6 ml, p < 0.01), but similar width (33 +/- 9 vs 29 +/- 5 mm, p > 0.05). In blind SPA, successful attempts were associated with the presence of bladder dullness on percussion (odds ratio 29). CONCLUSION: This study confirms that ultrasound-guided SPA has a high success rate. Blind SPA could also be equally successful with appropriate preparation. Ultrasound-guided SPA is recommended when the bladder depth exceeds 3 cm, or the bladder length exceeds 3.7 cm. If an ultrasound machine is not available, blind SPA may be an alternative, with attention being paid to prehydration and the demonstration of bladder dullness by percussion. PMID- 12113319 TI - Factors relating to the cardiac sequelae of Kawasaki disease one month after initial onset. AB - This study aimed to determine the risk factors related to the presence of cardiac sequelae 1 mo after initial onset and to examine the preventive effect of the early administration of high-dose gamma-globulin (GG) on cardiac sequelae in patients with Kawasaki disease. Patients treated with high-dose GG of 2000 +/- 100 mg kg(-1) were selected as subjects from the 15th nation-wide survey in Japan. Univariate and logistic multiple variable analyses were used to test the effects of background variables such as age and gender, variables relating to laboratory findings such as the percentage of neutrophil leucocytes, and variables relating to the GG treatment on the presence of cardiac sequelae. The odds ratios were significantly higher for males (1.48), those younger than 1 y of age (1.71), recurrent cases (2.42), and those with a low haematocrit (<32.5%) (1.45) and high percentage of neutrophil leucocytes (>68%) (1.63). The odds ratio was low for those who started GG administration in less than 6 d from onset between the patients with and without cardiac sequelae. The odds ratio for the duration of GG treatment was not significantly different between those with and without cardiac sequelae. CONCLUSION: Patients who received early administration of GG, less than 6 d from onset of the disease, had a lower risk than those received GG more than 6 d from the onset. The percentage of neutrophil leucocytes and the haematocrit level are useful indicators in predicting the development of cardiac sequelae. PMID- 12113320 TI - Preschool children with developmental coordination disorder: a short-term follow up of motor status at seven to eight years of age. AB - At a population-based screening of 5-6-y-old children, 37 children were identified as having definite or borderline motor difficulties consistent with developmental coordination disorder. When 7-8 y old they were re-examined to determine their current motor status and to compare it with that at the age of 5 6 y. The motor status was investigated from three perspectives: by a motor test, by the use of a self-perceived motor competence scale, and from the parents' descriptions. On a group level the children had not changed their motor status when 7-8 y old, according to the total scores in the Movement ABC motor test and the Perceived Motor Competence scale. On an individual level most children with definite motor difficulties remained in their category regarding motor difficulties at the follow-up, while most children with borderline motor difficulties did not. The parents' descriptions indicated an association between willingness to engage in physical activities and degree of motor difficulties. CONCLUSION: Over a period of 1.5 y most children with definite motor difficulties continued to have such difficulties. The parents' descriptions of their children's motor status and development were in agreement with the children's motor status as measured by a motor test, but were not in total accordance with the children's self-perceived competence. PMID- 12113321 TI - Comorbidity in children with severe developmental language disability. AB - In a cohort of 2359 children, screened for severe developmental language disability (DLD) at 3 y of age, 45 children were identified as true positives. The development, concerning DLD and comorbidity of 41 of these children still living in the municipality of Uppsala was followed up to school start. Criteria for comorbidity were: (a) suspected or diagnosed neuropsychiatric/neurodevelopmental disability according to information from the Child Habilitation Centre and the Child Psychiatric Centre or (b) low performance IQ, signs of activity/distractibility problems according to a psychologist's examination. By school start, 61% of the children with severe DLD were identified with comorbidity. CONCLUSION: Severe DLD is often combined with other disabilities within the neuropsychiatric/neurodevelopmental spectra. The comorbidity might not be obvious at 3 y of age--the age at which severe DLD is effectively identified by the 3-y screening programme. This in turn stresses the necessity of multidisciplinary teamwork both at the referral level and during the therapy work. PMID- 12113322 TI - Screening for fragile X syndrome: results from a school for mentally retarded children. AB - Fragile X syndrome is the most common inherited form of familial mental retardation. The purpose of this study was to identify yet unrecognized fragile X individuals and to estimate the frequency of both the FRAXA and FRAXE forms of the disease in a population of mentally retarded children attending a special school in Croatia. The results are reported of molecular screening of 114 children with mild to severe mental retardation. Three individuals (2.6%) with the FRAXA form of the fragile X syndrome and one boy (0.9%) with FRAXE mental retardation were detected; a total of four newly diagnosed fragile X families were identified. Closer clinical examination revealed that behavioural and speech disturbances were clearly present among all fragile X cases (both FRAXA and FRAXE), indicating that these features could be additional diagnostic criteria for the preselection of individuals at risk. CONCLUSION: Fragile X screening among mentally retarded children attending a special school should be highly encouraged to reveal the cause of mental retardation and to detect yet unrecognized fragile X individuals. The frequency of fragile X syndrome in a such population in Croatia was found to correlate with similar results from previous studies. However, since at the time of diagnosis all affected families had a second or even a third child born, earlier diagnosis should be considered to provide greater benefit to fragile X families. PMID- 12113323 TI - Multiple doses of secretin in the treatment of autism: a controlled study. AB - Dramatic effects on autistic behaviour after repeated injections of the gastrointestinal hormone secretin have been referred in a number of case reports. In the absence of curative and effective treatments for this disabling condition, this information has created new hope among parents. Although controlled studies on the effect of mainly one single dose have not documented any effect, many children still continue to receive secretin. Six children enrolled in a double blind, placebo-controlled crossover study in which each child was its own control. Human synthetic secretin, mean dose 3.4 clinical units, and placebo were administered intravenously in randomized order every 4th wk, on three occasions each. The measurement instruments were the visual analogue scale (VAS) and the aberrant behaviour checklist (ABC). Statistically significant differences were found for placebo in 3 out of 6 children and for secretin in one child, using parental ratings only (VAS scores). Differences were small and lacked clinical significance, which was in accordance with the overall impression of the parents and teachers and visual inspection of graphs. CONCLUSION: In this placebo controlled study, multiple doses of secretin did not produce any symptomatic improvement. PMID- 12113324 TI - Impact of topical oils on the skin barrier: possible implications for neonatal health in developing countries. AB - Topical therapy to enhance skin barrier function may be a simple, low-cost, effective strategy to improve outcome of preterm infants with a developmentally compromised epidermal barrier, as lipid constituents of topical products may act as a mechanical barrier and augment synthesis of barrier lipids. Natural oils are applied topically as part of a traditional oil massage to neonates in many developing countries. We sought to identify inexpensive, safe, vegetable oils available in developing countries that improved epidermal barrier function. The impact of oils on mouse epidermal barrier function (rate of transepidermal water loss over time following acute barrier disruption by tape-stripping) and ultrastructure was determined. A single application of sunflower seed oil significantly accelerated skin barrier recovery within 1 h; the effect was sustained 5 h after application. In contrast, the other vegetable oils tested (mustard, olive and soybean oils) all significantly delayed recovery of barrier function compared with control- or Aquaphor-treated skin. Twice-daily applications of mustard oil for 7 d resulted in sustained delay of barrier recovery. Moreover, adverse ultrastructural changes were seen under transmission electron microscopy in keratin intermediate filament, mitochondrial, nuclear, and nuclear envelope structure following a single application of mustard oil. CONCLUSION: Our data suggest that topical application of linoleate-enriched oil such as sunflower seed oil might enhance skin barrier function and improve outcome in neonates with compromised barrier function. Mustard oil, used routinely in newborn care throughout South Asia, has toxic effects on the epidermal barrier that warrant further investigation. PMID- 12113325 TI - Food intake and oral sucrose in preterms prior to heel prick. AB - To investigate the soothing effect of feeding on infants in distress, the effects of 2 mL 15% and 1 mL 25% sucrose given orally 2 min before heel prick in fasting preterms to reduce the pain response were assessed. The effects of milk intake by nasogastric tube were also assessed once during the last hour before heel prick, and the effects of milk intake by nasogastric tube once during the last hour before heel prick together with 1 mL 25% sucrose given orally 2 min before heel prick. The pain response was measured as changes in crying time, behavioural state, skin conductance and heart rate. Each group included 12 healthy preterm infants with a median gestational age of 32 wk and a median postnatal age of 14 d. These infants were randomly studied twice; one in connection with the intervention and once after being given sterilized water. Differences in the measured variables before and during heel prick showed that only the crying time was reduced when the infants received milk or 25% sucrose prior to heel prick (p < 0.05). If the infants received milk and 25% sucrose before heel prick, the crying time and the level of behavioural state were reduced (p < 0.05). The increase from before to during heel prick in skin conductance (number and amplitude of the waves) and heart rate correlated with the crying time (p < 0.01). PMID- 12113326 TI - Postnatal urinary conductance measurement in preterm infants. AB - The aim of this study was to determine whether serial urinary conductance measurements can be used to estimate reliably the end of the transition period of negative sodium balance in preterm infants. The relationship between urine conductance, measured by a conductance meter, and urine sodium concentration was determined in 109 pooled samples of urine obtained from 14 preterm infants during the transitional period of fluid balance. It was shown by linear regression analysis that urine sodium concentration (mmol l(-1)) = 0.78 x urine conductance 1.25. Urine sodium concentrations derived from the above formula were concordant with urine sodium measured directly when used to calculate daily sodium balance in all 14 infants. CONCLUSION: Urine conductance can be accurately measured at the cotside by neonatal nurses and used to identify the timing of the postnatal transition from negative to positive sodium balance in preterm infants. These findings can help in making decisions on the introduction of postnatal sodium administration to preterm infants. PMID- 12113327 TI - Epinephrine treatment of hypotension in very low birthweight infants. AB - The aim of this study was to examine the influence of a continuous infusion of epinephrine (adrenaline) on mean arterial blood pressure (MABP), heart rate, urine output and base deficit in very low birthweight infants (VLBWI) with systemic hypotension. In VLBWI who received an infusion of epinephrine for at least 12 h the mean urine output, administered fluid volume, base deficit and administered buffer 12 h before and 12 h during the infusion were recorded. If the infusion was shorter, but given for at least 2 h, the mean heart rate and MABP 2 h before and 2 h during the infusion were recorded. Thirty-one infants with a gestational age of 26 (23-30) wk [median (minimum-maximum)] and birthweight 690 (390-1310) g were included in this retrospective chart review. The patients received an infusion of epinephrine at a postnatal age of 3 (1-21) d. The doses ranged between 0.05 and 2.6 microg kg(-1) per minute within the first 24 h of administration. Three of 31 infants received epinephrine on 2 different occasions. The MABP [+7 (-1 to 13) mmHg, p=0.000001] and the heart rate [+10 (-10 to 42) bpm, p=0.000036] increased significantly (n = 34), whereas total volume administration and urine output remained the same between the 2 periods (Wilcoxon matched pairs test). The base deficit increased significantly [-3 ( 10.2 to 2.6), p = 0.0014, n = 19] without a change in the administration of buffer. CONCLUSION: The infusion of epinephrine increased the MABP and the heart rate without decreasing urine output in VLBWI with hypotension not responding to a dopamine infusion up to 15 microg kg(-1) per minute. A potential adverse effect was an increase in metabolic acidosis. PMID- 12113328 TI - First-time mothers' satisfaction with early encounters with the nurse in child healthcare: home visit or visit to the clinic? AB - The aim of this study was to describe first-time mothers' views of satisfaction with their first encounter with the nurse, in order to investigate differences between home visits and clinic visits and between high/middle and low socioeconomic classification (SEC). A nation-wide postal questionnaire sent to 800 first-time mothers yielded the data for statistical analysis. Data were collected using a modified version of the questionnaire "Quality of Care from the Patient's Perspective", the part concerning child healthcare. The results showed that mothers who had received home visits were more content with the encounter than were mothers who had to visit the clinic. This particularly concerned advice on breastfeeding, being able to talk to the nurse in peace and quiet, and the fact that the nurse took time and was personal. In contrast, the mothers who had received a home visit were less content with the competence of the nurse when she examined the child. Mothers of low SEC were less satisfied with the first encounter than were mothers of high/middle SEC with regard to several points. CONCLUSION: Home visits were shown to have advantages over visits to the clinic. Mothers of low SEC were less satisfied with the first encounter with the nurse than were mothers in the high/middle SEC. PMID- 12113329 TI - Breastfeeding, baby friendliness and birth in transition in North Western Russia: a study of women's perceptions of the care they receive when giving birth in six maternity homes in the cities of Archangelsk and Murmansk, 1999. AB - Women's own views on the quality of the birthing care they received were recorded in a small study in the cities of Archangelsk and Murmansk in February 1999. Six maternity wards took part; one hospital had already been designated as a Baby Friendly Hospital (BFH) according to the strict global criteria of the WHO/UNICEF recommended Baby Friendly Hospital Initiative (BFHI). Two of the hospitals had made profound changes in feeding routines and were by their own reckoning close to achieving this distinction, and were included in the BFH group. Three maternity wards were far from being in compliance with the BFHI approach and were grouped as the Non-Baby-Friendly Hospitals (NBFH). A total of 180 newly delivered mothers answered a 60-item questionnaire about their birthing and breastfeeding experiences. The questions were chosen from an existing protocol, the WEB (Women's Experiences of Birth) developed by one of the authors (BC). The study was part of an informal evaluation of five years of BFHI activities in the Barents Region, supported by Norway, and also aimed at recording any positive carry-over effect of the BFHI into obstetric routines as a whole. CONCLUSION: It was found that the project definitely had had an impact; feeding practices at the BFH were markedly closer to the international BFHI recommendations than at the NBFH. BFH mothers, however, reported suffering from breastfeeding problems just as often or more so than NBFH mothers. Possible explanations are discussed; it is concluded that this cross-sectional study may depict a transitory situation in the BFH. At the two hospitals not yet assessed, although staff felt that they had made profound changes, they may not yet have grasped the full extent and stringency of the changes required. The study shows that, despite good will, some practical details had not yet been worked out, resulting in a mixed outcome for the mothers. There was no noticeable carry-over of the attitudes and basic ideas of the project into obstetric care, either in the BFH or in the NBFH, so changes in this area may require separate strategies. PMID- 12113330 TI - Severe sleep problems in infancy associated with subsequent development of attention-deficit/hyperactivity disorder at 5.5 years of age. AB - The aim of this five-year prospective study was to follow and compare a group of children with severe sleep problems in infancy with a control group regarding development of symptoms of attention-deficit/hyperactivity disorder (ADHD). Factors in infancy were sought which were associated with development of ADHD in later childhood. A total community sample of 2518 infants aged 6-18 mo was approached with a questionnaire, and 83% responded. Data from the collection procedure point to a non-selective dropout. A case group of those children aged 6 12 mo who fulfilled specific criteria for severe and chronic sleep problems (n = 27) was compared with a control group of equal size, matched for age and gender. At the age of 5.5 y, seven of the children in the sleep problem group met the criteria for the diagnosis of ADHD based on an in-depth assessment by a multidisciplinary team. None of the control children qualified for the diagnosis. This difference is statistically significant. Comparisons between the children with ADHD and the rest of the problem group showed that given severe infant sleep problems, the following characteristics in infancy were associated with subsequent diagnosis of ADHD: psychosocial problems in the family, bedtime struggles and long sleep latency at bedtime. Owing to the small sample size, the results are considered preliminary. CONCLUSION: Approximately one in four children with severe sleep problems in infancy will later qualify for the diagnosis of ADHD. Infants with severe sleep problems, especially in combination with behavioural problems, high activity level and psychosocial problems in the family, deserve attention since neurodevelopmental problems seem to be prevalent in this category of children. PMID- 12113331 TI - Population-based rates of severe respiratory syncytial virus infection in children with and without risk factors, and outcome in a tertiary care setting. AB - The aim of this study was to make a population-based estimate of the risk of hospitalization and complications during virologically confirmed respiratory syncytial virus (RSV) infection in relation to established risk factors, and an estimation of additional risk factors and outcome as seen in a tertiary care referral centre. During a period of 12 y, all children with virologically confirmed RSV infection were included. Recorded complications were: admission to the intensive care unit, mechanical ventilation, death and later hospitalization for wheezing. In total, 1503 cases were identified, 1354 of which originated from the population defined by the catchment area. There was a biannual seasonal variation with late small outbreaks alternating with early large ones. The hospitalization rates for infants without risk factors were 0.8 and 1.4% during the 2 epidemic types. They were 1.6-3.2% for infants born preterm (<33 gestational wk), 2.9-7.0% for children under 2 y old with chronic lung disease of prematurity and 2.8-6.4% for infants with congenital heart disease. The presence of siblings in the family more than doubled the risk of hospitalization. Later hospitalization for wheezing occurred in 8.4 and 4.9% of children without risk factors over and under the age of 2 mo, respectively (p < 0.001). CONCLUSION: This study found lower population rates of hospitalization and complications than have previously been reported. The seasonal variation and the presence of siblings in the home influenced these rates by factors of 2. PMID- 12113333 TI - European Society for Neonatology: training program for the neonatologist in Europe. PMID- 12113332 TI - Vaccination coverage estimates by EPI cluster sampling survey of children (18-24 months) in Flanders, Belgium. AB - A random cluster sample according to the EPI cluster sampling technique was conducted in 1999 in Flanders (North Belgium) to ascertain the vaccination coverage of 18 to 24-mo-old children. Polio is the only mandatory vaccine. Diphtheria-tetanus-pertussis (DTP), Haemophilus influenzae type b (Hib), hepatitis B (HB) and measles-mumps-rubella (MMR) are included in the recommended schedule of vaccinations. For Hib and HB, a minimal cost was charged. Professional interviewers conducted interviews with the parents of 1110 children randomly selected in 89 municipalities. Analysis was conducted on the results of 1005 children. The coverage level (95% confidence interval) for the full schedule was 96% (95-97) for polio, 89% (87-91) for DTP, 78% (74-82) for Hib, 68% (64-72) for HB and 83% (81-87) for MMR. The vaccinations were administered by the regional children's health organization (70%), paediatricians (17%) and GPs (11%). No sociodemographic factors could be associated with vaccination coverage. One province showed significantly (p < 0.01) lower vaccination coverage levels compared with those of the other four provinces for DTP (91% vs 82%), Hib (78% vs 53%), HB (73% vs 46%) and MMR (87% vs 66%). CONCLUSION: There is a need for more and better information about vaccination for parents as well as for the healthcare providers. PMID- 12113334 TI - Low colonic obstruction due to Opuntia ficus indica seeds: the aftermath of enjoying delicious cactus fruits. PMID- 12113335 TI - Intra-lesional coagulation in haemangiomas. PMID- 12113336 TI - Adsorption equilibria of butyl- and amylbenzene on monolithic silica-based columns. AB - The adsorption isotherms of butyl- and amylbenzene on silica monolithic columns were measured by frontal analysis. The external, internal and total porosities of these columns were determined by inverse size-exclusion chromatography. The adsorption isotherms are concave upward in the entire concentration range investigated. They were fitted to the anti-Langmuir model, an unusual model in liquid-solid and liquid-liquid phase equilibria. Band profiles under overloaded conditions were recorded. They were in good agreement with the profiles calculated using th,e lumped pore diffusion model of chromatography and these adsorption isotherms. PMID- 12113337 TI - Evaluation of size-exclusion chromatography and size-exclusion electrochromatography calibration curves. AB - Size-exclusion chromatography (SEC) and size-exclusion electrochromatography (SEEC) are chromatographic techniques used to determine molecular mass (weight) distributions (MWD) of polymers. One important step in the data treatment to derive MWD parameters is the modelling of the calibration curves. The calibration curves applied in SEC and SEEC are generally not linear. In this study the modelling of calibration curves is being examined. Different polynomial models have been evaluated and compared, not only for model fit but also for their predictive properties. It was found that sometimes a straight line and sometimes a third-order polynomial model were best. The best model across the effective range (also called linear range) is not always found to be a straight line. The SEEC curves were found to have considerably higher prediction errors than the SEC ones. Reduction of the number of calibration standards to five or six did not greatly affect the predictive properties of the calibration curves, neither in SEC nor in SEEC. PMID- 12113338 TI - Determining the molar mass of a plasma substitute succinylated gelatin by size exclusion chromatography-multi-angle laser light scattering, sedimentation equilibrium and conventional size exclusion chromatography. AB - The clinical effectiveness of succinylated gelatin as a plasma substitute depends strongly on its molar mass, determined conventionally by size exclusion chromatography (SEC). This study evaluates different SEC calibration standards in comparison with two independent "absolute" methods for determining the weight average molar mass (M(w)) of a succinylated gelatin sample. SEC calibrated using succinylated gelatin fractions correlated well with size exclusion chromatography multi-angle laser light scattering (SEC-MALLS) and sedimentation equilibrium whereas SEC calibrated with unmodified gelatin, sodium polystyrene sulfonates or pullulans overestimated M(w) by over 20%. Universal calibration was equivocal. The problems associated with the preparation of succinylated gelatin fractions suggest that an absolute method such as SEC-MALLS may be a more suitable choice for determining the M(w) in succinylated gelatins. PMID- 12113340 TI - On-line pH modification of carbonate eluents using an electrolytic potassium hydroxide generator for ion chromatography. AB - Classical gradient elution, based on the application of a gradient pump used for mixing two or more prepared eluent components in pre-determined concentrations, was replaced by a chromatography system equipped with an isocratic pump and an electrolytic KOH generator. The isocratic pump delivered a constant concentration eluent composed of pure hydrogencarbonate solution. Carbonate ions, the main component of carbonate/hydrogencarbonate-based eluents, were formed by titration of hydrogencarbonate with KOH formed on-line in the electrolytic KOH generator. By changing the concentration of electrolytically-generated KOH, the eluent composition could be changed from pure hydrogencarbonate to a carbonate/hydrogencarbonate buffer, and finally to a carbonate/hydroxide-based eluent. The described system was tested to achieve pH-based changes of retention behavior of phosphate under constant inflow eluent composition conditions. PMID- 12113339 TI - Characterization and comparison of the chromatographic performance of conventional, polar-embedded, and polar-endcapped reversed-phase liquid chromatography stationary phases. AB - We have evaluated and compared the performance of several conventional C18 phases with those possessing either a polar-endcapping group or a polar-embedded group within the primary alkyl ligand and found distinct differences in the chromatographic behavior among the three groups, as well as a high degree of variability within each group. The trend is for the polar-endcapped phases to display similar hydrophobic retention characteristics as the conventional C18 columns, but to express higher hydrogen bonding capacities and silanol activity. The polar-embedded phases displayed the opposite behavior, with a greatly reduced hydrophobic nature compared to the conventional and polar-endcapped C18 phases, and also a very much reduced silanol activity. Most interestingly, it appears that ionic or dipole interactions play a significant role in the overall retention behavior of the polar-embedded phases towards basic and acidic analytes. PMID- 12113341 TI - Ion-pair reversed-phase liquid chromatography-electrospray mass spectrometry for the analysis of underivatized small peptides. AB - The single run analysis of 23 small peptides (principally glycyl and lysyl dipeptides) is performed by ion-pair reversed-phase liquid chromatography coupled with evaporative light scattering detection or electrospray (tandem) mass spectrometry. Several perfluorinated carboxylic acid homologues are evaluated with an octadecyl silica stationary phase (Supelcosil ABZ+ Plus). Among the perfluorocarboxylic acids tested the nonafluoropentanoic acid and the tridecafluoroheptanoic acid gave the best results. Special attention was paid to the separation of isomer/isobar dipeptides (e.g., Gly-Ile, Gly-Leu, Leu-Gly, as well as Gly-Gln, Gly-Lys, etc.) which is usually necessary in spite of the high specificity of mass spectrometry. Before LC-MS analysis, ion-spray fragmentation as well as optimization of MS parameters of the analysed peptides was investigated. The optimum collision energy of glysyl peptides, Ala-Gln, Asp-Asp and Asp-Asp-Asp (13-18 eV) was different from that of the lysyl peptides, Tyr-Glu and oxidised glutathione (25-32 eV). Limits of detection varied from 0.1 to 1.2 mg l(-1) for simple MS and 0.05 to 25 mg l(-1) for tandem MS. PMID- 12113342 TI - Effect of the mobile phase composition on the separation and detection of intact proteins by reversed-phase liquid chromatography-electrospray mass spectrometry. AB - Various buffers (ammonium acetate, ammonium formate, and ammonium hydrogencarbonate), acids (formic acid, acetic acid, heptafluorobutyric acid, and trifluoroacetic acid), and bases (ammonium hydroxide and morpholine) covering the range from 2 to 11.5 have been investigated for their performance in the separation of proteins by reversed-phase liquid chromatography (RPLC) and in their detection by electrospray mass spectrometry (ESI-MS). These additives were first tested for the detection of standard proteins by ESI-MS by flow-injection analysis (FIA). Those additives yielding the highest signals were employed for the separation of standard proteins by using three different reversed-phase columns: two C18 columns (4.6 mm I.D. and 2.1 mm I.D.) and one perfusion column (2 mm I.D.). The sensitivity of the LC-MS system was evaluated with the column giving the best results and with those LC eluents enabling the LC separation of the proteins and also yielding the highest MS signals. For that purpose, calibration curves were compared for both LC-MS and FIA-MS. Formic acid was the additive yielding the highest responses in FIA-MS and trifluoroacetic acid (TFA) gave the best separation and recovery of the proteins. However, problems related to poor recovery of the proteins in the column when formic acid was used and the significant signal suppression observed in MS when TFA was employed, made neither of them suitable for the sensitive detection of the proteins in LC-MS. PMID- 12113343 TI - Determination of pesticide residues in coconut water by liquid-liquid extraction and gas chromatography with electron-capture plus thermionic specific detection and solid-phase extraction and high-performance liquid chromatography with ultraviolet detection. AB - Two simple methods were developed to determine 11 pesticides in coconut water, a natural isotonic drink rich in salts, sugars and vitamins consumed by the people and athletes. The first procedure involves solid-phase extraction using Sep-Pak Vac C18 disposable cartridges with methanol for elution. Isocratic analysis was carried out by means of high-performance liquid chromatography with ultraviolet detection at 254 nm to analyse captan, chlorothalonil, carbendazim, lufenuron and diafenthiuron. The other procedure is based on liquid-liquid extraction with hexane-dichloromethane (1:1, v/v), followed by gas chromatographic analysis with effluent splitting to electron-capture detection for determination of endosulfan, captan, tetradifon and trichlorfon and thermionic specific detection for determination of malathion, parathion-methyl and monocrotophos. The methods were validated with fortified samples at different concentration levels (0.01-12.0 mg/kg). Average recoveries ranged from 75 to 104% with relative standard deviations between 1.4 and 11.5%. Each recovery analysis was repeated at least five times. Limits of detection ranged from 0.002 to 2.0 mg/kg. The analytical procedures were applied to 15 samples and no detectable amounts of the pesticides were found in any samples under the conditions described. PMID- 12113344 TI - Experimental investigation of the behavior of gas phase simulated moving beds. AB - The preparative, continuous gas chromatographic separation of the enantiomers of the inhalation anaesthetic enflurane has been studied on a chiral stationary phase based on octakis(3-O-butanoyl-2,6-di-O-n-pentyl)-gamma-cyclodextrin, dissolved in polysiloxane SE-54 and coated on Chromosorb particles. This has been carried out in a gas chromatographic simulated moving bed (GC-SMB) unit, equipped with eight columns, yielding a total volume of 1.128 l. With reference to this particular system, the separation performance and the behavior of GC-SMB units have been analyzed in depth. We have varied the internal flow-rates in the four sections of the unit in the range between about 1 and about 5 std l/min, the temperature between 20 and 45 degrees C, the switch time between about 2 and about 14 min, and the feed concentration in the range 0.32-0.90 mol%. Similarities and differences in the behavior of GC-SMBs as compared to conventional liquid phase SMBs have been described, and discussed. Operating conditions leading to more than 99% purity in one or both outlet stream have been identified, together with those achieving optimal throughput. Under such optimal conditions, about 20 g of each enflurane enantiomer with enantiomeric purity larger than 98% have been prepared. PMID- 12113345 TI - Dynamic sonication-assisted solvent extraction of organophosphate esters in air samples. AB - A new system for extracting solid samples based on dynamic sonication-assisted solvent extraction (DSASE) is described. The technique is highly efficient with respect to both time and solvent consumption. In tests reported here, organophosphate esters were extracted from air sampling filters in 3 min with an extraction volume of 600 microL of solvent. Furthermore, it was possible to replace a previously used chlorinated solvent with a halogen-free solvent mixture. The sample was placed in a cartridge through which fresh solvent was pumped continuously. A restrictor connected to the outlet of the cartridge allowed the system to be used at a temperature of 70 degrees C without reaching the boiling point of the solvent. Both spiked and non-spiked native samples were used for the evaluation, which clearly revealed a stronger analyte-matrix interaction in native samples. The DSASE technique was shown to recover larger amounts of organophosphate esters from native samples, compared to a static method. DSASE was applied to air samples collected in a lecture hall and from above a computer monitor. PMID- 12113346 TI - Steric mass-action model for dye-ligand affinity adsorption of protein. AB - The steric mass-action (SMA) model has been reported in the literature for ion exchange and metal-affinity interaction adsorption equilibrium of proteins. In this work, an SMA model was developed for protein adsorption equilibrium to dye ligand affinity adsorbent, Cibacron Blue-modified Sepharose CL-6B (CB-Sepharose). Static adsorption experiments with bovine serum albumin as a model protein were carried out to determine the model parameters, that is, equilibrium constant (K), characteristic number of binding sites (n), and steric factor (sigma). It was found that the linear model parameters, K and n decreased with the increase of ionic strength, while the nonlinear parameter, sigma, increased with ionic strength and the dye-ligand concentration. Thus, expressions correlating these parameters with the dye-ligand concentration and/or ionic strength were derived. With these correlations, the SMA model gave promising results in predicting protein adsorption isotherms. Thus, it is considered that the model would be useful in the theoretical analysis of dye-ligand affinity chromatography. PMID- 12113347 TI - Non-linear calibration functions in ion chromatography with suppressed conductivity detection using hydroxide eluents. AB - The linearity of calibration curves in ion chromatography with suppressed conductivity detection using hydroxide eluents was investigated. Theoretical calibration curves were derived for strong electrolytes and weak monobasic acids and the results compared with experimental data. At low concentrations up to 1 micromol l(-1) the autoprotolysis of water induces left-curved calibration functions even for strong electrolytes like nitrate. The experimental data are best described by a quadratic function, the differences between linear and quadratic regression being up to 10%. At higher concentrations the calibration curves for strong electrolytes are linear. Due to incomplete dissociation, the calibration curves for weak mono- and dibasic acids show a right curvature. Thus, depending on the analyte and the concentration range of interest, analysts should carefully choose between a linear and a quadratic regression function. PMID- 12113348 TI - The lost tribe of hydrogeologists? PMID- 12113349 TI - Perspectives on Japanese ground water resources. PMID- 12113350 TI - Ground water age. PMID- 12113351 TI - The water budget myth revisited: why hydrogeologists model. AB - Within the ground water community, the idea persists that if one can estimate the recharge to a ground water system, one then can determine the size of a sustainable development. Theis addressed this idea in 1940 and showed it to be wrong-yet the myth continues. The size of a sustainable ground water development usually depends on how much of the discharge from the system can be "captured" by the development. Capture is independent of the recharge; it depends on the dynamic response of the aquifer system to the development. Ground water models were created to study the response dynamics of ground water systems; it is one of the principal reasons hydrogeologists model. PMID- 12113352 TI - Prediction of diffusion coefficients in porous media using tortuosity factors based on interfacial areas. AB - Determination of aqueous phase diffusion coefficients of solutes through porous media is essential for understanding and modeling contaminant transport. Prediction of diffusion coefficients in both saturated and unsaturated zones requires knowledge of tortuosity and constrictivity factors. No methods are available for the direct measurement of these factors, which are empirical in their definition. In this paper, a new definition for the tortuosity factor is proposed, as the real to ideal interfacial area ratio. We define the tortuosity factor for saturated porous media (tau5) as the ratio S/S(o) (specific surface of real porous medium to that of an idealized capillary bundle). For unsaturated media, tortuosity factor (tau(a)) is defined as a(aw)/a(aw),o (ratio of the specific air-water interfacial area of real and the corresponding idealized porous medium). This tortuosity factor is suitably measured using sorptive tracers (e.g., nitrogen adsorption method) for saturated media and interfacial tracers for unsaturated media. A model based on this new definition of tortuosity factors, termed the interfacial area ratio (IAR) model, is presented for the prediction of diffusion coefficients as a function of the degree of water saturation. Diffusion coefficients and diffusive resistances measured in a number of saturated and unsaturated granular porous media, for solutes in dilute aqueous solutions, agree well with the predictions of the IAR model. A comparison of permeability of saturated sands estimated based on tau(s) and the same based on the Kozeny-Carman equation confirm the usefulness of the tau(s) parameter as a measure of tortuosity. PMID- 12113353 TI - Rapid evolution of redox processes in a petroleum hydrocarbon-contaminated aquifer. AB - Ground water chemistry data collected over a six-year period show that the distribution of contaminants and redox processes in a shallow petroleum hydrocarbon-contaminated aquifer has changed rapidly over time. Shortly after a gasoline release occurred in 1990, high concentrations of benzene were present near the contaminant source area. In this contaminated zone, dissolved oxygen in ground water was depleted, and by 1994 Fe(III) reduction and sulfate reduction were the predominant terminal electron accepting processes. Significantly, dissolved methane was below measurable levels in 1994, indicating the absence of significant methanogenesis. By 1996, however, depletion of solid-phase Fe(III) oxyhydrox ides in aquifer sediments and depletion of dissolved sulfate in ground water resulted in the onset of methanogenesis. Between 1996 and 2000, water chemistry data indicated that methanogenic metabolism became increasingly prevalent. Molecular analysis of 16S-rDNA extracted from sediments shows the presence of a more diverse methanogenic community inside as opposed to outside the plume core, and is consistent with water-chemistry data indicating a shift toward methanogenesis over time. This rapid evolution of redox processes reflects several factors including the large amounts of contaminants, relatively rapid ground water flow (approximately 0.3 m/day [approximately foot/day]), and low concentrations of microbially reducible Fe(III) oxyhydroxides ( approximately 1 micromol/g) initially present in aquifer sediments. These results illustrate that, under certain hydrologic conditions, redox conditions in petroleum hydrocarbon-contaminated aquifers can change rapidly in time and space, and that the availability of solid-phase Fe(III)-oxyhydroxides affects this rate of change. PMID- 12113354 TI - A subregional-scale method to assess aquifer vulnerability to pesticides. AB - A method to predict aquifer vulnerability to pesticide contamination at the subregional scale was developed. The assessment method was designed to incorporate relevant hydrologic and pesticide-transport information and to use generally available data. The method assumes steady-state advection of pesticides in the vadose zone, including sorption and biological decay. The solution is presented as a vulnerability index (VI) that increases as the aquifer vulnerability increases. The hydrologic input data for the VI model are obtained from the soil survey geographic database. Pesticides were grouped into three leachability classes using a leachability ratio (half-life divided by organic carbon partition coefficient). Pesticide transformation is assumed to occur in the surface layer. The influence of vertical transport in the remainder of the vadose zone has been incorporated by applying a multiplying factor to the VI that varies with depth to ground water. Hydraulic conductivity is used as a surrogate for soil-water velocity for practical purposes. The performance of the VI model is evaluated using ground water data from Weld County, Colorado. The model is demonstrated to be successful at predicting relative vulnerability, defined as the magnitude of pesticide concentration and percent of wells in a unit that exhibit a pesticide detection. Areas of low, medium, and high vulnerability are assigned. Results indicate that the vulnerability classifications are most dependent on the leachability ratio, hydraulic conductivity, and organic carbon content. PMID- 12113355 TI - Numerical examination of the factors controlling DNAPL migration through a single fracture. AB - The migration of five dense nonaqueous phase liquids (DNAPLs) through a single fracture in a clay aquitard was numerically simulated with the use of a compositional simulator. The effects of fracture aperture, fracture dip, matrix porosity, and matrix organic carbon content on the migration of chlorobenzene, 1,2-dichloroethylene, trichloroethylene, tetra-chloroethylene, and 1,2 dibromoethane were examined. Boundary conditions were chosen such that DNAPL entry into the system was allowed to vary according to the stresses applied. The aperture is the most important factor of those studied controlling the migration rate of DNAPL through a single fracture embedded in a clay matrix. Loss of mass to the matrix through diffusion does not significantly retard the migration rate of the DNAPL, particularly in larger aperture fractures (e.g., 50 microm). With time, the ratio of diffusive loss to the matrix to DNAPL flux into the fracture approaches an asymptotic value lower than unity. The implication is that matrix diffusion cannot arrest the migration of DNAPL in a single fracture. The complex relationships between density, viscosity, and solubility that, to some extent, govern the migration of DNAPL through these systems prevent accurate predictions without the use of numerical models. The contamination potential of the migrating DNAPL is significantly increased through the transfer of mass to the matrix. The occurrence of opposite concentration gradients within the matrix can cause dissolved phase contamination to exist in the system for more than 1000 years after the DNAPL has been completely removed from the fracture. PMID- 12113356 TI - Analytical model for aquifer response incorporating distributed stream leakage. AB - An analytical model of stream/aquifer interaction is proposed that predicts drawdown in an aquifer with leakage from a finite-width stream induced by pumping from a well. The model is formulated based on the assumptions of stream partial penetration, a semipervious streambed, and distributed recharge across a finite width stream. Advantages of the analytical solution include its simple structure, consisting of the Theis well function with integral modifications. The solution is derived for the semi-infinite domain between the stream and pumping well, which is of primary interest to hydrogeologists. Previous stream/aquifer analytical models are compared to the analytical solution based on dimensionless drawdown profiles. Drawdown in the aquifer near a wide stream was found to be less than that predicted by a solution that ignored stream width. Deviations between the proposed analytical solutions and previous solutions increase as stream width increases. For a hypothetical stream/aquifer system, the proposed analytical solution was equivalent to prior solutions when the ratio of the distance between the stream and aquifer to the stream width was greater than 25. This analytical solution may provide improved estimates of aquifer and streambed leakage parameters by curve fitting experimental field drawdown data. PMID- 12113357 TI - Methods to determine storativity of infinite confined aquifers from a recovery test. AB - Starting from the equations of Theis and Cooper-Jacob, two new mathematical methods are proposed for interpreting the residual drawdown data for an infinite confined aquifer. Under Theis' assumptions and using the Cooper-Jacob approximation, the principal aquifer characteristics of transmissivity, pumping storativity, and recovery storativity are expressed without any correction or additional assumption. An actual case is used for illustration and confirms the validity of proposed equations and methods. PMID- 12113358 TI - Michigan basin regional ground water flow discharge to three Great Lakes. AB - Ground water discharge to the Great Lakes around the Lower Peninsula of Michigan is primarily from recharge in riparian basins and proximal upland areas that are especially important to the northern half of the Lake Michigan shoreline. A steady-state finite-difference model was developed to simulate ground water flow in four regional aquifers in Michigan's Lower Peninsula: the Glaciofluvial, Saginaw, Parma-Bayport, and Marshall aquifers interlayered with the Till/"red beds," Saginaw, and Michigan confining units, respectively. The model domain was laterally bound by a continuous specified-head boundary, formed from lakes Michigan, Huron, St. Clair, and Erie, with the St. Clair and Detroit River connecting channels. The model was developed to quantify regional ground water flow in the aquifer systems using independently determined recharge estimates. According to the flow model, local stream stages and discharges account for 95% of the overall model water budget; only 50% enters the lakes directly from the ground water system. Direct ground water discharge to the Great Lakes' shorelines was calculated at 36 m3/sec, accounting for 5% of the overall model water budget. Lowland areas contribute far less ground water discharge to the Great Lakes than upland areas. The model indicates that Saginaw Bay receives only approximately 1.13 m3/sec ground water; the southern half of the Lake Michigan shoreline receives only approximately 2.83 m3/sec. In contrast, the northern half of the Lake Michigan shoreline receives more than 17 m3/sec from upland areas. PMID- 12113359 TI - The role of capillary forces in steady-state and transient seepage flows. AB - Steady and transient seepage problems were investigated numerically using a dimensionless formulation for water flow in variably saturated, two-dimensional anisotropic and homogenous porous media. The dimensionless formulation combines the aspect ratio and the anisotropy ratio in one dimensionless parameter, M, and accounts for capillarity effects using the ratio of the domain height to the height of capillary fringe as another dimensionless parameter, alpha. Two domain geometries were considered: rectangular and trapezoidal. It was found that M and alpha play major roles in the development and the height of the seepage face. The height of the steady-state seepage face increased with the decreasing value of alpha. The effects of capillarity on the development of the transient seepage face were investigated by lowering the open water level on one side of the domain at various uniform velocities while the other side was kept at a constant value. It was found for most scenarios that decoupling between the water table and the open water level first occurred for domains with large alpha. In other words, the seepage face heights of these systems were larger than those with a small alpha. These results are the opposite of the steady-state results and are due to the fact that drainage of the pores was the major mechanism controlling the drop of the transient water table, especially at earlier times. A criterion for the decoupling of the water table from the falling open water level (i.e., the formation of the transient seepage face) was developed by Dracos (1965). It states that decoupling does not occur as long as the falling speed, VF, of the open water level is less than Vd = K(o) sin2 B/phi where K(o) is the saturated hydraulic conductivity of the soil, phi is the porosity, and beta is the angle of the exit face with the horizontal (i.e., beta = pi/2 implies a vertical face). Our investigation revealed that decoupling occurs for smaller falling velocities than Vd, and is caused by, among others, the fact that the water table is not tangent to the exit face but rather intersects it at a nonzero angle. PMID- 12113360 TI - Localized thermal anomalies in haloclines of coastal Yucatan sinkholes. AB - A temperature spike is reported in the haloclines of three Yucatan sinkholes along a 1 km NW-SE transect from 5 to 4 km inland from the Caribbean coast. The temperature spike decreases in magnitude from 3.5 degrees C to 0.2 degrees C, approaching the coast. The anomaly does not vary diurnally and does not extend down into the underlying sea water. These conditions are inconsistent with explanations such as radiation absorption within the halocline, in situ microbially mediated sulfate reduction within the halocline and the underlying sea water, and sulfide oxidation by photosynthetic purple and green bacteria within the halocline. One explanation consistent with the shape and halocline location of the temperature spike involves a localized sea water convection cell operating near the coast. Cold sea water from the Caribbean Sea enters the coastal limestone at depths of a few hundred meters and heats up because of the geothermal gradient, buoyantly rising in vertical fractures within the unconfined aquifer. Blocked by the less dense fresh water, the movement stops in the halocline where the warm sea water mixes with brackish water. The convection cycle would be completed by the coastward movement of cooling brackish water. The observed temperature anomalies could possibly be snapshots of this warm layer moving coastward. PMID- 12113361 TI - Flow depletion in a small stream caused by ground water abstraction from wells. AB - The interaction between a gaining stream and a water-table aquifer is studied at an outwash plain. The aquifer is hydraulically well connected to the stream. Pumping tests were carried out in 1997 and 1998 in two wells 60 m from the stream, screening different depths of the aquifer. Drawdown was measured on both sides of the stream. Hydraulic head, drawdown, and stream depletion data were analyzed using numerical flow models. Similar models were fitted to each of two different data sets: Model A was fitted to steady-state hydraulic head and streamflow gain data not influenced by pumping; and model B was fitted to drawdown data measured during the 1998 pumping test. Each calibrated model closely fits its calibration data; however, predictions were biased if model A was used to predict the calibration data of model B, and vice versa. To further test the models, they were used to predict streamflow depletion during the two pumping tests as well as the drawdown during the 1997 test. Neither of these data were used for calibration. Model A predicted the measured depletions fairly accurately during both tests, whereas the predicted drawdowns in 1997 were significantly larger than actually measured. Contrary to this, the 1997 drawdowns predicted by model B were nearly unbiased; the predicted depletions deviate significantly from the measured depletions in 1997, but they compare well with the observations in 1998. Thus, although field work and analyses were extensive and done carefully to develop a ground water flow model that could predict both drawdown and streamflow depletion, the model predictions are biased. Analyses indicate that the deviations between model and data may be because of error in the models' representations of either the release of water from storage or of the hydrology in the riparian zone. PMID- 12113362 TI - Role of ground water in geomorphology, geology, and paleoclimate of the Southern High Plains, USA. AB - Study of ground water in the Southern High Plains is central to an understanding of the geomorphology, deposition of economic minerals, and climate change record in the area. Ground water has controlled the course of the Canadian and Pecos rivers that isolated the Southern High Plains from the Great Plains and has contributed significantly to the continuing retreat of the westward escarpment. Evaporative and dissolution processes are responsible for current plateau topography and the development of the signature 20,000 small playa basins and 40 to 50 large saline lake basins in the area. In conjunction with eolian processes, ground water transport controls the mineralogy of commercially valuable mineral deposits and sets up the distribution of fine efflorescent salts that adversely affect water quality. As the water table rises and retreats, lunette and tufa formation provides valuable paleoclimate data for the Southern High Plains. In all these cases, an understanding of ground water processes contributes valuable information to a broad range of geological topics, well beyond traditional interest in water supply and environmental issues. PMID- 12113363 TI - "A stream depletion field experiment," by Bruce Hunt, Julian Weir, and Bente Clausen, March-April 2001 issue, v. 39, no.2: 283-289. PMID- 12113364 TI - Water witching and dowsing. PMID- 12113365 TI - The evolution and current status of emergency medical services for children. AB - Emergency physicians have come to believe that comprehensive pediatric emergency care should be integrated into an overall medical system and organized regionally to address the special needs of children. Since our emergency medical systems have evolved in the care of adult trauma victims, we must look to that development for the origin of our present emergency medical services for children. Not until the 1970s did it became obvious that children, just as adults, should be included in an comprehensive emergency medical system for the care of their life threatening injuries. Since there is considerable overlap in the basic principles of trauma resuscitation and management of shock in children and adults, most children's regional trauma program developed as a part of an overall emergency medical system. The sharing of trauma facilities made it possible to utilize the special expertise of a very small number of pediatric surgeons who had trauma experience and to incorporate their skills into the broader concept of comprehensive regional trauma centers for children and adults. With further experience we soon realized that such emergency medical services for children's systems should include the following components: A two way communication system, a transport system with special equipment for the management of small children, a training program for first responders at the trauma site, a designated pediatric trauma center, a pediatric intensive care unit at the regional trauma center, a neurology/neurosurgery intermediate care unit at the regional center, a pediatric trauma rehabilitation unit and finally a pediatric trauma longterm rehabilitation and management unit for those with residual disabilities. With the further development of this concept of trauma units for children, pediatric surgeons, general surgeons involved in trauma care and pediatric emergency physicians have offered leadership to expand the emergency medical system for children to include life-threatening illness, as well as injuries. Thus, the organization of regional emergency medical services for children permits the highest quality management of children with life threatening injuries and illness. This then is the final product: an inclusive, comprehensive emergency medical system for children for all life threatening conditions, both trauma and serious illness. PMID- 12113366 TI - Practical points in evaluation and resuscitation of the injured child. AB - The ultimate goal of resuscitation of an injured child is delivery of oxygen to intracellular organelles in order to maintain aerobic metabolism. This can be obtained by following ATLS protocols with immediate attention to the "ABCDE's" and compulsive reevaluation of the adequacy of resuscitation maneuvers. After stabilization, seriously injured children should be transferred to trauma centers with established pediatric trauma programs utilizing preexisting transfer agreements and protocols. Pediatric trauma is indeed a team endeavor, requiring the coordinated expertise and teamwork of prehospital EMS providers, trauma team members, and the pediatric trauma and rehabilitation centers. With careful and compulsive communication and coordination, injured children can be returned to their families in better mental and physical condition than pre-injury with reasonable expectation of a full and productive life. PMID- 12113367 TI - Error and time delay in pediatric trauma resuscitation: addressing the problem with color-coded resuscitation aids. AB - This article concentrates on two pediatric specific factors operational in pediatrics therapeutics 1) the necessity to perform calculations; and 2) the lack of recognition of significant pediatric error. Resuscitation aids and their optimal function in pediatric trauma resuscitation are examined. PMID- 12113368 TI - Imaging of the injured child: important questions answered quickly and correctly. AB - Diagnostic imaging has assisted dramatically in the care of the injured child. The avoidance of unnecessary laparotomy in 35-85% of cases depending on the series reviewed is a major improvement in care. Additionally, non-operative treatment has clearly decreased the number of splenectomies children significantly undergo. Further improvements will only result in more accurate and focused interventions and continue to improve results. Physicians must remember, however, that trauma in children remains a surgical disease. Advances in imaging techniques have reduced operative intervention, but now children are exposed to far more potential risks and morbidity. Diagnoses evolve with time and the surgeon must be ready to take any injured child to the operating room for treatment when warranted at a moment's notice. PMID- 12113369 TI - Beyond the golden hour: avoiding the pitfalls from resuscitation to critical care. AB - The transition from resuscitation bay to intensive care unit is wrought with potential problems that can significantly affect patient outcome. Among these are hypothermia, injudicious fluid management, hypocarbia, incorrect drug dosing, and incomplete evaluation. Based on a comprehension of the ongoing pathophysiology associated with injury, steps can be taken to ensure that this process positions the patient on the beginning of the path to recovery and not further into the spiral of worsening organ system dysfunction. PMID- 12113370 TI - Critical care of the severely injured child. AB - It is the responsibility of the surgeon committed to care of these critically injured patients to understand the nature of injuries being treated, and to orchestrate this treatment in a manner that maximizes recovery, avoids unnecessary morbidity, and assures the injured child the best quality of life humanly possible. PMID- 12113371 TI - Management of childhood neurotrauma. AB - A summary of some of the more important aspects of brain, spinal, peripheral nerve and sport injuries of childhood is presented. Guidelines for the treatment of severe brain injury have been developed for adults, are currently employed with success to treat children, but much information still needs to be acquired about childhood brain injury so that better age specific treatment modalities could be implemented. The unique anatomy of the spine during childhood predisposes to cervical spinal injury without radiographic abnormality; immobilization is the primary treatment and a minority of cases require surgery. Peripheral nerve injuries are uncommon, often missed, and require skillful evaluation and early treatment by physical therapy and oftentimes surgery. Appreciation of the sequelae of cerebral concussion, education on proper sport techniques, body conditioning, and equipment upkeep are the mainstay of vigilant sport injury treatment and prevention. PMID- 12113372 TI - Infection control: avoiding the inevitable. AB - Infection, while a major cause of morbidity, should not be considered an inevitable consequence of injury. Good aseptic technique, compulsive attention to detail, and thorough understanding of the points addressed in the following list of critical points are the best guarantee that infection will not add avoidable morbidity to misfortune. Critical points regarding infectious problems in care of the injured child: 1. Polymicrobial infection is the rule with 50% of isolates being mixed aerobic and anaerobic bacteria. 2. It is a misnomer to consider antibiotic use in a pediatric trauma victim as prophylactic. Antimicrobials used in this setting are best considered adjunctive. 3. The major indication for anti infective therapy in pediatric trauma is an injury with a high probability of infection. 4. Antibiotics do not sterilize the wound or body cavity; they limit bacterial proliferation, thereby supplementing effective immune control. 5. Available studies suggest that 24 hours is as efficacious as a longer treatment duration in a purely adjunctive mode. 6. In bites inflicted by dogs and cats, Pasturella species are frequent. 7. Human bites may result in infection by Eikenella corrodens. 8. Based on this bacteriology, adjunctive intravenous ampicillin sulbactam or oral amoxicillin clavulanate are recommended for human and animal bites. 9. Tetanus prophylaxis is indicated in all significant soft tissue injuries. 10. Risk of osteomyelitis correlates directly with the extent of the associated soft tissue injury and vascular compromise. 11. The majority of infectious complications in the injured child are not a consequence of the injury itself, but rather in the treatment thereof. 12. In the injured child the most common nosocomial infection is lower respiratory followed by primary blood stream and the urinary tract. 13. The management of nosocomial pneumonia in the injured child is based on the time of diagnoses. Early evidence of pulmonary infection requires treatment with a third generation cephalosporin with or without an antistaphylococcal penicillin. Late pneumonia is treated with an aminoglycoside with or without an antipseudomonal added. 14. Catheter related infection is, in the injured child, overwhelmingly gram positive with coagulase negative staphlococcal species accounting for 30-60% of isolates. Staphlococcus aureus is responsible for an additional 15-20%. 15. The role of antibiotics in the prevention of catheter related meningitis is controversial. Recent adult studies suggest no advantage to their routine use. If utilized, they should only be employed prophylactically and not continued throughout the monitoring period. 16. Lack of response to treatment of sepsis may represent an inappropriate antimicrobial agent, improper dosage, inability to achieve adequate levels at the site of infection. (eg, CSF) fungal pathogen, and/or ongoing contamination or undrained purulent focus. PMID- 12113373 TI - Effective and efficient nutritional support for the injured child. AB - This article discusses the critical points in nutritional support of the injured child. Each applies currently understood basic science information to practical bedside care. PMID- 12113374 TI - Rehabilitation and outcome following pediatric traumatic brain injury. AB - The long-term outcome for a child who has sustained a traumatic brain injury must be viewed in the context of ongoing development and maturation. Although neuronal plasticity provides the potential for neuronal reorganization in a child's brain, it is the behavioral demands of the environment that allow the child to take advantage of this potential and to maximize recovery. Pediatric rehabilitation is the setting that provides the necessary experiences for stimulating neuronal reorganization following TBI. However, neuronal reorganization has a cost to long term development. The ultimate long-term impact of a TBI sustained in childhood depends on the child's ability to achieve developmental milestones following injury. Although injury-related and treatment-related factors are critical during the early stages of recovery, patient-related factors such as age-at-injury, developmental achievement at time of injury, maturation, and family involvement and resources impact the later stages of recovery. The process of pediatric rehabilitation following TBI is to provide an enriched, stimulating environment tailored to the needs of the child and based on real-word experiences. Early in the recovery process, pediatric rehabilitation is the setting that maximizes the potential for neuronal reorganization. Early rehabilitation also prepares the family for the child's long-term recovery and developmental needs. Involvement and training of family members early in the recovery process is critical for successful long-term outcome. Family members are the individuals best equipped to ensure treatment compliance and follow through with treatment recommendations, in maintaining treatment gains, and in generalizing treatment effects beyond the medical settings. Despite the life-long ramifications of childhood TBI, pediatric rehabilitation is the necessary step in promoting recovery and successful long term outcome. PMID- 12113375 TI - Practical aspects of performance improvement in pediatric trauma. AB - Pediatric trauma performance improvement programs may share some of the criteria tracked by their counterparts in the adult trauma world. However, some of the criteria must be specific to the unique diagnostic and therapeutic needs of children. Nine criteria are defined in terms of the critical issues, what information is required to evaluate the appropriateness of the care provided in regards to those issues, and acceptable thresholds for review. In addition, practical aspects of multi-disciplinary peer review in the performance improvement process is presented. PMID- 12113376 TI - The psychological aspects of pediatric trauma: perspectives on patient, family, and provider. AB - Pediatric trauma engenders multi-system injury--physiological as well as psychological. Not only does injury to a child affect the patient, but also the family system within which the child functions. Professional caregivers, too, are not immune to the emotional aftermath of pediatric traumatic injury. Effective management of the pediatric trauma patient must, therefore, include an understanding of the psychological aspects of injury--for the patient, the family, and the provider. This chapter seeks to delineate these variables in an effort to promote effective identification of the emotional components of injury and facilitation of necessary interventions in promoting positive overall injury outcomes. PMID- 12113377 TI - Caring for the injured children of our world: a global perspective. AB - This article describes several countries that began and continue active pediatric trauma programs, which can serve as examples for developing countries. Emergency medical services for children need to be developed so that the special needs of children with serious injuries are addressed. PMID- 12113378 TI - Evolution of dry powder inhaler design, formulation, and performance. AB - Many companies are now prioritizing the development of dry powder inhalers (DPIs) above pressurized formulations of asthma drugs. A well-designed DPI and an appropriate powder formulation can optimize the effectiveness of inhaled drug therapy. A DPI must be able to deliver medications effectively for most patients, and an ideal inhaler would provide a dose that does not vary with inspiratory flow rate. Recent regulatory guidelines, among which the U.S. FDA draft guidance is the most stringent, demand consistent dose delivery from an inhaler throughout its life and consistency of doses from one inhaler to another. However, the properties of free micronized powders often interfere with drug handling and with drug delivery reducing dose consistency. Recent advances in formulation technology can increase lung dose and reduce its variability. While a perfect DPI may never exist, both device and formulation technology are evolving to rectify perceived deficiencies in earlier systems. PMID- 12113379 TI - Differences in trigger factors and symptoms between patients with asthma-like symptoms and patients with asthma: development of a basis for a questionnaire. AB - Patients with asthma-like symptoms but with negative asthmatests are often misdiagnosed as having asthma and treated as asthmatics. They describe their trigger factors and symptoms very similar to those of patients with asthma. The aim of the study was to analyze differences in symptoms and trigger factors between asthma-like patients and asthmatics in order to elaborate a basis for a questionnaire for epidemiological and clinical use. A questionnaire with 54 questions about trigger factors and 137 questions about symptoms was sent to 40 patients with asthma-like symptoms and 40 with asthma, all consecutively selected from patients referred to an out-patient clinic for asthma and allergy for investigation of suspected asthma. Data were analyzed statistically in two steps using multiple logistic regression analysis. Significant differences were seen in several trigger factors and symptoms after the first analysis. After the second analysis, seven out of the 54 trigger factors and 22 out of the 137 symptoms emerged as those that most significantly discriminated between the two patient groups. These trigger factors and symptoms can be the basis of a new questionnaire with high discriminating power. Before using it, it is important to evaluate the best combination of variables, add some demographic variables and totestthe reliability and validity ofthis new questionnaire. PMID- 12113380 TI - Exhaled nitric oxide and exercise tolerance in severe COPD patients. AB - To answer the question as to whether pulmonary rehabilitation programs (PRP) induced increase in exercise tolerance (ET) is associated with increased levels of exhaled nitric oxide (eNO) in COPD patients of different degrees of severity, we designed a prospective and controlled study. Forty-seven stable COPD patients underwent an 8-week outpatient multidisciplinary PRP including supervised incremental exercise. Fractional eNO concentration (FE(NO)) and peak work-rate (W(peak) were assessed baseline (T-1), atthe end of 1-month run-in period (T0), and after (T1) the PRP. Lung function, walking test, health-related quality of life (HRQL) were also recorded. Patients were divided into three groups according to disease severity: 17 severe [FEV1 35 (5)% pred] COPD patients, seven of them with cor pulmonale; 15 mild [FEV1 78 (6)% pred], and 15 moderate [FEV1 56 (6)% pred] COPD patients. FE(NO) did not differ at T-1 and T0 (mean absolute change (SD): 0.03 (0.09) 95% CI-0.01, 0.16, 0.06 (1.03) 95% CI 0.03, 0.75 and 0.05 (0.06) 95% CI 0.02, 0.11 ppb in mild, moderate and severe patients, respectively). As compared to T0, both W(peak) (by 17,15 and 10%, respectively) and FE(NO) (by 29, 24 and 16%, respectively) significantly increased in all groups, but not in patients with cor pulmonale. A significant correlation between pre- and post-PRP changes in Wpeak and FE(NO) was found both in mild to moderate (r = 0.79, P < 0.00001) and severe (r = 0.76, P < 0.001) COPD patients. After a PRP, improvement in ET is associated with an increase in eNO also in most severe COPD patients, but not in those with cor pulmonale. PMID- 12113381 TI - Kinetics of lung clearance of 99mTc-DTPA in smoking patients with sarcoidosis compared to healthy smokers. AB - Investigation of lung clearance of 99mTc-labelled diethylene triamine penta acetic acid (DTPA) in smoking sarcoid patients has been impeded by difficulties to differ between pathology of clearance kinetics caused by sarcoidosis and by smoking. This study explores the kinetics of lung clearance of 99mTc-DTPA in 15 current smokers with intrathoracic sarcoidosis. The results are compared with findings from 16 healthy smokers. Measurements of lung clearance over 180 min, i.e. longer than usual, revealed in II of the sarcoid patients a bi-exponential lung clearance course, which is pathologic. All healthy smokers also showed a bi exponential lung clearance. In the analysis of the bi-exponential curve an initial fast, and a slow clearance component could be separated. The smokers with sarcoidosis had a significantly higher elimination rate of the slow component than the healthy smokers. Thus, analysis of the late part of the lung clearance curve may be rewarding in smoking sarcoid patients. The study shows that lung clearance of 99mTc-DTPA may be a method useful also in smoking patients with sarcoidosis. PMID- 12113382 TI - Comparison of second controller medications in addition to inhaled corticosteroid in patients with moderate asthma. AB - The objective of this study was to compare the efficacy and safety of the second controller medications (long-acting beta2-agonist, leukotriene receptor antagonist and sustained-release theophylline) used in addition to inhaler corticosteroid treatment in moderate persistent asthma. A total of 64 patients with asthma, in the moderate persistent asthma category, were divided into three groups. Patients, all of whom were concurrently using inhaled corticosteroid (Budesonide 400 microg twice daily), were treated for 3 months with either inhaled formoterol 9 microg twice daily (first group), oral zafirlukast 20 mg twice daily (second group), or sustained-release theophylline 400 mg once daily (third group). All of the patients were subjected to assessments on the subject of peak expiratory flow (PEF) variability, forced expiratory volume in 1 sec (FEV1), asthma symptom scores (daytime and night-time), supplemental terbutalin use, asthma exacerbations and adverse events. Over the 3-month treatment period. In all of the three groups, significant improvements were recorded in the lung function, asthma symptom scores and supplemental terbutalin use criteria, as a result oftreatments applied. Formoterol treatment resulted in significantly greater and earlier improvements compared with the other two groups in several criteria: PEF variability (17.9 +/- 2.5; 21.9 +/- 3.2; 23.7 +/- 3.3; P < 0.001); asthma symptom score (daytime) (1.6 +/- 0.5; 1 +/- 0.5; 2.0 +/- 0,5; P < 0.05); asthma symptom score (night-time) (1.2 +/- 0.4; 2.2 +/- 0.5; 16 +/- 0.6; P < 0001); and supplement alter butalin use (1.2 +/- 0.3; 1.8 +/- 0.5; 1.7 +/- 0.5; P < 0.05). However, at the end of the treatment, in all of the three groups studied, improvements were attained in overall asthma control and there was no statistical difference among the groups. Although there were no side effects which required the discontinuation of the treatment, it was observed that the maximum side effect was in the second group (20%, 31.6% and 20%, respectively). In conclusion, in patients who still have symptoms on treatment with inhaled corticosteroids, the addition of a long-acting beta2-agonist, leukotriene antagonists or sustained-release theophylline to the treatment is a logical approach, and, in addition to inhaled corticosteroids, any one of these second controller medications may be chosen in patients with moderate asthma. PMID- 12113383 TI - Individual dietary intervention in patients with COPD during multidisciplinary rehabilitation. AB - Dietary intervention studies in COPD patients often are short-term inpatient studies where a certain amount of extra energy is guaranteed. The aim of this study was to evaluate the effect of an 1 year individual multifaceted dietary intervention during multidisciplinary rehabilitation. Eighty-seven patients with severe COPD, not demanding oxygen therapy were included, 24 of them served as controls. A dietary history interview was performed at baseline and at study end. Dietary advice given were based on results from the dietary history and socio economic status. The intervention group was divided into three parts; NW: normal weight (dietary advice given aiming to weight maintenance), OW: overweight (weight-reducing advice) and UW: underweight (dietary advise based on an energy- and protein-rich diet). RESULTS: UW-group: Eighty-one per cent of the patients gained weight or kept a stable weight. OW-group: Fifty-seven per cent lost more than 2 kg NW-group: Seventy-six per cent kept a stable weight or gained weight. Increased dietary intake from baseline was seen for energy protein, carbohydrates and certain micronutrients (P < 0.05) in the UW group. Six minutes walking distance increased by approximately 20 m in both NW (P < 0.05) and UW patients. To conclude, slight, but uniform, indications of positive effects of dietary intervention during multidisciplinary rehabilitation was seen. Dietary intervention in underweight COPD patients might be a prerequisite for physical training. PMID- 12113384 TI - Long-term follow-up of untreated patients with sleep apnoea syndrome. AB - Obstructive sleep apnoea (OSA) is a common disorder with numerous potential sequelae. Although the majority of these consequences can be reduced with appropriate treatment, only limited data exist regarding the natural progression ofthis disorder in untreated individuals. We hereby report a long-term follow-up of all untreated patients (n = 40) followed-up in the Technion Sleep Clinic, using both subjective and objective measurements. In addition, we report a long term follow-up of 11 patients who attempted dietary weight loss. The average time interval between the first and second polysomnographies for the untreated group was 5.0 +/- 2.8 yrs, and 2.5 +/- 2.3 yrs for the weight reduction group. There was no significant change in Body Mass Index (BMI) or Respiratory Disturbance Index (RDI) between the two Polysomnographic (PSG) evaluations in the untreated patients. However, eight patients developed hypertension (n=5) or ischaemic heart disease (IHD) (n=3) between the two evaluations. RDI, age and BMI at the time ofthe initial evaluation were not predictive of changes in RDI, snoring intensity or minimal oxygen saturation. However, the patients who developed hypertension/IHD had significantly higher RDI than the patients who did not (46 +/- 27 vs. 23 +/- 17 h(-1), P < 0.005). In the weight-loss group, BMI decreased by a mean of 3.1 kg m(-2), and RDI decreased by 20events h(-1), P<0.05 for both. There was a significant correlation between the weight loss and improvement in RDI (R = 0.75, P = 0.005). We conclude that in untreated obstructive sleep apnoea patients RDI does not necessarily increase over time, but associated hypertension or ischaemic heart disease may develop. When weight loss is successfully achieved, sleep apnoea significantly improves with a high correlation between the extent of weight loss and the improvement in apnoea status. PMID- 12113385 TI - Intensified microbiological investigations in adult patients admitted to hospital with lower respiratory tract infections. AB - The objective of this study was to investigate the diagnostic yield of a programme with intensified microbiological investigations in immunocompetent adult patients with lower respiratory tract infections (LRTI). Patients in the study group were included prospectively and consecutively from September 1st 1997 to May 31st 1998 and were compared with a control group from the preceding year. A total of 67 adult patients were included in the study group and they were compared with 122 adult patients in the control group. The study group underwent fibre-optic bronchoscopy (FOB) with bronchoalveolar lavage (BAL). Only 7% in the historic control group were discharged with an aetiological diagnosis of their infections; while the diagnostic yield in the study group increased to 51% of patients. In the study group the presence of new infiltrates on chest X-ray increased the detection of a microbiological aetiology from 37% with no infiltrates to 62% with infiltrates and recent antibiotic therapy reduced the detection of a microbiological cause of infection from 61% in 36 patients who had not received antibiotic therapy to 39% in 31 patients who had received recent antibiotic therapy prior to microbiological sampling. Patients in the study group with known aetiology had higher values of inflammatory markers than patients with unknown aetiology. For Streptococcus pneumoniae infection culture and urine antigen detection were complimentary depending on recent antibiotic therapy since seven of eight culture-positive patients had not received antibiotic therapy within 72 h prior to investigation, while all four patients positive for urine antigens from S. pneumoniae had received antibiotic therapy within 72 h of urine sampling. In conclusion intensified microbiologic investigations increase the diagnostic yield from 7% to 51% of patients in the study group with an aetiologic diagnosis. Routine FOB with BAL had no apparent effect on clinical outcome and seems only justified in selected patients with severe LRTI with infiltrates on chest X-ray and signs of severe inflammation where a high diagnostic yield is achieved. PMID- 12113387 TI - Lack of additional effect of adjunct of assisted ventilation to pulmonary rehabilitation in mild COPD patients. AB - Different modalities of assisted ventilation improve breathlessness and exercise tolerance in patients with chronic obstructive pulmonary disease (COPD). The aim of this study was to evaluate the effects of the addition of assisted ventilation during exercise training on the outcome of a structured pulmonary rehabilitation programme (PRP) in COPD patients. Thirty-three male patients with stable COPD (mean (SD) forced expiratory volume in 1 s (FEV1) 44 (16) % pred), without chronic ventilatory failure, undergoing a 6-week multidisciplinary outpatient PRP including exercise training, were randomised to training during either mask proportional assist ventilation (PAV: 18 patients) or spontaneous breathing (SB: 15 patients). Assessment included exercise tolerance, dyspnoea, leg fatigue, and health-related quality of life (HRQL). Five out of 18 patients (28%) in the PAV group dropped out due to lack of compliance with the equipment. Both groups showed significant post-PRP improvements in exercise tolerance (peak work rate difference: 20 (95% Cl 2.4-37.6) and 14 (3.8% CI to 24.2) W in PAV and SB group, respectively), dyspnoea and leg fatigue, but not in HRQL, without any significant difference between groups. It is concluded that with the modality and in the patients assessed in this study assisted ventilation during training sessions included in a multidisciplinary PRP was not well tolerated by all patients and gave no additional physiological benefit in comparison with exercise training alone. PMID- 12113386 TI - Effect of ozone on bronchial mucosal inflammation in asthmatic and healthy subjects. AB - Epidemiological studies suggestthat asthmatics are more affected by ozone than healthy people. This study tested three hypotheses (1) that short-term exposure to ozone induces inflammatory cell increases and up-regulation of vascular adhesion molecules in airway lavages and bronchial tissue 6 h after ozone exposure in healthy subjects; (2) these responses are exaggerated in subjects with mild allergic asthma; (3) ozone exacerbates pre-existent allergic airways inflammation. We exposed 15 mild asthmatic and 15 healthy subjects to 0.2 ppm of ozone or filtered air for 2 h on two separate occasions. Airway lavages and bronchial biopsies were obtained 6 h post-challenge. We found that ozone induced similar increases in bronchial wash neutrophils in both groups, although the neutrophil increase in the asthmatic group was on top of an elevated baseline. In healthy subjects, ozone exposure increased the expression of the vascular endothelial adhesion molecules P-selectin and ICAM- 1, as well as increasing tissue neutrophil and mast cell numbers. The asthmatics showed allergic airways inflammation at baseline but ozone did not aggravate this at the investigated time point. At 6 h post-ozone-exposure, we found no evidence that mild asthmatics were more responsive than healthy to ozone in terms of exaggerated neutrophil recruitment or exacerbation of pre-existing allergic inflammation. Further work is needed to assess the possibility of a difference in time kinetics between healthy and asthmatic subjects in their response to ozone. PMID- 12113389 TI - Evaluation of the life satisfaction questionnaire (LSQ) using structural equation modelling (SEM). AB - The life satisfaction questionnaire (LSQ) was developed for use in conventional and complementary/anthroposophic care to assess the quality of life/life satisfaction of Swedish women with breast cancer. The first attempt to test the reliability and validity was in a sample of women with breast cancer (n = 362), the second in a random sample of Swedish women (n = 257). A theoretical model with six latent and 34 manifest variables was formulated. The aim of the present study was to perform confirmatory factor analyses using structural equation modelling. The software STREAMS was used. An additional sample of men (n = 263) was randomly selected from the Swedish population register. Confirmatory factor analyses were performed for the combined sample of women and men (n = 520) and for the women with breast cancer. The result of the confirmatory factor analyses showed that the factor structure of the original model was confirmed. The factors were called physical symptoms (PS), sickness impact (SI), quality of everyday activities (QDA), socio-economic situation (SES), quality of family relation (QFA), and quality of close friend relationship (QFR). PMID- 12113388 TI - Stability of time trade-off utilities for health states associated with the treatment of prostate cancer. AB - BACKGROUND: Patients diagnosed with localized prostate cancer face several treatment options. Patient preferences for treatment side effects often dominate the decision making process. We proposed to learn more about the nature of patient preferences, or utilities, for these side effects. METHODS: Two hundred and fifteen men were consecutively enrolled from three institutions for assessment after prostate needle biopsy. Baseline and 6 month follow-up assessments were done using the University of California, Los Angeles Prostate Cancer Index (UCLA PCI), and a laptop utility assessment application, U-Titer II. Patient utility was assessed for current pelvic functions as well as hypothetical pelvic dysfunctions. We calculated stability of utility scores and correlations between utility scores and UCLA PCI scores. RESULTS: Utility scores for current pelvic functions exhibited a significant 'ceiling effect.' Utility scores for current pelvic functions and hypothetical impaired states were stable after 6 months in patients with negative biopsies. In patients who underwent treatment, utility for current sexual function decreased by 0.13 units (p < 0.00) and utility for current urinary function decreased by 0.09 units (p < 0.01). Utility for hypothetical stress urinary incontinence rose in men with a >25-point drop in UCLA PCI score. CONCLUSION: Utilities for some 'current' pelvic functions decreased in tandem with UCLA PCI scores in men who experienced >25-point changes in these scores. Utilities for some 'hypothetical' pelvic dysfunctions rose as men began to actually experience functional changes in those areas. PMID- 12113391 TI - Validation of the KDQOL-SF: a dialysis-targeted health measure. AB - BACKGROUND: In evaluations of dialysis therapy, an assessment of health-related quality of life (HRQOL) is often important. The aim of this study was to determine the basic psychometric properties, reliability and validity of the short form of the KDQOL i.e. the KDQOL-SF, a dialysis-targeted instrument, and to assess its ability to detect changes over time. METHODS: In a prospective cohort study (Netherlands Cooperative Study on the Adequacy of Dialysis, NECOSAD), all new adult ESRD patients in 32 different Dutch centers were consecutively enrolled. Demographic, clinical and HRQOL data were obtained 3 and 12 months after the start of chronic dialysis therapy. RESULTS: The reliability of the KDQOL-SF was supported by test results that were above the recommended minimal values. Validity of KDQOL-SF was confirmed by the hypothesized positive correlations of the overall health rating and renal function, and by the negative correlations between the number of comorbidities and dialysis dose. Moreover, dialysis-targeted dimensions were more sensitive in detecting relevant differences pertaining to kidney diseases than generic dimensions. The KDQOL-SF was able to detect clinical changes over time. CONCLUSIONS: The psychometric properties of the KDQOL-SF were good, and the different dialysis-targeted dimensions were informative with a high reliability and validity. These results support the application of the KDQOL-SF in studies evaluating dialysis therapy. PMID- 12113390 TI - Health-related quality of life in type 1 diabetes without or with symptoms of long-term complications. AB - OBJECTIVE: To measure subjective health-related quality of life (HRQoL) of patients with type 1 diabetes and describe the influence of symptoms of diabetes related long-term complications on HRQoL. METHODS: The 15-D health-related quality of life measure (15D) was used to measure HRQoL of a representative sample of Finnish insulin-treated patients expected to have type 1 diabetes. Background data were gathered with a separate questionnaire. A tobit (censored regression) model was constructed to estimate the effects of symptoms of complications on HRQoL and to separate these effects from those of other health problems and aging. RESULTS: The 15D scores declined markedly with increasing age, and the prevalence of symptoms of long-term complications increased. The tobit regression model showed that these symptoms have a significant negative influence on HRQoL. The model explained over 50% of the variation in the 15D scores. CONCLUSIONS: High prevalence of symptoms of long-term complications combined with their significant negative influence on HRQoL causes substantial losses in terms of quality of life and utility from both individual and societal perspectives. Thus, the importance of secondary prevention, i.e., prevention of complications by better metabolic control, and also the so-far theoretic possibility to prevent type 1 diabetes itself is emphasized. PMID- 12113392 TI - Vision-specific health-related quality of life: content areas for nursing home residents. AB - Nursing home residents have a high prevalence of remediable visual impairment and blindness. Future research on the effectiveness of providing eye care to nursing home residents will need to include a vision-targeted health-related quality of life (HRQOL) instrument appropriate for this population. The purpose of this study was to identify the core content areas for such an instrument. In-depth interviews on vision-related issues were conducted with 40 residents. Interviews were audio-taped, transcribed, and coded using a standardized protocol. Binocular distance and near visual acuity were assessed using the resident's 'walking around' correction to examine whether one vision-specific HRQOL measure could address the needs of residents with 'good' and 'poor' vision. Overall 1070 vision related comments were identified. Residents mentioned 315 problem comments that were grouped into 13 categories, including ocular symptoms (18% of comments), reading (15%), general vision (13%), psychological distress (12%), and activities of daily living (ADLs) (7%). Compared to published data on vision-specific content areas most relevant to community based persons, nursing home residents focused more on ocular symptoms and basic ADLs, with no mention of issues related to driving, home care, and finances. The majority of categories mentioned did not differ on the proportion of comments made- by those with 'good' and 'poor' visual acuity, suggesting that one vision-specific HRQOL instrument would be appropriate for residents with varying levels of visual acuity. Future work will focus on developing a vision-specific HRQOL instrument for nursing home residents. PMID- 12113393 TI - Chronic fatigue and chronic fatigue syndrome: a co-twin control study of functional status. AB - Chronic fatigue syndrome (CFS) and the symptom of chronic fatigue may be accompanied by substantial functional disability. A volunteer sample of twins discordant for fatigue was identified from throughout the US. Fatigued twins were classified using three increasingly stringent definitions: (1) > or = 6 months of fatigue (119 pairs); (2) CFS-like illness based on self-report of the Centers for Disease Control and Prevention CFS research definition criteria (74 pairs); and (3) CFS assessed by clinical examination (22 pairs). Twins with chronic fatigue were compared with their unaffected co-twins on the eight standard scales and two physical and mental component summary scales from the medical outcomes study short-form health survey (SF-36). Substantial impairment was observed for fatigued twins across all levels of fatigue, while scores in the healthy twins were similar to US population values. Mean scores among fatigued twins on the physical and mental component summary scales were below 97 and 77%, respectively, of the US population scores. Diminished functional status was found across increasingly stringent classifications of fatigue and was associated with a dramatic decrement in physical functioning. The symptom of fatigue has a pronounced impact on functional status, especially in the domain of physical functioning. PMID- 12113394 TI - Femoral neck fractures in the elderly: functional outcome and quality of life according to EuroQol. AB - The main purpose of this prospective study was to investigate the functional outcome and health-related quality of life according to EuroQol (EQ-5D) after a femoral neck fracture treated with internal fixation in relatively healthy elderly patients. We also aimed to validate the use of the EQ-5D in routine clinical follow-ups of this group of patients. The inclusion criteria were more than 65 years of age, absence of severe cognitive dysfunction, living independently, and unhindered walking ability preoperatively. The mean follow-up period was 17 months. The rated prefracture EQ-5Dindex scores showed good correspondence with the scores of an age-matched Swedish reference population. The EQ-5Dindex scores decreased from 0.78 before the fracture (based on recall) to 0.59 at 4 months and 0.51 at 17 months after surgery. The decrease was significantly larger among patients with fracture healing complications. There was a good correlation between the EQ-5Dindex scores and other outcome measures such as pain, mobility, independence in ADL and independent living status. The questionnaire response rate (EQ-5D) was 89-100% on different follow-up occasions. The EQ-5D appears to be an easy-to use instrument even for elderly patients with femoral neck fractures. Changes in the quality of life may be useful to identify patients who might benefit from reoperation, i.e. arthroplasty. The EQ-5D also appears to be a relevant clinical end-point in outcome studies. PMID- 12113396 TI - The equivalence of English and Chinese SF-36 versions in bilingual Singapore Chinese. AB - OBJECTIVE: To assess the equivalence of English and Chinese versions of the SF 36. METHODS: Using a crossover design with block randomisation and stratification by age, identical English or Chinese questionnaires containing the English (UK) and Chinese (HK) SF-36 versions were administered 3-16 days apart to 168 free living, bilingual, ethnic Chinese volunteers in Singapore. Item level equivalence of both versions was assessed by comparing item means and orderings within each scale. Scale level equivalence was assessed by comparing internal consistency (Cronbach's alpha), results of factor analysis and mean scale scores (using paired t-tests and intra-class correlations). RESULTS: Item and scale level comparisons supported the equivalence of both versions. For both the versions, item means, item ordering and Cronbach's alpha were similar, and factor analysis yielded two factors with similar factor loadings. There was no clinically important difference in mean scale scores for seven of eight scales, and intra class correlations were excellent/good for five scales (0.69-0.77) and moderate for three scales (0.55-0.57). CONCLUSION: English (UK) and Chinese (HK) SF-36 versions are equivalent in bilingual Singapore Chinese. Our data suggest that SF 36 scores from English- and Chinese-speaking subjects may be combined in studies using the SF-36, increasing the power and representativeness of such studies. PMID- 12113395 TI - The quality of life questionnaire for cancer patients treated with anticancer drugs (QOL-ACD): validity and reliability in japanese patients with advanced non small-cell lung cancer. AB - The quality of life questionnaire for cancer patients treated with anticancer drugs (QOL-ACD), which consists of four domains (functional, physical, mental, and psychosocial) and a global face scale, was developed as a generic questionnaire for Japanese cancer patients undergoing chemotherapy. We examined the validity and reliability of this questionnaire in Japanese patients with advanced non-small-cell lung cancer (NSCLC), who participated in two randomized phase III trials. After excluding two items, one showing low test-retest reliability and the other showing poor convergent validity for the target population, Cronbach's alpha coefficients ranged from 0.795 to 0.897 and the intra-class correlation coefficients ranged from 0.612 to 0.866. These results confirmed the high reliability of the questionnaire. The results of factor analysis provided strong support for the domain structure used in the questionnaire. Each of the four domains had a moderate to strong association with important clinical variables, such as performance status or weight loss, and correlation analysis showed that the face scale provided an appropriate measure of the global quality of life. These results indicated that the QOL-ACD is potentially useful for clinical research on Japanese patients with advanced NSCLC. PMID- 12113397 TI - G-protein beta3-subunit gene variant, blood pressure and erythrocyte sodium/lithium countertransport in essential hypertension. AB - Recently, a C825T polymorphism in the gene coding for the beta3 subunit of G proteins (GNB3) has been described in cells from patients with essential hypertension and enhanced Na+/H+ exchange activity. This study aims to evaluate the association between the 825T allele and activity of erythrocyte sodium/lithium countertransport (Na+/Li+ CT) and other sodium transport systems in red blood cells from patients with essential hypertension. A group of 77 patients (36 male, 41 female; aged 51.7 +/- 1.1 years) was studied. The maximal rates (Vmax) of Na+/Li+ CT, Na+/K+/Cl- cotransport and Na+K+ ATPase were evaluated in erythrocytes from all the patients. They were genotyped for the C825T polymorphism by a polymerase chain reaction (PCR) method, followed by digestion with BseDI. Body mass index (BMI) was higher in CT+TT patients than in CC patients (28.9 +/- 0.5 vs. 27.0 +/- 0.7 kg/m2; P=0.023). Hypertensives with the T allele (CT+TT genotypes) showed significantly higher systolic blood pressure (BP) values (156.9 +/- 2.1 vs. 148.9 +/- 2.8 mmHg; P=0.024), whereas differences in diastolic BP did not reach statistical significance (96.4 +/- 1.0 vs. 94.0 +/- 1.1 mmHg; P=0.120). No differences in the Vmax of Na+/Li+ CT between the genotypes was seen (CC: 236 +/- 19 and CT+TT 277 +/- 23 mmol/L cells per h; P=0.221). Similarly, no differences were detected in the Vmax of erythrocyte Na+/K+/Cl- cotransport and Na+K+ ATPase among the genotypes. There was no appreciable association between the G-protein beta3-subunit C825T polymorphism and erythrocyte Na+/Li+ CT and other sodium transport systems in the hypertensive patient sample studied; however, those with the T allele were more obese and had more severe systolic hypertension. PMID- 12113398 TI - Hypothesis for the influence of fixatives on the chromatin patterns of interphase nuclei, based on shrinkage and retraction of nuclear and perinuclear structures. AB - Nuclear chromatin patterns are used to distinguish normal and abnormal cells in histopathology and cytopathology. However, many chromatin pattern features are affected by aspects of tissue processing, especially fixation. Major effects of aldehyde and/or ethanol fixation on nuclei in the living state include shrinkage, chromatin aggregation and production of a 'chromatinic rim'. The mechanisms of these effects are poorly understood. In the past, possible mechanisms of fixation induced morphological change have been considered only in terms of the theoretical model of the nucleus, which involves only a random tangle of partly unfolded chromosomes contained within the nuclear membrane. Such a model provides no basis for chromatin to be associated with the nuclear envelope, and hence no obvious clue to a mechanism for the formation of the 'chromatinic rim' in fixed nuclei. In recent years, two new models of nuclear structure have been described. The nuclear membrane-bound, chromosomal-domain model is based on the discoveries of chromatin-nuclear membrane attachments and of the localisation of the chromatin of each chromosome within discrete, exclusive parts of the nucleus (the 'domain' of each partly unfolded chromosome). The nuclear matrix/scaffold model is based on the discovery of relatively insoluble proteins in nuclei, which it suggests forms a 'matrix' and modulates gene expression by affecting transcription of DNA. Here, a hypothesis for fixation-associated chromatin pattern formation based mainly on the first model but partially relying on the second, is presented. The hypothesis offers explanations of the variations of appearance of nuclei according to fixation (especially air-drying versus wet fixation with formaldehyde, glutaraldehyde or ethanol); the appearances of the nuclei of more metabolically active versus less metabolically active cells of the same type; the appearances of nuclei after fixation with osmium tetroxide; and of the marked central clearing ('egg-shell' or 'orphan Annie' appearance) of tumour nuclei of papillary carcinoma of the thyroid gland. A similar process may underlie the phenomenon of 'chromatin margination' seen in apoptosis. Various tests of the hypothesis, such as time-lapse confocal microscopy of living nuclei during fixation, are suggested. The significance of the theory is that it suggests that chromatin patterns could be investigated in terms of qualitative and quantitative aspects of nuclear components, and hence be related to the results of studies of the structure and function of nuclei in health and disease. PMID- 12113399 TI - Safranin staining of Cyclospora cayetanensis oocysts not requiring microwave heating. PMID- 12113400 TI - Lectins as targeting agents--the in vitro binding of lectins to lesions in the eye and mouth. PMID- 12113401 TI - Re-use of a stripped cDNA microarray. PMID- 12113402 TI - Lipoprotein (a) in an immigrant Indian population sample in Australia. PMID- 12113403 TI - Enteropathogenic Escherichia coli infection: history and clinical aspects. AB - Diarrhoeagenic Escherichia coli remains an important cause of diarrhoeal disease worldwide. In terms of global public health, enteropathogenic E. coli (EPEC) and enterotoxigenic E. coli are the most important. However, enterohaemorrhagic E. coli has emerged as a cause of disease in developed countries in recent years, and a number of large outbreaks have been reported. Therefore, the importance of research into diarrhoeagenic E. coli remains an important issue. EPEC is the most widespread of the diarrhoeagenic E. coli and provides a good virulence model for other E. coli infections, as well as other pathogenic bacteria. Although the virulence mechanisms of E. coli are now better understood, there remains much to be learned before effective treatments can be developed. Type III secretion mechanisms, the locus of enterocyte effacement and various toxins are all involved in the pathogenesis of the various diarrhoeagenic E. coli and may provide targets for future therapies. This review aims to provide an update on the worldwide problem of diarrhoeagenic E. coli by focusing on EPEC, and describes the history of the organism, its incidence and the clinical aspects of infection. PMID- 12113405 TI - Data in abstracts of research articles. Are they consistent with those reported in the article? PMID- 12113404 TI - Impact of host genetics on susceptibility to human Chlamydia trachomatis disease. AB - Evidence that host genetic factors play a major role in susceptibility or resistance to many infectious diseases is increasing, due to major advances in genetic epidemiological methodology. Recent human genome mapping information and the identification of a large number of candidate genes provide the tools for such studies. The information obtained is important for understanding the pathogenesis of disease and for the design of preventive and therapeutic strategies. In the study of Chlamydia trachomatis disease pathogenesis, much research focuses on how bacterial factors modulate the immune response and thus contribute to the disease process. It is likely, however, that host factors also play a role, and therefore susceptibility to disease is the result of an environmental effect set against a background of genetic factors. This review outlines the evidence for the contribution of host genetic factors to susceptibility to C. trachomatis disease in humans. PMID- 12113406 TI - Effect of staff attitudes on quality in clinical microbiology services. AB - Technical quality of the work of clinical pathology laboratories is monitored regularly by both internal and external sources. Among the factors that might affect quality, laboratory staff attitudes are rarely considered. In this study, the psychological concepts of 'job satisfaction' and 'climate' are measured among microbiology biomedical scientists in the United Kingdom. A self-report questionnaire was developed and distributed (between November 1998 and February 1999) to biomedical scientists in 161 microbiology laboratories throughout the UK From 2415 questionnaires distributed, 931 replies were received--a response rate of 39%. A separate set of questions covering customer service and participation in internal and external quality assurance schemes was sent to laboratory managers. Biomedical scientists reported lower job satisfaction than did medical technologists in a previous study in the USA. Perception of climate was influenced by several demographic factors, the most important of which being the size of the laboratory. Optimal number of staff in a department was found to be less than 30. Aggregation of climate scores from members of the same department showed that a positive laboratory climate was important for good performance in internal and external measures of technical quality. For the best service, laboratory climate must be supported by a staff perception that the department is committed to enhancing quality--a climate for laboratory quality. PMID- 12113407 TI - Bacterial enteropathogens and factors associated with seasonal episodes of gastroenteritis in Nsukka, Nigeria. AB - Each year, between April and October, many children of school age and some young adults in Nsukka, Nigeria suffer from gastroenteritis. The period covers the rainy season in this part of Africa, when manured farmland occasionally is flooded. In view of the number of people suffering diarrhoea and occasionally low grade fever, it became necessary to investigate the nature of the bacterial agents responsible. Between April and October (1996-1998), 500 loose or watery stools were collected from patients, the ages of which ranged from one month to 31 years. Stools that contained parasites were excluded from the study. Samples were cultured on 5% blood agar and 1% egg-yolk agar (both containing 10 microg/mL ampicillin), MacConkey agar, Shigella Salmonella agar and in alkaline peptone water. Bacterial growths were identified using standard bacteriological procedures. Drinking water and some fruit and vegetables prevalent during this period of the year also were cultured. Of the 500 stool samples tested, 138 (27.6%) grew a range of organisms including Aeromonas hydrophila (65 [13%]), Salmonella spp. (55 [11%]), Shigella spp. (9 [1.8%]) and enteropathogenic Escherichia coli (9 [1.8%]). Drinking water and some vegetables grew Pseudomonas aeruginosa and Enterococcus faecalis, respectively. The highest isolation rate occurred during June and July, corresponding to the period of greatest flooding of arable land. Although no enteropathogens were isolated from the fruit and vegetables examined, they contained E. faecalis--an organism found in faeces. Our findings failed to explain why 72% of the samples grew no bacterial enteropathogens. PMID- 12113408 TI - Haematological influences of potassium adaptation in normotensive and renally hypertensive Wistar rats. AB - Dietary potassium is known to cause reduction in blood pressure in several models of hypertension in human and animal studies but its haematological effects are not known. Here, experiments are designed to study the haematological effects of potassium adaptation (achieved by administering 0.75% KCl solution in drinking water for five weeks) in Wistar rats. The animals are divided into four groups comprising controls, potassium-adapted, renal hypertensive, and renal hypertensive with later adaptation to potassium. Packed cell volume (PCV) and platelet count (PC), whole blood and plasma viscosities, and platelet aggregation in the presence of sodium nitroprusside, levcromakalim, and glibenclamide, are studied. Results showed comparable PCV and PC in all groups. While relative whole blood viscosity was significantly higher (P<0.05) in the hypertensive group, relative plasma viscosity was similar in all groups. Adaptation significantly reduced (P<0.05) the tendency of platelets to aggregate to collagen. Sodium nitroprusside significantly reduced (P<0.05) the pro-aggregatory effects of collagen only in the control group. Neither of the potassium-channel modulators (levcromakalim, glibenclamide) caused any significant alteration in platelet response to collagen at the concentrations studied. Although these results suggest that potassium adaptation may not affect haemorheology, the reduced ability of platelets to aggregate--by mechanisms not clearly understood--has implications for reduced thromboembolism and the attendant cardiovascular sequelae. PMID- 12113409 TI - Clinical relevance of intracellular cytokines IL-6 and IL-12 in acute pancreatitis, and correlation with APACHE III score. AB - Pro-inflammatory cytokines are involved in the pathogenesis of acute pancreatitis (AP). Here, we measure and correlate clinically the percentages of peripheral blood mononuclear cells (PBMC) that contain interleukin (IL)-6 and IL-12 and compare these with acute physiology and chronic health evaluation (APACHE III) scores in 30 patients with AP. Severity of AP is determined according to the Atlanta criteria. Patients with severe AP (n = 15) had significantly higher IL-6 values compared to those with mild AP (n = 15). IL-12 levels correlated well with aetiological factors (alcohol and biliary pathology) in patients with AP. Correlation was seen between IL-6 value and APACHE score in severe AP. A score of 30 points was used as the cut off between mild (<30) and severe (>30) cases, with a sensitivity of 80% and specificity of 100%. Cut off percentages for IL-6- and IL-12-positive PBMCs were >25% (positive predictive value [PPV]: 100%) and >9% (PPV: 70%), respectively. Based on these results, it would seem logical to use both APACHE III score and IL-6 percentage to assess severity in patients with AP. PMID- 12113410 TI - Association between tyrosine hydroxylase polymorphisms and left ventricular structure in young normotensive men. AB - Tyrosine hydroxylase (TH) is a rate-limiting enzyme for catecholamine biosynthesis. Increased sympathetic activity is associated with an increased left ventricular (LV) mass. However, the influence of TH gene polymorphisms on LV structure and function has yet to be investigated. Here, we analyse the association of Val-81-Met and tetranucleotide TCAT repeat TH polymorphisms with LV structure and function (assessed by echocardiography) in 108 normotensive men aged < or = 35 years (mean age: 25+/-4 years) with body mass index (BMI) < or = 30 kg/m2 (mean BMI: 23+/-3 kg/m2). The distribution of genotypes was VV homozygotes (n=42), VM heterozygotes (n=49) and MM homozygotes (n=17). The Val-81 Met polymorphism showed significant linkage disequilibrium with the TCAT polymorphism (P<0.0001). No differences were seen between the subgroups with respect to age, BMI and blood pressure. Compared with the VV and VM genotypes, subjects with the MM genotype showed significantly (all P<0.05) increased LV cavity diameter (VV: 52.8+/-3.9 mm, VM: 52.9+/-3.6 mm, MM: 56.1+/-3.2 mm), global LV mass (VV: 159+/-31 g, VM: 165+/-36 g, MM: 187+/-30 g) and LV mass index (VV: 81+/-14 g/m2, VM: 84+/-17 g/m2, MM: 93+/-12 g/m2). No differences were seen between the subgroups in parameters of LV function. In addition, plasma epinephrine and norepinephrine levels were comparable in the three subgroups. The results suggest an important association between the MM genotype of Val-81-Met TH gene polymorphism and increased LV cavity dimension and mass in a young normotensive male population, indicating an important role for genetic determination of the sympathetic system in LV growth. PMID- 12113411 TI - Threonines at position 174 and 235 of the angiotensinogen polypeptide chain are related to familial history of hypertension in a Spanish-Mediterranean population. AB - This study investigates the association between the allelic distribution of two polymorphisms of the angiotensinogen (AGT) gene (T174M and M235T in the polypeptide chain) and blood pressure (BP) in a Mediterranean population in the south-west of Europe. The sample consists of 1322 participants from urban and rural areas, from the province of Albacete (218,462 inhabitants), located in the south-east of Spain. The subsample of this study, adjusted by age (over 18 years old) and sex, consists of 401 individuals. A case-control study is conducted which analyses 205 individuals from the group with the highest BP (fifth quintile) and 196 from the group with the lowest BP (first quintile). In addition, a comparative and associated analysis of these polymorphisms with BP level and family history of hypertension is carried out. The T174 allele proved to be more common in the fifth quintile group, although not statistically so. When the presence of threonine was analysed in both polymorphism positions (174 and 235), the TTTT genotype was found to be more common in the fifth quintile than in the first quintile. Moreover, the TTTT genotype was significantly more common in individuals with a family history of hypertension, indicating that it could be considered a predisposing factor to high BP in individuals from such families. In addition, the T174M-T235T genotype was more common in the first quintile group, and showed significant association (P=0.05) with the group that had no family history of hypertension. PMID- 12113412 TI - The role of laparoscopic adhesiolysis in the treatment of patients with chronic abdominal pain or recurrent bowel obstruction. AB - BACKGROUND: Major abdominal operations result in random and unpredictable scar tissue formation. Intraabdominal scar tissue may contribute to recurrent episodes of bowel obstruction, chronic abdominal pain, or both. Laparoscopic adhesiolysis may provide relief of symptoms in patients with prior abdominal surgery with chronic abdominal pain or recurrent bowel obstruction. METHODS: Between September 1996 and April 1999, 35 patients underwent laparoscopic adhesiolysis. Fifteen of the patients had adhesiolysis in conjunction with other major laparoscopic procedures and were excluded from the study. Twenty of the patients who underwent adhesiolysis only were retrospectively assessed for symptomatic relief as well as peri-operative morbidity and mortality. RESULTS: Two of 20 patients were not available for long-term follow-up. In the 18 remaining patients, laparoscopic adhesiolysis was performed on 13 patients with abdominal pain and 5 patients with recurrent bowel obstruction. The follow-up period ranged from 1 to 32 (mean 11) months. Sixteen of the 18 (88.9%) operations were completed laparoscopically. Two operations were converted to open for partial enterectomy. An additional enterotomy was repaired laparoscopically. All 3 operative complications were encountered in patients operated on during hospitalization for active bowel obstruction. No mortalities or blood transfusions occurred. One patient required rehospitalization for nonoperative management of an intraabdominal hematoma. Fourteen of the 18 (77.8%) had subjective improvement in their quality of life after operation. Only 1 patient has required repeat adhesiolysis. CONCLUSIONS: Laparoscopic adhesiolysis is a safe and effective management option for patients with prior abdominal surgery with chronic abdominal pain or recurrent bowel obstruction not attributed to other intraabdominal pathology. Laparoscopic intervention in patients with active bowel obstruction may increase the risk of operative complications. PMID- 12113413 TI - Laparoscopic sacrocolpopexy, hysterectomy, and burch colposuspension: feasibility and short-term complications of 77 procedures. AB - OBJECTIVE: To report our first cases of laparoscopic sacropexy and assess the feasibility and short-term complications. METHODS: We retrospectively studied 77 laparoscopic sacral colpopexies performed from June 1996 to May 1998. Suspension was reinforced with 2 strips of synthetic mesh. Five patients had previously undergone hysterectomy, and 4 others had experienced failure of surgery for prolapse of the uterus. RESULTS: Laparoscopy was performed in 83 women with symptomatic prolapse of the uterus. Six cases required conversion to laparotomy because of technical difficulties. All of the remaining 77 patients underwent laparoscopic sacropexy that included anterior and posterior mesh reinforcement. Subtotal laparoscopic hysterectomy was performed in 60 cases, laparoscopic Burch colposuspension in 74, and levator myorrhaphy via a vaginal approach in 55. Operative time decreased from 292 to 180 minutes as experience was gained. The main operative complications were 1 rectal and 2 bladder injuries. Three patients required reoperations for hematoma or hemorrhage. One patient complained of chronic inflammation of the cervix, and another experienced rejection of the posterior mesh 6 months after the operation. Mean follow-up was 343 days. Three other patients required reoperation, 1 for a third-degree cystocele and 2 for recurrent stress incontinence. CONCLUSION: Laparoscopic sacrocolpopexy is feasible. Operative time and postoperative complications are related to the surgeon's experience but remain comparable to those noted in laparotomy. Long term assessment is required to confirm the results of this procedure. PMID- 12113414 TI - Laparoscopic surgery in the pregnant patient--one surgeon's experience in a small rural hospital. AB - OBJECTIVES: The laparoscopic treatment of urgent surgical conditions that develop in pregnant patients has not been extensively addressed in the current literature. It is a potential issue to which surgeons, especially rural surgeons, should give careful consideration, prior to being faced with an urgent situation during the delivery process. This report details 1 surgeon's experience over a 5 year period with laparoscopic surgery in the pregnant patient, primarily laparoscopic cholecystectomy, at a small rural Nebraska hospital. METHODS: Eleven laparoscopic operations were conducted in 10 patients. RESULTS: One patient underwent 2 separate operations: cholecystectomy at 6 weeks gestation and reduction of ovarian torsion/appendectomy at 20 weeks. One patient, at term, underwent combination cesarian delivery and laparoscopic cholecystectomy. Three patients underwent laparoscopic cholecystectomy during their third trimester of pregnancy. All patients had severe signs and symptoms that threatened successful term gestation and/or failed attempts at conservative medical management aimed at delaying cholecystectomy until after delivery. One complication occurred involving uterine perforation with a blunt 10-mm port canula. No fetal injury occurred, and after initial recovery from the cholecystectomy, the baby was successfully delivered later in the pregnancy via cesarian delivery without adverse sequelae. CONCLUSION: Urgent laparoscopic operations can be carried out successfully in pregnant patients throughout their pregnancy, even in remote locations lacking immediate on-site availability of subspecialty care. The surgeon must be skilled in surgical obstetrics and well trained and experienced in advanced laparoscopic techniques. It is recommended that the same lines of communication and referral for subspecialty involvement be in place as would be required in the management of premature delivery of pregnant patients without surgically urgent disease. Such lines of communication should be developed before the actual need arises. The rural surgeon must have a plan of action well in advance of that first encounter or any subsequent complication. PMID- 12113415 TI - Laparoscopic pelvic lymphadenectomy in the surgical treatment of endometrial cancer: results of a multicenter study. AB - OBJECTIVE: To analyze the results and determine the contribution of laparoscopic pelvic lymphadenectomy in the surgical treatment of women with endometrial cancer and compare with the open technique. METHODS: A prospective multicenter study was carried out on 120 women who underwent laparoscopic surgery (96 women) and open procedures (24 women) for endometrial cancer between April 1996 and March 2000. RESULTS: Four patients whose laparoscopic surgery was completed by laparotomy were excluded from the study. The other 92 laparoscopic procedures were successfully completed. Laparoscopically assisted surgical staging (LASS) was performed based on the grade of the tumor and the depth of myometrial invasion. Sixty-seven of the patients underwent hysterectomy, bilateral salpingooophorectomy (BSO), and pelvic lymphadenectomy, and 25 women also had para-aortic lymph node sampling dissection. Eleven of these patients had positive pelvic or para-aortic nodes. The mean operating time for the laparoscopic procedure was significantly longer (173.8 min, P < 0.0001) than the time for the open procedure (135.0 min). The rate of complications was similar in both groups. The recovery time was significantly reduced (P < 0.0001). CONCLUSION: The laparoscopic approach to hysterectomy and lymphadenectomy for early stage endometrial carcinoma is an attractive alternative to the abdominal surgical approach. The advantages of laparoscopically assisted surgical staging are patient related. Because the abdominal incision is avoided, the recovery time is reduced. Laparoscopic pelvic lymph node dissection is a procedure that is appropriate, when applicable. PMID- 12113416 TI - Videoscopic Heller myotomy with intraoperative endoscopy promotes optimal outcomes. AB - BACKGROUND AND OBJECTIVES: Minimally invasive surgical techniques are applicable to achalasia, but the optimum approach to intraoperative assessment of adequacy of myotomy remains unestablished. We set out to show that videoscopic Heller myotomy with concurrent endoscopy ensures adequacy of myotomy while limiting postoperative clinically apparent reflux. METHODS: Seventy-eight consecutive patients with achalasia underwent videoscopic Heller myotomy with concomitant endoscopy between 1992 and 1998. Fundoplication was not routinely undertaken. RESULTS: Preoperative symptoms consisted of dysphagia (100%), emesis/regurgitation (68%), heartburn (58%), and postprandial chest pain (49%). Following myotomy, significant improvement (P < 0.0001) was seen in dysphagia (43%), postprandial chest pain (13%), and emesis/regurgitation (9%) at a mean follow-up of 33+/-2.2 months. Mean reflux score (scale 0 to 5) improved from 3.7+/-0.3 to 1.5+/-0.2 (P < 0.0001). Improvement in symptoms was reported in 96% of patients. Fundoplication was used in 8 patients as part of hiatus reconstruction (n = 6) or repair of esophageal perforation (n = 2). CONCLUSIONS: Intraoperative endoscopy during videoscopic Heller myotomy guides the extent and adequacy of myotomy. By utilizing a focused dissection with preservation of the natural antireflux mechanisms around the gastroesophageal junction and limiting the extent of myotomy along the cardia, postoperative reflux symptoms are minimized. We advocate concomitant endoscopy during Heller myotomy to guide myotomy and submit that routine fundoplication is clinically unnecessary. PMID- 12113417 TI - What is the real value of antireflux surgery? AB - OBJECTIVE: The purpose of this study was to evaluate the changes in esophageal physiology that are produced after laparoscopic surgery in patients with gastroesophageal reflux disease (GERD). METHODS: From May 1996 until January 2000, 13 patients with GERD underwent antireflux laparoscopic surgery. In 8 patients, preoperative manometric studies showed motility disorders characterized by a decrease in the percentage of primary peristaltic waves (32% average), a reduction in the pressure of the waves (40 mm Hg average), and a decrease in the percentage of the physiological waves (7.4% average). Laparoscopic Toupet fundoplication was the surgical procedure used in all cases, without complications and with a good postoperative course. Esophageal manometry was performed 8 weeks after the operation in 7 patients. RESULTS: The results revealed an increase in the percentage of primary peristaltic waves (76.4% average) (P = 0.05906 Wilcoxon Test); an increase in the wave pressure (57 mm Hg average) (P = 0.1056); and an increase in the percentage of the physiological waves (45.8% average) (P = 0.05906). CONCLUSION: Our final conclusion was that antireflux laparoscopic surgery, in this specific case the Toupet (partial) fundoplication, induced recovery in esophageal motility in those patients with peristaltic alterations due to reflux. This plays an important role in disease control because the recovery of esophageal peristalsis allows an increase in its emptying and reduces the possibility of esophageal damage by reflux episodes that could persist even though a fundoplication was constructed. PMID- 12113418 TI - Primary thoracoscopic evaluation of pleural effusion with local anesthesia: an alternative approach. AB - OBJECTIVE: The development of a thoracoscopically assisted technique to be performed with the patient under local anesthesia for both diagnostic and therapeutic purposes when treating pleural effusions and empyemas in high-risk surgical patients. METHODS: Twenty patients with pleural effusion or empyema who were also determined to be at high risk for complications following a thoracotomy, pleural biopsy, general anesthesia, or all of these, underwent placement of a thoracoscope while under local anesthesia followed by thoracic fluid drainage, pleural biopsy, and pleurodesis as required. Patients were retrospectively evaluated for a variety of factors including personal history, pre-existing medical conditions, and pre- and postoperative course. RESULTS: The average age of the patients was 59 years (18 to 89) with a 55% male/45% female sex distribution. Patients had this procedure as a consequence of malignancy (50%), empyema (30%), spontaneous pneumothorax (10%), bronchiectasis (50%), or heart failure (5%). The average duration of the procedure was 62 minutes (20 to 190), with an average of 861 mL of fluid drainage, and 114 mL of estimated blood loss. The tube thoracostomy was usually removed on the sixth (0 to 13) postprocedure day. This procedure was well tolerated by the patients with the majority of pain management being achieved with patient controlled analgesia (58%). The direct complication rate was 10%, with 2 patients requiring endotracheal intubation. CONCLUSION: This novel thoracoscopic procedure represents an acceptable alternative to the traditional treatment of pleural effusions and empyema with comparable outcome parameters and morbidity. This technique may eventually become the standard of care for the treatment of pleural effusions. PMID- 12113419 TI - Laparoscopic cholecystectomy: relationship of pathology and operative time. AB - OBJECTIVE: Controversy exists regarding the use and timing of laparoscopic cholecystectomy in the treatment of both acute and chronic cholecystitis. Acute advocates claim to avoid fibrosis and potential dissection injuries, whereas chronic proponents avoid poor visualization due to edema and possible conversion. This study of both acute and chronic cholecystitis cases examines the relationships between pathology, operative time, and outcome of laparoscopic cholecystectomy. METHODS: A retrospective review of medical records and pathology of acute (n = 9) and chronic (n = 62) laparoscopic cholecystectomy cases, performed by 2 surgeons from 1995 to 1999 was undertaken. Using multiple regression techniques, the relationship between operative time and age, sex, race, presenting symptoms, and degree of pathologic cholecystitis was evaluated. RESULTS: One case of acute gangrenous cholecystitis required conversion. None of the chronic cases required conversion. In single variable analysis, abnormal liver function tests, chronic inflammation, wall thickness, and number of stones were each predictive of longer operative time. However, in the multiple regression, abnormal liver function tests were the only clinical factor that remained a predictor of operative time (16 minutes longer, P = 0.05). Time from presentation to operation had no effect on operative time. Twelve patients had preoperative endoscopic retrograde cholangiopancreatography, and 4 had choledocholithiasis (acute n = 1, chronic n = 3). Two chronic patients required postoperative endoscopy for a cystic duct leak (n = 1) and choledocholithiasis (n = 1). The adjusted average operative time for acute and chronic cases was similar (93 versus 74 minutes, P > 0.05). CONCLUSION: Laparoscopic cholecystectomy can be done safely for both acute and chronic cholecystitis with similar operative times. Abnormal liver function tests are associated with longer operative time. Time lapse between presentation and operation has no effect on operative time or outcome. PMID- 12113421 TI - Laparoscopic surgery in very acute cholecystitis. AB - The objective of this study was to demonstrate the safety and feasibility of laparoscopic cholecystectomy in empyematous or gangrenous cholecystitis. During the period from August 1998 to April 2000, we operated laparoscopically on 64 patients, without any selection, in which we established, preoperatively or intraoperatively, the diagnosis of empyematous or gangrenous cholecystitis using clinical criteria (fever, leukocytosis, persistent pain, abdominal tenderness or guarding), echographic findings and intraoperative or pathological aspects of the gallbladder. The operations were performed by experienced surgeons skillful in advanced laparoscopic procedure. We concluded successfully 59 operations. The five conversions were due to dense adhesions because of previous gastric surgery in 3 cases, to the lack of recognizing the anatomy of the biliary tree in one case and to a choledoco-duodenal fistula in the last case. No mortality and a very low morbidity with a short hospital stay, were noted in our study. We consider patients with very acute cholecystitis to be candidates for a laparoscopic approach. PMID- 12113420 TI - The laparoscopic challenge of cholecystitis. AB - OBJECTIVE: To evaluate the efficacy of laparoscopic cholecystectomy (LC) in acute and chronic cholecystitis and compare it with the open procedure. METHODS: This is a 5-year retrospective analysis performed at our hospital. Surgical treatment of gallbladder disease was performed in 1003 patients. Acute cholecystitis was present in 120 (11.9%) patients, and chronic cholecystitis was present in 830 (88.1%). Acute patients underwent surgery within 72 hours of symptom onset. The patients selected for LC or open cholecystectomy (OC) depended on the severity of disease, comorbid factors, and surgeon's preference. We reviewed age, sex, operating time, length of stay, perioperative complications, conversion rates, and cost. RESULTS: Patients chosen to undergo LC for acute cholecystitis tended to be younger females. Patients treated with LC for acute or chronic cholecystitis on average had a shorter operating time and length of hospital stay when compared with patients treated with OC (P < 0.005 by ANOVA with Bonferroni). Conversion rates (CR) for all LC decreased considerably from the first to the fifth year: 9% in 1995 (10/103), 9% 1996 (22/232), 4% in 1997 (8/188), 2% in 1998 (5/226) and 2% in 1999 (5/193). CONCLUSIONS: LC appears to be a reliable and cost effective procedure for acute and chronic cholecystitis; however, the surgical approach should be chosen with caution because of the potential spectrum of technical difficulties. CR is also improving as surgeons' experience broadens. PMID- 12113422 TI - Laparoscopic total mesorectum excision. AB - The main controversy of colon-rectal laparoscopic surgery comes from its use as a cancer treatment. Two points deserve special attention: the incidence of port site tumor implantation and the possibility of performing radical cancer surgery, such as total mesorectum excision. Once these points are addressed, the laparoscopic approach will be used routinely to treat rectal cancer. To clarify these points, 32 patients with cancer of the lower rectum participated in a special protocol that included preoperative radiotherapy and laparoscopic total mesorectum excision. All data were recorded. At the same time, all data recorded from the experience of a multicenter laparoscopic group (Brazilian Colorectal Laparoscopic Surgeons - 130 patients with tumor of the lower rectum) were analyzed and compared with the data provided by our patients. Analysis of the results suggests that a laparoscopic approach allows the same effective resection as that of conventional surgery and that preoperative irradiation does not influence the incidence of intraoperative complications. The extent of lymph nodal excision is similar to that obtained with open surgery, with an average of 12.3 lymph nodes dissected per specimen. The rate of local recurrence was 3.12%. No port site implantation of tumor was noted in this series of patients with cancer of the lower rectum. PMID- 12113423 TI - The direct trocar technique: an alternative approach to abdominal entry for laparoscopy. AB - OBJECTIVE: The direct trocar technique is an alternative to Veress needle insertion and open laparoscopy for accessing the abdominal cavity for operative laparoscopy. We review our approach to abdominal entry in 1385 laparoscopies performed between September 1993 and June 2000 by our group at Stanford University Hospital, a tertiary Medical Center. METHODS: We performed a retrospective chart review of 1385 patients who underwent operative laparoscopy during the study years. The mode of abdominal entry, patient demographics, and complications were reviewed. RESULTS: The transumbilical direct trocar entry method was used in 1223 patients. In 133 patients, the Veress needle insertion technique was used. Open laparoscopy was used in 22 patients. Three (0.21%) major complicadons occurred: 1 enterotomy, 1 omental herniation, and 1 bowel hemiation. One complication was related to primary access (0.072%) in a patient who had an open laparoscopy. She sustained an enterotomy during placement of the primary trocar. The bowel was repaired laparoscopically. No trocar-related injuries occurred among the 1223 patients in whom the direct trocar entry technique was used. One patient had an omental herniation and required a repeat laparoscopy on postoperative day 2. The second patient had a repeat laparoscopy on the 12th postoperative day to repair a bowel herniation. None of our patients required a laparotomy. No vascular injuries occurred. CONCLUSION: Based on our experience, the direct trocar technique is a safe approach to abdominal entry for laparoscopic surgery. PMID- 12113424 TI - "Endoview" project of intrapartum endoscopy. AB - INTRODUCTION: The change in obstetrical practices over the last decade in favor of trials of labor in patients with uterine scars has resulted in increased incidences of uterine ruptures. Although neither repeat cesarean delivery nor a trial of labor is risk free, evidence from a large multicenter study shows vaginal birth after the cesarean (VBAC) is associated with shorter hospital stays, fewer postpartum blood transfusions, and a decreased incidence of postpartum maternal fever. The uterine rupture remains the most serious complication associated with VBAC. Factors associated with uterine rupture include excessive exposure to oxytocin, dysfunctional labor, and a history of more than 1 cesarean delivery.2 Because uterine rupture may be a life-threatening event, intrapartum surveillance and the ability to perform an emergency surgery are both necessary when trial of labor is allowed. Until now, no early symptoms pathognomonic to uterine rupture had been described. We share our experiences with the novel approach to the problem - an intrapartum endoscopy. MATERIALS AND METHODS: Endoscopic examination was accomplished by using the intraoperational fiberscope (Olympus and Endoview system (Costa Mesa, CA, USA). A gas-sterilized 25-cm long fiberscope is introduced into the amniotic cavity through the cervical canal after rupture of the membranes. The distance between the fiberscope and the object varies from 3 to 50 mm. The fiberscope has a separate channel for the fluid infusion (normal saline) throughout the procedure; the surgeon looks through the eyepiece directly and exhibits control over the flexible scope. The duration of endoscopy is less than 15 minutes. The inserting of the endoscopic device is very similar to that of insertion of an intrauterine pressure catheter. The IRB Committees of both participating institutions approved the study protocol. Twenty-eight patients with an unknown or poorly documented site of the uterine scar were included in the study. An ultrasound examination had been performed on all patients prior to endoscopy to assess fetal wellbeing and placental location. The ages of the patients ranged from 21 to 38 years. Eighteen women had 1 previous cesarean delivery, and 10 had 2. The performance of intrapartum endoscopy did not interfere with fetal monitoring; 21 fetuses were monitored externally, 7 internally. Indications for previous cesarean deliveries were as follows: fetal distress in 11 cases, failure to progress in labor in 8, placenta previa in 2, and unknown in 7. Twenty-one patients delivered vaginally; 7 had had repeat cesarean deliveries. All neonates were born in satisfactory condition. The Apgar scores at 1 minute varied from 7 to 9 and at 5 minutes from 8 to 10. The integrity of the uterine wall was assessed by manual postpartum uterine exploration in each case of vaginal delivery and by visualization and palpation of the scar site in each abdominal delivery. RESULTS: The lower uterine segment and contractile portion of the anterior uterine wall were visualized successfully in all patients. In 25 patients, the presumed scar site looked totally indistinguishable from the rest of the lower uterine segment and anterior uterine wall. Two scars were identified as vertical in 2 patients who were delivered by a repeat abdominal operation. A vertical scar appears as a groove running in a cephalad-caudad direction from the lower uterine segment into the contractile portion of the anterior uterine wall. The usefulness of the intrapartum endoscopy is best demonstrated by the following case reports (2 of 28 study cases). PMID- 12113425 TI - Gastrointestinal hemorrhage from a small bowel polypoid hemangioma. AB - OBJECTIVES: Occult gastrointestinal bleeding can originate from the foregut, midgut, or hindgut. The evaluation of the foregut and hindgut are well established. Cases that involve bleeding from the midgut present a much more significant challenge in terms of detection and treatment. Methods of evaluation include small bowel endoscopy, arteriography, and gastrointestinal contrast studies. The differential diagnosis includes arteriovenous malformations, angiodysplasia, ulcers, and small bowel tumors. We will demonstrate that both the evaluation and treatment of these lesions may be accomplished using minimally invasive techniques. METHODS: A case of occult gastrointestinal bleeding from a polypoid hemangioma located in the distal jejunum is presented. Diagnosis and treatment was accomplished using angiographic localization with laparoscopic resection. RESULTS: Laparoscopic small bowel resection after angiographic localization was successful in removing the jejunal polypoid hemangioma. The patient experienced no further gastrointestinal bleeding. CONCLUSIONS: We will discuss the technique of localization and treatment used in this unusual case. A laparoscopic approach is an appropriate and beneficial treatment modality in a bleeding midgut lesion provided the lesion can be localized preoperatively and an oncologic resection is maintained. PMID- 12113426 TI - Port-site closure: a new problem, an old device. AB - OBJECTIVE: Trocar-site incisional hernias and their complications are reported in 1% to 6% of patients. Such hernias are attributed to the difficulty of applying standard suturing techniques to wound closure. We report our experience with a simple device, the Deschamps ligature needle. METHODS: The Deschamps needle has a handle and a tip (sharp or blunt), with an opening to pass suture. The blunt tip is very effective for closing trocar sites. Disposable needles are obviously sharp, but can bend on the needle holder and break in a deep small incision. The Deschamps needle is a rigid, noncutting instrument that can be forced through fascia and peritoneum (around the surgeon's fingertip) avoiding loss of pneumoperitoneum. A full-thickness closure is accomplished. We perform closure under direct vision through the scope. Tactile sense is provided by the surgeon's finger. The last trocar site is closed in the same manner without the scope. RESULTS: We have used the Deschamps needle since 1992 in all (1400) laparoscopic procedures. We close 10-mm and 5-mm trocar sites and have not observed wound dehiscence or hernias at these sites. CONCLUSION: The Deschamps needle is effective in preventing incisional hernias and wound dehiscence. It is cost effective. Disposable, single-use devices vary in price from $30 to $75 each. The Deschamps needle is sold in Italy at approximately $35 each. Considering that it may have been in the trays of most operating rooms for years (as in our case), and the number of procedures performed, we conclude that the real cost of this instrument is almost negligible. PMID- 12113427 TI - Rabbit tissue model (RTM) harvesting technique. AB - A method for creating a tissue model using a female rabbit for laparoscopic simulation exercises is described. The specimen is called a Rabbit Tissue Model (RTM). Dissection techniques are described for transforming the rabbit carcass into a small, compact unit that can be used for multiple training sessions. Preservation is accomplished by using saline and refrigeration. Only the animal trunk is used, with the rest of the animal carcass being discarded. Practice exercises are provided for using the preserved organs. Basic surgical skills, such as dissection, suturing, and knot tying, can be practiced on this model. In addition, the RTM can be used with any pelvic trainer that permits placement of larger practice specimens within its confines. PMID- 12113428 TI - Laparoscopic-assisted operation for familial adenomatous polyposis patients-two case reports. AB - OBJECTIVE: We describe herein the results of 2 laparoscopic operations to treat patients with familial adenomatous polyposis (FAP). METHODS: Two female FAP patients, aged 32 and 29 years old, were treated with restorative proctocolectomy and total colectomy with ileorectal anastomosis (hand-assisted laparoscopic surgery), respectively. RESULTS: The operative time was 360 minutes for the restorative proctocolectomy and 150 minutes for the total colectomy with ileorectal anastomosis. The blood loss was 500 cc for the restorative proctocolectomy and minimal for the total colectomy patient. The return of bowel movements took 3 days for each patient, and no complication occurred. Patients were discharged on the 15th and 7th postoperative days. CONCLUSION: A laparoscopic approach for restorative proctocolectomy or total colectomy with ileorectal anastomosis is safe and technically feasible, and provides good cosmesis. PMID- 12113429 TI - Competency and the six core competencies. PMID- 12113431 TI - Social epidemiology. PMID- 12113432 TI - Normative vs. attitudinal considerations in breastfeeding behavior: multifaceted social influences in a developing country context. AB - The aim of the paper is to test the basic assumptions underlying the theory of reasoned action (TRA) for exclusive breastfeeding behavior taking place in the rather complicated social environment of women who have just given birth. The paper aims (i) to argue that normative rather than attitudinal considerations are more important in engaging the correct breastfeeding behavior, and (ii) to demonstrate that the TRA concept of social norm should be treated as a multi layered construct which involves several enabling factors in predicting complex behaviors such as breastfeeding. Data were collected in three phases as part of a prospective cohort follow-up design. The first phase of data collection was conducted in the hospital with mothers after the delivery. Two follow-up questionnaires were administered at the end of the first and second months. Results did not confirm the assertions of the TRA. Logistic regression models and multiple regression analyses indicated that intention and belief/attitude measures taken at the time of birth did not predict end-of-first-month full breastfeeding behavior. Overall, results revealed that intention by itself was not a strong determinant of breastfeeding unless it was conditioned by enabling factors such as social support and subjective norms regarding breastfeeding. PMID- 12113433 TI - Utilization of health facilities and trained birth attendants for childbirth in rural Bangladesh: an empirical study. AB - The majority of births in rural Bangladesh are carried out in unhygienic conditions by relatives and traditional birth attendants (TBAs). This results in a high incidence of maternal and infant mortality that could be reduced if childbirth were to occur in health centers or under the supervision of trained TBAs (TTBAs). In this paper, we examined factors associated with utilization of modern health resources for childbirth in 39 villages of Bangladesh. We followed a retrospective survey research design to collect relevant information from couples who experienced childbirth during a two-year period from July 1, 1995 to June 1997. The data indicate that slightly over 11% of the deliveries were performed by trained personnel with the rest attended by TBAs. Multivariate analysis clearly shows that delivery complications was the most significant factor determining the use of modern health care resources for childbirth, followed by parental education, and pre-natal care. We conclude that quick response to delivery complications and improved access to hospitals and TTBAs can reduce the risk of infant and maternal mortality and morbidity in rural Bangladesh. PMID- 12113430 TI - Adhesions and adhesiolysis: the role of laparoscopy. AB - BACKGROUND: Adhesions commonly result from abdominal and pelvic surgical procedures and may result in intestinal obstruction, infertility, chronic pain, or complicate subsequent operations. Laparoscopy produces less peritoneal trauma than does conventional laparotomy and may result in decreased adhesion formation. We present a review of the available data on laparoscopy and adhesion formation, as well as laparoscopic adhesiolysis. We also review current adjuvant techniques that may be used by practicing laparoscopists to prevent adhesion formation. DATABASE: A Medline search using "adhesions," "adhesiolysis," and "laparoscopy" as key words was performed for English-language articles. Further references were obtained through cross-referencing the bibliography cited in each work. DISCUSSION: The majority of studies indicate that laparoscopy may reduce postoperative adhesion formation relative to laparotomy. However, laparoscopy by itself does not appear to eliminate adhesions completely. A variety of adjuvant materials are available to surgeons, and the most recent investigation has demonstrated significant potential for intraperitoneal barriers. Newer technologies continue to evolve and should result in clinically relevant reductions in adhesion formation. PMID- 12113434 TI - Nurse-physician communication and quality of drug use in Swedish nursing homes. AB - The objective was to explore the impact of the quality of nurse-physician communication on the quality of psychotropic drug use in Swedish nursing homes, while controlling for resident mix and other nursing home characteristics. Data were collected from a sample of 36 Swedish nursing homes providing care for 1645 residents. Drug use data, along with residents' demographic characteristics, were obtained from residents' medication administration lists. Ward nurses reported other residents' characteristics (e.g. diagnosis and frequency of behavioral problems), and facility characteristics were obtained from head nurses. The quality of drug use was assessed and cross-sectional relationships among study variables were compared. Outcome measures included two drug use quality scores reflecting selection of drug and polymedicine. To assess behavioral problems, a list of the most commonly observed problems was created through a number of steps, including focus groups in the target population. Furthermore, a valid and reliable scale for assessing communication quality was developed. This measure was included in a survey administrated to nurses in the 36 facilities. There was a remarkable variation in the quality of drug use according to the two drug measures. As predicted, the quality of drug use was positively associated with the quality of nurse-physician communication and with regular multidisciplinary team discussions addressing drug therapy and negatively associated with prevalence of behavioral disturbances among residents. Facility size, level of staffing, resident's diagnostic mix, and demographic mix were unrelated to the two drug quality measures. Future efforts to improve the quality of drug use in long term facilities should consider ways of improving communication skills and communication routines among health care professionals. PMID- 12113435 TI - Individual and neighbourhood determinants of social participation and social capital: a multilevel analysis of the city of Malmo, Sweden. AB - The aim of this study was to analyse the impact of neighbourhood on individual social capital (measured as social participation). The study population consisted of 14,390 individuals aged 45-73 that participated in the Malmo diet and cancer study in 1992-1994, residing in 90 neighbourhoods of Malmo, Sweden (population 250,000). A multilevel logistic regression model, with individuals at the first level and neighbourhoods at the second level, was performed. The study analysed the effect (intra-area correlation and cross-level modification) of the neighbourhood on individual social capital after adjustment for compositional factors (e.g. age, sex, educational level, occupational status, disability pension, living alone, sick leave, unemployment) and, finally, one contextual migration factor. The prevalence of low social participation varied from 23.0% to 39.7% in the first and third neighbourhood quartiles, respectively. Neighbourhood factors accounted for 6.3% of the total variance in social participation, and this effect was reduced but not eliminated when adjusting for all studied variables (-73%), especially the occupational composition of the neighbourhoods ( 58%). The contextual migration variable further reduced the variance in social participation at the neighbourhood level to some extent. Our study supports Putnam's notion that social capital, which is suggested to be an important factor for population health and possibly for health equity, is an aspect that is partly contextual in its nature. PMID- 12113436 TI - The relative influence of individual, social and physical environment determinants of physical activity. AB - Environmental determinants of health are receiving growing attention in the literature, although there is little empirical research in this area. The Study on Environmental and Individual Determinants of Physical Activity (known as the SEID project) was a social ecological project that examined the relative influence of individual, social environmental and physical environmental determinants of recreational physical activity. It involved a community survey of 1803 healthy workers and home-makers aged 18-59 years living in a 408 km2 area of metropolitan Perth, Western Australia. Physical environmental determinants were mainly conceptualised as spatial access to popular recreational facilities. Overall, 59% of respondents exercised as recommended. Recreational facilities located near home were used by more respondents than facilities located elsewhere. The most frequently used facilities were informal: the streets (45.6%); public open space (28.8%) and the beach (22.7%). The physical environment's directs the influence on exercising as recommended was found to be secondary to individual and social environmental determinants. Nevertheless, accessible facilities determined whether or not they were used and in this way, support and enhance the achievement of recommended levels of physical activity behaviour by providing opportunities. The results suggest that access to a supportive physical environment is necessary, but may be insufficient to increase recommended levels of physical activity in the community. Complementary strategies are required that aim to influence individual and social environmental factors. Given the popularity of walking in the community, it is recommended that greater emphasis be placed on creating streetscapes that enhance walking for recreation and transport. PMID- 12113437 TI - Asking patients about violence: a survey of 510 women attending social and health services in Trieste, Italy. AB - Violence against women is frequent, and has serious consequences for their physical and mental health. Until now, a common response of health services and professionals to victims has been the denial of the violence. The aims of this study were to estimate the prevalence of present and past violence among women attending social and health services in Trieste (Italy) for any reasons, and to evaluate the feasibility of asking them about violence. Five public health care facilities were involved: a hospital-based Emergency Department, two "Consultorio Familiare", and two community-based Social service centers. The final sample consisted of 510 women; the response rate among eligible women was 76% across all facilities. The study revealed a high prevalence of different kinds of violence, mostly perpetrated by men well known to the victim. Among the women interviewed, 10.2% had experienced physical/sexual violence in the last 12 months, regardless of perpetrator. Violence by a male partner or former partner, occurred in 6.4% of women; by other relatives, 1.6% of women; and by "other" persons 3.3% of women. The results demonstrated the feasibility of "asking about violence" in all cases; moreover, those women who had been abused were often eager to talk about it. A systematic approach to the issue of violence is a necessary condition for responding appropriately to the needs of women who have experienced or are experiencing violence. Nevertheless, to avoid the replication of a common bias against victims of violence, health and social personnel should be appropriately trained before "asking all cases" becomes a policy within health and social services. PMID- 12113438 TI - 'The public is too subjective': public involvement at different levels of health care decision making. AB - There are a number of impulses towards public participation in health care decision making including instrumentalist, communitarian, educative and expressive impulses and the desire for increased accountability. There has, however, been little research looking systematically at the public's preferences for being involved in particular types of rationing decisions, nor indeed, has there been a critical examination of the degree of involvement desired by the public. The research reported here uses findings from focus groups and in-depth interviews to explore these questions. Eight focus groups were conducted with a total of 57 informants, four amongst randomly selected members of the public and four with informants from health and non-health related organisations. Nineteen interviews were conducted to allow the elaboration of focus group comments, to probe views more deeply and to pursue emerging themes. The findings show variations in the willingness of members of the public to be involved in health care decisions and consistency across the different forms of the public as represented by the focus groups with randomly selected citizens and pre-existing organisations. There was a strong desire in all the groups for the public to be involved both at the system and programme levels, with much less willingness to be involved at the individual level. At the system and programme levels informants generally favoured consultation, without responsibility for decisions, but with the guarantee that their contribution would be heard and that decisions taken following consultation would be explained. At the patient level informants felt that the public should participate only by setting criteria for deciding between potential beneficiaries of treatment. The public has much to contribute, particularly at the system and programme levels, to supplement the inputs of health care professionals. PMID- 12113439 TI - "If there were a war tomorrow, we'd find the money": contrasting perspectives on the rationing of health care. AB - In spite of the substantial academic effort being devoted to the subject of health care rationing, there is little clarity about the views of those working in health care who have to implement rationing nor about the views of citizens who are (potentially) affected by the rationing of care. This paper reports the findings of a study conducted using focus groups and semi-structured interviews to explore and compare beliefs about rationing among citizens and those with a role in the health service (service informants) within the context of health care provision in the UK. Citizen and service informants both identified external pressures on the resources available for health care including technological improvement, the ageing population and increasing public expectations. Citizens, however, also identified such factors as the political choice to provide insufficient funds to the health service and the wasteful use of resources that are available. The predominant view was that these latter factors were amenable to change and thus that there are alternatives to the rationing of care. Some citizens accepted that some health care rationing might be necessary. Service informants, on the other hand, were cynical about prospects of increased funding and viewed further reductions in management as untenable. For them, rationing was an inevitability to be managed. A number of these informants felt that rationing should become more explicit, suggesting that openness made rationing both more democratic and more practical. Others, however, believed that explicit rationing would have a number of uncomfortable implications. The paper concludes by suggesting that if rationing is to become more explicit, its inevitability, as perceived by those working within health care, will have to be communicated to citizens. PMID- 12113440 TI - The paradoxical use of interpreting in psychiatry. AB - Changes in the official status of African languages in South Africa suggested an examination of the impact of multi-lingualism on the practice of institutional psychiatry. For a range of theoretical and institutional reasons, a 'language gap' between clinician and patient can be rendered irrelevant in terms of the routine production of psychiatric texts in which 'symptoms' are described and 'cases' are constructed. In contrast to the way in which the role of interpreting is obscured in some hospital settings, it is highlighted in forensic settings. Here the extent of the dependency of the clinician on the interpreter is made more visible. Through this ethnographic exploration of the institutional management of multi-lingualism, the status of 'the patient who requires interpreting' emerges as an institutional construct, being determined in large measure by the routines of institutional practice. The requirements of the institution that the patient move through the process of a hospital admission, and the different requirements of each stage of this process, inform the decision as to whether interpreting is necessary. Furthermore, the differing requirements of the members of the multi-disciplinary teams renders the status of 'the patient who requires interpreting' as variable and contested. Through this analysis the institutional management of multi-lingualism emerges as a site at which discourses of race in psychiatry are reproduced. PMID- 12113441 TI - Social integration of children with epilepsy in rural India. AB - Only a small fraction of people with epilepsy in developing countries has access to medical facilities. Even with effective treatment, their psychosocial needs are often overlooked in the absence of obvious disability. In rural areas, community-based rehabilitation programmes assist in the integration of people with disabilities into employment and the community. However, the functional impairment associated with epilepsy is not well recognised in intervention programmes in developing countries. We report, for the first time, the social activities of children with epilepsy and their peers in rural India. We employed a cross-sectional design using a new age and sex-specific social activity questionnaire. Population screening in the context of a community-based rehabilitation programme identified 88 children with epilepsy and 250 randomly selected controls. A trained interviewer administered the questionnaire to mothers in Bengali. Girls' activities were principally domestic, whilst boys' were mostly outdoors and involved peers. All groups of children with epilepsy had significant social deficits, equally for boys and girls in the age range from 2 to 18 years (p < 0.05). Boys with epilepsy had limited peer group activities, and parents conferred fewer responsibilities to school age and adolescent children compared to controls. The nature and degree of deficits were beyond the constraints imposed by neurological impairment. Our findings in pre-schoolers were consistent with parental attitudes of overprotection found in previous research. We conclude that social integration needs active and early promotion among children with epilepsy. The assessment of remediable risk and protective factors in the family and community is an important practical area for research in community-based rehabilitation. PMID- 12113442 TI - Drug utilisation and self-medication in rural communities in Vietnam. AB - Reportedly 40-60% of people in Vietnam depend on self-medication. To assess the current situation of self-medication practices as compared with medication given by health professionals in rural areas in Vietnam, we conducted a cross sectional survey at household level. A total of 505 women with at least one child younger than 5 years of age were interviewed in their homes about their drug utilisation practices and attitudes toward medication, by using structured questionnaires. Of the 505 households, 138 stocked drugs for anticipated illness in the future. A total of 96 different antibiotics (in terms of generic type) were kept at 76 households. These antibiotics were kept mainly for coughs and diarrhoea. The self medication group was twice as likely to use antibiotics than the other group. In addition, self-medication practice was increased when a mother kept medicines in the house. This study revealed that mistaken beliefs about medicines and undesirable attitudes toward medication were prevalent. Mothers used antibiotics as if such drugs were panaceas. In this context, there was insufficient public health education, no control over pharmaceutical promotion, and no efficient drug policy and regulation. More attention should be given to consumers and patients as the ultimate users of drugs so that they can access accurate information, assess the reliability of information and ask necessary questions. PMID- 12113443 TI - Science writers' reactions to a medical "breakthrough" story. AB - In numerous incidences, the news coverage of medical research has incited unjustified optimism or fear. The medical literature provides an archive of the scientific community's condemnation of these misleading reports, but little is known about how they are judged by newsmakers. This study explored science writers' reactions to a controversial New York Times story that inflated the hopes of thousands of cancer patients. More than 60 science writers in the US, Canada, and Great Britain participated in a 12-day email discussion triggered by the Times article. We analyzed 255 of these email postings and coded (1) positive and negative critiques of the Times story, (2) references to the article's repercussions including the creation of false hope, (3) attributions of responsibility for the resulting public misunderstanding, and (4) suggestions to improve the public's comprehension of medical research news. The participating science writers generally responded negatively to the controversial article: 83% of the critiques were unfavorable. In addition, the science writers in the sample were cognizant and concerned about the impact of their work on the public, and accepted the largest share of the responsibility for the false hope created by the news coverage of medical research. Finally, the suggestions offered by respondents to improve the public's understanding of medical research news were similar to those proposed by the scientific community. Thus, some commonality exists between how scientists and science writers believe the news coverage of medical research could be improved. PMID- 12113444 TI - International health in the 21st century: trends and challenges. PMID- 12113445 TI - Social inequalities in health within countries: not only an issue for affluent nations. AB - While interest in social disparities in health within affluent nations has been growing, discussion of equity in health with regard to low- and middle-income countries has generally focused on north-south and between-country differences, rather than on gaps between social groups within the countries where most of the world's population lives. This paper aims to articulate a rationale for focusing on within- as well as between-country health disparities in nations of all per capita income levels, and to suggest relevant reference material, particularly for developing country researchers. Routine health information can obscure large inter-group disparities within a country. While appropriately disaggregated routine information is lacking, evidence from special studies reveals significant and in many cases widening disparities in health among more and less privileged social groups within low- and middle- as well as high-income countries: avoidable disparities are observed not only across socioeconomic groups but also by gender, ethnicity, and other markers of underlying social disadvantage. Globally, economic inequalities are widening and, where relevant information is available, generally accompanied by widening or stagnant health inequalities. Related global economic trends, including pressures to cut social spending and compete in global markets, are making it especially difficult for lower-income countries to implement and sustain equitable policies. For all of these reasons, explicit concerns about equity in health and its determinants need to be placed higher on the policy and research agendas of both international and national organizations in low-, middle-, and high-income countries. International agencies can strengthen or undermine national efforts to achieve greater equity. The Primary Health Care strategy is at least as relevant today as it was two decades ago: but equity needs to move from being largely implicit to becoming an explicit component of the strategy, and progress toward greater equity must be carefully monitored in countries of all per capita income levels. Particularly in the context of an increasingly globalized world, improvements in health for privileged groups should suggest what could, with political will, be possible for all. PMID- 12113446 TI - Putting equity in health back onto the social policy agenda: experience from South Africa. AB - Over the past decade, international health policy debates have been dominated by efficiency considerations. There has been a recent resurgence of interest in health equity, including consideration of the notions of vertical equity and procedural justice. This paper explores the possible application of these notions within the context of South Africa, a country in which inequities in income and social service distribution between 'racial' groups were systematically promoted and entrenched during four decades of minority rule, guided by apartheid and related policies. With the transition to a democratic government in 1994, equity gained prominence on the South African social policy agenda. Health equity has been awarded a particularly high priority, not least of all because the health sector is seen as vehicle for achieving rapid equity gains. In addition, many of the other equity-promoting social sector policies (such as improved access to housing and water and sanitation services) have been motivated on the basis of their potential health equity gains. The South African experience since 1994 provides useful insights into factors which may facilitate or constrain health equity progress. In particular, the constitutional entitlement to health and civil society action to maintain health equity's place on the social policy agenda are seen as important facilitating factors. Certain health sector programmes have also been developed which are intended preferentially to benefit those who have been historically dis-advantaged, and which thus support vertical equity goals. However, there have been no efforts to promote cross-subsidisation between the private and public health sectors, and initial efforts to promote coherency in social policies (through the Reconstruction and Development Programme) appear not to have been sustained. In addition, macro-economic policies (particularly the highly ambitious budget deficit reduction targets of the government) are likely to undermine some of the equity-promoting social policy initiatives. Most importantly, the potential inter-relationship of vertical equity and procedural justice goals has not been adequately recognised. As a result, and despite policy rhetoric, this paper concludes that health equity goals are critically dependent on the central involvement of the dis-advantaged in decision-making about who should receive priority, what services should be delivered and how equity-promoting initiatives should be implemented. PMID- 12113447 TI - Staking a claim for claims: a case study of resource allocation in Australian Aboriginal health care. AB - There have been numerous ways in which the notion of equity has been put forward in the literature. This reflects the fact that equity is essentially driven by values and is therefore subject to individual interpretation and preferences. Deciding between these various value judgements is however outside the scope of economic analysis, as conventionally defined. This poses a problem for the examination of issues of resource allocation in Aboriginal health services in Australia, where equity, very clearly, has a role to play. One possibility for moving forward on this issue is the adoption of a 'claims' approach where the emphasis is on the explicit recognition of the values to be employed in the 'equitable' allocation of resources. This involves teasing out the principles by which, under various approaches, resources are allocated differentially across groups (e.g. under resource allocation formulae, the criterion of 'need' as measured by SMRs can be viewed to be a basis for a 'claim' over resources). The commonly cited 'basic needs approach' is then used in the paper as a case in point to illustrate how such underlying principles may be identified and then assessed. In relation to the issue of equity in Aboriginal health services, there are a number of possible standards for equity which seem to have a significant degree of community acceptance. The paper discusses ways in which they can be applied to the problem of deciding how to allocate resources in Aboriginal health. PMID- 12113448 TI - Reshaping the state from above, from within, from below: implications for public health. AB - The modern state is being reshaped by multiple forces acting simultaneously. From above, the state is actively constrained by agreements promoted by international agencies and by the power of multinational corporations. From within, the state is being reshaped by increasing trends toward marketization and by problems of corruption. From below, the state's role is being diminished by the expansion of decentralization and by the rising influence of non-governmental organizations. This article explores these three sets of processes--from above, from within, and from below--and suggests some implications for public health. Public health professionals require an understanding of the changing nature of the state, because of the consequences for thinking about the metaphors, solutions, and strategies for public health. PMID- 12113449 TI - Regulating incentives: the past and present role of the state in health care systems. AB - The desire of national policymakers to encourage entrepreneurial behavior in the health sector has generated not only a new structure of market-oriented incentives, but also a new regulatory role for the State. To ensure that entrepreneurial behavior will be directed toward achieving planned market objectives, the State must shift modalities from staid bureaucratic models of command-and-control to more sensitive and sophisticated systems of oversight and supervision. Available evidence suggests that this structural transformation is currently occurring in several Northern European countries. Successful implementation of that shift will require a new, intensive, and expensive strategy for human resources development, raising questions about the financial feasibility of this incentives-plus-regulation model for less-well-off CEE/CIS and developing countries. PMID- 12113450 TI - Improving basic health service delivery in low-income countries: 'voice' to the poor. AB - Public social services, such as basic health care, represent the effective option for the poor, especially in the rural areas of low-income countries. The quality of such services are at present extremely deficient, largely due to resource constraints and lack of political will to make them function effectively. The state can no longer provide the comprehensive services it has in the past and which were highly successful in a number of 'high-achieving' developing countries. Yet, the state must turn priority attention to providing public services for the poor, in order to close the widening gap between rich and poor. It needs to do this in partnership with the population it aims to reach, through effective linkage with grass-roots organizations and with the support of non governmental organizations. Giving 'voice' and participation to the population can not only increase the resource base for public services, but can also significantly improve the accountability of providers and lead to a cost effective option for the poor. PMID- 12113451 TI - Financing reforms of public health services in China: lessons for other nations. AB - Financing reforms of China's public health services are characterised by a reduction in government budgetary support and the introduction of charges. These reforms have changed the financing structure of public health institutions. Before the financing reforms, in 1980, government budgetary support covered the full costs of public health institutions, while after the reforms by the middle of the 1990s, the government's contribution to the institutions' revenue had fallen to 30-50%, barely covering the salaries of health workers, and the share of revenue generated from charges had increased to 50-70%. These market-oriented financing reforms improved the productivity of public health institutions, but several unintended consequences became evident. The economic incentives that were built into the financing system led to over-provision of unnecessary services, and under-provision of socially desirable services. User fees reduced the take-up of preventive services with positive externalities. The lack of government funds resulted in under-provision of services with public goods' characteristics. The Chinese experience has generated important lessons for other nations. Firstly, a decline in the role of government in financing public health services is likely to result in decreased overall efficiency of the health sector. Secondly, levying charges for public health services can reduce demand for these services and increase the risk of disease transmission. Thirdly, market-oriented financing reforms of public health services should not be considered as a policy option. Once this step is made, the unintended consequences may outweigh the intended ones. Chinese experience strongly suggests that the government should take a very active role in financing public health services. PMID- 12113452 TI - Research capacity strengthening in the South. AB - Active promotion of evidence-based decision-making at all levels of the health field is a necessary step in the direction of improving the health of the population. Recent studies have shown that the burden of disease in developing countries is high particularly the burden of infectious, communicable and non communicable diseases and health problems of mothers and children. There is presently, a mismatch between this increased disease and health burden and the technical and human capacity of developing countries to use existing knowledge and to generate new knowledge to combat these diseases and health problems. It is therefore necessary to assist developing countries to build indigenous research capability so they can undertake studies in their own national settings the results of which will lead to the development of appropriate control strategies in their countries. Building indigenous research capacity will enable developing country scientists to translate results of studies carried out elsewhere into their individual national settings. Eventually results of such studies will increase the global knowledge base about the particular health problems and contribute to finding appropriate solutions to them. The research will, finally, increase knowledge-based decision-making by their health leadership of the country. This paper has set out to describe some experiences in capacity strengthening over the last few decades and to propose from these, mechanisms for building these capacities in a sustainable manner. This paper has described the steps in capability strengthening with special emphasis on identification of trainees, their training and deployment on return. The paper has described mechanisms of research sustainability including creation of suitable career structures, remuneration of researchers and the importance of building up suitable infrastructure for research to meet increasing demands and competence. The place of partnerships South-South, South-North and networking has been stressed. Finally, the paper calls for greater involvement by policy makers in developing countries in the entire capacity building process. They should set highly focussed research priorities, identify competence not already existing and proceed to fill these gaps along the lines described. PMID- 12113453 TI - Placing gender at the centre of health programming: challenges and limitations. AB - In this paper we argue that a gender analysis is fundamental to health and health planning. We begin with a definition of gender and related concepts including equity and equality. We discuss why gender is key to understanding all dimensions of health including health care, health seeking behaviour and health status, and how a gender analysis can contribute to improved health policies and programming. Despite the many reasons for incorporating gender issues in health policies and programmes many obstacles remain, including the lack of attention to gender in the training of health professionals and the lack of awareness and sensitivity to gender concerns and disparities in the biomedical community. We argue that the key to placing gender values firmly in place in Health for All renewal is a change in philosophy at all levels of the health sector and suggest ways in which such a change can be implemented in the areas of policy, research, training and practical programmes and interventions. PMID- 12113454 TI - The rhetoric of sector-wide approaches for health development. AB - The past decade has witnessed an increasing concerns over the effectiveness of project-based development assistance and the promotion of sector-wide approaches (SWAps) to health as a means to increase donor collaboration, consolidate local management of resources and undertake the policy and systems reform necessary to achieve a greater impact on health issues. The concept has gained the support of both the World Bank and the World Health Organisation. as well as key bilateral donors, and dominates current initiatives in development assistance for health. This paper examines the proposal of SWAps as rhetoric, and seeks to understand how that rhetoric functions, despite the variable application of its constituent elements and the range of contexts in which it operates. PMID- 12113455 TI - Expression of functional recombinant human lysozyme in transgenic rice cell culture. AB - Using particle bombardment-mediated transformation, a codon-optimized synthetic gene for human lysozyme was introduced into the calli of rice (Oryza sativa) cultivar Taipei 309. The expression levels of recombinant human lysozyme in the transformed rice suspension cell culture approached approximately 4% of total soluble protein. Recombinant human lysozyme was purified to greater than 95% homogeneity using a two-step chromatography process. Amino acid sequencing verified that the N-terminus of the mature recombinant human lysozyme was identical to native human lysozyme. This indicates that the rice RAmy3D signal peptide was correctly cleaved off from the human lysozyme preprotein by endogenous rice signal peptidase. Recombinant human lysozyme was found to have the same molecular mass, isoelectric point and specific activity as native human lysozyme. The bactericidal activity of recombinant human lysozyme was determined by turbidimetric assay using Micrococcus lysodeikticus in 96-well microtiter plates. The bactericidal activity of lysozyme on gram-negative bacteria was examined by adding purified lysozyme to mid-log phase cultures of E. coli strain JM109. In this study, significant bactericidal activity was observed after E. coli cells were exposed to recombinant human lysozyme for 60 min. Both native and recombinant human lysozyme displayed the same thermostability and resistance to degradation by low pH. The potential for using rice-derived lysozyme as an antimicrobial food supplement, particularly for infant formula and baby foods, is discussed. PMID- 12113456 TI - Inducible, reversible hair loss in transgenic mice. AB - Telogen effluvium is a common type of hair loss. Although the morphological changes associated with telogen effluvium have been well characterized, the underlying molecular mechanisms remain unknown, and no animal models have been developed. We report here that inducible transgenic mice expressing high levels of the transcription factor, tTA (tetracycline transactivator), plus a reporter luciferase gene, show a reversible hair loss phenotype. Skin of these mice exhibits an increase in the number of hair follicles at the telogen phase, but a decreased number of follicles at the anagen phase. These changes resemble skin pathology seen in patients with telogen effluvium, which suggests that the inducible transgenic mice may be useful as a model for this disorder. Moreover, since overexpression of several other transgenes failed to cause skin pathology, the present findings also indicate types of molecular abnormalities that may cause reversible hair loss. PMID- 12113457 TI - Expression of an antisense GIGANTEA (GI) gene fragment in transgenic radish causes delayed bolting and flowering. AB - A late-flowering transgenic radish has been produced by the expression of an antisense GIGANTEA (GI) gene fragment using a floral-dip method. Twenty-five plants were dipped into a suspension of Agrobacterium carrying a 2.5 kb antisense GI gene fragment from Arabidopsis, along with the gusA and bar reporter genes, all under the control of a CaMV 35S promoter. From a total of 1462 seeds harvested from these floral-dipped plants, 16 Basta-resistant T1 plants were found to have GUS activity (transformation efficiency of 1.1%). Southern analysis confirmed the integration of one or two copies of the gusA gene in these herbicide-resistant plants. Expression of the GI gene in T1 plants was much reduced compared to both wildtype plants and plants transformed with pCAMBIA3301 (positive control). In the progenies of eleven T1 plants analysed (T2 generation), all lines showed a significant delay in both bolting and flowering times compared to wildtype and positive control plants, and that, the level of GI transcript was inversely proportional to the time of bolting and flowering. At a maximum, bolting and flowering times were delayed by 17 and 18 days respectively, compared to wildtype plants (in positive control plants, the delay was 23 and 26 days, respectively). Ten of the 11 lines exhibited a significant reduction in plant height compared to wildtype and positive control plants. This study provides evidence that down-regulation of the GI gene by co-suppression could delay bolting in a cold-sensitive long-day (LD) plant. Production of late flowering germplasms of radish may allow this important crop to be cultivated over an extended period and also provide further food to the famine countries of S/E Asia. PMID- 12113458 TI - Temporal and spatial expression of biologically active human factor VIII in the milk of transgenic mice driven by mammary-specific bovine alpha-lactalbumin regulation sequences. AB - Hemophilia A is one of the major inherited bleeding disorders caused by a deficiency or abnormality in coagulation factor VIII (FVIII). Hemophiliacs have been treated with whole plasma or purified FVIII concentrates. The risk of transmitting blood-borne viruses and the cost of highly purified FVIII are the major factors that restrict prophylaxis in hemophilia therapy. One of the challenges created by the biotechnology revolution is the development of methods for the economical production of highly purified proteins in large scales. Recent developments indicate that manipulating milk composition using transgenesis has focused mainly on the mammary gland as a bioreactor to produce pharmaceuticals. In the present study, a hybrid gene containing bovine alpha-lactalbumin and human FVIII cDNA was constructed for microinjection into the pronuclei of newly fertilized mouse eggs. The alphaLA-hFVIII hybrid gene was confirmed to be successfully integrated and stably germ-line transmitted in 12 (seven females/five males) lines. Western-blot analysis of milk samples obtained from eight of the transgenic founders and F1 offspring indicated that the recombinant hFVIII was secreted into the milk of the transgenic mice. The concentrations of rFVIII ranged from 7.0 to 50.2 microg/ml, over 35-200-fold higher than that in normal human plasma. Up to 13.4 U/ml of rFVIII was detected in an assay in which rFVIII restored normal clotting activity to FVIII-deficient human plasma. PMID- 12113459 TI - Anther-specific expression of ipt gene in transgenic tobacco and its effect on plant development. AB - Isopentenyl transferase (ipt) gene from Agrobacterium tumefaciens T-DNA was placed under the control of a TA29 promoter which expresses specifically in anther. The chimeric TA29-ipt gene was transferred to tobacco plants. During flowering, mRNA of the ipt gene in the anthers of the transgenic plants accumulated and the level of iPA + iPs increased 3-4-fold in the leaves, petals, pistils, and stamens compared with those in the wild type plants. This cytokinin increase affected various aspects in development indicating that the alterations of endogenous cytokinin level by using anther-specific expression of the TA29-ipt gene affected morphology, floral organ systems and reproductivity of the transgenic plants. PMID- 12113460 TI - An ES-like cell line from the marine fish Sparus aurata: characterization and chimaera production. AB - Embryonic stem (ES) cells provide a unique tool for cell-mediated gene transfer and targeted gene mutations due to the possibility of in vitro selection of desired genotypes. When selected cells contribute to the germ line in chimaeric embryos, transgenic animals may be generated with modified genetic traits. Though the ES cell approach has up to now been limited to mice, there is an increasing interest to develop this technology in both model and commercial fish species, with so far promising results in the medaka and zebrafish. In this study, we present evidence regarding a long-term stable cell line (SaBE-1c), derived from embryonic cells of the aquaculture marine fish Sparus aurata which has been characterized for (i) cell proliferation, (ii) chromosome complement, (iii) molecular markers, and (iv) in vitro tests of pluripotency by alkaline phosphatase (AP) staining, telomerase activity, and induced cell differentiation. These cells have proved their pluripotent capacities by in vitro tests. Furthermore, we have demonstrated their ability to produce chimaeras and to contribute to the formation of tissues from all three embryonic germ layers. These features suggest that SaBE-1c cells have the potential for multiple applications for the ES technology in fish, with the added value of originating from an economically important species. PMID- 12113461 TI - Plant-specific promoter sequences carry elements that are recognised by the eubacterial transcription machinery. AB - During evolution the promoter elements from prokaryotes and eukaryotes have developed differently with regard to their sequence and structure, implying that in general a transfer of eukaryotic promoter sequences into prokaryotes will not cause an efficient gene expression. However, there have been reports on the functionality of the 35S promoter from cauliflower mosaic virus (CaMV) in bacteria. We therefore decided to experimentally investigate the capability of plant promoter sequences to direct gene expression in various bacteria. Accordingly, we tested ten different plant-specific promoters from Solanum tuberosum, Nicotiana tabacum, CaMV, Agrobacterium tumefaciens, and A. rhizogenes for their ability to initiate transcription in five different eubacterial species (Escherichia coli, Yersinia enterocolitica, A. tumefaciens, Pseudomonas putida, and Acinetobacter sp. BD413). To monitor the strength of the plant-specific promoters in bacteria we created fusions between these promoters and the coding region of the luciferase genes from Vibrio harveyi and measured the luminescence in the bacteria. Heterologous gene expression was observed in 50% of the combinations analysed. We then mapped the transcription start site caused by one of the plant-specific promoters, the ST-LS1 promoter from S. tuberosum, in these bacterial species. The location of the mapped transcription start site indicated that the sequences of the plant promoter themselves were recognised by the bacterial transcription apparatus. The recognition of plant-specific promoter sequences by the bacterial RNA polymerase was further confirmed by site-directed mutagenesis of the ST-LS1 promoter and the analysis of the effects of the mutations on the strength of gene expression in E. coli. Using these mutants in our reporter assays we could localise the sequences of the ST-LS1 promoter serving as -10 region in E. coli. The results of our study show that promoter sequences are much less specific than is generally assumed. This is of great importance for our knowledge about the evolution of gene expression systems and for the construction of optimised expression vectors. PMID- 12113463 TI - E. coli chromosomal DNA in a transgene locus created by microprojectile bombardment in tobacco. PMID- 12113462 TI - The tomato RNA-directed RNA polymerase has no effect on gene silencing by RNA interference in transgenic mice. AB - Double-stranded RNA (dsRNA) has been shown to interfere with the function of specific genes in various invertebrate species. The application of dsRNA interference (RNAi) in vertebrates (zebrafish and mouse) is still limited to embryos and it is not clear whether the method is generally applicable. Using a transgenic mouse model we investigated whether a stably inherited dsRNA introduced as a transgene can interfere with the expression of a specific target gene in erythroid tissue during development. In our globin gene system we do not observe any specific RNA interference. We, therefore, also introduced another gene that may be involved in a mechanism of post transcriptional gene silencing (PTGS), namely RNA-dependent RNA polymerase (RdRP) that was proposed to be involved in producing RNAs that trigger PTGS in plants. However, even though the tomato RdRP is catalytically active in erythroid tissue, no RNAi was observed. PMID- 12113464 TI - Selection of transgenic Xenopus laevis using antibiotic resistance. AB - We previously established lines of transgenic Xenopus laevis expressing green fluorescent protein (GFP) or GFP fusion proteins in the rod photoreceptors of their retinas under control of the X. laevis opsin promoter, which permits easy identification of transgenic animals by fluorescence microscopy. However, GFP tags can alter the properties of fusion partners, and in many circumstances a second selectable marker would be useful. The transgene constructs we used also encode a gene that confers resistance to the antibiotic G418 in cultured mammalian cells. In this study, we show that F2 transgenic offspring of these animals are more resistant to G418 toxicity than their non-transgenic siblings, as are primary transgenic X. laevis. G418 resistance can be used as a selectable marker in transgenic X. laevis, and possibly other aquatic transgenic animals. PMID- 12113465 TI - Pregnancy, low back pain, and manual physical therapy interventions. PMID- 12113466 TI - The use of a modified classification system in the treatment of low back pain during pregnancy: a case report. AB - STUDY DESIGN: Case study. OBJECTIVE: To describe the use of a classification approach in the evaluation and treatment of a pregnant patient with low back pain (LBP). BACKGROUND: The patient was a 28-year-old primigravida in her 20th week of pregnancy. She presented with a chief complaint of LBP without precipitating trauma. Her pain limited her sitting to 20 minutes or less and restricted her ability to bend forward. METHODS AND MEASURES: This patient was treated 4 times during a period of 2 weeks. The patient was classified as stage 1 extension syndrome. Because of the patient's pregnancy, treatment with active extension exercises commonly prescribed for this syndrome was deemed inadequate. Therefore, manual joint mobilization was applied to the symptomatic vertebral segment. Additional intervention included moist heat, soft tissue mobilization to the thoracolumbar paraspinals, manual stretching of the hip flexors, abdominal bracing, and wall squat exercises. RESULTS: After 4 treatments, the patient was able to bend forward without pain, sit longer than 1 hour without discomfort, and work with minimal discomfort. She improved from a stage 1 classification to a stage 3 classification. CONCLUSION: This case illustrates the use of a classification system to guide physical therapy intervention. It also demonstrates an effective and safe use of manual techniques in the treatment of a pregnant patient. PMID- 12113467 TI - Thoracolumbar proprioception in individuals with and without low back pain: intratester reliability, clinical applicability, and validity. AB - STUDY DESIGN: Repeated measures design of active spinal position sense in individuals with and without low back pain (LBP). OBJECTIVES: Reproducibility and validity evaluation of thoracolumbar proprioception measurement. BACKGROUND: Proprioception studies in peripheral joints and the spine suggest that there may be proprioception deficits due to injury, pain, or degeneration. Kinesthetic retraining may be useful in rehabilitation of patients with LBP, but appropriate measures are required to objectively quantify spinal proprioception. METHODS AND MEASURES: Active-target reproduction in the sagittal, horizontal, and coronal planes was assessed (3 separate occasions for 18 asymptomatic volunteers and 2 occasions for 62 patients with LBP). Repositioning accuracy was expressed as absolute errors (AE) and variable errors (VE). Reliability was analyzed with intraclass correlation coefficient (ICC) and precision with standard error of measurement (SEM) and calculation of the smallest detectable difference (SDD) index. Repeated measures ANOVA and correlations were used for within-group comparisons and discriminant analysis for between-group comparisons. RESULTS: Reproducibility was better for the asymptomatic group, with AE for flexion and rotation being the most reliable (ICC = 0.76-0.80, SEM = 0.91 degrees-1.34 degrees). SDDs were high for all tests, suggesting limited clinical applicability. Reproducibility for the same tests was poor-moderate (ICC = 0.31 0.64, SEM = 0.45 degrees-3.90 degrees) for the patient group. AE for right-side rotation could discriminate between subject groups with 83.3% specificity but only 54.8% sensitivity. CONCLUSIONS: Proprioception testing, with the methods employed, did not demonstrate good measurement properties in a sample of patients with recurrent LBP. Neither could it sufficiently discriminate between individuals with and without LBP. Possible reasons for these findings are discussed. PMID- 12113468 TI - Reliability by surgical status of self-reported outcomes in patients who have shoulder pathologies. AB - STUDY DESIGN: A test-retest design was used to evaluate the reliability of the self-report sections of 4 shoulder pain and disability scales. OBJECTIVE: The objective of the study was to compare interitem consistency and test-retest reliability by surgical status (postoperative versus nonoperative) and to evaluate the effect of surgical status in the prediction of retest scores. BACKGROUND: Patients and healthcare providers evaluate shoulder status based on self-evaluations of pain and disability. Shoulder outcome measures have been developed that include self-reports, but the properties of these measures have not been assessed by surgical status. METHODS AND MEASURES: A questionnaire containing self-report sections of 4 shoulder scales was administered to study participants twice with 1 week between administrations. The outcome measures examined were the: (1) University of California at Los Angeles (UCLA) Shoulder Score; (2) Constant-Murley Scale (CMS); (3) American Shoulder and Elbow Society (ASES) Shoulder Index; and (4) Shoulder Pain and Disability Index (SPADI). Intraclass correlation coefficients (ICC) were calculated to estimate the test retest reliability of each of the scales and subscales. The interitem consistencies of the multi-item subscales were assessed using Cronbach's alpha. The effect of surgical status on shoulder outcome scale reliability was evaluated using a general linear models approach. RESULTS: The interitem consistency estimates for the multi-item scales were high with both operative and nonoperative participants (0.88 to 0.96). With the exception of the satisfaction subscale of the UCLA Shoulder Score for the nonsurgical group, the estimated intraclass coefficients ranged from 0.51 to 0.91. The prediction of UCLA satisfaction and ASES-disability, pain, and total retest scores was improved with the addition of surgical status into a regression model. CONCLUSIONS: The examined scales exhibited good internal consistency across surgical status. The postsurgical sample's reproducibility estimates tended to be higher than those of the nonsurgical sample. Reliability of shoulder outcome scales can be affected by patient surgical status. PMID- 12113469 TI - Assessment of isokinetic muscle strength in women who are obese. AB - STUDY DESIGN: Cross-sectional study of isokinetic trunk and knee muscle strength in women who are obese. OBJECTIVE: To provide reference values, to identify variables that affect peripheral muscle strength, and to provide recommendations for isokinetic testing of trunk and knee muscles in women who are obese and morbidly obese. BACKGROUND: The assessment of peripheral muscle strength is useful for the quantification of possible loss of strength, for exercise prescription, and for the evaluation of the effect of training programs in obese individuals. METHODS AND MEASURES: Isokinetic trunk and leg muscle strength was assessed in 241 women who were obese (18-65 years, body mass index (BMI) > or = 30 kg/m2). Trunk flexion and extension peak torque (PT) was measured using the Cybex TEF dynamometer; trunk rotation (TR) PT was measured using the Cybex TORSO dynamometer; and knee flexion/extension (KFE) PT was measured using the Cybex 350 dynamometer. Body composition was assessed using the bioelectrical impedance method; physical activity was assessed using the Baecke questionnaire; and peak VO2 was assessed using an incremental exercise capacity test on a bicycle ergometer. To identify variables related to muscle strength, Pearson correlations were computed and a stepwise multiple regression analysis was performed. RESULTS: Pearson correlation coefficients of all strength measurements at 60 degrees/s revealed low-to-moderate negative associations with age and positive associations with mass, height, fat free mass (FFM), and peak VO2 (P < 0.05), except for gravity-uncorrected trunk extension strength, which was not related to mass. The sports index of the Baecke questionnaire was associated with TR PT (r = 0.20, P < 0.01) and KFE PT (r = 0.18, P < 0.05). CONCLUSION: The weight of the trunk accounts largely for the measured trunk extensor and flexor strength in women who are obese. Contributing variables of isokinetic trunk flexion and extension strength in women who are obese are age, height, and FFM; whereas sports activities and aerobic fitness are contributing factors for trunk rotational and knee extension strength. Recommendations for measuring isokinetic muscle strength in individuals who are obese are provided. PMID- 12113470 TI - Bioflavonoids: proanthocyanidins and quercetin and their potential roles in treating musculoskeletal conditions. AB - As a clinician treating musculoskeletal conditions, one is continually in search of safe and more effective treatment methods that will hasten tissue healing. Chronic inflammation has been shown to cause connective tissue degradation. Typically, nonsteroidal anti-inflammatory drugs (NSAIDs) and/or corticosteroids are used to control the inflammatory process, however, long-term use has been associated with potentially serious side effects. The purpose of this article is to introduce and describe literature on 2 natural compounds, namely, proanthocyanidin (PCO) and quercetin, which are 2 specific types of bioflavonoids, and to discuss their potential benefits in treating musculoskeletal conditions. There is evidence to suggest that flavonoids may be beneficial to connective tissue for several reasons, which include the limiting of inflammation and associated tissue degradation, the improvement of local circulation, as well as the promoting of a strong collagen matrix. An overview of bioflavonoids as well as relevant research, safety issues, absorption, and specific sources of PCO and quercetin in foods and through supplementation is included. PMID- 12113471 TI - Differential diagnosis and treatment of subcalcaneal heel pain: a case report. PMID- 12113472 TI - Activated leukocyte cell adhesion molecule (CD166/ALCAM): developmental and mechanistic aspects of cell clustering and cell migration. AB - Activated leukocyte cell adhesion molecule (ALCAM/CD166) is a member of the immunoglobulin superfamily and belongs to a recent subgroup with five extracellular immunoglobulin-like domains (VVC2C2C2). ALCAM mediates both heterophilic (ALCAM-CD6) and homophilic (ALCAM-ALCAM) cell-cell interactions. While expressed in a wide variety of tissues, ALCAM is usually restricted to subsets of cells involved in dynamic growth and/or migration, including neural development, branching organ development, hematopoiesis, immune response and tumor progression. Recent structure-function analyses of ALCAM hint at how its cytoskeletal anchoring and the integrity of the extracellular immunoglobulin-like domains may regulate complex cellular properties in regard to cell adhesion, growth and migration. Accumulating evidence suggests that ALCAM expression may reflect the onset of a cellular program for homeostatic control of growth saturation, which induces either growth arrest or cell migration when the upper limits are exceeded. PMID- 12113473 TI - Human Kruppel-like factor5/KLF5: synergy with NF-kappaB/Rel factors and expression in human skin and hair follicles. AB - In this report we describe the identification of Kruppel-like factor 5 (KLF5/BTEB2) in a yeast one-hybrid screen using a keratinocyte-specific, NF kappaB binding site as bait. The KLF5 cDNA encodes a larger protein of 457 aa rather than the earlier reported protein of 209 aa. The full-length KLF5 functions as a transactivator in HepG2 cells, and the stimulation of cells with 12-0-tetradecanoylphorbol-13-acetate (TPA) can modulate its transcriptional activity. Overexpression of KLF5 leads to an increase in the TPA response from VLTRE, a TPA-inducible enhancer element that shows keratinocyte specificity with respect to Rel/NF-kappaB binding. The KLF5-mediated transcriptional increase is not observed in the presence of overexpressed NF-kappaB inhibitor, IkappaBalpha. Cotransfection of KLF5 and the p65 subunit of NF-kappaB, results in a synergistic transactivation of the VLTRE-luciferase reporter. The KLF5 mRNA and the protein is expressed in keratinocytes and throughout the adult human epidermis. Its expression is especially strong in the matrix and the inner root sheath cuticle layer of the hair follicle, sebaceous glands and sweat glands. Considering the TPA-responsiveness and expression pattern, we propose that KLF5 like another member of its family KLF4/GKLF may play an important role in skin morphogenesis and carcinogenesis potentially via its interaction with NF-kappaB factors. PMID- 12113474 TI - Mammalian meiotic telomeres: composition and ultrastructure in telomerase deficient mice. AB - During early meiotic prophase chromosome ends become attached to the nuclear envelope, a process that is essential for faithful homologue pairing and segregation. The factors involved in this attachment are largely unknown. Here we investigated the possible involvement of telomere chromatin by using late generation (G5 and G6) Terc-/- mice. These mice lack telomerase activity and show progressive telomere shortening with increasing mouse generations. We show here that in meiotic chromosome ends of late generation Terc-/- mice telomeric TTAGGG repeats and the TRF1 telomere-binding protein are significantly reduced or below detection level. In spite of this, electron microscopy showed no apparent structural differences at the attachment sites of meiotic chromosomes to the nuclear envelope between wild-type and G6 Terc-/- meiocytes. These results suggest, as already shown in yeast, that most telomere chromatin is dispensable for proper attachment of mammalian meiotic chromosome ends to the nuclear envelope. PMID- 12113475 TI - Chromatin condensation, cysteine-rich protamine, and establishment of disulphide interprotamine bonds during spermiogenesis of Eledone cirrhosa (Cephalopoda). AB - During spermiogenesis in Eledone cirrhosa a single protamine substitutes for histones in nuclei of developing spermatids. This protein displays a peculiar primary structure. It contains 22.6 mol% cysteine residues (19 cysteines in 84 residues). This makes it the most cysteine-rich protamine known. The proportion of basic residues is relatively low (arginine 36.9 mol%, lysine 19.0 mol%). The protamine of E. cirrhosa condenses spermiogenic chromatin in a pattern which comprises fibres with a progressively larger diameter and lamellae that finally undergo definitive coalescence. We have also performed a study that estimates the number of interprotamine disulphide bonds formed during the process of spermiogenic chromatin condensation by means of sequential disappearance of MMNA (monomaleimido-nanogold) labelling. During the first step of spermiogenesis, protamines are found spread over very slightly condensed chromatin with their cysteines in a reactive state (protamine-cys-SH). From this stage the interprotamine disulphide bonds are established in a progressive way. First they are formed inside the chromatin fibres. Subsequently, they participate in the mechanism of fibre coalescence and finally, in the last step of spermiogenesis, the remaining free reactive -SH groups of cysteine form disulphide bonds, thus promoting a definitive stabilization of the nucleoprotein complex in the ripe sperm nucleus. PMID- 12113476 TI - Microtubule dynamics differentially regulates Rho and Rac activity and triggers Rho-independent stress fiber formation in macrophage polykaryons. AB - Multinucleated giant cells (MNGC) derived from avian peripheral blood monocytes present a dense microtubular network emanating from peripherally located centrosomes. We were interested to study how microtubule and F-actin cytoskeletons cooperate in MNGC to maintain cell shape. Microtubule depolymerization by nocodazole triggered the reorganization of the F-actin cytoskeleton in MNGC that is normally organized into podosomes, cortical actin filaments and membrane ruffles. After nocodazole treatment, F-actin was redistributed into unusual transverse fibers associated with focal adhesion plaques. When microtubules were allowed to repolymerize after nocodazole removal, F-actin appeared transiently, together with the small GTPase Rac, in large membrane ruffles. Using affinity precipitation assays, we show that microtubule depolymerization leads to activation of Rho and inhibition of Rac, whereas microtubule repolymerization induces Rac activation and Rho inhibition. Thus, the level of microtubule polymerization inversely regulates Rho and Rac activity in MNGC. Moreover, using C3 exoenzyme, a known inhibitor of Rho, we demonstrate that both the F-actin fiber formation in response to microtubule depolymerization and the formation of membrane ruffles after microtubule repolymerization occur in C3 treated MNGC, indicating that Rho is not required for these events. PMID- 12113477 TI - Subtype-specific neuronal differentiation of PC12 cells transfected with alpha2 adrenergic receptors. AB - Cells of the PC12 rat pheochromocytoma cell line acquire characteristics of sympathetic neurons under appropriate treatment. Stably transfected PC12 cells expressing individual alpha2-adrenergic receptor (alpha2-AR) subtypes were used to assess the role of alpha2-ARs in neuronal differentiation and to characterise the signalling pathways activated by the alpha2-AR agonist epinephrine in these cells. The effects of alpha2-AR activation were compared with the differentiating action and the signalling mechanisms of nerve growth factor (NGF). Epinephrine induced neuronal differentiation of PC12alpha2 cells through alpha2-AR activation in a subtype-dependent manner, internalization of all human alpha2-AR subtypes, and activation of mitogen-activated protein kinase (MAPK) and the serine threonine protein kinase Akt. Epinephrine and NGF showed synergism in their differentiating effects. The MAPK kinase (MEK-1) inhibitor PD 98059 abolished the differentiating effect of epinephrine indicating that the differentiation is dependent on MAPK activation. Activating protein-1 (AP-1) DNA-binding activity was increased after epinephrine treatment in all three PC12alpha2 subtype clones. Evaluation of the potential physiological consequences of these findings requires further studies on endogenously expressed alpha2-ARs in neuronal cells. PMID- 12113478 TI - Comparison between human and animal isolates of Shiga toxin-producing Escherichia coli O157 from Australia. AB - There is very little human disease associated with enterohaemorrhagic Escherichia coli O157 in Australia even though these organisms are present in the animal population. A group of Australian isolates of E. coli O157:H7 and O157:H- from human and animal sources were tested for the presence of virulence markers and compared by XbaI DNA macrorestriction analysis using pulsed-field gel electrophoresis (PFGE). Each of 102 isolates tested contained the gene eae which encodes the E. coli attaching and effacing factor and all but one carried the enterohaemolysin gene, ehxA, found on the EHEC plasmid. The most common Shiga toxin gene carried was stx2c, either alone (16%) or in combination with stx1 (74%) or stx2 (3%). PFGE grouped the isolates based on H serotype and some clusters were source specific. Australian E. coli O157:H7 and H- isolates from human, animal and meat sources carry all the virulence markers associated with EHEC disease in humans therefore other factors must be responsible for the low rates of human infection in Australia. PMID- 12113479 TI - An outbreak of gastroenteritis in Osaka, Japan due to Escherichia coli serogroup O166:H15 that had a coding gene for enteroaggregative E. coli heat-stable enterotoxin 1 (EAST1). AB - In an outbreak of gastroenteritis on 23 July 1996, in Osaka, Japan, 54 of 91 persons who had attended a meeting the previous day became ill. Escherichia coli O166:H15 was isolated from stool specimens of patients (29/33, 88%). Laboratory tests for other bacterial pathogens and viruses were negative. The E. coli 0166 organisms did not adhere to HEp-2 cells in a localized, diffuse, or enteroaggregative manner. The organisms did not express known enterotoxigenic E. coli (ETEC) colonization factors. In polymerase chain reaction tests, the bacteria did not have coding genes for shigatoxin of enterohemorrhagic E. coli (EHEC), heat-labile, or heat-stable enterotoxin of ETEC, attachment and effacement (eaeA) of EPEC, or invasion (invE) of enteroinvasive E. coli (EIEC). Consequently, they could not be assigned to any of the recognized diarrhoeagenic groups of E. coli: EPEC, ETEC, EHEC, EIEC, enteroaggregative E. coli (EAggEC), or diffusely adhering E. coli. However, the organisms possessed the EAggEC heat stable enterotoxin (EAST1) gene. To our knowledge, this is the first report of an outbreak caused by E. coli that did not have well-characterized virulence genes other than EAST1. The isolates showed the same DNA banding pattern in pulsed field gel electrophoresis after digestion with the restriction enzymes XbaI or NotI. Three O166:H15 strains isolated from two sporadic cases and another outbreak during 1997-8 were distinct, indicating that multiple clones have spread already. We propose that diarrhoeal specimens should be examined for E. coli possessing the EAST1 gene. PMID- 12113480 TI - Molecular epidemiology of Salmonella typhimurium isolates from human sporadic and outbreak cases. AB - The molecular epidemiology of a representative collection of sporadic foreign and domestically acquired Salmonella Typhimurium (S. Typhimurium) isolates from Norwegian patients in 1996-9 was studied by numerical analysis of pulsed-field gel electrophoresis (PFGE) profiles. Three subclusters (E5, F1 and G1) comprised 47% of the 102 sporadic isolates investigated and 45% of the domestically acquired isolates fell in subclusters E5 and F1. Distinct seasonal and geographic variations were evident for these strains which have been responsible for both local outbreaks (E5) and a national epidemic (F1) where salmonella-infected hedgehogs and birds constituted the suggested primary source of infection. Subcluster G1 was dominated by imported multi-resistant definitive type (DT) 104 isolates. All multi-resistant isolates contained integron-associated gene cassette-structures. This study presents valuable information on the relative significance, geographic distribution and antibiotic resistance features of distinct S. Typhimurium clones causing human salmonellosis among Norwegians. PMID- 12113481 TI - The seasonal distribution of campylobacter infection in nine European countries and New Zealand. AB - In all temperate countries campylobacter infection in humans follows a striking seasonal pattern, but little attention has been given to exploring the epidemiological explanations. In order to better characterize the seasonal patterns, data from nine European countries and New Zealand have been examined. Several European countries with weekly data available showed remarkably consistent seasonal patterns from year to year, with peaks in week 22 in Wales, week 26 in Scotland, week 32 in Denmark, week 30 in Finland and week 33 in Sweden. In Europe, the seasonal peak was most prominent in Finland and least prominent in Scotland and Austria. In New Zealand the seasonality was less consistent since the peak was more prolonged. Possible explanations for the seasonal peaks are discussed. Research into the causes of campylobacter seasonality should help considerably in elucidating the sources of human infection. PMID- 12113482 TI - Streptococcus pyogenes prtFII, but not sfbI, sfbII or fbp54, is represented more frequently among invasive-disease isolates of tropical Australia. AB - Streptococcus pyogenes (group A streptococcus) strains may express several distinct fibronectin-binding proteins (FBPs) which are considered as major streptococcal adhesins. Of the FBPs, SfbI was shown in vitro to promote internalization of the bacterium into host cells and has been implicated in persistence. In the tropical Northern Territory, where group 4 streptococcal infection is common, multiple genotypes of the organism were found among isolates from invasive disease cases and no dominant strains were observed. To determine whether any FBPs is associated with invasive disease propensity of S. pyogenes, we have screened streptococcal isolates from bacteraemic and necrotizing fasciitis patients and isolates from uncomplicated infections for genetic endowment of 4 FBPs. No difference was observed in the distribution of sfbII, fbp54 and sfbI between the blood isolates and isolates from uncomplicated infection. We conclude that the presence of sfbI does not appear to promote invasive diseases, despite its association with persistence. We also show a higher proportion of group A streptococcus strains isolated from invasive disease cases possess prtFII when compared to strains isolated from non-invasive disease cases. We suggest that S. pyogenes may recruit different FBPs for different purposes. PMID- 12113484 TI - Listeria monocytogenes meningitis: serotype distribution and patient characteristics in The Netherlands, 1976-95. AB - Two hundred and seven cases of listeria meningitis that occurred in The Netherlands over 20 years were reviewed to study associations between Listeria monocytogenes serotype, age, underlying disease, and outcome. The mean annual incidence per 100,000 population was 0.12 in 1981-90, decreasing to 0.07 in 1991 5. Underlying disease was present in 50% of non-neonatal patients, most often haematological malignancy (15%) and the use of immunosuppressive therapy (14%). The meningitis-related case fatality rate was 16%; a significantly higher rate was associated with the presence of underlying disease (30%) or age > or = 70 years (29%). Serotype 4b was most frequent (65%) and L. monocytogenes types 1/2a, 1/2b, or 1/2c (30% of cases) were significantly more often isolated from non neonatal patients with underlying disease, suggesting a higher virulence of listerial serotype 4b. PMID- 12113485 TI - The burden of Helicobacter pylori infection in England and Wales. AB - The prevalence of active infection with Helicobacter pylori in the general population of England and Wales was estimated using high reactivity for specific IgG in serum ELISA as a marker. A total of 10,118 anonymized residues of serum samples collected in 1986 and 1996 from persons aged 1-84 years were used. Estimated prevalence of active infection varied by region and was highest in London. Prevalence was related to decade of birth and increased from 4-3% in those born during the 1980s to 30% in those born before 1940. An estimated total of 7.5 million people living in England and Wales have an active infection and analysis by decade of birth showed no significant difference between samples collected in 1986 and 1996. These data suggest H. pylori infection is becoming less common, is acquired at an early age and is unlikely to be resolved unless suitable antimicrobial treatment is sought. PMID- 12113483 TI - Distribution of emm genotypes and superantigen genes of Streptococcus pyogenes isolated in Japan, 1994-9. AB - The purpose of this study was to examine characteristic profiles of Streptococcus pyogenes clinical isolates isolated in Japan during 1994-9. Genotyping of the M protein (emm typing) revealed that emm types 12 and 28 were the most common among 316 isolates. Most of the emm12 isolates were isolated from mucosa, while emm58 and emm89 were from skin. Moreover, the emm3 isolates were dominant in invasive infections. The distribution of 6 superantigen genes showed that all isolates harboured the mf gene and many had the speG gene. Invasive isolates were shown to have the ssa gene at a higher rate (76%) than noninvasive (37%). The distribution of superantigens was significantly different between emm types, but not between isolation sites. These results suggest that the distribution of emm types is related to isolation site, whereas superantigen distribution is related to clinical features of S. pyogenes infections. PMID- 12113486 TI - Case-control studies of sporadic cryptosporidiosis in Melbourne and Adelaide, Australia. AB - Few studies have assessed risk factors for sporadic cryptosporidiosis in industrialized countries, even though it may be numerically more common than outbreaks of disease. We carried out case-control studies assessing risk factors for sporadic disease in Melbourne and Adelaide, which have water supplies from different ends of the raw water spectrum. In addition to examining drinking water, we assessed several other exposures. 201 cases and 795 controls were recruited for Melbourne and 134 cases and 536 controls were recruited for Adelaide. Risk factors were similar for the two cities, with swimming in public pools and contact with a person with diarrhoea being most important. The consumption of plain tap water was not found to be associated with disease. This study emphasizes the need for regular public health messages to the public and swimming pool managers in an attempt to prevent sporadic cryptosporidiosis, as well as outbreaks of disease. PMID- 12113487 TI - A community survey of self-reported gastroenteritis undertaken during an outbreak of cryptosporidiosis strongly associated with drinking water after much press interest. AB - We took the opportunity provided by a large outbreak of cryptosporidiosis in the North West of England to investigate the potential impact of recall bias on strength of association and on estimates of outbreak size. We conducted a community-based survey of 4 towns within the outbreak area and 4 control towns. A postal questionnaire was sent to 120 homes, chosen at random from the local telephone directory, in each of the 8 towns. Although not statistically significant, the prevalence of self-reported diarrhoeal disease was marginally higher in the control towns than in the outbreak towns. There was a very strong association between self-reported diarrhoea and drinking water consumption in both control and outbreak areas. The impact of recall bias in outbreak investigations is much greater than previously thought. Identification of the cause of outbreaks should not be based solely on case-control studies conducted after the press has reported the outbreak. Such evidence is likely to be unreliable and give falsely significant associations between water consumption and disease. PMID- 12113488 TI - Chronic Strongyloides stercoralis infection in Laotian immigrants and refugees 7 20 years after resettlement in Australia. AB - During the period 1974-91 large numbers of Southeast Asian immigrants and refugees were resettled in Western countries, including Australia. Health screening during this period demonstrated that intestinal parasite infections were common. A cross-sectional survey of 95 Laotian settlers who arrived in Australia on average 12 years prior to the study was conducted to determine if chronic intestinal parasite infections were prevalent in this group. Twenty-three participants had positive Strongyloides stercoralis test results (22 with positive serology, including I with S. stercoralis larvae detected in faeces and another with larvae and equivocal serology). Of these 23 participants, 18 (78%) had an elevated eosinophil count. Two patients had eggs of Opisthorchis spp. identified by faecal microscopy. The detection of chronic strongyloidiasis in Laotian settlers is a concern because of the potential serious morbidity associated with this pathogen. PMID- 12113489 TI - Influenza and pneumococcal vaccine distribution and use in primary care and hospital settings in Scotland: coverage, practice and policies. AB - A survey of the coverage, distribution and the factors associated with use of influenza and pneumococcal vaccines among general practitioners (GPs) in primary care and in hospital settings was carried out in 53 general practices in Scotland taking part in the 'Continuous Morbidity Recording' (CMR) programme. The annual vaccine distribution increased substantially among 53 general practices from 1993 to 1999 and in Scotland as a whole from 1984 to 1999. From the questionnaire, overall coverage was 43% (95% CI 38-48) for influenza vaccine in the 2000-1 season and 13% (95% CI 9-16) for pneumococcal vaccine in the last 5 year period, in high-risk patients recommended for these vaccines by the Department of Health (DoH). Influenza vaccine coverage was highest in the elderly (65 years of age and above) at 62% (95% CI 59-74). Although pneumococcal vaccination is not currently recommended for all elderly, coverage of this vaccine was also higher in this group (22%, 95% CI 16-29). In the majority of patients (influenza vaccine, 98% and pneumococcal vaccine, 94%), vaccination was carried out in general practice. Only 2% of patients had received pneumococcal vaccination in a hospital setting. The level of influenza and pneumococcal vaccination varied with the level of deprivation. Most GPs considered that the responsibility for influenza and pneumococcal vaccination lay with them. Forty-five percent of GPs reported having a written policy with set target for influenza vaccination and 11% for pneumococcal vaccination. PMID- 12113490 TI - The changing epidemiological pattern of hepatitis A in England and Wales. AB - Sera from an age-stratified sample of 4188 individuals, submitted for diagnostic purposes to 15 public health laboratories in England and Wales in 1996, were tested for hepatitis A antibody. The serological profiles were consistent with declining incidence in the past. This hypothesis was tested by comparing the serological profiles of Ashford, Leeds and Preston public health laboratories with those from sera collected during a previous study in the same laboratories in 1986/7. A comparison of equivalent 10 year birth cohorts revealed that significant hepatitis A seroconversion had only continued in Ashford. However, it is probable that most seroconversions are due to vaccination and immigration rather than continuing viral transmission. Further population-based surveys collecting more in-depth social and demographic data are needed to confirm the main factors influencing hepatitis A seroprevalence and to explain the regional differences. PMID- 12113491 TI - Epidemiology of hepatitis B infection among the Nicobarese--a mongoloid tribe of the Andaman and Nicobar Islands, India. AB - Andaman and Nicobar Islands, Union Territory of India, is home to six primitive tribes. Preliminary serological studies carried out earlier among the four accessible tribes revealed that hepatitis B virus (HBV) infection is hyper endemic among them. The present study was carried out to understand important modes of transmission and to identify possible risk factors associated with HBV infection among the Nicobarese tribe. The epidemiology of HBV infection in these islands appears to be distinct with a high prevalence of the chronic carrier state (22.2%) associated with a comparable proportion of the population being anti-HBs positive (26.3%). More than half of the HBsAg and anti-HBs negative individuals have anti-HBc antibodies. Age, past history of hospital admission, intramuscular injections and number of carriers in the tuhet were found to be significantly associated with exposure to hepatitis B virus. Horizontal transmission through close contact with carriers and perinatal route appears to be an important mode of transmission of HBV in this community. Besides this, use of unsafe injections represents an independent risk factor for acquiring HBV infection in this population. Introducing HBV vaccine in the infant immunization programme and improving injection safety would help to control the infection in the tribal community of these islands. PMID- 12113492 TI - Prevalence of hepatitis C among injectors in Scotland 1989-2000: declining trends among young injectors halt in the late 1990s. AB - We previously reported a continual decline in anti-HCV prevalence among young injectors from Glasgow and Lothian between 1990 and 1997. The original study was extended to ascertain if the anti-HCV prevalence among injectors from Glasgow, Lothian, Tayside and Grampian had changed since 1997. Residual sera from injectors who had undergone attributable anti-HIV testing were tested anonymously for anti-HCV. In all four regions, no significant changes in prevalence were found among those aged < 25 years during the late 1990s (Glasgow 1997-9/00: 43% 41%; Lothian 1997-9: 13%-17%; Tayside 1997-9: 45%-35%; Grampian 1996-9: 28%-29%). Among those aged > or = 25 years, significant decreases in prevalence were only observed in Glasgow (1997-9/00: 79%-72%, P = 0.03) and Lothian (1997-9: 54%-45%, P = 0.05). The findings highlight that existing harm reduction measures, acknowledged as having helped to reduce the spread of HCV, are not sufficient to bring this epidemic under control and reduce transmission to sporadic levels. PMID- 12113494 TI - Human papillomaviruses in Amerindian women from Brazilian Amazonia. AB - We evaluated the prevalence of human papillomavirus (HPV) infection in Amerindian women from a tribe in Brazilian Amazonia. Demographic data, pap smears and cervical samples for HPV DNA detection by polymerase chain reaction (PCR) were obtained for women aged above 10 years old. In total, 79 (85.9%) out of 92 eligible women who lived there were interviewed; all women already had engaged in sexual activity. Seventy-eight and 49 women allowed collection of pap smears and PCR samples, respectively. Cytological signs of HPV infection were observed in 11 patients; 6 of these were probed for HPV infection and 1 shown to be HPV 16. Overall prevalence of HPV infection detected by PCR was 14.3%. Three patients presented high-risk HPV DNA types:two HPV 16 and one co-infection of HPV 16 and 58. Cervical infection by oncogenic HPV types occurs in Amerindian women and cervical cancer screening should be a priority in this setting. PMID- 12113493 TI - Assessment of varicella underreporting in Italy. AB - We conducted a study to assess the degree of varicella underreporting in Italy, and its distribution by age group and geographical area. Underreporting in individuals from 6 months to 20 years of age was computed as the ratio between the varicella seroprevalence in 1996 and the 1996 lifetime cumulative incidence based on statutory notifications. The degree of underreporting at the national level was 7.7 (95% CI 7.4-7.9); underreporting was greater in older age groups and in southern Italy. Quantification of underreporting can contribute to better understanding of the burden of varicella and to evaluating the potential impact of mass vaccination. PMID- 12113495 TI - Modelling equine influenza 1: a stochastic model of within-yard epidemics. AB - This paper demonstrates that a simple stochastic model can capture the features of an epidemic of equine influenza in unvaccinated horses. When the model is modified to consider vaccinated horses, we find that vaccination dramatically reduces the incidence and size of epidemics. Although occasional larger outbreaks can still occur, these are exceptional. We then look at the effects of vaccination on a yard of horses, and in particular at the relationship between pre-challenge antibody level and quantity of virus shed when challenged with the virus. While on average, a high antibody level implies that less virus will be shed during the infectious period, we identify a high degree of heterogeneity in the response of horses with similar pre-challenge antibody levels. We develop a modified model that incorporates some heterogeneity in levels of infectivity, and compare this with the simpler model. PMID- 12113496 TI - Genealogical analyses of rabies virus strains from Brazil based on N gene alleles. AB - Thirty rabies virus isolates from cows and vampire bats from different regions of Sao Paulo State, Southeastern Brazil and three rabies vaccines were studied genetically. The analysis was based on direct sequencing of PCR-amplified products of 600 nucleotides coding for the amino terminus of nucleoprotein gene. The sequences were checked to verify their genealogical and evolutionary relationships and possible implication for health programmes. Statistical data indicated that there were no significant genetic differences between samples isolated from distinct hosts, from different geographical regions and between samples collected in the last two decades. According to the HKA test, the variability observed in the sequences is probably due to genetic drift. Since changes in genetic material may produce modifications in the protein responsible for immunogenicity of virus, which may eventually cause vaccine failure in herds, we suggest that continuous efforts in monitoring genetic diversity in rabies virus field strains, in relation to vaccine strains, must be conducted. PMID- 12113497 TI - Comparative epidemiology of scrapie outbreaks in individual sheep flocks. AB - Data recording the course of scrapie outbreaks in 4 sheep flocks (2 in Cheviot sheep and 2 in Suffolks) are compared. For each outbreak the data on scrapie incidence and sheep demography and pedigrees cover periods of years or decades. A key finding is that the incidence of clinical cases peaks in sheep 2-3 years old, despite very different forces-of-infection. This is consistent with age-specific susceptibility of sheep to scrapie, as has been reported for cattle to bovine spongiform encephalopathy and for humans to variant Creutzfeldt-Jakob disease. Scrapie incidence was higher in ewes than rams and at certain times of years, though these effects were not consistent between flocks. There was no evidence for high levels of vertical transmission. PMID- 12113498 TI - Prevalence of Salmonella typhimurium infection in Norwegian hedgehog populations associated with two human disease outbreaks. AB - Faecal carriage of salmonella was investigated in 320 hedgehogs from Moss municipality in south-eastern Norway, Askoy, Bergen and Os municipalities in central-western Norway, and five municipalities in south-western and central Norway. The sampling in Moss was carried out 1 year after a human outbreak of salmonellosis, whereas the sampling in Askoy, Bergen and Os was carried out during a human outbreak. Both outbreaks were caused by Salmonella Typhimurium 4,5,12:i:1,2. No salmonella were detected in the hedgehogs from south-western (0/115) and central (0/24) Norway. Thirty-nine percent (39/99) of the animals sampled on Jeloy, and 41% (34/82) of those from Askoy, Bergen and Os, carried S. Typhimurium 4,5,12:i:1,2. The PFGE profile of isolates from hedgehogs and human beings were identical within each of the two outbreak areas. A significantly higher carrier rate of S. Typhimurium occurred among hedgehogs sampled at feeding places, compared to those caught elsewhere. The salmonella-infected hedgehog populations most likely constituted the primary source of infection during both of the human disease outbreaks, and the Norwegian hedgehog is suggested as a reservoir host of S. Typhimurium 4,5,12:i:1,2. PMID- 12113499 TI - The impact of sporadic campylobacter and salmonella infection on health and health related behaviour: a case control study. AB - The aim of the work was to explore the impact on general and psychological health of those with a proven bacterial gastrointestinal infection and to compare this with controls from whom no bacterial pathogen was identified. A case control study was conducted using an interviewer-administered questionnaire. Thirty-nine cases from whose faeces salmonella or campylobacter had been cultured were compared with matched controls. Reported gastrointestinal symptoms, general health and self-reported hygiene practices were compared. At the time of acute illness the General Household Questionnaire suggested similar levels of morbidity, though by follow up the controls were substantially more likely to be distressed. Cases were more likely to have changed their food preparation practices, to avoid certain eating places and to have been given advice about food preparation. In this small study a positive diagnosis of salmonella or campylobacter seems to have had a reassuring effect when compared with those for whom no diagnosis was made. PMID- 12113500 TI - Prevalence of the genes for shigella enterotoxins 1 and 2 among clinical isolates of shigella in Israel. AB - Two enterotoxins, shigella enterotoxin 1 (SHET1) and shigella enterotoxin 2 (SHET2) have been recently characterized and are believed to play a role in the clinical manifestation of shigellosis. One hundred and twenty-one isolates of Shigella spp. of 13 different serotypes and variants and 10 isolates of enteroinvasive Escherichia coli (EIEC) isolated in Israel, were examined by polymerase chain reaction for the presence of SHET1 and SHET2 genes. SHET1 was only prevalent among isolates of S. flexneri 2a while SHET2 was found in all the serotypes that were tested except for several isolates of S. flexneri 1b that lost their virulence plasmid during storage. In addition, we found that the S. flexneri 2a vaccine strain T-32 Istrati contains the gene encoding for SHET1 but not that encoding for SHET2, suggesting that the latter is located within a large deletion occurring in the 140 Mda plasmid of this S. flexneri 2a non-invasive vaccine strain. PMID- 12113501 TI - Fixation of Cs to marine sediments estimated by a stochastic modelling approach. AB - Dumping of nuclear waste in the Kara Sea represents a potential source of radioactive contamination to the Arctic Seas in the future. The mobility of 137Cs ions leached from the waste will depend on the interactions with sediment particles. Whether sediments will act as a continuous permanent sink for released 137Cs, or contaminated sediments will serve as a diffuse source of 137Cs in the future, depends on the interaction kinetics and binding mechanisms involved. The main purpose of this paper is to study the performance of different stochastic models using kinetic information to estimate the time needed for Cs ions to become irreversibly fixed within the sediments. The kinetic information was obtained from 134Cs tracer sorption and desorption (sequential extractions) experiments, conducted over time, using sediments from the Stepovogo Fjord waste dumping site, on the east coast of Novaya Zemlya. Results show that 134Cs ions interact rapidly with the surfaces of the Stepovogo sediment, with an estimated distribution coefficient Kd(eq) of 300 ml/g (or 13m2/g), and the 134Cs ions are increasingly irreversibly fixed to the sediment over time. For the first time, stochastic theory has been utilised for sediment-seawater systems to estimate the mean residence times (MRTs) of Cs ions in operationally defined sediment phases described by compartment models. In the present work, two different stochastic models (i) a Markov process model (MP) being analogous to deterministic compartment models, and (ii) a semi-Markov process model (SMP) which should be physically more relevant for inhomogeneous systems, have been compared. As similar results were obtained using the two models, the less complicated MP model was utilised to predict the time needed for an average Cs ion to become irreversibly fixed in the Stepovogo sediments. According the model, approximately 1100 days of contact time between Cs ions and sediments is needed before 50% of the 134Cs ion becomes fixed in the irreversible sediment phase. while about 12.5 years are needed before 99.7% of the Cs ions are fixed. Thus, according to the model estimates the contact time between 137Cs ions leached from dumped waste and the Stepovogo Fjord sediment should be about 3 years before the sediment will act as an efficient permanent sink. Until then a significant fraction of 137Cs should be considered mobile. The stochastic modelling approach provides useful tools when assessing sediment-seawater interactions over time, and should be easily applicable to all sediment-seawater systems including a sink term. PMID- 12113502 TI - Radioactive contamination in the marine environment adjacent to the outfall of the radioactive waste treatment plant at ATOMFLOT, northern Russia. AB - RTP "ATOMFLOT" is a civilian nuclear icebreaker base located on the Kola Bay of northwest Russia. The objectives of this study were to determine the distributions of man-made radionuclides in the marine environment adjacent to the base, to explain the form of the distributions in sediments and to derive information concerning the fate of radionuclides discharged from ATOMFLOT. Mean activity concentrations (d.w.) for surface sediment, of 63 Bq kg(-1 137Cs, 5.8 Bq kg(-1) 90Sr and 0.45 Bq kg(-1 239,240)Pu were measured. Filtered seawater activity levels were in the range of 3--6.9 Bq m(-3) 137Cs, 2.0-11.2 Bq m(-3) 90Sr, and 16-40 m Bq m(-3), 239,240Pu. Short-lived radionuclides were present at sediment depths in excess of 10cm indicating a high degree of sediment mixing. Correlations of radionuclide activity concentrations with grain-size appear to be absent; instead, the presence of relatively contaminated sediment appears to be related to the existence of radioactive particles. PMID- 12113503 TI - Transfer of 137Cs and 60Co from irrigation water to a soil-tomato plant system. AB - An experiment has been performed at the nuclear power plant of Garigliano (Caserta, Italy), aiming at the measurement of transfer factors of 137Cs and 60Co radionuclides from the irrigation water to a soil-plant system, with particular attention to the influence on such transfers of the irrigation technique (ground or aerial). Tomato plants were irrigated weekly with water contaminated with 137Cs and 60Co (about 375 Bq/m2 week), using both irrigation techniques. After 13 weeks, fruits, leaves, stems, roots and soil were sampled, and radionuclide concentrations were measured by high-resolution gamma spectroscopy. It was found that the activity allocated to the plant organs is significantly dependent upon the irrigation technique, amounting to 2.1% and 1.6% of the activity given in the cultivation for aerial treatment and 0.4% and 0.3% for the ground treatment, for 137Cs and 60Co respectively. The activity absorbed by plants is allocated mainly in leaves (> 55%), while less then 10% is stored in the fruits, for both irrigation techniques. Transfer factors (soil-plant and irrigation water-plant) of tomato plants and of weeds have been determined for 137Cs and 60Co, as well as for natural 40K in the soil. PMID- 12113504 TI - Natural radioactivity in the scale of water well pipes. AB - The natural radioactivity of 226Ra and 228Ra in scale samples taken from pipes used in several local water wells was investigated. The results showed 226Ra activities to be varying from 1284 to 3613 Bq/kg whereas, the 228Ra concentrations did not show any significant variation, all being low, below 30 Bq/kg. The 222Rn exhalations from these scale samples were also measured and compared with the 226Ra contents. The average ratio of 222Rn/226Ra was 31%. Chemical analyses showed that the main constituent of the scale samples was iron. The radiation dose rates from the pipes and scale were up to 100nSv/h. Although not a major hazard this could present a long-term risk if the scale materials were handled indiscriminately. PMID- 12113505 TI - Modelling the long-term dynamics of radiocaesium in closed lakes. AB - During the years after the Chernobyl accident the radioceasium activity concentration in most contaminated aquatic ecosystems decreased markedly. Lakes with no permanent inflows and outflows (closed lakes), however, still present a radioecological problem which is expected to continue for some time. In this paper, a mechanistic model for the long-term prediction of radiocaesium behaviour in closed lakes is developed. The model of Prokhorov (Radiokhimiya (Radiochemistry) 11 (1969) 317) was modified to describe the effects of bottom sediment bioturbation, surface runoff from the catchment and suspended solids formation and sedimentation. The model input parameters are the effective diffusion coefficient in bottom sediments, depth of the completely mixed layer, the distribution coefficient in the sediment-water system, the runoff coefficient, sedimentation rate, and deposition density. Values of all these parameters can be independently estimated or measured in a short-term experiment. Given negligible runoff and sedimentation, the dynamics of radiocaesium in lake water is described by a simple equation with only one unknown parameter. This allows us to make long-term predictions on the basis of a series of measurements carried out during the relatively short period. The model was tested against 137Cs activity concentrations measured between 1993 and 1999 in Svyatoe lake in the Bryansk region of Russia. Calculated and measured activity concentrations are in good agreement. PMID- 12113507 TI - In situ gamma-ray spectrometry in forests: determination of kerma rate in air from 137Cs. AB - A method is presented to determine the kerma rate in air from 137Cs due to Chernobyl fallout in forests. In situ gamma-ray spectra from several forest sites in Russia, in the Ukraine and in Southern Germany are evaluated with the aim of deducing the ratio of primary and forward scattered photons for 137Cs. With this ratio and the results of Monte-Carlo simulations of photon transport the contribution of scattered photons to the total kerma is assessed successfully. Scattered photons contribute between 42% and 50% to the total kerma rate from radiocesium, which is less than according values for grassland areas. The contribution of radiocesium to the total kerma rate varies between 40% and 90%. whereas radiocesium stored in the forest biomass contributes only a few percent. The mean mass depth of radiocesium ranges from 2.6 to 6.4 g cm(-2) in the forest soils. PMID- 12113506 TI - Frequency distributions of 137Cs in fish and mammal populations. AB - We collected fish and mammals in several radioactively contaminated locations in the Chornobyl Exclusion Zone and analyzed them for 137Cs content. Frequency distributions were built for populations of channel catfish, yellow-necked mice and bank voles. We combined our data with similar data from several other studies to demonstrate the relationship between the standard deviations and means of 137Cs of fish and mammal populations. The frequency distributions of 137Cs in populations of fish and mammals are not normal, as indicated by the strong relationship between standard deviation and mean. Distributions for mammals are more skewed than those for fish. Fish and mammals probably use their environments in fundamentally different ways. The highest concentrations and thus greatest risks are therefore confined to relatively few individuals in each population. PMID- 12113508 TI - Current contamination by 137Cs and 90Sr of the inhabited part of the Techa river basin in the Urals. AB - Previous discharges of radioactivity from the Mayak Production Association plant in the Urals have resulted in considerable radionuclide contamination of the Techa River, and consequent high radiation doses during the late 1940s and 1950s to residents of villages along the Techa river. The most contaminated villages close to the site were evacuated in the period 1954-1962. The objective of this recent study was to conduct a preliminary assessment of the current radioactive contamination of soil, vegetation and foodstuffs in the two remaining villages closest to the Mayak site, Muslyumovo and Brodokalmak. The highest contamination levels in soil were found in the floodplain at 5.5 MBq m(-2) for 137Cs and 1.0 MBq m(-2) for 90Sr. Radionuclide contamination in soil of the villages was much lower, but exceeded that expected from global fallout. Data from 1207 measurements of 137Cs in milk and 1180 for 90Sr in milk for the period 1992-1999 were collated. There was no change with time in the 90Sr or 137Cs activity concentration in milk over the measured period. There were significantly higher 137Cs activity concentrations in milk sampled during the housed winter period in Muslyumovo compared with the grazing summer period, but compared to that for Brodokalmak or for either settlement for 90Sr. The highest measured activity concentrations in food products of 137Cs and 90Sr were found in river fish, waterfowl, poultry and milk. The measured activity concentrations of 137Cs and 90Sr in some animal products were higher than that expected from soil and vegetation from fields and pasture in the villages (not including the floodplain) confirming that the highly contaminated floodplains are contributing to contamination of some animal products. PMID- 12113510 TI - The contributions of library and information services to hospitals and academic health sciences centers: a preliminary taxonomy. AB - OBJECTIVES: This article presents a taxonomy of the contributions of library and information services (LIS) in hospitals and academic health sciences centers. The taxonomy emerges from a study with three objectives: to articulate the value of LIS for hospitals and academic health sciences centers in terms of contributions to organizational missions and goals, to identify measures and measurable surrogates associated with each LIS contribution, and to document best practices for communicating the value of LIS to institutional administrators. METHODS: The preliminary taxonomy of LIS contributions in hospitals and academic health sciences centers is based on a review of the literature, twelve semi-structured interviews with LIS directors and institutional administrators, and a focus group of administrators from five academic, teaching, and nonteaching hospitals. RESULTS: Derived from the balanced scorecard approach, the taxonomy of LIS contributions is organized on the basis of five mission-level concepts and fifteen organizational goals. LIS contributions are included only if they have measurable surrogates. CONCLUSIONS: The taxonomy of LIS contributions offers a framework for the collection of both quantitative and qualitative data in support of communicating the value of LIS in hospitals and academic health sciences centers. PMID- 12113509 TI - Determining value. PMID- 12113511 TI - Outreach to community organizations: the next consumer health frontier. AB - Outreach by health sciences librarians to various constituencies helps ensure that the information needs of these groups are adequately met. In the past, librarians have extended outreach to isolated rural health care providers, urban clinicians not affiliated with libraries, and consumers. One logical next step in outreach efforts is to partner with existing community groups who seek to improve the health of residents concerning such areas as lead contamination and rising asthma rates. This article describes a National Network of Libraries of Medicine funded project, where librarians worked with a group of grass roots community organizations, schools, and clinics. Knowledge gained from past outreach projects was applied with mixed success because of the unique nature of community groups where many unanticipated issues arose. Lessons learned while extending outreach library services to community groups are described. PMID- 12113513 TI - Students and overdue books in a medical library. AB - At the University of Ilorin Medical Library, sixty-one randomly selected medical students with overdue books were surveyed using a questionnaire with a view to (1) finding out why they had not returned the library books in their possession, (2) determining their perceptions of eight given overdue measures, and (3) seeking suggestions on how else to reduce overdue books. Most of the overdue books were as a result of (1) the students not finishing with the books and (2) the students being forgetful. Providing for renewals was the most favored overdue measure, while the need for increased multiple copies and extended loan periods for students were also stressed. Thus, a notice urging readers to return or renew borrowed library books was mounted on the issue desk as a reminder to all readers borrowing books. The library is being automated, which will facilitate timely generation and sending of overdue notices. More copies of some titles were purchased, while a copy each of others was transferred to the reserve collection. The need for an extended loan period will require further investigation, while the judicious use of other overdue measures to complement providing for renewals is recommended. PMID- 12113512 TI - Database searches for qualitative research. AB - Interest in the role of qualitative research in evidence-based health care is growing. However, the methods currently used to identify quantitative research do not translate easily to qualitative research. This paper highlights some of the difficulties during searches of electronic databases for qualitative research. These difficulties relate to the descriptive nature of the titles used in some qualitative studies, the variable information provided in abstracts, and the differences in the indexing of these studies across databases. PMID- 12113515 TI - Use of Web-based library resources by medical students in community and ambulatory settings. AB - PURPOSE: The purpose was to evaluate the use of Web-based library resources by third-year medical students. SETTING/PARTICIPANTS/RESOURCES: Third-year medical students (147) in a twelve-week multidisciplinary primary care rotation in community and ambulatory settings. METHODOLOGY: Individual user surveys and log file analysis of Website were used. RESULTS/OUTCOMES: Twenty resource topics were compiled into a Website to provide students with access to electronic library resources from any community-based clerkship location. These resource topics, covering subjects such as hypertension and back pain, linked to curriculum training problems, full-text journal articles, MEDLINE searches, electronic book chapters, and relevant Websites. More than half of the students (69%) accessed the Website on a daily or weekly basis. Over 80% thought the Website was a valuable addition to their clerkship. DISCUSSION/CONCLUSION: Web-based information resources can provide curriculum support to students for whom access to the library is difficult and time consuming. PMID- 12113514 TI - When less is more: a practical approach to searching for evidence-based answers. AB - The information needs of practicing clinicians are distinct from the needs of students, researchers, or nonclinical personnel. Clinicians seek information to stay current with new relevant medical developments and to find answers to patient-specific questions. The volume of available information makes clinicians' tasks of rapidly identifying high-quality studies daunting. New tools evaluate the rigor and relevance of information and summarize it in the form of synthesized clinical answers. These sources have the opposite focus of many other information tools in that they strive to provide less information rather than more. With the development of these sources of validated and refined information, a new search approach is needed to locate clinical information in which speed is the benchmark. The existing medical literature, including these new refinement tools, can be conceptualized as a pyramid, with the most useful information, based on validity and relevance, placed at the apex. Use of this hierarchy allows searchers to drill down through progressive layers until they find their answers. Librarians can play a significant role in evaluating the ever-increasing variety of these synthesized resources, placing them into the searching hierarchy, and training clinicians to search from the top down. PMID- 12113516 TI - Collection analysis techniques used to evaluate a graduate-level toxicology collection. AB - Collections librarians from academic libraries are often asked, on short notice, to evaluate whether their collections are able to support changes in their institutions' curricula, such as new programs or courses or revisions to existing programs or courses. With insufficient time to perform an exhaustive critique of the collection and a need to prepare a report for faculty external to the library, a selection of reliable but brief qualitative and quantitative tests is needed. In this study, materials-centered and use-centered methods were chosen to evaluate the toxicology collection of the University of Saskatchewan (U of S) Library. Strengths and weaknesses of the techniques are reviewed, along with examples of their use in evaluating the toxicology collection. The monograph portion of the collection was evaluated using list checking, citation analysis, and classified profile methods. Cost-effectiveness and impact factor data were compiled to rank journals from the collection. Use-centered methods such as circulation and interlibrary loan data identified highly used items that should be added to the collection. Finally, although the data were insufficient to evaluate the toxicology electronic journals at the U of S, a brief discussion of three initiatives that aim to assist librarians as they evaluate the use of networked electronic resources in their collections is presented. PMID- 12113517 TI - MedReach: building an Area Health Education Center medical information outreach system for northwest Ohio. AB - In collaboration with regional partners in northwest Ohio, the Area Health Education Center (AHEC) program at the Medical College of Ohio (MCO) at Toledo is reaching out to underserved areas, helping to provide educational opportunities to health care professionals in these communities. This paper describes the development of MedReach, a medical information outreach system that connects regional AHEC sites to MCO via the Internet. MedReach provides physicians and other health care professionals access and support to search computerized textbooks and databases for current information on medical diagnoses, treatments, and research. A unique aspect of the MedReach project is that users are able to receive personal help with information retrieval by calling or emailing MCO's outreach librarian. Periodically, the AHEC program and the Mulford Library at MCO also sponsor an educational program, titled "Medical Applications of Computers," for regional practitioners. Current feedback on both the medical information outreach system and the educational program has been positive. PMID- 12113518 TI - Reading factor: a new bibliometric criterion for managing digital libraries. PMID- 12113519 TI - Finding the evidence: teaching medical residents to search MEDLINE. PMID- 12113520 TI - Collaboration to teach graduate students how to write more effective theses. PMID- 12113521 TI - Access to information in war. PMID- 12113522 TI - Medical indexing outside the National Library of Medicine. PMID- 12113523 TI - Update on InfoRetriever software. PMID- 12113524 TI - Papers of two former Medical Library Association presidents cataloged in Philadelphia. PMID- 12113525 TI - Linda A. Watson, Medical Library Association President, 2002-2003. PMID- 12113526 TI - Cell proliferation and apoptosis during histogenesis of the guinea pig and rabbit cerebellar cortex. AB - Cell proliferation and apoptosis are essential for development of the nervous system. In this study we have investigated the histogenesis of the cerebellar cortex in guinea pig (a precocial species) and rabbit (an altricial species) at different stages of pregnancy and postnatal life. Proliferating cells were identified after labeling with antibodies against the proliferating cell nuclear antigen (PCNA) and/or the Ki-67 antigen. Apoptotic cells were visualized in situ by the TUNEL method and by immunodetection of cleaved caspase 3 and 9. In guinea pigs, both proliferating and apoptotic cells were detected during pre-natal life (E0-E40). Conversely, cell proliferation and apoptosis in rabbits were temporally restricted to early postnatal weeks (P0-P20). In both species cell proliferation was mainly linked to differentiation and migration of the granule cells. In both species, the majority of cells undergoing programmed cell death likely corresponded to granule cells. They were mainly detected in the external granular layer, and were by far more common than previously reported in other locations of the postnatal brain. This study shows that apoptosis is a shared process of cell death during cerebellar development in both altricial and precocial animals, and that there is a direct spatial and temporal correlation between cell proliferation and death in two mammals with different time tables in cerebellar maturation. PMID- 12113527 TI - Analysis of morphogenetic potential of caudal spinal cord in Triturus carnifex adults (Urodele amphibians) subjected to repeated tail amputations. AB - The present research was aimed at testing whether the extraordinary morphogenetic and histogenetic potential exhibited in the regenerating new tail remains constant even after repeated amputation or whether it changes as a result of the mechanisms responsible for the regenerative process. Particular attention was focused on regeneration of the spinal cord and ganglia. For this purpose, tail regeneration in adult specimens of Triturus carnifex subjected to repeated amputation (up to 7 times) was compared with that of control animals subjected to a single amputation. Results show that although it slowed down the morphogenetic and differentiative phase, repeated amputation did not significantly alter either the morphogenetic or the histogenetic potential of the ependymal layer of the regenerating spinal cord. The latter result leads to hypothesized that the cells of the ependymal layer of the stump, which are responsible for the formation of the apical ampulla and the ependymal tubule inside the regenerative blastema, do not derive from undifferentiated reserve elements triggered after tail amputation but rather from differentiated ependymal elements that dedifferentiate after the trauma and re-acquire embryonic potential. If this regeneration were actually to take place at the expense of the reserve elements, the continual regenerative processes induced by the repeated amputation would lead to the increasing depletion of these elements and a consequent reduction in regenerative capacity. PMID- 12113529 TI - Physiological asymmetry of trunk ranging and pelvis motility: an anatomo functional study in 80 healthy subjects. AB - A key feature in physiotherapeutic treatment of patients with motion disturbances is the appropriate ranging of the trunk and pelvis motility. Eighty subjects randomly selected and free from known pathology of the muscular-skeletal and/or of the neurological system classed into four groups according to the age and the sex have been assessed, by using a new, simple and easy administrable tool. Our results demonstrate that the new measurement tool showed a very low intra- and inter-observer variability, that healthy subjects showed a more adduced and elevated right scapula if compared to the contralateral one and, as regard as the pelvic motion, a broader joint excursion in passive motion compared with active motion in the overall group, a broader joint excursion in young subjects compared with elderly ones, and a broader joint excursion in female subjects compared with males subjects. In conclusion our study allowed to identify a range of physiological asymmetry and pelvis motility. Such a range of physiological asymmetry might be useful as a reference for the physiotherapists. PMID- 12113528 TI - Immunochemical and immunocytochemical expression of protein kinase c isoenzymes alpha, delta, epsilon and zeta in primary adherent cultures of chick chondrocytes. AB - The family of protein kinase C (PKC) comprises serine/threonine isoenzymes involved in various biological processes, including cell proliferation and differentiation. On the bases of previous investigations performed by us on the expression of various PKC isoforms in the endochondral ossification process of the vertebral column, the aim of the present work was to investigate the expression of various PKC-isoenzymes in chick primary chondrocyte cultures i.e. the most used chondrocyte culture model in vitro. Immunochemical and immunocytochemical experiments were performed to detect the expression of PKC alpha, -delta, -epsilon and -zeta. Chondrocyte cultures were examined two weeks after cell collection from tibiae of 6-day old chick embryos. By means of morphological observations associated with the immunocytochemical expression of type II collagen, two different cell phenotypes were identified, i.e. fibroblast like and polygonal-roundish-shaped cells. As far as PKC-isoenzyme expression was concerned, PKC-zeta revealed a stronger immunochemical and immunocytochemical expression; PKC-alpha exhibited a positivity less marked than PKC-zeta, whereas PKC-delta and -epsilon were less expressed in this culture stage. It is reasonable that a major role could be played by PKC-alpha and -zeta in this phase of the chondrogenic process, whereas PKC-delta and -epsilon could be involved in different stages of chondrocyte differentiation in vitro. PMID- 12113531 TI - Brachial plexus imaging. PMID- 12113530 TI - Isokinetic knee joint test in "gonalgia sine materia". AB - Fifty-four subjects, aged between 20 and 35 years, divided into two subgroups, respectively 30 healthy subjects (17 males and 13 females) and 24 subjects with "gonalgia sine materia" (13 males and 11 females) underwent isokinetic exercise test in order to compare their dominant limb with the not dominant one as regard as the strength of extensor and flexor muscles of the knee. No statistically significant difference was found in any of the studied parameters in the comparison between the dominant limb and the not dominant one, both within the subgroup of healthy subjects and within the subgroup of subjects with "gonalgia sine materia". Authors conclude that psychological features may play a preeminent role in the genesis, as well as in the maintenance of "gonalgia sine materia", thus confirming previous data available in medical literature. PMID- 12113533 TI - Temporary abdominal coverage and reclosure of the open abdomen: frequently asked questions. PMID- 12113532 TI - Pelvic fractures: epidemiology and predictors of associated abdominal injuries and outcomes. AB - BACKGROUND: Pelvic fractures are often associated with major intraabdominal injuries or severe bleeding from the fracture site. OBJECTIVE: To study the epidemiology of pelvic fractures and identify important risk factors for associated abdominal injuries, bleeding, need for angiographic embolization, and death. METHODS: Trauma registry study on pelvic fractures from blunt trauma. Stepwise logistic regression was used to identify risk factors of severe pelvic fractures, associated abdominal injuries, need for major blood transfusion, therapeutic embolization, and death from pelvic fracture. Adjusted relative risks and 95% confidence intervals were derived. RESULTS: There were 16,630 trauma registry patients with blunt trauma, of whom 1,545 (9.3%) had a pelvic fracture. The incidence of abdominal injuries was 16.5%, and the most common injured organs were the liver (6.1%) and the bladder and urethra (5.8%). In severe pelvic fractures (Abbreviated Injury Scale [AIS] > or =4), the incidence of associated intraabdominal injuries was 30.7%, and the most commonly injured organs were the bladder and urethra (14.6%). Among the risk factors studied, motor vehicle crash is the only notable risk factor negatively associated with severe pelvic fracture. Major risk factors for associated liver injury were motor vehicle crash and pelvis AIS > or = 4. Risk factors of major blood loss were age > 16 years, pelvic AIS > or =4, angiographic embolization, and Injury Severity Score (ISS) > 25. Age> 55 years was the only predictor for associated aortic injury. Factors associated with therapeutic angiographic embolization were pelvic AIS > or =4 and ISS > 25. The overall mortality was 13.5%, but only 0.8% died as a direct result of pelvic fracture. The only pronounced risk factor associated with mortality was ISS>25. CONCLUSIONS: Some epidemiological variables are important risk factors of severity of pelvic fractures, presence of associated abdominal injuries, blood loss, and need of angiography. These risk factors can help in selecting the most appropriate diagnostic and therapeutic interventions. PMID- 12113534 TI - A statewide population-based study of gender differences in trauma: validation of a prior single-institution study. AB - BACKGROUND: Women usually have lower mortality rates than men do at any age. This pattern is observed for most causes of death from chronic diseases. Significant controversy still exists about gender differences in outcomes in trauma. We previously reported no differences in in-hospital mortality based on gender in a large single-institution study (n= 18,892) that had a significant limitation in that it was not population based. This current study was performed to validate our earlier findings in a separate, statewide, population-based dataset of trauma victims. STUDY DESIGN: Prospective data were collected on 22,332 trauma patients (18,432 blunt, 3,900 penetrating) admitted to all trauma centers (n = 26) in Pennsylvania over 24 months (January 1996 to December 1997). Gender differences in in-hospital mortality were determined for the entire dataset and for the subsets of blunt and penetrating injury patients. A second analysis examined all blunt injury patients and excluded all patients with a hospital length of stay of less than 24 hours, eliminating patients who expired soon after admission. The null hypothesis was that female gender is protective in trauma outcomes. RESULTS: Multiple logistic regression analysis identified age (odds ratio [OR] 1.03, confidence interval [CI] 1.02 to 1.03), Injury Severity Score (OR 1.06, CI 1.05 to 1.06), non-Caucasian race (OR 1.72, CI 1.39 to 2.15), blunt injury type (OR 0.327, CI 0.26 to 0.41), and Revised Trauma Score (OR 0.44, CI 0.41 to 0.47) as independent predictors of in-hospital mortality in trauma. Preexisting diseases, including cardiac disease (OR 1.53, CI 1.12 to 2.09) and malignancy (OR 4.08, CI 1.64 to 10.17), were also identified as independent predictors of in-hospital mortality in trauma. Female gender was not associated with decreased mortality (OR 0.83, CI 0.67 to 1.03, p = 0.093). A second multiple regression analysis in blunt trauma patients admitted for longer than 24 hours (which eliminated early deaths and patients with minor injuries) determined that in-hospital mortality was not significantly different in male or female blunt trauma patients stratified by Injury Severity Score and age. The same factors that were predictive of in-hospital mortality in the total dataset were also significant in this secondary analysis. CONCLUSIONS: These population-based data confirm that female gender does not adversely affect in-hospital mortality in trauma when patients are appropriately stratified for other variables, including Injury Severity Score and age, that do significantly affect outcomes. PMID- 12113535 TI - When the sun can set on an unoperated bowel obstruction: management of malignant bowel obstruction. PMID- 12113536 TI - Intrahepatic gas. PMID- 12113537 TI - "Giant diverticulum" sigmoid colon. PMID- 12113538 TI - Surgical techniques in right laparoscopic donor nephrectomy. PMID- 12113539 TI - Rectourethral fistulae: the perineal approach. PMID- 12113540 TI - Intraoperative localization of parathyroid glands with gamma counter probe in primary hyperparathyroidism: a prospective study. AB - BACKGROUND: Technetium 99m-sestamibi imaging might be the best method to localize abnormal parathyroid glands. No studies to date have compared preoperative imaging and intraoperative gamma probe localization in patients with primary hyperparathyroidism. STUDY DESIGN: This prospective study included 20 arbitrarily selected patients with primary hyperparathyroidism, verified by elevated serum ionized calcium and intact parathyroid hormone concentrations and low serum phosphatase level. Each patient underwent both preoperative imaging study of the parathyroid glands with technetium 99m-sestamibi (dose 740MBq) and intraoperative localization with a handheld gamma probe. Full collar exploration served as the gold standard. RESULTS: Hypercalcemia and hypophosphatemia normalized in each patient. A single parathyroid adenoma was confirmed histologically in 16 and hyperplasia (4 abnormal glands) in 4 patients. None of the patients had multiple adenomas. The sensitivity of the preoperative scan was 81% (13 of 16 patients) in adenoma patients and 100% (4 of 4 patients) in hyperplasia. The corresponding specificity was 88% and 100%. Intraoperatively only 8 of 16 adenomas were correctly detected (sensitivity 50%), and none of the hyperplastic glands were correctly detected. CONCLUSIONS: In unselected patients with primary hyperparathyroidism, preoperative technetium 99m-sestamibi imaging is more accurate than intraoperative gamma probe detection in localizing abnormal parathyroid glands. PMID- 12113541 TI - Outcomes of simultaneous resection of synchronous esophageal and extraesophageal carcinomas. AB - BACKGROUND: Adequate extent of surgical resection of simultaneous primary esophageal and extraesophageal carcinomas is controversial. STUDY DESIGN: Clinicopathologic records and treatment outcomes of 57 patients undergoing simultaneous resection of both synchronous esophageal and extraesophageal carcinomas (SC group) were reviewed and compared with those of 316 patients receiving esophagectomy for solitary esophageal carcinoma (EC group). RESULTS: Mortality and morbidity rates were 3.5% and 45.6% in the SC group, and 3.2% and 44.3% in the EC group, respectively. No significant difference was detected in either of the rates between the two patient groups. The overall 5-year survival rate of the SC group was 40%. Survival of the patients undergoing curative resection of both esophageal and extraesophageal tumors (n = 30) was significantly better than that of the patients receiving palliative resection of at least one of the two tumors in the SC group (n=27)(5-year survival, 54.2% versus 19.9%, respectively)(p < 0.01). Survival of the SC group patients undergoing curative resection of both tumors (n = 30) did not differ from that of the EC group patients receiving curative esophagectomy (n = 182)(5-year survival rates, 54.2% versus 60.0%, respectively). CONCLUSIONS: Simultaneous resection of synchronous esophageal and extraesoprhageal carcinomas can be safely performed, and complete tumor clearance of both tumors is needed for favorable long-term results. PMID- 12113542 TI - Gum chewing enhances early recovery from postoperative ileus after laparoscopic colectomy. AB - BACKGROUND: Postoperative ileus limits early hospital discharge for patients who have undergone laparoscopic procedures. Sham feeding has been reported to enhance bowel motility. Here, the effect of gum chewing is evaluated as a convenient method to enhance postoperative recovery from ileus after laparoscopic colectomy. STUDY DESIGN: A total of 19 patients who underwent elective laparoscopic colectomy for colorectal cancer participated in the study. Each patient was randomly assigned to one of two groups: a gum-chewing group (n = 10, mean age 58.6 years, range 50 to 71 years) or a control group (n = 9, mean age 60.6 years, range 45 to 80 years). The patients in the gum-chewing group chewed gum three times a day from the first postoperative AM until oral intake. The times of the first passage of flatus and defecation were recorded precisely. RESULTS: The first passage of flatus was seen, on average, on postoperative day 2.1 in the gum chewing group and on day 3.2 in the control group (p < 0.01). The first defecation was 2.7 days sooner in the gum-chewing group (postoperative day 3.1) than in the control group (5.8 days; p< 0.01). All patients tolerated gum chewing on the first operative AM. The postoperative hospital stays for the gum-chewing and control groups were 13.5+/-3.0 days and 14.5+/-6.1 days, respectively. CONCLUSIONS: Gum chewing aids early recovery from postoperative ileus and is an inexpensive and physiologic method for stimulating bowel motility. Gum chewing should be added as an adjunct treatment in postoperative care because it might contribute to shorter hospital stays. PMID- 12113543 TI - Locoregional recurrence and survival after curative resection of adenocarcinoma of the colon. AB - BACKGROUND: There is wide variability in reported locoregional recurrence rates after curative resection of adenocarcinoma of the intraperitoneal colon, and there is no universally accepted surgical technique regarding length of the resected specimen or extent of lymphadenectomy. The aim of this study was to determine the disease-free survival, locoregional failure, and perioperative morbidity of patients undergoing curative resection of colon adenocarcinoma. STUDY DESIGN: The records of 316 consecutive patients undergoing curative resection for primary adenocarcinoma of the intraperitoneal colon between 1990 and 1995 were reviewed. Locoregional recurrence was defined as disease at the anastomosis or in the adjacent mesentery, peritoneum, retroperitoneum, or carcinomatosis. The product-limit method (Kaplan-Meier) was used to analyze survival and tumor recurrence. RESULTS: The study population comprised 167 men and 149 women, mean age 70+/-12 years (range 22 to 95 years). Median followup was 63+/-25 months. Five-year disease-free survival was 84% overall. Disease-free survival paralleled tumor stage: stage I, 99% (n = 73); stage II, 87% (n = 151); stage III, 72% (n = 92). The predominant pattern of tumor recurrence was distant failure only. Overall locoregional recurrence (locoregional and locoregional plus distant) at 5 years was 4%. Locoregional recurrence paralleled tumor stage: stage I, 0%; stage II, 2%; stage III, 10%. Of the 12 patients who suffered locoregional recurrence, 9 (75%) had T4 primary tumors, N2 nodal disease, or both. Major and minor complications occurred in 93 patients (29%) including: anastomotic leak or intraabdominal abscess (n = 4, 1%); hemorrhage (n = 8, 3%); cardiac complications (n= 17, 5%); pulmonary embolism (n=4, 10%); death (n=2, 1%). Multivariate analysis (Cox proportional hazards) revealed that the only independent predictor of disease-free survival and locoregional control was tumor stage. CONCLUSION: Longterm survival and locoregional control can be achieved for patients with colon cancer, with low morbidity. In the absence of adjacent organ invasion and N2 nodal disease, locoregional recurrence should be a rare event. Just as for rectal cancer, the technical aspects of colectomy for colon cancer deserve renewed attention. PMID- 12113544 TI - Hepatocyte protection by a protease inhibitor against ischemia/reperfusion injury of human liver. AB - BACKGROUND: Total clamping of the hepatic pedicle can induce profound hepatic ischemia/reperfusion (I/R) injury, which remains a potentially lethal problem after hepatectomy. STUDY DESIGN: The purpose of this study was to evaluate the efficacy of a protease inhibitor in ameliorating I/R injury of the human liver. In a prospective, randomized, clinical study, 66 patients who underwent liver resection under conditions of continuous inflow occlusion were randomly assigned to three groups: 25 patients were given a synthetic protease inhibitor (gabexate mesilate [GM], 2.0 mg/kg/hr) intravenously starting 24 hours before surgery until postoperative day 3 (preop GM group); 16 were similarly given GM at the beginning of surgery (intraop GM group); and 25 served as controls (without GM group). Laboratory data and intraoperative and postoperative variables were analyzed and plasma levels of cytokines--tumor necrosis factor-alpha (TNF-alpha), interleukin 1beta (IL-1beta), and interleukin-6 (IL-6)--were measured to determine the relationship between surgical stress and hepatic I/R injury. RESULTS: The three groups of patients were similar in terms of age, gender, preoperative assessments, hepatic inflow occlusion time (approximately 50 minutes), extent of resection (proportion of major and minor hepatectomy), and background liver conditions. Preoperative administration of gabexate mesilate (preop GM group) substantially ameliorated hepatic I/R injury as compared with the other patients (intraop and without GM groups); postoperative serum transaminase levels were notably decreased in association with marked suppression of IL-6 levels in blood circulation during liver surgery. This was accompanied by a lower rate of postoperative complications and no mortality. Gabexate mesilate pretreatment abrogated the positive correlation between postreperfusion hepatocyte injury and hepatic ischemia time. CONCLUSIONS: Preoperative administration of GM is useful for preventing I/R injury of the human liver, accompanied by suppression of the plasma proinflammatory cytokine IL-6. PMID- 12113545 TI - The vaginal-psoas suspension repair of uterovaginal prolapse. AB - BACKGROUND: This article introduces a novel technique for the abdominal suspension of the prolapsed vagina. A Gore-Tex (WL Gore & Associates, Flagstaff, AZ) graft interposition between the vaginal apex and the right psoas muscle is performed under direct visualization to effect the repair. STUDY DESIGN: Between June 1998 and June 2001, seven consecutive cases of uterovaginal prolapse were repaired abdominally using the vaginal-psoas technique as outlined here. Patients were followed postoperatively for recurrent prolapse, urinary incontinence, and the development of new or progressive pelvic support defects. RESULTS: The mean operative time was 134 minutes, and the mean blood loss was 160mL for all procedures. One patient was lost to followup after 2 months. The remaining six patients have a median followup of 22 months (range 4 to 36 months), and in these patients, there have been no instances of recurrent prolapse or urinary dysfunction. One patient developed a progression of her rectocele. There were no peroperative or postoperative complications directly related to the procedure. Specifically, there were no instances of hemorrhage, transfusion, small bowel obstruction, graft erosion, or genitofemoral neuropathy. CONCLUSIONS: The vaginal psoas colposuspension appears to be a simple, efficacious technique for the repair of uterovaginal prolapse. No significant morbidity directly related to the procedure has occurred to date. Further studies with longer followup will be required in the future before definitive conclusions can be reached. PMID- 12113546 TI - What's new in general thoracic surgery. PMID- 12113547 TI - What's new in otolaryngology-head and neck surgery. PMID- 12113548 TI - The unresolved problem of recurrent saphenofemoral reflux. PMID- 12113549 TI - Pain control in outpatient surgery. PMID- 12113551 TI - Upregulation of co-stimulatory molecule expression and dendritic cell marker (CD83) on B cells in periodontal disease. AB - T cells and their cytokines are well known for their important role in the pathogenesis of periodontitis. To date, the role of antigen presenting cells (APCs), which are known to be critical in the regulation of T cell response, has been poorly investigated in periodontitis. In this study, we analyzed the expression of co-stimulatory molecules (CD80 and CD86) and CD83, which is a marker of mature dendritic cells, on gingival cells that were isolated from severe periodontitis tissues, with the use of flow cytometry. Significant upregulation of CD86 and CD83 expression was detected in periodontitis lesions, and most of this occurred on B cells. In vitro peripheral blood mononuclear cell cultures showed that stimulation with different periodontopathic bacteria, that included Porphyromonas gingivalis, Actinobacillus actinomycetemcomitans, Prevotella intermedia, and Actinomyces viscosus, upregulated both CD86 and CD83 expression on B cells. Therefore, the presence of plaque bacteria may be responsible for the enhanced expression seen in vivo on gingival B cells. APC function by bacterial-activated B cells was further investigated using allogeneic mixed leukocyte reactions. After 24 h culture with either A. actinomycetemcomitans or P. gingivalis, these activated B cells performed as potent APCs in mixed leukocyte reactions, and they stimulated T cells to produce high levels of gamma interferon and minimal interleukin-5. In conclusion, periodontopathic bacterial-induced B cell activation with upregulation of CD86 and CD83 may be associated with enhanced APC function. The results of this study suggest, therefore, that infiltrated gingival B cells have a possible role as APCs in the regulation and maintenance of local T cell response in periodontitis. PMID- 12113550 TI - Effects of TNFalpha and prostaglandin E2 on the expression of MMPs in human periodontal ligament fibroblasts. AB - Accumulating evidence indicates that TNFalpha plays an important role in the pathogenesis of periodontitis, but the effect of TNFalpha on the degradation of the periodontal ligament is not well understood. This study used reverse transcriptase-PCR to investigate the effects of TNFalpha on matrix metalloproteinase (MMP) mRNA expression in human periodontal ligament fibroblasts. TNFalpha increased MMP-1, MMP-3 and MMP-13 mRNA levels in both a time-dependent (0-24 h) and a dose-dependent (0.1-10 ng/ml) manner. TNFalpha also increased COX-2 mRNA levels. Because elevation of COX-2 mRNA levels enhances the production of prostaglandins, we therefore investigated whether endogenous prostaglandins are involved in the MMP mRNA expression that is enhanced by TNFalpha. Pretreatment with the selective COX-2 inhibitor, NS-398, increased MMP 13 mRNA levels, while prostaglandin E2 and dibutyryl cyclic AMP decreased MMP-13 mRNA levels. Neither MMP-1 nor MMP-3 mRNA levels were affected by these chemicals. These findings indicate that prostaglandin E2 has a lowering effect on TNFalpha-enhanced MMP-13 mRNA levels, and that this effect is dependent on cAMP. Our results suggest that TNFalpha participates in periodontal ligament destruction by stimulating the production of MMPs (MMP-1, MMP-3 and MMP-13), while endogenous prostaglandin E2 has a negative feedback role in TNFalpha enhanced MMP-13 production. PMID- 12113552 TI - The role of LFA-1 in osteoclast development induced by co-cultures of mouse bone marrow cells and MC3T3-G2/PA6 cells. AB - Interactions between leukocyte function-associated antigen-1 (LFA-1) and intercellular adhesion molecule-1 (ICAM-1) influence the development of osteoclasts. However, little is known about how these adhesion molecules are involved in the process of osteoclast development. This study evaluated the role of LFA-1 and its ligands in osteoclast development and bone resorption. Co cultures of bone marrow cells from LFA-1-deficient mice and MC3T3-G2/PA6 (PA6) cells were cultured in the presence of 1alpha,25(OH)2D3 and dexamethasone for 7 days. The number of TRAP-positive cells that were generated by bone marrow cells from LFA-1-deficient mice was smaller than that generated by bone marrow cells from wild-type mice. In addition, the bone-resorbing activity of osteoclast-like cells that were generated from LFA-1-deficient mice was lower than that generated by osteoclast-like cells from wild-type mice. Immunofluorescence flow cytometry showed that osteoclast stromal PA6 cells expressed the cell adhesion molecules, ICAM-1 and VCAM-1. When monoclonal antibodies to mice VCAM-1, CD11b or CD18 were added separately to the co-culture system, the number of TRAP-positive cells that were generated from LFA-1-deficient mice was 20-30% smaller than that generated from wild-type mice. The formation of TRAP-positive cells from both LFA-1 deficient and wild-type mice was especially inhibited by anti-CD18 antibody, in comparison to the addition of normal IgG serum. These results suggest that LFA-1 adhesion molecules play a role in osteoclast development by affecting adhesion between stromal cells and osteoclast progenitors before the occurrence of ODF-ODF receptor signaling. CD18 appears to be a key adhesion molecule in cell-to-cell contacts during the early stage of osteoclast development. PMID- 12113553 TI - Morphological evaluation of combined effects of cyclosporin and nifedipine on gingival overgrowth in rats. AB - It has previously been shown that, while cyclosporin A (CsA) and nifedipine both cause gingival overgrowth in the rat, the combined use of these drugs increases the severity of overgrowth. The aim of this study was to describe the histometry and densities of fibroblasts, collagen fibers and vessels in the gingival tissue of rats that were treated with CsA and nifedipine, either alone or in combination. Rats were treated for 60 days with a daily subcutaneous injection of 10 mg/kg body weight of CsA and/or with 50 mg/kg body weight of nifedipine added to the chow. The results confirmed that CsA causes a more severe overgrowth than nifedipine, and that the combined use of these drugs increases the overgrowth severity. All the rat groups that were studied showed that, as the severity of overgrowth increased, there was a parallel increase in fibroblasts and collagen, and a decrease in vessel content. Therefore, independently of whether the gingival overgrowth was caused by CsA alone, nifedipine alone, or both treatments in combination, the fibroblast and collagen density increased in parallel with the severity of the overgrowth. PMID- 12113554 TI - Regulation of matrix metalloproteinase production by cytokines, pharmacological agents and periodontal pathogens in human periodontal ligament fibroblast cultures. AB - Matrix metalloproteinases (MMPs). produced by both infiltrating and resident cells of the periodontium, play a role in physiologic and pathologic events. It is recognized that an imbalance between activated MMPs and their endogenous inhibitors leads to pathologic breakdown of the extracellular matrix during periodontitis. To date, little is known about the regulation of MMP synthesis and secretion in human periodontal ligament fibroblasts (PDLFs). The purpose of this study was to examine the effects of cytokines, pharmacological agents (protein synthesis inhibitor and protein kinase C inhibitors) and predominant periodontal pathogens (Actinobacillus actinomycetemcomitans and Porphyromonas gingivalis) on MMP production in human PDLFs using gelatin zymography. The gelatin zymograms revealed that the main gelatinase secreted by human PDLFs migrated at 72 kDa and represents MMP-2. Minor gelatinolytic bands were also observed at 92 kDa regions that correspond to MMP-9. We found that A. actinomycetemcomitans, P. gingivalis and IL-1alpha can elevate MMP-2 secretion in human PDLFs. These results indicate that periodontal pathogens and inflammatory cytokines play an important role in tissue destruction and disintegration of extracellular matrix in periodontal diseases. Thus, activation of MMPs may be one of the distinct host degradative pathways in the pathogenesis of periodontitis. In addition, H7, staurosporine, cycloheximide and TGF-beta could suppress MMP-2 production. Agents that target protein synthesis or the protein kinase C pathway in human PDLFs inhibit MMP-2 production, and such inhibition may contribute to the pathogenesis of periodontal inflammation. Taken together, these findings suggest a possible new therapeutic approach, involving the use of drugs that modify host-response mechanisms to suppress or inhibit MMP-mediated tissue destruction. PMID- 12113555 TI - Hard tissue formation on dentin surfaces applied with recombinant human bone morphogenetic protein-2 in the connective tissue of the palate. AB - The purpose of this study was to evaluate whether hard tissue might be formed on dentin surfaces applied with recombinant human bone morphogenetic protein-2 (rhBMP-2) in palatal connective tissue. Fifty-eight dentin blocks were prepared from rat roots, demineralized with 24% EDTA (pH 7.0), applied with 0, 50 and 100 microgram/ml rhBMP-2, and labeled as groups 0, 50 and 100. The dentin blocks were then transplanted into palatal connective tissue of rats, and specimens were prepared at two and four weeks after surgery for histologic and histomorphometric examinations. The results showed that the percentage of newly formed hard tissue in relation to the total dentin block surface length in groups 0, 50 and 100 was 0.0%, 2.8% and 4.4% at two weeks, and 0.0%, 1.6% and 12.8% at four weeks, respectively. New hard tissue formation in groups 50 and 100 was significantly promoted as compared to group 0 (p < 0.01). These findings thus indicate that rhBMP-2 application to dentin enhanced new hard tissue formation on dentin surfaces in the connective tissue of the palate. PMID- 12113556 TI - Treatment with SRL172 (heat-killed Mycobacterium vaccae) inhibits progression of established experimental periodontal disease in Wistar rats. AB - Periodontal disease is accompanied by a change in the periodontal bacterial flora, and an increase in Th2 cytokine expression. Therefore, preparations that can down-regulate Th2 responses and increase Th1 responses to bacteria may have therapeutic effects. SRL172, a preparation of heat-killed Mycobacterium vaccae (NCTC11659), has been shown to have these properties in man and animals and, in a previous study, a single subcutaneous injection of this material 13 days before application of ligatures to the right second molars of Wistar rats strikingly reduced subsequent destruction of the tooth-supporting tissues. The same material has therefore been tested in a therapeutic protocol, in rats with ongoing periodontal disease. A silk ligature was placed round the maxillary right second molar in the gingival sulcus on day 0. This was removed after two weeks, and the animals immediately received 0.1 mg SRL172 s.c. or saline. One week later (i.e. at three weeks), a boost of 1.0 mg SRL172 or saline was given. Animals were sacrificed at eight weeks (i.e. five weeks after the booster dose). Treatment of ongoing periodontal disease with SRL172 significantly reduced fibre loss (p = 0.046 by histometry) and bone loss (p = 0.0008 by radiography) on the ligatured side, and reduced fibre loss (p = 0.0086) on the control side. Thus SRL172 is an effective treatment in this model. As SRL172 has undergone extensive safety testing in man, these results justify clinical studies in periodontal disease, and such studies are now planned. PMID- 12113557 TI - Recognition of the carbohydrate modifications to the RgpA protease of Porphyromonas gingivalis by periodontal patient serum IgG. AB - Periodontal infections by Porphyromonas gingivalis are associated with a sustained systemic IgG antibody response and elevations in local antibody synthesis to this organism. One of the targets of this response is a protease, RgpAcat, which is an important virulence determinant of this organism. Recently, we demonstrated that this molecule is glycosylated and that the glycan chains are immunologically related to P. gingivalis lipopolysaccharide (LPS) (Curtis et al., Infect Immun 1999;62:3816-3823). In the present study, we examined the role of these glycan additions in the immune recognition of RgpAcat, by sera from adult periodontal patients (n = 25). Serum IgG antibody levels to P. gingivalis W50, RgpAcat and LPS and to recombinant RgpA were determined by enzyme-linked immunosorbant assay (ELISA). No correlation was observed between the antibody levels to RgpAcat from P. gingivalis and the recombinant form of this enzyme expressed in Escherichia coli. However, a strong association was found between the recognition of LPS and the wild-type enzyme (R = 0.8926; p = 0.0005). Incorporation of LPS into the ELISA led to a significant reduction (mean 25%; range 0.8-43%, SD = 15; p < 0.05) in the recognition of RgpAcat, but had no effect on the recognition of control antigens. Deglycosylation of RgpAcat led to the abolition of immune recognition by patient serum IgG, which suggests that the glycan additions to this molecule are the principal targets of the immune response. Therefore, glycosylation of the RgpAcat protease may play an important role in immune evasion by shielding the primary structure from immune recognition. PMID- 12113558 TI - Simple and rapid detection of serum antibody to periodontopathic bacteria by dot blotting. AB - The aim of this study was to detect the specific immunoglobulin G antibodies against periodontopathic bacteria by dot blotting. In the procedure used, bacterial preparations were blotted on a nitrocellulose membrane. After blocking the nonspecific binding sites, the diluted serum was blotted onto the preparations. The membrane was immersed in secondary antibodies and then in substrate buffer. The colored blots were then evaluated. To test the reliability of this procedure, 20 serum samples were examined for antibody: ten for anti Porphyromonas gingivalis antibody, and the other ten for anti-Actinobacillus actinomycetemcomitans antibody. Five samples out of each set of ten had previously been confirmed as having high enzyme-linked immunosorbant assay (ELISA) titers to the antigen, while the other five had been confirmed as having average titer levels. Both whole-cell sonic extracts and fimbriae of P. gingivalis were used as antigens in the dot blotting, in order to compare their use as antigens in assays of the patients' sera. ELISA was also used to measure anti-P. gingivalis antibody titers. For the measurement of IgG antibodies against A. actinomycetemcomitans, formalin-killed whole cells were used. Fifty serum samples were examined for IgG antibodies against A. actinomycetemcomitans by dot blotting and ELISA. With both antigens, after 4 h, coloration of blots was more clearly visible for the high-titer sera than for the average-titer sera. The intensity of coloration of the blots for P. gingivalis and A. actinomycetemcomitans showed correlation with the ELISA titers. A particularly significant correlation was shown when P. gingivalis fimbriae were used as antigen. These results suggest that this dot blot method is a simple and rapid means of detection of serum antibodies, and that it shows promise as a chair-side assay method. PMID- 12113559 TI - The volatile fraction of cigarette smoke induces alterations in the human gingival fibroblast cytoskeleton. AB - Several in vitro investigations have indicated that the particulate phase of cigarette smoke, such as nicotine, affects many cell types, including gingival fibroblasts. However, few studies have been performed on the effects of the volatile fraction on the cellular structures that are involved in cell functions, such as adhesion and proliferation. Since the survival and reproduction of gingival fibroblasts are fundamental in maintaining the integrity of the oral connective tissue, as well as in wound healing, the effects on the cytoskeleton of acrolein and acetaldehyde, which are the volatile fractions of cigarette smoke, were examined in vitro for human gingival fibroblasts (HGFs). HGF strains that were taken from healthy subjects with non-inflamed-gingiva were utilized in this investigation. The cells were incubated in the presence of different concentrations of acrolein and acetaldehyde. Cell adhesion and viability were evaluated after incubation for 3 h and 5 days, respectively. The influence on cytoskeletal structures (tubulin, actin and vimentin intermediate filaments) was investigated with the indirect immunofluorescence technique. The results show that both substances produced similar effects, which resulted in a dose-dependent inhibition of HGF adhesion and viability. Disturbance of the HGF cytoskeleton consisted of disruption of the microtubules, actin filaments and vimentin microfilaments, which was accompanied by alterations to cell shape. Our experimental findings suggest that the volatile fractions of cigarette smoke, such as acrolein and acetaldehyde, have a cytotoxic effect on HGFs, with the result that they lose their capacity for adhesion and proliferation. The consequences of this could be impairment of the maintenance, integrity and remodelling of the oral connective tissue. According to our morphological evidence, these findings show that cigarette smoke can lead to the development and progression of periodontal disease, and indicate the need for appropriate therapy. PMID- 12113560 TI - Lacrimal function and ocular complications in patients treated with systemic isotretinoin. AB - PURPOSE: To evaluate the effect on lacrimal function and ocular complications in patients with severe acne vulgaris during systemic treatment with 13-cis-retinoic acid (isotretinoin). METHODS: Forty patients with acne vulgaris were treated with systemic isotretinoin at dosages of 0.5-1 mg/kg per day for two months. Full ophthalmologic examination, Schirmer I test, fluorescein break-up (BUT) and microbiological investigations of the conjunctival flora were done before, during the second month and at least one month after the end of the treatment. RESULTS: The average Schirmer values before and after the treatment were 21.6 mm/5 minutes (SD +/- 7.01) and 18.48 mm/5 minutes (SD +/- 7.87) respectively. After the treatment BUT was less than 10 seconds in 50% of the patients and 55% had blepharitis. Subjective symptoms like dryness, itching and contact lens intolerance occurred in 42.5% and colonization of the conjunctiva by Staphylococcus aureus increased significantly during treatment (p= 0.031). All abnormal findings disappeared one month after the cessation of treatment. DISCUSSION: Isotretinoin causes signs and symptoms of dry eye, probably by reducing meibomian gland function, but ocular complications are generally not serious when low doses are used for a limited time, and are reversible after discontinuation. PMID- 12113562 TI - Comparison of the effects of argon and neodymium:YAG laser iridotomy on cytokines in the rabbit aqueous humor. AB - PURPOSE: We compared the effects in rabbits of iridotomy using the argon or Nd:YAG laser on cytokines such as interleukin 1-beta (IL-1beta), interleukin 6 (IL-6), interleukin 8 (IL-8), and tumor necrosis factor-alpha (TNF-alpha). METHODS: Twenty pigmented rabbits (20 eyes) underwent argon and Nd:YAG iridotomy under general and topical anesthesia. Group 1 (10 right eyes) was treated with the argon laser and group 2 (10 right eyes) with the Nd:YAG laser, using an equal number of shots and the same laser parameters for each group. Left eyes in both groups were evaluated as controls. Aqueous humor specimens were collected from each eye preoperatively and 1 and 3 days postoperatively. RESULTS: Aqueous IL-6 and TNF-alpha levels rose more on day 1 in the Nd:YAG group than the argon group. IL-6 and TNF-alpha levels were significantly higher on days 1 and 3 than the pre operative and control levels (p<0.05). TNF-alpha levels on day 1 were significantly lower in the Nd:YAG than the argon group. There were no significant differences between the two laser groups for IL-6 on days 1 and 3. IL-1beta and IL-8 did not change CONCLUSIONS: Our findings suggest that cytokines, particularly IL-6 and TNF-alpha, may be inflammatory mediators in the early inflammation following argon and Nd:YAG laser iridotomies. These results also indicate that cytokines contribute to the acute effects of Nd:YAG and argon laser applications on inflammation. PMID- 12113561 TI - Limbal-conjunctival autograft transplantation for recurrent pterygium. AB - PURPOSE: To assess the usefulness of limbal-conjunctival autograft transplantation (LCAT) for the treatment of recurrent pterygium. PATIENTS AND METHODS: Seventeen eyes with advanced recurrent pterygium underwent LCAT All had already been treated at least twice either by simple excision (n=15) or by conjunctival rotation autograft (n=2). Three eyes (17.65%) had symblepharon at the time of surgery, so LCAT was combined with amniotic membrane transplantation. The autograft was taken from the supero-lateral part of the same eye and transferred to the area where the pterygium had been excised. RESULTS: During 6 18 months of follow-up no postoperative complications occurred. In 15 eyes (88.24%) no pterygium recurrence was recorded; recurrence occurred in two eyes (11.76%) after 8 and 5 months. In three eyes with a combined symblepharon formation, remission of both pterygium and symblepharon growth was obtained. CONCLUSIONS: LCAT seems to be a promising and safe procedure for recurrent pterygium. PMID- 12113563 TI - A simplified technique for non-penetrating deep sclerectomy. AB - PURPOSE: To present a modified technique for non-penetrating deep sclerotomy (NPDS) with limited use of sharp instruments. MATERIALS AND METHODS: A special fine spatula is used to separate the Descemet membrane from stromal tissue afterpreparing superficial and deep scleral flaps and opening the Schlemm's canal. Another specially designed spatula is used to protect the membrane when excising the overlying trabecular and stromal tissue. The other steps of the surgery are identical with the original Kozlov method. The modified surgery was performed by one surgeon experienced in NPDS, and three others starting their transition to this technique under the guidance of the first. Sixty-six patients (66 eyes) with primary open-angle glaucoma (51) or pseudo-exfoliature glaucoma (15) were treated with NPDS. RESULTS: Two microperforations (3%) of the Descemet membrane were the only intraoperative complications. They did not require iridotomy or any special treatment. In the early postoperative period one 2-mm hyphema was observed, and in five cases there was transient hypotony. In one other case there was a choroidal detachment, which resolved spontaneously in less than two weeks. IOP the day after surgery ranged from 5 to 13 mmHg (mean 9.6) and after one year from 10 to 22 mmHg (mean 14.6). Late complications and IOP control were similar to conventional NPDS. CONCLUSIONS: These procedures performed by one experienced and three inexperienced surgeons gave satisfactory results and the modified method can be recommended for surgeons beginning nonpenetrating glaucoma surgery, and should shorten their learning curve. PMID- 12113564 TI - Effect of intravenous droperidol on intraocular pressure and retrobulbar hemodynamics. AB - PURPOSE: Topically-applied dopamine antagonists reduce intraocular pressure (IOP) and inrease retinal blood flow in animal models. We examined the acute effects of intravenous infusion of a dopamine blocker (droperidol) on these parameters in healthy humans. METHODS: Sixteen subjects free from ocular or systemic disease (mean age 33 +/- 10 yrs) received either 5 mg i.v. droperidol over 5 minutes, or i.v. saline placebo in double-masked fashion. IOP was determined 30 and 60 minutes later, while color Doppler imaging was used to determine flow velocities in the ophthalmic, central retinal, and nasal and temporal posterior ciliary arteries 60 minutes after drug infusion. RESULTS: 30 minutes after drug infusion, IOP was reduced 6.0 mmHg as compared with baseline (p<0.001); after 60 minutes, IOP remained reduced by 3.7 mmHg (p<0.001). Placebo had no effect on IOP. While droperidol slightly elevated blood pressure and increased the calculated ocular perfusion pressure, the drug reduced visual acuity and contrast sensitivity (p<0.05). Droperidol elevated peak systolic velocity in the central retinal and nasal posterior ciliary arteries, without changing end-diastolic velocity or the resistance index in either of these vessels. Droperidol had no effect on flow velocities in the ophthalmic artery or the temporal posterior ciliary artery. CONCLUSIONS: The rapid and marked ocular hypotension resulting from intravenous droperidol suggests that this agent may prove useful in the management of acute ocular hypertension. The retrobulbar changes consequent to the ocular tension reduction likely represent autoregulatory responses to altered ocular perfusion pressure. PMID- 12113565 TI - Phacofragmentation without perfluorocarbon liquid for dislocated crystalline lenses or lens fragments after phacoemulsification. AB - PURPOSE: To evaluate the clinical outcome of vitrectomy with phacofragmentation without perfluorocarbon liquid (PFCL) in the management of dislocation of the crystalline lens, caused by trauma, hereditary disorders, or hypermature cataracts, and lens fragments due to phacoemulsification. METHODS: A prospective study was conducted to evaluate 30 eyes of 29 patients who had undergone standard pars plana vitrectomy with phacofragmentation without PFCL in the vitreous cavity, for the removal of dislocated crystalline lenses or lens fragments, from January 1998 to July 2000. All cases were followed for more than six months. RESULTS: Final best corrected visual acuity of 0.5 or better increased from 0% preoperatively to 36.7% postoperatively, and 0.2 or better rose to 76.7%. The mean IOP was significant reduced, from 26.35 mmHg to 12.75 mmHg. No intraoperative complications occurred. Although two eyes (6.7%) developed retinal detachment, one (3.3%) had cystoid macular edema (CME) and one (3.3%) had a transient intraocular pressure increase (to 25 mmHg) postoperatively, all others had a favorable outcome. CONCLUSIONS: Standard pars plana vitrectomy with phacofragmentation without PFCL in the vitreous cavity is a safe, simple, and effective method for removing a dislocated crystalline lens or lens fragments, with good visual outcome. PMID- 12113566 TI - Comparison of the clinical performance of Healon 5 and Healon in phacoemulsification. AB - PURPOSE: Healon 5 is a high-molecular-mass fraction of sodium hyaluronate. Its density endows it with a number of viscoelastic characteristics. In this prospective, randomised clinical study we compared the performance of Healon 5 and Healon in phacoemulsification. SETTING: Institute of Ophthalmology, University of Modena and Reggio Emilia, Italy. METHODS: Two groups of patients underwent phacoemulsification and intraocular lens (IOL) implantation. In the first 27 patients Healon 5 was used as viscoelastic substance during surgery, and in the second 27 Healon was used. The surgeons subjective comments on the performance of these viscoelastic agents were recorded at the different steps of surgery: injection, capsulorhexis, phacoemulsification, IOL implantation, removal of viscoelastic agent and trasparency throughout the operation. The surgeon's overall impression of the viscoelastics during the whole operation was noted. Tonometry and endothelial cell count were performed in all patients before and after operation. RESULTS: There was no statistical difference between the two groups as regards visual acuity, ocular pressure and endothelial damage. Healon 5 showed excellent ability to maintain the anterior chamber during capsulorhexis, phacoemulsification and IOL implantation. Removal time with Healon 5 was not appreciably longer than Healon. CONCLUSIONS: Healon 5 emerges as a very interesting viscoelastic substance. Visibility is better if the anterior chamber is filled completely. Removal is easier if it is aspirated while moving the irrigation aspiration tip with circular movements over the top and around the border of the IOL. PMID- 12113567 TI - Posterior capsular opacification in pseudophakic eyes with a silicone or acrylic intraocular lens. AB - PURPOSE: To compare posterior capsular opacification in eyes with IOL of two different materials--silicone or acrylic. METHODS: Eighty consecutive eyes undergoing cataract surgery were prospectively randomized in two groups, 40 eyes receiving a silicone (Sl--30NB) and 40 eyes an acrylic (Acrysof MA60BM) intraocular lens (IOL). The same surgeon performed phacoemulsification and the intraocular lens (PHACO IOL) operation in all cases. Patients were re-examined on the first postoperative day, after one week, four months, and 1-2.4 years. Seven eyes were lost to late control. RESULTS: Clinically significant posterior capsular opacification (PCO) (including eyes with capsulotomy already performed) was equally common in both groups; 25% in the silicone group and 19% in the acrylic group (p=0.53). The posterior capsule remained clear in 61% of the silicone and 76% of the acrylic IOL eyes (p=0.18). In the whole study group, 29% of eyes with and 14% without concurrent ocular diseases had significant PCO (p=0.13). In the silicone IOL group, PCO was more common in eyes with concurrent ocular diseases (44%) than eyes without other diseases (10%) (p=0.049). Eyes with acrylic IOL showed no difference in significant PCO, with or without other diseases (18% and 20%, respectively). CONCLUSIONS: In a consecutive series of 80 cataract eyes central PCO was equally common in eyes receiving a silicone or an acrylic IOL. In the silicone IOL group, however, significant PCO was more common if there was concurrent ocular disease, while with the acrylic IOL concurrent ocular disease did not seem to increase the risk of PCO. PMID- 12113568 TI - Original three-point fixation technique for sutured posterior chamber intraocular lens. AB - PURPOSE AND METHODS: In order to avoid the complications associated with posterior chamber intraocular lens (IOL) scleral fixation, the authors have developed an original surgical technique by which the IOL is secured at the ciliary sulcus by suturing the haptics to the sclera in three points (at the 3, 5 and 9 o'clock positions). This technique was utilized for secondary IOL implantation in 21 aphakic eyes. The mean follow-up was 18 months, range 6-28 months. RESULTS: All eyes that underwent secondary implants had equal or better visual acuity postoperatively; none developed serious intra- or postoperative complications. No tilt or decentration of the IOL was observed postoperatively. DISCUSSION: The technique described appeared easy to perform and produced good visual outcomes with stable transscleral fixation of the IOL. PMID- 12113569 TI - Epidemic endophthalmitis after cataract surgery. AB - PURPOSE: We analyzed the results of pars plana vitrectomy in group of patients with a view to establishing risk factors, optimal therapy, surgical technique and the best timing of the pars plana vitrectomy. METHODS: Eight patients presented features of bacterial endophthalmitis within two days of cataract extraction. We examined the relations between visual outcome and the identity of infecting species, optimal therapy, surgical technique and vitrectomy timing. RESULTS: Pseudomonas aeruginosa was a pathogenic microbe. Unfortunately the source of infection was not found. The long-term (9-12 months) results are not good. Final visual acuity of all eight patients oscillated between 20/200 and no light perception. Three patients were first treated with intravitreal ATB application and vitrectomy followed with 36 hours lateney. Their final VA was no light perception in two patients and hand motion in one. The outcome was better in five patients operated immediately after the onset of endophthalmitis. Final visual acuity in this group was between hand motion and 20/200. CONCLUSIONS: Visual prognosis in cases of endophthalmitis is closely related to the type of infecting organism, the visual acuity at presentation, and the speed of progression of inflammatory signs. The need for prompt vitrectomy as the only chance of retaining at least basic visual functions is fully demonstrated. PMID- 12113570 TI - Eye health care in Hungary. AB - PURPOSE: To describe eye health-care services provided by in patient Departments of Ophthalmology in Hungary as of 1998. METHODS: A standardised questionnaire was sent to all Hungarian in-patient ophthalmic departments. The response rate was 100%, and data from six university departments and 56 hospitals with ophthalmic units are summarised. RESULTS: During the 12-month period, a total of 78,008 ophthalmic operations were performed, by 489 ophthalmologists. They worked in 62 in-patient ophthalmic units having a total of 1952 beds. The cataract rate was 3564 operations per million inhabitants; intraocular lens implantation was performed on 97% of the cases. There were 591 corneal transplants, 1698 operations for retinal detachment and 510 vitrectomies for diabetic eye complications. CONCLUSIONS: The level of ophthalmic care in Hungary, judged on the basis of key objective parameters including number of ophthalmic specialists, number of ophthalmic beds, and the rates of surgery, generally conforms to standards prevailing in Western Europe, in spite of financial difficulties and the consequent lack of investment in new equipment and instruments, both major and minor. PMID- 12113571 TI - Ocular complications associated with brucellosis in an endemic area. AB - PURPOSE: To report the ocular manifestations associated with brucellosis in an endemic area. METHODS: We prospectively evaluated 147 patients with the diagnosis of brucellosis between May 1996 to May 2000 and recorded the ocular and systemic findings. The diagnosis was based on clinical findings, positive serological and bacteriological tests (Brucella agglutination test: over 1/160 titer, blood culture). RESULTS: Thirty-eight patients (26.0%) with brucellosis had ocular complications: conjunctivitis in 26 (17.7%), anterior uveitis in six (4.1%), posterior uveitis in one (0.7%), dacryoadenitis in two (1.4%), episcleritis in three (2.1%). Three of the seven patients with uveitis had spondylitis associated with brucellosis. Osteoarticular complications in brucellosis were more frequent in the patients with ocular involvement though the difference was not statistically significant compared with patients without ocular involvement. CONCLUSIONS: Ocular manifestations are frequent in brucellosis so an ophthalmologic examination should be routinely performed in patients with brucellosis in endemic areas. PMID- 12113572 TI - Divergence insufficiency and demyelinating disorder. AB - PURPOSE: To describe a divergence insufficiency in a young woman. METHODS: Case report. RESULTS: A 38-year-old woman presented with a episode of homonymus horizontal diplopia at distance. She was orthophoric at near but had esotropia at distance. Neurological evaluation was normal but multiple demyelinating lesions were shown in the magnetic resonance scan, with increased intrathecal Ig G production. Double vision improved after corticosteroid mega-doses. CONCLUSIONS: An acute onset of diplopia due to divergence insufficiency in a young adult may be associated with a demyelinating disorder. PMID- 12113573 TI - Dacryoadenitis as the earliest presenting manifestation of systemic Wegener's granulomatosis. AB - PURPOSE: To report a case presenting with dacryoadenitis as the earliest manifestation of systemic Wegener's granulomatosis (WG). DESIGN: Observational case report. METHODS: A 41-year-old woman initially presented symptoms of bilateral dacryoadenitis. She subsequently developed upper and lower respiratory tract involvement, scleritis and keratitis. RESULTS: Cytoplasmic antineutrophil antibody (c-ANCA) titer was positive. The lacrimal gland and lung biopsies were consistent with WG. The patient responded well to cyclophosphamide and prednisolone. CONCLUSIONS: Dacryoadenitis maybe the earliestpresenting manifestation of WG andprompt immunosuppressive chemotherapy may control it preventing the limited disease from progressing to a complete form and reducing its morbidity and mortality. PMID- 12113574 TI - A case of lichen planus-lupus erythematosus overlap syndrome with eyelid involvement. AB - PURPOSE: To report a case of lichen planus-lupus erythematosus overlap syndrome with eyelid involvement. Lichen planus and lupus erythematosus infrequently coexist in the same patients. Ocular involvement has rarely been reported for both diseases. CASE REPORT: We describe a case of lichen planus-lupus erythematosus overlap syndrome with eyelid involvement. Histopathologic and immunofluorescent studies were done on buccal, lip, left conjunctival, malar, auricular and scalp lesions. The immunopathologic features of the conjunctiva, buccal mucosa and lip were consistent with lichen planus, while those of the malar, auricular and scalp lesions favoured lupus erythematosus. RESULTS: The patient was successfully treated with hydroxychloroquine 200 mg/day and all lesions responded to therapy within weeks. CONCLUSIONS: This is a rare example of two coexisting autoimmune disease entities: lichen planus of the oral mucosa, lip, eyelid and discoid lupus erythematosus of the skin. To our knowledge, this is the first reported case of lichen planus-lupus erythematosus overlap syndrome with eyelid involvement. PMID- 12113575 TI - Macular phototrauma after cataract extraction and multifocal lens implantation: case report. AB - PURPOSE: To report a patient who developed photic injury after cataract surgery and multifocal MF4 lens implantation METHODS: A 41-year-old caucasian woman without antecedents of interest was subjected to left catarct surgery involving phacoemulsification with capsular sac implantation of a new type of multifocal lens. A coaxial light microscope was used for surgery. The operation was filmed and anterior pole photographs were obtained. RESULTS: Two days after the operation, the patient noted a paracentral scotoma and hand movement visual acuity in the left eye. Indirect ophthalmoscopy revealed an oval, hyperpigmented macular lesion compatible with phototoxic maculopathy. CONCLUSIONS: Phototoxic injury to the macula may occur after cataract extraction. Implantation of an intraocular lens is an important factor in the producion of maculopathy, on account of its light-focusing effect on the retina. This effect was probably increased in our patient by the use of a new autofocus multifocal lens. PMID- 12113576 TI - Production of a monoclonal antibody directed against the recombinant SEL1L protein. AB - SEL1L, highly similar to the C elegans sel-1 gene, is a recently cloned human gene whose function is under investigation. SEL1L is differentially expressed in tumors and normal tissues and seems to play a role in tumor growth and aggressiveness. We used the recombinant N-terminus of the SEL1L protein to immunize a Balb/c mouse and produce a monoclonal antibody. A hybridoma secreting an antibody specifically reacting on the SEL1L recombinant fragment was selected. This monoclonal antibody, named MSel1, recognizes the SEL1L protein by Western blotting, immunofluorescence and immunohistochemistry on normal and tumor cells. MSel1 is able to recognize SEL1L even on archival tumor specimens and is therefore particularly appropriate to study SEL1L involvement in tumor progression. PMID- 12113577 TI - Serum and cerebrospinal fluid human chorionic gonadotropin (hCG) and alpha fetoprotein (AFP) in intracranial germ cell tumors. AB - We report a retrospective study on serum and cerebrospinal fluid (CSF) alpha fetoprotein (AFP) and beta-human chorionic gonadotropin (betahCG) determination in a series of 30 patients bearing intracranial germ cell tumors. At diagnosis five patients had high serum and CSF AFP levels. No patient had positive serum AFP and negative CSF AFP or vice versa. Twelve of 30 patients had serum betahCG levels above 5 mlU/mL, eight had high betahCG only in CSF, and ten were completely negative. During treatment and follow-up both markers were accurate indicators of the response to therapy, decreasing rapidly and often becoming normal already after the first phase of treatment. We conclude that these two markers, and mostly betahCG, may be useful in the diagnosis and monitoring of the response to therapy of patients with intracranial germ cell tumors. PMID- 12113578 TI - Vascular endothelial growth factor levels in serum and plasma following esophageal cancer resection--relationship to platelet count. AB - In patients with cancer circulating vascular endothelial growth factor (VEGF) may be tumor-derived and have prognostic significance. Activated platelets may also be a source of VEGF, releasing it in serum formation. Debate exists as to whether serum or plasma VEGF (S-VEGF, P-VEGF) is the most appropriate surrogate marker of tumor angiogenesis. As healing wounds produce VEGF that can spill over into the circulation, we aimed to investigate the potential confounding effects of cancer surgery on both perioperative S-VEGF and P-VEGF levels and to evaluate their relationship with platelet count. S-VEGF, P-VEGF and platelet counts were measured in 23 patients undergoing esophageal cancer resection. Samples were taken preoperatively and six weeks following surgery. Seven patients were also sampled on postoperative days 1, 5 and 10. VEGF was assayed using a commercial enzyme linked immunosorbent assay. S-VEGF and P-VEGF both rose after surgery (S VEGF; day 5: 1017 [446-1224] pg/mL and day 10: 1231 [626-2046] pg/mL versus pre op: 329 [189-599] pg/mL. P-VEGF; day 1: 55 [46-104] pg/mL and day 10: 58 [20-154] pg/mL versus pre-op: 23 [13-46] pg/mL), falling towards preoperative levels by six weeks. Platelet count correlated with S-VEGF (rho=0.281; p<0.05, Spearman's rank) and P-VEGF (rho=0.330; p<0.01, Spearman's rank). Platelets may contribute to VEGF levels in plasma as well as in serum. The effects of surgery on S-VEGF or P-VEGF levels are mainly transient. Care must be exercised when interpreting circulating VEGF levels in the early postoperative period. PMID- 12113579 TI - Clinical significance of pepsinogen C tumor expression in patients with stage D2 prostate carcinoma. AB - Pepsinogen C is an aspartyl protease mainly involved in the digestion of proteins in the stomach, and an androgen-inducible protein in breast cancer cells. The aims of this study were to evaluate the expression and clinical significance of this enzyme in the primary tumors of prostate cancer patients with bone metastasis who were scheduled to receive antiandrogenic therapy. This study was prospectively performed in 28 stage D2 prostate cancer patients who, after diagnosis, received maximum androgen blockade. Pepsinogen C tumor expression was analyzed in samples (24 from needle biopsy cylinders and four from transurethral resection specimens) from primary tumors using an immunohistochemical assay. Twelve prostate carcinomas (42.8%) were positive for pepsinogen C. Pepsinogen C was a significant prognostic factor to predict a longer overall survival in the patients of our study (p<0.01). Pepsinogen C can be a new prognostic factor and a useful biological marker of androgen dependency in prostate cancer. PMID- 12113581 TI - The half-life of thymidine kinase 1 in serum measured by ECL dot blot: a potential marker for monitoring the response to surgery of patients with gastric cancer. AB - Thymidine kinase 1 in serum (STK1) of patients with gastric cancer was determined by two methods: ECL dot blot and radioactivity assay. Both measurements showed significantly different values for preoperative STK1 and healthy STK1 (p=0.012 for ECL dot blot and p=0.003 for the radioactivity assay). The preliminary results of ECL dot blot STK1 measurement showed that in tumor-free subjects the level of the enzyme was significantly reduced to 52.7% 35 days after surgery (n=8, p=0.0106). The decrease in STK1 levels in the tumor-free subjects paralleled the decline of the half-life of the STK1 enzyme. In patients with distant metastases (n=6) the enzyme level had increased to 173% 35 days postoperatively. By contrast, with the radioactivity assay no significant differences in thymidine kinase activity for 0-day-postoperative patients and 35 day-postoperative tumor-free patients was found (p=0.329). The activity decreased to 80% in 35-day-postoperative patients with metastatic disease. We suggest that the value of the half-life of STK1 measured by ECL dot blot can be used as a potential marker for monitoring the response to surgery in patients with gastric or other cancers one month after surgery. PMID- 12113580 TI - Serum chromogranin-alpha immunoradiometric assay in the diagnosis of pheochromocytoma. AB - BACKGROUND: The diagnosis of pheochromocytoma is based on laboratory tests that demonstrate an increase in urinary excretion of catecholamines or their metabolites. Chromogranin A (CgA) is a member of the granin family and is widely distributed in neuroendocrine cells and particularly in chromaffin adrenal cells. Consequently, serum CgA increases in patients affected by pheochromocytoma and other diseases of the chromaffin system. AIM: This study investigated the performance of serum CgA assay in the diagnosis of pheochromocytoma and compared serum CgA with 24-hour urinary epinephrine (E), norepinephrine (NE), vanillylmandelic acid (VMA) and metanephrines (MNs). METHODS: We enrolled 15 patients with histologically proven pheochromocytoma; 100 healthy blood donors and 148 patients with essential hypertension were enrolled as controls. Serum CgA was assayed by a specific immunoradiometric method (IRMA). Urinary tests were done with high performance liquid chromatography (HPLC). RESULTS: Circulating CgA showed a higher sensitivity (1.00), specificity (0.96) and accuracy (0.96) than all other tests. Serum levels of CgA clearly increased from blood donors and patients with essential hypertension to patients with pheochromocytoma (p<0.0001). Furthermore, a strong relationship between serum CgA and tumor mass was found (p<0.0001). In conclusion, our data suggest that the CgA assay might be used as a single test for the diagnosis of pheochromocytoma. PMID- 12113583 TI - Monitoring anti-p53 antibodies in patients with laryngeal squamous cell carcinoma: response to surgical treatment. PMID- 12113582 TI - Interleukin-6 is increased in breath condensate of patients with non-small cell lung cancer. AB - Despite recent advances in the diagnosis and treatment of non-small cell lung cancer (NSCLC), most patients still present with advanced stage disease at the time of diagnosis. Recent studies suggest that IL-6 is involved in the development of lung cancer. The aim of the present study was to investigate whether the measurement of IL-6 levels in the breath condensate of NSCLC patients could be used to bring forward the moment of diagnosis and to monitor the progression of the disease. Twenty patients with histological evidence of NSCLC (14 men and 6 women, age 63+/-8 years) and 15 healthy controls (8 men and 7 women, age 45+/-6 years) were enrolled in the study. IL6 was measured in the exhaled breath condensate of patients and controls by means of a specific enzyme immunoassay kit. Higher concentrations of exhaled IL-6 were found in NSCLC patients (9.6+/-0.3 pg/mL) than in controls (3.5+/-0.2 pg/mL). A statistically significant difference was observed between patients with NSCLC at different stages: higher concentrations of IL-6 (10.9+/-0.5 pg/mL) were found in patients with metastatic disease than in those with stage III (9.7+/-0.4 pg/mL), stage II (8.9+/-0.3 pg/mL) and stage I disease (7.9+/-0.3 pg/mL). These findings suggest that the measurement of IL-6 in the breath condensate of patients with NSCLC could be proposed as a parameter to take into account in early diagnosis and disease monitoring. PMID- 12113584 TI - Bravais-Pearson and Spearman correlation coefficients: meaning, test of hypothesis and confidence interval. PMID- 12113585 TI - Analysis of PSA velocity in 1666 healthy subjects undergoing total PSA determination at two consecutive screening rounds. AB - The study purpose was to assess PSA velocity (PSAV) in healthy subjects in order to establish a reliable cutoff for the differential diagnosis of prostate cancer in a screening setting. We studied a series of 1666 healthy men aged 55 to 74 years undergoing two total PSA determinations at a four-year interval within a population-based randomized screening trial at the Centro per lo Studio e la Prevenzione Oncologica of Florence. First and second screening round PSA assays (PSA1 and PSA2) were carried out with the same method and by the same laboratory. PSAV (PSA1-PSA2/year) was determined in non-cancer subjects in the overall series or in specific age and PSA subgroups, and in subjects with cancer detected at the second screening round. Average PSAV in 1648 non-cancer subjects was 0.07 ng/mL/year (range -2.18+5.99, 95% CI 0.05-0.09); at least one third of subjects showed a decrease in PSA (negative PSAV), mostly of limited magnitude and in the low PSA range. Average PSAV in the 18 cancer patients was 1.16 ng/mL/year (range 0.10-5.6, 95% CI 0.56-1.77), which is significantly higher (p<0.01) than in non cancer subjects. None of the cancer patients showed a PSA decrease over time. Whatever cutoff was taken for PSAV, its power to discriminate cancer was limited: in particular the previously used PSAV cutoff of 0.75 ng/mL/year would have included only 42 of the 1648 non-cancer subjects (specificity 97.5%) but excluded eight of the 18 cancer patients (sensitivity 55.5%). At best, with the adopted screening protocol PSAV (cutoff 0.10 ng/mL/year) could have spared 27.9% of non cancer subjects with PSA > or =2.5 ng/mL further diagnostic assessment and 22.7% of non-cancer subjects with PSA > or =4 ng/mL random sextant biopsy, while missing no cancers. This study provides a reliable estimate of PSAV based on a large unbiased population sample. PSAV is widely variable over time, particularly at low PSA values. PSAV might be of value as an indicator for diagnostic assessment and random sextant biopsy in a screening setting. PMID- 12113586 TI - Effect of antibiotic treatment on serum PSA and percent free PSA levels in patients with biochemical criteria for prostate biopsy and previous lower urinary tract infections. AB - BACKGROUND: Controversy exists as to the influence of inflammatory foci on total and free prostate-specific antigen (PSA) concentrations. The objective was to analyze the biological variations of PSA and percent free PSA (%f-PSA) in patients with biochemical criteria for prostate biopsy (PSA higher than 4 ng/mL and normal rectal examination) and compare them with the variation induced by antibiotic treatment in a cohort of patients with a history of lower urinary tract infections and no clinical evidence of prostatitis. METHODS: Ninety patients with a history of lower urinary tract infections, non-suspicious digital rectal examination and PSA between 4 and 20 ng/mL were analyzed. PSA concentration and %f-PSA were determined. Forty-five patients were treated with three weeks of ofloxacin, following which marker determination was repeated. All patients underwent ultrasound-controlled transrectal six-core prostate biopsy. RESULTS: Sixty-seven patients presented benign prostatic hyperplasia (BPH) (30 with prostatitic foci) and 23 cancer. Significant variations in PSA (6.97 ng/mL vs. 5.82 ng/mL, p=0.001) and %f-PSA (14.84% vs. 17.53%, p=0.01) were found only in the treated patients. These differences were significant for patients with BPH associated prostatitic foci and not for patients with BPH or cancer. The tendency was for PSA to decrease (15 treated patients with PSA <4 ng/mL vs. six non treated patients) and for %f-PSA to increase. The median variation of %f-PSA was greater than that of PSA. When the cutoff for %f-PSA was set at 25%, 18.9% of unnecessary biopsies after the first determination and 20% after the second could be avoided. By associating the reduction in PSA, up to 46% could be avoided in treated patients. CONCLUSION: Biochemical criteria for prostate biopsy may be modified in patients with a history of lower urinary tract infections due to variations greater than those explained by intraindividual biological variations, and may be influenced by the antibiotic treatment. These results suggest that subclinical inflammatory foci may influence PSA and %f-PSA. PMID- 12113587 TI - Insulin-like growth factor II (IGF-II) immunohistochemistry in breast cancer: relationship with the most important morphological and biochemical prognostic parameters. AB - Recent in situ hybridization experiments have shown a high content of IGF-II mRNA in breast cancer stroma. The aim of this study was to examine the relationship between IGF-II protein expression and several prognostic parameters in 75 infiltrating ductal carcinomas (IDC) of the breast. Tissue sections were evaluated for proliferative activity, IGF-II protein, ER, PgR, p53, and p21 expression using immunohistochemical procedures. The degree of stromal proliferation was assessed. Menopausal status, axillary lymph node involvement and nuclear grade were known. Thirty-five patients (44.3%) were premenopausal and 47 (62.6%) had lymph node metastases. Marked stromal proliferation was found in 34 (45.3%) specimens and high nuclear grade in 20 (26.5%). Eighteen tumors (24%) showed no IGF-II immunostaining. In the positive cases, IGF-II was detected both in the tumor stroma and in the cytoplasm of epithelial cancer cells: a high IGF II content was found in 12 specimens (16.0%), a low content in 14 (18.7%) and a moderate content in 31 (41.3%). Twenty-four tumors (32.0%) showed high proliferative activity. Both ER and PgR were expressed in the nucleus of cancer cells: 49 tumors (65.3%) were ER positive (ER+) and 34 (45.3%) PgR positive (PgR+). p21 protein was detected in 37 tumors (49.6%) and p53 in 12 (16%). IGF-II protein was not correlated with menopausal status, lymph node metastases, nuclear grade, proliferative activity, ER or p53. In contrast, IGF-II correlated strongly with stromal proliferation (p=0.008), PgR (p=0.03) and p21 (p=0.01). This study demonstrates that in IDC of the breast IGF-II protein is expressed in the epithelium and stroma of the majority of tumors and is correlated with stromal amount, PgR and p21 expression. These preliminary results indicate that IGF-II expression in breast cancer is connected with two important regulators of breast cancer growth and differentiation. PMID- 12113588 TI - Prognostic value of plasminogen activator inhibitors in breast cancer. AB - The prognosis of cancer is primarily dependent on its potential to invade and metastasize. Data from both preclinical and clinical studies strongly suggest that serine proteases, as well as their inhibitors and receptor, play a central role in the processes leading to metastasis. We therefore investigated the prognostic value of plasminogen activator inhibitors type 1 (PAI-1) and type 2 (PAI-2) and the combination of both inhibitors in 332 patients with operable breast cancer. PAI-1 and PAI-2 content was measured in the primary tumor cytosols using an enzyme-linked immunosorbent assay. For PAI-1 the median value (3.9 ng/mg protein) was used as cutoff, while the optimized cutoff for PAI-2 (6.5 ng/mg protein) was obtained using the log-rank statistic. After a median follow-up of 46 months 96 (29%) patients relapsed. In univariate analysis patients with a high PAI-1 or a low PAI-2 content had an increased risk of relapse. The difference was statistically significant for PAI-1 (p<0.0001) and almost statistically significant for PAI-2 (p=0.057). Stage, tumor size, differentiation grade, lymph node status and hormone receptor status also showed significant univariate impact on disease-free survival (DFS). In multivariate analysis (Cox model) PAI-1 (p<0.0001, RR=2.78), PAI-2 (p=0.0075, RR=2.17), UICC stage (p=0.0014, RR=2.2), differentiation grade (p=0.0097, RR=1.91) and nodal status (p<0.0001, RR=2.9) retained their significance. When both inhibitors were combined the worst prognosis was observed in patients with simultaneous high PAI-1 and low PAI-2 levels, whereas low PAI-1 in combination with high PAI-2 values indicated a very favorable prognosis. In conclusion, our study showed that both PAI-1 and PAI-2 had independent prognostic value in breast cancer. Combination of both inhibitors further improved the differentiation of patients with respect to prognosis. PMID- 12113589 TI - Diuretics and dopamine for the prevention and treatment of acute renal failure: a critical reappraisal. AB - The efficacy of diuretics and dopamine in preventing and treating acute renal failure (ARF) is still debated, although these drugs are widely used in clinical practice. The present review addresses this debate providing an update on available data. There are very few well controlled and sufficiently large trials testing these agents in different ARF clinical settings, with either a preventive or a curative goal. Unfortunately, most findings rule out any favourable effects of diuretics, mannitol, atrial natriuretic peptides, and dopamine. In fact, at least in some clinical conditions, they can even worsen the risk of ARF or aggravate its course. PMID- 12113590 TI - Health related quality of life (HRQOL) in the elderly on renal replacement therapy. AB - Various opinions have been presented about the influence of age on the health related quality of life (HRQOL) of patients on renal replacement therapy (RRT). Some authors sustain that age worsens HRQOL, while others show the opposite. It has been seen that a psychological adjustment occurs with aging, even when chronic illness is also present. In addition, elderly patients appear to adapt better to RRT than younger ones. PMID- 12113591 TI - Salt intake and kidney disease. AB - We have reviewed the role of salt intake in kidney diseases, particularly in relation to renal hemodynamics, renal excretion of proteins, renal morphological changes and progression of chronic renal failure. High salt intake may have detrimental effects on glomerular hemodynamics, inducing hyperfiltration and increasing the filtration fraction and glomerular pressure. This may be particularly important in elderly, obese, diabetic or black patients, who have a high prevalence of salt-sensitivity. Changes in salt intake may influence urinary excretion of proteins in patients with essential hypertension, or diabetic and non diabetic nephropathies. Moreover, high sodium intake may blunt the antiproteinuric effect of various drugs, including angiotensin-converting-enzyme inhibitors and calcium antagonists. Experimental studies show a direct tissue effect of salt on the kidney, independent of its ability to increase blood pressure, inducing hypertrophy, fibrosis and a decrease in glomerular basement membrane anionic sites. However, no firm conclusion can be drawn about the relationship between salt consumption and progression of chronic renal failure, because most information comes from conflicting, small, retrospective, observational studies. In conclusion, it would appear that restriction of sodium intake is an important preventive and therapeutic measure in patients with chronic renal diseases of various origin, or at risk of renal damage, such as hypertensive or diabetic patients. PMID- 12113593 TI - Scleroderma at end stage renal disease in the United States: patient characteristics and survival. AB - BACKGROUND: The patient characteristics and mortality associated with scleroderma have not been characterized for a national sample of end stage renal disease (ESRD) patients. METHODS: 364,317 patients in the United States Renal Data System initiated on ESRD therapy between 1 January 1992 and 30 June 1997 with valid causes of ESRD were analyzed in an historical cohort study of scleroderma. RESULTS: Of the study population, 820 (0.22%) had scleroderma. The mean age of patients with scleroderma was 56.38 +/- 13.93 years vs. 60.48 +/- 16.51 years for patients with other causes of ESRD (p<0.01 by Student's t-test). In histogram analysis, there were two age peaks: 45-49 and 65-69. In logistic regression, patients with scleroderma, compared to patients with other causes of ESRD, were significantly more likely to be women, Caucasian, younger, and more likely to have congestive heart failure but less likely to have ischemic heart disease, stroke, and receive predialysis erythropoietin. The unadjusted two-year survival of patients with scleroderma during the study period was 49.3% vs. 63.8% in all other patients (adjusted hazard ratio, 1.96, 95% CI 1.70-2.26, p=0.0001 by Cox Regression). CONCLUSIONS: Among patients with ESRD, the demographics of patients with scleroderma were similar to those of patients with scleroderma in the general population. Patients with scleroderma had decreased survival compared to patients with other causes of ESRD, despite being equally likely to be wait listed and receive renal transplantation adjusted for other factors. PMID- 12113592 TI - Aldosterone and PAI-1: implications for renal injury. AB - Aldosterone modulates cardiovascular and renal injury and fibrosis in animal models. This review explores the hypothesis that aldosterone causes injury and fibrosis, in part, through effects on plasminogen activator inhibitor-1, the major physiological inhibitor of plasminogen activators in vivo. Aldosterone interacts with angiotensin II to increase plasminogen activator inhibitor-1 expression in vitro and in vivo. Plasminogen activator inhibitor-1, by inhibiting the production of plasmin from plasminogen, tips the balance in favor of extracellular matrix accumulation and promotes fibrosis. Aldosterone receptor antagonism decreases both PAI-1 expression and fibrosis in animal models. These findings have implications for the clinical management of cardiovascular and renal disease. PMID- 12113594 TI - Risk factors for pulmonary embolism in chronic dialysis patients. AB - BACKGROUND: Risk factors for pulmonary embolism (PE) in end stage renal disease (ESRD) patients have not been studied in a large population. METHODS: 375,152 patients in the United States Renal Data System initiated on dialysis between 1 January 1992 and 30 June 1997 were analyzed in an historical cohort study of hospitalized PE (ICD9 Code 415.1x) occurring prior to receipt of renal transplant. Cox regression models were used to analyze risk factors for PE in dialysis. Dialysis modality was analyzed in an intention to treat fashion, thus patients who changed modalities later were considered to have remained on the same modality. RESULTS: The incidence of pulmonary embolism did not increase over time. Independent risk factors for hospitalizations for PE were similar to those in the general population (older age, females, systemic lupus erythematosus, lower risk for Asians) with the addition of peritoneal dialysis (vs. hemodialysis, adjusted odds ratio 1.56, 95% CI 1.15-2.13), polycystic kidney disease, and congestive heart failure. Notably, in Cox regression analysis, no relation was seen with baseline laboratory results (hematocrit, serum albumin, serum creatinine) or comorbidity (except congestive heart failure) and PE risk. Dialysis patients with PE had increased mortality (hazard ratio 1.20, 95% confidence interval 1.08-1.33). CONCLUSIONS: The incidence of PE did not increase significantly in ESRD patients from 1992-1997. PE were associated with increased mortality. Peritoneal dialysis patients may have higher risk of PE than hemodialysis patients, and other high-risk groups were identified. PMID- 12113595 TI - Hospitalizations for bacterial septicemia in patients with end stage renal disease due to diabetes on the renal transplant waiting list. AB - The incidence and risk factors for hospitalizations for bacterial septicemia, a serious cause of morbidity and mortality in end stage renal disease (ESRD), have been studied separately for patients on chronic dialysis and after renal transplantation, but have not been compared directly. Using data from the USRDS, we studied 11,369 patients with ESRD due to diabetes enrolled on the renal and renal-pancreas transplant waiting list from 1 July 1994-30 June 1997. Cox non proportional hazards regression models were used to calculate adjusted, time dependent hazard ratios (HR) for time to hospitalization for bacterial septicemia (ICD9 Code 038.x). In Cox Regression analysis, renal transplantation was independently associated with a shorter time to bacterial septicemia (HR 1.22, 95% confidence interval, 1.05-1.40). In addition, renal transplantation was associated with a higher rate of sepsis due to gram-negative organisms (HR 3.32, 95% CI 2.614.23) and urinary tract infection (10.43, 95% CI 6.72-16.17) compared with patients still on the renal transplant waiting list. The relative risk of sepsis increased with time after renal transplantation. Renal transplantation was associated with a significantly higher risk and different spectrum of bacterial septicemia than maintenance dialysis, and the risk of sepsis did not decrease over time. PMID- 12113596 TI - Hospitalizations for bacterial pneumonia after renal transplantation in the United States. AB - PURPOSE: Bacterial pneumonia has been cited as the leading cause of infectious death in renal transplant recipients but has not been studied in a national transplant population. SUBJECT AND METHODS: Retrospective analysis of the incidence, risk factors and mortality of hospitalized bacterial pneumonia (ICD9 Code 481.x486.x) for 33,479 renal transplant recipients in the United States Renal Data System transplanted from 1 July 1994-30 June 1997. RESULTS: Among all transplant recipients, 4.7% were hospitalized for a primary discharge diagnosis of pneumonia in the study period (2.86 episodes per 100 person years). 9.9% had bronchoscopy and 4.8% had open lung biopsy. A specific etiology was not identified in 72.5% of patients. The hospitalization rate for pneumonia and hazard for mortality due to hospitalized pneumonia were both constant over time. In logistic regression analysis, pneumonia prior to transplant (odds ratio 1.73, 95% confidence interval, 1.32-2.26), older recipient age, diabetes, delayed graft function, rejection (occurring at any time after transplant during the time of the study), duration of pre-transplant dialysis, and positive recipient cytomegalovirus serology were associated with pneumonia. In Cox Regression, hospitalization for pneumonia was associated with greater risk of mortality (hazard ratio 1.64, 95% CI, 1.42-1.89). CONCLUSIONS: Renal transplant recipients with a previous history of pneumonia are at increased risk for subsequent pneumonia, which is associated with substantially decreased patient survival. Given the low rate of specific etiologies identified in this study, invasive diagnosis may be underutilized in this population. PMID- 12113597 TI - Assessment of the efficiency of treatment of dyslipidaemia in renal outpatients. AB - BACKGROUND: Lipid abnormalities, together with other co-existent risk factors, contribute to the accelerated atherosclerotic process, and consequently to the high incidence of cardiovascular disease, observed in end-stage renal patients. The objectives of this study were to determine the prevalence of dyslipidaemia and to assess how it is managed in a range of patients at four UK renal units. METHODS: Patients with renal disease were recruited from the outpatient clinics of 4 UK hospitals. Individuals meeting the entry criteria were required to provide one sample of venous blood following a 12-hour fast. Lipid profiles were measured for each patient. RESULTS: The study population consisted of 677 patients of which 276 (40.8%) were pre-dialysis patients with existing renal disease, 233 patients (34.4%) were receiving haemodialysis and 168 patients (24.8%) were receiving CAPD. Analysis showed that 64% of all patients had hypercholesterolaemia (defined according to Joint British Societies guidelines as LDL-cholesterol [LDL-C] >3.0 mmol/L [115 mg/dL] and/or total cholesterol [TC] >5.0 mmol/L [190 mg/dLl). Despite the high incidence of hypercholesterolaemia, only 16% of study participants were receiving lipid-lowering therapy. An LDL-C goal of <3.0 mmol/L (115 mg/dL) was achieved in 50.0% of patients receiving lipid lowering treatment. CONCLUSIONS: The widespread failure to treat hypercholesterolaemia in patients with renal dysfunction given their high risk of future cardiovascular events is a major cause for concern. The observation that many dialysis patients receiving lipid-lowering therapy had not achieved recommended LDL-C and TC levels suggests that more efficient treatment of dyslipidaemia may be indicated in this patient population. PMID- 12113598 TI - Stopping smoking slows accelerated progression of renal failure in primary renal disease. AB - BACKGROUND: Cigarette smoking accelerates the progression of renal failure in primary kidney diseases. It is not known, however, whether stopping smoking slows this accelerated loss of renal function. METHODS: 45 patients with progressive primary nephropathies (glomerulonephritis or tubulointerstitial nephritis) and moderate renal failure were encouraged to stop heavy cigarette smoking (1-2 packs per day); 26 patients refused to change their smoking habits (current smokers), and 16 successfully stopped (ex-smokers) during the 24 month study period. Carboxyhemoglobin and creatinine clearance were measured every six months. The primary end-point of the study was end-stage renal disease requiring renal replacement therapy. RESULTS: There were no significant differences between the two patient groups in demographic, renal and treatment characteristics at the start of the study. Current and ex-smokers had similar rates of decline of creatinine clearance during the 24 months before the investigations. Compared to ex-smokers or matched non-smoking renal patients, permanent smokers had a significantly faster decline in creatinine clearance during the two-year study period. Renal replacement therapy had to be started in six smokers, but only in one ex-smoker and none of the non-smokers during the study period. CONCLUSIONS: Giving up smoking slowed the rapid progression of renal failure, but did not reverse the loss of renal function induced by smoking. We recommend that increased efforts should be made to encourage renal patients to give up smoking in order to prolong dialysis-free kidney survival. PMID- 12113599 TI - Ionised and total serum magnesium in renal transplant patients. AB - BACKGROUND: Hypomagnesemia in renal transplant patients is almost always documented through total serum values (MgT), but it has recently become user friendly to assay the biologically active, ionised fraction (Mg++). We verified the prevalence of true ionised magnesemia and the correspondence between total and ionised Mg assays in our transplanted patients, taking into account renal Mg excretion and the possible role of other reputed factors of hypomagnesemia (cyclosporine, secondary hyperparathyroidism and acid-base balance). METHODS: Thirty-eight transplanted patients (25M/13F, aged 41 +/- 11 years) and 38 age and sex matched controls were enrolled. Blood chemistries included: ionised Mg and Ca, total Mg and Ca, phosphate, creatinine, albumin, bicarbonate, alkaline phosphatase, parathyroid hormone and, in patients, cyclosporine (CyA). A 24-h urine collection (for Ca and Mg) and a fasting spot sample (for pH, Mg, Ca, phosphate, creatinine) were also obtained. RESULTS: Patients with mild renal failure (creatinine: Cr=1.75 +/- 0.83 mg/dL), mild persistent secondary hyperparathyroidism and almost normal tubular acidification capacity had MgT lower than controls (0.76 +/- 0.08 vs 0.82 +/- 0.08 mmol/L; p<0.002), with 10 cases (26%) of total hypomagnesemia. Mg++ was also significantly low (0.51 +/- 0.08 vs 0.53 +/- 0.05 mmol/L; p<0.03), but there were only four cases (10%) of true ionised hypomagnesemia. MgT and Mg++, although correlated (with a low r value: =0.49; p<0.001), showed poor correspondence in individual patients and MgT was not useful to identify cases of true ionised hypomagnesemia. Neither assay correlated with renal function. Daily urinary excretion of Mg was normal (3.5 +/- 1.3 vs 3.0 +/- 0.24 mmol/day; n.s.), with no case of definite hypomagnesuria. Fasting excretion fraction (EF) of Mg, calculated with both assays, was increased in approximately 60% of patients (EF(MgT) 4.9 +/- 2.6 vs 2.32 +/- 0.7%; p<0.0001; EF(Mg++) 7.74 +/- 4.9 vs 3.63 +/- 1.18%; p<0.0001) and positively correlated with serum Cr (r=0.62; p<0.0001 with EF(MgT); and r=0.467; p<0.005 with EF(Mg++) but not with CyA. Neither Mg assay correlated with serum CyA, calcium, phosphate, PTH or bicarbonate. CONCLUSIONS: In long term renal transplant patients not taking diuretics, the prevalence of true ionised hypomagnesemia is low. Renal insufficiency, typically associated with Mg retention, is the major cause of increased EF(Mg) and, as such, plays an antagonistic role to CyA and other factors of renal Mg wasting. Because MgT and Mg++ are not closely related, assay of the ionised fraction seems advisable in case of total hypomagnesemia. However, because diagnosis of depletion can hardly rely on serum assay alone, a fuller evaluation (urinary excretion and other clinical and biochemical signs of hypomagnesemia) is suggested before diagnosis is made. PMID- 12113600 TI - Crescentic glomerulonephritis in Egypt: clinical and histopathological risk factors. AB - We followed up 128 patients with crescentic glomerulonephritis (CGN), having sufficient clinical and histopathological data for a mean period of 34 +/- 28 months. There were 49 males and 79 females, mean age 22.7 +/- 14 years. We studied the effects of clinical, laboratory and histopathological parameters on kidney function and survival at the end point of the study. Multivariate analysis indicated that serum creatinine at presentation, nephrotic range proteinuria during the follow-up period, percentage of glomeruli with crescents, percentage of fibrous crescents and absence of cellular infiltration were significant risk factors affecting kidney function at the end of the study. The only risk factor significantly correlated with mortality was serum creatinine at last follow-up. PMID- 12113601 TI - Urinary glycosaminoglycan and proteoglycan excretion in normoalbuminuric patients with type 1 diabetes mellitus. AB - BACKGROUND: Diabetic nephropathy may be related to an abnormal metabolism of glycosaminoglycans (GAG) in the glomerular basement membrane (GBM). The first manifestation of nephropathy is microalbuminuria, whose appearance indicates a loss of GBM selectivity. The present study evaluated whether GAG excretion becomes abnormal in parallel with microalbuminuria, and whether such abnormalities are also present in normoalbuminuric diabetic patients. METHODS: We measured urinary GAG excretion in 60 patients with type 1 (insulin-dependent) diabetes mellitus and in 22 healthy subjects. GAG were isolated from 24-h urine using ion-exchange chromatography on DEAE Sephacel. GAG composition was determined by cellulose acetate electrophoresis and expressed as percentages by densitometric scanning of Alcian Blue stained strips. RESULTS: On subgrouping for albuminuric status and glyco-metabolic control, we found high urinary GAG concentrations in all except the normoalbuminuric patients with good glyco metabolic control. The urinary GAG electrophoretic pattern showed alterations in chondroitin sulphate (CS) and heparan sulphate (HS) relative contents. A higher frequency of low sulphated chondroitin sulphate-proteoglycan (LSC-PG) was observed in all patients, including those with normoalbuminuria and good glyco metabolic control. CONCLUSIONS: This urinary pattern may be indicative of an abnormal GBM metabolism. Since GAG play an important role in GBM permeability, these anomalies might consequently represent a first step towards selective changes of GBM in type 1 diabetes mellitus. PMID- 12113602 TI - Differential diagnosis of bacterial infection and inflammatory response in kidney diseases using procalcitonin. AB - BACKGROUND: Early diagnosis of bacterial infection in renal patients remains difficult. Common laboratory parameters, such as white blood cell (WBC) count, erythrocyte sedimentation rate, and C-reactive protein (CRP) may be affected by the underlying disease, uremia or its extracorporeal treatment, or by immunosuppressive drugs. Procalcitonin (PCT) may be useful for the detection of systemic bacterial infections in patients with end-stage renal disease (ESRD) undergoing renal replacement therapy, but elevated PCT concentrations have also been found in a significant number of uremic patients without signs of infection. METHODS: We tested whether measurements of PCT levels help distinguish the chronic inflammation in renal diseases from invasive bacterial infections. Serum levels of PCT were compared with the corresponding serum C-reactive protein (CRP) concentrations and WBC counts in 197 patients with different stages of renal disease: Group I) 32 patients with chronic renal failure (serum creatinine 2-6 mg/dL); group II) 31 patients with a functioning renal transplant receiving standard immunosuppressive regimens; group III) 76 clinically stable patients with ESRD undergoing hemodialysis (HD); group IV) 23 patients with chronic renal failure (CRF) due to systemic autoimmune disease; group V) 35 patients with proven systemic bacterial infection and CRF. RESULTS: PCT levels were within the normal range (< 0.5 ng/mL) in patients with CRF and renal transplant patients without any clinical evidence of bacterial infection, regardless of the degree of renal failure and the underlying disorders. In 22 out of 76 stable HD patients, PCT levels were above the upper limit of normal, but 97% of these values were below the proposed cut-off for chronic HD patients of < 1.5 ng/mL. CRP levels were elevated in 17 of 32 patients with CRF (mean +/- SD: 0.57 +/- 0.49 mg/dL), in 10 of 31 renal transplant patients (0.41 +/- 0.55 mg/dL), in 16 of 23 patients with autoimmune disorders (2.78 +/- 3.21 mg/dL) and in 42 of 76 patients treated by HD (0.64 +/- 0.58 mg/dL). In patients with CRF and systemic bacterial infections, both PCT and CRP were markedly elevated (PCT 61.50 +/- 115.4 ng/mL, CRP 14.50 +/- 10.36 mg/dL), but in contrast to PCT, CRP values overlapped in infected and non-infected patients. CONCLUSIONS: Our data indicate that PCT levels are not significantly affected by loss of renal function, immunosuppressive agents or autoimmune disorders. Thus, significantly elevated PCT concentrations offer good sensitivity and specificity for the early diagnosis of systemic bacterial infection in patients with CRF or patients with ESRD treated by HD. CRP concentrations may be useful indicators for inflammation in patients with renal diseases, but have low specificity for the diagnosis of bacterial infection. PMID- 12113603 TI - Oxidative stress markers in hepatitis C infected hemodialysis patients. AB - BACKGROUND: Oxidative stress in hemodialysis (HD) patients or hepatitis C virus (HCV) infections may be related to increased production of free radicals. We assessed the effect of HCV infection on the oxidative stress markers, malondialdehyde (as TBARS), protein carbonyl content and protein sulfhydryl groups, in chronic HD patients. METHODS: Twenty HCV infected patients (9 men and 11 women, age 44.8 +/- 14.3 years) and 10 non-HCV infected patients (6 men and 4 women, age 55.6 +/- 14.3 years) receiving regular HD were recruited. The average hemodialysis duration was 30 +/- 8 months for HCV (+) patients and 14 +/- 8 months for HCV (-) patients. Controls consisted of healthy subjects. RESULTS: Serum TBARS and carbonyl content were significantly elevated in HCV(-) patients (p<0.001, p<0.05) and in HCV(+) patients (p<0.001, p<0.001) vs. Controls. There was also a significant difference in serum TBARS and carbonyl content between HCV(-) patients and HCV(+) patients (p<0.001, p<0.05). Serum protein sulfhydryl groups in HCV(-) and HCV(+) patients were the same, but significantly lower than in Controls (p<0.001, p<0.001). When HCV(+) patients were divided into two sub groups, one with shorter (13.4 +/- 8 months; n=7) and the other with longer (40.3 +/- 8 months; n=13) duration of HD treatment, no differences were found between subgroups. CONCLUSION: HCV infection may aggravate oxidative stress in HD patients. PMID- 12113604 TI - BK polyomavirus interstitial nephritis in a renal transplant patient with no previous acute rejection episodes. AB - A renal transplant patient treated with tacrolimus and mycophenolate-mofetil (MMF) developed progressive graft function deterioration 10 months after transplantation. Biopsy of the graft showed severe, focally accentuated interstitial inflammation with focal tubulitis and tubular necrosis, and medium severe interstitial fibrosis with focal tubular atrophy. Glomerular and vascular structures were preserved. On careful examination, in some sections, tubular epithelial cells showed a definite increase with deformation of the nuclear shape, chromatin irregularities with peripheral dislocation and inclusion bodies. These cytopathic changes suggested polyoma virus infection ("decoy cells"). Subsequent screening of the urinary sediment confirmed the presence of many "decoy cells". Immunohistochemical analysis of the biopsy showed many tubular cells were strongly positive for the SV 40 antigen, specific for BK polyoma virus. A diagnosis of interstitial nephritis due to BK polyoma virus was made, though the coexistence of cellular rejection could not be excluded. At variance with previous reports, our patient had not had repeated episodes of rejection before biopsy or heavy immunosuppressive treatment, such as ALG, OKT3, after transplantation. This case shows that even in the absence of vigorous anti rejection therapy an immunosuppressive regimen based on tacrolimus and MMF may involve the risk of BK polyoma virus- associated interstitial nephritis. PMID- 12113605 TI - Statin specific toxicity in organ transplant recipients: case report and review of the literature. AB - Statins are widely prescribed to organ transplant recipients with hyperlipidemia. We report the case of a cardiac transplant recipient who developed severe rhabdomyolysis and acute renal failure after being switched from pravastatin to simvastatin. The patient's other medications included cyclosporin A and diltiazem. Unlike pravastatin, the metabolism and tissue concentrations of simvastatin--and of other statins - can be greatly affected by these drugs. The propensity of the individual statins to interact with drugs commonly prescribed to transplant recipients is reviewed. PMID- 12113606 TI - Hereditary nephritis with macrothrombocytopenia: phenotypic variety and the genotypic defect. AB - A number of patients present to nephrologists with end-stage renal failure of unknown cause and many have small kidneys, making renal biopsy inadvisable. A small number may have clues to the diagnosis of hereditary nephritis, as in the patient we present here. The propositus was a 22-year-old man, who was admitted to our nephrology ward because of recently discovered renal insufficiency. Ultrasound examination revealed bilateral small kidneys. He was severely hypertensive; audiometry and ophthalmic examinations were normal. Thrombocytopenia with giant platelets was observed in peripheral blood smears. Basophilic cytoplasmic inclusions (Dohle-like bodies) were present in neutrophil and basophilic granulocytes. A family history of nephropathy associated with macrothrombocytopenia was obtained. Epstein syndrome was diagnosed, a rare autosomal dominant disorder. He started hemodialysis and subsequently received a living donor kidney transplant (from his mother). Molecular genetics has considerably clarified the field of hereditary nephritis associated with macrothrombocytopenia by demonstrating that these syndromes involve a similar molecular defect. It was first shown that these syndromes were linked to the same locus on chromosome 22q. Then the gene involved--encoding non-muscle myosin heavy chain 9 (MYH9)--was identified. This entity ("MYH9 disease") must be clearly differentiated from Alport syndrome (type IV collagen disease). In conclusion, this case serves to remind us: 1) that in patients presenting late to nephrologists with bilateral small kidneys, the diagnosis can still be made in some instances on the basis of other clinical signs known to be associated with recognized inherited syndromes; 2) that there are various types of inherited thrombocytopenia associated with nephritis; screening for nephritis is mandatory in all of them. PMID- 12113607 TI - Painful cutaneous lesions, renal failure and urgent parathyroidectomy. AB - We describe two patients with end stage renal failure who presented with painful skin lesions, which rapidly progressed to become necrotic and gangrenous. The diagnosis was calciphylaxis, a rare disorder due to calcification and luminal fibrosis of small and medium sized cutaneous and systemic vessels. Both patients had tertiary hyperparathyroidism. An urgent parathyroidectomy was performed on one patient, which relieved her symptoms; the other required local surgery but refused parathyroidectomy and died. PMID- 12113608 TI - Laudanosine, an atracurium and cisatracurium metabolite. AB - Laudanosine is a metabolite of the neuromuscular-blocking drugs atracurium and cisatracurium with potentially toxic systemic effects. It crosses the blood-brain barrier and may cause excitement and seizure activity. Its interest in recent years has increased because of the recognized interaction with gamma-aminobutyric acid, opioid and nicotinic acetylcholine receptors. It has been shown to produce analgesia in animals. In the cardiovascular system, high plasma concentrations produce hypotension and bradycardia. In hepatic failure, its elimination half life is prolonged but only moderate accumulation occurs in adults, whereas in infants and children plasma concentration are greater. In patients undergoing liver transplantation, laudanosine concentrations are increased during preanhepatic, anhepatic and postanhepatic stages. Patients with renal failure have higher plasma concentrations and a longer mean elimination half-life. In pregnancy, laudanosine crosses the placental barrier. The mean transplacental transfer is 14% of maternal blood concentrations. Except for prolonged administration of atracurium in intensive care units, laudanosine accumulation and related toxicity seem unlikely to be achieved in clinical practice. When cisatracurium is used, plasma concentrations of laudanosine are lower. Further studies are needed, especially around the interactions with gamma-aminobutyric acid, opioid and nicotinic acetylcholine receptors. PMID- 12113609 TI - Thiopental impairs neutrophil oxidative response by inhibition of intracellular signalling. AB - BACKGROUND AND OBJECTIVE: Thiopental in clinically relevant concentrations inhibits the oxidative function of neutrophils, whereas only very high, non therapeutic concentrations of methohexital induce a similar effect. The study characterized the molecular basis of this differential action of oxy- and thiobarbiturates on neutrophils. METHODS: Neutrophils were incubated in vitro with thiopental or methohexital using concentrations within the therapeutic range. Neutrophil responses were induced using different stimuli: N-formyl-L methionyl-L-leucyl-L-phenylalanine (FMLP), C5a and 1,2-dioctanoyl-sn-glycerol (DiC8-DAG). FMLP and C5a bind to specific G-protein-coupled receptors that share the same second messenger cascade. In contrast, DiC8-DAG, an activator of protein kinase C, bypasses the signal transduction pathway downstream of the receptors. Hydrogen peroxide production by neutrophils was assessed using flow cytometry. To characterize the localization of the interaction site, FMLP receptor expression and cytosolic-free calcium were further analysed. RESULTS: FMLP and C5a-induced hydrogen peroxide production were both significantly impaired by thiopental, but not by methohexital. When postreceptor signalling was bypassed, by stimulation with DiC8-DAG, neither thiopental nor methohexital affected hydrogen peroxide production. Additionally, neither of the barbiturates impaired the cytosolic Ca2+ response. CONCLUSIONS: We conclude that neither protein kinase C nor the hydrogen peroxide-generating enzymes are affected by thiopental or methohexital. The unimpaired Ca2+ response suggests that the function of the receptors and G proteins were also unimpaired. Taken together, this indicates that the site of action of thiopental is in the cellular signalling upstream of protein kinase C. PMID- 12113610 TI - Flurbiprofen does not change the bispectral index and 95% spectral edge frequency during total intravenous anaesthesia with propofol and fentanyl. AB - BACKGROUND AND OBJECTIVE: Previous studies have shown that general anaesthetic agents modulate the production of hypothalamic prostaglandins (PG) D2 and E2, which are mediators of sleep and wakefulness respectively. Although flurbiprofen, a cyclo-oxygenase inhibitor, is used clinically as a non-steroidal anti inflammatory agent and postoperative analgesic, it reduces prostaglandin production. Thus, this agent may affect the depth of sedation during general anaesthesia. In this study, we examined if flurbiprofen affects the bispectral index, which correlates with sedation levels. METHODS: Fifteen patients who underwent elective surgery under total intravenous anaesthesia with propofol and fentanyl were studied. The sedation level was monitored using a bispectral index monitor. On attainment of stable haemodynamics and a bispectral index, patients were given flurbiprofen axetil 50 mg intravenously. A bispectral index and 95% spectral edge frequency were recorded before and 5, 10, 15, 20 and 30 min after flurbiprofen axetil intravenously. RESULTS: Bispectral indexes of 51.7 (95% CI: 47.3-56.8), 51.7 (47.1-56.3), 51.3 (46.3-56.3), 50.3 (45.8-54.2), 48.9 (43.6 54.1) and 50.3 (45.5-55.2) at 0, 5, 10, 15, 20, 30 min after flurbiprofen axetil intravenously were observed. There was no change in this or 95% spectral edge frequency. CONCLUSIONS: Clinical dose of flurbiprofen axetil does not alter the bispectral index and 95% spectral edge frequency under total intravenous anaesthesia with propofol and fentanyl. PMID- 12113611 TI - Acute normovolaemic haemodilution beyond a haematocrit of 25%: ratio of skeletal muscle tissue oxygen tension and cardiac index is not maintained in the healthy dog. AB - BACKGROUND AND OBJECTIVE: The study investigated the effect of acute normovolaemic haemodilution on haemodynamics, blood flow and oxygen transport variables with regard to skeletal muscle tissue oxygenation in a canine model. METHODS: Twenty foxhounds were anaesthetized, mechanically ventilated with 30% oxygen in air and underwent first-step normovolaemic haemodilution with Ringer's lactate solution to haematocrit (Hct) 30 and 25% and second-step acute normovolaemic haemodilution with 6% Hetastarch 70,000/0.5 to Hcts of 20, 15 and 10%. Catheters were inserted into femoral arteries and veins and into the pulmonary artery for measurements ofhaemodynamics, temperature, and sampling of arterial and mixed-venous blood. A flow probe was placed around the left femoral artery. Skeletal muscle tissue oxygen tension (tPO2) was measured in the gastrocnemius muscle using a stepwise driven polarographic needle probe creating histograms from 200 single tPO2 measurements. RESULTS: Until a Hct of 25% was reached, the heart rate, mean arterial pressure, global and muscular oxygen delivery and consumption remained constant, while the cardiac index and oxygen extraction ratio were significantly increased when compared with baseline. The median tPO2 was significantly decreased at Hcts 15 and 10%, despite increased cardiac index and regional blood flow. The ratio of tPO2 and cardiac index as a marker for efficiency of acute normovolaemic haemodilution started to decline beyond Hcts of 25% (change of slope). CONCLUSIONS: In acute normovolaemic haemodilution to the level of Hct of 25%, the ratio between tPO2 and cardiac index decreases in the healthy dog, indicating an uneconomic relation at the threshold of Hct of 25%. PMID- 12113612 TI - Manipulations in glycogen metabolism and the failure to influence infarct size in the ischaemic rabbit heart. AB - BACKGROUND AND OBJECTIVE: Myocardial ischaemic preconditioning is characterized by a reduction in the rate of glycolysis. Brief myocardial ischaemia also reduces the glycogen content of the heart. The first objective was to determine whether augmenting glucose oxidation by activation of the pyruvate dehydrogenase complex would prevent the infarct limitation of ischaemic preconditioning. The second part of the study evaluates whether glycogen depletion before ischaemia mimics the infarct-limiting effect of ischaemic preconditioning. METHODS: Dichloroacetate (300 + 150 mg kg(-1)), an activator of the pyruvate dehydrogenase complex, was administered intravenously in the anaesthetized open-chest rabbit. All animals underwent 45 min of regional ischaemia and 3 h of reperfusion. Ischaemic preconditiong was elicited by 5 min of coronary occlusion. Control rabbits, those with ischaemic preconditioning with no dichloroacetate, received a saline vehicle. An isolated perfused rabbit heart model was employed to test the second hypothesis. Hearts were depleted of glycogen by perfusing them with a substrate-free buffer. Infarction was assessed by triphenyl tetrazolium chloride and area at risk determined with fluorescent particles. RESULTS: (a) Pyruvate dehydrogenase complex activation experiments. Treatment with dichloroacetate alone did not alter infarct size (58 +/- 7% control vs. 60 +/- 5% dichloroacetate). Addition of dichloroacetate did not attenuate the infarct limiting effect of ischaemic preconditioning as evidenced by a similar reduction in infarct size in the ischaemic preconditioning group (22 +/- 5%) and in the ischaemic preconditioning + dichloroacetate group (27 +/- 7%). (b) Glycogen depletion experiments. Compared with control hearts with a normal glycogen content (4.84 +/- 0.15 mg g(-1) wet weight), glycogen depleted and ischaemic preconditioning hearts had reduced glycogen content before ischaemia (2.15 +/- 0.26, 1.62 +/- 0.17 mg g(-1) wet weight, respectively; P < 0.01). Glycogen depletion did not reduce infarct size: 25.0 +/- 4.5% cf. 27.9 +/- 3.4% in the control group. However, ischaemic preconditioning resulted in a significant reduction of infarct size (11.5 +/- 2.3% vs. 27.9 +/- 3.4% control; P < 0.01). CONCLUSIONS: Augmentation of oxidative glycolysis by dichloroacetate in in situ rabbit hearts does not alter the effect of ischaemic preconditioning, and glycogen depletion in the isolated rabbit heart does not influence infarct size after subsequent coronary occlusion. PMID- 12113613 TI - Effect of three different doses of urapidil on blood glucose concentrations in the streptozotocin diabetic rat. AB - BACKGROUND AND OBJECTIVE: Diabetes mellitus associated with hypertension often causes perioperative complications. The alpha1-adrenoceptor antagonist/5 hydroxytryptamine-1A receptor agonist urapidil is an approved drug used in hypertension and hypertensive emergencies. 5-Hydroxytryptamine-1A (5-HT1A) receptor agonists impair glucose metabolism. To evaluate a possible dose dependent hyperglycaemic effect of urapidil due to its 5-HT1A receptor agonistic properties, the effect of three doses of urapidil on hyperglycaemia in the streptozotocin diabetic rat was investigated. METHODS: Male Wistar-Kyoto rats were made diabetic by streptozotocin and randomly allocated to the following daily treatments for 7 days (n = 6 each): urapidil 6 mg kg(-1), urapidil 20 mg kg(-1), urapidil 60 mg kg(-1), insulin 4 IU kg(-1) subcutaneously. One diabetic group and one non-diabetic healthy group served as controls. RESULTS: Treatment for 7 days with urapidil 20 mg kg(-1) and urapidil 60 mg kg(-1) reduced mean glucose concentrations significantly (urapidil-20: 15.6 +/- 1.1 mmol L(-1), P = 0.023; urapidil-60: 15.8 +/- 0.8 mmol L(-1), P = 0.04) compared with diabetic controls (20.9 +/- 0.8 mmol L(-1)), whereas those after urapidil 6 mg kg(-1) were similar to diabetic controls. Insulin treatment normalized blood glucose concentrations. CONCLUSIONS: The alpha1-adrenoceptor antagonist/5-HT1A receptor agonist urapidil has no hyperglycaemic effect on experimental diabetes mellitus, even in high doses, despite its 5-HT1A receptor agonistic properties. PMID- 12113614 TI - Can computers help in assessing children's postoperative pain? Initial validation of a computer-assisted interview. AB - BACKGROUND AND OBJECTIVE: Computers offer the potential for the assessment of children who have difficulties in communication and cannot describe pain using conventional approaches. Such approaches must be reliable and valid. As a preliminary step towards this goal, the validity and reliability of a computer assisted pain assessment for children (MacInterview) was assessed using children with no known disabilities who had undergone surgery. METHODS: MacInterview uses body outlines and a range of different pain representations with scaling for size and intensity, and associated emotion. Following piloting with non-clinic children, the experience of acute postoperative pain was assessed for 30 children undergoing adenotonsillectomy or tonsillectomy the day following surgery using the MacInterview and three existing standardized assessment measures. Each child self-reported their current experience of postoperative pain on two occasions 30 min apart, and retrospectively on pain the previous evening in the immediate postoperative period. RESULTS: Analyses indicated good performance of MacInterview, showing positive correlations between 0.65 and 0.88 with standardized pain-intensity measures, and test-retest reliability was 0.9. Face validity was high, and children enjoyed the procedure. CONCLUSIONS: The procedure shows promise and is likely to merit further development for children showing difficulties in communication. PMID- 12113615 TI - Comparison of different priming techniques on the onset time and intubating conditions of rocuronium. AB - BACKGROUND AND OBJECTIVE: The aim was to compare the effects of two different priming doses and priming intervals with the standard intubating dose of rocuronium on the onset time and intubation conditions. METHODS: After induction of anaesthesia, 75 patients were randomly assigned to one of five groups. Patients in Group 1 received a priming dose of rocuronium 0.06 mg kg(-1) followed 2 min later by rocuronium 0.54 mg kg(-1), Group 2 received a priming dose of 0.10 mg kg(-1) followed 2 min later by a rocuronium injection of 0.50 mg kg(-1). Group 3 was given a priming dose of 0.06 mg kg(-1) followed 3 min later by administration 0.54 mg kg(-1), where Group 4 received a priming dose of 0.10 mg kg(-1) followed 3 min later by injection of 0.50 mg kg(-1). Group 5 received a placebo injection followed 3 min later by rocuronium 0.60 mg kg(-1). RESULTS: Priming with a 3 min priming interval shortened the onset time of rocuronium irrespective of the dosage of (P < 0.001). Clinical duration of action was significantly longer after priming in Group 4 than in Group 5. Clinically acceptable intubation conditions were obtained in all patients. CONCLUSIONS: Priming with a 3 min priming interval was effective when rapid tracheal intubation with rocuronium was necessary. However, priming with rocuronium should be used carefully with special attention given to the possibility of hypoxia and aspiration of gastric contents in awake patients. PMID- 12113616 TI - Effects of adding alfentanil or atracurium to lidocaine solution for intravenous regional anaesthesia. AB - BACKGROUND AND OBJECTIVE: The addition of alfentanil or atracurium to lidocaine solution for intravenous regional anaesthesia of the arm may have advantages with respect to improved muscle relaxation and better analgesia. The study investigates these possibilities. METHODS: We investigated 33 patients. Plain lidocaine solution was administered to Group 1 (n = 11). Alfentanil (0.5 mg) and atracurium (3 mg) were added to the lidocaine solution in Groups 2 (n = 11) and 3 (n = 11), respectively. The onset of sensory and motor block, intra- and postoperative pain scores, and the duration of postoperative analgesia were evaluated. RESULTS: There was a significant difference in the speed of the onset of sensory block in the hand, but not at the tourniquet site. The onset of the motor block, intra- and postoperative pain scores, and the duration of postoperative analgesia were similar in all groups. CONCLUSIONS: No clinical benefits of adding alfentanil or atracurium to lidocaine solution for intravenous regional anaesthesia of the arm could be shown. PMID- 12113618 TI - Epidural anaesthesia for arthroscopic knee surgery in a patient with Takayasu's arteritis. PMID- 12113617 TI - Acute abdomen after coronary artery bypass surgery masked by thoracic epidural analgesia. PMID- 12113619 TI - Workforce and regional distribution of anaesthesiologists in Japan. PMID- 12113620 TI - Insidious hypothyroidism unmasked after operation. PMID- 12113621 TI - Measurement of mean free paths for inelastic electron scattering of Si and SiO2. AB - The effects of accelerating voltage and collection angle on the mean free path for all inelastic electron scattering (lambdap), which is an important parameter for determining specimen thickness by using electron energy-loss spectroscopy, were investigated with crystalline Si and amorphous SiO2. First, thickness of Si film was measured with the convergent-beam electron diffraction method, while thickness of SiO2 particles was estimated from their spherical shape. Then from electron energy-loss spectra, lambdap was evaluated for Si film and SiO2 particles by changing the accelerating voltage (100 to approximately 300 kV) and the collection angle for the scattered electrons. Under the condition of no objective aperture, lambdap for Si film and SiO2 particles was found to increase with the increase of accelerating voltage and to take values of 180+/-6 nm (Si) and 247+/-8 nm (SiO2) at 300 kV. Also, it was found that lambdap in both cases decreases drastically with the increase of collection angle in the range smaller than 25 mrad, while it tends to take a constant value at the collection angle larger than 25 mrad at 200 kV. PMID- 12113622 TI - Single pole-piece objective lens with electrostatic bipotential lens for SEM. AB - This report describes the characteristics for the development of a compound lens that consists of a single pole-piece objective lens and an electrostatic bipotential lens. By applying a relatively small voltage of around 1 kV to the specimen and the bipotential lens, the image quality for low acceleration voltage is improved to a condition better than with just a single pole-piece lens. Even if the wafer is tilted to a large angle, the electric field near the specimen does not become asymmetrical, and there is no occurrence of astigmatic aberration or a reduction of the secondary electron signal. Therefore, 300 mm diameter wafers can be tilted with large angles to observe patterns, particles and defects with high-resolution SEM. Lastly, when the specimen is not tilted, a topographic image of the specimen surface can be obtained by detecting the secondary electron with dual detectors. PMID- 12113623 TI - Morphology of melt-crystallized syndiotactic polypropylene. AB - A transmission electron microscopy (TEM) study was made to investigate the mechanism of isothermal crystallization from melt for syndiotactic polypropylene (sPP) with a syndiotactic pentad fraction of 0.92. The TEM morphologies of RuO4 stained and ultrathin-sectioned specimens revealed a clear assignment of lamellar crystals with different crystal thicknesses to their corresponding melting temperatures and types of crystal structures. It has been found that isolated, single crystal lamellae with cell III structure were grown at 140 degrees C by a plausible crystal growth mechanism with regime I. Fringed-micellar type of lamellae with cell II structure as thin as c. 4 nm were formed in the interstices of the thick chain-folded lamellae by the subsequent quenching. PMID- 12113624 TI - A specimen-drift-free EDX mapping system in a STEM for observing two-dimensional profiles of low dose elements in fine semiconductor devices. AB - We developed a specimen-drift-free energy-dispersive X-ray (EDX) mapping system in a scanning transmission electron microscope (STEM) to improve the sensitivity and spatial resolution of EDX elemental mapping images. The amount of specimen drift was analysed from two STEM images before and after specimen drift by using the phase-correlation method, and was compensated for with an image-shift deflector of the STEM by the displacement of the scanning electron beam. We applied this system to observe the two-dimensional distribution of low dose arsenic in silicon semiconductor devices. The sensitivity of the elemental mapping was improved to several tenths atomic % for arsenic atoms while maintaining a spatial resolution of 2 nm. PMID- 12113625 TI - Differential expression of MHC class II antigens and cathepsin L by subtypes of cortical epithelial cells in the rat thymus: an immunoelectron microscopic study. AB - To better comprehend the thymic microenvironment, it is necessary to identify the antigenic profile of cortical thymic epithelial cells (cTECs) that are involved in the development of major histocompatibility complex (MHC)-restricted T cells. Ultrastructurally, cTECs can be classified into four morphologically distinct subtypes: subcapsular/perivascular (EC1), pale (EC2), intermediate (EC3) and dark (EC4) cells. Several immunohistochemical studies were done on cTECs at the light and electron microscopic levels, but not with reference to the above subtypes. In the present paper, we analysed the expression of MHC class II antigen and cathepsin L by individual cTEC subtypes at the electron microscopic level. We show that (1) MHC class II antigens are expressed on the cell surfaces except on the basal surface of EC1, both on the cell surface and in intracytoplasmic vacuoles of EC2, and only in the intracytoplasmic vacuoles of EC3 and EC4, and (2) that cathepsin L is expressed strongly and uniformly throughout the cytoplasm of EC2, but weakly and non-uniformly in the cytoplasm of EC1, EC3 and EC4. These results show that MHC class II antigen expression and cathepsin L expression is heterogeneous in cTEC subtypes and suggest that EC2 might play a significant role in the development of CD4+ T cells. PMID- 12113626 TI - Remodelling of capillary networks around muscle fibres in the extensor digitorum longus muscle of the normal aged rat. AB - Structural changes of capillaries around muscle fibres following their degeneration and regeneration were further examined in the extensor digitorum longus muscle of the 24-month-old normal rat. Bundles of muscle fibres were divided into three types: muscle bundles consisting of large muscle fibres exclusively more than 35 microm in diameter (type 1), various-sized muscle fibres ranging from 10 to 60 microm in diameter (type 2) and only small muscle fibres 20 40 microm in diameter (type 3). The mean number of capillaries around a muscle fibre was extremely high in the type 3 muscle bundle (4.83) and much lower in the type 2 muscle bundle (2.72) compared with that in the type 1 muscle bundle (3.48). Capillaries in the type 1 muscle bundle were round or oval in shape and were of the continuous type. In the type 2 muscle bundle, capillaries around large degenerating muscle fibres showed an irregularly compressed shape and the scaffolds of basal laminae were often found around them, being a result of the destruction of capillaries. On the other hand, small-sized capillaries less than 5 microm in diameter, being possibly regenerating capillaries, were found around small (probably regenerating) muscle fibres and often had a small number (less than 10) of fenestrae. Capillaries in the type 3 muscle bundle, similar in shape and size to those in the type 1, frequently branched or joined, but some of them were partially destroyed. These findings suggest that capillaries degenerate and regenerate to remodel capillary networks around the muscle fibres following their degeneration and regeneration, and that to effectively supply oxygen and nutrients to regenerating muscle fibres, capillaries temporarily form fenestrae and then the capillary networks become dense by sprouts from the existing capillaries, but excess capillaries may be gradually destroyed following maturation of the muscle fibres. PMID- 12113627 TI - Measurement of the repeat period of myelin sheath using ultrathin frozen sections. AB - The myelin sheath of peripheral nerves was observed by transmission electron microscopy (TEM) using plastic-embedded sections and ultrathin frozen sections. Repeat distances of myelin sheaths were measured in high-powered electron micrographs. The ultrathin frozen sections showed a longer repeat distance than the plastic-embedded sections. The ultrathin frozen sections were thought to contain fewer artefacts, as they had not been subject to dehydration and embedding. It is known that broken myelin sheaths are often observed under conventional TEM. It is thought that these procedures cause contraction and partial destruction of the myelin sheath. PMID- 12113628 TI - Simultaneous collection of topographic and fluorescent images of barley chromosomes by scanning near-field optical/atomic force microscopy. AB - The present study investigated the applicability of scanning near-field optical/atomic force microscopy (SNOM/AFM) to observations of YOYO-1 stained barley chromosomes. Using this technique, topographic and fluorescent images in the same portion were obtained simultaneously, thus enabling the precise analysis of fluorescent structures in relation to the morphology of chromosomes. In YOYO-1 stained chromosomes, the fluorescent intensity roughly reflected the local amount and/or density of DNA in chromosomes. R-banding of chromosomes stained with YOYO 1 and methyl green was also observed clearly as bright spots of various sizes by this microscope. Thus, the SNOM/AFM is expected to become a useful tool for analysing the structure and function of chromosomes more precisely than before. PMID- 12113629 TI - Results of use of metformin and replacement of starch with saturated fat in diets of patients with type 2 diabetes. AB - OBJECTIVE: To improve glycemic control by substituting saturated fat for starch, to identify any adverse effect on lipids masked by the extensive use of metformin and lipid-lowering drugs, and to attempt to separate dietary effects from effects of multiple drugs. METHODS: We undertook a retrospective review of medical records of patients who completed 1 year of follow-up after dietary prescription. The study subjects included 151 patients in the diet group (whose dietary instructions included high saturated fat but starch avoidance) and 132 historical control subjects (who were allowed unlimited monounsaturated fat but had restriction of starch in their diets). RESULTS: Hemoglobin A1c (HbA1c) levels improved in both study groups (-1.4 +/- 0.2% [P<0.001]; 95% confidence interval [CI], -1.9 to -0.9). Use of metformin was associated with a decrease in HbA1c ( 0.12 +/- 0.003%/mo [P<0.001]; 95% CI, -0.17 to -0.07). The diet group had an additional decrease of -0.7 +/- 0.2% (P<0.001; 95% CI, -1.1 to -0.3). Weight increase was associated with the use of insulin (+0.3 +/- 0.07 kg/mo [P<0.001]; 95% CI, 0.2 to 0.5), sulfonylurea (+0.18 +/- 0.06 kg/mo [P<0.01]; 95% CI, 0.05 to 0.30), and troglitazone (+0.7 +/- 0.2 kg/mo [P<0.005]; 95% CI, 0.3 to 1.2). Although not statistically significant, metformin therapy showed a trend for weight loss (-0.14 +/- 0.08 kg/mo; P = 0.07). An additional weight loss was noted in the diet group (-2.65 +/- 0.62 kg [P<0.001]; 95% CI, -3.87 to -1.44). Hydroxymethylglutaryl-coenzyme A reductase inhibitor use was associated with reduced total cholesterol level (-1.7 +/- 0.6 mg/dL per month [P<0.005]; 95% CI, 2.9 to -0.5). The diet group had an additional decrease of -13.0 +/- 4.5 mg/dL (P<0.001; 95% CI, -21.9 to -4.1). No significant effect of the diet on triglyceride, low-density lipoprotein, or high-density lipoprotein levels was detected. CONCLUSION: Addition of saturated fat and removal of starch from a high monounsaturated fat and starch-restricted diet improved glycemic control and were associated with weight loss without detectable adverse effects on serum lipids. PMID- 12113630 TI - Glycohemoglobin assessment program: glycated hemoglobin and epidemiologic variables in patients with type 2 diabetes. AB - OBJECTIVE: To obtain data on glycated hemoglobin (HbA1c) levels and a variety of epidemiologic variables in patients with type 2 diabetes living in the western United States. METHODS: This study was a noncomparative, multicenter, epidemiologic survey. Data were collected from consecutively enrolled patients at nine separate primary-care sites. Patients were eligible for inclusion if they had type 2 diabetes, were between 35 and 70 years old, and had received oral antidiabetic therapy, insulin, or both for at least 3 months before enrollment. RESULTS: Of 602 patients enrolled in the study, 588 were included in the final univariate analyses. The overall mean HbA1c level was 8.2%; however, only 20.4% of patients achieved the American Diabetes Association (ADA) strict HbA1c target of <7.0%. On the other extreme, only 18.9% of patients had HbA1c levels of greater than or equal to 9.5%. Patients treated with insulin had the highest HbA1c levels. Combination therapies were used in 59% of patients, and only 12% were treated with insulin alone. CONCLUSION: The mean HbA1c level in this study is lower than in prior surveys. The use of combination therapy for the management of type 2 diabetes has increased, and the use of insulin as monotherapy has decreased. Although more patients need to reach the ADA HbA1c target of <7.0%, extremely high HbA1c levels are less common than they were in the past. This finding suggests that HbA1c levels are declining, although many patients still have values above 7.0%. PMID- 12113631 TI - Performance of the provider satisfaction inventory to measure provider satisfaction with diabetes care. AB - OBJECTIVE: To develop and validate an inventory to measure provider satisfaction with diabetes management. METHODS: Using the Mayo Clinic Model of Care, a review of the literature, and expert input, we developed a 4-category (chronic disease management, collaborative team practice, outcomes, and supportive environment), 29-item, 7-point-per-item Provider Satisfaction Inventory (PSI). For evaluation of the PSI, we mailed the survey to 192 primary-care and specialized providers from 8 practice sites (of whom 60 primary-care providers were participating in either usual or planned diabetes care). The Cronbach a score was used to assess the instrument's internal reliability. Participating providers indicated satisfaction or dissatisfaction with management of chronic disease by responding to 29 statements. RESULTS: The response rate was 58%. In each category, the Cronbach a score ranged from 0.71 to 0.90. Providers expressed satisfaction with patient-physician relationships, with the contributions of the nurse educator to the team, and with physician leadership. Providers were dissatisfied with their ability to spend adequate time with the patient (3.6 +/- 1.4), their ability to give patients with diabetes necessary personal attention (4.1 +/- 1.2), the efficient passing of communication (4.3 +/- 1.2), and the opportunities for input to change practice (4.3 +/- 1.6). No statistically significant difference (P = 0.12) was found in mean total scores between planned care (5.0 +/- 0.5) and usual care (4.7 +/- 0.6) providers. Moreover, no significant differences were noted across practice sites. CONCLUSION: The PSI is a reliable and preliminarily valid instrument for measuring provider satisfaction with diabetes care. Use in research and quality improvement activities awaits further validation. PMID- 12113632 TI - Bone mineral density screening: assessment of influence on prevention and treatment of osteoporosis. AB - OBJECTIVE: To assess the effect of bone mineral density (BMD) screening on the decision to initiate preventive or therapeutic measures for osteoporosis. METHODS: We offered low-cost BMD screening by dual-energy x-ray absorptiometry of the lumbar spine and proximal femur in conjunction with National Osteoporosis Week. In an effort to assess whether the availability of the BMD measurements resulted in any medical action by the participants of the screening, we conducted a retrospective telephone survey 9 to 12 months after the screening. RESULTS: In response to a newspaper promotion, 350 subjects underwent BMD screening during a 3- to 4-month period. Of these 332 female and 18 male participants, 83% were Caucasian, 10% were Asian, 5% were Hispanic, and 2% were African American. The mean age was 60 +/- 11 years (range, 29 to 93). Osteoporosis (T-score > or = 2.5) was present in 24% and osteopenia (T-score of -1 to -2.49) in 47% of the subjects. A report was sent to the participant and, if requested, also to a specified physician. Of the 350 participants, 249 (238 women and 11 men) responded to the telephone survey. Of these respondents, 63% had sought medical consultation after the BMD screening. Results of the BMD study led to an increase in calcium intake in 32% of female respondents (48% of those with osteoporosis). After BMD measurement, use of osteoporosis therapy approved by the US Food and Drug Administration increased from 38% to 78% of those with osteoporosis. CONCLUSION: These results suggest that low-cost BMD screening is highly effective in increasing awareness of osteoporosis, prompting medical consultation, and initiating measures for prevention and treatment of osteoporosis. PMID- 12113633 TI - Response of bone mineral density to once-weekly administration of risedronate. AB - OBJECTIVE: To evaluate the changes in bone mineral density (BMD) and the tolerability associated with treatment with risedronate, 30 mg once weekly. METHODS: Risedronate, 30 mg, was administered once weekly before breakfast to patients with osteoporosis or osteopenia. All patients were also treated with calcium, 1,500 mg daily, and supplemental vitamin D. Patients receiving treatment with other antiresorptive agents were not excluded. BMD was assessed with dual energy x-ray absorptiometry (DEXA) at baseline and 1 year. RESULTS: After a mean of 11.6 +/- 0.4 months, mean BMD had increased at the lumbar spine and total hip, respectively, by 5.7% (P<0.001) and 2.9% (P<0.001) among all patients, by 4.8% (P<0.001) and 2.2% (P<0.04) among 23 treated with risedronate alone, and by 6.7% (P<0.003) and 3.6% (P<0.004) among 21 receiving risedronate and other antiresorptive agents. Linear regression analysis of the relationship between baseline T-score and BMD increment at the lumbar spine and femoral neck showed a statistically significant negative correlation, an indication of an enhanced response to treatment in patients with lower baseline T-scores. Adverse events possibly associated with risedronate therapy were recorded in 3 of the 70 enrolled patients (dyspepsia in 2 and urticaria in 1). Of 44 patients with follow up DEXA scans, 12 (27%) had not tolerated alendronate (10 mg daily) previously, but they tolerated once-weekly risedronate therapy without difficulty. CONCLUSION: A once-weekly 30-mg regimen of risedronate increases BMD when administered alone or with other antiresorptive agents. Few adverse events possibly associated with risedronate therapy were noted. PMID- 12113634 TI - Combination of gemfibrozil and orlistat for treatment of combined hyperlipidemia with predominant hypertriglyceridemia. AB - OBJECTIVE: To present a case of combined hyperlipidemia with predominant hypertriglyceridemia unresponsive to conventional diet and single-agent drug therapy but successfully treated with a combination of gemfibrozil and orlistat. METHODS: We describe a nonobese Asian Indian man with combined hyperlipidemia. Predominant hypertriglyceridemia was unresponsive to conventional therapy. Orlistat was added to the maximal dose of gemfibrozil, and baseline lipid profiles were compared with posttreatment values after repeated challenges with each drug individually and in combination. The relevant literature was also reviewed. RESULTS: At baseline, the patient's serum triglyceride level was 766 mg/dL and total cholesterol level was 241 mg/dL. On repeated measurements 4 months later, these values were 959 mg/dL and 309 mg/dL, respectively. With use of a reduced-fat diet and gemfibrozil (600 mg orally twice a day), serum triglyceride levels were 830 mg/dL and 909 mg/dL on two different occasions. Combination treatment with the same dosage of gemfibrozil and orlistat at 120 mg orally three times a day reduced triglyceride levels to 279 mg/dL and 244 mg/dL on two separate occasions. Rechallenges with drug monotherapy yielded triglyceride levels of up to 1,159 mg/dL with gemfibrozil alone and of up to 896 mg/dL with orlistat alone. A reduction of serum triglyceride levels to 269 mg/dL and 224 mg/dL occurred when combined treatment with both gemfibrozil and orlistat was reinstituted on two additional occasions. CONCLUSION: The combination of gemfibrozil and orlistat was extremely effective in reducing serum triglyceride levels in this patient with combined hyperlipidemia and predominant hypertriglyceridemia, whereas either one of these agents, when used alone, was ineffective. Determining the mechanisms of this synergy will necessitate further investigation. Additional studies of the use of the gemfibrozil-orlistat combination in patients who have combined hyperlipidemia with predominant hypertriglyceridemia are needed. PMID- 12113635 TI - Immobilization-related hypercalcemia after renal failure in burn injury. AB - OBJECTIVE: To describe a patient with hypercalcemia presumably due to immobilization in the setting of burn injury and acute renal failure. METHODS: We present a case report of a man who sustained a severe burn injury and then had renal failure and hypercalcemia. An additional series of patients with burns and immobilization was assessed for the presence of hypercalcemia. RESULTS: In a 43 year-old man with burns on 65% of his body surface area, acute renal failure developed. Renal function failed to return, and he continued to require hemodialysis. Because of the severity and extensiveness of his burns, he remained immobilized. Serum calcium levels were low during the early part of the hospitalization. On the 57th day, generalized tonic-clonic seizures developed, and he was found to have a high ionized calcium level (1.41 mmol/L). Low values were recorded for intact parathyroid hormone (2 pg/mL), 25-hydroxyvitamin D (5 ng/mL), and 1,25-dihydroxyvitamin D (4 pg/mL). Persistent and recurrent hypercalcemia eventually responded to pamidronate and calcitonin. Other than immobilization, we could identify no predisposing factors such as confounding illnesses or medications that could have caused the hypercalcemia. A review of serum ionized calcium levels in 50 consecutive patients admitted to a burn unit and immobilized for at least 20 days failed to reveal any episodes of persistent hypercalcemia. CONCLUSION: In our patient with burns and renal failure, symptomatic hypercalcemia was most likely attributable to prolonged immobilization. As patients with catastrophic illnesses survive for longer periods, additional problems such as hypercalcemia from immobilization may occur. PMID- 12113636 TI - Langerhans' cell histiocytosis involving the pituitary, thyroid, lung, and liver. AB - OBJECTIVE: To report the development, in an adult patient, of Langerhans' cell histiocytosis (LCH) involving the thyroid and most probably the pituitary gland, lungs, and liver. METHODS: We present a case report of a 29-year-old woman who requested a medical consultation because of polyuria and was found to have pituitary dysfunction. We describe the subsequent follow-up, which revealed progressive liver disease that necessitated transplantation and also the presence of a goiter, and discuss the unpredictable pathologic features of LCH. RESULTS: After the diagnosis of central diabetes insipidus and partial hypopituitarism, the patient was diagnosed 2 years later with sclerosing cholangitis. The hepatic involvement was progressive, and she required transplantation and immunosuppression. Three months before liver transplantation, a goiter was discovered. Fine-needle aspiration of the thyroid revealed infiltration by LCH. The goiter decreased in size after chemotherapy with vinblastine and prednisone. Computed tomography of the chest showed bilateral thin-walled cysts, consistent with eosinophilic granulomas. CONCLUSION: In patients with central diabetes insipidus and pituitary stalk thickening on imaging studies, LCH should be considered in the differential diagnosis. Other hormonal deficiencies may be present initially or may evolve after many years. Thus, continual surveillance is necessary. In addition, an ongoing potential exists for involvement of other organ systems such as the thyroid, liver, and lungs. We suggest consideration of LCH as a possible cause of a goiter in such patients. In our patient, it remains to be seen what effect the immunosuppressive therapy for the liver transplant has on the LCH disease process. PMID- 12113637 TI - Gonadotropin-secreting pituitary tumor associated with hypersecretion of testosterone and hypogonadism after hypophysectomy. AB - OBJECTIVE: To review gonadotropin-secreting pituitary tumors and report the rare case of one of these tumors that caused high serum testosterone concentrations, followed by hypogonadism after hypophysectomy. METHODS: A case report is presented of a 61-year-old man who had decreased vision in his left eye, found by computed tomography of the sella to be attributable to a soft tissue pituitary mass with upward extension that caused elevation and deviation of the optic chiasm. Endocrine and pathologic evaluations are presented, and the treatment and follow-up course are discussed. RESULTS: Endocrine evaluation revealed a serum follicle-stimulating hormone (FSH) of 72.48 mIU/mL, luteinizing hormone (LH) of 31.65 mIU/mL, prolactin of 26.42 ng/mL, and total testosterone of 15.24 ng/mL (all values higher than the normal ranges). A soft tissue mass (3.2 by 2.5 by 1.2 cm) with negative immunocytochemical staining for prolactin and growth hormone but positive staining for synaptophysin, FSH, and LH was removed. One month postoperatively, the patient's chief complaints were a decrease in penile size and erectile dysfunction. Endocrine evaluation revealed a decreased LH of <0.3 mIU/mL, total testosterone of <0.2 ng/mL, and FSH of 4.3 mIU/mL. Three months later with testosterone replacement therapy, testosterone levels normalized, LH was <0.3 mIU/mL, and FSH was 3.9 mIU/mL. Thyroid function and adrenal function were normal before and after surgical intervention. CONCLUSION: This rare case indicates that gonadotropin tumors can produce a functional LH that can increase serum testosterone levels. PMID- 12113638 TI - Lithium-associated thyroiditis. AB - OBJECTIVE: To report an unusual case of thyrotoxicosis caused by "silent thyroiditis" in a lithium-treated patient and to summarize all prior case reports of lithium-associated thyroiditis and compare them with our current case. METHODS: In addition to reporting our case, we undertook a MEDLINE search of all case reports of lithium-associated thyroiditis from 1978 until the present. All reported cases of lithium-associated thyroiditis must have had documented low thyroid radioiodine uptake to be included. RESULTS: A 52-year-old man with a history of bipolar disorder, who had been treated with lithium carbonate for 15 years, was admitted because of delusional mania. Although he had discontinued his lithium therapy 3 months before admission, he had noted symptoms of hyperthyroidism at least 1 month before admission. He was diagnosed with thyrotoxicosis due to silent thyroiditis on the basis of a high free thyroxine level, suppressed thyrotropin level, and low thyroid radioiodine uptake. We found only 10 other case reports of lithium-associated thyrotoxicosis due to silent thyroiditis. CONCLUSION: Thyrotoxicosis caused by silent thyroiditis may be associated with lithium therapy and is likely to be underreported. The pathogenic mechanism for such cases of thyroiditis is still unclear. PMID- 12113639 TI - Primary adrenal insufficiency caused by disseminated histoplasmosis: report of two cases. PMID- 12113640 TI - Nonsteroidal anti-inflammatory drugs and reversible female infertility: is there a link? AB - Nonsteroidal anti-inflammatory drugs (NSAIDs) are frequently prescribed to women of child-bearing age. Three case series highlight the possibility of a link between NSAIDs and reversible infertility. The pharmacological target of NSAIDs is cyclo-oxygenase (COX), which catalyses the first rate-limiting step in the production of prostaglandins. COX-2, one of two isoenzymes, is active in the ovaries during follicular development. Its inhibition is thought to cause luteinised unruptured follicle (LUF) syndrome, an anovulatory condition characterised by clinical signs of ovulation but in the absence of follicular rupture and ovum release. The evidence linking regular NSAID use to reversible LUF syndrome comes from animal studies and three clinical studies. COX-2 deficient mice have severely compromised ovulation in the presence of apparently normal follicular development. Experimental administration of prostaglandins induced ovulation in rabbits and this was blocked by the administration of indomethacin. The three clinical studies demonstrated the induction of delayed follicular rupture or LUF in previously ovulating women by the administration of NSAIDs. A link can therefore be identified between NSAID use and reversible female infertility and NSAID withdrawal should be considered prior to or concurrent with fertility investigations. PMID- 12113641 TI - Genotyping of drug targets: a method to predict adverse drug reactions? AB - In the last decades, advances in molecular biology have led to modern pharmacogenetics, which started as a science that focused on investigating drug metabolising enzymes and genetic determinants of pharmacokinetic variability. As more evidence has become available on the structure of drug targets and the genes coding for them, increasing attention has been directed towards pharmacodynamic explanations of variability in therapeutic response as well as in the risk for adverse drug reactions. Traditionally, genetic drug safety research has focused on variations in single genes whose functions are known to be related to given adverse drug reactions. A few such examples, malignant hyperthermia, the long QT syndrome, venous thromboembolic disease, tardive dyskinesia, and drug addiction, are presented in this article. In the future, results from the Human Genome Project together with tools such as DNA microarray technology, high-output screening systems and advanced bioinformatics, will permit a more thorough elucidation than is currently possible of the genetic components of adverse drug reactions. By screening for a large number of single nucleotide polymorphisms (SNPs), SNP patterns associated with adverse drug reactions can be discovered even though the functions of the SNPs as such are completely unknown. On the basis of these findings, it can be expected that pharmacogenetic research will identify situations where a drug should be avoided in certain individuals in order to reduce the risk for adverse drug reactions. If so, it will be feasible to use molecular diagnostics to select drugs that are safe for the individual patient. PMID- 12113643 TI - Safety of unoprostone isopropyl as mono- or adjunctive therapy in patients with primary open-angle glaucoma or ocular hypertension. AB - This review summarises the safety of unoprostone isopropyl (both at the 0.12 and 0.15% concentrations) instilled twice daily in patients with primary open-angle glaucoma (POAG) or ocular hypertension (OH). For unoprostone 0.15%, combined data from two 12-month comparative monotherapy studies are reported, as well as data from three adjunctive therapy studies and two special population studies. With unoprostone monotherapy, most adverse events were mild or moderate and transient in nature. Less than 7% of unoprostone-treated patients discontinued therapy due to an adverse event. The most common adverse events associated with unoprostone were burning/stinging, burning/stinging directly upon drug instillation, ocular itching, and conjunctival hyperaemia. Unoprostone had no clinically notable effects on vital signs, laboratory profiles, or comprehensive ophthalmic examinations. One of 659 unoprostone 0.15%-treated patients had a change in iris colour after 12 months of monotherapy. Except for a higher incidence of burning/stinging and burning/stinging upon instillation, unoprostone was comparable to timolol 0.5% twice daily and betaxolol 0.5% twice daily. No safety concerns were raised with use of unoprostone as adjunctive therapy. Unoprostone had no significant effect on exercise-induced heart rate in healthy subjects or on pulmonary function in patients with mild-to-moderate asthma. The safety profile of unoprostone 0.15% was consistent with published information on the 0.12% formulation. In conclusion, unoprostone has an excellent safety profile in patients with POAG or OH. PMID- 12113644 TI - Safety and tolerability of eprosartan in combination with hydrochlorothiazide. AB - The ideal antihypertensive drug should be effective in reducing blood pressure, but have a low incidence of adverse effects. Angiotensin II receptor blockers, such as eprosartan, are as effective as ACE inhibitors in reducing blood pressure, but lack the main adverse effect of ACE inhibitors, namely cough. Eprosartan has been shown to be well tolerated with a placebo-like adverse-effect profile. When given as monotherapy it is effective in reducing blood pressure; however, some patients require additional blood pressure control, which may be provided by combination therapy. Indeed, the combination of eprosartan and the thiazide diuretic hydrochlorothiazide has been shown to be effective in further reducing blood pressure in patients not optimally responding to eprosartan monotherapy. This article reviews the safety and tolerability of eprosartan in combination with hydrochlorothiazide from 17 studies of 1899 patients with hypertension and normotensive volunteers. Of these studies, four were controlled with patients receiving a fixed-dose combination, six were long-term, open-label, and another four were controlled studies with hydrochlorothiazide being given to eprosartan non-responders. The other three studies included healthy subjects receiving the combination of eprosartan and hydrochlorothiazide. There was a high completion rate in all studies evaluated. Most of the patients receiving eprosartan 600mg in combination with hydrochlorothiazide 12.5mg daily completed the studies, which supports acceptance of this combination therapy by patients. The most frequently reported adverse events in these combination studies were headache, dizziness, myalgia, and upper respiratory tract infection in patients with hypertension. The majority of adverse events were mild to moderate in intensity, and were not considered to be related to study treatment. The adverse event that was more common in patients receiving combination therapy compared with those receiving monotherapy was dizziness. This adverse event may be due to hydrochlorothiazide as it has previously been observed in patients taking thiazide diuretics. In healthy volunteers, the most frequently reported adverse events were headache, dizziness, and upper respiratory tract infection. However, none of these adverse events were considered related to study medication. In summary, the combination of eprosartan/hydrochlorothiazide is well tolerated, both as short- and long-term therapy, with most adverse events occurring early. The most frequent adverse events were headache, dizziness, and upper respiratory infection, which would be expected based on the safety profile of each of the components. Therefore, the combination of eprosartan with hydrochlorothiazide can be effectively and safely used in patients not adequately responding to eprosartan monotherapy. PMID- 12113642 TI - Comparative tolerability of therapies for ulcerative colitis. AB - This article reviews the clinical pharmacology, adverse events, and comparative tolerability of the drugs commonly available for treating ulcerative colitis. Synthetic glucocorticoids are the most commonly used conventional corticosteroids in the treatment of ulcerative colitis. Corticosteroids can be expected to impact on every organ system and most metabolic activities of the body. Suppression of the hypothalamic-pituitary-adrenal axis is common, but reversible, with conventional corticosteroids, but not with newer topically-acting corticosteroids. A serious complication of corticosteroids in children is growth retardation. The frequent adverse effects associated with the use of corticosteroids have prompted the development of a new group of rectal agents with equivalent efficacy and a more benign adverse event profile such as prednisolone metasulfobenzoate, fluticasone propionate, tixocortol pivalate, beclomethasone dipropionate and budesonide. The incidence of adverse effects related to the use of sulfasalazine (5-aminosalicylic acid plus sulfapyridine) is high and is dose related. The most frequently reported adverse effect is intolerance, not allergy, and relates to the sulfapyridine moiety correlating with the acetylator phenotype. Tolerance to 5-aminosalicylic acid by 80 to 90% of those patients allergic to, or intolerant of, sulfasalazine has given further evidence suggesting that the sulfa moiety is responsible for much of the toxicity of sulfasalazine. However, 10 to 20% of patients who are sulfasalazine intolerant have similar reactions to 5-aminosalicylic acid formulations, indicating that the 5-aminosalicylic acid moiety is responsible for adverse events in some patients taking sulfasalazine. Adverse effects resulting from treatment with azathioprine and mercaptopurine can be divided into two categories: allergic-type reactions that appear to be dose-independent and nonallergic-type reactions that are probably dose- and metabolism-dependent. It is well established now that genotype and thiopurine methyltransferase activity have an important impact on the rate of adverse effects during azathioprine or mercaptopurine therapy. Adverse effects resulting from high dose cyclosporin therapy for inflammatory bowel disease include: renal insufficiency, hypertension, opportunistic infections, seizures, paresthesias, tremor, headache, gingival hyperplasia, hypertrichosis, and anaphylaxis with intravenous cyclosporin. In contrast, the incidence of adverse events was relatively low when low-dose oral cyclosporin was used. The incidence of adverse events associated with any of the medications used in the treatment of ulcerative colitis is difficult to assess and it is therefore hard to make a comparative evaluation. The broadening of the drug regimen available to the clinician has advanced our knowledge about the disease, and further development of more effective, less toxic agents can be anticipated in the future. PMID- 12113645 TI - Approaches toward repeated supratherapeutic doses of paracetamol in children: a survey of medical directors of poison centres in North America and Europe. AB - BACKGROUND: During the last few years there has been an increase in the number of reports of liver failure associated with prolonged paracetamol (acetaminophen) administration in children for therapeutic reasons. OBJECTIVE: To describe the approach taken by medical directors of poison centres regarding the treatment of repeated supratherapeutic doses of paracetamol in children. METHODS: Questionnaires mailed to the medical directors of 76 poison centres in North America and 48 poison centres in Europe asked respondents to comment on the management of a hypothetical case of a child who had received repeated doses of paracetamol at a daily dose of 90 mg/kg during a febrile illness and who presented to the emergency department with mildly elevated serum transaminase levels. RESULTS: The response rate was 62% for North American centres and 44% for European centres. There was a wide range of answers regarding the maximal safe daily dose of paracetamol. For the case described, 71% of the respondents suggested measuring paracetamol serum concentration. Fifty-four percent suggested treating the patient with acetylcysteine and 35% suggested not treating the patient. CONCLUSIONS: Our study showed that there is little agreement among medical directors of poison centres on the toxic threshold for chronic exposure to paracetamol in children and on how these cases should be managed. PMID- 12113646 TI - Epidermolysis bullosa: new and emerging trends. AB - Epidermolysis bullosa is a family of inherited blistering skin disorders characterized by blister formation in response to mechanical trauma. Major types of epidermolysis bullosa include epidermolysis bullosa simplex, hemidesmosomal epidermolysis bullosa, junctional epidermolysis bullosa, and dystrophic epidermolysis bullosa. Current treatment for epidermolysis bullosa consists of supportive care for skin and other organ systems and entails a combination of wound management, infection support for chronic wounds, surgical management as needed, nutritional support, and preventative screening for squamous cell carcinoma in recessive dystrophic epidermolysis bullosa. The regimen must be tailored specifically to the severity and extent of skin and systemic involvement in each case. Recent studies have identified specific protein and genetic abnormalities for most epidermolysis bullosa subtypes. These new advancements in the understanding of molecular pathophysiology have provided much of the basis for current efforts to develop effective gene and protein therapy for epidermolysis bullosa. PMID- 12113647 TI - Topical noncorticosteroid immunomodulation in the treatment of atopic dermatitis. AB - At present, the first-line drugs for treating atopic dermatitis are topical corticosteroids. They are effective when used short-term; however, long-term use of the corticosteroids is associated with suppressive effects on the connective tissue, seen as skin atrophy or resistance to therapy. Currently, two topical noncorticosteroid immunomodulators tacrolimus (FK506) and pimecrolimus (SDZ ASM 981) are under development, or already on the market in some countries for atopic dermatitis. These two compounds show structural similarity. In T lymphocytes they bind to the same cellular receptor, the FK-binding protein (FKBP) or macrophilin 12. Tacrolimus shows a 3-fold greater affinity to FKBP compared with pimecrolimus. The tacrolimus/ pimecrolimus-FKBP complex further binds to calcineurin, an enzyme vital for the early activation of T cells. The consequence of calcineurin binding is a lack of activation of both T helper cell types 1 and 2. Further effects of these compounds have been suggested on other inflammatory cells, such as Langerhans cells and mast cells/basophils. In contrast to corticosteroids, no suppressive effects on connective tissue cells have been observed. Taken together, treatment of inflammation results in healing of the barrier function of the skin. This again results in reduced bioavailability of the drug, as compared with systemic use. Placebo-controlled studies have shown the efficacy of both tacrolimus (at 0.03 and 0.1%) and pimecrolimus (at 0.6 and 1%). The main adverse event in these studies has been a burning sensation and increased pruritus at the site of application. Typically, these adverse events are observed only during the first days of treatment. Long-term safety studies, of up to one year, have not revealed any new adverse events. So far, long-term use of topical noncorticosteroid compounds has not been associated with signs of immune deficiency. Although there is currently no evidence for clinically relevant, prolonged adverse effects, some of these, such as an increased risk of photocarcinogenesis, need to be monitored. There is evidence from tacrolimus studies that monotherapy results in better long-term results when compared with combination therapy with corticosteroids. Tacrolimus and pimecrolimus could replace topical corticosteroids as the first-line treatment of atopic dermatitis. PMID- 12113648 TI - Pustular skin disorders: diagnosis and treatment. AB - The differential diagnosis for pustular skin disorders is extensive. The distribution of the lesions and the age of the patient are characteristics that may provide strong clues to the etiology of cutaneous pustular eruptions. In adults, generalized pustular dermatoses include pustular psoriasis, Reiter's disease and subcorneal pustular dermatosis. Medications can cause generalized pustular eruptions, such as in the case of acute generalized exanthematous pustulosis; or more localized reactions, such as acneiform drug eruptions, which usually involve the face, chest and back. Localized pustular eruptions are seen on the hands and feet in adults with pustulosis palmaris et plantaris and acrodermatitis continua (both of which may be variants of psoriasis); on the face in patients with acne vulgaris, rosacea, and perioral dermatitis; and on the trunk and/or extremities in patients with folliculitis. A separate condition known as eosinophilic folliculitis occurs in individuals with advanced human immunodeficiency disease. Severely pruritic, sterile, eosinophilic pustules are found on the chest, proximal extremities, head and neck. Elevated serum immunoglobulin E and eosinophilia are often concurrently found. In neonates, it is especially important to make the correct diagnosis with respect to pustular skin disorders, since pustules can be a manifestation of sepsis or other serious infectious diseases. Generalized pustular eruptions in neonates include erythema toxicum neonatorum and transient neonatal pustular melanosis, both of which are non-infectious. Pustules are seen in infants with congenital cutaneous candidiasis, which may or may not involve disseminated disease. Ofuji's syndrome is an uncommon generalized pustular dermatosis of infancy with associated eosinophilia. As in adults, neonates and infants may develop acne or scabies infestations. In this article, we review the most common pustular dermatoses and offer a systematic approach to making a diagnosis. We also report the most up-to date information on the treatment of these various cutaneous pustular conditions. PMID- 12113649 TI - Current concepts in the management of patients with melanoma. AB - Melanoma is a significant health problem. Despite public education and free cancer screenings, the incidence and mortality of melanoma continues to rise; however, many currently diagnosed melanomas are thin lesions, suggesting that education and awareness is having an impact. In addition, there are still subsets of patients who need increased surveillance in order to increase their survival. Although large congenital nevi may be precursors of melanoma, small and medium congenital nevi have an insignificant risk for melanoma development. Large congenital nevi, which are axial in location, appear to be more likely to develop melanoma and are associated with melanocytosis and melanoma of the CNS, both of which portend a poor prognosis. Recently, the recommended margins of excision have become more conservative so that many of the surgical defects can be closed primarily. Lymphoscintigraphy and sentinel node biopsy have replaced elective node dissections, thus decreasing the morbidity associated with the surgical management of melanoma. Although controversy still exists as to whether or not sentinel lymph node biopsy alters a patient's prognosis, it has been shown to be a powerful prognostic indicator. Although most melanomas are managed by routine surgical excision, other modalities are sometimes employed. For example, cryosurgery or radiation therapy may be indicated in the frail, elderly individual with a large facial lentigo maligna. Mohs surgery is the treatment of choice for head and neck melanomas and those located in areas where maximum preservation of tissue is required and for desmoplastic and acral lentiginous melanomas. Much more work remains in the area of adjuvant therapy, chemotherapy, and immunotherapy. Dacarbazine remains the drug of choice in disseminated melanoma, but remissions are usually short lived. Interleukin and biochemotherapy has yielded good results but the percentage benefiting is small. Although high dose interferon increases disease-free and overall survival in some patients, it remains a controversial drug which is not easily tolerated. In the new staging system for melanoma, ulceration is second only to Breslow's thickness. In transit (satellite) lesions have also been included in this new system. The new system also recognizes that patients with only microscopic metastatic nodal disease fare better than patients with clinically enlarged metastatic nodes and that it is the number of nodes involved with metastases, not their size, that determines the patient's prognosis. Except for lesions <1mm thick, the Clark's level of invasion has been de-emphasized. PMID- 12113650 TI - Topical use of dexpanthenol in skin disorders. AB - Pantothenic acid is essential to normal epithelial function. It is a component of coenzyme A, which serves as a cofactor for a variety of enzyme-catalyzed reactions that are important in the metabolism of carbohydrates, fatty acids, proteins, gluconeogenesis, sterols, steroid hormones, and porphyrins. The topical use of dexpanthenol, the stable alcoholic analog of pantothenic acid, is based on good skin penetration and high local concentrations of dexpanthenol when administered in an adequate vehicle, such as water-in-oil emulsions. Topical dexpanthenol acts like a moisturizer, improving stratum corneum hydration, reducing transepidermal water loss and maintaining skin softness and elasticity. Activation of fibroblast proliferation, which is of relevance in wound healing, has been observed both in vitro and in vivo with dexpanthenol. Accelerated re epithelization in wound healing, monitored by means of the transepidermal water loss as an indicator of the intact epidermal barrier function, has also been seen. Dexpanthenol has been shown to have an anti-inflammatory effect on experimental ultraviolet-induced erythema. Beneficial effects of dexpanthenol have been observed in patients who have undergone skin transplantation or scar treatment, or therapy for burn injuries and different dermatoses. The stimulation of epithelization, granulation and mitigation of itching were the most prominent effects of formulations containing dexpanthenol. In double-blind placebo controlled clinical trials, dexpanthenol was evaluated for its efficacy in improving wound healing. Epidermal wounds treated with dexpanthenol emulsion showed a reduction in erythema, and more elastic and solid tissue regeneration. Monitoring of transepidermal water loss showed a significant acceleration of epidermal regeneration as a result of dexpanthenol therapy, as compared with the vehicle. In an irritation model, pretreatment with dexpanthenol cream resulted in significantly less damage to the stratum corneum barrier, compared with no pretreatment. Adjuvant skin care with dexpanthenol considerably improved the symptoms of skin irritation, such as dryness of the skin, roughness, scaling, pruritus, erythema, erosion/fissures, over 3 to 4 weeks. Usually, the topical administration of dexpanthenol preparations is well tolerated, with minimal risk of skin irritancy or sensitization. PMID- 12113652 TI - Cutaneous drug reaction case reports: from the world literature. PMID- 12113651 TI - Spotlight on topical pimecrolimus in atopic dermatitis. AB - Pimecrolimus (SDZ ASM 981), an ascomycin derivative, is a nonsteroid, has anti inflammatory activity, and has demonstrated efficacy in reducing symptoms of atopic dermatitis in adult and pediatric patients when applied topically. Compared with vehicle, topical pimecrolimus 1.0% cream was significantly more effective at reducing symptoms of atopic dermatitis, as measured by the Eczema Area and Severity Index (EASI), in infants aged 3 to 23 months, children aged 2 to 17 years, and adults. The median reductions from baseline in the total EASI score in adults after treatment with pimecrolimus 1.0% or corresponding vehicle twice daily for 3 weeks were 47 and 0%, respectively. In infants and children, treatment with pimecrolimus 1.0% twice daily for 6 weeks resulted in significant decreases in mean EASI scores compared with vehicle. The severity of pruritus was significantly reduced in patients of all age groups after topical treatment with pimecrolimus 1.0% cream. Compared with vehicle, the incidence of eczematous flares was also reduced by intermittent long-term use of topical pimecrolimus 1.0% in adults, children and infants. Sixty-one percent of children treated with pimecrolimus for 1 year completed the first 6 months of treatment without experiencing a flare, compared with 35% of patients who received vehicle. Furthermore, the use of topical corticosteroids for the treatment of uncontrolled flares in adults, children and infants was lower in the pimecrolimus groups than in the vehicle groups. Topical pimecrolimus 1.0% cream is well tolerated in atopic dermatitis patients of all age groups. There were no clinically relevant systemic adverse events reported from any of the studies in patients with atopic dermatitis. The most frequently reported adverse events pertained to application site reactions, such as burning and a feeling of warmth. In conclusion, topical pimecrolimus 1.0% cream has shown efficacy in the treatment of mild to moderate atopic dermatitis in infants, children and adults. Although tolerability data concerning infants and children have not yet been published in full, the drug appears to be well tolerated in all age groups, and there have been no reports of clinically relevant systemic adverse events. Furthermore, pimecrolimus has shown no potential for skin atrophy, a problem commonly associated with treatment with topical corticosteroids. Topical pimecrolimus 1.0% provides a promising and well tolerated treatment option in the management of infants, children and adults with mild to moderate atopic dermatitis. PMID- 12113653 TI - Small interfering RNAs as a tool to assign Rho GTPase exchange-factor function in vivo. AB - Rho GTPases control a complex network of intracellular signalling pathways. Whereas progress has been made in identifying downstream signalling partners for these proteins, the characterization of Rho upstream regulatory guanine nucleotide exchange factors (GEFs) has been hampered by a lack of suitable research tools. Here we use small interfering RNAs (siRNAs) to examine the cellular regulation of the RhoB GTPase, and show that RhoB is activated downstream of the epidermal-growth-factor receptor through the Vav2 exchange factor. These studies demonstrate that siRNAs are an ideal research tool for the assignment of Rho GEF function in vivo. PMID- 12113654 TI - Characterization of cationic lipid DNA transfection complexes differing in susceptability to serum inhibition. AB - BACKGROUND: Cationic lipid DNA complexes based on DOTAP (1,2-dioleoyl-3 (trimethyammonium) propane) and mixtures of DOTAP and cholesterol (DC) have been previously optimized for transfection efficiency in the absence of serum and used as a non-viral gene delivery system. To determine whether DOTAP and DC lipid DNA complexes could be obtained with increased transfection efficiency in the presence of high serum concentrations, the composition of the complexes was varied systematically and a total of 162 different complexes were analyzed for transfection efficiency in the presence and absence of high serum concentrations. RESULTS: Increasing the ratio of DOTAP or DC to DNA led to a dose dependent enhancement of transfection efficiency in the presence of high serum concentrations up to a ratio of approximately 128 nmol lipid/microg DNA. Transfection efficiency could be further increased for all ratios of DOTAP and DC to DNA by addition of the DNA condensing agent protamine sulfate (PS). For DOTAP DNA complexes with ratios of < or = 32 nmol/microg DNA, peak transfection efficiencies were obtained with 4 microg PS/microg DNA. In contrast, increasing the amount of PS of DC complexes above 0.5 microg PS/microg DNA did not lead to significant further increases in transfection efficiency in the presence of high serum concentrations. Four complexes, which had a similar high transfection efficiency in cell culture in the presence of low serum concentrations but which differed largely in the lipid to DNA ratio and the amount of PS were selected for further analysis. Intravenous injection of the selected complexes led to 22-fold differences in transduction efficiency, which correlated with transfection efficiency in the presence of high serum concentrations. The complex with the highest transfection efficiency in vivo consisted of 64 nmol DC/ 16 microg PS/microg DNA. Physical analysis revealed a predicted size of 440 nm and the highest zeta potential of the complexes analyzed. CONCLUSIONS: Optimization of cationic lipid DNA complexes for transfection efficiency in the presence of high concentrations of serum led to the identification of a DC complex with high transduction efficiency in mice. This complex differs from previously described ones by higher lipid to DNA and PS to DNA ratios. The stability of this complex in the presence of high concentrations of serum and its high transduction efficiency in mice suggests that it is a promising candidate vehicle for in vivo gene delivery. PMID- 12113655 TI - Transjugular renal biopsy in high-risk patients: an American case series. AB - BACKGROUND: In the United States, transjugular renal biopsies using the Quickcore side cut needle system have previously been described primarily for transjugular renal biopsy in patients with concurrent liver and kidney disease. METHODS: We describe transjugular renal biopsy with the Quickcore trade mark system in 9 patients with nephrotic syndrome and contraindications to percutaneous renal biopsy, who underwent biopsy between 23 October 1996 and 12 April 2001. The most common contraindication was oral anticoagulation with coumadin (40%). Other contraindications included horseshoe kidney, severe renal failure, and spontaneous coagulopathy. A 62 cm straight catheter and 60 cm side-cut Quickcore biopsy needle were used to obtain cortical tissue. Packing of the biopsy tract with Gelfoam was used for venographically identified capsular perforation. RESULTS: Ten procedures were performed on 9 patients with one requiring re-biopsy (5% of all renal biopsies performed at our institution). There were 9 transjugular renal biopsy and one combined liver-kidney biopsy. A mean of 4 +/- 2 passes were made, with a mean of 3 +/- 1 cores obtained per procedure. Histologic diagnosis was made in 90% of biopsies and in 100% of patients. Two patients developed transient hydronephrosis associated with gross hematuria; both required transfusion. Capsular perforation occurred in 90%. One patient died of bacterial sepsis, unrelated to the biopsy, several days after the procedure. CONCLUSIONS: Transjugular renal biopsy appears to be efficacious in high-risk patients, for whom the percutaneous approach is contraindicated, including patients on oral anticoagulation. The transfusion rate in the present study was similar to other American reports using this technique. PMID- 12113657 TI - Occurrence of 3-chloro-propane-1,2-diol (3-MCPD) and related compounds in foods: a review. AB - A critical review of the occurrence of 3-chloro-propane-1,2-diol (3-MCPD) in foods not known to contain hydrolysed vegetable proteins is presented. The review covers the properties and chemistry of 3-MCPD and the current methods of analysis in foodstuffs. The results of UK surveys of 3-MCPD occurrence in both retail foods and commercial food ingredients are discussed with particular reference to cereal, meat and dairy products. The possible mechanisms for the formation and decay of 3-MCPD in foods are suggested. The review does not cover the detailed toxicology of 3-MCPD and its occurrence in hydrolysed vegetable proteins, which have been considered elsewhere, nor possible issues such as in-vivo formation. PMID- 12113656 TI - Loss of cellular adhesion to matrix induces p53-independent expression of PTEN tumor suppressor. AB - BACKGROUND: The tumor suppressor gene PTEN has been found mutated in many types of advanced tumors. When introduced into tumor cells that lack the wild-type allele of the gene, exogenous PTEN was able to suppress their ability to grow anchorage-independently, and thus reverted one of the typical characteristics of tumor cells. As these findings indicated that PTEN might be involved in the regulation of anchorage-dependent cell growth, we analyzed this aspect of PTEN function in non-tumor cells with an anchorage-dependent phenotype. RESULTS: We found that in response to the disruption of cell-matrix interactions, expression of endogenous PTEN was transcriptionally activated, and elevated levels of PTEN protein and activity were present in the cells. These events correlated with decreased phosphorylation of focal adhesion kinase, and occurred even in the absence of p53, a tumor suppressor protein and recently established stimulator of PTEN transcription. CONCLUSIONS: In view of PTEN's potent growth-inhibitory capacity, we conclude that its induction after cell-matrix disruptions contributes to the maintenance of the anchorage-dependent phenotype of normal cells. PMID- 12113658 TI - Development of methods for the determination of vitamins A, E and beta-carotene in processed foods based on supercritical fluid extraction: a collaborative study. AB - New methodologies based on supercritical fluid extraction (SFE) have been developed for the determination of fat-soluble vitamins in processed foods. The results obtained so far indicate that SFE is well suited to extraction of fat soluble vitamins from food products, although validation work is required to establish accuracy and precision. The vitamins investigated were A, E and beta carotene, and the processed foods were UHT milk, milk powder, minced meat, liver paste, infant formula, canned baby food and margarine. Extraction equipment employed analyte collection on either a solid-phase trap or in a solvent. After extraction, the samples were saponified and the vitamins determined using reversed-phase liquid chromatography with ultraviolet or fluorescence detection. Sample throughput was at least 12 samples day(-1), i.e. at least twice the number achievable with a conventional extraction methodology. The detection limits for the vitamins in different processed foods were well below 0.1 microg g(-1). Recoveries (in comparison with vitamin levels obtained using conventional solvent extraction) were close to 100% for experienced personal with access to modern automatic equipment. To reach this level, it was necessary to protect the vitamins with an antioxidant during the different steps of the analysis procedure, to add methanol or ethanol to the extraction cell to facilitate the analyte extraction from the food matrix, and when using a solid-phase trap, to employ a fractionated extraction-elution procedure to prevent breakthrough losses. The developed methods were tested in a validation exercise between five laboratories, which had taken part in the method development, and in an intercomparison between 10 laboratories including laboratories with less experience of vitamin determination. The within-laboratory RSD was generally < or = 11%. The average of the between-laboratory relative standard deviation (RSD) was about 23% in the validation, and increased to about 40% in the intercomparison. Ruggedness tests performed at different steps of the project showed that different types and models of equipment did not give large differences in recoveries. Thus, the increasing RSD can largely be ascribed to differences in experience in vitamin analysis of the participants. PMID- 12113659 TI - Estimates of the mean per capita daily intake of benzoic and sorbic acids in Brazil. AB - The daily intakes of benzoates and sorbates from selected food categories were estimated in Brazil in 1999. The Budget method was used as a first screening procedure for the estimation of the safety aspects of the maximum permitted levels of benzoates and sorbates established by the Brazilian food legislation. This screening indicated that benzoates should be further investigated. In a second step, the daily intakes of these preservatives were assessed by combining measured levels of these additives with national food consumption data derived from a household economic survey and a packaged good market survey. Benzoate and sorbate levels in soft drinks, fruit juices, margarine, yoghurt and cheese were determined by HPLC with a photodiode array detector (detection at 228 nm for benzoic acid, 260 nm for sorbic acid). The estimated intakes of benzoates and sorbates for the average consumer were below the ADIs, ranging from 0.3 to 0.9 and 0.2 to 0.3 mg kg(-1) body weight, respectively. Soft drinks were identified as the main source of benzoates representing >80% of the estimated intake. PMID- 12113660 TI - Research on the origin, and on the impact of post-harvest handling and manufacturing on the presence of ochratoxin A in coffee. AB - The major risk factors and processing steps that can lead to contamination of green coffee with ochratoxin A (OTA) have been identified. Surveys of the green coffee production chain indicate that Aspergillus ochraceus and A. carbonarius are the most potent OTA producers on coffee. Both have been successfully grown in vitro on green coffee and coffee cherries, respectively, producing high amounts of OTA (5-13 mg kg(-1)). The so-called dry processing of coffee, which is cherry drying, was identified as one of the steps during which OTA formation can take place, particularly under humid tropical conditions. Cherries contain sufficient amounts of water to support mould growth and OTA formation during the initial 3-5 days of drying on the outer part of the cherries. Not surprisingly, after dehulling, husks can be highly contaminated with OTA, as also indicated by its enhanced concentration in soluble coffees adulterated with husks and parchment. A minimum water activity of 0.80 (about 14% MC) is required for in vitro OTA production on green coffee, a fact that does not rule out the possibility of OTA contamination due to improper transportation and storage of green coffee. However, this appears not to be a major route for OTA contamination of coffee. OTA contamination can clearly be minimized by following good agricultural practice and a subsequent post-harvest handling consisting of appropriate techniques for drying, grading, transportation and storage of green coffee; these procedures are well established. PMID- 12113661 TI - Occurrence of aflatoxin in commodities imported into Qatar, 1997-2000. AB - The occurrence of aflatoxin in commodities imported into Qatar was investigated from 1999 to 2000. During the 4 years, 351 samples of susceptible commodities were analysed. Aflatoxin was detected in 71 (20%) samples in the range 0.1-20 microg kg(-1) and in 50 (14%) samples above the permitted level of 20 microg kg( 1). The highest incidence and levels of aflatoxin contamination were recorded in pistachio without shell followed by pistachio with shell. Aflatoxin levels >20 microg kg(-1) in the pistachio samples varied from 8.7 to 33%. The highest level of total aflatoxin found in pistachio without shell was 289 microg kg(-1). A few samples of corn and corn products (three of 54 analysed), peanut and peanut products (nine of 42 analysed) and other nuts like almond, walnut and cashew (one of 40 analysed) were found contaminated with low levels (0.1-20 microg kg(-1)) of aflatoxins. Only one sample of custard powder and one sample of roasted peanut were found with aflatoxin >20 microg kg(-1) PMID- 12113662 TI - Occurrence of aflatoxins and fumonisins in Incaparina from Guatemala. AB - The occurrence of aflatoxins and fumonisins in Incaparina was investigated. Incaparina is a mixture of corn and cottonseed flour with added vitamins, minerals and a preservative. It has been marketed as a high-protein food supplement, particularly for children on protein-deficient diets. According to estimates, 80% of Guatemalan children in their first year are given Incaparina to provide an adequate diet. Eight samples of Incaparina manufactured in Guatemala were collected. Five were from three different geographical locations in the USA and three were from Guatemala. Seven were examined for fungal contamination and analysed for aflatoxins and fumonisins. Aspergillus flavus was the predominant fungus in all samples purchased in the USA and in one sample purchased from Guatemala, whereas Fusarium verticillioides was present in only two samples (one from the USA and one from Guatemala). All samples contained aflatoxins, ranging from 3 to 214 ng g(-1) and <2 to 32ng g(-1) for aflatoxin B(1) and aflatoxin B(2), respectively; and one sample contained aflatoxin G(1) (7 ng g(-1)). Total aflatoxins present ranged from 3 to 244 ng g(1). All samples contained fumonisins, ranging from 0.2 to 1.7 microg g(-1), <0.1 to 0.6 microg g(-1), and <0.1 to 0.2 microg g(-1) for fumonisins B(1), fumonisin B(2), and fumonisin B(2), respectively. Total fumonisins present ranged from 0.2 to 2.2 microg g(-1). The identity of aflatoxin B(2) was confirmed using both the chemical derivatization method and liquid chromatographic (LC)/mass spectrometric (MS) analysis. Appropriate regulatory action was recommended for the import of Incaparina and has been in effect since 22 December 1998. PMID- 12113663 TI - Fumonisin B(1) in maize harvested in Iran during 1999. AB - The fumonisin B(1) (FB(1)) contamination of maize collected in two areas of Iran during 1999 was determined. The 20 maize samples from Mazandaran Province, situated on the Caspian littoral of Iran, consisted of random samples of farmers' lots and were all contaminated with FB(1) at a mean level of 3.18 mg kg(-1) (range 0.68-7.66 mg kg(-1)). The 10 samples (of the same maize cultivar) from Isfahan Province in central Iran were purchased as maize cobs in local retail markets and had mean FB levels of 0.22 mg kg(-1) (mean of all samples, 6/10 samples positive, range <0.01-0.88 mg kg(-1)). The FB levels in Mazandaran, an area of high oesophageal cancer, were significantly (p < 0.0001) higher than the FB levels found in maize from Isfahan, an area of low oesophageal cancer in Iran. PMID- 12113664 TI - Removal of common Fusarium toxins in vitro by strains of Lactobacillus and Propionibacterium. AB - This study was conducted to examine the ability of selected strains of Lactobacillus and Propionibacterium to remove common Fusarium toxins, trichothecenes, from liquid media. The trichothecenes studied were deoxynivalenol (DON), 3-acetyldeoxynivalenol (3-AcDON), nivalenol (NIV), fusarenon (FX), diacetoxyscirpenol (DAS), T-2 toxin (T-2) and HT-2 toxin (HT-2). The Lactobacillus rhamnosus strain GG (LGG), Lactobacillus rhamnosus strain LC-705 (LC-705) and Propionibacterium freudenreichii ssp. shermanii JS (PJS) were incubated in PBS buffer containing 20 microg toxin ml(-1) for 1h at 37 degrees C, and after centrifugation the concentration of the toxins was measured in the supernatant fraction. Both viable and heat-killed forms of LGG and PJS were more efficient than LC-705 in removing the toxins from the liquid media. LGG and PJS removed four of the seven tested toxins (the removal varying from 18 to 93%) and LC-705 two toxins (10-64%). Of the toxins, 3-AcDON was not removed by any of the bacteria; HT-2 was removed by the non-viable LGG and also slightly by non-viable LC-705; DAS was removed by all three bacteria tested. Binding is postulated as the possible mechanism of the removal, since no difference was observed between the ability of viable and heat-killed bacteria in removing the trichothecenes, and no degradation products of the toxins were detected by gas chromatography (GC)-mass spectrometry (MS) analysis. It is concluded that significant differences exist in the ability of the bacteria to bind trichothecenes in vitro. PMID- 12113666 TI - [Present status and evaluation of interventional therapy for primary hepatocellular carcinoma]. PMID- 12113665 TI - Probabilistic intake assessment and body burden estimation of dioxin-like substances in background conditions and during a short food contamination episode. AB - The objective was to perform a dioxin body burden estimate based on a probabilistic intake assessment of PCDDs, PCDFs and dioxin-like PCBs because of the so-called 1999 'Belgian dioxin incident'. Monte Carlo simulation techniques were used to combine detailed 7-day food intake data on the individual level from a sample of 14-18-year-old adolescents with 'background' and 'incident-related' food contamination data. In background conditions, 3% of the adolescents had an intake <1 pg TEQ kg(-1) bw day(-1), while 85% had <4 pg TEQ kg(-1) bw day(-1). Milk and other dairy products were the basic source of dioxin-like contaminants, while fish constituted the main source at the higher percentiles of intake. During the dioxin incident, the estimated median dioxin intake showed a moderate increase. At the 99th percentile, the highest intake level, and the 95% upper bound uncertainty level, peak body burden-23.73 ng TEQ kg(-1) bw-remained below body burdens that in the experimental animal or in man are accompanied by a population-based observable increase in the incidence of adverse effects. The 1999 Belgian dioxin incident most likely did not affect public health in Belgium in a measurable way, although exceptions remain possible on the individual level. PMID- 12113667 TI - [Prognostic factors influencing survival in patients with large hepatocellular carcinoma receiving combined transcatheter arterial chemoembolization and radiotherapy]. AB - OBJECTIVE: To observe the long-term effects of combined transcatheter arterial chemoembolization (TACE) and radiotherapy for patients with large hepatocellular carcinoma (HCC) and to analyze the prognostic factors. METHODS: A total of 107 patients with large unresectable HCC (the largest diameter of tumor ranged from 5 to 18 cm) were treated with TACE followed by external-beam irradiation. Acute effects and survival rates were observed. The Cox proportional hazards model was used to analyze the prognostic factors. RESULTS: An objective response was achieved in 48.6% of the cases. The cumulative survival rates at 1, 3, and 5 years were 59.4%, 28.4%, and 15.8%, respectively. The tumor number and irradiation dose were the independent prognostic factors. The cumulative survival rates of the patients with a solitary lesion (75.8%, 43.9%, and 26.8% at 1, 3, and 5 years, respectively) were significantly higher than those with multiple lesions (31.3%, and 5.0% at 1 and 3 years, respectively, P=0.0005). The survival rates of the patients received irradiation above 40 Gy (95.8%, 74.7%, and 37.4% at 1, 3, and 5 years, respectively) were significantly higher than those received 20~40 Gy (60.9%, 20.7%, and 10.3%, respectively) and those received radiation lower than 20 Gy (26.7%, 7.1%, and 7.1%, respectively, P=0.0001). CONCLUSIONS: Combined TACE with radiotherapy is a promising treatment for large unresectable HCC. The number of tumor is the most important clinical prognostic factor. Delivering the highest irradiation dose within the tolerance of the liver is the key to improve the long-term effect. PMID- 12113668 TI - [Intratumoral microvessel density and expression of vascular endothelial growth factor in hepatocellular carcinoma after chemoembolization]. AB - OBJECTIVE: To investigate intratumoral microvessel density (MVD) and expression of vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (BFGF) in hepatocellular carcinoma (HCC) after transcatheter arterial chemoembolization (TACE) and to evaluate their significance. METHODS: MVD and expression of VEGF and BFGF in cancerous tissues were examined in forty specimens resected from patients with HCC using immunohistochemical methods. Among these patients, 20 patients received 1 to 7 treatments of TACE prior to II-phase surgical resection (TACE group), the other 20 patients were treated by operation without receiving any other treatment preoperatively (surgical group). There was no significant difference in clinical features between the two groups. MVD was assessed by counting immunostained endothelial cells within a certain area, and staining intensity of VEGF was assessed quantitatively with computer-assisted image analyzer. The expression of BFGF was determined by cell-positive or cell negative. RESULTS: The average MVD was 130.51 75.5 in TACE group and 152.35 58.80 in surgical group. There was no significant difference between the two groups (t= 1.021, P=0.341). Staining intensity of VEGF was 645.60 543.27 in TACE group, higher than in surgical group (158.28 188.48, t=281, P<0.001). BFGF-positive rate was 35% in TACE group and 40% in surgical group. There was no significant difference (x(2)=0.107, P=0.744). CONCLUSIONS: The results indicate that survived cancerous tissue has rich vascularity and the expression of VEGF of the cancerous cells can be enhanced by TACE which may play an important role in reestablishment of blood supply to tumor after TACE. PMID- 12113670 TI - [Localization and distribution of Kupffer cells in hepatocellular carcinoma]. PMID- 12113669 TI - [Hyperthermal lipiodol embolization and thermocoagulation for the treatment of primary hepatocellular carcinoma]. AB - OBJECTIVE: To explore the efficacy of hyperthermal lipiodol embolization and thermocoagulation for the treatment of primary hepatocellular carcinoma. METHODS: One hundred and thirty-one cases were randomized into two groups: the hyperthermal dilute lipiodol embolization group (63 cases) and the chemoembolization group (68 cases). With Seldinger's method, We first placed the catheter to the targeting vessel superselectively and then put the hyperthermal dilute lipiodol (110 degrees C) 10~30ml to the tumor vessels to IV degree for the former group; gave the lipiodol-epirubicin emulsion by the same way to the latter group. RESULTS: The rate of tumor minification and AFP normalization in the hyperthermal lipiodol embolization group was higher than that in the lipiodol epirubicin embolization group. The side effects and the liver damage were mild in the former group. The survival time of the patients in the former group was longer than that in the latter group. CONCLUSIONS: Embolization of the tumor vessels with hyperthermal dilute lipiodol is more thorough due to its better fluidity. The thermocoagulation of the hyperthermal dilute lipiodol becomes stronger for its higher specific heat. It is therefore a good technique for the treatment of primary hepatocellular carcinoma. PMID- 12113671 TI - [Immune responses of dendritic cells after loaded with cytotoxicity T lymphocyte epitope based peptide of human alpha-fetoprotein (hAFP)]. AB - OBJECTIVE: To study the immune responses of lymphocytes after activated by dendritic cells (DCs) loaded with cytotoxicity T lymphocyte (CTL) epitope based peptide of human alpha-fetoprotein (hAFP, 218-226 LLNQHACAV). METHODS: Get high purity DCs by cultured plastic-adherent monocytes isolated from healthy donor of HLA-A2(+) peripheral blood with granulocyte-monocyte colony-stimulating factor (GM-CSF) and interleukin-4 (IL-4) for 7 days. Stimulate self-lymphocytes with DCs that loaded with CTL epitope based peptide of hAFP under the culture medium contains interleukin-2 (IL-2) for 7 days. Analyse IL-12 and TNF in culture medium and also the specific lysis activity of lymphocytes against four strains of primary hepatocellular carcinoma cells. RESULTS: After stimulated by DC loaded with CTL epitope based peptide derived from hAFP, lymphocytes appeared a good characteristics and the culture medium of activated lymphocytes contained a high level Th1 type cytokines of IL-12 and TNF. Activated lymphocytes not only specifically lysed HLA-A2(+) HepG2 line but also had the cytotoxicity against other three primary hepatocellular carcinoma cell lines and T2 target cell loaded with peptide of hAFP. CONCLUSIONS: The results of this research supply the basic materials for the DC based vaccine with HLA-A2 restricted peptide epitope derived from hAFP against AFP positive primary hepatocellular carcinoma. PMID- 12113672 TI - [Effect of glycine on the expression of CD(14) gene and protein of hepatic tissue in the course of developing cirrhosis of rats]. AB - OBJECTIVE: To observe the effect of glycine on the expression of CD(14) mRNA and protein of hepatic tissue in the course of developing cirrhosis of rats. METHODS: The cirrhotic model of Wistar rats was established by complex pathogens, who were respectively fed with control diets and control diets adding glycine (1g/d, giving by intragastric infusion) or 5% glycine containing diets at the same time. The rats were sacrificed at 2, 4, and 8 weeks, respectively. Hepatic tissues were collected to measure the expression of CD(14) mRNA and CD(14) protein by the reverse transcriptase-polymerase chain reaction (RT-PCR) and western blot analysis. RESULTS: The expression of CD(14) mRNA and CD(14) protein in the hepatic tissue of fatty liver and cirrhotic rats fed with diets containing glycine was weaker than their control groups, and the expression of CD(14) mRNA and protein was the weakest in 8 weeks cirrhotic rats fed with the diets. CONCLUSIONS: Glycine can markedly downregulate the expression of CD(14) mRNA and CD(14) protein in hepatic tissues of cirrhotic rats. PMID- 12113673 TI - [Establishment and optimization of rat models for hepatic oval cells proliferation]. AB - OBJECTIVE: To establish a rat model for hepatic oval cell proliferation and to observe the relationship between 2-acetaminofluorene (AAF) dosage and oval cell proliferation in the rat liver. METHODS: Male Wistar rats weighing 150 g received daily oral gavage of AAF for 4 days before operation and up to 7 days after operation. Two-thirds hepatectomy was performed on the 5th day and the gavage was not performed on the day of operation. AFF was given with the dosage of 2.5 mg/kg, 5 mg/kg, 10 mg/kg, 15 mg/kg, and 20 mg/kg body weight. Animals in control group were given saline. Three rats from each group were killed every 2~3 days after hepatectomy and liver slices were fixed and processed for routine histology and immunohistochemistry. RESULTS: Hepatic oval cells were not observed in the liver of controls and only a few were detected in the liver of 2.5 mg/kg and 5 mg/kg groups. However, obvious oval cell proliferation was seen in the liver of 10 mg/kg, 15 mg/kg, and 20 mg/kg groups. Hepatic oval cells were stained positive for cytokeratin 19, OV6, vimentin and proliferating cell nuclear antigen (PCNA). CONCLUSIONS: Satisfactory rat models for hepatic oval cell proliferation can be obtained using our scheme when AAF is dosed at 10~20 mg/kg body weight. PMID- 12113674 TI - [Study on inhibiting factor of Na(+)-K(+)ATP enzyme in fulminant hepatitis]. PMID- 12113675 TI - [Quantitation of alpha-fetoprotein messenger RNA in peripheral blood of nude mice and its relationship with tumor recurrence and metastasis after curative resection of hepatocellular carcinoma]. AB - OBJECTIVE: To assess the level of alpha-fetoprotein (AFP) messenger RNA (mRNA) in peripheral blood of nude mice, and to study its relationship with tumor recurrence and metastasis after curative resection of hepatocellular carcinoma (HCC). METHODS: The metastatic model of human HCC in nude mice LCI-D20 was used in this study. Curative resection was performed at 10th day after tumor implantation in 28 nude mice. Interferon alpha-1b (IFN alpha-1b) was administered subcutaneously from the next day after resection, at doses of 3 10(7)U/kg/d (8 nude mice), 1.5 10(7) U/kg/d (8 nude mice) respectively in the treatment groups, and normal saline alone in the controlled group (12 nude mice). Thirty-five days after treatment, one milliliter of peripheral blood was taken and AFP mRNA was quantitatively analyzed using TaqMan real-time quantitative RT-PCR. The mice were sacrificed. The size of recurrent tumor was measured and the presence of lung metastases was observed. RESULTS: The liver recurrent rate, lung metastatic rate and positivity of AFP mRNA in the controlled group were all 100% (12/12), whereas it was 62.5% (5/8), 0% (0/8), 87.5% (7/8) respectively in the IFN alpha-1b 1.5 10(7)U/kg/d treated group. The recurrent tumor in liver of the IFN alpha-1b 1.5 10(7)U/kg/d treated group was much smaller than that of the controlled group (25 mm(3) 2 mm(3) vs 1143 mm(3) 3 mm(3), t =9.27, P<0.01), and the level of AFP mRNA was also lower than that of the controlled group [(85 6)copies/mug vs (955 2) copies/mug, t =4.33, P<0.01]. In the IFN alpha-1b 3 10(7)U/kg/d treated group, there was only one recurrent tumor (0.5 mm(3)), no lung metastasis, and the positivity of AFP mRNA was 0% (?(2)=11.67, P<0.01). CONCLUSIONS: These results suggest that the level of AFP mRNA in peripheral blood may indicate recurrence and/or metastasis after curative resection of HCC. TaqMan real time quantitative RT-PCR is a very sensitive and convenient method for detecting circulating cancer cells. PMID- 12113676 TI - [Clinical characteristics and psychotropic therapy of depression and anxious symptoms of patients with HBV infection]. PMID- 12113677 TI - [Prophylactic and therapeutic effect of oxymatrine on D-galactosamine-induced rat liver fibrosis]. AB - OBJECTIVE: To investigate the prophylactic and therapeutic effect of oxymatrine on experimental liver fibrosis and to reveal its mechanism. METHODS: By establishing D-galactosamine-induced rat liver fibrosis model, we observed the effect of oxymatrine on serum and tissue biochemical indexes, content of liver hydroxyline, expression of TGF?1 mRNA and changes of tissue pathology. RESULTS: There was a decline of liver hydroxyline and serum AST and ALT in oxymatrine group compared to those of the D-GalN group. The hydroxyline content in oxymatrine pretreatment group was (0.50 0.11)mug/mg compared with (0.99 0.14)mug/mg in D-GalN group (t=8.366, P<0.01). The content in oxymatrine treatment group was (0.44 0.04)mug/mg compared with 0.70 0.06 in D-GalN group (t=9.839, P<0.01). The SOD activity was (149.81 15.28) NU/mg in oxymatrine pretreatment group and (95.22 16.33) NU/mg in the model group (t=7.309, P<0.01); (157.68 19.54) NU/mg in the treatment group compared with (119.88 14.94) NU/mg in the model group (t=4.348, P<0.01). MDA in the pretreatment group was (2.06 0.17) nmol/mg, lower than (4.57 0.37) nmol/mg in the model group (t=17.529, P<0.01). In the treatment group, it was (1.76 0.24)nmol/mg, lower than (3.10 0.17) nmol/mg in the model group (t=12.697, P<0.01). TGF?1 mRNA reduced in the pretreatment and treatment groups as compared with that in the model group (0.21 0.01 vs 0.50 0.01, t=48.665, P<0.01; 0.18 0.02 vs 0.38 0.01, t=22.464, P<0.01). Electron microscopy showed that oxymatrine group had milder hepatocyte degeneration and less fibrosis accumulation than did the model group. Microscopy revealed wide septa expansion from the portal area to the central venous, piecemeal and confluent necrosis and pseudo-nodular formation in part of the lobular in the model group. While in oxymatrine group these lesions were much improved. CONCLUSIONS: Oxymatrine shows prophylactic and therapeutic effect in D galactosamine induced rat liver fibrosis. This is partly by protecting hepatocyte and suppressing fibrosis accumulation through anti-lipoperoxidation. PMID- 12113678 TI - [Effect of compound 861 on tissue inhibitor of metalloprotenase 1 gene expression of HSC-T6 cells]. AB - OBJECTIVE: To observe the in vitro effect of compound 861 (Cpd 861) on tissue inhibitor of metalloprotenase 1 (TIMP1) mRNA levels of HSC-T6 cell. METHODS: HSC T6 cells were exposed in different concentrations of Cpd 861 (0.25~1.0 mg/ml) for 48 hours. The TIMP1 level was measured by the quantitative reverse-transcription polymerase chain reaction (RT-PCR). RESULTS: The TIMP1 mRNA levels of HSC-T6 cells at different concentrations of Cpd 861 were lower (2.50 0.71, 0.50 0.01, 0.11 0.03) than those of the normal control (3.78 0.67, P<0.05 or P<0.01). CONCLUSIONS: The antifibrotic mechanism of Cpd 861 is partly due to its downregulation on TIMP1 mRNA levels of HSC-T6 cells. PMID- 12113679 TI - [Effects of genistein on the fenestrae, proliferation and nitric oxide synthesis of liver sinusoidal endothelial cells from carbon tetrachloride-induced experimental hepatic fibrosis rats]. AB - OBJECTIVE: To investigate the effects of genistein, a tyrosine protein kinase inhibitor, on the fenestrae, proliferation and nitric oxide (NO) synthesis of the liver sinusoidal endothelial cells from CCl(4)-induced hepatic fibrosis rats in vitro. METHODS: By in situ collagenase perfusion and two-step percoll gradient centrifugation, SECs were isolated and cultured from normal and CCl(4)-treated Wistar rats. The fenestrae of SECs were observed by the scanning electron microscopy, and the SECs cell proliferation was determined by the MTT assay. The concentrations of NO in the cultured medium of SECs were detected indirectly by measurement of nitrates and nitrites (the stable products of NO) using the nitrate reduction method. RESULTS: Scanning electron microscopic studies revealed that the number of fenestrae in SECs from all stage of hepatic fibrotic rats was decreased markedly as compared with the SECs from normal controls; however, no obvious changes in the fenestrae of SECs were observed after treated with genistein (100 mumol/L) for 24 hours. After treated with 100 mumol/L genistein for 24 hours, the cell proliferation rates of SECs from all stages of hepatic fibrosis were decreased significantly was compared with the control group (P<0.05). The synthesis of NO by SECs from all stages of hepatic fibrosis was markedly lower than those of normal controls. Treatment with 100 mumol/L genistein for 24 hours could increase the synthesis of NO by SECs from the early stage (stage I) of fibrosis; however, this effect of genistein was not observed in SECs from stage II or III of fibrosis at this concentration. CONCLUSIONS: The results suggest that genistein may play an important role in regulating the function of SECs. PMID- 12113680 TI - [Luteolin inhibits proliferation and collagen synthesis of hepatic stellate cells]. AB - OBJECTIVE: To investigate the effect of luteolin on the proliferation and collagen expression of hepatic stellate cells. METHODS: The effect of luteolin on proliferation and collagen synthesis of hepatic stellate cells isolated from the liver of Wistar rats were determined by (3)H-TdR and (3)H-Pro, and procollagen gene expression was also detected by DIG-labeled gene probe and in situ hybridization. RESULTS: The proliferation and collagen synthesis were significantly and dose-dependently inhibited by luteolin when the concentrations reached 10 micromol/L and 20 micromol/L respectively (t=2.542, P<0.05; t=3.650, P<0.01). The type I, III procollagen mRNA expression was decreased by 25 micromol/L luteolin, in which the type I procollagen mRNA was reduced with statistical significance (x(2)=6.850, P<0.01). CONCLUSIONS: Luteolin inhibits the proliferation and collagen expression of hepatic stellate cells in vitro. It may have a preventive or therapeutic role in liver fibrosis. PMID- 12113681 TI - [Expression of lipopolysaccharide binding protein and lipopolysaccharide receptor CD14 in experimental alcoholic liver disease]. AB - OBJECTIVE: To observe the expression of lipopolysaccharide binding protein (LBP) and CD14 mRNA in alcohol-induced liver disease (ALD) and evaluate the relationship between the expression of LBP and CD14 mRNA and the severity of liver injury in alcoholic-fed rats. METHODS: Twenty Wistar rats were divided into two groups: ethanol-fed group and control group. Ethanol-fed group were fed ethanol (by intragastric infusion of 500 ml/L ethanol orally, dose of 5~12 g/kg/d) and control group received dextrose instead of ethanol. Rats of both groups were sacrificed at 4 weeks and 8 weeks, respectively. Levels of endotoxin and alanine transaminase (ALT) in blood were measured, and liver pathology was observed by light and electronic microscopy. Expression of LBP and CD14 mRNA in liver tissues were determined with the reverse transcription polymerase chain reaction (RT-PCR) analysis. RESULTS: Plasma endotoxin levels were increased significantly in ethanol-fed rats [(129 21) pg/ml and (187 35) pg/ml at 4 weeks and 8 weeks] than in control rats [(48 9) pg/ml and (53 11) pg/ml, respectively, t=11.2, 11.6, P<0.05]. Mean values for plasma ALT levels were increased dramatically in ethanol-fed rats after 4 weeks and 8 weeks [(112 15) U/L and (147 22) U/L, respectively] than in the control animals [(31 12)U/L and (33 9)U/L, respectively, t=5.9, 20.6, P<0.05]. In liver sections from ethanol-fed rats, there was marked pathological changes (steatosis, cell infiltration and necrosis). In the control rats, there was no significant difference in the levels of LBP and CD14 mRNA at the two time points. In ethanol-fed rats, ethanol administration led to a significant increase in LBP and CD14 mRNA levels as compared with the control group (P<0.05). CONCLUSIONS: Ethanol administration lead to a significant increase in endotoxin levels of the serum and LBP and CD14 mRNA expression in liver tissues in ethanol- fed rats when compared with the control rats. Increase of LBP and CD14 mRNA expression may result in greater sensitivity to endotoxin and thus lead to liver injury. PMID- 12113682 TI - [HCV RNA assessment by PCR technique for screening post-transfusion HCV infection among blood donors]. AB - OBJECTIVE: To survey the application of PCR for screening HCV RNA from blood donations within the window period. METHODS: According to a standardized method, 12 blood banks organized by the National Center for Clinical Laboratories collected and prepared about ten thousands specimens. The specimens were tested with two different kits. RESULTS: Among the 7173 specimens A group, 21 were PCR positive for HCV RNA. The positive rate was 0.29%. There were not positive for HCV RNA among 7477 specimens (B group). CONCLUSIONS: It is feasible to use the PCR screening for the detection of HCV RNA of blood donations but is unnecessary to standardize the specimen collection and the kit selection. PMID- 12113683 TI - [Clinical research of patients with acute or chronic hepatic failure treated with molecular adsorbent recirculating system]. AB - OBJECTIVE: To summarize the experience of a single treatment using molecular adsorbent recirculating system (MARS) in patients with acute-on-chronic liver failure. METHODS: Twenty-five eases treated by MARS-artificial liver were followed up and reviewed. RESULTS: The levels of serum total bilirubin, non conjugated bilirubin and blood ammonia were significantly decreased from (618.51 200.68) mmol/L to (390.81 146.02) mmol/L (t=2.729, P<0.01), (490.03 163.39) mmol/L to (303.28 113.06) mmol/L (t =2.516, P<0.01), and (152.44 82.62)mmol/L to (84.80 13.30)mmol/L (t=2.174, P<0.05), respectively. Prothrombin activity was significantly increased from 70.55% 32.39% to 93.63% 14.20% (t=1.728, P<0.05) in patients during a single 6 h treatment with MARS. No difference was presented in the markers of liver zymogram, serum protein, kidney function, electrolyte, blood routine and blood gas analysis before and after the MARS. Thirteen of 17 patients have been cured or improved, 4 died, and the survival rate was 76.5%. CONCLUSIONS: MARS is a safe and an effective treatment for patients with liver failure. PMID- 12113684 TI - [Effects of lamivudine and thymosin alpha1 combination therapy on patients with chronic hepatitis B]. PMID- 12113685 TI - [Detection and clinical significance of serum antimitochondrial antibody in patients with viral hepatitis or primary biliary cirrhosis]. PMID- 12113686 TI - [Relationship between serum TGF beta-1 with chronic hepatitis B involving in liver cell function and liver biopsy fibrosis]. PMID- 12113687 TI - [Antiproliferative effect of IH764-3 derived from Salvia miltiorrhiza in cultured rat hepatic stellate cells. ]. PMID- 12113688 TI - [Fulminant viral hepatitis: mice model and its clinical implications]. PMID- 12113689 TI - [Effects of nuclear transcriptional factors on hepatic stellate cells' activation]. PMID- 12113690 TI - [The role of hepatic sinusoids in local control of the immune response]. PMID- 12113691 TI - [Changes of GR and HSP70 and their relationships with secondary hepatic damage after severe trauma]. PMID- 12113692 TI - [Clinical application of molecular adsorbent recirculating system-artificial liver support system]. PMID- 12113693 TI - [Hepatic stem cells in liver diseases]. PMID- 12113694 TI - [Progress in the research on the HBV mutants associeted with lamivudine therapy]. PMID- 12113695 TI - [Platelet antiaggregation treatment in the aftermath of GUSTO IV, TARGET, TACTICS, and CURE trials]. AB - Various clinical studies have shown that the risk associated with acute coronary syndrome can be reduced by pharmacological treatment with platelet glycoprotein IIb-IIIa receptor inhibitors. The treatment with these drugs is beneficial in conjunction with conventional medical treatment, during angioplasty and in patients undergoing aortocoronary bypass, as indicated by the results of the CAPTURE, PRISM, and PURSUIT studies. However, shortly after the European Society of Cardiology recommended systematic use of glycoprotein IIb-IIIa inhibitors in the management of high-risk patients, the report of negative results in the GUSTO IV and TARGET trials caused this position to be reconsidered. The need for studies to better characterize the mechanism of action of these antithrombotic agents is evident. Studies in vitro have demonstrated that prolonged treatment with these drugs can cause patients to remain outside the therapeutic range by causing platelet receptors to change configuration and have more affinity for fibrinogen. In view of these findings, the results of the GUSTO IV and TARGET studies, far from demonstrating the ineffectiveness of the antiplatelet aggregation agents, could be interpreted as a failure in the design of the therapeutic strategy. On the other hand, the results of the CURE study provide new evidence of the benefits of the use of clopidogrel in patients with acute coronary syndrome undergoing medical or revascularization treatment (PCI-CURE group). The debate about the usefulness of platelet glycoprotein IIb-IIIa inhibitors, alone or in combination with clopidogrel in certain circumstances remains open. The evidence available to date is inconclusive and the guidelines for the management of patients with acute coronary syndrome should be updated. PMID- 12113696 TI - [Direct stenting: the decline of balloon?]. PMID- 12113697 TI - [Direct Stenting without Predilation: a Single-Center Experience with 1,000 Lesions]. AB - INTRODUCTION: Direct stenting has been shown to save costs, procedural time, radiation, and contrast use. We analyze the results of direct stenting in daily practice. MATERIAL AND METHODS: We retrospectively analyzed the interventions in the first 1,000 lesions that were treated with direct stenting at our center. Primary success, dissection, need for additional dilation, embolism, stent loss, and side branch occlusion were the variables assessed. RESULTS: Direct stenting was attempted in 1,000 lesions in 784 patients (age 63 11 years, females 21%, diabetes 37%). Primary or rescue angioplasty was performed in 8%. One or more thrombi were found in 16%, bifurcation in 9%, calcification in 5%, angulation in 2.3%, and tortuosity in 3.2%. The reference diameter was 3.0 0.5 mm. The primary success rate was 93.1%. Failure of direct stenting (6.9%) was associated with the circumflex artery in 38%, calcification in 26%, angulation in 22%, and tortuosity in 31%. In 39 lesions, additional dilation with different balloons was required. Additional stenting was required for dissection in 40 lesions and secondary to incomplete coverage of the lesion in 27. Thrombus embolism occurred in 7 lesions, 6 of them with a previously visible thrombus and one in a vein graft. Stent embolisms occurred in 6 cases, 4 of which were retrieved. Four side branches became occluded, but 2 of them were recovered at the end of the procedure. CONCLUSIONS: Direct stenting is a safe technique with low percentage of dissection, need for postdilation, thrombus embolism, and side br PMID- 12113698 TI - [Head-up tilt test potentiated with nitroglycerin. What is the optimal duration of the test after administration of the drug?]. AB - INTRODUCTION AND OBJECTIVES: Numerous variations of the head-up tilt-table test potentiated with nitrates have been reported. After the administration of nitroglycerin, between 10 and 25 min of continued tilting have been recommended. The aim of this study was to assess the optimal duration of the pharmacological phase of the head-up tilt-table test potentiated with sublingual administration of nitroglycerin spray (NTG-TT). METHOD: The records of 498 consecutive NTG-TT were reviewed. Our protocol consisted of a 20-min drug-free phase at a 60o angle. If syncope does not develop, 400 microgram of sublingual nitroglycerin spray is administered and the patient continues to be tilted for a further 25 min. The test results and time to a positive response were analyzed. RESULTS: The result of NTG-TT was positive in 288 procedures, most of them after nitroglycerin administration (255, 88.5%). The mean time to a positive response was 10.7 6.7 and 5.0 2.4 min during the control and pharmacological phases respectively. Most positive responses were concentrated in the 3 to 5 min after drug administration. The time to syncope after nitroglycerin administration was over 10 min in 9 patients and 15 min in only 2 patients. CONCLUSIONS: The duration of the pharmacological phase of NTG-TT using the described protocol can be reduced to 15 min without loss of sensitivity. A further reduction to 10 min would decrease the rate of positive responses by a small amount and might be considered clinically acceptable. PMID- 12113699 TI - [Indications for catheter ablation and results in Andalusia, Spain]. AB - INTRODUCTION AND OBJECTIVES: We report the results of the first Catheter Ablation Registry of the Arrhythmia Working Group of the Andalusian Society of Cardiology (AWGASC) for 2000. METHODS: The register includes information about the ablation procedures performed in 2000, which was collected retrospectively and submitted voluntarily by four out of six cardiac electrophysiology laboratories of the AWGASC. A total of 424 patients (mean age 45 18 years; 50% men) were included. Twelve patients underwent two different ablation procedures, bringing the total number of procedures to 436. The overall success rate (based on current criteria), success rate by procedure, in-hospital mortality, and major complications are reported. RESULTS: The type and distribution of the ablation procedures were atrioventricular nodal re-entry tachycardia ablation, 34%; accessory pathway ablation, 39%; ventricular tachycardia ablation, 8%; atrial tachycardia ablation, 3%; atrioventricular junctional ablation, 9%, and cavo tricuspid isthmus ablation, 9%. The overall success rate was 94% (range 97.8% to 87.4% in different laboratories), rate of major complications 1.1% (range 0% to 3.7%), and overall mortality 0.23% (1 patient). CONCLUSIONS: These findings summarize the indications and results of catheter ablation procedures performed in 2000 at four cardiac electrophysiology laboratories in Andalusia. This is the first multicenter registry in Spain. PMID- 12113700 TI - [Familial dilated cardiomyopathy in patients transplanted for idiopathic dilated cardiomyopathy]. AB - OBJECTIVE: To evaluate the prevalence, clinical features, and pattern of inheritance of familial dilated cardiomyopathy (DCM) in heart transplant patients. PATIENTS AND METHOD: Patients with idiopathic DCM who had undergone heart transplantation were invited to participate. Patients with alcohol abuse were excluded. A clinical evaluation, 12-lead ECG, echocardiogram, blood tests, and DNA extraction were performed in patients and relatives. Familial DCM was defined as the presence of at least one relative with idiopathic DCM. Possible familial DCM was considered when at least one relative had left ventricular enlargement (LVE) (> 112% predicted LVEDD). RESULTS: One hundred and ninety-nine relatives of 43 families were studied. DCM was familial in 11 probands (25.6%) and possibly familial in 11 (25.6%). Fifteen relatives had DCM (7.5%), 26 (13.1%) LVE, and 5 (2.5%) hypertrophic cardiomyopathy. The pattern of inheritance was autosomal dominant in most families. Five probands (3 with familial DCM) had antecedents of consanguinity and possible recessive inheritance. Six probands (14%, 1 with familial DCM) had relatives with conduction system defects. Creatine kinase was moderately increased in 9 relatives (4.5%), 3 of them with LVE. Fifteen patients had at least moderate alcohol intake. Three of them had familial DCM (relatives without alcohol abuse) and 6 had possible familial DCM. CONCLUSIONS: The prevalence of familial DCM is high in patients who undergo heart transplant. Left ventricular enlargement, conduction system abnormalities, and elevated creatine kinase may be early markers of familial disease. Hypertrophic cardiomyopathy is present in some relatives of patients with idiopathic DCM. Familial DCM is present in patients with a previous diagnosis of alcoholic DCM. PMID- 12113701 TI - [A comparative study of the follow-up and hemodynamics in vivo of 21 mm Carpentier-Edwards supra-annular and perimount bioprostheses]. AB - INTRODUCTION AND OBJECTIVES: Analysis and comparison of the clinical performance and hemodynamics in vivo of 21 mm Carpentier-Edwards supra-annular (CESA) and Perimount (CEPM) aortic bioprostheses. METHODS: A follow-up study was made of 40 patients implanted a 21 mm CESA (n = 21) or CEPM (n = 19) prosthesis between October 1992 and September 1997. All eligible survivors (14 CESA, 12 CEPM) were assessed echocardiographically. RESULTS: There were no significant differences between models in the effective orifice area (1.6 cm2 for CESA, 1.44 cm2 for CEPM), peak flow rate (rest: 2.5 m/s for CESA, 2.3 m/s for CEPM; post-dobutamine: 3.4 m/s for CESA, 3.3 m/s for CEPM), mean flow rate (rest: 1.7 m/s for CESA, 1.6 m/s for CEPM; post-dobutamine: 2.5 m/s for CESA, 2.2 m/s for CEPM), peak gradient (rest: 28.3 mmHg for CESA, 21.6 mmHg for CEPM; post-dobutamine: 48.4 mmHg for CESA, 41.6 mmHg for CEPM), and mean gradient (rest: 15.8 mmHg for CESA, 12.0 mmHg for CEPM; post-dobutamine: 28.5 mmHg for CESA, 22.5 mmHg for CEPM). CONCLUSION: In our experience, these two prosthetic models have similar hemodynamic characteristics in small aortic annuli. PMID- 12113702 TI - [Interleukin-10 and coronary disease]. AB - Understanding of the pathophysiology of atherosclerosis has changed markedly over the past few decades. It is now widely accepted that inflammation plays a fundamental role in the genesis and development of atherosclerosis. Inflammatory mechanisms also appear to determine clinical presentation and disease outcome. Atherosclerotic lesions have high concentrations of inflammatory cells (T lymphocytes and activated macrophages) as well as an abundance of pro inflammatory cytokines [interleukin (IL)-1, IL-6, IL-8, interferon-gamma, tumor necrosis factor-alpha, etc.] that modulate local inflammatory responses. These may also alter plaque stability and facilitate the development of acute cardiovascular events. The role of anti-inflammatory cytokines in this context remains to be studied. IL-10 is an anti-inflammatory cytokine synthesised by T lymphocytes and macrophages and has other anti-inflammatory effects. IL-10 expression within human atherosclerotic plaques has been demonstrated and animal experiments have shown that low levels of IL-10 lead to the development of extensive and unstable atherosclerotic lesions. Currently available evidence suggests a potential protective role for IL-10 in atherosclerosis. This new perspective on coronary disease as a chronic inflammatory process may open new avenues for the management of ischemic heart disease. PMID- 12113704 TI - [Images in cardiology. Cardiac amyloidosis]. PMID- 12113703 TI - [Prognosis of diabetic patients with ischemic cardiopathy]. AB - The prevalence of diabetes in Spain is about 6% and increases with age and obesity. Diabetes is present in approximately 25% of patients with coronary heart disease (CHD). Pre-diabetic and diabetic patients have a higher incidence of CHD and poorer prognosis, with high short- and long-term mortality. The protective effect of pre-menopause status is suppressed by diabetes. Diabetes has a synergic effect with other cardiovascular risk factors. Primary prevention in diabetic patients should be approached as in non-diabetic post-infarction patients. In diabetes, a healthy life-style and strict control of blood sugar and the other cardiovascular risk factors, particularly hypertension, is mandatory. PMID- 12113705 TI - [Angina related to 5-Fluorouracil]. AB - Treatment with 5-fluorouracil is common in oncological patients. Side effects on bone marrow, skin, and mucous membranes have been reported. Cardiotoxicity, which is less predictable, can be life-threatening. Manifestations include angina, arrhythmias, infarction, heart failure and cardiogenic shock. The toxic mechanisms that might be involved have been much discussed but have not yet been clearly established. Current evidence supports the possibility of a metabolic effect in common with the cascade secondary to ischemia due to coronary disease. Based on a case report, we discuss the usual clinical presentation, treatment and prognosis. Finally we make recommendations for managing patients being treated with 5-fluorouracil. PMID- 12113706 TI - [Cardiac injury after indirect lightning strike]. AB - Lightning strike is one of the most frequent causes of death due to natural phenomena. In such cases, cardiac injury is the main cause of death, with type of lesion varying by type of impact. We report the case of a 29-year-old woman who was struck indirectly by lightning. Upon hospital admission, she showed both the echocardiographic disturbances characteristic of direct impact and electrocardiographic disturbances. Both types of change resolved spontaneously. After describing the case, we briefly review the literature on echo and electrocardiographic disturbances after lightning strike. PMID- 12113707 TI - [Percutaneous thrombin injection for closure of femoral pseudoaneurysms: preliminary experience]. AB - An arterial pseudoaneurysm is an uncommon complication of cardiovascular procedures associated with considerable morbidity and increased hospital costs. Percutaneous thrombin injection is one approach to therapy. We describe our initial experience with this technique in 3 patients, with special attention to the utility of sonographic guidance. In all cases complete closure was achieved, although one patient required additional brief extrinsic compression with the ultrasound probe. PMID- 12113708 TI - [Acute pulmonary edema with normal coronary arteries: mechanism identification by ergonovine stress echocardiography]. AB - Coronary spasm is a constriction of the epicardial coronary arteries that produces myocardial ischemia. It is considered the main mechanism of the dynamic coronary artery stenosis. The standard method for diagnosing coronary spasm is the ergonovine test during diagnostic coronary angiography. Another test currently used is stress echocardiography with intravenous ergonovine injection. We present the case of a patient with angina, acute pulmonary edema and normal angiographic coronary arteries in which stress echocardiography with ergonovine demonstrated transient severe mitral regurgitation. PMID- 12113709 TI - [Coronary Spasm after Administration of Propranolol during Dobutamine Stress Echocardiography]. AB - Dobutamine stress echocardiography, a highly useful and safe challenge test for myocardial ischemia, is being used increasingly. We report the case of a 37-year old man with rest angina, repolarization abnormalities in precordial leads and normal coronary arteries who was referred for dobutamine-atropine stress echocardiography, which was negative for ischemia. However, after testing, upon injection of propranolol, the patient suffered chest pain associated with ST elevation and severe regional systolic abnormalities. After intravenous nitroglycerin administration, chest pain and electrocardiographic abnormalities disappeared quickly, and systolic motion became normal. This complication was interpreted as a coronary spasm. We discuss the causes for the spasm and the role that might have been played by the drugs employed. PMID- 12113710 TI - [Why was our manuscript rejected?]. PMID- 12113712 TI - [Heart failure: from clinical trials to everyday clinical practice]. PMID- 12113713 TI - [Primary angioplasty: this balloon is of general interest, indeed]. PMID- 12113714 TI - [Dobutamine instead of exercise stress test to detect coronary artery disease. Is electrocardiographic information enough?]. PMID- 12113715 TI - [Baseline characteristics and determinants of outcome in a patient population admitted for heart failure to a general hospital]. AB - INTRODUCTION AND OBJECTIVES: To assess baseline characteristics, management patterns, and clinical outcomes after 18 months in patients diagnosed as heart failure in a tertiary hospital in Catalonia, Spain. METHODS: The records of all 265 patients admitted to the Hospital General Vall d'Hebron from July through December 1998 with a diagnosis of heart failure who met study criteria were identified and analyzed. Patients were interviewed by telephone 18 months later. RESULTS: The mean age of the study population was 75 12 years, 42% were male, 19% were admitted for causes other than heart failure, and 62% had significant comorbidity. Ventricular function was assessed in 68% (preferentially patients with a better prognosis), and was considered normal in 41%. Angiotensin converting enzyme inhibitors or angiotensin II antagonists were used in 54%, and beta-blockers in 4%. The 18-month mortality was 46% (77% cardiac mortality). Multivariate predictors of death were older age, severe or previous heart failure, and serious comorbidity. At 18 months, 69% of survivors were in functional classes I or II. CONCLUSIONS: 1) As in other geographic areas, patients in this study were an older population with poor survival; 2) local patterns of care definitely need improvement; 3) comorbidity is important for prognosis, and 4) a significant proportion of survivors enjoy an acceptable quality of life long after discharge. PMID- 12113716 TI - [Hospitalized congestive heart failure patients with preserved versus abnormal left ventricular systolic function]. AB - OBJECTIVES: To compare the clinical characteristics of hospitalized patients with congestive heart failure and left ventricular dysfunction versus normal systolic function. METHODS: Clinical records of all admissions with a heart failure diagnostic code over a one-year period were reviewed retrospectively. Of 1,953 admissions, 595 were excluded because they did not fulfill diagnostic criteria. RESULTS: A total of 1,069 patients had 1,358 admissions with confirmed heart failure (1.27 admissions/patient). Of them, 706 patients (66%) had an echocardiographic study and 381 (54%) had ventricular dysfunction. Ventricular dysfunction was associated with previous myocardial infarction (OR = 5.8), left bundle-branch block (OR = 5.0), male sex (OR = 2.0), and smoking (OR = 1.8). Meanwhile, a negative association existed with age (OR = 0.97), previous valve surgery (OR = 0.46) and atrial fibrillation (OR = 0.49). Patients with ventricular dysfunction had more hospitalizations in the cardiology department and received more vasodilators, aspirin, and nitrates on discharge. The prescription of angiotensin converting enzyme inhibitors prescription to patients with ventricular dysfunction increased with the severity of ventricular dysfunction and was more frequent in patients admitted to the cardiology department. Systolic dysfunction increased hospital mortality (OR = 2.9). CONCLUSIONS: Patients admitted with heart failure and systolic dysfunction had a different clinical profile than patients with a normal ejection fraction. Seven clinical variables predicted the presence of systolic dysfunction. Patients with ventricular dysfunction had more hospital mortality and were prescribed vasodilators, aspirin, and nitrates more often on discharge. PMID- 12113717 TI - [Program of coronary angioplasty in acute myocardial infarction in the region of Murcia (Spain). APRIMUR Registry]. AB - INTRODUCTION AND OBJECTIVES: The geographic characteristic and healthcare facilities of the region of Murcia, Spain, are enough to assure that coronary angioplasty can be carried out in acute myocardial infarction according to current guidelines. The development of a regional program for coronary intervention in acute myocardial infarction may increase the number of patients who would benefit from reperfusion therapy in general and primary angioplasty in particular. MATERIAL AND METHODS: The program was initiated in April 2000 and had four steps: 1) Establishment of primary angioplasty as the treatment of choice of acute myocardial infarction in the regional reference hospital. 2) Application of phase 1 to a second hospital located 10 kilometers away from the reference hospital. 3) Extension of phase 1 to the entire city of Murcia. 4) Provision of facilities for coronary angioplasty in acute myocardial infarction to all patients in the region. RESULTS: Between January 2000 and August 2001, 392 angioplasties were performed for acute myocardial infarction. Primary angioplasty was performed in 92% and 85% of the patients with an indication for reperfusion therapy in phase 1 and 2, respectively. The median delay (indication to beginning of procedure) was 25 and 35 minutes in phases 1 and 2, respectively. Total mortality was 11,5% (5,2% in patients without shock at admission). CONCLUSIONS: The design of a regional program of primary angioplasty may increase the number of patients who receive reperfusion therapy in compliance with current recommendations for the treatment of acute myocardial infarction. PMID- 12113718 TI - [Time intervals in primary angioplasty from onset of symptoms until restoration of blood flow]. AB - INTRODUCTION AND OBJECTIVE: A limitation to the widespread use of primary angioplasty is delayed reperfusion. Most current data are from clinical trials and there is little information about the use of primary angioplasty in clinical practice. The objective of this study was to analyze the duration of each stage leading to primary angioplasty in a hospital where it is the treatment of choice for acute myocardial infarction. PATIENTS AND METHOD: Prospective observational study of patients admitted to our hospital from April 2000 to August 2001 for acute myocardial infarction with an indication for reperfusion. The time intervals from onset of symptoms until the end of angioplasty were analyzed. RESULTS: Primary angioplasty was performed in 201 of 218 patients with an indication for reperfusion (92%). Median values (percentiles 25-75) were: Time 1 (onset of symptoms-hospital arrival): 91 (50-150) minutes. Time 2 (hospital arrival-call to interventional team): 20 (10-49) minutes. Time 3: (call to interventional team-team arrival): 15 (0-20) minutes. Time 4: (team arrival patient arrival at the catheterization laboratory): 10 (5-15) minutes. Time 5 (patient arrival-opening of coronary artery): 20 (15-30) minutes. Time 6 (opening of coronary artery-TIMI III flow): 10 (0-25) minutes. CONCLUSIONS: The most time consuming stage in primary angioplasty was from the onset of symptoms until patient arrival at the hospital (Time 1). Inside the hospital, the most time consuming stage was the diagnosis and decision to perform angioplasty (Time 2). The rates of primary angioplasty could be increased if delays in reperfusion were reduced with respect to those considered acceptable in current practice guidelines. PMID- 12113719 TI - [Long-term follow-up of patients with proximal left anterior descending coronary artery stenosis treated with stent]. AB - INTRODUCTION AND OBJECTIVE: Patients with lesions of the proximal left anterior descending coronary artery are a special high-risk group. In the present study we analyzed the efficacy and safety of coronary stenting in such lesions and the factors related to a less favorable prognosis in long-term follow-up. METHODS: Ninety-eight consecutive patients with severe left anterior descending artery stenosis were enrolled, all with coronary angioplasty and elective stenting. Clinical follow-up was carried out annually in all patients by personal interview or telephone contact. The incidence of death, new infarction, anginal status, and new revascularization procedures was registered. Clinical, angiographic, and procedural variables were analyzed to identify predictors of long term prognosis. RESULTS: Mean follow-up was 38 11 months. There was only one major periprocedural complication, which required urgent surgery. Five deaths were registered, 3 of non-cardiac and 2 of cardiac origin. Twenty-five patients developed angina and 11 underwent a new revascularization of the proximal left anterior descending coronary artery (6 surgical and 5 angioplasty). Two patients had new anterior myocardial infarction. At 60 months the major cardiac event-free rate was 83.7% and the cardiac death-free rate was 98%. The use of two stents and the association of diabetes-hypertension-hypercholesterolemia were associated with a less favorable prognosis in our population. CONCLUSIONS: Stenting of left anterior descending coronary stenosis was safe and effective in a long-term analysis. The survival rate was high and the incidence of new revascularization was low. PMID- 12113720 TI - [Dobutamine stress test. Specificity and sensitivity of continuous ST-segment monitoring in 12 simultaneous standard ECG leads]. AB - INTRODUCTION AND OBJECTIVES: This study was designed to evaluate the specificity and sensitivity of the dobutamine stress test with continuous ST-segment monitoring in 12 standard leads. PATIENTS AND METHOD: We analyzed 75 patients, 36 with unstable angina, 22 post-myocardial infarction angina, 3 after successful angioplasty, 6 chronic stable angina, and 8 atypical chest pain (with normal coronary arteries). All of them underwent coronary angiography (coronary lesions were considering significant with > 70%). Beta-blocking agents, calcium antagonists, and nitrates were discontinued for 48 h before the test. A 12-lead ECG was acquired automatically and ST-segment changes were monitored. Dobutamine infusion started at 5 microgram/kg/min and was increased at 3-minute intervals to 10 microgram, 20 microgram, and a maximum of 40 microgram/kg/min. If heart rate did not reach 85% of the theoretical maximum, 0.5-1 mg atropine was given without discontinuing dobutamine infusion. RESULTS: Fifty patients (67%) had abnormal coronary arteries (excluding vessels with 100% obstruction) and 25 patients (33%) had normal arteries. In the group of patients with coronary lesions, the test was abnormal in 90% and normal in 10%. In the group of patients with normal arteries, the test was abnormal in 16% and normal in 84%. Consequently, the test had 90% sensitivity, 84% specificity, and 10% false negative and 16% false positive results. CONCLUSIONS: Our results showed that the dobutamine/ECG test was a simple, effective, and safe bedside tool for diagnosing the severity of coronary disease. PMID- 12113721 TI - [Prognostic value of fibrinogen in patients admitted with suspected unstable angina and non-q-wave myocardial infarction]. AB - INTRODUCTION AND OBJECTIVE: In recent years, the relation between biological markers of inflammation and prognosis in patients suffering from acute coronary syndromes has been investigated. The aim of this study was to evaluate the association between baseline fibrinogen concentrations and the development of clinical events in patients admitted with suspicion of unstable angina and non-Q wave myocardial infarction. MATERIAL AND METHOD: Levels of fibrinogen at enrollment were analyzed in 325 consecutive patients with acute coronary syndromes. Fibrinogen values were divided into tertiles and the incidence of clinical events was evaluated at each level. The combination of death and/or myocardial infarction was the main endpoint. RESULTS: Fibrinogen levels were significantly higher in patients who subsequently had myocardial infarction, cardiac death, or both during follow up. The probabilities of death and/or myocardial infarction were 6%, 13%, and 29% (p < 0.0001), respectively, in patients grouped by fibrinogen tertiles (304, 305-374 and 375 mg/dl). Multivariate predictors of combined events were age, previous angina, ST-segment depression in the admission ECG, and fibrinogen into tertiles. The adjusted hazard ratio (95% CI) for patients in the upper tertile was 4.8 (1.6-14; p = 0.004). CONCLUSIONS: High fibrinogen levels were related to a less favorable long term or short-term outcome in patients admitted for suspicion of unstable angina and non-Q-wave myocardial infarction. This association persists after adjustment for other classical risk factors such as age, prior angina, and ST-segment depression in the ECG. PMID- 12113722 TI - [2002 Update of the Guidelines of the Spanish Society of Cardiology for Unstable Angina/Without ST-Segment Elevation Myocardial Infarction]. AB - Since the Spanish Society of Cardiology Clinical Practice Guidelines on Unstable Angina/Non-Q-Wave Myocardial Infarction were released in 1999, the conclusions of several studies that have been published make it advisable to update current clinical recommendations. The main findings are related to the developing role of Chest Pain Units in the management and early risk stratification of acute coronary syndromes in the emergency department; new information concerning the efficacy of glycoprotein IIb/IIIa inhibitors, clopidogrel and low-molecular weight heparins in the pharmacological treatment of acute coronary syndromes without ST-segment elevation; and the role of early invasive strategy in improving the prognosis of these patients. The published evidence is reviewed and the corresponding clinical recommendations for the management of acute coronary syndromes without persistent ST-segment elevation are updated. PMID- 12113724 TI - [Helicobacter pylori: a new cardiovascular risk factor?]. AB - There is increasing evidence that certain microbial agents may have an etiopathogenic role in the development of atherothrombosis. Helicobacter pylori, a bacterium that causes peptic ulcer disease, has been suggested as one of the microbes involved in the development of atherothrombosis. This hypothesis is based on the following observations: a) a higher prevalence of Helicobacter pylori infection in patients with coronary artery disease, myocardial infarction, or cerebrovascular disease; b) the coincidence of Helicobacter pylori infection and cardiovascular risk factors, such as serum cholesterol and triglyceride concentrations and plasma fibrinogen; c) Helicobacter pylori seropositivity correlates with acute-phase proteins associated with higher risk of coronary disease, such as C-reactive protein, and d) controversial PCR studies indicating the presence of Helicobacter pylori in atheromas. Analysis of the scientific evidence suggests that Helicobacter pylori infection could indirectly contribute to the development and severity of atherothrombosis and cardiovascular disease. PMID- 12113725 TI - [Epidemiology of diabetes and its non-coronary complications]. AB - Diabetes mellitus is among the diseases with great impact on health and society, not only for its high prevalence but also for its chronic complications and high mortality. The most precise method to investigate the prevalence of diabetes is by oral glucose tolerance testing. In Spain, the prevalence of diabetes in the 30 65 year-old population is estimated to be 6.5% among 30-to-65- year old, and 10.3% among the 30-to-89 year-old population. The ratio of known to unknown diabetes ranges from 1:3 to 2:3. The incidence of diabetes mellitus type 2 in Spain is 8/1000 persons per year, and the incidence of type 1 is 11 to 12 cases per 100,000 persons per year. The prevalence of chronic complications varies according to type of diabetes, time since onset and degree of metabolic control: neuropathy 25%, retinopathy 32% and nephropathy 23%. Diabetes is one of the most important causes of death in Spain, occupying third place for women and seventh for men. PMID- 12113726 TI - [Bilateral pulmonary veins atresia]. PMID- 12113727 TI - [Pulmonary hypertension associated with human immunodeficiency virus infection: a review of 4 cases]. AB - Several cardiorespiratory diseases can complicate human immunodeficiency virus (HIV) infection. Primary pulmonary hypertension is a rare clinical disorder with a poor prognosis. We describe this syndrome in four HIV- positive patients who were examined in our hospital. PMID- 12113728 TI - [Giant cell myocarditis. A case report]. AB - Giant cell myocarditis is a rare disease of unknown origin that is probably autoimmune in nature; the prognosis is poor and death often ensues unless a heart transplant is performed. Several cases responding to immunosuppressive therapy have been recently reported, however. We describe a patient who developed fulminant heart failure requiring heart transplantation. Examination of the explanted heart confirmed the diagnosis of giant cell myocarditis. PMID- 12113729 TI - [Middle aortic syndrome caused by Takayasu's disease and treated by stent implantation: a report of medium-term follow-up]. AB - A 13-year-old girl with middle aortic syndrome caused by Takayasu's disease was treated by balloon angioplasty of the right renal artery stenosis and the implantation of 3 stents, 2 in the stenosed thoracic segment and 1 in the abdominal segment of the aorta. Spiral computed tomography one and two years after the stents were inserted showed that the disease had progressed despite treatment with immunosuppressants. PMID- 12113730 TI - [Transient ST Elevation: a finding that may be frequent in percutaneous atrial septal defect closure in adults]. AB - Transient ST elevation in inferior leads has been described as a rare complication during percutaneous atrial septal defect closure. We present a series of adult patients who underwent percutaneous atrial septal defect closure with the Amplatzer device and in whom transient ST changes were observed frequently. PMID- 12113731 TI - [Ventricular pacing after atrioventricular node ablation]. PMID- 12113733 TI - [What is an efficient health technology in Spain?]. AB - INTRODUCTION: Despite the growing recognition of the potential applications of cost-effectiveness assessments, a criterion to establish what is an efficient health technology does not exist in Spain. The objective of this work is to describe the limits and the criteria used in Spain to recommend the adoption of health interventions. METHOD: A review of the economic evaluations of health technologies published in Spain from 1990 to 2001 was conducted. Complete economic assessments in which the cost-effectiveness ratio was expressed as cost per life-year gained (LYG), cost per quality-adjusted-life-year (QALY) or cost per saved live were selected. Those interventions in which the authors established recommendations (adoption or rejection) and the criteria used were analyzed. RESULTS: Twenty (20%) of the 100 complete economic evaluations fulfilled the selection criteria. In16 studies, the results were expressed as cost per LYG, in 6 studies as cost per QALY and in 1 as cost per saved live. A total of 82 health interventions were assessed and some kind of recommendation was established in 44 of them. All technologies with a cost-effectiveness ratio lower than 30,000 euros (5 million pesetas) per LYG were recommended for adoption by the authors. Up to that limit there was no a clear tendency. CONCLUSIONS: Although the results must be interpreted with much precaution, given the limitations of the study, the limits of cost-effectiveness presented in this work could be a first reference to which would be an efficient health intervention in Spain. PMID- 12113734 TI - [Disagreement among hospital and community studies evaluating the same research question]. AB - The goal of this review is to delineate some of reasons that justify the lack of consistency between hospital-based and community research. The main reasons for the differences are the selection of the hospital population and information based on clinical chart (its lack of uniformity and the treatment of the not available data). The reasons for lack of consistency are divided according to the type of research question: frequency, diagnosis, etiology, prognosis and treatment-prevention. The way a hospital population is selected justifies discrepancies regarding frequency and prognosis. As regards diagnosis, differences are mainly due the prevalence of disease. In the ascertainment of causality several biases are more common in hospital-based research, such as detection bias, protopathic (both producing an away-from-null estimate), and inclusion bias (diminishing the strength of association). Examples taken from the medical literature are offered to illustrate each bias. Regarding treatment prevention problems arise from external validity, as clinical trials are less prone to bias; this latter situation is exemplified with an assessment of vaccine efficacy in both patients and healthy population. The frequency of citation of bias was assessed by a Medline search; in hospital studies detection bias and confounding by indication were more often quoted than in non-hospital research (RR = 2.71; 95% CI; 1.69-4.37; RR = 1.76; 95% CI, 0,90-3,42, respectively). Lastly, several recommendations are given to increase the validity of hospital based research. PMID- 12113735 TI - [Write a scientific paper the easy way]. PMID- 12113736 TI - [Glass ceiling and slippery stairs? Gender inequalities and strategies for change in the Spanish Society of Public Health and Health Services Administration]. AB - In scientific societies, as in other social fields, women's participation in decision making is lower than that of men. We describe the situation in SESPAS (Spanish Society of Public Health and Health Services Administration) where, despite representing a 40% of its members, very few women have been in positions in which decisions are taken or in those of professional recognition. The process of change implemented during recent years and some of the effects of the actions taken are presented. Making the existing inequalities known has generated debate and interest in the intervention. A gender and public health working group was set up. In the last two years more women have been promoted to more senior positions in SESPAS. PMID- 12113738 TI - [The reporting of diseases. A one-century tradition]. PMID- 12113740 TI - [How should we assess the real severity of COPD?]. PMID- 12113741 TI - [The epidemiology of COPD and asthma exacerbations in a general hospital]. AB - This retrospective study identified the clinical and epidemiological characteristics of patients coming to the emergency room of our hospital with exacerbated asthma or chronic obstructive pulmonary disease (COPD) in 1993 and 1994. We followed a previously established protocol to review the case histories of patients from both years. The 1,592 exacerbations we identified in 1,209 asthmatics accounted for 0.9% of all emergency visits, with a mean of 2.2 1.6 (0 9) visits daily. The 2,106 exacerbations of COPD in 1,208 patients accounted for 1.2% of all emergencies, with a mean 2.9 (1-12) visits per day. The mean age was 51.2 (14-93) years for asthmatics and 70.3 (29-96) years for COPD patients. Of asthmatics, 69.8% were women and 30.1% were men, whereas 91.4% of COPD patients were men. The readmission rate was 3.4% for asthmatics and 4.8% for COPD patients. The hospitalization rate was 26.7% for asthmatics and 49.4% for COPD exacerbation patients. The hospital saw 22.6% of asthma exacerbations between midnight and 8 a.m. and 41.6% of COPD exacerbations during the same time frame. For both diseases, more emergencies occurred in winter. Correlation between asthma and COPD and declared influenza cases in the community were r = 0.63 (p < 0.001) for asthma and r = 0.83 (p < 0.0001) for COPD. Our findings underline the considerable emergency care burden generated by exacerbations of obstructive airway diseases and suggest that community acquired respiratory infections are usually the underlying cause. PMID- 12113742 TI - [Importance of serum interleukin-6 as a mediator of systemic inflammation in patients with alpha-1 antitrypsin deficiency]. AB - The objective of this study was to determine whether high concentrations of circulating interleukin-6 (IL-6) and/or the soluble receptor of IL-6 (SRIL-6) may mediate systemic inflammatory activity in patients with alpha-1 antitrypsin deficiency (AATD). To that end we assessed serum concentrations of IL-6 and SRIL 6 for 7 patients with AATD in stable phase. The patients' mean age was 51 years (SD 5.2); mean FEV1% was 35.5% (SD 15%). IL-6 and SRIL-6 concentrations were compared with those of 23 non-AATD patients with COPD but with similar changes in lung function (mean age 63 years, SD 10.1; FEV1% 38.3%, SD 11%). The AADT patients had mean IL-6 concentrations of 4.7 pg/mL (interquartile range [IR( 4.0) and RSIL-6 levels of 129.1 ng/mL (IR 31.5). The COPD patients had IL-6 concentrations of 4.1 pg/mL (IR 4.2) and SRIL-6 levels of 140.8 ng/mL (IR 71). No significant differences between the AADT group and the COPD group were observed for either cytokine (non-parametric Mann Whitney U test, p > 0.05). Only one AADT patient had an IL-6 concentration that was higher than normal. In conclusion, the serum IL-6 and SRIL-6 concentrations of patients with AADT are not different from those of patients with COPD, similarly altered respiratory function and normal alpha-1 antitrypsin levels. These results do not point to a role for alpha-1 antitrypsin in systemic inflammatory stimulation in patients with AADT. PMID- 12113743 TI - [Evaluation of a workplace anti-smoking program at a company with 640 employees]. AB - OBJECTIVES: Awareness of the health risks of passive smoking for non-smokers has led to the development of workplace interventions for smokers, although they are still few in our setting. The objective of this study was to evaluate the efficacy of an anti-smoking program among the workers of a company, in function of changes in the pre- and post-intervention prevalence of smoking in the study population. METHOD: A pre-intervention questionnaire on smoking (prevalence and attitudes) was administered at a company with 640 employees (92% men, 8% women). An anti-smoking program was carried out inside the company during working hours over the next nine months. The questionnaire was then repeated (post intervention) in order to evaluate its efficacy based on changes. For employees who answered both questionnaires, paired variables were analyzed. RESULTS: The pre-intervention questionnaire was answered by 388 employees (60%), 357 men (92%) and 31 women (8%); their mean age was 48.4 years (SD 9.36). The prevalence of smoking was 55%, including daily smokers, sporadic smokers and others. The mean number of cigarettes per day was 17.86 (SD 2.45). The mean level of nicotine dependence measured by the Fagerstrom test was 3.3 (SD 2.8) out of 10. The post intervention questionnaire was answered by 206 employees (32%). Among employees who answered both questionnaires (83), analysis of paired data showed a decrease in the prevalence of smoking of 4% and in the number of cigarettes per day from 17.4 (pre-intervention) to 16.4 (post-intervention). The differences were not statistically significant. CONCLUSIONS: Although the workplace is an appropriate setting for anti-smoking interventions that would contribute to improving the health of smokers, such programs are currently underutilized. PMID- 12113744 TI - [Applications of non-invasive mechanical ventilation in patients receiving endotracheal ventilation]. PMID- 12113745 TI - [Foreign bodies]. PMID- 12113746 TI - [Diagnosis and treatment of pulmonary arteriovenous fistulas]. AB - Pulmonary arteriovenous fistulas are rare malformations often associated with Rendu-Osler-Weber (ROW) disease. Morbidity and mortality are significant and arise from hemorrhagic and neurological complications.We report the cases of two patients, mother and son, with earlier diagnoses of ROW disease, who suffered, respectively, a stroke and a brain abscess with massive pulmonary thromboembolism. Helicoid computed axial tomographic scans demonstrated pulmonary arteriovenous fistulas in both. Given these findings, we performed embolotherapy. PMID- 12113747 TI - [Pulmonary capillary hemangiomatosis: a rare cause of pulmonary hypertension]. AB - Pulmonary capillary hemangiomatosis (PCH) is a rare cause of pulmonary hypertension characterized by capillary proliferation infiltrating the structures of the pulmonary parenchyma. Although veins are particularly involved, proliferation also affects bronchiolar, interstitial and other structures. We report a case of PCH in a 70-year-old man. Pulmonary artery hypertension was demonstrated by echocardiogram and angiography. Severe emphysema could be seen in a computed tomographic scan of the thorax, even though spirometric values indicated that airflow obstruction was mild. Dyspnea and respiratory insufficiency progressed with marked shunting until death. Tissue inspection at the autopsy revealed capillary proliferation in the alveolar walls with occasional oviform protrusions into air spaces or around small vessels and bronchioles. Endothelial cells in newly formed vessels were not atypical and mitosis was scarce; p53 expression was negative and Ki67 proliferation slight, indicating that PCH is not a neoplastic process as has sometimes been suggested. PMID- 12113752 TI - Food-borne intestinal trematode infections in the Republic of Korea. AB - A total of 19 species of food-borne intestinal trematodes have been reported in humans in the Republic of Korea. They include 12 species of the Heterophyidae, Metagonimus yokogawai, M. takahashii, M. miyatai, Heterophyes nocens, H. heterophyes (imported), H. dispar (imported), Heterophyopsis continua, Pygidiopsis summa, Stellantchasmus falcatus, Centrocestus armatus, Stictodora fuscata, and S. lari; four species of the Echinostomatidae, Echinostoma hortense, E. cinetorchis, Echinochasmus japonicus, and Acanthoparyphium tyosenense; and one species each of the Neodiplostomidae, Neodiplostomum seoulense, Plagiorchiidae, Plagiorchis muris, and Gymnophallidae, Gymnophalloides seoi. Fresh water fish harbor the metacercarial stage of M. yokogawai, M. takahashii, M. miyatai, C. armatus, E. hortense, E. cinetorchis, E. japonicus, or P. muris. Brackish water fish serve as the second intermediate hosts for H. nocens, H. continua, P. summa, S. falcatus, S. fuscata, and S. lari. Brackish water bivalves are the source of infection with A. tyosenense. Tadpoles and frogs are the second intermediate hosts for N. seoulense, but the major source of human infection is the grass snake Rhabdophis tigrina, a paratenic host. The metacercariae of G. seoi are observed in oysters. The natural definitive hosts are, in most cases, mammals such as rats, cats and dogs. However, several species (C. armatus, S. lari, E. japonicus, A. tyosenense, and G. seoi) have birds as natural definitive hosts. Host-parasite relationships, pathogenesis and pathology, immunity, clinical aspects, differential diagnosis, treatment, prevention and control of these intestinal trematodes are briefly discussed. PMID- 12113748 TI - [Spontaneous bacterial empyema]. PMID- 12113753 TI - Expression of excretory and secretory protein genes of Trichinella at muscle stage differs before and after cyst formation. AB - By adapting a semi-quantitative reverse transcriptase-PCR (RT-PCR) method, we investigated kinetics of gene expression at different developmental stages of Trichinella spiralis and T. pseudospiralis. The analyzed genes included four kinds of excretory and secretory (ES) proteins, a heat shock protein (HSP) and a DNA binding protein and showed that T. spiralis and T. pseudospiralis expressed ES proteins in a stage-specific manner. The gene encoding a 43 kDa ES protein was expressed by muscle larvae, either pre-cyst or post-cyst larvae. The genes encoding: the 53 kDa ES protein of T. spiralis; 53 kDa ES protein of T. pseudospiralis; and 19.6 kDa ES protein of T. spiralis were expressed by post cyst larvae and adult worms, but not expressed by pre-cyst larvae or newborn larvae. The results showed that pre-cyst larvae and post-cyst larvae are similar but different in the expression of 53 and 19.6 kDa ES proteins. On the other hand, genes of housekeeping proteins, such as HSP and the DNA binding protein, were expressed at all stages although there were some differences in the expression level. PMID- 12113754 TI - Genotypic assignment of infection by Dirofilaria repens. AB - Dirofilariasis is a parasitic disease, which if treated inappropriately due to misdiagnosis, can cause unwanted complications particularly when the infection is located in the breast. The numerous obstacles that can cause misdiagnosis of dirofilariases by standard morphological procedures prompted the development of a Dirofilaria repens-specific direct polymerase chain reaction (PCR)-based diagnostic approach using freshly infected dog blood. Reliable amplification of nematode DNA from formalin-fixed infected human specimens by this method is only possible from relatively fresh biological material, preserved in the fixative for up to 20 days. We report here our first case of dirofilariasis since the development of PCR genotyping, where the pathogen was morphologically unrecognizable and the diagnosis was based exclusively on DNA amplification. We complete our methodological contribution to the clinical laboratory diagnosis of dirofilariasis by presenting two more cases, where the primary genotypic assignment of infection by D. repens was further confirmed by conventional morphological means. PMID- 12113755 TI - Neoechinorhynchus qatarensis sp. n. (Acanthocephala: Neoechinorhynchidae) from the blue-barred flame parrot fish, Scarus ghobban Forsskal, 1775, in Qatari waters of the Arabian Gulf. AB - Neoechinorhynchus qatarensis sp. n. (Acanthocephala: Neoechinorhynchidae) is described from the marine blue-barred flame parrot fish Scarus ghobban Forsskal, 1775 in the Arabian Gulf waters off the coast of Qatar. The new species is distinguished from all other species of the genus Neoechinorhynchus by its unique reproductive system and a glandular structure associated with the proboscis receptacle in both males and females. Other uncommon features of N. qatarensis are also discussed. PMID- 12113756 TI - Enhanced UVfemale1 tumor growth in CBF1 mice infected with Schistosoma mansoni due to modulation of Th1-like responses. AB - Effects of Schistosoma mansoni infection on anti-tumor immunity were examined in CBF1 mice with ultraviolet-induced UVfemale1 fibrosarcoma cells. Although many laboratory established tumor cells had rejection mechanisms independent of CD4(+) T cells, we confirmed that CD4(+) cells had significant roles in rejection of UVfemale1 cells in the syngeneic CBF1 mice. When we prepared two CBF1 mouse groups, S. mansoni-infected and schistosome-free, the former group showed up regulation of Th2-like response to UVfemale1 cells, whereas the latter group mice showed rather type 1-dominant patterns. Cytotoxic activity against UVfemale1 cells tested in vitro, which was attributed to CD8(+) cells, was significantly weaker in S. mansoni-infected mice compared with infection-free mice. In tumor challenge experiments in vivo, we observed that rapid and complete rejection of UVfemale1 cells required the presence of CD8(+) T cells. Under only CD4-depleted situation, survival of tumor cells in schistosome-free mice was prolonged up to 1 month or more. Under the presence of both CD4(+) and CD8(+) cells, S. mansoni infected mice rejected the challenged UVfemale1 cells as was seen in normal mice. However, when CD8(+) cells were depleted from S. mansoni-infected mice, inoculated UVfemale1 cells grew more rapidly than in infection-free mice. Our results suggest that functionally polarized cytokine patterns in schistosome infected hosts promote rapid tumor growth. PMID- 12113757 TI - A theoretical analysis of the relations between the risk of congenital toxoplasmosis and the annual infection rates with a convincing argument for better public intervention. AB - In a theoretical analysis of the present study, we quantitatively indicate a potential threat of congenital toxoplasmosis to Japanese young women by the use of a simple mathematical model or a special case of the well-known catalytic infection model. For introducing a risk function of congenital toxoplasmosis, an annual infection rate, r, was divided into r(1), the rate before age a(0 < a < 15), and r(2), the rate after age a. Presuming the values of r(1), r(2) and a on the basis of the current situation of Toxoplasma infection in Japan, simulation analyses were performed with the mathematical model. As the simulation clearly demonstrated, Japanese young women are potentially facing a threat of congenital toxoplasmosis, although the current risk of it is relatively lower. From the viewpoint of risk management, public intervention programs are required. Based on our analyses, public intervention programs can be classified into two groups: group 1 for women before age a and group 2 for those after age a, and each group is expected to give a different kind of effect to the risk of congenital toxoplasmosis. The present study implies that a certain public intervention program could augment the risk, inadvertently. PMID- 12113759 TI - Study of the ethiological agent of gnathostomosis in Nayarit, Mexico. AB - In order to clarify the specific identity of the etiological agent of human gnathostomosis in Nayarit State, Mexico, morphological and molecular studies were conducted on advanced third stage larvae obtained from human and fish tissue. Cathorops fuerthii from Agua Brava lagoons complex, was the only fish species found to be infected among four species surveyed. Morphological variability does not allow specific identification of the larvae. Internal transcribed spacer 2 of the ribosomal DNA was sequenced for six larvae (five from fish, one from human tissue). Low divergence in the sequences of Nayarit larvae and Gnathostoma binucleatum (0.24% or less) indicate that the larvae examined belong to this species. PMID- 12113758 TI - Strain-specific difference in amino acid sequences of trypanosome alternative oxidase. AB - Cyanide-insensitive trypanosome alternative oxidase (TAO) is the terminal oxidase of the respiratory chain of long slender bloodstream forms of the African trypanosome, which causes sleeping sickness in human and nagana in cattle. TAO has been targeted for the development of anti-trypanosomal drugs because it does not exist in the host. The cDNA for TAO has been cloned from Trypanosoma brucei brucei EATRO110 strain and has been used for further characterization. In this study, we found amino acid sequence of the C-terminal part of TAO from the strain that we are using, T. b. brucei TC221, is considerably different from that of the EATRO110 strain. PMID- 12113760 TI - A praziquantel-ineffective fascioliasis case successfully treated with triclabendazole. AB - Human fascioliasis cases in Japan have been reported infrequently and the most appropriate therapy for this disease remains to be determined. This case report describes a patient with the infection unsuccessfully treated with multiple high doses of praziquantel for over 1 year and eventually cured by the administration of triclabendazole in two single doses of 10 mg/kg and 12.5 mg/kg 5 months apart. PMID- 12113761 TI - A more efficient Big Blue protocol improves transgene rescue and accuracy in a adduct and mutation measurement. AB - Transgenic mutational systems have provided researchers with an invaluable tool, allowing the measurement of both spontaneous and induced mutations. The Big Blue transgenic rodent mutagenesis system developed by Stratagene (La Jolla, CA) uses a lambda shuttle vector carrying lacI as the mutational target gene. A common criticism of the Big Blue system is that it relies on visual screening to detect mutants rather than positive selection, which is employed in more recently developed systems. The lack of positive selection, however, has provided the Big Blue system with a unique advantage, as it allows for the dynamic quantification of mutation fixation, repair, and adduct stability, since both pre-mutagenic DNA adducts and mutations can readily be quantified [Proc. Natl. Acad. Sci. U.S.A. 97 (2000) 11391]. Improvements to the standard Big Blue assay protocol are required for the visualization of mutant plaques resulting from pre-mutagenic damage, as these can appear much lighter in color than the lightest color control mutant (CM0). This increase in detection has been achieved by the development of a protocol that now permits the effective measurement of repair and mutation fixation utilizing the Big Blue system. This new protocol has also addressed efficiency, allowing for a two-fold increase in the number of plaques produced per packaging reaction and a decrease in both phage migration and plaque size, permitting a greater than three-fold increase in plating density. The implementation of this protocol will make the Big Blue assay more economical and less demanding than before, while providing researchers with an efficient means to measure both repair and mutation in this system. PMID- 12113762 TI - Response of the phiX174 am3, cs70 transgene to acute and chronic ENU exposure: implications for protocol design. AB - Studies of other transgenic assays have shown that time after treatment is a very important variable in the analysis of mutation frequencies but that eventually a plateau frequency is reached, indicating that the mutations are neutral. This neutrality is very important for the design of both experiments and testing protocols. Here we show that the phiX174 am3, cs70 transgene gives qualitatively similar results to the other transgenes studied after exposure of the mice to N ethyl-N-nitrosourea. In the small intestine, the mutant frequency induced by an acute dose did not change significantly from 10 to 70 days post-treatment, indicating that the mutations induced are, indeed, neutral. Likewise, the mutant frequency increased linearly with duration of exposure to ENU at a constant rate. Mutant frequencies obtained were 10 times higher from the chronic exposure than produced by a nearly lethal acute dose. As in previous comparisons of a transgene and the endogenous Dlb-1 locus in the small intestine, the chronic exposure was much more effective at increasing the sensitivity of the transgene than of the endogenous gene. The Dlb-1 locus shows more complex kinetics in this strain, as in others, with mutations initially accumulating at a slower rate, indicating a differential repair of genetic damage. PMID- 12113763 TI - Copper redox-dependent activation of 2-tert-butyl(1,4)hydroquinone: formation of reactive oxygen species and induction of oxidative DNA damage in isolated DNA and cultured rat hepatocytes. AB - The biotransformation of butylated hydroxyanisole (BHA), a possible carcinogenic food antioxidant, includes o-demethylation to 2-tert-butyl(1,4)hydroquinone (TBHQ) which can subsequently be oxidized to 2-tert-butyl(1,4)paraquinone (TBQ). In this study, we have examined the capacity of Cu, a nuclei- and DNA-associated transition metal, to mediate the oxidation of TBHQ. In phosphate buffered saline (PBS), autooxidation of TBHQ to TBQ was not detectable, while Cu(II) at micromolar concentrations strongly catalyzed the oxidation of TBHQ to TBQ. Oxidation of TBHQ by Cu(II) was accompanied by the utilization of O(2) and the concomitant generation of H(2)O(2). Using electron spin resonance spectroscopy, it was observed that Cu(II) mediated the one electron oxidation of TBHQ to a semiquinone anion radical. The formation of a semiquinone anion radical, the utilization of O(2) and the generation of H(2)O(2) and TBQ could be completely blocked by bathocuproinedisulfonic acid (BCS) and reduced glutathione (GSH), two Cu(I)-chelators. 4-Pyridyl-1-oxide-N-tert-butylnitrone (POBN)-spin trapping experiments showed that the reaction of TBHQ with Cu(II) resulted in the generation of POBN-CH(3) and POBN-CH(OH)CH(3) adducts in the presence of dimethyl sulfoxide (DMSO) and ethanol, respectively, suggesting the formation of hydroxyl radical or a similar reactive intermediate. The formation of POBN-CH(3) adduct from the TBHQ/Cu(II)+DMSO could be completely inhibited by catalase, GSH or BCS, indicating that the hydroxyl radical or its equivalent is generated from the interaction of H(2)O(2) with Cu(I). Incubation of supercoiled phiX-174 plasmid DNA with the TBHQ/Cu(II) resulted in extensive DNA strand breaks, which could be prevented by catalase or BCS. Incubation of rat hepatocytes with TBHQ in PBS led to increased formation of 8-hydroxy-2'-deoxyguanosine (8-OHdG) in nuclear DNA. The TBHQ-induced formation of 8-OHdG was markedly reduced in the presence of cell permeable Cu(I)-specific chelator, bathocuproine or neocuproine, suggesting that a Cu(II)/Cu(I) redox mechanism may also be involved in the induction of oxidative DNA damage by TBHQ in hepatocytes. Taken together, the above results conclusively demonstrate that the activation of TBHQ by Cu(II) results in the formation of TBQ, semiquinone anion radical and reactive oxygen species (ROS), and that the ROS formed may participate in oxidative DNA damage in both isolated DNA and intact cells. These reactions may contribute to the carcinogenicity as well as other biochemical activities observed with BHA in animals. To our knowledge this study provides the first evidence that endogenous cellular Cu may be capable of bioactivating TBHQ, leading to oxidative DNA damage in cultured cells. PMID- 12113764 TI - Induction of micronuclei and erythrocyte alterations in the catfish Clarias batrachus by 2,4-dichlorophenoxyacetic acid and butachlor. AB - The micronucleus test (MNT) in fish erythrocytes has increasingly been used to detect the genotoxic effects of environmental mutagens and its frequency is considered to reflect the genotoxic damage to cells, mainly the chromosomes. Besides, morphologically altered erythrocyte is taken as an index of cytotoxicity. Both parameters were used in the present study by two herbicides, 2,4-dichlorophenoxyacetic acid (2,4-D, in 25, 50 and 75ppm concentrations) and 2 chloro-2,6-diethyl-N-(butoxymethyl) acetanilide (butachlor, in 1, 2 and 2.5ppm concentrations) for genotoxic and cytotoxic endpoints. The study was carried out by an in vivo method on peripheral erythrocytes of catfish Clarias batrachus using multiple sampling times (48, 72 and 96h). Cytogenetic preparations were made by haematoxylin-eosin staining technique. Pycnotic and granular micronuclei (MN) were consistently observed irrespective of chemical tested. A wide range of altered cells was also observed. Echinocytes accompanied by altered nuclei and vacuoles were prominent feature of 2,4-D, whereas, anisochromasia and anisocytosis of erythrocytes were characteristic of butachlor. Increase in MN as well as altered cells frequencies were significant. A positive dose-response relationship in all exposures and sampling times was observed. Herbicides used were found to be genotoxic as well as cytotoxic in this fish. The suitability of the adopted parameters for the screening of the aquatic genotoxicants is discussed. PMID- 12113765 TI - A comparison between mouse and fish micronucleus test using cyclophosphamide, mitomycin C and various pesticides. AB - A comparative analysis between mouse and fish erythrocyte micronuclei (MN) assays was carried out using cyclophosphamide, mitomycin C and various pesticides such as alliete, brestanid, decis 25 CE (deltamethrin), kelthane 480 CE (dicofol), roundup (glyphosate), imazapyr and thiram. The aim of this study was to evaluate the fish species Tilapia rendalli as a suitable organism for the detection of genotoxicants in water. The clastogens cyclophosphamide and mitomycin C induced MN in both test-systems. Insecticides: decis 25 CE increased T. rendalli MN frequencies at doses of 1.0 and 5.0mg/kg, but not at the highest dose, and in mice there was no MN induction. Kelthane 480 CE also induced a significant MN frequency in T. rendalli, but not in mice. Fungicides: alliete and brestanid induced MN only in T. rendalli, while thiram was negative in both assays. Herbicides: imazapyr induced MN in T. rendalli at the maximum tolerated dose only, while roundup induced MN at three dosed levels. In mice both herbicides presented negative results. This study revealed that fish MN assay can be used as a genotoxicological test-system since some methodological particularities were observed. PMID- 12113766 TI - Mutagenicity of ground water (in the bore-holes) in Ararat Valley (Armenia) detected by Trad-SHM bioassay. AB - The genotoxicity of ground water from four bore-holes of different depths (40 120m) in the Ararat valley (Armenia) used both for drinking and irrigation was investigated. The frequency of recessive somatic mutations was determined using the Tradescantia-stamen-hair-mutation (Trad-SHM) test. The Tradescantia clone 02 was used. The pink mutation events (PMEs) were increased by 3.18-6.81-fold in comparison with the control depending both on the depth of subterranean water location and the increase of Na(+) ion concentration in these water samples. The peak frequency was found in water from the 40-45m depth. The deeper the bore holes, the lower the mutagenicity of water and the concentration of Na(+) ions. Different types of mutant sector arrangements and their frequencies changed depending on the subterranean water depth. PMID- 12113767 TI - Mutagenicity of sulfoscanate: a comparative study. AB - The mutagenic activity of sulfoscanate (SSC) (4-isothiocyanate-4'-nitrodiphenyl sulphide) has been compared with that of the following reported drugs: (a) nitroscanate (NSC) (4-isothiocyanate-4'-nitrodiphenyl ether) which is a veterinary anthelmintic drug and (b) amoscanate (ASC) (4-isothiocyanate-4' nitrodiphenyl amine) which is effective against schistosomes. SSC has been found to be a very potent mutagen towards TA98 and TA100 inducing 26.0 and 475.5revertants/nmole, respectively. NSC was found to induce mutations at a rate of 11.1 and 21.5revertants/nmole in TA98 and TA100, respectively. ASC was found to be non-mutagenic as such, but the urine of animals given the drug displayed mutagenicity. When SSC was tested in TA98/1,8-DNP(6), deficient in O acetyltransferase, the activity decreased to 10.0revertants/nmole. However, in case of NSC the mutagenic activity was reduced to 0.24revertants/nmole, indicating the importance of O-acetyltransferase in generating N acetoxyarylamine. In TA98NR, deficient in nitroreductase, the mutagenicity of SSC and NSC was totally absent. The positional isomers of SSC, 4-isothiocyanate-3' nitro- and 4-isothiocyanate-2'-nitrodiphenyl sulphide, were found to be non mutagenic in both TA98 and TA100. Our comparison of the mutagenic activity of SSC, NSC and ASC indicates that the pattern of activity is SSC>NSC>ASC. PMID- 12113768 TI - An exploratory analysis of multiple mutation spectra. AB - The last decade has witnessed a remarkable increase in the number of mutations identified both in human disease-related genes and mutation reporter genes including those in mammalian cells and transgenic animals. This has led to the curation of a number of computerised databases, which make mutation data freely available for analysis. A primary interest of both the clinical researcher and the genetic toxicologist is determination of location and types of mutation within a gene of interest. Collections of mutation data observed for a disease related gene or, for a gene exposed to a particular chemical, permits discovery of regions of sequence along the gene prone to mutagenesis and may provide clues to the origin of a mutation. The principal tool for visualising the distribution pattern of mutant data along a gene is the mutation spectrum: the distribution and frequency of mutations along a nucleotide sequence. In genetic toxicology, the current wealth of mutation data available allows us to construct many mutation spectra of interest to investigate the mutagenic mechanisms and mutational sites for one or a group of mutagens. Using the multivariate statistical methods principal components analysis (PCA) and cluster analysis (CA) we have tested the ability of these methods to establish the underlying patterns within and between 60 UV-induced, mitomycin C-induced and spontaneous mutations in the supF gene. The spectra were derived from human, monkey and mouse cells including both repair efficient and repair deficient cell lines. We demonstrate and support the successful application of multivariate statistical methods for exploring large sets of mutation spectra to reveal underlying patterns, groupings and similarities. The methods clearly demonstrate how different patterns of spontaneous and UV-induced supF mutation spectra can result from variation in plasmid, culture medium, species origin of cell line and whether mutations arose in vivo or in vitro. PMID- 12113769 TI - Genotoxicity assessment of the antiepileptic drug AMP397, an Ames-positive aromatic nitro compound. AB - AMP397 is a novel antiepileptic agent and the first competitive AMPA antagonist with high receptor affinity, good in vivo potency, and oral activity. AMP397 has a structural alert (aromatic nitro group) and was mutagenic in Salmonella typhimurium strains TA97a, TA98 and TA100 without S9, but negative in the nitroreductase-deficient strains TA98NR and TA100NR. The amino derivative of AMP397 was negative in wild-type strains TA98 and TA100. AMP397 was negative in a mouse lymphoma tk assay, which included a 24h treatment without S9. A weak micronucleus induction in vitro was found at the highest concentrations tested in V79 cells with S9. AMP397 was negative in the following in vivo studies, which included the maximum tolerated doses of 320mg/kg in mice and 2000mg/kg in rats: MutaMouse assay in colon and liver (5x320mg/kg) at three sampling times (3, 7 and 31 days after the last administration); DNA binding study in the liver of mice and rats after a single treatment with [14C]-AMP397; comet assay (1x2000mg/kg) in jejunum and liver of rats, sampling times 3 and 24h after administration; micronucleus test (2x320mg/kg) in the bone marrow of mice, sampling 24h after the second administration. Based on these results, it was concluded that AMP397 has no genotoxic potential in vivo. In particular, no genotoxic metabolite is formed in mammalian cells, and, if formed by intestinal bacteria, is unable to exert any genotoxic activity in the adjacent intestinal tissue. These data were considered to provide sufficient safety to initiate clinical development of the compound. PMID- 12113770 TI - Statistical calculation of detection limits for DNA adducts using the 32P postlabeling assay with a standard addition procedure. AB - The 32P-postlabeling assay is widely used for the analysis of DNA adducts. Some adducts can be detected with very high sensitivity but quantification can be unreliable, particularly if it is based only on comparison with unmodified nucleotides (relative adduct labeling, RAL values). Furthermore, guidelines to calculate detection limits for adduct concentrations are lacking. This is particularly important for human biomonitoring studies of environmental exposures, where a low adduct level can remain undetected. Reports of null results of toxicity studies should always include a limit of detection, indicating the effect magnitude that would have produced, with a given probability of false negative (type II error), a statistically significant increase (type I error). Here, we report on a procedure based on t-statistics to calculate two types of detection limits, the "critical level (CL)" and the "detection level (DL)". The first is the size of the difference between exposed and controls required to achieve statistical significance. The second is the size of the difference that will be detected with a chosen probability of a false negative. For the degrees of freedom (d.f.) to be used for the t-values, a general formula is given so that different standard deviations and group sizes of control and exposed groups can be handled. A sample calculation of the whole procedure is shown, using the null data for the formation of a particular adduct in lung DNA of styrene-treated mice, analyzed by 32P-postlabeling. The procedure takes into account: (i) TLC-specific background radioactivity; (ii) variability within the control and exposed groups; and (iii) confidence limits for the factor to convert 32P-radioactivity to amounts of adduct. The latter step incorporates the variance of the differences between the samples and replicates spiked with adduct standard. A statement such as follows is the result: the concentration of the alpha-isomer adduct of styrene 7,8-oxide at the O(6)-position of guanine in mouse lung DNA would have to be at least 12 adducts per 10(8) nucleotides to be detected in the given experiment on a 5% level (type I error), with a probability of 5% to miss an existing effect (type II error). PMID- 12113771 TI - The inhibitory effect of the fungicides captan and captafol on eukaryotic topoisomerases in vitro and lack of recombinagenic activity in the wing spot test of Drosophila melanogaster. AB - In studies on the mechanisms of mutagenic and carcinogenic action of captan and captafol-related chloroalkylthiocarboximide fungicides, two effects were tested: (i) the effect of both compounds on the activity of eukaryotic topoisomerases I and II in vitro, and (ii) their mutagenic and recombinagenic activity in the somatic mutation and recombination test (SMART) in wing cells of Drosophila melanogaster. Only captafol inhibited the activity of topoisomerase I (10-20% inhibition of activity in the range of 10-100microM). In contrast, both chemicals decreased the activity of topoisomerase II already at 1microM concentration (50 and 20% inhibition of activity by captafol and captan, respectively).Genotoxicity was tested in vivo by administrating both compounds by acute (3h) and chronic feeding (48h) of 3-day-old larvae. In acute feeding, captan and captafol demonstrated positive results only for small single and total spots in 10-100mM exposure concentration range. Both chemicals were inconclusive for large single spots, as well as for twin spots. In chronic treatment, captan showed positive results only for small single and total spots at 2.5 and 5mM concentrations. Captafol gave inconclusive results over all concentrations tested. The results of the acute treatment experiments which have been performed at very high doses (50% toxicity at higher doses) indicate very weak overall mutagenic activity of both test fungicides. PMID- 12113773 TI - The presence of greater than 210 MNBNC at the time of post-treatment sample evaluation showed complete response to treatment. PMID- 12113772 TI - Biomonitoring of exposure to urban air pollutants: analysis of sister chromatid exchanges and DNA lesions in peripheral lymphocytes of traffic policemen. AB - In order to elucidate the health effects of occupational exposure to traffic fumes, a few biomarkers of early genetic effect were investigated in Rome traffic policemen. One hundred and ninety healthy subjects engaged in traffic control (133 subjects) or in office work (57 subjects) participated the study. For all subjects, detailed information on smoking habits and other potential confounders were recorded by questionnaires. Average exposure of the study groups to benzene and other aromatic hydrocarbons was evaluated in a parallel exposure survey. All workers were genotyped for the following metabolic polymorphisms: CYP1A1 (m1, m2, and m4 variants), CYP2E1 (PstI and RsaI), NQO1 (Hinf1), GSTM1 and GSTT1 (null variants). In this paper, the results of the analysis of sister chromatid exchanges (SCE) in peripheral lymphocytes, and DNA damage by alkaline (pH 13) comet assay in mononuclear blood cells are reported. No statistically significant difference in the frequency of SCE or high frequency cells (HFC) was observed between traffic wardens and office workers (controls), despite the significantly higher exposure to benzene of the former (average group exposure 9.5 versus 3.8microg/m(3), 7h TWA). Conversely, both SCE per cell and HFC were highly significantly (P<0.001) increased in smokers compared to nonsmokers, showing a significant correlation (P<0.001) with the number of cigarettes per day. Multiple regression analyses of data, with metabolic polymorphisms, smoking habits, alcohol consumption, age, gender, and family history of cancer as independent variables, showed that smoking habits, and possibly the CYP2E1 variant genotypes, were the main factors explaining the variance of both SCE and HFC. Within smokers, an association of borderline significance between the CYP1A1 variant genotypes and increased SCE (P=0.050) and HFC (P=0.090) was found. This effect was mainly observed in light smokers (<15 cigarettes per day). The analysis of DNA damage by comet assay did not highlight any statistically significant difference between the exposed and control workers. Moreover, no significant model explaining tail moment variance was obtained by multiple regression analysis using the independent variables shown above. On the whole, these results indicate that exposure to moderate air pollution levels does not result in a detectable increase of genetic damage in blood cells. This evidence does not rule out any possibility of adverse effects, but strongly suggests that in urban residents life-style related factors, such as tobacco smoking, give the prevailing contribution to individual genotoxic burden. PMID- 12113775 TI - Competitive quantitative measurement of the AMPA receptor gene expression at the single cell level. AB - Our laboratory has developed a competitive reverse transcriptase polymerase chain reaction (RT-PCR) procedure to analyse the mRNA expression of the alpha-amino-3 hydroxy-5-methylisoxazole-4-propionate (AMPA) receptor subunits in single cells. By the use of an internal RNA standard competing equally with the four subunit's mRNA, we have analysed 283 whole single hippocampus CA1 cells from adult rat brain. The cells were sampled from three groups of animals: one control group, one group subjected to preconditioning ischemia, and one group subjected to global cerebral ischemia. After reverse-transcription and PCR-amplification of mRNA in the cells, the PCR product was digested using subunit specific endonucleases and quantified by Cy-5 fluorescence. The median mRNA copy numbers achieved from control rats were 290, 247, 207, and 16 GluR1-4, respectively. PMID- 12113776 TI - Quantifying presynaptic terminals at the light microscope level in intact and deafferented central nervous structures. AB - Quantification of presynaptic terminals often requires laborious techniques that involve tissue preparation for ultrastructural analysis. Modern preembedding immunohistochemical techniques provide a high morphological resolution at the light microscope level, thus allowing us to identify immunostained presynaptic boutons using specific antibodies. When absolute density of boutons (D(a)) is analysed for comparison between control and deafferented nervous tissue, quantification may be distorted due to tissue shrinkage that follows deafferentiation. The magnitude of this effect must be, therefore, estimated to correct quantitative data. Using the superior colliculus (SC) as a model, an easily applicable protocol to quantify the density of small size labelled particles in control and deafferented nervous tissue is described. This protocol was used to analyse the effect of neonatal and adult enucleation on the adult pattern of cholinergic input to the rat SC. Statistical treatment of data demonstrated that neonatal enucleation caused a drastic increase in bouton density in the visual collicular layers, stratum zonale (SZ) and stratum griseum superficiale (SGS). The same lesion carried out in adult animals caused an increase in the bouton density exclusively in the SZ. PMID- 12113779 TI - A combined immunohistochemical and autoradiographic method to detect midbrain dopaminergic neurons and determine their time of origin. AB - The present paper describes an efficient tyrosine hydroxylase immunohistochemical method combined with [3H]thymidine autoradiography to identify, in the same tissue section, substantia nigra pars compacta dopaminergic neurons and quantitatively determine their neurogenetic timetables both in control animals and in homozygous weaver mice. The experimental animals were the offspring of pregnant dams injected with [3H]thymidine on embryonic days 11-12, 12-13, 13-14 and 14-15. Preservation of tyrosine hydroxylase immunoreactivity, as well as the establishment of the best conditions for the [3H]thymidine autoradiography, were attained after many trials in which fixatives, buffers, antibody dilution and conditions for photographic emulsion, developer and fixer were tested. The proposed combined technique reveals that the detection of tyrosine hydroxylase positive neurons is accurate, unambiguous and has a low interassay variability. Moreover, it did not influence posterior [3H]thymidine autoradiography, which was highly reproducible and presented very low background. The method described here allows us to demonstrate that the neurogenetic timetables of midbrain dopaminergic neurons were different between control and homozygous weaver mice in the mesencephalic area studied. PMID- 12113777 TI - Subcellular localization of GABA receptors in the central nervous system using post-embedding immunohistochemistry. AB - The following detailed protocol can be applied to demonstrate the localization of GABA receptors in CNS neurons at the ultrastructural level. While others have investigated receptors at the electron microscopic level using immunocytochemical techniques, the appearance of the tissue is usually poor and analyses of the distribution of receptors is limited. The methodology described in this paper allows for optimal preservation of the tissue while retaining immunogenicity. It does this, in part, by utilizing a balanced salt solution washout in conjunction with fixation. When the ionic composition of a fixative solution differs from extracellular fluids, like in most fixation protocols for electron microscopy, ultrastructural changes may occur in the tissue. Balanced salt solutions, like the Tyrode solution used here, helps maintain the normal extracellular environment allowing the fixing agent to reach sufficient concentration to bring about permanent and more optimal fixation even when reduced amounts of glutaraldehyde are required to preserve antigenicity. Therefore, unlike many protocols for post-embedding immunoelectronmicroscopy, this method allows for superior preservation of tissue ultrastructure compared to results previously published by others. PMID- 12113778 TI - Methods to examine molecular changes and behavioral effects of drug administration. AB - Our laboratory has developed an integrative approach to study the molecular changes and behavioral effects of drug administration consisting of a combination of quantitative real-time reverse transcription polymerase chain reaction, RNA isolation and differential display, in situ hybridization, place preference conditioning and high-performance liquid chromatography. Although the techniques are not novel, this multi-systems approach allows for the examination of gene expression changes following the administration of drugs of abuse such as cocaine, and allows for an analysis of behavior and neurochemistry of gene knockout mice. As a result of this combination of techniques, we have been able to determine the expression, location and function of the CD81 protein. Specifically, CD81 was induced exclusively in the nucleus accumbens by cocaine treatment. Subsequent behavioral testing of CD81 knockout mice revealed these mice displayed altered sensitivity to cocaine. PMID- 12113780 TI - Isolating vessels from the mouse brain for gene expression analysis using laser capture microdissection. AB - Studies of gene expression often examine a pool of RNA extracted from the diverse cell types making up a tissue. We have developed a method for isolating vessels from the brain in order to understand the changes occurring in the vessels during the pathogenesis of cerebral malaria. Vessels were visualised by incubating sections of mouse brain with a substrate for alkaline phosphatase. Vessels were collected by laser capture microdissection and the specificity was monitored by measuring the expression of cell-specific markers. RNA from the captured vessels was highly enriched in mRNA for genes associated with endothelial cells and pericytes. Measurement of indoleamine 2,3-dioxygenase mRNA indicated it was possible to detect changes in gene expression, due to malaria infection, occurring specifically within the vessels. Laser capture microdissection can be used to study changes in gene expression occurring at the blood-brain barrier. PMID- 12113781 TI - A simplified method for combined immunohistochemistry and in-situ hybridization in fresh-frozen, cryocut mouse brain sections. AB - A method is described to perform combined immunohistochemistry and in situ hybridization in mouse brain sections. The protocol is specific to sections mounted on glass slides. In contrast to earlier methods that require either paraffin embedding or perfusion of the brain with paraformaldehyde, this protocol can be carried out on fresh-frozen, cryostat cut post-fixed sections. This simple and concise protocol increases the applicability of the technique as the RNAse free immunodetection of antigen is useful by itself for immunologically identifying specific cells of interest and then examining gene expression in those cells using techniques such as real-time PCR and microarray analysis. The use of fresh-frozen, cryocut sections enables reliable detection of easily perturbable post-translational modifications such as phosphorylation and improves the quality of results obtained in subsequent in situ hybridization by reducing the background signal and interference from lower cell layers. Inducible transgenic mice that express either a dominant negative mutant form of the cAMP response element binding protein (mCREB) or CREB, in discrete brain regions, were used in this study. The combined immunohistochemistry and in situ hybridization protocol was used to examine colocalization of enkephalin or dynorphin mRNA, both downstream targets of CREB-mediated gene expression, in cells expressing transgenic mCREB or CREB. PMID- 12113782 TI - Distinction between supraspinatus, infraspinatus and subscapularis tendon tears with ultrasound in 332 surgically confirmed cases. AB - The aim of this retrospective study was to determine the diagnostic value of preoperative ultrasonography for the characterisation of size and location of the involved tendons. in 332 consecutive patients who underwent surgery, all preoperative ultrasonographic reports were reviewed and ultrasound (US) and surgical findings were compared. Ultrasound criteria for cuff tears were complete nonvisualisation of the cuff tendons or localised absence and focal discontinuity. In all but 12 cases, US diagnoses corresponded with intraoperative findings (sensitivity 98%, confidence interval 95.1-99.3; specificity 93%, CI 85.7-97.1; accuracy 97%, CI 93.8-98.1). Size and location of the tear were correctly predicted in 69 of 96 cases (accuracy 87%, sensitivity 89%, specificity 87%). US demonstrated less extensive tears than observed at surgery in 18%. Ultrasonography was highly accurate and sensitive for detecting rotator cuff tears, but seems to be a method of limited value for evaluation of the size of cuff tears, in particular, for the detection of small tears. PMID- 12113783 TI - Prenatal depiction of cystic hygroma using three-dimensional ultrasound. AB - Prenatal diagnosis of fetal cystic hygroma is very important in clinical medicine. In this series, we report our work of detecting cystic hygroma using three-dimensional (3-D) ultrasound (US). We reviewed our computer database of prenatal diagnosis on cystic hygroma in National Cheng Kung University Hospital from May 1995 to June 2000. All the fetuses were initially scanned by a high resolution, real-time, 2-D US scanner and subsequently by a 3-D US scanner. In total, 23 cases of fetal cystic hygroma were diagnosed in utero. The range of gestational age at prenatal diagnosis by US was between 11 and 24 weeks, and 91% were diagnosed before 21 weeks. Among them, 8 cases (35%) were Turner syndrome (45,X), and 10 cases (43%) were complicated with hydrops fetalis. Although the diagnostic rates by 2-D US and 3-D US were both 100% (23 of 23), notably, 3-D US can provide additional vivid illustrations in 3-D after various modes of reconstruction, but 2-D US cannot. In conclusion, 3-D US may add novel visual depiction of the lesion in 3-D after reconstruction and, thus, assists substantially in prenatal consultation. PMID- 12113784 TI - Follow-up ultrasonography for late neck metastases of head and neck cancer. AB - Ultrasonographic examinations to detect late cervical lymph node metastases were performed at follow-up examinations for 52 patients with head and neck cancer who had not received neck dissection. Using diagnostic criteria we established previously, we accurately diagnosed 24 nodes in 10 patients as late metastases; 21 of these were nonpalpable. Of 12 nodes in 8 patients that were diagnosed as nonmetastatic using ultrasound (US) before radical treatment of the primary tumor, 11 showed an increase in the minimal diameter, 9 showed an internal echo pattern shift from homogeneous to heterogeneous and, in 7, the echogenic hilus changed from present to absent. A diagnostic equation was used to calculate a predictive value lambda for 146 nodes. Of these, 5 of 25 with a lambda value of 1 3) beta-D-Gal-(1-->3)-beta-D-Gal-(1-->4)-beta-D-Xyl-(1-->O-SerGlyTrpProAspGly (1), which was designed as a probe for glycan elongation toward heparin, was synthesized in a stereocontrolled manner. PMID- 12113805 TI - Combinatorial synthesis and biological evaluation of isoxazole-based libraries as antithrombotic agents. AB - The 3-substituted phenyl-5-isoxazolecarboxaldehydes have been identified as activated aldehydes for the generation of isoxazole-based combinatorial libraries on solid phase through automation. Three highly functionalized isoxazole-based libraries comprising of 32, 96 and 45 compounds each have been synthesized in parallel format using Baylis Hillman reaction, Michael addition, reductive amination and alkylation reactions. With an objective of lead generation all the three libraries were evaluated for their antithrombin activity in vivo. PMID- 12113806 TI - Synthesis and affinity studies of himbacine derived muscarinic receptor antagonists. AB - A series of himbacine (1)-related analogues has been prepared featuring three different isomeric configurations with respect to the B-ring (a, b and natural c) and three different interconnecting two-carbon unsaturated units [natural (E) ene, (Z)-ene, and yne]. The study of the binding affinities of the nine resulting compounds, including synthetic (+)-himbacine (3c), towards the M(1)-M(4) muscarine receptor subtypes revealed that analogues 3a and 5c display a promising 10-fold selectivity for the M(2) receptor as compared to the M(1) receptor. PMID- 12113808 TI - Synthesis of C-8 alkylamino substituted pyrrolo[2,1-c][1,4]benzodiazepines as potential anti-cancer agents. AB - The design and facile synthesis of C-8 alkylamino substituted pyrrolo[2,1 c][1,4]benzodiazepines is described. These have been prepared by linking the amines at C-8 position with propane spacer to improve solubility in water, and their in vitro cytotoxicity studies have been carried out. PMID- 12113807 TI - Photo-oxygenation of geraniol: synthesis of a novel series of hydroxy functionalized anti-malarial 1,2,4-trioxanes. AB - Photo-oxygenation of geraniol 2, an abundantly available allylic alcohol, furnished a mixture of mono- and di-hydroperoxy products; the latter have been used for the preparation of a novel series of hydroxy-functionalized anti malarial 1,2,4-trioxanes (7a-d, 8a-d). PMID- 12113809 TI - Synthesis and anti-influenza virus activity of 4-guanidino-7-substituted Neu5Ac2en derivatives. AB - Substitution of 7-OH by small hydrophobic groups on zanamivir resulted in the retaining of low nanomolar inhibitory activities against not only influenza A virus sialidase but also influenza A virus in cell culture. These compounds were prepared by treatment of the corresponding 7-substituted sialic acids derived from 4-modified N-acetyl-D-mannosamine (ManNAc) using enzyme-catalyzed aldol condensation. PMID- 12113810 TI - Synthesis and anti-influenza virus activity of 7-O-alkylated derivatives related to zanamivir. AB - A series of 7-alkyl ether derivatives related to zanamivir were synthesized using direct alkylation of the C-7 alcohol of sialic acid. Alkyl ether moiety of less than 12 carbons in length showed low nanomolar inhibitory activity against influenza A virus sialidase. Furthermore, their moiety improved influenza A virus plaque reduction activity compared to zanamivir. However, removal of the 8,9-diol of the 7-O-alkyl derivatives resulted in loss of antiviral potency. This result suggests that 8,9-diol must play an important role in binding with both influenza A and B virus sialidases. PMID- 12113811 TI - Synthesis and anti-influenza evaluation of polyvalent sialidase inhibitors bearing 4-guanidino-Neu5Ac2en derivatives. AB - We synthesized polyvalent sialidase inhibitors bearing 4-guanidino-Neu5Ac2en analogues on the polyglutamic acid back bone, via a spacer of alkyl ether at the C-7 position. These multivalent conjugates 9 and 10 showed enhancement of antiviral activity against infuenza A virus and more potent efficacy in vivo relative to a monomeric sialidase inhibitor. PMID- 12113812 TI - Design and synthesis of C-8 linked pyrrolobenzodiazepine-naphthalimide hybrids as anti-tumour agents. AB - The facile synthesis of C-8 linked pyrrolobenzodiazepine-naphthalimide hybrid analogues is described. The compounds are prepared with varying degrees of linker length in order to probe the structural requirements for optimal in vitro anti tumour activity. Some of these new hybrid compounds showed higher cytotoxic activity than the existing natural and synthetic pyrrolo[2,1 c][1,4]benzodiazepines. PMID- 12113813 TI - 2,4-Disubstituted pyrroles: synthesis, traceless linking and pharmacological investigations leading to the dopamine D4 receptor partial agonist FAUC 356. AB - Solution-phase synthesis and a solid-phase supported approach to piperazinylmethyl substituted pyrroles are described. Receptor binding studies and the measurement of D4 ligand efficacy led to the ethynylpyrrole 1d (FAUC 356) exerting selective D4 binding and substantial ligand efficacy (66%, EC(50)=1.9nM). This activity profile might be of interest for the treatment of ADHD. PMID- 12113814 TI - Synthesis and PTP1B inhibition of 1,2-naphthoquinone derivatives as potent anti diabetic agents. AB - A new series of 1,2-naphthoquinone derivatives was synthesized by various synthetic methods and evaluated for their ability to inhibit protein tyrosine phosphatase 1B (PTP1B). 1,2-Naphthoquinone derivatives with substituent at R(4) position showed submicromolar inhibitory activity, and compound 24 demonstrated 10- to 60-fold selectivity against the tested phosphatases. Also, several 4-aryl 1,2-naphthoquinone derivatives with substituents at R(3), R(6), R(7), or/and R(8) showed submicromolar inhibitory activity and good plasma stability. PMID- 12113815 TI - Novel diphenylalkyl piperazine derivatives with dual calcium antagonistic and antioxidative activities. AB - Two types of novel diphenylalkyl piperazine derivatives containing the thio or aminopropanol moiety substituted by phenyl or benzyl group were synthesized, and evaluated for their calcium antagonistic and antioxidative activities. These compounds showed apparent inhibitions against KCl-induced contractions in isolated rat aorta. Among them, phenylamino compound 9 and benzylamino compound 13 also possessed potent inhibitory activities against auto-oxidative lipid peroxidations in canine brain homogenates. Two representative compounds 3a and 9 were evaluated for their inhibitory activities against KCl-induced contractions in isolated canine arteries (basilar, coronary, mesenteric, and renal). Both compounds showed the most potent inhibitions to basilar artery. PMID- 12113816 TI - The synthesis and effect of fluorinated chalcone derivatives on nitric oxide production. AB - Dimethoxy- and trimethoxychalcone derivatives, with various patterns of fluorination, were synthesized and evaluated for their influence on nitric oxide production. Some of them, chalcones 1, 5, 7, 10, 11 and 17, inhibited NO production with an IC(50) in the submicromolar range; 17 is especially noteworthy because of its potency (IC(50) 30nM). These effects were not the consequence of a direct inhibitory action on enzyme activity but the inhibition of enzyme expression. PMID- 12113817 TI - Synthesis and anti-tumor activity of novel combretastatins: combretocyclopentenones and related analogues. AB - A series of 2-(3,4,5-trimethoxyphenyl)-3-arylcyclopent-2-ene-1-ones (8a-8e) and their related analogues, including pentenone 9a, pentenol 10a, pentene 12a, and furane 15, were synthesized and evaluated for cytotoxicity against murine and human tumor cell lines. Compounds 8a-c, 8e and 9a showed strong cytotoxicity with IC(50) values in the range of 8-34ng/mL. Compound 8e exhibited significant anti tumor activity in BDF1 mice bearing Lewis lung carcinoma cells with an inhibition ratio of 59%. PMID- 12113819 TI - Synthesis and application of an auxiliary group for chemical ligation at the X gly site. AB - An efficient synthesis of an auxiliary group, the 2-mercapto-4,5-dimethoxybenzyl (Dmmb) moiety, to form a Gly-building block is presented. The building block was incorporated into peptides to study the reaction with thiobenzyl-activated derivatives. The target peptides have been obtained by standard chemical ligation reaction, followed by TMSBr-assisted cleavage to remove the auxiliary group. Prior to Dmmb removal, under acidic conditions an unexpected rearrangement was observed and evidence for a mechanism is provided. PMID- 12113818 TI - Inhibition of ADP-triggered blood platelet aggregation by diadenosine polyphosphate analogues. AB - The synthesis and biological evaluation of new diadenosine polyphosphate analogues on blood platelet aggregation are reported. The most active are compounds with a sulfur atom replacing one or both non-bridging oxygens at phosphorus bound to adenosyl residues and hydroxymethyl groups of bis(hydroxymethyl)phosphinic acid. PMID- 12113820 TI - Novel human metabolites of the angiotensin-II antagonist tasosartan and their pharmacological effects. AB - Three novel metabolites of the angiotensin-II (A-II) receptor antagonist tasosartan have been identified in humans, and the syntheses and pharmacologic profiling of these metabolites are reported. Each metabolite bound the human A-II receptor with IC(50)s between 20 and 45nM. The in vivo effects of these compounds in attenuating the pressor response to angiotensin-II challenge in anesthetized rats were also investigated. An unsaturated diol metabolite exhibited in vivo efficacy at intravenous doses of 1 and 3mg/kg, while the other metabolites, both carboxylic acids, had no significant effect at the same doses. PMID- 12113821 TI - A solid-phase approach towards the synthesis of PDE5 inhibitors. AB - PDE5 inhibitors based upon the xanthine scaffold 8 have been expediently synthesized using a solid-phase route. Attachment of the xanthine scaffold 8 using the Rink chloride linker 4 and N-1 functionalization using Mitsunobu chemistry is described. A library of compounds was produced in multi-milligram quantities with excellent purities and acceptable yields. The compounds were tested for their PDE5 inhibitory activity. PMID- 12113822 TI - Base pairing properties of 8-oxo-7,8-dihydroadenosine in cDNA synthesis by reverse transcriptases. AB - Incorporation of nucleotides opposite 8-oxo-7,8-dihydroadenosine (8-oxoA) in oligonucleotides with dNTPs by three reverse transcriptases (AMV-, MMRV-, RAV2 RT) in cDNA synthesis was studied. Guanine as well as thymine was incorporated preferentially by all reverse transcriptases. In the melting temperature experiment, 8-oxoA and 8-oxoA-Me formed base pairs with thymine and guanine with similar stabilities. PMID- 12113823 TI - A convenient synthesis of delta(7,8)-morphinan-6-one and its direct oxidation to 14-hydroxy-delta(7,8)-morphinan-6-one. AB - Synthesis of Delta(7,8)-morphinan-6-one by Grewe cyclization and bromoketalization reaction as crucial steps is described. Introduction of a hydroxyl group at 14-position is demonstrated by direct oxidation with MnO(2) in the presence of silica gel. PMID- 12113824 TI - Discovery of non-zwitterionic GABA(A) receptor full agonists and a superagonist. AB - Numerous previous studies of GABA(A) receptor ligands have suggested that GABA(A) receptor agonists must be zwitterionic and feature an intercharge separation similar to that of GABA (approx. 4.7-6A). In this communication we demonstrate that appropriately functionalized GABA amides are partial, full, or superagonists, despite their non-zwitterionic structure. PMID- 12113826 TI - Novel cyclourethane-derived HIV protease inhibitors: a ring-closing olefin metathesis based strategy. AB - A series of novel macrocyclic urethanes incorporating a (R)-hydroxyethylamine isostere was designed and synthesized. Ring size and substituent efffects have been investigated. Cyclourethanes containing 14- to 16-membered rings exhibited low nanomolar inhibitory potencies against HIV-1 protease. PMID- 12113825 TI - A comparison of the binding of three series of nicotinic ligands. AB - A total of 24 aryl-substituted analogues of nicotine (1a) and two related series of nicotinic ligands, aminomethylpyridines 3 and ether analogues 8, were examined to determine if they bind at alpha4beta2 nACh receptors in a common manner. A modest correlation (r=0.785) was found between the affinities of the nicotine analogues and derivatives of 3, but little correlation (r=0.348) was found with analogues 8. However, a modest correlation (r=0.742) exists between the binding of analogues 3 and 8. It seems that 1-series and 8-series compounds bind differently but that the 3-series compounds share some intermediate binding similarity with both. PMID- 12113827 TI - Translocation of the 5-alkoxy substituent of 2,5-dialkoxyarylalkylamines to the 6 position: effects on 5-HT(2A/2C) receptor affinity. AB - Positional modification of 2,5-dimethoxyamphetamine analogues has been studied. Specifically, the 5-alkoxy substituent was translocated to the 6-position of the phenyl nucleus. Methoxy groups were also constrained by incorporation into appended dihydrofuran and furan rings. 2,6-Dimethoxy-4-methylamphetamine had an approximately 3-fold lower affinity for the 5-HT(2A) receptor compared to the parent 2,5-dimethoxy-4-methylamphetamine (DOM). The rigid compound based on the 2,3,5,6-tetrahydrobenzo[1,2-b;5,4-b']difuran nucleus and the aromatic analogue containing the benzo[1,2-b;5,4-b']difuran nucleus possessed an approximate 7- and 27-fold increase in affinity, respectively, compared to 2,6-dimethoxy-4 methylamphetamine, the non-rigid, positional isomer. PMID- 12113828 TI - N-[2-(Indan-1-yl)-3-mercapto-propionyl] amino acids as highly potent inhibitors of the three vasopeptidases (NEP, ACE, ECE): In vitro and In vivo activities. AB - We have previously reported the design of a lead compound 1a for the joint inhibition of neprilysin (NEP, EC 3.4.24.11), angiotensin converting enzyme (ACE, EC 3.4.15.1) and endothelin converting enzyme (ECE-1, EC 3.4.24.71), three metallopeptidases which are implicated in the regulation of fluid homeostasis and vascular tone. We report here the synthesis and biological activities of analogues derived from this lead with inhibitory potencies in the nanomolar range for the three enzymes. Compounds 8b and 15c are the most potent triple inhibitors described to date. PMID- 12113829 TI - Inhibition of feline immunodeficiency virus (FIV) replication by DNA binding polyamides. AB - Two DNA minor-groove binding polyamides 1 and 2 were designed and synthesized and evaluated for inhibition of FIV-34TF10 replication. Both 1 and 2 decreased the replication of FIV-34TF10 by 75% by acting at the level of the virus but outside of the LTR or env region. PMID- 12113830 TI - Inhibition of Src kinase activity by 4-anilino-7-thienyl-3 quinolinecarbonitriles. AB - Based on a screening lead from a yeast-based assay to identify Src family kinase inhibitors, a series of 4-anilino-7-thienyl-3-quinolinecarbonitriles was prepared. When the thiophene ring was substituted with a water-solubilizing group in a 2,5-, 3,5- or 2,4-pattern, potent inhibition of Src kinase activity was observed. PMID- 12113831 TI - Convergent synthesis of an inner core GPI of sperm CD52. AB - An inner core of the GPI anchor of sperm CD52 antigens was synthesized by a highly convergent process using specially modified inositol, glucosamine and phospholipid as key building blocks. This paper also presents a new and efficient procedure to prepare 1,2,6-differentiated derivatives of inositol for GPI syntheses. PMID- 12113832 TI - 2-(2-Hydroxy-3-alkoxyphenyl)-1H-benzimidazole-5-carboxamidine derivatives as potent and selective urokinase-type plasminogen activator inhibitors. AB - The development of potent and selective urokinase-type plasminogen activator (uPA) inhibitors based on the lead molecule 2-(2-hydroxy-3-ethoxyphenyl)-1H benzimidazole-5-carboxamidine (3a) is described. PMID- 12113833 TI - 4-Aminoarylguanidine and 4-aminobenzamidine derivatives as potent and selective urokinase-type plasminogen activator inhibitors. AB - The structure-based design of potent and selective urokinase-type plasminogen activator (uPA) inhibitors with 4-aminoarylamidine or 4-aminoarylguanidine S1 binding groups, is described. PMID- 12113834 TI - The synthesis and biological evaluation of a series of potent dual inhibitors of farnesyl and geranyl-Geranyl protein transferases. AB - We have prepared a series of potent, dual inhibitors of the prenyl transferases farnesyl protein transferase (FPTase) and geranyl-geranyl protein transferase I (GGPTase). The compounds were shown to possess potent activity against both enzymes in cell culture. Mechanistic analysis has shown that the compounds are CAAX competitive for FPTase inhibition but geranyl-geranyl pyrophosphate (GGPP) competitive for GGPTase inhibiton. PMID- 12113835 TI - Structure-activity relationships of non-imidazole H(3) receptor ligands. Part 1. AB - SAR studies for novel non-imidazole containing H(3) receptor antagonists with high potency and selectivity for rat H(3) receptors are described. A high throughput screening lead, A-923, was further elaborated in a systematic manner to clarify a pharmacophore for this class of aryloxyalkyl piperazine based compounds. PMID- 12113836 TI - Structure-activity relationships of non-imidazole H(3) receptor ligands. Part 2: binding preference for D-amino acids motifs. AB - Structure-activity relationship studies on novel non-imidazole, D-amino acid containing ligands of histamine 3 receptors are presented. A-304121 is a D alanine piperazine amide with high affinity at the rat H(3) receptor. PMID- 12113837 TI - Synthesis and biological evaluation of a spongistatin AB-spiroketal analogue. AB - The synthesis of a simplified analogue of the potent, cytotoxic tubulin depolymerizing agent spongistatin 1, based on the AB spiroketal framework, is presented. The new structural analogue is an extension of a recently described spongistatin congener reported to disrupt microtubules in breast cancer cells in vitro and to alter the microtubule assembly reaction. Cytotoxicity data on the new structural analogue, as well as the parent congener, are reported. We found no significant cytotoxic or antitubulin activity with either compound. PMID- 12113840 TI - Cardiopulmonary resuscitation using the cardio vent device in a resuscitation model. AB - To compare the "Bellows on Sternum Resuscitation" (BSR) device that permits simultaneous compression and ventilation by one rescuer with two person cardiopulmonary resuscitation (CPR) with bag-valve-mask (BVM) ventilation in a single blind crossover study performed in the laboratory setting. Tidal volume and compression depth were recorded continuously during 12-min CPR sessions with the BSR device and two person CPR. Six CPR instructors performed a total of 1,894 ventilations and 10,532 compressions in 3 separate 12-min sessions. Mean tidal volume (MTV) and compression rate (CR) with the BSR device differed significantly from CPR with the BVM group (1242 mL vs. 1065 mL, respectively, p = 0.0018 and 63.2 compressions per minute (cpm) vs. 81.3 cpm, respectively, p = 0.0076). Error in compression depth (ECD) rate of 9.78% was observed with the BSR device compared to 8.49% with BMV CPR (p = 0.1815). Error rate was significantly greater during the second half of CPR sessions for both BSR and BVM groups. It is concluded that one-person CPR with the BSR device is equivalent to two-person CPR with BVM in all measured parameters except for CR. Both groups exhibited greater error rate in CPR performance in the latter half of 12-min CPR sessions. PMID- 12113838 TI - Substituted acrylamides as factor Xa inhibitors: improving bioavailability by P1 modification. AB - To overcome the low bioavailability of our substituted acrylamide P1 benzamidine factor Xa inhibitors reported previously, neutral and less basic groups were used to replace the benzamidine. As a result, a series of P1 aminoisoquinoline substituted acrylamide Xa inhibitors was identified to be potent, selective, and orally bioavailable. Modification of P4 moiety of these compounds further improved their pharmacokinetic properties. PMID- 12113839 TI - Preparation of soluble and insoluble polymer supported IBX reagents. AB - A series of soluble and insoluble polymer supported versions of the versatile oxidizing reagent IBX has been prepared. Each of the reagents were evaluated for their efficiency in the conversion of benzyl alcohol to benzaldehyde. Results from this study were that the soluble, non-crosslinked polystyrene supported IBX reagent gave the best rate of conversion to benzaldehyde, while the macroporous polymer supported IBX resin provided a superior rate of conversion to benzaldehyde when compared with a gel type resin. The macroporous IBX reagent was also shown to convert a series of alcohols to the corresponding aldehydes and ketones. PMID- 12113841 TI - Descriptive epidemiology of a cluster of hand injuries from snowblowers. AB - The objective of the study is to describe the nature and type of snowblower related hand injuries, including long term follow up. A retrospective study was performed of a cluster of patients injured by snowblower use, presenting to an urban Emergency Department (ED) following a major snowstorm. For each patient, demographic data and subsequent records from outpatient management were reviewed. Interviews were conducted 3 years later to assess long-term outcomes. Eleven patients, all male, presented to the ED with snowblower injuries, all to the hand. Injuries included soft tissue lacerations and partial to complete amputations. Ten patients were treated and released and one was admitted. In a 3 year follow-up, few patients reported major sequela. Almost half reported persistent pain, or minor disability. In conclusion, in our series, snowblower injuries resulted exclusively in trauma to the hand. Most injuries can be managed on an outpatient basis. Many patients experience persistent symptoms. All patients had entirely preventable injuries. PMID- 12113842 TI - An evaluation of the risk for latex allergy in prehospital EMS providers. AB - Hospital health care providers are increasingly being diagnosed as latex sensitive or allergic. Little is established on incidence or risks to prehospital health care providers. A written survey of EMT-DCs and EMT-Ps was done anonymously using established risk stratification questions to identify factors that indicate higher potential for developing latex allergies. There were 666 surveys distributed with 580 (87%) returned completed. Of the respondents, 533 were male (91%) with 510 (87%) reporting more than 5 years of field experience. Of the survey participants, 435 (75%) were EMT-DC level and 145 (25%) were EMT-P level. We found that latex sensitivities and allergies are present in our population, with an 8% incidence of latex allergies in EMT-DCs and 18% in EMT-Ps. A greater number of respondents report having factors that have been established to be associated with increased risk for latex allergies, indicating the need for more vigilant monitoring for the development of such reactions. PMID- 12113843 TI - Fatal hyperkalemia related to combined therapy with a COX-2 inhibitor, ACE inhibitor and potassium rich diet. AB - We describe the case of a 77-year old mildly hypertensive woman with no underlying renal disease who was admitted to the Emergency Department (ED) in a comatose state with fever. The patient had been on low dose enalapril and a potassium rich diet. Five days before admission, rofecoxib, a new selective COX-2 inhibitor nonsteroidal anti-inflammatory drug (NSAID), was added for leg pain. She was found to have severe hyperkalemia and died 90 min after her arrival. We cannot absolutely determine whether the COX-2 inhibitor was the dominant contributor to the development of hyperkalemia or the combination itself, with an intercurrent infection and some degree of dehydration. Physicians should be aware of this possible complication and only prescribe NSAIDs, including the new COX-2 drugs, to the elderly under close monitoring of kidney function and electrolyte tests. PMID- 12113844 TI - Airway compromise after routine alpha-hydroxy facial peel administration. AB - High concentration alpha-hydroxy facial "peels" are widely used for cosmetic anti aging purposes. We report a case of secondary burns to the neck and face resulting in airway compromise after a professionally applied citric acid facial treatment. PMID- 12113846 TI - Thrombocytopenia and altered mental status in an HIV-positive woman. AB - We present a case of thrombotic thrombocytopenic purpura (TTP) in a human immunodeficiency virus (HIV)-positive woman with altered mental status. Altered mental status with thrombocytopenia may be due to many causes, including consumptive coagulopathy, systemic lupus erythematosis, infection, and as side effects of commonly used anti-seizure medications. Of these, platelet transfusion is ineffective or specifically contraindicated in the consumptive coagulopathies, including TTP. TTP should be considered in all patients with altered mental status or neurologic dysfunction, thrombocytopenia, and hemolytic anemia to prevent morbidity and mortality. PMID- 12113845 TI - Fulminant infection and toxic shock syndrome caused by Streptococcus pyogenes. AB - Two patients presented to the Emergency Department (ED) with features of toxic shock syndrome, including hypotension, acute respiratory distress syndrome (ARDS), renal and hepatic insufficiency and disseminated intravascular coagulation (DIC). Computed tomography (CT) scan identified the source of infection in one patient. At laparotomy, pelvic peritonitis and massive edema of the pelvic retroperitoneal tissue was found. The other patient had myonecrosis of the forearm necessitating amputation. Intra-operative cultures of tissue in each case yielded Streptococcus pyogenes, Group A. These patients were treated early with clindamycin and intensive supportive care as well as surgery, and both made a full recovery. Because of the necessity of early recognition of the varied presentation of these infections, the clinical features as well as essential interventions are emphasized. We review the pathophysiology of invasive Group A streptococcal infection to increase awareness of these uncommon but fulminant and often lethal infections. PMID- 12113847 TI - Hypothermia from prolonged immersion: biophysical parameters of a survivor. AB - We report a case of survival following prolonged immersion and hypothermia. The patient survived for over 9 h in open water, after his vessel capsized and sank in the Pacific Ocean off the coast of Northern California. Water temperature on the day of the sinking was 14.4 degrees C (58.0 degrees F). Although he did have adequate flotation, the patient did not wear a survival suit. On initial physical examination in the Emergency Department (ED), the patient's rectal temperature was 30.0 degrees C (86.0 degrees F). With active rewarming, his temperature returned to normal (37.0 degrees C (98.6 degrees F)) within 5 h. Body fat of the patient was 19.6%, near the 50th percentile for his age (19.0%). Surface/volume ratio of the patient (.0228 m(2)/L) was 19% smaller than a predicted average (.0282 m(2)/L). We believe that the patient's large body habitus contributed to survival and that surface/volume ratio was likely the biophysical variable most closely associated with decreased cooling. PMID- 12113848 TI - An unusual presentation of stridor: blunt pediatric laryngotracheal trauma. AB - A case report is presented to review pediatric blunt laryngotracheal trauma. The common presenting symptoms of stridor and respiratory distress and their implications for the pediatric trauma patient are discussed. Use of computed tomography and direct endoscopy of the airway should help optimize airway management in these patients. PMID- 12113849 TI - Color doppler in the diagnosis of ectopic pregnancy in the emergency department: is there anything beyond a mass and fluid? AB - Pregnant patients with first trimester complications are a common presentation in many Emergency Departments (EDs). The burden of disproving the existence of an ectopic pregnancy falls on the Emergency Physician (EP). This may be a difficult task depending on the availability of specialty backup and radiologic services. For more than a decade EPs have documented use of bedside ultrasonography for ruling out cases of ectopic pregnancy. Now, with the entry of newer technology into many emergency ultrasound programs, color Doppler is available to an increasing number of EPs. Color Doppler is used as an adjunct to diagnosing ectopic pregnancy by both radiology and gynecology practitioners and can at times provide critical information regarding early pregnancy. Presented are three cases of ectopic pregnancy diagnosed on the basis of abnormal color Doppler findings, and a discussion of the technique. PMID- 12113850 TI - Deaths associated with inappropriate intravenous colchicine administration. AB - Intravenous (IV) colchicine is occasionally prescribed for the treatment of acute gouty arthritis. The Food and Drug Administration (FDA) recently received a report of death in a patient that was associated with inappropriate IV dosing of colchicine. This report prompted further investigation of other deaths associated with IV colchicine use in the FDA Adverse Event Reporting System (AERS) and the medical literature. A total of 20 deaths were identified. Eight patients were females, 11 were males, and the gender was unknown in 1. In all cases, the recommended maximum cumulative dose of 2 to 4 mg during a course of therapy was exceeded. Dose reductions are recommended in patients with renal or hepatic disease and in the elderly. All reported adverse events were associated with colchicine toxicity, including thrombocytopenia, leukopenia, pancytopenia, agranulocytosis, aplastic anemia, acute renal failure, and disseminated intravascular coagulopathy. Death occurred within 1 to 40 days after drug administration. Therapeutic guidelines exist for use of IV colchicine and these guidelines should be followed to prevent serious toxicities and death. PMID- 12113851 TI - Paroxysmal supraventricular tachycardia induced by oral stimulation: a case report and review of swallow-induced dysrhythmias. AB - We present the case of a 54-year-old man who experienced reproducible paroxysmal supraventricular tachycardia (SVT) with simple oral stimulation. The tachycardia was felt to be focal atrial fibrillation, and the patient was placed on propafenone with good results. There are no previous known cases of this exact condition. However, this may represent a variant of swallow-induced tachycardia. Case reports involving swallow-induced tachycardia with speculated mechanisms and treatments are discussed. PMID- 12113852 TI - Point of care testing in the emergency department. AB - Point of care (POC) testing in the Emergency Department (ED) is becoming more common. The implementation and maintenance of POC testing in the ED, however, is a complex issue. We performed a systematic review of the English language literature published between 1985 and June 2001 with a focus on POC testing and ED application. Articles that addressed the following were included in the review: implementation of POC testing, maintenance and regulation of POC testing, and application of POC testing. Current POC technology has been found to be reliable in various patient care settings, including the ED. Cost and connectivity issues are complex and difficult to assess, making these the greatest barriers to the full acceptance of POC testing in the ED. Patient care issues must be weighed against the cost of implementing POC testing and supporting the infrastructure needed to maintain this technology in the ED. PMID- 12113853 TI - A young woman with altered mental status. PMID- 12113854 TI - Diagnosis of ruptured ectopic pregnancy by bedside ultrasonography. PMID- 12113855 TI - The utility of ultrasound in a case of pericardial tamponade. PMID- 12113857 TI - Tension pneumoperitoneum--a complication of colonoscopy: recognition and treatment in the emergency department. PMID- 12113858 TI - Rupture of a previously normal spleen in association with enoxaparin: an unusual cause of shock. PMID- 12113856 TI - A critically ill young man with septic emboli to the lungs. PMID- 12113860 TI - Home administration of charcoal: can mothers administer a therapeutic dose? PMID- 12113861 TI - Silent ischemia. PMID- 12113862 TI - The unique educational value of emergency medicine student interest groups. AB - Student interest groups offer many additional educational opportunities for medical students. The discipline of Emergency Medicine is uniquely positioned to provide medical students with additional resources that may enhance student involvement in clinical and community projects. Because of Emergency Medicine's strong intrinsic ties with both clinical medicine and the surrounding community, students can use Emergency Medicine student interest groups to implement programs designed to help them gain clinical exposure, fuel research ideas, serve the local community, and most importantly, better themselves as medical students. Such opportunities can be advantageous to medical students entering any medical specialty. Thus, Emergency Medicine student interest groups are a unique and valuable educational resource for all medical students, providing many opportunities for students to enhance their medical education. PMID- 12113865 TI - Scientific and clinical challenges in the management of urinary tract infections. PMID- 12113866 TI - Epidemiology of urinary tract infections: incidence, morbidity, and economic costs. AB - Urinary tract infections (UTIs) are considered to be the most common bacterial infection. According to the 1997 National Ambulatory Medical Care Survey and National Hospital Ambulatory Medical Care Survey, UTI accounted for nearly 7 million office visits and 1 million emergency department visits, resulting in 100,000 hospitalizations. Nevertheless, it is difficult to accurately assess the incidence of UTIs, because they are not reportable diseases in the United States. This situation is further complicated by the fact that accurate diagnosis depends on both the presence of symptoms and a positive urine culture, although in most outpatient settings this diagnosis is made without the benefit of culture. Women are significantly more likely to experience UTI than men. Nearly 1 in 3 women will have had at least 1 episode of UTI requiring antimicrobial therapy by the age of 24 years. Almost half of all women will experience 1 UTI during their lifetime. Specific subpopulations at increased risk of UTI include infants, pregnant women, the elderly, patients with spinal cord injuries and/or catheters, patients with diabetes or multiple sclerosis, patients with acquired immunodeficiency disease syndrome/human immunodeficiency virus, and patients with underlying urologic abnormalities. Catheter-associated UTI is the most common nosocomial infection, accounting for >1 million cases in hospitals and nursing homes. The risk of UTI increases with increasing duration of catheterization. In noninstitutionalized elderly populations, UTIs are the second most common form of infection, accounting for nearly 25% of all infections. There are important medical and financial implications associated with UTIs. In the nonobstructed, nonpregnant female adult, acute uncomplicated UTI is believed to be a benign illness with no long-term medical consequences. However, UTI elevates the risk of pyelonephritis, premature delivery, and fetal mortality among pregnant women, and is associated with impaired renal function and end-stage renal disease among pediatric patients. Financially, the estimated annual cost of community-acquired UTI is significant, at approximately $1.6 billion. PMID- 12113867 TI - The etiology of urinary tract infection: traditional and emerging pathogens. AB - The microbial etiology of urinary infections has been regarded as well established and reasonably consistent. Escherichia coli remains the predominant uropathogen (80%) isolated in acute community-acquired uncomplicated infections, followed by Staphylococcus saprophyticus (10% to 15%). Klebsiella, Enterobacter, and Proteus species, and enterococci infrequently cause uncomplicated cystitis and pyelonephritis. The pathogens traditionally associated with UTI are changing many of their features, particularly because of antimicrobial resistance. The etiology of UTI is also affected by underlying host factors that complicate UTI, such as age, diabetes, spinal cord injury, or catheterization. Consequently, complicated UTI has a more diverse etiology than uncomplicated UTI, and organisms that rarely cause disease in healthy patients can cause significant disease in hosts with anatomic, metabolic, or immunologic underlying disease. The majority of community-acquired symptomatic UTIs in elderly women are caused by E coli. However, gram-positive organisms are common, and polymicrobial infections account for up to 1 in 3 infections in the elderly. In comparison, the most common organisms isolated in children with uncomplicated UTI are Enterobacteriaceae. Etiologic pathogens associated with UTI among patients with diabetes include Klebsiella spp., Group B streptococci, and Enterococcus spp., as well as E coli. Patients with spinal cord injuries commonly have E coli infections. Other common uropathogens include Pseudomonas and Proteus mirabilis.Recent advances in molecular biology may facilitate the identification of new etiologic agents for UTI. The need for accurate and updated population surveillance data is apparent, particularly in light of concerns regarding antimicrobial resistance. This information will directly affect selection of empiric therapy for UTI. PMID- 12113868 TI - The optimal use of diagnostic testing in women with acute uncomplicated cystitis. AB - Acute uncomplicated cystitis is a common and costly disorder in women, and there is considerable variation in the diagnostic strategies currently used in clinical practice. Because the diagnosis of cystitis can be established in most patients using the history alone, the clinician's responsibility is to determine which patients require additional diagnostic testing. Patients with typical symptoms (i.e., dysuria, frequency, urgency, hematuria), without risk factors for complicated infection or pyelonephritis, and without a history of vaginal discharge, have a very high probability of cystitis and are appropriate candidates for empiric treatment. It is more difficult, however, to rule out infection in patients with suspected cystitis. Because the prevalence of culture proven infection is very high in women who present with >/=1 symptom, and because the treatment threshold for this condition is low, a urine culture is generally required to rule out infection in patients with atypical symptoms, even in the presence of a negative dipstick test. In population-based, before-and-after studies, use of diagnostic algorithms has been shown to significantly decrease the use of urinalysis, urine culture, and office visits while increasing the percentage of patients who receive recommended antibiotics. These strategies have substantially reduced the cost of managing cystitis without an increase in adverse events or a decrease in patient satisfaction. Randomized controlled trials are needed to more closely examine the outcomes, costs of care, and patient satisfaction from different diagnostic and management strategies. PMID- 12113869 TI - Addressing antibiotic resistance. AB - Management of uncomplicated urinary tract infections (UTIs) has traditionally been based on 2 important principles: the spectrum of organisms causing acute UTI is highly predictable (Escherichia coli accounts for 75% to 90% and Staphylococcus saprophyticus accounts for 5% to 15% of isolates), and the susceptibility patterns of these organisms have also been relatively predictable. As a result, empiric therapy with short-course trimethoprim-sulfamethoxazole (TMP SMX) has been a standard management approach for uncomplicated cystitis.However, antibiotic resistance is now becoming a major factor not only in nosocomial complicated UTIs, but also in uncomplicated community-acquired UTIs. Resistance to TMP-SMX now approaches 18% to 22% in some regions of the United States, and nearly 1 in 3 bacterial strains causing cystitis or pyelonephritis demonstrate resistance to amoxicillin. Fortunately, resistance to other agents, such as nitrofurantoin and the fluoroquinolones, has remained low, at approximately 2%. Preliminary data suggest that the increase in TMP-SMX resistance is associated with poorer bacteriologic and clinical outcomes when TMP-SMX is used for therapy. As a result, these trends have necessitated a change in the management approach to community-acquired UTI. The use of TMP-SMX as a first-line agent for empiric therapy of uncomplicated cystitis is only appropriate in areas where TMP-SMX resistance prevalence is <10% to 20%. In areas where resistance to TMP-SMX exceeds this rate, alternative agents need to be considered. PMID- 12113870 TI - Urinary tract infection: traditional pharmacologic therapies. AB - Urinary tract infections (UTIs) are common bacterial infections, particularly in women. Antimicrobial therapy is seldom indicated for asymptomatic infection, but antimicrobial therapy is usually indicated for amelioration of symptoms. Management of acute uncomplicated UTI (cystitis) is generally straightforward, with a predictable distribution of uropathogens isolated. First-line treatment of acute uncomplicated UTI has traditionally involved a 3-day regimen of trimethoprim-sulfamethoxazole (TMP-SMX) or TMP alone for patients with sulfa allergies. Increasing resistance among community-acquired Escherichia coli to TMP SMX worldwide has led to a reassessment of the most appropriate empiric therapy for these infections. Alternative first-line agents include the fluoroquinolones, nitrofurantoin, and fosfomycin. Factors to be considered in the selection of appropriate antimicrobial therapy include pharmacokinetics, spectrum of activity of the antimicrobial agent, resistance prevalence for the community, potential for adverse effects, and duration of therapy. Ideal antimicrobial agents for UTI management have primary excretion routes through the urinary tract to achieve high urinary drug levels. In addition, there are special considerations in the management of UTI among selected populations, including postmenopausal and pregnant women, and for women with frequent recurrent UTIs. PMID- 12113871 TI - The expanding role of fluoroquinolones. AB - There has been a growing rate of resistance among common urinary tract pathogens, such as Escherichia coli, to traditional antimicrobial therapies including the "gold standard" trimethoprim-sulfamethoxazole (TMP-SMX). Consequently, fluoroquinolone antimicrobial agents have taken on an expanding management role for UTIs. In fact, the recent Infectious Diseases Society of America clinical management guidelines for UTI recommend fluoroquinolones as first-line therapy for uncomplicated UTI in areas where resistance is likely to be of concern. Fluoroquinolones have demonstrated high bacteriologic and clinical cure rates, as well as low rates of resistance, among most common uropathogens. There are currently 7 fluoroquinolones with indications for UTI in the United States. However, only 3 are commonly used: levofloxacin, ciprofloxacin, and, to a lesser extent, gatifloxacin. Many of the fluoroquinolone agents have once-daily dosing regimens, enhancing patient adherence. In addition, levofloxacin and gatifloxacin have same-dose bioequivalency between their intravenous and oral formulations, allowing for "switch" or step-down therapy from parenteral to oral formulations of the same agent at the same dose. Fluoroquinolones are indicated for the management of acute uncomplicated UTIs, as well as complicated and severe UTI and pyelonephritis, in adults. They are the first-line treatment of acute uncomplicated cystitis in patients who cannot tolerate sulfonamides or TMP, who live in geographic areas with known resistance >10% to 20% to TMP-SMX, or who have risk factors for such resistance. Fluoroquinolone properties include a broad spectrum of coverage, low rates of resistance, and good safety profiles. PMID- 12113872 TI - Urinary tract infection at the age extremes: pediatrics and geriatrics. AB - Urinary tract infections (UTIs) are common and generally benign conditions among healthy, sexually active young women without long-term medical sequelae. In contrast, UTIs are more complicated among those individuals at either end of the age spectrum: infants/young children and geriatrics. UTI in children younger than 2 years has been associated with significant morbidity and long-term medical consequences, necessitating an extensive and somewhat invasive imaging evaluation to identify possible underlying functional or anatomic abnormalities. Pediatric UTI should be considered complicated until proved otherwise, and treatment should reflect the severity of signs and symptoms. Management in the acutely ill child frequently involves parenteral broad-spectrum antimicrobial agents, and less ill children can be treated with trimethoprim- sulfamethoxazole (TMP-SMX), beta lactams, and cephalosporins.UTI among older patients (>65 years) may be complicated by comorbidities, the baseline presence of asymptomatic bacteriuria, and benign urinary symptoms that can complicate diagnosis. The etiology of UTI encompasses a broader spectrum of infecting organisms than is seen among younger patients and includes more gram-positive organisms. Symptomatic UTI is generally more difficult to treat than among younger populations. Management should be conservative, of longer treatment durations, and cover a broad spectrum of possible uropathogens. Oral or parenteral treatment with a fluoroquinolone for 7 days is the preferred empiric approach. TMP-SMX can also be considered a first line agent in women only, but only if the pathogen is known to be TMP-SMX sensitive. PMID- 12113873 TI - Pathogenesis of bacteriuria and infection in the spinal cord injured patient. AB - Spinal cord injury (SCI) produces profound alterations in lower urinary tract function. Incontinence, elevated intravesical pressure, reflux, stones, and neurological obstruction, commonly found in the spinal cord-injured population, increase the risk of urinary infection. The overall rate of urinary infection in SCI patient is about 2.5 episodes per patient per year. Despite improved methods of treatment, urinary tract morbidity still ranks as the second leading cause of death in the SCI patient.SCI removes the ability of the pontine micturition center and higher centers in the brain to inhibit, control, or coordinate the activity of the vesicourethral unit. As a result, a patient with complete quadriplegia is typically unaware of bladder activity. Bladder contraction is accompanied by vesicosphincter dyssynergia instead of sphincter relaxation. It is widely accepted that intermittent catheterization, when compared with indwelling catheters, reduces the risk of urinary tract infection (UTI) in SCI patients and is the preferred method of bladder drainage in this patient population. Attempts at eliminating bacteriuria associated with indwelling or intermittent catheters have generally been unsuccessful. There is now appreciation of the fact that a creeping adherent biofilm of bacteria frequently ascends through the luminal and external surfaces of an indwelling catheter, often within 8 to 24 hours, leading to bacterial adherence to the bladder surface and correlating with symptomatic infection. The use of antimicrobial agents to clear or prevent bacteriuria in patients on indwelling or intermittent catheterization has had mixed success. Treatment for asymptomatic bacteriuria in SCI patients remains controversial. SCI patients with symptomatic urinary infections should be treated with the most specific, narrowest spectrum antibiotics available for the shortest possible time. Guidelines for selecting antimicrobial agents in SCI patients are similar to guidelines for the treatment of complicated urinary infections in the general population. Characteristics of the quinolones make them well suited to treating UTI in the SCI patient. PMID- 12113874 TI - Urinary tract infections in patients with diabetes. AB - Results of various epidemiologic studies suggest that bacteriuria and urinary tract infection (UTI) occur more commonly in women with diabetes than in women without this disease. Similar findings have been demonstrated for asymptomatic bacteriuria (ASB), with ASB being a risk factor for pyelonephritis and subsequent decline in renal function. Although ASB is not associated with serious health outcomes in healthy patients, further research needs to be undertaken regarding the impact of ASB in patients with diabetes. Patients with diabetes often have increased complications of UTI, including such rare complications as emphysematous cystitis and pyelonephritis, fungal infections (particularly Candida species), and increased severity and unusual manifestations (e.g., gram negative pathogens other than Escherichia coli). Anatomic and functional abnormalities of the urinary tract are also associated with diabetes. Such abnormalities result in greater instrumentation of the urinary tract, thereby increasing the risk of secondary UTI. In addition, these abnormalities complicate UTI and require specialized treatment strategies. There is a greater likelihood of UTI affected by antimicrobial resistance or atypical uropathogens, and the risk of upper tract involvement is increased. Pre- and posttherapy urine cultures are therefore indicated. The initial choice of empiric antimicrobial therapy should be based on Gram stain and urine culture. Choice of antibiotic therapy should integrate local sensitivity patterns of the infecting organism. Fluoroquinolones are a reasonable empiric choice for many patients with diabetes. For seriously ill patients, including patients infected with Pseudomonas spp., such agents as imipenem, ticarcillin-clavulanate, and piperacillin-tazobactam may also be considered. Treatment of ASB in patients with diabetes is often recommended to prevent the risk of symptomatic UTI. However, the management of ASB in patients with diabetes is complex, with no single preferred approach. PMID- 12113876 TI - The effects of phytoestrogen supplementation in postmenopausal women. PMID- 12113877 TI - Three-dimensional sonography for assessment of morphology and vascularization of the fetus and placenta. AB - OBJECTIVE: To analyze the potential of three-dimensional power Doppler sonography in morphologic and functional assessment of the fetus and placenta. METHODS: Review of the recent literature on three-dimensional sonography in early pregnancy and in the second and third trimester. RESULTS: Three-dimensional sonography plays an important role in obstetrics predominantly for assessing fetal anatomy. Multiplanar images and rotation of the object allow systematic review of anatomic structures, such as limb buds, cerebral cavities, cord insertion, stomach, and bladder. Using this modality, volumes of the gestational sac, yolk sac, and fetal organs can be obtained easily. Three-dimensional power Doppler sonography has the potential to study the intervillous and placental circulation and evaluate the development of the embryonic and fetal cardiovascular system. CONCLUSIONS: Three-dimensional ultrasound imaging complements pathologic and histologic evaluation of the developing embryo, giving rise to the new term "three-dimensional sonoembryology." It is evident that three dimensional ultrasonography improves the visualization of the normal and abnormal fetal anatomy giving a realistic impression of the extent of the defects. These data are useful not only for obstetricians but also for pediatricians and pediatric surgeons. PMID- 12113878 TI - Effect of long-term, postasphyxial administration of magnesium sulfate on immunostaining of microtubule-associated protein-2 and activated caspase-3 in 7 day-old rat brain. AB - OBJECTIVE: To test whether magnesium inhibits apoptosis during hypoxia-ischemia (HI) in the developing brain, we studied the effect of long-term magnesium treatment on an early marker of neuronal damage (loss of microtubule-associated protein-2; MAP-2) and a marker of apoptosis (activated caspase-3) after HI in newborn rats. METHODS: Seven-day-old rat pups (n = 107) were exposed to unilateral carotid artery ligation and 2 hours of hypoxia (8% oxygen in 92% nitrogen). Magnesium was administered by a micro-osmotic pump implanted in the back, at an infusion rate of 75 mg/kg per hour for 3 days. Neuronal loss in the cerebral cortex and hippocampus was evaluated by loss of MAP-2 at 24 and 48 hours after HI and visually ranked by a semiquantitative, three-point scale (mild, moderate, and severe). Caspase-3 activation was evaluated similarly in an adjacent section. Area and severity of the damaged lesion were compared between the two staining methods and between magnesium and controls, by chi(2) test and Fisher test. RESULTS: Three-day magnesium administration reduced loss of MAP-2 (from 33 of 50 to 21 of 45, P =.06) and activation of caspase-3 (from 34 of 50 to 21 of 45, P =.04) in the cerebral cortex after HI. There was no significant change in these immunostainings in the hippocampal regions. In the hippocampal CA3, the severity ranked by activated caspase-3 staining was significantly less than that of MAP-2 staining. CONCLUSION: Long-term, post-HI treatment with magnesium inhibited caspase-3 activation and MAP-2 immunostaining, suggesting that magnesium inhibited apoptotic neuronal death of HI in 7-day-old rats. PMID- 12113879 TI - Placental expression of insulin-like growth factor receptor-1 and insulin receptor in the growth-restricted fetal rat. AB - OBJECTIVE: The type 1 insulin-like growth factor receptor (IGF-R1), which resembles the insulin receptor (INS-R), is expressed abundantly in the placenta. The regulation of fetal growth by IGFs and insulin might reflect their actions on the placenta. This study compared the placental expression of IGF-R1 and INS-R in growth-restricted and normal pregnancies. METHODS: Fetal growth restriction was produced in Sprague-Dawley rats by bilateral ligation of the uterine artery on day 19 (term = 21.5 days). Fetuses were delivered by hysterotomy on day 20, and placental samples were frozen. Fetuses that had been subjected to a sham operation of maternal laparotomy without uterine artery manipulation were studied as matched controls. IGF-R1 and INS-R message was assessed by reverse transcription and polymerase chain reaction amplification of extracted placental RNA. The receptors also were assessed by Western blotting of extracted placental protein. Measurements for the growth-restricted group were expressed as a multiple of that of the matched sham litter. RESULTS: Placental IGF-R1 messenger RNA (mRNA) level was significantly lower in the growth-restricted fetuses (0.66 x sham [0.47-0.93], P <.05), but INS-R mRNA was not different (0.89 x sham [0.63 1.3], P >.5). IGF-R1 protein transcript was similarly reduced in the growth restricted fetuses (0.78 x sham [0.69-0.88], P <.005), but the INS-R protein transcript was not (0.98 x sham [0.69-1.4], P >.8). Birth weights were significantly less in the growth-restricted group (3.06 +/- 0.07 versus 3.29 +/- 0.06 g, P =.016); placental weights were not (0.54 +/- 0.02 versus 0.54 +/- 0.02 g, P =.90). CONCLUSIONS: The placenta responds to decreased nutrient delivery with decreased expression of IGF-R1. This would reduce the growth-promoting effects of insulin-like growth factors, which include augmentation of placental lactogen production and glucose and amino acid transport. INS-R was unaffected, which suggests that placental response to insulin is less important than its response to insulin-like growth factors in growth restriction. PMID- 12113880 TI - Interferon gamma antagonizes interleukin-1beta-induced cyclooxygenase-2 expression and prostaglandin E(2) production in human myometrial cells. AB - OBJECTIVE: To evaluate the effect of interferon gamma (IFNgamma) on interleukin 1beta (IL-1) and tumor necrosis factor-alpha (TNFalpha)-promoted prostaglandin E(2) (PGE(2)) production and to investigate the interaction of IFNgamma and IL-1 on cyclooxygenase-2 (COX-2) expression, as well as nuclear factor-kappaB (NF kappaB) activation in human myometrial cells. METHODS: An immortalized human myometrial cell line was cultured in Dulbecco modified Eagle medium (DMEM) fortified with 10% (v/v) fetal calf serum (FCS) in multiwell plates until near confluency. Twenty-four hours before the start of the experiments, the medium was replaced with FCS-free medium containing 0.5% bovine serum albumin. Prostaglandin E(2) was determined in the medium with a specific radioimmunoassay having a sensitivity of 10 pg. The COX-2 and NF-kappaB inhibitory protein (IkappaB) protein levels were measured in cell extracts by Western blot. RESULTS: Cell cultures primed with IFNgamma produced significantly less (P <.05-.01) PGE(2) in response to cytokines than cells exposed to IL-1, TNF-alpha, or the combination of the two. This result corresponded to a similar inhibition of IkappaB degradation (a prerequisite of NF-kappaB activation) as well as COX-2 protein steady state levels. CONCLUSION: Interferon gamma acts as a partial antagonist of IL-1 signaling in this cell model at a site upstream from the activation of the NF-kappaB pathway, causing a partial inhibition of COX-2 expression and PGE(2) production. PMID- 12113882 TI - Estrogen receptor alpha and progesterone receptor A and B concentration and localization in the lower uterine segment in term parturition. AB - OBJECTIVE: To determine the localization and concentrations of estrogen receptor alpha and progesterone receptors A and B in the lower uterine segment during term parturition. METHODS: Biopsies were taken from 70 patients during nonelective cesarean delivery. The patients were at different stages of cervical dilatation (<2 cm, 2-3.9 cm, 4-6 cm, >6 cm) and different duration of labor (< or =6 hours, >6-12 hours, >12 hours). The receptor concentrations were determined with solid phase immunoassays, and their localization was investigated immunohistochemically. RESULTS: Estrogen receptor alpha concentration decreased significantly from 2.12 fmol/mg protein at less than 2 cm dilatation to 1.08 fmol/mg (4-6 cm) but tended to increase at greater than 6 cm. Progesterone receptor A and B concentration was 84.7 fmol/mg at less than 2 cm dilatation, decreased significantly to 36.6 fmol/mg (2-3.9 cm), and increased again with further dilation. Concentrations of both receptors did not depend on duration of labor. By immunohistochemistry only progesterone receptor A and B was found to be expressed by endothelial and smooth muscle cells of the vessels, stromal fibroblasts, smooth muscle cells in the myometrium, and glandular epithelial cells. Regardless of the extent of cervical dilatation, expression of progesterone receptors A and B was marked. CONCLUSION: A decrease in estrogen receptor alpha and progesterone receptor A and B concentration in the early phase of first stage labor may play a role in cervical dilation at term. PMID- 12113881 TI - Effects of intra-amniotic endotoxin on lung structure and function two months after term birth in sheep. AB - OBJECTIVE: Intra-amniotic endotoxin causes chorioamnionitis and results in improved lung function after preterm delivery in sheep. Our aim was to determine the effects on lung structure and function after term birth in lambs exposed to intra-amniotic or intra-allantoic endotoxin. METHODS: At 119 days' gestation, pregnant ewes bearing singleton fetuses received intra-amniotic or intra allantoic injections of either saline (allantoic, n = 1; amniotic, n = 10) or Escherichia coli (055:B5) endotoxin (allantoic, n = 5; amniotic, n = 7). Amniotic or allantoic fluid was aspirated for white blood cell counts 24 hours after the injections. Ewes (n = 20) were allowed to deliver spontaneously. At 8 weeks' postnatal age we measured ventilation, lung volumes, and compliance in the offspring and collected their lungs for morphologic analysis. RESULTS: Higher amniotic or allantoic cell counts were confined to the fluid space into which endotoxin was injected. Saline injections did not increase amniotic or allantoic white blood cell counts. Gestation length, birth weight, and postnatal growth were unaffected by endotoxin treatments. Lung volumes and compliances at 8 weeks of age were not different between saline-treated and endotoxin-treated groups. Lung morphometry was not significantly altered by endotoxin, with one minor exception: interlobular septal volume was increased by intra-amniotic endotoxin, but this effect had no functional consequences. CONCLUSIONS: Intra-amniotic or intra-allantoic endotoxin caused a localized inflammatory response but did not cause preterm delivery or intrauterine growth restriction. The functional improvements and corresponding structural alterations in the lungs of preterm lambs, reported previously in this model, were not associated with improved or impaired lung function or marked alterations in lung structure 2 months after birth at term. PMID- 12113883 TI - Levels of maternal plasma corticotropin-releasing factor and urocortin during labor. AB - OBJECTIVE: Corticotropin-releasing factor (CRF) is produced by the placenta and intrauterine tissues and secreted in increasing amounts from early to term pregnancy. In the presence of labor, a more incisive increase in CRF levels has been described, and women with preterm labor or those destined to have premature delivery have higher midpregnancy CRF levels than those who deliver at term. Urocortin is a 40-amino acid peptide belonging to the CRF family, expressed by human trophoblast and fetal membranes, which has the same biologic effects as CRF. Acting on the same CRF receptors, urocortin stimulates myometrial contractility and ACTH and prostaglandin release from cultured human placental cells. Because no data exist about urocortin levels in the maternal circulation at parturition, we investigated whether maternal plasma urocortin and CRF levels change according to cervical dilatation in healthy pregnant women at term labor. METHODS: In a cross-sectional study of labor, a single maternal blood sample was collected from healthy pregnant women at term (n = 40); in a second longitudinal study, plasma samples were collected longitudinally in a subset of patients (n = 8) throughout labor, according to a Bishop score evaluation. RESULTS: Both maternal plasma CRF and urocortin levels were higher in labor than those previously reported during pregnancy, but they did not change significantly during the different stages of labor when evaluated longitudinally. Some patients showed a trend toward increasing levels, whereas others had variable concentrations. CONCLUSION: Neither CRF nor urocortin levels changed during the progression of spontaneous labor. PMID- 12113884 TI - A pilot study of the effects of phytoestrogen supplementation on postmenopausal endometrium. AB - OBJECTIVE: This study was designed to assess endometrial histology in postmenopausal women not taking hormone replacement therapy, to evaluate side effects and efficacy of phytoestrogens in treating menopause-associated symptoms, and to determine whether 6 months of phytoestrogen supplementation altered endometrial histology. METHODS: We performed a prospective, double-blinded, randomized, placebo-controlled trial comparing the effects of 6 months of dietary phytoestrogen supplementation versus placebo in postmenopausal women. Baseline endometrial biopsies were performed and, if adequate, nonhyperplastic, noncancerous, and nonovulatory, subjects were randomly assigned to receive daily placebo or soy cereal supplementation for 6 months. Study subjects completed baseline and weekly dietary, symptom, and side effect logs. Repeat endometrial biopsies were obtained at 6 months. RESULTS: Subjects were recruited from January 1998 through June 2000. Twenty-seven subjects were randomized, and 19 completed the study. One (3.7%) baseline endometrial sample was weakly proliferative. All other baseline and final biopsies were consistent with atrophic, inactive endometrium. The maximum risk of endometrial stimulation with phytoestrogens is 35%. Hot flushes, night sweats, and vaginal dryness were significantly less severe at the final week of the study compared with baseline in the placebo group. Insomnia was more common in the treated group. There were no other statistically significant differences in symptoms or side effects. CONCLUSION: Phytoestrogens did not cause stimulation of the endometrium. Insomnia was more frequent over the 6-month study in the soy group, whereas hot flushes, night sweats, and vaginal dryness improved from baseline in the placebo group but not in the soy group. PMID- 12113885 TI - Regulation of Fas ligand expression by estrogen in normal ovary. AB - OBJECTIVE: The clinical significance of the Fas/Fas ligand (FasL) system in hormone-sensitive carcinomas such as breast and ovary has been reported. However, only a few studies have investigated the potential hormonal regulation of its expression. In this study, we evaluated the expression of FasL in normal ovarian tissue during the normal female reproductive cycle with the goal of identifying potential hormones that can regulate FasL expression. METHODS: We used Western blot analysis to examine the expression of FasL in the rat ovary throughout the natural estrous cycle. We employed Western blot and reverse transcriptase polymerase chain reaction to study hormonal regulation of FasL in human ovarian epithelial cells and normal ovarian tissues. RESULTS: FasL protein expression levels change in the ovary during the female reproductive cycle. FasL protein appeared intensively in estrus, declined sharply in metestrous, further decreased to a very low level in diestrus, and was absent in proestrus. Because the protein expression pattern of FasL in the cycling ovary was similar to the estrogen receptor beta expression pattern, we examined the effect of estrogen on the level of FasL protein and found that estrogen indeed upregulates the expression of FasL protein and mRNA levels in ovarian epithelial cells as well as in normal ovarian tissues. Furthermore, we showed that the estrogen-induced increase in the FasL protein and mRNA levels could be abolished by 4-hydroxytamoxifen, which suggests that the observed increase in FasL expression was mediated by estrogen receptor. CONCLUSION: Our findings support the hypothesis that the expression of FasL in normal ovary is hormonally sensitive and could have a key role in the physiology of normal ovarian tissue. PMID- 12113886 TI - Synthesis of analogs of L-valacyclovir and determination of their substrate activity for the oligopeptide transporter in Caco-2 cells. AB - L-Valacyclovir, a prodrug of acyclovir, is a substrate for the peptide transporter (PepT1) in the intestinal mucosa, which accounts for its higher than expected oral bioavailability. The substrate activity of L-valacyclovir for PepT1 is surprising, particularly when one considers that the molecule has the structural features of a nucleoside rather than a peptide. In an attempt to better understand the structure-transport relationships (STR) for the interactions of L-valacyclovir with PepT1, analogs of this molecule with structural changes in the guanine moiety were synthesized and their substrate activity for PepT1 in Caco-2 cell monolayers was determined. The analogs synthesized include those that had the guanine moiety of L-valacyclovir substituted with purine, benzimidazole, and 7-azaindole. All three analogs (purine, benzimidazole, and 7-azaindole) exhibited affinity for PepT1 in binding studies, but only the purine analog (as the L-valine ester) showed PepT1 associated transcellular transport across Caco-2 cell monolayers. The benzimidazole and 7-azaindole analogs (as their L-valine esters) were rapidly metabolized by esterase when applied to the apical surface of Caco-2 cells, which probably explains their low penetration as the intact prodrugs via PepT1. PMID- 12113887 TI - A convenient synthesis by microwave heating and pharmacological evaluation of novel benzoyltriazole and saccharine derivatives as 5-HT(1A) receptor ligands. AB - A series of novel 1,2,3-4-benzoyltriazole and saccharine derivatives were designed and synthesized by microwave heating. They were evaluated on a battery of receptors, including serotonin 5-HT(1A,) 5-HT(2A) and 5-HT(2C), and the most interesting compounds were further evaluated on dopaminergic D(1), D(2) and adrenergic alpha(1), alpha(2) receptors. Conventional and microwave heating of the reactions were compared. Synthesis by microwave heating gave the desired compounds in better yields than those obtained by conventional heating. The overall times for the syntheses were considerably reduced. All compounds displayed moderate affinity for 5-HT(1A) receptor. The most interesting compound 33 showed a high affinity (K(i)=93 nM) which was combined with no affinity on the other receptors considered. PMID- 12113888 TI - Tissue-specific regulation of expression and activity of P-glycoprotein in adjuvant arthritis rats. AB - Cyclosporine A and steroids are effective against rheumatoid arthritis and also known as substrates of P-glycoprotein (P-gp). We investigated the effect of arthritis on the hepatic and intestinal P-gp activity in rats, and substantiated the expression level of the hepatic P-gp. Doxorubicin was used as a P-gp substrate. Cumulative biliary excretion and intestinal exsorption of doxorubicin following intravenous administration were compared between adjuvant arthritis (AA) and normal rats. Intestinal P-gp activity was also investigated by intestinal everted sac method, and hepatic P-gp was detected by FITC-labeled antibody and visualized using a confocal laser microscope system. Biliary clearance of doxorubicin in AA rats was significantly decreased from that in normal rats. The expression level of the hepatic P-gp in AA rats was very low compared to normal rats, indicating down-regulation. Intestinal exsorption clearance was not different between AA and normal rats. Permeability of doxorubicin across intestinal everted sac was comparable between AA and normal rats, corresponding to in vivo study. In AA rats, hepatic P-gp activity was decreased due to the reduction of expression level, but intestinal P-gp activity was not changed. Different regulation systems may be involved in liver and intestine. PMID- 12113889 TI - Evaluation of the surfactant properties of ascorbyl palmitate sodium salt. AB - In this paper we report on the physicochemical surface properties of ascorbyl palmitate (Asc16) and of its sodium salt (Asc16Na) with a view to their use as surfactants. Asc16Na was synthesized from ascorbyl palmitate by neutralizing the OH groups in position 3 of the ascorbyl ring. The acid-base properties, thermal analysis and stability of Asc16Na monomers were determined. Self-assembling parameters of micellar aggregates in aqueous dispersions through critical micellar concentration (CMC) and critical micellar temperature (CMT) were measured. Asc16Na micellar dispersions efficiently solubilize poorly soluble drugs such as phenacetin and griseofulvin, and enhance their apparent solubility in aqueous environments. Stability tests showed that Asc16Na is more unstable than ascorbyl palmitate. Ascorbyl palmitate and its sodium salt are insoluble at room temperature in water, but their solubilities strongly depend on temperature, and largely increase above the CMT. Although Asc16Na is insoluble at room temperature, it is more soluble than Asc16, and its CMT significantly lowers in the undissociated acidic form. The apparent solubilities of phenacetin and griseofulvin are increased in Asc16Na aqueous solutions. The Asc16Na potential use as surfactant is restricted by its low stability in water, therefore the addition of some antioxidant species is necessary. PMID- 12113890 TI - Evaluation of the role of intestinal and liver metabolism in the conversion of two different ester prodrugs of sanfetrinem to the parent drug in vitro and in vivo using different rat tissues and a surgically prepared rat model. AB - To improve oral absorption of sanfetrinem, a broad-spectrum, beta-lactamase stable antibiotic, two different ester prodrugs have been selected. Both prodrugs proved to be readily hydrolyzed after absorption before reaching the systemic circulation. The objective of this study was to evaluate the role of intestinal and liver metabolism in the conversion of the two prodrugs into the active compound. In vitro experiments were performed in different rat tissues involved in the absorption process. Moreover data obtained with in vitro experiments have been integrated with data obtained in vivo using a surgically prepared rat model which allows for the measurement of the amount of intact prodrug that overcomes the intestinal mucosa and its presence in the portal vein. Both prodrugs proved to be readily cleaved by jejunum and liver microsomes. The rates of ester hydrolysis with these two tissues were 10- to 30-fold higher than those calculated in intestinal juice at pH 7.4 and about 100-fold higher than in buffer at pH 5.5. These data suggest that both the intestinal wall and liver could play an important role in the conversion of the two prodrugs in active parent compound. In the in vivo experiment, relative to sanfetrinem levels, very low concentrations of intact esters were measured in the portal vein blood, indicating that the two prodrugs are nearly completely hydrolyzed to the active drug by the intestinal wall. In conclusion this study demonstrated that the intestinal epithelium plays a major role in the conversion of the two prodrugs into sanfetrinem. The liver, despite its high esterase activity seems to be only marginally involved. PMID- 12113891 TI - Eudragit RS100 nanosuspensions for the ophthalmic controlled delivery of ibuprofen. AB - Topical application of non-steroidal anti-inflammatory drugs on the eye is a common treatment used to contrast the miosis induced by surgical traumas, such as cataract extraction. With the aim of improving the availability of sodium ibuprofen (IBU) at the intraocular level, IBU-loaded polymeric nanoparticle suspensions were made from inert polymer resins (Eudragit RS100). The nanosuspensions were prepared by a modification of the quasi-emulsion solvent diffusion technique using variable formulation parameters (drug-to-polymer ratio, total drug and polymer amount, stirring speed). Nanosuspensions had mean sizes around 100 nm and a positive charge (zeta-potential of +40/+60 mV), this makes them suitable for ophthalmic applications. Stability tests (up to 24 months storage at 4 degrees C or at room temperature) or freeze-drying were carried out to optimize a suitable pharmaceutical preparation. In vitro dissolution tests indicated a controlled release profile of IBU from nanoparticles. In vivo efficacy was assessed on the rabbit eye after induction of an ocular trauma (paracentesis). An inhibition of the miotic response to the surgical trauma was achieved, comparable to a control aqueous eye-drop formulation, even though a lower concentration of free drug in the conjunctival sac was reached from the nanoparticle system. Drug levels in the aqueous humour were also higher after application of the nanosuspensions; moreover, IBU-loaded nanosuspensions did not show toxicity on ocular tissues. PMID- 12113892 TI - Near-infrared FT-Raman spectroscopy as a rapid analytical tool for the determination of diltiazem hydrochloride in tablets. AB - This study was performed to develop a fast and reliable analytical method for the quantitative determination of diltiazem hydrochloride in tablets. HPLC is currently the preferred method, but is time consuming due to extensive sample preparation. FT-Raman spectroscopy was used to quantitatively analyse diltiazem hydrochloride in commercially available tablets (Tildiem) and in experimental tablets prepared at lab-scale. The percentage of diltiazem hydrochloride in each tablet was determined by calculating-after vector normalisation-the total peak area of the spectral band between 1625 and 1560 cm(-1). No spectral interference from tablet excipients was seen at this spectral band. After FT-Raman spectroscopy the same samples were analyzed based on the HPLC method described in the USP XXIV. The drug dosage per tablet obtained from the vibrational spectroscopy method correlated well with the results obtained using HPLC analysis for both the commercial tablets (HPLC: 63.57+/-0.13 mg; Raman: 63.28+/-0.26 mg (n=50)) and the experimental tablets (HPLC: 181.02+/-0.25 mg; Raman: 181.22+/ 0.35 mg (n=50)). FT-Raman is a reliable alternative for the HPLC method to quantify the amount of diltiazem hydrochloride in tablets. The spectroscopic method is faster because it eliminates sample pre-treatment. Furthermore the FT Raman method has an additional advantage of not requiring solvents. PMID- 12113893 TI - Binding and release of drugs into and from thermosensitive poly(N-vinyl caprolactam) nanoparticles. AB - Three model drug substances, the beta-blocking agents nadolol and propranolol and a choline-esterase inhibitor tacrine, were used in order to determine how different drug molecules affect the behavior of thermally responsive polymer nanoparticles composed of poly(N-vinylcaprolactam) (PVCL). Pure PVCL particles in water exist in a swollen state at room temperature, but the size of the particles decreases discontinuously when the temperature is raised above the volume phase transition temperature. At temperatures above this transition temperature, water is expelled out from the nanoscopic hydrogel particles. Light scattering studies revealed that the more hydrophobic drug substances, propranolol and tacrine, considerably swell the PVCL-microgel. The more hydrophilic drug, nadolol, decreased the transition temperature of PVCL particles, whereas the transition temperature values of pure PVCL particles and that of the added propranolol and tacrine were quite similar. Attenuated drug release results showed that the beta blocking agents were tightly bound to the microgel, and this was more evident at higher temperatures. On the contrary, the release of tacrine across the cellulose membrane was increased when PVCL particles were present. Thus, both physical and chemical properties of the drugs clearly affected their binding to PVCL particles and the release of drugs was affected by the temperature. PMID- 12113894 TI - Monitoring ibuprofen enantiomers released from polymeric systems. AB - Two methacrylic derivatives of ibuprofen (N-[4-[2-(4 isobutylphenyl)propionyloxy]phenyl] methacrylamide (MAI) and 2-[(4 isobutylphenyl)propionyloxy]ethyl methacrylate (MEI)) were used together with 2 hydroxyethyl methacrylate (HEMA) to synthesize four polymeric materials: two hydrophobic homopolymers, PMAI and PMEI, and two hydrophilic copolymers containing 70% (w/w) HEMA, MAI-HEMA 30 and MEI-HEMA 30. The enantiomeric determination of R- and S-IBU released from these four systems has been carried out by capillary electrophoresis. Release of R- and S-IBU was monitored during in vitro assays done at 37 degrees C at pH 7.4 and 10 in buffered solutions and rat plasma. There is a hydrolytical activation in plasma and at pH 10 compared to pH 7.4; moreover, the release rate from the copolymers is much higher than from the homopolymers as a consequence of the greater hydrophilic character. A slight excess of the S-enantiomer of IBU is observed in all the experiments, being more relevant at higher release rates, i.e. copolymers at pH 10. PMID- 12113895 TI - Dose-response measures of rCBF and subjective changes following procaine in healthy female volunteers. AB - The intravenous administration of procaine shows relatively specific activation of limbic structures. Several investigators have utilized this property of procaine to probe limbic system dysfunction in neuropsychiatric disorders. The dose of procaine utilized in human studies varies significantly, however, and the optimal dose of procaine as a limbic probe has not been demonstrated. In two 10 individual groups of healthy female volunteers, we assessed the regional cerebral blood flow (rCBF) response, by single-photon emission computed tomography (SPECT), to saline and 1.38 mg/kg procaine (Group I), and saline, 0.5 mg/kg and 1.0 mg/kg procaine (Group II). Compared to saline, 0.5 mg/kg procaine produced minimal rCBF changes, 1.0 mg/kg procaine induced both limbic and non-limbic activation, and 1.38 mg/kg procaine showed relatively specific rCBF limbic activation. Subjective responses increased in a dose-response manner. We conclude that a dose of 1.38 mg/kg procaine provides a more limited and specific activation of limbic structures than 1.00 mg/kg procaine and thus may be more useful as a specific probe of limbic function. PMID- 12113896 TI - Alterations in MAP2 immunocytochemistry in areas 9 and 32 of schizophrenic prefrontal cortex. AB - A variety of lines of converging evidence implicate the prefrontal cortex (PFC) in schizophrenia. Studies employing Nissl stains have suggested that PFC dendrites may be atrophic in schizophrenia; however, Nissl stains do not reveal dendrites. We employed MAP2 immunocytochemistry, which stains dendrites to examine cortical layers III and V in two areas of the PFC (areas 9 and 32). Occipital cortex (area 17) was examined as a control region. Tissues from seven schizophrenics and seven non-psychiatric controls were examined. Immunostaining was quantitated by area fraction analysis. MAP2 area fraction was decreased in both layers in both regions of PFC, but not in occipital cortex. Area 9 exhibited a 42% reduction in layer V and a 36% reduction in layer III. Area 32 exhibited a 31% reduction in layer V and a 36% reduction in layer III. Neither region exhibited a significant change in the density of pyramidal cells. These data are consistent with the hypothesis of a schizophrenia-associated decrease in dendritic material in the PFC. PMID- 12113897 TI - MR signal intensity of gray matter/white matter contrast and intracranial fat: effects of age and sex. AB - Signal intensity (SI) values of gray- and white-matter brain regions of interest (ROIs) were obtained from T(2)- and proton density-weighted magnetic resonance (MR) images of 58 normal subjects aged 22-82 years (31 females, 52.3+/-18.8 years; 27 males, 54.1+/-18.1 years). Sampled ROIs included the caudate, putamen, thalamus, orbitofrontal gyrus, gyrus rectus, uncus, frontal white matter, anterior and posterior corpus callosum, cranial-cervical junction fat, and retroorbital fat. Effects of age and sex on SI were examined using repeated measures analysis of covariance. For both T(2)- and proton density-weighted acquisitions, a significant inverse relationship between age and SI was observed for the ratio of all summed gray-matter ROIs divided by summed white-matter ROIs. This relationship was additionally observed for ratios of both subcortical gray/white matter and cortical gray/white matter. Females compared with males had significantly lower cortical gray/white matter ratios on T(2)-weighted scans. Differences in SI were observed between cranial-cervical junction fat and retroorbital fat on both acquisitions, with females showing significantly higher values for cranial-cervical junction fat and males showing higher values for retroorbital fat. Implications for brain morphometry, the use of fat as a reference standard, and other issues in neuroimaging are discussed. PMID- 12113898 TI - Occipital hypoperfusion in a patient with psychogenic visual disturbance. AB - A patient with bilateral diminished visual acuity and blurred vision was given the diagnosis of psychogenic visual disturbance after extensive psychiatric and ophthalmological examinations. Single photon emission computed tomography with Tc 99m-ethylcysteinate dimer demonstrated remarkably reduced regional cerebral blood flow (rCBF) in the bilateral occipital lobes. More specifically, rCBF was reduced in the visual association areas, but not in the primary visual areas. These findings suggest that functional suppression of the visual association area is associated with the development of psychogenic visual disturbance. PMID- 12113899 TI - Haloperidol abolished glutamate release evoked by photic stimulation of the visual cortex in rats. AB - There is evidence that systemic administration of haloperidol, a dopamine receptor blocker, attenuates visual cortex evoked potentials. However, there is scarce information on cortical neurochemical changes associated with haloperidol effects on visual function. The present experiment was designed to investigate: (1) the effect of photic stimulation on glutamate release in the visual cortex; and (2) whether systemic administration of haloperidol would affect those neurochemical changes. Microdialysis probes were implanted in the occipital cortex. Glutamate levels were measured every 30 s using capillary zone electrophoresis. Extracellular glutamate levels increased to about 282% 30 s after photic stimulation started and remain elevated for the 3 min that the photic stimulation lasted. Haloperidol (1.5 and 5 mg/kg, i.p.) completely suppressed the increased of glutamate efflux during photic stimulation. Finally, it was also found that the highest dose of haloperidol (5 mg/kg) did not change glutamate basal levels. The results are discussed with reference to possible dopaminergic actions on the visual system function. PMID- 12113900 TI - Calcium currents in rat entorhinal cortex layer II stellate and layer III pyramidal neurons in acute brain slice. AB - The entorhinal cortex (EC) is an essential relay station in neocortical - hippocampal information transfer and memory functions. Layer II stellate and layer III pyramidal neurons show specific damage in Alzheimer's disease and epilepsy, respectively. Using whole cell patch clamp recording in rat brain slices we here demonstrate that high voltage activated Ca(2+)-currents (I(Ca)(2+)) are about 1.6-fold bigger in stellate cells than in pyramidal neurons while current density is equal in both cell types. In stellate cells I(Ca)(2+) shows stronger inactivation with depolarization, block of I(Ca)(2+) by Ni(2+) (300 and 600 microM) is more effective, and this block decreases more for currents evoked from a less negative holding potential than in layer III pyramidal neurons. These data indicate distinct molecular composition of Ca(2+) channels and can partially explain stronger increases of [Ca(2+)](i) during 10 Hz firing activity in EC pyramidal versus stellate neurons. PMID- 12113901 TI - Developmental organization of the glossopharyngeal nucleus in the embryonic chick brainstem slice as revealed by optical sectioning recording. AB - We traced developmental changes in the ventro-dorsal distribution pattern of glossopharyngeal nerve (N. IX) responses by applying an optical sectioning method to thick slice preparations dissected from E4 to E8 chick embryos. We identified the motor and sensory nuclei related to the glossopharyngeal nerve in the rostral and caudal focal planes, respectively. In the E4 and E5 preparations, the motoneuronal responses appeared on the central part of the stimulated side of the brainstem. As development proceeded to E6, the response area became localized on the dorsal region. The change in the ventro-dorsal distribution pattern was similar to that observed in the vagus nerve-related nuclei, suggesting that there might be an essential process underlying functional organization of the brainstem nuclei. PMID- 12113902 TI - Cholera toxin B subunit labeling in lamina II of spinal cord dorsal horn following chronic inflammation in rats. AB - We have investigated the effect of inflammation on the labeling pattern of cholera toxin B subunit (CTB)-conjugated horseradish peroxidase, an A-fiber marker, by an intra-sciatic nerve injection of the tracer. Following chronic inflammation in one hind paw in rats, there was substantial CTB labeling in lamina II of the spinal dorsal horn, which is normally absent. However, there was no change in the labeling pattern of wheat germ agglutinin or fluoride resistant acid phosphatase/thiamine monophosphatase, two C-fiber markers. The CTB labeling in lamina II after peripheral nerve injury has been interpreted as central sprouting of A-fibers or uptake of the tracer by injured C-fibers. Our results suggest that chronic inflammation and nerve injury may share some common mechanisms in generating allodynia and hyperalgesia. PMID- 12113903 TI - Long-term potentiation and adenosine sensitivity are unchanged in the AS/AGU protein kinase Cgamma-deficient rat. AB - The AS/AGU rat is a spontaneously occurring mutation which exhibits locomotor abnormalities, reduced tyrosine hydroxylase levels in substantia nigra and lower extracellular levels of dopamine, making it a valuable model for some human locomotor disorders, and spontaneous chronic degeneration. The molecular defect is an absence of protein kinase Cgamma (PKCgamma), an enzyme suggested to play a role in synaptic plasticity. We have therefore examined long-term potentiation (LTP) in hippocampal slices from the mutant animals compared with the normal control strain of Albino Swiss rat. In the CA1 region, LTP was of the same magnitude in mutant and control animals, and the presynaptic inhibitory effects of adenosine were unchanged in naive slices or following LTP. Paired-pulse inhibition and facilitation were normal. It is concluded that the absence of PKCgamma in this strain does not modify synaptic plasticity or presynaptic sensitivity to adenosine. PMID- 12113904 TI - Absence of painful neuropathy after chronic oral fluoride intake in Sprague Dawley and Lou/C rats. AB - The possibility that chronicle oral ingestion of fluoride-rich water could modify peripheral pain sensitivity was studied in two strains of adult rats, Sprague Dawley and Lou/C rats. Sodium fluoride was given orally in water to male Sprague Dawley (75 and 150 ppm) and Lou rats (150 ppm) for 15 and 27 weeks, respectively. Using classical behavioural evaluation methods of pain symptoms, only slight tendencies to a thermal hyperalgia and a mechanical allodynia were observed in Sprague-Dawley rats. PMID- 12113905 TI - P-Glycoprotein-mediated efflux of phenobarbital, lamotrigine, and felbamate at the blood-brain barrier: evidence from microdialysis experiments in rats. AB - Although a series of new antiepileptic drugs (AEDs) have been launched in the last two decades, drug-refractoriness remains a major problem concerning 20-30% of epileptic patients. The fact that most patients with refractory epilepsy are resistant to several AEDs acting via different targets points to an involvement of unspecific mechanisms like changes in local uptake of AEDs in the epileptic focus region. Increased expression of multidrug transporters has been reported in epileptogenic brain tissue from pharmacoresistant patients undergoing epilepsy surgery. However, only limited information exists on the extent to which AEDs are transported by multidrug transporters like P-glycoprotein (PGP). In the present study, the effect of PGP inhibition by verapamil on brain access of the AEDs phenobarbital, lamotrigine, and felbamate was investigated by in vivo microdialysis in rats. Local perfusion of verapamil via the microdialysis probe increased the concentration of the three AEDs in the extracellular fluid of the cerebral cortex in a significant manner. The data indicate that overexpression of PGP in epileptic tissue is likely to limit brain access of the AEDs phenobarbital, lamotrigine, and felbamate, thus favoring the hypothesis that multidrug transporters play a crucial role in the phenomenon of drug-refractory epilepsy. PMID- 12113906 TI - Relationship among (1)H-magnetic resonance spectroscopy, brain volumetry and genetic polymorphisms in humans with memory impairment. AB - We investigated the relationship among neuroanatomical, neurochemical and genetic variables in 44 subjects with age-related memory impairment. Hydrogen magnetic resonance spectroscopy was used to determine N-acetyl/creatine (NAA/Cr) concentrations in basal ganglia and medial temporal regions. Volumetric measures were obtained for caudate nucleus and hippocampus. Genetic polymorphisms examined included apolipoproteins (APO) E and CI, angiotensin converting enzyme and dopamine D2 receptor TaqI genes. Age was found to be negatively correlated with hippocampal and basal ganglia volumes, but not with neurochemical values. Multiple regression analyses showed that the APOC1 polymorphism was the only variable which predicted NAA/Cr values in basal ganglia. NAA/Cr metabolites in the medial temporal lobe but not in the basal ganglia region were related with lower performance in verbal memory. PMID- 12113907 TI - Cerebral transcriptome analysis of transgenic mice overexpressing erythropoietin. AB - Erythropoietin (EPO) and its receptor are expressed in the brain, and one of its roles appears to be neuroprotection. This study investigates whether chronic overexpression of EPO changes brain mRNA expression in the brains of transgenic mice. Therefore, cerebral mRNA expression was investigated in transgenic mice overexpressing EPO. Microarray analysis revealed an upregulation (2.8- to 3.6 fold) of N-acetylglucosamine-6-O-sulfotransferase (prolongation of the EPO effect), a translocase of the inner mitochondrial membrane (mitochondrial matrix import of nuclear encoded proteins), a mitochondrial ribosomal protein (mitochondrial protein translation), and a peroxisomal biogenesis factor (formation of peroxisomes). In conclusion, components of oxidative metabolism pathways were activated at the level of transcription which could be related to neuroprotective effects of EPO or could indicate compromised tissue. PMID- 12113908 TI - Cortico-muscular coupling in a human subject with mirror movements--a magnetoencephalographic study. AB - We studied cortico-muscular coupling in a 15-year-old male suffering from congenital mirror movements (MMs) of hands. Cortex-muscle coherence was analyzed between magnetoencephalographic signals and the electromyograms (EMGs) recorded from both hands and feet during uni- and bilateral isometric contractions. Regardless of the side of the intended contraction, the motor cortex contralateral to the contraction was coupled to the muscles of both hands at 20 25 Hz. No coupling was found from the other, ipsilateral hemisphere. EMGs of the two hands were coupled during both intended uni- and bilateral contractions, but only during unilateral contractions could the coupling solely be explained by cortical activation. We suggest that our subject's MMs result from activation of an ipsilateral corticospinal projection, with involvement of additional synchronizing mechanisms at the subcortical, brainstem, or spinal level during bilateral contraction. PMID- 12113909 TI - CpG-oligodeoxynucleotide rejection of a neuroblastoma in A/J mice does not induce a paraneoplastic disease. AB - Oligodeoxynucleotides containing CpG motifs (CpG-ODN) are powerful immunostimulating agents that are currently entering clinical trials in various human diseases. Concerns exist about potential auto-immune diseases triggered by such treatment. We thus investigated whether tumor rejection induced by CpG-ODN treatment could lead to a harmful auto-immune reaction against the nervous system (neurological paraneoplastic disease) at the time of acute tumor rejection, or in long-term surviving animals. Mice bearing established neuroblastomas were treated with intra-tumoral injections of CpG-ODN, resulting in tumor inhibition and tumor rejection in one-third of the animals. Immunocytochemistry and Western blot studies revealed no specific anti-neuronal antibodies. None of the animals developed neurological disabilities and histological studies of the nervous system were normal. CpG-ODN can therefore trigger neuroblastoma rejection without inducing neurological paraneoplastic disease. PMID- 12113910 TI - Apelin-immunoreactivity in the rat hypothalamus and pituitary. AB - With the use of an antiserum against human apelin-36, apelin-immunoreactivity (irAP) was detected in neurons and cell processes of the supraoptic nucleus (SO), paraventricular nucleus (PVH), accessory neurosecretory nuclei (Acc) and suprachiasmatic nucleus. Strongly labeled cells/processes were noted in the internal layer of the median eminence, infundibular stem, anterior and posterior pituitary. Double-labeling the sections with goat polyclonal neurophysin I antiserum and rabbit polyclonal apelin-antiserum revealed a population of magnocellular neurons in the PVH, SO and Acc expressing both irAP and neurophysin I-immunoreactivity (irNP), the latter being a marker of oxytocin-containing neurons. By inference, the AP-positive but irNP-negative magnocellular neurons could be vasopressin-containing. The presence of irAP in certain hypothalamic nuclei and pituitary suggests that the peptide may be a signaling molecule released from the hypothalamic-hypophysial axis. PMID- 12113911 TI - Activation of spinal group I metabotropic glutamate receptors in rats evokes local glutamate release and spontaneous nociceptive behaviors: effects of 2 methyl-6-(phenylethynyl)-pyridine pretreatment. AB - Intrathecal (i.t.) administration of the group I metabotropic glutamate receptor (mGluR) agonist (RS)-3,5-dihydroxyphenylglycine ((RS)-3,5-DHPG) to rats produces an immediate display of spontaneous nociceptive behaviors (SNBs) persisting for up to 10 h after injection (NeuroReport 7 (1996) 2743). The mechanisms underlying these behavioral effects are not entirely understood but may include enhanced release of glutamate within the dorsal horn of the spinal cord. The current experiments used microdialysis in awake moving animals to test: (1), whether i.t. (S)-3,5-DHPG increases the local release of glutamate at doses that also induce SNBs; and (2), whether the effects on glutamate release (as well as SNBs) can be blocked by pretreatment with the mGluR5 selective antagonist 2-methyl-6 (phenylethynyl)-pyridine (MPEP). Male Sprague-Dawley rats were implanted with a microdialysis probe inserted into the i.t. space of the spinal cord (J. Neurosci. Methods 62 (1995) 43) and then tested under i.t. drug conditions (0.01, 0.1 and 1 mM (S)-3,5-DHPG) following a 2-3 day recovery period. As predicted, local application of (S)-3,5-DHPG via the microdialysis probe increased the release of glutamate in a dose-dependent manner. Significant SNBs were also noted in the 0.1 and 1 mM groups in a manner paralleling the onset and duration of the glutamate response. Pretreatment with MPEP (55 mg/kg, intraperitoneally) blocked glutamate release to the 0.1 mM dose of (S)-3,5-DHPG, and also decreased the proportion of animals displaying SNBs in this dose group. No effects of MPEP were seen against the higher dose of (S)-3,5-DHPG (1 mM). These results suggest that stimulation of spinal mGluR5 leads to glutamate release within the spinal cord, a response that may in part account for the nociceptive behaviors evoked by i.t. (S)-3,5-DHPG. PMID- 12113912 TI - Entrainment impaired, masking spared: an apparent genetic abnormality that prevents circadian rhythm entrainment to 24-h lighting cycles in California mice. AB - Lighting cycles can influence the expression of daily activity rhythms in two ways: by entraining the circadian pacemaker that normally drives this rhythm, and by directly affecting the expression of activity itself, thereby 'masking' the influence of the pacemaker. We describe a California mouse (Peromyscus californicus) in which these processes are dissociated. Circadian rhythms of wheel-running activity were recorded continuously while the animal was housed in a standard light/dark cycle and in constant darkness. This animal expressed a normal circadian rhythm that failed to entrain to the light/dark cycle, but was completely masked during the light phase. This animal's phenotype appears to have a genetic basis, since the progeny of selective crosses of his descendants showed similar abnormalities. These mice are the first example of animals expressing apparently normal circadian rhythms that are not entrained by light, but that still show potent masking responses to light exposure. PMID- 12113913 TI - Activation of mitochondrial ATP-sensitive potassium channels prevents neuronal cell death after ischemia in neonatal rats. AB - Activation of mitochondrial ATP-sensitive potassium channels (mK(ATP)) has been shown to protect against cell death following ischemia/reperfusion in the heart but not in brain. We examined whether mK(ATP) activation with diazoxide (DIZ) prevents neuronal cell death following hypoxia-ischemia (HI) in 7-day-old rat pups. Rat pups were subjected to HI (left carotid ligation; 8% O(2); 2.5 h), following administration of vehicle, 1.9 mg/kg DIZ, 3.8 mg/kg DIZ or DIZ plus 10 mg/kg 5-hydroxydecanoic acid (mK(ATP) antagonist). Total infarct volume was reduced from 99.8+/-2.7% in vehicle animals to 80.6+/-4.2% in 3.8 mg/kg DIZ treated animals (n=85, P<0.05). Western blotting showed K(ATP) subunits concentrated in mitochondria. Fluorescent studies indicated DIZ directly depolarized the mitochondria. In conclusion, selective opening of mK(ATP) prior to HI results in neuroprotection in immature rats. PMID- 12113914 TI - Facilitation of the spinal H-reflex by auditory stimulation in dyslexic adults. AB - Dyslexic subjects show a variety of mild sensory and motor deficits that have been assumed to reflect dysfunction of the large-diameter 'magnocells' in different parts of the brain. Hearing as a warning sense relies on rapidly conducting fibers, and on the basis of the magnocellular deficit theory, we wondered whether auditory alerting would be weakened in dyslexic adults. We quantified the strength of sound-induced spinal facilitation in seven dyslexic and eight normal-reading adults by measuring the amplitudes of H-reflex, a monosynaptic spinal reflex, after loud binaural sounds. The audiospinal facilitation was of similar strength in dyslexic and control adults, indicating normal auditory alerting via cerebrospinal pathways. The slightly prolonged facilitation in dyslexics agrees with the dyslexics' general sluggishness of sensorimotor processing. PMID- 12113916 TI - Increased prevalence of type 2 diabetes mellitus among psychiatric inpatients with bipolar I affective and schizoaffective disorders independent of psychotropic drug use. AB - BACKGROUND: There are numerous reports of abnormal glucose metabolism, including increased rates of type 2 diabetes mellitus, in psychiatric patients. It remains unclear, however, whether there is an intrinsic relationship between abnormal glucose metabolism and particular psychiatric disorders, because the relationship is complicated by treatment with psychotropic medications that promote weight gain and hyperglycemia. This study aimed to clarify this relationship. METHODS: The medical records of 243 inpatients, aged 50-74 years, with diagnoses of major depression, bipolar I disorder, schizoaffective disorder, schizophrenia, and dementia were reviewed. Psychiatric and type 2 diabetes mellitus diagnoses, medications, body mass index (BMI), age, gender, and race were recorded. Diabetes rates were compared to age, race, and gender-matched rates in the US general population. RESULTS: Rates of type 2 diabetes mellitus were: schizoaffective (50%) > bipolar I (26%) > major depression (18%) = dementia (18%) > schizophrenia (13%) (p < 0.006). Diabetic patients had a higher mean BMI (p = 0.01), but not a significantly higher use of psychotropic medications previously reported to be associated with new-onset type 2 diabetes (e.g., phenothiazines, clozapine, olanzapine). Logistic regression revealed that psychiatric diagnosis and BMI were the only significant and independent predictors of diabetes diagnosis. Compared to national norms, diabetes rates were significantly elevated only in bipolar I affective and schizoaffective patients. LIMITATIONS: This study was a retrospective chart review of older, hospitalized patients. CONCLUSIONS: This is the first published study to show an increased prevalence of type 2 diabetes mellitus among psychiatric patients with particular psychiatric illnesses independent of the effects of age, race, gender, medication, and body mass. This finding, which requires replication in a larger scale, prospective study, suggests an intrinsic relationship between abnormal glucose metabolism and bipolar I affective and schizoaffective disorders. PMID- 12113917 TI - Bipolar-panic comorbidity in the general population: prevalence and associated morbidity. AB - BACKGROUND: The goal of this study was to determine the prevalence, psychiatric comorbidity, panic symptomotology and physical morbidity associated with the co occurrence of bipolar disorder and panic attacks in the general population. METHOD: Data were drawn from the National Comorbidity Survey, a community-based household sample (n=8098) representative of the US adult population aged 15-54. F tests were used to determine differences in sociodemographic characteristics, panic symptoms, physical and psychiatric morbidity between individuals with bipolar disorder with and without co-occurring panic attacks (lifetime). Multiple logistic regression analyses were used to identify sociodemographic and psychiatric correlates of the co-occurrence of bipolar disorder and panic attacks. RESULT: The co-occurrence of bipolar disorder and panic attacks was associated with earlier onset of panic attack [17.1 (8.7) years vs. 22.0 (10.3) years, F=8.3, df=1, 429, P=0.004] and significantly greater panic symptomotology (P<0.0001). Comorbid substance dependence [OR=5.8 (2.6, 13.0)], specific phobia [OR=3.3 (1.4, 7.6)], and GAD [OR=2.9 (1.3, 6.8)] were each independent correlates of the co-occurrence bipolar disorder and panic attacks. CONCLUSIONS: These findings are consistent with, and extend, results from clinical studies showing that panic attacks are not uncommon, and are associated with significantly elevated rates of comorbid psychopathology, among individuals with bipolar disorder in the general population. Future studies that examine the relationship between panic attacks and bipolar disorder using a longitudinal design may be helpful in improving our understanding of the mechanism of this association. PMID- 12113918 TI - Psychiatric treatment seeking and psychosocial impairment among young adults with depression. AB - BACKGROUND: We report data on 1-year prevalence and comorbidity of depression, related impairment, treatment need, and psychiatric treatment among young adults. METHODS: A sample of young urban adults (n=245) mean age 21.8 years was screened from a baseline population of 706 high-school students and given a semistructured clinical interview to evaluate 12-month prevalence of depression, psychosocial functioning according to DSM-IV GAF scale, need for psychiatric treatment, and use of mental health services. RESULTS: One in 10 young adults suffered from depression with associated psychosocial impairment, the female-to-male-ratio being approximately 2:1. Most depressive disorders were comorbid with other DSM IV disorders, depression usually occurring secondary to other disorders. Comorbidity was related to impairment, treatment need, and treatment contacts. Less than half of the depressed young adults had ever contacted mental health services, and less than one-third reported treatment contacts during the index episode. Males were less likely than females to report previous treatment contacts or intention to refer to mental health services for their problems, but treatment contacts during the index episode were reported equally often by both sexes. CONCLUSIONS: A minority of the severely depressed young adults with associated impairment had sought treatment. Except for subjects with dysthymia, no gender difference emerged in treatment contact rates during the 12-month depression episode. Comorbidity showed important clinical implications by its relation to severity of depression and treatment contacts. PMID- 12113915 TI - Are the neural substrates of memory the final common pathway in posttraumatic stress disorder (PTSD)? AB - A model for the posttraumatic stress disorder (PTSD) as a disorder of memory is presented drawing both on psychological and neurobiological data. Evidence on intrusive memories and deficits in declarative memory function in PTSD-patients is reviewed in relation to three brain areas that are involved in memory functioning and the stress response: the hippocampus, amygdala, and the prefrontal cortex. Neurobiological studies have shown that the noradrenergic stress-system is involved in enhanced encoding of emotional memories, sensitization, and fear conditioning, by way of its effects on the amygdala. Chronic stress also affects the hippocampus, a brain area involved in declarative memories, suggesting that hippocampal dysfunction may partly account for the deficits in declarative memory in PTSD-patients. Deficits in the medial prefrontal cortex, a structure that normally inhibits the amygdala, may further enhance the effects of the amygdala, thereby increasing the frequency and intensity of the traumatic memories. Thus, by way of its influence on these brain structures, exposure to severe stress may simultaneously result in strong emotional reactions and in difficulties to recall the emotional event. This model is also relevant for understanding the distinction between declarative and non declarative memory-functions in processing trauma-related information in PTSD. Implications of our model are reviewed. PMID- 12113919 TI - Social adjustment in in-patients with affective disorders: predictive factors. AB - We assessed social adjustment in 145 depressed in-patients using the self reporting Social Adjustment Scale (42-item version) to evaluate the contribution of demographic and clinical variables and examine social functioning at different levels of depression. Our results indicate that the presence of a psychopathology in association with interpersonal sensitivity, hostility and perceived social support aspects -- and not the severity of current depressive symptoms -- were the most important factors affecting social adjustment. As expected, social disturbances are more pronounced in severe depressives who experience difficulties in all areas: by contrast, patients with low depressive symptom levels do not appear to be maladjusted, by comparison with a community sample. PMID- 12113920 TI - Deliberate self harm patients with depressive disorders: treatment and outcome. AB - BACKGROUND: Depression is the most common psychiatric disorder in deliberate self harm (DSH) patients and in those who commit suicide. The aim of this study was to examine the treatment received by DSH patients with depression and their progress following DSH. METHODS: A representative sample of 106 patients with an ICD-10 depressive episode who presented to a general hospital following an episode of DSH were investigated in terms of their treatment before and after the episode and their outcome at follow-up. RESULTS: Prior to the index episode of DSH, 39 patients (36.8%) were receiving treatment from the psychiatric services and a further 35 (33.0%) were receiving treatment for mental health problems from their general practitioner. Fifty-two patients (49.1%) were receiving antidepressants (in therapeutic dosages in 93.6%). After the episode of DSH 94 patients (88.7%) were offered treatment with the psychiatric services, either as a new referral or continuation of treatment they were receiving prior to DSH. Of the patients who were followed-up, 36.3% remained in contact with the psychiatric services, 52.3% showed poor compliance with recommended treatment and 60.2% no longer fulfilled the diagnostic criteria for depression. Almost one-third reported a further episode of DSH during the follow-up period. LIMITATIONS: The nature and quality of non-physical treatments provided by the psychiatric services was not investigated. Reports of the treatment provided by general practitioners, the timing of improvement in symptoms and compliance with treatment largely relied on patients' self report. CONCLUSIONS: All patients presenting following DSH need to be carefully screened for depressive illness. Randomised controlled studies need to be conducted on DSH patients with depression to determine which treatments are effective. PMID- 12113921 TI - Patterns of sensitisation in the course of affective illness. A life-charting study of treatment-refractory depressed patients. AB - BACKGROUND: A 'sensitisation' process over time has been suggested by Post and collaborators--the affective illness course shows a tendency towards more frequent, deeper, and less stress-related episodes over time. The main aim of the present study was to test the sensitisation hypothesis using a Swedish Life Charting program. METHODS: Thirty patients with treatment-refractory affective disorder, of whom four had bipolar I disorder, were first interviewed using a semistructural interview manual covering episodes, treatment and stress. All previous psychiatric records were then recruited. Information from the records and from the interview was coalesced into individual, retrospective life charts. RESULTS: Twenty patients showed a sensitisation course and 10 patients showed a non-sensitisation course. In both groups, almost 90% of illness episodes had undergone treatment. Time spent in illness since onset of the affective disorder was about 33% for the sensitisation group and more than 50% for the non sensitisation group. LIMITATIONS: The retrospective nature of the study is a limitation. The results apply to patients with severe treatment-refractory affective disorder and may not be generalisable to general patients with less severe mood disorders. CONCLUSIONS: Our results partially validate the Post hypothesis of affective sensitisation in demonstrating this phenomenon in more than half of our affectively recurring patients. However, a substantial minority were clearly non-sensitisers showing a stable but more malignant illness course. Future studies need to elucidate whether these two groups benefit from different kinds of treatment. PMID- 12113922 TI - Seasonal affective disorder in eight groups in Turkey: a cross-national perspective. AB - OBJECTIVE: Previous estimates of the prevalence of seasonal affective disorder (SAD) in community-based samples generally originated from western countries. We report prevalence rates in eight groups from four latitudes in Turkey. METHOD: Seasonal Pattern Assessment Questionnaire (SPAQ) was distributed to the community based samples from eight different locations at four latitudes in Turkey. The prevalence rates of winter SAD and subsyndromal SAD (S-SAD) were estimated for the four groups at the same latitudes by using SPAQ responses. RESULTS: We distributed 3229 SPAQs, had an overall response rate of 54.16% and 1749 SPAQs were included in the analyses. Seasonality was reported as a problem by 549 subjects (31.57%) of our 1749 respondents. Prevalence of winter SAD and S-SAD are estimated as 4.86 and 8.35%, respectively, for the whole group. Prevalence rates were determined for each center and for four latitudes (two centers at the same latitude were grouped as one). In Adana-Gaziantep (lt. 37), Izmir-Elazig (lt. 38), Eskisehir-Ankara (lt. 39) and Trabzon-Edirne (lt. 41), the prevalence rates for winter SAD were 6.66, 2.25, 8.00 and 3.76%, respectively. CONCLUSIONS: Our prevalence estimates of winter SAD are similar to those found in previous community-based studies at the same latitudes; no correlation was found between latitude and prevalence of winter SAD, which could be related to the sampling methodology or to the fact that there were only 5 degrees of difference between the latitudes. PMID- 12113923 TI - Thyroid echogenicity in manic-depressive patients receiving lithium therapy. AB - BACKGROUND: Lithium is known to induce subclinical or overt hypothyroidism and changes of thyroid volume in manic-depressive patients. Little is known about alterations of thyroid echogenicity due to drug-induced dysfunction. METHODS: Twenty manic-depressive patients receiving lithium therapy for at least 6 months and 20 euthyroid volunteers without any antidepressive medication as control group, matched in age and gender, were investigated by laboratory measurements and thyroid ultrasonography including standardized grey scale analyses in representative regions of interest (ROI). RESULTS: Thyroid function was normal in all patients (mean FT4 1.1+/-0.2 ng/dl, mean TSH 1.6+/-0.9 micro U/ml) and controls (mean FT4 1.5+/-0.4 ng/dl, TSH 1.1+/-0.3 micro U/ml). Except for two patients, no thyroid autoantibody levels could been detected. Thyroid volumetry revealed significant higher mean values for the lithium treated patients (16.9+/ 11.9 ml) compared with the controls (11.4+/-4.5 ml, P<0.05) with a considerable number of goiters (six patients vs. one control). Thyroid echogenicities in both groups were similar (patients 23.9+/-3.7 grey scales, Grauwerteinheiten = GWE, controls 24.2+/-1.3 GWE) and did not depend on the size of the organs. CONCLUSION: Lithium treatment contributes to increased thyroid volumes, probably due to inhibition of thyroid function and TSH upregulation, but not to changes of thyroid echo levels in patients with still euthyroid function. Further echogenicity studies on patients with lithium-induced overt hypothyroidism and autoimmune activity will be of special interest. PMID- 12113924 TI - A family study of early-onset bipolar I disorder. AB - BACKGROUND: Relatives of early-onset bipolar probands have greater risk for affective disorders than those of adult-onset bipolar probands. METHODS: Relatives of 50 adolescent bipolar I probands and 36 adult-onset bipolar probands (onset > or = 25 years) were assessed using the Family Interview for Genetic Studies (FIGS) by a qualified psychiatrist blind to the proband status. Morbid risk was calculated using Weinberg's method of age correction. RESULTS: Relatives of early-onset probands had significantly greater risk for affective disorders compared to the relatives of adult-onset bipolar probands. CONCLUSIONS: Early onset bipolar disorder is more familial than the adult bipolar disorder. IMPLICATIONS: Subdivision of bipolar disorder according to age-at-onset may identify homogeneous subtypes useful for genetic studies. LIMITATIONS: Patients were recruited from a major psychiatric hospital. The family history method was used to collect information about relatives. PMID- 12113925 TI - The influence of depressive state features on trait measurement. AB - BACKGROUND: Certain personality characteristics may change during depression, reflecting temporary states, while others remain stable. This study investigates the stability of personality during depression. Identification of depression sensitive personality traits may help to elucidate the mechanisms that influence course and outcome of depression. METHODS: For 42 ambulatory and 38 day care patients with a depressive disorder, depression severity and personality characteristics were measured repeatedly during a 12 week-period. The total HDRS score was indicative of depression severity, while the NEO-FFI was used for measurement of basic personality traits and the IPS for depression-specific traits. RESULTS: In 12 weeks, depression severity significantly decreased for both patient groups. The Big Five traits Neuroticism and Extraversion and all but one aspect of Interpersonal sensitivity showed depression-related changes towards the normal range. Openness, Altruism and Conscientiousness remained stable. LIMITATIONS: The number of patients excluded from the study due to missing data is substantial. Furthermore, it was not possible to control for treatment influence due to a double-blind design. CONCLUSIONS: Neuroticism, Extraversion and Interpersonal sensitivity scores are likely to reflect state components during a depression, while the stability of Openness, Conscientiousness and Altruism scores indicates pure personality traits. PMID- 12113926 TI - Demonstration of spread by Mycobacterium tuberculosis bacilli in A549 epithelial cell monolayers. AB - We developed an in vitro tissue-culture model to analyze the process involved in mycobacterial spread through lung epithelial cell monolayers. A549 cells were infected with low numbers of viable Mycobacterium tuberculosis bacilli expressing the gfp gene. Subsequent addition of a soft agarose overlay prevented the dispersal of the bacilli from the initial points of attachment. By fluorescence microscopy the bacteria were observed to infect and grow within the primary target cells; this was followed by lysis of the infected cells and subsequent infection of adjacent cells. This process repeated itself until an area of clearing (plaque formation) was observed. The addition of amikacin after initial infection did not prevent intracellular growth; however, subsequent plaque formation was not observed. Plaque formation was also observed after infection with Mycobacterium bovis BCG bacilli, but the plaques were smaller than those formed after infection with M. tuberculosis. These observations reinforce the possibility that cell-to-cell spreading of M. tuberculosis bacilli, particularly early in the course of infection within lung macrophages, pneumocytes, and other cells, may be an important component in the infectious process. PMID- 12113927 TI - Identification and analysis of the amylase-binding protein B (AbpB) and gene (abpB) from Streptococcus gordonii. AB - The binding of salivary amylase to Streptococcus gordonii has previously been shown to involve a 20-kDa amylase-binding protein (AbpA). S. gordonii also releases an 82-kDa protein into the supernatant that binds amylase. To study this 82-kDa component, proteins were precipitated from bacterial culture supernatants by the addition of acetone or purified amylase. Precipitated proteins were separated by SDS-PAGE and transferred to a sequencing membrane. The P2 kDa band was then sequenced, yielding a 25 N-terminal amino acid sequence, CGFIFGRQLTADGSTMFGPTEDYP. Primers derived from this sequence were used in an inverse PCR strategy to clone the full-length gene from S. gordonii chromosomal DNA. An open reading frame of 1959 bp was noted that encoded a 652 amino acid protein having a predicted molecular mass of 80 kDa. The first 24 amino acid residues were consistent with a hydrophobic signal peptide, followed by a 25 amino acid N-terminal sequence that shared identity (24 of 25 residues) with the amino acid sequence of purified AbpB. The abpB gene from strains of S. gordonii was interrupted by allelic exchange with a 420-bp fragment of the abpB gene linked to an erythromycin cassette. The 82-kDa protein was not detected in supernatants from these mutants. These abpB mutants retained the ability to bind soluble amylase. Thus, AbpA, but not AbpB, appears sufficient to be the major receptor for amylase binding to the streptococcal surface. The role of AbpB in bacterial colonization remains to be elucidated. PMID- 12113928 TI - The malate dehydrogenase of Ralstonia eutropha and functionality of the C(3)/C(4) metabolism in a Tn5-induced mdh mutant. AB - The Tn5-induced mutant VG12 of Ralstonia eutropha HF39, which was isolated in this study, revealed an interesting phenotype: it grew on fructose and pyruvate as well as autotrophically like the wild-type, whereas growth on tricarboxylic acid intermediates and glyoxylic acid was reduced, and no growth occurred if acetate, propionate or levulinate were provided as carbon source. Tn5 was mapped in a gene encoding an NAD(H)-dependent malate dehydrogenase (MDH), and MDH activity was strongly diminished in VG12. Furthermore, the mdh gene was cloned, sequenced and heterologously expressed in Escherichia coli, conferring significantly higher specific MDH activity to the recombinant strain. The phenotype of VG12 sheds light on the C(3)/C(4) metabolism of R. eutropha, which mediates between the Entner-Doudoroff pathway and the tricarboxylic acid cycle (TCC), demonstrating that enzymes catalyzing the conversion of C(3) and C(4) metabolites can circumvent the metabolic disruption of the TCC in VG12 and that the phosphoenolpyruvate carboxykinase serves a dual and important function. PMID- 12113929 TI - Phylogenetic analysis of the genus Aquaspirillum based on 16S rRNA gene sequences. AB - Phylogenetic analysis of 15 species of the genus Aquaspirillum based on 16S rRNA gene (rDNA) sequences indicated that the genus Aquaspirillum is phylogenetically heterogeneous and the species could be divided into four groups as follows: Aquaspirillum serpens, the type species of this genus, A. dispar and A. putridiconchylium are situated in the family Neisseriaceae; members of the second group, A. gracile, A. delicatum, A. anulus, A. giesbergeri, A. sinuosum, A. metamorphum and A. psychrophilum, are included in the family Comamonadaceae; the two members of the third group, A. arcticum and A. autotrophicum, are included in the family Oxalobacteriaceae; and members of the fourth group, A. polymorphum, A. peregrinum, and A. itersonii, are included in the alpha-subdivision of Proteobacteria. Thus, phylogenetic studies indicated that all the species excepting A. serpens, the type species, should be transferred to distinct genera. PMID- 12113930 TI - A molecular marker diagnostic of a specific isolate of an arbuscular mycorrhizal fungus, Gigaspora margarita. AB - To investigate the auto-ecology of a strain of Gigaspora margarita in a commercial inoculum, we found a pair of PCR primers amplifying a sequence of 235 bp diagnostic of the isolate. We designed an oligonucleotide probe based on the DNA sequence. The combination of PCR and the probing successfully detected the diagnostic sequence from both DNA preparations of single spores and colonized roots. This protocol enabled us to distinguish the isolate among several isolates from Japan, Nepal and the USA. PMID- 12113931 TI - Control of autogenous activation of Herbaspirillum seropedicae nifA promoter by the IHF protein. AB - Analysis of the expression of the Herbaspirillum seropedicae nifA promoter in Escherichia coli and Herbaspirillum seropedicae, showed that nifA expression is primarily dependent on NtrC but also required NifA for maximal expression under nitrogen-fixing conditions. Deletion of the IHF (integration host factor)-binding site produced a promoter with two-fold higher activity than the native promoter in the H. seropedicae wild-type strain but not in a nifA strain, indicating that IHF controls NifA auto-activation. IHF is apparently required to prevent overexpression of the NifA protein via auto-activation under nitrogen-fixing conditions in H. seropedicae. PMID- 12113932 TI - Asp 187 and Phe 190 residues in lethal factor are required for the expression of anthrax lethal toxin activity. AB - Anthrax toxin consists of three proteins, protective antigen, lethal factor, and edema factor. Protective antigen translocates lethal factor and edema factor to the cytosol of mammalian cells. The amino-termini of lethal factor and edema factor have several homologous stretches. These regions are presumably involved in binding to protective antigen. In the present study we have determined the role of one such homologous stretch in lethal factor. Residues 187AspLeuLeuPhe190 were replaced by alanine. Asp187Ala and Phe190Ala were found to be non-toxic in combination with protective antigen. Their protective antigen-binding ability was drastically reduced. We propose that Asp187 and Phe190 are crucial for the expression of anthrax lethal toxin activity. PMID- 12113933 TI - Pheno- and genotypic properties of streptococci of serological group B of canine and feline origin. AB - In the present study streptococci of serological group B isolated from canines (n=48) and felines (n=7) were comparatively investigated with group B streptococci from humans and bovines for cultural, biochemical and serological properties for antibiotic resistancies and by molecular analysis. An identification was performed with group B-specific antiserum, biochemical reactions, by PCR amplification and subsequent endonuclease digestion of the 16S rRNA gene and by amplification of species-specific parts of the 16S rDNA the 16S 23S rDNA intergenic spacer region and the CAMP factor gene cfb. Phenotypic similarities of group B streptococci of canine and feline origin with group B streptococci from humans and differences to group B streptococci of bovine origin could be observed in lactose fermentation, serotype patterns, pigmentation, growth properties of the bacteria in fluid medium and soft agar, hemagglutination reactions and in minocycline and tetracycline resistance. A negative hyaluronidase plate test, a hylB amplicon with a size of 4.6 kb and an insertion sequence 1548 could be observed among canine, feline and human group B streptococci of serotype III. The remaining hyaluronidase positive strains, also including all isolates of bovine origin, had a hylB gene with a size of 3.3 kb. Further genotypic differences could be observed in the occurrence of the genes lmb and scpB which appeared generally among canine, feline and human group B streptococci, but less pronounced among bovine isolates of this species. According to the presented data group B streptococci of canine and feline origin seemed to be more related to human than to bovine isolates of this species possibly indicating some epidemiological relation. PMID- 12113934 TI - Identification and characterization of a novel Chlamydia trachomatis reticulate body protein. AB - The genome of the obligate intracellular bacterium Chlamydia trachomatis comprises 894 genes predicted by computer-based analysis. As part of a large scale proteome analysis of C. trachomatis, a small abundant protein encoded by a previously unrecognized novel 204-bp open reading frame was identified by tandem mass spectrometry. No homology of this protein was observed to proteins from other organisms. The protein was conserved in C. trachomatis but not found in Chlamydia pneumoniae. Using proteomics, we show that the expression of the protein is initiated at the middle of the developmental cycle. The protein is rapidly degraded and is only present in reticulate or intermediate bodies, suggesting a possible function in the intracellular stage of C. trachomatis development. We have termed the protein '7-kDa reticulate body protein'. PMID- 12113935 TI - A distinct physiological state of Lactococcus lactis cells that confers survival against a direct and prolonged exposure to severe stresses. AB - When exponential phase cultures of Lactococcus lactis were directly exposed to severe stresses (acid, bile salt, heat, and hydrogen peroxide) for a prolonged period, most of the cells were quickly killed, however, a small number of the cells, approximately 0.01% of the population, was found to survive. How these 'survivor' cells might have survived the stresses, when other supposedly-the-same cells could not, was investigated. The cultures were not exposed to any mild stresses prior to the exposure to the severe stresses, and therefore adaptation can be ruled out as the cause of survival. When the survivor cells were re cultured and re-exposed to the same severe stresses a similar pattern of survival was displayed, indicating that the survivor cells were not stress-resistant mutants. Furthermore, the survivor cells displayed typical growth kinetics once they were freed of the stresses. The survivor cells appear to be in a distinct physiological state, because when they were tested against a second stress they exhibited significantly greater survival against that stress than the normal cells exposed to the same stress. Also, cells at different time points of synchronously growing culture displayed different levels of survival against stress. It is proposed that the difference in survival of exponential phase cells is due to the difference in the protein makeup of cells at different stages of the cell cycle. PMID- 12113936 TI - Construction of fusion vectors of corynebacteria: expression of glutathione-S transferase fusion protein in Corynebacterium acetoacidophilum ATCC 21476. AB - A series of fusion vectors containing glutathione-S-transferase (GST) were constructed by inserting GST fusion cassette of Escherichia coli vectors pGEX4T 1, -2 and -3 in corynebacterial vector pBK2. Efficient expression of GST driven by inducible tac promoter of E. coli was observed in Corynebacterium acetoacidophilum. Fusion of enhanced green fluorescent protein (EGFP) and streptokinase genes in this vector resulted in the synthesis of both the fusion proteins. The ability of this recombinant organism to produce several-fold more of the product in the extracellular medium than in the intracellular space would make this system quite attractive as far as the downstream processing of the product is concerned. PMID- 12113937 TI - Chemical stress-responsive genes from the lignin-degrading fungus Phanerochaete chrysosporium exposed to dibenzo-p-dioxin. AB - The stress-responsive genes expressed against the exogenous addition of dibenzo-p dioxin from the lignin-degrading basidiomycete, Phanerochaete chrysosporium, were determined utilizing a differential display reverse transcription-PCR technique. Six cDNA fragments, exhibiting a high homology with various proteins from other microorganisms, were identified via a BLAST search; that is, NADH-ubiquinone oxidoreductase (NUO), ATP/ADP carrier, uric acid-xanthine permease, manganese superoxide dismutase, 3-hydroxybutyryl-coenzyme A dehydrogenase, and cytoskeletal protein. The expression of NUO was also up-regulated by catechol and trihydroxybenzene but not by dibenzofuran, suggesting that NUO expression was initiated by the formation of quinone products through the reaction of extracellular one-electron oxidizing enzymes with dibenzo-p-dioxin. PMID- 12113938 TI - Identification of a type III secretion system in uropathogenic Escherichia coli. AB - To determine virulence-related genes in uropathogenic Escherichia coli (UPEC) showing invasiveness to T-24 bladder cancer cells, genomic subtractive hybridization was performed between a highly invasive and a less invasive strain. Forty-nine DNA fragments were isolated from the invasive strain. One of them showed homology with Salmonella invA gene. By chromosomal walking of the strain, a type III secretion system that has been described in E. coli O157:H7 was identified on the genome of the invasive strains. Three strains out of 100 UPEC isolates had a type III secretion system inserted at 64 min of the chromosome, corresponding to E. coli K-12 MG1655. This finding suggested that the type III secretion system could play a part in uropathogenicity of UPEC. PMID- 12113939 TI - Identification and characterization of the Escherichia coli envC gene encoding a periplasmic coiled-coil protein with putative peptidase activity. AB - PM61 is a chain-forming envC strain of Escherichia coli with a leaky outer membrane. It was found to have an oversized penicillin-binding protein 3, which was the result of an IS4 insertion in the prc gene. The other properties of PM61 were caused by the envC mutation. We cloned the envC (yibP) gene and identified the mutation site, causing a single residue substitution, H366Y, in the PM61 envC allele. The gene product was predicted to be a periplasmic protein having coiled coil structure in the N-terminal region and homology to lysostaphin in the C terminal region. Overexpression of envC inhibited cell growth, and overexpression of the PM61 mutant allele caused cell lysis. Disruption of the chromosomal envC caused the same defects as the envC point mutation, indicating the gene is dispensable for growth but important for normal septation/separation and cell envelope integrity. PMID- 12113940 TI - Secretion of endoxylanase A from Penicillium purpurogenum by Saccharomyces cerevisiae transformed with genomic fungal DNA. AB - Saccharomyces cerevisiae was transformed with a genomic library from Penicillium purpurogenum, and an endoxylanase-producing yeast clone (named 44A) that grows on xylose or xylan as sole carbon source was isolated. This yeast synthesizes xynA mRNA and secretes endoxylanase A to culture media when grown on xylan or xylose, but not glucose. Analysis by pulse-field gel electrophoresis and sequencing indicates that xynA, including its eight introns, has been inserted into the yeast genome. It was shown by sequencing that clone 44A is able to correctly splice xynA introns. This is the first successful attempt to express a fungal endoxylanase gene in yeast with correct intron splicing. PMID- 12113941 TI - De novo synthesis of amino acids by the ruminal anaerobic fungi, Piromyces communis and Neocallimastix frontalis. AB - Anaerobic fungi are an important component of the cellulolytic ruminal microflora. Ammonia alone as N source supports growth, but amino acid mixtures are stimulatory. In order to evaluate the extent of de novo synthesis of individual amino acids in Piromyces communis and Neocallimastix frontalis, isotope enrichment in amino acids was determined during growth on (15)NH(4)Cl in different media. Most cell N (0.78 and 0.63 for P. communis and N. frontalis, respectively) and amino acid N (0.73 and 0.59) continued to be formed de novo from ammonia when 1 g l(-1) trypticase was added to the medium; this concentration approximates the peak concentration of peptides in the rumen after feeding. Higher peptide/amino acid concentrations decreased de novo synthesis. Lysine was exceptional, in that its synthesis decreased much more than other amino acids when Trypticase or amino acids were added to the medium, suggesting that lysine synthesis might limit fungal growth in the rumen. PMID- 12113942 TI - Efflux of chromate by Pseudomonas aeruginosa cells expressing the ChrA protein. AB - The ChrA protein of Pseudomonas aeruginosa plasmid pUM505 confers resistance to chromate. Using an in vitro system, we reported [Alvarez, A.H. et al. (1999) J. Bacteriol. 181, 7398-7400] that chromate resistance is based on energy-dependent efflux of chromate. It is shown here that ChrA determines in vivo efflux of 51CrO(4)(2-) as well. Chromate-loaded cell suspensions of P. aeruginosa strain PAO1 harboring recombinant plasmid pEPL1, which expresses the ChrA protein, showed accelerated efflux of 51CrO(4)(2-) as compared to the plasmidless chromate sensitive derivative. After a 10-min loading, about 40% of 51CrO(4)(2-) was lost from resistant cells in 15 min. Chromate efflux by resistant cells showed a typical saturation kinetics with an apparent K(m) of 82+/-11 microM chromate and a V(max) of 0.133+/-0.009 nmol chromate min(-1) (mg protein)(-1). Oxyanions sulfate and molybdate inhibited chromate efflux in a concentration-dependent fashion, whereas arsenate and ortho-vanadate had no significant effect on chromate release. Inhibition of chromate extrusion by valinomycin, nigericin, and carbonyl cyanide m-chlorophenylhydrazone, but not by oligomycin or dicyclohexylcarbodiimide, indicated that chromate efflux was driven by the membrane potential. PMID- 12113943 TI - Pythium terrestris, a new species isolated from France, its ITS region, taxonomy and its comparison with related species. AB - Pythium terrestris (F-78) was isolated from soil samples taken in Lille in northern France. Its morphology resembles that of Pythium rostratum, and Pythium longandrum, a recently described species. However the antheridial and sporangial characteristics of this new species are unique. The fungus does not sporulate, the sporangium germinates directly into mycelium through germ tubes. The oogonia of P. terrestris are normally provided with hypogynous and monoclinous antheridia, at times the monoclinous antheridial branches wrap around the oogonia forming a complicated knot. Morphological features of this new species, together with the sequences of the internal transcribed spacer (ITS) region of its nuclear ribosomal DNA and its comparison with related species are discussed here. PMID- 12113944 TI - A partially 3-O-methylated (1-->4)-linked alpha-D-galactan and alpha-D-mannan from Pleurotus ostreatoroseus Sing. AB - The two main water-soluble extracellular polysaccharides produced by the basidiomycete fungus Pleurotus ostreatoroseus Sing were isolated and purified. They were characterized using 13C, 1H, and 1H, 13C HMQC NMR spectroscopy, methylation analysis, and Smith degradation. One was a mannan having a main chain of (1-->6)-linked alpha-D-mannopyranosyl residues, almost all of which were branched at O-2 with side chains of different lengths, containing 2-O- and 3-O linked mannopyranosyl units. The other was a partially 3-O-methylated (1-->4) linked alpha-D-galactopyranan, a structure that has not been previously described. PMID- 12113945 TI - Increases in amino-cupric-silver staining of the supraoptic nucleus after sleep deprivation. AB - Sleep deprived rats undergo a predictable sequence of physiological changes, including changes in skin condition, increased energy expenditure, and altered thermoregulation. Amino-cupric-silver staining was used to identify sleep deprivation related changes in the brain. A significant increase in staining was observed in the supraoptic nucleus (SON) of the hypothalamus of rats with high sleep loss (>45 h) vs. their yoked controls. Follow-up experiments showed that staining was not significantly different in rats sleep deprived for less than 45 h, suggesting that injurious sleep deprivation-related processes occur above a threshold quantity of sleep loss. These anatomical changes suggest that the effects of sleep deprivation may be related to protein metabolism in certain brain regions. PMID- 12113947 TI - Characteristics of electroacupuncture-induced analgesia in mice: variation with strain, frequency, intensity and opioid involvement. AB - The present study was conducted to evaluate the characteristics of electroacupuncture (EA)-induced analgesia in mice. Three inbred strains of mice (DBA/2, C57BL/6J, BALB/c) and three outbred strains (ICR, LACA, NIH) were used in the experiment. Two pairs of metallic needles were inserted into acupoints ST 36 and SP 6 connected to an electric pulse generator. EA parameters were set as constant current output with alteration of a positive and negative square wave, 0.6 ms in pulse width for 2 Hz and 0.3 ms for 100 Hz. Tail-flick latencies evoked by radiant heat were measured before, during and after EA stimulation. We found that (1) DBA/2 mice showed a significantly more potent analgesic effect than the other five strains in response to both 100 and 2 Hz EA. In this case, the intensities were 1.0-2.0-2.0 mA, 10 min for each intensity totally 30 min. (2) EA analgesia increased as the intensity of stimulation increased from 0.5 to 2.0 mA, but it remained at this plateau when the intensity further increased from 2.0 to 3.0 mA. (3) 10.0 mg x kg(-1) naloxone was needed to block the analgesic effect induced by 2 Hz EA of 2.0 mA, but to block that by 100 Hz, 25.0 mg x kg(-1) was necessary. (4) A positive correlation was observed between analgesia induced by morphine at the dose of 5.0 mg x kg(-1) and by 100 Hz EA in two tested strains DBA/2 and C57BL/6J. In conclusion, EA induces reliable, strain-dependent analgesia in mice. The naloxone-reversibility of EA, a measure of whether it is opioid or non-opioid mediated, is dependent upon intensity and frequency. PMID- 12113946 TI - Quantitative autoradiographic mapping of opioid receptors in the brain of delta opioid receptor gene knockout mice. AB - Using quantitative receptor autoradiography we have determined if deletion of the delta-opioid receptor gene (Oprd1) results in compensatory changes in the expression of other opioid receptors. Gene targeting was used to delete exon 1 of the mouse delta-opioid receptor gene and autoradiography was carried out on brains from wild-type, heterozygous and homozygous knockout mice. Delta-opioid receptors were labeled with [(3)H]deltorphin I (7 nM), mu- with [(3)H]DAMGO (4 nM), and kappa- with [(3)H]CI-977 (2.5 nM) or [(3)H]bremazocine (2 nM in the presence of DPDPE and DAMGO) and non-specific binding determined with naloxone. [(3)H]Deltorphin I binding was reduced by approximately 50% in heterozygous animals. In homozygous animals specific binding could only be detected after long term film exposure (12 weeks). Regions exhibiting this residual [(3)H]deltorphin I binding correlated significantly with those demonstrating high levels of the mu receptor and were abolished in the presence of the mu-agonist DAMGO. Autoradiographic mapping showed significant overall reductions in [(3)H]DAMGO and [(3)H]CI-977 binding throughout the brain following loss of both copies of the Oprd1 gene. In contrast, overall levels of [(3)H]bremazocine binding were higher in brains from -/- than +/+ mice. Our findings suggest that residual [(3)H]deltorphin I binding in the brain of delta-receptor gene knockout mice is the result of cross-reactivity with mu-sites and that there are no delta-receptor subtypes derived from a different gene. Changes in mu- and kappa-receptor labeling suggest compensatory changes in these subtypes in response to the absence of the delta-receptor. The differences in [(3)H]CI-977 and [(3)H]bremazocine binding indicate these ligands show differential recognition of the kappa-receptor. PMID- 12113948 TI - Reactivity to object and spatial novelty is normal in older Ts65Dn mice that model Down syndrome and Alzheimer's disease. AB - Ts65Dn mice, a model for Down syndrome and Alzheimer's disease, have a spontaneous age-related reduction of cholinergic markers in medial septal neurons, hippocampal abnormalities, and an age-related learning deficit in a task that requires an intact hippocampus. Others have shown that when normal rodents explored an open field with objects, they detected the displacement of some of the familiar objects within the arena (spatial novelty) and the presence of a new object (object novelty); whereas rodents with hippocampal, fornix, or neonatal selective basal forebrain cholinergic lesions were impaired in detecting spatial, but not object, novelty. In this study, both control and Ts65Dn mice responded to both the spatial and object changes. This unexpected finding could have several explanations. One may be related to recent studies that suggest that only rats with neonatal, but not adult, basal forebrain cholinergic 192 IgG-saporin lesions are impaired in reacting to spatial novelty. PMID- 12113949 TI - Excitatory effects of dopamine on subthalamic nucleus neurons: in vitro study of rats pretreated with 6-hydroxydopamine and levodopa. AB - Increased output from the subthalamic nucleus (STN) following chronic dopamine depletion has been linked to the rigidity and tremor seen in Parkinson's disease (PD). We used extracellular microelectrode recordings from rat brain slices to investigate effects of dopamine on STN neurons. In brain slices prepared from rats that received unilateral 6-hydroxydopamine (6-OHDA) treatment, the spontaneous firing rate of STN neurons was reduced by 63%, and the firing pattern was more irregular, compared to STN neurons from normal rats. However, treatment with levodopa (50 mg/kg, i.p., daily) for 4 weeks normalized the firing rate and pattern of STN neurons in the 6-OHDA-treated rats. Dopamine (3-300 microM), added to the superfusate, significantly increased the firing rates of STN neurons in a concentration-dependent fashion, and also produced a more regular firing pattern in 6-OHDA-lesioned tissue. This excitatory effect of dopamine was mimicked by a D2 receptor agonist (quinpirole), and was reduced by the D2 antagonists haloperidol, clozapine and sulpiride. Antagonists of the D1 receptor (SCH-23390) and ionotropic glutamatergic receptors (CNQX and AP5) could not block the effect of dopamine on firing rate. These results suggest that dopamine exerts a direct excitatory influence on STN neurons via the activation of D2-like receptors. PMID- 12113950 TI - Oxyhemoglobin produces necrosis, not apoptosis, in astrocytes. AB - OBJECTIVE: Subarachnoid blood, resulting from traumatic brain injury or subarachnoid hemorrhage, has been linked with cell injury and stress gene induction. We investigated whether oxyhemoglobin (OxyHb), a major component in blood clots, exerts a cytotoxic effect on cultured astrocyte cells, and the pattern of cell death. METHODS: A murine astrocyte cell line was used (passages 28-35). Cell growth studies were performed 24, 48, and 72 h after exposure to OxyHb (1, 10, and 30 microM). Western blot analysis of poly adenosine diphosphate [ADP]-ribose polymerase (PARP) cleavage and TUNEL stain analysis were performed to determine the presence of apoptosis. Cells treated with OxyHb were also evaluated with transmission electron microscopy to determine changes that may have occurred at the ultra-structural level. RESULTS: OxyHb (10-30 microM), after 72-h incubation, inhibited cell growth. Western blot analysis of PARP and TUNEL staining for the presence of apoptosis were essentially negative in all groups. Ultrastructural analysis revealed an abundance of necrosis and random occurrences of apoptosis in a few cells. CONCLUSION: Cultured astrocytes exposed to OxyHb causing cell growth inhibition could possibly be a result of cellular cytotoxicity and necrosis. PMID- 12113951 TI - Intracerebroventricular administration of alpha-melanocyte stimulating hormone increases phosphorylation of CREB in TRH- and CRH-producing neurons of the hypothalamic paraventricular nucleus. AB - Changes in circulating leptin levels, as determined by nutritional status, are important for the central regulation of neuroendocrine axes. Among these effects, fasting reduces TRH gene expression selectively in the hypothalamic paraventricular nucleus (PVN), which can be reversed by leptin administration. Intracerebroventricular (i.c.v.) infusion of alpha-MSH recapitulates the effects of leptin on hypophysiotropic TRH neurons, completely restoring proTRH mRNA to levels in fed animals despite continuation of the fast, making alpha-MSH a candidate for mediating the central effects of leptin. As alpha-MSH binds to a G protein coupled receptor that activates cAMP and alpha-MSH stimulates the TRH promoter through the phosphorylation of the transcription factor CREB in vitro, we determined whether i.c.v. injection of alpha-MSH to rats regulates phosphorylation of CREB, specifically in hypophysiotropic TRH neurons of PVN. As alpha-MSH also induces the activation of CRH gene expression in the PVN, we further determined whether i.c.v. injection of alpha-MSH regulates the phosphorylation of CREB in hypophysiotropic CRH neurons. In vehicle-treated animals, only rare neurons contained nuclear phospho-CREB (PCREB) immunoreactivity in the parvocellular PVN. I.c.v. injection of 10 microg alpha MSH dramatically increased the number of PCREB-immunolabeled cell nuclei in the PVN in fasted groups at 10 min postinjection, particularly in the medial, periventricular, anterior and ventral parvocellular subdivisions, whereas a moderate increase of PCREB immunoreactivity was observed at 30 min and PCREB immunoreactivity was lowest at 1 h postinfusion. Double immunolabeling with proTRH antiserum revealed that following i.c.v. alpha-MSH infusion at 10 min, the majority of TRH neurons contained PCREB in the anterior (71%), medial (83%) and periventricular (63%) parvocellular subdivisions. The percentage of double labeled TRH neurons declined at 30 min and 1 h post alpha-MSH infusion. Similarly, only 16% of CRH neurons of the medial parvocellular neurons contained PCREB nuclei in vehicle treated animals, whereas 10 min following alpha-MSH infusion the percentage of CRH neurons colocalizing with the PCREB rose to 54%, then fell to 37 and 17% at 30 and 60 min postinfusion, respectively. These data demonstrate that i.c.v. alpha-MSH administration increases the phosphorylation of CREB in several subdivisions of the PVN including TRH and CRH neurons in the medial and periventricular parvocellular subdivisions, suggesting that phosphorylation of CREB may be necessary for alpha-MSH-induced activation of the TRH and CRH genes. The increase in PCREB in the anterior and ventral parvocellular subdivisions of the PVN, regions linked to nonhypophysiotropic functions such as autonomic regulation, would also imply a role for these neurons in anorectic and energy wasting responses of melanocortin signaling. PMID- 12113952 TI - Modulation of serotonergic projection from dorsal raphe nucleus to basolateral amygdala on sleep-waking cycle of rats. AB - Putative serotonergic dorsal raphe nucleus (DRN) neurons display a dramatic role in the modulation of behavior. However, it is not clear how this modulation is mediated. The present study investigated the modulatory effects of serotonergic projection of the DRN to the basolateral amygdala (BLA) on the sleep-waking cycle using polysomnograph (PSG) in rats. DRN microinjection of kainic acid (KA) caused insomnia immediately. From the third day, however, slow wave sleep (SWS) and paradoxical sleep (PS) increased markedly. DRN microinjection of p chlorophenylalanine (PCPA, once a day for 2 days), which inhibits the synthesis of serotonin (5-HT), led to similar effect to KA administration. The percent of sleep-wakefulness began to change on the third day after PCPA microinjection into the DRN, and the effect was most significant on the sixth day. The percent of sleep-wakefulness started to resume on the seventh day. SWS and PS were reduced after excitation of DRN neurons by microinjection of L-glutamate (L-Glu) into the DRN. Preapplication of the nonselective 5-HT receptor antagonist methysergide (MS) into bilateral BLA blocked the effect of DRN microinjection of L-Glu. Furthermore, bilateral BLA microinjection of 5-hydroxytryptophan (5-HTP), the precursor of 5-HT, on the sixth day after microinjection of PCPA into the DRN, could reverse the effect of PCPA microinjection. These results indicate that the modulation of the DRN on sleep is partially mediated by the serotonergic projection of the DRN to the BLA. PMID- 12113954 TI - Protection by rapid chemical preconditioning of stressed hippocampal slice: a study of cellular swelling using optical scatter imaging. AB - It has been demonstrated that anoxic preconditioning protects against a subsequent 'lethal' injury in the hippocampal slice. The goal of this paper was to test the hypothesis that chemical preconditioning could help reduce the cellular swelling observed in excitotoxically injured hippocampal slices. The control slice was given a 10-min insult of 100 microM N-methyl-D-aspartate (NMDA) to simulate ischemic injury, followed by 30-min perfusion of standard Ringers solution. Cellular swelling was observed with a microscope designed to image light scatter changes resulting from cellular swelling. After the control NMDA injury, the average peak scatter change for CA1, CA3 and DG regions was 31.0 +/- 3.4, 22.4 +/- 4.8 and 27.6 +/- 4.6%, respectively. The peak scatter change of the overall slice was 26.0 +/- 3.6%. The experimental slices were preconditioned by three short 100 microM NMDA insults of 15 s each separated by 10 min of standard Ringers solution perfusion. The slices then received 10 min of 'lethal' injury by 100 microM NMDA. It was observed that the overall scatter signal, as a measure of cellular swelling, was reduced by 8.0% (P<0.05, n=11) after preconditioning. A regional heterogeneity in the responses was also observed. Cellular swelling in CA1, CA3 and DG were reduced by 9.8% (P<0.001, n=11), 9.2% (P<0.005, n=11) and 7.7% (P<0.05, n=11), respectively, when compared to the control. This study presents experimental evidence that short episodes of preconditioning may protect against acute cellular swelling under ischemic conditions. PMID- 12113953 TI - Noise-induced compensation for postural hypotension in primary autonomic failure. AB - Noise can have a beneficial effect on sensory neurological systems, enhancing detection of small afferent signals and thereby improve efferent neural responses. We hypothesized whether a similar mechanism would facilitate impaired neural transmission associated with neurological disease, and tested whether addition of external noise to baroreceptor signaling could improve blunted autonomic efferent responses to a postural challenge in patients with primary autonomic failure (PAF). Five PAF patients were tested, one in duplicate and another triplicate, for their transient responses of heart rate (measured from electrocardiographic RR intervals; RRIs) and systolic (SBP) and diastolic (DBP) blood pressures to either 30 degrees or 60 degrees head-up tilt, with and without continuous application of beat-to-beat Gaussian white noise to the carotid sinus baroreceptors. Also, the effects of noise were compared with those by a continuous positive pressure applied to the carotid sinus baroreceptors. The data were fit to a first order model to evaluate the speed (by the time constant; tau) and the magnitudes (by the steady state gains; Gs) of RRI and blood pressure responses. The PAF patients exhibited marked drops in SBP and DBP and a blunted increase in heart rate upon transition from a supine to a head-up position. Addition of noise, not the continuous positive pressure, to the arterial baroreceptors significantly (P<0.05) increased the G in RRI and diminished the Gs in SBP and DBP, though the time courses (taus) of both the RRI and blood pressure responses were unaffected. The addition of external noise to baroreceptor signaling ameliorated the marked postural hypotension seen in patients with PAF. PMID- 12113956 TI - Noninvasive measurement of cerebral bioimpedance for detection of cerebral edema in the neonatal piglet. AB - The association of sustained cerebral edema with poor neurological outcome following hypoxia-ischaemia in the neonate suggests that measurement of cerebral edema may allow early prediction of outcome in these infants. Direct measurements of cerebral impedance have been widely used in animal studies to monitor cerebral edema, but such invasive measurements are not possible in the human neonate. This study investigated the ability of noninvasive cerebral impedance measurements to detect cerebral edema following hypoxia-ischaemia. One-day-old piglets were anaesthetized, intubated and ventilated. Hypoxia was induced by reducing the inspired oxygen concentration to 4-6% O(2). Noninvasive cerebral bioimpedance was measured using gel electrodes attached to the scalp. Cerebral bioimpedance was also measured directly by insertion of two silver-silver chloride electrodes subdurally. Noninvasive and invasive measurements were made before, during and after hypoxia. Whole body impedance was measured to assess overall fluid movements. Intracranial pressure was measured continuously via a catheter inserted subdurally, as an index of cerebral edema. There was good agreement between noninvasive and invasive measurements of cerebral impedance although externally obtained responses were attenuated. Noninvasive measurements were also well correlated with intracranial pressure. Whole body impedance changes did not account for increases in noninvasively measured cerebral impedance. Results suggest that noninvasive cerebral impedance measurements do reflect intracranial events, and are able to detect cerebral edema following hypoxia-ischaemia in the neonate. PMID- 12113955 TI - Involvement of endogenous opioid systems in nociceptin-induced spinal antinociception in rats. AB - The present study investigates the involvement of opioid receptors in the antinociceptive effects of nociceptin in the spinal cord of the rat. Intrathecal administrations of 5 and 10 nmol of nociceptin significantly increase the withdraw response latencies to noxious thermal and mechanical stimulations. This nociceptin-induced antinociceptive effect is significantly attenuated by intrathecal injection of (Nphe(1))nociceptin(1-13)-NH(2), a selective antagonist of the nociceptin receptor (opioid receptor-like receptor ORL1), indicating an ORL1 receptor-mediated mechanism. This antinociceptive effect is also significantly attenuated by intrathecal injections of naloxone (a nonselective opioid receptor antagonist), naltrindole (a selective delta-opioid receptor antagonist), and beta-funaltrexamine (a selective mu-opioid receptor antagonist) in a dose-dependent manner, but not by the selective kappa-opioid receptor antagonist norbinaltorphimine. Since it is unlikely that nociceptin acts by direct binding to opioid receptors, these results suggest a possible interaction between the nociceptin/ORL1 and opioid systems in the dorsal horn of the rat spinal cord. PMID- 12113957 TI - Post-insult exposure to (+/-) kavain potentiates N-methyl-D-aspartate toxicity in the developing hippocampus. AB - Kavapyrone extracts of the pepper plant Piper methysticum Forst. have been reported to be pharmacologically active in the brain by modulating the function of several ionotropic receptor systems and voltage-sensitive ion channels. While kavapyrones have previously demonstrated neuroprotective effects against several forms of neurotoxicity, the possibility remains that perturbed function of neuronal ion transport may prove to be neurotoxic in some instances. The present studies were designed to examine the effects of the kavapyrone, (+/-) kavain, on viability of organotypic hippocampal explants exposed to the excitotoxin N-methyl D-aspartate (NMDA). Exposure to (+/-) kavain (1-600 microM) for 24 h did not alter neuronal viability in the CA1, CA3, or dentate gyrus regions of hippocampal explants. However, higher concentrations of (+/-) kavain (> or =300 microM) produced marked neurotoxicity in the lacunosum moleculare layer of the hippocampus. One hour of exposure to NMDA (20 microM) produced significant neuronal death in both the CA3 and CA1 pyramidal cell regions, effects prevented by co-exposure to MK-801 (30 microM). Co-exposure of explants to (+/-) kavain (1 100 microM) with NMDA did not alter the severity of NMDA-induced neurotoxicity. However, exposure of NMDA-treated explants to (+/-) kavain (> or =10 microM) for 24 h after insult produced significant increases in neurotoxicity in the CA1 and dentate gyrus regions of explants. In conclusion, while the kavapyrone (+/-) kavain is neurotoxic only at high concentrations when exposed alone to the developing hippocampus, it appears to adversely affect neuronal recovery following excitotoxic insults. PMID- 12113958 TI - Nitric oxide and nitric oxide synthases in the fetal cerebral cortex of rats following transient uteroplacental ischemia. AB - The effect of transient uteroplacental ischemia on nitric oxide (NO) levels, enzymatic activity, and expression of NO synthase (NOS) isoforms was studied in fetal rat brains. Fetuses were subjected to ischemia by clamping the uterine arteries for 5 min on gestational day 17 (GD17). At different times after ischemia, fetuses were delivered by Cesarean section under anesthesia to obtain the brains. Transient uteroplacental ischemia produced a time dependent increase in nitrite levels in the brain, reaching a maximum value (300 +/- 25% of baseline) 24 h after uterine artery occlusion and remaining elevated as long as 48 h. Significantly increased nitrite levels were found in the cerebral cortex but not in the mesencephalon and cerebellum. The ischemia-induced increment in nitrite levels was totally blocked by either L-NAME (10 mg/kg) or AMT (0.65 mg/kg) administered i.p. 1 h before uterine artery occlusion. Both Ca(2+) dependent and Ca(2+)-independent NOS activities in the cerebral cortex remained significantly increased with respect to controls after 24 h following the ischemia. Reverse transcriptase-polymerase chain reaction showed augmented levels of mRNAs for both nNOS and iNOS when compared with controls at 8 h after ischemia. At 36 h, nNOS mRNA returned to basal levels whereas eNOS mRNA levels increased and iNOS mRNA remained elevated. Our results show that the three NOS isoforms participate in increasing NO levels after transient ischemia and suggest a biphasic and differential regulation of the expression of constitutive NOS isoforms in the rat cerebral cortex. PMID- 12113959 TI - Endogenous gonadal hormones modulate behavioral and neurochemical responses to acute and chronic cocaine administration. AB - Recent evidence demonstrates that there are sex differences in behavioral responses to cocaine. Further, reproductive gonadal hormones (estrogen, progesterone and testosterone) have been further implicated in mediating some of the cocaine-induced alterations. To better understand sex differences and the role of gonadal hormones in cocaine-induced locomotor and stereotypic behavior, intact and gonadectomized male and female Fischer rats were randomly assigned to either chronic cocaine (15 mg/kg) or saline treatments for 14 days followed by a challenge administration (7 days after the last cocaine/saline administration). Locomotor (ambulatory and rearing) and stereotypic activities were measured on days 1, 7 and 14 as well as after withdrawal/challenge with cocaine. Overall, intact female rats consistently showed a rapid (acquired by day 7) and longer lasting (persistent through the challenge dose) sensitization for all locomotor behaviors than any of the other groups. In contrast, intact males developed sensitization of these locomotor activities only in response to chronic cocaine administration, and after withdrawal and drug challenge the sensitization to cocaine-induced locomotor activity was no longer present. In female rats, gonadectomy affected ambulatory activity but not total and rearing responses after acute, sub-acute, chronic and challenge response to cocaine. On the other hand, castrated male rats were affected in cocaine-induced ambulatory activity but not rearing activity. In intact male rats, cocaine-induced stereotypic activity was rapidly and persistently sensitized after 7 days of cocaine administration, where gonadectomized male rats developed sensitization to cocaine induced stereotypic activity only after a challenge cocaine administration. Although cocaine induced stereotypic activity, no statistically significant differences were observed between intact and ovariectomized female rats or throughout the different lengths of cocaine administration. After a challenge of cocaine, corticosterone levels were induced in all experimental groups. Moreover, no differences in levels of benzoylecgonine, a cocaine metabolite, were observed. Similar to our previous observations after acute cocaine administration, after challenge of cocaine, an increase in progesterone and a decrease in testosterone levels were observed in intact females and males, respectively. In summary, endogenous hormones seem to be involved in the behavioral activation and development of sensitization to cocaine. PMID- 12113961 TI - Conditioned place preference to morphine in cannabinoid CB1 receptor knockout mice. AB - Recent reports have suggested an involvement of the brain cannabinoid system in the morphine-reward pathway. To address this question we evaluated whether CB1 receptor knockout mice would show a conditioned place preference to morphine. CB1 receptor knockout mice developed a strong place preference to 4 and 8 mg/kg morphine, similar to that in wild-type Swiss-Webster mice. This data thus does not support a contribution of the brain cannabinoid system to morphine reward. PMID- 12113960 TI - Nitric oxide accounts for an increased glycolytic rate in activated astrocytes through a glycogenolysis-independent mechanism. AB - The possible interference of nitric oxide (NO) in glucose metabolism was studied in activated astrocytes. Lipopolysaccharide (LPS) treatment triggered a NO mediated increase in glucose consumption and lactate production, suggesting an enhanced rate of glycolysis. Active glycogen synthesis was also observed after LPS treatment, but NO synthase inhibition was unable to prevent this effect. These results strongly suggest that endogenously-formed NO stimulates glycolysis through a glycogenolysis-independent mechanism in astrocytes. PMID- 12113963 TI - Endothelin B receptor deficiency augments neuronal damage upon exposure to hypoxia-ischemia in vivo. AB - The role of functional endothelin-B (ETB)-receptors on neuronal survival upon hypoxia-ischemia (HI) has been investigated in 14-day-old ETB-receptor-deficient spotting lethal (sl/sl) and wildtype (+/+) rats. Carotid ligation followed by exposure to 8% oxygen for 2 h produced distinct cortical and hippocampal neuronal damage. Damage severity 24 h after HI was mild to intermediate in +/+ rats whereas large cortical infarcts and profound apoptosis of the hippocampus evolved in sl/sl rats. The number of apoptotic cells in the dentate 24 h after HI amounted to 30 +/- 7 cells/0.1 mm(2) in sl/sl compared to 9 +/- 3 cells/0.1 mm(2) in wildtype rats (mean +/- S.E.M., n=10-11, P=0.0093). In-vitro hypoxia (15 h) resulted in a comparable increase in cell death in primary pure neuronal hippocampal cultures from both groups (49.8 +/- 1.6% in sl/sl, 51.4 +/- 0.9% in +/+, mean +/- S.E.M., n=5, P=0.0560). To conclude, absence of functional ETB receptors is associated with an increased susceptibility to HI in-vivo, which is not intrinsic to neurons. Antagonism of ETB receptors seems not to be desirable in ischemic stroke. PMID- 12113962 TI - Reduced aversion to oral capsaicin following neurotoxic destruction of superficial medullary neurons expressing NK-1 receptors. AB - Aversion to capsaicin (0.1-10 ppm) was assessed using a two-bottle paired preference paradigm, before and after intracisternal injection of substance P conjugated to saporin (SP-SAP) to ablate neurons in superficial medullary and cervical dorsal horn that express NK-1 receptors. Before SP-SAP, there was a concentration-dependent decrease in consumption of capsaicin with a threshold of 0.1-0.3 ppm. Following SP-SAP, significantly more capsaicin solution was consumed at 1- and 10-ppm concentrations. These results support a role for substance P in the mediation of high, but not low, levels of capsaicin-induced oral irritation. PMID- 12113965 TI - Improving the assessment of gestational age in a Zimbabwean population. AB - OBJECTIVES: To evaluate the performance and the utility of using birthweight adjusted scores of Dubowitz and Ballard methods of estimating gestational age in a Zimbabwean population. METHOD: The Dubowitz and the Ballard methods of estimating gestational age were administered to 364 African newborn infants with a known last menstrual period (LMP) at Harare Maternity Hospital. RESULTS: Both methods were good predictors of gestational age useful in differentiating term from pre-term infants. Our regression line was Y((LMP gestational age))=23.814+0.301*score for the Dubowitz and Y((LMP gestational age))=24.493+0.420*score for the Ballard method. Addition of birthweight to the regression models improved prediction of gestational age; Y((LMP gestational age))=23.512+0.219*score+0.0015*grams for Dubowitz and Y((LMP gestational age))=24.002+0.292*score+0.0016*grams for Ballard method. CONCLUSIONS: We recommend the use of our birthweight-adjusted maturity scales; the Dubowitz for studies of prematurity, and the Ballard for routine clinical practice. PMID- 12113967 TI - Post-partum maternity 'blues' as a reflection of newborn nursing care in Japan. AB - OBJECTIVES: To investigate the prevalence of post-partum 'blues' in mothers whose babies are cared for in a newborn nursery, compared with mothers providing rooming-in care. METHODS: Japanese normal primiparous women were prospectively studied from 1998 to 1999. The newborns of these mothers were managed in the newborn nursery or by rooming-in care. To diagnose maternity 'blues' and 'depression', the Stein's Questionnaire and the Edinburgh Postnatal Depression Scale were used. RESULTS: Ninety-seven and 93 women were managed by newborn nursery care and by rooming-in care, respectively. Of these women, a total of 181 women were considered for analysis. 'Blues' was noted in 31 of 92 mothers (33.7%) receiving newborn nursery care and in 18 of 89 (20.2%) receiving rooming-in care with a significant difference (P<0.05), and in 49 of 181 (27.1%) as a whole. The daily Stein's scores changed significantly during the 10 days post-partum in each group (P<0.0001). Post-partum 'depression' was observed in three mothers (3.4%) in the newborn nursery care group and in four (4.8%) of the rooming-in care group, not a significant difference, and in seven (4.1%) as a whole. CONCLUSION: Maternity 'blues' is experienced by 25% or more of Japanese primiparous women delivering healthy babies via uncomplicated delivery. The system of newborn nursery infant care may be a potential causal factor for maternity 'blues', although it should be confirmed by a prospective randomized trial. PMID- 12113966 TI - Induction of labor with prostaglandin E2 vaginal tablets in parous and grandmultiparous patients with previous cesarean section. AB - OBJECTIVES: To compare the outcome of induction of labor with prostaglandin E2 vaginal tablets between lower parity (parity 1-5) and grandmultiparous (parity >5) patients with a history of one previous lower segment cesarean section. METHODS: A prospective study of 113 patients conducted at King Faisal Military Hospital, Khamis Mushayt, Saudi Arabia during a 5-year period spanning January 1995 to December 1999. RESULTS: There were no statistically significant differences in the two groups regarding mean maternal age, dose of prostaglandin used, gestation at delivery, mean birth weight, P>0.05. Syntocinon augmentation was used in 16 (21.9%) of the lower parity patients compared with 8 (20.0%) in the grandmultiparas but this was not statistically significant, (P=0.677). However, there was a statistically significant difference in the cesarean section rate between the two groups, P=0.019. Although no cases of uterine hyperstimulation were recorded, there was one rupture of the uterus in each of the two groups of patients; 1.36% and 2.5%, respectively, but this was not statistically significant, P=1.000. CONCLUSIONS: The complications of induction of labor with prostaglandin E2 vaginal tablets in grandmultiparous patients with previous cesarean section were similar to those with lower parity but the cesarean section rate was significantly higher. However, larger studies are needed for validation. PMID- 12113968 TI - Digital examination compared to trans-perineal ultrasound for the evaluation of anal sphincter repair. AB - OBJECTIVE: To assess the adequacy of a third- or a fourth-degree laceration repair by comparing digital and trans-perineal ultrasound measurements. METHOD: During a 4-year period, 34 subjects without prior history of anal sphincter injury or fecal incontinence underwent ultrasound measurements of external anal sphincter muscle diameter and perineal length, which were compared to measurements obtained by digital examination. RESULTS: Pearson's correlation coefficients for comparing the digital external sphincter examination to trans perineal ultrasonography, and the digital perineal examination to trans-perineal ultrasonography were 0.88 and 0.40, respectively. Patients (n=4/34) whose external sphincter was identified as less than 1 cm by digital examination were found to have an external sphincter diameter of less than 1 cm by trans-perineal ultrasound. CONCLUSION: The digital perineum examination is a reliable method of measuring the external sphincter thickness and perineal body length immediately after primary repair. PMID- 12113970 TI - Gestational diabetes: implications of variation in diagnostic criteria. PMID- 12113969 TI - Adjuvant chemotherapy in stage I uterine endometrial carcinoma. AB - OBJECTIVE: We have assessed prognostic factors and the efficacy of adjuvant chemotherapy in stage I uterine endometrial carcinoma. METHODS: 251 primary surgically treated stage I patients were studied. Prognostic factors were evaluated and 5-year and 10-year survival rates were compared in patients with lymph-vascular space invasion to investigate whether adjuvant chemotherapy improves survival. RESULTS: The overall 5-year and 10-year survival rates were 94% and 93%. Multivariate analysis indicates that lymph-vascular space invasion is the most significant prognostic factor in both 5- and 10-year survival rates (P<0.001 at both times) and stage/depth of invasion is significant for the 10 year survival rate (P=0.04). Of 54 patients with lymph-vascular space invasion, statistically significant differences were observed in 10-year survival rate (P=0.02) between patients who had surgery followed by adjuvant chemotherapy (n=23) and patients who had surgery alone (n=31). Toxicities were mild to moderate (30%). CONCLUSIONS: The clinical importance of lymph-vascular space invasion and the efficacy of adjuvant chemotherapy were confirmed. This observation warrants a larger comparative study with adjuvant chemotherapy. PMID- 12113971 TI - Comparison of two transverse abdominal incisions for cesarean delivery. PMID- 12113972 TI - Perineal massage during pregnancy in primiparous women. PMID- 12113973 TI - Factors affecting the choice of delivery site and incorporation of traditional birth customs in a refugee camp, Thailand. PMID- 12113974 TI - A retrospective study on the reproductive outcome of the septate uterus corrected by hysteroscopic metroplasty. PMID- 12113975 TI - The risk of vaginal wall prolapse following sacro-spinous fixation. PMID- 12113976 TI - Age at diagnosis and other prognosticators in epithelial ovarian cancers. PMID- 12113977 TI - The Hyperglycemia and Adverse Pregnancy Outcome (HAPO) Study. AB - OBJECTIVE: The objective of the Hyperglycemia and Adverse Pregnancy Outcome (HAPO) Study is to clarify unanswered questions on associations of maternal glycemia, less severe than overt diabetes mellitus, with risks of adverse pregnancy outcome. This report describes the background and design of the HAPO Study. METHODS: HAPO is a 5-year investigator-initiated prospective observational study that will recruit approximately 25000 pregnant women in 10 countries. HAPO utilizes a Central Laboratory for measurement of key metabolic variables, a Clinical Coordinating Center, a Data Coordinating Center, and an independent Data Monitoring Committee. Glucose tolerance is assessed by a 75 g 2-h OGTT at 24-32 weeks' gestation. Results are unblinded to the woman and her caregivers if: fasting plasma glucose >5.8 mmol/l, 2-h plasma glucose >11.1 mmol/l or any plasma glucose <2.5 mmol/l. Random plasma glucose measurement is performed at 34-37 weeks or if symptoms suggest hyperglycemia; results are unblinded for values > or = 8.9 mmol/l. Sociodemographic and health history data are collected via questionnaire and medical record abstraction. Maternal blood is obtained for measurement of serum C-peptide and hemoglobin A1c (HbA(1C)), cord blood for serum C-peptide and plasma glucose, and a capillary specimen is taken between 1 and 2 h following delivery for neonatal plasma glucose. Neonatal anthropometrics are obtained, and follow-up data are collected at 4-6 weeks post-delivery. The primary outcomes to be assessed in relation to maternal glycemia are cesarean delivery, increased fetal size (macrosomia/LGA/obesity), neonatal morbidity (hypoglycemia), and fetal hyperinsulinism. PMID- 12113979 TI - Quantitative determination of a selective alpha-1a receptor antagonist in human plasma by high-performance liquid chromatography with tandem mass spectrometric detection. AB - Solid-phase extraction, utilizing a 96-well plate format, was used to isolate an alpha-1a receptor antagonist and internal standard from human plasma. Following the isolation procedure, the analyte and internal standard were separated and detected using reversed-phase HPLC coupled with atmospheric pressure chemical ionization (APCI) mass spectrometry operated in the positive ion multiple reaction monitoring (MRM) mode. Based upon the peak area ratio (analyte: internal standard) the analyte was quantified over a concentration range of 0.02-2 ng/ml. Assay validation results including parameters such as precision and accuracy are presented. The validated method was subsequently used to support human pharmacokinetic studies. PMID- 12113978 TI - New and rapid fully automated method for determination of tazobactam and piperacillin in fatty tissue and serum by column-switching liquid chromatography. AB - A sensitive and rapid HPLC assay for determining tazobactam and piperacillin in fatty tissue and serum is described. While the common methods need liquid-liquid extraction before the injection in a automated column switching HPLC, the new method works by direct injection of the filtered tissue extract or diluted serum in a automated column switching HPLC without any other pre-treatment. This was performed by the use of a NH2-precolumn and enrichment/transfer at different pH level. During the analyses, the NH2-precolumn was automatically regenerated with acetonitrile-water. The chromatogram peaks for piperacillin and tazobactam were identified by the retention time and quantified by peak area. The calibration curve was linear between 1 and 16 microg/ml. The quantification limit of tazobactam was about 1 microg/ml in fatty tissue extracts and in diluted serum (calculated for pure serum 2 microg/ml), respectively. For piperacillin it was less. The described procedure allows sample clean-up and determination of the antibiotic within 35 min. The chromatograms with this easy sample treatment had the same quantity of matrix peaks and in contrast to liquid-liquid extraction no loss of piperacillin. Because of the automatically rinsing of the NH2-precolumn during the chromatographic separation, more than 50 different biological samples could be measured with one NH2-precolumn without loss of performance. PMID- 12113980 TI - Behaviour of vanadate and vanadium-transferrin complex on different anion exchange columns. Application to in vivo 48V-labelled rat serum. AB - The behaviour of free [48V]vanadate and [48V]vanadium-transferrin complex was investigated on five different anion-exchange columns (Mono Q 5/5 HR, Hitrap Q HP, Sepharose Q FF, Sepharose DEAE FF and Hitrap Q XL). The recovery of both V compounds was quantitative. The peak shape and retention time of vanadate varied according to the type of column. The vanadium-transferrin complex also showed different elution patterns depending on the type of column. Especially in case of the Sepharose Q FF, Mono Q 5/5 HR and Hitrap Q XL columns the vanadium transferrin binding was degraded during elution on the column. The results clearly prove that care should be taken as to the choice of column for speciation purposes of vanadium compounds in order to prevent various artefacts showing up in the chromatograms. A Hitrap Q HP column was used to fractionate different vanadium compounds in rat serum. PMID- 12113982 TI - Determination of mirtazapine and its demethyl metabolite in plasma by high performance liquid chromatography with ultraviolet detection. Application to management of acute intoxication. AB - Mirtazapine is a new centrally acting noradrenergic and specific serotonin antidepressant, with an active demethyl metabolite. For toxicological purposes, a specific and accurate RP-HPLC assay was developed for the simultaneous plasma determination of these compounds. A linear response was observed over the concentration range 50-500 ng/ml. A good accuracy (bias <10%) was achieved for all quality controls, with intra-day and inter-day variation coefficients less than 8.3%. The lower limit of quantification was 20 ng/ml, without interferences with endogenous or exogenous components. This rapid method (run time <12 min) was used to manage three intoxications involving mirtazapine. PMID- 12113981 TI - Simultaneous quantitation of fatty acids, sterols and bile acids in human stool by capillary gas-liquid chromatography. AB - A simple method for the simultaneous gas-liquid chromatographic quantitation of fatty acids, sterols and bile acids from human fecal samples is described. The various compounds are directly converted into the n-butyl ester-trimethylsilyl ether derivatives, without prior isolation from the stool. Under these conditions, fecal bile acid derivatives are well resolved from each other and from those of fecal fatty acids and sterols without overlaps. The method was found to be reproducible and recoveries were similar to those obtained after exhaustive solvent extraction of fecal sterols, fatty acids and bile acids. Optimum derivatization conditions that allowed maximum recovery of fecal components with minimal destruction and application of the method for simultaneous bile acid, fatty acid and sterol analysis in human stool are described. PMID- 12113983 TI - Quantitation of the flavonoid wogonin and its major metabolite wogonin-7 beta-D glucuronide in rat plasma by liquid chromatography-tandem mass spectrometry. AB - This study described the application of liquid chromatography-tandem mass spectrometry for the quantitation of wogonin and its major metabolite in rat plasma. Only one conjugated metabolite with glucuronic acid was identified by chromatographic and electrospray multi-stage mass spectrometric assay. A derivatization reaction with 2-chlorethanol further demonstrated that the metabolite was wogonin-7 beta-D-glucuronide (W-7-G), not wogonin-5 beta-D glucuronide. Other conjugated metabolites, e.g., sulfates and glucosides, were not detected. The plasma concentration of free wogonin was determined using atmospheric pressure chemical ionization source in the selected reaction monitoring mode. The method had a lower limit of quantitation of 0.25 ng/ml for wogonin, which offered increased sensitivity, selectivity and speed of analysis over an existing method. Incubation of the plasma samples with beta-glucuronidase allows the quantitation of W-7-G. This quantitation method was successfully applied to a preclinical pharmacokinetic study of wogonin and its major metabolite, W-7-G, after an oral administration of 5 mg/kg wogonin to rats. PMID- 12113984 TI - Application of size-exclusion chromatography-inductively coupled plasma mass spectrometry for fractionation of element species in seeds of legumes. AB - Fractionation of soluble species of P, Mn, Fe, Co, Ni, Cu, Zn, Se and Mo in pea and lentil seeds was made by on-line hyphenation of size-exclusion chromatography (SEC) and inductively coupled plasma mass spectrometry. Seed samples were extracted with 0.02 mol l(-1) Tris-HCl buffer solution, pH 7.5. SEC was performed on Superdex 75 and Superdex Peptide columns (300 x 10 mm) with the same buffer solution as the mobile phase. Monitoring of oxide ion 47(PO)+ was used for detection of phosphorus compounds. Other elements were detected as ions of 55Mn, 57Fe, 59Co, 62Ni, 65Cu, 66Zn, 82Se and 95Mo nuclides. Elements in individual elution zones were quantified using external calibration. Complete chromatographic recoveries of elements were found in cases of phosphorus, nickel and copper. Substantial parts of manganese and zinc, as well as traces of cobalt, selenium and molybdenum are retained on the column. Injection of EDTA solution removes these elements from the column. Chromatographic profiles of pea and lentil samples are very similar for all elements except Mo. Main element species in the high-molecular-mass region (approx. 190,000 rel. mol. mass unit) were detected in case of Fe. Low-molecular-mass species (<2000 rel. mol. mass unit) as major element forms are typical for Cu and Zn. PMID- 12113985 TI - Validated liquid chromatographic method for the determination of bexarotene in human plasma. AB - A new liquid chromatographic method is described for the determination of the anti-tumour agent bexarotene in human plasma over the range 0.500-1500 ng/ml, using 1 ml of sample. Sample preparation consists of liberating the analyte from plasma lipids by adding acetonitrile, followed by acidification of the plasma and liquid extraction using a mixture of isoamyl alcohol and pentane or hexane. Separation and quantitation are performed by reversed-phase column liquid chromatography with fluorescence detection. Parameters affecting the performance of these steps are discussed. Validation results on linearity, selectivity, accuracy, precision, recovery and stability are shown, as well as the application of the method to samples from clinical trials. PMID- 12113986 TI - Enantioselective determination of (R)- and (S)-sotalol in human plasma by on-line coupling of a restricted-access material precolumn to a cellobiohydrolase I-based chiral stationary phase. AB - A liquid chromatographic column-switching method for the enantioselective determination of (RS)-sotalol in plasma was developed and validated. The method is based on the on-line coupling of a precolumn filled with the restricted access material LiChrospher ADS to a cellobiohydrolase I-based chiral stationary phase (CSP). The plasma samples were injected onto the precolumn using a mobile phase containing 1% methanol in 10 mM phosphate buffer at pH 7.4 for 10 min for the removal of matrix components. The analytes were transferred to the CSP for their enantiomeric separation by backflushing the precolumn with 15% 2-propanol in 10 mM phosphate buffer (pH 7.0) including 0.05 mM EDTA. The quantitative determination of the sotalol enantiomers was possible upon addition of the internal standard (S)-atenolol. The method was validated showing a good linearity in the concentration range from 25 to 1000 microg l(-1) for each enantiomer. The average values of the intra- and inter-day variability were 1.17% and 3.42%, respectively, for (R)-sotalol and 1.24% and 1.99%, respectively, for (S)-sotalol. The applicability of the method to real world samples has been proven by means of two pharmacokinetic studies. They revealed that the pharmacokinetic properties of the sotalol enantiomers do not differ significantly neither for healthy young volunteers after single dose application nor for elder patients in the steady state. PMID- 12113987 TI - Sensitive determination method of estradiol in plasma using high-performance liquid chromatography with electrochemical detection. AB - The improvement in the sensitive determination method of estradiol using HPLC with electrochemical detection is described. The improvement was due to the optimization of the potential applied to the electrode of the analytical cell and employment of a guard cell. The detection conditions were optimized from the electrochemical properties of estradiol in acidic and alkaline eluents. The employment of the guard cell drastically decreased the background noise without any reduction in the response of estradiol, and contributed to improvement in the sensitivity. The optimized method combined with pretreatment by liquid-liquid extraction was applied to the determination of estradiol in rat plasma. The detection limit of 8 pg for the standard solution and 24 pg for the plasma sample, which was about 6-8-fold more sensitive compared to the previous reports, was attained. PMID- 12113988 TI - Validated capillary gas chromatographic-mass spectrometric assay to determine 2 methylcitric acid I and II levels in human serum by using a pulsed splitless injection procedure. AB - BACKGROUND: Despite some clinical applications of 2-methylcitric acid (2-MCA) determination in urine and amniotic fluid, a diagnostic use of 2-MCA levels in serum is not common practice. This could be related to the complexity of the assay, or possibly to unawareness of other feasible clinical applications. METHODS: The levels of the diastereomers 2-MCA I and II in human serum were determined by GC-MS based on a method using a pulsed splitless injection technique. A stable isotope dilution principle was modified considering the diastereomer ratio and impurities of the internal standard. Precision parameters as well as recovery rates of the assay were determined. Reference intervals for 2 MCA(total), 2-MCA I and II levels were obtained in 52 healthy volunteers (31 female, 21 male, mean age 41.7+/-14.4 years). RESULTS: 2-MCA was readily detected in each sample of serum, as well as in urine, cerebrospinal fluid and amniotic fluid. The limit of detection was 10 nmol/l for 2-MCA(total). The internal standard showed a diastereomer ratio of 2-MCA II-d3 to 2-MCA I-d3 of 0.83+/-0.05, its chemical purity had to be corrected to 90.5+/-0.5%. In concentrations of 446, 750 and 1256 nmol/l 2-MCA(total), recovery rates of 98.5, 93.7 and 88% with a mean intra-assay RSD of 1.5% were determined. The day-to-day precision was 10% RSD (SD 40 nmol/l) for 2-MCA(total) obtained with a pooled serum sample at a concentration of 401 nmol/l 2-MCA(total) over a period of 5 months (n=17). The normal range for 2-MCA(total) in human serum was calculated as 81-266 nmol/l confirming previous findings. CONCLUSIONS: The GC-MS assay using a pulsed splitless injection procedure ensures a good response to differing concentrations of 2-MCA in various specimens. Considering exact determination of the diastereomer ratio as well as the purity of the internal standard, the assay offers good precision and recovery for 2-MCA I and II levels in serum. PMID- 12113990 TI - Determination of 9-nitrocamptothecin and its metabolite 9-aminocamptothecin in human plasma using high-performance liquid chromatography with ultraviolet and fluorescence detection. AB - A high-performance liquid chromatography assay is described for the determination of the investigational anti-cancer drug 9-nitrocamptothecin (9-NC) and its metabolite 9-aminocamptothecin (9-AC) as the total of their lactone and carboxylate forms. The sample pre-treatment consisted of a deproteinisation step and a quantitative acid-catalyzed conversion of all 9-NC and 9-AC into their lactone forms and a subsequent solid-phase extraction. Redissolved extracts were analyzed on a Prodigy analytical column, using a mixture of methanol-phosphate buffer (pH 2.5). Detection was concomitantly performed with UV for 9-NC and fluorimetrically for 9-AC. The lower limit of quantifications were 10 ng/ml and 2.5 ng/ml for the determination of 9-NC and 9-AC, respectively, using 500 microl of plasma. The presented method was successfully applied to a clinical pharmacokinetic study. PMID- 12113989 TI - Simultaneous determination of indinavir, ritonavir and lopinavir (ABT 378) in human plasma by high-performance liquid chromatography. AB - An isocratic reversed-phase high-performance liquid chromatographic method with ultraviolet detection at 205 nm has been validated for the determination of indinavir, ritonavir and lopinavir (ABT 378) in human plasma. The ritonavir analogue A-86093.0 was used as internal standard. Good chromatographic separation was achieved using a stainless steel column packed with 5 microm Phenomenex phenyl hexyl material operated at 40 degrees C, and a mobile phase consisting of acetonitrile-10 mM potassium phosphate buffer (50:50, v/v). The calibration curve for indinavir was linear over the range of 50 to 1000 microg/l while the ritonavir and lopinavir calibration curves were linear over the range of 100 to 15,000 microg/l. The lower limit of quantitations for indinavir, ritonavir and lopinavir were 50, 100 and 100 microg/l, respectively, using 500 microl of human plasma. The validation data showed that the assay is sensitive, specific and reproducible for determination of indinavir, ritonavir and lopinavir. This method is being used in a therapeutic drug monitoring service to quantitate these therapeutic agents in patients infected with human immunodeficiency virus. PMID- 12113991 TI - Application of capillary zone electrophoresis to the analysis and to a stability study of cephalosporins. AB - The applicability of capillary zone electrophoresis (CZE) for the determination of cephalosporin antibiotics has been studied. In the case of the separation conditions optimised for fourteen cephalosporins, the precision of migration times was smaller than 1.3% RSD, and the values of the limit of detection ranged between 0.42 and 1.62 microg/ml. The proposed CZE method was applied to study the stability of cephalosporins in water at different temperatures (+25, +4 and -18 degrees C). It was established that the degradation of most cephalosporins was not higher than 20% at room temperature within 4 h of dissolution of these antibiotics. PMID- 12113992 TI - Simple liquid chromatography-electrospray ionization mass spectrometry method for the routine determination of salmon calcitonin in serum. AB - A simple liquid chromatography-electrospray ionization mass spectrometry (LC-ESI MS) method was developed for the quantification of salmon calcitonin (sCT) in serum. Serum samples from rats and dogs were deproteinized and freeze-dried. The residue was then reconstituted with 57% acetonitrile in water containing 0.1% trifluoroacetic acid and 0.005% benzalkonium chloride. A 20-microl aliquot of the reconstituted solution was injected onto a polymer based RP-C18 column, The outlet was connected to an ion-trap mass spectrometer equipped with an ESI source, and spectra were recorded in a positive-ion, selected-ion monitoring mode. The limit of quantification of the method was 10 ng/ml. Biexponential curves were observed for the temporal serum concentration of sCT following intravenous administration of sCT to rats (100 microg/kg) and dogs (250 microg/kg), resulting in reasonable pharmacokinetic parameters. The present method appears applicable to routine analysis of serum sCT in pharmacokinetic studies with good selectivity, accuracy and precision. PMID- 12113993 TI - Flavor experiences during formula feeding are related to preferences during childhood. AB - As part of a program of research designed to investigate the long-term effects of early feeding experiences, the present study exploited the substantial flavor variation inherent in three classes of commercially available infant formulas and determined whether flavor preferences during childhood differed as a function of the class of formula (i.e., milk, soy, hydrolysate) that 4- to 5-year-old children were fed during their infancy. Age appropriate, game-like tasks that were fun for children and minimized the impact of language development were used to examine their preferences for a wide range of food-related odor qualities including infant formulas, as well as the flavor of milk-based and hydrolysate formulas and plain, sour- and bitter-flavored apple juices. Formula type influenced children's flavor preferences when tested several years after their last exposure to the formula. When compared to children who were fed milk-based formulas (n=27), children fed protein hydrolysate formulas (n=50) were more likely to prefer sour-flavored juices, as well as the odor and flavor of formulas, and less likely to make negative facial expressions during the taste tests. Those fed soy formulas (n=27) preferred the bitter-flavored apple juice. That the effects of differential formula feeding also modified children's food preferences is suggested by mothers' reports that children fed hydrolysate or soy formulas were significantly more likely to prefer broccoli than were those fed milk formulas. These data are consistent with the hypothesis that flavor experiences influence subsequent flavor preferences even several years following the early experience. PMID- 12113994 TI - Indications of improved cognitive development at one year of age among infants born very prematurely who received care based on the Newborn Individualized Developmental Care and Assessment Program (NIDCAP). AB - BACKGROUND AND OBJECTIVE: Care based on the Newborn Individualized Developmental Care and Assessment Program (NIDCAP) has been reported to exert a positive impact on the development of prematurely born infants. The aim of the present investigation was to determine the effect of such care on the 1-year development of infants born with a gestational age of less than 32 weeks. METHODS: All surviving infants (11 in the NIDCAP group and 9 in the control group) were assessed employing the Bayley Scales of Infant Development at 1 year of corrected age. RESULTS: The Mental Developmental Index (MDI) of children who had received care according to NIDCAP was higher [88 (72-114)] [median (range)] than the corresponding value for the control children [78 (50-82)] (p=0.01). The odds ratio for being alive with an MDI>80 was 14 (95% CI; 1.4-141.5) in favour of the intervention group. However, the Psychomotor Developmental Indices (PDI) were not significantly different [85 (61-108) and 69 (50-114), respectively] (p=0.23). CONCLUSION: Our findings indicate that care based on the NIDCAP might have a positive impact on the cognitive development of infants born very prematurely. PMID- 12113995 TI - Sleep-wake patterns in preterm infants and 6 month's home environment: implications for early cognitive development. AB - AIMS: This study examined the relationship between early organization of sleep wake states and developmental outcome at 6-month-old premature infants. STUDY DESIGN: This was a prospective randomized study that evaluated the sleep-wake states of healthy premature infants in the nursery environment for two successive 72-h periods, at 32 and 36 weeks gestational age. SUBJECTS: Thirty-four healthy premature infants. OUTCOME MEASURES: Three sleep-wake parameters: percent of quiet sleep, activity level and total amount of sleep, were studied with miniature activity monitors attached to the infant's ankles. The rearing environments of the infants were also assessed at 6 months of age, using the HOME Inventory. Finally, child developmental status was recorded by means of the Mental Development Index (MDI) of the Bayley Scales for Infant Development, at a chronological age of 6 months. RESULTS: Lower total time spent in night sleep, higher mean level of night activity level, and a later rich home environment were all predictive of higher Bayley scores (MDI) at a chronological age of 6 months. Regression analysis indicated that early biological maturity was more strongly related to the child's developmental status than later home environment, although both contributed to the prediction. CONCLUSIONS: These results suggest that biological factors may be significant predictors early in development, whereas the impact of environmental influences increases with development. PMID- 12113996 TI - Muscle power development during the first year of life predicts neuromotor behaviour at 7 years in preterm born high-risk infants. AB - The aim of the study was to find if neurological function during the first year of life could predict neuromotor behaviour at 7 years of age in children born preterm with a high risk. A follow-up study of neuromotor behaviour in 52 children at a mean age of 3, 6, 12 months (corrected age) and 7 years was performed. All children were born with a gestational age less than 32 weeks and/or a birthweight under 1500 g and the infants were categorised according to their medical history in the three highest categories of the 'Neonatal Medical Index' (NMI, from category I to V, from few to serious complications). In addition, neonatal cerebral ultrasound abnormalities were used to divide the infants further into the different NMI categories. At 3 and 6 months, the relationship between active and passive muscle power was measured in shoulders, trunk and legs and (a)symmetry between right and left was noted. The results at 3 and 6 months were ranged from 1 for optimal to 5 for poor muscle power regulation. At 12 months of age, a neurological examination was done with special emphasis on the assessment of postural control, spontaneous motility, hand function and elicited infantile reactions with special attention to (a)symmetry. Outcome at 12 months was expressed as percentage of the optimal score on each subcategory. At 7 years, the motor behaviour study based on Touwen's examination for minor neurological dysfunction was performed. This investigation focuses on different functions, such as hand function, quality of walking, posture, passive muscle tone, coordination and diadochokinesis. The outcome was expressed as percentage of the optimal score on the combined subcategories. The best prediction of neuromotor behaviour at 7 years was assessed with stepwise linear multiple regression, using as potential predictors perinatal factors and outcome of motor behaviour at the corrected age of 3, 6 and 12 months. At 7 years none of the children scored 100% on the combined subcategories, 15 children (29%) scored between 75% and 99%, whereas 15 children scored less than 50%. Neuromotor behaviour at 7 years could be predicted by the NMI categorisation and gender with a sensitivity of 92% (specificity 47%; positive and negative predictive value 81% and 70%). No direct relation was found between neuromotor behaviour and cerebral ultrasound classification only, days on the ventilator and/or continuous positive airway pressure, birthweight, gestational age and dysmaturity. The best predictor of neuromotor behaviour at 7 years was the combination of outcome of muscle power in shoulders and legs at 3 months and postural control at 12 months, taking into account the gender of the child (sensitivity 95%; specificity 40%; positive predictive value 80%; negative predictive value 75%). PMID- 12113997 TI - Relation between oligohydramnios and spinal flexion in the human fetus. AB - BACKGROUND: Oligohydramnios, a severe reduction in the volume of amniotic fluid, is associated with fetal lung hypoplasia but underlying processes are unclear. Studies in sheep suggest that oligohydramnios may lead to lung hypoplasia by causing increased flexion of the fetal spine, but this has not been demonstrated in the human, which has a different uterine anatomy. AIMS: Our aims were to quantify spinal flexion in the human fetus and to determine the relationship between oligohydramnios and the degree of spinal flexion. SUBJECTS AND METHODS: In 35 pregnancies, we used ultrasonography to assess amniotic fluid volume and to image the fetal spine so as to provide an index of mean spinal radius between the upper thoracic and lumbar spine. In 17 pregnancies with evidence of oligohydramnios resulting from premature rupture of membranes (mean +/- SEM gestation 28.8 +/-1.4 weeks), the mean radius of fetal spinal curvature between the upper thorax and sacrum was compared with values from 18 control fetuses with normal amounts of amniotic fluid (at 28.9 +/- 1.6 weeks). RESULTS: In each fetus, the spine could be imaged and the mean radius of curvature calculated. Oligohydramnios was associated with a significantly increased degree of fetal spinal flexion compared to controls. CONCLUSION: In human pregnancy, oligohydramnios is associated with increased flexion of the fetal spine, which is likely due to reduced uterine volume. This could contribute to the development of fetal lung hypoplasia, fetal immobility and other fetal anomalies. PMID- 12113998 TI - Brain electrical tomography in depression: the importance of symptom severity, anxiety, and melancholic features. AB - BACKGROUND: The frontal lobe has been crucially involved in the neurobiology of major depression, but inconsistencies among studies exist, in part due to a failure of considering modulatory variables such as symptom severity, comorbidity with anxiety, and distinct subtypes, as codeterminants for patterns of brain activation in depression. METHODS: Resting electroencephalogram was recorded in 38 unmedicated subjects with major depressive disorder and 18 normal comparison subjects, and analyzed with a tomographic source localization method that computes the cortical three-dimensional distribution of current density for standard electroencephalogram frequency bands. Symptom severity and anxiety were measured via self-report and melancholic features via clinical interview. RESULTS: Depressed subjects showed more excitatory (beta3, 21.5-30.0 Hz) activity in the right superior and inferior frontal lobe (Brodmann's area 9/10/11) than comparison subjects. In melancholic subjects, this effect was particularly pronounced for severe depression, and right frontal activity correlated positively with anxiety. Depressed subjects showed posterior cingulate and precuneus hypoactivity. CONCLUSIONS: While confirming prior results implicating right frontal and posterior cingulate regions, this study highlights the importance of depression severity, anxiety, and melancholic features in patterns of brain activity accompanying depression. PMID- 12113999 TI - Proton magnetic resonance spectroscopic imaging in pediatric major depression. AB - BACKGROUND: Neurobiologic abnormalities in dorsolateral prefrontal cortex (DLPFC) are believed to be involved in the pathophysiology of major depressive disorder (MDD). Although MDD commonly emerges during childhood and adolescence, to our knowledge, no prior study has examined the DLPFC in pediatric patients with MDD. METHODS: In this study, choline compounds (Cho), N-acetylaspartate (NAA), and creatine/phosphocreatine (Cr) were measured in left and right DLPFC using a multislice proton magnetic resonance spectroscopic imaging sequence with validated phantom replacement methodology in 11 treatment-naive MDD patients, 10 16 years of age, and 11 case-matched healthy control subjects. RESULTS: A significant increase in Cho was observed in left but not right DLPFC in MDD patients versus control subjects (32.5% higher). No significant differences in NAA or Cr were observed between case-control pairs. CONCLUSIONS: These results provide new evidence of localized functional neurochemical marker alterations in left DLPFC in pediatric MDD. Our results must be considered preliminary, however, given the small sample size. PMID- 12114000 TI - Regional prefrontal gray and white matter abnormalities in bipolar disorder. AB - BACKGROUND: Previous magnetic resonance imaging (MRI) studies indicate that compared with healthy volunteers, patients with bipolar disorder have structural and functional abnormalities in the prefrontal cortex. The aim of this study was to investigate differences in prefrontal subregions between bipolar patients and healthy subjects. METHODS: Bipolar patients hospitalized for a manic episode (n = 17), and demographically matched healthy volunteers (n = 12) were recruited. Contiguous 1-mm coronal T1-weighted MRI slices were obtained using a Picker 1.5 Tesla scanner. The gray and white matter volumes of five prefrontal subregions of interest were measured: superior, middle, inferior, cingulate, and orbital. RESULTS: Bipolar patients had smaller left prefrontal gray matter volumes, specifically in the middle and superior subregions and smaller right prefrontal gray matter volumes, specifically in the inferior and middle subregions. White matter differences were not observed in any of the prefrontal subregions. CONCLUSIONS: The results suggest that bipolar patients have subregion-specific gray matter volume reductions in the prefrontal cortex as compared to healthy subjects. Further investigations into the role of specific prefrontal subregions in bipolar disorder are warranted. PMID- 12114001 TI - Disruption of posterior brain systems for reading in children with developmental dyslexia. AB - BACKGROUND: Converging evidence indicates a functional disruption in the neural systems for reading in adults with dyslexia. We examined brain activation patterns in dyslexic and nonimpaired children during pseudoword and real-word reading tasks that required phonologic analysis (i.e., tapped the problems experienced by dyslexic children in sounding out words). METHODS: We used functional magnetic resonance imaging (fMRI) to study 144 right-handed children, 70 dyslexic readers, and 74 nonimpaired readers as they read pseudowords and real words. RESULTS: Children with dyslexia demonstrated a disruption in neural systems for reading involving posterior brain regions, including parietotemporal sites and sites in the occipitotemporal area. Reading skill was positively correlated with the magnitude of activation in the left occipitotemporal region. Activation in the left and right inferior frontal gyri was greater in older compared with younger dyslexic children. CONCLUSIONS: These findings provide neurobiological evidence of an underlying disruption in the neural systems for reading in children with dyslexia and indicate that it is evident at a young age. The locus of the disruption places childhood dyslexia within the same neurobiological framework as dyslexia, and acquired alexia, occurring in adults. PMID- 12114002 TI - Stimulation of protein kinase a activity in the rat amygdala enhances reward related learning. AB - BACKGROUND: Drug addiction in humans is associated with abnormal metabolic activity within the amygdala and heightened control of behavior by drugs and drug related (conditioned) stimuli. Drug-induced neuroadaptations, including activation of cAMP (cyclic adenosine monophosphate)-dependent protein kinase A (PKA), within the amygdala may contribute to the synaptic plasticity and reward related learning that underlies pathologic behavior in addicted individuals. METHODS: In this study, we tested the hypothesis that stimulation of PKA activity within the rat amygdala would facilitate the acquisition of Pavlovian approach behavior, a measure of reward-related learning. RESULTS: Intraamygdala infusions of Sp-cAMPS (which activates PKA) produced concentration-dependent enhancements of the acquisition of approach to a conditioned stimulus that predicted water availability; intraamygdala infusions of cholera toxin (which elevates cAMP levels) produced a similar effect. Conversely, intraamygdala infusions of Rp cAMPS, an inhibitor of PKA, impaired acquisition of approach behavior. CONCLUSIONS: Together, these data demonstrate that stimulation of PKA activity in the amygdala can facilitate reward-related learning and suggest that neuroadaptative changes in the PKA pathway within this brain region may be a mechanism by which chronic drug abuse alters the control of behavior by drug associated stimuli. PMID- 12114003 TI - Reduced hippocampal volume and total white matter volume in posttraumatic stress disorder. AB - BACKGROUND: Reduced hippocampal volumes in posttraumatic stress disorder (PTSD) patients are thought to reflect specific changes of this structure. Previous magnetic resonance imaging (MRI) studies have not consistently examined indices of overall brain atrophy, therefore it cannot be completely ruled out that hippocampal changes are explained by whole-brain atrophy. The purpose of this study was to assess hippocampal and whole-brain volume in civilian PTSD. METHODS: Twelve subjects with PTSD and 10 control subjects underwent brain MRI. Hippocampal volumes were visually quantified using a computerized volumetric program. Whole-brain volumes were obtained with automated k-means-based segmentation. RESULTS: No differences were found in intracranial volumes (ICV). Subjects with PTSD had higher cerebrospinal fluid (CSF)/ICV ratios and lower white matter/ICV ratios, consistent with generalized white matter (WM) atrophy. The effect of age on CSF/ICV was more pronounced in the PTSD group. Subjects with PTSD had smaller absolute and normalized bilateral hippocampal volumes. These differences persisted after adjusting for lifetime weeks of alcohol intoxication. Posttraumatic stress disorder and depression scores correlated negatively with left hippocampal volume, but PTSD scores were a better predictor of hippocampal volumes. CONCLUSIONS: Our results replicate previous findings of reduced hippocampal volume in PTSD but also suggest independent, generalized, white matter atrophy. PMID- 12114004 TI - rCBF differences between panic disorder patients and control subjects during anticipatory anxiety and rest. AB - BACKGROUND: Our goal was to identify brain structures involved in anticipatory anxiety in panic disorder (PD) patients compared to control subjects. METHODS: Seventeen PD patients and 21 healthy control subjects were studied with H(2)(15)O positron emission tomography scan, before and after a pentagastrin challenge. RESULTS: During anticipatory anxiety we found hypoactivity in the precentral gyrus, the inferior frontal gyrus, the right amygdala, and the anterior insula in PD patients compared to control subjects. Hyperactivity in patients compared to control subjects was observed in the parahippocampal gyrus, the superior temporal lobe, the hypothalamus, the anterior cingulate gyrus, and the midbrain. After the challenge, the patients showed decreases compared to the control subjects in the precentral gyrus, the inferior frontal gyrus, and the anterior insula. Regions of increased activity in the patients compared to the control subjects were the parahippocampal gyrus, the superior temporal lobe, the anterior cingulate gyrus, and the midbrain. CONCLUSIONS: The pattern of regional cerebral blood flow activations and deactivations we observed both before and after the pentagastrin challenge was the same, although different in intensity. During anticipatory anxiety more voxels were (de)activated than during rest after the challenge. PMID- 12114005 TI - Neuropeptide-Y, cortisol, and subjective distress in humans exposed to acute stress: replication and extension of previous report. AB - BACKGROUND: We previously reported that stress-related release of cortisol and neuropeptide-Y (NPY) were significantly and positively associated in U.S. Army soldiers participating in survival training. Furthermore, greater levels of NPY were observed in individuals exhibiting fewer psychologic symptoms of dissociation during stress. This study tested whether these findings would be replicated in a sample of U.S. Navy personnel participating in survival school training. METHODS: Psychologic as well as salivary and plasma hormone indices were assessed in 25 active duty personnel before, during, and 24 hours after exposure to U.S. Navy survival school stress. RESULTS: Cortisol and NPY were significantly and positively associated during stress and 24 hours after stress; NPY and norepinephrine (NE) were significantly and positively related during and 24 hours after stress. There was a significant, negative relationship between psychologic distress and NPY release during stress. Finally, psychologic symptoms of dissociation reported at baseline predicted significantly less NPY release during stress. CONCLUSIONS: These data replicate our previous studies demonstrating that acute stress elicits NPY release and that this release is positively associated with cortisol and NE release. These data also replicate our previous finding that greater levels of NPY release are associated with less psychologic distress suggesting that NPY confers anxiolytic activity. PMID- 12114006 TI - A further support to the hypothesis of a link between serotonin, autism, and the cerebellum. PMID- 12114008 TI - A mixed-effects time-to-event analysis of the relationship between first lactation lameness and subsequent lameness in dairy cows in the UK. AB - Data for 611 second-lactation and 251 third-lactation cows were examined using mixed-effects time-to-event models to determine the shape of the hazard, quantify relative risk and estimate herd- and sire-level variation in time to lameness. The semi-parametric Cox and fully parametric Weibull models were suggested from univariable Kaplan-Meier plots. Time to all-lameness, claw-horn lameness and skin lameness were modelled. Explanatory variables were season of current-lactation calving, age at first calving and first-lactation lameness history (whether all lameness or claw-horn and skin lameness). In mixed-effects models of lactation-2 lameness, previously lame cows had a significantly increased hazard (hazard ratio (HR)=2.0 for all types of lameness and HR=3.2 for claw-horn lameness) compared to those not previously lame. These relationships were less marked in the third lactation. There was little evidence for an effect of age at first calving, whilst possible differences between calving seasons were observed. The hazard function suggested that the rate of lameness was roughly flat across each lactation. Herd-level variation was more evident for infectious foot diseases. The contribution of the sire increased with parity and might be important for sole ulcer and white-line disease. PMID- 12114009 TI - Risk factors for Aujeszky's-disease seropositivity of swine herds of a region of northern Italy. AB - Vaccination programs to control Aujeszky's-disease virus (ADV) using gE-deleted vaccines are being considered in several European countries. Knowledge of factors influencing ADV-seropositivity for vaccinated herds might contribute to the success of these programs. A multivariable analysis of ADV-seropositivity in 1248 swine herds (332 farrow-to-finish, 260 farrow-to-feeder and 656 finishing herds) in the Emilia-Romagna region of Italy revealed that (1) high pig density (number of pigs in a 6-km radius), (2) gilt purchasing, and (3) increased number of fattening pigs were risk factors for farrow-to-finish herds. In farrow-to-feeder herds, ADV-seropositivity was related to (1) increased number of breeders, (2) heavy-gilt purchasing, and (3) increased pig density. In finishing herds, (1) increased herd size was related to ADV-seropositivity, whereas (2) periodic rearing suspension was protective. PMID- 12114010 TI - Ecto-, endo- and haemoparasites in free-range chickens in the Goromonzi District in Zimbabwe. AB - A cross-sectional study determined the prevalence of ecto-, endo- and haemoparasites in free-range chickens from the Goromonzi District, Zimbabwe. Fifty young and 50 adult birds were selected randomly. All chickens harboured ecto- and endoparasites, and 32% were infected with haemoparasites. Eight different ectoparasites were identified; the more prevalent ones had the following prevalences (young, %; adult, %): Argas persicus (6; 14), Cnemidocoptes mutans (6; 32), Echidnophaga gallinacea (72; 74), Goniocotes gallinae (0; 22), Menacanthus stramenius (90; 88) and Menopon gallinea (24; 66). The prevalences of C. mutans, G. gallinae and M. gallinae were higher in adults compared to young chickens. The mean (+/-S.D.) number of helminth species per chicken was 6.7+/-2.0 for young chickens and 6.4+/-2.0 for adult chickens with a range of 1-10 for young chickens and a range of 1-11 for adult chickens. The most prevalent nematodes identified were (with prevalence in % for young/adult birds): Allodapa suctoria (76; 72), Ascaridia galli (48; 24), Gongylonema ingluvicola (28; 56), Heterakis gallinarum (64; 62) and Tetrameres americana (70; 62). For cestodes the prevalences were: Amoebotaenia cuneata (60; 68), Hymenolepis spp. (62; 80), Raillietina echinobothrida (66; 34), Raillietina tetragona (94; 100) and Skrjabinia cesticillus (50; 76). The young chickens had higher prevalences of A. galli and R. echinobothrida compared to adults, but lower prevalence of G. ingluvicola and S. cesticillus. Eimeria spp. oocysts were isolated in 36% of 47 investigated samples. The prevalence was 47% for young chickens and 18% for adult chickens. Prevalences (in %) of haemoparasites in young and adult chickens were: Aegyptinella pullorum (7; 6), Leucocytozoon sabrazesi (3; 1), Plasmodium gallinaceum (8; 6) and Trypanosoma avium (2; 3). PMID- 12114011 TI - A simulation of Johne's disease control. AB - A dynamic and stochastic simulation model (the "JohneSSim model") was developed to evaluate the economic and epidemiological effects of different strategies for control of paratuberculosis in dairy herds. Animals occupy one of the six defined infection states; the spread of Johne's disease is modeled with five infection routes. Many different dairy farm situations can be simulated. Control strategies that can be simulated are: (1) test-and-cull; (2) calf hygiene management; (3) vaccination and (4) grouping of animals. Losses are caused by: (1) reduced milk production; (2) diagnosis and treatment costs; (3) lower slaughter value of cows and (4) sub-optimal culling. The benefits were calculated as reduction in the losses caused by Johne's disease; the costs of each strategy were calculated on the basis of actual costs of each item; and net present value (NPV) was calculated as benefits minus costs. Herd and prevalence data from The Netherlands and Pennsylvania, USA were used. In both situations, a low true mean prevalence within 20 years could be reached only when all calf management tools were applied. The Dutch control program (PPN) was on average economically attractive (with or without labor costs, the average NPV was Euro 1183 and 12,397, respectively). In Pennsylvania, contract heifer rearing and improved calf hygiene reduced the prevalence effectively and had large economic benefits (US$ 43,917 for 20-year period) if the calves were sent to the heifer facility while very young. Validation with data from 21 infected Dutch dairy farms (as well as face validation: comparison of the results of the JohneSSim model with experiences of Johne's experts) supported the basic assumptions in the model. PMID- 12114013 TI - A meta-analysis comparing the effect of vaccines against Mycoplasma hyopneumoniae on daily weight gain in pigs. AB - Our aims were to evaluate the published literature concerning the effect of swine vaccination against Mycoplasma hyopneumoniae on the average daily weight gain (ADWG). This was done by re-evaluating the influence of selected factors on ADWG by a meta-analysis of published studies from 1991 to 1999, fulfilling certain inclusion criteria. With ADWG as the outcome, an analysis of variance was performed for such variables as treatment, vaccination schedule, age during study, housing system and publication quality. Each clinical trial was considered as a random effect and the numbers of pigs in each trial were weightings. Of 63 published studies, 16 describing three commercial vaccines fulfilled the criteria for the meta-analysis. Due to few studies with one of the vaccines (n=3), only two vaccines were included. Vaccinated pigs gained an average of 592g (S.E.=15) with Stellamune and 590g (S.E.=15) with Suvaxyne compared to non-vaccinated pigs that gained an average of 569g (S.E.=14)(P<0.01) when adjusted for age during the study. Vaccine type, vaccination schedule, housing system and publication quality were not significantly associated with ADWG. PMID- 12114014 TI - Probability of and risk factors for introduction of infectious diseases into Dutch SPF dairy farms: a cohort study. AB - A 2-year cohort study was conducted to investigate the probability of disease introduction into Dutch dairy farms. The farms were tested regularly for diseases and were visited biannually to collect management data. Ninety-five specific pathogen-free (SPF) dairy farms were selected from a database of bovine herpesvirus type 1 (BHV1)-free farms to study the probability of, and risk factors for, introduction of BHV1, bovine viral diarrhoea virus (BVDV), Salmonella enterica subsp. enterica serotype Dublin (S. dublin), and Leptospira interrogans serovar hardjo (L. hardjo). Although most of the 95 SPF farms had a low risk on introduction of infectious diseases, one disease was introduced into 12 farms and two diseases were introduced into one farm. Three farms experienced an outbreak of BHV1, one farm an outbreak of L. hardjo, two farms BVDV, six farms S. dublin, and one farm both BHV1 and S. dublin. The total incidence rate was 0.09 (0.06-0.12) per herd-year at risk. The results suggest that the "non outbreak" farms were significantly more closed than the "outbreak" farms. Direct animal contacts with other cattle should be avoided and professional visitors should be instructed to wear protective clothing before handling cattle. PMID- 12114012 TI - Prevalence and interrelationships of hoof lesions and lameness in Swedish dairy cows. AB - The prevalence of hoof lesions and lameness in 4899 heifers and cows was determined at claw trimming one time in a cross-sectional study on 101 Swedish dairy farms, 1996-1998. The percentage of affected animals was 41% for heel-horn erosion, 30% for sole haemorrhages, 27% for erosive dermatitis, 21% for abnormal claw shape, 14% for white-line haemorrhages, 8.8% for white-line fissures, 8.6% for sole ulcers, 3.3% for double soles, 2.3% for verrucose dermatitis, and 1.8% for interdigital hyperplasia (IH). Seventy-two percent of all animals had at least one hoof lesion. The prevalence of lameness was 5.1%; most hoof lesions did not cause lameness. Differences between herds were substantial; the herd specific, animal-level prevalence of lesions ranged from 25 to 98% and of lameness from 0 to 33%. Sole haemorrhages were found in all herds. The proportion of variance at the between-herd level was particularly high for heel-horn erosion (62%) and abnormal claw shape (54%). Strong correlations between lesions were found within hooves (and animals), e.g. for heel-horn erosion and dermatitis (Spearman's rank correlation, r(s)=0.36 and 0.37, respectively) and for sole and white-line haemorrhages (r(s)=0.25 and 0.28). Most hoof lesions affected hind and front hooves bilaterally, whereas the correlation between hind and front hooves generally was lower. Herds that ranked high for prevalence of sole ulcer also ranked high for sole haemorrhages and for abnormal claw shape and herds that ranked high for dermatitis also ranked high for heel-horn erosion, verrucose dermatitis and IH. Abnormal claw shape was strongly associated with sole ulcer (r(s)=0.41 at cow level)-suggesting the importance of maintaining a correct claw shape for the prevention of hoof-horn lesions. PMID- 12114016 TI - BRICHOS: a conserved domain in proteins associated with dementia, respiratory distress and cancer. AB - A novel domain (the BRICHOS domain) of approximately 100 amino acids has been identified in several previously unrelated proteins that are linked to major diseases. These include BRI(2), which is related to familial British and Danish dementia (FBD and FDD); Chondromodulin-I (ChM-I), related to chondrosarcoma; CA11, related to stomach cancer; and surfactant protein C (SP-C), related to respiratory distress syndrome (RDS). In several of these, the conserved BRICHOS domain is located in the propeptide region that is removed after proteolytic processing. Experimental data suggest that the role of this domain could be related to the complex post-translational processing of these proteins. PMID- 12114015 TI - Wnt signal transduction: kinase cogs in a nano-machine? PMID- 12114017 TI - Understanding nuclear order. PMID- 12114022 TI - Oxidative stress shortens telomeres. AB - Telomeres in most human cells shorten with each round of DNA replication, because they lack the enzyme telomerase. This is not, however, the only determinant of the rate of loss of telomeric DNA. Oxidative damage is repaired less well in telomeric DNA than elsewhere in the chromosome, and oxidative stress accelerates telomere loss, whereas antioxidants decelerate it. I suggest here that oxidative stress is an important modulator of telomere loss and that telomere-driven replicative senescence is primarily a stress response. This might have evolved to block the growth of cells that have been exposed to a high risk of mutation. PMID- 12114023 TI - rRNA modifications and ribosome function. AB - The development of three-dimensional maps of the modified nucleotides in the ribosomes of Escherichia coli and yeast has revealed that most (approximately 95% in E. coli and 60% in yeast) occur in functionally important regions. These include the peptidyl transferase centre, the A, P and E sites of tRNA- and mRNA binding, the polypeptide exit tunnel, and sites of subunit-subunit interaction. The correlations suggest that many ribosome functions benefit from nucleotide modification. PMID- 12114024 TI - Cell cycle and death control: long live Forkheads. AB - The FOXO family of Forkhead transcription factors, FKHR (FOXO1), FKHR-L1 (FOXO3a) and AFX (FOXO4), are regulated by the phosphoinositide-3-kinase-protein-kinase-B (PI3K-PKB/c-Akt) pathway. Direct phosphorylation by PKB results in cytoplasmic retention and inactivation, inhibiting the expression of FOXO-regulated genes, which control the cell cycle, cell death, cell metabolism and oxidative stress. This pathway appears to be well conserved throughout evolution. In the nematode Caenorhabditis elegans, it affects lifespan and controls dauer formation. Recent discoveries about FOXO regulation by PI3K-PKB signalling suggest that the PI3K PKB-FOXO pathway might participate in similar processes in higher eukaryotes. PMID- 12114025 TI - Molybdenum and tungsten in biology. AB - Molybdenum is the only second-row transition metal that is required by most living organisms, and the few species that do not require molybdenum use tungsten, which lies immediately below molybdenum in the periodic table. Because of their unique chemical versatility and unusually high bioavailability these two transition metals have been incorporated into the active sites of enzymes over the course of evolution. Enzymes that contain molybdenum or tungsten continue to be discovered and several crystal structures have become available recently. This new structural information has been complemented by spectroscopic and kinetic methods, as well as computational approaches. Together, these studies provide an increasingly detailed view of the reaction mechanisms and the correlation between the electronic structure of the active site and catalytic function, one of the fundamental goals in metallobiochemistry. PMID- 12114026 TI - Protein quality control: U-box-containing E3 ubiquitin ligases join the fold. AB - Molecular chaperones act with folding co-chaperones to suppress protein aggregation and refold stress damaged proteins. However, it is not clear how slowly folding or misfolded polypeptides are targeted for proteasomal degradation. Generally, selection of proteins for degradation is mediated by E3 ubiquitin ligases of the mechanistically distinct HECT and RING domain sub-types. Recent studies suggest that the U-box protein family represents a third class of E3 enzymes. CHIP, a U-box-containing protein, is a degradatory co-chaperone of heat-shock protein 70 (Hsp70) and Hsp90 that facilitates the polyubiquitination of chaperone substrates. These data indicate a model for protein quality control in which the interaction of Hsp70 and Hsp90 with co-chaperones that have either folding or degradatory activity helps to determine the fate of non-native cellular proteins. PMID- 12114027 TI - The discovery of polyribosomes. PMID- 12114030 TI - The economics of HIV in Africa. PMID- 12114031 TI - Two decades of progress in preventing vascular disease. PMID- 12114032 TI - Evidence of success in HIV prevention in Africa. PMID- 12114033 TI - Medical marijuana--moving beyond the smoke. PMID- 12114034 TI - Face transplantation--fantasy or the future? PMID- 12114036 TI - MRC/BHF Heart Protection Study of cholesterol lowering with simvastatin in 20,536 high-risk individuals: a randomised placebo-controlled trial. AB - BACKGROUND: Throughout the usual LDL cholesterol range in Western populations, lower blood concentrations are associated with lower cardiovascular disease risk. In such populations, therefore, reducing LDL cholesterol may reduce the development of vascular disease, largely irrespective of initial cholesterol concentrations. METHODS: 20,536 UK adults (aged 40-80 years) with coronary disease, other occlusive arterial disease, or diabetes were randomly allocated to receive 40 mg simvastatin daily (average compliance: 85%) or matching placebo (average non-study statin use: 17%). Analyses are of the first occurrence of particular events, and compare all simvastatin-allocated versus all placebo allocated participants. These "intention-to-treat" comparisons assess the effects of about two-thirds (85% minus 17%) taking a statin during the scheduled 5-year treatment period, which yielded an average difference in LDL cholesterol of 1.0 mmol/L (about two-thirds of the effect of actual use of 40 mg simvastatin daily). Primary outcomes were mortality (for overall analyses) and fatal or non-fatal vascular events (for subcategory analyses), with subsidiary assessments of cancer and of other major morbidity. FINDINGS: All-cause mortality was significantly reduced (1328 [12.9%] deaths among 10,269 allocated simvastatin versus 1507 [14.7%] among 10,267 allocated placebo; p=0.0003), due to a highly significant 18% (SE 5) proportional reduction in the coronary death rate (587 [5.7%] vs 707 [6.9%]; p=0.0005), a marginally significant reduction in other vascular deaths (194 [1.9%] vs 230 [2.2%]; p=0.07), and a non-significant reduction in non vascular deaths (547 [5.3%] vs 570 [5.6%]; p=0.4). There were highly significant reductions of about one-quarter in the first event rate for non-fatal myocardial infarction or coronary death (898 [8.7%] vs 1212 [11.8%]; p<0.0001), for non fatal or fatal stroke (444 [4.3%] vs 585 [5.7%]; p<0.0001), and for coronary or non-coronary revascularisation (939 [9.1%] vs 1205 [11.7%]; p<0.0001). For the first occurrence of any of these major vascular events, there was a definite 24% (SE 3; 95% CI 19-28) reduction in the event rate (2033 [19.8%] vs 2585 [25.2%] affected individuals; p<0.0001). During the first year the reduction in major vascular events was not significant, but subsequently it was highly significant during each separate year. The proportional reduction in the event rate was similar (and significant) in each subcategory of participant studied, including: those without diagnosed coronary disease who had cerebrovascular disease, or had peripheral artery disease, or had diabetes; men and, separately, women; those aged either under or over 70 years at entry; and--most notably--even those who presented with LDL cholesterol below 3.0 mmol/L (116 mg/dL), or total cholesterol below 5.0 mmol/L (193 mg/dL). The benefits of simvastatin were additional to those of other cardioprotective treatments. The annual excess risk of myopathy with this regimen was about 0.01%. There were no significant adverse effects on cancer incidence or on hospitalisation for any other non-vascular cause. INTERPRETATION: Adding simvastatin to existing treatments safely produces substantial additional benefits for a wide range of high-risk patients, irrespective of their initial cholesterol concentrations. Allocation to 40 mg simvastatin daily reduced the rates of myocardial infarction, of stroke, and of revascularisation by about one-quarter. After making allowance for non compliance, actual use of this regimen would probably reduce these rates by about one-third. Hence, among the many types of high-risk individual studied, 5 years of simvastatin would prevent about 70-100 people per 1000 from suffering at least one of these major vascular events (and longer treatment should produce further benefit). The size of the 5-year benefit depends chiefly on such individuals' overall risk of major vascular events, rather than on their blood lipid concentrations alone. PMID- 12114037 TI - MRC/BHF Heart Protection Study of antioxidant vitamin supplementation in 20,536 high-risk individuals: a randomised placebo-controlled trial. AB - BACKGROUND: It has been suggested that increased intake of various antioxidant vitamins reduces the incidence rates of vascular disease, cancer, and other adverse outcomes. METHODS: 20,536 UK adults (aged 40-80) with coronary disease, other occlusive arterial disease, or diabetes were randomly allocated to receive antioxidant vitamin supplementation (600 mg vitamin E, 250 mg vitamin C, and 20 mg beta-carotene daily) or matching placebo. Intention-to-treat comparisons of outcome were conducted between all vitamin-allocated and all placebo-allocated participants. An average of 83% of participants in each treatment group remained compliant during the scheduled 5-year treatment period. Allocation to this vitamin regimen approximately doubled the plasma concentration of alpha tocopherol, increased that of vitamin C by one-third, and quadrupled that of beta carotene. Primary outcomes were major coronary events (for overall analyses) and fatal or non-fatal vascular events (for subcategory analyses), with subsidiary assessments of cancer and of other major morbidity. FINDINGS: There were no significant differences in all-cause mortality (1446 [14.1%] vitamin-allocated vs 1389 [13.5%] placebo-allocated), or in deaths due to vascular (878 [8.6%] vs 840 [8.2%]) or non-vascular (568 [5.5%] vs 549 [5.3%]) causes. Nor were there any significant differences in the numbers of participants having non-fatal myocardial infarction or coronary death (1063 [10.4%] vs 1047 [10.2%]), non-fatal or fatal stroke (511 [5.0%] vs 518 [5.0%]), or coronary or non-coronary revascularisation (1058 [10.3%] vs 1086 [10.6%]). For the first occurrence of any of these "major vascular events", there were no material differences either overall (2306 [22.5%] vs 2312 [22.5%]; event rate ratio 1.00 [95% CI 0.94-1.06]) or in any of the various subcategories considered. There were no significant effects on cancer incidence or on hospitalisation for any other non-vascular cause. INTERPRETATION: Among the high-risk individuals that were studied, these antioxidant vitamins appeared to be safe. But, although this regimen increased blood vitamin concentrations substantially, it did not produce any significant reductions in the 5-year mortality from, or incidence of, any type of vascular disease, cancer, or other major outcome. PMID- 12114038 TI - Photographic negative shadow of pulmonary oedema. PMID- 12114039 TI - Assessment of a pilot antiretroviral drug therapy programme in Uganda: patients' response, survival, and drug resistance. AB - BACKGROUND: Little is known about how to implement antiretroviral treatment programmes in resource-limited countries. We assessed the UNAIDS/Uganda Ministry of Health HIV Drug Access Initiative--one of the first pilot antiretroviral programmes in Africa--in which patients paid for their medications at negotiated reduced prices. METHODS: We assessed patients' clinical and laboratory information from August, 1998, to July, 2000, from three of the five accredited treatment centres in Uganda, and tested a subset of specimens for phenotypic drug resistance. FINDINGS: 912 patients presented for care at five treatment centres. We assessed the care of 476 patients at three centres, of whom 399 started antiretroviral therapy. 204 (51%) received highly active antiretroviral therapy (HAART), 189 (47%) dual nucleoside reverse transcriptase inhibitors (2NRTI), and six (2%) NRTI monotherapy. Median baseline CD4 cell counts were 73 cells/microL (IQR 15-187); viral load was 193817 copies/mL (37013-651 716). The probability of remaining alive and in care was 0.63 (95% CI 0.58-0.67) at 6 months and 0.49 (0.43-0.55) at 1 year. Patients receiving HAART had greater virological responses than those receiving 2NRTI. Cox's proportional hazards models adjusted for viral load and regimen showed that a CD4 cell count of less than 50 cells/microL (vs 50 cells/microL or more) was strongly associated with death (hazard ratio 2.93 [1.51 5.68], p=0.001). Among 82 patients with a viral load of more than 1000 copies/mL more than 90 days into therapy, phenotypic resistance to NRTIs was found for 47 (57%): 29 of 37 (78%) who never received HAART versus 18 of 45 (40%) who received HAART (p=0.0005). INTERPRETATION: This pilot programme successfully expanded access to antiretroviral drugs in Uganda. Identification and treatment of patients earlier in the course of their illness and increased use of HAART could improve probability of survival and decrease drug resistance. PMID- 12114040 TI - Declining HIV-1 incidence and associated prevalence over 10 years in a rural population in south-west Uganda: a cohort study. AB - BACKGROUND: In Uganda, there have been encouraging reports of reductions in HIV-1 prevalence but not in incidence, which is the most reliable measure of epidemic trends. We describe HIV-1 incidence and prevalence trends in a rural population based cohort between 1989 and 1999. METHODS: We surveyed the adult population of 15 neighbouring villages for HIV-1 infection using annual censuses, questionnaires, and serological surveys. We report crude annual incidence rates by calendar year and prevalence by survey round. FINDINGS: 6566 HIV-1 seronegative adults were bled two or more times between January, 1990, and December, 1999, contributing 31984 person years at risk (PYAR) and 190 seroconversions. HIV-1 incidence fell from 8.0 to 5.2 per 1000 PYAR between 1990 and 1999 (p=0.002, chi(2) for trend). Significant sex-specific and age-group specific reductions in incidence were evident. Incidence was 37% lower for 1995 99 than for 1990-94 (p=0.002, t-test). On average, 4642 adult residents had a definite HIV-1 serostatus at each yearly survey round. HIV-1 prevalence fell significantly between the first and tenth annual survey rounds (p=0.03, chi(2) for trend), especially among men aged 20-24 years (6.5% to 2.2%) and 25-29 years (15.2% to 10.9%) and women aged 13-19 years (2.8% to 0.9%) and 20-24 years (19.3% to 10.1%) (all p<0.001, chi(2) for trend). INTERPRETATION: Our findings of a significant drop in adult HIV-1 incidence in rural Ugandans give hope to AIDS control programmes elsewhere in sub-Saharan Africa where rates of HIV-1 infection remain high. PMID- 12114041 TI - Effect of T-cell peptides derived from Fel d 1 on allergic reactions and cytokine production in patients sensitive to cats: a randomised controlled trial. AB - BACKGROUND: Some patients with asthma who are allergic to cats and are injected intradermally with short, overlapping, T-cell peptides derived from Fel d 1 develop late asthmatic reactions to the peptides, which are associated with a reduction in late-phase skin reactions induced by whole allergens and bronchial hyporesponsiveness to the peptides on the second injection. We aimed to ascertain the effect of multiple injections on the magnitude of the early and late phase skin reactions to intact allergens. METHODS: After a 9-week run-in period, we randomly assigned patients with asthma and allergies to cats to receive either Fel d 1 peptides (90 microg in increasing divided doses) or placebo. The primary outcome was late-phase cutaneous reactions to whole cat dander. Outcomes were measured at baseline, 4-8 weeks, and 3-9 months. Analysis was by intention to treat. FINDINGS: 16 patients were randomly assigned to the peptides, and eight to placebo. All patients completed the course of injections. Four of the 16 patients on Fel d 1 peptides had initial late asthmatic reactions, but could be desensitised to the higher dose of peptide. Patients in the peptide group but not the placebo group had a significant reduction in the size of their late reaction to whole cat dander between baseline and both follow-ups, but the difference between groups was not significant (first follow-up, difference -422.8 mm(2) [95% CI -1115.0 to 269.4], p=0.43; second follow-up -1180.8 mm(2) [-2216.8 to -144.8], p=0.058). The size of the late reaction to Fel d 1 significantly differed between treatment groups at both follow-ups. At second follow-up, the size of the early reaction to Fel D 1, but not to whole cat dander was significantly reduced in those on peptides compared with those on placebo. The concentration of interferon gamma and of interleukin 4 and 13, and the amount of proliferation, significantly decreased between baseline and second follow-up, and the concentration of interleukin 10 was significantly higher in patients on peptides, however, none of these values differed significantly between groups. Patients on peptides had a significantly greater decrease in the concentration of interferon gamma and interleukin 13, and in the amount of proliferation between baseline and first follow-up than did those on placebo. INTERPRETATION: Several, short, overlapping Fel d 1 T-cell peptides have potential in treatment of cat allergy. PMID- 12114042 TI - Treatment of anisakiasis with albendazole. PMID- 12114043 TI - How practical are recommendations for dietary control in phenylketonuria? AB - In patients with phenylketonuria, blood phenylalanine concentration during childhood is the major determinant of cognitive outcome. Guidelines provide age related recommendations for treatment. To ascertain the extent to which these aims are achievable, we audited results from four centres for the years 1994 2000. The median proportion of samples with phenylalanine concentrations above those recommended was less than 30% for those younger than age 10 years but almost 80% for those aged 15 years and older. Similarly, the median frequency of blood sampling, expressed as a proportion of that recommended, was more than 80% for patients younger than 10 years but less than 50% by age 15 years. Our results indicate the difficulty of maintaining control in phenylketonuria, especially in older rather than younger children. PMID- 12114044 TI - Adiponectin and development of type 2 diabetes in the Pima Indian population. AB - Adiponectin is a collagen-like circulating protein secreted by adipocytes that is proposed to mediate obesity-related resistance to insulin. In a case-control series, we assessed the role of adiponectin in later development of type 2 diabetes in 70 patients who later developed type 2 diabetes and 70 controls, matched for body-mass index, age, and sex. Cases and controls were taken from the longitudinal study of health in the Pima Indian population. At baseline, the concentration of adiponectin was lower in cases than in controls (p=0.01) and individuals with high concentrations of this protein were less likely to develop type 2 diabetes than those with low concentrations (incidence rate ratio 0.63 [95% CI 0.43-0.92]; p=0.02). PMID- 12114045 TI - Chloroquine-resistant Plasmodium malariae in south Sumatra, Indonesia. AB - Oral chloroquine is the treatment of choice for uncomplicated Plasmodium malariae infections worldwide. We did a prospective 28-day in-vivo assessment of the efficacy of chloroquine for treatment of P malariae on Legundi Island in Lampung Bay, Sumatra, Indonesia. Of 28 patients, one had recurrent parasitaemia on day 28, and two had persistent parasitaemia to day 8. Whole-blood chloroquine and desethylchloroquine concentrations were at ordinarily effective levels (> or = 100 microg/L) on day 8 in both cases of persistent parasitaemia. These findings suggest that clinical resistance to chloroquine by P malariae occurs in the Indonesian archipelago of southeast Asia. PMID- 12114046 TI - Low profile for health at Kananaskis G8 summit. PMID- 12114050 TI - USPSTF releases guidelines on breast-cancer chemoprevention. PMID- 12114052 TI - WHO urges more research into acrylamide in food. PMID- 12114053 TI - UN study expresses concern over national antiabortion policies. PMID- 12114056 TI - Tanzania rail crash highlights lack of medical resources for disasters. PMID- 12114057 TI - Uganda hopes Fund will bring long-term progress. PMID- 12114058 TI - Shadow on the continent: public health and HIV/AIDS in Africa in the 21st century. AB - Approaches to the prevention and control of the HIV/AIDS epidemic in Africa have been heavily based on early experiences and policies from industrialised countries, where the disease affects specific risk groups. HIV/AIDS has been dealt with differently from other sexually transmitted or lethal infectious diseases, despite being Africa's leading cause of death. In this review, we discuss the evolution of the global response to the epidemic, and the importance of redefining HIV/AIDS in Africa as a public health and infectious disease emergency. We discuss reconsideration of policies and practice around HIV testing and partner notification, and emphasise the need for an increased focus on treatment. Human-rights based approaches to HIV/AIDS prevention might have reduced the role of public health and social justice, which offer a more applied and practical framework for HIV/AIDS prevention and care in Africa's devastating epidemic. PMID- 12114059 TI - Always something new from Africa. PMID- 12114060 TI - Can we reverse the HIV/AIDS pandemic with an expanded response? AB - HIV/AIDS has reached pandemic proportions, and is one of the leading causes of death worldwide. In 2001, the Declaration of Commitment on HIV/AIDS set out several aims with respect to reducing the effect and spread of HIV/AIDS, and an expanded response in low-income and middle-income countries was initiated. Here we examine the potential effect of the expanded global response based on analyses of epidemiological data, of mathematical models of HIV-1 transmission, and a review of the impact of prevention interventions on risk behaviours. Analyses suggest that if the successes achieved in some countries in prevention of transmission can be expanded to a global scale by 2005, about 29 million new infections could be prevented by 2010. PMID- 12114061 TI - The Ugandan success story? Evidence and claims of HIV-1 prevention. PMID- 12114062 TI - Improvement of the study, analysis, and reporting of adverse events associated with antiretroviral therapy. PMID- 12114063 TI - HIV/AIDS prevention and treatment. PMID- 12114064 TI - HIV/AIDS prevention and treatment. PMID- 12114065 TI - HIV/AIDS prevention and treatment. PMID- 12114068 TI - HIV/AIDS prevention and treatment. PMID- 12114069 TI - Who is fooling us? PMID- 12114070 TI - Non-steroidal anti-inflammatory drugs and risk of serious coronary heart disease. PMID- 12114071 TI - Non-steroidal anti-inflammatory drugs and risk of serious coronary heart disease. PMID- 12114073 TI - Human rights for people registering as organ donors. PMID- 12114074 TI - Data for older adult populations in sub-Saharan Africa. PMID- 12114075 TI - Mobilisation of Cuban doctors in developing countries. PMID- 12114076 TI - The battle of the sexes in sports. PMID- 12114090 TI - Myocardial contrast echocardiography in acute myocardial infarction: room for improvement in performance and interpretation. PMID- 12114091 TI - Weak concordance between wall motion and microvasculature status after acute myocardial infarction: study with myocardial contrast echocardiography in real time with power modulation. AB - AIMS: The microvasculature damage after myocardial infarction has important implications. The hypothesis of the study was that wall motion abnormalities and microcirculation status do not necessarily match after myocardial infarction, and therefore the study of only myocardial wall motion could offer an incomplete evaluation in these patients. METHODS: Wall motion and myocardial perfusion assessed by contrast echocardiography were evaluated by two different blinded investigators in 29 patients with recent (<1 week) myocardial infarction. Myocardial perfusion was assessed in real-time using power modulation after Optison (1.5-3.0 ml) intravenous administration. RESULTS: One hundred and ninety nine segments could be adequately evaluated. Of these, 54 (27%) were akinetic. Regarding contrast opacification, 134 segments (67%) had a normal perfusion, whereas the remaining 65 (33%) had an impaired (n=37, 19%) or absent (n= 28, 14.1%) perfusion. Concordance between presence of akinesia and abnormal contrast opacification was only moderate (kappa index 0.42) and agreement only occurred in 116 segments (58%). Fourteen per cent of normoquinetic segments had an impaired perfusion, whereas 35% of akinetic segments had a preserved perfusion. Correlation between the proportion of segments with akinesia and the proportion of segments with impaired perfusion was moderate (r=0.41), and there was no correlation between the proportion of segments with akinesia and the percentage of segments with absent perfusion. CONCLUSION: There is a weak association between regional systolic function and myocardial perfusion after myocardial infarction, as assessed by real-time contrast myocardial echocardiography using power modulation. PMID- 12114092 TI - Real time contrast echocardiography--a new bedside technique to predict contractile reserve early after acute myocardial infarction. AB - AIMS: Power pulse inversion echocardiography is a new technique by which contrast microbubbles can be visualised in real time within the myocardium, enabling simultaneous assessment of myocardial function and microvascular integrity, which is a prerequisite for myocardial viability. We aimed to determine whether microvascular integrity using power pulse inversion can be used to predict contractile reserve early after myocardial infarction. METHODS AND RESULTS: We studied 19 stable patients 5.1(1.6) days after presentation using low dose dobutamine stress echocardiography and power pulse inversion using slow bolus intravenous injections of Optison. A 16-segment left ventricular model was used to define wall thickening at baseline and following low dose dobutamine infusion (1, normal; 2, reduced; 3, absent), and contrast opacification (1, homogeneous; 2, heterogenous or reduced; 3, absent). The techniques were compared on a segment by-segment basis to determine whether microvascular integrity (contrast opacification score of 1 or 2) could predict contractile reserve (any improvement during low dose dobutamine infusion) in segments that were akinetic at rest. Follow-up echocardiography was performed one month later. RESULTS: Ninety-four (31%) of the 304 segments were akinetic at rest, and 22 (23%) of these demonstrated contractile reserve. In 87 (92%) of the resting akinetic segments contrast opacification could be adequately determined, and of these 20 (23%) showed microvascular integrity. The negative and positive predictive value of microvascular integrity for determining contractile reserve was 90% and 65%, respectively, and 92% and 59% respectively for predicting recovery of function. CONCLUSION: Power pulse inversion can be used at rest to determine myocardial function and simultaneously to predict contractile reserve of akinetic segments in patients early after myocardial infarction. This technique has the potential to provide a bedside assessment of myocardial viability. PMID- 12114093 TI - Perforated aneurysms of left side valves during active infective endocarditis complicating hypertrophic obstructive cardiomyopathy. AB - The most frequent site of vegetative lesion in patients with hypertrophic cardiomyopathy is anterior mitral leaflet, due to chronic endocardial trauma arising from systolic anterior motion. We describe three cases of serious infective endocarditis complicated lesions (vegetation, aneurysm and perforation) on aortic and mitral valves, in patients with obstructive hypertrophic cardiomyopathy. In particular, we observed how severe valvular damage and dysfunction, combined with particular hemodynamic conditions, are followed by adverse clinical outcome. We performed transthoracic echocardiogram and transoesophageal echocardiography studies to define morphologic and hemodynamic features of infection, deciding the proper therapy and we planned the echocardiographic follow-up. PMID- 12114094 TI - Left ventricular diastolic dysfunction in congenital chronic anaemias during childhood as determined by comprehensive echocardiographic imaging including acoustic quantification. AB - AIMS: To evaluate prospectively the left ventricular performance in thalassaemia major and sickle cell disease using comprehensive echocardiographic imaging including acoustic quantification during early childhood. METHODS AND RESULTS: Twenty-three patients with thalassaemia and 26 patients with sickle cell disease underwent echocardiographic examination including M-mode, 2-D, Doppler and acoustic quantification. All patients were matched for age, sex, weight and height with 20 normal controls. All patients were below 13 years of age. Thalassaemia and sickle cell disease patients were significantly anaemic when compared with normals (P<0.0001). All patients had normal left ventricular systolic parameters. Acoustic quantification-derived left ventricular volumes, filling rates, and emptying rates were not different in thalassaemia patients from controls. Left ventricular volumes, however, were larger in sickle cell disease patients than in controls. In contrast, by Doppler technique, left ventricular filling occurs mainly in early diastole (E wave) in thalassaemia patients and mainly in late diastole (A wave) in sickle cell disease patients, (P=0.03 and 0.01 respectively). E/A ratio was lower and diastolic filling period was shorter than normal in sickle cell disease but not in thalassaemia patients. Patients in both groups had left ventricular mass (determined by M-mode) significantly higher than normal (P<0.0001). CONCLUSION: The left ventricular systolic performance is well preserved in patients with chronic anaemia due to thalassaemia major and sickle cell disease during early childhood. In both diseases, however, there is left ventricular hypertrophy and measurable abnormalities in the diastolic filling detected by Doppler. Such changes do not fit a specific cardiomyopathic pattern due to diastolic dysfunction i.e. restrictive physiology vs delayed relaxation. Acoustic quantification of left ventricular diastolic parameters (filling rates) was less sensitive than Doppler in detecting these early diastolic abnormalities in both diseases. PMID- 12114095 TI - Usefulness of contrast agents in the diagnosis of left ventricular pseudoaneurysm after acute myocardial infarction. AB - BACKGROUND AND OBJECTIVE: The diagnosis of left ventricular pseudoaneurysm after acute myocardial infarction is usually based on echocardiography. However, this technique may have limitations in some patients, especially in cases with suboptimal acoustic window. The objective of this study was to evaluate the usefulness of contrast echocardiography in the diagnosis of left ventricular pseudoaneurysm after myocardial infarction. METHODS AND RESULTS: The study population comprises six patients in whom a two-dimensional echocardiography showed an image consistent with left ventricular pseudoaneurysm. Levovist (Schering) 4gr was administered i.v. to more clearly visualize the blood flow from the left ventricle to the left ventricular pseudoaneurysm cavity in all patients. Infarct location was anterior in five patients, and posterolateral in one. No patient had received thrombolysis or primary angioplasty during the acute phase. The transthoracic echocardiographic study showed an echo-free space adjacent to left ventricle in all patients. In four cases, the diagnosis of left ventricular pseudoaneurysm was made before contrast administration. In the remaining two patients, the definite diagnosis was made only after Levovist administration. CONCLUSION: In the diagnosis of postinfarction left ventricular pseudoaneurysm, the administration of contrast agents may be of help in the correct visualization of the blood flow from the left ventricle to the left ventricular pseudoaneurysm cavity, and may allow a definite diagnosis to be obtained in some patients. PMID- 12114097 TI - Mitral inflow and mitral annular motion velocities in patients with mitral annular calcification: evaluation by pulsed Doppler echocardiography and pulsed Doppler tissue imaging. AB - AIMS: We evaluated the relationship between the mitral inflow velocities by pulsed Doppler echocardiography and mitral annular motion velocities by pulsed Doppler tissue imaging in patients with mitral annular calcification. METHODS AND RESULTS: Fifty-three patients with mitral annular calcification were divided into two groups: severe mitral annular calcification (n=15, mitral annular calcification bigger than or equal 5mm in width) and mild mitral annular calcification (n=38, mitral annular calcification <5mm in width). In addition, 20 patients with hypertensive heart disease (HHD group) and mild left ventricular hypertrophy but no mitral annular calcification and 30 normal individuals (normal group) were studied. The early diastolic mitral inflow velocity (E) was higher in the severe mitral annular calcification group (0.75+/-0.26 m/s) than in the HHD and normal groups (mild mitral annular calcification, 0.65+/-0.21; HHD, 0.57+/ 0.24; normal, 0.55+/-0.15m/s), and the late diastolic mitral inflow velocity (A) was higher in the severe mitral annular calcification group (1.24+/-0.23 m/s) than in the other three groups (mild mitral annular calcification, 0.96+/-0.20; HHD, 0.84+/-0.23; normal, 0.75+/-0.13 m/s). In contrast, the early and late diastolic annular velocities (Ea, Aa) were lower in the severe mitral annular calcification group (Ea: 5.7+/-2.2; Aa: 11.9+/-4.4 cm/s) than in the other three groups (Ea: mild mitral annular calcification, 8.3+/-2.5; HHD, 7.7+/-2.2; normal, 9.0+/-1.8 cm/s; Aa: mild mitral annular calcification, 14.2+/-4.1; HHD, 14.3+/ 2.8; normal, 14.2+/-2.1cm/s). Mitral valve area was smaller in the severe mitral annular calcification group (2.6+/-1.0 cm(2)) than in the other three groups (mild mitral annular calcification, 3.1+/-0.7; HHD, 4.1+/-0.7; normal, 4.2+/-0.9 cm(2)). In the mitral annular calcification and normal groups, the A correlated inversely with mitral valve area (r=-0.67, P<0.01) and directly with severity of mitral annular calcification (r=0.65, P<0.01), and the Ea correlated inversely with left ventricular wall thickness (r=-0.37, P<0.01) and severity of mitral annular calcification (r=-0.45, P<0.01). CONCLUSION: Patients with severe mitral annular calcification have higher mitral inflow velocities due to mitral annular restriction and lower mitral annular velocities caused by decreased mitral annular motion and abnormal left ventricular relaxation. PMID- 12114096 TI - Coronary microcirculation in essential hypertension: a quantitative myocardial contrast echocardiographic approach. AB - AIMS: The aims of the present study were: (a) to demonstrate whether quantitative myocardial contrast echocardiography can detect the increase in coronary flow induced by dipyridamole infusion vasodilation through the myocardial opacification due to the transit of microbubbles, both at rest and after dipyridamole induced vasodilation; (b) to explore the coronary microcirculatory function before and after dipyridamole in two different models: asymptomatic and relatively young hypertensive patients with a mild degree of left ventricular hypertrophy, and healthy controls. METHODS AND RESULTS: Two groups of strictly age-matched males were studied (case-control study): 10, relatively young and asymptomatic essential hypertensive patients with a mild degree of left ventricular hypertrophy with a normal left ventricular function, and 10 healthy controls. The main findings were: the microbubbles' appearance area was significantly lower in hypertensive patients than in controls (P<0.05) because of a significantly lower time to peak. The peak intensity at rest was higher in hypertensives than in controls (P<0.05); but the per cent increase after vasodilatory stimulus was significantly higher in controls (+71% in controls vs +31% in hypertensives; P<0.05). The microbubbles' disappearance area was comparable in both groups at rest; the per cent increase of this parameter after dipyridamole was significantly higher in controls (+124%) than in hypertensives (+90%) (P<0.05). The results achieved in this study documented that the coronary microcirculation in hypertensive patients presenting a mild degree of left ventricular hypertrophy, explored with quantitative myocardial contrast echocardiography, showed a different behaviour in comparison with controls, in the vasodilatory response to dipyridamole. CONCLUSION: The coronary microcirculation in hypertensives showed a reduced vasodilation capacity of the resistance arterioles under dipyridamole induced vasodilatation, and a possible impairment of the endothelium dependent vasodilation. This happened despite an increase in the left ventricular mass, where the relation between capillary bed distribution and hypertrophied myocardium (rarefaction phenomenon) is not completely respected. PMID- 12114098 TI - Right ventricular diastolic dysfunction in arterial systemic hypertension: analysis by pulsed tissue Doppler. AB - AIMS: This study analyses right ventricular longitudinal function in arterial systemic hypertension by pulsed tissue Doppler. METHODS AND RESULTS: Thirty normotensives and 30 hypertensives, free of cardiac drugs, underwent standard Doppler echocardiography and pulsed tissue Doppler of right ventricular lateral tricuspid annulus and left ventricular lateral mitral annulus. By tissue Doppler, systolic and diastolic measurements were obtained. Hypertensives had higher left ventricular mass and impaired Doppler diastolic indexes, without changes of global systolic function. Tissue Doppler showed reduction of right ventricular E/A ratio and prolongation of relaxation time in comparison with controls (both P<0.00001). In the overall population, the length of tissue Doppler derived right ventricular relaxation time was positively related to right ventricular anterior wall thickness while right ventricular E/A ratio was positively related to E/A ratio of left ventricular mitral annulus (both P<0.00001). These relations remained significant even after adjusting for clinical and echocardiographic confounders by separate multivariate models. CONCLUSIONS: Arterial systemic hypertension is associated to right ventricular longitudinal diastolic dysfunction. This dysfunction involves the prolongation of active relaxation, which is independently associated with the degree of right ventricular hypertrophy and the impairment of passive wall properties, which is mainly due to ventricular interaction occurring under left ventricular pressure overload conditions. PMID- 12114099 TI - Doppler echocardiographic evaluation of right ventricular diastolic function in hypertrophic cardiomyopathy. AB - AIMS: Left ventricular diastolic function in patients with hypertrophic cardiomyopathy has been adequately studied. In contrast there are few studies concerning right ventricular diastolic function in hypertrophic cardiomyopathy. We studied right ventricular diastolic function in patients with hypertrophic cardiomyopathy using Doppler echocardiography. METHODS AND RESULTS: We studied 20 patients with hypertrophic cardiomyopathy (mean age 43.6+/-13.8 years) and 20 healthy volunteers (control group, mean age 43+/-13.8 years). We calculated left ventricular and right ventricular diastolic indices using pulsed Doppler echocardiography. Hypertrophic cardiomyopathy patients compared with controls had significantly lower right ventricular-E/A ratio (1.01+/-0.40 vs 1.30+/-0.28, P<0.04), significantly prolonged right ventricular isovolumic relaxation time (170+/-72 vs 32+/-23 ms, P<0.001), and also significantly prolonged right ventricular deceleration time (160+/-58 vs 118+/-35 ms, P<0.01). There was also strong significant correlation between right ventricular deceleration time and left ventricular deceleration time (r=0.78), right ventricular-E/A ratio and left atrial filling fraction (r=-0.55) and between right atrial filling fraction and left atrial filling fraction (r=0.75). CONCLUSIONS: Right ventricular diastolic function in patients with hypertrophic cardiomyopathy is impaired, reflecting abnormal relaxation. Right ventricular diastolic indices correlate well with those of left ventricle. PMID- 12114100 TI - Lipomatus hypertrophy of the atrial septum and prominent crista terminalis appearing as a right atrial mass. AB - In these case reports, transthoracic echocardiography suggested the presence of a right atrial mass. However, subsequent transoesophageal echocardiography revealed that the 'right atrial mass' was actually a lipomatous hypertrophied atrial septum in combination with a prominent crista terminalis. An understanding of the anatomy and the echocardiographic appearance of a lipomatous hypertrophied atrial septum appearing with a prominent crista terminalis will minimize the misdiagnosis of these structures. PMID- 12114102 TI - Interventricular septal tumour. PMID- 12114101 TI - Cor triatriatum sinistrum and persistent left superior vena cava: an original association. AB - Cor triatriatum sinistrum is a rare congenital heart disease usually diagnosed in symptomatic children. Symptoms depend on the degree of obstruction to pulmonary venous return with pulmonary hypertension and other associated abnormalities. Persistent left superior vena cava is quite a common congenital heart disease (about 0.5% in healthy populations). It should be suspected every time a dilated coronary sinus is detected at the echo examination. Transthoracic and transoesophageal examinations visualize the site and the size of the fibrous membrane as well as the degree of obstruction, and allow the evaluation of pulmonary pressures that are very important clues for prognosis and therapy. This case report describes the clinical signs and the diagnostic ultrasound findings evaluated in comparison with magnetic resonance imaging, a well-defined gold standard in heart disease of this uncommon congenital association. PMID- 12114103 TI - Metastatic malignant melanoma presenting as congestive heart failure: diagnosis by transoesophageal echocardiography. PMID- 12114104 TI - Weakness of a giant: mutations of the sarcomeric protein titin. AB - In 2002, three reports described for the first time mutations in the sarcomeric protein titin associated with dilated cardiomyopathy in humans. Despite different locations (Z-line region, Z-I transitional zone, N2B region, half A band region) all mutations resulted in heart failure. In addition, an N2B mutation was found in zebrafish embryos with ventricular dilatation and cardiac insufficiency. It is concluded that titin mutations have significant functional consequences and need to be studied intensively in the future. PMID- 12114105 TI - Tales of the cryptic: unveiling more angiogenesis inhibitors. AB - Over the past few years, there has been a dramatic increase in the identification of anti-angiogenic fragments from larger proteins with unrelated functions. These cryptic anti-angiogenic molecules are largely derived from matrix components such as collagen and fibronectin, as well as from circulating proteins and some intracellular proteins. Here, we discuss the relevance of these developments in terms of their physiological roles and possible therapeutic applications. PMID- 12114106 TI - Focus on cancer prevention. AB - Tumor Prevention and Genetics 2002, the 2nd International Conference and 7th Annual Meeting of The International Society of Cancer Chemoprevention (ISCaC) was held in St. Gallen, Switzerland, 14-16 February 2002. PMID- 12114110 TI - Dual action of glatiramer acetate (Cop-1) in the treatment of CNS autoimmune and neurodegenerative disorders. AB - Protective autoimmunity is the body's defense mechanism against destructive self compounds such as those commonly associated with neurodegenerative disorders. Autoimmune disease and neurodegenerative disorders can thus be viewed as two extreme manifestations of the same process. Therefore, when designing therapy, it is important to avoid an approach that will cure the one by invoking the other. One way to stop, or at least slow down, the progression of neurodegeneration without risking development of an autoimmune disease is by boosting protective autoimmunity in a well-controlled way. Copolymer 1 (Cop-1), an approved drug for the treatment of multiple sclerosis, can be used as a treatment for autoimmune diseases and as a therapeutic vaccine for neurodegenerative diseases. We propose that the protective effect of Cop-1 vaccination is obtained through a well controlled inflammatory reaction, and that the activity of Cop-1 in driving this reaction derives from its ability to serve as a 'universal antigen' by weakly activating a wide spectrum of self-reactive T cells. PMID- 12114112 TI - PEDF: anti-angiogenic guardian of ocular function. AB - Sight-threatening eye diseases can be caused and exacerbated by the aberrant growth of new blood vessels. Recent work indicates that this neovascularization not only is a response to a rise in the local concentration of molecules that induce such angiogenesis but also requires a fall in the levels of endogenous molecules that inhibit angiogenesis. One of the most potent of these endogenous inhibitors is pigment epithelium-derived factor (PEDF), which serves as a survival factor for neuronal components of the eye as well as an essential inhibitor of the growth of ocular blood vessels. Its anti-angiogenic activity is selective in that it is effective against newly forming vessels but spares existing ones, and it is reversible. The molecular basis for this delicate control of endothelial cells is beginning to be understood and strategies to test the ability of PEDF to ameliorate or prevent vessel damage in the eye are developing rapidly. PMID- 12114111 TI - Edible plant vaccines: applications for prophylactic and therapeutic molecular medicine. AB - The use of edible plants for the production and delivery of vaccine proteins could provide an economical alternative to fermentation systems. Genes encoding bacterial and viral antigens are faithfully expressed in edible tissues to form immunogenic proteins. Studies in animals and humans have shown that ingestion of transgenic plants containing vaccine proteins causes production of antigen specific antibodies in serum and mucosal secretions. In general, the technology is limited by low expression levels for nuclear-integrated transgenes, but recent progress in plant organelle transformation shows promise for enhanced expression. The stability and immunogenicity of orally delivered antigens vary greatly, which necessitates further study on protein engineering to enhance mucosal delivery. These issues are discussed with regard to the further development of plant-based vaccine technology. PMID- 12114113 TI - Cell replacement therapy for type 1 diabetes. AB - Replacement of the insulin-producing pancreatic islet beta cells represents the ultimate treatment for type 1 diabetes. Recent advances in islet transplantation underscore the urgent need for developing alternatives to human tissue donors, which are scarce. Two possible approaches are the expansion of differentiated beta cells by reversible immortalization and the generation of insulin-producing cells from embryonic or adult stem cells. It is possible that new insights into endocrine pancreas development will ultimately lead to manipulation of progenitor cell fate towards the beta-cell phenotype of insulin production, storage and regulated secretion. Both allogeneic and autologous surrogate beta cells are likely to require protection from recurring autoimmunity. This protection might take the form of tolerization, cell encapsulation, or cell engineering with immunoprotective genes. If successful, these approaches could lead to widespread cell replacement therapy for type 1 diabetes. PMID- 12114114 TI - Transcription factors in bone: developmental and pathological aspects. AB - The skeleton in vertebrates is composed of bone and cartilage, which contains three specific types: osteoblasts and osteoclasts in bone and chondrocytes in cartilage. Like other cell types in the body, skeletal cell differentiation is controlled by multiple transcription factors at various stages of their development. Cbfa1 and Osx, a newly identified zinc-finger containing protein, are osteoblast-specific transcription factors. Loss of function of either one of them leads to absence of bone in mammals. Here, we discuss transcription factors involved in controlling the differentiation of osteoclasts, such as Pu.1 and nuclear factor (NF)-kappaB, and chondrocytes, such as Sox proteins. Finally, recent progress in identifying mutations in transcription factors affecting skeletal patterning and development is also described. PMID- 12114115 TI - Mouse models for human DNA mismatch-repair gene defects. AB - The mammalian DNA mismatch-repair genes belong to a family of genes that comprise several homologs of the Escherichia coli mutS and mutL genes. The observation that mutations in the two human repair genes MSH2 and MLH1 are responsible for hereditary nonpolyposis colorectal cancer, as well as a significant number of sporadic colorectal cancers, raises several questions about the role of these proteins and their family members in the initiation and progression of colorectal cancer. To address these questions, mice with inactivating mutations in all the known mutS and mutL homologs have been generated. The development of these mouse lines has permitted the systematic analysis of the role of each gene in the repair process and has underscored their significance in mutation avoidance and cancer susceptibility. These analyses were critical for our understanding of the function of these genes at the organismal level and also revealed an essential role for some of the DNA mismatch-repair genes in mammalian meiosis. PMID- 12114116 TI - Watching real-time metastasis in vivo. PMID- 12114117 TI - Blood-bank testing for infectious diseases: how safe is blood transfusion? AB - Remarkable progress has been made in transfusion safety from infection over the past three decades. Donor deferrals for at-risk behaviors, the introduction of more-sensitive viral-screening assays and the recent introduction of nucleic-acid amplification technology have nearly eliminated transmission of HIV and hepatitis C virus (HCV) by blood transfusion in North America. Nevertheless, risks of other infectious agents for which such robust screening tools have not been developed, such as bacteria and parasites, still remain. As a result of these successes, the non-infectious risks such as misidentification of patients and inadequate and inappropriate transfusion have become the primary sources of transfusion risk. PMID- 12114120 TI - Feasibility of off-pump ASD closure using real-time 3-D echocardiography. AB - BACKGROUND: If surgeons could "see " through blood and cardiac chamber walls, it would ultimately be unnecessary to open the heart or use cardiopulmonary bypass to perform procedures such as atrial septal defect (ASD) closure or mitral valve repair. Conventional echocardiography generates cross-sectional images that are not satisfactory as the only visualization for surgical procedures, and current 3 Dimensional (3-D) CT and echo systems take several minutes to compose and process a single still frame. Recently, however, the first system for real-time 3-D echocardiography has been developed. This study examines whether real-time 3-D echocardiography can provide images of sufficient anatomic definition, depth perception, and image resolution to substitute for optical visualization in performing ASD closures. METHODS: A prototype Volumetrics 3-D echocardiographic system was evaluated in a water bath on a complex-surfaced standard reference model to determine the image resolution and define the ideal imaging parameters. A static image and views of sutures being placed with an endoscopic needle driver and two commercial suture placement devices were evaluated at multiple angles and distances from the target. The resulting images were graded by a blinded reviewer. Once the best imaging parameters were determined, five porcine ASDs were closed with interrupted sutures, running sutures, or a pursestring suture using only echo visualization. RESULTS: The highest quality images were obtained with the probe at a distance of 4-6 cm and at angles perpendicular or 45 degrees forward to the target. Spatial and temporal resolutions were adequate to suture all ASDs closed under only echo guidance. CONCLUSIONS: The evaluated real-time 3 D echo system provided adequate spatial and temporal information to act as a guide for surgical procedures. PMID- 12114121 TI - Epicardial ablation on the beating heart: progress towards an off-pump maze procedure. AB - BACKGROUND: The treatment of atrial fibrillation with unipolar radiofrequency (RF) catheter ablation techniques has been fraught with difficulty. This study was designed to evaluate the potential advantages of bipolar RF energy and its ability to create transmural linear lesions on the beating heart. METHODS: A right thoracotomy was performed on eight adult sheep. A bipolar RF device was inserted and targeted tissue was clamped between the instrument arms. Ablation was performed on the beating heart. Energy was delivered until the tissue conductance between the two electrodes became constant. RESULTS: A total of 30 lesions were performed. Average ablation time for all lesions was 9.2+/-3.9 seconds with a mean peak temperature of 48.7+/-5.8 OC. Pacing studies demonstrated 100%(30/30) isolation and staining showed that all lesions were transmural. CONCLUSIONS: Epicardially delivered bipolar RF energy was able to reproducibly isolate atrial myocardium. As opposed to other energy sources, lesions were always transmural and can be created within seconds. On-line measurement of conductance correlates with lesion transmurality. This technology has the potential to perform a beating heart MAZE procedure. PMID- 12114122 TI - Thoracic epidural anesthesia for cardiac surgery via left anterior thoracotomy in the conscious patient. AB - BACKGROUND: Cardiac surgery is perceived to be maximally invasive and fraught with complications. In response to this perception, cardiothoracic surgeons have been refining traditional techniques to minimize their invasive nature. Epidural anesthesia has been used safely and effectively for numerous surgical procedures to reduce morbidity associated with general anesthesia. In hopes of achieving a similar result, we set out to determine the feasibility of using thoracic epidural anesthesia for limited cardiac surgery through a left anterior thoracotomy for patients who were awake and spontaneously breathing. METHODS: A high thoracic epidural technique was used in all cases. In each instance, the chest was entered through a small left anterior thoracotomy. The procedures included minimally invasive direct coronary artery bypass (MIDCAB) and transmyocardial revascularization (TMR). These procedures were performed in routine fashion using standard techniques. Pulmonary function tests were performed preoperatively, and the adequacy of respiratory function was serially monitored throughout each operation. The epidural catheters were left in place for 24 hours after operation for pain control. RESULTS: A total of 10 operations were performed. These included 7 MIDCAB, 2 TMR and 1 MIDCAB/TMR hybrid. The mean preoperative forced expiratory volume for one second (FEV1) was 1.9 liters. Significant intra-operative hypoxia or hypercarbia was not seen. One patient required intubation during the procedure for restlessness not associated with hypoxia. Two others required brief periods of assisted ventilation. All procedures were completed without incident. The mean operating time and length of stay were 70 minutes and 4.7 days. Postoperative pain control and patient satisfaction were excellent. CONCLUSIONS: Thoracic epidural anesthesia for limited cardiac surgical procedures by means of a left anterior thoracotomy is feasible, even in patients with diminished pulmonary function. Furthermore, this method offered no significant technical hurdles. Nevertheless, the applicability of this technique to other procedures remains unclear. We believe that these results warrant controlled comparison of regional versus general anesthesia for limited cardiac surgery. PMID- 12114123 TI - Clinical outcomes in coronary artery bypass graft surgery: comparison of off-pump and on-pump techniques. AB - BACKGROUND: A consecutive series of patients undergoing first-time coronary artery bypass graft (CABG) surgery were analyzed and the impact of off-pump surgery was evaluated. METHODS: From January 1, 2000 to December 31, 2000, 367 patients underwent isolated first-time CABG surgery. One hundred and twenty underwent off-pump CABG (Group A, 32.7%) and 247 underwent conventional on-pump CABG (Group B, 67.3%). Five patients were converted during operation and were included in Group A. The pre-operative characteristics, intra-operative details, and post-operative course were analyzed in the two groups. All patients were followed up between 11 and 23 months (median 18 months) after operation by telephone interviews or questionnaire survey. RESULTS: Early mortality was 2.1% (group A, 0.83%; group B, 2.83%), with the difference not being statistically significant. The incidence of post-op stroke (group A, 1.66%; group B, 3.66%), renal failure (group A, 2.5%; group B, 5.66%), and gastrointestinal complications (group A, 1.66%; group B, 1.21%) was likewise not significantly different in the two groups. However, the patients in group A had a statistically significant lower incidence of low cardiac-output (group A 13.3%; group B 29.5%; p = 0.002), atrial fibrillation (group A 11.66%; group B 30.36%; p<0.001), blood product transfusion (group A 39.66%; group B 89.87%; p<0.001), time on ventilator (group A, 5.96 hrs; group B, 10.31 hrs; p<0.001), and post-op hospital stay (group A, 7.79 days; group B, 9.81 days, p<0.001). Medium-term results (recurrence of angina, late mortality, cardiovascular events, and need for revascularization) were similar in the two groups. CONCLUSIONS: Off-pump CABG results in a decreased incidence of complications in the immediate post-op period with comparable results in the medium term. PMID- 12114124 TI - Multiple off-pump coronary revascularization with "aorta no-touch " technique using composite and sequential methods. AB - BACKGROUND: Although off-pump coronary artery bypass grafting (OPCAB) has been widely applied in patients who are considered high risk for cardiopulmonary bypass (CPB), there is still a risk of stroke during the operation because of the ascending aortic partial clamp for proximal anastomosis. In the present study, we report the initial results of an "aorta no-touch " technique using an in-situ graft and composite and sequential grafting methods. METHODS: Between March 2000 and April 2001, 120 patients underwent OPCAB with this technique. The age of patients ranged from 47 to 86 years, with a mean age of 65.7 +/- 8.7 years. On average, 3.12 +/- 0.77 grafts per patient were completed. More than 4 distal anastomoses were performed in 32 patients (27%). As in-situ grafts, 140 internal thoracic arteries (ITAs) and 9 gastroepiploic arteries were used. The radial artery (RA) was used as a Y composite graft in 91 patients, as an I composite graft in 25, and as a K composite graft in 2. Sequential bypass grafting was performed using the RA in 60 patients, the saphenous vein (SV) in 6, the ITA in 4, and the gastroepiploic artery (GEA) in 3. Arterial grafts were used in 92% (345/374) of total bypass grafts. Distal anastomosed sites were 119 left anterior descending arteries (LADs) (32%), 90 posterolateral branches (24%), 64 posterior descending arteries (17%), 49 diagonal branches (13%), 39 obtuse marginal branches (10%), and 13 right coronary arteries (3.5%). RESULTS: There was no operative death or stroke. Early postoperative angiography revealed 95.5% (321/336) graft patency with 100% patency (119/119) of ITA to LAD grafts. Graft patency of the ITAs and RAs (98.5% and 95.9%) were significantly better than that of the GEA (79.0%, p = 0.0064 and p = 0.030) and saphenous vein (82.3%, p = 0.011 and p = 0.048). CONCLUSION: OPCAB performed with the aorta no-touch technique using an in-situ graft and composite and sequential grafting methods provides excellent early clinical results and graft patency. PMID- 12114125 TI - Technical aids to performing thoracoscopic robotically-assisted internal mammary artery harvesting. AB - OBJECTIVE: This report outlines the procedures and technical aids used for performing thoracoscopic internal mammary artery (IMA) harvesting in a series of 308 patients. METHODS: As a part of atraumatic coronary artery bypass (ACAB) operations, thoracoscopic IMA harvests (294 left, 14 right, and 12 bilateral) were performed in 308 consecutive patients. Single-lung ventilation and carbon dioxide insufflation were employed in all cases to facilitate exposure and dissection. A voice-activated robotic arm controlled the camera view, and harvesting was accomplished with the electrocautery on a low setting. RESULTS: Harvest time decreased from a mean of 58.4 minutes in the first fifty procedures to 29.4 minutes in the last fifty procedures. There were no significant complications as a result of this technique, and no patients required a conversion to sternotomy as a result of IMA injury. CONCLUSIONS: Thoracoscopic internal mammary artery harvesting is an essential basic skill for cardiac surgeons interested in performing minimally invasive and atraumatic coronary bypass procedures. Thoracoscopic IMA harvesting can be successfully performed with the use of the technical aids and procedures outlined in this report. PMID- 12114126 TI - Evaluation of acute traumatic changes of the coronary artery wall after robotically assisted endoscopic coronary artery bypass grafting. AB - BACKGROUND: There is concern that the technical limitations of robotic systems used in endoscopic coronary artery bypass grafting (CABG) may lead to increased trauma of the anastomotic site. To examine this issue, we compared the acute traumatic changes of the coronary artery wall caused by conventional manual suturing and robotically assisted suturing for anastomoses using the ZEUS telemanipulator (Computer Motion Inc., Goleta, CA) in a laboratory setting. METHODS: Coronary artery bypass grafting was performed on isolated porcine hearts. Fifteen anastomoses (with harvested porcine right coronary artery (RCA) segments) were carried out using the ZEUS microsurgical telemanipulator (group Z), while 15 further anastomoses were performed with a conventional manual technique (group M) using Gore-Tex CV-8 suture material. Specimens were taken from each anastomotic site and from native parts of the left anterior descending artery (LAD) (control group). Morphological changes of the cellular and fibrous components of the lamina intima and lamina media, and the shape and maximum diameter of the puncture mark, were examined by light microscopy (LM), transmission electron microscopy (TEM), and scanning electron microscopy (SEM). Vascular endothelial damage and denudation were graded on a score from 1 to 5. RESULTS: In each group, 14 specimens were evaluated. SEM findings showed a significantly higher degree of endothelial denudation in group Z and group M compared to the control group, while group Z was significantly more affected than group M. Likewise, the maximum diameter of the puncture mark was significantly larger in group Z than in group M. TEM and LM studies supported these results. In addition, LM revealed that in five specimens of group Z the shape of the stitch through the artery wall was not cylindrical, as in the other cases, but was asymmetrical and displayed a superficial furrow on the side of the vascular lumen. CONCLUSION: The results indicate that there is an increased incidence of damage to the coronary artery wall caused by the microsurgical telemanipulator. Further studies are necessary to determine whether the differences between conventional and robotic-assisted suturing techniques will have an effect on the long-term outcome of coronary artery bypass grafting. PMID- 12114127 TI - A new and simplified method for coronary and graft imaging during CABG. AB - BACKGROUND: Improvements in percutaneous catheter interventions and new technical demands in the practice of coronary surgery have increased the need for an accurate and easy-to-use imaging modality for validating the quality of bypass grafts in the operating room. This report examines the initial clinical use of fluorescent cardiac imaging, a technology that uses indocyanine green (ICG) with a portable imaging device to visualize coronary anatomy and grafts intraoperatively. METHODS: The modality was evaluated at two institutions in 20 patients undergoing non-emergent CABG or MIDCAB with respect to safety, feasibility of use, and image quality. Images were generated and acquired with a portable laser diode/infrared camera device after injection of 0.5 ml of ICG (0.5 5.0 mg/ml) either intravenously, via the antegrade cardioplegia cannula, or via the cardiopulmonary bypass circuit. RESULTS: There were no ICG- or imaging device related complications. The technology was easy-to-use during conventional CABG as well as MIDCAB and adequately demonstrated coronary anatomy, filling of the grafts, and graft patency in all but two patients. In one patient, the use of the modality resulted in the intraoperative recognition and revision of a non functioning graft. CONCLUSION: This technology is user-friendly in the operating room, appears to be safe, provides good-quality images of coronary anatomy and grafts, and holds promise as an intraoperative graft validation tool for conventional and minimally invasive CABG. PMID- 12114128 TI - Port-Access cardiac surgery: from a learning process to the standard. AB - BACKGROUND: Port-Access surgery has been one of the most innovative and controversial methods in the spectrum of minimally invasive techniques for cardiac operations and has been widely used for the treatment of several cardiac diseases. The technique was introduced in our center to evaluate its efficacy in reproducing standardized results without an additional risk. METHODS: Endovascular cardiopulmonary bypass (CPB) through femoral access and endoluminal aortic occlusion were used in 129 patients for a variety of surgical procedures, all of which were video-assisted. A minimal (4-6 cm) anterior thoracotomy through the fourth intercostal space was used in all cases as the surgical approach. RESULTS: More than 96% of the planned cases concluded as true Port-Access procedures. Mean CBP and crossclamp times were 87.2 min. +/- 51.2 (range of 10 457) and 54.9 min. +/- 30.6 (range of 10-190), respectively. Hospital mortality for the overall group was 1.5%, and mitral valve surgery had a 2.2% hospital death rate. The incidence of early neurological events was 0.7%. Mean extubation time, ICU stay, and total length of hospital stay were 5 hours +/- 6 hrs. (range of 2-32), 12 hours +/- 11.8 hrs. (range of 5-78), and 7 days +/- 7.03 days (range of 1-72), respectively. CONCLUSIONS: Our experience indicates that the Port- Access technique is safe and permits reproduction of standardized results with the use of a very limited surgical approach. We are convinced that this is a superior procedure for certain types of surgery, including isolated primary or redo mitral surgery, repair of a variety of atrial septal defects (ASDs), and atrial tumors. It is especially useful in high-risk patients, such as elderly patients or those requiring reoperation. Simplification of the procedure is nevertheless desirable in order to further reduce the time of operation and to address other drawbacks. PMID- 12114129 TI - A new live animal training model for off-pump coronary bypass surgery. AB - Training models are needed to perform accurate off-pump coronary artery bypass (OPCAB) surgery and to test evolving new technologies like minimally invasive devices and robotics. We describe a simple, effective and reproducible live animal training model to perform multiple arterial anastomoses on the beating heart that would maximize the use of available resources for training purposes. PMID- 12114130 TI - Proceedings of the First International Symposium on Beating Heart Surgery. December 4, 1999. Belo Hroizonte, Brazil. PMID- 12114131 TI - Minimally invasive direct coronary artery bypass grafting (MIDCAB) and off-pump coronary artery bypass grafting (OPCAB): two techniques for beating heart surgery. AB - OBJECTIVE: Coronary bypass surgery can be performed less invasively by avoiding cardiopulmonary bypass (CPB). We present our experiences with beating heart bypass surgery performed through a minithoracotomy or sternotomy. METHODS: From May 1997 to September 1999, 340 patients were included in a prospective study. Of these patients, 111 (group 1) underwent minimally invasive direct coronary artery bypass grafting (MIDCAB) through an antero-lateral minithoracotomy, and 229 (group 2) had off-pump coronary artery bypass grafting (OPCAB) through a full sternotomy. A pressure stabilizer was used for MIDCAB and a suction stabilizer for OPCAB surgery. Early postoperative angiography was performed on 48% of patients in group 1 and 45% of those in group 2. Statistical analysis was applied to compare the variables from both groups and a probability value of less than 0.05 was considered significant. RESULTS: In all MIDCAB grafts, revascularization was performed by a single left internal mammary artery (LIMA) graft to the left anterior descending coronary artery (LAD). This procedure was completed in 96.4% of patients without CPB. Conversion to sternotomy was necessary for one patient (0.9%). In the OPCAB group, an average of 1.7 grafts per patient were revascularized, of which 98 were single, 99 double, and 32 triple. Of the OPCAB group, 12% of patients were redo operations and 17% had severe comorbidities. Conversion to CPB was necessary for 10 patients (4.4%) because of hemodynamic instability. No cerebrovascular accident (CVA) was seen in any group. There were no hospital deaths in the MIDCAB group, but there were three deaths (1.3%) in the OPCAB group. Age, previous bypass surgery, and severe comorbidities did not influence early mortality. Early postoperative reoperation due to graft failure was necessary for three patients (2.7%) after MIDCAB and for three patients (1.3%) after OPCAB. Confirmed by angiography, the early postoperative total graft patency rate was 96.2% in the MIDCAB group and 96.6% in the OPCAB group; the perfect patency rate (no stenosis greater than 50%) was 92.4% and 93.1%, respectively. CONCLUSIONS: Coronary bypass surgery without the use of CPB is feasible and safe, and offers good early results. Nevertheless, MIDCAB grafting is a challenging technique and should only be performed in selected patients with favorable coronary anatomy. On the other hand, with the sternotomy approach, exposure of all vessels was well tolerated and made complete revascularization feasible. OPCAB can be performed safely even on high-risk patients. PMID- 12114133 TI - Total myocardial revascularization without cardiopulmonary bypass: a reality. AB - OBJECTIVE: From January 1995 to June 1999 our surgical team at ICORP, Fortaleza, Ceara, Brazil, performed coronary artery surgery without cardiopulmonary bypass as a routine procedure. A total of 897 operations were performed during this period, 91.8 % (824) of them without cardiopulmonary bypass. The purpose of the present study is to evaluate the results of these 824 patients with regard to duration of hospitalization, age, sex, number of grafts, reoperations performed, morbidity and mortality. All patients underwent previous coronary arteriography. METHODS: With patients admitted in the hospital for elective or emergency myocardial revascularization, all the coronary arteries were bypassed, possibly without cardiopulmonary bypass, including the marginal branches of the circumflex artery. Basic statistical analysis has been performed over the above-mentioned variables. RESULTS: The patients' ages ranged from 35 to 88 years, with a mean age of 61.2. The average duration of hospitalization was seven days. In all, 1,738 grafts were implanted, the number in individual patients varying from one to four (average of 2.1 per patient). The incidence of procedure-related complications was 3.15% (26 patients). Twenty-three patients (2.8%) died in the early postoperative period. CONCLUSIONS: Considering the data obtained from this study, we conclude that the procedure can be used in the vast majority of patients undergoing coronary artery surgery (compared to the similar studies of patients operated on with cardiopulmonary bypass). PMID- 12114132 TI - Ministernotomy in myocardial revascularization without cardiopulmonary bypass: technical aspects and early results. AB - OBJECTIVE: This study attempts to evaluate the feasibility of ministernotomy in beating heart coronary surgery, with special emphasis on technical aspects. METHODS: From September 1997 to September 1999, 137 patients were scheduled for off-pump coronary surgery in our institution. In 61 cases requiring revascularization of the left anterior descending artery (LAD) and right coronary artery (RCA) systems, the approach was either a reversed "L-shaped " ministernotomy (56 patients) or a "T-shaped " ministernotomy (five patients). Mean age of the ministernotomy patients was 64 +/- 10 years, and 17 of the patients were female. The mean left ventricular ejection fraction (LVEF) was 60 +/- 11% (<35% in four patients), and 32 patients (52.5%) had one-vessel disease while 29 (47.5%) had 2-vessel or 3-vessel diseases. There were seven (11.4%) urgent procedures. For these procedures, we used devices that we designed ourselves for sternal retraction and coronary stabilization. RESULTS: Five patients (8.2%) needed conversion to another method due to hemodynamic instability or ischemia, while 56 of the patients completed the procedure. Fifty one patients (91.1%) had a single graft on the LAD, four (7.1%) had a double graft on the LAD and the right or diagonal coronary artery, and one (1.8%) had a triple graft on the LAD and two diagonal branches. Mean coronary occlusion times and operative times were 12.1 +/- 2.7 and 152 +/- 33 minutes, respectively. Mean creatine kinase value was 29.8 +/- 24.6. One patient died of acute myocardial infarction, and one patient had temporary acute renal failure. Mean in-hospital stay was 5.2 +/- 1.9 days. Of the 18 patients (32.1%) who had postoperative angiographic control (range of 1 to 13 months), 17 showed patent anastomoses, and one required percutaneous transluminal coronary angioplasty (PTCA) of the anastomosis on the LAD. Mean follow-up time for all patients was 10.8 +/- 6.4 months. Freedom from any kind of repeat procedure was 98.2%. CONCLUSION: Ministernotomy is a safe approach for patients not requiring grafts on the circumflex system. The possibility of multiple grafting and the easy conversion to a conventional surgical method make ministernotomy a preferable approach for minimally invasive coronary surgery. PMID- 12114134 TI - Ambulatory care education: what we know and what we don't. PMID- 12114136 TI - An efficient and effective teaching model for ambulatory education. AB - Teaching and learning in the ambulatory setting have been described as inefficient, variable, and unpredictable. A model of ambulatory teaching that was piloted in three settings (1973-1981 in a university-affiliated outpatient clinic in Portland, Oregon, 1996-2000 in a community outpatient clinic, and 2000-2001 in an outpatient clinic serving Dartmouth Medical School's teaching hospital) that combines a system of education and a system of patient care is presented. Fully integrating learners into the office practice using creative scheduling, pre rotation learning, and learner competence certification enabled the learners to provide care in roles traditionally fulfilled by physicians and nurses. Practice redesign made learners active members of the patient care team by involving them in such tasks as patient intake, histories and physicals, patient education, and monitoring of patient progress between visits. So that learners can be active members of the patient care team on the first day of clinic, pre-training is provided by the clerkship or residency so that they are able to competently provide care in the time available. To assure effective education, teaching and learning times are explicitly scheduled by parallel booking of patients for the learner and the preceptor at the same time. In the pilot settings this teaching model maintained or improved preceptor productivity and on-time efficiency compared with these outcomes of traditional scheduling. The time spent alone with patients, in direct observation by preceptors, and for scheduled case discussion was appreciated by learners. Increased satisfaction was enjoyed by learners, teachers, clinic staff, and patients. Barriers to implementation include too few examining rooms, inability to manipulate patient appointment schedules, and learners' not being present in a teaching clinic all the time. PMID- 12114138 TI - A collaborative model for supporting community-based interdisciplinary education. AB - Development and support of community-based, interdisciplinary ambulatory medical education has achieved high priority due to on-site capacity and the unique educational experiences community sites contribute to the educational program. The authors describe the collaborative model their school developed and implemented in 2000 to integrate institution- and community-based interdisciplinary education through a centralized office, the strengths and challenges faced in applying it, the educational outcomes that are being tracked to evaluate its effectiveness, and estimates of funds needed to ensure its success. Core funding of $180,000 is available annually for a centralized office, the keystone of the model described here. With this funding, the office has (1) addressed recruitment, retention, and quality of educators for UME; (2) promoted innovation in education, evaluation, and research; (3) supported development of a comprehensive curriculum for medical school education; and (4) monitored the effectiveness of community-based education programs by tracking product yield and cost estimates needed to generate these programs. The model's Teaching and Learning Database contains information about more than 1,500 educational placements at 165 ambulatory teaching sites (80% in northern New England) involving 320 active preceptors. The centralized office facilitated 36 site visits, 22% of which were interdisciplinary, involving 122 preceptors. A total of 98 follow-up requests by community-based preceptors were fulfilled in 2000. The current submission-to-funding ratio for educational grants is 56%. Costs per educational activity have ranged from $811.50 to $1,938, with costs per preceptor ranging from $101.40 to $217.82. Cost per product (grants, manuscripts, presentations) in research and academic scholarship activities was $2,492. The model allows the medical school to balance institutional and departmental support for its educational programs, and to better position itself for the ongoing changes in the health care system. PMID- 12114137 TI - Development of a handheld computer documentation system to enhance an integrated primary care clerkship. AB - Documentation systems are used by medical schools and residency programs to record the clinical experiences of their learners. The authors developed a system for their school's (Dartmouth's) multidisciplinary primary care clerkship (family medicine, internal medicine, pediatrics) that documents students' clinical and educational experiences and provides feedback designed to enhance clinical training utilizing a timely data-reporting system. The five critical components of the system are (1) a valid, reliable and feasible data-collection instrument; (2) orientation of and ongoing support for student and faculty users; (3) generation and distribution of timely feedback reports to students, preceptors, and clerkship directors; (4) adequate financial and technical support; and (5) a database design that allows for overall evaluation of educational outcomes. The system, whose development began in 1997, generated and distributed approximately 150 peer-comparison reports of clinical teaching experiences to students, preceptors, and course directors during 2001, in formats that are easy to interpret and use to individualize learning. The authors present report formats and annual cost estimate comparisons of paper- and computer-based system development and maintenance, which range from $35,935 to $53,780 for the paper based system and from $46,820 to $109,308 for the computer-based system. They mention ongoing challenges in components of the system. They conclude that a comprehensive documentation and feedback system provides an essential infrastructure for the evaluation and enhancement of community-based teaching and learning in primary care ambulatory clerkships, whether separate or integrated. PMID- 12114139 TI - Assessing quality and costs of education in the ambulatory setting: a review of the literature. AB - PURPOSE: Time-pressured interactions with little direct observation or feedback characterize teaching in ambulatory settings. The authors report findings from the literature on teaching and learning in the ambulatory setting and propose opportunities for further research that addresses these barriers. METHOD: The authors searched 1995-1999 databases for all empirical studies that focused on research conducted in ambulatory settings. Publications were reviewed for evidence of inclusion criteria. Findings were sorted into categories previously described for defining and evaluating quality of ambulatory care educational programs. RESULTS: Most studies were conducted in departments of internal medicine (40%), focused on medical students (43%), and took place in a single program (77%), making generalizations difficult. Students and residents are learning in ambulatory environments, and the types of patients they encounter are likely to prepare them for practice. Patient care outcomes have emerged as a measure of learning. Teachers may be the single most important factor, yet they lack self-confidence as teachers. Community-based preceptors teach because of enjoyment of teaching and the opportunity to stay current. However, none of the studies addressed the impact of the Medicare documentation requirements on satisfaction with teaching. Teaching settings cost about one third more than non teaching settings to operate. CONCLUSION: This review identifies many gaps in our knowledge of effective clinical teaching practices, and of learning environments in which that teaching takes place. The predominance of single-institution studies limits generalizability of current findings. A prioritized research agenda should be established and funded, focusing on improving the efficiency and effectiveness of teaching and learning in ambulatory settings. PMID- 12114140 TI - An analysis of students' clinical experiences in an integrated primary care clerkship. AB - PURPOSE: Combining complementary clinical content into an integrated clerkship curriculum should enhance students' abilities to develop skills relevant to multiple disciplines, but how educational opportunities in primary care ambulatory settings complement each other is unknown. The authors conducted an observational analytic study to explore where opportunities exist to apply clinical skills during a 16-week integrated primary care clerkship (eight weeks of family medicine, four weeks of ambulatory pediatrics, and four weeks of ambulatory internal medicine). METHOD: Using handheld computers, students recorded common problems, symptoms, and diagnoses they saw. The students also recorded information about the educational process of the clerkship. Two data files were created from the database. Descriptive statistics were used to characterize the students' clerkship experiences, and ANOVA was used to evaluate differences among these blocks within the clerkship. RESULTS: Students encountered different frequencies of presenting symptoms, the majority of which occurred in pediatrics (23.2 per student per week versus 16.3 in medicine and 16.8 in family medicine; p =.01). Students provided more behavioral change counseling in family medicine (5.2 episodes per student per week versus 4.2 and 2.0 in internal medicine and pediatrics, respectively; p =.01), and they performed more clinical procedures in family medicine (1.9 per student per week versus 0.6 and 1.1 in pediatrics and internal medicine, respectively; p =.001). Students were more likely to encounter specific conditions in internal medicine (35.3 per student per week versus 30.0 and 21.4 in family medicine and pediatrics, respectively; p =.01). Elements of the teaching and learning processes also differed by clerkship. CONCLUSIONS: Very little overlap was found in symptoms, conditions, procedures, and other educational opportunities in the ambulatory pediatrics, internal medicine, and family medicine blocks that constitute the integrated primary care clerkship. The blocks provided different and complementary learning opportunities for students. These findings will assist in clerkship planning and in guiding students to seek opportunities that will ensure educational excellence. PMID- 12114142 TI - Communication with deaf and hard-of-hearing people: a guide for medical education. AB - Some physicians may be insufficiently prepared to work with the many patients who have hearing loss. People with hearing loss constitute approximately 9% of the U.S. population, and the prevalence is increasing. Patients with hearing loss and their physicians report communication difficulties; physicians also report feeling less comfortable with these patients. Although communication with patients plays a major role in determining diagnoses and management, little attention is given to teaching medical students and residents the skills necessary to facilitate communication when hearing loss is involved. The need for these skills will increase with the expected rise in the number of such patients. The author presents the rationale for including information about hearing loss in curricula on patient-doctor communication, and suggests curricular content, including background regarding hearing loss and techniques that can enhance the physician's ability to listen to (that is, "hear") and learn about the stories of these patients. PMID- 12114141 TI - Evidence for longitudinal ambulatory care rotations: a review of the literature. AB - PURPOSE: Block ambulatory rotations and longitudinal ambulatory care experiences are now common in U.S. medical schools, but little is known about their efficacy. Through a structured review of the medical literature from 1966 through March 2000, the authors summarize the characteristics of, the evidence for, and the evaluation of longitudinal ambulatory care rotations. METHOD: The authors searched Medline using the terms "outpatients," "continuity of patient care," "ambulatory care," "mentors," "preceptorship," "graduate medical education," "curriculum," and "clinical clerkship" cross-matched to "medical students" and "internship and residency" for literature published from 1966 through March 2000. They narrowed the list to only articles containing empirical outcome data focusing on medical students' experiences in longitudinal ambulatory care rotations. Each study was scored to assess its strengths and weaknesses. RESULTS: Seven articles met the search criteria. The articles identified the benefits of longitudinal ambulatory care experiences, including developing effective patient interactions and understanding chronic diseases. There were little or no differences in the students' overall knowledge acquisition when those with longitudinal experiences were compared with those in block rotations. DISCUSSION: Although longitudinal ambulatory care experiences are now common in medical schools, evidence supporting their widespread implementation is sparse. Few studies employ rigorous methods to evaluate educational outcomes. Research to identify benefits and costs, improve the quality and consistency of the students' experiences, and develop other innovative ways of teaching and learning ambulatory care is needed. PMID- 12114143 TI - The challenge of teaching rehabilitative care in medical school. AB - Rehabilitative care has gained importance because the population is aging, and improved acute and chronic medical care saves and prolongs lives but leaves some patients with temporary or permanent physical impairments. However, despite its importance, the teaching and learning of rehabilitative care in medical school lag behind medical education relating to acute and chronic care. The authors analyze the broad scope of rehabilitative care and the need to include it in the medical school curriculum. They also discuss advantages for students and their patients of learning rehabilitative care in the undergraduate curriculum and suggest methods to improve teaching it. PMID- 12114144 TI - The professor and the outpatient department. 1966. PMID- 12114145 TI - Learning medicine with a learning disability: reflections of a survivor. PMID- 12114148 TI - The training and career paths of fellows in the National Research Service Award (NRSA) Program for Research in Primary Medical Care. AB - PURPOSE: To describe the training and career paths of fellows in the National Research Service Award (NRSA) Program for Research in Primary Medical Care. METHOD: All fellows who graduated from 25 NRSA programs nationally between July 1988 and June 1997 (n = 215) were mailed a questionnaire. Personal characteristics, fellowship experiences, and current professional positions were compared between faculty researchers, faculty clinician-educators, and individuals who were not in full-time academic positions. RESULTS: A total of 146 NRSA graduates (68%) completed the survey. A mean of four years had elapsed since their fellowships. Of the respondents, 36% were faculty researchers, 32% were faculty clinician-educators, and 32% were not on full-time faculties. Faculty researchers did not differ from the other groups in demographics or acquisition of advanced degrees, but they were more often general internists than general pediatricians, family physicians, or from other disciplines (p =.002). Fellowship graduates spent a mean of 29% of their training in course work and 38% conducting research. Faculty researchers spent a greater proportion of their fellowship conducting research (46% versus 34% for clinician-educators and 31% for those not on full-time faculties, respectively, p <.0001). They were also more productive in terms of subsequent publications and grant acquisitions. CONCLUSIONS: Only a minority of those completing NRSA programs held positions as faculty researchers. The preponderance of general internists among researchers may indicate problems in the capacity of general pediatrics and family medicine to support primary care research. The amounts of direct research time during these fellowships may need to be increased to enhance the likelihood of subsequent research success. PMID- 12114149 TI - Premed survival: understanding the culling process in premedical undergraduate education. AB - PURPOSE: Why undergraduate students pursue or drop a premedical curriculum has received only scant attention. In this study the authors attempted to uncover reasons why students either persevere in their premedical studies or seek alternative careers. METHOD: Using convenience sampling, the authors surveyed 97 undergraduates at a small liberal arts college from November 2000 to March 2001. Of those surveyed, 44 were former premed students who completed a three-page questionnaire about why they had decided not to become physicians; 53 premed students completed a two-page questionnaire about their career aspirations. RESULTS: The response rate was 100%. Premed students were attracted to the field by the intellectual stimulation and the power to help others, yet most were also very concerned about being in debt, dealing with patients who might die, and the compatibility of medicine with having a family. Women students were more concerned than the men about having only limited time to become acquainted with patients on a social level (71% of women versus 45% of men: p =.05). The decision of students to forgo a career as a physician was shaped by apprehensions regarding the years of work required in residency, the need to be on call, unacceptably low grades, and the realization that other attractive career options are available. Of those who said low grades were a deciding factor, most (78%) named organic chemistry as the single course that had affected their plans. Students who acknowledged the role of their poor performance in organic chemistry were more likely to be dissatisfied with their change in plans than were those who did not identify this course as influential (44% versus 29%). CONCLUSIONS: Although the sampling technique and sample size severely limit the authors' ability to generalize their findings, the data offer a starting point for those interested in the reasons for the drop in medical school applicants. The authors state the fact that most former premed students admitted organic chemistry had played a significant role in the change in their career plans deserves attention, and it may be time to consider whether a single course should contribute to eliminating persons who might otherwise excel as physicians. PMID- 12114151 TI - Standard setting: a comparison of case-author and modified borderline-group methods in a small-scale OSCE. AB - PURPOSE: To compare cut scores resulting from the case-author method and the modified borderline-group method (MBG) of standard setting in an undergraduate objective structured clinical examination (OSCE), and to review the feasibility of using the MBG method of standard setting in a small-scale OSCE. METHOD: Sixty one fourth-year medical students underwent a ten-station OSCE examination. For the eight stations used in this study, cut scores were established using the case author and MBG methods. Cut scores and pass rates were compared for individual stations and the entire exam. RESULTS: The case-author and MBG methods of standard setting produced different cut scores for the entire examination (5.77 and 5.31, respectively) and for each station individually. The percentage of students failing the examination based on the case-author cut score was 42.2%, and based on the MBG cut score it was 15.25%. CONCLUSIONS: The case-author and MBG methods of standard setting produced different cut scores in an undergraduate OSCE. Overall, the MBG method was the more credible and defensible method of standard setting, and appeared well suited to a small-scale OSCE. PMID- 12114150 TI - The effect of candidates' perceptions of the evaluation method on reliability of checklist and global rating scores in an objective structured clinical examination. AB - PURPOSE: Process-oriented global ratings, which assess "overall performance" on one or a number of domains, have been purported to capture nuances of expert performance better than checklists. Pilot data indicate that students change behaviors depending on their perceptions of how they are being scored, while experts do not. This study examines the impact of the students' orientation to the rating system on OSCE scores and the interstation reliability of the checklist and global scores. METHOD: A total of 57 third- and fourth-year medical students at one school were randomly assigned to two groups and performed a ten station OSCE. Group 1 was told that scores were based on checklists. Group 2 was informed that performance would be rated using global ratings geared toward assessing overall competence. All candidates were scored by physician-examiners who were unaware of the students' orientations to the rating system and who used both checklists and global rating forms. RESULTS: A mixed two-factor ANOVA identified a significant interaction of rating form by group (F(1,55) = 5.5, p <.05), with Group 1 (checklist-oriented) having higher checklist scores but lower global scores than did Group 2 (oriented to global ratings). In addition, Group 1 had higher interstation alpha coefficients than did Group 2 for both global scores (0.74 versus 0.63) and checklist scores (0.63 versus 0.40). CONCLUSIONS: The interaction effect on total exam scores suggests that students adapt their behaviors to the system of evaluation. However, the lower reliability coefficients for both forms found in the process-oriented global-rating group suggest that an individual's capacity to adapt to the system of global rating forms is relatively station-specific, possibly depending on his or her expertise in the domain represented in each station. PMID- 12114152 TI - Study month or vacation? Preparing for USMLE Step 2. AB - Students often request a "study month" prior to taking the USMLE Step 2 to maximize their performance on the exam. This report questions the utility of this approach. PMID- 12114153 TI - Reflective practice skills in undergraduates. AB - Critical self-appraisal skills are fundamental to professional competence. This study evaluated the educational contribution of self-assessment during individual students' projects. PMID- 12114154 TI - Health science learning academy: a successful "pipeline" educational program for high school students. AB - OBJECTIVE: The objective of the Health Professions Partnership Initiative is to increase the number of underrepresented minority Georgia residents who become health care professionals by (1) creating a pipeline of well-qualified high school and college students interested in health care careers, (2) increasing the number of well-qualified applicants to medical and other health professions schools, and (3) increasing the number of underrepresented minority students at the Medical College of Georgia (MCG). DESCRIPTION: The Health Professions Partnership Initiative at MCG was created in 1996 by collaboration among the MCG Schools of Medicine and Nursing, two Augusta high schools attended primarily by underrepresented minority students, three historically black colleges and universities, the Fort Discovery National Science Center of Augusta, community service organizations, and MCG student organizations. The project was funded by the Association of American Medical Colleges and The Robert Wood Johnson Foundation. The high school component, the Health Science Learning Academy (HSLA), was designed to strengthen the students' educational backgrounds and interest in professional careers as evidenced by increased standardized test scores and numbers of students entering college and health professions schools. Additional goals included a system to track students' progress throughout the pipeline as well as professional development sessions to enrich faculty members' knowledge and enhance their teaching expertise. The HSLA began with ninth-grade students from the two high schools. During its second year, funding from the Health 1st Foundation allowed inclusion of another high school and expansion to ninth grade through twelfth grade. The HSLA's enrichment classes meet for three hours on 18 Saturday mornings during the academic year and include computer interactive SAT preparation and English composition (tenth grade); biology, algebra, calculus, and English composition (eleventh grade); and advanced mathematics and biology (twelfth grade). DISCUSSION: The ultimate solution to the paucity of underrepresented minority physicians resides largely in successful pipeline programs that expand the pool of well-qualified applicants, matriculants, and graduates from medical schools. Intermediate results of the HSLA support the success of the program. Since its creation in the 1996-1997 academic year, 203 students have participated in the HSLA and all 38 (from the original two schools) who completed the four-year program have enrolled in college. The mean SAT score for students who completed the HSLA program was 1,066, compared with a mean of 923 for all college-bound students in the participating schools. The mean increases in SAT scores for students who completed the four-year program were.5% (1,100 to 1,105) for students attending a magnet high school and 18% (929 to 1,130) for students attending the comprehensive high school. The mean overall increases in SAT scores for students in the two high schools were 1% (1,044 to 1,048) and 9.1% (765 to 834), respectively. The HSLA is accomplishing its goals and, while it is too early to know if these students will participate in MCAT preparatory programs and apply to medical and other health professions schools, their sustained commitment and enthusiasm bode well for continued success. PMID- 12114155 TI - Educational assessment center techniques for entrance selection in medical school. AB - OBJECTIVE: Dutch higher education is freely accessible for those who have proper high school qualifications. However, admission to medical schools has been limited by government to regulate manpower planning. Selection has been carried out by a national lottery approach since 1972, but in 2000, the Dutch government asked medical schools to experiment with qualitative selection procedures at their own institutions. The University Medical Center Utrecht School of Medical Sciences has used a technique derived from assessment-center approaches to assist in the medical school admission process. Dutch assessment centers use observation procedures in which candidates act in simulated activities that are characteristic of the vacant position. DESCRIPTION: In April 2001, 61 candidates for 23 places were invited for selection days. After a selection interview, candidates were asked to perform activities that are characteristic of course requirements: (1) studying a three-to-five page text about diagnostic and therapeutic procedures of disease A during one hour; (2) explaining the studied procedures to another candidate and receiving information about disease B, studied by this other candidate, during one hour; (3) answering the questions of a standardized patient about disease A in 15 minutes; and (4) answering the questions of a standardized patient about disease B in 15 minutes. A three-person selection committee behind a one-way screen observed the two 15-minute interviews with the standardized patients. The selection committee independently scored content quality of the information that was given to the standardized patients as well as the quality of attitude towards and communication with both patients. The average scores for these three criteria were weighted equally to arrive at a total score. In addition, each candidate received a score resulting from the interview with the other candidate who explained disease B. This score was combined with the other three to a final score. DISCUSSION: The Utrecht medical curriculum may be viewed as a hybrid PBL program. Integration of basic and clinical sciences, patient contacts from the start, training of skills in communication with standardized patients, physical examination, extensive small group teaching, structured independent studying, and collaboration to prepare for short presentations to peers were all characteristic of the medical school curriculum. Thus, the assessment-center technique reflected the characteristics of the medical school curriculum. First analyses showed satisfactory reliabilities of the three scores (0.79 to 0.92); the average agreement between raters was 0.60. Correlation analysis between scores supported the internal convergent and discriminant validity of the assessment activities. The predictive validity remains to be studied. PMID- 12114156 TI - Linking cultural competency and community service: a partnership between students, faculty, and the community. AB - OBJECTIVE: The purpose of the migrant health initiative is to give medical students the opportunity to provide clinical services, at appropriate levels of training, to a population that reflects a different ethnic and economic background than medical students typically see in the clinical setting. This initiative integrates concepts of cultural competency with experiential learning. DESCRIPTION: The migrant health initiative provides an infrastructure for a cultural competency educational program in the first two years of medical school within an essentials of clinical medicine course (ECM). The ECM course emphasizes the impacts of family, society, and community on the delivery of patient health care. Experience in the provision of clinical care to migrant workers provides an exceptional opportunity to expose students to a medically underserved, diverse group of people and to provide care to persons with a different language. The program was developed from grass-roots initiatives of students and the region's Area Health Education Center (AHEC). Historically, migrant workers provide the majority of the labor for harvesting onions in southeast Georgia. In April 2000, the regional AHEC organized a one-day clinic for migrant workers, staffed by one local physician and allied health students. Care was provided to over 400 laborers. As a result of the response by the migrant workforce, the AHEC developed a partnership with the local community to expand the health care services to this underserved group. Five medical students, working with the school's associate dean for curriculum and local AHEC, had observed a migrant health clinic organized by the AHEC and were seeking ongoing community service opportunities. Based on the interest and enthusiasm of the students, a faculty supervised migrant health elective was developed for first- and second-year students. The number of students who wanted to take the elective exceeded the available opportunities. The ECM course will be enhanced with an integrated and longitudinal curriculum that focuses on migrant health care; the revised ECM course will provide students with the knowledge, skills, attitudes, and behaviors to care for individuals from different cultures. First-year students will be able to volunteer to work in the program. Second-year students will participate in at least one migrant health clinic, traveling only three hours but providing a different world with medical care. In addition, the opportunities for medical students to participate in a community health initiative and to work with nursing and allied health students will enhance their public health knowledge and their team and leadership skills. DISCUSSION: The partnership between students, faculty, and the community provides the mechanism to thoughtfully develop and integrate cultural issues and experiences into the curriculum. Students have recently received a Caring for Community five-year grant from the Association of American Medical Colleges. Program expansions will continue into the third-year medicine clerkship and include a senior elective. The program expansions will result in a migrant health initiative that will be coordinated; comprehensive; and expand student knowledge, skills, and experiences in cultural health care. PMID- 12114157 TI - Community medicine in action: an integrated, fourth-year urban continuity preceptorship. AB - OBJECTIVE: To provide an opportunity for fourth-year students at the University of Wisconsin Medical School in Madison to immerse in urban community medicine during a 34-week program. This experience enhances the integrity of the fourth year as well as merges medicine and public health perspectives in medical education as called for by the Medicine and Public Health Initiative. DESCRIPTION: A limited number of fourth-year Wisconsin medical students have the opportunity to select a one-year, continuity-based preceptorship at the Milwaukee clinical campus with a focus in one of three domains: family medicine, internal medicine, or women's health. Students participate in the following clinical activities: a one-year, integrated preceptorship (one to three half days per week in a primary preceptor's office), medicine subinternship, senior surgery clerkship, selectives (16-20 weeks of clerkships relevant to preceptorship focus area), and one month of out-of-city electives. Complementing this community-based clinical experience is the opportunity to develop an increased appreciation for urban community health issues and resources by participating in a required urban community medicine clerkship and a mentored student scholarly project focusing on an aspect of urban community medicine and population health. All students begin the year in July with a four-week urban community medicine clerkship, which is based on the St. Luke's family practice residency's community medicine rotation and arranged by residency faculty. They conduct a "windshield survey" of a Milwaukee neighborhood, observing health hazards and identifying assets, and then present these observations to others in the clerkship. During this first month, students are introduced to the work of a variety of social service agencies, the Milwaukee City Health Department, and the Aurora Health Care/UW community clinics, which serve the state's most diverse zip codes. They meet with providers and researchers who share their expertise in infectious disease, preventive medicine, perinatal epidemiology, domestic violence, sexual assault, and disease management. Students develop increased understanding of barriers to health and personal resilience by listening to focus groups conducted with homeless men and undocumented Latino women. They participate in a resident and faculty development retreat on enhancing community medicine knowledge and skills. By August, students select an advisor and outline a project designed to expand understanding in the areas of urban population health research, community health education, professional education, or health intervention planning and evaluation. Faculty members at the Center for Urban Population Health work closely with the students throughout the year, which includes two weeks in the spring that are dedicated to intensive work on the projects. DISCUSSION: This fourth-year, urban community based preceptorship is designed to provide students with an alternative fourth year that integrates skill development in clinical and community medicine, offers a continuity primary care experience, and showcases innovative urban health resources and role models. It is hoped that these students will pursue graduate medical education in Milwaukee, incorporate a population perspective in their practice, and choose to work in neighborhoods that are currently underserved. PMID- 12114158 TI - Ambulatory rounds: a venue for evidence-based medicine. AB - OBJECTIVE: The format of inpatient morning reports and ward rounds is infrequently applied in ambulatory medical education. Published reports, however, suggest that this format provides for learner-centered, case-based discussions rather than topic-based lectures in the ambulatory setting.(1) We developed an ambulatory morning report with the specific objective of enhancing evidence-based medical inquiry among our pediatrics housestaff. DESCRIPTION: We developed a pediatric encounter form (PEF) by adapting and modifying an instrument described by Paccione et al.(2) The PEF was to be used by residents to document pertinent information and unanswered questions about patients seen during each ambulatory clinic session. Prompts were provided for documenting the patient's primary complaints, the patient's disposition, and questions that the resident needed answered. The PEF was piloted among a group of residents and faculty. The final version incorporated both resident and faculty input. Each resident was asked to complete a PEF for a maximum of two patients per clinic session. We did not direct residents as to what types of questions to formulate. All completed forms were maintained in a central folder. Next, we instituted a one-hour "Ambulatory Rounds" seminar once a week at lunch-time. During these seminars, faculty selected PEF cases from the previous week for discussion. Residents presented the cases and discussed the reasons behind the formulation of their questions. Faculty facilitated and guided residents toward resources for answering their questions. Faculty also helped residents to reformulate their questions to reflect an evidence-based medicine approach. At the end of each seminar, residents elected to research specific questions and present brief reports at the next seminar. To test the hypothesis that residents will formulate a higher proportion of evidence-based medicine (EBM) questions over time, we collected and analyzed 445 questions asked by 12 residents between July 2000 and August 2001. We categorized questions into EBM and non-EBM questions based on faculty assessment. We performed a trend analysis using chi-square to compare questions from July 2000 (as reference value) with the six-month periods of August 2000 to January 2001 and February to August 2001. By the end of the observation period, the proportion of EBM questions had significantly increased from 13% in July 2000 to 28% in the first six-month period and 59% in the second six-month period (p < 0.001). DISCUSSION: We describe a new application of outpatient morning reports. This format has been very well received. Housestaff gave the ambulatory rounds an average rating of 4.3 (out of 5) on a Likert scale. Our experience suggests that this format not only provides a forum for case-based learning but can be successfully used to enhance the principles of evidence-based medicine among residents. PMID- 12114159 TI - Leadership Opportunities with Communities, the Medically Underserved, and Special Populations (LOCUS). AB - OBJECTIVES: The Leadership Opportunities with Communities, the Underserved, and Special Populations (LOCUS) Program aims to improve medical students' leadership knowledge and skills, to improve self-awareness and motivation for community service, and to provide models for students to integrate community service into their medical careers. DESCRIPTION: The LOCUS program was established as a longitudinal, extracurricular student opportunity at the University of Wisconsin Medical School in the fall of 1998. Up to 15 new students each year are selected for the program through an application and interview process during their first or second year of medical school. Students remain in the program from acceptance until graduation from medical school. Nearly 50 students have enrolled in the program to date. LOCUS fellows are matched with a physician mentor, participate in core curriculum activities, and complete a longitudinal community service project. Mentors are community generalist physicians who have integrated community service into their own careers. Students participate in their mentors' clinical practices one afternoon a month during the first two years, and mentors serve as role models and provide guidance for students' projects and career development. The program administration and staff are supported through federal predoctoral training and Area Health Education Centers (AHEC) grants. The LOCUS core curriculum is delivered through a series of retreats, workshops, and seminars that emphasize active learning methods and include approximately 20 hours of scheduled activities per academic year. The curriculum addresses concepts of leadership in relation to one's self and in relation to others. Students are introduced to methods of self-reflection and develop their own vision and mission statements. Students also discuss the importance of compassion, self-care, striving for balance, avoiding burnout, and being realistic about what they can accomplish. Students practice strategies for working with teams, organizing meetings, working with media, taking political action, and resolving conflicts. They acquire community health skills such as assessing the health needs of a defined population; engaging community members' participation in health program development; and selecting priorities, designing interventions, and measuring the progress of community health care. Working in small teams, LOCUS fellows apply and refine their leadership skills through design and completion of a community health service project. Students can design their own projects or work on projects designed by community partners. The projects have addressed a variety of community health needs, such as parenting support for teen mothers, teaching health education for residents of group homes, and providing free sports physical exams for uninsured youth. DISCUSSION: This pilot program demonstrates that motivated students can develop leadership skills and address unmet community health needs while they progress through medical school. LOCUS students, staff, and physicians provide a social network that includes opportunities, encouragement, reflection, and problem solving. Student and mentor satisfaction with the program has been high. Future challenges include securing long-term funding, refining the core curriculum, assessing the impact of the program on participants, and improving the quality of projects through community partnerships. LOCUS strives to kindle the fires of altruism and community service so they are not extinguished as students progress through medical training. PMID- 12114160 TI - Mixing it up: integrating evidence-based medicine and patient care. AB - OBJECTIVE: To teach internal medicine residents to use evidence-based medicine (EBM) in their interactions with patients by creating curricula that integrate EBM into clinical topics in internal medicine. DESCRIPTION: The last several years have brought the wide-spread inclusion of EBM in internal medicine training programs in the United States. However, EBM is often taught as an independent topic and is poorly integrated into the clinical teaching of trainees. Most EBM education occurs in a journal-club format, focusing on question development, searching, and critical appraisal. The challenge of discussing the evidence with patients is rarely addressed. We set out to integrate EBM teaching into new curricula in women's health, addiction medicine, and topics in anticoagulation. During the first of two ambulatory blocks of the year, residents participate in an EBM seminar series in which they present cases, generate questions, and critically appraise the evidence. Second-year residents present articles on therapy or diagnosis and third-year residents present articles on diagnosis, meta analysis, or decision and economic analysis. Both the women's health curriculum and the anticoagulation curriculum are presented during the second ambulatory block of the year as four half-day small-group seminars. The women's health curriculum is presented to the second-year residents and the anticoagulation curriculum is presented to the third-year residents. Both curricula are case based and emphasize essential skills in patient care, including interview techniques, sensitivity to psychosocial issues, and skills in evidence-based patient care. Teaching EBM is not identified to the residents as a goal of these curricula; instead EBM, psychosocial medicine, and communication skills are woven into the content material and taught in the context of the broader subject matter. Learners are expected to integrate these concepts into actual practice. The curricula utilize clinical vignettes and role-plays to link EBM concepts such as number needed to treat or decision analysis to real-patient decisions. Residents are also asked to apply the evidence in their own patient encounters for further discussion at later sessions. Simpler concepts of therapy and diagnosis are covered during the second year in women's health and the more complex concepts of meta-analysis; decision analysis, and economic analysis are covered during the third year in anti-coagulation. DISCUSSION: The women's health curriculum was introduced in the spring of 2000; the anticoagulation curriculum was introduced in the spring of 2001. Both have been well received and seem to have impacted the ability of our housestaff to incorporate EBM into patient care. Currently under development in this series is a curriculum in addiction medicine for interns that will use a similar approach to provide an overview of EBM topics and their integration into the flow of patient care. We feel that these educational programs have helped EBM to bridge the gap between the classroom and the exam room. PMID- 12114161 TI - A picture is worth more than a thousand words: enhancement of a pre-exam telephone consultation in dermatology with digital images. AB - OBJECTIVE: In addition to the assessment and the management of patients with skin diseases, a considerable portion of dermatology residency involves examining clinical images and generating differential diagnoses from these images. This training, though helpful for recognizing manifestations of rare disorders, goes unused by most practicing dermatologists after certification. In contrast, dermatology residents learn and master verbal descriptions of skin diseases and continue to use this skill throughout their careers. However, problems arise when a dermatologist is not available and a non-dermatologist attempts to verbally describe a skin condition. An accurate description of a cutaneous disorder can facilitate effective triage management of a patient when a dermatologist is not available. Unfortunately, an inaccurate description by the referring provider can lead to diagnostic bias and ineffective, or even harmful, initial treatment. In recent years, digital photography has facilitated the electronic transfer of clinical images over distances. However, despite the promise that this technique shows in providing teledermatologic services to specialty-underserved areas and the availability of low-cost digital cameras, telephone consultation is still the standard of care when a dermatologist is not available. The purpose of this study is to compare the reliability of dermatologic consultations that use the telephone with that of dermatologic consultations that use both the telephone and digital images. DESCRIPTION: After patient approval, an acute care provider randomly assigned patients with skin disorders of unclear etiology to two groups, with and without digital images. The acute care provider then performed an exam and took the patient's history. Telephone data, with or without digital images, were then presented to the consulting dermatologist, who formulated a pre physical exam differential diagnosis and treatment plan. The consulting dermatologist immediately examined the patient in person and refined the diagnosis and management. The confidence in diagnosis, both before and after the in-person exam, was compared in the patient group with digital images and in the patient group without digital images using a five-point scale (1 = no confidence, 5 = most confident). DISCUSSION: The consulting dermatologist evaluated 12 patients (six with digital images and six without digital images). In the patient group with digital images, the consulting dermatologist's confidence in diagnosis varied very little from before to after the in-person exam (from no change in five cases to a one-point increase in the sixth case). In the patient group without digital images, the consulting dermatologist's confidence level increased significantly from before to after the in-person exam. This led to therapy changes for three of the six patients in the patient group without digital images, versus two of the six patients in the patient group with digital images. This study indicates that an acute care provider's verbal description of a skin condition may be less reliable compared with a provider's verbal description combined with digital images. Telephone-only descriptions may also lead to management discrepancies more frequently than telephone descriptions with digital images. This has at least two implications for medical education: (1) need for support of formal teaching of the language of dermatology to non-dermatologists and (2) justification of the time spent in two-dimensional clinical image interpretation by dermatology residents in light of digital image technology. PMID- 12114162 TI - Teaching residents humanistic skills in a colposcopy clinic. AB - OBJECTIVE: The aim of this project is to add formal training and evaluation of humanistic skills for residents in a colposcopy clinic. Medical education of residents in office procedures has generally focused on proficiency in a particular technical skill and acquisition of clinical knowledge regarding management options. The literature supports assessing pattern recognition with visual images and observing a specific number of procedures to achieve proficiency. Little attention has been given to training residents to demonstrate sensitivity during procedures. However, we believe this is a key element, as the procedure involves anxious patients who are fearful of cancer and fearful of pain in their most private body parts. Further, women who have painful or unpleasant colposcopy procedures are less likely to follow up for further evaluation of treatment and may be at higher risk to develop cervical cancer. DESCRIPTION: Colposcopy training for gynecology residents, family medicine residents, medical students, and emergency room sexual assault nurses is provided at a colposcopy clinic that performs more than 400 procedures per year. The majority of women are referred for colposcopy from outside clinics, and more than one third are non English-speaking. We have added to our traditional didactics and case instruction curriculum a session dedicated to training residents in humanistic skills related to performing colposcopy. Topics of discussion include (1) how to ask sensitive questions regarding risk factors for cervical dysplasia, (2) how to explain the procedure and obtain consent, especially with non-English-speaking women, (3) how to educate patients about human papilloma virus (HPV) and prevention of cervical cancer, (4) how to speak to the patient and the attending physician during the procedure to reduce rather than increase anxiety, (5) how to provide comfort measures during colposcopy (e.g., using a warm speculum, using local anesthesia, hiding instruments from patient view), and (6) how to review findings during and after procedures. The discussion includes errors that previous residents have made and approaches that create effective, culturally sensitive role models. The training session participants are given feedback regarding statements or actions that may make patients anxious or less comfortable during and following clinical procedures. Residents are evaluated at the end of each rotation by faculty who rate their technical skills, visual pattern recognition, knowledge on the subject, and humanistic qualities. DISCUSSION: The training session participants have responded favorably to feedback about observed interactions with patients during procedures. Indications are that this training may improve the quality of care given by providers during colposcopy and other sensitive procedures. To document the curriculum's effect on patient satisfaction and care, we plan to pilot the use of a patient survey regarding physicians' humanistic qualities. PMID- 12114163 TI - A women's health track for internal medicine residents using evidence-based medicine. AB - OBJECTIVES: Evidenced-based medicine has established itself as an integral part of medical education and practice. The explosion of new knowledge in women's health and the need to teach this to internal medicine residents in an evidence based fashion have presented a challenge to medical educators. To address this need, we developed and implemented an evidence-based women's health curriculum to be used in addition to clinical training in a women's health center for internal medicine residents. The objectives of the curriculum are to (1) define and utilize basic evidence-based medicine concepts to critically analyze women's health literature, (2) understand recent innovations in women's health from an evidence-based viewpoint, (3) gain clinical experience in women's health, and (4) apply evidence-based medicine to the clinical practice of women's health. DESCRIPTION: We designed our curriculum based on recommendations from the National Academy of Women's Health Medical Education, the American Board of Internal Medicine, the Fifth Report of the Council on Graduate Medical Education, and the results of needs assessments of internal medicine residents at our institution. Using Medline to create a women's health bibliography, an extensive literature search was performed on the following topics: osteoporosis, breast cancer, hormone replacement therapy, domestic violence, coronary artery disease in women, menopause, headaches, substance abuse in women, urinary incontinence, dementia, sexual dysfunction, and evidence-based medicine. Peer-reviewed journal articles were compiled by subject matter for placement in our clinic's resource center and were entered into a computerized database that will link with online journals and be available for electronic access. Most articles were selected based on the criteria of data published since 1990, and randomized, double blinded, placebo-controlled studies were given preference. Weekly 45-minute sessions preceding the resident clinic in the women's health center are held in a journal-club format to review literature in a systematic fashion. Faculty and residents review and analyze one to two articles weekly. Content experts provide context and clinical expertise to resident discussions. Clinical questions, such as "Should I prescribe hormone replacement therapy to my postmenopausal patient?" are addressed in each session. Evidence-based medicine core concepts are reviewed and applied; these core concepts include the number needed to treat, absolute risk reduction, and relative risk. DISCUSSION: The women's health curriculum, weekly conferences, and clinical experience serve to update residents and clinicians in women's health literature, to exchange ideas for the improvement of women's health as it is taught in internal medicine, and to further elucidate the evidence behind what we practice and teach. The curriculum equips physicians to provide patients with solid, evidence-based interpretations of new scientific knowledge to discern truth from fallacy. PMID- 12114164 TI - Improving residents' interviewing skills by group videotape review. AB - OBJECTIVE: Internal medicine residency training programs typically emphasize biomedical learning, but relatively few provide opportunities for residents to improve outpatient interviewing skills or to address challenging patient encounters. Even fewer programs provide resources to assess patient-resident relationship skills. To address these issues, we developed a curriculum that is designed to enhance patient-centered interviewing techniques in residents. DESCRIPTION: At the UCSF-VA PRIME residency program, interns on ambulatory block attend three didactic sessions that introduce basic medical interviewing techniques, including elicitation of patient concerns and mutual agenda setting, biopsychosocial interviewing, and conflict resolution in the patient-physician relationship. These sessions are taught using role-play exercises and interactive, case-based discussions. Second- and third-year residents on ambulatory block participate in an ongoing small-group seminar in which they present videotapes of their interviews with patients. To make these tapes, residents arrange to see patients in a clinic room where a videotape recorder is mounted on the wall. Patients who consent to be videotaped complete a form stating that the tapes are to be used solely for educational purposes. Twice a year, each of our 16 residents selects one encounter that highlights personal learning goals related to interviewing and presents those learning goals with associated videotape clips in a 90-minute seminar. Each seminar is devoted to two residents, who facilitate a discussion of the effectiveness of the interview and solicit feedback about potential methods for improvement. DISCUSSION: When the seminars were originally developed, we anticipated that this innovative combination of traditional individual videotape review with small-group learning would encourage self-directed learning. Indeed, over the last three years, residents have become more confident with their interviewing capabilities and less self-conscious about showing their own videotaped interviews. As a result, the seminars have unexpectedly evolved into a highly sophisticated series of learning modules, in which residents seek their most challenging patient encounters to videotape and show to the group. Residents have presented complicated scenarios involving critical patient-physician conflicts, somatizing patients, cross-cultural communication difficulties, overzealous family members, patients with substance abuse, and bad-news interviews. These dilemmas represent fundamental management challenges that are difficult to discuss in a more didactic format, and the immediate case-based nature of the interviews makes these often-emotional issues come alive. The group videotape reviews also give residents opportunities to reflect on their own interviewing encounters, to observe other interviewing styles and techniques, and to provide support to their fellow residents after particularly emotional interviews. An ancillary benefit of these exercises is that we have now developed a library of challenging interviews, which are easily accessible for further teaching seminars. Our residents consider this learning experience to be one of the most positive of their residency and valuable for their professional development. Residents report that this small-group seminar series has markedly improved their communication with patients, and they now clamor for the opportunity to present interviewing dilemmas. We believe that similar curricula can be readily instituted at other residency programs. PMID- 12114165 TI - Teaching medical residents about teenagers: an introductory curriculum in adolescent medicine. AB - OBJECTIVE: Adolescents in the United States have high morbidity rates, which are attributable to injuries, behavioral disorders, sexually transmitted diseases, and unplanned pregnancies. This has led to a call to action for health care educators to better prepare future practitioners to meet adolescent health care needs. Although pediatrics residency programs have required one-month curricula dedicated to adolescent medicine, many internal medicine (IM) residency programs do not have such requirements despite an American College of Physicians position paper recognizing the importance of internists' providing health care to adolescents. Thus, an introductory curriculum in adolescent medicine was developed for a community hospital IM residency program. The curriculum was designed to train IM residents to effectively interview, provide preventive care for, and evaluate common medical problems of older adolescents (ages 16-21) in an outpatient setting. DESCRIPTION: The curriculum was based on the results of an extensive needs assessment, which included surveys sent to practicing internists and current IM residents. It included three units, each a series of one-hour lessons delivered via morning report or grand rounds sessions. The first unit was interviewing, which covered specific interviewing questions, communication methods, and confidentiality issues. The second unit was preventive medicine care, which addressed immunizations, injury prevention, and adolescent drug and alcohol use. The third unit was diagnosis and management of common problems, which focused on topics frequently encountered during adolescent outpatient visits, including acne, sexually transmitted diseases, reproductive health care needs, and menstrual abnormalities. Other topics pertinent to the care of adolescents, notably depression, eating disorders, and sports medicine, were covered by previously established curricula, and thus omitted. This curriculum was created through the support of the Michigan State University Primary Care Faculty Development Fellowship Program, and was reviewed by curricular and adolescent medicine content experts. The residency program director supported its implementation over the course of two academic years. The pilot unit interviewing was conducted in February 2001, during two one-hour morning report sessions. In the first session, a short lecture outlining interviewing concepts and principles was given, followed by several case-based studies describing office presentations of adolescent patients. In the second session, an actual office visit was simulated; the residents observed adolescent volunteers being interviewed, and were then given an opportunity to ask the patients further questions. Residents completed a brief quiz and evaluation survey at the end of the pilot unit. DISCUSSION: Twenty-one of 40 residents participated in the pilot unit. Evaluations were overall very positive. Quiz scores confirmed that the residents achieved the desired learning objectives. Given these results from the pilot unit, the remaining two units of the curriculum have been integrated into the residency curriculum. Additional faculty members have been selected to deliver future sessions and support this important educational activity. The adolescent medicine curriculum can be used as an model by other IM residency programs for teaching adolescent health care, with an emphasis on both the knowledge base of adolescent issues and awareness of the unique skills necessary for the establishment of a physician-patient relationship between internists and older adolescents. PMID- 12114166 TI - Implementing a medicine-spirituality curriculum in a community-based internal medicine residency program. AB - OBJECTIVE: To promote greater sensitivity to and heightened awareness of the relevance and therapeutic potential of integrating medicine and spirituality in the healing process of patients cared for by our medical residents. Strategies for clear, effective, and empathetic communication are integrated into the curriculum. DESCRIPTION: With the support of The University of Massachusetts Medical School Macy Initiative in health communication, funded by the Josiah Macy, Jr. Foundation, we have fully implemented a medicine-spirituality curriculum as an integral aspect of our residency program. Current strategies include (1) new house officers participate in the workshop "Communicating Bad News," which is based on a videotaped interaction and experiential role-play about the challenging "art" of sharing bad and often traumatic news; (2) a monthly lecture series that looks at various aspects of religious and spiritual practices and their implications on science and health with topics including the following: taking a spiritual history, exploring world religious views from a Judeo-Christian perspective, studying Eastern philosophies such as Buddhism and Hinduism, and discussing cultural diversity's effect on how people understand and cope with illness; (3) residents receive a comprehensive, evidence-based syllabus that encompasses all of the medical literature relating to spirituality, religion and health; (4) local hospice professionals give end-of-life care lectures about pain management, palliation, advanced directives, and ethical implications; (5) our residents spend one or two days per year with our pastoral care leaders and one to two days per year with our hospice team; (6) monthly ward rounds with a faculty member who emphasizes the spiritual dimension of a particular case and the faith-based resources in our hospital and community. DISCUSSION: Traditionally, graduate medical education has not emphasized the importance of spirituality as a "target" for routine inquiry, understanding, and sharing in the context of patient care. We are beginning to see that residents need to be aware of the relationship between spirituality and health, as a consequence of this curriculum. Because the curriculum is seamlessly integrated into a preexisting infrastructure (e.g., noon conferences, ambulatory off-site experiences, walk rounds, etc.), it has been relatively easy to implement. Focusing on the literature has also provided a "scientific door" that has made this more palatable. Over time, we will foster a growing alliance of the medical and faith communities in our rural area. This has potent implications for community health initiatives. Two of our residents have already volunteered to give talks at local congregations. Spirituality and religion are sensitive and personal areas that can be awkward to embrace and openly discuss. By remaining sensitive and respectful of all views, we strive to diminish the obstacles and enable a more provocative, enlightening residency experience. As a consequence, we are forced to reconsider what it is to be a "healer" and what it is to be "healed." Annual verbal and written feedback will allow us to refine our curriculum. I anticipate this to be a permanent aspect of our residents' training. PMID- 12114167 TI - A communication assessment and skill-building exercise (CASE) for first-year residents. AB - OBJECTIVE: Good communication skills are essential for residents entering postgraduate education programs. However, these skills vary widely among medical school graduates. This pilot program was designed to create opportunities for (1) teaching essential interviewing and communication skills to trainees at the beginning of residency, (2) assessing resident skills and confidence with specific types of interview situations, (3) developing faculty teaching and assessment skills, (4) encouraging collegial interaction between faculty and new trainees, and (5) guiding residency curricular development. DESCRIPTION: During residency orientation, all first-year internal medicine residents (n = 26) at the University of Minnesota participated in the communication assessment and skill building exercise (CASE). CASE consisted of four ten-minute stations in which residents demonstrated their communication skills in encounters with standardized patients (SPs) while faculty members observed for specific skills. Faculty and SPs were oriented to the educational purposes and goals of their stations, and received instructions on methods of providing feedback to residents. With each station, residents were provided one and a half minutes of direct feedback by the faculty observer and the SP. The residents were asked to deal with an angry family member, to counsel for smoking cessation, to set a patient-encounter agenda, and to deliver bad news. A resident's performance was analyzed for each station, and individual profiles were created. All residents and faculty completed evaluations of the exercise, assessing the benefits and areas for improvement. DISCUSSION: Evaluations and feedback from residents and faculty showed that most of our objectives were accomplished. Residents reported learning important skills, receiving valuable feedback, and increasing their confidence in dealing with certain types of stressful communication situations in residency. The activity was also perceived as an excellent way to meet and interact with faculty. Evaluators found the experience rewarding, an effective method for assessing and teaching clinical skills, a faculty development experience for themselves in learning about structured practical skills exercises, and a good way to meet new interns. The residency program director found individual resident performance profiles valuable for identifying learning issues and for guiding curricular development. Time constraints were the most frequently cited area for improvement. The exercise became feasible by collaborating with the medical school Office of Education-Educational Development and Research, whose mission is to collaborate with faculty across the continuum of medical education to improve the quality of instruction and evaluation. The residency program saved considerable time, effort, and expense by using portions of the medical school's existing student skills-assessment programs and by using chief residents and faculty as evaluators. We plan to use CASE next year with a wider variety of physician-patient scenarios for interns, and to expand the program to include beginning second- and third-year residents. Also, since this type of exercise creates powerful feedback and assessment opportunities for instructors and course directors, and because feedback was so favorable from evaluators, we will encourage participation in CASE as part of our faculty educational development program. PMID- 12114168 TI - Training community-responsive physicians. AB - OBJECTIVE: The "community-responsive" primary care provider has a population health perspective and is prevention-oriented, culturally competent, collaborative, and an active community leader and patient advocate. To encourage residents to value this level of community involvement and possess the requisite knowledge and skills, St. Luke's family practice residency program has developed a longitudinal community medicine curriculum designed to teach the four domains of physician-community involvement: (1) insight into sociocultural aspects of patient care, (2) familiarity with community health resources, (3) community oriented primary care skills, and (4) community involvement.(1) Training physicians with the desire and skills to practice in medically underserved communities is a program goal. DESCRIPTION: The three-year community medicine curriculum begins during residency orientation with a windshield survey of the communities served by the campus-related clinics. During the first year, all residents participate in a four-week community medicine rotation. By providing health education and clinical services to diverse populations in community clinics, agencies, and schools, they begin to develop community health improvement skills, while observing role models, developing advocacy skills, expanding cultural awareness, and experiencing interdisciplinary collaboration. At the end of the first year, residents select a community clinical site, where they will see patients and develop a community health-improvement project during the last two years of training. A required "capstone presentation" describes the scope and nature of each resident's project by focusing on process and outcome measures. Innovative qualitative evaluation tools include a written portfolio of reflections and sequential "video journaling." An attribute-based progression matrix developed by Alverno College was adapted to facilitate serial identification and tracking of resident growth in eight domains: communication, analysis, problem solving, aesthetic responsiveness, global perspectives, valuing in decision making, social interaction, and effective citizenship.(2) Support for this program is provided by HRSA, Wisconsin AHEC, Aurora Health Care, and the medical school. DISCUSSION: A recent program graduate, now a faculty member, demonstrated the potential for this educational strategy by developing a local "Reach Out and Read" program. Targeting literacy as a factor related to the cycle of poverty and poor health, she implemented an intervention in which residents read with their pediatric patients during each visit and give the child a book to take home. As each residency class implements health-promoting interventions, this longitudinal community medicine residency curriculum will improve community health through "service learning," as well as develop a cadre of young physicians who practice community-responsive clinical medicine and have the skills and confidence to choose to serve underserved populations. PMID- 12114169 TI - E-conferencing for delivery of residency didactics. AB - OBJECTIVE: While didactic conferences are an important component of residency training, delivering them efficiently is a challenge for many programs, especially when residents are located in multiple sites, as they are at Wayne State University School of Medicine in the Department of Family Medicine. Our residents find it difficult to travel from our hospitals or rotation sites to a centralized location for conferences. In order to overcome this barrier, we implemented distance learning and electronically delivered the conferences to the residents. DESCRIPTION: We introduced an Internet-delivered, group-learning interactive conference model in which the lecturer is in one location with a group of residents and additional residents are in multiple locations. We launched the project in July 2001 using external company meeting services to schedule, coordinate, support, and archive the conferences. Equipment needed in each location consisted of a computer with an Internet connection, a telephone line, and a LCD projector (a computer monitor sufficed for small groups). We purposely chose simple distance-learning technology and used widely available equipment. Our e-conferencing had two components: (1) audio transmission via telephone connection and (2) visual transmission of PowerPoint presentations via the Internet. The telephone connection was open to all users, allowing residents to ask questions or make comments. Residents chose a conference location depending on geographic proximity to their rotation locations. Although we could accommodate up to 50 sites, we focused on a small number of locations in order to facilitate interaction among residents and faculty. Each conference session is archived and stored on the server for one week so those residents whose other residency-related responsibilities precluded attendance can view any conferences they have missed. DISCUSSION: E-conferencing proved to be an effective method of delivering didactics in our residency program. Its many advantages included ease of use, cost-efficiency, and wide availability of equipment. Residents had the advantage of both geographic and temporal independence. Our e-conferences were interactive, and in addition to a PowerPoint presentation, faculty provided Web sites and hyperlinks for references. Initial problems included slow-speed connection, the requirement for digital materials, and the need for residents and faculty to adjust to a new learning method. There was also a need for increased coordination at the sites and reliance on electronic communication. To assess the effectiveness of the program, residents completed knowledge pre- and post-tests and a conference evaluation form. We also monitored conference attendance rates. Preliminary results indicated positive resident attitudes toward distance learning and significant increases in conference attendance. To objectively evaluate this instructional delivery method, we will compare residents' knowledge gains in the face-to-face instructor group with those of the group to which the lecture is broadcast. Ultimately, we are hoping to offer this educational opportunity to other family practice residency programs in the area, to medical students interested in family medicine, and to community family physicians for continuing medical education. We are considering the addition of streaming video to the presentations in the future, once the bandwidth of the Internet connections is sufficient. PMID- 12114170 TI - Pediatric residents as learners and teachers of evidence-based medicine. AB - OBJECTIVES: Residents must learn to assess the medical literature and apply it clinically. We designed and implemented a curriculum to support resident acquisition and use of skills required for critical review and clinical application of evidence from the pediatrics literature. The experience provided an opportunity for residents to observe, demonstrate under supervision, and practice evidence-based skills using the "see one, do one, practice many" approach. DESCRIPTION: We revised our residents' evidence-based pediatrics journal club curriculum to provide specific learning objectives to be accomplished by residents during the three years of residency. Curriculum objectives address study design, definition of the research question, characterization of variable type and choices of statistical methods, attention to sample size, diagnostic test characteristics, identification of sources of bias, and generalization and specific application of research findings to our clinical setting. We implemented a three-stage approach to skills learning. First year residents participate in a monthly journal-club presented by the second-year residents ("see one"). During the second year, residents present noon conferences based on the curriculum objectives. The second-year resident meets with a faculty member to select a journal article for discussion, identify key curriculum objectives illustrated by the study, and choose pertinent clinical epidemiology references. The resulting presentation is given as a noon conference to an audience of residents, students, and faculty ("do one"). During the remainder of residency, residents continue to attend a monthly journal club that equips them with new skills and allows them to take a more meaningful part in the discussions ("practice many"). Curriculum faculty attend the monthly journal club to help direct the discussion of the article to fulfill curriculum objectives. DISCUSSION: Skills are best acquired in an environment which promotes active learning supervised by experts and provides frequent opportunities to practice the skills. Residents have responded positively to our curriculum and have presented high-quality conferences. Evaluation data being collected now (resident self-assessments and evaluations of the experience, faculty assessments of presentations, pre- and post-second year written assessments of knowledge) will inform us if the desired outcomes are being attained. Over the first two years of implementation of this curriculum, we have observed that it takes no more faculty time to supervise resident preparation and presentation than it would for the faculty to prepare and present material concerning similar curriculum objectives in a lecture or conference format. Additional benefits for residents include creating materials for presentation with experienced faculty, making presentations for peer groups, and assuming the role of teacher. PMID- 12114171 TI - Development of an office-based curriculum of common pediatric primary care skills for residents. AB - OBJECTIVE: Basic primary care skills-such as injections and hearing screening-are commonly absent from residency curricula, yet competence in these skills is required by residency accrediting organizations. To meet this need at our program, an office-based curriculum of common pediatric primary care skills was developed and piloted in a resident continuity practice. DESCRIPTION: Based on a needs assessment, an eight-unit curriculum was developed to teach residents basic ambulatory primary care skills. The program was designed as a skills supplement to existing primary care curricula and includes hearing screening, vision screening, developmental screening, injections, venipuncture, urinalysis, in office rapid testing, and analysis of skin scrapings. Each unit lasted one month, using a "skill of the month" format during continuity clinic. During the month, residents received instruction and demonstration, practiced the skill, and tested to document competence. A pilot of the unit "Intramuscular, Subcutaneous, and Intradermal Injections," was conducted at the Pediatric Primary Care Center of Cincinnati Children's Hospital with 26 pediatrics residents. Fourteen residents participated in the pilot prior to testing and a comparison group of 12 was tested without the pilot experience. The pilot occurred over two weeks. The first week, a 20-minute training session was held at the beginning of continuity clinic to discuss a procedure checklist for injections and allow residents to practice with a mannequin. Throughout the remainder of the continuity clinic during the pilot, residents administered injections to their patients, following the procedure checklist and under supervision by medical assistants. At the conclusion of the second week, residents were evaluated with a written test and a practicum. The 12 residents in the control group were tested identically. DISCUSSION: This pilot demonstrated that it is feasible to teach primary care skills to residents in the office setting. In our pilot, the test group performed 61% better on a written test and 64% better on a practical test when compared with the control group (p <.0001). Residents who participated in the pilot felt the methods used were appropriate and effective and that the skills taught were important. Additionally, they found the pilot did not interfere with the operation of the continuity clinic. The procedure checklist proved to be an effective and simple method of instructing a psychomotor skill. Conducting the educational sessions at the beginning of clinic was difficult due to interruptions and tardiness. While other methods, such as noon conferences, may also be effective, instruction in the actual clinical setting appeared to better demonstrate the importance, practicality, and relevance of the skill. The residents were more enthusiastic during this office-based curriculum than a typical resident conference. We conclude that this model is an effective and practical method to teach primary care skills in a clinical setting. Our success with the pilot unit has been encouraging, and we plan to develop and test the remaining units of the curriculum. PMID- 12114172 TI - Colonoscopy curriculum development and performance-based assessment criteria on a computer-based endoscopy simulator. AB - OBJECTIVE: Computer-based colonoscopy simulators (CBCS) have been developed and are being introduced into the training environment. The ability of these simulators to replicate the dimensions of patient-based diagnostic colonoscopy is good. However, the benefit of simulators to either learners or their patients has not yet been established. We describe a process by which a CBCS curriculum and CBCS-based performance criteria were established for first-year gastroenterology fellows at the Mayo Clinic in Rochester, Minnesota. DESCRIPTION: We used a commercially available CBCS (AccuTouch Endoscopy Simulator, Immersion Medical, Gaithersburg, MD), which consists of a specialized colonoscope that is inserted into a computer-based module with a screen showing the colonic lumen of a virtual patient. A tutorial and six cases of varying complexity are available on the CBCS. Performance variables that are measured by the simulator include the time to complete the procedure, the distance that the scope was advanced, the degree to which the mucosa was adequately visualized, the possible complications such as colonic perforation, and the level of pain experienced by the simulated patient. To begin, we established ideal performance standards by measuring the above variables for ten "expert" faculty colonscopists who completed two cases on the CBCS. Next, we measured CBCS performance standards for five partially trained colonoscopists. Finally, two non-physician gastrointestinal assistants, without prior endoscopic training, were asked to practice on the simulator to determine the time and procedure frequency required to improve their CBCS proficiency. By calculating average performance standards within each of these three groups, we were able to estimate the number of CBCS cases and minimal performance standards for new trainees. Based on the learning curves for novice colonoscopists as well as the performances of partially trained and expert colonoscopists, we speculated that if CBCS training were to be beneficial, the benefit would most likely occur at the early stages of training. The curriculum we developed consists of viewing a one-hour, multimedia tutorial, which describes the procedure and various colonoscopy techniques. This is followed by nine hours of hands-on CBCS experience, during which time the trainee will complete approximately 25 CBCS colonoscopies. Before advancing to live-patient colonoscopies, the trainee must meet certain performance standards on specific CBCS cases. These standards include the ability to view the entire colon in less than 15 minutes with minimal pain and no complications. DISCUSSION: If effective, this new colonoscopy training curriculum should result in improved competency at patient-based colonoscopy, particularly in the early stages of training. To address this question, first-year gastroenterology fellows at Mayo Clinic have been randomized into two groups: (1) a group that experiences a tutorial with hands-on CBCS curriculum and (2) a group that experiences a tutorial only. Their performances at patient-based colonoscopy as well as surveys of patient satisfaction will be measured and analyzed to determine what, if any, benefit is provided by CBCS. PMID- 12114173 TI - Assessing the needs of residency program directors to meet the ACGME general competencies. AB - OBJECTIVE: New accreditation requirements for residency training programs require residents to have educational experiences that allow them to demonstrate competency in the following areas: (1) patient care, (2) medical knowledge, (3) practice-based learning and improvement, (4) interpersonal and communication skills, (5) professionalism, and (6) systems-based practice. Residents' competence must be assessed with dependable measures. Residency training program directors (PDs) need assistance in complying with these new requirements. DESCRIPTION: Using a survey modified from Michigan State University, we asked PDs to rate their current understanding of and preparation for the general competencies and to provide written comments. PDs of the 47 ACGME-accredited programs received e-mailed instructions to complete the Web-based survey. Twenty four PDs (51%) complied by the deadline. The mean ratings were calculated from a five-point scale (1 = strongly disagree, major impediment or least useful, 5 = strongly agree, not an impediment, or most useful). PDs felt they were informed (3.45) and understood (3.67) the requirements, but they were not well prepared to meet them (2.95). The perceived impediments to implementation included amount of PD time (2.27), amount of residents' protected time for the curriculum (2.30), amount of residency support staff (2.73), lack of expertise in curriculum development (2.73) and evaluation (2.41), and lack of funding for resources other than personnel (2.91). PDs rated types of assistance that would be helpful: developing workshops or presentations on curriculum development and evaluation techniques (3.82), developing curricula (4.14), providing one-on-one consultation (4.23), receiving examples of materials, methods, and ideas from other programs (4.41), and describing evaluation methods/instruments (4.50). Written comments stated that time to concentrate on the topic, release time from clinical responsibilities, and technical computer support would be helpful. Of the six competency areas, PDs were most interested in receiving assistance in developing curricular materials for the competencies of systems-based practice (4.50), professionalism (4.36), and practice-based learning and improvement (4.27). PDs were most interested in receiving assistance in developing evaluations for practice-based learning and improvement (4.59), professionalism (4.59), interpersonal and communication skills (4.45), and systems-based practice (4.36). PDs responded that they currently use written faculty evaluations to assess all six general competency areas. DISCUSSION: Results of the survey indicate that PDs require assistance to comply with the new ACGME requirements. Curricular materials and valid and reliable evaluation methods need to be developed. In order to assist PDs, the following activities are under way: (1) PDs are members of a listserve for sharing ideas and examples of curricular and evaluation materials; (2) PDs attend a monthly seminar series that provides practical information for curricular material development and specific evaluation methods, including indications for use and feasibility; (3) educators from our Office of Educational Development provide individual consultations with each PD; (4) PDs participate in an eight hour workshop with practical sessions for developing curricular materials and evaluations; and (5) two institution-wide assessments are being developed: a patient-satisfaction survey and a 360-degree evaluation to assess communication skills and professionalism. PMID- 12114174 TI - Education on-demand: the development of a simulator-based medical education service. AB - OBJECTIVES: Clinical medical education depends on the availability of instructive patient encounters, or "good teaching cases." While all medical students hope to see enough patients of sufficient scope and variety, exposure to good teaching cases has been traditionally limited by time and chance. Students may graduate from medical school without having seen a number of important cases, each of which may represent a knowledge gap they will carry forward into internship and future patient care. Recently, however, the advent of high-fidelity patient simulators has enabled instructors to recreate realistic patient scenarios in a standardized fashion. Using the simulator, we wanted to create a medical education service-like any other clinical teaching service, but designed exclusively to help students fill in the gaps in their own education, on demand. We hoped to mitigate the inherent variability of standard clinical teaching, and to augment areas of deficiency. DESCRIPTION: Using a Human Patient Simulator(TM) (Medical Education Technologies, Inc.), which is equipped with a transmitted voice and reactive eyes, chest movements and breath sounds, heart tones and palpable pulses, a multidisciplinary group of physicians and educators designed a simulator-based medical education service. The premise was that students should have the ability to realistically practice and discuss medicine with a physician mentor at any time they wish, with full access to simulator-enabled cardiac monitoring, diagnostic resources, pharmacologic agents, and invasive procedures. Students were informed of the program by e-mail and by course instructors. A dedicated pager was established for the on-call physician-educator, and the number disseminated to students. Physician-educators included faculty members scheduled for dedicated teaching time, and senior residents participating in a medical education elective. On-call physicians were responsible for fielding educational requests, developing appropriate scenarios, and scheduling instructional time. Upon arriving at the skills lab for their appointments, students would proceed to interview, evaluate, and treat the mannequin-simulator as if it were a real patient, using the instructor for assistance as needed. All students participated in an educational debriefing after each session. Instructors could also request formal observation and feedback on their teaching style, in collaboration with an existing faculty development program. DISCUSSION: Students enjoy the opportunity to practice medicine on-demand with dedicated clinical mentoring by a practicing physician. Course directors are interested in scheduling simulator time to help bring to life tutorial-based teaching cases and other course material for their students. By offering a medical education elective for residents, we have bolstered the pool of available instructors, provided a valuable learning experience for residents as teachers, and fostered additional opportunities for collaboration between the medical school and clinical training sites. Customized, realistic clinical correlates are now readily available for students and teachers, allowing reliable access to "the good teaching case." PMID- 12114175 TI - A real-time computer model to assess resident work-hours scenarios. AB - OBJECTIVE: To accurately model residents' work hours and assess options to forthrightly meet Residency Review Committee-Internal Medicine (RRC-IM) requirements. DESCRIPTION: The requirements limiting residents' work hours are clearly defined by the Accreditation Council for Graduate Medical Education (ACGME) and the RRC-IM: "When averaged over any four-week rotation or assignment, residents must not spend more than 80 hours per week in patient care duties."(1) The call for the profession to realistically address work-hours violations is of paramount importance.(2) Unfortunately, work hours are hard to calculate. We developed an electronic model of residents' work-hours scenarios using Microsoft Excel 97. This model allows the input of multiple parameters (i.e., call frequency, call position, days off, short-call, weeks per rotation, outpatient weeks, clinic day of the week, additional time due to clinic) and start and stop times for post-call, non-call, short-call, and weekend days. For each resident on a rotation, the model graphically demonstrates call schedules, plots clinic days, and portrays all possible and preferred days off. We tested the model for accuracy in several scenarios. For example, the model predicted average work hours of 85.1 hours per week for fourth-night-call rotations. This was compared with logs of actual work hours of 84.6 hours per week. Model accuracy for this scenario was 99.4% (95% CI 96.2%-100%). The model prospectively predicted work hours of 89.9 hours/week in the cardiac intensive care unit (CCU). Subsequent surveys found mean CCU work hours of 88, 1 hours per week. Model accuracy for this scenario was 98% (95% CI 93.2-100%). Thus validated, we then used the model to test proposed scenarios for complying with RRC-IM limits. The flexibility of the model allowed demonstration of the full range of work-hours scenarios in every rotation of our 36-month program. Demonstrations of status-quo work-hours scenarios were presented to faculty as well as real-time demonstrations of the feasibility, or unfeasibility, of their proposed solutions. The model clearly demonstrated that non-call (i.e., short-call) admissions without concomitant decreases in overnight call frequency resulted in substantial increases in total work hours. Attempts to "get the resident out" an hour or two earlier each day had negligible effects on total hours and were unrealistic paper solutions. For fourth-night-call rotations, the addition of a "golden weekend" (i.e., a fifth day off per month) was found to significantly reduce work hours. The electronic model allowed the development of creative schedules for previously third-night call rotations that limit resident work hours without decreasing continuity of care by scheduling overnight call every sixth night alternating with sixth-night short-call rotations. DISCUSSION: Our electronic model is sufficiently robust to accurately estimate work hours on multiple and varied rotations. This model clearly demonstrates that it is very difficult to meet the RRC-IM work-hours limitations under standard fourth-night-call schedules with only four days off per month. We are successfully using our model to test proposed alternative scenarios, to overcome faculty misconceptions about resident work-hours "solutions," and to make changes to our call schedules that both are realistic for residents to accomplish and truly diminish total resident work hours toward the requirements of the RRC-IM. PMID- 12114176 TI - Interactive computer simulations of knee-replacement surgery. AB - OBJECTIVE: Current surgical training programs in the United States are based on an apprenticeship model. This model is outdated because it does not provide conceptual scaffolding, promote collaborative learning, or offer constructive reinforcement. Our objective was to create a more useful approach by preparing students and residents for operative cases using interactive computer simulations of surgery. Total-knee-replacement surgery (TKR) is an ideal procedure to model on the computer because there is a systematic protocol for the procedure. Also, this protocol is difficult to learn by the apprenticeship model because of the multiple instruments that must be used in a specific order. We designed an interactive computer tutorial to teach medical students and residents how to perform knee-replacement surgery. We also aimed to reinforce the specific protocol of the operative procedure. Our final goal was to provide immediate, constructive feedback. DESCRIPTION: We created a computer tutorial by generating three-dimensional wire-frame models of the surgical instruments. Next, we applied a surface to the wire-frame models using three-dimensional modeling. Finally, the three-dimensional models were animated to simulate the motions of an actual TKR. The tutorial is a step-by-step tutorial that teaches and tests the correct sequence of steps in a TKR. The student or resident must select the correct instruments in the correct order. The learner is encouraged to learn the stepwise surgical protocol through repetitive use of the computer simulation. Constructive feedback is acquired through a grading system, which rates the student's or resident's ability to perform the task in the correct order. The grading system also accounts for the time required to perform the simulated procedure. We evaluated the efficacy of this teaching technique by testing medical students who learned by the computer simulation and those who learned by reading the surgical protocol manual. Both groups then performed TKR on manufactured bone models using real instruments. Their technique was graded with the standard protocol. The students who learned on the computer simulation performed the task in a shorter time and with fewer errors than the control group. They were also more engaged in the learning process. DISCUSSION: Surgical training programs generally lack a consistent approach to preoperative education related to surgical procedures. This interactive computer tutorial has allowed us to make a quantum leap in medical student and resident teaching in our orthopedic department because the students actually participate in the entire process. Our technique provides a linear, sequential method of skill acquisition and direct feedback, which is ideally suited for learning stepwise surgical protocols. Since our initial evaluation has shown the efficacy of this program, we have implemented this teaching tool into our orthopedic curriculum. Our plans for future work with this simulator include modeling procedures involving other anatomic areas of interest, such as the hip and shoulder. PMID- 12114177 TI - Teaching and testing physical examination skills without the use of patients. AB - OBJECTIVE: To design a cardiopulmonary physical exam curriculum that does not involve the use of patients. Bedside teaching is becoming a lost art, and the use of alternative methods of instruction such as simulation has become increasingly important. Simulators have been shown to enhance physical examination skills of students and physicians in training.(1) DESCRIPTION: In 1995, a program was started to improve cardiopulmonary physical diagnosis and the teaching of auscultation at the University of Texas Medical Branch at Galveston (UTMB). The teaching manikin "Harvey" played a vital role in the development of the new curriculum. In 1997, UTMB adopted an organ-based approach to the basic science curriculum. The cardiopulmonary module in the basic science curriculum was a ten week course taught in the second year of medical school. The physical diagnosis section of that course involved six instructional hours; four of the six hours were dedicated to cardiac auscultation and two hours to pulmonary auscultation. Only simulators and CD-ROMs were used for instruction. The 184 second-year medical students at UTMB were formed into small groups for instruction and practice. Although "Harvey" was an effective teaching tool, other simulators had to be developed for testing students' skills after instruction. It would be very difficult to administer a skills OSCE for 184 students without the development of several smaller transportable simulators. A commercially available blood pressure simulator from the Medical Plastics Laboratory, Inc., Gatesville, TX, was used to test the accuracy of students' blood pressure readings. Small auscultation transducers combined with a palpable pulse simulator, developed by one of the authors (WT) in collaboration with Andries Acoustics, Spicewood, TX, were used to efficiently test students' proficiency in cardiopulmonary auscultation. Digital simulated cardiopulmonary sounds were recorded onto a standard CD-ROM mini-disc and transmitted to the small transducers. Students used their own stethoscopes for auscultation. The targeted skills were efficiently tested in one hour of testing time per student. DISCUSSION: This cardiopulmonary instructional module was well received by the second-year medical students. In the skills OSCE, 80% of the students accurately measured systolic and diastolic blood pressure to within 5 mm Hg. Cardiopulmonary auscultation proficiency results showed average recognition of 60% for cardiac abnormalities and 88% for pulmonary sounds. Developing auscultation transducers with pulse simulation capability ensured that students could identify systole. Therefore, heart murmurs and sounds could be timed with the cardiac cycle. We found the results from the skills OSCE encouraging. Most students demonstrated reasonable competency in the skills taught, and the new transportable simulators performed well. The six-hour instructional module was meant to prepare students for their bedside teaching during the third year of medical school. The significant cost of the "Harvey" simulator may be a barrier to its widespread use for teaching. Therefore, continued development of smaller transportable simulators for teaching and testing purposes is important. PMID- 12114178 TI - Incorporating simulators in a standardized patient exam. AB - OBJECTIVE: Using simulated patients during a clinical skills exam that involves many students has the advantage of standardizing the delivery of historical data. One major disadvantage is the inability to standardize the physical exam findings. We designed a simulated patient exam that incorporates simulated abnormal physical exam findings. DESCRIPTION: The simulated patient exam case was divided into three separate stations: (1) the simulated patient's history, (2) the simulated physical exam, and (3) the presentation station. Dyspnea was chosen as the chief complaint because of the broad differential of possible cardiac and pulmonary auscultatory findings. In the first station, students obtained historical data from the standardized simulated patient. Students were graded on their ability to ask appropriate historical questions. Trained observers were used to verify the numbers of historical cues obtained by the students. The second station consisted of simulated physical exam findings. Students first measured the blood pressure on a commercially available blood pressure simulator arm from the Medical Plastics Laboratory, Inc., Gatesville, TX. Students then auscultated an abnormal digital heart sound and pulmonary sound from a small auscultation transducer developed by Andries Acoustics, Spicewood, TX. Students also palpated a simulated pulse from a newly developed pulse transducer. Digital cardiopulmonary sounds and pulse data were recorded onto a CD-ROM disc and transmitted to the small transducers via a CD-ROM disc player. Students used their own stethoscopes to auscultate cardiopulmonary sounds from the small transducers. The students were graded in the second station on their ability to accurately measure a blood pressure, identify abnormal cardiopulmonary digital sounds, and finally describe a peripheral pulse. In the third station, students presented the historical data and physical exam findings to a faculty member, and then provided a differential diagnosis list based on their key findings from the other two stations. A total of 171 students (n = 171) completed the simulated patient exam. Each student completed the exam in 45 minutes. DISCUSSION: In our simulated patient exam, students were evaluated not only on their data-gathering skills for key historical findings but also on the ability to correctly identify key physical exam findings such as abnormal cardiopulmonary sounds. Key physical exam findings were then integrated into the clinical decision-making process, which was presented in the faculty presentation station. Simulated patients with abnormal cardiopulmonary findings can be used for testing purposes. However, cardiac auscultatory abnormalities such as the ventricular S3 gallop are difficult to find and usually occur in a decompensated state such as heart failure. Other physical exam findings such as pulmonary crackles and wheezes also occur in decompensated conditions. Therefore, the use of simulators during a simulated patient exam offers the possibility of introducing several abnormal physical exam findings without having an unstable patient present in an exam setting. Further, the use of simulated physical exam findings allows for complete standardization of a clinical-simulated patient exam. PMID- 12114179 TI - An innovative partnership in service. AB - OBJECTIVE: Stimulated by the need for better alignment of educational content and goals with evolving societal needs, practice patterns, and scientific developments, many medical schools are implementing new and creative educational experiences for students. Tulane University School of Medicine and Apple Computers have established an innovative partnership in which Apple laptop computers support and enhance students' service learning projects. The partnership also provides a unique opportunity to meet the Medical School Objectives Project (MSOP) objectives in Medical Informatics and Population Health, as outlined in Report II.(1) DESCRIPTION: Apple Computers has a commitment to the New Orleans community as part of its corporate strategic plan to support educational programs at all levels; Tulane has a longstanding commitment to and experience with student-led service learning as part of the Foundations in Medicine Course.(2) Senior administrative personnel from Tulane and Apple discussed these common interests, resulting in a partnership to enhance the potential impact on the community served. Apple agreed to donate 20 G3 Powerbooks and a complete set of the Apple Learning series of software to support new and ongoing service-learning projects. A committee of Tulane faculty and students, information technology staff, and an Apple representative developed the project. To maximize students' access to the laptops while managing the administration's liability, the laptops were identically configured with standardized software packages (database development and maintenance, Web access, word processing, presentation development and execution, automated backup, and individual project access to protected server space). To maximize the use of the laptops, students from the service-learning organizations can check out the laptops on a just-in-time basis, because the projects have different needs over time. Student-service leaders are currently defining and developing the exact uses for the laptops. We anticipate that this project will enhance the administrative management of service-learning programs (e.g., schedules, directions to sites), the presentation of educational programs (e.g., teaching in schools), the creation of new media to support programs (e.g., our restaurant choking program has a partnership with the American Heart Association to create a video and training manual to be used nationwide), and data tracking (e.g., sites and clients served, outcomes achieved). Students' use of the laptops should support the achievement of several of the MSOP Report II Medical Informatics objectives. To assess that, all first-year medical students are completing a pre- and post-project survey based on those objectives. DISCUSSION: The availability of laptops and software should significantly enhance the service-learning programs. The students participating should gain important skills in the use of computer technology related to their roles as lifelong learners, educators and communicators, researchers, and managers.(1) We plan to report the results of the pre- and post-project surveys once they have been completed. Students' feedback on the project has been very positive, and we hope it can serve as a model for other medical school, corporate, and community partnerships. PMID- 12114182 TI - The promise of translational physiology. PMID- 12114180 TI - Electronic procedure logs: taking it further. AB - OBJECTIVE: To create an electronic procedural logbook with enhanced user interactivity and usefulness as an educational resource. DESCRIPTION: From our own work on electronic student logbooks and other studies, it is clear that compliance with complete and valid data entry remains a challenge. Without direct and visible benefit, students are reluctant to spend time logging all their cases. We developed an interactive procedural logbook using Microsoft Embedded Visual Basic and Metrowerks CodeWarrior. Interface design focused on rapid data entry with minimum requirement for text, and field-level automation where possible. We loaded it onto a mixed platform of personal digital assistants (PDAs)--Compaq iPaq Pocket PCs and HandEra 330 PalmOS devices. Our rural residents were supplied with the devices of their choice. Various built-in educational reference resources included: (1) contextual help options about each procedure, which contained diagrams and pictures; (2) diagnostic and fee coding so they could see how poorly some procedures pay; and (3) Quick Tips relevant to each procedure, which can be easily modified by the preceptor. Preceptor evaluations and comments can be entered rapidly. Using built-in database conduits, data are automatically collected from each device on every HotSync with the desktop. Data can then be collated and analyzed using Microsoft Access or via secure Web access. DISCUSSION: Improved compliance has been dramatic-one resident logged 250 procedures in just two months. However, not all residents have been successful in establishing seamless synchronization, and the resulting data loss has caused frustration. The evidence indicates the need to implement central data collection and backup right from the outset. Central data collection provides many advantages. The program director has better information for future applications. Preceptor evaluations are now spread over many interactions and yet can be collated and analyzed. Quick Tips have been very popular-we have been able to collect the tips and redistribute them. Focus-group feedback from the residents has shown that the rich data in the logbook's reference component improved its usefulness and popularity as an educational tool. Choice of device type is important for user acceptance because devotees of one platform are reluctant to switch to another. Cross-platform development does slow the process but is increasingly easy with the latest software design tools, such as AppForge. These new tools have enabled us to explore further improvements in data entry. Digital ink provides the ability to capture annotated diagrams and preceptor signatures. Voice input is built in with these devices, and our software now allows for voice annotation for more detailed commentary by preceptors or student. The compressed digital sound file is collected along with the data and transcribed centrally (on-device voice recognition is not feasible yet). Point-of care accessibility has been the key attraction of using these devices for logging encounter data. This project demonstrates that it is important to explore all multimodal interactive capabilities to provide a truly rich educational tool. PMID- 12114183 TI - Oxygen free radicals and heart failure: new insight into an old question. PMID- 12114185 TI - Antioxidant responses to oxidant-mediated lung diseases. AB - Reactive oxygen species (ROS) and reactive nitrogen species (RNS) are generated throughout the human body. Enzymatic and nonenzymatic antioxidants detoxify ROS and RNS and minimize damage to biomolecules. An imbalance between the production of ROS and RNS and antioxidant capacity leads to a state of "oxidative stress" that contributes to the pathogenesis of a number of human diseases by damaging lipids, protein, and DNA. In general, lung diseases are related to inflammatory processes that generate increased ROS and RNS. The susceptibility of the lung to oxidative injury depends largely on its ability to upregulate protective ROS and RNS scavenging systems. Unfortunately, the primary intracellular antioxidants are expressed at low levels in the human lung and are not acutely induced when exposed to oxidative stresses such as cigarette smoke and hyperoxia. However, the response of extracellular antioxidant enzymes, the critical primary defense against exogenous oxidative stress, increases rapidly and in proportion to oxidative stress. In this paper, we review how antioxidants in the lung respond to oxidative stress in several lung diseases and focus on the mechanisms that upregulate extracellular glutathione peroxidase. PMID- 12114184 TI - Induction of oxidative stress and disintegrin metalloproteinase in human heart end-stage failure. AB - Collagen degradation is required for the creation of new integrin binding sites necessary for cell survival. However, a complete separation between the matrix and the cell leads to apoptosis, dilatation, and failure. Previous studies have demonstrated increased metalloproteinase activity in the failing myocardium. To test the hypothesis that disintegrin metalloproteinase (DMP) is induced in human heart end-stage failure, left ventricle tissue from ischemic cardiomyopathic (ICM, n = 10) and dilated cardiomyopathic (DCM, n = 10) human hearts were obtained at the time of orthotopic cardiac transplant. Normal (n = 5) tissue specimens were obtained from unused hearts. The levels of reduced oxygen species (ROS) were 12 +/- 2, 25 +/- 3, and 16 +/- 2 nmol (means +/- SE, P < 0.005) in normal, ICM, and DCM, respectively, by spectrofluorometry. The percent levels of endothelial cells were 100 +/- 15, 35 +/- 19, and 55 +/- 11 in normal, ICM, and DCM, respectively, by CD31 labeling. The levels of nitrotyrosine by Western analysis were significantly increased, and endothelial nitric oxide (NO) by the Griess method was decreased in ICM and DCM compared with normal tissue. The synthesis and degradation of beta(1)-integrin and connexin 43 were significantly increased in ICM and DCM compared with normal hearts by Western analysis. Levels of DMP were increased, and levels of cardiac inhibitor of metalloproteinase (CIMP) were decreased. Aggrecanase activity of DMP was significantly increased in ICM and DCM hearts compared with normal. These results suggest that the occurrence of cardiomyopathy is significantly confounded by the increase in ROS, nitrotyrosine, and DMP activity. This increase is associated with decreased NO, endothelial cell density, and CIMP. In vitro, treatment of CIMP abrogated the DMP activity. The treatment with CIMP may prevent degradation of integrin and connexin and ameliorate heart failure. PMID- 12114186 TI - Maintenance of the mouse type II cell phenotype in vitro. AB - The purpose of this study was to identify culture conditions for maintenance of isolated mouse type II cells with intact surfactant protein (SP) and phospholipid production. Type II cells were isolated from 6-wk-old mice and cultured on Matrigel matrix-rat tail collagen (70:30 vol/vol) in bronchial epithelial cell growth medium minus hydrocortisone plus 5% charcoal-stripped FBS and 10 ng/ml keratinocyte growth factor. Under these conditions, type II cells actively produced surfactant phospholipids and proteins for at least 7 days. Synthesis and secretion of surfactant phospholipids and SP-A, -B, -C, and -D declined on day 1 of culture but recovered by day 3, reaching levels comparable to or exceeding freshly isolated cells by day 5. Abundant lamellar bodies were readily apparent in cells examined on days 5 and 7, and a surfactant pellet was recovered by centrifugation of media harvested on each day of culture. Secretion of SP-B, SP C, and phosphatidylcholine was stimulated by phorbol 12-myristate 13-acetate and was inhibited by compound 48/80. When tested with a bubble surfactometer, surfactant secreted by type II cells on day 5 of culture lowered surface tension to 5.2 +/- 2.3 mN/m. This is the first description of the synthesis and secretion of a functional surfactant complex by mouse type II cells after 7 days in primary culture. PMID- 12114187 TI - Molecular cloning of actin filament-associated protein: a putative adaptor in stretch-induced Src activation. AB - Mechanical stretch-induced activation of c-Src is an important step for signal transduction of stretch-induced fetal rat lung cell proliferation. This process appears to be mediated through actin filament-associated protein (AFAP), encoded by a gene originally cloned from the chicken. In the present study, we cloned the rat AFAP gene from fetal rat lungs. Its mRNA and protein are differentially expressed among various tissues. The protein is colocalized with actin filaments in fetal rat lung epithelial cells and fibroblasts. Mechanical stretch increased tyrosine phosphorylation of rat AFAP and its binding to c-Src within the initial several minutes. Src SH2 and SH3 binding motifs are highly conserved in the AFAP proteins (from chicken, rat to human). On the basis of the molecular structure of AFAP protein, we speculate that it is an adaptor in mechanical stretch-induced activation of c-Src. A novel model of mechanoreception is proposed. PMID- 12114188 TI - Dexamethasone-induced changes in lung function are not prevented by concomitant treatment with retinoic acid. AB - Alveolarization is impaired in rats treated with dexamethasone (Dex) on postnatal days 4-13, but concomitant treatment with all-trans retinoic acid (RA) increases alveolar number. To determine whether morphological changes induced by Dex and/or RA predict changes in lung function at 1 mo, we assessed resting breathing parameters, dynamic compliance, ventilation required to maintain O(2) saturation at > or = 90%, and pressure-volume curves of air-filled lungs. During resting breathing, mean tidal volume per gram was greater in Dex + RA-treated rats than in controls (P < 0.05). Dynamic compliance was also greater in Dex- and Dex + RA treated rats than in controls or RA-treated rats (P < 0.02). In Dex- and Dex + RA treated rats, we observed increased hysteresis ratios (P < or = 0.006), air trapping (P < 0.05), and lung volumes at 5 and 13.5 cmH(2)O pressure (P < 0.001) and decreased elastic recoil (P < 0.007). The effect of Dex on elastic recoil was greater in female than in male rats (P = 0.006). Despite impaired septation, O(2) saturation was not compromised in Dex- or Dex + RA-treated rats. Thus lung function changes induced by Dex treatment during alveolarization were not prevented by concomitant treatment with RA. PMID- 12114189 TI - Role of adipocyte differentiation-related protein in surfactant phospholipid synthesis by type II cells. AB - Adipocyte differentiation-related protein (ADrP) is an intrinsic lipid storage droplet protein that is highly expressed in lung. ADrP localizes to lipid storage droplets within lipofibroblasts, pulmonary cells characterized by high triacylglycerol, which is a precursor for surfactant phospholipid synthesis by alveolar type II epithelial (EPII) cells. The developmental pattern of ADrP mRNA and protein expression in lung tissue parallels triacylglycerol accumulation in rat lung. ADrP mRNA levels are relatively high in isolated lipofibroblasts, accounting for the high ADrP expression in lung. Isolated EPII cells, which do not store neutral lipids but derive them from lipofibroblasts, have low levels of ADrP mRNA expression. ADrP is found around lipid droplets in cultured lipofibroblasts, but not in EPII cells isolated from developing rat lung. After coculture with lipofibroblasts, EPII cells acquired ADrP, which associates with lipid droplets. Furthermore, (3)H-labeled triolein in isolated ADrP-coated lipid droplets is a tenfold better substrate for surfactant phospholipid synthesis by cultured EPII cells than (3)H-labeled synthetic triolein alone. Antibodies to ADrP block transfer of neutral lipid. These data suggest a role for ADrP in this novel mechanism for the transfer of lipid between lipofibroblasts and EPII cells. PMID- 12114190 TI - Inhibition of inwardly rectifying K(+) channels by cGMP in pulmonary vascular endothelial cells. AB - Endothelial barrier dysfunction is typically triggered by increased intracellular Ca(2+) concentration. Membrane-permeable analogs of guanosine 3',5'-cyclic monophosphate (cGMP) prevent disruption of endothelial cell integrity. Because membrane potential (E(m)), which influences the electrochemical gradient for Ca(2+) influx, is regulated by K(+) channels, we investigated the effect of 8 bromo-cGMP on E(m) and inwardly rectifying K(+) (K(IR)) currents in bovine pulmonary artery and microvascular endothelial cells (BPAEC and BMVEC), using whole cell patch-clamp techniques. Both cell types exhibited inward currents at potentials negative to -50 mV that were abolished by application of 10 microM Ba(2+), consistent with K(IR) current. Ba(2+) also depolarized both cell types. 8 Bromo-cGMP (10(-3) M) depolarized BPAEC and BMVEC and inhibited K(IR) current. Pretreatment with Rp-8-cPCT-cGMPS or KT-5823, protein kinase G (PKG) antagonists, did not prevent current inhibition by 8-bromo-cGMP. These data suggest that 8 bromo-cGMP induces depolarization in BPAEC and BMVEC due, in part, to PKG independent inhibition of K(IR) current. The depolarization could be a protective mechanism that prevents endothelial cell barrier dysfunction by reducing the driving force for Ca(2+) entry. PMID- 12114191 TI - Lung alveoli: endogenous programmed destruction and regeneration. AB - Mammalian alveoli, complex architectural and cellular units with dimensions that are linked to the organism's O2 consumption (VO2), are thought to be destroyed only by disease and not to spontaneously regenerate. Calorie restriction of adult mammals lowers VO2, and ad libitum refeeding returns VO2 to pre-calorie restriction values. We took advantage of these relationships and tested the hypothesis in adult mice that calorie restriction (two-thirds reduction for 2 wk) followed by ad libitum refeeding (3 wk) would cause alveolar destruction and regeneration, respectively. Calorie restriction diminished alveolar number 55% and alveolar surface area 25%. Refeeding fully reversed these changes. Neither manipulation altered lung volume. Within 72 h, calorie restriction increased alveolar wall cell apoptosis and diminished lung DNA (approximately 20%). By 72 h of refeeding, alveolar wall cell replication increased and lung DNA rose to amounts in mice that were never calorie restricted. We conclude that adult mice have endogenous programs to destroy and regenerate alveoli, thereby raising the danger of inappropriate activation but the possibility of therapeutic induction, if similar programs exist in humans. PMID- 12114192 TI - Maternal loading with very low-density lipoproteins stimulates fetal surfactant synthesis. AB - We examined whether administration of very low-density lipoproteins (VLDL) to pregnant rats increases surfactant phosphatidylcholine (PtdCho) content in fetal pre-type II alveolar epithelial cells. VLDL-triglycerides are hydrolyzed to fatty acids by lipoprotein lipase (LPL), an enzyme activated by heparin. Fatty acids released by LPL can incorporate into the PtdCho molecule or activate the key biosynthetic enzyme cytidylyltransferase (CCT). Dams were given BSA, heparin, VLDL, or VLDL with heparin intravenously. Radiolabeled VLDL given to the pregnant rat crossed the placenta and was distributed systemically in the fetus and incorporated into disaturated PtdCho (DSPtdCho) in pre-type II cells. Maternal administration of VLDL with heparin increased DSPtdCho content in cells by 45% compared with control (P < 0.05). VLDL produced a dose-dependent, saturable, and selective increase in CCT activity. VLDL did not significantly alter immunoreactive CCT content but increased palmitic, stearic, and oleic acids in pre-type II cells. Furthermore, hypertriglyceridemic apolipoprotein E knockout mice contained significantly greater levels of DSPtdCho content in alveolar lavage and CCT activity compared with either LDL receptor knockout mice or wild type controls that have normal serum triglycerides. Thus the nutritional or genetic modulation of serum VLDL-triglycerides provides specific fatty acids that stimulate PtdCho synthesis and CCT activity thereby increasing surfactant content. PMID- 12114193 TI - Urokinase induces its own expression in Beas2B lung epithelial cells. AB - The urokinase-type plasminogen activator (uPA) interacts with its receptor (uPAR) to promote local proteolysis as well as cellular proliferation and migration. These functions contribute to the pathogenesis of lung inflammation and remodeling as well as the growth and invasiveness of lung neoplasms. In this study, we sought to determine if uPA alters its own expression in lung epithelial cells. Using immunoprecipitation and Western and Northern blotting techniques, we found that uPA treatment enhanced uPA expression in Beas2B lung epithelial cells in a time- and concentration-dependent manner. The induction of uPA expression is mediated through its cell surface receptor uPAR and does not require uPA enzymatic activity. The amino-terminal fragment of uPA, lacking the catalytic domain, is sufficient to induce uPA expression. The serine protease plasmin and the protease inhibitor aprotinin failed to alter uPA-mediated uPA expression, whereas alpha-thrombin potentiated the response. Pretreatment of Beas2B cells with a tyrosine kinase inhibitor, herbimycin, suggests that activation of tyrosine kinase(s) is involved in the uPA-mediated uPA expression. Induction of uPA expression by exposure of lung-derived epithelial cells to uPA is a newly defined pathway by which this protease could influence expression of local fibrinolytic activity and other uPA-dependent cellular responses germane to lung inflammation or neoplasia. PMID- 12114194 TI - Liquid secretion inhibitors reduce mucociliary transport in glandular airways. AB - Because of its possible importance in cystic fibrosis (CF) pulmonary pathogenesis, the effect of anion and liquid secretion inhibitors on airway mucociliary transport was examined. When excised porcine tracheas were treated with ACh to induce gland liquid secretion, the rate of mucociliary transport was increased nearly threefold from 2.5 +/- 0.5 to 6.8 +/- 0.8 mm/min. Pretreatment with both bumetanide and dimethylamiloride (DMA), to respectively inhibit Cl(-) and HCO secretion, significantly reduced mucociliary transport in the presence of ACh by 92%. Pretreatment with the anion channel blocker 5-nitro-2-(3 phenylpropylamino)benzoic acid similarly reduced mucociliary transport in ACh treated airways by 97%. These agents did not, however, reduce ciliary beat frequency. Luminal application of benzamil to block liquid absorption significantly attenuated the inhibitory effects of bumetanide and DMA on mucociliary transport. We conclude that anion and liquid secretion is essential for normal mucociliary transport in glandular airways. Because the CF transmembrane conductance regulator protein likely mediates Cl(-), HCO, and liquid secretion in normal glands, we speculate that impairment of gland liquid secretion significantly contributes to defective mucociliary transport in CF. PMID- 12114195 TI - Oxidative stress and inflammation contribute to lung toxicity after a common breast cancer chemotherapy regimen. AB - Delayed pulmonary toxicity syndrome after high-dose chemotherapy (HDC) and autologous hematopoietic support occurs in up to 64% of women with advanced-stage breast cancer. Using a similar, but nonmyeloablative, HDC treatment regimen in mice, we found both immediate and persistent lung injury, coincident with marked decreases in lung tissue glutathione reductase activity and accompanied by increases in lung oxidized glutathione, bronchoalveolar lavage (BAL) lipid peroxidation, and BAL total cell counts. Most interestingly, at 6 wk, BAL total cell counts had increased fourfold, with lymphocyte cell counts increasing >11 fold. A single supplemental dose of glutathione prevented early lung injury at 48 h but showed no lung-protective effects at 6 wk, whereas single doses of other thiol-sparing agents (Ethyol and glutathione monoethyl ester) showed no benefit. These data suggest that this HDC regimen results in acute and persistent lung toxicity, induced in part by oxidative stress, that culminates with an acute lung cellular inflammatory response. Continuous glutathione supplementation and/or attenuation of the delayed pulmonary inflammatory response may prove beneficial in preventing lung toxicity after the use of these chemotherapeutic agents. PMID- 12114196 TI - Regulation of heme oxygenase-1 by nitric oxide during hepatopulmonary syndrome. AB - During hepatopulmonary syndrome caused by liver cirrhosis, pulmonary endothelial nitric oxide (NO) synthase (NOS) expression and NO production are increased. Increased NO contributes to the blunted hypoxic pressor response (HPR) during cirrhosis and may induce heme oxygenase-1 (HO-1) expression and carbon monoxide (CO) production, exacerbating the blunted HPR. We hypothesized that NO regulates the expression of HO-1 during cirrhosis, contributing to hepatopulmonary syndrome. Cirrhosis was induced in rats by common bile duct ligation (CBDL). Rats were studied 2 and 5 wk after CBDL or sham surgery. Lung HO-1 expression was elevated 5 wk after CBDL. Liver HO-1 was increased at 2 wk and remained elevated at 5 wk. In catheterized rats, the blunted HPR was partially restored by HO inhibition. Rats treated with the NOS inhibitor N(G)-nitro-L-arginine methyl ester for the entire 2- or 5-wk duration had normalized HO-1 expression and HPR. These data provide in vivo evidence for the NO-mediated upregulation of HO-1 expression and support the concept that hepatopulmonary syndrome is multifactorial, involving not only NO, but also HO-1 and CO. PMID- 12114197 TI - PI3K is required for proliferation and migration of human pulmonary vascular smooth muscle cells. AB - Human vascular smooth muscle cell proliferation and migration contribute to vascular remodeling in pulmonary hypertension and atherosclerosis. The precise mechanisms that regulate structural remodeling of the vessel wall remain unknown. This study tests the hypothesis that phosphatidylinositol 3-kinase (PI3K) activation is both necessary and sufficient to mediate human pulmonary vascular smooth muscle (PVSM) cell proliferation and migration. Microinjection of human PVSM cells with a dominant-negative class IA PI3K inhibited platelet-derived growth factor (PDGF)-induced DNA synthesis by 65% (P < 0.001; chi(2) analysis) compared with cells microinjected with control plasmid, whereas microinjection of cells with a constitutively active class IA PI3K (p110*-CA) was sufficient to induce DNA synthesis (mitotic index of p110*-CA-microinjected cells was 15% vs. 3% in control cells; P < 0.01). Transfection of PVSM cells with p110*-CA was also sufficient to promote human PVSM cell migration. In parallel experiments, stimulation of human PVSM cells with PDGF induced PI3K-dependent activation of Akt, p70 S6 kinase, and ribosomal protein S6 but not mitogen-activated protein kinase. PDGF-induced proliferation and migration was inhibited by LY-294002. These results demonstrate that PI3K signaling is both necessary and sufficient to mediate human PVSM cell proliferation and migration and suggest that the activation of PI3K may play an important role in vascular remodeling. PMID- 12114198 TI - MAPK activation is involved in posttranscriptional regulation of RSV-induced RANTES gene expression. AB - Airway epithelial cells represent the primary cell target of respiratory syncytial virus (RSV) infection. They actively participate in the lung immune/inflammatory response that follows RSV infection by expressing chemokines, small chemotactic cytokines that recruit and activate leukocytes. Regulated on activation, normal T cell expressed, and presumably secreted (RANTES) is a member of the CC chemokine subfamily and is strongly chemotactic for T lymphocytes, monocytes, basophils, and eosinophils, cell types that are present or activated in the inflammatory infiltrate that follows RSV infection of the lung. RSV infection of airway epithelial cells induces RANTES expression by increasing gene transcription and stabilizing RNA transcripts. The signaling pathway regulating RANTES gene expression after RSV infection has not been determined. In this study, we examined the role of extracellular signal-regulated kinase (ERK) and p38, members of the mitogen-activated protein (MAP) kinase (MAPK) family, in RSV induced RANTES production. RSV infection of alveolar epithelial cells induced increased phosphorylation and catalytic activity of ERK and the upstream kinases Raf-1 and MAP ERK kinase. Induction of the MAP signaling cascade required a replication-competent virus. RSV infection of alveolar epithelial cells also induced activation of p38 MAPK. Inhibition of ERK and p38 activation significantly reduced RSV-induced RANTES mRNA and protein secretion without affecting RANTES gene transcription or transcription factor activation. These results indicate that the MAPK signaling cascade regulates RANTES production in alveolar epithelial cells through a posttranscriptional mechanism. PMID- 12114199 TI - Mechanisms of bradykinin-mediated dilation in newborn piglet pulmonary conducting and resistance vessels. AB - Bradykinin (BK) is a potent dilator of the perinatal pulmonary circulation. We investigated segmental differences in BK-induced dilation in newborn pig large conducting pulmonary artery and vein rings and in pressurized pulmonary resistance arteries (PRA). In conducting pulmonary arteries and veins, BK-induced relaxation is abolished by endothelial disruption and by inhibition of nitric oxide (NO) synthase with nitro-L-arginine (L-NA). In PRA, two-thirds of the dilation response is L-NA insensitive. Charybdotoxin plus apamin and depolarization with KCl abolish the L-NA-insensitive dilations, findings that implicate the release of endothelium-derived hyperpolarizing factor (EDHF). However, endothelium-disrupted PRA retain the ability to dilate to BK but not to ACh or A-23187. In endothelium-disrupted PRA, dilation was inhibited by charybdotoxin. Thus in PRA, BK elicits dilation by multiple and duplicative signaling pathways. Release of NO and EDHF contributes to the response in endothelium-intact PRA; in endothelium-disrupted PRA, dilation occurs by direct activation of vascular smooth muscle calcium-dependent potassium channels. Redundant signaling pathways mediating pulmonary dilation to BK may be required to assure a smooth transition to extrauterine life. PMID- 12114201 TI - Interaction of matrix with integrin receptors is required for optimal LPS-induced MAP kinase activation. AB - Exposure of macrophages to endotoxin [lipopolysaccharide (LPS)] results in a cascade of events resulting in the release of multiple inflammatory and anti inflammatory mediators. The Toll-like receptor (TLR) 4 complex is the major receptor that mediates LPS signaling. However, there is evidence that other surface molecules may play a complementary role in the TLR-induced events. Integrin receptors are one class of receptors that have been linked to LPS signaling. This study investigates the role of macrophage integrin receptors in the activation of mitogen-activated protein (MAP) kinases by LPS. In conditions where macrophages were not permitted to adhere to matrix or a tissue culture surface, we found a decrease in LPS signaling as documented by a marked reduction in tyrosine phosphorylation of whole cell proteins. This was accompanied by a significant decrease in extracellular signal-regulated kinase and c-Jun NH(2) terminal kinase MAP kinase activation. Inhibition of integrin signaling, with EDTA or RGD peptides, decreased LPS-induced MAP kinase activity. The functional consequence of blocking integrin signaling was demonstrated by decreased LPS induced tumor necrosis factor-alpha production. These observations demonstrate that, in addition to the TLR receptor complex, optimal LPS signaling requires complementary signals from integrin receptors. PMID- 12114200 TI - Role of 5,6-epoxyeicosatrienoic acid in the regulation of newborn piglet pulmonary vascular tone. AB - We examined the responses of newborn piglet pulmonary resistance arteries (PRAs) to 5,6-epoxyeicosatrienoic acid (5,6-EET), a cytochrome P-450 metabolite of arachidonic acid. In PRAs preconstricted with a thromboxane A(2) mimetic, 5,6-EET caused a concentration-dependent dilation. This dilation was partially inhibited by the combination of charybdotoxin (CTX) and apamin, inhibitors of large and small conductance calcium-dependent potassium (K(Ca)) channels, and was abolished by depolarization of vascular smooth muscle with KCl. Disruption of the endothelium significantly attenuated the dilation, suggesting involvement of one or more endothelium-derived vasodilator pathways in this response. The dilation was partially inhibited by nitro-L-arginine (L-NA), an inhibitor of nitric oxide synthase (NOS), but was unaffected by indomethacin, a cyclooxygenase (COX) inhibitor. The combined inhibition of NOS and K(Ca) channels with L-NA, CTX, and apamin abolished 5,6-EET-mediated dilation. Similarly, combined inhibition of NOS and COX abolished the response. We conclude that 5,6-EET is a potent vasodilator in newborn piglet PRAs. This dilation is mediated by redundant pathways that include release of nitric oxide (NO) and COX metabolites and activation of K(Ca) channels. The endothelium dependence of this response suggests that 5,6-EET is not itself an endothelium-derived hyperpolarizing factor (EDHF) but may induce the release of one or more endothelium-derived relaxing factors, such as NO and/or EDHF. PMID- 12114202 TI - Glutathione and other low-molecular-weight thiols relax guinea pig trachea ex vivo: interactions with nitric oxide? AB - The aim of this study was to determine the effects of glutathione (GSH) on trachea smooth muscle tension in view of previously reported interactions between GSH and nitric oxide (NO) (Gaston B. Biochim Biophys Acta 1411: 323-333, 1999; Kelm M. Biochim Biophys Acta 1411: 273-289, 1999; and Kharitonov VG, Sundquist AR, and Sharma VS. J Biol Chem 270: 28158-28164, 1995) and the high (millimolar) concentrations of GSH in trachea epithelium (Rahman I, Li XY, Donaldson K, Harrison DJ, and MacNee W. Am J Physiol Lung Cell Mol Physiol 269: L285-L292, 1995). GSH and other thiols (1.0-10 mM) dose dependently decreased the tension in isolated guinea pig tracheas. Relaxations by GSH were paralleled with sevenfold increased nitrite levels (P < 0.05) in the tracheal effluent, suggesting an interaction between GSH and NO. However, preincubation with a NO scavenger did not reduce the relaxations by GSH or its NO adduct, S-nitrosoglutathione (GSNO). Inhibition of guanylyl cyclase inhibited the relaxations induced by GSNO, but not by GSH. Blocking potassium channels, however, completely abolished the relaxing effects of GSH (P < 0.05). Preincubation of tracheas with GSH significantly (P < 0.05) suppressed hyperreactivity to histamine as caused by removal of tracheal epithelium. These data indicate that GSH plays a role in maintaining tracheal tone. The mechanism is probably an antioxidative action of GSH itself rather than an action of NO or GSNO. PMID- 12114203 TI - Glutathione prevents inhibition of fibroblast-mediated collagen gel contraction by cigarette smoke. AB - Cigarette smoke, the major risk factor for the development of emphysema, contains over 4,700 chemical compounds, including free radicals and other oxidants (10(14)/puff). An imbalance between oxidants and antioxidants has been proposed in the pathogenesis of chronic obstructive pulmonary disease. Inhibition of repair processes has been suggested to be one pathway contributing to the development of emphysema. We hypothesized that cigarette smoke inhibition of repair might result from a shift of the oxidant/antioxidant balance in favor of oxidants. To evaluate this hypothesis, N-acetyl-L-cysteine (NAC), which serves as a substrate for glutathione (GSH) production, and buthionine sulfoximine (BSO), which inhibits GSH production, were incubated in the presence and absence of cigarette smoke extract (CSE) with fibroblasts in three-dimensional collagen gels. Neither agent alone altered gel contraction. CSE inhibition of gel contraction, however, was mitigated by NAC and potentiated by BSO. Parallel effects were observed on cigarette smoke inhibition of fibronectin production and mRNA expression as well as by changes in intracellular GSH content. Pretreatment of fibroblasts with NAC or BSO resulted in similar effects, suggesting that neither agent was acting directly on smoke but, rather, was altering cellular response to smoke. In conclusion, smoke inhibition of fibroblast repair, as reflected by collagen gel contraction and fibronectin production, may be modulated by intracellular GSH levels. PMID- 12114204 TI - TNF-alpha inhibits SP-A gene expression in lung epithelial cells via p38 MAPK. AB - Surfactant protein A (SP-A), the major lung surfactant-associated protein, mediates local defense against pathogens and modulates inflammation in the alveolus. Tumor necrosis factor (TNF)-alpha, a proinflammatory cytokine, inhibits SP-A gene expression in lung epithelial cells. Inhibitors of the phosphatidylinositol 3-kinase pathway, i.e., wortmannin, LY-294002, and rapamycin, did not block the inhibitory effects of TNF-alpha on SP-A mRNA levels. An inhibitor of the p44/42 mitogen-activated protein kinase (MAPK) pathway, PD 98059, was also ineffective. PD-169316 and SB-203580, inhibitors of p38 MAPK, blocked the TNF-alpha-mediated inhibition of SP-A mRNA levels. TNF-alpha increased the phosphorylation of p38 MAPK within 15 min. Anisomycin, an activator of p38 MAPK, increased p38 MAPK phosphorylation and decreased SP-A mRNA levels in a dose-dependent manner. Finally, TNF-alpha increased the phosphorylation of ATF 2, a transcription factor that is a p38 MAPK substrate. We conclude that TNF alpha downregulates SP-A gene expression in lung epithelial cells via the p38 MAPK signal transduction pathway. PMID- 12114205 TI - Prostacyclin analogs inhibit fibroblast migration. AB - The controlled accumulation of fibroblasts to sites of inflammation is crucial to effective tissue repair after injury. Either inadequate or excessive accumulation of fibroblasts could result in abnormal tissue function. Prostacyclin (PGI(2)) is a potent mediator in the coagulation and inflammatory processes. The aim of this study was to investigate the effect of PGI(2) on chemotaxis of human fetal lung fibroblasts (HFL-1). Using the blind well chamber technique, we found that the PGI(2) analog carbaprostacyclin (10(-6) M) inhibited HFL-1 chemotaxis to human plasma fibronectin (20 microg/ml) 58.0 +/- 13.2% (P < 0.05) and to platelet derived growth factor (PDGF)-BB (10 ng/ml) 48.7 +/- 4.6% (P < 0.05). Checkerboard analysis demonstrated that carbaprostacyclin inhibits both directed and undirected migration. The inhibitory effect of the carbaprostacyclin was concentration dependent and blocked by the cAMP-dependent protein kinase (PKA) inhibitor KT-5720, suggesting that a cAMP-PKA pathway may be involved in the process. Two other PGI(2) analogs, ciprostene and dehydro-15-cyclohexyl carbaprostacyclin (both 10(-6) M), significantly inhibited fibroblast migration to fibronectin. In summary, PGI(2) appears to inhibit fibroblast chemotaxis to fibronectin and PDGF-BB. Such an effect may contribute to the regulation of fibroblasts in wound healing and could contribute to the pathogenesis of diseases characterized by abnormal tissue repair remodeling. PMID- 12114206 TI - Physiological properties and functions of Ca(2+) sparks in rat intrapulmonary arterial smooth muscle cells. AB - Ca(+) spark has been implicated as a pivotal feedback mechanism for regulating membrane potential and vasomotor tone in systemic arterial smooth muscle cells (SASMCs), but little is known about its properties in pulmonary arterial smooth muscle cells (PASMCs). Using confocal microscopy, we identified spontaneous Ca(2+) sparks in rat intralobar PASMCs and characterized their spatiotemporal properties and physiological functions. Ca(2+) sparks of PASMCs had a lower frequency and smaller amplitude than cardiac sparks. They were abolished by inhibition of ryanodine receptors but not by inhibition of inositol trisphosphate receptors and L-type Ca(2+) channels. Enhanced Ca(2+) influx by BAY K8644, K(+), or high Ca(2+) caused a significant increase in spark frequency. Functionally, enhancing Ca(2+) sparks with caffeine (0.5 mM) caused membrane depolarization in PASMCs, in contrast to hyperpolarization in SASMCs. Norepinephrine and endothelin 1 both caused global elevations in cytosolic Ca(2+) concentration ([Ca(2+)]), but only endothelin-1 increased spark frequency. These results suggest that Ca(2+) sparks of PASMCs are similar to those of SASMCs, originate from ryanodine receptors, and are enhanced by Ca(2+) influx. However, they play a different modulatory role on membrane potential and are under agonist-specific regulation independent of global [Ca(2+)]. PMID- 12114207 TI - Primary inflammation in human cystic fibrosis small airways. AB - Most cystic fibrosis (CF) patients die of lung failure, due to the combined effects of bacterial infection, neutrophil-mediated inflammation, and airway obstruction by hyperviscous mucus. To this day, it remains unclear where and how this pathological vicious circle is initiated in vivo. In particular, it has proven difficult to investigate whether inflammatory pathways are dysregulated in CF airways independently of infection. Also, the relative involvement of large (tracheobronchial) vs. small (bronchiolar) airways in CF pathophysiology is still unclear. To help address these issues, we used an in vivo model based on the maturation of human fetal CF and non-CF small airways in severe combined immunodeficiency mice. We show that uninfected mature CF small airway grafts, but not matched non-CF controls, undergo time-dependent neutrophil-mediated inflammation, leading to progressive lung tissue destruction. This model of mature human small airways provides the first clear-cut evidence that, in CF, inflammation may arise at least partly from a primary defect in the regulation of neutrophil recruitment, independently of infection. PMID- 12114209 TI - GM-CSF expression by human lung microvascular endothelial cells: in vitro and in vivo findings. AB - Recently, many findings indicate that granulocyte-macrophage colony-stimulating factor (GM-CSF) plays an important role in the pathogenesis of acute and chronic lung diseases. In the present paper, the production of this cytokine in human pulmonary microvascular endothelial cells (HPMEC) is investigated. In an in vitro study, quiescent HPMEC did not express GM-CSF, either at the transcriptional or at the protein level. After activation for 4 h with tumor necrosis factor (TNF) alpha (30/300 U/ml), lipopolysaccharide (LPS; 0.1/1 microg/ml), or interleukin (IL)-1 beta (100 U/ml), a significant release of GM-CSF was measured by enzyme linked immunosorbent assay, with a time-dependent increase over 72 h. IL-8 (4, 16, or 64 ng/ml) or IL-1 beta at a concentration of 10 U/ml did not induce the release of GM-CSF. Human umbilical vein endothelial cells (HUVEC) and the angiosarcoma cell line HAEND served as reference cell lines. GM-CSF release in HPMEC was significantly (P < 0.025-0.05) less inducible by IL-1 beta than in HUVEC. A constitutive expression of GM-CSF by HAEND was observed. Additionally, GM-CSF expression in vivo by the lung microvasculature was confirmed by immunohistochemistry in lung tissue. To our knowledge, this is the first report of the ability of human pulmonary endothelial cells to synthesize and release GM CSF. These results support the hypothesis that the lung microvasculature via the production of GM-CSF is a potential contributor to the cytokine network in lung diseases. This could be of particular importance in the pathogenesis of the acute respiratory distress syndrome in which endothelial dysfunction plays a central pathogenetic role. PMID- 12114208 TI - Injury, inflammation, and remodeling in fetal sheep lung after intra-amniotic endotoxin. AB - Chorioamnionitis is frequent in preterm labor and increases the risk of bronchopulmonary dysplasia. We hypothesized that intra-amniotic endotoxin injures the lung in utero, causing a sequence of inflammation and tissue injury similar to that which occurs in the injured adult lung. Preterm lamb lungs at 125 days gestational age were evaluated for indicators of inflammation, injury, and repair 5 h, 24 h, 72 h, and 7 days after 4 mg of intra-amniotic endotoxin injection. At 5 h, the epithelial cells in large airways expressed heat shock protein 70, and alveolar interleukin-8 was increased. Surfactant protein B (SP-B) decreased in alveolar type II cells at 5 h, and SP-B in lung tissue and alveolar lavage fluid increased by 72 h. By 24 h, neutrophils were recruited into the large airways, and cell death was the highest. Alveolar type II cells decreased by 25% at 24 h, and proliferation was highest at 72 h, consistent with tissue remodeling. Intra amniotic endotoxin caused surfactant secretion, inflammation, cell death, and remodeling as indications of lung injury. The recovery phase was accompanied by maturational changes in the fetal lung. PMID- 12114210 TI - GATA-6 is required for maturation of the lung in late gestation. AB - GATA-6, a member of the GATA family of zinc finger proteins, is the only family member known to be expressed in the epithelial cells of the developing airway epithelium. To determine the role of GATA-6 in lung morphogenesis, a chimeric fusion protein containing GATA-6 and the strong transcriptional inhibitor, engrailed, was conditionally expressed in mice under control of a doxycycline inducible transgene. Expression of GATA-6-engrailed was initiated at embryonic day (E) 6-7 by treatment of the dam with doxycycline. Although branching morphogenesis of the proximal airways proceeded normally to E16.5, maturation of terminal airways and alveoli that normally occurs before birth was inhibited. At E17.5-18.5, aquaporin-5 mRNA and type I cell marker-alpha staining, both markers of type I cells, were decreased. Homogeneous distribution of the thyroid transcription factor-1, decreased expression of surfactant proteins, delayed thinning of the walls of the peripheral airways, and lack of squamous differentiation of epithelial cells were observed in the lung periphery after expression of GATA-6-engrailed. Activity of GATA-6 is required for maturation of the gas exchange area before birth. PMID- 12114212 TI - Enhanced nitric oxide and reactive oxygen species production and damage after inhalation of silica. AB - In previous reports from this study, measurements of pulmonary inflammation, bronchoalveolar lavage cell cytokine production and nuclear factor-kappa B activation, cytotoxic damage, and fibrosis were detailed. In this study, we investigated the temporal relationship between silica inhalation, nitric oxide (NO), and reactive oxygen species (ROS) production, and damage mediated by these radicals in the rat. Rats were exposed to a silica aerosol (15 mg/m(3) silica, 6 h/day, 5 days/wk) for 116 days. We report time-dependent changes in 1) activation of alveolar macrophages and concomitant production of NO and ROS, 2) immunohistochemical localization of inducible NO synthase and the NO-induced damage product nitrotyrosine, 3) bronchoalveolar lavage fluid NO(x) and superoxide dismutase concentrations, and 4) lung lipid peroxidation levels. The major observations made in this study are as follows: 1) NO and ROS production and resultant damage increased during silica exposure, and 2) the sites of inducible NO synthase activation and NO-mediated damage are associated anatomically with pathological lesions in the lungs. PMID- 12114211 TI - Role of heme oxygenases in sepsis-induced diaphragmatic contractile dysfunction and oxidative stress. AB - Heme oxygenases (HOs), essential enzymes for heme metabolism, play an important role in the defense against oxidative stress. In this study, we evaluated the expression and functional significance of HO-1 and HO-2 in the ventilatory muscles of normal rats and rats injected with bacterial lipopolysaccharide (LPS). Both HO-1 and HO-2 proteins were detected inside ventilatory and limb muscle fibers of normal rats. Diaphragmatic HO-1 and HO-2 expressions rose significantly within 1 and 12 h of LPS injection, respectively. Inhibition of the activity of inducible nitric oxide synthase (iNOS) in rats and absence of this isoform in iNOS(-/-) mice did alter sepsis-induced regulation of muscle HOs. Systemic inhibition of HO activity with chromium mesoporphyrin IX enhanced muscle protein oxidation and hydroxynonenal formation in both normal and septic rats. Moreover, in vitro diaphragmatic force generation declined substantially in response to HO inhibition both in normal and septic rats. We conclude that both HO-1 and HO-2 proteins play an important role in the regulation of muscle contractility and in the defense against sepsis-induced oxidative stress. PMID- 12114213 TI - Nerve growth factor and nerve growth factor receptors in respiratory syncytial virus-infected lungs. AB - Nerve growth factor (NGF) controls sensorineural development and responsiveness and modulates immunoinflammatory reactions. Respiratory syncytial virus (RSV) potentiates the proinflammatory effects of sensory nerves in rat airways by upregulating the substance P receptor, neurokinin 1 (NK(1)). We investigated whether the expression of NGF and its trkA and p75 receptors in the lungs is age dependent, whether it is upregulated during RSV infection, and whether it affects neurogenic inflammation. Pathogen-free rats were killed at 2 (weanling) to 12 (adult) wk of age; in addition, subgroups of rats were inoculated with RSV or virus-free medium. In pathogen-free rats, expression of NGF and its receptors in the lungs declined with age, but RSV doubled expression of NGF, trkA, and p75 in weanling and adult rats. Exogenous NGF upregulated NK(1) receptor expression in the lungs. Anti-NGF antibody inhibited NK(1) receptor upregulation and neurogenic inflammation in RSV-infected lungs. These data indicate that expression of NGF and its receptors in the lungs declines physiologically with age but is upregulated by RSV and is a major determinant of neurogenic inflammation. PMID- 12114214 TI - Constitutively active ErbB4 and ErbB2 mutants exhibit distinct biological activities. AB - ErbB4 is a member of the epidermal growth factor receptor(EGFR) family of tyrosine kinases, which includes EGFR/ErbB1, ErbB2/HER2/Neu, and ErbB3/HER3. These receptors play important roles both in normal development and in neoplasia. For example, deregulated signaling by ErbB1 and ErbB2 is observed in many human malignancies. In contrast, the roles that ErbB4 plays in tumorigenesis and normal biological processes have not been clearly defined. To identify the biological responses that are coupled to ErbB4, we have constructed three constitutively active ErbB4 mutants. Unlike a constitutively active ErbB2 mutant, the ErbB4 mutants are not coupled to increased cell proliferation, loss of contact inhibition, or anchorage independence in a rodent fibroblast cell line. This suggests that ErbB2 and ErbB4 may play distinct roles in tumorigenesis in vivo. PMID- 12114215 TI - Expression profiling of lineage differentiation in pluripotential human embryonal carcinoma cells. AB - Pluripotential human embryonal carcinoma (EC) cell linesundergo differentiation programs resembling those occurring in embryonal stem cells during development. Expression profiling was performed during the terminal differentiation of the EC cell line, NTera2/Clone D1 by all-trans-retinoic acid. Time-response analysis via clustering of >12,000 human transcripts revealed distinct stages in the transition from an EC cell to neuronal progenitor cells expressing patterning markers compatible with posterior hindbrain fates followed by the appearance of immature postmitotic neurons with an evolving synaptic apparatus. Global analysis of gene expression allows monitoring cell fate and differentiation of EC cells in vitro and may provide insight into human embryonal stem cell development. PMID- 12114216 TI - Adenoviral delivery of an antisense RNA complementary to the 3' coding sequence of transforming growth factor-beta1 inhibits fibrogenic activities of hepatic stellate cells. AB - Liver fibrosis occurs as a consequence of the transdifferentiationof hepatic stellate cells into myofibroblasts and is associated with an increased expression and activation of transforming growth factor (TGF)-beta1. This pluripotent, profibrogenic cytokine stimulates matrix synthesis and decreases matrix degradation, resulting in fibrosis. Thus, blockade of synthesis or sequestering of mature TGF-beta1 is a primary target for the development of antifibrotic approaches. The purpose of this study was to investigate whether the administration of adenoviruses constitutively expressing an antisense mRNA complementary to the 3' coding sequence of TGF-beta1 is able to suppress the synthesis of TGF-beta1 in culture-activated hepatic stellate cells. We demonstrate that the adenoviral vehicle directs high-level expression of the transgene and proved that the transduced antisense is biologically active by immunoprecipitation, Western blot, quantitative TGF-beta1 ELISA, and cell proliferation assays. Additionally, the biological function of the transgene was confirmed by analysis of differential activity of TGF-beta1-responsive genes using cell ELISA, Northern blotting, and by microarray technology, respectively. Furthermore, we examined the effects of that transgene on the expression of TGF beta2, TGF-beta3, collagen type alpha1(I), latent transforming growth factor binding protein 1, types I and II TGF-beta receptors, and alpha-smooth muscle actin. Our results indicate that the administration of antisense mRNA offers a feasible approach to block autocrine TGF-beta1 signaling in hepatic stellate cells and may be useful and applicable in future to the treatment of fibrosis in chronic liver diseases. PMID- 12114218 TI - Macroeconomics and health. PMID- 12114217 TI - The role of growth factors in the activity of pharmacological differentiation agents. AB - Bryostatin-1 inhibits acute myeloid leukemia (AML) in vitroat doses that stimulate the growth of normal hematopoietic progenitors.Although bryostatin-1 has a number of distinct biological activities, those specifically responsible for its antileukemic activity are unclear. We found that bryostatin-1 (10(-8) M) inhibited cell cycling at G(1), induced phenotypic evidence of differentiation, and limited the clonogenic growth of both AML cell lines and patient specimens. This activity was markedly enhanced by granulocyte/macrophage-colony stimulating factor, whereas growth factor-neutralizing antibodies completely inhibited both the differentiating and antileukemic activities of bryostatin-1. Cell cycle inhibition and growth factors were also required for the differentiating activities of two unrelated agents, hydroxyurea and phenylbutyrate. These data suggest that many pharmacological differentiating agents require both cell cycle arrest and lineage-specific growth factors for full activity and may explain why these agents have demonstrated only limited clinical efficacy. PMID- 12114219 TI - A time for global health. PMID- 12114220 TI - A learning world for the Global Fund. PMID- 12114221 TI - Primary prevention of coronary heart disease. PMID- 12114222 TI - Growth hormone in growth hormone deficiency. PMID- 12114223 TI - Diabetic nephropathy. PMID- 12114227 TI - Hopes rise for patients with drug resistant HIV. PMID- 12114228 TI - GPs should use nurses more to lighten their load. PMID- 12114229 TI - Doctors face trial for manslaughter as criminal charges against doctors continue to rise. PMID- 12114230 TI - UK has highest reduction in deaths from lung and breast cancer. PMID- 12114231 TI - Exponent of "male menopause" censured by GMC. PMID- 12114232 TI - New BMA president compares a third of NHS care to care in the developing world. PMID- 12114233 TI - UK government should stop recruiting doctors from abroad. PMID- 12114234 TI - Prospective multicentre randomised trial of tension-free vaginal tape and colposuspension as primary treatment for stress incontinence. AB - OBJECTIVE: To compare tension-free vaginal tape with colposuspension as primary treatment for stress incontinence. DESIGN: Multicentred randomised comparative trial. SETTING: Gynaecology or urology departments in 14 centres in the United Kingdom and Eire, including university teaching hospitals and district general hospitals. PARTICIPANTS: 344 women with urodynamic stress incontinence; 175 randomised to tension-free vaginal tape and 169 to colposuspension MAIN OUTCOME MEASURES: Assessment before treatment and at six months postoperatively with the SF-36, the Bristol female lower urinary tract symptoms questionnaire, the EQ-5D health questionnaire, a one week urinary diary, one hour perineal pad test, cystometry, and, in some centres, urethral profilometry. RESULTS: 23 women in the colposuspension group and 5 in the vaginal tape group withdrew before surgery. No significant difference was found between the groups for cure rates: 115 (66%) women in the vaginal tape group and 97 (57%) in the colposuspension group were objectively cured (95% confidence interval for difference in cure -4.7% to 21.3%). Bladder injury was more common during the vaginal tape procedure; postoperative complications, in particular delayed resumption of micturition, were more common after colposuspension. Operation time, duration of hospital stay, and return to normal activity were all longer after colposuspension than after the vaginal tape procedure. CONCLUSION: Surgery with tension-free vaginal tape is associated with more operative complications than colposuspension, but colposuspension is associated with more postoperative complications and longer recovery. Vaginal tape shows promise for the treatment of urodynamic stress incontinence because of minimal access and rapid recovery times; cure rates at six months were comparable with colposuspension. PMID- 12114235 TI - Adult height after long term treatment with recombinant growth hormone for idiopathic isolated growth hormone deficiency: observational follow up study of the French population based registry. AB - OBJECTIVE: To evaluate the efficacy of recombinant growth hormone for increasing adult height in children treated for idiopathic isolated growth hormone deficiency. DESIGN: Observational follow up study. SETTING: Population based registry. PARTICIPANTS: All 2852 French children diagnosed as having isolated idiopathic growth hormone deficiency whose treatment started between 1987 and 1992 and ended before 1996. MAIN OUTCOME MEASURES: Change in height between the start of treatment and adulthood; classification of patients according to whether treatment was completed as scheduled or stopped early. RESULTS: Adult height was obtained for 2165 (76%) patients. The mean dose of growth hormone at start of treatment was 0.42 IU/kg/week. Height gain was 1.1 (SD 0.9) standard deviation (SD) scores, resulting in an adult height of -1.6 (0.9) SD score (girls, 154 (5) cm; boys, 167 (6) cm). Patients who completed the treatment gained 1.0 (0.7) SD score of height in 3.6 (1.4) years. Patients with treatments stopped early gained 0.6 (0.6) SD score in 2.7 (1.4) years while receiving treatment and a further 0.4 (0.9) SD score after the end of treatment. Most of the variation in height gain was explained by regression towards the mean, patients' characteristics, and delay in starting puberty. Severe growth hormone deficiency was associated with better outcome. Each year of treatment was associated with a gain of 0.2 SD score(1.3 cm). CONCLUSION: The effect of growth hormone is unclear in many patients treated for so called idiopathic isolated growth hormone deficiency. Most of the patients have pubertal delay and a spontaneous growth potential, which must be taken into account when measuring the effect and cost effectiveness of treatments. Growth hormone deficiency should be clearly distinguished from pubertal delay, and criteria should restrict the definition to patients with severely and permanently altered growth hormone secretion as our results support the use of growth hormone in such patients. Long term trials are required for most patients currently treated. PMID- 12114236 TI - Familial, psychiatric, and socioeconomic risk factors for suicide in young people: nested case-control study. AB - OBJECTIVE: To estimate the risk of suicide in young people related to family and individual psychiatric and socioeconomic factors. DESIGN: Population based nested case-control study. SETTING: Data from longitudinal Danish registers. CASES AND CONTROL: 496 young people aged 10-21 years who had committed suicide during 1981 97 in Denmark and 24, 800 controls matched for sex, age, and time. MAIN OUTCOME AND MEASURES: All suicides in Denmark compared with controls; parents and siblings identified from population based registers; inpatient information from discharge registers of national hospitals; and socioeconomic data from administrative registers. RESULTS: Parental factors associated with an increased risk of suicide in young people were suicide or early death, admission to hospital for a mental illness, unemployment, low income, poor schooling, and divorce, as well as mental illness in siblings and mental illness and short duration of schooling in the young people themselves. The strongest risk factor was mental illness in the young people. The effect of the parents' socioeconomic factors decreased after adjustment for a family history of mental illness and a family history of suicide. CONCLUSIONS: Recognising mental illness in young people and dealing with it appropriately could help prevent suicides. The high relative risk associated with a low socioeconomic status of the parents may be confounded and overestimated if not adjusted for mental illness and suicide in the family. PMID- 12114237 TI - Interaction between warfarin and topical miconazole cream. PMID- 12114238 TI - Screening for cardiovascular risk: public health imperative or matter for individual informed choice? PMID- 12114239 TI - What is newsworthy? Longitudinal study of the reporting of medical research in two British newspapers. AB - OBJECTIVE: To assess the characteristics of medical research that is press released by general medical journals and reported in newspapers. DESIGN: Longitudinal study. DATA SOURCES: All original research articles published in Lancet and BMJ during 1999 and 2000. MAIN OUTCOME MEASURES: Inclusion of articles in Lancet or BMJ press releases, and reporting of articles in Times or Sun newspapers. RESULTS: Of 1193 original research articles, 517 (43%) were highlighted in a press release and 81 (7%) were reported in one or both newspapers. All articles covered in newspapers had been press released. The probability of inclusion in press releases was similar for observational studies and randomised controlled trials, but trials were less likely to be covered in the newspapers (odds ratio 0.15 (95% confidence interval 0.06 to 0.37)). Good news and bad news were equally likely to be press released, but bad news was more likely to be reported in newspapers (1.74 (1.07 to 2.83)). Studies of women's health, reproduction, and cancer were more likely to be press released and covered in newspapers. Studies from industrialised countries other than Britain were less likely to be reported in newspapers (0.51 (0.31 to 0.82)), and no studies from developing countries were covered. CONCLUSIONS: Characteristics of articles were more strongly associated with selection for reporting in newspapers than with selection for inclusion in press releases, although each stage influenced the reporting process. Newspapers underreported randomised trials, emphasised bad news from observational studies, and ignored research from developing countries. PMID- 12114240 TI - Chronic renal disease. PMID- 12114241 TI - Antenatal screening for rubella-infection or immunity? PMID- 12114243 TI - Globalisation and the challenges to health systems. PMID- 12114242 TI - Organising care for chronic illness. PMID- 12114244 TI - Economic modelling before clinical trials. PMID- 12114245 TI - Consultants' new contract. Shurely shome mishtake? PMID- 12114246 TI - Gold for the NHS. No natural limitation exists on demand for services free at point of supply. PMID- 12114247 TI - Postcode prescribing is alive and well in Scotland. PMID- 12114248 TI - vCJD: the epidemic that never was. New variant Creutzfeldt-Jakob disease: the critique that never was. PMID- 12114250 TI - Evolving general practice consultation in Britain. Increasing consultation time may not be straightforward. PMID- 12114249 TI - Has medicalisation of childbirth gone too far? Regional analgesia in labour permits childbirth without fear. PMID- 12114251 TI - The lot of airline pilots and consultants is not so different. PMID- 12114252 TI - Altered neuroendocrine immune (NEI) networks in rheumatology. PMID- 12114253 TI - Perspectives on the relationship of adrenal steroids to rheumatoid arthritis. AB - An expanded model of RA is presented that incorporates cumulative multifactorial processes operating over a prolonged physiological phase prior to initial clinical manifestations. During this phase, progressive imbalances in the homeostasis of core neuroendocrine, immunological, and microvascular systems are believed to occur. Normal adrenal function plays an essential role in helping to maintain homeostasis of core systems in health. In RA, chronic adrenal hypocompetence is suspected to occur in a minority subset of females who have younger clinical onset and males who have associated low serum testosterone levels. Chronic, relative glucocorticoid insufficiency is believed to contribute to development of inflammatory manifestations in RA patients. Androgenic deficiencies, particularly of gonadal origin in males, may also contribute to RA, particularly its decreased anabolic features. Precise influences of hypocompetent adrenal steroid function on long-term modeling of the immunological compartment and control of microvascular activation processes are not well understood. These complex mechanisms need to be elucidated for better understanding of the physiopathogenesis of RA. Nevertheless, at a clinical level, sufficient data are currently available to endorse further controlled studies of early clinical onset patients and prospective investigations to determine more definitively the roles of adrenal (and gonadal) steroids in subsets of RA patients and unaffected susceptible persons in the population. PMID- 12114254 TI - Neuroimmunoendocrinology of the rheumatic diseases: past, present, and future. AB - Adaptation to stressful stimuli, maintenance of homeostasis, and ultimately, survival require bidirectional feedback communication among components of the stress response and immune and endocrine systems. Substantial progress has been made in delineating molecular, cellular, and systemic physiologic mechanisms underlying this communication, particularly mechanisms that target the immune system. For example, our understanding of the immunomodulatory activities of numerous neuroendocrine mediators, such as cortisol, estrogen, testosterone, DHEA, catecholamines, corticotropin-releasing hormone, and adenosine, has advanced substantially. Substantial progress has also been made in defining how abnormalities involving these factors may contribute to the initiation, progression, and severity of autoimmune rheumatic diseases, particularly rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE). For RA, the available data support the view that inflammatory and immune system inhibitory mechanisms, involving the hypothalamic-pituitary-adrenal (HPA) axis and sympathetic nervous system are deficient. Age, gender, and reproductive status acting, in part, through gonadal hormonal effects on disease susceptibility genes also appear likely to modulate the inhibitory stress response systems and immune function. Animal model data also have provided direct evidence that many autoimmune disease regulatory genes are gender influenced. For SLE, a growing body of recent data indicates that estrogens and androgens exert contrasting effects on B-lymphocytes (i.e., estrogens enhance and testosterone suppresses autoantibody production). These observations provide potential new insights into SLE pathogenesis and gender differences in prevalence. Continued investigation will refine our understanding of these observations and will uncover even more extensive interactions of the nervous, immune, and endocrine systems. Moreover, it is highly likely that improved understanding of these interactions will translate into improved therapy for the rheumatic diseases. PMID- 12114255 TI - Neural-immune interactions in health and disease. AB - Many lines of research have established the numerous routes by which the immune and central nervous systems (CNS) communicate. The CNS signals the immune system via hormonal and neuronal pathways and the immune system signals the CNS through similar routes via immune mediators and cytokines. The primary hormonal pathway by which the CNS regulates the immune system is the hypothalamic-pituitary adrenal (HPA) axis, through the hormones of the neuroendocrine stress response. The sympathetic nervous system regulates immune system function primarily via adrenergic neurotransmitters released through neuronal routes. Neuroendocrine regulation of immune function is essential for survival during stress or infection and to modulate immune responses in inflammatory disease. Glucocorticoids are the main effector endpoint of the neuroendocrine response system. PMID- 12114256 TI - Fifty years of experience with cortisone therapy in the study and treatment of rheumatoid arthritis. AB - In 1948 the U.S. rheumatologist Phillip S. Hench administered cortisone for the first time to a patient with rheumatoid arthritis (RA), thereby discovering the therapeutic effects of glucocorticoids. He published this observation together with Kendall, Slocumb, and Polly in 1949, and they received, along with Reichstein and Kendall, the Nobel Prize in Medicine or Physiology in 1950. However, as early as 1949, he rejected the idea that steroids were of etiological significance for RA, and instead stressed their unique place as a tool for pathophysiological research. The discovery of the glucocorticoid receptor and its genomic effects disclosed that there are no qualitative differences between the effects of endogenous cortisol and exogenously applied synthetic glucocorticoids, since all effects are transmitted via the same receptor. Later came the discovery that the hypothalamo-pituitary-adrenal axis is stimulated by cytokines after activation of the immune system. Glucocorticoids are not only the most effective antiphlogistic and immune-suppressive substances with instant effect, but they also show, with low-dosage long-term treatment, clear antiproliferative effects on the cartilage and bone destroying pannus in RA. Little is still known about the precise mechanisms of actions of glucocorticoids in general, and specifically when rheumatic autoimmune diseases are involved. The high effectiveness of these substances and their direct effects via the genomic glucocorticoid receptor allows us to anticipate that uncovering their mechanisms of action will shed deeper insight into the pathomechanisms of these diseases. The use of TNFalpha blockers in the treatment of rheumatoid arthritis and Crohn's disease, with their dramatic immediate effects, comparable with those of the glucocorticoids but without the side effects of the latter, points us in that direction. PMID- 12114257 TI - Mechanisms of corticosteroid resistance in rheumatoid arthritis: a putative role for the corticosteroid receptor beta isoform. AB - Corticosteroids (CSs) have potent immunosuppressive effects and are commonly used to treat a range of immunological and inflammatory diseases such as rheumatoid arthritis (RA). These effects are mediated by the ability of CSs to modulate gene expression. CSs act by binding to the CS receptor (CR), which exists as alpha and beta isoforms. Only CRalpha binds CS. CRbeta functions as an endogenous inhibitor of CS and is expressed in several tissues. The CS/CRalpha complex binds to the glucocorticosteroid response element in the nucleus and also interferes with AP-1 and NF-kappaB binding. Thus, CSs inhibit the transcription of AP-1 and NF-kappaB inducible genes, such as interleukin (IL)-2, IL-6, IL-8, IL-1beta, and tumor necrosis factor (TNF) alpha, as well as T-cell proliferation. In clinical practice, a proportion of RA patients do not respond adequately to CS therapy. On this basis, RA patients can be divided on clinical grounds and on the ability of CSs to inhibit concanavalin A (conA)-induced peripheral blood T-cell proliferation in vitro into CS-sensitive (SS) and CS-resistant (SR) subgroups. The in vitro defined SS and SR subgroups correlate with the clinical responses to CS therapy. The mechanisms of the SR in RA patients remain unknown but may include the following: dysregulation of CRalpha function, alterations in the intracellular signaling mechanisms and/or utilization of various other cellular activation pathways, perturbations of the cytokine milieu, and inhibition of lipocortin. In SR subjects, CSs fail to significantly inhibit conA-induced IL-2 and IL-4 secretion and LPS-induced IL-8, IL-1beta secretion in vitro. CS therapy fails to reduce the circulating levels of IL-8, IL-1beta, and TNFalpha in SR RA patients. Peripheral blood mononuclear cells (PBMCs) from SR significantly overexpress activated NF-kappaB and IkappaBalpha. In vitro CSs fail to significantly inhibit conA-induced NF-kappaB activation in PBMCs from SR RA patients. Our preliminary observations show enhanced CRbeta expression by PBMCs from SR RA patients. It is most likely that other molecular mechanisms such as enhanced AP-1 expression are involved, and we currently are investigating such possibilities. PMID- 12114258 TI - Immune homeostasis requires several biologic factors including glucocorticoid hormones. AB - Immunological tolerance can be achieved by several mechanisms including suppressor cells, soluble factors, and neurohormonal mediators. On the cellular level, we isolated a population of CD8+CD28- T cells capable of inhibiting anti CD3 mAb-induced proliferation of autologous peripheral blood mononuclear cells in an HLA-I nonrestricted manner via production of IFN-gamma and IL-6. Interestingly, CD8+CD28- T cells from systemic lupus erythematosus patients with active disease do not display this inhibitory activity and show a marked imbalance between inhibitory (IL-6) and stimulatory (IL-12) cytokines. For soluble factors, we studied soluble HLA molecules (sHLAs) and double-stranded DNA (ds-DNA). Soluble HLA-I (sHLA-I) molecules induce soluble Fas ligand (sFasL) secretion and trigger apoptosis in phytohemagglutin (PHA)-activated Fas+ T cells. Double-stranded DNA binds to HLA-II molecules and inhibits HLA-II-mediated antigen presentation. On the neurohormonal side, we focused our attention on the immunological activity of corticosteroids (CTSs). CTSs inhibit recirculation of CD4+ T cells, suppress the proliferation and immunological function of activated T cells, and induce apoptosis of activated lymphocytes. Taken together, these data suggest the presence of a complex network of immunoregulatory mechanisms in which CTSs play a strong role supporting their recognized efficacy in the treatment of inflammatory and immunological diseases. PMID- 12114259 TI - Glucocorticoid receptor downregulation in early diagnosed rheumatoid arthritis. AB - In patients with rheumatoid arthritis (RA) with a mean duration of six years, glucocorticoid receptor (GR) downregulation has been reported without change in cortisol levels. This phenomenon might play a role in the etiopathogenesis of RA. If this is the case, GR downregulation should also be found in early-diagnosed RA. We compared GR expression and cortisol levels of early RA patients with those of sex and age matched healthy controls. In early (disease duration less than one year) RA patients (52F/29M; mean [SE] age 63 +/- 2 years) and in 39 controls (23F/16M; mean age 63 +/- 2 years) blood samples were taken between 8:00 and 10:00 a.m. GR expression (GR number), cortisol levels (fluorescence polarization immunoassay, FPIA), ESR, CRP, and the presence of swollen and painful knees were determined. RA was diagnosed according to the ACR classification criteria. In early RA a significantly lower GR expression was found compared to controls. Interestingly, cortisol levels were also significantly lower. Both findings were due to the observed differences in the female population. In the male population no differences were found. No correlations were found among parameters of disease activity, nor was there a relation between GR expression and serum cortisol levels. In female patients with early-diagnosed RA a decrease in glucocorticoid receptor expression was found as well as a decrease in cortisol levels. Since normal glucocorticoid receptor expression was found in the male population with early-diagnosed RA, it remains questionable if glucocorticoid receptor expression is causally involved in pathogenesis of RA. PMID- 12114260 TI - Differential male and female adrenal cortical steroid hormone and cortisol responses to interleukin-6 in humans. AB - Evidence from experimental animal studies show that sex hormones influence the glucocorticoid response to a variety of inflammatory and noninflammatory stimuli. In this study we assessed gender differences in the response of ACTH and cortisol in normal young male and female humans following intravenous infusion of human IL 6 in various dosages. Males presented a significantly stronger ACTH production in response to IL-6 than females. Peak cortisol response, however, was similar in males and females. Cortisol/ACTH ratios were significantly higher in females than in males, both at baseline and after each of the IL-6 dosages. These results suggest that an effective glucocorticoid response requires similar levels of IL-6 in males and females. However, they also suggest that the adrenals of males and females have different sensitivities to ACTH (higher in females) and possibly also to direct IL-6 stimulation. PMID- 12114261 TI - Mechanisms of glucocorticoid action in bone. AB - Glucocorticoids cause profound effects on bone cell replication, differentiation, and function. Glucocorticoids increase bone resorption by stimulating osteoclastogenesis by increasing the expression of RANK ligand and decreasing the expression of its decoy receptor, osteoprotegerin. In accordance with the increase in bone resorption, glucocorticoids stimulate the expression of collagenase 3 by posttranscriptional mechanisms. The most significant effect of glucocorticoids in bone is an inhibition of bone formation. This is because of a decrease in the number of osteoblasts and their function. The decrease in cell number is secondary to a decrease in osteoblastic cell replication and differentiation, and an increase in the apoptosis of mature osteoblasts. Glucocorticoids decrease osteoblastic function directly and indirectly through the modulation of growth factor expression, receptor binding, or binding protein levels. Clinically, patients with glucocorticoid-induced osteoporosis (GIOP) develop bone loss in the first few months of glucocorticoid exposure, and modest doses of glucocorticoids increase the risk of fractures of the spine and hip. Bisphosphonates inhibit bone resorption and prevent and revert the bone loss that follows glucocorticoid exposure. Anabolic agents, such as parathyroid hormone, stimulate bone formation and can increase bone mass in GIOP. PMID- 12114262 TI - Glucocorticoids in rheumatoid arthritis: effects on erosions and bone. AB - Recently, four prospective placebo-controlled studies have further evaluated the disease-modifying properties of glucocorticoids in the treatment of rheumatoid arthritis. These studies irrefutably show that the use of (low) doses of glucocorticoids leads to a significant retardation of the progression of erosions, especially in early rheumatoid arthritis. This effect on erosions seems more impressive and probably more persistent than the well-known relief during low-dose glucocorticoid therapy of symptoms, such as pain, stiffness, and joint scores. The management of the (side) effects of glucocorticoids on bone has clearly improved in the last years. These two developments lead to a further optimizing of glucocorticoid treatment in patients with rheumatoid arthritis. PMID- 12114263 TI - Cortisol, dehydroepiandrosterone sulfate, and androstenedione levels in patients with polymyalgia rheumatica during twelve months of glucocorticoid therapy. AB - This paper aims to evaluate adrenal gland hormone levels in patients with polymyalgia rheumatica (PMR) during glucocorticoid (GC) therapy. A lower than expected basal production of cortisol was found in active and glucocorticoid untreated PMR patients, particularly females. The abrupt onset of PMR with clinical features similar to those of the steroid-withdrawal syndrome or adrenal insufficiency, as well as the clinical response to GC therapy in elderly people already age-disposed to an inadequate adrenal and anti-inflammatory response, might represent the most significant pathophysiological basis of the disease. PMID- 12114264 TI - Interactions between glucocorticoids and cytokines in the bone microenvironment. AB - Cytokines belonging to the so-called interleukin-6 (IL-6) or gp130 cytokine family, notably IL-6 and IL-11, are known as pro-resorptive cytokines, in that they promote osteoclastogenesis. Glucocorticoid (GC)-induced osteoporosis is admittedly the most frequent secondary osteoporosis. The pathogenesis still has many unresolved issues. Although the effects of GCs on cytokine production and recognition have been extensively studied, little is known about the effects of cytokines on GC action at the target level. We have focused on the effects of IL 6 and IL-11 on specific binding by type II GC receptors (GRs) in two human osteoblast-like cell lines (Saos-2 and MG-63) that have remarkably different constitutive expression of these cytokines and GRs as well. We have provided evidence that IL-6 upregulates GR binding sites, while IL-11 downregulates these sites, as determined by radioligand binding assay and Scatchard analysis. GR affinity (K(d)) did not change after exposure to both cytokines. A number of experiments were consistent with the view that in human osteoblast-like cells, cytokines of the IL-6 family have autocrine modulatory effects on GRalpha (GRbeta is a variant that does not bind specifically in our method). Complex effects of GCs on the system(s) of proinflammatory/anti-inflammatory cytokines and conversely of these cytokines on GC action could account for the dynamics of bone loss in patients given GCs and conceivably having high concentrations of these cytokines in the bone microenvironment. PMID- 12114265 TI - Regulation of leukocyte-endothelial interactions by glucocorticoids. AB - Glucocorticoids (GCs) are steroid molecules endowed with powerful anti inflammatory and immunosuppressive properties. Traditionally, the anti inflammatory action of GC has been largely ascribed to the synthesis of lipocortin-1 (now know as annexin I), while the immunosuppressive effect has been linked to the inhibition of several immune functions and the synthesis of important cytokines and chemokines. In addition to these modes of action, there is a mounting body of evidence suggesting that GCs can also inhibit cell adhesion events, which also play a crucial role in the inflammatory/ immune response. The mechanisms by which GCs modulate cell adhesion are complex and multifactorial. It is now clear that GCs can directly regulate cell adhesion molecule (CAM) gene transactivation through the classical glucocorticoid receptor (GR) pathways. These involve interference with activation/ transcription factors such as AP-1 and NF-kappaB, as well as binding of the GC-GR complex to specific DNA sequences, called glucocorticoid response element "GRE," with ensuing CAM gene inhibition. In addition to these "genomic" mechanisms, there is increasing recognition of alternative modalities of action of GC that are independent from modulating gene expression and for this reason defined as "non-genomic." These are characterized by a rapid response (seconds/minutes) and insensitivity to inhibitors of gene transcription and protein synthesis. The non-genomic effects could be due to direct physicochemical interactions with cell membrane constituents including ion channels and membrane associated proteins. This would lead to inhibition of intracellular signaling pathways involved in CAM activation and cytoskeleton reorganization essential for cell adhesion and locomotion. PMID- 12114266 TI - Corticotropin-releasing hormone signaling in synovial tissue vascular endothelium is mediated through the cAMP/CREB pathway. AB - Modulation of locally produced corticotropin-releasing hormone (CRH) is a component of the cytokine network in human inflammatory arthritis. CRH signaling, through the CRH-receptor subtype R1alpha, may play a role in both vascular changes and pathologic mechanisms associated with joint inflammation. Furthermore, the peripheral actions of CRH may be mediated in part through the NURR subfamily of nuclear orphan receptors. The aim of this study was to establish the signaling mechanisms through which CRH receptor-mediated responses contribute to gene regulation in inflamed synovial vasculature. Immunohistochemical analysis of serial rheumatoid arthritis (RA) tissue sections demonstrates CRH and NURR1 expression in the synovial lining layer, subsynovial lining layer, and the vascular endothelium. The identical pattern of immunolocalization confirms that NURR1 is produced at the same synovial sites shown to produce CRH. The distribution of specific NURR1 staining on the synovial vasculature parallels that observed for CRH-R1 expression. Using primary synovial tissue endothelial cells, we demonstrate that CRH induces specific CREB-1 and ATF 2 binding to the NURR1 promoter. We further provide evidence that CRH signaling can be mimicked by activation of cAMP/PKA/CREB using forskolin in primary human microvascular endothelial cells. These data indicate that the CRH receptor dependent inflammatory response in synovial tissue endothelium is mediated through the cAMP/CREB signaling pathway. PMID- 12114268 TI - Estrogens and the vascular endothelium. AB - Estrogens exert important regulatory functions on vessel wall components, which may contribute to the increased prevalence and severity of certain chronic inflammatory and autoimmune diseases in females and the lower cardiovascular risk observed in premenopausal women. Endothelial cells have been recently identified as targets for estrogens, and estrogen receptors have been demonstrated in endothelial cells from various vascular beds. This review focuses on the regulatory function of estrogens in endothelial cell responses relevant to vessel inflammation, injury, and repair; estrogen effects on nitric oxide production and release; estrogen modulation of endothelial cell adhesion molecule expression; and estrogen regulation of angiogenesis. The mechanisms through which estrogen regulates endothelial cell functions are complex and involve both genomic and nongenomic mechanisms. PMID- 12114267 TI - Androgens and estrogens modulate the immune and inflammatory responses in rheumatoid arthritis. AB - Generally, androgens exert suppressive effects on both humoral and cellular immune responses and seem to represent natural anti-inflammatory hormones; in contrast, estrogens exert immunoenhancing activities, at least on humoral immune response. Low levels of gonadal androgens (testosterone/dihydrotestosterone) and adrenal androgens (dehydroepiandrosterone and its sulfate), as well as lower androgen/estrogen ratios, have been detected in body fluids (that is, blood, synovial fluid, smears, salivary) of both male and female rheumatoid arthritis patients, supporting the possibility of a pathogenic role for the decreased levels of the immune-suppressive androgens. Several physiological, pathological, and therapeutic conditions may change the sex hormone milieu and/or peripheral conversion, including the menstrual cycle, pregnancy, the postpartum period, menopause, chronic stress, and inflammatory cytokines, as well as use of corticosteroids, oral contraceptives, and steroid hormonal replacements, inducing altered androgen/estrogen ratios and related effects. Therefore, sex hormone balance is still a crucial factor in the regulation of immune and inflammatory responses, and the therapeutical modulation of this balance should represent part of advanced biological treatments for rheumatoid arthritis and other autoimmune rheumatic diseases. PMID- 12114269 TI - Relationships between glucocorticoids and gonadal steroids in rheumatoid arthritis. AB - Gender and sex hormones are strongly related to the incidence and progression of autoimmune rheumatic diseases. Although sex steroids have been shown to have direct effects on the immune system, their influence in vivo may be mediated via interactions with third party systems including the hypothalamic-pituitary adrenal axis. Such interactions are well demonstrated in experimental animals. In humans, there is increasing, although indirect, evidence that these interactions also occur. Possible interactions at the cell and gene level, with mutual antagonism or synergy between cortico- and gonadal steroids, open new exciting hypotheses that await clarification. PMID- 12114271 TI - Hormone replacement therapy and chronic graft-versus-host disease activity in women treated with bone marrow transplantation for hematologic malignancies. AB - Several lines of experimental evidence suggest that sex hormones may influence the development and activity of chronic graft-versus-host disease (cGVHD), which frequently occurs in patients undergoing allogeneic bone marrow transplantation (ABMT). Following ABMT, young women are commonly treated with hormone replacement therapy (HRT) because of irreversible gonadal failure. It seemed therefore worthwhile to investigate the effects of this therapy on the activity of cGVHD. Premenopausal women treated with ABMT for hematological malignancies between January 1997 and December 2000 were evaluated for cGVHD activity. They were divided into two groups, depending on whether or not they were treated with HRT. Seventy-one women qualified for the present study: 39 received HRT (treated group), while 32 did not (controls). In both groups of patients, cGVHD activity score was comparable before the start of HRT. No differences were observed in cGVHD activity score between the HRT group and controls after 3, 6, 12, and 24 months from the start of HRT. Furthermore, HRT did not induce any increase in the cGVHD activity score in the treated group of patients at any time from the start of HRT. According to present data, HRT did not appear to influence the activity of cGVHD in young women who underwent ABMT for hematological malignancies. Therefore, we can safely propose this therapy for women with gonadal failure after ABMT. PMID- 12114270 TI - Influence of cytokines and growth factors on distinct steroidogenic enzymes in vitro: a short tabular data collection. AB - Cytokines (IL-1, IL-6, IL-8, IL-11, TNF, IFN-gamma, and TGF-beta) and growth factors (EGF, bFGF, aFGF, and KGF) play an important role in modulation of hormone secretion by directly influencing specific enzyme steps of steroidogenesis in various endocrine cell types. For this tabular data collection, the following enzyme steps were considered: steroidogenic acute regulatory protein (StAR), side chain cleavage enzyme (P450scc), 3 beta hydroxysteroid dehydrogenase, 17-alpha-hydroxylase/17,20-lyase (P450c17), 17-beta hydroxysteroid-dehydrogenase, aromatase complex, 5-alpha-reductase, P450c21, DHEAS sulfatase, and DHEA sulfotransferase. This collection summarizes the current information on how the mentioned cytokines and growth factors influence particular enzyme steps. PMID- 12114272 TI - A role for sex steroids in autoimmune diseases: a working hypothesis and supporting data. AB - In recent years there has been a continuingly increasing interest in novel research subjects, as yet poorly explored, either because they relate to aspects previously thought to be marginal with respect to classical fields of investigation, or because they require both specialized competence and intense cross-talk by researchers from disparate areas. The potential interaction between immunity and cancer has generated a remarkable number of studies, including those related to the newly explored immune-neuro-endocrine system. In this paper, we review a few autoimmune diseases as examples of a mutual relationship between immune diseases and malignancies. We also review our previous studies on patients with rheumatoid arthritis (RA). In particular, aiming to define the hormone responsive or -sensitive status of synovial tissues and cells, we have inspected different endocrine end-points, including (1) high- and low-affinity sites of androgen and estrogen binding; (2) the activity of key enzymes of steroid metabolism; and (3) the hormonal profile of synovial fluids as an indication of local endocrine milieu. Overall, our data provide convincing evidence for synovial macrophage-like cells and a subset of T lymphocytes to be considered as target cells for gonadal steroids. This provides a basis for developing new strategies for alternative treatments of RA and possibly unveils novel perspectives in both research and the clinic for other autoimmune diseases as well. In addition, the association of autoimmunity and cancer may disclose promising new avenues of research linking steroid hormones, the immune system, and malignant transformation. PMID- 12114273 TI - Modulation of cell growth and apoptosis by sex hormones in cultured monocytic THP 1 cells. AB - Several authors have reported the regulation of apoptotic phenomena by sex hormones in different cell lines, including T lymphocytes and mononuclear cells. Since androgens can modulate the programmed cell death in responsive cell lines, we decided to investigate the induction of apoptosis in THP-1 cells following their differentiation into macrophage-like cells and exposure to sex hormones. In addition, we decided to evaluate the proto-oncogene Bax and Fas (CD 95) and cleaved PARP (poly-adp-ribose-polymerase) expression in the same cultured cells. The results showed for the first time the dose-/time-dependent regulation of the apoptotic event in human monocytic THP-1 cells treated with different concentrations of androgens. No significant changes were observed for estrogen treated and unstimulated control cells. In particular, the cells, after stimulation with androgens but not with estrogens, were found to be positive for the proto-oncogene Bax, Fas, and for cleaved subunits of PARP expression as demonstrated with different assays including immunocytochemical assay and Western blot analysis. In conclusion, these results support the possibility of sex hormone modulation of apoptosis in macrophage-like cells, with interesting therapeutic perspectives in rheumatoid arthritis. PMID- 12114274 TI - Androgen deficiency, Meibomian gland dysfunction, and evaporative dry eye. AB - OBJECTIVE: We have recently discovered that women with primary and secondary Sjogren's syndrome are androgen-deficient. We hypothesize that this hormone insufficiency contributes to the meibomian gland dysfunction, tear film instability, and evaporative dry eye that are characteristic of this autoimmune disorder. If our hypothesis is correct, we predict: (1) that androgens regulate meibomian gland function, control the quality and/or quantity of lipids produced by this tissue, and promote the formation of the tear film's lipid layer; and (2) that androgen deficiency, due to an attenuation in androgen synthesis (e.g., during Sjogren's syndrome, menopause, aging, complete androgen-insensitivity syndrome [CAIS] and anti-androgen use), will lead to meibomian gland dysfunction and evaporative dry eye. The following studies were designed to test these predictions. METHODS: Experimental procedures included clinical studies, animal models, and histological, biochemical, molecular biological, and biomedical engineering techniques. RESULTS: Our results demonstrate that: (1) androgens regulate the meibomian gland. This tissue contains androgen receptor mRNA, androgen receptor protein within acinar epithelial cell nuclei, and Types 1 and 2 5alpha-reductase mRNAs. Moreover, androgens appear to modulate lipid production and gene expression in mouse and/or rabbit meibomian glands; and (2) androgen deficiency may lead to meibomian gland dysfunction, altered lipid profiles in meibomian gland secretions, tear film instability, and evaporative dry eye. Thus, we have found that anti-androgen therapy in men is associated with meibomian gland disease, a decreased tear film breakup time, and functional dry eye. Furthermore, we have discovered that androgen receptor dysfunction in women with CAIS is associated with meibomian gland changes and a significant increase in the signs and symptoms of dry eye. Of interest, we have also found that androgen deficiency is associated with significant and striking alterations in the neutral and polar lipid patterns of human meibomian gland secretions. CONCLUSIONS: Our findings show that the meibomian gland is an androgen target organ and that androgen deficiency may promote meibomian gland dysfunction and evaporative dry eye. Overall, these results support our hypothesis that androgen deficiency may be an important etiologic factor in the pathogenesis of evaporative dry eye in women with Sjogren's syndrome. PMID- 12114275 TI - Do estrogen and progesterone play a role in the dry eye of Sjogren's syndrome? PMID- 12114276 TI - Delayed neutrophil apoptosis induced by synovial fluid in rheumatoid arthritis: role of cytokines, estrogens, and adenosine. AB - The fate of neutrophils at sites of inflammation, where these cells are likely exposed to both anti- and proapoptotic influences, needs to be clarified. To investigate this issue, we studied the survival of neutrophils in the presence of articular fluids from RA joints before and after immune complex activation. Eight of eleven samples of RA synovial fluid studied were found to inhibit spontaneous and immune complex-stimulated neutrophil apoptosis. No relationships were found between GM-CSF and TNF-alpha concentrations measured on each sample of synovial fluid studied and the levels of neutrophil apoptosis detectable in the presence of the same synovial fluid. Furthermore, no activity on neutrophil survival was observed at either physiologic or pharmacologic concentrations of estradiol. On the contrary, the synovial fluid anti-apoptotic activity correlates (r(2) = 0.8818, p < 0.0001) with the adenosine detected at concentrations in each sample ranging from 18.7 to 52.4 microM. Finally, synovial fluids were incapable of interfering with neutrophil activation evaluated as superoxide anion production. Our results suggest that the microenvironment of rheumatoid synovial fluid is a proinflammatory milieu responsible for the in loco persistence of activated and long-surviving neutrophils. PMID- 12114278 TI - Post-menopause is the main risk factor for developing isolated pulmonary hypertension in systemic sclerosis. AB - In scleroderma patients, isolated pulmonary hypertension (PHT) has been associated with selected HLA haplotypes, severe impairment of the diffusing capacity for carbon monoxide and the diagnosis of CREST. Most patients with CREST have a late-age onset of the disease, corresponding to the perimenopausal or postmenopausal period. We conducted a retrospective cohort study to determine the role of post-menopause and of the other known clinical and biological markers in the development of isolated pulmonary hypertension in Italian patients with systemic sclerosis. 189 female patients with scleroderma who had no ecographic signs of pulmonary hypertension (PHT) and radiographic signs of lung fibrosis at the first visit and did not develop significant pulmonary fibrosis during the observation time were included. Sixty-three out of 189 patients (33.3%) presented isolated pulmonary hypertension. A severe impairment of diffusing capacity for carbon monoxide at admission was found to be an early predictive element for its development. An increased risk was associated with postmenopausal condition (RR = 5.2, p = 0.000), CREST syndrome (RR = 2.8, p = 0.001) and haplotype HLA-B35 (RR = 2.8; p = 0.002). A significant positive interaction between postmenopausal condition and either HLA-B35 (RR = 15.2; p = 0.000) or the diagnosis of CREST (RR = 14.1; p = 0.000) was found. Postmenopausal condition alone or in combination with HLA-B35 and CREST syndrome is the main risk-factor for developing primary pulmonary hypertension in scleroderma patients. This suggests that hormonal replacement therapy could play a role in preventing isolated PHT in patients with systemic sclerosis. PMID- 12114277 TI - Antiproliferative-antiinflammatory effects of methotrexate and sex hormones on cultured differentiating myeloid monocytic cells (THP-1). AB - Methotrexate (MTX) is believed to exert both antiproliferative and antiinflammatory effects in a dose-related manner in a majority of rheumatoid arthritis (RA) patients along with an abrupt flare of the disease after drug discontinuation. To investigate the antiproliferative and antiinflammatory actions of MTX and the combined action of sex hormones, we evaluated these effects in differentiated monocytic myeloid cells (THP-1) prestimulated with testosterone (T) or 17-beta estradiol (E2). The effects of MTX and T combined treatment (T/MTX) on THP-1 cells showed a significant inhibition of cell proliferation when compared with E2/MTX- treated cells or controls: 53% at 72 h versus E2-treated cells; 58% at 96 h versus E2-treated cells; and 41% versus controls, respectively. Bax and Fas CD95 expression was found increased in T treated cells: 14% T at 48 h vs. E(2)-treated cells and controls; 45% T at 72 h versus E2-treated cells and controls; 97% at 96 h versus E2-treated cells and 37% versus controls for Bax: 33%, 41%, and 42% T versus E2-treated cells for Fas. Moreover, a significant decrease of IL-12 levels in T/MTX treated cells was found at any time when compared to E2-treated cells. In summary, the association of testosterone and MTX compared to MTX alone suggests possible synergistic actions. Therefore, the enhancing antiinflammatory effects exerted by androgens might represent a further explanation for the reduced frequency of inflammatory diseases in male subjects. PMID- 12114279 TI - Effects of stress on serum prolactin levels in patients with systemic lupus erythematosus. AB - The results of a study of effects of stress on serum prolactin (PRL) levels in patients with SLE reveal statistically significant differences in serum PRL level readings in samples taken over a short period of time, thus corroborating the need to take into account PRL stress induction during sample withdrawals and to interpret the values obtained, especially where moderate idiopathic hyperprolactinemia was detected. To eliminate any external stress factors, it is advisable to take PRL samples repeatedly and in perfectly resting patients. PMID- 12114280 TI - Prolactin and IgG-prolactin complex levels in patients with rheumatic arthritis. PMID- 12114281 TI - Serum prolactin concentrations in male patients with rheumatoid arthritis. AB - Altered serum prolactin (PRL) levels have been reported in autoimmune diseases; however, data of serum PRL concentrations in rheumatoid arthritis (RA) are contradictory. We evaluated the PRL status in men affected by RA and the possible relationships among serum PRL levels, bone mass, and disease activity. We investigated 29 men affected by RA and 30 age- and sex-matched controls. All patients were evaluated for serum PRL levels, parameters of disease activity, and bone mineral density (BMD) at L2-L4 and the femoral neck. Serum PRL levels were found significantly higher in men with RA than in controls (p = 0.001). High serum PRL levels were significantly correlated with duration of RA and some laboratory parameters of RA disease activity. A negative correlation between femoral BMD and serum PRL levels were found (r = -0.821, p = 0.001). Male patients affected by RA showed high serum PRL levels. The serum PRL concentration was found to be increased in relation to the duration and the activity of the disease. Serum PRL levels do not seem to have any relationship with the BMD, at least in RA. PMID- 12114282 TI - Somatotropic, lactotropic and adrenocortical responses to insulin-induced hypoglycemia in patients with rheumatoid arthritis. AB - Neuroendocrine mechanisms have been suggested to play an important role in the onset and progression of rheumatoid arthritis (RA). The aim of this study was to evaluate hypothalamic-pituitary functions in RA patients by measurement of hormone responses to insulin-induced hypoglycemia. Insulin-hypoglycemia (Actrapid HM 0.1 IU/kg, i.v. as a bolus) was induced in 17 male patients and in 11 age-, gender-, and weight-matched healthy subjects. Concentrations of growth hormone (GH), prolactin (PRL) and cortisol were analyzed in plasma. PRL release after thyreoliberin stimulation (TRH, 200 g, i.v.) was determined in 21 patients with active forms of RA and in 12 control subjects to evaluate pituitary lactotropic response. In RA patients, basal concentrations of glucose, GH, PRL, and cortisol were in the normal range and they were comparable to those in the control group. Stress of hypoglycemia induced significant elevation of GH, PRL, and cortisol concentrations in all groups. Cortisol responses to hypoglycemia were comparable in patients and in control subjects. GH release during hypoglycemia was increased (p < 0.05) and PRL response was attenuated (p < 0.05) in RA patients versus control subjects. After TRH administration, PRL response was the same in patients as in healthy subjects. In conclusion, the present study revealed an altered hypothalamic-pituitary function in patients with RA, namely, an enhanced somatotropic and reduced lactotropic activation in response to insulin-induced hypoglycemia. Basal hormone levels and cortisol release during hypoglycemia were similar to those in healthy subjects. PMID- 12114283 TI - Melatonin in rheumatoid arthritis: synovial macrophages show melatonin receptors. AB - The pineal neurohormone melatonin is widely recognized as exerting important immunoenhancing effects that act on specific receptors in immunocompetent cells. This action results in stimulation of cytokine production in lymphocytes and macrophages. Here we report that the nocturnal plasma concentration of melatonin in rheumatoid arthritis (RA) patients is higher than in healthy controls. Furthermore, melatonin is present in the synovial fluid of RA patients and synovial macrophages express a specific binding site. We suggest that melatonin may exert a disease-promoting role in RA. PMID- 12114284 TI - Melatonin serum levels in rheumatoid arthritis. AB - The pineal hormone melatonin (MLT) exerts a variety of effects on the immune system. MLT activates immune cells and enhances inflammatory cytokine and nitric oxide production. Cytokines are strongly involved in the synovial immune and inflammatory response in rheumatoid arthritis (RA) and reach the peak of concentration in the early morning, when MLT serum level is higher. Nocturnal MLT serum levels were evaluated in 10 RA patients and in 6 healthy controls. Blood samples were obtained at 8 and 12 p.m., as well as at 2, 4, 6, and 8 a.m. MLT serum levels at 8 p.m. and 8 a.m. were found to be higher in RA patients than in controls (p < 0.05). In both RA patients and healthy subjects, MLT progressively increased from 8 p.m. to the first hours of the morning, when the peak level was reached (p < 0.02). However, MLT serum level reached the peak at least two hours before in RA patients than in controls (p < 0.05). Subsequently, in RA patients, MLT concentration showed a plateau level lasting two to three hours, an effect not observed in healthy controls. After 2 a.m., MLT levels decreased similarly in both RA patients and healthy subjects. Several clinical symptoms of RA, such as morning gelling, stiffness, and swelling, which are more evident in the early morning, might be related to the neuroimmunomodulatory effects exerted by MLT on synovitis and might be explained by the imbalance between cortisol serum levels (lower in RA patients) and MLT serum levels (higher in RA patients). PMID- 12114286 TI - Stress hormones, proinflammatory and antiinflammatory cytokines, and autoimmunity. AB - Recent evidence indicates that glucocorticoids and catecholamines, the major stress hormones, inhibit the production of proinflammatory cytokines, such as interleukin (IL)-12, tumor necrosis factor (TNF)-alpha, and interferon (IFN) gamma, whereas they stimulate the production of antiinflammatory cytokines, such as IL-10, IL-4, and transforming growth factor (TGF)-beta. Thus, systemically, an excessive immune response, through activation of the stress system, stimulates an important negative feedback mechanism, which protects the organism from an "overshoot" of proinflammatory cytokines and other products of activated macrophages with tissue-damaging potential. Conversely, in certain local responses and under certain conditions, stress hormones actually may boost regional immune responses, through induction of TNF-alpha, IL-1, and IL-8, and by inhibiting TGF-beta production. Therefore, conditions that are associated with significant changes in stress system activity, such as acute or chronic stress, cessation of chronic stress, severe exercise, and pregnancy and the postpartum period, through modulation of the systemic or local pro/antiinflammatory cytokine balance, may suppress or potentiate autoimmune diseases activity and/or progression. PMID- 12114285 TI - Melatonin participates in the control of testosterone secretion from rat testis: an overview of our experience. AB - The androgen milieu plays an important role in the control of immunity, and melatonin (MLT) is known to modulate the immune response as well. Our recent studies have focused on the role of MLT in controlling androgen secretion from rat testis. The presence of MLT receptors, their modulation, and the effect of MLT on steroidogenesis have been studied on Percoll-purified rat Leydig cells (LCs) cultured in vitro. MLT receptors present in adult rat LCs (B(max) = 46.7 +/ 3.5 fmol/mg protein; K(d) = 88.7 +/- 6.2 pmol/l) are coupled through a pertussis toxin-sensitive G-protein and are downregulated by prolonged exposure to MLT itself. When acutely added for three hours, MLT (0.4-400 nM) inhibits T secretion in response to LH, forskolin, and GnRH, but sensitizes cAMP-dependent T secretion when present in the preincubation media (16 h). The acute inhibition of adenylyl cyclase and the block of 17-20-desmolase (with increased 17-OH-progesterone) and of calcium release from the intracellular stores, followed by sensitization of the adenylyl cyclase activity in the long term, are described as mechanisms of MLT action. Ultimately, our studies suggest that MLT is likely to control the immune response not only through its direct effects, but also by finely modulating the androgen milieu. PMID- 12114287 TI - Role of interleukin-6 in stress response in normal and tumorous adrenal cells and during chronic inflammation. AB - Interleukin-6 (IL-6) is the end-product of a cytokine signaling cascade and is secreted by specialized immune cells during inflammation. It has a great influence on many functions, including differentiation, stimulation, and activation of immune cells, or other cells of neuroendocrine origin. Thus, IL-6 serves as a key messenger in its communication with the neuroendocrine system, and serves as a potent activator of the hypothalamic-pituitary-adrenal axis at all levels. Changes in the levels of expression of this cytokine and its receptor have been observed during chronic inflammatory disease, and have been associated with tumorigenesis. Therefore, we studied the effect of IL-6 on normal and adenomatous human adrenal cells in vitro. The expression of IL-6 receptor mRNA was quantified within the same tissue. IL-6 potently stimulated cortisol secretion from dispersed normal human adrenal cells. We found immunoreactivity for the IL-6 receptor on cultured cells and paraffin-embedded sections of adrenal tissues. Further, there was a more pronounced expression of IL-6 mRNA in adrenal adenomas of patients with Cushing's syndrome, compared to normal human adrenals. Despite this fact, the sensitivity of cells of adenomatous adrenal glands to IL-6 was significantly decreased relative to cells from normal controls. These results were confirmed employing the permanent adrenocortical cancer cell line model NCI H295. We infer that the loss of responsivity of tumorous adrenal cells to IL-6, and in part corticotropin, is an important step in the process of adrenal tumorigenesis by which regulation by differentiating proteins is bypassed. PMID- 12114288 TI - Effects of chronic food restriction stress and chronic psychological stress on the development of adjuvant arthritis in male long evans rats. AB - The aim of the present experiments was to study the effect of stress of chronic food restriction (FR) and of repeated psychological challenge (PS) on the development of adjuvant arthritis in the Long Evans male rats. In the FR series, four groups of animals were compared: non-treated control (C) and arthritic (AA) rats, both with free access to food and water, and two analogous groups with a 40% food restriction-FR and AA-FR. All animals were killed 22 days after injection of cFA. In the PS series, stress was induced by random daily exposures of the rats to isolation, over-crowding, food/water deprivation, foot shock, tilting, fear for 14 days before cFA injection and 12 days thereafter (groups: C, AA, PS, and AA-PS). Arthritis causes swelling of the hindpaw, which was prevented in the AA-FR group. PS causes more severe disease symptoms: AA-PS rats had more severe hindpaw swelling than AA rats. Forty percent food restriction associated with elevated CORT levels mitigated inflammatory parameters activated during AA. PS worsened the disease. These results suggested that activated CORT is not the only cause of disease suppression, but some metabolic changes during FR play a role. PMID- 12114289 TI - Fatigue in daily life in patients with primary Sjogren's syndrome and systemic lupus erythematosus. AB - Fatigue is a well-recognized complaint with major impact on daily life in primary Sjogren's syndrome (PSS) and systemic lupus erythematosus (SLE). Previous research has not taken into account several crucial aspects of fatigue. This study examined various aspects of fatigue in the daily life of patients with PSS and SLE and in healthy controls. We compared age-adjusted, repeated measurements of fatigue across the day of female patients with SLE (n = 20, mean age 43.4 +/- 11.3), with PSS (n = 28, mean age 53.7 +/- 13.9) and healthy participants (n = 30, mean age 50.5 +/- 13.4). General and physical fatigue was significantly higher in patients than in healthy participants. Groups did not differ with respect to average levels of reduced motivation or mental fatigue. Both general and physical fatigue and reduced activity varied significantly during the day. Adjusting for depressive symptoms, groups showed significantly different time courses during the day. In healthy participants and patients with SLE, fatigue first decreased and then increased, whereas a rather opposite course-at least for the first part of the day-was observed in patients with PSS. Using an ecologically valid assessment method, we demonstrated substantially higher levels of daily fatigue in SLE and PSS patients as compared to healthy participants, thereby jeopardizing these patients' quality of life. The effect of disease on variations in fatigue over the day should be the subject of further inquiry, especially as it might clarify underlying mechanisms. PMID- 12114290 TI - Immunological responses are not abnormal in symptomatic Gulf War veterans. AB - The underlying etiology and pathogenesis of Gulf War veterans' illnesses continue to be under intense investigation. Reports have suggested the basis for these illnesses may be an altered immune system, but compelling evidence is lacking. We sought to determine whether in vitro immune responses were abnormal in symptomatic Gulf War veterans relative to matched controls. A randomized case control study was conducted by blinded comparison of laboratory measures of in vitro immune responses in blood samples obtained from veterans in an outpatient facility of a Veterans Affairs medical center. Symptomatic Gulf War veterans with otherwise undefined illnesses (52 symptomatic subjects), asymptomatic Gulf War veterans (31 asymptomatic controls), and veterans who had applied for disability compensation and had not participated in the Gulf War (21 disability controls) represented the volunteer sample. In vitro cellular and humoral immune responses were measured to detect functional abnormalities in antigen presenting cells (autologous mixed leukocyte reactions and expression of interleukin (IL)-1beta, IL-6, IL-10, and tumor necrosis factor-alpha); T cells (lymphocyte proliferation using the polyclonal T-cell activators phytohemagglutinin and Concanavalin A; primary immune responses in allogeneic mixed leukocyte reactions; secondary immune response using the recall antigens tetanus toxoid, Candida albicans, and anthrax vaccine; and soluble IL-2 receptor expression); type-1 T-helper cells (gamma interferon expression); type-2 T-helper cells (IL-4 and IL-10 expression); and B cells (polyclonal B-cell activator pokeweed mitogen-induced immunoglobulin production). In general, immune response measures did not differ significantly between groups. Heightened responses observed in the disability control group (sporadically greater responses to one mitogen and two antigens) and the Gulf War participation control group (greater recall responses to anthrax vaccine) did not suggest impaired immune cell function in symptomatic veterans when compared with controls. We conclude that in vitro immunological responses are not abnormal in symptomatic Gulf War veterans. PMID- 12114291 TI - Mechanisms of pain in arthritis. AB - Inflammation in the joint causes peripheral sensitization (increase of sensitivity of nociceptive primary afferent neurons) and central sensitization (hyperexcitability of nociceptive neurons in the central nervous system). The processes of sensitization are thought to be the basis of arthritic pain that appears as spontaneous pain (joints at rest) and hyperalgesia (augmented pain response on noxious stimulation and pain on normally nonpainful stimulation). Sensitization also facilitates efferent neuronal processes through which the nervous system influences the inflammatory process. Peripheral sensitization is produced by the action of inflammatory mediators such as bradykinin, prostaglandins, neuropeptides, and cytokines which activate corresponding receptors in proportions of nerve fibers. In addition, the expression of receptors, for example, bradykinin and neurokinin 1 receptors, is upregulated during inflammation. The development of hyperexcitability of spinal cord neurons is produced by various transmitter/receptor systems that constitute and modulate synaptic activation of the neurons. The key transmitter is glutamate that activates N-methyl-d-aspartate (NMDA) and non-NMDA receptors on spinal cord neurons. Blockade of these receptors prevents and reduces central sensitization. Excitatory neuropeptides (substance P and calcitonin gene-related peptide) further central sensitization. Central sensitization also is facilitated by mediators that have complex actions (e.g., prostaglandin E(2)). Spinal PGE(2) binds to receptors at presynaptic endings of primary afferent neurons (thus influencing synaptic release) and to receptors on postsynaptic spinal cord neurons. The administration of PGE(2) to the spinal cord surface produces changes of responsiveness of spinal neurons similar to peripheral inflammation, and spinal indomethacin to the spinal cord attenuates development of hyperexcitability significantly. PMID- 12114293 TI - Effects of norepinephrine on oxygen consumption of quiescent and activated human peripheral blood mononuclear cells. PMID- 12114292 TI - Immunopathogenesis of rheumatic diseases in the context of neuroendocrine interactions. AB - Growing evidence supports the hypothesis that alterations of the stress response and interactions between the neuroendocrine and immune systems contribute to the pathogenesis of rheumatic diseases such as rheumatoid arthritis (RA). In particular, the hypothalamus-pituitary-adrenal (HPA) axis and the autonomic nervous system (ANS) are of special interest. Polymorphisms of the corticotropin releasing hormone (CRH)-regulating region have been described recently. These polymorphisms are differentially distributed in RA patients and healthy subjects of various ethnic origin, thus supporting the hypothesis that they represent a new genetic marker for RA susceptibility. The decreased expression of beta(2) adrenergic receptors (beta(2)-R) on lymphatic cells in rheumatic diseases like RA, together with an impaired influence of catecholamines on immune function in these patients, further underlines the concept of a dysfunction of the ANS in rheumatic diseases. Results from work in this field will provide more insight into the pathogenesis of RA and help to establish novel therapies for this chronic rheumatic disease. PMID- 12114294 TI - Involvement of the hypothalamic-pituitary-adrenal axis in children with oligoarticular-onset idiopathic arthritis. AB - Adult patients with rheumatic arthritis and other rheumatic disorders show inappropriate cortisol secretion and peculiar CRH promoter gene polymorphisms. So far, no data are available about this topic in children with juvenile idiopathic arthritis (JIA). We have studied a series of 13 prepubertal patients (10 female, 3 male) affected with oligoarticular JIA (o-JIA) without clinical and biological signs of disease activity (ESR and IL-6). ACTH plasma concentrations were significantly increased at 8 a.m. in o-JIA patients, whereas no differences were found in cortisol plasma concentrations. The ACTH/cortisol ratio was significantly increased in o-JIA patients with respect to the normal population both at 8 a.m. and at noon. DHEAS and testosterone plasma concentration did not statistically differ in the two populations. The genetic study was aimed at defining the prevalence of polymorphisms A1 and A2 in o-JIA patients, but we failed to find allelic or genotypic differences. Our study suggests the presence of a partial resistance to ACTH with a dysregulated pattern of secretion also in inactive o-JIA patients. These preliminary data need further confirmation in larger pediatric studies. PMID- 12114295 TI - Corticotropin-releasing factor modulates cardiovascular and pupillary autonomic reflexes in man: is there a link to inflammation-induced autonomic nervous hyperreflexia? AB - In two recent studies, we found autonomic nervous hyperreflexia in subjects with chronic inflammatory diseases, particularly, in those subjects with higher degrees of systemic inflammation. Since corticotropin-releasing factor (CRF) is induced by inflammatory stimuli and acts within the brain to change neuroendocrine and autonomic activity, we investigated CRF modulation of standard autonomic nervous reflexes. Fifteen healthy subjects were administered 100 microg CRF i.v., which led to a short-term increase of heart rate (p < 0.001) and a decrease in systolic and diastolic blood pressure (p < 0.050). These changes were accompanied by a reduction in heart rate variation at rest (p = 0.010) and during the respiratory sinus arrhythmia test (p = 0.019), and a reduction of pupillary latency time (p = 0.038). In further 21 normal subjects we studied the effect of prednisolone treatment on autonomic nervous function (100 mg/d during three days -> CRF reduction), which resulted in an increase of heart rate (p < 0.001), increase of heart rate variation during the respiratory sinus arrhythmia test (p < 0.001), increase in pupillary latency time (p = 0.012), a increase in maximal pupillary area (p = 0.002), and a decrease in diastolic blood pressure (p = 0.002). In conclusion, induction of a low central CRF content by prednisolone leads to a marked hyperreflexia in respiratory sinus arrhythmia and hyporeflexia in the latency time test. It is obvious that CRF can regulate autonomic reflexes. Possibly, central CRF status may also influence autonomic reflexes during chronic inflammation due to chronically changed central CRF concentration. PMID- 12114296 TI - Neuropeptides in experimental and degenerative arthritis. AB - Classical symptoms of both inflammatory and degenerative arthritides may contribute to neurogenic responses like wheal, flare, edema, and pain. Rheumatoid arthritis (RA) is an autoimmune disease with an immunogenetic background. Neurogenic inflammation has been considered to play an essential role in RA, in part because of the symmetrical involvement (cross-spinal reflexes) and the predominant involvement of the most heavily innervated small joints of the hands and the feet (highly represented in the hominiculus). In contrast, osteoarthritis (OA) is considered to arise as a result of degeneration of the hyaline articular cartilage, which secondarily results in local inflammation and pain. However, it is possible that the age-related and predominant (compared to nociceptive nerves) degeneration of the proprioceptive, kinesthetic and vasoregulatory nerves can represent the primary pathogenic events. This leads to progressive damage of tissue with extremely poor capacity for self-regeneration. Inflammation, be it primary/autoimmune or secondary/degenerative, leads to peripheral sensitization and stimulation, which may further lead to central sensitization, neurogenic amplification of the inflammatory responses and activation of the neuro-endocrine axis. Neuropeptides serve as messengers, which modulate and mediate the actions in these cascades. Accordingly, many neuropeptides have been used successfully as experimental treatments, most recently VIP, which effectively controlled collagen induced arthritis in mice. Therefore, it can safely be concluded that better treatment/control of disease activity and pain can be achieved by blocking the cascade leading to initiation and/or amplification of inflammatory process combined with effects on central nociceptive and neuroendocrine responses. PMID- 12114297 TI - Psychoneuroimmunology: new avenues of research for the twenty-first century. AB - Psychoneuroimmunology (PNI) investigates the relations between the psychophysiological and immunophysiological dimensions of living beings. PNI brings together researchers in a number of scientific and medical disciplines, including psychology, the neurosciences, immunology, physiology, pharmacology, psychiatry, behavioral medicine, infectious diseases, and rheumatology. All are scientists with profound interest in interactions between the nervous and immune systems, and the relation between behavior and health. Despite the variety in domains and approaches to research, the outcome common to all research endeavors is the discovery of new information, of uncovered facts, of novel evidence, which contributes to the continuing generation of knowledge. In this paper we discuss psychoneuroimmune aspects of some conditions that are not routinely immediately associated with immunity, such as the condition of being the caregiver of somebody suffering from dementia; the effect on the brain-body modulations of aluminum, a metal that is not a component of the human body; and insomnia, a fairly common but disturbing disease, that even today lacks an effectual treatment. PMID- 12114298 TI - Brain functional involvement by perfusion SPECT in systemic sclerosis and Behcet's disease. AB - Involvement of the central nervous system is frequent in systemic immune-mediated diseases, such as systemic lupus erythematosus, Behcet's disease (BD), and systemic sclerosis (SSc). Structural brain examinations, such as computed tomography and magnetic resonance imaging, may be useful in diagnosing and following-up these cerebral syndromes in some cases, but they are more often inconclusive. Single photon emission computed tomography (SPECT) with perfusion tracers is a powerful method that can disclose brain involvement in many clinical situations, even in patients with subtle neurological symptoms. In fact, perfusion tracers can disclose regional hypoperfusion caused by both ischemia due to vascular narrowing and neuronal metabolic derangement due to direct neuronal damage. The latter phenomenon occurs because the blood flow to the brain is strictly regulated by metabolic demands, so that hypometabolism is reflected by hypoperfusion in most instances. SPECT findings in 42 mainly neurologically asymptomatic patients with SSc and in eight mainly neurologically symptomatic patients with BD are reported. SPECT was shown to be very sensitive and disclosed brain functional deficits in approximately half of SSc patients without neurological complaints and in all patients with BD, who had various neurological symptoms but usually inconclusive structural brain examinations. PMID- 12114299 TI - Neurological and neuroendocrine-cytokine inter-relationship in the antiphospholipid syndrome. AB - Although many neurological deficits have been described in the antiphospholipid syndrome (APS), only stroke is well established and accepted as a diagnostic criterion in the disease. We presently review clinical data obtained from large series of cases regarding stroke, dementia, epilepsy, chorea, migraine, white matter disease, and behavioral changes in APS, or linked-to-laboratory criteria such as antiphospholipid antibodies (aPL). The contribution of animal models to our understanding of these manifestations of APS is stressed, especially regarding the cognitive and behavioral aspects for which we have established model systems in the mouse. These models utilize immunization of mice with beta(2)-glycoprotein1, a central autoantigen in APS, which induces persistent high levels of aPLs. These mice develop hyperactive behavior after a period of four months, as well as deficits in learning and memory, and are potentially valuable as a system in which to study the pathogenesis and treatment of cognitive and behavioral aspects of APS. We have developed another model, in which IgGs from APS patients induce depolarization of brain synaptoneurosomes, and which may serve as a model for the pathogenesis of epilepsy in APS. Hormonal changes are another potential CNS manifestation of APS and this may be potentially linked to the systemic and central effects of cytokines such as interleukin-3. Better understanding of the link between APS and neurological or neuroendocrine manifestations other than stroke will reveal whether they can be used as clinical criteria for the diagnosis of APS and, it is hoped, lead to better treatment. PMID- 12114300 TI - Decreased catecholamine-induced cell death in B lymphocytes from patients with rheumatoid arthritis. AB - The expression of beta2-adrenergic receptors (beta2-R) on B lymphocytes and agonist-induced cAMP production is reduced in patients with rheumatoid arthritis (RA). To further study functional consequences of the diminished beta2-R density on B lymphocytes in RA patients, agonist-induced cell death was evaluated and compared to healthy controls. B lymphocytes from patients with RA and healthy controls were activated with anti-IgM-antibody. Coincubation was carried out with isoprenaline (iso, 0.001-10 microM). Apoptotic and necrotic cells were determined using Annexin-V and propidium-iodide staining. beta2-R-induced cell death in B cells from healthy volunteers was stimulated after 24 h (medium, 21.2 +/- 1.6%; iso, 34.6 +/- 4.4%; increase 59.3 +/- 10.1%). However, in RA patients the increase in cell death following beta2-R stimulation (21.8 +/- 8.9%) was significantly impaired (p = 0.02). Our data demonstrate that catecholamine induced cell death after stimulation of beta2-R on B lymphocytes is decreased in RA patients, possibly contributing to the pathogenesis of the disease. PMID- 12114301 TI - Psychoneuroimmunology in oral biology and medicine: the model of oral lichen planus. AB - Rheumatoid arthritis involves psychoneuroendocrine-immunopathological comorbidities. In the stoma, patients with rheumatoid arthritis frequently show signs of periondontal disease consequent to elevated levels of crevicular proinflammatory cytokines. It is not clear whether rheumatoid arthritis may manifest in association with immunopathological manifestations of the oral soft mucosa. Oral lichen planus (OLP), first described by E. Wilson in 1859, is a T cell-mediated inflammatory disease whose lesions characteristically lack B cells, plasma cells, immunoglobulin. or complement. It is increasingly well characterized and recognized as a model for psychoneuroimmunology research in oral biology and medicine. To date, we have shown an association between changes in hypothalamic-pituitary-adrenal (HPA) regulation, systemic markers of cellular immunity and mood states, with clinical stages of OLP (i.e., atrophic vs. erosive vs. bullous lesions). We report significant associations (p < 0.05) between the stage of OLP, HPA deregulation, and altered distribution and functional responses of naive CD4(+) cells. We emphasize the need to study in greater details the psychoneuroendocrine-immune inter-relationships in OLP, and we propose a novel neuroimmune hypothesis for OLP. PMID- 12114302 TI - Expression of IL-15 and IL-15 receptor isoforms in select structures of human fetal brain. AB - IL-15, a key cytokine linking innate and acquired immunity, is expressed in many cell types and tissues. Recent data indicate constitutive expression of IL-15 in human neural cell lines and tissues. The aim of the present study was to examine the expression patterns of mRNA encoding IL-15 and IL-15 receptor alpha (IL 15Ralpha) isoforms in select structures of human fetal brain. We report that mRNA for IL-15 and IL-15Ralpha isoforms were expressed in all tested brain structures: cerebral cortex, cerebellum, hippocampus, and thalamus. However, the levels of IL 15 and IL-15Ralpha mRNA were higher in the hippocampus and cerebellum in comparison with cortex and thalamus. Moreover, higher levels of cytosol in comparison with membrane-bound IL-15 isoform were present in all brain structures. The constitutive, but distinct, expression of IL-15 and its receptors in select human fetal brain structures suggests that IL-15 plays a role in their development and physiology. PMID- 12114303 TI - The use of pupillometry in joint and connective tissue diseases. AB - The central and peripheral nervous systems are variably affected in the rheumatic diseases. Automated standardized infrared pupillometry allows the safe, noninvasive assessment of the pupillary innervation. Pupillometry has already been used in studying the autonomic nervous system (ANS) in various rheumatic diseases. In systemic lupus erythematosus, the irideal parasympathetic branch of ANS was more affected then the sympathetic branch. In Sjogren's syndrome, signs of pupillary parasympathetic denervation have been reported. In rheumatoid arthritis, pupil parasympathetic dysfunction has been shown to correlate with ocular dryness. In systemic sclerosis (SSc), both sympathetic and parasympathetic irideal impairment have been demonstrated. Beside providing autonomic innervation, sensory nerves fibers are able to control iris diameter. Exogenous ocular instillation of substance P (SP), a sensory neuropeptide, can determine an omathropine-resistant, non-cholinergic myosis, acting on specific receptors present on the iris sphincter muscle. We first studied pupillary SP-ergic responsiveness in SSc, evaluating substance P (SP)-stimulated pupillary diameters by pupillometry. A higher basal and SP-stimulated myosis was found in lSSc versus both dSSc and controls, whereas no differences existed between dSSc and controls. From the literature, the pupillary parasympathetic nervous system seems to be more affected than the sympathetic branch of ANS in the rheumatic diseases characterized by an inflammatory status. However, we found in SSc both sympathetic and parasympathetic pupil control to be equally impaired. From our experience, we conclude that pupillary nervous control is differently affected in the two subsets of SSc, and that the SP-ergic system seems to be impaired only in lSSc. PMID- 12114304 TI - Opioid peptides endomorphin-1 and endomorphin-2 in the immune system in humans and in a rodent model of inflammation. AB - Endomorphin (EM)-1 and EM-2 are tetrapeptides with high affinity and selectivity for the micro-opioid receptor. We have utilized specific radioimmunoassays to characterize EM-1 and EM-2 in immune tissues from normal human subjects and from rats with adjuvant arthritis (AA). PBLs from three normal human subjects contained 248, 13, and 303 pg EM-1 per 100 million cells, whereas EM-2 was measured in two subjects at 69 and 588 pg per 100 million cells. In AA rats, EM-1 (but not EM-2) contents in the spleen and thymus were elevated compared with levels in tissues from non-AA controls. EM-1 was detectable in five of eight samples of synovial tissue from inflamed hind paws, whereas EM-2 was detectable in two of eight synovial extracts. Neither EM-1 nor EM-2 were detectable in synovial tissue from non-AA rats. To our knowledge, this is the first report of endomorphins in normal human immune cells. Increased endomorphin expression or uptake in peripheral tissues in a rodent model of chronic inflammation provides potential for endomorphins to selectively modulate chronic inflammation in mammals. PMID- 12114305 TI - Cytokines, acute-phase proteins, and hormones: IL-1 and TNF-alpha production in contact-mediated activation of monocytes by T lymphocytes. AB - The cytokine network is a homeostatic system that has to be perceived in an analogous fashion to the acid/base equilibrium. The level of any cytokine in biological fluids can be interpreted correctly only by taking into account the levels of other synergistic cytokines, of their respective inhibitors, and of each cytokine receptor. Due to their potent activities in many different processes (including cell growth and differentiation, development, and repair processes leading to the restoration of homeostasis), the cytokine activities have to be tightly controlled by natural inhibitory mechanisms. Since one of the main functions of cytokines is to mediate interactions between the immune and inflammatory system, it is thought that chronic immuno-inflammatory diseases might be caused in part by the uncontrolled production of cytokines. Depending on the stage of inflammation or the biological effect determined, the same cytokine might be pro- or anti-inflammatory. This applies, for instance, to IL-4, IL-10, and TGFbeta. An important mechanism that triggers the production of pro inflammatory cytokines in chronic inflammatory diseases is the direct cellular contact between stimulated T cells and monocyte-macrophages. This mechanism is blocked at the systemic level by the "negative" acute-phase protein apolipoprotein A-I (apo A-I). The levels of expression of cytokines and cytokine inhibitors and acute-phase proteins are ruled by hormones. Estrogens as well as androgens inhibit the production of IL-1beta and TNF-alpha on monocyte macrophages. However, androgens antagonize estrogen stimulatory effects on apo A I synthesis by the liver. Other studies suggest that estradiol is more inhibitory to Thl cytokines (e.g., IFNgamma, IL-2), while testosterone is inhibitory to Th2 cytokines (e.g., IL-4). Cytokines also control the axis of the hypothalamic hypophyseal-adrenal glands as well as the sexual hormones. The discrepancy between studies would suggest that the mechanisms are different in physiological and pathophysiological conditions. PMID- 12114307 TI - Carotid atherosclerosis in patients with rheumatoid arthritis: a preliminary case control study. AB - A preliminary study was undertaken to investigate the severity of atherosclerotic lesions of neck arteries in patients with rheumatoid arthritis (RA) in comparison with matched controls. The working hypothesis derives from the recent view that the atherosclerotic plaque is essentially an inflammatory lesion. Thus, patients affected by a chronic inflammatory disease, such as RA, might show acceleration of atherosclerosis despite treatment with anti-inflammatory drugs. In 19 patients with RA the prevalence of atherosclerosis of internal carotid arteries, as seen on high-resolution duplex-scanning of neck arteries, was significantly (p < 0.05) higher than in 19 controls, whereas no significant difference was found for the other arteries. Factors underlying this finding are not known; increased levels of homocysteine and other thiol compounds that may enhance atherosclerosis in RA patients deserve further investigation. Moreover, more specific factors of RA are currently being investigated. In fact, immune complexes fixing C1q bind to endothelial receptors, thus triggering an upregulation of adhesion molecules, such as E-selectin and intercellular and vascular cell adhesion molecules 1, on the endothelium surface. PMID- 12114306 TI - Osteoporosis with underlying connective tissue disease: an unusual case. AB - A 44-year-old male was initially seen by dermatologists, who noted an erythematous rash on sun-exposed areas, the back, shoulders, and upper arms. There was associated muscle weakness and significant weight loss. Investigation revealed mildly raised aspartate and alanine transaminases but normal creatine kinase. Inflammatory indices and antinuclear antibodies (ANAs) were normal. Biopsy of the rash was reported as consistent with either dermatomyositis (DM) or acute lupus erythematosus. A diagnosis of DM was made, and prednisolone was given with improvement of the rash but deteriorating myopathy. The patient was referred to the rheumatology department, and further history revealed multiple vertebral fractures after falling from standing height; these had occurred six months prior to starting steroids. Besides smoking he had no other risk factors for osteoporosis. Examination showed normal muscle strength, no muscle tenderness, and no joint abnormality. Repeat muscle enzymes were normal, and ANAs were now 1 : 100, but dsDNA antibodies and extractable nuclear antigens were normal. Investigations for osteoporosis revealed a hypergonadotrophic hypogonadism picture. Further examination indicated scanty pubic and auxiliary hair, small testicles, and mild gynecomastia. He is married, though has no children of his own. The hormonal profile raised the possibility of Klinefelter's syndrome, which was subsequently confirmed with karyotyping of 47 XXY. Hypogonadism has been established as a cause of osteoporosis in males, and in this case would explain the occurrence of fractures in the absence of other major risk factors. Systemic lupus erythematosus has been recognized in association with Klinefelter's syndrome; in view of the normal muscle enzymes, his rash is most likely due to acute discoid lupus with androgen deficiency causing muscle weakness. PMID- 12114308 TI - Blunted ACTH and cortisol responses to systemic injection of corticotropin releasing hormone (CRH) in fibromyalgia: role of somatostatin and CRH-binding protein. AB - Thirteen female patients suffering from fibromyalgia (FM) and thirteen female age matched controls were intravenously injected with a bolus dose of 100 microg corticotropin-releasing hormone (CRH), and the evoked secretion pattern of ACTH, cortisol, somatostatin, and growth hormone (GH) was followed up for two hours, together with the plasma levels of CRH. The increases of ACTH and cortisol following CRH were not significantly different between controls and FM patients. The increase of plasma CRH following its injection was significantly higher in FM patients and lasted about 45 min, paralleled by an increase of somatostatin with a similar time course. Basal GH levels were significantly lower in FM patients. GH increased in FM patients 90 min after injection of CRH, coincident with decreasing CRH and somatostatin levels, while GH levels in controls rather decreased with the lowest values occurring 90 min after CRH. The results support the concept that the hormonal secretion pattern frequently observed in FM patients is primarily caused by CRH, possibly as a response to chronic pain and stress. The elevated levels of CRH in the circulation of FM patients suggest elevated levels of CRH-binding protein, which could explain why the levels of ACTH and cortisol between controls and FM following CRH do not differ. PMID- 12114309 TI - Involvement of the glucocorticoid receptor in the pathogenesis of rheumatoid arthritis. AB - The glucocorticoid receptor (GR) is a ligand-inducible transcription factor which controls the expression of several genes. Its cognate ligand, the glucocorticoids, induces receptor activation by binding to the cytoplasmic located receptor, ultimately leading to translocation of the receptor/hormone complex into the nucleus and the regulation of gene activity. Because glucocorticoids are widely used for suppression of inflammation in rheumatoid arthritis (RA), we investigated whether the expression level of GR is correlated with RA. We designed a study to detect the total amount of GR in lymphocytes of untreated RA patients, glucocorticoid-treated RA patients, and healthy controls. We observed a significant change in the expression levels of GR. Untreated RA patients exhibited a significantly higher amount of GR than the healthy controls, whereas glucocorticoid-treated RA patients showed a strongly decreased receptor density. These results seem to reflect a functional dysregulation of the HPA axis and may lead to a better understanding of the pathogenesis of RA. PMID- 12114310 TI - Regulation of apoptosis in endocrine autoimmunity: insights from Hashimoto's thyroiditis and Graves' disease. AB - Dysregulation of apoptosis is associated with the pathogenesis of organ-specific autoimmune diseases, through altered target organ susceptibility. Apoptosis signaling pathways can be initiated through activation of death receptors such as Fas. A comparative analysis of the expression of Fas and FasL, the antiapoptotic molecule Bcl-2, and apoptosis in both thyrocytes and thyroid-infiltrating lymphocytes (TILs) from patients with either Graves' disease (GD) or Hashimoto's thyroiditis (HT) was performed. GD thyrocytes expressed less Fas than HT thyrocytes, whereas GD TILs had higher levels of Fas and FasL than HT TILs. GD thyrocytes expressed higher levels of Bcl-2 compared with HT thyrocytes. The opposite pattern was observed in GD (low Bcl-2) and HT (high Bcl-2) TILs. Consistently, thyrocyte apoptosis was marked in HT and poor in GD thyroids, and TIL apoptosis was marked in GD and poor in HT. Our findings suggest that in GD thyroid the regulation of Fas/FasL/Bcl-2 favors apoptosis of infiltrating lymphocytes. Moreover, the reduced levels of Fas/FasL and increased levels of Bcl 2 should favor thyrocyte survival and hypertrophy associated with stimulatory thyroid-stimulating hormone receptor antibodies. In contrast, the regulation of Fas/FasL/Bcl-2 expression in HT can promote thyrocyte apoptosis via homophylic Fas-FasL interactions, and a gradual reduction in thyrocyte numbers leading to hypothyroidism. Fas-mediated apoptosis may be a general mechanism of cell damage in destructive organ-specific autoimmunity. PMID- 12114311 TI - Serum osteocalcin levels in premenopausal rheumatoid arthritis patients. AB - The objective of this study was to assess the occurrence of generalized bone loss in rheumatoid arthritis (RA) patients and to evaluate the factors influencing bone loss, in particular, the usefulness of bone turnover markers. Twenty-five premenopausal patients (mean age, 40 x 5 years) with active RA were compared with 27 age-matched premenopausal patients with RA but without active disease and 30 age-matched healthy premenopausal controls. Serum concentrations of osteocalcin, intact parathyroid hormone (PTH), spot urine concentrations of crosslinked N telopeptidases of type 1 collagen (NTX), and deoxypyridinoline (DPD) were detected by ELISA and radioimmunoassay. Serum osteocalcin levels were found to be significantly lower (p < 0.001) in patients with active RA compared with patients without active RA and controls. Similarly, serum intact PTH was significantly lower (p < 0.01) in patients with active RA than in patients with RA without active disease and controls. Spot urine concentrations of NTX and DPD were significantly higher (p < 0.01) in active RA patients than in patients with nonactive RA and controls. Positive correlations between osteocalcin and marker of disease activity were found to be significant (p < 0.01). There were no significant correlations between serum intact PTH, urine concentrations of NTX and DPD, and markers of inflammation. This study suggests that generalized bone loss occurs in active RA and is characterized by an evident bone resorption correlated with the high levels of inflammation. PMID- 12114312 TI - Prolactin/cortisol ratio in rheumatoid arthritis. AB - Prolactin (PRL) and glucocorticoids are hormones involved in the regulation of the immune system. Rheumatoid arthritis (RA) is an inflammatory condition that presents a diurnal rhythm of disease activity. PRL/cortisol ratio, and IL-1beta and TNF-alpha levels were determined in patients with RA and in control subjects at 0600, 1000, 1400, 1800, 2200, and 0200 hours. In patients with RA we observed higher PRL/cortisol ratio at 0200 hours, whereas IL-1beta and TNF-alpha reached their highest serum levels at 0200 and 0600 hours. In patients with RA we observed an imbalance in favor of proinflammatory hormones as opposed to levels of antiinflammatory hormones during nocturnal hours together with increased levels of IL-1beta and TNF-alpha of the diurnal rhythm of disease activity. PMID- 12114313 TI - TRPC4 knockout mice: the coming of age of TRP channels as gates of calcium entry responsible for cellular responses. PMID- 12114314 TI - Hypoxia and HIF-1alpha stability: another stress-sensing mechanism for Shc. PMID- 12114315 TI - nNOS-containing perivascular nerves: stranger by the minute. PMID- 12114316 TI - Induction of bone-type alkaline phosphatase in human vascular smooth muscle cells: roles of tumor necrosis factor-alpha and oncostatin M derived from macrophages. AB - Inflammatory cells such as macrophages and T lymphocytes play an important role in vascular calcification associated with atherosclerosis and cardiac valvular disease. In particular, macrophages activated with cytokines derived from T lymphocytes such as interferon-gamma (IFN-gamma) may contribute to the development of vascular calcification. Moreover, we have shown the stimulatory effect of 1alpha,25-dihydroxyvitamin D3 (1,25(OH)2D3) on in vitro calcification through increasing the expression of alkaline phosphatase (ALP), an ectoenzyme indispensable for bone mineralization, in vascular smooth muscle cells. Therefore, we hypothesized that macrophages may induce calcifying phenotype, especially the expression of ALP in human vascular smooth muscle cells (HVSMCs) in the presence of IFN-gamma and 1,25(OH)2D3. To test this hypothesis, we used cocultures of HVSMCs with human monocytic cell line (THP-1) or peripheral blood monocytes (PBMCs) in the presence of IFN-gamma and 1,25(OH)2D3. THP-1 cells or PBMCs induced ALP activity and its gene expression in HVSMCs and the cells with high expression of ALP calcified their extracellular matrix by the addition of beta-glycerophosphate. Thermostability and immunoassay showed that ALP induced in HVSMCs was bone-specific enzyme. We further identified tumor necrosis factor alpha (TNF-alpha) and oncostatin M (OSM) as major factors inducing ALP in HVSMCs in the culture supernatants of THP-1 cells. TNF-alpha and OSM, only when applied together, increased ALP activities and in vitro calcification in HVSMCs in the presence of IFN-gamma and 1,25(OH)2D3. These results suggest that macrophages may contribute to the development of vascular calcification through producing various inflammatory mediators, especially TNF-alpha and OSM. PMID- 12114317 TI - 1alpha,25-dihydroxyvitamin D3 induces vascular smooth muscle cell migration via activation of phosphatidylinositol 3-kinase. AB - The steroid hormone 1alpha,25-dihydroxyvitamin D3 [1alpha, 25-(OH)2D3] promotes vascular smooth muscle cell (VSMC) growth and calcification, but the precise mechanism by which 1alpha, 25-(OH)2D3 regulates VSMC migration is unknown. In rat aortic SMCs, we found that 1alpha, 25-(OH)2D3 (0.1 to 100 nmol/L) induced a dose dependent increase in VSMC migration. This response required the activation of phosphatidylinositol 3-kinase (PI3 kinase) because 1alpha, 25-(OH)2D3-induced migration was completely abolished by the PI3 kinase inhibitors, LY294002 (10 micromol/L) or wortmannin (30 nmol/L). Furthermore, the RNA polymerase inhibitor, 5,6-dichlorobenzimidazole riboside (50 micromol/L), did not affect 1alpha, 25 (OH)2D3-induced VSMC migration, suggesting that gene transcription is not involved in this rapid response. Using analogs of 1alpha, 25-(OH)2D3, which have been characterized for their abilities to induce either transcriptional or nontranscriptional responses of 1alpha, 25-(OH)2D3, we found that 1alpha,25 dihydroxylumisterol, which is a potent agonist of the rapid, nongenomic responses, was equipotent with 1alpha, 25-(OH)2D3 in inducing PI3 kinase activity and VSMC migration. Moreover, 1beta, 25-(OH)2D3, which specifically antagonizes the nongenomic actions of 1alpha, 25-(OH)2D3, abolished 1alpha, 25-(OH)2D3 induced PI3 kinase activity and VSMC migration, whereas the inhibitor of the genomic actions of vitamin D, (23S)-25-dehydro-1alpha-OH-D3-26,23-lactone, did not affect these responses. These results indicate that 1alpha, 25-(OH)2D3 induces VSMC migration independent of gene transcription via PI3 kinase pathway, and suggest a possible mechanism by which 1alpha, 25-(OH)2D3 may contribute to neointima formation in atherosclerosis and vascular remodeling. PMID- 12114318 TI - Novel vascular endothelial growth factor binding domains of fibronectin enhance vascular endothelial growth factor biological activity. AB - Interactions between integrins and growth factor receptors play a critical role in the development and healing of the vasculature. This study mapped two binding domains on fibronectin (FN) that modulate the activity of the angiogenic factor, vascular endothelial growth factor (VEGF). Using solid-phase assays and surface plasmon resonance analysis, we identified two novel VEGF binding domains within the N- and C-terminus of the FN molecule. Native FN bound to VEGF enhanced endothelial cell migration and mitogen-activated protein (MAP) kinase activity, but FN that is devoid of the VEGF binding domains failed to do so. Coprecipitation studies confirmed a direct physical association between VEGF receptor-2 (Flk-1) and the FN integrin, alpha5beta1, which required intact FN because FN fragments lacking the VEGF binding domains failed to support receptor association. Thrombin-activated platelets released intact VEGF/FN complexes, which stimulated endothelial cell migration and could be inhibited by soluble high affinity VEGF receptor 1 and antibodies to alpha5beta1 integrin. This study demonstrates that FN is potentially a physiological cofactor for VEGF and provides insights into mechanisms by which growth factor receptors and integrins cooperate to influence cellular behavior. PMID- 12114320 TI - Hypoxic induction of the hypoxia-inducible factor is mediated via the adaptor protein Shc in endothelial cells. AB - Tyrosine kinase cascades may play a role in the hypoxic regulation of hypoxia inducible factor (HIF)-1. We investigated the role of tyrosine kinase phosphorylation and of the Shc/Ras cascade on hypoxic HIF-1 stabilization. Exposure of human umbilical vein endothelial cells to hypoxia results in HIF protein stabilization as early as 10 minutes, with a maximum at 3 hours, and also in Shc tyrosine phosphorylation, with a maximum at 10 minutes. To test whether Shc directly mediates hypoxia-induced HIF stabilization, human umbilical vein endothelial cells were transfected with a dominant-negative Shc mutant (dnShc), resulting in significantly reduced HIF protein levels compared with control. Similar results were obtained with cells transfected with dominant-negative Ras, a known downstream effector of Shc. Hypoxia-induced Ras activity was significantly reduced in cells transfected with dnShc compared with control levels, indicating that Ras indeed acts downstream from Shc. Moreover, cells pretreated with a specific Raf-1 kinase inhibitor, a known downstream effector of Ras, exhibited reduced HIF protein levels. To examine the functional consequences of Shc in hypoxic signaling, HIF-1 ubiquitination, protein stabilization, and endothelial cell migration were assessed. Overexpression of dnShc increased ubiquitination of HIF-1 and reduced the half-life of the protein. Moreover, dnShc, dominant-negative Ras, or the Raf-1 kinase inhibitor significantly inhibited migration under hypoxia. Thus, Shc in concert with Ras and Raf-1 contributes to hypoxia-induced HIF-1alpha protein stabilization and endothelial cell migration. PMID- 12114319 TI - Role of sterol regulatory element binding proteins in the regulation of Galpha(i2) expression in cultured atrial cells. AB - We have previously demonstrated that growth of embryonic chick atrial cells in medium supplemented with lipoprotein-depleted serum (LPDS) resulted in a coordinate increase in the expression of genes involved in the parasympathetic response of the heart (the M2 muscarinic receptor; the alpha-subunit of the heterotrimeric G protein, Galpha(i2); and the inward rectifying K+ channel protein, GIRK1) and a marked increase in the negative chronotropic response of atrial cells to muscarinic stimulation. In the present study, we demonstrate that regulation of Galpha(i2) promoter activity by LPDS is mediated by the binding of a sterol regulatory element binding protein (SREBP) to a sterol regulatory element (SRE) in the Galpha(i2) promoter. Deletion and point mutation of this putative SRE interfered with the regulation of the Galpha(i2) promoter by SREBP and LPDS. Furthermore gel shift assays demonstrated that point mutations in the putative Galpha(i2) SRE markedly inhibited the binding of purified SREBP to oligonucleotides containing the Galpha(i2) SRE sequence. The expression of a dominant-negative SREBP mutant interfered with LPDS stimulation of Galpha(i2) promoter activity. Finally, we demonstrate that SREBP-1 is markedly more potent than SREBP-2 for the stimulation of Galpha(i2) promoter activity, suggesting that SREBP1 may play a role in the regulation of Galpha(i2) expression. These are the first data to demonstrate SREBP regulation of a protein not involved in lipid homeostasis and suggest a new relationship between lipid metabolism and the parasympathetic response of the heart. PMID- 12114321 TI - Phosphatidylinositol 3-kinase functionally compartmentalizes the concurrent G(s) signaling during beta2-adrenergic stimulation. AB - Compartmentation of intracellular signaling pathways serves as an important mechanism conferring the specificity of G protein-coupled receptor (GPCR) signaling. In the heart, stimulation of beta2-adrenoceptor (beta2-AR), a prototypical GPCR, activates a tightly localized protein kinase A (PKA) signaling, which regulates substrates at cell surface membranes, bypassing cytosolic target proteins (eg, phospholamban). Although a concurrent activation of beta2-AR-coupled G(i) proteins has been implicated in the functional compartmentation of PKA signaling, the exact mechanism underlying the restriction of the beta2-AR-PKA pathway remains unclear. In the present study, we demonstrate that phosphatidylinositol 3-kinase (PI3K) plays an essential role in confining the beta2-AR-PKA signaling. Inhibition of PI3K with LY294002 or wortmannin enables beta2-AR-PKA signaling to reach intracellular substrates, as manifested by a robust increase in phosphorylation of phospholamban, and markedly enhances the receptor-mediated positive contractile and relaxant responses in cardiac myocytes. These potentiating effects of PI3K inhibitors are not accompanied by an increase in beta2-AR-induced cAMP formation. Blocking G(i) or Gbetagamma signaling with pertussis toxin or betaARK-ct, a peptide inhibitor of Gbetagamma, completely prevents the potentiating effects induced by PI3K inhibition, indicating that the pathway responsible for the functional compartmentation of beta2-AR-PKA signaling sequentially involves G(i), Gbetagamma, and PI3K. Thus, PI3K constitutes a key downstream event of beta2-AR-G(i) signaling, which confines and negates the concurrent beta2-AR/G(s)-mediated PKA signaling. PMID- 12114322 TI - Increased NADPH oxidase activity, gp91phox expression, and endothelium-dependent vasorelaxation during neointima formation in rabbits. AB - Reactive oxygen species including superoxide and hydrogen peroxide are important mediators in atherogenesis. We investigated the enzymatic source of vascular superoxide and its role in endothelium-dependent vasorelaxation during neointima formation. Silastic collars positioned around carotid arteries of rabbits for 14 days induced neointimal thickening. Using lucigenin-enhanced chemiluminescence, superoxide production was detectable in collared artery sections, but not in controls, only after inactivation of endogenous Cu2+/Zn2+-superoxide dismutase (Cu2+/Zn2+-SOD) with diethyldithiocarbamate (DETCA). Dihydroethidium staining indicated that endothelium and adventitia were the major sites of superoxide generation. Superoxide production in DETCA-treated collared arteries was enhanced further by NADPH and was inhibited by diphenyleneiodonium, suggesting NADPH oxidase was the source of the radical in collared arteries. Moreover, real-time PCR demonstrated 11-fold higher expression of the gp91phox subunit of NADPH oxidase in collared arteries than in controls. In vascular reactivity studies, endothelium-dependent vasorelaxation to acetylcholine did not differ between collared and control sections. However, treatment with DETCA reduced relaxations to acetylcholine in collared rings, but not in controls. NADPH further reduced relaxations to acetylcholine in DETCA-treated collared sections, but not in controls. In DETCA/NADPH-treated collared rings, sensitivity to nitroprusside, in contrast to acetylcholine, exceeded that of controls. Moreover, further treatment of such rings with exogenous Cu2+/Zn2+-SOD restored acetylcholine relaxations without altering nitroprusside responses. Thus, early neointimal lesions induced by periarterial collars are associated with elevated gp91phox expression and increased NAPDH-oxidase-dependent superoxide production in endothelium and adventitia. However, endothelium-dependent vasorelaxation is largely preserved due to the actions of Cu2+/Zn2+-SOD and increased smooth muscle sensitivity to nitric oxide. PMID- 12114323 TI - Alpha7-nicotinic acetylcholine receptors on cerebral perivascular sympathetic nerves mediate choline-induced nitrergic neurogenic vasodilation. AB - It has been suggested in isolated porcine cerebral arteries that stimulation by nicotine of alpha7-nicotinic acetylcholine receptors (alpha7-nAChRs) on sympathetic nerves, but not direct stimulation of parasympathetic nitrergic nerves, caused nitrergic neurogenic dilation. Direct evidence supporting this hypothesis has not been presented. The present study, which used in vitro tissue bath and confocal microscopy techniques, was designed to determine whether choline, a selective agonist for alpha7-nAChRs, induced sympathetic-dependent nitrergic dilation of porcine basilar arterial rings. Choline and several nAChR agonists induced exclusive relaxation of basilar arterial rings without endothelium. The relaxation was blocked by tetrodotoxin, nitro-L-arginine, guanethidine, and beta2-adrenoceptor antagonists. Furthermore, the relaxation was blocked by methyllycaconitine and alpha-bungarotoxin (preferential alpha7-nAChR antagonists) and mecamylamine but was not affected by dihydro-beta-erythroidine (a preferential alpha4-nAChR antagonist). Confocal microscopic study demonstrated that choline and nicotine induced significant calcium influx in cultured porcine superior cervical ganglionic cells but failed to affect calcium influx in cultured sphenopalatine ganglionic cells, providing direct evidence that choline and nicotine did not act directly on the parasympathetic nitrergic neurons. The increased calcium influx in superior cervical ganglionic cells was attenuated by alpha-bungarotoxin and methyllycaconitine but not by dihydro-beta-erythroidine. These results support our hypothesis that activation of alpha7-nAChRs on cerebral perivascular sympathetic nerves causes calcium influx and the release of norepinephrine, which then act on presynaptic beta2-adrenoceptors located on the neighboring nitrergic nerve terminals, resulting in NO release and vasodilation. Endogenous choline may play an important role in regulating cerebral sympathetic activity and vascular tone. PMID- 12114324 TI - Impairment of store-operated Ca2+ entry in TRPC4(-/-) mice interferes with increase in lung microvascular permeability. AB - We investigated the possibility that the TRPC gene family of putative store operated Ca2+ entry channels contributes to the increase in microvascular endothelial permeability by prolonging the rise in intracellular Ca2+ signaling. Studies were made in wild-type (wt) and TRPC4 knockout (TRPC4(-/-) mice and lung vascular endothelial cells (LECs) isolated from these animals. RT-PCR showed expression of TRPC1, TRPC3, TRPC4, and TRPC6 mRNA in wt LECs, but TRPC4 mRNA expression was not detected in TRPC4(-/-) LECs. We studied the response to thrombin because it is known to increase endothelial permeability by the activation of G protein-coupled proteinase-activated receptor-1 (PAR-1). In wt LECs, thrombin or PAR-1 agonist peptide (TFLLRNPNDK-NH2) resulted in a prolonged Ca2+ transient secondary to influx of Ca2+. Ca2+ influx activated by thrombin was blocked by La3+ (1 micromol/L). In TRPC4(-/-) LECs, thrombin or TFLLRNPNDK-NH2 produced a similar initial increase of intracellular Ca2+ secondary to Ca2+ store depletion, but Ca2+ influx induced by these agonists was drastically reduced. The defect in Ca2+ influx in TRPC4(-/-) endothelial cells was associated with lack of thrombin-induced actin-stress fiber formation and a reduced endothelial cell retraction response. In isolated-perfused mouse lungs, the PAR-1 agonist peptide increased microvessel filtration coefficient (K(f,c)), a measure of vascular permeability, by a factor of 2.8 in wt and 1.4 in TRPC4(-/-); La3+ (1 micromol/L) addition to wt lung perfusate reduced the agonist effect to that observed in TRPC4(-/-). These results show that TRPC4-dependent Ca2+ entry in mouse LECs is a key determinant of increased microvascular permeability. PMID- 12114326 TI - Lung cancer screening: conundrum or contumacy? PMID- 12114325 TI - Osteopontin plays an important role in the development of medial thickening and neointimal formation. AB - Osteopontin (OPN) is a soluble secreted phosphoprotein that binds with high affinity to several integrins and it has been found at the site of atherosclerotic lesions. However, the role of OPN expression in vivo is still poorly understood. To investigate the physiological role of OPN in detail, we generated transgenic mice (Tg) overexpressing the OPN gene under control of the cytomegalovirus enhancer/chicken beta-actin promoter. We detected OPN mRNAs in almost all tissues of 3 lines of Tg mice by Northern blotting. The serum levels of OPN were significantly higher in Tg than in non-Tg mice (782+/-107 versus 182+/-44 ng/mL; P<0.001). Compared with non-Tg mice, a 73% (88+/-6 versus 51+/-7 microm; P<0.001) and 94% (126+/-15 versus 73+/-11 microm; P<0.0001) increase in the medial thickness of the aorta was determined in Tg mice at 16 and 32 weeks after birth. However, we found no evidence of inflammatory cells adhering to endothelial cells, intimal hyperplasia, or calcification in any region of Tg mice without artery injury. We then investigated the effect of cuff-induced injury to the femoral artery. The intimal thickening in Tg mice increased 2.9-fold more than that in non-Tg mice (4.9+/-1.9 versus 1.7+/-0.4 microm; P=0.022). The expression of OPN induces both medial thickening without injury and neointimal formation after injury, thus suggesting that OPN plays a role in the development of atherosclerosis, vascular remodeling, and restenosis after angioplasty in vivo. PMID- 12114327 TI - Lung cancer screening, once again. PMID- 12114328 TI - Quality of life after lung cancer surgery: a forgotten outcome measure. PMID- 12114329 TI - Lessening the punch of heparin-induced thrombocytopenia. PMID- 12114330 TI - Does splinting from thoracic bone ischemia and infarction contribute to the acute chest syndrome in sickle cell disease? PMID- 12114331 TI - The quick and the dead: the importance of rapid evaluation of infiltrates in the immunocompromised patient. PMID- 12114333 TI - Lung cancer screening using low-dose spiral CT: results of baseline and 1-year follow-up studies. AB - STUDY OBJECTIVE: To evaluate the feasibility of lung cancer screening using low dose spiral CT as a part of annual health examinations. DESIGN: Nonrandomized, screening practice. METHODS: From April 1998 to August 2000, CT screening was performed as a part of annual health examinations on a total of 7,956 individuals who belonged to the Hitachi Employee's Health Insurance Group. Of those participants, 5,568 were rescreened 1 year later. When a noncalcified solitary pulmonary nodule (SPN) >or= 8 mm was detected on CT screening, a detailed CT scan was carried out approximately 1 month later. RESULTS: During the baseline screening, a total of 2,865 noncalcified SPNs were detected among the 7,956 participants. Primary lung cancer was histologically confirmed in 40 patients (41 lesions). The prevalence was 0.44% of all participants from the baseline, and 0.07% from the repeated screening. Thirty-five of 41 tumors were stage I. Current or former smokers represented only 17 of 40 cases. The detection rate was rather high in female participants. CONCLUSION: Low-dose spiral CT seems to be a promising method for screening early lung cancer as a part of annual health examinations. Female and nonsmoking subjects should be included in the baseline screening. However, for yearly repeat screening, the participants may be selected on the basis of gender, smoking history, and results at the baseline screening. PMID- 12114334 TI - What happens to patients undergoing lung cancer surgery? Outcomes and quality of life before and after surgery. AB - OBJECTIVE: To compare baseline preoperative and 6-month postoperative functional health status and quality of life in patients undergoing lung cancer resection. METHODS: Lung cancer surgery patients from three hospitals were administered the Short-Form 36 Health Survey (SF-36) and the Ferrans and Powers' quality-of-life index (QLI) before surgery and 6 months after surgery. Preoperative, intraoperative, hospital stay, and 6-month postoperative clinical data were collected. All p values or= 50% were independent predictors for the secondary end point. CONCLUSION: The ACE DD genotype is associated with increased midterm mortality and cardiac morbidity after CABG. PMID- 12114336 TI - Heparin-induced thrombocytopenia: temporal pattern of thrombocytopenia in relation to initial use or reexposure to heparin. AB - STUDY OBJECTIVES: Heparin-induced thrombocytopenia (HIT) is an immune-mediated adverse drug reaction associated with a decrease of platelet counts that usually begins after at least 5 days of heparin treatment. Uncertainty exists about the risk of early onset of HIT (ie, < 5 days) in relation to previous heparin exposure. We therefore analyzed the temporal pattern of thrombocytopenia in patients with laboratory-confirmed HIT to assess whether patients with previous heparin exposure have an increased risk of early onset of HIT. DESIGN: Platelet count patterns in patients with a laboratory-confirmed diagnosis of HIT were examined in a retrospective chart review of a clinical study database. The onset of thrombocytopenia < 100 x 10(9)/L associated with the current heparin treatment (mainly unfractionated heparin) was analyzed using nonparametric maximum likelihood estimation. RESULTS: A total of 119 patients with 125 treatment episodes were assessed: HIT developed in 79 patients during initial exposure to heparin, and in 46 patients during reexposure. Early onset (< 5 days) of thrombocytopenia was associated with very recent heparin exposure. Patients reexposed to heparin within 3 months had an earlier onset of thrombocytopenia as compared to patients reexposed to heparin after 3 months (4.9 +/- 4.4 days vs 11.5 +/- 5.5 days [mean +/- SD], p = 0.001). There was no difference between onset on thrombocytopenia < 100 x 10(9)/L in patients reexposed to heparin within 3 to 12 months and after 1 year (9.7 +/- 6.4 days vs 12.3 +/- 5.2 days, p = 0.41). Whether platelet counts were obtained daily or less regularly did not affect the analysis. CONCLUSION: Early onset of thrombocytopenia in HIT is associated with recent heparin treatment (< 3 months). In contrast, for patients who did not receive heparin within the previous 3 months, HIT is an unlikely explanation for thrombocytopenia that occurs within the first 5 days. PMID- 12114337 TI - Breathing patterns during vaso-occlusive crisis of sickle cell disease. AB - STUDY OBJECTIVES: To determine the effect of sickle cell pain and its treatment on patients' breathing patterns, and to compare the effect of thoracic cage pain to pain at other sites. DESIGN: Prospective, observational study. SETTING: Sickle Cell Center Day Hospital. PATIENTS: Twenty-five patients with sickle cell disease admitted to the Sickle Cell Center Day Hospital for treatment of vaso-occlusive crisis (VOC) [10 patients with chest (thoracic cage) pain]. INTERVENTIONS: Breathing patterns were measured by respiratory inductive plethysmography. Tidal breathing data, including respiratory rate, tidal volume (VT), minute ventilation, and the rib cage contribution to VT, were collected at baseline and then following treatment with opioid analgesia. MEASUREMENTS AND RESULTS: The patients with chest pain had smaller V(Ts) at baseline than those with pain at other sites (355 +/- 37 mL vs 508 +/- 141 mL, p = 0.003), and higher respiratory rates (23.2 +/- 8.2 breaths/min vs 17.6 breaths/min, p = 0.03). These differences became insignificant following opioid treatment. Six patients had respiratory alternans (four patients in the chest pain group, and two patients with pain at other sites). All cases of respiratory alternans resolved following opioid administration. CONCLUSIONS: Patients with VOC and chest pain have more shallow, rapid breathing than patients with pain elsewhere. Analgesia reduces these differences. As pain-associated shallow breathing and maldistribution of ventilation may contribute to the pathogenesis of acute chest syndrome, these results support the need for adequate pain relief and monitoring of ventilatory patterns during the treatment of VOC. PMID- 12114338 TI - A 6-month, placebo-controlled study comparing lung function and health status changes in COPD patients treated with tiotropium or salmeterol. AB - BACKGROUND: Tiotropium, a once-daily anticholinergic, and salmeterol represent two inhaled, long-acting bronchodilators from different pharmacologic classes. A trial was designed to examine the efficacy and safety of both compounds with multiple outcome measures, including lung function, dyspnea, and health-related quality of life (HRQoL) in patients with COPD. METHODS: A 6-month, randomized, placebo-controlled, double-blind, double-dummy, parallel-group study of tiotropium, 18 microg once daily via dry-powder inhaler, compared with salmeterol, 50 microg bid via metered-dose inhaler, was conducted in patients with COPD. Efficacy was assessed by 12-h monitoring of spirometry, transition dyspnea index (TDI), and the St. George's Respiratory Questionnaire (SGRQ). RESULTS: A total of 623 patients participated (tiotropium, n= 209; salmeterol, n = 213; and placebo, n = 201). The groups were similar in age (mean, 65 years), gender (75% men), and baseline FEV(1) (mean, 1.08 +/- 0.37 L; percent predicted, 40 +/- 12% [+/- SD]). Compared with placebo treatment, the mean predose morning FEV(1) following 6 months of therapy increased significantly more for the tiotropium group (0.14 L) than the salmeterol group (0.09 L; p < 0.01). The average FEV(1) (0 to 12 h) for tiotropium was statistically superior to salmeterol (difference, 0.08 L; p < 0.001). Tiotropium improved TDI focal score by 1.02 U compared with placebo (p = 0.01), whereas there was no significant change in TDI focal score with salmeterol (0.24 U). Tiotropium was superior to salmeterol in improving TDI focal score (p < 0.05). At 6 months, the mean improvement in SGRQ total score vs baseline was tiotropium, - 5.14 U (p < 0.05 vs placebo); salmeterol, - 3.54 U (p = 0.4 vs placebo); and placebo, - 2.43 U. A statistically higher proportion of patients receiving tiotropium achieved at least a 4-U change in SGRQ score compared to patients receiving placebo. Both active drugs reduced the need for rescue albuterol (p < 0.0001). CONCLUSIONS: Tiotropium once daily produces superior bronchodilation, improvements in dyspnea, and proportion of patients achieving meaningful changes in HRQoL compared to twice-daily salmeterol in patients with COPD. PMID- 12114339 TI - The short-term effect of a rollator on functional exercise capacity among individuals with severe COPD. AB - STUDY OBJECTIVES: This study was conducted to examine the short-term effects of using a rollator on functional exercise capacity among individuals with COPD and to characterize which individuals benefit most from its use. DESIGN: Repeated measures randomized crossover design using the 6-min walk test (6MWT) as the primary outcome measure. SETTING: Respiratory rehabilitation center. PATIENTS: Forty stable subjects who had received a diagnosis of COPD. INTERVENTIONS: Two 6MWTs were performed on each study day. One 6MWT was performed unaided, and the other was performed with a rollator. The order was randomized on the first day and reversed on the second day. RESULTS: Use of the rollator was associated with a significant reduction in dyspnea (p < 0.001) and duration of rest (reduction for the total group, 19 s; and reduction for those who walked < 300 m unaided, 40 s; p = 0.001) during the 6MWT. For subjects who walked < 300 m unaided, there was also a significant improvement in distance walked (p = 0.02). No changes were found for the measures of cardiorespiratory function or gait (p > 0.05). The requirement to rest during an unaided 6MWT was a significant predictor of improved functional exercise capacity with the use of the rollator (p < 0.005). The majority of subjects whose unaided 6MWT distance was < 300 m preferred using the rollator to walking unaided. CONCLUSIONS: Use of a rollator was effective in improving functional exercise capacity by reducing dyspnea and rest duration among stable individuals with severe COPD. Individuals who walked < 300 m and individuals who required a rest during an unaided 6MWT benefited the most from using a rollator in terms of reduced dyspnea, reduced rest time, and improved distance walked. PMID- 12114340 TI - Biochemical efficacy and safety of a new pooled human plasma alpha(1) antitrypsin, Respitin. AB - BACKGROUND: Augmentation therapy with pooled human plasma-derived alpha(1) antitrypsin (AAT) has been shown to have biochemical efficacy in restoring serum AAT levels above the protective threshold. Also, clinical efficacy has been suggested. OBJECTIVE: To evaluate the bioequivalence of a new solvent detergent treated preparation of pooled human plasma-derived AAT (proposed name Respitin; Alpha Therapeutic Corporation; Los Angeles, CA) to the commercially available preparation (Prolastin; Bayer Corporation; West Haven, CT), we conducted a randomized controlled trial. METHODS: Eligible subjects were adults (> 18 years of age) who had never smoked or were ex-smokers, had severe deficiency of AAT, and had fixed airflow obstruction (ie, postbroncholdilator FEV(1) of 30 to 80% of predicted values and/or diffusing capacity of the lung for carbon monoxide [DLCO] of < 70% of predicted values with evidence of emphysema on a CT scan). Of the 28 subjects recruited, 26 completed the 12-week comparison. Participants were randomized to receive Respitin (60 mg/kg once weekly; 14 subjects) or Prolastin (60 mg/kg once weekly; 14 subjects), and recipients of Prolastin then crossed over to receive Respitin thereafter for the 24-week duration of the study. RESULTS: The primary efficacy criteria were satisfied for equivalence to comparator (ie, the ratio of mean trough serum levels for Respitin/Prolastin at weeks 8 to 11 exceeded the efficacy criterion [0.905; p = 0.0206] as did the slope of the mean trough level over weeks 11 to 23 [-0.003 micromol per week]). In Respitin recipients, the trough serum antineutrophil elastase capacity at week 7 and at weeks 8 to 11 was also equivalent to the comparator, as was the rise in AAT levels in epithelial lining fluid from baseline to week 7. The levels of urinary elastin degradation products showed little appreciable change for > 24 weeks, and no difference between compared groups was shown through week 12. Adverse events were similarly infrequent in compared groups. Finally, neither spirometry measurements nor DLCO showed a significant change through 24 weeks. CONCLUSIONS: We conclude that this new solvent detergent-treated pooled human plasma-derived AAT (Respitin) demonstrates biochemical equivalence to Prolastin and that this new drug is well-tolerated. PMID- 12114341 TI - Effects of nasal pressure support on ventilation and inspiratory work in normocapnic and hypercapnic patients with stable COPD. AB - OBJECTIVES: To assess and compare the effect of nasal continuous positive airway pressure (nCPAP), inspiratory pressure support (PSV), and bilevel positive airway pressure (biPAP) on ventilatory parameters and inspiratory work (WOB) in normocapnic and hypercapnic patients with stable COPD. METHODS: While administering nasal pressure support to 10 normocapnic and 10 hypercapnic patients with COPD, we measured airflow and volume with a pneumotachograph as well as esophageal and gastric pressures under nCPAP, PSV, and biPAP conditions. RESULTS: nCPAP had no influence on ventilatory parameters but decreased WOB and transdiaphragmatic work (Wdi) at 10 cm H(2)O of pressure in both groups. With PSV and biPAP, ventilatory parameters increased proportionally to the inspiratory applied pressure. WOB and Wdi decreased significantly in both groups while increasing the pressure support. A similar decrease was observed during biPAP proportionally to the level of pressure support. The diaphragmatic pressure-time product decreased similarly in both groups during PSV and biPAP. CONCLUSION: The ventilatory response under nCPAP, PSV, and biPAP conditions is similar in hypercapnic and normocapnic patients with stable COPD; PSV and biPAP increase ventilatory parameters and improve Wdi. On the contrary, nCPAP improves WOB but does not increase ventilatory parameters. PMID- 12114342 TI - Effect of a nonrebreathing exhalation valve on long-term nasal ventilation using a bilevel device. AB - STUDY OBJECTIVE: To determine whether an exhalation valve designed to minimize rebreathing improves daytime or nocturnal gas exchange or improves symptoms compared with a traditional valve during nocturnal nasal ventilation delivered using a bilevel pressure ventilation device. DESIGN: Prospective direct comparison trial with each patient sequentially using both valves, during a 2 week run-in period with a traditional valve, a 2-week trial with the nonrebreathing valve, and a 2-week washout period with the traditional valve. SETTING: Outpatient pulmonary function laboratory and home nocturnal monitoring. PATIENTS: Seven patients who received long-term (> 1 year) nocturnal nasal bilevel pressure ventilation with an expiratory pressure of 1 year. PMID- 12114344 TI - Effects of intermittent negative pressure ventilation on effective ventilation in normal awake subjects. AB - RATIONALE: Previous studies have shown that an increase in inspiratory pressure during nasal intermittent positive pressure ventilation (IPPV) does not result in increased effective minute ventilation (E) due to glottic interference. STUDY OBJECTIVES: To test the consequences of increases in negative pressure ventilation (NPV) on V(E). MATERIAL AND METHODS: Eight healthy awake subjects underwent NPV delivered by an iron lung. First, NPV was started at a respirator frequency (f) of 15 cycles per minute with an inspiratory negative pressure (INP) of - 15 cm H(2)O (F15-P15). Then, f was increased to 20 cycles per minute and INP was kept at - 15 cm H(2)O. Next, f was kept at 20 cycles per minute and INP was reduced to - 30 cm H(2)O (F20-P30). Finally, f was decreased to 15 cycles per minute and INP was kept at - 30 cm H(2)O. At each step and for each breath, effective tidal volume (VT), V(E), and end-tidal carbon dioxide pressure were measured. In three subjects, the glottis width was assessed using fiberoptic bronchoscopy. RESULTS: From spontaneous breathing to the first step of NPV (F15 P15), we observed an inhibition of the phasic inspiratory diaphragmatic electromyogram concomitant to a significant increase in V(E) (p < 0.0005). For the group as a whole, the increase in mechanical ventilation (from F15-P15 to F20 P30) resulted in significant increases in VT and V(E) leading to hypocapnia (p < 0.0005). Moreover, the glottis width did not decrease with the increase in mechanical ventilation. CONCLUSIONS: We conclude that in normal awake subjects, NPV allowed a significant increase in V(E). These results differ from those previously obtained with nasal IPPV in which the glottic width interferes with the delivered mechanical ventilation. PMID- 12114345 TI - Management of acute proximal deep vein thrombosis: pharmacoeconomic evaluation of outpatient treatment with enoxaparin vs inpatient treatment with unfractionated heparin. AB - OBJECTIVES: A landmark Canadian randomized controlled clinical trial compared treatment of acute proximal vein thrombosis via low-molecular-weight heparin (LMWH) [enoxaparin] administered primarily at home with IV unfractionated heparin (UH) in the hospital. Results demonstrated equivalent safety and efficacy for home care with enoxaparin with a reduction in cost. Our objective was to validate these findings in the routine practice setting of a US health maintenance organization. DESIGN: Retrospective analysis of medical and administrative records of health-plan members meeting inclusion-exclusion criteria of the Canadian trial during the period from 1995 to 1998. SETTING: Staff-model health maintenance organization serving New Mexico. PATIENTS: Persons presenting as outpatients from 1995 to 1996 or from 1997 to 1998 with acute, proximal deep vein thrombosis (DVT) diagnosed by duplex ultrasonography. INTERVENTIONS: Initial anticoagulant therapy of IV UH administered in the hospital (from 1995 to 1996 group, n = 64) or subcutaneous LMWH (enoxaparin) administered primarily at home (from 1997 to 1998 group, n = 65), followed by warfarin therapy. RESULTS: No statistically significant differences were observed in the number of recurrent venous thromboembolic events (p = 0.36) or bleeding events (p = 1.0). Mean +/- SD cost per patient was 9,347 dollars +/- 8,469 in the enoxaparin group compared with 11,930 dollars +/- 10,892 in the UH group, a difference of - 2,583 dollars (95% bootstrap-adjusted asymmetrical confidence interval, - 6,147 dollars, + 650 dollars). CONCLUSIONS: Retrospective replication of the Canadian study in a US routine (managed) care setting found similar clinical and economic outcomes. Treatment of acute proximal DVT with enoxaparin in a primarily outpatient setting can be accomplished safely and yields savings through avoidance or minimization of inpatient stays. PMID- 12114346 TI - Detection of pelvic vein thrombosis by magnetic resonance angiography in patients with acute pulmonary embolism and normal lower limb compression ultrasonography. AB - STUDY OBJECTIVE: In patients with proven acute pulmonary embolism (PE), a systematic search for "residual" deep vein thrombosis (DVT) using venography or compression duplex ultrasonography (CDUS) of the lower limbs is negative in 20 to 50% of patients. We hypothesized that undetectable pelvic vein thrombosis (from the external iliac vein to the inferior vena cava) could account for a substantial proportion of patients with negative CDUS findings. Using a noninvasive test, magnetic resonance angiography (MRA), the objective of the study was to assess the prevalence of pelvic DVT in patients with acute PEs and normal findings on lower limb CDUS. DESIGN: Prospective study. SETTING: A 35-bed respiratory unit in a 680-bed Parisian teaching hospital. PATIENTS: From June 1995 to October 1996, 24 patients (mean age, 49 years; age range, 18 to 83 years) with acute PEs and normal findings on lower limb CDUS underwent pelvic MRA. MEASUREMENTS AND RESULTS: MRA disclosed pelvic DVT in seven patients (29%). The common iliac vein was involved in five patients. Internal iliac vein (hypogastric) thrombosis was imaged in two patients, but no patients had DVT limited to this vein. Three patients underwent subsequent venography studies that confirmed the MRA findings. In three other patients, a new MRA at the end of anticoagulant therapy showed the resolution of the DVT. CONCLUSIONS: Our data support the view that, among patients with negative findings on CDUS, a substantial proportion of the DVTs that are responsible for PE originates in the pelvic veins. This study provides additional arguments to suggest that MRA might become the reference test for the exploration of pelvic DVT. PMID- 12114347 TI - Physicians' judgments of survival after medical management and mortality risk reduction due to revascularization procedures for patients with coronary artery disease. AB - STUDY OBJECTIVE: s: To assess the accuracy of physicians' judgments of survival probability for medically managed patients with coronary artery disease (CAD), and of the absolute risk reduction of mortality due to coronary artery bypass grafting (CABG) or percutaneous transluminal coronary angioplasty (PTCA) for such patients; and relationships among these judgments and the physicians' propensity to perform revascularization. DESIGN: Two surveys (for three-vessel or two-vessel CAD) for patients presenting with stable CAD, currently managed medically, and without other life-limiting problems. SETTING: Multiple educational conferences, 1996-1997. PARTICIPANTS: Conference attendees. MEASUREMENTS AND RESULTS: Main outcomes were proportions of patients for whom the physicians would recommend revascularization (CABG for three-vessel CAD, CABG or PTCA for two-vessel CAD), and judgments of the proportions of medically managed patients who would be alive after 5 years, 7 years, and 11 years, and of absolute risk reduction of mortality due to CABG (or PTCA for two-vessel CAD). At least one half of the participants judged the survival rate of medically managed patients with three-vessel or two vessel CAD to be less than the lowest rates supported by the best available evidence. More than one fourth judged the absolute risk reduction due to CABG to be higher than the highest values based on such evidence. Physicians' propensity to perform revascularization correlated inversely with their judgments of survival given medical management, and with their judgments of absolute risk reduction due to revascularization. CONCLUSIONS: Physicians may overuse revascularization because of excessive pessimism about survival of medically managed patients, and excessive optimism about the survival benefits of revascularization. PMID- 12114348 TI - ECG discrimination between right and left circumflex coronary arterial occlusion in patients with acute inferior myocardial infarction: value of old criteria and use of lead aVR. AB - STUDY OBJECTIVES: Prior studies have proposed several ECG criteria for identifying the culprit artery in patients with acute inferior myocardial infarction (MI). We applied each criterion to our patients to assess its utility. In doing so, we discovered a previously unreported, but highly useful, criterion utilizing lead aVR. STUDY DESIGN: Retrospective review. PATIENTS: Thirty consecutive patients with symptoms of acute MI, ST-segment elevation in the inferior ECG leads, an appropriate rise and fall of creatine kinase and troponin I levels, and coronary arteriography within 7 days of the onset of symptoms. MEASUREMENTS: The ECG recorded within 24 h of the onset of symptoms that had the most prominent ST-segment changes was analyzed. In the 12 standard leads and in lead V(4)R, ST-segment elevation or depression was measured 0.06 s after the J point. RESULTS: Four previously described criteria were useful in identifying the right coronary artery (RCA) or the left circumflex coronary artery (LCX) as the culprit: ST-segment elevation in lead I, ST-segment more or less elevated in lead II than in lead III, ST-segment elevation >or= 0.5 mm in lead V(4)R, and various combinations of ST-segment elevation or depression in leads V(1) and V(2). A new criterion was found to be at least as useful as any previously described: the presence and amount of ST-segment depression in lead aVR. CONCLUSIONS: At least five different ST-segment criteria help to identify the RCA or the LCX as the culprit artery in patients with acute inferior MI. One of these, the amount of ST segment depression in lead aVR, has not been reported previously and needs validation in a larger study. PMID- 12114349 TI - Noninvasive evaluation of pulmonary capillary wedge pressure by BP response to the Valsalva maneuver. AB - STUDY OBJECTIVES: To determine the BP response to the Valsalva maneuver (VM) at baseline and after changes in therapy and to compare this response to the invasively measured pulmonary capillary wedge pressure (PCWP). DESIGN: Comparison of the BP response to the VM with invasively measured PCWP. In a subset of patients, direct PCWP and pulse amplitude ratio (PAR) measurements were repeated (mean +/- SD) 3.2 +/- 4.5 months later after adjusting the therapy. SETTING: Tertiary-care center. PATIENTS: Forty-two stable patients (8 women; mean age, 58 +/- 13 years) undergoing right heart catheterization who were in sinus rhythm. MEASUREMENTS: PAR calculated between the end and the beginning of the VM using the last two beats and the first three beats of the straining phase and simultaneous measurement of PCWP. RESULTS: There was a highly significant correlation between the invasively measured PCWP (range, 2 to 32 mm Hg) and the PAR (range, 0.28 to 1.15; R(2) = 0.75; p < 0.001). In addition, changes of PCWP during follow-up (-16 to 13 mm Hg) were well-correlated (R(2) = 0.93; p < 0.001; n = 11) with changes in PAR (-0.44 to 0.47). The administration of medication (eg, beta-blockers, amiodarone, angiotensin-converting enzyme inhibitor, and digoxin) did not influence the results. CONCLUSIONS: PCWP and changes during therapy can be estimated noninvasively by measuring the PAR during the VM with acceptable accuracy in stable patients with cardiac conditions. Thus, this method may be a useful tool in detecting an elevated PCWP and hemodynamic response to therapy. PMID- 12114350 TI - Bronchoprotective effects of leukotriene receptor antagonists in asthma: a meta analysis. AB - STUDY OBJECTIVE: Cysteinyl leukotrienes are important proinflammatory mediators in the pathogenesis of asthma. Since bronchial hyperresponsiveness is a noninvasive surrogate marker of asthmatic airway inflammation, we evaluated the bronchoprotection afforded by leukotriene receptor antagonists (LTRAs). DESIGN: Systematic review of randomized, placebo-controlled trials in which LTRAs were administered for >or= 5 days. Studies in which active drug was administered as a first-line or second-line therapy were used. SETTING: MEDLINE, BIDS, and Cochrane Library data registers. MEASUREMENTS: The doubling dose/dilution difference that caused a 20% fall in the FEV(1) between LTRA and placebo. RESULTS: Thirteen trials (353 subjects) fulfilled eligibility criteria. Combining the results the overall weighted estimated protection amounted to a 0.85 doubling dose shift (95% confidence interval, 0.69 to 1.02). CONCLUSION: Since the estimated protection amounted to almost one doubling dose, this reinforces the role of LTRAs as anti inflammatory therapy in asthma. PMID- 12114351 TI - Effects of adding either a leukotriene receptor antagonist or low-dose theophylline to a low or medium dose of inhaled corticosteroid in patients with persistent asthma. AB - STUDY OBJECTIVES: To evaluate the effect of adding zafirlukast or low-dose theophylline to a beclomethasone dipropionate (BDP) extra-fine hydrofluoroalkane aerosol on bronchial hyperresponsiveness as the primary outcome variable. METHODS: Twenty-four patients with mild-to-moderate asthma were studied using a randomized crossover design with the following three treatment blocks: (1) beclomethasone, 100 microg/d, alone for the first 2 weeks followed by 400 microg/d alone for the next 2 weeks; (2) beclomethasone, 100 microg/d, followed by 400 microg/d, with the addition of zafirlukast, 20 mg bid; (3) beclomethasone, 100 microg/d, followed by 400 microg/d, with the addition of theophylline, 200 to 300 mg bid. Measurements were made after 2 and 4 weeks of each treatment and at pretreatment baseline. RESULTS: The mean trough plasma theophylline concentration was 6.7 mg/L, coinciding with the anti-inflammatory target range (ie, 5 to 10 mg/L). The provocative dose of methacholine causing a 20% fall in FEV(1) (as doubling dose difference from baseline) was significantly (p < 0.05) greater with beclomethasone, 100 microg, plus zafirlukast (1.1 doubling dose) but not with beclomethasone, 100 microg, plus theophylline (0.7 doubling dose) compared to beclomethasone, 100 microg alone (0.4 doubling dose), but not compared to beclomethasone, 400 microg alone (1.1 doubling dose). There were also significant (p < 0.05) differences between beclomethasone, 100 microg, plus zafirlukast (but not BDP, 100 microg, plus theophylline) vs beclomethasone, 100 microg, alone in terms of nitric oxide level, midexpiratory phase of forced expiratory flow, and peak expiratory flow. There were no further significant improvements observed with the addition of zafirlukast or theophylline to beclomethasone, 400 microg. CONCLUSIONS: A leukotriene receptor antagonist, but not low-dose theophylline, conferred significant additive anti-inflammatory effects to therapy with a low dose inhaled corticosteroid but not to that with a medium dose of an inhaled corticosteroid. Thus, optimizing the dose of inhaled corticosteroid as monotherapy would seem to be the logical first step, which is in keeping with current guidelines. PMID- 12114352 TI - Continuous vs intermittent beta-agonists in the treatment of acute adult asthma: a systematic review with meta-analysis. AB - BACKGROUND: Since the late 1980s, there has been considerable clinical and academic interest in the use of continuous aerosolized bronchodilators for the treatment of patients with acute asthma. These studies have suggested that this therapy is safe, is at least as effective as intermittent nebulization, and may be superior to intermittent nebulization in patients with the most severely impaired pulmonary function. OBJECTIVES: To determine whether continuous nebulization offered an advantage over intermittent nebulization for the treatment of adults with acute asthma in the emergency department (ED). DESIGN: Systematic review of randomized controlled trials of adults with acute asthma. OUTCOMES: Change in pulmonary function tests as primary outcome, and admissions to the hospital and side effects as secondary outcomes. RESULTS: Six studies including 393 adults with acute asthma were selected. No significant differences were demonstrated between the two delivery methods in terms of pulmonary function measures obtained after 1 h of treatment (standardized mean difference [SMD], 0.15; 95% confidence interval [CI], -0.35 to 0.05) and after 2 to 3 h of treatment (SMD, -0.19; 95% CI, -0.39 to 0.01). No significant heterogeneity was demonstrated (p > 0.5). At the end of treatment, there was a significantly greater decrease in pulse rate when the continuous nebulizer was used (weighted mean difference [WMD], -6.82; 95% CI, -8.67 to -3.90 beats/min; chi(2), 2.55; degrees of freedom [df], 4; p = 0.6). Additionally, the analysis showed a significant decrease of serum potassium concentration with the use of intermittent nebulization (WMD, 0.12; 95% CI, 0.24 to 0.01 mmol/L; chi(2), 0.5; df, 2; p = 0.8). However, this finding was obtained on the analysis of only two trials. Finally, at the end of the study period, no significant differences were identified between patients treated with continuous or intermittent nebulization with respect to hospital admission (relative risk, 0.68; 95% CI, 0.33 to 1.38; chi(2), 2.06; df, 1; p = 0.2). CONCLUSIONS: Overall, this review supports the equivalence of continuous and intermittent albuterol nebulization in the treatment of acute adult asthma. PMID- 12114353 TI - Vascular involvement in exercise-induced airway narrowing in patients with bronchial asthma. AB - STUDY OBJECTIVES: The bronchial microcirculation has the potential to contribute to the pathophysiologic mechanisms of exercise-induced bronchoconstriction (EIB) in asthmatic patients. This study was designed to determine whether increase in airway vascular permeability is associated with the severity of EIB in asthmatic patients. DESIGN: Cross-sectional analysis. SETTING: University hospital. PARTICIPANTS: Twenty-five asthmatic patients and 12 normal control subjects. INTERVENTIONS: All asthmatics performed an exercise test, and the percentage of maximal fall in FEV(1) and the area under the curve of the percentage fall in FEV(1) plotted against time for 30 min (AUC(0-30)) were determined. MEASUREMENTS AND RESULTS: The inflammatory indexes, NO levels, and airway vascular permeability index (ratio of albumin concentrations in induced sputum and serum) were examined in all subjects. The airway vascular permeability index was significantly higher in EIB-positive asthmatics (0.031 +/- 0.009) than in EIB negative asthmatics (0.020 +/- 0.005, p = 0.0011) and normal control subjects (0.008 +/- 0.003, p < 0.0001). We also found that there was a significant correlation between NO levels in induced sputum and the airway vascular permeability index (r = 0.525, p = 0.0101). Moreover, the airway vascular permeability index was significantly correlated with the severity of EIB (percentage of maximal fall in FEV(1) [r = 0.761, p = 0.0002], AUC(0-30) [r = 0.716, p = 0.0005]). However, this index was not significantly correlated with magnitude of eosinophilic inflammation. CONCLUSION: Our findings suggest that increased airway vascular permeability due to excessive production of NO correlates with the severity of EIB in asthmatics, and that assessment of albumin flux in airway lining fluid stimulated by hypertonic saline solution is a sensitive predictor of the severity of EIB. PMID- 12114354 TI - Association of asthma severity and bronchial hyperresponsiveness with a polymorphism in the cytotoxic T-lymphocyte antigen-4 gene. AB - OBJECTIVES: Cytotoxic T-lymphocyte antigen (CTLA)-4 is a homolog of CD28, which is expressed only on activated T cells. It binds to accessory molecule B7 and mediates T-cell-dependent immune response. Signaling through CTLA-4 may down regulate type 1 T-helper cell proliferation; moreover, some studies suggest that CTLA-4 might also deliver a positive signal to type 2 T-helper cell activation. Disruption of this delicate balance of immune regulation may lead to autoimmune diseases or atopic diseases. To evaluate the possible role of CTLA-4 polymorphisms in bronchial asthma, we investigated the association between polymorphisms (exon 1 +49 A/G, promoter -318 C/T) and atopy, asthma severity, and bronchial hyperresponsiveness in bronchial asthma patients and a group of healthy control subjects. PATIENTS: Eighty-eight asthmatic patients and 88 healthy control subjects were studied. MEASUREMENTS AND RESULTS: Asthma severity assessment, methacholine challenge, allergy skinprick test, and serum total IgE measurements were performed. The genotypes of the CTLA-4 promoter (-318 C/T) and exon 1 (+49 A/G) in all subjects were determined using the polymerase chain reaction and restriction fragment length polymorphism. The CTLA-4 promoter (-318 C/T) polymorphism was shown to be associated with asthma severity, but not with asthma, atopy, or bronchial hyperresponsiveness. A significant association was found between severe asthma and the T allele (p = 0.037). The CTLA-4 exon 1 (+49 A/G) polymorphism was shown to be associated with bronchial hyperresponsiveness, but not with asthma, atopy, or asthma severity. Asthmatic patients of the GG genotype had more hyperresponsive airways than those with the AG or AA genotype (p = 0.019). CONCLUSIONS: The CTLA-4 promoter (-318 C/T) T allele may serve as a clinically useful marker of severe asthma. The CTLA-4 exon 1 (+49 A/G) polymorphism may have a disease-modifying effect in asthmatic airways. PMID- 12114355 TI - Intrabreath diffusing capacity of the lung in healthy individuals at rest and during exercise. AB - BACKGROUND: Traditional approaches to measuring the diffusing capacity of the lung for carbon monoxide (DLCO) treat the lung as a single, well-mixed compartment and produce a single value for DLCO to represent an average diffusing capacity of the lung (DL). Because DL distribution in the lung is inhomogeneous, and changes in the DL in diseased lungs may be regional, measuring regional DL, especially during exercise, may be more sensitive in detecting pulmonary vascular diseases. OBJECTIVES: To characterize regional changes in DL in healthy individuals from rest to exercise, and to provide normal references for future studies in pulmonary vascular disorders. METHODS: We reanalyzed DLCO and phase III CH(4) slopes that were obtained during a slow, single exhalation at rest and during exercise in our extended database of 105 healthy individuals. DLCO profiles between 20% and 80% of exhaled vital capacity (VC) (ie, the intrabreath DLCO) were analyzed by calculating the average DLCO measured at midlung volume (ie, 30 to 45% of exhaled VC [DLCOMLV]) and by fitting the whole curve with a third-order polynomial equation. RESULTS: DLCO decreased nonlinearly by approximately 30%, from 20 to 80% of exhaled VC at rest. DLCO during exercise was greater than that at rest, and the increase was similar at all lung volumes. The CH(4) slopes at rest and during exercise were similar. Prediction equations based on regressions on age, sex, and height were computed for resting and exercise DLCOMLV and the phase III CH(4) slope (an index of ventilation distribution). CONCLUSIONS: Capillary recruitment/dilation during exercise in healthy individuals is a uniform process throughout the lungs. Our analyses provide a database for a noninvasive method that can incorporate exercise to evaluate the volume-dependent distribution of DLCO in lung diseases. PMID- 12114356 TI - Detection of wheezing during maximal forced exhalation in patients with obstructed airways. AB - STUDY OBJECTIVES: Wheezing is a common clinical finding in patients with asthma and COPD during episodes of severe airway obstruction, and can also be heard in normal subjects during forced expiratory maneuvers; however, the properties of wheezing are difficult to perceive and quantify during auscultation. We therefore developed and evaluated a new technique for recording and analyzing wheezing during forced expiratory maneuvers in a group of patients with obstructed airways (asthma, COPD) and a control group of healthy subjects. MATERIAL AND METHODS: Sixteen patients with asthma (9 men and 7 women), 6 patients with COPD (6 men), and 15 healthy subjects (7 men and 8 women) were enrolled. The patients had moderate-to-severe obstruction (FEV(1) of 40 to 53% predicted). A contact sensor on the trachea was used to record sound during forced expiratory maneuvers. Wheeze detection was carried out by a modified algorithm in a frequency-time space after applying the fast Fourier transform. RESULTS: More wheezes were recorded in patients with obstructed airways than in control subjects: asthma patients, 8.4 +/- 6.4 wheezes; COPD patients, 10.4 +/- 6.1 wheezes; and control subjects, 2.9 +/- 2.0 wheezes (mean +/- SD). The mean frequency of all detected wheezes was higher in control subjects than in patients with obstructed airways (asthma patients, 560.9 +/- 140.8 Hz; COPD patients, 669.4 +/- 250.1 Hz; and control subjects, 750.7 +/- 175.7 Hz). The total number of wheezes after terbutaline inhalation changed more in patients with obstructed airways than in control subjects. CONCLUSIONS: The new method that we describe for studying airway behavior during forced expiratory maneuvers is able to identify and analyze wheeze segments generated in patients with obstructed airways, as evidenced by the greater number of wheezes detected in the patient group, the main finding of this study. This method clearly and objectively identifies the presence of obstructive disease. PMID- 12114357 TI - Use of inspiratory muscle strength training to facilitate ventilator weaning: a series of 10 consecutive patients. AB - BACKGROUND AND PURPOSE: We instituted a low-repetition, high-intensity inspiratory muscle strength training (IMST) program and progressively longer spontaneous breathing periods (SBPs) in a group of medically complex patients who were dependent on mechanical ventilation (MV) and had failed to wean. CASE DESCRIPTIONS: IMST was provided to 10 consecutive patients (four men, six women; mean [+/- SD] age, 59 +/- 15 years) who had failed to wean from MV by conventional methods for >or= 7 days. Prior to initiating IMST, patients had received MV support for a mean of 34 +/- 31 days. Daily IMST consisted of four sets of six breaths through a threshold inspiratory muscle trainer that had been set at an intensity to yield an exertion rating of 6 to 8 of a maximal value of 10. At the start of IMST, patients were tolerating 2.1 +/- 3.4 consecutive hours of SBPs. The duration of the SBPs was increased daily, as tolerated. Patients were considered to have been weaned from MV when they were able to breathe without MV support for 24 consecutive hours. OUTCOMES: After 44 +/- 43 days of IMST, 9 of 10 patients were weaned from MV. The initial IMST pressure was 7 +/- 3 cm H(2)O, and it was increased to 18 +/- 7 cm H(2)O (p < 0.05). DISCUSSION: These results indicate that an IMST protocol that produces significant increases in threshold training pressure, in combination with progressive SBPs, aids in weaning patients from MV. Although promising, these preliminary observations must be tested in a controlled trial. PMID- 12114358 TI - Influence of gender and inspiratory muscle training on the perception of dyspnea in patients with asthma. AB - BACKGROUND: Men and women respond differently to asthma. PATIENTS AND METHODS: Maximal inspiratory mouth pressure (P(Imax)), beta(2)-agonist consumption, and perception of dyspnea (POD) were measured in 22 women and 22 men with mild persistent-to-moderate asthma. Next, the women were randomized into two groups: those who received inspiratory muscle training and those who received sham training. The training ended when the P(Imax) of the training group was equal to that of the male subjects. POD was then measured once again. RESULTS: Baseline P(Imax) was significantly lower (p < 0.01) while POD and mean daily beta(2) agonist consumption were significantly higher in the female subjects. P(Imax) reached the level of the male subjects at the end of the 20th week of training. The increase in the P(Imax) was associated with a statistically significant decrease in mean daily beta(2)-agonist use and in POD to a similar level as in male subjects. CONCLUSIONS: POD and mean daily beta(2)-agonist consumption in asthmatic women are significantly higher, and the P(Imax) significantly lower, than that of their male counterparts. When the P(Imax) of female subjects following training is equal to that in male subjects, the differences in POD and mean daily beta(2)-agonist consumption disappear. PMID- 12114359 TI - Endobronchial hamartoma. AB - OBJECTIVES: To describe clinical, endoscopic, radiographic, and follow-up characteristics of a series of patients in whom endobronchial hamartoma (EH) had been diagnosed. METHODS: Retrospective study of all cases of hamartoma diagnosed by bronchial biopsy between 1974 and 1997 in a tertiary referral hospital in Madrid, Spain. RESULTS: EH was diagnosed 47 patients during the study period. Four patients were excluded from the study because no clinical history was available. We analyzed the cases of 43 patients (37 men and 6 women), with a mean (+/- SD) age of 62 +/- 12 years. Seven patients had a concurrent lung neoplasm, and the EH was an incidental endoscopic finding. Among the other 36 patients, 31 had a new onset of respiratory symptoms, most commonly, recurrent respiratory infections in 16 patients (44%) and hemoptysis in a further 12 patients (33.4%). Chest radiograph findings were abnormal in 38 of 43 patients. At bronchoscopy, the lesions were equally distributed throughout the right and left lungs with no clear lobar predilection. Endobronchial obstruction was evident in 26 patients (72.2%) without concurrent neoplasm, 17 of whom underwent resection with a rigid bronchoscope and laser, with total resolution in 13 patients. Partial resolution was achieved in four patients, two of whom needed a second endoscopic procedure. Five patients were treated with open lung surgery. Clinical and endoscopic follow up was performed in 23 patients at 1 to 73 months (mean, 17 months), and recurrence was found in 4 patients. CONCLUSION: EH frequently produces respiratory complaints and radiographic abnormalities. Patients with endobronchial obstructions had satisfactory responses to endoscopic therapy. PMID- 12114360 TI - Laryngotracheoscopic findings in long-term follow-up after Griggs tracheostomy. AB - OBJECTIVE: Analysis of laryngotracheoscopic findings of the upper airway tract following percutaneous tracheostomy using the technique according to Griggs. DESIGN: Retrospective cohort study PATIENTS: Nineteen of 32 long-term surviving patients (mean follow-up duration, 17 months; range, 11 to 23 months) underwent a modified Griggs tracheostomy during their stay in the ICU following cardiothoracic surgery. INTERVENTIONS: Nineteen patients gave their informed consent for laryngotracheoscopy to localize and assess the percutaneous dilatational tracheostomy (PDT) puncture site, to evaluate the laryngotracheal morphology, and to quantify tracheal stenosis if present. In addition, specific symptoms of the upper airway tract were evaluated. RESULTS: At the time of examination, no clinically relevant cases of stenoses were found, although one patient had undergone surgical revision of the PDT for extensive granulation prior to our examination. The endoscopic examination revealed that 12 of 19 patients (63%) had tracheal stenoses > 10%, and 2 patients had tracheal stenoses > 25%. In 7 of 19 patients (32%), the cricoid cartilage was affected by the PDT site. Despite endoscopic guidance during PDT, the location of the puncture site was found to vary greatly. CONCLUSION: In contrast to recent reports on the long term outcome after Griggs PDT, we found tracheal stenoses > 10% in 63% of our patients. The grade of stenosis depended mainly on the puncture site of the PDT. Based on these results, we would emphasize the importance of adequate endoscopic guidance during PDT. Further studies are required in order to clarify the risk of long-term complications arising after PDT using the technique of Griggs. PMID- 12114361 TI - Effect of a short course of clarithromycin therapy on sputum production in patients with chronic airway hypersecretion. AB - STUDY OBJECTIVE: Long-term administration of macrolide antibiotics reduces sputum production in patients with chronic airway diseases, probably by inhibiting airway inflammation. The objective of the present study was to determine the acute effects of a macrolide on airway chloride secretion and sputum production. METHODS: We first investigated the effect of erythromycin treatment on chloride diffusion potential difference (CPD) across tracheal mucosa in vivo. Next, we conducted a double-blind, parallel-group study examining the effect of 7 days of treatment with clarithromycin (400 mg/d), amoxicillin (1,500 mg/d), or cefaclor (750 mg/d) in patients with chronic bronchitis or bronchiectasis without apparent respiratory infection. RESULTS: IV administration of erythromycin decreased the CPD of rabbit tracheal mucosa in a dose-dependent manner. Treatment of patients with clarithromycin decreased sputum production, whereas amoxicillin and cefaclor treatment had no effect. The percentage of patients whose sputum decreased > 30% from baseline (responders) was 38% in the clarithromycin group, 7% in the amoxicillin group, and 0% in the cefaclor group. During treatment with clarithromycin, the sputum solid composition increased and chloride concentration decreased in responders, but these changes were not observed in nonresponders. CONCLUSION: Short-term administration of 14-membered macrolide reduces chronic airway hypersecretion, presumably by inhibiting chloride secretion and the resultant water secretion across the airway mucosa. PMID- 12114362 TI - Pharmacokinetics and bioavailability of aerosolized tobramycin in cystic fibrosis. AB - STUDY OBJECTIVES: To describe the pharmacokinetics and bioavailability of inhaled tobramycin (TOBI; Chiron Corporation; Seattle, WA), 300-mg dose, delivered by a nebulizer (PARI LC Plus; Pari Respiratory; Richmond, VA) and a compressor (Pulmo Aide, model 5650D; DeVilbiss Health Care; Somerset, PA) in cystic fibrosis (CF) patients during the pivotal phase III trials. DESIGN: Data from two identical, 24 week, randomized, double-blind, placebo-controlled, parallel-group studies. SETTING: US sites randomized 258 patients with CF to receive tobramycin, 300 mg twice daily, in three 28-day on/28-day off treatment cycles. MEASUREMENT: Tobramycin sputum concentrations were assessed 10 min after the first and last doses were administered in the 20-week study. Serum tobramycin concentrations were assessed before and 1 h after the first and last doses had been administered. The population estimate of the apparent clearance was used to estimate the bioavailability fraction. RESULTS: The mean peak sputum concentration was 1,237 microg/g. About 95% of patients achieved sputum concentrations > 25 times the minimum inhibitory concentration of the Pseudomonas aeruginosa isolates. One hour after the dose, the mean serum concentration was 0.95 microg/mL. Tobramycin did not accumulate in the sputum or serum over the course of the study. Pharmacokinetic data were best represented by a two compartment model with biexponential decay and slope estimates comparable to those following parenteral administration. The estimated systemic bioavailability after aerosol administration was 11.7% of the nominal dose. CONCLUSIONS: The administration of tobramycin, 300 mg bid, in a 28-day off/28-day on regimen produced low serum tobramycin concentrations, reducing the potential for systemic toxicity. High sputum concentrations ensure efficacious antibiotic levels at the site of the infection. Inhaled tobramycin significantly improved the therapeutic ratio over that of parenteral aminoglycosides. PMID- 12114363 TI - Thalidomide for chronic sarcoidosis. AB - STUDY OBJECTIVES: Thalidomide therapy has been shown to modify granulomatous diseases, such as tuberculosis and leprosy. Lupus pernio is a skin manifestation of sarcoidosis that does not remit spontaneously, and was used as a marker of efficacy of thalidomide for sarcoidosis. DESIGN: An open-label, dose-escalation trial of thalidomide. SETTING: Patients were seen at one of four specialized sarcoidosis clinics in the United States. PATIENTS: Fifteen patients with lupus pernio and other manifestations of sarcoidosis unresponsive to prior therapy were enrolled. INTERVENTIONS: Skin lesions were assessed with visual examination by the treating physician, and photographic evaluation by a blinded panel of physicians reviewing photographs of the lesions before and after therapy. MEASUREMENTS AND RESULTS: Fourteen patients completed 4 months of therapy. All patients experienced some improvement in their skin lesions subjectively, and 10 of 12 evaluable patients showed improvement using photograph scoring. Five patients were better after 1 month (treated with 50 mg/d of thalidomide), seven more patients improved after 2 months (treated with 100 mg/d of thalidomide in the second month), and two patients required an additional month of 200 mg of thalidomide to achieve a response. Patients reported increased somnolence (n = 9), numbness (n = 7), dizziness (n = 2), constipation (n = 6), rash (n = 1), and increasing shortness of breath (n = 1). One patient discontinued therapy because of new-onset dyspnea, due to probably unrelated new-onset congestive heart failure. CONCLUSION: Thalidomide was an effective form of treatment for chronic cutaneous sarcoidosis. The drug was well tolerated and may be a useful alternative to systemic corticosteroids. PMID- 12114364 TI - Outcomes for patients with sarcoidosis awaiting lung transplantation. AB - STUDY OBJECTIVES: To describe the population of patients with sarcoidosis listed for orthotopic lung transplantation (OLT) in the United States, and to determine outcomes for these subjects relative to persons awaiting OLT for idiopathic pulmonary fibrosis (IPF). DESIGN: Retrospective analysis of the United Network for Organ Sharing transplant database over the period between January 1995 and December 2000. PATIENTS: All patients listed for OLT with an underlying diagnosis of either sarcoidosis or IPF. MEASUREMENTS AND RESULTS: During the study period, 427 patients with sarcoidosis and 2,115 patients with IPF were registered on the list for OLT. Demographically, the patients with sarcoidosis were younger and more likely to be female African Americans than were patients with IPF. Pulmonary function was worse in patients with sarcoidosis. The mean FVC was 42.6% of predicted, as compared to 45.0% of predicted in patients with IPF (p = 0.0044). The FEV(1) also differed between the populations (36.0% vs 46.0% of predicted for patients with sarcoidosis and IPF, respectively; p < 0.0001). Only 30.1% of patients with sarcoidosis and 32.4% of patients with IPF lacked functional limitations. For the subset of patients with hemodynamic data available, the mean pulmonary artery pressure was significantly higher in the sarcoidosis population (34.4 mm Hg vs 25.6 mm Hg, respectively; p < 0.0001). Neither the pulmonary capillary wedge pressure nor the cardiac index differed between the groups. Patients with sarcoidosis were less likely to receive a transplant. Approximately 30% of patients with sarcoidosis underwent OLT, compared to 37.3% of IPF patients (p = 0.0102). For those who did undergo transplantation, the median wait until OLT was 803 days for patients with sarcoidosis compared to 555 days for patients with IPF (p < 0.0001). Mortality rates were similar in both groups. In the sarcoidosis group, 28.1% of patients died, compared to 31.1% of patients with IPF (p = not significant). CONCLUSIONS: Patients with sarcoidosis are at as high a risk for mortality as patients with IPF while awaiting transplantation. Nonetheless, patients with sarcoidosis are less likely to undergo OLT. Pulmonary hypertension is a major concern in patients with advanced sarcoidosis awaiting transplantation. PMID- 12114365 TI - Validity of scoring systems to predict risk of prolonged mechanical ventilation after coronary artery bypass graft surgery. AB - STUDY OBJECTIVE: Two scoring systems, (the Spivack scoring system [SSS] and the cardiac risk score [CRS]), have been proposed to predict the risk of prolonged mechanical ventilation (PMV) after coronary artery bypass graft surgery (CABG). The primary objective of this study was to validate the efficacy of these scoring systems to predict the risk of PMV. DESIGN: Prospective observational study. SETTING: Cardiovascular surgical ICU. PATIENTS: Three hundred forty-eight patients underwent CABG. Following surgery, patients were extubated by a standardized respiratory weaning protocol. MEASUREMENTS AND RESULTS: Forty-nine percent of patients had SSS > 0 and had significantly longer duration of mechanical ventilation. The sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of the SSS for failure to extubate at 48 h are 80, 49, 9%, and 98%, respectively. Two hundred thirty-two patients (67.5%), 101 patients (29%), and 12 patients (3.5%) had a CRS of 0 to 4, 5 to 8, and > 8, respectively. Patients with lower scores had shorter duration of mechanical ventilation. The sensitivity, specificity, PPV, and NPV of the CRS for failure to extubate at 10 h are 42, 73, 47% and 69%, respectively. CONCLUSION: The SSS may be used as a preoperative screening tool. A simple questionnaire that includes history of unstable angina, diabetes, congestive heart failure, and smoking prior to hospital admission can be used to calculate the SSS. Patients with SSS 24 h and PMV > 48 h, respectively. MEASUREMENTS AND RESULTS: One hundred sixty-seven patients (41.75%), 27 patients (6.75%), and 21 (5.25%) patients, respectively, could not be extubated at 8, 24, and 48 h. Depressed level of consciousness was the most common reason for FTE in 58 of 167 patients (34.7%). The main cause of depressed level of consciousness was prolonged sedation due to anesthetic agents (51 patients; 30.5%). Hypoxemia was the most common cause for PMV for > 24 h (15 patients) and PMV > 48 h (13 patients). The causes of hypoxemia were cardiogenic and noncardiogenic pulmonary edema, pneumonia, and "hypoxemia of unknown etiology." Tachypnea due to acid-base disturbances was a reason for FTE and PMV for > 24 h in 27 and 3 patients, respectively. Cardiovascular instability was a rare reason for FTE. Postoperative bleeding was a cause for PMV in 18 patients. Four patients had more than a single reason for FTE at each assessment. Different causes have a variable effect on the duration of mechanical ventilation. CONCLUSION: The causes of PMV are heterogeneous, vary with time, and have a variable impact on the duration of mechanical ventilation required after the patient undergoes CABG surgery. PMID- 12114367 TI - Prognostic factors of non-HIV immunocompromised patients with pulmonary infiltrates. AB - STUDY OBJECTIVES: To assess the outcome and the prognostic factors in 200 non-HIV immunocompromised patients with pulmonary infiltrates (PIs). DESIGN: Prospective observational study. SETTING: An 800-bed university hospital. PATIENTS: Two hundred non-HIV immunocompromised patients (hematologic malignancies, 79 patients; hematopoietic stem cell transplants [HSCTs], 61 patients; and solid organ transplants, 60 patients). METHODS: Investigation of prognostic factors related to mortality using a multiple logistic regression model. RESULTS: Specific diagnosis of the PI was obtained in 78% of the cases (infectious origin was determined in 74%). The overall mortality rate was 39% (78 of 200 patients). Patients with HSCT had the highest mortality rate (53%). A requirement for mechanical ventilation (odds ratio [OR], 28; 95% confidence interval [CI], 9 to 93), an APACHE (acute physiology and chronic health evaluation) II score of > 20 (OR, 5.5; 95% CI, 2 to 14.7), and a delay of > 5 days in establishing a specific diagnosis (OR, 3.4; 95% CI, 1.2 to 9.6) were the variables associated with mortality at the multivariate analysis. The subgroup analysis based on underlying disease confirmed the prognostic significance of these variables and the infectious etiology for the PI. CONCLUSIONS: Mortality in immunocompromised patients is high, particularly in patients undergoing HSCT. Achieving an earlier diagnosis potentially may improve the mortality rate of these patients. PMID- 12114368 TI - Clinical importance of delays in the initiation of appropriate antibiotic treatment for ventilator-associated pneumonia. AB - STUDY OBJECTIVES: To determine the influence of initially delayed appropriate antibiotic treatment (IDAAT) on the outcomes of patients with ventilator associated pneumonia (VAP). SETTING: Medical ICU of Barnes-Jewish Hospital, St. Louis, a university-affiliated urban teaching hospital. PATIENTS: One hundred seven consecutive patients receiving mechanical ventilation and antibiotic treatment for VAP. INTERVENTIONS: Prospective patient surveillance and data collection. MEASUREMENTS AND RESULTS: All 107 patients eventually received treatment with an antibiotic regimen that was shown in vitro to be active against the bacterial pathogens isolated from their respiratory secretions. Thirty-three patients (30.8%) received antibiotic treatment that was delayed for >or= 24 h after initially meeting diagnostic criteria for VAP. These patients were classified as receiving IDAAT. The most common reason for the administration of IDAAT was a delay in writing the antibiotic orders (n = 25; 75.8%). The mean time (+/- SD) interval from initially meeting the diagnostic criteria for VAP until the administration of antibiotic treatment was 28.6 +/- 5.8 h among patients classified as receiving IDAAT, compared to 12.5 +/- 4.2 h for all other patients (p < 0.001). Forty-four patients (41.1%) with VAP died during their hospitalization. Increasing APACHE (acute physiology and chronic health evaluation) II scores (adjusted odds ratio, 1.13; 95% confidence interval, 1.09 to 1.18; p < 0.001), presence of malignancy (adjusted odds ratio, 3.20; 95% confidence interval, 1.79 to 5.71; p = 0.044), and the administration of IDAAT (adjusted odds ratio, 7.68; 95% confidence interval, 4.50 to 13.09; p < 0.001) were identified as risk factors independently associated with hospital mortality by logistic regression analysis. CONCLUSION: These data suggest that patients classified as receiving IDAAT are at greater risk for hospital mortality. Clinicians should avoid delaying the administration of appropriate antibiotic treatment to patients with VAP in order to minimize their risk of mortality. PMID- 12114369 TI - Chronic intermittent asphyxia impairs rat upper airway muscle responses to acute hypoxia and asphyxia. AB - BACKGROUND: Obstructive sleep apnea (OSA) is a major clinical disorder that is characterized by multiple episodes of upper airway obstruction due to the failure of the upper airway dilator muscles to maintain upper airway patency. This results in chronic intermittent asphyxia (CIA) due to repetitive apneas, but very little is known about the effects of CIA on upper airway muscle function. OBJECTIVE: To test the hypothesis that CIA affects upper airway muscle activity and electromyogram (EMG) responses to acute hypoxia and asphyxia. DESIGN: Record upper airway EMG responses to acute hypoxia and asphyxia in control and CIA treated rats. SETTING: Department of Physiology, Royal College of Surgeons in Ireland, Dublin, Ireland. MEASUREMENTS: Sternohyoid (SH) muscle and diaphragm (DIA) muscle EMG activities were recorded in both groups during normoxia, hypoxia (7.5% O(2) in N(2)), and asphyxia (7.5% O(2) and 3% CO(2)) under pentobarbitone anesthesia. RESULTS: Baseline SH EMG activity was significantly elevated in the CIA-treated rats compared to the controls, whereas DIA EMG activity was similar in the two groups. In addition, CIA significantly reduced SH EMG but not DIA EMG responses to acute hypoxia and asphyxia. CONCLUSIONS: The elevated upper airway muscle activity associated with OSA in humans during wakefulness is due at least in part to CIA. We propose that a reduction in the response of upper airway dilator muscles to acute asphyxia following upper airway obstruction is likely to cause further asphyxic insult, leading to a vicious feed-forward cycle exacerbating the condition. Our results suggest that CIA contributes to the pathophysiology of sleep-disordered breathing. PMID- 12114370 TI - Efficacy of dye-stained enteral formula in detecting pulmonary aspiration. AB - STUDY OBJECTIVE: To determine the extent to which a mixture of human gastric juice and enteral formula stained with two concentrations of FD&C Blue No. 1 food dye (0.8 and 1.5 mL/L) is visible in suctioned tracheobronchial secretions following three forced small-volume pulmonary aspirations over a 6-h period in an animal model. DESIGN: Experimental 2 x 3 repeated measures. SETTING: Animal laboratory and an acute care hospital. PARTICIPANTS: Ninety New Zealand white rabbits weighing approximately 3 kg each, and 90 acutely ill adults who furnished gastric juice. INTERVENTIONS: A mixture of human gastric juice and enteral formula stained with 0.8 or 1.5 mL of dye per liter was instilled intratracheally over a 30-min period into anesthetized intubated animals at baseline, 2 h, and 4 h. A total of 0.4 mL/kg of the mixture was instilled at each session. Ninety minutes after each instillation, suctioned secretions were examined for visible dye and blood. MEASUREMENTS AND RESULTS: Dye was visible in 46.3% of the secretions (125 of 270). The concentration of dye had no significant effect on dye visibility. Blood that was present in 114 of 270 of the secretions (42.2%) interfered with dye visibility in all but two secretions. For reasons unknown, even in the absence of blood, dye visibility decreased from 90.2% (55 of 61 secretions) after the first aspiration event to only 61% (25 of 41 secretions) after the third aspiration event. CONCLUSIONS: Findings from this animal model study do not support the use of the dye method to detect repeated small-volume aspirations. For clinicians who choose to use the dye method in selected situations, it appears that a dye concentration of 0.8 mL/L may be as effective in detecting aspiration as a 1.5 mL/L concentration. PMID- 12114371 TI - Establishment of 15 cancer cell lines from patients with lung cancer and the potential tools for immunotherapy. AB - BACKGROUND: Since lung cancer is the major cause of death not only in Japan but in many other industrialized countries, the development of new therapeutic modalities is quite important. In patients with melanoma, immunotherapy with some tumor antigens has been shown to result in tumor regression. However, little is known about specific immune responses and tumor antigens in lung cancer, due to difficulty in establishing appropriate lung cancer cell lines. In order to resolve these difficulties, we tried to establish and characterize lung cancer cell lines as useful tools for the analysis of tumor-specific immune responses in patients with lung cancer. MATERIALS AND METHODS: We tried to establish lung cancer cell lines from 549 patients with resectable lung cancer and from 21 patients with pleural and pericardial effusions or lymph node metastasis. We characterized the established cell lines after the induction of tumor-specific cytotoxic T lymphocytes (CTLs), and analyzed both the major histocompatibility complex (MHC) class I and class II molecules on their surfaces. RESULTS: We succeeded in establishing 15 lung cancer cell lines from 570 specimens (2.6%). The success rate of the establishment of lung cancer cell lines was significantly higher in patients at such advanced stages as MHC III and IV than in those at MHC stages I and II (p = 0.004). MHC class I molecules were expressed in 12 of 15 cell lines (80%), while MHC class II molecules were found in 3 of 15 cell lines (20%) on their cell surfaces by flow cytometry. A haplotype loss of MHC class I antigens was found in 6 of 15 cell lines (40%). Although CTLs were induced in only two of eight cell lines tried by stimulation with nontransduced autologous tumor cell lines, CTLs were successfully induced in all of eight cell lines tested by stimulation with CD80-transfected autologous tumor cells. CONCLUSIONS: These results suggested that the tumor antigens recognized by CTLs could thus exist in the tumor cells derived from many lung cancer patients. It is, therefore, possible that antigen-specific immunotherapies may be potentially effective for patients with lung cancer by adoptive transfer of CTLs, as well as by vaccine therapy using tumor-specific antigens. PMID- 12114372 TI - A randomized, placebo-controlled trial of a leukotriene synthesis inhibitor in patients with COPD. AB - STUDY OBJECTIVE: Patients with COPD classically have neutrophilic bronchial inflammation and raised airway concentrations of the neutrophil chemoattractant leukotriene B(4) (LTB(4)). A small phase II trial was conducted to assess the effects of a leukotriene synthesis inhibitor on bronchial inflammation in patients with stable COPD. DESIGN: A randomized, double-blind, placebo controlled, parallel-group study. SETTING: Respiratory medicine department of a university hospital. PATIENTS AND INTERVENTION: Seventeen patients with chronic bronchitis and COPD (mean FEV(1), 35.5% predicted; SD, 14.8% predicted) were randomized to receive 14 days of the oral leukotriene synthesis inhibitor BAYx1005 (500 mg bid) or placebo. MEASUREMENTS AND RESULTS: Spontaneous sputum samples obtained at baseline and at the end of treatment were assayed for LTB(4), myeloperoxidase (an indirect marker of neutrophil numbers and/or activation), and chemotactic activity (Boyden chamber). After 14 days, there were no significant differences (p > 0.05) in absolute LTB(4) concentrations between the two treatment groups. However, BAYx1005 treatment produced a significantly greater median reduction in LTB(4) of - 3.1 nM (interquartile range [IQR], - 9.6 to - 0.2 nM) vs 3.0 nM (IQR, - 0.3 to 8.5 nM) [p = 0.001], with concentrations decreasing from 8.0 nM (IQR, 4.3 to 24.4 nM) at baseline to 4.2 nM (IQR, 1.9 to 11.9 nM) at the end of treatment (p = 0.03). There were no changes in the placebo group and no differences in sputum myeloperoxidase concentration or chemotaxis between the two treatment arms (p > 0.05). CONCLUSIONS: This small study suggests that a leukotriene synthesis inhibitor can produce modest reductions in some measures of neutrophilic bronchial inflammation in patients with COPD. This class of anti inflammatory agent requires further study in larger numbers of patients to determine clinical benefit. PMID- 12114373 TI - Public health implications of voters' attitudes regarding statewide tobacco policy. AB - BACKGROUND: Tobacco use remains the most preventable cause of death and disability in the United States. Public opinion regarding tobacco use is not only an important barometer of the likelihood of effective tobacco-control legislation, but also identifies ongoing public health educational needs. Because > 63,000 children become new smokers annually in Pennsylvania, we chose to evaluate the statewide public health tobacco perspective in order to help tailor future public policy interventions. STUDY DESIGN AND SETTING: Registered voters were randomly contacted in a statewide telephone survey. To reduce response bias, an independent polling firm conducted the 643 structured interviews. RESULTS: Most respondents were >or= 45 years old (55%), female (54%), and had at least some college education (62%). Twenty-eight percent (95% confidence interval [CI], 25 to 32%) were current tobacco users, and 38% (95% CI, 34 to 42%) had lost family members or friends to smoking-related disease. Ninety-two percent (95% CI, 90 to 94%) expressed "concern" about adolescent tobacco use, but only 46% (95% CI, 42 to 50%) believed that government needed to do more. Of respondents opposed to government involvement, 65% (95% CI, 61 to 68%) believed it was an improper role for government, or that there are more important non-health government priorities. When framed more personally, 80% (95% CI, 77 to 83%) indicated that elected officials have a responsibility to "dedicate a significant portion of tobacco settlement" to prevention. Still, 28% (95% CI, 25 to 32%) would oppose laws restricting smoking in establishments frequented by youth. CONCLUSIONS: Prior public health education initiatives have been effective in shaping the tobacco-related health concerns of Pennsylvania voters. As expected, the overwhelming majority of respondents are concerned about youth tobacco use and agree that money should be spent on tobacco-control initiatives. In contrast, many are reluctant to support "government" involvement in what is still seen as a personal issue. Future public health initiatives should focus on this dichotomy and should highlight the utility of an integrated policy approach to tobacco control. PMID- 12114374 TI - The problem with diagnosis related group 475. AB - The reimbursement of hospital Part A cost by Medicare under the original prospective payment system (PPS) created serious financial problems for hospitals in many areas but was especially serious in patients receiving mechanical ventilation. Subsequent revisions of the PPS corrected some of the financial burden of the cost of such patients when the patient required tracheostomy for prolonged ventilator care, Diagnosis Related Group (DRG) 483. The problem that remains, however, is that only patients with a medical diagnostic classification (MDC) within the MDC 4 area, Diseases and Disorders of the Respiratory System, qualify for DRG 475, the only DRG other than DRG 483 that recognizes the cost of mechanical ventilation. This study evaluates the Part A costs of medical and surgical patients who received mechanical ventilation for >or= 3 days during 1 year at Saint Marys Hospital and Rochester Methodist Hospital in Rochester, MN, who did not qualify for DRG 475. The analysis of the financial effect of these patients under the Medicare system reveals a significant monetary loss and this is compared to other payor groups. PMID- 12114375 TI - Fungal endocarditis, 1995-2000. AB - One hundred fifty-two cases of fungal endocarditis (FE) were identified in the English-language literature between January 1, 1995, and June 30, 2000. Although the median age of patients (44 years) was relatively young, injection drug use was identified as a risk factor in only 4.1% of cases. Other factors, including underlying cardiac abnormalities (47.3%), prosthetic valves (44.6%), and central venous catheters (30.4%), were more commonly identified as predisposing conditions and reflect the changing epidemiology of the syndrome. Unfortunately, mortality remains unacceptably high, particularly for patients with Aspergillus related FE. Novel therapies are needed to improve patient outcomes. PMID- 12114376 TI - Clinical, diagnostic, and management perspectives of aortic dissection. AB - The incidence of aortic dissection ranges from 5 to 30 cases per million people per year, depending on the prevalence of risk factors in the study population. Although the disease is uncommon, its outcome is frequently fatal, and many patients with aortic dissection die before presentation to the hospital or prior to diagnosis. While pain is the most common symptom of aortic dissection, more than one-third of patients may develop a myriad of symptoms secondary to the involvement of the organ systems. Physical findings may be absent or, if present, could be suggestive of a diverse range of other conditions. Keeping a high clinical index of suspicion is mandatory for the accurate and rapid diagnosis of aortic dissection. CT scanning, MRI, and transesophageal echocardiography are all fairly accurate modalities that are used to diagnose aortic dissection, but each is fraught with certain limitations. The choice of the diagnostic modality depends, to a great extent, on the availability and expertise at the given institution. The management of aortic dissection has consisted of aggressive antihypertensive treatment, when associated with systemic hypertension, and surgery. Recently, endovascular stent placement has been used for the treatment of aortic dissection in select patient populations, but the experience is limited. The technique could be an option for patients who are poor surgical candidates, or in whom the risk of complications is gravely high, especially so in the patients with distal dissections. The clinical, diagnostic, and management perspectives on aortic dissection and its variants, aortic intramural hematoma and atherosclerotic aortic ulcer, are reviewed. PMID- 12114377 TI - Improved survival and higher mortality: the conundrum of lung cancer screening. PMID- 12114378 TI - Effects of a contemporary, exercise-based rehabilitation and cardiovascular risk reduction program on coronary patients with abnormal baseline risk factors. AB - Phase II cardiac rehabilitation programs are associated with improvements in exercise tolerance, coronary risk factors, and psychosocial well-being. Nevertheless, previous reports have generally evaluated the global effectiveness of these programs (ie, on all subjects, collectively), which may serve to camouflage or attenuate the impact of these interventions on specific patient subsets. In this study, we investigated the effectiveness of a contemporary, exercise-based cardiac rehabilitation program that included a cardiovascular risk reduction intervention, using a computerized database on 117 patients (average age, 66.5 years; 68% men; 96% white) who completed pre-phase II and post-phase II evaluations. Exercise training involved three 45- to 60-min sessions per week at minimum of 40 to 50% to a maximum of 75% oxygen uptake for 6 to 8 weeks. The effectiveness of the exercise training program was substantiated by significant (p 1.8 million. Between February 1994 and September 2001, seven patients with tuberculous tracheobronchial stenosis (mean age, 43 years) underwent a total of 11 dilatations with placement of 10 straight stents and 1 Y stent. Under general anesthesia, all patients underwent rigid bronchoscopy and dilatation of the stenosis with placement of a Dumon stent. There were no deaths. One patient developed a pneumothorax. Two patients experienced migration of the stent, which required reintervention for adjustment of position of the stent. The stents were left in situ for a mean period of 32 months. There was marked improvement in dyspnea in all patients after the procedure, as determined by visual analog scale. Endoscopic dilatation with placement of a silicone stent is an effective treatment for patients with tuberculous tracheobronchial stenosis. PMID- 12114386 TI - Central bronchopleural fistulas closed by bronchoscopic injection of absolute ethanol. AB - Five consecutive bronchopleural fistulas (BPFs) were successfully treated by injecting absolute ethanol directly into the submucosal layer of the fistula under flexible bronchoscopic observation. No complications occurred as a result of this treatment. Our nonsurgical treatment may be very useful to reduce the costs of and duration of hospitalization and to improve the patient's quality of life. This is the first report of the bronchoscopic closure of BPFs by injecting absolute ethanol, and we would recommend this treatment as a first-line therapy for patients with a postoperative central BPF with an orifice that is < 3 mm in diameter. PMID- 12114387 TI - Severe erythroderma as a complication of continuous epoprostenol therapy. AB - Epoprostenol is a vasodilator that is produced by vascular endothelial cells and is currently the "gold standard" therapy for patients with severe primary pulmonary hypertension or pulmonary hypertension secondary to collagen vascular disease. Hypersensitivity to the drug has not been reported. We report a case of a patient with pulmonary hypertension and undifferentiated connective tissue disease who, after 2 months of treatment with epoprostenol, presented with rapidly progressive erythema, scaling, nausea and vomiting, and fever. Test results from a skin biopsy specimen were consistent with a drug reaction. The patient' condition improved after rapid tapering of her epoprostenol and administration of corticosteroids. Epoprostenol may be associated rarely with severe erythroderma. PMID- 12114388 TI - Factors influencing the serum osmolar gap. PMID- 12114389 TI - Behcet disease, Adamantiades-Behcet disease, or Hippocrates-Adamantiades-Behcet disease? PMID- 12114390 TI - Does rinsing the mouth before expectoration improve sputum specimen quality? PMID- 12114391 TI - Immersion pulmonary edema in special forces combat swimmers. PMID- 12114392 TI - Exercise testing in the consulting room. PMID- 12114393 TI - Underwater photography in the human airway: use of saline solution to visualize peripheral tumors and evaluate airway rigidity. PMID- 12114394 TI - Transient lactic acidosis as a side effect of inhaled salbutamol. PMID- 12114395 TI - Self-induced subcutaneous emphysema and pneumomediastinum. PMID- 12114396 TI - Peroxisome proliferator-activated receptor-gamma expression in lung. PMID- 12114399 TI - A new marker determining clonal outgrowth. PMID- 12114400 TI - Apoptosis: target of cancer therapy. AB - Recent knowledge on apoptosis has made it possible to devise novel approaches, which exploit this process to treat cancer. In this review, we discuss in detail approaches to induce tumor cell apoptosis, their mechanism of action, stage of development, and possible drawbacks. Finally, the obstacles yet to be overcome and the perspectives for potential clinical use of apoptosis-triggering approaches in cancer therapy in the future are assessed. PMID- 12114401 TI - Validation of the pharmacodynamics of BMS-247550, an analogue of epothilone B, during a phase I clinical study. AB - The primary aims of this study were to evaluate the timecourse and dose response of microtubule bundle formationin peripheral blood mononuclear cells (PBMCs) and to correlate these data with BMS-247550 pharmacokinetics. The data presented here were obtained from 17 patients enrolled in a Phase I trial who received five dose levels of BMS-247550 (7.4-59.2 mg/m(2)), given as a 1-h infusion once every 3 weeks. Plasma drug exposure or area under the curve (AUC), and tubulin bundle formation in PBMCs were assessed in cycles 1 and 2. Similar analyses were also performed on tumor biopsies from one eligible patient. PBMCs exhibited dramatic microtubule bundle formation 1 h after infusion that declined by 24 h, showing a positive correlation with AUC((0-24)) for cycles 1 and 2. A similar pattern of tubulin bundle formation also was observed in a smaller proportion of breast tumor cells from one patient who exhibited a partial response to BMS-247550. This patient's tumor expressed multidrug resistance (MDR1) and MDR-associated protein (MRP1), and in addition poly(ADPribose) polymerase cleavage, a marker of cell death, was observed within 23 h after drug infusion. This patient was also heterozygous for a novel polymorphism at the extreme COOH terminus of beta tubulin (Gly 437 Gly/Ser), although the relevance of the polymorphism to the response is unknown. In summary, microtubule bundle formation in PBMCs occurs within 1 h of treatment with BMS-247550 and is related to plasma AUC. Similar bundle formation was seen in one tumor sample, despite expression of MDR1 and MRP1. Cell death occurred 23 h after peak microtubule bundle formation in these tumor cells. These findings validate in vitro pharmacodynamic observations. PMID- 12114403 TI - Infusion of CD4+ donor lymphocytes induces the expansion of CD8+ donor T cells with cytolytic activity directed against recipient hematopoietic cells. AB - PURPOSE: Donor lymphocyte infusions (DLIs) provide effectivetherapy for patients with relapsed chronic myeloid leukemia after allogeneic bone marrow transplantation. Previous studies have suggested that depletion of CD8+ T cells from the infused donor lymphocytes can reduce the incidence of graft-versus-host disease associated with DLI without reducing antileukemia activity. In this situation however, the immune effector cells responsible for tumor rejection have not been identified. The goal of this study was to characterize these effector populations. EXPERIMENTAL DESIGN: We studied three representative patients with relapsed chronic myeloid leukemia who achieved complete molecular remission after receiving CD8+ T-cell-depleted DLI from HLA-identical sibling donors. Effector T cells were characterized in patient samples after in vitro stimulation and functional assessment. T-cell clones relevant to the immune response were then isolated and further characterized. RESULTS: Analysis of peripheral blood samples collected after DLI indicated the presence of a high frequency of circulating host-reactive cytolytic CD8+ T cells secreting IFN-gamma. These HLA class I restricted CTLs were specific for recipient minor histocompatibility antigens (mHAs) because they did not recognize target cells of donor origin. One CTL clone was further expanded in vitro and shown to recognize a broadly expressed mHA presented by HLA-B5701. Using a molecular approach, we demonstrated that this clone was expanded in peripheral blood and marrow after DLI. It was not detected before DLI. CONCLUSIONS: These results indicate that CD4+ DLI elicits a potent allogeneic response mediated by mHA-specific CD8+ T cells. PMID- 12114402 TI - Active specific immunotherapy with a beta-human chorionic gonadotropin peptide vaccine in patients with metastatic colorectal cancer: antibody response is associated with improved survival. AB - PURPOSE: Human chorionic gonadotropin (hCG) is produced by colorectalcancers and may play a role in its progression. The clinical and immunological effects of a synthetic vaccine targeting beta-hCG composed of the COOH terminal peptide of beta-hCG (CTP37) conjugated to diphtheria toxoid (DT) was investigated in patients with metastatic colorectal cancer. EXPERIMENTAL DESIGN: Seventy-seven patients from 12 centers were randomly divided into two vaccine dose groups. CTP37-DT was formulated in an emulsion and administered i.m. on days 0, 28, and 70. Patients were evaluated for toxicity, overall survival, and antibody response to hCG and to DT. RESULTS: Immunizations were well tolerated with no patients requiring cessation of the injections. Anti-hCG antibody was detected in 56 of the 77 patients treated. Significant differences in antibody response and survival were not observed between the two dose groups. An intention-to-treat analysis of all vaccinated patients showed a median survival of 34 weeks. Patients who developed anti-hCG antibody levels higher than or equal to the median value exhibited a median survival of 45 weeks compared with 24 weeks for patients who developed anti-hCG antibody levels lower than the median (P = 0.0002). In contrast, no significant difference was observed when comparing survival based upon the level of DT antibody that developed (P = 0.80). CONCLUSIONS: Vaccination with CTP37-DT induced anti-hCG antibodies in most patients with advanced colorectal cancer. Anti-hCG antibody induction was associated with longer overall survival. CTP37-DT has an excellent safety profile and warrants further study in patients with colorectal cancer. PMID- 12114404 TI - Frequent germ-line succinate dehydrogenase subunit D gene mutations in patients with apparently sporadic parasympathetic paraganglioma. AB - PURPOSE: Recently, familial paraganglioma (PGL) was shown to be caused bymutations in the gene encoding succinate dehydrogenase subunit D (SDHD). However, the prevalence of SDHD mutations in apparently sporadic PGL is unknown. We studied the frequency and spectrum of germ-line and somatic SDHD mutations in patients with parasympathetic PGL. EXPERIMENTAL DESIGH: We studied 57 unselected patients who developed parasympathetic PGLs (n = 105 tumors) and who were treated between 1987 and 1999 at the Erasmus MC (Rotterdam, the Netherlands). Thirty eight (67%) of these patients (n = 51 tumors) lacked a family history of parasympathetic PGL. We used conformation-dependent gel electrophoresis and sequence determination analysis of germ-line and tumor DNA to identify SDHD mutations. We compared the clinical and molecular characteristics of sporadic and hereditary PGLs. RESULTS: Three different SDHD germ-line mutations were identified in 32 of the 57 (56%) patients. These included 19 of 19 (100%) patients with familial PGL and also 13 of 38 (34%) patients with apparently sporadic PGL. All three mutations were characterized as missense mutations (D92Y, L95P, and L139P) in highly conserved regions of the SDHD gene and were not observed in 200 control alleles. No somatic mutations were found. CONCLUSIONS: Germ-line mutations of the SDHD gene are present in a significant number of patients with apparently sporadic parasympathetic PGL. Somatic SDHD mutations do not play a significant role in the sporadic form of this tumor. Genetic testing for SDHD germ-line mutations should be considered for every patient presenting with this tumor, even if a personal or family history of PGL is absent, to allow appropriate clinical management. PMID- 12114405 TI - Suppression of type I interferon signaling proteins is an early event in squamous skin carcinogenesis. AB - PURPOSE: IFN-based therapy has been shown to be active in the treatmentof squamous cell carcinoma (SCC) of the skin, the most aggressive form of non melanoma skin cancer. Based largely on this activity, we began programmatically examining the expression of IFN-stimulated gene factor 3 (ISGF-3) proteins (signal transducers and activators of transcription 1alpha/beta, signal transducers and activators of transcription 2, and p48), which are important mediators of IFN-alpha signaling, in skin premalignancy and SCC. Our previous preliminary studies suggested suppression of some or all of the ISGF-3 proteins in skin SCC. EXPERIMENTAL DESIGN: To determine the timing of the suppression of IFN-alpha signaling proteins in squamous skin carcinogenesis, we have now compared ISGF-3 expression by immunohistochemical staining in biopsies of actinic keratosis, a form of skin premalignancy, and matched normal skin. RESULTS: We observed a significant decrease in expression of one or more ISGF-3 proteins in 76% of patients with actinic keratosis (19 of 25 patients). In addition, we found a suppression of one or more ISGF-3 proteins in 67% of skin SCC patients tested (12 of 18 patients), confirming our previous observations. CONCLUSIONS: These data have led to the hypothesis that the suppressed expression of ISGF-3 proteins and consequent reduction in responsiveness to endogenous IFN likely are an early event in skin carcinogenesis. PMID- 12114406 TI - Cytogenetic evidence that circulating epithelial cells in patients with carcinoma are malignant. AB - PURPOSE: Numerous studies of circulating epithelial cells (CECs)have been described in cancer patients, and genetic abnormalities have been well documented. However, with one exception in colorectal cancer, there has been no report of matching the genetic abnormalities in the CECs with the primary tumor. The purpose of this investigation was to determine (a) whether CECs in patients including those with early tumors are aneusomic and (b) whether their aneusomic patterns match those from the primary tumor, indicating common clonality. EXPERIMENTAL DESIGN: Thirty-one cancer patients had CECs identified by immunofluorescence staining using a monoclonal anti-cytokeratin antibody. Their CECs were analyzed by enumerator DNA probes for chromosomes 1, 3, 4, 7, 8, 11, or 17 by dual or tricolor fluorescence in situ hybridization. Touch preparations of the primary tumor tissue were available from 17 of 31 patients and hybridized with the same set of probes used to genotype the CECs. RESULTS: The number of CECs from each patient ranged from 1-92 cells/cytospin. CECs showed abnormal copy numbers for at least one of the probes in 25 of 31 patients. Touch preparations from the primary tumors of 13 patients with aneusomic CECs were available. The pattern of aneusomy matched a clone in the primary tumor in 10 patients. CONCLUSIONS: We conclude that the vast majority of CECs in breast, kidney, prostate, and colon cancer patients are aneusomic and derived from the primary tumor. PMID- 12114408 TI - Inactivation of the antiapoptotic phosphatidylinositol 3-kinase-Akt pathway by the combined treatment of taxol and mitogen-activated protein kinase kinase inhibition. AB - Paclitaxel (Taxol) activates a number of signal transduction pathways that lead to apoptosis. In contrast, paclitaxel also activates cell survival pathways, such as the Raf-mitogen-activated protein kinase kinase (MEK)-extracellular signal regulated kinase (ERK) pathway. Previously, we have shown that inhibition of MEK combined with paclitaxel treatment causes an impressive enhancement of apoptosis in various tumor cell lines. Here, we find that the combination of paclitaxel with a MEK inhibitor leads to a dramatic inactivation of the antiapoptotic Akt (protein kinase B) kinase. The decrease in Akt is not reflected at the protein or mRNA level but rather attributed to kinase inactivation. To confirm that inactivation of Akt is significant, a constitutively active Akt mutant was introduced and shown to reverse tumor cell apoptosis. Further analysis upstream of Akt shows that treatment with the combination of paclitaxel and MEK inhibitor down-regulates PI3K activity more than either agent alone. The direct pharmacological inhibition of phosphatidylinositol 3-kinase (PI3K) similarly enhances paclitaxel-induced tumor apoptosis in a dose-dependent manner. Our results suggest the combination of paclitaxel and MEK inhibitor leads to down regulation of the PI3K-Akt signaling in addition to the proapoptotic effects of paclitaxel and MEK inhibitor alone. Overall, these findings render the combined use of paclitaxel with MEK inhibitors, or paclitaxel with PI3K inhibitors, as a promising new strategy for cancer chemotherapy. PMID- 12114407 TI - The tumor suppressor gene LKB1 is associated with prognosis in human breast carcinoma. AB - PURPOSE: LKB1 (also called STK11) is a recently identified tumor suppressor gene in which its mutation can lead to Peutz-Jeghers syndrome, characterized by gastrointestinal polyps and cancers of different organ systems. Weak expression of this gene does occur at a certain frequency in sporadic breast cancer. This indicates that LKB1 gene may relate to the tumorigenesis of breast cancer. EXPERIMENTAL DESIGN: To investigate the function of the LKB1 gene in sporadic breast cancer, we reintroduced LKB1 into breast cancer cell lines which lack the LKB1 gene. Also, we examined the LKB1 protein expression in human breast cancer samples. RESULTS: We found that reintroducing LKB1 into breast cancer cell lines suppresses cell growth by G(1) cell cycle block. The LKB1-mediated G(1) cell cycle arrest is caused by up-regulation of the expression of p21(WAF1/CIP1) in breast cancer MDA-MB-435 cells. We also demonstrated that low LKB1 protein expression correlates with higher histological grade (P = 0.013), larger tumor size (P = 0.001), progesterone receptor status (P = 0.048), and presence of lymph node metastasis (P = 0.003). Furthermore, LKB1 low expression was associated with a higher relapse rate (P = 0.002) and a worse OS (P = 0.008). CONCLUSIONS: LKB1 plays a role in tumor suppressor function in human breast cancer. LKB1 expression may be a useful prognostic marker in human breast cancer. PMID- 12114410 TI - Measurement of perfusion in stage IIIA-N2 non-small cell lung cancer using H(2)(15)O and positron emission tomography. AB - PURPOSE: As the interest in antiangiogenesis therapy in oncology is rising, the need for in vivo techniques to monitor such therapy is obvious. Measurement of tumor perfusion using positron emission tomography and H(2)(15)O potentially is such a technique. The objective of the present study was to assess whether it is feasible to measure perfusion in vivo in non-small cell lung cancer (NSCLC) using H(2)(15)O and positron emission tomography. EXPERIMENTAL DESIGN: Fifteen dynamic H(2)(15)O and [(18)F]2-fluoro-2-deoxy-D-glucose ((18)FDG) studies were performed in 10 patients with stage IIIA-N2 NSCLC. Blood flow (BF) data were correlated with simplified methods of analysis (tumor:normal tissue ratio and standardized uptake value) and with glucose metabolism (MR(glu)). RESULTS: (18)FDG data were required for accurate definition of tumor and mediastinal lymph node metastases. There was large intertumor variation in BF. Correlation of simplified methods of analysis with quantitative BF was poor. In addition, BF and MR(glu) were not correlated. CONCLUSION: Measurement of BF in NSCLC using H(2)(15)O and (18)FDG is feasible. Simple uptake analysis, however, cannot be used as an indicator of perfusion. Whether BF can be used for response monitoring needs to be evaluated in a large patient study, where results can be compared with outcome. PMID- 12114409 TI - Coordinate expression of apoptosis-associated proteins in human breast cancer before and during chemotherapy. AB - PURPOSE: Induction of apoptosis is a key factor in the response of tumors to chemotherapy. Laboratory studies have established many of the factors that regulate and execute apoptosis, but the significance of these in human tumors is poorly understood. Therefore, the relationship between key components of this machinery was examined in primary human breast carcinomas before and 24 h after the initiation of chemotherapy. EXPERIMENTAL DESIGN: Apoptosis was measured using the terminal deoxynucleotidyl transferase-mediated nick end labeling assay, and proliferation was assessed using the anti-Ki67 antibody MIB-1. Monospecific polyclonal antibodies were used for immunohistochemical detection of Bcl-2, Bax, XIAP, activated (cleaved) caspase 3 and 6, and cleaved DNA Fragmentation Factor 40 (DFF40) using paraffin-embedded tissues. RESULTS: Before treatment, a significant correlation was found between apoptosis and proliferation (r = 0.64, P < 0.0001), between caspases 3 and 6 (r = 0.49, P = 0.004) and between cleaved DFF40 and active caspases 3 (r = 0.66, P < 0.0001) or 6 (r = 0.47, P = 0.006). Before treatment, expression of inhibitor of apoptosis protein, XIAP, also correlated positively with cleaved caspase 3 (r = 0.64, P < 0.0001), caspase 6 (r = 0.36, P = 0.04), and DFF40 (r = 0.61, P = 0.0001). At 24 h after chemotherapy, significant increases in apoptosis and decreases in proliferation were observed, with the degree of increase in apoptosis inversely associated with decrease in proliferation. Chemotherapy-induced increases in Bax were correlated with increases in cleaved DFF40 (r = 0.54, P = 0.0008), but no other variables showed significant change at 24 h after initiation of chemotherapy. CONCLUSION: The pretreatment biomarker relationships suggest parallel cleavage and activation of these executioner proteins in breast cancer and that XIAP may maintain cell survival in the face of caspase activation. The findings provide in vivo evidence in human breast cancer that chemotherapy induces an apoptotic program characterized by up-regulation of Bax and cleavage of caspase substrate DFF40. PMID- 12114411 TI - Phase I and pharmacokinetic study of once daily oral administration of S-1 in patients with advanced cancer. AB - PURPOSE: To determine the maximum tolerated dose, dose-limiting toxicities(DLTs), and pharmacokinetics of S-1, a combination of tegafur, 5-chloro-2,4 dihydroxypyridine (CDHP), and oxonic acid, administered once daily in patients with advanced cancer. EXPERIMENTAL DESIGN: Eighteen patients with refractory malignancies were treated with S-1 administered once daily for 21 consecutive days, followed by a 1-week break. Of 16 evaluable patients, 6 were treated at a dose of 50 mg/m(2)/day, and 10 were treated at 60 mg/m(2)/day. RESULTS: DLTs were observed in 1 of 6 evaluable patients treated with 50 mg/m(2)/day and in 4 of 10 evaluable patients treated with 60 mg/m(2)/day. DLTs included diarrhea, nausea/vomiting, fatigue, and hyperbilirubinemia. The maximum tolerated dose was 50 mg/m(2)/day. Pharmacokinetic data are consistent with potent modulation of 5 fluorouracil (5-FU) by CDHP, with prolonged half-life and 5-FU AUC at least 10 fold higher than reported in previous studies of equitoxic doses of tegafur modulated by uracil. Pharmacodynamic analysis demonstrated a correlation between diarrhea grade and both 5-FU C(max) (r = 0.57, P < 0.05) and 5-FU area under the curve (r = 0.74, P < 0.01). CONCLUSIONS: The recommended Phase II dose of S-1 administered once daily for 21 consecutive days of 28 is 50 mg/m(2). The pharmacokinetic data presented provide evidence of 5-FU modulation by CDHP. Pharmacodynamic analyses suggest that the utility of pharmacology-based dosing of S-1 should be explored in future trials. Evaluation of once-daily dosing of S-1 in malignancies for which fluoropyrimidines have known antitumor activity is warranted. PMID- 12114412 TI - Phase I trial and correlative laboratory studies of bryostatin 1 (NSC 339555) and high-dose 1-B-D-arabinofuranosylcytosine in patients with refractory acute leukemia. AB - A Phase I trial has been conducted in patients with refractory/relapsed acute leukemia in which escalating doses of the protein kinase C (PKC) activator and down-regulator bryostatin 1 (NSC399555), administered as a 24-h continuous infusion on days 1 and 11, were given immediately before and after a split course of high-dose 1-beta-D-arabinofuranosylcytosine (HiDAC; 1.5 g/m(2) every 12 h x 4) administered on days 2 and 3, and 9 and 10. The bryostatin 1 maximally tolerated dose (MTD) was identified as 50 microg/m(2), with myalgias representing the major dose-limiting toxicity (DLT). Other DLTs included prolonged neutropenia and thrombocytopenia, and hepatotoxicity. Of the 23 patients who completed their course of therapy and were fully evaluable for response, the large majority of whom had unfavorable prognostic characteristics, 4 complete remissions (CRs) were obtained. An additional 3 patients were treated at a 3 g/m(2) ara-C (1-beta-D arabinofuranosylcytosine) dose level to determine whether this HiDAC dose could be administered in conjunction with bryostatin 1. All 3 of these patients experienced DLT, and this dose was considered above the MTD. However, one of the latter patients, who was heavily pretreated, also achieved a CR that persisted 5+ months without maintenance. Finally, 1 patient post-HiDAC and autologous bone marrow transplantation achieved a 5+ month leukemia-free survival although she did not meet the criteria for a CR because of persistent transfusion requirements. Correlative laboratory studies performed on blasts from 9 patients revealed that in vivo administration of bryostatin 1 resulted in variable effects on total blast PKC activity, including decreases in 4 samples, increases in 2, and no change in 3. Previous in vivo bryostatin 1 exposure also exerted disparate effects on the extent of apoptosis observed in blasts exposed to ara-C ex vivo, although increases were noted in a subset of patient samples. Interestingly, in vivo administration of bryostatin 1 by itself induced lethality in some patient specimens. No clear relationship between the in vivo effects of bryostatin 1 on blast PKC activity and the extent of ara-C-related apoptosis that occurred ex vivo was apparent. Together, these findings demonstrate that bryostatin 1 can be safely administered as a continuous infusion before and after a split course of HiDAC in patients with refractory leukemia, and identify the bryostatin 1 MTD as 50 microg/m(2) when given by this schedule. Furthermore, the achievement of several CRs in the setting of a Phase I trial in which many patients had particularly high-risk features (e.g., short initial remission, previous HiDAC or autologous bone marrow transplantation, and multiple previous salvage regimens) suggests that this regimen has activity in acute leukemia and warrants additional investigation. PMID- 12114413 TI - Phase I and pharmacokinetic study of DX-8951f (exatecan mesylate), a hexacyclic camptothecin, on a daily-times-five schedule in patients with advanced leukemia. AB - PURPOSE: DX-8951f is a novel hexacyclic camptothecin-analogue topoisomerase I inhibitor with both in vitro antileukemic activity and myelosuppression as a dose limiting toxicity in solid tumor Phase I studies. DX-8951f is active in a human acute myeloid leukemia (AML) severe combined immunodeficient mouse model. In a leukemia Phase I study, we investigated the toxicity profile and pharmacokinetics of DX-8951f in patients with primary refractory or relapsed AML or acute lymphocytic leukemia, myelodysplastic syndromes, or chronic myelogenous leukemia in blastic phase (CML-BP). EXPERIMENTAL DESIGN: DX-8951f was given as an i.v. infusion over 30 min daily for 5 or 7 days. The starting dose was 0.6 mg/m(2)/day for 5 days (3.0 mg/m(2)/course). Courses were given every 3-4 weeks according to toxicity and antileukemic efficacy. RESULTS: Twenty-five patients (AML, 21 patients; myelodysplastic syndrome, 1 patient; acute lymphocytic leukemia, 2 patients; CML-BP, 1 patient) were treated. Stomatitis was the dose-limiting toxicity, occurring in two of two patients treated at 1.35 mg/m(2)/day for 5 days, two of three treated at 1.2 mg/m(2)/day for 5 days, and one of six treated at 0.9 mg/m(2)/day for 7 days. The recommended Phase II dose was 0.9 mg/m(2)/day for 5 days. The pharmacokinetics of DX-8951 was linear and well fit by a two compartment model. CONCLUSIONS: Phase II studies are warranted to further define the activity of DX-8951f in patients with hematological malignancies. PMID- 12114414 TI - A phase I study of pivaloyloxymethyl butyrate, a prodrug of the differentiating agent butyric acid, in patients with advanced solid malignancies. AB - Pivaloyloxymethyl butyrate (AN-9), an acyloxyalkyl ester prodrug of butyric acid (BA), has demonstrated greater potency than BA at inducing malignant cell differentiation and tumor growth inhibition and has demonstrated more favorable toxicological, pharmacological, and pharmaceutical properties than BA in preclinical studies. The principal objective of this study was to determine the feasibility of administering AN-9 as a 6-h i.v. infusion daily for 5 days every 3 weeks in patients with advanced solid malignancies. The study also sought to determine the principal toxicities and maximum tolerated dose of AN-9 on this intermittent schedule, as well as the effects of AN-9 on fetal hemoglobin production, a parameter indicative of RBC differentiation. None of the 28 patients treated with 85 total courses of AN-9 at dosages ranging from 0.047 to 3.3 g/m(2)/day every 3 weeks experienced dose limiting toxicity. Mild to moderate nausea, vomiting, hepatic transaminase elevation, hyperglycemia, fever, fatigue, anorexia, injection site reaction, diarrhea, and visual complaints were observed. Dose escalation of AN-9 was limited by the maximum feasible volume of its intralipid formulation vehicle that could be administered safely on this schedule, resulting in a maximum deliverable dose of 3.3 g/m(2)/day. There was no consistent increase in fetal hemoglobin with AN-9 treatment. A partial response was observed in a previously untreated patient with metastatic non-small cell lung cancer. Additional disease-directed clinical evaluations of AN-9 are necessary to establish the breadth of its antitumor activity and to assess its role as an effective differentiating agent. PMID- 12114415 TI - A phase I pharmacologic and pharmacodynamic study of pyrazoloacridine given as a weekly 24-hour continuous intravenous infusion in adult cancer patients. AB - PURPOSE: Pyrazoloacridine (PZA) is an investigational nucleic acid binding agent that inhibits the activity of topoisomerases I and II through a mechanism distinct from other topoisomerase poisons. PZA shows schedule-independent cytotoxicity against tumor cells, whereas host toxicity is greater with shorter infusions. We assessed the clinical toxicities and pharmacologic effects of PZA given as a 24-h i.v. infusion weekly for 3 of 4 weeks. EXPERIMENTAL DESIGN: Thirty-two adult patients with solid tumors received PZA at five dose levels (100 351 mg/m(2)). Plasma samples were obtained at the end of the PZA infusion at all of the dose levels, with extended sampling in a cohort treated at the recommended dose. RESULTS: Dose-limiting granulocytopenia and mucositis occurred in 2 of 6 patients at 351 mg/m(2), but lower doses were well tolerated. No responses were seen, but 28% had stable disease for > or =3 months. Plasma levels strongly correlated with the degree of granulocytopenia. Extended pharmacokinetics in 7 patients treated with 281 mg/m(2) indicated the following averages: maximum plasma level, 1.6 microM; area under the plasma concentration-time curve, 56 microM.h; terminal half-life, 27 h; urinary recovery, 17% over 72 h. DNA fragmentation in post-PZA bone marrow mononuclear cells was seen in 9 of 28 samples (all at > or =281 mg/m(2)). CONCLUSIONS: Unlike other schedules of PZA, neurotoxicity and thrombocytopenia were not problematic with a weekly 24-h infusion of PZA. The recommended Phase II dose is 281 mg/m(2), which was well tolerated. Both end of infusion plasma levels and presence of DNA damage correlated with granulocyte toxicity. PMID- 12114416 TI - A phase I and pharmacokinetic study of SAM486A, a novel polyamine biosynthesis inhibitor, administered on a daily-times-five every-three-week schedule in patients with Advanced solid malignancies. AB - PURPOSE: SAM486A is a novel inhibitor of the polyamine biosynthetic enzyme S adenosylmethionine decarboxylase (SAMDC). This study was performed to characterize the toxicity profile and the pharmacological behavior and to determine the maximum tolerated dose (MTD) of SAM486A administered by a 1-h i.v. infusion daily for 5 days every 3 weeks in patients with advanced cancer. EXPERIMENTAL DESIGN: Twenty-three patients received 46 cycles of SAM486A at dose levels ranging from 3.6 to 202.8 mg/m(2)/day. SAM486A plasma concentrations were measured during the first cycle for pharmacokinetic and pharmacodynamic evaluations. Paired tumor biopsy specimens pre- and posttreatment were obtained in 1 patient to assess the impact of SAM486A on intratumoral enzymes and metabolites involved in the polyamine biosynthetic pathway. RESULTS: The dose limiting toxicity of SAM486A on this schedule was myelosuppression. Nonhematological toxicities, including nausea, vomiting, anorexia, and fatigue, were mild to moderate in severity. The MTD of SAM486A was 102.4 mg/m(2)/day. Pharmacokinetic analyses demonstrated a rapid initial decrease in plasma drug concentrations at the end of infusion, followed by a long terminal elimination phase with a mean (+/- SD) terminal elimination half-life of 65.4 +/- 55.6 h. Dose and area under the concentration-time curve correlated with the appearance of grade 4 neutropenia with correlation coefficients of 0.70 and 0.69, respectively. Analysis of paired tumor biopsy specimens taken before and after SAM486A treatment in 1 patient with metastatic melanoma revealed decreased SAMDC activity, increased ornithine decarboxylase activity, increased levels of putrescine, and depleted levels of decarboxylated S-adenosylmethionine and spermine, all of which are consistent with the proposed mode of action of SAM486A. CONCLUSIONS: SAM486A was well tolerated on this schedule of administration with the MTD established at 102.4 mg/m(2)/day. Neutropenia was dose-limiting and correlated with dose and area under the concentration-time curve. Pharmacodynamic assessment of tumoral tissues in 1 study patient demonstrated changes in the levels of polyamines and their biosynthetic enzymes consistent with SAMDC inhibition. PMID- 12114417 TI - Treatment of philadelphia chromosome-positive, accelerated-phase chronic myelogenous leukemia with imatinib mesylate. AB - PURPOSE: Imatinib mesylate, a specific Bcr-Abl tyrosine kinase inhibitor, has shown encouraging activity in chronic myelogenous leukemia (CML). EXPERIMENTAL DESIGN: We treated 237 patients (median age, 50 years; age range, 18-82 years) with Philadelphia chromosome (Ph)-positive accelerated-phase CML with oral imatinib mesylate at daily doses of 400 mg (26 patients) or 600 mg (211 patients) and evaluated response and survival characteristics in univariate and multivariate analyses. RESULTS: Among the 200 patients with accelerated-phase CML for whom follow-up was 3 months or more, rates of complete and partial hematological response were 80% and 10%. Cytogenetic responses were evident in 90 patients [45%; complete response in 47 patients (24%) and partial response (Ph 1 34%) in 21 patients (11%)]. The estimated 18-month survival rate was 73%. The estimated complete hematological response rate at 18 months was 68%; that for cytogenetic response was 82%. In multivariate analyses, a diagnosis-to-treatment interval of 3 years or more, splenomegaly, and peripheral blasts predicted poor major cytogenetic response; age >60 years, marrow basophilia, and clonal evolution predicted poor survival. The 600-mg drug dose was associated with better cytogenetic response and survival in univariate analysis (P < 0.01) but not in multivariate analysis. Landmark analysis showed that achieving a cytogenetic response at 3 months or a major cytogenetic response (Ph < 35%) at 6 months was associated with better long-term survival. Seven of 15 patients who were in a second chronic phase achieved major cytogenetic response. The incidence of severe nonhematological toxic effects was 23%; drug discontinuation for severe toxicity was needed in 3% of patients. CONCLUSIONS: Imatinib mesylate was active against Ph-positive, accelerated-phase CML, and the prognostic factors identified in this study could aid in tailoring treatment strategies to specific risk groups. PMID- 12114418 TI - Imatinib mesylate for Philadelphia chromosome-positive, chronic-phase myeloid leukemia after failure of interferon-alpha: follow-up results. AB - We treated 261 patients with Philadelphia chromosome (Ph)-positive chronic myeloid leukemia (CML) in chronic phase after failure of IFN-alpha with the Bcr Abl tyrosine kinase inhibitor imatinib mesylate (400 mg/day given p.o.) and analyzed hematological and cytogenetic responses, long-term prognosis, factors associated with achievement of major cytogenetic response and survival, and comparative survival in similar patients treated with other regimens. Median patient age was 55 years; 34% were 60 years or older, and median chronic-phase duration was 33 months. Overall, 94% achieved a complete hematological response, and 71% had a cytogenetic response [major (Ph+ cells <35%) in 62% and complete in 45%]. At a median follow-up of 17 months, 241 patients (92%) were still taking imatinib mesylate; estimated 18-month freedom from progression and survival rates were 93 and 96%. Multivariate analysis of factors associated with major cytogenetic response identified long chronic phase, marrow basophilia, high percentage of Ph+ cells before therapy, and prior hematological resistance to IFN alpha as being adverse factors. This model was used to generate good-, intermediate- and poor-risk subgroups who had estimated major cytogenetic response rates of 93, 53, and 34%, respectively. Univariate analysis in terms of survival identified leukocytosis, high percentages of peripheral and marrow blasts, marrow basophilia, and the presence of cytogenetic clonal evolution as being adverse factors. Achieving a cytogenetic response at 3 or 6 months of therapy was associated with prolonged survival. In a subset analysis, survival rates among 161 patients with Ph-positive CML after hematological or cytogenetic failure after IFN-alpha who had been treated with imatinib mesylate were better than those for similar patients treated previously with other regimens. In summary, imatinib mesylate is highly effective in chronic-phase CML after IFN alpha failure. We identified pretreatment and treatment-associated factors that were associated with higher major cytogenetic response rates and with improved survival. PMID- 12114419 TI - Phase II randomized study of ISIS 3521 and ISIS 5132 in patients with locally advanced or metastatic colorectal cancer: a National Cancer Institute of Canada clinical trials group study. AB - BACKGROUND: Because treatment of metastatic colon cancer is noncurative, new treatments are needed. This trial evaluated the antitumor effects of two targeted anticancer agents: (a) ISIS 3521, an antisense inhibitor of the protein kinase C alpha; and (b) ISIS 5132, an antisense inhibitor of c-raf kinase in patients untreated previously with recurrent or metastatic colorectal carcinoma. PATIENTS AND METHODS: All patients had colorectal adenocarcinoma with measurable disease and no prior chemotherapy for metastatic disease. Patients were randomized to receive either ISIS 3521 or ISIS 5132 at a dose of 2 mg/kg/day as a continuous i.v. infusion 21 of 28 days. Cycles were repeated as long as progression was not seen, and doses of both agents were modified according to toxic effects. A two arm study design was used with each study arm considered independently. Steady state blood levels of both antisense molecules were measured on days 8, 15, and 22 of the first cycle of therapy. RESULTS: Thirty-seven eligible patients were enrolled, and 32 were evaluable for response (17 receiving ISIS 3521 and 15 receiving ISIS 5132). No responses were noted. Four of the patients receiving ISIS 3521 had stable disease, and 5 patients receiving ISIS 5132 were stable. CONCLUSION: Neither ISIS 5132 nor ISIS 3521given in the dose and schedule studied induced objective responses in untreated colorectal cancer patients. PMID- 12114421 TI - Altered irinotecan pharmacokinetics in pediatric high-grade glioma patients receiving enzyme-inducing anticonvulsant therapy. AB - PURPOSE: The purpose of this study was to determine the effect of enzyme-inducing anticonvulsants (EIAs) on the disposition of irinotecan and metabolites in pediatric patients with high-grade glioma. EXPERIMENTAL DESIGN: Pediatric patients with newly diagnosed high-grade glioma were enrolled on this study between March 1999 and February 2001. During course 1, irinotecan was administered as a 60-min i.v. infusion at a dosage of 20 mg/m(2)/day for 5 days of 2 consecutive weeks. On days 1 and 12 of course 1, we collected serial plasma samples to measure the concentrations of the lactone and total forms of irinotecan and its metabolites SN-38 (7-ethyl-10-hydroxycamptothecin), SN-38 glucuronide (7-ethyl-10-[3,4,5-trihydroxy-pyran-2-carboxylic acid]camptothecin), and 7-ethyl-10-[4-N-(5-aminopentanoic acid)-1-piperidino]carbonyloxycamptothecin. RESULTS: Thirty-one patients were enrolled. In patients receiving EIAs, the area under the concentration versus time curve (AUC) of irinotecan lactone and SN-38 lactone was significantly lower (P = 0.01 and P = 0.002, respectively), and the irinotecan lactone clearance was significantly higher (P = 0.0003), as compared with those in patients who received no EIAs. The glucuronidation ratio was higher (P = 0.0009), and the ratio of SN-38 AUC to irinotecan AUC was lower (P = 0.02) in patients who received EIAs. Two patients receiving EIAs tolerated increased irinotecan dosages of 30 and 40 mg/m(2)/day without toxicity. One patient receiving EIAs experienced grade 3 diarrhea when the dosage of irinotecan was increased to 60 mg/m(2)/day. CONCLUSIONS: EIAs increase the clearance of irinotecan and cause a decrease in systemic exposure to the active metabolite SN 38. Patients who are receiving irinotecan and who require anticonvulsants should be placed on non-EIA therapy, when possible. PMID- 12114420 TI - Phase I clinical and pharmacokinetic study of NSC 655649, a rebeccamycin analogue, given in both single-dose and multiple-dose formats. AB - PURPOSE: NSC 655649 was given in both single- and multiple-dose formats, to characterize maximum tolerated dose (MTD), toxicity, and pharmacokinetic profile. EXPERIMENTAL DESIGN: Patients with advanced malignancies were treated with escalating doses of NSC 655649 in either a single-dose format (step 1) or a multiple-dose format (step 2). In step 1, NSC 655649 was given as a 30-60 min infusion. In step 2, the NSC 655649 dose was divided into three consecutive daily doses. Plasma and urine were sampled to assess the pharmacokinetic and excretory characteristics of NSC 655649. A total of 12 patients were enrolled at the MTD for the purpose of gender equity. RESULTS: Forty-three patients were treated with NSC 655649 for a total of 108 cycles in step 1, and 26 patients were treated for a total of 41 cycles in step 2. The MTD for both steps 1 and 2 was determined to be 572 mg/m(2). Myelosuppression was the dose-limiting toxicity. Local venous irritation was generally grade 1-2 in severity but could only be adequately prevented by administration of study drug through central i.v. access. One patient with adenocarcinoma of unknown primary experienced a partial response on step 1. Four patients experienced stable disease of >100 days duration. CONCLUSIONS: NSC 655649 may be safely given at an MTD of 572 mg/m(2) in both single-dose and multiple-dose formats. Optimally, this drug should be administered through central i.v. access. PMID- 12114422 TI - Bone marrow angiogenesis in 400 patients with monoclonal gammopathy of undetermined significance, multiple myeloma, and primary amyloidosis. AB - PURPOSE: To determine whether bone marrow (BM) angiogenesis progressively increases along the spectrum of plasma cell disorders ranging from monoclonal gammopathy of undetermined significance (MGUS) to advanced myeloma. EXPERIMENTAL DESIGN: Four hundred patients with the following disorders were studied: MGUS (76 patients); smoldering (indolent; early-stage) multiple myeloma (SMM; 112 patients); newly diagnosed, active multiple myeloma (MM; 99 patients); relapsed (advanced) multiple myeloma (RMM; 26 patients); and primary amyloidosis (AL; 87 patients). Forty-two normal control BM samples were studied for comparison. BM angiogenesis was studied in a blinded manner by immunohistochemical staining for CD34 to identify microvessels. RESULTS: The median (range) microvessel density (MVD) per x400 high power field was 1.3 (0-11) in the controls, 1.7 (0-10) in AL, 3 (0-23) in MGUS, 4 (1-30) in SMM, 11 (1-48) in newly diagnosed MM, and 20 (6-47) in RMM; P < 0.001. MVD was significantly higher in MGUS, SMM, newly diagnosed MM, and RMM compared with controls and AL; P < 0.001. MVD was not significantly different between controls and AL. By grading, high-grade angiogenesis was present in 0% of controls and AL, 1% of MGUS, 3% of SMM, 29% of newly diagnosed MM, and 42% of RMM; P < 0.001. MVD correlated with the BM plasma cell labeling index (rho = 0.46, P < 0.001) and BM plasma cell percentage (rho 0.5, P < 0.001). Survival was 28 months in SMM and newly diagnosed MM with high-grade angiogenesis, compared with 53 months for those with low- and intermediate-grade angiogenesis; P = 0.02. CONCLUSIONS: BM angiogenesis progressively increases along the spectrum of plasma cell disorders, from the more benign MGUS stage to advanced myeloma, indicating that angiogenesis may be related to disease progression. PMID- 12114423 TI - DNA methylation of multiple promoter-associated CpG islands in adult acute lymphocytic leukemia. AB - PURPOSE: Aberrant methylation of promoter-associated CpG islands is an epigenetic oncogenic mechanism. The objective of this study was to define the methylation characteristics of patients with acute lymphocytic leukemia (ALL). EXPERIMENTAL DESIGN: Using bisulfite-PCR followed by restriction enzyme digestion (COBRA), we have analyzed the methylation status of 10 promoter-associated CpG islands in 80 untreated adult patients with ALL. RESULTS: Mean methylation density of MDR1, THBS2, MYF3, ER, p15, THBS1, CD10, C-ABL, and p16 was 24.5%, 20.8%, 17.6%, 16.1%, 11.3%, 8.9%, 4.5%, 3.7%, and 1.3% respectively. p73 was methylated in 17 of 80 cases (21.2%). A total of 86.2% of the cases had methylation of at least one gene, and 42.5% of the cases had methylation of three or more genes. MDR1 methylation was inversely correlated with age (P = 0.01). CD10 methylation inversely correlated with CD10 expression (P = 0.0001). Methylation of MDR1 and THBS1 was inversely associated with the presence of the Philadelphia chromosome, whereas C-ABL methylation correlated with the presence of the p210 variant of the Philadelphia chromosome. In univariate analysis, methylation of THBS1 was associated with a favorable outcome (P = 0.02), whereas methylation of p73, p15, and C-ABL was associated with a trend toward worse prognosis. CONCLUSIONS: Aberrant DNA methylation of promoter-associated CpG islands is very common in adult ALL and potentially defines subgroups with distinct clinical and biological characteristics. PMID- 12114424 TI - Suppression of rho B expression in invasive carcinoma from head and neck cancer patients. AB - PURPOSE: In contrast to Ras small GTPases, which contribute to human malignancy when overexpressed or constitutively activated, convincing evidence for the involvement of Ras homologous (Rho) GTPases in human cancer is still missing. In cell culture and animal models, RhoB antagonizes malignant transformation, but no data are available regarding the expression of RhoB in human tumors. In this study, we have analyzed the status of the RhoB protein and the closely related family member RhoA in human head and neck squamous cell carcinomas. EXPERIMENTAL DESIGN: Protein immunoexpression was quantitated by image analysis in the context of tumor invasion and differentiation. To account for possible individual variations, expression levels of RhoB and RhoA were evaluated in the tumor and its adjacent nonneoplastic tissue. Potential gene deletions or mutations were assessed by PCR and RT-PCR. RESULTS: RhoB expression is readily detected in normal epithelium, carcinomas in situ, and well-differentiated tumors, but it becomes weak to undetectable as tumors become deeply invasive and poorly differentiated. In contrast, Ki67 (proliferation marker) and RhoA protein levels increase with tumor progression. Furthermore, whereas in nonneoplastic keratinocytes RhoB is localized mainly in the nucleus, in carcinomas RhoB is predominantly located in the cytoplasm. RhoB gene deletions or mutations were not found. CONCLUSIONS: These results give additional support to the notion that RhoB may play a tumor suppressive role in squamous cell carcinomas of the head and neck. The lack of RhoB expression in deeply invasive carcinoma argues against inhibition of RhoB farnesylation as a mediator of farnesyltransferase inhibitors' antitumor activity. PMID- 12114425 TI - CA125 response is associated with estrogen receptor expression in a phase II trial of letrozole in ovarian cancer: identification of an endocrine-sensitive subgroup. AB - PURPOSE: This study was an open-label Phase II trial of the aromatase inhibitor letrozole (Femara) in patients with relapsed ovarian cancer with evaluation of possible biological markers for response. EXPERIMENTAL DESIGN: 60 patients were treated with letrozole (2.5 mg daily) at the time of CA125 relapse. Disease response was assessed by Union International Contre Cancer (UICC) criteria and by CA125 measurement. Estrogen receptor (ER), progesterone receptor, epidermal growth factor receptor, erbB2, and HSP27 were measured by immunohistochemistry in paraffin-fixed material obtained from the primary tumors at initial surgery. RESULTS: 50 patients were evaluable by UICC criteria, and although no complete or partial responses were obtained, 10 patients had stable disease on scan for at least 12 weeks. CA125 responses were evaluable in 54 patients. A partial marker response (>50% decrease) was seen in 5, and the marker remained stable in an additional 14 patients (25% increase). Tumors from the UICC stable disease group had significantly higher ER (P = 0.027) and progesterone receptor (P = 0.0066) values than the progressive disease group, and a combination of these was strongly associated with stable disease (P < 0.0001). Using CA125 criteria, comparison of the CA125 stable/responding disease with progressive disease indicated that tumors with higher ER (P = 0.013), lower erbB2 (P = 0.026), and higher epidermal growth factor receptor (P = 0.009) were associated with CA125 stable/responsive disease. CONCLUSIONS: These results imply that letrozole treatment can produce disease stabilization and CA125 responses that in turn are linked to higher levels of ER expression. These data suggest the presence of an endocrine-sensitive group that could be targeted in future studies. PMID- 12114426 TI - The expression of carbonic anhydrase II in hematological malignancies. AB - PURPOSE: Carbonic anhydrases (CAs) are key enzymes that regulate acid-base homeostasis in both normal and pathological conditions. Recent studies have shown that they are functionally involved in the growth and invasion of cancer cells. However, there are only a few publications on CAs in hematological malignancies. EXPERIMENTAL DESIGN: Here we investigated the presence of CA isozymes in six malignant hematopoietic cell lines and malignant blast cells of bone marrow samples collected from patients with acute myeloid leukemia, acute lymphoblastic leukemia, or chronic myelomonocytic leukemia. RESULTS: Because three of the malignant hematopoietic cell lines expressed CA II, we also set out to examine its expression in a series of bone marrow samples. Positive reactions were found in 16 of 26 cases (62%) of acute myeloid leukemia, 11 of 15 cases (73%) of acute lymphoblastic leukemia, and 1 of 2 cases (50%) of chronic myelomonocytic leukemia. CONCLUSIONS: The results indicate that CA II expression is not restricted to only one cell lineage but may result from a genetic aberration that occurs in both myeloid and lymphatic lineages or in their progenitor cell. Because CA II is expressed in most patients with leukemic blast cells, CA inhibitors may prove to be of value as an adjunct to chemotherapy for such cancers. PMID- 12114427 TI - Methylation microarray analysis of late-stage ovarian carcinomas distinguishes progression-free survival in patients and identifies candidate epigenetic markers. AB - PURPOSE: The purpose of this study was to profile methylation alterations of CpG islands in ovarian tumors and to identify candidate markers for diagnosis and prognosis of the disease. EXPERIMENTAL DESIGN: A global analysis of DNA methylation using a novel microarray approach called differential methylation hybridization was performed on 19 patients with stage III and IV ovarian carcinomas. RESULTS: Hierarchical clustering identified two groups of patients with distinct methylation profiles. Tumors from group 1 contained high levels of concurrent methylation, whereas group 2 tumors had lower tumor methylation levels. The duration of progression-free survival after chemotherapy was significantly shorter for patients in group 1 compared with group 2 (P < 0.001). Differential methylation in tumors was independently confirmed by methylation specific PCR. CONCLUSIONS: The data suggest that a higher degree of CpG island methylation is associated with early disease recurrence after chemotherapy. The differential methylation hybridization assay also identified a select group of CpG island loci that are potentially useful as epigenetic markers for predicting treatment outcome in ovarian cancer patients. PMID- 12114428 TI - Vascular endothelial growth factor splice variants and their prognostic value in breast and ovarian cancer. AB - PURPOSE: Vascular endothelial growth factor (VEGF) is a promotor for tumor angiogenesis, and is known to be elevated in breast and ovarian cancers. Through alternative splicing six VEGF isoforms were identified. We studied VEGF isoform expression in breast and ovarian cancer cell lines, as well as in breast carcinomas and ovarian tumors, and correlated the expression pattern with the in vitro invasiveness of the breast carcinoma cell lines and the clinicopathologic characteristics of the tumors. EXPERIMENTAL DESIGN: Reverse transcription-PCR and automated laser fluorescence fragment analysis were used to determine the expression of each splice variant. This method allowed the detection of all of the splice variants simultaneously, especially VEGF145 for the first time in tumor tissue. RESULTS: VEGF121 and VEGF165 were the most dominantly expressed variants in all of the tumor samples and cell lines investigated. VEGF145 was very weakly or not expressed in breast and ovarian cancers. Statistical analysis showed no correlation between VEGF splice variant expression in the tumors and histological type, differentiation grade, tumor size, Federation Internationale des Gynaecologistes et Obstetristes, and nodal status from cancer patients. There was also no correlation between the invasive capacity of breast cell lines and VEGF isoform expression. CONCLUSIONS: Even though expression levels of VEGF have been shown to be important for tumor invasion and progression, the present data indicate no relation of VEGF isoform pattern with invasion and progression. PMID- 12114429 TI - Mitochondrial C-tract alteration in premalignant lesions of the head and neck: a marker for progression and clonal proliferation. AB - PURPOSE: Although mitochondrial DNA mutations have been described recently in many different tumor types, the nature and timing of such alterations remain unclear. In an effort to further examine the role of mitochondrial DNA mutations in carcinogenesis, we examined 137 premalignant lesions of the head and neck from 93 patients for DNA alterations in the poly-cytosine tract (C-tract) of the displacement loop, discovered recently to be a hot spot of mitochondrial DNA alteration. EXPERIMENTAL DESING: All premalignant lesions were tested using a length-based PCR assay, which amplified the C-tract region of mitochondrial DNA. Somatic microsatellites at six loci were also tested on a subset of patients with metachronous or synchronous lesions found to possess a mitochondrial C-tract alteration. RESULTS: Thirty-four of 93 (37%) patients harbored lesions that displayed a C-tract alteration. There was a clear increase in incidence from histologically benign hyperplasia (22%) to squamous carcinoma in situ (62%: P < 0.01). We also tested synchronous dysplastic lesions, metachronous dysplastic lesions, and normal epithelium adjacent to dysplastic epithelium with this assay. In most cases, the mitochondrial C-tract status identified a clonal relationship between these lesions. Genomic microsatellites also confirmed that a clonal relationship was present in many of these cases. CONCLUSIONS: Mitochondrial DNA alterations in the head and neck occur in the earliest premalignant lesions and demonstrate a rising incidence that parallels histological severity. These alterations are valuable as additional markers of histopathological progression. PMID- 12114430 TI - Clinicopathological significance of loss of heterozygosity on chromosome 13q in hepatocellular carcinoma. AB - PURPOSE: Allelic loss is the most frequently genetic alteration found in hepatocellular carcinoma (HCC). Previous genome-wide studies have indicated that chromosome 13q is one of the most frequently affected chromosomes. However, reports on detailed deletion mapping as well as detailed clinicopathological correlation are scanty. EXPERIMENTAL DESIGN: We performed high-density allelotyping on chromosome 13q in HCC from 60 patients and investigated the correlation between allelic losses on chromosome 13q and the clinicopathological features. RESULTS: Allelic loss at one or more of the 29 microsatellite markers was found in 28 (47%) of the 60 HCCs. Allelic losses were more frequently found in tumors with larger size or in tumors at more advanced tumor stages (P = 0.015 and 0.012, respectively). These two clinicopathological features were also significantly associated with the accumulation of allelic losses in terms of fractional allelic loss index (P = 0.028 and 0.018, respectively). In addition, subchromosomal regions located at 13q12.3-14.1 and 13q32 were found to be significantly associated with advanced tumor stages and larger tumor size, respectively (P = 0.008 and 0.007). CONCLUSIONS: The overall findings suggested that allelic losses on 13q might play an important role in contributing to a more aggressive tumor behavior. Putative tumor suppressor genes might be harbored at 13q12.3-14.1 and 13q32, and inactivation of these genes via allelic losses might enhance tumor progression in HCC. PMID- 12114431 TI - Overexpression of epidermal growth factor and hepatocyte growth factor receptors in a proportion of gastrinomas correlates with aggressive growth and lower curability. AB - PURPOSE: Growth factor receptor expression and activation, particularly for epidermal growth factor (EGF) and hepatocyte growth factor (HGF), in many endocrine and nonendocrine tumors is important in determining tumor recurrence, growth, and aggressiveness. Whether this is true of neuroendocrine tumors such as gastrinomas is unclear. EXPERIMENTAL DESIGN: To address this question, we analyzed the extent of EGFR and HGFR expression in gastrinomas from 38 patients with Zollinger-Ellison syndrome and correlated it with clinical and tumor characteristics. EGFR (n = 38) and HGFR (n = 22) mRNA levels were determined by competitive PCR, and immunohistochemistry was performed on a subset. RESULTS: In each of the gastrinomas studied, detectable levels of EGFR and HGFR mRNA were present. Low levels of EGFR protein expression were detected in 40% of gastrinomas and HGFR protein expression in 90%. EGFR mRNA expression varied by 1050-fold and HGFR by 375-fold. Eighteen percent of gastrinomas overexpressed EGFR mRNA and 14% overexpressed HGFR mRNA, compared with normal pancreas. Maximal EGFR and HGFR mRNA levels were 4- and 1.2-fold increased and correlated with the presence of liver metastases (P = 0.034) and decreased long-term curability (P = 0.027) but not tumor location, size, or tumor functional characteristics. CONCLUSIONS: These above results indicate that EGFR and HGFR mRNA are universally expressed in gastrinomas. Furthermore, each is overexpressed in a minority (15 20%) of the gastrinomas, and the overexpression correlates with aggressive growth and lower curability. PMID- 12114432 TI - Low ERCC1 expression correlates with prolonged survival after cisplatin plus gemcitabine chemotherapy in non-small cell lung cancer. AB - PURPOSE: Overexpression of the excision repair cross-complementing 1 (ERCC1) gene, which is crucial in the repair of cisplatin (CDDP)-DNA adducts, is reported to negatively influence the effectiveness of CDDP-based therapy for gastric and ovarian cancers. Recent evidence indicates that Gemcitabine (Gem) may modulate ERCC1 nucleotide excision repair activity, and down-regulation of DNA repair activity by ERCC1 antisense RNA reportedly inhibits synergism of CDDP/Gem. We investigated whether ERCC1 mRNA expression levels were associated with clinical outcomes after treatment with a combination Gem/CDDP regimen for patients with advanced stage non-small cell lung cancer (NSCLC). EXPERIMENTAL DESIGN: Response and survival were correlated with the level of ERCC1 expression in 56 patients with advanced (stage IIIb or IV) NSCLC treated as part of a multicenter randomized trial with Gem 1250 mg/m(2) days 1 and 8 plus CDDP 100 mg/m(2) on day 1 every 3 weeks. mRNA was isolated from paraffin-embedded pretreatment primary tumor specimens, and relative expression levels of ERCC1/beta-actin were measured using a quantitative reverse transcription-PCR (Taqman) system. RESULTS: ERCC1 expression was detectable in all tumors. There were no significant differences in ERCC1 levels by gender, age, performance status, weight loss, or tumor stage. The overall response rate was 44.7%. There were no significant associations between ERCC1 expression and response. Median overall survival was significantly longer in patients with low ERCC1 expression tumors (61.6 weeks; 95% confidence interval, 42.4-80.7 weeks) compared to patients with high expression tumors (20.4 weeks, 95% confidence interval, 6.9-33.9 weeks). ERCC1 expression, Eastern Cooperative Oncology Group performance status, and presence of weight loss were significant prognostic factors for survival in a Cox proportional hazards multivariable analysis. CONCLUSIONS: These data suggest that ERCC1 expression is a predictive factor for survival after CDDP/Gem therapy in advanced NSCLC. Although there was a trend toward decreased response with high ERCC1 mRNA levels, this difference failed to reach statistical significance. This result may reflect the impact of Gem and the requirement for ERCC1 expression for CDDP/Gem synergism or may be attributable to the relatively small patient sample size in this study. Prospective studies of ERCC1 as a predictive marker for activity of CDDP-based regimens in NSCLC are warranted. PMID- 12114434 TI - Proteomic analysis of lung adenocarcinoma: identification of a highly expressed set of proteins in tumors. AB - PURPOSE: The goal of this study was to identify potential protein markers in lung adenocarcinomas. EXPERIMENTAL DESIGN: A series of 93 lung adenocarcinomas (64 stage I and 29 stage III) and 10 uninvolved lung samples were examined for quantitative differences in protein expression using two-dimensional PAGE. Candidate proteins were identified using matrix-assisted laser desorption/ionization mass spectrometry or peptide sequencing. The levels of the individual isoforms of nine proteins found to be overexpressed in the lung tumors were examined. Potential mechanisms for overexpression were examined by comparing mRNA expression levels, assessed using oligonucleotide arrays, to the protein values in the same samples. RESULTS: Antioxidant enzyme AOE372, ATP synthase subunit d (ATP5D), beta1,4-galactosyltransferase, cytosolic inorganic pyrophosphatase, glucose-regulated M(r) 58,000 protein, glutathione-S-transferase M4, prolyl 4-hydroxylase beta subunit, triosephosphate isomerase, and ubiquitin thiolesterase (UCHL1) were identified as being significantly overexpressed in lung adenocarcinomas. The expression of these proteins was increased from 1.4- to 10.6-fold as compared with uninvolved lung tissue. The expression of the individual protein isoforms was correlated with 10 clinicopathological variables as well as with each gene's mRNA level in the same sample. Both isoforms of glucose-regulated M(r) 58,000 protein were found to be significantly correlated with their mRNA expression profiles (P < 0.05), indicating that increased transcription likely underlies the increased expression of these proteins. CONCLUSIONS: Two-dimensional PAGE and mass spectrometry can identify proteins showing increased expression in lung adenocarcinoma. The association of specific isoforms of these proteins with clinical variables and understanding the regulation of their expression will aid in determination of their potential use as biomarkers in this cancer. PMID- 12114433 TI - Differential expression of FEZ1/LZTS1 gene in lung cancers and their cell cultures. AB - PURPOSE: The FEZ1/LZTS1 (FEZ1) gene, located on chromosome 8p22 (8p22), was identified recently as a candidate tumor suppressor gene. Because loss of heterozygosity at 8p21-22 is a frequent event in lung cancers, we studied FEZ1 alteration in short-term cultures of resected lung cancer tumors and cell lines. EXPERIMENTAL DESIGN: We examined FEZ1 expression in 17 non-small cell lung cancer (NSCLC), 19 small cell lung cancer (SCLC) cell lines, and 6 pairs of short-term cultures of resected NSCLCs and accompanying nonmalignant bronchial cells (NBECs) by reverse transcription-PCR and Western blotting. To investigate the mechanism for silencing, cells were cultured with 5-aza-2'-deoxycytidine or trichostatin A. We screened for genomic mutations by PCR-single-strand conformational polymorphism. RESULTS: Thirteen of 17 NSCLC (76%) and 3 of 19 SCLC (16%) of cell lines showed absent expression (P = 0.001). Of the paired NSCLC-NBEC cultures, 3 of 6 showed loss of expression in tumor cell cultures. In the cell lines retaining expression, the amplicon products in SCLCs were more intense than those of NSCLCs and NBECs. Expression of FEZ1 was not restored by 5-aza-2' deoxycytidine and trichostatin A. Although FEZ1 expression was moderately correlated with loss of heterozygosity of specific microsatellite makers at 8p21 22 in NSCLC cell lines, it was strongly correlated to D8S261 and LPL loci in SCLC cell lines. No mutation was found within cording region of FEZ1 by PCR-single strand conformational polymorphism. CONCLUSIONS: We found differential FEZ1 expression in NSCLC and SCLC cell lines, and the absent expression in 3 of 6 short-term cultures of NSCLC tumors. FEZ1 may be related to tumorigenesis of lung cancer. PMID- 12114435 TI - Serum osteoprotegerin levels in healthy controls and cancer patients. AB - PURPOSE: Osteoprotegerin (OPG) is a novel secreted member of the tumor necrosis factor receptor superfamily. In vitro, OPG blocks osteoclastogenesis in a dose dependent manner. Serum OPG levels were assayed in cancer patients and healthy control subjects using an ELISA. RESULTS: OPG levels in healthy controls were significantly higher in sera (0.17 ng/ml) than in plasma (0.14 ng/ml). OPG levels did not differ by age in either control group. Serum was available from patients with solid tumors (n = 145), hematological malignancies (n = 111), benign hematological disorders (n = 35), and rheumatologic diseases (n = 60). When adjusted for age and sex, there was no significant OPG elevation in the sera of patients with solid tumors compared with controls (0.2 versus 0.18 ng/ml). When analyzed by site of primary malignancy within the solid tumor patient group, serum OPG elevations were observed only in patients with colorectal cancer (0.29 ng/ml; P < 0.0001) and pancreatic cancer (0.35 ng/ml; P < 0.0001). When analyzed by site of metastasis within the solid tumor patient group, significant elevations in serum OPG were observed only in patients with liver metastases (0.29 ng/ml) and soft tissue metastases (0.21 ng/ml) but not in patients with bone or lung metastases. Within the hematological malignancy group, serum levels of OPG were significantly lower in patients with multiple myeloma (0.12 ng/ml) but were elevated in patients with Hodgkin's disease (0.29 ng/ml) and Non Hodgkin's Lymphoma (0.24 ng/ml; P = 0.048). CONCLUSIONS: Although some patients with malignancy have significant elevations of circulating OPG, these concentrations do not approach the level that would be expected to suppress osteoclast function. PMID- 12114436 TI - Genetic classification of colorectal cancer based on chromosomal loss and microsatellite instability predicts survival. AB - PURPOSE: Colorectal cancers harbor one of two distinct alterations, unilateral chromosomal loss as evidenced by a loss of heterozygosity (LOH) and microsatellite instability (MSI), as represented by the widespread insertion or deletion of simple repeat nucleotides. We investigated the relationships between the clinicopathological features and microsatellite alterations (LOH and MSI) of 168 colorectal cancers. EXPERIMENTAL DESIGN: The concerted and individual effects of various chromosomal losses on survival were comparatively analyzed using a reference panel of 40 microsatellite markers in eight cancer-related chromosomes, 3p, 4p, 5q, 8p, 9p, 13q, 17p, and 18q. RESULTS: Of the 168 colorectal cancers tested, 29 (17%) with high-frequency MSI were associated with good survival (P < 0.05). The extent of LOH detected in 139 (83%) cases without MSI was classified as low level involving three or fewer arms (35%), moderate level involving four arms (22%), or high level involving five or more arms (43%). High-level loss correlated with earlier onset, lymphatic invasion, and rectal location, whereas low-level loss was more common in proximal colon and stages I and II (P < 0.05). The survival curve and multivariate analysis identified high- and low-level chromosomal loss as the most significant predictor of poor and good survival, respectively (log-rank test, P < 0.0001), in patients with stage II (hazard ratio, 6.27; 95% confidence interval, 1.99-19.7; P = 0.0017) and those with stage III (hazard ratio, 10.89; 95% confidence interval, 2.54-46.77; P = 0.0013). Moderate chromosomal loss showed dual prognostic values associated with favorable stage II and unfavorable stage III. Single chromosomal losses tended to play a role as a part of the concerted chromosomal function. CONCLUSION: The classification of colorectal cancer based on chromosomal loss and MSI provides a prognostic index that reflects tumor pathobiology. PMID- 12114437 TI - Validation of the fluorinated 2-nitroimidazole SR-4554 as a noninvasive hypoxia marker detected by magnetic resonance spectroscopy. AB - PURPOSE: Tumor hypoxia is associated with poor prognosis and a more malignant tumor phenotype. SR-4554, a fluorinated 2-nitroimidazole, is selectively bioreduced and bound in hypoxic cells. We present validation studies of SR-4554 as a noninvasive hypoxia marker detected by fluorine-19 magnetic resonance spectroscopy ((19)F MRS) in the P22 carcinosarcoma, a tumor with clinically relevant hypoxia levels. EXPERIMENTAL DESIGN: Tumor-bearing female severe combined immunodeficient mice received SR-4554 at 180 mg/kg. Pharmacokinetic studies of parent SR-4554 in plasma and tumors were performed using high performance liquid chromatography-UV. Total SR-4554 (parent SR-4554 and bioreduction products) was monitored in tumor by (19)F MRS using a 4.7 T spectrometer, with continuous acquisition for up to 5 h. A parameter of total SR 4554 retention, the 3-h (19)F retention index ((19)FRI) was determined. Tumor pO(2), assessed polarographically, was decreased (5 mg/kg hydralazine or 100 mg/kg combretastatin A-4 phosphate) or increased [1 l/min carbogen (5% CO(2), 95% O(2)) plus 500 mg/kg nicotinamide], and the corresponding (19)FRI was measured. RESULTS: Comparative HPLC-UV- and MRS-derived assessments of parent and total SR 4554, respectively, indicated that concentrations of total SR-4554 consistently exceeded parent SR-4554, the differential increasing with time. This indicates formation and retention of SR-4554 bioreduction products in tumor, confirming the presence of hypoxia. The (19)FRI was higher in hydralazine- and combretastatin treated animals compared with unmodulated animals (P = 0.004 and 0.15, respectively) and animals receiving carbogen and nicotinamide (P = 0.0001 and 0.005, respectively). Significant correlations were demonstrated between mean (19)FRI and polarographic pO(2) parameters (P < 0.002). CONCLUSIONS: Retention of hypoxia-related SR-4554 bioreduction products can be detected in the clinically relevant P22 tumor by (19)F MRS, and the (19)FRI correlates with polarographically measured pO(2). These findings support the use of SR 4554 as a noninvasive hypoxia marker. PMID- 12114438 TI - Enhancing immunogenicity of a CTL epitope from carcinoembryonic antigen by selective amino acid replacements. AB - PURPOSE: Many epitopes from tumor antigens recognized by CTLs canbe poorly immunogenic. This low immunogenicity can be improved by carrying out amino acid replacements in their sequence. We have applied this strategy to enhance the immunogenicity of the HLA-A2-restricted CTL epitope CEA691 (IMIGVLVGV) from carcinoembryonic antigen (CEA), which is expressed by a wide variety of tumors. EXPERIMENTAL DESIGN: Substituted peptides from CEA691 were synthesized and tested in HLA-A2-binding assays, and in recognition by CEA691-specific CTL. Selected peptides were used to induce CTL responses in vivo in HLA-A2Kb transgenic mice and in vitro with human cells. RESULTS: Our experiments afforded several peptides with enhanced binding and/or recognition by CTL specific of CEA691. However, when HLA-A2Kb mice were immunized with these peptides only a few induced a CTL response that cross-reacted with CEA691. Additional replacement of their NH(2) terminal amino acid by Y (tyrosine) afforded peptides YMIGMLVGV and YMIGVLLGV with enhanced in vivo and in vitro immunogenicity than CEA691. Indeed, they activated a greater number of precursor cells that recognized CEA691-pulsed cells and tumor cells expressing CEA. CONCLUSIONS: Our results widen the possibility of treating CEA-expressing tumors with enhanced efficacy. PMID- 12114439 TI - Novel anti-CD30 recombinant immunotoxins containing disulfide-stabilized Fv fragments. AB - PURPOSE: To develop a novel targeting reagent to CD30 expressed on Hodgkin'sdisease and anaplastic large cell lymphoma, we made a panel of recombinant immunotoxins specific for CD30 using Fvs of newly produced anti-CD30 monoclonal antibodies (MAbs) and a M(r) 38,000 truncated mutant of Pseudomonas exotoxin. EXPERIMENTAL DESIGN: A group of MAbs against CD30 was produced and characterized for their reactivity and epitopes. Recombinant immunotoxins were made using the Fv genes cloned from the hybridomas. Their cytotoxic activities were examined on various CD30-positive cell lines. RESULTS: Six MAbs were produced. All reacted with recombinant soluble CD30 and to a CD30-Fc fusion protein, and bound to native CD30 expressed on Hodgkin's lymphoma-derived cell lines. The epitopes of the six MAbs were classified into two groups by a mutual competition assay for the binding to CD30 on cells. Sequencing the cDNAs revealed that all of the variable chains are unique except one valiable light that is shared by two MAbs. We made four disulfide stabilized Fv-based recombinant immunotoxins, in which the valiable heavy, which is genetically fused with truncated mutant of Pseudomonas exotoxin, forms a disulfide bond with the valiable light. The purified immunotoxins bound to recombinant soluble CD30 immobilized on a biosensor chip with K(d)s of 4-400 nM. Fluorescence-activated cell sorter analysis confirmed their specific binding. In vitro cytotoxicity tests showed that the immunotoxins specifically kill a variety of CD30-positive lymphoma cell lines as well as CD30-transfected A431 cells. The IC(50) ranged from 0.3 to 100 ng/ml. CONCLUSIONS: Four anti-CD30 disulfide stabilized Fv immunotoxins were successfully produced. Two of these showed good cytotoxic activity to various CD30-positive cell lines. These newly produced immunotoxins should be additionally evaluated for the treatment of CD30-positive lymphomas. PMID- 12114440 TI - Acquisition of agonistic properties of nonsteroidal antiandrogens after treatment with oncostatin M in prostate cancer cells. AB - PURPOSE: Interleukin-6 (IL-6), a proinflammatory cytokine the serum andtissue levels of which are elevated in prostate cancer patients, activates the androgen receptor (AR) in a ligand-independent and synergistic manner. Oncostatin M (OSM) is an IL-6 type cytokine that regulates the growth of prostate cancer cells in a paracrine fashion. The present study was designed to investigate the regulation of AR expression and function by OSM, as well as the efficacy of the nonsteroidal antiandrogens hydroxyflutamide and bicalutamide in the inhibition of AR-mediated signal transduction. EXPERIMENTAL DESIGN: Expression of the OSM receptor-beta in the prostate cancer cell lines LNCaP, PC-3, and DU-145 was investigated by reverse transcription-PCR. DU-145 and PC-3 cells were cotransfected with an androgen-responsive gene and AR cDNA. Reporter gene activity was measured after treatment with androgen and/or OSM in the absence or presence of antiandrogens or protein kinase inhibitors. AR expression after OSM treatment was assessed by Western blot. RESULTS: OSM receptor-beta expression was higher in DU-145 and PC-3 than in LNCaP cells. OSM caused ligand-independent activation of the AR in DU-145 cells, and the maximal activation was 62% of that induced by the synthetic androgen methyltrienolone. In the presence of OSM, hydroxyflutamide behaved as an AR agonist. Bicalutamide down-regulated AR activation caused by OSM only at a concentration of 1 microM. The inhibitor of the protein kinase A signaling pathway PKI and dn signal transducers and activators of transcription (STAT) 3 showed no effect on AR activation by OSM. The inhibitor of the MAPK pathway, PD 98059, caused only a minor down-regulation of OSM-induced reporter gene activity. OSM did not change AR expression in DU-145 cells transfected with AR cDNA. CONCLUSIONS: OSM is a member of the IL-6 family of cytokines, which causes ligand independent activation of the AR without altering receptor expression. In contrast to AR activation by IL-6, nonsteroidal AR antagonists act as agonists in the presence of OSM. This may be attributable to recruitment of different intermediary signal transduction proteins by OSM and IL-6, respectively. The acquisition of agonistic properties of AR blockers in the presence of OSM might compromise use of these drugs in prostate cancer treatment. PMID- 12114441 TI - Clinicopathological significance of epigenetic inactivation of RASSF1A at 3p21.3 in stage I lung adenocarcinoma. AB - PURPOSE: Chromosome 3p is deleted frequently in various types of human cancers, including lung cancer. Recently, the RASSF1A gene was isolated from the 3p21.3 region homozygously deleted in lung and breast cancer cell lines, and it was shown to be inactivated by hypermethylation of the promoter region in lung cancers. In this study, we investigated the pathogenetic and clinicopathological significances of RASSF1A methylation in the development and/or progression of lung adenocarcinoma. EXPERIMENTAL DESIGN: Association of RASSF1A methylation with clinicopathological features, allelic imbalance at 3p21.3, p53 mutations, and K ras mutations was examined in 110 stage I lung adenocarcinomas. RESULTS: Thirty five of 110 (32%) tumors showed RASSF1A methylation. RASSF1A methylation was dominantly detected in tumors with vascular invasion (P = 0.0242) or pleural involvement (P = 0.0305), and was observed more frequently in poorly differentiated tumors than in well (P = 0.0005) or moderately (P = 0.0835) differentiated tumors. Furthermore, RASSF1A methylation correlated with adverse survival by univariate analysis (P = 0.0368; log-rank test) as well as multivariate analysis (P = 0.032,; risk ratio 2.357; 95% confidence interval, 1.075-5.169). The correlation between RASSF1A methylation and allelic imbalance at 3p21.3 was significant (P = 0.0005), whereas the correlation between RASSF1A methylation and p53 mutation was borderline (P = 0.0842). However, there was no correlation or inverse correlation between RASSF1A methylation and K-ras mutation (P = 0.2193). CONCLUSIONS: These results indicated that epigenetic inactivation of RASSF1A plays an important role in the progression of lung adenocarcinoma, and that RASSF1A hypermethylation appears to be a useful molecular marker for the prognosis of patients with stage I lung adenocarcinoma. PMID- 12114442 TI - Inhibition of nuclear factor kappaB activation in PC3 cells by genistein is mediated via Akt signaling pathway. AB - Prostate cancer is the second leading cause of cancer-related deaths in menin the United States. Genistein is a prominent isoflavonoid found in soy products and has been proposed to be responsible for lowering the rate of prostate cancer in Asians. However, the molecular mechanism(s) by which genistein elicits its effects on prostate cancer cells has not been fully elucidated. We have previously shown that genistein induces apoptosis and inhibits the activation of nuclear factor kappaB (NF-kappaB) pathway, which could be mediated via Akt signaling pathway, the most important survival pathway in cellular signaling. In this study, we investigated whether there is any cross-talk between Akt and NF kappaB during genistein-induced apoptosis in PC3 prostate cancer cells. We found that genistein inhibits cell growth and induces apoptotic processes in PC3 prostate cancer cells but not in nontumorigenic CRL-2221 human prostate epithelial cells. Immunoprecipitation, kinase assays, and Western blot analysis showed that genistein specifically inhibits Akt kinase activity and abrogates the epidermal growth factor-induced activation of Akt in prostate cancer cells. NF kappaB DNA-binding analysis and transfection studies with Akt cDNA and NF-kappaB Luc constructs revealed that Akt transfection results in the induction of NF kappaB activation and this is completely inhibited by genistein treatment. Moreover, genistein abrogated the epidermal growth factor-induced activation of NF-kappaB, which was mediated via Akt signaling pathway. From these results, we conclude that the inhibition of Akt and NF-kappaB activity and their cross-talk provide a novel mechanism by which genistein inhibits cell growth and induces apoptotic processes in tumorigenic but not in nontumorigenic prostate epithelial cells. PMID- 12114443 TI - The effect of second-line antiestrogen therapy on breast tumor growth after first line treatment with the aromatase inhibitor letrozole: long-term studies using the intratumoral aromatase postmenopausal breast cancer model. AB - PURPOSE: The aromatase inhibitors letrozole and anastrozole have been approvedrecently as first-line treatment options for hormone-dependent advanced breast cancer. Although it is established that a proportion of patients who relapse on first-line tamoxifen therapy show additional responses to aromatase inhibitors, it has not been determined whether tumors that acquire resistance to aromatase inhibitors in the first line remain sensitive to second-line therapy with antiestrogens. The aim of this study was to determine whether aromatase transfected and hormone-dependent MCF-7Ca human breast cancer cells remain sensitive to antiestrogens after: (a) long-term growth in steroid-depleted medium in vitro; and (b) long-term treatment with the aromatase inhibitor letrozole in vivo. METHODS: In the first approach, a variant of the MCF-7Ca human breast cancer cell line was selected that had acquired the ability to grow in estrogen depleted medium after 6-8 months of culture. Steroid-deprived UMB-1Ca cells were analyzed for aromatase activity levels, hormone receptor levels, and sensitivity to estrogens and antiestrogens in vitro and in vivo. In the second approach, established MCF-7Ca breast tumor xenografts were treated with letrozole 10 microg/day for 12 weeks followed by 100 microg/day for 25 weeks until tumors acquired the ability to proliferate in the presence of the drug. Long-term letrozole-treated tumors were then transplanted into new mice, and the effects of antiestrogens and aromatase inhibitors on tumor growth were determined. RESULTS: Steroid-deprived UMB-1Ca breast cancer cells continued to express aromatase activity at levels comparable with the parental cell line. However, compared with MCF-7Ca cells, UMB-1Ca cells expressed elevated levels of functionally active estrogen receptor. The growth of UMB-1Ca cells in vitro was inhibited by the antiestrogens tamoxifen and faslodex and tumor growth in vivo was inhibited by tamoxifen. In the second approach, the time for MCF-7Ca tumor xenografts to approximately double in volume after being treated sequentially with the increasing doses of letrozole was thirty-seven weeks. Long-term letrozole-treated tumors continued to express functionally active aromatase. When transplanted into new mice, growth of the long-term letrozole-treated tumors was slowed by tamoxifen and inhibited more effectively by faslodex. Tumor growth was refractory to the aromatase inhibitors anastrozole and formestane but, surprisingly, showed sensitivity to letrozole. CONCLUSIONS: Steroid-deprived UMB-1Ca human breast cancer cells selected in vitro and long-term letrozole-treated MCF-7Ca breast tumor xenografts remain sensitive to second-line therapy with antiestrogens and, in particular, to faslodex. This finding is associated with increased expression of functionally active estrogen receptor after steroid-deprivation of MCF-7Ca human breast cancer cells in vitro. PMID- 12114444 TI - Frequent administration of angiogenesis inhibitor TNP-470 (AGM-1470) at an optimal biological dose inhibits tumor growth and metastasis of metastatic human transitional cell carcinoma in the urinary bladder. AB - PURPOSE: The angiogenic inhibitor TNP-470 (AGM-1470, O-chloracetyl-carbamoyl fumagillol) has been reported to inhibit the growth of human transitional cell carcinoma (TCC) in the urinary bladder. However, it is still unknown whether TNP 470 inhibits metastasis of TCC. Here, we identify an efficient protocol using TNP 470, and optimize its antitumor and antimetastatic effects on human TCC in the urinary bladder. EXPERIMENTAL DESIGN: In vitro, the human metastatic TCC cell line 253J B-V and human umbilical vascular endothelial cells were treated with TNP-470, and examined for cell growth and protein production of angiogenic factors. To study in vivo effects of TNP-470, 253J B-V cells were implanted orthotopically into athymic nude mice. TNP-470 was administered in several dosing and scheduling regimens, and its effects on tumor growth, metastasis, intratumor neovascularization, and mRNA expression of angiogenic factors were determined in both nonestablished and established tumors. RESULTS: In vitro treatment with TNP 470 inhibited cell growth and production of basic fibroblast growth factor protein in 253J B-V and human umbilical vascular endothelial cells in a dose dependent manner. In vivo daily administration of TNP-470 significantly inhibited tumor growth (P < 0.001), metastasis (P < 0.05), intratumor neovascularization (P < 0.005), and mRNA expression of basic fibroblast growth factor and MMP-9 (P < 0.005), in both nonestablished and established tumors. Increasing the daily dose did not increase the effect on tumor growth, metastasis, and angiogenesis; however, the drug-induced toxicity did increase in a dose-dependent manner. CONCLUSIONS: Frequent administration of TNP-470 at an optimal biological dose provided maximal antitumor and antimetastatic effects of human TCC of the urinary bladder. It may function by down-regulating angiogenic factors. PMID- 12114445 TI - Synergistic activation of the androgen receptor by bombesin and low-dose androgen. AB - PURPOSE: Neuropeptide growth factors such as bombesinare implicated in progression to androgen-independent prostate cancer (PC). We examined the impact of bombesin on androgen receptor (AR)-mediated gene expression. EXPERIMENTAL DESIGN: The AR together with the AR-responsive probasin ARR(3)tk-luc or PSA-pPUE ELB-luc promoter was cotransfected into Swiss 3T3 and PC-3 cells, both of which express high-affinity bombesin receptors; the cells were incubated with bombesin (0-50 nM) and dihydrotestosterone (DHT; 0-10 nM), and luciferase activities were measured. DHT increased transcription approximately 40-fold at doses of 1 and 10 nM but had no effect at 10 pM. Bombesin alone, or with 1 or 10 nM DHT, did not further increase transcription. However, 5 nM bombesin and 10 pM DHT, doses that by themselves had no effect, resulted in a approximately 20 fold increase in transcription (P < 0.005). This synergistic effect was blocked by bombesin receptor antagonists and recombinant neutral endopeptidase, which hydrolyzes bombesin. Bombesin and DHT together also increased binding of nuclear extracts from PC-3 cells transfected with AR to a consensus androgen response element in mobility shift assays and increased the level of secreted prostate-specific antigen in LNCaP cell supernatant compared with DHT or bombesin alone. Immunoprecipitation of AR from (32)P-labeled LNCaP cells revealed that 5 nM bombesin + 10 pM DHT induced AR phosphorylation comparable with 1 nM DHT, whereas bombesin or 10 pM DHT alone did not. CONCLUSIONS: These data indicate that bombesin can synergize with low (castrate) levels of DHT to induce AR-mediated transcription and suggest that neuropeptides promote AR-mediated signaling in androgen-independent prostate cancer. PMID- 12114446 TI - A randomized controlled trial of octreotide pamoate long-acting release and carboplatin versus carboplatin alone in dogs with naturally occurring osteosarcoma: evaluation of insulin-like growth factor suppression and chemotherapy. AB - PURPOSE: The purpose of this research was to determine whether insulin-like growth factor (IGF) suppression, using a long-acting analogue of somatostatin (OncoLAR, octreotide pamoate long-acting release), will decrease chemotherapy resistance by eliminating an important survival signal to osteosarcoma (OSA) cells in a relevant naturally occurring cancer model. EXPERIMNETAL DESIGN: We conducted a randomized, blinded, placebo-controlled preclinical study in pet dogs with naturally occurring OSA. The study compared primary tumor necrosis and apoptosis, and survival of pet dogs receiving OncoLAR and carboplatin chemotherapy compared with dogs receiving placebo and carboplatin. RESULTS: Dogs receiving OncoLAR had suppression of serum IGF levels by approximately 43% without toxicity. No differences in primary tumor necrosis, apoptosis, tumor IGF mRNA expression, or survival were seen between the dogs receiving OncoLAR plus chemotherapy compared with OncoLAR alone. CONCLUSION: The suppression of IGF levels by the extent and/or duration achieved in the trial was not sufficient to improve chemotherapy-related antitumor effects in pet dogs with OSA. PMID- 12114447 TI - Synergistic therapy of human ovarian carcinoma implanted orthotopically in nude mice by optimal biological dose of pegylated interferon alpha combined with paclitaxel. AB - The purpose of this study was to optimize the antitumor andantiangiogenic activities of pegylated IFN-alpha (PEG-IFN-alpha)alone or in combination with paclitaxel against SKOV3ip1 human ovarian cancer cells growing orthotopically in female nude mice. Seven days after the i.p. implantation of tumor cells, groups of mice (n = 10) were injected s.c. once per week (for 4 weeks) with different doses of PEG-IFN-alpha (3,500, 7,000, 35,000, and 350,000 units). PEG-IFN-alpha at 7,000 units significantly decreased tumor incidence and volume. At doses exceeding 7,000 units, PEG-IFN-alpha was less efficacious. In another set of studies conducted 7 days after the i.p. implantation of SKOV3ip1 cells, groups of mice (n = 10) received (once per week for 4 weeks) either s.c. administrations of PEG-IFN-alpha (7,000 units), i.p. injections of paclitaxel (100 microg/wk), or a combination of PEG-IFN-alpha and paclitaxel. The mice were killed 7 days after the last treatment, and tumor burden was assessed. Administration of PEG-IFN alpha at the optimal biological dose (7,000 units) in combination with paclitaxel significantly decreased angiogenesis and progressive growth of human ovarian carcinoma cells in a synergistic fashion. The combination therapy produced the most significant inhibition in expression of the proangiogenic molecules basic fibroblast growth factor and matrix metalloproteinase-9. Decreased microvessel density, decreased proliferating cell nuclear antigen staining, and increased endothelial cell apoptosis also correlated with therapeutic success. Collectively, the data suggest that combining the optimal biological dose of PEG IFN-alpha with paclitaxel may provide a novel and effective approach to the treatment of human ovarian carcinoma. PMID- 12114448 TI - Methotrexate intracellular disposition in acute lymphoblastic leukemia: a mathematical model of gamma-glutamyl hydrolase activity. AB - Methotrexate (MTX) is an antifolate that is widely used for the treatment of childhood acute lymphoblastic leukemia (ALL) and a number of other malignant and nonmalignant diseases. Within cells, MTX is metabolized to more active methotrexate polyglutamates (MTXPG), and these polyglutamates are subsequently cleaved in lysosomes by gamma-glutamyl hydrolase (GGH). GGH is reported to act as either an endopeptidase or an exopeptidase, exhibiting species differences in these functions. To better define the in vivo functions of GGH in human leukemia cells, we characterized GGH activity with different MTXPG substrates (MTX with three to five glutamates) in human T- and B-lineage leukemia cell lines, and in primary leukemia cells from newly diagnosed patients with ALL. Parameters estimated from fitting a series of hypothetical mathematical models to the data revealed that the experimental data were best fit by a model where GGH simultaneously cleaved multiple glutamyl residues, with highest activity at cleaving the outermost or two outermost residues from a polyglutamate chain. The model also revealed that GGH has a higher affinity for longer chain polyglutamates. Together, these findings provide new insights to the intracellular disposition of MTX in human ALL cells, and provides a mechanism based model for characterizing differences among patients and genetic subtypes of ALL. PMID- 12114449 TI - Src family kinase inhibitor PP2 restores the E-cadherin/catenin cell adhesion system in human cancer cells and reduces cancer metastasis. AB - The E-cadherin/catenin cell adhesion system is often down-regulatedin epithelial tumors. This is thought to play an important role in cancer invasion and metastasis. Restoring this system may enable suppression of the metastatic spread of cancer. This study examined the effect of Src family kinase inhibitor PP2 on E cadherin-mediated cell-cell adhesion and metastatic potentials. In cell aggregation assays, PP2 stimulated the aggregation of colon, liver, and breast cancer cells. In vitro cultures of cancer cells showed that PP2 induced strong cell-cell contact. Immunoblot analysis showed that PP2 enhanced E cadherin/catenin expression and that increased E-cadherin/catenin proteins were strongly associated with the actin cytoskeleton. Northern blot studies indicated that the observed increase of E-cadherin/catenin protein content was due to their increased gene expression. After the spleens of severe combined immunodeficient mice were inoculated with cancer cells, treatment with PP2 for 3 weeks markedly reduced the rate of liver metastasis, compared with the control counterparts. Our data demonstrate that PP2 can activate the functioning of the E-cadherin-mediated cell adhesion system, which is associated with the suppression of metastasis in cancer cells. Thus, selective inhibition of Src activation may be potentially useful in the prevention of cancer metastasis. PMID- 12114450 TI - Pharmacokinetics of intrathecal gemcitabine in nonhuman primates. AB - PURPOSE: Gemcitabine is an excellent candidate for regional therapy. We quantified cerebrospinal fluid (CSF) and plasma concentrations of gemcitabine and its inactive metabolite, 2',2'-difluorodeoxyuridine (dFdU), in nonhuman primates given intrathecal gemcitabine. EXPERIMENTAL DESIGN: Three nonhuman primates received 5 mg of gemcitabine via lateral ventricle. CSF was sampled from the fourth ventricle in all of the animals and the lumbar space in one, and one had plasma sampled. One animal had ventricular CSF sampled after receiving 5 mg intralumbar gemcitabine. Gemcitabine and dFdU were measured by high-performance liquid chromatography. Three additional animals had 5 mg intralumbar gemcitabine administered weekly for 4 weeks and were monitored for toxicity. RESULTS: At 37 degrees C in vitro, gemcitabine was stable in CSF. Ventricular delivery of gemcitabine produced peak ventricular CSF gemcitabine concentrations of 297 +/- 105 microg/ml. After 6 h, the concentrations were <0.03 microg/ml. Intrathecal gemcitabine was rapidly and extensively converted to dFdU. CSF dFdU concentrations increased to 82 microg/ml at 1 h and then declined to very low values by 24 h. After intraventricular administration, CSF gemcitabine and dFdU area(s) under the curve (AUC) were 251 +/- 85 and 249 +/- 88 microg/ml x h. Intralumbar gemcitabine produced lower ventricular CSF gemcitabine and dFdU concentrations than did intraventricular gemcitabine. The plasma gemcitabine AUC associated with 5 mg of intraventricular gemcitabine was 2 mg/ml x h, which was >200-fold lower than the CSF gemcitabine AUC in the same animal. Transient CSF pleocytosis was the only toxicity observed. CONCLUSIONS: Our results demonstrate a large pharmacokinetic advantage of intrathecal gemcitabine and support a planned Phase I clinical trial of this dosing strategy. PMID- 12114451 TI - Significant growth inhibition of human lung cancer cells both in vitro and in vivo by the combined use of a selective cyclooxygenase 2 inhibitor, JTE-522, and conventional anticancer agents. AB - This study reports that a selective COX-2 inhibitor JTE-522inhibits both in vitro and in vivo growth of human lung cancer cells as a single agent. Furthermore, the adjunct use of JTE-522 is shown to significantly enhance treatment efficacy of conventional anticancer drugs not only in vitro but also in vivo without causing any noticeable side effects. Indeed, IC(50)s of various anticancer agents in vitro were reduced by up to 70%, whereas the combination therapy of JTE-522 with docetaxel and vinorelbine inhibited tumor growth in vivo by 65 and 55%, respectively. Taken together, these findings suggest that the use of a selective COX-2 inhibitor in the treatment of lung cancer may be promising, especially because of its enhancement of the treatment efficacy of conventional anticancer agents without compromising quality of life. PMID- 12114452 TI - Selective inhibition of the epidermal growth factor receptor by ZD1839 decreases the growth and invasion of ovarian clear cell adenocarcinoma cells. AB - The mechanism that regulates the growth of ovarian clearcell adenocarcinoma (CCA) are not well understood. We investigated the role of several growth factors that bind to membrane tyrosine kinase receptors and added them to the ovarian CCA cell lines KK, RMG-1, and HAC-II to evaluate their effect on growth and cellular invasion. Epidermal growth factor and transforming growth factor-alpha significantly stimulated the growth and invasion of CCA cell lines in vitro. ZD1839, an epidermal growth factor receptor-tyrosine kinase inhibitor, decreased the growth and invasion of CCA cell lines in vitro and in vivo inhibited the growth of xenografts of the CCA cell line RMG-1. Severe combined immunodeficient mice bearing RMG-1 xenografts treated with ZD1839 survived for longer than the untreated control group. From these findings, we conclude that ZD1839 may offer a new and effective treatment for ovarian CCA. PMID- 12114453 TI - A new type of antimetastatic peptide derived from fibronectin. AB - PURPOSE: We found previously that fibronectin (FN) has a cryptic functional site (YTIYVIAL sequence within the 14th type III repeat) opposing cell adhesion to extracellular matrix. A 22-mer FN peptide containing this site, termed FNIII14, inhibits beta1 integrin-mediated adhesion without binding to integrins. The present study shows that FNIII14 has the potential to prevent lymphoma cell metastasis. EXPERIMENTAL DESIGN: Antimetastatic effect of FNIII14 has been evaluated through in vitro or in vivo experiments. RESULTS: FNIII14 inhibited the integrin alpha4beta1-mediated B lymphoma Ramos cell adhesion to VCAM-1 on venule endothelial cells, as well as to FN. Murine T lymphoma L5178Y-ML25 cells, which are known to metastasize to liver and spleen, preferentially adhered to vitronectin (VN) and migrated toward VN concentration gradients. FNIII14 abrogated both the integrin alphavbeta3-mediated adhesion and migration of L5178Y ML25 cells. Inhibition of the alphavbeta3mediated L5178Y-ML25 cell adhesion by FNIII14 was reversed by phenylarsine oxide, a protein tyrosine phosphatase inhibitor. In addition, FNIII14 abrogated the VN-stimulated tyrosine phosphorylation of intracellular signaling proteins, including focal adhesion kinase (p125(FAK)) and paxillin, suggesting that such a diversity of FNIII14 effects might be because of the negative regulation of p125(FAK) and paxillin tyrosine phosphorylation, which has been involved in adhesion signals transduced by different integrins. The in vivo experiment using a murine metastasis model showed that FNIII14 would inhibit liver and spleen metastases of L5178Y-ML25 cells at a dose much lower than that of RGDS. CONCLUSIONS: FNIII14 might be applicable as a new type of antimetastatic agent distinct from integrin-binding peptides. PMID- 12114455 TI - Orthodontic speciality training in the UK. PMID- 12114454 TI - Chemoembolization of the lung improves tumor control in a rat model. AB - PURPOSE: The novel method of organ-specific drug application we present here is unilateral chemoembolization of the lung by injecting the pulmonary artery with degradable starch microspheres and cytotoxic drugs to improve tumor control in lung metastases. EXPERIMENTAL DESIGN: In a solitary metastasis rat model (CC531 adenocarcinoma), we studied the clinical and histological tumor response as well as subacute toxicity of the lung. Fourteen days after tumor induction, animals were randomized into five groups. Groups I and II served as controls. Group III received carboplatin i.v. (45 mg/kg). Isolated lung perfusion with buffered starch solution and carboplatin (15 mg/kg) was installed in group IV. Chemoembolization with carboplatin (15 mg/kg) was performed in group V. RESULTS: Seven days later, the difference in the tumor volume before and after treatment was +422 mm(3) (+/-226) in group I, +697 mm(3) (+/-423) in group II, +70 mm(3) (+/-31) in group III, -8 mm(3) (+/-17) in group IV, and -17 mm(3) (+/-16) in group V (P < 0.05 groups IV and V versus groups I, II, and III). No pleural spread was observed in groups IV and V. Histologically, the area of tumor necrosis was largest in group IV. Mild alveolar cell hyperplasia, pulmonary edema, and hemorrhage without subacute fibrotic changes were noted in all groups. CONCLUSION: This is the first study to perform chemoembolization of the lung. Compared with i.v. therapy, chemoembolization was more effective without serious toxicity. Its efficacy was comparable with that of isolated lung perfusion but less stressful for a possible clinical application. PMID- 12114456 TI - The British Orthodontic Society medal of the Intercollegiate M.Orth. of the Royal College of Surgeons of London and Glasgow 2001 and the William Houston medal of the M.Orth. of the Royal College of Surgeons of Edinburgh 2001. PMID- 12114457 TI - Unilateral distalization of a maxillary molar with sliding mechanics: a case report. AB - INTRODUCTION: A unilateral Class II relationship could arise due to early loss of an upper second deciduous molar on one side during the mixed dentition period. This would allow the mesial drift of the molars, which may block the eruption of the second premolar. METHODS AND RESULTS: A 15-year 8-month-old male patient presented with a Class II molar relationship on the right, and Class I canine and molar relationship on the left side. His E was extracted when he was 5 years old. The 54 were impacted and the 3 was ectopically positioned due to the space loss from the mesial migration of the 76. In addition 21 1 were in cross-bite. Skeletally he had Class III tendency with low MMPA. He presented with a straight profile and retruded upper lip. For maxillary molar distalization, a newly developed 'Keles Slider' was used. The appliance was composed of one premolar and two molar bands, and the anchorage unit was composed of a wide Nance button. 46 were connected to the Nance button and, therefore, included into the anchorage unit. The point of distal force application was close to the centre of resistance of the 6 and parallel to the occlusal plane. Ni-Ti coil springs were used and 200 g of distal force was applied. Seven months later the space required for eruption of the permanent premolars and canine was regained, and the anterior cross-bite corrected. The appliance was removed and final alignment of the teeth was achieved with fixed appliances. CONCLUSION: At the end of the second phase treatment Class I molar and canine relationship was achieved on the both sides, the anterior cross-bite was corrected, inter-incisal angle was improved, and ideal overbite and overjet relationship was achieved. The active treatment time was 27 months. PMID- 12114458 TI - Orthodontic facebows: safety issues and current management. AB - Some patients treated with extra-oral traction provided by simple elasticated materials and a standard facebow have experienced problems with the standard facebow coming out of the buccal tubes at night and the catapult effect of the extra-oral traction. The disengagement of the facebow at night has affected the success of treatment and occasionally injured the patient. This paper draws on material from a variety of papers and lists the known causes and considers the associated safety issues. It also provides some clinical tips and makes several suggestions for the continued use of this very useful form of additional orthodontic anchorage. PMID- 12114460 TI - A cephalometric inter-centre comparison of growth in children with cleft lip and palate. AB - AIM: To examine whether the treatment provided by the Mount Vernon Cleft Team produces craniofacial growth outcomes comparable with that of the Oslo Team. LOCATION: Mount Vernon Hospital, Middlesex, UK. DESIGN: A retrospective cephalometric investigation. SUBJECTS: Seventy-five Mount Vernon children and 150 Oslo children with complete unilateral or bilateral clefts of the lip and palate METHOD: The subjects were matched for age, gender, and cleft type, and their radiographs were digitized. The radiographs from each site were grouped according to patient age (9-11 or 14-16) and cleft classification (bilateral/unilateral). Patients with associated craniofacial anomalies were excluded from the study. RESULTS: Of the four variables studied (SNA, SNPg, NGn, sNANsPG) significant differences in maxillary growth were noted for bilateral and unilateral cleft groups at 14-16 years of age. The soft tissue profile was significantly flatter in bilateral and unilateral Mount Vernon cases at 14-16 years. The craniofacial growth exhibited by the Mount Vernon patients demonstrated 3.9-5.1 degrees reduction in maxillary prominence with respect to the Oslo sample. The bilateral cases from Mount Vernon had greater anterior face heights at 14-16 years. CONCLUSION: The treatment provided by the Mount Vernon Cleft team leads to a reduced maxillary prominence in children aged 14-16 years compared with the Oslo sample. This reduction is statistically significant in unilateral cleft lip and palate. PMID- 12114461 TI - Effectiveness of community-based salaried orthodontic services provided in England and Wales. AB - OBJECTIVES: To assess the effectiveness of the salaried Community Orthodontic Services in England and Wales, using occlusal indices, and to determine the predictors of treatment outcome. DESIGN: A retrospective investigation. A random stratified sample of districts where Community Orthodontic Services are provided was selected and visited during 1997. METHOD: All community orthodontists in England and Wales, and CDS managers who could be identified were asked to take part in this study. A stratified random sample of 15 per cent of the districts where community orthodontic services were provided was selected and a sample of the records of treated patients was examined. RESULTS: The orthodontists in the sample were providing treatment for patients clearly in need of treatment. There were, however, some variations between districts. Similarly, when the effectiveness of treatment in terms of dento-alveolar change was evaluated, the mean change in PAR and percentage PAR reduction was high. Again, there were variations between the districts. CONCLUSIONS: The Community Orthodontic Service provides effective orthodontic treatment to many individuals clearly in need of that treatment. The most significant predictor of treatment outcome was the use of two-arch fixed appliances, which produced the best treatment outcome PMID- 12114462 TI - Are photographic records reliable for orthodontic screening? AB - AIM: The aim of the study was to evaluate the reliability of a panel of orthodontists for accepting new patient referrals based on clinical photographs. SAMPLE: Eight orthodontists from Greater Manchester, Lancashire, Chester, and Derbyshire observed clinical photographs of 40 consecutive new patients attending the orthodontic department, Hope Hospital, Salford. METHOD: They recorded whether or not they would accept the patient, as a new patient referral, in their department. Each consultant was asked to take into account factors, such as oral hygiene, dental development, and severity of the malocclusion. STATISTICS: Kappa statistic for multiple-rater agreement and kappa statistic for intra-observer reliability were calculated. RESULTS: Inter-observer panel agreement for accepting new patient referrals based on photographic information was low (multiple rater kappa score 0.37). Intra-examiner agreement was better (kappa range 0.34-0.90). CONCLUSION: Clinician agreement for screening and accepting orthodontic referrals based on clinical photographs is comparable to that previously reported for other clinical decision making. PMID- 12114463 TI - Experimental tooth movement under light orthodontic forces: rates of tooth movement and changes of the periodontium. AB - AIM: To investigate light forces for experimental tooth movement. METHOD: Light orthodontic forces of 1.2, 3.6, 6.5, and 10 g force (gf) were applied for 14 days to move rat molars, and the effects of the forces on the rate of tooth movement and changes of the periodontium were examined. RESULTS: In the early period, despite the different levels of force used in each group, there were no significant differences in tooth displacement. From hour 56 to day 14, the tooth displacement in the 1.2 gf group was significantly smaller than that in the other groups and the rate was nearly constant. The rates of tooth displacement in the 3.6, 6.5, and 10 gf groups fluctuated repeatedly, while the orthodontic forces gradually decreased. CONCLUSION: Experimental tooth movement in rats, tipping without friction under light forces, were either constant or fluctuated in cycles of several days' duration. This is in contradiction to the three-phases-theory of tooth movement described in previous investigations using heavy forces. PMID- 12114465 TI - How to do... a case report. PMID- 12114464 TI - Attitudes of UK consultants to teledentistry as a means of providing orthodontic advice to dental practitioners and their patients. AB - OBJECTIVE: To determine UK orthodontic consultants' attitudes to the provision of orthodontic advice to general dental practitioners by electronic means. DESIGN: Questionnaire. SETTING: Conducted by email and surface mail as appropriate in August 2000. SUBJECTS: All those UK NHS orthodontic consultants contained in the membership lists of the Consultant Orthodontists Group of the British Orthodontic Society. OUTCOME: An 86 per cent response was obtained from the 231 consultants. RESULTS: More than half (58 per cent) of the consultants were interested in providing an electronic diagnostic service for the general dental practitioners in their locality and 70 per cent were in favour of further research into this possibility. Provided this was mediated through their GDP, only 26% would oppose consultant advice being given electronically from a centralized source. CONCLUSIONS: A majority of UK orthodontic consultants support the concept of using teledentistry to make their advice more accessible to dentists and patients. PMID- 12114469 TI - Clinical photographs--the gold standard. AB - This survey was carried out to allow a minimum data set required for intra- and extra-oral photographs to be established. In 1999 a questionnaire was sent to members of the Angle Society of Europe to establish their current clinical practice with regard to extra- and intra-oral photography. The Angle Society was chosen because of their stated commitment to a high standard of record keeping and of clinical care. Results showed that a full series of extra- and intra-oral photographs were taken both before and after treatment, as well as stage photographs during treatment on many cases. The need for each of these photographs will be discussed in some detail, and recommendations will be made as to what would be considered the Gold standard before, during, and after a course of orthodontic treatment. PMID- 12114470 TI - Teaching and learning: an update for the orthodontist. AB - Developments in teaching and learning have implications for every orthodontist. This paper describes some of the theories of teaching and learning that have led to a quiet revolution in higher education. Developments have included the incorporation of self-directed and problem-based learning concepts, together with a more active and interactive role for the learner. The importance of these changes for orthodontic education is discussed. PMID- 12114472 TI - An ex vivo assessment of resin modified glass ionomer cement in relation to bonding technique by C. J. Larmour and D. R. Stirrups, J Orthod 28(3). PMID- 12114473 TI - The effect of a single BRCA2 mutation on cancer in Iceland. AB - OBJECTIVE: To estimate the risk of malignant diseases in families of probands with the same mutation in the BRCA2 gene. DESIGN: A cohort study using record linkage of a breast cancer family resource and the Icelandic Cancer Registry. SETTING: Iceland. SUBJECTS: Families of 995 breast cancer patients, from which 887 were tested for a single founder 999del5 mutation; 90 had the mutation and 797 did not. RESULTS: Relatives of probands with the mutation had significantly increased relative risk (RR) of breast cancer. For first degree relatives, the RR was 7.55 (95% CI 6.04 to 9.03) but was 1.72 (95% CI 1.49 to 1.96) in first degree relatives of probands without the mutation. For prostate and ovarian cancer, the first and second degree relatives of probands with the mutation had a significantly increased RR, but in families of probands without the mutation no significant familial risk was found. CONCLUSIONS: The 999del5 mutation in the BRCA2 gene explains a substantial proportion of familial risk of breast cancer in Iceland, but significant familial risk remains in relatives of probands without the mutation. For prostate and ovarian cancer, the mutation accounts for most of the familiality observed in families of breast cancer patients. PMID- 12114475 TI - Genetic and functional analysis of the von Hippel-Lindau (VHL) tumour suppressor gene promoter. AB - The VHL gatekeeper tumour suppressor gene is inactivated in the familial cancer syndrome von Hippel-Lindau disease and in most sporadic clear cell renal cell carcinomas. Recently the VHL gene product has been identified as a specific component of a SCF-like complex, which regulates proteolytic degradation of the hypoxia inducible transcription factors HIF-1 and HIF-2. pVHL is critical for normal development and mRNA expression studies suggest a role in nephrogenesis. Despite the importance of VHL in oncogenesis and development, little is known about the regulation of VHL expression. To investigate VHL promoter activity, we performed comparative sequence analysis of human, primate, and rodent 5' VHL sequences. We then proceeded to deletion analysis of regions showing significant evolutionary conservation between human and rat promoter sequences, and defined two positive and one negative regulatory regions. Analysis of specific putative transcription factor binding sites identified a functional Sp1 site, which was shown to be a regulatory element. Overlapping Sp1/AP2 sites were also identified and candidate E2F1 binding sites evaluated. Three binding sites for as yet unidentified transcription factors were mapped also. These investigations provide a basis for elucidating the regulation of VHL expression in development, the molecular pathology of epigenetic silencing of VHL in tumourigenesis, and suggest a possible link between Sp1, VHL, and nephrogenesis. PMID- 12114476 TI - Linkage of otosclerosis to a third locus (OTSC3) on human chromosome 6p21.3-22.3. AB - Clinical otosclerosis (OMIM 166800/605727) has a prevalence of 0.2-1% among white adults, making it the single most common cause of hearing impairment in this group. It is caused by abnormal bone homeostasis of the otic capsule with the consequent development of sclerotic foci that invade the stapedio-vestibular joint (oval window) interfering with free motion of the stapes. Impaired ossicular chain mobility results in a conductive hearing loss. We identified the first locus for otosclerosis (OTSC1) on chromosome 15 in 1998 and reported a second locus (OTSC2) on chromosome 7 last year. Here we present results of a genome wide linkage study on a large Cypriot family segregating otosclerosis. Results of this study exclude linkage to OTSC1 and OTSC2 and identify a third locus, OTSC3, on chromosome 6p. The defined OTSC3 interval covers the HLA region, consistent with reported associations between HLA-A/HLA-B antigens and otosclerosis. PMID- 12114477 TI - Close relatives: distant relations. PMID- 12114478 TI - Analysis of the phenotypic abnormalities in lymphoedema-distichiasis syndrome in 74 patients with FOXC2 mutations or linkage to 16q24. AB - INTRODUCTION: Lymphoedema-distichiasis syndrome (LD) (OMIM 153400) is a rare, primary lymphoedema of pubertal onset, associated with distichiasis. Causative mutations have now been described in FOXC2, a forkhead transcription factor gene. Numerous clinical associations have been reported with this condition, including congenital heart disease, ptosis, varicose veins, cleft palate, and spinal extradural cysts. SUBJECTS: We report clinical findings in 74 affected subjects from 18 families and six isolated cases. All of them were shown to have mutations in FOXC2 with the exception of one family who had two affected subjects with lymphoedema and distichiasis and linkage consistent with the 16q24 locus. RESULTS: The presence of lymphoedema was highly penetrant. Males had an earlier onset of lymphoedema and a significantly increased risk of complications. Lymphatic imaging confirmed the earlier suggestion that LD is associated with a normal or increased number of lymphatic vessels rather than the hypoplasia or aplasia seen in other forms of primary lymphoedema. Distichiasis was 94.2% penetrant, but not always symptomatic. Associated findings included ptosis (31%), congenital heart disease (6.8%), and cleft palate (4%). Other than distichiasis, the most commonly occurring anomaly was varicose veins of early onset (49%). This has not been previously reported and suggests a possible developmental role for FOXC2 in both venous and lymphatic systems. This is the first gene that has been implicated in the aetiology of varicose veins. CONCLUSION: Unlike previous publications, the thorough clinical characterisation of our patients permits more accurate prediction of various phenotypic abnormalities likely to manifest in subjects with FOXC2 mutations. PMID- 12114479 TI - Novel autosomal dominant mandibulofacial dysostosis with ptosis: clinical description and exclusion of TCOF1. AB - BACKGROUND: Treacher Collins syndrome (TCS), the most common type of mandibulofacial dysostosis (MFD), is genetically homogeneous. Other types of MFD are less common and, of these, only the Bauru type of MFD has an autosomal dominant (AD) mode of inheritance established. Here we report clinical features of a kindred with a unique AD MFD with the exclusion of linkage to the TCS locus (TCOF1) on chromosome 5q31-q32. METHODS: Six affected family members underwent a complete medical genetics physical examination and two affected subjects had skeletal survey. All available medical records were reviewed. Linkage analysis using the markers spanning the TCOF1 locus was performed. One typically affected family member had a high resolution karyotype. RESULTS: Affected subjects had significant craniofacial abnormalities without any significant acral changes and thus had a phenotype consistent with a MFD variant. Distinctive features included hypoplasia of the zygomatic complex, micrognathia with malocclusion, auricular abnormalities with conductive hearing loss, and ptosis. Significantly negative two point lod scores were obtained for markers spanning the TCOF1 locus, excluding the possibility that the disease in our kindred is allelic with TCS. High resolution karyotype was normal. CONCLUSIONS: We report a kindred with a novel type of MFD that is not linked to the TCOF1 locus and is also clinically distinct from other types of AD MFD. Identification of additional families will facilitate identification of the gene causing this type of AD MFD and further characterisation of the clinical phenotype. PMID- 12114481 TI - Linkage stratification and mutation analysis at the Parkin locus identifies mutation positive Parkinson's disease families. PMID- 12114482 TI - Screening of TCOF1 in patients from different populations: confirmation of mutational hot spots and identification of a novel missense mutation that suggests an important functional domain in the protein treacle. PMID- 12114483 TI - Molecular studies in 10 cases of Rubinstein-Taybi syndrome, including a mild variant showing a missense mutation in codon 1175 of CREBBP. PMID- 12114484 TI - Q829X, a novel mutation in the gene encoding otoferlin (OTOF), is frequently found in Spanish patients with prelingual non-syndromic hearing loss. PMID- 12114485 TI - Localisation of the Y chromosome stature gene to a 700 kb interval in close proximity to the centromere. PMID- 12114486 TI - Deletion of 9p associated with gonadal dysfunction in 46,XY but not in 46,XX human fetuses. PMID- 12114487 TI - Maternal uniparental disomy 12 in a healthy girl with a 47,XX,+der(12)(:p11- >q11:)/46,XX karyotype. PMID- 12114488 TI - Predictive testing in the context of pregnancy: experience in Huntington's disease and autosomal dominant cerebellar ataxia. PMID- 12114490 TI - Management of women with a family history of breast cancer in the North West Region of England: training for implementing a vision of the future. PMID- 12114489 TI - Concerns of women presenting to a comprehensive cancer centre for genetic cancer risk assessment. PMID- 12114491 TI - Breakpoint analysis of a familial balanced translocation t(2;8)(q31;p21) associated with mesomelic dysplasia. PMID- 12114492 TI - Novel BRCA2 mutation in a Polish family with hamartoma and two male breast cancers. PMID- 12114493 TI - Detection of large rearrangements of exons 13 and 22 in the BRCA1 gene in German families. PMID- 12114494 TI - Comorbid VHL and SCA2 mutations in a large kindred: confounding diagnosis of neurological dysfunction caused by CNS VHL vascular tumours versus SCA2 atrophic neurodegeneration. PMID- 12114495 TI - Germline mutations in the von Hippel-Lindau (VHL) gene in patients from Poland: disease presentation in patients with deletions of the entire VHL gene. PMID- 12114496 TI - Identification of 13 new mutations in the ACVRL1 gene in a group of 52 unselected Italian patients affected by hereditary haemorrhagic telangiectasia. PMID- 12114497 TI - Alkaptonuria in the Dominican Republic: identification of the founder AKU mutation and further evidence of mutation hot spots in the HGO gene. PMID- 12114498 TI - INSL3/Leydig insulin-like peptide activates the LGR8 receptor important in testis descent. AB - Several orphan G protein-coupled receptors homologous to gonadotropin and thyrotropin receptors have recently been identified and named as LGR4-8. INSL3, also known as Leydig insulin-like peptide or relaxin-like factor, is a relaxin family member expressed in testis Leydig cells and ovarian theca and luteal cells. Male mice mutant for INSL3 exhibit cryptorchidism or defects in testis descent due to abnormal gubernaculum development whereas overexpression of INSL3 induces ovary descent in transgenic females. Because transgenic mice missing the LGR8 gene are also cryptorchid, INSL3 was tested as the ligand for LGR8. Here, we show that treatment with INSL3 stimulated cAMP production in cells expressing recombinant LGR8 but not LGR7. In addition, interactions between INSL3 and LGR8 were demonstrated following ligand receptor cross-linking. Northern blot analysis indicated that the LGR8 transcripts are expressed in gubernaculum whereas treatment of cultured gubernacular cells with INSL3 stimulated cAMP production and thymidine incorporation. The present study identified the ligand for an orphan G protein-coupled receptor based on common phenotypes of ligand and receptor null mice. Demonstration of INSL3 as the ligand for LGR8 facilitates understanding of the mechanism of testis descent and allows studies on the role of INSL3 in gonadal and other physiological processes. PMID- 12114499 TI - HIV-1 Tat interaction with RNA polymerase II C-terminal domain (CTD) and a dynamic association with CDK2 induce CTD phosphorylation and transcription from HIV-1 promoter. AB - Human immunodeficiency virus, type 1 (HIV-1), Tat protein activates viral gene expression through promoting transcriptional elongation by RNA polymerase II (RNAPII). In this process Tat enhances phosphorylation of the C-terminal domain (CTD) of RNAPII by activating cell cycle-dependent kinases (CDKs) associated with general transcription factors of the promoter complex, specifically CDK7 and CDK9. We reported a Tat-associated T-cell-derived kinase, which contained CDK2. Here, we provide further evidence that CDK2 is involved in Tat-mediated CTD phosphorylation and in HIV-1 transcription in vitro. Tat-mediated CTD phosphorylation by CDK2 required cysteine 22 in the activation domain of Tat and amino acids 42-72 of Tat. CDK2 phosphorylated Tat itself, apparently by forming dynamic contacts with amino acids 15-24 and 36-49 of Tat. Also, amino acids 24-36 and 45-72 of Tat interacted with CTD. CDK2 associated with RNAPII and was found in elongation complexes assembled on HIV-1 long-terminal repeat template. Recombinant CDK2/cyclin E stimulated Tat-dependent HIV-1 transcription in reconstituted transcription assay. Immunodepletion of CDK2/cyclin E in HeLa nuclear extract blocked Tat-dependent transcription. We suggest that CDK2 is part of a transcription complex that is required for Tat-dependent transcription and that interaction of Tat with CTD and a dynamic association of Tat with CDK2/cyclin E stimulated CTD phosphorylation by CDK2. PMID- 12114500 TI - L-phenylalanine and NPS R-467 synergistically potentiate the function of the extracellular calcium-sensing receptor through distinct sites. AB - The extracellular calcium (Ca(2+)(o))-sensing receptor (CaR) can be potentiated by allosteric activators including calcimimetics and l-amino acids. In this study, we found that many mutations had differential effects on the functional modulation of the CaR by these two allosteric activators, supporting the idea that these modulators act through distinct sites. 10 mm l-phenylalanine and 1 microm NPS R-467, submaximal doses of the two agents, each elicited similar modulation of R185Q. However, there are different relative potencies for these two modulators with some receptors being more responsive to l-phenylalanine and others being more responsive to NPS R-467. The responsiveness of the CaR to Ca(2+)(o) appears to be essential to observe the potentiating action of l phenylalanine but not of NPS R-467 on the receptor. NPS R-467 reduces the Hill coefficients of the wild-type as well as mutant receptors, suggesting that engagement of all Ca(2+) binding sites is not required when the receptor is activated by NPS R-467. In contrast, l-phenylalanine has little effect on the Hill coefficients of mutant receptors. The two-site model is further supported by the observation that these two classes of modulators exert a synergistic effect on CaRs with inactivating mutations that are responsive to both modulators. PMID- 12114501 TI - A copper-responsive transcription factor, CRF1, mediates copper and cadmium resistance in Yarrowia lipolytica. AB - The dimorphic yeast Yarrowia lipolytica is more resistant to high copper concentrations than Saccharomyces cerevisiae. This differential tolerance to copper ions has been observed in several strains arising from non-related isolates. To investigate the molecular basis of this resistance, we obtained several copper-sensitive mutants. By complementation of one of them, we isolated the YlCRF1 gene encoding for a copper-binding transcription factor of 411 amino acids homologous to ScAce1p, CgAmt1p, and ScMac1p. Naturally occurring copper sensitive strains lack the CRF1 allele. The YlCRF1 transcript is not induced by the addition of copper to the medium. Gene disruption demonstrated that YlCRF1 is responsible for a 4- to 5-fold increase in Y. lipolytica copper tolerance. We further show that strain Deltacrf1 is more sensitive to cadmium but not to other metals. The role of YlCrf1p as a copper-sensitive transcription factor is supported by the finding that the protein is immunolocalized in the nucleus during growth in copper-supplemented but not in copper-free medium. However, in contrast to the S. cerevisiae strain mutated in the metallothionein transcription activator ACE1, Y. lipolytica strain Deltacrf1 is still able to increase metallothionein (MTP) mRNA levels in response to copper addition. CRF1 deletion does not affect superoxide dismutase (SOD) activity either. Our data suggest the existence of one or more different target genes for Crf1p, other than MTP or SOD1, and support its role as a novel copper-responsive transcription factor involved in metal detoxification. PMID- 12114502 TI - Essential role of cAMP-response element-binding protein activation by A2A adenosine receptors in rescuing the nerve growth factor-induced neurite outgrowth impaired by blockage of the MAPK cascade. AB - We found in the present study that stimulation of the A(2A) adenosine receptor (A(2A)-R) using an A(2A)-selective agonist (CGS21680) rescued the blockage of nerve growth factor (NGF)-induced neurite outgrowth when the NGF-evoked MAPK cascade was suppressed by an MEK inhibitor (PD98059) or by a dominant-negative MAPK mutant (dnMAPK). This action of A(2A)-R (designated as the A(2A)-rescue effect) can be blocked by two inhibitors of protein kinase A (PKA) and was absent in a PKA-deficient PC12 variant. Activation of the cAMP/PKA pathway by forskolin exerted the same effect as that by A(2A)-R stimulation. PKA, thus, appears to mediate the A(2A)-rescue effect. Results from cAMP-response element-binding protein (CREB) phosphorylation at serine 133, trans-reporting assays, and overexpression of two dominant-negative CREB mutants revealed that A(2A)-R stimulation led to activation of CREB in a PKA-dependent manner and subsequently reversed the damage of NGF-evoked neurite outgrowth by PD98059 or dnMAPK. Expression of an active mutant of CREB readily rescued the NGF-induced neurite outgrowth impaired by dnMAPK, further strengthening the importance of CREB in the NGF-mediated neurite outgrowth process. Moreover, simultaneous activation of the A(2A)-R/PKA/CREB-mediated and the phosphatidylinositol 3-kinase pathways caused neurite outgrowth that was not suppressed by a selective inhibitor of TrkA, indicating that transactivation of TrkA was not involved. Collectively, CREB functions in conjunction with the phosphatidylinositol 3-kinase pathway to mediate the neurite outgrowth process in PC12 cells. PMID- 12114503 TI - Elevated Akt phosphorylation as an indicator of renal tubular epithelial cell stress. AB - Characterization of the phosphoinositide 3-kinase-signaling pathway in a human renal tubular epithelial cell (TEC) line HKC-8 revealed high levels of Akt phosphorylation in serum-starved cultures. In contrast to Erk1/2, little additional phosphorylation of Akt was observed after cytokine or serum stimulation. Replacement of the conditioned medium attenuated Akt phosphorylation such that 90 min after the addition of warmed serum-free media, Akt phosphorylation had fallen sufficiently to allow an epidermal growth factor stimulated increase to be detected readily. Although the mechanism by which the phosphoinositide 3-kinase/Akt pathway is activated in serum-starved TEC is unknown, the mediator responsible is secreted from these cells. Thus, conditioned media removed from a dish of quiescent TECs stimulated Akt phosphorylation in washed TEC cultures within 10 min. Biochemical characterization of the bioactive agent identified a heat labile factor of small apparent molecular mass. The basal level of Akt phosphorylation observed in serum-starved cultures was inhibited by wortmannin at concentrations that demonstrated its dependence on 3 phosphoinositide synthesis (IC(50) = 8 nm). Regular removal of conditioned media from TEC cultures and its replacement with serum free media resulted in a sustained attenuation of Akt phosphorylation. Interestingly, after 5 days of this treatment, washed TEC cultures contained a greater number of viable cells than cultures maintained in conditioned media throughout. This observation was not explained by a difference in the rate of DNA synthesis. Instead, the number of cells undergoing apoptosis increased markedly in the unwashed cultures. Consequently, we propose that in HKC-8 cells Akt phosphorylation is up-regulated in an effort to minimize cell death. This stress-activated response is initiated by a factor secreted into the conditioned medium that stimulates the phosphoinositide 3-kinase signaling pathway. PMID- 12114504 TI - Matrix metalloproteinases cleave connective tissue growth factor and reactivate angiogenic activity of vascular endothelial growth factor 165. AB - Vascular endothelial growth factor (VEGF), a potent angiogenic mitogen, plays a crucial role in angiogenesis under various pathophysiological conditions. We have recently demonstrated that VEGF(165), one of the VEGF isoforms, binds connective tissue growth factor (CTGF) and that its angiogenic activity is inhibited in the VEGF(165).CTGF complex form (Inoki, I., Shiomi, T., Hashimoto, G., Enomoto, H., Nakamura, H., Makino, K., Ikeda, E., Takata, S., Kobayashi, K. and Okada, Y. (2002) FASEB J. 16, 219-221). In the present study, we further examined the susceptibility of the VEGF(165).CTGF complex to matrix metalloproteinases (MMP-1, -2, -3, -7, -9, and -13), ADAMTS4 (aggrecanase-1), and serine proteinases, and evaluated the recovery of the angiogenic activity of VEGF(165) after the treatment. Among the MMPs, MMP-1, -3, -7, and -13 processed CTGF of the complex into the major NH(2)- and COOH-terminal fragments, whereas VEGF(165) was completely resistant to the MMPs. On the other hand, elastase and plasmin cleaved both CTGF and VEGF(165) of the complex, but they were completely resistant to ADAMTS4. By digestion of the immobilized VEGF(165).CTGF complex with MMP-3 or MMP 7, both NH(2)- and COOH-terminal fragments of CTGF were dissociated and released from the complex into the liquid phase. The in vitro angiogenic activity of VEGF(165) blocked in the VEGF(165).CTGF complex was reactivated to original levels after CTGF digestion of the complex with MMP-1, -3, and -13. Recovery of angiogenic activity was further confirmed by in vivo angiogenesis assay using a Matrigel injection model in mice. These results demonstrate for the first time that CTGF is a substrate of MMPs and that the angiogenic activity of VEGF(165) suppressed by the complex formation with CTGF is recovered through the selective degradation of CTGF by MMPs. MMPs may play a novel role through CTGF degradation in VEGF-induced angiogenesis during embryonic development, tissue maintenance, and/or pathological processes of various diseases. PMID- 12114505 TI - SNAP-25 traffics to the plasma membrane by a syntaxin-independent mechanism. AB - SNAREs (soluble N-ethylmaleimide-sensitive factor attachment protein receptors) are essential for vesicle docking and fusion. SNAP-25, syntaxin 1A, and synaptobrevin/vesicle-associated membrane protein (VAMP) are SNARE proteins that mediate fusion of synaptic vesicles with the plasma membrane. It has been proposed that interactions of SNAP-25 with syntaxin 1A are required for initial membrane attachment of SNAP-25 (Vogel, K., Cabaniols, J.-P., and Roche, P. (2000) J. Biol. Chem. 275, 2959-2965). However, we have shown previously that residues 85-120 of the SNAP-25 interhelical domain, which do not interact with syntaxin, are necessary and sufficient for palmitoylation and plasma membrane localization of a green fluorescent protein reporter molecule (Gonzalo, S., Greentree, W. K., and Linder, M. E. (1999) J. Biol. Chem. 274, 21313-21318). To clarify the role of syntaxin in membrane targeting of SNAP-25, we studied a SNAP-25 point mutant (G43D) that does not interact with syntaxin. SNAP-25 G43D/green fluorescent protein was palmitoylated and localized at the plasma membrane. Newly synthesized SNAP-25 G43D had the same kinetics of membrane association as the wild-type protein. Furthermore, expression of a cytosolic mutant syntaxin 1A did not interfere with SNAP-25 membrane interactions or palmitoylation in the neuronal cell line NG108-15. Exogenously expressed SNAP-25 targets efficiently to the plasma membrane in cells of neuronal origin but only partially in HeLa cells, a neurosecretion-incompetent line. This phenotype was not rescued when syntaxin 1A was co-expressed with SNAP-25. Our data support a syntaxin-independent mechanism of membrane targeting for SNAP-25. PMID- 12114506 TI - Specific orientation and two-dimensional crystallization of the proteasome at metal-chelating lipid interfaces. AB - The potential of a protein-engineered His tag to immobilize macromolecules in a predictable orientation at metal-chelating lipid interfaces was investigated using recombinant 20 S proteasomes His-tagged in various positions. Electron micrographs demonstrated that the orientation of proteasomes bound to chelating lipid films could be controlled via the location of their His tags: proteasomes His-tagged at their sides displayed exclusively side-on views, while proteasomes His-tagged at their ends displayed exclusively end-on views. The activity of proteasomes immobilized at chelating lipid interfaces was well preserved. In solution, His-tagged proteasomes hydrolyzed casein at rates comparable with wild type proteasomes, unless the His tags were located in the vicinity of the N termini of alpha-subunits. The N termini of alpha-subunits might partly occlude the entrance channel in alpha-rings through which substrates enter the proteasome for subsequent degradation. A combination of electron micrographs and atomic force microscope topographs revealed a propensity of vertically oriented proteasomes to crystallize in two dimensions on fluid lipid films. The oriented immobilization of His-tagged proteins at biocompatible lipid interfaces will assist structural studies as well as the investigation of biomolecular interaction via a wide variety of surface-sensitive techniques including single molecule analysis. PMID- 12114507 TI - Trafficking of L-type calcium channels mediated by the postsynaptic scaffolding protein AKAP79. AB - Accurate calcium signaling requires spatial and temporal coordination of voltage gated calcium channels (VGCCs) and a variety of signal transduction proteins. Accordingly, regulation of L-type VGCCs involves the assembly of complexes that include the channel subunits, protein kinase A (PKA), protein kinase A anchoring proteins (AKAPs), and beta2-adrenergic receptors, although the molecular details underlying these interactions remain enigmatic. We show here, by combining extracellular epitope splicing into the channel pore-forming subunit and immunoassays with whole cell and single channel electrophysiological recordings, that AKAP79 directly regulates cell surface expression of L-type calcium channels independently of PKA. This regulation involves a short polyproline sequence contained specifically within the II-III cytoplasmic loop of the channel. Thus we propose a novel mechanism whereby AKAP79 and L-type VGCCs function as components of a biosynthetic mechanism that favors membrane incorporation of organized molecular complexes in a manner that is independent of PKA phosphorylation events. PMID- 12114508 TI - Sequence-specific recognition of collagen triple helices by the collagen-specific molecular chaperone HSP47. AB - HSP47 is a molecular chaperone that plays an unknown role during the assembly and transport of procollagen. Our previous studies showed that, unlike most chaperones, HSP47 interacts with a correctly folded substrate. We suggested that HSP47 either stabilizes the correctly folded collagen helix from heat denaturation or prevents lateral aggregation prior to its transport from the endoplasmic reticulum. In this study we have addressed the role of triple helix stability in the binding of HSP47 to procollagen by expressing procollagen molecules with differing thermal stabilities and analyzing their ability to interact with HSP47 within the endoplasmic reticulum. Our results show that HSP47 interacts with thermostable procollagen molecules, suggesting that helix stabilization is not the primary function of HSP47 and that the interaction of HSP47 with procollagen depends upon the presence of a minimum of one Gly-X-Arg triplet within the triple helical domain. Interestingly, procollagen chains containing high proportions of stabilizing triplets formed triple helices and interacted with HSP47 even in the absence of proline hydroxylation, demonstrating that recognition does not depend upon this modification. Our results support the view that HSP47 functions early in the secretory pathway by preventing the lateral aggregation of procollagen chains. PMID- 12114509 TI - Differential acquisition of antigenic peptides by Hsp70 and Hsc70 under oxidative conditions. AB - Hsp70 and Hsc70 are two chaperones of high homology expressed under contrasting situations. Hsc70 is constitutively expressed and poorly stress-inducible, whereas Hsp70 is unabundant in normal physiological situations and strongly induced under oxidative stress. In the present study we show that the chaperoning activity of purified Hsp70 and Hsc70 is minimal under reducing conditions and increases in environments that mimic oxidative stress. Association with peptides is more pronounced for Hsp70 than for Hsc70 in every condition tested and is accompanied with a gradual change in secondary structure during oxidation. The binding of peptides to Hsp70 and Hsc70 under oxidative conditions is not reversible by treatment with a reducing agent, confirming that other chaperone associated factors are required for substrate release. These findings support the idea that formation of HSP70-peptide complexes and possibly their immunogenicity is enhanced in conditions of stress. PMID- 12114510 TI - Discriminating between the activities of human neutrophil elastase and proteinase 3 using serpin-derived fluorogenic substrates. AB - Human neutrophil elastase (HNE) has long been linked to the pathology of a variety of inflammatory diseases and therefore is a potential target for therapeutic intervention. At least two other serine proteases, proteinase 3 (Pr3) and cathepsin G, are stored within the same neutrophil primary granules as HNE and are released from the cell at the same time at inflammatory sites. HNE and Pr3 are structurally and functionally very similar, and no substrate is currently available that is preferentially cleaved by Pr3 rather than HNE. Discrimination between these two proteases is the first step in elucidating their relative contributions to the development and spread of inflammatory diseases. Therefore, we have prepared new fluorescent peptidyl substrates derived from natural target proteins of the serpin family. This was done because serpins are rapidly cleaved within their reactive site loop whether they act as protease substrates or inhibitors. The hydrolysis of peptide substrates reflects the specificity of the parent serpin including those from alpha-1-protease inhibitor and monocyte neutrophil elastase inhibitor, two potent inhibitors of elastase and Pr3. More specific substrates for these proteases were derived from the reactive site loop of plasminogen activator inhibitor 1, proteinase inhibitors 6 and 9, and from the related viral cytokine response modifier A (CrmA). This improved specificity was obtained by using a cysteinyl residue at P1 for Pr3 and an Ile residue for HNE and because of occupation of protease S' subsites. These substrates enabled us to quantify nanomolar concentrations of HNE and Pr3 that were free in solution or bound at the neutrophil surface. As membrane-bound proteases resist inhibition by endogenous inhibitors, measuring their activity at the surface of neutrophils may be a great help in understanding their role during inflammation. PMID- 12114512 TI - Identification of Reverb(alpha) as a novel ROR(alpha) target gene. AB - The nuclear receptor superfamily comprises a large number of ligand-activated transcription factors that are involved in numerous biological processes such as cell proliferation, differentiation, and homeostasis. ROR(alpha) (NR1F1) and Reverb(alpha) (NR1D1) are two members of this family whose biological functions are largely unknown. In addition, no ligand has been yet identified for these two receptors; therefore, they are referred as orphan receptors. Here, we show that ROR(alpha) and Reverb(alpha) are expressed with a similar tissue distribution and are both induced during the differentiation of rat L6 myoblastic cells. Ectopic expression of ROR(alpha)1 in L6 cells significantly induces Reverb(alpha) expression as demonstrated by Northern blot analysis. Using reverse transcription PCR to analyze Reverb(alpha) gene expression from staggerer mice, we found that there was a significant reduction of Reverb(alpha) mRNA in the skeletal muscle comparing it with the wild-type mice, which suggests that ROR(alpha) is involved in the regulation of Reverb(alpha) gene expression. Transient transfection assays using the Reverb(alpha) promoter demonstrate that ROR(alpha) regulates the Reverb(alpha) gene at the transcriptional level. Furthermore, mutagenesis experiments indicate that ROR(alpha) regulates Reverb(alpha) transcription via a monomeric ROR response element located in the Reverb(alpha) gene promoter. Electrophoretic mobility shift assays show that ROR(alpha) binds strongly to this site in a specific-manner. Finally, overexpression of GRIP-1/TIF-2, but not SRC 1, potentiates ROR(alpha)-stimulated Reverb(alpha) promoter activity in transient transfection experiments. Together, our results identify Reverb(alpha) as a novel target gene for ROR(alpha). PMID- 12114513 TI - The role of cAMP-dependent signaling in receptor-recognized forms of alpha 2 macroglobulin-induced cellular proliferation. AB - Ligation of alpha(2)-macroglobulin receptors by receptor-recognized forms of alpha(2)-macroglobulin (alpha(2)M*) activates various signaling cascades and promotes cell proliferation. It also elevates cAMP in murine peritoneal macrophages. We now report that a significant elevation of cAMP-response element binding protein (CREB) occurs in alpha(2)M*-stimulated cells, and this effect is potentiated by isobutylmethylxanthine, dibutyryl-cAMP, or forskolin. An alpha(2)M* concentration-dependent rapid increase in phosphorylated CREB at Ser(133) also occurred, a necessary event in its activation. Inhibition of Ca(2+)/calmodulin kinase, protein kinases A and C, tyrosine kinases, ribosomal S6 kinase, farnesyl transferase, extracellular signal-regulated kinases 1/2, phosphatidylinositol 3-kinase, or p38 mitogen-activated protein kinase markedly reduce alpha(2)M*-induced phosphorylation of CREB, indicating a role for the p21(ras)-dependent and phosphatidylinositol 3-kinase signaling pathways in regulating CREB activation by alpha(2)M*. Finally, silencing the CREB gene by transfecting cells with a homologous gene sequence double-stranded RNA drastically reduced the expression of CREB and blocked the ability of alpha(2)M* to promote macrophage cell division. We conclude that cAMP-dependent signal transduction as well as other signaling cascades are essential for alpha(2)M* induced cell proliferation. PMID- 12114514 TI - Molecular mechanisms of pre-T cell receptor-induced survival. AB - En route to maturing as T cell receptor (TCR) alphabeta-expressing cells, the development of thymocytes is contingent on expression of a pre-TCR complex comprising a TCRbeta chain paired with a surrogate TCRalpha chain, pre-Talpha (pTalpha). The pre-TCR has been proposed to promote cell survival, proliferation, differentiation, and lineage commitment. However, the precise molecular mechanisms governing this variety of effects remain elusive. Here, we present a cellular system designed to biochemically dissect signals elicited upon pre-TCR expression. Using the T cell line 4G4 stably transfected with one of the two known pTalpha isoforms or selective pTalpha deletion mutants and TCRbeta, we were able to observe that expression of a functional pre-TCR complex is sufficient to control the levels of surface Fas protein, the stimulation of mitogen-activated and stress-regulated kinases, and the activation status of the p53 antioncogene. We demonstrate that this regulation has a major impact on the expression of important regulators of apoptosis, such as Bcl-2 family members, and the cell cycle, such as p21(WAF). Furthermore, we show here that cells expressing a functional pre-TCR are more resistant to different types of DNA damage-induced apoptosis and that these effects are contingent on an intact cytoplasmic tail of pTalpha. We finally propose that the presence of a functional pre-TCR complex triggers many intracellular pathways capable of driving and ensuring thymocyte survival in the presence of DNA damage. PMID- 12114511 TI - Activation of protein kinase C beta II by the stereo-specific phosphatidylserine receptor is required for phagocytosis of apoptotic thymocytes by resident murine tissue macrophages. AB - We showed previously that protein kinase C (PKC) is required for phagocytosis of apoptotic leukocytes by murine alveolar (AMo) and peritoneal macrophages (PMo) and that such phagocytosis is markedly lower in AMo compared with PMo. In this study, we examined the roles of individual PKC isoforms in phagocytosis of apoptotic thymocytes by these two Mo populations. By immunoblotting, AMo expressed equivalent PKC eta but lower amounts of other isoforms (alpha, betaI, betaII, delta, epsilon, mu, and zeta), with the greatest difference in betaII expression. A requirement for PKC betaII for phagocytosis was demonstrated collectively by phorbol 12-myristate 13-acetate-induced depletion of PKC betaII, by dose-response to PKC inhibitor Ro-32-0432, and by use of PKC betaII myristoylated peptide as a blocker. Exposure of PMo to phosphatidylserine (PS) liposomes specifically induced translocation of PKC betaII and other isoforms to membranes and cytoskeleton. Both AMo and PMo expressed functional PS receptor, blockade of which inhibited PKC betaII translocation. Our results indicate that murine tissue Mo require PKC betaII for phagocytosis of apoptotic cells, which differs from the PKC isoform requirement previously described in Mo phagocytosis of other particles, and imply that a crucial action of the PS receptor in this process is PKC betaII activation. PMID- 12114516 TI - Crystal structure of the F87W/Y96F/V247L mutant of cytochrome P-450cam with 1,3,5 trichlorobenzene bound and further protein engineering for the oxidation of pentachlorobenzene and hexachlorobenzene. AB - We reported previously that the F87W/Y96F/V247L mutant of cytochrome P-450cam (CYP101) from Pseudomonas putida catalyzed the rapid oxidation of lightly chlorinated benzenes, but pentachlorobenzene oxidation was slow (Jones, J. P., O'Hare, E. J., and Wong, L. L. (2001) Eur. J. Biochem. 268, 1460-1467). In the present work, we determined the crystal structure of this mutant with bound 1,3,5 trichlorobenzene. The substrate was bound to crystallographically independent CYP101 molecules in at least three different orientations, which were distinguished by the angle between the benzene ring and the porphyrin, and one orientation contained an Fe-Cl interaction. In another orientation, the substrate was almost parallel to the heme, with a C-H bond closest to the iron. The enzyme/substrate contacts suggested that the L244A mutation should promote the binding of pentachlorobenzene and hexachlorobenzene by creating space to accommodate the extra chlorines. The F87W/Y96F/L244A/V247L mutant thus designed was found to oxidize pentachlorobenzene at a rate of 82.5 nmol (nmol CYP101)(-1) min(-1), 45 times faster than the F87W/Y96F/V247L parent mutant. The rate of hexachlorobenzene oxidation was increased 200-fold, to 2.0 min(-1). Both substrates are oxidized to pentachlorophenol, which is degraded by micro organisms. In principle, the F87W/Y96F/L244A/V247L mutant could have applications in the bioremediation of polychlorinated benzenes. PMID- 12114515 TI - Isolation and functional analysis of the mouse RXRgamma1 gene promoter in anterior pituitary cells. AB - The retinoid X receptor (RXR) isoform RXRgamma has limited tissue expression, including brain, skeletal muscle, and anterior pituitary gland. Within the anterior pituitary gland, RXRgamma expression is limited primarily to the thyrotropes. In this report, we have isolated approximately 3 kb of 5'-flanking DNA of the mouse RXRgamma1 gene. We have identified the major transcription start site in the thyrotrope-derived TtT-97 cells. Transient transfection studies show that a 1.4-kb promoter fragment has full promoter activity in TtT-97 cells. This promoter has much less activity in thyrotrope-derived alphaTSH cells, pituitary derived GH3 somatomammotropes, and non-pituitary CV-1 cells. None of these cell lines has detectable RXRgamma1 mRNA. A previous report has identified a non consensus direct repeat (DR-1) element in the RXRgamma2 gene promoter region that mediates stimulation of promoter activity by 9-cis-retinoic acid (9-cis-RA). Inspection of the RXRgamma1 promoter region revealed a non-consensus DR-1 element at -232 bp from the transcription start site. Interestingly, RXRgamma1 promoter activity was suppressed 50% by 9-cis-RA in the TtT-97 thyrotropes. Further experiments in non-pituitary cells showed that suppression of RXRgamma1 promoter activity was RXR-dependent. Mutagenesis of the DR-1 element abrogated suppression of promoter activity by 9-cis-RA, suggesting that this negative regulation requires both RXR and this specific DR-1 element. In summary, we have isolated the mouse RXRgamma1 gene promoter region and identified the major start site in thyrotropes. Promoter activity is uniquely suppressed by 9-cis-RA through a DR-1 element. Isolation and characterization of the mouse RXRgamma1 promoter region provides a tool for further investigation focusing on thyrotrope-specific gene expression as well as negative regulation of genes by retinoic acid. PMID- 12114518 TI - Modulation of a brain voltage-gated K+ channel by syntaxin 1A requires the physical interaction of Gbetagamma with the channel. AB - Recently we suggested that direct interactions between voltage-gated K(+) channels and proteins of the exocytotic machinery, such as those observed between the Kv1.1/Kvbeta channel, syntaxin 1A, and SNAP-25 may be involved in neurotransmitter release. Furthermore, we demonstrated that the direct interaction with syntaxin 1A enhances the fast inactivation of Kv1.1/Kvbeta1.1 in oocytes. Here we show that G-protein betagamma subunits play a crucial role in the enhancement of inactivation by syntaxin 1A. The effect caused by overexpression of syntaxin 1A is eliminated in the presence of chelators of endogenous betagamma subunits in the whole cell and at the plasma membrane. Conversely, enhancement of inactivation caused by overexpression of beta(1)gamma(2) subunits is eliminated upon knock-down of endogenous syntaxin or its scavenging at the plasma membrane. We further show that the N terminus of Kv1.1 binds brain synaptosomal and recombinant syntaxin 1A and concomitantly binds beta(1)gamma(2); the binding of beta(1)gamma(2) enhances that of syntaxin 1A. Taken together, we suggest a mechanism whereby syntaxin and G protein betagamma subunits interact concomitantly with a Kv channel to regulate its inactivation. PMID- 12114517 TI - Leptin promotes biliary cholesterol elimination during weight loss in ob/ob mice by regulating the enterohepatic circulation of bile salts. AB - Leptin administration to obese C57BL/6J (ob/ob) mice results in weight loss by reducing body fat. Because adipose tissue is an important storage depot for cholesterol, we explored evidence that leptin-induced weight loss in ob/ob mice was accompanied by transport of cholesterol to the liver and its elimination via bile. Consistent with mobilization of stored cholesterol, cholesterol concentrations in adipose tissue remained unchanged during weight loss. Plasma cholesterol levels fell sharply, and microscopic analyses of gallbladder bile revealed cholesterol crystals as well as cholesterol gallstones. Surprisingly, leptin reduced biliary cholesterol secretion rates without affecting secretion rates of bile salts or phospholipids. Instead, cholesterol supersaturation of gallbladder bile was due to marked decreases in bile salt hydrophobicity and not to hypersecretion of biliary cholesterol per se, such as occurs in humans during weight loss. In addition to regulating bile salt composition, leptin treatment decreased bile salt pool size. The smaller, more hydrophilic bile salt pool was associated with substantial decreases in intestinal cholesterol absorption. Within the liver, leptin treatment reduced the activity of 3-hydroxy-3 methylglutaryl-CoA reductase, but it did not change activities of cholesterol 7alpha-hydroxylase or acyl-CoA:cholesterol acyltransferase. These data suggest that leptin regulates biliary lipid metabolism to promote efficient elimination of excess cholesterol stored in adipose tissue. Cholesterol gallstone formation during weight loss in ob/ob mice appears to represent a pathologic consequence of an adaptive response that prevents absorption of biliary and dietary cholesterol. PMID- 12114519 TI - Selective effects of calcium chelators on anterograde and retrograde protein transport in the cell. AB - Calcium plays a regulatory role in several aspects of protein trafficking in the cell. Both vesicle fusion and vesicle formation can be inhibited by the addition of calcium chelators. Because the effects of calcium chelators have been studied predominantly in cell-free systems, it is not clear exactly which transport steps in the secretory pathway are sensitive to calcium levels. In this regard, we have studied the effects of calcium chelators on both anterograde and retrograde protein transport in whole cells. Using both cytochemical and biochemical analyses, we find that the anterograde-directed exit of vesicular stomatitis virus G protein and the retrograde-directed exit of Shiga toxin from the Golgi apparatus are both inhibited by calcium chelation. The exit of vesicular stomatitis virus G from a pre-Golgi compartment and the exit of Shiga toxin from an endosomal compartment are sensitive to the membrane-permeant calcium chelator 1,2-bis(2-amino phenoxy)ethane-N,N,N',N'-tetraacetic acid-tetrakis (acetoxymethyl ester) (BAPTA-AM). By contrast, endoplasmic reticulum exit and endocytic internalization from the plasma membrane are not affected by BAPTA. Together, our data show that some, but not all, trafficking steps in the cell may be regulated by calcium. These studies provide a framework for a more detailed analysis of the role of calcium as a regulatory agent during protein transport. PMID- 12114521 TI - The inhibitory function in human progesterone receptor N termini binds SUMO-1 protein to regulate autoinhibition and transrepression. AB - Although most studies of progesterone receptors (PR) and their two isoforms, PR-A and PR-B, focus on transcriptional stimulation, the receptors exhibit important inhibitory properties. Autoinhibition refers to an inhibitory function located in the PR N terminus, whose deletion increases transcriptional activity at least 6 10-fold. Transrepression refers to the ability of PR-A to suppress the transcriptional activity of PR-B and other nuclear receptors, including estrogen receptors. Self-squelching refers to the observation in transient transfection assays that increasing receptor concentrations paradoxically decrease transcriptional activity. Using a series of N-terminal deletion mutants constructed in both PR isoforms, we have mapped their autoinhibitory and transrepressor activities to a small ubiquitin-like modifier (SUMO-1) protein consensus-binding motif, (387)IKEE, located in the N terminus upstream of AF1. Self-squelching does not involve this site. SUMO-1 binds PR covalently at (387)IKEE, but only if the C-terminal, liganded, hormone-binding domain is also present. A single point K388R mutation within the (387)IKEE motif in either PR-A or PR-B leads to a loss of autoinhibitory and transrepressor functions of the liganded, full-length receptors. We conclude that autoinhibition and transrepression involve N-terminal sumoylation combined with intramolecular N/C terminal communication. PMID- 12114520 TI - The N-terminal domains of syntaxin 7 and vti1b form three-helix bundles that differ in their ability to regulate SNARE complex assembly. AB - The SNAREs syntaxin 7, syntaxin 8, vti1b, and endobrevin/VAMP8 function in the fusion of late endosomes. Although the core complex formed by these SNAREs is very similar to the neuronal SNARE complex, it differs from the neuronal complex in that three of the four SNAREs contain extended N-terminal regions of unknown structure and function. Here we show that the N-terminal regions of syntaxin 7, syntaxin 8, and vti1b contain well folded alpha-helical domains. Multidimensional NMR spectroscopy revealed that in syntaxin 7 and vti1b, the domains form three helix bundles resembling those of syntaxin 1, Sso1p, and Vam3p. The three-helix bundle domain of vti1b is the first of its kind identified in a SNARE outside the syntaxin family. Only syntaxin 7 adopts a closed conformation, whereas in vti1b and syntaxin 8, the N-terminal domains do not interact with the adjacent SNARE motifs. Accordingly, the rate of SNARE complex assembly is retarded about 7-fold when syntaxin 7 contains its N-terminal domain, whereas the N-terminal domains of vti1b and syntaxin 8 have no influence on assembly kinetics. We conclude that three-helix bundles represent a common fold for SNARE N-terminal domains, not restricted to the syntaxin family. However, they differ in their ability to adopt closed conformations and thus to regulate the assembly of SNARE complexes. PMID- 12114522 TI - Involvement of an upstream stimulatory factor as well as cAMP-responsive element binding protein in the activation of brain-derived neurotrophic factor gene promoter I. AB - The use of different brain-derived neurotrophic factor (BDNF) gene promoters results in the differential production of 5'-alternative transcripts, suggesting versatile functions of BDNF in neurons. Among four BDNF promoters I, II, III, and IV (BDNF-PI, -PII, -PIII, and -PIV), BDNF-PI was markedly activated, as well as BDNF-PIII, by Ca(2+) signals evoked via neuronal activity. However, little is known about the mechanisms for the transcriptional activation of BDNF-PI. Using rat cortical neurons in culture, we assigned the promoter sequences responsible for the Ca(2+) signal-mediated activation of BDNF-PI and found that the Ca(2+) responsive elements were located in two separate (distal and proximal) regions and that the DNA sequences in the proximal region containing cAMP-responsive element (CRE), which is overlapped by the upstream stimulatory factor (USF) binding element, were largely responsible for the activation of BDNF-PI. CRE binding protein (CREB) family transcription factors and USF1/USF2 bind to this overlapping site, depending upon their preferred sequences which also control the magnitude of the activation. Overexpression of dominant negative CREB or USF reduced the BDNF-PI activation. These findings support that not only CREB but also USF1/USF2 contributes to Ca(2+) signal-mediated activation of BDNF-PI through the recognition of an overlapping CRE and USF-binding element. PMID- 12114523 TI - Analysis of the regulation of the A33 antigen gene reveals intestine-specific mechanisms of gene expression. AB - The A33 antigen is a transmembrane protein expressed almost exclusively by intestinal epithelial cells. The level of its expression is robust and uniform throughout the rostrocaudal axis of the human and mouse intestines. In the colon, strong expression is found in the basolateral membranes of both the proliferating cells in the lower regions of the crypts and the differentiating cells in the upper regions of crypts. Similarly, in the small intestine, the protein is highly expressed by all the epithelial cells in the crypts and by the differentiated cells migrating over the villi. Thus, the A33 antigen has emerged as a definitive marker for all intestinal epithelial cells, irrespective of cell lineage and differentiation status. To understand the molecular mechanisms mediating this rare tissue-specific expression pattern, we undertook a comprehensive analysis of the 5'-regulatory region of the human A33 antigen gene. This allowed us to point to positive cis-regulatory elements incorporating consensus Kruppel-like factor and caudal-related homeobox (CDX)-binding sites, located just upstream from the human A33 antigen transcription start site, as being important for the intestine specific expression pattern of this gene. Further analysis provided evidence that the A33 antigen gene may be one of only a few target genes to be described thus far for the intestine-specific homeobox transcription factor, CDX1. Taken together, our data lead us to propose that the activity of CDX1 is pivotal in mediating the exquisite, intestine-specific expression pattern of the A33 antigen gene. PMID- 12114524 TI - Alpha-conotoxin GIC from Conus geographus, a novel peptide antagonist of nicotinic acetylcholine receptors. AB - Many venomous organisms produce toxins that disrupt neuromuscular communication to paralyze their prey. One common class of such toxins comprises nicotinic acetylcholine receptor antagonists (nAChRs). Thus, most toxins that act on nAChRs are targeted to the neuromuscular subtype. The toxin characterized in this report, alpha-conotoxin GIC, is a most striking exception. The 16-amino acid peptide was identified from a genomic DNA clone from Conus geographus. The predicted mature toxin was synthesized, and synthetic toxin was used in all studies described. alpha-Conotoxin GIC shows no paralytic activity in fish or mice. Furthermore, even at concentrations up to 100 microm, the peptide has no detectable effect on the human muscle nicotinic receptor subtype heterologously expressed in Xenopus oocytes. In contrast, the toxin has high affinity (IC(50) approximately 1.1 nm) for the human alpha3beta2 subunit combination, making it the most neuronally selective nicotinic antagonist characterized thus far. Although alpha-conotoxin GIC shares some sequence similarity with alpha-conotoxin MII, which is also a potent alpha3beta2 nicotinic antagonist, it is much less hydrophobic, and the kinetics of channel block are substantially different. It is noteworthy that the nicotinic ligands in C. geographus venom fit an emerging pattern in venomous predators, with one nicotinic antagonist targeted to the muscle subtype (thereby causing paralysis) and a second nicotinic antagonist targeted to the alpha3beta2 nAChR subtype (possibly inhibiting the fight-or flight response). PMID- 12114526 TI - Oxygen-independent coproporphyrinogen-III oxidase HemN from Escherichia coli. AB - In bacteria the oxygen-independent coproporphyrinogen-III oxidase catalyzes the oxygen-independent conversion of coproporphyrinogen-III to protoporphyrinogen-IX. The Escherichia coli hemN gene encoding a putative part of this enzyme was overexpressed in E. coli. Anaerobically purified HemN is a monomeric protein with a native M(r) = 52,000 +/- 5,000. A newly established anaerobic enzyme assay was used to demonstrate for the first time in vitro coproporphyrinogen-III oxidase activity for recombinant purified HemN. The enzyme requires S-adenosyl-l methionine (SAM), NAD(P)H, and additional cytoplasmatic components for catalysis. An oxygen-sensitive iron-sulfur cluster was identified by absorption spectroscopy and iron analysis. Cysteine residues Cys(62), Cys(66), and Cys(69), which are part of the conserved CXXXCXXC motif found in all HemN proteins, are essential for iron-sulfur cluster formation and enzyme function. Completely conserved residues Tyr(56) and His(58), localized closely to the cysteine-rich motif, were found to be important for iron-sulfur cluster integrity. Mutation of Gly(111) and Gly(113), which are part of the potential GGGTP S-adenosyl-l-methionine binding motif, completely abolished enzymatic function. Observed functional properties in combination with a recently published computer-based enzyme classification (Sofia, H. J., Chen, G., Hetzler, B. G., Reyes-Spindola, J. F., and Miller, N. E. (2001) Nucleic Acids Res. 29, 1097-1106) identifies HemN as "Radical SAM enzyme." An appropriate enzymatic mechanism is suggested. PMID- 12114525 TI - Inhibitory cross-talk between estrogen receptor (ER) and constitutively activated androstane receptor (CAR). CAR inhibits ER-mediated signaling pathway by squelching p160 coactivators. AB - Estrogen receptor (ER) activity can be modulated by the action of other nuclear receptors. To study whether ER activity is altered by orphan nuclear receptors that mediate the cellular response to xenobiotics, cross-talk between ER and constitutive androstane receptor (CAR), steroid and xenobiotic receptor, or peroxisome proliferator-activated receptor gamma was examined in HepG2 cells. Of these receptors, CAR substantially inhibited ER-mediated transcriptional activity of the vitellogenin B1 promoter as well as a synthetic estrogen responsive element (ERE)-containing promoter. Treatment with an agonist of CAR, 1,4-bis-(2 (3,5-dichloropyridoxyl))benzene, potentiated CAR-mediated transcriptional repression. In contrast, an antagonist of CAR, androstenol, alleviated the repression effect. Although CAR interacted with the ER in solution, CAR did not interact with the ER bound to the ERE. CAR/retinoid X receptor bound to the ERE but with much lower affinity than ER. Incremental amounts of CAR elicited a progressive reduction of the ER activity induced by the p160 coactivator glucocorticoid receptor interacting protein 1 (GRIP-1). In turn, increasing amounts of GRIP-1 progressively reversed the depression of ER activity by CAR. An agonist or antagonist of CAR potentiated or alleviated, respectively, the CAR mediated repression of the GRIP-1-enhanced ER activity, which is consistent with the ability of theses ligands to increase or decrease, respectively, the interaction of CAR with GRIP-1. A CAR mutant that did not interact with GRIP-1 did not inhibit ER-mediated transactivation. Our data demonstrate that xenobiotic nuclear receptor CAR antagonizes ER-mediated transcriptional activity by squelching limiting amounts of p160 coactivator and imply that xenobiotics may influence ER function of female reproductive physiology, cell differentiation, tumorigenesis, and lipid metabolism. PMID- 12114527 TI - Dynamics of chromophore binding to Lhc proteins in vivo and in vitro during operation of the xanthophyll cycle. AB - Three plant xanthophylls are components of the xanthophyll cycle in which, upon exposure of leaves to high light, the enzyme violaxanthin de-epoxidase (VDE) transforms violaxanthin into zeaxanthin via the intermediate antheraxanthin. Previous work () showed that xanthophylls are bound to Lhc proteins and that substitution of violaxanthin with zeaxanthin induces conformational changes and fluorescence quenching by thermal dissipation. We have analyzed the efficiency of different Lhc proteins to exchange violaxanthin with zeaxanthin both in vivo and in vitro. Light stress of Zea mays leaves activates VDE, and the newly formed zeaxanthin is found primarily in CP26 and CP24, whereas other Lhc proteins show a lower exchange capacity. The de-epoxidation system has been reconstituted in vitro by using recombinant Lhc proteins, recombinant VDE, and monogalactosyl diacylglycerol (MGDG) to determine the intrinsic capacity for violaxanthin-to zeaxanthin exchange of individual Lhc gene products. Again, CP26 was the most efficient in xanthophyll exchange. Biochemical and spectroscopic analysis of individual Lhc proteins after de-epoxidation in vitro showed that xanthophyll exchange occurs at the L2-binding site. Xanthophyll exchange depends on low pH, implying that access to the binding site is controlled by a conformational change via lumenal pH. These findings suggest that the xanthophyll cycle participates in a signal transduction system acting in the modulation of light harvesting versus thermal dissipation in the antenna system of higher plants. PMID- 12114528 TI - Chemical synthesis of poliovirus cDNA: generation of infectious virus in the absence of natural template. AB - Full-length poliovirus complementary DNA (cDNA) was synthesized by assembling oligonucleotides of plus and minus strand polarity. The synthetic poliovirus cDNA was transcribed by RNA polymerase into viral RNA, which translated and replicated in a cell-free extract, resulting in the de novo synthesis of infectious poliovirus. Experiments in tissue culture using neutralizing antibodies and CD155 receptor-specific antibodies and neurovirulence tests in CD155 transgenic mice confirmed that the synthetic virus had biochemical and pathogenic characteristics of poliovirus. Our results show that it is possible to synthesize an infectious agent by in vitro chemical-biochemical means solely by following instructions from a written sequence. PMID- 12114529 TI - Predictive identification of exonic splicing enhancers in human genes. AB - Specific short oligonucleotide sequences that enhance pre-mRNA splicing when present in exons, termed exonic splicing enhancers (ESEs), play important roles in constitutive and alternative splicing. A computational method, RESCUE-ESE, was developed that predicts which sequences have ESE activity by statistical analysis of exon-intron and splice site composition. When large data sets of human gene sequences were used, this method identified 10 predicted ESE motifs. Representatives of all 10 motifs were found to display enhancer activity in vivo, whereas point mutants of these sequences exhibited sharply reduced activity. The motifs identified enable prediction of the splicing phenotypes of exonic mutations in human genes. PMID- 12114530 TI - Steering attosecond electron wave packets with light. AB - Photoelectrons excited by extreme ultraviolet or x-ray photons in the presence of a strong laser field generally suffer a spread of their energies due to the absorption and emission of laser photons. We demonstrate that if the emitted electron wave packet is temporally confined to a small fraction of the oscillation period of the interacting light wave, its energy spectrum can be up- or downshifted by many times the laser photon energy without substantial broadening. The light wave can accelerate or decelerate the electron's drift velocity, i.e., steer the electron wave packet like a classical particle. This capability strictly relies on a sub-femtosecond duration of the ionizing x-ray pulse and on its timing to the phase of the light wave with a similar accuracy, offering a simple and potentially single-shot diagnostic tool for attosecond pump probe spectroscopy. PMID- 12114531 TI - Dynamical model of long-term synaptic plasticity. AB - Long-term synaptic plasticity leading to enhancement in synaptic efficacy (long term potentiation, LTP) or decrease in synaptic efficacy (long-term depression, LTD) is widely regarded as underlying learning and memory in nervous systems. LTP and LTD at excitatory neuronal synapses are observed to be induced by precise timing of pre- and postsynaptic events. Modification of synaptic transmission in long-term plasticity is a complex process involving many pathways; for example, it is also known that both forms of synaptic plasticity can be induced by various time courses of Ca(2+) introduction into the postsynaptic cell. We present a phenomenological description of a two-component process for synaptic plasticity. Our dynamical model reproduces the spike time-dependent plasticity of excitatory synapses as a function of relative timing between pre- and postsynaptic events, as observed in recent experiments. The model accounts for LTP and LTD when the postsynaptic cell is voltage clamped and depolarized (LTP) or hyperpolarized (LTD) and no postsynaptic action potentials are evoked. We are also able to connect our model with the Bienenstock, Cooper, and Munro rule. We give model predictions for changes in synaptic strength when periodic spike trains of varying frequency and Poisson distributed spike trains with varying average frequency are presented pre- and postsynaptically. When the frequency of spike presentation exceeds approximately 30-40 Hz, only LTP is induced. PMID- 12114532 TI - Characterization of the expression of MHC proteins in human embryonic stem cells. AB - Human embryonic stem (ES) cells are pluripotent cells that may be used in transplantation medicine. These cells can be induced to differentiate into cells from the three embryonic germ layers both in vivo and in vitro. To determine whether human ES cells might be rejected after transplantation, we examined cell surface expression of the MHC proteins in these cells. Our results show very low expression levels of MHC class I (MHC-I) proteins on the surface of human ES cells that moderately increase on in vitro or in vivo differentiation. A dramatic induction of MHC-I proteins was observed when the cells were treated with IFN gamma but not with IFN-alpha or -beta. However, all three IFNs induced expression of MHC-I proteins in differentiated human ES cells. MHC-II proteins and HLA-G were not expressed on the surface of undifferentiated or differentiated cells. Ligands for natural killer cell receptors were either absent or expressed in very low levels in human ES cells and in their differentiated derivatives. In accordance, natural killer cytotoxic assays demonstrated only limited lysis of both undifferentiated and differentiated cells. To initiate a histocompatibility databank of human ES cells, we have isotyped several of the published ES cell lines for their human leukocyte antigens. In conclusion, our results demonstrate that human ES cells can express high levels of MHC-I proteins and thus may be rejected on transplantation. PMID- 12114533 TI - Modulating influence on HIV/AIDS by interacting RANTES gene variants. AB - RANTES (regulated on activation normal T cell expressed and secreted), a ligand for the CC chemokine receptor 5, potently inhibits HIV-1 replication in vitro. We tested the influence of four RANTES single nucleotide polymorphism (SNP) variants and their haplotypes on HIV-1 infection and AIDS progression in five AIDS cohorts. Three SNPs in the RANTES gene region on chromosome 17 (403A in the promoter, In1.1C in the first intron, and 3'222C in the 3' untranslated region) are associated with increased frequency of HIV-1 infection. The common In1.1C SNP allele is nested within an intronic regulatory sequence element that exhibits differential allele binding to nuclear proteins and a down-regulation of gene transcription. The In1.1C allele or haplotypes that include In1.1C display a strong dominant association with rapid progression to AIDS among HIV-1-infected individuals in African-American, European-American, and combined cohorts. The principal RANTES SNP genetic influence on AIDS progression derives from the down regulating RANTES In1.1C allele, although linkage disequilibrium with adjoining RANTES SNPs including a weaker up-regulating RANTES promoter allele (-28G), can modify the observed epidemiological patterns. The In1.1C-bearing genotypes account for 37% of the attributable risk for rapid progression among African Americans and may also be an important influence on AIDS progression in Africa. The diminished transcription of RANTES afforded by the In1.1C regulatory allele is consistent with increased HIV-1 spread in vivo, leading to accelerated progression to AIDS. PMID- 12114534 TI - A 76-kb duplicon maps close to the BCR gene on chromosome 22 and the ABL gene on chromosome 9: possible involvement in the genesis of the Philadelphia chromosome translocation. AB - A patient with a typical form of chronic myeloid leukemia was found to carry a large deletion on the derivative chromosome 9q+ and an unusual BCR-ABL transcript characterized by the insertion, between BCR exon 14 and ABL exon 2, of 126 bp derived from a region located on chromosome 9, 1.4 Mb 5' to ABL. This sequence was contained in the bacterial artificial chromosome RP11-65J3, which in fluorescence in situ hybridization experiments on normal metaphases was found to detect, in addition to the predicted clear signal at 9q34, a faint but distinct signal at 22q11.2, where the BCR gene is located, suggesting the presence of a large region of homology between the two chromosomal regions. Indeed, blast analysis of the RP11-65J3 sequence against the entire human genome revealed the presence of a stretch of homology, about 76 kb long, located approximately 150 kb 3' to the BCR gene, and containing the 126-bp insertion sequence. Evolutionary studies using fluorescence in situ hybridization identified the region as a duplicon, which transposed from the region orthologous to human 9q34 to chromosome 22 after the divergence of orangutan from the human-chimpanzee-gorilla common ancestor about 14 million years ago. Recent sequence analyses have disclosed an unpredicted extensive segmental duplication of our genome, and the impact of duplicons in triggering genomic disorders is becoming more and more apparent. The discovery of a large duplicon relatively close to the ABL and BCR genes and the finding that the 126-bp insertion is very close to the duplicon at 9q34 open the question of the possible involvement of the duplicon in the formation of the Philadelphia chromosome translocation. PMID- 12114536 TI - Heterogeneous nucleation of supercooled water, and the effect of an added catalyst. AB - The statistics of liquid-to-crystal nucleation are measured rigorously by using a recently developed automated lag-time apparatus (ALTA). A single sample, in this case a sample of pure water both with and without an (insoluble) AgI crystal, is repeatedly cooled, nucleated, and thawed. Analysis of the data, coupled with a second kind of experiment, shows that the statistics of nucleation are consistent with a first-order kinetic mechanism over a wide range of supercooling temperatures. The limitations of classical nucleation theory are exhibited. Our analysis unifies many related experiments in biology, physics, chemistry, and chemical engineering. PMID- 12114535 TI - Identification of two cerebral malaria resistance loci using an inbred wild derived mouse strain. AB - Malaria is a complex infectious disease in which the host/parasite interaction is strongly influenced by host genetic factors. The consequences of plasmodial infections range from asymptomatic to severe complications like the neurological syndrome cerebral malaria induced by Plasmodium falciparum in humans and Plasmodium berghei ANKA in rodents. Mice infected with P. berghei ANKA show marked differences in disease manifestation and either die from experimental cerebral malaria (ECM) or from hemolytic anemia caused by hyperparasitemia (HP). A majority of laboratory mouse strains so far investigated are susceptible to ECM; however, a number of wild-derived inbred strains show resistance. To evaluate the genetic basis of this difference, we crossed a uniquely ECM resistant, wild-derived inbred strain (WLA) with an ECM susceptible laboratory strain (C57BL/6J). All of the (WLA x C57BL/6J) F(1) and 97% of the F(2) progeny displayed ECM resistance similar to the WLA strain. To screen for loci contributing to ECM resistance, we analyzed a cohort of mice backcrossed to the C57BL/6J parental strain. A genome wide screening of this cohort provided significant linkage of ECM resistance to marker loci in two genetic regions on chromosome 1 (chi(2) = 18.98, P = 1.3 x 10(-5)) and on chromosome 11 (chi(2) = 16.51, P = 4.8 x 10(-5)), being designated Berr1 and Berr2, respectively. These data provide the first evidence of loci associated with resistance to murine cerebral malaria, which may have important implications for the search for genetic factors controlling cerebral malaria in humans. PMID- 12114538 TI - Spatial patterns in ant colonies. AB - The origins of large-scale spatial patterns in biology have been an important source of theoretical speculation since the pioneering work by Turing (1952) on the chemical basis of morphogenesis. Knowing how these patterns emerge and their functional role is important to our understanding of the evolution of biocomplexity and the role played by self organization. However, so far, conclusive evidence for local activation-long-range inhibition mechanisms in real biological systems has been elusive. Here a well-defined experimental and theoretical analysis of the pattern formation dynamics exhibited by clustering behavior in ant colonies is presented. These experiments and a simple mathematical model show that these colonies do indeed use this type of mechanism. All microscopic variables have been measured and provide the first evidence, to our knowledge, for this type of self-organized behavior in complex biological systems, supporting early conjectures about its role in the organization of insect societies. PMID- 12114537 TI - Retrograde trans-synaptic transfer of green fluorescent protein allows the genetic mapping of neuronal circuits in transgenic mice. AB - The function of the nervous system is a consequence of the intricate synaptic connectivity of its neurons. Our understanding of these highly complex networks has profited enormously from methods used over the past two decades that are based on the mechanical injection of tracer molecules into brain regions. We have developed a genetic system for the mapping of synaptic connections during development of the mammalian central nervous system and in the mature brain. It is based on the transsynaptic transfer of green fluorescent protein (GFP) in the brains of mice using a fusion protein with a nontoxic fragment of tetanus toxin (TTC) expressed in defined neurons. These transgenic mice allowed us to visualize neurons, at single-cell resolution, that are in synaptic contact by the detection of GFP in interconnected circuits. Targeted genetic expression with a specific promoter permitted us to transfer GFP to defined subsets of neurons and brain regions. GFP-TTC is coexpressed with a lacZ reporter gene to discriminate neurons that produce the tracer from cells that have acquired it transneuronally. The marker shows selective transfer in the retrograde direction. We have used electron microscopic detection of GFP to define the ultrastructural features of the system. Our work opens up a range of possibilities for brain slice and in vivo studies taking advantage of the fluorescence of GFP. We point the way toward the use of powerful multiphoton technology and set the stage for the transsynaptic transfer of other proteins in the brains of mice. PMID- 12114539 TI - mda-7 (IL-24) Mediates selective apoptosis in human melanoma cells by inducing the coordinated overexpression of the GADD family of genes by means of p38 MAPK. AB - Subtraction hybridization identified melanoma differentiation-associated gene-7 (mda-7) as a gene induced during terminal differentiation in human melanoma cells. On the basis of structure, chromosomal localization and cytokine-like properties, mda-7 is classified as IL-24. Administration of mda-7/IL-24 by means of a replication-incompetent adenovirus (Ad.mda-7) induces apoptosis selectively in diverse human cancer cells without inducing harmful effects in normal fibroblast or epithelial cells. The present studies investigated the mechanism underlying this differential apoptotic effect. Infection of melanoma cells, but not normal immortal melanocytes, with Ad.mda-7 induced a time- and dose-dependent increase in expression, mRNA and protein, of a family of growth arrest and DNA damage (GADD)-inducible genes, which correlated with induction of apoptosis. Among the members of the GADD family of genes, GADD153, GADD45 alpha, and GADD34 displayed marked, and GADD45 gamma showed minimal induction. Treatment of melanoma cells with SB203580, a selective inhibitor of the p38 mitogen-activated protein kinase (MAPK) pathway, effectively inhibited Ad.mda-7-induced apoptosis. Additional support for an involvement of the p38 MAPK pathway in Ad.mda-7 mediated apoptosis was documented by using an adenovirus expressing a dominant negative mutant of p38 MAPK. Infection with Ad.mda-7 increased the phosphorylation of p38 MAPK and heat shock protein 27 in melanoma cells but not in normal immortal melanocytes. In addition, SB203580 effectively inhibited Ad.mda-7-mediated induction of the GADD family of genes in a time- and dose dependent manner, and it effectively blocked Ad.mda-7-mediated down-regulation of the antiapoptotic protein BCL-2. Inhibition of GADD genes by an antisense approach either alone or in combination also effectively blocked Ad.mda-7-induced apoptosis in melanoma cells. These results support the hypothesis that Ad.mda-7 mediates induction of the GADD family of genes by means of the p38 MAPK pathway, thereby resulting in the selective induction of apoptosis in human melanoma cells. PMID- 12114540 TI - Human influenza viruses activate an interferon-independent transcription of cellular antiviral genes: outcome with influenza A virus is unique. AB - We examine the IFN-alpha/beta-independent activation of cellular transcription that constitutes an early antiviral response of cells against influenza A and B viruses, which cause widespread epidemics in humans. We show that influenza B virus induces the synthesis in human cells of several mature mRNAs encoded by genes containing an IFN-alpha/beta-stimulated response element (ISRE). Consequently, the IFN regulatory factor-3 transcription factor, which is required for the transcription of ISRE-controlled genes, is activated after influenza B virus infection. The production of these cellular mRNAs, some of which encode antiviral proteins, is independent of not only IFN-alpha/beta, but also viral protein synthesis. These mature cellular antiviral mRNAs are not produced after infection with influenza A virus, but IFN regulatory factor-3 is activated and the transcription of the ISRE-controlled p56 gene is induced. Consequently, like other newly synthesized cellular premRNAs in influenza A virus-infected cells, the posttranscriptional processing of premRNAs encoded by ISRE-controlled genes is inhibited. Previous work has established that such posttranscriptional inhibition is mediated by the viral NS1A protein. This unique, global countermeasure against the early, IFN-alpha/beta-independent antiviral response of cells may be an important factor in the pathogenicity of influenza A virus infection. PMID- 12114541 TI - Monoallelic expression and methylation of imprinted genes in human and mouse embryonic germ cell lineages. AB - Imprinting is an epigenetic modification leading to monoallelic expression of some genes, and disrupted imprinting is believed to be a barrier to human stem cell transplantation, based on studies that suggest that epigenetic marks are unstable in mouse embryonic germ (EG) and embryonic stem (ES) cells. However, stem cell imprinting has not previously been examined directly in humans. We found that three imprinted genes, TSSC5, H19, and SNRPN, show monoallelic expression in in vitro differentiated human EG-derived cells, and a fourth gene, IGF2, shows partially relaxed imprinting at a ratio from 4:1 to 5:1, comparable to that found in normal somatic cells. In addition, we found normal methylation of an imprinting control region (ICR) that regulates H19 and IGF2 imprinting, suggesting that imprinting may not be a significant epigenetic barrier to human EG cell transplantation. Finally, we were able to construct an in vitro mouse model of genomic imprinting, by generating EG cells from 8.5-day embryos of an interspecific cross, in which undifferentiated cells show biallelic expression and acquire preferential parental allele expression after differentiation. This model should allow experimental manipulation of epigenetic modifications of cultured EG cells that may not be possible in human stem cell studies. PMID- 12114542 TI - Genetic basis for systems of skeletal quantitative traits: principal component analysis of the canid skeleton. AB - Evolution of mammalian skeletal structure can be rapid and the changes profound, as illustrated by the morphological diversity of the domestic dog. Here we use principal component analysis of skeletal variation in a population of Portuguese Water Dogs to reveal systems of traits defining skeletal structures. This analysis classifies phenotypic variation into independent components that can be used to dissect genetic networks regulating complex biological systems. We show that unlinked quantitative trait loci associated with these principal components individually promote both correlations within structures (e.g., within the skull or among the limb bones) and inverse correlations between structures (e.g., skull vs. limb bones). These quantitative trait loci are consistent with regulatory genes that inhibit growth of some bones while enhancing growth of others. These systems of traits could explain the skeletal differences between divergent breeds such as Greyhounds and Pit Bulls, and even some of the skeletal transformations that characterize the evolution of hominids. PMID- 12114543 TI - Internal IgH class switch region deletions are position-independent and enhanced by AID expression. AB - Ig heavy chain class switch recombination (CSR) involves a recombination/deletion mechanism that exchanges the expressed C(H) gene with a downstream C(H) gene. CSR is mediated by highly repetitive switch (S) region sequences and requires the activation-induced deaminase (AID). The S region 5' of the C mu gene (S mu) can undergo high-frequency internal deletions in normal B cells and B cell lines activated for CSR, although the relationship of these deletions and CSR has not been elucidated. In this study, we introduced constitutively transcribed S mu or S gamma 2b regions into a pro-B cell line that can be activated for AID expression, CSR, and endogenous S mu deletions. We find that randomly integrated S region transcription units in these cells also undergo increased levels of internal rearrangements after cellular activation, indicating that the deletion process is independent of location within the Ig heavy chain locus and potentially AID-promoted. To test the latter issue, we generated hybridomas from wild-type and AID-deficient activated B cells and assayed them for internal S mu deletions and S region mutations. These studies demonstrated that efficient intra S region recombination depends on AID expression and that internal S region deletions are accompanied by frequent mutations, indicating that most S region deletions occur by the same mechanism as CSR. PMID- 12114544 TI - Endo-beta-N-acetylglucosaminidase, an enzyme involved in processing of free oligosaccharides in the cytosol. AB - Formation of oligosaccharides occurs both in the cytosol and in the lumen of the endoplasmic reticulum (ER). Luminal oligosaccharides are transported into the cytosol to ensure that they do not interfere with proper functioning of the glycan-dependent quality control machinery in the lumen of the ER for newly synthesized glycoproteins. Once in the cytosol, free oligosaccharides are catabolized, possibly to maximize the reutilization of the component sugars. An endo-beta-N-acetylglucosaminidase (ENGase) is a key enzyme involved in the processing of free oligosaccharides in the cytosol. This enzyme activity has been widely described in animal cells, but the gene encoding this enzyme activity has not been reported. Here, we report the identification of the gene encoding human cytosolic ENGase. After 11 steps, the enzyme was purified 150,000-fold to homogeneity from hen oviduct, and several internal amino acid sequences were analyzed. Based on the internal sequence and examination of expressed sequence tag (EST) databases, we identified the human orthologue of the purified protein. The human protein consists of 743 aa and has no apparent signal sequence, supporting the idea that this enzyme is localized in the cytosol. By expressing the cDNA of the putative human ENGase in COS-7 cells, the enzyme activity in the soluble fraction was enhanced 100-fold over the basal level, confirming that the human gene identified indeed encodes for ENGase. Careful gene database surveys revealed the occurrence of ENGase homologues in Drosophila melanogaster, Caenorhabditis elegans, and Arabidopsis thaliana, indicating the broad occurrence of ENGase in higher eukaryotes. This gene was expressed in a variety of human tissues, suggesting that this enzyme is involved in basic biological processes in eukaryotic cells. PMID- 12114546 TI - The GSK3-like kinase BIN2 phosphorylates and destabilizes BZR1, a positive regulator of the brassinosteroid signaling pathway in Arabidopsis. AB - Brassinosteroids (BRs) are a class of steroid hormones essential for normal growth and development in plants. BR signaling involves the cell-surface receptor BRI1, the glycogen synthase kinase-3-like kinase BIN2 as a negative regulator, and nuclear proteins BZR1 and BZR2/BES1 as positive regulators. The interactions among these components remain unclear. Here we report that BRs induce dephosphorylation and accumulation of BZR1 protein. Experiments using a proteasome inhibitor, MG132, suggest that phosphorylation of BZR1 increases its degradation by the proteasome machinery. BIN2 directly interacts with BZR1 in yeast two-hybrid assays, phosphorylates BZR1 in vitro, and negatively regulates BZR1 protein accumulation in vivo. These results strongly suggest that BIN2 phosphorylates BZR1 and targets it for degradation and that BR signaling causes BZR1 dephosphorylation and accumulation by inhibiting BIN2 activity. PMID- 12114545 TI - Calpain-dependent cleavage of cain/cabin1 activates calcineurin to mediate calcium-triggered cell death. AB - Cain/cabin1 is an endogenous inhibitor of calcineurin (Cn), a calcium-dependent serine/threonine phosphatase involved in various cellular functions including apoptosis. We show here that during apoptosis cain/cabin1 is cleaved by calpain at the carboxyl terminus to generate a cleavage product with a molecular mass of 32 kDa as a necessary step leading to Cn-mediated cell death. Mouse cain/cabin1 was identified from a thymus cDNA library by an in vitro substrate-screening assay with calpain. Exposure of Jurkat cells to the calcium ionophore, induced cain/cabin1 cleavage and cell death, accompanied by activation of calpain and Cn. The calpain inhibitors, calpeptin and zLLY, suppressed both -induced cain/cabin1 cleavage and Cn activation, indicating that Cn activation and cain/cabin1 cleavage are calpain-dependent. Expression of cain/cabin1 or a catalytically inactive Cn mutant [CnA beta(2)(1-401/H160N)] and treatment with FK506 reduced induced cell death. In vitro calpain cleavage and immunoprecipitation assays with deletion mutants of cain/cabin1 showed that cleavage occurred in the Cn-binding domain of cain/cabin1, indicating that the cleavage at its C terminus by calpain prevented cain/cabin1 from binding to Cn. In addition, in vitro binding assays showed that cain/cabin1 bound to the Cn B-binding domain of Cn A. Taken together, these results indicate that calpain cleaves the calcineurin-binding domain of cain/cabin1 to activate Cn and elicit calcium-triggered cell death. PMID- 12114548 TI - Spectral profiling for the simultaneous observation of four distinct fluorescent proteins and detection of protein-protein interaction via fluorescence resonance energy transfer in tobacco leaf nuclei. AB - The control of subcellular localization of proteins and their interaction with other partners in vivo are important parameters that provide clues to their function and regulation. The ability to simultaneously track multiple protein species with high resolution should provide a valuable assay system to study and characterize various types of posttranslational control pathways. In this work, we established the system and a method involving "spectral profiling" for the resolution of four different fluorescent protein tags in the same viewing field using digital imaging technology. With these techniques, we have (a) developed new derivatives of mGFP5, which is commonly used in the plant field, that are about three times brighter; (b) demonstrated that four spectrally distinct fluorescent proteins (cyan, green, yellow, and red) that are fused to a transcription factor could be stably expressed in nuclei and distinguished in tobacco (Nicotiana tabacum) mesophyll cells; and (c) shown that interaction between partners of a dimeric transcription factor can be detected by measuring fluorescence resonance energy transfer. These technologies should help one to study protein-protein interactions efficiently, especially for nuclear proteins under in vivo conditions. PMID- 12114547 TI - A postsynaptic transient K(+) current modulated by arachidonic acid regulates synaptic integration and threshold for LTP induction in hippocampal pyramidal cells. AB - Voltage-gated ion channels in the dendrites and somata of central neurons can modulate the impact of synaptic inputs. One of the ionic currents contributing to such modulation is the fast inactivating A-type potassium current (I(A)). We have investigated the role of I(A) in synaptic integration in rat CA1 pyramidal cells by using arachidonic acid (AA) and heteropodatoxin-3 (HpTX3), a selective blocker of the Kv4 channels underlying much of the somatodendritic I(A). AA and HpTX3 each reduced I(A) by 60-70% (measured at the soma) and strongly enhanced the amplitude and summation of excitatory postsynaptic responses, thus facilitating action potential discharges. HpTX3 also reduced the threshold for induction of long-term potentiation. We conclude that the postsynaptic I(A) is activated during synaptic depolarizations and effectively regulates the somatodendritic integration of high-frequency trains of synaptic input. AA, which can be released by such input, enhances synaptic efficacy by suppressing I(A), which could play an important role in frequency-dependent synaptic plasticity in the hippocampus. PMID- 12114550 TI - Molecular distinction between alternative oxidase from monocots and dicots. PMID- 12114551 TI - Nitric oxide is required for root organogenesis. PMID- 12114549 TI - Ethanol vapor is an efficient inducer of the alc gene expression system in model and crop plant species. AB - We have demonstrated that low concentrations of ethanol vapor efficiently induce the alc gene expression system in tobacco (Nicotiana tabacum cv Samsun NN), potato (Solanum tuberosum cv Solara), and oilseed rape (Brassica napus cv Westar). For many situations, this may be the preferred method of induction because it avoids direct application of comparatively high concentrations of an ethanol solution. Although induction was seen with less than 0.4 microM ethanol vapor, maximal induction of the chloramphenicol acetyl transferase gene was achieved after 48 h in leaves of tobacco plants enclosed with 4.5 microM ethanol vapor. In the absence of ethanol, there is no detectable gene expression. Treatment of potato tubers with ethanol vapor results in uniform beta glucoronidase (GUS) expression. Vapor treatment of a single oilseed rape leaf resulted in induction of GUS in the treated leaf only and (14)C-ethanol labeling in tobacco confirmed that the inducer was not translocated. In contrast, enclosure of the roots, aerial parts, or whole plant with ethanol vapor resulted in induction of GUS activity in leaves and roots. The data reported here broaden the utility of the alc system for research and crop biotechnology. PMID- 12114552 TI - The treasure trove of algal chloroplast genomes. Surprises in architecture and gene content, and their functional implications. PMID- 12114553 TI - Genetic manipulation of vacuolar proton pumps and transporters. PMID- 12114555 TI - Genome properties of the diatom Phaeodactylum tricornutum. AB - Diatoms are a ubiquitous class of microalgae of extreme importance for global primary productivity and for the biogeochemical cycling of minerals such as silica. However, very little is known about diatom cell biology or about their genome structure. For diatom researchers to take advantage of genomics and post genomics technologies, it is necessary to establish a model diatom species. Phaeodactylum tricornutum is an obvious candidate because of its ease of culture and because it can be genetically transformed. Therefore, we have examined its genome composition by the generation of approximately 1,000 expressed sequence tags. Although more than 60% of the sequences could not be unequivocally identified by similarity to sequences in the databases, approximately 20% had high similarity with a range of genes defined functionally at the protein level. It is interesting that many of these sequences are more similar to animal rather than plant counterparts. Base composition at each codon position and GC content of the genome were compared with Arabidopsis, maize (Zea mays), and Chlamydomonas reinhardtii. It was found that distribution of GC within the coding sequences is as homogeneous in P. tricornutum as in Arabidopsis, but with a slightly higher GC content. Furthermore, we present evidence that the P. tricornutum genome is likely to be small (less than 20 Mb). Therefore, this combined information supports the development of this species as a model system for molecular-based studies of diatom biology. The nucleotide sequence data reported has been deposited in GenBank Nucleotide Sequence Database (dbEST section) under accession nos. BI306757 through BI307753. PMID- 12114554 TI - Expression profiling of reciprocal maize hybrids divergent for cold germination and desiccation tolerance. AB - Recombinant inbred lines (RILs) derived from B73 x M017 were screened for cold germination (CG) and desiccation tolerance (DT) phenotypes. Reciprocal F(1) hybrids were made between divergent RILs, and hybrids that showed differential phenotypes (parent-of-origin effect) for CG or DT were selected for profiling mRNA and protein expression. mRNA and proteins were extracted from embryo axes of seed germinated for 11 d at 12.5 degrees C in the dark and developing embryos at 40% seed moisture (R5 stage) for CG and DT, respectively. GeneCalling analysis, an open-ended mRNA profiling method, identified 336 of 32,496 and 656 of 32,940 cDNA fragments that showed >or=1.5-fold change in expression between the reciprocal F(1) hybrids for CG and DT, respectively. Protein expression map (PEM) analysis, an open-ended two-dimensional polyacrylamide gel electrophoresis, identified 117 of 2,641 and 205 of 1,876 detected proteins to be differentially expressed with >or=1.5-fold change between the reciprocal F(1) hybrids in CG and DT samples, respectively. A subset of these proteins was identified by tandem mass spectrometry followed by database query of the spectra. The differentially expressed genes/proteins were classified into various functional groups including carbohydrate and amino acid metabolism, ion transporters, stress and defense response, polyamine metabolism, chaperonins, cytoskeleton associated, etc. Phenotypic analysis of seed from self-pollinated ears of the reciprocal F(1) hybrids displayed small differences compared with the reciprocal hybrids themselves, suggesting a negligible effect of cytoplasmic factors on CG and DT traits. The results provide leads to improving our understanding of the genes involved in stress response during seed maturation and germination. PMID- 12114556 TI - Methyl jasmonate induces traumatic resin ducts, terpenoid resin biosynthesis, and terpenoid accumulation in developing xylem of Norway spruce stems. AB - Norway spruce (Picea abies L. Karst) produces an oleoresin characterized by a diverse array of terpenoids, monoterpenoids, sesquiterpenoids, and diterpene resin acids that can protect conifers against potential herbivores and pathogens. Oleoresin accumulates constitutively in resin ducts in the cortex and phloem (bark) of Norway spruce stems. De novo formation of traumatic resin ducts (TDs) is observed in the developing secondary xylem (wood) after insect attack, fungal elicitation, and mechanical wounding. Here, we characterize the methyl jasmonate induced formation of TDs in Norway spruce by microscopy, chemical analyses of resin composition, and assays of terpenoid biosynthetic enzymes. The response involves tissue-specific differentiation of TDs, terpenoid accumulation, and induction of enzyme activities of both prenyltransferases and terpene synthases in the developing xylem, a tissue that constitutively lacks axial resin ducts in spruce. The induction of a complex defense response in Norway spruce by methyl jasmonate application provides new avenues to evaluate the role of resin defenses for protection of conifers against destructive pests such as white pine weevils (Pissodes strobi), bark beetles (Coleoptera, Scolytidae), and insect-associated tree pathogens. PMID- 12114557 TI - Novel insight into vascular, stress, and auxin-dependent and -independent gene expression programs in strawberry, a non-climacteric fruit. AB - Using cDNA microarrays, a comprehensive investigation of gene expression was carried out in strawberry (Fragaria x ananassa) fruit to understand the flow of events associated with its maturation and non-climacteric ripening. We detected key processes and novel genes not previously associated with fruit development and ripening, related to vascular development, oxidative stress, and auxin response. Microarray analysis during fruit development and in receptacle and seed (achene) tissues established an interesting parallelism in gene expression between the transdifferentiation of tracheary elements in Zinnia elegans and strawberry. One of the genes, CAD, common to both systems and encoding the lignin related protein cinnamyl alcohol dehydrogenase, was immunolocalized to immature xylem cells of the vascular bundles in the strawberry receptacle. To examine the importance of oxidative stress in ripening, gene expression was compared between fruit treated on-vine with a free radical generator and non-treated fruit. Of 46 genes induced, 20 were also ripening regulated. This might suggest that active gene expression is induced to cope with oxidative stress conditions during ripening or that the strawberry ripening transcriptional program is an oxidative stress-induced process. To gain insight into the hormonal control of non climacteric fruit ripening, an additional microarray experiment was conducted comparing gene expression in fruit treated exogenously with auxin and control fruit. Novel auxin-dependent genes and processes were identified in addition to transcriptional programs acting independent of auxin mainly related to cell wall metabolism and stress response. PMID- 12114558 TI - Effect of yeast CTA1 gene expression on response of tobacco plants to tobacco mosaic virus infection. AB - The response of tobacco (Nicotiana tabacum L. cv Xanthi-nc) plants with elevated catalase activity was studied after infection by tobacco mosaic virus (TMV). These plants contain the yeast (Saccharomyces cerevisiae) peroxisomal catalase gene CTA1 under the control of the cauliflower mosaic virus 35S promoter. The transgenic lines exhibited 2- to 4-fold higher total in vitro catalase activity than untransformed control plants under normal growth conditions. Cellular localization of the CTA1 protein was established using immunocytochemical analysis. Gold particles were detected mainly inside peroxisomes, whereas no significant labeling was detected in other cellular compartments or in the intercellular space. The physiological state of the transgenic plants was evaluated in respect to growth rate, general appearance, carbohydrate content, and dry weight. No significant differences were recorded in comparison with non transgenic tobacco plants. The 3,3'-diaminobenzidine-stain method was applied to visualize hydrogen peroxide (H(2)O(2)) in the TMV infected tissue. Presence of H(2)O(2) could be detected around necrotic lesions caused by TMV infection in non transgenic plants but to a much lesser extent in the CTA1 transgenic plants. In addition, the size of necrotic lesions was significantly bigger in the infected leaves of the transgenic plants. Changes in the distribution of H(2)O(2) and in lesion formation were not reflected by changes in salicylic acid production. In contrast to the local response, the systemic response in upper noninoculated leaves of both CTA1 transgenic and control plants was similar. This suggests that increased cellular catalase activity influences local but not systemic response to TMV infection. PMID- 12114559 TI - Immunohistochemistry of active gibberellins and gibberellin-inducible alpha amylase in developing seeds of morning glory. AB - Gibberellins (GAs) in developing seeds of morning glory (Pharbitis nil) were quantified and localized by immunostaining. The starch grains began to be digested after the GA contents had increased and reached a plateau. Immunohistochemical staining with the antigibberellin A(1)-methyl ester antiserum, which has high affinity to biologically active GAs, showed that GA(1) and/or GA(3) were localized around starch grains in the integument of developing young seeds, suggesting the participation of GA-inducible alpha-amylase in this digestion. We isolated an alpha-amylase cDNA (PnAmy1) that was expressed in the immature seeds, and using an antibody raised against recombinant protein, it was shown that PnAmy1 was expressed in the immature seeds. GA responsiveness of PnAmy1 was shown by treating the young fruits 9 d after anthesis with GA(3). RNA blot and immunoblot analyses showed that PnAmy1 emerged soon after the rapid increase of GA(1/3). An immunohistochemical analysis of PnAmy1 showed that it, like the seed GA(1/3), was also localized around starch grains in the integument of developing young seeds. The localization of GA(1/3) in the integument coincident with the expression of PnAmy1 suggests that both function as part of a process to release sugars for translocation or for the further development of the seeds. PMID- 12114560 TI - The liverwort contains a lectin that is structurally and evolutionary related to the monocot mannose-binding lectins. AB - A mannose (Man)-binding lectin has been isolated and characterized from the thallus of the liverwort Marchantia polymorpha. N-terminal sequencing indicated that the M. polymorpha agglutinin (Marpola) shares sequence similarity with the superfamily of monocot Man-binding lectins. Searches in the databases yielded expressed sequence tags encoding Marpola. Sequence analysis, molecular modeling, and docking experiments revealed striking structural similarities between Marpola and the monocot Man-binding lectins. Activity and specificity studies further indicated that Marpola is a much stronger agglutinin than the Galanthus nivalis agglutinin and exhibits a preference for methylated Man and glucose, which is unprecedented within the family of monocot Man-binding lectins. The discovery of Marpola allows us, for the first time, to corroborate the evolutionary relationship between a lectin from a lower plant and a well-established lectin family from flowering plants. In addition, the identification of Marpola sheds a new light on the molecular evolution of the superfamily of monocot Man-binding lectins. Beside evolutionary considerations, the occurrence of a G. nivalis agglutinin homolog in a lower plant necessitates the rethinking of the physiological role of the whole family of monocot Man-binding lectins. PMID- 12114561 TI - Leucine-derived cyano glucosides in barley. AB - Barley (Hordeum vulgare) seedlings contain five cyano glucosides derived from the amino acid L-leucine (Leu). The chemical structure and the relative abundance of the cyano glucosides were investigated by liquid chromatography-mass spectrometry and nuclear magnetic resonance analyses using spring barley cultivars with high, medium, and low cyanide potential. The barley cultivars showed a 10-fold difference in their cyano glucoside content, but the relative content of the individual cyano glucosides remained constant. Epiheterodendrin, the only cyanogenic glucoside present, comprised 12% to 18% of the total content of cyano glucosides. It is proposed that the aglycones of all five cyano glucosides are formed by the initial action of a cytochrome P450 enzyme of the CYP79 family converting L-Leu into Z-3-methylbutanal oxime and subsequent action of a less specific CYP71E enzyme converting the oxime into 3-methylbutyro nitrile and mediating subsequent hydroxylations at the alpha-, as well as beta- and gamma-, carbon atoms. Presence of cyano glucosides in the barley seedlings was restricted to leaf tissue, with 99% confined to the epidermis cell layers of the leaf blade. Microsomal preparations from epidermal cells were not able to convert L [(14)C]Leu into the biosynthetic intermediate, Z-3-methylbutanal-oxime. This was only achieved using microsomal preparations from other cell types in the basal leaf segment, demonstrating translocation of the cyano glucosides to the epidermal cell layers after biosynthesis. A beta-glucosidase able to degrade epiheterodendrin was detected exclusively in yet a third compartment, the endosperm of the germinating seed. Therefore, in barley, a putative function of cyano glucosides in plant defense is not linked to cyanide release. PMID- 12114562 TI - The barley MLO modulator of defense and cell death is responsive to biotic and abiotic stress stimuli. AB - Lack of the barley (Hordeum vulgare) seven-transmembrane domain MLO protein confers resistance against the fungal pathogen Blumeria graminis f. sp. hordei (Bgh). To broaden the basis for MLO structure/function studies, we sequenced additional mlo resistance alleles, two of which confer only partial resistance. Wild-type MLO dampens the cell wall-restricted hydrogen peroxide burst at points of attempted fungal penetration of the epidermal cell wall, and in subtending mesophyll cells, it suppresses a second oxidative burst and cell death. Although the Bgh-induced cell death in mlo plants is spatially and temporally separated from resistance, we show that the two processes are linked. Uninoculated mutant mlo plants exhibit spontaneous mesophyll cell death that appears to be part of accelerated leaf senescence. Mlo transcript abundance increases in response to Bgh, rice (Oryza sativa) blast, wounding, paraquat treatment, a wheat powdery mildew-derived carbohydrate elicitor, and during leaf senescence. This suggests a broad involvement of Mlo in cell death protection and in responses to biotic and abiotic stresses. PMID- 12114563 TI - Heterology expression of the Arabidopsis C-repeat/dehydration response element binding factor 1 gene confers elevated tolerance to chilling and oxidative stresses in transgenic tomato. AB - In an attempt to improve stress tolerance of tomato (Lycopersicon esculentum) plants, an expression vector containing an Arabidopsis C-repeat/dehydration responsive element binding factor 1 (CBF1) cDNA driven by a cauliflower mosaic virus 35S promoter was transferred into tomato plants. Transgenic expression of CBF1 was proved by northern- and western-blot analyses. The degree of chilling tolerance of transgenic T(1) and T(2) plants was found to be significantly greater than that of wild-type tomato plants as measured by survival rate, chlorophyll fluorescence value, and radical elongation. The transgenic tomato plants exhibited patterns of growth retardation; however, they resumed normal growth after GA(3) (gibberellic acid) treatment. More importantly, GA(3)-treated transgenic plants still exhibited a greater degree of chilling tolerance compared with wild-type plants. Subtractive hybridization was performed to isolate the responsive genes of heterologous Arabidopsis CBF1 in transgenic tomato plants. CATALASE1 (CAT1) was obtained and showed activation in transgenic tomato plants. The CAT1 gene and catalase activity were also highly induced in the transgenic tomato plants. The level of H(2)O(2) in the transgenic plants was lower than that in the wild-type plants under either normal or cold conditions. The transgenic plants also exhibited considerable tolerance against oxidative damage induced by methyl viologen. Results from the current study suggest that heterologous CBF1 expression in transgenic tomato plants may induce several oxidative-stress responsive genes to protect from chilling stress. PMID- 12114564 TI - Regulation of squalene synthase, a key enzyme of sterol biosynthesis, in tobacco. AB - Squalene synthase (SS) represents a putative branch point in the isoprenoid biosynthetic pathway capable of diverting carbon flow specifically to the biosynthesis of sterols and, hence, is considered a potential regulatory point for sterol metabolism. For example, when plant cells grown in suspension culture are challenged with fungal elicitors, suppression of sterol biosynthesis has been correlated with a reduction in SS enzyme activity. The current study sought to correlate changes in SS enzyme activity with changes in the level of the corresponding protein and mRNA. Using an SS-specific antibody, the initial suppression of SS enzyme activity in elicitor-challenged cells was not reflected by changes in the absolute level of the corresponding polypeptide, implicating a post-translational control mechanism for this enzyme activity. In comparison, the absolute level of the SS mRNA did decrease approximately 5-fold in the elicitor treated cells, which is suggestive of decreased transcription of the SS gene. Study of SS in intact plants was also initiated by measuring the level of SS enzyme activity, the level of the corresponding protein, and the expression of SS gene promoter-reporter gene constructs in transgenic plants. SS enzyme activity, polypeptide level, and gene expression were all localized predominately to the shoot apical meristem, with much lower levels observed in leaves and roots. These later results suggest that sterol biosynthesis is localized to the apical meristems and that apical meristems may be a source of sterols for other plant tissues. PMID- 12114565 TI - The white clover enod40 gene family. Expression patterns of two types of genes indicate a role in vascular function. AB - Enod40 is one of the genes associated with legume nodule development and has a putative role in general plant organogenesis. We have isolated a small enod40 gene family from white clover (Trifolium repens), with three genes designated Trenod40-1, Trenod40-2, and Trenod40-3, all containing the conserved enod40 regions I and II. Trenod40-1 and Trenod40-2 share over 90% homology in the transcribed regions and high levels of similarity in their upstream regulatory sequences. Trenod40-1 and Trenod40-2 are similar to the enod40 genes of legumes forming indeterminate nodules (group II) and are predominantly expressed in nodules. Trenod40-3 shares only 32.8% identity with Trenod40-1 and Trenod40-2 within the transcribed region. Trenod40-3 is similar to the enod40 genes of legumes with determinate nodules (group I) and is not predominantly expressed in nodules. To our knowledge, this is the first report of both group I- and group II type enod40 genes being expressed in a single legume species. In situ hybridization studies revealed that Trenod40 genes were highly expressed in non symbiotic tissues, particularly in stolon nodes during nodal root and lateral shoot development. High levels of Trenod40 transcripts were also present in the vascular bundles of mature plant organs, mainly at sites of intensive lateral transport, suggesting a role in vascular tissue function. The expression pattern of Trenod40 genes was analyzed further using Trenod40 promoter-gus fusions in transgenic white clover and tobacco (Nicotiana tabacum), indicating that white clover and tobacco share the regulatory mechanisms for Trenod40-1/2 promoters and some aspects of Trenod40-3 regulation. PMID- 12114566 TI - Distinct physiological roles of fructokinase isozymes revealed by gene-specific suppression of Frk1 and Frk2 expression in tomato. AB - There are two divergent fructokinase isozymes, Frk1 and Frk2 in tomato (Lycopersicon esculentum Mill.) plants. To investigate the physiological functions of each isozyme, the expression of each fructokinase mRNA was independently suppressed in transgenic tomato plants, and the respective phenotypes were evaluated. Suppression of Frk1 expression resulted in delayed flowering at the first inflorescence. Suppression of Frk2 did not effect flowering time but resulted in growth inhibition of stems and roots, reduction of flower and fruit number, and reduction of seed number per fruit. Localization of Frk1 and Frk2 mRNA accumulation by in situ hybridization in wild-type tomato fruit tissue indicated that Frk2 is expressed specifically in early tomato seed development. Fruit hexose and starch content were not effected by the suppression of either Frk gene alone. The results collectively indicate that flowering time is specifically promoted by Frk1 and that Frk2 plays specific roles in contributing to stem and root growth and to seed development. Because Frk1 and Frk2 gene expression was suppressed individually in transgenic plants, other significant metabolic roles of fructokinases may not have been observed if Frk1 and Frk2 play, at least partially, redundant metabolic roles. PMID- 12114567 TI - The serine-rich N-terminal domain of oat phytochrome a helps regulate light responses and subnuclear localization of the photoreceptor. AB - Phytochrome (phy) A mediates two distinct photobiological responses in plants: the very-low-fluence response (VLFR), which can be saturated by short pulses of very-low-fluence light, and the high-irradiance response (HIR), which requires prolonged irradiation with higher fluences of far-red light (FR). To investigate whether the VLFR and HIR involve different domains within the phyA molecule, transgenic tobacco (Nicotiana tabacum cv Xanthi) and Arabidopsis seedlings expressing full-length (FL) and various deletion mutants of oat (Avena sativa) phyA were examined for their light sensitivity. Although most mutants were either partially active or inactive, a strong differential effect was observed for the Delta6-12 phyA mutant missing the serine-rich domain between amino acids 6 and 12. Delta6-12 phyA was as active as FL phyA for the VLFR of hypocotyl growth and cotyledon unfolding in Arabidopsis, and was hyperactive in the VLFR of hypocotyl growth and cotyledon unfolding in tobacco, and the VLFR blocking subsequent greening under white light in Arabidopsis. In contrast, Delta6-12 phyA showed a dominant-negative suppression of HIR in both species. In hypocotyl cells of Arabidopsis irradiated with FR phyA:green fluorescent protein (GFP) and Delta6-12 phyA:GFP fusions localized to the nucleus and coalesced into foci. The proportion of nuclei with abundant foci was enhanced by continuous compared with hourly FR provided at equal total fluence in FL phyA:GFP, and by Delta6-12 phyA mutation under hourly FR. We propose that the N-terminal serine-rich domain of phyA is involved in channeling downstream signaling via the VLFR or HIR pathways in different cellular contexts. PMID- 12114568 TI - Combinatorial interaction of cis elements specifies the expression of the Arabidopsis AtHsp90-1 gene. AB - The promoter region of the Arabidopsis AtHsp90-1 gene is congested with heat shock elements and stress response elements, as well as with other potential transcriptional binding sites (activating protein 1, CCAAT/enhancer-binding protein element, and metal regulatory element). To determine how the expression of this bona fide AtHsp90-1 gene is regulated, a comprehensive quantitative and qualitative promoter deletion analysis was conducted under various environmental conditions and during development. The promoter induces gene expression at high levels after heat shock and arsenite treatment. However, our results show that the two stress responses may involve common but not necessarily the same regulatory elements. Whereas for heat induction, heat shock elements and stress response elements act cooperatively to promote high levels of gene expression, arsenite induction seems to require the involvement of activating protein 1 regulatory sequences. In stressed transgenic plants harboring the full-length promoter, beta-glucuronidase activity was prominent in all tissues. Nevertheless, progressive deletion of the promoter decreases the level of expression under heat shock and restricts it predominantly in the two meristems of the plant. In contrast, under arsenite induction, proximal sequences induce AtHsp90-1 gene expression only in the shoot meristem. Distally located elements negatively regulate AtHsp90-1 gene expression under unstressed conditions, whereas flower specific regulated expression in mature pollen grains suggests the prominent role of the AtHsp90-1 in pollen development. The results show that the regulation of developmental expression, suppression, or stress induction is mainly due to combinatorial contribution of the cis elements in the promoter region of the AtHsp90-1 gene. PMID- 12114569 TI - PROLIFERATING INFLORESCENCE MERISTEM, a MADS-box gene that regulates floral meristem identity in pea. AB - SQUAMOSA and APETALA1 are floral meristem identity genes from snapdragon (Antirrhinum majus) and Arabidopsis, respectively. Here, we characterize the floral meristem identity mutation proliferating inflorescence meristem (pim) from pea (Pisum sativum) and show that it corresponds to a defect in the PEAM4 gene, a homolog of SQUAMOSA and APETALA1. The PEAM4 coding region was deleted in the pim 1 allele, and this deletion cosegregated with the pim-1 mutant phenotype. The pim 2 allele carried a nucleotide substitution at a predicted 5' splice site that resulted in mis-splicing of pim-2 mRNA. PCR products corresponding to unspliced and exon-skipped mRNA species were observed. The pim-1 and pim-2 mutations delayed floral meristem specification and altered floral morphology significantly but had no observable effect on vegetative development. These floral-specific mutant phenotypes and the restriction of PIM gene expression to flowers contrast with other known floral meristem genes in pea that additionally affect vegetative development. The identification of PIM provides an opportunity to compare pathways to flowering in species with different inflorescence architectures. PMID- 12114570 TI - Targeting tryptophan decarboxylase to selected subcellular compartments of tobacco plants affects enzyme stability and in vivo function and leads to a lesion-mimic phenotype. AB - Tryptophan decarboxylase (TDC) is a cytosolic enzyme that catalyzes an early step of the terpenoid indole alkaloid biosynthetic pathway by decarboxylation of L tryptophan to produce the protoalkaloid tryptamine. In the present study, recombinant TDC was targeted to the chloroplast, cytosol, and endoplasmic reticulum (ER) of tobacco (Nicotiana tabacum) plants to evaluate the effects of subcellular compartmentation on the accumulation of functional enzyme and its corresponding enzymatic product. TDC accumulation and in vivo function was significantly affected by the subcellular localization. Immunoblot analysis demonstrated that chloroplast-targeted TDC had improved accumulation and/or stability when compared with the cytosolic enzyme. Because ER-targeted TDC was not detectable by immunoblot analysis and tryptamine levels found in transient expression studies and in transgenic plants were low, it was concluded that the recombinant TDC was most likely unstable if ER retained. Targeting TDC to the chloroplast stroma resulted in the highest accumulation level of tryptamine so far reported in the literature for studies on heterologous TDC expression in tobacco. However, plants accumulating high levels of functional TDC in the chloroplast developed a lesion-mimic phenotype that was probably triggered by the relatively high accumulation of tryptamine in this compartment. We demonstrate that subcellular targeting may provide a useful strategy for enhancing accumulation and/or stability of enzymes involved in secondary metabolism and to divert metabolic flux toward desired end products. However, metabolic engineering of plants is a very demanding task because unexpected, and possibly unwanted, effects may be observed on plant metabolism and/or phenotype. PMID- 12114571 TI - Overexpression of cytosolic glutamine synthetase. Relation to nitrogen, light, and photorespiration. AB - In plants, ammonium released during photorespiration exceeds primary nitrogen assimilation by as much as 10-fold. Analysis of photorespiratory mutants indicates that photorespiratory ammonium released in mitochondria is reassimilated in the chloroplast by a chloroplastic isoenzyme of glutamine synthetase (GS2), the predominant GS isoform in leaves of Solanaceous species including tobacco (Nicotiana tabacum). By contrast, cytosolic GS1 is expressed in the vasculature of several species including tobacco. Here, we report the effects on growth and photorespiration of overexpressing a cytosolic GS1 isoenzyme in leaf mesophyll cells of tobacco. The plants, which ectopically overexpress cytosolic GS1 in leaves, display a light-dependent improved growth phenotype under nitrogen-limiting and nitrogen-non-limiting conditions. Improved growth was evidenced by increases in fresh weight, dry weight, and leaf soluble protein. Because the improved growth phenotype was dependent on light, this suggested that the ectopic expression of cytosolic GS1 in leaves may act via photosynthetic/photorespiratory process. The ectopic overexpression of cytosolic GS1 in tobacco leaves resulted in a 6- to 7-fold decrease in levels of free ammonium in leaves. Thus, the overexpression of cytosolic GS1 in leaf mesophyll cells seems to provide an alternate route to chloroplastic GS2 for the assimilation of photorespiratory ammonium. The cytosolic GS1 transgenic plants also exhibit an increase in the CO(2) photorespiratory burst and an increase in levels of photorespiratory intermediates, suggesting changes in photorespiration. Because the GS1 transgenic plants have an unaltered CO(2) compensation point, this may reflect an accompanying increase in photosynthetic capacity. Together, these results provide new insights into the possible mechanisms responsible for the improved growth phenotype of cytosolic GS1 overexpressing plants. Our studies provide further support for the notion that the ectopic overexpression of genes for cytosolic GS1 can potentially be used to affect increases in nitrogen use efficiency in transgenic crop plants. PMID- 12114573 TI - The physiology and biophysics of an aluminum tolerance mechanism based on root citrate exudation in maize. AB - Al-induced release of Al-chelating ligands (primarily organic acids) into the rhizosphere from the root apex has been identified as a major Al tolerance mechanism in a number of plant species. In the present study, we conducted physiological investigations to study the spatial and temporal characteristics of Al-activated root organic acid exudation, as well as changes in root organic acid content and Al accumulation, in an Al-tolerant maize (Zea mays) single cross (SLP 181/71 x Cateto Colombia 96/71). These investigations were integrated with biophysical studies using the patch-clamp technique to examine Al-activated anion channel activity in protoplasts isolated from different regions of the maize root. Exposure to Al nearly instantaneously activated a concentration-dependent citrate release, which saturated at rates close to 0.5 nmol citrate h(-1) root( 1), with the half-maximal rates of citrate release occurring at about 20 microM Al(3+) activity. Comparison of citrate exudation rates between decapped and capped roots indicated the root cap does not play a major role in perceiving the Al signal or in the exudation process. Spatial analysis indicated that the predominant citrate exudation is not confined to the root apex, but could be found as far as 5 cm beyond the root cap, involving cortex and stelar cells. Patch clamp recordings obtained in whole-cell and outside-out patches confirmed the presence of an Al-inducible plasma membrane anion channel in protoplasts isolated from stelar or cortical tissues. The unitary conductance of this channel was 23 to 55 pS. Our results suggest that this transporter mediates the Al induced citrate release observed in the intact tissue. In addition to the rapid Al activation of citrate release, a slower, Al-inducible increase in root citrate content was also observed. These findings led us to speculate that in addition to the Al exclusion mechanism based on root citrate exudation, a second internal Al tolerance mechanism may be operating based on Al-inducible changes in organic acid synthesis and compartmentation. We discuss our findings in terms of recent genetic studies of Al tolerance in maize, which suggest that Al tolerance in maize is a complex trait. PMID- 12114572 TI - Leaf senescence and starvation-induced chlorosis are accelerated by the disruption of an Arabidopsis autophagy gene. AB - Autophagy is an intracellular process for vacuolar bulk degradation of cytoplasmic components. The molecular machinery responsible for yeast and mammalian autophagy has recently begun to be elucidated at the cellular level, but the role that autophagy plays at the organismal level has yet to be determined. In this study, a genome-wide search revealed significant conservation between yeast and plant autophagy genes. Twenty-five plant genes that are homologous to 12 yeast genes essential for autophagy were discovered. We identified an Arabidopsis mutant carrying a T-DNA insertion within AtAPG9, which is the only ortholog of yeast Apg9 in Arabidopsis (atapg9-1). AtAPG9 is transcribed in every wild-type organ tested but not in the atapg9-1 mutant. Under nitrogen or carbon-starvation conditions, chlorosis was observed earlier in atapg9-1 cotyledons and rosette leaves compared with wild-type plants. Furthermore, atapg9-1 exhibited a reduction in seed set when nitrogen starved. Even under nutrient growth conditions, bolting and natural leaf senescence were accelerated in atapg9-1 plants. Senescence-associated genes SEN1 and YSL4 were up regulated in atapg9-1 before induction of senescence, unlike in wild type. All of these phenotypes were complemented by the expression of wild-type AtAPG9 in atapg9-1 plants. These results imply that autophagy is required for maintenance of the cellular viability under nutrient-limited conditions and for efficient nutrient use as a whole plant. PMID- 12114574 TI - Selective activation of the developmentally regulated Ha hsp17.6 G1 promoter by heat stress transcription factors. AB - Using two well-characterized heat stress transcription factors (Hsfs) from tomato (Lycopersicon peruvianum; LpHsfA1 and LpHsfA2), we analyzed the transcriptional activation of the Ha hsp17.6 G1 promoter in sunflower (Helianthus annuus) embryos. In this system, we observed transient promoter activation only with LpHsfA2. In contrast, both factors were able to activate mutant versions of the promoter with improved consensus Hsf-binding sites. Exclusive activation by LpHsfA2 was also observed in yeast (Saccharomyces cerevisiae) without other Hsfs and with a minimal Cyc1 promoter fused to the Ha hsp17.6 G1 heat stress cis element. Furthermore, the same promoter mutations reproduced the loss of activation selectivity, as observed in sunflower embryos. The results of in vitro binding experiments rule out differential DNA binding of the two factors as the explanation for the observed differential activation capacity. We conclude that the specific sequence of this heat stress cis-element is crucial for Hsf promoter selectivity, and that this selectivity could involve preferential transcriptional activation following DNA binding. In sunflower embryos, we also observed synergistic transcriptional activation by co-expression of LpHsfA1 and LpHsfA2. Mutational analyses of the Ha hsp17.6 G1 promoter, combined with in vitro binding assays, suggest that mixed oligomers of the two factors may be involved in promoter activation. We discuss the relevance of our observations for mechanisms of developmental regulation of plant heat stress protein genes. PMID- 12114575 TI - Effect of short-term N(2) deficiency on expression of the ureide pathway in cowpea root nodules. AB - Root systems of 28-d-old cowpea (Vigna unguiculata L. Walp cv Vita 3: Bradyrhizobium sp. strain CB756) plants bearing nitrogen-fixing nodules in sand culture were exposed to an atmosphere of Ar:O(2) (80:20, v/v) for 48 h and then returned to air. Root systems of control plants were maintained in air throughout. Nodules were harvested at the same times in control and Ar:O(2) treated root systems. Activities of two enzymes of de novo purine synthesis, glycinamide ribonucleotide transformylase (GART; EC 2.1.2.2), aminoimidazole ribonucleotide synthetase (AIRS; EC 6.3.3.1), uricase (EC 1.7.3.3), and phosphoenolpyruvate carboxylase (PEPC; EC 4.1.1.31) were measured together with the protein level of each using immune-specific polyclonal antibodies. AIRS activity and protein both declined to very low levels within 6 h in Ar:O(2) together with a decline in transcript level of pur5, the encoding gene. GART activity, protein, and transcript (pur3) levels were relatively stable. Uricase activity declined in Ar:O(2) as rapidly as AIRS activity but the protein was stable. PEPC activity showed evidence of increased sensitivity to inhibition by malate but the protein level was stable. The data indicate that the flux of fixed N from bacteroids (N(2)-fixing nodule bacteria) is in some way associated with transcriptional control over pur5 and possibly also catabolism of AIRS protein. In contrast, there is limited posttranslational control over GART and PEPC and close posttranslational control over uricase activity. The significance of these different levels of regulation is discussed in relation to the overall control of enhanced expression of plant enzymes in the cowpea symbiosis. PMID- 12114576 TI - Dhurrin synthesis in sorghum is regulated at the transcriptional level and induced by nitrogen fertilization in older plants. AB - The content of the cyanogenic glucoside dhurrin in sorghum (Sorghum bicolor L. Moench) varies depending on plant age and growth conditions. The cyanide potential is highest shortly after onset of germination. At this stage, nitrogen application has no effect on dhurrin content, whereas in older plants, nitrogen application induces an increase. At all stages, the content of dhurrin correlates well with the activity of the two biosynthetic enzymes, CYP79A1 and CYP71E1, and with the protein and mRNA level for the two enzymes. During development, the activity of CYP79A1 is lower than the activity of CYP71E1, suggesting that CYP79A1 catalyzes the rate-limiting step in dhurrin synthesis as has previously been shown using etiolated seedlings. The site of dhurrin synthesis shifts from leaves to stem during plant development. In combination, the results demonstrate that dhurrin content in sorghum is largely determined by transcriptional regulation of the biosynthetic enzymes CYP79A1 and CYP71E1. PMID- 12114577 TI - Phosphite, an analog of phosphate, suppresses the coordinated expression of genes under phosphate starvation. AB - Phosphate (Pi) and its analog phosphite (Phi) are acquired by plants via Pi transporters. Although the uptake and mobility of Phi and Pi are similar, there is no evidence suggesting that plants can utilize Phi as a sole source of phosphorus. Phi is also known to interfere with many of the Pi starvation responses in plants and yeast (Saccharomyces cerevisiae). In this study, effects of Phi on plant growth and coordinated expression of genes induced by Pi starvation were analyzed. Phi suppressed many of the Pi starvation responses that are commonly observed in plants. Enhanced root growth and root to shoot ratio, a hallmark of Pi stress response, was strongly inhibited by Phi. The negative effects of Phi were not obvious in plants supplemented with Pi. The expression of Pi starvation-induced genes such as LePT1, LePT2, AtPT1, and AtPT2 (high-affinity Pi transporters); LePS2 (a novel acid phosphatase); LePS3 and TPSI1 (novel genes); and PAP1 (purple acid phosphatase) was suppressed by Phi in plants and cell cultures. Expression of luciferase reporter gene driven by the Pi starvation induced AtPT2 promoter was also suppressed by Phi. These analyses showed that suppression of Pi starvation-induced genes is an early response to addition of Phi. These data also provide evidence that Phi interferes with gene expression at the level of transcription. Synchronized suppression of multiple Pi starvation induced genes by Phi points to its action on the early molecular events, probably signal transduction, in Pi starvation response. PMID- 12114578 TI - Brassinosteroid-regulated gene expression. AB - Major brassinosteroid (BR) effects such as BR-induced growth are mediated through genomic pathways because RNA synthesis inhibitors and protein synthesis inhibitors interfere with these processes. A limited number of BR-regulated genes have been identified hitherto. The majority of genes (such as BRU1, CycD3, Lin6, OPR3, and TRIP-1) were identified by comparisons of BR-treated versus control treated plants. However, altered transcript levels after BR application may not reflect normal physiological events. A complementary approach is the comparison of BR-deficient plants versus wild-type plants. No artificial treatments interfere with endogenous signaling pathways, but a subset of phenotypic alterations of phytohormone-deficient plants most probably is secondary. To identify genes that are subject to direct BR regulation, we analyzed CPD antisense and dwf1-6 (cbb1) mutant plants. Both show a mild phenotype in comparison with BR-deficient mutants such as cpd/cbb3, det2, and dwf4. Plants were grown under two different environments to filter out BR deficiency effects that occur only at certain environmental conditions. Finally, we established expression patterns after BR treatment of wild-type and dwf1-6 (cbb1) plants. Ideally, a BR-regulated gene displays a dose-response relationship in such a way that a gene with decreased transcript levels in BR-deficient plants is BR inducible and vice versa. Expression profile analysis of above ground part of plants was performed by means of Affymetrix Arabidopsis Genome Arrays. PMID- 12114580 TI - Glyphosate-resistant goosegrass. Identification of a mutation in the target enzyme 5-enolpyruvylshikimate-3-phosphate synthase. AB - The spontaneous occurrence of resistance to the herbicide glyphosate in weed species has been an extremely infrequent event, despite over 20 years of extensive use. Recently, a glyphosate-resistant biotype of goosegrass (Eleusine indica) was identified in Malaysia exhibiting an LD(50) value approximately 2- to 4-fold greater than the sensitive biotype collected from the same region. A comparison of the inhibition of 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) activity by glyphosate in extracts prepared from the resistant (R) and sensitive (S) biotypes revealed an approximately 5-fold higher IC(50)(glyphosate) for the (R) biotype. Sequence comparisons of the predicted EPSPS mature protein coding regions from both biotypes revealed four single-nucleotide differences, two of which result in amino acid changes. One of these changes, a proline to serine substitution at position 106 in the (R) biotype, corresponds to a substitution previously identified in a glyphosate-insensitive EPSPS enzyme from Salmonella typhimurium. Kinetic data generated for the recombinant enzymes suggests that the second substitution identified in the (R) EPSPS does not contribute significantly to its reduced glyphosate sensitivity. Escherichia coli aroA- (EPSPS deficient) strains expressing the mature EPSPS enzyme from the (R) biotype exhibited an approximately 3-fold increase in glyphosate tolerance relative to strains expressing the mature EPSPS from the (S) biotype. These results provide the first evidence for an altered EPSPS enzyme as an underlying component of evolved glyphosate resistance in any plant species. PMID- 12114579 TI - Investigation of the microheterogeneity and aglycone specificity-conferring residues of black cherry prunasin hydrolases. AB - In black cherry (Prunus serotina Ehrh.) seed homogenates, (R)-amygdalin is degraded to HCN, benzaldehyde, and glucose by the sequential action of amygdalin hydrolase (AH), prunasin hydrolase (PH), and mandelonitrile lyase. Leaves are also highly cyanogenic because they possess (R)-prunasin, PH, and mandelonitrile lyase. Taking both enzymological and molecular approaches, we demonstrate here that black cherry PH is encoded by a putative multigene family of at least five members. Their respective cDNAs (designated Ph1, Ph2, Ph3, Ph4, and Ph5) predict isoforms that share 49% to 92% amino acid identity with members of glycoside hydrolase family 1, including their catalytic asparagine-glutamate-proline and isoleucine-threonine-glutamate-asparagine-glycine motifs. Furthermore, consistent with the vacuolar/protein body location and glycoprotein character of these hydrolases, their open reading frames predict N-terminal signal sequences and multiple potential N-glycosylation sites. Genomic sequences corresponding to the open reading frames of these PHs and of the previously isolated AH1 isoform are interrupted at identical positions by 12 introns. Earlier studies established that native AH and PH display strict specificities toward their respective glucosidic substrates. Such behavior was also shown by recombinant AH1, PH2, and PH4 proteins after expression in Pichia pastoris. Three amino acid moieties that may play a role in conferring such aglycone specificities were predicted by structural modeling and comparative sequence analysis and tested by introducing single and multiple mutations into isoform AH1 by site-directed mutagenesis. The double mutant AH ID (Y200I and G394D) hydrolyzed prunasin at approximately 150% of the rate of amygdalin hydrolysis, whereas the other mutations failed to engender PH activity. PMID- 12114581 TI - Differential accumulation of dimethylallyl diphosphate in leaves and needles of isoprene- and methylbutenol-emitting and nonemitting species. AB - The biosynthesis and emission of volatile plant terpenoids, such as isoprene and methylbutenol (MBO), depend on the chloroplastic production of dimethylallyl diphosphate (DMAPP). To date, it has been difficult to study the relationship of cellular DMAPP levels to emission of these volatiles because of the lack of a sensitive assay for DMAPP in plant tissues. Using a recent DMAPP assay developed in our laboratories, we report that species with the highest potential for isoprene and MBO production also exhibit elevated light-dependent DMAPP production, ranging from 110% to 1,063%. Even species that do not produce significant amounts of volatile terpenoids, however, exhibit some potential for light-dependent production of DMAPP. We used a nonaqueous fractionation technique to determine the intracellular distribution of DMAPP in isoprene-emitting cottonwood (Populus deltoides) leaves; approximately 65% to 70% of the DMAPP recovered at midday occurred in the chloroplasts, indicating that most of the light-dependent production of DMAPP was chloroplastic in origin. The midday concentration of chloroplastic DMAPP in cottonwood leaves is estimated to be 0.13 to 3.0 mM, which is consistent with the relatively high K(m)s that have been reported for isoprene synthases (0.5-8 mM). The results provide support for the hypothesis that the light dependence of isoprene and MBO emissions is in part due to controls over DMAPP production. PMID- 12114582 TI - Cloning and characterization of the abscisic acid-specific glucosyltransferase gene from adzuki bean seedlings. AB - The glycosylated forms of abscisic acid (ABA) have been identified from many plant species and are known to be the forms of ABA-catabolism, although their (physiological) roles have not yet been elucidated. ABA-glucosyltransferase ( GTase) is thought to play a key role in the glycosylation of ABA. We isolated an ABA-inducible GTase gene from UDP-GTase homologs obtained from adzuki bean (Vigna angularis) seedlings. The deduced amino acid sequence (accession no. AB065190) showed 30% to 44% identity with the known UDP-GTase homologs. The recombinant protein with a glutathione S-transferase-tag was expressed in Escherichia coli and showed enzymatic activity in an ABA-specific manner. The enzymatic activity was detected over a wide pH range from 5.0 to 9.0, the optimum range being between pH 6.0 and 7.3, in a citrate and Tris-HCl buffer. The product from racemic ABA and UDP-D-glucose was identified to be ABA-GE by gas chromatography/mass spectrometry. The recombinant GTase (rAOG) converted 2-trans (+)-ABA better than (+)-S-ABA and (-)-R-ABA. Although trans-cinnamic acid was slightly converted to its conjugate by the GTase, (-)-PA was not at all. The mRNA level was increased by ABA application or by water stress and wounding. We suggest that the gene encodes an ABA-specific GTase and that its expression is regulated by environmental stress. PMID- 12114583 TI - The effects of abiotic factors on induced volatile emissions in corn plants. AB - Many plants respond to herbivory by releasing a specific blend of volatiles that is attractive to natural enemies of the herbivores. In corn (Zea mays), this induced odor blend is mainly composed of terpenoids and indole. The induced signal varies with plant species and genotype, but little is known about the variation due to abiotic factors. Here, we tested the effect of soil humidity, air humidity, temperature, light, and fertilization rate on the emission of induced volatiles in young corn plants. Each factor was tested separately under constant conditions for the other factors. Plants released more when standing in dry soil than in wet soil, whereas for air humidity, the optimal release was found at around 60% relative humidity. Temperatures between 22 degrees C and 27 degrees C led to a higher emission than lower or higher temperatures. Light intensity had a dramatic effect. The emission of volatiles did not occur in the dark and increased steadily with an increase in the light intensity. An experiment with an unnatural light-dark cycle showed that the release was fully photophase dependent. Fertilization also had a strong positive effect; the emission of volatiles was minimal when plants were grown under low nutrition, even when results were corrected for plant biomass. Changes in all abiotic factors caused small but significant changes in the relative ratios among the different compounds (quality) in the induced odor blends, except for air humidity. Hence, climatic conditions and nutrient availability can be important factors in determining the intensity and variability in the release of induced plant volatiles. PMID- 12114584 TI - Proteome analysis of grain filling and seed maturation in barley. AB - In monocotyledonous plants, the process of seed development involves the deposition of reserves in the starchy endosperm and development of the embryo and aleurone layer. The final stages of seed development are accompanied by an increase in desiccation tolerance and drying out of the mature seed. We have used two-dimensional gel electrophoresis for a time-resolved study of the changes in proteins that occur during seed development in barley (Hordeum vulgare). About 1,000 low-salt extractable protein spots could be resolved on the two-dimensional gels. Protein spots were divided into six categories according to the timing of appearance or disappearance during the 5-week period of comparison. Nineteen different proteins or protein fragments in 36 selected spots were identified by matrix-assisted laser-desorption ionization time of flight mass spectrometry (MS) or nano-electrospray tandem MS/MS. Some proteins were present throughout development (for example, cytosolic malate dehydrogenase), whereas others were associated with the early grain filling (ascorbate peroxidase) or desiccation (Cor14b) stages. Most noticeably, the development process is characterized by an accumulation of low-M(r) alpha-amylase/trypsin inhibitors, serine protease inhibitors, and enzymes involved in protection against oxidative stress. We present examples of proteins not previously experimentally observed, differential extractability of thiol-bound proteins, and possible allele-specific spot variation. Our results both confirm and expand on knowledge gained from previous analyses of individual proteins involved in grain filling and maturation. PMID- 12114585 TI - Salt induction of fatty acid elongase and membrane lipid modifications in the extreme halotolerant alga Dunaliella salina. AB - In studies of the outstanding salt tolerance of the unicellular green alga Dunaliella salina, we isolated a cDNA for a salt-inducible mRNA encoding a protein homologous to plant beta-ketoacyl-coenzyme A (CoA) synthases (Kcs). These microsomal enzymes catalyze the condensation of malonyl-CoA with acyl-CoA, the first and rate-limiting step in fatty acid elongation. Kcs activity, localized to a D. salina microsomal fraction, increased in cells transferred from 0.5 to 3.5 M NaCl, as did the level of the kcs mRNA. The function of the kcs gene product was directly demonstrated by the condensing activity exhibited by Escherichia coli cells expressing the kcs cDNA. The effect of salinity on kcs expression in D. salina suggested the possibility that salt adaptation entailed modifications in the fatty acid composition of algal membranes. Lipid analyses indicated that microsomes, but not plasma membranes or thylakoids, from cells grown in 3.5 M NaCl contained a considerably higher ratio of C18 (mostly unsaturated) to C16 (mostly saturated) fatty acids compared with cells grown in 0.5 M salt. Thus, the salt-inducible Kcs, jointly with fatty acid desaturases, may play a role in adapting intracellular membrane compartments to function in the high internal glycerol concentrations balancing the external osmotic pressure. PMID- 12114586 TI - Gibberellic acid, synthetic auxins, and ethylene differentially modulate alpha-L Arabinofuranosidase activities in antisense 1-aminocyclopropane-1-carboxylic acid synthase tomato pericarp discs. AB - Alpha-L-Arabinofuranosidases (alpha-Afs) are plant enzymes capable of releasing terminal arabinofuranosyl residues from cell wall matrix polymers, as well as from different glycoconjugates. Three different alpha-Af isoforms were distinguished by size exclusion chromatography of protein extracts from control tomatoes (Lycopersicon esculentum) and an ethylene synthesis-suppressed (ESS) line expressing an antisense 1-aminocyclopropane-1-carboxylic synthase transgene. alpha-Af I and II are active throughout fruit ontogeny. alpha-Af I is the first Zn-dependent cell wall enzyme isolated from tomato pericarp tissues, thus suggesting the involvement of zinc in fruit cell wall metabolism. This isoform is inhibited by 1,10-phenanthroline, but remains stable in the presence of NaCl and sucrose. alpha-Af II activity accounts for over 80% of the total alpha-Af activity in 10-d-old fruit, but activity drops during ripening. In contrast, alpha-Af III is ethylene dependent and specifically active during ripening. alpha Af I released monosaccharide arabinose from KOH-soluble polysaccharides from tomato cell walls, whereas alpha-Af II and III acted on Na(2)CO(3)-soluble pectins. Different alpha-Af isoform responses to gibberellic acid, synthetic auxins, and ethylene were followed by using a novel ESS mature-green tomato pericarp disc system. alpha-Af I and II activity increased when gibberellic acid or 2,4-dichlorophenoxyacetic acid was applied, whereas ethylene treatment enhanced only alpha-Af III activity. Results suggest that tomato alpha-Afs are encoded by a gene family under differential hormonal controls, and probably have different in vivo functions. The ESS pericarp explant system allows comprehensive studies involving effects of physiological levels of different growth regulators on gene expression and enzyme activity with negligible wound-induced ethylene production. PMID- 12114587 TI - The role of ethylene and wound signaling in resistance of tomato to Botrytis cinerea. AB - Ethylene, jasmonate, and salicylate play important roles in plant defense responses to pathogens. To investigate the contributions of these compounds in resistance of tomato (Lycopersicon esculentum) to the fungal pathogen Botrytis cinerea, three types of experiments were conducted: (a) quantitative disease assays with plants pretreated with ethylene, inhibitors of ethylene perception, or salicylate; (b) quantitative disease assays with mutants or transgenes affected in the production of or the response to either ethylene or jasmonate; and (c) expression analysis of defense-related genes before and after inoculation of plants with B. cinerea. Plants pretreated with ethylene showed a decreased susceptibility toward B. cinerea, whereas pretreatment with 1-methylcyclopropene, an inhibitor of ethylene perception, resulted in increased susceptibility. Ethylene pretreatment induced expression of several pathogenesis-related protein genes before B. cinerea infection. Proteinase inhibitor I expression was repressed by ethylene and induced by 1-methylcyclopropene. Ethylene also induced resistance in the mutant Never ripe. RNA analysis showed that Never ripe retained some ethylene sensitivity. The mutant Epinastic, constitutively activated in a subset of ethylene responses, and a transgenic line producing negligible ethylene were also tested. The results confirmed that ethylene responses are important for resistance of tomato to B. cinerea. The mutant Defenseless, impaired in jasmonate biosynthesis, showed increased susceptibility to B. cinerea. A transgenic line with reduced prosystemin expression showed similar susceptibility as Defenseless, whereas a prosystemin-overexpressing transgene was highly resistant. Ethylene and wound signaling acted independently on resistance. Salicylate and ethylene acted synergistically on defense gene expression, but antagonistically on resistance. PMID- 12114588 TI - Arabidopsis mutants deficient in diacylglycerol acyltransferase display increased sensitivity to abscisic acid, sugars, and osmotic stress during germination and seedling development. AB - Arabidopsis seeds store triacylglycerol (TAG) as the major carbon reserve, which is used to support postgerminative seedling growth. Diacylglycerol acyltransferase (DGAT) catalyzes the final step in TAG synthesis, and two isoforms of DGAT have previously been identified in Arabidopsis. It has been shown that DGAT1 plays an important role in seed development because Arabidopsis with mutations at the TAG1 locus accumulate less seed oil. There is also evidence showing that DGAT1 is active after seed germination. The aim of this study is to investigate the effect of mutations of DGAT1 on postembryonic development in Arabidopsis. We carried out detailed analyses of two tag1 mutants in different ecotypic backgrounds of Arabidopsis. Results show that during germination and seedling growth, seed storage TAG degradation was not affected in the tag1 mutants. However, sugar content of the mutant seedlings is altered, and activities of the hexokinases are significantly increased in the tag1 mutant seedlings. The tag1 mutants are also more sensitive to abscisic acid, glucose, and osmotic strength of the medium in germination and seedling growth. PMID- 12114589 TI - Excess copper predisposes photosystem II to photoinhibition in vivo by outcompeting iron and causing decrease in leaf chlorophyll. AB - Photoinhibition of photosystem II was studied in vivo with bean (Phaseolus vulgaris) plants grown in the presence of 0.3 (control), 4, or 15 microM Cu(2+). Although photoinhibition, measured in the presence of lincomycin to block concurrent recovery, is faster in leaves of Cu(2+)-treated plants than in control leaves, thylakoids isolated from Cu-treated plants did not show high sensitivity to photoinhibition. Direct effects of excess Cu(2+) on chloroplast metabolism are actually unlikely, because the Cu concentration of chloroplasts of Cu-treated plants was lower than that of their leaves. Excess Cu in the growth medium did not cause severe oxidative stress, collapse of antioxidative defenses, or loss of photoprotection. Thus, these hypothetical effects can be eliminated as causes for Cu-enhanced photoinhibition in intact leaves. However, Cu treatment lowered the leaf chlorophyll (Chl) concentration and reduced the thylakoid membrane network. The loss of Chl and sensitivity to photoinhibition could be overcome by adding excess Fe together with excess Cu to the growth medium. The addition of Fe lowered the Cu(2+) concentration of the leaves, suggesting that Cu outcompetes Fe in Fe uptake. We suggest that the reduction of leaf Chl concentration, caused by the Cu-induced iron deficiency, causes the high photosensitivity of photosystem II in Cu(2+)-treated plants. A causal relationship between the susceptibility to photoinhibition and the leaf optical density was established in several plant species. Plant species adapted to high-light habitats apparently benefit from thick leaves because the rate of photoinhibition is directly proportional to light intensity, but photosynthesis becomes saturated by moderate light. PMID- 12114590 TI - Cold-regulated cereal chloroplast late embryogenesis abundant-like proteins. Molecular characterization and functional analyses. AB - Cold acclimation and freezing tolerance are the result of complex interaction between low temperature, light, and photosystem II (PSII) excitation pressure. Previous results have shown that expression of the Wcs19 gene is correlated with PSII excitation pressure measured in vivo as the relative reduction state of PSII. Using cDNA library screening and data mining, we have identified three different groups of proteins, late embryogenesis abundant (LEA) 3-L1, LEA3-L2, and LEA3-L3, sharing identities with WCS19. These groups represent a new class of proteins in cereals related to group 3 LEA proteins. They share important characteristics such as a sorting signal that is predicted to target them to either the chloroplast or mitochondria and a C-terminal sequence that may be involved in oligomerization. The results of subcellular fractionation, immunolocalization by electron microscopy and the analyses of target sequences within the Wcs19 gene are consistent with the localization of WCS19 within the chloroplast stroma of wheat (Triticum aestivum) and rye (Secale cereale). Western analysis showed that the accumulation of chloroplastic LEA3-L2 proteins is correlated with the capacity of different wheat and rye cultivars to develop freezing tolerance. Arabidopsis was transformed with the Wcs19 gene and the transgenic plants showed a significant increase in their freezing tolerance. This increase was only evident in cold-acclimated plants. The putative function of this protein in the enhancement of freezing tolerance is discussed. PMID- 12114591 TI - Ethylene emission and responsiveness to applied ethylene vary among Poa species that inherently differ in leaf elongation rates. AB - A plant's ability to produce and respond to ethylene is essential for its vegetative growth. We studied whole-shoot ethylene emission and leaf growth responses to applied ethylene in four Poa spp. that differ inherently in leaf elongation rate and whole-plant relative growth rate. Compared with the fast growing Poa annua and Poa trivialis, the shoots of the slow-growing species Poa alpina and Poa compressa emitted daily 30% to 50% less ethylene, and their leaf elongation rate was more strongly inhibited when ethylene concentration was increased up to 1 microL L(-1). To our surprise, however, low ethylene concentrations (0.02-0.03 microL L(-1)) promoted leaf growth in the two slow growing species; at the same concentrations, leaf elongation rate of the two fast growing species was only slightly inhibited. All responses were observed within 20 min after ethylene applications. Although ethylene generally inhibits growth, our results show that in some species, it may actually stimulate growth. Moreover, in the two slow-growing Poa spp., both growth stimulation and inhibition occurred in a narrow ethylene concentration range, and this effect was associated with a much lower ethylene emission. These findings suggest that the regulation of ethylene production rates and perception of the gas may be more crucial during leaf expansion of these species under non-stressful conditions and that endogenous ethylene concentrations are not large enough to saturate leaf growth responses. In the two fast-growing species, a comparatively higher ethylene endogenous concentration may conversely be present and sufficiently high to saturate leaf elongation responses, invariably leading to growth inhibition. PMID- 12114592 TI - Transfer specificity of detergent-solubilized fenugreek galactomannan galactosyltransferase. AB - The current experimental model for galactomannan biosynthesis in membrane-bound enzyme systems from developing legume-seed endosperms involves functional interaction between a GDP-mannose (Man) mannan synthase and a UDP-galactose (Gal) galactosyltransferase. The transfer specificity of the galactosyltransferase to the elongating mannan chain is critical in regulating the distribution and the degree of Gal substitution of the mannan backbone of the primary biosynthetic product. Detergent solubilization of the galactosyltransferase of fenugreek (Trigonella foenum-graecum) with retention of activity permitted the partial purification of the enzyme and the cloning and sequencing of the corresponding cDNA with proof of functional identity. We now document the positional specificity of transfer of ((14)C)Gal from UDP-((14)C)Gal to manno oligosaccharide acceptors, chain lengths 5 to 8, catalyzed by the detergent solubilized galactosyltransferase. Enzymatic fragmentation analyses of the labeled products showed that a single Gal residue was transferred per acceptor molecule, that the linkage was (1-->6)-alpha, and that there was transfer to alternative Man residues within the acceptor molecules. Analysis of the relative frequencies of transfer to alternative Man residues within acceptor oligosaccharides of different chain length allowed the deduction of the substrate subsite recognition requirement of the galactosyltransferase. The enzyme has a principal recognition sequence of six Man residues, with transfer of Gal to the third Man residue from the nonreducing end of the sequence. These observations are incorporated into a refined model for enzyme interaction in galactomannan biosynthesis. PMID- 12114593 TI - Influence of the diadinoxanthin pool size on photoprotection in the marine planktonic diatom Phaeodactylum tricornutum. AB - The pool size of the xanthophyll cycle pigment diadinoxanthin (DD) in the diatom Phaeodactylum tricornutum depends on illumination conditions during culture. Intermittent light caused a doubling of the DD pool without significant change in other pigment contents and photosynthetic parameters, including the photosystem II (PSII) antenna size. On exposure to high-light intensity, extensive de epoxidation of DD to diatoxanthin (DT) rapidly caused a very strong quenching of the maximum chlorophyll fluorescence yield (F(m), PSII reaction centers closed), which was fully reversed in the dark. The non-photochemical quenching of the minimum fluorescence yield (F(o), PSII centers open) decreased the quantum efficiency of PSII proportionally. For both F(m) and F(o), the non-photochemical quenching expressed as F/F' - 1 (with F' the quenched level) was proportional to the DT concentration. However, the quenching of F(o) relative to that of F(m) was much stronger than random quenching in a homogeneous antenna could explain, showing that the rate of photochemical excitation trapping was limited by energy transfer to the reaction center rather than by charge separation. The cells can increase not only the amount of DT they can produce, but also its efficiency in competing with the PSII reaction center for excitation. The combined effect allowed intermittent light grown cells to down-regulate PSII by 90% and virtually eliminated photoinhibition by saturating light. The unusually rapid and effective photoprotection by the xanthophyll cycle in diatoms may help to explain their dominance in turbulent waters. PMID- 12114595 TI - Sharing agriculture's genetic bounty. PMID- 12114594 TI - Differential messenger RNA gradients in the unicellular alga Acetabularia acetabulum. Role of the cytoskeleton. AB - The unicellular green alga Acetabularia acetabulum has proven itself to be a superior model for studies of morphogenesis because of its large size and distinctive polar morphology. The giant cell forms an elongated tube (a stalk of up to 60 mm in length), which at its apical pole makes whorls of hairs, followed by one whorl of gametophores in the shape of a cap. At its basal pole, the cell extends into a rhizoid wherein the single nucleus is positioned. In this study, we have determined the level of specific messenger RNAs in the apical, middle, and basal regions using reverse transcriptase-PCR methodology. Four mRNA classes were distinguished: those that were uniformly distributed (small subunit of Rubisco, actin-1, ADP-glucose, centrin, and alpha- and beta-tubulin), those that expressed apical/basal (calmodulin-4) or basal/apical gradients (calmodulin-2 and a Ran-G protein), and those with development-specific patterns of distribution (mitogen-activated protein kinase, actin-2, and UDP-glucose-epimerase). Restoration of the apical/basal calmodulin-4 mRNA gradient after amputation of the apical region of the cell requires the nucleus and was abolished by cytochalasin D. Accumulation of actin-1 mRNA in the vicinity of the wound set by the amputation needs, likewise, the presence of the nucleus and was also inhibited by cytochalasin. This suggests that actin microfilaments of the cytoskeleton are involved in directed transport and/or anchoring of these mRNAs. PMID- 12114596 TI - Public health. Agreement unlocks loan for TB and AIDS treatment in Russia. PMID- 12114597 TI - Cancer risk. Nudge from Congress prompts NCI review. PMID- 12114598 TI - Paleoanthropology. First member of human family uncovered. PMID- 12114599 TI - Public health. AIDS researcher named CDC chief. PMID- 12114600 TI - Mathematics: NSF to double number of math institutes. PMID- 12114601 TI - Virology. Active poliovirus baked from scratch. PMID- 12114602 TI - Embryonic stem cells. Stem cells not so stealthy after all. PMID- 12114603 TI - Solar system origins. Diamond dust dearth raises doubts. PMID- 12114605 TI - NINDS delves into drug development. PMID- 12114604 TI - Neuroscience. Animal studies raise hopes for spinal cord repair. PMID- 12114606 TI - Geoscience. Data dilemma: stow it, or kiss it goodbye. PMID- 12114608 TI - High-energy physics. Shadowy 'weak force' steps into the light. PMID- 12114607 TI - Research management. Big facilities account is big headache for NSF. PMID- 12114609 TI - Human genetics. A rational view of insurance and genetic discrimination. PMID- 12114610 TI - Human genetics. Before it's too late--addressing fear of genetic information. PMID- 12114611 TI - Cell biology. PARP-1--a perpetrator of apoptotic cell death? PMID- 12114612 TI - Microbiology. A binding contract for anthrax. PMID- 12114613 TI - Paleoclimate. The glacial tropical Pacific--not just a west side story. PMID- 12114614 TI - Development. Sociogenomics takes flight. PMID- 12114615 TI - Applied physics. Squeezing x-ray photons. PMID- 12114616 TI - Paleoclimate. Earth's long-term memory. PMID- 12114617 TI - Another phocine distemper outbreak in Europe. PMID- 12114618 TI - Super ENSO and global climate oscillations at millennial time scales. AB - The late Pleistocene history of seawater temperature and salinity variability in the western tropical Pacific warm pool is reconstructed from oxygen isotope (delta18O) and magnesium/calcium composition of planktonic foraminifera. Differentiating the calcite delta18O record into components of temperature and local water delta18O reveals a dominant salinity signal that varied in accord with Dansgaard/Oeschger cycles over Greenland. Salinities were higher at times of high-latitude cooling and were lower during interstadials. The pattern and magnitude of the salinity variations imply shifts in the tropical Pacific ocean/atmosphere system analogous to modern El Nino-Southern Oscillation (ENSO). El Nino conditions correlate with stadials at high latitudes, whereas La Nina conditions correlate with interstadials. Millennial-scale shifts in atmospheric convection away from the western tropical Pacific may explain many paleo observations, including lower atmospheric CO2, N2O, and CH4 during stadials and patterns of extratropical ocean variability that have tropical source functions that are negatively correlated with El Nino. PMID- 12114619 TI - El Nino-like pattern in ice age tropical Pacific sea surface temperature. AB - Sea surface temperatures (SSTs) in the cold tongue of the eastern equatorial Pacific exert powerful controls on global atmospheric circulation patterns. We examined climate variability in this region from the Last Glacial Maximum (LGM) to the present, using a SST record reconstructed from magnesium/calcium ratios in foraminifera from sea-floor sediments near the Galapagos Islands. Cold-tongue SST varied coherently with precession-induced changes in seasonality during the past 30,000 years. Observed LGM cooling of just 1.2 degrees C implies a relaxation of tropical temperature gradients, weakened Hadley and Walker circulation, southward shift of the Intertropical Convergence Zone, and a persistent El Nino-like pattern in the tropical Pacific. This is contrasted with mid-Holocene cooling suggestive of a La Nina-like pattern with enhanced SST gradients and strengthened trade winds. Our results support a potent role for altered tropical Pacific SST gradients in global climate variations. PMID- 12114620 TI - Two-dimensional x-ray waveguides and point sources. AB - We show that resonant coupling of synchrotron beams into suitable nanostructures can be used for the generation of coherent x-ray point sources. A two dimensionally confining x-ray waveguide structure has been fabricated by e-beam lithography. By shining a parallel undulator beam onto the structure, a discrete set of resonant modes can be excited in the dielectric cavity, depending on the two orthogonal coupling angles between the beam and the waveguide interfaces. The resonant excitation of the modes is evidenced from the characteristic set of coupling angles as well as the observed far-field pattern. The x-ray nanostructure may be used as coherent x-ray point sources with a beam cross section in the nanometer range. PMID- 12114621 TI - Spin-polarized resonant tunneling in magnetic tunnel junctions. AB - Insertion of a thin nonmagnetic copper Cu(001) layer between the tunnel barrier and the ferromagnetic electrode of a magnetic tunnel junction is shown to result in the oscillation of the tunnel magnetoresistance as a function of the Cu layer thickness. The effect is interpreted in terms of the formation of spin-polarized resonant tunneling. The amplitude of the oscillation is so large that even the sign of the tunnel magnetoresistance alternates. The oscillation period depends on the applied bias voltage, reflecting the energy band structure of Cu. The results are encouraging for the development of spin-dependent resonant tunneling devices. PMID- 12114622 TI - Spontaneous organization of single CdTe nanoparticles into luminescent nanowires. AB - Nanoparticles of CdTe were found to spontaneously reorganize into crystalline nanowires upon controlled removal of the protective shell of organic stabilizer. The intermediate step in the nanowire formation was found to be pearl-necklace aggregates. Strong dipole-dipole interaction is believed to be the driving force of nanoparticle self-organization. The linear aggregates subsequently recrystallized into nanowires whose diameter was determined by the diameter of the nanoparticles. The produced nanowires have high aspect ratio, uniformity, and optical activity. These findings demonstrate the collective behavior of nanoparticles as well as a convenient, simple technique for production of one dimensional semiconductor colloids suitable for subsequent processing into quantum-confined superstructures, materials, and devices. PMID- 12114623 TI - Role for stearoyl-CoA desaturase-1 in leptin-mediated weight loss. AB - Leptin elicits a metabolic response that cannot be explained by its anorectic effects alone. To examine the mechanism underlying leptin's metabolic actions, we used transcription profiling to identify leptin-regulated genes in ob/ob liver. Leptin was found to specifically repress RNA levels and enzymatic activity of hepatic stearoyl-CoA desaturase-1 (SCD-1), which catalyzes the biosynthesis of monounsaturated fatty acids. Mice lacking SCD-1 were lean and hypermetabolic. ob/ob mice with mutations in SCD-1 were significantly less obese than ob/ob controls and had markedly increased energy expenditure. ob/ob mice with mutations in SCD-1 had histologically normal livers with significantly reduced triglyceride storage and VLDL (very low density lipoprotein) production. These findings suggest that down-regulation of SCD-1 is an important component of leptin's metabolic actions. PMID- 12114624 TI - Multiple roles of Arabidopsis VRN1 in vernalization and flowering time control. AB - Arabidopsis VRN genes mediate vernalization, the process by which a long period of cold induces a mitotically stable state that leads to accelerated flowering during later development. VRN1 encodes a protein that binds DNA in vitro in a non sequence-specific manner and functions in stable repression of the major target of the vernalization pathway, the floral repressor FLC. Overexpression of VRN1 reveals a vernalization-independent function for VRN1, mediated predominantly through the floral pathway integrator FT, and demonstrates that VRN1 requires vernalization-specific factors to target FLC. PMID- 12114625 TI - Generation of self-incompatible Arabidopsis thaliana by transfer of two S locus genes from A. lyrata. AB - Transitions from cross-fertilizing to self-fertilizing mating systems have occurred frequently in natural and domesticated plant populations, but the underlying genetic causes are unknown. We show that gene transfer of the stigma receptor kinase SRK and its pollen-borne ligand SCR from one S-locus haplotype of the self-incompatible and cross-fertilizing Arabidopsis lyrata is sufficient to impart self-incompatibility phenotype in self-fertile Arabidopsis thaliana, which lacks functional orthologs of these genes. This successful complementation demonstrates that the signaling cascade leading to inhibition of self-related pollen was maintained in A. thaliana. Analysis of self-incompatibility will be facilitated by the tools available in this species. PMID- 12114626 TI - Evolution of the gene network underlying wing polyphenism in ants. AB - Wing polyphenism in ants evolved once, 125 million years ago, and has been a key to their amazing evolutionary success. We characterized the expression of several genes within the network underlying the wing primordia of reproductive (winged) and sterile (wingless) ant castes. We show that the expression of several genes within the network is conserved in the winged castes of four ant species, whereas points of interruption within the network in the wingless castes are evolutionarily labile. The simultaneous evolutionary lability and conservation of the network underlying wing development in ants may have played an important role in the morphological diversification of this group and may be a general feature of polyphenic development and evolution in plants and animals. PMID- 12114627 TI - Evolution and development of sex differences in cooperative behavior in meerkats. AB - In cooperatively breeding birds, where helpers of both sexes assist with the provisioning and upbringing of offspring who are not their own, males tend to contribute more than females to rearing young. This sex difference has been attributed to paternity uncertainty, but could also occur because males contribute more where they are likely to remain and breed in their group of origin. In contrast to most birds, female meerkats (Suricata suricatta) are more likely to breed in their natal group than males. We show that female meerkat helpers contribute more to rearing young than males and that female helpers feed female pups more frequently than males. Our results suggest that sex differences in cooperative behavior are generated by sex differences in philopatry and occur because females derive greater direct benefits than males from raising recruits to their natal group. These findings support the view that direct, mutualistic benefits are important in the evolution of specialized cooperative behavior. PMID- 12114628 TI - Leg patterning driven by proximal-distal interactions and EGFR signaling. AB - wingless and decapentaplegic signaling establishes the proximal-distal axis of Drosophila legs by activating the expression of genes such as Distalless and dachshund in broad proximal-distal domains during early leg development. However, here we show that wingless and decapentaplegic are not required throughout all of proximal-distal development. The tarsus, which has been proposed to be an ancestral structure, is instead defined by the activity of Distalless, dachshund, and a distal gradient of epidermal growth factor receptor (EGFR)-Ras signaling. Our results uncover a mechanism for appendage patterning directed by genes expressed in proximal-distal domains and possibly conserved in other arthropods and vertebrates. PMID- 12114629 TI - Mediation of poly(ADP-ribose) polymerase-1-dependent cell death by apoptosis inducing factor. AB - Poly(ADP-ribose) polymerase-1 (PARP-1) protects the genome by functioning in the DNA damage surveillance network. PARP-1 is also a mediator of cell death after ischemia-reperfusion injury, glutamate excitotoxicity, and various inflammatory processes. We show that PARP-1 activation is required for translocation of apoptosis-inducing factor (AIF) from the mitochondria to the nucleus and that AIF is necessary for PARP-1-dependent cell death. N-methyl-N'-nitro-N nitrosoguanidine, H2O2, and N-methyl-d-aspartate induce AIF translocation and cell death, which is prevented by PARP inhibitors or genetic knockout of PARP-1, but is caspase independent. Microinjection of an antibody to AIF protects against PARP-1-dependent cytotoxicity. These data support a model in which PARP-1 activation signals AIF release from mitochondria, resulting in a caspase independent pathway of programmed cell death. PMID- 12114631 TI - Computer-Assisted Microscope Analysis of Feulgen-Stained Nuclei in Gonadotroph Adenomas and Null-Cell Adenomas of the Pituitary Gland. AB - The current classification of clinically nonfunctioning pituitary adenomas is based on immunocytochemical and ultrastructural studies. However, a number of cases have less distinctive features that cannot be easily conformed with the prevailing morphologic classifications. The diagnostic information contributed by the determination of the nuclear DNA content (DNA ploidy level) and quantitative chromatic pattern description as opposed to the morphofunctional diagnosis in clinically nonfunctioning adenomas was consequently investigated in a series of 71 pituitary adenomas, including 31 null-cell adenomas, 35 gonadotropin adenomas, and 5 nonfunctioning adenomas that were not examined by electron microscopy. DNA ploidy level (8 variables) and quantitative chromatin pattern description (30 variables) were carried out by means of the computer-assisted microscope analysis of 80-1600 Feulgen-stained nuclei analyzed/case. The diagnostic information contributed by the 38 quantitative variables was determined by multifactorial statistical analysis (i.e., Discriminant Analysis). This computer-assisted classification significantly differentiated nulLcell adenomas from gonadotropin adenomas (p = 0.0025). In addition, it was able to differentiate three major subtypes of nonfunctioning adenomas on the basis of their immunohistochemical profiles. These were the immunonegative adenomas, the follicle-stimulating hormone (FSH)-positive adenomas, and the a-subunit (a) and/or luteiwizing hormone (LH)-positive adenomas (p < 0.0001 to p < 0.001). We thus suggest that the cytometric image analysis of Feulgen-stained nuclei can contribute on the discrimination of subtypes of clinically nonfunctioning pituitary adenomas. PMID- 12114633 TI - Endothelial Regulation of Vascular Repair: Role of bFGF in Paracrine Pathways. AB - Vascular repair following injury is mediated by both endothelial and smooth muscle cells often through paracrine pathways. Basic fibroblast growth factor (bFGF) is present at sites of vascular injury. The role of bFGF in regulating reendothelialization through an effect on centrosome redistribution in cell migration is discussed. The role of bFGF in neointimal formation, especially as it relates to smooth muscle cell proliferation, is reviewed. It is concluded that bFGF appears to be an important agent regulating the early responses of the artery wall to injury. PMID- 12114630 TI - Pituitary Sex Steroid Receptors: Localization and Function. AB - The pituitary contains estrogen receptor (ER), progesterone receptor (PR), and androgen receptor (AR). In accordance with immunocytochemistry, it is agreed that sex hormone receptors reside into the nucleus. All three receptors are found predominantly in gonadotrophs and lactotrophs, and less frequently in other cell types. ER plays a major role in prolactin (PRL) production and lactotroph proliferation, and protracted estrogen administration induces lactotroph hyperplasia and adenoma in rodents. Most research on PR and AR is focused on their role in the fine-tuning of gonadotropin secretion during estrous cycle. Contrary to the effect in nontumorous pituitary, estrogens can inhibit the proliferation of transplantable rat pituitary tumors and of cell lines derived from them. In humans, despite the presence of ER in all types of adenohypophysial tumors, the role of estrogen in tumor cell proliferation is still unclear. Few results indicate that tumor growth is stimulated by estrogen, and inhibited by progesterone and androgen. Novel data reveal that steroid hormones can act directly on plasma membrane or via other receptors, and interact with growth factors, oncogenes, and other transcription factors. The mechanisms by which steroid hormones control cell proliferation remain a major challenge for future research. PMID- 12114632 TI - Growth Factors. AB - Growth factors are polypeptides that interact with specific cellular receptors leading to many different biological responses. There are various families of growth factors that have similar biochemical structures. Although many growth factors stimulate cell proliferation, a few have primarily inhibitory functions, such as transforming growth factor-p (TGFB). Growth factors regulate various modes of action of endocrine tissues, including autocrine, paracrine, and endocrine functions. Recent studies have shown that growth factors also regulate various proteins in the cell cycle, and may have a direct or indirect effect on cell proliferation. For example, TGFB regulates various inhibitory cell-cycle proteins, including p27(kip1) and p15(INK4B). Molecular analyses of growth factors, including the cloning and sequencing of specific growth factor receptors, have contributed greatly to our understanding of the role these factors play in cellular homeostasis and neoplastic development. PMID- 12114634 TI - Molecular Pathology of the Pituitary Adenomas. AB - Molecular techniques have been applied to the study of pituitary adenomas by many investigators. We have used nonisotopic in situ hybridization (ISH) to analyze pituitary adenomas in our laboratory. These studies have provided insight into mRNA production by various adenomas, and have contributed toward our new proposed clinical and cytofunctional classification of pituitary adenomas. PMID- 12114635 TI - Paraganglioma-Like Adenomas of the Thyroid (PLAT): Incidental Lesions with Unusual Features in a Patient with Nodular Goiter. AB - We discovered two well-demarcated nodules incidentally in a thyroid removed because of a nodular goiter. Histologically, the nodules showed a pattern of paraganglioma or so-called paraganglioma-like adenoma of the thyroid (PLAT), with lobules of polygonal and oval cells in a vascular stroma, but the immunohistochemical markers typical of paraganglioma, including chromogranin, synaptophysin, Leu 7 and 5-100, and thyroglobulin, characteristic of PLAT, were negative in the tumor cells. C-cell markers calcitonin and somatostatin were also negative. Stain for neuron-specific enolase (NSE), however, showed a distinctive pattern of reactivity within cells at the periphery of the lobules, whereas all tumor cells stained positively for keratins. Stain for carcinoembryonic antigen showed a focal interstitial pattern that corresponded to small intercellular spaces filled by microvilli identified ultrastructurally. This pattern of immunohistochemical staining has not been previously described in paraganglioma or in PLAT, and may have implications about the origin and nature of these controversial entities. PMID- 12114636 TI - Multiple Endocrine Neoplasms in a Patient with AL Amyloidosis-Associated Plasma Cell Dyscrasia. AB - Reports on patients with systemic amyloidosis-associated plasma cell dyscrasia (PCD) who have multiple endocrine tumors are very rare. Here we describe such a case. The patient was a 74-yr-old man with amyloid light-chain (AL) amyloidosis associated PCD who had a null-cell adenoma of the pituitary and a latent papillary carcinoma of the thyroid gland. as well as a tubular adenoma of the sigmoid colon. The amyloid protein deposits reacted with the antibody to the n immunoglobulin light chain. Since PCD by itself may be a risk factor for the development of subsequent neoplasms, the careful clinical evaluation of PCD patients is recommended. PMID- 12114637 TI - Mediastinal Parathyroid Tumor: Giant Adenoma or Carcinoma? AB - A 71-yr-old woman presented with progressive weakness, bone pain, polydipsia, and epigastric pain. Laboratory studies established the diagnosis of primary hyperparathy roidism with excessively elevated levels of parathyrod hormone (PTH) complicated by renal failure and anemia. Preoperative imaging using (99m)technetium hexakis 2-methoxy-isobutylisonitrile (MIBI) demonstrated an area of intense uptake in the mediastinum, which on T(2) -weighted magnetic resonance imaging revealed the presence of a hyperintense mediastinal mass of 25 mm in diameter adjacent to the ascending aorta Surgical exploration and resection of the mass were performed, and histological examination of the tumor revealed solid masses of epithelial cells growing in a trabecular pattern, hyaline bands, and low mitotic activity. Immunohistochemical staining of the tumor specimens using monoclonal mouse antihuman antibodies revealed markedly positive immunoreactivity of tumor cells for p53 protein and absence of nuclear immunoreactivity for the retinoblastoma tumor-suppressor protein, findings consistent with parathyroid carcinoma. Improved imaging techniques and advances in molecular pathology of parathyroid disorders may help to improve clinical management of patients with parathyroid neoplasia. PMID- 12114640 TI - Deficiency of Phenylethanolamine N-Methyltransferase in Norepinephrine-Producing Pheochromocytoma. AB - Pheochromocytoma is a catecholamine (CA)-producing tumor that is classified into two types: the norepinephrine (NE) and the mixed NE and epinephrine type (E-type) from plasma CA levels. Phenylethanolamine N-methyltransferase (PNMT) is the terminal enzyme in CA synthesis; it catalyzes the synthesis of E from NE. It is not known whether the absence of immunoreactive PNMT is paralleled by a lack of PNMT mRNA. The mRNA of tyrosine hydroxylase (TI-I) and PNMT in seven pheochromocytomas, five NE-type and two E-type tumors, were examined by Northern blot analysis and in situ hybridization (ISH) technique. TH mRNA was detected in all tumors but PNMT mRNA was limited only to the E-type tumors. In addition to our previous immunohistochemical study of 70 pheochromocytomas and paragangliomas in which all pheochromocytomas had cells immunoreactive to TH, but PNMT was expressed only in E-type, we concluded that NE-type pheochromocytoma lacks PNMT both at the mRNA and protein levels, resulting in an inability to produce E. The essential difference between NE-type and E-type pheochromocytoma is that the NE type lacks PNMT. PMID- 12114639 TI - Possible Relation of p53 and mdm-2 Oncoprotein Expression in Thyroid Carcinoma: A Molecular-Pathological and Immunohistochemical Study on Paraffin-Embedded Tissue. AB - Routinely processed tissues from a series of benign and malignant thyroid lesions were immunohistochemically investigated with antibodies against p53 and mdm-2. p53 was immunolocalized in <10% of nuclei in 2/80 nodular goiters, 2/60 follicular adenomas, 26/68 follicular carcinomas, 7/40 papillary carcinomas, 3/10 "insular" carcinomas, and 10/31 anaplastic carcinomas. More than 10% positively stained nuclei were found in 2 widely invasive follicular, 2 insular, and 15 anaplastic carcinomas. All p53-positive cases showed a concomitant immunohistochemical mdm-2 expression; an immunohistochemical colocalization on serial section was demonstrated in 12 anaplastic carcinomas. Screening by polymerase chain reaction single-strand conformation polymorphism (PCR-SSCP) analysis of these 12 cases revealed no relevant mutations in the coding regions of exons 2-11 of the p53 gene. Additionally, 1 follicular adenoma, 6 follicular carcinomas (4 minimally and 2 widely invasive), 1 papillary, and 2 poorly differentiated insular carcinomas were mdm-2 positive without immunohistochemically detectable p53 expression. These results provide evidence that wild-type p53 expression in thyroid carcinomas may be associated with mdm-2 induced formation of stable complexes. However, the role of p53 mutations and p53 protein inactivation owing to other factors (e.g., mdm-2) in the progression of thyroid carcinomas is still poorly understood. PMID- 12114638 TI - Proliferation Markers and Their Uses in the Study of Endocrine Tumors. AB - A growing body of literature supports the view that the proliferative activity (PA) of tumor cells is an important prognostic indicator for a variety of different tumors. We examined the role of PA in diagnosis and prediction or malignancy of endocrine tumors (ETs) of pituitary gland, pancreas, thyroid, parathyroid glands, adrenal glands, paraganglia, gastroenteric tract, and lung. The data in the literature indicate that the assessment of PA is not a diagnostic indicator of malignancy especially at the individual case level, whereas it can be useful for identifying subsets of malignant tumors with different aggressiveness potential, as well as for choosing therapeutic options in metatstatic lesions. We hope that, in the near future, multiparametric approaches including PA markers, cell growth and differentiation factors, and oncogenes will yield valuable information for diagnosis and prognosis of ETs also in individual cases. PMID- 12114641 TI - Amino Acid Composition of Proteins Extracted from Endemic Goiter Glands. AB - Proteins were isolated and characterized in thyroid tissue of six patients from Brazil with endemic goiter. Biochemical studies of these thyroidal proteins included gel filtration, electrophoresis, and amino acid analysis. In addition to thyroglobulin, two of the most abundant proteins found in all goiters studied had molecular weights of 68 and 14 kDa. One protein was identified as albumin based on its immunoreactivity with antibodies to serum albumin. The other protein was identified as a hemoglobin subunit using reverse-phase high-performance liquid chromatography (H PLC). Identification was confirmed by partial N-terminal amino acid sequencing that showed several nonconservative differences in thyroid albumin when compared to human serum albumin (HSA). Biochemical findings were correlated with iodine and hormone contents in serum and thyroglobulin from these patients. Based on these findings, we suggest that hemoglobin and albumin are taken up from the blood by the hyperplastic thyroid tissue. Albumin would be processed by the thyroid follicular cell, becoming iodinated and released into the circulation. PMID- 12114642 TI - The Immunophenotype of Pituitary Adenomas. AB - Although the production of pituitary hormones by adenohypophysial tumors has been studied extensively, an examination of the immunophenotype of pituitary adenomas using a broad spectrum of antibodies has not been previously investigated. We studied 23 pituitary adenomas using a large panel of antibodies to determine if these tumors exhibited a common immunophenotype. Various neuroendocrine markers, synaptophysin, neuron-specific enolase (NSE), and the intermediate filament protein, low-mol-wt keratin were expressed in most examples. There was, however, differential expression of chromogranin A in that few prolactin (PRL) and adenocorticotrophic hormone (ACTH) adenomas stained positively, whereas all other adenoma subtypes were reactive. The ACTH adenomas had a unique profile with positive staining for galanin, neurophysiri, vasopressin, and ubiquitin. These results indicate that (1) pituitary adenomas do not express a single "generic" immunophenotype; (2) synaptophysin is the most reliable and best broad spectrum marker for pituitary adenomas; (3) the neuroendocrine granule marker chromogranin A is useful in the identification of null cell adenoma, a tumor that usually does not stain for anterior pituitary tumors; and (4) among pituitary tumors, ACTH adenomas have a unique immunoprofile. PMID- 12114643 TI - Vanishing Psammoma Bodies in the Anterior Pituitary of the Human Newborn: An Immunohistochemical and Histometric Study. AB - Adenohypophyses of human newborns contain characteristic psammoma bodies. Their numbers are maximal within 2 weeks of the neonatal period and diminish thereafter. They are very rare in infant pituitaries, seeming to disappear by shrinkage in that there is a significant direct correlation between their number and size. The bodies were found to contain a high concentration of endogenous peroxidase, thus suggesting that the enzyme may be responsible for their disappearance. A statistical majority of psammoma bodies were located within follicular lumens. By immunohistochemistry, the follicular epithelium surrounding psammoma bodies showed immunoreactivity for various pituitary hormones. Light microscopy demonstrated that adenohypophysial cells surrounding psammoma bodies contain randomly scattered granules or globules exhibiting peroxidase activity. Extrusion of such granules into follicular lumens may play a role in the genesis of the concretions. The conspicuous lamellar nature of the calcified psammoma bodies suggests that waves of calcium deposition occur during their morphogenesis. Despite histologic similarities, the histochemical characteristics of this type of psammoma body differ from those in other organs as well as from the calcification encountered in prolactin (PRL)-producing pituitary adenomas. PMID- 12114644 TI - The Anterior Pituitary in Hemochromatosis. AB - Pituitary changes in the case of a 69-year-old man with hemochromatosis are reported. The patient died of complications of hepatocellular carcinoma. The pituitary removed at autopsy was studied by histology, histochemistry, immunocytochemistry, electron microscopy, and X-ray diffraction. Preferential localization of iron deposits was demonstrated within gonadotrophs, which, at the ultrastructural level, displayed selective, severe cellular injury. X-ray diffraction revealed the deposition of iron-accumulated lysosomes. Iron storage also was noted in stellate cells. We consider selective injury of pituitary gonadotrophs to be the basis of hypogonadism in iron-overloaded states. PMID- 12114645 TI - Expression of the Neural Cell Adhesion Molecule NCAM by Peptide- and Steroid Producing Endocrine Cells and Tumors: Alternatively Spliced Forms and Polysialylation. AB - The adhesive properties of neural cell adhesion molecules (NCAMs) can be modified by alternative splicing of the primary transcript or by posttranslational modifications, such as sialylation. In this article, we describe distinct forms of alternative splicing and posttranslational modification of the extracellular domain of NCAM of various endocrine tissues and derived tumor cells of the rat and of steroid- and peptide-hormone producing endocrine cells in humans. NCAM-140 is the major isoform expressed in the rat adrenal gland, adenohypophysis, and in granulosa and granulosa-lutein cells. NCAM-180 is predominant in the neurohypophysis. Polysialylated NCAM is expressed in different endocrine tissues and tumor cells of the rat. Different amounts of NCAM mRNA containing the "extra exon" VASE at the exon 7/8 splice boundary were detected in endocrine cells of rats. Human granulosa cells in culture undergo luteinization. During this process, the VASE-containing NCAM isoform is supplemented by an alternatively spliced isoform without this insert. Thus, modifications of NCAM may be important for adhesive interactions in normal and neoplastic endocrine cells. In addition, the differential expression and the alternative splicing of NCAM during luteinization of granulosa cells raise the possibility that NCAM could be involved in folliculogenesis and the formation of the corpus luteum in humans. PMID- 12114646 TI - Smooth Muscle Cell Abnormalities Associated with Gastric ECL Cell Carcinoids. AB - The histologic and immunohistochemical study of 45 ECL cell gastric carcinoids and of the extratumoral gastric mucosa revealed four variants of smooth muscle cell abnormalities: (1) hypertrophy of muscularis mucosae trapped within the tumors, a finding occurring in 76.5% of cases; (2) proliferation of stromal smooth muscle cells originating from the muscularas mucosae and mostly associated with tumor invasion of the submucosa (seen in 93.9% of cases with abundant stromal component of the tumors); (3) occurrence of frequent, prominent aggregates of smooth muscle cells in the lamina propria of the antral (but not of the fundic) mucosa of the stomach (found in 41.7% of cases); and (4) increased thickness of the extratumoral muscularis mucosae in the fundic (but not in the antral) mucosa of patients with gastric carcinoids. In addition, localized muscle cell proliferation was also associated with foci of micronodular hyperplasia of endocrine cells in the extratumoral mucosa. These findings were neither observed in control cases of gastric adenocarcinoma, gastric peptic ulcer, and duodenal peptic ulcer (10 unselected cases from each group) nor were they observed in 10 subjects with normal gastric mucosa collected at autopsy. With the possible exception of the increased thickness of the extratumoral fundic muscularis mucosae, which may be influenced by the mucosal inflammatory process, it is suggested that the present findings represent a proliferative response of smooth muscle cells to basic fibroblastic growth factor whose production by gastric carcinoids and their precursor lesions has recently been demonstrated. PMID- 12114647 TI - Vasculature in Nontumorous Hypophyses, Pituitary Adenomas, and Carcinomas: A Quantitative Morphologic Study. AB - Vascular supply is essential for tumor proliferation and metastasis formation. Correlation was noted between vascular density and tumor size as well as metastases in several tumor types. The aim of the present study was to assess vascular density in nontumorous hypophyses, pituitary adenomas, primary pituitary carcinomas, and carcinomas metastatic to the pituitary. Twenty nontumorous hypophyses, 87 endocrinologically active or inactive pituitary adenomas, 8 primary pituitary carcinomas, 8 metastatic carcinomas, and 10 randomly selected noninvasive and 6 invasive adenomas were included in the study. Tissues were fixed in formalin, embedded in paraffin, cut, stained with hematoxylin and eosin, PAS, and immunostained for adenohypophysial hormones as well as Factor VIII related antigen using the streptavidin-biotin-peroxidase complex method. Four counts were performed: percentage of capillary area, number of vessels per field, percentage of endothelial cells, and number of endothelial cells per field. The results show that pituitary adenomas have significantly lower vascular densities as compared to nontumorous adenohypophyses. Prolactin-producing adenomas removed from untreated patients have the highest counts and growth hormone-producing adenomas the lowest counts. However, the observed differences among adenoma types are not of statistical significance. No differences are noted between noninvasive and invasive tumors. Primary pituitary carcinomas show no significant increase in vascular densities. Some metastatic tumors exhibit high vascularity. It can be concluded that pituitary adenomas have a limited capacity to induce angiogenesis. Lack of significant angiogenesis may play a role in the slow pace of pituitary tumor growth and rarity of metastases. PMID- 12114648 TI - Basic Fibroblast Growth Factor Expression by Two Prolactin and Thyrotropin Producing Pituitary Adenomas. AB - We investigated the expression of basic fibroblast growth factor (bFGF) in an aggressive type of PRL and TSH-producing pituitary adenoma. Immunocytochemistry and electron microscopy were used to characterize the tumors removed from two patients. lmmunoassays were used to measure hormone and bFGF levels in vitro and in vivo. Reverse transcription-polymerase chain reaction (RT-PCR) was used to detect bFGF mRNA expression by these tumors. Morphologically, these tumors were characterized by an unusual plurihormonal pattern with expression of PRL and TSH and the ultrastructural characteristics of the silent subtype 3 adenoma; in addition, both adenomas displayed marked interstitial fibrosis. bFGF was measurable in the circulation of these patients ranging from 7.5-20.5 pg/mL (normal < 1 pg/mL). bFGF concentrations were reduced following surgical adenomectomy. bFCF in culture media was present in concentrations of 197-387 pg/24h/10(5) cells. bFGF mRNA expression was identified in both adenomas examined. bFGF levels were unaltered in the culture media and in the serum of patients following GnRH and TRH treatment. In conclusion, the expression of bFGF by these plurihormonal pituitary adenomas suggests the possibility that it may play a role in the development of fibrosis and tumor cell proliferation of this unusual type of pituitary neoplasm. PMID- 12114649 TI - Amyloid Deposits in Pituitaries and Pituitary Adenomas: Immunohistochemistry and In Situ Hybridization. AB - The patterns of deposition and immunoreactivity of interstitial amyloid were studied in 11 pituitary glands obtained at autopsy and 9 surgically resected pituitary adenomas using Congo red staining and a panel of antisera directed against 5 major amyloid fibril proteins and all pituitary hormones. The deposition pattern of amyloid in pituitary glands differed from that in adenomas but all amyloid deposits showed an immunostaining with anti-amyloid X-light chain. The remaining antisera were immunonegative. In situ hybridization using an oligodeoxyribonucleotide-probe complementary to the mRNA coding for the constant region of human X-light chain yielded no hybridization signals in the pituitaries or pituitary adenomas, excluding local synthesis and secretion of immunoglobulins. Since no case studied suffered from generalized AX-amyloidosis and adsorption of immunoglobulins to the unknown amyloid fribril protein of the pituitary seems to be unlikely, crossreaction of the polyclonal antisera with an undefined antigen is probable. The similar immunostaining properties of amyloid deposits in "normal" pituitaries and pituitary adenomas suggest they both originate from the same precursor protein. PMID- 12114650 TI - Immunoreactive Characteristics and Classification of Hyperparathyroidism. AB - Hyperparathyroidism is caused mainly by three different conditions: namely, secondary hyperplasia, primary hyperplasia, and adenoma with only a few cases due to carcinomas. Histological distinction among these diseases is still difficult. In an attempt to characterize the three conditions, 17 cases from patients with hyperparathyroidism and 12 with normal glands were investigated immunohistochemically using antibodies against PTH, PTHrP, Ki-67 (MIB-1), and chromogranin A. The normal glands showed a diffuse staining pattern for PTH, and focal staining for PTHrP and for chromogranin A. Secondary hyperplasia demonstrated either focal, diffuse, or mixed staining in one gland with the three antibodies. For the primary hyperplasias and adenomas, the cases could be divided into two groups. The first group (group I), including 1 case of primary hyperplasia and 3 cases of adenoma, showed a homogeneous staining pattern with all three antibodies. A heterogeneous staining pattern similar to secondary hyperplasia was found in the other 8 cases that formed the second group (group II). There were five types of cytologic staining patterns after immunostaining. In secondary hyperplasia and group II, several patterns appeared simultaneously. On the contrary, only one pattern was found in group I. The proliferative index (P1) from Ki-67 staining of group I was also significantly higher than in group II. A lower P1 was observed in the normal glands. The present results indicate that different immunohistochemical characteristics exist in primary hyperplasias and in adenomas. PMID- 12114651 TI - Excessive Endothelial Cell Proliferation Occurring in an Organizing Thyroid Hematoma: Report of a Case and Review of the Literature. AB - The authors report a case of a thyroid hematoma containing an exuberant endothelial cell proliferation. No underlying vascular neoplasm was apparent and the hematoma occurred in a substernal goiter from a 90-yr-old male. Review of the literature revealed four similar previously reported cases of endothelial cell proliferation found in thyroid hematomas for a total of five cases. A fine needle aspiration or biopsy procedure antedated the definitive surgery in four patients and may be responsible for the formation of the hematoma in these cases. Areas of the endothelial proliferation resembled papillary endothelial hyperplasia (Masson's hemangioma) in four cases and cavernous hemangioma in three. However, these lesions occurred outside of vascular lumina and instead were intimately associated with fibrinous material of the hematoma. In addition, two cases contained foci of endothelial cells with nuclear atypia or intracytoplasmic lumens requiring distinction from angiosarcoma. These vigorous endothelial proliferations most likely represent alterations in the organization of a thyroid hematoma, a process similar to papillary endothelial hyperplasia, which is usually intravascular. Primary vascular neoplasms of the thyroid are exceedingly rare. Nevertheless, awareness of exuberant endothelial cell proliferation as a potential, albeit rare, manifestation of a thyroid hematoma should prompt careful attention to distinguish this reactive process from vascular neoplasms such as hemangioma or angiosarcoma. PMID- 12114652 TI - Lymphocytic Hypophysitis Presenting as a Pituitary Tumor in a 63-Year-Old Man. AB - This report describes a case of lymphocytic hypophysitis in a 63-year-old man who presented with symptoms of a pituitary mass lesion associated with hypothyroidism and hypogonadism. Postoperative endocrinologicaI testing demonstrated gonadotropic, thyrotropic, and corticotropic hypopituitarism, and the patient was commenced on replacement therapy with hydrocortisone and levothyroxine. Histological examination of the pituitary tissue obtained by transsphenoidal surgery revealed lymphocytic hypophysitis without evidence of a pituitary adenoma. The vast majority of patients with lymphocytic hypophysitis are women particularly during pregnancy and the puerperium. Until recently only four men were reported in the literature. The pathogenesis of lymphocytic hypophysitis is uncertain but autoimmune mechanisms are possibly involved. PMID- 12114655 TI - Proprotein-Processing Endoprotease Furin and its Substrate Parathyroid Hormone Related Protein Are Coexpressed in Insulinoma Cells. AB - Parathyroid hormone-related protein (PTHrP) is frequently produced in pancreatic endocrine tumors. PTHrP is synthesized as the precursor pro-PTHrP and undergoes a series of posttranslational processing reactions, among which cleavage of a 12 amino acid sequence from its precursor is crucial for the biological activation of PTHrP. This cleavage is catalyzed by furin, a proprotein-processing endoprotease that cleaves the consensus sequence -Arg-X-(Lys/Arg)-Arg-X-. We previously reported that furin is highly expressed in rat pancreatic islets during the perinatal stage and that the expression of furin in pancreatic-cells induces faster cell growth. From this, we postulated that furin may be co expressed with PTHrP in insulinomas. We immunostained insulin, PTHrP, and furin in 21 human pancreatic endocrine tumors: 10 insulinomas, 5 VIPomas, 4 gastrinomas, and 2 somatostatinomas. Of these 21 endocrine tumors, furin was positively stained in all 10 insulinomas. Likewise, PTHrP was detected in the same insulinomas. We found one VIPoma and one gastrinoma contained a few insulin positive cells scatteringly, which were also positive for furin and PTHrP. But other non-insulinoma endocrine tumors did not display furin and PTHrP positivity. We conclude that furin and its substrate pro-PTHrP are co-expressed specifically in insulinomas. PMID- 12114654 TI - Expression of Hepatocyte Growth Factor (HGF) and its Receptor (MET) in Medullary Carcinoma of the Thyroid. AB - The tyrosine kinase receptor encoded by the MET oncogene has the unusual property of mediating the invasive growth of epithelial cells upon binding with the hepatocyte growth factor (HGF). The MET/HGF receptor is known to be overexpressed in thyroid carcinomas originated from follicular cells, but has not been reported in C cell tumors. To investigate the role of HGF and of its receptor (encoded by MET oncogene) in medullary carcinoma of the thyroid (MCT), we studied the expression of HGF and MET in 20 cases by means of different techniques. By RT PCR, HGF mRNA was found in 10/20 cases, while MET mRNA presence was demonstrated in 8/20, of which 7 also expressed HGF. Northern blot analysis and in situ hybridization were performed in selected cases and confirmed RT-PCR data in some cases, although the lower sensitivity of these procedures did not allow the identification of all RT-PCR positive cases. By immunohistochemistry (using specific monoclonal antibodies) HGF was demonstrated in 8/9 RT-PCR positive cases and a monoclonal to MET immunostained 5/6 RT-PCR positive cases. All receptor positive cases also expressed the ligand in the same tumor cell population. These findings demonstrate MET and HGF co-expression in a subset of MCT, in which autocrine/paracrine circuits may be active. No correlation was found between HGF/MET expression and clinico-pathological parameters, except for the more common multifocality of HGF/MET positive MCT. Whether the potential activation of MET in MCT is responsible for local invasion and malignant evolution is to be further investigated, especially in multifocal and aggressive tumors. PMID- 12114656 TI - Octreotide Effect on Growth Hormone and Somatostatin Subtype 2 Receptor mRNAs of the Human Pituitary Somatotroph Adenomas. AB - Octreotide, a somatostatin analog used to treat acromegalic patients harboring a pituitary tumor, acts via somatostatin subtype 2 receptor (SSTR2) and causes significant decrease of circulating GH levels and sometimes mild to moderate tumor shrinkage. To further elucidate the mechanism of octreotide action, we studied GH and SSTR2 mRNAs by in situ hybridization in densely and sparsely granulated somatotroph adenomas removed by surgery from 14 treated and 14 untreated patients. Only in densely granulated adenomas were the GH and SSTR2 mRNA signals mildly decreased relative to untreated matched adenomas. The decrease of GH mRNA in densely granulated somatotroph adenomas suggests that they may have a more favorable response to octreotide therapy than sparsely granulated tumors. PMID- 12114653 TI - Hormonally Functional Ovarian Neoplasms. AB - Hormonally functional ovarian neoplasms are those tumors that secrete one or more hormones that are clinically manifested in the patient. The hormone production may have implications for the diagnosis, management or treatment of the patient. Hormonally functional ovarian neoplasms include tumors that belong to various histologic categories and produce a variety of hormonal effects. Functional ovarian tumors most commonly produce steroid hormones, and such tumors frequently belong in the sex cord-stromal and steroid cell categories. In addition, a wide variety of peptide hormones may be produced by ovarian tumors. Although in most instances the neoplastic cells themselves produce the hormones, a wide variety of tumors may induce their stroma to produce steroid hormones. The stroma of ovarian tumors is derived from the ovarian stroma and may, on occasion, resemble specialized ovarian stroma and its derivatives. Cells resembling luteinized stromal cells or luteinized theca cells may be present and appear to be responsible for the resultant hormone secretion. PMID- 12114659 TI - Columnar Cell Carcinoma of the Thyroid: MIB-1 Immunoreactivity as a Prognostic Factor. AB - We report a case of columnar cell carcinoma of the thyroid. A 47-year-old Japanese man had a nonencapsulated thyroid mass that infiltrated the surrounding tissues extensively. Seventeen months after thyroidectomy he died of respiratory failure resulting from tracheal invasion. An autopsy showed distant metastases to the liver, lung, esophagus, and pancreas. Histologically, the thyroid mass consisted of tall columnar atypical cells with marked nuclear stratification, About one-fifth of tumor cells were immunopositive for M18-1. The MIB-1 -positive index of our case was extremely high, compared with that of ordinary papillary carcinoma. This case indicates that biological growth activity in columnar cell carcinoma may be similar to that of undifferentiated carcinoma of the thyroid, since the MIB-1 -positive index is close to each other. PMID- 12114657 TI - From Endocrinology to Intracrinology. AB - Recently, in situ formation of active sex steroids at the sites of their actions from biologically inactive precursors in the circulation have been demonstrated to play very important roles in sex steroid-dependent neoplasms. These tissues in which the conversion occurs are designated as intracrine tissues and their mechanisms of actions can be designated as intracrinology in contrast to endocrinology. Aromatase, which converts serum androgens to estrone, and 17B hydroxysteroid dehydrogenase I, which is involved primarily in the conversion of estrone to estradiol, are two major enzymes which function in the in situ formation of biologically active estrogens from circulatory androgens. In human estrogen-dependent neoplasms, including breast, endometrioid endometrial, and common epithelial ovarian carcinoma, we recently demonstrated overexpression of aromatase, especially in stromal cells at sites of frank invasion possibly under a new promoter usage and that of 17B-hydroxysteroid dehydrogenase I in these carcinoma cells. These estrogen-dependent carcinomas are considered to have a common characteristic in estrogen metabolism (i.e., the expression of aromatase in the stromal cells and of 17B-hydroxysteroid dehydrogenase I in the epithelial cells). With these in situ mechanisms of generating biologically active estrogens from circulating androgens, that is, "intracrine manner," these estrogen dependent neoplasms can exert estrogenic actions on carcinoma cells despite low circulating serum estrogen levels, as observed in postmenopausal women. Evaluation of intracrine mechanisms can provide new insights into various estrogen-related biological phenomena in humans. PMID- 12114658 TI - Nonradioactive In Situ Hybridization: Recent Techniques and Applications. AB - In situ hybridization (ISH) has become a standard method for the localization of nucleic acid sequences in chromosomes, single cells, and tissue sections. Nonradioactive ISH has not only eliminated the problems associated with radioactive probes but has also achieved a higher degree of resolution. Advances in probe preparation and labeling methods have facilitated the general application of ISH. In combination with immunohistochemistry, ISH can provide histological information on gene activity at the DNA, mRNA, and protein levels. Some nonradioactive ISH can simultaneously detect nucleic acid sequences in the same tissue or in a chromosome spread. Advances in ISH technology, including use of the polymerase chain reaction offer both a high sensitivity allowing detection of low levels of gene expression and the cytological localization of gene sequences. PMID- 12114660 TI - Prevalence and Prognostic Significance of Neuroendocrine Differentiation in Colorectal Carcinomas. AB - The prognostic significance of neuroendocrine differentiation in colorectal carcinoma is uncertain. We analyzed 289 moderately differentiated (grades II and Ill) colorectal carcinomas for neuroendocrine differentiation by immunohistochemistry and in situ hybridization. The tumors were divided into three groups based on the presence of and the numbers of neuroendocrine cells, with group I having no neuroendocrine cells, group II having <1 positive cell/mm(2), and group Ill with >1 positive cell/mm(2). In situ hybridization with probes for chromogranin A and B detected almost twice as many neuroendocrine cells as did immunostaining with an antibody for chromogranin A. There was no prognostic difference associated with the presence or absence of neuroendocrine differentiation in this group of moderately differentiated carcinomas. These results indicate that the presence of neuroendocrine cells detected by expression of chromogranin protein or mRNA does not influence prognosis in moderately differentiated colorectal carcinomas. PMID- 12114661 TI - CD26 (Dipeptidyl Aminopeptidase IV) Expression in Normal and Diseased Human Thyroid Glands. AB - The objective of the present study was to investigate CD26 (dipeptidyl aminopeptidase IV) expression in normal and diseased thyroids and its relation to differentiation and cell proliferation. CD 26 was also evaluated as a possible marker of malignancy in thyroid neoplasias. A total of 38 normal thyroids and 117 diseased thyroids (neoplastic and non-neoplastic) were evaluated. CD26 and thyroglobulin (Tg) expression was determined by analyzing at least 200 cells/specimen. A minimum of 500 cells/specimen were counted to calculate the MIB 1-positive cell rate expressed as a percentage of total nucleated epithelial cells. CD26 expression was absent in all thyroids from fetuses and children. Among the adults, 7.1 % had CD26 expression only in oncocytic metaplastic areas. In 3 of the 7 elderly subjects, CD26 expression was present in 0.2-90% of epithelial cells. CD26 expression was observed in all diseased thyroids. Since this enzyme is also expressed in benign conditions, it is not useful as a marker of malignancy. There was no relationship between CD26 and Tg expression. The MIB 1-positive cell rate was found to be low for all kinds of thyroid tissues, and when the cell proliferation rate was analyzed according to CD26 expression, a greater cell proliferation rate was found in CD26-positive differentiated (follicular and papillary) carcinomas than in CD26-negative carcinomas. These results demonstrate that expression of this enzyme is related to the proliferative activity of follicular cells. PMID- 12114662 TI - Adrenocorticotropin-Producing Pituitary Carcinoma with Expression of c-erbB-2 and High PCNA Index: A Comparative Study with Pituitary Adenomas and Normal Pituitary Tissues. AB - Pituitary carcinomas are very rare neoplasms with a poor prognosis. We report a case of Cushing's disease resulting from a pituitary carcinoma in a 22-yr old female, who died of massive hepatic failure. At autopsy, there was invasion of the parasellar structures and vasculature by the tumor, which stained positively only for ACTH. There were two metastatic nodules in the liver, which also stained positively for ACTH. When compared to other cases of Cushing's disease (n = 52), other pituitary adenomas (n = 292). and normal pituitary tissues (n = 21), the pituitary carcinoma was the only one with c-erbB-2 membrane staining in both the sellar-located tissue and liver metastasis. C-erbB-2 staining was present in the cytoplasm of a variable number of cells in 40% of the invasive adenomas (n = 103), while only 1.2% of the noninvasive tumors (n = 241) expressed this protein (p < 0.001). No particular immunohistological type preferentially expressed this protein. In normal pituitary tissues, 10% of the cells expressed cytoplasmic c erbB-2. A higher index of proliferating cell nuclear antigen (PCNA) in the primary tumor and liver metastasis (10%) was also found compared to other ACTH secreting adenomas (invasive, 3.4 -t 1 S% vs 1 ii +/- 1.5% in noninvasive) and other pituitary tumors (invasive, 2.9 +/- 1.5% vs 1.5 +/- 1.3% in noninvasive). The PCNA index was significantly higher in invasive tumors than in noninvasive adenomas (p = 0.004). PCNA staining was negative in normal pituitary tissues. Staining for p53, pRB and p(2ras) was negative in the carcinoma and liver metastasis. We suggest that the c-erbB-2 membrane pattern and a higher PCNA index may indicate a worse prognosis in adenohypophyseal neoplasia. PMID- 12114663 TI - Immunodetection of Insulin-Like Growth Factor I (IGF-l) in Normal and Pathological Adrenocortical Tissue. AB - IGF-I is one of the peptides that participates in normal adrenocortical cell growth and, possibly, in the genesis and/or maintenance of tumors and hyperplasias of the adrenal cortex. An immunohistochemical technique was used for the analysis of IGF-l expression in eight control and four hyperplastic adrenals, 11 adrenal cortical adenomas, and 18 adrenal cortical carcinomas. A large number of IGF-I positive cells with granular cytoplasmic (GC) staining pattern was found in the reticularis layer of control adrenal tissues. Fifty percent of the hyperplasias had the GC pattern and the other 50% a mixed pattern; in 64% of the tumors, the adenomas showed a LM (linear membrane) pattern, while adenocarcinomas usually showed a GC pattern (94%). Approximately 75% of the hyperplasias had between 10 and 50% of IGF-l positive cells, while adenomas and carcinomas had over 50% of IGF-I positive cells in 64% and 83% of the samples, respectively. The size of the tumors with 50% positive cells, compared with those with less than 50%, was, on average, greater, but no statistical differences between cell positivity and corresponding clinical syndrome were observed. Thus, detection of IGF-l in control and pathological adrenal tissue suggests participation of this growth factor in cell function and/or growth and proliferation. PMID- 12114664 TI - Lymphocytic Hypophysitis: Light and Electron Microscopic Findings and Correlation to Clinical Appearance. AB - Light and electron microscopic findings of six cases of lymphocytic hypophysitis of a collective of 2500 surgical specimens are presented. Five cases were obtained by surgery, one case was obtained from autopsy. Light microscopy revealed an infiltration of mature lymphocytes and plasma cells in the interstitium, partly in the acini, as well as in the posterior lobe and the capsule. The structure of the remaining anterior lobe was normal. The final stage of lymphocytic hypophysitis is fibrosis, which was seen in all cases to varying degrees. The infiltrate consisted mainly of Tiymphocytes, being positive for CK 45 RO and CD 43. Immunocytochemical staining revealed different proportions of residual adenohypophyseal cells. Mainly prolactin reactive cells were observed as were growth hormone reactive cells. By electron microscopy some ruptured acini and damaged adenohypophyseal cells could be seen. Few pituitary cells contained enlarged lysosomal bodies or oncocytic changes. lnflammation causing enlargement of the pituitary leads to patients often being operated under the preoperative diagnosis of a tumor of the sellar region. It is important to identify this special type of hypophysitis, as a different course of treatment is required. PMID- 12114665 TI - Concurrent Pheochromocytoma, Paraganglioma, Papillary Thyroid Carcinoma, and Desmoid Tumor: A Case Report with Analyses at the Molecular Level. AB - Reports on the association of papillary thyroid carcinoma with paraganglionic or desmoid tumors have appeared infrequently. The former setting usually affects middle-aged females; the latter is typical of familial adenomatous polyposis. We report the case of a 69-yr-old man in whom two abdominal masses had been instrumentally detected following an access of abdominal pain. Save for a moderate hypertension, he was asymptomatic and an impalpable thyroid nodule was detected by ultrasonography. A high urinary noradrenaline output and cytology of the masses raised the suspicion of pheochromocytoma. At laparotomy, an adrenal pheochromocytoma and a paracaval paraganglioma were excised. Subsequently, hemithyroidectomy was performed, and histopathology revealed papillary microcarcinoma. A nodule of desmoid tumor was also removed from the abdominal wall. An analysis of RET, APC, and TP53 gene mutations, and of RET and NTRK1 gene rearrangements, yielded negative results. No in vitro transforming activity was detected in the tumor DNA when assayed in transfection experiments. The lack of a consistent family history also made unlikely the possibility of identifying the putative germline defect by linkage analyses. Should this unusual aggregation of tumors represent a new entity, a number of genetic alterations have now been excluded. PMID- 12114666 TI - Pituitary Adenoma with Cholesterol Clefts. AB - Histologically, cholesterol clefts are often observed in craniopharyngioma, Rathke's cleft cyst, and various granulomas. However, pituitary adenomas with cholesterol clefts are rare. A 46-year-old woman developed visual field disturbance. She had no history of severe headache that would suggest pituitary apoplexy. She presented with homonymous bitemporal hemianopsia and galactorrhea. Blood prolactin level was 63.1 ng/mL Other hypophysial hormone levels were within normal range. Magnetic resonance imaging revealed a pituitary tumor with intratumoral cyst. The cyst showed high intensity on T1- and T2-weighted images. The tumor was demonstrated with iso intensity on T1-weighted image and with high intensity on 12-weighted image. She underwent trans-sphenoidal surgery. The tumor was soft, with yellowish, oily fluid, probably the cyst content. By light microscopy with hematoxylin and eosin staining, a typical chromophobic adenoma of the pituitary was identified. Immunostaining revealed immunoreactivity for ACTH in several cells. Many cholesterol clefts and several hemosiderin pigment containing macrophages were observed. Electron microscopy demonstrated a pituitary adenoma with sparse and small secretory granules and numerous lysosomes. The cyst was most likely caused by focal hemorrhagic infarction, followed by the formation of cholesterol crystals, the appearance of hemosiderin containing macrophages, foreign body product cells, and accumulation of lysosomes. PMID- 12114667 TI - Relevance of Somatostatin Receptors and Other Peptide Receptors in Pathology. AB - Receptors for regulatory peptides can be overexpressed by several diseases, in particular by neoplasms. This review summarizes the current status of knowledge in this field, on the basis of in vitro receptor studies and with emphasis on receptors for somatostatin as well as for substance P (SP), vasoactive intestinal polypeptide (VIP), and cholecystokinin. It evaluates the existing and potential clinical implications of the findings for diagnosis and therapy and discusses the role of the pathologist in this context. PMID- 12114668 TI - Immunohistochemical Localization of p53 in Human Thyroid Neoplasms: Correlation with Biological Behavior. AB - Molecular analyses of thyroid tumors have documented mutations in the tumor suppressor p53 gene almost exclusively in anaplastic carcinomas. In contrast, immunohistochemistry has localized p53 in differentiated papillary and follicular thyroid cancers. To establish the significance of p53 immunolocalization in these lesions, 78 thyroid tumors of follicular derivation were examined. All tumors were classified by strict criteria and the extent of tumor was determined morphologically. Immunohistochemical staining for p53 was performed on paraffin sections of formalin-fixed tumor tissue. The results of staining were correlated with diagnosis, tumor extent and clinical outcome. Immunopositivity for p53 was diffuse and strong in all five anaplastic carcinomas examined. There was no staining in five of six follicular adenomas. Four of nine follicular carcinomas had some degree of nuclear staining, but this was focal; all nine tumors were confined to the thyroid at the time of examination. Of 49 papillary carcinomas, 26 were intrathyroidal, and 7 of these were occult; there was no p53 positivity in any occult lesion and only S of the 19 palpable lesions stained. In contrast, among 23 papillary carcinomas with extra thyroidal extension or metastases, only 9 were negative for p53 immunoreactivity. Five of seven tall cell papillary carcinomas and one of two insular carcinomas had p53 immunopositivity and this correlated with aggressive behavior. These results support the tumorigenic role of p53 mutations postulated for anaplastic thyroid carcinomas and indicate that localization of p53 by immunohistochemistry is a useful prognostic index of clinical behavior in differentiated thyroid carcinomas of follicular cell derivation. PMID- 12114669 TI - Proliferation in Adrenocortical Tumors: Correlation with Clinical Outcome and p53 Status. AB - There appears to be a relationship between mitotic activity and malignant behavior in adrenocortical tumors, and carcinomas with a high mitotic index may have a poorer prognosis. This has been investigated further by quantifying and comparing the Ki-67 index using antibody MIB-1 in a series of 14 adrenocortical adenomas and 40 carcinomas. The levels have been correlated with survival and disease-free survival in carcinomas and with evidence of abnormal p53 expression as detected by immunohistochemistry. Nevertheless, many carcinomas have a low level of proliferation that may reflect varying abnormalities within the regulation of both cell division and apoptosis. Expression of bcl-2 protein, an inhibitor of apoptosis has therefore also been examined. The Ki-67 index in carcinomas was significantly higher than in adenomas, but below 4% there was overlap. There was no significant difference in survival between carcinomas with MIB-1 index <3% and those greater, but the lower group had significantly longer disease-free survival (p = 0.02). There was no significant difference between p53 immunopositive and p53 immunonegative carcinomas. No tumor showed immunopositivity for bcl-2 protein. It is concluded that MIB-1 index may contribute additional prognostic information in adrenocortical tumors. Inhibition of apoptosis by bcl-2 does not appear to play a role in tumor growth. PMID- 12114670 TI - Clinical, Pathological, and Molecular Studies of Two Families with Iodide Organification Defect. AB - Two unrelated families (CA and NA) in which an iodide organification defect (lOD) was present in two siblings of each family were studied. These patients had congenital goiters with hypothyroidism and a positive perchlorate discharge test. Examination of the thyroid tissue revealed no thyroid peroxidase (TPO) activity. Histologic findings were consistent with a microfollicular pattern of hyperplasia. Moderate cellular atypia was present, characterized by nuclear pleomorphism and hyperchromatism. Full length thyroglobulin was purified by gel filtration, but was not iodinated. Immunohistochemical studies using a polyclonal anti-human thyroid peroxidase (hTPO) antibody confirmed the presence of immunoreactive TPO protein in the thyroid tissues. Samples of normal and affected individuals were studied with respect to the presence of various fragments using TPO probes of varying sizes. The two affected siblings from family CA were homozygous for fragments 3.9, 4.6, and 7.0 kb (8g111) and 2.3 and 2.9 kb (Ta ql), whereas the parents were heterozygous. In the other family (NA), the Bg/ll digestion and TPO-31 hybridization revealed an interesting and informative polymorphism. The parents showed two different polymorphic patterns: the father had a 5.0/4.6 kb pattern and the mother a 4.7/4.5 kb pattern. However, the two affected siblings showed the same heterozygotic allelic pattern at 4.5/4.6 kb. The restriction fragment length polymorphism detected in these two families suggests an association between the TPO gene and an lOD. Results suggest that in these dyshormonogenetic tissues an altered TPO protein molecule is being synthesized, without detectable in vitro activity, but visible by immunostaining techniques in the goitrous tissue. Mutations in the TPC gene sequence are most likely associated with these changes. PMID- 12114671 TI - Cathepsin B and Cathepsin D Expression in Follicular Adenomas and Carcinomas of the Thyroid Gland. AB - Biologic features that distinguish follicular adenomas (FAs), minimally invasive follicular carcinomas (MICs), and extensively invasive follicular carcinomas (EICs) of the thyroid gland are not well understood. Endogenous proteases, including cathepsin B (CB) and cathepsin D (CD), have been linked to tumor progression in other malignancies and may be important in the different biologic behavior of these follicular thyroidal lesions. Archival paraffin-embedded tissue sections from 16 FAs, 12 MICs, and 4 EICs were studied for immunohistochemical expression of CR and CD. Percent of tumor staining, intensity of staining, and intracytoplasmic staining pattern were assessed. Increased intensity of staining with CD was observed in follicular carcinomas compared to adenomas and was greatest in the EIC (p < 0.04). An increase in diffuse cytoplasmic staining pattern with CD (p < 0.008) and CB (p < 0.02) was observed in follicular carcinomas compared to FAs. The increased intensity of staining with CD in the follicular thyroid carcinomas and the increased diffuse cytoplasmic staining pattern with CD and CR in these carcinomas suggest that these proteinases may play a role in their propensity to invade and metastasize and, therefore, their more aggressive behavior. Furthermore, this diffuse cytoplasmic staining suggests an altered intracellular processing of these proteinases in the carcinomas. PMID- 12114672 TI - Adenomatoid Tumor of the Right Adrenal Gland in a Patient with AIDS. AB - An autopsy case with an incidentally discovered adenomatoid tumor (AT) arising in the right adrenal gland of a 34-yr-old man with AIDS is presented. The immediate cause of death was disseminated coccidioidomycosis. The affected right adrenal gland was partially substituted by a firm 3.O cm nodule enclosed by cortical adrenal tissue. Histologically, the tumor had the typical appearance of those adenomatoid neoplasms described in the genital tract. The mesothelial origin of the neoplasm was confirmed by immunopositive cells for low weight cytokeratin and vimentin. Uncommon neoplasms in the adrenal glands of AIDS patients include leiomyosarcomas, leiomyomas, and malignant nerve sheath tumors. This report may represent the first case of an adrenal gland AT in a patient with AIDS, and probably the third well-documented histologically and immunohistochemically adrenal gland AT. PMID- 12114673 TI - Cushing's Syndrome from an Ectopic Pituitary Adenoma with Peliosis: A Histological, Immunohistochemical, and Ultrastructural Study and Review of the Literature. AB - Ectopic pituitary adenoma (EPA) is rare and, to the authors' knowledge, its association with peliosis has not yet been described. The case of a 38-yr-old woman with clinical and biochemical evidence of Cushing's syndrome is reported. Magnetic resonance imaging (MRI) disclosed a normal pituitary and a separate mass in the sphenoid sinus. The surgically removed portion of the sellar pituitary contained no adenoma. There was only Crooke's hyaline change in the corticotrophs, indicating exposure to glucocorticoid excess. By histology, the mass in the sphenoid sinus was a congested, chromophobic, partly basophilic, periodic acid-Schiff (PAS)-positive pituitary adenoma composed of pleomorphic, adrenocorticotropic hormone (ACTH)-positive, corticotrophs. There was focal immunopositivity for MIB-1 and proliferating cell nuclear antigen (PCNA). Electron microscopy confirmed the diagnosis of corticotroph adenoma. A striking finding, consistent with the diagnosis of peliosis, was the presence of multiple large blood-filled spaces lacking an endothelial lining. The capillaries were dilated, but often appeared empty and the fenestrated endothelium exhibited discontinuities. The cause of peliosis is obscure. It may be that the venous outflow was impaired in this case leading to capillary dilation, congestion, hyperpermeability, rupture, and accumulation of blood in extravascular spaces. PMID- 12114674 TI - Epithelial Cyst of Thyroid. AB - A benign thyroid cyst lined by thyroid follicular epithelial cells lacking immunoreactivity to thyroglobulin is reported. The patient had a 3 yr history of a large cyst in the lateral anterior neck, with no symptoms of hyperthyroidism or hyperparathyroidism. Water-clear fluid was repeatedly aspirated; analysis indicated the absence of parathyroid hormone (PTH0), T3, and T4. Hemithyroidectomy revealed a simple epithelial cyst. Such cysts are rare, comprising approx 1% of thyroid nodules. A differential diagnosis of cystic neck lesions is presented and the possible pathogenesis of this cyst is discussed. PMID- 12114675 TI - A Plurihormonal TSH-Secreting Pituitary Microadenoma: Report of a Case with an Atypical Clinical Presentation and Transient Response to Bromocriptine Therapy. AB - Inappropriate secretion of thyrotropin (TSH) is a rare cause of hyperthyroidism, and it is caused by either a TSH-producing pituitary adenoma (usually a macroadenoma) or to selective pituitary resistance to thyroid hormone. The case of a 31-yr-old male who presented with clinical features of thyrotoxicosis, including episodes of thyrotoxic paralysis, and a thyroid profile characterized by free hyperthyroxinemia and hypertriiodothyronemia with a nonsuppressed, inadequately normal TSH is reported. Dynamic testing showed both, lack of TSH stimulation by thyroid-releasing hormone (TRH), and lack of suppression by T3, consistent with autonomous TSH secretion. Pituitary MRI revealed a microadenoma. Seventy five percent of the patients serum TSH immunoreactivity eluted as u subunit in Sephadex G-100 chromatography. A diagnosis of TSH-secreting microadenoma was established, and the patient was treated successfully with bromocriptine, which resulted in both clinical and biochemical resolution of his hyperthyroidism. Two months later, he became hyperthyroid again during bromocriptine therapy. Octreotide was started with adequate control of his symptoms and normalization of his free T4 level. He eventually underwent transsphenoidal surgery with successful resection of a chromophobic microadenoma which immunostained for TSH, growth hormone (GH), luteinizing hormone (LN), and follicle-stimulating hormone (FSH). One month postoperatively he is clinically and biochemically euthyroid on no medications. PMID- 12114678 TI - Apoptosis and Expression of the Proto-Oncogenes bcl-2 and p53 and the Proliferation Factor Ki-67 in Human Medullary Thyroid Carcinoma. AB - Apoptosis and Immunoreactivity for bcl-2, p53, and Ki-67 were studied in 21 patients with meduilary thyroid carcinoma (MTC). The DNA nick end labeling method was used to assess apoptosis. The relationships between the different factors were analyzed, as were their relations to clinicopathologic data, including survival. More than 80% of the tumors harbored apoptotic cells. Tumors in individuals who had died of the MTC disease had a higher percentage of apoptosis. All cases demonstrated immunoreactivity to bcl-2 disease-free individuals had a higher rate than those with recurrent disease. No obvious pattern could be discerned in the relation of p53 or Ki-67 to the outcome of the disease. An inverse correlation between bcl-2 and apoptosis (r=-0.81; p < 0.01) was demonstrated. bcl-2 was significantly (p = 0.014) associated with apoptosis even after taking both p53 and Ki-67 into consideration, but these two factors were unrelated to apoptosis. None of the factors studied were correlated to crude survival, either in univarlate or in muiltivariate analyses. This study established that bcl-2 immunoreactivity is closely associated with apoptosis in MTC, suggesting that a down-regulation of the bcl-2 protein is related to a more aggressive growth rate and might be a useful marker for the evaluation of MTC. PMID- 12114679 TI - Intraepithelial Neutrophils in Thyroid Fine-Needle Aspiration: A Portent of Aggressive Thyroid Cancer? AB - We report three patients with papillary thyroid carcinoma in whom fine-needle aspiration (FNA) showed neutrophils within tumor cells. All three patients presented with large neck masses; at excision, two proved to be tall cell variants of papillary cancer. Nodal metastasis, extrathyroidal extension, and vascular invasion were found in both cases. One patient has experienced recurrent disease; the other has an increasing thyroglobulin titer but no clinically appreciable recurrence. The third patient refused further therapy, but brain metastases were noted clinically; this patient died of disease. In each case, FNA showed tumor clusters with characteristic nuclear features, papillary groups, and psammoma bodies. Neutrophils were present in the cytoplasm of tumor cells in the absence of necrosis. Immunostaining for proliferating cell nuclear antigen (PCNA), MIB-1 (Ki-67), and p53 tumor suppressor gene product was markedly positive. lntraepithelial neutrophils have not been previously reported in differentiated thyroid tumors. We postulate that these neoplasms produce specific leukocyte-attracting cytokines analogous to those produced by anaplastic and poorly differentiated thyroid carcinomas. We believe the finding of intraepithelial leukocytes in the absence of necrosis in thyroid FNA specimens represents a characteristic of clinically aggressive differentiated papillary neoplasms; in our small series, each represented a tall cell variant of papillary carcinoma. PMID- 12114676 TI - The Pathology of Medullary Carcinoma of the Thyroid: Review of the Literature and Personal Experience on 62 Cases. AB - The aim of the present review is to analyze the numerous pathological patterns of medullary carcinoma of the thyroid (MCT) and discuss the problems of differential diagnosis with other thyroid and nonthyroid tumors. In addition, morphological parameters and phenotypic features were related to the clinical outcome. The recent literature was reviewed and compared with the features of 62 MCTs observed at our institution. The most common patterns of growth are trabecular, alveolar and spindle cells, but MCT can mimic virtually all other primary thyroid tumors and some nonthyroid neoplasms. This heterogeneity has no proven implications for prognosis, but is of relevance for diagnostic purposes, as the differential diagnosis of MCT can be difficult in nonclassical cases. In agreement also with the literature data, no relationship between histopathological parameters and outcome was found in our series, although clinically aggressive tumors had a more advanced stage at presentation. Immunocytochemica demonstration of calcitonin is apparently the only valid criterion for a correct typing of MCT. Chromogranin A is an additional sensitive marker of MCT and parallels calcitonin expression in the majority of cases. PMID- 12114677 TI - Pituitary Carcinomas. AB - Pituitary carcinomas are defined by their metastatic growth. Most of them also invade into surrounding tissues. They should be classified by the site of their metastases (cerebrospinal, systemic, or combined) and by the presumable cell type of origin, respectively with the hormone being demonstrable by immunohistochemistry (adrenocorticotrophic hormone [ACTH], prolactin [PRL], growth hormone [GH], hormone-negative). Pituitary carcinomas develop from invasive adenomas. Nearly all tumors had been treated by surgery or X-ray before they metastasized. Since 1976, 37 cases demonstrated with modern methods were reported: 23 had metastasized into the brain or meninges, 10 showed extracerebral metastases, and 4 showed both types of metastases. In our collection of pituitary tumors, three carcinomas (0.13%) were identified: two with systemic metastases (one ACTH secreting and one PRL secreting) and one with meningeal dissemination and ACTH production. The diagnosis of pituitary carcinomas should be based on four criteria: a demonstrable metastasis, identification of the primary tumor as a pituitary tumor, similarity between the structure and immunohistological marker expression of metastasis and primary tumor, and exclusion of an alternative primary tumor. PMID- 12114680 TI - Recurrence in Parathyroid Hyperplasias Owing to Secondary Hyperparathyroidism is Predicted by Morphological Patterns and Proliferative Activity Values. AB - The histological pattern and the cell proliferative activity (as detected by Ki 67 immunostaining) of a series of 50 parathyroid hyperplasias (PTHs) secondary to renal failure were studied to assess their value in predicting recurrence of hyperparathyrodism (HPT). On account of their clinical evolution, these cases were divided into two groups, recurrent HPT (23 cases) and nonrecurrent HPT (27 cases). A nodular growth pattern (as opposed to diffuse) was the prevalent one and was observed in 20 (74%) cases of now recurrent HPT and in 22 (95.6%) cases of recurrent HPT, a statistically significant difference (p < 0.05). The Ki-67 proliferative fraction was 1 9% in recurrent HPT cases, as compared with 0.81% in nonrecurrent HPT, a difference which was statistically significant (p = 0.001). We conclude that a nodular pattern of growth and an elevated Ki-67 proliferative fraction (>1.5%) in PTH are both associated with a higher risk of recurrence (4.30) of HPT. PMID- 12114681 TI - Localization of Epidermal Growth Factor (EGF) and Epidermal Growth Factor Receptor (EGFr) in Human Pituitary Adenomas and Nontumorous Pituitaries: An Immunocytochemical Study. AB - Epidermal growth factor (EGF) and epidermal growth factor receptor (EGFr) were investigated by immunocytochemistry (ICH) in 57 human pituitary adenomas and 10 nontumorous autopsy pituitaries. EGF immunoreactivity was demonstrated in 24 adenomas (42%), representing 23 functioning tumors and 1 nonfunctioning tumor of oncocytic type, and in all nontumorous pituitaries. Among 40 tumors, EGFr was found positive in 15 functioning adenomas (37.5%), representing 50% of them. The presence of both EGF and EGFr was found mainly in corticotroph adenomas (60%) and less frequently in somatotroph and lactotroph adenomas (20%). ICH on serial sections with EGF or EGFr and adrenocorticotrophic hormone (ACTH) or S-100 protein revealed that EGF and EGFr are localized specifically in corticotrophs and EGFr in stellate cells of nontumorous adenohypophysis. These results confirm the presence of EGF and EGFr in human pituitary adenomas and nontumorous pituitaries and highlight their frequent occurrence in hormone-producing adenomas. Further work is required to explore the possiblity that EGF and EGFr play a role in hormone production, release, and tumor progression. PMID- 12114682 TI - Molecular Screening for RET Proto-Oncogene Mutations in a German MEN2A Pedigree. AB - Multiple endocrine neoplasia type 2A (MEN2A), a dominantly inherited cancer syndrome, is defined by the presence of medullary thyroid carcinoma (MTC), pheochromocytoma (pheo), and primary hyperparathyroidism (p-HPT). Along with multiple endocrine neoplasia type 2B (MEN2B) and familial medullary thyroid carcinoma (FMTC), it is associated with germline mutations of the RETproto oncogene localized in 10q11.2. In FMTC and MEN2A, point mutations result in the substitution of one of five Cys residues in the extracellular domain of RET. In a larger pedigree from Saarland, several individuals were observed with C-cell thyroid carcinoma. We screened 16 members of this extended family by single strand conformation polymorphism analysis (SSCP), polymerase chain reaction (PCR), followed by restriction enzyme analysis, and by sequencing the mutated regions. In 7 family members, all of whom had been earlier operated on because of MTC, a DNA transition from T to C was observed, causing an amino acid substitution Cys(634)Arg. Nine members of the kindred did not carry the mutation and may be excluded from yearly biochemical testing. One of these persons seems to have been unnecessarily operated on owing to a borderline pentagastrin test. PMID- 12114683 TI - Expression and Regulation of Transforming Growth Factor B1 in Cultured Normal and Neoplastic Rat Pituitary Cells. AB - Pituitary prolactin (PRL) cell gene expression and proliferation are regulated by hormones and growth factors. Transforming growth factor beta (TGFB) and blast growth factor (bFGF) have been implicated in the regulation of antenor pituitary function. To study the roles of TGFB and bFGF in anterior pituitary cell function, we analyzed normal and neoplastic pituitary cells in serum-free media. The various isoforms of TGFB and TGFB receptor types I, II, and III were also analyzed by reverse transcription-polymerase chain reaction (RT-PCR) in pituitary cells. Transforming growth factor beta 1 (TGFB1) stimulated PRL expression and PRL cell proliferation in normal pituitary. TGFB1 stimulated PRL expression, but inhibited proliferation in the growth hormone (GH) and PRL-producing GH(3) cells. Estradiol 17 B (E(2)) and bFGE stimulated PRL gene expression in normal pituitary and GH(3) cells, whereas E(2) inhibited and bFGF stimulated TGFB1 mRNA levels in normal pituitary PRL cells, but not in GH(3) cells. Both normal pituitary and GH(3) cells expressed the mRNAs for TGFB1, TGFB2, and TGFB3 isoforms and for TGFB receptors I, II, and III. These results indicate that there is a relative loss of regulatory control by growth factors in neoplastic GH(3) cells compared to normal pituitary PRL cells. PMID- 12114684 TI - Pituitary Corticotroph Adenoma in a Woman with Long-Standing Addison's Disease: A Histologic, immunocytochemical, Electron Microscopic, and In Situ Hybridization Study. AB - A 64-year-old woman with long-standing Addison's disease owing to destructive immune adrenalitis presented with hyperpigmentation and progressively increasing blood adrenocorticotrophic hormone (ACTH) levels. Magnetic resonance imaging demonstrated a pituitary microadenoma, which was removed by transsphenoidal surgery and investigated by histology, immunocytochemistry, transmission electron microscopy, and in situ hybridization (ISH). The morphologic studies revealed a basophilic, periodic acid-Schift (PAS)-positive pituitary adenoma immunoreactive for ACTH and B-endorphin and in several cells for a-subunit. By transmission electron microscopy, the tumor was a densely granulated corticotroph adenoma, which, by ISH, expressed pro-opiomelanocortin (POMC) mRNA. The lack of corticotroph hyperplasia in the nontumorous adenohypophysis was an intriguing finding. Corticotroph adenomas in patients with long-standing Addison's disease were very rarely examined by morphology. Our report includes a detailed morphologic analysis and is the first demonstration of POMC mRNA in the tumor cells using ISH. The question of whether the adenoma was related to increased secretory activity secondary to protracted hypocorticism or developed independently unrelated to deranged endocrine homeostasis remains unresolved. The lack of corticotroph hyperplasia in the nontumorous adenohypophysis favors the interpretation that hypothalamic stimulation played no major role in adenoma formation in our case. PMID- 12114685 TI - Heat-Shock Stress-Response Proteins in Endocrine Pathology. AB - Heat-shock proteins (HSPs), also known as stress-response proteins, represent an evolutionarily conserved class of glycoproteins; members of this protein family are also known as "molecular chaperones." HSPs are constitutively expressed, and most are overproduced in response to a nonlethal thermal shock or other stressful conditions. They are implicated in several cell functions; they likely act in association with steroid receptors at the level of receptor-DNA interactions. Various types of HSPs have been found in endocrine glands, hormone-dependent tissues, and neoplasms. At present, their exact role remains obscure. HSPs may serve as tumor markers of prognostic significance; they may also have diagnostic and therapeutic uses. PMID- 12114687 TI - Monoclonal Antibodies Recognizing Normal and Neoplastic Human Adrenal Cortex. AB - Four monoclonal antibodies (MAb) were generated by immunization of mice with dispersed cells from normal human adrenal gland (Na) and adrenocortical adenoma causing cortisol excess (Ac). Immunohistochemically reacted cryosections revealed differential labeling of the normal cortical parenchyma, and immunofluorescence on dispersed cells displayed that Ac5 alone labeled the cell surface. Immunoprecipitation demonstrated that the antibodies recognized apparently different structures of 51-88 kDa. Immunohistochemical examination of several normal human tissues substantiated restricted reactivity, especially for the Na2 and Na7 antibodies, and that the adrenal medulla was not stained by any of the antibodies. The antibodies recognized the vast majority of the parenchymal cells of cortical adenomas (n = 21). Each antibody also reacted with all adrenocortical carcinomas (n = 17), and the staining generally was most intense and extensive with Na7. Analysis of other pathological human tissues revealed highly restricted reactivity for the Na2 antibody. Na2 and Na5 failed to stain 17 renal cell carcinomas. None of the antibodies recognized pheochromocytomas. These antibodies may lead to improved histological recognition and characterization of human adrenal lesions. PMID- 12114686 TI - HGH, PRL, and ACTH Gene Expression in Clinically Nonfunctioning Adenomas Detected with Nonisotopic In Situ Hybridization Method. AB - In situ hybridization (ISH), which can manifest the specific gene expression of anterior pituitary hormones (mRNA), as well as immunohistochemistry (IHC), is needed to clarify the endocrine function of pituitary adenomas. With the aid of nonisotopic ISH, which has several advantages over isotopic ISH, we examined the expression of pituitary hormone mRNAs in 14 clinically nonfunctioning adenomas, which were considered to be a subtype of gonadotroph adenomas. Gene expression of growth hormone (GH; 4/14), prolactin (PRL; 5/14), adrenocorticotroph hormone (ACTH; 4/14), and gonadotropin were detected with our nonisotopic ISH studies. It is suggested from our ISH studies that some clinically nonfunctioning adenomas are composed of hormone (or subunit) producing cells and may be derived from plurihormonal primordial stem cells. PMID- 12114688 TI - Immunohistochemical Localization of Chromostatin and Pancreastatin, Chromogranin A-Derived Bioactive Peptides, in Normal and Neoplastic Neuroendocrine Tissues. AB - Despite the widespread distribution of chromogranin A (CgA) in neuroendocrine tissues, the biological function of CgA has not yet been elucidated. The primary amino acid sequence of CgA, elucidated by cDNA analysis, has been revealed to include several pairs of basic amino acid residues that are homologous to the bioactive peptides, such as pancreastatin (PST) and chromostatin (CST). Using antibodies for human PST and CST, the immunohistochemical localization of these peptides was investigated in neuroendocrine tissues, including human pituitary glands, pancreas, adrenal medulla, various types of neuroendocrine neoplasms (13 pheochromocytomas, 10 medullary thyroid carcinomas, 11 pancreatic endocrine tumors, and 19 carcinoid tumors), and the cell line QGP-1N derived from human somatostatin-producing pancreatic endocrine tumor. Variable immunoreactive intensities of PST and CST were seen, but both peptides were detectable in all neuroendocrine tissues and in most of the neoplasms. Immunoreactivity for both PST and CST was observed in 100 and 73%, respectively, of pancreatic endocrine tumors, all pheochromocytomas, and 80 and 40%, respectively, of medullary thyroid carcinomas, as well as all nonrectal carcinoid tumors. In rectal carcinoids, cells immunoreactive for PST and CST were sparse. The distribution of PST and CST was similar to that of CgA, and it is considered that these peptides are simultaneously processed from CgA, and may play roles in autocrine and paracrine regulation on various hormones in addition to their previously known functions. PMID- 12114689 TI - Proliferation Markers and EGF in ACTH-Secreting Adenomas and Carcinomas of the Pituitary. AB - Pituitary carcinomas are only defined by their metastatic growth, which may be intracranial or systemic. To establish further morphological and immunohistochemical differences between pituitary carcinomas and adenomas, 19 ACTH-secreting adenomas (10 non invasive and 9 invasive) and 2 ACTH-secreting carcinomas with their metastases were studied for expression of the intermediate filaments keratin and vimentin and the tumor-associated antigens Ki67, proliferating cell nuclear antigen (PCNA), epidermal growth factor (EGF), cathepsin D, p53, and carcinoembryonic antigen (CEA). Immunohistochemistry was performed using avidin-biotin techniques on formalin-fixed, paraffin-embedded tissue. With the exception of one noninvasive pituitary adenoma, one carcinoma, and the metastases, all tumors contained keratin; none contained vimentin. All tumors stained negative for CEA and p53. Eleven (58.5%) adenomas and both pituitary carcinomas contained Ki67-positive nuclei; 14 (74%) adenomas and one carcinoma revealed PCNA. No correlation was found between the two markers. Seven (38%) adenomas showed a labeling index <1 % for cathepsin D, whereas none of the carcinomas or metastases did so. EGF was found in 7 (38%) adenomas and in both carcinomas. A tendency to a higher rate of EGF positivity in the invasive adenomas was observed. The metastases showed a higher labeling index, and far more intense staining results for Ki67, PCNA, and EGF than the primary tumor. The metastases also had a higher proliferation rate and growth factor content than the carcinoma itself. PMID- 12114690 TI - Failure of Partial Hypophysectomy as Definitive Treatment in Cushing's Disease Owing to Nodular Corticotrope Hyperplasia: Report of Four Cases. AB - Nodular corticotrope hyperplasia is a rare pathology causing Cushing's syndrome owing to a primary pituitary disease or ectopic CRH production. In this study, we evaluated the laboratory and pathological findings and results of transsphenoidal pituitary surgery in four patients with Cushing's disease. Dynamic tests of pituitary-adrenal function (dexamethasone suppression, metyrapone, CRH, and DDAVP tests) were done before and after transsphenoidal pituitary surgery. Plasma and total urinary cortisol, serum 11-deoxycortisol, and plasma ACTH were determined by RIA. Hormonal dynamic tests and radiologic studies were compatible with a pituitary ACTH source. The transsphenoidal surgery revealed the presence of corticotrope hyperplasia confirmed by immunoperoxidase stain and a preserved reticulum framework in the removed pituitary tissue of these four patients. The pituitary surgery led to a short period of improvement in two of the patients (1 and 4), a 3-yr remission in one patient (patient 2), and no improvement in one (patient 3). We conclude that although our patients appear to have inadequate suppression with high-dose dexamethasone, there is no way to diagnose this pathology presurgically, and that total hypophysectomy, bilateral adrenalectomy, and irradiation are the only alternatives for definitive treatment. A CRH secreting ectopic tumor could not be found in our patients either before or after surgery in the follow-up period. PMID- 12114691 TI - Pituitary Folliculo-Stellate-Like Cells Stimulate Somatotroic Pituitary Tumor Growth in Nude Mice. AB - A new cell line (TtT/GF) established from a murine pituitary thyrotropic tumor having characteristics similar to those of pituitary folliculo-stellate cell (FS cell) was implanted into nude mice together with cells from a rat pituitary somatotrophic tumor cell line (MtT/S) to determine whether the former enhances pituitary tumor growth. For as long as 2-3 mo after implantation, MtT/S cells implanted either alone or together with fibroblasts formed either no tumors or only very small tumors in the nude mice. In contrast, all of the nude mice that had received MtT/S cells implanted together with TtT/GF cells developed large tumors. Furthermore, the mice bearing the MtT/S and TtT/GF implants showed a significantly higher body weight and serum growth hormone level than those bearing only MtT/S cells or a combination of MtT/S cells and fibroblasts. The TtT/GF cell line itself had no tumorigenicity during the experimental period. Therefore, the TtT/GF cell line as a model of FS cells enhanced pituitary endocrine cell tumor formation. Additionally, immunocytochemistry showed that TtT/GF cells positive for glial fibrillary acidic protein (GFAP) or S-100 protein were present in the parenchymatous tissue elements or connective tissue surrounding the tumor nests. In the parenchymatous tissue, the TtT/GF cells exhibited a stellate appearance and surrounded neighboring tumor cells with their long cell processes. These results suggest that TtT/GF cells can serve as a model for pituitary FS cells, and are capable of stimulating pituitary tumor growth either by modifying the microenvironment or producing growth factors. PMID- 12114692 TI - Cystic Squamous Cell Carcinoma of the Thyroid A Possible New Subgroup of Intrathyroidal Epithelial Thymoma. AB - In the thyroid gland, both squamous cell carcinoma and intrathyroidal epithelial thymoma are rare tumors with squamous features, but their prognoses differ; the former is usually fatal, and the latter is favorable. We describe a 38-yr-old female patient with a cystic tumor diagnosed cytologically as squamous cell carcinoma, who was treated with subtotal thyroidectomy. The tumor was located intrathyroidally in the middle third of the left lobe, and no invasion to the surrounding tissues was found. Histologic examination disclosed a cyst lined by epithelial cells with squamous cell differentiation. The tumor cells had ill defined cell borders and large nuclei with prominent nucleoli. Invasion of the tumor capsule, a few mitoses, and infiltration of lymphocytes in the tumor periphery were noted, but neither necrosis nor metastasis to lymph nodes was observed. The patient has been free from recurrence for 3 yr after thyroid lobectomy. We propose that this type of cystic intrathyroidal squamous cell carcinoma may represent a cystic variant of intrathyroidal epithelial thymoma. PMID- 12114694 TI - Vascularity in Nontumorous Human Pituitaries and Incidental Microadenomas: A Morphometric Study. AB - The aim of the present study was to determine the microvascular angioarchitecture in the lateral and central portions of the anterior lobe as well as of the posterior lobe. The possible association between vascularity and age, sex, and pregnancy was also examined. In addition the vascular density of incidental microadenomas was investigated and compared to that of nontumorous gland. Blood vessels of 120 nontumorous pituitaries and 11 incidental microadenomas obtained at autopsy were examined by immunohistochemistry using the endothelial marker CD 34. Microvascular density (MVD) and microvessel surface density (MSD) were determined by morphometry using an automatic computer image analysis system. MVD and MSD were higher in the anterior than in the posterior lobe. Age, gender, or pregnancy did not affect the angioarchitecture of these sites. No statistical differences in MVD and MSD were observed between central and lateral areas of the anterior lobe, although MVD appeared to be lower in the lateral zones. A marked difference was noted in the vascularity of microadenomas compared to nontumorus tissue; both MVD and MSD were significantly lower in adenomatous tissue. Our studies suggest that the capillary network of the anterior and posterior lobes differ. They also indicate that the microvascular architecture is not significantly affected by age, gender, or pregnancy. Lack of significant angiogenesis in pituitary microadenomas may underlie the low growth rate and infrequency of metastasis of pituitary adenomas. PMID- 12114693 TI - Bcl-2 Gene Family in Endocrine Pathology: A Review. AB - BcI-2 is a member of a large multigene family, which includes genes that can inhibit or promote apoptosis. The regulation of apoptosis is achieved by homo- or heterodimerization of their proteins through four highly conserved domains. Bcl-2 protein is a strong cell death suppressor in a wide range of cell types and under a variety of stimuli. Bcl-2 and the other members of this family are differentially expressed in the endocrine glands and disregulation of their expression seems to contribute to the neoplastic transformation in these organs. The significance of bcl-2 and the related proteins for endocrine pathology at the experimental and clinical level is reviewed in this article. PMID- 12114695 TI - Prognostic Significance of p27, Ki-67, and Topoisomerase lla Expression in Clinically Nonfunctioning Pancreatic Endocrine Tumors. AB - Nonfunctioning islet cell tumors or pancreatic endocrine tumors are the most common type of malignant islet cell tumor. Although previously detected usually at an advanced stage because of mass effect, the early detection rate of small localized disease has been increasing. To date it has been difficult to predict the clinical behavior in localized regional nonfunctioning tumors. To investigate potential markers predicting malignancy and poor prognosis in nonfunctioning pancreatic endocrine tumors, we analyzed the expression of Ki-67, topoisomerase Ila (Topolla), and p27, as well as a variety of clinicopathologic parameters in 76 cases of nonfunctioning islet cell tumors (23 benign cases and 53 malignant cases). Ki-67, Topolla. and p27 labeling indices were significantly different between benign and malignant tumors. Expression of Ki-67, Topolla, and p27 were associated with survival in patients with a malignant tumor in a univariate setting. However, only p27 and Topolla were jointly associated with survival in multivariate analysis. Immunohistochemical staining for p27, Topolla, and Ki-67 can be helpful in the diagnosis of nonfunctioning pancreatic endocrine tumor. Analysis of p27 and Topolla may also have potential utility as prognostic factors for malignant tumors. PMID- 12114696 TI - Nuclear Accumulation of B-Catenin in Human Endocrine Tumors: Association with Ki 67 (MIB-1) Proliferative Activity. AB - B -Catenin is closely associated with carcinoma invasion/metastasis and poor survival. Recent studies have demonstrated that abnormal expression of B catenin, especially its nuclear accumulation, also plays an important role in wingless/Wnt signaling pathway. In this study, we evaluated immunohistochemically the nuclear localization of B -catenin in a total of 93 human-endocrine-related tumors including 1 medullary carcinoma (thyroid gland), 12 parathyroid tumors, 22 carcinoid tumors (digestive tract and liver), 7 islet cell tumors, 26 adrenocortical tumors, 13 neuroblastoma (adrenal gland), and 12 pheochromocytoma (adrenal gland), and also studied genetic alterations of the B -catenin gene. Nuclear accumulation of B -catenin was frequently detected in 8 of 22 (36%) carcinoid tumors and 2 of 7 (29%) islet cell tumors. No genetic alteration in exon 3 of the B -catenin gene encoding serine/threonine rich domain, which was phosphorylated by GSK-3 B, was detected in any groups of the endocrine tumors. However, nuclear accumulation of B -catenin in carcinoid tumors was significantly correlated with the proliferative marker Ki-67 (MIB-1) labeling index (p <0.001). Our findings suggest that nuclear transfer and accumulation of the B -catenin may contribute in the tumorigenesis of carcinoid tumor as an oncoprotein. PMID- 12114698 TI - Analysis of the Cycilin D1/p16/pRb Pathway in Parathyroid Adenomas. AB - Cyclin D1/p16/pRb pathway controlling G1-S cell cycle checkpoint is frequently altered in human tumors. Currently, scarce data are available for parathyroid tumors. We have studied 46 parathyroid adenomas (PTAs) and 12 normal parathyroid glands (PTGs) by immunohistochemistry with cyclin D1 (CD1), p16, pRb, and Ki-67 antibodies. We observed CD1 expression in 89%, p16 in 70%, and pRb in 100% of PTAs. Statistically significant differences with normal PTGs were found only concerning p16 expression (p <0.05). Proliferating rate (Ki-67) was always low, although significantly higher than in normal PTGs. Our findings demonstrate the presence of alterations in the CD1/p16/pRb pathway in PTAs, consisting in p16 overexpression apparently unrelated to pRb downregulation. On the other hand, we did not find significant differences in CD1 expression between PTAs and normal PTGs, suggesting CD1 overexpression could be a physiological event in parathyroid tissue. PMID- 12114697 TI - p53/MDM2 Pathway Aberrations in Parathyroid Tumors: p21(WAF-1) and MDM2 Are Frequently Overexpressed in Parathyroid Adenomas. AB - Parathyroid adenomas (PTAs) are the main cause of primary hyperparathyroidism. Cell cycle regulation in normal parathyroid tissue (NPT) and PTA remains largely unknown. We have systematically explored several components involved in the p53/MDM2/p19(ARF) pathway in PTA and compared the results were with NPT. Forty six PTA and 12 NPT were immunostained with anti-p21(WAF-1), MDM2, p53, and p27(KP1) antibodies. The slides were processed by cytometry and the results were statistically analyzed using nonparametric methods (Mann-Whitney test). p2l(WAF 1) and MDM2 expression were significantly higher in PTA compared with NPT (p < 0.05). The opposite results were found for p27(KIP1) (p< 0.05). Occasional p53 staining was found in some PTA, albeit no significant difference was found in comparison with NPT. In conclusion, MDM2 and p2l(WAF-1) are the proteins more overexpressed in PTA. These findings are surprising taking into account the benign nature of PTA, making them suitable candidates for further molecular analysis. PMID- 12114700 TI - Electron Microscopic and Confocal Laser Scanning Microscopic Observation of Subcellular Organelles and Pituitary Hormone mRNA: Application of Ultrastructural In Situ Hybridization and Immunohistochemistry to the Pathophysiological Studies of Pituitary Cells. AB - Nonradioisotopic electron microscopic (EM) in situ hybridization (ISH) (EM-SH) with biotinylated oligonucleotide probes is utilized for the ultrastructural visualization of pituitary hormone mRNA in rat pituitary cells. EMISH is an important tool for clarifying the intracellular localization of mRNA and the exact site of specific hormone synthesis on the rough endoplasmic reticulum. The simultaneous visualization of mRNA and encoded protein in the same cell using preembedding EM-ISH and subsequent postembedding immunoreaction with protein A colloidal gold complex can provide an important clue for elucidating the intracellular correlation of mRNA translation and secretion of translated protein. Another focus of this review is the utilization of a recently developed imaging system of confocal laser scanning microscopy (CLSM). The combination of CLSM and image analysis system (lAS) enables us to visualize an individual dimensional image of the intracellular distribution of mRNA and subcellular organelles successfully at any optional cross sections of light microscopic ISH studies, and can be another useful tool for the ultrastructural ISH study of mRNA. PMID- 12114701 TI - Immunocytochemical Localization of Glucose Transporter-2 (GLUT-2) in Pancreatic Islets and Islet Cell Tumors. AB - Glucose is a major metabolic fuel in mammals and is transported into organs and cells by a facilitated diffusion which involves binding of glucose to glucose transporters (GLUTs). Among several GLUTs so far indentified, GLUT-2 is specifically localized immunocytochemV cally in beta-islet cells. Using immunocytochemical staining, normal pancreases and 27 cases of islet cell tumors, including insulinomas, gastrinomas, glucagonomas, pancreatic polypeptide-omas (PPomas), and a nonfunctioning islet cells tumor, were systematically stained for four different pancreatic hormones, gastrin, and GLUT-2. GLUT-2 staining in beta islet cells was more diffuse than that of insulin immunostaining, and corresponded with the positive staining in the lateral segments of beta-cell plasma membrane, that faced adjacent beta-cells. Glucagon, somatostatin (SRIF) and PP cells stained weakly for GLUT-2, weaker than that of beta-cells. Some nonbeta cells, especially extra-islet PP cells were not stained for GLUT-2. Among islet cell tumors, insulinomas stained less strongly for GLUT-2 than normal beta cells from the adjacent normal pancreas. Gastrinomas, glucagonomas, and PPomas stained weaker than insulinomas. Even nonfunctioning islet cell tumors were weakly stained for GLUT-2. The positive staining for GLUT-2 observed for islets cells and all islet tumors is consistent with the notion that all pancreatic islet cells and islet cell tumors utilize glucose as a major fuel, requiring transporter-facilitated diffusion of glucose into the cells of normal organ and their tumors. PMID- 12114702 TI - Thyroid Peroxidase Expression in Diseased Human Thyroid Glands. AB - Histoenzymologic studies of the human thyroid have been restricted to attempts to differentiate benign from malignant tumors and, to our knowledge, there is no systematic study on the distribution and behavior of enzymes for several types of lesions of the thyroid gland. The aim of the present study was to investigate thyroid peroxidase (IPO) expression in diseased thyroids, as well as to study the role of this enzyme as a parameter to define the biological behavior of thyroid lesions. A total of 1 17 neoplastic and nonneoplastic lesions of the thyroid were evaluated. The expression of TPO was determined by the analysis of at least 200 cells/specimen. A rate of 80% of positive cells was considered a threshold for TPO positivity. TPO expression was detected in all nonneoplastic lesions of thyroid as well as in 78.8% of the adenomas. We also observed positivity for TPO in 20% of malignant lesions. Therefore, these findings do not allow separation of benign and malignant lesions of the thyroid based in the expression of TPO. PMID- 12114699 TI - CARD In Situ Hybridization: Sights and Signals. AB - During the last decade, several strategies have been developed to improve the detection sensitivity of in situ hybridization (ISV) by amplification of either target nucleic acid sequences prior to ISH (e.g., in situ PCRX or the detection signals after the hybridization procedures (signal amplification). Here we outline the principles of tyramide signal amplification using the catalyzed reporter deposition (CARD) technique, summarize applications as well as possible limitations of CARD 15K, and discuss some future directions of in situ nucleic acid detection using this amplification strategy. PMID- 12114703 TI - Vascular Endothelial Growth Factor (VEGF) Expression in Human Pituitary Adenomas and Carcinomas. AB - Vascular endothelial growth factor (VEGF) is a key mediator of endothelial cell proliferation, angiogenesis, and vascular permeability. Little is known about its expression in human pituitary adenomas. We examined 148 human pituitary adenomas for VEGF protein expression by immunohistochemistry. The strongest immunoreactivity was present in GH adenomas, corticotroph, silent corticotroph. silent subtype 3, and nononcocytic null cell adenomas. GH adenomas treated with octreotide stained less intensely than did untreated tumors. Relatively weak staining was present in PRL, gonadotroph, thyrotroph, and oncocytic null cell adenomas in the same sections showed evidence of down-regulation of VEGE protein expression in adenomas. Pituitary carcinomas usually had stronger staining than adenomas. In situ hybridization studies with oligonucleotide probes showed positive staining in all groups with stronger staining in GH, ACTH, TSH, and gonadotroph adenomas and in pituitary carcinomas. These results indicate that VEGF expression is more prominent in certain adenoma subtypes, that decreased expression occurs in adenomas as compared to nontumorous pituitary and that carcinomas show increased VEGF expression relative to adenomas suggesting up regulation of VEGF during pituitary tumor progression. PMID- 12114704 TI - Xanthomatous Hypophysitis: A Novel Entity of Obscure Etiology. AB - We report a case of xanthomatous hypophysitis, a recently described entity of obscure etiology affecting the pituitary gland, in a 43-yr-old woman, Histologically it is characterized by infiltration of the anterior pituitary by foamy histiocytes which are strongly immunoreactive for CD68 (histiocytic marker) and are immunonegative for 5100 and CD 1 a. Electron microscopy revealed histiocytes with abundant cytopasmic lipid droplets and membrane bound vacuoles. Fragments of intact anterior pituitary with preserved vascw lar and reticulin networks are seen. Xanthomatous hypophysitis resembles neoplasm on clinical and radiologic grounds. PMID- 12114705 TI - The Clinicopathologic Significance of Unilateral Adrenal Cortical Hyperplasia: Report of an Unusual Case and a Review of the Literature. AB - We report a case of unilateral primary adrenal cortical hyperplasia associated with hyperaldosteromsm in a 39-yr-old Chinese man. The patient presented with hypertension hy pokalemia, primary hyperaldosteronism(1) suppressed renin activity, and was complicated with aortic dissection. The aortic dissection required emergency realignment while unilateral laparoscopic adrenalectomy was performed subsequently Pathologic examination of the adrenal lesion revealed multiple cortical nodules (up to 1 4 cm in diameter). No telomerase activity was detected in the adrenal lesion. A review of the literature showed that unilateral adrenal cortical hyperplasia has a predilection for the left side occurring more often in males. We conclude that unilateral adrenal cortical hyperplasia is a rare but unique entity and that unilateral adrenalectomy is the treatment of choice. PMID- 12114707 TI - Metastatic Follicular Variant of Papillary Thyroid Carcinoma Manifested as a Primary Renal Neoplasm. AB - Although autopsies performed on patients with cancer have shown 4 6 to 7.6% of metastases to the kidneys, the preoperative clinical identification remains rare. The primary sources of metastases to the kidney are in decreasing order of frequency: breast, lung, intestine, contralateral kidney, stomach, ovary, cervix, pancreas, uterus, and pro tate. As far as we know, only seven cases of malignant neoplasms of the thyroid gland metastatictothe kidney have been reported [1]. Herein, we report the eighth case of this event and the second of a follicular variant of papillary thyroid carcinoma whose initial manifestation was hematuria and flank pain. PMID- 12114706 TI - Myelolipoma: A New Adrenal Finding in Carney's Complex? AB - Pigmented nodular cortical hyperplasia. a rare cause of Cushing's syndrome, is characterized by resistance to inhibition with dexamethasone and normal sized adrenal glands with multiple, small, pigmented nodules. The disorder may be a component of a syndrome inherited as an autosomal dominant pattern that includes intra- and extracardiac myxomas, lentiginous lesions, blue nevi, other functional endocrire tumors, and perppheral nerve tumors (Carney's complex). We report a patient in whom bilateral myelolipomas were found, in addition to the usual features of this complex. A 29-yr old man was admitted to the hospital for Cushing's syndrome of probably more than 15 yr duration. Physical examination showed diffuse facial hyperchromatic macules. 02-05 cm, predominantly around the lips and on the palmar surfaces of the fingers. Results with dexamethasone suppression nocturnal testing (1 and 8 mg) were compatible with an adrenal adenoma. The computed tomography (CT) of the sella turcica was normal. Adrenal CT showed a tumor in the left gland with a double component one solid and another suggestive of fat, consistent with an angiomyelolipoma. Following 5 wk treatment with ketoconazole, 800 mg per day po, serum cortisol decreased to 5.9 Ag/dL. morning and evening, respectively. Bilateral adrenalectomy was performed. Pathologic examination revealed pigmented nodular cortical hyperplasia and a dominant myelolipoma in the left adrenal. A microscopic myelolipoma was identified in the right adrenal. An echocardiogram showed a mass on the posterior wall of the left ventricle which was a myxoma. Study of the patient's family disclosed two sisters with facial lentigines. Echocardiograms were performed on all available first degree relatives: all were normal. Nocturnal inhibition with dexamethasone revealed that one of the patient's sisters with lentigines also had hypercortisolism. Myelolipoma has been reported in association to Cushing syndrome in humans and experimentally after pituitary extracts in animals. The relationship between this finding and the Carney's complex remain elusive. PMID- 12114708 TI - ACTH-Producing Cholangiocarcinoma Associated with Cushing's Syndrome. AB - A 61-year~old woman was admitted to the hospital with clinical manifestations of Cushing's syndrome. The ACTH level was 1340 pglmL, the urinary free cortisol level > 900 pg/mL, and the serum K+ levels 21 meqlL. The brain/pituitary MRI and thoracic CT scan were normal. Gastroscopy, colonoscopy, and small bowel follow through were normal. Abdominal CT and MRI showed normal adrenals, but dilated gallbladder with numerous gallstones, as well as peripancreatic and hepatoduodenal lymphadenopathy. A large meta-static deposit and three smaller lesions were also seen in the liver. Because of the poor respiratory function tests and the severe hypokalaemia, laparoscopy under local anaesthesia was performed. Following the procedure the patient became gradually jaundiced and thus underwent exploratory laparotomy. Locally advanced cholangiocarcinoma was found, infiltrating the liver hilum, with multiple small bilateral liver metastatic deposits. Acute cholecystitis with pericholecystic abscess was also found. Cholecystostomy as well as gallbladder, liver and hilar node biopsies were performed. Histopathology showed liver adenocarcinoma of bile duct origin, while immunocytochemistry revealed scattered, chromogranin A positive cells, some of them strongly immunoreactive for ACTH. Small clusters of chromogranin A positive cells were also found to be immunoreactive for CRH, but not for ACTH. PMID- 12114709 TI - Catalyzed Reporter Deposition Method for Amplifying Endocrine Products. AB - The catalyze reporter deposition (CARD) method is a new technique to enhance signal detection in immunostaining, immunoelectron microscopy, and in situ hybridization. This technique, which uses a biotinylated tyramine amplification reagent, has been applied to endocrine cells and tumors, and has shown a marked increase in sensitivity in detecting small amounts of protein antigens in endocrine cells and tumors. Although some technical problems remain before widespread application of this method can be used the potential for more in depth understanding of normal and neoplastic endocrine tissue by CARD analysis is largely unexplored at this time. PMID- 12114710 TI - Discordant Expression of Ii and HLA-DR in Thyrocytes: A Possible Pathogenetic Factor in Hashimoto's Thyroiditis. AB - Hashimoto's thyroiditis is the archetype of organ-specific autoimmune disease. The key pathogenetic feature is the activation of thyroid-specific T-cells by properly presented endogenous thyroid antigens. There is strong indication that thyrocytes themselves present self-antigens, based on the finding of antigen presenting human leukocyte antigen-DR (HLA-DR) molecules on the thyrocytes in Hashimoto's thyroiditis. Because class ll-associated invariant chain (Ii) is tightly bound to the antigen-binding groove of the DR molecules in the endoplasmic reticulum and is found to compete with the endogenous peptides for binding with DR. it is speculated that Ii prevents endogenous peptides from binding to, and being presented by, DR on the cell surface. We hypothesize that in Hashimoto's thyroiditis, Ii is insufficient, leaving DR molecules open for endogenous peptides, In this article, we test our hypothesis of discordance between DR and Ii in the thyroid epithelial cells in Hashimoto's thyroiditis by immunohistochemistry. Formalin-fixed, paraffin-embedded archival tissue from eight cases of Hashimoto's thyroiditis and four cases of normal thyroid specimen were randomly selected and immunostained with monoclonal antibodies (MAbs) against HLA-DR and Ii using avidin-biotin-peroxidase method. We found that in all eight cases of Hashimoto's thyroiditis. Ii is significantly reduced relative to DR. The results support our hypothesis that presentation of self-antigen may be a result of insufficient Ii in the thyrocytes in Hashimoto's thyroiditis. PMID- 12114712 TI - The Thyroid in Acquired Immunodeficiency Syndrome. AB - Forty-seven thyroids obtained at autopsy from patients with acquired immunodeficiency syndrome (AIDS) with no clinical manifestations of thyroid disease were analyzed systematically in order to determine the frequency and the major pathological characteristics of thyroid involvement in these individuals. The glands were obtained from 38 men and 9 women aged (on average) 33.6 yr. The specimens were weighed, measured, and evaluated after fixation in formalin Histological examination was performed on at least 10 macroscopically normal and altered areas. The anatomopathological lesions detected in 29 glands (613%) were chronic nonspecific focal inflammation (482%); mycobacteriosis and colloid goiter (172%); histoplasmosis, cryptococcosis, and lipomatosis (133%), and paracoccidioidomycosis and hyperplastic nodules (3A%). Although thyroid disease had not been clinically diagnosed, thyroid involvement was elevated (613%) and in 14 cases (293%) it was related to the immunodeficiency, with mycobacteria being the most common opportunistic agents. There appears to be no report of the association of lipomatosis with AIDS, although this was a frequent finding in the present study (13 7%), exceeding by far the rates reported in the literature (1 2%). Thus, thyroid lesions are frequent in AIDS patients, occurring in two thirds of the patients studied, especially those with disseminated infection. PMID- 12114711 TI - Analysis of TGF-B and TGF-B-RII in Thyroid Neoplasms from the United States, Japan, and China. AB - Transforming growth factor B (TGF-B) has an inhibitory effect on cell proliferation in various cells and tumors, so loss of TG-B-receptor (TGF-B-R) may lead to increase proliferative activity in these tumors. We compared the expression of TGF-B and TGF-B-Rll in a group of thyroid neoplasms from the United States, Japan, and China to determine if there were differences in the expression of this growth factor or its receptors in various tumor types from different countries. A total 108 neoplastic thyroids from the United States, 42 from Japan, and 46 from China were analyzed for TGF-B1, TGF-B3, and TGF-B-Rll by in situ hybridization with riboprobes. TGF-jB-RII expression was also examined by immunohistochemistry. TGF-B1 mRNA was expressed in all neoplastic thyroids from all three countries except for one anaplasti carcinoma (ACA). TGF-B3 expression was lowest in follicular carcinomas (FCA) from all three countries (30/42; 71%). TGF-B-RII was much lower in FCA from Japan (112; 50%) and China (6/11; 55%) compared to cases from the United States (26/29; 90%). TGF-B-RII expression in papillary carcinoma (PCA) was also lower in carcinomas from Japan (21/28; 75%) and China (23/30; 77%) compared to the United States (24/25; 96%). Most ACA from the United States (25/30; 83%) and from China (3/3; 100%) were positive for TGF-B Rll. Immunohistochemical analysis for TGF-B-RII protein expression showed the highest levels in follicular adenomas (FA) (38/38; 100%) with decreased immunoreactivity in FCA (36142; 86%). PCA (66/83; 80%), and ACA (14/33; 42%). These findings suggest that loss of TGF-B--RII may be important in thyroid tumor progression and that environmental/geographic factors may play a role in the variable expression of TGF-B--RII in thyroid malignancy. PMID- 12114713 TI - Pathological Findings in Dyshormonogenetic Goiter with Defective Iodide Transport. AB - An adult male patient (38 yr old) with a large congenital goiter with hypothyroidism was suspected of having a defective iodide (L) transport mechanism based on low thyroid uptake and a very low salivary/plasma ratio. Moreover the high serum levels of TSH (104 uU/mL) declined to 7.2 uU/mL with a corresponding normalization of serum total T4 concentration, after 40 d of treatment with Lugolts solution (6 mg L/d). Thyroid surgery was performed because a fine-needle aspiration biopsy of a nodule revealed atypical cells associated with the presence of a large compressive goiter (150 g). Pathologic examination indicated histological findings compatible with the hyperplastic pattern with predominant microfollicular aspect. Immunostaining for other specific thyroid proteins thyroid peroxidase (TPO) and Tg, indicated normal expression of both transcripts. Electron microscopy (x13,000) showed the ultrastructural aspects of a hyperactive follicular cell with folding of the basal membrane. Sequencing of the entire sodium/iodide (Na/I) symporter (NIS) cDNA derived from thyroidal mRNA revealed a homozygous substitution of the normal cytosine in nucleotide 1163 with an adenine, resulting in a stop signal at codon 272. This nonsense mutation produces a truncated NIS symporter protein without iodide transport activity. Although the propositus is homozygotic for the NIS-272X expression of one normal allele in the heterozygotic son, mother, and paternal aunt is sufficient to maintain normal thyroid function. PMID- 12114714 TI - Hyperplastic and Tumorous Lesions of the Adrenals in an Unselected Autopsy Series. AB - In an unselected consecutive postmortem series, 512% of the adrenals showed cortical nodules with diameters up to 1 mm in 13%, between 1 and 2 mm in 28% and < 2 mm in 12%. In 22% of cases only one nodule was present; whereas in 15% two nodules and in 17% more than three nodules were found. A cortical adenoma was detected in 5% of cases. Adenomas were smaller than 6 mm in 1% and < 15 mm also in 1% of the entire collection. For differentiation of nodules criteria listed in Table 4 were used. Correlation with clinical data revealed a significantly higher number of nodules and an adenoma more frequently in hypertensive patients. Metastases were found in 19% of all cases with malignant tumors. Twenty-three percent of the metastases were smaller than 1 mm and 20% were < 10 mm in diameter. Adrenal metastases usually indicated generalized dissemination of the tumor. PMID- 12114716 TI - A Minimally Invasive Approach to the Management of Bronchial Carcinoid Tumors Associated with Ectopic Cushing's Syndrome. AB - Cushing's Syndrome is usually the result of a pituitary corticotroph or an adrenocortical adenoma. Rarely, an ectopic carcinoid tumor can elaborate sufficient amounts of adrenocorticotropic hormone (ACTH) to result in cortisol excess and Cushing's Syndrome. The diagnosis and management of these unusual neoplasms remains problematic. We describe two cases of Cushing's Syndrome where the diagnosis of ectopic disease was suspected on the basis of inferior petrosal sinus sampling (IPSS). We also describe a minimally invasive thoracoscopic approach to the resection of pulmonary carcinoid tumors. We believe that this technique offers a significant improvement over conventional thoracotomy for the management of potentially compromised patients with Cushing's Syndrome, while still permitting definitive histologic confirmation of the diagnosis. PMID- 12114715 TI - Fat-Cell Metaplasia in the Adrenal Cortex: Incidence, Structure, and Correlation to Basic Diseases in a Postmortem Series. AB - In a postmortem series of adrenals from 497 patients 25 cases (5%) showed foci of fat-cell metaplasia or bone-marrow metaplasia in the zona fasciculata or the zona reticularis. There was only one focus in 45% of adrenals with metaplasia, but in 36.4%. more than six foci were present. The sizes varied between 0.036 and 0.64 mm. The foci were composed of mature, mostly univacuolar, fat cells and various amounts of myeloid cells The incidence of fat-cell metaplasia or of bone marrow metaplasia correlated with arterial hypertension and severe coronary heart disease. In 76% of cases, nodular hyperplasia was demonstrable, but the metaplasia were more often found adjacent to, rather than within, the nodules. They appear to be related to metaplasia in adrenocortical tumors and to myelolipomas. The common pathogenesis is thought to be based on focal necroses in combination with local endocrine stimulation. PMID- 12114717 TI - Renal Cell Carcinoma Metastatic to the Thyroid Gland: A Comparative Molecular Study Between the Primary and the Metastatic Tumor. AB - A case of renal cell carcinoma metastatic to the thyroid gland is presented. The thyroid tumor showed typical features: clear cells arranged in nests with a prominent sinusoidal vascular pattern. The histologic appearance was identical to the renal tumor removed 6 yr before. A comparative molecular study between the primary and the metastatic tumor showed a common profile with a loss of heterozygosity at identical loci on chromosome 3, which provided further support to the metastatic nature of the thyroid tumor. However, the frequent existence of allele losses on chromosome 3p in both renal cell carcinomas and primary thyroid follicular carcinomas rest some force from such a molecular approach to the differential diagnosis of thyroid tumors that contain a predominant population of clear cells. PMID- 12114718 TI - Remodeling of Hyperplastic Pituitaries in Hypothyroid us-Subunit Knockout Mice After Thyroxine and 1713-Estradiol Treatment: Role of Apoptosis. AB - Hyperplasia of pituitary thyrotrophs is often associated with hypothyroidism. In this study. the effects of thyroxine and 1 7B-estradiol on thyrotroph hyperplasia was analyzed using a hypothyroid mouse model resulting from targeted disruption of the glycoprotein hormone a-subunit (aSU) gene, which leads to lack of functional thyroid-stimulating hormone (TSH), luteinizing hormone (LH), and follicle-stimulating hormone (FSH) and underdevelopment of the thyroid and gonads. Thyroxine replacement for 2 mo resulted in a decrease in the relative percent of thyrotrophs and an increase of lactotrophs and somatotrophs numbers to normal values. A twofold increase in the relative percent of gonadotrophs was observed compared to wild-type mouse pituitary. Treatment for 2 mo with 17B estradiol led to an increase in lactotroph numbers to normal levels, but had no influence on thyrotroph hyperplasia. Rearrangement of the hyperplastic pituitary phenotype after hormonal replacement proceeded without any evidence of pituitary cell necrosis. A slight increase in apoptotic cell death was observed in hormone treated pituitaries, and this was localized to TSH cells by double-labeling experiments. Chronic thyroxine treatment resulted in increased expression of Bcl 2 protein in hypertrophied pituitary cells, whereas 17f3-estradiol increased expression of Bad protein in prolactin cells. These results suggest that apoptotic cell death is involved in reversal of thyrotroph hyperplasia in the presence of thyroid hormone. Thyroxine and 17-estradiol may influence cell death in this model by regulating expression of the Bcl-2 protein family in a celltype specific manner. PMID- 12114719 TI - Primitive Neuroectodermal Tumor of the Meninges: An Histological, Immunohistochemical, Ultrastructural, and Cytogenetic Study. AB - We report a case of primitive neuroectodermal tumor (PNET) arising from the meninges in a 30-yr-old female patient whose clinical and radiological features were consistent with meningioma. The neoplasm was composed of primitive small, round cells, growing in sheets and nests in continuity with meningeal layers. Ultrastructurally, the neoplastic cells were characterized by large and regular nuclei, primitive cytoplasm with pools of glycogen, and lack of dense core granules. The neuroectodermal nature of the tumor was confirmed by positive immunostaining for vimentin, neurofilaments, neuron specific enolase (NSE), and 013 (an antibody raised against MIC2 antigen). Further support to the diagnosis was obtained by reverse transcriptase-polymerase chain reaction (RT-PCR) detection of Chromogranin A and Secretogranin II genes. t(1 1;22) translocation was also observed by RT-PCR, a finding that was not recorded in previously described intracranial PNET. The tumor followed a malignant course, recurring and spreading to chest wall and sacroiliac region over a 10-yr period. The meningeal location enlarges the topographic spectrum of intracranial PNET, and this tumor has to be considered in the differential diagnosis with meningioma. PMID- 12114721 TI - Proliferative Activity in Pancreatic Endocrine Tumors: Association with Function, Metastases, and Survival. AB - Endocrine tumors of the pancreas are slow-growing lesions, yet one-third to one half will metastasize. It is generally accepted that histopathologic features do not reliably predict metastatic potential or outcome. We investigated whether proliferative activity, as determined by MIB-l labeling, correlated with tumor type, metastasis, or patient survival. Formalin-fixed sections of pancreatic endocrine tumors were immunohistochemically stained for the MIB-l antibody against Ki-67 using the avidin-biotin complex technique. Labeling index (LI) was determined by counting 1000 consecutive tumor cells in an area of greatest staining intensity at x400 and expressed as a percentage. The study group included 37 patients, including 10 gastrinomas, 9 insulinomas, 4 glucagonomas, 2 VlPomas, and 12 nonfunctioning tumors. Twenty-one patients had metastases, primarily to regional lymph nodes and the liver. Five patients had MEN I. MIB-1 LI was significantly greater in the nonfunctioning tumors (mean 20S%) than in the functioning tumors (mean 51%) (p = 0.01). LI for functional tumors (insulinomas 64%, glucagonoma 44%, gastrinomas 32%, VlPomas 32%) were similar to each other, MIB-1 was significantly higher in those tumors that metastasized (mean 15.6%) compared to those that did not (mean 31%), (p = 0.04). All tumors with MIB-1 LI >10% developed metastases. Logistic regression showed that MIB-1 was a significant predictor of metastases (p = 0.003) after adjusting for functional status. MIB-1 LI also correlated with outcome in that those patients with MIB-1 LI >/=10% had a mean survival of 19 mo compared to 72 mo for those with levels <10% (p = 0.0001). Results of the proportional hazards model showed that MIR-1 remained a significant (p = 0.03) and independent predictor of survival times after adjustment for tumor size and functional status. Higher MIB-1 LI values were significantly associated with shorter survival times. In conclusion, MIB-1 LI appears to be a useful indicator of metastatic potential and is predictive of outcome in PET. PMID- 12114720 TI - Pituitary Insulin: Insulin-Like Growth Factors. AB - Insulin-like growth factor (IGF)-l and IGF-ll peptides as well as their mRNAs are produced in many organs, including the pituitary. Although IGFl and IGFII peptides are localized in endocrine cells of the anterior pituitary, IGF-l mRNA can be detected throughout the adenohypophysis, and IGF-ll mRNA is abundant in intermediate and neural lobes. It is well-established that both circulating and intrinsic IGF-l are negative regulators of pituitary GH production. Other functions of intrinsic IGFs in normal and tumorous pituitary are just emerging. IGF-l may play a role in the stimulation of PRL synthesis and mediation of proliferative effects of estrogen on lactotroph. Compared with IGF-l, the function of IGE-Il has not been clarified so far. The growth-promoting actions of IGFs are mediated by IGF-l receptor. The role of local and circulating IGFBPs in pituitary are not yet documented. IGFBPs in other tissues have inhibitory and stimulatory effects on IGFs, and can act independently from the IGFs as well. IGFs have been reported to promote cell proliferation in many tumors. However, the extent to which IGFs contribute to pituitary tumor development and growth remains obscure. PMID- 12114722 TI - Immunocytochemical Investigations on the Vascularization of Pituitary Adenomas. AB - Blood vessels within pituitary adenomas were visualized using the immunocytochemical reaction for Factor VIII (von Willebrand Factor), a specific marker of the vascular endothelium. The number of immunopositive vascular profiles were counted and expressed as a mean number per one microscopic field. The results were related to the type of adenoma, established on the basis of immunocytochemical investigation using the antibodies against pituitary hormones or a-subunit (a-SU). It was found that the richest vascularization occurred in adenomas expressing follicle-stimulating hormone (FSH). The possible role of FSH in pituitary angiogenesis is discussed. PMID- 12114725 TI - Fine-Needle Aspiration in the Evaluation of Thyroid Neoplasms. AB - Fine-needle aspiration (FNA) is a safe, rapid, and accurate diagnostic tool. Although it continues to gain acceptance, the pace is slow. Probably, if more pathologists master the basics (i.e., learn how to obtain a good sample), this simple technique could be utilized to its fullest advantage. If the sample is not adequate or representative of the lesion, the diagnosis will not be correct. Based on personal experience, we believe that suction should be minimal when obtaining thyroid aspirates. Cytologic diagnostic criteria for the most common neoplasms of the thyroid gland are provided. PMID- 12114724 TI - Investigation of the Presence of Apotipoprotein E, G lycosaminoglycans, Basement Membrane Proteins, and Protease inhibitors in Senile interstitial Amyloid of the Pituitary. AB - The aim of our study was to test whether local- or organ-limited interstitial amyloid of the pituitary is associated with the presence of glycosaminoglycans, basement membrane proteins, protease inhibitors, and apolipoprotein E (apo E), as previously observed in other amyloid syndromes. Serial sections from amyloidotic and nonamyloidotic autopsy pituitaries of patients age 85 yr and over were stained with Congo red, Alcian blue, and, applying immunohistochemistry, with antibodies directed against fibronectin, collagen IV, laminin, apo E, a(1) antitrypsin and a(1)-antichymotrypsin. Interstitial amyloid was deposited in the immediate vicinity of capillaries and around the acini of the anterior lobe. Glycosaminoglycans were found in capillaries and around the acini of both nonamyloidotic and amyloidotic glands and they were also related spatially to amyloid deposits. Immunostaining of nonamyloidotic and amyloidotic glands demonstrated the presence of fibronectin, collagen IV, and laminin, which was related to basement membranes (fibronectin, collagen IV, and laminin), interstitium, and serum (fibronectin only). In amyloidotic glands, each basement membrane protein presented with an additional spatial relationship to amyloid deposits. Apo E was found in amyloidotic cases only within the amyloid deposits. The results are consistent with the presence of glycosaminoglycans, basement membrane proteins, and apo E in local interstitial amyloid deposits of the pituitary, as previously described in other amyloid syndromes, such as inflammatory related AA-amyloidosis or AB-amyloidosis related to Alzheimer's disease. PMID- 12114723 TI - Pituitary Changes in Ataxia-Telangiectasia Syndrome: An Immunocytochemical, In Situ Hybridization, and DNA Cytometric Study of Three Cases. AB - Ataxia-telangiectasia (AT) syndrome (cerebellar ataxia, oculocutaneous telangiectasias, immunodeficiency, susceptibility to infections, and neoplasia) is associated with cyto- and nucleomegaly in several organ systems. Our aim was to determine (1) whether such cellular abnormalities in the pituitary selectively involve specific cell types, and (2) the proliferation and DNA ploidy status of such cells. Three AT autopsy pituitaries were studied by histology, immunohistochemistry (pituitary hormones, MIB-1, p53 protein), in situ hybridization (pituitary hormones), and Feulgen stain image analysis for ploidy. Results indicated that, in adenohypophyses the scattered pleomorphic, bizarre nuclei were mainly those of somatotrophs and corticotrophs, growth hormone (GH), or adrenocorticotropic hormone (ACm) immunoreactive and expressing the GH or ACTH gene, respectively. Cyto- and nucleomegaly were less frequent in other secretory cells but were also noted in pituicytes of the posterior lobe. Affected cells were immunonegative for MIB-1 and for p53 protein. Image morphometric DNA analysis showed the bizarre cells to be aneuploid with complex histogram patterns, including many nuclei with DNA contents >8 n. No adenomas were found. We conclude that in AT adenohypophyseal cells with cyto- and nucleomegaly, as well as pleomorphism, synthesize and store adenohypophyseal hormones, mainly GH or ACTH. They and affected pituicytes are nonproliferative and are aneuploid. PMID- 12114726 TI - The Relationship of Lymphocytic Thyroiditis to the Development of Thyroid Carcinoma. AB - There are many reported predisposing factors for thyroid papillary carcinoma, including genetic factors [1] previous irradiation [2,3], abnormal iodine intake [4-6], hyperthyroidism [7], pregnancy [8], and a dyshormonogenetic state [9]. However, whether there is a causative link between lymphocytic thyroiditis and thyroid papillary carcinoma remains controversial. Most reports indicating some association have suffered from methodologic problems. In particular, race, age, and sex differences in susceptibility to chronic lymphocytic thyroiditis and thyroid cancer have not been well-controlled in previous studies. We have therefore carefully evaluated previous studies and tried to determine if there is a definite relation between lymphocytic thyroiditis and thyroid papillary carcinoma histo-pathologically. PMID- 12114728 TI - RET Proto-Oncogene and Thyroid Cancer. AB - The RET proto-oncogene has not only conclusively been identified as responsible for the three subtypes of the inherited cancer syndrome multiple endocrine neoplasia type 2 (MEN-2) but also shown to be involved in the molecular evolution of sporadic medullary and papillary thyroid carcinoma as well as Hirschsprung's disease. A variety of recent studies have elucidated the pathophysiological mechanisms leading to neoplastic disease and we now understand that dominant activating germline mutations lead to MEN-2A, MEN-2B, and familial MTC; somatic mutations to sporadic medullary thyroid carcinoma; RET rearrangements to papillary thyroid carcinoma; and inactivating alterations to Hirschsprung's disease. The clinical significance, however, of RET alterations especially in sporadic thyroid tumors is still controversial and therapeutic concepts in MEN-2 gene carriers only start to emerge. This article is a short summary of the recent findings on the structure and physiology of the RET proto-oncogene and its role in familial and sporadic thyroid cancer. PMID- 12114727 TI - Minimally Invasive Follicular Carcinoma. AB - The criteria for the diagnosis of follicular carcinoma of the thyroid and in particular of the "minimally invasive" variant are discussed. The introduction of a new terminology reflecting the indolent behavior of most of the thyroid neoplasms morphologically resembling follicular adenomas, and currently diagnosed as carcinomas on the sole basis of capsular invasion, is recommended. PMID- 12114729 TI - TGF-B and Estrogen Regulate P27(Kip1) and Cyclin D(1) in Normal and Neoplastic Rat Pituitary Cells. AB - Pituitary hyperplasia and tumor growth are regulated by various hormones and growth factors. Estrogen (E(2)) stimulates pituitary cell proliferation and prolactin (PRL) production. Estrogen also regulates transforming growth factor-B (TGF-B) effects in the pituitary. IGF-B in turn regulates various cell cycle proteins including p15 and p27(Kip1) (p27). To better understand the regulatory role of growth factors and hormones in the cell cycle we analyzed cyclin D(1), cyclin E, and p27 expression in normal and neoplastic rat pituitary cells. An in vitro analysis using cultured normal pituitary cells and GH(3) tumor cells and an in vivo analysis of estrogen-treated normal pituitary and implanted GH(3) cells were performed. Semiquantitative RT-PCR was used to analyze mRNA expression for cyclin D(1) cyclin E, and p27 in cultured pituitary cells and E(2)-treated pituitaries in vivo, Cyclin D(1) and p27 were localized in the nuclei of normal pituitary cells by immunocytochemistry (ICC). Very weak or absent immunostaining for cyclin D(1) and p27 was present in GH(3) cells. Both normal pituitary and GH(3) cells had strong nuclear staining for cyclin E. Normal pituitary had a 20 fold greater amount of cyclin D mRNA and a 3-fold greater amount of p27 mRNA compared to GH cells, whereas GH cells had slightly (1.5-fold) more cyclin E than normal pituitary cells. Treatment in vivo stimulated cell proliferation and decreased cyclin D(1) mRNA levels in normal pituitary. GH(3) tumor cells, implanted subcutaneously in the same animal, showed increased proliferation after E(2) treatment, but there was no change in cyclin B(1) mRNA in GH(3) cells. Cyclin E and p27 mRNA levels did not change significantly in normal pituitary or in GH(3) cells after E(2) treatment in vivo. Treatment of normal pituitary cells with 10(-9)M TGF-B1 for 3 d in vitro led to significant decreases in cyclin B(1) and p27 mRNAs (p < 0.05 ), whereas cyclin E levels were unchanged. These results indicate that cyclin B(1) and p27 mRNAs are present at significantly higher levels in normal pituitary compared to GH(3) cells, and that both E(2) and TGF-B1 can down-regulate cyclin B(1) mRNA levels in normal pituitary cells, suggesting that these factors regulate G1 to S phase transition in pituitary cells. The lower levels of specific cell cycle regulators in GH cells may explain the decreased regulatory control by E(2) in GH(3) tumor cells. PMID- 12114730 TI - Pancreatic Neuroendocrine Carcinoma Metastatic to the Breast as Part of the Multiple Endocrine Neoplasia Type 1 Syndrome. AB - Breast metastases from nonmammary tumors are rare. We report here the first case of pancreatic neuroendocrine carcinoma metastatic to the breast in a patient with possible multiple endocrine neoplasia type 1. The diagnosis was supported by histological examination, immunohistochemistry, and ultrastructural analysis. This observation emphasizes the importance of clinical data for an accurate diagnosis, especially during intraoperative examination. When pathologists are faced with an unusual breast tumor larger than 2 cm, we would recommend freezing and/or saving pieces of tissue for ultrastructural analysis, which might help in the diagnosis. PMID- 12114732 TI - Interphase Analysis of Chromosome 11 in Human Pituitary Somatotroph Adenomas by Direct Fluorescence In Situ Hybridization. AB - Chromosome 11 numerical abnormalities were detected by fluorescence in situ hybridization (FISH) technique in four surgically removed pituitary somatotroph adenomas from patients clinically associated with acromegaly. The tumors were diagnosed by histology. immunocytochemistry, and electron microscopy, and included two densely granulated somatotroph (DG-SM) and two sparsely granulated somatotroph (SG-SM) adenomas. For demonstration of chromosome 11, the direct FISH technique was applied on imprints from fresh adenoma tissue fixed in acetone using an a-satellite specific centromeric probe. The slides were studied with a fluorescence microscope and the percentages of positive nuclei with aberrant fluorescent signals were counted. All tumors exhibited numerical chromosomal abnormalities in 8-23% of their cell population and included nuclei containing 1 3 extra chromosome 11 copies. The SC-SM adenomas exhibited more prominent abnormalities compared with the DG-SM adenomas. We conclude that numerical chromosome 11 abnormalities represent a frequent event among somatotroph adenomas with a tendency for higher frequency in SC-SM adenomas. PMID- 12114733 TI - Nitric Oxide Synthase in Human Parathyroid Glands and Parathyroid Adenomas. AB - Nitric oxide (NO) is a novel gaseous intercellular transmitter thought to play important physiological roles in the regulation of blood flow and hormone secretion in, for example, the pituitary, the thyroid, and the endocrine pancreas. Whether nitric oxide synthase (NOS) is present in the human parathyroid glands has not yet been demonstrated. In the present study, histologically normal, but functionally suppressed human parathyroid glands and parathyroid adenomas from patients with primary hyperparathyroidism were investigated by immunocytochemistry with antibodies against neuronal NOS and by reduced nicotinamide adenine dinucleotide phosphate (NADPH) diaphorase histochemistry. We also used H&E to identify the NOS-immunoreactive cells. Immunocytochemistry demonstrated the presence of neuronal-type NOS in a subpopulation of glandular cells, identified as oxyphilic cells, in both normal parathyroid glands and adenomas. NADPH-diaphorase staining visualized NOS in the endothelium of blood vessels and in glandular cells, corresponding to those containing immunoreactive NOS. In addition, we found NADPIH-diaphorase staining in many chief cells. Our results indicate that both glandular cells and vascular endothelium in human parathyroid glands and adenomas express NOS. There is thus a morphological substrate for locally produced NO that may be involved in the regulation of parathyroid blood flow and hormone secretion. PMID- 12114735 TI - Lipomatous Changes in Adrenocortical Adenomas: Report of Two Cases. AB - Rare cases of myelolipomas associated with adrenocortical lesions responsible for Cushing or Conn syndromes have been described. We report two additional cases of extensive lipomatous changes in incidentally discovered adrenocortical adenomas, which were preoperatively interpreted as myelolipomas on the basis of radiologic findings. Microscopically, the adenomas were composed of sheets and nests of adrenocortical cells, and extensive areas of mature adipose tissue admixed with a bland stromal infiltration of small cells. The impression was that myeloid cells were present, featuring a myelolipoma associated with a clear cell adenoma of the adrenal cortex, but specific immunocytochemical markers of myeloid lineage were not reactive in the small cell component, and these cells consisted of small lymphocytes. The lipomatous tissue may represent a degenerative phenomena within an adrenocortical adenoma or may be an additional neoplastic component of the tumor. Irrespective of their origin, extensive (myelo)lipomatous changes in adrenocortical tumors can lead to misinterpretations in the preoperative work-up of patients with adrenal masses. PMID- 12114734 TI - Assessment of Mitotic Activity in Pituitary Adenomas and Carcinomas. AB - Assessment of mitotic activity represents one of the oldest and most routinely used histopathologic methods of evaluating the biological aggressiveness of human tumors. In the case of pituitary tumors, however, the relevance of this approach as a means of gauging tumor behavior remains ill-defined. In this article, the relationship between the mitotic index and biological aggressiveness of pituitary tumors was evaluated in a series of 54 pituitary adenomas and 6 primary pituitary carcinomas. All tumors were fully classified by immunohistochemistry and electron microscopy; adenomas were further stratified on the basis of their invasion status, the latter being defined as gross, operatively, or radiologically apparent infiltration of dura or bone. Mitotic figures were present in 11 tumors, 10 being either invasive adenomas or pituitary carcinomas. A significant association between the presence of mitotic figures and tumor behavior was noted, as evidenced by progressive increments in the proportion of cases expressing mitotic figures in the categories of noninvasive adenoma, invasive adenoma, and pituitary carcinoma (3.9, 21.4, and 66.7%, respectively; Fisher's exact test, two tailed, p < 0.001). The mitotic index, however, appeared to be a less informative parameter, being extremely low in all cases (mean = 0.016% +/- 0.005 [+/- SEMI). Although the mean mitotic index in pituitary carcinomas (0.09% +/- 0.035) was significantly higher than the mean mitotic index of either noninvasive adenomas (0.002% +/- 0.002) or invasive adenomas (0.013% +/- 0.005), no practical threshold value capable of distinguishing these three groups was evident. Comparison of the mitotic index with Ki-67 derived growth fractions in these tumors revealed a significant but weak linear correlation (r = 0.41, p < 0.01). These data suggest that when, mitotic figures are present, they do provide some indication of the behavior and invasive potential of pituitary tumors. For routine diagnostic purposes, however, the discriminating power of this parameter is somewhat limited, being superseded by alternative and more informative methods of growth fraction determination such as that provided by the Ki-67 immunolabeling. PMID- 12114736 TI - Localization of Activin A/EDF in the Normal and Diabetic Rat Pancreas: Immunohistochemical and In Situ Hybridization Studies. AB - We examined the immunohistochemical localization of activin A/erythroid differentiation factor (EDF) in the pancreases of normal and diabetic rats with insulin-dependent diabetes mellitus or noninsulin-dependent diabetes mellitus to elucidate how activin A/EDF modulates insulin secretion. In both the normal and the diabetic pancreas, all of the cells staining with antirecombinant human (rh) activin A/EDF antiserum were insulin-producing B-cells, and they gradually decreased in number with the development of diabetic changes in both types of diabetic rats. No rh activin AIEDF immunoreactivity was detected in A-, D-, or pancreatic polypeptide (PR) cells in the islets, or in the exocrine cells. The in situ hybridization (ISH) method showed that activin A/EDF mRNA is localized in activin A/EDF-positive cells. These data indicated that activin A/EDF localizes in insulin-producing B-cells of normal and diabetic pancreases, and may act as an autocrine modulator of insulin secretion. PMID- 12114737 TI - The Pituitary in Isolated ACTH Deficiency: A Histologic, Immunocytochemical, and In Situ Hybridization Study. AB - A 74-year-old man presented in a near terminal state with progressive generalized muscular weakness, gastrointestinal disturbances, and lethargy. Investigations revealed hypotension, hyponatremia, hypoglycemia, and low plasma cortisol concentration accompanied by undetectable plasma adrenocorticotropic hormone (ACTH) level. The patient died shortly after admission to hospital, with adrenocortical failure being the provisional cause of death. Autopsy disclosed profound bilateral atrophy of adrenal cortices with evidence of a mild focal inflammatory reaction. The pituitary gland appeared normal on both gross and histologic examinations. There was no histologic evidence of inflammation, fibrosis, or adenohypophysial cell hyperplasia. By immunocytochemistry, no ACTH and B-endorphin immunoreactive cells were identified in the adenohypophysis. In situ hybridization (ISH) for pro-opiomelanocortin (POMC) mRNA yielded conclusively negative results. The case presented here was regarded as isolated ACTH deficiency. Although the remaining pituitary functions were not assessed, clinical and morphologic findings strongly support the supposition that aside from ACTH deficiency, secretory function of other pituitary hormones was preserved. This is the first case in which the pituitary was studied by immunocytochemistry and ISH. The possible pathogenetic mechanisms accounting for the isolated ACTH deficiency are discussed. PMID- 12114738 TI - The Pituitary in Gigantism. AB - To compare the pituitary pathology of gigantism to that of acromegaly, 19 surgically resected lesions were studied from 10 males and 9 females, ages 13-49 (mean, 19 yr) with excessive height (>/=95th percentile), onset of disease prior to puberty, elevated growth hormone (GH) levels despite glucose suppression, and a pathologically confirmed GH-producing pituitary mass. One patient had MEN-I. The lesions included 18 adenomas and 1 case of pure hyperplasia. The median, mean, and range of serum GH and prolactin (PRL) levels were 64, 235, 5-1000 ng/mL and 47, 146, 29-770 ng/mL, respectively. Of the 8 adenoma specimens accompanied by nontumoral pituitary (i.e., tissue wherein the presence of hyperplasia was assessable), 3 (37%) demonstrated both. Of the 18 tumors, 78% were macroadenomas and 22% were grossly invasive; their immunophenotypes included GH (5%), GH and PRL (19%), and GHPRL and a glycoprotein hormone, usually TSH and/or a-subunit (76%). Of the 10 adenoma-containing lesions subject to electron microscopy (EM), 2 consisted of GH cells alone; 2 of mammosomatotroph (MS) cells alone; 1 of GH and MS cells; 1 of GH and PRL cells; 2 of GH, PRL, and MS cells; 1 of GH, PRL, and glycoprotein cells; and 1 was a subtype 3 adenoma. Ultrastructurally, GH cells and/or MS cells predominated in these lesions. Immuno-EM of one CH and PRL cell and of one GH-PR-MS tumor showed GH and PRL to be present not only in single cells but within the same granules. Nine of 12 adenoma-associated lesions subject to combined in situ hybridization (ISH) and immunostaining showed double labeling for PRL (or GH) mRNA and for GH (or PRL), respectively, features indicating MS differentiation. In the 4 lesions exhibiting hyperplasia, either alone (1) or in association with adenoma (3), EM showed MS cells in 3, and immuno-EM as well as combined immunohistochemistry and ISH showed double labeling for GH and PRL in both of the 2 cases studied. In summary, although in terms of their tinctorial characteristics and tumor size, the lesions of giants resemble those of acromegalics, those of the former are less often invasive and glycoprotein hormone containing, and more often contain ultrastructurally distinctive MS cells. The high frequency of adenoma with hyperplasia (37%) and the occurrence of hyperplasia alone (6%) is of particular notice since this finding is rare in patients with acromegaly. Hyperplasia is, however, seen in ectopic GH-releasing hormone production and the McCune-Albright syndrome. We conclude that the presence of MS is not rare in the pituitary lesions of patients with gigantism. Their presence may be a reflection of either hypothalamic dysfunction or of an intrinsic abnormality of pituitary cells, PMID- 12114731 TI - Pulmonary Neuroendocrine Cells and Lung Development. AB - Pulmonary neuroendocrine cells produce bioactive peptides such as gastrin releasing peptide (GRP) at high levels in developing fetal lung. The role of GRP and other peptides in promoting branching morphogenesis, cell proliferation, and cell differentiation during lung organogenesis is reviewed. Possible roles for bioactive peptides derived from these cells in the pathophysiology of perinatal lung disorders are discussed. PMID- 12114739 TI - Detection of Heterozygous Mutation in the Retinoblastoma Gene in a Human Pituitary Adenoma Using PCR-SSCP Analysis and Direct Sequencing. AB - Genomic deoxyribonucleic acid from surgical specimens of 25 pituitary adenomas was screened for the presence of mutations in the tumor suppressor gene retinoblastoma gene, using polymerase chain reaction and single-strand conformation polymorphism analysis, followed by direct deoxyribonucleic acid sequencing. Mutation causing an amino acid change was found in one of the 25 pituitary adenomas. The mutation site was in exon 19 (codon 621) of the retinoblastoma gene. In addition, there were three types of silent mutations in introns of the gene. The patient in whom the retinoblastoma mutation was identified had a tumor with high clinical malignancy, a high percentage of c-myc protein-labeled cells, and a diagnosis of plurihormonal pituitary adenoma based on the presence of cells immunoreactive for five pituitary hormones. This article suggests that point mutation of retinoblastoma gene is rare in human pituitary adenomas but may provide a marker for aggressive pituitary adenoma. PMID- 12114740 TI - HLA-D Antigen Expression and Langerhans' Cell Infiltrates in Thyroid Tumors. AB - Papillary carcinomas (PCs) of thyroid are among the most common but least aggressive human malignancies. The factors explaining the indolence of these tumors are unknown but host-tumor immune interactions may play a role. This study was designed to determine if there is morphologic evidence of these. Frozen tissues collected from 21 PCs, 4 follicular adenomas (FAs), 4 follicular carcinomas (FCs), and 11 nodular hyperplasias (NHs) were stained immunohistochemically for HlLA-D antigens, lymphocyte, and macrophage markers; results were graded numerically. Paraffin-embedded tumors (35 PCs, 10 FAs, and 10 FCs) were stained for 5-100 protein to detect Langerhans' cells (LCs). Diffuse staining for HLA-D antigens and heavy mononuclear infiltrates were found more commonly in PCs compared to follicular neoplasms (FNs) or NHs. No consistent relationship was found between lymphocyte/macrophage infiltrates and expression of HLA-D antigens. The largest number of LCs was in PCs (median 11.8 cells/standard microscopic field [c/smf]), fewer cells were found in FA (3.7 c/smf), and the least in FC (0.05 c/smf). Features of host-tumor interaction including HLA-D expression and infiltrates with lymphocyte macrophages and LC are more strongly expressed in PC than other tumors. This may play a role in explaining their biological behavior. PMID- 12114742 TI - Immunohistochemical Heterogeneity Within Clinically Nonfunctioning Pituitary Adenomas. AB - To investigate whether adenohypophysial hormone expression is heterogeneous within individual clinically nonfunctioning pituitary adenomas, immunohistochemical examinations were performed on tissues obtained by multiple sampling of 11 adenomas. Stained sections were assessed by morphometric image analysis as well as semiquantitative estimation under microscopy. All tumors except one were immunopositive for one or more gonadotropins. Results were divided into five grades based on the proportion of immunoreactive cells per section. Semiquantitative estimation showed only a one-grade difference among samples from the same tumor in four cases for FSHB and in two cases for LHB. These qualitative similarities between multiple samples were confirmed by morphometric image analysis. From the practical standpoint of making a diagnosis of nonfunctioning pituitary adenoma, it is not necessary to take into account immunohistochemical heterogeneity within an individual tumor, and immunohistochemical findings in a given sample obtained at surgery can be regarded as representative of the entire adenoma. PMID- 12114743 TI - Immunolocalization of Parathyroid Hormone in Human Parathyroid Glands with Special References to Microwave Antigen Retrieval. AB - The subcellular localization of parathyroid hormone (PTH) in the normal human parathyroid glands with particular reference to microwave antigen retrieval was investigated using peroxidase-labeled PTH antibody, immunohistochemical, and immunoelectron microscopic methods. The results revealed that PTH granules existed mainly as pro-PTH on the trans side of Golgi and in the regions adjacent to Golgi apparatus. Only a small proportion of secretory granules were stored near the plasma membrane. Microwave irradiation was essential for the immunodetection of PTH. As the irradiative time extended from 1 to 30 min, the staining intensity increased, and the subcellular preservation decreased. Microwave irradiation for 15 mm (with the sections in citrate buffer) with a power output of 500 W is the most ideal for PTH antigen retrieval, as well as for subcellular preservation. PMID- 12114741 TI - Insulin-Like Growth Factor I in Human Thyroid Tissue: Specific Localization by Immunohistochemistry and In Situ Hybridization. AB - Insulin-like growth factor I and II (IGF-l and IGF-ll) have been implicated in the replication of normal thyroid follicular cells in vitro. This study evaluates the distribution and abundance of immunoreactive IGF-l by histochemical analysis in human thyroid tissue with different histopathologic characteristics. We used two types of highly specific and sensitive polyclonal rabbit anti-IGF-l antibodies and one monoclonal antibody (MAb) with the immunoperoxidase technique on sections of 25 glands harboring adenomatous goiter; 11 glands with follicular adenoma (FA); 45 glands with thyroid carcinoma of papillary, follicular, and undifferentiated types; and 18 glands with Graves' disease. Immunoreactive IGF-l was present in some thyroid follicular cells of all thyroid tissues examined. The percentage of cells staining positively varies among the different processes, being lowest in normal thyroid tissues and highest in all thyroid carcinomas. The cytoplasmic pattern of IGF-l immunoreactivity also varied among the different thyroid conditions. Furthermore, using nonradioactive in situ hybridization (ISH) we detected IGF-l mRNA in the thyroid cells of adenomatous goiter. The expression was higher in the histologically hyperplastic areas. These findings provide further support for an autocrine and/or paracrine role of IGF-l in the function and/or growth of normal thyroid follicular cells and suggest that IGF-l may play a role in the dysfunctional growth of thyroid follicular cells in adenomatous goiter, thyroid carcinoma, and Graves' hyperthyroidism. PMID- 12114744 TI - An Unusual Case of Multiple Gastric Carcinoids Associated with Diffuse Endocrine Cell Hyperplasia and Parietal Cell Hypertrophy. AB - We describe a case of multiple gastric carcinoid tumors in a 47 year-old Japanese man. The patient had markedly elevated serum gastrin levels (>800 pg/mL), which were suppressed by secretin-pancreozymin administration. In the partial gastrectomy specimen, a total of 19 carcinoids arose from diffuse linear and micronodular hyperplasia of the oxyntic mucosal endocrine cells. The carcinoid cells were chromogranin A-positive. Except for a very small number of serotonin positive cells in several carcinoids, none of the 19 reacted with a battery of antibodies to other bioactive neuroendocrine substances. The most prominent findings in this case were peculiar fundic glands that were distended with a proteinaceous substance and lined with large hypertrophic parietal cells. At the ultra-structural level, these cells showed poorly developed intracytoplasmic canaliculi and vesicotubular profiles, yet their large cytoplasm had numerous mitochondria. On the basis of our histological and ultrastructural findings we suggest that an intrinsic HCI secretion abnormality of the parietal cells may be responsible for the patient's hypergastranemia. Since there have been no reports on similar parietal cells changes, it is possible that our case may represent a pathological entity not previously described. PMID- 12114745 TI - An Atypical Acidophil Cell Line Tumor Showing Focal Differentiation Toward Both Growth Hormone and Prolactin Cells. AB - We report a case of giant pituitary adenoma in a child. Computerized tomography (CT) scan revealed a suprasellar extension tumor mass with hydrocephalus. There was no clinical evidence of acromegaly, gigantism, and other hormonal symptoms. Endocrinologic studies showed within normal value of serum growth hormone (GH: 4.2 ng/mL) and slightly increased levels of prolactin (PRL: 78 ng/mL) and other pituitary hormone values were within normal range. On suppression test by bromocryptin, both GH and PRL levels were reduced. Histopathological findings revealed that the tumor consisted of predominantly chromophobic and partly eosinophilic adenoma cells. Immunohistochemical staining detected GH and PRL in a small number of distinctly different adenoma cells, respectively. Nonradioactive in situ hybridization (ISH) also showed GH and PRL mRNA expression in identical immunopositive cells. Electron microscopy (EM) demonstrated adenoma cells with moderate or small numbers of two types of dense granules and without fibrous body which are characteristic of sparsely granulated GH-cell adenomas. The adenoma does not fit into any classification but may be an atypical acidophil cell line tumor showing focal differentiation toward both GH and PRL cells. PMID- 12114747 TI - Pheochromocytoma in von hippel-lindau disease: distinct histopathologic phenotype compared to pheochromocytoma in multiple endocrine neoplasia type 2. AB - Pheochromocytomas are rare neuroendocrine tumors that arise from chromaffin tissue. In a small subset of patients, pheochromocytomas occur as a manifestation of von Hippel- Lindau (VHL) disease. The histology of VHL-associated pheochromocytomas has not been reported in detail. In this article, we describe histopathologic features of 14 pheochromocytomas in eight patients with VHL disease and demonstrate that VHL-associated pheochromocytomas have a distinct histologic phenotype as compared with pheochromocytomas in patients with multiple endocrine neoplasia type 2 (MEN 2). VHL tumors are characterized by a thick vascular tumor capsule; myxoid and hyalinized stroma; round, small to medium tumor cells intermixed with small vessels; predominantly amphophilic and clear cytoplasm; absence of cytoplasmic hyaline globules; and lack of nuclear atypia or mitoses. In contrast to MEN 2, there is no extratumoral adrenomedullary hyperplasia in the VHL adrenal gland. Our findings of a distinct histologic phenotype of VHL pheochromocytoma may further help in subdividing patients who clinically present with multiple, bilateral pheochromocytomas. PMID- 12114748 TI - Localization of myosin XVA in endocrine tumors of gut and pancreas. AB - The myosin superfamily includes conventional and unconventional myosin proteins. Among unconventional myosins, myosin XVA has recently been characterized, and it has been suggested that it may be involved in cytoplasmic organelle movement, including secretory granules in pituitary cells and pituitary adenomas. In this study, we investigated the expression of myosin XVA protein and mRNA in normal endocrine cells and in a series of 53 endocrine tumors of the gut and pancreas. Myosin XVA was expressed in rare normal endocrine cells of the gut and in almost all pancreatic islet cells. In addition, myosin XVA was detected in several cells of all endocrine tumors investigated, and its expression was not related to malignancy, type, site, or functional status of tumors. These results indicate that myosin XVA protein and mRNA are widely distributed in endocrine cells of the gut and pancreas. Although the role of this protein in endocrine cells is unknown, previous studies suggest that it may have a role in secretory granule movement and/or hormone secretion. PMID- 12114749 TI - Autopsy findings in diabetic patients: a 27-yr clinicopathologic study with emphasi on opportunistic infections and cancers. AB - Diabetes mellitus has become a growing epidemic in the Asia-Pacific region. The aims of this study were to determine at autopsy the prevalence and characteristics of pathologic lesions in patients with diabetes mellitus. The 13,215 autopsy reports in our institution were examined for the diagnosis of diabetes mellitus. In patients with diabetes mellitus, the demographic data and the different pathologic lesions noted were analyzed. Diabetes mellitus was found in 820 patients (426 men and 394 women), comprising 6.2% of all autopsies. The two most common types of disease were cardiovascular diseases and infections, found in 69 and 53% of diabetic patients, respectively. Bacterial infection, in particular tuberculosis, was the most common type of infection noted. Localized and disseminated fungal infections were also common. In addition, urinary tract diseases were noted in 48%, hepatobiliary tract lesions in 42%, central nervous system disorders in 25%, and tumors in 29% of the diabetic patients. Malignant tumors were more often seen than benign tumors (18 vs 11% of patients, respectively). Many of the tumors were adenocarcinomas, and the most common neoplastic lesions were carcinomas of the lung, pancreas, liver, large intestine, stomach, and esophagus. Diabetic complications and associated diseases are common problems in this population. Adequate health care resources are needed for their prevention and treatment. PMID- 12114746 TI - RET/PTC rearrangement in thyroid tumors. AB - Rearrangement of the RET gene, also known as RET/PTC rearrangement, is the most common genetic alteration identified to date in thyroid papillary carcinomas. The prevalence of RET/PTC in papillary carcinomas shows significant geographic variation and is approx 35% in North America. RET/PTC is more common in tumors in children and young adults, and in papillary carcinomas associated with radiation exposure. There are at least 10 different types of RET/PTC, all resulting from the fusion of the tyrosine kinase domain of RET to the 5' portion of different genes. RET/PTC1 and RET/PTC3 are the most common types, accounting for >90% of all rearrangements. There is some evidence that different types of RET/PTC may be associated with distinct biologic properties of papillary carcinomas. RET/PTC1 tends to be more common in tumors with typical papillary growth and microcarcinomas and to have a more benign clinical course, whereas RET/PTC3 in some populations shows a strong correlation with the solid variant of papillary carcinoma and more aggressive tumor behavior. RET/PTC has recently been found in hyalinizing trabecular adenomas of the thyroid gland, providing molecular evidence in favor of this tumor to be a variant of papillary carcinoma. The occurrence of RET/PTC in Hashimoto thyroiditis and thyroid follicular adenomas and hyperplastic nodules reported in several studies has not been confirmed in other observations and remains controversial. PMID- 12114750 TI - Expression of E-cadherin, b-catenin, and Ki-67 in goblet cell carcinoids of the appendix: an immunohistochemical study with clinical correlation. AB - Goblet cell carcinoid (GCC) of the appendix is a rare entity, of which both the histogenesis and biologic behavior remain controversial, and prognostic tools and therapeutic strategies for this unusual tumor have yet to be defined. The aim of this study was to analyze expression of E-cadherin and b-catenin in GCCs of the appendix with long-term follow-up data as related to the expression of Ki-67 proliferation marker to provide a rationale for treatment guidelines. We analyzed the expression of E-cadherin, b-catenin, and Ki-67 in 11 GCCs of the appendix and control groups of typical carcinoids of the large intestine (n = 29), well to moderately differentiated adenocarcinomas of the colon (n = 10), poorly differentiated adenocarcinomas of the colon (n = 12), and normal appendiceal tissues (n = 10). There was no significant difference between the GCCs and normal appendiceal tissues regarding the expression of E-cadherin or b-catenin (p = 0.297 and 0.103, respectively). The percentage of positive GCC cells ranged between 0.52 and 10.35% (4.27 +/- 0.80), and only one case had a score >10%. Metastatic tumor spread and death were found in high MIB-1 labeling index (LI) cases of GCC (>3%). Our findings suggest that the behavior of the majority of GCCs might be indolent and different from adenocarcinomas because of the preserved expression of E-cadherin and b-catenin and relatively low MIB-1 LI. However, some of these tumors act aggressively and MIB-1 LI might be a good parameter to determine the therapeutic procedure. PMID- 12114751 TI - Mature cystic teratoma involving adrenal gland. AB - A mature cystic teratoma presented as an adrenal mass in a 57-yr-old woman. The tumor was found to be predominantly paraadrenal but focally interrupted the adrenal cortex so that an intraadrenal origin could not be ruled out. Similar lesions have been reported extremely rarely and should be considered in the differential diagnosis of hormonally silent adrenal tumors. The findings of rimlike calcification and fatty density on computed tomography may be helpful diagnostically. PMID- 12114752 TI - Simultaneous medullary and papillary microcarcinoma of thyroid in a patient with secondary hyperparathyroidism. AB - The simultaneous occurrence of two neoplasms of different cellular origin in one organ is a known but rare event. Such a situation occurs in the thyroid when medullary and follicular carcinoma with differentiation develops. Several cases of transitional tumors have been reported, but the simultaneous occurrence of a carcinoma with medullary and follicular differentiated carcinoma is rare. In all patients, tumors were suspected after local examination of the thyroid. We report on a patient in whom a papillary microcarcinoma was detected on the left side and a medullary carcinoma on the right side while the patient was undergoing surgery for secondary hyperparathyroidism. Both tumors were confirmed by immunohistology; regional lymph nodes were free of tumor. Although the simultaneous occurrence of papillary and medullary carcinoma may have been a simple coincidence, the patient's history offers room for further speculation; he had chronic replicative hepatitis C and a 13-yr history of immunosuppression following renal transplantation. Therefore, a common oncogenic stimulus may have been involved. In the final analysis, the reason for both malignancies could not be clearly established. PMID- 12114753 TI - Medullary thyroid carcinoma, follicular variant. AB - We report here a 48-yr-old woman presenting with a solitary thyroid nodule in the left lobe of the thyroid. The aspiration cytology of the nodule was reported as follicular neoplasia and she underwent surgery. Frozen section was suspicious for medullary thyroid carcinoma and a total thyroidectomy was performed. The pathology report revealed medullary thyroid carcinoma, follicular variant. Immunohistochemical analysis was negative for thyroglobulin and positive for calcitonin. A few patients with this variant have been reported in the literature, mainly diagnosed by immunohistochemical features of the tumor. In light of the limited information we have obtained from the literature, it is reasonable to emphasize that these cases should be distinguished from the mixed medullary- follicular thyroid carcinomas and medullary carcinomas with entrapped follicles. Immunohistochemical examination with calcitonin and thyroglobulin is also essential. PMID- 12114754 TI - Expression of Growth Factors in Normal and Neoplastic Pituitary Tissues. AB - Many growth factors are expressed in normal pituitary cells and pituitary tumors. They are involved in gene expression for pituitary hormones and in cell proliferation. Some appear to be important for prognosis or treatment. Strong overexpression of some growth factors may indicate a more rapid growth. The significance of the different growth factors for pituitary function and pathology is discussed. PMID- 12114755 TI - Evaluation of Endocrine Neoplasms Using Fine Needle Aspiration Biopsy. PMID- 12114756 TI - Localization of Hepatocyte Growth Factor and Its Receptor met in Endocrine Cells and Related Tumors of the Gut and Pancreas: An Immunohistochemical Study. AB - Hepatocyte growth factor (HGF), a stimulator of angiogenesis and cell migration, regulates the growth of a wide variety of cells by binding to its high-affinity receptor met and is involved in the growth and aggressiveness of several tumors. In this study we investigated the expression of HGF and met in normal endocrine cells and related neoplasms of the gut and pancreas to verify their possible role in tumor pathogenesis, growth, and aggressiveness. Normal tissues and 60 different endocrine tumors were immunostained using specific antibodies directed against HGF, met, and various hormones. HGF immunoreactivity (IR) was found in antroduodenal G cells, rectal enterochromaffin (EC) cells, and pancreatic A and B cells, whereas met IR was detected in antral EC and C cells, and in pancreatic B cells; 46 of 60 tumors examined were positive for HGF, and they were mainly represented by ECL-, EC-, and L-cell neoplasms. met IR was identified in 50/60 tumors of various phenotypes. HGF and met coexpression was found in 42/60 cases, most of which were represented by EC-cell tumors. HGF/met coexpression was significantly more frequent in ileocolonic EC-cell tumors, which in the majority of cases were malignant, than in appendiceal EC-cell tumors, which were all benign. Our results demonstrated, for the first time, that HGF and met are specifically distributed in normal gut and pancreatic endocrine cells and, in addition, suggest that HGF and met may be implicated as autocrine/paracrine factors regulating the growth of gastroenteropancreatic endocrine tumors, mainly of ileocolonic EC-cell carcinoids. PMID- 12114757 TI - Cell Cycle Regulatory Proteins p27(kip), Cyclins Dl and E and Proliferative Activity in Oncocytic (Hurthle Cell) Lesions of the Thyroid. AB - Cyclins are prime cell-cycle regulators central to the control of cell proliferation in eukaryotic cells. The formation of cyclin/cyclin-dependent kinases (CDK) complexes activates the kinases and initiates a cascade of events, which directs cells through the cell cycle. CDK inhibitors (CDKIs) such as p27(kip1) inhibit cyclln-CDK complexes and function as negative regulators of the cell cycle. Previous studies have shown that p27(kip1) is decreased In malignant relative to benign thyroid tumors, but its role and Interaction with other cell cycle regulatory proteins have not been well established In oncocytic lesions of the thyroid. We studied the expression of p27(kip1), cyclins D1 and E, and Ki67 In 20 cases of oncocytic adenoma (AD). 6 cases of oncocytic carcinoma (CA). 8 cases of Hashimoto's thyroiditis (HT). and 9 cases of nodular goiter with oncocytic change (NG) by Immunohistochemlstry. In the latter two lesions only oncocytic cells were evaluated. The positive staining was stratified Into four groups. Statistical analysis was done using the Kruslcal-Wallis one-way analysis of variance test, and, when significant the Dunn multiple-comparisons procedure was used to determine pairwise differences. AllI 20 AD were p27(kip1) posItive, 10 were 4+, 2 were 3+, and the remaining 8 were 1+. In contrast all 6 CA showed 4+ p27(kip1) staining, of the 8 HT 2 were 4+, two 3+, three1+, and I was negative.All 9 NG were p27 positive, 7 showed 4+, one 3+, and one 1+ staining. On pairwise comparison differences in p27(kip1) staining between AD and CA and between HT and CA were statistically significant (p=0.0243 and p=0.0142, respectively). In all but one case Ki67 expression was either very low (<3%) or negative. No significant differences were seen in the expression of cyclin D1 or cyclin E among the groups observed. In conclusion, the increased p27(kip1) expression in malignant oncocytlc tumors relative to benign oncocytic lesions is unlike any other malignant progression reported in the thyroid and other organ systems in the body. This may reflect on the biologic nature of the oncocytic cells of the thyroid and the significance of this finding remains to be established. Proliferative activity as studied by Ki67 immunostalning was not helpful in distinguishing benign from malignant oncocytic tumors. PMID- 12114758 TI - Effect of Dopamine Agonists on Lactotroph Adenomas of the Human Pituitary. AB - Dopamine (DA) agonists cause reduction of blood prolactin level and tumor shrinkage in most patients with lactotroph adenoma. Our aim was to investigate the cellular mechanism of tumor shrinkage by determining mitotic, MIB-1, p27, and apoptotic indices, as well as microvessel density (MVD), surface microvessel density (SMD). ploidy, and other nuclear parameters. Surgically removed lactotroph adenomas were selected from 29 patients, of whom 19 were treated with oral bromocriptine (BEC), long-acting injectable BEC (BEC-LAR), or quinagolide and 10 were untreated. In treated adenomas mitotic and MIB-1 indices were lower, whereas the apoptotic indices were not significantly higher compared to untreated adenomas. The decrease in MIB-1 labeling reached significance in adenomas exposed to quinagolide (p<0.05). Aside from the BEC-LAR treated group, wherein p27 expression was significantly reduced (p<0.05), p27 expression did not differ significantly between the treated and untreated groups. MVD density was significantly lower in the treated adenomas, whereas the decrease in SMD did not attain significance. The DNA ploidy and most other nuclear parameters did not differ significantly in the two groups. In conclusion, reduction of mitotic and MIB-1 indices indicates that suppression of cell proliferation contributes to tumor shrinkage, whereas p27 protein expression and apoptosis play no major role in the adenoma involution. Further studies are required to explain the effect of DA agonists on MVD and SMD. PMID- 12114759 TI - Combined "Mixed Medullary-Follicular" and "Papillary" Carcinoma of the Thyroid with Lymph Node Metastasis. AB - We report a case of combined "mixed medullary follicular" and "papIlary" carcinoma of the thyroid that occurred in a 44 yr-old Japanese woman. The grossly single 3 cm tumor was histologically composed of both mixed medullary follicular carcinoma and papillary carcinoma, which abutted against each other with a clear border between two components. Immunohistochemically, the component of medullary carcinoma was positive for calcitonin and carcinoembryonic antigen (CEA), and the follicular carcinoma and papillary carcinoma components were positive for thyroglobulin. Lymph node metastasis was also noted. The patient has been alive without recurrence for 20 yr. To the best of our knowledge, this is the first reported case in the literature. We report this unique case of thyroid carcinoma and review related thyroid malignancies. PMID- 12114760 TI - Cushing's Syndrome Caused by a Well-Differentiated Ileal Neuroendocrine Carcinoma. AB - We report a very rare case of Cushing's syndrome caused by an ileal adrenocorticotrophin-secreting well-differentiated endocrine carcinoma with liver metastases. A 62-yr-old woman presented with clinical and biological signs suggestive of paraneoplastic Cushing's syndrome Radiological investigations, including magnetic resonance imaging of the pituitary gland, chest and abdominal computerized tomography scan, smalI bowel barium study and pancreatic endoscopic scan, 111 -pentetreotide scintigraphy (octreoscan), esogastroduodenoscopy, and colonoscopy did not detect the source of the ectopic adrenocorticotropic hormone (ACTH) secretior but showed a few liver nodules. Because a 10 mo-long ketoconazole treatment was not effective, a bilateral adrenalectomy was performed in order to control the Cushing's syndrome, and a liver nodule biopsy diagnosed a metastatic neuroendocrine tumor. Subsequently, a second laparotomy disclosed a 1.8 cm ileal neuroendocrine tumor, cosecreting ACTH and serotonin on immunohistochemistry with many liver metastases. ACTH and secretion by intestinal neuroendocrine tumors is rare, and the secretion is a most always clinicailly silent. Cushing's syndrome caused by an intestinal low grade neuroendocrine tumor is even rarer, with only two previous cases reported in the literature. Our observation underlines the difficulty of localizing the source of ectopic ACTI-I secretion in paraneoplastic Cushing's syndrome. PMID- 12114761 TI - Gallbladder Adenocarcinoma with Florid Neuroendocrine Cell Nests and Extensive Paneth Cell Metaplasia. AB - We report a unique case of gallbladder adenocarcinoma associated with florid neuroendocrine cell nests and extensive Paneth cell metaplasia that has not been described previously. The patient was a 79-yr-old woman with a pedunculated, polypoid mass in the gallbladder. Microscopically, the mass was composed at tumor cells showing tubular and papillary growth patterns, consistent with well differentiated adenocarcinoma. One-third or more of the tumor cell showed Paneth cell appearance. Goblet cell-type tumors were also intermingled. In addition, neuroendocrine cell nests, that were connected to the neoplastic glands, were scattered throughout the stroma. lmmunohistochemically, the labeling index of MIB 1 in adenocarcinoma cells including Paneth cell-type carcinoma cells was approx 40%. Neuron-specific enolase, chromogranin A, and synaptophysin were positive in the neuroendocrine cells forming solid nests and intermingled within neoplastic glands. They were immunopositive for serotonin but negative for insulin, glucagon, somatostatin, and pancreatic polypeptide (PP). Although MIB-1-positive neuroendocrine cell nests were very few with weak staining, we think that the neuroendocrine cell nests were neoplastic in nature. The formation of the multifocal neuroendocrine nests may be a consequence of the trophic effects of unknown substance(s), which can promote serotonin producing neuroendocrine cells to proliferate. We postulate that Paneth cell-type carcinoma cells may be intimately related to such substance(s) in our case. PMID- 12114762 TI - Cystic Medullary Thyroid Carcinoma: Report of a Case with Morphological and Clinical Correlations. AB - Cystic lesions of the thyroid are common findings. Although many thyroid cysts are of benign, some cases of hemorrhagic degenerative changes occur in neoplastic nodules, mostly follicular neoplasms and papillary carcinomas. The occurrence of hemorrhagic changes in medullary carcinomas has never been documented with aspirative cytological and histological pictures to the best of our knowledge. A case of medullary thyroid carcinoma with a large central hemorrhagic cyst is described, and the literature regarding the pathogenesis of this regression and the occurrence of cystic neoplasms in the thyroid is reviewed. PMID- 12114763 TI - Human Insulinomas: Clinical, Cellular, and Molecular Aspects. AB - Insulinoma is the most frequently encountered functioning endocrine pancreatic tumor in humans. In this overview we summarize morphological and clinical features of insulinomas, report about the proinsulin-insulin conversion in normal and neoplastic B-cells, discuss the new classification, the criteria of malignancy, and the clonal composition of endocrine pancreatic tumors, and outline recent findings on the molecular pathology of these tumors. PMID- 12114765 TI - Neoplasms Causing Nonhyperinsulinemic Hypoglycemia. AB - Non-islet cell tumor hypoglycemia (NICTH) is uncommon. Many of the tumors associated with NICTH are mesenchymal tumors, although carcinomas are also involved in some cases. High serum levels of insulin-like growth factor II (IGF II) have been associated with NICTH. Analysis of 4 pituitary tumors, 2 adrenocortical tumors, 42 solitary fibrous tumors, and 23 other mesenchymal tumors for IGF-II protein and mRNA showed that most mesenchymal tumors expressed IGF-II protein and mRNA, although only 4 of 48 patients had associated hypoglycemia. Tumor size was related to IGF-II production. Tumors less than 5 cm were usually negative for IGF-II mRNA, whereas 92.3% of tumors greater than 9 cm were positive for IGF-II mRNA. These results show that IGF-II mRNA and protein can be readily detected in many tumors, even when the tumors are not associated with clinical hypoglycemia. The expression and production of this growth factor cannot accurately predict patients with clinical evidence of hypoglycemia. PMID- 12114764 TI - Origin and Genetic Background of Insulinomas. AB - Insulin-producing B cell tumors (insulinomas) are the most frequent functioning endocrine tumors of the pancreas. Available experimental evidence suggests that the islet B cell is the most likely cell of origin of insulinomas, while the duct endocrine cell should be considered if rearrangement of the pancreatic parenchyma occurs. Data on the genetic background of insulinomas suggest that the B cell tumor development may result from alteration of several genes, including the multiple endocrine neoplasia type 1 (MEN1) gene. PMID- 12114767 TI - Clinicopathologic and Prognostic Significance of the Expression of Mucins, Simple Mucin Antigens and Histo-Blood Group Antigens in Papillary Thyroid Carcinoma. AB - We have previously analyzed the expression of MUC1, underglycosylated MUC1, MUC2, MUC5AC, Tn, sialyl Tn, Lewis(a), sialyl Lewis(a), Lewis(x), and sialyl Lewis(x) in papillary thyroid carcinoma (PTC). The present study of 26 thoroughly scrutinized cases of PTC with "good" or "bad outcome" and a minimum of 10 yr of follow-up (or until death of the patients) was undertaken in an attempt to find if there is any relationship between the expression of the aforementioned antigens and the clinicopathologic and morphologic features of the cases and to evaluate the prognostic significance of the immunohistochemical pattern of PTC. We observed a significant or suggestive association between the expression of MUC1, underglycosylated MUC1, MUC2, Lewis(a), and Lewis(x) and the older age of the patients, and a suggestive association between Lewis(x) expression and lymph node metastasis and venous invasion. There was also a strong correlation between extrathyroid invasion, lymph node metastasis, intrathyroid dissemination, and venous invasion and patients outcome thus demonstrating, once more, the prominence of the classic clinicopathologic and morphologic features in the prognostic evaluation of PTC. The immunohistochemical study of mucins, simple mucin antigens, and histo-blood group antigens did not provide additional prognostic information, but for the significant association of Lewis(x) immunoreactivity to the group of "bad outcome." PMID- 12114766 TI - Focal and Diffuse Beta Cell Changes in Persistent Hyperinsulinemic Hypoglycemia of Infancy. AB - In recent years our understanding of the changes in the endocrine pancreas in persistent hyperinsulinemic hypoglycemia in infancy, a form of congenital hypoglycemia, has increased considerably, in terms of both morphological classification and molecular pathogenesis. This review summarizes the current state of knowledge about the pathological lesions in the pancreas and their relationship to recently reported molecular findings. PMID- 12114768 TI - Papillary Thyroid Carcinoma Overexpresses Fully and Underglycosylated Mucins Together with Native and Sialylated Simple Mucin Antigens and Histo-Blood Group Antigens. AB - We studied the immunohistochemical expression of mucins (MUC1, underglycosylated MUC1, MUC2, MUC5AC, and MUC6), simple mucin antigens (Tn, sialyl Tn, and T), and histo-blood group antigens (type 1-Lewis(a) and sialyl Lewis(a) type 2-Lewis(x) and sialyl Lewis(x)) in a series of 26 papillary thyroid carcinomas (PTC), 6 follicular carcinomas, and a control group of 32 cases of "normal" thyroid parenchyma adjacent to the tumors. PTC expressed more often, more intensively, and more extensively every antigen but MUC6, which was not observed in any case. The expression of MUC5AC was also extremely rare. MUC1 expression was related to the expression of underglycosylated MUC1, MUC2, Lewis(a), and sialyl Lewis(a). A trend toward an association between the expression of MUC1 and that of type 2 histo-blood group antigens was also observed. Whenever there was a dissociation between the expression of type 1 and type 2 Lewis antigens, MUC1 appeared closely related to type 1 and independent from type 2 histo-blood group antigens. We conclude that MUC1 plays a pivotal, though not exclusive, role in the glycosylation features of well differentiated thyroid carcinomas. Despite the prominent expression of mucins and carbohydrate antigens in PTC, no significant differences were observed between PTC and follicular carcinoma thus ruling out the possibility of using the aforementioned antigens as diagnostic markers per se. PMID- 12114769 TI - Superoxide Dismutase in Human Adrenal and its Disorders: A Correlation with Development and Neoplastic Changes. AB - In adrenal glands, oxidative free radicals are synthesized in the course of hormonal production, and enzyme superoxide dismutase (SOD) is considered to scavenge these harmful superoxide radicals and, subsequently, to protect the cells. We studied immunohistochemical localization of Mn (manganese)-SOD and Cu,Zn (copper-zinc)-SOD in human adrenal and its disorders from fetus to adult obtained from autopsy or surgery in order to examine the possible biological significance of these two enzymes. In fetal adrenal (n = 4), Cu,Zn-SOD and Mn-SOD were detected only in the fetal cortex. In adrenal glands from children (n = 21) to adults (n = 15), Mn-SOD immunoreactivity was exclusively detected in adrenal medulla, whereas Cu,Zn-SOD immunoreactivity was present only in adrenocortical parenchymal cells, weakly in the zona glomerulosa, and markedly in the zona reticularis. There were no differences in relative immunointensity and/or patterns of immunolocalization of these two SODs among different age groups. Both Cu,Zn-SOD and Mn-SOD immunoreactivity were detected in compact tumor cells of adrenocortical adenoma (n = 16). Marked immunoreactivity of both Cu,Zn-SOD and Mn SOD was detected in adrenocortical carcinoma (n = 11) and pheochromocytoma (n = 5). These results indicate that Cu,Zn-SOD and Mu-SOD may play different roles as a scavenger or antioxidants in normal human adrenal glands, i.e., Cu,Zn-SOD as a scavenger of toxic superoxide radicals generated during steroidogenesis and Mn SOD during catecholamine production. Cu,Zn-SOD and Mn-SOD immunoreactivities detected in adrenal neoplasms are also considered to represent altered expression of these enzymes associated with neoplastic transformation, as reported in other human malignancies. PMID- 12114770 TI - Mitotic Counts in Rat Adenohypophysial Thyrotrophs and Somatotrophs: Effects of Short-Term Thyroidectomy, Thyroxine, and Thyrotropin-Releasing Hormone. AB - The question of whether thyroxine (T(4)) and thyrotropin-releasing hormone (TRH) affect mitoses in pituitary thyrotrophs (Tt) and somatotrophs (St) of hypothyroid rats was investigated. Fifteen day thyroidectomized (Tx) rats were used. Groups of Tx animals received T(4) or TRH or both. Except 6 and 24 h TRH groups, the animals were sacrificed 12 h after injections. Unoperated euthyroid rats served as controls. In Tx group adenohypophysial mitoses were significantly increased. T(4) diminished mitoses in Tx rats. Mitotic counts were decreased in 6 and 24 h Tx groups, but increased in 12 h TRH group. TRH plus T(4) in Tx animals had a synergistic effect on adenohypophysial mitoses. In unoperated controls few mitoses were observed in Tt and more mitoses in St. In Tx rats more mitoses were seen in Tt than in St. T(4) alone failed to reduce mitoses in Tt but increased them in St. We concluded that T(4) affects Tt and St replication. In normal rats mitoses occur mainly in St. In Tx rats mitotic activity increased in Tt. TRH plus T(4) have a synergistic mitogenic effect on St. T(4) but not TRH affects St replication. It appears that the presence of T(4) is necessary for St multiplication. PMID- 12114771 TI - Composite Pheochromocytoma-Ganglioneuroma of the Adrenal Gland: An Uncommon Entity with Distinctive Clinicopathologic Features. AB - Background: Adrenal composite pheochromocytoma-ganglioneuroma is uncommon. The aim of this study is to investigate the characteristics of patients with this tumor. Methods: Histologic features of 46 pheochromocytomas diagnosed over a 28 year period (1971-1998) were reviewed. The clinical records, pathologic features, and ploidy patterns of patients with composite pheochromocytoma-ganglioneuroma were studied. Cases reported in the literature were also reviewed and compared with typical pheochromocytomas. Results: Four patients (two men; two women) with composite pheochromocytoma-ganglioneuromas were found. The tumors comprised 8.7% of pheochromocytomas. The two male patients with composite tumors were younger than the two female patients (age = 32, 52 vs 73, 75, respectively). The radiologic, gross, and microscopic appearances of the tumors were heterogeneous. One patient had malignant tumor with distant metastases. Flow cytometric analysis showed that the pheochromocytoma component was aneuploid and the ganglioneuroma portion was diploid. A review of the literature showed 31 adrenal composite pheochromocytoma-ganglioneuromas. Composite tumors were seen in older patients and were bigger than typical pheochromocytomas. Conclusion: Composite pheochromocytoma-ganglioneuromas had distinctive clinicopathologic features. Life long clinical and biochemical follow-up of these patients is essential in view of the non-diploid DNA pattern. PMID- 12114772 TI - Pigmented Granules in Functional Black Adenoma of the Adrenal Gland: A Histochemical and Ultrastructural Study. AB - A black adenoma of the adrenal gland was laparoscopically removed from a 61-year old Japanese female who had clinical and laboratory findings characteristic of Cushing's syndrome. The tumor consisted of polygonal cells that contained numerous brown pigmented granules of various sizes by routine hematoxylin-eosin staining. The histochemical study showed that the pigment had the characteristics of lipofuscin, not of melanin or neuromelanin. Electron microscopic study revealed tumor cells with two types of pigmented granules. These results show that there might be differences in the lipid metabolism of individual tumor cells in black adenomas of adrenal; which suggests an interesting histogenesis for these granules. PMID- 12114773 TI - Plurihormonal Bronchial Carcinoid Associated with Ectopic Cushing's Syndrome. AB - A 31-year-old woman presented with progressive weight gain, facial acne, round facies, hirsutism, and secondary amenorrhea. Her plasma cortisol, urinary free cortisol, and plasma ACTH were elevated. CT scan of abdomen revealed bilateral diffuse adrenal enlargement. MRI of pituitary failed to identify a lesion. CT scan of chest revealed an 8 mm nodule in the lower lobe of the left lung. The diagnosis of ectopic Cushing's syndrome was made. The lung tumor was surgically removed. The tumor measured up to 1.5 cm in diameter. By light microscopy, a tumor with characteristic features of bronchial carcinoid was noted. Immunostains were positive for neuron-specific enolase, synaptophysin, chromogranin, low molecular-weight keratin, ACTH, beta endorphin, corticotropin-releasing hormone, bombesin, alpha subunit, and somatostatin. Electron microscopy revealed an endocrine neoplasm. The secretory granules displayed great variation subdividing the endocrine neoplasm. The secretory granules displayed great variation subdividing the cell population into an undetermined number of phenotypes. In situ hybridization demonstrated positivity for pro-opiomelanocortin mRNA in the tumor cells. Postoperatively, plasma cortisol had fallen. The patient remained symptom-free one year later. The case presented here was regarded as a plurihormonal bronchial carcinoid associated with ectopic Cushing's syndrome. This unique plurihormonal bronchial carcinoid tumor produced three hormones, ACTH, CRH, and bombesin, that may have contributed to the patient's ectopic Cushing's syndrome. PMID- 12114774 TI - GH and PRL-Producing Pituitary Adenoma with Neuron-Like Differentiation. AB - A pituitary adenoma with neuron-like differentiation in the sella turcica is reported. Sections of the tumor showed a mixture of adenoma cells, ganglionic cells, and neuropil-like structures by light microscopy. Both pituitary adenoma cells and large cells recognized as ganglionic cells by H&E were strongly immunoreactive for both growth hormone (GH) and prolactin (PRL), which indicated that these large cells had properties similar to those of pituitary adenoma cells. Furthermore, electron microscopy (EM) revealed characteristic low electron dense secretory granules as well as GH-type large electron-dense secretory granules in adenoma cells, neuropils, and swollen bulbs of neuronal endings, which indicated that these three populations may be of the same origin. Furthermore, we could not find typical cell bodies of ganglionic cells by EM. These results are consistent with a hypothesis that attempts to explain the origin of the neuronal components by the neuronal differentiation of adenoma cells. Thus, the best designation of our tumor may be "pituitary adenoma with neuron-like differentiation." PMID- 12114775 TI - Clonality of Endocrine Proliferative Lesions: A Critical Reappraisal. AB - The distinction between nodular hyperplasia and benign tumors of the endocrine system is problematical. Although numerous parameters including lesional size, multicentricity and histological features have been suggested as distinguishing criteria, none of these is absolute. Analyses based on X-chromosome inactivation have provided conflicting results with respect to the clonal origins of these lesions, and at least some lesions conventionally classified as hyperplastic nodules appear to be monoclonal, Although clonality has been generally equated with neoplasia, it is likely that clonal expansion of genetically normal cells can occur in the endocrine system as a result of a variety of growth of promoting stimuli. Multiparametric studies employing markers for X-chromosome inactivation together with methods for identification of unique non-X-linked genetic alterations will be required to resolve the many questions relating to the pathogenesis of endocrine proliferative lesions. PMID- 12114776 TI - Clonality in Thyroid Nodules: The Hyperplasia-Neoplasia Sequence. AB - The evaluation of clonality in nodular proliferations of the thyroid gland has identified a large proportion of monoclonal nodules even in the setting of nonendemic nodular goiter The literature on this topic is reviewed and an attempt is made to explain the available data. In the absence of a clear understanding of the biological impact and potential of these molecular events, a recommendation is made for continued use of current histomorphologic nomenclature when diagnosing such thyroid lesions in the day-to-day practice of surgical pathology. PMID- 12114777 TI - Clonality Analysis of Benign Parathyroid Lesions by Human Androgen Receptor (HUMARA) Gene Assay. AB - Benign conditions of the parathyroid gland have been classified as adenomas and hyperplasias. These entities however are difficult to distinguish when only a single gland is enlarged. Adenomas are defined as neoplastic clonal growths whereas hyperplasias are considered to be reactive processes of polyclonal origin. In order to analyze the clonal pattern of these lesions, we have studied hyperplasias and adenomas of parathyroid glands from women by the human androgen receptor (HUMARA) assay, a recently reliable and highly-lnformative technique based on the X-chromosome inactivation pattern in females. Samples consisted of formalin-fixed as well as frozen tissues. Informativeness with HUMARA marker was 87% (13/15 cases). All hyperplasias (5/5) and 6/8 adenomas yielded polyclonal results, since two alleles of similar intensity appeared when the lesion was HpaIl-digested. Two parathyroid adenomas had a loss of one X-alIeIe for the HUMARA gene and they were interpreted as monoclonal. These results show that parathyroid hyperplasias and adenomas, considered as multigland or monogland involvement diseases respectively, may be both polyclonal in origin, and that only a small subset of adenomas is found to be clonal. Consequently, clonality analysis cannot allow a clear distinction between these two entities as classically diagnosed. A different approach should be considering hyperplasia or adenoma when a polyclonal or monoclonal result has been obtained by clonality analysis. PMID- 12114779 TI - Fine-Needle Aspiration Cytology of Papillary Hurthle Cell Carcinoma with Lymphocytic Stroma "Warthin-Like Tumor" of the Thyroid. AB - Papillary Hurthle cell carcinoma with lymphocytic stroma is a recent addition to the list of variants of papillary carcinoma of the thyroid. We report the aspiration cytology and histology findings of this tumor arising in two patients. The smears were cellular, and revealed Hurthle cells arranged in three dimensional groups, papillary fragments, and as singly dispersed cells with a prominent intimately associated inflammatory component of lymphocytes and few plasma cells. The Hurthle cells were pleomorphic and showed granular eosinophilic cytoplasm, eccentrically oriented nuclei with prominent nucleoli. Nuclear features of papillary carcinoma were present among both the cellular groups and scattered cells. The histologic examination showed a circumscribed papillary tumor comprising Hurthle cells and a brisk inflammatory component filling the stalks of papillae, These findings were consistent with a papillary Hurthle cells carcinoma with lymphocytic stroma, the so-called Warthin-like tumor of the thyroid. Hurthle cells admixed with inflammatory cells in cytology preparations can be seen in Hurthle cell nodules or neoplasms arising in a background of chronic lymphocytic thyroiditis. We suggest that a careful search for nuclear features may be helpful in diagnosing this variant of papillary carcinoma. PMID- 12114778 TI - Clonality Studies in the Analysis of Adrenal Medullary Proliferations: Application Principles and Limitations. AB - Clonality remains as the hallmark of neoplasms. A dual genetic approach using markers nonrelated (e.g., X-chromosome inactivation assays) and related to the malignant transformation (such as loss of heterozygosity analyses of tumor suppressor genes) would provide useful clonality information from early and advanced tumor stages, respectively. Tumor progression and clonal selection would result in genetic instability and heterogeneous expression of those molecular markers related to the malignant pathway. Therefore, only the coexistence of multiple genetic abnormalities would support the clonal nature as an expression of convergent cell selection. Considering those facts, the currently available evidence on tumorigenesis and clonality in the adrenal medulla can be summarized as follows: 1. Multistep tumorigenesis defines the evolution of pheochromocytomas, as evidenced by the presence of several genetic alterations. 2. Both the significant association of nonrandom genetic alterations (specially 1p and 22q interstitial deletions) and the topographic accumulation of genetic deletions at the peripheral tumor compartment support a convergent clone selection for these neoplasms. 3. Although many genetic loci show nonrandom abnormalities, the most frequently involved locates on chromosome 1p regardless of genetic tumor background (sporadic or inherited predisposition). 4. Most pheochromocytomas should begin as monoclonal proliferations that do not always correlate with histopathologic features, particularly in inherited tumor syndromes. 5. Early histopathologic stages, described as adrenal medullary hyperplasias, are defined by hyperproliferative features in animal models and monoclonal patterns in the adrenal nodules from patients with MEN-2a. PMID- 12114780 TI - The ret-Activating Ligand GDNF Is Differentiative and Not Mitogenic for Normal and Neoplastic Human Chromaffin Cells In Vitro. AB - Activating mutations of the receptor tyrosine kinase, ret, are associated with multiple endocrine neoplasia type 2A (MEN 2A). However, the mechanisms leading to tumor development are unclear. Glial-derived neurotrophic factor (GDNF) activates wild-type ret via interaction with a second receptor, GFR a-l. We have utilized GDNF to stimulate normal and neoplastic chromaffin cells in order to ask whether ret activation is mitogenic. Cells from three normal adult adrenal medullas, one sporadic pheochromocytoma, and three MEN-2A pheochromocytomas were labeled with bromodeoxyuridine (BrdU) for 12 d in the presence or absence of GDNF or nerve growth factor (NGF), which is known to stimulate neurite outgrowth, but not proliferation in human chromaffin and pheochromocytoma cell cultures. Responses to GDNF and NGF were comparable, except for two MEN-2A pheochromocytomas that responded minimally to GDNF and robustly to NGF. These tumors responded to GDNF biochemically, as measured by phosphorylation of mitogen-activated protein kineses, despite their weak morphological responses. Our findings suggest that activation of ret may not be sufficient to produce chromaffin cell hyperplasia or neoplasia directly by stimulating cell proliferation. However the possibility that altered cell-cell or cell-substrate interactions might cause responses to become differ entiative rather than proliferative in vitro has not been ruled out. We also demonstrate, for the first time, that at least some human pheochromocytomas with an MEN-2A ret mutation respond to a normal ret ligand. This responsiveness could be mediated by a remaining normal ret allele or by other mechanisms. PMID- 12114781 TI - Thyroid Peroxidase and Thyroglobulin Expression in Normal Human Thyroid Glands. AB - The objective of the present study was to investigate thyroid peroxidase (TPO) and thyroglobulin (Tg) expression in normal thyroid tissue. Thirty-eight normal thyroids from fetuses(1) children, adults, and elderly subjects were evaluated. TPO and Tg expression was determined by analyzing at least 200 cells/specimen. A value of 80% was considered to be the threshold for TPO positivity. In terms of age, TPO expression occurs in fetuses along with follicle differentiation and is absent in solid follicles. TPO staining was cytoplasmic, and was more intense in the apical membrane in tissue from fetuses, children, and young adults. A perinuclear rang was found in tissue from older adults and from the elderly subjects. There was concomitant TPO and Tg expression in all groups studied. Tg expression was observed in all normal thyroids from all age groups. Tg expression was extremely variable (10-100% of the cells) and fainter than TPO expression. PMID- 12114782 TI - Percoll Density Gradient-Enriched Populations of Rat Pituitary Cells: Interleukin 6 Secretion, Proliferative Activity, and Nitric Oxide Synthase Expression. AB - We used a discontinuous Percoll density gradient centrifugation to prepare enriched populations of prolactin (PRL), growth hormone (GH), and folliculo stellate (FS) cells from rat anterior pituitaries in order to characterize these various cell populations. After cell dissociation and centrifugation, enriched PRL cells (55% of total cells as determined by immunocytochemistry [ICC] were present in Fraction 1 (Fr1) (density ([d]) = 1.059). Fr2 (d=1.071) had enriched S100-positive FS cells (31% of total cells), but enriched GH cell (60% of total cells) were present in Fr3 (d=1.094). Interleukin 6 (IL-6) was secreted mainly by enriched PRL cells in Fr1 (350 pg/mL/106 cells) and Fr2 (194 pg/mL/106 cells), and much less by the enriched GH cells inFr3 (16 pg/mL/106). Proliferation studies with combined 3H-thymidine and ICC for pituitary hormones showed that only the PRL cell had significant prolifereative activity. Immunostaining showed that immediately after separation, all three isoforms of nitric oxide synthase (NOS) were expressed anterior pituitary cells. After 3 d of culture, there was a marked increase in nuclear staining for neuronal NOS (nNOS) in all three fractions, whereas inducible NOS (iNOS) and endothelial NOS (eNOS) rexpression did not change significantly. These results indicate that: 1. Enriched populations of PRL, FS, and GH pituitary cells can be readily obtained with a rapid discontinuous percoll density separation procedure. 2. PRL cells from different fractions of the gradient show differenet proliferation rates and IL-6 secretion varied in different enriched cell populations. 3. Although all three isoforms of NOS were expressed in rat pituitary cells, nNOS is the prindipal isoform in anterior pituitary cells, and its expression was icreased after 3 d of culture of anterior pitutuitary cells. PMID- 12114783 TI - Clear-Cell Follicular Adenoma of Ectopic Thyroid in the Submandibular Region. AB - A case of clear-cell follicular adenoma arising in ectopic thyroid tissue is reported. The 2.0-cm tumor arose in the submandibular region in a 29-yr-old female. The diagnosis was established on the basis of light microscopic morphology, a positive thyroglobulin immunohistology, and the presence of normal thyroid tissue surrounding the mass. Preoperative computed tomography (CT) scan and postoperative ultrasound studies revealed a normal orthotopic thyroid gland. No additional tumors have since been detected. The patient is free of recurrent or metastatic disease 54 mo following excision of the mass. Only eight previously published reports have described ectopic thyroid tissue in the submandibular region, all but one of which lacked an orthotopic thyroid gland. In this article, we describe the pathological features of our case and review the existing literature on the subject. PMID- 12114784 TI - Adrenal Pseudocysts: Evidence of Their Posthemorrhagic Nature. AB - Hemorrhagic adrenal pseudocysts are uncommon nonneoplastic lesions that have been reported as secondary to intraparenchymal hemorrhage or alternatively related to endothelial (vascular) cysts. Ultrastructural and ammunohistochemical evidence in support of the latter has been presented, but the exact nature of hemorrhagic adrenal pseudocysts remains poorly defined. We evaluated six surgical specimens of hemorrhagic adrenal pseudocysts using immunohistochemical staining for CD31 and CD34, as well as conventional histochemistry. All six cases had hemorrhagic contents within a wall of variable thickness possessing focal areas of linear, disrupted elastin, and smooth muscle. Three cases demonstrated extensive thrombosis with organization, including papillary endothelial hyperplasia, simulating angiosarcoma. In these cases, CD3I and CD34 staining decorated areas of papillary endothelial hyperplasia as well as foci of the internal cyst lining, whereas the other cases were negative for both antibodies. Of interest is the history of FNA prior to surgical resection in three cases of hemorrhagic adrenal pseudocysts, two of which showed papillary endothelial hyperplasia. The presence of papillary endothelial hyperplasia and our immunohistochemical findings support the conclusion that adrenal pseudocysts are posthemorrhagic and derive from vascular disruption. Furthermore, FNA or other interventional studies may be associated with papillary endothelial hyperplasia in hemorrhagic adrenal pseudocysts. PMID- 12114785 TI - Composite Paraganglioma-Ganglioneuroma of the Urinary Bladder: A Clinicopathologic, Immunohistochemical, and UItrastructural Study of a Case and Review of the Literature. AB - Urinary bladder paragangliomas are rare. An 81-yr-old woman was admitted because of whole-stream painless hematuria. Investigations revealed a pedunculated bladder tumor Pathological examination showed a biphasic tumor, composite paraganglioma-ganglio neuroma. The two separate components were documented by both immunohistochemical and ultrastructural studies. A review of the English literature on urinary bladder paragangliomas showed that the present case is the first case with this unique feature documented in detail, and the patient is the oldest one being reported. PMID- 12114786 TI - Thirty-Year Survival with an Untreated Malignant Duodenal Somatostatinoma. AB - Gastrointestinal somatostatinomas are rare. Only 56 cases with duodenal origin have been reported in the literature. The long-term course of an untreated tumor like this is unknown. We report a case of a male patient who lived 30 yr with an untreated metastatic duodenal somatostatinoma. This case suggests that a duodenal somatostatinoma may be of low malignant potential and that expectancy when treating this tumor might be indicated. PMID- 12114787 TI - Angiogenesis in Endocrine Neoplasms. AB - Angiogenesis promotes the growth of tumors, because it both facilitates oxygenation and nutrient flow, and removes metabolic waste. During the past two decades, as the importance of tumor vascularity became recognized, angiogenesis and the relationship between blood vessels and tumor progression received increasing attention. It was found that isolated tumor tissues failed to expand beyond few millimeters unless vascularized, whereupon vascularization they exhibited a rapid growth. Extensive research focusing on the relationship between tumor proliferation and the formation of new vessels has initially been undertaken to assess the role of angiogenesis in the progression of breast carcinomas. Significant results emerged from these investigations, and similar studies were extended to other tumor types, such as melanomas, cervical and pulmonary carcinomas, and so on. It is of note that angiogenesis as it relates to endocrine tumors has so far been limited to pituitary neoplasms, thyroid carcinomas, and pheochromocytomas. The purpose of the article is to provide a brief review of angiogenesis and to summarize available data regarding its role in the growth off endocrine neoplasms. PMID- 12114788 TI - Insulin-Dependent Diabetes Mellitus: Islet Changes in Relation to Etiology and Pathogenesis. AB - Type and type II diabetes are the most common types of diabetes. The ratio of type I to type II diabetes is about 1:9. Type I diabetes is caused by absolute insulin deficiency and is therefore referred to as insulin-dependent diabetes. The disease becomes manifest clinically in childhood or adolescence ("juvenile diabetes"), though onset in adulthood is increasingly being observed. Morphologically a subtotal (>80%) to total loss of B-cells in the pancreatic islets occurs. Lymphocytic insulitis, which disappears after the B-cells have been totally destroyed, is pathogneumonic of type I diabetes. This insulitis is an expression of an autoimmune event that is triggered by a multitude of factors. An important factor appears to be a genetic predisposition (human leukocyte antigen [HLA] DR3/DR4/DQ8) in connection with as-yet-unknown environmental factors (e.g., viruses). Autoantibodies, such as islet cell cytoplasmic antibodies (ICA). insulin autoantibodies (IAA) and/or autoantibodies to the gamma aminobutyric acid (GABA)-synthesizing enzyme glutamic acid carboxylase (GAD), are already detectable in a prediabetic phase, though it is not possible to predict the time of clinical onset. The course of the disease is dependent on age. Young children require insulin therapy sooner than juveniles or adults. PMID- 12114789 TI - Dyshormonogenetic Goiter: A Clinicopathologic Study of 56 Cases. AB - Dyshormonogenetic goiters (DG) are genetically determined thyroid hyperplasias due to enzyme defects in thyroid-hormone synthesis. We report 56 cases of DG occurring in 34 females and 22 males. The patients age ranged from newborn to 52 yr (median 16), 75% of the cases occurring before the age of 24. All patients presented with clinically evidence of goiter except for two patients that were diagnosed at autopsy. Hypothyroidism was documented before the histological diagnosis was made in 36 patients (64%). The thyroid gland was enlarged and multinodular in all cases, weighing up to 600g. Microscopically, the most common alteration consisted of markedly cellular nodules exhibiting a variety of architectural appearances, the solid and/or microfollicular patterns predominating. Papillary proliferations and an insular growth pattern were also present. Fibrosis was a common finding; in some instances it was very conspicuous, resulting in irregularities at the edge of the nodules simulating capsular invasion. Other constant features included marked nuclear atypia and minimal amount of colloid. In 18% of the cases, the degree of hyperplasia and atypia were such as to result in a mistaken diagnosis of follicular, papillary, medullary, or undifferentiated carcinoma. Three of the glands contained incidental small neoplasms fulfilling the criteria of papillary microcarcinoma; one of them was multicentric. The presence in a thyroid gland of the combination of these morphologic features should suggest the diagnosis of dyshormonogenetic goiter. The only other condition we are aware of that can result in a similar microscopic picture is iatrogenic goiter resulting from the administration of antithyroidal agents. PMID- 12114790 TI - TGFB, TGFB Receptors, Ki-67, and p27(Kip)l Expression in Papillary Thyroid Carcinomas. AB - Although most papillary thyroid carcinomas behave as low-grade neoplasms and are generally associated with a good prognosis, some subgroups of these neoplasms represent more aggressive variants. In order to determine if differences in the behavior of these papillary carcinomas were related to expression of growth factors or cell-cycle proteins, we analyzed a series of papillary carcinomas including the conventional or usual type (n = 27), tall cell (n = 27), diffuse sclerosing (n = 5), and columnar cell (n = 2) variants for expression of transforming growth factor beta (TGB), TGB receptors (TGB-RI and II, the proliferation marker Ki-67, and for the cell-cycle inhibitory protein p27(Kip)1 (p27). All groups of thyroid tumors expressed TGFB and TGFB-RI and RlI by immunohistochemical staining. There was a marked increase in the Ki-67 labeling index after staining with antibody MIB-1 in the columnar cell tumors compared to the other groups, but this difference was not significant because of the small number of tumors in this group. The cell-cycle inhibitory protein p27 was expressed in all groups and was not significantly different between groups. Normal thyroid cells had a higher labeling index for p27 compared to papillary carcinomas. These results indicate that TGFB and TGFB receptors I and II are commonly expressed in the usual and in variant forms of papillary thyroid carcinomas, and that there is decreased expression of p27 protein in all of these neoplasms compared to normal thyroid. The biological basis for the more aggressive behavior of these variants of papillary thyroid carcinoma remains uncertain. PMID- 12114791 TI - Carbohydrate Antigens as Oncofetal Antigens in Papillary Carcinoma of the Thyroid Gland. AB - The expression of simple mucin-type antigens, Lewis type-1 antigens, and Lewis type-2-related antigens was evaluated by immunohistochemistry in a series of nine fetal thyroid glands. The aim of the study was to establish whether the previously reported expression of carbohydrate antigens in thyroid carcinomas represents a "de novo" expression or a form of the so-called oncofetal expression. We have detected simple mucin-type antigens and Lewis type-2-related antigens in fetal thyroid tissue. Lewis type-I antigens were not found. Taking the results of this study together with those previously reported, we conclude that the presence of Lewis type I antigens in thyroid carcinomas appears to be a "de novo" expression, whereas the constant and strong expression of simple mucin type antigens and Lewis type-2-related antigens represents a re-expression (oncofetal expression) of antigens already present during fetal life. PMID- 12114792 TI - Matrix Metalloproteinases and Tissue Inhibitors of Metalloproteinases in Pituitary Adenomas: Possible Markers of Neuroendocrine Cells. AB - Matrix metalloproteinases (MMPs) are involved in remodeling processes and have been immunocytochemically localized in some endocrine glands and their tumors. Using anterior pituitary gland and pituitary adenomas, immunocytochemical localization of MMP-2 (gelatinase-A), -9 (gelatinase-B), tissue inhibitor of metalloproteinase (TIMP)-1 and -2 was performed. Normal anterior-pituitary cells all contain MMPs and lesser amount of TIMPs, whereas far fewer MMPs and TIMPs are identified in anterior pituitary adenomas. There is no correlation between pituitary hormone and MMPs-TIMPs localization, thus MMP-TIMP homeostasis may not be involved in hormone synthesis and secretion of anterior pituitary cells and their adenomas, Because MMPs and TIMPs are more abundantly and specifically localized in pituitary cells and their adenomas, MMPs and TIMPs may be included as markers for endocrine cells, including anterior-pituitary cells. PMID- 12114794 TI - Metastatic Psammomatous Somatostatinoma of the Pancreas Causing Severe Ketoacidotic Diabetes Cured by Surgery. AB - A case of somatostatin-producing pancreatic tumor associated with severe insulin dependent diabetes mellitus and ketoacidotic coma is reported. The tumor, a 10-cm expansile mass arising from the pancreatic tail of a 70-yr-old woman, was first detected by ultrasonography, performed because of abdominal pain, and subsequently confirmed by computed tomography and fine-needle tumor aspiration. Pathologic investigation showed a predominantly solid-trabecular structure with scattered microacini and psammomatous bodies. A large proportion of tumor cells expressed somatostatin and/or calcitonin. Following resection of the primary tumor and three peripancreatic lymph nodes with metastases, the patient recovered rapidly from her diabetic syndrome and remained in substantially good health during a subsequent 8-yr follow-up period, without evidence of tumor recurrence. PMID- 12114793 TI - Effects of Propyithiouracil (PTU) Administration on the Synthesis and Secretion of Thyroglobulin in the Rat Thyroid Gland: A Quantitative Immuno-electron Microscopic Study Using Immunogold Technique. AB - To clarify the effects of an antithyroid drug on the kinetics of thyroglobulin synthesis, secretion, and reabsorption in the thyroid follicles, propylthiouracil (PTU) was administered to rats and the thyroid glands were examined by a refined post-embedding immunogold technique during and after withdrawal of PTU. Seven-wk old male Wistar rats were administered with S mg of PTU/d through a gastric tube, and sacrificed at 1 and 2 wk of administration and at 1, 2, and 3 d, and 1, 2, 3, and 4 wk, after discontinuation. The administration of PTU caused a remarkable dilatation of the rER and Golgi apparatus, but these areas gradually recovered after withdrawal of PTU. During the experiment, no significant change in the density of thyroglobulin (Tg) was observed except for a transient increase immediately after withdrawal of PTU. The expression of Tg on subapical vesicles (SV) and follicular colloid took a relatively parallel course; increasing during administration of PTU and decreasing with a transient peak immediately after treatment was discontinued. In contrast to the remarkable changes in the morphology of compartments involved in Tg synthesis, the development of colloid droplets and formation of secondary lysosomes were suppressed during and after discontinuing administration of PTU. However, the basic pattern of the gradient of Tg density among the cellular compartments was essentially retained in the experimental group. Thus the present immunoelectron-microscopic study provided evidence that administration of PTU stimulates the synthesis and secretion of Tg in the follicular epithelium in vivo, and, also, suppresses reabsorption and degradation of Tg. Further, it was speculated that the density gradient of Tg among the compartments involved in Tg synthesis, secretion and storage is regulated by an unknown constitutive mechanism and not by the thyroid-stimulating hormone (TSH)-TSH receptor-mediated system. PMID- 12114795 TI - Primary Lymphoma of Pituitary Gland: A Neoplasm of Acquired Malt? AB - A 66-yr-old man presented with presyncopal episodes, dizziness, anorexia, nausea, and weight loss and was noted to have low blood pressure with a postural drop and sparse eyebrows. Laboratory investigations revealed evidence of hypopituitarism. Magnetic resonance imaging (MRI) revealed a non-enhancing mass arising from the adenohypophysis. The neurohypophysis was displaced laterally but appeared otherwise normal. The lesion was thought to be a nonfunctioning pituitary adenoma and a trans-sphenoidal hypophysectomy was performed. Histologically, this lesion was a diffuse large B-cell lymphoma that had features of a high grade mucosa associated lymphoid tissue (MALT)-type lymphoma arising in association with low grade B-cell lymphoma of MALT type. There was no clinical or radiological evidence of lymphoma in other sites and there was no evidence of an immunocompromised state. Only one previous case of primary malignant lymphoma of the pituitary has been reported and this patient presented with compression of the optic chiasm. We describe the clinical and pathological features of a patient who presented with hypopituitarism and was found to have a pituitary lymphoma. This is the first reported case of a pituitary lymphoma presenting with pituitary failure and the first case characterized by lymphocyte-marker studies that confirmed it to be a B-cell lymphoma. PMID- 12114796 TI - Thyroid Carcinoma in Japan and the West: Similarities and Differences. AB - Geographic or ethnic differences in the incidence of thyroid carcinoma, as well as in the histologic distribution of thyroid carcinoma between Japan and Western countries, have been described but are still unclear. The recent establishment of histologic criteria for the diagnosis of thyroid carcinoma by the WHO committee has facilitated the comparison of clinicopathological data of patients with thyroid carcinoma all over the world. The aim of the present review article is to clarify the epidemiological and clinicopathological differences of thyroid carcinoma between Japan and Western countries. We found recently no significant differences in the incidence, mortality, and histologic distribution of thyroid carcinoma between Japan and Western countries; this was contrary to our expectation. This is likely attributable to westernization of the Japanese diet, standardized medical levels, and international standardization of histologic criteria of thyroid carcinoma. PMID- 12114797 TI - Adrenal Medullary Nodules in Beckwith-Wiedemann Syndrome Resemble Extra-Adrenal Paraganglia. AB - Clues to mechanisms regulating development and tumorigenesis may be provided by studies of unusual diseases. Beckwit-Wiedemann syndrome (BWS) is a rare congenital disorder apparently related to abnormal regulation of insulin-like growth factor-2 (IGF-2) production. IGF2 mRNA has been previously localized to the chief cells of extra-adrenal paraganglia and to adult, but not fetal, adrenal medulla. Expression of IGF-2 by neuroblastomas has been hypothesized to reflect extra-adrenal paraganglionic differentiation. In the adrenals of a fetus with 8W5, we have observed both increased numbers of chromaffin cells and organoid nodules resembling extra-adrenal paraganglia. Immunoreactive IGF-2 was observed in both cell types, but was also observed in chromaffin cells in the normal fetal adrenal. The findings suggest autocrine or paracrine influences of IGF-2 in regulating the number and phenotype of cells derived from sympathoadrenal precursors in the developing adrenal medulla as well as in extra-adrenal paraganglia. These results have implications for the interpretation of data from neuroblastoma studies. PMID- 12114798 TI - Midgut Carcinoids and Solid Carcinomas of the Intestine: Differences in Endocrine Markers and p53 Mutations. AB - Fifteen midgut carcinoid tumors and 5 solid carcinomas of the intestine with carcinoid-like morphological features were evaluated histochemically and immunohistochemically with respect to various endocrine markers and expression of mutant p53 protein. Direct sequencing of the p53 gene after PCR amplification was carried out on microdissected cells from all tumors. All investigated carcinoid tumors showed chromogranin and argentaffin reaction, but lacked nuclear immunostaining with p53 antibodies. In 14 immunohistochemically negative midgut carcinoid tumors, no mutations were identified. One carcinoid tumor devoid of p53 staining was, however, found to contain mutation in exon 6 of the p53 gene. In contrast, the solid carcinomas were essentially chromogranin negative but all displayed clearly positive p53 staining in a variable number of cell nuclei. Sequence analysis of exons 5-8 of the p53 gene in the 5 carcinomas showed mutations in exons 6 and 7 in 2 tumors and in exon 8 in 2 other tumors, while no mutation was detected in the fifth tumor. The carcinoid tumors and the solid carcinomas of the small intestine are thought to derive histogenetically from endocrine cells and enterocytes, respectively. The present results substantiate that divergent mechanisms operate in the development of the two tumor types. PMID- 12114799 TI - Tissue Localization of Nerve Growth Factor Receptors: trk A and Low-Affinity Nerve Growth Factor Receptor in Neuroblastoma, Pheochromocytoma, and Retinoblastoma. AB - The immunohistochemical localization of nerve growth factor receptor (NGFR)-high affinity NGFR (trk A) and low-affinity NGFR (LNGFR)-was investigated in 23 neuroblastoma group tumors, 18 pheochromocytomas, 2 mixed neuroendocrine-neural tumors, and 16 retinoblastomas. trk A was expressed in the tumor cells of all neuroblastomas, pheochromocytomas, and retinoblastomas. Immunoreactive intensity was especially strong in the larger ganglionic tumor cells of ganglioneuroblastoma and ganglioneuroma. Messenger RNA (mRNA) of trk A was also strongly expressed in the ganglionic cells of ganglioneuroblastomas and chromaffin cells of pheochromocytomas by in situ hybridization method. LNGFR was negative in the tumor cells of neuroblastoma; however, it showed strong immunoreactivity in ganglionic tumor cells and Schwann cells of ganglioneuroblastoma/ganglioneuroma, and sustentacular cells of pheochromocytoma. Although normal retina expressed both trk A and LNG FR, tumor cells of retinoblastoma were positive for only trk A but negative for LNGFR. Such differences in the expression of trk A and LNGFR may reflect neuronglial interactions in the survival and maturation of the sympathetic nerves, retina, and tumors in these tissues. PMID- 12114800 TI - Expression of Simple Mucin Type Antigens and Lewis Type 1 and Type 2 Chain Antigens in the Thyroid Gland: An Immunohistochemical Study of Normal Thyroid Tissues, Benign Lesions, and Malignant Tumors. AB - In order to characterize the pattern of expression of carbohydrate structures in several types of thyroid tissues and to evaluate the putative usefulness of the detection of such antigens in diagnostic surgical pathology, we undertook the immunohistochemical study of simple mucin type antigens (T, Tn, and sialyl Tn), Lewis type I antigens (Lewis a, sialyl Lewis a, and Lewis b), and Lewis type 2 related antigens (precursor type 2, H type 2, Lewis x, sialyl Lewis x, and Lewis y) in thyroid samples obtained from 65 patients. The material consisted on paraffin sections of normal thyroid (n = 43), benign lesions (13 goiters/hyperplastic lesions and 15 adenomas), and malignant tumors (12 follicular carcinomas and 27 papillary carcinomas, 5 of which had lymph node metastases) of the thyroid follicular epithelium. Tn, T, and precursor type 2 antigens were the only antigens that were detected and very rarely in normal thyroid. Benign lesions were similar to normal thyroid despite displaying a higher prevalence of immunoreactivity for several antigens of the three groups. Thyroid carcinomas presented a significantly higher level of expression of all types of simple mucin, Lewis type 1, and Lewis type 2 antigens than the normal thyroid and benign lesions. The expression of sialyl Tn was restricted to malignant tumors, and the expression of sialyl Lewis x was closely associated, though not exclusively, to papillary carcinomas. The immunoreactivity was stronger and the number of positive cases was higher in papillary than in follicular carcinomas. No differences were found between primary tumors and the respective metastases. The existence of distinct patterns of expression of carbohydrate antigens in different types of thyroid lesions points to the usefulness of the detection of some of these antigens in thyroid surgical pathology. The putative role of such antigens in the peculiar metastatic properties of thyroid carcinomas remains unsettled. PMID- 12114801 TI - Macrofollicular Variant of Papillary Thyroid Carcinoma: A Case and Control Analysis. AB - The planimetric, flow cytometric, and immunohistochemical characteristics of the macrofollicular variant of papillary thyroid carcinoma (MFVPTC) have not been reported before. The clinical, morphological, immunohistochemical, planimetric, and flow cytometric characteristics of six cases of the MFVPTC and six of the follicular variant of papillary thyroid carcinoma (FVPTC) were analyzed. Patients had undergone surgical treatment. The mean age was 38 (range 29-64 yr), and five were women. Tumors had a mean size of 3.2 cm (range 0.3-4.5 cm). Half were originally diagnosed as goiter. Macrofollicles had a mean diameter of 345.5 um, perimeter of 1237 um, and area of 13,779 um(2), with nuclear changes of PTC. Mean follow-up was 107 mo (range 12-277), and neither lymph node metastases nor recurrence were seen. Differences in diameter, perimeter, and area between the macrofollicular and follicular variants were found. Follicular neoplastic cells were thyroglobulin and 5-100 protein positive in macrofollicles and normofollicles. All were negative to cytokeratin and to high-mol-wt keratin. All tumors were diploid. There were no significant differences in follow-up, DNA content, nor immunohistochemical reactivity. Differences in diameter (p < 0.00006), perimeter (p < 0.0001), and area (p < 0.001) were observed. It is important to recognize this variant because it could be misdiagnosed as benign thyroidal lesions. PMID- 12114803 TI - Parathyroid Hemangioma: A Report of Two Cases. AB - Two cases of intraparathyroid hemangioma, associated with hyperparathyroidism, are reported. The first case showed a typical capillary hemangioma morphology with small branching vascular channels, almost completely replacing the gland's architecture. The second case was a 2-mm cavernous hemangioma associated with glandular hyperplasia. This is, to our knowledge, the first time that this type of lesion is described. PMID- 12114802 TI - Peripheral Nerve Sheath Tumors of the Thyroid Gland: A Series of Four Cases and a Review of the Literature. AB - Primary peripheral nerve sheath tumors (PNSTs) of the thyroid gland are exceptionally rare. Two schwannomas and two malignant PNSTs (MPNSTs), arising primarily within the thyroid gland, were identified in the files of the Endocrine Tumor Registry at the Armed Forces Institute of Pathology. The patients included two females, age 69 and 80 yr, and two males, age 18 and 33 yr. The patients presented with a mass in the thyroid gland confined to a single lobe of the thyroid without involvement of the cervical neck region. None of the patients had a history of neurofibromatosis. The benign tumors were encapsulated, one of them cystic, with the characteristic cellular and nuclear features of schwannomas. The MPNSTs were invasive tumors, effacing the thyroid parenchyma, with a fascicular pattern of growth composed of neural appearing cells with increased cellularity, increased mitotic activity, and with focal necrosis. Immunoreactivity for 5100 protein and vimentin was seen in all tumors. The patients with schwannomas, treated only by surgical resection, were alive without evidence of disease, over a period of 5-33 yr. Both patients with MPNSTs died of the disease 8 mo and 42 mo, respectively, with widely disseminated disease. Primary thyroid PNSTs are exceptionally rare tumors. MPNSTs, in this limited experience, have a fatal outcome irrespective of aggressive adjuvant therapy. PMID- 12114804 TI - Overview of Neuroendocrine Cells and Tumors. AB - The diffuse neuroendocrine system (DNES) is composed of cells and tumors with secretory granules ranging from 50-400 nm in diameter. Members of the DNES commonly stain for chromogranin and synaptophysin by immunohistochemistry and may express a variety of peptide hormones. Recent studies have shown that the proprotein convertases (proconvertases or PCs) are good broad-spectrum markers for members of the DNES. Gene expression can be readily detecting in neuroendocrine cells and tumors by in situ hybridization (ISH). Newer techniques such as in situ polymerase chain reaction (PCR) can be used to detect gene products that are expressed in low copy numbers in neuroendocrine cells. The concept of multidirectional differentiation is an important notion that helps to explain multiple patterns of phenotypic expression observed in some neuroendocrine tumors. PMID- 12114806 TI - Follicular Carcinoma Associated with Minocycline-Induced Black Thyroid. AB - Grossly evident black pigmentation of the thyroid has been observed in a number of patients, most of whom have a history of chronic ingestion of minocycline. In the majority of reported cases, no thyroid dysfunction or lesion has been noted, although rare instances of papillary carcinoma have been described. We describe a patient with a history of chronic minocycline ingestion, who presented with a neck mass of recent onset. Histologic examination of the thyroid revealed diffuse pigment deposition, typical for that seen in association with minocycline ingestion. Also present was a 3.2-cm follicular neoplasm with capsular and vascular permeation, consistent with minimally invasive follicular thyroid carcinoma. This represents the first report of follicular carcinoma associated with minocycline-induced black thyroid (MIBT). PMID- 12114805 TI - Neuroendocrine Tumors of the Head and Neck: A Selected Review with Emphasis on Terminology. AB - Virtually every variant of neuroendocrine neoplasia can occur, at least rarely, in the head and neck region. This review focuses on the terminology surrounding neuroendocrine carcinomas of the larynx and their distinction from morphologically similar but biologically distinctive neoplasms. It is suggested that rare typical laryngeal carcinoids be labeled as such. There is little evidence that these lesions are part of a morphologic continuum. In contrast, more common "carcinoid-like" carcinomas, previously referred to as "atypical carcinoids" are more appropriately labeled as "moderately differentiated neuroendocrine carcinomas." These neoplasms should, in turn, be distinguished from "small cell neuroendocrine carcinomas," although these latter two neoplasms do represent a morphologic and behavioral spectrum. Light microscopic and immunohistochemical features distinguishing neuroendocrine carcinomas of the larynx from paraganglioma, metastatic medullary carcinoma, malignant melanoma, and basaloid squamous cell carcinoma are presented. The second portion of this review outlines the clinicopathologic features of two head and neck neoplasms exhibiting varying degrees of neuroendocrine differentiation. Olfactory neuroblastomas have well-developed neuroendocrine differentiation, almost invariably arise from the olfactory mucosa, typically exhibit low-grade cytologic features, and may have protracted clinical course with an approximately 50% overall 5-yr survival. In contrast, sinonasal undifferentiated carcinoma is a microscopically high-grade neoplasm with minimal, abortive neuroendocrine features, a highly aggressive clinical course, and virtually 100% mortality. They can arise throughout the sinonasal region. PMID- 12114807 TI - Bilateral Thyroid and Ultimobranchial Medullary Carcinoma. AB - The ultimobranchial bodies in human embryos develop from the fourth and fifth branchial pouch complexes along with thymic and parathyroid tissue. They become incorporated within the lateral thyroid lobes and are believed to be involved in the development of C-cells. We report a case of an unusual bilateral thyroid and neck prelaryngeal medullary carcinoma in a 23-year-old male patient who belongs to a multiple endocrine neoplasia type 2a (MEN type 2a) family with thyroid tumors and pheochromocytomas. The medullary carcinoma was located in an abnormal cystic structure that seems to be a remnant of the ultimobranchial body (UBB) in the neck. Within the contralateral thyroid lobe, the medullary carcinoma was associated with C-cell hyperplasia. PMID- 12114808 TI - Apoptosis in Endocrine Glands. AB - Apoptosis or programmed cell death, is a phenomenon with ultrastructural and biochemical characteristics, which is thought to be distinctive from ordinary necrosis. Shrinkage of cells associated with crescent clumps of heterochromatin and formation of membrane-bound apoptotic bodies are thought to represent distinguishing morphologic features. Internucleosomal cleavage of DNA strands reveals a characteristic ladder pattern in gel electrophoresis. Apoptosis is mediated by an active regulatory mechanism, constitutively expressed in normal and neoplastic cells. bcL2, bcl-x, bax, and APO-1/Fas (CD 95) genes are specifically involved in the apoptotic process. Rat thymocytes exposed to glucocorticoids represent a useful model to study cell death. Steroids and peptide hormones play a role in the regulation of apoptosis. Although there is a great interest in monitoring apoptotic process in endocrine cells and their tumors, only a few studies address apoptosis in endocrine glands so far. One goal of future investigation should be directed to explore therapeutic applications. PMID- 12114809 TI - Molecular Diagnosis of Multiple Endocrine Neoplasia (MEN) in Paraffin-Embedded Specimens. AB - In this article, we summarize our recent findings on rearranged during transfection (RE7) mutations in a series of 46 sporadic as well as multiple endocrine neoplasia (MEN) type 2- associated tumors and present results of our family screening efforts to identify MEN 2 and MEN 1 gene carriers. A nonisotopic polymerase chain reaction-based single-strand conformation polymorphism (PCR SSCP) analysis and heteroduplex gel electrophoresis method was used to screen DNA extracted from archival specimens of 22 patients with MEN 2-associated and 24 patients with sporadic tumors for mutations in RETexons 1O, 11, 13, and 16. Point mutations were identified by nonisotopic cycle sequencing of PCR products using an automated DNA sequencer. We found six different missense germ line mutations at cysteine residues encoded by exons 10 and 11 in all patients with MEN 2A or familial medullary thyroid carcinoma (FMTC). The frequency of mutations at codon 634 was higher in patients with MEN 2A than with FMTC and a (63)Cys - Arg mutation was associated with parathyroid disease. A germline Met -* Thr point mutation at codon 918 of the RETtyrosine kinase domain encoded by exon 16 was identified in all MEN 2B patients. Nonpredicted inheritable medullary thyroid carcinomas (MTCs) were detected in two patients and a mosaic postzygotic mutation was found in one additional patient. Tumor-specific (somatic) Met - Thr point mutations at codon 918 were identified in 5 of 13 sporadic MTCs and 2 of 8 sporadic pheochromocytomas (PCCs). The remaining sporadic tumors lacked mutations in all four RET exons tested. In exon 13, a nucleic acid polymorphism (CTT/CTG; Leu) at codon 769 was identified, which is present in approx 40% of the examined population. Our study demonstrates that the molecular methods used are not only suitable to identify asymptomatic individuals at risk for MEN 2A, FMTC, and MEN 2B, but also to distinguish sporadic from inherited tumors using archival tissue specimens; and that more tumors than clinically expected are inheritable, indicating the need for genetic analysis of all MTC and PCC patients. PMID- 12114810 TI - Immunohistochemical Analysis of the Cell Cycle-Associated Antigens Ki-67 and Retinoblastoma Protein in Parathyroid Carcinomas and Adenomas. AB - The morphologic distinction between parathyroid carcinoma and adenoma can be a difficult diagnostic problem. We analyzed nuclear immunoreactivity for the cell cycle-associated antigen Ki-67 with monoclonal antibody (MAb) MIB-1 and for retinoblastoma (RB) protein with two polyclonal antisera in 24 parathyroid carcinomas and 35 adenomas, which were formalin fixed and paraffin embedded to determine if these antibodies could assist in distinguishing between carcinomas and adenomas. In addition, 10 cases of parathyroid hyperplasia and 5 cases of normal parathyroids were examined as control tissues. The Ki-67 labeling index was significantly higher in parathyroid carcinomas compared to adenomas (7.1 +/- 1.0% vs 2.4 +/- 0.2%, p <0.001). No patient with a parathyroid adenoma, parathyroid hyperplasia, or normal parathyroid gland had a Ki-67 labeling index >5.3%. Analysis of the primary tumors from patients with recurrent carcinomas and from those with nonrecurrent carcinomas showed a higher mean Ki-67 labeling index (7.8 +/- 1.5% vs 5.2 +/- 1.1%) in the former group. Although these differences were not statistically significant, the RB protein immunoreactivity was not useful in distinguishing between parathyroid carcinomas and adenomas in paraffin tissue sections. These results indicate that nuclear immunoreactivity for the cell cycle-associated antigen Ki-67 may be another useful method to assist in distinguishing parathyroid carcinomas from adenomas. PMID- 12114811 TI - A Serum-Free System for Primary Cultures of Human Pituitary Adenomas. AB - We report the successful use of a serum-free culture system for primary cultures of human pituitary adenomas. The system utilizes histiotypic suspension culture with low protein-binding membrane inserts that enable cells to retain their three dimensional tissue configuration, closely mimicking the growth pattern in vivo. A serum-free defined medium was developed with CMRL-1969 (Connaught, Willowdale, Ontario, Canada) supplemented with 0.375% albumin bovine Fraction V, 5 ug/mL insulin, 5 ug/mL transferrin, 5 ng/mL sodium selenite, 30 ug/mL putrescine, 6.85 x 10(11)M hydrocortisone, and 3.7 x 10'(11)Mtri-iodothyronine (T(3)). We analyzed eight surgically resected human pituitary adenomas. Basal pituitary hormone secretion measured by radioimmunoassay of pituitary hormones was compared with hormone hypersecretion in vivo and with control cells of the same tumors cultured in CMRL-1969 with 10% fetal calf serum. The light microscopic, immunocytochemical and ultrastructural morphology of cells cultured in this serum-free histiotypic system was compared with cells cultured in serum-supplemented media and with cells cultured on collagen-coated plastic; all cultured cells were compared with the morphology of surgically resected tissues of the same specimens. Basal pituitary hormone secretion during 24-hour incubations correlated with the clinical patterns of hormone excess; the data were similar in serum-enriched and serum-free cultures, however, hormone secretion decreased less rapidly in the serum-free cultures. Cells maintained in the histiotypic culture system closely resembled the corresponding surgically resected tumor using the morphologic parameters and were better preserved than those plated in collagen-coated plastic wells. This comparative study indicates that this serum-free histiotypic culture system provides an ideal method of examining pituitary adenomas in vitro without altering the profile of hormone secretion and cell morphology documented in vivo. This system can be used to examine the production and effects of a wide range of hormones and growth factors that have been implicated as causative agents in pituitary tumorigenesis. PMID- 12114812 TI - Incidence, Pathology, and Recurrence of Pituitary Adenomas: Study of 647 Unselected Surgical Cases. AB - The incidence of various types of unselected pituitary adenomas based on correlation of pathologic and clinical data was assessed. We investigated 647 cases of unselected pituitary adenomas, which were surgically removed between 1980 and 1993. All cases were examined by immunohistochemistry and electron microscopy. The mean age of patients was 44.0 years with 40.0 years for women (55.2%) and 49.1 years for men (44.8%). Age distribution indicated a remarkable sex difference: 52.4% of women and 26.8% of men were between 21 and 40 years at the time of surgery. Based on immunohistochemistry and electron microscopy, prolactin (PRL) cell adenomas represented 263% of tumors, growth hormone (GH) cell adenomas 12.5%, adrenocorticotrophic hormone (ACTH) cell adenomas 12.4%, oncocytomas 12,4%, and gonadotroph cell adenomas 9.4%. Seventy-three percent of the prolactinomas occurred in women and 73.8% of the oncocytomas were found in men. The incidence of pediatrics pituitary adenomas was 4.6%. All 647 cases were followed up; the mean follow-up period was 96.6 months. In 40 patients (6.2%), the adenoma recurred. Recurrence was common in functioning ACTH cell adenomas (8 cases: 9.5%) followed by silent adenomas (7 cases: 25.9%). Recurrence was noted after 2-96 months (average 28.7 months) following surgery. The shortest remission period was found in a patient with oncocytoma followed by a patient with prolactinoma. PMID- 12114814 TI - Characterization of Pheochromocytomas in a Mouse Strain with a Targeted Disruptive Mutation of the Neurofibromatosis Gene Nf1. AB - Patients with neurofibromatosis type 1 (NF1) show an increased frequency of pheochromocytomas. The NF1 gene encodes a GTPase-activating protein that controls the activity of ras proteins in intracellular signaling. A mouse strain with a knockout mutation of Nf1, the murine counterpart of NF1, has recently been constructed. This mutation, designated Nf1(n31), has been shown to be associated with the frequent development of pheochromocytomas in heterozygous animals. Pheochromocytomas are extremely rare in wild-type mice. We have characterized the tumors to assess their relevance as a model for human pheochromocytomas. The frequency of pheochromocytomas was determined in inbred compared to outbred mice carrying the Nf1(31) mutation. Paraffin sections of pheochromocytomas from seven mice were stained immunohistochemically for the catecholamine biosynthetic enzymes, tyrosine hydroxylase (TH), and phenylethanolamine-N-methyltransferase (PNMT) to infer their profiles of catecholamine synthesis, and for chromogranin A (CGA) to infer their content of secretory granules. Cultured cells from a representative tumor were studied in vitro to assess proliferation and neuronal differentiation. Pheochromocytomas arose in approx 15% of Nf1(n31) mice with a mixed genetic background, but were absent in inbred mice. Approximately one fourth of the tumors were bilateral. The tumors exhibited variable morphology. All included cells that appeared well differentiated and resembled normal chromaffin cells in that they expressed TH, PNMT, and CGA. Focal neuronal differentiation was also observed. In cell culture, the tumor cells ceased to proliferate and the majority underwent terminal differentiation into TH-positive cells with neuronal morphology. The phenotype of pheochromocytomas in mice with the Nf1(31) mutation resembles that of human pheochromocytomas, particularly with respect to their ability to produce epinephrine, as inferred from positive staining for PNMT. The tumors also resemble both normal and neoplastic human adrenal medulla with respect to their extensive differentiation into neuron-like cells in vitro. This change in phenotype may be related to ras activation. These neoplasms may be valuable both as models for the pathobiology of adrenal medullary neoplasia, and as a source of epinephrine-producing pheochromocytoma cell lines, for which adequate models currently do not exist. PMID- 12114815 TI - Lymphocytic Adenohypophysitis Associated with Rathke's Cleft Cyst. AB - During the 8th month of her first pregnancy, a 40-year-old female suffered from visual disturbances. After treatment of pericarditis, which appeared 1 month after a normal delivery, she was referred to the neurosurgical department. She showed bitemporal hemianopsia, disturbance of visual acuity, and hypopituitarism. Initial computed tomography (CT) image showed a solid pituitary mass with suprasellar extension. However, 2 months later, the CT image changed to an enlarged partially cystic lesion. Transsphenoidal exploration of the sella demonstrated lymphocytic adenohypophysitis coexistent with Rathke's cleft cyst. To our knowledge, such an association has never been reported previously. Presurgical diagnosis of lymphocytic adenohypophysitis still remains difficult and surgical intervention is necessary for definitive diagnosis. However, special attention is needed for the histological diagnosis of this lesion, particularly in clinically atypical cases. PMID- 12114813 TI - Y Chromosomal Sequences Identified in Gonadal Tissue of Two 45,X Patients with Turner Syndrome. AB - We examined excised gonadal tissue obtained from two 45,X patients for evidence of Y chromosomal material. Both patients had features atypical for individuals with Turner syndrome, a large dysgerminoma in patient 1 and clitoromegaly in patient 2. Southern blot analysis of polymerase chain reaction (PCR)-amplified DNA was performed for five Y chromosome-specific probes (SRY, ZFY. DYZ3, KALY, and DYZ1). Fluorescence in situ hybridization (FISH) with a combination probe specific for the DYZ1/DYZ3 loci was utilized. For both patients, Southern blot analysis of PCR-amplified DNA with primers for the SRY gene was positive. No signals were detected with the other Y chromosome-specific probes for patient 1. For patient 2, positive signals were obtained for all-Y-specific probes. FISH was negative in the gonadal specimen from patient 1, while rare cells were positive in the sections from patient 2. Turner syndrome and mixed gonadal dysgenesis may represent different points on a continuum of disorders of sexual differentiation. Although the risk for gonadal tumors is considered to be low in patients with Turner syndrome, prospective evaluation is critical to ascertain: The frequency of somatic cell mosaicism for cell lines carrying Y chromosomal material, and how the presence of Y chromosomal material in patients with Turner syndrome affects the propensity for virilization and gonadal neoplasms. PMID- 12114816 TI - Endocrine Cell Replacement of Oxyntic Glands in Zollinger-Ellison Syndrome: A Role for Female Sex Hormones? AB - A 48-year-old woman with Zollinger-Ellison syndrome (ZES), but no evidence of multiple endocrine neoplasia (MEN)-1 syndrome, developed an unusually florid evolution of enterochromaffin-like (ECL) cell hyperplasia, which led to extensive replacement of oxyntic glands by endocrine tissue and resulted in the disappearance of the patient's gastric acid hypersecretion with antisecretory treatment no longer required. The patient's previous history included breast cancer, treated with surgery and 5 years of antiestrogen therapy, and bilateral granulosathecal cell tumor of the ovary. In addition, increased circulating levels of 17 B-estradiol (17BE) and progesterone, possibly depending on concomitant development of liver cirrhosis, were also found. On the basis of these associations, it is suggested that abnormalities in the domain of female sex hormones, with a potential synergistic role of liver dysfunction, may be involved in the florid evolution of hypergastrinemia-driven proliferation of ECL cells observed in the present case. PMID- 12114817 TI - p27(kip1) and Other Cell-Cycle Protein Expression in Normal and Neoplastic Endocrine Tissues. AB - Endocrine tumors are often diagnostic challenges. Recent studies have suggested that proteins that regulate cell cycle may have diagnostic and prognostic utility in endocrine tumors. p27(kip1) (p27) is a cyclin-dependent kinase inhibitor that regulates the transition from the G1 to the S phase of the cell cycle. p27 and other cell-cycle proteins are becoming increasingly important in assessing the biologic behavior of endocrine neoplasms, classifying hyperplastic and neoplastic endocrine tissues, and in the progression of endocrine tumors. p27 appears to separate normal from neoplastic endocrine tissues and, in some cases, benign from malignant endocrine tumors. However, there is overlap in p27 expression among individual cases of benign and malignant tumors, and p27 has only limited prognostic utility in endocrine tumors compared to some epithelial tumors such as breast and prostate neoplasms. Thus, more effective molecular and cellular markers that can be used for diagnostic and prognostic purposes in endocrine pathology are needed. PMID- 12114819 TI - Genetic Changes in Chromosomes 1p and 17p in Thyroid Cancer Progression. AB - Little is known about the genetic alterations that occur during the progression of thyroid neoplasms. To understand better the biology of thyroid tumors, we investigated several genetic loci in benign and malignant thyroid neoplasms. Forty-one thyroid tumors (6 adenomas, 16 papillary, 14 follicular, and 5 anaplastic carcinomas) were studied. Normal and tumor cells were microdissected from paraffin-embedded tissues. DNA was used for polymerase chain reaction-based loss of heterozygosity (LOH) analysis with the following markers: D1S243 (1p35 36), D1S165 (1p36) and D1S162 (1p32), TP53 (17p13), and INT-2 (11q13). Immunohistochemistry for Ki-67 was performed. The Ki-67 labeling index (LI) was the percentage of positive tumor cells. LOH at 1p was seen in 2 of 5 (40%) informative cases of anaplastic carcinoma (2 of 2 at D1S162 and 1 of 2 at D1S165) and in 2 of 11 (18%) informative cases of follicular carcinoma (2 of 7 at D1S243, 2 of 7 at D1S1654, and 1 of 6 at D1S162). One anaplastic (20%) and two follicular carcinomas (14%) had LOH in at least two of the 1p loci analyzed. None of the adenomas and papillary carcinomas had LOH at these loci. LOH at 17p and 11q13 were infrequent. Ki-67 LI was 1.4, 7, 16, and 65% in adenomas, papillary, follicular, and anaplastic carcinomas, respectively. Allelic loss at 1p may occur in aggressive types of thyroid carcinoma and may be a marker of poor prognosis. LOH at 1p may represent a late genetic event in thyroid carcinogenesis. LOH at 17p and 11q13 (MEN gene locus) is uncommon in thyroid neoplasms. PMID- 12114818 TI - The Art and Applications of Fluorescence In Situ Hybridization in Endocrine Pathology. AB - Fluorescence in situ hybridization (FISH) or molecular cytogenetics is currently recognized as a reliable, sensitive, and reproducible technique for identifying the copy number and structure of chromosomes. FISH combines molecular genetics with classic cytogenetics and allows simultaneous morphologic evaluation on a single slide. Centromeric DNA probes are used to detect specific chromosomes and telomeric probes to demonstrate all chromosomes. Sequence-specific probes can localize in situ a single gene copy on a specific chromosome locus. FISH allows cytogenetic investigation of metaphase spreads and interphase nuclei. Several protocols have been proposed to analyze preparations from fresh samples or archival material. Comparative genomic hybridization (CGH) is a novel cytogenetic technique, which combines FISH with automatic digital image analysis. Comparative analysis of the hybridization products of tumor DNA and reference DNA with normal metaphase chromosomes, each labeled with color different fluorochrome, can retrieve chromosomal imbalances of the entire genome in a single experiment. FISH and CGH are powerful morphologic tools in understanding physiologic mechanisms and in resolving problems of the pathogenesis of several diseases. These techniques shed light on the cytogenetic background in many endocrinological disorders, providing a better understanding of the activities and alterations of endocrine cell function. PMID- 12114821 TI - Combined Riedel's Disease and Fibrosing Hashimoto's Thyroiditis: A Report of Three Cases with Two Showing Coexisting Papillary Carcinoma. AB - Extensive sclerosis of the thyroid gland can be seen in both benign and malignant conditions. The benign sclerosing lesions of the thyroid include Riedel's disease and fibrosing Hashimoto's thyroiditis. Although these conditions usually occur separately, rarely can they occur simultaneously. In malignant lesions, papillary thyroid carcinoma and anaplastic carcinoma of the thyroid can be associated with extensive sclerosis leading to partial or total effacement of the tumor. We report on three cases that showed simultaneous occurrence of Riedel's disease and fibrosing Hashimoto's thyroiditis. Two of these cases also showed papillary carcinoma (one case of Warthin's-like papillary carcinoma and one case of classic type). All patients were females (age range 32-67 yr) and presented with elevated antithyroglobulin antibodies. Two patients presented with a solitary thyroid mass, and from these one had multiple bilateral neck nodes and a paravertebral mass. The third patient presented with a multinodular gland adherent to neck structures. All patients underwent total thyroidectomy. Histologic sections showed extensive replacement of the thyroid parenchyma with dense keloidal fibrosis, intermixed well-developed lymphoid follicles, and scattered lymphocytes and plasma cells. In all cases the fibrotic process extended beyond thyroid capsule with involvement of the perithyroidal soft tissues and skeletal muscle consistent with Riedel's disease. One case showed a classic papillary carcinoma with bilateral lymph node metastases, and the other showed a Warthin's-like papillary carcinoma. In both cases the papillary cancers were surrounded by dense sclerosis. Immunohistochemical stains for B- and T-markers and immunoglobulin light chains showed a polyclonal population of the lymphoid cells. The simultaneous occurrence of Riedel's disease and fibrosing Hashimoto's thyroiditis is rare and most likely represents a coincidental phenomenon, because both of these conditions are distinct clinicopathologic entities. PMID- 12114820 TI - Expression of the von Hippel-Lindau Tumor Suppressor Gene in Nonneoplastic and Neoplastic Lesions of the Thyroid. AB - Alterations of the von Hippel-Lindau (VHL) gene, which is supposed to act as a tumor suppressor gene, can cause hereditary tumors associated with the VHL syndrome and are found in different sporadic cancers as well. While VHL protein is distinctly detectable in thyroid follicles, so far its expression in nonneoplastic and neoplastic lesions of the thyroid has not been investigated comprehensively. To illuminate the role of VHL for thyroid tumorigenesis, we investigated 12 follicular adenomas; 22 follicular carcinomas; 11 papillary carcinomas; 6 poorly differentiated carcinomas (PDTCs); 9 undifferentiated carcinomas (UTCs); 8 medullary carcinomas; 13 cases with nonneoplastic as well as normal thyroid tissue of 10 patients with antibodies against VHL, vascular endothelial growth factor (VEGF); and the proliferation marker MIB1 immunohistochemically; and selected cases by Western blot analysis. VHL was clearly expressed in nonneoplastic lesions and differentiated tumors derived from follicular epithelium, diminished in PDTCs and very weakly or not detectable in UTCs (p = 0.001), nonneoplastic, and neoplastic C-cells. Although slightly increased in certain differentiated tumors, VEGF was found to be reduced in UTCs as well. In summary, VHL is expressed differently in nonneoplastic and neoplastic lesions of the thyroid in proportion to the level of differentiation. VHL gene alterations appear to be a late event in tumorigenesis of the thyroid and a reduction in VHL protein expression is associated with a loss of differentiation and increased aggressiveness in thyroid tumors. There is no apparent inverse correlation between VHL and VEGF expression as described for other sporadic carcinomas. Therefore, the role of VHL for angiogenesis and the molecular basis of the inactivation of VHL in thyroid tumors remains to be elucidated. PMID- 12114822 TI - Immunocytochemical Localization of Prohormone Convertase 1/3 and 2 in Thyroid C Cells and Medullary Thyroid Carcinomas. AB - Prohormone convertases (PCs) are the key enzymes in the regulated pathways for the post-translational processing of peptide hormones and are involved in converting larger prohormones to smaller, biologically active hormones. Immunolocalization of PC1/3 and PC2 was performed with thyroid C-cells from normal thyroid glands and medullary thyroid carcinomas (MTCs); thyroid C-cells were not consistently positive for PCs, whereas MTCs were consistently positive for PCs. Positive staining for PCs included tumor cell nests adjacent to the main MTCs in cases of multiple endocrine neoplasia type 2. Procalcitonin was originally detected in MTCs and a large amount of procalcitonin was present together with abundant PCs in MTCs. Thus, immunocytochemical staining for PCs may be another characteristic of MTCs. PMID- 12114823 TI - Silver-Staining Nucleolar Organizer Region Quantification in Pituitary Adenomas. AB - Nucleolar organizer regions (NORs) are segments of DNA, encoding for ribosomal RNA. They are associated with argyrophilic proteins and, thus, they can be localized through silver staining. A correlation has been shown between the number, the size, or the intranuclear localization of AgNORs, and the proliferative activity of cells. The aim of this study was to examine numerous features of AgNORs in pituitary adenomas and to relate them to immunohistochemical typing of tumor. Histologic slides from 32 pituitary tumors and one normal pituitary were silver-stained and analyzed with a computerized system for microscopic image analysis, supported by an AgNORmeter95 program. All the tumors were also immunocytochemically characterized. We have found that gonadotropinomas, when compared with pleurihormonal adenomas, revealed a lower proportion of nuclei with a single AgNOR and a higher percentage of marginal dots. Recurrent adenomas, when compared with primary adenomas, showed a higher proportion of nuclei with three AgNOR dots, a larger total area of dots in the nuclei, and a higher standard deviation of the AgNOR dot area in the nucleus. Adenomas immunopositive for prolactin, when compared with immunonegative ones, showed a larger mean area of the AgNOR dot, a larger area of the biggest dot in the nucleus, and a higher proportion of nuclei within a single dot. These results suggest that the estimated parameters of AgNOR dots differ according to tumor aggressiveness and to the hormone immunopositivity of pituitary adenomas. PMID- 12114824 TI - Insulin and Glucagon mRNA Expression and Prohormone Convertase Immunoreactivity in Normal and Neoplastic Pancreatic Endocrine Tissue. AB - Prohormone convertase (PC) 1/3 and PC2 are neuroendocrine-specific enzymes that convert prohormones to active hormones. To learn more about the role of these PCs in normal and neoplastic islet cells, we analyzed a series of pancreatic endocrine tumors to determine the role of PC1/3 and PC2 in islet cell hormone processing. In situ hybridization with insulin and glucagon probes and immunostaining with antibodies to PC1/3 and PC2 were done with 6 normal pancreases, 33 insulinomas, and 7 glucagonomas. The intensity of the reactions was graded from 0 to 3+. Normal islets stained strongly for both proinsulin and proglucagon mRNAs. Insulinomas and glucagonomas had readily detected hormone mRNAs for proinsulin and proglucagon, respectively. Normal pancreatic insulin cells stained weakly for PC1/3 (1+) and PC2 (1-2+), and glucagon cells stained weakly for PC1/3 (1-2+) and strongly for PC2 (2-3+). Both insulinomas and glucagonomas stained strongly for PC2 (2-3+) with less intense staining for PC1 (1-2+). These results indicate that PC2 is more highly expressed in insulin- and glucagon-producing pancreatic islet cell tumors and that there is increased expression of PC2 in insulinomas compared to normal insulin-producing cells. PMID- 12114825 TI - Correlation Between Clinical and Histological Analyses in Retroocular Connective Tissues and Extraocular Muscles from Patients with Graves' Ophthalmopathy. AB - Twenty-two patients with Graves' ophthalmopathy underwent biopsy, and two patients had both eyes biopsied. The samples for the control group (n = 4) were obtained during routine non-thyroid-related corrective strabismus surgery. Ophthalmological evaluation with clinical activity score (CAS), endocrinological evaluation, and ultrasound were used in our study. Correlation between clinical and histological analyses in connective tissues and extraocular muscles from patients with Graves' ophthalmopathy was done. The echography results disclosed an enlargement in all extraocular recti muscles with the exception of one patient. Periodic acid-Schiff and Giemsa stains revealed a moderate number of mast cells in the endomysial connective tissue, none of which displayed significant degranulation. There were no signs of muscle cell damage. Fifteen of the biopsies showed weak cellular reactions with only scattered inflammatory cells. Furthermore, the inflammatory process may be localized and not equally distributed throughout the muscle. Thus, the biopsies might not be representative for the whole muscle. Statistical significance analysis was found when sex and CAS were compared (p = 0.001683) using the Fisher Test. In conclusion, our investigation indicates a pleomorphic pattern of histologic findings in connective tissue and extraocular muscles in Graves' ophthalmopathy. PMID- 12114826 TI - Neurofilaments in Thyrotrophs of Hypothyroid Rats: An Immunohistochemical Study. AB - We examined by immunohistochemistry the expression of 200-kDa (NF-H) neurofilaments in pituitary thyrotrophs of rats, made hypothyroid by propylthiouracil (PTU) administration. Rats were sacrificed at 3, 7, 14, and 28 d of PTU administration as well as at 3, 7, and 14 d after interruption of a 14-d PTU treatment. The pituitaries were fixed, dehydrated, and embedded in paraffin. Consecutive sections were prepared for the immunohistochemical demonstration of thyroid-stimulation hormone and NF-H. NF-H were found in some thyrotrophs of the adenohypophysis of control rats. NF-H were expressed in thyroidectomy cells of hypothyroid rats. Discontinuation of PTU treatment led to recovery from cytologic alterations in the thyroidectomy cells expressing NF-H. PMID- 12114827 TI - Functional salutogenic mechanisms of the brain. AB - Neuroscientists are typically interested in the brain in relation to disease, but much could also be learned by studying the brain in relation to health. The brain has processes, functional salutogenic mechanisms, that contribute to health by enabling one's outlook on life to benefit one's health. For example, the belief that things will work out as well as can reasonably be expected is a key aspect of the outlook of people who tend to stay well even when in potentially stressful situations. Believing in God, feeling happy, being mutually in love, and expecting things to change for the better are also outlooks that can be salutogenic. Beliefs need not even be rational or realistic in order for them to be salutogenic, as shown by phenomena such as faith healing and the placebo effect. Thus, the brain responds to stimuli and interprets them, mainly without one's awareness, in ways that can enhance one's well-being. Although little is presently known concerning neuropathways of functional salutogenic mechanisms, further research on relations between salutogenesis and brain function can be expected to provide new strategies for improving health worldwide. PMID- 12114828 TI - Media appeals for directed altruistic living liver donations: lessons from Camilo Sandoval Ewen. AB - Recently, a Canadian couple sought media help to find a person who was willing to donate part of his or her liver to their infant son, Camilo Sandoval Ewen, who suffered from biliary atresia. Although Canadian transplant programs do not perform transplants from non-emotionally related living donors, they are accepted by some U.S. transplant centers, particularly when donated in a nondirected fashion. This paper examines the ethical questions raised by media appeals for directed altruistic living donations of solid organs, specifically liver lobes, and argues that media-directed altruistic donations are ethically problematic. While they can and do succeed, they raise ethical concerns regarding (1) who has access to the media; (2) who will be successful in their media campaigns; and (3) how coercion and inducement by brokers (families or institutions) can be prevented. Justice requires that the media adopt a policy not to cover organ appeals by families, despite their human interest value, and that transplant centers develop a unified policy restricting directed living donations to those who have an emotional relationship. PMID- 12114830 TI - A natural history of "agonist". AB - This paper constructs a brief history of the biochemical term agonist by exploring the multiple meanings of the root agon in ancient Greek literature and describing how agonist first appeared in the scientific literature of the 20th century in the context of neurophysiologists' debates about the existence and properties of cellular receptors. While the narrow scientific definition of agonist may appear colorless and dead when compared with the web of allusions spun by the ancient Greek agon, the scientific power and creativity of agonist actually resides precisely in its exact, restricted meaning for biomedical researchers. PMID- 12114829 TI - Bioethical pluralism and complementarity. AB - This essay presents complementarity as a novel feature of bioethical pluralism. First introduced by Neils Bohr in conjunction with quantum physics, complementarity in bioethics occurs when different perspectives account for equally important features of a situation but are mutually exclusive. Unlike conventional approaches to bioethical pluralism, which attempt in one fashion or another to isolate and choose between different perspectives, complementarity accepts all perspectives. As a result, complementarity results in a state of holistic, dynamic tension, rather than one that yields singular or final moral judgments. PMID- 12114831 TI - Thomas Kuhn is alive and well: the evolutionary relationships of simple life form -a paradigm under siege ? AB - A Kuhnian framework is used to analyze the current controversy over whether two or three fundamental types of life forms exist. Until the 1980s, all life was classified into two primary forms: eukaryotes and prokaryotes. Using molecular sequencing data, Carl Woese suggested the archaebacteria constituted a third domain. In the mid-1990s, Radhey S. Gupta challenged the three-domain hypothesis. While this dispute may seem to be a purely technical debate over the analysis of protein and nucleic acid sequence data, the controversy encompasses broader issues such as the aims of classification and the role of microorganisms in the biosphere. At the heart of this dispute is what kinds of data are relevant to constructing an overall taxonomy. The prestige of molecular biology played a large role in why the three-domain hypothesis was accepted so readily, but supporters of the two-domain hypothesis argue that the fossil record, morphology, and cell physiology should all play a role in taxonomy. This case study provides a good example of a paradigm shift in the making, demonstrating that issues beyond the raw data will be significant factors in deciding whether the three domain hypothesis will prevail or a new classificatory scheme will emerge. PMID- 12114832 TI - Vignettes of medical school during the war, 1942-1945. PMID- 12114833 TI - Medical validity in Eastern and Western traditions. AB - When comparing biomedical treatments and traditional treatments from China and India, validity is established by a randomized placebo-controlled trial (RPCT). While the advantages of RPCT are uncontestable, its requirements handicap validation of treatments from Eastern traditions, which are integrated within a worldview that encompasses natural philosophy, theology, empirically based clinical experience, and spiritual and moral tenets. RPCT evolved during a time when comparatively little was known about Eastern medical traditions; about the effects of emotional, cognitive, and cultural factors on healing; or about the evolutionary biology and psychology of innate mind/body mechanisms, such as those producing placebo responses, which may constitute evolved adaptations naturally selected during human evolution. A fair way of both securing the advantages provided by RPCT and balancing them with a methodology that ensures safe and efficacious treatments from other traditions can be facilitated by examining medical validity in light of generalizations from evolutionary psychology and the cultural study of medicine. PMID- 12114834 TI - The fault lines of academic medicine. AB - Unprecedented advances in biomedical research and the upheaval in health care economics have converged to cause seismic changes in the traditional organization of medical schools and academic health centers. This process is particularly evident in departments of internal medicine. The activities and functions of academic medicine are in the midst of separation and realignment along lines that do not honor historical departmental and divisional boundaries. The organization of a successful medical school or department must be dynamic, constantly serving its constituents to accommodate progress and change and to promote optimal structure for academic productivity. PMID- 12114835 TI - The original "Jaws" attack. PMID- 12114836 TI - Advances in modern electrocardiographic equipment for long-term ambulatory monitoring. AB - Electrocardiographic ambulatory "Holter" monitoring (AECG) is an essential tool in the diagnostic evaluation of patients with cardiac arrhythmias. Recent advances in digital Holter technology have improved the quality of the ECG signals and new dedicated algorithms have expanded the clinical application of software-based AECG analysis systems. Due to the availability of inexpensive large storage capacities, very long-term (weeks to months) continuous high quality AECG monitoring will soon be available, together with devices for long term long-distance telemetric surveillance for high-risk cardiac patients, utilizing trans-telephonic transmission of ECG data. New digital recorders will also have the capability for multichannel simultaneous recordings (currently from 3 to 8 simultaneous leads). Multichannel digital recordings will allow the recording of different biological signals by appropriate sensors, such as respiratory frequency, peripheral oxygen tension, arterial pulse pressure, EEG, and others. This will transform conventional AECG in ambulatory policardiography, allowing the comprehensive evaluation of patients with complex disorders, such as heart failure or sleep apnea syndromes. By this global approach, Holter analysis becomes a real "noninvasive electrophysiological test," to identify potential risk factors for life-threatening cardiac arrhythmias. PMID- 12114837 TI - Update on intensive care ECG and cardiac event monitoring. AB - This brief review is aimed primarily as a resource for the clinician and summarizes recent advancements in electrocardiographic monitoring in the intensive care unit. Emphasis is placed on recent advances in ICU ECG and cardiac event monitoring with particular attention to arrhythmia detection in patients following myocardial infarction. Specific topics addressed include: clinical indicators of impending arrhythmic events and sudden death, signal averaged ECG, QT dispersion, ST segment fluctuation, T-wave alternans, QT interval beat-to-beat variability, heart rate variability, and advances in automated arrhythmia detection. PMID- 12114838 TI - Electrocardiographic ST-segment changes during acute myocardial ischemia. AB - The recognition and management of patients with acute coronary syndromes has relied to a large extent on the standard 12-lead electrocardiogram (ECG) for assessing ST-segment changes associated with ischemia. The purpose of this review is to show both the capabilities and the limitations of the 12-lead ECG in recognizing ischemia, and to seek alternative electrocardiographic leads, optimized for detection of ischemia originating in different regions of the ventricular myocardium. Three such leads are proposed-based on the results obtained by electrocardiographic body-surface mapping performed during ischemia induced by balloon-inflation coronary angioplasty. A survey of recent clinical studies shows that the electrocardiographic manifestations of acute myocardial ischemia observed during coronary angioplasty are in agreement with the ST segment measurements in admission ECGs of patients with acute myocardial infarction. PMID- 12114839 TI - State of the art in stress testing and ischaemia monitoring. AB - Electrocardiography remains the most widely used method for detecting myocardial ischemia. ST segment abnormalities in the resting 12-lead electrocardiogram in subjects with angina and coronary risk factors seem to definitely indicate ischemic heart disease and an adverse prognosis. ST depression during exercise testing is the first line provocative test for ischemic heart disease although it has a mean sensitivity of only 68% and a slightly higher specificity (77%). The presence or absence of chest pain in patients with an ischemic ST response to exercise testing does not change the risk of future ischemic events. However, ST depression during the recovery period is associated with increased risk both for acute coronary events and coronary death, whereas silent ischemia during recovery is an even stronger predictor than during exercise. The amplitude of ST depression has not been documented to reflect the magnitude of ischemia. Therefore, new methods are under investigation such as adding R and Q wave amplitude criteria, maximal ST/heart rate slope, linear regression analysis of the heart rate related change in ST depression and a score integrating ST segment amplitude and slope changes. The demonstration of episodic ST segment depressions in the ambulatory setting, even without accompanying chest pain, are an expression of transient ischemia and such episodes seem to represent a poor prognosis. In the hospital setting, ST depression detected by continuous monitoring is related to the clinical outcome. ST segment monitoring during the first 6-9 hours after coronary care unit admission provides important prognostic information on-line and considerably improves early risk stratification. Such continuous ST monitoring overcomes some of the limitations of static monitoring, as it improves the likelihood of capturing the maximal point of ST deviation, as well as early episodes of reocclusion that are manifest as recurrent ST elevation. PMID- 12114840 TI - Electrocardiogram of the failing heart. AB - In the failing heart general specific (e.g., Q-waves after acute myocardial infarction, persistent ST-elevations in post-myocardial infarction left ventricular aneurysm) and unspecific ECG changes (e.g., left bundle branch block, right bundle branch block, ST-T-alterations due to digitalis glycosides or antiarrhythmic drugs) may be seen in the conventional 12-lead ECG. In addition, atrial and ventricular tachy-arrhythmias may be detected and quantified by 24 hour-Holter ECG recordings, that may be relevant for a worse prognosis of patients with congestive heart failure. Heart rate variability as the most relevant derived ECG parameter of sympathetic tone fluctuations may be of important prognostic significance in congestive heart failure patients. An abnormal signal averaged P-wave duration may predict the incidence of atrial fibrillation, as may apply to QRS-prolongation and/or ventricular late potentials in the signal averaged ECG for the incidence of serious life-threatening ventricular tachy-arrhythmias or death from pump failure. Last but not least, cardiac repolarization abnormalities may be detected by QT dispersion-, QT-/QTc fluctuation- or T-wave alternans studies, but the true prognostic significance of these parameters for predicting sudden cardiac death or death from pump failure in patients with congestive heart failure remains unclear. PMID- 12114841 TI - P wave assessment: state of the art update. AB - Diagnostic (mapping) and therapeutic (ablation, pacing) advances have provided insight into atrial depolarization processes and new developments in P wave analysis. Information about interatrial pathways is important to the understanding of interatrial conduction delay. A standardized method for P wave analysis is necessary for the development of a clinical role for management of patients with paroxysmal atrial fibrillation using signal-averaged P wave analysis and P wave dispersion. Algorithms for predicting localization of ectopic P waves may facilitate catheter ablation. P wave changes due to pacing at different atrial sites may be useful for permanent pacing for prevention of atrial fibrillation. Introduction of these developments into clinical practice should allow better prevention and treatment of atrial arrhythmias and could have considerable impact in view of their high frequency especially in the older population. PMID- 12114842 TI - Future concepts in P wave morphological analyses. AB - The idea of analyzing P wave inscriptions to discriminate between patients with different diseases is not new. Whereas standard approaches deal mainly with an analysis of the duration of the P wave, there is an increasing interest for the P wave morphology. Detailed analysis of the low voltage P wave will require a hardware setup that enables high-quality signal acquisition with low noise interference in combination with an ECG subtraction technique. The feasibility of the surface ECG to detect the presence of an abnormal electrophysiological substrate in the atrial myocardium as well as to localize ectopic atrial rhythms will be enhanced by use of P wave morphological analysis. PMID- 12114843 TI - Monitoring of atrial tachyarrhythmias. AB - Atrial fibrillation, the most common tachyarrhythmia, significantly reduces the quality of life of patients and is associated with 15% of strokes. With the aging of the population, it is beginning to reach epidemic proportions. Guidelines for management of atrial tachyarrhythmias exist but in order to implement them, it needs early diagnosis and execution of treatment strategies. Atrial tachyarrhythmias are treated by drugs, ablative therapies, implantable electrical devices or external Cardioversion. An ideal monitoring systems should diagnose the tachyarrhythmia early and provide adequate information to help define the appropriate treatment strategy. Presently, external monitoring systems are routinely used for diagnosis. Although they are effective in patients with palpitations, their value is significantly limited in asymptomatic patients or those with other symptoms as well as for evaluating the efficacy and safety of treatment. Many of these limitations can be addressed with chronically implantable devices because of their accurate detection and ability to store large amounts of data. Presently, implantable devices are indicated for patients with Bradyarrhythmias and Tachyarrhythmias who can benefit from the electrical therapy provided by the devices. About a third to half of these patients have concurrent atrial tachyarrhythmias. The extensive information available from some of the recently available devices should provide significant value for diagnosing atrial tachyarrhythmias, optimizing treatment strategies and improving morbidity and mortality. This needs to be investigated with prospective trials. In the future, if the diagnostic information can be shown to have significant value, these devices may be implanted for their diagnostic capabilities only. PMID- 12114844 TI - Non-invasive characteristics of atrial fibrillation: the value of Holter recordings for the treatment of AF. AB - Atrial fibrillation (AF) is the most common arrhythmia in man associated with significant morbidity and excess mortality. AF can be 'lone' but is frequently associated with underlying heart disease while in some patients a genetic cause has been identified. In the past decade our knowledge about the mechanisms of AF and our options for (non)pharmacological treatment of AF have increased importantly. Since the success rate of drug therapy is frequently disappointing "hybrid therapy" is often necessary (e.g., drugs in combination with cardioversion, pacemaker implantation or an ablation procedure). Therapy should focus on identifying the specific substrate (underlying heart disease) and triggers for AF in each patient, making a more individualized therapy possible. For this, non-invasive testing becomes more and more important. Holter recordings may show focal activity (monomorphic atrial premature beats, atrial tachycardia) or other supraventricular arrhythmias (AV-nodal reentrant tachycardia, circus movement tachycardia) which can successfully be treated. In addition, AF may transiently convert to atrial flutter (AFL), either spontaneously or after administration of (class IC) drugs. Recent studies have shown that ablation of the flutter circuit or ectopic activity can cure AF in many of these patients. PMID- 12114845 TI - Assessing heart rate variability from real-world Holter reports. AB - Real world clinical Holter reports are often difficult to interpret from a heart rate variability (HRV) perspective. In many cases HRV software is absent. Step-by step HRV assessment from clinical Holter reports includes: making sure that there is enough usable data, assessing maximum and minimum heart rates, assessing circadian HRV from hourly average heart rates, and assessing HRV from the histogram of R-R intervals and from the plot of R-R intervals or heart rate vs. time. If HRV data are available, time domain HRV is easiest to understand and less sensitive to scanning errors. SDNN (the standard deviation of all N-N intervals in ms) and SDANN (the standard deviation of the 5-min average of N-N intervals in ms) are easily interpreted. SDNN < 70 ms post-MI is a cut point for increased mortality risk. Two times ln SDANN is a good surrogate for ln ultra low frequency power and can be compared with published cut points. SDNNIDX (the average of the standard deviations of N-N intervals for each 5-min in ms) < 30 ms is associated with increased risk in patients with congestive heart failure. RMSSD (the root mean square of successive N-N interval difference in ms) < 17.5 ms has also been associated with increased risk post-myocardial infarction. Frequency domain HRV values are often not comparable to published data. However, graphical power spectral plots can provide additional information about whether the HRV pattern is normal and can also identify some patients with obstructive sleep apnea. PMID- 12114846 TI - Clinical implications of present physiological understanding of HRV components. AB - Time and frequency domain analysis of heart rate variability (HRV) is a non invasive technique capable of providing information on autonomic modulation of the sinus node and of stratifying risk after myocardial infarction and in heart failure. One of the basic assumptions used to explain the negative predictive value of reduced HRV was the concept that overall HRV was largely dependent on vagal mechanisms and that a reduction in HRV could reflect an increased sympathetic and a reduced vagal modulation of sinus node; i.e., an autonomic imbalance favouring cardiac electrical instability. This initial interpretation was challenged by several findings indicating a greater complexity of the relationship between neural input and sinus node responsiveness as well as the possible interference with non neural mechanisms.Nevertheless, the prognostic value of time and geometric parameters of HRV has been consistently confirmed. More complex is the interpretation of spectral parameters particularly when they are computed on 24-hour recordings. Under controlled conditions, instead, the computation of low and high frequency components and of their ratio seems to provide information on sympatho-vagal balance in normal subjects as well as in most patients with preserved left ventricular function, thus providing an unique tool to investigate neural control mechanisms. More recently, analysis on nonlinear dynamics of HRV has been utilized to describe the fractal-like characteristics of the variability signal and has been shown to identify patients at risk for sudden cardiac death. In conclusion, in spite of an incomplete understanding of the physiological significance of HRV parameters, this non invasive methodology is of substantial utility to evaluate autonomic control mechanisms and to identify patients with an increased cardiac mortality. PMID- 12114848 TI - A phenomenon of heart-rate turbulence, its evaluation, and prognostic value. AB - Identification of high-risk cardiac patients is crucial for stratification strategies and prevention of cardiovascular events, including death. Single ventricular premature beat triggers some oscillations in cardiac cycle duration (the shortening followed by the lengthening of the cycle intervals) in healthy subjects and low-risk patients with ischaemic heart disease and/or heart failure. This phenomenon is called heart-rate turbulence (HRT). It was shown in retrospective and prospective studies that the absence of HRT is associated with increased risk of subsequent mortality in cardiac patients. HRT can be quantified by two variables: turbulence onset (TO), describing an early acceleration phase, and turbulence slope (TS), describing a late deceleration phase of heart rate after ventricular premature beat. Both TO and TS are independent one from another and from other conventional risk predictors. The combination of TO and TS seems to be the strongest Holter-based risk predictor and has some addictive predictive value to left ventricular ejection fraction, heart rate variability, and the averaged diurnal heart rate and baroreflex sensitivity. In addition, HRT has a predictive value in patients treated with beta-blockers and amiodarone. Moreover, it is thought that HRT is mediated by baroreflex and therefore can be used as a non-invasive measure of its sensitivity and autonomic nervous system function. Blunted HRT can be observed in diabetic patients with autonomic dysfunction and in patients with atropine-blocked vagal nerve activity. Moreover, it seems that a diurnal variation of HRT exists because it is better expressed during sleep. However, the use of HRT is limited to patients with dominant sinus rhythm and the presence of single ventricular beat. Nevertheless, the assessment of HRT is an inexpensive and simple method and can be performed with a routine ambulatory 24 hour ECG recording. PMID- 12114847 TI - Clinical applicability of heart rate variability analysis by methods based on nonlinear dynamics. AB - Analysis of heart rate (HR) variability has become an important widely used method for assessing cardiac autonomic regulation. Conventionally, HR variability has been analyzed with time and frequency domain methods. Analysis of HR dynamics by methods based on nonlinear systems theory has opened a novel approach for studying the abnormalities in HR behavior. Recent studies have shown that these measures, particularly scaling analysis methods of HR dynamics, are altered among various patients populations with cardiovascular diseases, and they provide prognostic information. Altered long-term scaling properties of HR dynamics and more random short-term HR fluctuation has been observed, e.g., among patients with previous myocardial infarction, and these alterations have been shown to be associated with increased mortality rate. A relatively large body of data indicate that altered scaling properties of R-R intervals are physiologically deleterious. These findings support the notion that some nonlinear methods, such as scaling and complexity measures, give clinically valuable information for risk stratification among various patient populations. This article provides a review of our current knowledge of the usefulness of dynamical measures of HR fluctuation. PMID- 12114849 TI - Mechanisms involved in heart rate turbulence. AB - Proper understanding of the mechanisms involved in heart rate turbulence (HRT) may offer an explanation of why it is such a potent postinfarction risk stratifier. This article reviews the physiological background of ventriculophasic sinus arrhythmia-a phenomenon which shares some underlying physiological features with HRT including cardiac autonomic regulation. It is now believed that HRT is principally triggered by a transient loss of vagal efferent activity in response to the missed baroreflex afferent input due to ventricular premature beat-induced haemodynamically inefficient ventricular contraction. Studies are summarized which support more or less directly this hypothesis. The physiology of early acceleration and late deceleration of heart rate after a ventricular premature beat is discussed. Qualitatively different but otherwise quantitatively uniform postectopic dynamics of systolic blood pressure after ventricular premature beats is demonstrated in subjects with normal and abnormal left ventricular function. It is concluded that the slope of late deceleration of heart rate after ventricular premature beats can serve as a reasonable surrogate for baroreflex sensitivity. PMID- 12114850 TI - Circadian rhythm in the cardiovascular system: considerations in non-invasive electrophysiology. AB - Most cardiovascular activities show a circadian rhythm, as do electrophysiological phenomenon. Under the influence of both external stimuli and endogenous homoeostatic mechanisms, cardiac electrophysiological properties change diurnally and enable the cardiovascular system adapt to rest-exercise cycles. According to recent reports, almost all non-invasive electrophysiological phenomena, such as electrocardiographic indices, cardiac refractoriness and conduction, pacing and defibrillation threshold, heart rate variability indices, and even Q-T dispersion and T-wave alternans, show diurnal variability. Furthermore, many of these changes are clinically significant and may affect results of some diagnostic studies. These characteristics of the cardiovascular system require us keep in mind the "time" factor any time we analyze electrophysiological results and make clinical decisions. PMID- 12114851 TI - QT interval measurements. AB - The QT interval, which represents duration of ventricular electrical systole, i.e., the time required for completion of both ventricular depolarization and repolarization, has been a parameter of particular interest in cardiology. However, the relationship between duration of cellular action potentials and the QT interval recorded at the body surface is very complex. As a result, the QT interval is difficult to measure with precision. First, there is inherent imprecision in identifying the end of the T wave because of incomplete understanding of the recovery process and its projection on the body surface. Second, significant variation both in the onset of the QRS complex and the end of the T wave among some ECG leads provides different QT values depending on the leads selected for measurement. Third, technical factors such as paper speed and sensitivity influence QT measurements with higher paper speed leading to shorter interval values and higher sensitivity resulting in QT prolongation. The above problems do not appear to be solved by automatic QT measurement techniques, which have been found to be less accurate in cardiac patients than in healthy controls. In conclusion, we should accept that QT interval remains merely a gross measure of ventricular electrical systole and/or repolarization and we should not expect significant improvement in accuracy of traditional QT interval measurements. Rather, in clinical research, methods examining the shape or amplitude of the T wave and its changes related to heart rate should be exploited. PMID- 12114852 TI - Physiological mechanisms influencing cardiac repolarization and QT interval. AB - Cardiac repolarization primarily results from activation of outward currents carried by potassium ions which are heterogeneously distributed throughout the myocardial layers which can lead to changes in various phases of cardiac cycle. QT interval is a reflector of cardiac repolarization and can be influenced by heart rate, autonomic nervous system, gender and time of the day. T wave alternans (TWA), specifically the discordant pattern is a feature of abnormal cardiac repolarization that can be used as a predictor of ventricular arrhythmias. PMID- 12114853 TI - Individual patterns of QT/RR relationship. AB - In clinical practice, an imprecision introduced by ad hoc selected heart rate correction formula of the QT interval is unlikely to lead to erroneous conclusions if all borderline cases are carefully considered. On the contrary, in clinical investigations (e.g., studies of drug effects) the over- or undercorrection of QTc may lead to significant and systematic bias with both false positive and false negative findings. None of the previously published "global" heart rate correction formulae has been universally successful because the QT/RR relationship is different between different subjects and a formula that corrects the QT interval for heart rate acceptably in one individual may be very misleading in another individual. Moreover, it has been recently established that the QT/RR patterns not only exhibit a substantial inter-subject variability but also a high intra-subject stability. Thus, in precise investigations, individual QT/RR relationship should be first established in each subject and subsequently translated into individual heart rate correction formula. PMID- 12114854 TI - QTc interval in the assessment of cardiac risk. AB - In the United States alone 300,000-400,000 people die of sudden cardiac death every year. Much of this mortality is assumed to be caused by ventricular tachyarrhythmias. Prolonged QTc reflect cardiac repolarization prolongation and/or increased repolarization inhomogenity known to be associated with increased risk of arrhythmias. The paper gives a review of the possibilities to assess the risk of ventricular arrhythmia and/or cardiac death from QTc. Prolonged QTc may hold independent prognostic importance for mortality in common diseases as ischemic heart disease and diabetes mellitus where as the prognostic importance in heart failure and arterial hypertension is more uncertain. In more rare diseases as the inborn long QT syndrome the QT interval gives not only important hint to the diagnosis but the magnitude also provides information on prognosis. QTc has probably no independent prognostic importance in hypertrophic cardiomyopathy or in the arrhythmogenic right ventricular disease. The degree of QTc prolonging during treatment with QTc prolonging drugs is prognostic for the risk of ventricular arrhythmia in form of torsade de pointes and QTc prolonging drugs should probably not be prescribed for patients with a QTc greater than 460 ms and withdrawn if QTc exceeds 500 ms during treatment. Data from the DIAMOND study suggest that QTc can be used to point out those heart failure patients who will benefit from antiarrhythmic therapy. PMID- 12114855 TI - Why did QT dispersion die? AB - BACKGROUND: Considerable controversy exists about the meaning of QT dispersion (QTD). The working hypothesis of the present paper was that the necessary although not sufficient condition for the validity of QTD concept is the association of QTD with nondipolar voltage (NDPV) in T waves of the 12-lead ECG. METHODS AND RESULTS: ECGs of 4,890 subjects, 966 with coronary heart disease (CHD) and 3,844 considered CHD-free were processed using computer programs for measurement of the ratio of the first two eigenvalues (E2/E1), nondipolar voltage (NDPV), terminal T wave direction and ECG estimate of left ventricular mass (LVM). The mean NDPV in T wave was 11 muV (SD 3.9), with 6 muV (SD 1.3) in terminal 40 ms. NDPV alone explained only 6% and NDPV, E2/E1 and LVM combined 13% of QTD variance. There was a modest increase in the fraction of subjects with QTD >60 ms among subjects with NDPV in terminal T > 7 muV compared to those with NDPV >/=7 muV (15% vs. 10%). A more profound increase was associated with terminal T wave direction deviating from normal (37% vs. 12% among those with normal direction), reflecting dipolar rather than nondipolar components. CONCLUSIONS: The association between QTD and NDPV is weak, and QTD is unlikely to represent any meaningful myocardial repolarization event in the interval domain. It seems more logical to use direct measurement of NDPV as a potential marker of localized dispersion and heterogeneity of ventricular repolarization for evaluation of the risk of adverse cardiac events. PMID- 12114856 TI - Remodeling of cardiac repolarization: mechanisms and implications of memory. AB - Memory is a well established property of biological organisms, allowing them to adapt to their environment and respond to novel stimuli. Sensitization occurs in response to a noxious stimulus, and increases the behavioral response to subsequent stimuli. In contrast, habituation occurs in response to an innocuous stimulus, and decreases the behavioral response to subsequent stimuli. Therefore, the response of an organism to a stimulus does not simply depend on the stimulus, but also on previous stimuli that the organism has received. Similarly, the response of the heart to a stimulus does not simply depend on the stimulus, but also on previous patterns of depolarization and repolarization, due to electrical remodeling. Electrical remodeling, the persistent change in electrophysiological properties of myocardium in response to a change in rate or activation sequence, has been well described in atria. It can be induced by rapid pacing or atrial fibrillation (AF), and results in shortened atrial refractory period and increased susceptibility to atrial arrhythmias. These changes have been associated with alterations of potassium and calcium currents. However, the fundamental mechanisms responsible for triggering changes in channel expression in response to alterations in rate and activation sequence in AF are poorly understood. Even less is known about electrical remodeling in ventricle. Rapid ventricular pacing or an alteration of ventricular activation sequence produces persistent changes in heterogeneity of repolarization and, in contrast to atria, a prolongation of action potential duration. Ventricular electrical remodeling is responsible for "T-wave memory," which is observed commonly in patients after periods of altered activation sequence (e.g., chronic pacing). These changes have been associated with alterations of potassium currents, specifically I(to), implying that electrical remodeling is heterogeneously expressed in the different cell types across the transmural wall. Finally, remodeling of gap junctions may also play a prominent role in action potential changes during remodeling. The signal transduction pathways through which a change in rate or activation sequence triggers changes in the expression of ionic currents are being actively investigated. PMID- 12114857 TI - T-wave patterns associated with the hereditary long QT syndrome. AB - Mutations involving 6 different ion-channel genes have been identified in subjects with the hereditary Long QT Syndrome. These gene mutations result in structural and functional changes in ion-channel proteins with resultant alterations in potassium and sodium repolarization currents that affect the morphologic features of electrocardiographic repolarization. This review highlights the genotype-phenotype associations related to ventricular repolarization that have been reported in the LQTS literature, with particular focus on ECG T-wave patterns in LQT1, LQT2, and LQT3 genotypes. PMID- 12114858 TI - Practical use of T wave morphology assessment. AB - QT dispersion (QTd) has not proven to be a useful marker derived from the 12-lead electrocardiogram (ECG) for stratification of patients at risk for sudden cardiac death. To overcome its methodological shortcomings, novel ECG variables of T wave morphology have been proposed. The total cosine R-to-T (TCRT), T wave morphology dispersion, T wave loop dispersion, normalized T wave loop area, as well as absolute and relative T wave residuum evaluating non-dipolar ECG signal contents were evaluated in two clinical studies involving post myocardial infarction (MI) patients and US veterans with cardiovascular disease. In 280 post MI patients with 27 events over a mean follow-up of 32 months, TCRT and T wave loop dispersion were independent predictors of mortality. In 813 male US veterans with cardiovascular disease the absolute and relative T wave residua were independent predictors of patient risk during a long-term follow-up of more than 10 years. On Cox regression analysis, age, presence of left ventricular hypertrophy (LVH) and left ventricular ejection fraction (LVEF) were also predictors of survival. The latter study in US veterans therefore was the first to demonstrate that a novel parameter characterizing heterogeneity of ventricular repolarization within the 12-lead surface ECG permits risk stratification in patients with cardiovascular disease. All of the ECG variables are easily accessible from digital 12-lead surface ECG recordings using custom computer programs. They may prove useful to identify risk patients that benefit from the implantable cardioverter defibrillator (ICD). PMID- 12114859 TI - Clinical value of T-wave alternans assessment. AB - Microvolt-level T-wave alternans (TWA) is a new arrhythmia risk marker to assess subtle changes in repolarization that has been introduced for arrhythmia risk stratification. Recent experimental studies have demonstrated that it reflects a heartrate dependent increased spatial dispersion of repolarization associated with unidirectional conduction block, and reentry that may result in the occurrence of ventricular fibrillation. Clinical studies have convincingly demonstrated that TWA is closely related to arrhythmia induction in the electrophysiology (EP) laboratory as well as to the occurrence of spontaneous ventricular tachyarrhythmias in patients undergoing EP study. Subsequent studies showed that TWA-assessed noninvasively-is predictive of future arrhythmic events in patients with implanted ICDs as well as for ventricular tachyarrhythmias in patients with congestive heart failure without a prior history of arrhythmias. There is still controversy, however, about the predictive value of TWA in patients following acute myocardial infarction (MI). Several studies which differ in patient selection, pharmacologic treatment of the patients, and endpoint definitions, have reported conflicting results. Therefore, studies with a large number of unselected patients after acute MI on optimal treatment according to contemporary therapeutic guidelines as well as of patients with reduced left ventricular ejection fraction following MI are needed to define its role with regard to identifying patients who may benefit from primary preventive ICD therapy. Future research should also focus on evaluation of alternative methods to increase heart rate (i.e., pharmacological stimulation) in an attempt to reduce the proportion of incomplete tests in patients with insufficient increase in heart rate during exercise testing. PMID- 12114860 TI - Optimizing ambulatory ECG monitoring of T-wave alternans for arrhythmia risk assessment. AB - Considerable scientific data support the potential value of T-wave alternans (TWA) as an index of vulnerability to ventricular fibrillation. This chapter summarizes our state of knowledge regarding the use of routine ambulatory ECGs to evaluate TWA and discusses recent methodologic approaches designed to optimize AECG-based TWA analysis for arrhythmia risk stratification. Newer methods, including the nonspectral technique of Modified Moving Average analysis, appear promising in detecting TWA during the changing conditions associated with daily activities. The Modified Moving Average approach does not require specialized electrodes and is not encumbered by the need to achieve target heart rates, as is the case for conventional spectral-based methods. Guidelines are provided for evaluating latent cardiac electrical instability using AECG-based TWA testing. These recent developments make possible the TWA analysis of ambulatory ECGs not only in prospective trials but also in vast stores of archival data. PMID- 12114861 TI - [Early transvaginal measurement of cephalic index for the detection of fetuses at risk for Down syndrome]. AB - BACKGROUND: The aim of this study was to evaluate the utility of cephalic index screening in early pregnancy for the identification of fetuses at risk for trisomy 21. METHODS: Measurements of cephalic index, calculated as the ratio between biparietal diameter/occipito-frontal diameter, were obtained in 36 Down syndrome fetuses and the values were compared with normal data in the same gestational periods considered. RESULTS: Cephalic index was found to show fairly constant values throughout the period evaluated. The measurements obtained in Down syndrome fetuses were similar to those obtained in euploid fetuses. CONCLUSIONS: The data obtained show that in early pregnancy cephalic index cannot be considered a useful tool in the detection of fetuses at risk for Down syndrome. PMID- 12114862 TI - Screening and treatment for cervical intraepithelial neoplasia (CIN) in HIV infected women. AB - BACKGROUND: To determine: 1) whether the pathology correlates with the degree of immunosuppression, 2) whether there is a relation between pathology and antiretroviral therapies, 3) whether Papanicolau (Pap) smears correlate with colposcopic and histologic findings, 4) whether there is rapid genital disease progression after standard gynaecologic care. METHODS: Retrospective study. Immunologic, gynaecologic and virologic data were extracted either from patients charts or from laboratory testing. RESULTS: At first visit Pap smears resulted normal in 43.7% of the women, 8.4% of the patients had reactive and reparative changes, 2.8% atypical cells of undetermined significance, 33.8% low-grade squamous intraepithelial lesions and 11.3% high-grade squamous intraepithelial lesions. Patients with a normal PAP smear had higher CD4 cell count (318+/-191 cells/mL) compared to patients with squamous intraepithelial lesions (297+/-116 cells/mL) but the difference was not statistically significant (Mann-Whitney test). The distribution of cervical dysplasia was found to be similar regardless of antiretroviral therapy (chi(2) test). The sensitivity and specificity of Pap tests for detecting CIN were 94 and 80%. Twenty-two per cent of surgically treated women had persistent or recurrent disease. CONCLUSIONS: Lower CD4+ cell counts are not predictive of the presence of cervical dysplasia. All HIV-infected women, independently from their immunological and clinical conditions, need regular Pap smears with appropriate follow-up for abnormal cervicovaginal cytology, this could prevent nearly all deaths from cervical cancer. PMID- 12114863 TI - [Cervico-vaginal infections. Study of a population in the Turin area]. AB - BACKGROUND: Aim of this study is to determine the prevalence, in the Turin area, of the pathogens chiefly involved in the genesis of the most common infections of the female genitalia. METHODS: This is a retrospective study. During the period of time beginning from January 1, 1997 and ending December 31, 1999, we examined 10,249 women from 14 years to 54 years of age, who were seen at the out-patient diagnosis service of the Sant'Anna Hospital. The patients' cervical specimens were screened for common germs, Candida spp., Trichomonas vaginalis, Bact. Vaginosis, Chlamydia trachomatis, Microplasms, and Neisseria gonorrhea. The prevalence of each micro-organism was found. The obtained data were cross referenced with the risk factors in the clinical history of each patient. The chi(2) test with a C.I. of 95% was used for the statistical evaluations. RESULTS: It is shown by a detailed analysis of the data in our possession that an anamnesis oriented mainly for the evaluation of the various risk factors would be more effective than one oriented for the presence of a subjective symptomatology, since many of these infections are either weakly symptomatic or totally asymptomatic (especially in the case of C. trachomatis), as has been many times underlined in the international literature. CONCLUSIONS: Greater attention should be given to the collection of anamnestic information in order to more precisely target for examination those subjects at greater risk of contracting a sexually transmitted infection. PMID- 12114864 TI - Risk of genital prolapse and urinary incontinence due to pregnancy and delivery. A prospective study. AB - BACKGROUND: Recent literature shows conflicting results regarding this subject. Using a prospective study, we analysed the possible effects of pregnancy and delivery upon the pelvic floor support. METHODS: In a group of 344 patients who received our phone-call 3 months after delivery, only 58 accepted the investigation, and came for an exam. We looked out for pathologies such as genital prolapse and stress urinary incontinence (IUS). During the exam we analysed: vagino-perineal scars; descensus of the vaginal walls and of the uterus; dyspareunia; urinary frequency and urgency; urge Incontinence and IUS; weakening of pelvic floor muscles. RESULTS: We objectively identified in cystocele the prevalent "anatomic" damage, and in IUS, the most frequent "functional" damage. We then tried to find a statistical correlation between these pathologies and the most important risk factors cited in the literature. CONCLUSIONS: Through the systematic analysis of the obtained data, we thus identified the most important risk factors that lead to the development of these pathologies: operative delivery, pluriparity, heavy work, high BMI in mothers and newborns. The results that emerged from our study lead to some remarks of interest and discussion. PMID- 12114865 TI - [Therapy for cervical intraepithelial neoplasia and fertility]. AB - BACKGROUND: We reviewed the case records of patients of childbearing age treated with various types of surgical techniques for cervical intraepithelial neoplasia (CIN) to determine the impact of surgical treatment on their fertility. METHODS: Between 1983 and 1997 a total of 486 women with CIN received surgical treatment at out unit. Laser vaporization was used in 196 cases, cold-knife conization in 163 and REP in 127. The outcome of the various treatments was then compared. RESULTS: Independent of the surgical technique used, the percentage of pregnancies achieved after surgery was high: 93.33 and 96.66% of patients treated with laser vaporization and REP, respectively, and 87.69% of those who received cold-knife conization. The differences did not reach statistical significance nor were significant differences observed in the number of abortions or in the method of birth delivery (spontaneous, Cesarean section). However, a higher percentage of premature births was noted among women who received cold-knife conization (31.57%), which was statistically significant in the comparison among the three groups. CONCLUSIONS: The results from our study indicated which techniques for the treatment of CIN may be preferable. Compared with the other two techniques, cold-knife conization bears higher costs (hospitalization, general anesthesia) and has been superceded by laser vaporization and REP as evaluated in this series. When cold-knife conization must be used, cerclage of the cervix uteri should be performed in the event of future pregnancy. In contrast, laser vaporization and REP can be performed in an outpatient setting with local anesthesia. These techniques, because they are conservative, afford the advantages of complete lesion removal and maintenance of reproductive capability. Another important consideration is that REP is less costly and allows histological examination of the surgical specimen. PMID- 12114866 TI - [Foetal-maternal alloimmunizations in the South-East area of the Venice province]. AB - BACKGROUND: This study report the results obtained in a retrospective analysis of the foetal-maternal alloimmunizations observed from 1993 to 1999 in the South East area of the Venice province. METHODS: The data concerning 17,000 pregnancy observed in this area from 1993-1999 have been collected. For each pregnancy data concerning maternal ABO, Rh, Kk and IAT as well as foetal ABO, Rh, Kk and DAT were available. Further data (mainly antibodies concentration and specificity) were available if a foetal-maternal alloimmunization was detected and if transfusional support was given after the birth. RESULTS: The authors observed 465 alloimmunizations (prevalence 2.7%): 381 (82%) of these were due to an ABO foetal-maternal incompatibility, 23 due to D incompatibility and the other 61 due to other blood groups antigens. Only 6 cases needed transfusional support: 5 exchange transfusion (a patient needed 2 exchanges) and a delayed transfusion. CONCLUSIONS: Foetal-maternal alloimmunizations are today a rare but not exceptional event (about 3% of pregnancy), the great majority of these alloimmunizations are due to an ABO incompatibility. Despite the prevention of alloimmunization in D negative women by using Rh immune globulin, anti-D alloimmunization is still observed. A great number of other blood groups antigens are involved in foetal-maternal alloimmunization mainly within the Rh system (CcEe, etc.). In the authors' experience the great majority of foetal-maternal alloimmunizations were clinically silent, only 6 cases (1.3% of patients with a positive DAT) needed transfusional therapy. PMID- 12114868 TI - [Water birth. Clinical and practical advice]. AB - Water labor and/or birth are more and more requested and with the increased number of pregnant women choosing water-birth it is necessary to define advice for a safe use of this method of birth-assistance, in order to prevent any possible connected complication. The criteria for admission or exclusion to water labor and/or birth, correlated with maternal-fetal clinical aspects, hygienic aspects, legal-medicine aspects, are indicated. The structural, technical, organizing, specific requirements, in addition those of every delivery room, and considered important for water birth places, are underlined. PMID- 12114867 TI - [The perimenopause. Problems and therapeutic changes]. AB - The female life period in which hormonal share begins to drop and the first menopausal clinical symptoms occur, is called perimenopausal period. During this female life phase, frequently, time of regular function and time of ovarian dysfunctions occur, with a limited luteal phase and an estrogen production fall. So, perimenopausal clinical symptoms begin: hot flushes, tiredness and libido decrease; to these problems, others can be connected to inadequate luteal phase, as breast pain, nervousness and body increase. Therefore, it's possible to affirm that the perimenopause is a particularly delicate period, either because it represents a transient moment to climateric phase, or because it's possible to detect a great hormonal instability, fundamental step for clinical problems. In our investigation we discuss this problem, explain the causes and the possible remedies to delay the onset of symptoms or to treat hormonal perimenopausal modifications. PMID- 12114869 TI - Antenatal diagnosis and obstetric management of cystic hygroma occurring in twin pregnancies. A case report. AB - The lymphatic system has an early development in the embryo. Usually, its development is complete by the 40th postconceptional day. If this connection fails to develop, lymphatic stasis and dilatation of the lymphatic channels may occur, causing a number of pathologies such as: lymphangiomas, lymphangiectasis and cystic hygromas. Prenatal diagnosis can be made during the first trimester of pregnancy by ultrasonographic examination. A case of a twin pregnancy associated with cystic hygroma and bilateral hypoplasia of lower and upper limbs of both foetuses without chromosomal abnormalities is reported. PMID- 12114870 TI - [HRT in post-menopausal women: endometrial histology and bleeding patterns]. AB - BACKGROUND: In this open prospective study the correlation between bleeding patterns and endometrial histology has been evaluated in 101 postmenopausal women after 2 years of continuous sequential hormone replacement therapy (HRT). METHODS: All patients received continuous transdermal 17-b-estradiol supplementation, 0.05 mg/daily, with cyclic progestogen for 12 days every month. The progestogen was: dydrogesterone 10 mg/daily (56 cases); nomegestrol 5 mg/daily (15 cases); MAP 10 mg/daily (15 cases); norethisterone 0.25 mg/daily (15 cases). The changes in the characteristics of bleeding pattern and endometrial biopsy were performed in 90 of 101 patients, at the 10-12th of progestogen therapy. RESULTS: The endometrial pattern was secretory in 60 cases, proliferative in 5 and atrophic in 22. In 3 cases the endometrial histology showed a simple hyperplasia. The bleeding generally starts 2 days after the end of progestogen therapy (13th +/- 2.9 day), with a mean duration of 4 days (4 +/- 2.8); in 21 patients (~20%) the bleeding is reduced. The endometrial histological characteristics haven't any influence on the bleeding pattern. CONCLUSIONS: In this study there was a low incidence of simple hyperplasia (3%), but the characteristics of bleeding don't permit to suspect this hyperplasia. PMID- 12114872 TI - Circulating tumor markers and nuclear medicine imaging modalities: breast, prostate and ovarian cancer. AB - Clinical oncologists have always shown great interest in circulating tumor markers. There are several markers that in the clinical routine are a signal of particular tumor types; some of them are strictly tissue-specific such as prostatic specific antigen (PSA) for prostatic cancer, AFP and HCG for germ cell tumors of the testis and ovary, others such as CA 15.3, CA125, CEA or cytokeratins are less specific since their elevations can be found in different varieties of cancers even if they are preferentially associated to a certain tumor type, thus are considered markers for breast, ovarian cancer and colon adenocarcinoma. The most useful clinical applications of these parameters is their determination during the follow-up of the treated patients, in order to detect the tumor recurrence early, and also to evaluate the evolution of the disease by monitoring the treatment responses. During follow-up, increasing levels of tumor markers can be observed even several months before the clinical demonstration of cancer recurrence. The association of tumor marker tests with imaging modalities can lead to several advantages: the first is to confirm the diagnosis of relapses, possibly before the appearence of the related clinical symptoms due to tumor growth; the second is to localize the sites of lesions, while tumor markers provide only a general indication of the existence of metastases; the third is to make possible a correct whole body restaging. In the assessment of cancer response tumor markers are often very reliable and their changes are faster than the morphological ones. Among all the imaging modalities, nuclear medicine plays an important role in detecting recurrences and metastatic localizations as it is able to investigate functional rather than morphological aspects of tumors, and provide different information in comparison to morphologic imaging. In addition, the scintigraphic techniques offer the possibility to evaluate treatment responses, confirming or not the information from biochemical changes. This review aims to show some examples (breast, prostate and ovarian cancer) in which the combination of nuclear medicine imaging modalities and tumor marker tests is proposed for clinical practice. The advantages and some critical aspects are discussed on the basis of the clinical findings and the most important clinical indications are described. PMID- 12114873 TI - PET-imaging in tumors of the reproductive trac. AB - There is increasing evidence that metabolic imaging with positron-emission tomography (PET) using fluor-18 labeled fluorodeoxyglucose (18F FDG) is highly accurate for in vivo detection of a variety of malignancies. This quality gives FDG-PET an important role in the detection of malignant tumors and their metastases as well as for differentiation of tumors of unknown etiology. In the male and female reproductive tract, whole body imaging with FDG-PET is in particular capable of visualizing lymph-node and distant metastases before these changes become apparent on conventional cross-sectional imaging modalities. According to the incidence of tumors in the reproductive tract, FDG-PET-imaging has been evaluated in prostate cancer, ovarian cancer, cervical and testicular cancer. The role of PET is discussed with respect to the current management of patients. The presented data indicate that FDG-PET is more accurate for lymph node staging in cervical cancer and testicular cancer. In ovarian cancer, FDG-PET may be helpful for detection of tumor recurrence. The role of FDG-PET is questionable in prostate cancer, due to the low metabolic activity of this type of cancer. Carbon-11 labeled acetate and carbon-11 or fluor-18 labeled choline are more promising than FDG for detection of recurrence in prostate cancer. In all other tumors of the reproductive tract there is limited experience with PET for a final conclusion. PMID- 12114874 TI - Early diagnosis of recurrent breast cancer with FDG-PET in patients with progressive elevation of serum tumor markers. AB - BACKGROUND: The aim of this work is to assess the diagnostic value of positron emission tomography (PET) with 18F-fluorodeoxyglucose (FDG), in the early detection of tumour recurrence in already treated breast cancer patients in apparent complete remission and with a progressive elevation of tumour markers CEA and/or CA 15.3 without any other clinical or instrumental signs of relapses. METHODS: The author studied 45 women (mean age 58+/-12, range 35-80 years) with histological diagnosis of breast cancer who underwent a tumour marker-guided whole body FDG-PET. All patients were in remission, without any other clinical or instrumental signs of relapses, except for the progressive elevation of CA 15.3 and/or CEA, tested during the follow-up. FDG-PET results were controlled by pathology when histological sampling was possible, by other conventional imaging modalities (US, X-rays, CT, MRI) and/or by clinical follow-up up to 12 months at least. RESULTS: FDG-PET findings were evaluated in 38 patients: 27 resulted positive. Among these 27 PET positive patients 24 were true positive and 3 false positive. Tumour marker guided FDG-PET was also able to discover 3 unknown neoplasms not visualized by other modalities. PET revealed 54 sites of intense focal FDG uptake. The anatomical distribution of these sites was 19 skeleton, 18 lymph node basins, 5 liver, 5 pelvic region, 1 lung, 1 pericardium, 1 pleura, 1 contralateral breast, 2 peritoneum and 1 thyroid bed. Forty-eight of these 54 sites of FDG accumulation were confirmed to be metastases. FDG-PET resulted negative in 11 patients and only in 2 of them the other diagnostic modalities were able to discover metastatic lesions; we had 9 true negative and 2 false positive RESULTS. On the basis of our investigation the performances of tumour marker guided FDG-PET per patient are as follows: sensitivity 92% (24/26), specificity 75% (9/12), positive predictive value 89% (24/27), negative predictive value 82% (9/11), accuracy 87% (33/38). CONCLUSIONS: This study demonstrated the clinical utility of tumour marker-guided PET in the follow-up of breast cancer patients. This diagnostic approach allowed to modify the clinical management in those patients in whom a tumor relapse or unexpected primary neoplasm was discovered. PMID- 12114875 TI - Qualitative and quantitative comparison between images obtained with filtered back projection and iterative reconstruction in prostate cancer lesions of (18)F FDG PET. AB - BACKGROUND: Recently, iterative reconstruction with segmented attenuation corrections (IRSAC) has been introduced for reconstruction of (18)F-FDG PET images. IRSAC produces images that are more pleasing to the eye, but qualitative and quantitative comparisons between IRSAC and filtered back projection (FBP) have not been reported for metastatic cancer. Since quantitative data has been widely used as an adjunct to interpretation of PET scans, comparison between IRSAC and FBP is needed. The purpose of this study was to compare image quality and the maximum standardized uptake value (SUVmax) obtained with FBP and with IRSAC in metastatic lesions from prostate cancer. METHODS: Twenty (18)F-FDG PET scans (10 baseline and 10 follow-up) were performed in 10 patients with prostate cancer (ages 66-85 yrs, mean 73.6 yrs). Acquisition began 45 min after injection of 370 MBq of (18)F-FDG. Images were reconstructed using FBP and IRSAC, and submitted to visual and quantitative analysis. SUVmax was obtained for all metastases, on FBP and IRSAC. A Jaszczak phantom study was also performed. RESULTS: IRSAC images showed better image quality than FBP especially in regions of high activity concentrations. IRSAC detected 106 lesions on both baseline and follow-up scans, while FBP detected 100 and 95 lesions on baseline and follow-up scans, respectively. Therefore, 17 more lesions were seen on IRSAC. The mean SUVmax values on baseline scans for FBP and IRSAC were systematically different, at 4.46+/-1.99 and 5.13+/-2.67, respectively. On follow-up scans values were 3.89+/-1.72 for FBP and 4.29+/-1.93 for IRSAC. Comparison of FBP with IRSAC on baseline and follow-up scans were statistically significant (baseline: paired "t" test p=0.0017; follow-up: paired "t"-test p=0.0008). Phantom studies reveal that these differences can be explained by the type of reconstruction filters used, and IRSAC was more accurate than FBP. CONCLUSIONS: IRSAC detects smaller volumes in phantoms, patient images are easier to interpret and more metastatic lesions were detected. In addition, IRSAC provides reproducible quantitative data, comparable to data provided by FBP. IRSAC SUV and FBP SUV are in close agreement but there was a statistically significant difference between the two, and therefore threshold values of SUV will probably need to be re-determined with IRSAC, and are likely to be 10 to 19% higher than currently reported. PMID- 12114876 TI - The role of (111)In Capromab Pendetide (Prosta-ScintR) immunoscintigraphy in the management of prostate cancer. AB - (111)In Capromab Pendetide (ProstaScintR) is a whole murine antibody that is reactive with prostate specific membrane antigen (PSMA), a glycoprotein on the surface of normal and abnormal prostate epithelium. It has proven to be of great value in assisting management decisions in prostate cancer patients who initially present with high risk for metastatic spread, or who develop a picture of recurrent disease after surgery or radiation therapy. Patterns of metastatic lymphatic spread have correlated well with autopsy reports in the literature. Unfortunately, other imaging study and/or histologic confirmation of scintigraphic findings has been difficult to obtain. Prostascint's role in predicting durable complete response (DCR) in postoperative patients having salvage radiotherapy to their prostate fossa is very promising. Further investigative work in larger patient populations is needed to confirm these early results. PMID- 12114877 TI - Internal mammary node lymphoscintigraphy and biopsy in breast cancer. AB - BACKGROUND: In patients with breast cancer, sentinel nodes (SNs) are detected outside the axilla in 1-2% of cases after superficial injection of radiocolloid in the breast. We investigated whether deep injection of tracer visualized internal mammary chain lymph (IMC) nodes more often, and assessed the impact of IMC status on disease staging. METHODS: A total of 400 patients were enrolled in this trial. The study group included 200 patients with T1-T2 N0 breast cancer in an inner quadrant. Radio tracer was injected superficially in 100 (group A), and deeply under the tumor in the others (group B). If an IMC took up tracer in group B patients it was biopsied. An additional 200 patients with outer quadrant lesions were also studied lymphoscintigraphically following superficial (100 patients) or deep (100 patients) injection, but IMC nodes were not biopsied as this would have required an additional surgical excision. RESULTS: An SN was visualized in the IMC in 65.6% of inner quadrant patients after deep injection and in 2.1% after superficial injection. In outer quadrant patients, deep injection visualized an SN in the IMC in 10% of cases. The IMC SN was located mainly in the 2nd and 3rd intercostal spaces. Radioguided IMC biopsy was performed in 62 patients. Node removal proved simple and risks insignificant. Stage migration occurred in 8% of cases. CONCLUSIONS: Deep injection allows SN localization in the IMC in 65% of inner quadrant breast lesions. Biopsy of the axillary plus IMC resulted in stage migration in 8% of patients. It is unclear whether this additional information can lead to better survival. PMID- 12114879 TI - A review of the efficacy of bone scanning in prostate and breast cancer. AB - Bone scintigraphy has provided valuable data in the assessment and management of neoplastic disease since being first described in the early 1960s. There have been many developments in imaging techniques and radiopharmaceuticals over the years allowing more reliable detection of metastatic spread to bone. Other imaging modalities are also evolving roles in the detection of metastatic spread including computed tomography (CT), magnetic resonance imaging (MRI), and positron emission tomography (PET). Despite this, isotope bone scans continue to have a central role in detection and surveillance of bone metastases in breast and prostate cancer. Paralleling developments in imaging there have been enormous changes in the treatment options available for cancers of the breast and prostate that have metastasised to bone. Bone specific treatments including radionuclides and bisphosphonates as well as high dose chemotherapy provide potential improvement in disease control. There is also evidence that earlier treatment of bone metastases may prolong survival. This increases the need for efficient methods of detection and monitoring of disease. In this article we discuss the efficacy of bone scintigraphy in breast and prostate cancer from the point of view of staging, systematic follow-up of asymptomatic patients, evaluation of symptomatic patients and the assessment of response to therapy. PMID- 12114878 TI - Radioguided occult lesion localisation (ROLL) and surgical biopsy in breast cancer. Technical aspects. AB - BACKGROUND: The surgical management of non-palpable breast lesions remains controversial. At our Institute we have introduced a new technique, radioguided occult lesion localisation (ROLL) to replace standard methods and overcome their disadvantages. In this paper technical aspects of ROLL and results on a large series of patients are reported. METHODS: We analysed 812 consecutive patients with 816 non-palpable breast lesions detected mammographically or ultrasonically. (99m)Tc-labelled particles of human serum albumin (7-10 MBq) in 0.2 ml saline were injected into the lesion under stereotactic mammographic or ultrasonic guidance. Mammography and scintigraphy were then performed. With ultrasound guidance only scintigraphic control was necessary. The excision biopsy was carried out with the aid of a hand-held gamma-detecting probe, and entire removal of the lesion was verified by X-ray of the specimen. RESULTS: The tracer was correctly positioned initially in 772/816 (94.6%) cases and at second attempt in another 2. In 42/816 (5.1%) cases, lesion localisation had to be repeated using a traditional approach. X-ray demonstrated the lesion was entirely removed in 770/774 (99.5%) cases. Pathological examination revealed 367 (47.4%) benign lesions and 407 (52.6%) cancers. The cancers were treated by conservative breast surgery in 99.5% of cases. CONCLUSIONS: We concluded that ROLL enables the surgeon to remove occult breast lesions easily and reliably. PMID- 12114880 TI - A historical survey of scintigraphic methodology for evaluation of the reproductive tract. AB - Scintigraphic techniques have contributed to many aspects of our understanding of genital and reproductive physiology and pathophysiology. Few of these methods have become mainstream diagnostic techniques; nonetheless, their flexible, physiologic, and intrinsically quantitative nature have contributed information and insights not readily available by other means. The techniques discussed in this review measure various dynamic processes within the body, including blood flow, variation of blood volume, and lymphatic and fallopian tube transport. Dynamic measurement of these processes exploits nuclear medicine's ability to radiolabel and monitor substances while preserving normal physiologic behavior. Consideration of these methods will potentially stimulate future development and application of radionuclide techniques to emerging questions in the fields of reproductive and genital physiology. PMID- 12114881 TI - Use of umbilical artery base excess: algorithm for the timing of hypoxic injury. AB - Intrapartum asphyxia is responsible for only a small proportion of cerebral palsy cases, although obstetricians are often held accountable. Umbilical cord pH and blood gas values provide valuable information regarding the status of the infant at birth; base excess determination quantifies the magnitude of metabolic acidosis, the putative risk factor for central neurologic injury. Human and animals studies have confirmed normal values of base excess before labor, and consistent rates of base excess change in relation to the degree of fetal hypoxemia or heart rate patterns. Thus, the combination of assumed base excess values before labor and measured values after birth, together with an assessment of degrees of fetal hypoxemia during labor, permits an interpolation of fetal base excess values throughout the course of labor. Because threshold levels of base excess (eg, -12 mmol/L) have been associated with an increased risk of neonatal neurologic injury, this approach provides a framework for the assessment of fetal heart rate tracings during labor and, potentially, the timing of hypoxic/ischemic injury. PMID- 12114882 TI - A comprehensive ethical framework for fetal research and its application to fetal surgery for spina bifida. AB - The transition from innovation to standard of care for fetal surgery should be guided by ethical considerations. We provide an ethical framework that identifies criteria for the investigation of fetal surgery. This framework addresses the initiation and assessment of clinical trials to determine whether they establish a standard of care, describes an appropriate informed consent process, considers whether selection criteria should include the abortion preferences of the pregnant woman, and considers whether physicians have an obligation to offer referral to such investigation. This ethical framework, in a clinically comprehensive fashion, takes account of the physician's obligations to the fetal patient, the pregnant woman, and future fetal and pregnant patients. We apply this framework to clinical investigations of fetal surgery for spina bifida. PMID- 12114883 TI - Can cervicography be improved? An evaluation with arbitrated cervicography interpretations. AB - OBJECTIVE: The purpose of this study was to estimate the optimal performance of cervicography. We compared an arbitrated cervigram classification with an arbitrated referent diagnosis of cervical neoplasia. STUDY DESIGN: From an initial group of 8460 women, a stratified sample of cervigrams from 3645 women and histologic information from 414 women underwent arbitration. Interobserver agreement was assessed for cervicography and the referent diagnosis. Sensitivity, specificity, and predictive values were estimated for initial and arbitrated cervicography results, compared with the initial and arbitrated referent diagnoses. RESULTS: For the detection of arbitrated high-grade lesions or cancer, arbitrated cervicography yielded an overall sensitivity of 63.9% and a specificity of 93.7%. Significantly higher sensitivity was associated with younger age and age-related visual characteristics. CONCLUSION: Optimization of the cervigram classification improved performance over a single interpretation in this population but suggested the limits of static visual screening. PMID- 12114884 TI - Expression of homeobox gene transcripts in trophoblastic cells. AB - OBJECTIVE: This study was conducted to examine the dynamics of homeobox gene expression in the differentiation of trophoblasts as a key to the understanding of the regulatory mechanisms that are involved in placental development. STUDY DESIGN: Expression of homeobox genes was examined in primary trophoblastic cells and in the BeWo choriocarcinoma model cell lines by molecular and immunocytochemistry techniques. RESULTS: We demonstrated the expression of 3 homeobox genes (HOX B6, HOX C6, and HOX A11) in primary trophoblastic cells. BeWo cells showed an expression pattern similar to that of the primary cell lines. In both primary trophoblasts and BeWo cells, the HOX A11 gene, but not the HOX B6 or HOX C6 genes, were found to down-regulate with differentiation from single- to multinucleate giant cells. CONCLUSION: This study demonstrates a novel expression pattern for HOX A11 gene in trophoblastic differentiation and suggests that the down-regulation of HOX A11 may be necessary for the differentiation of cytotrophoblasts into syncytiotrophoblasts. PMID- 12114885 TI - Effect of long-term hormone replacement therapy on plasma homocysteine in postmenopausal women: a randomized controlled study. AB - OBJECTIVE: The purpose of this study was to investigate the long-term effect of hormone replacement therapy on total homocysteine and to study whether there was any difference in effect between opposed and unopposed hormone replacement therapy or whether the methylenetetrahydrofolate reductase C677T polymorphism was associated with the effect of hormone replacement therapy on total homocysteine. STUDY DESIGN: Two hundred nine healthy postmenopausal women were randomized to hormone replacement therapy (n = 103) or no substitution (n = 106) 5 to 7 years earlier. RESULTS: Women who received hormone replacement therapy had significantly lower total homocysteine concentrations than women in the control group; median total homocysteine values were 8.6 micromol/L and 9.7 micromol/L, respectively, in a per-protocol analysis (P =.02). The effect was comparable in all methylenetetrahydrofolate reductase genotypes, and no difference between unopposed and opposed hormone replacement therapy could be demonstrated. Similar results were obtained when an intention-to-treat analysis was performed. CONCLUSION: Long-term hormone replacement therapy results in lower total homocysteine concentrations in all methylenetetrahydrofolate reductase genotypes without demonstrable difference in effect between unopposed and opposed hormone replacement therapy. PMID- 12114887 TI - Predicting postoperative voiding efficiency after operation for incontinence and prolapse. AB - OBJECTIVE: The purpose of this study was to determine the postoperative postvoid residual volume that predicts a voiding efficiency after operation for incontinence and prolapse. STUDY DESIGN: Ninety-nine patients met the criteria for inclusion. The patients' bladders were filled retrograde through gravity, through a Foley catheter, with 300 mL of sterile water, or by subjective fullness. The catheter was then removed, and the patient was asked to void spontaneously into a measured urine collection device ("Texas hat"). A postvoid residual was then calculated (amount voided minus amount inserted). The Foley catheter was reinserted for patients with a postvoid residual of >50%. These patients were instructed to follow-up in the office in 1 week, at which time the test was repeated. This process continued until the patient had a postvoid residual of < or = 50% or was taught intermittent self-catheterization. RESULTS: Ninety patients were available for follow-up. The overall failure rate (defined as cases that required reinsertion of a catheter) was 7.8%. No patient with a postvoid residual of < or = 32% required reinsertion of a catheter. CONCLUSION: With the use of this simple bedside test, voiding efficiency was predicted in 92% of patients who voided > or = 50% of the amount inserted and in 100% of patients who voided > or = 68%. PMID- 12114886 TI - Duloxetine versus placebo in the treatment of stress urinary incontinence. AB - OBJECTIVE: The purpose of this study was to assess the efficacy and safety of duloxetine, a selective inhibitor of serotonin and norepinephrine reuptake, in the treatment of stress urinary incontinence. STUDY DESIGN: A double-blind, randomized, placebo-controlled study was conducted in 553 women aged 18 to 65 years with a predominant symptom of stress urinary incontinence. Subjects were randomized to placebo (n = 138 women) or duloxetine at one of three doses (20 mg/d, n = 138 women; 40 mg/d, n = 137 women; or 80 mg/d, n = 140 women). Outcome variables that were assessed after 12 weeks of treatment included incontinence episode frequency recorded in a real-time diary and answers provided to the Patient Global Impression of Improvement scale and the Incontinence Quality of Life questionnaire. RESULTS: Duloxetine was associated with significant and dose dependent decreases in incontinence episode frequency that paralleled improvements that were observed in the Patient Global Impression of Improvement scale and the Incontinence Quality of Life questionnaire. The median incontinence episode frequency decrease with the use of the pooled diary analysis with placebo was 41% compared with 54% for duloxetine 20 mg per day (P =.06), 59% for duloxetine 40 mg per day (P =.002), and 64% for duloxetine 80 mg per day (P <.001). One half of the subjects at the 80 mg per day dose had a > or = 64% reduction in incontinence episode frequency (P <.001 vs placebo); 67% had > or = 50% reduction (P =.001 vs placebo). These improvements were observed despite significant concurrent dose-dependent increases in the average voiding interval in the duloxetine groups compared with the placebo group. Similar statistically significant improvements were demonstrated in a subgroup of 163 subjects who had more severe stress urinary incontinence (> or = 14 incontinence episode frequency per week; 49%-64% reduction in incontinence episode frequency in the duloxetine groups compared with 30% in the placebo group). Discontinuation rates for adverse events were 5% for placebo and 9%, 12%, and 15% for duloxetine 20, 40, and 80 mg per day, respectively (P =.04). Nausea was the most common symptom that led to discontinuation. None of the adverse events that were reported were considered to be clinically severe. CONCLUSION: This trial provides evidence for the efficacy and safety of duloxetine as a pharmacologic agent for the treatment of stress urinary incontinence. PMID- 12114888 TI - Familial incidence of urinary incontinence. AB - OBJECTIVE: The purpose of this study was to evaluate whether urinary incontinence is more common in family members of women with incontinence compared with continent individuals. STUDY DESIGN: Women who were examined at 2 different outpatient facilities over a 2-year period received a questionnaire that covered social, behavioral, and medical issues. They were also asked whether they had a family member who complained of urinary incontinence and, if so, the degree of the relationship. Subjects were excluded for the following reasons: not mentally competent, difficulty understanding the written English language, and a history of bladder cancer or of acquired or congenital neuropathy. Statistical analyses were conducted with chi-squared tests for differences between groups; a probability of <.05 was defined as significant. RESULTS: A total of 833 women received the questionnaire; 667 women answered the question about urinary incontinence in family members. These 667 women were divided into 3 groups: group I, 441 incontinent women from the first facility; group II, 112 continent women from the first facility; and group III, 114 continent women from the second facility. Women with at least 1 relative with urinary incontinence were 34.9% in group I, 16.1% in group II, and 5.3% in group III. This difference was statistically significant. In a comparison of group I and group II, the odds that an incontinent woman had at least 1 relative with incontinence were 2.6 times higher (95% CI, 1.50-4.48); comparing group I and group III, the odds were 9.6 times higher (95% CI, 4.17-22.25). CONCLUSION: In our study population, women with urinary incontinence were more likely to have at least 1 family member also with incontinence when compared with women who were continent. PMID- 12114889 TI - Morphometric analysis of smooth muscle in the anterior vaginal wall of women with pelvic organ prolapse. AB - OBJECTIVE: The purpose of this study was to compare the smooth muscle content of the anterior vaginal wall in normal women and women with pelvic organ prolapse. STUDY DESIGN: Specimens were taken from the apex of the anterior vaginal cuff after abdominal hysterectomy from 28 women with pelvic organ prolapse and 12 control subjects. Smooth muscle cells of the anterior vaginal wall were identified by immunohistochemistry with antibodies to smooth muscle alpha-actin. Morphometric analysis was used to determine the fractional area of nonvascular smooth muscle in the muscularis in histologic cross-sections of the anterior vaginal wall. RESULTS: The fractional area of nonvascular vaginal smooth muscle in the muscularis of women with prolapse was significantly decreased compared with that of control subjects. This decreased fraction of smooth muscle in the anterior vaginal wall was not related to age, race, or stage of prolapse. In women with prolapse, vaginal smooth muscle content was most diminished in specimens from postmenopausal women with no estrogen replacement. The fractional area of muscularis smooth muscle was also decreased significantly in premenopausal women with prolapse. CONCLUSION: The fraction of smooth muscle in the muscularis of the anterior vaginal wall is significantly decreased in women with pelvic organ prolapse compared with normal control subjects. PMID- 12114890 TI - Innervation of the female levator ani muscles. AB - OBJECTIVE: The objective of this study was to characterize the innervation of the human female levator ani muscles. STUDY DESIGN: Detailed dissections of the peripheral innervation of the iliococcygeal, pubococcygeal, puborectal, and coccygeal muscles were performed in 12 fresh-frozen female cadavers (aged, 32-100 years) with the use of transabdominal, gluteal, and perineal approaches. Both the pudendal nerve and the sacral nerve roots that enter the pelvis from the cephalic side were followed from their origin at the sacral foramina to their termination. Pelvic floor innervation was described with reference to fixed bony landmarks, particularly the coccyx, the ischial spine and the inferior pubis. Photographs were taken, and nerve biopsies were performed to confirm the gross findings histologically. Biopsy specimens were stained with Masson's trichrome. RESULTS: In each dissection, a nerve originated from the S3 to S5 foramina (S4 alone, 30%; from S3 and S4, 40%; from S4 and S5, 30%), crossed the superior surface of the coccygeal muscle (3.0 +/- 1.4 cm medial to the ischial spine [range, 1.0-4.2 cm]), traveled on the superior surface of the iliococcygeal muscle innervating it at its approximate midpoint, and continued on to innervate both the pubococcygeal and puborectal muscles at their approximate midpoint. The pudendal nerve originated from the S2 to S4 foramina, exited the pelvis through the greater sciatic foramen, traversed Alcock's canal, and branched to innervate the external anal sphincter, the external urethral sphincter, the perineal musculature, the clitoris, and the skin. Despite specific attempts to locate pudendal branches to the levator ani, none could be demonstrated. Nerve biopsy specimens that were obtained at gross dissection were confirmed histologically. CONCLUSION: Gross dissections suggest that the female levator ani muscle is not innervated by the pudendal nerve but rather by innervation that originates the sacral nerve roots (S3-S5) that travels on the superior surface of the pelvic floor (levator ani nerve). Because definitive studies (eg, nerve transection or neurotracer studies) cannot be performed in humans, further studies that will use appropriate animal models are necessary to confirm and extend our findings. PMID- 12114891 TI - Regulation of matrix metalloproteinase expression by estrogen in fibroblasts that are derived from the pelvic floor. AB - OBJECTIVE: The purpose of this study was to determine whether estrogen suppresses matrix metalloproteinase-2 and -9 proenzyme expression by fibroblasts that are derived from the supportive connective tissue of the pelvic floor. STUDY DESIGN: A primary fibroblast culture that was developed from a biopsy specimen of the arcus tendineus was treated with interleukin-1 beta (10-15 ng/mL), transforming growth factor-beta 1 (5-15 ng/mL), 17 beta-estradiol (10(-9)-10(-7) mol/L), and Imperial Chemical Industries (ICI) 182 780 (10(-8)-10(-6) mol/L). Cellular and extracellular protein were analyzed by Western blotting and substrate zymography, respectively, for the effect of each treatment on the amount of pro-matrix metalloproteinase-2 and -9 and the membrane type 1 matrix metalloproteinase protein. RESULTS: Both cellular and extracellular pro-matrix metalloproteinase-2 protein were increased by transforming growth factor-beta1 (P =.01) and decreased by estradiol (P <.001) and ICI 182 780 (P =.02 and.002, respectively). Membrane type 1 matrix metalloproteinase was not affected by estradiol, ICI 182 780, interleukin-1 beta, or transforming growth factor-beta 1. Extracellular pro matrix metalloproteinase-9 was increased by the cytokines interleukin-1 beta (P <.001) and transforming growth factor-beta1 (P <.001) and decreased by estradiol (P <.001) and ICI 182 780 (P <.001). CONCLUSION: The proenzymes of the tissue degrading matrix metalloproteinases -2 and -9 are decreased by 17-beta estradiol and ICI 182 780. PMID- 12114892 TI - Prevalence of comorbid psychiatric illness and its impact on symptom perception, quality of life, and functional status in women with urinary incontinence. AB - OBJECTIVE: The purpose of this study was to determine the prevalence and impact of major depression and panic disorder in women with urinary incontinence. STUDY DESIGN: Participants were 218 consecutive women with urinary incontinence over a 14-month period. Major depression and panic disorder diagnoses, symptom perception, incontinence-specific quality of life, functional status, and urinary incontinence type were assessed. RESULTS: The overall prevalence of major depression and panic disorder was 16% and 7%, respectively. In a comparison to patients with stress urinary incontinence, the odds of having major depression were 9.2 for patients with urge and 13.5 for patients with mixed urinary incontinence. Although clinically similar to patients who did not have depression, patients with depression rated their urinary incontinence as significantly more severe and had greater quality of life and functional status impairment. CONCLUSION: Current major depression and panic disorder are highly prevalent in women with urinary incontinence. Patients with urge and mixed urinary incontinence are significantly more likely to have coexistent psychiatric illness. Comorbid major depression significantly impacts a patient's urinary incontinence symptom reporting, incontinence-specific quality of life, and functional status. PMID- 12114893 TI - Patient-centered goals for pelvic floor dysfunction surgery: what is success, and is it achieved? AB - OBJECTIVE: The purpose of this study was to describe patient-identified goals for pelvic floor dysfunction surgical procedures and patient-reported achievement of those goals. STUDY DESIGN: Thirty-three consecutive patients scheduled for pelvic floor dysfunction surgical procedures completed a preoperative questionnaire on which they listed up to 5 personal goals for surgical outcomes. At 6- and 12-week follow-up, patients reported the degree (rated 1-5) to which each goal had been met (1 = strongly disagree that the goal was met; 5 = strongly agree). Age, race, vaginal parity, previous pelvic surgical procedures, pelvic floor dysfunction diagnosis, pelvic floor dysfunction surgical procedure, and perioperative complications were also recorded. Goals were categorized as being primarily related to symptom relief, increasing activity, general or other health concerns, social relationships and self-image, or physical appearance. Rates of self reported goal achievement at 6 and 12 weeks were calculated, and differences in the proportion of goals achieved by category were assessed. RESULTS: Women reported a mean of 3.6 goals; 24 of 33 women (72.7%) listed > or = 4 goals. Of the 119 goals listed, 51 goals (42.9%) dealt with urinary or bowel symptoms; 36 goals (30.3%) dealt with improving activity; 15 goals (12.6%) dealt with general health concerns; 14 goals (11.8%) dealt with social relationships and self-image, and 3 goals (2.5%) dealt with physical appearance. Twenty-seven of the 33 women (81.8%) listed symptom relief; 22 women (66.7%) listed at least 1 activity related goal; 14 women (42.4%) listed general or longer term health, and 11 women (33.3%) listed a social or self-image goal. Of the 119 goals listed, women agreed or strongly agreed that 88 goals (73.9%) were met at 6 weeks, and 101 goals (84.9%) were met at 12 weeks (chi(2) = 3.7 for difference between proportion at 6 and 12 weeks; P =.054). At 6 weeks, women agreed or strongly agreed that most goals had been met for activity, symptoms, general health, and appearance, but not for social/self-image goals (chi(2) = 24.9; P <.001). However, by 12 weeks, women agreed or strongly agreed that most goals had been met in all categories. CONCLUSION: Women who undergo pelvic floor dysfunction surgical procedures have a variety of desired subjective outcomes. Goals that relate to social roles, sexuality, and self-image may take longer to successfully achieve than other types of goals. Longer-term follow-up is crucial to determine whether initial improvements have been maintained. Assessment of patient goals is quick and easy and may help clinicians better understand and care for their patients. PMID- 12114894 TI - Fascial and muscular abnormalities in women with urethral hypermobility and anterior vaginal wall prolapse. AB - OBJECTIVE: Our purpose was to assess the structural integrity of individual elements of the urethral and anterior vaginal wall support system. STUDY DESIGN: Notes were made during retropubic operations for cystourethrocele and stress incontinence in 71 women aged 52 +/- 12.4 (SD) years. Vaginal support was assessed with the Baden-Walker system with the following average findings: urethra 1.9 +/- 0.6, bladder 1.9 +/- 1.0, apex 0.8 +/- 1.1, upper posterior wall 0.3 +/- 0.8, and rectocele 1.1 +/- 0.7. The presence of the following features was noted: paravaginal defect, integrity of the pubic and ischial attachments of the arcus tendineus fascia pelvis (ATFP), appearance of the ATFP on the sidewall, and abnormalities in the pubococcygeal muscle. RESULTS: Paravaginal defects were present in 87.3% on the left and in 88.7% on the right. Detachment of the ATFP from the pubic bone was present in 1.4% (left) and 2.8% (right). The ATFP was detached from the ischial spine in 97.6% (left) and 95.1% (right). Remnants of the ATFP were present on the sidewall in 62% (left) and 63% (right). Of these, 9% extended one fourth the distance to the spine, 21% one half the distance, 3% three fourths the distance, and 17% all the way to the spine. The pubococcygeal muscle was visibly normal in 45% (left) and 39% (right). It showed localized atrophy in 22% (left) and 30% (right) and generalized atrophy in 22.5% (left) 30.0% (right). CONCLUSION: The ATFP usually detaches from the ischial spine, but not from the pubis; slightly less than half of these women have visibly abnormal levator ani muscles. PMID- 12114895 TI - Pelvic organ support in nulliparous pregnant and nonpregnant women: a case control study. AB - OBJECTIVE: Our purpose was to compare pelvic organ support in nulliparous pregnant and nonpregnant women at a single institution. STUDY DESIGN: This was a case-control study. Pregnant patients and nonpregnant control subjects were matched according to age and race. Subjects underwent pelvic organ support evaluation by use of the pelvic organ prolapse quantification (POPQ) examination as part of routine prenatal or gynecologic care. The Pearson chi(2) statistic was used for statistical analysis, with a P value of 5% set for significance. RESULTS: A total of 21 pregnant and 21 nonpregnant nulliparous women between the ages of 18 and 29 years were included. All patients in the nonpregnant group had a POPQ stage of 0 or 1, whereas 47.6% of the pregnant subjects had POPQ stage 2 (P <.001). Individual components of the POPQ examination were compared. Significant differences were noted for points Aa and Ba, Ap and Bp, and PB and TVL. CONCLUSIONS: In nulliparous women, pregnancy is associated with increased POPQ stage compared with nonpregnant control subjects. PMID- 12114896 TI - P2X(3) receptor subunit messenger RNA expression in the female mouse bladder after oophorectomy with or without estrogen replacement. AB - OBJECTIVE: This investigation was undertaken to determine whether the sensory neuron adenosine triphosphate receptor subunit (P2X(3)) messenger RNA expression is altered in female mouse bladders after surgical oophorectomy with or without estrogen replacement. STUDY DESIGN: The mean relative concentrations of the P2X(3) receptor in 30 female mouse bladders (10 sham operated, 10 oophorectomized, and 10 oophorectomized with estrogen replacement) were determined with quantitative reverse transcription-polymerase chain reaction analysis. RESULTS: P2X(3) expression increased after surgical oophorectomy (0.91 +/- 0.16 vs 1.04 +/- 0.11, P =.048). However, P2X(3) expression after oophorectomy and immediate estrogen replacement did not differ from that of oophorectomy alone (1.11 +/- 0.15 vs 1.04 +/- 0.11, P =.206). CONCLUSIONS: The P2X(3) sensory neuron receptor messenger RNA expression is increased after oophorectomy but is not influenced by subsequent estrogen replacement. This has clinical significance because sensory neuron receptors may be associated with certain forms of bladder dysfunction. PMID- 12114897 TI - Open Burch urethropexy has a low rate of perioperative complications. AB - OBJECTIVE: Our purpose was to report perioperative complication rates with open Burch urethropexy in a tertiary care teaching center. STUDY DESIGN: One hundred fifty-one consecutive hospital and clinic charts of women who underwent Burch urethropexy for genuine stress incontinence between January 1995 and April 2000 were reviewed. Statistical analysis included the chi(2) test of association for nominal data and the Mann-Whitney test for comparison of population medians. RESULTS: All women had urodynamically proved genuine stress incontinence and 34% had detrusor instability. Most women (81%) had concomitant surgery for pelvic organ prolapse. Sixty-three percent had prior pelvic surgery with 32% having had a urethropexy, needle suspension, or anterior colporrhaphy. Perioperative complications were uncommon. The rate of lower urinary tract injury from Burch urethropexy was <1%. Two cystotomies (1.3%) were associated with surgery for prolapse, and one Burch suture (0.7%) was noted in the bladder on routine intraoperative cystoscopy. One woman (0.7%) received a single unit of blood after surgery, whereas two women had postoperative ileus. Incisional complications were most common (3%) with five cases of cellulitis and one incisional separation. One woman has prolonged voiding dysfunction and continues to perform intermittent self-catheterization 3 years after surgery. Eight percent of women had de novo detrusor instability. No women had perioperative deep venous thromboses, pulmonary emboli, adverse drug reactions, myocardial infarctions, or peripheral nerve injuries. Twenty-nine women (19%) had Burch retropubic urethropexy without concomitant surgery. There were no lower urinary tract injuries, prolonged catheterizations, or development of de novo detrusor instability in this group. The only perioperative complication was a single case of incisional cellulitis (3%). CONCLUSION: Open Burch urethropexy has a low rate of perioperative complications. The minimal morbidity of open Burch urethropexy in a teaching setting makes it the preferred teaching technique for this procedure. PMID- 12114898 TI - Differences in pelvic floor area between African American and European American women. AB - OBJECTIVE: This study tests the null hypothesis that the size of the pelvic opening spanned by the pelvic floor is the same in African American and European American women. STUDY DESIGN: Forty African American female pelvises were age matched with 40 European American female pelvises from the Hamann-Todd collection at the Cleveland Museum of Natural History. The distances between the anchoring points of the pelvic floor to the bony pelvis (pubis anteriorly, ischial spines laterally, and inferior lateral angle of the sacrum posteriorly) were measured on each half of the pelvis. Measurements from left and right halves were averaged. The cross-sectional area of the pelvic floor was calculated from these dimensions. The bi-ischial line divided the total area into anterior and posterior pelvic floor areas. Analyses taking into account differences in stature by dividing individual dimensions by height were also performed. Group differences were compared with the Student t test and the Mann-Whitney rank sum test. RESULTS: African American women had a 5.1% smaller pelvic floor area than European American women (889.6 cm(2) vs 937.0 cm(2), 5.1% P =.037). This was attributable to a 10.4% smaller posterior area (365.3 cm(2) vs 407.6 cm(2), 10.4% P =.016), whereas the anterior areas were similar (524.3 cm(2) vs 529.3 cm(2), P =.61). The following measured distances were smaller in African American women: ischial spine to inferior sacral angle (5.4 cm vs 5.9 cm, P =.016) and bi-ischial diameter (10.0 cm vs 10.6 cm, P =.004). These distances remained significant after height was controlled. CONCLUSIONS: In African American women, the posterior pelvic floor area is 10.4% smaller than in European American women, resulting in a 5.1% smaller total pelvic floor area. PMID- 12114899 TI - The standardisation of terminology of lower urinary tract function: report from the Standardisation Sub-committee of the International Continence Society. PMID- 12114900 TI - A longitudinal study of biochemical variables in women at risk of preeclampsia. AB - OBJECTIVE: The purpose of this study was to characterize gestational profiles of biochemical markers that are associated with preeclampsia in the blood of pregnant women in whom preeclampsia developed later and to compare these markers with the markers of women who were delivered of small-for-gestational-age infants without preeclampsia and with women who were at low risk for the development of preeclampsia. STUDY DESIGN: This was a prospective case control study. The subjects were women at risk of preeclampsia who were enrolled in the placebo arm of a clinical trial. Indices of antioxidant status, oxidative stress, placental and endothelial function, and serum lipid concentrations were evaluated from 20 weeks of gestation until delivery in 21 women in whom preeclampsia developed later, in 17 women without preeclampsia who were delivered of small-for gestational-age infants, and in 27 women who were at low risk for the development of preeclampsia. RESULTS: Ascorbic acid was reduced early in preeclampsia and small-for-gestational-age pregnancies. Leptin, placenta growth factor, the plasminogen activator inhibitor (PAI-1)/PAI-2 ratio, and uric acid were predictive of the development of preeclampsia. CONCLUSION: Gestational profiles of several markers were abnormal in the group with preeclampsia, and some of the markers that may prove useful in the selective prediction of preeclampsia were identified. PMID- 12114901 TI - Antimicrobial factors in the cervical mucus plug. AB - OBJECTIVE: The cervical mucus plug is positioned between the microbe-rich vagina and the normally sterile uterine cavity, which suggests a host defense function, but few relevant data are available. We analyzed the composition and antimicrobial activity of cervical mucus plugs. STUDY DESIGN: Cervical mucus plugs were collected from healthy women at delivery. Groups of plugs were randomly selected for electrolyte analysis, antimicrobial activity assays against group B Streptococcus, Escherichia coli, Candida albicans, and assays of known antimicrobial polypeptides. RESULTS: Both intact cervical mucus plugs and their aqueous extracts exhibited antimicrobial activity against aerobic microbes, in the order of potency: group B Streptococcus > E coli > C albicans. Semiquantitative Western blotting of extracts showed that secretory leukoprotease inhibitor, lysozyme, lactoferrin, and neutrophil defensins were present at concentrations that were sufficient for antimicrobial activity. CONCLUSION: The cervical mucus plug is not only a mechanical but also a chemical barrier to infection that ascends from the vagina. PMID- 12114902 TI - First magnetomyographic recordings of uterine activity with spatial-temporal information with a 151-channel sensor array. AB - OBJECTIVE: The purpose of this study was to examine the feasibility of recording the spatial-temporal magnetomyographic activity from the pregnant uterus with the use of the newly developed 151-channel noninvasive device, known as the superconducting quantum interference device array for reproductive assessment. STUDY DESIGN: Uterine magnetomyographic signals were recorded from 10 pregnant subjects with the 151-channel sensor array curved to fit the pregnant abdomen. The recording sessions were 16 minutes in length, with a sampling rate of 250 Hz. RESULTS: Uterine activity bursts were successfully recorded with the superconducting quantum interference device array for reproductive assessment system. By obtaining a contour plot of the magnetic field distribution, we were able to localize the areas of activation over the uterus during a contraction. Also, it was possible to calculate the time delay in the propagation of the activity across the uterus. CONCLUSION: Using superconducting quantum interference device array for reproductive assessment system, we have established the feasibility of recording uterine contractile activity with spatial-temporal resolution that is high enough to determine the regions of localized activation and propagation over the uterus. PMID- 12114903 TI - Pregnancy in women who undergo long-term hemodialysis. AB - OBJECTIVE: Pregnancy is rare in women who require long-term hemodialysis, and pregnancy outcome with a live birth has a low success rate. The purpose of this study was to describe the treatment of pregnancy and the outcome in a series of patients undergoing long-term hemodialysis treatment. STUDY DESIGN: A total of 15 women who were undergoing long-term hemodialysis treatment who had 18 pregnancies during the period from 1990 to 2000 were included in this study. All the women had been undergoing hemodialysis for a mean of 5.3 years before pregnancy, except for one woman who began hemodialysis at 16 weeks of gestation. When conception was confirmed, the risks of pregnancy were explained to the couple and to the medical team. Almost all hemodialysis were performed with a high-flux dialyzer and with volume-controlled ultrafiltration. During pregnancy the hemodialysis schedule was increased to 4 hours 4 to 6 times weekly, with a blood flow rate of 250 to 300 mL/min and a dialysate flow rate of 500 to 600 mL/min. Data on changes in dialysis regimen, biochemistry, blood pressure control, use of erythropoietin, medical complications, obstetric regimen, and perinatal problems were collected. RESULTS: Elective abortion was performed in 5 of the 18 pregnancies. Thirteen pregnancies were treated, with 12 live births, of which 9 infants survived. There was 1 intrauterine fetal death and 3 neonatal deaths. No fetal anomaly was detected. The mean gestational age at delivery was 32 weeks (range, 23-36 weeks). The mean newborn weight was 1542 g (range, 512-1660 g), with intrauterine growth restriction in 7 of the 9 cases. Anemia was treated with recombinant human erythropoietin and/or transfusion in all cases. Of the 15 women undergoing hemodialysis treatment, elevated blood pressure was complicated in 13 pregnancies, in which 7 were treated with antihypertensive drugs. Polyhydramnios occurred in 6 of 9 surviving live births and was partially relieved after hemodialysis. Cesarean delivery was performed in 6 of 13 deliveries. All of the women recovered after delivery to their prepregnancy dialysis therapy levels. CONCLUSION: Although pregnancy remains risky in women who are undergoing long term dialysis, advances in dialysis, obstetrics, and neonatal treatment have led to an improved success rate. Our data from 1990 to 2000 showed a 60% success rate. PMID- 12114904 TI - Polymorphism in the interleukin-1 gene complex and spontaneous preterm delivery. AB - OBJECTIVE: We examined the association between preterm delivery and polymorphisms at position +3953 of the interleukin-1 beta gene (IL1B+3953) and in intron 2 of the interleukin-1 receptor antagonist gene (IL1RN). STUDY DESIGN: This was a case control study that involved 52 pregnancies that resulted in spontaneous preterm delivery before 34 weeks of gestation and 197 pregnancies that resulted in birth at term. Polymorphisms were determined by polymerase chain reaction and restriction fragment length polymorphism analysis. RESULTS: Homozygous carriage of IL1B+3953 allele 1 by fetuses of African descent was associated with a risk of preterm delivery (P =.033). Fetuses of Hispanic descent that carried IL1RN allele 2 were found to be at an increased risk for preterm premature rupture of membranes and subsequent preterm delivery(P =.021; odds ratio, 6.5; 95% CI, 1.25 37.7). CONCLUSION: There are associations of spontaneous preterm delivery with the fetal carriage of IL1B+3953*1 and IL1RN*2 alleles in African and Hispanic populations, respectively. PMID- 12114905 TI - Prolonged mild fetal hypoxia up-regulates type I nitric oxide synthase expression in discrete areas of the late-gestation fetal sheep brain. AB - OBJECTIVE: Our purpose was to study the effects of prolonged mild hypoxemia on type I nitric oxide synthase (NOS) messenger RNA, protein, and enzymatic activity in the fetal sheep brain. STUDY DESIGN: Pregnant sheep were randomly allocated to receive maternal nitrogen (n = 8) or compressed air (controls, n = 5) to reduce fetal brachial artery PO(2) by 25% for 5 days. Type I NOS mRNA (determined by ribonuclease protection assay) protein (determined by Western blot) and enzymatic activity (determined by citrulline assay) were measured in the hippocampus, striatum, cerebellum, and frontal cortex. Data are presented as mean +/- SEM and were compared by means of one-way analysis of variance or two-sample t test. RESULTS: The reduction in maternal inspired oxygen concentration decreased fetal PO(2) by 26% and oxygen content by 25% without an associated change in PCO(2) or pH. Fetal hypoxemia increased type I NOS mRNA by threefold in the striatum and by 2-fold in the frontal cortex and cerebellum, but it did not change mRNA expression in the hippocampus (P <.05). Type I NOS protein and catalytic activity increased only in the striatum (P <.05). CONCLUSION: Prolonged mild hypoxemia has a differential effect on type I NOS mRNA in fetal sheep brain areas. Type I NOS protein and catalytic activity significantly increased only in the striatum. Our data suggest that fetal type I NOS gene expression is regulated at transcriptional, post-transcriptional, and translational levels. PMID- 12114906 TI - Neurodevelopmental outcome of premature infants after antenatal phenobarbital exposure. AB - OBJECTIVE: We previously demonstrated that antenatal phenobarbital does not decrease the risk of intracranial hemorrhage or early death in premature infants. The objective of the present study was to evaluate the impact of antenatal phenobarbital exposure on the neurodevelopmental outcome of premature infants born to women who were participating in the randomized clinical trial of antenatal phenobarbital exposure. STUDY DESIGN: Infants were evaluated at 18 to 22 months corrected age with a standard neurologic examination and the Bayley scales of infant development measuring the mental developmental index and the psychomotor developmental index. RESULTS: Of the 578 infants <34 weeks of gestational age who were born to women who were enrolled in the primary study, 7 infants died after discharge from the neonatal intensive care unit, and 135 infants were lost to follow-up. Infants who were lost to follow-up had a higher mean birth weight and gestational age and a lower maternal education, but the rates of intracranial hemorrhage were comparable to those infants who were evaluated. Among the infants who were evaluated (n = 436; 76%), the mean birth weight and gestational age, maternal education, and frequency and distribution of intracranial hemorrhage were similar in the antenatal phenobarbital exposed and placebo groups. Eighteen infants (8%) in the antenatal phenobarbital exposed group and 21 infants (11%) in the placebo group had cerebral palsy (P = not significant). There was no difference between the 2 groups in either the median Bayley II mental developmental index (85 in the antenatal phenobarbital and 86 in the placebo group) or the Psychomotor Developmental Index (91 in the antenatal phenobarbital and 91 in the placebo group). Infants with intracranial hemorrhage (23%) had significantly lower mental developmental index and psychomotor developmental index scores than infants with no intracranial hemorrhage, independent of antenatal phenobarbital exposure. In the total cohort of 436 infants, the presence of intracranial hemorrhage or periventricular leukomalacia was associated with lower mental developmental index and psychomotor developmental index scores; the presence of increasing birth weight, maternal education, and a complete course of antenatal steroids was associated with a higher mental developmental index score. CONCLUSION: Antenatal phenobarbital exposure did not favorably or adversely affect the neurodevelopmental outcome of premature infants at 18 to 22 months of age. PMID- 12114907 TI - Fetal umbilical vascular response to chronic reductions in uteroplacental blood flow in late-term sheep. AB - OBJECTIVE: The present study was designed to determine the effects of chronic reduction in uterine blood flow (UBF) on umbilical blood flow (UmbBF) and fetal cardiovascular hemodynamics and oxygenation. STUDY DESIGN: Sixteen sheep with singleton pregnancies were instrumented on gestational day (GD) 110; an externally adjustable vascular occluder was placed on the common internal iliac artery. UBF in control animals rose from 867 +/- 61 mL/min on GD 115 to 1520 +/- 158 mL/min (n = 8) on GD 138, whereas UBF in restricted animals was maintained at 750 +/- 50 mL/min (n = 8). RESULTS: UmbBF in control animals increased from 472 +/- 25 mL/min to 744 +/- 58 mL/min over the study period from GD 115 to GD 138. This was associated with normal gestational increases in fetal arterial pressure and significant reductions in calculated umbilical vascular resistance. Although restricted animals initially had a similar UmbBF on GD 115, UmbBF rose only to 545 +/- 43 mL/min over the study period (control vs restricted, P <.008). Although fetal arterial pressure showed normal gestational changes, umbilical vascular resistance failed to decrease over gestation in restricted animals as it did in control animals. Fetal heart rate and oxygenation showed normal changes in both groups. CONCLUSION: Chronic reduction in UBF prevents umbilical vascular resistance from undergoing normal gestational decreases, leading to significantly lower UmbBF. This altered umbilical perfusion pattern would be expected to significantly affect fetal delivery of oxygen and nutrients and ultimately fetal growth. PMID- 12114908 TI - The pharmacokinetics of glyceryl trinitrate with the use of the in vitro term human placental perfusion setup. AB - OBJECTIVE: The purpose of this study was to determine the pharmacokinetics of glyceryl trinitrate across the in vitro term human perfused placenta. STUDY DESIGN: Peripheral placental lobules (n = 6) were dually perfused. The maternal side was perfused with glyceryl trinitrate (100 nmol/L) for 90 minutes. Serial samples from the fetal venous and maternal venous catheters were collected and assayed for glyceryl trinitrate and its vasoactive metabolites (1,2- and 1,3 glyceryl dinitrate) with gas chromatography. Fetal arterial perfusion pressure was continuously measured throughout. Data are expressed as the mean +/- SEM, with 1-way analysis of variance followed by a Newman-Keuls post-hoc test (P <.05). RESULTS: The mean steady-state fetal venous:maternal side ratio of glyceryl trinitrate concentration was 18.5% +/- 3.7%. The 1,2-glyceryl dinitrate and 1,3-glyceryl dinitrate levels in the fetal venous samples and maternal venous samples were less than the lower end of the sensitivity range of the assay (<5 nmol/L). Fetal arterial perfusion pressure did not change with glyceryl trinitrate administration. CONCLUSION: The glyceryl trinitrate in the fetal venous sample was approximately 18.5% of the glyceryl trinitrate in the maternal side in this preparation. The presence of glyceryl dinitrates in the maternal venous samples and the fetal venous samples indicates the capacity for placental biotransformation of glyceryl trinitrate. The data demonstrate that glyceryl trinitrate, at a tocolytic concentration, has the ability to cross the placenta but was not found to affect fetal arterial perfusion pressure. PMID- 12114909 TI - Chronic in utero plasma hyperosmolality alters hypothalamic arginine vasopressin synthesis and pituitary arginine vasopressin content in newborn lambs. AB - OBJECTIVE: Arginine vasopressin is synthesized in the hypothalamus and secreted by the posterior pituitary gland in response to plasma hypertonicity. Previous studies suggest that in utero and neonatal exposure of rat pups to prolonged alterations of plasma osmolality may permanently alter (imprint) arginine vasopressin synthesis and secretion, thus adult responses to osmotic challenges. Little is known, however, of the potential for imprinting of neuroendocrinologic systems in precocial species. In view of the frequent occurrence of altered maternal and fetal plasma tonicity (eg, maternal dehydration, hyperemesis), we sought to determine the effect of prolonged maternal hypertonicity on arginine vasopressin gene expression and pituitary gland content in neonatal sheep. STUDY DESIGN: Pregnant ewes at 119 +/- 3 days of gestation were water restricted to achieve and maintain plasma hypertonicity (10-20 mOsm/kg above baseline level) until normal term delivery. Newborns were provided maternal nursing ad libitum. Within 24 hours after birth, study neonatal lambs (n = 6) and age-matched control neonatal lambs (n = 5) were killed, and the pituitary gland and hypothalamus were removed and frozen immediately. Pituitary arginine vasopressin content was determined by radioimmunoassay, and hypothalamic arginine vasopressin gene expression was quantified with Northern blot. Differences in pituitary arginine vasopressin content and hypothalamic arginine vasopressin gene expression (arginine vasopressin/ beta-actin ratio) between study and control newborn lambs were analyzed by unpaired t test. RESULTS: In response to maternal water restriction, maternal plasma osmolality increased from 307 +/- 0.9 mOsm/kg to 325 +/- 1.3 mOsm/kg, and plasma sodium increased from 147 +/- 1.3 mEq/L to 156 +/- 1.2 mEq/L. The maternal plasma hyperosmolality and hypernatremia were maintained until normal term delivery. At the time of death, study (in utero dehydrated) lambs had higher plasma sodium (150 +/- 0.4 mEq/L vs 146.5 +/- 1.5 mEq/L; P <.05) and chloride (112.8 +/- 1.0 mEq/L vs 108.5 +/- 1.5 mEq/L; P <.05) levels, and lower potassium (4.5 +/- 0.2 mEq/L vs 5.5 +/- 0.3 mEq/L; P <.05) concentrations than control newborn lambs. Both newborn groups had similar plasma osmolality (320.0 +/- 1.3 mOsm/kg vs 318.0 +/- 3.4 mOsm/kg). Total pituitary arginine vasopressin content was significantly greater in the study than in the control newborn lambs (8.3 +/- 2.8 microg vs 1.6 +/- 1.3 microg; P <.05). Conversely, hypothalamic arginine vasopressin messenger RNA levels were lower in the study newborn lambs than in the control newborn lambs (arginine vasopressin/beta-actin ratio: 0.29 +/- 0.01 vs 0.68 +/- 0.15; P <.05). CONCLUSION: Despite the presence of plasma hypernatremia, prolonged elevation of fetal plasma tonicity increases newborn pituitary arginine vasopressin content yet decreases hypothalamic arginine vasopressin gene expression. The present study suggests that prolonged prenatal exposure to plasma hypertonicity may imprint the hypothalamic-pituitary arginine vasopressin regulatory system. PMID- 12114911 TI - Regulation of matrix metalloproteinases (type IV collagenases) and their inhibitors in the virgin, timed pregnant, and postpartum rat uterus and cervix by prostaglandin E(2)-cyclic adenosine monophosphate. AB - OBJECTIVE: Our purpose was to examine whether the type IV collagenases (metalloproteinase [MMP]-2 and MMP-9) and their inhibitors (TIMP-1 and TIMP-2) are regulated by a prostaglandin E(2) (PGE(2))-cyclic adenosine monophosphate (cAMP) mechanism in nonpregnant virgin, preterm and term pregnant and postpartum rats. STUDY DESIGN: Sprague-Dawley rats were infused with either saline solution or PGE(2) over 24 hours or were noninfused. Plasma and tissue were analyzed for cAMP, MMP-2 and MMP-9, and TIMP-1 and TIMP-2. RESULTS: PGE(2) evoked elevations in plasma and tissue levels of cAMP and MMP-2 in the preterm and term pregnant rats. MMP-9 levels were elevated in the preterm plasma and uterus, whereas in the term pregnant and postpartum rats, MMP-9 levels were increased in the cervix. CONCLUSIONS: MMP levels in the pregnant and postpartum rat uterus and cervix are regulated in part by a PGE(2)-cAMP mechanism. Specifically, MMP-9 may be involved in preterm labor, cervical maturity, and involution of the postpartum uterus. PMID- 12114912 TI - Inhibition of system A amino acid transport activity by ethanol in BeWo choriocarcinoma cells. AB - OBJECTIVE: Our purpose was to investigate the influence of ethanol on system A amino acid transporter in BeWo cells. STUDY DESIGN: BeWo cells were cultured in the absence or presence of ethanol. The function of system A was monitored by the transport of alpha-(methylamino)isobutyric acid. Messenger RNA levels for system A were assessed by Northern analysis. RESULTS: Treatment of BeWo cells with ethanol reduced the activity of system A. The effect was dose and treatment time dependent. The decrease in system A activity was 38% +/- 3% at 0.75% ethanol with a 16-hour treatment time. The activities of several other transporters tested were not affected. The effect on system A activity was associated with a decrease in the maximal velocity of the transport system without affecting the substrate affinity. Ethanol did not alter the messenger RNA levels for system A. CONCLUSION: Exposure of BeWo cells to ethanol significantly reduces the function of system A. This finding has potential implications that may be relevant to the pathogenesis of the fetal alcohol syndrome. PMID- 12114913 TI - False-positive serum human chorionic gonadotropin results: causes, characteristics, and recognition. AB - False-positive serum human chorionic gonadotropin results are estimated to occur in 1 in 10(3) to 1 in 10(4) tests. Most of these false-positive results are due to interference by non-human chorionic gonadotropin substances (especially human luteinizing hormone and anti-animal immunoglobulin antibodies) and the detection of pituitary human chorionic gonadotropin. The false-positive human chorionic gonadotropin measurements are characterized by serum levels that are generally <1000 mIU/mL, the absence of human chorionic gonadotropin in the urine, nonparallelism of the human chorionic gonadotropin levels in serial dilutions of the serum with the human chorionic gonadotropin standard, results that are not consistent with the clinical or operative findings, the absence of a substantial change in the serum levels over time or after therapy, and the finding of a negative result when an alternative type of human chorionic gonadotropin assay is used. An algorithm is presented to aid in the recognition of false-positive human chorionic gonadotropin results in patients. PMID- 12114910 TI - A novel method for the detection of Down syndrome with the use of four serum markers. AB - OBJECTIVE: The purpose of this study was to evaluate the use of the Lens culinaris agglutinin-reactive alpha-fetoprotein ratio for the detection of fetal Down syndrome in combination with traditional serum markers. STUDY DESIGN: We obtained maternal serum from 530 women with unaffected pregnancies and 31 women who were pregnant with a fetus with Down syndrome at 14 to 20 weeks of gestation. Various combinations of Lens culinaris agglutinin-reactive alpha-fetoprotein ratio, alpha-fetoprotein, human chorionic gonadotropin, and unconjugated estriol were evaluated by score, without regard for the maternal age-related fetal Down syndrome risk. RESULTS: The best combination of serum markers, according to our scoring method, was a combination of all 4 markers. This combination showed a sensitivity of 83.9%, with a 5.1% false-positive rate for Down syndrome. Moreover, in older women, high sensitivity was obtained without increasing the false-positive rate. CONCLUSION: The screen using all 4 markers showed high sensitivity in all age groups without increasing the false-positive rate. PMID- 12114914 TI - Metabolic treatment of pregnancy and postdelivery period in a patient with cobalamin A disease. AB - OBJECTIVE: We report the successful treatment of a woman with Cobalamin A disease with hydroxycobalamin injections during pregnancy and delivery and 3 months after delivery. Urine and plasma methylmalonic acid levels served to adjust therapy before and after delivery. The mother had no untoward metabolic complications and gave birth to a normal baby. PMID- 12114915 TI - Treatment of a case of primary retroperitoneal mucinous cystadenocarcinoma: is adjuvant hysterectomy and bilateral salpingo-oophorectomy justified? AB - OBJECTIVE: We present a case of primary retroperitoneal mucinous cystadenocarcinoma in a 38-year-old woman. STUDY DESIGN: The tumor was resected with a segment of adjacent descending colon. Five years after the operation, the patient is well, without evidence of recurring disease, based on clinical investigation and modern imaging techniques. RESULTS: In the light of the literature, it appears most likely that this rare tumor is caused by coelomic metaplasia. On the basis of the histopathologic findings in our case and the reports from the literature, we recommend radical tumor excision en bloc with all infiltrated adjacent structures. CONCLUSION: Added removal of unaffected uterus and adnexes makes young women infertile and climacteric and is not yet validated by long-term results. PMID- 12114916 TI - Uncomplicated erosion of polytetrafluoroethylene grafts into the rectum. AB - Synthetic materials are frequently used in gynecologic surgical procedures to repair pelvic floor hernias and prolapse and to form barriers to adhesion formation. Mesh erosion into the vagina and lower urinary tract are known complications. We report 2 cases of polytetrafluoroethylene mesh erosion into the rectum. PMID- 12114917 TI - Effects of sublingual nitroglycerin on human uterine contractility during the active phase of labor. AB - Nitroglycerin is administered intravenously in acute obstetric emergencies to relax the uterus. However, complications (eg, hypotension, acute uterine bleeding) are frequent, which prompted a search for alternative routes of administration. We hypothesized that the sublingual administration of nitroglycerin would reduce uterine tone and contractility with few complications. Intrauterine pressure was measured in 12 women who were actively laboring (>4 cm dilatation, regular contractions) with epidural analgesia and who were alert and responsive throughout the study. In a double-blind fashion, subjects were randomized to receive either placebo or sublingual nitroglycerin (3 doses, 800 microg each) 10 minutes apart. The obstetric anesthesiologist continuously monitored maternal blood pressure and fetal heart rate. Cervical dilatation was assessed at the beginning and the end of the protocol. The area under the intrauterine pressure curve (integral) was used to estimate uterine contractility. Intrauterine pressure was analyzed before the randomization code was broken. Nitroglycerin did not alter the intrauterine pressure integral after the first dose (placebo, 3147 mm Hg x s [95% CI, 2206-4088] vs nitroglycerin, 4146 mm Hg x s [95% CI, 2451-5841]; P =.22), second dose (placebo, 3123 mm Hg x s [95% CI, 2447-3799] vs nitroglycerin, 3611 mm Hg x s [95% CI, 2723-4499]; P =.28), or third dose (placebo, 3303 mm Hg x s [95% CI, 2616-3990] vs nitroglycerin, 3810 mm Hg x s [95% CI, 2306-5314]; P =.45). Cervical dilation, basal uterine tone, duration and frequency of uterine contractions, or fetal heart rhythm remained unaffected. Maternal mean arterial pressure decreased significantly after nitroglycerin was administered. All women were delivered vaginally without intervention. Three doses of sublingual nitroglycerin (800 microg per dose) reduce neither uterine activity nor tone, despite lowering maternal blood pressure. If a clinical option, sublingual nitroglycerin will require a higher dose, which would place mother and fetus at risk for complication. PMID- 12114918 TI - Nonbilharzial bladder carcinoma complicating pregnancy--treatment with bacille Calmette-Guerin. AB - Bladder carcinoma in situ is often treated with intravesical bacille Calmette Guerin. Unable to find a report of this therapy in pregnancy, we describe such a case. Intravesical bacille Calmette-Guerin immunotherapy may be safely administered during the second trimester in an appropriately selected patient. PMID- 12114920 TI - Postpartum hemorrhage with concurrent massive inferior epigastric artery bleeding after cesarean delivery. AB - Postpartum hemorrhage with concurrent massive inferior epigastric artery bleeding after cesarean delivery is unusual. In the current case, besides embolization of the bilateral hypogastric arteries, recognition and successful embolization of a clinically overlooked active bleeding source in the abdominal wall was crucial to effecting a cure. PMID- 12114919 TI - Decreasing the plasma triglyceride level in hypertriglyceridemia-induced pancreatitis in pregnancy: a case report. AB - We report a case of hypertriglyceridemia-induced pancreatitis in pregnancy in which continuous intravenous heparin was used effectively and safely to lower the plasma triglyceride level within 24 hours. PMID- 12114921 TI - Case report and review of the perinatal implications of maternal lithium use. AB - The purpose of this study was to review the use of lithium in pregnancy and its effects on the neonate. This was a case study and review of the published literature. Lithium is commonly used in the treatment of psychiatric disorders, specifically bipolar depression. Bipolar disorders that require treatment with lithium demand special consideration when the woman becomes pregnant. Reported neonatal problems with maternal lithium therapy include Ebstein's anomaly, poor respiratory effort and cyanosis, rhythm disturbances, nephrogenic diabetes insipidus, thyroid dysfunction, hypoglycemia, hypotonia and lethargy, hyperbilirubinemia, and large-for-gestational-age infants. Lithium can have adverse effects on the fetus and newborn infant, but data suggest normal behavioral patterns in childhood. PMID- 12114922 TI - Doppler ultrasonographic evaluation of twins discordant for Rh alloimmunization. AB - Minimally invasive methods have been used successfully to diagnose and treat maternal red blood cell alloimmunization. We present a case of a dizygotic twin pregnancy discordant for the Rh allele. Serial Doppler ultrasonography helped to diagnose the fetus that was at-risk for anemia caused by Rh alloimmunization and to direct treatment. PMID- 12114923 TI - Multiple foreign body erosions after laparoscopic colposuspension with mesh. AB - Laparoscopic colposuspension with mesh and tackers is gaining popularity for the treatment of stress incontinence because of shorter operator learning curves and increased scarring and fibrosis with surgical success rates comparable to suturing techniques. We report two cases of multiple foreign body erosions requiring reoperation after this technique. PMID- 12114924 TI - The prophylactic forceps operation. 1920. PMID- 12114925 TI - Predicting ambulation by checking leg withdrawal in fetuses with spina bifida. PMID- 12114926 TI - Neurobiology of diurnal periodicity for fetal heart rate variability. PMID- 12114927 TI - Defining the relationship between obstetricians and maternal-fetal medicine specialists. PMID- 12114928 TI - PTEN mutation analysis in uterine leiomyosarcomas and mixed mullerian sarcomas. PMID- 12114929 TI - Selective photocoagulation of monochorionric twin pregnancy. PMID- 12114930 TI - Patients with severe eclampsia benefit from long-term epidural analgesia. PMID- 12114931 TI - A light in the distance. PMID- 12114932 TI - Clean versus sterile technique. PMID- 12114933 TI - Pouchitis: part 2: treatment options and their effectiveness. PMID- 12114934 TI - Historical research and WOC nursing: a strange and wonderful relationship. PMID- 12114935 TI - Sexual counseling by the ET nurse: if not you, then who? PMID- 12114936 TI - Current home care expenditures for persons with leg ulcers. AB - OBJECTIVE: The purpose of this study was to gain a better understanding of the home care expenditures incurred in providing care to the population with leg ulcers. DESIGN: The study was designed as a descriptive survey and was conducted over a 4-week period during March 1999. SETTING AND SUBJECTS: Persons in a large Ontario urban center with an ulcer below the knee, including the foot, who were receiving nursing services in the home, were eligible for inclusion in the study. INSTRUMENTS: A leg assessment tool, a supply usage form, and a visiting nurses log (all developed by the researchers for the study) were used to collect data. METHODS: Home care nurses visited all clients and completed an in-depth assessment of their social, medical, and leg ulcer history. Legs were inspected, an ankle brachial pressure index score was determined, and ulcers were examined and measured. For each nursing visit, supply usage, travel and treatment times, and mileage were tracked. RESULTS: During the study period, 2270 visits were made (mean treatment time = 26 minutes, mean travel time = 17 minutes) costing $80.62 (Canadian dollars). Supply costs were $21.06. The regional annual home care expenditures were conservatively estimated to be $1.3 million. CONCLUSION: Costs could potentially be reduced by cutting the 40% visit time attributed to travel, decreasing the visit frequency to clients with minimal drainage, and attention to "best practice." PMID- 12114937 TI - The rectal trumpet: use of a nasopharyngeal airway to contain fecal incontinence in critically ill patients. AB - OBJECTIVE: Our objective was to determine if a nasopharyngeal airway (rectal trumpet) could be used as a fecal containment device with less trauma than traditional devices, such as a fecal incontinence pouch or balloon rectal catheter. DESIGN: A single-subject clinical series was used. SETTING AND SUBJECTS: A nonrandom sample of critically ill adult and geriatric patients (n = 22) with ongoing fecal incontinence who were receiving care in an intensive care and intermediate care unit in a university teaching hospital was used. INSTRUMENTS: Direct observation, medical record review, a questionnaire, and interviews were used. METHODS: The bedside nurses identified patients as study candidates. Clinical findings were documented in the medical record. The nurses providing patient care completed questionnaires. MAIN OUTCOME MEASURES: Main outcome measures were parameters related to efficacy, practicality, and complications of use of the rectal trumpet: stool containment, skin and anal sphincter integrity, patient comfort, and ease of insertion. RESULTS: All 22 patients (100%) had containment or improved containment of stool. Observable healing or restoration of skin integrity occurred in 90% of the patients with acquired skin injury (n = 20). None of the patients suffered any change in tone or damage to the anal sphincter. Although 41% of the patients experienced discomfort with insertion of the rectal trumpet, 86% had no discomfort while it was maintained in position. Insertion of the rectal trumpet was rated as easy by 84% of the responding nurses (n = 63). CONCLUSIONS: Use of a rectal trumpet was well tolerated by patients and practical for nurses. Incontinence was contained and no untoward effects were noted. Benefits to the patient included wound healing and improved comfort. PMID- 12114938 TI - Irritable bowel syndrome: more than a gut feeling. AB - Irritable bowel syndrome (IBS) is a common functional bowel disorder that affects approximately 8% to 20% of the populations of industrialized countries. In most countries the majority of patients seeking health care services for IBS are women. During the past 10 years there has been a remarkable growth in both basic and clinical research on IBS. At the same time, new drugs are currently under investigation for their efficacy in the management of IBS. This review is intended to cover the current research related to the etiology and pathophysiology of IBS, as well as the diagnosis and management of this condition. Because nurses are frequently at the forefront of assessing and managing patients with this condition, nursing care issues are addressed. PMID- 12114939 TI - Patient with compartment syndrome of the lower extremity. PMID- 12114941 TI - Dr Isaac Hays and Dr Isaac Israel Hayes: their adventures in two worlds. PMID- 12114942 TI - Joseph Rogers (1820-1889). PMID- 12114943 TI - Charles Meller and John Kirk: medical practitioners and practice on Livingstone's Zambesi expedition, 1858-64. PMID- 12114944 TI - Sir John Parkinson (1885-1976). PMID- 12114945 TI - William Mestrezat (1883-1928): oenologist, physician, neurochemist. PMID- 12114946 TI - E V McCollum: the man. PMID- 12114947 TI - Alexander Gordon of Aberdeen and the discovery of the contagiousness of puerperal fever. PMID- 12114948 TI - George Johnson (1818-1896). PMID- 12114949 TI - Dr A S Thomson: ethnographer and historian. PMID- 12114950 TI - Hua Tuo, the Chinese god of surgery. PMID- 12114951 TI - Did Ramanujan have Asperger's disorder or Asperger's syndrome? PMID- 12114952 TI - Professor Hugo Carl Plaut: pioneer bacteriologist and mycologist. PMID- 12114953 TI - The evolution of medical science: a biographical analysis. PMID- 12114954 TI - The Barons Larrey: Dominique Jean (1766-1842); Hippolyte (1808-1895). PMID- 12114956 TI - A Novel Myotoxin from the Venom of Trimeresurus mucrosquamatus. AB - A novel myotoxin, designated TMPB, was purified from the venom of Trimeresurus mucrosquamatus by Sephadex G-100 superfine gel chromatography and fast protein liquid chromatography (FPLC). The N-terminal sequence of 24 amino acid residues was determined by protein sequencer. The sequence similarities between TMPB and other two phospholipase A(2) (PLA2s) previously purified from the same venom were 41.7% and 54.2%, respectively, but TMPB showed no detectable PLA(2) hydrolytic activity. Its molecular weight was estimated to be 16 000 by reducing SDS-PAGE and isoelectric point was determined to be 9.2 by isoelectric focusing electrophoresis. TMPB exhibited strong myotoxicity and platelet aggregation inhibiting activity, and the two activities could all be inhibited by heparin. PMID- 12114957 TI - Investigation of Protein-DNA Interactions in Enhancer II and Core Promoter of HBV by in vivo Footprinting. AB - The protein-DNA interactions in the enhancer II (ENII) and core promoter regions of hepatitis B virus (HBV) were investigated by the ligation-mediated polymerase chain reaction (LMPCR) in vivo footprinting in HepG2.2.15 cell line. Major footprints are evident at the enhancer II and core promoter regions. In particular, proteins appear to interact with the sites for Sp1, C/EBP, HNF4, HNF3, HNF1 and hB1F, as well as the TATA-like element. It confirmed that these protein-DNA interactions defined in vitro play important roles in the regulation of 3.5 kb RNAs transcription of HBV in vivo. Notably, three novel sites nt 1774 1789, nt 1801-1818 and nt 1835-1843 were found to be protected or hypersensitive, indicating that they were also involved in protein-DNA interactions in vivo. Functional analysis using CAT reporter gene demonstrated that the site from nt 1801 to 1818 was important for the function of HBV core promoter. PMID- 12114958 TI - A Novel System for Hyper Expression and Rapid Purification of Arginyl-tRNA Synthetase from Escherichia coli. AB - The gene argS encoding arginyl-tRNA synthetase (ArgRS) in Escherichia coli has been cloned into the vector pMFT7-5 which allows overproduction of ArgRS. The specific activity of the synthetase in the crude extract of E.coli JM109(DE3) transformant containing the plasmid pMFT7-argS was approximately 2 500 times that of JM109(DE3) (the host strain without the vector). The overproduced synthetase can be purified to homogeneity with the specific activity of 36 000 units/mg under native condition within one day by a two-step purification system of DEAE Sepharose CL-6B Fast Flow and Blue Sepharose CL-6B chromatographies with an overall yield of 69%. Moreover, this system makes it very easy to incorporate efficiently the expensive labeled amino acids into this enzyme. This novel system can conveniently provide a large amount of purified enzyme with a much higher specific activity than before which may be beneficial to NMR and crystallographic studies on this enzyme. PMID- 12114959 TI - Expression and Biochemical Characterization of Acidic Phospholipase A(2)I from Agkistrodon acutus. AB - A cDNA encoding acidic phospholipase A(2)I(A.aAPLA(2)I)from Agkistrodon acutus was inserted into a bacterial expression vector and effectively expressed in E.coli RR1. The protein was produced as insoluble inclusion bodies. After partial purification by washing the inclusion bodies with Triton X-100, denaturing and refolding, the renatured recombinant protein was purified by FPLC column Superose(TM)12. The enzymatic acti-vity and platelet aggregation inhibiting effect of the expressed A.aAPLA(2)I is close to those of denatured-refolded native acidic PLA(2) from Agkistrodon halys Pallas, and has the same hemolytic activity as denatured-refolded basic phospholipase A(2) from Agkistrodon halys Pallas. The roles of various amino acid residues in the enzymatic activity and pharmacological activities of phospholipase A2 are discussed. PMID- 12114960 TI - SELEX Screening and Characterization of Small RNA Molecules That Specifically Bind the Reactive Blue Dye. AB - A 73-base single-stranded DNA library with a T7 promoter was chemically synthesized, in which 20 consecutive bases were completely randomized. After PCR amplification and T7 RNA polymerase transcription of the DNA random library, the in vitro transcribed RNA random library was subjected to 8 successive rounds of reactive blue dye column selection by SELEX method, resulting in a sharp increase of the percentage of the transcribed RNA pool that could bind reactive blue dye, from less than 0.03% in the first round pool to 22.4% in the eighth-round pool. Cloning and sequencing of the enriched RNAs pool led to three groups of RNA molecules in terms of molecular lengths. The correlation between the binding capacities and their corresponding secondary structures of the screened RNAs suggests that the RNA helix is the main motif in the interaction between RNAs and reactive blue dye. These results demonstrate that it is possible to find RNA candidates with a given function in a shorter RNA random library so that it is easier to follow the structure and function relationship. In addition, longer RNA sequences could theoretically be derived from short RNA sequences through a process of motif combination in the course of molecular evolution. PMID- 12114961 TI - Cloning of a Gene in Rat Brain Induced to Be Expressed Differently by AVP(4-8) with Differential Display PCR. AB - As a metabolite of arginine-vasopressin, AVP(4-8) has been shown to have potent memory-enhancing activity and induce a series of physiological and biochemical events in rat brain. The technique known as differential display polymerase chain reaction(DD-PCR)was applied to explore genes induced to be expressed differ-ently by AVP(4-8) in rat hippocampus. Nine different primer pairs were used to perform DD-PCR after reverse transcription, and more than ten differential display fragments were observed. The most remarkable different fragment, dd1, was further cloned and sequenced. Homology searching in Genebank showed that dd1 may be a new gene. Moreover, the results from Northern blot confirmed that dd1 is indeed a gene induced by AVP(4-8). PMID- 12114962 TI - Construction and Expression of hTNFalpha-hTGF3 Fused Gene. AB - Human tumor necrosis factor was fused with hTGF3, the third loop region of hTGFalpha specifically binding to EGF receptor, through a flexible linker by recombinant DNA techniques. The constructed plasmid containing hTNFalpha-hTGF3 fused gene, pSB92-TLT, was expressed under the control of P(L) promoter and after the induction at 42 degrees it gave a high level of expression of the fused gene, ranging in 50%-60% of total bacterial protein. 95% of the expressed product was inclusion body, and only 5% was soluble. When the fused gene was constructed into the constitutive plasmid pLC, a new plasmid controlled by P(L) promoter but without cIts857, and expressed at 30 degrees, the expression yield reached 50% and a high proportion of soluble protein, 35% of total bacterial protein, was obtained. Western blot analysis indicated that the fusion protein could specifically bind to anti-hTNFalpha antibody. Comparing to original hTNFalpha, the fused hTNFalpha derivative showed about equal cytotoxicity to L929 cell line and higher cytotoxicity to some human tumor cell lines. This shows that the fusion protein TLT may be promising in application. PMID- 12114963 TI - Functional Difference between N Domain and C Domain of hEGF and hTGF-alpha. AB - By exchanging the N domain and C domain of hEGF and hTGF-alpha genes by PCR, two chimeras E-TGF(EGF(1-32)-TGF-alpha(34-50))and T-EGF(TGF-alpha(1-33)-EGF(33 53))were constructed. The wild and chimeric molecules were expressed in E.coli under phoA system. The expressed hEGF, hTGF-alpha and two chimeras were purified. The EGF receptor competitive binding affinity of the four molecules was hEGF > hTGF-alpha and E-TGF > T-EGF and the cell proliferation stimulating activity of them was hTGF-alpha and E-TGF > T-EGF > hEGF. The result suggests that the N domain of hEGF and hTGF-alpha may play a major role in receptor binding activity and C domain of them may be responsible for stimulating cell proliferation. PMID- 12114964 TI - Effect of High Hydrostatic Pressure on Activity of Restriction Endonucleases. AB - The effect of high hydrostatic pressure on the activities of type II restriction enzymes HindIII and XbaI in digesting plasmid pSPORT1 was studied. The endonuclease activity of HindIII and XbaI at 37 degrees were gradually inhibited by increasing pressure and completely inhibited at 200 and 180 MPa, respectively. No obvious irreversible effect was observed for HindIII after suffering high pressure, while a considerable irreversible inactivation was observed for XbaI. The standard molar volume changes for HindIII and XbaI estimated from the inhibition of endonuclease activity at different pressures were 213 and 103 ml/mol, respectively. It was also concluded that pressurization did not change the substrate sequence specificity of both HindIII and XbaI. PMID- 12114965 TI - Telomere Shortening of MCF-7 Cells Caused by Antisense Telomerase cDNA. AB - pAdE(1)CMVITREXneo is a novel adenovirus vector which can integrate foreign genes into genome of host cells. The cDNA of human telomerase RNA was integrated inversely into the vector via antisense oligonucleotide technique. Antisense recombinant virus (vAdT-AAV) was obtained by cotransfecting pAdT and pBHG(11) into 293 cells. The telomere length of breast cancer MCF-7 cells was found shortened after the transfection by vAdT-AAV. PMID- 12114966 TI - Molecular Cloning and Expression of the cDNA for Disintegrin from Agkistrodon acutus. AB - In order to clone novel and valuable genes from natural resources and develop genetic engineering drugs, total RNA was extracted from the venom gland of Agkistrodon acutus by one step method. The cDNAs for low molecular weight metalloproteinase were amplified by RT-PCR and cloned into the pGEMT vector. Their sequence were determined. One of the deduced amino acid sequences included a metalloproteinase and a disintegrin (acugrin) at the carboxyl terminus. This result further confirms that metalloproteinase and disintegrin are derived from the proenzyme of snake venom metalloproeinase. Acugrin exhibits a certain amino acid homology with other disintegrins from snake venoms. Acugrin has four disulfide bonds and a consensus motif RGD. The cDNA for the acugrin was cloned into the plasmid pT7ZZa and expressed in E.coli. ZZ-acugrin fusion protein could strongly inhibit aggregation of platelet induced by ADP and the IC(50) was about 4 nmol/L. PMID- 12114967 TI - Molecular Cloning of MAPK Gene Family Using Synthetic Oligonucleotide Probe. AB - MAPK(mitogen activated protein kinase) is a kind of Ser/Thr protein kinase. The MAPKs play an important role in several different signal transduction pathways. The MAPKs may also have a role in morphorgenesis of Candida albicans. An oligonucleotide probe was used to screen novel MAPKs in C. albicans. All MAPKs shared high homogeneity in their eleven kinase subdomains, especially subdoman VII and VIII. In subdomain VII, nearly all MAPKs have the same KIDFGLAR sequence, and the two known MAPKs in C. albicans CEK1 and MKC1 have only one different nucleotide in that DNA sequence. This probe was hybridized with C. albicans genomic DNA. Under stringent conditions, the probe could only hybridize with CEK1 and MKC1 gene fragment. But when hybridized at 40 degrees in non-SDS solution, two novel bands appeared. This condition was used to screen SC5314 DNA library, and many positive clones with different hybridization density were obtained. The strongest hybridization clones were identified to contain CEK1 and MKC1 gene. From the stronger positive hybridization clones, two novel genes were identified. The first gene, named CRK1(CDC2-related protein kinase 1), shared high homogeneity to MAPKs, but was not of them. It is closest to SGV1 from S. cerevisiae (with homology 47%) and PITALRE from human (with homology 41%), both of which are CDC2-related protein kinases. The second gene called CEK2(Candida albicans extracelluar signal-regulated kinase 2) is a novel MAPK of Candida albicans, which shares the highest identity with CEK1 and its S. cerevisiae homologs, FUS3 and KSS1, two redundant MAPKs in yeast pheromone response and morphogenesis. PMID- 12114968 TI - Functions of CRK1 Gene of Candida albicans as Studied by Gene Knock-out. AB - In the course of screening MAPK related protein kinase genes from Candida albicans DNA library, a CRK1 (CDC2-related protein kinase) gene was identified. Deletion of the CRK1 gene significantly decreased the growth. rate of cells. CRK1 knock out strains precipitated more rapidly in YPD liquid medium. The CRK1 null mutant stimulated hyphae formation in Lee's liquid medium at pH 4.5 and 25 degrees, which is the medium that maintains the wild type Candida albicans strain in yeast form. These phenomena suggest that CRK1 gene may be involved in cell cycle, flocculation and morphogenesis of Candida albicans. Overexpression of the half length CRK1 gene could induce the invasive growth into agar faintly, the cells that was capable of the invasive growth also changed their cell shapes to long, thin pseudohyphal-like cells, but the full-length CRK1 had only weak effect. The Crk1p might promote the invasive growth or cell elongation through a route different from the MAPK signal transduction pathway. PMID- 12114969 TI - Synthesis and Biological Activity of Human Calcitonin Analogue. AB - The potency of salmon calcitonin (sCT) is higher but the structural homology between sCT and human calcition (hCT) is only 50%. Based on the comparison of the structure between sCT and hCT, we have designed and synthesized a hCT analogue (mhCT-2) by solid phase method, using air oxidation in diluted solution to obtain a peptide with intramolecular disulfide bond. Through HPLC purification, we obtained a capillary electrophoresis-homogeneous mhCT, results of analysis of its mass spectrum and N-terminal sequencing were in accordance with the theoretical values. The results of calcitonin bioassay by estimating the blood calcium levels in rats showed that the potency of mhCT-2 was around 2 000 IU/mg, one order of magnitude higher than that of hCT. In RIA assay, we have found that the immunoactivity of mhCT-2 and hCT was much different because of their different binding abilities to anti-hCT antibody. This indicated that the conformation of mhCT-2 was changed as compared with hCT. In rat osteoporosis model, the results showed that pharmacologic effects of mhCT-2 was the same as that of sCT. The synthetic mhCT-2 seems promising to be a clinically useful peptide with high potential in osteoporosis therapy, because it is similar in biological properties to, but less immunogenic than sCT. PMID- 12114970 TI - Site-directed Mutagenesis of the Active Center of Penicillin Acylase from E. coli ATCC 11105. AB - Site-directed mutagenesis and chemical modification were performed at Ser290 of the penicillin G acylase from E. coli ATCC11105. The Ser290 was substituted by Cys or Secys. Wild type and mutant proteins were purified, and the activities and kinetic constants of penicillin acylases for hydrolysis and synthesis were determined, respectively. Although their K(m) values were not changed, the k(cat) values of the thiol-PGA and seleno-PGA were decreased from 135s(-1) to 0.63s(-1) and 0.38s(-1) against NIPAB, and from 34.38s(-1) to 0.23s(-1) and 0.06s(-1) against penicillin G. Contrary to Choi's report(Choi K S (et al. J Bacteriology), 1992, 10 6270-6276), we found that hydrolysis activity was certainly kept in the mutant of penicillin acylase. In addition, the specific activities of synthesis were decreased by 5-fold and 20-fold, respectively. PMID- 12114971 TI - Structure of Cytoplasmic Polyhedrosis Virus from Bombyx mori. AB - The structures of full and empty capsids of CPV were studied by negative staining and electron cryomicroscopy and computer reconstruction techniques. By comparing the structures and biochemical compositions, the CPV was identified as a single layered capsid with its five structural proteins located on it. This single capsid is arranged according to T=1 icosahedral symmetry with 12 turret-like spikes at its icosahedral vertices. The empty and full CPV show identical capsid but differ inside. The dense and ordered genomic dsRNA is located inside the full CPV. The internal space of the empty CPV has almost no electron density except for 12 electron densities attributed the transcriptional enzyme complexes extending inward from the base part of CPV spikes. PMID- 12114972 TI - Cloning and Nucleotide Sequencing of Prolyl Endopeptidase Gene from Aeromonas punctata subsp. punctata. AB - Prolyl endopeptidase activity was found in Aeromonas punctata subsp. Punctata. The genomic DNA was partially digested with EcoRI and the recovered 8-16 kb DNA fragments were inserted into the EcoRI site of plasmid pUC19, and were transformated into Escherichia coli DH5alpha. The resulted clones were screened by using Benzyloxycarbonyl-Gly-Pro-beta-naphthylamide, the specific substrate of prolyl endopeptidase and a positive clone was obtained. The 12 kb insertion fragment of recombinant plasmid was digested with HincII and subcloned. The PEP gene was found in the 3.5 kb HincII/EcoRI fragment. Nucleotide sequence of the gene was completely sequenced by Auto Sequencer. The complete gene consisted of 2 073 bp corresponding to 690 amino acid residues with a calculated molecular weight of 76 467 Da. The amino acid sequence was 92.3% 53.2% 33.5% 33.2% and 20.5% homologous to those of Aeromonas hydrophila, Flavobacterium meningosepticum, porcine brain, human lymphocytes and Pyrococcus furiosus respectively. From a survey of sequence homology with other members of the prolyl endopeptidase family, the amino acid residues involved in the catalytic triad were deduced to be Ser(538) Asp(622) and His(657). PMID- 12114973 TI - Regulation of Phospholipase D Activity in Human Hepatocacinoma Cells by Protein Kinases and D-sphingosine. AB - The effects of some inhibitors of protein kinase C(PKC) and tyrosine protein kinase(TPK)as well as the antibodies to PKC isotypes on the activity of phosphatidylcholine-specific phospholipase D(PLD)in 7721 human hepatocarcinoma cells were determined in order to study the regulation of PKC and TPK on PLD in these cells. It was found that all of the four inhibitors of PKC (chelerythrine, H-7, calphostin C and stausporine) and two inhibitors of TPK (tyrphostin 46 and genistein) partially inhibited the basal activity of PLD. Among them, the inhibition rates of staurosporine and calphostin C were the highest. The effects of TPK inhibitors were less than that of PKC inhibitors. When the inhibitors of PKC and TPK were added in combination, the inhibitory effect was greater than that used separately. A well known physiological inhibitor of PKC, D-sphingosine, did not show any inhibition, but did show stimulation on PLD activity. The mechanism is probably related to the transformation of D-sphingosine to D sphingosine 1-phosphate, a stimulator of PLD via the activation TPK (and probably also PKC). The stimulating effects of both D-sphingosine and D-sphingosine 1 phosphate were blocked by TPK inhibitors and other PKC inhibitors. Among the 3 common PKC isotypes in human hepatocarcinoma cells, PKCalpha and PKCbetaI may be the main isotypes of PKC in the regulation of PLD. PMID- 12114974 TI - Roles of Various Regions of the HBV Core Promoter in the Transcription of Precore and Pregenomic RNA. AB - To determine the potential functions of different regions in the HBV core promoter, several mutants containing B3 and/or C deletion (p3.6IIdeltaB3, p3.5IIdeltaB3, p3.6IIdeltaC, p3.5IIdeltaB3C) have been constructed based on the two HBV transcription units (p3.6II and p3.5II) which contained ENII/Cp and Cp immediate upstream of the linear HBV genome, respectively. The functional effects of deletions were assessed with respect to antigen production, viral mRNA synthesis, and viral DNA replication. When HepG2 cells were transfected with C deletion mutants, HBeAg became undetectable in the supernatant while the expression of HBcAg in the cell lysate was retained. Consistently, precore RNA disappeared but the synthesis of pregenomic RNA and viral DNA replication could still be observed. On the other hand, deletions of B3 fragment reduced the expression of HBeAg and HBcAg significantly, while maintained the synthesis of precore RNA, pregenomic RNA and viral progeny DNA. The results strongly suggest that both fragment C and fragment D serve as minimal promoter elements for the transcription initiation of 3.5 kb RNAs. Notably, the D fragment which contains a novel nuclear factor binding site close to the start site of pregenomic RNA, is critical for the basal transcription of pregenomic RNA, which has not been reported previously. B3 plays an important role in the efficient transcription of precore and pregenomic RNAs, but is not indispensable for their basal transcription. PMID- 12114975 TI - The Regulation of Xmyf-5 Gene, a Muscle Determinant, Expression in Xenopus Laevis. AB - Xmyf-5 gene is a member of the myogenic regulatory genes important for Xenopus muscle cell commitment and differentiation. To analyze the mechanism involved in regulating Xmyf-5 gene activity in the process of myogenesis, the 4.9 kb 5' upstream sequence of Xmyf-5 was identified. LacZ gene could be activated in the presomitic mesoderm in stage 14 embryos and myotomes in stage 25 embryos when it was driven by the 4.9 kb sequence and introduced into embryos at 2 cell stage. The Luciferase reporter gene activities driven by deletion constructs suggested that there are at least one enhancer between 4.9 kb and 3.5 kb of Xmyf-5 5' upstream DNA and one negatively regulative factor between 2.0 kb and 0.9 kb of the Xmyf-5 5' upstream DNA. In addition, sequence analysis indicated that a 100 bp fragment 5' upstream of Xmyf-5 contained conserved regulative elements such as OLS which were found in other vertebrates. PMID- 12114976 TI - Secretory Expression of Human Insulin in Methylotrophic Yeast Pichia pastoris. AB - The porcine insulin precursor (PIP) gene and its derivative form sp-PIP gene, which had a nona-peptide (called spacer peptide, sp) added at the 5' terminus of PIP gene, were inserted into the plasmid pPIC9 of Pichia pastoris to obtain secretory plasmid pPIC9/PIP and pPIC9/sp-PIP, respectively. P.pastoris GS115 was transformed by pPIC9/PIP or pPIC9/sp-PIP and the high-copy strains, P39(-sp) and S51(+sp), were selected by dot-blotting. The expression levels of PIP and sp-PIP were 10 mg/L and 40 mg/L in 1 L shake flask, respectively, indicating that the spacer peptide could increase the expression. The expression level of PIP (sp PIP) in P.pastoris was higher than that of PIP in S.cerevisiae and K.lactis reported in this laboratory. The expression level of sp-PIP was 250 mg/L in 10 L fermentor. Recombinant human insulin was obtained by means of transpeptidation of PIP or sp-PIP. The receptor binding capacity is identical with that of porcine insulin. In vivo biological activity of the recombinant human insulin is 27 IU/mg. PMID- 12114977 TI - Application of Free-flow Electrophoresis to the Separation of Murine Lymphocytes. AB - Free-flow electrophoresis offers not only the all-in-solution separation process but also very gentle separation conditions, and it can be used for continuous separation and preparation, so it has been widely applied in the fields of biochemistry and cell biology. Using this technique to separate insoluble particles such as cells, both high separation efficiency and high activity and viability could be obtained. Murine lymphocytes were separated in low ionic strength triethanolamine buffer by free-flow electrophoresis, the cell fractions were detected and characterized by means of UV spectrometry, immune fluorescence labeling and flow cytometry. Results indicated that T and B lymphocytes were well separated with high viability. PMID- 12114978 TI - Preparation and Structural Analysis of Sulfated Derivatives of Lentinan. AB - A series of sulfated derivatives of lentinan were prepared by the modified Wolfrom method. Sulfates content analysis indicated that the proportion of sulfate substitution depended on the reaction time and the ratio of chlorsulfonic acid and pyridine in the ester reagent. Methylation analysis by Needs method and Hakomori method was proved not to be suitable for the structural analysis of sulfated lentinan. (13)C-NMR data showed that sulfates were located on the carbon 6 in all sulfated derivatives of lentinan showing the higher reactivity of the hydroxyls on C-6 position than those at other position. PMID- 12114979 TI - Human &mgr; Opioid Receptor Expressed in Sf9 Insect Cells was Functionally Coupled to Endogenous G Protein. AB - Human &mgr; opioid receptor(H&mgr;OR) with a tag of six consecutive histidines at its carboxyl terminus overexpressed in recombinant baculovirus infected Sf9 insect cells was demonstrated to be functionally coupled to endogenous G proteins. Na(+) and GTP could reduce the affinity binding of etorphine and Ohmefentanyl(Ohm) to H&mgr;OR overexpressed in Sf9. The binding of (35)S GTPgammaS to Sf9 cell membranes containing H&mgr;OR was stimulated by DAGO and Ohm. This stimulation could be blocked by naloxone or the pretreatment of PTX. Also, DAGO and Ohm could inhibit the increasing of intracellular cAMP stimulated by forskolin. The results showed H&mgr;OR expressed in Sf9 insect cells were functionally coupled to endogenous PTX sensitive G(i/o) proteins. PMID- 12114980 TI - High Expression of Penicillin G Acylase Gene from Bacillus megaterium in Bacillus subtilis. AB - The penicillin G acylase gene (pga) amplified by PCR from Bacillus megaterium was subcloned into an expressing vector pPZW103 (P43 as promoter). The recombinant plasmid was transferred into Bacillus subtilis DB104. Penicillin G acylase production in the B. subtilis transformant was 3-6 u/ml, higher than that of published recombinant strains. Penicillin G acylase production was induced by phenylacetic acid in B. megaterium, whereas the enzyme was produced constitutively in the B. subtilis transformant carrying B. megaterium pga. The recombinant strain showed high stability in antibiotic-free medium for 10 days. Enzyme in crude broth was purified by Al(2)O(3) chromatography and phenyl Sepharose CL-4B hydrophobic chromatography and the total yield is 79%. The purified enzyme with specific activity of 52 u/mg can be directly immobilized for use. PMID- 12114981 TI - Characteristics of BmK AS Binding to Rat Brain and Cockroach Nerve Cord Synaptosomes. AB - BmK AS, one novel neurotoxic polypeptide from Chinese scorpion Buthus martensi Karsch was labeled by (125)I with INDOGEN method. Iododerivative was used in binding experiments on mammal and insect synaptosomes. BmK AS binds specifically to a single class of non-interacting binding sites with high affinity (K(d)=1.49 nmol/L) and low capacity (B(max)=1.39 nmol/g) on rat brain synaptosomes. Likewise, BmK AS also binds specifically to a single class of non-interacting binding sites with high affinity (K(d)=0.79 nmol/L) and low capacity (B(max)=6.60 nmol/g) on cockroach nerve cord synaptosomes. It indicates that there are special target receptors for BmK AS type scorpion neurotoxins on both mammal and insect excitable cell membranes. PMID- 12114982 TI - Molecular Design, Cloning, Expression and Purification of NH(2)-terminal Mutant of Staphylokinase deltaNSak AB - The 3-D structure of human microplasminogen was predicted by the method of homology modeling. The binding sites of staphylokinase and microplasminogen were determined by high-resolution genetic docking. This model is consistent with several known experimental properties of staphylokinase. To elucidate the function of NH(2)-terminus of staphylokinase for further rational reconstruction, a NH(2)-terminal-deletion mutant of staphylokinase(deltaNSak) was designed. The soluble deltaNSak was achieved with fibrinolytic activity. deltaNSak was purified by two-step ion exchange chromatography. The purity was over 95% and the specific activity of deltaNSak was 9x10(4) IU/mg. These data suggest that the 15 NH(2) terminal amino acids of staphylokinase are not required for its plasminogen activating potential and the computer model is reasonable. PMID- 12114983 TI - N-cadherin Invovles in P19 Cell in vitro Neuronal Differentiation. AB - N-cadherin plays an important role in the early morphogenesis of neural tissue. For exploring the possible role of N-cadherin in neuronal differentiation in vitro, we chose RA induced P19 cell neuronal differentiation as a model system. Using RT-PCR, we examined the expression pattern of N-cadherin during P19 cell neuronal differentiation. The results showed that N-cadherin expression was increased rapidly during RA induction with its highest expression level at day 3 of neuronal differentiation, then down-regulated along with neuronal maturation. This expression pattern was similar with CNS development in vivo. Then, we constructed a N-cadherin expression-vector and transfected in P19 cells. The stable transfected cell lines, N-cadherin/P19 were selected, and we found that, after aggregated 4 days, the N-cadherin overexpression P19 cells could differentiate into neuron-like cells without the RA induction. The immunocytochemical staining showed that these neuron-like cells expressed neuron specific marker, NF160. These results suggest that N-cadherin gene is involved in P19 neuron differentiation. PMID- 12114984 TI - Progress in Protein Translocation across Mitochondrial Membrane. AB - About 98% of the 1 000 or so mitochondrial proteins are encoded in the nucleus and synthesized on cytosolic ribosomes as precursors, which are then imported into mitochondria. In the present article the import machinery and pathway of mitochondrial proteins located in the matrix or inner membrane are described. Apocytochrome c, precursor of cytochrome c does not possess a cleavable N terminal presequence and no proteinaceous component responsible for its import has ever been identified. A quite unique translocation pathway of apocytochrome c is also reviewed. PMID- 12114985 TI - Structure and Function of Vascular Endothelial Growth Factor and Its Related Proteins. AB - Vascular endothelial growth factor(VEGF) is a highly specific mitogen promoting the formation of blood vessels in embryogenesis and wound healing. It is also a potent inducer of vascular permeability. It is a member of the cystine knot growth factor superfamily. A detailed structural and functional characterization of the interactions between VEGF and its receptors is a prerequisite for the design of molecule antagonists. These structural characterizations and biological properties make VEGF an important research object in the fields of neovascularization of ischemia tissues, prognosis of cancers, tumor metastasis, and the gene therapy. PMID- 12114986 TI - A Novel Oxidized Low Density Lipoprotein-binding Protein in Macrophage. AB - Scavenger receptor A(SR-A)on the mouse peritoneal macrophages(MPM) mediates the endocytic uptake of oxidized low density lipoproteins(ox-LDL). However, using ligand blotting and immunoblotting, a novel macrophage membrane protein binding to ox-LDL with estimated molecular mass of 92 kD was found. This membrane protein could not bind to ac-LDL. Its binding to ligands was not affected by reducing reagents either. Preincubation with medium containing neuraminidase dramatically decreased binding of the 92 kD membrane protein to ox-LDL. Excess amounts of ox LDL could completely block the binding of (125)I ox-LDL to 92 kD membrane protein, but polyI, dextran sulfate and fucoidin showed partial competitive inhibition effects to the binding. These results suggest that the novel 92 kD membrane protein plays important role in the MPM uptake of ox-LDL. PMID- 12114987 TI - Simultneous Expression of CS3 Colonization Factor Antigen and Cholera Toxin B Subunit in Salmonella typhi. AB - A host-plasmid balancing system was established based on asd gene in an avirulent strain of Salmonella typhi to express enterotoxigenic E.coli surface antigen CS3 and V.cholerae toxin subunit B(CTB). The plasmid can be stably maintained in the host and can express CS3 and CTB in the host cell without any antibiotic selection, although expression level and growth characteristics of the recombinant strain expressing either CS3 or CTB are superior to that of the recombinant strain which expresses both of the antigens. Antibo-dies against CS3 and CTB can be detected in sera of mice immunized with recombinant bacteria either orally or subcutaneously, and mice immunized subcutaneously can be protected from challenging with virulent strain of Salmonella typhi. This work may be helpful in constructing multivalent recombinant vaccines for prevention of bacterial diarrhea. PMID- 12114988 TI - Group II Phospholipase A(2) Activity and Its Molecular Regulation in Rat Heart during Sepsis. AB - Changes of activity and content of myocardial group II phospholipase A(2) and its mRNA transcription and stability during rat sepsis were investigated. Results showed that, compared with control group,myocardial group II phospholipase A(2) activity in early and late sepsis decreased by 25.0%(P<0.05) and increased by 47.6%(P<0.01),respectively group II phospholipase A(2) protein concentration reduced by 27.0% and augmented by 48.0%(P<0.01),respectively. Myocardial group II phospholipase A(2) mRNA transcription rate and content showed similar two-phases changes. The mRNA transcription rate during early and late sepsis decreased by 45.0% and increased by 70.0%(P<0.01),respectively. The mRNA content decreased by 34.1% in early sepsis and increased by 157.0% in late sepsis(P<0.01), respectively. The half-life of group II phospholipase A(2) mRNA remained unchanged notably during early and late stage of sepsis. These data suggest that myocardial group II phospholipase A(2) activity decreased in early stage of sepsis and increased in its late stage, and these changes were regulated transcriptionally. PMID- 12114989 TI - Cloning, Expression, Characterization and Application of Bst DNA Polymerase Large Fragment. AB - DNA sequencing using thermostable DNA polymerase is very valuable in molecular biology. From a specific strain of Bacillus stearothermophilus, the gene of the Bst DNA polymerase large fragment which had no 5'-3' exonuclease activity was cloned and recombined into an expression vector pYZ23,and the constructed plasmid was introduced into Escherichia coli JF1125,then the expressed protein was isolated and purified. The genetic engineered product shows characteristics similar with the purified natural Bst DNA polymerase large fragment. It will benefit the wide application of Bst DNA polymerase and the study of the relationship between the structure and function of Bst DNA polymerase. PMID- 12114990 TI - Effect of IL-3 on Globin Gene Expression in a Human Erythroleukemia Cell Line. AB - A human erythroleukemia cell line(K562 cells)was used as a model to study the effect of interleukin-3(IL-3) on human globin gene expression in the cells. The results showed that the beta-globin gene was not expressed in uninduced K562 cells however, it was expressed when K562 cells were induced for 3 or 5 days by IL-3. The expression of alpha- and gamma-globin gene were not much different between IL-3 induced and uninduced K562 cells. Using the method of benzidine dyeing, it was observed that the percentage of blue cells was significantly increased when K562 cells were induced by IL-3. It suggested that IL-3 could not only activate the exp-ression of beta-globin gene in K562 cells, but also induce the synthesis of hemoglobin. Therefore, IL-3 may play a critical role in inducing K562 cells to terminal differentiation. PMID- 12114991 TI - HPLC and MS Studies of Orientation of Melittin on Phospholipid Membrane. AB - The orientation of melittin binding to liposomes with or without transmembrane potential was investigated. With a combination of high performance liquid chromatography and mass spectrometry, the membrane association state of melittin was detected by analyzing its trypsin-digested products. It was found that the enzyme could access all the proteolytic sites on protein binding to the liposome without membrane potential, while one proteolytic site on the N-terminal of the protein was blocked after it binding to the liposome with negative transmembrane potential. The results showed that melittin changed its orientation in the latter case. PMID- 12114992 TI - Effect of Protein Kinase C Inhibitor on Scavenger Receptor in Human U937 Macrophage-like Cells. AB - In order to investigate effects of cell protein phosphorylation on scavenger receptor, human U937 macrophage-like cells were treated with protein kinase C inhibitor staurosporine, then the cells were incubated with (125)I ox-LDL or ox LDL, and the cellular degradation of (125)I ox-LDL, its binding to receptor and the internalization of cell surface ox-LDL receptor complex as well as the accumulation of lipids within cells were measured separately. Moreover, the effects of the drug on expression of cell surface receptor were observed by means of autoradiography. The results indicated that staurosporine could enhance U937 cells to bind lipids and stimulate scavenger receptor expression, and could reduce degradation of lipids by U937 cells and the accumulation of cholesterol within the cells. It suggests that the function of scavenger receptors may be correlated with cell protein phorsphorylation. PMID- 12114993 TI - Binding of Insulin Receptor Substrate 1 PH Domain to Protein Kinase C. AB - To screen for novel ligands of insulin receptor substrate 1(IRS-1)PH domains, to investigate the role of the PH domains in signal transduction processes and to examine association of the PH domain with protein kinase C(PKC), rat liver was employed to clone the gene encoding for the PH domains. Total RNAs were isolated and purified from fresh liver and mRNAs were reversely transcribed into cDNAs. After PCR the fragments of the DNA were cloned into vector pUC19. The sequence of the fusion gene was confirmed by sequencing, and the gene was correctly expressed in E.coli as fusion protein with glutathione S- transferase(GST). The fusion protein was purified by glutathione agarose beads, then was incubated with the lysate of Jurkat cells. After SDS-PAGE, the proteins were transferred to PVDF membrane, and an anti-PKC antibody was used to detect binding between the PH domain with PKC. The sequence of the gene encoding for the PH domains was confirmed to be correct, and the PH domain was successfully expressed in E.coli JM 109 in soluble form. Western blots confirmed the binding of the PH domain with PKC in vitro. In conclusion, purified IRS-1 PH domain GST fusion protein was obtained and its biological activity was confirmed. PMID- 12114994 TI - Sequence Analysis of SOD Gene of Bombyx mori Nuclear Polyhedrosis Virus and Its Expression in E.coli. AB - Superoxide dismutases scavenge superoxide radicals and protect cells from oxidative stress. SOD gene of Bombyx mori nuclear polyhedrosis virus has been cloned by PCR, and is expressed in E.coli with the activity of SOD being 576.13 u/mL. DNA sequence analysis shows that SOD gene of BmNPV encodes 151 amino acids. The homology between BmNPV and AcNPV of the nucleotide sequences of SOD gene is 97.2%, and 56% between BmNPV and human SOD1. PMID- 12114995 TI - Dispersive X-ray Spectroscopy Analysis of Porous beta-TCP Bioceramics after Implantation into Femur. AB - In this paper, energy dispersive X-ray spectroscopy and element ratio of implanted bioceramic, interface and rabbit femur when the bioceramic implanted into femur were measured by two different scan electron microscopy (SEM) EDXA modes. The changes of element ratio and components of materials and interface implanted into femur were compared. Thorough studies were made on the transformation of TCP from lifeless materials into living bones after implantation, by using SEM-EDXA, Raman spectra and infrared spectra. These results provided rich evidence for understanding biodegration and bone bonding mechanism of calcium phosphate bioceramics. PMID- 12114996 TI - Cellobiose Dehydrogenase Inhibition of Polymerization of Phenolic Compounds and Enhancing Lignin Degradation by Lignina. AB - The kinetic behavior of cellobiose dehydrogenase (CDH) was investigated by steady state initial velocity studies. Variation in the concentration of one substrate led to changes in K(m) and V(max) of the other substrate. The results were consistent with a ping-pong mechanism. In the presence of cellobiose, CDH could reduce many oxidized products catalyzed by soybean hull peroxidase (SHP). The oxidation product of 1-hydroxybenzotriazole (HBT) catalyzed by SHP inactivated the enzyme itself however, CDH could prevent SHP from inactivation by reducing the oxidation product of HBT. CDH could also inhibit the polymerization of phenolic compounds catalyzed by SHP. It was found that the addition of CDH could enhance kraft pulp lignin degradation by ligninases. PMID- 12114997 TI - Isolation and Identification of Membrane Proteins Binding to Transfer RNA. AB - Crude extract of total membrane proteins of yeast was obtained by a method of phase separation in Triton X-114. The membrane extract was applied to the affinity column on which yeast total tRNAs were covalently coupled to hydrazinyl Sepharose 4B. By eluting with increasing concentration gradient of (NH(4))(2)SO(4), eluted proteins were found between concentrations of (NH(4))(2)SO(4) 0.1 mol/L to 0.3 mol/L. By gel mobility-shift assays, it was also observed that there were more than two mobility-shift bands on the gel electropherogram after incubation of sample of (32)P-labeled tRNA with the eluent proteins and the reaction mixtures were analyzed by non-denaturing polyacrylamide gel electrophoresis. These results confirm that these eluted proteins contain two major tRNA specific binding proteins. PMID- 12114998 TI - Protein and Peptide Identification by Capillary Zone Electrophoresis-Tandem Mass Spectrometry. AB - A method for highly sensitive protein and peptide identification by capillary zone electrophoresis/electrospray ionization-tandem mass spectrometry (CZE/ESI/MS/MS) was described. A mixture of peptides Met-enkephalin and Leu enkephalin was analyzed, and their amino acid sequences were identified, respectively, using CZE/MS/MS. Peptide mapping for a tryptic digest of horse cytochrome c was performed by the same method. The results indicated that almost all peptides generated by tryptic digestion were identified, and their sequences were interpreted by their collision-induced (CID) mass spectra. Applying the uninterpreted CID spectra of the protein digest to searching in sequence database by the SEQUEST program could identify proteins conveniently and rapidly. The samples consumed in the CZE/MS/MS experiments were at picomole level, so this method is quite suitable for quality control of recombinant proteins and trace amount protein identification in the proteome research. PMID- 12114999 TI - An Evolutionary Trace Method for Functional Prediction of Genomes. AB - It is essential for functional genomics to develop an accurate and efficient functional prediction method of genomes. Here, a new method is suggested, that is based on the fact that ortholog-specific motif is an evolutionary trace, in which the functional prediction of genomics is carried out. First, orthologous sets in a family were constructed using evolutionary analysis then functional motifs for each orthologous set were found out and a database consisted of these motifs was built. When this database is completed, unknown genes can be predicted accurately and quickly by searching it. The pilot study on five families proves that our method is feasible. PMID- 12115000 TI - Mechanism of Differentiation of HEL Cells Induced by Hydroxyurea. AB - HEL cells, a human erythroleukemia cell line, mainly express fetal globin gene(gamma) and small amount of the embryonic(epsilon) globin gene, but not the adult (beta) globin gene. Our previous studies demonstrated that hydroxyurea could induce HEL cells to express adult (beta) globin gene and induce HEL cells to terminal differentiation. In order to reveal the differentiation mechanism of HEL cells induced by hydroxyurea, GMSA was performed to examine the binding of nuclear extracts from hydroxyurea induced and uninduced HEL cells to the positive control region in the 5'-flanking sequence of human beta-globin gene and to the synthesized GATA oligonucleotides. Our results showed that, in the nuclear proteins from the induced HEL cells, the binding of GATA-1 to GATA motif was increased. However, the binding of GATA-2 to GATA motif was decreased. Besides, our data showed that a YY1-like protein was decreased quickly in hydroxyurea induced HEL cells. Western blotting analysis was used to analyze variation of GATA proteins in nuclei of HEL cells. Results revealed that the amount of GATA-2 was significantly decreased while the amount of GATA-1 was increased after HEL cells were induced for 12 h. These results indicate that GATA-1 may play a critical role in inducing HEL cells to terminal differentiation. The YY1-like protein and GATA-2 might inhibit HEL cells from terminal differentiation. PMID- 12115001 TI - Glucocorticoids Modulate G-protein alpha-subunit Levels in Rat Hypothalamus. AB - Glucocorticoids (Gc) affect neuroendocrine function of hypothalamic-pituitary adrenal axis (HPAA), especially of the hypothalamus. Synthesis and secretion of neuronal hormones of hypothalamus are associated with G-protein-linked signalling system. Here, the regulation by Gc of the levels of several G-protein alpha subunits in rat hypothalamus was studied with Western blot method. Results showed that Gc downregulated Galphas1, Galphas2,Galphai and Galphaq with different magnitudes, being 14.37,15.66,14.61 and 12.37 per cent, respectively. However, Gc did not affect Galphao expression. Our results provided important information for understanding how Gc alters neuroendocrine function through G-protein-linked signalling system. PMID- 12115002 TI - Syp Y279, Y304 Can Mediate the Binding of Bcr-Abl to Grb2 and Other Proteins. AB - In this study we obtained 3 mutants of Syp cDNA Y279F, Y304F and Y546F by using the method of site-directed mutation in vitro. We then inserted them into the pXM mammalian expression vector, and transferred the plasmids into K562 cells. Western Blot has found WT/ 3MT Syp are expressed in the K562 cells. Immunoprecipitation and Immunoblot have found that WT, Y279F, Y304F and Y546F Syp can bind directly to the Bcr-Abl in vivo. However, we found that mutation of Syp Y279F can block the binding of Grb2 SH2 to Syp mutation of Syp Y304F can block the binding of Shc SH2 to Syp in vitro. As a adaptor, Syp can mediate binding of Bcr-Abl to Shc and Grb2.Grb2 bind to Syp pY279, Shc bind to Syp pY304 in vitro. PMID- 12115003 TI - The Feasibility of Using Proteome Expression Profile for Genome Annotation. AB - By investigating into the expression data from ECO2DBASE (Edition 6),the feasibility of using proteome expression profile for genome annotation was tested. Based on our newly developed CRC (cellular role cluster) method,79 proteins extracted from ECO2DBASE were clustered into 4 CRCs. Function related proteins tend to be clustered into same CRC. Total 9 aminoacyl-tRNA synthetases were clustered into CRC2, whereas 4 heat-shock proteins into CRC3. These results indicate with enough proteome expression data and the efficient algorithm, proteome expression profile can provide very important information for genome annotation, while this kind of information is sequence-independent. PMID- 12115004 TI - Analysis of N-Terminal Nucleotide Sequence of NDV Fusion Protein. AB - Fusion protein of Newcastle disease virus is a glycoprotein with antigenicity. The cDNA of fusion gene of NDV strain F48E8 was cloned by RT-PCR, and the sequence of F48E8 was compared with other two virulent strains, Australia victoria and Italien. Results show that the sequences of N-terminus are very similar, and cysteine residues and the potential glycosylation sites are relatively conserved. However, there are a few differences in the region of signal peptides. PMID- 12115005 TI - Expression, Purification and Characterization of Regulatory Subunit of Type I cAMP-dependent Protein Kinase in E.coli. AB - Regulatory subunit of type I cAMP-dependent protein kinase (RI) possesses two cAMP-binding sites with high affinity for cAMP. In the current study, human RI cDNA fragment coding for residue 93-381 was cloned from neuronal cells. After subcloning into pET30a, the recombinant RI was expressed in E.coli BL21. Considerable amount of soluble RI subunit was produced in BL21 upon induction of IPTG. RI was purified to homogeneity over 95% after His-bind metal chelation chromatography. Approximately 8 mg pure RI was obtained from 0.1 liter of BL21 culture. Human RI prepared with this method binds to cAMP specifically and therefore can be used in cAMP assay. PMID- 12115006 TI - Potential GM-CSF Antagonists Selected from a Peptide Phage Display Library. AB - Peptide phage display libraries have been successfully applied in areas of mapping antibody epitope, finding ligands for enzymes, receptors, and many other molecules. But it has been demonstrated to be very difficult to select cytokine binders from peptide phage display libraries probably because cytokine is not so sticky as antibody that there are rare chances of capturing peptide phages during biopanning. A pVIII-based peptide phage display library was panned with the cytokine GM-CSF and some GM-CSF binding clones were selected based on high throughput screening (HTS) method and confirmed by ELISA and micropanning assays. These cytokine-binders may be utilized in affinity chromatography in cytokine downstream processing and even act as potential antagonists of GM-CSF if their affinity are further improved through secondary library strategy. PMID- 12115007 TI - A Discriminator Base in tDNA(Trp). AB - The discriminator base G73 was one of the major identity elements of tRNA(Trp). Aminoacylation assay and CD spectrum of tDNA(Trp)-NCCrA demonstrated that under the same pH, the change of discriminator base affected tRNA conformation at the 3' terminus while tDNA(Trp) with the same discriminator base exhibited different conformation and aminoacylation activity under various pH conditions. Our results showed that it was the conformation of tRNA, but not the base itself, that determined the accurate recognition between tRNA and its cognate synthetase. PMID- 12115008 TI - Activity of Lipase in Reversed Micelles. AB - The enzyme activity of lipase from Candida rugosa in AOT isooctane reversed micelles was investigated. Effect of pH, temperature, substrate concentration, waterpool (R value), choice of organic solvent, surfactant concentration on the activity of lipase were studied in detail. Results indicate that the enzyme in reversed micelles follows the Michaelis-Menten dynamics temperature optimum=32.5 degrees, pH optimum=7.0, and maximum activity is reached at the concentration of olive oil of 40%. The enzyme activity was dependent on R (molar ratio of water to surfactant) and maximum activity was obtained at R=11. PMID- 12115009 TI - Preparation, Characteristics and Toxicity of Ricin-containing Galactosyl Ceramide Liposomes. AB - Ricin-containing galactosyl ceramide liposomes (ricin-GCLs) lyophilized product was made by the dehydration-rehydration vesicles(DRV) method and its toxicity to human heptocarcinoma cell line 7721 and to mice was investigated. The results indicated that the maximum encapsulation efficiency of ricin was 74.1%. The distribution of ricin-GCLs' sizes accords with intravenous injection standard and changes little in half a year at 4 degrees. In the cytotoxicity assay, the ratio of ricin-GCLs' to free ricin's IC50 was 1.00:3.89, which indicated that the ricin entrapped in GCLs was more effective than the free one in killing human heptocarcinoma cell line 7721. In vivo, the mice's LD(50) of ricin-GCLs ((41.8+/ 9.33) &mgr;g/kg)is more than that of free ricin ((30.6+/-8.49) &mgr;g/kg) by 36.29%. So, ricin entrapped in GCLs promotes its cytotoxicity and reduces its toxic side-effect. PMID- 12115010 TI - Separation and Characterization of Oligodeoxynucleotides by FPLC. AB - The purity of the antisense drugs synthesized by DNA synthesizer should be determined effectively. At pH 12,the oligodeoxynucleotides (ODNs) ranged from 19 bases to 21 bases can be separated successfully on the anion exchange column MONO Q by elution with a NaCl gradient. The experiments proved that the fast protein liquid chromatography (FPLC) was a useful tool for isolation and identification of the antisense drugs. PMID- 12115012 TI - Oxygen-binding Properties of Complexes between Dextran and Pyridoxal 5-Phosphate Modified Porcine Hemoglobin. AB - It was demonstrated by oxygen equilibrium curve that the pyridoxal 5 phosphate(PLP) modified porcine deoxyhemoglobin(pHbbeta) had lower oxygen affinity than that of stroma-free porcine hemoglobin(pHb). Accourding to analysis of SDS-PAGE, the porcine hemoglobin derivatives were not cross-linked between its subunits. A conjugate was synthesized between pHbbeta and p-toluenesolfonyl chloride-actived dextran. The oxygen affinity of Dx-pHbbeta was high than that of pHbbeta, but still lower than that of pHb. Judged by cellulose acetate film electrophoresis, the mobility of Dx-pHbbeta was apparently different from that of pHbbeta. Dx-pHbbeta has characterized absorbance peak in UV spectrum, which can be used to analysis the binding ratio between Dx and pHbbeta. PMID- 12115011 TI - Analysis of Phylogenetic Relationships Among Twelve Yeast Species. AB - Partial sequences of 25 S ribosomal DNA for nine Candida yeast and one Kluyveromyces yeast strains were cloned and determined. We compared them with two published 25 S rDNA sequences of Saccharomyces cerevisiae and Candida albicans. An evolutionary tree of the twelve species was inferred from about 370 sites of 5' end of 25 S ribosomal DNA using the methods of neighbor-joining and bootstrap. The molecular data indicate a very close affinity between Candida kefyr CBS834 and Kluyveromyces cicerisporus CBS4857. PMID- 12115013 TI - Genetic polymorphisms. PMID- 12115014 TI - Old drugs, new bottles, therapeutic advantage? PMID- 12115016 TI - Double-blind, placebo-controlled study of the efficacy and tolerability of nimesulide administered orally in acute bronchial asthma. AB - In this double-blind, placebo-controlled study, children with acute exacerbation of bronchial asthma between the ages of 1 and 12 years not responding to conventional therapy with bronchodilators and injectable steroids were enrolled. A total of 60 children (two groups of 30 each) was studied. The overall response to therapy was assessed based on the guidelines and recommendations of the National Heart, Lung and Blood Institute. The efficacy parameters included respiratory and heart rates, degree of dyspnea, accessory muscle usage, color, wheeze, and degree of oxygen saturation. Children with moderate to severe exacerbation received either nimesulide suspension 1.5 mg/kg per dose or identical placebo orally as per random protocol. To assess the clinical progress, all the efficacy parameters were reassessed after 30 minutes and 1, 2, and 6 hours. A significant difference was observed in the overall assessment of response at 1, 2, and 6 hours in the two treatment groups. A greater number of children showed a good overall response in the nimesulide group compared with the placebo group at 1, 2, and 6 hours (P <.01). No side effects were reported in any of the patients in either group. None of the patients was withdrawn prematurely from either group. It is evident from the current study that nimesulide showed good efficacy and tolerability. Therefore, nimesulide could be administered to asthmatic patients whenever there is a need for such therapy. PMID- 12115015 TI - In vivo evaluation of interaction between aqueous seed extract of Garcinia kola Heckel and ciprofloxacin hydrochloride. AB - The effect of Garcinia kola seed extract (100 mg/kg) on the pharmacokinetic and antibacterial effects of ciprofloxacin hydrochloride (40 mg/kg) was studied. The results (mean +/- SEM) indicated that concurrent administration of both agents significantly (P < 0.05) decreased average serum concentration, peak serum concentration, and elimination rate of ciprofloxacin HCl, whereas the half-life and clearance rate were increased. The decrease in clearance rate was not significant. There was no difference in time to peak plasma concentration of ciprofloxacin HCl in both groups (n = 5), which occurred at 1 hour. However, the peak plasma concentration of ciprofloxacin HCl was 46.90 +/- 9.50 microg/mL in the group that received ciprofloxacin HCl alone as against 35.80 +/- 9.30 microg/mL noted in the group that received both agents (difference of 22.24%). At 2.5 hours and longer, the values were higher in the group that received both agents, but these were not statistically significant. The reciprocal serum inhibitory titer (SIT) was 33.33 and 50.00% higher in group that received ciprofloxacin HCl alone at 1 and 2.5 hours, respectively; the highest value for both groups being at 1 hour. In contrast, at 4 hours, the value of reciprocal SIT was 66.67% higher in the group that received both agents and at 24 hours, the value was zero for both groups. The observed pharmacokinetic and antibacterial interactions at various time interval indicate biphasic interaction. The interaction was antagonistic at 1 and 2.5 hours, but exhibited potentiation at 4 hours. The precise mechanism underlying the observed biphasic interaction is not fully understood. PMID- 12115017 TI - A randomized, double-blind, controlled trial of vitamin C in the management of hypertension and lipids. AB - The effect of vitamin C on blood pressure is not well established. This is a randomized, double-blind control trial. Eligible patients were followed for 8 months. Patients were randomized to 500, 1000, or 2000 mg vitamin C. During each visit, a history including medication change was obtained and standardized blood pressure measurements were performed. A 1-week dietary diary was filled out before each visit. Multiple regression analysis and subsequent multiple comparisons were used for data analysis. Fifty-four patients satisfied our criteria and agreed to participate. Thirty-one patients (mean age, 62 +/- 2 years; 52% men, 90% whites) were randomized to the three doses of vitamin C. Overall compliance was 48 +/- 2%. Both mean systolic blood pressure (SBP) and diastolic blood pressure (DBP) decreased during the vitamin C supplementation phase [mean SBP dropped by 4.5 +/- 1.8 mm Hg (P <.05) and DBP by 2.8 +/- 1.2 mm Hg (P <.05)]. There was no difference between the three vitamin C groups (P =.48). This effect was significant for only 1 month of supplementation, but the trend persisted. There was no reported intolerance to vitamin C. There was no change in lipid levels after 6 months of treatment. Vitamin C supplementation lowers blood pressure in mildly hypertensive patients. There is no additional benefit for a higher dose than 500 mg daily. The effect of vitamin C is most likely to be only short term. PMID- 12115018 TI - Long-term clinical outcome of elderly patients with reflux esophagitis: a six month to three-year follow-up study. AB - Although the prevalence of reflux esophagitis is known to increase with age, data on the long-term outcome of esophagitis in elderly patients are scarce. We sought to evaluate the clinical outcome of elderly patients with esophagitis 6 months to 3 years after diagnosis and to identify specific prognostic indicators of a poor outcome. This was a long-term (6 months to 3 years) follow-up study. Patients older than 65 years of age diagnosed as having reflux esophagitis healed after acute treatment (2 to 4 months) were included in the study. Clinical examinations and upper gastrointestinal endoscopy were performed every 6 months for the first year and annually thereafter. After healing, no therapy was prescribed; in the event of symptom recurrence, a maintenance therapy consisting either of H2 blockers or proton pump inhibitors (PPI) was prescribed. At baseline and during follow-up, the following clinical parameters were recorded: gender, age, the presence of symptoms (heartburn, acid regurgitation, epigastric/chest pain), type and dose of the maintenance therapy, nonsteroidal antiinflammatory drug use; gastric Helicobacter pylori infection, diagnosis of hiatal hernia, and/or Barrett's esophagus. The chi-square test, the Kaplan-Meier test, and Cox's proportional hazards regression analysis were used for statistical analyses. Included in the final analysis were 138 patients (M/F, 81/57; mean age, 79.7 years; range, 66-97). The numbers of patients in need of maintenance therapy were 47 of 69 (68.1%) after 6 months, 29 of 58 (50%) after 12 months, 17 of 39 (43.6%) after 24 months, and 12 of 26 (46.1%) after 36 months of follow-up. A significantly higher esophagitis relapse rate was found in patients not treated compared with subjects who were in maintenance therapy: 59% versus 8.5% (P <.0001) at 6 months, 65.5% versus 20.7% at 12 months (P <.002), 63.6% versus 11.7% at 24 months (P =.003), and 57.1% versus 8.3% at 36 months (P =.02). No significant difference in relapse rate was found in patients treated with H2 blockers versus PPIs (21.7% versus 10%). The Cox model demonstrated that no maintenance treatment (P =.00001), the presence of typical symptoms (P =.00001), the presence of hiatal hernia (P =.03), and a high severity grade of esophagitis at baseline (P =.009) were risk factors for relapse of esophagitis. In elderly subjects, esophagitis relapse occurs in a high percentage of cases, particularly in patients not treated with antisecretory drugs. The presence of typical symptoms, hiatal hernia, and a severe grade of esophagitis are risk factors for relapse. The most effective measure for minimizing the occurrence of relapse is a maintenance therapy with antisecretory drugs. PMID- 12115019 TI - Single-dose oral activated charcoal in the treatment of the self-poisoned patient: a prospective, randomized, controlled trial. AB - Oral activated charcoal (OAC) is a universally accepted treatment of the overdose patient. Although the benefits of OAC have been suggested, there are no conclusive clinical data indicating that OAC affects outcome in overdose patients. This study was a prospective, randomized, controlled trial to determine the effects of OAC treatment in the self-poisoned adult patient. Adult patients presenting to the emergency department (ED) with a history of oral overdose were assigned to treatment with OAC (50 g) or supportive care only on an even-odd day protocol. Patients did not undergo gastric evacuation procedures in the ED. The outcome measures were clinical deterioration, length of stay in the ED or hospital, and complication rate. Over a 24-month period, 1479 patients were entered into the study. There were no significant differences in outcome parameters between the OAC treatment group and controls when comparing the length of intubation time, length of hospital stay, and the complication rates associated with the overdose. There was a higher incidence of vomiting and longer length of ED stay associated with OAC treatment. The results of this study indicated that oral drug overdose patients do not require gastric evacuation or charcoal administration. OAC provided no additional benefit to supportive care alone, was associated with a higher incidence of vomiting and a longer length of ED stay, and did not improve clinical outcome. PMID- 12115020 TI - Genotyping and phenotyping the cytochrome p-450 enzymes. AB - With estimates of the percentage of pharmaceuticals that are subject to metabolism by the cytochrome P-450 enzymes (CYPs) in excess of 80%, the relative activities of these enzymes in various subpopulations and even in individual patients can have important ramifications in matters ranging from dose selection to prediction of toxicity to suitability of a new chemical entity (NCE) for continued drug development. The interindividual variation in CYP activities can be profound, and the differences may be due to environmental/physiologic factors, genetic factors, or both. With regard to the process of drug development, it would be useful to know as early in the development process as possible which CYPs are likely to process a NCE, the likely interindividual variation in the processing of a NCE by CYPs, which CYP activities are likely to be altered by a NCE, and the magnitude by which CYP activity is likely to be altered by a NCE. The latter two, in particular, will be useful in predicting drug interactions between the NCE and currently available drugs. For purposes of establishing treatment regimens that are maximally effective and minimally toxic, it follows that advance knowledge of probable CYP activities could be helpful. To the extent that phenotypic expression of CYP activity corresponds to CYP genotype, it may be possible a priori to design optimized therapeutic regimens for selective CYP substrates based on knowledge of a patient's CYP genotype. Because the expression of CYP activity is determined predominantly by prevailing environmental/physiologic conditions, tailoring drug therapy to meet individual patient needs can require knowledge of a patient's CYP phenotype. Strategies for genotyping and phenotyping CYP-450 activity are discussed with special attention paid to in vivo phenotyping methods. PMID- 12115021 TI - Furosemide: progress in understanding its diuretic, anti-inflammatory, and bronchodilating mechanism of action, and use in the treatment of respiratory tract diseases. AB - Accumulated experimental and clinical data suggest that adrenocorticosteroids and/or endogenous ouabain-like substances may play an important role in the mechanism of furosemide diuretic action. It was reported that the drug is highly bound in the adrenals, lungs, kidney, spleen, and liver. In patients with liver cirrhosis, furosemide exerted a markedly decreased natriuretic effect compared with normal subjects, and the plasma levels of circulating endothelin and atrial natriuretic factor (ANF) were significantly elevated. In neonates, after administration of furosemide, the urinary excretion of endothelin-1 and aldosterone increased markedly, and it is known that endothelin may release ANF and aldosterone in a dose-dependent manner. Furosemide was used to stimulate zona glomerulosa, whereas ANF decreased the production of steroids in zona glomerulosa and fasciculata cell culture owing to stimulation by various factors. Because the concomitant use of ANF and furosemide appeared to be diuretically effective in newborns after cardiac surgery, one may suggest that furosemide competes with ANF for its effects on the adrenals. Furosemide administered by inhalation exerted a protective effect on allergic and perennial nonallergic rhinitis and was effective in preventing the postsurgical recurrence of nasal polyposis. The drug can also be used as an antiasthmatic agent. In preterm ventilator-dependent infants with chronic lung disease, aerosolized furosemide improved pulmonary function with no marked effect on diuresis. In adults and children with asthma, furosemide exerted a protective effect against bronchoconstriction induced by several indirect stimuli similar to that of disodium cromoglycate or nedocromil. Aerosolized furosemide had a preventive effect also on bronchoconstriction induced by inhaled lysine acetylsalicylate in patients with aspirin-sensitive asthma. In high-dose beclomethasone-dependent asthma, inhaled lysine acetylsalicylate and furosemide exerted a mutually potentiating antiasthmatic activity, allowing considerable sparing of the inhaled steroid. It is proposed that this effect may be explained by the corticosteroid-sparing action of lysine released from the lysine acetylsalicylate molecule because similar beneficial effects were also obtained after the concomitant use of epsilon-aminocaproic acid (whose chemical structure is almost the same as that of lysine) and prednisone. Furosemide exhibited an anti-inflammatory effect through inhibition of production and release of cytokines interleukin (IL)-6, IL-8, and tumor necrosis factor alpha from peripheral mononuclear cells, which may have a beneficial effect on local inflamed tissue imbalance in the ratio of different cytokines, thus improving the sensitivity of target cells to endogenous glucocorticosteroids. PMID- 12115024 TI - HLA-Cw6 specificity and polymorphic residues are associated with susceptibility among Chinese psoriatics in Taiwan. AB - Previous serotyping in Chinese patients has failed to confirm an association between HLA-Cw6 and psoriasis. As serotyping has proved to be less valuable in the determination of HLA-C, genotyping of HLA-C in 68 Taiwanese psoriasis vulgaris (PSV) patients was performed using polymerase chain reaction/sequence specific oligonucleotide probe hybridization (PCR-SSOPH) of HLA-Cw genes. Compared to 213 non-PSV control subjects, HLA-Cw6 was significantly associated with PSV (16.18% of PSV patients vs 5.16% of controls; odds ratio 3.53, Pc/=4 mm) were present in 14 mice in the UV group as compared to only one mouse in the UV-MAL group. In mice treated on one side with MAL and the other side with vehicle, the delay in the appearance of tumour was only observed on the side treated with MAL, suggesting that a local rather than systemic effect was responsible for this phenomenon. In vivo fluorescence spectroscopy and quantitative fluorescence microscopy showed that there was a preferential accumulation of protoporphyrin IX in tumours as compared to adjacent UV-exposed skin and normal skin at the time of light exposure. In conclusion, topical MAL followed by light under suberythematous conditions delayed the appearance of UV induced skin tumours without increasing mortality or morbidity, and thus acted in a prophylactic manner. PMID- 12115028 TI - Okadaic acid-induced EGF receptor and MAP kinase activation does not require reactive oxygen species in primary keratinocytes. PMID- 12115029 TI - Genetic characterization of the Dyscalc locus. AB - Calcification occurs frequently in the development of atherosclerotic lesions, and studies in mice have indicated a genetic contribution. We now show that one genetic factor contributing to aortic calcification is the Dyscalc locus, previously shown to contribute to myocardial calcification. Thus, the Dyscalc locus, on proximal mouse Chromosome (Chr) 7, segregated with vascular calcification in a large cross between susceptible strain DBA/2J and resistant strain C57BL/6J. Further evidence was observed by analysis of recombinant inbred strains derived from various susceptible and resistant parental strains. Myocardial and vascular calcifications are importantly influenced by multiple modifier loci as well as the Dyscalc gene, making fine mapping of Dyscalc difficult. In order to allow more detailed genetic and biochemical characterization of Dyscalc, we have identified congenic strains containing the Dyscalc locus from resistant strain C57BL/10 on the background of susceptible strain C3H/DiSnA. The congenic strains exhibit little or no myocardial or vascular calcification, unlike the background HcB C3H strain, and the calcification segregated as a Mendelian factor, allowing finer mapping of Dyscalc. PMID- 12115031 TI - Identification of seven loci for static glucokinesis and dynamic glucokinesis in mice. AB - Non-insulin-dependent diabetes mellitus (NIDDM) is characterized by a breakdown of glucose homeostasis and is responsible for serious complications in various organs and vessels. Most of the genetic factors of NIDDM are yet unknown. Here, we identified two types of genetic factors that regulate homeostasis of blood glucose by measuring various pharmacokinetic parameters, some of which are used in the non-compartment analysis of drug metabolism in 340 F(2) progeny from the NIDDM model KK-A(y)/Ta Jcl mouse strain, and in non-diabetic PWK strain. We define "static glucokinesis" as the regulation of homeostasis that occurs during glucose deprivation, and "dynamic glucokinesis" as that during glucose stress; for instance, glucose tolerance test. Quantitative trait locus analysis revealed eight loci involved in the regulation of glucose homeostasis on chromosomes 7 ( Nidd1k), 2 ( Nidd2k), 1 ( Nidd3k, Nidd4k, and Nidd5k), 11 ( Nidd6k), 5 ( Nidd7k) (named Nidd1k through Nidd7k), and 4 ( Bwt1k). Nidd1k, Nidd4k, and Nidd7k were novel loci associated with NIDDM in mice. Nidd1k, Nidd2k, Nidd3k, and Nidd4k had linkage to factors characteristic of both static and dynamic glucokinesis. Nidd5k and Nidd6k showed linkage specific to markers of dynamic glucokinesis, and Nidd7k had linkage specific to static glucokinesis. Bwt1k was linked to obesity. Thus, the genetic factors for static glucokinesis and those for dynamic glucokinesis partially overlapped. PMID- 12115032 TI - Application of interval haplotype analysis facilitates efficient mapping of the mutation causing osteopetrosis in tl rats. PMID- 12115030 TI - Comparative sequence analysis in eight inbred strains of the metastasis modifier QTL candidate gene Brms1. PMID- 12115033 TI - Detailed chromosomal and radiation hybrid mapping in the proximal part of rat Chromosome 10 and gene order comparison with mouse and human. AB - The rat provides valuable and sometimes unique models of human complex diseases. To fully exploit the rat models in biomedical research, it is important to have access to detailed knowledge of the rat genome organization as well as its relation to the human genome. Rat Chromosome 10 (RNO10) harbors several important cancer-related genes. Deletions in the proximal part of RNO10 were repeatedly found in a rat model for endometrial cancer. To identify functional and positional candidate genes in the affected region, we used radiation hybrid (RH) mapping and single- and dual-color fluorescence in situ hybridization (FISH) techniques to construct a detailed chromosomal map of the proximal part of RNO10. The regional localization of 14 genes, most of them cancer-related ( Grin2a, Gspt1, Crebbp, Gfer, Tsc2, Tpsb1, Il9r, Il4, Irf1, Csf2, Sparc, Tp53, Thra1, Gh1), and of five microsatellite markers ( D10Mit10, D10Rat42, D10Rat50, D10Rat72, and D10Rat165) was determined on RNO10. For a fifteenth gene, Ppm1b, which had previously been assigned to RNO10, the map position was corrected to RNO6q12-q13. PMID- 12115034 TI - Comparative mapping of human Chromosome 19 with the chicken shows conserved synteny and gives an insight into chromosomal evolution. AB - Human Chromosome 19 (HSA19) is virtually completely sequenced. A complete physical contig map made up of BACs and cosmids is also available for this chromosome. It is, therefore, a rich source of information that we have used as the basis for a comparative mapping study with the chicken. Various orthologs of genes known to map to HSA19 have been mapped in the chicken. Five chicken microchromosomes (two of which were previously undefined) are seen to show conserved synteny with this chromosome, along with individual gene homologs on Chr 1 and another tiny microchromosome. Compared with the mouse, which has 12 chromosomal regions homologous to HSA19, the chicken genotype displays fewer evolutionary rearrangements. The ancestral nature of the chicken karyotype is demonstrated and may prove to be an excellent tool for studying genome evolution. PMID- 12115035 TI - An extensive and comprehensive radiation hybrid map of bovine Chromosome 15: comparison with human Chromosome 11. AB - An extensive and comprehensive radiation hybrid map of bovine Chromosome 15 (BTA15) was built with 42 anonymous markers, 3 ESTs, and 49 genes. This work allows us to refine the comparative map between human Chromosome (Chr) 11 (HSA11) and BTA15. Four blocks with a similar gene content and relatively good gene order conservation were identified. The discrepancies are concentrated on closely positioned genes for which discrimination is not possible between mapping resolution limits in either the human or the bovine maps and true local inversions. Using the gene order similarity and the human physical map as starting point, we estimated the overall physical length of BTA15 to be around 75.3 Mb. The INRA bovine BAC library was screened for all the markers ordered on the bovine map, which will provide anchors for future efforts in the construction of a physical map of the bovine genome.Finally, this map contains the majority of publicly available polymorphic markers described for BTA15 and integrates those with comparative mapping information. It should, therefore, constitute a powerful tool in the identification of relevant candidate genes in regions of BTA15 harboring economic trait loci. PMID- 12115036 TI - A radiation hybrid map for the bovine Y Chromosome. AB - Screening a bovine Y Chromosome-specific DNA library resulted in 34 new microsatellites, six of which mapped to the pseudoautosomal region (PAR), and 28 localized to the Y-specific region. These microsatellites, together with 23 markers previously mapped to the bovine Y Chr, were scored on a 7000-rad cattle hamster radiation hybrid (RH) panel. Retention frequency of individual markers ranged from 18.5% to 76.5% with an average of 48.4%. Markers with high retention frequency (>55%) were found to exist in multiple copies on the Y Chr. Thirteen markers were placed on the PAR RH map with the AmelY gene proximal to the pseudoautosomal boundary and 46 markers, including Sry and Tspy gene, on the Y specific region of the RH map. The microsatellites developed and mapped in this work will be useful for comparative mapping of cattle, sheep, and goat, studying the origin, evolution, and migration of bovidae species and provide an initial platform to develop a high-resolution map of the Y Chr and positional cloning of Y-specific genes. PMID- 12115037 TI - Computational analysis of composite regulatory elements. AB - Combinatorial regulation is a powerful mechanism for generating specificity in gene expression, and it is thought to play a pivotal role in the formation of the complex gene regulatory networks found in higher eukaryotes. The term "Composite Element" (CE) refers to a minimal functional unit where protein-DNA and protein protein interactions contribute to a highly specific pattern of gene transcriptional regulation. Identification of composite elements will help to better understand gene regulation networks. Experimentally identified CEs are limited in number, and the currently available CE database COMPEL is based on such published information. Here, based on the statistical analysis of over represented adjacent transcription factor binding sites, we describe a computational method to predict composite regulatory elements in genomic sequences. The algorithm proved to be efficient for extracting composite elements that had been experimentally confirmed and documented in the COMPEL database. Furthermore, putative new composite elements are predicted based on this method, and we have been able to confirm some of our predictions which are not included in the COMPEL database by searching published information. PMID- 12115038 TI - Assessment of sediments in complex freshwater river systems. PMID- 12115039 TI - Assessment of injury to fish and wildlife resources in the Grand Calumet River and Indiana Harbor Area of Concern, USA. AB - This article is the second in a series of three that describes the results of a Natural Resource Damage Assessment (NRDA) conducted in the Grand Calumet River and Indiana Harbor Area of Concern (IHAOC). The assessment area is located in northwest Indiana and was divided into nine reaches to facilitate the assessment. This component of the NRDA was undertaken to determine if fish and wildlife resources have been injured due to exposure to contaminants that are associated with discharges of oil or releases of other hazardous substances. To support this assessment, information was compiled on the chemical composition of sediment and tissues; on the toxicity of whole sediments, pore water, and elutriates to fish; on the status of fish communities; and on fish health. The data on each of these indicators were compared to regionally relevant benchmarks to assess the presence and extent of injury to fish and wildlife resources. The results of this assessment indicate that injury to fish and wildlife resources has occurred throughout the assessment area, with up to five distinct lines of evidence demonstrating injury within the various reaches. Based on the frequency of exceedance of the benchmarks for assessing sediment and tissue chemistry data, total polychlorinated biphenyls is the primary bioaccumulative contaminant of concern in the assessment area. It is important to note, however, that this assessment was restricted by the availability of published bioaccumulation-based sediment quality guidelines, tissue residue guidelines, and other benchmarks of sediment quality conditions. The availability of chemistry data for tissues also restricted this assessment in certain reaches of the assessment area. Furthermore, insufficient information was located to facilitate identification of the substances that are causing or substantially contributing to effects on fish (i.e., sediment toxicity, impaired fish health, or impaired fish community structure). Therefore, substances not included on the list of COCs cannot necessarily be considered to be of low priority with respect to sediment injury (e.g., metals, polycyclic aromatic hydrocarbons, alkanes, alkenes, organochlorine pesticides, phthalates, dioxins, and furans, etc.). PMID- 12115040 TI - An assessment of injury to sediments and sediment-dwelling organisms in the Grand Calumet River and Indiana Harbor Area of Concern, USA. AB - This article is the first in a series of three that describe the results of a Natural Resource Damage Assessment (NRDA) conducted in the Grand Calumet River and Indiana Harbor Area of Concern (IHAOC). The assessment area is located in northwest Indiana and was divided into nine reaches to facilitate the assessment. This component of the NRDA was undertaken to determine if sediments and sediment dwelling organisms have been injured due to exposure to contaminants that have accumulated in sediments as a result of discharges of oil or releases of other hazardous substances from industrial, municipal, and nonpoint sources. To support this assessment, information was compiled on the chemical composition of sediment and pore water; on the toxicity of whole sediments, pore water, and elutriates; and on the status of benthic invertebrate communities. The data on each of these indicators were compared to regionally relevant benchmarks to assess the presence and extent of injury to surface water resources ( i.e., sediments) or biological resources ( i.e., sediment-dwelling organisms). The results of this assessment indicate that sediment injury has occurred throughout the assessment area, with up to four distinct lines of evidence demonstrating injury within the various reaches. The primary contaminants of concern ( i.e., those substances that are present at concentrations that are sufficient to cause or substantially contribute to sediment injury) include metals, polycyclic aromatic hydrocarbons, and total polychlorinated biphenyls. PMID- 12115041 TI - Toxicity assessment of sediments from the Grand Calumet River and Indiana Harbor Canal in Northwestern Indiana, USA. AB - The objective of this study was to evaluate the toxicity of sediments from the Grand Calumet River and Indiana Harbor Canal located in northwestern Indiana, USA. Toxicity tests used in this assessment included 10-day sediment exposures with the amphipod Hyalella azteca, 31-day sediment exposures with the oligochaete Lumbriculus variegatus, and the Microtox Solid-Phase Sediment Toxicity Test. A total of 30 sampling stations were selected in locations that had limited historic matching toxicity and chemistry data. Toxic effects on amphipod survival were observed in 60% of the samples from the assessment area. Results of a toxicity test with oligochaetes indicated that sediments from the assessment area were too toxic to be used in proposed bioaccumulation testing. Measurement of amphipod length after the 10-day exposures did not provide useful information beyond that provided by the survival endpoint. Seven of the 15 samples that were identified as toxic in the amphipod tests were not identified as toxic in the Microtox test, indicating that the 10-day H. azteca test was more sensitive than the Microtox test. Samples that were toxic tended to have the highest concentrations of metals, polycyclic aromatic hydrocarbons (PAHs), and polychlorinated biphenyls (PCBs). The toxic samples often had an excess of simultaneously extracted metals (SEM) relative to acid volatile sulfide (AVS) and had multiple exceedances of probable effect concentrations (PECs). Metals may have contributed to the toxicity of samples that had both an excess molar concentration of SEM relative to AVS and elevated concentrations of metals in pore water. However, of the samples that had an excess of SEM relative to AVS, only 38% of these samples had elevated concentration of metals in pore water. The lack of correspondence between SEM-AVS and pore water metals indicates that there are variables in addition to AVS controlling the concentrations of metals in pore water. A mean PEC quotient of 3.4 (based on concentrations of metals, PAHs, and PCBs) was exceeded in 33% of the sediment samples and a mean quotient of 0.63 was exceeded in 70% of the thirty sediment samples from the assessment area. A 50% incidence of toxicity has been previously reported in a database for sediment tests with H. azteca at a mean quotient of 3.4 in 10-day exposures and at a mean quotient of 0.63 in 28-day exposures. Among the Indiana Harbor samples, most of the samples with a mean PEC quotient above 0.63 ( i.e., 15 of 21; 71%) and above 3.4 ( i.e., 10 of 10; 100%) were toxic to amphipods. Results of this study and previous studies demonstrate that sediments from this assessment area are among the most contaminated and toxic that have ever been reported. PMID- 12115042 TI - Suppression of pyrite oxidation by iron 8-hydroxyquinoline. AB - One of the important approaches to prevent pyrite (FeS(2)) oxidation and subsequent formation of acid mine drainage (AMD) is to create a surface coating on pyrite. In this study, a coating of iron 8-hydroxyquinoline was formed by leaching pyrite with a 0.10 M H(2)O(2)/0.0034 M 8-hydroxyquinoline solution; stability of the coated pyrite was tested under various pH and temperature conditions. The results showed that iron 8-hydroxyquinoline coating could significantly suppress further pyrite oxidation by both chemical (H(2)O(2)) and biological ( e.g., Thiobacillus ferrooxidans) processes. At pH from 3.0 to 5.0 and temperature from 10-40 degrees C, the amount of SO(4)(2-) leached out by 0.10 M H(2)O(2) from the coated pyrite samples was 54.8-70.1% less than that from the uncoated controls. The oxidation of pyrite followed a pseudo-zero-order kinetics under the constant concentration of H(2)O(2). In the presence of microorganisms, sulfate leached out of the uncoated pyrite in 1 year accounted for 5.32% of the total pyrite in the system, with a concurrent pH drop to 2.35 under the ambient room temperatures. In contrast, the amount leached out from the coated samples was only 0.15% of the total pyrite and the final pH was 5.48. Thus, the coating decreased the leachability of pyrite by 97% in the inoculated systems. In comparison to the more widely studied iron phosphate coating, the advantage of iron 8-hydroxyquinoline coating was that it inhibited both chemical and biological pyrite oxidation, whereas iron phosphate coating could only inhibit chemical pyrite oxidation. PMID- 12115043 TI - Characterization of estrogenic activity of riverine sediments from the Czech Republic. AB - Extracts of sediments from rivers in an industrialized area in the Czech Republic were used to evaluate suitability of a simple in vitro bioassay system to detect estrogen receptor (ER)-mediated activity in the complex mixture. Total estrogenic activity was detected by measuring luciferase activity in a stably transfected cell line containing an estrogen-responsive element linked to a luciferase reporter gene. For appropriate interpretation of ER-mediated activity, the effect of sediment extracts on the cell cytotoxicity was assessed at the same time. All sediment samples elicited considerable estrogenic activity. Fractionation of the extracts along with bioassay testing and subsequent instrumental analysis allowed the estrogenic fractions to be identified. The Florisil fraction, which was intermediate in polarity, was the most estrogenic. Instrumental analysis documented that the concentration of the degradation products of alkylphenol ethoxylates did not occur at sufficient concentrations to account for the estrogenic activity. Mass-balance calculations and testing of fractions confirmed that certain polycyclic aromatic hydrocarbons (PAHs) or their metabolites were the most likely compounds contributing to estrogenicity. Some other compounds, such as PCNs and PAH derivatives, that were present in the first and second fraction were tested for their potential estrogenic activity. Their ER-mediated activity and contribution to the overall responses of the complex extracts were very low. The concentrations of 17beta-estradiol present in the bioassay media was an important factor for the evaluation of (anti)estrogenicity of single compound(s) or complex mixtures. PMID- 12115044 TI - Incorporation of toxicity tests into the Turkish industrial discharge monitoring systems. AB - A toxicity evaluation is an important parameter in wastewater quality and in the monitoring of discharged effluents. Some organic and inorganic compounds at toxic levels have been detected in industrial discharges, resulting in plant upsets and discharge permit violations. In some cases, even though the effluent does not exceed the chemical-specific discharge limits, the results of toxicity tests show potential toxicity. Knowledge of the toxicity of effluents can benefit treatment plant operators in optimizing plant operation, protecting receiving water quality, and establishing sewer discharge permits. In the Turkish regulations only toxicity dilution factor with fish is part of the toxicity monitoring program of permissible wastewater discharge. This study investigated the acute toxicity of pulp-paper, leather, and petrochemical industry wastewaters by traditional and enrichment toxicity tests and emphasized the importance of toxicity tests in wastewater discharge regulations. The enrichment toxicity tests are novel applications indicating whether there is potential toxicity or stimulation conditions. Different organisms were used, including bacteria (Floc and Coliform bacteria) algae (Chlorella sp.), fish (Lepistes sp.), and protozoan species (Vorticella sp.) to represent four trophic levels. The toxicity test results were compared with chemical analyses to identify the pollutants responsible for the toxicity in the effluent wastewater samples. Toxicity of the effluents could not be explained by using physicochemical analyses in five cases for the leather and four cases for the pulp-paper and petrochemical industries. The results clearly showed that the use of bioassay tests produce additional information about the toxicity potential of industrial discharges and effluents. PMID- 12115045 TI - Fate and effects of the triazinone herbicide metribuzin in experimental pond mesocosms. AB - Metribuzin is a triazinone herbicide that is widely used for the control of grasses and broad-leaved weeds in soybeans, sugarcane, and numerous other crops. Metribuzin is highly toxic to freshwater macrophytes and algae under laboratory conditions (median plant EC(50) = 31 microg/L; n = 11 species) but has not been studied under controlled outdoor conditions. We conducted a 6-week study to examine the aquatic fate and effects of metribuzin in 0.1-ha outdoor aquatic mesocosms. Mesocosms (n = 2 per treatment) were treated with metribuzin at one of five concentrations: 0, 9, 19, 38, or 75 microg/L. Concentrations were selected to bracket known laboratory effect concentrations and to reflect calculated edge of-field concentrations. The dissipation half-life of metribuzin in water was 5 days. Metribuzin had no statistically significant effects on water quality, periphyton biomass, macrophyte biomass, macrophyte species composition, fish survival, or fish growth at treatment levels ranging up to and including 75 microg/L. Although metribuzin is highly toxic to freshwater macrophytes and algae under laboratory conditions, it poses little risk to nontarget aquatic plants due to the short aqueous dissipation half-life. The findings also demonstrate that current herbicide risk assessment procedures used in the registration process could benefit from empirical assessments of the fate of chemicals under realistic environmental conditions. PMID- 12115046 TI - Effects of metals on seed germination, root elongation, and coleoptile and hypocotyl growth in Triticum aestivum and Cucumis sativus. AB - A simple, fast, and easy-to-perform method was carried out for the quantification of the inhibitory effects of metals on wheat and cucumber. The method uses seed germination, root elongation, and hypocotyl and coleoptile growth in these plants as parameters in the presence of varying concentrations of metals. Metals selected for this study were Hg, Cd, Co, Cu, Pb, and Zn. Although effective concentrations of these metals for a certain degree of inhibition were different, both plants had a reduced seed germination rate, root, and hypocotyl or coleoptile length with increasing concentrations. Mercury was determined to be the most inhibitory metal on these parameters. This metal caused a complete inhibition of germination in wheat and cucumber seeds at certain concentrations- >or=1.5 mM in cucumber and at 1.7 mM in wheat. No other metal caused this kind of inhibition even at the highest concentration (8.0 mM) applied. Though this metal possessed a higher inhibition of germination in cucumber than in wheat seeds, the inhibitory effects of other metals used were the reverse, being higher in wheat. With some exceptions, all metals in selected concentrations caused a significant ( p < 0.01 or p < 0.05) decrease in germination rate of both plants compared to control group seeds. PMID- 12115047 TI - Lethal body residue of chlorophenols and mixtures of chlorophenols in benthic organisms. AB - The lethal body residue (LBR) of a few chlorophenol congeners were measured in the oligochaete worm Lumbriculus variegatus, and the LBR of pentachlorophenol was measured also in a midge, Chironomus riparius larvae. LBR is defined as the concentration of the compound in the organism, on molar basis, to cause death, and the LBR(50) is defined as the calculated LBR value to cause a 50% mortality in population after a given time. Groups of 30 or 40 organisms were exposed to different chlorophenol concentrations in artificial soft fresh water to achieve differential mortality. Exposure times were either 24 h or 48 h. In addition to exposures with individual congeners, mixtures of chlorophenols were also tested. After each exposure, the surviving organisms were collected and the body burden of chlorophenols was measured by gas chromatography with electron capture detection. The measured body burden was related to the percent mortality in the group. The trichlorophenols and pentachlorophenol have a 48-h LBR(50) of 0.45 0.66 micromol/g wet weight in L. variegatus. The 48-h LBR(50) of pentachlorophenol for C. riparius was 0.15 micromol/g wet weight, indicating a slight difference in the sensitivity of these two species. The 48-h LBR(50) of 2,3,4,6-tetrachlorophenol is 0.91 micromol/g wet weight, and the value for 2,6 dichlorophenol is 1.2 micromol/g wet weight in L. variegatus. The 48-h LBR(50)s of the chlorophenol mixtures ranged from 0.50 to 0.83 micromol/g wet weight, demonstrating an additive toxicity. PMID- 12115048 TI - Bioavailability and toxicokinetics of (14)C-lindane (gamma-HCH) in the enchytraeid Enchytraeus albidus in two soil types: the aging effect. AB - Enchytraeids are important members of the soil fauna living in the true soil layer instead of the humus like most earthworms, resulting in a different interaction with chemicals in soil. It is well known that the detectable concentration of contaminants in the soil cannot fully predict a biological effect; therefore bioaccumulation studies are of great importance. In this study the bioaccumulation pattern of the pesticide lindane ([(14)C]gamma-HCH) in Enchytraeus albidus was analyzed in two different soil types, an artificial Organisation for Economic Co-operation and Development (OECD) soil and a natural agricultural soil. The effect of aging on bioaccumulation and bioavailability was also analyzed. Aging experiments consisted of having the same procedures at different times after soil contamination: Immediately after contamination and 1, 2, 4, 6, and 10 months after that. Major differences occurred within the first month. Considering the overall data (10 months), it is possible to observe that in OECD soil there is a correlation between concentration in soil and in the organisms, but that was not observed for the natural soil. The decrease patterns for concentration in soil and in watery soil extracts were different: monophasic behavior occurred in OECD soil, and a biphasic pattern was measured in natural soil. It was observed that the bioavailable ( i.e., water-extractable) fraction of lindane in the natural soil decreased faster than the total soil concentration. In natural soil the decrease of concentration appeared faster than in OECD soil; this may be related to the lower amount of organic matter content. PMID- 12115049 TI - Toxicity of select beta adrenergic receptor-blocking pharmaceuticals (B-blockers) on aquatic organisms. AB - One class of pharmaceutical compounds identified in U.S. and European waters are the B-adrenergic receptor blocking compounds (B-blockers). However, little information is available on the potential aquatic toxicity of these compounds. Therefore, Hyalella azteca, Daphnia magna, Ceriodaphnia dubia, and Oryias latipes (Japanese medaka) were exposed to metoprolol, nadolol, and propranolol to determine potential toxicity. Average 48-h LC(50) for propranolol to H. azteca was 29.8 mg/L. The no-observed-effects concentration (NOEC) and lowest-observed effects concentration (LOEC) for propranolol affecting reproduction of H. azteca were 0.001 and 0.1 mg/L, respectively. The average propranolol and metoprolol 48 h LC(50)s for D. magna were 1.6 and 63.9 mg/L, respectively. C. dubia 48-h LC(50)s were 0.85 and 8.8 mg/L for propranolol and metoprolol, respectively. The NOEC and LOEC of propranolol affecting reproduction in C. dubia were 0.125 and 0.25 mg/L, respectively. In O. latipes, the propranolol 48-h LC(50) was 24.3 mg/L. Medaka growth was decreased at 0.5 mg/L propranolol. A 2-week medaka reproductive study indicated significant changes in plasma steroid levels; however, no changes in the average number of eggs produced or number of viable eggs which hatched was observed. In a 4-week follow-up propranolol exposure, the total number of eggs produced by medaka and the number of viable eggs that hatched were decreased at concentrations as low as 0.5 microg/L. Based on this study and the expected aqueous environmental exposure levels, adverse effects of propranolol to invertebrate populations is unlikely; however, further reproductive studies are need to elucidate the risk to teleosts. PMID- 12115050 TI - Relation between growth and the heavy metal concentration in organs of bream Abramis brama L. populating Lake Balaton. AB - The effect of growth and physical condition on the level of heavy metals accumulated in the organs of common bream (Abramis brama L.) populating Lake Balaton was investigated on samples collected in October 1999 and May 2000 from two well-separable sites regarding their trophic state and pollution impact (western and eastern basins). The average metal concentrations in the organs of fish varied in the following ranges: Cd, 0.39-1.98; Cu, 1.73-57.3; Hg, 0.02-0.13; Pb, 0.39-3.15; and Zn, 12.7-159.3 microg/g dry weight. The highest Cd, Cu, Pb and Zn concentrations were detected in the gill and liver of fish, whereas the highest Hg concentrations were measured in the muscle. The maximum metal concentrations in the muscle of bream were on average below the maximum permissible levels for human consumption. Significant positive correlation was found among the heavy metal load of bream and their instantaneous growth rate; hardly any connection was observed related to the physical condition of samples. The relatively low metal concentrations of the ambient water and their poor correlation with the heavy metal load of bream, indicates that for the mature stages of this fish species the metal uptake from food is predominant, and thus the heavy metal load of fish reflect more the pollution state of the sediment and its biota, rather than that of the ambient water. PMID- 12115051 TI - Global distribution of halogenated dimethyl bipyrroles in marine mammal blubber. AB - Four halogenated dimethyl bipyrroles (HDBPs), hypothesized to be naturally produced, were quantitated in marine mammal blubber from a number of species obtained from various locations worldwide. HDBPs were found in samples from all locations studied. Concentrations of total HDBPs (SigmaHDBPs) ranged from 0.4 ng/g lipid weight in ringed seals (Phoca hispida) from the White Sea to 2,540 ng/g lipid weight in Dall's porpoise (Phocoenoides dalli) from the northwestern North Pacific Ocean. At their highest levels, SigmaHDBPs made up 11% of the total quantitated organohalogen body burden of adult male Dall's porpoises. In two beluga (Delphinapterus leucas) data subsets, it was found that males contained significantly higher concentrations of SigmaHDBPs than females. No significant effects of age or sex on SigmaHDBPs were observed in harbor seal (Phoca vitulina) and bowhead whale (Balaena mysticetus) data subsets. The geographical distribution of concentrations did not resemble that of the ubiquitous anthropogenic organohalogen, polychlorinated biphenyl congener CB-153. Higher concentrations of HDBPs and different patterns of congeners were observed in samples from Pacific as opposed to non-Pacific Ocean influenced environments. Concentrations of HDBPs in beluga from the Arctic and St. Lawrence River were similar. Their high abundance in north Pacific Ocean biota and widespread occurrence suggest that HDBPs undergo extensive transport from a source located primarily in the Pacific Ocean. Evidence from HDBP congener patterns indicates that both ocean currents and atmospheric transport likely play a role in the movement of HDBPs. These results imply that HDBPs and anthropogenic organohalogens have different sources and support the natural production hypothesis. PMID- 12115052 TI - Assembly of membrane-bound respiratory complexes by the Tat protein-transport system. AB - The Tat protein-export system serves to translocate folded proteins, often containing redox cofactors, across the bacterial inner membrane. Substrate proteins are directed to the Tat apparatus by distinctive N-terminal signal peptides containing a consensus SRRxFLK 'twin-arginine' motif. Here we review recent studies of the Tat system with particular emphasis on the assembly of membrane-bound respiratory complexes. We discuss the connection between Tat targeting and topological organisation of the complexes and consider the role of chaperone proteins in cofactor insertion and Tat targeting. The crystal structure of Escherichia coli formate dehydrogenase-N demonstrates that some Tat substrates are integral membrane proteins. Sequence analysis suggests that one-quarter of all traffic on the E. coli Tat pathway is inner-membrane proteins. PMID- 12115053 TI - Genetics and control of CO(2) assimilation in the chemoautotroph Ralstonia eutropha. AB - The nutritional versatility of facultative autotrophs requires efficient overall control of their metabolism. Most of these organisms are Proteobacteria that assimilate CO(2) via the highly energy-demanding Calvin-Benson-Bassham reductive pentose-phosphate cycle. The enzymes of the cycle are encoded by cbb genes organized in cbb operons differing in size and composition, although conserved features are apparent. Transcription of the operons, which may form regulons, is strictly controlled, being induced during autotrophic but repressed to varying extents during heterotrophic growth of the bacteria. The chemoautotroph Ralstonia eutropha is one of the organisms studied extensively for the mechanisms involved in the expression of cbb gene systems. CbbR is a LysR-type transcriptional regulator and the key activator protein of cbb operons. The cbbR gene is typically located adjacent and in divergent orientation to its cognate operon. The activating function of CbbR seems to be modulated by metabolites signaling the nutritional state of the cell to the cbb system. Phosphoenolpyruvate is such a signal metabolite acting as a negative effector of R. eutropha CbbR, whereas NADPH has been proposed to be a coactivator of the protein in two other chemoautotrophs, Xanthobacter flavus and Hydrogenophilus thermoluteolus. There is evidence for the participation of additional regulators in cbb control. In the photoautotrophs Rhodobacter capsulatus and Rhodobacter sphaeroides, response regulator RegA of the global two-component signal transduction system RegBA serves this function. It is conceivable that specific variants of cbb control systems have evolved to ensure their optimal integration into regulatory networks operating in the diverse autotrophs characterized by different metabolic capabilities. PMID- 12115054 TI - Role of SdiA in Salmonella enterica serovar Typhimurium physiology and virulence. AB - sdiAin Salmonella enterica serovar Typhimurium encodes a protein belonging to the LuxR family of transcriptional regulators. Initial computer analysis revealed the presence of a fur box 19 bp upstream of the start codon of the sdiA gene and a helix-turn-helix motif in the carboxy-terminal part of the SdiA protein typical for transcriptional regulators. Deletion of the furbox resulted in twofold increase of sdiA transcription. Furthermore, addition of dipyridyl, an iron chelator, to culture media increased sdiA transcription to the level observed in the fur box mutant, confirming that sdiA is suppressed in the presence of iron. When S. entericasv. Typhimurium was grown in conditioned medium, sdiA transcription was repressed to 30% of that in cells grown in fresh LB broth; this repression was independent of the fur box. Oral infection of mice with the strain lacking the helix-turn-helix domain of SdiA indicated increased virulence of this S. entericasv. Typhimurium mutant. sdiA, dually controlled by iron concentration and culture-density-derived signals, may therefore play an important role in S. entericasv. Typhimurium virulence regulation. PMID- 12115055 TI - Cloning and analysis of the simocyclinone biosynthetic gene cluster of Streptomyces antibioticus Tu 6040. AB - The biosynthetic gene cluster of the aminocoumarin antibiotic simocyclinone D8 was cloned by screening a cosmid library of Streptomyces antibioticusTu 6040 with a heterologous probe from a gene encoding a cytochrome P450 enzyme involved in the biosynthesis of the aminocoumarin antibiotic novobiocin. Sequence analysis of a 39.4-kb region revealed the presence of 38 ORFs. Six of the identified ORFs showed striking similarity to genes from the biosynthetic gene clusters of the aminocoumarin antibiotics novobiocin and coumermycin A(1). Simocyclinone also contains an angucyclinone moiety, and 12 of the ORFs showed high sequence similarity to biosynthetic genes of other angucyclinone antibiotics. Possible functions within the biosynthesis of simocyclinone D8 could be assigned to 23 ORFs by comparison with sequences in GenBank. Experimental proof for the function of the identified gene cluster was provided by a gene inactivation experiment, which resulted in the abolishment of the formation of the aminocoumarin moiety of simocyclinone. Feeding of the mutant with the aminocoumarin moiety of novobiocin led to a new, artificial simocyclinone derivative. PMID- 12115056 TI - Deletion analysis of the C-terminal region of the alpha-amylase of Bacillus sp. strain TS-23. AB - The alpha-amylase from Bacillus sp. strain TS-23 is a secreted starch hydrolase with a domain organization similar to that of other microbial alpha-amylases and an additional functionally unknown domain (amino acids 517-613) in the C-terminal region. By sequence comparison, we found that this latter domain contained a sequence motif typical for raw-starch binding. To investigate the functional role of the C-terminal region of the alpha-amylase of Bacillus sp. strain TS-23, four His(6)-tagged mutants with extensive deletions in this region were constructed and expressed in Escherichia coli. SDS-PAGE and activity staining analyses showed that the N- and C-terminally truncated alpha-amylases had molecular masses of approximately 65, 58, 54, and 49 kDa. Progressive loss of raw-starch-binding activity occurred upon removal of C-terminal amino acid residues, indicating the requirement for the entire region in formation of a functional starch-binding domain. Up to 98 amino acids from the C-terminal end of the alpha-amylase could be deleted without significant effect on the raw-starch hydrolytic activity or thermal stability. Furthermore, the active mutants hydrolyzed raw corn starch to produce maltopentaose as the main product, suggesting that the raw-starch hydrolytic activity of the Bacillus sp. strain TS-23 alpha-amylase is functional and independent from the starch-binding domain. PMID- 12115057 TI - A novel streptomycete lipase: cloning, sequencing and high-level expression of the Streptomyces rimosus GDS(L)-lipase gene. AB - An extracellular lipase from Streptomyces rimosus R6-554W has been recently purified and biochemically characterized. In this report the cloning, sequencing, and high-level expression of its gene is described. The cloned DNA contained an ORF of 804 bp encoding a 268-amino-acid polypeptide with 34 amino acid residues at the amino terminus of the sequence that were not found in the mature protein. The theoretical molecular mass (24.172 kDa) deduced from the amino acid sequence of the mature enzyme was experimentally confirmed. This lipase showed no overall amino acid sequence similarity to other lipases in the databases. However, two hypothetical proteins, i. e. putative hydrolases, derived from the genome sequencing data of Streptomyces coelicolor A3(2), showed 66% and 33% identity. In addition, a significant similarity to esterases from Streptomyces diastatochromogenes and Aspergillus terreus was found. Sequence analysis revealed that our novel S. rimosus lipase containing a GDS(L)-like consensus motif belongs to family II of lipolytic enzymes, previously unrecognized in Streptomyces. When the lipase gene was expressed in a S. rimosus lipase-deficient strain harboring the lipase gene on a high-copy-number vector, lipase activity was 22-fold higher than in the original strain. PMID- 12115058 TI - Phylogeny of green sulfur bacteria on the basis of gene sequences of 16S rRNA and of the Fenna-Matthews-Olson protein. AB - The phylogeny of green sulfur bacteria was studied on the basis of gene sequences of the 16S rRNA and of the Fenna-Matthews-Olson (FMO) protein. Representative and type strains (31 strains total) of most of the known species were analyzed. On the basis of fmoA gene sequences from Chlorobium tepidum ATCC 49652(T) and Chlorobium limicola DSM 249(T) available from the EMBL database, primers were constructed that allowed sequence analysis of a major part of the fmoAgene. The largely congruent phylogenetic relationship of sequences of the fmoA gene and of 16S rDNA gives considerable support to the phylogeny of green sulfur bacteria previously suggested on the basis of 16S rDNA sequences. Distinct groups of strains were recognized on the basis of 16S rDNA and FMO sequences and supported by characteristic signature amino acids of FMO. Marine strains formed clusters separate from freshwater strains. The resulting phylogenetic grouping and relationship of the green sulfur bacteria do not correlate with their current taxonomic classification. PMID- 12115059 TI - A chimeric Anabaena/ Escherichia coli KdpD protein (Anacoli KdpD) functionally interacts with E. coli KdpE and activates kdp expression in E. coli. AB - The kdpFABC operon, coding for a high-affinity K(+)-translocating P-type ATPase, is expressed in Escherichia coli as a backup system during K(+) starvation or an increase in medium osmolality. Expression of the operon is regulated by the membrane-bound sensor kinase KdpD and the cytosolic response regulator KdpE. From a nitrogen-fixing cyanobacterium, Anabaena sp. strain L-31, a kdpDgene was cloned (GenBank accession no. AF213466) which codes for a KdpD protein (365 amino acids) that lacks both the transmembrane segments and C-terminal transmitter domain and thus is shorter than E. coli KdpD. A chimeric kdpD gene was constructed and expressed in E. coli coding for a protein (Anacoli KdpD), in which the first 365 amino acids of E. coli KdpD were replaced by those from Anabaena KdpD. In everted membrane vesicles, this chimeric Anacoli KdpD protein exhibited activities, such as autophosphorylation, transphosphorylation and ATP-dependent dephosphorylation of E. coli KdpE, which closely resemble those of the E. coli wild-type KdpD. Cells of E. coli synthesizing Anacoli KdpD expressed kdpFABC in response to K(+) limitation and osmotic upshock. The data demonstrate that Anabaena KdpD can interact with the E. coliKdpD C-terminal domain resulting in a protein that is functional in vitro as well as in vivo. PMID- 12115061 TI - Diversity of rhizobia from nodules of the leguminous tree Gliricidia sepium, a natural host of Rhizobium tropici. AB - The Rhizobium species that nodulate the legume tree Gliricidia sepium were analyzed by phenotypic characteristics (including nodule formation in different hosts), PCR-RFLP patterns and sequences of 16S rRNA genes, multilocus enzyme electrophoresis, and plasmid patterns. Strains of Rhizobium tropici type A and B, Sinorhizobium spp., and Rhizobium etli bv. phaseoli were encountered in G. sepium nodules and their presence depended on the site sampled. PMID- 12115060 TI - The role of the fatty acid beta-oxidation multienzyme complex from Pseudomonas oleovorans in polyhydroxyalkanoate biosynthesis: molecular characterization of the fadBA operon from P. oleovorans and of the enoyl-CoA hydratase genes phaJ from P. oleovorans and Pseudomonas putida. AB - In order to investigate the role of the putative epimerase function of the beta oxidation multienzyme complex (FadBA) in the provision of (R)-3-hydroxyacyl-CoA thioesters for medium-chain-length polyhydroxyalkanoate (PHA(MCL)) biosynthesis, the fadBA(Po) operon of Pseudomonas oleovorans was cloned and characterized. The fadBA(Po) operon and a class-II PHA synthase gene of Pseudomonas aeruginosa were heterologously co-expressed in Escherichia coli to determine whether the putative epimerase function of FadBA(Po) has the ability to provide precursors for PHA accumulation in a non-PHA-accumulating bacterium. Cultivation studies with fatty acids as carbon source revealed that FadBA(Po) did not mediate PHA(MCL) biosynthesis in the E. coli wild-type strain harboring a PHA synthase gene. However, PHA accumulation was strongly impaired in a recombinant E. coli fadB mutant, which harbored a PHA synthase gene. These data indicate that in pseudomonads FadBA does not possess the inherent property, based on a putative epimerase function, to provide the ( R)-enantiomer of 3-hydroxyacyl-CoA efficiently and that other linking enzymes are required to efficiently channel intermediates of beta-oxidation towards PHA(MCL) biosynthesis. However, the phaJ gene from P. oleovorans and from Pseudomonas putida, both of which encoded a 3- Re enoyl-CoA hydratase, was identified. The co-expression of phaJ(Po/Pp) with either a class-II PHA synthase gene or the PHA synthase gene from Aeromonas punctata in E. coli revealed that PhaJ(Po/Pp) mediated biosynthesis of either PHA(MCL), contributing to about 1% of cellular dry mass, or of poly(3 hydroxybutyrate- co-3-hydroxyhexanoate), contributing to 3.6% of cellular dry mass, when grown on decanoate. These data indicate that FadBA(Po)does not mediate the provision of (R)-3-hydroxyacyl-CoA, which resembles FadBA of non-PHA accumulating bacteria, and that 3- Re enoyl-CoA hydratases are required to divert intermediates of fatty acid beta-oxidation towards PHA biosynthesis in P. oleovorans. PMID- 12115062 TI - The diagnostic criteria for primary osteoporosis and the incidence of osteoporosis in China. PMID- 12115063 TI - Molecular pathogenesis of tumorigenesis in sporadic parathyroid adenomas. PMID- 12115064 TI - Effects of skeletal loading on bone mass and compensation mechanism in bone: a new insight into the "mechanostat" theory. AB - We have suggested that: (i) osteocalcin carboxylation is related to bone material properties (bone quality), and (ii) impairment of bone material properties could be compensated by bone mass increase. The aim of the present prospective study was to investigate whether the effects of skeletal loading on bone mass were associated with the compensation mechanism between bone mass and bone material properties. The subjects were 56 healthy female volunteers aged around 50 years. They were classified into pre- and postmenopausal groups, and each group was then subdivided into a no-exercise (control) and a vertical jumping exercise group. Bone mineral density (BMD) was measured at baseline and after the 6-month study period. Urinary gamma-carboxyglutamate (Gla), a possible parameter of osteocalcin carboxylation, was also measured at baseline. Among the premenopausal women, hip BMD in the exercise group increased significantly in comparison to the control value. Among the postmenopausal women, however, there was no significant difference in the BMD change between the control and the exercise group. In addition, the baseline urinary Gla level showed an inverse correlation with the change in whole body BMD in the premenopausal exercise group. These results suggest that: (i) estrogen plays a certain role in the high-impact exercise induced bone gain, and (ii) the effects of skeletal loading on bone mass are involved in the compensation mechanism, i.e., bone gain due to high-impact exercise becomes greater in accordance with the degree of deterioration in bone material properties. Our concept of the compensation mechanism could provide a new insight into the understanding of the skeleton's adaptability to load bearing. PMID- 12115065 TI - Assessment of bone feature parameters from lumbar trabecular skeletal patterns using mathematical morphology image processing. AB - In this study, bone feature parameters were obtained from digital lumbar vertebral images to determine their potential usefulness in assessing binary trabecular skeletal patterns. The system consisted of a magnification radiographic technique, computed radiography (CR), microfocused X-ray computed tomography (mu-CT), and mathematical morphology image processing. A digital CR image was produced, using a ten-times magnified lumbar vertebral cancellous bone block (1.5 x 1.5 x 1.5 cm(3)). The information was then subjected to mathematical morphology image processing, to extract eight binary trabecular skeletal patterns having different continuities as the operation number ( n) increased. These skeletal patterns were used as test patterns and analyzed quantitatively by calculation of the skeletal pixel percentage (SKP) and star volume analysis (Vsk, volume of skeletal trabecular elements; Vsp, volume of nonskeletal elements). Then the binary skeletal pattern data were converted into the mu-CT format, using a general personal computer, and the images were quantitatively analyzed with mu CT built-in image analysis software for microstructural indices, fractal dimension analysis, and node-strut analysis. The SKP and Vsk showed an expected decrease as n increased. Concurrently, the Vsp showed an expected increase as n increased. The skeletal thickness (Sk.Th) showed a constant value at n = 1 to 4 and n = 5 to 7, but decreased in a stepwise fashion at n = 1 and n = 5. The skeletal number (Sk.N), bone area fraction (B.Ar/T.Ar) and bone perimeter fraction (B.Pm/T.Ar), decreased as n increased. Skeletal space (Sk.Sp) and perimeter-to-area ratio (B.Pm/B.Ar) increased with increasing n. In the fractal dimension analysis, the values decreased with increasing n and showed changes similar to the image observations. In fact, the SKP, star volume analysis, Sk.N, B.Ar/T.Ar, B.Pm/T.Ar, Sk.Sp, and B.Pm/B.Ar all closely mirrored the observational analysis of the images. Linkage of the skeletal structure as functions of the following parameters could be quantitatively demonstrated. The parameters used were node (Nd) and terminus (Tm). Variations in the total strut length (TSL) and Tm could demonstrate quantitative changes in skeletal structure. These results indicate that the system consisting of a combination of a magnification radiographic technique, CR, mu-CT, and mathematical morphology image processing may be a useful tool for quantitative skeletal structure analysis and the structural assessment of lumbar vertebrae, for the assessment of skeletal structural changes. It is important to choose suitable parameters for the desired structural changes. PMID- 12115066 TI - A time course of bone response to jump exercise in C57BL/6J mice. AB - Exercise, by way of mechanical loading, provides a physiological stimulus to which bone tissue adapts by increased bone formation. The mechanical stimulus due to physical activity depends on both the magnitude and the duration of the exercise. Earlier studies have demonstrated that jump training for 4 weeks produces a significant bone formation response in C57BL/6J mice. An early time point with significant increase in bone formation response would be helpful in: (1) designing genetic quantitative trait loci (QTL) studies to investigate genes regulating the bone adaptive response to mechanical stimulus; and (2) mechanistic studies to investigate early stimulus to bone tissue. Consequently, we investigated the bone structural response after 2, 3, and 4 weeks of exercise with a loading cycle of ten jumps a day. We used biochemical markers and peripheral quantitative computed tomography (pQCT) of excised femur to measure bone density, bone mineral content (BMC), and area. Four-week-old mice were separated into control ( n = 6) and jump groups ( n = 6), and the latter groups of mice were subjected to jump exercise of 2-week, 3-week, and 4-week duration. Data (pQCT) from a mid-diaphyseal slice were used to compare bone formation parameters between exercise and control groups, and between different time points. There was no statistically significant change in bone response after 2 weeks of jump exercise as compared with the age-matched controls. After 3 weeks of jump exercise, the periosteal circumference, which is the most efficient means of measuring adaptation to exercise, was increased by 3% ( P < 0.05), and total and cortical area were increased by 6% ( P < 0.05) and 11% ( P < 0.01), respectively. Total bone mineral density (BMD) increased by 11% ( P < 0.01). The biggest changes were observed in cortical and total BMC, with the increase in total BMC being 12% ( P < 0.01). Interestingly, the increase in BMC was observed throughout the length of the femur and was not confined to the mid-diaphysis. Consistent with earlier studies, mid-femur bone mass and area remained significantly elevated in the 4-week exercise group when compared with the control group of mice. The levels of the biochemical markers osteocalcin, skeletal alkaline phosphatase, and C-telopeptide were not significantly different between the exercise and control groups, indicating the absence of any systemic response due to the exercise. We conclude that a shorter exercise regimen, of 3 weeks, induced a bone response that was greater than or equal to that of 4 weeks of jump exercise reported earlier. PMID- 12115067 TI - Maintenance of trabecular structure and bone volume by vitamin K(2) in mature rats with long-term tail suspension. AB - Bone volume loss is one of the major health problems during long-term spaceflight. We examined the effects of vitamin K(2) on bone abnormalities in tail-suspended mature male Sprague-Dawley rats (13 weeks old). In this model, increased bone resorption and sustained suppression of bone formation resulted in progressive bone loss in 4 weeks, which simulates bone changes in humans during spaceflight. A significant decrease in bone mineral density (BMD), as well as a decreased mineral apposition rate (MAR), increased number of osteoclasts per bone perimeter (N.Oc/B.Pm), and increased osteoclast surface per bone surface (Oc.S/BS) in the suspended group was effectively prevented by vitamin K(2), given orally (menatetrenone, 22 mg/kg body weight). Microfocus computed tomography (CT) and node-strut analyses revealed that the volume and structure of trabecular bone were maintained near normal by the vitamin K(2) treatment. A recent report has suggested the abnormal metabolism or action of vitamin K in a microgravity environment, and our data therefore suggest that vitamin K(2) may be useful for the prevention of bone loss and for the maintenance of normal trabecular structure during spaceflight. PMID- 12115068 TI - Microstructural properties of bone in rat vertebra after long-term clodronate treatment. AB - Bisphosphonates (BPs) are known to increase bone mineral density, but it is not known how this increase manifests at low hierarchic levels of the bone structure. The present study aimed to clarify the effects of the long-term use of clodronate on the microstructure and chemical composition of bone. The second lumbar vertebral body (L2) in growing rats, subjected to 32 weeks' treatment with clodronate at either a therapeutic dose of 2 mg/kg, or a high dose of 10 mg/kg, or physiological saline (control group), was studied by scanning electron microscopy for morphology, by backscattered electron image (BSE) for density, and by energy dispersive spectrometry for material analysis. BSE images showed that the degree of mineralization in the different areas of trabecular bone of the vertebral body varied in both the control and the study groups, but this variation seemed to be different in the control and study groups. BSE analysis showed that there was more high-density bone (white area) in the low-dose clodronate group than in the controls, but the difference between the high-dose clodronate group and the control group was not significant. The density of the white area (high-density bone) was slightly increased in the low-dose clodronate group. There were no differences in the density of the gray area (low-density bone) between the groups. Neither the distribution of Ca, P, or Mg, nor the total mineral content, was affected by the clodronate treatment. Our results indicate that long-term clodronate treatment at the therapeutic level increases the proportion of high-density bone in the vertebral body in non-osteoporotic rats. PMID- 12115069 TI - Transient relief of metastatic cancer bone pain by oral administration of etidronate. AB - The aims of the present study were to determine whether patients with painful bone metastases from primary cancer sites showed a higher level of a bone resorption marker than those with no evidence of skeletal-related events, and to clarify the efficacy of oral administration of etidronate for pain due to bone metastases and bone resorption. Thirty outpatients with cancer were recruited: 10 with pain due to bone metastasis from the primary cancer site; lung (4), prostate (3), and breast (3) (M group), and 20 with primary cancer of the stomach (11), colon (4), breast (3), lung (1), and bladder (1) with no such evidence of skeletal-related events (non-M group). None of the patients in the M group either needed morphine for pain relief or had hypercalcemia, although all of them had been taking nonsteroidal anti-inflammatory drugs (NSAIDs). During the study, they continued taking NSAIDs, as they had before the study. The level of urinary cross linked N-telopeptides of type I collagen (NTx) at baseline was significantly higher in the M group than in the non-M group ( P < 0.01). Oral administration of etidronate (400 mg/day for 2 weeks) to patients in the M group significantly reduced bone pain 2 and 12 weeks after the start of treatment; however, the pain relief effect was diminished 12 weeks after the start of treatment, despite a significant decrease in urinary NTx level ( P < 0.05 by one-way analysis of variance [ANOVA] with repeated measurements). The present study provides evidence suggesting that patients with painful bone metastases from primary cancer sites may have a higher level of urinary NTx than those with no evidence of skeletal related events, and that oral administration of etidronate at the dose we used may have the potential to transiently relieve their bone pain by decreasing abnormally raised bone resorption. Although the present study had a small sample size, and had no placebo controls, the results may be useful, especially as they raise additional questions that could stimulate further research in Japan. PMID- 12115070 TI - Treatment for patients with postmenopausal osteoporosis who have been placed on HRT and show a decrease in bone mineral density: effects of concomitant administration of vitamin K(2). AB - We have been using hormone replacement therapy (HRT) as the treatment of choice for menopausal symptoms and osteoporosis. Estrogen increases bone mineral density (BMD) for 2 or 3 years, and only maintains BMD thereafter. In the present study, we investigated whether BMD improved with HRT in combination with vitamin K(2). Ninety-four patients with postmenopausal osteoporosis were studied. All patients were placed on HRT for more than 1 year. Ten patients whose BMD had increased up to a plateau and showed a decreasing trend thereafter while they were receiving HRT were placed on HRT in combination with vitamin K(2). The long-term BMD (L(2) L(4)) profiles of those receiving HRT alone, and the effects of HRT in combination with vitamin K(2) on the BMD profiles were examined. The rate of change in BMD (mean +/- SE) of all patients who underwent HRT alone was 2.9 +/- 1.2% 1 year after the initiation of HRT, 4.6 +/- 1.2% 2 years later, and 5.4 +/- 1.2% 3 years later. The BMD in these patients, which reached a peak increase after 3 years, no longer increased from the following year. When vitamin K(2) was administered concomitantly to 10 patients for 12 months because of the decrease in BMD, their BMD (mean +/- SE) increased significantly, from 0.734 +/- 0.021 g/cm(2) to 0.783 +/- 0.026 g/cm(2) ( P < 0.03; paired- t test). We conclude that HRT in combination with vitamin K(2) significantly improved BMD that was on the decrease after HRT alone. Therefore, as a supplementary drug for postmenopausal osteoporosis, vitamin K(2) is a good therapeutic option for patients who are placed on HRT. PMID- 12115071 TI - Chemotherapy did not enhance the anti-osteolytic effects of bisphosphonate in multiple myeloma bone disease. PMID- 12115072 TI - Spironolactone for the treatment of hypercalciuric hypocalcemia. AB - A patient with hypocalcemia and hypomagnesemia, and renal wasting of both cations, is presented. This patient had a dramatic calcium response to treatment with spironolactone. The possible diagnoses and mechanisms of response are discussed. It is concluded that the patient had a calcium-sensing receptor mutation, and that treatment with spironolactone induced an effect on renal handling of calcium in this condition. It is suggested that this treatment be tried in other similar patients. PMID- 12115073 TI - Endoscopic buccal mucosal grafting to the anterior glottic web: a case report. AB - Anterior glottic web (AGW) is a challenge for otolaryngologists to treat because of the wide range of symptoms and the involvement of the patient's airway. We were presented with an 18-year-old male patient suffering from exertional dyspnea and moderate dysphonia. Two attempts at microlarnygeal surgery, using a CO(2) laser to try to lysis the AGW, failed to achieve satisfactory results. Further examination indicated AGW, with compromised bilateral vocal fold movement and voice generation capabilities. A two-stage endoscopic buccal mucosal grafting (EBMG) was recommended to accomplish anterior commissure reconstruction to treat the epithelial loss resulting from AGW. In our postoperative assessment, the patient's symptoms had improved dramatically, with clear, smooth voice production without dyspnea. Therefore, EBMG is believed to be a technically viable and feasible approach to treating AGW and is also recommended as an effective alternative to treatments such as laryngofissure and tracheostomy. PMID- 12115074 TI - Voice analysis and videolaryngostroboscopy in patients with Parkinson's disease. AB - Parkinson's disease (PD) is clinically characterized by a resting tremor, bradykinesia, cogwheel phenomenon, rigidity, disorder of postural reflexes and especially changes in voice and speech. We studied 30 PD patients who were treated with dopamine and 20 normal subjects as the control group. The parameters of vocal fold edges, glottal closure, vertical levels of cords, amplitude of vibration, mucosal wave, vibratory behavior, phase symmetry, ventricular folds and movements, periodicity, arytenoids and thick mucous were evaluated by videolaryngostroboscopy. The Unified Parkinson's Disease Rating Scale was applied to the patient group. The voices of the patients were evaluated by the Dr.Speech 4 and Spectra-PRO computer programs. Maximum phonation time, fundamental frequency, amplitude and the harmonic-to-noise ratio were recorded and compared with those of the control group. The abnormal videolaryngostroboscopic findings were more frequent in the PD group (70% versus 45%; P<0.05). Voice analysis showed significant differences in the parameters such as maximum phonation time, maximum fundamental frequency, the frequency range and the harmonic-to-noise ratio. We thought that these methods and parameters yielded sufficient information for diagnosis and follow-up of vocal function in patients with PD. PMID- 12115075 TI - Immunohistochemical expression of VEGF and VEGF receptors in nasal polyps as compared to normal turbinate mucosa. AB - The factors involved in the development of chronic inflammation and edema in nasal polyps remain to be clarified. The expression of vascular endothelial growth factor (VEGF) has been described in plasma cells, suggesting that plasma cells may play a major role in the development of edema in nasal polyps through the production of VEGF. We performed immunohistochemical analysis using specific antibodies to VEGF and to the known VEGF receptors, VEGFR-1 and VEGFR-2, on paraffin sections of human nasal polyps ( n=11) and controls of human mucosa of the normal middle turbinates ( n=6). In normal turbinate mucosa, sporadic immunostaining for VEGF was observed throughout the endothelial cells of the small veins and arteries. VEGFR-1 and VEGFR-2 expression was faint in the healthy turbinates. In nasal polyp tissues, strong immunostaining for VEGF was found in the endothelium of blood vessels and in the infiltrating perivascular inflammatory cells. Fibroblasts also stained for VEGF. Strong immunolabeling to VEGFR-1 was evident in the vascular endothelium, whereas weak to moderate VEGFR-1 staining was generally confined to scattered mononuclear round cells. Mononuclear round cells and the endothelium of capillaries revealed immunoreactivity to VEGFR 2. These findings support a role for VEGF and its receptors, VEGFR-1 and VEGFR-2, in the development and perpetuation of edema and angiogenesis in nasal polyps. PMID- 12115076 TI - Acoustic neuroma surgery and tinnitus. AB - The objectives of this study were to assess the effect that acoustic neuroma surgery has on tinnitus and to investigate possible predictors (tumour size and patients' ages at operation) as well as to ascertain if the overall quality of life in patients with acoustic neuromas is affected by their tinnitus. A questionnaire was sent to randomly selected patients post acoustic-neuroma surgery. This was based on the Glasgow Benefit Inventory and contained a standardised series of four functional gradings for tinnitus. In this study, 51 patients from a total of 68 returned the questionnaire, and there was a follow-up period of between 1 and 3 years following the acoustic neuroma surgery. The age at operation, size of the tumour and overall quality of life were correlated with the impact of surgery on tinnitus. Statistical analysis used the one-way analysis of variance, chi-square test, one-way analysis by ranks and Spearman Rank Correlations. Significance was accepted at the P<0.05 level. Overall, 30 (58.8%) of the patients had tinnitus preoperatively in comparison to 34 (66.6%) postoperatively. After surgery, tinnitus became better in 8 (16%) patients, 28 (55%) did not experience any change, and 15 (29%) became worse. Neither tumour size nor age at the time of the operation had a statistically significant association with the impact of surgery on tinnitus. There was no statistically significant association between changes in tinnitus status and changes in the quality of life following the operation ( P>0.05). A significant percentage of patients with acoustic neuromas, approximately 60%, suffer from tinnitus preoperatively, and this number may increase slightly postoperatively. It remains unpredictable which patients will improve, which will show no change and which will deteriorate as age and tumour size do not seem to be associated with the impact of surgery on tinnitus. The results also suggest that tinnitus may be of relatively minor importance in the overall quality of life of patients following acoustic neuroma surgery. However, candidates for surgery should be thoroughly informed about the possible effect of the operation on their tinnitus status. PMID- 12115078 TI - Arachnoid cyst of the cranial posterior fossa causing sensorineural hearing loss and tinnitus: a case report. AB - Arachnoid cysts are developmental collections of cerebrospinal fluid covered by layers of arachnoidal epithelium and are usually located in the middle cranial fossa. Localizations in the posterior fossa are uncommon and generally remain asymptomatic or cause vague and non-specific symptoms. We here report the unusual case of a young patient with an arachnoid cyst of the posterior fossa that had become manifest in the form of left-sided hypoacusia and tinnitus. Audiometric evaluation, auditory brainstem responses and transient-evoked otoacoustic emissions tests revealed a progressive monolateral sensorineural retrocochlear hearing loss with cochlear involvement. Magnetic-resonance imaging showed an arachnoid cyst of the cerebellar convexity compressing the cerebellar hemisphere and thus the homolateral cerebellopontine angle. Because of the progressive worsening of the retrocochlear impairment after a 6-month wait-and-see period, the patient finally underwent endoscopic cyst decompression. This was judged to be successful as it stopped the progression of hearing loss and the impairment of the auditory brainstem responses and made the tinnitus more tolerable. PMID- 12115077 TI - Ear injury caused by a sticky-tipped applicator. AB - We here report a relatively rare case of traumatic injury of the tympanic membrane caused by a sticky-tipped applicator as well as some characteristics of this sticky-tipped applicator that were examined experimentally. This rare case was very unusual in that the injury was caused by external force applied from the medial to lateral direction during very careful cleaning (no unexpected force). Although removal of cerumen by a sticky substance seems to be an excellent idea, the present case shows that the stickiness that facilitates removal of the cerumen may be dangerous for thin, dry skin such as the tympanic membrane. Actually, the experimental considerations indicated that the stickiness was affected by the surface condition of the objective: soft, dry conditions increase it, while on the contrary, moist, dusty and oily conditions reduce it. The stickiness of the applicator, which was adjusted to use in the ear canal with an oily surface, appeared to be too large for a dry tympanic membrane. It is recommended that the sticky substance should be removed from the tip area or reduced by application of oil or powder before use. PMID- 12115079 TI - Degeneration of the stria vascularis during development in melanocyte-deficient mutant rats (Ws/Ws rats). AB - Developmental changes in the stria vascularis of white spotting (Ws) rats were examined by scanning and transmission electron microscopes and by diaminobenzidine-staining techniques. The stria of Ws/Ws homozygote rats was found to have both pigmented and non-pigmented portions. While the pigmented portions possessed intermediate cells in the same manner as the stria of wild +/+ rats, the non-pigmented portions lacked the cells. Examination at 1, 2, 3, 4, 6, 8 and 14 weeks after birth revealed a progressive degeneration in the marginal cells and strial capillaries in the non-pigmented portions. At 1 week, no significant differences were seen in the marginal cells of any of the rats examined. At 2 weeks, the basolateral infoldings of the marginal cells were seen to be well developed and adult-like in the pigmented portions of Ws/Ws rats and in +/+ rats. In the non-pigmented portions, the basolateral infoldings of the marginal cells appeared well developed; however, vacant spaces were seen around the basolateral infoldings. At 3 weeks, the basolateral infoldings of the marginal cells in the non-pigmented portions had become more atrophic, and the empty spaces around the basolateral infoldings had enlarged. Also, the marginal cells themselves had become flatter or thinner. These findings became more prominent at 4 weeks and 6 weeks. At 8 weeks and 14 weeks after birth, the marginal cells appeared markedly flat, and no basolateral infoldings were seen in the non-pigmented portions. Pigmented portions of the stria in Ws/Ws rats, on the other hand, showed normal development throughout this period. A DAB-staining examination of the stria capillary net in Ws/Ws rats showed it to be well developed at 3 weeks in both pigmented and non-pigmented portions. At 8 weeks, a thickening of the capillary basement membrane was apparent. The above findings lead the authors to believe that intermediate cells play an important role in the development and maintenance of marginal cells and the strial capillary system. PMID- 12115080 TI - A tissue-engineering model for the manufacture of auricular-shaped cartilage implants. AB - The established surgical methods of external ear reconstruction using autogenous tissue represent the current state of the art. Because of the limited possibilities for shaping conventional harvested autogenous rib cartilage, the cosmetic results of auricular reconstruction are frequently unsatisfactory. Tissue engineering could represent an alternative technique for obtaining a precisely shaped cartilage implant that avoids donor site morbidity and unsatisfactory cosmetic results. In this study, the reliability and quality of a tissue-engineering model for the manufacture of auricular-shaped human cartilage implants was investigated, focusing on the feasibility of the manufacturing process and the in vivo and in vitro maturation of an extracellular cartilage like matrix. Implants were molded within an auricular-shaped silicone cylinder, and human nasal septal chondrocytes crosslinked by human fibrin within bioresorbable PGLA-PLLA polymer scaffolds were used. After an in vitro incubation of up to 6 weeks, defined fragments of the prefabricated auricular-shaped construct were implanted subcutaneously on the backs of nude mice for at least 6 to 12 weeks ( n=7). Scaffolds without cell loading served as controls. Macroscopic and histochemical examination after 3 and 6 weeks in vitro showed a solid compound of homogenously distributed chondrocytes within the polymer scaffold, leading only to a limited pericellular matrix formation. Analysis after 6 and 12 weeks of in vivo maturation demonstrated a solid tissue compound and neocartilage formation with the presence of cartilage-specific matrix components. Implants obtained shape and size during the entire period of implantation. The model of cartilage implant manufacturing presented here meets all biocompatible requirements for in vitro prefabrication and in vivo maturation of autogenous, individually shaped cartilage transplants. PMID- 12115081 TI - Papillary thyroid carcinoma metastasis to the parapharyngeal space. AB - Thyroid papillary carcinoma presenting as a pharyngeal mass is a rare clinical occurrence and has only been reported sporadically. We present here two patients who had papillary carcinoma metastasis of the thyroid gland to the parapharyngeal space. Each patient had a different clinical progress. Upward lymphatic spread of the tumor to involve the parapharyngeal space via the lateral retropharyngeal nodes was indicated. This pattern of spread is in keeping with Rouviere's description of a direct lymphatic pathway from the thyroid gland to the retropharyngeal nodes and parapharyngeal space. PMID- 12115082 TI - Diagnostic strategies in cervical carcinoma of an unknown primary (CUP). AB - In patients with cervical cancer of an unknown primary (CUP), no established concept exists for the necessary diagnostic procedures. In order to find the primary tumor, extensive diagnostic steps are generally recommended; however, they are often not performed consistently. In the current study, we consistently used a diagnostic algorithm and analyzed its consequences on patients' prognoses. We retrospectively studied 57 patients who were found to have a cervical metastasis of the upper- or midneck and an unknown primary tumor after routine examination of the head and neck region. Patients were analyzed for the value of applied diagnostic measures, tumor classification, survival rates and frequencies of subsequent lymph node or distant metastases after the initial treatment. Our results showed that a diagnostic algorithm (lymph node biopsy, rigid panendoscopy with systematic biopsies of suspect regions as well as blind biopsies of endoscopically inconspicuous regions, including the tongue base and nasopharynx and bilateral tonsillectomy) led to the detection of 14 occult oropharyngeal and 5 nasopharyngeal primary tumors in the patients. These tumors were primarily diagnosed as CUP. Oropharyngeal tumors either grew submucosally or were so small that only microscopic evaluation of the entire tonsil uncovered the tumor. Imaging procedures (X-ray, ultrasound, CT, MRT and FDG-PET) as well as gynecological, urological and gastroenterological consultations did not reveal the primary tumors in any of the cases. The 3-year survival rate for the patients with occult oropharyngeal primary tumors was 100% after treatment, while the patients in which our diagnostic schedule did not reveal a primary tumor showed a survival rate of 58%. The prognosis of all of the patients with cervical carcinoma metastasis was dependent on the initial nodal stage. Metachronous metastasis after completion of the initial treatment was prognostically infaust, while secondary detection of the primary tumor was worthwhile during follow-up as long as further treatment options were offered. The prognosis of patients with cervical carcinoma metastases of the upper- and midneck is much more favorable than that of patients with a CUP syndrome of other localizations. Identification of an occult pharyngeal tumor is prognostically relevant, since it opens up the possibility of specific locoregional treatment. In patients with cervical CUP, blind but systematic pharyngeal biopsies, including bilateral tonsillectomy, should be performed. PMID- 12115084 TI - Facial electroneurography on the contralateral side in unilateral Bell's palsy. AB - In this study, ten patients who exhibited severe unilateral Bell's palsy of the House-Brackmann grade V underwent facial electroneurography (ENoG) on the contralateral, healthy side. Serial ENoG was conducted in seven consecutive sessions within 6 months at a given current intensity level of stimulation. According to our results, all the patients presented a rise in the maximum compound-action potential (MCAP) amplitude on the healthy side within 20 to 45 days from the onset of the palsy and shortly after the onset of the recovery of the facial function. This was attributed to the central contralateral compensatory process, which restores balanced facial function. Based on our data, a hypothetical model is shown, which demonstrates the clinical course of the contralateral MCAP values and reflects the plasticity effect of the central nervous system after the onset of Bell's palsy. PMID- 12115083 TI - The effects of caloric vestibular stimulation on EEGs in patients with central vertigo. AB - The EEGs of 30 patients with central vertigo were examined by caloric vestibular stimulation (CVS). The control group consisted of 30 persons without vertigo disturbances. In the standard procedure (and after hyperventilation), there was no statistically significant difference between the two groups of patients. In comparison to the EEGs of healthy persons, CVS produced statistically significant differences ( P<0.001) in the EEGs of patients with vertigo. PMID- 12115085 TI - Tuberculin skin testing surveillance of health care personnel. AB - To estimate the incidence of and assess risk factors for occupational Mycobacterium tuberculosis transmission to health care personnel (HCP) in 5 New York City and Boston health care facilities, performance of prospective tuberculin skin tests (TSTs) was conducted from April 1994 through October 1995. Two-step testing was used at the enrollment of 2198 HCP with negative TST results. Follow-up visits were scheduled for every 6 months. Thirty (1.5%) of 1960 HCP with >/=1 follow-up evaluation had TST conversion (that is, an increase in TST induration of >/=10 mm). Independent risk factors for TST conversion were entering the United States after 1991 and inclusion in a tuberculosis-contact investigation in the workplace. These findings suggest that occupational transmission of M. tuberculosis occurred, as well as possible nonoccupational transmission or late boosting among foreign-born HCP who recently entered the United States. These results demonstrate the difficulty in interpreting TST results and estimating conversion rates among HCP, especially when large proportions of foreign-born HCP are included in surveillance. PMID- 12115086 TI - Risk factors for late-onset nosocomial pneumonia caused by Stenotrophomonas maltophilia in critically ill trauma patients. AB - Patients with nosocomial pneumonia caused by Stenotrophomonas maltophilia often receive inadequate empiric antibiotic therapy, potentially increasing mortality. Knowledge of the risk factors associated with S. maltophilia pneumonia may better guide the selection of empiric antibiotic therapy. Potential risk factors for S. maltophilia pneumonia were retrospectively analyzed for critically ill trauma patients with late-onset gram-negative pneumonia. The effects of S. maltophilia infection and inadequate empiric antibiotic therapy on patient outcomes were also assessed. By multivariate analysis, S. maltophilia pneumonia was found to be associated with cefepime exposure and tracheostomy in patients with a single pneumonia episode and with higher Injury Severity Score and pulmonary contusion in patients with multiple pneumonia episodes. S. maltophilia pneumonia was associated with increased patient morbidity; only inadequate empiric antibiotic therapy was associated with a higher mortality rate. In critically ill trauma patients with late-onset ventilator-associated pneumonia and these risk factors, empiric antibiotic therapy should include agents active against S. maltophilia. PMID- 12115087 TI - Injection drug use facilitates hepatitis C virus infection of peripheral blood mononuclear cells. AB - Infection of peripheral blood mononuclear cells (PBMCs) with hepatitis C virus (HCV) has been demonstrated and has been found to play a role in relapse of HCV disease and vertical transmission of HCV. Injection drug use is thought to impair function of the immune system and induce tolerance to viruses; therefore, HCV infection of PBMCs could be more likely to occur in injection drug users (IDUs) with HCV infection. Of 108 women who tested negative for human immunodeficiency virus type 1 and positive for HCV RNA, 51 had a history of injection drug use and 57 had no known risk factor for HCV infection. HCV infection was found, by nested reverse-transcription polymerase chain reaction analysis, in the PBMCs of 33 IDUs and of 13 non-IDUs (P=.00003). No correlation was found between infection of the PBMCs and HCV genotype or virus load. Route of transmission and viral factors, as well as immunologic dysfunction, may play a role in viral tropism. PMID- 12115088 TI - Risk Factors Associated with Helicobacter pylori Infection among Children in a Defined Geographic Area. AB - Factors influencing the pattern of Helicobacter pylori infection among children living in adjacent urban and rural areas of northern Sardinia, Italy, were compared. The seroprevalence of H. pylori infection was 22% (625 of 2810 children) in the study population and was significantly higher among children in rural areas (37%) than in urban areas (13%) (odds ratio [OR], 3.8; 95% confidence interval [CI], 3.2-4.7; P<.005). This difference was consistent within each age group. In rural areas, children who had dogs were at greatest risk for H. pylori infection (OR, 1.8; 95% CI, 1.3-2.6; P<.05). No association was seen between H. pylori seropositivity and a history of breast-feeding. Urban children attending day care centers had a higher prevalence of infection (17%) than did those who never attended (12%) (OR, 1.5; 95% CI, 1.1-2.0; P<.05). The epidemiology of H. pylori infection is complex; even within the same geographic area, different factors influence acquisition of H. pylori infection. PMID- 12115089 TI - Natural history of human calicivirus infection: a prospective cohort study. AB - We investigated the natural history of human Calicivirus infection in the community. Clinical information was obtained from 99 subjects infected with Norwalk-like viruses (NLV) and 40 subjects infected with Sapporo-like viruses (SLV) in a prospective, community-based cohort study. NLV infection was common in all age groups, whereas SLV infection was mainly restricted to children aged <5 years. Symptoms lasted for a median of 5 and 6 days for NLV and SLV infections, respectively. Disease was characterized by diarrhea during the first 5 days (87% of patients with NLV infection and 95% of patients with SLV infection) and vomiting on the first day (74% for NLV and 60% for SLV). Vomiting was less common in children aged <1 year (59% for NLV and 44% for SLV) than it was among children aged >/=1 year (>75% for NLV and >67% for SLV). Overall, NLV was detected in 26% of patients up to 3 weeks after the onset of illness. This proportion was highest (38%) for children aged <1 year. SLV shedding subsided after 14 days. These data show that the durations of disease and viral shedding of caliciviruses are longer than has been described elsewhere. Therefore, the impact of these infections may have been underestimated. PMID- 12115090 TI - Outcome of and prognostic factors for herpes simplex encephalitis in adult patients: results of a multicenter study. AB - Management of herpes simplex encephalitis (HSE) has been considerably improved by the availability of acyclovir therapy and rapid polymerase chain reaction (PCR) based diagnostic assays. Prognostic factors for this rare affliction are, however, misestimated. We conducted a large retrospective multicenter study that included 93 adult patients in whom HSE was diagnosed by PCR from 1991 through 1998 and who were treated with intravenous acyclovir. Among the 85 patients assessed at 6 months, 30 (35%) had a poor outcome, which led to death in 13 patients (15%) and severe disability in 17 (20%). The outcome was favorable for 55 patients (65%). A multivariate analysis identified 2 factors that were found to be independently associated with poor outcome: a Simplified Acute Physiology Score II >/=27 at admission and a delay of >2 days between admission to the hospital and initiation of acyclovir therapy. Early administration of antiviral therapy is the only parameter that can be modified to improve the prognosis of patients with HSE. PMID- 12115091 TI - A randomized, double-blind study of the efficacy of a 10- or 20-day course of sodium stibogluconate for treatment of cutaneous leishmaniasis in United States military personnel. AB - The recommended treatment for cutaneous leishmaniasis is pentavalent antimony at a dosage of 20 mg/kg/day for 20 days. Some studies conducted in locales in which Leishmania is endemic have suggested that shorter courses of treatment may be as efficacious. We conducted a randomized, double-blind, placebo-controlled study of 10 versus 20 days of sodium stibogluconate (SSG) in United States military personnel who contracted cutaneous leishmaniasis while serving overseas; 19 patients received SSG for 10 days (and placebo for 10 days), and 19 patients received SSG for 20 days. Cure rates were 100% (19 of 19 patients) in the 10-day group and 95% (18 of 19 patients) in the 20-day group. Side effects were more common among patients who received 20 days of therapy. In this group of otherwise healthy young adults, SSG at a dosage of 20 mg/kg/day for 10 days appears to have been therapeutically equivalent and less toxic than the standard 20-day course. PMID- 12115093 TI - Coagulation abnormalities in dengue hemorrhagic Fever: serial investigations in 167 Vietnamese children with Dengue shock syndrome. AB - The pathophysiological basis of hemorrhage in dengue infections remains poorly understood, despite the increasing global importance of these infections. A large prospective study of 167 Vietnamese children with dengue shock syndrome documented only minor prolongations of prothrombin and partial thromboplastin times but moderate to severe depression of plasma fibrinogen concentrations. A detailed study of 48 children revealed low plasma concentrations of the anticoagulant proteins C, S, and antithrombin III, which decreased with increasing severity of shock, probably because of capillary leakage. Concurrent increases in the levels of thrombomodulin, tissue factor, and plasminogen activator inhibitor type 1 (PAI-1) indicated increased production of these proteins. Thrombomodulin levels suggestive of endothelial activation correlated with increasing shock severity, whereas PAI-1 levels correlated with bleeding severity. Dengue virus can directly activate plasminogen in vitro. Rather than causing true disseminated intravascular coagulation, dengue infection may activate fibrinolysis primarily, degrading fibrinogen directly and prompting secondary activation of procoagulant homeostatic mechanisms. PMID- 12115092 TI - Epidemiology of invasive group a streptococcus disease in the United States, 1995 1999. AB - Severe invasive group A streptococcal (GAS) disease is believed to have reemerged during the past 10-20 years. We conducted active, laboratory, population-based surveillance in 5 US states (total population, 13,214,992). From 1 July 1995 through 31 December 1999, we identified 2002 episodes of invasive GAS (3.5 cases per 100,000 persons). Rates varied by age (higher among those <2 or >/=65 years old), surveillance area, and race (higher among black individuals) but did not increase during the study period. The 5 most common emm types (1, 28, 12, 3, and 11) accounted for 49.2% of isolates; newly characterized emm types accounted for 8.9% of isolates. Older age; presence of streptococcal toxic shock syndrome, meningitis, or pneumonia; and infection with emm1 or emm3 were all independent predictors of death. We estimate that 9600-9700 cases of invasive GAS disease occur in the United States each year, resulting in 1100-1300 deaths. PMID- 12115094 TI - Osteomyelitis in elderly patients. AB - In elderly persons, osteomyelitis is second only to soft-tissue infection as the most important musculoskeletal infection. Acute osteomyelitis is usually acquired hematogenously, and the most common pathogen is Staphylococcus aureus. Acute osteomyelitis can usually be cured with antimicrobial therapy alone. In contrast, chronic osteomyelitis may be caused by S. aureus but is often due to gram negative organisms. The causative organism of chronic osteomyelitis is identified by culture of aseptically obtained bone biopsy specimens. Because of the presence of infected bone fragments without a blood supply (sequestra), cure of chronic osteomyelitis with antibiotic therapy alone is rarely, if ever, possible. Adequate surgical debridement is the cornerstone of therapy for chronic osteomyelitis, and cure is not possible without the removal of all infected bone. PMID- 12115095 TI - Economic evaluation of immunization strategies. AB - Resources used to provide health care are vast but not limitless. When clinicians are asked to participate in decisions for large groups of patients (in a managed care context, in an institution, or at the level of local health authorities), the balance between consumption of resources and the benefits of an intervention is important. Clinicians may use cost-effectiveness and cost-benefit studies to inform such decisions (but not to make them). Because of differences in methods, the presentation of results, and country-specific parameters, economic evaluations of the same vaccination strategy by different groups may have divergent results. In this article, we review methodologic issues, limitations, and ethical considerations related to economic evaluations of immunization strategies, focusing on immunizations associated with travel. PMID- 12115096 TI - Helicobacter pylori: consensus and controversy. AB - Helicobacter pylori is uniquely adapted to colonize the human stomach. Infection leads to a range of subclinical and clinical outcomes that depend on properties of the infecting strain, the host, and the environment. Eradication therapy is indicated for infected persons who develop peptic ulcer disease or gastric lymphoma or who are beginning long-term treatment with nonsteroidal anti inflammatory drugs. However, treatment may worsen gastroesophageal reflux disease and increase the risk of esophageal cancer. H. pylori infections can be diagnosed noninvasively and can be eradicated with approximately 85% success by a variety of multidrug, 7-14-day regimens. Unfortunately, antibiotic resistance is affecting treatment effectiveness in the United States and abroad. A more complete understanding of the variation in H. pylori pathogenesis should lead to clearer recommendations about treatment for infected persons who have neither peptic ulcer disease nor gastric lymphoma. PMID- 12115097 TI - Human immunodeficiency virus in correctional facilities: a review. AB - It is estimated that up to one-fourth of the people living with human immunodeficiency virus (HIV) infection in the United States pass through a correctional facility each year. The majority of persons who enter a correctional facility today will return home in the near future. Most inmates with HIV infection acquire it in the outside community; prison does not seem to be an amplifying reservoir. How correctional health services deal with the HIV-infected person has important implications to the overall care of HIV-infected people in the community. Routine HIV testing is well accepted. Combination antiretroviral therapy has been associated with a reduction in mortality in prisons. A link between area HIV specialists and correctional health care providers is an important partnership for ensuring that HIV-infected patients have optimal care both inside prison and after release. PMID- 12115098 TI - Does patient sex affect human immunodeficiency virus levels? AB - We undertook a critical epidemiological review of the available evidence concerning whether women have lower levels of human immunodeficiency virus (HIV) RNA than do men at similar stages of HIV infection. The 13 studies included in this analysis reported viral load measurements in HIV-infected men and women at a single point in time (cross-sectional studies) or over time (longitudinal studies). Seven of the 9 cross-sectional studies demonstrated that women had 0.13 0.35 log(10) ( approximately 2-fold) lower levels of HIV RNA than do men, despite controlling for CD4(+) cell count. Four longitudinal studies revealed that women had 0.33-0.78 log(10) (2- to 6-fold) lower levels of HIV RNA than do men, even when controlling for time since seroconversion. Adjustment for possible confounders of the relationship between sex and viral load, including age, race, mode of virus transmission, and antiretroviral therapy use, did not change this outcome. This finding is significant, because viral loads are frequently used to guide the initiation and modification of antiretroviral therapy. PMID- 12115099 TI - Comparative analysis of commercial assays for the detection and quantification of Human Immunodeficiency Virus Type 1 (HIV-1) RNA in plasma from patients infected with HIV-1 subtype C. AB - The commercial assays commonly used to quantify plasma human immunodeficiency virus type 1 (HIV-1) RNA in clinical settings were designed to assess HIV-1 subtype B. We compared the performance of 4 commercial assays (Amplicor versions 1.0 and 1.5 [Roche]; Quantiplex [Chiron]; and NASBA HIV-1 RNA QT [Organon Teknika]) in detecting and quantifying HIV-1 RNA in plasma from HIV-infected persons from Zambia, an area where HIV-1 subtype C is predominant. Each assay detected plasma HIV-1 RNA, but they do not all measure statistically similar quantities of plasma HIV-1 RNA. PMID- 12115100 TI - BK virus-associated hemorrhagic cystitis in a Human Immunodeficiency Virus infected patient. AB - BK virus-associated hemorrhagic cystitis is a common clinical problem in bone marrow transplant recipients but is considered rare in other immunosuppressed patient populations. We describe a human immunodeficiency virus-infected patient with non-Hodgkin's lymphoma in whom BK virus-associated hemorrhagic cystitis developed; viruria was quantitated in urine by immunocytochemistry, and the patient showed no response to cidofovir. PMID- 12115101 TI - Multifocal vasculopathy due to Varicella-Zoster Virus (VZV): serial analysis of VZV DNA and intrathecal synthesis of VZV antibody in cerebrospinal fluid. AB - Recognition of multifocal vasculopathy due to varicella-zoster virus (VZV) is often problematic. We describe a human immunodeficiency virus-infected patient who had progressive central nervous system disease for >3 months. Both VZV DNA and antibody were detected in cerebrospinal fluid (CSF) specimens; serial polymerase chain reaction analyses confirmed the diagnosis and guided the duration of therapy. Reduced ratios of VZV antibody in serum to that in CSF were also demonstrated. PMID- 12115102 TI - Furuncular myiasis: a simple and rapid method for extraction of intact Dermatobia hominis larvae. AB - We report a case of furuncular myiasis complicated by Staphylococcus aureus infection and beta-hemolytic streptococcal cellulitis. The Dermatobia hominis larva that caused this lesion could not be extracted using standard methods, including suffocation and application of lateral pressure, and surgery was contraindicated because of cellulitis. The botfly maggot was completely and rapidly extracted with an inexpensive, disposable, commercial venom extractor. PMID- 12115103 TI - Early detection of central nervous system tuberculosis with the gen-probe nucleic Acid amplification assay: utility in an inner city hospital. AB - Central nervous system tuberculosis is a serious clinical problem, the treatment of which is sometimes hampered by delayed diagnosis. We investigated the utility of the Gen-Probe nucleic acid amplification assay for the rapid diagnosis of tuberculous meningitis and as a noninvasive method of identifying intracranial tuberculoma. PMID- 12115105 TI - Garlic supplements and saquinavir. PMID- 12115106 TI - Prevention of nosocomial fungal infection: the French approach. PMID- 12115108 TI - Role of Non-O157:H7 Escherichia coli in hemolytic uremic syndrome. PMID- 12115109 TI - Linezolid-induced pancytopenia. PMID- 12115111 TI - Thrombocytopenia secondary to linezolid administration: what is the risk? PMID- 12115112 TI - Safety of lactobacillus strains as probiotic agents. PMID- 12115113 TI - Linezolid-induced pure red blood cell aplasia. AB - We report a case of reversible pure red blood cell aplasia that developed in a patient who had received 8 weeks of linezolid therapy. PMID- 12115114 TI - Staphylococcus lugdunensis infection sites: predominance of abscesses in the pelvic girdle region. AB - We used a screening protocol to identify Staphylococcus lugdunensis in clinical specimens with pure or predominant growth of coagulase-negative staphylococci. S. lugdunensis was isolated from 17 patients in a 12-month period and was judged to be the cause of infection in 14 cases. In 13 cases, the patient had a soft-tissue abscess, and in 9 of these, the abscess was located in the pelvic girdle region, which suggests that this may be the natural habitat of S. lugdunensis. PMID- 12115115 TI - Breakthrough trichosporonosis in a bone marrow transplant recipient receiving caspofungin acetate. AB - We report a case of invasive trichosporonosis due to Trichosporon beigelii originating in the left wrist of a bone marrow transplant recipient who was receiving caspofungin acetate as prophylaxis against invasive Aspergillus infection. While the patient's neutrophil count was recovering, treatment with fluconazole and amphotericin B lipid complex resulted in a complete clinical response. PMID- 12115116 TI - Stability studies of recombinant Saccharomyces cerevisiae in the presence of varying selection pressure. AB - A recombinant yeast plasmid carrying the Ieu2 gene for auxotrophic complementation and a reporter gene for beta-galactosidase under the control of Gal10 promoter was studied in Saccharomyces cerevisiae. Growth, product formation, and plasmid stability were studied in defined, semi-defined, and complex media. The biomass concentration and specific activity were higher in complex medium than in defined medium, which was selective for the growth of plasmid-containing cells, leading to a 10-fold increase in volumetric activity. However, plasmid instability was very high in complex media with 50% plasmid-free cells emerging in the culture within 75 h of cultivation. In order to control instability, the growth rates of the plasmid-containing and plasmid-free cells were determined in semi-defined media, which consisted of defined medium supplemented with different concentrations of yeast extract. Below a critical concentration of yeast extract (0.05 g/L), the plasmid-containing cells had a growth rate advantage over the plasmid-free cells. This was possibly because, at this concentration of yeast extract, the availability of leucine became the rate determining factor in the specific growth rate of plasmid-free cells. A feeding strategy was designed which maintained a low concentration of the residual yeast extract in the medium and thus continuously provided the plasmid-containing cells with a competitive advantage over the plasmid-free cells. This resulted in high stability as well as high cell density under non-selective conditions, which led to a 10-fold increase in the volumetric activity compared to that achieved in defined selective media. A simple mathematical model was formulated to verify the experimental data. The important state variables and process parameters, i.e., biomass concentration, beta-galactosidase expression, sucrose consumption, yeast extract consumption, and specific growth rates of the two cell populations, were evaluated. These variables and parameters along with the differential equations based on material balances as well as the experimental results obtained were used in a mathematical model for the fed-batch cultivation. These correctly verified the experimental data and clearly illustrated the concept behind the success of the fed-batch strategy under yeast extract starvation. PMID- 12115117 TI - Use of isolated cyclohexanone monooxygenase from recombinant Escherichia coli as a biocatalyst for Baeyer-Villiger and sulfide oxidations. AB - The performance, in Baeyer-Villiger and heteroatom oxidations, of a partially purified preparation of cyclohexanone monooxygenase obtained from an Escherichia coli strain in which the gene of the enzyme was cloned and overexpressed was investigated. As model reactions, the oxidations of racemic bicyclo[3.2.0]hept-2 en-6-one into two regioisomeric lactones and of methyl phenyl sulphide into the corresponding (R)-sulphoxide were used. Enzyme stability and reuse, substrate and product inhibition, product removal, and cofactor recycling were evaluated. Of the various NADPH regeneration systems tested, 2-propanol/alcohol dehydrogenase from Thermoanerobium brockii appeared the most suitable because of the low cost of the second substrate and the high regeneration rate. Concerning enzyme stability, kosmotropic salts were the only additives able to improve it (e.g., half-life from 1 day in diluted buffer to 1 week in 1 M sodium sulphate) but only under storage conditions. Instead, significant stabilization under working conditions was obtained by immobilization on Eupergit C (half-life approximately 2.5 days), a procedure that made it possible to reuse the catalyst up to 16 times with complete substrate (5 g x L(-1)) conversion at each cycle. Reuse of free enzyme was also achieved in a membrane reactor but with lower efficiency. Water organic solvent biphasic systems, which would overcome substrate inhibition and remove from the aqueous phase, where reaction takes place, the formed product, were unsuccessful because of their destabilizing effect on cyclohexanone monooxygenase. More satisfactory was continuous substrate feeding, which shortened reaction times and, very importantly, yielded in the case of bicyclo[3.2.0]hept-2-en-6-one (10 g x L(-1)) both lactone products with high optical purity (enantiomeric excess > or = 96%), which was not the case when all of the substrate was added in a single batch. PMID- 12115118 TI - Mechanism of adaptation of an atypical alkaline p-nitrophenyl phosphatase from the archaeon Halobacterium salinarum at low-water environments. AB - Enzymes suspended in organic solvents represent a versatile system for studying the involvement of water in catalytic properties and their flexibility in adapting to different environmental conditions. The extremely halophilic alkaline p-nitrophenylphosphate phosphatase from the archaeon Halobacterium salinarum was solubilized in an organic medium consisting of reversed micelles of hexadecyltrimethylammoniumbromide in cyclohexane, with 1-butanol as cosurfactant. Hydrolysis of p-nitrophenylphosphate was nonlinear with time when the enzyme was microinjected into reversed micelles that contained substrate. These data are consistent with a kinetic model in which the enzyme is irreversibly converted from an initial form to a final stable form during the first seconds of the encapsulation process. The model features a rate constant (k) for that transition and separate hydrolysis rates, v(1) and v(2), for the two forms of the enzyme. The enzyme conversion may be governed by the encapsulation process. PMID- 12115119 TI - Measuring the pH environment of DNA delivered using nonviral vectors: implications for lysosomal trafficking. AB - The degradation of DNA in lysosomes represents a major obstacle to efficient nonviral gene delivery. The rational design of vectors that overcome this obstacle requires a better understanding of the lysosomal barrier to gene delivery, which in turn requires a means to investigate this intermediate step. To this end, we developed a technique to measure the pH environment of delivered DNA, from which the degree to which vectors avoided trafficking to acidic Iysosomes could be determined. The measured average pH of DNA delivered using poly-L-lysine (PLL) polyplexes was 4.5, suggesting that PLL polyplexes were trafficked to acidic lysosomes. Other vectors could avoid or buffer the pH of Iysosomes as DNA delivered using Lipofectamine Plus, polyethylenimine (PEI), linear polyethylenimine (LPEI), and two degradable poly(beta-amino ester)s (poly 1 and poly-2) had average pH values of 7.1, 5.9, 5.0, 6.7, and 6.4, respectively. PMID- 12115120 TI - Lipase-catalyzed acidolysis of menhaden oil with conjugated linoleic acid: effect of water content. AB - The effect of the water content on the lipase-catalyzed (Candida rugosa) interesterification (acidolysis) of menhaden oil with conjugated linoleic acid was studied for amounts of added water ranging from 0-4% (w/w). The rate of the acidolysis reaction increased with increasing water content, but the corresponding percentage of n-3 fatty acids liberated also increased. The implications of water content for minimization of the release of n-3 fatty acid residues while maximizing incorporation of CLA are discussed. PMID- 12115121 TI - Rationally designed fluorescently labeled sulfate-binding protein mutants: evaluation in the development of a sensing system for sulfate. AB - Periplasmic binding proteins from E. coli undergo large conformational changes upon binding their respective ligands. By attaching a fluorescent probe at rationally selected unique sites on the protein, these conformational changes in the protein can be monitored by measuring the changes in fluorescence intensity of the probe which allow the development of reagentless sensing systems for their corresponding ligands. In this work, we evaluated several sites on bacterial periplasmic sulfate-binding protein (SBP) for attachment of a fluorescent probe and rationally designed a reagentless sensing system for sulfate. Eight different mutants of SBP were prepared by employing the polymerase chain reaction (PCR) to introduce a unique cysteine residue at a specific location on the protein. The sites Gly55, Ser90, Ser129, Ala140, Leu145, Ser171, Val181, and Gly186 were chosen for mutagenesis by studying the three-dimensional X-ray crystal structure of SBP. An environment-sensitive fluorescent probe (MDCC) was then attached site specifically to the protein through the sulfhydryl group of the unique cysteine residue introduced. Each fluorescent probe-conjugated SBP mutant was characterized in terms of its fluorescence properties and Ser171 was determined to be the best site for the attachment of the fluorescent probe that would allow for the development of a reagentless sensing system for sulfate. Three different environment-sensitive fluorescent probes (1,5-IAEDANS, MDCC, and acylodan) were studied with the SBP171 mutant protein. A calibration curve for sulfate was constructed using the labeled protein and relating the change in the fluorescence intensity with the amount of sulfate present in the sample. The detection limit for sulfate was found to be in the submicromolar range using this system. The selectivity of the sensing system was demonstrated by evaluating its response to other anions. A fast and selective sensing system with detection limits for sulfate in the submicromolar range was developed. PMID- 12115122 TI - Monitoring of complex industrial bioprocesses for metabolite concentrations using modern spectroscopies and machine learning: application to gibberellic acid production. AB - Two rapid vibrational spectroscopic approaches (diffuse reflectance-absorbance Fourier transform infrared [FT-IR] and dispersive Raman spectroscopy), and one mass spectrometric method based on in vacuo Curie-point pyrolysis (PyMS), were investigated in this study. A diverse range of unprocessed, industrial fed-batch fermentation broths containing the fungus Gibberella fujikuroi producing the natural product gibberellic acid, were analyzed directly without a priori chromatographic separation. Partial least squares regression (PLSR) and artificial neural networks (ANNs) were applied to all of the information-rich spectra obtained by each of the methods to obtain quantitative information on the gibberellic acid titer. These estimates were of good precision, and the typical root-mean-square error for predictions of concentrations in an independent test set was <10% over a very wide titer range from 0 to 4925 ppm. However, although PLSR and ANNs are very powerful techniques they are often described as "black box" methods because the information they use to construct the calibration model is largely inaccessible. Therefore, a variety of novel evolutionary computation based methods, including genetic algorithms and genetic programming, were used to produce models that allowed the determination of those input variables that contributed most to the models formed, and to observe that these models were predominantly based on the concentration of gibberellic acid itself. This is the first time that these three modern analytical spectroscopies, in combination with advanced chemometric data analysis, have been compared for their ability to analyze a real commercial bioprocess. The results demonstrate unequivocally that all methods provide very rapid and accurate estimates of the progress of industrial fermentations, and indicate that, of the three methods studied, Raman spectroscopy is the ideal bioprocess monitoring method because it can be adapted for on-line analysis. PMID- 12115123 TI - Contribution of aerial hyphae of Aspergillus oryzae to respiration in a model solid-state fermentation system. AB - Oxygen transfer is for two reasons a major concern in scale-up and process control in industrial application of aerobic fungal solid-state fermentation (SSF): 1) heat production is proportional to oxygen uptake and it is well known that heat removal is one of the main problems in scaled-up fermenters, and 2) oxygen supply to the mycelium on the surface of or inside the substrate particles may be hampered by diffusion limitation. This article gives the first experimental evidence that aerial hyphae are important for fungal respiration in SSF. In cultures of A. oryzae on a wheat-flour model substrate, aerial hyphae contributed up to 75% of the oxygen uptake rate by the fungus. This is due to the fact that A. oryzae forms very abundant aerial mycelium and diffusion of oxygen in the gas-filled pores of the aerial hyphae layer is rapid. It means that diffusion limitation in the densely packed mycelium layer that is formed closer to the substrate surface and that has liquid-filled pores is much less important for A. oryzae than was previously reported for R. oligosporus and C. minitans. It also means that the overall oxygen uptake rate for A. oryzae is much higher than the oxygen uptake rate that can be predicted in the densely packed mycelium layer for R. oligosporus and C. minitans. This would imply that cooling problems become more pronounced. Therefore, it is very important to clarify the physiological role of aerial hyphae in SSF. PMID- 12115124 TI - Effect of oxygen transfer on glycerol biosynthesis by an osmophilic yeast Candida magnoliae I(2)B. AB - The influence of oxygen on glycerol production by an osmophilic yeast, Candida magnoliae I(2)B, was studied in a bioreactor. Oxygen transfer rates (OTRs) and volumetric oxygen transfer coefficients (k(L)a) were determined at different aeration and agitation rates. Cell growth as well as glycerol production was strongly affected by oxygen supply. Improvement in OTRs resulted in increased cell growth and glycerol yield. However, at high OTRs, there was a reduction in glucose uptake rate, indicating Pasteur Effect, and glycerol accumulation was also reduced at k(L)a of 253 h(-1). The availability of oxygen per unit of cell mass was found to be the most important factor that controlled cell growth, glucose uptake, and glycerol yield. The overall productivity and yield of glycerol could be related with k(L)a. The biosynthesis of glycerol was found to both growth- and non-growth-associated, although glycerol was mainly produced in post-exponential phase. PMID- 12115125 TI - Glass micromodel study of bacterial dispersion in spatially periodic porous networks. AB - Successful implementation of bioremediation clean-up strategies depends on accurate predictions of the transport of bacteria within the subsurface. In this study, etched flat-plate glass micromodels were used to examine bacterial transport in a homogenous network. These networks were created by acid-etching interconnected channels into a glass plate and then fusing it to an unetched plate forming semi-cylindrical pores. The transparent nature of the micromodel allows for both qualitative observations of the bacteria within the pores and quantitative measurements of their concentration. The micromodels are designed to allow establishment of a well-characterized step change in bacterial concentration (Escherichia coli NR50) within the network. During the experiments, bacteria are dispersed through the network by flow. Light scattering is used to detect the change in turbidity within the pores as the bacteria travel through the network. The change in turbidity is used to construct breakthrough curves and spatial concentration profiles of bacteria within the network. The breakthrough curves are fit to the one-dimensional advection/dispersion equation to determine dispersion coefficients at different interstitial fluid velocities. From the breakthrough curves, dispersion coefficients were reproducible for replicate experiments over a range of velocities in the advection-dominated regime. The dispersivity values for two network designs resembling an interconnecting capillary network and a spatially periodic network of cylinders were 0.28 and 0.33 cm respectively, which are slightly greater than the literature values found for other pore networks. Experiments were also conducted within the diffusion dominated regime to examine the effects of bacterial motility on dispersion. The accumulation of bacteria on the pore walls became significant at the low flow rates and extended experimental times thereby rendering the use of light scattering to determine concentrations ineffective. Bacterial chemotaxis, created by a self-imposed oxygen gradient, was also observed in the micromodel under stagnant fluid conditions. PMID- 12115126 TI - Arylsulfotransferase from Clostridium innocuum-A new enzyme catalyst for sulfation of phenol-containing compounds. AB - Arylsulfotransferase (AST, EC 2.8.2.22), an enzyme capable of sulfating a wide range of phenol-containing compounds was purified from a Clostridium innocuum isolate (strain 554). The enzyme has a molecular weight of 320 kDa and is composed of four subunits. Unlike many mammalian and plant arylsulfotransferases, AST from Clostridium utilizes arylsulfates, including p-nitrophenyl sulfate, as sulfate donors, and is not reactive with 3-phosphoadenosine-5'-phosphosulfate (PAPS). The enzyme possesses broad substrate specificity and is active with a variety of phenols, quinones and flavonoids, but does not utilize primary and secondary alcohols and sugars as substrates. Arylsulfotransferase tolerates the presence of 10 vol% of polar cosolvents (dimethyl formamide, acetonitrile, methanol), but loses significant activity at higher solvent concentrations of 30 40 vol%. The enzyme retains high arylsulfotransferase activity in biphasic systems composed of water and nonpolar solvents, such as cyclohexane, toluene and chloroform, while in biphasic systems with more polar solvents (ethyl acetate, 2 pentanone, methyl tert-butyl ether, and butyl acetate) the enzyme activity is completely lost. High yields of AST-catalyzed sulfation were achieved in reactions with several phenols and tyrosine-containing peptides. Overall, AST studied in this work is a promising biocatalyst in organic synthesis to afford efficient sulfation of phenolic compounds under mild reaction conditions. PMID- 12115127 TI - A novel model of solute transport in a hollow-fiber bioartificial pancreas based on a finite element method. AB - Extravascular bioartificial pancreas based on hollow fiber seems to be a promising treatment of diabetes mellitus. However, solutes mass-transport limitations in such a device could explain its lack of success. To determine critical device parameters, we have developed a novel tridimensional model based on finite element method for glucose, insulin, and oxygen diffusion around an islet of Langerhans encapsulated in a hollow-fiber section. A glucose ramp stimulation was applied outside the fiber and diffused to the islet. Concomitantly, a stationary oxygen partial pressure was applied outside the fiber, and determined local oxygen partial pressure on the islet environment. An insulin secretion model stimulated by a glucose concentration ramp and corrected by the local oxygen partial pressure was also implemented. Insulin secretion by the islet was thus computed as a response to glucose signal. The model predictions notably showed that the fiber radius had to be small enough to favor a fast response for insulin secretion and to ensure a maximal oxygen partial pressure in the islet environment. Besides the effect of fiber radius, a better islet oxygenation could be achieved by adjustments on the islet density, i.e., on the fiber length dedicated to a single islet. These hints should allow the future proposal of an optimal design for an implantable bioartificial pancreas. PMID- 12115128 TI - Microbial synthesis of semiconductor CdS nanoparticles, their characterization, and their use in the fabrication of an ideal diode. AB - Cadmium sulfide nanoparticles were synthesized intracellularly by a Schizosaccharomyces pombe strain when challenged with 1 mM cadmium in solution. The nanoparticles, a known semiconducting material, exhibited an absorbance maximum at 305 nm. X-ray scattering data showed that the nanoparticles had a Wurtzite (Cd(16)S(20))-type hexagonal lattice structure and most of the nanoparicles were in the size range of 1-1.5 nm. The nanoparticles were used in the fabrication of a heterojunction with poly (p-phenylenevinylene). The diode exhibited approximately 75 mA/cm(2) current at 10 V when forward biased and the breakdown occurred at approximately 15 V in the reverse biased mode. These characteristics are considered ideal for a diode. PMID- 12115129 TI - Penicillin acylase-catalyzed ampicillin synthesis using a pH gradient: a new approach to optimization. AB - The penicillin acylase-catalyzed synthesis of ampicillin by acyl transfer from D (-)-phenylglycine amide (D-PGA) to 6-aminopenicillanic acid (6-APA) becomes more effective when a judiciously chosen pH gradient is applied in the course of the process. This reaction concept is based on two experimental observations: 1) The ratio of the initial synthesis and hydrolysis rates (V(S)/V(H)) is pH-dependent and exhibits a maximum at pH 6.5-7.0 for a saturated solution of 6-APA; 2) at a fixed 6-APA concentration below saturation, V(S)/V(H) increases with decreasing pH. Optimum synthetic efficiency could, therefore, be achieved by starting with a concentrated 6-APA solution at pH 7 and gradually decreasing the pH to 6.3 in the course of 6-APA consumption. A conversion of 96% of 6-APA and 71% of D-PGA into ampicillin was accomplished in an optimized procedure, which significantly exceeds the efficiency of enzymatic synthesis performed at a constant pH of either 7.0 or 6.3. PMID- 12115130 TI - Intermolecular Overhauser effects in fluoroalcohol solutions of cyclo alanylglycine. AB - Interactions between the diketopiperazine cyclo-alanylglycine and four fluorinated alcohols in water-fluoroalcohol mixtures were examined by (1)H[(19)F] intermolecular nuclear Overhauser effects (NOE) experiments. The alcohols studied were trifluoroethanol, hexafluoroacetone trihydrate, 1,1,1,3,3,3 hexafluoroisopropanol and perfluoro-t-butanol. The experimental methods used permit detection of solvent-solute NOEs of 0.1% or less. Solute and solvent diffusion coefficients were determined and apparent molecular radii of the fluoroalcohols estimated. Using these data, it was shown that observed (1)H[(19)F] intermolecular NOEs are consistent with expectations based on theory. A method for extending conventional theory to take into account the shape of a solute and the exposure of its hydrogens to solvent is described. This approach gives reasonable agreement with experimental results, particularly if it is assumed that solute-solvent interactions take place in such a way that the fluorines of a fluoroalcohol are preferentially oriented toward the solute during solute-solvent encounters. The results support the suggestion that intermolecular (1)H[(19)F] NOEs may become a useful tool for studies of peptide and protein conformations in fluoroalcohol-water solvent mixtures. PMID- 12115131 TI - Factors governing 3(10)-helix vs alpha-helix formation in peptides: percentage of C(alpha)-tetrasubstituted alpha-amino acid residues and sequence dependence. AB - As an additional step toward the dissection of the factors responsible for the onset of 3(10)-helix vs alpha-helix in peptides, in this paper we describe the results of a three-dimensional (3D) structural analysis by x-ray diffraction of the N(alpha)-acylated heptapeptide alkylamide mBrBz-L-Iva-L-(alphaMe)Val-L-Abu-L (alphaMe)Val-L-(alphaMe)Phe-L-(alphaMe)Val-L-Iva-NHMe characterized by a single (L-Abu3) C(alpha)-trisubstituted and six C(alpha)-tetrasubstituted alpha-amino acids. We find that in the crystal state this peptide is folded in a mixed helical structure with short elements of 3(10)-helix at either terminus and a central region of alpha-helix. This finding, taken together with the published NMR and x-ray diffraction data on the all C(alpha)-methylated parent sequence and its L-Val2 analog (also the latter heptapeptide has a single C(alpha) trisubstituted alpha-amino acid) strongly supports the view that one C(alpha) trisubstituted alpha-amino acid inserted near the N-terminus of an N(alpha) acylated heptapeptide alkylamide sequence may be enough to switch a regular 3(10) helix into an essentially alpha-helical conformation. As a corollary of this work, the x-ray diffraction structure of the N(alpha)-protected, C-terminal tetrapeptide alkylamide Z-L-(alphaMe)Val-L-(alphaMe)Phe-L-(alphaMe)Val-L-Iva NHMe, also reported here, is clearly indicative of the preference of this fully C(alpha)-methylated, short peptide for the 3(10)-helix. As the same terminally blocked sequence is mixed 3(10)/alpha-helical in the L-Abu3 heptapeptide amide but regular 3(10)-helical in the tetrapeptide amide and in the parent heptapeptide amide, these results point to an evident plasticity even of a fully C(alpha)-methylated short peptide. PMID- 12115132 TI - Determination of alpha-helix and beta-sheet stability in the solid state: a solid state NMR investigation of poly(L-alanine). AB - The relative stability of alpha-helix and beta-sheet secondary structure in the solid state was investigated using poly(L-alanine) (PLA) as a model system. Protein folding and stability has been well studied in solution, but little is known about solid-state environments, such as the core of a folded protein, where peptide packing interactions are the dominant factor in determining structural stability. (13)C cross-polarization with magic angle spinning (CPMAS) NMR spectroscopy was used to determine the backbone conformation of solid powder samples of 15-kDa and 21.4-kDa PLA before and after various sample treatments. Reprecipitation from helix-inducing solvents traps the alpha-helical conformation of PLA, although the method of reprecipitation also affects the conformational distribution. Grinding converts the secondary structure of PLA to a final steady state mixture of 55% beta-sheet and 45% alpha-helix at room temperature regardless of the initial secondary structure. Grinding PLA at liquid nitrogen temperatures leads to a similar steady-state mixture with 60% beta-sheet and 40% alpha-helix, indicating that mechanical shear force is sufficient to induce secondary structure interconversion. Cooling the sample in liquid nitrogen or subjecting it to high pressure has no effect on secondary structure. Heating the sample without grinding results in equilibration of secondary structure to 50% alpha-helix/50% beta-sheet at 100 degrees C when starting from a mostly alpha helical state. No change was observed upon heating a beta-sheet sample, perhaps due to kinetic effects and the different heating rate used in the experiments. These results are consistent with beta-sheet approximately 260 J/mol more stable than alpha-helix in solid-state PLA. PMID- 12115133 TI - Structural analysis of peptide helices containing centrally positioned lactic acid residues. AB - The effect of insertion of lactic acid (Lac) residues into peptide helices has been probed using specifically designed sequences. The crystal structures of 11 residue and 14-residue depsipeptides Boc-Val-Val-Ala-Leu-Val-Lac-Leu-Aib-Val-Ala Leu-OMe (1) and Boc-Val-Ala-Leu-Aib-Val-Ala-Leu-Val-Lac-Leu-Aib-Val-Ala-Leu-OMe (3), containing centrally positioned Lac residues, have been determined. The structure of an 11-residue peptide Boc-Val-Ala-Leu-Aib-Val-Ala-Leu-Aib-Val-Ala Leu-OMe (2), analog of a which is an amide previously determined Lac-containing depsipeptide, Boc-Val-Ala-Leu-Aib-Val-Lac-Leu-Aib-Val-Ala-Leu-OMe I. L. Karle, C. Das, and P. Balaram, Biopolymers, Vol. 59, (2001) pp. 276-289], is also reported. Peptide 1 adopts a helical fold, which is stabilized by mixture of 4-->1 and 5- >1 hydrogen bonds. Peptide 2 adopts a completely alpha-helical conformation stabilized by eight successive 5-->1 hydrogen bonds. Peptide 3 appears to be predominately alpha-helical, with seven 5-->1 hydrogen bonds and three 4-->1 interaction interspersed in the sequence. In the structure of peptide 3 in addition to water molecules in the head-to-tail region, hydration at an internal segment of the helix is also observed. A comparison of five related peptide helices, containing a single Lac residue, reveals that the hydroxy acid can be comfortably accommodated at interior positions in the helix, with the closest C=O...O distances lying between 2.8 and 3.3 A. PMID- 12115134 TI - Structural studies of starches with different water contents. AB - The proportion of double helices in starches from a series of pea [rb, rug4-b, rug3-a, and lam-c mutants, and the wild type (WT) parental line], potato and maize (normal and low amylose), and wheat (normal) lines, ranged from about 30 50% on a dry weight basis. In relatively dry starch powders, only about half of the double helices were in crystalline order, this proportion being higher for A type than for B-type starches. Using starch from WT pea as an example, it was found that increasing water content results in an increase in total crystallinity. When the water content was raised to a level similar to that in excess water, the proportion of crystallinity was close to the proportion of double helices (DH). Measuring crystallinity in starches with a high water content is difficult using traditional methods such as x-ray diffraction. A method was developed, therefore, for determining starch structural characteristics in excess water by measuring the enthalpy of gelatinization transition in quasi-equilibrium differential scanning calorimetry (DSC) experiments. It is suggested that DH% = DeltaH(sp)/DeltaH(DH) x 100%, where DeltaH(sp) and DeltaH(DH) represent the specific enthalpies of gelatinisation transition, DeltaH(sp) being measured as J/g dry starch weight and DeltaH(DH) as J/g DH, in starch. Studies on potato and maize starches in excess water and in 0.6M KCl showed, respectively, that DeltaH(DH) was 36.3 and 35.6 J/g for B-type polymorphs and 33.0 and 35.0 J/g for A-type polymorphs. For C-type starches, such as those from pea, intermediate values of DeltaH(DH), related to the proportions A-/B-polymorphs, should be used. The type of crystallinity in starch can be determined by the shift in peak temperature for thermograms in excess water and in excess 0.6M KCl. For B-polymorphs this shift was found to be approximately 2-3 degrees C and for A-polymorphs approximately 7-12 degrees C. The ratio between ordered areas with both A- and B-polymorphs can be determined from the enthalpies of disruption of each area. These enthalpies can be obtained by deconvolution of bimodal thermograms produced by C-type starches in excess 0.6M KCl. This methodical approach can be applied to all starches that give a sharp gelatinisation thermogram in excess water. Using a range of methods, including DSC, it was found that starch granules from the mutant peas are constructed in a similar way to those from the WT, with B-polymorphs in the centre and A polymorphs at the periphery of all granules. The proportion of A/B-polymorphs, however, differed between the mutants. It was found that in addition to increasing the total crystallinity, increasing the water content within the granules also resulted in an increase in the proportion of B-polymorphs. PMID- 12115135 TI - A conformational analysis of leucine enkephalin as a function of pH. AB - The conformations of Leu enkephalin in aqueous solution have been investigated as a function of pH using molecular dynamics simulations. The simulations suggest the peptide backbone exists as a mixture of folded and unfolded forms (approximately 50% each) at neutral pH, but is always unfolded at low or high pH. The folded form at neutral pH possesses a 2 --> 5 hydrogen bond and a close head to tail separation. No significant intramolecular hydrogen bonding of the carbonyl oxygens was observed in either the folded or unfolded forms of the peptide. Analysis of the Gly carbonyl oxygens and terminal groups indicated that, while the conformational population distribution of Leu enkephalin did vary noticeably as a function of pH, their hydration was essentially independent of pH and in agreement with the available NMR data. Further study indicated that the unfolded state of the peptide was not random in nature and consisted of one major unfolded backbone arrangement stabilized by a persistent hydrophobic interaction between the side chains of Tyr and Leu. PMID- 12115136 TI - Selectivity in heavy metal- binding to peptides and proteins. AB - The metal-binding affinities and three-dimensional structures of three synthetic 18-residue peptides with sequences derived from that of the highly conserved metal-binding motif MXCXXC found in many heavy metal-binding proteins were determined. A change in register of the cysteines and alanines of the sequence from the periplasmic mercury-binding protein, MerP, i.e., CAAC, CACA, and CCAA, affects the specificity of metal binding, in particular, the peptide with vicinal cysteines binds only mercury. The three-dimensional structures of the mercury bound forms of the three peptides determined in solution by NMR spectroscopy peptides differ considerably, even though they are all linear bicoordinate complexes. The three-dimensional structure of the peptide with CAAC bound to Cd(II) demonstrates that the metal-binding loop is malleable enough to accommodate modes of coordination other than linear bicoordinate. PMID- 12115137 TI - Solution structure of a D,L-alternating oligonorleucine as a model of double stranded antiparallel beta-helix. AB - Conformational characteristics of alternating D,L linear peptides are of particular interest because of their capacity to form transmembrane channels with different transport properties, as some natural antibiotics do. Single- and double-stranded beta-helical structures are common for alternating D,L peptides. The stability of the beta-helix depends on several structural factors, such as the backbone peptide length, type and position of side chains, and nature of terminal groups. The NMR and molecular dynamics solution conformation of a synthetic alternating D,L-oligopeptide with 15 norleucines (XVMe) has been used as a model to get insight in to the conformational features of double-stranded beta-helix structures. The NH chemical shift values (delta(NH)) and long-range nuclear Overhauser effects (NOE) cross peaks, in particular interstrand connectivities, clearly point to an antiparallel double-stranded beta-helix for the XVMe major conformation in solution. An extensive set of distances (from NOE cross peaks) and H-bonds (from delta(NH)) has been included in the molecular dynamics calculations. The experimental NMR data and theoretical calculations clearly indicate that the most probable conformation of XVMe in solution is a double-strand antiparallel beta(5.6) increasing decreasing-helix structure. PMID- 12115138 TI - Importance of mutant position in Ramachandran plot for predicting protein stability of surface mutations. AB - Understanding the mechanisms by which mutations affect protein stability is one of the most important problems in molecular biology. In this work, we analyzed the relationship between changes in protein stability caused by surface mutations and changes in 49 physicochemical, energetic, and conformational properties of amino acid residues. We found that the hydration entropy was the major contributor to the stability of surface mutations in helical segments; other properties responsible for size and volume of molecule also correlated significantly with stability. Classification of coil mutations based on their locations in the (phi-psi) map improved the correlation significantly, demonstrating the existence of a relationship between stability and strain energy, which indicates that the role of strain energy is very important for the stability of surface mutations. We observed that the inclusion of sequence and structural information raised the correlation, indicating the influence of surrounding residues on the stability of surface mutations. Further, we examined the previously reported "inverse relationship" between stability and hydrophobicity, and observed that the inverse hydrophobic effect was generally applicable only to coil mutations. The present study leads to a simple method for predicting protein stability changes caused by amino acid substitutions, which will be useful for protein engineering in designing novel proteins with increased stability and altered function. PMID- 12115139 TI - Structural transformation and aggregation of human alpha-synuclein in trifluoroethanol: non-amyloid component sequence is essential and beta-sheet formation is prerequisite to aggregation. AB - Amyloid-like aggregation of alpha-synuclein and deposit in Lewy bodies are thought to be the major cause of Parkinson's disease. Here we describe the secondary structural transformation and aggregation of human alpha-synuclein and its C-terminus truncated fragments in trifluoroethanol. Proteins containing the NAC (non-amyloid component) segment undergo a three-state transition: from native random coil to beta-sheet and to alpha-helical structure, while the NAC deficient fragment and gamma-synuclein undergo a typical two-state coil-to-alpha transition. The beta-sheet form is highly hydrophobic that strongly binds to 1 anilinonaphthalene-8-sulfonic acid (ANS) and is prone to self-aggregation. The results suggest that the NAC sequence is essential to beta-sheet formation and the aggregation originates from the beta-sheet intermediate, which may be implicated in the pathogenesis of Parkinson's disease. PMID- 12115140 TI - Vibrational CD (VCD) and atomic force microscopy (AFM) study of DNA interaction with Cr3+ ions: VCD and AFM evidence of DNA condensation. AB - The interaction of natural calf thymus DNA with Cr(3+) ions was studied at room temperature by means of vibrational CD (VCD) and infrared absorption (ir) spectroscopy, and atomic force microscopy (AFM). Cr(3+) ion binding mainly to N(7) (G) and to phosphate groups was demonstrated. Psi-type VCD spectra resembling electronic CD (ECD) spectra, which appear during psi-type DNA condensation, were observed. These spectra are characterized mainly by an anomalous, severalfold increase of VCD intensity. Such anomalous VCD spectra were assigned to DNA condensation with formation of large and dense particles of a size comparable to the wavelength of the probing ir beam and possessing large scale helicity. Atomic force microscopy confirmed DNA condensation by Cr(3+) ions and the formation of tight DNA particles responsible for the psi-type VCD spectra. Upon increasing the Cr(3+) ion concentration the shape of the condensates changed from loose flower-like structures to highly packed dense spheres. No DNA denaturation was seen even at the highest concentration of Cr(3+) ions studied. The secondary structure of DNA remained in a B-form before and after the condensation. VCD and ir as well as AFM proved to be an effective combination for investigating DNA condensation. In addition to the ability of VCD to determine DNA condensation, VCD and ir can in the same experiment provide unambiguous information about the secondary structure of DNA contained in the condensed particles. PMID- 12115141 TI - DNA persistence length revisited. AB - DNA restriction fragments ranging from 79 to 789 base pairs in length have been characterized by transient electric birefringence (TEB) measurements at various temperatures between 4 and 43 degrees C. The DNA fragments do not contain runs of four or more adenine residues in a row and migrate with normal electrophoretic mobilities in polyacrylamide gels, indicating that they are not intrinsically curved or bent. The low ionic strength buffers used for the measurements contained 1 mM Tris Cl, pH 8.0, EDTA, and variable concentrations of Na(+) or Mg(2+) ions. The rotational relaxation times were obtained by fitting the TEB field-free decay signals with a nonlinear least-squared fitting program; the decay of the birefringence was monoexponential for fragments < or = 241 base pair (bp) in length and multiexponential for larger fragments. The terminal relaxation times, characteristic of the end-over-end rotation of the DNA molecules, were then used to determine the persistence length (p) and hydrodynamic radius (r) of DNA as a function of temperature and ionic strength, using several different hydrodynamic models. The specific values obtained for p and r are model dependent. The wormlike chain model of P. J. Hagerman and B. H. Zimm (Biopolymers 1981, Vol. 20, pp. 1481-1502) combined with the revised Broersma equation (J. Newman et al., Journal of Mol Biol 1997, Vol. 116, pp. 593-606) appears to be the most suitable for describing the flexibility of DNA in low ionic strength solutions. The values of p and r obtained from the global least squares fitting of this equation are independent of DNA length, and the deviations of the individual values from the average are reasonably small. The consensus r value calculated for DNA in various low ionic strength solutions containing 1 mM Tris buffer is 14.7 +/- 0.4 A at 20 degrees C. The consensus p values decrease from 814 approximately 564 A in solutions containing 1 mM Tris buffer plus 0.2-1 mM NaCl and decrease still further to 440 A in solutions containing 0.2 mM Mg(2+) ions. The persistence length exhibits a shallow maximum at 20 degrees C and decreases slowly upon either increasing or decreasing the temperature, regardless of the model used to fit the data. By contrast, the consensus values of the hydrodynamic radius are independent of temperature. The calculated persistence lengths and hydrodynamic radii are compared with other data in the literature. PMID- 12115142 TI - Influence of netropsin's charges on the minor groove width of d(CGCGAATTCGCG)2. AB - The exact understanding of the interaction of minor groove binding drugs with DNA is of interest due to their importance as transcription controlling drugs. In this study we performed four molecular dynamics simulations, one of the uncomplexed d(CGCGAATTCGCG)(2) dodecamer and three simulations of the DNA complexed with the minor groove binder netropsin. The charged guanidinium and amidinium ends of the small ligand were in one simulation formally uncharged, in the second one normally charged, and in the third simulation we doubled the charges of the two ends. So we are able to filter out the influence the charges exert on the DNA structure. The positive charges reduce the width of the minor groove showing that charges are able to modify the groove width by charge neutralization of the negative phosphate groups. The quality of the used force field was successfully tested by comparing the results of the uncomplexed dodecamer with already reported NMR and x-ray studies. Thus our simulations should be able to describe the minor groove width of DNA in a correct manner underlying the validity of the results. PMID- 12115143 TI - Solid-state NMR relaxation studies of Australian spider silks. AB - Solid-state NMR techniques were used to study two different types of spider silk from two Australian orb-web spider species, Nephila edulis and Argiope keyserlingi. A comparison of (13)C-T(1) and (1)H-T(1rho) solid-state NMR relaxation data of the Ala Calpha, Ala Cbeta, Gly Calpha, and carbonyl resonances revealed subtle differences between dragline and cocoon silk. (13)C-T(1rho) and (1)H-T(1) relaxation experiments showed significant differences between silks of the two species with possible structural variations. Comparison of our data to previous (13)C-T(1) relaxation studies of silk from Nephila clavipes (A. Simmons et al., Macromolecules, 1994, Vol. 27, pp. 5235-5237) also supports the finding that differences in molecular mobility of dragline silk exist between species. Interspecies differences in silk structure may be due to different functional properties. Relaxation studies performed on wet (supercontracted) and dry silks showed that the degree of hydration affects relaxation properties, and hence changes in molecular mobility are correlated with functional properties of silk. PMID- 12115145 TI - Peptide fragments as models to study the structure of a G-protein coupled receptor: the alpha-factor receptor of Saccharomyces cerevisiae. AB - The alpha-factor tridecapeptide initiates mating in Saccharomyces cerevisiae upon interaction with Ste2p, its cognate G-protein coupled receptor (GPCR). This interaction is being used as a paradigm for understanding the structure and mechanism of activation of GPCRs by medium-sized peptides. In this article, the use of fragments of Ste2p to study its structure is reviewed. Methods of synthesis of peptides corresponding to both extramembranous and transmembrane domains of Ste2p are evaluated and problems that are encountered during synthesis and purification are described. The results from conformational analyses of the peptide fragments using fluorescence spectroscopy, CD, infrared spectroscopy, and NMR spectroscopy in organic-aqueous mixtures and in the presence of detergent micelles and lipid bilayers are critically reviewed. The data obtained to date provide biophysical evidence for the structure of different domains of Ste2p and indicate that peptides corresponding to these domains have unique biophysical tendencies. The studies carried out on Ste2p fragments indicate that valuable information concerning the structure of the intact receptor can be obtained by studying peptide fragments corresponding to domains of these polytopic integral membrane proteins. PMID- 12115146 TI - Controls exerted by the Aib residue: helix formation and helix reversal. AB - The helix forming properties of the achiral alpha-amino isobutyric residue (Aib) have been demonstrated by numerous crystal structure analyses of designed and naturally occurring peptides containing one or more Aib residues in the sequence. Experimental and computational results concerning the type of helix obtained, whether the 3(10)-helix with 4 --> 1 type hydrogen bonds or the alpha-helix with 5 --> 1 hydrogen bonds or mixtures of the two, have been published. This paper deals with residues that, if inserted into a sequence, could perturb the helix forming propensity afforded by the presence of Aib residues. Examples of structures will be presented in which Pro, Hyp, Gly-Gly, d-Ala-Gly, and Lac have been centrally placed in the sequence. In addition to the formation of helices, detailed experimentally obtained conformation information is presented for the role of the Aib residue in reversing the sense of the helix (the Schellman motif) with the consequent formation of the 6 --> 1 type hydrogen bond or a solvated 6 - > 1 hydrogen bond. Data are presented for 13 molecules with helix reversals at the C-terminus or near the center of the sequence. PMID- 12115147 TI - Stabilization of alpha-helix structure by polar side-chain interactions: complex salt bridges, cation-pi interactions, and C-H em leader O H-bonds. AB - It is generally understood that helical proteins are stabilized by a combination of hydrophobic and packing interactions, together with H-bonds and electrostatic interactions. Here we show that polar side-chain interactions on the surface can play an important role in helix formation and stability. We review studies on model helical peptides that reveal the effect of weak interactions between side chains on helix stability, focusing on some nonclassical side-chain-side-chain interactions: complex salt bridges, cation-pi, and C-H em leader O H-bonding interactions. Each of these can be shown to contribute to helix stability, and thus must be included in a comprehensive catalogue of helix stabilizing effects. The issue of the structure of the unfolded states of helical peptides is also discussed, in the light of recent experiments showing that these contain substantial amounts of polyproline II conformation. PMID- 12115148 TI - Alzheimer beta-amyloid peptides: structures of amyloid fibrils and alternate aggregation products. AB - The amyloidoses are a heterogeneous group of diseases, which are characterized by the local or systemic deposition of amyloid. At the root of these diseases are changes in protein conformation where normal innocuous proteins transform into insoluble amyloid fibrils and deposit in tissues. The amyloid fibrils of Alzheimer's disease are composed of the Abeta peptide and deposit in the form of senile plaques. Neurodegeneration surrounds the amyloid deposits, indicating that neurotoxic substances are produced during the deposition process. Whether the neurotoxic species is the amyloid fibril or a fibril precursor is a current area of active research. This review focuses on advancements made in elucidating the molecular structures of the Abeta amyloid fibril and alternate aggregation products of the Abeta peptide formed during fibrillogenesis. PMID- 12115149 TI - Malignant lymphoma presenting as Tietze's syndrome. PMID- 12115150 TI - Focusing interventions for disability among patients with rheumatoid arthritis. PMID- 12115151 TI - Medication toxicity among patients with ankylosing spondylitis. AB - OBJECTIVE: To determine the role of medication toxicity in the discontinuation of antirheumatic treatment among patients with ankylosing spondylitis (AS), and to compare the toxicity of different medications. METHODS: In a prospective longitudinal study of 241 patients with AS, we examined the duration of treatment and discontinuations due to side effects of new courses of sulfasalazine, methotrexate, ibuprofen, naproxen, indomethacin, diclofenac, piroxicam, nabumetone, and celecoxib. RESULTS: Of the 241 patients, 167 reported having a new treatment course of either sulfasalazine (n = 49), methotrexate (n = 19), ibuprofen (n = 105), naproxen (n = 57), indomethacin (n = 50), diclofenac (n = 38), piroxicam (n = 34), nabumetone (n = 27), or celecoxib (n = 25), for a total of 404 new treatment courses. Side effects were reported in 6.7% (ibuprofen) to 47.3% (methotrexate) of the courses. Between 2% (ibuprofen) and 23.5% (piroxicam) of courses were discontinued due to toxicity. For each medication, the duration of treatment was most often limited by factors other than toxicity. The time to drug discontinuation for any reason and the time to discontinuation due to toxicity did not differ between sulfasalazine and methotrexate. The time to drug discontinuation for any reason did not differ among nonsteroidal antiinflammatory drugs (NSAIDs), but discontinuations due to toxicity occurred earlier with piroxicam than with other NSAIDs. CONCLUSION: Although medication toxicity is common among patients with AS, it is an uncommon cause of discontinuation of antirheumatic treatment. PMID- 12115152 TI - Use of a numerical rating scale as an answer modality in ankylosing spondylitis specific questionnaires. AB - OBJECTIVES: To determine the agreement of scores on the original visual analog scale (VAS) or Likert scale of the Bath Ankylosing Spondylitis Disease Activity Index (BASDAI), Bath Ankylosing Spondylitis Functional Index (BASFI), and Dougados Functional Index (DFI) with scores on a numerical rating scale (NRS). To assess the reproducibility and responsiveness of the instruments with the original scale and NRS. METHODS: Five hundred thirty-six patients with ankylosing spondylitis from the Netherlands, Mexico, and Switzerland completed a questionnaire in which all questions from the BASDAI, BASFI, and DFI were presented twice in random order with an 11-point NRS and either a 10-cm VAS (BASDAI and BASFI) or a 5-point Likert scale (DFI). Agreement of scores using Bland-Altman plots and intraclass correlation coefficients (ICCs), reproducibility using ICCs, and responsiveness were assessed. RESULTS: Large variability between the scores on the original scales and the NRS was found in individual questions of all 3 questionnaires, although total scores showed ICCs of at least 0.88. Reproducibility of all answer modalities showed low ICCs in individual questions, but moderate to good ICCs in total scores (Dutch group 0.62 0.89; Mexican group 0.53-0.72). Moderate to large effects (0.48-1.04) were found in responsiveness scores in the 3 questionnaires. No major differences in reproducibility and responsiveness between the answer modalities were found. CONCLUSION: Although large variability between the scores on the original answer scales and the NRS was observed, the BASDAI, BASFI, and DFI can be administered with an NRS, which does not show important differences compared with the original scales. PMID- 12115153 TI - One-year open-label trial of thalidomide in ankylosing spondylitis. AB - OBJECTIVES: To test in a large open study whether thalidomide is potentially useful in treating ankylosing spondylitis, and to see if thalidomide induces any change in expression of genes in peripheral blood mononuclear cells (PBMC). METHODS: Thirty male patients with treatment-refractory ankylosing spondylitis were recruited into a 12-month open study using thalidomide at a dosage of 200 mg/day. Seven indices were measured as primary outcome measures, and 6 other indices as secondary outcome measures. Transcripts in the PBMC of some of these patients were first screened with microarray, and then measured with reverse transcriptase-polymerase chain reaction. RESULTS: Twenty-six patients completed the study. Of these, 80% showed a >20% improvement in 4 of 7 primary indices. Sharp declines in several parameters were noticed at 3-6 months. Nine patients became pain-free. There was also a statistically significant decrease in tumor necrosis factor alpha transcripts in the PBMC. CONCLUSION: Thalidomide is a reasonably promising drug in treatment-resistant ankylosing spondylitis. PMID- 12115154 TI - Statistical presentation and analysis of ordered categorical outcome data in rheumatology journals. AB - OBJECTIVE: To assess the appropriateness of presentation of summary measures and analysis of ordered categorical (ordinal) data in three rheumatology journals in 1999, and to consider differences between basic and clinical science articles. METHODS: Six hundred forty-four full-length articles from the 1999 editions of 3 rheumatology journals were evaluated for inclusion of an ordinal outcome. Articles were classified as basic or clinical science, and the appropriateness of presentation and analysis of the ordinal outcome were assessed. Chi-square tests were used to evaluate difference in percentages. RESULTS: Ordinal outcomes were identified in 175 (27.2%) of 644 articles. Only 69 (39.4%) had appropriate data presentation, and 111 (63.4%) had appropriate data analysis. Appropriate presentation was seen less commonly in the basic science rather than the clinical science articles, but differences in the occurrence of appropriate analysis were not seen. CONCLUSION: Ordinal data are common in rheumatology articles, but presentation usually does not conform to recommended guidelines. PMID- 12115155 TI - Does the label "fibromyalgia" alter health status, function, and health service utilization? A prospective, within-group comparison in a community cohort of adults with chronic widespread pain. AB - OBJECTIVE: To determine if assigning the label of fibromyalgia (FM) to individuals with chronic widespread pain has a significant effect on long-term health status, function, and health service utilization. METHODS: In the London Fibromyalgia Epidemiology Study, 100 individuals with FM were identified by screening 3,395 non-institutionalized adults. Only 28 of the 100 had been previously diagnosed with FM; for 72, the diagnostic label was new. All 28 with prediagnosed FM were female compared with 58 of the 72 newly diagnosed cases. In a prospective, within-group comparison, we compared previously non-labeled FM cases at study entry (prelabeling) and at 18 and 36 months followup (postlabeling) with respect to general health status, fibromyalgia-related symptoms, and all items from the Fibromyalgia Impact Questionnaire (FIQ) (including total FIQ score, and several measures of health service utilization) to see if health status, function, and health services utilization had changed, using paired t-tests. We also compared percentage reporting work disability at baseline and 18 months using Pearson's chi(2). RESULTS: Fifty-six (78%) of the original 72 newly diagnosed FM cases were available for reassessment at 18 months, and 43 (60%) at 36 months. Although physical functioning decreased slightly over time, there also was a statistically significant improvement in satisfaction with health, and newly diagnosed FM cases reported fewer symptoms and major symptoms over the long term. No other differences in clinical status or health service use occurred over time. CONCLUSION: The FM label does not have a meaningful adverse affect on clinical outcome over the long term. Further research is necessary to document the short- and long-term effect of labeling in the chronic pain patient. PMID- 12115156 TI - Factors of importance for work disability in women with fibromyalgia: an interview study. AB - OBJECTIVE: Fibromyalgia symptoms such as continuous pain, tiredness, hyperalgesia, and allodynia limit gainful employment. The present study examines which factors influence the decision to remain in a work role for women with fibromyalgia. This information is important for the individual and for public finances. METHODS: Qualitative interviews were performed with 39 women, 19 of whom were gainfully employed and 20 who had stopped working. The transcribed interviews were analyzed and divided into categories and subcategories. RESULTS: Four categories appear, at societal and individual levels, that were regarded as important by the women for remaining in a work role. CONCLUSIONS: The ability to remain at work depends not only on limitations in work capacity, but also on the capacity of society to adjust work environments and work tasks. More individual solutions are needed to allow women with fibromyalgia to maintain work roles. PMID- 12115157 TI - The genetic contribution to carpal tunnel syndrome in women: a twin study. AB - OBJECTIVE: To assess the relative genetic and environmental contribution to carpal tunnel syndrome (CTS) using a classic twin study of monozygotic (MZ) and dizygotic (DZ) twins. METHODS: The study group comprised unselected female twin pairs, between 20 and 80 years of age, from the St Thomas' UK Adult Twin Registry. Individuals completed a questionnaire that included details on potential risk factors for CTS. The diagnosis of CTS was made using a standardized hand pain diagram and validated criteria. The genetic contribution to CTS was assessed using variance component and regression methods, the heritability was adjusted for environmental confounders. The role of individual risk factors was assessed by a nested case-control study. RESULTS: An overall prevalence of 14.2% for CTS was found in a population of 4,488 females, comprising 867 MZ and 970 DZ twin pairs, and 814 singletons. The concordance for CTS was significantly higher in MZ compared with DZ twins (case-wise concordance values of 0.35 and 0.24 respectively, with a significantly increased MZ:DZ ratio of 1.48; P = 0.03). Modeling produced a heritability estimate of 0.46 (95% CI 0.34-0.58) that was essentially unchanged after adjustment for environmental risk factors including age, body mass index, physical activities, and hormonal/reproductive factors. No major influence of any individual risk factor was seen in the case-control analysis of 520 cases and 3,154 controls, apart from a modest association with menopausal status with an increased risk of 1.53 and 1.43 in the peri and postmenopausal groups. There was no overall effect of age or body mass index. CONCLUSION: This is the first study to explore the genetic component of CTS. Our data show that up to half of the liability to CTS in women is genetically determined, and this appears to be the single strongest risk factor, with only minor contributions from known environmental factors. Further studies should focus on genetic mechanisms that may lead to tests for susceptibility and detection of those at risk of developing CTS. PMID- 12115158 TI - A randomized controlled trial to evaluate the slow-acting symptom-modifying effects of colchicine in osteoarthritis of the knee: a preliminary report. AB - OBJECTIVE: To determine if colchicine added to nimesulide may have a beneficial effect on osteoarthritis (OA) of the knee. METHODS: Colchicine 0.5 mg twice daily or placebo was added to nimesulide (a nonsteroidal antiinflammatory drug) in 36 patients with OA of the knee in a randomized, double-blind, placebo-controlled trial over a 5-month period. RESULTS: The 30% improvement rate at 20 weeks was higher in the colchicine group than in the control group receiving placebo, as measured by total Western Ontario and McMaster University Osteoarthritis scores (57.9% versus 23.5%) and visual analog scale for index knee pain (52.6% versus 17.6%) (primary measures of response). The significance persisted on combined analysis by Mantel-Haenszel test (P = 0.062). Comparison of means also showed significant improvement in the colchicine group versus the control group in a multivariate analysis performed using T(2) test (P = 0.0115). CONCLUSION: Among patients with OA of the knee, the group receiving colchicine plus nimesulide exhibited significantly greater symptomatic benefit at 20 weeks than did the control group receiving nimesulide plus placebo. PMID- 12115159 TI - Physician treatment preferences in rheumatoid arthritis of differing disease severity and activity: the impact of cost on first-line therapy. AB - OBJECTIVE: To conduct a pilot study to identify rheumatologists' treatment preferences for first-line rheumatoid arthritis (RA) therapy and determine whether pharmacoeconomic variables modify physician choice(s). METHODS: A questionnaire describing 3 different RA scenarios was mailed to American College of Rheumatology members within 4 geographic regions of the US. Physicians were asked to identify their choice(s) of first-line therapy for each of the cases, first taking cost into consideration, second without considering the influence of cost, and third identifying the therapy that would be chosen for either themselves or a family member. RESULTS: Three hundred seventy-five questionnaires out of a total of 994 (37.7%) were returned between 3/12/00 and 4/25/00. Hydroxychloroquine was the most commonly cited medication for a mild disease activity/severity presentation, and methotrexate for a moderate-to-severe disease activity/severity presentation. For the severe disease activity/severity presentation, when cost was not considered, 217 (65%) rheumatologists included new disease-modifying antirheumatic drugs (leflunomide, etanercept, and infliximab) in their choice of first-line agents; this number decreased to 47 (14%) when cost was a consideration. CONCLUSION: Pharmacoeconomics appear to play a dominant role in rheumatologists' choice of treatment regimens, at times contrary to the physician's perception of the effectiveness of a drug. Future studies should address physician preferences in more depth with respect to cost and its various components. PMID- 12115160 TI - Psychological interventions for rheumatoid arthritis: a meta-analysis of randomized controlled trials. AB - OBJECTIVE: To carry out a systematic review of the literature examining the efficacy of psychological interventions (e.g., relaxation, biofeedback, cognitive behavioral therapy) in the treatment of rheumatoid arthritis (RA). METHODS: Studies that met the following criteria were included: random assignment, wait list or usual care control condition; publication in peer-reviewed journals; treatment that included some psychological component beyond simply providing education information; and separate data provided for patients with RA if subjects with conditions other than RA were included. Two investigators independently extracted data on study design, sample size and characteristics, type of intervention, type of control, direction and nature of the outcome(s). RESULTS: Twenty-five trials met the inclusion criteria. Methodologic quality was assessed, and effect sizes were calculated for 6 outcomes. Significant pooled effect sizes were found postintervention for pain (0.22), functional disability (0.27), psychological status (0.15), coping (0.46), and self efficacy (0.35). At followup (averaging 8.5 months), significant pooled effect sizes were observed for tender joints (0.33), psychological status (0.30), and coping (0.52). No clear or consistent patterns emerged when effect sizes for different types of treatment and control conditions were compared, or when higher quality trials were compared to lower quality ones. Findings do, however, suggest that these psychological interventions may be more effective for patients who have had the illness for shorter duration. CONCLUSIONS: Despite some methodologic flaws in the literature, psychological interventions may be important adjunctive therapies in the medical management of RA. PMID- 12115161 TI - Association of interleukin-4 and interleukin-1B gene variants with Larsen score progression in rheumatoid arthritis. AB - OBJECTIVE: To perform a genetic association study using markers in the interleukin-1 (IL-1) gene cluster and the IL-4/IL-4 receptor system genes, seeking evidence for involvement in the onset or the erosive outcome of rheumatoid arthritis (RA). METHODS: We tested the allelic distribution of IL-1A (+4845), IL-1B (-511), IL-1B (+3954), IL-1RN (+2018), IL-4 variable number of tandem repeat (VNTR), and IL-4R (+1902) in 233 patients with RA, 99 with polymyalgia rheumatica, and 148 ethnically matched controls. We analyzed the frequency of these gene variants in respect to presence of disease, but also to the degree of radiologic erosions (Larsen score) as a function of disease duration in 157 patients who had available radiographs of both hands. RESULTS: None of the 6 genetic polymorphisms was significantly different in frequency between RA patients and healthy controls or patients with polymyalgia rheumatica. Among RA patients, the rarer (#2) alleles of IL-4 VNTR and IL-1B (-511) were both associated with a milder Larsen score progression: The slope of Larsen progression in the rare allele groups diverged significantly from those of the frequent allele groups after approximately 20 years of disease duration (P < 0.001). CONCLUSION: None of the markers tested were shown to be associated with increased or decreased risk of RA. The rarer alleles of IL-4 VNTR and IL-1B ( 511) appear to be associated with a less severe course in RA of long duration. PMID- 12115162 TI - Systemic lupus erythematosus in three ethnic groups. X. Measuring cognitive impairment with the cognitive symptoms inventory. AB - OBJECTIVE: To determine the factor structure of the Cognitive Symptoms Inventory (CSI) in patients with systemic lupus erythematosus (SLE) participating in a multiethnic longitudinal study of outcome, the Lupus in Minority populations, Nature versus nurture (LUMINA) study. METHODS: LUMINA patients of Hispanic (n = 48), African American (n = 64), and Caucasian (n = 44) ethnicity who had a study visit (enrollment or followup) between January 1 and September 30, 2000 were included. Patients completed the CSI, a 21-item self-report measure of cognitive function. Sociodemographic, clinical, immunologic, psychosocial, and behavioral variables were ascertained per protocol and as previously described. Data were analyzed with SPSS. The factor structure of the CSI was determined using the principal axis method with oblique rotation as decided by Gorsuch. All factors having an Eigenvalue greater than 1 were considered. A 4-factor solution was derived that accounted for 42.6% of the common variance. The correlations between patient factor scale scores and variables from the demographic, clinical, psychosocial, and behavioral domains were then examined. RESULTS: The four factors and their respective variance are, Attention/Concentration (28.8%), Pattern Recognition/Activity Management (5.7%), Intermediate Memory (4.7%), and Initiation of Executive Functions (3.4%); each factor correlated with the total CSI score. Overall, patients' factor scale scores were positively and significantly correlated with other measures of cognitive dysfunction such as the Systemic Lupus Activity Measure (neuromotor domain) or the Systemic Lupus International Collaborating Clinics Damage Index (neurocognitive impairment), as well as with measures of fatigue, maladaptive coping skills, poor mental functioning, poor social support, and helplessness. They were, however, not correlated with sociodemographic or clinical variables. CONCLUSIONS: In addition to demonstrating that the CSI can be used to measure cognitive impairment in patients with SLE in the research setting, we have determined a 4-factor solution for the CSI that appears to have adequate metric properties. At present, the CSI may best be used as a screen for difficulties in daily activities involving intermediate memory, concentration, attention, and executive function. Nevertheless, further work with the CSI items and factor scales is necessary to establish internal and test-retest reliability of the factor scales; and provide additional evidence of the convergent and predictive validity of these scales in larger samples of patients from each ethnic subgroup. PMID- 12115163 TI - Effect of Wegener's granulomatosis on work disability, need for medical care, and quality of life in patients younger than 40 years at diagnosis. AB - OBJECTIVES: To evaluate the effects of Wegener's granulomatosis (WG) on employment status, work disability, and need for medical care of 60 consecutive WG patients aged < or = 40 years at diagnosis. METHODS: Sixty WG patients (26 male, 34 female) with a median age of 36 years (range 17-48 years) and a median duration of disease of 39 months (range 0-228 months) completed self-administered questionnaires on hospitalization, medical care, and employment status plus the Medical Outcomes Study-Short Form-36 (SF-36) estimating their health-related quality of life. RESULTS: Thirty-two of the 60 patients reported full- or part time employment more than 3 years after diagnosis. Only 14 of the 51 patients employed at diagnosis (27%) were currently receiving a permanent work disability pension due to WG. Two additional patients had lost work because of WG. Women who were employed at diagnosis had a nearly 3-fold higher risk of losing their jobs compared with men (P = 0.0006). There were no differences with regard to age at diagnosis, disease duration, disease severity, or education level between employed and unemployed patients. Employed patients had missed a median of 14 workdays (range 0-18 days) due to WG within the past 12 months. More than half of all patients (33 of 60) had been hospitalized during the previous 12 months because of WG. Ninety-three percent of all patients had visited their physician once or more per month, more than half of them at least once per week, regardless of employment status, severity of disease, or type of current medication. Unemployed WG patients experienced significant reductions in social and physical function and in their perceived degree of general health as assessed by the SF 36. CONCLUSIONS: Twenty-seven percent of WG patients younger than age 40 who were employed at diagnosis received permanent work disability within a disease duration of 39 months. Unemployment is followed by a considerable reduction in disease-related quality of life compared with employed patients, independent of severity and extent of disease. Furthermore, because patients were followed closely by an interdisciplinary team, a high rate of hospitalization and frequent visits to physicians resulted. PMID- 12115165 TI - The disablement process in rheumatoid arthritis. PMID- 12115164 TI - High rate of renal relapse in 71 patients with Wegener's granulomatosis under maintenance of remission with low-dose methotrexate. AB - OBJECTIVE: To examine the long-term efficacy of low-dose intravenous methotrexate (MTX) with and without concomitant glucocorticoids (GC) for remission maintenance in patients with generalized Wegener's granulomatosis (WG) in an open-label, prospective, standardized trial. METHODS: After induction of remission by cyclophosphamide and GC, 71 patients (41 males, 30 female) with initially generalized WG received low-dose methotrexate at 0.3 mg/kg body weight once weekly. At study-start 55 of 71 (77.5%) patients were on low-dose GC (mean 5.9 mg/day) which was tapered during the study. All patients underwent interdisciplinary staging at 3-month (and later at 6-month) intervals to assess disease activity and extent as well as side effects. End points were the first relapse or the end of study (January 2001). RESULTS: Within a mean followup period of 25.2 months, 26 patients (36.6%) experienced a relapse after a mean of 19.4 months. Seventeen (65.4%) of these 26 patients had terminated GC therapy at the time of relapse. There was no difference in relapse rates among patients with and without concomitant GC at study start. Relapses occurred mainly in the initially involved organ systems, preferentially in the ear, nose and throat tract in 18 of 26 patients and the kidney in 16 of 26 patients. One renal relapse presented as rapid, progressive glomerulonephritis with lethal outcome. Further, 14 relapses were accompanied by a significant rise in creatinine values. In 15/26 patients the relapse was paralleled or preceded by a significant rise of antineutrophil cytoplasmic antibody titer. Two patients ceased MTX prematurely because of persistent leukopenia. CONCLUSION: Weekly MTX is a well tolerated therapy for long-term maintenance of remission. However, one-third of the patients relapsed during ongoing MTX treatment, irrespective of whether they were still receiving GC. Because more than half of the relapses affected the kidney, close monitoring is indispensable. PMID- 12115166 TI - Pathogenesis of lupus. PMID- 12115167 TI - Increased frequency of malignancy found in patients presenting with new-onset polymyalgic symptoms suggested to have polymyalgia rheumatica. PMID- 12115168 TI - Nonmedication approaches in hand osteoarthritis. PMID- 12115169 TI - Comment on prescribing tumor necrosis factor inhibitors. PMID- 12115170 TI - CD154-CD40 interactions mediate differentiation to plasma cells in healthy individuals and persons with systemic lupus erythematosus. PMID- 12115171 TI - Epitope spreading in systemic lupus erythematosus: identification of triggering peptide sequences. PMID- 12115172 TI - Vaccination for Lyme disease: cost-effectiveness versus cost and value. PMID- 12115173 TI - Etanercept versus methotrexate in patients with early rheumatoid arthritis: two year radiographic and clinical outcomes. AB - OBJECTIVE: To compare the clinical and radiographic outcomes in patients with rheumatoid arthritis (RA) who received monotherapy with either etanercept or methotrexate (MTX) for 2 years and to assess the safety of this therapy. METHODS: In the Enbrel ERA (early rheumatoid arthritis) trial, 632 patients with early, active RA were randomized to receive either twice-weekly subcutaneous etanercept (10 mg or 25 mg) or weekly oral MTX (mean dosage 19 mg per week) for at least 1 year in a double-blind manner. Following the blinded phase of the trial, 512 patients continued to receive the therapy to which they had been randomized for up to 1 additional year, in an open-label manner. Radiograph readers remained blinded to treatment group assignment and the chronologic order of images. RESULTS: At 24 months, more 25-mg etanercept patients than MTX patients met American College of Rheumatology 20% improvement criteria (72% and 59%, respectively; P = 0.005), and more had no increase in total score and erosion scores on the Sharp scale (P = 0.017 and P = 0.012, respectively). The mean changes in total Sharp score and erosion score in the 25-mg etanercept group (1.3 and 0.66 units, respectively) were significantly lower than those in the MTX group (3.2 and 1.86 units, respectively; P = 0.001). Significantly more patients in the 25-mg etanercept group (55%) than in the MTX group (37%) had at least 0.5 units of improvement in the Health Assessment Questionnaire disability index (P < 0.001). Fewer patients in the etanercept group than in the MTX group experienced adverse events or discontinued treatment because of adverse events. CONCLUSION: Etanercept as monotherapy was safe and was superior to MTX in reducing disease activity, arresting structural damage, and decreasing disability over 2 years in patients with early, aggressive RA. PMID- 12115174 TI - The relationship of serum infliximab concentrations to clinical improvement in rheumatoid arthritis: results from ATTRACT, a multicenter, randomized, double blind, placebo-controlled trial. AB - OBJECTIVE: To investigate the relationship between serum concentrations of infliximab, a monoclonal anti-tumor necrosis factor alpha antibody, and clinical improvement from infliximab therapy for rheumatoid arthritis (RA). METHODS: Multiple blood samples were obtained from each of 428 subjects with active RA who were enrolled in a multicenter, randomized, double-blind, placebo-controlled trial (ATTRACT [Anti-Tumor Necrosis Factor Trial in Rheumatoid Arthritis with Concomitant Therapy]) evaluating the clinical efficacy and safety of infliximab therapy. Serum levels of infliximab were measured by enzyme-linked immunosorbent assay. Dose-response trends were analyzed using generalized logistic regression techniques. Pharmacokinetic modeling was used to predict the serum concentrations of infliximab after simulated infusions using doses and dosing intervals not evaluated in the trial. RESULTS: At week 54, 26% of the subjects receiving 3 mg/kg infliximab every 8 weeks had undetectable trough serum levels of infliximab, a significantly greater proportion than in the other 3 treatment groups (P < 0.001). Increased magnitude of American College of Rheumatology (ACR) response (measured by the ACR-N, a continuous measure of clinical improvement derived from the ACR 20% response criteria) and greater reduction from baseline in serum C-reactive protein level were both associated with higher trough serum concentrations of infliximab (P < 0.001), as was less progression of radiographic joint damage (P = 0.004), providing support for a dose-response relationship. Pharmacokinetic models predicted that decreasing the dosing interval from 8 weeks to 6 weeks would yield higher trough serum levels of infliximab than increasing the dose by 100 mg. CONCLUSION: These results suggest that some patients with RA may benefit from infliximab given at higher doses than 3 mg/kg or more frequently than every 8 weeks. PMID- 12115175 TI - Hydroxychloroquine concentration-response relationships in patients with rheumatoid arthritis. AB - OBJECTIVE: A dose-response relationship for hydroxychloroquine (HCQ), in terms of the proportion of patients achieving the Paulus 20% criteria for improvement, had previously been observed in patients with rheumatoid arthritis (RA) receiving a 6 week loading regimen of 400, 800, or 1,200 mg HCQ daily. This present retrospective analysis was performed to investigate possible relationships between the blood HCQ and HCQ-metabolite concentrations and measures of efficacy and toxicity. In addition, we sought to ascertain whether further investigation of HCQ/HCQ-metabolite levels might lead to testing of one of these substances as a new antirheumatic drug. METHODS: Patients with active RA (n = 212) began a 6 week, double-blind trial comparing 3 different doses of HCQ at 400, 800, or 1,200 mg/day, followed by 18 weeks of open-label HCQ treatment at 400 mg/day. Patients were repeatedly evaluated for treatment efficacy and toxicity. Blood samples were available from 123 patients for analysis of HCQ, desethylhydroxychloroquine (DHCQ), desethylchloroquine (DCQ), and bisdesethylchloroquine (BDCQ) levels using high-performance liquid chromatography. Achievement of the modified Paulus 20% improvement criteria for response in RA was used as the primary efficacy parameter. Spontaneously reported adverse events were categorized and analyzed as toxicity outcome variables. The relationship between response (efficacy and toxicity) and drug levels was evaluated using logistic regression analysis. RESULTS: The subset of patients with blood concentration data was equivalent to the larger study population in all demographic and outcome characteristics. The mean HCQ, DHCQ, and DCQ elimination half-lives were 123, 161, and 180 hours, respectively. There was a positive correlation between the Paulus 20% improvement criteria response and blood DHCQ concentrations during weeks 1-6 (P < 0.001). A potential relationship between ocular adverse events and BDCQ levels was found (P = 0.036). Logistic regression analysis of adverse events data showed that adverse gastrointestinal events were associated with higher HCQ levels (P = 0.001-0.021) during weeks 1, 2, and 3. CONCLUSION: There is a weak, but predictable, relationship between blood DHCQ concentrations and efficacy of treatment with HCQ. In addition, there is an association between gastrointestinal adverse events and elevated blood HCQ concentrations. Further investigation of these relationships is warranted to see if DHCQ may be introduced as a new antirheumatic drug. PMID- 12115176 TI - Costimulatory blockade in patients with rheumatoid arthritis: a pilot, dose finding, double-blind, placebo-controlled clinical trial evaluating CTLA-4Ig and LEA29Y eighty-five days after the first infusion. AB - OBJECTIVE: T cells are involved in the pathogenesis of rheumatoid arthritis (RA). In animal models of autoimmune diseases, blockade of costimulatory molecules on antigen-presenting cells has been demonstrated to be effective in preventing or treating this disease by preventing T cell activation. To date, the effect of costimulatory blockade in patients with RA is unknown. The goal of this multicenter, multinational study was to determine the safety and preliminary efficacy of costimulatory blockade using CTLA-4Ig and LEA29Y in RA patients who have been treated unsuccessfully with at least 1 disease-modifying agent. METHODS: CTLA-4Ig, LEA29Y (0.5, 2, or 10 mg/kg), or placebo was administered intravenously to 214 patients with RA. Patients received 4 infusions of study medication, on days 1, 15, 29, and 57, and were evaluated on day 85. The primary end point was the proportion of patients meeting the American College of Rheumatology 20% improvement criteria (ACR20). All patients were monitored for treatment safety and tolerability. RESULTS: CTLA-4Ig and LEA29Y infusions were well tolerated at all dose levels. Peri-infusional adverse events were carefully monitored, and showed similar incidence across all dose groups with the exception of headaches, which were slightly more frequent in the 2 treatment groups. The incidence of discontinuations due to worsening of RA was 19%, 12%, and 9% at 0.5, 2, and 10 mg/kg, respectively, in the CTLA-4Ig-treated patients and 3%, 3%, and 6% at 0.5, 2, and 10 mg/kg, respectively, in the LEA29Y-treated patients (versus 31% in the placebo group). ACR20 responses on day 85 had increased in a dose dependent manner (23%, 44%, and 53% of CTLA-4Ig-treated patients and 34%, 45%, and 61% of LEA29Y-treated patients at 0.5, 2.0, and 10 mg/kg, respectively, versus 31% of placebo-treated patients). CONCLUSION: Both of the costimulatory blocking molecules studied were generally safe and well tolerated. As compared with placebo, both CTLA-4Ig and LEA29Y demonstrated efficacy in the treatment of RA. PMID- 12115177 TI - Heterogeneity between men and women in the influence of the HLA-DRB1 shared epitope on the clinical expression of rheumatoid arthritis. AB - OBJECTIVE: To test the hypothesis that the influence of the HLA-DRB1 shared epitope (SE) on the clinical manifestations of rheumatoid arthritis (RA) differs between men and women. METHODS: We assessed 777 consecutive RA patients for age at disease onset, articular manifestations, subcutaneous nodules, laboratory and radiographic findings, and treatment received. We typed HLA-DRB1 alleles by polymerase chain reaction-sequence-specific primer amplification and categorized the number of SE-containing alleles. We used regression models to adjust comparisons between the sexes for age and clustering by recruitment center, and included SE x sex interaction terms to look for heterogeneity between men and women in the effect of the SE. RESULTS: Among the 777 RA patients, 548 (71%) were women. Men and women differed significantly in the adjusted frequency of SE positivity (women 71.4% versus men 78.4%; P < or = 0.001). The SE was associated with a younger age at symptom onset and RA diagnosis among men, but not among women. The SE likewise had a significant adverse effect on joint tenderness, swelling, and deformity among men only. The SE was associated with a higher erythrocyte sedimentation rate in women and more frequent positivity for rheumatoid factor among both men and women. CONCLUSION: There is heterogeneity between men and women in the effect of the SE on RA susceptibility and clinical expression. Further research is needed to understand the mechanism of this heterogeneity. PMID- 12115179 TI - Elevated levels of platelet microparticles are associated with disease activity in rheumatoid arthritis. AB - OBJECTIVE: Platelets are involved in various thrombotic events, often by means of platelet-derived microparticles (PMPs). It is likely that platelets are also involved in inflammation. Because inflammatory processes play a central role in rheumatoid arthritis (RA), we sought to determine whether PMPs are present in this disease. METHODS: This descriptive, cross-sectional study included 19 RA patients and 10 healthy controls. Nine of the patients had active RA (erythrocyte sedimentation rate [ESR] > or =28 mm/hour and/or C-reactive protein [CRP] level > or =28 mg/liter, > or =9 painful joints, and > or =6 swollen joints), and 10 had inactive disease (ESR < or =27 mm/hour, CRP < or =27 mg/liter, no tender joints, and no swollen joints). Platelet counts and PMP numbers were determined using cell counter and flow cytometry, respectively. RESULTS: Platelet counts in the 3 groups were similar. However, levels of PMPs in RA patients were significantly higher than those in healthy controls (median 616 versus 118 x 10(6)/liter; P = 0.005). PMP levels were higher in patients with active RA than in those with inactive RA (median 2,104 versus 504 x 10(6)/liter; P > 0.05). Moreover, PMP levels correlated with disease activity (r = 0.67, P = 0.05). CONCLUSION: PMPs are associated with RA, and PMP levels are correlated with disease activity. Thus, platelets probably play a part in the inflammatory process of RA by means of PMPs. Given the importance of PMPs in cardiovascular diseases, this may be one reason for the enhanced cardiovascular morbidity and mortality in RA. PMID- 12115178 TI - Increased thickness of the arterial intima-media detected by ultrasonography in patients with rheumatoid arthritis. AB - OBJECTIVE: To determine whether arterial wall thickening is advanced in rheumatoid arthritis (RA) patients compared with healthy controls by measuring the intima-media thickness (IMT) of the common carotid and femoral arteries, and to evaluate the factors associated with arterial IMT in patients with RA. METHODS: We studied 138 RA patients and 94 healthy controls (matched for age, sex, and other major risk factors for atherosclerosis). IMT was measured on digitized still images of the common carotid and femoral arteries obtained by high-resolution ultrasonography (10-MHz in-line Sectascanner). Laboratory variables relevant to RA activity were measured by routine methods. The degree of RA progression was assessed by scoring (Larsen method) metacarpophalangeal (MCP) joints on hand radiographs. Activities of daily living were determined by a modified Health Assessment Questionnaire (M-HAQ) score, and physical activity levels were assessed by ultrasound measurement of the calcaneus (expressed as the osteo-sono assessment index [OSI] Z score). RESULTS: Common carotid and femoral artery IMTs were significantly higher (P < 0.05) in RA patients (mean +/- SD 0.641 +/- 0.127 and 0.632 +/- 0.125 mm, respectively) compared with controls (0.576 +/- 0.115 and 0.593 +/- 0.141 mm, respectively). Multiple regression analysis revealed a significant association between RA and the common carotid artery IMT. Moreover, the common carotid artery IMT in RA patients was positively associated with disease duration, the MCP joint Larsen score, and the M-HAQ score, and was negatively associated with the calcaneus OSI Z score. No significant association was found between corticosteroid treatment and common carotid artery IMT. CONCLUSION: RA patients exhibited greater thickness of the common carotid and femoral arteries than healthy controls. The duration and severity of RA and decreased activities of daily living, but not corticosteroid treatment, were independently associated with the increased arterial wall thickness. PMID- 12115180 TI - Necessary role of phosphatidylinositol 3-kinase in transforming growth factor beta-mediated activation of Akt in normal and rheumatoid arthritis synovial fibroblasts. AB - OBJECTIVE: Rheumatoid arthritis is a disease that, pathologically, is characterized by the progressive growth and invasion of the synovial pannus into the surrounding cartilage and bone. Many cytokines, including transforming growth factor beta1 (TGFbeta1), have been implicated in this process, but their mode of action is incompletely understood. The goal of the present study was to better understand the downstream signaling pathways of TGFbeta in fibroblasts. METHODS: The role of phosphatidylinositol 3-kinase (PI 3-kinase) was determined by chemical inhibition with LY294002 or wortmannin. Activation of protein kinase B (Akt), c-Jun N-terminal kinases (JNKs), and extracellular signal-regulated kinases (ERKs) was evaluated by Western blot analysis using phospho-specific antibodies. RESULTS: Exposure of fibroblasts to TGFbeta rapidly induced activation of a kinase, Akt, that is known to inhibit apoptosis by a variety of pathways. Activation of Akt was blocked by the specific PI 3-kinase inhibitor, LY294002, indicating that TGFbeta-mediated phosphorylation of Akt was dependent on PI 3-kinase activation. This activation pathway was relatively selective for Akt, since inhibition of PI 3-kinase failed to substantially modify activation of ERKs or JNKs in synovial fibroblasts. Inhibition of the PI 3-kinase/Akt pathway resulted in impaired proliferation of synovial fibroblasts and partial attenuation of the protective effect of TGFbeta on Fas-mediated apoptosis. CONCLUSION: TGFbeta exerts its growth and antiapoptotic effects on fibroblasts, at least in part, by activation of the PI 3-kinase/Akt pathway. PMID- 12115181 TI - FLICE-inhibitory protein expression in synovial fibroblasts and at sites of cartilage and bone erosion in rheumatoid arthritis. AB - OBJECTIVE: Rheumatoid arthritis (RA) is a chronic inflammatory disease characterized by a hyperplastic synovial tissue, inflammatory infiltrates, and a progressive destruction of cartilage and bone. FLICE-inhibitory protein (FLIP) prevents the association of caspase 8 with FADD and thus exerts an antiapoptotic effect through inhibition of Fas-mediated apoptosis. We undertook this study to examine the expression of FLIP in RA, osteoarthritic (OA), and normal synovial tissues. METHODS: We investigated the expression of FLIP (long form) in 5 RA, 2 OA, and 2 normal synovial tissue samples. A 393-bp fragment was amplified from complementary DNA obtained from cultured RA synovial fibroblasts (RASF) by reverse transcription-polymerase chain reaction (RT-PCR). Using in situ hybridization, the expression of FLIP messenger RNA (mRNA) in paraffin-embedded synovial tissue sections was investigated semiquantitatively by analyzing the lining layer, the sublining, and sites of invasion. Immunohistochemistry with anti-CD68 antibodies was performed on serial tissue sections to further characterize the cell types expressing FLIP. In addition, quantitative expression of FLIP was measured by real-time PCR. RESULTS: RT-PCR revealed the expression of FLIP mRNA in all RA and OA samples tested. Using in situ hybridization in synovial tissue, FLIP was detected in all 5 RA samples and in 1 of 2 OA samples, but in neither of the 2 normal control samples. In RA, FLIP expression could be found in both the lining and sublining layers; most importantly, it could also be identified at sites of cartilage invasion and bone destruction. Moreover, quantitative PCR analysis showed 50% higher FLIP expression in RASF than in OASF. CONCLUSION: The expression of antiapoptotic FLIP in RA synovial tissue and in synovial fibroblasts suggests the idea of a novel pathway in RA that potentially extends the lifespan of cartilage- and bone-degrading synovial cells, thus contributing to the progression of joint destruction. PMID- 12115182 TI - Association of the interleukin-1 gene cluster on chromosome 2q13 with knee osteoarthritis. AB - OBJECTIVE: To investigate whether the interleukin-1 (IL-1) ligand gene cluster at 2q13 encodes for genetic susceptibility to primary osteoarthritis (OA). METHODS: Seven single-nucleotide polymorphisms (SNPs) and a variable-number tandem repeat (VNTR) polymorphism from within the IL-1 ligand genes IL1A, IL1B, and IL1RN were genotyped in a cohort of 557 OA cases and 557 age-matched controls. RESULTS: None of the variants demonstrated association in the unstratified data set. However, when cases were stratified according to sex and site of disease (hip or knee), 4 SNPs showed marginal evidence for association (P < 0.1) in knee cases (n = 136) and male knee cases (n = 58). For 2 of these SNPs, evidence for association was enhanced when probands from 60 knee-only affected sibling pair families were genotyped and combined with the original knee cases (P < or = 0.05). Further analysis revealed that the associated alleles at 2 of these SNPs were markers for the same haplotype, the frequency of which was significantly elevated when knee cases and knee probands were combined (P = 0.01, odds ratio [OR] 1.4) and when male knee cases and male knee probands were combined (P = 0.009, OR 1.7). Furthermore, linkage analysis of 2q revealed suggestive evidence for linkage to the IL-1 gene clusters in affected sibling pairs concordant for knee OA but no evidence for linkage in affected sibling pairs concordant for hip OA. CONCLUSION: The IL-1 ligand cluster encodes for susceptibility to knee OA but not to hip OA, highlighting the genetic heterogeneity of this common, complex disease. PMID- 12115183 TI - The alpha5beta1 integrin provides matrix survival signals for normal and osteoarthritic human articular chondrocytes in vitro. AB - OBJECTIVE: Chondrocyte cell death may play an important role in the development of arthritis. The goal of the present study was to evaluate the role of the extracellular matrix (ECM) in promoting chondrocyte survival via signals through the integrin family of ECM receptors. METHODS: Chondrocytes were isolated by sequential enzymatic digestion from normal ankle cartilage of organ donors and from osteoarthritic (OA) knee tissue obtained from patients undergoing total knee replacement. Cell survival in monolayer and in suspension culture was measured using fluorescent labels after treatment with specific integrin-blocking antibodies and echistatin, a disintegrin peptide. A quantitative enzyme-linked immunosorbent assay for histone-associated DNA fragments and morphologic evaluation by electron microscopy were used to evaluate apoptosis. RESULTS: Freshly isolated chondrocytes died when plated in serum-free media at low density on poly-L-lysine, but showed >95% survival on fibronectin (FN). A monoclonal blocking antibody to the alpha5-integrin subunit (FN receptor) significantly inhibited survival on FN, whereas control antibodies had no effect. Likewise, treatment of freshly isolated chondrocytes in serum-free alginate-suspension culture with the alpha5-blocking antibody resulted in cell death in a dose dependent manner, with 20 microg/ml of the antibody reducing normal chondrocyte survival to 20% of that in controls, and OA chondrocyte survival to 23% of that in controls. Antibody inhibition of alphav and alpha1 integrins or treatment with echistatin did not cause cell death. Addition of insulin-like growth factor 1 (IGF-1; 100 ng/ ml) was not able to improve survival of alpha5-antibody-treated cells. However, treatment with 10% fetal bovine serum improved normal chondrocyte survival to 98% (a 5.1-fold increase) and OA chondrocyte survival to 64% (a 2.8 fold increase). Cell death due to alpha5 inhibition was associated with apoptosis. CONCLUSION: These results demonstrate that chondrocyte survival signals are transmitted via the alpha5beta1 FN receptor. Inhibition of matrix survival signals mediated by alpha5beta1 also inhibits the ability of IGF-1 to promote survival, suggesting that IGF-1-mediated survival signaling may require a cosignal from alpha5beta1. PMID- 12115184 TI - Regional quantification of cartilage type II collagen and aggrecan messenger RNA in joints with early experimental osteoarthritis. AB - OBJECTIVE: Accurate assessment of chondrocyte metabolism is a potentially valuable indicator of cartilage health in osteoarthritis (OA). This study was conducted to explore the anabolic metabolism of chondrocytes early in the natural history of an experimental canine model of OA. METHODS: Competitive reverse transcription-polymerase chain reaction was used to calculate the copy number of type II collagen and the messenger RNA (mRNA) levels of aggrecan core protein in articular cartilage samples obtained from different regions of the femorotibial joint 12 and 39 weeks after cruciate transection. RESULTS: Gene expression of both type II collagen and aggrecan in cartilage samples obtained from experimental joints at both intervals after surgery was elevated significantly compared with that in samples from contralateral control joints. The number of mRNA copies per microgram of DNA of aggrecan exceeded that of type II collagen in control cartilage, but the copy number of type II collagen mRNA exceeded that of aggrecan in OA cartilage. Thus, the ratio of type II collagen-to-aggrecan mRNA copy number (normalized to DNA) was shown to be characteristically altered in cartilage with experimental OA. CONCLUSION: Accurate assessment of multiple gene products in small samples of cartilage taken from focal areas of a joint can be used diagnostically for monitoring chondrocyte metabolism and possibly for staging at least the early phases of this joint disease. PMID- 12115185 TI - Pathologic indicators of degradation and inflammation in human osteoarthritic cartilage are abrogated by exposure to n-3 fatty acids. AB - OBJECTIVE: To determine if n-3 polyunsaturated fatty acid (PUFA) supplementation (versus treatment with n-6 polyunsaturated or other fatty acid supplements) affects the metabolism of osteoarthritic (OA) cartilage. METHODS: The metabolic profile of human OA cartilage was determined at the time of harvest and after 24 hour exposure to n-3 PUFAs or other classes of fatty acids, followed by explant culture for 4 days in the presence or absence of interleukin-1 (IL-1). Parameters measured were glycosaminoglycan release, aggrecanase and matrix metalloproteinase (MMP) activity, and the levels of expression of messenger RNA (mRNA) for mediators of inflammation, aggrecanases, MMPs, and their natural tissue inhibitors (tissue inhibitors of metalloproteinases [TIMPs]). RESULTS: Supplementation with n-3 PUFA (but not other fatty acids) reduced, in a dose dependent manner, the endogenous and IL-1-induced release of proteoglycan metabolites from articular cartilage explants and specifically abolished endogenous aggrecanase and collagenase proteolytic activity. Similarly, expression of mRNA for ADAMTS-4, MMP-13, and MMP-3 (but not TIMP-1, -2, or -3) was also specifically abolished with n-3 PUFA supplementation. In addition, n-3 PUFA supplementation abolished the expression of mRNA for mediators of inflammation (cyclooxygenase 2, 5-lipoxygenase, 5-lipoxygenase-activating protein, tumor necrosis factor alpha, IL-1alpha, and IL-1beta) without affecting the expression of message for several other proteins involved in normal tissue homeostasis. CONCLUSION: These studies show that the pathologic indicators manifested in human OA cartilage can be significantly altered by exposure of the cartilage to n-3 PUFA, but not to other classes of fatty acids. PMID- 12115187 TI - Role of the Fcgamma receptor IIa polymorphism in susceptibility to systemic lupus erythematosus and lupus nephritis: a meta-analysis. AB - OBJECTIVE: To assess the impact of the Fcgamma receptor type IIa (FcgammaRIIa) R/H131 polymorphism on the risk for systemic lupus erythematosus (SLE) and development of lupus nephritis. METHODS: A meta-analysis was performed based on the Medline and Embase databases (last retrieval August 2001), assessment of bibliographies of pertinent articles, and additional data gathered after contact with primary investigators. RESULTS: A total of 25 comparisons from 17 studies involving R/H131 genotyping of 1,405 patients with lupus nephritis, 1,709 SLE patients without nephritis, and 2,580 non-SLE controls were included. No association between RR genotype and risk of lupus nephritis relative to both other genotypes (odds ratio [OR] 1.05, 95% confidence interval [95% CI] 0.88 1.27) was demonstrated in the total meta-analysis or in any racial subgroup. The RR genotype was more frequent in SLE patients as a whole (OR 1.30, 95% CI 1.10 1.52) and in SLE patients without nephritis (OR 1.27, 95% CI 1.04-1.55) compared with disease-free controls. A potential dose-response relation between the R131 allele and the risk of SLE was also identified, with an OR of 1.23 for RR versus RH (95% CI 1.03-1.46). The OR was 1.55 for RR versus HH (95% CI 1.21-1.98). There was no significant heterogeneity between racial subgroups. The population attributable fractions of SLE cases due to the FcgammaRIIa-R131 allele were 13%, 40%, and 24% in subjects of European, African, and Asian descent, respectively. CONCLUSION: The FcgammaRIIa-R/H131 polymorphism represents a significant risk factor for SLE but has no clear effect on susceptibility for lupus nephritis. PMID- 12115186 TI - The effect of anti-CD40 ligand antibody on B cells in human systemic lupus erythematosus. AB - OBJECTIVE: To determine the immunologic effects of anti-CD40 ligand (anti-CD40L) therapy in 5 patients with systemic lupus erythematosus nephritis who participated in an open-label study of a humanized anti-CD40L monoclonal antibody. METHODS: Serum and peripheral blood mononuclear cells were obtained before, during, and after treatment, and the frequency of Ig and anti-DNA antibody-secreting B cells was analyzed by enzyme-linked immunospot assay and by analysis of Epstein-Barr virus (EBV)-transformed B cell lines. To determine the effect of treatment on somatic mutation of Ig genes, reverse transcriptase polymerase chain reaction was performed on messenger RNA from 4 patients, using primers specific for the DP-47 heavy chain gene and for IgG. Finally, B cell phenotype was investigated using flow cytometry. RESULTS: Even a brief period of treatment with anti-CD40L markedly reduced the frequency of IgG and IgG anti-DNA antibody-producing B cells, and these changes persisted for several months after cessation of treatment. To confirm these findings, EBV-transformed B cell lines were screened from each of 3 patients, and a 10-fold decrease in anti-DNA antibody-secreting cell lines was found after treatment in all 3 patients. Few differences in mutation patterns were observed before and after treatment; however, the frequency of germline-encoded DP-47 sequences was significantly increased before treatment and normalized following treatment. Flow cytometric analysis of B cells revealed expansion of a CD27-/IgD- B cell subset in some of the patients, which did not change with treatment. CONCLUSION: These are the first mechanistic studies of the effect of anti-CD40L therapy in human autoimmune disease. The results suggest that further studies of CD40L blockade are warranted. PMID- 12115188 TI - Anti-La/SSB antibodies transported across the placenta bind apoptotic cells in fetal organs targeted in neonatal lupus. AB - OBJECTIVE: To determine whether La and/or Ro epitopes on apoptotic cells in fetal organs that are targeted in neonatal lupus syndrome (NLS) are accessible for binding by autoantibodies in vivo, we traced the fate of transplacental autoantibodies in a murine passive transfer model. METHODS: Pregnant mice at day 15 of gestation (E15) were injected intraperitoneally with human anti-Ro/La positive sera or control sera, and transplacental transfer of human autoantibodies was tested by enzyme-linked immunosorbent assay with recombinant antigens. Multiple cryostat sections at the level of the heart of E17 fetuses were visualized simultaneously for human IgG binding and apoptosis (TUNEL) under confocal microscopy. Serial paraffin sections of E17 and E19 fetuses were examined for histologic evidence of inflammation. RESULTS: Human IgG anti-52-kd Ro, anti-60-kd Ro, and anti-La autoantibodies were transported efficiently into the fetal circulation. Human IgG-apoptotic cell complexes were detected in the heart (atrial trabeculae and atrioventricular node), skin, liver, and newly forming bone of fetuses from mothers injected with anti-Ro/La sera but not control sera. The IgG binding was fetal-specific and organ-specific; transplacental autoantibodies did not bind to apoptotic cells in the fetal thymus, lung, brain, or gut. The complexes were not associated with an inflammatory reaction. Injection of mothers with affinity-purified anti-La autoantibodies (but not anti-Ro/La Ig depleted of anti-La) revealed an identical location of IgG binding to apoptotic cells in the fetuses. CONCLUSION: This is the first study to demonstrate that transplacental anti-La autoantibodies bind specifically to apoptotic cells in selected fetal organs in vivo, similar to the organ involvement in NLS. We hypothesize that additional factors are required to promote proinflammatory clearance of IgG-apoptotic cell complexes and subsequent tissue damage. PMID- 12115189 TI - Autologous blood stem cell transplantation in refractory systemic lupus erythematosus with severe pulmonary impairment: a case report. AB - For patients with severe forms of autoimmunity, including systemic lupus erythematosus (SLE), purging autoreactive T cells from the immune repertoire by transplanting autologous hematopoietic stem cells (ASCT) is a therapeutic option. We describe an 18-year-old woman with SLE who had been treated with corticosteroids, azathioprine, cyclophosphamide (CYC), and immunopheresis for 4 years, during which time mechanical ventilation for lupus pneumonitis had been repeatedly required. After the patient was conditioned by administration of CYC and antithymocyte globulin, a total of 8.87 x 10(6) purified CD34+ cells per kg of body weight was infused. Hematopoietic regeneration was observed within 9 days. Twenty-one months after ASCT, the patient continues to be in complete clinical remission, with no signs of SLE-related disease activity and without any immunosuppressive medications. Her pulmonary function has returned to normal. Although a longer followup is required for assessment of the durability of response, the patient's course indicates that ASCT may be a way to reinduce tolerance in patients with SLE. PMID- 12115190 TI - Suppression of tumor necrosis factor alpha-induced matrix metalloproteinase 9 production in human salivary gland acinar cells by cepharanthine occurs via down regulation of nuclear factor kappaB: a possible therapeutic agent for preventing the destruction of the acinar structure in the salivary glands of Sjogren's syndrome patients. AB - OBJECTIVE: Our previous results suggested that suppression of tumor necrosis factor alpha (TNFalpha)-induced matrix metalloproteinase 9 (MMP-9) could prevent the destruction of acinar tissue in the salivary glands of patients with Sjogren's syndrome (SS). The present study was undertaken to investigate the effect of cepharanthine on the suppression of TNFalpha-induced MMP-9 production in NS-SV-AC, an SV40-immortalized normal human acinar cell clone. METHODS: After pretreatment with or without cepharanthine, NS-SV-AC cells were treated with TNFalpha alone or with a combination of TNFalpha and cepharanthine. The expression of MMP-9 was then examined at the protein and messenger RNA levels. In addition, the effect of cepharanthine on the morphogenetic behavior of NS-SV-AC cells cultured on type IV collagen-coated dishes in the presence of TNFalpha was examined. RESULTS: Although TNFalpha induced the production of MMP-9 in NS-SV-AC cells, this production was greatly suppressed when cells were pretreated with cepharanthine, followed by treatment with both TNFalpha and cepharanthine. In addition, cepharanthine suppressed the TNFalpha-stimulated NF-kappaB activity by partly preventing the degradation of IkappaBalpha protein in NS-SV-AC cells. When NS-SV-AC cells were seeded on type IV collagen-coated dishes in the presence of both TNFalpha and plasmin, type IV collagen interaction with the cells was lost and the cells entered apoptosis. However, pretreatment with cepharanthine restored the aberrant in vitro morphogenesis of the NS-SV-AC cells. CONCLUSION: These results may indicate a molecular mechanism by which cepharanthine is able to protect against the destruction of the acinar structure in salivary glands from patients with SS. PMID- 12115191 TI - Antifibroblast antibodies from systemic sclerosis patients are internalized by fibroblasts via a caveolin-linked pathway. AB - OBJECTIVE: Fibroblast activation is a crucial event in the development of systemic sclerosis (SSc). Antifibroblast autoantibodies (AFAs), detectable in the sera of SSc patients, are able to induce a proinflammatory phenotype on cultured fibroblasts. This study was undertaken to investigate the mechanisms of the interaction between AFAs and living fibroblasts. METHODS: We coupled to fluorescein 1) IgG purified from AFA-positive and AFA-negative SSc sera (as assessed by cellular enzyme-linked immunosorbent assay) and 2) single healthy donor and pooled normal IgG. The interaction of IgG with living cultured fibroblasts from healthy individuals and from a patient with SSc was visualized by real-time confocal microscopy. Intracellular colocalization of caveolin and internalized AFA-positive IgG was assessed by immunofluorescence. RESULTS: AFA positive IgG bound to living fibroblasts and was internalized with a cytoplasmic fibrillar pattern, in contrast to AFA-negative IgG. In the IgG tested, no correlation with antinuclear antibody activity was found. Preincubation of fibroblasts with normal IgG did not affect internalization. Internalized AFA positive IgG colocalized with caveolin, and internalization was entirely inhibited by disassembling fibroblast caveolae with filipin. CONCLUSION: The finding that both normal and pathologic fibroblasts specifically internalized AFA positive, but not AFA-negative, IgG demonstrates that AFAs in SSc patient sera interact with constitutively expressed membrane molecules on fibroblasts, via an Fc-independent mechanism. The results of colocalization and inhibition experiments suggest that microdomains containing caveolin are involved in the interaction between AFAs and fibroblasts. These data, together with the reported ability of AFAs to activate fibroblasts, provide evidence for a role of AFAs in the pathogenesis of SSc. PMID- 12115192 TI - Autoantibodies to fibroblasts induce a proadhesive and proinflammatory fibroblast phenotype in patients with systemic sclerosis. AB - OBJECTIVE: Fibroblasts play a major role in the development of systemic sclerosis (SSc), and the occurrence of serum autoantibodies reacting with fibroblast plasma membrane antigens in SSc has been reported. This study was undertaken to investigate whether IgG from SSc sera that react with human fibroblasts modulates the fibroblasts' function. METHODS: Sera from 69 patients with SSc (28 with limited cutaneous SSc [lcSSc] and 41 with diffuse cutaneous SSc [dcSSc]), 30 patients with sarcoidosis, and 50 matched healthy controls were examined. We evaluated antibody binding to human skin and lung fibroblasts by cell-based enzyme-linked immunosorbent assay (ELISA), indirect immunofluorescence, and flow cytometry. We further investigated the ability of purified IgG to modulate 1) intercellular adhesion molecule 1 (ICAM-1) expression, 2) U937 cell adhesion to fibroblasts, and 3) fibroblast steady-state messenger RNA (mRNA) levels of interleukin-1alpha (IL-1alpha), IL-beta, and IL-6, and IL-6 protein production. RESULTS: Of 69 SSc sera tested by cell-based ELISA, 58% bound to normal skin and lung fibroblasts. The prevalence of binding was significantly higher in dcSSc than in lcSSc (P < 0.05). Only IgG from SSc sera that were positive for antifibroblast antibody (AFA) induced a dose-dependent up-regulation of ICAM-1 expression and IL-6 production, enhancement of U937 cell adhesion, and increased levels of IL-1alpha, IL-1beta, and IL-6 mRNA in fibroblasts. Up-regulation of ICAM-1 mediated by AFA IgG was inhibited by the addition of IL-1 receptor antagonist, indicating an autocrine activation loop. CONCLUSION: Our findings confirm the presence of AFAs in SSc sera and demonstrate, for the first time, that autoantibodies reacting with fibroblast surface molecules act as an extrinsic stimulus inducing fibroblast activation in vitro. PMID- 12115193 TI - A gene in the telomeric HLA complex distinct from HLA-A is involved in predisposition to juvenile idiopathic arthritis. AB - OBJECTIVE: Juvenile idiopathic arthritis (JIA) is associated with particular alleles at 3 different HLA loci: HLA-A, HLA-DR/DQ, and HLA-DP. These associations are independent of each other (i.e., they cannot be explained by the known linkage disequilibrium between alleles at these loci). The purpose of this study was to look for additional JIA susceptibility genes in the HLA complex. METHODS: One hundred two Norwegian JIA patients and 270 healthy individuals, all carrying the DQ4;DR8 haplotype, were investigated by scanning approximately 10 megabases of DNA covering the HLA complex with microsatellite polymorphisms. An expectation maximization algorithm was used to estimate haplotype frequencies, and the distribution of microsatellite alleles on the high-risk DQ4;DR8 haplotype was compared between patients and controls, to exclude effects secondary to linkage disequilibrium with these susceptibility genes. RESULTS: Allele 5 at the microsatellite locus D6S265 (D6S265* 5), 100 kb centromeric of HLA-A, showed a strong positive association with the disease (odds ratio 4.7, corrected P < 10( 6)). Haplotype analysis demonstrated that the D6S265*5 association was not caused by linkage disequilibrium to the gene encoding HLA-A*02, which has previously been reported to be associated with JIA. Instead, our data suggested that a gene in linkage disequilibrium with D6S265*5, but distinct from HLA-A*02, is involved in the predisposition to JIA. CONCLUSION: We found that D6S265*5 could be a marker for an additional susceptibility gene in JIA which is distinct from A*02, adding to the risk conferred by DQ4;DR8. PMID- 12115194 TI - The efficacy and safety of intraarticular corticosteroid therapy for coxitis in juvenile rheumatoid arthritis. AB - OBJECTIVE: To study the efficacy and safety of intraarticular triamcinolone hexacetonide (IATH) for the treatment of coxitis in patients with juvenile rheumatoid arthritis (JRA). METHODS: Fifty consecutive patients with JRA and coxitis were studied prospectively. Forty-eight children received IATH in 67 arthritic hips. The remaining 2 children exhibited 3 cases of femoral head necrosis (FHN) at the initial assessment and were only followed up; both were receiving long-term systemic steroids. After a minimum of 2 years, the study was concluded with a final evaluation that included magnetic resonance imaging. RESULTS: In 39 of 67 hip joints (58%), remission of the coxitis for a period of 2 years was obtained through a single administration of IATH, while another 12 hip joints showed remission of coxitis after repeated TH injections (total remission rate 76%). We observed 2 patients with FHN following IATH. Both of these children were receiving long-term systemic steroids. During the period between onset of JRA and screening assessment for this study, the children exhibited 2.4 cases of FHN per 100 patient-years, while 1.5 cases of FHN per 100 patient-years were observed between IATH treatment and final followup. All 5 observed cases of FHN occurred among the 20 children who received long-term systemic steroids, while no necrosis occurred in the 30 children who did not receive systemic corticosteroids (P = 0.009 by Fisher's exact test). CONCLUSION: IATH for juvenile rheumatoid coxitis was an effective treatment that did not increase the rate of FHN. Systemic steroids, however (or their covariable, severity of JRA), may increase the risk of FHN in JRA. PMID- 12115196 TI - Takayasu arteritis: utility and limitations of magnetic resonance imaging in diagnosis and treatment. AB - OBJECTIVE: Previous studies have confirmed the poor correlation of symptoms, signs, and levels of acute-phase reactants with disease activity in approximately 50% of all patients with Takayasu arteritis (TA). Invasive angiographic studies demonstrate vessel lumen anatomy, but do not provide qualitative information about the vessel wall. Moreover, sequential invasive angiographic studies expose patients to high-dose ionizing radiation and catheter/procedure-related morbidity. The aim of the present study was to determine the utility of new developments in vascular magnetic resonance (MR) technology in patients with TA. METHODS: Electrocardiogram-gated "edema-weighted" MR was used to evaluate the aorta and its primary branches with regard to the vascular lumen, vessel wall anatomy, and vessel wall edema in 24 TA patients (77 studies). Inclusion criteria were age <50 years and features of TA on both clinical examination and invasive angiographic studies. Patients were stratified based on clinical and laboratory indications of having either unequivocally active disease, inactive disease, or uncertain disease status. RESULTS: MR revealed vessel wall edema in 94% (17 of 18), 81% (13 of 16), and 56% (24 of 43) of studies obtained during periods of unequivocally active disease, uncertain disease activity, and apparent clinical remission, respectively. Westergren erythrocyte sedimentation rate and C-reactive protein values did not correlate with either the clinical assessment of disease activity or MR evidence of vascular edema. The frequency of presumed vascular inflammation (edema), as assessed by MR, in patients who appeared to be in remission was similar to the reported frequency of new angiographic lesions and histopathologic evidence of active disease in surgical specimens from patients thought to be in remission. However, the presence of edema within vessel walls did not consistently correlate with the occurrence of new anatomic changes found on subsequent studies. CONCLUSION: Inconsistencies in the presence or absence of vessel edema and subsequent anatomic changes have cast doubt on the utility of edema-weighted MR imaging as a sole guide to disease activity and treatment in TA. In this study, the greatest utility of MR was in providing a safe, noninvasive means of assessing changes in vascular anatomy. PMID- 12115195 TI - Role of NOD2 variants in spondylarthritis. AB - OBJECTIVE: To investigate the role of the gene NOD2 in susceptibility to, and clinical manifestations of, ankylosing spondylitis (AS). METHODS: A case-control study of NOD2 polymorphisms known to be associated with Crohn's disease (CD) (Pro(268)Ser, Arg(702)Trp, Gly(908)Arg, and Leu(1007)fsinsC) was performed in 229 cases of primary AS with no diagnosed inflammatory bowel disease (IBD), 197 cases of AS associated with IBD (referred to as colitic spondylarthritis; comprising 78 with CD and 119 with ulcerative colitis [UC]), and 229 ethnically matched, healthy controls. Associations between NOD2 polymorphisms and several clinical features of AS, including disease severity assessed by questionnaire and age at spondylarthritis onset, were also investigated. Exclusion linkage mapping of chromosome 16 was performed in a separate group of 185 multicase families with AS. RESULTS: An association was identified between Gly(908)Arg and UC spondylarthritis (P = 0.016, odds ratio [OR] 4.6, 95% confidence interval [95% CI] 1.3-16), and a nonsignificant trend with a similar magnitude was observed in association with CD spondylarthritis (P = 0.08, OR 3.9, 95% CI 0.8-18). The Pro(268)Ser variant was inversely associated with UC spondylarthritis (P = 0.003, OR 0.55, 95% CI 0.37-0.82), but not with CD spondylarthritis. No association was demonstrated between NOD2 variants and primary AS, or between other variants of NOD2 and either UC or CD spondylarthritis. Carriage of the Pro(268)Ser polymorphism was associated with greater disease activity as measured by the Bath Ankylosing Spondylitis Disease Activity Index (P = 0.002). Although patients with CD had a younger age at spondylarthritis onset than did those with UC (22.4 years versus 26.4 years; P = 0.01), no association was noted between the NOD2 variants linked with CD and age at spondylarthritis onset. In primary AS, the presence of a gene with a magnitude of association >2.0 was excluded (exclusion logarithm of odds score less than -2.0), and no association was observed with the microsatellite D16S3136. CONCLUSION: NOD2 variants do not significantly affect the risk of developing primary AS, but may influence susceptibility to, and clinical manifestations of, colitic spondylarthritis. PMID- 12115197 TI - Monosodium urate monohydrate crystal-induced inflammation in vivo: quantitative histomorphometric analysis of cellular events. AB - OBJECTIVE: To quantify the inflammatory cell response in rat air pouch pseudosynovial membrane during monosodium urate monohydrate (MSU) crystal-induced inflammation. METHODS: In the rat air-pouch model, we used a computer-assisted histomorphometric method to quantify cell distributions, based on cell linear densities, in histologic sections of membranes from pouches injected with MSU or saline. The volume, white blood cell (WBC) count, and histamine content of the pouch exudates were determined at several time points. RESULTS: Injection of 10 mg of MSU crystals into the pouch produced an acute exudate. After peaking at 24 hours, the exudate volume and WBC count decreased spontaneously over the next 3 days, simulating the self-limited course of acute gout. Membrane thickness followed a parallel course. Membrane polymorphonuclear cell (PMN) linear densities were closely correlated with exudate WBC counts, suggesting PMN recruitment from the subintimal synovial membrane. Both monocyte/macrophage and mast cell linear densities increased in the subintimal layer 2 hours after crystal injection (P = 0.038 and P = 0.03, respectively, versus controls), whereas PMN linear densities showed 2 peaks, one at 4 hours and the other 24 hours. The exudate histamine content peaked 6 hours after crystal injection, when mast cell linear densities were minimal in the membranes, suggesting mast cell degranulation. CONCLUSION: An increase in monocyte/macrophage and mast cell densities in the membrane preceded the PMN influx in the pouch membrane and exudate, suggesting that mast cells may be involved in the early phase of MSU crystal-induced inflammation, at least in this rat model. PMID- 12115198 TI - Cost-effectiveness analysis of the Lyme disease vaccine. AB - OBJECTIVE: A vaccine for Lyme disease was approved in 1998 for use in the US. Given the high cost of the vaccine, the low risk of Lyme disease in many areas, and the largely curable nature of the disease, the cost-effectiveness of the vaccine in various risk groups is uncertain. This study was undertaken to examine the cost-effectiveness of the Lyme disease vaccine and the factors that influence its cost-effectiveness. METHODS: We constructed a Markov decision-analysis model to evaluate the clinical effectiveness and cost-effectiveness of the Lyme disease vaccine in populations at various levels of risk for the disease. The probabilities of clinical events and costs were estimated from reports in the literature. Sensitivity analyses assessed the impact of potential variations of parameters on model results. RESULTS: At the average national incidence of Lyme disease (0.0067%), the incremental cost-effectiveness of vaccination was approximately $1,600,000 per case averted when a yearly booster was given for 10 years after the standard initial vaccination regimen of 3 inoculations at 0, 1, and 12 months. For populations with an annual Lyme disease incidence of 1% (the incidence in several well-defined geographical areas of the US), the incremental cost-effectiveness was approximately $9,900 per case averted. Disease incidence had to exceed 10% before vaccination with yearly boosters became both more effective and more cost saving than no vaccination. CONCLUSION: The Lyme disease vaccine is cost-effective only for individuals who live in areas where Lyme disease is endemic and who are frequently exposed to ticks. PMID- 12115200 TI - Escherichia coli heat-labile enterotoxin B subunit prevents autoimmune arthritis through induction of regulatory CD4+ T cells. AB - OBJECTIVE: The receptor-binding B subunit of Escherichia coli heat-labile enterotoxin (EtxB) is a highly stable, nontoxic protein that is capable of modulating immune responses. This study was conducted to determine whether mucosal administration of EtxB can block collagen-induced arthritis (CIA) and to investigate the mechanisms involved. METHODS: Clinical arthritis in DBA/1 mice was monitored following mucosal administration of EtxB on 4 occasions. The dependence of disease prevention on receptor binding by EtxB and the associated alterations to the immune response to type II collagen (CII) were assessed. Adoptive transfer experiments and lymph node cell cocultures were used to investigate the underlying mechanisms. RESULTS: Both intranasal and intragastric delivery of EtxB were effective in preventing CIA; a 1-microg dose of EtxB was protective after intranasal administration. A non-receptor-binding mutant of EtxB failed to prevent disease. Intranasal EtxB lowered both the incidence and severity of arthritis when given either at the time of disease induction or 25 days later. EtxB markedly reduced levels of anti-CII IgG2a antibodies and interferon-gamma (IFNgamma) production while not affecting levels of IgG1, interleukin-4 (IL-4), or IL-10. Disease protection could be transferred by CD4+ T cells from treated mice, an effect that was abrogated upon depletion of the CD25+ population. In addition, CD4+CD25+ T cells from treated mice were able to suppress anti-CII IFNgamma production by CII-primed lymph node cells. CONCLUSION: Mucosal administration of EtxB can be used to prevent or treat CIA. Modulation of the anti-CII immune response by EtxB is associated with a reduction in Th1 cell reactivity without a concomitant shift toward Th2. Instead, EtxB mediates its effects through enhancing the activity of a population of CD4+ regulatory T cells. PMID- 12115199 TI - C3-Tat/HIV-regulated intraarticular human interleukin-1 receptor antagonist gene therapy results in efficient inhibition of collagen-induced arthritis superior to cytomegalovirus-regulated expression of the same transgene. AB - OBJECTIVE: To achieve disease-inducible expression of recombinant antiinflammatory proteins in order to allow autoregulation of drug dose by natural homeostatic mechanisms. METHODS: We compared the inducible 2-component expression system (C3-human immunodeficiency virus/transactivator of transcription [C3-Tat/HIV]) with the constitutive cytomegalovirus (CMV) promoter in the polyarticular collagen-induced arthritis (CIA) model in mice. DBA/1 mice were immunized with bovine type II collagen and were given boosters on day 21. On day 22, mice were injected intraarticularly with the adenoviral vectors AdCMVLuc, AdCMVhIL-1Ra, AdC3-Tat/HIV-Luc, or AdC3-Tat/HIV-hIL-1Ra. The injected knee joints and hind paws were then scored for signs of arthritis, and knee joint histology was compared. RESULTS: The CMV-driven interleukin-1 receptor antagonist (IL-1Ra) expression resulted in a high constitutive expression and amelioration of CIA. C3 Tat/HIV-driven IL-1Ra expression could be detected only on days 24, 29, and 35. Fourteen days after injection of the vectors, CIA was significantly better inhibited by the C3-Tat/HIV-driven IL-1Ra expression compared with the CMV-driven IL-1Ra expression. Moreover, prevention of CIA in the knee joints also prevented CIA in the untreated hind paws. CONCLUSION: Our data demonstrate for the first time the feasibility of an inducible expression system for local production of IL 1Ra for treatment of arthritis in the CIA model. PMID- 12115201 TI - Fenofibrate inhibits reactive amyloidosis in mice. AB - OBJECTIVE: To examine the effects of the lipid-lowering agent fenofibrate on experimental AA amyloidosis and on serum amyloid A (SAA) levels. METHODS: Fenofibrate was administered orally in a mouse model of amyloidosis, which is induced by injections of amyloid-enhancing factor and Freund's complete adjuvant. Fenofibrate was given for 3 weeks, including a 1-week course before induction of amyloidosis. Splenic amyloid deposits were evaluated histologically, and SAA levels were measured. RESULTS: Fenofibrate inhibited the formation of splenic amyloid deposits and suppressed the elevation of SAA levels. CONCLUSION; Fenofibrate inhibits experimental amyloidosis by reducing levels of the precursor SAA. PMID- 12115204 TI - Nephrotic syndrome associated with anti-tumor necrosis factor alpha therapy in a patient with rheumatoid arthritis: comment on the article by Charles et al. PMID- 12115202 TI - The role of intravenous immunoglobulin therapy in mediating skin fibrosis in tight skin mice. PMID- 12115205 TI - Sample sizes estimated in clinical trials using either a composite index (ASAS response criteria) or single outcome variables in ankylosing spondylitis. PMID- 12115207 TI - No evidence of an association between the interleukin-10 promoter GCC allele and primary Sjogren's syndrome: comment on the article by Hulkkonen et al. PMID- 12115210 TI - Clinical images: Hydroxychloroquine-associated mucocutaneous hyperpigmentation. PMID- 12115213 TI - Reflections on September 11, 2001. PMID- 12115214 TI - Fibromyalgia: where are we a decade after the American College of Rheumatology classification criteria were developed? PMID- 12115216 TI - Criteria for early rheumatoid arthritis: from Bayes' law revisited to new thoughts on pathogenesis. PMID- 12115215 TI - Abnormal T cell signal transduction in systemic lupus erythematosus. PMID- 12115220 TI - Etanercept in the treatment of adult patients with Still's disease. AB - OBJECTIVE: To evaluate the safety and efficacy of etanercept in the treatment of adult patients with Still's disease. METHODS: Twelve adult patients who met criteria for Still's disease and had active arthritis were enrolled in a 6-month open-label trial of etanercept given in biweekly doses of 25 mg. The mean disease duration at study entry was 10.7 years. All patients had been treated unsuccessfully with other disease-modifying antirheumatic drugs. Efficacy was evaluated according to American College of Rheumatology (ACR) improvement criteria, and adverse events were recorded. RESULTS: Ten patients successfully completed the study; 2 withdrew due to disease flare. In 4 patients, the dosage of etanercept was increased from 25 mg biweekly to 25 mg 3 times per week. Seven patients met ACR 20% response criteria. Of these 7 responders, 4 met ACR 50% response criteria and 2 met ACR 70% response criteria. Among the 3 patients with systemic features of Still's disease (fever and rash), improvement in these features was seen in 1; the arthritis did not improve in any of these 3 patients. Except in the 2 patients who withdrew due to disease flare (rash, fever, and arthritis), no other significant adverse events occurred. CONCLUSION: In this initial study of etanercept therapy for Still's disease in the adult, this treatment resulted in improvement in the arthritis and was well tolerated. Additional trials should be performed to elucidate the effects of tumor necrosis factor inhibitors in Still's disease. PMID- 12115219 TI - Treatment of rheumatoid arthritis with methotrexate and hydroxychloroquine, methotrexate and sulfasalazine, or a combination of the three medications: results of a two-year, randomized, double-blind, placebo-controlled trial. AB - OBJECTIVE: To compare the efficacy of combination therapy with methotrexate (MTX) and hydroxychloroquine (HCQ), MTX and sulfasalazine (SSZ), and MTX, HCQ, and SSZ in patients with rheumatoid arthritis (RA). METHODS: RA patients (n = 171) who had not previously been treated with combinations of the study medications were randomized to receive 1 of the 3 treatment combinations in this 2-year, double blind, placebo-controlled protocol. HCQ was given at a dosage of 200 mg twice a day. The dosage of MTX was accelerated from 7.5 mg/week to 17.5 mg/week in all patients who were not in remission. Similarly, the dosage of SSZ was escalated from 500 mg twice a day to 1 gm twice a day in patients who were not in remission. The primary end point of the study was the percentage of patients who had a 20% response to therapy according to the American College of Rheumatology (ACR) criteria at 2 years. RESULTS: Intent-to-treat analysis revealed that patients receiving the triple combination responded best, with 78% achieving an ACR 20% response at 2 years, compared with 60% of those treated with MTX and HCQ (P = 0.05) and 49% of those treated with MTX and SSZ (P = 0.002). Similar trends were seen for the ACR 50% response, with 55%, 40%, and 29% of patients in the 3 treatment groups, respectively, achieving these results at 2 years (P = 0.005 for the triple combination group versus the MTX and SSZ group). All combination treatments were well-tolerated. Fourteen patients (evenly distributed among the 3 groups) withdrew from the protocol because of symptoms that were potentially related to the study medication. CONCLUSION: The triple combination of MTX, SSZ, and HCQ is well-tolerated, and its efficacy is superior to that of the double combination of MTX and SSZ and is marginally superior to that of the double combination of MTX and HCQ. PMID- 12115221 TI - Assessment of proximal finger joint inflammation in patients with rheumatoid arthritis, using a novel laser-based imaging technique. AB - OBJECTIVE: To evaluate a newly developed laser-based imaging technique for the study of soft tissue changes and acute inflammatory processes of the proximal interphalangeal (PIP) joints in patients with rheumatoid arthritis (RA). METHODS: A novel imaging device was developed which allows the transillumination of PIP joints using laser light in the near-infrared wavelength range. In a first clinical followup study, a total of 72 PIP joints of 22 patients with RA and 64 PIP joints of 8 healthy controls were examined both clinically and with the new laser device. At baseline and at followup after a mean of 6 weeks, clinical signs of synovitis, the joint circumference, and the degree of pain were assessed for each PIP joint in order to determine the clinical degree of inflammation. Different features were extracted from the laser images and evaluated by a neural network. RESULTS: At baseline, 72 PIP joints in the RA patients showed clinical signs of inflammation. At followup, 45 PIP joints showed clinical improvement, 13 showed steady active inflammation, and 14 showed deterioration compared with the first visit. None of the 64 PIP joints in the healthy individuals showed any signs of synovitis. The inflammatory status of 60 of the 72 RA joints examined was classified correctly by laser examination and joint circumference determination, giving a sensitivity of 80%, a specificity of 89%, and an accuracy of 83% in detecting inflammatory changes in affected joints. Laser data and joint circumference determination of healthy joints at followup resulted in an accuracy of 85% in reproducing the image. CONCLUSION: The new laser-based imaging technique allows the transillumination of PIP joints and gives information about the inflammatory status of the joint after processing through a neural network. Our data indicate that laser imaging may provide additional information in the early diagnosis of an inflammatory joint process and may prove particularly useful in assessing acute joint inflammation at followup. PMID- 12115223 TI - Presence of autoantibodies to the glycolytic enzyme alpha-enolase in sera from patients with early rheumatoid arthritis. AB - OBJECTIVE: To identify a new autoantigen/autoantibody population in rheumatoid arthritis (RA) sera. METHODS: Following a population-based recruitment effort, 255 patients with very early arthritis (median disease duration 4 months) were studied using different clinical, biologic, and radiologic assessments. After a followup period of 1 year, patients were classified as having RA (n = 145), non RA rheumatic diseases (n = 70), and undifferentiated arthritis (n = 40). Patients' sera were analyzed by one-dimensional (1D) and 2D Western blotting. The recognized 50-kd protein was analyzed by matrix-assisted laser desorption ionization-time-of-flight (MALDI-TOF) mass spectrometry (MS). RA serum reactivities were evaluated against the recombinant protein synthesized by an in vitro coupled transcription-translation system. RESULTS: On 1D Western blots, 36 of the 145 RA sera bound to a 50-kd polypeptide. On 2D Western blots, anti-50-kd+ RA sera recognized a triplet of isoelectric point 6.5-7.0 and a molecular mass of 50 kd. The 3 spots of the triplet were analyzed by MALDI-TOF MS and were shown to correspond to human alpha-enolase. A goat anti-enolase antiserum, which recognized a band comigrating with the 50-kd antigen on 1D Western blots, gave a labeling pattern on 2D Western blots similar to that observed with anti-50-kd+ RA sera. Among the 36 RA sera that identified alpha-enolase in protein maps, only 8 recognized the recombinant (unmodified) alpha-enolase. The specificity of anti alpha -enolase antibodies for RA was 97.1%. Half of the anti-alpha -enolase positive RA patients were negative for both rheumatoid factor and antifilaggrin antibodies. The presence of anti-alpha-enolase antibodies was the greatest predictive factor of radiologic progression in the first 66 RA patients included. CONCLUSION: Autoantibodies to alpha-enolase, an enzyme of the glycolytic pathway, are present in the sera of patients with very early RA and have potential diagnostic and prognostic value for RA. PMID- 12115222 TI - Identification of citrullinated rheumatoid arthritis-specific epitopes in natural filaggrin relevant for antifilaggrin autoantibody detection by line immunoassay. AB - OBJECTIVE: To identify immunodominant epitopes in natural filaggrin that are reactive with antifilaggrin autoantibodies (AFA) in the sera of patients with rheumatoid arthritis (RA) and to explore their use in a diagnostic assay format. METHODS: Based on the results of epitope mapping of human natural filaggrin as well as molecular modeling and computational chemistry, synthetic peptides together with recombinant citrullinated filaggrin were evaluated by a line immunoassay (LIA) for AFA detection. Diagnostic performance was assessed using 336 RA and 253 disease control sera and was compared with that of reference methods. RESULTS: Several immunoreactive epitopes were identified in natural filaggrin, all of which contained at least 1 citrulline residue. Three antigenic substrates, including 2 synthetic peptides and recombinant citrullinated filaggrin showing maximal reactivity on LIA, were finally selected. Using the 3 antigen LIA3, overall sensitivity, specificity, and positive predictive value for RA were 65.2%, 98.0%, and 89.1%, respectively, compared with 61.9%, 98.8%, and 92.8% using the 2-antigen LIA2 (without recombinant protein). Thirty-seven percent of the rheumatoid factor (RF)-negative RA samples (30 of 81) were AFA positive by LIA2, and 52 of 54 RF-positive control samples had no AFA detected on LIA2. Higher specificity and sensitivity were obtained by LIA2 versus anti-RA33 immunoblot, whereas good agreement was observed with antikeratin antibody testing. LIA performed significantly better than AFA immunoblotting using natural filaggrin, at a specificity level of 99% (P = 0.0047). CONCLUSION: Citrullinated residues are present in immunoreactive epitopes of natural human filaggrin. AFA can be readily detected by citrullinated peptides in an LIA-based test, resulting in high specificity and positive predictive value for RA. The LIA could serve as a user-friendly alternative to existing immunofluorescence tests and AFA immunoblot techniques. Given its complementarity to RF, this test can be a valuable tool in the differential diagnosis of arthritis. PMID- 12115224 TI - Cyclosporine inhibition of vascular endothelial growth factor production in rheumatoid synovial fibroblasts. AB - OBJECTIVE: To determine the antiangiogenic effect of cyclosporin A (CSA) in rheumatoid arthritis (RA). METHODS: We investigated the effect of CSA on the production of vascular endothelial growth factor (VEGF) by rheumatoid synovial fibroblasts. Fibroblast-like synoviocytes (FLS) were prepared from the synovial tissues of RA patients, and cultured in the presence of CSA. The production of VEGF by FLS was measured in culture supernatants by enzyme-linked immunosorbent assay. The VEGF messenger RNA (mRNA) expression and activator protein 1 (AP-1) binding activity for VEGF transcription were determined by polymerase chain reaction and electrophoretic mobility shift assay, respectively. RESULTS: CSA dose-dependently inhibited both constitutive and transforming growth factor beta induced VEGF production at the protein and mRNA levels. The suppressive action of CSA on VEGF synthesis was calcineurin dependent, as evidenced by a comparable inhibition by FK-506. Agonists of cAMP, 3-isobutyl-1-methylxanthine and N-2-O dibutyryl-cAMP, mimicked the effect of CSA on VEGF production, while a cAMP antagonist, 2',3'-dideoxyadenosine, abrogated the effect of CSA. A gel mobility shift assay showed that the inhibitory effect of CSA was associated with decreased AP-1 binding activity to the VEGF promoter, in a cAMP-dependent manner. CONCLUSION: CSA may exert an antiangiogenic effect by inhibiting AP-1-mediated VEGF expression in rheumatoid synovial fibroblasts. PMID- 12115225 TI - Macrophage subpopulations in rheumatoid synovium: reduced CD163 expression in CD4+ T lymphocyte-rich microenvironments. AB - OBJECTIVE: The cell surface glycoprotein CD163 is a member of the cysteine-rich scavenger receptor family, highly specific for leukocytes of the mononuclear phagocyte lineage. In vitro, it is induced by glucocorticoids, interleukin-6 (IL 6), and IL-10 and down-regulated by interferon-gamma (IFNgamma), indicating that it has a role in antiinflammatory or other immunomodulatory pathways. We assessed CD163 expression in microenvironments within rheumatoid arthritis (RA) synovium to clarify the relationships among CD4+ T lymphocytes, IFNgamma, and macrophage function in RA. METHODS: Double immunofluorescence and serial immunoenzymatic studies were performed on normal, osteoarthritic, and RA synovium and tonsil with antibodies to CD163, CD45, CD68, CD14, CD3, CD4, CD8, CD19, and IFNgamma. RESULTS: CD163 was observed on all CD14+ cells in synovium and tonsil with the exception of cells within larger T lymphocyte clusters in synovium and within tonsillar follicles. All brightly CD14+ cells in or around vessel walls (interpreted as immigrant monocytes) were CD163+. CD163 labeled fewer cells than did CD68 in synovial intima, but all CD45+ intimal cells were CD163+. CD4+,IFNgamma+ T lymphocytes in RA synovium were chiefly localized within clusters containing CD68+, CD163- cells. CONCLUSION: Within RA synovium, CD163 has major advantages as a macrophage marker and does not appear to be restricted to "mature" macrophages. CD163 discriminates between synovial macrophages and synovial intimal fibroblasts, which also stain positively for CD68 in diseased tissue. PMID- 12115226 TI - High prevalence of lateral knee osteoarthritis in Beijing Chinese compared with Framingham Caucasian subjects. AB - OBJECTIVE: We recently reported that, despite their thinness, elderly subjects in Beijing, China had an equal prevalence (in men) or higher prevalence (in women) of both radiographic and symptomatic knee osteoarthritis (OA) compared with that in the Framingham, Massachusetts cohort of elderly subjects. Our objective was to evaluate whether Chinese subjects might have more medial disease than do Caucasians, given a report of varus alignment in the knee joints of Chinese elderly. METHODS: We compared the prevalence of medial and lateral radiographic knee OA and measured anatomic alignment in the knees of elderly subjects from the Beijing and Framingham cohorts. Both studies recruited a random sample of the population. The Beijing OA Study used the Framingham OA Study protocol for radiographs. Anteroposterior weight-bearing films were read for Kellgren and Lawrence (K/L) grade and joint space narrowing (JSN; 0-3 scale) in each compartment. Medial disease was defined when radiographs showed a K/L grade >or=2 and medial JSN >or=1, and lateral disease was assessed in a comparable manner. Using knee-specific analyses, we compared the prevalence of medial and lateral knee OA after adjusting for age, body mass index, and the correlation between the 2 knees. Restricting the analyses to knees with JSN >or=2 and comparing the proportion of OA knees with medial disease and lateral disease yielded similar results. We assessed alignment in knees from 100 persons without OA, measuring the angle subtended by the femoral and tibial anatomic axes. RESULTS: We studied 1,781 Chinese subjects ages 60-88 years, and 1,084 Framingham subjects ages 63-93 years. Whereas medial OA was less prevalent among the Beijing men, lateral OA was more than twice as prevalent among the Beijing men and women, compared with that in the Framingham elderly. For example, of all knees with radiographic OA, 28.5% of Beijing women's knees had lateral OA versus 11% of Framingham women's knees (P < 0.001), and among men, 32.3% of Beijing men's knees versus 8.8% of Framingham men's knees (P < 0.001) had lateral OA. Alignment was more valgus in the Beijing men than in the Framingham men (mean 4.5 degrees versus 2.7 degrees valgus, respectively; P < 0.001), but no differences in alignment were evident in the women. CONCLUSION: In this first attempt to compare the characteristics of OA in different racial groups, we conclude that, opposite to expectations, Chinese subjects have much more lateral OA than do Caucasian subjects in the Framingham cohort, a predilection possibly explained in the men by differences in anatomic alignment. PMID- 12115227 TI - Knee pain reduces joint space width in conventional standing anteroposterior radiographs of osteoarthritic knees. AB - OBJECTIVE: A suspected, but heretofore undemonstrated, limitation of the conventional weight-bearing anteroposterior (AP) knee radiograph, in which the joint is imaged in extension, for studies of progression of osteoarthritis (OA) is that changes in knee pain may affect extension, thereby altering the apparent thickness of the articular cartilage. The present study was undertaken to examine the effect of changes in knee pain of varying magnitudes on radiographic joint space width (JSW) in the weight-bearing extended and the semiflexed AP views, in which radioanatomic positioning of the knee was carefully standardized by fluoroscopy. METHODS: Fifteen patients with knee OA underwent a washout of their analgesic/nonsteroidal antiinflammatory drug (NSAID) agents (duration 5 half lives), after which standing AP and semiflexed AP knee radiographs of both knees were obtained. Examinations were repeated 1-12 weeks later (median 4.5 weeks, mean 6.0 weeks), after resumption of analgesic/NSAID therapy. Knee pain was measured with the pain subscale of the Western Ontario and McMaster Universities Osteoarthritis (WOMAC) Index (Likert scale). JSW was measured with a pair of calipers and a magnifying lens. Mixed model analyses of variance were used to test the significance of changes in pain and JSW within and between 2 groups of knees with mild-to-moderate radiographic severity of OA: (a) "flaring knees," in which the patient rated standing knee pain as severe or extreme after the washout and in which pain decreased to any degree after resumption of analgesics and/or NSAIDs (n = 12) and (b) "nonflaring knees," in which standing knee pain was absent, mild, or moderate after the washout or did not decrease after resumption of treatment (n = 15). RESULTS: After reinstitution of treatment, WOMAC pain scores decreased significantly in both flaring and nonflaring knees (-44%; P < 0.0001 and -18%; P < 0.01, respectively). After adjustment for the within-subject correlation between knees, mean JSW (+/-SEM) in the extended view of the flaring OA knee increased significantly from the first to second examination (0.20 +/- 0.06 mm; P = 0.005). In contrast, the change in adjusted mean JSW in the extended view of the nonflaring OA knee was negligible (-0.04 +/- 0.04 mm) and significantly smaller than that observed in flaring knees (P < 0.01). Mean JSW in the semiflexed AP view was unaffected by the severity or responsiveness of standing knee pain in flaring and nonflaring OA knees. CONCLUSION: JSW in weight bearing extended-view radiographs of highly symptomatic OA knees can be altered significantly by changes in joint pain. In clinical trials and in epidemiologic studies of OA progression that use this radiographic technique, longitudinal variations in pain may confound changes in the apparent thickness of the articular cartilage. PMID- 12115228 TI - Small nerve fiber involvement in systemic lupus erythematosus: a controlled study. AB - OBJECTIVE: To determine if patients with systemic lupus erythematosus (SLE) may have a peripheral neuropathy involving unmyelinated and small, myelinated nerve fibers, by immunostaining epidermal nerve fibers (ENF) in skin biopsy samples for the panaxonal marker, protein gene product 9.5 (PGP 9.5). METHODS: Fifteen consecutive and nonselected SLE patients and 15 age- and sex-matched controls were included in the study. The age of the patients ranged from 25 years to 65 years, with a mean +/- SD age of 47.3 +/- 10.2 years and a disease duration of 2 28 years (mean +/- SD 14.8 +/- 8.6 years). Two 3-mm skin biopsy samples were obtained with a punch needle approximately 10 cm superior to the lateral malleolus of the right leg and immunostained with 0.1% rabbit polyclonal antibodies to human PGP 9.5. The number of ENF per millimeter was counted and recorded as the mean +/- SD of counts in six 50-microm sections, 3 from each of the 2 biopsy samples. RESULTS: The mean number of ENF per mm in patients with SLE was 8.0 +/- 1.5 (range 5.0-9.9), while the matched controls had 12.2 +/- 3.8 ENF per mm (range 6.8-18.6) (P = 0.0006). CONCLUSION: This study indicates that a small fiber involvement in patients with SLE may be responsible for the prevalent neuropathic symptoms and impaired warm sense that is observed in such patients. PMID- 12115229 TI - A serologic marker for fetal risk of congenital heart block. AB - OBJECTIVE: To analyze the humoral immune response to Ro/SSA and La/SSB antigens in detail, in order to identify markers in mothers at high risk of having children with congenital heart block (CHB). METHODS: Serum samples were obtained from 9 Ro/La-positive mothers who gave birth to affected children, from their 8 newborns with CHB, and from 26 Ro/La-positive mothers whose children were healthy. Antibodies against Ro 52-kd, Ro 60-kd, and La were analyzed by enzyme linked immunosorbent assay and immunoblotting, using recombinant proteins and synthetic peptides. RESULTS: IgG anti-Ro 52-kd antibodies were detected in all mothers who gave birth to children with CHB, as well as in their affected children, but were less frequent and at lower levels in control mothers. Fine mapping revealed a striking difference in which the response in mothers with affected children was dominated by antibodies to amino acids 200-239 of the Ro 52 kd protein (P = 0.0002), whereas the primary activity in control mothers was against amino acids 176-196 (P = 0.001). Furthermore, 8 of 9 mothers of children with CHB had antibody reactivity against amino acids 1-135 of the Ro 52-kd protein, containing 2 putative zinc fingers reconstituted under reducing conditions. CONCLUSION: The results suggest that development of CHB is strongly dependent on a specific antibody profile to Ro 52-kd, which may be a useful tool to identify pregnant Ro/La-positive women at risk of delivering a baby with CHB. Close monitoring of mothers at high risk would enable early detection of a block that is still developing and allow early treatment to combat more serious complications. PMID- 12115230 TI - Fcgamma receptor gene polymorphisms in Japanese patients with systemic lupus erythematosus: contribution of FCGR2B to genetic susceptibility. AB - OBJECTIVE: Human low-affinity Fcgamma receptors (FcgammaR) constitute a clustered gene family located on chromosome 1q23, that consists of FcgammaRIIA, IIB, IIC, IIIA, and IIIB genes. FcgammaRIIB is unique in its ability to transmit inhibitory signals, and recent animal studies demonstrated a role for FcgammaRIIB deficiency in the development of autoimmunity. Genetic variants of FcgammaRIIA, IIIA, and IIIB and their association with systemic lupus erythematosus (SLE) have been extensively studied in various populations, but the results were inconsistent. To examine the possibility that another susceptibility gene of primary significance exists within the FcgammaR region, we screened for polymorphisms of the human FCGR2B gene, and examined whether these polymorphisms are associated with SLE. METHODS: Variation screening of FCGR2B was performed by direct sequencing and polymerase chain reaction (PCR)-single-strand conformation polymorphism methods using complementary DNA samples. Genotyping of the detected polymorphism was done using genomic DNA, with a specific genotyping system based on nested PCR and hybridization probing. Association with SLE was analyzed in 193 Japanese patients with SLE and 303 healthy individuals. In addition, the same groups of patients and controls were genotyped for the previously known polymorphisms of FCGR2A, FCGR3A, and FCGR3B. RESULTS: We detected a single-nucleotide polymorphism in FCGR2B, (c.695T>C), coding for a nonsynonymous substitution, Ile232Thr (I232T), within the transmembrane domain. The frequency of the 232T/T genotype was significantly increased in SLE patients compared with healthy individuals. When the same patients and controls were also genotyped for FCGR2A-131R/H, FCGR3A 176V/F, and FCGR3B-NA1/2 polymorphisms, FCGR3A-176F/F showed significant association. Two-locus analyses suggested that both FCGR2B and FCGR3A may contribute to SLE susceptibility, while the previously reported association of FCGR3B was considered to be secondary and derived from strong linkage disequilibrium with FCGR2B. CONCLUSION: These results demonstrate the association of a new polymorphism of FCGR2B (I232T) with susceptibility to SLE in the Japanese. PMID- 12115231 TI - Expression of recombination activating genes 1 and 2 in peripheral B cells of patients with systemic lupus erythematosus. AB - OBJECTIVE: To analyze immunoregulatory abnormalities in patients with systemic lupus erythematosus (SLE) by assessing the expression of messenger RNA (mRNA) for types 1 and 2 recombination activating genes (RAG) in the peripheral blood of patients with active SLE. METHODS: We examined B cell populations and also individual B cells from patients with SLE for the expression of RAG mRNA. RESULTS: Analysis of bulk mRNA indicated that RAG1 and RAG2 mRNA were found routinely in peripheral B cells of patients with active SLE, but not in healthy subjects. When assessed on a single-cell basis, there was a 3-fold increase in the frequency of RAG1- and RAG2-expressing B cells in SLE patients compared with healthy subjects. Notably, B cells expressing both RAG1 and RAG2 mRNA expressed only IgD mRNA, but not IgG mRNA. Fifty percent of RAG-expressing B cells also expressed VpreB mRNA, whereas all expressed CD154 mRNA. Phenotypic analysis indicated that RAG-expressing B cells were activated, mature B cells. CONCLUSION: These results indicate that RAG expression is up-regulated in peripheral IgD+ and VpreB+ B cells of patients with active SLE. These cells may contribute to the immunoregulatory abnormalities in patients with SLE. PMID- 12115232 TI - Apoptotic U1-70 kd is antigenically distinct from the intact form of the U1-70-kd molecule. AB - OBJECTIVE: To determine whether immune responses to an apoptotically modified form of a human lupus autoantigen can be distinguished from immune responses to the intact form of the same antigen. METHODS: Immunoblot and enzyme-linked immunosorbent assay techniques were used to test human autoimmune sera for the presence of antibodies to apoptotic forms of the U1- 70-kd small nuclear RNP antigen, while antibody recognition of intact U1-70 kd was blocked. RESULTS: poptosis-specific U1-70-kd antibodies were identified by immunoblot in 15 of 29 sera with antibodies to intact U1-70 kd and in 2 of 25 sera without measurable antibodies to intact U1-70 kd. Bacterially produced, purified, caspase-cleaved U1 70 kd without additional posttranslational modifications was a target of apoptosis-specific antibodies in 3 of 9 U1-70-kd-positive sera tested. CONCLUSION: The apoptotic form of U1-70 kd displays B cell epitopes that are not displayed on the intact form of U1-70 kd. Caspase cleavage in the absence of additional posttranslational modifications is sufficient to induce the display of some of these epitopes. Immunity to apoptotically modified proteins can develop against caspase-cleaved forms or against forms that undergo additional posttranslational modification. PMID- 12115233 TI - T cell autoimmunity to histones and nucleosomes is a latent property of the normal immune system. AB - OBJECTIVE: Investigators in this study undertook to determine whether in vitro antigen-responsive immune (polyomavirus T antigen [T-ag]- specific) and autoimmune (histone-specific) T cells from normal individuals share structural and genetic characteristics with those from patients with systemic lupus erythematosus (SLE). METHODS: Histone-specific T cells were generated by stimulation of peripheral blood mononuclear cells (PBMCs) with nucleosome-T-ag complexes and were subsequently maintained by pure histones. T-ag-specific T cell clones were initiated and maintained by T-ag. The frequencies of circulating histone- and T-ag-specific T cells were determined in healthy individuals and in SLE patients by limiting dilution of PBMCs. T cell receptor (TCR) gene usage and variable-region structures were determined by complementary DNA sequencing. These sequences were compared between T-ag- and histone-specific T cells and between normal individuals and SLE patients for each specificity. RESULTS: Individual in vitro-expanded histone- and T-ag-specific T cells from normal individuals displayed identical TCR V(alpha) and/or V(beta) chain third complementarity determining region (CDR3) sequences, indicating that they were clonally expanded in vivo. The frequencies of in vitro antigen-responsive T-ag- or histone-specific T cells from normal individuals were similar to those from SLE patients. Although heterogeneous for variable-region structure and gene usage, histone-specific T cells from healthy individuals and SLE patients selected aspartic and/or glutamic acids at positions 99 and/or 100 of the V(beta) CDR3 sequence. CONCLUSION: Autoimmune T cells from healthy individuals can be activated by nucleosome- T-ag complexes and maintained by histones in vitro. Such T cells possessed TCR structures similar to those from SLE patients, demonstrating that T cell autoimmunity to nucleosomes may be a latent property of the normal immune system. PMID- 12115235 TI - Clinical images: Epidural lipomatosis in a 14-year-old boy with systemic lupus erythematosus. PMID- 12115234 TI - Demethylation of ITGAL (CD11a) regulatory sequences in systemic lupus erythematosus. AB - OBJECTIVE: Inhibition of T cell DNA methylation causes autoreactivity in vitro and a lupus-like disease in vivo, suggesting that T cell DNA hypomethylation may contribute to autoimmunity. The hypomethylation effects are due, in part, to overexpression of lymphocyte function-associated antigen 1 (LFA-1) (CD11a/CD18). Importantly, T cells from patients with active lupus have hypomethylated DNA and overexpress LFA-1 on an autoreactive subset, suggesting that the same mechanism could contribute to human lupus. The present study investigated the nature of the methylation change that affects LFA-1 expression in vitro and in human lupus. METHODS: Bisulfite sequencing was used to determine the methylation status of the ITGAL promoter and flanking regions in T cells from lupus patients and healthy subjects, and in T cells treated with DNA methylation inhibitors. "Patch" methylation of promoter sequences in reporter constructs was used to determine the functional significance of the methylation changes. RESULTS: Hypomethylation of specific sequences flanking the ITGAL promoter was seen in T cells from patients with active lupus and in T cells treated with 5-azacytidine and procainamide. Patch methylation of this region suppressed ITGAL promoter function. CONCLUSION: DNA methylation changes occur in specific sequences that regulate LFA-1 expression in lupus T cells and in the hypomethylation model, indicating that altered methylation of specific genes may play a role in the pathogenesis of lupus. PMID- 12115236 TI - Quantitative computed tomography of the lumbar spine, not dual x-ray absorptiometry, is an independent predictor of prevalent vertebral fractures in postmenopausal women with osteopenia receiving long-term glucocorticoid and hormone-replacement therapy. AB - OBJECTIVE: To determine which measurement of bone mineral density (BMD) predicts vertebral fractures in a cohort of postmenopausal women with glucocorticoid induced osteoporosis. METHODS: We recruited 114 subjects into the study. All had osteopenia of the lumbar spine or hip, as demonstrated by dual x-ray absorptiometry (DXA), and were receiving long-term glucocorticoids and hormone replacement therapy (HRT). Measurements of BMD by DXA of the lumbar spine, hip (and subregions), and forearm (and subregions), quantitative computed tomography (QCT) of the spine and hip (n = 59), and radiographs of the thoracolumbar spine were performed on all subjects to assess prevalent vertebral fractures. Vertebral fracture prevalence, as determined by morphometry, required a >or=20% (or >or=4 mm) loss of vertebral body height. Demographic information was obtained by questionnaire. Multiple regression and classification and regression trees (CART) analyses were used to assess predictors of vertebral fracture. RESULTS: Twenty six percent of the study subjects had prevalent fractures. BMD of the lumbar spine, total hip and hip subregions, as measured by QCT, but only the lumbar spine and total hip, as measured by DXA, were significantly associated with prevalent vertebral fractures. However, only lumbar spine BMD as measured by QCT was a significant predictor of vertebral fractures. CART analysis showed that a BMD value <0.065 gm/cm(3) was associated with a 7-fold higher risk of fracture than a BMD value >or=0.065 gm/cm(3). CONCLUSION: In postmenopausal women with osteoporosis induced by long-term glucocorticoid treatment who are also receiving HRT, BMD of the lumbar spine as measured by QCT, but not DXA, is an independent predictor of vertebral fractures. PMID- 12115237 TI - Viral interleukin-10 gene inhibition of inflammation, osteoclastogenesis, and bone resorption in response to titanium particles. AB - OBJECTIVE: To evaluate the potential of viral interleukin-10 (vIL-10) gene therapy as an approach to prevent wear debris-induced inflammation, osteoclastogenesis, and bone resorption as it relates to periprosthetic osteolysis in patients with total joint replacements. METHODS: Replication defective adenovirus vectors expressing vIL-10 (AdvIL-10) or LacZ (AdLacZ) target genes were used to transduce fibroblast-like synoviocytes (FLS) in vitro, and the effects of these cells on wear debris-induced proinflammatory cytokine production and receptor activator of nuclear factor kappaB ligand + macrophage colony stimulating factor splenocyte osteoclastogenesis were assessed by enzyme-linked immunosorbent assay and tartrate-resistant acid phosphatase assay. The effects of AdvIL-10 administration on wear debris-induced osteolysis in vivo were analyzed using the mouse calvaria model, in which AdLacZ was used as the control. RESULTS: In the presence of AdLacZ-infected FLS, titanium particle-stimulated macrophages exhibited a marked increase in secretion of tumor necrosis factor alpha (TNFalpha) (6.5-fold), IL-6 (13-fold), and IL-1 (5-fold). Coculture with AdvIL-10 transduced FLS suppressed cytokine secretion to basal levels, while addition of an anti-IL-10 neutralizing antibody completely blocked this effect. The vIL-10 transduced FLS also inhibited osteoclastogenesis 10-fold in an anti-IL-10 sensitive manner. In vivo, titanium implantation resulted in a 2-fold increase in osteoclasts (P < 0.05) and in a 2-fold increase in sagittal suture area (P < 0.05). This increase over control levels was completely blocked in mice receiving intraperitoneal injections of AdvIL-10, all of whom had measurable serum vIL-10 levels for the duration of the experiment. Immunohistochemistry demonstrated reduced cyclooxygenase 2 and TNFalpha expression in AdvIL-10-infected animals. CONCLUSION: This study demonstrates that gene delivery of vIL-10 inhibits 3 processes critically involved in periprosthetic osteolysis: 1) wear debris induced proinflammatory cytokine production, 2) osteoclastogenesis, and 3) osteolysis. PMID- 12115239 TI - Increased urinary F2-isoprostanes in systemic sclerosis, but not in primary Raynaud's phenomenon: effect of cold exposure. AB - OBJECTIVE: F2-isoprostanes are free radical-dependent arachidonic acid metabolites that are used as clinical markers of lipid peroxidation in systemic sclerosis (SSc) and other microvascular diseases. The objectives of this study were to determine whether the basal urinary levels of F2-isoprostane in SSc patients differ from those in patients with primary Raynaud's phenomenon (RP) and to investigate whether F2-isoprostane formation correlates with the cutaneous microvascular perfusion decrease following cold exposure in SSc patients, patients with primary RP, and healthy controls. METHODS: Eleven women with RP secondary to SSc, 11 women with primary RP, and 11 healthy women were exposed to decreasing room temperature, from 25 degrees C to 15 degrees C, for 40 minutes. Urine samples were obtained before and after the test for gas chromatography/electronic impact mass spectrometry quantification of 15-F(2t) isoprostane (15-F(2t)-IsoP; also called isoprostaglandin F(2alpha) type III). Cutaneous blood flow was monitored using a laser Doppler perfusion imager. RESULTS: The mean +/- SEM urinary 15-F(2t)-IsoP levels at baseline in SSc patients (178 +/- 32 pmoles/mmole of creatinine) were 1.9 times higher than those in healthy controls (95 +/- 11 pmoles/mmole of creatinine) and 1.7 times higher than those in patients with primary RP (107 +/- 19 pmoles/mmole of creatinine) (P < 0.05 for controls and patients with primary RP versus SSc patients). No significant correlation was found between basal urinary 15-F(2t)-IsoP levels and the temperature or cutaneous blood flow decrease in response to the whole-body cooling. Furthermore, the 15-F(2t)-IsoP response to the cooling test was not correlated with the cutaneous blood flow decrease. CONCLUSION: Lipid peroxidation is increased in SSc patients, but not in patients with primary RP. Cold exposure leads to a significant but small increase in 15-F(2t)-IsoP levels that is independent of the cutaneous blood flow decrease. F2-isoprostane quantification may be an interesting pharmacologic tool for monitoring responses to antioxidant treatment in SSc patients. PMID- 12115238 TI - A multicenter, randomized, double-blind, placebo-controlled trial of adjuvant methotrexate treatment for giant cell arteritis. AB - OBJECTIVE: To evaluate treatment with methotrexate (MTX) in patients with newly diagnosed giant cell arteritis (GCA) to determine if MTX reduces GCA relapses and cumulative corticosteroid (CS) requirements and diminishes disease- and treatment related morbidity. METHODS: This was a multicenter, randomized, double-blind study. Over 4 years, 16 centers from the International Network for the Study of Systemic Vasculitides enrolled patients with unequivocal GCA. The initial treatment was 1 mg/kg/day (97% of surface FcgammaRIIIb from neutrophils previously treated with tumor necrosis factor alpha to mobilize intracellular stores of receptor. This treatment profoundly inhibited activation of primed neutrophils by soluble ICs of the type found in diseased rheumatoid joints, but had no effect on phagocytosis and killing of serum-opsonized S aureus. CONCLUSION: FcgammaRIIIb plays a major role in the secretion of toxic products in response to ICs, but little or no role in the phagocytosis and killing of serum-opsonized bacteria. The selective suppression of effector neutrophil function is therefore possible. FcgammaRIIIb, or its intracellular signaling pathway, is a potential therapeutic target in inflammatory diseases such as rheumatoid arthritis, because disruption of its function should decrease inflammatory tissue damage, but not jeopardize host protection against infection. PMID- 12115245 TI - Protoporphyrin IX photodynamic therapy for synovitis. AB - OBJECTIVE: To determine the conditions for synovial accumulation of protoporphyrin IX (PpIX) and photodynamic therapy (PDT)-induced synovial cytotoxicity in vitro and in vivo. METHODS: Synovial tissues were obtained from mice with antigen-induced arthritis (AIA) and incubated with different concentrations of 5-aminolevulinic acid hexyl ester (h-ALA), a PpIX precursor. Following photoexcitation, cell death in synovial tissues was evaluated by Sytox green fluorescence. PDT was performed after intraarticular injection of h-ALA into the knee joints of mice with AIA, and its effect on joint inflammation was assessed by technetium scintigraphy and histology. Synovial biopsy samples were obtained from patients with osteoarthritis (OA; n = 9) and rheumatoid arthritis (RA; n = 7) and studied for PDT-induced cytotoxicity in vitro. RESULTS: Conversion of h-ALA to PpIX was observed in inflamed synovium in mice and humans. Cytotoxicity was confirmed by Sytox green staining in samples subjected to PDT. In the AIA model, injection of affected knees with h-ALA prior to PDT led to a statistically significant reduction of joint damage in the irradiated joints. The preferential transformation of h-ALA to PpIX in inflammatory tissues was confirmed in human synovial biopsy tissues, where RA samples showed higher tissue concentrations of PpIX following incubation with h-ALA than did OA samples. Fluorescence microscopy showed that PpIX was localized to the synovial lining layer, endothelial cells, and macrophages and induced cell death after PDT. CONCLUSION: Our findings suggest that PDT based on the accumulation of PpIX in the synovial membrane may be a rational basis for photodynamic synovectomy in arthritic diseases. PMID- 12115244 TI - Analysis of the function, expression, and subcellular distribution of human tristetraprolin. AB - OBJECTIVE: The zinc-finger protein tristetraprolin (TTP) has been demonstrated to regulate tumor necrosis factor alpha (TNFalpha) messenger RNA (mRNA) instability in murine macrophages. We sought to develop a model system to characterize the effects of human TTP (hTTP) on TNFalpha 3'-untranslated region (3'-UTR)-mediated expression. We also generated a specific polyclonal antibody against hTTP that enabled the examination of the subcellular distribution of hTTP and its RNA binding in vivo. METHODS: Transfection of reporter gene constructs were used to functionally characterize the role of hTTP in regulating TNFalpha expression in a 3'-UTR-dependent manner. An immunoprecipitation reverse transcription-polymerase chain reaction technique, immunoblotting, immunocytochemistry, and sucrose density fractionation were used to identify and localize hTTP. RESULTS: We found that hTTP interacted with human TNFalpha mRNA in the cytoplasm. The presence of the TNFalpha 3'-UTR was sufficient to confer binding by TTP in vivo. This interaction resulted in reduced luciferase reporter gene activity in a TNFalpha 3'-UTR adenine-uridine-rich element (ARE)-dependent manner. Immunoblotting and immunocytochemistry indicated that endogenous and transfected hTTP localized to the cytoplasm. Results of sucrose density fractionation studies were consistent with a polysomal location of hTTP. In rheumatoid synovium, hTTP expression was restricted to cells in the synovial lining layers. CONCLUSION: Through the development of an antiserum specific for hTTP, we have been able to demonstrate that hTTP binds specifically to the TNFalpha 3'-UTR and reduces reporter gene expression in an ARE-specific manner. These studies establish that hTTP is likely to function in a similar, if not identical manner, in the posttranscriptional regulation of TNFalpha. Understanding the posttranscriptional regulation of TNFalpha biosynthesis is important for the development of novel treatment strategies in rheumatoid arthritis. PMID- 12115246 TI - Local interleukin-12 gene transfer promotes conversion of an acute arthritis to a chronic destructive arthritis. AB - OBJECTIVE: To determine whether local overexpression of interleukin-12 (IL-12), a pleiotropic cytokine that promotes the development of naive T cells into Th1 cells, could aggravate murine streptococcal cell wall (SCW)-induced arthritis, a model of acute arthritis. METHODS: C57BL/6 mice were injected intraarticularly with saline or with 10(7) plaque-forming units of control vector (Ad5del70-3) or IL-12 vector (AdmIL-12.1) into the right knee joint 1 day before intraarticular injection of 25 microg of SCW fragments. The development of joint swelling, changes in chondrocyte proteoglycan (PG) synthesis, and joint destruction were examined thereafter. RESULTS: In normal joints, high levels of IL-12 (20 ng/ml on day 1) could be detected after application of the AdmIL-12.1 vector. After 14 days, expression of IL-12 was still found locally, but IL-12 alone did not induce protracted inflammation. Local expression of IL-12, in combination with SCW, markedly aggravated SCW-induced arthritis, as determined by enhanced joint swelling and prolonged inhibition of chondrocyte PG synthesis. Histologic examination on day 21 showed a chronic inflammatory process, with persistent cartilage PG depletion, cartilage erosion, and VDIPEN neoepitope expression (indicative of metalloproteinase activation). The mixture of IL-12 with SCW fragments did not lead to a chronic destructive process in mice deficient for recombination-activating gene 2, indicating the involvement of lymphocytes. In addition, systemic flare of smoldering SCW arthritis, produced by intravenous injection of SCW fragments, was only seen in the AdmIL-12/SCW group. CONCLUSION: These results indicate that local overexpression of IL-12 promotes conversion of an acute arthritis to a chronic destructive immune-mediated process, which is more susceptible to flares. PMID- 12115247 TI - Two NOD Idd-associated intervals contribute synergistically to the development of autoimmune exocrinopathy (Sjogren's syndrome) on a healthy murine background. AB - OBJECTIVE: The NOD mouse is genetically predisposed to the development of at least 2 autoimmune diseases, autoimmune diabetes and autoimmune exocrinopathy (AEC). More than 19 chromosomal intervals (referred to as Idd regions) that contribute to diabetes susceptibility in the NOD mouse model have been identified, but only 2 chromosomal intervals (associated with Idd3 and Idd5) have been shown to control sialadenitis. In the present study, we bred the Idd3 and Idd5 chromosomal intervals from NOD mice into non-autoimmune C57BL/6 mice to determine if these intervals recreate a Sjogren's syndrome (SS)-like phenotype. METHODS: C57BL/6.NODc3 mice carrying Idd3 and C57BL/6.NODc1t mice carrying Idd5 were crossed and intercrossed to generate a C57BL/6.NODc3.NODc1t mouse line homozygous for the Idd3 and Idd5 chromosomal intervals on an otherwise disease resistant genetic background. C57BL/6.NODc3.NODc1t mice were evaluated for biochemical, pathophysiologic, and immunologic markers characteristic of the SS like phenotype present in the NOD mouse. RESULTS: C57BL/6.NODc3.NODc1t mice fully manifested the SS-like phenotype of the NOD mouse, including decreased salivary and lacrimal gland secretory flow rates, increased salivary protein content due in part to less fluid, aberrant proteolytic enzyme activity, decline in amylase activity, appearance of autoantibodies to exocrine gland proteins, and glandular lymphocytic focal infiltrates. Loss of secretory function occurred more rapidly in C57BL/6.NODc3.NODc1t mice (by 12 weeks of age) than in NOD mice (by 16 weeks of age). No signs of insulitis or autoimmune (type 1) diabetes were observed in the C57BL/6.NODc3.NODc1t mice. CONCLUSION: Genes located within the 2 chromosomal intervals Idd3 and Idd5 appear necessary and sufficient for manifestation of AEC. We propose that this murine model of SS-like disease be designated C57BL/6.NOD Aec1Aec2. Identification of specific genes within the Aec1 and Aec2 genetic regions should help elucidate the mechanism(s) underlying SS-like disease. PMID- 12115249 TI - CARD15/NOD2 analyses in spondylarthropathy. PMID- 12115248 TI - Immunization of naive BALB/c mice with human beta2-glycoprotein I breaks tolerance to the murine molecule. AB - OBJECTIVE: Immunization of naive mice with beta2-glycoprotein I (beta2GPI) leads to the generation of pathogenic anticardiolipin antibodies associated with clinical manifestations of the antiphospholipid syndrome (APS). The aim of this study was to determine whether immunization of naive mice with human beta2GPI, which shares homology with mouse beta2GPI molecules, breaks tolerance to murine beta2GPI and leads to the generation of anti-mouse beta2GPI. METHODS: Twenty-four female BALB/c mice were immunized in the footpads with 10 microg of human beta2GPI. Twelve age- and sex-matched BALB/c mice were immunized in the same manner with Freund's complete adjuvant and served as controls. The reactivity of whole sera, polyclonal IgG, and affinity-purified anti-beta2GPI IgG antibodies against human, bovine, and mouse beta2GPI was evaluated by enzyme-linked immunosorbent assay. RESULTS: High titers of anti-human beta2GPI IgG antibodies were detected 1 month after immunization. Progressively increasing titers against murine and bovine beta2GPI were recorded 1-4 months after injection. CONCLUSION: Immunization of mice with human beta2GPI resulted in the generation of antibodies reacting with human, bovine, and murine beta2GPI. The loss of tolerance to mouse beta2GPI is attributable to the high interspecies homology of beta2GPI. These results may point to molecular mimicry as a possible cause of APS. PMID- 12115251 TI - American College of Rheumatology donation of educational materials to international rheumatology community. PMID- 12115253 TI - Regression of subacute cutaneous lupus erythematosus in a patient with rheumatoid arthritis treated with a biologic tumor necrosis factor alpha-blocking agent: comment on the article by Pisetsky and the letter from Aringer et al. PMID- 12115255 TI - Close relationship between systemic lupus erythematosus and thrombotic thrombocytopenic purpura in childhood: comment on the article by Brunner et al. PMID- 12115256 TI - Increased levels of interleukin-18 in patients with systemic lupus erythematosus: comment on the article by Shibatomi et al. PMID- 12115258 TI - Autoimmune diseases may result from inappropriate RNA polymerase III transcription: comment on the article by Neidhart et al. PMID- 12115261 TI - More evidence on the dysautonomic nature of fibromyalgia: the association with short stature. PMID- 12115259 TI - Folic acid and folinic acid supplementation during low-dose methotrexate therapy for rheumatoid arthritis: comment on the article by van Ede et al. PMID- 12115262 TI - Clinical images: Dialysis-associated uremic tumoral calcinosis. PMID- 12115267 TI - Ganglionic eminence of the human fetal brain--new vistas. AB - This review deals with recent findings concerning the complex functions of the ganglionic eminence (GE), which represents a conspicuous domain of the telencephalic proliferative zone and persists nearly throughout fetal life. The GE not only contains precursor neurons of the basal ganglia, it also contributes significantly to the population of interneurons in the cerebral cortex and to a population of thalamic neurons. The latter migrate through a distinct transient structure, the gangliothalamic body (GTB). The GE also represents an intermediate target for growing thalamic axons (on their way to the cerebral cortex) and cortical axons (on their way to the thalamus). In developmental neuropathology the GE plays an important role in prematurely born infants. The pathogenesis of GE bleedings is discussed with regard to the abundant expression of interleukin-6 (IL-6) receptors on GE cells. The consequences of such bleedings are discussed in view of cellular responses, such as the induction of leukemia inhibitory factor (LIF) expression in GE cells after hemorrhage. PMID- 12115268 TI - Claudin localization in cilia of the retinal pigment epithelium. AB - Using immunocytochemistry and confocal microscopy we demonstrate that claudin immunoreactivity is a novel marker for retinal pigment epithelial cilia. Claudin immunoreactivity obtained by polyclonal anti-claudin 1 antibody, which could crossreact with claudin 3, was colocalized with acetylated tubulin immunoreactivity in cultured human retinal pigment epithelial cells. Claudin immunoreactivity associated with the retinal pigment epithelium (RPE) cilia was more intense than was claudin-immunoreactivity in the junctional complex. Approximately two-thirds of the RPE cells in the rat contain cilia that are immunoreactive with acetylated tubulin on postnatal day 1, and a significant portion of these cilia label with the anti-claudin 1 antibody. Cilia decrease in frequency over subsequent postnatal days, and are absent by postnatal day 30. As RPE cilia decrease in number during postnatal rat development, claudin immunoreactivity is lost earlier than acetylated tubulin, suggesting that the loss of claudin may initiate RPE cilium degeneration. Claudin-immunoreactivity was not evident in cilia of photoreceptor cells, epithelia of nasal mucosa, small intestine, or colon, suggesting that claudin may be a unique molecule in RPE cilia. These data suggest that cilia of the RPE, unlike cilia on other cell types, contain claudin, and that this molecule may play an important and specific role in the function and/or maintenance of RPE cilia. PMID- 12115269 TI - Formation of short-lived multinucleated giant cells (MGCS) from cultured gilthead seabream macrophages. AB - Monocytes/macrophages obtained from the head kidney and peritoneal cavity of gilthead seabream (Sparus aurata) were cultured using plates from three different manufacturers, and were maintained under different conditions. The effects on the morphology and fusion of monocytes/macrophages of initial cell loading, removal of non-adherent cells at different times after plating, and addition of serum and antibiotics were evaluated by light microscopy, and transmission (TEM) and scanning (SEM) electron microscopy. Despite variations in adherence, the behaviour and the morphological changes in kidney monocytes/macrophages were similar in all three types of plates. When foetal calf serum (FCS) was added to the incubation medium, most of the cells resembling monocytes/macrophages were connected by cytoplasmic extensions that formed bridges after 24 hr in culture. After 30 hr, the monocytes/macrophages started to fuse, forming multinucleated giant cells (MGCs) which gradually increased in size until the culture was 4-5 days old. After 5 days the MGCs started to die, and after a week most had disappeared from the cultures. Cells incubated with medium without serum showed changes similar to those fed with FCS, but some cells survived for 3 weeks. The addition of fish serum to the medium appeared to accelerate all processes: the monocytes/macrophages and MGCs died after 3 days in culture. Antibiotics had no apparent effect on the cultures. Removal of non-adherent cells at different times after plating did not appear to affect cell fusion. Coating the wells with extracellular matrix proteins reduced adherence but did not inhibit cell fusion. Curiously, not all macrophages fused with MGCs, and, unlike MGCs, these macrophages phagocytosed sheep red blood cells (SRBCs). Peritoneal macrophages also fused and formed MGCs in culture, similarly to kidney cells. PMID- 12115270 TI - Conduction system abnormalities in rat embryos induced by maternal hyperthermia. AB - Maternal hyperthermia induces severe malformations in the central nervous system (CNS) in both humans and laboratory animals. These phenomena are accompanied by apoptotic cell death, especially in the developing CNS. Cardiovascular malformations in conjunction with skeletal and CNS abnormalities have been reported in embryos of laboratory animals. In rats, hyperthermic treatment at 43 degrees C for 15 min at day 9 of pregnancy induced various severe external malformations in embryos, such as exencephaly, spina bifida, microphthalmia, anophthalmia, facial cleft or defect, generalized edema, and cardiovascular abnormalities. Examination of the embryonic heart revealed abnormal formation of the conduction system. Although hyperthermia causes marked hemodynamic defects, we could not obtain direct proof of a link between hemodynamic alteration by hyperthermia and malformations of the conduction system. PMID- 12115271 TI - Complete discontinuity of the distal fistula tract from the developing gut: direct histologic evidence for the mechanism of tracheoesophageal fistula formation. AB - The embryogenesis of tracheoesophageal anomalies remains controversial. The purpose of this study was to better define the embryogenesis of developing esophageal atresia with tracheoesophageal fistula (EA/TEF), with specific attention to the controversial issue of whether a discontinuity exists in the foregut during its development of EA/TEF. Pregnant outbred rats were injected with adriamycin (2 mg/kg i.p.) on days 6-9 of gestation (E6-E9). At E12.5 and 13.5, microdissection of the entire foregut was performed. Foreguts were examined by phase microscopy, and serial, precisely transverse sections were created for hematoxylin and eosin (H&E) staining. Gross microdissection of the developing foregut at E12.5 (n = 9) revealed a blind-ending, bulbous fistula tract arising from the middle branch of the tracheal trifurcation (as seen by direct and phase microscopy). No connection with the gut could be appreciated at E12.5, but by E13.5 (n = 10) there was an obvious connection between the fistula and the stomach. Serial H&E transverse sections also demonstrated a blind-ending fistula tract arising from the trachea at E12.5. This fistula tract was clearly discontinuous from the developing stomach, which appeared much further caudal to the end of the fistula tract. These results strongly support a model of experimental TEF wherein the fistula tract arises from a trifurcation of the trachea, and (only during a specific gestational window between days 12.5 and 13.5) there is discontinuity between the fistula tract and the stomach. By day 13.5, the fistula joins with the stomach anlage. These observations in the developing EA/TEF should help to resolve the controversy about the mechanism of EA/TEF formation. PMID- 12115272 TI - Bone density of the arm and forearm as an age indicator in specimens of stranded striped dolphins (Stenella coeruleoalba). AB - The age of odontocetes living in the wild is determined mainly by analysis of dentine layers in sections of the teeth. We examined a series of specimens from striped dolphins (Stenella coeruleoalba, Meyen, 1833) that had stranded along the Italian coast of the Mediterranean sea. The present study analyzes and describes bone density in the arm and forearm of the stranded specimens, and correlates the data with total body length of the animal and age as determined by the number of dentine layers in sections of the teeth. According to our model, age can be predicted on the basis of bone density and total body length of the stranded animal. This is the first study to use bone density as a biological parameter to understand the wear and tear of life in the sea. The results suggest that bone density is a new tool for recording age in wild odontocetes. PMID- 12115273 TI - Sexual dimorphism in digit-length ratios of laboratory mice. AB - The lengths of the index finger (2D) and ring finger (4D) are sexually dimorphic in humans, and men have a smaller 2D:4D ratio compared to women. Prenatal androgens appear to be important in the development of the 2D:4D sex difference, since it has been reported in children as young as 2 years old, and since humans exposed to supernormal prenatal androgen levels display a smaller 2D:4D ratio. We tested whether another mammalian species in which the process of peripheral sexual differentiation is androgen-dependent might also show a sex difference in digit ratios. The 2D:4D ratio of adult outbred mice was calculated for both the left and right rear paws. A sex difference was observed in the right rear paw: female mice had a larger 2D:4D ratio than did males. We also found this difference in prepubescent weanling mice. This sex difference is in the same direction as that observed in humans, and suggests that sexual dimorphism in digit length ratios is a feature common to many, if not all, mammals. The mouse may therefore be a useful animal model for studying the factors that influence finger length patterns, which have recently been correlated with several specific behaviors and disease predispositions in humans. PMID- 12115275 TI - Functional gliding spaces of the dorsal side of the human hand. AB - The clinical and functional importance of gliding spaces of the hand (e.g., their role in the spread of infection or as a consideration in reconstructive surgery) has been repeatedly emphasized. However, only a few studies have provided details regarding the connective tissue spaces in the metacarpal region of the dorsal side of the human hand. The aim of the present study was to analyze the morphology and elucidate the anatomic relation of functional gliding spaces in the metacarpal region on the dorsal side of the human hand in order to provide a better understanding of function, and of clinical disorders and their treatment. To delineate these spaces we used a plastic (Acrifix 90) injection method. Twenty fixed and unfixed cadaver hands were subcutaneously injected with Acrifix 90 (a methacrylate) into the metacarpophalangeal transitional region and into the tendon sheaths of the extensor muscles. Different colors were used to distinguish one injected plastic solution from another. The spreading pattern of the injected medium was analyzed by careful dissection. To delineate the exact bordering structures and the topography of the injected spaces, two hands were plastinated using the E12/E6 technique (von Hagens et al., Anat Embryol 1987;175:411-421), and one hand was injected and embedded in Technovit 7100 for histological investigations. Injecting the plastic into the metacarpophalangeal transitional region of fingers II-IV in a disto-proximal direction, the solution spreads along the surface of the separate extensor tendons. It then coalesces 1-2 cm proximal to the injection points to form a continuous plastic plate, which protrudes between and on top of the previous injected tendon sheaths. In no case was a communication between the paratendinous tissue and the tendon sheaths observed. Laterally, the injected solution is delimited at the radial side of the extensor tendon of the second finger and at the ulnar side of the extensor tendon of the fourth finger. Using the described technique at the fifth finger yields a plastic plate that extends from the injection point to the tendon sheath. However, in two specimens a connection between the plastic injected into the tendon sheath of the fifth finger, and the distal injected solution was observed. PMID- 12115274 TI - Three-dimensional architecture of the myosalpinx in the mare as revealed by scanning electron microscopy. AB - The three-dimensional architecture of the myosalpinx in the mare was investigated by means of scanning electron microscopy (SEM) after removal of interstitial connective tissue with NaOH digestion. In the extramural portion of the tubo uterine junction (TUJ), isthmus, and ampulla, the myosalpinx architecture is represented by a unique muscular structure which runs from the mesosalpinx to the base of the inner mucous folds. This unique muscular structure consists mainly of bundles of muscular fibers independent of one another, which show a multiple spatial arrangement and form a complex network. Such a muscular architecture is likely more suitable for stirring rather than pushing the embryos and gametes through the Fallopian tube. PMID- 12115276 TI - Morphological characteristics of the vomeronasal organ of the newborn Asian elephant (Elephas maximus). AB - The 6-week-old Asian elephant (Elephas maximus) has a well-documented precocious flehmen response to pheromones, suggesting that the pheromone-detecting vomeronasal organ (VNO) is functional very early in the life of this species. To further document this, the VNOs of two newborn elephants were examined in situ and analyzed by light microscopy (LM) to ascertain their structural maturity at birth. A tubular, cartilage-encased VNO was located along the anterior base of each side of the nasal septum. Its rostral end was connected to a duct to the roof of the mouth; the caudal end was attached to a well-defined vomeronasal nerve projecting toward the brain. LM revealed distinctive differences in the mucosae bordering the horseshoe-shaped lumen: a concave, sensory mucosa, and a convex, nonsensory mucosa. Small groups of receptor neurons were observed among ciliated columnar cells in the sensory epithelium. Numerous unmyelinated nerve bundles and blood vessels filled the underlying lamina propria (LP) and a small section of the vomeronasal nerve was conspicuous at one edge. The nonsensory mucosa manifested a thinner epithelium that principally consisted of ciliated columnar cells, some of which showed a granular cytoplasm, and a conspicuous row of basal cells. The LP was replete with acinar glands and ducts that opened into the lumen. This study shows that the VNO of the newborn elephant has reached an advanced stage of structural maturity, closely resembling that of the adult. Its composition supports the view that flehmen at 6 weeks delivers pheromones to a functional VNO. PMID- 12115278 TI - Button osteoma: its etiology and pathophysiology. AB - The present study investigates a circumscribed bony overgrowth on the cranial vault, known as button osteoma (BtO) and referred to here as button lesion (BtL). We discuss its anthropological implications. Data on its histology, location, and population distribution (by age, race, and gender) are provided. Microscopically, BtL is composed of well-organized dense lamellated bone which is poorly vascularized and with very few osteocytes. It forms a dome-shaped roof over an underlying diploeized area which includes the ectocranial table. The frequency of BtL is similar in modern (37.6%) and archaeological (41.1%) populations, in blacks, whites, males, and females, and correlates with age. It is rare in nonhuman primates. Fifty-five percent of the human skulls studied by us had BtL only on the parietal, 23.6% on the frontal, and 3.6% on the occipital bones. Fifteen percent had BtL on both the frontal and parietal bones. No lateral preference was found. Most skulls with BtL (64.1%) had only one lesion, 20.4% had two BtL, and 15.4% demonstrated multiple BtL. The average number of button osteomas on an affected skull was 1.97. The frequency of large osteomas (0.5-1.0 cm) was similar in young and old age groups. The demographic characteristics of BtL, mainly its high frequency among ancient and modern populations, its independence of sex and race, its scarcity in other primates, and the fact that its macro- and microstruture are indicative of an hamartoma (and not an osteoma or exostosis) suggest an evolutionary background to the phenomenon. PMID- 12115277 TI - Serpens endocrania symmetrica (SES): a new term and a possible clue for identifying intrathoracic disease in skeletal populations. AB - This paper describes a phenomenon in the endocranial plate, which we have termed "serpens endocrania symmetrica" (SES), and discusses its value as a diagnostic tool. The affected discolored bone area exhibits disruption of the endocranial surface, lending it a maze-like appearance. Histological sections demonstrate that the process is limited to the most superficial portion of the endocranium, with no diploic and ectocranial involvement (sinus areas excepted). Adult skulls (n = 1,884) from the Hamann-Todd collection (HTH), housed at the Cleveland Museum of Natural History, were utilized for the present study. SES was recognized in 32 of the 1,884 skulls studied (1.7%). The frequency of SES among individuals reported to have died from tuberculosis (TB) was 4.4%. The rate of SES in the non TB sample was only 0.53%. The locations were as follows: limited to sinus area, 28.1%; calvarium (excluding the sinuses), 46.9%; sinus + calvarium, 25.0%. SES was bilateral in 90.9% of cases. Twenty-five of the 32 individuals (78.1%) with SES in the HTH collection had tuberculosis specifically listed as the cause of death. Six of the other 7 individuals had infections other than TB. In 29 of the 32 individuals with SES, infection involved structures within the thorax. As SES was also associated with another osteological phenomenon known to represent pulmonary disease, i.e., hypertrophic osteoarthropathy (HOA; 68.0% of SES individuals also had HOA), SES may be of diagnostic value in paleopathology for the recognition of intrathoracic disease, and perhaps tuberculosis. PMID- 12115279 TI - Origins of primate locomotion: gait mechanics of the woolly opossum. AB - The locomotion of primates differs from that of other mammals in three fundamental ways. During quadrupedal walking, primates use diagonal sequence gaits, protract their arms more at forelimb touchdown, and experience lower vertical substrate reaction forces on their forelimbs relative to their hindlimbs. It is widely held that the unusual walking gaits of primates represent a basal adaptation for movement on thin, flexible branches and reflect a major change in the functional role of the forelimb. However, little data on nonprimate arboreal mammals exist to test this notion. To that end, we examined the gait mechanics of the woolly opossum (Caluromys philander), a marsupial convergent with small-bodied prosimians in ecology, behavior, and morphology. Data on the footfall sequence, relative arm protraction, and peak vertical substrate reaction forces were obtained from videotapes and force records for three adult woolly opossums walking quadrupedally on a wooden runway and a thin pole. For all steps recorded on both substrates, woolly opossums always used diagonal sequence walking gaits, protracted their arms beyond 90 degrees relative to horizontal body axis, and experienced peak vertical substrate reaction forces on forelimbs that were significantly lower than on hindlimbs. The woolly opossum is the first nonprimate mammal to show locomotor mechanics that are identical to those of primates. This case of convergence between primates and a committed fine-branch, arboreal marsupial strongly implies that the earliest primates evolved gait specializations for fine-branch locomotion, which reflect important changes in forelimb function. PMID- 12115280 TI - Out of the mouths of baboons: stress, life history, and dental development in the Awash National Park hybrid zone, Ethiopia. AB - The techniques of dental histology provide a method for reconstructing much of the life history of an individual, as accentuated increments visible in polarized light microscopy record incidents of physiological stress during the formation of dental tissues. Combined with counts of the normal periodic growth increments, they provide a means of reconstructing the chronology of dental development, age at death, and the ages at which stress occurs. In this study, we determine age at death and reconstruct the chronology of dental development in two male anubis baboons from Uganda and two female baboons from the Awash National Park hybrid zone. For the female baboons, we used the dates of death and rainfall records for the region to determine date of birth, ages at periods of physiological stress, dates at which these stresses occurred, and rainfall amounts for those months. Ages determined histologically for each specimen are comparable to ages estimated from dental emergence schedules and dental scores for wild baboons. Crown formation times are longer than those reported in radiographic studies of captive yellow baboons. Age at initiation of crown formation is similar to that reported for radiographic studies, but ages at completion of crown formation are consistently later. The pattern of stresses is similar in the two female baboons, suggesting that individual life history intersects with local ecology to produce a pattern of accentuated increments occurring during the weaning process and at the onset of menarche, as well as during the first postweaning dry and rainy periods. PMID- 12115281 TI - Aggressive and neutral interventions in conflicts in captive groups of brown lemurs (Eulemur fulvus fulvus). AB - Third-party interventions in conflicts have revealed complexity in primate social relationships. This type of intervention has seldom been analyzed in prosimians, although many of these species exhibit complex (multimale/multifemale) social organizations. The present study on captive brown lemurs (Eulemur fulvus fulvus) shows that dominant individuals were more likely to intervene in conflicts. Both males and females intervened aggressively in conflicts. Female aggressive interventions occurred mainly on behalf of close kin, whereas males mainly intervened on behalf of juveniles. This study also provides the first record of neutral or peaceful interventions in lemurs. Although females intervened neutrally, almost all neutral interventions were by dominant males. Dominant males intervened in conflicts neutrally more often than aggressively, principally in conflicts between adults and juveniles or between juveniles. Neutral interventions by males always ended the conflicts and were often followed by affiliative contacts between participants (intervenors and opponents). In lemurs, female interventions can be explained by kin selection, while the nature of dominant males' interventions suggests a control role. Interventions by males on behalf of juveniles may increase the formers' fitness. PMID- 12115283 TI - Longitudinal analysis of length growth in the chimpanzee (Pan troglodytes). AB - The adolescent growth spurt in linear dimension in humans is considered to be unique among mammals, but few comparative studies have been done, even on chimpanzees. Growth of the summed length of crown to rump, thigh, and leg was studied longitudinally in 12 chimpanzees. We took body weight growth and reproductive maturation into consideration. Reproductive maturation was monitored by the swelling of sexual skin and menarche in females, and by testicular development in males. We applied two relationships found in humans between body length growth and the environment to the chimpanzees. The first relationship was the robustness of the growth spurt, meaning that the spurt is absent only in individuals under the most severe environmental pressure. Subjects maturing in a favorable or even mediocre environment are anticipated to show the growth spurt. The second relationship was catch-up growth, where, when the environment is ameliorated, growth may be accelerated to attain the target size. Catch-up growth at the end of the juvenile period may mimic the adolescent growth spurt. Results showed that subjects living under favorable conditions did not exhibit a growth spurt, and that it was only the subjects who had delayed growth in the juvenile period that showed a spurt in adolescence, the period when reproductive maturation occurred. Although we have concluded that chimpanzees do not have an adolescent growth spurt, except in cases of catch-up growth, this does not mean that they have a different growth pattern from that of humans. The absence of a growth spurt may be associated with adaptations to chimpanzee patrilineal society, where adolescent males are incorporated into the adult hierarchy at a low rank. PMID- 12115282 TI - Genetic drift and gene flow in a prehistoric population of the Azapa valley and coast, Chile. AB - The present paper studies the evolutionary process operating on prehistoric groups from the Azapa valley and coast (northern Chile). The sample consists of 237 crania from the Archaic Late, Early Intermediate, Middle, Late Intermediate, and Late periods. Six metric variables were used, which were transformed to eliminate the special environmental component and to increase the proportion of genetic variance. Population structure was assessed using a method based on quantitative genetic theory, which predicts a lineal relationship between average within-group phenotypic variance and group distance to the population centroid. Results indicate that 17.5% of the total variance accounts for special environmental variance. An excess in extraregional genetic flow is observed in the population corresponding to the Early Intermediate period in the valley. A reduced differentiation is observed between Archaic and Early Intermediate coastal groups, as well as between the latter and the inhabitants of the valley in the same period. Genetic differences between both areas increased substantially since the Middle period. Evidence indicates that long-range gene flow was lower on the coast than in the valley, the lowest estimated Fst being 0.0199 for the total population (coast and valley), 0.0111 for the coastal population, and 0.0057 for the valley. Results are discussed in terms of regional archeological and ethnohistorical evidence, and a microevolutionary model is presented to account for the biological history of the population. PMID- 12115284 TI - Influence of lean body mass on performance differences of male and female distance runners in warm, humid environments. AB - The purpose of this investigation was to evaluate the influence of lean body mass (LBM) and body weight (BW) on the thermoregulatory responses and endurance performance of male and female athletes in warm, humid environments. Ten (5 males, 5 females) healthy, moderately trained athletes with varying physiques performed a self-paced 30-min run on a motorized treadmill in warm (30 degrees C), humid (60% relative humidity) conditions, with the aim of running the greatest distance possible. Males completed one trial, while females completed two trials, one in each of the follicular (Fol) and luteal (Lut) phases of the menstrual cycle in a randomized fashion. There were no significant differences among groups for distance run (males, 5.2 +/- 0.4 km; Fol, 4.9 +/- 0.1 km; Lut, 4.7 +/- 0.1 km). However, following analysis of covariance accounting for LBM and BW, the distances run were significantly different. The adjusted means for distance run after accounting for LBM were 3.4 km for males (P < 0.05), 5.9 km for Fol, and 5.6 km for Lut. Adjusted means accounting for BW resulted in run distances of 6.5 km for males (P < 0.05), 4.2 km for Fol, and 4.0 km for Lut. Thermoregulatory responses such as rectal and skin temperatures were similar among groups. Avenues of heat loss and gain were altered relative to the menstrual cycle phase. The results suggest that one reason for the disparity in performance between male and female athletes over similar race distances might in part be related to unequal body characteristics and in particular to differences in LBM. PMID- 12115285 TI - Remembering Malthus: a preliminary argument for a significant reduction in global human numbers. PMID- 12115286 TI - Early experience with the artificial nipple. AB - The present study examined the newborn pup's responsiveness to artificial nipples that differed in length and width. In a series of experiments, pups were: exposed to a long (5.0 mm), thin (1.0 mm) nipple or a short (2.0 mm), fat (2.5 mm) nipple and their immediate behavioral responses including oral grasping were recorded and quantified (Experiment 1); preexposed to one artificial nipple and then reexposed to another artificial nipple to determine how early nipple experience affected subsequent responding to the artificial nipple (Experiment 2); videotaped to characterize details of oral grasping and to develop a technique for measuring depth of nipple entry into the pup's mouth (Experiment 3a); provided early nipple experience to see how depth of nipple entry into the pup's mouth changed when pups were reexposed to another artificial nipple (Experiment 3b); and exposed to an artificial nipple to determine how early nipple experience changed the pattern of oral grasping and ingestion of milk from a surrogate nipple (Experiment 4). PMID- 12115287 TI - The effects of age and isolation period on two phases of behavioral response to foot shock in isolation-reared rats. AB - Using rats as subjects, the effects of a period of isolation and the subjects' age during isolation on the response to foot shock were systematically examined in three experiments. Both the thresholds of shock that evoked a jumping response and that evoked a thrashing response were measured. The results suggest that the threshold for jumping response decreased when rats were isolated during the postweaning stage of development. On the other hand, the threshold for thrashing response decreased when the subjects were isolated for more than 39 days, independently of their age during isolation. Possible causes of the differential effects of these two factors are discussed in relation to the developmental process of these two behavioral indices. PMID- 12115288 TI - Mutual influence of the maternal hen's food calling and feeding behavior on the behavior of her chicks. AB - It is widely accepted that brooding hens attract their chicks to food by food calling, but until now, the concurrent behavior of hens and chicks has never been investigated in depth. This study provides a detailed analysis of both the behavior of brooded chicks and the distance to their mother in relation to her feeding sequences, and whether they contained food calling. Our results revealed that brooding hens utter food calls while pecking, especially when their chicks are not feeding and/or have been at some distance for several seconds. Chicks' response to their mother's feeding activities was more pronounced in the presence than in the absence of food call. Chicks responded to this call by approaching their mother and increasing their pecking; their response became more efficient as they grew older. We thus consider food calls as an arousal vocalization that directs the chicks' attention to a food item chosen by a hen. PMID- 12115289 TI - Morphine administration selectively facilitates social play in common marmosets. AB - Common marmosets (Callithrix jacchus) form extended families, and several cohorts of young may reside together. Play is shown extensively among the offspring. We hypothesized that opiate activity modulates social play, and predicted that administration of morphine (0.5 mg/kg) would facilitate social play, whereas pretreatment with naloxone (0.5 mg/kg) would block morphine's effects. Morphine administration was associated with significantly increased social play, and the effect of morphine was attributable to the focal subject, since play initiated by others was unaffected by treatment. Non-social categories of play, such as object manipulation and locomotor play, and affiliative behaviors, such as time spent huddled, were unaffected by treatment. Twittering and play face, behaviors used by young during play, also increased after morphine administration. Pretreatment with naloxone blocked morphine's effects. Total activity was significantly increased by morphine administration. We conclude that social play is specifically facilitated by opiate activation, whereas other categories of play behavior and social behavior were unaffected by morphine. Thus, social play represents a distinct category of social behavior in juvenile common marmosets with regulatory processes that are unique from other types of social behavior. PMID- 12115291 TI - Individual housing during the play period results in changed responses to and consequences of a psychosocial stress situation in rats. AB - In the present study, the consequences of individual rearing during the play period on adult responses to aggression were investigated in a complex social setting. In a group of either socially or individually reared rats, an aggressor was repeatedly introduced. Separate control groups of individually and socially reared rats were not exposed to an aggressor. To allow an interpretation of the altered reactivity to aggression in terms of (in)efficient or (in)adequate behavior, not only the responses in the presence of the aggressor but also the consequences of the altered reactivity on subsequent behavior in the absence of the aggressor were observed. We demonstrated that a higher number of injuries, more agitation of the aggressor, and more ultrasonic distress vocalizations accompanied the altered responses of individually reared rats in the presence of the aggressor. In the absence of the aggressor, individually reared rats displayed less social stress-reducing behaviors (play and social grooming) than socially reared rats. It was concluded that individually reared rats display a less adequate response to aggression in this social context. PMID- 12115290 TI - Differences in behavior and monoamine laterality following neonatal clomipramine treatment. AB - Postnatal treatment between 8 to 21 days of age with clomipramine (15 mg/kg, twice daily) produces an animal model that has many of the behavioral hallmarks of depression. In this study, we investigated the enduring behavioral and neurochemical effects of this early treatment in adult animals. Locomotor activity was increased in clomipramine-treated males, but not females, relative to vehicle-treated subjects. Increases in anxiety-like behavior in the elevated plus maze also were observed in clomipramine-exposed adults, but no sex differences were detected. Clomipramine-treated animals had shifts in the laterality of monoamines in limbic regions with lower serotonin levels on the right side while vehicle-treated animals had lower serotonin on the left side. The lateralization of dopamine content demonstrated the same pattern. This decline in monoaminergic content is consistent with clinical studies demonstrating decrements in serotonin as well as alterations in the lateralization of function in individuals with major depressive order. PMID- 12115293 TI - Prenatal ethanol exposure and spatial navigation: effects of postnatal handling and aging. AB - Prenatal ethanol exposure results in spatial navigation deficits in young and mid aged animals. In contrast, postnatal handling attenuates spatial deficits that emerge with age in animals that are not handled. Therefore, we investigated the ability of handling to attenuate spatial deficits in animals prenatally exposed to ethanol (E). Sprague-Dawley male offspring from E, pair-fed (PF), and control (C) groups were handled (H) or nonhandled (NH) from 1 to 15 days of age and tested on the Morris water maze at 2 or 13 to 14 months of age. In young animals, H-E males had longer latencies to locate the submerged platform, and E animals, across handling conditions, showed altered search patterns compared to their PF and C counterparts. Mid-aged animals had longer latencies than young animals, with no differences among E, PF, and C animals. However, corticosterone levels were higher in mid-aged E than in C males. Handling did not attenuate impairments associated with either prenatal ethanol exposure or aging. PMID- 12115294 TI - What does fetal movement predict about behavior during the first two years of life? AB - This study evaluated whether motor activity prior to birth is predictive of motor behavior and temperament in neonates, infants, and toddlers. Three measures of fetal motor activity (activity level, amplitude, and number of movements) were collected at 24, 30, and 36 weeks of gestation in 52 healthy fetuses using Doppler-based actography. Postnatal data collection included a neurobehavioral assessment at 2-weeks postpartum (n = 41), and laboratory-based behavioral observations at 1 and 2 years of age (ns = 35). Individual stability in motor activity was present during gestation. Predictive relations between fetal movement and neonatal behavior were inconsistent; significant but small positive associations were detected between motor behavior at 36 weeks and neonatal irritability and motor development. Fetal activity level at 36 weeks was positively associated with observed 1-year activity level for boys (but inversely related for girls) and maternal report of activity level at 2 years. Fetal movement was consistently and negatively predictive of distress to limitations at 1 year and behavioral inhibition at 2 years, accounting for 21 to 43% of the variance in these measures. Intrafetal variability in motor behavior makes this a relatively unstable metric for prediction to neonatal maturational outcomes, which are relatively constrained, but fetal motor activity appears to predict temperament attributes related to regulatory behaviors in early childhood. PMID- 12115295 TI - Transnatal olfactory continuity in the rabbit: behavioral evidence and short-term consequence of its disruption. AB - This study investigates the role of prenatal odor learning on postnatal adaptive orientation responses in the newborn rabbit. Preference tests revealed that pups are equally attracted to the odors of placentae and colostrum (Experiments 1-4), suggesting that an odor continuity may exist between the fetal and neonatal environments. To test some predictions derived from this hypothesis, we manipulated the odor of the diet of pregnant-lactating does to control the chemical niches of their perinates. Fetuses exposed in this way to the odor of cumin (C) were selectively attracted as neonates to the odor of pure C (Experiment 6). Prenatal exposure to C also was followed, to a certain extent, by enhanced attraction to C odor in the placenta or colostrum from females which had consumed it (Experiments 5 & 7). Finally, the functional implications of perinatal odor continuity were tested by disrupting it. The odor component of the feto-neonatal transitional environment revealed indeed to affect the ability of certain pups to gain colostrum and milk at the very first sucking opportunities (Experiment 8). PMID- 12115296 TI - The effects of age and sex on mental rotation performance, verbal performance, and brain electrical activity. AB - This study examined the effects of age and sex on mental rotation performance, verbal performance, and brain-wave activity. Thirty-two 8-year-olds (16 boys) and 32 college students (16 men) had EEG recorded at baseline and while performing four computerized tasks: a two-dimensional (2D) gingerbread man mental rotation, a 2D alphanumeric mental rotation, of three-dimensional (3D) basketball player mental rotation, and lexical decision making. Additionally, participants completed a paper- and pencil water level task and an oral verbal fluency task. On the 2D alphanumeric and 3D basketball player mental rotation tasks, men performed better than boys, but the performance of women and girls did not differ. On the water level task, men performed better than women whereas there was no difference between boys and girls. No sex differences were found on the 2D gingerbread man mental rotation, lexical decision-making, and verbal fluency tasks. EEG analyses indicated that men exhibited left posterior temporal activation during the 2D alphanumeric task and that men and boys both exhibited greater left parietal activation than women and girls during the 2D gingerbread man task. On the 3D basketball player mental rotation task, all participants exhibited greater activation of the right parietal area than the left parietal area. These data give insight into the brain activity and cognitive development changes that occur between childhood and adulthood. PMID- 12115298 TI - Atypical squamous cells of undetermined significance: a continuing controversy. PMID- 12115297 TI - Twenty-four-hour-old lambs rely more on maternal behavior than on the learning of individual characteristics to discriminate between their own and an alien mother. AB - Lambs can discriminate their own mother from an alien dam on the first day of life, suggesting the recognition of individual physical characteristics of the mother. Alternatively, their choice may depend on behavioral differences existing between the ewes because of their maternal selectivity. To clarify this, the ability of 24-hr-old lambs to discriminate between their own and an alien mother, that were either intact and accept only their own lamb at nursing (i.e., selective, n = 19) or anosmic, which nurse indiscriminately alien lambs as well as their own (i.e., nonselective, n = 24), was assessed by a 5-min, two-choice test. With intact dams, lambs spent significantly more time next to their own mother whereas this was not so in the presence of anosmic dams. Furthermore, in the intact group, the vocal activity by their own mother differed from that by the alien dam while this was not so in anosmic ewes. We conclude that 24-hr-old lambs rely more on the behavior of the ewes to select their dam than on their individual physical characteristics. PMID- 12115299 TI - Reducing or eliminating use of the category of atypical squamous cells of undetermined significance decreases the diagnostic accuracy of the Papanicolaou smear. AB - BACKGROUND: The diagnosis of "atypical squamous cells of undetermined significance" (ASCUS) is controversial, not only for the clinical utility of its subcategories (favor reactive vs. favor dysplasia), but for its very existence as an expression of uncertainty. In the current study, the authors investigated the impact of reducing and eliminating this category on the sensitivity and predictive values for detecting squamous intraepithelial lesions (SILs). METHODS: One hundred cervical Papanicolaou (Pap) smears originally diagnosed as ASCUS, all of which had histologic follow-up, were reviewed blindly and reclassified as either negative (within normal limits/benign cellular changes), low-grade SIL (LSIL), or high-grade SIL (HSIL) by 1 reviewer who eliminated ASCUS as a diagnostic possibility entirely. A second reviewer reinterpreted the cases but attempted to use the ASCUS diagnosis (favor reactive or favor dysplasia) sparingly. All histologic diagnoses were reviewed, and an adjudicated final diagnosis was established. Reviewed smear interpretations were correlated with the histologic diagnosis (original, reviewed, and adjudicated). Statistical analysis was performed using the Fisher exact test. RESULTS: Thirty-eight women had histologically confirmed SIL (21 LSIL cases and 17 HSIL cases [including 1 case of endocervical adenocarcinoma]); 31 of these 38 cases originally were classified as ASCUS, not otherwise specified, 1 case was classified as ASCUS favor reactive, and 6 cases were classified as ASCUS favor dysplasia. The reviewer who used the ASCUS diagnosis sparingly reclassified the smears as negative (62 cases); ASCUS, favor reactive (3 cases); ASCUS, favor dysplasia (13 cases); LSIL (19 cases); and HSIL (3 cases). The reviewer who eliminated the ASCUS category reclassified the smears as negative (59 cases), LSIL (29 cases), and HSIL (12 cases). The rate of SIL/HSIL in those cases interpreted as abnormal was 38%/17% originally, 42%/24% with a reduced ASCUS interpretation, and 37%/17% when the ASCUS category was eliminated. In those ASCUS smears that were reclassified as negative, the SIL/HSIL rate was 35%/13% with the reduced ASCUS interpretation and 39%/17% when the ASCUS category was eliminated. The sensitivity for detecting a SIL/HSIL was reduced from 100%/100% for the original ASCUS interpretation to 42%/53% for the reduced ASCUS interpretation to 39%/41% with the elimination of the ASCUS interpretation. CONCLUSIONS: Although in the current study utilization of the ASCUS diagnosis was found to result in a 62% negative or reactive outcome on biopsy, a significant number of patients with SIL were detected (38% in the current series, 17% with HSIL). Despite the improved correlation with negative biopsies, reducing or eliminating the ASCUS diagnosis appears to decrease the sensitivity of the Pap smear significantly and appears to be no better than chance at predicting a diagnosis of SIL on biopsy, including HSIL. PMID- 12115300 TI - Micropapillary serous carcinoma of the ovary: cytomorphologic characteristics in peritoneal/pelvic washings. AB - BACKGROUND: Micropapillary serous carcinoma (MPSC), a recently described entity, is an ovarian tumor with a distinctive histologic architecture that lacks a destructive infiltrative growth pattern and behaves like a low-grade neoplasm. The purpose of this study was to determine if specific cytomorphologic features were associated with this tumor in peritoneal/ pelvic washings. METHODS: Eight cases of MPSC were retrieved from the cytopathology files at The Johns Hopkins Hospital. Patients ranged in age from 31 to 74 years (mean, 58 years). A cytomorphologic comparison was made with pelvic washings of eight cases of papillary serous carcinoma (PSC) of the ovary. RESULTS: MPSC demonstrated small but well formed papillary fragments (generally < 30 cells) composed of monotonous, relatively small epithelial cells, often with multiple nucleoli. Single, large atypical cells were seldom present and were seen in less than one half of cases. Cellularity was generally high and slide background was clear with minimal inflammatory cells. In comparison, PSC, in addition to the smaller papillary fragments, also exhibited larger more complex papillary fragments (generally > 30 cells) composed of pleomorphic, hyperchromatic cells often with single prominent nucleoli. Single, large tumor cells exhibiting eccentric atypical nuclei or multinucleation were present in high concentration in the majority of PSCs. Psammoma bodies were observed in one half of cases in both tumor types. CONCLUSIONS: Although MPSC shares cytomorphologic similarities with PSC, it can be diagnosed adequately in peritoneal/ pelvic washings. Careful interpretation of the subtle cytologic differences seen in the two tumor types may facilitate the differentiation of these neoplasms for a more appropriate management of the patient. PMID- 12115301 TI - The effects of the current World Health Organization/International Society of Urologic Pathologists bladder neoplasm classification system on urine cytology results. AB - BACKGROUND: Historically, the diagnostic accuracy of urine cytology for the detection of urothelial carcinoma has been low, particularly for low-grade lesions. In 1998, the World Health Organization and International Society of Urologic Pathologists (WHO/ISUP) established a new classification system for urothelial neoplasms. It has been postulated that the accuracy of urine cytology for the detection of low-grade carcinomas should improve, because the current WHO/ISUP system classifies bladder lesions that lack any significant cytologic atypia as papillary urothelial neoplasms of low malignant potential (PUNLMP). Also, the accuracy of urinary cytology for the detection of high-grade lesions is expected to decrease, because the criteria for high-grade lesions is lowered, placing most of the previous WHO Grade 2 tumors into the current high-grade carcinoma category. METHODS: One hundred bladder biopsies that were classified according to the previous WHO classification system were examined by one pathologist and were reclassified according to the new WHO/ISUP scheme. All biopsies had corresponding urine specimens that had been diagnosed previously by a different cytopathologists. The patients' previous WHO and current WHO/ISUP surgical diagnoses were compared with the corresponding urine cytology. In addition, cytospins obtained from 40 patients with a histologic diagnosis of low grade urothelial carcinoma (n = 20 patients) and high-grade urothelial carcinoma (n = 20 patients) according to the WHO/ISUP classification system were reviewed to identify any outstanding cytologic features. RESULTS: According to the original WHO classification system, there were 26 patients with Grade 1 transitional cell carcinoma (TCC), 61 patients with Grade 2 TCC, and 13 patients with Grade 3 TCC. The corresponding cytology was positive in 11 of 26 Grade 1 tumors, 28 of 61 Grade 2 tumors, and 12 of 13 Grade 3 tumors. After the reclassification, there was 1 papilloma, 12 PUNLMP lesions, 50 low-grade urothelial carcinomas, and 37 high-grade carcinomas. The cytology was positive in 0 of 1 papillomas, 5 of 7 PUNLMP lesions, 18 of 50 low-grade carcinomas, and 28 of 37 high-grade carcinomas. In addition, morphologic uniformity and cytoplasmic homogeneity (50.0% and 45.0%, respectively) were seen more commonly in low-grade bladder tumors. CONCLUSIONS: The diagnostic accuracy of urine cytology for high grade lesions decreased, as expected; however, cytologic detection of low-grade urothelial carcinomas was remarkably lower than expected. In addition, no outstanding cytologic features could be identified to make a definitive diagnosis. PMID- 12115302 TI - Imprint cytologic features of pulmonary sclerosing hemangioma: comparison with well-differentiated papillary adenocarcinoma. AB - BACKGROUND: Sclerosing hemangiomas (SH) of the lung are uncommon tumors and are thought to be benign. However, histogenesis of these tumors has not yet been characterized adequately. Moreover, there are few reports dealing with their cytologic features, and it is generally considered difficult to make accurate diagnoses of sclerosing hemangiomas that have a predominantly papillary pattern. METHODS: Cytologic features were analyzed for 15 sclerosing hemangiomas, and cytologic features of sclerosing hemangioma were compared with features of 22 cases of well-differentiated papillary adenocarcinoma classified as pathologic Stage 1A. RESULTS: Blood and round cells were observed more frequently in SH than in adenocarcinomas (P < 0.05), whereas necrosis was seen more frequently in adenocarcinomas than in SH (P < 0.05). The presence of nucleoli, nuclear indentations, irregularities of nuclear margins, nuclear polymorphisms, and high nuclear-cytoplasmic (NC) ratios of tumor cells were observed less frequently in SH. Polynuclear (having three or more nuclei) tumor cells were observed only in adenocarcinoma cases. In morphometric studies, the nuclear areas, cytoplasmic areas, NC ratios, long axes of nuclei, short axes of nuclei, and nuclear rotundity ratios were significantly higher in adenocarcinoma cells than in SH cells (P < 0.05). CONCLUSIONS: The presence of polymorphous cells and tumor cells with bland nuclei are characteristic cytologic findings associated with sclerosing hemangioma. It is possible to make accurate diagnoses for SH cases preoperatively by careful cytologic characterization. PMID- 12115303 TI - Intraoperative assessment of sentinel lymph nodes in patients with breast carcinoma: accuracy of rapid imprint cytology compared with definitive histologic workup. AB - BACKGROUND: As sentinel lymph node biopsy (SNB) becomes a new surgical standard in the treatment of patients with breast carcinoma, there is an emergent need for a fast and accurate method with which to assess the SN intraoperatively, so a decision can be made regarding whether to perform axillary lymph node dissection during primary surgery. In the current study, the authors performed a prospective investigation of the relative merits of imprint cytology for that purpose. METHODS: Seventy-six patients with T1-T2 breast carcinoma were included after undergoing successful SNB. SNs were freshly sectioned at 2-mm intervals and imprint smears were obtained from all cut surfaces. The smears were examined using a rapid May-Grunwald-Giemsa stain variation, and the SNs were judged to be positive or negative for metastases. SNs later were submitted for paraffin embedding and serial sectioning. Both hematoxylin and eosin stained and cytokeratin (CK) immunostained sections were examined. The postoperative evaluation of the SNs was taken as the gold standard. RESULTS: Intraoperative cytology showed a sensitivity of 67.7%, a specificity of 100%, an accuracy of 86.8%, and a negative predictive value of 81.8%. The majority of false-negative cases (8 of 10 cases) were due to micrometastasis in the SNs that were discovered only after exhaustive examination with serial sectioning and CK immunostaining. CONCLUSIONS: The results of the current study demonstrate that the accuracy of imprint cytology is high enough to warrant its use for intraoperative SN assessment. If the findings are negative, axillary lymph node dissection can be omitted. Only a few patients with SN micrometastasis may require reoperation. PMID- 12115304 TI - Fine-needle aspiration cytology of articular and periarticular lesions. AB - BACKGROUND: The cytologic diagnosis of joint and articular surface-based lesions traditionally has been accomplished by examination of fluids or effusions. Although exfoliative cytology remains an accurate diagnostic test, not all joint based lesions will produce effusions that are amenable to this type of examination. Fine-needle aspiration (FNA) represents an excellent alternative to traditional cytologic or histologic methods of diagnosis in joint pathology. METHODS: The authors reviewed FNA materials for the period 1992-2001 from lesions of joint spaces and periarticular soft tissues. All diagnoses based on cytologic materials that were included in this study were confirmed with histologic follow up. Cytologic and histologic materials were prepared using standard methods. RESULTS: The authors found six relatively common lesions that were amenable to diagnosis by FNA. These included rheumatoid nodule, gouty tophi, ganglion cysts, pigmented villonodular synovitis, synovial chondromatosis, and synovial sarcoma. There are potential pitfalls in discriminating gout from pseudogout and synovial chondromatosis from chondrosarcoma. CONCLUSIONS: In most instances, mass producing lesions of the joint space or the periarticular soft tissues can be diagnosed successfully by FNA. The common lesions are easily recognizable and are cytologically distinctive. PMID- 12115305 TI - Cytologic findings of angioimmunoblastic T-cell lymphoma: analysis of 16 fine needle aspirates over 9-year period. AB - BACKGROUND: Peripheral T-cell lymphoma often represents an important diagnostic pitfall in fine-needle aspiration biopsy due to the heterogeneous cell population present. A classic example of this group is angioimmunoblastic T-cell lymphoma (AILD-T). The fine-needle aspiration cytology of this relatively well-defined histologic subtype of T-cell lymphoma is rarely described in the literature. METHODS: The authors reviewed 16 fine-needle aspirates of AILD-T from 9 patients in Queen Mary Hospital and Pamela Youde Nethersole Eastern Hospital, Hong Kong, over a 9-year period from early 1993 to mid-2001. The morphologic features seen in cytology smears and/or cell block sections were correlated with histologic and immunohistochemical findings of excisional biopsy specimens. RESULTS: The smears and cytospin preparations showed a heterogeneous population of hematolymphoid cells, including small lymphocytes; nondescript, medium-sized lymphoid cells; immunoblasts; plasma cells; eosinophils; and reticulum cells, including follicular dendritic cells. In general, tingible body macrophages were not identified. Conversely, follicular dendritic cells were discernible easily in most cases and sometimes were admixed intimately with lymphoid cells, forming dendritic cell-lymphocyte complexes. There also were large lymphoid tissue fragments containing a scaffold of arborizing small vessels. Pleomorphic cells with high mitotic activity or lymphoid cells with clear cytoplasm were not identified. The cell block sections often showed an intimate admixture of small lymphocytes, plasma cells, eosinophils, and reticulum cells amid a background of reticulin fibers. Lymphoid follicles with well-developed germinal centers were never found. The features seen in cytologic preparations were reminiscent of those seen in histologic sections of the corresponding lymph node excisional biopsies. CONCLUSIONS: though ancillary investigative methods, including flow cytometry and molecular study, are of limited value in fine-needle aspiration cytology assessment of AILD-T due to the heterogeneous cell population present, recognition of the peculiar combination of cytologic features, especially in the right clinical setting, should provide a clue about the diagnosis. A high index of suspicion is essential to avoid a false negative diagnosis of reactive lymphadenopathy. PMID- 12115307 TI - Real-time, high-definition, three-dimensional microscopy for evaluating problematic cervical Papanicolaou smears classified as atypical squamous cells of undetermined significance. AB - BACKGROUND: The perceived inadequacies of the cervical Papanicolaou (Pap) smear have been attributed to sampling, screening, or interpretive errors. Within this type of cytologic preparation, there are thick cell clusters in which the cells are obscured. It may not possible to evaluate these areas by conventional microscopy. The authors clinically tested the hypothesis that high-definition, three-dimensional (3-D) microscopy based on multiple oblique illumination (MOI), with its ability to penetrate into thick areas, would be useful in evaluating problematic cervical Pap smears, particularly those diagnosed as atypical squamous cells of undetermined significance (ASCUS). METHODS: ASCUS Pap smears and corresponding surgical biopsy specimens were evaluated prospectively using standard, axially illuminated microscopes and a new high-definition, 3-D microscope employing MOI. The Pap smears were reviewed in a blinded fashion with both types of microscopy. The rendered diagnoses were then compared with the subsequent tissue biopsies, which also were blinded, as the definitive end point. RESULTS: It was immediately apparent that the high-definition, 3-D MOI microscope had better resolution compared with the standard microscopes. Pap smears and biopsy diagnoses were correlated significantly for MOI (P < 0.001), and there were significant improvements (P = 0.0108) in accuracy when 3-D, high-definition microscopy was compared with conventional microscopy. The authors found no statistically significant correlation between ASCUS diagnoses that were rendered by using standard microscopes compared with the subsequent biopsy. CONCLUSIONS: Due to enhanced visualization through thick cell clusters, an increased depth of field, light penetration, and resolution, high-definition, 3-D microscopy based on MOI produced superior accuracy compared with conventional light microscopy in evaluating cervical Pap smears. PMID- 12115306 TI - Endoscopic ultrasound-guided fine-needle aspiration in 179 cases: the M. D. Anderson Cancer Center experience. AB - BACKGROUND: Recently, endoscopic ultrasound-guided fine-needle aspiration (EUS FNA) has emerged as a diagnostic adjunct for small pancreatic lesions and abdominal and mediastinal lymph node diseases. DESIGN: During a 21-month period, we performed 179 EUS-FNAs in 166 consecutive patients; these data are the subject of this study. An average of 2.6 needle passes were obtained and aspiration was performed most commonly in the pancreas (162 cases, 91%). The FNA smears were reviewed using six diagnostic categories (negative for malignancy/nondiagnostic [NND], atypia, suspicious for malignancy, benign tumor/cyst, neuroendocrine neoplasm [NEN], and carcinoma). The review diagnosis was correlated with the histologic diagnosis made on resection or surgical biopsy specimens in 70 cases. Up to 17 months of clinical follow-up were sought for the cases with a negative or inconclusive FNA diagnosis and no diagnostic tissue confirmation (81 cases). RESULTS: The review FNA diagnoses were as follows: NND (49 cases), atypia (17 cases), suspicious for malignancy (12 cases), benign tumor/cyst excluding NEN (10 cases), NEN (6 cases), carcinoma (85 cases). Follow-up methods included resection (49 cases), surgical biopsy (21 cases), repeat FNA or brushing cytology (28 cases), and clinical follow-up only (81 cases). Of the 49 NND cases, 23 (47%) had positive follow-up results (i.e., false-negative diagnosis) that were confirmed by tissue diagnosis (resection/surgical biopsy in 11 cases [48%] and repeat FNA/brushing in 12 cases [52%]). These included pancreatic/ampullary adenocarcinoma in 20 cases, esophageal squamous carcinoma in 1 case, and NEN in 2 cases. Follow-up also revealed carcinoma in all 12 suspicious cases and 13 pancreatic adenocarcinomas and 1 microcystic adenoma in 14 of the 17 atypical cases. Overall, repeat computed tomography (CT)-guided FNA samples yielded a definite diagnosis in four atypical and seven NND cases, whereas EUS-FNA results provided a definite diagnosis in three cases in which CT-guided FNA failed and in two cases in which ampullary biopsy failed. No false-positive cases were identified. The false-negative rate due to inadequate sampling was 13.2%. Sensitivity (including cases with inadequate cellularity and nondiagnostic aspirates) was 81.7% and specificity was 100%. None of the factors evaluated (lesion characteristics, aspiration site, and tumor type) significantly influenced diagnostic results. CONCLUSION: EUS-FNA is a valuable diagnostic and staging tool with high specificity and sensitivity. Negative or nondiagnostic cases on EUS-FNA require further diagnostic work for a definitive diagnosis when clinical or radiographic findings do not correlate with the FNA results. PMID- 12115308 TI - mRNA detection of tumor-rejection genes BAGE, GAGE, and MAGE in peritoneal fluid from patients with ovarian carcinoma as a potential diagnostic tool. AB - BACKGROUND: It has been found that the members of the BAGE, MAGE, and GAGE gene families are expressed almost exclusively in neoplastic tissues. Normal tissues, except testis and placenta, are negative. Therefore, the expression of these genes may serve as a useful diagnostic marker in detecting malignant disease. The involvement of the serous cavities by malignant neoplasms has important therapeutic and prognostic implications. Accordingly, the diagnosis of peritoneal spread of ovarian carcinoma plays an important role for both initial and second look staging procedures. In some patients, however, a definite diagnosis cannot be established by morphologic or immunocytologic examination alone. Detection of tumor specific gene expression may be a sensitive additional tool in these settings. METHODS: The authors studied the gene expression observed in 44 ascites specimens. Gene expression was evaluated by reverse transcription-polymerase chain reaction analysis and sequencing. RESULTS: Of 44 ascites specimens, the expression of BAGE, MAGE-1, MAGE-3, and GAGE-1/2 was recognized in 17 specimens (63%), 2 specimens (7%), 8 specimens (30%), and 8 specimens (30%) with histologically proven ovarian carcinoma, respectively. Expression of the MAGE and GAGE genes was not observed in patients with nonneoplastic disease, whereas BAGE expression was seen in one patient with cirrhosis. CONCLUSIONS: These findings show that testing for BAGE, GAGE-1/2, MAGE-1, and MAGE-3 transcriptional activity in ascites specimens results in high sensitivity in diagnosing malignant ascites. PMID- 12115309 TI - Paget disease of the nipple: a multifocal manifestation of higher-risk disease. AB - BACKGROUND: The treatment of Paget disease by mastectomy has been challenged recently in favor of breast-conserving techniques. A large series of patients treated with mastectomy has been reviewed to assess the feasibility of less radical surgery. METHODS: The cases of 70 women with a clinical diagnosis of Paget disease were reviewed. The type, grade, receptor and node status, and the mammographic and pathologic extent of the underlying breast malignancy were determined. The survival of patients with invasive disease was compared with matched controls without Paget disease. RESULTS: The underlying malignancy was invasive in 58% of cases. Despite the fact that only one third of women presented with a palpable mass, the malignancy was frequently extensive, being confined to the retroareolar region in only 25% of cases. The true extent of the disease was underestimated by mammography in 43% of cases. Of the patients with ductal in situ carcinoma, 96.5% had high-grade carcinomas and 100% had invasive carcinomas of high cytonuclear grade. Overexpression of the c-erb-B2 oncogene was detectable in 83% of cases. Patients with Paget disease had a significantly worse survival than matched controls, but this difference was eliminated if they were also matched for c-erb-B2 status. CONCLUSIONS: Paget disease is often associated with extensive underlying malignancy, which is difficult to assess accurately either clinically or mammographically. As a consequence, cone excision of the nipple would have resulted in incomplete excision in 75% of cases. The underlying disease is of high grade and is frequently c-erb-B2 positive with a resulting poor prognosis. Aggressive local and systemic treatment would seem to be merited. PMID- 12115310 TI - Carcinoma detection at the breast examination center of Harlem. AB - BACKGROUND: Breast carcinoma is one of the leading causes of excess mortality rates in Harlem, an inner-city neighborhood with the highest mortality rates and worst life expectancy in New York City. This study reports the results of a breast carcinoma screening and diagnostic program in Harlem. METHODS: Retrospective review was performed of a database of 49,750 visits to the Breast Examination Center of Harlem from 1995 to 2000. During this period, 181 breast carcinomas were diagnosed in 178 women. The medical records of these 178 women were reviewed to determine the method of detection, stage, and treatment. RESULTS: Among these women, 89% were black or Hispanic, 45% had no medical insurance, and 38% had incomes below federal poverty guidelines. Breast carcinoma stage, known for 167 carcinomas, was Stage 0 in 38 (23%), Stage I in 38 (23%), Stage II in 63 (38%), Stage III in 24 (14%), and Stage IV in 4 (2%). Fifty-six cases (34%) were minimal breast carcinomas. Of 181 breast carcinomas, 122 (67%) were palpable and 59 (33%) were nonpalpable, detected only by mammography in asymptomatic women. Nonpalpable, as opposed to palpable, breast carcinomas were significantly more likely to be ductal carcinoma in situ (30 of 55 [54%] vs. 8 of 112 [7%], P < 0.0000001) or minimal breast carcinoma (39 of 55 [71%] vs. 17 of 112 [15%], P = 0.0000001) and were more likely to be treated with breast conserving surgery (47 of 56 [84%] vs. 76 of 110 [69%], P < 0.04). CONCLUSIONS: A breast carcinoma screening and diagnostic program has been established in Harlem, a traditionally underserved area in New York City. Early, curable breast carcinomas were detected but outreach remains a challenge, particularly for the uninsured. PMID- 12115311 TI - Sentinel lymph node biopsy in patients with ductal carcinoma in situ: a proposal. PMID- 12115312 TI - Ethnicity related differences in the survival of young breast carcinoma patients. AB - BACKGROUND: African-American women face an increased risk of early-onset breast carcinoma compared to white American women, and breast carcinoma has been reported to be particularly aggressive in premenopausal women. METHODS: Surveillance, Epidemiology, and End Results Program data were analyzed for 507 African-American and 1378 white patients from Detroit diagnosed with breast carcinoma under the age of 40 between 1990 and 1999. RESULTS: The proportion of in situ disease detected in African-American patients between 1995 and 1999 nearly doubled compared to the 1990-1994 interval (11.3% compared to 6.4%) but was consistently lower than the proportion of in situ disease seen in white patients for the same intervals (15.7% and 16.4% respectively). Evaluation of patients with invasive disease revealed that African-American patients had larger mean tumor size (3.4 cm versus 2.6 cm; P < 0.001), lower rates of localized disease (42.4% versus 52.1%; P < 0.001), higher rates of estrogen receptor negativity (61.9% versus 44.4%; P < 0.001), and higher proportions of medullary tumors (5.8% versus 3.3%; P = 0.021). Cox proportional hazards survival analysis adjusted for age, tumor size, nodal status, hormone receptor status, and histology showed higher mortality rates for African-American patients at all disease stages. Relative risk of death for African-American patients was 1.94 in patients with localized disease (95% confidence interval [CI], 1.23-3.05), 1.58 for regional disease (95% CI = 1.18-2.11), and 2.32 for distant disease (95% CI = 1.15-4.69). CONCLUSIONS: These findings show that young African-American breast carcinoma patients face an increased mortality risk. Additional studies evaluating risk and treatment response in this subset of patients are warranted. PMID- 12115313 TI - GLI3 is not mutated commonly in sporadic medulloblastomas. AB - BACKGROUND: Medulloblastoma is a malignant, invasive embryonic tumor of the cerebellum. Sonic hedgehog (SHH) is a secreted glycoprotein that has a major role in the developing cerebellum. Activation of the SHH pathway resulting from mutations in the PATCH gene, which is an inhibitor of the pathway, are associated with hereditary and sporadic medulloblastomas. The GLI3 protein is another negative regulator of SHH signaling. The authors hypothesized that mutations in GLI3 may be associated with meduloblastomas. METHODS: The authors describe a patient with hereditary Greig syndrome, which was caused by mutations in GLI3, and medulloblastoma. Another such patient was described in the literature. They also sequenced the GLI3 gene, including all exon-intron boundaries, in an additional 12 sporadic medulloblastomas. RESULTS: The authors detected a new nonsense germline mutation in a child with Greig syndrome and medulloblastoma. This mutation generates a stop codon in position 809 of GLI3 that has been predicted to result in massive truncation of the protein. Several new polymorphisms, but no tumor-associated mutations, were found in sporadic tumors. CONCLUSIONS: Gli3 is mutated rarely in medulloblastoma. PMID- 12115314 TI - Association of prediagnosis endoscopy with stage and survival in adenocarcinoma of the esophagus and gastric cardia. AB - BACKGROUND: Barrett esophagus, a consequence of chronic gastroesophageal reflux disease, is a premalignant condition for adenocarcinoma of the esophagus and, possibly, the gastric cardia. However, the actual use and clinical impact of upper gastrointestinal endoscopy in screening and surveillance for Barrett esophagus are unknown. METHODS: A cohort included 1633 patients with adenocarcinoma (777 esophagus, 856 cardia) who were 70 years or older. They were diagnosed between 1993 and 1996 and were identified from the Surveillance, Epidemiology and End Results program registry. All claims for upper endoscopy and a diagnosis of Barrett esophagus from 1991 through 1 year before diagnosis were identified from linked Medicare files. RESULTS: One or more upper endoscopies before diagnosis were performed in 9.7% of patients (13.0% esophagus, 6.8% cardia) and a diagnosis of Barrett esophagus was present in only 3.7% of patients. A shift toward earlier stage at diagnosis was observed in patients with previous endoscopy or Barrett diagnosis. For example, 62% of patients with esophageal and 49% of patients with cardia tumors who underwent previous endoscopy presented with in situ or local stage carcinoma, compared with 35% and 27% of other patients, respectively. Receipt of endoscopy was also associated with a reduced risk of death for esophageal adenocarcinoma (relative hazard 0.73, 95% confidence interval 0.57-0.93; P = 0.01), but not for adenocarcinoma of the cardia. CONCLUSIONS: Receipt of upper endoscopy at least 1 year before diagnosis of adenocarcinoma, which may reflect prediagnosis screening, was associated with an earlier tumor stage and improved survival. These data support the role of endoscopic screening and surveillance for Barrett esophagus and highlight the underdiagnosis of populations at risk. PMID- 12115315 TI - Socioeconomic and demographic disparities in treatment for carcinomas of the colon and rectum. AB - BACKGROUND: The current study examined the relationship between socioeconomic and demographic factors and type of treatment for carcinomas of the colon and rectum. The National Institutes of Health and the National Cancer Institute recommend surgery followed by adjuvant chemo- and/or radiotherapy for Stage III colon and Stages II and III rectal carcinomas. METHODS: The authors linked Washington State's cancer registry and hospital discharge records and U.S. census data to assess socioeconomic and demographic factors related to treatment, controlling for clinical factors. RESULTS: Compared to colon carcinoma patients under age 65 years, patients aged 75-84 years and 85 years or older were at higher risk for a treatment plan of surgery without adjuvant therapy (adjusted odds ratio [OR] = 2.5, 95% confidence interval [CI] = 1.3-4.7; OR = 14.1, CI = 6.3-31.4, respectively). Risk of no adjuvant therapy was more than doubled for patients in zip codes in the lowest quartile of per capita income compared to the top three quartiles (OR = 2.3, CI = 1.5-3.4) and for those with Medicare compared to private insurance (OR = 2.2, CI = 1.3-3.8). Older patients with rectal carcinoma were also at higher risk of a treatment plan that did not include adjuvant therapy. CONCLUSIONS: The current findings suggest disparities in the provision of recommended medical procedures related to socioeconomic and demographic factors. PMID- 12115316 TI - Vascular endothelial growth factor overexpression is correlated with von Hippel Lindau tumor suppressor gene inactivation in patients with sporadic renal cell carcinoma. AB - BACKGROUND: A high frequency of genetic alterations of the von Hippel-Lindau (VHL) gene and overexpression of the vascular endothelial growth factor (VEGF) gene have been observed independently in human sporadic renal cell carcinoma (RCCs), but to the authors' knowledge the association between the two has not been characterized in primary sporadic RCC. In the current study the authors report the simultaneous comparison of the biallelic inactivation status of the VHL gene and VEGF expression levels in patients with sporadic RCC. METHODS: DNA and RNA were extracted from 27 sporadic RCC samples. Mutation was analyzed by direct sequencing of the amplified VHL DNA and cDNA. Loss of heterozygosity (LOH) of the gene was analyzed at three polymorphic markers. The VEGF mRNA was measured using Northern blot analysis. RESULTS: Mutations of the VHL gene were found in 14 of 27 RCC samples (51.9%). LOH analysis by a VHL-intragenic polymorphic marker and 2 extragenic microsatellite markers, D3S1560 and D3S1317, showed that LOH occurred in 10 of 15 RCC samples (66.7%). Overexpression of VEGF mRNA was observed in 17 of 27 RCC cases (63.0%): 15 of the 18 RCC samples estimated to have at least 1 hit, but only 2 of the 6 RCC samples with 0-1 hit, and none of the 3 RCC samples in which the VHL gene was not inactivated. CONCLUSIONS: VEGF overexpression was found to be correlated with both monoallelic and biallelic VHL inactivation. Alteration of the VHL gene is believed to cause angiogenesis in RCC cases through the overexpression of VEGF. PMID- 12115317 TI - Mental health in men treated for early stage prostate carcinoma: a posttreatment, longitudinal quality of life analysis from the Cancer of the Prostate Strategic Urologic Research Endeavor. AB - BACKGROUND: The current study was conducted to assess posttreatment changes in the mental components of health related quality of life in prostate carcinoma patients during the two years following diagnosis and management with radical prostatectomy, pelvic irradiation, or watchful waiting. METHODS: The authors studied the mental domains of general health related quality of life in 452 men recently diagnosed with early stage prostate carcinoma and treated with radical prostatectomy, pelvic radiation, or watchful waiting. Outcomes were assessed with the RAND 36-Item Health Survey, a validated health-related quality of life instrument that includes four mental domains. To minimize the influence of potentially confounding factors, the authors adjusted for age, comorbidity, prostate specific antigen (PSA) at diagnosis, and biopsy Gleason score. All subjects were drawn from CaPSURE, a national, longitudinal cohort. RESULTS: By 6 12 months after treatment, the active treatment groups began to show differences in mental health and vitality. By 15 months, surgery and radiation patients scored differently in all four mental domains. Over time, the gaps between mental domain scores grew wider among the treatment groups, with surgery patients performing the best, radiation patients performing the worst, and watchful waiting patients falling in between. CONCLUSIONS: The mental health profiles differ for patients undergoing surgery, radiation, or watchful waiting for early stage prostate carcinoma. Men with more serious disease, as evidenced by higher PSA levels or more aggressive histology, tended to worry more about it. Older men performed better, while sicker men performed worse, even though the older men tended to be sicker. PMID- 12115319 TI - Fas/Fas ligand system and apoptosis induction in testicular carcinoma. AB - BACKGROUND: Tumor-infiltrating, Fas ligand (FasL)-expressing lymphocytes are able to eliminate Fas-bearing tumor cells by apoptosis induction. Activated cytotoxic T-cells that express Fas may enter apoptosis in the presence of FasL tumor cells. To date, no studies of patients with testicular carcinoma have correlated the differential expression of Fas and FasL in both cell types with the corresponding apoptotic index (AI). METHODS: Fas and FasL were investigated immunohistochemically in paraffin embedded tissue sections from 25 patients with nonseminomatous testicular tumors. The percentages of positive cells and the ratios of Fas cells to FasL cells were correlated with the AI of tumor cells and lymphocytes, respectively, using Spearman correlations. RESULTS: No association was found between the rate of FasL positive cells and AI of the other cell type or between the rate of Fas positive cells and the AI of the same cell type. Ratios between Fas positive cells and FasL positive cells were not correlated with the AI; however, a significant positive correlation was found between the AI of tumor cells and the AI of lymphocytes. CONCLUSIONS: It seems unlikely that the Fas/FasL system is responsible for immune escape of the tumor in testicular carcinoma. Rather, the significant positive correlation between the AIs of tumor cells and lymphocytes implicate a previously unknown mechanism of apoptosis induction in both cell types. PMID- 12115318 TI - Bladder tumor markers for monitoring recurrence and screening comparison of hyaluronic acid-hyaluronidase and BTA-Stat tests. AB - BACKGROUND: One of the goals of a noninvasive test for bladder carcinoma screening would be to reduce surveillance cystoscopies among patients with a history of bladder carcinoma. In addition, an accurate bladder carcinoma marker could be used to screen a high-risk population. The authors examined the efficacy of the hyaluronic acid-hyaluronidase (HA-HAase) and BTA-Stat tests to detect and predict bladder carcinoma recurrence and tested their specificity for bladder carcinoma screening. METHODS: Over a four year period, the authors prospectively collected 225 urine specimens from 70 bladder carcinoma patients and analyzed them by the HA-HAase test. Tumors were identified during 178 visits, and in 47 specimens there was no evidence of disease (NED). Twenty six of these 70 patients were randomly selected to have the BTA-Stat test (111 surveillance visits). In a separate study, 401 former Department of Energy (DOE) workers, who are likely to be at a higher risk for bladder carcinoma, were screened by the HA-HAase and BTA Stat urine tests. RESULTS: The HA-HAase test had an approximately 91.0% sensitivity, 70% specificity, 87% accuracy, 92% positive predictive value (PPV), and 67% negative predictive value (NPV) in the 70 bladder carcinoma patients. There were 14 false-positives; however, 6 of these had recurred in approximately 5 months. Only 4 out of 33 NED cases recurred in that time period (chi-square = 5.43; degrees of freedom [DF] = 1; P = 0.0198). Thus, a false-positive HA-HAase test carried a significant risk of recurrence within five months (relative risk [RR] = 3.5; odds ratio [OR] = 5.44). In a direct comparison, the HA-HAase and BTA Stat had 94% and 61% sensitivity, 63% and 74% specificity, 87% and 64% accuracy, 89% and 88% PPV, and 77% and 38% NPV, respectively. While 6 of the 10 false positive on the HA-HAase test recurred in 5 months (chi-square = 9.6; DF = 1; P = 0.004), only 1 of the 7 false-positives on the BTA-Stat test recurred in that time period (chi-square = 0.096; DF = 1; P = 0.756). The RR and OR for the HA HAase test were 10.2 and 24, and for the BTA-Stat, 1.4 and 1.5, respectively. In the DOE worker screening study, the HA-HAase and BTA-Stat had 14% (56 out of 401) and 16.7% (67 out of 401) positive rates, respectively. Sixty three percent of the positives on the BTA-Stat test, but only 25% of the positives on the HA-HAase test, had benign urologic conditions. None of the biomarker positive cases with clinical follow-up (n = 29) had evidence of bladder carcinoma. CONCLUSIONS: The HA-HAase test is efficient and superior to the BTA-Stat for detecting and predicting bladder carcinoma recurrence. Noninvasive tests with low false positive rates could be used for bladder carcinoma screening in high-risk populations (e.g., those with occupational exposure to carcinogens or smokers). PMID- 12115320 TI - Complications from treatment for prostate carcinoma among men in the Detroit area. AB - BACKGROUND: Aggressive treatment of early stage prostate carcinoma (PC) is limited primarily to two modalities: radical prostatectomy (RP) and external beam radiation therapy (RT). The authors conducted a population-based study of Detroit area men with localized PC to determine the outcome of bowel, urinary, and sexual function after aggressive treatment. METHODS: Men with PC were identified through the Metropolitan Detroit Cancer Surveillance System, a member of the National Cancer Institute Surveillance, Epidemiology, and End Results Program. Patients participated in interviews about their pretreatment bowel, urinary, and sexual function approximately 9 months after treatment. The same men were asked identical questions about their function an average of 2 years after treatment. Treatment outcomes were compared for men who underwent RP and men who received RT. RESULTS: Of 501 men, 398 (79.4%) participated in both interviews, 304 of whom (76.4%) had localized PC and had been treated at least 1 year previously (median, 688 days). One hundred thirty men underwent RP, and 115 men received RT. The proportion of men in the RP group who reported an increase in incontinence symptoms was significant (53.8% compared with 19.2% in the RT group; P < 0.001). Men in the RT group reported increased loose stools between the pretreatment and post-treatment interviews (5.2% vs. 29.6%; P < 0.001). Men in both the RT group and the RP group reported increases in impotence from 40% to > 75% (P < 0.001 for both). Men in the RT group were 3.6 times more likely to have bowel incontinence compared with men in the RP group (odds ratio [OR], 3.61; 95% confidence interval [95% CI], 1.54-8.47). Urinary incontinence (OR, 2.87; 95% CI, 1.52-5.44) and erection difficulty (OR, 3.98; 95% CI, 1.35-11.70) were more likely among men in the RP group. CONCLUSIONS: Although patients may have recalled their baseline function as better than it was, the current results are consistent with other population-based studies of treatment outcomes among men with localized PC. They indicate that the side effects associated with treatment are greater than those based on case series. Physicians and patients should be aware of these population based outcomes and should use them as part of the decision-making process regarding the treatment options for men with PC. PMID- 12115321 TI - Improved survival with perilymphatic interleukin 2 in patients with resectable squamous cell carcinoma of the oral cavity and oropharynx. AB - BACKGROUND: The current randomized, multicenter, Phase III trial was conducted to determine whether the disease free interval and overall survival of patients with T2-T4,N0-N3,M0 squamous cell carcinoma (SCC) of the oral cavity or oropharynx could be extended through the combination of surgery (and radiotherapy, if required) with perilymphatic recombinant IL-2 (rIL-2). METHODS: Patients with a resectable T2-T4,N0-N3,M0 SCC of the oral cavity and oropharynx were assigned randomly to receive surgery and radiotherapy or to receive IL-2, surgery, and radiotherapy. Five thousand units of rIL-2 were injected around the ipsilateral cervical lymph node chain daily for 10 days before surgery. After surgery, contralateral 5-day rIL-2 courses were administered monthly for 1 year. The differences in disease free and overall survival between the two groups of patients were evaluated statistically. RESULTS: Two hundred two patients finished the study. No significant complications related to rIL-2 were encountered, and surgery and radiotherapy were not hampered by its prior administration. Multivariate analysis conducted to determine the extent to which survival was influenced by rIL-2 and the other variables showed that rIL-2 significantly lengthened disease free survival (P < 0.01) and that this resulted in longer overall survival (P < 0.03). CONCLUSIONS: The data emerging from this trial indicate that perilymphatic administration of low, nontoxic doses of rIL-2 is a simple and manageable way to delay recurrences of SCC. PMID- 12115322 TI - An exploration of the pretreatment coping strategies of patients with carcinoma of the head and neck. AB - BACKGROUND: Patients with head and neck carcinoma (HNC) often face exhaustive and debilitating treatment as well as physical and functional residual effects, such as disfigurement, compromised speech, dry mouth, and difficulties swallowing. Understanding how patients cope with these challenges is important in comprehensive care of patients with HNC. METHODS: Seventy-nine patients with HNC were assessed for quality of life (QOL) and coping strategy. Measures included the Functional Assessment of Cancer Therapy-Head and Neck, the Performance Status Scale for Head and Neck Cancer Patients, and the Ways of Coping-Cancer Version. Coping strategies were summarized and related to patient demographics and QOL. RESULTS: The results suggested that patients with HNC used a wide range of coping strategies, with social support seeking behaviors representing the greatest proportion of total coping effort (25%). The use of avoidant coping strategies (both cognitive and behavioral escape) was associated with poorer overall QOL. CONCLUSIONS: Although further examination of these issues in larger groups of patients with HNC is warranted, the current findings suggest the adaptability of this group of patients and the potential benefit of social support-based assistance or intervention. PMID- 12115323 TI - Prognostic significance of intrahepatic lymphatic invasion in patients with hepatic resection due to metastases from colorectal carcinoma. AB - BACKGROUND: Intrahepatic spread from liver metastases of colorectal carcinoma has been well described; however, its prognostic value after hepatectomy is controversial. To clearly determine factors predicting survival after hepatectomy in such patients, the authors evaluated 14 clinicopathologic factors of liver metastasis from colorectal carcinoma with special reference to intrahepatic lymphatic invasion. METHODS: The authors retrospectively analyzed data obtained from 67 consecutive patients who underwent hepatectomy for liver metastasis from colorectal carcinoma. Intrahepatic spread was classified into discreet categories that were evaluated separately: invasion to the portal vein, hepatic vein, bile duct, and lymphatic or perineural space. Overall survival and disease free survival periods were examined as functions of clinicopathologic determinants by univariate and multivariate analyses. RESULTS: Intrahepatic spread was found in a total of 28 (43.1%) of the 65 evaluable cases. Portal vein invasion was found in 15 (23.1%) of these cases, hepatic vein invasion in 3 (4.6%), bile duct invasion in 10 (15.4%), and intrahepatic lymphatic invasion in 10 (15.4%). Five year overall and disease free survival rates after hepatectomy were 33.4% and 28.5%, respectively. A short interval (< 12 months) from treatment of primary colorectal carcinoma to liver metastasis and the presence of intrahepatic lymphatic invasion significantly and adversely affected the overall and disease free survival rates. CONCLUSIONS: Intrahepatic lymphatic invasion was shown statistically to be an independent predictor of recurrence and death after hepatectomy in patients with liver metastases from primary colorectal carcinoma. PMID- 12115325 TI - The clinical course of nonsmall cell lung carcinoma in survivors of Hodgkin disease. AB - BACKGROUND: The objective of this study was to document the natural history of second lung carcinomas, common second tumors that arise in survivors of Hodgkin disease (HD). METHODS: The data bases of the Memorial Sloan-Kettering Cancer Center were searched to retrieve those patients who were listed with a diagnosis of both lung carcinoma and HD. Information was extracted regarding their HD (including age at diagnosis and treatment received) and their lung carcinoma (including smoking history, latency from HD, histology, disease stage, treatment received, treatment response, and survival). RESULTS: Twenty-one lung carcinomas were diagnosed in 19 patients, with a median latency of 13 years from the time of diagnosis of HD. Only five patients underwent complete resection, and four patients were alive and disease free at the last follow-up. In contrast, the median survival of 14 patients with unresectable disease was 3 months. No major objective responses were documented after chemotherapy. Poor performance status and prior thoracic radiotherapy limited treatment in patients with advanced disease. All patients had either received radiotherapy to the chest for HD or had a history of smoking; 74% of patients had both risk factors for the development of lung carcinoma. CONCLUSIONS: In patients with a history of HD, survival after the development of lung carcinoma is poor. Because surgical resection can lead to long-term survival, early detection is crucial. HD survivors, especially those with a history of smoking, should undergo careful surveillance for second primary lung carcinomas and other diseases. Patients who are diagnosed with HD should abstain from smoking. Physicians should assess specifically the smoking status of all HD patients and prescribe a smoking cessation program. PMID- 12115324 TI - Serum tissue polypeptide specific antigen as a noninvasive prognostic indicator for early recurrence of hepatocellular carcinoma after curative resection. AB - BACKGROUND: Serum tissue polypeptide specific antigen (TPS) is a useful cell proliferation marker in diagnosing and monitoring patients with a variety of malignancies. The objective of this study was to determine the usefulness of serum TPS as a noninvasive prognostic factor for early recurrence of hepatocellular carcinoma (HCC) after patients undergo curative resection. METHODS: Serum TPS levels were measured by monoclonal TPS immunoradiometric assay in 54 patients shortly before they underwent curative resection for HCC. The recurrence time was correlated with the TPS level and with other prognostic factors using the log-rank test in univariate analysis and a Cox regression in multivariate analysis. Receiver operating characteristic analysis was performed to examine the power of the various prognostic factors to distinguish between patients with recurrent tumors and patients who were disease free. RESULTS: Patients who had higher levels of TPS (>or= 150 U/L) had earlier recurrences compared with patients who had lower levels of TPS (< 150 U/L; P = 0.016) in univariate analysis. Tumor size, the number of tumors, portal vein invasion, and the resection margins also were associated significantly with the time to tumor recurrence (P = 0.015, P = 0.004, P = 0.003, and P = 0.003, respectively). Serum alpha-fetoprotein was not a significant risk factor for tumor recurrence. In multivariate analysis, the TPS level, tumor size, and resection margins were independent prognostic factors (P = 0.025, P = 0.018, and P = 0.016, respectively). The inclusion of TPS in addition to tumor size and resection margins increased the rate of corrective prediction from 0.72 to 0.80. CONCLUSIONS: The current study demonstrated that the preoperative serum TPS level was a significant factor in predicting early recurrence of HCC after curative resection. Patients with high serum TPS levels warrant more aggressive treatment and close follow-up after they undergo tumor resection. PMID- 12115326 TI - Phase I trial of subcutaneous recombinant human interleukin-2 in patients with metastatic melanoma. AB - BACKGROUND: Interleukin-2 (IL-2) has activity in metastatic melanoma when given in high doses by the intravenous (IV) route, but its side effects and effectiveness when given in intermediate to high doses by the subcutaneous (SC) route have not been studied adequately. This study sought to determine the maximum tolerated dose (MTD) of IL-2 administered once daily by the SC route. METHODS: Outpatients with progressive metastatic melanoma after chemotherapy were enrolled in a Phase I trial of IL-2 administered SC daily for 5 days per week for 4 consecutive weeks, repeated at 6-week intervals. Patients were instructed to drink at least 2 L of fluid daily. IL-2 pharmacokinetic studies were performed at the two highest dose levels. Toxicity was recorded weekly using the National Cancer Institute Common Toxicity Criteria. Response was assessed at 6-week intervals. RESULTS: Three patients, 6 patients, 6 patients, and 4 patients received a median of 2 courses of SC IL-2 at dose levels of 6 MIU/m(2), 9 MIU/m(2), 12 MIU/m(2), and 15 MIU/m(2), respectively. Failure to maintain adequate fluid intake was responsible for 2 episodes of syncope at the 9 MIU/m(2) dose level and for 2 incidents of reversible prerenal azotemia at the 15 MIU/m(2) dose level. IL-2 treatment was resumed in these patients without incident. At the 15 MIU/m(2) dose level, 2 patients had severe headaches, depression, and visual hallucinations requiring discontinuation of treatment. Cough and fluid retention at the end of the third and fourth weeks at the 15 MIU/m(2) dose level approximated the symptoms reported by inpatients treated by continuous IV infusion at 9 MIU/m(2) on the same schedule. There was a partial response and a complete response in subcutaneous disease at the 12 MIU/m(2) and 15 MIU/m(2) dose levels, respectively, each lasting < 2 months. Plasma IL-2 levels after SC injection of 1000-5000 pg/mL reached maximum by 3 hours and were detectable for up to 48 hours after administration. The half-lives for SC IL-2 absorbance and clearance were 1.6 hours and 5.2 hours, respectively, and the calculated area under the curve was 30,584 pg/mL x hour. CONCLUSIONS: SC IL-2 was well tolerated and had high sustained bioavailability at the higher doses studied. The MTD for a daily SC regimen was 12 MIU/m(2) and is recommended for future studies. PMID- 12115327 TI - Predictors of skin self-examination performance. AB - BACKGROUND: Skin self-examination (SSE) may reduce the death rate from melanoma by as much as 63%. Enhancing SSE performance may reduce mortality and morbidity. This study determined predictors of SSE performance in a population of individuals who were at risk of developing melanoma or nonmelanoma skin carcinoma (NMSC). METHODS: Patients (n = 200) were asked about their knowledge of the warning signs, their sense of the importance of SSE to them, their attitude about and confidence in their ability to perform SSE, and their impression of their partner's comfort and ability with assisting in the skin examination. The interval since last skin examination, the number of physician visits (nondermatologist and dermatologist), the number and type of skin malignancies, the time since initial diagnosis, the number of skin biopsies, and health insurance status were determined from the medical records for the prior 3 years. RESULTS: Seventy percent of participants performed SSE. The three strongest predictors of SSE performance were attitude, having dermatology visits with skin biopsies and at least one skin carcinoma in the previous 3 years, and confidence in performance (P = 0.0001). Other predictors of SSE performance were perceived risk (P = 0.0001), knowledge (P = 0.004), and younger age (P = 0.045). CONCLUSIONS: Annual skin examination by physicians and monthly SSE by patients reinforce one another in promoting early detection. In this high-risk population, the dermatologist reinforced SSE performance by biopsy of skin lesions that were skin malignancies. People have intimate knowledge of their own skin and bear the consequences for failure to detect and treat skin carcinoma early; thus, monthly SSE becomes relevant as a personal health-promotion habit. PMID- 12115328 TI - Multicenter trial of low-dose paclitaxel in patients with advanced AIDS-related Kaposi sarcoma. AB - BACKGROUND: Treatment options are limited for patients with advanced acquired immunodeficiency syndrome (AIDS)-related Kaposi sarcoma (AIDS-KS) whose disease has progressed after receiving therapy with liposomal anthracyclines or combination chemotherapy with doxorubicin (Adriamycin), bleomycin, and vincristine (ABV). This study was performed to assess the safety and efficacy of a novel dose and schedule of paclitaxel in patients with AIDS-KS who failed to respond to previous systemic chemotherapy. METHODS: This was an open-label, multicenter Phase II study. Eligible patients had advanced AIDS-KS consisting of at least 25 mucocutaneous lesions, visceral disease, or lymphedema, and had failed to respond to at least one previous systemic chemotherapy regimen. Patients were treated with paclitaxel at a dose of 100 mg/m(2) given intravenously over 3 hours, every 2 weeks. Primary efficacy end points were tumor response, time to progression, time to treatment failure, and survival. Quality of life and adverse events were evaluated using the Symptom Distress Scale (SDS) and the World Health Organization Toxicity Criteria, respectively. RESULTS: One hundred and seven male patients with advanced AIDS-KS were enrolled from nine participating sites. The median entry CD4+ lymphocyte count was 41/mm(3) (range 0 1139). Previous treatment regimens included ABV in 52, liposomal daunorubicin in 49, and liposomal doxorubicin in 40 patients. Forty-one patients (38%) received two or more previous chemotherapy regimens. Protease inhibitor use during the study was reported by 82 (77%) patients overall; 47 patients (44%) were receiving a protease inhibitor before study entry. Complete or partial response was documented in 60 patients (56%). The median duration of response was 8.9 months. Major response rate was similar when comparing patients not on a protease inhibitor at the time of response (59%) with patients on a protease inhibitor at time of response (54%). However, protease inhibitor use had a significant impact on survival (P = 0.04). Grade 4 neutropenia was reported in 35% of patients; other life-threatening side effects were uncommon. Significant improvements were seen in the total quality of life scores measured by the SDS, including significant improvement in KS-related symptoms such as facial disease, tumor associated edema, and pulmonary involvement. CONCLUSION: Paclitaxel given every 2 weeks induces major tumor regression in the majority of patients with advanced KS who failed to respond to previous systemic chemotherapy. Paclitaxel is associated with significant improvement in quality of life with acceptable toxicity and should be considered as an effective treatment option for patients with advanced KS. PMID- 12115329 TI - Changing patient perceptions of the side effects of cancer chemotherapy. AB - BACKGROUND: Quality-of-life (QoL) issues have become increasingly important as the number of newly diagnosed patients with cancer increases and survival improves. In 1983, Coates et al. reported a survey of patient perceptions of the side effects of cancer chemotherapy and showed the importance of including patient feedback for the accurate assessment of QoL (Eur J Cancer Clin Oncol. 1983;19:203-208.). The authors carried out a similar survey in 100 patients with cancer with the objectives of 1) investigating the changes in patient perceptions that have occurred and 2) evaluating the impact of new treatments on the profile of chemotherapy side effects among patients receiving anticancer drugs. METHODS: One hundred patients attending the outpatient Medical Oncology Department of the Pitie Salpetriere Hospital Group were surveyed between August 1998 and February 2000 by trained interviewers who were blinded to the patients' treatment. Patients identified all side effects associated with their treatment using a set of 45 cards that named physical side effects (Group A) and a set of 27 cards that named nonphysical side effects (Group B), and the patients ranked these side effects according to severity. The top 5 cards from each group were then combined, and the resulting 10 cards were rated again by severity, regardless of group. Results were analyzed for the entire cohort and for demographic, social, and clinical subgroups. RESULTS: The participants included 65 women and 35 men; the most common malignancies were breast carcinoma (40 patients), gastrointestinal carcinoma (19 patients), lung carcinoma (7 patients), and ovarian carcinoma (9 patients). Patients rated affects my family or partner as the most severe side effect, alopecia was second, and fatigue was the third most severe. Effects on work or home responsibilities, effects on social activities, and loss of interest in sex were ranked fourth, fifth, and sixth, respectively. The results contrasted with those of Coates et al., in which affects my family or partner was ranked 10th, and fatigue was ranked 8th. CONCLUSIONS: Patient perceptions of the side effects of cancer chemotherapy have changed markedly. In the current study, fatigue and psychosocial QoL concerns predominated, compared with emesis, nausea, and negative reactions to the treatment visit in the original survey. The current findings are consistent with the progress that has been made in reducing certain chemotherapy-associated toxicities. Fatigue, however, although it often is related to anemia and is treatable with recombinant human erythropoietin, remains a major concern. The emotional, social, and sexual consequences of cancer treatment present continuing challenges in efforts to optimize QoL and to develop effective supportive care. PMID- 12115330 TI - Patterns of Care Study quantitative evaluation of the quality of radiotherapy in Japan. AB - BACKGROUND: Quality assurance (QA) of clinical practice is important for any medical specialty. Programs based on the Patterns of Care Study (PCS) have been developed to compare the quality of radiotherapeutic care at individual institutions, with the national average representing the process and outcome of radiotherapy. The feasibility of these programs was analyzed. METHODS: Calculation programs for the national average and standard score were developed to evaluate quantitatively the process and outcome of radiotherapy at individual institutions as well as at the national level. The programs were used to evaluate the quality of radiotherapy for 561 esophageal carcinoma patients surveyed in the Japanese PCS. RESULTS: As a representative example of QA measurement, the national average for the 5-year survival rate for these patients in the nonsurgery group was 5%. The regional averages for those in academic and nonacademic institutions were 9% and 1%, respectively (P = 0.0142), showing a significant difference between these two institutional strata. The standard score compared with the national average for institution No.105, for example, was 16.3 (P < 0.0001), with the positive value indicating that the outcome at this institution was significantly higher than the national average. The corresponding figure compared with the regional average was -0.3 (P = 0.7391), with the negative value indicating the outcome is not superior to the regional average of academic institutions. CONCLUSIONS: These programs make it possible to compare quantitatively the quality of radiation therapy at individual institutions with the national and regional averages. They should also be useful for nationwide QA projects in radiation oncology as well as in other medical specialities. PMID- 12115331 TI - Pretreatment prognostic factors and treatment results in children with hepatoblastoma: a report from the German Cooperative Pediatric Liver Tumor Study HB 94. AB - BACKGROUND: In the past 20 years, a dramatic improvement in the prognosis of patients with hepatoblastoma (HB) has been achieved by combining surgery with chemotherapy in several national and international trials. A worldwide, unsolved problem remains the treatment of patients with advanced or metastatic HB. METHODS: The German Cooperative Pediatric Liver Tumor Study HB 94 was a prospective, multicenter, single-arm study. The study ran from January 1994 to December 1998. The protocol assessed the efficiency of chemotherapy consisting of cisplatin, ifosfamide, and doxorubicin (CDDP/IFO/DOXO) and/or etoposide and carboplatin (VP16/CARBO). The prognostic significance of the surgical strategy, pretreatment factors, and tumor characteristics for disease free survival (DFS) were analyzed. RESULTS: Sixty-nine children with HB were treated in the HB 94 study. The median follow-up of survivors was 58 months (range, 32-93 months). Fifty-three of 69 patients (77%) remained alive, and 16 of 69 patients (23%) died. Long-term DFS was as follows: 26 of 27 patients had Stage I HB, 3 of 3 patients had Stage II HB, 19 of 25 patients had Stage III HB, and 5 of 14 patients had Stage IV. A complete resection of the primary tumor was achieved in 54 of 63 patients (86%). Six children (8%) had no surgical treatment. Twenty-two tumors were resected primarily, and 41 children underwent surgery after initial chemotherapy. Two children underwent liver transplantation. There was no perioperative death. Forty-eight children received primary chemotherapy with CDDP/IFO/DOXO. Forty-one of 48 children achieved partial remission after CDDP/IFO/DOXO. Eighteen children with advanced or recurrent HB underwent VP16/CARBO chemotherapy, with a response achieved by 12 children. The relevant pretreatment prognostic factors were growth pattern of the liver tumor (P = 0.0135), vascular tumor invasion (P = 0.0039), occurrence of distant metastases (P = 0.0001), initial alpha-fetoprotein level (P = 0.0034), and surgical radicality (P < 0.0001). CONCLUSIONS: The current results underline the necessity of preoperative chemotherapy in all children with HB. Complete tumor resection is one of the main prognostic factors. PMID- 12115332 TI - Cognitive functioning and quality of life in long-term adult survivors of bone marrow transplantation. AB - BACKGROUND: The late neurotoxic effects of bone marrow transplantation (BMT) on cognitive functioning and quality of life (QOL) were investigated in a consecutively treated cohort of long-term adult survivors. METHODS: Progression free patients treated with BMT or peripheral stem cell grafts for a hematologic malignancy at least 2 years before study participation were examined with a comprehensive battery of neuropsychological tests and questionnaires for QOL and mood states. The results of the neuropsychological tests were compared with healthy population norms. RESULTS: Forty patients were included, 87.5% of whom had undergone an allogeneic transplantation. All received total body irradiation up to 12 Gy (in two fractions). Assessment took place 22-82 months after BMT. Mild to moderate cognitive impairment was found in 24 patients (60%). Compared with healthy population norms, selective attention and executive function, information processing speed, verbal learning, and verbal and visual memory were most likely to be affected. The mean score for the total patient group revealed that these patients scored significantly lower on the information processing speed task compared with expected scores obtained from the normal population. The main predictors for poor neuropsychological performance were fatigue, global health, and educational level. Other correlations with moderate to severe cognitive impairment were subjective cognitive complaints, physical functioning, social functioning, overall mood states, and employment status. CONCLUSIONS: These data indicate that BMT may lead to cognitive complaints and late cognitive deficits in long-term adult survivors. Cognitive functioning should therefore be used as an outcome parameter in BMT studies. PMID- 12115333 TI - Decreasing incidence rates of primary central nervous system lymphoma. AB - BACKGROUND: Incidence rates of primary central nervous system lymphomas (PCNSL) in the U.S. were reported to have increased dramatically during the 1980s and early 1990s. Recent reports portray a continuation of this trend. With potential etiologic factors related to immunosuppression changing, the authors hypothesized that the incidence of PCNSL would be stabilizing. METHODS: The authors analyzed age specific, gender specific, and race specific PCNSL incidence rates from 1973 1998 using data from the Surveillance, Epidemiology, and End Results (SEER) program. To estimate the impact of the acquired immunodeficiency syndrome (AIDS) and organ transplantation on PCNSL trends, the authors evaluated incidence data from the Centers for Disease Control and Prevention HIV [human immunodeficiency virus]/AIDS Surveillance Report, and the United Network for Organ Sharing. RESULTSPCNSL incidence rates decreased from a peak of 10.2 per 1 million person years in 1995 to 5.1 per 1 million person-years in 1998, a decrease that was largely attributable to a decrease in the incidence of the disease in males age or= 60 years essentially has remained unchanged since 1994, at approximately 16 per 1 million person-years. Since the early 1980s, regardless of age, PCNSL incidence rates were higher in males than in females, and higher in blacks than in whites. Concordant with PCNSL trends, AIDS rates have decreased since their peak in 1993. However, the rate of solid organ transplantations has increased steadily since 1990. CONCLUSIONS: In contrast to recent reports of a progressively increasing incidence, the authors found that PCNSL rates have been decreasing in the majority of demographic groups in the U.S since the mid-1990s. A notable exception was observed in the highest PCNSL risk group, those age >or= 60 years. PMID- 12115334 TI - Rapid titration with intravenous morphine for severe cancer pain and immediate oral conversion. AB - BACKGROUND: Cancer pain emergencies presenting with severe excruciating pain require a rapid application of powerful analgesic strategies. The aim of the current study was to evaluate a method of rapid titration with intravenous morphine to achieve relief of cancer pain of severe intensity. METHODS: Forty nine consecutive patients admitted to a Pain Relief and Palliative Care Unit for severe and prolonged pain were enrolled in the study. Pain was evaluated on a numeric scale of 0-10 (0 indicated no pain and 10 indicated excruciating pain). After the initial assessment (T0), an intravenous line was inserted and boluses of morphine (2 mg every 2 minutes) were given until the initial signs of significant analgesia were detected or severe adverse effects occurred (T1). A continuous reassessment was warranted and the effective total dose administrated intravenously was assumed to last approximately 4 hours and was calculated for 24 hours. The dose immediately was converted to oral morphine (a 1:3 ratio for low doses and a 1:2 ratio for high doses). RESULTS: Data from 45 patients was analyzed. A significant decrease in pain intensity was achieved in a mean of 9.7 minutes (95% confidence interval [95% CI], 7.4-12.1 minutes), using a mean dose of intravenous morphine of 8.5 mg (95% CI, 6.5-10.5 mg). The doses administered rapidly were converted to oral morphine and pain control was maintained until the patient's discharge, which occurred in a mean of 4.6 days (95% CI, 4.1-5.2 days). The incidence of adverse effects was minimal. CONCLUSIONS: The results of the current study demonstrate that cancer pain emergencies can be treated rapidly in the majority of cancer patients with an acceptable level of adverse effects. Intravenous administration of morphine requires initial close supervision and continuity of medical and nursing care. PMID- 12115337 TI - Polymerase chain reaction-based diagnosis of bone marrow involvement in 170 cases of non-Hodgkin lymphoma. AB - BACKGROUND: Up to the current time, diagnosis of bone marrow (BM) involvement in non-Hodgkin lymphoma (NHL) has been based on morphologic findings. Polymerase chain reaction (PCR) for antigen receptor gene rearrangements has the potential to increase the detection sensitivity of minimal degrees of BM involvement. The authors therefore assessed PCR-based clonalities of BM concurrently with morphology from 170 cases with NHL and evaluated the usefulness of comparative analysis of clonalities between bilateral BMs and the lymph node and the clinical significance of PCR based clonalities of BM. METHODS: Bilateral BM clot sections of 170 cases and 47 lymph nodes were tested for immunoglobulin heavy chain gene rearrangement or T-cell receptor gamma gene rearrangement according to the B- or T-lineage of the lymph node. RESULTS: When compared with morphology, the results of PCR showed an unexpectedly low positive concordance rate of 61.0% for B-cell NHL and 57.1% for T-cell NHL. When the clonality of BM was compared with that of lymph nodes in B-cell NHL, bilateral clonalities of BM showed high concordance with the clonality of the lymph nodes. PCR-based clonality did not show significant impact on survival. CONCLUSIONS: Morphology remains the gold standard in the evaluation of BM involvement by NHL. Although the comparative analysis of BM clonality and that of the lymph nodes is considered a valuable tool that increases the reliability of clonality, PCR-based clonality of BM does not significantly add to the sensitivity of diagnosing BM involvement by NHL. PMID- 12115338 TI - International prognostic index-based outcomes for diffuse large B-cell lymphomas. AB - BACKGROUND: We present the results of doxorubicin-based chemotherapy with or without involved-field radiotherapy for patients with diffuse large B-cell lymphoma (DLBCL) according to the international prognostic index (IPI). METHODS: From September 1988 through December 1996, 294 patients with Stage I-IV Working Formulation large B-cell or T-cell lymphomas were treated prospectively on two protocols at our center. Diagnoses were reclassified subsequently according to the new World Health Organization classification. Two-hundred and twenty-four patients had DLBCL, including 24 patients with primary mediastinal large B-cell lymphoma. Treatment consisted of a median of six cycles of cyclophosphamide, doxorubicin, vincristine, and prednisone (CHOP) chemotherapy with or without involved-field radiotherapy (median dose, 39.6 Gy). RESULTS/CONCLUSIONS: The median length of follow-up among surviving patients was 5.0 years. Patient subgroups differed from each other in terms of progression-free (P = 0.003), cause-specific (P = 0.003), and overall (P = 0.001) survival rates when analyzed by IPI. Five-year progression-free, cause-specific, and overall survival rates for 212 patients with an IPI of 0-2 were 73%, 84%, and 82%, respectively, versus only 37%, 33%, and 32% for 12 patients with an IPI of 3-4. To improve our results, we are conducting clinical trials with young DLBCL patients and with patients who are older than 60 years. The young DLBCL patients, who have more than two adverse prognostic features, are receiving high-dose chemotherapy and autologous stem cell rescue. The patients who are older than 60 years, regardless of IPI, are receiving rituximab immunotherapy and liposomal CHOP chemotherapy with or without involved-field radiotherapy. PMID- 12115339 TI - A problem-solving approach to stress reduction among younger women with breast carcinoma: a randomized controlled trial. AB - BACKGROUND: Previous research indicates that younger women (i.e., 50) and that coping style is significantly related to the psychosocial adjustment of women with this disease. The purpose of this study was to evaluate through a randomized controlled trial the effectiveness of a problem-solving training intervention designed to empower women with breast carcinoma to cope with a range of difficulties when diagnosed in mid-life. METHODS: The study population consisted of women aged 50 years or younger who had no prior history of breast carcinoma, were diagnosed with Stage I-IIIA tumors, and for whom a first course of chemotherapy had been initiated recently. The intervention consisted of two in person and four telephone sessions with an oncology nurse who provided problem solving skills training and informational materials to the women over a 12-week period. All subjects were assessed for physical and psychosocial adjustment through telephone and mailed surveys at baseline, at 4 -months, and at 8 months. RESULTS: Of 183 eligible women, 164 participated (a 90% participation rate), 149 of whom completed the study (a 91% completion rate). The subjects had significantly lower unmet needs and better mental health at the 4-month assessment. The intervention significantly decreased the number and severity of difficulties experienced by women with average or good problem-solving skills at 8 months, but was not effective in alleviating or resolving the problems encountered by women with poor problem-solving skills, relative to the control group. CONCLUSIONS: We conclude that this problem-solving therapy-based home care training intervention is an effective method of helping the majority of women with breast carcinoma to reduce the stresses associated with the diagnosis and treatment of cancer in mid-life. PMID- 12115340 TI - Significance of secondary ultrasonographic endometrial thickening in postmenopausal tamoxifen-treated women. AB - BACKGROUND: Ultrasonography has a limited value in endometrial assessment for identification of endometrial pathologies in postmenopausal tamoxifen-treated patients. METHODS: We compared the rate of endometrial pathologies and the mean +/- SD of endometrial thickness diagnosed after the first and second transvaginal ultrasonographic studies performed on 55 postmenopausal tamoxifen-treated patients with secondary endometrial thickening (Group I). This rate was also compared with 46 similar patients without secondary thickening (Group II). We also compared the mean +/- SD of endometrial thickness detected in various ultrasonographic studies, as well as various clinical features. RESULTS: A significantly higher rate of endometrial pathologies, including two cases of endometrial cancer identified in gynecologically asymptomatic patients (3.6%), was diagnosed in Group I after the second study compared with the first study (52.7% and 9.1%, respectively; P = 0.001) and compared with those diagnosed after the second study in Group II (30.4%; P = 0.03). There was a significant increase (74.7 +/- 115%) in endometrial thickness after the second study compared with the first study performed on Group I (10.7 +/- 5.53 mm and 16.59 +/- 5.53 mm, respectively; P = 0.0001) and a significant difference in endometrial thickness demonstrated in the second study performed on Groups I and II (16.59 +/- 5.53 mm and 11.4 +/- 3.91 mm, respectively; P = 0.001). There were no significant differences in the time elapsed since the diagnosis of breast carcinoma and from the beginning of tamoxifen treatment to the performance of the first ultrasonographic study as well as the time elapsed between the first and second studies performed. CONCLUSIONS: A significant increase (> 50%) in secondary endometrial thickening, measured ultrasonographically, in postmenopausal tamoxifen-treated patients, is associated with a high rate of endometrial pathologies, including endometrial cancer. PMID- 12115341 TI - Assessment of histologic features and expression of biomarkers in predicting pathologic response to anthracycline-based neoadjuvant chemotherapy in patients with breast carcinoma. AB - BACKGROUND: There is significant variability in the response of tumors to neoadjuvant chemotherapy, and the underlying mechanism for this variability is unknown. In this study, the authors investigated the roles of tumor nuclear grade, mitotic activity, and biomarker expression profiles in predicting the pathologic response of breast tumors to preoperative chemotherapy. METHODS: Eighty-two patients with breast carcinoma participated in two clinical trials and were treated with neoadjuvant chemotherapy, which consisted of either a conventional dose of fluorouracil, doxorubicin, and cyclophosphamide (FAC) or dose-escalated FAC. The mean age of the patients was 46 years (range, 24-69 years). Nuclear grade, mitotic activity, and biomarker profile (Her2-neu and mitosin expression patterns) in pretreatment tumors were correlated with the postchemotherapy pathologic response. RESULTS: Twelve patients (15%) had a complete pathologic response (CPR), 23 patients (28%) had a near complete response (NCR), and 47 patients (57%) had significant residual disease present either at the primary site or in the axillary lymph nodes. The authors found that the nuclear grade and mitotic activity of pretreatment tumors were correlated significantly with CPR and NCR (P = 0.002 and P = 0.004). Mitosin also was correlated significantly with CPR and NCR (P = 0.028). A higher percentage of patients with Her2-neu-positive tumors had a CPR or an NCR (P = 0.152). CPR and NCR were not correlated significantly with disease stage (P = 0.186) or lymph node positivity (P = 0.498). CONCLUSIONS: The current results indicate that tumor nuclear grade and tumor proliferative activity (mitotic activity and mitosin immunostaining) of pretreatment tumors in patients with breast carcinoma may serve as important indicators for the pathologic responsiveness of tumors to neoadjuvant, anthracycline-based chemotherapy. PMID- 12115342 TI - Treatment and prognosis of thymic carcinoma: a retrospective analysis of 40 cases. AB - BACKGROUND: Thymic carcinomas are rare neoplasms, and information regarding the results of treatment and possible prognostic factors in patients with these tumors is limited. METHODS: The records of 40 patients with histologically confirmed thymic carcinoma who were treated between 1984 and 1998 were reviewed. Twenty-seven patients were treated with surgical resection followed by radiotherapy with or without chemotherapy, and the remaining 13 patients were treated with radiotherapy with or without chemotherapy. The median follow-up time for the 13 surviving patients was 87 months (range, 44-193 months). RESULTS: The 5-year and 10-year actuarial overall survival rates in all patients were 38% and 28%, respectively. On univariate analysis, complete resection, Karnofsky performance status (KPS), histology, and Masaoka stage at the time of diagnosis were found to have a significant impact on overall survival, whereas on multivariate analysis, complete resection, KPS, and histology were found to be significant prognostic factors. With regard to the degree of resection, 12 of 16 patients (75%) treated with complete resection were alive and free of disease at the time of last follow-up whereas 1 of 24 patients (4%) treated with incomplete resection or biopsy still was alive. Among 12 surviving patients treated with complete resection, 8 with resectable tumors at the time of presentation all had low-grade histology (squamous cell carcinoma) and were treated successfully with complete resection and postoperative radiotherapy with or without adjuvant chemotherapy. The remaining four patients with unresectable tumors at the time of presentation were treated successfully with neoadjuvant chemotherapy, complete resection, and postoperative radiotherapy. CONCLUSIONS: The results of the current study indicate that multimodal treatment, especially complete resection and postoperative radiotherapy with or without chemotherapy, is a curative therapy for thymic carcinomas. PMID- 12115343 TI - Impact of thrombophilic gene mutations on thrombosis risk in patients with gastrointestinal carcinoma. AB - BACKGROUND: Patients with malignancies have an increased risk for thromboembolic events due to the release of tissue factor by the tumor, damage to the vessel wall, and immobilization. Moreover, tumors may improve their growth and metastatic spread by utilizing the coagulation system. To date, no information is available on the additional role of prothrombotic mutations in these patients. METHODS: The prevalence of the factor V Leiden mutation (FVL) and the prothrombin G20210A mutation and of homozygosity for the methylenetetrahydrofolate reductase (MTHFR) C677T substitution has been analyzed in a cohort of 175 patients with gastrointestinal adenocarcinoma by the polymerase chain reaction-restriction fragment length polymorphism technique. RESULTS: 6.9% of the patients were heterozygous for FVL, 5.7% were heterozygous for the prothrombin mutation, and 9.7% were homozygous for the MTHFR C677T mutation was detected in 9.7% of patients. Compared with the normal population, we found an increased prevalence of the prothrombin G20210A substitution (5.7% vs. 0.8%, P = 0.028). Thromboses were absent in 147 patients (Group A), whereas 28 of the patients suffered from thromboses during the period following tumor diagnosis (Group B). In Group A, 6.8% of the patients and 21.4% of the patients in Group B had a thrombosis before the diagnosis of cancer (P = 0.025, odds ratio [OR] 3.7). Heterozygous FVL was present in 4.8% of the patients in Group A and in 17.9% of the patients in Group B (P = 0.026, OR 4.4). In patients with thromboses before the detection of the tumor, the risk was elevated 6.3-fold (25.0% vs 5.0%, P = 0.015). Heterozygosity for the prothrombin mutation and homozygosity for the MTHFR C677T substitution did not increase the incidence of thromboses. CONCLUSIONS: We demonstrated a significant effect of FVL on thrombosis in patients with malignant disease. Most thromboses occurred during the first months after tumor diagnosis, implicating diagnostic and therapeutic procedures as important nongenetic risk factors for venous thromboembolism. Our data also indicate that the prothrombin G20210A mutation may be a possible cofactor in cancer pathogenesis. PMID- 12115344 TI - Phosphorylation of Akt/PKB is required for suppression of cancer cell apoptosis and tumor progression in human colorectal carcinoma. AB - BACKGROUND: Akt/protein kinase B (PKB), which is included in phosphatidyl inositol-3-OH kinase (PI3K) signaling, controls many intracellular processes, such as the suppression of apoptosis and the promotion of the cell cycle. Therefore, the authors investigated phosphorylated Akt (Ser473) in colorectal carcinomas to reveal the role of PI3K signaling during the development of colorectal carcinoma. METHODS: Expression of phosphorylated Akt (Ser473) in two colon carcinoma cell lines (DLD-1 and Colo205) and 65 human colorectal carcinomas was analyzed using western blotting and immunohistochemistry, respectively. Growth inhibition and induction of apoptosis caused by LY294002, a specific inhibitor of PI3K, were also examined in these cell lines. In tumor samples, the level of cell proliferation activity (Ki-67) and number of apoptotic bodies (single stranded DNA) were determined by counting positive cells. RESULTS: LY294002 significantly affected the proliferation and apoptosis of Colo205 cells, suggesting an association with the low phosphorylation level of Akt protein. Immunohistochemic analysis showed that 46% of the tumors had a high level of expression of phosphorylated Akt with a close association with Ki-67 proliferative activity (P < 0.001) and the number of apoptotic bodies (P < 0.001). Akt phosphorylation was also correlated with some clinicopathologic parameters of the malignancies, including depth of invasion, infiltration to venous vessels, lymph node metastasis, and clinicopathologic stage. CONCLUSIONS: The phosphorylated Akt level can monitor cell growth and resistance to apoptosis, indicating that activation of Akt plays an important role during the progression of colorectal carcinomas by helping promote cell growth and rescue cells from apoptosis. These findings also suggest the possibility of using LY294002 for treatment of colorectal carcinoma. PMID- 12115345 TI - 7-Hydroxytryptophan, a novel, specific, cytotoxic agent for carcinoids and other serotonin-producing tumors. AB - BACKGROUND: Carcinoids and small cell lung carcinomas stimulate their growth in an autocrine manner by releasing serotonin, an effect that is blocked by selective serotonergic receptor antagonists that, unfortunately, exert undesirable side effects on serotonergic central nervous function. Moreover, conventional chemotherapeutic agents, such as streptozocin, fluorouracil, cyclophosphamide, and doxorubicin, which target tumor cells directly, have produced disappointing results in the treatment of patients with these tumors in the advanced stage. Therefore, there is still a need for more specific and potent chemotherapeutic agents in the fight against serotonin-producing tumors. METHODS: The authors synthesized 7-hydroxytryptophan to test its chemotherapeutic value in cell culture, using a system consisting of serotonin-producing and nonproducing cell lines. RESULTS: The authors chose tryptophan hydroxylase, the rate-limiting enzyme of serotonin biosynthesis, which is expressed highly in small cell lung carcinomas and carcinoids, as a target for the induction of cellular suicide by chemotherapy. They found that this otherwise substrate specific enzyme was capable of metabolizing in situ a harmless tryptophan analogue, 7 hydroxytryptophan, to a potent toxin, 5,7-dihydroxytryptamine, a conversion blocked by the specific tryptophan hydroxylase inhibitor parachlorophenylalanine. CONCLUSIONS: These data suggest that 7-hydroxytryptophan may be a highly specific chemotherapeutic compound against serotonin-producing tumors that also interferes with the autocrine capabilities of serotonin synthesis. PMID- 12115346 TI - Prediction of pelvic lymph node metastasis by the ratio of cathepsin B to stefin A in patients with prostate carcinoma. AB - BACKGROUND: Pathologic grade and/or histologic score, extraprostatic extension indicated by invasion of the prostatic capsule, margin, and/or seminal vesicles by prostate cancer cells, serum total prostate-specific antigen (PSA), free PSA, complexed PSA levels and/or their ratios, regional pelvic lymph node metastases, and clinical staging have been used to diagnose and monitor the treatment of prostate carcinoma (PC) patients. The Gleason grading system is also used to grade/score a patient's stage of disease, with lower to higher scores indicating progression of PC. However, Gleason's system cannot be used to distinguish biologically aggressive PCs within a single Gleason score. Our objective was to identify subpopulations (or clones) of aggressive prostate cancers within an individual Gleason score by utilizing biological molecule(s) that also facilitate cancer cell invasion to prostatic stroma and metastasis to the lymph nodes. MATERIALS AND METHODS: Specimens were collected from 97 patients with PC and from 8 patients with benign prostatic hyperplasia. These patients had not been treated with hormonal and/or chemotherapeutic agents before undergoing a prostatectomy at the Minneapolis Veterans Affairs Medical Center. Formalin-fixed, paraffin or paraplast-embedded prostate tissue sections were stained with hematoxylin and eosin for pathologic diagnosis and adjacent sections were stained for for immunohistochemical study. We also collected data on age, race, extraprostatic extension, margin status, seminal vesicle, and lymph node invasion by cancer cells, clinical stage at prostatectomy, and mortality/survival data, including the available presurgery and postsurgery serum total PSA and prostatic acid phosphatase concentrations in patients. Immunohistochemical localization of mouse or rabbit anti-cathepsin B (CB) antibody IgG and mouse antihuman stefin (cystatin) A IgG was quantified using a computer-based image analysis system equipped with Metamorph software. RESULTS: CB and stefin A identified aggressive and less aggressive clones of PCs within an individual Gleason score. Tumors with a Gleason Score of 6 that are similar histologically and morphologically were heterogeneous with respect to the ratios of CB to stefin A (CB > stefin A, CB = stefin A, and CB < stefin A). We also found a significant positive association (P = 0.0066) between ratios of CB and stefin A (CB > stefin A) and the incidence of pelvic lymph node metastases, but not with ratios of CB less than stefin A and/or ratios of CB equal to stefin A. Patients with Gleason 7 PCs had a higher incidence of positive lymph nodes than those with Gleason Score 6 tumors. Our data indicated that mortality rates increased in patients when the ratios of CB were greater than stefin A. CONCLUSIONS: PC within an individual Gleason score is a heterogeneous tumor that contains clones or subpopulations of aggressive and less aggressive tumors that can be defined by the ratios of CB to stefin A. PC with an aggressive clone can be identified when the ratio of CB is greater than that of stefin A. Less aggressive clones are identified when the ratio of CB is less than that of stefin A or when the ratio of CB is equal to that of stefin A. The ratios of CB to stefin A can be used in the differential diagnosis and treatment of patients with PC. This is the first report to identify phenotypes of aggressive and less aggressive PCs within a Gleason score. PMID- 12115347 TI - Prognostic impact of extensive parenchymal invasion pattern in pT3 renal pelvic transitional cell carcinoma. AB - BACKGROUND: Pathologic T3 renal pelvic transitional cell carcinoma exhibits various patterns of invasion. The authors investigated the prognostic impact of three patterns of invasion of pT3 renal pelvic transitional cell carcinoma. METHODS: Of 212 patients who underwent surgery for renal pelvic transitional cell carcinoma, 70 with pT3 disease were eligible for the main analyses. The candidate predictors of prognosis included patient age, gender, lesion laterality, tumor grade, perioperative cisplatin-based systemic chemotherapy, lymph node involvement, vascular involvement, and patterns of invasion. Invasion patterns were classified as fat invasion, ductal involvement, or parenchymal invasion. RESULTS: Mean postoperative followup was 33.5 months (range, 1-136 months). On univariate analysis, gender, lymph node involvement, vascular involvement, and extensive parenchymal invasion each had a significant impact on the cause specific survival rate. A multivariate analysis using Cox stepwise regression revealed that extensive parenchymal involvement was the strongest prognostic predictor (P = 0.0004, hazard ratio = 5.59). Lymph node involvement (P = 0.0175, hazard ratio = 3.14) and gender (P = 0.0361, hazard ratio = 2.42) were other weaker predictors. Statistically, pT3 disease without extensive parenchymal invasion had a prognosis similar to that of lower stage disease, and pT3 disease with extensive parenchymal invasion had a prognosis similar to that of pT4 disease. CONCLUSIONS: Extensive parenchymal invasion has a strong prognostic impact in renal pelvic transitional cell carcinoma. pT3 disease should be subclassified into two separate entities, that with and that without extensive parenchymal invasion, in view of prognosis. PMID- 12115348 TI - Microsatellite instability and mutation analysis of candidate genes in unselected sardinian patients with endometrial carcinoma. AB - BACKGROUND: Microsatellite instability (MSI) is due mostly to a defective DNA mismatch repair (MMR). Inactivation of the two principal MMR genes, hMLH1 and hMSH2, and the PTEN tumor suppressor gene seems to be involved in endometrial tumorigenesis. In this study, Sardinian patients with endometrial carcinoma (EC) were analyzed to assess the prevalence of both the mutator phenotype (as defined by the presence of MSI and abnormal MMR gene expression at the somatic level) and the hMLH1, hMSH2, and PTEN germline mutations among patients with MSI positive EC. METHODS: Paraffin embedded tissue samples from 116 consecutive patients with EC were screened for MSI by polymerase chain reaction-based microsatellite analysis. Immunohistochemistry (IHC) with anti-hMLH1 and anti-hMSH2 antibodies was performed on MSI positive tumor tissue sections. Germline DNA was used for mutational screening by denaturing high-performance liquid chromatography analysis and automated sequencing. RESULTS: Thirty-nine patients with EC (34%) exhibited MSI; among them, 25 tumor samples (64%) showed negative immunostaining for hMLH1/hMSH2 proteins (referred to as IHC negative). No disease-causing mutation within the coding sequences of the hMLH1/hMSH2 and PTEN genes was found in patients with EC who had the mutator phenotype (MSI positive and IHC negative), except for a newly described hMLH1 missense mutation, Ile655Val, that was observed in 1 of 27 patients (4%). Although MSI was more common among patients with advanced-stage EC and increased as the tumor grade increased, no significant correlation with disease free survival or overall survival was observed among the two groups (MSI positive or MSI negative) of patients with EC. CONCLUSIONS: In patients with MSI positive EC, epigenetic inactivations rather than genetic mutations of the MMR genes seem to be involved in endometrial tumorigenesis. No prognostic value was demonstrated for MSI in patients with EC. PMID- 12115350 TI - High-density allelotyping of chromosome 8p in hepatocellular carcinoma and clinicopathologic correlation. AB - BACKGROUND: Allelic deletions are frequent genetic alterations in patients with hepatocellular carcinoma (HCC). METHODS: To evaluate the allelic losses on chromosome 8p in HCC patients and define their clinicopathologic significance, we performed high-density allelotyping on 8p in 60 patients with HCC and analyzed the clinicopathologic correlation. RESULTS: Using 24 microsatellite markers, allelic losses on 8p were frequent. Loss of heterozygosity (LOH) at one or more loci was observed in 34 (57%) HCC patients. When the allelic losses were compared between groups categorized by clinicopathologic variables, significant correlation was found between tumors with interstitial losses and larger tumor size (> 5 cm; P = 0.026). In addition, allelic loss at D8S298 at 8p22 was associated closely with venous permeation, tumor microsatellite formation, and larger tumor size (P = 0.019, 0.024, and 0.007, respectively). LOH at locus D8S1721 at 8p23.1 was seen more frequently in nonencapsulated tumors (P = 0.007) and LOH at D8S1771 at 8p21.3 was associated with a larger tumor size and poorer cellular differentiation (P = 0.018 and 0.049, respectively). CONCLUSIONS: Allelic losses on 8p are frequent in HCC patients. Association of allelic losses at specific loci on 8p with a more aggressive tumor behavior suggests that loss/inactivation of putative tumor suppressor gene(s) located at these regions may confer a tumor growth advantage and contribute to the progression of HCC. PMID- 12115349 TI - Multimodal intensification therapy for previously untreated advanced resectable squamous cell carcinoma of the oral cavity, oropharynx, or hypopharynx. AB - BACKGROUND: An intensified treatment regimen for previously untreated Stage III and IV resectable oral cavity, oropharyngeal, or hypopharyngeal squamous cell carcinoma was analyzed to assess disease control, patient compliance, and toxicity. METHODS: Forty three patients with previously untreated, advanced, resectable squamous cell carcinoma of the oral cavity, oropharynx, or hypopharynx were enrolled in a prospective Phase II institutional clinical trial at a tertiary care comprehensive cancer center. This regimen was a continuum of multimodal treatment in a contracted time interval. It included preoperative slightly accelerated hyperfractionated radiotherapy with concurrent cisplatin, followed immediately with surgery and intraoperative radiotherapy, and completed with early postoperative weekly paclitaxel (beginning on Day 6 after surgery), two additional cisplatin cycles, and concurrent once daily radiotherapy beginning on Day 28 after surgery. RESULTS: The current trial was designed to reduce the toxicity of the systemic therapy while maintaining or improving locoregional/distant disease control and patient compliance. There were 43 patients enrolled, and the range of time at risk was 2.6 to 24.7 months (median, 14.6 months). Of the 43 registered patients, 43 were evaluable. The locoregional (100%) and systemic (93%) disease control rates were excellent, with low rates of patient noncompliance (21%) and reduced levels of toxicity. CONCLUSIONS: An intensive treatment regimen that improves disease control and treatment compliance is clearly feasible for this patient population. Future plans include modifications to continue to reduce toxicity and expansion to a multi-center Phase II trial to determine if the single institutional results can be duplicated. PMID- 12115351 TI - A phase II trial of gemcitabine in patients with advanced hepatocellular carcinoma. AB - BACKGROUND: There is no effective systemic therapy for patients with hepatocellular carcinoma. A recent trial reported a moderate antitumor activity for gemcitabine among Asian patients with advanced hepatocellular carcinoma. This led to our examination of the efficacy and tolerability of the drug in a population of U.S. patients. METHODS: Thirty patients with measurable, unresectable, or metastatic hepatocellular carcinoma who had received at least one previous form of systemic therapy were enrolled. All patients were required to have adequate major organ function and performance status. Patients received gemcitabine (1000 mg/m(2) intravenously over 30 minutes weekly) for 3 consecutive weeks followed by a 1-week rest. Patients were assessed radiographically every 8 weeks. RESULTS: All 30 patients were evaluable for response and toxicity. Ninety cycles of therapy were administered (median 2, range 1-8). No complete or partial responses were observed. Nine patients (30%) had stable disease (median duration 7.4 months, range 2-17). Median survival for all 30 patients was 6.9 months (95% confidence interval, 4.5-13.5) and the 1-year survival rate was 40%. Mild hematologic toxicity occurred. Two patients (7%) developed Grade 4 neutropenia and one patient (3%) experienced Grade 3 thrombocytopenia. There were no episodes of febrile neutropenia. One patient who had previously undergone orthotopic liver transplantation developed hemolytic-uremic syndrome that resolved with discontinuation of chemotherapy and plasmapheresis. CONCLUSIONS: Although generally well tolerated, gemcitabine had minimal effect in patients with advanced hepatocellular carcinoma. PMID- 12115352 TI - A natural history of melanomas and dysplastic nevi: an atlas of lesions in melanoma-prone families. AB - BACKGROUND: Few long-term clinical and histologic data for melanocytic lesions have been available based on the mutation status of families at an increased risk of melanoma. In the current study, the authors describe the clinical and histologic features of dysplastic nevi and melanoma over time in families at an increased risk of melanoma with differing germline mutations in CDKN2A, CDK4, or not yet identified genes. METHODS: Thirty-three families with > 2 living members with invasive melanoma were evaluated clinically and followed prospectively for up to 25 years. All the participants were evaluated by the same study team at the Clinical Center of the National Institutes of Health or in local clinics. After informed consent was obtained, family members (n = 844) were examined and photographed. Blood was obtained for genetic studies; genotyping for CDKN2A and CDK4 was performed. Sequential photographs of melanocytic lesions were taken as part of the clinical evaluations. When melanocytic lesions were removed, the histology was reviewed. Representative photographs and photomicrographs were selected for six classes of lesions and three mutation groups. RESULTS: All the families were found to have members with dysplastic nevi and melanoma; 17 had mutations in CDKN2A, 2 had mutations in CDK4, and 14 had no mutations in either gene identified. The majority of dysplastic nevi either remain stable or regress; few change in a manner that should cause concern for melanoma. With careful surveillance, melanomas can be found early. CONCLUSIONS: The melanomas and dysplastic nevi that were found to occur in the study families did not appear to vary by the type of mutation identified in the families. PMID- 12115354 TI - High-grade gliomas in patients with prior systemic malignancies. AB - BACKGROUND: The current study was conducted to characterize the impact of a prior malignancy on the diagnosis, treatment, and outcome of high-grade glioma. METHODS: A retrospective study of 21 patients with a histologic diagnosis of glioblastoma multiforme (GBM), anaplastic astrocytoma (AA), or anaplastic oligodendroglioma (AO) after a prior diagnosis of solid tumor or hematologic malignancy was conducted. Glioma histology (GBM vs. AA/AO), patient age ( 60 years), and extent of resection (biopsy vs. subtotal vs. complete) were evaluated for their prognostic influence. RESULTS: Of the 21 patients studied, 17 had GBM, 3 had AA, and 1 patient had high-grade AO. There were 25 systemic carcinomas diagnosed in 21 patients (18 solid tumors including breast carcinoma, prostate carcinoma, and melanoma and 7 hematologic malignancies). The glioma occurred within a previous radiation field in only three patients, two of whom had solid tumors and one of whom had a childhood hematologic malignancy. Surgical resection was the initial treatment for the brain tumor in 17 patients, and the majority of patients received radiation therapy and adjuvant chemotherapy. Four patients who initially were misdiagnosed as having brain metastases received whole brain radiation therapy as their initial treatment, thereby compromising optimal care. The overall median survival for all the patients in the current study was 14 months (range, 1-44 months) from the time of brain tumor diagnosis. The extent of resection was found to be the only prognostic variable that was associated with survival (P = 0.03). CONCLUSIONS: Secondary glioma may occur in patients as a consequence of therapy for a prior malignancy, but most often represents a sporadic event. The outcome and recommended treatment are identical to those for patients with primary gliomas. Accurate diagnosis is essential; neuroimaging often is suggestive of a primary brain tumor and should initiate surgical intervention so that histopathology can be obtained early and appropriate treatment instituted. PMID- 12115353 TI - Hepatocyte growth factor is associated with poor prognosis of malignant gliomas and is a predictor for recurrence of meningioma. AB - BACKGROUND: Hepatocyte growth factor (HGF) is a cytokine that participates in multiple cell functions; it promotes proliferation, motility, and morphogenesis of epithelial cells. Some malignant tumors, such as breast carcinoma, bronchogenic carcinoma, and multiple myeloma, overexpress it and its receptor. Hepatocyte growth factor is also present in normal astrocytes; therefore, it is important to investigate whether HGF participates in the pathophysiology of malignant gliomas and other brain tumors. Intratumoral concentration of HGF in human intracranial neoplasms was measured and correlated with prognosis, tumor recurrence, vasogenic edema, cell proliferation index, and vascular density. METHODS: Hepatocyte growth factor concentration was measured in 62 intracranial tumors, including 16 anaplasic astrocytomas (AA), 16 glioblastoma multiformes (GM), 11 meningiomas, 9 hypophyseal adenomas, 7 oligodendrogliomas, and 3 cordomas, and in 4 samples of nonneoplastic brain tissue. The following parameters were correlated with HGF values: survival and tumor recurrence, cell proliferation index and vascular density as determined by immunohistopathologic analysis, and peritumoral edema as seen by magnetic resonance imaging. RESULTS: Hepatocyte growth factor concentration (pg/mL) was significantly higher in malignant gliomas (AA and GM) than in adenomas, oligodendrogliomas, and nonneoplastic brain tissue, but it was similar to that of meningiomas. Mean survival of patients with AA was 16.5 +/- 3.6 months and for patients with GM 12.3 +/- 1.3 months. Hepatocyte growth factor concentration was higher in GM than in AA (15,844 +/- 2504 vs. 7499 +/- 1703, P = 0.0375) and was correlated with the cell proliferation index and with poor prognosis. Likewise, mean tumoral concentration of HGF was higher in meningiomas that relapsed than in those without recurrence (22,887 +/- 6489 vs. 2090 +/- 497, P = 0.008). CONCLUSIONS: Intratumoral concentration of HGF in gliomas is associated with malignancy and poor prognosis. High HGF is also found in meningiomas and is related with long term recurrence. The current findings suggest that the routine measurement of HGF may be used as a predictive factor for planning therapeutic strategies in both malignant gliomas and meningiomas. The potential use of HGF inhibitors or antagonists for therapy of these tumors should be explored. PMID- 12115355 TI - Phase II trial of gemcitabine in advanced sarcomas. AB - BACKGROUND: Care for patients with advanced sarcomas is mainly palliative. Gemcitabine, a nucleoside antimetabolite, is an analog of deoxycytidine that has shown antitumor activity in several tumors. The aim of the current study was to determine the clinical activity of gemcitabine in patients with sarcomas. METHODS: The authors evaluated gemcitabine in patients with histologically confirmed sarcomas; one prior exposure to chemotherapy treatment was allowed. Prior radiation was allowed if given to non-indicator lesions. Treatment consisted of gemcitabine 1250 mg/m(2) intravenously over 30 minutes, every week x three, cycles repeated q28 days. RESULTS: Twenty nine of 30 patients were evaluable; one patient refused to initiate study treatment. The mean age was 50 years (range, 22-81 years); 59% were male, and 35% had an Eastern Cooperative Oncology Group performance status of 0 (vs. 1 or 2). Patients were histologically classified as leiomyosarcoma (seven gastrointestinal, four retroperitoneal, two inferior vena caval, three of the extremity, and two uterine), synovial (two patients), malignant fibrous histiocytoma (two patients), fibrosarcoma (one patient), osteosarcoma (two patients), liposarcoma (one patient), hemangiosarcoma (one patient), or giant cell (one patient). Patients received an average of two cycles (range, one to eight). Eighty three percent of patients discontinued treatment due to progression and 14% due to toxicity/refusal. Hematologic toxicities >or= Grade 3 were seen in 32% of patients and consisted of leukopenia and thrombocytopenia. Anorexia (Grade 1/2 in 6 patients, Grade 3 in 1 patient), nausea (Grade 1/2 in 7 patients, Grade 3 in 1 patient), and lethargy (Grade 1/2 in 19 patients) were the most frequently observed nonhematologic toxicities. One patient experienced Grade 3 edema and muscle infarction. A different patient experienced unexplained Grade 3 chest pain. One partial response was observed in a uterine leiomyosarcoma patient lasting at least three months. Overall response rate was 3% (95% confidence interval [CI]: 0-15). Median time -to progression was 2.1 months (95% CI: 1.8-3.0). CONCLUSIONS: The current gemcitabine regimen demonstrated acceptable levels of toxicity, but it failed to produce the number of responses needed to justify expansion of the current study. This regimen is not recommended for advanced sarcomas. PMID- 12115356 TI - Antifungal prophylaxis for severely neutropenic chemotherapy recipients: a meta analysis of randomized-controlled clinical trials. AB - BACKGROUND: The overall clinical efficacy of the azoles antifungal agents and low dose intravenous amphotericin B for antifungal chemoprophylaxis in patients with malignant disease who have severe neutropenia remains unclear. METHODS: Randomized-controlled trials of azoles (fluconazole, itraconazole, ketoconazole, and miconazole) or intravenous amphotericin B formulations compared with placebo/no treatment or polyene-based controls in severely neutropenic chemotherapy recipients were evaluated using meta-analytical techniques. RESULTS: Thirty-eight trials that included 7014 patients (study agents, 3515 patients; control patients, 3499 patients) were analyzed. Overall, there were reductions in the use of parenteral antifungal therapy (prophylaxis success: odds ratio [OR], 0.57; 95% confidence interval [95% CI], 0.48-0.68; relative risk reduction [RRR], 19%; number requiring treatment for this outcome [NNT], 10 patients), superficial fungal infection (OR, 0.29; 95% CI, 0.20-0.43; RRR, 61%; NNT, 12 patients), invasive fungal infection (OR, 0.44; 95% CI, 0.35-0.55; RRR, 56%; NNT, 22 patients), and fungal infection-related mortality (OR, 0.58; 95% CI, 0.41-0.82; RRR, 47%; NNT, 52 patients). Invasive aspergillosis was unaffected (OR, 1.03; 95% CI, 0.62-1.44). Although overall mortality was not reduced (OR, 0.87; 95% CI, 0.74-1.03), subgroup analyses showed reduced mortality in studies of patients who had prolonged neutropenia (OR, 0.72; 95% CI, 0.55-0.95) or who underwent hematopoietic stem cell transplantation (HSCT) (OR, 0.77; 95% CI, 0.59-0.99). The multivariate metaregression analyses identified HSCT, prolonged neutropenia, acute leukemia with prolonged neutropenia, and higher azole dose as predictors of treatment effect. CONCLUSIONS: Antifungal prophylaxis reduced morbidity, as evidenced by reductions in the use of parenteral antifungal therapy, superficial fungal infection, and invasive fungal infection, as well as reducing fungal infection-related mortality. These effects were most pronounced in patients with malignant disease who had prolonged neutropenia and HSCT recipients. PMID- 12115357 TI - Higher oxidation and lower antioxidant levels in peripheral blood plasma and bone marrow plasma from advanced cancer patients. AB - BACKGROUND: Bone marrow (BM) is an important tissue in the generation of immunocompetent and peripheral blood cells. The precursors of hematopoietic cells in BM undergo continuous proliferation and differentiation and are highly vulnerable to acute and chronic oxidative stress. Little is known about the oxidant and antioxidant status in the BM of untreated patients with nonhematologic tumors. In this study, oxidative stress was evaluated in peripheral blood plasma (PBP) and BM plasma (BMP) from lung carcinoma (LC) and breast carcinoma (BC) patients. METHODS: The sample included 13 consecutive untreated LC patients, 15 BC patients, and 11 healthy controls. Luminol-dependent chemiluminescence was used to evaluate oxygen radical generation by peripheral blood neutrophils. Lipid oxidation, assessed by 2-thiobarbituric acid-reactive substances (TBARS), and alpha-tocopherol, beta-carotene, and total ubiquinol-10 levels were determined in PBP and BMP. RESULTS: In LC and BC patients, neutrophil chemiluminescence was higher (128% and 264%, respectively) than in controls (P < 0.05). In cancer patients, TBARS levels were higher in both PBP (51% and 243% for LC and BC patients, respectively) and BMP (66% and 305% for LC and BC patients, respectively) than in plasma from controls (P < 0.01). alpha-Tocopherol and total ubiquinol-10 levels were significantly lower in BMP from BC patients compared with controls. In BC patients, alpha-tocopherol content in PBP was significantly lower than in controls. CONCLUSIONS: Untreated cancer patients presented an imbalance between oxidant generation and lipid-soluble antioxidant levels in favor of the former. PMID- 12115358 TI - The Bleeding Risk Index: a clinical prediction rule to guide the prophylactic use of platelet transfusions in patients with lymphoma or solid tumors. AB - BACKGROUND: The correlation between platelet count and bleeding has been well described, although no formal methods for applying this information to clinical decisions are available. The authors developed a clinical prediction rule to guide the prophylactic use of platelet transfusions among patients with lymphoma or solid tumors. METHODS: The Bleeding Risk Index (BRI) was developed from logistic regression analysis of a randomly selected 750-chemotherapy cycle derivation set using data from Day 1 of cycles. The sensitivity and specificity of a BRI-based prophylaxis strategy were compared in a 512-cycle validation set with two strategies based on initiation of prophylaxis when platelet counts fell below thresholds of 20,000 per microL or 10,000 per microL. RESULTS: Factors that were predictive of bleeding included any prior episode of bleeding (odds ratio [OR], 5.6; 95% confidence interval [95% CI], 2.2-14.0), treatment with a drug affecting platelet function (OR, 5.1; 95% CI, 2.0-12.6), bone marrow metastases (OR, 4.3; 95% CI, 1.7-10.8), a baseline platelet count < 75,000 per microL (OR, 3.5; 95% CI, 1.4-8.9), genitourinary or gynecologic malignancy (OR, 3.3; 95% CI, 1.3-8.2), a Zubrod performance status score > 2 (OR, 3.4; 95% CI, 1.4-8.5), and treatment with agents that were highly toxic to the bone marrow (OR, 2.2; 95% CI, 1.0-5.4). Compared with 20,000 and 10,000 platelet threshold strategies, the BRI based strategy provided the best trade-off between sensitivity for major bleeding episodes (80%) and specificity for any bleeding (84%). CONCLUSIONS: Patients with lymphoma or solid tumors who are at high risk of bleeding can be identified reliably on Day 1 of a chemotherapy cycle. An individualized, BRI-based approach to bleeding prophylaxis provides a highly sensitive and specific alternative to traditional, nonindividualized platelet threshold strategies. PMID- 12115359 TI - Vinorelbine in previously treated advanced childhood sarcomas: evidence of activity in rhabdomyosarcoma. AB - BACKGROUND: Vinca alkaloids have proved active against a number of pediatric malignancies. The aim of this study was to assess the feasibility and effectiveness of using vinorelbine in previously treated pediatric patients with advanced sarcomas. METHODS: From September 1998 to August 2001, 33 previously treated patients with progressive sarcoma were treated: 13 had rhabdomyosarcomas, 5 had other soft tissue sarcomas, 9 had Ewing sarcomas, and 6 had osteosarcomas. Vinorelbine was given intravenously on Days 1 and 8 of a 21-day treatment cycle. Four patients with uncontrolled pain or central nervous system invasion received concurrent radiotherapy and were only evaluated for toxicity. RESULTS: One hundred seventy-eight treatment cycles were administered (median of four per patient, range 1-20). Grade 3 to 4 neutropenia occurred in 63% of patients, Grade 3 anemia in 9%, and Grade 3 thrombocytopenia in 3%. Nonhematological toxicity was mild or moderate, i.e., always lower than Grade 3, with the exception of one child who experienced paralytic ileus. Twenty-eight patients were assessable for response. Eight patients had a partial response, one patient had a minor response, and nine patients had stable disease. Objective responses were observed in 6 of 12 patients with rhadbomyosarcomas (five of six of the alveolar subtype), in one of five patients with osteosarcomas, and in one of seven patients with Ewing sarcomas. The median duration of response was 10 months (range, 3+ to 20). CONCLUSIONS: Vinorelbine has a favorable toxicity profile with evidence of biological activity in already heavily treated pediatric patients with sarcomas. In particular, the objective response rate obtained for patients with alveolar rhabdomyosarcoma seems very promising. Due to the few cases considered here, further Phase II studies are needed to establish a potential role of vinorelbine in the treatment of these tumors. PMID- 12115360 TI - Clear cell sarcoma of tendons and aponeuroses in pediatric patients: a report from the Italian and German Soft Tissue Sarcoma Cooperative Group. AB - BACKGROUND: Clear cell sarcoma (CCS) of tendons and aponeuroses is extremely rare in childhood and little information is available on its clinical management. Originally believed to be a type of melanoma of soft tissue origin, CCS is now considered a distinct clinicopathologic entity that behaves like a high-grade soft tissue sarcoma. We report on a series of 28 pediatric patients treated from 1980 to 2000 by the Soft Tissue Sarcoma Italian Cooperative Group and the German Cooperative Group. METHODS: Patients were treated with a multimodality therapeutic approach. Surgical resection was complete in 17 patients (mutilating in 3), radiotherapy was administered to 8 patients, and 20 patients received chemotherapy. RESULTS: After a median follow-up of 102 months (range, 19-238 months), the 5-year and event-free survival rates were 66.4% and 63.3%, respectively. Seventeen patients were alive in first remission, two were alive in second remission, and nine had died of disease. The response to chemotherapy in the 7 evaluable patients included one partial remission, one minor response, and five no responses. Radiotherapy contributed to achieving local control in four of six Intergroup Rhabdomyosarcoma Study (IRS) Group II patients. Statistically significant differences in outcome were evident according to IRS group, tumor size, and site. CONCLUSIONS: Our study confirms the aggressive behavior of CCS. Complete surgical resection represents the mainstay of treatment, and even the only treatment for patients with small tumors. Radiotherapy may control microscopic residual disease after surgery. Chemotherapy is ineffective and the prognosis is unfavorable for patients with unresectable and large tumors. PMID- 12115361 TI - Evaluation of chemotherapy response in pediatric bone sarcomas by [F-18] fluorodeoxy-D-glucose positron emission tomography. AB - BACKGROUND: Response to neoadjuvant chemotherapy is a significant prognostic factor for osteosarcoma (OS) and the Ewing sarcoma family of tumors (ESFT). Conventional radiographic imaging does not discriminate between responding and nonresponding osseous tumors. [F-18]-fluorodeoxy-D-glucose (FDG) positron emission tomography (PET) is a noninvasive imaging modality that accurately predicts histopathologic response in patients with various malignancies. To describe the FDG PET imaging characteristics and to determine the correlation between FDG PET imaging and chemotherapy response in children with bone sarcomas, we reviewed our single institution experience. METHODS: Thirty-three pediatric patients with OS or ESFT with osseous primary sites were evaluated by FDG PET. All patients received standard neoadjuvant chemotherapy. FDG PET standard uptake values before (SUV1) and after (SUV2) chemotherapy were analyzed and correlated with chemotherapy response assessed by histopathology in surgically excised tumors. Twenty-six patients had SUV1, SUV2, and surgical excision. RESULTS: Although the mean SUV1 in children with OS or ESFT were similar (8.2. vs. 5.3, P = 0.13), mean SUV2 for OS patients was greater than the values for ESFT patients (3.3 vs. 1.5, P = 0.01). All ESFT patients and 28% of OS patients had a favorable histologic response to chemotherapy (>or= 90% necrosis). Combining ESFT and OS patients, both SUV2 and the ratio of SUV2 to SUV1 (SUV2:SUV1) were correlated with histologic response (P = 0.01 for both comparisons). CONCLUSION: FDG PET evaluation of pediatric bone sarcomas demonstrated significant alteration in response to neoadjuvant chemotherapy. SUV2 and SUV2:SUV1 correlated with histopathologic assessment of response and potentially could be used as a noninvasive surrogate to predict response in patients. PMID- 12115362 TI - Radiation-induced cavernous hemangiomas of the brain: a late effect predominantly in children. AB - INTRODUCTION: The induction of cavernomas as a consequence of brain irradiation was first suspected in 1994 and has been controversial since that time. METHODS: Between 1986 and 2000, 189 cerebral cavernomas were diagnosed in the Neurosurgical Department of the University of Heidelberg; of those patients, 5 had received prior radiation therapy. The ages of these 5 patients were compared with those of the 184 others with naturally occuring cavernomas. In an examination of 40 patients with cavernomas occurring after radiation (the 5 mentioned above, plus 35 from the literature) the age distribution was investigated, and a possible relationship between radiation dosage and latency interval to diagnosis of cavernoma was examined. RESULTS: Almost one in four of the patients under 15 years of age diagnosed with a cerebral cavernoma in the Neurosurgical Department of the University of Heidelberg had received prior radiation. In 40 patients with cavernomas and prior radiation (5 from Heidelberg, 35 from the literature), there was a clear accumulation in the age group of 10-19 years (50%). Most of those patients had received radiation in the first 10 years of life. The accumulation of cavernomas after radiation in childhood could not be explained by a greater frequency of radiation exposure in children compared to adults. In children up to 10 years of age at the time of radiation therapy, a dose of 3000 cGy and higher was followed by a shorter latency interval to incidence of cavernoma (P = 0.0018). In patients older than 10 years at the time of radiation, postradiation cavernomas only occurred when dosage was 3000 cGy or greater. CONCLUSION: These results indicate a correlation between radiation and cavernoma, particularly in children under 10 years of age at the time of radiation therapy. In adults, cavernomas after radiation rarely occur, and then only after higher radiation dosages (3000 cGy or more). PMID- 12115363 TI - Clinical relevance of internal tandem duplication of the FLT3 gene in childhood acute myeloid leukemia. AB - BACKGROUND: Recently, an internal tandem duplication of the FLT3 gene (FLT3/ITD) was found in approximately 20% of adult acute myeloid leukemia (AML) cases and associated with a poor outcome. However, there are few studies on FLT3/ITD in childhood AML, and the clinical significance of FLT3/ITD is thus unclear. METHODS: FLT3/ITD was analyzed in 80 children with de novo AML. The genomic DNA polymerase chain reaction (PCR) assay was performed to identify FLT3/ITD. Genescan analysis to determine the allelic distribution was then performed for those PCR products with aberrant bands. Direct sequencing of PCR products was also carried out in each sample with FLT3/ITD. RESULTS: The incidence of FLT3/ITD was 11.3% (9 out of 80 patients) in AML, with 25% (3 out of 12 patients) in acute promyelocytic leukemia (APL) and 8.8% (6 out of 68 patients) in non-M3 AML. The size of duplicated fragments varied from 21 base pairs (bp) to 75 bp, and the mutant to wild type ratio of FLT3 ranged from 0.28 to 16.60 in the nine patients with FLT3/ITD. The incidence of FLT3/ITD in childhood AML in patients > 10 years of age was 24%, compared to 5% of those patients 25 years living in Copenhagen County, Denmark. The responses to questionnaires and the results of examinations, including the Test of Psychic Vulnerability, were collected during 1982-1984 and during 1991-1992. The observed numbers of cancers were compared with the numbers that would have been expected if the cohort members had experienced the same risk of cancer as the population of Copenhagen County. Regression analyses were performed with the Cox proportional hazards model to adjust for well-known risk factors for cancer. RESULTS: A total of 403 cancers were observed, and 412.02 were expected, yielding a standardized incidence ratio of 0.98 (95% confidence interval [95% CI], 0.88 1.19). The authors did not observe a significant increase in the risk of site specific cancers. The risk for cancer was not influenced by the type of vulnerability in a multivariate analysis (hazards ratio, 1.16; 95% CI, 0.85 1.57). CONCLUSIONS: The authors found no increased risk for cancer among psychically vulnerable persons compared with nonvulnerable persons; however, the results indicate that behavior and certain life-style factors may be determined by personality, which, in turn, may determine the risk for cancer. PMID- 12115365 TI - Primary malignant neoplasms of the appendix: a population-based study from the surveillance, epidemiology and end-results program, 1973-1998. AB - BACKGROUND: Cancer of the appendix is an uncommon disease that is rarely suspected rarely before surgery. Although several case series of these tumors have been published, little research has been anchored in population-based data on cancer of the appendix. METHODS: This analysis included all actively followed cases of appendiceal neoplasms reported to the National Cancer Institute's Surveillance, Epidemiology and End-Results (SEER) program between 1973 and 1998. Tumors were classified as "colonic type" adenocarcinoma, mucinous adenocarcinoma, signet ring cell carcinoma, goblet cell carcinoid, and "malignant carcinoid" (SEER only collects data on carcinoids specifically classified as malignant). We compared incidence, overall survival and survival rates by extent of disease at diagnosis. RESULTS: Between 1973 and 1998, 2117 appendiceal malignancies were reported to the SEER program, of which 1645 cases were included in the analysis. Age-adjusted incidence of cancer of the appendix was 0.12 cases per 1,000,000 people per year. Demographic characteristics of patients with goblet cell carcinoid tumors were midway between those of patients with malignant carcinoid and all types of adenocarcinomas. After controlling for age and extent of disease at diagnosis, the overall survival rate for patients diagnosed between 1983 and 1997 (n = 1061) was significantly worse for those with signet ring cell carcinoma than for those with any other tumor type (P < 0.01). In addition, overall survival rates were better for patients with malignant carcinoid (P = 0.01). CONCLUSIONS: Demographic characteristics of patients with cancer of the appendix vary by histology. Except for signet ring cell carcinoma and malignant carcinoid, the extent of disease at time of diagnosis is a more important predictor of survival than histology. PMID- 12115366 TI - Breast and cervical carcinoma screening practices among women in rural and nonrural areas of the United States, 1998-1999. AB - BACKGROUND: Prior studies have suggested that women living in rural areas may be less likely than women living in urban areas to have had a recent mammogram and Papanicolau (Pap) test and that rural women may face substantial barriers to receiving preventive health care services. METHODS: The authors examined both breast and cervical carcinoma screening practices of women living in rural and nonrural areas of the United States from 1998 through 1999 using data from the Behavioral Risk Factor Surveillance System. The authors limited their analyses of screening mammography and clinical breast examination to women aged 40 years or older (n = 108,326). In addition, they limited their analyses of Pap testing to women aged 18 years or older who did not have a history of hysterectomy (n = 131,813). They divided the geographic areas of residence into rural areas and small towns, suburban areas and smaller metropolitan areas, and larger metropolitan areas. RESULTS: Approximately 66.7% (95% confidence interval [CI] = 65.8% to 67.6%) of women aged 40 years or older who resided in rural areas had received a mammogram in the past 2 years, compared with 75.4% of women living in larger metropolitan areas (95% CI = 74.9% to 75.9%). About 73.0% (95% CI = 72.2% to 73.9%) of women aged 40 years or older who resided in rural areas had received a clinical breast examination in the past 2 years, compared with 78.2% of women living in larger metropolitan areas (95% CI = 77.8% to 78.7%). About 81.3% (95% CI = 80.6% to 82.0%) of 131,813 rural women aged 18 years or older who had not undergone a hysterectomy had received a Pap test in the past 3 years, compared with 84.5% of women living in larger metropolitan areas (95% CI = 84.1% to 84.9%). The differences in screening across rural and nonrural areas persisted in multivariate analysis (P < 0.001). CONCLUSIONS: These results underscore the need for continued efforts to provide breast and cervical carcinoma screening to women living in rural areas of the United States. PMID- 12115367 TI - Prognostic factors for death after an isolated local recurrence in patients with early-stage breast carcinoma. AB - BACKGROUND: The authors analyzed the outcome of patients with early-stage breast carcinoma after an isolated local recurrence, taking into account initial tumor characteristics and the type of initial treatment and local salvage treatment. METHODS: One hundred five patients were studied who presented with a breast tumor measuring or= 70 years vs. < 70 years; OR = 4.06, P < 0.001), and the number of examined nodes (10 resected lymph nodes vs. 0-9; OR = 0.57, P < 0.001). CONCLUSIONS: The current study strongly suggests that staging is not reliable when fewer than 10 lymph nodes are examined. The number of examined lymph nodes should be used as a stratification criterion in clinical trials and as an adjustment variable in survival studies. PMID- 12115374 TI - Prognostic relevance of immunohistochemically detected lymph node micrometastasis in patients with gastric carcinoma. AB - BACKGROUND: Micrometastases consisting of one to a few cells in lymph nodes resected during gastrectomy are difficult to identify using conventional hematoxylin and eosin (H&E) stains. It has been shown that immunostaining for cytokeratins is effective in detecting lymph node micrometastasis in a variety of human tumors, but only a few previous reports demonstrated its use in the treatment of patients with early and advanced gastric carcinoma, and those reports had conflicting results. METHODS: In this study, 3625 regional lymph nodes that were dissected in gastrectomy specimens from 153 patients with early stage gastric carcinoma (46 patients) and advanced gastric carcinoma (107 patients) were immunostained with the anticytokeratin cocktail AE1/3 for micrometastasis (median, 23 lymph nodes; range, 8-66 lymph nodes). Micrometastasis (MM) was defined as a single tumor cell or clusters of tumor cells that were missed on conventional examination with H&E stains but were detected by immunostaining with broad-spectrum anticytokeratin antibodies. RESULTS: Lymph node metastasis (LNM) was detected in 609 lymph nodes (17%) by H&E staining. MM was identified in another 191 of the remaining lymph nodes (6.3%) from 75 patients. Twenty-eight of those patients were up-staged. There was a significant correlation between MM and depth of tumor invasion (P < 0.01). Patients with MM had a decreased 5-year survival rate (49%) compared with patients without MM (76%) for both early and advanced gastric carcinoma. The effect of MM on survival was most pronounced for patients in the Stage I and LNM negative group. CONCLUSIONS: Immunohistochemical examination using broad-spectrum anticytokeratin antibodies increased the detection rate of LNM and had a significant impact on staging and survival in patients with gastric carcinoma. PMID- 12115375 TI - Prognostic value of cyclin B1 in patients with esophageal squamous cell carcinoma. AB - BACKGROUND: It has been reported that p53 regulates the G2-M checkpoint transition through cyclin B1, and it has been suggested that p53 plays an important role in the development and progression of various malignancies. The objective of the current study was to clarify the role of the cell cycle regulators, cyclin B1 and p53, in patients with esophageal squamous cell carcinoma (ESCC). METHODS: Tissue samples from 71 patients with ESCC were included in the current study. Expression levels of cyclin B1 and p53 in samples of normal squamous epithelium, dysplasia, and tumor cells from patients with ESCC were analyzed by immunohistochemistry. RESULTS: Several cells in the basement layer of normal epithelium expressed cyclin B1. The number of cyclin B1 positive cells tended to increase as the degree of dysplasia increased from low grade to high grade. More than 20% of tumor cells were cyclin B1 positive in 38 patients (49.3%). Several clinicopathologic parameters, including macroscopic configuration (P < 0.01), pathologic tumor status (P < 0.05), pathologic lymph node status (P < 0.001), pathologic metastatic status (P < 0.01), tumor stage (P < 0.0001), and invasion of lymphatic vessels (P < 0.05), were correlated with the overexpression of cyclin B1. Elevated expression levels of cyclin B1 also were correlated with a poor prognosis in patients with ESCC in univariate analysis (P < 0.0001) and multivariate analysis (P = 0.0135). In contrast, p53 expression exhibited no significant correlation with the level of cyclin B1 expression and was not associated with prognostic parameters in patients with ESCC. CONCLUSIONS: These findings suggest that cyclin B1 is involved in the pathogenesis of carcinoma of the esophagus and that elevated levels of cyclin B1 expression, but not p53 expression, may indicate a poor prognosis for patients with ESCC. PMID- 12115377 TI - Follow-up surveillance strategies for genitourinary malignancies. AB - BACKGROUND: Genitourinary cancers account for more than 20% of all malignancies in the United States. These cancers do not usually yield rapid mortality, thereby necessitating longer-term surveillance strategies. METHODS: A review and analysis of relevant studies were performed. Follow-up strategies are proposed to reflect effective methods to detect recurrent prostate, bladder, renal, and testicular cancers. Cost analysis was performed using Medicare reimbursement rates. RESULTS: For genitourinary tumors, follow-up tests can be planned rationally based on detection rates and patterns. Tumor grade and stage drive follow-up strategies, along with therapeutic implications of detecting a recurrence. Symptomatic recurrences often obviate the need for radiographic tests and can minimize costs. Stage- specific plans for these four urologic malignancies are outlined specifically. CONCLUSIONS: Not all surveillance approaches have been critically tested for follow-up of genitourinary tumors, but ample data are available to propose sound medical and economic strategies. PMID- 12115376 TI - Histologic indices in biopsy specimens for estimating the probability of extended local spread in patients with rectal carcinoma. AB - BACKGROUND: Although precise preoperative assessment of the extent of local cancer spread is important to determine the appropriate treatment strategy, imaging modalities have not been sufficient. The aim of this study was to establish effective preoperative indices that would predict the degree of local spread in patients with rectal carcinoma. METHODS: In specimens from 437 patients with advanced rectal carcinoma, the submucosal horizontal invasive frontal region was examined histologically with reference to three unfavorable characteristics: 1) tumor "budding", 2) poor differentiation, and 3) vascular invasion. In addition, a transanal submucosal biopsy, which targets the tumor edge, was performed on 85 patients to verify the utility of preoperative evaluation of these parameters. RESULTS: Multivariate logistic analysis showed that three unfavorable parameters had independent impact on the degree of nodal involvement. These parameters related significantly to the number of lymph nodes involved, the development of extranodal tumor deposits, circumferential surgical margin involvement, and lateral pelvic lymph node metastases. Regarding patients without unfavorable parameters as a standard, the odds ratio of pelvic recurrence was 1.8 (0.9-3.4) in patients with one unfavorable parameter and 5.3 (2.7-10.2) in patients with multiple unfavorable parameters. Based on the transanal biopsy, the submucosal invasive frontal region could be estimated in 73 patients (85.9%). Among these cases, the multiple unfavorable parameters were relevant to an increased risk of extensive local spread. In addition, pelvic recurrence developed in 36% of patients with multiple unfavorable parameters (no-risk patients, 5%; single-risk patients, 13%). CONCLUSION: Histology in the submucosal invasive frontal region reflects the extent of local spread and can be evaluated preoperatively by transanal biopsy, which should become a useful tool for therapy selection for patients with advanced rectal carcinoma. PMID- 12115378 TI - The economic burden of prostate cancer, California, 1998. AB - BACKGROUND: Prostate cancer is the most common malignancy diagnosed among men in the United States. This article reviews previous studies of the annual cost of all cancers and of prostate cancer in the United States and California and estimates the direct and indirect costs of prostate cancer in California in 1998. METHODS: Hospitalization costs, including costs of primary and secondary diagnoses of prostate cancer, were derived from the California Hospital Discharge data set (CHDS). Charges were converted to costs using hospital specific cost-to charge ratios and an imputed cost for Health Maintenance Organization hospitalizations. Other direct medical costs were derived from the 1997 Medical Expenditure Panel Survey. Indirect mortality costs are the product of the number of deaths and the expected value of a male's future earnings taking into account age at death, earning patterns at successive ages, labor force participation, imputed value of housekeeping services, and a 3% discount rate. RESULTS: Prostate cancer direct health care costs in California were estimated at 180 million dollars, and lost productivity from premature death was estimated at 180 million dollars, for a total cost of 360 million dollars in 1998. The disease is largely one of older men; hospitalization costs account for three-fifths of total direct costs, and Medicare and private health insurance share almost equally in paying for hospital care. CONCLUSIONS: It is critical to identify cost-effective screening efforts that permit early detection of prostate cancer to reduce illness, premature deaths, and the high costs of prostate cancer. PMID- 12115379 TI - Evaluation of urine CYFRA 21-1 for the detection of primary and recurrent bladder carcinoma. AB - BACKGROUND: The urinary concentration of soluble cytokeratin 19 fragments, measured by the CYFRA 21-1 assay, may be used for the noninvasive, early detection of bladder carcinoma. METHODS: This prospective study examined urine samples from 325 patients. The authors included 152 patients who presented with hematuria or irritative voiding symptoms (Group 1), 107 patients who were under surveillance after undergoing transurethral resection of bladder carcinoma (Group 2), 46 patients with urinary tract pathology other than bladder carcinoma (Group 3), and 20 healthy participants (Group 4). The urine concentration of CYFRA 21-1 was measured by an immunoradiometric assay. The patients in Groups 1 and 2 underwent cytoscopy and urine cytopathology. Biopsies were obtained if a tumor was seen on cytoscopy or if there was a suspicion of carcinoma in situ (CIS). RESULTS: The optimal cut-off concentration for the detection of primary bladder tumors, 4.9 microg/L, resulted in a sensitivity of 79.3% and a specificity of 88.6%. The optimal threshold for the detection of recurrent bladder tumors (excluding patients who had been treated with intravesical bacillus Calmette Guerin [BCG]), 4.04 microg/L, resulted in a sensitivity of 76.2% and a specificity of 84.2%. There was no significant advantage for centrifugation of the urine samples or for determination of the creatinine concentration in the urine samples. The CYFRA 21-1 assay of urine samples provided a three-fold greater sensitivity compared with the sensitivity of cytology for detecting Grade 1 transitional cell tumors. CYFRA 21-1 detected 91.9% of Grade 3 tumors, 100% of CIS, and 92.8% of invasive bladder tumors (T2 or higher classification). The CYFRA 21-1 assay detected all tumors that had positive cytology with the exception of only one tumor. Conversely, the assay identified 71% of primary tumors and 65% of recurrent tumors that were missed by cytopathology. Urinary stones, infection, and previous intravesical BCG immunotherapy caused many false positive results. CONCLUSIONS: The urinary CYFRA 21-1 assay is a useful test for the noninvasive detection of bladder carcinoma and for surveillance of patients who were not treated previously with BCG. It may be used in combination with urine cytology and bladder ultrasound. Multi-institutional trials are required to compare the accuracy as well as the cost of this combination of tests with cystoscopy. PMID- 12115380 TI - Facing a prostate cancer diagnosis: who is at risk for increased distress? AB - BACKGROUND: The psychological adaptation to a cancer diagnosis is characterized by significant distress. Although distress levels in cancer patients have been reported to be the highest shortly after diagnosis and before treatment, few attempts have been made to study emotional adjustment during the diagnostic period of prostate cancer patients. This study's purpose was to determine whether differences in distress levels can be attributed to differences in diagnostic status, optimism, and/or coping strategies. METHODS: This study followed patients across 4 weeks, from prebiopsy to 2 weeks postdiagnosis, using these two time points as measurements. Data were collected between 1995 and 1998 at the Miami and Palo Alto VA Medical Center urology clinics. Biopsies were performed on 101 men (ages 46-87) to determine whether prostate cancer was present. These men completed prebiopsy and postdiagnosis questionnaires. RESULTS: Of optimism, coping, and cancer status, the only significant predictor of increased distress at postdiagnosis was dispositional avoidance at prebiopsy for both cancer and noncancer groups. CONCLUSIONS: Although these findings complement other studies linking avoidance with increased distress, the similarities between the cancer and noncancer groups underscore the need to consider both of these groups during the first few weeks of the prostate cancer diagnostic process. PMID- 12115381 TI - Serum arginase activity in postsurgical monitoring of patients with colorectal carcinoma. AB - BACKGROUND: Colorectal carcinoma (CRC) is one of the most common malignancies. In the current work, the role of arginase as a diagnostic marker in patients with recurrent CRC and colorectal liver metastases (CRCLM) was studied. METHODS: Arginase activity was monitored in serum from 40 patients with primary CRC and from 100 patients with CRCLM. Blood was taken before and after patients underwent tumor resection. Studies were conducted for 3 years. RESULTS: Preoperative arginase activity in serum from patients with CRC and CRCLM was much greater compared with the arginase activity in serum from healthy control blood donors. One and two cut-off levels of increased arginase activity were observed in patients with CRC and CRCLM, respectively. After patients underwent tumor resection, the arginase activity decreased to normal values in both patients with CRC and patients with CRCLM. Activity levels remained low in patients who did not develop recurrent CRC or CRCLM (first or second). In patients who developed subsequent recurrences or metastases that appeared after surgery, during 3 years of surveillance, a significant rise in serum arginase activity was observed. The clinical prognosis for patients was worst when the postoperative serum arginase activity was very high, because those patients more often developed second liver metastases or died. CONCLUSIONS: The authors conclude that the determination of serum arginase activity may be a complementary test to confirm the occurrence of CRC and may be useful for the early diagnosis of patients who develop recurrent CRC and/or CRCLM. PMID- 12115382 TI - Comparative immunohistochemical studies of bcl-2 and p53 proteins in benign and malignant ovarian endometriotic cysts. AB - BACKGROUND: A number of histologic and epidemiologic studies have suggested an association between endometriosis and ovarian carcinoma. Some reports have described a transition from endometriosis to atypical endometriosis to carcinoma. Using immunohistochemistry, the authors compared staining patterns in benign endometriotic cysts with ovarian tumors and the endometriotic cyst lining from which they arose, in an attempt to identify sequential or etiologic correlations. METHODS: One hundred thirteen formalin-fixed, paraffin-embedded sections were studied (30 benign ovarian endometriotic cysts, 24 endometriotic cysts containing endometrioid carcinomas, 19 endometriotic cysts harboring clear cell carcinomas, and 40 ovarian papillary serous cystadenocarcinomas). All sections were immunostained with anti-bcl-2 and anti-p53 antibodies using the streptavidin biotin method. RESULTS: bcl-2 was reported to stain 23% of benign endometriotic cysts, 67% of endometrioid carcinomas, 73% of clear cell carcinomas, and 50% of papillary serous carcinomas. Approximately 42% of benign endometriotic lesions adjacent to the endometrioid carcinoma and 73% adjacent to clear cell carcinomas were found to stain for bcl-2 (p = 0.274 [not significant (NS)] and P = 0.008, respectively). p53 staining was negative in the benign endometriotic cyst group and was positive in 37-55% of the group with tumors. p53 staining was positive in 25% of the benign endometriotic lesions next to the endometrioid carcinoma and in 9% of the benign endometriotic lesions next to clear cell carcinoma (P = 0.014 and P = 0.239 [NS], respectively). CONCLUSIONS: The results of the current study suggest that alterations in bcl-2 and p53 may be associated with the malignant transformation of endometriotic cysts. PMID- 12115383 TI - Ribosomal DNA methylation in patients with endometrial carcinoma: an independent prognostic marker. AB - BACKGROUND: Surgery is curative for the majority of patients with endometrial carcinoma: Consequently, the use of adjuvant therapy has been controversial. More effective methods to identify patients who are at risk for recurrence and who would benefit from adjuvant therapy are needed. The objective of this study was to investigate the prognostic significance of ribosomal DNA (rDNA) methylation in patients with endometrial carcinoma. METHODS: rDNA methylation was assessed in 215 endometrial tumors by Southern blot analysis of HpaII-digested DNA using a probe corresponding to the 5.8 and 28S ribosomal subunits. Tumor rDNA methylation status was correlated with patient outcome. RESULTS: The majority of tumors demonstrated high levels of rDNA methylation (rDNA-high; 74%). Both disease free survival and overall survival were significantly worse for patients with low level rDNA methylation (rDNA-low; P < 0.0001). African-American patients were more likely to have rDNA-low tumors than Caucasian patients (P = 0.002). Among the subpopulation of patients with endometrial carcinoma for whom the use of adjuvant therapy is most controversial (148 women with Stage I-II endometrioid tumors), survival was significantly worse for the patients with rDNA-low tumors (P < 0.0001); and, using multivariate analyses, tumor rDNA level was the only significant prognostic factor for both disease free survival and overall survival (hazard ratios, 11.0 and 26.3, respectively; P < 0.01 for both). CONCLUSIONS: rDNA methylation is an independent prognostic indicator for patients with endometrial carcinoma that may serve to identify women with early-stage disease at who are at high risk for recurrence. Racial differences in DNA methylation may explain the historically observed disparity in survival between African-American patients and Caucasian patients. PMID- 12115384 TI - Docetaxel/cisplatin as first-line chemotherapy in patients with head and neck carcinoma: a phase II trial. AB - BACKGROUND: The objective of this Phase II study was to assess the clinical activity and toxicity of docetaxel (D) and cisplatin (P) in patients with locally advanced unresectable, metastatic, or recurrent squamous cell carcinoma of the head and neck (SCCHN). PATIENTS: Of 34 patients, 30 were eligible for treatment with D 80 mg/m(2) on Day 1 and P 70 mg/m(2) on Day 2. Therapy was repeated every 3 weeks. At the start of chemotherapy, the tumors had the following extensions: locoregional, n = 15; distant metastatic, n = 2; and relapse, n = 13. RESULTS: Overall, the rate of objective responses in the population of all eligible patients based on an intention-to-treat analysis was 53%, with a 95% confidence interval (CI; 34.33-71.66%). Two patients had complete disease remission (pathologic), 4 patients had complete disease remission (clinical), 10 patients had partial disease remission, 3 patients had no change in disease status, and 7 patients had disease progression. The duration of objective response was median 5+ months (range 3-8+ months). Eleven patients (37%) had Grade 4 granulocytopenia and three patients (10%) had Grade 3 granulocytopenia (grades were based on the classification of the National Cancer Institute of Canada-Common Toxicity Criteria). Six patients died of septicemia. CONCLUSIONS: Overall, the combination of D and P represents a highly active chemotherapeutic regimen for the treatment of patients with SCCHN. However, because of the high toxicity of this regimen, prophylactic administration of antibiotics and hematopoietic growth factors is essential as is a three-day corticosteroid premedication regimen. Above all, this combination of drugs is not recommended for treatment of patients with a World Health Organization performance status of >1. PMID- 12115385 TI - Uridine phosphorylase is a potential prognostic factor in patients with oral squamous cell carcinoma. AB - BACKGROUND: Uridine phosphorylase (UPase) catalyzes the reversible phosphorolysis of uridine to uracil. The expression levels and the enzymatic activity of UPase are reported to be higher in human solid tumors than in adjacent normal tissues. However, to the authors' knowledge the clinical significance of UPase expression as determined by immunohistochemical analysis has not been demonstrated in human malignancies. METHODS: The authors prepared the antibody against UPase, examined the staining of UPase in 72 cases of oral squamous cell carcinoma (SCC) with immunohistochemical analysis using the antibody, and analyzed its relation to clinical and pathologic factors. RESULTS: UPase stained mainly within the invasive edges of tumors and macrophages. UPase-positive staining was observed in 56 of the 72 tumors (77.8%). High staining of UPase (defined as > 50% of cells in a tumor biopsy that are positive for UPase) in primary tumors frequently was associated with the presence of metastasis to lymph nodes (P = 0.007 by the Fischer exact test) and with lower overall survival (P = 0.03 by the log-rank test). CONCLUSIONS: The high rate of UPase staining in biopsies from patients with oral SCC may be a prognostic marker for these individuals. PMID- 12115386 TI - Squamous cell carcinoma of the oropharynx: surgery, radiation therapy, or both. AB - BACKGROUND: The treatment of patients with squamous cell carcinoma (SCC) of the oropharynx remains controversial. No randomized trial has addressed adequately the question of whether surgery (S), radiation therapy (RT), or combined treatment is most effective. METHODS: Treatment results from North American academic institutions that used S with or without adjuvant RT (S +/- RT) or used RT alone or followed by neck dissection (RT +/- ND) for patients with SCC of the tonsillar region or the base of tongue were compiled through a MEDLINE search (from 1970 to August, 2000) and from the references cited in each report. Studies were eligible for inclusion if they contained direct, actuarial (life-table), or Kaplan-Meier calculations for the following end points: local control, local regional control, 5-year absolute survival, 5-year cause specific survival, or severe or fatal treatment complications. Weighted average results, which took into account series size, were calculated for each end point for the purposes of treatment comparison. Results and conclusions were based on data from 51 reported series, representing the treatment of approximately 6400 patients from the United States and Canada. RESULTS: The results for patients with SCC of the base of tongue who underwent S +/- RT versus RT +/- ND, respectively, were as follows: local control, 79% versus 76% (P = 0.087); local-regional control, 60% versus 69% (P = 0.009); 5-year survival, 49% versus 52% (P = 0.2); 5-year cause specific survival, 62% versus 63% (P = 0.4); severe complications, 32% versus 3.8% (P < 0.001); and fatal complications, 3.5% versus 0.4% (P < 0.001). The results for patients with SCC in the tonsillar region who underwent S +/- RT versus RT +/- ND, respectively, were as follows: local control, 70% versus 68% (P = 0.2); local regional control, 65% versus 69% (P = 0.1); 5-year survival, 47% versus 43% (P = 0.2); 5-year cause specific survival, 57% versus 59% (P = 0.3); severe complications, 23% versus 6% (P < 0.001); and fatal complications, 3.2% versus 0.8% (P < 0.001). CONCLUSIONS: The information in this article provides a useful benchmark for evidence-based counseling of patients with SCC of the oropharynx. The rates of local control, local-regional control, 5-year survival, and 5-year cause specific survival were similar for patients who underwent S +/- RT or RT +/ ND, whereas the rates of severe or fatal complications were significantly greater for the S +/- RT group. Furthermore, available data on the functional consequences of treatment suggest the superiority of RT +/- ND. The authors conclude that RT +/- ND is preferable for the majority of patients with SCC of the oropharynx. PMID- 12115388 TI - Cisplatin, tegafur, and leucovorin: a moderately effective and minimally toxic outpatient neoadjuvant chemotherapy for locally advanced squamous cell carcinoma of the head and neck. AB - BACKGROUND: To evaluate the efficacy and toxicity of cisplatin, tegafur, and leucovorin as neoadjuvant chemotherapy (CT) for patients with advanced, nonmetastatic squamous cell carcinoma of the head and neck (SCCHN). METHODS: Patients with SCCHN according to World Health Organization (WHO) performance status of 2 or less and adequate organ function were enrolled. The CT regimen (PTL) was 50 mg/m(2) cisplatin (P) on Day 1, 800 mg per day oral tegafur (T), and 60 mg per day oral leucovorin (L) for 14 days. The CT was administered at outpatient clinics for 14-day cycles. PTL was initiated with the intent of organ preservation and it was continued for a maximum of six cycles before locoregional therapy. Reevaluation after three cycles led to the termination of CT when the response was less than a partial response. CT was discontinued immediately upon evidence of tumor progression or excessive toxicity. RESULTS: From March 1996 through July 1999, 97 patients were enrolled consecutively. All participants were men with a median age of 56 years (range, 37-70 years). The primary tumor sites were the tongue base, 14, and the hypopharynx, 83. Sixteen percent of the tumors were Stage III, 84% were Stage IV, 62% were Stage T4, and 44% were Stage N2-3. The median number of CT cycles was six. On an intent-to-treat basis, 26 patients (27%) achieved complete responses and 32 patients (33%) achieved partial responses. The overall response rate was 60% (95% confidence interval, 50-70%). The most common toxicities of WHO Grade 3 or higher included (percent of patients): anemia, 8.3%; stomatitis, 6.3%; thrombocytopenia, 3.1%; and vomiting, 3.1%. With a median follow-up period of 3 years, the overall survival and disease free survival rates were 40% and 38%, respectively. Organ preservation was achieved in 70% (29 of 37) of the surviving patients. CONCLUSION: The outpatient PTL regimen was a moderately effective and minimally toxic CT for SCCHN. PTL should be studied further in combination with other active agents or radiotherapy for patients with SCCHN. PMID- 12115387 TI - The role of diet and specific micronutrients in the etiology of oral carcinoma. AB - BACKGROUND: Carcinoma of the oral cavity is one of the most common cancers worldwide. Tobacco smoking and the consumption of alcoholic beverages are significant risk factors but to the authors' knowledge the role of nutrition is not adequately understood. The authors undertook an epidemiologic study of oral carcinoma occurring in Greece, where tobacco smoking and alcohol consumption are common but the incidence of the disease is among the lowest reported in Europe. METHODS: One hundred six patients with histologically confirmed incident oral carcinoma and an equal number of control subjects matched for age and gender were studied. Dietary information was assessed through a validated extensive food frequency questionnaire and the data were analyzed using conditional logistic regression. RESULTS: After adjustment for energy intake, tobacco smoking, and alcohol consumption, there was evidence that the consumption of cereals, fruits, dairy products, and added lipids (which in Greece are represented mostly by olive oil) was found to be associated inversely with the risk of oral carcinoma. Only with respect to meat and meat products was there adequate evidence of a positive association with the risk of oral carcinoma. Among the micronutrients studied, riboflavin, magnesium, and iron appeared to be correlated inversely with the disease. CONCLUSIONS: Fruits, cereals, dairy products, and olive oil appear to convey protection against oral carcinoma and their effects may be mediated through higher intakes of riboflavin, iron, and magnesium. The low incidence of oral carcinoma reported in Greece may be explained in part by the higher consumption of the food groups and micronutrients that appear to protect against the disease. PMID- 12115389 TI - Chronic myelogenous leukemia in T cell lymphoid blastic phase achieving durable complete cytogenetic and molecular remission with imatinib mesylate (STI571; Gleevec) therapy. AB - BACKGROUND: A T cell lymphoid blastic phase of chronic myelogenous leukemia (CML) is a rare occurrence, with only a few reported cases worldwide. Standard therapy for such patients is undetermined. Imatinib mesylate, a Bcr-Abl tyrosine kinase inhibitor, has shown activity in CML. METHODS: The authors report on a patient with CML and marrow as well as extramedullary nodal T cell lymphoid blastic phase who was treated with imatinib mesylate. RESULTS: The patient achieved complete morphologic and cytogenetic remission within two months of therapy. Competitive quantitative polymerase chain reaction analysis of marrow cells was negative after 15 months. Response had lasted for 26+ months at the time of writing. CONCLUSIONS: The current data suggest that imatinib mesylate may produce long term event free survival in patients with T-cell lymphoid blastic phase CML. Its potential role alone or in combinations should be further explored in this condition. PMID- 12115390 TI - Adenosquamous carcinoma of the gallbladder warrants resection only if curative resection is feasible. AB - BACKGROUND: Adenosquamous/squamous cell carcinoma of the gallbladder generally has been considered a lethal disease. The objective of this study was to clarify the effectiveness of resection for patients with adenosquamous/squamous cell carcinoma of the gallbladder. METHODS: Twenty-nine patients who underwent resection for either adenosquamous carcinoma (n = 28 patients) or squamous cell carcinoma (n = 1 patient) were analyzed retrospectively. Sixteen patients underwent radical resection, whereas the other patients underwent primary tumor resection alone. To elucidate the factors that influenced postresectional survival, 10 variables (age, gender, presence or absence of gallstones, size of the primary tumor, lymphatic vessel invasion, blood vessel invasion, perineural invasion, TNM stage, residual tumor status, and type of resection) were examined. RESULTS: Twenty-three patients (79.3%) were categorized with T3 or T4 tumors that invaded adjacent organs. The outcome after undergoing radical resection (cumulative 5-year survival rate, 48.6%) was significantly better compared with the outcome of patients after undergoing primary tumor resection alone (cumulative 3-year survival rate, 7.7%; P = 0.004). The outcome after undergoing resection was better in 14 patients who had no residual tumor (cumulative 5-year survival rate, 62.9%) compared with the outcome in 15 patients who had residual tumor (cumulative 5-year survival rate, 0%; P < 0.001). Univariate analysis revealed that residual tumor status (P < 0.001), type of resection (P = 0.004), patient age (P = 0.012), and blood vessel invasion (P = 0.017) were significant prognostic factors. Residual tumor status (P = 0.026) was the only significant independent prognostic factor. CONCLUSIONS: Adenosquamous/squamous cell carcinoma of the gallbladder warrants resection only if potentially curative (R0) resection is feasible. PMID- 12115391 TI - Mental state as a possible independent prognostic variable for survival in patients with advanced lung carcinoma. AB - BACKGROUND: Although psychologic factors have been reported to influence the progression of cancer, this theory remains controversial. A prospective study of patients with advanced lung carcinoma was performed to explore the influence of the patient's mental state on survival. METHODS: The patient's mental state was assessed with the Tokyo University Egogram. In a preliminary study, the egograms of long-term survivors (survival > 3 years) with TNM Stage IIIB or Stage IV lung carcinoma were compared with the egograms of consecutive, newly diagnosed lung carcinoma patients (controls). Next, in a prospective study, 123 patients with nonsmall cell lung carcinoma and 56 patients with small cell lung carcinoma (Stage IIIB or Stage IV; Eastern Cooperative Oncology Group performance status of 0 or 1) completed the egogram. Based on the results of the preliminary study, the subjects in the prospective study were divided into Group A (Free Child [FC] >or= 50th percentile and Adapted Child [AC] < 50th percentile) and Group B (FC < 50 percentile or AC >or= 50 percentile). The survival of the two groups was compared. The Cox proportional hazards model was used to determine the joint effect of the patient's mental state and other prognostic factors. RESULTS: In the preliminary study, the FC score of the long-term survivors was significantly higher and the AC score was significantly lower than those of the controls. In the prospective study, the survival of Group A was significantly longer than that of Group B both in the nonsmall cell lung carcinoma and small cell lung carcinoma patients (P = 0.002 and P = 0.005, respectively, by the log-rank test). Multivariate analysis demonstrated that after adjustment for clinical factors, being in Group A was a significant predictor of survival both in the nonsmall cell and small cell lung carcinoma patients. CONCLUSIONS: The results of the current study demonstrate that the mental state of the patient as assessed by the egogram may have prognostic significance in patients with advanced lung carcinoma. PMID- 12115392 TI - Intraoperative evaluation of sentinel lymph nodes for metastatic melanoma by imprint cytology. AB - BACKGROUND: Sentinel lymph node (SLN) biopsy has revolutionized lymph node staging in patients with malignant melanoma. Intraoperative evaluation is a new addition to the SLN procedure that allows for a one-step regional lymph node dissection to be performed when the SLN biopsy findings are positive. To date, several studies have evaluated the use of intraoperative frozen sectioning to evaluate the SLN in patients with melanoma. The literature pertaining to the use of intraoperative imprint cytology (IIC) to evaluate the SLN in melanoma patients is scant and to the authors' knowledge studies published to date are relatively small. The purpose of the current study was to evaluate the utility of IIC in patients undergoing SLN for melanoma. METHODS: A total of 235 SLN biopsies from 93 patients with malignant melanoma and 3 patients with atypical Spitz nevi were examined by IIC after SLN biopsy using a double indicator technique. The SLNs were bisected and a pair of imprints were made from each half. One imprint from each half was stained with hematoxylin and eosin (H & E) whereas its counterpart was stained with Diff-Quik. Paraffin-embedded permanent sections were examined using multiple H & E stained sections from the SLNs in conjunction with immunohistochemical staining for S-100 and HMB-45 proteins. RESULTS: A total of 235 SLNs were excised from 93 patients (2.5 SLNs per patient). On a per patient basis, metastases were identified in 21 patients (23%) on permanent section evaluation. Of these 21 patients, 8 were detected by IIC (sensitivity of 38%). The negative predictive value was 85%. No false-positive results were identified (specificity of 100%). The positive predictive value was 100%. The overall accuracy of the intraoperative evaluation was 86%. Patients found to have positive SLNs by IIC went on to undergo lymphadenectomy under the same anesthetic. CONCLUSIONS: The sensitivity and specificity of IIC are similar to those of intraoperative frozen-section evaluation. Therefore, IIC appears to be a viable alternative to frozen sectioning when intraoperative evaluation is required. IIC evaluation of SLN makes a single surgical procedure possible for patients with malignant melanoma who are undergoing SLN. PMID- 12115393 TI - Low natural killer activity and central nervous system disease as a high-risk prognostic indicator in young patients with hemophagocytic lymphohistiocytosis. AB - BACKGROUND: Familial hemophagocytic lymphohistiocytosis HLH (FHL) is fatal, unless patients are rescued with hematopoietic stem cell transplantation (SCT). Although the molecular identification of FHL now is possible at least in part from perforin gene study, many cases escape detection or never are tested due to the lack of specific hallmarks, making diagnosis difficult. To the authors' knowledge, it remains to be determined whether persistently low natural killer cell (NK) activity and a high incidence of central nervous system (CNS) disease increase the probability of FHL. METHODS: The authors analyzed 42 HLH patients age < 2 years, 13 of whom developed overt CNS disease and 5 of whom demonstrated persistently deficient NK activity (Group 1). The remaining 24 patients had no CNS disease and had NK activity of moderate decrease to within the normal range (Group 2). RESULTS: In Group 1, CNS symptoms were detected in 6 cases within 1 month and between 4.5-9 months in 6 other patients. In these cases, spotty lesions demonstrating a high T2 signal in the white matter were noted on brain magnetic resonance imaging. The survival was significantly poor for patients in Group 1 unless they were rescued with SCT, which was performed in 5 of the 13 patients with CNS disease and in all 5 patients with persistent NK activity deficiency. SCT was successful in 9 patients, with no CNS sequelae reported after the transplantation. Conversely, the prognosis of the 24 patients in Group 2 was better and only 1 patient required SCT. CONCLUSIONS: Very young HLH patients (age < 2 years) who are at high risk of fatal FHL with persistently deficient NK activity and/or overt CNS disease require appropriate SCT to reverse CNS disease and achieve a complete cure. PMID- 12115394 TI - Prevention of cisplatin-induced acute and delayed emesis by the selective neurokinin-1 antagonists, L-758,298 and MK-869. AB - BACKGROUND: Recent studies have suggested that antiemetic therapy with a triple combination of the neurokinin-1 receptor antagonist MK-869, a serotonin (5-HT(3)) antagonist, and dexamethasone provides enhanced control of cisplatin-induced emesis compared with standard therapy regimens. The authors compared the antiemetic activity of a dual combination of MK-869 and dexamethasone with that of a standard dual combination of ondansetron and dexamethasone to characterize further the efficacy and tolerability profile of MK-869. METHODS: This was a multicenter, double-blind, randomized, active agent-controlled study of 177 cisplatin-naive patients with malignant disease. On Day 1, MK-869 was given intravenously as its water-soluble prodrug, L-758,298. Patients were randomized to one of three groups as follows. Group I received L-758,298 100 mg intravenously (i.v.), then dexamethasone 20 mg i.v., and cisplatin >or= 70 mg/m(2) on Day 1 followed by 300 mg MK-869 (tablet) orally on Days 2-5; Group II received L-758,298 100 mg i.v., then dexamethasone 20 mg i.v., and cisplatin >or= 70 mg/m(2) on Day 1 followed by placebo on Days 2-5; and Group III received ondansetron 32 mg i.v., then dexamethasone 20 mg i.v., and cisplatin >or= 70 mg/m(2) on Day 1 followed by placebo on Days 2-5. Emesis was recorded over Days 1 5 in a diary. Nausea was assessed every 24 hours by visual analog scale. Additional medication was available for emesis or nausea at any time. The primary efficacy parameters of interest were the proportion of patients without emesis and the proportion without emesis or rescue therapy on Day 1 (acute phase) and on Days 2-5 (delayed phase). RESULTS: No serious adverse events were attributed to L 758,298 or MK-869. On Day 1, the proportions of patients with no emesis and no use of rescue medication were 44% of patients in Group I, 36% of patients in Group II, 40% of patients in Groups I and II combined, and 83% of patients in Group III (P < 0.001 for Group III vs. the combined Groups I and II). The proportions of patients with no emesis and no use of rescue medication on Days 2 5 were 59% of patients in Group I, 46% of patients in Group II, and 38% of patients in Group III (P < 0.05 for Group I vs. Group III). The proportions of patients who were without emesis on Day 1 were 49% of patients in Group I, 47% of patients in Group II, and 84% of patients in Group III (P < 0.01 for Group I or II vs. Group III). On Days 2-5, however, the proportions of patients who were without emesis on Days 2-5 were 65% of patients in Group I, 61% of patients in Group II, and 41% of patients in Group III (P < 0.05 for Group I or II vs. Group III). Nausea scores in the acute phase were lower for Group III than for Group I, Group II, or Groups I and II combined (P < 0.05), although there was no significant difference among groups either for the delayed phase or overall for Days 1-5. CONCLUSIONS: Although the L-758,298 and dexamethasone combination reduced acute (Day 1) emesis compared with historic rates, dual therapy with ondansetron and dexamethasone was superior in controlling acute emesis. Continued dosing with MK-869 may enhance control of other measures of delayed emesis, such as the use of rescue medication, although confirmation is required before a definitive conclusion may be drawn. MK-869 given as dual therapy with dexamethasone was superior to ondansetron with dexamethasone for the control of delayed emesis (Days 2-5) and control of the need for rescue medication on Days 2 5. PMID- 12115395 TI - Quantitation of GD2 synthase mRNA by real-time reverse transcription-polymerase chain reaction: utility in bone marrow purging of neuroblastoma by anti-GD2 antibody 3F8. AB - BACKGROUND: Antigen ganglioside GD2 is expressed abundantly on neuroblastoma (NB) cells. Anti-GD2 monoclonal antibody (MoAb) 3F8 kills NB cells by complement dependent cytotoxicity and antibody-dependent cellular cytotoxicity. Its utility in bone marrow (BM) purging is evaluated by a real-time reverse transcription polymerase chain reaction (RT-PCR) assay to quantify the mRNA of GD2 synthase, the key enzyme in GD2 synthesis. METHODS: From 1990 to 1993, 10 patients with relapsed/refractory Stage 4 NB participated in a pilot study. In these patients, MoAb 3F8 was used to purge tumor cells from harvested BM that had 5% or less tumor content by immunofluorescence (IF). Subsequently, 31 Stage 4 NB patients who underwent treatment on the N7 protocol (1994-1999) had their BM, which was in remission, purged by 3F8 before (131)I-3F8 myeloablative radioimmunotherapy. GD2 positive tumor cells before and after purging were quantified by real-time quantitative RT-PCR of GD2 synthase. RESULTS: GD2 positivity by IF was found before purging in six of eight patients in the pilot study. Five of six patients became negative postpurging. Of 31 patients on the N7 protocol, the more sensitive real-time quantitative RT-PCR detected GD2 synthase mRNA in the BM samples of 7 patients even though the prepurge BM samples were negative by histology and IF. Six of the seven BM samples became negative after 3F8 purging. Marker positivity before purging was statistically significant in predicting overall survival (P = 0.04), but not progression-free survival (P = 0.1). In vitro hematopoietic stem cell recovery and the median time to engraftment were acceptable. CONCLUSION: Tumor cell depletion quantified by real-time RT-PCR demonstrated efficacy of MoAb 3F8 in BM purging. PMID- 12115397 TI - Incidence of cancer among patients with knee implants in Sweden, 1980-1994. AB - BACKGROUND: As knee implants become more common, it is important to study their potential health risks. We investigated cancer occurrence in a nationwide population-based cohort of 30,011 patients who underwent knee replacement surgery in Sweden from 1980 to 1994. METHODS: Patients were followed from 1 year after the date of their surgery through December 31, 1995, accruing 122,616 person years of observation. The average follow-up time was 4.3 years, with 2365 patients followed for 10 years or more. RESULTS: Overall cancer incidence was not elevated compared with the general population of Sweden (standardized incidence ratio [SIR] = 1.03; 95% confidence interval [CI] = 0.98-1.08). A reduced rate for all respiratory cancers (SIR = 0.73; 95% CI = 0.59-0.91) and for lung cancer (SIR = 0.73; 95% CI = 0.58-0.91) was found among both men and women. Elevated rates were found for prostate (SIR = 1.20; 95% CI = 1.06-1.34) and bone cancer (SIR = 6.00; 95% CI = 1.24-17.52) in men. The bone cancer excess was based on three observed cases, two of which occurred at a site unrelated to the implant and the site of the third tumor is unknown. Rates of connective tissue cancer and leukemia-lymphoma were not elevated significantly among knee implant recipients. Long-term follow-up (>or= 10 years) did not show a significant excess risk for all cancer (SIR = 1.10; 95% CI = 0.86-1.38) or for any site-specific cancer, including bone cancer, lymphoma, or leukemia. Subgroup analyses for patients with rheumatoid arthritis produced results similar to the overall results. CONCLUSIONS: This epidemiologic study of cancer risk among patients with knee implants is the largest to date. It provides evidence that the incidence of cancer among patients with knee implants is similar to that of the general population. Continued follow-up of this cohort is warranted to evaluate further potential long-term effects of these implants. PMID- 12115396 TI - A prospective study evaluating the response of patients with unrelieved cancer pain to parenteral opioids. AB - BACKGROUND: The initiation of continuous parenteral (subcutaneous or intravenous) opioids or a change of opioid (opioid rotation) are treatment options for patients who fail on oral or transdermal opioids. There are insufficient data on the efficacy of these strategies, and comparative data are unavailable. METHODS: The authors prospectively evaluated the efficacy of the start of parenteral opioids in 100 patients with cancer pain who failed on conventional opioids. Pain intensity was rated at rest and during movement from 0 to 10 and was categorized as mild (0-4), moderate (5-6), or severe (7-10): Clinically important pain control was defined as a decrease >or= 2 points in pain intensity and pain intensity < 7. Pain control was evaluated on the second day and again when a clinical decision was made to continue or change parenteral opioid treatment after a median of 6 days. The presence of side effects (absent, mild, moderate, or severe) was evaluated. RESULTS: The mean pain intensity at rest decreased significantly from 6.3 to 4.4 at 48 hours and to 3.4 at the end of treatment. The mean pain intensity during movement decreased significantly from 8.4 to 5.7 at 48 hours and to 4.6 at the end of treatment. Clinically important pain control at rest was seen in 52% of patients at 48 hours, in 71% of patients at the end of treatment; and clinically important pain control during movement was seen in 43% of patients at 48 hours and in 61% of patients at the end of treatment. The proportion of patients with mild pain increased significantly both at rest and during movement. Side effects were present in 78% of patients, and they resolved completely in 32% of patients. The median intravenous morphine equivalent dose increased from 80 mg per day to 135 mg per day at 48 hours and to 201 mg per day at the end of treatment. Results were not different for opioid rotation or for change of route only, nor did the start of antitumour treatment influence the results. In 34% of patients, it was decided to rotate to a second-line parenteral opioid or to start either spinal analgesia or a sedation procedure after a median of 6 days. During follow-up, 18% of patients who were dismissed with parenteral opioids (and 6% of all patients) needed a further change of treatment. CONCLUSIONS: Parenteral opioids improved the balance between analgesia and side effects in patients with cancer pain who failed on conventional opioids, with an important improvement seen in 71% of patients. On the basis of this study, it is concluded that parenteral opioids are a good alternative to spinal opioids. Furthermore, it is suggested that a change of route alone is as effective as opioid rotation. PMID- 12115398 TI - Familial breast carcinoma risks by morphology: a nationwide epidemiologic study from Sweden. AB - BACKGROUND: Familial risks in patients with breast carcinoma have not been assessed by morphologic types of medically verified cancers. Reliable data on familial risks would help to establish prevention programs and guide clinical decisions. METHODS: We used the nationwide Swedish Family-Cancer Database to calculate standardized incidence ratios (SIRs) and 95% confidence intervals (CIs) for invasive and in situ breast carcinomas in women with mothers and sisters. This database has information on 10.2 million individuals and on more than 13,000 morphology-specific breast carcinomas. RESULTS: SIRs for all invasive breast carcinomas were 1.82 (95% CI 1.71-1.93) for breast carcinoma in the mother and 1.89 (1.70-2.01) for breast carcinoma in a sister. The respective risks were 1.81 and 1.85 for a mother and sisters with ductal breast carcinoma. The SIRs were equally for lobular, tubuloductal, comedo, and mucinous breast carcinomas. However, the SIRs for lobular carcinoma were lower than those for the ductal type, whereas the opposite trend was noted for the comedo and mucinous type; none of the differences were significant. The risks for all morphologic types were highest when both a mother and a sister were affected, SIR 3.19 (2.36-4.22). The risks for in situ breast carcinomas were 2.09 (1.78-2.44) for an affected mother, 2.24 (1.88-2.85) for an affected sister, and 5.23 (2.59-9.39) when both a mother and a sister were affected. CONCLUSIONS: The data suggest that the familial risk of breast carcinoma is independent of the morphologic type. The higher risks in in situ cancer may be due to medical surveillance. The risks were identical from a mother or sister proband, suggesting that recessive effects are unlikely as a heritable cause of breast carcinoma. PMID- 12115400 TI - Protein-protein interactions in concentrated electrolyte solutions. AB - Protein-protein interactions were measured for ovalbumin and for lysozyme in aqueous salt solutions. Protein-protein interactions are correlated with a proposed potential of mean force equal to the free energy to desolvate the protein surface that is made inaccessible to the solvent due to the protein protein interaction. This energy is calculated from the surface free energy of the protein that is determined from protein-salt preferential-interaction parameter measurements. In classical salting-out behavior, the protein-salt preferential interaction is unfavorable. Because addition of salt raises the surface free energy of the protein according to the surface-tension increment of the salt, protein-protein attraction increases, leading to a reduction in solubility. When the surface chemistry of proteins is altered by binding of a specific ion, salting-in is observed when the interactions between (kosmotrope) ion-protein complexes are more repulsive than those between the uncomplexed proteins. However, salting-out is observed when interactions between (chaotrope) ion-protein complexes are more attractive than those of the uncomplexed proteins. PMID- 12115401 TI - Ultra scaledown to predict filtering centrifugation of secreted antibody fragments from fungal broth. AB - Extracellularly expressed anti-hen egg lysozyme single-chain antibody fragments (scFv) produced by Aspergillus awamori were recovered using filtering centrifugation. Two filtering centrifuges with 0.5- and 30-L capacities were used to represent laboratory- and pilot-scale equipment, respectively. Critical regime analysis using the computational fluid dynamics (CFD) technique provided information about the local energy dissipation rates in both units. Experimental data indicated loss of scFv activity for energy dissipation rates above about 2.0 x 10(4) W kg(-1). This loss of activity increased in the presence of gas-liquid interfaces during filtering centrifugation. An ultra scaledown filtering centrifuge with a maximum working volume of 35 mL was designed to mimic the operating conditions identified by the critical regime analysis for the laboratory- and pilot-plant-scale units. The recovered scFv activity levels and the separation performance of the three units were comparable when operated at equal maximum energy dissipation rates. PMID- 12115402 TI - Effect of complexing agents on reduction of Cr(VI) by Desulfovibrio vulgaris ATCC 29579. AB - The reduction of Cr(VI) at the expense of molecular hydrogen was studied using resting cells of Desulfovibrio vulgaris ATCC 29579 in anaerobic resting cell suspensions in MOPS buffer. Bioreduction occurred only in the presence of ligands or chelating agents (CO32-, citrate, NTA, EDTA, DTPA). The stimulatory effect of these ligands on the rate of Cr(VI) reduction was correlated (r = 0.988) with the strength of the ligand/chelate complex of Cr(III). The data are examined with respect to likely solution and redox equilibria in the ionic matrix of the carrier solution, and with respect to the potential for bioremediation of Cr(VI). PMID- 12115403 TI - The response of GS-NS0 myeloma cells to single and multiple pH perturbations. AB - Animal cells are cultured in several types of vessels at laboratory and industrial scale the most common being the stirred tank and the air-lift. Economically, it is preferable to culture animal cells at the largest possible scale but the perceived sensitivity of animal cells to hydrodynamic shear has, until now, limited the aeration and agitation rates used. This has been reported to cause inhomogeneities in operational parameters such as dissolved oxygen concentration, temperature and pH. pH is of special interest during the latter stages of many animal cell fermentation because alkali additions, used for pH control, can cause large local pH perturbations of varying size and duration. The effect of single and multiple pH perturbations on the cell growth of a widely used GS-NS0 mouse myeloma cell line grown in batch culture was investigated. The effect of perturbation amplitude and duration was investigated using a single stirred tank reactor (STR). In the single STR system cells were subjected to one pH 8.0 or 9.0 perturbation ranging in duration from 0-90 minutes. No measurable decrease in viable cell number was seen for pH 8.0 perturbations of any duration whereas pH 9.0 perturbations lasting for 10 minutes caused a 15% decrease in viable cell number. The proportion of viable cells decreased with increasing perturbation time and a 90-minute exposure killed all of the cells. The effect of multiple pH perturbations on GS-NS0 cells was investigated using two connected STR's. More specifically the number of perturbations and the perturbation frequency were investigated. Cells were subjected to between 0 and 100 perturbations at pH 8.0; the time between each perturbation (frequency) was 6 minutes and each perturbation lasted for 200 seconds. Viable cell number decreased with increasing perturbation number, with 100 perturbations causing death of 27.5% of cells. Cells were also exposed to 10 perturbations at pH 9.0, each of 200 second duration at frequencies of either 6, 18 or 60 minutes. Approximately 8 times more cells were killed with perturbations at a 6-minute frequency (28.3% cell death) than at a 60-minute frequency (3.4% cell death). PMID- 12115404 TI - The effect of ajmalicine spiking and resin addition timing on the production of indole alkaloids from Catharanthus roseus cell cultures. AB - The potential for the feedback inhibition of indole alkaloid synthesis was investigated by spiking suspension cultures of Catharanthus roseus with 0, 9, or 18 mg/L ajmalicine on day 0. The production of ajmalicine, catharanthine, and serpentine were inhibited in a dose-dependent manner. The inhibition was transient as the exogenous ajmalicine was ultimately either metabolized in the medium or within the cell. The addition of neutral resin has previously been shown to enhance ajmalicine production. To minimize product inhibition and product metabolism, Amberlite XAD-7 resin was added to immobilized cultures of C. roseus starting on either day 0, 5, or 15, and fresh resin was exchanged for spent resin every 5 days. The addition of resin did not decrease the viability of the culture. Growth was reduced only in cultures with resin added on day 0. Alkaloid production was enhanced to different extents by the timing of resin addition, suggesting that feedback inhibition or product metabolism was present throughout the culture period. Ajmalicine recovery was nearly 100% when the resin was added initially either on day 0 or day 5. Ajmalicine recovery was reduced to 55% when the resin was added later in the culture period starting on day 15, presumably because of resin saturation or the inaccessibility of alkaloids trapped in the vacuole. Delaying the addition of XAD-7 resin until 5 days after the start of the culture resulted in the highest improvement in ajmalicine production, i.e approximately 70% and also resulted in the complete recovery of ajmalicine from the cell. PMID- 12115405 TI - A two-phase model for water and heat transfer within an intermittently-mixed solid-state fermentation bioreactor with forced aeration. AB - A two-phase dynamic model is developed that describes heat and mass transfer in intermittently-mixed solid-state fermentation bioreactors. The model predicts that in the regions of the bed near the air inlet there can be significant differences in the air and solid temperatures, while in the remainder of the bed the gas and solid phases are much closer to equilibrium, although there can be differences in water activity of around 0.05. The increase in the temperature of the gas as it flows through the bed means that it is impossible to prevent the bed from drying out, even if saturated air is used at the air inlet. The substrate can dry to water activities that severely limit growth, unless the bed is intermittently mixed, with the addition of water to bring the water activity back to the desired value. Under the conditions assumed for the simulation, which was designed to mimic the growth of Aspergillus niger on corn, two mixing events were necessary, one at 17.4 and the other at 27.9 h. Even though such a strategy can minimize the restriction of growth by water-limitation, temperature limitation remains a problem due to the rapid heating dynamics. The model is obviously a useful tool that can be used to guide scale-up and to test control strategies. Such a model, describing the non-equilibrium situation between the gas and solid phases, has not previously been proposed for solid-state fermentation bioreactors. Models in the literature that assume gas-solid temperature and moisture equilibrium cannot describe the large temperature differences between the gas and solid phase which occur within the bed near the air inlet. PMID- 12115406 TI - Differential rates of gene expression monitored by green fluorescent protein. AB - The use of green fluorescent protein (GFP) as a reporter gene has made a broad impact in several areas, especially in studies of protein trafficking, localization, and expression analysis. GFP's many advantages are that it is small, autocatalytic, and does not require fixation, cell disruption, or the addition of cofactors or substrates. Two characteristics of GFP, extreme stability and chromophore cyclization lag time, pose a hindrance to the application of GFP as a real-time gene expression reporter in bioprocess applications. In this report, we present analytical methods that overcome these problems and enable the temporal visualization of discrete gene regulatory events. The approach we present measures the rate of change in GFP fluorescence, which in turn reflects the rate of gene expression. We conducted fermentation and microplate experiments using a protein synthesis inhibitor to illustrate the feasibility of this system. Additional experiments using the classic gene regulation of the araBAD operon show the utility of GFP as a near real-time indicator of gene regulation. With repetitive induction and repression of the arabinose promoter, the differential rate of GFP fluorescence emission shows corresponding cyclical changes during the culture. PMID- 12115407 TI - Fermentation strategies for improved heterologous expression of laccase in Pichia pastoris. AB - Improved expression of recombinant laccase by Pichia pastoris carrying the lcc1 cDNA isolated from Trametes versicolor was achieved by optimization of the cultivation conditions in a fermentor equipped with a methanol sensor system. The results indicated that the activity obtained in fermentor cultivations was at least 7 times higher than in shake-flask cultures. Three different strategies for fermentor cultivations were compared: A (30 degrees C, 1.0% methanol), B (20 degrees C, 1.0% methanol), and C (20 degrees C, 0.5% methanol). The laccase activity, particularly the specific activity, could be improved by decreasing the cultivation temperature. The mechanisms behind the temperature effect on the laccase activity may be ascribed to poor stability, release of more proteases from dead cells, and folding problems at higher temperature. The results showed that the methanol concentration had a marked effect on the production of active heterologous laccase. A fivefold higher volumetric laccase activity was obtained when the methanol concentration was kept at 0.5% instead of 1.0%. The detrimental effect of methanol on the production of recombinant laccase may be attributed to lower laccase stability, a higher proteolytic activity, and folding problems due to higher growth rate at 1.0% methanol. PMID- 12115409 TI - Anchorage of cyclodextrin glucanotransferase on the outer membrane of Escherichia coli. AB - The gene encoding cyclodextrin glucanotransferase (CGTase) was successfully cloned from B. macerans by PCR. A recombinant plasmid pCS005 with a gene encoding the Lpp-OmpA-CGTase trifusion protein was constructed and transformed into E. coli for the surface display of CGTase. Results of immunoblotting analysis and protease accessibility on the fractionated cell membranes confirmed that the Lpp OmpA-CGTase trifusion protein was successfully anchored on the outer membrane of E. coli. However, only 50% of the membrane-anchored trifusion proteins were displayed on the outer surface of E. coli with the remaining 50% un-translocated. The low efficiency of surface display is attributed to the large size of CGTase. Only a trace amount of CGTase activity was detected for both the whole cells and the cell debris fractions. Because the results of the protease accessibility study suggested that the trypsin-resistant conformation of CGTase was preserved in the membrane-anchored CGTase, we believe that the lack of enzyme activity is mainly due to the inaccessibility of the CGTase active site, near the N-terminus, for substrate molecules. It can be estimated that the critical size for surface display of protein in E. coli is approximately 70 kDa. PMID- 12115408 TI - Protein-mediated isolation of plasmid DNA by a zinc finger-glutathione S transferase affinity linker. AB - The sequence-specific affinity chromatographic isolation of plasmid DNA from crude lysates of E. coli DH5alpha fermentations is addressed. A zinc finger-GST fusion protein that binds a synthetic oligonucleotide cassette containing the appropriate DNA recognition sequence is described. This cassette was inserted into the SmaI site of pUC19 to enable the affinity isolation of the plasmid. It is shown that zinc finger-GST fusion proteins can bind both their DNA recognition sequence and a glutathione-derivatized solid support simultaneously. Furthermore, a simple procedure for the isolation of such plasmids from clarified cell lysates is demonstrated. Cell lysates were clarified by cross-flow Dean vortex microfiltration, and the permeate was incubated with zinc finger-GST fusion protein. The resulting complex was adsorbed directly onto glutathione-Sepharose. Analysis of the glutathione-eluted complex showed that plasmid DNA had been recovered, largely free from contamination by genomic DNA or bacterial cell proteins. PMID- 12115410 TI - Polymer surfaces derivatized with poly(vinyl-N-hexylpyridinium) kill airborne and waterborne bacteria. AB - A facile methodology has been developed for covalently derivatizing the surfaces of common materials with a designed antibacterial polycation, poly(vinyl-N pyridinium bromide), wherein the first, key step involves surface coating with a nanolayer of silica. Various commercial synthetic polymers derivatized in this manner become bactericidal-they kill up to 99% of deposited, from either an aerosol or an aqueous suspension, Gram-positive and Gram-negative bacteria on contact. PMID- 12115411 TI - Selective recovery of lactate dehydrogenase using affinity foam. AB - Selective isolation of lactate dehydrogenase (LDH) from porcine muscle extract was studied using foam generated from the vigorous stirring of a non-ionic surfactant, Triton X-114 derivatized with Cibacron blue. The cloud point of the surfactant-dye conjugate was higher than that of the native Triton X-114, and also the foam prepared from the affinity surfactant was more rigid taking a longer time to collapse. The equilibrium dissociation constant between pure LDH and surfactant-dye conjugate was 5.0 microM as compared to the value of 2.2 microM for the enzyme and free dye as measured by differential spectroscopy. The isolation procedure involved mixing of the porcine muscle extract with the affinity foam, separating and collapsing the foam, and warming the solution formed to 37 degrees C to yield the surfactant-dye phase and an aqueous phase containing the enzyme. The effect of surfactant concentration and protein load on enzyme recovery and purification was investigated. Under optimal conditions, LDH was quantitatively recovered with high purification factor in a very short time. Both recovery and purification were higher when foam prepared from an equivalent mixture of surfactant-dye conjugate and unmodified surfactant was used. The selectivity of interaction between LDH and detergent-dye conjugate was confirmed by lowered recovery when NADH was included during the binding step. PMID- 12115412 TI - Low-temperature electron microscopy for the study of polysaccharide ultrastructures in hydrogels. I. Theoretical and technical considerations. AB - The high-pressure freezing (HPF) technique was applied to the cryo-immobilization of alginate gels and the quality of the freezing analyzed on a TEM by comparison of the segregation pattern of samples of decreasing thickness. Dynamic simulations of heat transfer within an idealized slab of pure water surrounded by two walls of aluminium were performed to illustrate the effect of the heat transfer coefficient by convection on the cooling rate of the sample. Heat transfer coefficients in liquid nitrogen and liquid propane at ambient pressure were measured using a carefully characterized thermocouple and the values incorporated as parameters in heat-transfer simulations to compare the efficiency of the plunge-freezing technique with the high-pressure freezing technique. Values of the heat-transfer coefficient in liquid nitrogen and liquid propane, calculated between 273 K and 173 K were 670 and 18420 W/m(2)/K, respectively. Based on TEM observations and the results of heat-transfer simulations, the HPF technique was adapted to the cryo-fixation of 50-microm-thick alginate gels. The occurrence of artifacts was rejected because no differences were observed in the pattern of cryo-fixed and freeze-substituted samples of various thickness, with and without ethanol as cryo-protectant. A sample thickness of 50 microm was found to ensure an adequate preservation of structures as small as a few nanometers, as verified by TEM and SEM observations. Finally, DSC measurements on alginate solutions and alginate beads revealed that under the experimental conditions (0 3%), alginate cannot be considered to be an efficient cryo-protectant. PMID- 12115413 TI - Low-temperature electron microscopy for the study of polysaccharide ultrastructures in hydrogels. II. Effect of temperature on the structure of Ca2+ alginate beads. AB - Calcium alginate beads were thermally treated at temperatures ranging from 25 degrees C to 130 degrees C for periods of up to 30 minutes. Important modifications to the structure of the alginate beads were shown to be a function of the temperature and period of incubation at each temperature. Modifications to the alginate beads included reduction in size, mechanical resistance, and molecular weight cut-off with increasing temperature and incubation period. Thus, heating 700 microm calcium alginate beads for 20 min at 130 degrees C resulted in a 23% reduction in diameter, 70% increase in mechanical resistance, and 67% reduction in molecular weight cut-off. Incubation of calcium alginate beads containing 2 x 10(6) kDa blue dextran for 20 min at 130 degrees C resulted in no detectable loss of either dye or alginate. This indicates the shrinkage of the beads was due to re-arrangement of the alginate chains within the beads, coupled with loss of water. This hypothesis was verified by direct visual observation of calcium alginate beads before and after thermal treatment using cryo-scanning electron microscopy (cryo-SEM). Unlike other microscopy methods cryo-SEM offers the advantage of extremely rapid freezing which preserves the original structure of the alginate network. As a result cryo-SEM is a powerful tool for studies of hydrogel and capsule structure and formation. Differential scanning calorimetry (DSC) showed that the water entrapped in 2% alginate beads was present in a single state, irrespective of the thermal treatment. This result is attributed to the low alginate concentration used to form the beads. PMID- 12115414 TI - Application of a pore-blockage--cake-filtration model to protein fouling during microfiltration. AB - Although protein fouling is a critical factor governing the performance of microfiltration systems, there have been relatively few studies comparing the fouling behavior of different proteins. Flux-decline data were obtained for the filtration of bovine serum albumin, lysozyme, pepsin, immunoglobulin G, and myoglobin through polycarbonate track-etch membranes. The data were analyzed using a recently developed model that accounts for simultaneous pore blockage and cake formation. The model was in very good agreement with the data for all five proteins, demonstrating the general applicability of this new theoretical framework. The initial fouling due to pore blockage is directly related to the concentration of protein aggregates in solution, which was measured independently by quasi-elastic light scattering. The results provide important insights into the mechanisms of protein fouling during microfiltration. PMID- 12115415 TI - Affinity separation using an Fv antibody fragment-"smart" polymer conjugate. AB - Poly(N-isopropylacrylamide), or PNIPAAm, is considered a "smart" polymer because it sharply precipitates when heated above a critical temperature, about 32 degrees C in water, and redissolves when cooled. Conjugates made of PNIPAAm and IgG antibodies also exhibit the same critical temperature behavior. Interestingly, antigens that are complexed with these conjugates can also be phase-separated along with the conjugates. In this work, we conjugated PNIPAAm for the first time to the immunoglobulin Fv fragment, the smallest fragment of an antibody that still retains the antigenic affinity of the whole antibody. For our studies, we used an Fv fragment that strongly binds hen egg white lysozyme (HEL). The purified Fv fragment-polymer conjugate precipitated at the same temperature as did the pure polymer. After addition of the conjugate to a mixture containing HEL and after thermal separation of the conjugate at 37 degrees C, the amount of HEL in solution was reduced by as much as 80%. We were able to demonstrate the reversibility of the separation through three cycles of precipitation and dissolution. It was also possible to recover free HEL by thermal separation of the conjugate in the presence of an eluant, 50 mM diethylamine. The conjugate can then be recycled for second use. In conclusion, immunoseparations can be performed using smart polymer conjugates made with just the variable domains of an antibody. Unlike whole antibodies, fragments of antibodies can be produced in Escherichia coli, allowing easier genetic engineering of the antibody and tailoring of the conjugate. PMID- 12115416 TI - Antibody production by a hybridoma cell line at high cell density is limited by two independent mechanisms. AB - Our previous attempt to model the stationary phase of production-scale hollow fiber bioreactors using a scaled-down micro hollow-fiber bioreactor resulted in a predicted antibody production rate that was three- to fourfold lower than the actual value (Gramer and Poeschl, 2000). Medium limitations were suspected as the reason for the discrepancy. In this study, various increases in medium feed rate were implemented in the micro bioreactor by increasing the diameter of the silicone tubing that houses the hollow fibers. Because larger diameter tubing may induce oxygen limitations, we also explored the effect of medium recirculation to enhance oxygenation. Antibody production in the micro bioreactor increased both as a result of increased medium supply and due to medium recirculation. However, these parameters increased antibody production through two independent mechanisms. The increased medium supply resulted in a higher cell-specific antibody production rate, but not a higher viable cell density. Medium circulation resulted in the support of a higher viable cell density, but had little effect on the cell-specific secretion rate. The two mechanisms of enhanced antibody production were additive, demonstrating that simultaneous parameters can limit antibody production by this cell line in a hollow-fiber system. When the medium feed and circulation rates were increased to a volumetrically proportional scale, scale-up predictions from the micro bioreactor matched the actual data from the production-scale system to within 15%. These data demonstrate the usefulness of the micro bioreactor for characterizing cell growth and limiting mechanisms at high cell densities. PMID- 12115417 TI - Three-dimensional simulation of grain mixing in three different rotating drum designs for solid-state fermentation. AB - A previously published two-dimensional discrete particle simulation model for radial mixing behavior of various slowly rotating drums for solid-state fermentation (SSF) has been extended to a three-dimensional model that also predicts axial mixing. Radial and axial mixing characteristics were predicted for three different drum designs: (1) without baffles; (2) with straight baffles; and (3) with curved baffles. The axial mixing behavior was studied experimentally with video- and image-analysis techniques. In the drum without baffles and with curved baffles the predicted mixing behavior matched the observed behavior adequately. The predicted axial mixing behavior in the drum with straight baffles was predicted less accurately, and it appeared to be strongly dependent on particle rotation, which was in contrast to the other drum designs. In the drum with curved baffles complete mixing in the radial and axial direction was achieved much faster than in the other designs; that is, it was already achieved after three to four rotations. This drum design may therefore be very well suited to SSF. It is concluded that discrete particle simulations provide valuable detailed knowledge about particle transport processes, and this may help to understand and optimize related heat and mass transfer processes in SSF. PMID- 12115418 TI - Predicting amino acid residues responsible for enzyme specificity solely from protein sequences. AB - We describe a general, modular method for developing protocols to identify the amino acid residues that most likely define the division of a protein superfamily into two subsets. As one possibility, we use PROBE to gather superfamily members and perform an ungapped alignment. We then use a modified BLOSUM62 substitution matrix to determine the discriminating power of each column of aligned residues. The overall method is particularly useful for predicting amino acids responsible for substrate or binding specificity when no structures are available. We apply our method to three pairs of protein classes in three different superfamilies, and present our results, some of which have been experimentally verified. This approach may accelerate the elucidation of enzymic substrate specificity, which is critical for both mechanistic insights into biocatalysis and ultimate application. PMID- 12115419 TI - High-gradient magnetic affinity separation of trypsin from porcine pancreatin. AB - We introduce a robust and scale-flexible approach to macromolecule purification employing tailor-made magnetic adsorbents and high-gradient magnetic separation technology adapted from the mineral processing industries. Detailed procedures for the synthesis of large quantities of low-cost defined submicron-sized magnetic supports are presented. These support materials exhibit unique features, which facilitate their large-scale processing using high magnetic field gradients, namely sufficiently high magnetization, a relatively narrow particle size distribution and ideal superparamagnetism. Following systematic optimization with respect to activation chemistry, spacer length and ligand density, conditions for preparation of effective high capacity (Q(max) = 120 mg g(-1)) strongly interacting (Kd < 0.3 microm) trypsin-binding adsorbents based on immobilized benzamidine were established. In small-scale studies approximately 95% of the endogenous trypsin present in a crude porcine pancreatin feedstock was recovered with a purification factor of approximately 4.1 at the expense of only a 4% loss in alpha-amylase activity. Efficient recovery of trypsin from the same feedstock was demonstrated at a vastly increased scale using a high-gradient magnetic separation system to capture loaded benzamidine-linked adsorbents following batch adsorption. With the aid of a simple recycle loop over 80% of the initially adsorbed trypsin was recovered in-line with an overall purification factor of approximately 3.5. PMID- 12115420 TI - Conversion and toxicity characteristics of formaldehyde in acetoclastic methanogenic sludge. AB - An unadapted mixed methanogenic sludge transformed formaldehyde into methanol and formate. The methanol to formate ratio obtained was 1:1. Formaldehyde conversion proceeded without any lag phase, suggesting the constitutive character of the formaldehyde conversion enzymes involved. Because the rate of formaldehyde conversion declined at increased formaldehyde additions, we hypothesized that some enzymes and/or cofactors might become denatured as a result of the excess of formaldehyde. Furthermore, formaldehyde was found to be toxic to acetoclastic methanogenesis in a dual character. Formaldehyde toxicity was partly reversible because once the formaldehyde concentration was extremely low or virtually removed from the system, the methane production rate was partially recovered. Because the degree of this recovery was not complete, we conclude that formaldehyde toxicity was partly irreversible as well. The irreversible toxicity likely can be attributed to biomass formaldehyde-related decay. Independent of the mode of formaldehyde addition (i.e., slug or continuous), the irreversible toxicity was dependent on the total amount of formaldehyde added to the system. This finding suggests that to treat formaldehyde-containing waste streams, a balance between formaldehyde-related decay and biomass growth should be attained. PMID- 12115421 TI - Production of lipolyzed butteroil by a calf pregastric esterase immobilized in a hollow fiber reactor: III. Effect of glycerol. AB - Lipolysis of butter oil in a hollow fiber reactor containing an immobilized calf pregastric esterase was studied at 40 degrees C, a pH of 6.0, and glycerol concentrations of 0, 150, and 500 g/L in the buffer solution. The concentrations of 10 fatty acid species in the lipolyzed product were determined using high performance liquid chromatography. The rate of loss of enzyme activity and the relative selectivities of this esterase depended on the glycerol concentration. By contrast, the overall rate of release of fatty acids was not affected by the glycerol concentration. Loss of enzyme activity was modeled using first-order kinetics. The models for deactivation and reaction kinetics were fit simultaneously to the data. The model was successful in describing the rates of release of all 10 fatty acid species for a range of space times from 0 to 25 h. The parameters of the model were tested for dependence on glycerol concentration. PMID- 12115422 TI - Advantages of using non-isothermal bioreactors for the enzymatic synthesis of antibiotics: the penicillin G acylase as enzyme model. AB - A new hydrophobic and catalytic membrane was prepared by immobilizing Penicillin G acylase (PGA, EC.3.5.1.11) from E. coli on a nylon membrane, chemically grafted with butylmethacrylate (BMA). Hexamethylenediamine (HMDA) and glutaraldehyde (Glu) were used as a spacer and coupling agent, respectively. PGA was used for the enzymatic synthesis of cephalexin, using D(-)-phenylglycine methyl ester (PGME) and 7-amino-3-deacetoxycephalosporanic acid (7-ADCA) as substrates. Several factors affecting this reaction, such as pH, temperature, and concentrations of substrates were investigated. The results indicated good enzyme binding efficiency of the pre-treated membrane, and an increased stability of the immobilized PGA towards pH and temperature. Calculation of the activation energies showed that cephalexin production by the immobilized biocatalyst was limited by diffusion, resulting in a decrease of enzyme activity and substrate affinity. Temperature gradients were employed as a way to reduce the effects of diffusion limitation. Cephalexin was found to linearly increase with the applied temperature gradient. A temperature difference of about 3 degrees C across the catalytic membrane resulted into a cephalexin synthesis increase of 100% with a 50% reduction of the production times. The advantage of using non-isothermal bioreactors in biotechnological processes, including pharmaceutical applications, is also discussed. PMID- 12115423 TI - Kinetics of benzene biotransformation under microaerophilic and oxygen-limited conditions. AB - A special microbial consortium adapted to degrade petroleum hydrocarbons at limited availability of oxygen, transformed benzene, a highly toxic and carcinogenic contaminant of groundwater and soil, at low initial dissolved oxygen (DO) concentrations of 0.05-2 mg/L. The employed initial concentrations of dissolved oxygen were considerably lower than the previously reported values. Under these conditions, the overall transformation of benzene ranged from 34% +/- 1.7% to 100%, considerably higher than the theoretical predictions for complete mineralization of benzene based on the requirement of 3.08 mg oxygen/mg benzene. Unlike biotransformation that proceeded at the lowest examined DO concentration of 0.05 mg/L, the mineralization of benzene, defined by its conversion to CO(2) and water, required a minimum DO concentration of 0.2 mg/L. The mineralization of benzene under microaerophilic conditions (DO < 2 mg/L), ranged from 0.83% +/- 0.06% to 89% +/- 1.3%, which was less than the theoretical predictions at any given initial DO concentration. The regulatory effects of dissolved oxygen concentration or its partial pressure on the activities of enzymes catalyzing the biotransformation of aromatic hydrocarbons was postulated to account for the reduced mineralization of benzene. The ratio between the transformed benzene and the consumed oxygen increased with the decrease of initial DO concentration, reaching a value of 2.8, considerably higher than the theoretical value of 0.33 obtained for a complete aerobic oxidation of benzene. Phenol was the major and the most stable intermediate metabolite during the biotransformation of benzene at low concentrations of DO. While benzene transformation stopped after the depletion of oxygen in the experimental system, phenol continued to accumulate under strictly anaerobic conditions, indicating its formation from an alternative carbon source, possibly biomass. PMID- 12115424 TI - A two-step, one-pot enzymatic synthesis of cephalexin from D-phenylglycine nitrile. AB - A cascade of two enzymatic transformations is employed in a one-pot synthesis of cephalexin. The nitrile hydratase (from R. rhodochrous MAWE)-catalyzed hydration of D-phenylglycine nitrile to the corresponding amide was combined with the penicillin G acylase (penicillin amidohydrolase, E.C. 3.5.1.11)-catalyzed acylation of 7-ADCA with the in situ-formed amide to afford a two-step, one-pot synthesis of cephalexin. D-Phenylglycine nitrile appeared to have a remarkable selective inhibitory effect on the penicillin G acylase, resulting in a threefold increase in the synthesis/hydrolysis (S/H) ratio. 1,5-Dihydroxynaphthalene, when added to the reaction mixture, cocrystallized with cephalexin. The resulting low cephalexin concentration prevented its chemical as well as enzymatic degradation; cephalexin was obtained at 79% yield with an S/H ratio of 7.7. PMID- 12115428 TI - Metabolic pathway analysis of a recombinant yeast for rational strain development. AB - Elementary mode analysis has been used to study a metabolic pathway model of a recombinant Saccharomyces cerevisiae system that was genetically engineered to produce the bacterial storage compound poly-beta-hydroxybutyrate (PHB). The model includes biochemical reactions from the intermediary metabolism and takes into account cellular compartmentalization as well as the reversibility/irreversibility of the reactions. The reaction network connects the production and/or consumption of eight external metabolites including glucose, acetate, glycerol, ethanol, PHB, CO(2), succinate, and adenosine triphosphate (ATP). Elementary mode analysis of the wild-type S. cerevisiae system reveals 241 unique reaction combinations that balance the eight external metabolites. When the recombinant PHB pathway is included, and when the reaction model is altered to simulate the experimental conditions when PHB accumulates, the analysis reveals 20 unique elementary modes. Of these 20 modes, 7 produce PHB with the optimal mode having a theoretical PHB carbon yield of 0.67. Elementary mode analysis was also used to analyze the possible effects of biochemical network modifications and altered culturing conditions. When the natively absent ATP citrate-lyase activity is added to the recombinant reaction network, the number of unique modes increases from 20 to 496, with 314 of these modes producing PHB. With this topological modification, the maximum theoretical PHB carbon yield increases from 0.67 to 0.83. Adding a transhydrogenase reaction to the model also improves the theoretical conversion of substrate into PHB. The recombinant system with the transhydrogenase reaction but without the ATP citrate-lyase reaction has an increase in PHB carbon yield from 0.67 to 0.71. When the model includes both the ATP citrate-lyase reaction and the transhydrogenase reaction, the maximum theoretical carbon yield increases to 0.84. The reaction model was also used to explore the possibility of producing PHB under anaerobic conditions. In the absence of oxygen, the recombinant reaction network possesses two elementary modes capable of producing PHB. Interestingly, both modes also produce ethanol. Elementary mode analysis provides a means of deconstructing complex metabolic networks into their basic functional units. This information can be used for analyzing existing pathways and for the rational design of further modifications that could improve the system's conversion of substrate into product. PMID- 12115429 TI - Optimal temperature and photoperiod for the cultivation of Agardhiella subulata microplantlets in a bubble-column photobioreactor. AB - The optimal temperature and illumination photoperiod requirements for the phototrophic growth of a novel microplantlet suspension culture derived from the macrophytic marine red alga Agardhiella subulata were determined. The optimal growth temperature was 24 degrees C. The effects of illumination light-dark (LD) photoperiod (hour of light:hours of darkness within a 24 h cycle) on biomass production was studied within a bubble-column photobioreactor. The 4.5 cm diameter photobioreactor was maintained at near-saturation conditions with respect to light flux (38 mciromol photons m(-2) s(-1)), nutrient medium delivery (20% nutrient replacement per day), and CO(2) delivery (0.35 mmol CO(2) L(-1) h( 1)) so that the cumulative effects of photodamage on the cell density versus time curve at photoperiods approaching continuous light could be observed. Biomass production was maximized at 16:8 LD, where biomass densities exceeding 3.6 g dry cell mass L(-1) were achieved after 60 days in culture. Biomass production was proportional to photoperiod at low fractional photoperiods (< or =10:14 LD), but high fractional photoperiods approaching continuous light (> or = 20:4 LD) shut down biomass production. Biomass production versus time profiles under resource saturated cultivation conditions were adequately described by a cumulative photodamage growth model, which coupled reversible photodamage processes to the specific growth rate. Under light-saturated growth conditions, the rate constant for photodamage was kd = 1.17 +/- 0.28 day(-1) (+/-1.0 SE), and the rate constant for photodamage repair was kr = 5.12 +/- 0.95 day(-1) (+/-1.0 SE) at 24 degrees C. PMID- 12115430 TI - Biodegradation of crude oil across a wide range of salinities by an extremely halotolerant bacterial consortium MPD-M, immobilized onto polypropylene fibers. AB - The bacterial consortium MPD-M, isolated from sediment associated with Colombian mangrove roots, was effective in the treatment of hydrocarbons in water with salinities varying from 0 to 180 g L(-1). Where the salinity of the culture medium surpassed 20 g L(-1), its effectiveness increased when the cells were immobilized on polypropylene fibers. Over the range of salinity evaluated, the immobilized cells significantly enhanced the biodegradation rate of crude oil compared with free-living cells, especially with increasing salinity in the culture medium. Contrary to that observed in free cell systems, the bacterial consortium MPD-M was highly stable in immobilized systems and it was not greatly affected by increments in salinity. Biodegradation was evident even at the highest salinity evaluated (180 g L(-1)), where biodegradation was between 4 and 7 times higher with immobilized cells compared to free cells. The biodegradation of pristane (PR) and phytane (PH) and of the aromatic fraction was also increased using cells immobilized on polypropylene fibers. PMID- 12115431 TI - Development of (R)-4-hydroxymandelonitrile synthesis in an aqueous-organic biphasic stirred tank batch reactor. AB - A relatively new hydroxynitrile lyase-catalyzed reaction was optimized to be suitable for rapid and efficient development of a full-scale production process. The conversion of 4-hydroxybenzaldehyde into (R)-4-hydroxymandelonitrile, catalyzed by Prunus amygdalus hydroxynitrile lyase, was carried out in a biphasic system of aqueous buffer (pH 5.5) and methyl tert-butyl ether and is described with a process model. The process model consists of a description of the reaction kinetics, mass transfer kinetics, and mass balances for both the aqueous and the organic phase. Values are determined for the equilibrium constant, the enzyme kinetic parameters, the lumped mass transfer coefficient for benzaldehyde, and the partition coefficients. By using estimated prices of enzyme and reactor use, the optimum aqueous phase volume fraction and required enzyme concentration were calculated at a temperature of 20 degrees C for a batch-operated stirred tank reactor. According to the process model it was possible to convert 90% of the 4 hydroxybenzaldehyde into (R)-4-hydroxymandelonitrile with 95% enantiomeric excess. The price optimum for this reaction was found at an aqueous phase volume of 17% of the total volume. The required enzyme concentration to meet the targets was 28.6 g/L aqueous phase. At the predicted optimum, the synthesis was performed experimentally and the results were in accordance with the simulation regarding the extent of conversion and the enantiomeric excess. PMID- 12115432 TI - Metabolic behavior of immobilized Candida guilliermondii cells during batch xylitol production from sugarcane bagasse acid hydrolyzate. AB - Candida guilliermondii cells, immobilized in Ca-alginate beads, were used for batch xylitol production from concentrated sugarcane bagasse hydrolyzate. Maximum xylitol concentration (20.6 g/L), volumetric productivity (0.43 g/L. h), and yield (0.47 g/g) obtained after 48 h of fermentation were higher than similar immobilized-cell systems but lower than free-cell cultivation systems. Substrates, products, and biomass concentrations were used in material balances to study the ways in which the different carbon sources were utilized by the yeast cells under microaerobic conditions. The fraction of xylose consumed to produce xylitol reached a maximum value (0.70) after glucose and oxygen depletion while alternative metabolic routes were favored by sub-optimal conditions. PMID- 12115433 TI - The measurement of Bacillus mycoides spore adhesion using atomic force microscopy, simple counting methods, and a spinning disk technique. AB - An atomic force microscope has been used to study the adhesion of Bacillus mycoides spores to a hydrophilic glass surface and a hydrophobic-coated glass surface. AFM images of spores attached to the hydrophobic-coated mica surface allowed the measurement of spore dimensions in an aqueous environment without desiccation. The spore exosporium was observed to be flexible and to promote the adhesion of the spore by increasing the area of spore contact with the surface. Results from counting procedures using light microscopy matched the density of spores observed on the hydrophobic-coated glass surface with AFM. However, no spores were observed on the hydrophilic glass surface with AFM, a consequence of the weaker adhesion of the spores at this surface. AFM was also used to quantify directly the interactions of B. mycoides spores at the two surfaces in an aqueous environment. The measurements used "spore probes" constructed by immobilizing a single spore at the apex of a tipless AFM cantilever. The data showed that stretching and sequential bond breaking occurred as the spores were retracted from the hydrophilic glass surface. The greatest spore adhesion was measured at the hydrophobic-coated glass surface. An attractive force on the spores was measured as the spores approached the hydrophobic-coated surface. At the hydrophilic glass surface, only repulsive forces were measured during the approach of the spores. The AFM force measurements were in qualitative agreement with the results of a hydrodynamic shear adhesion assay that used a spinning disk technique. Quantitatively, AFM measurements of adhesive force were up to 4 x 10(3) times larger than the estimates made using the spinning disk data. This is a consequence of the different types of forces applied to the spore in the different adhesion assays. AFM has provided some unique insights into the interactions of spores with surfaces. No other instrument can make such direct measurements for single microbiological cells. PMID- 12115434 TI - Inducible expression of Bcl-XL restricts apoptosis resistance to the antibody secretion phase in hybridoma cultures. AB - B-cell hybridomas are widely used to produce monoclonal antibodies via large scale cell culture. Unfortunately, these cells are highly sensitive to apoptotic death under conditions of nutrient deprivation observed at the plateau phase of batch cultures. Previous work has indicated that constitutive high-level expression of antiapoptotic genes in hybridoma cells could delay apoptosis, resulting in higher cell densities and prolonged viability. However, the constitutive high-level expression of antiapoptotic genes has been shown to have detrimental effects on genomic stability of other types of cultured cells. Inducible gene expression may be used to avoid this problem. In the present study, we first constructed an expression vector in which the promoter of a mammalian metallothionein (MT) gene drives the expression of bcl-XL in response to metal exposure. The vector was then used to exogenously control the expression of bcl-XL in D5 hybridoma cells. Our data show that stably transfected D5 cells (4G1.D9) expressed high levels of Bcl-X(L) following overnight exposure to ZnSO(4) concentrations (50 to 100 microM) that did not affect control cells. The level of Bcl-X(L) expressed after ZnSO(4) induction was sufficient to prevent apoptosis experimentally induced by cycloheximide and allowed 4G1.D9 cells to grow at higher densities and remain viable for prolonged periods in suboptimal culture conditions. The use of inducible bcl-XL expression permits extension of the viability of cultured B-cell hybridomas during the antibody secretion phase without the adverse genetic effects associated with constitutive long-term bcl-XL expression. PMID- 12115436 TI - Integration of reactive membrane extraction with lipase-hydrolysis dynamic kinetic resolution of naproxen 2,2,2-trifluoroethyl thioester in isooctane. AB - Lipases immobilized on polypropylene powders have been used as the biocatalyst in the enantioselective hydrolysis of (S)-naproxen from racemic naproxen thioesters in isooctane, in which trioctylamine was added to perform in situ racemization of the remaining (R)-thioester substrate. A detailed study of the kinetics for hydrolysis and racemization indicates that increasing the trioctylamine concentration can activate and stabilize the lipase as well as enhance the racemization and non-stereoselective hydrolysis of the thioester. Effects of the aqueous pH value and trioctylamine concentration on (S)-naproxen dissociation and partitioning in the aqueous phase as well as the transportation in a hollow fiber membrane were further investigated. Good agreements between the experimental data and theoretical results were obtained when the dynamic kinetic resolution process was integrated with a hollow fiber membrane to reactively extract the desired (S) naproxen out of the reaction medium. PMID- 12115435 TI - Catabolic enzyme levels in bacteria grown on binary and ternary substrate mixtures in continuous culture. AB - Bacteria grow on multicomponent substrates in most natural and engineered environments. To advance our ability to model bacterial growth on such substrates, axenic cultures were grown in chemostats at a low specific growth rate and a constant total energy flux on binary and ternary substrate mixtures and were assayed for key catabolic enzymes for each substrate. The substrates were benzoate, salicylate, and glucose, and the enzymes were catechol 1,2 dioxygenase, gentisate 1,2-dioxygenase, and glucose-6-phosphate dehydrogenase, respectively. The binary mixtures were salicylate with benzoate and salicylate with glucose. Measurements were also made of oxygen uptake rate by whole cells in response to each substrate. The effects of the substrate mixture on the oxygen uptake rate paralleled the effects on the measured enzymes. Catechol 1,2 dioxygenase exhibited a threshold response before synthesis occurred. Below the threshold flux of benzoate through the chemostat, either basal enzyme levels or nonspecific enzymes kept reactor concentrations too low for enzyme synthesis. Above the threshold, enzyme levels were linearly related to the fraction of the total energy flux through the chemostat due to benzoate. Gentisate 1,2 dioxygenase exhibited a linear response to the salicylate flux when mixed with benzoate, but a threshold response when mixed with glucose. Glucose-6-phosphate dehydrogenase activity increased in direct proportion to the glucose flux through the chemostat over the entire range studied. The results from two ternary mixtures were consistent with those from the binary mixtures. PMID- 12115437 TI - Virus safety of a porcine-derived medical device: evaluation of a viral inactivation method. AB - The goal of this study was to evaluate the efficacy of a virus-inactivating process for use during the preparation of porcine-derived extracellular matrix biomaterials for human clinical implantation. Porcine small intestine, the source material for the tissue-engineered, small intestinal submucosa (SIS) biomaterial, was evaluated. Relevant enveloped, non-enveloped, and model viruses representative of different virus families were included in the investigation: porcine parvovirus (PPV), porcine reovirus, murine leukemia retrovirus (LRV), and porcine pseudorabies (herpes) virus (PRV). Samples of small intestine were deliberately inoculated with approximately 1 x 10(7) plaque-forming units (PFU) of virus which were thereafter exposed to a 0.18% peracetic acid/4.8% aqueous ethanol mixture for time periods ranging from 5 minutes to 2 hours. Enveloped viruses were more easily inactivated than non-enveloped viruses, but material processed for 30 minutes or longer inactivated all of the viruses. D(10) values were calculated and used to extrapolate the extent of inactivation after 2 hours. Viral titers were reduced by more than 14.0 log(10) PPV, 21.0 log(10) reovirus, 40.0 log(10) PRV, and 27.0 log(10) LRV, meeting international standards for viral sterility. These results demonstrate that treatment of porcine small intestine with a peracetic acid/ethanol solution leads to a virus-free, non-crosslinked biomaterial safe for xenotransplantation into humans. PMID- 12115438 TI - Phase separation rates of aqueous two-phase systems: correlation with system properties. AB - The kinetics of phase separation in aqueous two-phase systems have been investigated as a function of the physical properties of the system. Two distinct situations for the settling velocities were found, one in which the light, organic-rich (PEG) phase is continuous and the other in which the heavier, salt rich (phosphate) phase is continuous. The settling rate of a particular system is a crucial parameter for equipment design, and it was studied as a function of measured viscosity and density of each of the phases as well as the interfacial tension between the phases. Interfacial tension increases with increasing tie line length. A correlation that describes the rate of phase separation was investigated. This correlation, which is a function of the system parameters mentioned above, described the behavior of the system successfully. Different values of the parameters in the correlation were fitted for bottom-phase continuous and top-phase-continuous systems. These parameters showed that density and viscosity play a role in the rate of separation in both top continuous- and bottom continuous-phase regions but are more dominant in the continuous top-phase region. The composition of the two-phase system was characterized by the tie line length. The rate of separation increased with increasing tie line length in both cases but at a faster rate when the bottom (less viscous) phase was the continuous phase. These results show that working in a continuous bottom-phase region is advantageous to ensure fast separation. PMID- 12115439 TI - Active site titration as a tool for the evaluation of immobilization procedures of penicillin acylase. AB - Native and immobilized preparations of penicillin acylase from Escherichia coli and Alcaligenes faecalis were studied using an active site titration technique. Knowledge of the number of active sites allowed the calculation of the average turnover rate of the enzyme in the various preparations and allowed us to quantify the contribution of irreversible inactivation of the enzyme to the loss of catalytic activity during the immobilization procedure. In most cases a loss of active sites as well as a decrease of catalytic activity per active site (turnover rate) was observed upon immobilization. Immobilization techniques affected the enzymes differently. The effect of increased loading of penicillin acylase on the average turnover rate was determined by active site titration to assess diffusion limitations in the carrier. PMID- 12115440 TI - Effect of electrical charges and fields on injury and viability of airborne bacteria. AB - In this study, the effects of the electric charges and fields on the viability of airborne microorganisms were investigated. The electric charges of different magnitude and polarity were imparted on airborne microbial cells by a means of induction charging. The airborne microorganisms carrying different electric charge levels were then extracted by an electric mobility analyzer and collected using a microbial sampler. It was found that the viability of Pseudomonas fluorescens bacteria, used as a model for sensitive bacteria, carrying a net charge from 4100 negative to 30 positive elementary charges ranged between 40% and 60%; the viability of the cells carrying >2700 positive charges was below 1.5%. In contrast, the viability of the stress-resistant spores of Bacillus subtilis var. niger (used as simulant of anthrax-causing Bacillus anthracis spores when testing bioaerosol sensors in various studies), was not affected by the amount of electric charges on the spores. Because bacterial cells depend on their membrane potential for basic metabolic activities, drastic changes occurring in the membrane potential during aerosolization and the local electric fields induced by the imposed charges appeared to affect the sensitive cells' viability. These findings facilitate applications of electric charging for environmental control purposes involving sterilization of bacterial cells by imposing high electric charges on them. The findings from this study can also be used in the development of new bioaerosol sampling methods based on electrostatic principles. PMID- 12115441 TI - Thin film rheology and lubricity of hyaluronic acid solutions at a normal physiological concentration. AB - Using a surface forces apparatus to measure forces, and optical (multiple beam) interferometry to measure surface shapes and separations (to +/-1 A), the normal, viscous, and shear (lubrication) forces between smooth mica surfaces in aqueous hyaluronic acid (HA) solutions were measured. The experimental conditions of loading pressures, pH, and HA concentration were set to closely correspond to physiological human knee-joint conditions. From the force and optical (refractive index) measurements, it was concluded that, like other negatively charged polyelectrolytes, HA does not naturally adsorb on the mica surface which is hydrophilic and weakly negatively charged at physiological conditions: the polymer solution exhibits the bulk viscosity (22.5 +/- 1.5 cP) for films thicker than about 0.4 miccrom of the polymer, whereas for thinner films, the viscosity decreases monotonically toward the value of the pure electrolyte solution (1 cP) as HA is extruded from between the surfaces. This is indicative of a repulsive "depletion" interaction of HA with each mica surface and to a weakly attractive polymer-mediated force between the two surfaces. Thus, free HA in synovial fluid is not expected to act as a good "boundary lubricant." Relaxation measurements on approaching and receding surfaces in HA solutions were also performed, and it is shown that the presence of HA in the bulk solution can improve "hydrodynamic" modes of lubrication, for example, by assuaging the compression stroke. The study includes information that is beneficial to researchers working with biomaterials viscosupplementation devices. PMID- 12115442 TI - Bioactivity of three CaO-P2O5-SiO2 sol-gel glasses. AB - Three gel glasses containing 25 mol % of CaO and SiO(2) + P(2)O(5) contents (in mol %): 75 + 0 (S75); 72.5 + 2.5 (S72.5P2.5); and 70 + 5 (S70P5), respectively, were obtained, characterized, and studied when soaked in a simulated body fluid (SBF). The influence of composition in both textural properties (surface area and porosity) and in vitro behavior of glasses was studied. In as prepared S72.5P2.5 and S70P5 glasses, crystalline phosphate nuclei were detected through XRD and FTIR. In addition, N(2) adsorption and Hg porosimetry measurements showed that the surface area increased, whereas the pore volume and the pore diameter decreased as P(2)O(5) in glasses increased. These variations were explained on the basis of the withdrawal of calcium from the glass silica network, due to the calcium-phosphorous bonding. In vitro studies showed that the three compositions were bioactive, because an apatite layer was formed after soaking in SBF. S75 presented the highest initial reactivity but the lowest crystallization rate of the apatite-like phase. For S72.5P2.5, and S70P5 the amorphous calcium phosphate formation was slower than for S75, but the crystallization of apatite was observed after shorter periods in SBF. Furthermore, after 7 days of soaking, the layer thickness decreased as P(2)O(5) in glasses increased. PMID- 12115443 TI - Flow properties of liquid calcium alginate polymer injected through medical microcatheters for endovascular embolization. AB - The flow properties of liquid calcium alginate injections were investigated for application in endovascular embolization. Alginate shear properties were assessed with a rheometer and a controlled injection system. The experimental results were used to model the flow properties and predict alginate's flow characteristics within various medical microcatheter delivery systems. The results suggest that alginates undergo shear-thinning effects with increasing shear. A flow comparison of 2.0 wt % alginate and a Newtonian fluid (82 cP) injected from the same microcatheter had similar flow rates at low injection pressure (100 kPa). However, at high injection pressure (2100 kPa), the alginate was injectable at a flow rate 100% higher than was the Newtonian fluid. Further analysis of injections through microcatheters resulted in a flow model for predicting viscosity changes, flow rates, and injection pressures of liquid alginate at medium-to-high shear rates. The predicted injection pressures and flow rates had an average variance of less than 15% from that of the experimental flow data. This study indicates that calcium alginate has the requisite flow properties for successful delivery to vascular lesions via endovascular injection. Possible uses of alginates include treating arteriovenous malformations (AVMs), aneurysms, blood flow to tumors, and vascular hemorrhages. PMID- 12115444 TI - Ion leaching from dental ceramics during static in vitro corrosion testing. AB - Dental ceramics are often called inert materials. It can be hypothesized, however, that differences in the composition, microstructure, and environmental conditions will affect the degree of corrosion degradation in an aqueous environment. The aims of the study were, therefore, to study the ion dissolution from glass-phase ceramics, with or without crystalline inclusions, and from all crystalline ceramics and to compare the effects of different corrosion media. Ceramic specimens were produced from glass-phase and oxide ceramics and given an equivalent surface smoothness, after which they were subjected to in vitro corrosion (Milli-Q water at 37 +/- 2 degrees C for 18 h and 4% acetic acid solution at 80 +/- 2 degrees C for 18 h, respectively). The temperature of the corrosion solution was slowly increased until it reached 80 +/- 2 degrees C to reduce the risk of microcrack formation at the surface. The analyses of ion leakage were performed with inductively coupled plasma optical emission spectroscopy. A large number of inorganic elements leached out from the various dental ceramics. The major leaching elements were sodium and potassium; in the acid-corrosion experiments, there were also magnesium, silicon, and aluminum and, on a lower scale, yttrium, calcium, and chromium. The various glass-phase ceramics displayed significant differences in ion leakage and significantly higher leakage values than all-crystalline alumina and zirconia ceramics. No significant difference in dissolution was found between high and low-sintering glass-phase ceramics or between glass-phase ceramics with high volume fractions of crystallites in the glass phase in comparison with those with lower crystalline content. It can be concluded, therefore, that none of the dental ceramics studied are chemically inert in an aqueous environment. PMID- 12115445 TI - Porous polymer/bioactive glass composites for soft-to-hard tissue interfaces. AB - Porous composites consisting of a polysulfone (or cellulose acetate) matrix and bioactive glass particles were prepared by phase separation techniques. Microstructures were designed for potential application as an interconnect between artificial cartilage and bone. The effects of polymer type, concentration and molecular weight, as well as bioactive glass size and content, on the microstructures of the composites were studied. The composites have asymmetric structures with dense top layers and porous structures beneath. The microstructural features depend most strongly on the type of polymer, the interaction between the polymer and bioactive glass, and the glass content. The dense top layer could be removed by abrasion to make a structure with large pores (20-150 microm) exposed. Composites were immersed in simulated body fluid at body temperature. The growth of hydroxycarbonate apatite inside and on the composites demonstrates their potential for integration with bone. Composite modulus and break strength increased with increasing glass content due to the change in composition and pore content. PMID- 12115446 TI - Interfacial tensile strength between polymethylmethacrylate-based bioactive bone cements and bone. AB - We have developed two types of polymethylmethacrylate (PMMA)-based bioactive bone cements containing bioactive glass beads (designated GBC) or apatite-wollastonite containing glass-ceramic powder (designated AWC) as the filler. A new method was used to evaluate the bone-cement interfacial strength of these bioactive bone cements. Two types of bioactive bone cements (GBC and AWC) and PMMA cement (CMW 1) were put in a frame attached to the smooth tibial metaphyseal cortex of the rabbit and polymerized in situ. The load required to detach the cement from the bone was measured at 4, 8, and 16 weeks after implantation. The interfacial tensile strength of GBC and AWC showed significantly higher values than PMMA cement from 4 weeks, and increased with time. For GBC, strength reached a maximum value of 12.39 +/- 1.79 kgf 16 weeks after implantation. Histological examination of rabbit tibiae up to 16 weeks demonstrated no intervening layer between the bioactive bone cements and the bone, whereas fibrous tissue was observed at the interface between the PMMA cement and the bone. From this study, we conclude that PMMA-based bioactive bone cements have a relatively higher adhesiveness at the interface than the conventionally used PMMA cement, showing potential as a promising alternative. PMID- 12115448 TI - Polymer--calcium phosphate cement composites for bone substitutes. AB - The use of self-setting calcium phosphate cements (CPCs) as bioresorbable bone replacement implant materials presently is limited to non-load-bearing applications because of their low compressive strength relative to natural bone. The present study investigated the possibility of strengthening a commercially available CPC, alpha-BSM, by incorporating various water-soluble polymers into the cement paste during setting. Several polyelectrolytes, poly(ethylene oxide), and the protein bovine serum albumin (BSA) were added in solution to the cement paste to create calcium phosphate-polymer composites. Composites formulated with the polycations poly(ethylenimine) and poly(allylamine hydrochloride) exhibited compressive strengths up to six times greater than that of pure alpha-BSM material, with a maximum value reached at intermediate polymer content and for the highest molecular weight studied. Composites containing BSA developed compressive strengths twice that of the original cement at protein concentrations of 13-25% by weight. In each case, XRD studies correlate the improvement in compressive strength with reduced crystallite dimensions, as evidenced by a broadening of the (0,0,2) reflection. This suggests that polycation or BSA adsorption inhibits crystal growth and possibly leads to a larger crystal aspect ratio. SEM results indicate a denser, more interdigitated microstructure. The increased strength was attributed to the polymer's capacity to bridge between multiple crystallites (thus forming a more cohesive composite) and to absorb energy through plastic flow. PMID- 12115447 TI - Effects of dental resin components on vascular reactivity. AB - The frequent use of resins in dentistry has raised the question of their compatibility with oral tissues. The present study was undertaken to determine the effects of the resin components methyl methacrylate (MMA), hydroxyethyl methacrylate (HEMA), and triethylene glycol dimethacrylate (TEGDMA) on the reactivity of blood vessels using the isolated rat aorta as a tissue model. MMA, HEMA, and TEGDMA caused endothelium-dependent and -independent relaxation of rat aortic rings in a concentration-related manner. The endothelium-dependent responses of the tissues to all the resins were significantly attenuated by N nitro-L-arginine methyl ester (L-NAME), indicating the involvement of nitric oxide. The vasorelaxant effects of both MMA and TEGDMA on the intact and denuded aortae were markedly inhibited by indomethacin, providing evidence for the role of prostanoids in these responses. Glybenclamide selectively attenuated TEGDMA induced relaxation of the tissues with and without endothelium to a similar extent, suggesting the activation of vascular smooth muscle K(ATP) channels by this resin. It is concluded that MMA, HEMA, and TEGDMA interfere with the function of blood vessels by inducing vasorelaxation via different mechanisms, which, depending upon the type of resin, may at least involve the release of nitric oxide and prostanoid(s), and the activation of smooth muscle K(ATP) channels. These phenomena may play a role in tissue homeostasis and certain pathophysiological conditions associated with the application of resin materials to the oral environment. PMID- 12115449 TI - Extended bioactivity in the proximity of hydroxyapatite ceramic surfaces induced by polarization charges. AB - Surface charges of electrically polarized hydroxyapatite (HAp) ceramics were demonstrated to enhance osteoconductivity in the wide gap of the canine bone whereas the bone formation processes varied according to the charge polarity. Cell reactions in the vicinity of the charged surfaces of HAp ceramics were not phagocytic absorption but rather bone formation by stimulated osteoblasts surrounding newly formed bone 7 days after implantation. The bone formation in direct contact with negatively charged ceramic surfaces suggests that the negative charges enhanced the osteobonding ability of the HAp ceramics. In the vicinity of the positively charged HAp surface, the formation of bones derived from osteoid tissues entirely occupied the 0.2-0.3-mm gaps between the HAp and the bone at 7 days. Surface charges induced by electrical polarization significantly cooperated with the innate bioactivity of HAp and reconstructed the wide bone defects more promptly than did the nonpolarized HAp ceramic surface. PMID- 12115450 TI - Characterization of a novel calcium phosphate/sulphate bone cement. AB - Apatitic cements have shown excellent biocompatibility and adequate mechanical properties but have slow resorption in the human body. To assure that new bone tissue grows into the bone defect, a certain porosity is necessary although hard to achieve in injectable cements with suitable mechanical properties. An attempt was made by mixing alpha-tricalcium phosphate (alpha-TCP), calcium sulphate hemihydrate (CSH) and an aqueous solution containing 2.5 wt% of Na(2)HPO(4). The aim was to obtain a material containing two phases: a) one apatitic phase (calcium-deficient hydroxyapatite; CDHA) and b) one resorbable phase (calcium sulphate dihydrate; CSD). alpha-TCP and CSH mixtures were produced at relative intervals of 20 wt%. The liquid-to-powder (L/P) ratio to obtain a paste was 0.32 mLg(-1). The highest compressive strength (34 MPa) was obtained for the pure alpha-TCP sample. The strength was, in a first approximation, directly correlated to the weight proportions of the powders. X-ray diffraction analysis showed that the relative intensity for CDHA increased linearly, and the one for CSD decreased exponentially, when the amount of alpha-TCP increased. Thus, CSH ceased to transform to CSD when the amount of alpha-TCP increased. Observations in environmental scanning electron microscopy confirmed the X-ray diffraction results. CSH-crystals (100 microm) were embedded in the HA-matrix permitting gradual porosity in the material when resorbed. PMID- 12115451 TI - Effects of substratum surface topography on the organization of cells and collagen fibers in collagen gel cultures. AB - This study investigated the orientation of fibroblasts and collagen cultured on microfabricated grooved or smooth titanium surfaces, as well as on tissue culture polystyrene, in the presence or absence of collagen gels. The gels were first added either to the confluent fibroblast culture on the surface (cell-gel condition) or to the fibroblasts were suspended within the collagen gel and then placed onto the surface (gel condition). Cells and collagen were observed with differential interference, polarization, and confocal laser scanning microscopy. Although the smooth surfaces had no effect on cell orientation in the gel for the first 2 weeks of culture, cells did orient with grooves regardless of the culture conditions. There was evidence for orthogonal multilayering of cells under the cell-gel condition at 4 weeks, and collagen alignment reflected cell alignment. The interaction of the collagen gel with the surface depended on whether the cell gel or the gel condition was employed. In the former condition, the gel contracted toward the substratum, whereas the gel condition resulted in the formation of a ring of collagen loosely attached to the substratum. These results suggest that the order in which fibroblasts encounter substratum and extracellular matrix can influence the eventual matrix-cell interactions, and that substratum topography can influence matrix and cell orientation in zones not immediately in contact with the surface. PMID- 12115452 TI - The effect of polyethylene particle phagocytosis on the viability of mature human macrophages. AB - Macrophages are the major cell type observed in the inflammatory membrane retrieved at implant revision surgery. In this study, mature human monocyte derived macrophages (MDM) were adapted to a previously established in vitro model to examine the influence of high-density polyethylene (HDPE) particulate (4-10 microm) on MDM viability. HDPE particles were suspended in soluble type I collagen, which subsequently was solidified on glass coverslips. Mature human macrophages, derived from differentiating peripheral blood monocytes on polystyrene for 10 days, were incubated in culture media on collagen controls and collagen-particle substrata for 31 days. Histologic analysis demonstrated that MDMs were in contact with the particles at 2 h. The majority of the particles were associated with the cells within 24 h. Based on electron microscopy, those cells associated with the particles appeared to be morphologically activated rather than necrotic or apoptotic. Assessment of cell viability revealed no differences among the groups at 24 h, but at 31 days significantly more viable cells and higher DNA values were found associated with the particle groups versus the collagen controls. The histologic results validate human mature MDMs as a clinically relevant cell type for study of the role of polyethylene particulate in aseptic loosening. The cell viability results indicate that phagocytosis of HDPE is not toxic to MDMs but in fact prolongs MDM survival. The long-lived MDMs may play a role in perpetuating chronic inflammation surrounding implants. PMID- 12115453 TI - The biological effects of antiadhesion agents on activated RAW264.7 macrophages. AB - The objective of this study is to determine the biological effects of various antiadhesion agents on macrophages, which play an essential role in wound healing and adhesion. To determine these effects, RAW264.7 macrophages were activated with lipopolysaccharide in the presence of antiadhesion agents: oxidized regenerated cellulose (oxyC), sodium hyaluronate (HA), dexamethasone (Dex), or chondroitin sulfate (CS). The release of nitric oxide (NO), vascular endothelial growth factor (VEGF), interleukin-6 (IL-6), or matrix metalloproteinases (MMPs) from RAW264.7 was measured. We found that oxyC reduced the release of NO, IL-6, MMP-2, and MMP-9, whereas it enhanced the release of VEGF. HA reduced the release of MMP-2, whereas it enhanced the release of VEGF and NO. HA exhibited no significant effect on the release of IL-6 or MMP-9. Dex reduced the release of NO, VEGF, IL-6, MMP-2, and MMP-9. CS reduced the release of VEGF, IL-6, and MMP 2, although it had no significant effect on the release of NO and MMP-9. Antiadhesion agents, which have been clinically used as physical barriers, modulated the functions of macrophages. PMID- 12115454 TI - Tribological investigation of oriented HDPE. AB - The possibility to control the wear properties of high-density polyethylene (HDPE) material at an early processing stage is explored. Wear measurements of cold roll-drawn HDPE with two different draw ratios were carried out for three sliding planes, each in two directions. The dependence of the wear properties on the degree and direction of orientation was investigated. The experiments were performed in a pin-on-disc machine in a dry environment. The tribo-couple consisted of HDPE plates versus a standardised diamond coated steel disc. The results show that the wear resistance of cold roll-drawn HDPE differ widely, by a factor up to 6, depending on the sliding direction relative to the drawing direction. The material has a significantly better wear resistance when the sliding direction was perpendicular to the processing direction. The best wear resistance was in the end plane and it was improved by a factor up to 3.6 when the draw ratio was increased from 2 to 4. These results indicate that molecular orientation by polymer processing is a promising method to improve the wear properties and decrease the wear debris production of HDPE. PMID- 12115456 TI - Kinetic study of citric acid influence on calcium phosphate bone cements as water reducing agent. AB - Most of the research performed on calcium phosphate bone cements (CPBCs) has dealt with the improvement of bone cement formulations for new, demanding bone filling applications. In particular, the development of injectable bone cements is of real interest for the biomedical community. The aim of this work was to study the effect of citric acid on the injectability and the setting properties of alpha-tricalcium phosphate-based cements. A comparative kinetic study was performed on cements with and without citric acid relating the hardening curves and the hydration rates using a mathematical approach. Citric acid behaved as a fluidificant during the first stages of the cement mixing. The dissolution precipitation reactions of the alpha-tricalcium phosphate were retarded with the addition of citric acid and the compressive strength at saturation increased. In conclusion, citric acid can behave as a water-reducing admixture. PMID- 12115455 TI - Adsorbed poly(ethyleneoxide)-poly(propyleneoxide) copolymers on synthetic surfaces: spectroscopy and microscopy of polymer structures and effects on adhesion of skin-borne bacteria. AB - Poly(ethyleneoxide)-copoly(propyleneoxide) (PEO-PPO) polymer coatings were evaluated for their resistance to the attachment of the marker organism Serratia marcescens and the skin-borne bacteria Staphylococcus epidermidis. The copolymers were adsorbed onto poly(styrene) films-chosen as simplified physicochemical models of skin surfaces-and their surface characteristics probed by contact angle goniometry, attenuated total reflectance-Fourier transform infrared (ATR-FTIR), atomic force microscopy (AFM), and X-ray photoelectron spectroscopy (XPS). These functional surfaces were then presented to microbial cultures, bacterial attachment was assessed by fluorescence microscopy and AFM, and the structures of the polymer films examined again spectroscopically. Surface characterization data suggest that the adsorbed copolymer was partially retained at the surface and resisted bacterial attachment for 24 h. Quantitative evaluation of cell attachment was carried out by scintillation counting of (14)C-labeled microorganisms in conjunction with plate counts. The results show that a densely packed layer of PEO-PPO copolymer can reduce attachment of skin commensals by an order of magnitude, even when the coating is applied by a simple adsorptive process. The work supports the hypothesis that adhesion of microorganisms to biological substrates can be reduced if a pretreatment with an appropriate copolymer can be effected in vivo. PMID- 12115458 TI - Effect of surface treatment of NiTi alloy on its corrosion behavior in Hanks' solution. AB - The effect of surface modification of NiTi shape memory alloy on its corrosion behavior in Hanks' solution was determined. The near-equi-atomic super elastic NiTi (Ni 55.8 wt %) alloy used for this study was provided by Memry USA. The surfaces of heat-treated samples were modified by mechanical polishing (MP), electropolishing (EP), and electropolishing followed by chemical passivation (CP). As-heat-treated samples with straw-colored oxide finishes (SCO) and blue colored oxide finishes (BO) also were included in the study. Surface analysis was performed using auger electron spectroscopy (AES), atomic force microscopy (AFM), and contact angle measurements (CAM). It was shown that surface roughness increased in the order CP < EP < SCO < BO < MP. The nickel release within the five groups of NiTi samples, as determined by atomic absorption spectrophotometry, reduced in time over the measured period. The level of Ni ions released over a 25-day immersion period was highest in the SCO sample (0.002 microg/day). This Ni level is negligible compared with the daily intake of Ni in an ordinary diet. The auger electron spectroscopy (AES) analyses indicated that before immersion in Hanks' physiologic solution, the main surface composition of all the samples was titanium and nickel, with a small amount of oxygen, carbon, and sulphur as contaminants. And the surface oxide thickness of the different samples increased in the order CP < EP < MP < BO < SCO. On the other hand, for the electrodes treated under the same conditions, the mean breakdown potential value decreased in the order BO > MP > CP > EP > SCO while the corrosion current density and rate increased in the order CP < SCO < EP < BO < MP. PMID- 12115459 TI - Control of surface mineralization using collagen fibrils. AB - Surface structure in the form of roughness and organized patterning can affect osteoblastic adhesion and proliferation. This study investigates the effect of reconstituted collagen fibrils on the deposition pattern of a homogeneous inorganic mineral (sodium chloride). The patterns were monitored from nanometer to millimeter scales using atomic force and light microscopies. Initially, mineral deposits formed blocks following the contour of the collagen fibrils. At later times, dendritic structures formed. This demonstrates that collagen fibrils can affect the surface deposition pattern of saline minerals. It is also shown that collagen fibril diameter and the stoichiometry of the inorganic and organic phases effect the surface distribution of minerals. PMID- 12115457 TI - Delivery of insulin from hydroxyapatite ceramic microspheres: preliminary in vivo studies. AB - The major limiting factor in utilizing protein drugs for sustained delivery is the lack of suitable delivery systems. Ceramic hydroxyapatite microspheres are biocompatible and utilized for the purification of proteins. As a preliminary study, we have investigated the possibility of using hydroxyapatite ceramic microspheres loaded with insulin as an implantable delivery system in rats. With this limited test, it was shown that the loaded insulin was active and able to suppress the blood glucose level in normal rats. PMID- 12115460 TI - Effects of ion concentration and pH on hydroxyapatite deposition from aqueous solution onto titanium by the thermal substrate method. AB - A new hydrocoating method (the thermal substrate method) has been proposed for coating calcium phosphates, such as hydroxyapatite, onto titanium substrates in an aqueous solution. The influences of several solution properties on the thermal substrate method were examined. The solutions used included 3 mmol dm(-3) Ca(H(2)PO(4))(2) and 7 mmol dm(-3) CaCl(2) as a reference concentration solution. The ion concentration was changed from 0.1 to 2 times with respect to the reference concentration. The experimental studies were conducted under the following conditions: temperature = 140 degrees C, heating time = 10-20 min., pH = 4-8 and Ca/P = 0.0167-16.7. The type of precipitate changed, depending on the pH and ion concentration. In the reference solution with pH > 6, predominantly hydroxyapatite was precipitated onto titanium. By contrast, only CaHPO(4) was formed in the solution of pH 4. In the solution with an ion concentration of one tenth the reference solution, CaHPO(4) was also precipitated. The addition of H(3)PO(4) to the 0.1-times solution accelerated the precipitation rate of HA. It is suggested that the PO(4) (3-) concentration was insufficient to form HA in the Ca/P = 1.67 solution. PMID- 12115462 TI - Force spectroscopy study of the adhesion of plasma proteins to the surface of a dialysis membrane: role of the nanoscale surface hydrophobicity and topography. AB - A mechanochemical study of the process of adhesion of plasma proteins to the surface of dialysis membranes was carried out with a scanning force microscope (SFM) in the force spectroscopy mode. Three representative blood plasma proteins (fibronectin, fibrinogen, and albumin) covalently were grafted to a SFM probe, and the adhesion forces of these proteins to cellulosic and synthetic dialysis membranes were measured. The experiment was tailored to apply a controlled load on the protein molecules adsorbed onto the surface in order to simulate the squeezing forces exerted on them during blood filtration. The de-adhesion forces, measured using this new approach for studying the interaction between a protein and dialysis membranes, suggest that the membrane's topography, at a nanometer scale, plays a critical role in the adhesion process. This result was strongly supported by parallel experiments performed on a flattened glass surface with the same dominant hydrophilic character as dialysis membranes. In contrast, a hydrophobic polystyrene surface led to de-adhesion forces at least one order of magnitude greater, overwhelming any possible shape recognition process between the protein molecules and the surface. PMID- 12115461 TI - Feasibility study using a natural compound (reuterin) produced by Lactobacillus reuteri in sterilizing and crosslinking biological tissues. AB - Bioprostheses derived from biological tissues have to be fixed and subsequently sterilized before they can be implanted in humans. Currently available crosslinking agents and sterilants used in the fixation or sterilization of biological tissues such as glutaraldehyde and formaldehyde are all highly cytotoxic, which may impair the biocompatibility of bioprostheses. Therefore, it is desirable to provide an agent suitable for use in biomedical applications that is of low cytotoxicity and may form sterile and biocompatible crosslinked products. To achieve this goal, a natural compound (reuterin), produced by Lactobacillus reuteri in the presence of glycerol, was used by our group. It is known that reuterin has antibacterial, antimycotic, and antiprotozoal activities. Additionally, as in the case with formaldehyde, reuterin may react with the free amino groups in biological tissues by using its aldehyde functional group. Therefore, it was speculated that reuterin could be used as a crosslinking agent and a sterilant for biological tissues in the same way as glutaraldehyde and formaldehyde. In the study, the production of reuterin, produced by Lactobacillus reuteri under control conditions, was reported. Preparative chromatography was used to purify reuterin. Also, the minimal inhibitory concentration and minimal bactericidal concentration of reuterin and its antimicrobial activity on a contaminated tissue were investigated. In addition, the cytotoxicity of reuterin was evaluated. Glutaraldehyde, the most commonly used sterilant in the sterilization of biological tissues, was employed as a control. Furthermore, the feasibility of using reuterin as a crosslinking agent in fixing biological tissues was studied. Fresh and the glutaraldehyde-fixed tissues were used as controls. The results obtained in the minimal inhibitory concentration and minimal bactericidal concentration studies and in the sterilization study of a contaminated tissue indicated that the antimicrobial activity of reuterin is significantly superior to its glutaraldehyde counterpart. In addition, the results obtained in the 3-(4,5-dimethylthiazol-yl)-2,5-diphenyltetrazolium bromide assay showed that reuterin is significantly less cytotoxic than glutaraldehyde. Additionally, it was found that reuterin is an effective crosslinking agent for biological tissue fixation. The reuterin-fixed tissue had comparable free amino group content, denaturation temperature, and resistance against enzymatic degradation as the glutaraldehyde-fixed tissue. In conclusion, the results obtained in this study indicate that reuterin is an effective agent in the sterilization and fixation of biological tissues. PMID- 12115463 TI - Application of magnetic resonance microscopy to tissue engineering: a polylactide model. AB - Absorbable polymers are unique materials that find application as temporary scaffolds in tissue engineering. They are often extremely sensitive to histological processing and, for this reason, studying fragile, tissue-engineered constructs before implantation can be quite difficult. This research investigates the use of noninvasive imaging using magnetic resonance microscopy (MRM) as a tool to enhance the assessment of these cellular constructs. A series of cellular, polylactide constructs was developed and analyzed using a battery of tests, including MRM. Distribution of rat aortic smooth muscle cells within the scaffolds was compared as one example of a tissue engineering MRM application. Cells were loaded in varying amounts using static and dynamic methods. It was found that the cellular component was readily identified and the polymer microstructure readily assessed. Specifically, the MRM results showed a heterogeneous distribution of cells due to static loading and a homogenous distribution associated with dynamic loading, results that were not visible through biochemical tests, scanning electron microscopy, or histological evaluation independently. MRM also allowed differentiation between different levels of cellular loading. The current state of MRM is such that it is extremely useful in the refinement of polymer processing and cell seeding methods. This method has the potential, with technological advances, to be of future use in the characterization of cell-polymer interactions. PMID- 12115464 TI - Sequential robust design methodology and X-ray photoelectron spectroscopy to analyze the grafting of hyaluronic acid to glass substrates. AB - Sequential Robust Design experiments and X-ray photoelectron spectroscopic (XPS) studies were performed to examine the immobilization of hyaluronic acid (HA) on glass substrates chemisorbed with N-(2-aminoethyl)-3-aminopropyl-trimethoxysilane (EDS). Numerous reaction conditions were investigated, including the concentrations of 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC), N-hydroxysulfosuccinimide (Sulfo-NHS), and HA, and the reaction buffer type, concentration, and pH. The elemental surface compositions of carbon and silicon (C/Si ratio) were used to assess the extent of HA immobilization, leading to the identification of critical HA-binding reaction conditions and the determination of an optimum surface chemistry. The optimum chemistry consisted of 200 mM EDC, 50 mM Sulfo-NHS, 10 mM N-(2-hydroxyethyl)piperazine-N'-(2 ethanesulfonic acid) (HEPES) buffer at a pH of 7.0, and 3 mg/mL HA. This work emphasizes the advantages of using Robust Design methods over traditional statistical experimental design, particularly when large numbers of variables are examined and costly analytical techniques are employed. PMID- 12115465 TI - Initial interaction of U2OS cells with noncoated and calcium phosphate coated titanium substrates. AB - From previous studies, we know that calcium phosphate (CaP) coated implants stimulate bone formation compared to uncoated implants. Nevertheless, the mechanism by which substrate surface characteristics affect cell function is unclear. In this study, we examined the initial interaction (30 min to 24 h) of U2OS cells with titanium substrates with or without a CaP coating. The effect of substrate roughness was also studied. When cell attachment was studied, we found that cells attached more readily to rough than to smooth surfaces. Also, more cells attached to the uncoated than to the CaP coated surface. After 24 h, cell numbers were similar for all substrate surfaces. Further, cells spread to a larger area on noncoated titanium than on the CaP coated substrates. At 24 h, the sequence of cell size was smooth titanium > rough titanium > CaP coated titanium. Shape measurements showed differences in cell shape between the cells on the different materials only at 7 h, not at different culture times. Cells expressed alpha2, alpha3, alpha5, alpha6, alphav, and beta1 subunits. Expression of alpha1, alpha4, alphavbeta3, beta3, beta4, and beta7 was extremely low or was not found. The beta1 integrin expression was higher on the coated than on the noncoated titanium at 3 h, but not on the other studied times. Expression of alpha2, alpha5, alpha6, and alphav expression was found to be upregulated at 24 h compared to earlier culture times on coated titanium, but not on uncoated titanium substrates. From this we conclude that the surface characteristics of a material (roughness and composition) can affect the initial interaction of cells with the material. PMID- 12115466 TI - Gene expression analysis of osteoblastic cells contacted by orthopedic implant particles. AB - Particles generated from orthopedic implants through years of wear play an essential role in the aseptic loosening of a prosthesis. We have investigated the biocompatibility of these orthopedic particles on different osteoblast-like cells representative of different stages of osteoblast maturation. We found the particles induced a caspase-dependent apoptosis of osteoblasts, with less mature osteoblasts being the most susceptible. An analysis of gene expression was performed on the less mature osteoblasts, which were in contact with the particles. We found that the particles had a profound impact on genes that code for inflammatory cytokines and genes involved in controlling the nuclear architecture. Results from this study suggest that the peri-implant osteolysis after a total joint replacement can be due in part to a decrease of bone formation and not solely to an overstimulation of bone resorption as is generally proposed. Development of new drugs that promote normal bone formation and osteoblast survival would possibly control peri-implant osteolysis, resulting in a better prognosis for patients with orthopedic implants. PMID- 12115467 TI - The sintered microsphere matrix for bone tissue engineering: in vitro osteoconductivity studies. AB - A tissue engineering approach has been used to design three-dimensional synthetic matrices for bone repair. The osteoconductivity and degradation profile of a novel polymeric bone-graft substitute was evaluated in an in vitro setting. Using the copolymer poly(lactide-co-glycolide) [PLAGA], a sintering technique based on microsphere technology was used to fabricate three-dimensional porous scaffolds for bone regeneration. Osteoblasts and fibroblasts were seeded onto a 50:50 PLAGA scaffold. Morphologic evaluation through scanning electron microscopy demonstrated that both cell types attached and spread over the scaffold. Cells migrated through the matrix using cytoplasmic extensions to bridge the structure. Cross-sectional images indicated that cellular proliferation had penetrated into the matrix approximately 700 microm from the surface. Examination of the surfaces of cell/matrix constructs demonstrated that cellular proliferation had encompassed the pores of the matrix by 14 days of cell culture. With the aim of optimizing polymer composition and polymer molecular weight, a degradation study was conducted utilizing the matrix. The results demonstrate that degradation of the sintered matrix is dependent on molecular weight, copolymer ratio, and pore volume. From this data, it was determined that 75:25 PLAGA with an initial molecular weight of 100,000 has an optimal degradation profile. These studies show that the sintered microsphere matrix has an osteoconductive structure capable of functioning as a cellular scaffold with a degradation profile suitable for bone regeneration. PMID- 12115468 TI - Interaction of oxidation and crosslinking in gamma-irradiated ultrahigh molecular weight polyethylene. AB - The interaction between oxidation and crosslinking in gamma-irradiated ultrahigh molecular-weight polyethylene with and without artificial aging was studied. The effect of the atmosphere during irradiation (air vs. low oxygen) occurred primarily within about 0.5 mm of the surface, that is, the depth to which oxygen had diffused when the polyethylene specimen was machined and when it was irradiated. Irradiation in the presence of oxygen induced oxidation instead of crosslinking, so that the level of crosslinking achieved was lower than that which normally would occur at the same dose in the absence of oxygen. Subsequent artificial aging reduced the gel content (crosslinking) and had a maximal effect on the surface and subsurface regions for the gamma-air and gamma-low oxygen polyethylenes, respectively. Thus the storage environments and durations prior to irradiation and prior to artificial aging must be taken into account when attempting to duplicate the oxidation-crosslinking profiles that occur with actual implants in clinical use. In addition, the oxidation mechanisms initiated by the artificial aging method used in this study (i.e., heating in air to 80 degrees C) initiated somewhat different oxidative reactions from those that occur during prolonged shelf life at room temperature or in vivo. In particular, the formation of a peak of oxidation below the free surface of the polyethylene is due to the combined effects of the distribution of residual free radicals and the diffusion gradient of the oxygen. The interactive relationship between oxidation and crosslinking characterized in the present study provides a fundamental basis for understanding the wear behavior of gamma-sterilized components in past clinical use. It also provides guidelines for the development of polyethylenes with improved resistance to oxidation and wear, with particular relevance to estimation of the amount of crosslinking need- ed to potentially eliminate the clinical problem of osteolysis. PMID- 12115469 TI - Histological characterization of the early stages of bone morphogenetic protein induced osteogenesis. AB - On the basis of currently available knowledge, we hypothesize that the initial bone formation, as induced by bone morphogenetic protein (BMP), is influenced by the chemical composition and three-dimensional spatial configuration of the used carrier material. Therefore, in the current study, the osteoinductive properties of porous titanium (Ti) fiber mesh with a calcium phosphate (Ca-P) coating (Ti CaP), insoluble bone matrix (IBM), fibrous glass membrane (FGM), and porous particles of hydroxy apatite (PPHAP) loaded with rhBMP-2 were compared in a rat ectopic assay model at short implantation periods. Twelve Ti-CaP, 12 IBM, 12 FGM, and 12 PPHAP implants, loaded with rhBMP-2, were subcutaneously placed in 16 Wistar King rats. The rats were sacrificed at 3, 5, 7, and 9 days post-operative, and the implants were retrieved. Histological analysis demonstrated that IBM and Ti-CaP had induced ectopic cartilage and bone formation by 5 and 7 days, respectively. However, in PPHAP, bone formation and cartilage formation were seen together at 7 days. At 9 days, in Ti-CaP, IBM, and PPHAP, cartilage was seen together with trabecular bone. At 9 days, in FGM, only cartilage was observed. Quantitative rating of the tissue response, using a scoring system, demonstrated that the observed differences were statistically significant (Wilcoxon rank sum test, p < 0.05). We conclude that IBM, CaP-coated Ti mesh, FGM, and PPHAP provided with rhBMP-2 can indeed induce ectopic bone formation with a cartilaginous phase in a rat model at short implantation periods. Considering the different chemical composition and three-dimensional spatial configuration of the carrier materials used, these findings even suggest that endochondral ossification is present in rhBMP-2-induced osteogenesis, even though the amount of cartilage may differ. PMID- 12115470 TI - Immunogenicity of polymeric implants: long-term antibody response against polyester (Dacron) following the implantation of vascular prostheses into LEW.1A rats. AB - Implanted biomaterials trigger acute and chronic inflammatory responses directly correlated to the central role of phagocytic cells at the host-implant interface. This study was designed to evaluate specific humoral immune responses following repeated intraperitoneal implantations of collagen-impregnated polyester (Dacron) prosthetic segments into LEWIS rats. Serum antibody detection was performed by enzyme immunoassay with the prosthetic segments as a target. Cutoff values for antibody positivity were greater than or equal to the 99th percentile for control rats. Polymer immunoglobiulin G (IgG) antibodies were significantly increased (p < 0.05) by repeated implantation and were subsequently followed until experimental day 293. Antibody formation was significantly enhanced through the application of complete Freund's adjuvant in combination with the first implantation. All rats within this group were antibody-positive on day 53, but only 6 of 10 animals that received the prosthesis without the adjuvant were. After preincubation of sera with bovine collagen type I (solid phase adsorbed or in solution), polymer antibody binding was discovered not to be diminished, indicating that the IgG antibodies detected were not directed against the prosthesis impregnation. Furthermore, a significant correlation was obtained between polymer antibody binding to collagen-impregnated and nonimpregnated prostheses (r(s) = 0.797, p < 0.001). There was no substantiated correlation between antibody binding to polyester and to an irrelevant polymer (Tecoflex EG 80). We conclude that specific polymer antibodies may indeed provide an additional parameter for biocompatibility testing as well as a possible serological marker of an inflammatory response to implants. PMID- 12115472 TI - Surface modification by alkali and heat treatments in titanium alloys. AB - Pure titanium and titanium alloys are normally used for orthopedic and dental prostheses. Nevertheless, their chemical, biological, and mechanical properties still can be improved by the development of new preparation technologies. This has been the limiting factor for these metals to show low affinity to living bone. The purpose of this study is to improve the bone-bonding ability between titanium alloys and living bone through a chemically activated process and a thermally activated one. Two kinds of titanium alloys, a newly designed Ti-In-Nb Ta alloy and a commercially available Ti-6Al-4V ELI alloy, were used in this study. In this study, surface modification of the titanium alloys by alkali and heat treatments (AHT), alkali treated in 5.0M NaOH solution, and heat treated in vacuum furnace at 600 degrees C, is reported. After AHT, the effects of the AHT on the bone integration property were evaluated in vitro. Surface morphologies of AHT were observed by optical microscopy (OM) and scanning electron microscopy (SEM). Chemical compositional surface changes were investigated by X-ray diffractometry (XRD), energy dispersive spectroscopy (EDS), and auger electron spectroscopy (AES). Titanium alloys with surface modification by AHT showed improved bioactive behavior, and the Ti-In-Nb-Ta alloy had better bioactivity than the Ti-6Al-4V ELI alloy in vitro. PMID- 12115471 TI - Biocompatibility and in vivo gentamicin release from bioactive sol-gel glass implants. AB - Biomaterial pieces, with suitable osteogenic properties for use in the treatment of bone defects and the capability to avoid bone infections, have been synthesized. These materials are composed exclusively of gentamicin sulfate and bioactive SiO(2)-CaO-P(2)O(5) sol-gel glass (previously synthesized). Implant processing was achieved by uniaxial and isostatic pressure of the components mixture. After implanting the pieces into rabbit femur, we studied (i) the antibiotic release, determining the concentration in proximal and distal bone, liver, kidney, and lung as a function of time; and (ii) the bone growth resulting from the glass reactivity in the biologic environment. The results indicate that the implants are good carriers for local gentamicin release in the osseous tissue, exhibiting excellent biocompatibility and bone integration. Moreover, these implants are able to promote bone growth during their resorption process. PMID- 12115473 TI - Effect of fibrin sealant composition on human neutrophil chemotaxis. AB - The use of fibrin sealants offers one of the most physiologically compatible approaches to preventing postoperative adhesions. Although a number of fibrin sealant formulations have been developed, little is known about how the various components of these preparations affect the wound-healing process. Because one of the key steps in wound healing is the migration of phagocytic leukocytes, such as neutrophils, into the site of injury, we performed studies to characterize systematically the effects of various fibrin sealant components on neutrophil chemotaxis. Using a transwell chemotaxis assay, we found that increasing fibrin concentration resulted in an inhibition of the ability of the cells to migrate through the clots in a dose-dependent manner, and at fibrin clot concentrations >2.0 mg/mL chemotaxis was completely blocked. Factor XIII crosslinking of the clots also had a significant impact on neutrophil chemotaxis, and sealant preparations deficient in Factor XIII allowed neutrophil migration at much higher fibrin concentrations. The presence of various other fibrin sealant components such as plasminogen and fibrinolysis inhibitors (aprotinin and tranexamic acid) did not have any significant effects on the ability of neutrophils to migrate through fibrin clots as compared to control clots without these components. Overall, these studies show that the composition of fibrin sealant preparations can significantly affect neutrophil migration into the site of injury, which could possibly influence the wound healing process. PMID- 12115474 TI - Attachment, spreading, and matrix formation by human gingival fibroblasts on porous-structured titanium alloy and calcium polyphosphate substrates. AB - Porous calcium polyphosphate (CPP) structures represent promising resorbable implant systems that can promote anchorage to connective tissues. Previous studies focused on chondrocyte interactions with CPP, but there are limited data on interactions of soft connective tissue cells with these materials. We studied attachment, spreading, and matrix formation by human gingival fibroblasts when cultured on amorphous and crystalline CPP. Comparison with porous Ti6Al4V substrates of similar volume percent, porosity, and pore size distribution provided evaluations of fibroblast interactions with rapid, moderate, and nonbiodegradable systems, respectively. Cells were incubated on substrates in medium containing ascorbic acid and evaluated at 3, 24, 48, 72, and 96 h after plating. Attached cell counts, cytoplasmic actin filament area, and immunostained extracellular type 1 collagen or fibronectin were quantified by morphometric analyses using epifluorescence microscopy. Cell morphology and substrate interactions were evaluated by scanning electron microscopy. Spreading, attachment, and matrix production were similar for both CPP substrates. In contrast, titanium alloy substrates exhibited threefold more attachment and twofold more spreading than CPP substrates. The area per cell of immunostained extracellular collagen and fibronectin was similar for the three different substrates. The results indicate that the crystallinity and, hence, degradation rate of CPP substrates does not substantially affect the interactions of fibroblasts with CPP materials but that compared with titanium alloy substrates, spreading and attachment are inhibited. PMID- 12115475 TI - Synthesis, characterization, and cytocompatibility of elastomeric, biodegradable poly(ester-urethane)ureas based on poly(caprolactone) and putrescine. AB - The engineering of tissue for mechanically demanding applications in the cardiovascular system is likely to require mechanical conditioning of cell scaffold constructs prior to their implantation. Scaffold properties amenable to such an application include high elasticity and strength coupled with controllable biodegradative and cell-adhesive properties. To fulfill such design criteria, we have synthesized a family of poly(ester-urethane)ureas (PEUUs) from polycaprolactone and 1,4-diisocyanatobutane. Lysine ethyl ester (Lys) or putrescine was used as chain extenders. To encourage cell adhesion, PEUUs were surface modified with radio-frequency glow discharge followed by coupling of Arg Gly-Asp-Ser (RGDS). The synthesized PEUUs were highly flexible, with breaking strains of 660-895% and tensile strengths from 9.2-29 MPa. Incubation in aqueous buffer for 8 weeks resulted in mass loss, from >50% (Lys chain extender) to 10% (putrescine chain extender). Human endothelial cells cultured for 4 days with medium containing the degradation products from PEUUs with either the Lys or putrescine chain extender showed no toxic effects. Cell adhesion was 85% of that measured on tissue-culture polystyrene for unmodified PEUU surfaces (p < 0.01) and >160% (p < 0.001) of polystyrene on RGDS-modified PEUUs. These biodegradable PEUUs demonstrate potential for future application as cell scaffolds in cardiovascular tissue-engineering or other soft-tissue applications. PMID- 12115476 TI - Identification of a new proliferation-associated protein NET-1/C4.8 characteristic for a subset of high-grade cervical intraepithelial neoplasia and cervical carcinomas. AB - A large number of genes are known to be differentially expressed at distinct steps of carcinogenesis. By using a cell culture model system for cervical cancer, we had previously identified several genes that were more strongly expressed when comparing normal cervical epithelium with cervical intraepithelial neoplasia (CIN) and cervical cancer. In our study, we show that one of these genes, C4.8, is identical to NET-1, which is a new member of the tetraspanin family of proteins. By generating a mouse polyclonal antiserum against the major extracellular domain of the protein, we could detect NET-1/C4.8 expression both after ectopic expression of the gene in cell cultures and in cryostat sections of cervical biopsies. Moreover, immunohistochemic analyses of normal cervical epithelium, metaplasia, condyloma and CIN of different severity suggest that NET 1/C4.8 expression is associated with neoplastic cell proliferation. Notably, expression of the protein throughout the entire epithelium is only evident for a subset of CIN3. The potential importance for this gene in cervical carcinogenesis is underlined by an invariably strong expression in all undifferentiated squamous cell cancers examined. This indicates that this gene may be of prognostic value. PMID- 12115478 TI - Rat Muc4 (sialomucin complex) reduces binding of anti-ErbB2 antibodies to tumor cell surfaces, a potential mechanism for herceptin resistance. AB - Muc4 (also called sialomucin complex), the rat homolog of human MUC4, is a heterodimeric glycoprotein complex that consists of a peripheral O-glycosylated mucin subunit, ASGP-1, tightly but noncovalently linked to a N-glycosylated transmembrane subunit, ASGP-2. The complex is expressed in a number of normal, vulnerable epithelial tissues, including mammary gland, uterus, colon, cornea and trachea. Muc4/SMC is also overexpressed or aberrantly expressed on a number of human tumors including breast tumors. Overexpression of Muc4/SMC has been shown to block cell-cell and cell-matrix interactions, protect tumor cells from immune surveillance and promote metastasis. In addition, as a ligand for ErbB2, Muc4/SMC can potentiate phosphorylation of ErbB2 and potentially alter signals generated from this receptor. Using A375 human melanoma cells and MCF7 human breast adenocarcinoma cells stably transfected with tetracycline regulatable Muc4, we have investigated whether overexpression of Muc4/SMC can repress antibody binding to cell surface-expressed ErbB2. Overexpression of Muc4/SMC does not affect the level of ErbB2 expression in either cell line, but it does reduce binding of a number of anti-ErbB2 antibodies, including Herceptin. Interestingly, overexpression of ErbB2 does not block binding of other unrelated antibodies of the same isotype, suggesting that the reduction in ErbB2 antibody binding is due to complex formation of Muc4/SMC and ErbB2. Furthermore, capping of Muc4/SMC with anti-Muc4/SMC antibodies reduces antibody binding to ErbB2 instead of increasing binding, again suggesting that reduced antibody binding to ErbB2 is due to steric hindrance from complex formation of Muc4/SMC and ErbB2. Thus, overexpression of Muc4/SMC on tumor cells may have both prognostic and therapeutic relevance. PMID- 12115477 TI - Transcript profiling of enzymes involved in detoxification of xenobiotics and reactive oxygen in human normal and simian virus 40 T antigen-immortalized oral keratinocytes. AB - The metabolic detoxification capacity may critically regulate the susceptibility of human tissues to cancer development. We used standardized and quantitative, reverse transcription-polymerase chain reaction (StaRT-PCR) and microarray chip techniques to analyze transcript levels of multiple detoxification enzymes in cultured normal human oral keratinocytes (NOK) and the Siman virus 40 T antigen immortalized oral keratinocyte line SVpgC2a, viewing the latter as a model of a benign tumor state. With good agreement between the 2 methodologies, NOK and SVpgC2a were found to express transcripts for cytochrome P450 enzymes (CYPs), factors related to CYP induction and enzymes involved in conjugation reactions or detoxification of reactive oxygen. The cell types expressed similar levels of CYP 2B6/7, CYP 2E1, P450 oxidoreductase, the aryl hydrocarbon receptor nuclear translocator, sulfotransferase 1A1, sulfotransferase 1A3, epoxide hydrolase, glutathione S-transferase M3, glutathione S-transferase pi and catalase, superoxide dismutase 1, glutathione peroxidase 1 and glutathione peroxidase 3. In contrast, SVpgC2a exhibited comparatively higher levels of CYP1A1, 1B1, aryl hydrocarbon receptor, glutathione S-transferase M1, 2, 4, 5, glutathione S transferase theta 1 and superoxide dismutase 2 and comparatively lower levels of UDP glycosyltransferase 2 and microsomal glutathione S-transferase 1. Some transcripts, e.g., CYP 2A6/7, were not detected by either standard, non quantitative RT-PCR or the above methods, whereas others were barely quantifiable by StaRT-PCR, i.e., were present at 1-10 molecules/10(6) molecules of actin. Overall, the expression analysis demonstrated presence of mRNA for multiple enzymes involved in foreign compound metabolism and detoxification pathways, including several enzymes not previously reported for oral epithelium. Generally, the comparison of NOK from 2 individuals indicated relatively similar transcript levels of these enzymes. In contrast, differences between NOK and SVpgC2a, e.g., for CYP1B1, may reflect alteration caused by immortalization and aid identification of early stage tumor markers in oral epithelium. PMID- 12115479 TI - Stromelysin-3 expression by mammary tumor-associated fibroblasts under in vitro breast cancer cell induction. AB - To understand better the influence of the host stromal phenotype on stromal expression of stromelysin-3 (ST3) in breast cancer, we have investigated ST3 expression by host stromal cells isolated from 9 different primary breast carcinomas. These tumor-associated fibroblasts were cocultivated with 3 epithelial cancer cell lines of mammary origin (MDA-MB-231, SK-BR-3 and MCF-7), as well as with normal human mammary epithelial cells (NME and 184A1) and keratinocytes, using both direct and indirect coculture systems. ST3 expression was demonstrated by both in situ hybridization and immunocytochemistry. The results showed that ST3 expression by stromal cells was cancer-specific. Indeed, ST3 expression by tumor-associated stromal cells was induced by 3 malignant cancer cell lines (MDA-MB-231, SK-BR-3 and MCF-7), whereas no ST3 was expressed under normal mammary epithelial cell stimulation. ST3 expression was weak or absent in unstimulated tumor-associated fibroblasts. However, after direct coculture with cancer cells, expression of ST3 transcripts reappeared in 8 of the 9 cases and was observed only in fibroblasts located in close contact with tumor cells. Under similar coculture conditions and using the same cancer cell line stimulation, ST3 expression was, however, quite variable among these 9 cases, reflecting the difference of protease expression observed on the sections of the original tumors. Tumor induction of ST3 expression was much more important by direct cell-cell contact than by indirect stimulation and was not influenced by the addition of basic fibroblast growth factor (bFGF) and anti-bFGF to the culture medium. Our results suggest that the host stromal cell phenotype may significantly influence host stromal cell protease expression under cancer cell stimulation. PMID- 12115480 TI - Involvement of alphavbeta 3 integrin and disruption of endothelial fibronectin network during the adhesion of the human ovarian adenocarcinoma cell line IGROV1 on the human umbilical vein cell extracellular matrix. AB - Like the majority of tumor cells, ovarian cancer cell growth is critically dependent on their neovascularization. Adhesion molecules and cellular events that lead to ovarian tumor cell interactions with endothelial extracellular matrix surrounding the vasculature are poorly identified. To understand the role of alphavbeta3 integrin and its ligand fibronectin in this process, we used in vitro coculture models with IGROV1 human ovarian adenocarcinoma cell line and human umbilical vein endothelial cells (HUVEC). Adhesion assays revealed a strong ability of IGROV1 cells to adhere to HUVEC-ECM. alphavbeta3 is mainly implicated and seems to cooperate with alpha5beta1 integrin in this event. Immunofluorescence staining revealed the presence of alphavbeta3 and alpha5beta1 in IGROV1 cells adhering on HUVEC-ECM at regions of cell sub-stratum contacts. Furthermore, our data showed the absence of fibronectin staining in IGROV1 cells and the disruption of the HUVEC-ECM fibrillar fibronectin network under IGROV1 cell influence. In situ experiments in ovarian neoplastic tissue corroborated the absence of fibronectin in the tumor and its strong detection in vasculature. These findings suggest the active participation of alphavbeta3 and alpha5beta1 integrins and the reorganization of endothelial fibronectin during the adhesion of IGROV1 cells to HUVEC-ECM whereas IGROV1 cells seem to be unable to synthesize fibronectin. Thus, fibronectin integrin receptors expressed by ovarian tumor cells and endothelial fibronectin may be of importance in ovarian carcinoma neovascularization and during tumor-vasculature interactions. PMID- 12115481 TI - Possible role of galectin-9 in cell aggregation and apoptosis of human melanoma cell lines and its clinical significance. AB - Galectin-9 expression was examined in 6 human melanoma cell lines. Among them, MM BP proliferated with colony formation, but MM-RU failed. RT-PCR analysis revealed evident expression of galectin-9 mRNA in MM-BP but not in MM-RU. MM-BP expressed galectin-9 protein both on the surface and in the cytoplasm, whereas MM-RU expressed it only weakly in the cytoplasm. Exogenous galectin-9 induced in vitro both cell aggregation and apoptosis of MM-RU proliferating without colony formation. Association of galectin-9 expression in melanoma cells with prognosis of the patients bearing melanocytic tumors was further examined. Galectin-9 protein was strongly and homogeneously expressed in melanocytic nevi, but down regulated in melanoma cells especially in metastatic lesions. High galectin-9 expression was inversely correlated with the progression of this disease, suggesting that high galectin-9 expression in primary melanoma lesions links to a better prognosis. PMID- 12115482 TI - DNA topoisomerase IIalpha predicts progression-free and overall survival in pediatric malignant non-brainstem gliomas. AB - Malignant non-brainstem glioma (MNBG) is a rare pediatric brain tumor. The prognosis for children harboring this lesion remains largely unpredictable. Assessment of histologic features alone only provides a marginal insight into the biologic behavior of these lesions. Hence, the identification of novel molecular markers capable of characterizing these lesions more accurately with respect to their biologic aggressiveness is definitely needed. Our current study examined the expression of nuclear DNA topoisomerase IIalpha (TIIalpha), a novel marker of cell cycle turnover and a determinant of tumor cell resistance to chemotherapy, in a series of 17 archival pediatric MNBGs. TIIalpha expression was found to extend over a wide range in the study cohort (3.9-69.1%). A cutoff labeling index of 12% was found to define 2 prognostic subgroups (TIIalpha <12 vs. >or=12) with profoundly different 5-year progression-free survival (60% vs. 8%; p = 0.0108, log-rank test) and overall survival (100% vs. 8%; p = 0.0038) rates. TIIalpha expression was significantly linked to MIB-1 antibody labeling of the Ki-67 nuclear antigen (R = 0.919, p < 0.001). A high TIIalpha labeling index remained associated with short progression-free survival (p = 0.022) and overall survival (p = 0.022) in multivariate analysis (Cox regression). In conclusion, considering that TIIalpha expression was not related to histopathologic grade, biological characteristics as assessed by TIIalpha labeling may complement the information obtained by tumor morphology as a means of improving the accuracy of patient prognosis prediction. PMID- 12115483 TI - Vasculogenic mimicry and pseudo-comedo formation in breast cancer. AB - Tumors require a blood supply for growth and hematogenous metastases. Until recently, most research in this area has focused on the role of angiogenesis, the recruitment of new vessels into a tumor from preexisting vessels. Previously, in a study of breast cancer (IBC), in which we used established inflammatory breast cancer (IBC) xenografts (WIBC-9) originating from a patient with IBC (Shirakawa et al., Cancer Res 2001:61:445-451), we reported observing vasculogenic mimicry (VM), a condition in which bloodstreams within cancer tissue are not accompanied by a lining of endothelial cells (ECs) (Shirakawa et al., Cancer Res 2002:62:560 566). In the present study, we examined 331 surgically resected breast cancer specimens for evidence of VM, using immunohistochemistry and laser-captured microdissection (LCM) followed by nested reverse transcriptase polymerase chain reaction (RT-PCR). Surprisingly, 7.9% (26 specimens) of the 331 specimens exhibited evidence of VM. Of these 26 VM specimens, 84.6% (22 specimens) exhibited pseudo-comedo formation. RT-PCR analysis of 8 microdissected typical VM specimens revealed expression of Tie-2, Flt-1, thrombin receptor and CD31 in 63, 50, 0 and 0% of specimens, respectively. In contrast, results of RT-PCR analysis of 8 specimens from non-VM tumors were negative for expression of these genes. The 26 VM cases tended to have a higher percentage of hematogenous recurrence (p = 0.059) and a lower percentage of 5-year survival (p = 0.071) than the 305 non VM cases. However, there were no significant differences in tumor size, lymph node metastasis, estrogen receptors or progesterone receptors between the 2 groups (p > 0.1). Our results suggest that the existence of VM increases the likelihood of hematogenous metastases and is in inverse proportion to prognosis. PMID- 12115484 TI - Loss of heterozygosity on chromosome 10q is associated with earlier onset sporadic colorectal adenocarcinoma. AB - Recent studies have shown that loss of heterozygosity (LOH) on chromosome 10q is a frequent event in a number of tumour types including colorectal cancers. Because previous studies have used markers located mainly distally on chromosome 10, we have examined 114 sporadic colorectal adenocarcinomas for LOH using a panel of 9 highly polymorphic microsatellite markers spanning the long arm of chromosome 10. Using microdissected tumour material, LOH of one or more chromosome 10q markers was a frequent event (75 of 114; 66%). The highest frequency of loss (42 of 96; 44%) was observed at the marker D10S1790 located at 10q21.1. The mean age of presentation, of patients with LOH of D10S1790 was significantly (p = 0.0006) lower (67.1 years) compared to patients with retention of this marker (73.5 years). When we compared frequency of loss at this marker in patients presenting before 70 years of age (68%) to those above 70 years (23%) we observed a significant difference (p < 0.0001). Statistical analysis between loss, or instability at other markers and clinicopathological features did not show any significant associations. In addition LOH at D10S1790 was infrequent in adenomas (2 of 20; 10%) compared to adenocarcinomas (42 of 96; 44%) (p = 0.0047), suggesting that loss within this region is a late event in colorectal tumorigenesis. The association of loss at D10S1790 and an earlier age of presentation in adenocarcinomas suggests that this locus may harbor a tumour suppressor gene(s), which affects the rate of colorectal tumour progression. Identification of this region of genetic loss further refines our understanding of the paradigm in this tumour type of multiple-steps responsible for initiation and progression. PMID- 12115485 TI - Analysis of mutations and identification of several polymorphisms in the putative promoter region of the P34CDC2-related CDC2L1 gene located at 1P36 in melanoma cell lines and melanoma families. AB - Chromosome 1 abnormalities are the most commonly detected aberrations in many cancers including malignant melanoma. Partial deletions and an allelic loss of the chromosome 1p36 region observed in melanoma indicate the presence of putative tumor suppressor gene(s) in this region. A candidate gene, CDC2L1, which encodes PITSLRE proteins related to p34(cdc2), is mapped to 1p36. To determine whether CDC2L1 mutation is involved in melanoma development, we examined 20 melanoma cell lines and 11 members of melanoma-prone families linked to chromosome 1p36. Mutation analysis throughout the entire coding region of the CDC2L1 gene revealed only 1 mutation (C-->T at nucleotide location 97 of exon 7, Ser-->Leu) in the melanoma cell line UACC 903 out of 20 melanoma cell lines and 6 melanoma cases. However, 4 polymorphic nucleotide changes, C-48T, G-53C, T-103C and T-210C, in the putative promoter region of CDC2L1 were identified. The 4 variants were located within or beside the conserved binding sites of transcription factors TCF11, MZF1 and TAAC box, indicating their potential effects on the regulation of CDC2L1 expression. No aberrant methylation of the CDC2L1 CpG island in the promoter region was observed by sodium bisulfite genomic sequencing. These results indicate that mutations are rare in the CDC2L1 gene in these melanoma cell lines and melanoma families and that the aberrant cytosine methylation of the CDC2L1 CpG island is not the mechanism of CDC2L1 repression in melanoma. The contribution of 4 promoter polymorphisms to the transcriptional regulation of the gene and its association with melanoma warrants further investigation. PMID- 12115486 TI - CT7 (MAGE-C1) antigen expression in normal and neoplastic tissues. AB - CT7 (MAGE-C1) is a member of the cancer testis (CT) antigen family. The present study describes the generation of CT7-33, a monoclonal antibody (MAb) to CT7, and the preliminary protein expression analysis of CT7 in normal tissues and in a limited number of neoplastic lesions. CT7-33 was effective in frozen as well as formalin-fixed, paraffin-embedded tissues, and immunohistochemistry/reverse transcriptase polymerase chain reaction (RT-PCR) co-typing demonstrated antibody specificity. CT7-33 immunoreactivity in normal adult tissues is restricted to testicular germ cells. In neoplastic lesions, CT7-33 immunostaining is confined to tumor cells, and the frequency of CT7 protein expression mostly parallels previous mRNA analyses. Whereas colorectal and renal cell carcinomas, as well as sarcomas, exhibit poor or no CT7-33 staining, carcinomas of the mammary gland and ovary, nonsmall cell lung carcinoma and metastatic melanomas exhibit a high incidence of CT7 protein expression. However, as seen in previous analyses of other CT antigens, the expression pattern is mostly heterogeneous, and tumors with more than 50% of tumor staining are only infrequently encountered. In summary, our study presents a new serologic reagent for the analysis of CT7 on a protein level and confirms what is known with regard to the expression pattern of other CT antigens in tumors: frequent heterogeneity of antigen expression. PMID- 12115487 TI - Genistein, a soy isoflavone, induces glutathione peroxidase in the human prostate cancer cell lines LNCaP and PC-3. AB - Genistein is a major component of soybean isoflavone and has multiple functions resulting in antitumor effects. Prostate cancer is 1 of the targets for the preventive role of genistein. We examined the effect of genistein on human prostate cancer (LNCaP and PC-3) cells. Proliferation of both cell lines was inhibited by genistein treatment in a dose-dependent manner. To obtain the gene expression profile of genistein in LNCaP cells, we performed cDNA microarray analysis. The expression of many genes, including apoptosis inhibitor (survivin), DNA topoisomerase II, cell division cycle 6 (CDC6) and mitogen-activated protein kinase 6 (MAPK 6), was downregulated. Expression levels were increased more than 2-fold in only 4 genes. The glutathione peroxidase (GPx)-1 gene expression level was the most upregulated. Quantitative real-time polymerase chain reaction revealed significant elevation of transcript levels of GPx-1 in both LNCaP and PC 3 cells. Upregulation of gene expression levels accompanied elevation of GPx enzyme activities. In contrast, no significant changes were observed in the gene expression levels and enzyme activities of the other antioxidant enzymes, superoxide dismutase and catalase. GPx activation might be one of the important characteristics of the effects of genistein on prostate cancer cells. PMID- 12115489 TI - Time trends incidence of both major histologic types of esophageal carcinomas in selected countries, 1973-1995. AB - The purpose of our study was to examine the incidence patterns of 2 major histologic types of esophageal cancer, in selected countries world-wide and to identify components of birth cohort, period and age as determinants of observed time trends using regression modeling. The roles of temporal changes in specification of histology of tumors and of classification of cancers at the gastroesophageal junction as esophageal or gastric in origin were taken into consideration. In all, 56,426 esophageal cancer cases were included. The results indicate that the incidence rate of squamous cell carcinoma of the esophagus has been relatively stable in most of the countries analyzed, although increasing trends were observed in Denmark and the Netherlands (Eindhoven) among men and in Canada, Scotland and Switzerland among women. There was a significant increase in the incidence of esophageal adenocarcinomas in both sexes in the United States (among whites and blacks), Canada and South Australia and in 6 European countries (Scotland, Denmark, Iceland, Finland, Sweden and Norway). In France the increase was limited to men and in Switzerland the increase was observed only in women. Modeling was unable to distinguish which trends were the results of changes in risk between generations (as cohort effects), or changes in all age groups simultaneously (as a period effect). PMID- 12115488 TI - Diagnostic accuracy of stereotactic large-core needle biopsy for nonpalpable breast disease: results of a multicenter prospective study with 95% surgical confirmation. AB - Stereotactic large-core needle biopsy is increasingly applied for the diagnosis of nonpalpable breast disease. Our study examines whether this minimally invasive technique is sufficiently accurate to replace surgical breast biopsy. In a prospective multicenter study, 973 consecutive women with 1,029 nonpalpable breast lesions were offered stereotactic 14-gauge needle biopsy. If the needle biopsy yielded breast cancer, the patient was offered therapeutic surgery. Surgical biopsy was proposed in cases of needle biopsies without malignancy. An expert panel reviewed all discrepancies in histologic diagnosis between the needle biopsy and open biopsy. Forty-five patients withdrew from participation and 113 (11%) planned needle biopsy procedures were cancelled. Of the 871 successful biopsy procedures, 95% were confirmed surgically. In 13 cases (1.5%), insufficient material was obtained for histologic assessment. Fifty-five percent of the needle biopsies were diagnosed as malignant (290 invasive cancers, 190 ductal carcinoma in situ). Thirteen of the 322 lesions (4%, 95% CI 2-7%) with a benign needle biopsy diagnosis contained malignancy after surgery. Six of the 26 (23%, 95% CI 9-44%) lesions with a high-risk diagnosis (atypical ductal or lobular hyperplasia or lobular carcinoma in situ) were diagnosed as malignant after surgery. Five of the 30 lesions containing normal breast tissue held malignancy (17%, 95% CI 6-35%). Guidelines for the management of different categories of needle biopsy diagnoses were made. Application of these guidelines to the present findings resulted in sensitivity and specificity rates of 97% (95% CI 95-98%) and 99% (95% CI 97-100%), respectively. Stereotactic large-core needle biopsy is an accurate diagnostic instrument for nonpalpable breast disease. It may safely replace needle localised open-breast biopsy provided that high-risk and normal breast tissue diagnoses are followed by needle or open-breast biopsy. PMID- 12115490 TI - A variant of the DNA repair gene XRCC3 and risk of squamous cell carcinoma of the head and neck: a case-control analysis. AB - Individuals differ in their ability to repair DNA damage induced by carcinogens. Studies have shown that polymorphisms in DNA repair genes contribute to individual variation in DNA repair capacity and cancer risk. In a hospital-based case-control study, we tested the hypothesis that a C to T variant (Thr241Met) of DNA repair gene X-ray repair cross-complementing group 3 (XRCC3) is associated with risk of developing squamous cell carcinoma of the head and neck (SCCHN). We genotyped for this variant in 367 non-Hispanic white patients newly diagnosed with SCCHN and 354 frequency-matched cancer-free controls. Compared with the XRCC3 18067CC and 18607CT genotypes, the variant XRCC3 18067TT genotype was associated with a non-statistically significantly increased risk of SCCHN (adjusted odds ratio [ORadj], 1.36; 95% confidence interval [CI], 0.89-2.08), but this risk was significantly increased among female subjects (ORadj 2.23, 95% CI, 1.00-4.98) and current smokers (ORadj, 2.26; 95% CI, 1.02-4.99). These findings suggest that the variant XRCC3 18067TT genotype may not play a major role in the etiology of SCCHN but may contribute to a subset of SCCHN. Larger studies are needed to verify these findings. PMID- 12115491 TI - Cancer risks in twins: results from the Swedish family-cancer database. AB - Twin studies on cancer have addressed two general questions, one about the possible carcinogenic effects of twinning and the second about heritable effects of cancer. The first question is answered by comparing the occurrence of cancer in twins to that in singletons; the second is answered in probandwise analysis of monozygotic twins compared to dizygotic twins or siblings. We used the nationwide Swedish Family-Cancer Database on 10.2 million individuals and 62,574 0-66-year old twins to calculate standardized incidence ratios (SIRs) and 95% confidence intervals (CIs) for all main cancer compared to cancer in singletons. In probandwise analysis, the SIR was calculated for the co-twin of an affected twin. The overall risk of cancer in same or opposite sex twins was at the level of the risk for singletons. Testicular cancer was increased among same sex twins and all twins to an SIR of 1.43. Melanoma was decreased in these groups of twins to an SIR of 0.84. Some other cancer sites were increased or decreased in some groups of twins, but none in all twins. The SIR of breast cancer was 1.01 and 1.04 in same and opposite sex twins, respectively. Probandwise analysis showed increased risks for Hodgkin's disease in males and breast cancer and childhood acute lymphoid leukemia among females. The data on this unselected population of twins suggest that twinning per se is not a risk factor of cancer. However, because twins are smaller than singletons at birth, some possible effects could be masked by such differences. In utero hormonal exposures may be related to the risk of testicular cancer. The protective effects in melanoma may be due to socioeconomic factors. PMID- 12115492 TI - Sophoranone, extracted from a traditional Chinese medicine Shan Dou Gen, induces apoptosis in human leukemia U937 cells via formation of reactive oxygen species and opening of mitochondrial permeability transition pores. AB - Screening of various natural products in a search for novel inducers of apoptosis in human leukemia cells led us to identify the strong apoptosis-inducing activity in a fraction extracted with methanol from the roots of Sophora subprostrata Chun et T. Chen. We purified the compound that induced apoptosis in human leukemia cells and identified it as sophoranone. Sophoranone inhibited cell growth and induced apoptosis in various lines of cells from human solid tumors, with 50% inhibition of growth of human stomach cancer MKN7 cells at 1.2 +/- 0.3 microM. The growth-inhibitory and apoptosis-inducing activities of sophoranone for leukemia U937 cells were very much stronger than those of other flavonoids, such as daidzein, genistein and quercetin. At the early stages of treatment of U937 cells with sophoranone, reactive oxygen species were formed, mitochondrial permeability pores were opened and cytochrome c was released from mitochondria. Cytochrome c was also released upon treatment of isolated mitochondria with sophoranone. Inhibitors of complexes III and IV, but not complexes I and II, of the mitochondrial respiratory chain prevented the release of cytochrome c from isolated mitochondria by sophoranone, as well as the induction of apoptosis in U937 cells in response to sophoranone. Our results indicate that sophoranone might be a unique apoptosis-inducing anticancer agent that targets mitochondria. PMID- 12115494 TI - Transformation by inorganic arsenic compounds of normal Syrian hamster embryo cells into a neoplastic state in which they become anchorage-independent and cause tumors in newborn hamsters. AB - Arsenic is a known human carcinogen, but little evidence exists for its carcinogenicity in animals. In order to investigate the ability of inorganic arsenics to transform normal cells into a neoplastic state, mass cultures of normal, diploid Syrian hamster embryo (SHE) cells exposed to various concentrations of sodium arsenite or sodium arsenate for 48 hr were continually passaged and tested for neoplastic transformation, as determined by anchorage independent growth in semisolid agar and tumorigenicity in newborn hamsters. Twenty-one of 22 (96%) untreated, control cultures senesced by 20 passages. While 1 culture escaped senescence, it did not acquire the ability to either grow in semisolid agar or form tumors in animals. Ten of 14 (71%) cultures exposed to sodium arsenite or sodium arsenate escaped senescence. Nine of the 10 (90%) arsenic-treated immortal cultures acquired the anchorage-independent phenotype. Five of 5 anchorage-independent cultures examined were tumorigenic. Two of 3 morphologically transformed colonies induced by sodium arsenate also acquired the ability to grow in semisolid agar when isolated. Amplification of the c-myc or c Ha-ras oncogene was detected in 3 of 5 and 4 of 5 tumorigenic cell lines, respectively. Both c-myc and c-Ha-ras were amplified even in a preneoplastic, anchorage-dependent cell line, but neither was amplified in 6 of 9 anchorage independent cell lines. Overexpression of c-myc and c-Ha-ras mRNA was observed in most of the neoplastically transformed cell lines but not in the preneoplastic cell line. Experiments using the methylation-sensitive restriction endonuclease isoschizomers HpaII and MspI revealed hypomethylation of c-myc and c-Ha-ras in the 5'-CCGG sequence of arsenic-exposed cell lines but not in the parental SHE cells or a spontaneously transformed cell line. Thus, inorganic arsenics induce neoplastic transformation of normal, diploid mammalian cells. Overexpression of oncogenes by DNA hypomethylation may participate in the arsenic-induced neoplastic transformation of mammalian cells. PMID- 12115495 TI - Promiscuous expression of Epstein-Barr virus genes in Burkitt's lymphoma from the central African country Malawi. AB - Primary BL in Malawian children has a very high frequency association, approaching 100%, with the human herpesvirus EBV. A detailed study carried out on viral gene expression in these tumours, using both fresh material and methanol fixed FNAs, showed, contrary to prediction, that most belong to a variant "class II" latency category, with lytic cycle-related genes also expressed. That is, in addition to EBNA1 expression, membrane proteins (LMP1/2A), immediate early (BZLF1) and early (IR2 and IR4) genes, a putative viral oncogene (BARF1), CST (BART) antisense transcripts and the viral bcl-2 homologue are expressed in a high proportion of the BLs. Most, but not all, express the small viral (EBER) RNAs. Two other significant observations were made: (i) in addition to expression of cellular cytokine (IL-10) transcripts in all tumours investigated, the normally silent viral IL-10 homologue was expressed in some tumours; (ii) whereas EBNA1 expression from its restricted Qp promoter was generally observed, the nonrestricted Cp/Wp promoter was also active in some tumours. Viral gene expression in the Malawian [endemic (e)] BLs appears to be more promiscuous than predicted from other studies, but expression accords with the cytopathologic picture of eBLs as a rapidly proliferating cell population accompanied by considerable necrosis, and a clinically diverse disease. A small-scale study of relapse Malawian BLs revealed a different picture of viral association, more akin to systemic BL than eBL, where EBV appears to be absent or present only at very low levels. The significance of these findings is considered. PMID- 12115497 TI - Expression of androgen receptor and 5alpha-reductases in the human normal endometrium and its disorders. AB - Androgen metabolism and actions are considered to play a very important role in the development and progression of the normal human endometrium and its disorders. Details regarding androgen actions in these tissues, however, have not been well studied. We first immunolocalized the androgen receptor (AR) and 5alpha reductases, which catalyze the conversion of testosterone to the bioactive and potent androgen, 5alpha-dihydrotestesterone (DHT), in 18 normal cycling human endometria, 27 endometrial hyperplasia and 46 endometrioid endometrial adenocarcinomas. We also examined the mRNA expression of AR and 5alpha-reductases in 7 cases of endometrioid endometrial adenocarcinomas using reverse transcription polymerase chain reaction (RT-PCR). In the normal human endometrium, AR was immunolocalized predominantly in stromal cells of the proliferative phase of the menstrual cycle and in epithelial cells of the secretory phase, whereas 5alpha-reductase types 1 and 2 immunoreactivities were detected in the cytoplasm of epithelial cells but not in stromal cells throughout all phases of the menstrual cycle. In endometrial hyperplasia, the median labeling index (LI) for AR was 48.1%, whereas positive immunostaining for 5alpha reductase Type 1 and Type 2 was detected in only 1 case. In endometrial carcinoma, AR immunoreactivity was detected in the nuclei of carcinoma cells and the number of positive cases was 39/44 (88.6%). Median LI for AR was 36.1%. Immunoreactivity for 5alpha-reductase Type 1 and Type 2 was detected in the cytoplasm of carcinoma cells and the number of positive cases was 37/44 cases (84.1%) and 34/44 (77.3%) for 5alpha-reductase Types 1 and 2, respectively. A significant positive correlation was detected between 5alpha-reductase Type 1 and Type 2 immunoreactivity (p < 0.001). AR LI was not correlated with the presence or absence of Type 1 and Type 2 5alpha-reductases. Results from our RT-PCR studies were consistent with those of immunohistochemistry. These results suggest that DHT may play more important roles than testosterone in the regulation of androgen action in endometrial cancer and normal human endometrium, especially in the secretory phase, in which both AR and 5alpha-reductase are increased. Androgenic actions may be also regulated predominantly by serum testosterone and not by DHT in endometrial hyperplasia because of the absence of 5alpha-reductases in the site of its actions. PMID- 12115496 TI - EBV-expressing AGS gastric carcinoma cell sublines present increased motility and invasiveness. AB - Tumor invasion marks a critical point in cancer progression; it is a harbinger of morbidity and mortality. Thus, the cellular events that enable the invasive phenotype are under intense investigation. Epstein-Barr virus (EBV) is associated with a number of cancers, including Burkitt lymphoma (BL) and nasopharyngeal carcinoma (NPC) and is suspected to contribute to their tumorigenesis. On average, 8% of gastric carcinomas have been shown to carry this virus. To explore whether the presence of EBV in gastric carcinoma contributes to tumor progression in this predominantly invasive carcinoma, we examined a panel of 2 in vitro EBV infected human gastric cancer cell line sublines and their mock-infected AGS parental control line. We found EBV infection caused a marked increase in transmigration of a Matrigel barrier (415% and 303%, p < 0.05, for the 2 infected lines). This correlated with increased motility of these sublines (233% and 140%, p < 0.05). As this pattern of increased motility leading to a more pronounced enhancement of invasion has been noted in other tumor cells, we explored the roles of autocrine signaling pathways previously implicated in carcinoma motility and invasion. Inhibitors to the epidermal growth factor receptor (EGFR) (PD153035), phospholipase C (PLC) (U73122), extracellular-signal regulated kinase (ERK)/mitogen-activated protein kinase (MAPK) (PD089035) and PI-3 kinase (Wortmannin) were not informative. These data suggest that EBV increases migration of AGS cells by a mechanism independent of these autocrine growth factor-induced pathways. Instead, we found that the EBV-infected cells presented increased focal adhesion kinase (FAK) phosphorylation. These findings suggest a role for integrin-mediated signaling in promoting EBV-associated invasiveness. PMID- 12115498 TI - Flavopiridol inhibits vascular endothelial growth factor production induced by hypoxia or picolinic acid in human neuroblastoma. AB - Human neuroblastoma (NB) tumors elaborate angiogenic peptides, and enhanced angiogenesis correlates with their aggressive behavior, metastatic spread and poor clinical outcome. Hence, inhibition of angiogenic factor production may represent a potential therapeutic target for NB treatment. There is currently little information regarding the stimuli that control NB production of angiogenic mediators. In this study, we analyzed the effects of hypoxia, a common feature of solid tumors and a major drive to tumor angiogenesis, and of PA, a tryptophan catabolite produced under inflammatory conditions and endowed with several biologic properties, on the production of the angiogenic activator VEGF by advanced-stage human NB cell lines. We demonstrate that both stimuli are potent inducers of VEGF expression and secretion. VEGF upregulation by PA involved iron chelation because iron sulfate prevented this effect whereas the iron-chelating agent DFX induced VEGF production. Conversely, the CDK inhibitor Flp completely blocked VEGF induction by hypoxia. This effect occurred as early as 3 hr after stimulation and did not require de novo protein synthesis. Moreover, Flp exerted similar inhibitory activity on VEGF induction by PA or DFX, suggesting that this compound targets an essential step in the signaling pathway that leads to VEGF expression. Our findings demonstrate that PA can modulate angiogenic factor production by tumor cells and establish the importance of Flp as an inhibitor of VEGF production by human NB. PMID- 12115499 TI - Common and differential chemokine expression patterns in rs cells of NLP, EBV positive and negative classical Hodgkin lymphomas. AB - Hodgkin lymphoma (HL) is characterized by a minority of neoplastic cells, the so called Reed-Sternberg (RS) cells and a vast majority of reactive cells. RS cells produce chemokines that can attract subsets of peripheral blood cells into HL tissues. To gain insight in the chemokines involved in HL, 16 chemokines were selected based on their ability to recruit different subsets of cells. Five HL, 5 non-HL-derived cell lines, 22 HL, 5 non-HL and 3 control tissues were analyzed by reverse transcriptase-polymerase chain reaction (RT-PCR). Products for 13 of these 16 chemokines were detected in 1 or more of the cell lines tested. No or only very faint signals were obtained in HL for CXCL12, CCL7 and CCL8, but CXCL10, CCL5, CCL13, CCL17 and CCL22 were highly or differentially expressed in HL cell lines and tissues. Immunohistochemistry was performed with antibodies reactive with the latter 5 chemokines on paraffin sections of 21 cases of HL. CCL17 and CCL22 had the highest signals in RS cells at gene expression and at protein levels. CCL17 was specific for the classic HL subtypes, whereas CCL22 also had low signals in NLP samples, as well as in some non-HL. CXCL10 was expressed in a large proportion of HL cases with a predominant expression in EBV positive cases. The results indicate that RS cells produce a complex pattern of chemokines that are involved in the recruitment of reactive cells and contribute to the paradox of an extensive but ineffective host immune response. PMID- 12115500 TI - Signal processing in migrating T24 human bladder carcinoma cells: role of the autocrine interleukin-8 loop. AB - T24 human bladder carcinoma cells reveal a high locomotor activity (70% locomoting cells) within a 3-dimensional collagen matrix. This high migratory activity is induced by an autocrine engagement of the interleukin-8 receptor A, as was shown by antibodies neutralizing the secreted interleukin-8. Treatment of the cells with these specific antibodies reduced the locomotor activity by half. The intracellular signal transduction underlying the interleukin-8-induced T24 locomotion involves the activity of protein tyrosine kinases (PTKs), the phospholipase Cgamma (PLCgamma) and the protein kinase C (PKC), as proven by the use of specific enzyme inhibitors. These results suggest the following model for the regulatory signal transduction of interleukin-8-induced human T24 bladder carcinoma cell migration: The engagement of the interleukin-8-receptor, a receptor of the serpentine family, leads to the beta-arrestin-mediated activation of PTKs. These kinases phosphorylate the PLCgamma, which generates the second messengers diacylglycerol (DAG) and inositol-1,4,5-trisphosphate (IP(3)). DAG activates the PKC, whereas IP(3) mediates the release of calcium from the endoplasmatic reticulum. By means of confocal laser microscopy, we observed an oscillation of the cytosolic calcium concentration in migrating T24 cells, which were loaded with the calcium-dye fluo-3/AM. Here, we report on a new autocrine function of secreted interleukin-8 and the intracellular signal transduction leading to the regulation of cytosolic calcium and to a migratory tumor cell phenotype. PMID- 12115502 TI - CHC1-L, a candidate gene for prostate carcinogenesis at 13q14.2, is frequently affected by loss of heterozygosity and underexpressed in human prostate cancer. AB - Loss of heterozygosity (LOH) at chromosome 13q14 is one of the most recurrent anomalies observed in sporadic prostate tumors. This LOH is believed to unmask recessive mutations that inactivate a tumor-suppressor gene(s) which otherwise regulates normal cell growth and suppresses abnormal cell proliferation. Identification of potential tumor-suppressor genes within the deleted region is a way of indicating putative pathways of prostate cancer development and progression. The main target that disappears or is downregulated as a result of 13q14 loss remains to be identified. Therefore, our first concern was to find a gene located in the 13q14 region whose transcription is reduced. CHC1-L, for chromosome condensation 1-like, is mapped to the smallest common deleted region. CHC1-L expression is significantly reduced in prostate tumors compared to normal prostate tissues (p = 0.0002). In 21 of 36 (58%) primary prostate tumors studied, CHC1-L expression was reduced at least 2-fold, as measured by real-time quantitative RT-PCR; 18 of the tumors (50%) showed 13q14 LOH for at least 1 of the 5 polymorphic markers that we studied in the region, and 14 (78%) of these were among the tumors underexpressing CHC1-L. CHC1-L is alternatively spliced at its 5' end to produce 2 isoforms, of 551 and 526 aa. Analyses of CHC1-L integrity and of the quantitative expression of its variants indicate that the observed underexpression in prostate tumors is related to reduced expression of the 551 aa isoform. Although CHC1-L is not the obvious candidate given its only known homology, to RCC1, a guanine nucleotide exchange factor for the Ras-related GTPase Ran, the frequent significant decrease observed in its expression in prostate cancer associated with the difference in frequency of CHC1-L variant isoforms between normal and neoplastic prostate tissues places it in a pivotal role or possibly adjacent to a gene that has that role in prostate cancer evolution. PMID- 12115501 TI - Wnt/beta-catenin signaling suppresses apoptosis in low serum medium and induces morphologic change in rodent fibroblasts. AB - Wnt/beta-catenin signaling plays important roles in tumorigenesis in certain tumors as well as during development. However, the mechanism of tumorigenesis mediated by this signaling remains to be elucidated. We investigated the response of rodent fibroblasts to activation of Wnt/beta-catenin signaling by treatment with conditioned medium containing soluble Wnt-3a protein (W3a-CM) and by expression of a constitutive active beta-catenin gene harbored by an adenovirus vector. W3a-CM induced transcriptional activation of a beta-catenin/T-cell factor (Tcf)-responsive promoter in rodent fibroblasts such as NIH3T3, Rat-1, Swiss3T3 and Balb3T3 cells. In these cells, an increase in saturation density and an inhibition of apoptosis and/or promotion of growth in low-serum medium were induced by treatment with W3a-CM. In Rat-1 cells, morphologic changes were also induced. All these alterations were reversible. Moreover, the inhibition of apoptosis of NIH3T3 cells in low-serum medium and the morphologic changes in Rat 1 cells, but not the increase in saturation density, were also induced by ectopic expression of a constitutive active beta-catenin gene. These results suggested that activation of Wnt/beta-catenin signaling induces inhibition of apoptosis and morphologic changes in these cells. PMID- 12115503 TI - MSI in endometrial carcinoma: absence of MLH1 promoter methylation is associated with increased familial risk for cancers. AB - Loss of DNA mismatch repair occurs in a variety of malignancies and is associated with genome-wide instability of microsatellite repeats, a molecular phenotype referred to as microsatellite instability (MSI). MSI is a consistent feature of colorectal and endometrial tumors from patients with hereditary non-polyposis colorectal cancer (HNPCC). Sporadic colorectal and endometrial cancers that exhibit MSI frequently have methylation of the MLH1 promoter. We undertook a detailed family and medical history study to compare family cancer risk for women with MSI-positive and -negative endometrial cancers. The MLH1 promoter methylation status was determined for all cancers. Family histories were developed for 80 probands (40 with MSI-positive and 40 with MSI-negative tumors). The numbers of reported cancers in first- and second-degree relatives of the 2 groups were similar. There was a modest increase in familial cancer clustering for MSI-positive probands. When MSI-positive tumors were subclassified according to MLH1 promoter methylation, a clear association between methylation status and familial cancer risk was evident. Women with MSI-positive endometrial cancers in which the MLH1 promoter was unmethylated had a 7-fold relative risk (RR) of demonstrating familial clustering of cancers [RR 7.07 (95% confidence interval 2.29-21.81)]. The women with MSI-positive, MLH1-unmethylated tumors were significantly younger than the rest of the study population (56.1 years vs. 65.4, p < or = 0.01). Age of onset and tumor MSI not associated with MLH1 promoter methylation may point to women with a genetic susceptibility to malignancies. PMID- 12115504 TI - CD40 ligation downregulates EBNA-2 and LMP-1 expression in EBV-transformed lymphoblastoid cell lines. AB - Epstein-Barr virus (EBV) drives the proliferation of human B cells in vitro and during primary infection in vivo. The transformed immunoblasts express nuclear proteins EBNA1-6, transcribed from the Cp/Wp promoter, and the membrane proteins LMP-1, -2A and -2B (lymphoblastoid type of latency). EBV persists through life in resting memory B cells with a restricted type of latency in the absence of the Cp/Wp promoter activity. Since CD40 crosslinking can reportedly inhibit the growth of EBV-transformed lymphoblastoid cell lines (LCLs), we have examined the effect of CD40 ligation on the expression of EBNAs and LMP-1 and on Cp EBV promoter activity together with several phenotypic markers. CD40 crosslinking led to a partial downregulation of EBNA-2, EBNA3-6 and LMP-1 in LCLs, paralleled by downregulation of Cp promoter activity. It also induced upregulation of the germinal center marker CD77 on the LCL cells. Our findings suggest that the encounter of proliferating EBV-transformed immunoblasts with CD40L, as would occur when normal B cells generate memory cells in germinal centers, may switch the viral transcription program from the full lymphoblastoid to a more restricted latency program in a proportion of cells. This would permit virus persistence in the B-cell memory compartment. PMID- 12115505 TI - Activation of Kupffer cells inhibits tumor growth in a murine model system. AB - Kupffer cells, a liver organ-specific macrophage, play an important role in preventing the development of malignant tumors. The mechanism responsible for their tumoricidal activities is not completely known. In our study, we established in vivo models involving a rat malignant cell line, rat Kupffer cells and tumor implantation in nude mice. A series of relevant in vitro experiments were also carried out to determine possible pathways. LPS-activated Kupffer cells produced significant amounts of NO, TNFalpha and IFNgamma. Malignant cells treated with either Kupffer cells or culture supernatant of the activated Kupffer cells had an increase in caspase-8 activity. Implanted tumors originated from malignant cells treated with either Kupffer cells or culture supernatant of the activated Kupffer cells grew much smaller than those from malignant cells without treatment or treated with control supernatants. The alteration of anti-apoptotic Bcl-2 was inversely associated with the change of pro-apoptotic caspase-8 and their levels in the tumor tissues matched the size of the tumors and treatments they received. It appeared that the above changes resulted in an increase in cellular DNA damage and apoptosis seen in malignant cells. Therefore, Kupffer cells execute their anti-tumor effect via increasing the production of NO, TNFalpha and IFNgamma and these cytotoxic molecules inhibit the growth of tumor by damaging cellular DNA and inducing apoptosis that was featured by downregulation of Bcl-2 but upregulation of caspase-8. PMID- 12115506 TI - Expression of UPA and UPAR is associated with the clinical course of urinary bladder neoplasms. AB - The expression of urokinase plasminogen activator (uPA) and its receptor (uPAR) mRNA was determined in 194 subjects with newly detected bladder neoplasms, selected from a larger population-based series. An association was found between uPA and uPAR expression (n = 172; Spearman r(s) = 0.60, p < 0.001). Both uPA and uPAR mRNA levels were higher in muscle invasive (T2+) tumors than in noninvasive mucosal tumors (Ta) or those invading submucosa (T1). The relative hazard ratios (RHRs) for cancer-specific death associated with elevated expression (95% CI), adjusted for age and gender in a Cox proportional hazard model, were 1.8 (1.0 3.3) for uPA (upper quartile cut-line), 2.2 (1.3-4.0) for uPAR (median quartile cut-line) and 2.5 (1.3-4.9) for uPA + uPAR. An RHR for metastatic disease of 4.0 (1.6-9.9) was observed for uPAR. Restricting the analyses to T2+ tumors, the corresponding figures were: 2.1 (1.1-3.9) for uPA, 1.6 (0.8-3.3) for uPAR and 2.5 (1.1-5.6) for both. We conclude that expression of uPA and uPAR is associated with the clinical behaviour of bladder neoplasms, possibly providing means for refined staging of muscle invasive tumors and target proteins for novel therapies. PMID- 12115507 TI - A case-control study of stomach cancer in Mumbai, India. AB - Stomach cancer incidence rates are much lower in India than elsewhere, but the stomach remains one of the 10 leading sites of cancer in both sexes in most of the metropolitan registries. This is an unmatched case-control study of stomach cancer carried out at Tata Memorial Hospital (TMH), Mumbai. Our purpose was to identify the association of tobacco and alcohol use, occupational hazards, diet, consumption of beverages like tea and coffee, the living environment, cooking media and literacy with stomach cancer. Our study included 170 stomach cancer cases and 2,184 hospital controls interviewed during the period 1988-1992. Tobacco chewing, bidi or cigarette smoking and alcohol drinking did not emerge as high risk factors for stomach cancer. Consumption of dry fish at least once a week compared to never or once a every 2 weeks showed a 12-fold excess risk (OR = 12.4, 95% CI 7.0-22.1, p < 0.0001) for stomach cancer among the nonvegetarian food items considered. A protective effect of tea consumption (OR = 0.4, 95% CI 0.2-0.9, p = 0.03), showing 59% reduction in risk, was identified, which could be of use for possible control and prevention of this cancer. PMID- 12115508 TI - Effect of type of screening laboratory on population-based occurrence of cervical lesions in Finland. AB - The incidence of cervical cancer decreased in Finland over a 30-year period because of an effective screening program, but in the beginning of the 1990s it began to increase. Reasons for such an increase are variable: changes in sexual habits, shortcomings in attendance for screening and possibly variation in laboratory quality. We evaluated the impact of 3 laboratories in the greater Helsinki area on screening performance and on the incidence of invasive cervical cancer and preinvasive cervical lesions in the target population. We studied time trends, geographic differences in attendance and detection rates from screening and the incidence of invasive cancer in the greater Helsinki area (population about 1 million) during the 1990s, when screening was reorganized from the Cancer Society of Finland laboratory to the municipal one (Helsinki) and to a private laboratory (Espoo), while in Vantaa screening remained with the same Cancer Society laboratory. The attendance rate for screening increased during the study period in all 3 cities. The numbers of cytologically suspected and histologically confirmed precancerous lesions found, including severe lesions, decreased significantly with the change of laboratory in Espoo; but in Helsinki and Vantaa, they increased. The overall incidence of invasive cervical cancer increased in all cities in the age groups screened but mostly in Espoo. The rather rapid changes and variation in trends in the number of screening findings cannot be explained by changes in etiologic factors or attendance. They may be related more to the quality of the laboratory performance and perhaps to the criteria used in cytology and colposcopy. A well-organized auditing system is proposed to maintain high quality in screening. PMID- 12115509 TI - Grade of malignancy of cervical cancer in regions of Uganda with varying malarial endemicity. AB - As in a prior study on malignant lymphomas, 3 and 6 areas of Uganda showing low and high malarial endemicity, respectively, were selected for analysis and the data retrieved from the Kampala Cancer Registry, which in the 1960s and 1970s collected cases of cancer through a widely used free biopsy service from the whole country. Overall incidence rates were derived from 924 cases from the 12 year period 1964-1975. For reasons of economy, grade of tumour was determined only in cases pertaining to the 6-year period 1968-1973. Of 457 cases, 304 could be reviewed histologically. Only the group of squamous cell carcinomas (84.9%, 258 cases) was large enough for subsequent geographic analysis. High incidence rates of CC were found in areas with high malarial endemicity, whereas low incidence rates occurred where malaria was either frequent or rare. A correlate to malarial infection was the proportion of high-grade carcinomas irrespective of the overall incidence of CC. With high prevalence of malaria and high CPRs of 35 74%, the relative share of high-grade cancer amounted to 50-67%. Where malaria was rare with low CPRs of 8-11%, these values were lower and varied only from 25 39% with a similar range of 14%. Geographic agreement between malarial endemicity and the PI of high-grade cancer was high in the 9 study areas and only slightly lower than for BL, for which the association with malaria is beyond doubt. Compared to areas with little malaria, the RR for the incidence of high-grade carcinomas in areas with severe malaria was increased. The value was 2.04 with a 95% confidence interval of 1.37-3.04. Attributable to secondary immunodeficiency, lifelong exposure to malaria may result in excess frequency of high-grade malignant tumours not only in the group of malignant lymphomas but also in CC. PMID- 12115510 TI - Risk of pancreatic cancer in relation to alcohol drinking, coffee consumption and medical history: findings from the Japan collaborative cohort study for evaluation of cancer risk. AB - We evaluated the associations of such lifestyle factors as alcohol drinking, coffee consumption and medical history with risk of death from pancreatic cancer in a large-scale prospective cohort study [the Japan Collaborative Cohort Study for Evaluation of Cancer Risk (JACC study)] in Japan. Subjects were 110,792 (46,465 men and 64,327 women) inhabitants who were enrolled from 45 areas throughout Japan. At baseline, a self-administered questionnaire was used to obtain information on lifestyle factors and medical history. Cox proportional hazard models were used to calculate relative risks. During the follow-up period (mean +/- SD 8.1 +/- 1.8 years), 225 deaths due to pancreatic cancer were identified. Overall, neither alcohol nor coffee intake was associated with risk of death from pancreatic cancer. Heavy coffee consumption (> or =4 cups/day), however, may increase the risk. Men who reported a history of diabetes mellitus and women who reported a history of gallstone/cholecystitis were at significantly (2-fold) increased risk of death from pancreatic cancer. PMID- 12115512 TI - Improved survival in both histologic types of oesophageal cancer in Sweden. AB - The prognosis among patients diagnosed with oesophageal cancer is poor with an overall 5-year survival close to 5% in most countries. Improved diagnostic and surgical strategies might influence the survival, however. We investigated the observed and relative survival among all patients in Sweden diagnosed with oesophageal adenocarcinoma (n = 1,441) or squamous cell carcinoma (n = 6395) from 1961-1996 with follow-up to December 1997. Observed survival rates were calculated by the life-table method. Relative survival rates were computed as the ratio of the observed to the expected survival. The expected survival was inferred from the survival among the entire Swedish population in the same age, sex and calendar year strata. The 5-year observed survival rate for adenocarcinoma increased from a stable figure close to 4% during the entire period 1961-1989 to 10.5% during 1990-1996. Similarly, the 5-year relative survival rate was stable around 5% during 1961-1989, but during 1990-1996 the survival was increased to 13.7%. For squamous cell carcinoma, the survival improved slightly by each decade, starting with 3.8% 5-year observed survival in 1961-1969 to 7.0% during 1990-1996. Similarly, the 5-year relative survival improved from 5.0% to 8.9% during the study period. In conclusion, the survival rates for both oesophageal adenocarcinoma and squamous cell carcinoma have increased significantly during the 1990s compared to those in the previous 3 decades (p < 0.001). PMID- 12115511 TI - Rapidly rising breast cancer incidence rates among Asian-American women. AB - In recent years, breast cancer incidence rates have fluctuated over relatively short time spans; examination of these patterns can provide etiologic clues and direction for prevention programs. Asian-American women are generally considered to be at lower risk of breast cancer than other ethnic groups. However, their rates are typically based on an aggregation of ethnic Asian populations, which may obscure important ethnic differences in risk. Detailed analyses of the trends in ethnic-specific incidence rates will provide more information than when ethnicities are combined. Los Angeles County, California, the most populous and probably the most ethnically diverse county in the United States, has a large multi-ethnic Asian-American population. Trends in invasive female breast cancer incidence were examined using data from the Los Angeles Cancer Surveillance Program, the population-based cancer registry covering the County. Although overall breast cancer incidence rates remained stable in the late 1980s and early 1990s, data for the most recent 5-year period suggest that incidence may again be increasing for Asian-American and non-Hispanic white women over age 50 (estimated annual percent change = 6.3%, p < 0.05 and 1.5%, p < 0.05, respectively), although little change has occurred among black and Hispanic women. Invasive breast cancer incidence rates for Asian-American ethnic groups are heterogeneous and, for most, are increasing. In Los Angeles County, rates for Japanese-American women have increased rapidly since 1988 and are now approaching rates for non Hispanic white women. Rates among Filipinas, who have historically had higher rates than their other Asian-American counterparts, are not increasing as rapidly as rates for Japanese women, but remain relatively high. Breast cancer risk among women of Japanese and Filipino ancestry is twice that of Chinese and Korean women. Asian women, who commonly have low breast cancer rates in their native countries, typically experience increasing breast cancer incidence after immigrating to the United States. Ethnic-specific incidence rates show that Japanese-Americans, the first Asian population to immigrate to Los Angeles County in large numbers and the most acculturated, have experienced a rapid increase in breast cancer incidence. Japanese-American rates in Los Angeles County may have already surpassed those of non-Hispanic whites if recent trends have continued unabated. PMID- 12115513 TI - NS-398, a selective cyclooxygenase 2 inhibitor, inhibited cell growth and induced cell cycle arrest in human hepatocellular carcinoma cell lines. AB - Cyclooxygenase 2 (COX-2) has been suggested to be associated with liver carcinogenesis. Several reports have shown that NSAIDs inhibit the growth of hepatocellular carcinoma cell lines. There is little evidence of how COX-2 inhibitors regulate the proliferation of hepatocellular carcinoma cells or the mechanism involved. In our study, we investigated the growth-inhibitory mechanism of a selective COX-2 inhibitor, NS-398, in 4 hepatocellular carcinoma cell lines by studying cell growth, COX-2 and proliferating cell nuclear antigen (PCNA) expression, cell cycle distribution and the evidence of apoptosis. NS-398 inhibited the growth of all 4 cell lines in a time- and dose-dependent manner and the inhibitory effects were independent of the level of COX-2 protein expression. PCNA expression was downregulated by NS-398 in a dose-independent manner. NS-398 caused cell cycle arrest in the S phase with a reduction in cell numbers and cell accumulation in the G0/G1 phase, for all 4 cell lines. No evidence of apoptosis was observed in our present study. Our findings suggest that a selective COX-2 inhibitor might serve as an effective tool for the chemoprevention and treatment of hepatocellular carcinomas. A reduction in cell number in the S phase may be an important event in cell cycle arrest caused by NS-398 in hepatocellular carcinoma cell lines. PMID- 12115514 TI - Post-menopausal hormonal therapy and gallbladder cancer risk. AB - The relation between post-menopausal hormone replacement therapy (HRT) and gallbladder cancer was analyzed in women above age 45 years, using data of a case control study conducted in Italy between 1985 and 1997, on 31 incident, histologically confirmed cases and 3,702 controls in hospital for acute, non neoplastic conditions. The multivariate odds ratio (OR) was 3.2 (95% confidence interval: 1.1-9.3) for those who had ever used HRT and the OR tended to rise with longer duration. Although based on small numbers, due to the rarity of the disease, these findings provide the first direct epidemiological evidence of an association between HRT and gallbladder cancer. PMID- 12115515 TI - Unexpected sensitivity to radiation of fibroblasts from unaffected parents of children with hereditary retinoblastoma. AB - The response to ionizing radiation was examined in diploid skin fibroblasts derived from 5 patients with hereditary type retinoblastoma as well as their parents. Unexpected sensitivity to cell killing, as measured by clonogenic survival, as well as enhanced radiation-induced G(1) arrest were observed in at least 1 parental fibroblast strain in all 5 families. In all cases, parental strains were equally or more radiosensitive than the probands. The mutation of the retinoblastoma gene (RB) determined in 4 of 5 probands was either absent from the parental cells, as expected from the negative family histories, or identical, in 1 father who was a known carrier. In the fifth family, the family history was negative for retinoblastoma. We hypothesize that the increased parental cell sensitivity to radiation suggests the presence of an as yet unrecognized genetic event occurring in 1 or both parents of children with retinoblastoma. Whether it increases mutability of the RB locus or other loci or interacts with RB is conjectural. PMID- 12115518 TI - Increased incidence rate of colorectal tumors due to the intake of a soluble dietary fiber in rat chemical carcinogenesis can be suppressed by substituting partially an insoluble dietary fiber for the soluble one. AB - In epidemiologic studies on human colorectal tumors, results on the relative protective effect of soluble and insoluble fibers are not consistent. We studied in this work the effect in rats of feeding guar gum or guar gum together with cellulose on the incidence of colorectal tumors induced by 1,2-dimethylhydrazine. The results were as follows: (i) The enhancement of tumor formation by feeding solely guar gum (guar gum group) was suppressed completely when two-thirds of the guar gum was replaced with cellulose (cellulose-guar gum group). The odds ratio for tumor formation was 0.075 (95% CI 0.006-0.936, p = 0.044) for guar gum group vs. no fiber control and 0.833 (0.134-5.167, p = 0.83) for cellulose-guar gum group vs. the control. (ii) In both groups, serum cholesterol and triglyceride levels decreased significantly compared to the no fiber control group, and fecal excretion of total bile acids almost doubled. (iii) In guar gum group rats, the deconjugation activity (beta-glucuronidase, beta-glucosidase) was higher than the control or cellulose-guar gum group rats. (iv) The amount of cecal short-chain fatty acids was almost double in guar gum group rats compared to the cellulose guar gum group or the control rats, and pH of the cecal content of the guar gum group rats had a tendency to be lower. (v) The concentration of fecal secondary bile acids was extremely low in the younger rats of the guar gum group. From these results, it seemed significant to study the cancer preventive effect of the mixed feeding to experimental animals of water-soluble and insoluble fibers instead of the singular feeding. PMID- 12115519 TI - Metabolism and DNA binding of the environmental pollutant 6-nitrochrysene in primary culture of human breast cells and in cultured MCF-10A, MCF-7 and MDA-MB 435s cell lines. AB - The environmental pollutant 6-nitrochrysene (6-NC) is a potent mammary carcinogen in the rat. To determine if the results obtained in 6-NC-treated rodents can be applicable to humans, we examined its metabolic activation in primary cultures of human breast cells prepared from tissues obtained from reduction mammoplasty from 3 women and in a cultured, immortalized human mammary epithelial cell line (MCF 10A), as well as estrogen-dependent (MCF-7) and estrogen-independent (MDA-MB 435s) human breast cancer cell lines. Metabolites identified following 24 hr incubations of [(3)H]6-NC (2.5, 5.0 and 10 microM) with human breast cells were derived from ring-oxidation (trans-1,2-dihydroxy-1,2-dihydro-6-nitrochrysene [1,2 DHD-6-NC]) and nitro-reduction (6-aminochrysene [6-AC]); chrysene-5,6-quinone (C 5,6-Q) was also detected. Levels of metabolites (pmol/mg protein) varied greatly depending on the concentration of 6-NC and the individual breast tissue used; 1,2 DHD-6-NC, ranged from not detected to 15.6 +/- 1.0; 6-AC, from 11.5 +/- 4.0 to 155 +/- 10.2; C-5,6-Q, from 18.3 +/- 10.8 to 196.7 +/- 15.4. Qualitatively similar metabolic profiles were obtained upon incubation of [(3)H]6-NC with MCF 10A, MCF-7 and MDA-MB-435s. We also detected 1,2-dihydroxy-6-aminochrysene (1,2 DH-6-AC; ranged from not detected to 50.4 +/- 9.8). Some of the metabolites identified in our study are known to be proximate carcinogenic forms of 6-NC in rodents. MCF-7 was the most efficient cell line in catalyzing 6-NC to genotoxic metabolites, and we demonstrated that the major DNA adduct is chromatographically identical to that found in the mammary gland of rats treated by gavage with 6-NC and that obtained from the incubation of [(3)H]1,2-DHD-6-NC with MCF7 cells or from nitro-reduction of 1,2-DHD-6-NC in the presence of 2'-deoxyguanosine 5' monophosphate in vitro. This is the first report to demonstrate the ability of human breast cells, MCF-10A, and breast cancer cell lines to activate 6-NC to metabolites that can damage DNA. PMID- 12115517 TI - Ipriflavone inhibits osteolytic bone metastasis of human breast cancer cells in a nude mouse model. AB - Osteolytic bone metastasis is a frequent problem in the treatment of cancer. Ipriflavone, a synthetic isoflavone that inhibits osteoclastic bone resorption, has been used for the treatment of osteoporosis in some countries. Some other isoflavones also exhibit an antitumor effect in vitro and in vivo. Here, we studied the effects of ipriflavone on osteolytic bone metastasis of MDA-231 human breast cancer cells injected intracardially into athymic nude mice (ICR-nu/nu). Daily oral administration of ipriflavone at 12 mg/mouse significantly inhibited the development of new osteolytic bone metastases (p < 0.05) and the progression of established osteolytic lesions (p = 0.01), prolonging the life of tumor bearing mice (p = 0.01 vs. control). In addition, ipriflavone reduced the number of osteoclasts at the bone-cancer interface with no severe adverse effects on the host. In vitro, ipriflavone inhibited the proliferation and DNA synthesis of MDA 231 cells and blocked the ligand-induced phosphorylation of Tyr(845) of the EGFR. Ipriflavone did not promote apoptosis of MDA-231 cells. Our results show that ipriflavone not only directly inhibits the growth of cancer cells but also reduces osteoclasts to prevent the soft tissue tumor burden and osteolytic bone metastases. These findings raise the possibility that ipriflavone may be of use as a therapeutic agent against osteolytic bone metastasis. PMID- 12115520 TI - Acceleration of invasive activity via matrix metalloproteinases by transfection of the estrogen receptor-alpha gene in endometrial carcinoma cells. AB - It is well known that the functions of reproductive organs are regulated by sex steroids and their receptors and it is hypothesized that the progression of neoplasms that originate from the reproductive organs is influenced by them. However, the correlation between sex steroids and tumor progression, especially tumor invasion, is not well known in endometrial carcinoma. In our study, we focused on the influence of estrogen and its receptor in invasion and matrix metalloproteinases (MMPs), which are known to be important in tumor invasion, as well as on endometrial carcinoma cells. The growth of Ishikawa cells, to which an estrogen receptor-alpha expressing vector was transfected, was accelerated by 17 beta-estradiol as was the acceleration of the expression of cyclin D1. By invasion assay, in conditions with 17 beta-estradiol, the invasiveness of Ishikawa cells was enhanced. Furthermore, according to the accelerated invasiveness, the expression of MMP-1, -7 and -9 and Ets-1 was enhanced. These results suggest that activation of ER-alpha by estrogen results in tumor progression by stimulating cell growth and invasiveness via acceleration of the expression of MMPs. PMID- 12115522 TI - HRAS1 variable number of tandem repeats polymorphism and risk of bladder cancer. AB - The HRAS1 variable number of tandem repeats (VNTR) polymorphism, 1 kb downstream from the HRAS1 gene, has been reported to be associated with risk of various cancers. To examine whether individuals with rare HRAS1 VNTR alleles are at increased risk of bladder cancer we carried out a case control study with 230 bladder cancer cases and 203 hospital-based controls frequency-matched on ethnicity, gender and age. For genotyping we used a PCR-based long-gel electrophoretic assay that provides precise allele size discrimination. We did not find evidence of a strong overall effect of the HRAS1 VNTR on bladder cancer risk. Genotype data for whites and blacks were analyzed separately, but the number of black subjects was too small to estimate meaningful odds ratios. Compared to white subjects with 2 common alleles, the odds ratio (OR) for white subjects with 1 rare allele was 0.9 (95% confidence interval (CI) = 0.5-1.4) and for those with 2 rare alleles OR = 1.7 (95% CI = 0.6-5.4). HRAS1 genotype may be related to the prognosis of bladder cancer, however, because incident cases, i.e., newly diagnosed cases had a higher frequency of rare alleles than did prevalent cases, i.e., cases already existing at the time of recruitment. Repeating the analyses with incident cases only (n = 53), the OR for subjects with 1 rare allele was 1.2 (95% CI = 0.6-2.4) and for those with 2 rare alleles 3.2 (95% CI = 0.8-13.7). The number of incident cases was too small to draw firm conclusions on a possible association with a subgroup of tumors with a poor prognosis. Published 2002 Wiley-Liss, Inc. PMID- 12115521 TI - Spontaneously formed tumorigenic hybrids of Meth A sarcoma and macrophages grow faster and are better vascularized than the parental tumor. AB - Macrophages and Meth A sarcoma cells spontaneously fuse and give rise to tumorigenic hybrid cell lines with a mixed phenotype. We report here that the hybrid tumors grow faster and have a strikingly better developed vasculature than the parent sarcoma. Thus, electron microscopy and immunohistochemical analysis revealed that in the most active areas of neovascularization, the tumors that emerged from inocula of monoclonal hybrid cell populations had a microvessel density nearly twice that of Meth A tumors after 1 week of growth. Moreover, the proportion of vessels associated with pericytes, detected by staining for smooth muscle alpha-actin, was 3 times higher in the hybrid tumors, attesting to the more advanced differentiation of their vasculature. The collagenous stroma component was also more extensive in the hybrid tumors. Concentration of the angiogenic proteins vascular endothelial growth factor (VEGF) and transforming growth factor-beta (TGF-beta) were significantly higher in supernatants of hybrid cell cultures compared with Meth A cultures. These observations indicate that the growth advantage of the hybrid tumors over the parental sarcoma is due to a higher angiogenic capacity. Because the malignant features of many tumors correlate with angiogenesis and because macrophages are known to be major producers of angiogenic factors, our data open the possibility that the intense neovascularization of highly aggressive cancers in some cases reflects the acquisition of macrophage traits by heterotypic cell fusion. PMID- 12115523 TI - Mucin phenotype and microsatellite instability in early multiple gastric cancers. AB - Clinicopathologically, multiple gastric cancers (MGCs) are reported to involve predominantly intestinal-type adenocarcinoma and frequently to be associated with severe intestinal metaplasia. However, there are few reports concerning the characteristic biomarkers of early MGCs. The aim of our study was to identify the cellular lineage defined by mucin phenotypes and the relationships among mucin phenotypes, background mucosa and microsatellite instability (MSI) of early MGCs. We examined mucin phenotypes of 63 surgically resected carcinomas from 25 patients with early MGCs and 39 early solitary gastric cancers (SGCs) by immunohistochemical analysis using a panel of monoclonal antibodies. MSI and the degree of intestinal metaplasia (IM) on the background mucosa were also examined. In early MGCs, the incidence of cancer exhibiting the gastric phenotype (G-type) was 59% (37 of 63 cancers), which was higher than that in early SGCs (23%, 9 of 39 cancers). There was a significant difference between the distributions of mucin phenotypes in early MGCs and early SGCs (p = 0.001). Whereas half of the G type cancers in early MGCs were related to severe IM, none of the G-type cancers in early SGCs were related to severe IM. In the early MGCs, MSI was observed in 21 of 63 cancers (33.3%). In contrast, MSI was observed in only 3 of the 39 (7.7%) early SGCs, indicating a significant difference between these 2 groups (p < 0.01). Our results suggest that the characteristic features of early MGCs are the gastric mucin dominant phenotype and high frequency of MSI. PMID- 12115524 TI - Effects of cytochrome P450 (CYP) 2A6 gene deletion and CYP2E1 genotypes on gastric adenocarcinoma. AB - Cytochrome P450 (CYP) 2A6 and CYP2E1 are enzymes with a high ability to activate a nitrosamine, 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), to its potent and ultimate carcinogens. The polymorphic CYP2A6 and CYP2E1 have been implicated in increased susceptibility to certain malignancies. In our study, 120 Japanese patients with gastric adenocarcinoma and 158 healthy controls were compared for frequencies of CYP2A6 and CYP2E1 genotypes. The frequency with which the subjects carried homozygotes of the CYP2A6 gene deletion allele, which causes lack of the enzyme activity, was significantly higher in the gastric cancer patients than in the healthy control subjects (OR = 3.14, 95% confidence interval (95% CI) = 1.05-9.41). Subdividing gastric adenocarcinoma according to tumor differentiation, patients with the well-differentiated type were 4.9-fold more likely to have the CYP2A6 homozygote deletion genotype (OR = 4.91, 95% CI 1.17 20.52). Stratifying by smoking status, we did not find the risk of CYP2A6 gene deletion allele in gastric adenocarcinoma. The CYP2E1 polymorphism detected by RsaI was not significantly different between gastric adenocarcinoma patients (40.8%) and the control population (44.3%). No statistically significant changes were observed when the CYP2E1 genotype was examined relative to tumor differentiation and smoking status. These results suggest that the CTY2A6 deletion is associated with gastric adenocarcinoma among Japanese populations. PMID- 12115525 TI - Expression of N-terminally truncated isoforms of CDP/CUX is increased in human uterine leiomyomas. AB - Genetic analyses and mRNA expression studies have implicated CUTL1 as a candidate tumor-suppressor gene in uterine leiomyomas and breast cancers. However, modulation of CDP/Cux, the protein encoded by CUTL1, does not agree with this notion. The activity of CDP/Cux, which is the DNA binding subunit of HiNF-D, was upregulated as normal cells progressed into S phase and constitutively elevated in several tumor cell lines. Activation of CDP/Cux at the G(1)/S transition involved the proteolytic processing of the protein to generate a shorter isoform. Uterine leiomyomas represent a unique reagent for molecular analysis because they are resected as homogeneous tumor tissue together with the adjacent normal myometrium and they are often very large. In the present study, proteins were isolated from 16 pairs of matched tumors and adjacent myometrium and analyzed by Western blot and electrophoretic mobility shift assays. Strikingly, in 11/16 tumors, the steady-state level of small CDP/Cux isoforms was increased compared to normal control tissue. Where tested, a corresponding increase in CDP/Cux stable DNA binding activity was observed. DNA sequencing analysis of CUTL1 cDNAs from 6 leiomyomas, including 4 with LOH of CUTL1, did not reveal any gross rearrangement or point mutations. Altogether these findings suggest that CUTL1 is probably not the tumor suppressor on 7q22. Moreover, the frequent increase in smaller CDP/Cux isoforms indicates that molecular events associated with the truncation of CDP/Cux proteins may be selected in uterine leiomyomas. PMID- 12115526 TI - SH2D1A expression in Burkitt lymphoma cells is restricted to EBV positive group I lines and is downregulated in parallel with immunoblastic transformation. AB - The SH2 domain containing SH2D1A protein has been characterized in relation to the X-linked lymphoproliferative disease (XLP), a primary immunodeficiency that leads to serious clinical conditions after Epstein-Barr virus (EBV) infection. The SH2D1A gene is mutated in the majority of XLP patients. We previously detected SH2D1A in activated T and NK cells, but not in B lymphocytes. We have found SH2D1A protein in Burkitt lymphoma (BL) lines, but only in those that carried EBV and had a Group I (germinal center) phenotype. All the EBV-carrying Group III (immunoblastic) and the EBV-negative BL lines tested were SH2D1A negative. Motivated by these differences, we studied the impact of EBV and the cellular phenotype on SH2D1A expression. We approached the former question with BL sublines after both the loss of the virus and subsequent reinfection. We also tested original EBV-negative BL lines carrying transfected EBV genes, such as EBNA1, EBNA2, EBNA6, EBER1, 2 and LMP1, respectively. In our experiments, no direct relationship could be seen between EBV and SH2D1A expression. We modified the phenotype of the Group I BL cells by LMP1 transfection or CD40 ligation. The phenotypic changes, indicated by expression of immunoblastic markers, e.g., SLAM, were accompanied by downregulation of SH2D1A. It seems, therefore, that the presence of EBV and the phenotype of the cell together regulate SH2D1A expression in the BL cells. It is possible that SH2D1A is expressed in a narrow window of B cell development represented by germinal center cells. PMID- 12115527 TI - Renal cell carcinoma-associated antigen G250 encodes a naturally processed epitope presented by human leukocyte antigen-DR molecules to CD4(+) T lymphocytes. AB - We previously identified an HLA-A2.1-restricted epitope within the RCC-associated antigen G250 that is recognized by CTLs. Using DCs of healthy individuals, which were loaded with overlapping 20 mer G250-derived peptides, we here report the induction of CD4(+) T cells that recognize the G250 peptide of amino acids 249 268. Moreover, naturally processed G250 protein is readily recognized by these G250-specific CD4(+) T cells in the context of HLA-DR molecules. Interestingly, peptide G250:249-268 overlaps the previously identified HLA-A2.1-restricted G250 epitope recognized by CTLs. These data and the high prevalence of G250 in RCC patients make peptide G250:249-268 a potential target in peptide-based vaccines to induce both CD4(+) and CD8(+) T-cell responses in patients. PMID- 12115528 TI - IFN-gammaupregulates apoptosis-related molecules and enhances Fas-mediated apoptosis in human cholangiocarcinoma. AB - Human cholangiocarcinoma is a malignancy with no effective therapy and a poor prognosis. Previously, we demonstrated that cultured human cholangiocarcinoma cell lines heterogeneously express Fas on their surface, resulting in 2 subpopulations, Fas-high and Fas-low cells. Fas-low cells are resistant to apoptosis induced by Fas antibody and the calmodulin antagonists tamoxifen and trifluoperazine and are tumorigenic in nude mice (Pan et al., Am J Pathol 1999;155:193-203). Here, we show that IFN-gamma enhances apoptosis in both Fas high and Fas-low cells. IFN-gamma upregulates many apoptosis-related molecules, including Fas, caspase-3, caspase-4, caspase-7, caspase-8 and Bak, in both cell lines. Pretreatment with IFN-gamma facilitated Fas-mediated caspase cleavage, cytochrome c release and Bax translocation. The ability of IFN-gamma to inhibit tumorigenesis of Fas-low cells was demonstrated in nude mice. Intratumoral injection of IFN-gamma decreased tumor volumes by 78%. These findings indicate that IFN-gamma modulates the apoptotic pathway by upregulating apoptosis-related genes. This renders tumorigenic Fas-low cholangiocarcinoma cells nontumorigenic and sensitive to Fas apoptosis, thus representing a possible therapeutic modality. PMID- 12115529 TI - Expression of maspin predicts poor prognosis in breast-cancer patients. AB - The tumor suppressor gene maspin has been reported to inhibit the invasiveness and motility of breast cancer cells. It has been reported that maspin is expressed in normal human mammary epithelial cells but is downregulated during cancer progression, and that p53 could induce maspin expression by transcriptional activation. However, to date, the clinical significance of maspin expression and its correlation with p53 protein expression in human breast cancer patients have not been elucidated. One hundred and sixty-eight female patients diagnosed with invasive ductal carcinoma, who had undergone a mastectomy (154 patients) or breast-conserving surgery (14 patients), were followed up for 15-119 months (median: 87 months) postoperatively. Immunoreactivity for maspin and p53 antibodies with paraffin-embedded carcinoma tissue was investigated using labeled streptavidin-biotin methods. Tumors with more than 20% of positive cells were considered positive for the expression of maspin. The expression of maspin in carcinoma cells was found in 27.4% (46 of 168) and significantly correlated with larger tumor size (p = 0.008), higher histologic grade (p = 0.0001) and positive p53 status (p = 0.003). A significant inverse relationship was observed between the expression of maspin and estrogen receptor (p = 0.0004) or progesterone receptor status (p = 0.02). Univariate analysis by log-rank test revealed a significant association between the expression of maspin and shorter relapse-free survival (p < 0.0001) and overall survival (p < 0.0001). According to Cox's multivariate analysis, the expression of maspin had the most significant effect in relapse-free survival (p < 0.0001) and overall survival (p < 0.0001) followed by lymph node status. In turn, the expression of maspin in 58 cases of ductal carcinoma in situ were also investigated to explore whether the downregulation of maspin through cancer progression are true or not. However, there were no positive cases in our series. These results seem to be contrary to previous reports defining maspin as a tumor suppressor gene. Although more precise characterization of the maspin expression, especially gene analysis is essential, the present investigation suggests that the expression of maspin is not downregulated through malignant progression and that the immunohistochemic detection of maspin in carcinoma cells may be helpful for selecting the group of breast cancer patients with an aggressive phenotype. PMID- 12115530 TI - Inhibitory effects of the standardized extract (DA-9601) of Artemisia asiatica Nakai on phorbol ester-induced ornithine decarboxylase activity, papilloma formation, cyclooxygenase-2 expression, inducible nitric oxide synthase expression and nuclear transcription factor kappa B activation in mouse skin. AB - Artemisia asiatica Nakai has been used in traditional Asian medicine for the treatment of inflammatory and other disorders. Previous studies have revealed that the formulated ethanol extract (DA-9601) of A. asiatica has pronounced antioxidative and antiinflammatory activities and exhibits cytoprotective effects against experimentally induced gastrointestinal, hepatic and pancreatic damage. In the present study, we assessed the inhibitory effect of DA-9601 on tumor promotion, which is closely linked to inflammatory tissue damage. As an initial approach to evaluating the possible antitumor-promoting potential of DA-9601, its effects on TPA-induced ear edema were examined in female ICR mice. Pretreatment of the inner surface of the mouse ear with DA-9601 30 min prior to topical application of TPA inhibited ear edema at 5 hr. TPA-stimulated expression of epidermal COX-2 and iNOS was also mitigated by topical application of the same extract. Moreover, DA-9601 abrogated the TPA-mediated activation of NF-kappa B/Rel and AP-1 in mouse epidermis. Suppression of epidermal NF-kappa B by DA-9601 appeared to be mediated in part through inhibition of I kappa B alpha degradation, thereby blocking the nuclear translocation of p65, the functional subunit of NF-kappa B. DA-9601 also significantly suppressed TPA-induced ODC activity and papilloma formation in mouse skin. Taken together, these findings suggest that DA-9601 derived from A. asiatica possesses potential chemopreventive activities. PMID- 12115531 TI - Cytokine serum levels in soft tissue sarcoma patients: correlations with clinico pathological features and prognosis. AB - We investigated the correlations between serum levels of selected proinflammatory, hematopoietic and angiogenic cytokines and soluble cytokine receptors with the clinico-pathological features and prognosis in soft tissue sarcoma patients. Serum levels of 9 cytokines (TNFalpha, IL-1ra, IL-6, IL-8, IL 10, M-CSF, G-CSF, VEGF, bFGF) and 4 free cytokine receptors (sIL-2R alpha, sIL 6R, TNFRI, TNFRII) were measured by means of an enzyme-linked immunoadsorbent assay kit in 156 soft tissue sarcoma patients before the treatment and in 50 healthy controls. Serum levels of 10 cytokines and cytokine receptors were also assayed during patients' follow-up after the treatment. Significantly elevated pretreatment serum levels of 11/13 cytokines and cytokine receptors were found in sarcoma patients, as compared to healthy controls. In 40.4% of patients 6 or more cytokines and cytokine receptors (most frequently: TNF RI, IL-6, IL-8) were elevated in parallel. Serum levels of IL-6, sIL-2R, VEGF, M-CSF and TNF RI correlated significantly with tumor size and serum levels of IL-8 and IL-6 were significantly higher in patients with Grade 2/3 vs. Grade 1 tumors. We did not observe any significant differences in cytokine serum levels between patients with primary and recurrent tumors and patients with and without distant metastases. Using univariate analysis, overall survival (OS) in all patients was affected by tumor size (<5 cm vs. 5-10 cm vs. >10 cm), tumor grade (G1 vs. G2/3), presence of metastases, pretreatment serum levels of 8 cytokines (IL-6, IL-8, IL 10, sIL-2R, TNF RI, TNF RII, M-CSF, VEGF) and the number of cytokines increased (0-1 vs. 2-5 vs. < or = 6). Elevated serum levels of IL-6, IL-8, IL-10 and sIL-2R alpha, high tumor grade and larger tumor size strongly correlated with shorter disease-free survival (DFS). Multivariate analysis identified G2/3 tumor grade (p = 0.001), the presence of metastases (p = 0.004), elevated IL-6 serum level (p = 0.02), elevated IL-8 serum level (p = 0.048) and the number of cytokine serum levels above upper cut-off values (p = 0.01) as the independent prognostic factors related to OS, and G2/3 tumor grade (p = 0.005) and increased IL-6 serum level (p = 0.035) as independent prognostic factors related to DFS. In a group of patients without metastases (M0) higher tumor grade, elevated serum level of IL-6 and TNF RII, and the number of elevated cytokine serum levels correlated independently with poor survival. We found a significant decrease of several cytokine serum levels in patients after treatment (IL-1ra, IL-6, IL-8, IL-10, TNF RII, M-CSF) [p < 0.05]. Persistently elevated serum level of IL-6 after the treatment has also shown negative prognostic significance for OS (univariate analysis). Serum levels of some proinflammatory, hematopoietic and angiogenic cytokines and cytokine receptors are elevated, frequently in parallel, in a large percentage of soft tissue sarcoma patients. Significant correlations of serum cytokine levels with tumor size and grade suggest that some of these cytokines may be directly or indirectly involved in the progression of soft tissue sarcomas. Serum assays of IL-6, IL-8 and TNF RII before or after the treatment may be useful in establishing soft tissue sarcoma patients prognosis. PMID- 12115532 TI - Overexpression of Bcl-2 in squamous cell carcinoma of the larynx: a marker of radioresistance. AB - Squamous cell carcinoma of the larynx can be treated using radiotherapy or surgery, either alone or in combination. Radiotherapy is preferred for early stage tumours, as it spares the larynx and therefore preserves speech and swallowing. Unfortunately, approximately 15% of tumours treated this way will prove to be radioresistant, as manifest by tumour recurrence within the original radiotherapy field over the ensuing 12 months. By causing extensive DNA damage, radiotherapy aims to induce apoptosis and tumour regression. Our hypothesis was that defects in the mechanisms that recognise DNA damage, induce cell cycle arrest or control apoptosis, either alone or in combination, may be responsible for radioresistance. We therefore undertook an immunohistochemic analysis of pretreatment biopsies of radioresistant (n = 8) and radiosensitive (n = 13) laryngeal tumours. To minimise the impact of confounding factors, strict inclusion criteria were observed; all tumours were of the glottic subsite and all recurrences developed within 12 months of radiotherapy at the site of the original tumour. The expression of key proteins involved in DNA damage recognition (p53), cell cycle arrest (ATM, p16 and p21/WAF1) and apoptosis (Bcl-2 and BAX) were studied. Ki-67 was also assessed as a marker of cell proliferation to exclude low mitotic rate as a cause of radioresistance. A statistically significant correlation was observed between overexpression of Bcl-2 and radioresistance (p = 0.003, Fisher's exact test). We hypothesise that overexpression of the anti-apoptotic protein Bcl-2 allows tumour cells with extensive radiation-induced DNA damage to continue proliferating; the absence of an appropriate apoptotic response manifests clinically as radioresistance. PMID- 12115533 TI - A population-based familial aggregation analysis indicates genetic contribution in a majority of renal cell carcinomas. AB - The etiology of RCC is incompletely understood and the inherited genetic contribution uncertain. Although there are rare mendelian forms of RCC stemming from inherited mutations, most cases are thought to be sporadic. We sought to determine the extent of familial aggregation among Icelandic RCC patients in general. Medical and pathologic records for all patients diagnosed with RCC in Iceland between 1955 and 1999 were reviewed. This included a total of 1,078 RCC cases, 660 males and 418 females. With the use of an extensive computerized database containing genealogic information on 630,000 people in Iceland during the past 11 centuries, several analyses were conducted to determine whether the patients were more related to each other than members drawn at random from the population. Patients with RCC were significantly more related to each other than were subjects in matched groups of controls. This relatedness extended beyond the nuclear family. RRs were significantly greater than 1.0 for siblings, parents and cousins of probands. RRs were 2-3 for first-degree relatives and 1.6 for third degree relatives. The risk of RCC is significantly higher for members of the extended family of an affected individual, as well as the nuclear family. Our results indicate that germline mutations are significantly involved in what has been defined as sporadic RCC. PMID- 12115534 TI - Specific immunotherapy against occult cancer metastases. AB - We investigated the efficacy of a preparation containing High Five (H5) insect cells infected with recombinant baculovirus encoding the murine interferon-beta gene (H5BVIFN-beta) against established primary tumors and occult lung metastases. Injection of live or lyophilized H5BVIFN-beta into established subcutaneous tumors of the highly metastatic murine UV-2237m fibrosarcoma or K 1735M2 melanoma in syngeneic mice eradicated both primary tumors and preexisting lung metastases. The therapeutic effects of H5BVIFN-beta were not observed in nude mice and were diminished in syngeneic mice depleted of CD4(+) and CD8(+) T cells. Immunohistochemical staining showed that tumors injected with H5BVIFN-beta were densely infiltrated by CD4(+) and CD8(+) T cells in mice with normal CD4/CD8 complement. These data demonstrate that, unlike most immunologic approaches in which prophylaxis can be achieved but eradication of established tumor is rare, lyophilized preparations of H5BVIFN-beta can serve as a novel immunotherapy against both primary tumors and their occult metastases. PMID- 12115535 TI - ICAM-1 expression and the soluble ICAM-1 level for evaluating the metastatic potential of gastric cancer. AB - ICAM-1 plays an important role in cell-cell and cell-extracellular matrix interactions, especially tumor invasion and cytotoxicity of lymphocytes. In the present study, the relationship between metastasis of gastric cancer and ICAM-1 expression by cancer cells or the serum level of s-ICAM-1 was (s-ICAM-1) was examined. ICAM-1 was detected by immunohistochemic staining in 49.0% of 108 patients with gastric cancer. The ICAM-1 expression rate was higher at a more advanced stage, based on lymph node metastasis, being 46.9% in node-negative and 56.1% in node-positive cases. In patients with liver metastasis, the rate was 90.9%, while it was 43.3% in patients without liver metastasis (p < 0.05). The serum s-ICAM-1 level was 262.1 ng/ml (median 205.5, range 176.0-271.0) in healthy subjects and 391.5 ng/ml (median 317.5, range 148.7-1,768.0) in gastric cancer patients (p < 0.001). The serum s-ICAM-1 level was significantly higher in patients with liver metastasis than in patients without liver metastasis (p < 0.0001). In addition, positive ICAM-1 expression cases had significantly higher s ICAM-1 levels than negative ones, 408.9 +/- 188.4 and 308.1 +/- 88.1 ng/ml, respectively. These results suggested that ICAM-1 was overexpressed in cancer cells and released as s-ICAM-1, which would promote hematogenous metastasis by suppressing local anticancer immunity. PMID- 12115536 TI - The antifungal drug ciclopirox inhibits deoxyhypusine and proline hydroxylation, endothelial cell growth and angiogenesis in vitro. AB - The hypusine biosynthetic steps represent novel targets for intervention in cell proliferation. Hypusine is a rare amino acid, formed posttranslationally in one cellular protein, eIF5A, and is essential for cell proliferation. Deoxyhypusine hydroxylase, the metalloenzyme catalyzing the final step in hypusine biosynthesis, and prolyl 4-hydroxylase, a non-heme iron enzyme critical for collagen processing, can be inhibited by small chelating molecules that target their essential metal atom. We examined the effects of 5 compounds (ciclopirox, deferiprone, deferoxamine, mimosine and 2,2'-dipyridyl) on these protein hydroxylases in HUVECs, on cell proliferation and on angiogenesis using 2 model assays: tube-like vessel formation on Matrigel and the chick aortic arch sprouting assay. These compounds inhibited cellular deoxyhypusine hydroxylase in a concentration-dependent manner, but their efficacy varied widely in the following order: ciclopirox--> deferoxamine-->2,2'-dipyridyl-->deferiprone- >mimosine (IC(50) 5-200 microM). Inhibition of DNA synthesis, following the same order (IC(50) 10-450 microM), correlated with G(1) arrest of the cell cycle. These compounds also inhibited proline hydroxylation and maturation of collagen in HUVECs and caused inhibition of angiogenesis in vitro. Of the compounds tested, ciclopirox was by far the most effective inhibitor of HUVEC proliferation and angiogenesis. The strong antiangiogenic activity of this readily available antifungal drug along with its antiproliferative effects suggests a new potential application for ciclopirox in the treatment of solid tumors. PMID- 12115538 TI - Genetic susceptibility and gastric cancer risk. AB - The aim of the present paper is to review and evaluate, in a comprehensive manner, the most recent published evidence on the contribution of genetic susceptibility to gastric cancer risk in humans. We have identified all studies available in MEDLINE published up to October 2001. Only studies carried out in humans and comparing gastric cancer cases with at least 1 standard control group were included in the analysis. We were able to find 31 articles based on 25 case control studies carried out in Caucasian, Asian and African populations. Most of the studies assess the effect of genes involved in detoxifying pathways (n = 12) and inflammatory responses (n = 7). The most widely studied is the GSTM1 null polymorphism. Only a very few studies have evaluated the risk of gastric cancer associated with genes acting on mucosa protection, oxidative damage and DNA repair. The most consistent results are the increased gastric cancer risk associated with IL1B and NAT1 variants, which may account for up to 48% of attributable risk of gastric cancer. Only polymorphisms at HLA-DQ, TNF and CYP2E genes may confer some protective effect against gastric cancer. The most important limitations that preclude definitive conclusions are (i) the lack of appropriate control of potential sources of bias (only 5 population-based studies have been published so far); (ii) the low number of cases analyzed (14 studies included fewer than 99 cases); and (iii) the low number of studies (n = 3) offering concomitant analysis of genetic susceptibility and exposure to relevant cofactors (Helicobacter pylori infection, diet and smoking). We conclude that the scientific data on the role of genetic factors in gastric cancer risk are promising. The lack of association reported so far should be considered with caution due to significant limitations in study design. Cohort studies taking into account simultaneously the different genetic and environmental factors potentially involved in gastric tumorigenesis are needed to ascertain not only the relative contribution of these factors to tumor development but also the contribution of their putative interactions. PMID- 12115539 TI - Impairment of stimulation by estrogen of insulin-activated nitric oxide synthase in human breast cancer. AB - Nitric oxide (NO) is reported to have several important effects in the control of neoplasm. We have reported before the presence of an insulin-activated constitutive form of membrane-bound nitric oxide synthase (IANOS) in various cells. Since the insulin-induced NO synthesis by IANOS could have important consequences on the pathophysiology of neoplastic cells, the role of estrogen on the activity of IANOS in malignant and nonmalignant breast tissue as well as in erythrocytes in breast cancer patients was determined. It was found that the IANOS activity of nonmalignant breast tissue was maximally stimulated by 4-fold over the basal activity in the presence of physiologic amounts of estrogen (8-32 nM). The enzymic activity was, however, inhibited by estrogen both below and above this range when compared to appropriate controls. In contrast, both the basal IANOS activity and the stimulatory effect of estrogen was markedly impaired in malignant breast tissue and in erythrocytes in these patients. It was also noted that tamoxifen, a widely used nonsteroidal compound in breast cancer, mimicked estrogen both in the stimulation and in the inhibition of IANOS activity in both of the tissues. These results indicated the probable existence of a novel pathway for estrogen effect independent of nuclear receptor for the stimulation of IANOS activity that might have important consequences in breast cancer and suggested that some of the beneficial effects of tamoxifen could be related to its estrogen-mimicking effect on IANOS independent of hormone-responsive elements sequence in the DNA. PMID- 12115540 TI - RPR-115135, a farnesyltransferase inhibitor, increases 5-FU- cytotoxicity in ten human colon cancer cell lines: role of p53. AB - A new non peptidic farnesyltransferase inhibitor, RPR-115135, in combination with 5-FU was studied in 10 human colon cancer cell lines (HCT-116, RKO, DLD-1, Colo 320, LoVo, SW-620, HT-29, HCT-15, Colo-205 and KM-12) carrying several mutations but well characterized for p53 and Ras status. We found that there was a slight tendency (not statistically significant) for the p53 inactivated cells to be less sensitive to 5-FU after 6 days continuous treatment. Simultaneous administration of RPR-115135 and 5-FU, at subtoxic concentrations, resulted in a synergistic enhancement of 5-FU cytotoxicity in the p53 wildtype cells (HCT-116, RKO, DLD-1, Colo-320, LoVo). In the p53 mutated cells (SW-620, HT-29, HCT-15, Colo-205, KM 12) the effect was very complicated. In HCT-15 the combination resulted in antagonism, in KM-12 in antagonism or in synergy (at different concentrations) and in SW-620, HT-29 and Colo-205 cells in synergy but only when 5-FU was administered at high concentrations. Growth inhibition could be accounted for on the basis of a specific cell cycle arrest phenotype (G2-M arrest), as assayed by flow cytometry, only in the p53 functioning cell lines. The combination RPR 115135 + 5-FU increases apoptotic events only in these cell lines. In the mutated cell lines no major alterations on cell cycle arrest phenotype and no induction of apoptosis was observed. Although RPR-115135 can potentiate the effect of 5-FU in cells in which p53 function is disrupted, these data suggest strongly that RPR 115135 significantly enhances the efficacy of 5-FU only when p53 is functioning. PMID- 12115541 TI - Protein phosphatase-2A regulates endothelial cell motility and both the phosphorylation and the stability of focal adhesion complexes. AB - Solid cancers must stimulate expansion of the vascular network for continued growth. The process of angiogenesis involves endothelial cell migration so as to reorganize into vessel structures. The extent of cellular motility is regulated in part by the balance between serine/threonine kinases and protein phosphatases. In the present study, we show a decline in the activity of the serine/threonine phosphatase PP-2A in endothelial cells whose motility is stimulated by exposure to medium conditioned by either murine LLC cells or human HNSCC cells. Inhibition of endothelial cell PP-2A pharmacologically by treatment with okadaic acid also stimulated endothelial cell motility. Identification of mechanisms by which PP-2A inhibition might stimulate endothelial cell motility focused on proteins of the focal adhesions. Inhibition of PP-2A caused hyperphosphorylation of the paxillin serine residues and dephosphorylation of its tyrosine residues, dissolution of FAK/Src/paxillin complexes and decreased phosphorylation of the inhibitory Y529 residue of Src, suggesting increased Src activity. Inhibition of Src activity prevented the stimulation of PP-2A-inhibited cell motility. Our results suggest an interrelationship between tumor inhibition of PP-2A, dissolution of focal adhesion complexes and stimulated motility of endothelial cells. PMID- 12115542 TI - Retinoic acid modulates the ability of macrophages to participate in the induction of the angiogenic phenotype in head and neck squamous cell carcinoma. AB - Angiogenesis, an essential step in the development of neoplasia, is a complex process that involves the interaction of tumor cells with stromal cells. Tumor associated macrophages (TAMs) can participate in the induction of angiogenesis and are of prognostic value in some neoplasms. Specimens from head and neck squamous cell carcinomas (HNSCC) often contain large numbers of TAMs. In addition, experimental evidence has demonstrated that HNSCC tumor cells can attract and activate macrophages to participate in the expression of the angiogenic phenotype. These findings suggest that antiangiogenic therapies for HNSCC must include strategies that will block the recruitment of macrophages into the tumor microenvironment. We investigated the ability of retinoic acid (RA) to modulate the ability of tumor cells to recruit and activate monocytes for participation in tumor angiogenesis. Owing to a decrease in the secretion of MCP 1 and transforming growth factor-beta 1 (TGF-beta 1), tumor cells treated with RA were unable to induce peripheral blood monocyte (PBM) chemotaxis. Also, as a result of the decrease in TGF-beta 1 secretion, RA-treated tumor cells were unable to activate macrophages for secretion of vascular endothelial growth factor (VEGF) and interleukin-8 (IL-8). In addition to its affects on tumor cells, RA also directly altered the ability of monocytes to participate in the tumor angiogenesis process. PBM exposed to RA were unable to migrate toward inducers of PBM such as MCP-1 and TGF-beta 1. Finally, RA decreased the ability of tumor-activated macrophages to secrete IL-8 and VEGF. These data demonstrate alternative mechanisms by which RA may modulate angiogenesis in the tumor microenvironment. In addition, it underscores the necessity to develop antiangiogenic treatment protocols that can block each of the ways in which new blood vessel growth is induced in tumor microenvironments. PMID- 12115543 TI - P53 is the strongest predictor of survival in high-risk primary breast cancer patients undergoing high-dose chemotherapy with autologous blood stem cell support. AB - Our purpose was to determine the predictive value of tumor biologic parameters in patients with HRPBC who received HDCT with ASCT as first-line treatment. From September 1992 to May 2000, 149 stage II or III HRPBC patients were enrolled in a single-arm trial using a tandem HDCT regimen followed by ASCT. Her2/neu, p53, Ki67 and bcl-2 protein expression was studied using immunohistochemic staining on formalin-fixed, paraffin-embedded primary tumor sections. DNA content of tumor cells (DNA index) and tumor cell proliferation (SPF) were measured by DNA flow cytometry. The relationship between these tumor biologic parameters, on the one hand, and DFS, DDFS and OS, on the other, was analyzed. With a median follow-up of 43 months (range 7-106), p53 protein accumulation (p = 0.000004), negative combined hormone receptor status (p = 0.003) and Her2/neu overexpression (p = 0.02) were significant negative predictors of OS in univariate analysis. A poorer DFS was associated with p53 positivity (p = 0.04) and nodal ratio > or = 0.8 (p = 0.008). Poorer DDFS was associated with p53 positivity (p = 0.03). In multivariate analysis, Her2/neu overexpression (RR = 3.86, 95% CI 1.48-10.1, p = 0.006) and p53 overexpression (RR = 6.06, 95% CI 2.22-16.52, p < 0.001) proved to be independent predictors of adverse OS. p53 overexpression was the only independent predictor of DFS (RR = 2.21, 95% CI 1.07-4.57, p = 0.03). p53 overexpression and Her2/neu overexpression are independent negative predictors of survival in HRPBC treated with HDCT. The adverse impact of these biologic features was probably not altered by HDCT. For HRPBC patients with tumors not overexpressing Her2/neu or p53, HDCT may be an appropriate approach to achieve long-term survival and tumor control. PMID- 12115545 TI - p53 Codon 72 polymorphism in gastric cancer susceptibility in patients with Helicobacter pylori-associated chronic gastritis. AB - p53 codon 72, which produces variant proteins with an arginine (Arg) or proline (Pro), has been reported to be associated with cancers of the lung, esophagus and cervix. However, there have been no reports on the p53 codon 72 polymorphism in gastric cancer susceptibility in patients with Helicobacter pylori-associated chronic gastritis (H. pylori-CG). We, therefore, examined the polymorphism in 117 gastric cancer patients (72 intestinal type and 45 diffuse type) with H. pylori CG and 116 H. pylori-CG patients without gastric cancer as controls. Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis was performed to analyze the p53 codon 72 polymorphism. The crude genotypic frequencies in the gastric cancer patients were similar to those of the controls. However, when gastric cancers were classified by histologic subtype, the Pro/Pro was more frequent in the patients with diffuse type gastric cancer than in the controls (22.2% of cases vs. 12.1% of controls). The Pro/Pro genotype was associated with a 2.98-fold higher risk of diffuse-type cancer compared to the Arg/Arg genotype (95% confidence interval [CI] 1.07-8.32, p = 0.038). These results suggest that the Pro/Pro genotype at p53 codon 72 contributes to susceptibility for diffuse-type gastric cancer in patients with H. pylori-CG. The p53 codon 72 polymorphism may serve as the genetic marker for the risk assessment of the diffuse-type gastric cancer development in patients with H. pylori-CG. PMID- 12115544 TI - Overexpression of the Wilms' tumor gene WT1 in de novo lung cancers. AB - Expression of the Wilms' tumor gene WT1 in de novo lung cancer was examined using quantitative real-time RT-PCR and immunohistochemistry. Overexpression of the WT1 gene was detected by RT-PCR in 54/56 (96%) de novo non-small cell lung cancers examined and confirmed by detection of WT1 protein with an anti-WT1 antibody. Overexpression of the WT1 gene was also demonstrated in 5/6 (83%) de novo small cell lung cancers by immunohistochemistry. Furthermore, when the WT1 gene was examined for mutations by direct sequencing of genomic DNA in 7 lung cancers, no mutations were found. These results suggest that the nonmutated, wild-type WT1 gene plays an important role in tumorigenesis of de novo lung cancers and may provide us with the rationale for new therapeutic strategies for lung cancer targeting the WT1 gene and its products. PMID- 12115547 TI - Transcriptional profiling of a human papillomavirus 33-positive squamous epithelial cell line which acquired a selective growth advantage after viral integration. AB - Alterations in gene expression represent key events in carcinogenesis. We have studied HPV-induced cervical carcinogenesis, using an HPV-33-positive cell line (UT-DEC-1) established from a low-grade vaginal dysplasia (VAIN-I). Early-passage cells contained HPV-33 in episomal form, but these were superseded at later passages by cells carrying only integrated virus. To gain insight into the biologic significance of HPV integration, we compared the level of gene expression in normal vaginal keratinocytes, early-passage and late-passage UT-DEC 1 cells, using cDNA microarrays. Total RNA was isolated from cells by CsCl gradient centrifugation, reverse-transcribed with MMLV reverse transcriptase and labeled with alpha-(32)P ATP. A cDNA microarray expression profile analysis was performed with Clontech's Human Cancer 1.2 cDNA expression array kit. The 16 upregulated genes (cut-off 2-fold), identified by comparing both cell types to control keratinocytes, appeared to support cell-cycle progression or to be functional in mitosis. These included, e.g., MCM4 DNA replication licensing factor, cdc2p34 and chromatin assembly factor 1 p48 subunit. Downregulated genes (44 altogether) interfered with apoptosis and cell adhesion, including the apoptosis-inducing genes FRAP, Bik and caspase-9 precursor. The most significant differences between the late and early passages (29 and 46 constantly up- and downregulated genes without any fluctuation) were overexpression of the transcription factors E2F5 with its dimerization partner DP1, NF-kappa B and serine/threonine kinases and underexpression of enzymes of the MAPK pathway. Acquisition of a selective growth advantage after viral integration might be explained by a major shift from a MAPK pathway to cell-cycle dysregulation (G(2)/M). PMID- 12115546 TI - Broadened ligand responsiveness of androgen receptor mutants obtained by random amino acid substitution of H874 and mutation hot spot T877 in prostate cancer. AB - In a subset of endocrine therapy-resistant prostate cancers, amino acid substitutions H874Y, T877A and T877S, which broaden ligand specificity of the ligand binding domain (LBD) of the androgen receptor (AR), have been detected. To increase our knowledge of the role of amino acid substitutions at these specific positions in prostate cancer, codons 874 and 877 were subjected to random mutagenesis. AR mutants were screened in a yeast readout system for responsiveness to 5 alpha-dihydrotestosterone, progesterone and dehydroepiandrosterone. At position 874, only the histidine to tyrosine substitution could broaden AR ligand specificity. At position 877, 4 ligand specificity broadening substitutions were found: T877A, T877S, T877C and T877G. The latter 2 were not found in prostate cancer. The AR mutants were tested in mammalian (Hep3B) cells for responsiveness to 13 different ligands. All mutants displayed their own ligand specificity spectrum. Importantly, AR(H874Y) and AR(T877A) could be activated by cortisol. According to the 3-dimensional structure of the AR LBD, T877 interacts directly with the 17 beta-hydroxyl group of androgens. All amino acid substitutions identified at position 877 had smaller side chains than the threonine in the wild-type receptor, indicating that increased space in the ligand binding pocket is important in broadened ligand specificity. Because H874 does not interact directly with the ligand, its substitution by a tyrosine is expected to change the ligand binding pocket conformation indirectly. For T877C and T877G substitutions, 2-point mutations are required, and for H874Y, T877A and T877S substitutions, only a 1-point mutation is sufficient. This most likely explains that the latter 3 have been found in prostate cancer. PMID- 12115548 TI - Prevalence of human papillomavirus in cervical cancer: a multicenter study in China. AB - A large-scale epidemiologic survey on the prevalence of different types of human papillomavirus (HPV) in cervical cancer in China is indicated because of the implications for the development of diagnostic probes and vaccines against cervical cancer. A total of 809 cervical cancer specimens were collected from 5 regions in China including Shanghai, Guangzhou, Sichuan, Beijing and Hong Kong. HPV DNA was detected in 83.7% of the specimens. HPV-16 was present in 79.6%, HPV 18 in 7.5%, HPV-52 in 2.6% and HPV-58 in 3.8% of all HPV-positive specimens. The prevalences of HPV-16 and HPV-18 in Hong Kong were 61.7 and 14.8%, respectively, representing a lower HPV-16 and a higher HPV-18 proportion compared with the other regions. HPV-16 remained the most common HPV infection in both squamous cell carcinoma (SCC) and adenocarcinoma (AC). The proportion of HPV-18 infection was significantly higher in AC than in SCC. PMID- 12115549 TI - A sequence repeat in the insulin-like growth factor-1 gene and risk of breast cancer. AB - Insulin-like growth factor-1 (IGF-I), a potent mitogen, is hypothesized to influence breast cancer risk. In 3 previous studies, a polymorphism in the IGF-1 gene (sequence repeat length) was associated with plasma IGF-I level. We evaluated prospectively the relationships among a (CA)(n) repeat polymorphism in the IGF-1 gene, IGF-I level and breast cancer risk in a nested case-control study conducted within the Nurses' Health Study. Blood samples were collected in 1989 1990; up to June 1994, we identified 463 cases of breast cancer. One to 2 controls were selected per case, matched by age, menopausal status, postmenopausal hormone use, month and time of day of blood collection and fasting status, for a total of 622 controls. Although no significant trend was observed, plasma IGF-I levels were significantly lower among controls, with no copy of the 19 allele, compared with those homozygous for the 19 (CA)(n) repeat length (146 and 173 ng/ml, respectively; p-value for pairwise mean comparison = 0.005). In conditional logistic regression, controlling for established breast cancer risk factors, we observed no significant association between (CA)(n) repeat length genotype and risk of breast cancer [compared with repeat genotype 19/19-18/19 genotype relative risk (RR) = 0.96, 95% confidence interval (CI) = 0.56-1.64; 18/20 genotype RR = 0.92, 95% CI = 0.39-2.19; 19/20 genotype RR = 1.16, 95% CI = 0.82-1.64; 19/21 genotype RR = 0.69, 95% CI = 0.42-1.14; 20/20 genotype RR = 0.55, 95% CI = 0.28-1.10; 20/21 genotype RR = 0.72, 95% CI = 0.29-1.79]. Results did not vary substantially when evaluated according to menopausal status, tumor receptor status or category of other breast cancer risk factors. Although a modest association cannot be excluded, our data do not support an important relation between this IGF-1 gene polymorphism and breast cancer risk. PMID- 12115551 TI - Cancer incidence among Sami in Northern Finland, 1979-1998. AB - The Sami population living in Northern Finland represents a specific genetic background and a way of life that is different from other Finns. A cohort of 2,100 Sami and 4,174 non-Sami people from the 2 northernmost municipalities of Finland on 31 December 1978 was identified from the national Population Register and followed up through the Finnish Cancer Registry for cancer incidence during 1979-1998. There were 111 cancer cases among the Sami, while the expected number based on the average cancer incidence in the Finnish population was 173. Among the non-Sami cohort members, there were 226 cases of cancer vs. 224 expected cases. The Sami had significantly decreased incidence of cancers of the prostate [standardised incidence ratio (SIR) 0.25; 95% confidence interval (CI) 0.08-0.58] and breast (SIR 0.36; 95% CI 0.14-0.73), similarly for both localised and nonlocalised tumours. Low SIRs were also observed for bladder cancer (SIR 0.28; 97% CI 0.03-0.99) basal cell carcinoma of the skin (SIR 0.12; 95% CI 0.03-0.30) and other nonmelanoma skin cancers (SIR 0; 95% CI 0-0.63). In contrast to other subcategories of the Sami, the Skolts, whose lifestyle stems from areas that now belong to Russia, showed a nonreduced overall cancer risk and a significantly elevated risk for stomach cancer (SIR 3.8; 95% CI 1.5-7.8). The low cancer incidence among the other Sami populations in Finland cannot be fully explained by their specific way of life. It seems likely that the Sami ethnicity carries a reduced cancer incidence level. Although many Sami have been exposed to radioactive fallout from the nuclear weapon tests via their reindeer-rich diet, this does not seem to affect their cancer risk. PMID- 12115550 TI - Tamoxifen and toremifene treatment of breast cancer and risk of subsequent endometrial cancer: a population-based case-control study. AB - A population-based case-control study was performed to evaluate the risk of endometrial cancer related to tamoxifen or toremifene treatment. All patients with breast cancer diagnosis since 1980 in Finland who subsequently developed an endometrial cancer by the end of 1995 and 3 matched controls were identified among the 38,000 breast cancer patients of the Finnish Cancer Registry database. Detailed information on treatment of breast cancer and potential confounders was collected from hospital records. The OR for tamoxifen treatment (59 cases), adjusted for significant cofactors (increased risk associated with obesity, low parity and PR positivity) was 2.9 (95% CI 1.8-4.7). The OR for toremifene (3 cases) was 0.9 (95% CI 0.3-3.9). The OR related to adjuvant tamoxifen treatment reached its maximum 2-5 years after the beginning of treatment (OR 5.1, 95% CI 2.1-13), while the OR for tamoxifen used for palliative treatment of advanced breast cancer was especially high after a lag of over 5 years (OR 9.5, 95% CI 2.5 36). The risk increase due to tamoxifen was slightly higher if the age at initiation was below 55, and risk was more pronounced among patients with well differentiated endometrial cancer than patients with cancers of clinical grades 2 or 3. According to our results, treatment with tamoxifen increases the risk of endometrial cancer. Due to the rare use of toremifene up to the mid-1990s, the risk assessment concerning it was inconclusive. PMID- 12115552 TI - Increased emotional distress in daughters of breast cancer patients is associated with decreased natural cytotoxic activity, elevated levels of stress hormones and decreased secretion of Th1 cytokines. AB - DBCP who are aware of their increased risk of developing breast cancer may suffer from high emotional distress. Chronic stress may interfere with NCA and low NCA is associated with increased cancer risk. We studied 80 DBCP and 47 age- and education-matched healthy females (controls). Heparinized venous blood (30 ml) was drawn from all subjects between 8 and 9 A.M., and each participant answered a set of psychologic questionnaires. In addition, the first-morning urine sample was collected. DBCP scored significantly higher in emotional distress compared to controls. Levels of stress hormones in DBCP were higher and in vitro secretion of IL-2, IL-12 and IFN-gamma lower compared to controls. NCA against NK-resistant (MCF-7, COLO-205, U937) and NK-sensitive (K562) cell lines was significantly lower in DBCP and much less augmented by in vitro preincubation with IL-2 or IL 12 compared to controls. NCA and in vitro Th1 cytokine secretion were inversely correlated with the degree of emotional distress and the level of stress hormones in blood or urine. High emotional distress and elevated levels of stress hormones are associated with impaired immune surveillance functions in DBCP. This may contribute to the increased risk of DBCP to develop breast cancer. An interventional trial to enhance coping and reduce stress levels may help to decrease the risk for breast cancer onset in DBCP. PMID- 12115553 TI - Food groups and laryngeal cancer risk: a case-control study from Italy and Switzerland. AB - Besides tobacco and alcohol, diet has been thought to be associated with laryngeal cancer risk. We thus analyzed the role of various food groups, as well as specific seasoning fats, in a case-control study conducted in Northern Italy and the Swiss Canton of Vaud from 1992 to 2000. Our study included 527 incident, histologically confirmed cases and 1,297 frequency-matched controls, selected among patients admitted to the same hospitals as cases for acute, nonneoplastic conditions, unrelated to smoking, alcohol consumption and long-term modifications of diet. The subjects' usual diet was investigated through a validated food frequency questionnaire, including 78 foods and beverages. Odds ratios (OR) and 95% confidence intervals (CI) were estimated using unconditional multiple logistic regression models. After adjustment for major confounding factors, a significant trend of increasing risk was observed for eggs (OR = 1.7 for the highest compared to the lowest quintile), red meat (OR = 3.1), processed meat (OR = 1.7), fish (OR = 1.6) and sugars (OR = 1.6). Significant inverse associations were observed for pulses (OR = 0.7), raw vegetables (OR = 0.2), cooked vegetables (OR = 0.3), citrus fruit (OR = 0.6) and other fruit (OR = 0.5). In regard to seasoning fats, a significant reduction of cancer risk was observed for olive oil (OR = 0.4) and specific seed oils (OR = 0.6), while mixed seed oils were directly associated with laryngeal cancer risk (OR = 2.2). Our study suggests that increasing vegetables and fruit, decreasing meat consumption and perhaps substituting olive oil or specific seed oils for other types of seasoning lipids might help reduce laryngeal cancer risk. PMID- 12115554 TI - Personality characteristics and the risk of breast cancer: a prospective cohort study. AB - Various personality characteristics have been suggested to increase the risk of breast cancer but reliable epidemiologic data on this issue are limited. We prospectively investigated the relationship between personality characteristics and the risk of breast cancer in 12,499 Finnish women aged 18 years or more. In health questionnaires in 1975 and 1981, these women completed at least one of the following personality scales: Eysenck extroversion, Bortner type A behaviour and author-constructed measure of hostility. They also reported about other potential breast cancer risk factors. From 1976-1996, 253 cases of breast cancer were identified by record linkage with the Finnish Cancer Registry. Proportional hazard models were used to estimate hazard ratios (HR) and 95% confidence intervals (CI). The multivariable HRs of breast cancer for women with intermediate level (scores 3-6) and high level (7-9) of extroversion in 1975 were 1.18 (95% CI 0.87-1.60) and 0.97 (95% CI 0.64-1.47), respectively, compared to those with low level (0-2). These results remained unaltered when the level of extroversion was determined as the average of the 1975 and 1981 reports. There was also no increase in breast cancer risk in relation to type A behaviour and hostility. Furthermore, we observed no substantial joint effects of personality characteristics on the risk of breast cancer. In conclusion, our data do not support the existence of an important role for personality in the aetiology of breast cancer. These findings are reassuring to those who have believed the contrary. PMID- 12115555 TI - Pharmacokinetic and tumor-seeking properties of recombinant and nonrecombinant anti-carcinoembryonic antigen antibody fragments. AB - Production of recombinant antibody fragments in bacterial expression systems results in intentional or fortuitous differences compared to the original products prepared by hybridoma technology. These differences may have significant effects not only on antigen-binding properties but also on pharmacokinetic and tumor-seeking properties. Our major goal was to investigate some of these possible differences. We produced in Escherichia coli an rFab' fragment containing only 1 cysteine residue in the hinge region; the fragment was derived from a mouse MAb (F6) specific for CEA. The rFab' had a slightly lower m.w. and a higher isoelectric point relative to the corresponding nonrecombinant fragment (pFab'). This was explained by the absence of N-glycosylation on the V kappa domain of rFab'. V kappa glycosylation had no significant effect on antibody binding affinity and kinetics. However, rFab' was eliminated from the circulation much faster than pFab', and the maximal dose accumulated in the tumor was reduced relative to pFab'. Thus, glycosylation appears to modify the targeting efficiency of antibody fragments. rF(ab')(2) fragments were obtained either spontaneously from the culture supernatant of E. coli or by chemical cross-linking [rcF(ab')(2)]. We observed improved tumor targeting with rcF(ab')(2) compared to rF(ab')(2), which could be explained by the greater stability of the thioether compared to the disulfide linkage. These results demonstrate that a single cysteine residue in the hinge region of rFab' is particularly well suited to prepare stable, chemically coupled, bivalent or bispecific antibodies, avoiding intrahinge disulfide bonding and thus achieving higher production yields. PMID- 12115556 TI - Family history of breast cancer, age and benign breast disease. AB - A major risk factor for breast cancer is having a first-degree family history of the disease. Benign breast disease (BBD), particularly atypical hyperplasia, is also associated with an increased risk of breast cancer. However, the relationship between family history of breast cancer and BBD is unclear. From 1989 through 1997, 80,995 participants in the Nurses' Health Study II were followed; 16,849 reported a first diagnosis of BBD. Pathology slides were reviewed for 1,465 women who reported having a tissue biopsy, and these were classified as nonproliferative BBD, proliferative BBD without atypia or atypical hyperplasia. Women with a family history of breast cancer were more likely to report a physician diagnosis of BBD [rate ratio (RR) = 1.38, 95% confidence interval (CI) 1.29-1.46]. The magnitude of this association declined with age from RR = 1.96 (95% CI 1.55-2.47) at 25-29 years to RR = 1.20 (95% CI 0.95-1.52) at age 45-50 years. Among women with proliferative disease, those with a family history of breast cancer were almost 3 times as likely to have atypia (prevalence odds ratio = 2.72, 95% CI 1.23-5.89) than those with no family history. In conclusion, women with a family history of breast cancer appear to be at increased risk of being diagnosed with BBD, in particular the high-risk types of BBD associated with a greatly increased risk of breast cancer. This link adds weight to the belief that BBD with atypia is a precursor or marker lesion for breast cancer. PMID- 12115558 TI - Strategies for vascular targeting in tumors. PMID- 12115559 TI - Prognostic value of P53 mutations in rectal carcinoma. AB - The influence of p53 mutations on the response to ionizing radiation and survival was retrospectively evaluated in patients treated with preoperative radiotherapy for rectal carcinoma. From 1989 to 1991, 86 rectal cancer patients treated by preoperative radiotherapy were included in this series. For all patients, endorectal sonography (to define ultrasonography TNM [uTNM]) was performed before treatment; 19 patients were classified as stage 1, 27 as stage 2 and 40 as stage 3. Response to radiotherapy (39 Gy in 13 fractions delivered in 17 days) was assessed by comparing the uT and the T obtained by histologic examination of the resected specimen (TNM classification). A rectal cancer biopsy was performed before treatment and enabled the search for p53 mutations by denaturing gradient gel electrophoresis (DGGE) and sequencing. The status of the p53 gene was correlated with the response to radiotherapy and survival. Forty-nine percent of the tumors presented abnormal DGGE profiles. The prevalence of p53 mutations was significantly higher in patients who did not respond to radiotherapy (63%) than in those who did respond (34%) (p < 0.01). Presence of a p53 mutation was associated with significantly shorter 5-year survival compared to patients without mutations (p < 0.02). In a multivariate analysis, p53 mutation status remained a prognostic factor independent of tumor posttreatment staging (p < 0.05). p53 status is an independent prognostic factor of response to radiotherapy and survival in rectal carcinoma. PMID- 12115561 TI - Desferrioxamine enhances AIDS-associated Kaposi's sarcoma tumor development in a xenograft model. AB - Iron is suspected to be involved in the induction and/or progression of various human tumors. More particularly, we have previously shown that iron may be involved in the pathogenesis of Kaposi's sarcoma (KS). We have also shown that the iron chelator desferrioxamine (DFO) has a potent anti-KS activity in vitro, suggesting that it may represent a potential therapeutic approach for the treatment of KS. The present study was designed to investigate the effect of DFO on the growth of human KS xenografts in immunodeficient mice. Unexpectedly, we found that mice treated with DFO (400 mg/kg, 3 times weekly) (n = 30) exhibited a marked enhancement of tumor growth compared with control mice (n = 33) (230 +/- 134 mm(2) versus 143 +/- 70 mm p < 0.01). No enhancement of tumor growth was seen in mice treated with iron-saturated DFO. At least 2 findings suggest that this paradoxic pro-KS activity occurred independently of mice iron stores. First, treatment with DFO had only a marginal effect on ferritin and hematocrit levels. Second, induction of effective iron depletion by an iron-poor diet (6.7 mg iron/kg diet) (n = 23) did not have a deleterious effect on the growth of the KS xenografts. The lesions obtained from the DFO-treated animals exhibited a significantly decreased apoptotic index (p < 0.05), indicating that some antiapoptotic mechanism induced by DFO may be operating in vivo to favour tumor growth. In conclusion, our data show that DFO has a stimulatory effect on KS growth in immunodeficient mice, suggesting that this drug is not indicated in patients with KS. PMID- 12115560 TI - Promoting effects of monomethylarsonic acid, dimethylarsinic acid and trimethylarsine oxide on induction of rat liver preneoplastic glutathione S transferase placental form positive foci: a possible reactive oxygen species mechanism. AB - Dimethylarsinic acid (DMA) is a major metabolite of inorganic arsenicals, which are epidemiologically significant chemicals in relation to liver cancer in mammals. The present study was conducted to determine the promoting effects of organic arsenicals related to DMA [monomethylarsonic acid (MMA) and trimethylarsine oxide (TMAO)] on rat liver carcinogenesis using a liver medium term bioassay (the Ito test). Male, 10-week-old, F344 rats were given a single i.p. injection of diethylnitrosamine at a dose of 200 mg/kg b.w. as an initiator. Starting 2 weeks thereafter they received 100 ppm of MMA, DMA or TMAO in their drinking water, or no supplement as a control, for 6 weeks. All animals underwent 2/3 partial hepatectomy in week 3 after initiation. Quantification of glutathione S-transferase placental form (GST-P)-positive foci as preneoplastic lesions in liver sections revealed significantly increased numbers and areas in all 3 treated groups compared with controls. Hepatic microsome cytochrome P-450 content was markedly increased with all 3 arsenic treatments. Markedly elevated CYP 2B1 protein levels and CYP 2B1/2 mRNA levels were thus observed in all cases. The potency of promotion was similar for MMA, DMA and TMAO. Since hydroxyradicals were found to be generated in the relatively early phase while methylated arsenicals were metabolized in liver, the resultant oxidative stress might have promoted lesion development. PMID- 12115562 TI - Molecular profiling of human chondrosarcomas for matrix production and cancer markers. AB - Chondrosarcoma is the second most common malignant bone tumor, characterized by production of abundant extracellular matrix resembling hyaline cartilage. To better understand the molecular pathogenesis of chondrosarcoma, we analyzed 12 chondrosarcomas for their production of connective tissue components and SOX9, a key regulator of normal chondrocyte differentiation. Furthermore, 10 chondrosarcoma samples were screened for additional changes in gene expression using cDNA array analysis. In Northern analysis, several tumors were found to express type II collagen mRNA at levels comparable to fetal cartilage used as a control. Interestingly, the highest levels of type II collagen mRNA were seen in 2 of the 3 grade 3 chondrosarcomas, which also exhibited the highest mRNA levels of SOX9 and "prechondrogenic" pro alpha 1(IIA) collagen. Expression of SOX9 in human chondrosarcomas is novel and suggests that chondrosarcomas originate from a multipotent stem cell committed to differentiation along the chondrogenic pathway. Results of the cDNA array analyses emphasize the heterogeneous nature of chondrosarcoma as no single transcript was systematically up- or downregulated in all tumors analyzed. Among the interesting changes observed was upregulation of decorin mRNA in 7 of the 10 tumors analyzed. Further studies are needed to determine whether decorin plays a role in the pathogenesis of chondrosarcoma. The cDNA arrays also revealed discrepancies from Northern and RNase protection analyses in transcript levels of matrix components, emphasizing the need to validate cDNA array data with other techniques. PMID- 12115563 TI - Impact of PTEN expression on the outcome of hepatitis C virus-positive cirrhotic hepatocellular carcinoma patients: possible relationship with COX II and inducible nitric oxide synthase. AB - PTEN, a novel tumor suppressor, functions as a regulator of both cell cycle progression and apoptosis. PTEN gene is frequently mutated or deleted in several malignancies including human hepatocellular carcinoma (HCC). The clinical significance and prognostic value of PTEN expression in HCC or in the surrounding non-cancerous parenchyma remain obscure. Using immunohistochemistry, we analyzed the PTEN protein expression in 46 tissue sections collected from surgically resected hepatitis C virus (HCV)-positive cirrhotic HCC patients. Although the surrounding normal liver tissue was strongly expressing PTEN in 42 cases (91.3%), the immunostaining intensity was low in 29 (63.1%) and high in 17 (36.9%) of the HCCs. Additionally a significant positive correlation was identified between low PTEN expression in the HCC and increased expression of iNOS and COX II in the surrounding liver. The overall survival was significantly longer for the HCC patients with high PTEN expression than patients with low PTEN expression. Univariate analysis revealed PTEN expression as an independent prognostic factor for patients survival. By Western blot analysis we also found that the Akt/PKB signaling, which is negatively regulated by PTEN, was upregulated in the HCCs in comparison to its expression in the surrounding liver tissue. These results demonstrate that downregulation of PTEN in the tumor is an important step in HCV positive cirrhotic hepatocarcinogenesis and might result in concomitant upregulation of iNOS and COX II in the surrounding liver in favor of tumor promotion. PMID- 12115564 TI - Effector mechanisms of norcantharidin-induced mitotic arrest and apoptosis in human hepatoma cells. AB - NCTD is a demethylated form of cantharidin with antitumor properties, which is now in use as a routine anticancer drug against hepatoma. However, there is limited information on the effect of NCTD on human cancer cells. In the present study, NCTD inhibited proliferation, caused mitotic arrest, then progressed to apoptosis within 96 hr in 3 human hepatoma cell lines: HepG2, Hep3B and Huh-7. NCTD treatment (5 microg/ml) enhanced the expression of Cdc25C and p21(Cip1/Waf1), increasing the phosphorylation of these 2 proteins. In addition, NCTD treatment induced an earlier increase in cyclin B1-associated histone H1 kinase activity within 48 hr, but an approximately 70% reduction of both protein level and kinase activity of cyclin B1 was observed at 72 hr. Treatment with NCTD significantly decreased the expression of p53 protein but did not affect the expression of Cdk1 and p27(Kip1). Moreover, NCTD treatment also increased the phosphorylation of Bcl-2 and Bcl-X(L) but did not affect the expression of Bax or Bad. Bcl-2 phosphorylation appears to inhibit its binding to Bax since less Bax was detected in immunocomplex with Bcl-2 in NCTD-treated HepG2 cells. In addition, NCTD treatment caused activation of caspase-9 and caspase-3, preceding DNA fragmentation and morphologic features of apoptosis. Pretreatment with the broad-spectrum caspase inhibitor z-VAD-fmk markedly inhibited NCTD-induced caspase-3 activity and cell death. These results suggest that phosphorylation of p21(Cip1/Waf1) and Cdc25C and biphasic regulation of cyclin B1-associated kinase activity may contribute to NCTD-induced M-phase cell-cycle arrest. Furthermore, the increase of p21(Cip1/Waf1), phosphorylation of Bcl-2 and Bcl-X(L), activation of caspase-9 and caspase-3 may be the molecular mechanism through which NCTD induces apoptosis. PMID- 12115565 TI - Expression of MUC1 splice variants in benign and malignant ovarian tumours. AB - MUC1 is expressed on the surface of ovarian cancer cells. Nine different splice variants of MUC1 have been described, but no study has reported on the expression of MUC1 isoforms in human ovarian cancer. Our study compares patterns of expression of MUC1 splice variants of malignant and benign ovarian tumours. Ovarian tissue samples were taken from patients with benign ovarian tumours (n = 34) and from patients who had surgery for primary (n = 47) or recurrent (n = 8) ovarian cancer. RT-PCR for MUC1 splice variants A, B, C, D, X, Y, Z, REP and SEC was performed and their expression compared to clinical and histopathologic parameters. Variants A, D, X, Y and Z were more frequently expressed in malignant than in benign tumours. All primary ovarian cancer cases were positive for variant REP but negative for variant SEC. No significant association of the expression of MUC1 splice variants with the response to chemotherapy or patient survival could be demonstrated. Expression of MUC1 splice variants A, D, X, Y, Z and REP is associated with the presence of malignancy, whereas expression of MUC1/SEC is associated with the absence of malignancy. PMID- 12115566 TI - Differential gene expression in breast cancer cell lines and stroma-tumor differences in microdissected breast cancer biopsies revealed by display array analysis. AB - To examine gene-expression patterning in late-stage breast cancer biopsies, we used a microdissection technique to separate tumor from the surrounding breast tissue or stroma. A DD-PCR protocol was then used to amplify expressed products, which were resolved using PAGE and used as probe to hybridize with representative human arrays and cDNA libraries. The probe derived from the tumor-stroma comparison was hybridized with a gene array and an arrayed cDNA library derived from a GCT of bone; 21 known genes or expressed sequence tags were detected, of which 17 showed differential expression. These included factors associated with epithelial to mesenchymal transition (vimentin), the cargo selection protein (TIP47) and the signal transducer and activator of transcription (STAT3). Northern blot analysis was used to confirm those genes also expressed by representative breast cancer cell lines. Notably, 6 genes of unknown function were restricted to tumor while the majority of stroma-associated genes were known. When applied to transformed breast cancer cell lines (MDA-MB-435 and T47D) that are known to have different metastatic potential, DD array analysis revealed a further 20 genes; 17 of these genes showed differential expression. Use of microdissection and the DD-PCR array protocol allowed us to identify factors whose localized expression within the breast may play a role in abnormal breast development or breast carcinogenesis. PMID- 12115567 TI - Suppression of growth and increased cellular attachment after expression of DAL-1 in MCF-7 breast cancer cells. AB - The differentially expressed in adenocarcinoma of the lung (DAL-1) gene, which shares significant homology with members of the 4.1/ezrin/radixin/moesin/neurofibromatosis 2 (ERM/NF2) protein family, has previously been shown to suppress growth in lung cancer cell lines. This gene localizes to chromosome band 18p11.3, which undergoes loss of heterozygosity (LOH) in nonsmall cell lung carcinomas and a significant proportion of ductal carcinomas in situ (DCIS) of the breast. This finding suggests that alteration of gene(s) (possibly DAL-1) within this chromosomal region may be important early in the progression of breast disease. We generated MCF-7 cell lines expressing DAL-1 constitutively or under the control of an inducible promoter and analyzed the effect of DAL-1 expression on growth. These investigations revealed that the DAL 1 protein suppresses the growth of MCF-7 cells and may do so in part through the induction of apoptosis. In addition, expression of DAL-1 increased attachment of these cells to a variety of extracellular matrices. This is the first evidence that the DAL-1 protein functions at the interface between cell adhesion and apoptosis in controlling cell growth. PMID- 12115568 TI - Aberrant methylation of the CDKN2a/p16INK4a gene promoter region in preinvasive bronchial lesions: a prospective study in high-risk patients without invasive cancer. AB - Among the identified factors involved in malignant transformation, abnormal methylation of the CDKN2A/p16(INK4a) gene promoter has been described as an early event, particularly in bronchial cell cancerization. Precancerous bronchial lesions (n = 70) prospectively sampled during fluorescence endoscopy in a series of 37 patients at high risk for lung cancer were studied with respect to the methylation status of the CDKN2A gene. Methylation-specific polymerase chain reaction was performed on DNA extracted from pure bronchial cell populations derived from biopsies and detection of p16 protein was studied by immunohistochemistry on contiguous parallel biopsies. Aberrant methylation of the CDKN2A gene promoter was found in 19% of preinvasive lesions and its frequency increased with the histologic grade of the lesions. Methylation in at least 1 bronchial site was significantly more frequent in patients with cancer history, although there was no difference in the outcome of patients with or without methylation in bronchial epithelium. The other risk factors studied (tobacco and asbestos exposure) did not influence the methylation status. There was no relationship between CDKN2A methylation and the evolutionary character of the lesions. Our results confirm that abnormal methylation of the CDKN2A gene promoter is an early event in bronchial cell cancerization, which can persist for several years after carcinogen exposure cessation, and show that this epigenetic alteration cannot predict the evolution of precancerous lesions within a 2-year follow-up. PMID- 12115569 TI - Geographic differences in invasive and in situ breast cancer incidence according to precise geographic coordinates, Connecticut, 1991-95. AB - To evaluate geographical variation of invasive and in situ breast cancer incidence rates using precise geographical coordinates for place of residence at diagnosis, latitude-longitude coordinates pertaining to 10,601 invasive and 1,814 in situ breast cancers for Connecticut women, 1991-95, were linked to US Census information on the 2,905 State census block groups. A spatial scan statistic was used to detect geographic excess or deficits in incidence and test the statistical significance of results, without prior assumptions about the size or location of such areas. The age adjusted invasive cancer incidence rate was 165.3/100,000 women/year. The spatial scan statistic identified 3 places with significantly low incidence rates and 4 places where rates were significantly high. The most probable location of low incidence was rural northeastern Connecticut where risk of disease, relative to elsewhere around the state, was 0.70 (p = 0.0001); the most probable place of elevated incidence was north central Connecticut where a relative risk of 1.34 (p = 0.002) was observed. Incidence of in situ disease was estimated to be significantly high for north central Connecticut (RR = 1.84; p = 0.0001). Geographic differences of invasive and in situ breast cancer incidence were observed. Examining cancer events at the lowest available level of data aggregation is beneficial in highlighting localized rate variations. Such information may enable public health officials to target additional resources for promoting breast cancer screening to specific locations. PMID- 12115570 TI - Determinants of cervical cancer rates in developing countries. AB - Although cervical cancer (CC) is a leading cause of cancer-related deaths in developing countries, incidence rates vary considerably, ranging from 3 to 61 per 10(5) females. Identifying determinants of high vs. low rates may suggest population-level prevention strategies. CC rates for 175 countries were obtained from the IARC. Country-specific behavioral, health, economic and demographic measures were obtained from United Nations agencies and other international organizations. Regression analyses performed for 127 low or medium developed countries identified both geography and religion as independently associated with high CC rates. Among behavioral measures, high fertility rates, early age at birth of first child and high teenage birth rates were significantly associated with high CC rates. Countries with high CC rates had fewer doctors per capita, less immunization coverage, more HIV infections and shorter life expectancies. CC rates also tended to be higher in countries with more spending on health and younger, less educated populations. Patterns of CC rates suggest that programmatic approaches, such as promoting delayed childbearing and sexual monogamy, may be appropriate interventions. For countries with high CC rates and some flexibility in their health-care budgets, a once-in-a-lifetime screen of women 30-50 years of age, using Pap smears, direct visual inspection and/or HPV DNA testing, may be cost-effective. Finally, relatively low immunization rates and a shortage of health-care workers in countries with high CC rates suggest potential challenges for introducing prophylactic HPV vaccines. PMID- 12115571 TI - Relation of childhood brain tumors to exposure of parents and children to tobacco smoke: the SEARCH international case-control study. Surveillance of Environmental Aspects Related to Cancer in Humans. AB - The etiology of childhood brain tumors (CBTs) remains unknown. Tobacco smoke contains several known carcinogens and can induce DNA adducts in human placenta and hemoglobin adducts in fetuses. We present the results of an international case-control study to evaluate the association between CBTs and exposure of parents and children to cigarette smoke. The study was undertaken as part of the SEARCH program of the IARC. Nine centers in 7 countries were involved. The studies mainly covered the 1980s and early 1990s. Cases (1,218, ages 0-19 years) were children newly diagnosed with a primary brain tumor; there were 2,223 population-based controls. Most mothers who agreed to participate were interviewed in person at home. Odds ratios (ORs) were calculated by unconditional logistic regression, adjusted for age, sex and center, for all types of CBT combined, 4 CBT histotypes, 5 age groups and each center. There was no association between the risk of brain tumors in the child and parental smoking prior to pregnancy, maternal smoking or regular exposure to others' cigarette smoke during pregnancy at home or at work, or passive smoking by the child during the first year of life. These results did not change considering the child's age at diagnosis, the histologic type of tumor or center. PMID- 12115572 TI - Attributable risks for familial breast cancer by proband status and morphology: a nationwide epidemiologic study from Sweden. AB - Population attributable factions (PAFs) show the proportion of the disease that could be prevented if the cause could be removed. The PAF for familial breast cancer has not been precisely determined. We used the nationwide Swedish Family Cancer Database on 10.2 million individuals and 190,000 mothers' and 26,000 daughters' breast cancers to calculate familial standardized incidence ratios (SIRs), proportion of cases with a family history and familial PAFs for all invasive and in situ and morphology-specific breast cancers in daughters who were 0-66 years old. The data were calculated by mother only, sister only or both as probands. More than 5,500 familial breast cancers were recorded. The familial SIRs for all invasive breast cancer were 1.79 by breast cancer in the mother only, 2.03 by breast cancer in a sister only and 2.82 by breast cancer in both a mother and sister. The familial PAFs were 3.61, 3.01 and 0.43%, respectively, giving a total PAF of 7.05%. Age-specific risks were shown for the mother and sister history of breast cancer. The PAF values decreased by age when the daughter had a mother history of breast cancer but not when she had a sister history. PAFs did not depend on the morphologic type of breast cancer. The data show that the familial PAF of breast cancer among a 0-66-year-old population of daughters was 7% and independent of the morphologic type. If contribution from the paternal side was allowed for, the PAF would be 11%. PMID- 12115573 TI - Multifunctional anti-angiogenic activity of the cyclic peroxide ANO-2 with antitumor activity. AB - Our focus was to develop an anti-angiogenic drug possessing the inhibitory activity of urokinase-type plasminogen activator (u-PA) production. During preliminary screening, the effects of 13 ozonides on the inhibition of u-PA production in human fibrosarcoma HT-1080 cells and on the inhibition of angiogenesis on chicken embryonic chorioallantoic membranes were determined. Of the ozonides tested, 9 inhibited in vitro u-PA production of HT-1080 cells and 7 of these 9 exhibited strong anti-angiogenic activity. Interestingly, 6 of the 13 ozonides also inhibited cathepsin B activity. 1-Phenyl-1, 4-epoxy-1H,4H naphtho[1,8-de][1, 2]dioxepin (ANO-2) potently inhibited cathepsin B (IC(50) = 0.47 microM) as well as u-PA production. Consequently, ANO-2 was selected for further study. ANO-2 inhibited tube formation by human umbilical vein endothelial cells cultured on Matrigel while exhibiting no cytotoxicity. Additionally, in vivo administration of ANO-2 inhibited angiogenesis induced by mouse Sarcoma-180 cells tested using the mouse dorsal air sac assay. Moreover, ANO-2 also suppressed primary tumor growth and reduced the number of pulmonary metastases caused by Lewis lung carcinoma cells in mice. These in vitro and in vivo activities indicate that ANO-2 has considerable potential as a new and potent anti-angiogenic drug that inhibits both u-PA production and enzymatic activity of cathepsins, indicating that ANO-2 may be a multifunctional inhibitor of angiogenesis. PMID- 12115575 TI - Supra-additive effect with concurrent paclitaxel and cisplatin in vulvar squamous cell carcinoma in vitro. AB - The effect of concurrent paclitaxel and cisplatin was tested in vitro in 5 vulvar squamous cell carcinoma (SCC) cell lines (UM-SCV-1A, -2, -4 and -7 and UT-SCV-3). Chemosensitivity was tested using the 96-well plate clonogenic assay. Paclitaxel concentrations used varied between 0.4 and 1.6 nM, and cisplatin concentrations varied between 0.1 and 0.9 microg/ml. These drug concentrations are clinically achievable. Survival data were fitted to the LQ model, and the area under the curve (AUC) value was obtained with numerical integration. The type of interaction was determined by comparing the AUC ratio of the 2 drugs with the survival fraction (SF) of paclitaxel alone. With all cell lines tested the growth inhibitory effect of simultaneous paclitaxel and cisplatin was at least additive. The effect of the tested combination on the UM-SCV-1A and UT-SCV-3 cell lines was clearly supra-additive with all paclitaxel concentrations tested, and the UM-SCV 4 and UM-SCV-7 cell lines exhibited a supra-additive effect with increasing paclitaxel concentrations. The degree of supra-additivity was dose-dependent in the UM-SCV-7 cell line with increasing synergy at higher paclitaxel doses. In the current study the combination of paclitaxel and cisplatin had a clear additive or supra-additive cytotoxic effect on the vulvar SCC cell lines, and it has been successfully used in other gynecologic malignancies; therefore concurrent paclitaxel and cisplatin also deserves further testing in clinical settings in advanced-stage vulvar carcinoma, which has a poor prognosis. PMID- 12115574 TI - Expression analysis of delta-catenin and prostate-specific membrane antigen: their potential as diagnostic markers for prostate cancer. AB - The current approach to prostate cancer diagnosis has major limitations including the inability of prostate-specific antigen (PSA) assays to accurately differentiate between prostate cancer and benign prostate hyperplasia (BPH) and the imprecision of transrectal ultrasound (TRUS) biopsy sampling. We have employed cDNA microarray screening to compare gene expression patterns in BPH and tumour samples to identify expression markers that may be useful in discriminating between these conditions. Screening of 3 individual cDNA arrays identified 8 genes with expression 3-fold greater in 6 tumour tissues than in 1 nontumour sample and 1 BPH sample. Real-time PCR was used to confirm the overexpression of these 8 genes and 12 genes selected from the literature against a panel of 17 tumours and 11 BPH samples. Two genes, delta-catenin (delta catenin; CTNND2) and prostate-specific membrane antigen (PSMA; FOLH1), were significantly overexpressed in prostate cancer compared to BPH. Prostate epithelial cells stained positively for delta-catenin and PSMA in our prostate cancer tissues, whereas the majority of our BPH tissues were negative for both markers. Thus we have identified delta-catenin (not previously associated with prostatic adenocarcinoma) and confirmed the potential of PSMA as potential candidates for the diagnosis and management of prostate cancer. PMID- 12115576 TI - Clinical use of serum TRA-1-60 as tumor marker in patients with germ cell cancer. AB - TRA-1-60 antigen has been related to the presence of embryonal germ cell carcinoma (EC) and carcinoma in situ. Our study further investigated the clinical efficacy of TRA-1-60 as a serum tumor marker for germ cell cancer in the testis. Three groups of patients with germ cell tumors were included: Group 1, 34 patients with disseminated disease (24 nonseminomatous germ cell tumors [NSGCT] and 10 seminomatous germ cell tumors [SGCT]); this group of patients were followed during the course of chemotherapy with measurements of TRA-1-60, HCG and AFP; Group 2, 28 patients with Stage I NSGCT (22 with embryonal carcinoma [EC] component and 6 without EC-component, median follow-up 15 months); and Group 3, 40 patients with Stage I pure SGCT (median follow-up 15 months). Seventy-eight percent of patients with disseminated EC-positive NSGCT had increased levels of TRA-1-60 before chemotherapy. After chemotherapy, levels of TRA-1-60 had dropped significantly (p < 0.01). Levels of TRA-1-60 did not normalize in 15% of NSGCT and 30% of SGCT patients after chemotherapy. This was not associated with recurrent disease. Approximately one-third of patients with Stage I NSGCT had increased values of TRA-1-60 during follow-up without having a relapse. Contrary to earlier reports TRA-1-60 is not at present useful as a tumor marker in patients with germ cell tumors. Although detecting a few early relapses the rate of false positive elevations in the tumor marker makes it unreliable in the clinical setting. Our study did confirm that elevated levels of TRA-1-60 were present in approximately 80% of patients with disseminated EC-positive NSGCT before start of chemotherapy and chemotherapy induced a significant decrease in levels of TRA-1-60. Thus, the TRA-1-60 antigen might still prove clinically useful provided that the reliability of the assay can be increased. PMID- 12115579 TI - Peripheral lymph node stromal cells can promote growth and tumorigenicity of breast carcinoma cells through the release of IGF-I and EGF. AB - The regional lymph nodes draining primary breast carcinomas are generally the first site to be invaded by disseminating tumor cells. The extent of lymph node involvement remains the most reliable indicator for staging and prognosis of breast cancer. We have investigated host-tumor interactions between breast carcinoma cells and the lymph node stroma, which may control the outcome of lymph node infiltration. In a previous study, we identified integrin-mediated cell adhesion as a correlate of the metastatic potential of human and rat carcinoma cells. Our present objective was to determine whether lymphatic stromal cells can affect cancer cell growth through the elaboration of growth-modulating factors. Two lymphatic stromal cell lines, ST-A4 and ST-B12, were established from normal rat lymph node stromal cell cultures. SFM conditioned by these cells increased the proliferation of human (Hs578T and MCF-7) and rat (TMT-081) breast carcinoma cells by up to 7-fold and augmented their ability to form colonies in semisolid agar by up to 41-fold. This effect was specific as normal, diploid human breast epithelial cells (Hs578Bst), a nontumorigenic, immortalized human breast epithelial cell line (MCF-10A) and a nonmetastatic rat mammary carcinoma cell line (MT-W9B) had either no or reduced responses. RT-PCR analysis revealed that both lymph node stromal cell lines expressed mRNA transcripts for multiple growth factors, including IGF-I, EGF, HGF and PDGF-alpha, and produced detectable levels of IGF-I, EGF and PDGF-alpha, as assessed by Western blotting. Antibody-mediated depletion assays identified IGF-I and EGF as the major mitogenic factors in the CM. The identification of these cells raises the possibility that the lymph node microenvironment may contribute actively to the process of cancer cell dissemination. PMID- 12115580 TI - Effect of DNA repair gene polymorphisms on BPDE-DNA adducts in human lymphocytes. AB - To determine whether variations in DNA repair genes are related to host DNA damage, we investigated the association between polymorphism in the XPD gene (codon 199, 312, 751) and the XRCC1 gene (codon 194, 399) and the presence of benzo(a)pyrene diolepoxide adducts to lymphocyte DNA (BPDE-DNA) in a group of male patients with incident lung cancer, all current smokers. BPDE-DNA adducts were analyzed by high-resolution gas chromatography-negative ion chemical ionization-mass spectrometry. XPD and XRCC1 genotypes were identified by PCR RFLP. XRCC1 and XPD genotypes did not affect the levels and proportion of detectable BPDE-DNA adducts. The patients were also genotyped for the GSTM1 polymorphism, given its role in the detoxification of BPDE. Individuals with the GSTM1 deletion had significantly higher levels of BPDE-DNA adducts when they were XPD-Asp312Asp+Lys751Lys than carriers of at least one variant allele. No such association was found with the XRCC1 genotypes. Because of the small study population (n = 60), further statistical analysis of possible gene-gene and gene environment would not be informative. This is the first study analysing the specific BPDE-DNA adduct in vivo with regard to polymorphic repair genes (XPD, XRCC1) and xenobiotic metabolizing gene (GSTM1). Our results raise the possibility that the XPD-Asp312Asp+Lys751Lys genotype may increase BPDE-DNA damage; this effect might be evident in individuals who are especially likely to have accumulated damage, probably because of lower detoxification capacity and high environmental exposure. PMID- 12115581 TI - Differential production of angiostatin by concomitant antitumoral resistance inducing cancer cells. AB - The phenomenon by which tumor-bearing hosts are capable of inhibiting secondary tumor implants or metastases, known as concomitant antitumoral resistance (CAR), is presumably due to antiangiogenesis at places distant from the primary tumor. Although angiostatin, a potent inhibitor of angiogenesis, has been reported to be one of the factors responsible for suppressing the growth of secondary tumors in mice bearing previous tumors, it has not been definitively proven yet. With the aim of investigating whether CAR-inducing cancer cells display a differential angiostatin production and to support the role ascribed to that molecule concerning the inhibition of secondary tumor implants, 5 tumor models with different CAR-inducing capacities were studied herein. One of the 2 human lung cancer cell lines analyzed revealed a strong CAR against secondary s.c. tumor implants in nude mice, and 2 of 3 of the murine mammary tumors used exhibited inhibitory effect on secondary s.c. and i.v. tumor inoculations in syngeneic hosts. Since angiostatin is a proteolytic fragment from plasminogen, we examined by Western blot the ability of all conditioned media collected from the tumor cells studied to convert plasminogen to angiostatin. An association between in vivo generation of CAR and in vitro conversion of plasminogen into angiostatin was found. Since different enzymatic mechanisms were described to explain the generation of angiostatin, we also studied gelatinase and urokinase-type plasminogen activator secretion in conditioned media by zymography. The conversion of plasminogen into angiostatin by conditioned media was mainly inhibited by broad-spectrum serine proteinase inhibitors, suggesting a possible role for 1 or more enzymes of that group in the process. These findings suggest the existence of a differential angiostatin generation by CAR-inducing cancer cells, providing additional support to previous data obtained by other authors. PMID- 12115582 TI - Novel camptothecin derivative BNP1350 in experimental human ovarian cancer: determination of efficacy and possible mechanisms of resistance. AB - The novel camptothecin derivative BNP1350 (7-[2-trimethylsilyl)ethyl]-20(S) camptothecin), also known as Karenitecin, has been developed for superior oral bioavailability and increased lactone stability. In our study, we describe the antiproliferative effects of BNP1350, SN-38 and topotecan in 4 human ovarian cancer cell lines. BNP1350 was found to be slightly more potent than SN-38 (p<0.01) and was considerably more potent than topotecan (p<0.01). We extended these studies to well-established human ovarian cancer xenografts in which we compared the growth inhibition induced by BNP1350 with that of topotecan given in equitoxic schedules. The growth inhibition in all 3 xenografts induced by BNP1350 was > or =75%, which was significantly better than that resulting from topotecan (p<0.05). We then selected BNP1350-resistant variants of the A2780 human ovarian cancer cell line, 2780K4 (resistance factor: 41) and 2780K32 (resistance factor: 90), to analyze possible resistance mechanisms. These variants exhibited cross resistance against all camptothecins tested. In comparison with 2780K4 cells, 2780K32 cells were relatively more resistant against SN-38, topotecan, DX-8951f and BNP1350. In addition, 2780K32 cells were highly cross-resistant against mitoxantrone. In both 2780K4 and 2780K32, the amount of topoisomerase I was not changed but the catalytic activity was reduced. Furthermore, 2780K32 cells clearly overexpressed the breast cancer resistance protein (BCRP), as demonstrated for both the gene and the protein. In contrast to topotecan, BNP1350 proved not to be a good substrate for BCRP. Overall, we conclude that BNP1350 offers advantages over topotecan expressed by high efficacy in experimental human ovarian cancer and poor affinity for BCRP. PMID- 12115583 TI - Distinct in vivo expression patterns of survivin splice variants in renal cell carcinomas. AB - Survivin, a novel member of the inhibitor of apoptosis protein (IAP) family, reduces the susceptibility of tumor cells to proapoptotic stimuli, thereby promoting tumor cell survival during tumor progression and treatment with anticancer drugs. Recently, we identified 2 novel alternative splice variants of survivin, survivin-2B and survivin-Delta Ex3, which differ in their antiapoptotic properties. Survivin-2B has lost its antiapoptotic potential and may act as a naturally occurring antagonist of antiapoptotic survivin and survivin-Delta Ex3. Because the in vivo expression of these splice variants in human cancer has not been analyzed so far, 57 renal cell carcinomas (RCCs) were explored using quantitative reverse transcriptase polymerase chain reaction. We found that all RCCs express survivin-Delta Ex3, survivin-2B and survivin, the latter being the dominant transcript. When we compared early and intermediate stages with late stages of clear cell RCCs, no significant changes in the expression levels of survivin and survivin-Delta Ex3 became evident. However, a significant decrease was observed for the mRNA ratio between survivin-2B and survivin in late tumor stages (p = 0.036). Chromophilic/papillary RCCs, which are known to be less aggressive than clear cell RCCs, did not show significantly lower expression levels of antiapoptotic survivin and survivin-Delta Ex3, compared with stage adjusted clear cell RCCs. Our study demonstrates for the first time in vivo expression of functionally different survivin variants and suggests a role of these survivin splice variants in the progression and clinical behavior of human RCCs. PMID- 12115584 TI - Involvement of human heat shock protein 90 alpha in nicotine-induced apoptosis. AB - There have been conflicting reports of the apoptotic effects of nicotine on human cells and those studies reporting nicotine-induced apoptosis have not unequivocally clarified the molecular mechanisms underlying the effect. However, we found here that human RSa cells, established from embryonic fibroblastic cells doubly infected with Rous sarcoma virus and Simian virus 40, underwent apoptosis when cultured with medium containing 0.06-0.6 microM nicotine. The apoptosis was assessed by cellular DNA fragmentation and caspase-3 protease activation. Viability of RSa cells was reduced by nicotine treatment, as analyzed by MTT assay and the reduction was lessened by combination treatment with a caspase-3 inhibitor, acetyl-L-aspartyl-L-glutamyl-L-valyl-L-aspart-1-al (Ac-DEVD-CHO). Levels of expression of heat shock protein 90 alpha (Hsp90 alpha) were found to be increased 20 min after the nicotine treatment, as analyzed by polymerase chain reaction-based mRNA differential display after Northern blotting analysis of mRNA amounts. Cellular contents of Hsp90 alpha were furthermore increased in the nicotine-treated RSa cells, as quantitated by Western immunoblot analysis. By contrast, in RSa cells treated with nicotine in combination with geldanamycin (GA), an inhibitor of Hsp90 alpha function, DNA fragmentation was not detected and caspase-3 protease activity levels were the same as those of mock-treated cells. Nicotine-induced caspase-3 activation and Hsp90 alpha expression, as well as suppression of the induction by GA, were also observed in a xeroderma pigmentosum patient-derived cell line, XP2OS cells. Thus, it was suggested that nicotine induces apoptosis, possibly via Hsp90 alpha expression, in human cells tested. PMID- 12115585 TI - Farnesyltransferase inhibitor, R115777, reverses the resistance of human glioma cell lines to ionizing radiation. AB - We investigated for the first time the ability of farnesyltransferase inhibitors (FTI) to radiosensitize human glioma. For this, human glioma cell lines were treated with the specific FTI, R115777, 48 hr prior to a 2Gy irradiation. The treatment with R115777 decreased by 45% the SF2 value of the more radioresistant glioma cell lines (SF763 and U87) without any significant effect on the radioresistance of the radiosensitive ones (SF767 and U251-MG). This radiosensitizer effect was due to the induction of post-mitotic necrotic cell death. We then tested the hypothesis that wild-type Ras or RhoB, which has been proposed as potential FTI target, could control the glioma radioresistance. For this, we expressed inducible dominant negative forms of Ras (RasN17) and RhoB (RhoBN19) in radioresistant U87 glioma cell line and analyzed the survival after irradiation of the obtained clones. While blocking Ras pathways by expression of RasN17 did not affect the SF2 value of the U87 glioma cell line, the expression of RhoBN19 dramatically reduced the cell survival after irradiation of these cells. Taken together, these data demonstrated that RhoB, but not Ras, is implicated in glioma radioresistance. Furthermore, the R115777 differential radiosensitizer effect underlines the potential therapeutic interest of using this drug as a radiosensitizer of human glioma. PMID- 12115586 TI - Expression and functional analysis of the anaplastic lymphoma kinase (ALK) gene in tumor cell lines. AB - The initial identification of the ALK gene, expressed as C-terminal part of the transforming fusion protein NPM-ALK in the t(2;5)(p23;q35) lymphoma-associated chromosomal translocation, revealed a novel receptor tyrosine kinase (RTK). In order to expand the knowledge on ALK expression in the human system, we examined a panel of human cell lines for ALK expression and found that transcription is completely repressed in cell lines of entodermal origin (0/21). Furthermore, full length receptor expression is absent in cell lines of the hematopoietic system with the exception of t(2;5)-associated anaplastic large cell lymphomas lines (ALCL), which are known to express chimeric NPM-ALK mRNA. Cell lines established from solid tumors of ectodermal origin, including melanoma and breast carcinoma, exhibited widespread mRNA expression of the ALK receptor at a broad range (53/64), an association which was found to be strongest in cell lines derived from neuroblastoma (6/6), glioblastoma (8/8) as well as in cell lines established from Ewing sarcoma (4/4) and retinoblastomas (2/2). Because of the reported involvement of neutrophin tyrosine kinase receptors in autocrine differentiation in neuroblastomas, we analyzed cell lines positive for full length or chimeric ALK protein for the presence of phoshotyrosine residues within the intracellular region of ALK. While the constitutive activation of chimeric NPM-ALK molecules could be shown, no evidence was found for induced or constitutively activated ALK receptors in neuroblastoma, melanoma or breast carcinoma cell lines. Although the receptor could be shown to be consistently expressed with exclusive specificity in tissues developed from the ectoderm, our results do not support any involvement of ALK in the stimulation of tumorigenic cell growth or differentiation so far, indicating that ALK expression is a physiologic rather than a pathologic phenomenon. PMID- 12115587 TI - Reduced expression of the insulin-induced protein 1 and p41 Arp2/3 complex genes in human gastric cancers. AB - Aberrantly methylated DNA fragments in a human gastric cancer were searched for by a genome-scanning method, methylation-sensitive-representational difference analysis (MS-RDA). Six DNA fragments flanked by CpG islands (CGIs) and hypermethylated in the cancer were isolated. Four of the 6 fragments possessed genes in their vicinities. Quantitative RT-PCR analysis of the 4 genes showed reduced expression of 2 genes in cancers: Insulin-induced protein 1 (INSIG1/CL-6) and p41 Arp2/3 complex (p41-Arc). As for INSIG1, a DNA fragment was derived from the edge of a CGI in the promoter region. The edge was methylated in 11 of 22 primary gastric cancers, whereas the center was not methylated in any cancer. INSIG1 expression was markedly reduced in 19 cancers, including the 11 cancers with the methylation. By 5-aza-2'-deoxycytidine treatment of 5 cell lines with the methylation of the edge, partial restoration of INSIG1 expression was detected only in 2 of them. These data indicated that, although the reduced INSIG1 expression in cancers was associated with the methylation at the edge of the CGI in the promoter region, the methylation was likely to be a secondary change. As for p41-Arc, a DNA fragment was derived from a CGI overlapping exon 8, and its methylation did not correlate with its expression. However, methylation of a CGI in the promoter region with a marked reduction of its expression was observed in 1 of the 22 primary cancers. INSIG1 and p41-Arc are known to be involved in cellular differentiation and morphology, respectively, and it was suggested that their reduced expressions might be involved in gastric cancer development or progression. PMID- 12115588 TI - MICA triggering signal for NK cell tumor lysis is counteracted by HLA-G1-mediated inhibitory signal. AB - MICA, a highly glycosylated membrane-anchored cell-surface MHC Class I-related chain, has recently been reported to activate NK cell cytolytic responses in epithelial tumors. Tumor cells may escape from NK lysis by counteracting NK cytotoxicity activating signals with inhibitory ones. Among the molecules that mediate an NK inhibitory signal, HLA-G1, a non-classical MHC Class I antigen, is of particular interest. HLA-G1 is ectopically expressed in various tumors, including melanoma and constitutes the major NK inhibitory ligand in the M8 melanoma cell line when coexpressed with HLA-A, -B, -C and -E molecules. We have evaluated the balance between 2 powerful signals that affect NK cell tumor lysis, one inhibitory and the other one activating, respectively HLA-G1 and MICA. For this purpose, we transfected the M8 melanoma cell line, which spontaneously expresses MICA, with HLA-G1 cDNA, using it as a target for the NKL effector. We carried out cytotoxicity assays, using antibodies that disrupt interactions between the MICA and HLA-G1 ligands and their respective NK effector counterparts, the NKG2D activating and ILT2 inhibitory receptors. Results showed that 1) MICA expressed in the M8 melanoma cell line triggered NK cell tumor lysis and 2) HLA-G1 coexpression mediated the inhibition of NK cytotoxicity by mitigating the MICA activating signal. HLA-G1 expression in a tumor cell line in which MICA is switched on would therefore appear to be a powerful way to turn off NK cells, supporting the emerging idea that the balance between positive and negative NK cytolysis signals critically influences tumor progression. PMID- 12115589 TI - Urinary level of 1,N(6) -ethenodeoxyadenosine, a marker of oxidative stress, is associated with salt excretion and omega 6-polyunsaturated fatty acid intake in postmenopausal Japanese women. AB - Excretion of 1,N(6)-ethenodeoxyadenosine (epsilon dA), a marker for lipid peroxidation (LPO)-derived DNA damage was analyzed in urine of nonsmoking postmenopausal women participating in a dietary intervention trial in Northern Japan. Hereby the efficacy of dietary consultation in reducing salt and increasing vitamin C and carotenes during 1 year was estimated. Thirty postmenopausal women, 60-69 years of age, from the intervention group and 30 age matched women from the control group were randomly selected. The subjects completed a self-administered diet history questionnaire and in the pre- and post intervention period 48 hr urine and fasting blood samples were collected. epsilon dA in urine was analyzed by an immuno-precipitation-high performance liquid chromatography-fluorescence detection method. epsilon dA excretion (/48 hr) in the 59 postmenopausal Japanese women with complete urine collection ranged from 12-226 pmol at the pre-intervention. At the pre-intervention, epsilon dA excretion was positively associated with urinary salt excretion (R = 0.33, p = 0.01) and omega-6 polyunsaturated fatty acid intake (%energy value, R = 0.28, p = 0.03) in the 59 women. The average epsilon dA excretion in the intervention group was 61 pmol at pre-intervention and 44 pmol at post-intervention (p = 0.14). In the control group, it was 58 pmol at pre-intervention and 75 pmol at post intervention (p = 0.24). During the intervention period, 18/29 (62%) of the subjects in the intervention group exhibited the decreased excretion and 10/26 (38%) in the control group (p = 0.08). Results from this pilot study suggest urinary epsilon dA as a potential biomarker of DNA damage possibly derived from salt-induced inflammation and LPO; further exploration of epsilon dA in human biomonitoring studies is warranted. PMID- 12115590 TI - Physical activity and risk for breast cancer a prospective cohort study among Swedish twins. AB - The epidemiologic association between physical activity and breast cancer has been corroborated in many studies. Some inconsistencies remain, possibly due to variation in life periods for exposure assessment, confounding and undetected effect modification. In our cohort study, we address some of these questions by taking into account physical activity in different periods of life and by investigating effect modification by birth cohort and body mass index (BMI). Altogether 9,539 same-sex twin women aged 42-70 years who answered a questionnaire about their work and leisure-time physical exercise from ages 25 to 50 during 1967 and 1970 were included in our cohort. During follow-up, 506 breast cancer cases occurred through 1997. We used multivariate Cox models to estimate relative risk (RR) with 95% confidence interval (CI). We found no associations between physical activity and breast cancer overall. Women born between 1901 and 1917 (aged 51-70 at baseline) who reported regular leisure-time activity had a borderline significant 40% lower risk compared with those who reported no activity (RR 0.6; 95% CI 0.4-1.0; test for trend, p = 0.07). This association appeared to be confined to women with a low BMI after the age of 50 and to women with a high BMI during the premenopausal period. We found no evidence that work activity reduces risk for breast cancer. The importance of physical activity for breast cancer risk seems to depend on birth cohort. The association may be limited to normal-weight postmenopausal women and overweight premenopausal women. PMID- 12115591 TI - Cancer incidence in a population-based cohort of patients with Wegener's granulomatosis. AB - Wegener's granulomatosis is necrotizing granulomatous vasculitis of unknown origin, which untreated has a high mortality within the first year of onset. The introduction of corticosteroids and cyclophosphamide in the treatment has considerably improved survival rates, but past studies have indicated an increased cancer risk, including an increased risk for urinary bladder cancer. No large assessment of the general cancer occurrence in Wegener's granulomatosis has been reported. The aim of our study was to assess the general incidence of cancer in patients with Wegener's granulomatosis and to put this in relation to the risk for bladder cancer. We identified a population-based cohort of 1,065 patients with Wegener's granulomatosis in the Swedish In-patient Register. Through linkage with the Swedish Cancer Register, we followed the cohort for cancer occurrence for up to 26 years. Standardized incidence ratios (SIR) between observed and expected numbers of cancers were used as a measure of relative risk. There was a 2-fold overall increased risk for cancer in the cohort. The increase was most pronounced for bladder cancer (SIR = 4.8; 95% CI 2.6-8.1), squamous cell skin cancer (SIR = 7.3; 95% CI 4.4-12), leukemias (SIR = 5.7; 95% CI 2.3-12) and for malignant lymphomas (SIR = 4.2; 95% CI 4.2-8.3). The results confirm previous indications of an increased risk for cancer of the urinary bladder but also points to increased risks for cancer at other sites. PMID- 12115592 TI - Cancer patterns in eastern India: the first report of the Kolkata cancer registry. AB - There are no population-based data available for the cancer patterns in Eastern India. This is the first report of cancer incidence in the region from the population-based cancer registry in Kolkata (Calcutta), the capital city of the state of West Bengal, India, for the period 1998-1999. The cancer registry collects data on all new cases of cancer diagnosed in the resident population of Kolkata. Since cancer is not a notifiable disease in India, registration is carried out by active data collection by the registry staff. The cancer registry staff visits 50 data sources comprising cancer hospitals, secondary and tertiary care hospitals, nursing homes, diagnostic laboratories and death registration offices; scrutinizes medical records and collects details on incident cancer cases. A customized version of CanReg-3 software was used for data entry and analysis. A total of 11,700 cases were registered during the 2-year period from 1 January 1998 to 31 December 1999. The overall age-adjusted (world population) incidence rates were 102.1 per 100,000 males and 114.6 per 100,000 females. The most frequently reported malignancies in males were lung cancer (16.3%), followed by cancers of the oral cavity (7.1%), pharynx (5.7%) and larynx (5.7%). In females, the most frequently reported malignancies were breast (22.7%) followed by uterine cervix (17.5%), gallbladder (6.4%) and ovary (5.8%). The data reported by the Kolkata cancer registry provide information on the cancer profile in Eastern India for the first time. The highest incidence rate of lung cancer in males in India is reported from Calcutta. A high risk of gallbladder cancer is observed in women. The observed cancer patterns indicate that tobacco-control measures and early detection of head and neck, breast and cervical cancers are of importance for cancer control in this population. PMID- 12115593 TI - Evaluation of downstaging in the detection of cervical neoplasia in Kolkata, India. AB - Unaided visual inspection or "downstaging" has been suggested as a potential alternative method for cervical cancer screening in developing countries. Our study was designed to evaluate the accuracy of downstaging to detect cervical neoplasia in a low-resource setting. A total of 6,399 women aged 30-64 years were screened with downstaging by trained nonmedical health workers. Two thresholds were used to define positive downstaging: "low threshold" when any visible abnormality on the cervix was considered positive and "high threshold" when selected abnormalities such as bleeding on touch, bleeding erosion, hypertrophied oedematous cervix, congested stippled cervix and growth or ulcer constituted the positive test. All women underwent a colposcopy examination. Biopsies were directed when colposcopy revealed abnormal lesions. True disease status was defined as histologically proven moderate dysplasia and worse lesions. Since all the participants received a diagnostic (reference) investigation (biopsy and/or colposcopy), sensitivity, specificity and predictive values were estimated directly. Low- and high-threshold downstaging were positive in 1,585 (24.8%) and 460 (7.2%) women, respectively. The sensitivities of low- and high-threshold downstaging to detect high-grade precursors and invasive cancers were 48.9% and 31.9%, respectively. The specificities were 75.8% and 93.3%, respectively. These results indicate that downstaging is not suitable as an independent primary screening modality for cervical neoplasia. PMID- 12115594 TI - Covariates and confounding in epidemiologic studies using metabolic gene polymorphisms. AB - The relationship between exposure and disease when biomarkers are introduced in an epidemiologic study is explored and summarized. In molecular epidemiologic studies, biologic measurements play a major role as markers of exposure, disease or susceptibility to disease and/or exposure. In this scenario, the definition and management of confounding factors may change. Sometimes the presence or activation of the biomarker is partially caused by the relevant environmental exposure, and therefore the 2 variables (exposure and biomarker) should not be always treated as confounders of each other. Models of exposure-disease association in the presence of biologic markers are presented. The concept of confounders is reviewed in light of the role of biomarkers in the pathway between exposure and disease. PMID- 12115595 TI - In vitro and in vivo activity of MT201, a fully human monoclonal antibody for pancarcinoma treatment. AB - In our study, a novel, fully human, recombinant monoclonal antibody of the IgG1 isotype, called MT201, was characterized for its binding properties, complement dependent (CDC) and antibody-dependent cellular cytotoxicity (ADCC), as well as for its in vivo antitumor activity in a nude mouse model. MT201 was found to bind its target, the epithelial cell adhesion molecule (Ep-CAM; also called 17-1A antigen, KSA, EGP-2, GA733-2), with low affinity in a range similar to that of the clinically validated, murine monoclonal IgG2a antibody edrecolomab (Panorex(R)). MT201 exhibited Ep-CAM-specific CDC with a potency similar to that of edrecolomab. However, the efficacy of ADCC of MT201, as mediated by human immune effector cells, was by 2 orders of magnitude higher than that of edrecolomab. Addition of human serum reduced the ADCC of MT201 while it essentially abolished ADCC of edrecolomab within the concentration range tested. In a nude mouse xenograft model, growth of tumors derived from the human colon carcinoma line HT-29 was significantly and comparably suppressed by MT201 and edrecolomab. The fully human nature and the improved ADCC of MT201 with human effector cells will make MT201 a promising candidate for the clinical development of a novel pan-carcinoma antibody that is superior to edrecolomab. PMID- 12115596 TI - Lovastatin potentiates antitumor activity of doxorubicin in murine melanoma via an apoptosis-dependent mechanism. AB - Lovastatin, a drug successfully used in the clinic to prevent and to treat coronary heart disease, has recently been reported to decrease the incidence of melanoma in lovastatin-treated patients. Lovastatin has also been proved to potentiate antitumor effects of both cisplatin and TNF-alpha in murine melanoma models. Recently, an augmented therapeutic effect of lovastatin and doxorubicin has been reported in 3 tumor models in mice. In our preliminary study lovastatin caused retardation of melanoma growth in mice treated with doxorubicin (Feleszko et al. J Natl Cancer Inst 1998;90:247-8). In the present report, we supplement our preliminary observations and demonstrate in 2 murine and 2 human melanoma cell lines that lovastatin effectively potentiates the cytostatic/cytotoxic activity of doxorubicin in vitro via an augmentation of apoptosis (estimated with PARP-cleavage assay, annexin V assay and TUNEL). The combined antiproliferative activity of lovastatin and doxorubicin was evaluated using the combination index (CI) method of Chou and Talalay, revealing synergistic interactions in melanoma cells exposed to lovastatin and doxorubicin. In B16F10 murine melanoma model in vivo, we have demonstrated significantly increased sensitivity to the combined treatment with both lovastatin (5 mg/kg for 14 days) and doxorubicin (4 x 1 mg/kg) as compared with either agent acting alone. Lovastatin treatment resulted also in significant reduction of the number of experimental metastasis in doxorubicin-treated mice. The results of our studies suggest that lovastatin may enhance the effectiveness of chemotherapeutic agents in the treatment of malignant melanomas. PMID- 12115597 TI - Treatment of peripheral blood with staurosporine increases detection of circulating carcinoembryonic antigen positive tumor cells. PMID- 12115600 TI - Hyperosmotic stimuli inhibit VCAM-1 expression in cultured endothelial cells via effects on interferon regulatory factor-1 expression and activity. AB - Transcriptional up-regulation of the VCAM-1 gene, induced by proinflammatory cytokines such as IL-1beta and TNF-alpha, requires activation of not only NF kappaB, but also involves interferon regulatory factor (IRF)-1. During a study of gene induction by mechanical stimuli in cultured human endothelial cells, we noted that medium hyperosmolarity appeared to influence cytokine-induced expression of VCAM-1. Indeed, addition of hyperosmotic, pathophysiologically relevant concentrations of NaCl effectively inhibited IL-1beta or TNF-alpha induction of VCAM-1, but not E-selectin, at the level of mRNA and cell surface protein. Because induction of both VCAM-1 and E-selectin by these cytokines is NF kappaB dependent, we investigated whether the inhibitory effect of hyperosmotic medium might involve IRF-1. Electrophoretic mobility shift assays of the VCAM-1 promoter demonstrated that hyperosmotic medium suppressed IL-1beta- or TNF-alpha activated binding activities of IRF-1, but not NF-kappaB, to their respective sites. Hyperosmotic medium also inhibited the expression of IRF-1 induced by TNF alpha or IFN-gamma. Furthermore, hyperosmotic medium inhibited TNF-alpha or IFN gamma induction of guanylate binding protein-1, another IRF-1-dependent gene. Taken together, hyperosmolarity selectively inhibits cytokine-induced VCAM-1 in endothelial cells, via an IRF-1-dependent mechanism. Thus, pathophysiological fluctuations in plasma osmolarity may influence certain endothelial-dependent components of the inflammatory response and host defense mechanisms. PMID- 12115601 TI - The impact of c-met/scatter factor receptor on dendritic cell migration. AB - Dendritic cells (DC) are professional antigen-presenting cells that possess both migratory properties and potent T cell stimulatory activity, and that allow the uptake of antigenic material inperipheral tissues and its subsequent presentation in the T cell areas of lymphoid organs. Thus motility represents a central property that is required for DC function. Here we report on the expression of the receptor tyrosine kinase c-met in DC. c-Met is the high affinity receptor for scatter factor (SF)/hepatocyte growth factor, and ligand-activated c-met exhibits mitogenic, morphogenic andmotogenic activity in vivo and in vitro. c-Met is signaling competent in DC since it is effectively tyrosine phosphorylated in response to SF ligand. It is demonstrated here that ligand-activated c-met regulates DC adhesion to the extracellular matrix component laminin but leaves antigen presenting function unaffected. Importantly, in ear sheet explant experiments activationof c-met by ligand induces emigration of cutaneous DC (Langerhans cell, LC) from skin, but SF is not a chemoattractant factor for DC. Our results suggest an important role of the c-met/SF system in DC/LC migration. PMID- 12115602 TI - Stability of the B cell antigen receptor modulates its signaling and antigen targeting functions. AB - The binding of antigens to the B cell antigen receptor (BCR) results in the initiation of signaling cascades and the internalization of the antigens for processing and presentation. Recent studies indicate that antigen binding destabilizes the BCR as a mechanism to down-regulate B cell responses. Two point mutations in the transmembrane domain of murine membrane IgM (mIgM) (YS to VV) weaken the interaction of mIgM with Igalpha/Igbeta heterodimer, resulting in a destabilized BCR. Using muYS/VV BCR, effects of the destabilized BCR on the functions of an endogenous wild-type mIgG(2a) BCR were analyzed. The muYS/VV BCR is defective in signaling and does not target antigens to late endocytic compartments for processing and presentation. Coligation of the muYS/VV BCR with the endogenous BCR interferes with antigen-targeting functions of the endogenous BCR. Thus, the destabilized BCR has a dominant effect, down-regulating the function of stable wild-type BCR. The ability of the destabilized BCR to influence the stable BCR may play an important role in turning off B cell responses for antigen-driven anergy and tolerance. PMID- 12115603 TI - The anti-apoptotic molecules Bcl-xL and Bcl-w target protein phosphatase 1alpha to Bad. AB - Bcl-xL and Bcl-w specifically interact with PP1alpha and Bad. A phosphatase activity sensitive to okadaic acid was detected in Bcl-xL, Bcl-w and Bad immunoprecipitates. Serine phosphorylation of Bcl-xL and Bcl-w correlates with the number of trimolecular complexes formed. Depletion of Bcl-xL and Bcl-w decreases the remaining Bad-associated phosphatase activity and association of protein phosphatase 1 (PP1)alpha to Bad. Bcl-xL and Bcl-w contain the R/K X V/I X F consensus motif shared by PP1 targeting subunits. This motif, in addition to F X X R X R motif, is involved in binding of Bcl-xL and Bcl-w to PP1alpha. Disruption of Bcl-xL/PP1alpha or Bcl-w/PP1alpha association strongly decreases Bad-associated phosphataseactivity and stability of trimolecular complexes. These results suggest that Bcl-xL and Bcl-w are PP1alpha targeting subunits and this trimolecular complex may be involved in the control of apoptosis. PMID- 12115604 TI - Sphingosine 1 phosphate induces the chemotaxis of human natural killer cells. Role for heterotrimeric G proteins and phosphoinositide 3 kinases. AB - We have examined the effect of sphingolipids on the chemotaxis of human natural killer (NK) cells. Messenger RNA for Edg-1, Edg-6 and Edg-8 but not Edg-3, are expressed in these cells. Sphingosine 1 phosphate (SPP), dihydro SPP (DHSPP) or the CC chemokine RANTES (CCL5), but not sphingosine induces the chemotaxis of these cells. Pertussis toxin inhibits the chemotaxis induced by these ligands. Permeabilization of NK cells with streptolysin O (SLO) and introduction of blocking antibodies to the heterotrimeric G proteins, showed that Galpha(i2), Galpha(s), Galpha(q/11) or Galpha(13) mediate the chemotaxis of SPP, whereas Galpha(i2), Galpha(o) or Galpha(q/11) mediate the chemotaxis of DHSPP. Galpha(i2), Galpha(o), Galpha(s), Galpha(q/11), Galpha(z) or Galpha(12 )mediates RANTES-induced NK cell chemotaxis. Further analysis showed that phosphoinositide 3 kinase (PI3K) inhibitors wortmannin and LY294002 inhibit NK cell chemotaxis induced by SPP, DHSPP or RANTES. Blocking antibody to PI3Kgamma inhibits the chemotaxis induced by the three ligands, whereas anti-PI3Kbeta was without effect. In contrast, SPP and DHSPP recruit PI3Kbeta isozyme into NK cell membranes, suggesting that although this isoform is not involved in chemotaxis, it is activated by these phospholipids. PMID- 12115605 TI - MyD-1 (SIRPalpha) regulates T cell function in the absence of exogenous danger signals, via a TNFalpha-dependent pathway. AB - Signal inhibitory regulatory proteins are a family of transmembrane glycoproteins involved in the negative regulation of receptor tyrosine kinase signaling pathways. MyD-1 is a recently described member of this family. In this report we have assessed the function of MyD-1 in regulating T cell function utilizing an anti-MyD-1-specific monoclonal antibody (mAb). We show that ligating MyD-1 on antigen-presenting cells (APC) inhibits subsequent T cell activation induced by either anti-CD3 mAb or by allogeneic APC but has no effect on responses to either tuberculin purified protein derivative or tetanus toxoid antigens. Moreover, we show that the inhibition of T cell responses is not due to any change of costimulatory molecule expression on APC. We observed that the production of TNFalpha, a cytokine that we have earlier identified as important in the mechanism of MyD-1 immune regulation, is inhibited by cross-linking of MyD-1. We further show that TNFalpha is critically important in the regulation of T cell responses in the absence of danger signals, and indeed addition of TNFalpha can overcome the inhibitory effects of anti-MyD-1 antibody. This information may lead to a better understanding of the regulation of T cell responses in the absence of danger and therefore offer a possible therapeutic target to modulate aberrant immune responses. PMID- 12115607 TI - Increased circulating CD11b+CD11c+ dendritic cells (DC) in aged BWF1 mice which can be matured by TNF-alpha into BLC/CXCL13-producing DC. AB - Dendritic cells (DC) play a pivotal role in regulating immune responses. We previously reported aberrant high production of B lymphocyte chemoattractant (BLC/CXCL13) by DC in aged BWF1 mice, amurine model for systemic lupus erythematosus (SLE). We describe here that CD11b+CD11c+ cells were markedly increased in the peripheral blood (PBL-DC) in aged BWF1, but not in similarly aged NZB or NZW mice. Part of PBL-DC showed a typical dendritic morphology and expressed MHC class II molecules, and had a weak, but significant antigen presenting ability in mixed lymphocytereaction. PBL-DC were chemoattracted to several chemokines in vitro including secondary lymphoid tissue chemokine (SLC), liver and activation-regulated chemokine (LARC), RANTES, macrophage inflammatory protein-1alpha, whereas splenic mature DC from aged BWF1 mice were preferentially chemoattracted towards SLC. BLC production was induced when PBL-DC were cultured in the presence of TNF-alpha for 3 days. BLC expression was also induced in bone marrow-derived DC when they were differentiated into mature DC in the presence of TNF-alpha and IL-1beta, while both IFN-alpha and IFN-gamma failed to induce BLC expression in bone marrow-derived DC. Since TNF-alpha expression is increased in aged BWF1 mice, DC recruitment in the circulation and maturation into BLC producing DC by TNF-alpha may play a pivotal role in the development of systemic autoimmune diseases. PMID- 12115606 TI - Bone marrow CD8 T cells are in a different activation state than those in lymphoid periphery. AB - In addition to its well-established use for hematopoiesis reconstitution, bone marrow is considered with increasing interest as a possible source of mature cells for adoptive therapies, in particular for the immunotherapy of cancer. Nevertheless, the peculiarities of bone marrow T cells in comparison with those in lymphoid periphery are still largely unknown. In this report, we show for the first time that bone marrow CD8 T cells are in a different activation state than those in peripheral lymphoid organs. Firstly, we observed that mouse bone marrow contains a significantly higher percentage of blasts within the CD8 T cells than either spleen or lymph nodes, yet such enrichment is not due to recent antigenic stimulation. Secondly, when we challenged bone marrow CD8 T cells from immunized mice with their antigen in vitro, they displayed a faster response than those from the spleen. Thus, we suggest that the bone marrow could be a preferential source of CD8 T cells for adoptive therapies in those cases in which highly active effectors are required. PMID- 12115608 TI - The follicular dendritic cell restricted epitope, FDC-M2, is complement C4; localization of immune complexes in mouse tissues. AB - We have identified the murine follicular dendritic cell (FDC) marker, FDC-M2, recognized by monoclonal antibody mAb209, as complement component C4. Consistent with this, FDC-M2 was detectable at sites of immune complex-mediated inflammatory disease. Analysis of FDC-M2 distribution in complement-deficient mice highlighted the differences in immune complex clearance between these mice, andshowed that a population of uncharacterized FDC-M2+ reticular and perivascular cells in the spleen, distinct from FDC, are also involved in immune complex capture and possibly retention. These results demonstrate that mAb209, in addition to its role as an FDC marker, is a valuable reagent for the analysis of complement deposition in vivo. PMID- 12115609 TI - Ligand-activated natural killer T lymphocytes promptly produce IL-3 and GM-CSF in vivo: relevance to peripheral myeloid recruitment. AB - Natural killer (NK) T cells are prominent for their prompt IL-4 and IFN-gamma production upon TCR ligation that enables them to influence acquired immune responses. In the present study we provide evidence that the regulatory functions of this particular T cell subset extend to the myeloid compartment of bone marrow and spleen through its production of hematopoietic growth factors. Bone marrow and spleen NKT cells responded to a single injection of their specific ligand alpha-galactosylceramide (alpha-GalCer) by producing both IL-3 and granulocyte macrophage colony stimulating factor (GM-CSF), whose colony-stimulating activity became detectable in the serum as early as 1 h post treatment. These cytokines were not produced in mice lacking NKT cells (CD1d-/-), whose exclusive involvement in this biological activity was further confirmed by intracellular immuno-staining. Growth factor production was accompanied by significant changes in the myeloid compartment of treated mice, namely mobilization of myeloid progenitors (colony-forming unit cells, CFU-C) and neutrophils from the bone marrow to the periphery. Taken together, our data support the notion that activated NKT cells influence innate immune responses by recruiting myeloid progenitors and granulocytes to the periphery through their production of hematopoietic growth factors. PMID- 12115610 TI - Critical role of antigen-specific antibody in experimental autoimmune encephalomyelitis induced by recombinant myelin oligodendrocyte glycoprotein. AB - The role of B cells and antibody in the pathogenesis of experimental autoimmune encephalomyelitis (EAE) remains controversial. We previously demonstrated that B cells are required for EAE to be induced by the 120-amino acid extracellular domain of myelin oligodendrocyte glycoprotein (MOG). In the present study, the role of B cells in MOG-induced EAE was further characterized. Passive transfer of activated B cells or serum from MOG-primed wild-type (WT) mice was found to reconstitute the ability for clinical and histological EAE to be induced in MOG immunized B cell-deficient mice. MOG-induced EAE did not occur with transfer of B cells that had been nonspecifically activated by lipopolysaccharide or isolated from naive or myelin basic protein (MBP)-primed WT mice. Likewise, MOG-primed serum, but not naive serum or serum from MBP-, Hen egg lysozyme-, or MOG(35-55) primed mice, led to EAE in B cell-/- animals. While both MOG-primed B cells and serum reconstituted the ability for disease induction, MOG-primed serum was much more efficient, leading to clinical and histological EAE similar to that seen in the WT. Injection of MOG serum into healthy B cell-/- mice 30 days after MOG immunization led to rapid appearance of clinical signs and CNS inflammation, indicating that an antigen-specific factor is necessary for initiation of CNS inflammation,and not just demyelination. These data strongly suggest that MOG specific antibody is critical to the initiation of MOG-induced murine EAE. PMID- 12115611 TI - IFN-gamma-independent synergistic effects of IL-12 and IL-15 induce anti-tumor immune responses in syngeneic mice. AB - IL-15 and IL-12 display anti-tumor activity in different models and IFN-gamma has been reported as a secondary mediator of both IL-12 and IL-15 effects. TS/A murine adenocarcinoma cells were engineered to secrete IL-12, IL-15 or both cytokines. TS/A cells secreting IL-15 (TS/A-IL-15) displayed a reduced tumorigenicity (50%) when implanted subcutaneously in syngeneic mice, while both TS/A-IL-12 and TS/A-IL-12/IL-15 were rejected by 100% of animals. In contrast, TS/A-IL-15 and TS/A-IL-12 were tumorigenic in syngeneic IFN-gamma knockout mice (100% and >90% of take rate, respectively), but TS/A-IL-12/IL-15 were completely rejected by 90% of these mice. All IFN-gamma-deficient mice rejecting TS/A-IL 12/IL-15 developed protective immunity against wild-type TS/A, as indicated by re challenge experiments. Immunohistochemical analysis of the TS/A-IL-12/IL-15 tumor rejection area in IFN-gamma-deficient mice showed a marked reactive cell infiltration constituted of CD8+ cells, granulocytes, NK cells, macrophages and dendritic cells associated with the expression of IL-1beta, TNF-alpha, GM-CSF, MCP-1 and MIP-2. In vivo depletion experiments showed that rejection of TS/A-IL 12/IL-15 cells required CD8+ T lymphocytes and also involved granulocytes, while CD4+ and NK cells played a minor role. These data show IFN-gamma-independent synergistic anti-tumor effects of IL-12 and IL-15, involving CD8+ cells and secondary chemokines and cytokines, such as TNF-alpha. PMID- 12115612 TI - Comparison of CD22 binding to native CD45 and synthetic oligosaccharide. AB - The B cell surface molecule CD22 is a member of the Siglec family. Siglecs possess a conserved membrane-distal immunoglobulin domain that mediates binding to sialylated glycoproteins or glycolipids. Although the structural basis of sialic acid recognition by Siglecs is quite well understood, the binding properties of the interaction between Siglecs and their native ligands have not been investigated. CD22 binding requires alpha2-6-linked sialic acid, which is mostly carried on N-glycans. One protein that carries such N-glycans is CD45. In this study we used surface plasmon resonance to perform thermodynamic and kinetic analysis of CD22 binding to native CD45. CD22 bound with a low affinity (K(d) 130 microM at 25 degrees C) and very fast kinetics (k(off) >or=18 s(-1), calculated k(on) >or=1.5 x 10(5) M(-1)s(-1)). Van't Hoff analysis revealed that binding was enthalpically driven at physiological temperatures, as is typical of most lectin carbohydrate interactions. Since there is evidence that CD22 binds preferably to CD45, even though many cell surface proteins carry alpha2-6-linked sialic acid, we compared the affinities of CD22 binding to CD45, to CD4 carrying alpha2-6 linked sialic acid, and to a synthetic alpha2-6-sialoglycoconjugate. The affinities did not differ substantially, suggesting that CD22 binds preferentially to CD45 not because the latter presents higher affinity ligands but because it carries multiple copies of thereof. PMID- 12115613 TI - CCR4 in human allergen-induced late responses in the skin and lung. AB - We studied the regulation of CCR4 expression in peripheral blood and in human models of cutaneous and pulmonary allergen challenge. CCR4 expression was detectable on freshly isolated CD4+ lymphocytes and in CD4+ and CD8+ T cell lines derived from blood of atopic donors. Numbers of CCR4+ cells were up-regulated in T cell lines expanded in the presence of IL-4. CCR4 mRNA was absent at baseline in normal subjects in lung and skin, but present at baseline in the lung of some atopics. Baseline expression of CCR4 mRNA and protein was higher in lung vs. skin, but allergen-induced increases in CCR4 mRNA+ cells were observed in both organs. CCR4 protein+ cells were present at higher levels after allergen challenge in atopics compared to normal subjects. CCR4 may be important in the recruitment of T lymphocytes at sites of allergic inflammation, in a non-organ specific manner. PMID- 12115614 TI - A comparative analysis of B cell-mediated myelin oligodendrocyte glycoprotein experimental autoimmune encephalomyelitis pathogenesis in B cell-deficient mice reveals an effect on demyelination. AB - We have investigated the role of B cells in myelin oligodendrocyte glycoprotein (MOG)-induced experimental autoimmune encephalomyelitis (EAE) using B cell deficient mice muMT) and mice bearing the X-linked immunodeficiency (xid). The mice were immunized with MOG(1-125 )in complete Freund's adjuvant but without use of pertussis toxin. B cell-deficient muMT mice on different genetic backgrounds (C57BL/10 and DBA/1 strains) developed EAE, although with a reduced clinical severity. Histological analyses revealed decreased demyelination in the central nervous system while the influx of inflammatory cells was similar or only slightly reduced as compared to B cell-sufficient control mice. Xid mice on the DBA/1 background also developed disease with a reduced disease severity. The anti MOG antibody response in the xid mice was decreased, while the T cell response to MOG was unaffected. We thus demonstrate that B cells are not critical for the development of MOG-induced EAE but contribute to the severity. The contribution of B cells to pathogenesis appears to be mainly through demyelination rather than through inflammation. PMID- 12115615 TI - Age- and tissue-specific differences in human germinal center B cell selection revealed by analysis of IgVH gene hypermutation and lineage trees. AB - The elderly produce increased levels of antibodies to autologous antigens and are less able to make high-affinity antibodies to foreign antigens. Ig gene hypermutation is integral to the affinity maturation process but previous studies of hypermutation with age have yielded conflicting results. The cells studied have represented post-germinal center (GC) populations and, therefore, the results may be complicated by possible differences in activation history. We studied Ig genes from GC B cells to elucidate which factors in the affinity maturation process change with age. Age-related changes in the pattern of hypermutation were seen, although the analysis of variable region heavy chain (VH) genes and their lineage trees shows that an alteration in the mechanism of somatic hypermutation is unlikely. The changes are due to founder cell effects and/or the process of selection. Striking tissue-specific differences were seen. All measurements indicated that selection of Ig genes may decrease in Peyer's patch GC but increase in splenic GC with age. These tissue-specific differences highlight the importance of considering the activation and effector sites when studying immune senescence. PMID- 12115616 TI - Bacterial CpG-DNA and lipopolysaccharides activate Toll-like receptors at distinct cellular compartments. AB - Recognition by innate immune cells of the pathogen associated molecular patterns (PAMP) lipopolysaccharide (LPS) from Gram-negative bacteria and bacterial CpG-DNA depends on Toll-like receptor4 (TLR4) and TLR9, respectively. To define differences in the response to these distinct PAMP we compared a key intracellular event, namely recruitment of myeloid differentiation marker 88 (MyD88) to the respective PAMP-initiated TLR signaling. Using MyD88-GFP fusion protein expressing macrophages we demonstrate that LPS and CpG-DNA trigger signaling from two different cellular locations: theformer at the cell membrane and the latter at the lysosomal compartment. While LPS does not require endocytosis to functionally associate with the membrane expressed TLR4/MD2 complex, internalization and endosomal maturation is conditional for CpG-DNA to activate TLR9. In support of these data TLR9 is not localized at the cell surface, but intracellularily. These data stress the need to characterize individual TLR at the very beginning of signal initiation in order to understand their diverse biological functions. PMID- 12115617 TI - The complementation of lymphotoxin deficiency with LIGHT, a newly discovered TNF family member, for the restoration of secondary lymphoid structure and function. AB - Highly organized lymphoid structures provide the intricate microenvironment essential for the mediation of the effective immune responses. Compared with lymphotoxin beta knockout mice (LTbeta-/-), LTbeta receptor knockout (LTbetaR-/-) mice present with more severely disorganized splenic structures, suggesting the potential involvement of another ligand. LIGHT, a newly identified TNF family member, is a costimulatory molecule for T cells and binds to LTbetaR and herpes virus entry mediator (HVEM) in vitro. Here, we show that the complementation of LTalpha-/- mice with a LIGHT transgene (LIGHT Tg/LTalpha-/-) leads to the restoration of secondary lymphoid tissue chemokine and T/B cell zone segregation. LIGHT Tg/LTalpha-/- mice also preserve dendritic cells, follicular dendritic cell networks, and germinal centers, though not the marginal zone. Consequently, IgG responses to soluble, but not particulate, antigens are restored, confirming the role of primary follicle and marginal zone in the responses to soluble and particulate antigens. The failure of the LIGHT transgene to rescue the defective splenic structures in LTbetaR-/- mice demonstrates that LIGHT can interact with LTbetaR in vivo. More severely disorganized splenic structures developed after blockade of endogenous LIGHT in LTbeta-/- mice. These findings uncover the potential interaction between LIGHT and one of its receptors, LTbetaR, in supporting even in the absence of LT the development and maintenance of lymphoid microenvironment. PMID- 12115618 TI - The differential contribution of granzyme A and granzyme B in cytotoxic T lymphocyte-mediated apoptosis is determined by the quality of target cells. AB - Complementary approaches with purified molecules or transfected cytolytic effector cells have suggested that both, granzyme A (gzmA) and granzyme B (gzmB), similarly contribute to CTL-mediatedand perforin (perf)-dependent apoptotic nuclear damage (DNA fragmentation) in target cells. Studies employing gzmA or gzmB single-knockout mice on the other hand indicated that gzmB is the prominent CTL effector molecule for the rapid induction of DNA fragmentation, with gzmA playing only a minor part. We have now taken ex vivo-derived virus-specific or in vitro generated alloreactive CTL from mice deficient in either gzmA or gzmB and a panel of three target cells to reinvestigate this unresolved issue. We show that rapid CTL-mediated DNA fragmentation of L1210.3 target cells is solely dependent on gzmB, whereas the DNA fragmentation of EL4.F15 target cells by the same CTL population is mainly induced by gzmA and only marginally by gzmB. Moreover, CTL induced apoptosis of a third target cell, MC57G, was partially dependent on both gzmA and gzmB activities. The differential contribution of the two gzms to apoptosis was further verified by their distinct sensitivity tocaspase inhibitors. The data suggest that both, gzmA and gzmB, have a similar potential to induce rapid perf-mediated apoptosis but that their individual contribution to the underlying intracellular processes is dictated by the quality of the target cell. PMID- 12115619 TI - Inhibition of caspase or FADD function blocks proliferation but not MAP kinase activation and interleukin-2-production during primary stimulation of T cells. AB - Caspases are instrumental in the implementation of apoptotic cell death, and caspase activation is in most investigated cases closely linked to apoptosis. Recent data demonstrate, however, that caspases are also activated during primary T cell activation in the absence of apoptosis. Here we provide evidence that caspase activity is required for some but not all aspects of T cell activation. CD3-triggered proliferation of mouse T cells was impaired in the presence of the pan-caspase-inhibitor Z-Val-Ala-Asp-fluoromethylketone (Z-VAD-fmk) and the number of cells entering the cell cycle was reduced. Costimulation by CD28 or externally added interleukin-2 (IL-2) failed to rescue proliferation. Re-stimulation of pre activated T cells, however, was not affected by Z-VAD-fmk. Intriguingly, CD3 induced production of IL-2 by primary T cells was not impaired in the presence of Z-VAD-fmk. Likewise, CD3-induced activation of mitogen-activated protein kinases was unaffected by Z-VAD-fmk and intracellular levels of inhibitory kappaBalpha were the same as in control cells. T cells transgenically expressing a dominant negative mutant of the caspase-adaptor Fas-associated molecule with death domain (FADD)/MORT1 displayed the same pattern of reaction, i.e. a reduced proliferative response but normal IL-2-production. These data show a distinct role of caspases during primaryT cell activation and provide evidence for a FADD-caspase-pathway not only in the induction of apoptosis but also of T cell proliferation. PMID- 12115620 TI - Membrane lymphotoxin contributes to anti-leishmanial immunity by controlling structural integrity of lymphoid organs. AB - Lymphotoxin (LT)alpha in combination with LTbeta forms membrane-bound heterotrimeric complexes with a crucial function in lymph node (LN) organogenesis and correct morphogenesis of secondary lymphoid tissue. To study the role of membrane LT (mLT) in lymphoid tissue organogenesis we generated an LTbeta deficient mouse strain on a pure genetic C57BL/6 background (B6.LTbeta-/-) and compared it to a unique series of LTalpha-, TNF- and TNF/LTalpha-gene-targeted mice on an identical genetic background (B6.LTalpha-/-, B6.TNF-/- and B6 TNF/LTalpha-/-). B6.LTbeta-/- mice lacked peripheral LN with the exception of mesenteric LN, and displayed a disturbed micro-architecture of the spleen, although less profoundly than LTalpha- or TNF/LTalpha-deficient mice. Radiation bone marrow chimeras (B6.WT-->B6.LTbeta-/- developed Peyer's patch (PP)-like lymphoid aggregates in the intestinal wall indicating a possible role for soluble LTalpha(3) in the formation of the PP anlage. After infection with Leishmania major, B6.LTbeta-/- mice developed a fatal disseminating leishmaniasis resulting in death after 8 to 14 weeks, despite the natural resistance of the C57BL/6 genetic background (B6.WT) mice to the parasite. Both, the cellular and the humoral anti-L. major immune responses were delayed and ineffective. However, the expression pattern of the key cytokines IFN-gamma and IL-12 were comparable in B6.WT and B6.LTbeta-/- mice. Infection of radiation bone marrow chimeras showed that it is the LTbeta-dependent presence of lymphoid tissue and not the expression of mLT itself that renders mice resistant to leishmaniasis. PMID- 12115621 TI - BAFF is a survival and maturation factor for mouse B cells. AB - Human B cell-activating factor (BAFF) induces mouse surface IgM+ B cells of the immature type from bone marrow and of the immature types 1 and 2 from spleen, as well as of the mature type from spleen to increased longevity in tissue culture. BAFF does so polyclonally and without inducing proliferation in any of these B cell subpopulations. BAFF induces phenotypic and functional maturation of immature to mature B cells so that all immature cells loose C1qRp (AA4.1, 493) expression and type 1 immature cells up-regulate IgD, CD21 and CD23. Immature B cells of types 1 and 2, upon pre-incubation with BAFF, change their reactiveness to Ig-specific antibodies so that they no longer enter apoptosis but now proliferate. However, BAFF does not seem to overcome negative selection of developing immature B cells in vitro. PMID- 12115622 TI - Increased IL-10 production during spontaneous apoptosis of monocytes. AB - Monocytes/macrophages undergo apoptosis and are in contact with apoptotic cells both in vitro and in vivo. The data show that monocytes undergoing spontaneous apoptosis in vitro change their cytokine production profile. We demonstrate that the lipopolysaccharide (LPS)-induced production of interleukin-10 (IL-10) is up regulated, while production of pro-inflammatory cytokines such as tumor necrosis factor-alpha (TNF-alpha) and interleukin-1beta (IL-1beta) is either not affected or reduced. These differences seen both at the protein and mRNA level directly correlate with the appearance of apoptotic cells in the culture. Flow cytometry analysis using double staining, surface with annexin V and intracellular with anti-IL-10, suggested that annexin V-negative monocytes are the predominant source of IL-10. Analysis of sorted populations of monocytes indicated that the increase in IL-10 synthesis appears to result from direct interactions between non-apoptotic and apoptotic cells at the time of stimulation. Also non-apoptotic, freshly isolated monocytes produced more IL-10 upon stimulation with LPS, Staphylococcus aureus or zymosan when apoptotic neutrophils were added to the culture. In contrast, monocyte-derived macrophages did not produce more IL-10 in the presence of apoptotic neutrophils. Finally, we found that the presence of apoptotic monocytes in the culture may influence specific immune responses. The data show that in the presence of annexin V-positive monocytes CD4-positive memory T cells produce less IFN-gamma upon stimulation with purified protein derivative of tuberculin, which could be partially reversed by anti-IL-10 neutralizing antibodies. We conclude that these findings might illustrate the mechanisms operating within an inflammatory site and play an important immunoregulatory role during the resolution of inflammation and specific immune responses. PMID- 12115623 TI - Regulatory Th2 response induced following adoptive transfer of dendritic cells in prediabetic NOD mice. AB - We previously demonstrated that immunotherapy with dendritic cells (DC) prevented diabetes development in prediabetic NOD mice and that this effect was optimal when using a stimulatory DC population generated from bone marrow cells cultured with granulocyte-macrophage colony-stimulating factor (GM-CSF) and IL-4. In this study, we have investigated the mechanism by which GM-CSF- and IL-4-cultured DC prevent diabetes in prediabetic NOD mice. Histological analysis of pancreatic tissue from DC-treated mice revealed a reduction in the severity of insulitis compared to controls. Analysisof the T cell response in DC-treated mice suggested a general shift towards a Th2-dominated response, as determined by cytokine production following either concanavalin A or anti-TCR stimulation. Furthermore, sorted CD45RB(lo) CD25+ CD4+ T cells from the spleen of DC-treated mice produced high amounts of Th2 cytokines following anti-TCR stimulation, suggesting that these cells are responsible for the apparent Th2 shift. We conclude that DC therapy may have corrected the immunoregulatory defect in the NOD mouse, thus restoring a balance between pathogenic Th1 cells and protective Th2 cells. PMID- 12115624 TI - Ly49i2 is an inhibitory rat natural killer cell receptor for an MHC class Ia molecule (RT1-A1c). AB - We have exploited strain-specific differences in the NK allorecognition repertoires to generate rat monoclonal antibodies against receptors involved in the control of allogeneic responses by rat NK cells. The monoclonal antibody STOK2 binds to a homodimeric glycoprotein that has been implicated as an inhibitory receptor for an MHC molecule in the PVG strain. In the present study, we haveidentified this glycoprotein as a novel rat Ly49 receptor (Ly49i2) containing an immunoreceptor tyrosine-based inhibitory motif. Ligation of the Ly49i2 receptor induces inhibitory signals, and Ly49i2 coprecipitates with the inhibitory tyrosine phosphatase SHP-1 in stably transfected RNK-16 cells. Moreover, it inhibits natural killing of lymphoblast targets and transfected fibroblast targets expressingthe classical MHC class Ia allele RT1-A1(c). Ly49i2, therefore, is an inhibitory receptor for specific MHC class Ia molecules, similar to inhibitory members of the mouse Ly49 family. PMID- 12115626 TI - T cell priming by dendritic cells: thresholds for proliferation, differentiation and death and intraclonal functional diversification. AB - The variables that influence priming of human naive CD4+ T cells by dendritic cells (DC) were dissected in vitro by analyzing the response to the bacterial superantigen toxicshock syndrome toxin or to alloantigens. We show that under conditions that force DC-T cell interactions a single DC can prime up to 20 naive T cells. Moreover, the strength of antigenic stimulation, as determined by DC numbers, antigen dose, TCR avidity and duration of DC-T cell interactions, drives the progressive differentiation of proliferating T cells from a non-effector CCR7+ stage, to an effector CCR7- stage and, eventually, to cell death. We also show that the proliferating CCR7+ and CCR7- populations share clonotypic sequences, demonstrating that the two cell fates can be generated within a single clone. Taken together these results indicate that the strength of antigenic stimulation regulates T cell progression through thresholds of proliferation, differentiation and death. However, the random nature of DC-T cell encounters introduces a critical stochastic element in T cell stimulation, which leads to the generation of cells endowed with distinct homing potentials and effector functions within a given T cell clone. PMID- 12115625 TI - TNFalpha-induced macrophage chemokine secretion is more dependent on NF-kappaB expression than lipopolysaccharides-induced macrophage chemokine secretion. AB - The transcription factor NF-kappaB is a pivotal intracellular regulator of many inflammatory responses and it has been proposed that it represents a potential therapeutic target. As chemokines are important for the progress of an inflammatory response by the recruitment of immuno-competent cells, the role NF kappaB plays in TNFalpha- or lipopolysaccharides (LPS)-induced chemokine secretion by human monocyte-derived macrophages was examined. Secretion of the CXC chemokines IL-8, GROalpha and ENA-78, induced by TNFalpha, was significantly suppressed by inhibiting NF-kappaB, using overexpression of IkappaBalpha. However, when induced by LPS the expression of these chemokines was unaffected. In contrast, expression of the CC chemokines MIP-1alpha, MCP-1 and RANTES inducedby TNFalpha or LPS was significantly inhibited by the overexpression of IkappaBalpha. Therefore, there appear to be different mechanisms regulating CC and CXC chemokine secretion by macrophages, depending on the stimulus and that TNFalpha and LPS can use different signaling mechanisms in macrophages to regulate chemokine synthesis. PMID- 12115627 TI - T cell receptor transgenic mice recognizing the immunodominant epitope of the Torpedo californica acetylcholine receptor. AB - Myasthenia gravis (MG) is an autoimmune disease caused by T cell-dependent antibody-mediated reduction of acetylcholine receptors (AChR) at the neuromuscular junction. Immunization of animals with Torpedo californica AChR (TAChR) results in an experimental model of MG. We used the variable regions of alpha and beta T cell receptor (TCR) genes recognizing an immunodominant peptide containing amino acids 146-162 from the alpha subunit of TAChR presented in the context of I-A(b) to generate TCR-transgenic mice. We found that the transgenic TCR was strongly positively selected and that transgenic T cells proliferated robustly to the immunodominant peptide and TAChR. Unexpectedly, there was a variable paucity of B cells in the blood and spleen from transgenic mice, which averaged about 16% of peripheral blood lymphocytes, compared to 55% in wild-type B6 mice. Unselected transgenic mice immunized with TAChR exhibited weak anti TAChR antibody responses. However, transgenic mice selected to have relatively higher B cell numbers produced anti-TAChR titers equal to B6 mice and a predominance of Th1-induced antibody isotypes were observed in certain experiments. The incidence and severity of clinical disease was variable following immunizations. These mice should be useful for studying the pathogenesis and treatment of MG. PMID- 12115628 TI - A short peptide at the C terminus is responsible for the nuclear localization of RAG2. AB - The RAG1 and RAG2 proteins are the lymphoid-specific factors essential for V(D)J recombination, the process that leads to the diversification of antigen receptors on B and T lymphocytes. Nucleolar/nuclear localization of RAG1 is mediated by four basic domains, which are the binding sites for the nuclear transport proteins SRP1 and RCH1, and by a nuclear localization signal (NLS) in the fifth basic domain. The C-terminal region of RAG2 from amino acids (aa) 417 to 484 shows a homology with the PHD domain of other proteins involved in chromatin mediated gene regulation by protein-protein interactions. Mutations in this domain were shown to be responsible for several diseases and in some case lead to altered subcellular localization of proteins. We found that the C-terminal PHD domain of RAG2 is not responsible for the nuclear localization of the protein. We report here the characterization of a region (aa 491-527) in the C-terminal domain of RAG2, downstream of the putative PHD domain, which directs the nuclear localization of the protein. PMID- 12115629 TI - Extension, retraction and contraction in the formation of a dendritic cell dendrite: distinct roles for Rho GTPases. AB - The morphology of antigen-presenting dendritic cells (DC) is characterized by the possession of numerous long arborizing processes known as dendrites. The formation of these processes by DC, both in the periphery and in lymphoid organs, is believed to contribute to the remarkable efficiency with which they take up, process and present antigen to T cells. However, the process of dendrite formation and the signaling pathways that lead to the formation of these dendrites remain obscure. In this study we describe an in vitro model in which human immature DC form long processes similar to those formed in vivo. The formation of these processes involves initial attachment of a cell protrusion to the extracellular matrix substrate, and subsequent movement of the cell body away from the adhesion site, leaving behind a long slender dendrite. Dendrite formation, but not their maintenance, was found to be dependent on the activity of Rho GTPases. More specifically, Cdc42 and Rac1 were both required for the migration step of process formation, promoting cell spreading and extension. In contrast, Rho, and its downstream effector p160ROCK, regulated the release of adhesions to the substratum, and associated cellular contraction. Consequently, inhibition of Rho/p160ROCK leads to the formation of longer dendrites. DC therefore coordinate adhesion and protrusion to perform a specialized process of cellular morphogenesis, which differentiates these cells from all other cells of the immune system and may contribute to their distinctive function. PMID- 12115633 TI - Natural mutants of the immune system: a lot to learn! PMID- 12115630 TI - CpG motifs of bacterial DNA exacerbate colitis of dextran sulfate sodium-treated mice. AB - Inflammatory bowel disease (IBD) is characterized by a dysregulated intestinal immune response with elevated levels of the Th1 cytokines TNF, IL-12 and IFN gamma. The luminal flora has been implicated as a major factor contributing to the initiation and perpetuation of chronic intestinal inflammation by as yet unknown mechanisms. Bacterial DNA contains unmethylated cytosine-guanosine dinucleotides (CpG) which strongly activate Th1-mediated immune responses. To test whether these CpG-motifs contribute to intestinal inflammation we treated mice with dextran-sulfate-sodium (DSS)-induced acute or chronic colitis for 5 days with CpG-containing oligodeoxynucleotides (CpG-ODN). Colonic inflammation was assessed by histological scoring. Colonic cytokine RNA was quantified by reverse transcription-PCR and cytokine secretion from mesenterial lymph node cells by ELISA. In chronic colitis, CpG-ODN treatment severely aggravated inflammation by 50%. Colonic expression of IFN-gamma and TNF was elevated (200- and 150-fold, respectively) and IFN-gamma and IL-12 secretion from lymph node cells was increased 5,000- and 8-fold, respectively, compared to GpG-ODN-treated controls. Similar effects were obtained in acute colitis. In conclusion, CpG motifs of bacterial DNA have proinflammatory activity by strengthening the Th1 arm of immunity in DSS-induced colitis, and might therefore play a significant role in the initiation and perpetuation of inflammation in IBD. PMID- 12115635 TI - Alternatively spliced forms of Igalpha and Igbeta prevent B cell receptor expression on the cell surface. AB - The B cell antigen receptor (BCR) includes an Igalpha/Igbeta heterodimer non covalently associated with surface immunoglobulin. Recently, variant Igalpha and Igbeta transcripts, arising from alternative mRNA splicing, have been reported. The present study examined the function of the potential products of these transcripts, by utilizing cDNA expression plasmids to reconstitute human BCR expression in transfected 293T cells. Spliced transcripts produced truncated proteins (deltaIgalpha and deltaIgbeta), that failed to form heterodimers with their full-length counterparts, and did not mediate transport of IgM to the cell surface. When overexpressed, both deltaIgalpha and deltaIgbeta acted as competitors of Igalpha and Igbeta, leading to down-modulated surface IgM expression, and retention of IgM in the endoplasmic reticulum. These findings document a possible novel mechanism for controlling BCR expression in B cells, based on up-regulated synthesis of components devoid of transport function. PMID- 12115634 TI - Chemokine production by natural killer cells from nonagenarians. AB - In this study we investigated whether purified NK cells, derived from a group of nonagenarian healthy subjects, were able to produce the chemokines MIP-1alpha, RANTES and IL-8, and also characterized the effect of IL-12 or IL-2 immunomodulatory cytokines (that are among the most effective inducers of NK lytic activity and soluble factor secretion) on the induction, in vitro, of these chemokines and on the modulation of the corresponding receptors. This study provides evidence that human NK cells from healthy subjects over 90 years old retain the ability to synthesize MIP-1alpha, Rantes and IL-8 chemotactic cytokines, that NK cells isolated from these subjects can be activated to significantly up-regulate the production of these chemokines in response to stimulation by IL-12 or IL-2 cytokines (even though production remains lower than that observed in young subjects), and that NK cells express the corresponding chemokine receptors. PMID- 12115636 TI - The number of long-lasting functional memory CD8+ T cells generated depends on the nature of the initial nonspecific stimulation. AB - The mechanism of generation of memory cytotoxic T cells (CTL) following immunization remains controversial. Using tumor protection and IFN-gamma ELISPOT assays in mice to detect functional CTL, we show that the initial effector CTL burst size after immunization is not directly related to the amount of functional memory CTL formed, suggesting that memory CTL are unlikely to arise stochastically from effector CTL. Induction of MHC class II-restricted T helper cells at the time of immunization by inclusion of a T helper peptide or protein in the immunogen, is necessary to generate memory CTL, although no T helper cell induction is required to generate effector CTL to a strong MHC class I-binding peptide. Host protective T cell memory correlates with the number of CTL epitope responsive IFN-gamma-secreting memory T cells as measured in an ELISPOT assay at the time of tumor challenge. We conclude that a different antigen presenting environment is required to induce long-lasting functional memory CTL, and non cognate stimulation of the immune system is essential to allow generation of a long-lasting host protective memory CTL response. PMID- 12115637 TI - Somatostatin is expressed in the murine thymus and enhances thymocyte development. AB - A functional interaction between the immune and the nervous system has been suggested, with neuropeptides acting as immunomodulators. Somatostatin (SOM) is a neuropeptide, mainly produced in the brain, that binds to five different receptors (SSTR). It is believed that SOM along with one of its receptors, SSTR2, is expressed in the murine thymus, although their exact localization is unresolved. We found that SOM is highly expressed in both cortical and medullary epithelial cells whereas its receptor SSTR2 is expressed on thymocytes. In order to elucidate its role in thymopoiesis, SOM was added in fetal thymic organ culture (FTOC) and found to increase thymocyte numbers and enhance maturation. SOM increased the cellular proliferation of total splenocytes but inhibited proliferation of thymocytes and purified splenic T cells. Furthermore, SOM was able to induce the migration of thymocytes. We also investigated the effect of four other neuropeptides in FTOC and found that, vasoactive intestinal peptide had a marginal effect, whereas substance P increased thymic cellularity, at intermediate but not at low or high concentrations. In contrast, both neuropeptide Y and calcitonin gene-related peptide reduced thymocyte numbers. This study supports the hypothesis for a role of neuropeptides, particularly somatostatin, in immune regulation and development. PMID- 12115638 TI - A juxta-membrane amino acid sequence of P-selectin glycoprotein ligand-1 is involved in moesin binding and ezrin/radixin/moesin-directed targeting at the trailing edge of migrating lymphocytes. AB - P-selectin glycoprotein ligand 1 (PSGL-1) is an adhesion receptor localized on the tips of microvilli that is involved in the rolling of neutrophils on activated endothelium. We found that PSGL-1 was concentrated at the uropod of chemokine-stimulated lymphoid cells. Dynamic fluorescence videomicroscopy analyses of migrating lymphocytes demonstrated that PSGL-1 and moesin redistributed towards the cellular uropod at the trailing edge of these cells, where activated ezrin/radixin/moesin (ERM) proteins were located. An eighteen amino acid sequence in the juxta-membrane region of the PSGL-1 cytoplasmic tail was found to be critical for uropod targeting and moesin binding. Substitution of S336, S348, and the basic cluster R337K338 by alanines within this region significantly impaired both moesin binding and PSGL-1 polarization. These results underline the role of moesin in the subcellular redistribution of PSGL-1 in lymphoid cells and make evident the importance of specific serine residues within the cytoplasmic tail of PSGL-1 for this process. PMID- 12115639 TI - Defective generation but normal maintenance of memory T cells in old mice. AB - The ability to maintain memory after encounter with antigen is one of the central features of the immune system. Immune memory generated during young age can be maintained well into old age. However, we know little about maintenance of immune memory generated during old age. In this study, we compared generation and maintenance of memory CD8 T cells in young (2-3-month-old) and aged (22-24-month old) mice following acute lymphocytic choriomeningitis virus infection. We found that young mice made a more vigorous primary T cell response and generated higher levels of memory cells than old mice. However, once generated, memory CD8 T cells were maintained at stable levels in both young and old mice for more than 5 months. Interestingly, the generation of a secondary effector response in vivo was again slightly compromised in the old mice. Taken together, these results show that generation of T cell responses is compromised in old age, but maintenance of the pool of memory T cells is not affected by the aging process. PMID- 12115640 TI - Identification and characterization of Xenopus CD8+ T cells expressing an NK cell associated molecule. AB - Growing evidence suggests that some immune responses are mediated not only by conventional and distinct NK cells and CTL, but also by T cell subsets expressing NK receptors and NK cell-associated molecules. Consistent with our previously published finding that the mAb 1F8 identifies non-T/non-B cells in Xenopus that effect NK-like killing in vitro, we now report that in vivo treatment with this mAb impairs rejection of transplanted MHC class I-negative tumor cells. However, we also find that the NK cell-associated molecule recognized by mAb 1F8 is expressed by a minor population of CD8+ T cells, in which fully rearranged TCRbeta mRNA of at least three different V families can be identified, by contrast, 1F8+/CD8- (NK) cells lack such TCRbeta message. Additionally, the expression of the NK cell-associated molecule can be induced in vitro by a transient submitogenic stimulation of naive CD8+ T cells with PMA and ionomycin. Such induced expression of 1F8 also occurs in alloantigen-activated CTL and is coincident with a down-regulation of MHC-specific cytotoxicity. Taken together, these new data suggest that regulation of CD8+ T cell activity involving NK cell associated molecules is a general and evolutionarily ancient phenomenon. PMID- 12115641 TI - Ly-6A is critical for the function of double negative regulatory T cells. AB - We have recently demonstrated that CD3+CD4-CD8- double negative (DN) T cells can down-regulate allogeneic immune responses both in vitro and in vivo by killing activated syngeneic CD8+ T cells. The goal of this study was to identify molecules that are crucial for DN T cell-mediated suppression. We demonstrate that Ly-6A (Sca-1) is highly expressed on DN T cells. Incubation with IL-10 significantly reduced Ly-6A expression and the function of DN T cells. DN T cell mediated killing was significantly reduced when Ly-6A was blocked.Ly-6A-deficient mice showed an accelerated allograft rejection when compared to wild-type controls. Furthermore we demonstrate that pretransplantation donor lymphocyte infusion (DLI) led to activation and proliferation of recipient DN T cells and prolongation of bm1-->B6 skin allograft survival. However, when the recipients were deficient in Ly-6A, the beneficial effect of DLI on allograft survival was abolished. Moreover, deficiency in Ly-6A did not affect the activation and proliferation of DN T cells. Rather, it impaired the ability of DN T cells to kill activated anti-donor CD8+ T cells. Taken together, our data indicate that Ly 6A plays a crucial role in DN T cell-mediated regulation in vitro and in vivo, perhaps by enhancing DN-CD8+ T cell signaling. PMID- 12115642 TI - The fate of duplicated major histocompatibility complex class Ia genes in a dodecaploid amphibian, Xenopus ruwenzoriensis. AB - The dodecaploid anuran amphibian Xenopus ruwenzoriensis represents the only polyploid species of Xenopus in which the full silencing of the extra copies of the major histocompatibility complex (MHC) has not occurred. Xenopus ruwenzoriensis is a recent polyploid that has evolved within one of the two tetraploid groups of Xenopus through allopolyploidization. Family studies of its MHC haplotype suggested a polysomic inheritance of the MHC class I and II genes. Four class Ia bands can be detected per individual in Southern blot analysis and, similarly, four different cDNA sequences are expressed per individual. The Xenopus class Ia sequences we analyzed belong to only one of the old class I lineages and show a homogenization of their alpha3 domain sequences. This homogenization occurred after speciation within the Xenopus ruwenzoriensis species, either due to gene conversion or inter-alleles/loci recombination.A re evaluation of the polymorphism of class Ia in Xenopus, by looking at the rate of non-synonymous versus synonymous substitutions, suggests that Xenopus MHC class Ia genes are not under strong overdominant selection. This is a rare situation among vertebrates. The observed polymorphism is most likely due to the interlocus genetic exchanges related to the peculiar mode of speciation of the genus. PMID- 12115643 TI - Active tuberculosis in Africa is associated with reduced Th1 and increased Th2 activity in vivo. AB - Activation of Th1 lymphocytes, IFN-gamma production and macrophage activation are crucial in defense against Mycobacteria. In developing countries, Th2 activation and IL-4 production have been associated in vitro with tuberculosis and with poor clinical outcome after treatment. Serological markers of Th1 [soluble lymphocyte activation gene (LAG)-3] and Th2 (IgE, solubleCD30, and CCL22/macrophage-derived chemokine) activity were measured in 414 HIV-negative tuberculosis patients from The Gambia and Guinee and in 414 healthy household and community controls. Measurements were repeated during treatment to assess the effect of therapy on Th1/Th2 ratio. At diagnosis, sLAG-3 levels were lower in patients than in community controls (p<0.0001), but were higher in household controls exposed to contact with patients than in community controls (p<0.0001). In comparison with community controls, patients had consistently higher levels of IgE, sCD30, and CCL22 (p<0.0001), whereas household controls had lower levels of indicators of Th2 activity (p<0.0001). After treatment, cured patients had higher levels of Th1 (p<0.0001) and lower levels of Th2 (p<0.0001) activity than patients who were not successfully treated or interrupted therapy. In Africa, tuberculosis is associated with low Th1 and high Th2 activity in vivo, whereas close exposure to tuberculosis is associated with a high Th1/Th2 ratio. Patients with favorable outcome after treatment exhibit a higher Th1/Th2 ratio compared to patients with poor clinical outcome. PMID- 12115644 TI - BCR mutants deficient in ligand-independent and more sensitive for ligand dependent signaling. AB - Signal transduction by the B cell antigen receptor (BCR) regulates development, survival and clonal expansion of B cells. The BCR complex comprises the membrane bound immunoglobulin molecule (mIg) and the Ig-alpha/Ig-beta heterodimer, and was shown to form oligomeric structures. In pervanadate (PV)-treated B cells, multiple proteins are tyrosine phosphorylated upon expression of the BCR, indicating that the BCR can signal in an antigen-independent fashion. We analyzed the signal transduction from BCR mutants which either have an altered heavy chain transmembrane region or lack the Ig-alpha cytoplasmic tail. In comparison to cells expressing the wild-type receptors, those with a mutant BCR respond to PV treatment with reduced and retarded tyrosine phosphorylation of substrate proteins. Conversely, the cells with mutant BCR are more sensitive to stimulation with low doses of antigen. These data suggest that a correctly assembled BCR complex is important for antigen-independent signaling and setting the threshold for antigen-dependent BCR activation. PMID- 12115645 TI - Antigen-specific T cell suppression by human CD4+CD25+ regulatory T cells. AB - Anergic/suppressive CD4+CD25+ T cells have been proposed to play an important role in the maintenance of peripheral tolerance. Here we demonstrate that in humans these cells suppress proliferation to self antigens, but also to dietary and foreign antigens. The suppressive CD4+CD25+ T cells display a broad usage of the T cell receptor Vbeta repertoire,suggesting that they recognize a wide variety of antigens. They reside in the primed/memory CD4+CD45RO+CD45RB(low) subset and have short telomeres, indicating that these cells have the phenotype of highly differentiated CD4+ T cells that have experienced repeated episodes of antigen-specific stimulation in vivo. This suggests that anergic/suppressive CD4+CD25+ T cells may be generated in the periphery as a consequence of repeated antigenic encounter. This is supported by the observation that highly differentiated CD4+T cells can be induced to become anergic/suppressive when stimulated by antigen presented by non-professional antigen-presenting cells. We suggest that besides being generated in the thymus, CD4+CD25+ regulatory T cells may also be generated in the periphery. This would provide a mechanism for the generation of regulatory cells that induce tolerance to a wide array of antigens that may not be encountered in the thymus. PMID- 12115646 TI - DNA vaccine using invariant chain gene for delivery of CD4+ T cell epitope peptide derived from Japanese cedar pollen allergen inhibits allergen-specific IgE response. AB - To establish a new immunotherapy for type I allergic diseases without allergic side effects, we attempted to develop a DNA vaccine encoding both a CD4+ T cell epitope site in a major Japanese cedar pollen allergen (Cry j 2) and an invariant chain (Ii) for the delivery of the epitope peptide into the MHC class II loading pathway. We constructed a plasmid DNA encoding the Ii mutant either by replacement of the core CLIP (class II-associated invariant chain peptide) with a peptide corresponding to the major Cry j 2 CD4+ T cell epitope in BALB/c mice, designated as p247-258 (pCPCJ2), or by fusion of the Ii with p247-258 at the C terminus (pIiCJ2). As expected, repeated inoculation of BALB/c mice with pCPCJ2 or pIiCJ2 induced no antibody response to Cry j 2. In contrast, intramuscular inoculation of BALB/c mice with pCPCJ2 or pIiCJ2 predominantly induced p247-258 specific Th1 cells, resulting in the inhibition of IgE response to subsequent Cry j 2 injections. Our results demonstrated that the plasmid DNA encoding the CD4+ T cell epitope and Ii can induce epitope-specific CD4+ T cell responses in vivo and the potential to regulate type I allergic reaction without allergic side effects. PMID- 12115647 TI - CD84 is up-regulated on a major population of human memory B cells and recruits the SH2 domain containing proteins SAP and EAT-2. AB - CD84 is a member of the CD2 subset of the immunoglobulin superfamily of cell surface receptors. Several members of this family are involved in the activation of T cells and NK cells. Although CD84 was originally cloned from a human B cell line cDNA library, very little is known regarding its biology on primary human leukocytes. We investigated the expression and biochemistry of CD84 on human B cells. CD84 was expressed on B cells in peripheral blood, spleen and cord blood. Two populations of splenic B cells could be resolved, CD84(lo) and CD84(hi). CD84(hi) B cells represented a subset of memory B cells as demonstrated by increased cell size, co-expression of the memory B cell-specific marker CD27, somatically mutated Ig variable region genes, and increased proliferation compared to CD84(lo) B cells. CD84 became rapidly phosphorylated on tyrosine residues following ligation with a specific monoclonal antibody and recruited the cytoplasmic adaptor proteins SAP and EAT-2. The ability of CD84 to undergo tyrosine phosphorylation and to recruit these SH2 domain-containing proteins suggests it may function in the activation of B cells, particularly memory cells, and its signal transduction pathway may utilize SAP and/or EAT-2. Thus, investigation of expression and function of CD84 and CD27 is likely to contribute to a greater understanding of the development and biology of memory B cells in normal and immunocompromised hosts. PMID- 12115648 TI - Tumor antigen drives a persistent oligoclonal expansion of CD8+ T cells in aged mice. AB - Oligoclonal T cell expansions (TCE) are common in old humans and mice, but it is not known whether the T cell response to a specific antigen is more restricted in old vs. young animals. Herein, we describe an enhanced and prolonged response of tumor antigen-specific CD8 cells in old mice identified by K(d)/peptide tetramers and Vbeta10 staining. At the onset of the response CD8 T cell numbers and Vbeta10+CD8+ cells at the site of tumor injection were lower in old mice, hinting that control of initial tumor growth may not be optimal. As further evidence of a dysregulated response in old mice, antibody titers to the tumor were deficient and the CD8 tumor antigen-specific response was greater and more prolonged in the blood and spleen. Old mice selected a more oligoclonal TCR repertoire based on TCRbeta chain CDR3 length analysis and sequences. Persistent expansions of Vbeta10+CD8+ cells in old mice had memory/activation phenotypes. This induced tumor antigen-specific response may represent a model for the spontaneous TCE observed with aging and demonstrates that the CD8 response to a defined peptide/MHC antigen is indeed more oligoclonal in old mice. PMID- 12115649 TI - Non-germ-line encoded residues are critical for effective antibody recognition of a poorly immunogenic neutralization epitope on glycoprotein B of human cytomegalovirus. AB - The capability of the antibody (Ab) repertoire to mount a response to appropriate epitopes on infectious agents will strongly affect the ability of the immune system to provide protection against disease. Certain epitopes may be poor inducers of immunity but are nevertheless able to promote biologically important protection when recognized by the immune repertoire. We have investigated the recognition by Ab of one such poorly immunogenic target, antigenic domain 2 (AD 2) on human cytomegalovirus glycoprotein B. To date, only two well-characterized human monoclonal Ab reactive with this epitope are known. To define parameters important for establishment of a human paratope recognizing this epitope, we created variants of the variable genes utilized by one of these Ab and used phage display technology to select Ab fragments with retained antigen specificity. We show here that residues in the first complementarity determining region of both the heavy and the light chain are involved in determining the AD-2 specificity and, in addition, that key mutations in the germ-line sequence are required for effective interaction with this epitope. Thus, the products of the human germ line IGHV3-30 and IGKV3-11 genes, the only V genes that have been demonstrated to participate in an AD-2 specific Ab response, do not have the intrinsic features required for high-affinity recognition of this epitope. We propose that the inability of the human germ-line gene-encoded Ab repertoire to directly recognize this and possibly other antigenic determinants results in their poor immunogenicity in vivo. This may favor responses to other epitopes, which have a high-affinity imprint in the human germ-line Ab repertoire. PMID- 12115650 TI - Lck tyrosine kinase is important for activation of the CD11a/CD18-integrins in human T lymphocytes. AB - CD11a/CD18 (beta2)-integrins are expressed on leukocytes and are involved in cell adhesion and signaling. Despite extensive studies the signaling pathways and molecular mechanisms involved in integrin regulation in T cells remain not completely understood. We have now studied the involvement of the tyrosine kinase Lck in the regulation of CD11a/CD18 function in Jurkat T cells. Using the Src family kinase inhibitor PP2, we found that CD3 ligation-induced adhesion to ICAM 1 was inhibited by PP2 at the same concentration required for complete inhibition of the MAP kinase pathway, implicating a role for Lck in integrin activation. We therefore used the Lck-deficient Jurkat cell line JCaM1.6 to further examine the involvement of Lck in integrin regulation. Interestingly, JCaM1.6 cells showed dramatically reduced levels of both CD3- and phorbol ester-induced adhesion to coated ICAM-1 as compared to normal Jurkat cells. By using flow cytometry and cell surface labeling, it was found that the surface expression of the CD11a/CD18 integrins was significantly lower in Lck-deficient T cells as compared to normal Jurkat cells. CD18 was expressed as a mature and an immaturely glycosylated form in Jurkat T cell lines, and predominantly the immature form, not associated with CD11a, was found in Lck-deficient cells. Retransfection of human Lck in JCaM1.6 cells restored adhesion. Thus, Lck is involved in regulating CD11a/CD18-integrins in T cells. PMID- 12115651 TI - B cells engineered to express Fas ligand suppress pre-sensitized antigen-specific T cell responses in vivo. AB - Inducing apoptosis of activated lymphocytes via Fas ligand (FasL, CD95) may be a useful strategy for the treatment of autoimmune diseases mediated by pathogenic T cells. We propose that B cells may be ideal tools for effective delivery of a FasL-mediated apoptotic signal to pathogenic T cells for a variety of reasons, including their unique ability to efficiently take up and present antigen to T cells that share the same specificity. Here, we demonstrate that B cell clones engineered to express CD95 can effectively suppress a systemic primed antigen specific T cell response in vivo. Intravenous injection of antigen-pulsed FasL expressing B cells eliminated antigen-specific (TCR transgenic) T cells from the draining lymph nodes within 12-60 h, and suppressed a delayed-type hypersensitivity response in an antigen-specific manner. These results indicate that B cells can be engineered to express FasL, and used to impair T cell function in vivo, suggesting that FasL-expressing B cells may be an effective tool for the treatment of established T cell-mediated autoimmune and inflammatory diseases. PMID- 12115652 TI - A reversible functional defect of CD8+ T lymphocytes involving loss of tetramer labeling. AB - We have observed that human CTL clones lose their specific cytolytic activity and cytokine production under certain stimulation conditions, while retaining an antigen-dependent growth pattern. These inactive CTL simultaneously lose their labeling by an HLA-peptide tetramer, even though the amount of TCR-CD3 at their surface is not reduced. The tetramer-negative cells recover tetramer staining and cytolytic activity after stimulation with tumor cells in the presence of a supernatant of activated lymphocytes. Our results suggest the existence of a new type of functional defect of CTL. They also indicate that tetramers may fail to reveal some CTL bearing the relevant TCR, even when such functionally arrested CTL retain the potential to participate in immune responses because their defect is reversible. PMID- 12115653 TI - Plasma extravasation induced by dietary supplemented histamine in histamine-free mice. AB - Histidine decarboxylase (HDC) synthesizes endogenous histamine from histidine in mammals. To evaluate the role of histamine in skin allergic reaction, we used HDC gene knockout mice lacking histamine. No plasma extravasation reaction was observed in HDC-/- mice after passive cutaneous anaphylaxis (PCA) test. Compound 48/80, a mast cell granule depletor, produced plasma extravasation inHDC+/+ mice but no extravasation in HDC-/- mice. Interestingly, orally administered histamine was distributed in the skin in HDC-/- mice and in these histamine-supplemented mice the plasma extravasation reaction was observed after the injection of compound 48/80 and the PCA test. Cultured bone marrow-derived mast cells of HDC-/ mice took up histamine from the histamine-supplemented medium into the secretory granules. The absorbed histamine was released in response to the same antigen and antibody combination used as in PCA test. In contrast to the immediate-type response, the delayed-type hypersensitive response, observed as a thickening of the ear skin after trinitrochlorobenzene challenge (following sensitization), showed no differences between HDC+/+ and HDC-/- mice. Therefore, among the allergic skin reactions, histamine is revealed to be an important mediator especially for the plasma extravasation in an immediate-type allergy model. PMID- 12115654 TI - Binding immune-stimulating oligonucleotides to cationic peptides from viral core antigen enhances their potency as adjuvants. AB - Priming specific Th1 immunity by recombinant hepatitis B core antigen (HBcAg) depends on its arginine (Arg)-rich, 34-36-residue-long C terminus, and nucleotides bound to it. This adjuvant activity intrinsic to HBcAg facilitates priming of Th1 immunity to co-delivered, unrelated antigens, such as hepatitis B surface antigen (HBsAg), or ovalbumin (OVA) that prime specific Th2 immunity when injected without adjuvants. We loaded immune-stimulating, CpG-containing oligodeoxynucleotides (ODN) to the HBcAg-derived Arg-rich peptides C-1 (HBcAg(150 157), RRRDRGRS) or C-2 (HBcAg(164-179), SPRRRRSQSPRRRRSQ). When these peptide/nucleotide complexes were co-injected into mice with HBsAg, hepatitis B precore antigen (HBeAg) or OVA, the animals developed strikingly higher serum IgG2 antibody titers and cytotoxic T lymphocyte responses than animals co injected with these antigens and 'free' (not peptide-bound) ODN. Potent Th1 promoting adjuvants can thus be synthesized that mimic priming of anti-viral immunity by natural nucleocapsid particles. PMID- 12115655 TI - Granulocyte colony-stimulating factor attenuates LPS-stimulated IL-1beta release via suppressed processing of proIL-1beta, whereas TNF-alpha release is inhibited on the level of proTNF-alpha formation. AB - In the presence of granulocyte colony-stimulating factor (G-CSF), the release of IL-1beta and TNF-alpha by LPS-stimulated human whole blood was suppressed. Via measurement of cytokine mRNA, inactive precursor and mature protein, we investigated whether this inhibition occurs at the transcriptional, translational or post-translational level of cytokine production. G-CSF inhibited IL-1beta release, but the formation of proIL-1beta was not attenuated, indicating that G CSF interferes with the proteolytic processing of proIL-1beta. Since the release of IL-1beta in LPS-stimulated whole blood was blocked by the caspase-1 inhibitor YVAD-cmk, processing of proIL-1beta appears to depend on caspase-1 activity. The conclusion that G-CSF inhibits caspase-1 activity was supported bythe finding that the release of IL-18 was also inhibited by G-CSF, similar to IL-1beta release. Intracellular caspase-1 activity in monocytes was measured by flow cytometry with the cell-permeablecaspase substrate Asp(2)-rhodamine. In the presence of G-CSF the cleavage of this substrate was inhibited by more than 50%. G-CSF had no effect on LPS-induced doubling of caspase-1 mRNA, indicating that G CSF affects caspase-1 activation and not its formation. For TNF-alpha another mechanism of G-CSF action was identified: TNF-alpha as well as proTNF-alpha formation were inhibited by G-CSF, butG-CSF had no influence on LPS-induced TNF alpha mRNA level. We therefore suggest that G-CSF causes translational silencing of LPS-induced TNF-alpha mRNA. PMID- 12115656 TI - Direct binding of C1q to apoptotic cells and cell blebs induces complement activation. AB - Deficiency of early components of the classical pathway of complement, particularly C1q, predisposes to the development of systemic lupus erythematosus. Several studies have suggested an association between the classical complement pathway and the clearance of apoptotic cells. Mice with a targeted deletion of the C1q gene develop a lupus-like renal disease, which is associated with the presence of multiple apoptotic bodies in the kidney. In the present study we demonstrate that highly purified C1q binds to apoptotic cells and isolated blebs derived from these apoptotic cells. Binding of C1q to apoptotic cells occurs via the globular heads of C1q and induces activation of the classical complement pathway, as shown by the deposition of C4 and C3 on the surface of these cells and on cell-derived blebs. In addition, for the first time, we demonstrate that surface-bound C1q is present on a subpopulation of microparticles isolated from human plasma. Taken together, these observations demonstrate that C1q binds directly to apoptotic cells and blebs derived therefrom and support a role for C1q, possibly in concert with C4 and C3, in the clearance of apoptotic cells and blebs by the phagocytic system. PMID- 12115658 TI - Identification and expression of Helios, a member of the Ikaros family, in the Mexican axolotl: implications for the embryonic origin of lymphocyte progenitors. AB - Transcription factors of the Ikaros gene family are critical for the differentiation of T and B lymphocytes from pluripotent hematopoietic stem cells. To study the first steps of lymphopoiesis in the Mexican axolotl, we have cloned the Helios ortholog in this urodele amphibian species. We demonstrated that the axolotl Helios contains a 144-bp deletion at the 5' end of the activation domain. Helios is expressed in both the thymus and spleen but not in the liver of the pre adult axolotl. During ontogeny, Helios transcripts are detected from neurula stage, before the apparition of the first Ikaros transcripts and the colonization of lymphoid tissues. Interestingly, Helios and Ikaros mRNA are found predominantly in the ventral blood islands of late tail-bud embryos. These results suggest that in contrast to the Xenopus and amniote embryos where two sites of hematopoiesis have been characterized, the ventral blood islands could be the major site of hematopoiesis in the axolotl. PMID- 12115657 TI - CD8+ T cell apoptosis induced by Escherichia coli heat-labile enterotoxin B subunit occurs via a novel pathway involving NF-kappaB-dependent caspase activation. AB - The B subunit of Escherichia coli heat-labile enterotoxin (EtxB) is a potent immunomodulatory molecule capable of treating and preventing autoimmune disease. These properties result from its ability to bind to glycolipid receptors, principally G(M1) ganglioside, and modulate immune cell function. EtxB receptor binding causes B cell activation, modulates monocyte cytokine secretion and triggers apoptosis of CD8+ T cells. These wide-ranging effects suggest that B subunit receptor interaction triggers signaling events affecting cellular differentiation. We have investigated the processes by which EtxB induces CD8+ T cell apoptosis. We show that receptor interaction by EtxB activates caspase-3 in CD8+ but not in CD4+ T cells. Inhibition of caspase-3 blocks the apoptotic process. EtxB induces the activation of NF-kappaB in both CD8+ and CD4+ T cells. The findings that (i) SN50, a peptide inhibitor of NF-kappaB nuclear translocation, prevents caspase-3 activation and subsequent apoptosis, and (ii) CD8+CD4- thymocytes from transgenic mice expressing a dominant-negative form of the IkappaBalpha protein were markedly less susceptible to EtxB-induced apoptosis than cells from wild-type mice, indicate that NF-kappaB is important in the induction of the apoptotic pathway. Further investigations revealed that while caspase-8 activity is detected concomitant to caspase-3, caspase-9 activation, following mitochondrial cytochrome c release, is detectable later on. These observations are consistent with death receptor-mediated signaling, however, experiments using lpr/lpr and p55 TNFR -/- mice rule out the involvement of Fas and the p55 TNF receptor, respectively. The data therefore indicate that EtxB mediated apoptosis occurs via a novel pathway involving NF-kappaB. PMID- 12115659 TI - Immunosuppressive activity of capsaicinoids: capsiate derived from sweet peppers inhibits NF-kappaB activation and is a potent antiinflammatory compound in vivo. AB - Capsiate and its dihydroderivatives are the major capsaicinoids of sweet pepper. These new capsaicinoids do not activate the vanilloid receptor type 1 (VR1) but they share with capsaicin (CPS)some biological activities mediated in a VR1 independent fashion. In this study we show that CPS and nordihydrocapsiate (CPT) inhibit early and late events in T cell activation, including CD69, CD25 and ICAM 1 cell surface expression, progression to the S phase of the cell cycle and proliferation in response to TCR and CD28 co-engagement. Moreover, both CPS and CPT inhibit NF-kappaB activation in response to different agents including TNF alpha. CPS itself does not affect the DNA-binding ability of NF-kappaB but it prevents IkappaB kinase activation and IkappaBalpha degradation in a dose dependent manner, without inhibiting the activation of the mitogen-activated protein kinases, p38, extracellular regulated kinase and c-Jun N-terminal protein kinase. Moreover, intraperitoneal pretreatment with CPT prevented mice from lethal septic shock induced by lipopolysaccharide. In a second model of inflammation CPT pretreatment greatly reduced the extensive damage in the glandular epithelium observed in the bowel of DSS-treated mice. Taken together, these results suggest that CPT and related synthetic analogues target specific pathways involved in inflammation, and hold considerable potential for dietary health benefits as well as for pharmaceutical development. PMID- 12115660 TI - T helper differentiation in resistant and susceptible B7-deficient mice infected with Leishmania major. AB - The contribution of the costimulatory molecules B7-1 and B7-2 to the in vivo differentiation of Th cells remains controversial. The infection of resistant and susceptible strains of mice with the parasite Leishmania major provides a well established model for studying in vivo differentiation of CD4+ T cells. We have infected B7-1/B7-2-deficient mice on the BALB/c and 129 background with L. major and subsequently examined different parameters of infection and cytokine responses upon restimulation of lymph node cells in vitro. BALB/c B7-2-deficient and B7-1/B7-2-double deficient mice are resistant to L. major, whereas BALB/c B7 1-deficient mice remain as susceptible as wild-type BALB/c mice. Differential expression of B7-1 and B7-2 can explain the distinct roles observed for these B7 costimulators in L. major infection. PMID- 12115661 TI - Reversal of tolerance induced by transplantation of skin expressing the immunodominant T cell epitope of rat type II collagen entitles development of collagen-induced arthritis but not graft rejection. AB - Collagen-induced arthritis (CIA) is induced in H-2(q) mice after immunization with rat type II collagen (CII). The immunodominant T cell epitope on heterologous CII has been located to CII256-270. We have previously shown that TSC transgenic mice, which express the heterologous epitope in type I collagen (CI), e.g. in skin, are tolerized against rat CII and resistant to CIA. In this study we transplanted skin from TSC transgenic mice onto non-transgenic CIA susceptible littermates to investigate whether introduction of this epitope to a naive immune system would lead to T cell priming and graft rejection or instead to tolerance and arthritis protection. Interestingly, TSC grafts were accepted and not even immunization of recipient mice with CII in adjuvant induced graft rejection. Instead, TSC skin recipients displayed a reduced T and B cell response to CII and were also protected from arthritis. However, additional priming could break arthritis protection and was accompanied by an increased T cell response to the grafted epitope. Strikingly, despite the regained T cell response, development of arthritis was not accompanied by graft rejection, showing that these immune-mediated inflammatory responses involve different mechanisms. PMID- 12115662 TI - Neutralization of IFN-inducible protein 10/CXCL10 exacerbates experimental autoimmune encephalomyelitis. AB - We examined the effect of a monoclonal antibody (mAb) against interferon (IFN) inducible protein 10 (IP-10)/CXCL10 on the development of experimental autoimmune encephalomyelitis (EAE) in rats induced by injecting xenogeneic brain homogenates into footpads. Treatment with neutralizing mAb against CXCL10 exacerbated EAE with increased infiltrating CD4+ cells in the central nervous system. Furthermore, the exacerbation by the mAb treatment was accompanied by less enlarged draining popliteal lymph nodes (LN) in parallel with cell number compared with those of EAE rats treated with control mAb, whereas other lymphoid organs such as the spleen and thymus were not significantly different between rats treated with anti-CXCL10 and the control mAb. Induction of gene expression of CXCL9/Mig and CXCL10 and their receptor CXCR3 was confirmed in the draining LN in EAE rats. Induction of the third CXCR3 ligand, CXCL11/I-TAC was not seen in the draining LN, whereas all three CXCR3 ligands and CXCR3 itself were markedly detected in the spinal cords following the development of EAE. These findings suggest that CXCL10 produced in the LN plays a specific inhibitory role in the development of Th1-mediated diseases such as EAE by holding sensitized and activated Th1s expressing CXCR3 in the draining LN. PMID- 12115663 TI - Induction of the chemokine receptor CXCR3 on TCR-stimulated T cells: dependence on the release from persistent TCR-triggering and requirement for IFN-gamma stimulation. AB - The chemokine receptor CXCR3 has been shown to play a key role in the recruitment of T cells to sites of inflammation such as allografts. Here, we investigated which signals and conditions areresponsible for CXCR3 induction. CXCR3 was induced on T cells that were stimulated with anti-CD3 plus anti-CD28 monoclonal antibodies and then recultured without any external stimuli. CXCR3 expression was inhibited when TCR stimulation was persistent in the reculture. CXCR3 induction also depended on the stimulation with IFN-gamma because CXCR3 expression was not induced in IFN-gamma-deficient T cells. The induction of another Th1 chemokine receptor CCR5 absolutely required IL-12 stimulation and STAT4 involvement. In contrast, CXCR3 was induced on STAT4-deficient T cells independently of IL-12 stimulation as long as IFN-gamma was produced as a result of potent TCR stimulation. These results show that CXCR3 induction on TCR-triggered T cells requires the release of these T cells from persistent TCR signaling and the stimulation with IFN-gamma and also indicate the differential regulatory mechanisms underlying the induction of two Th1 chemokine receptors. PMID- 12115664 TI - Differential activation of human and guinea pig complement by pentameric and hexameric IgM. AB - Human and mouse IgM can be polymerized as a hexamer in addition to a pentamer. Our previous work with mouse IgM measured activation of guinea pig complement by highly enriched preparations of hexamer and pentamer and showed that hexamer is >100-fold more active than pentamer. In this report pentamer and hexamer were compared for their capacity to activate complement in a homogeneic system, i.e. chimeric mouse V/human Cmu IgM pentamer and hexamer were assayed separately for their capacity to activate human (and guinea pig) complement. In both the homogeneic and the xenogeneic systems hexamer was more active than pentamer, but the magnitude of the difference between hexamer and pentamer depended on the complement source. Whereas chimeric hexamer activated guinea pig complement >100 fold more efficiently than did chimeric pentamer, this hexamer was only 4-13-fold more active than pentamer when assayed with human complement. Similarly, mouse hexamer, which was >100-fold more active than mouse pentamer with guinea pig complement, was only approximately 2-fold more active than mouse pentamer with human complement. Mouse hexameric and pentameric IgM were each approximately 20 fold more active with human complement than were the corresponding chimeric isoforms of IgM. PMID- 12115665 TI - CD5-mediated inhibition of TCR signaling during intrathymic selection and development does not require the CD5 extracellular domain. AB - CD5 functions as a negative regulator of TCR signaling during intrathymic T cell development, but it is not known if this negative regulatory function requires CD5 engagement of an extracellular ligand. The present study has specifically examined the role of the CD5 extracellular domain in T cell development by introducing into CD5-/- mice a chimeric CD5 molecule in which the extracellular domain of CD5 is replaced with the extracellular domain of human IL-2R p55 (Tac) for which no ligand exists in the mouse. We now report that CD5 mediated down regulation of TCR signaling during thymocyte development does not require the CD5 extracellular domain and, consequently, does not involve CD5 binding of an extracellular ligand in the thymus. PMID- 12115667 TI - Neurochemistry of nerve fibers apposing sympathetic preganglionic neurons activated by sustained hypotension. AB - Sympathetic preganglionic neurons (SPN) in rat spinal cord were activated by the reflex stimulation of bulbospinal sympathetic neuronal pathways after a nitroprusside-induced hypotension. Hypotension-sensitive SPN, identified by immunoreactivity (IR) to the product of the immediate early gene c-fos and to choline acetyltransferase, were localized in the intermediolateral cell column of thoracic and upper lumbar cord, particularly middle to lower thoracic cord. Putative neurotransmitters, or their markers, in varicose fiber networks around SPN were identified. Nearly all hypotension-sensitive (Fos-IR) SPN were apposed by varicose fibers immunoreactive for tyrosine hydroxylase, serotonin, substance P, or enkephalin. Neuropeptide Y (NPY)- or phenylethanolamine-N-methyl transferase (PNMT)-IR varicose fibers apposed Fos-IR SPN in the upper and middle thoracic spinal cord, but in lower thoracic segments some Fos-IR SPN lacked these appositions. In thoracic segment 12, 51% +/- 5% of Fos-IR SPN (n = 9 rats) lacked PNMT contacts and 25% +/- 3% of Fos-IR SPN (n = 8 rats) lacked NPY contacts. In contrast to other chemically defined afferents, galanin-IR varicose fibers apposed fewer than half of the Fos-IR SPN in the middle to lower thoracic cord. Neurotransmitters/neuromodulators that might influence the activity of SPN acting in the baroreflex-mediated control of blood pressure have been identified. Uniformity in the neurochemistry of some fibers making connections with Fos-IR SPN, regardless of their segmental origin, suggests that common sets of neurons provide convergent inputs to all hypotension-sensitive SPN. Other fibers show topographic differences in their contacts with Fos-IR SPN, suggesting that subgroups of hypotension-sensitive SPN are targeted by particular neuron groups. PMID- 12115668 TI - Distribution of myelinated and unmyelinated nerve fibers and their possible role in blood flow control in crotaline snake infrared receptor organs. AB - We used transmission electron microscopic montages to examine the composition of nerve bundles serving the infrared pit organs of two species of crotaline snakes, Agkistrodon blomhoffii and A. brevicaudus. In the three main bundles, the myelinated fibers totaled 2,200-3,700, and unmyelinated fibers 2,400. We also discovered for the first time two accessory bundles composed almost entirely of unmyelinated fibers running alongside the main bundles, containing an average total of 3,300 unmyelinated fibers vs. an average of 10 myelinated fibers. Thus, the average total of unmyelinated fibers was nearly twice that of myelinated fibers. To study the nature of the unmyelinated fibers, we did double staining immunohistochemistry with antibodies for substance P (SP) and vasoactive intestinal polypeptide (VIP) in combination with and without capsaicin pretreatment. SP and VIP immunoreactive varicose fibers ran straight toward the center of the pit membrane in parallel with arterioles and venules, and also formed a dense network around the periphery of the membrane. There were three types of fibers: fibers containing only SP, fibers containing only VIP, and fibers containing both peptides. SP-only fibers were distributed singly throughout the pit membrane and in small bundles around the periphery. SP+VIP fibers were distributed sparsely in the pit membrane and around its periphery. VIP-only fibers were distributed throughout the pit membrane and were of smaller diameter than SP and SP+VIP fibers. After treatment with capsaicin, most of the three types of varicose fibers disappeared from the central part of the pit membrane, but those around the periphery remained unaffected. The capsaicin sensitive fibers may be unmyelinated sensory types, and the unaffected ones may be autonomic nerve fibers. PMID- 12115669 TI - Differential localization of high- and low-molecular-weight variants of microtubule-associated protein 2 in the developing rat telencephalon. AB - Microtubule-associated protein 2 (MAP2) occurs in developing mammalian neuronal tissue as both high- and low-molecular-weight forms with temporally regulated expression. We studied the MAP2 expression in the developing rat telencephalon with monoclonal antibodies that recognized both the high- and low-molecular weight forms of MAP2 variants or that specifically recognized high-molecular weight forms of MAP2 variants. Differences in the staining patterns of these antibodies reflected differences in the distribution of the high- and low molecular-weight MAP2s. The immunoreactive sites of high- and low-molecular weight MAP2 had a more widespread distribution in the embryonic telencephalon than those of high-molecular-weight MAP2. Many bipolar cells in the ganglionic eminence (GE) and in the intermediate zone (IZ) of the neocortex showed low molecular-weight MAP2 immunoreactivity, but they showed weak or no high-molecular weight MAP2 immunoreactivity. Expression of mRNA containing exons common to high- and low-molecular-weight MAP2 was detected in the tangentially ellipsoidal cells in the IZ, but expression of mRNA containing an exon specific to high-molecular weight MAP2 was not detected in these cells by in situ hybridization. We interpreted these observations as indicating that the bipolar cells contained MAP2c preferentially, but contained MAP2a and MAP2b (MAP2a/b) at a very low or negligible level. The cells that expressed MAP2c preferentially among the MAP2 splicing variants composed 50% of the preplate cells, most of the MAP2-positive cells in the hippocampus and the corpus callosum. Double labeling by DiI staining and Dlx2 immunohistochemistry, or by Dlx2 and MAP2 immunohistochemistry, revealed that most of the Dlx2-positive cells in the IZ expressed MAP2c preferentially at embryonic day 16. Another double-labeling study revealed that most GAD-positive cells in the preplate were MAP2a/b positive, whereas most GAD-positive cells in the IZ expressed MAP2c preferentially, with only a negligible level of MAP2a/b immunoreactivity. We conclude that MAP2 immunoreactivity in the IZ was localized in the tangentially migrating neurons. The tangentially migrating neurons seemed to acquire MAP2a/b immunoreactivity as they entered the preplate or cortical plate and developed into mature neurons. Radially migrating neurons in the IZ were MAP2 negative. After entering to the preplate or the cortical plate, they became MAP2a/b positive as they developed into mature neurons. PMID- 12115670 TI - Neuroepithelial bodies: a morphologic substrate for the link between neuronal nitric oxide and sensitivity to airway hypoxia? AB - Currently, the significance of nitric oxide (NO) in the respiratory tract is a matter of great interest because NO is believed to play a major role in the physiological regulation of airway function but also in lung pathology. What is especially intriguing with respect to the present investigation, are reports that the pulmonary expression of neuronal NO synthase (nNOS) is altered as a result of airway hypoxia. We examined the possible relationship between intrapulmonary nitrergic structures and pulmonary neuroepithelial bodies (NEBs), chemoreceptor like epithelial cell groups that are known to have all necessary components for oxygen perception. Tyramide-enhanced immunostaining for nNOS was combined with known markers for NEBs in an ontogenetic study of rat lungs. From postnatal day 2 onward, nNOS-immunoreactive (-IR) neuronal cell bodies, present mainly in the lamina propria at all levels of intrapulmonary airways, were seen to give rise to remarkable intraepithelial terminal arborizations that invariably colocalized with NEBs. nNOS immunoreactivity was absent from the vagal calbindin D28k(CB) -IR and the spinal calcitonin gene-related peptide(CGRP) -IR extrinsic sensory nerve fiber populations that our group reported earlier to selectively contact NEBs. Quantitative analysis showed that all NEBs receiving nNOS-IR terminals were also contacted by CGRP-IR nerve fibers, whereas approximately 55% were additionally contacted by CB-IR nerves. The reported nitrergic neurons did not express the cholinergic marker vesicular acetylcholine transporter and were always surrounded by a basket of CGRP-IR nerve terminals. In conclusion, part of the pulmonary NEBs selectively receive extensive nitrergic nerve terminals that originate from intrinsic neurons. Together with literature data on lung physiology and pharmacology, some interesting suggestions for the functional significance of the association between pulmonary CGRP-IR NEBs, nNOS-IR neurons, and CGRP-IR afferents described in the present study, are discussed. PMID- 12115671 TI - Bladder contractility-related neurons in Barrington's nucleus: axonal projections to the spinal cord in the cat. AB - Barrington's nucleus projects directly to the sacral parasympathetic nucleus. The purpose of this study was to clarify whether neurons in Barrington's nucleus that increase their firing during bladder contractions project to the spinal cord and, if so, to which level(s) the axon reaches. Single units were recorded in Barrington's nucleus of cat with glass microelectrodes, and the termination level of descending axons was determined by antidromic stimulation of the spinal cord. Thirty-nine neurons projecting to the spinal cord were located in rostral parts of the dorsolateral pontine tegmentum, medial and ventral to the mesencephalic trigeminal tract. This finding is consistent with previous neuronal tracing studies. All neurons increased their firing rates during contraction associated with micturition. In 19 examined neurons, the most caudal level of the descending axon distributed between the L7 and the S3 level. Stimulation of the axon at this most caudal level resulted in antidromic spike latencies ranging between 19.5 msec and 45.0 msec. These antidromic latencies were much smaller than previously reported orthodromic conduction times between neurons in Barrington's nucleus and sacral preganglionic neurons innervating the bladder. The mean conduction velocity of the descending axon from the cell body to the border between Th13 and the L1 ranged between 7.2 m/sec and 27.7 m/sec. The decrease of the mean conduction velocity was observed at the lumbar as well as at the sacral segments, suggesting that axons issue collaterals to the lumbar level as well as to the sacral level. PMID- 12115673 TI - Localization of tyrosine hydroxylase and its messenger RNA in the brain of rainbow trout by immunocytochemistry and in situ hybridization. AB - This report describes the distribution of tyrosine hydroxylase (TH)-expressing structures in the brain of rainbow trout (Oncorhynchus mykiss). TH neurons have been localized by the use of two complementary techniques, immunocytochemistry and in situ hybridization of TH messenger RNA. Results obtained from in situ hybridization and immunocytochemistry were in agreement. TH cells were observed in many areas of the brain, with a higher density at the level of the olfactory bulbs where TH-positive neurons are abundant in the internal cell layer. In the telencephalon, two populations of TH neurons can be distinguished: one group is located in the area ventralis telencephali pars dorsalis, and the other group is located in the area ventralis telencephali pars ventralis and extends laterally in the area ventralis telencephali pars lateralis. Many labeled neurons are also seen in the preoptic area as well as in the hypothalamus, where several clusters of TH-positive cells are observed. Some of these neurons located in the paraventricular organ grow a short cytoplasmic extension directed to the ventricular wall and are known to be cerebrospinal fluid-contacting cells. The most caudal TH neurons are observed at the level of the locus caeruleus. At the level of the pituitary, TH-positive fibers are observed in the neurohypophysis. The TH-immunoreactive innervation at the level of the pituitary provides a neuroanatomic basis for the effects of dopamine and/or norepinephrine on the release of pituitary hormones in fish. PMID- 12115672 TI - Combination of gonadal steroid treatment and peripheral nerve grafting results in a peripheral motoneuron-like pattern of beta II-tubulin mRNA expression in axotomized hamster rubrospinal motoneurons. AB - Rubrospinal motoneurons (RSMN) represent a population of androgen receptor containing central motoneurons in rodents. In this study, the ability of testosterone propionate (TP), alone or in conjunction with a peripheral nerve graft (PNG), to alter the molecular program of injured RSMN was accomplished using betaII-tubulin cDNA probes and quantitative in situ hybridization (ISH). Initial fluoro-gold labeling experiments following a T1 hemisection established that, as in the rat, the hamster rubrospinal system is essentially crossed and that injured RSMN concentrate in the ventrolateral region of the red nucleus. In the second experimental series, adult gonadectomized male hamsters were subjected to a right T1 hemisection, with half of the operated animals immediately subcutaneously implanted with 1 10 mm TP Silastic capsule and the other half sham implanted. In a third experimental series, animals were subjected to T1 hemisection, followed by transplantation of a predegenerated autologous segment of peripheral nerve. Half of the animals in each group received TP implants at the time of spinal cord injury and PNG. Postoperative times were 2, 7, and 14 days (dpo). Quantitative ISH was performed using a betaII-tubulin-specific (33)P labeled cDNA probe, emulsion autoradiography, and computerized image analysis for grain counting. Injury alone resulted in a short-lived increase in betaII-tubulin mRNA expression in the RSMN at 2 dpo, with a significant decline to well below control values at 7 and 14 dpo. TP treatment or PNG alone attenuated, but did not prevent, the down-regulation of betaII-tubulin mRNA. In contrast, the combination of TP with a PNG sustained the injury-induced increase in betaII-tubulin mRNA levels throughout the postoperative period of 2, 7, and 14 dpo. The synergistic effects of the two treatment strategies confirm the importance of targeting multiple aspects of the injury response for therapeutic intervention. PMID- 12115674 TI - Distribution of neuropeptide Y Y1 receptors in rodent peripheral tissues. AB - Using a sensitive immunohistochemical technique, the localization of neuropeptide Y (NPY) Y1-receptor (Y1R)-like immunoreactivity (LI) was studied in various peripheral tissues of rat. Wild-type (WT) and Y1R-knockout (KO) mice were also analyzed. Y1R-LI was found in small arteries and arterioles in many tissues, with particularly high levels in the thyroid and parathyroid glands. In the thyroid gland, Y1R-LI was seen in blood vessel walls lacking alpha-smooth muscle actin, i.e., perhaps in endothelial cells of capillaries. Larger arteries lacked detectable Y1R-LI. A distinct Y1R-immunoreactive (IR) reticulum was seen in the WT mouse spleen, but not in Y1R-KO mouse or rat. In the gastrointestinal tract, Y1R-positive neurons were observed in the myenteric plexus, and a few enteroendocrine cells were Y1R-IR. Some cells in islets of Langerhans in the pancreas were Y1R-positive, and double immunostaining showed coexistence with somatostatin in D-cells. In the urogenital tract, Y1R-LI was observed in the collecting tubule cells of the renal papillae and in some epithelial cells of the seminal vesicle. Some chromaffin cells of adrenal medulla were positive for Y1R. The problem of the specificity of the Y1R-LI is evaluated using adsorption tests as well as comparisons among rat, WT mouse, and mouse with deleted Y1R. Our findings support many earlier studies based on other methodologies, showing that Y1Rs on smooth muscle cells of blood vessels mediate NPY-induced vasoconstriction in various organs. In addition, Y1Rs in other cells in parenchymal tissues of several organs suggest nonvascular effects of NPY via the Y1R. PMID- 12115675 TI - Complementary and layered expression of Ephs and ephrins in developing mouse inner ear. AB - The distributions of the Eph-class receptors EphA4 and EphB1, and their ligands ephrin-A2, ephrin-B1, and ephrin-B2, were analysed by immunostaining in the mouse inner ear. Complementary patterns of EphA4 and its potential ligand ephrin-A2 were found, with ephrin-A2 in many of the structures lining the cochlear duct and within the cochlear nerve cells, and EphA4 in the deeper structures underlying the cochlear duct and in the cells lining the nerve pathway. EphB1 and its potential ligands ephrin-B1 and ephrin-B2 showed a segregated layered expression in the lateral wall of the cochlear duct (the external sulcus), which together with EphA4 expressed in the area, form a four-layered structure with an alternating pattern of receptors and ligands in the different layers. This arrangement gives the potential for different bidirectional Eph-mediated interactions between each of the layers. The results suggest that the Eph system in the cochlea may have a role in maintaining cell segregation during phases of cochlear development. PMID- 12115676 TI - Up-regulation of tyrosine kinase (Trka, Trkb) receptor expression and phosphorylation in lumbosacral dorsal root ganglia after chronic spinal cord (T8 T10) injury. AB - Previous studies have demonstrated changes in urinary bladder neurotrophic factors after bladder dysfunction. We have hypothesized that retrograde transport of neurotrophin(s) from the bladder to lumbosacral dorsal root ganglia (DRG) may play a role in bladder reflex reorganization after spinal cord injury (SCI). In this study, we determined whether the expression of tyrosine kinase receptors (TrkA, TrkB) is altered in lumbosacral DRG after SCI through immunofluorescence techniques. Complete transection of the spinal cord (T8-T10) was performed in female Wistar rats (120-150 g), and animals were studied 5-6 weeks after SCI. One week before killing, Fast Blue (FB) was injected into the bladder to label bladder afferent cells in the L1, L2, L6, and S1 DRG. After SCI, a significant increase in the number of TrkA-immunoreactive (IR) positive cells was detected in the L6-S1 DRG (L6: 1.9-fold, P < or = 0.01; S1: 1.7-fold, P < or = 0.05) and in the L1 DRG (3.0-fold; P < or = 0.01) but not in the L4-L5 DRG compared with spinal-intact (control) rats. After SCI, a significant increase in the number of TrkB-IR cells was also detected in the L6-S1 DRG (L6: 2.2-fold, P < or = 0.01; S1: 1.5-fold, P < or = 0.05) and in the L1-L2 DRG (L1: 1.5-fold, P < or = 0.01; L2: 1.3-fold, P < or = 0.05) but not in the L4-L5 DRG compared with control rats. After SCI, the percentage of FB-labeled cells expressing TrkA immunoreactivity (approximately 68%) or TrkB immunoreactivity (approximately 65%) in L1 and L6 DRG significantly (P < or = 0.01) increased compared with control (20-30%) DRG. After SCI, the percentage of TrkA-IR cells expressing phosphorylated (p)-Trk immunoreactivity significantly increased (1.5- to 2.3-fold increase) in the L1, L6, and S1 DRG. The percentage of TrkB-IR cells expressing p-Trk immunoreactivity after SCI also increased (1.3-fold increase) in the L1 and L6 DRG. These results demonstrate that (1) TrkA and TrkB immunoreactivity is increased in bladder afferent cells after SCI and (2) TrkA and TrkB receptors are phosphorylated in DRG after SCI. Neuroplasticity of lower urinary tract reflexes after SCI may be mediated by both nerve growth factor and brain-derived neurotrophic factor. PMID- 12115677 TI - Differential expression of neuronal calcium sensor-1 in the developing chick retina. AB - Neuronal calcium sensor-1 (NCS-1) is a Ca(2+) binding protein that has been implicated in the regulation of neurotransmission and synaptogenesis. In this study we investigated the developmental expression and localization of NCS-1 in the chick retina. Single- and double-labeling experiments with three-dimensional reconstruction as well as ultrastructural data of the distribution of NCS-1 suggest that this protein is also involved in axonal process outgrowth. We found an early expression of NCS-1 in ganglion cells and their axons, in amacrine, and in horizontal cells, whereas photoreceptors were immunonegative at embryonic stages. In the early posthatching days we found strong immunostaining for NCS-1 in horizontal cells and their processes in the outer plexiform layer. In contrast, synaptic vesicle protein 2 (SV2) was prominent only in photoreceptor synaptic terminals. Ultrastructural analysis confirmed that NCS-1 was localized postsynaptically in horizontal cell processes, whereas presynaptic terminals were immunonegative. However, at late posthatching days we observed that photoreceptor ribbon synapses (from rods and/or cones) also expressed NCS-1. Thus the results support the notion that NCS-1 is involved in neuronal process outgrowth and is localized in pre- and postsynaptic compartments including mature photoreceptor synapses. PMID- 12115678 TI - Cellular localization and development of neuronal intranuclear inclusions in striatal and cortical neurons in R6/2 transgenic mice. AB - The cellular localization and development of neuronal intranuclear inclusions (NIIs) in cortex and striatum of R6/2 HD transgenic mice were studied to ascertain the relationship of NIIs to symptom formation in these mice and gain clues regarding the possible relationship of NII formation to neuropathology in Huntington's disease (HD). All NIIs observed in R6/2 mice were ubiquitinated, and no evidence was observed for a contribution to them from wild-type huntingtin; they were first observed in cortex and striatum at 3.5 weeks of age. In cortex, NIIs increased rapidly in size and prevalence after their appearance. Generally, cortical projection neurons developed NIIs more rapidly than cortical interneurons containing calbindin or parvalbumin. Few cortical somatostatinergic interneurons, however, formed NIIs. In striatum, calbindinergic projection neurons and parvalbuminergic interneurons rapidly formed NIIs, but they formed more gradually in cholinergic interneurons, and few somatostatinergic interneurons developed NIIs. Striatal NIIs tended to be smaller than those in cortex. The early accumulation of NIIs in cortex and striatum in R6/2 mice is consistent with the early appearance of motor and learning abnormalities in these mice, and the eventual pervasiveness of NIIs at ages at which severe abnormalities are evident is consistent with their contribution to a neuronal dysfunction underlying the abnormalities. That cortex develops larger NIIs than striatum, however, is inconsistent with the preferential loss of striatal neurons in HD but is consistent with recent evidence of early morphological abnormalities in cortical neurons in HD. That calbindinergic and parvalbuminergic striatal neurons develop large NIIs is consistent with a contribution of nuclear aggregate formation to their high degree of vulnerability in HD. PMID- 12115679 TI - Gamma-aminobutyric acidB receptors and the development of the ventromedial nucleus of the hypothalamus. AB - Gamma-aminobutyric acid (GABA) is a highly abundant neurotransmitter in the brain and the ligand for GABA(A), GABA(B), and GABA(C) receptors. Unlike GABA(A) and GABA(C) receptors, which are chloride channels, GABA(B) receptors are G-protein linked and alter cell-signaling pathways. Electrophysiological studies have found GABA(B) receptors in cultured embryonic hypothalamus, but the distribution of these receptors remains unknown. In the present study, we examined the expression of GABA(B) receptors in the ventromedial nucleus of the hypothalamus (VMH) during embryonic mouse development. GABA(B) receptors were present in the VMH at all ages examined, from embryonic day 13 to postnatal day 6. Using a brain slice preparation, we examined the effect of GABA(B) receptor activation on cell movement in the embryonic VMH as the nucleus forms in vitro. The GABA(B) receptor agonist baclofen decreased the rate of cell movement in a dose-dependent manner. Baclofen reduced cell movement by up to 56% compared with vehicle-treated controls. The percentage of cells moving per field and the angles of cell movement were not affected. With our previous findings of GABA(A) receptor activation, it is likely that GABA influences VMH development via multiple mechanisms. PMID- 12115680 TI - Evidence from V1 connections for both dorsal and ventral subdivisions of V3 in three species of New World monkeys. AB - We used patterns of connections of primary visual cortex (V1) to reevaluate differing proposals on the organization of extrastriate cortex in three species of New World monkeys. Several fluorescent tracers and the bidirectional tracer cholera toxin B subunit (CTB) were injected into dorsal V1 (representing the lower visual quadrant) and ventral V1 (representing the upper visual quadrant) of titi, squirrel, and owl monkeys. Labeled cells and terminals were plotted on brain sections cut parallel to the surface of flattened cortex and were related to architectonic boundaries. The results provided compelling evidence for both dorsal V3 with dorsal V1 connections and ventral V3 with ventral V1 connections. The connection pattern indicated that V3 represents the visual hemifield as a mirror image of V2. In addition, V3 could be recognized by a weak banding pattern in brain sections processed for cytochrome oxidase. V1 has connections with at least 12 subdivisions of visual cortex, with half of the connections involving V2 and 20% V3. Comparable results were obtained from all three species, suggesting that visual cortex is similarly organized. PMID- 12115681 TI - Primary afferent input to and receptive field properties of cells in rat lumbar area X. AB - In this study we examined the primary afferent input to rat area X of Rexed, and characterized sensory receptive fields (RFs) of the cells therein. This poorly understood area contains primary afferent fibres, some of which are arranged into a compact bundle beneath the central canal. Anterograde transport of the B fragment of cholera toxin (CTB) from the sciatic nerve showed a strictly ipsilateral projection to segments in L4 and L5 but both ipsi- and contralateral projections in L6 and more caudal segments. The response of cells in area X to mechanical cutaneous stimuli was recorded through extracellular microelectrodes in decerebrate, decerebrate-spinal, and urethane-anaesthetised preparations. The lateral edge of area X was marked by an abrupt change in the RFs: Lateral to area X in the dorsal horn, they were strictly unilateral and relatively small. At a mean of 90 microm from the midline, there was an abrupt expansion of the RFs to cover at least the entire ipsilateral dermatome. Within area X, 70% of the cells' RFs extended across the midline to include contralateral skin. In 35% of cells recorded in rats with intact spinal cords, the RF extended rostrally onto the forelimb. In a small number of cells, the RF included ear pinnae and nose. The precise function of area X cells remains unknown; although they have been shown to be involved in visceral reflexes, the fact that they receive convergent input from a wide variety of tissues and from local and remote body parts implies a more generalized, integrative function. PMID- 12115682 TI - Similarities and differences in the innervation of mystacial vibrissal follicle sinus complexes in the rat and cat: a confocal microscopic study. AB - Our confocal three-dimensional analyses revealed substantial differences in the innervation to vibrissal follicle-sinus complexes (FSCs) in the rat and cat. This is the first study using anti-protein gene product 9.5 (PGP9.5) immunolabeling and confocal microscopy on thick sections to examine systematically the terminal arborizations of the various FSC endings and to compare them between two species, the rat and the cat, that have similar-appearing FSCs but different exploratory behaviors, such as existence or absence of whisking. At least eight distinct endings were clearly discriminated three dimensionally in this study: 1) Merkel endings at the rete ridge collar, 2) circumferentially oriented lanceolate endings, 3) Merkel endings at the level of the ring sinus, 4) longitudinally oriented lanceolate endings, 5) club-like ringwulst endings, 6) reticular endings, 7) spiny endings, and 8) encapsulated endings. Of particular contrast, each nerve fiber that innervates Merkel cells at the level of the ring sinus in the rat usually terminates as a single, relatively small cluster of endings, whereas in the cat they terminate en passant as several large clusters of endings. Also, individual arbors of reticular endings in the rat ramify parallel to the vibrissae and distribute over wide, overlapping territories, whereas those in the cat ramify perpendicular and terminate in tightly circumscribed territories. Otherwise, the inner conical body of rat FSCs contains en passant, circumferentially oriented lanceolate endings that are lacking in the cat, whereas the cavernous sinus of the cat has en passant corpuscular endings that are lacking in the rat. Surprisingly, the one type of innervation that is the most similar in both species is a major set of simple, club-like endings, located at the attachment of the ringwulst, that had not previously been recognized as a morphologically unique type of innervation. Although the basic structure of the FSCs is similar in the rat and cat, the numerous differences in innervation suggest that these species would have different tactile capabilities and perceptions possibly related to their different vibrissa-related exploratory behaviors. PMID- 12115683 TI - Dopamine and nitric oxide control both flickering and steady-light-induced cone contraction and horizontal cell spinule formation in the teleost (carp) retina: serial interaction of dopamine and nitric oxide. AB - Adaptation to ambient light, which is an important characteristic of the vertebrate visual system, involves cellular and subcellular (synaptic) plasticity of the retina. The present study investigated dopamine (DA) and nitric oxide (NO) as possible neurochemical modulators controlling cone photomechanical movements (PMMs) and horizontal cell (HC) spinules in relation to steady and flickering light adaptation in the carp retina. Haloperidol (HAL; a nonspecific DA receptor blocker) or cPTIO (a NO scavenger) largely inhibited the cone PMMs and HC spinule formation induced by either steady or flickering light. These results suggested that both DA and NO could be involved in the light-adaptation changes induced by either pattern of input and that DA and NO effects may not be completely independent. The possibility that NO and DA interact serially was evaluated pharmacologically by cross-antagonist application (i.e., DA + cPTIO or NO + HAL). When a NO donor was coapplied with HAL to dark-adapted eyecups, normal light adaptive cone PMMs and HC spinules occurred. In contrast, when DA was applied in the presence of cPTIO, the dark-adapted state persisted. It was concluded 1) that DA and NO are both light-adaptive neurochemicals, released in the retina during either steady or flickering light; 2) that the effects of DA and NO on light adaptive cone PMMs and HC spinules do not occur in parallel; and 3) that NO and DA act mainly in series, specifically as follows: Light --> DA --> NO --> Cone PMMs + HC spinules. PMID- 12115684 TI - Association of the AMPA receptor-related postsynaptic density proteins GRIP and ABP with subsets of glutamate-sensitive neurons in the rat retina. AB - We used specific antibodies against two postsynaptic density proteins, GRIP (glutamate receptor interacting protein) and ABP (AMPA receptor-binding protein), to study their distribution in the rat retina. In the central nervous system, it has been shown that both proteins bind strongly to the AMPA glutamate receptor (GluR) 2/3 subunits, but not other GluRs, through a set of three PDZ domains. Western blots detected a single GRIP protein that was virtually identical in retina and brain, whereas retinal ABP corresponded to only one of three ABP peptides found in brain. The retinal distributions of GluR2/3, GRIP, and ABP immunoreactivity (IR) were similar but not identical. GluR2/3 immunoreactivity (IR) was abundant in both plexiform layers and in large perikarya. ABP IR was concentrated in large perikarya but was sparse in the plexiform layers, whereas GRIP IR was relatively more abundant in the plexiform layers than in perikarya. Immunolabel for these three antibodies consisted of puncta < or = 0.2 microm in diameter. The cellular localization of GRIP and ABP IR was examined by double labeling subclasses of retinal neuron with characteristic marker proteins, e.g., calbindin. GRIP, ABP, and GluR2/3 IR were detected in horizontal cells, dopaminergic and glycinergic AII amacrine cells and large ganglion cells. Immunolabel was absent in rod bipolar and weak or absent in cholinergic amacrine cells. By using the tyramide method of signal amplification, a colocalization of GluR2/3 was found with either GRIP or ABP in horizontal cell terminals, and perikarya of amacrine and ganglion cells. Our results show that ABP and GRIP colocalize with GluR2/3 in particular subsets of retinal neuron, as was previously established for certain neurons in the brain. PMID- 12115685 TI - Direct catecholaminergic-cholinergic interactions in the basal forebrain. III. Adrenergic innervation of choline acetyltransferase-containing neurons in the rat. AB - The central adrenergic neurons have been suggested to play a role in the regulation of arousal and in the neuronal control of the cardiovascular system. To provide morphological evidence that these functions could be mediated via the basal forebrain, we performed correlated light and electron microscopic double immunolabeling experiments using antibodies against phenylethanolamine N methyltransferase (PNMT) and choline acetyltransferase, the synthesizing enzymes for adrenaline and acetylcholine, respectively. Most adrenergic/cholinergic appositions were located in the horizontal limb of diagonal band of Broca, within the substantia innominata, and in a narrow band bordering the substantia innominata and the globus pallidus. Quantitative analysis indicated that cholinergic neurons of the substantia innominata receive significantly higher numbers of adrenergic appositions than cholinergic cells in the rest of the basal forebrain. In the majority of cases, the ultrastructural analysis revealed axodendritic asymmetric synapses. By comparing the number and distribution of dopamine beta-hydroxylase (DBH)/cholinergic appositions, described earlier, with those of PNMT/cholinergic interactions in the basal forebrain, it can be concluded that a significant proportion of putative DBH/cholinergic contacts may represent adrenergic input. Our results support the hypothesis that the adrenergic/cholinergic link in the basal forebrain may represent a critical component of a central network coordinating autonomic regulation with cortical activation. PMID- 12115686 TI - Sensory neuron addition in juvenile rat: time course and specificity. AB - The time course and specificity of neuron addition to lumbar dorsal root ganglia (DRGs) L(4)-L(6) of rats was investigated. By using methods validated by three dimensional reconstructions, profile counts in paraffin sections of nucleoli within a nucleus were 36% greater in 100-day-old (P100) rats than in 1-day-old (P1) rats. Adult values were reached by P50. Added neurons fell disproportionately into the population of neurons whose size was below that of the mean size within the ganglion. The biochemical characteristics of small neurons were used to determine whether added neurons fall into particular subpopulations. In DRGs, L(3) and L(4), the number of neurons immunoreactive to substance P (SP) or calcitonin-gene-related peptide (CGRP) or that bound the lectin isolectin B4 (IB4) was determined. Between P5 and P100, the number of SP stained neurons increased by 2,280 (40% increase), CGRP-stained neurons increased by 6,080 (70% increase), and IB4-stained neurons increased by 6,900 (90% increase). The increase in the number of neurons stained for CGRP or IB4 was more than twice the number of neurons found to be added to these ganglia, indicating that coexpression of these markers as well as neuron number may be developmentally regulated during postnatal life. PMID- 12115687 TI - GABABR1 receptor protein expression in human mesial temporal cortex: changes in temporal lobe epilepsy. AB - Immunocytochemistry was used to examine gamma-aminobutyric acid beta (GABA)(B)R1a b protein expression in the human hippocampal formation (including dentate gyrus, hippocampus proper, subicular complex, and entorhinal cortex) and perirhinal cortex. Overall, GABA(B)R1a-b immunostaining was intense and widespread but showed differential areal and laminar distributions of labeled cells. GABA(B)R1a b-immunoreactive (-ir) neurons were found in the three main layers of the dentate gyrus, the most intense labeling being present in the polymorphic layer, whereas the granule cells were moderately immunoreactive. Except for slight variations, similar distribution patterns of GABA(B)R1a-b immunostaining were found along the different subfields of the Ammon's horn (CA1-CA4). The highest density of GABA(B)R1a-b-ir neurons was localized in the stratum pyramidale, where virtually every pyramidal cell was intensely immunoreactive, including the proximal part of the apical dendrites. Within the subicular complex, a more intense GABA(B)R1a-b immunostaining was found in the subiculum than in the presubiculum or parasubiculum, especially in the pyramidal and polymorphic cell layers. In the entorhinal cortex, distribution of GABA(B)R1a-b immunoreactivity was localized mainly in both pyramidal and nonpyramidal cells of layers II, III, and VI and in the superficial part of layer V, with layers I, IV, and deep layer V being less intensely stained. In the perirhinal cortex, the most intense GABA(B)R1a-b immunoreactivity was located in the deep part of layer III and in layer V and was mainly confined to medium-sized and large pyramidal cells. Thus, the differential expression, but widespread distribution, of GABA(B)R1a-b protein found in the present study suggests the involvement of GABA(B) receptors in many circuits of the human hippocampal formation and adjacent cortical structures. Interestingly, the hippocampal formation of epileptic patients (n = 8) with hippocampal sclerosis showed similar intensity of GABA(B)R1a-b immunostaining in the surviving neurons located within or adjacent to those regions presenting neuronal loss than in the controls. However, surviving neurons in the granule cell layer of the dentate gyrus displayed a significant reduction in immunostaining in 7 of 8 patients. Therefore, alterations in inhibitory synaptic transmission through GABA(B) receptors appears to affect differentially certain hippocampal circuits in a population of epileptic patients. This reduction in GABA(B)R1a-b expression could contribute to the pathophysiology of temporal lobe epilepsy. PMID- 12115688 TI - Functional mapping of the rat olfactory bulb using diverse odorants reveals modular responses to functional groups and hydrocarbon structural features. AB - In an effort to understand the olfactory code of rats, we collected more than 1,500,000 measurements of glomerular activity in response to 54 odorants selected to provide differences in functional groups and hydrocarbon structure. Each odorant evoked a unique response pattern by differentially stimulating clusters of glomeruli, called modules. Odorants sharing specific aspects of their structure activated the same modules, allowing us to relate responses to structure across approximately 80% of the glomerular layer. The most obvious relationship was between the presence of particular oxygen-containing functional groups and the activity of glomeruli within dorsal modules. Functional group specific responses were observed for odorants possessing a wide range of hydrocarbon structure, including aliphatic, cyclic, and aromatic features. Even formic acid and acetone, the simplest odorants possessing acid or ketone functional groups, respectively, stimulated modules specific for these functional groups. At the same time, quantitative analysis of pattern similarities revealed relationships in activation patterns between odorants of similar hydrocarbon structure. The odorant responses were reliable enough to allow us to predict accurately specific aspects of odorant molecular structure from the evoked glomerular activity pattern, as well as predicting the location of glomerular activity evoked by novel odorants. PMID- 12115689 TI - Pattern of synaptic excitation and inhibition upon direction-selective retinal ganglion cells. AB - The distributions of excitatory and inhibitory synapses upon the dendritic arbor of a direction-selective retinal ganglion cell were compared by triple-labeling techniques. The dendrites were visualized by confocal microscopy after injection of Lucifer yellow. Excitatory inputs from bipolar cells were located by using antibodies against kinesin II, a component of synaptic ribbons. Inhibitory inputs were identified by antibodies against gamma-aminobutyric acid-A receptors. The combined images were examined by visual inspection and by formal, automated analyses, in a search for anisotropies that might contribute to a directional preference of the ganglion cell. Within the limits of our analysis, none was found. If an anatomic specialization underlies direction selectivity, it appears to lie in the geometry and spatial positioning of the neurons afferent to the ganglion cell and/or the microcircuitry among its afferent synapses. PMID- 12115690 TI - Patterns of expression of calcium binding proteins and neuronal nitric oxide synthase in different populations of hippocampal GABAergic neurons in mice. AB - We examined the expression of calcium binding proteins parvalbumin (PV), calretinin (CR), and calbindin D28K (CB), and neuronal nitric oxide synthase (nNOS) in gamma-aminobutyric acid (GABA)ergic neurons of the mouse hippocampus, with particular reference to areal and dorsoventral differences. First, we estimated the colocalization of the calcium binding proteins and nNOS. GABAergic neurons containing both PV and nNOS, i.e., PV-immunoreactive (-IR)/nNOS-IR neurons, were rare in Ammon's horn but frequent in the dentate gyrus (DG). CR IR/nNOS-IR neurons and CB-IR/nNOS-IR neurons were frequent in Ammon's horn but rare in the DG. In the entire hippocampus, the percentage of CR-IR neurons containing nNOS was significantly higher at the ventral level (44.3%) than at the dorsal level (17.0%). The percentage of CB-IR neurons containing nNOS was also significantly higher at the ventral level (42.3%) than at the dorsal level (29.3%). Next, we estimated the numerical densities (NDs) of calcium binding protein-containing GABAergic neurons. The ND of PV-IR neurons was comparable at the dorsal (1.16 x 10(3)/mm(3)) and ventral levels (1.23 x 10(3)/mm(3)), respectively. The ND of CR-IR neurons was less at the dorsal level (0.52 x 10(3)/mm(3)) than at the ventral level (0.64 x 10(3)/mm(3)). The ND of CB-IR neurons was also less at the dorsal level (0.91 x 10(3)/mm(3)) than at the ventral level (1.57 x 10(3)/mm(3)). Overall, approximately half of the GABAergic neurons contained one of the three calcium binding proteins (45% at the dorsal level and 47% at the ventral level). These data establish a baseline for examining potential roles of GABAergic neurons in hippocampal network activity in mice. PMID- 12115691 TI - Multiple mechanisms of rhabdom shedding in the lateral eye of Limulus polyphemus. AB - Rhabdom shedding in horseshoe crab lateral eye photoreceptors was studied with anti-opsin and anti-arrestin immunocytochemistry. Two, possibly three, distinct shedding mechanisms were revealed in animals maintained in natural lighting. Transient rhabdom shedding, triggered by dawn, is a brief, synchronous event that removes up to 10% of the rhabdom membrane. Whorls of rhabdomeral membrane break into vesicles and form compact multivesicular bodies. These debris particles are immunoreactive for opsin and are of a relatively uniform size, averaging approximately 2 microm(2) in area. Transient shedding requires that input from circadian efferent fibers to the retina precedes the light trigger, and cutting the optic nerve blocks efferent input and transient shedding. Light-driven rhabdom shedding is a progressive process. Rhabdomeral membrane is removed by coated vesicles that accumulate into loosely packed multivesicular bodies. These debris particles label with antibodies directed against opsin, arrestin, and adaptin, and they have a large distribution of sizes, averaging almost 6 microm(2) in area and ranging up to 25 microm(2) or more. The amount of rhabdomeral membrane removed by light-driven shedding has seasonal variation and depends on latitude. Light-driven shedding does not require circadian efferent input. A possible third shedding mechanism, light-independent shedding, is observed when transient shedding is blocked either by 48 hours of darkness or by cutting the optic nerve. Small particles, averaging 1.8 microm(2) in area, exhibiting opsin but not arrestin immunoreactivity can then be found in the cytoplasm surrounding the rhabdom. The nature of light-independent shedding is not yet clear. PMID- 12115692 TI - Hypothalamic inferior lobe and lateral torus connections in a percomorph teleost, the red cichlid (Hemichromis lifalili). AB - The neuroanatomic connections of the inferior lobe and the lateral torus of the percomorph Hemichromis lifalili were investigated by 1,1', dioctadecyl-3,3,3',3' tetramethylindo-carbocyanine perchlorate (DiI) tracing. The inferior lobe and the lateral torus both receive afferents from the secondary gustatory nucleus. Additional afferents reach the inferior lobe from the nucleus glomerulosus, nucleus suprachiasmaticus, dorsal and central posterior thalamic nucleus, nucleus lateralis valvulae, magnocellular part of the magnocellular nucleus of the preoptic region, caudal nucleus of the preglomerular region, posterior tuberal nucleus, area dorsalis of the telencephalon, and a tegmental nucleus (T2). Efferents from the inferior lobe and the lateral torus terminate in the dorsal hypothalamic neuropil and corpus mamillare. Furthermore, the inferior lobe projects to the medial nucleus of the lateral tuberal hypothalamus and perhaps makes axo-axonal synapses in the tractus tectobulbaris rectus. The inferior lobe and the torus lateralis have reciprocal connections with the preglomerular tertiary gustatory nucleus and posterior thalamic nucleus and are also mutually interconnected. The inferior lobe is also reciprocally connected with the medial nucleus of the preglomerular region, reticular formation and sparsely with the anterior dorsal thalamic and the ventromedial thalamic nuclei. Thus, whereas the lateral torus is exclusively connected with the gustatory system, the inferior lobe is of a multisensory nature. In comparison with the goldfish (Carassius auratus), the connectivity pattern of the inferior lobe of Hemichromis lifalili reflects its specialization with respect to the visual system, as it receives qualitative (i.e., dorsal posterior, anterior, and ventromedial thalamic nuclei) as well as quantitative (i.e., nucleus glomerulosus) additional visual input. PMID- 12115693 TI - HRP injection in lobule VI-VII of the cerebellar cortex reveals a bilateral inferior olive projection in granuloprival rats. AB - In immature rats, Purkinje cells receive synapses from multiple climbing fibers. During development, this multi-innervation regresses and only one climbing fiber innervates each Purkinje cell in the adult. The multi-innervation of immature rats is maintained in the adult if the precursors of the cerebellar granule cells are destroyed by early postnatal X-irradiation. The present study was undertaken to determine the origin of climbing fibers projecting to lobule VI-VII of the cerebellum in X-irradiated granuloprival rats. Olivary neurons were labelled by retrograde transport of wheat germ agglutinin conjugated to horseradish peroxidase, which was injected by iontophoresis in the right vermis of lobule VI VII. Three-dimensional reconstructions of the inferior olive were made for granuloprival and control rats. No significant variation in the shape and dimension of the olive was observed between the two groups. Labeled cells were found in the middle part of the median accessory olive (MAO). In control rats, stained cells were found only in the contralateral MAO, whereas in the granuloprival rats they were located in both the contralateral and the ipsilateral MAO. Homologous zones were marked in control and granuloprival rats in the middle part of MAO. In granuloprival rats, there was a symmetry in the distribution of the stained cells in the ipsi- and contralateral MAO along the three axes. Therefore, polyinnervation involves homologous regions of both inferior olivary nuclei. PMID- 12115694 TI - Cellular localization of the vesicular inhibitory amino acid transporter in the mouse and human retina. AB - Horizontal cells are classically thought to mediate lateral inhibition by gamma aminobutyric acid (GABA)-transporter mediated release. In the mammalian retina, however, GABA uptake and cloned GABA transporter were not detected in horizontal cells. Furthermore, the vesicular inhibitory amino acid transporter (VIAAT or VGAT) that loads GABA and glycine into synaptic vesicles was reported recently to be expressed in horizontal cells. To further assess synaptic transmission in mammalian horizontal cells, we examined the subcellular distribution of VIAAT in mouse and human retina by confocal microscopy with specific cell markers. VIAAT was observed in the mouse outer plexiform layer as punctate structures that localized in calbindin-positive horizontal cells. These structures were in close apposition with synaptophysin-, PSD-95-, dystrophin-, and bassoon-immunopositive photoreceptor terminals, suggesting that VIAAT is localized in horizontal cell tips at photoreceptor terminals. VIAAT-positive puncta were also in apposition to lectin-labeled cone terminals or dendrites of PKCalpha-immunopositive rod bipolar cells, indicating that VIAAT is expressed in horizontal cell tips at both rod and cone terminals. By contrast, only a very few puncta were observed in the human outer plexiform layer, whereas the inner plexiform layer remained labeled as in the mouse retina. When using adult human retinal cells in culture, horizontal cells identified by parvalbumin immunostaining were found to contain VIAAT, either at their terminals or throughout the entire cell similarly as in syntaxin immunopositive cells. These differences between human retinal tissue and cultured cells were attributed to VIAAT degradation in postmortem retinal tissue. VIAAT localization in mouse and human horizontal cells further support the role of inhibitory transmitters in lateral inhibition at the photoreceptor terminals. PMID- 12115695 TI - Spontaneous and augmented growth of axons in the primate spinal cord: effects of local injury and nerve growth factor-secreting cell grafts. AB - Little is known about molecular and cellular responses to spinal cord injury in primates. In this study, the normal milieu of the primate spinal cord was disturbed by multiple needle penetrations and cell injections in the mid-thoracic spinal cord; subsequent effects on local axons and expression of extracellular matrix (ECM) molecules were examined, together with effects of cellular delivery of nerve growth factor (NGF) to the injured region. Four adult rhesus monkeys each received injections of two grafts of autologous fibroblasts genetically modified to secrete human NGF, and, in control injection sites, two separate grafts of autologous fibroblasts transduced to express the reporter gene, beta galactosidase. Three months later, Schwann cells extensively infiltrated the region of localized injury and penetrated both NGF and control fibroblast grafts. Marked upregulation of several ECM molecules occurred, including chondroitin and heparan sulfate proteoglycans and type IV collagen, in or adjacent to all injection sites. Schwann cells were an apparent source of some ECM expression. Spinal cord sensory axons and putative coerulospinal axons extended into both graft types, but they penetrated NGF grafts to a significantly greater extent. Many of these axons expressed the cell adhesion molecule L1. Thus, extensive cellular and molecular changes occur at sites of localized primate spinal cord injury and grafting, attributable in part to migrating Schwann cells, and are accompanied by spontaneous axonal plasticity. These molecular and cellular events closely resemble those observed in the rodent spinal cord after injury. Furthermore, as in rodent studies, cellular delivery of a trophic factor significantly augments axonal plasticity in the primate spinal cord. PMID- 12115696 TI - Skin blood vessels are simultaneously innervated by sensory, sympathetic, and parasympathetic fibers. AB - Despite the known major role of skin blood vessel innervation in blood flow control, particularly in disease, little information on the co-innervation of blood vessels by sensory and autonomic fibers and the relationships of these fibers to one another is available. To fill this gap, we performed a light and electron microscopic analysis of the innervation of skin vessels by sensory and autonomic fibers by using the rat and monkey lower lips as a model. In rats, double-labeling immunocytochemistry revealed that combinations of fibers immunoreactive for substance P (SP) and dopamine-beta-hydroxylase (DbetaH), SP and vesicular acetylcholine transporter (VAChT), as well as DbetaH and VAChT occurred only around blood vessels in the lower dermis. All fiber types travelled in parallel and in close proximity to one another. In the upper dermis, blood vessels were innervated by SP-containing fibers only. Although nerve terminals displayed synaptic vesicles, synaptic specializations were never observed, suggesting that, in this territory, these fibers do not establish synaptic contacts. Quantification of the distance between the various immunoreactive terminals and their presumptive targets (smooth muscle cells and endothelial cells) revealed that both sympathetic and parasympathetic fibers were significantly closer to the endothelial cell layer and smooth muscle cells compared with sensory fibers. In monkeys, double-labeling immunocytochemistry was performed for SP-DbetaH and SP-VAChT only. The results obtained are similar to those found in rats; however, the fiber density was greater in monkeys. Our findings suggest that the regulation of skin microcirculation might be the result of the coordinated functions of sensory and autonomic fibers. PMID- 12115697 TI - Developmental changes in the spatial pattern of mesencephalic trigeminal nucleus (Mes5) neuron populations in the developing chick optic tectum. AB - The developing mesencephalic trigeminal nucleus (nucleus of the fifth cranial nerve; Mes5) is composed of four neuron populations: 1) the medial group, located at the tectal commissure; 2) the lateral group distributed along the optic tectum hemispheres; 3) a group outside the neural tube; and 4) a population located at the posterior commissure. The present work aims to elucidate the site of appearance, temporal evolution, and spatial distribution of the four Mes5 populations during development. According to detailed qualitative observations Mes5 neurons appear as a primitive unique population along a thin dorsal medial band of the mesencephalon. According to quantitative analyses (changes in cell density along defined reference axes performed as a function of time and space), the definitive spatial pattern of Mes5 neurons results from a process of differential cell movements along the tangential plane of the tectal hemispheres. Radial migration does not have a relevant developmental role. Segregation of medial and lateral group populations depends on the intensity of the lateral displacements. The mesenchymal population appears as an outsider subset of neurons that migrate from the cephalic third of the neural tube dorsal midregion to the mesenchymal compartment. This process, together with the intensive lateral displacements that the insider subset undergoes, contributes to the disappearance of this transient population. We cannot find evidence indicating that neural crest-derived precursors enter the neural tube and differentiate into Mes5 neurons. Our results can be better interpreted in terms of the notion that a dorsal neural tube progenitor cell population behaves as precursor of both migrating peripheral descendants (neural crest) and intrinsic neurons (Mes5). PMID- 12115698 TI - Red nucleus projections to distinct motor neuron pools in the rat spinal cord. AB - Despite being one of the more extensively investigated descending pathways of the rat spinal cord, the termination pattern and postsynaptic targets of the rubrospinal tract (RST) still present some unresolved issues. In addition to locomotor functions, the RST is implicated in the control of limb movements such as reaching and grasping. Although a strong RST projection onto interneurons of intermediate Rexed's laminae V and VI have been described through the entire length of the rat spinal cord, the existence of direct rubro-motoneuronal connections have not been demonstrated. In the present study, anterograde tracing of the rat RST with biotinylated dextran amine (BDA) was combined with injections of cholera toxin beta-subunit (CTbeta) into selected groups of forelimb muscles to analyze in detail the rubral projections to the forelimb areas of the cervical spinal cord. The double-staining procedure suggested a direct projection from the RST to specific populations of motoneurons. Three populations of forelimb muscles were distinguished, i.e., paw, "distal" muscles; forearm, "intermediate" muscles; and upper arm, "proximal" muscles. A somatotopic distribution of the corresponding motor neuron pools was present in the spinal cord segments C4-Th1. Rubrospinal axons were seen in close apposition to the distal and intermediate muscle motoneurons, but were consistently absent in the most ventrally situated motor column projecting to proximal muscles. Microstimulation of the red nucleus resulted in electromyographic responses with shorter latency in the distal forelimb muscles than in proximal muscles. These experiments support a specific, preferential role of the RST in distal forelimb muscle control. PMID- 12115699 TI - Cocaine- and amphetamine-regulated transcript peptide projections in the ventral midbrain: colocalization with gamma-aminobutyric acid, melanin-concentrating hormone, dynorphin, and synaptic interactions with dopamine neurons. AB - To date, cocaine- and amphetamine-regulated transcript (CART) peptides have been found to influence feeding, locomotor activity, and conditioned place preference. A common brain structure that could mediate these effects is the ventral tegmental area (VTA). For a better understanding of the anatomical substrates that might underlie CART peptides' role in these behaviors, we performed a series of experiments to elucidate the source, synaptic connectivity, and neurochemical content of CART peptide-immunoreactive (CARTir) terminals in the rat VTA. Double labeling immunofluorescence revealed that approximately 15% of CARTir terminals in the VTA contain the hypothalamic neuropeptide, melanin-concentrating hormone (MCH). Furthermore, CART peptides were also found to colocalize with GABA and, to a small extent, with dynorphin in nerve terminals in both the VTA and the substantia nigra (SN). In the VTA, CARTir terminals form both symmetric and asymmetric synapses onto dopaminergic and nondopaminergic distal dendrites, suggesting that various sources contribute to this innervation. About 30% of CARTir terminals in the VTA and only 15% in the SN appose or form synaptic contact with DA neurons, which support our previous data showing that GABAergic basal ganglia output neurons in the substantia nigra pars reticulata (SNr) receive strong CARTir input from the accumbens core. Results of these studies suggest that the most significant behavioral states influenced by CART peptides, feeding and locomotion, may be mediated by direct and/or indirect modulation of VTA dopaminergic neuronal activity. PMID- 12115700 TI - Organization of trigeminocollicular connections and their relations to the sensory innervation of the eyelids in the rat. AB - Relationships between the trigeminal component of blinking and the superior colliculus (SC) were studied in rats. To localize primary afferent eyelid projections in the sensory trigeminal complex, neuronal tracing experiments were performed as well as analysis of c-Fos protein expression after supraorbital (SO) nerve stimulation. Labelled nerve fibers were found to enter ventrally within the ipsilateral sensory trigeminal complex. Labelled boutons were observed at the junction of the principal nucleus (5P) and the pars oralis (5o) and in the pars caudalis (5c). The c-Fos immunoreactivity was observed in neurons located in the ipsilateral ventral parts of 5P, 5o, and the pars interpolaris (5i) and bilaterally in 5c. Injections in 5P, 5o, 5i, and 5c resulted in anterogradely labelled fibers, with a contralateral preponderance, within the intermediate and deeper SC layers. Injections in 5P or 5o showed anterogradely labelled nerve fibers, profusely terminating in small patches in the medial and central portions of SC layer 4. Subsequently, dense labelling was found in the lateral portion of SC layers 4-7, without patch-like organization. Injections in SC showed retrogradely labelled neurons predominantly within the contralateral part of the sensory trigeminal complex (28% in 5P, 20% in 5o, 50% in 5i, and 2% in 5c). Colocalization of the retrograde tracer after SC injections and c-Fos immunoreactivity in neurons demonstrated that some 5P, 5o, and 5i neurons receive SO nerve inputs and project to SC. This implies that intermediate and deeper SC layers receive sensory information from the eyelids and may be directly involved in the regulation of eye-eyelid coordination. PMID- 12115701 TI - Immunocytochemical localization of a Manduca sexta gamma-aminobutyric acid transporter. AB - Gamma-aminobutyric acid (GABA) is a major inhibitory neurotransmitter in insect central and peripheral nervous systems. Although much work has focused on the downstream targets of GABA, signal termination at insect GABAergic synapses has received very little attention. One of the major mechanisms of terminating synaptic transmission involves transport of the neurotransmitter molecules into presynaptic neurons or surrounding glia. Here we report the immunolocalization of a GABA transporter in the tobacco hornworm, Manduca sexta (MasGAT), using an affinity-purified antibody developed to the C-terminus. This is the first demonstration of an insect neurotransmitter transporter immunolocalization study. Results showed strong staining in the neuropil regions of embryonic, larval, and pharate adult central nervous system. Expression pattern in the pharate adult brain mostly mimicked that observed for GABA, with staining in parts of the optic and antennal lobes, mushroom body, lateral protocerebrum, and central complex. Certain longitudinal and lateral connectives of ganglia were observed to have immunostained fibers representing axons. These data support the view that GABA is involved in visual and olfactory processing in the insect brain. PMID- 12115702 TI - WDR1 colocalizes with ADF and actin in the normal and noise-damaged chick cochlea. AB - Auditory hair cells of birds, unlike hair cells in the mammalian organ of Corti, can regenerate following sound-induced loss. We have identified several genes that are upregulated following such an insult. One gene, WDR1, encodes the vertebrate homologue of actin-interacting protein 1, which interacts with actin depolymerization factor (ADF) to enhance the rate of actin filament cleavage. We examined WDR1 expression in the developing, mature, and noise-damaged chick cochlea by in situ hybridization and immunocytochemistry. In the mature cochlea, WDR1 mRNA was detected in hair cells, homogene cells, and cuboidal cells, all of which contain high levels of F-actin. In the developing inner ear, WDR1 mRNA was detected in homogene cells and cuboidal cells by embryonic day 7, in the undifferentiated sensory epithelium by day 9, and in hair cells at embryonic day 16. We also demonstrated colocalization of WDR1, ADF, and F-actin in all three cell types in the normal and noise-damaged cochlea. Immediately after acoustic overstimulation, WDR1 mRNA was seen in supporting cells. These cells contribute to the structural integrity of the basilar papilla, the maintenance of the ionic barrier at the reticular lamina, and the generation of new hair cells. These results indicate that one of the immediate responses of the supporting cell after noise exposure is to induce WDR1 gene expression and thus to increase the rate of actin filament turnover. These results suggest that WDR1 may play a role either in restoring cytoskeletal integrity in supporting cells or in a cell signaling pathway required for regeneration. PMID- 12115703 TI - Localisation of cannabinoid CB(1) receptor immunoreactivity in the guinea pig and rat myenteric plexus. AB - Activation of cannabinoid CB(1) receptors inhibits gastrointestinal motility, propulsion, and transit, whereas selective antagonism of these receptors has the opposite effects, suggesting the presence of endocannabinoid tone. Supporting evidence for presynaptic CB(1) receptors on myenteric neurons has been found in vitro. In this study, selective CB(1) receptor antibodies and neuronal markers were used to identify and characterise myenteric neurons expressing cannabinoid receptors. Whole mounts of rat and guinea pig myenteric preparations were dually labelled with antibodies against the CB(1) receptor and choline acetyltransferase, neurofilament proteins, calbindin, calretinin, synapsin I, microtubule-associated protein-2, calcitonin gene-related peptide, or substance P. The pattern of CB(1) receptor labelling and the neurochemical classification of CB(1) receptor-positive cells were markedly influenced by the species and fixation procedure. Virtually all choline acetyltransferase-immunoreactive myenteric neurons expressed CB(1) receptors in ganglia from both species. Subpopulations of neurons identified with calbindin, calretinin, and microtubule associated protein-2 did not express CB(1) receptors. A few calcitonin gene related peptide- and substance P-positive somata coexpressed CB(1) receptor immunoreactivity but showed little colocalisation on individual fibres. There was a close association between CB(1) receptor immunoreactivity and fibres labelled for synaptic protein, suggesting a role in the modulation of transmitter release. Functional responses to cannabinoids in the presence of hexamethonium suggest further that CB(1) receptors occur on excitatory motoneurons. In conclusion, CB(1) receptors are expressed on a variety of cholinergic sensory, interneuronal, and motor neurons in myenteric ganglia. PMID- 12115705 TI - Distribution of vesicular glutamate transporter mRNA in rat hypothalamus. AB - Two isoforms of the vesicular glutamate transporter, VGLUT1 and VGLUT2, were recently cloned and biophysically characterized. Both VGLUT1 and VGLUT2 specifically transport glutamate into synaptic vesicles, making them definitive markers for neurons using glutamate as a neurotransmitter. The present study takes advantage of the specificity of the vesicular transporters to afford the first detailed map of putative glutamatergic neurons in the rat hypothalamus. In situ hybridization analysis was used to map hypothalamic distributions of VGLUT1 and VGLUT2 mRNAs. VGLUT2 is clearly the predominant vesicular transporter mRNA found in the hypothalamus; rich expression can be documented in regions regulating energy balance (ventromedial hypothalamus), neuroendocrine function (preoptic nuclei), autonomic tone (posterior hypothalamus), and behavioral/homeostatic integration (lateral hypothalamus, mammillary nuclei). Expression of VGLUT1 is decidedly more circumspect and is confined to relatively weak labeling in lateral hypothalamic regions, neuroendocrine nuclei, and the suprachiasmatic nucleus. Importantly, dual-label analysis revealed no incidence of colocalization of VGLUT1 or VGLUT2 mRNAs in glutamic acid decarboxylase (GAD) 65-positive neurons, indicating that GABA neurons do not express either transporter. Our data support a major role for hypothalamic glutamatergic neurons in regulation of all aspects of hypothalamic function. PMID- 12115706 TI - Glutamate receptors at rod bipolar ribbon synapses in the rabbit retina. AB - In the mammalian retina, maximum sensitivity is achieved in the rod pathway, which serves dark-adapted vision. Rod bipolar cells carry the highly convergent rod input and make ribbon synapses with two postsynaptic elements in the inner retina. One postsynaptic neuron is the AII amacrine cell, which feeds the rod signal into the cone pathways. The other postsynaptic element is either an S1 or S2 amacrine cell. These two wide-field GABA amacrine cells both make reciprocal synapses with rod bipolar terminals but their individual roles are unknown. AII and S1/S2 dendrites come in close together and form a dyad opposing the presynaptic ribbon, which is the site of glutamate release. Therefore, two postsynaptic neurons sense the very same neurotransmitter yet serve different functions in the rod pathway. This functional diversity could be derived partly from the expression of different glutamate receptors on each postsynaptic element. In this study, we labeled all pre- and postsynaptic combinations and a signal-averaging method was developed to locate glutamate receptor subunits. In summary, GluR2/3 and GluR4 are expressed by AII amacrine cells but not by S1/S2 amacrine cells. In contrast, the orphan subunit delta1/2 is exclusively located on S1 varicosities but not on AII or S2 amacrine cells. These results confirm the prediction of divergence mediated by different glutamate receptors at the rod bipolar dyad. Each different amacrine cell type appears to express specific glutamate receptors. Finally, the differential expression of glutamate receptors by S1 and S2 may partly explain the need for two wide-field GABA amacrine cells with the same feedback connections to rod bipolar terminals. PMID- 12115707 TI - Localization of choline acetyltransferase (ChAT) immunoreactivity in the brain of a caecilian amphibian, Dermophis mexicanus (Amphibia: Gymnophiona). AB - The organization of the cholinergic system in the brain of anuran and urodele amphibians was recently studied, and significant differences were noted between both amphibian orders. However, comparable data are not available for the third order of amphibians, the limbless gymnophionans (caecilians). To further assess general and derived features of the cholinergic system in amphibians, we have investigated the distribution of choline acetyltransferase immunoreactive (ChAT ir) cell bodies and fibers in the brain of the gymnophionan Dermophis mexicanus. This distribution showed particular features of gymnophionans such as the existence of a particularly large cholinergic population in the striatum, the presence of ChAT-ir cells in the mesencephalic tectum, and the organization of the cranial nerve motor nuclei. These peculiarities probably reflect major adaptations of gymnophionans to a fossorial habit. Comparison of our results with those in other vertebrates, including a segmental approach to correlate cell populations across species, shows that the general pattern of organization of cholinergic systems in vertebrates can be modified in certain species in response to adaptative processes that lead to morphological and behavioral modifications of members of a given class of vertebrates, as shown for gymnophionans. PMID- 12115708 TI - Ultrastructure of endomorphin-1 immunoreactivity in the rat dorsal pontine tegmentum: evidence for preferential targeting of peptidergic neurons in Barrington's nucleus rather than catecholaminergic neurons in the peri-locus coeruleus. AB - Endomorphins are opioid tetrapeptides that have high affinity and selectivity for mu-opioid receptors (muORs). Light microscopic studies have shown that endomorphin-1 (EM-1) -containing fibers are distributed within the brainstem dorsal pontine tegmentum. Here, immunoelectron microscopy was conducted in the rat brainstem to identify potential cellular interactions between EM-1 and tyrosine hydroxylase (TH) -labeled cellular profiles in the locus coeruleus (LC) and peri-LC, an area known to contain extensive noradrenergic dendrites of LC neurons. Furthermore, sections through the rostral dorsal pons, from colchicine treated rats, were processed for EM-1 and corticotropin releasing factor (CRF), a neuropeptide known to be present in neurons of Barrington's nucleus. EM-1 immunoreactivity was identified in unmyelinated axons, axon terminals, and occasionally in cellular profiles resembling glial processes. Within axon terminals, peroxidase labeling for EM-1 was enriched in large dense core vesicles. In sections processed for EM-1 and TH, approximately 10% of EM-1 containing axon terminals (n=269) targeted dendrites that exhibited immunogold silver labeling for TH. In contrast, approximately 30% of EM-1-labeled axon terminals analyzed (n = 180) targeted CRF-containing somata and dendrites in Barrington's nucleus. Taken together, these data indicate that the modulation of nociceptive and autonomic function as well as stress and arousal responses attributed to EM-1 in the central nervous system may arise, in part, from direct actions on catecholaminergic neurons in the peri-LC. However, the increased frequency with which EM-1 axon terminals form synapses with CRF-containing profiles in Barrington's nucleus suggests a novel role for EM-1 in the modulation of functions associated with Barrington's nucleus neurons such as micturition control and pelvic visceral function. PMID- 12115709 TI - Distribution of tyrosine hydroxylase (TH) and dopamine beta-hydroxylase (DBH) immunoreactivity in the central nervous system of two chondrostean fishes (Acipenser baeri and Huso huso). AB - To obtain a better understanding of the evolution of the brain catecholaminergic systems of fishes, we have examined the distribution of catecholamine synthesizing enzymes in two species of sturgeon (Acipenser baeri and Huso huso) using antibodies against tyrosine hydroxylase (TH) and dopamine-beta -hydroxylase (DBH; only analyzed in Acipenser). Both sturgeons showed TH-immunoreactive (THir) neurons widely distributed in most regions of the brain, the highest number of THir cells being located in the forebrain (olfactory bulb, preoptic area, and posterior tuberculum). THir cells were also seen in other forebrain areas (retrobulbar area, dorsal and ventral telencephalic areas, hypothalamus, ventral thalamus, pretectal area) and in the brainstem (locus coeruleus, viscerosensory area, caudal reticular formation, and area postrema). Immunoreactive fibers and varicosities showed a wide distribution, being particularly abundant in the diencephalon and mesencephalon. DBH-immunoreactive (DBHir) cells were observed in the anterior tuberal nucleus, where these cells were TH-negative, and in the locus coeruleus and the caudal rhombencephalon (vagal reticular formation), where the DBHir cells were also THir. DBHir fibers were scarce in the telencephalon and very abundant in the diencephalon, mesencephalon, and rhombencephalon. The comparative analysis of the catecholaminergic systems of chondrosteans and those observed in other groups of fishes and tetrapods indicate a similar organization of many nuclei, as well as characteristics that are probably primitive, such as the presence of a large number of forebrain catecholaminergic groups. PMID- 12115711 TI - Understanding and treating PTSD: introduction. PMID- 12115710 TI - Vocal-acoustic circuitry and descending vocal pathways in teleost fish: convergence with terrestrial vertebrates reveals conserved traits. AB - Vocal behavior is multifaceted and requires that vocal-motor patterning be integrated at multiple brain levels with auditory, neuroendocrine, and other social behavior processes (e.g., courtship and aggression). We now provide anatomical evidence for an extensive vocal network in teleost fishes (Batrachoididae: Porichthys notatus; Opsanus beta) that is strongly integrated with neuroendocrine and auditory pathways and that exhibits striking similarities to the vocal-acoustic circuitry known for mammals. Biotin compound injections into neurophysiologically identified vocal regions of the forebrain (preoptic area and anterior hypothalamus) and of the midbrain (periaqueductal gray and paralemniscal tegmentum) reveal extensive connectivity within and between these regions, as well as reciprocal relationships with the auditory thalamus and/or auditory midbrain (torus semicircularis). Thus, specific components of the basal forebrain and midbrain are here designated as the forebrain vocal-acoustic complex (fVAC) and midbrain vocal-acoustic complex (mVAC), respectively. Biotin injections into the mVAC and a previously identified hindbrain vocal pattern generator likewise provide anatomical evidence for a distributed network of descending projections to the vocal pacemaker-motoneuron circuitry. Together, the present experiments establish a vocal-auditory-neuroendocrine network in teleost fish that links the forebrain and midbrain to the hindbrain vocal pattern generator (i.e., fVAC --> mVAC --> pattern generator) and provides an anatomical framework for the previously identified neuropeptide modulation of vocal activity elicited from the forebrain and midbrain, which contributes to the expression of sex- and male morph-specific behavior. We conclude with a broad comparison of these findings with those for other vertebrate taxa and suggest that the present findings provide novel insights into the structure of conserved behavioral regulatory circuits that have led to evolutionary convergence in vocal-acoustic systems. PMID- 12115712 TI - Research on posttraumatic stress disorder: epidemiology, pathophysiology, and assessment. AB - Posttraumatic stress disorder (PTSD) is a highly prevalent disorder in both clinical and community populations. This article reviews current knowledge about PTSD in order to assist clinicians in the diagnosis and treatment of reactions to traumatic life events. First, research findings are presented, followed by guidelines for the assessment of trauma and PTSD. Topics discussed include epidemiology, course, and comorbidity, as well as information processing and psychobiology. The review is limited to information about PTSD in adults, although some of the material may generalize to child and adolescent populations. PMID- 12115713 TI - Cognitive-behavior therapy for PTSD in rape survivors. AB - In recent years, new data have appeared, further suggesting the utility of cognitive-behavioral interventions for posttraumatic stress disorder (PTSD) subsequent to sexual assault. In this article, we present a model of cognitive behavioral treatment (CBT) for PTSD in rape survivors. Emotional-processing theory, which proposes mechanisms that underlie the development of disturbances following rape, is reviewed. A CBT-based therapy (Prolonged Exposure) is presented that entails education about common reactions to trauma, relaxation training, imaginal reliving of the rape memory, exposure to trauma reminders, and cognitive restructuring. Current research regarding the use of prolonged exposure is discussed. The case example of a young female rape survivor is described in detail, and her prior substance dependence and intense shame are highlighted. The therapy was successful in reducing the client's symptoms of PTSD, as well as her depressive symptoms, and these gains were maintained at a one-year follow-up assessment. PMID- 12115714 TI - Trauma focus group therapy for combat-related PTSD: an update. AB - Individual cognitive-behavioral therapy involving directed exposure to memories of traumatic events has been found to be effective in treating posttraumatic stress disorder. In this article, we present updated information on an alternative group form of exposure therapy: manualized trauma-focus group therapy (TFGT), designed as an efficient means of conducting directed exposure. We describe the cognitive-behavioral and developmental models from which the approach was derived, present an overview of session topics and a case illustration, provide guidelines for referring individuals to TFGT, and offer suggestions for future research. PMID- 12115715 TI - Brief psychodynamic treatment of PTSD. AB - This article describes a brief psychodynamic psychotherapy for adults suffering from PTSD following exposure to a single traumatic event, such as tragic bereavement, assault, or loss of a body part through surgery. It uses a supportive therapeutic relationship to uncover what the specific event and circumstances that follow mean to the individual and the obstacles to normal psychological processing of these events. Using this 12-session treatment model, therapists pay particular attention to the individual's current phase of response and the typical ways that the individual avoids threatening information. Making links among the recent trauma, earlier developmental experiences that may have rendered the individual vulnerable to the development of PTSD, and ways that conflicts are reenacted in the therapeutic dyad, dynamic therapists seek to help traumatized individuals re-establish a sense of coherence and meaning in their lives. A case illustration is provided to demonstrate the phases and techniques in this approach. PMID- 12115716 TI - Eye Movement Desensitization and Reprocessing (EMDR): information processing in the treatment of trauma. AB - Eye Movement Desensitization and Reprocessing (EMDR) is an efficacious and efficient treatment for posttraumatic stress disorder (PTSD). This article provides a brief overview of the findings of 20 controlled-outcome studies and describes Shapiro's Adaptive Information Processing model. This model posits that pathology results when distressing experiences are processed inadequately and hypothesizes that EMDR accelerates information processing, resulting in the adaptive resolution of traumatic memories. A detailed description of the eight phases of treatment highlights the procedures, assumptions, and clinical observations that currently guide EMDR clinical practice. A case study, with an in-session transcript, illustrates the application of EMDR to address the past events that have laid the groundwork for dysfunction, the present circumstances that elicit distress, and skills acquisition needed for adaptive functioning. PMID- 12115717 TI - Psychosocial treatment of posttraumatic stress disorder: a practice-friendly review of outcome research. AB - A review of the treatment research indicates that several forms of therapy appear to be useful in reducing the symptoms of posttraumatic stress disorder (PTSD). Strongest support is found for the treatments that combine cognitive and behavioral techniques. Hypnosis, psychodynamic, anxiety management, and group therapies also may produce short-term symptom reduction. Still unknown is whether any approach produces lasting effects. Imaginal exposure to trauma memories and hypnosis are techniques most likely to affect the intrusive symptoms of PTSD, while cognitive and psychodynamic approaches may address better the numbing and avoidance symptoms. Treatment should be tailored to the severity and type of presenting PTSD symptoms, to the type of trauma experience, and to the many likely comorbid diagnoses and adjustment problems. PMID- 12115718 TI - Ras family genes: an interesting link between cell cycle and cancer. AB - Ras genes are evolutionary conserved and codify for a monomeric G protein binding GTP (active form) or GDP (inactive form). The ras genes are ubiquitously expressed although mRNA analysis suggests different level expression in tissue. Mutations in each ras gene frequently were found in different tumors, suggesting their involvement in the development of specific neoplasia. These mutations lead to a constitutive active and potentially oncogenic protein that could cause a deregulation of cell cycle. Ras protein moderates cellular responses at several mitogens and/or differentiation factors and at external stimuli. These stimuli activate a series of signal transduction pathways that either can be independent or interconnected at different points. Recent observations begin to clarify the complex relationship between Ras activation, apoptosis, and cellular proliferation. A greater understanding of these processes would help to identify the factors directly responsible for cell cycle deregulation in several tumors, moreover it would help the design of specific therapeutic strategies, for the control on the proliferation of neoplastic cells. We summarize here current knowledge of ras genes family: structural and functional characteristics of Ras proteins and their links with cell cycle and cancer. PMID- 12115719 TI - Mitochondria, the killer organelles and their weapons. AB - Apoptosis is a cell-autonomous mode of death that is activated to eradicate superfluous, damaged, mutated, or aged cells. In addition to their role as the cell's powerhouse, mitochondria play a central role in the control of apoptosis. Thus, numerous pro-apoptotic molecules act on mitochondria and provoke the permeabilization of mitochondrial membranes. Soluble proteins contained in the mitochondrial intermembrane space are released through the outer membrane and participate in the organized destruction of the cell. Several among these lethal proteins can activate caspases, a class of cysteine proteases specifically activated in apoptosis, whereas others act in a caspase-independent fashion, by acting as nucleases (e.g., endonuclease G), nuclease activators (e.g., apoptosis inducing factor), or serine proteases (e.g., Omi/HtrA2). In addition, mitochondria can generate reactive oxygen species, following uncoupling and/or inhibition of the respiratory chain. The diversity of mitochondrial factors participating in apoptosis emphasizes the central role of these organelles in apoptosis control and unravels novel mechanisms of cell death execution. PMID- 12115720 TI - Interview with the retinoblastoma family members: do they help each other? AB - The ultimate destiny of a cell to undergo division, differentiation, survival, and death results from an intricate balance between multiple regulators including oncogenes, tumor suppressor genes, and cell cycle associated proteins. Deregulation of the cell cycle machinery switches the phenotype from a normal cell to a cancerous cell. Fundamental alterations of tumor suppressor genes may result in an unregulated cell cycle with the accumulation of mutations and eventual neoplastic transformation. As such, one may define cancer as a genetic disease of the cell cycle. In this review, we will emphasize our current understanding of how the cell cycle machinery maintains cellular homeostasis by studying the consequences of its deregulation. PMID- 12115721 TI - Transient and sustained ERK phosphorylation and nuclear translocation in growth control. AB - Growth stimulation and inhibition are both associated with tyrosine phosphorylation. We examined the effects of epidermal growth factor (EGF), a growth stimulant, and compound 5 (Cpd 5), a protein-tyrosine phosphatase (PTPase) inhibitor, which inhibits the growth of the same Hep3B hepatoma cells. We found that both EGF and Cpd 5 induced tyrosine phosphorylation of EGF receptor (EGFR) and ERK. However, the phosphorylation caused by EGF was transient and that caused by Cpd 5 was prolonged. Furthermore, Cpd 5 action caused a strong nuclear phospho ERK signal and induced phospho-Elk-1, a nuclear target of ERK activation, in contrast to the weak effects of EGF. An ERK kinase assay demonstrated that ERK activated by Cpd 5 could phosphorylate its physiological substrate, Elk-1. The MEK inhibitors PD098056 and U0126 abrogated both the induction by Cpd 5 of phospho-ERK, its nuclear translocation and phospho-Elk-1 and also antagonized its growth inhibitory effects. Furthermore, phospho-ERK phosphatase and phospho-Elk-1 activities were lost from nuclear extracts from Cpd 5 treated, but not EGF treated cells. In conclusion, the data show that Cpd 5 causes growth inhibition as a consequence of prolonged ERK and Elk-1 phosphorylation, likely a result of inhibition of multiple PTPases, including those acting on phospho-EGFR, on phospho-ERK, and on phospho-Elk-1, in contrast to the kinase driven transient activation resulting from EGF. PMID- 12115722 TI - Plasmin activates pro-matrix metalloproteinase-2 with a membrane-type 1 matrix metalloproteinase-dependent mechanism. AB - Membrane-type 1 matrix metalloproteinase (MT1-MMP) has been implicated as a physiological activator of progelatinase A (MMP-2). We previously reported that plasmin treatment of cells results in proMMP-2 activation and increased type IV collagen degradation. Here, we analyzed the role of MT1-MMP in plasmin activation of MMP-2 using HT-1080 cells transfected with MT1-MMP sense or antisense cDNA. Control, vector-transfected cells that expressed endogenous MT1-MMP, and antisense cDNA transfectants with very low levels of MT1-MMP did not activate proMMP-2. Conversely, cells transfected with sense MT1-MMP cDNA expressed high MT1-MMP levels and processed proMMP-2 to 68/66-kDa intermediate activation products. Control cells and MT1-MMP transfectants had much higher levels of cell associated MMP-2 than antisense cDNA transfectants. Addition of plasmin(ogen) to control or MT1-MMP-transfected cells generated active, 62-kDa MMP-2, but was ineffective with antisense cDNA transfectants. The effect of plasmin(ogen) was prevented by inhibitors of plasmin, but not by metalloproteinase inhibitors, implicating plasmin as a mechanism for proMMP-2 activation independent of the activity of MT1-MMP or other MMPs. Plasmin-mediated activation of proMMP-2 did not result from processing of proMT1-MMP and did not correlate with alpha(v)beta(3) integrin or TIMP-2 levels. Thus, plasmin can activate proMMP-2 only in the presence of MT1-MMP; however, this process does not require the catalytic activity of MT1-MMP. PMID- 12115723 TI - Recovery of cellular E-cadherin precedes replenishment of estrogen receptor and estrogen-dependent proliferation of breast cancer cells rescued from a death stimulus. AB - Loss of estrogen-responsiveness and impaired E-cadherin expression/function has been linked to increased metastatic potential of breast cancer cells. In this study, we report that proliferation of breast cancer cells can resume following removal of a toxic stimulus causing severe impairment of cell adhesion and estrogen responsiveness. This type of response was induced by okadaic acid (OA) in MCF-7 cells, and was accompanied by an almost complete block of DNA synthesis, loss of cell-cell contact and cell detachment from culture dishes, loss of estrogen receptor (ER), progesterone receptor (PR) and E-cadherin, whereas only a weak, if any, inhibition of protein synthesis could be observed. These responses were detected in MCF-7 cells after a 1-day treatment with 50 nM OA, and could be reversed if OA-treated cells were recovered in a culture medium devoid of the toxin, so that rescued cells resumed growth 8-12 days after replating. By pulse chase experiments, we found that protein synthesis was not significantly affected in rescued cells, whose DNA synthesis, instead, was almost completely blocked during the first days of MCF-7 cell rescue from OA treatment. We also analyzed E cadherin, mitogen activated protein kinase isoforms ERK1 and ERK2, Bcl-2 and BAX proteins during the rescue of MCF-7 cells from OA-induced cell death, and found that their expression followed temporally defined patterns. Cellular levels of E cadherin returned to control levels within the first days of the rescue, followed by ER, ERK1, and ERK2, and finally by Bcl-2 and BAX proteins. Under our experimental conditions, restoration of cell adhesion did not require a functional ER system, but recovery of a normal ER pool accompanied resumption of estrogen-dependent proliferation of OA-treated MCF-7 cells. PMID- 12115724 TI - Angiopoietin-2 plays an important role in retinal angiogenesis. AB - Angiopoietin 2 (Ang2) expression in the retina is increased during physiologic and pathologic neovascularization suggesting that it may be involved. In this study, we used Ang2-deficient mice to test that hypothesis. Mice deficient in Ang2 showed delayed and incomplete development of the superficial vascular bed of the retina, which develops primarily by vasculogenesis, and complete absence of the intermediate and deep vascular beds which develop by angiogenesis. In addition to incomplete retinal vascular development, Ang2-deficient mice showed lack of regression of the hyaloid vasculature, resulting in a phenotype that mimics infants with persistent fetal vasculature (PFV), a relatively common congenital abnormality. Exposure to high levels of oxygen resulted in partial regression of the retinal vessels, indicating that oxygen-induced regression of retinal vessels does not require Ang2. When these oxygen-exposed mice with few retinal vessels were moved to room air, there was no ischemia-induced retinal neovascularization. These data support the hypothesis that Ang2 plays a critical role in physiologic and pathologic angiogenesis, and physiologic, but not oxygen induced vascular regression. The data also suggest that infants with PFV should be examined for genetic modifications that would be expected to cause perturbations in Tie2 signaling. PMID- 12115725 TI - Enhanced activity of Ca2+-activated K+ channels by 1-[2-hydroxy-3-propyl-4-[(1H tetrazol-5-yl)butoxyl]phenyl] ethanone (LY-171883) in neuroendocrine and neuroblastoma cell lines. AB - The effects of LY-171883, an orally active leukotriene antagonist, on membrane currents were examined in pituitary GH(3) and in neuroblastoma IMR-32 cells. In GH(3) cells, LY-171883 (1-300 microM) reversibly increased the amplitude of Ca(2+)-activated K(+) current in a concentration-dependent manner with an EC(50) value of 15 microM. In excised inside-out patches recorded from GH(3) cells, the application of LY-171883 into cytosolic face did not modify single channel conductance of large-conductance Ca(2+)-activated K(+) (BK(Ca)) channels; however, it did increase the channel activity. The LY-171883-stimulated activity of BK(Ca) channels is dependent on membrane potential, and results mainly from an increase in mean open time and a decrease in mean closed time. However, REV-5901 (30 microM) suppressed the activity of BK(Ca) channels and MK-571 (30 microM) did not have any effect on it. Under the current-clamp condition, LY-171883 (30 microM) caused membrane hyperpolarization as well as decreased the firing rate of action potentials in GH(3) cells. In neuroblastoma IMR-32 cells, the application of LY-171883 (30 microM) also stimulated BK(Ca) channel activity in a voltage dependent manner. However, neither clofibrate (30 microM) nor leukotriene D(4) (10 microM) affected the channel activity in IMR-32 cells. Troglitazone (30 microM) decreased the channel activity, but ciglitazone (30 microM) enhanced it. This study clearly demonstrates that LY-171883 stimulates the activity of BK(Ca) channels in a manner unlikely to be linked to its blockade of leukotriene receptors or stimulation of peroxisome proliferator-activated receptors. The stimulatory effects on these channels may, at least in part, contribute to the underlying cellular mechanisms by which LY-171883 affects neuronal or neuroendocrine function. PMID- 12115726 TI - p115 Rho GTPase activating protein interacts with MEKK1. AB - Mammalian MAP/ERK kinase kinase 1 (MEKK1) was identified as a mammalian homolog of Ste11p of the yeast pheromone-induced mating pathway. Like Ste11p, MEKK1 is a MAP3 kinase linked to at least two MAP kinase cascades and regulatory events that require cytoskeletal reorganization. MEKK1 is activated by molecules that impact cytoskeletal function. MEKK1-/-cells are defective in cell migration, demonstrating that it is required for cell motility. MEKK1 has a 1,200 residue N terminal regulatory domain that interacts with a dozen identified proteins. Using part of the MEKK1 N-terminus in a yeast two-hybrid screen, we discovered a novel interaction with p115 Rho GTPase-activating protein (GAP). The p115 Rho GAP binds to MEKK1 in vitro and in intact cells. The p115 Rho GAP has selectivity for RhoA over other Rho family members. Expression of p115 Rho GAP reduces MEKK1-induced signaling to AP-1. The reduced activation of AP-1 is dependent on the association of MEKK1 with p115 Rho GAP, because deletion of the Rho GAP SH3 domain, which abrogates their interaction, restores the stimulatory effect of MEKK1 on AP-1 activity. Here we have identified an MEKK1 binding partner that offers a connection between this protein kinase and the machinery regulating cytoskeletal reorganization. PMID- 12115727 TI - CDK9 has the intrinsic property to shuttle between nucleus and cytoplasm, and enhanced expression of cyclin T1 promotes its nuclear localization. AB - CDK9 in association with cyclin T constitutes the P-TEFb complex that stimulates transcription elongation of RNAPII transcripts by phosphorylation of the CTD of RNAPII. Here we report subcellular distribution of P-TEFb in terms of localization of CDK9 and cyclin T1. We found that cyclin T1 is exclusively nuclear and it is present in nuclear-speckled structures. CDK9, albeit mainly nuclear, was also visualized in the cytoplasm. We determined that CDK9 is actively exported from the nucleus, and that leptomycin B (LMB), a specific inhibitor of nuclear export, inhibits this process. Interestingly, enforced expression of cyclin T1 enhances nuclear localization of CDK9. These findings reveal a novel control mechanism for the function of the P-TEFb complex. PMID- 12115728 TI - Anti-proliferative effects of 8-chloro-cAMP and other cAMP analogs are unrelated to their effects on protein kinase A regulatory subunit expression. AB - Conflicting reports have attributed 8-chloro-cAMP (Cl-cAMP)-mediated inhibition of tumor cell growth to either a toxic 8-chloro-adenosine (Cl-AdR) breakdown product or a Cl-cAMP-mediated decrease in ratio of Type I to Type II regulatory (R) subunits of protein kinase A (PKA). Using the MCF-7 human breast cancer and S49 mouse lymphoma cell lines as models, we show that the effects of Cl-cAMP and other cAMP analogs on growth and R subunit expression are unrelated. MCF-7 cell growth was insensitive to most analogs and inducers of cAMP, but was potently inhibited by Cl-cAMP acting through uptake and phosphorylation of its Cl-AdR breakdown product. Possible roles of adenosine receptors or P(2) purinoceptors in these Cl-cAMP-mediated growth effects were ruled out by studies with agonists and antagonists. Cholera toxin markedly decreased the ratio of Type I to Type II R subunits in MCF-7 cells without affecting growth, while growth inhibitory concentrations of Cl-cAMP or Cl-AdR had insignificant effects on this ratio. In S49 cells, where PKA activation is known to inhibit cell growth, PKA-deficient mutants retained sensitivity to both Cl-cAMP and the related 8-bromo-cAMP. Adenosine kinase (AK)-deficient S49 cells were inhibited only by higher concentrations of these 8-halogenated cAMP analogs. Of the commonly used cAMP analogs, only 8-(4-chlorophenylthio)-cAMP acted purely as a cyclic nucleotide having no effect on PKA-deficient cells, but strongly inhibiting both wild-type and AK-deficient cells. Where growth inhibitory concentrations of most cAMP analogs reduced RI expression in the AK-deficient mutant, a functionally equivalent concentration of (N(6), O(2'))dibutyryl-cAMP maintained or increased this expression. PMID- 12115729 TI - Induction or suppression of expression of cytochrome C oxidase subunit II by heregulin beta 1 in human mammary epithelial cells is dependent on the levels of ErbB2 expression. AB - The ErbB family of receptor kinases is composed of four members: epidermal growth factor receptor (EGFR/ErbB1), ErbB2/neu, ErbB3, and ErbB4. Amplification of the ErbB2/neu is found in about 30% of breast cancer patients and is associated with a poor prognosis. Heregulin (HRG) activates the ErbB2 via induction of heterodimerization with ErbB3 and ErbB4 receptors. With suppression subtractive hybridization, we demonstrated that the expression of cytochrome c oxidase subunit II (COXII) is HRG-responsive. Two nontransformed human mammary epithelial cell lines, the HB2 and the HB2(ErbB2) (the HB2 engineered to overexpress ErbB2), displayed an opposite response to HRG-mediated regulation. HRG upregulated mRNA expression of COXII in the HB2 cells, but suppressed COXII expression in the HB2(ErbB2) cells. A human breast cancer cell line (T47D), which expresses ErbB2 at a level similar to that of the HB2 cells, also responded to HRG by increasing COXII mRNA levels. Therefore, HRG regulation of COXII expression depends on the levels of ErbB2 expression. Furthermore, the expression of COXII was inversely correlated to the levels of ErbB2, i.e., the cells overexpressing ErbB2 exhibited lower COXII levels. HRG-evoked signal transduction differed between the cells with normal ErbB expression and the cells overexpressing ErbB2. The activation of both ERK and PI3-K was essential for HRG regulation of COXII, i.e., blockage of either pathway eliminated HRG-mediated alteration. This is the first report demonstrating that the expression of mitochondria-encoded COXII is HRG responsive. The levels of ErbB2 expression are decisive for the diverse biological activities of HRG. PMID- 12115730 TI - Stress kinase p38 mediates EGFR transactivation by hyperosmolar concentrations of sorbitol. AB - Activation of the epidermal growth factor receptor (EGFR) has been shown to occur by ligand-dependent and ligand-independent mechanisms. Different molecular mechanisms have been found to be responsible for ligand-independent receptor transactivation. Here, we show that hyperosmolar concentrations of sorbitol activate the EGFR in human keratinocytes. Experiments using specific inhibitors of EGFR phosphorylation show that the increased amount of activated receptors is the result of a decreased rate of dephosphorylation. Furthermore, sorbitol treatment results in a strong activation of stress kinase p38. Treatment of the cells with SB203580, a known inhibitor of p38 alpha and beta kinases, results in impairment of receptor activation, indicating that the stress kinase is involved in receptor activation modulation. This is further reinforced by experiments showing that addition of Toxin B, known to be an inhibitor of the small Rho GTPases rac1, cdc42, and Rho A/B, to the cells results in a strong induction of EGFR activation. Our results point, therefore, to a mechanism by which osmotic shock activates EGFR through the small Rho GTPases-p38 stress kinase pathway. PMID- 12115731 TI - Cellular response to oxidative stress: signaling for suicide and survival. AB - Reactive oxygen species (ROS), whether produced endogenously as a consequence of normal cell functions or derived from external sources, pose a constant threat to cells living in an aerobic environment as they can result in severe damage to DNA, protein, and lipids. The importance of oxidative damage to the pathogenesis of many diseases as well as to degenerative processes of aging has becoming increasingly apparent over the past few years. Cells contain a number of antioxidant defenses to minimize fluctuations in ROS, but ROS generation often exceeds the cell's antioxidant capacity, resulting in a condition termed oxidative stress. Host survival depends upon the ability of cells and tissues to adapt to or resist the stress, and repair or remove damaged molecules or cells. Numerous stress response mechanisms have evolved for these purposes, and they are rapidly activated in response to oxidative insults. Some of the pathways are preferentially linked to enhanced survival, while others are more frequently associated with cell death. Still others have been implicated in both extremes depending on the particular circumstances. In this review, we discuss the various signaling pathways known to be activated in response to oxidative stress in mammalian cells, the mechanisms leading to their activation, and their roles in influencing cell survival. These pathways constitute important avenues for therapeutic interventions aimed at limiting oxidative damage or attenuating its sequelae. PMID- 12115732 TI - Insights into electromagnetic interaction mechanisms. AB - Low frequency (< 300 Hz) electromagnetic (EM) fields induce biological changes that include effects ranging from increased enzyme reaction rates to increased transcript levels for specific genes. The induction of stress gene HSP70 expression by exposure to EM fields provides insight into how EM fields interact with cells and tissues. Insights into the mechanism(s) are also provided by examination of the interaction of EM fields with moving charges and their influence on enzyme reaction rates in cell-free systems. Biological studies with in vitro model systems have focused, in general, on the nature of the signal transduction pathways involved in response to EM fields. It is likely, however, that EM fields also interact directly with electrons in DNA to stimulate biosynthesis. Identification of an EM field-sensitive DNA sequence in the heat shock 70 (HSP70) promoter, points to the application of EM fields in two biomedical applications: cytoprotection and gene therapy. EM field induction of the stress protein hsp70 may also provide a useful biomarker for establishing a science-based safety standard for the design of cell phones and their transmission towers. PMID- 12115733 TI - Does p53 affect organismal aging? AB - The p53 protein plays a critical role in the prevention of cancer. It responds to a variety of cellular stresses to induce either apoptosis, a transient cell cycle arrest, or a terminal cell cycle arrest called senescence. Senescence in cultured cells is associated with augmented p53 activity and abrogation of p53 activity may delay in vitro senescence. Increasing evidence suggests that p53 may also influence aspects of organismal aging. Several mutant mouse models that display alterations in longevity and aging-related phenotypes have defects in genes that alter p53 signaling. Recently, my laboratory has developed and characterized a p53 mutant mouse line that appears to have an enhanced p53 response. These p53 mutants exhibit increased cancer resistance, yet have a shortened longevity and display a number of early aging-associated phenotypes, suggesting a role for p53 in the aging process. The nature of the aging phenotypes observed in this p53 mutant line is consistent with a model in which aging is driven in part by a gradual depletion of stem cell functional capacity. PMID- 12115734 TI - Progression elevated gene-3, PEG-3, induces genomic instability in rodent and human tumor cells. AB - Genomic instability is a fundamental component of cancer progression. Subtraction hybridization identified a novel rodent gene, progression elevated gene-3 (PEG-3) whose expression directly correlates with cancer aggressiveness and progression. Moreover, ectopic expression of PEG-3 in rodent or human tumor cells produces an aggressive transformed phenotype. We demonstrate that PEG-3 expression in rodent tumor cells correlates directly with genomic instability as characterized by alterations in chromosome composition and structure. Additionally, elevated endogenous or ectopic expression of PEG-3 in rodent and human tumor cells, respectively, enhances gene amplification, as monitored by resistance to methothrexate (MTX) and amplification of the dihydrofolate reductase (dhfr) gene. Stable expression of PEG-3 in normal cloned rat embryo fibroblast (CREF) cells marginally elevates MTX resistance, but morphology remains unaltered and anchorage independence is not induced, suggesting that these phenotypes are separable in immortal cells and gene amplification may precede the acquisition of morphological and oncogenic transformation. The present studies document that stable, inducible, and transient expression of PEG-3 in cancer cells augments genomic instability. In these contexts, one mechanism by which PEG-3 influences cancer progression may be by preferentially facilitating the development of genomic changes in evolving cancer cells. PMID- 12115735 TI - Epidermal expression of the full-length extracellular calcium-sensing receptor is required for normal keratinocyte differentiation. AB - The importance of the extracellular calcium-sensing receptor (CaR) in the stringent control of extracellular Ca(2+) concentration is well established. However, the presence of CaR in tissues not directly involved in regulating mineral ion homeostasis such as the epidermis suggests a role for CaR in other cellular functions. Although extracellular Ca(2+) regulates the differentiation of epidermal keratinocytes, the role of CaR in this process in the epidermis is not fully understood. In this study we showed using in situ hybridization and immunohistochemistry that CaR is expressed in suprabasal keratinocytes of the mammalian epidermis. We then evaluated the changes in epidermal keratinocyte morphology and differentiation in Casr(-/-) mice lacking the full-length CaR. These mice show increased expression of an alternatively spliced form of CaR which lacks acute Ca(2+)-signaling properties. The absence of the full-length CaR in the epidermis resulted in ultrastructural changes (abnormal keratohyalin granule formation and precocious lamellar body secretion) in the terminally differentiated granular keratinocytes. Furthermore, the expression of both mRNA and protein for the calcium inducible keratinocyte differentiation markers, filaggrin and loricrin, were down-regulated in the epidermis of Casr(-/-) mice, whereas the number of proliferating cells were increased even though the calcium gradient within the epidermis was enhanced. Our results demonstrate that the epidermal expression of the full-length CaR is required for the normal terminal differentiation of keratinocytes. PMID- 12115736 TI - CTGF/Hcs24, a hypertrophic chondrocyte-specific gene product, stimulates proliferation and differentiation, but not hypertrophy of cultured articular chondrocytes. AB - We previously reported that connective tissue growth factor/hypertrophic chondrocyte-specific gene product 24 (CTGF/Hcs24) stimulated the proliferation and differentiation of rabbit growth cartilage (RGC) cells in vitro. In this study, we investigated the effects of CTGF/Hcs24 on the proliferation and differentiation of rabbit articular cartilage (RAC) cells in vitro. RAC cells transduced by recombinant adenoviruses generating mRNA for CTGF/Hcs24 synthesized more proteoglycan than the control cells. Also, treatment of RAC cells with recombinant CTGF/Hcs24 (rCTGF/Hcs24) increased DNA and proteoglycan syntheses in a dose-dependent manner. Northern blot analysis revealed that the rCTGF/Hcs24 stimulated the gene expression of type II collagen and aggrecan core protein, which are markers of chondrocyte maturation, in both RGC and RAC cells. However, the gene expression of type X collagen, a marker of hypertrophic chondrocytes, was stimulated by rCTGF/Hcs24 only in RGC cells, but not in RAC cells. Oppositely, gene expression of tenascin-C, a marker of articular chondrocytes, was stimulated by rCTGF/Hcs24 in RAC cells, but not in RGC cells. Moreover, rCTGF/Hcs24 effectively increased both alkaline phosphatase (ALPase) activity and matrix calcification of RGC cells, but not of RAC cells. These results indicate that CTGF/Hcs24 promotes the proliferation and differentiation of articular chondrocytes, but does not promote their hypertrophy or calcification. Taken together, the data show that CTGF/Hcs24 is a direct growth and differentiation factor for articular cartilage, and suggest that it may be useful for the repair of articular cartilage. PMID- 12115737 TI - Transcription factors C/EBP-beta and -delta regulate IL-6 production in IL-1beta stimulated human enterocytes. AB - In recent studies, treatment with IL-1beta of cultured Caco-2 cells, a human intestinal epithelial cell line, resulted in transcriptional upregulation of IL-6 production. The role of C/EBP-beta and -delta in enterocyte IL-6 production is not known. Stimulation with IL-1beta of Caco-2 cells transiently transfected with a luciferase reporter plasmid containing a wild-type IL-6 promoter resulted in an approximately 3.5-fold increase in luciferase activity. This effect of IL-1beta was reduced by approximately 30% when the C/EBP binding site in the IL-6 promoter was mutated, supporting a role of C/EBP in the regulation of IL-6 production. When Caco-2 cells were treated with IL-1beta in the presence of the MAPK inhibitor, PD-98059, IL-6 mRNA and protein levels were reduced by the same concentrations of PD-98059 that inhibited C/EBP DNA binding activity in previous studies. Finally, overexpression of C/EBP-beta and -delta in IL-1beta-treated Caco-2 cells resulted in a 10-12-fold increase in IL-6 production. The results suggest that the beta and delta isoforms of the C/EBP family of transcription factors at least in part regulate IL-6 production in human intestinal epithelial cells. PMID- 12115738 TI - Continuous exposure of airway epithelial cells to hydrogen peroxide: protection by KGF. AB - Reactive oxygen species (ROS) increase permeability in the airway epithelium. Extended periods of oxidant exposure may be experienced by those suffering from chronic inflammation of the lungs, receiving supplemental oxygen, or living in areas with high levels of air pollution. We studied the effects of long-term, continuous exposure to hydrogen peroxide (H(2)O(2)) on the trans-epithelial electrical resistance (TER) across cultured monolayers of a transformed cell line of human bronchial epithelial cells, 16HBE14o- (16HBE). A TER perfusion system was employed to continuously monitor the TER without disturbing the tissue model. The TER decreased in a dose-dependent manner with increasing concentrations of H(2)O(2) (0.1, 0.5, and 1.0 mM), regardless of pre-incubation conditions. Cell cultures pre-treated with 50 ng/ml keratinocyte growth factor (KGF) showed a significant delay in oxidant-induced TER decreases caused by 0.1 mM H(2)O(2). Exposure to 0.1 mM H(2)O(2) for 350 min led to disruption of tight junction proteins, ZO-1 and occludin, but KGF treatment prevented this damage. The recovery of epithelial barrier function after exposure to oxidants was also studied. Tissue models exposed to 0.5 mM H(2)O(2) for 25 min showed complete recovery of TER after 20 h, independent of culture pre-treatment. In contrast, KGF pre-incubation enhanced the recovery of 16HBE cultures exposed for 50 min to 0.5 mM H(2)O(2). PMID- 12115740 TI - Primary human articular chondrocytes, dedifferentiated chondrocytes, and synoviocytes exhibit differential responsiveness to interleukin-4: correlation with the expression pattern of the common receptor gamma chain. AB - Interleukin (IL)-4, which exhibits potent anti-inflammatory activities, is of potential therapeutic value in destructive arthropathies. To further define the response of human joint cells to IL-4, we analyzed the ability of this cytokine to modulate the effects of IL-1beta and growth factors. Freshly isolated chondrocytes, dedifferentiated chondrocytes, and synoviocytes were treated with IL-4 before determination of nitric oxide (NO) and collagenase production in response to IL-1beta, or before proliferation assays in presence of IL-1beta, platelet-derived growth factor (PDGF), or transforming growth factor (TGF)-beta. IL-4 downregulated IL-1beta induced NO production in dedifferentiated chondrocytes and inhibited IL-1beta induced collagenase release, as well as IL 1beta and growth factor induced proliferation in dedifferentiated chondrocytes and synoviocytes. In contrast, IL-4 had no effect in freshly isolated primary chondrocytes and in cartilage explants. The lack of response to IL-4 in primary chondrocytes was associated with impaired signal transduction, as indicated by markedly decreased IL-4 dependent tyrosine phosphorylation of signal transducer and activator of transcription (STAT)-6. It also correlated with differences in the expression pattern of IL-4 receptor (IL-4R) subunits during chondrocyte dedifferentiation. Indeed, whereas the IL-4Ralpha and IL-13Ralpha' subunits were expressed in all cell types, expression of the common receptor gamma chain was restricted to freshly isolated chondrocytes. In conclusion, IL-4 downregulated IL 1beta-induced catabolic events and cell proliferation in dedifferentiated chondrocytes and synoviocytes, but had no effects in freshly isolated chondrocytes. The difference in IL-4 responsiveness between primary and dedifferentiated chondrocytes correlated with changes in proximal signaling events and in the expression pattern of IL-4R subunits during cell dedifferentiation. PMID- 12115739 TI - Cystathionine beta-synthase is coordinately regulated with proliferation through a redox-sensitive mechanism in cultured human cells and Saccharomyces cerevisiae. AB - Cystathionine beta-synthase (CBS) catalyzes the condensation of serine with homocysteine to form cystathionine and occupies a crucial regulatory position between the methionine cycle and the biosynthesis of cysteine by transsulfuration. Analysis of CBS activity under a variety of growth conditions indicated that CBS is coordinately regulated with proliferation in both yeast and human cells. In batch cultures of Saccharomyces cerevisiae, maximal CBS activities were observed in the exponential phase of cells grown on glucose, while growth-arrested cultures or those growing non-fermentatively on ethanol or glycerol had approximately 3-fold less activity. CBS activity assays and Western blotting indicated that growth-specific regulation of CBS is evolutionarily conserved in a range of human cell lines. CBS activity was found to be maximal during proliferation and was reduced two- to five-fold when cells became quiescent at confluence. In cultured HepG2 cells, the human CBS gene is induced by serum and basic fibroblast growth factor and is downregulated, but not abolished, by contact inhibition, serum-starvation, nutrient depletion, or the induction of differentiation. Consequently, for certain cell types, CBS may represent a novel marker of both differentiation and proliferation. The intracellular level of the CBS regulator compound, S-adenosylmethionine, was found to reflect the proliferation status of both yeast and human cells, and as such, constitutes an additional mechanism for proliferation-specific regulation of human CBS. Our data indicates that screening compounds for the ability to affect transsulfuration in cultured cell models must take proliferation status into account to avoid masking regulatory interactions that may be of significance in vivo. PMID- 12115741 TI - Epidermal growth factor stimulates proton efflux from chondrocytic cells. AB - Proton efflux from chondrocytes alters the extracellular pH and ionic composition of cartilage, and influences the synthesis and degradation of extracellular matrix. Epidermal growth factor (EGF) promotes chondrocyte proliferation during skeletal development and accumulates in the synovial fluid in rheumatoid arthritis. The purpose of this study was to investigate the effect of EGF on proton efflux from chondrocytes. When monitored using a Cytosensor microphysiometer, EGF was found to rapidly activate proton efflux from CFK2 chondrocytic cells and rat articular chondrocytes. The actions of EGF were concentration-dependent with half-maximal effects at 0.3-0.7 ng/ml. Partial desensitization and time-dependent recovery of the response were observed following repeated exposures to EGF. EGF-induced proton efflux was dependent on extracellular glucose, and inhibitors of Na(+)/H(+) exchange (NHE) markedly attenuated the initial increase in proton efflux. The response was diminished by inhibitors of phosphatidylinositol 3-kinase and phospholipase C, but not by inhibitors of MEK (MAPK/ERK kinase) or protein kinase A or C. Thus, EGF-induced proton efflux involves glucose metabolism and NHE, and is regulated by a discrete subset of EGF-activated signaling pathways. In vivo, proton efflux induced by EGF may lead to an acidic environment, enhancing turnover of cartilage matrix during development and in rheumatoid arthritis. PMID- 12115744 TI - Osteogenic stem cells and orthopedic engineering: summary and update. AB - The use of osteogenic stem cells or osteoprogenitors to reconstruct skeletal tissues is a popular area of research investigation with high potential for successful use of tissue-engineering principles in orthopedics. Recent studies demonstrate the migration of marrow-derived stem cells to skeletal sites and the proliferation and differentiation at local tissue sites and support possibilities for assessing the successful uses of human osteoprogenitors in the treatment of bone deficiency diseases. In addition, the development of gene therapy procedures in these and other conditions is now considered an achievable goal with the use of these primitive marrow cells. PMID- 12115742 TI - Development of selective tolerance to interleukin-1beta by human chondrocytes in vitro. AB - Interleukin-1 induces release of NO and PGE(2) and production of matrix degrading enzymes in chondrocytes. In osteoarthritis (OA), IL-1 continually, or episodically, acts on chondrocytes in a paracrine and autocrine manner. Human chondrocytes in chondron pellet culture were treated chronically (up to 14 days) with IL-1beta. Chondrons from OA articular cartilage were cultured for 3 weeks before treatment with IL-1beta (0.05-10 ng/ml) for an additional 2 weeks. Spontaneous release of NO and IL-1beta declined over the pretreatment period. In response to IL-1beta (0.1 ng/ml), NO and PGE(2) release was maximal on Day 2 or 3 and then declined to near basal level by Day 14. Synthesis was recovered by addition of 1 ng/ml IL-1beta on Day 11. Expression of inducible nitric oxide synthase (iNOS), detected by immunofluorescence, was elevated on Day 2 and declined through Day 14, which coordinated with the pattern of NO release. On the other hand, IL-1beta-induced MMP-13 synthesis was elevated on Day 3, declined on Day 5, and then increased again through Day 14. IL-1beta increased glucose consumption and lactate production throughout the treatment. IL-1beta stimulated proteoglycan degradation in the early days and inhibited proteoglycan synthesis through Day 14. Chondron pellet cultures from non-OA cartilage released the same amount of NO but produced less PGE(2) and MMP-13 in response to IL-1beta than OA cultures. Like the OA, IL-1beta-induced NO and PGE(2) release decreased over time. In conclusion, with prolonged exposure to IL-1beta, human chondrocytes develop selective tolerance involving NO and PGE(2) release but not MMP-13 production, metabolic activity, or matrix metabolism. PMID- 12115745 TI - Mineralization followup with the use of NMR spectroscopy and others. AB - High-resolution solid state NMR spectroscopy appears to be a powerful method for a better understanding of bone structures and bone substitutes and implants. It is particularly efficient to estimate osteoformation via bioceramic bone colonization. Pathological calcification occurring in bioprosthetic heart valves and breast prostheses can be characterized as a complement to X-ray spectrometry. PMID- 12115746 TI - The effect of the monomer-to-powder ratio on the material properties of acrylic bone cement. AB - The procedure percutaneous vertebroplasty consists of injecting polymethylmethacrylate cement into vertebral bodies for the treatment of osteoporotic compression fractures and tumors of the spine. Clinicians practicing vertebroplasty commonly alter the mixture of monomer-to-powder recommended by the manufacturer in an effort to decrease viscosity and increase the working time. The purpose of the current study was to measure the effect of varying the monomer to-powder ratio on the compressive material properties (compressive modulus, yield stress, and ultimate compressive strength) of the cement Simplex P (Stryker Howmedica-Osteonics, Rutherford, NJ). Cylindrical specimens were prepared using monomer-to-powder ratios of 0.45 to 1.00 mL/g and tested in compression. Peak compressive material properties occurred at the mixture ratio recommended by the manufacturer (0.5 mL/g) but decreased as the ratio of monomer to powder was increased. The material properties of specimens cured for 1 hour were significantly less than those for specimens cured for 24 hours. The monomer-to powder ratio affects the compressive material properties of cement. The clinical significance of these results with respect to vertebroplasty is yet to be determined. PMID- 12115747 TI - The influence of culture conditions on extracellular matrix proteins synthesized by osteoblasts derived from rabbit bone marrow. AB - The influence of culture conditions on the extracellular matrix (ECM) protein expressions of rabbit bone marrow stromal cells has been studied. The focus was on the effects of two kinds of sera, fetal calf serum (FCS) and Ultroser, on cells treated with dexamethasone. The induction of osteoblastic differentiation by dexamethasone addition is confirmed, particularly when cells are cultured in FCS. Bone marrow stromal cells produce alkaline phosphatase positive CFU-F and produce ECM with some mineralized nodules. Analysis by means of two-dimensional gel electrophoresis showed important changes in the composition of ECM proteins after dexamethasone treatment. Overexpression, underexpression, and new synthesized proteins were observed. The most significant modification was linked to the synthesis of four new proteins visible in the acidic area with a low molecular weight of around 17 kDa. These proteins did not correspond to those ECM proteins known to be induced by dexamethasone. Moreover, the effect of dexamethasone on osteoblastic differentiation induction appears very limited when cells are cultured in Ultroser compared to FCS. The protein pattern with Ultroser is different to that obtained with FCS. Cells cultured in Ultroser synthesized no new protein. The different behavior of cells according to the type of medium used is discussed in terms of the osteogenic factors present in the two different sera. PMID- 12115748 TI - The biodegradation mechanism of calcium phosphate biomaterials in bone. AB - This study was undertaken to understand the biodegradation mechanisms of calcium phosphate (Ca-P) biomaterials with different crystallization. Two types of sintered Ca-P porous ceramic (HA and beta-TCP) and a Ca-P bone cement (CPC) were implanted into cavities drilled in rabbit femoral and tibiae condyles. The results have shown that a material biodegradation was rapid in the beta-TCP and the CPC, but very weak in the HA. This biodegradation presented a decrease of material volume from the periphery to the center as well as a particle formation causing phagocytosis by numerous macrophages and multinucleated giant cells in the CPC. In the beta-TCP, there was a peripheral and central decrease of material volume as well as an absence of particle formation or visible phagocytosis. The process of biodegradation is considered to be directly influenced by the type of material crystallization. The sintered bioceramics processed at a high temperature exhibit good crystallization and are primarily degraded by a process dependent on interstitial liquids. However, the bone cement is formed by physicochemical crystallization and is degraded through a dissolution process associated with a cellular process. PMID- 12115749 TI - Bone colonization of beta-TCP granules incorporated in brushite cements. AB - Injectable calcium phosphate hydraulic cements are known to have a high clinical potential in bone reconstruction for mini-invasive orthopaedic surgery, interventional radiology, and rheumatology. Previous in vivo experiments in rabbit have shown that the presence of beta-TCP granules in injectable bone cement help maintain the transient biomechanical function of the implanted bone and promote the formation of good-quality new bone. Histomorphometric analysis of two brushite hydraulic cement (BHC) mixtures selected from previous results (referred to in this work as BHC-A and BHC-B) was performed at three postoperative delays (0, 12, and 24 weeks): histomorphometric analysis of bone colonization within beta-TCP shows that, just before implantation, the beta-TCP granule area is significantly higher in BHC-B; the residual granule area decreases steadily over time in BHC-A, whereas it goes through a maximum of 30% at 12 weeks in BHC-B; the residual granule porosity increases steadily up to 35% in BHC-A, whereas it goes through a maximum of 35% at 12 weeks and decreases somewhat until 24 weeks in BHC-B. New bone formation within granules appears higher in BHC-A (58% Area) compared to BHC-B (38% area) at 12 weeks. At 24 weeks bone colonization levels off in both cements at about 50% area. Irrespective of the cement matrix composition, beta-TCP granules contribute actively to the conduction of new bone formation. PMID- 12115750 TI - Influence of resinous monomers on the differentiation in vitro of human pulp cells into odontoblasts. AB - Odontoblasts are highly differentiated postmitotic cells, which under pathological conditions such as carious lesions and dental injuries may degenerate and be replaced by other pulp cells. A recent work showed that this physiological event can be reproduced in an in vitro assay system. The purpose of the present study was to evaluate the effects of resinous monomers on odontoblast differentiation in vitro. Pulp cores from extracted human third molars were cultured with beta-glycerophosphate (2 mM) and used to evaluate the effects of TEGDMA, HEMA, UDMA, and Bis-GMA on the differentiation of pulp fibroblasts into odontoblasts. The effect of the monomers was studied by evaluating the expression of several odontoblast specific genes. In the absence of monomers, mineral nodule formation was observed. Pulp cells contributing to the nodule formation synthesized type I collagen, osteonectin, and dentin sialoprotein (DSP). In addition, Fourier transform infrared microspectroscopy showed that the mineral and organic composition of the nodules were characteristic of dentin. When the monomers were added at nontoxic concentrations, the effects of HEMA and Bis-GMA were more evident than that of TEGDMA and UDMA on collagen 1, osteonectin, and DSP expression. However, all monomers significantly decreased DSP expression and completely inhibited the mineral nodule formation. PMID- 12115751 TI - Model of the mechanism of Ca loss by bones under microgravity and earth conditions. AB - A comparative characterization of crystal structure, morphology, sizes, and orientation in Ca phosphate precipitation from aqueous solutions, the mineral phase in bones, and mineral deposits on cardiac valves has been performed by high resolution transmission electron microscopy to model possible mechanisms of Ca loss by bones. Physiological changes occurring in organisms can lead to deep perturbations of the natural calcium phosphate supersaturation and its local distribution, which in turn influences the phase composition, morphology, and organization of the mineral phase. Formation of crystals with larger size or of two distinct phases instead of the single hydroxyapatite one can result in the deterioration of the Ca balance in bone and tissue destruction as well as the possible misorientation (or spread of orientation) between HAP crystals newly formed in the bone. PMID- 12115752 TI - Surface structure study of biological calcium phosphate apatite crystals from human tooth enamel. AB - A surface-structure study of human tooth enamel crystals has been carried out by high-resolution electron microscopy (HREM). The surfaces of several crystals have been examined and the surfaces of a single crystal are described here. The observations made on this crystal are similar to the observations made on the other crystals, although the difference of morphology in the crystals observed indicates that the growth control by matrix proteins takes only place on the [011 macro 0] surfaces of the crystals. The crystal described here is oriented along the [112 macro 0] direction, and the following surfaces have been analyzed: (11 macro 00), (011 macro 1), (01 macro 11) and (0001). A subsurface reconstruction is observed just below the surface of the crystal not bonded to the matrix and lying above the supporting film. Observation of the matrix surrounding the crystal shows the existence of poorly crystalline phases with a structure close to that of hydroxyapatite (OHAP). Finally, a comparison of the images of the (011 macro 0) surface with computer-simulated images calculated for several models of the OHAP surface structure shows that the surface itself is stoichiometric and contains both calcium and phosphate groups. The knowledge of the binding sites of proteins on biomineral crystals such as the ones found in human tooth enamel is of prime importance for the understanding of their growth process. In this study, the first structure determination of the surface of human tooth enamel crystals is presented. PMID- 12115753 TI - Development of a new ultrasonic technique for bone and biomaterials in vitro characterization. AB - Classical techniques based on bone imaging allow visual examination or provide quantitative parameters like bone mineral density, which is a mass per unit of surface. Unfortunately, these techniques are generally expensive. A new method of bone characterization that uses ultrasound techniques is presented in this article. This method is able to evaluate mechanical properties like Young's modulus E and cortical bone thickness with low-frequency transducers and does not require the use of a coupling medium. Some results of Young's modulus measurements are presented and compared to the literature values and the pulse echo method. PMID- 12115754 TI - Study of a hydraulic dicalcium phosphate dihydrate/calcium oxide-based cement for dental applications. AB - By mixing CaHPO(4) x 2H(2)O (DCPD) and CaO with water or sodium phosphate buffers as liquid phase, a calcium phosphate cement was obtained. Its physical and mechanical properties, such as compressive strength, initial and final setting times, cohesion time, dough time, swelling time, dimensional and thermal behavior, and injectability were investigated by varying different parameters such as liquid to powder (L/P) ratio (0.35-0.7 ml g(-1)), molar calcium to phosphate (Ca/P) ratio (1.67-2.5) and the pH (4, 7, and 9) and the concentration (0-1 M) of the sodium phosphate buffer. The best results were obtained with the pH 7 sodium phosphate buffer at the concentration of 0.75 M. With this liquid phase, physical and mechanical properties depended on the Ca/P and L/P ratios, varying from 3 to 11 MPa (compressive strength), 6 to 10 min (initial setting time), 11 to 15 min (final setting time), 15 to 30 min (swelling time), 7 to 20 min (time of 100% injectability). The dough or working time was over 16 min. This cement expanded during its setting (1.2-5 % according to Ca/P and L/P ratios); this would allow a tight filling. Given the mechanical and rheological properties of this new DCPD/CaO-based cement, its use as root canal sealing material can be considered as classical calcium hydroxide or ZnO/eugenol-based pastes, without or with a gutta-percha point. PMID- 12115755 TI - Slow-releasing potential of vancomycin-loaded porous hydroxyapatite blocks implanted into MRSA osteomyelitis. AB - Although antibiotic-loaded hydroxyapatite blocks have been used for the treatment of chronic osteomyelitis, their long-term potential for releasing antibiotic into human bones is not well known. Five patients with chronic osteomyelitis due to methicillin-resistant Staphylococcus aureus (MRSA) infection were effectively treated with local implantation of vancomycin-loaded hydroxyapatite blocks. Blocks were removed during the following reconstructive surgeries when the releasing capability of the blocks, and the bacteriocidal activity of the remaining vancomycin in these blocks could be evaluated. Vancomycin was rapidly released within 1 month after implantation, and by 3 months 90% of vancomycin had leaked from the blocks. At 18 months vancomycin still remained in a bacteriocidal form in the hydroxyapatite blocks, though the blocks had no releasing potential or the eluted vancomycin had been changed to a different form. Vancomycin-loaded porous hydroxyapatite blocks would be useful for the treatment of chronic osteomyelitis or implant-associated osteomyelitis due to MRSA. PMID- 12115756 TI - A scanning electron microscopy study of human osteoblast morphology on five orthopedic metals. AB - Despite the long-standing use of metals as orthopedic implants there still are unsolved problems with these materials and open questions about their behavior in a biological environment. Cell-culture studies provide a useful tool for investigations. In addition to the determination of biochemical or molecular biological parameters, the morphology of adhering cells reflects their interaction with the substrata. This article describes an investigation of the morphology of human osteoblasts on stainless steel, cobalt chromium alloy, commercially pure titanium, Ti-6Al-4V, and Ti-6Al-7Nb with surface designs similar to those used as clinical implants. A cell culture plastic surface was used as a control material. The materials were examined by scanning electron microscopy at different points of time. The cells spread, proliferated, and formed nodules on all test substrates in a time-dependent manner, without signs of a disturbing influence from any of the materials. On the smooth surfaces the cells showed a flattened fibroblast-like morphology and only slight differences could be detected. Therefore, the cellular morphology seems not to be markedly affected by the different chemical material compositions. In contrast, the titanium alloy with a rough, sandblasted surface induced a three-dimensional growth. This three-dimensional cellular network could be the basis for the known earlier differentiation of osteoblasts on rough surfaces in vitro and a better osseointegration in vivo. PMID- 12115757 TI - Ce-TZP/Al2O3 nanocomposite as a bearing material in total joint replacement. AB - The objectives of this study were to investigate the biocompatibility, phase stability, and wear properties of a newly developed Ce-TZP/Al(2)O(3) nanocomposite, as compared to conventional ceramics, and to determine whether the new composite could be used as a bearing material in total joint prostheses. In tests of mechanical properties, this composite showed significantly higher toughness than conventional Y-TZP. For biocompatibility tests, cylindrical specimens of both the Ce-TZP/Al(2)O(3) nanocomposite and monolithic alumina were implanted into the paraspinal muscles of male Wistar rats. The tissue reactions were almost the same, and at 24 weeks after implantation, thin fibrous capsules with almost no inflammation were observed around both of them. There were no significant differences in membrane thickness between the two ceramics. After hydrothermal treatment in 121 degrees C vapor for 18 h, the new composite showed complete resistance to aging degradation, whereas Y-TZP showed a phase transformation of 25.3 vol% (initial 0.4%) to the monoclinic form. According to the results of pin-on-disk tests, the wear rates of Ce-TZP/Al(2)O(3) nanocomposite and alumina were 0.55 +/- 0.04 x 10(-7) and 2.12 +/- 0.37 x 10( 7)mm(3)/Nm, respectively. The results of this study suggest that the Ce TZP/Al(2)O(3) nanocomposite is a promising alternative ceramic component for total joint replacement. PMID- 12115758 TI - Preparation of macroporous biodegradable PLGA scaffolds for cell attachment with the use of mixed salts as porogen additives. AB - In the present study, a mixture of ammonium-bicarbonate (NH(4)HCO(3)) and sodium chloride (NaCl) particles was used as a porogen additive to fabricate highly macroporous biodegradable poly(lactic-co-glycolic acid) (PLGA) scaffolds. A two step salt-leaching process was performed after the sample had become semisolidified. Compared to the standard solvent-casting/particulate-leaching (SC/PL) technique, the processing time of this approach was significantly shorter: Instead of several days, only half a day was required. In addition, the polymer/salts/solvent mixture can be easily handled and molded into scaffolds of any specific shape-for example, thin sheet, cylindrical, or bone-shaped-for special applications in tissue engineering. Our results demonstrate that these scaffolds have a highly interconnected open-pore structure as well as greater mechanical properties than those made using the standard SC/PL technique. Primary rat osteoblasts seeded into the scaffolds exhibited good seeding efficiency. The method presented here is a promising approach for fabricating scaffolds for tissue engineering applications. PMID- 12115760 TI - The hydrolytic degradation of polydioxanone (PDSII) sutures. Part II: Micromechanisms of deformation. AB - An investigation of morphological changes in hydrolytically degraded polydioxanone sutures (PDSII) during in situ tensile deformation is reported. These changes were followed with the use of real-time small-angle X-ray scattering with simultaneous tensile deformation experiments. The results are correlated with separately performed wide-angle X-ray scattering, and optical microscopy experiments on the effects of tensile strain. Interlamellar and interfibrillar amorphous regions appear to control the microdeformation and the shape of the stress-strain curves of PDSII sutures. The fibers deform initially by interlamellar separation and at larger strains by fibrillar slip. Fractures appear to be caused by the catastrophic growth of cracks. The degradation of interlamellar tie chains in PDSII sutures does not appear to significantly affect the progress of deformation, because deformation through interlamellar separation is largely unchanged by degradation. Conversely, hydration and scission of interfibrillar tie chains lowers the load required to propagate interfibrillar shear. The fibers fail at progressively lower strains as degradation proceeds. PMID- 12115759 TI - The hydrolytic degradation of polydioxanone (PDSII) sutures. Part I: Morphological aspects. AB - The morphological changes in polydioxanone sutures (PDSII), and the resulting changes in properties are explored as a function of in vitro degradation time with the use of small- and wide-angle x-ray scattering, optical microscopy, differential scanning calorimetry, dynamic mechanical thermal analysis, and measurements of tensile properties and mass change. There are significant changes to the properties of the suture on initial hydration. In the time range of 0-80 days investigated, two subsequent stages of degradation are apparent. Chain scission is thought to occur in the first stage, beginning as soon as the chains become hydrated. However, in this stage, the effects of scission on the physical properties of the suture are not apparent. The transition from the first (dormant) stage to the second (active) stage, in which changes to the physical structure and mechanical properties of the fiber become measurable, is gradual, with the first change occurring after about 15 days. The relationships between the changes in the microstructure and the properties are discussed and a general model for the degradation process proposed. PMID- 12115761 TI - Long-term quantification of the release of monomers from dental resin composites and a resin-modified glass ionomer cement. AB - This study quantified the release of monomers from polymerized specimens of four commercially available resin composites and one glass ionomer cement immersed in water:ethanol solutions. Individual standard curves were prepared from five monomers: (1) triethylene glycol dimethacrylate (TEGDMA), (2) 2-hydroxy-ethyl methacrylate (HEMA), (3) urethane dimethacrylate (UDMA), (4) bisphenol A glycidyl dimethacrylate (BISGMA), and (5) bisphenol A. The concentration of the monomers was determined at Days 1, 7, 30, and 90 with the use of electrospray ionization/mass spectrometry. Data were expressed in mean micromol per mm(2) surface area of specimen and analyzed with Scheffe's test (p<0.05). The following monomers were found in water: monomers (1) and (2) from Delton sealant, monomer (5) from ScotchBond Multipurpose Adhesive and Delton sealant, monomer (3) from Definite and monomer (4) from Fuji II LC, ScotchBond Multipurpose Adhesive, Synergy and Definite. All these monomers increased in concentration over time, with the exception of monomer (1) from Delton sealant. Monomers (3) and (5) were found in extracts of materials despite their absence from the manufacturer's published composition. All monomers were released in significantly higher concentrations in water:ethanol solutions than in water. The greatest release of monomers occurred in the first day. The effect of the measured concentrations of monomers (1-5) on human genes, cells, or tissues needs to be considered with the use of a biological model. PMID- 12115762 TI - Micromorphological changes in resin-dentin bonds after 1 year of water storage. AB - The purpose of this study was to evaluate the degradation of resin-dentin bonds after 1 year of water storage. Resin-dentin-bonded specimens were prepared with the use of an adhesive resin system (One-Step: Bisco). Half of the experimental specimens were sectioned perpendicular to the adhesive interface to produce a beam (adhesive area: 0.9 mm(2)) before being stored in distilled water at 37 degrees C for 1 year. The remaining half of the bonded specimens were sectioned into beams of similar dimensions after 1 year of water storage. Additional bonded specimens that had been stored in water for 24 h before sectioning into beams were used as controls. The beams in the two experimental groups and the control group were subjected to microtensile bond testing. Fractography was performed on all fractured beams with the use of FE-SEM. There were significant (p <.05) differences in bond strength among the control specimens (55.9 +/- 12.9 MPa), specimens that had been sectioned into beams after water storage (68.9 +/- 18.6 MPa), and specimens that had been sectioned into beams before water storage (28.1 +/- 9.3 MPa). Fractography revealed that the resin material was gradually extracted from the periphery to the center portion of the beam. This probably accounted for the decrease in bond strength after 1 year of water storage. PMID- 12115763 TI - A comparison of the shapes of hydroxyapatite implants before and after implantation. AB - Based on helical volume scan computed tomography (HVCT) data, it has been demonstrated previously that real-size models can be prepared with the binder-jet method with extremely high precision of various parts ranging from the outer structure of the cranio-maxillofacial bone to such fine parts as the cranial base and the orbital walls. The application of this method to clinical cases with large cranial bone defects has also been studied. The results showed the usefulness of the binder-jet method, employing starch for fixing, with which a model can be prepared faster and with less deformity caused by the weight of the material itself. Therefore, the binder-jet method is expected to attain greater clinical significance in this field. In this study the comparison between the shapes of implants before and after implantation revealed that the most important aspect of the design of hydroxyapatite (HAP) implants was determining the shapes of edges, especially its angles. That is, an implant needs very little trimming before being implanted to the patient's defective site if the shape of the edges of the implant can be designed and prepared to fix the incised bone surfaces properly. In addition, to obtain optimum results it was also shown that it is necessary to design optimal shapes of implants, taking into account thickness, porosity, and curvature, before processing the shapes of edges. It was found necessary to establish a technology and a method for producing HAP implants that perfectly fit cranial bone defects. For this purpose, it would be useful to make a template of the implants using the mirror-reversion and binder-jet methods. PMID- 12115764 TI - Porous structure of Purevision versus Focus Night&Day and conventional hydrogel contact lenses. AB - The surface and bulk structures of hydrogel contact lenses that contain siloxane moieties, Purevisiontrade mark (balafilcon A) and Focus(R)Night&Daytrade mark (lotrafilcon A), were investigated. Standard hydrogel lenses of low (Seequence(R)), medium (Acuvue(R)), and high water content (Precision UV(R)) were used as controls. All the lenses were dehydrated in a series of ethanol solutions of increased concentration, critical-point dried in CO(2), and sputter coated with gold/palladium before they were examined by scanning electron microscopy. Of all lenses examined, only the balafilcon lenses presented, in addition to the polymer network porosity characteristic of all hydrogels, a macroporous structure that was observed on the front and back surfaces, and in their bulk. The average diameter of the macropores appears to be much larger, from one to several orders of magnitude, than the network porosity of standard hydrogel lenses. The macropores might contribute to the gas and water permeability of these lenses, as well as to their mobility on the cornea. PMID- 12115765 TI - Adhesion barriers of carboxymethylcellulose and polyethylene oxide composite gels. AB - Composite gels and films of CMC and PEO have been used to separate healing tissues and have been demonstrated to reduce postsurgical adhesions in animal models of adhesion formation. Gels of CMC/PEO were studied here to elucidate the mechanism by which the combination of PEO with CMC is effective in reducing adhesions between tissues. CMC and PEO were demonstrated to undergo micro phase separation to form a two-phase system. Protein partitioning was measured in this system for albumin, fibrinogen, and gamma globulin. All of these proteins were found to partition preferentially into the CMC phase. When gels of CMC and PEO were examined for tissue adherence, the addition of PEO reduced the adherence of CMC to tissues. To further investigate the effects of PEO on tissue adherence of the gel, the extent of thrombus formation of citrated blood initiated by calcium chloride in CMC/PEO gels was measured in vitro. The extent of thrombus formation by CMC was reduced proportionally to the content of PEO in gels of CMC/PEO. A model was developed to explain how CMC and PEO contribute to the effectiveness of CMC/PEO gels that form a barrier between healing tissues to reduce postsurgical adhesions. In an open system PEO is released from the gel faster than CMC is dissolved, resulting in a shell structure with CMC coated by PEO. The PEO-rich outer layer functions to inhibit protein deposition and thrombus formation. The CMC-rich layer functions to anchor the gel to the tissue surface. PMID- 12115766 TI - Preparation of blood-compatible hollow fibers from a polymer alloy composed of polysulfone and 2-methacryloyloxyethyl phosphorylcholine polymer. AB - Blood-compatible hollow fibers were successfully prepared from a polymer alloy composed of polysulfone (PSf) and the 2-methacryloyloxyethyl phosphorylcholine (MPC) polymer. To improve the hydrophilicity, fouling-resistance characteristics, and blood compatibility of the PSf hollow fiber in a hemodialyzer, an MPC polymer that can be blended with PSf was synthesized in order to prepare the polymer alloy (PSf/MPC polymer). The contents of the MPC polymer blended in the PSf were 7 and 15 wt%. The PSf/MPC polymer hollow fiber could be prepared by both wet and dry-wet processing methods. The hollow fiber took an asymmetric structure, that is, the hollow-fiber membrane had a dense skin layer on the porous sponge-like structure. The mechanical strength was higher than that of conventional PSf hollow fibers for hemodialysis. The surface characterization of the PSf/MPC polymer hollow fiber by x-ray photoelectron spectroscopy revealed that the MPC units were concentrated at the surface. The permeability for solutes through the PSf/MPC polymer hollow fibers was measured for 4 h. The permeabilities of both a low-molecular-weight compound and protein were greater than those of the PSf hollow fibers. The amount of adsorbed protein was lower on the PSf/MPC polymer hollow fiber when compared to that of the PSf hollow fiber. Moreover, platelet adhesion was also effectively inhibited on the PSf/MPC polymer hollow fiber. Based on these results, the addition of the MPC polymer to the PSf is a very useful method to improve the functions and blood compatibility of the hollow fiber. PMID- 12115767 TI - Ultimate tensile strength of dentin: Evidence for a damage mechanics approach to dentin failure. AB - Dentin structure and properties are known to vary with orientation and location. The present study explored the variation in the ultimate tensile strength (UTS) of dentin with location in the tooth. Hourglass specimens were prepared from dentin located in the center, under cusps, and in the cervical regions of human molar teeth. These were tested in tension at various distances from the pulp. Median tensile strengths ranged from 44.4 MPa in the inner dentin near the pulp, to 97.8 MPa near the dentino-enamel junction (DEJ). This increase in the median UTS with distance from the pulp to the DEJ was statistically significant (P <.001). Of particular importance was the observation that the UTS measurements followed a Weibull probability distribution, with a Weibull modulus of about 4.5. The Weibull behavior of the UTS data strongly suggests that the large variances in fracture strength data result from a distribution of preexisting defects in the dentin. These findings justify a damage-mechanics approach to studies of dentin failure. PMID- 12115768 TI - Lactic acid based PEU/HA and PEU/BCP composites: Dynamic mechanical characterization of hydrolysis. AB - Lactic acid based poly(ester-urethane) (PEU-BDI) and its composites with 20 and 40 vol.% bioceramic filler were characterized prior to their use as biocompatible and bioabsorbable artificial bone materials. Morphological, dynamic mechanical properties, and degradation of these either hydroxyapatite or biphasic calcium phosphate containing composites were determined. Addition of particulate bioactive filler increased the composite stiffness and the glass transition temperature, indicating strong interactions between the filler and matrix. Materials were sterilized by gamma-irradiation, which reduced the average molecular weights by 30-40%. However, dynamic mechanical properties were not significantly affected by irradiation. Specimens were immersed in 0.85 w/v saline at 37 degrees C for 5 weeks, and changes in molecular weights, mass, water absorption, and dynamic mechanical properties were recorded. All the composite materials showed promising dynamic mechanical performance over the 5 weeks of hydrolysis. Average molecular weights of PEU-BDI and its composites did not change substantially during the test period. PEU-BDI retained its modulus values relatively well, and although the moduli of the composite materials were much higher, especially at high filler content, they exhibited faster loss of mechanical integrity. PMID- 12115769 TI - The influence of material and design features on the mechanical properties of transpedicular spinal fixation implants. AB - This study describes the finding and performance of mechanical strength and corrosion testing procedures for comparative examination of multiple internal transpedicular spine fixators. Seven different implant models from five different manufacturers were compared regarding their bending strength and fatigue resistance. Because of the unacceptably high levels of time and material that they require, ISO and ASTM testing standards are not applicable to comparative testing. In addition, there is a lack of knowledge about clinically defined and proven strength-limit values. Therefore, actual standard testing procedures have been modified and extended to corrosion testing. Overall, the effort necessary to obtain reproducible comparative data has been reduced significantly. Although a reduced number of implants of each type were available for destructive testing, the results revealed fundamental differences in the tested implants between different materials and design features. During fatigue testing some of the implants showed poor corrosion properties. Because spinal fixation implants tend to be used as long-duration implants, corrosion testing as well as comparative strength testing with clinically successful implant models should be performed as preclinical evaluations. PMID- 12115770 TI - Investigation of interfacial strength and its structure on the development of a new design of UHMWPE acetabular component. AB - Previous studies associated with the development of a new ultra-high-molecular weight polyethylene (UHMWPE) acetabular component have shown high interfacial tensile strengths through chemical and mechanical bonds between virgin UHMWPE or polymethylmethacrylate (PMMA) and PMMA/methylmethacrylate (MMA) monomer treated UHMWPE. Along with the interfacial strength, the mechanism of interfacial strength development has been investigated, correlating the interfacial strength to its structure, with the different molding temperatures or amount of PMMA in the treated UHMWPE. Of three different fracture patterns-adhesive, mixed, and cohesive-most fractures occurred in the mixed or cohesive mode, indicating either a strong interface or a weak bulk phase. Load-displacement plots from the interfacial tensile tests represented two distinct fracture patterns, suggesting the nature of interfacial structure. Comparison of theoretical and real interfacial strength showed a close match between the two strengths for the interface between PMMA and treated UHMWPE, but a large difference for the interface between UHMWPE and treated UHMWPE. This result hints that although the PMMA/treated UHMWPE interface develops its interfacial strength in a relatively simple mechanism of direct chemical bonds, the UHMWPE/treated-UHMWPE interface builds its strength in a complex way. PMID- 12115771 TI - Superabsorbent materials from shellfish waste--a review. AB - Increasing global demand for improved absorbent materials for body fluids in disposable medical and personal-care articles creates an incentive for new basic research and development of efficient absorbent materials and systems with additional benefits such as biodegradability or certain biomedical functions. Highly absorbing materials based on polyelectrolyte polymers can absorb up to 50 grams of body fluid per gram of dry mass. Currently available synthetic superabsorbents are not biodegradable in landfills and do not offer any value added functions to personal and medical-care products. Various academic and industrial research groups have put considerable amounts of effort and resources toward development of new absorbent materials from natural polymers, which would decompose in landfills. The basic substrates in these studies have been mainly polysaccharides, particularly cellulose and starch. The most common approach has involved converting these polymers into carboxymethyl derivatives followed by structural cross-linking. Commercial synthetic superabsorbent polymers as well as those derived from cellulose and starch are essentially polyanionic. On the other hand, polycationic absorbers seem to have potential functional advantages over the polyanionic counterparts. Chitin is the second abundant natural polymer, whose main derivative, chitosan, becomes polycationic in acid media. Currently, the main source of this polysaccharide is shellfish waste. This review provides basic information about new superabsorbent materials based on chitosan salts, their properties and preparation. PMID- 12115772 TI - Accutane cases: a teratogen information service's approach. PMID- 12115773 TI - Maternal diabetes and malformations. PMID- 12115775 TI - Exposure-disease continuum for 2-chloro-2'-deoxyadenosine (2-CdA), a prototype teratogen: induction of lumbar hernia in the rat and species comparison for the teratogenic responses. AB - BACKGROUND: The purine analog 2-chloro-2'-deoxyadenosine (2-CdA) caused ocular and limb defects in the mouse and rabbit. The current study examined the teratogenic potential of this drug in the rat and compared the adverse developmental outcomes with the other species. METHODS: Timed-pregnant Sprague Dawley rats were given a single intraperitoneal injection of various doses of 2 CdA ranging from 5-60 mg/kg, at gestational day (GD) 9.5 and GD 14. 2-CdA concentrations in maternal serum and embryos were measured by HPLC and termed fetuses were prepared for teratological examination. RESULTS: Full-litter resorption was seen in dams receiving 50 mg/kg of 2-CdA at GD 9.5, whereas post implantation loss was significantly increased and fetal weights significantly reduced at 40 mg/kg. Gross examination of the surviving fetuses revealed microphthalmia, a shortened body trunk and lumbar hernia, manifested by a soft mass protrusion at the lumbar region on one or both sides of the spine. Incidence of these defects increased in a dose-dependent fashion. Histological examination indicated that the hernia was associated with hypoplasia of the body wall, poorly developed skeletal muscle bundles surrounding the vertebral column in the lumbar region, and an absence of the lateral muscle groups that allowed protrusion of the abdominal viscera. The lumbar hernia was generally accompanied by spina bifida, deformed ribs and a wide spectrum of soft tissue-abnormalities that included kidney, genitourinary and heart defects. At GD 14, exposure to 2-CdA at 60 mg/kg produced oligodactyly in one of six litters. CONCLUSIONS: 2-CdA produced similar ocular defects in the rat and mouse, although the incidence was much lower in the former species. In contrast, the drug-induced lumbar hernia was only seen in the rat. These apparent disparities were not readily explained by species differences in pharmacokinetic parameters. the similarities between the teratological features of 2-CdA-induced lumbar hernia in the rat and the clinical description of lumbocostovertebral syndrome, however, may provide a key to unlock the etiology of this rare birth defect in humans. PMID- 12115776 TI - Effect of prenatal exposure to anticonvulsant drugs on dermal ridge patterns of fingers. AB - BACKGROUND: An altered frequency of specific dermal ridge patterns on fingertips, such as an increased number of arches, has been observed in children exposed in utero to anticonvulsants and other teratogens. Asymmetry of the distribution of dermal ridge patterns has been attributed to environmental exposures and genetic factors. METHODS: We evaluated all of the dermal ridge patterns of 66 children who had been exposed to either the anticonvulsant phenytoin alone or phenytoin and phenobarbital. We determined the frequency of each pattern, concordance between the fingers on the left and right hands, sex differences and total ridge counts in the drug-exposed children and compared them to the findings in 716 unexposed comparison children. The frequency of each pattern was established in comparison to the most common type of pattern (ulnar loop), which showed that there were alterations in the frequency of arches, radial loops and whorls on specific fingers. RESULTS: Eight (12.1%) of 66 children had three or more arch patterns, with all but one having been exposed to phenytoin and phenobarbital. Only one of these eight children was considered by the masked examiner to have fingernail hypoplasia. There was no evidence of asymmetry in the anticonvulsant exposed children. There were minor differences in the distribution of total ridge count. CONCLUSIONS: Subtle differences in several dermal ridge patterns, not just arch patterns, were present in anticonvulsant-exposed children, primarily in those exposed to polytherapy: phenytoin and phenobarbital. PMID- 12115777 TI - Further evidence for the role of free radicals in the limb teratogenicity of L NAME. AB - BACKGROUND: L-NAME (N(G)-nitro-(L)-arginine methyl ester), a nitric oxide synthase inhibitor, causes severe limb reduction malformations when gravid rats are treated intraperitoneally on gd-17. Hemorrhages, appearing within hours of L NAME administration, and defects at term can be significantly reduced by co treatment with PBN (alpha-phenyl-N-t-butylnitrone), a spin trap antioxidant. We have proposed that limb defects result from ischemia-reperfusion injury. We examine the role of xanthine oxidase and ROS formation in the limb effects of L NAME. METHODS: Gravidas were treated with L-NAME (50 mg/kg) in the presence or absence of allopurinol, a xanthine oxidase inhibitor. Spatial patterns of limb hemorrhage were determined promptly and at term as was digit length at the latter interval. Xanthine oxidase activities were assayed in control and treated limbs with and without allopurinol co-treatment. RESULTS: Allopurinol significantly reduced hemorrhage severity in a dose-responsive fashion when fetuses were examined at term. Higher doses of allopurinol significantly preserved digit length. Xanthine oxidase activities in fetal limb were significantly increased by L-NAME treatment whereas co-treatment with allopurinol restored activities to near-control levels. CONCLUSIONS: These findings support the role of excess reactive oxygen species (ROS) formation in L-NAME-induced limb reduction. We propose that nitric oxide (NO) depletion by L-NAME interferes with vascular integrity, and causes vasoconstriction. Resultant hypoxia stimulates superoxide formation and nitric oxide formation catalyzed by the inducible isoform of nitric oxide synthase. The reduction products of superoxide or the products of its reaction with nitric oxide oxidize or nitrate endothelial components resulting in limb reduction defects. PMID- 12115779 TI - Insufficient folic acid intake in the Netherlands: what about the future? AB - BACKGROUND: In 1993 all women of childbearing age in the Netherlands were advised to take a daily 0.5 mg folic acid pill to reduce the risk for neural tube defects. This study describes both recent and past awareness and use of folic acid supplements in relation to socio-economic status in the Northern Netherlands. The consequences of a recent report of the Dutch Health Council report will be discussed as well. METHODS: In the most recent cross-sectional study (November 2000), pregnant women filled out a questionnaire. Out of 473 women, 461 were willing to cooperate. The highest fulfilled level of education was taken as an indicator for socio-economic status. RESULTS: Seventy-seven percent (n = 357) of the respondents had heard about folic acid before being pregnant. Sixty-three percent (n = 289) knew about the protective effect for NTDs and 33% (n = 151) knew the entire advised period. Sixty-one percent (n = 265) of the respondents used folic acid in some part of the advised period and 36% (n = 164) used it in the entire advised period. Higher educated women knew more about folic acid and used it significantly more often in the periconceptional period than lower educated women. CONCLUSIONS: Because compliance to proper use of folic acid was poor, food fortification in the Netherlands must be seriously considered. The Dutch Health Council wants to limit the fortification of food products to those products that are especially aimed for women who wish to become pregnant. The fortification of specific products instead of staple foods is a missed chance to reduce NTDs and possibly other birth defects and cardiovascular defects as well. PMID- 12115778 TI - Prevalence of spina bifida and anencephaly during the transition to mandatory folic acid fortification in the United States. AB - BACKGROUND: In 1992, the United States Public Health Service recommended that all women of childbearing age consume 400 microg of folic acid daily. The Food and Drug Administration authorized the addition of synthetic folic acid to grain products in March 1996 with mandatory compliance by January 1998. The impact of these public health policies on the prevalence of neural tube defects needs to be evaluated. We sought to determine the prevalences of spina bifida and anencephaly during the transition to mandatory folic acid fortification. METHODS: Twenty-four population-based surveillance systems were used to identify 5,630 cases of spina bifida and anencephaly from 1995-99. Cases were divided into three temporal categories depending on whether neural tube development occurred before folic acid fortification (January 1995 to December 1996), during optional fortification (January 1997 to September 1998), or during mandatory fortification (October 1998 to December 1999). Prevalences for each defect were calculated for each time period. Data were also stratified by programs that did and did not ascertain prenatally diagnosed cases. RESULTS: The prevalence of spina bifida decreased 31% (prevalence ratio [PR] = 0.69, 95% confidence interval [CI] = 0.63-0.74) from the pre- to the mandatory fortification period and the prevalence of anencephaly decreased 16% (PR = 0.84, 95% CI = 0.75-0.95). Stratification by prenatal ascertainment did not alter results for spina bifida but did impact anencephaly trends. CONCLUSIONS: The decline in the prevalence of spina bifida was temporally associated with folic acid fortification of US grain supplies. The temporal association between fortification and the prevalence of anencephaly is unclear. PMID- 12115780 TI - Inertia on folic acid fortification: public health malpractice. PMID- 12115784 TI - Constructing representations of Karl Spencer Lashley. AB - I compare, contrast, and analyze two published constructions, a straightforward approach and a social constructivist perspective, of the life and work of psychobiologist Karl S. Lashley (1890-1958). Although it is clear at a general level that scientific endeavors are affected by extrascientific factors, particular concern is directed at the issues involved in demonstrating that the scientific work of an individual, in this case Lashley, is affected by specific extrascientific influences. There is insufficient evidence to conclude that Lashley's work was driven predominantly by racial and genetic determinist positions and that he was something other than the scientist he represented himself as being. I then discuss an unpublished personality theory approach to Lashley that suggests the influence of some personality characteristics on his work and helps to provide balance and perspective in constructing representations of Lashley. PMID- 12115785 TI - The depoliticization of Karl Lashley: A response to Dewsbury. AB - In this response to Donald Dewsbury's article, I address Dewsbury's criticisms of my interpretation of Karl Lashley by, first, disputing Dewsbury's characterization of my historiographic stance. I then go on to address Dewsbury's specific points, particularly regarding Lashley's racism and hereditarianism, which I argue are central to an understanding of the context of his science. I conclude by criticizing Dewsbury's proposed personality theory of Lashley on the grounds that it depoliticizes Lashley by psychologizing him. PMID- 12115787 TI - Toward a racial abyss: eugenics, Wickliffe Draper, and the origins of The Pioneer Fund. AB - The Pioneer Fund was created in 1937 "to conduct or aid in conducting study and research into problems of heredity and eugenics . . . and problems of race betterment with special reference to the people of the United States." The Fund was endowed by Colonel Wickliffe Preston Draper, a New England textile heir, and perpetuates his legacy through an active program of grants, some of the more controversial in aid of research on racial group differences. Those presently associated with the Fund maintain that it has made a substantial contribution to the behavioral and social sciences, but insider accounts of Pioneer's history oversimplify its past and smooth over its more tendentious elements. This article examines the social context and intellectual background to Pioneer's origins, with a focus on Col. Draper himself, his concerns about racial degeneration, and his relation to the eugenics movement. In conclusion, it evaluates the official history of the fund. PMID- 12115788 TI - Friedrich Albert Lange on neo-Kantianism, socialist Darwinism, and a psychology without a soul. AB - Friedrich Albert Lange was a German philosopher, political theorist, educator, and psychologist who outlined an objective psychology in the 1860s. This article shows how some of the most important worldviews of the nineteenth century (Kantianism, Marxism, and Darwinism) were combined creatively in his thought system. He was crucial in the development of neo-Kantianism and incorporated psycho-physiological research on sensation and perception in order to defend Kant's epistemological idealism. Based on a critique of phrenology and philosophical psychology of his time, Lange developed a program of a psychology without a soul. He suggested that only those phenomena that can be observed and controlled should be studied, that psychology should focus on actions and speech, and that for each psychological event the corresponding physical or physiological processes should be identified. Lange opposed introspection and subjective accounts and promoted experiments and statistics. He also promoted Darwinism for psychology while developing a socialist progressive-democratic reading of Darwin in his social theory. The implications of socialist Darwinism on Lange's conceptualization of race are discussed and his prominence in nineteenth century philosophy and psychology is summarized. PMID- 12115789 TI - Freud's "bad conscience": The case of Nietzsche's Genealogy. AB - This article develops the argument that Friedrich Nietzsche influenced several aspects of Freud's later writings by illustrating, in particular, the impact of Nietzsche's On the Genealogy of Morals on Freud's Civilization and its Discontents. The theoretical and conceptual schemes represented in Freud's Discontents are found to bear a remarkable similarity to Nietzsche's Genealogy on a number of highly specific points. It is suggested that "DAS ES," "Uber-ich," and "bad conscience," concepts central to Freud's moral theory of mind, are at least partly derived from Nietzsche. Moreover, Freud's phylogenetic theory of guilt is based upon premises found in Nietzsche, as are specific details relating to ideas on human prehistory and the ancestral family. Based on this evidence, a re-examination of the moral and social dimensions of Freud's "structural" model may be in order. PMID- 12115792 TI - The TNM system: our language for cancer care. PMID- 12115793 TI - Sole brachytherapy of the tumor bed after conservative surgery for T1 breast cancer: five-year results of a phase I-II study and initial findings of a randomized phase III trial. AB - BACKGROUND AND OBJECTIVES: The objectives of this study were to test the feasibility of sole interstitial high-dose-rate brachytherapy (HDR-BT) after breast-conserving surgery (BCS) for T1 breast cancer in a phase I-II study, and to present the initial findings of a phase III trial comparing the efficacy of tumor bed radiotherapy (TBRT) alone with conventional whole breast radiotherapy (WBRT). METHODS: Forty-five prospectively selected patients with T1 breast cancer undergoing BCS were enrolled into a phase I-II study of TBRT alone, using interstitial HDR implants. HDR-BT of 7 x 4.33 Gy (n = 8) and 7 x 5.2 Gy (n = 37) was delivered to the tumor bed. Based on the results of this phase I-II study, a further 126 patients were randomized to receive 50 Gy WBRT (n = 63) or TBRT alone (n = 63); the latter consisted of either 7 x 5.2 Gy HDR-BT (n = 46) or 50-Gy wide field electron irradiation (n = 17). Breast cancer related events and side effects were assessed. RESULTS: In the phase I-II study, at a median follow-up of 57 months, 2 (4.4%) local, 3 (6.7%) axillary, and 3 (6.7%) distant failures were observed. Two patients (4.4%) died of breast cancer. The 5-year probability of cancer-specific, relapse-free and local recurrence-free survival was 90.0%, 85.9%, and 95.6%, respectively. The cosmetic results were judged to be excellent in 44 of 45 patients (97.8%). Severe (higher than grade 2) skin sequelae or fibrosis was not found. Symptomatic fat necrosis occurred in one patient (2.2%). In the phase III study, at a median follow-up of 30 months, the locoregional tumor control was 100% in both arms. The 3-year probability of cancer-specific and relapse-free survival was 98.1% and 98.4% in the WBRT group and 100% and 94.4% in the TBRT group, respectively (P = NS). There was no significant difference between the two treatment arms regarding the incidence of radiation side effects. CONCLUSIONS: Five-year results of our phase I-II study prove that sole HDR-BT of the tumor bed with careful patient selection and adequate quality assurance is a feasible alternative to WBRT. However, long-term results of phase III trials are required to determine the equivalence of TBRT alone, compared with WBRT in the management of selected patients with early breast cancer. PMID- 12115795 TI - Sentinel node biopsy for breast cancer: does the number of sentinel nodes removed have an impact on the accuracy of finding a positive node? AB - BACKGROUND AND OBJECTIVES: The number of sentinel lymph nodes (SLNs) removed during biopsy may have an impact on the accuracy of finding a positive SLN. This study investigated various factors to determine if they had any significant correlation with the number of SLNs found during biopsy. In patients with positive SLNs, the nodes were then analyzed to determine which SLN contained metastasis. METHODS: For 263 patients with breast cancer who successfully underwent SLN biopsy, parameters such as tumor size, histologic characteristics, differentiation, and receptor status, patient age, breast quadrant, type of surgery, mapping with blue dye only or with radiocolloid, and whether biopsy was performed before or after tumorectomy were prospectively collected. These factors were analyzed to determine whether they had any significant correlation to the number of removed lymph nodes. Positive SLNs were ranked in the order they were removed and examined for which node contained the metastasis. RESULTS: During biopsy, a mean of 1.8 (range, 1-5) SLNs were found. The SLNs were negative in 158 patients and positive in 105. The number of SLNs removed was comparable between node-negative and node-positive patients, and none of the parameters analyzed was significantly related to the number of SLNs removed. Of the 105 patients with a positive SLN, the first SLN independently predicted the pathologic status of the axilla in 96 patients (91.4%; 95% CI 86.1-96.8), and the first and second SLN independently predicted the status in 104 patients (99%; 95% CI 97.2-100). Only one of 105 patients had metastasis in the third SLN removed. CONCLUSION: The pathologic status of the axilla was independently determined by removal of the first or first and second SLN in 99% of patients; removal of more than three SLNs did not increase the accuracy of finding a positive node. PMID- 12115796 TI - Influence of environment and healthcare structures on the survival of patients with colorectal cancer: a French population-based study. AB - BACKGROUND AND OBJECTIVES: Colorectal cancer is one of the highest-ranking cancers in France, both sexes combined, with 33000 new cases per year. To report on the practice and the efficiency of the healthcare system, an evaluation of the therapeutic management of colorectal cancer was carried out in the department of the Herault, in the south of France. METHODS: Cases of colorectal cancer in 1992 (344 colorectal cancer incidental cases) in the department of the Herault were reviewed. The diversity of the therapeutic choices and survival were evaluated for the different types of healthcare facilities (private hospitals, nonspecialized and specialized hospitals) and residential areas (rural, semi urban, urban). RESULTS: Two hundred seventy-one patients with colorectal cancer (78.8%) and 234 patients with colorectal cancer (67.7%) were respectively diagnosed and treated in the private sector. Sixteen cases (23.5%) in the public sector (29.7% in the university hospital) and 24 cases (19%) in the private sector involved surgical emergencies (peritonitis, intestinal obstructions) (P = 0.003). Radiotherapy was performed in 59% of patients with rectal cancer. Preoperative radiotherapy was used primarily in specialized hospitals (80% of radiated rectal cancer; P = 0.002), as opposed to postoperative radiotherapy, which was used predominantly in private hospitals (P = 0.005). Forty-five percent of the patients with colorectal cancer who had lymph node involvement have been treated with chemotherapy. In multivariate analysis, lymph node metastasis and the presence of metastases (Dukes stage) were the most important independent pejorative prognostic factors, followed by the initial treatment in nonspecialized hospitals, complicated colorectal cancer (intestinal obstruction, peritonitis), lack of histological differentiation, and rural and urban residential areas. CONCLUSIONS: Apart from independent prognostic factors, such as parietal, ganglionic, or metastatic extensions, the lack of histological differentiation, and the complicated forms, heterogeneity and inequality persist in the distribution, treatment for, and the survival of patients with colorectal cancer based on the type of healthcare facility and the living area of this French population. PMID- 12115797 TI - Hepatic arterial infusion of 5-fluorouracil and cisplatin for unresectable or recurrent hepatocellular carcinoma with tumor thrombus of the portal vein. AB - BACKGROUND AND OBJECTIVES: This study was designed to evaluate the efficacy of hepatic arterial infusion of 5-fluorouracil (5-FU) and cisplatin (CDDP) for unresectable or recurrent hepatocellular carcinoma (HCC) with tumor thrombus of the trunk or first branches of the portal vein (PVTT). METHODS: Seven unresectable or recurrent HCC patients with PVTT were enrolled in this study. A one-week course of chemotherapy consisted of intraarterial administration of CDDP (10 mg on days 1-5) for 1 h and 5-FU (250 mg on days 1-5) for 24 h, followed by cessation of administration for the subsequent 2 days (days 6 and 7). Three or more sequential courses of chemotherapy were given through an implanted reservoir. RESULTS: Serum alpha-fetoprotein (AFP) levels before the chemotherapy were >20 ng/ml in six of the seven patients. Serum AFP levels were decreased in four of the six patients after chemotherapy, including two patients (cases 1 and 7) whose AFP levels later returned to normal. Six of the seven patients had measurable lesions in the liver, with a response rate of 33%. In three of the seven patients (43%), PVTTs decreased in size or disappeared after chemotherapy. The mean and median survival periods of all patients were 8.0 and. 7.5 months, respectively. CONCLUSIONS: The chemotherapy described in this report is beneficial in terms of survival for HCC patients with PVTT for whom transcatheter arterial embolization or surgical treatment is contraindicated. PMID- 12115798 TI - Tumor response to arterial embolization hyperthermia and direct injection hyperthermia in a rabbit liver tumor model. AB - BACKGROUND AND OBJECTIVES: It is possible to arterially embolize or directly inject liver tumors in small animal models with ferromagnetic particles that generate hysteretic heating on exposure to an alternating magnetic field. The objective of this study was to compare the response of liver tumors to arterial embolization hyperthermia (AEH) and direct injection hyperthermia (DIH). METHODS: Ten rabbits containing experimental hepatic tumors were treated with AEH, and a second group of ten rabbits were treated with DIH. The tumors of both groups were heated to 43 degrees to 50 degrees C for 20 minutes. Tumor response, which was determined by measuring change in tumor volume and by comparison of tumor mass after treatment with the mass of untreated control tumors of the same age, was assessed 14 days after treatment. RESULTS: All tumors treated with AEH decreased in volume by 50% to 94% (P = 0.005), and their average mass (median 1.73 gm) was significantly less than that of untreated control tumors (median 8.01 gm, n = 20; P < 0.001). Three of the treated tumors were completely necrotic, while the remainder were at least 80% necrotic. Nine of the ten tumors treated with DIH increased in volume by at least 143% (P = 0.01), and their average mass (median 5.68 gm) was not significantly different from that of the untreated control tumors (P = 0.56). CONCLUSION: These results indicate that AEH is more effective than DIH at moderately elevated temperatures. This is probably because the more widespread particle distribution that can be achieved using arterial embolization results in more extensive and complete treatment of the tumor. PMID- 12115800 TI - Surgical lesioning of splanchnic nerves using wet needle radiofrequency thermoablation. PMID- 12115799 TI - Diagnosis and management of patients with thyroid nodules. AB - Fewer than 5% of all adults will have a palpable thyroid nodule, but this is still a large number of individuals who require evaluation. Although most thyroid nodules are a result of a benign disease process (more than 95%), the possibility of thyroid cancer is always a consideration. Important aspects of history taking with a patient in whom a thyroid nodule has been noted include age, gender, family history of thyroid or endocrine disease, prior low dosage head and neck radiation, recent hoarseness, dysphagia, and symptoms of hypermetabolism. Key features of evaluation by physical examination are the size and location of the thyroid abnormality, the degree of firmness of the nodule, the presence of other nodules in the thyroid, palpable cervical lymph nodes, vocal cord paresis or paralysis, and tachycardia and/or tremor. The major categories of thyroid abnormality in such patients include cysts, adenomas, thyroiditis, and cancer. Although radionuclide scans, ultrasound examination and computer tomography have all been employed in the assessment of thyroid nodules, and thyroid stimulating hormone assay is useful for confirming a euthyroid state, fine needle aspiration biopsy (FNAB) has proved to be the most efficient diagnostic tool. The findings from FNAB allow avoidance of operative treatment for a large portion of these patients with palpable thyroid nodules, but a diagnosis of "follicular neoplasm" on FNAB usually requires operation, despite the fact that many such patients do ultimately prove to have a benign lesion. The extent of operation in patients undergoing surgery will depend on the diagnostic findings before operation, but unilateral thyroid lobectomy is the minimum procedure when surgery is required. PMID- 12115802 TI - Treatment of neoplastic diseases of the sacrum. PMID- 12115801 TI - Radio-guided parathyroidectomy for primary hyperparathyroidism combined with video-assisted surgery using the solid-state, multi-crystal gamma camera. PMID- 12115803 TI - Examination of the solubilization of drugs by bile salt micelles. AB - The purpose of this review is to provide a critical examination of the reported solubilization of drugs by bile salt micelles. The underlying premise is that with better information regarding the inherent biological complexity, efforts to predict the oral bioavailability of drug will be enhanced. The common means of comparing the reported values was chosen to be the solubilization ratio. This is equal to the moles of drug solubilized per mole of bile salt. The values were segregated according to bile salt type, temperature, ionic strength, and the presence and absence of added lipids. Only segregation by bile salt type was pairwise statistically significant. From the solubilization ratios and the reported values of the aqueous solubility, the logarithms of the mole fraction micelle partition coefficients, log K(m/a), were calculated. The log K(m/w) was found to be correlated with the reported logarithm of the octanol/water partition coefficient. The rank order of slopes of the log K(m/a) as a function of log K(o/w) was cholate approximately taurodeoxycholate > glycocholate approximately taurocholate approximately glycodeoxycholate, with deoxycholate not being statistically different from the other data sets. The slope and intercept for the bile salt mixed micelle systems were 0.600 and 2.44, respectively, which were statistically indistinguishable from glycocholate, taurocholate, and glycodeoxycholate bile salt data. The existence of statistically significant correlations suggests that predicting the solubilization in the intestine may be possible with in vitro measurements if additional information is gathered in the appropriate micellar solutions. PMID- 12115804 TI - Reversal of multidrug resistance-associated protein-mediated daunorubicin resistance by camptothecin. AB - The multidrug-resistance (MR) status of camptothecin (CPT) was investigated in colon adenocarcinoma HT29 cells, leukemia K562, and breast carcinoma MCF7 cells expressing P-glycoprotein (Pgp) and/or MR-associated protein (MRP1). The concentration that induced 50% growth inhibition (IC(50)) against CPT was 0.14 and 0.20 microM in parental K562/WT and MCF7/WT cells, respectively. The drug resistant subline KH30 and MCF7/VP cells, which both overexpress MRP1, presented IC(50) values of 0.63 and 3.10 microM, respectively. The resulting resistance indexes were 3.80 and 12.50, respectively. However, in KH300 cells, a cell line that preferentially overexpresses Pgp, the IC(50) of CPT was 0.08 microM and thus did not exhibit resistance against CPT. In MCF7/DoX cells, preferentially overexpressing Pgp, but also a significant level of MRP1, the IC(50) of CPT was 0.64 microM and thus presented a resistance index of 3.26 against CPT. The cytotoxic effect of CPT was modulated in cells expressing MRP1 (MCF7/VP, HT29 cells) by the specific MRP1 modulators, probenecid and MK571. These results led us to consider CPT as a substrate for MRP1 and a potential modulator of MRP1 activity. To test this hypothesis, we examined the ability of nontoxic concentrations of CPT to sensitize MRP1-overexpressing cells to daunorubicin (DNR). In MCF7/VP and KH30 cells, nontoxic concentrations of CPT were able to enhance cytotoxicity of DNR and its nuclear accumulation. Sequential and simultaneous associations of CPT (100 nM) and DNR provided complete reversal of resistance, thus showing a synergistic effect in KH30 cells. However, simultaneous association (with 10 or 20 nM CPT) had an additive effect in MCF7/VP. These data suggest that CPT could be proposed as a candidate for the reversal of the MRP1 phenotype at clinically achievable concentrations. PMID- 12115805 TI - Effects of low-frequency ultrasound on the transdermal permeation of mannitol: comparative studies with in vivo and in vitro skin. AB - The in vivo and the in vitro correlation of the effects of low-frequency ultrasound (low-frequency sonophoresis, LFS) on the percutaneous penetration of mannitol, a model hydrophilic permeant, was investigated using three in vitro skin models (including full-thickness and split-thickness pig skin, and heat stripped human cadaver skin) and in vivo pig as the animal model. The central objective of this article was to identify the relevant in vitro skin models and ultrasound conditions that may be used in in vitro LFS studies to predict the effects of LFS in vivo on the transdermal delivery of hydrophilic permeants. In this article, by conducting comparative studies of the in vivo pig skin and of the three in vitro skin models under two LFS protocols (a constant ultrasound energy dose protocol, and a constant skin electrical resistance protocol), we demonstrated that: (1) under a constant ultrasound energy dose protocol (protocol A, 5 min LFS), no good correlation was observed between the in vivo skin and the in vitro skin models in terms of the measured skin permeabilities to mannitol. Moreover, the effects of LFS on the barrier functions of the in vivo pig skin, as measured by the enhancement ratio of the skin permeation rate of mannitol and by the reduction of the skin electrical resistance, are much more pronounced than those observed with the excised skin models in vitro; (2) under a constant skin electrical resistance protocol (protocol B) of LFS, a good correlation was found between the skin permeability to mannitol measured using the three in vitro skin models and that of the in vivo pig skin. This result indicates that by utilizing the skin electrical resistance as a quick indicator of the skin permeabilization state due to LFS, the three in vitro skin models can be utilized to predict the transport rate of mannitol across the in vivo skin during LFS; (3) by applying a recently developed skin porous-pathway theory, we demonstrated that within the range of LFS conditions examined, the three in vitro skin models exhibit similar transport properties to mannitol and similar skin effective pore radius values, and hence, represent equivalent skin models for the in vitro LFS studies in the case of hydrophilic permeants; (4) histological studies revealed that the LFS protocol that was shown to be efficacious in enhancing the skin penetration rate of mannitol across the in vivo pig skin, and was also utilized for the in vivo/in vitro skin comparative studies, is safe for the living skin; and (5) through measuring the skin concentration of mannitol in the presence and in the absence of the LFS treatment, we found that the LFS-induced flux enhancement outweighs the enhancement of the skin concentration of mannitol during the LFS studies both in vivo and in vitro. This result suggests that LFS represents a good method of enhancing the systemic absorption of hydrophilic permeants, while it does not significantly alter the vehicle-to-skin partition coefficient for the same class of permeants. PMID- 12115806 TI - Steric stabilization of liposomes by pH-responsive N-isopropylacrylamide copolymer. AB - The aim of this study was to characterize a pH-sensitive liposome formulation bearing a terminally alkylated N-isopropylacrylamide (NIPAM) copolymer with regard to its pH responsiveness, surface properties, and pharmacokinetics. The interacting forces between two lipid bilayers bearing the anchored NIPAM copolymer were measured with a surface force apparatus. The pH-triggered content release was evaluated in buffer before and after incubation in human serum. The pharmacokinetics was determined in rats following the intravenous injection of 67Ga-loaded liposomes with or without the polymer coating. The force measurements between lipid bilayers showed that NIPAM copolymers provide a steric barrier that was dependent on pH. The pH-sensitive liposomes maintained their pH sensitivity after incubation in serum. In vivo, the polymer-coated liposomes exhibited a prolonged circulation time in rats, with an area under the blood concentration time curve that is 1.6-fold higher than the control formulation. This study showed that liposomes can be rendered pH sensitive by anchoring a terminally alkylated NIPAM copolymer at their surface. At neutral pH, the polymer provides a steric barrier that increases the liposome circulation time in vivo. PMID- 12115807 TI - Diffusional release of a solute from a spherical reservoir into a finite external volume. AB - An exact solution has been obtained for the release kinetics of a solute from a spherical reservoir with the burst effect initial condition into a finite external volume. The exact solution is derived based on the time Laplace transform method. The results presented here indicate that as the external fluid volume increases, the cumulative release at any time and the releasable amount of the solute at infinite time increase. In addition, for a given external volume, as the polymeric coat thickness increases the fractional release at any time decreases. Experimentally, cumulative release profiles of theophylline microspheres coated with ethylene vinyl acetate copolymer into different external volumes agreed with the mathematical predictions. PMID- 12115808 TI - Solubilization of hydrophobic drugs in octanoyl-6-O-ascorbic acid micellar dispersions. AB - Alkanoyl-6-O-ascorbic acid esters are easily obtained from vitamin C, and produce self-assembled aggregates in water solutions, with an inner hydrophobic pool surrounded by an external hydrophilic shell. Compared to ascorbic acid, their solubility in oils and fats is greatly enhanced, while the peculiar antioxidant activity is retained in the polar head groups of such surfactants. In virtue of their amphiphilic nature, ascorbic acid-based supramolecular systems can dissolve relevant amounts of hydrophobic, poorly water soluble chemicals such as drugs, vitamins, and so on, and at the same time they provide a suitable shield against oxidative deterioration of valuable materials. In this article we report our study on the self-assembling properties of octanoyl-6-O-ascorbic acid in water, and on the solubilization of some lipophilic molecules in its dispersions. PMID- 12115809 TI - The influence of lipids on stereoselective pharmacokinetics of halofantrine: Important implications in food-effect studies involving drugs that bind to lipoproteins. AB - The objective of this study was to determine the effect of lipids on the pharmacokinetics of halofantrine enantiomers. Rats were given (+/-)-halofantrine HCl 2 mg/kg i.v., or 7 mg/kg orally. Some rats were rendered hyperlipidemic by intraperitoneal administration of poloxamer 407 1 g/kg, followed by (+/-) halofantrine HCl intravenously. In other normolipidemic rats, (+/-)-halofantrine was administered under fasted conditions, or after peanut oil given orally. Halofantrine enantiomer plasma concentrations were considerably (>10-fold) increased in hyperlipidemia. Decreases were noted in the clearance, volume of distribution and the unbound fraction in plasma of the hyperlipidemic rats. Peanut oil caused a significant 28% reduction in clearance of the (-), but not the (+) enantiomer (mean clearance reduced 11%) of halofantrine. After oral halofantrine, peanut oil resulted in a two- to threefold increase in the plasma area under the curves of halofantrine enantiomers. Halofantrine enantiomer pharmacokinetics are highly dependent upon plasma lipid concentrations. Oral lipids may result in a stereoselective interaction at the level of clearance. Because lipids may affect clearance of drugs that bind to lipoproteins, in determining bioavailability of such drugs in food-effect studies, reference intravenous groups should be included to separate true increase in bioavailability from the effects of decreased clearance. PMID- 12115810 TI - Pharmacokinetics and tissue distribution after intravenous administration of a single dose of amphotericin B cochleates, a new lipid-based delivery system. AB - Model independent pharmacokinetic analysis of intravenous (iv) amphotericin B cochleates (CAMB), a new lipid-based drug delivery system, in mice (0.625 mg/kg) shows a two-phase disposition profile in blood [area under the curve of concentration versus time from time zero to infinity (AUC(0-infinity)) = 1.01 microg. h/mL, half-life (t((1/2))) = 11.68 h, volume of distribution at steady state (V(ss)) = 9.59 L/kg, clearance (CL) = 10.36 mL/min/kg and mean residence time from time 0 to infinity (MRT(0-infinity)) = 15.41 h). In target tissues, maximum time (t(max)) ranged from 2 min (spleen and lung) to 10 min (liver) and lungs presented the highest AMB concentration (16.4 microg. h/g) followed by liver (8.56 microg/g), and spleen (6.63 microg/g). In addition, liver and spleen presented the longest elution half-life (75.03 and 66.71 h, respectively), MRT(0 infinity) (98.4 and 86.3 h, respectively), and AMB exposure:liver AUC(0-infinity) = 474 and 116.4 microg. h/g for the spleen. The large V(ss) and the extensive tissue AUC indicate large and efficient ability of cochleates to penetrate and deliver AMB. Differences in tissue uptake mechanism and pharmacokinetic data suggest a crucial role of macrophages in CAMB clearance from blood as well as an essential role of the liver and the spleen in AMB distribution to target tissues. PMID- 12115811 TI - Prediction of aqueous solubility of organic compounds using a quantitative structure-property relationship. AB - A quantitative structure-property relationship (QSPR) was developed for predicting the aqueous solubility of drug-like compounds from their chemical structures. A set of 321 structurally diverse drugs or related compounds, with their intrinsic aqueous solubility collected from literature, was used in this analysis. The data were divided into a training set (n = 267) for building the model and a randomly chosen testing set (n = 54) for assessing the predictive ability of the model. A series of molecular descriptors was calculated directly from chemical structures and a set of eight descriptors, including dipole moment, surface area, volume, molecular weight, number of rotatable bonds/total bonds, number of hydrogen-bond acceptors, number of hydrogen-bond donors and density, was chosen for the final model. The eight-descriptor model generated by multiple linear regression was further optimized by a genetic algorithm guided selection method. The model has a correlation coefficient (r) of 0.95 and a root-mean square (rms) error of 0.56 log unit. It predicts the solubility of testing set compounds with a reasonable degree of accuracy (r = 0.84 and rms = 0.86 log unit). The present model can serve as a tool for medicinal chemists to guide their early synthetic efforts in arriving at appropriate analogs. PMID- 12115812 TI - Dynamics of pharmaceutical amorphous solids: the study of enthalpy relaxation by isothermal microcalorimetry. AB - The structural relaxation time is a measure of the molecular mobility involved in enthalpy relaxation, and thus, is a measure of the dynamics of amorphous (glassy) pharmaceutical solids that determines physicochemical properties and reactivity of drugs in amorphous formulations. In this article we describe a novel method for characterization of structural relaxation using isothermal microcalorimetry, which directly measures the rate of heat release during the relaxation processes. The structural relaxation time is then obtained from a fit of the power data to the derivative version of the Kohlrausch-Williams-Watts (KWW) equation. The relaxation times of quenched and lyophilized samples of saccharides were studied using an isothermal microcalorimeter, the Thermal Activity Monitor (TAM). In addition to the KWW derivative function, a derivative equation of the modified stretched exponential function (MSE) was employed to evaluate TAM data. The later (MSE) appeared to have numerical advantages over the KWW equation. The data demonstrate, as expected, that structural relaxation times of amorphous solids depend on a number of variables, including nature of material, temperature, moisture content, thermal history, etc. Isothermal microcalorimetry with the TAM provides a very fast and reliable way to characterize the dynamics of glassy materials, which in many respects is superior to the conventional DSC approach. To the extent stability and structural relaxation dynamics in the glass are correlated, structural relaxation parameters derived by isothermal microcalorimetry may provide data useful for rational development of stable peptide and protein formulations and for the control of their processing. PMID- 12115813 TI - Physical stability of amorphous pharmaceuticals: Importance of configurational thermodynamic quantities and molecular mobility. AB - This work relates the thermodynamic quantities (Gc, Hc, and Sc) and the molecular mobility values (1/tau) of five structurally diverse amorphous compounds to their crystallization behavior. The model compounds included: ritonavir, ABT-229, fenofibrate, sucrose, and acetaminophen. Modulated temperature DSC was used to measure the heat capacities as a function of temperature for the amorphous and crystalline phases of each compound. Knowledge of the heat capacities and fusion data allowed calculation of the configurational thermodynamic quantities and the Kauzmann temperatures (T(K)) using established relationships. The molecular relaxation time constants (tau) were then calculated from the Vogel-Tammann Fulcher representation of the Adam-Gibbs model. Amorphous samples were heated at 1 K/min and a reduced crystallization temperature, defined as (Tc - Tg)/(Tm-Tg), was used to compare crystallization tendencies. Crystallization was observed for all compounds except ritonavir. The configurational free energy values (Gc) show that thermodynamic driving forces for crystallization follow the order: ritonavir > acetaminophen approximately fenofibrate > sucrose > ABT-229. The entropic barrier to crystallization, which is inversely related to the probability that the molecules are in the proper orientation, followed the order: ritonavir > fenofibrate > ABT-229 > acetaminophen approximately sucrose. Molecular mobility values, which are proportional to molecular collision rates, followed the order: acetaminophen > fenofibrate > sucrose > ABT-229 approximately ritonavir. Crystallization studies under nonisothermal conditions revealed that compounds with the highest entropic barriers and lowest mobilities were most difficult to crystallize, regardless of the thermodynamic driving forces. This investigation demonstrates the importance of both configurational entropy and molecular mobility to understanding the physical stability of amorphous pharmaceuticals. PMID- 12115814 TI - Elimination of polymer interference in chromatographic analysis of estradiol degradation products in a transdermal drug delivery formulation by proper selection of extraction solvents. AB - This article describes the proper selection of extraction solvents to eliminate interference from a polymer matrix to the quantitation of estradiol degradation products in a transdermal formulation by reversed-phase liquid chromatography. The separation is performed by gradient elution with acetonitrile and water as the mobile phase on Inertsil ODS columns. Severe band distortion and insufficient recovery are observed for two geometric degradation products (or impurities) when the sample is prepared by acetonitrile. It is anticipated that the poor resolution and recovery are caused by multiple retention processes due to the reversible binding of degradation products to the polymer matrix (or the impurity polymer interaction). This interaction is eliminated by adding formamide, a solvent that possesses similar properties to the matrix, in the extraction solvent. It is believed that the favorable interaction between formamide and the polymer matrix releases the impurity molecules, and they can then be separated by a single retention mode. It has been confirmed experimentally that the use of formamide in the extraction solvent not only sharpens the peaks tremendously, but also recovers the degradation products completely. PMID- 12115815 TI - Ultrasound-compacted indomethacin/polyvinylpyrrolidone systems: effect of compaction process on particle morphology and dissolution behavior. AB - Indomethacin (IMC)/polyvinylpyrrolidone systems were prepared under different technological conditions, using co-evaporation, kneading, traditional, and ultrasound (US) compaction. The materials thus obtained were milled and sieved and the powders were analyzed by using scanning electron microscopy to evaluate the morphology of the final particles and the fractal dimension of the particle contour. In the case of US-treated particles, scanning electron micrographs suggest that IMC could have partially covered the excipient granule surface, which appears lustrous and smooth, whereas after co-evaporation, the particles display a stratified structure. The external color of the granules, the hot stage microscopy examination, and the absence of the melting peak of the drug in thermograms supports the idea that IMC converts into an amorphous form under US discharge. Each technological treatment performed on the binary mixtures increases the dissolution rate of the drug, with respect to the pure drug and the physical mixture, but to a lesser extent than US compaction. US compaction and co evaporation produce comparable results in improving the release of the drug. Polyvinylpyrrolidone offers better results than beta-cyclodextrin in promoting the dissolution of IMC, when both systems are compacted under US. PMID- 12115816 TI - Prediction of steady-state skin permeabilities of polar and nonpolar permeants across excised pig skin based on measurements of transient diffusion: characterization of hydration effects on the skin porous pathway. AB - The applicability of a two-parameter Fickian diffusion model for predicting the skin steady-state permeability based on measurements of the transient transport of permeants across the skin was tested. Using five model permeants possessing different physicochemical properties and pig skin as the model membrane, the skin permeabilities predicted by the two-parameter Fickian diffusion model were compared with the measured skin permeabilities. Results show that the transient skin permeation profiles of the hydrophobic permeants, estradiol, testosterone, and dolichol, across split-thickness pig skin can be modeled adequately by the two-parameter Fickian diffusion model (with constant parameter values), and therefore, that this model can be utilized to shorten the experimental time required to determine the skin permeabilities of these compounds. However, the skin permeabilities of the highly hydrophilic permeants, mannitol and sucrose, predicted by the two-parameter Fickian diffusion model (with constant parameter values) were significantly lower than the experimentally determined values, indicating that the dominant skin pathway of polar permeants within the excised pig skin undergoes significant structural changes during the in vitro diffusion cell studies. Although the skin permeability values determined experimentally using the traditional steady-state method normally correspond to a highly hydrated skin sample, the two-parameter Fickian diffusion model enables an estimation of the skin permeability of the skin membrane at its less-hydrated state (a condition more representative of in vivo and clinical situations). Using the two-parameter Fickian diffusion model and a recently developed skin porous pathway theory, the effects of skin hydration on the skin porous pathway within the excised pig skin were characterized. Specifically, we found that hydration leads to induction of new pores/reduction of the tortuosity of existing pores within the excised pig skin during the 48 h diffusion cell studies conducted, while the skin average pore radii remain relatively constant (approximately 26 A) for up to 48 h. PMID- 12115817 TI - Interspecies scaling of biliary excreted drugs. AB - The objective of this study is to predict clearance of drugs in humans from animals which are excreted in the bile. Clearance (CL) of eight drugs known to be excreted in the bile were randomly selected from the literature. Scaling of CL was performed using at least three animal species. Using simple allometry, CL x mean life-span potential (MLP) or CL x brain weight, CLs of studied drugs were predicted in humans. The choice of one of the methods depended on the 'rule of exponents' as described by Mahmood and Balian. A 'correction factor' was calculated by adjusting bile flow rate based on the species body weight (bile flow = mL/day/kg body weight) or liver weight (bile flow = mL/day/kg liver weight). Using the 'rule of exponents' and combining it with the 'correction factor', the CLs of biliary excreted drugs were predicted in humans. Predicted CLs in humans from animals using simple allometry were several times higher for all eight drugs (% error [range] = 46-1703). Using the 'rule of exponents' and combining it with a 'correction factor' as described in this report provided a substantial improvement (% error [range] = 5-91) in the prediction of CL for biliary excreted drugs. The results of this study indicate that the CL of a biliary excreted drug may be overpredicted in humans and by applying the 'correction factor' employed here, the predictability of drug CL in humans from animal data may be significantly improved. PMID- 12115818 TI - Correlation between stratum corneum/water-partition coefficient and amounts of flufenamic acid penetrated into the stratum corneum. AB - The stratum corneum of various donors differs in particular in the composition of the lipoidal phase. Considering the drug amounts penetrating into the stratum corneum a simple methodology to correlate these differences in the stratum corneum composition with the drug amounts detectable within the stratum corneum is desirable. Penetration experiments investigating several incubation times were carried out with three different skin flaps using the Saarbruecken penetration model and the lipophilic model drug flufenamic acid. The drug amounts within the stratum corneum were obtained with the tape-stripping technique, while the drug amounts present in the deeper skin layers were achieved by cryosectioning. The stratum corneum/water-partition coefficient was determined with the same three skin flaps to characterize the lipoidal stratum corneum phase in general, and the differences were attributed to the different amounts of ceramides and sterols. In addition, for the lipophilic drug flufenamic acid, a direct linear correlation was found between the stratum corneum/water-partition coefficients and the drug amounts penetrated into the stratum corneum for all investigated time intervals (correlation coefficients of r(30 min) = 0.998, r(60 min) = 0.998 and r(180 min) = 0.987). In contrast to the stratum corneum/water-partition coefficients, the determination of a corresponding relationship for the stratum corneum and the deeper skin layers failed due to the reason that steady-state conditions could not be achieved for the deeper skin layers during the investigated time intervals. In summary, the stratum corneum/water-partition coefficients offer the possibility to predict drug amounts within the stratum corneum of different donor skin flaps without a time consuming determination of the lipid composition of the stratum corneum. PMID- 12115819 TI - Effects of mechanical processing on phase composition. AB - One factor that must be considered during drug development process is that various types of pharmaceutical manufacturing can alter the physical characteristics of the drug entity. These effects become particularly important during scale-up of processing operations, because new and unanticipated results can become manifest in systems of insufficient characterization. Any transformed drug substance or altered dosage form could exhibit an altered solubility or dissolution rate that might produce an undesirable bioavailability profile. Some of the more interesting mechanical manipulations that have the potential to yield problems include particle size reduction and compression, and such investigations are the focus of this minireview. PMID- 12115820 TI - Shear-induced degradation of plasmid DNA. AB - The majority of gene therapy clinical trials use plasmid DNA that is susceptible to shear-induced degradation. Many processing steps in the extraction, purification, and preparation of plasmid-based therapeutics can impart significant shear stress that can fracture the phosphodiester backbone of polynucleotides, and reduce biological activity. Much of the mechanistic work on shear degradation of DNA was conducted over 30 years ago, and we rely heavily on this early work in an attempt to explain the empirical observations of more recent investigations concerning the aerosolization of plasmids. Unfortunately, the sporadic reports of shear degradation in the literature use different experimental systems, making it difficult to quantitatively compare results and reach definitive mechanistic conclusions. In this review, we describe the forces imparted to DNA during shear stress, and use published data to quantitatively evaluate their relative effects. In addition, we discuss the effects of molecular weight, strain rate, particle size, flexibility, ionic strength, gas-liquid interfaces, and turbulence on the fluid flow degradation of supercoiled plasmid DNA. Finally, we speculate on computational methods that might allow degradation rates in different experimental systems to be predicted. PMID- 12115821 TI - Factors affecting the deposition of inhaled porous drug particles. AB - Recent findings indicate that the inhalation of large manufactured porous particles may be particularly effective for drug delivery. In this study, a mathematical model was employed to systematically investigate the effects of particle size, particle density, aerosol polydispersity, and patient ventilatory parameters on deposition patterns of inhaled drugs in healthy human lungs. Aerodynamically similar particles with densities of 0.1, 1.0, and 2.0 g/cm(3) were considered. Particle size distributions were defined with mass median aerodynamic diameters (MMADs) ranging from 1 to 3 microm and geometric standard deviations ranging from 1.5 to 2.5, representing particles in the respirable size range. Breathing rates of 30 and 60 L/min with tidal volumes of 500 to 3000 mL were assumed, simulating shallow to deep breaths from a dry powder inhaler. Particles with a high density and a small geometric diameter had slightly greater deposition fractions than particles that were aerodynamically similar, but had lower density and larger geometric size (typical of manufactured porous particles). This can be explained by the fact that particles with a small geometric diameter deposit primarily by diffusion, which is a function of geometric size but is independent of density. As MMAD increased, the effect of density on deposition was less pronounced because of the decreased efficiency of diffusion for large particles. These data suggest that polydisperse aerosols containing a significant proportion of submicron particles will deposit in the pulmonary airways with greater efficiency than aerodynamically similar aerosols comprised of geometrically larger porous particles. PMID- 12115822 TI - Rapid determination of liposome-water partition coefficients (Klw) using liposome electrokinetic chromatography (LEKC). AB - Liposome Electrokinetic Chromatography (LEKC) provides a simple and facile approach for determining liposome-water partition coefficients, Klw. LEKC is a Capillary Electrophoresis (CE) technique where liposomes are incorporated into a buffer solution and act as a pseudostationary phase providing sites of interaction for solutes. The retention factors of solutes in LEKC are directly proportional to Klw. This article describes how LEKC can be used to determine Klw for both neutral and charged solutes. The Klw values for a group of neutral aromatic compounds, beta-blockers, and other drugs are reported. In addition, the usefulness of two quantitative structure partition relationships (QSPR) for estimation of Klw is demonstrated. One is the logarithmic linear relationship between liposome water and octanol-water partition coefficients (Pow). The other is the Linear Solvation Energy Relationship (LSER). The calculated Klw values from the two QSPR agree nicely with the observed values and with one another. PMID- 12115824 TI - Effect of omeprazole on oral and intravenous RS-methadone pharmacokinetics and pharmacodynamics in the rat. AB - The effect of omeprazole on oral and intravenous (iv) RS-methadone pharmacokinetics and pharmacodynamics was studied in awake, freely moving rats, which were divided in four groups: oral RS-methadone (3 mg/kg) was given (a) to a control group (CO(oral)) (n = 65) and (b) to an omeprazole pretreated group (OP(oral)) (n = 77), and iv RS-methadone (0.35 mg/kg) was administered (c) to a control group (CO(iv)) (n = 86) and (d) to an omeprazole pretreated group (OP(iv)) (n = 86). Omeprazole (2 mg/kg) was given iv 2 h before RS-methadone. Plasma concentrations of RS-methadone (Cp) were determined by high-performance liquid chromatography and analgesic response by tail flick for 0-180 min (oral) and 0-120 min (iv). RS-Methadone rate of absorption (mean +/- SE) was faster in OP(oral) (k(01) = 0.31 +/- 0.04 min(-1)) than in CO(oral) (k(01) = 0.05 +/- 0.007 min(-1)), consequently plasma peak concentrations (C(max)) were greater (197.41 +/- 33.70 ng/mL versus 83.54 +/- 7.97 ng/mL) and the time to reach C(max) (t(max)) was shorter (11.23 +/- 1.32 min versus 39.18 +/- 1.74 min). Mean area under the Cp-time curve (AUC(0-infinity)) and hence bioavailability of oral RS methadone were increased by omeprazole without significant changes in the elimination. Omeprazole did not affect the pharmacokinetics of iv RS-methadone. The changes of the analgesic effect of RS-methadone as a function of time were similar in all four groups. In the CO(oral) group, Cp and analgesic effect were defined by the E(max) model. The relationship between Cp and drug effect in the OP(oral) group showed a counterclockwise hysteresis (k(e0) of 0.018 +/- 0.006 min(-1)). For the iv groups (CO(iv) and OP(iv)), the Cp-analgesic effect relationship was described by an E(max) sigmoid model and omeprazole did not affect the pharmacodynamic parameters. It is concluded that omeprazole causes an increase in the bioavailability of oral RS-methadone without modifying the analgesic response but affecting the Cp-effect relationship. PMID- 12115823 TI - The effect of laser treatment on skin to enhance and control transdermal delivery of 5-fluorouracil. AB - The effect of three lasers (i.e., the ruby, erbium:YAG, and CO2) on the ability to enhance and control skin permeation of 5-fluorouracil (5-FU) was studied in vitro. Light microscopic and ultrastructural (scanning electron microscopic) changes in the nude mouse skin were also compared for these lasers. The histological observations and permeation profiles of each laser differed because the three lasers produce different physical and physiologic effects when striking the skin. The skin permeation of 5-FU could be moderately promoted by a single photomechanical wave generated by the ruby laser (at 4.0 and 7.0 J/cm(2)) without adversely affecting the viability or structure of the skin. The stratum corneum (SC) layer in the skin was partly ablated by an erbium:YAG laser, resulting in a greater enhancement effect on skin permeation of 5-FU. The flux of 5-FU across erbium:YAG laser-treated skin was 53-133-fold higher than that across intact skin. Both SC ablation and a thermal effect may contribute to the effect of the CO2 laser on skin structure. Lower energies of the CO2 laser did not modulate 5 FU permeation. A 36-41-fold increase in 5-FU flux was observed after exposure to higher fluences (4.0 and 7.0 J/cm(2)) of the CO2 laser. Histological changes induced by both the erbium:YAG and CO2 lasers had completely recovered within 4 days. PMID- 12115826 TI - Theoretical and experimental investigations on the morphology of pharmaceutical crystals. AB - We investigated the morphologies of three polymorphs of 1,3-di(cyclopropylmethyl) 8-aminoxanthine, a compound of pharmaceutical importance. We compared the experimental morphologies with those predicted by theoretical methods. We also predicted the elastic constants of the three polymorphs. These results are used to help assess the stabilities of the three polymorphs and compare the abilities of theoretical methods to reproduce experimental morphologies. PMID- 12115825 TI - Terpenes in propylene glycol as skin-penetration enhancers: permeation and partition of haloperidol, Fourier transform infrared spectroscopy, and differential scanning calorimetry. AB - The respective alcoholic terpenes carvacrol, linalool, and alpha-terpineol were used at 5% w/v in propylene glycol (PG) to increase the in vitro permeation of haloperidol (HP) through human skin. The possible enhancement mechanism was then elucidated with HP-stratum corneum (SC) binding studies, Fourier transform infrared spectroscopy, and differential scanning calorimetry. The greatest increase in the permeation of HP was achieved with linalool followed by carvacrol and terpineol. HP permeation with linalool was predicted to reach a therapeutic plasma concentration and therapeutic daily-permeated amounts. Carvacrol increased lag time, which was attributed to slow redistribution of the enhancer within SC. Carvacrol increased the partition of the drug to the pulverized SC. Pure PG extracted lipids from SC but less than that achieved by the terpenes in PG. Terpenes extracted lipids to a similar extent. An increase in bilayer cohesion in the remaining lipids present in the SC could be attributed to the alignment of terpenes within the lipid bilayer. The higher permeation with linalool was attributed to its molecular orientation within the lipid bilayer. Terpenes showed different rates of SC dehydration but did not change the percentages of secondary structures of keratin. PMID- 12115827 TI - Development and in vivo evaluation of buccal tablets prepared using danazol sulfobutylether 7 beta-cyclodextrin (SBE 7) complexes. AB - The aim of the present work was to develop a mucoadhesive controlled-release formulation of danazol-sulfobutylether 7 beta-cyclodextrin (SBE 7) complex and to evaluate the feasibility of improving the bioavailability of danazol via the buccal route. Different types of polymers, polycarbophil (PC) and hydroxypropylmethyl cellulose (HPMC) were mixed with danazol-SBE 7 complex and compressed into tablets. These tablets were evaluated for their dissolution and mucoadhesion properties and for drug absorption in female beagle dogs. Increased mucoadhesion was observed for PC-containing tablets compared with HPMC tablets. As the concentration of polymer increased, drug release decreased, and PC containing tablets gave slower release compared to HPMC tablets. In vivo bioavailability performed in dogs showed that the perorally administered danazol SBE 7 complex and the danazol-SBE 7 (in PC matrix) buccal tablets had absolute bioavailabilities of 64% and 25%, respectively, that are significantly greater than 1.8% observed for the commercial formulation Danocrine. The increased bioavailability was attributed to the enhanced solubility consequent to complexation, and the possible avoidance of first-pass metabolism upon buccal administration. PMID- 12115828 TI - Physical characterization of a chitosan-based hydrogel delivery system. AB - Hydrogel formation from a mixture of biocompatible chitosan, beta-glycerol phosphate, and hydroxyethyl cellulose was studied. The rheological properties of the formed hydrogels were examined using a Bohlin VOR rheometer. The effect of hydrogel composition and temperature on both the gelation rate and the elastic strength of hydrogels was investigated, from which possible hydrogel formation mechanisms were inferred. The formed hydrogels as potential vehicles for delivering pilocarpine were examined. The advantages and disadvantages of chitosan/hydroxyethyl cellulose hydrogel as a delivery system are discussed. PMID- 12115829 TI - Characterization of ibuprofen as a nontraditional plasticizer of ethyl cellulose. AB - This study describes the characterization of the plasticizing properties of ibuprofen (IBP) on hot-melt extruded ethyl cellulose (EC). The thermal behavior of hot-melt extrudates containing 0, 5, 10, and 20% (w/w) IBP was evaluated using modulated temperature differential scanning calorimetry. By means of comparison, co-evaporates containing the same concentrations of IBP and EC, were also evaluated. Both methods yielded solid solutions having one glass transition temperature indicating compatibility between drug and polymer. A similar decrease in glass transition temperature was noticed with increasing IBP concentration in the solid solutions prepared via both methods, indicating its plasticizing effect. The plasticizing efficiency was of the same magnitude as for the traditionally used plasticizers. Infrared spectroscopy was performed for better understanding of the chemical interactions in the molecular dispersions and confirmed the existence of hydrogen bonds between IBP and EC. Overall, the study has highlighted the plasticizing properties of IBP on EC during hot-melt extrusion. PMID- 12115830 TI - Solubility, ionization, and partitioning behavior of unsymmetrical disulfide compounds: alkyl 2-imidazolyl disulfides. AB - Alkyl 2-imidazolyl disulfide compounds are novel antitumor agents, one of which is currently being evaluated in Phase I clinical trials. These molecules contain an unsymmetrical disulfide fragment, the lipophilic and electronic contributions of which are still not defined in the literature. Lipophilicity, ionization, and solubility of a number of alkyl 2-imidazolyl disulfides were studied. Based on the additivity of lipophilicity and ionization properties, the contribution of the unsymmetrical disulfide fragment to lipophilicity and ionization was elucidated. The unsymmetrical disulfide fragment contributed a Rekker's hydrophobic constant of 0.761 to the lipophilicity of these compounds and an approximated Hammett constant (sigma) of 0.30 to their ionization. The applicability of the general solubility equation (GSE) proposed by Jain and Yalkowsky in predicting the aqueous solubility of these analogs was evaluated. The GSE correctly ranked the aqueous solubilities of these compounds and estimated their log molar solubilities with an average absolute error of 0.35. PMID- 12115832 TI - Measuring the surface area of aluminum hydroxide adjuvant. AB - The traditional method of determining surface area, nitrogen gas sorption, requires complete drying of the sample prior to analysis. This technique is not suitable for aluminum hydroxide adjuvant because it is composed of submicron, fibrous particles that agglomerate irreversibly upon complete removal of water. In this study, the surface area of a commercial aluminum hydroxide adjuvant was determined by a gravimetric/FTIR method that measures the water adsorption capacity. This technique does not require complete drying of the adjuvant. Five replicate determinations gave a mean surface area of 514 m(2)/g and a 95% confidence interval of 36 m(2)/g for a commercial aluminum hydroxide adjuvant. The X-ray diffraction pattern and the Scherrer equation were used to calculate the dimensions of the primary crystallites. The average calculated dimensions were 4.5 x 2.2 x 10 nm. Based on these dimensions, the mean calculated surface area of the commercial aluminum hydroxide adjuvant was 509 m(2)/g, and the 95% confidential interval was 30 m(2)/g. The close agreement between the two surface area values indicates that either method may be used to determine the surface area of aluminum hydroxide adjuvant. The high surface area, which was determined by two methods, is an important property of aluminum hydroxide adjuvants, and is the basis for the intrinsically high protein adsorption capacity. PMID- 12115831 TI - Equilibrium loading of cells with macromolecules by ultrasound: effects of molecular size and acoustic energy. AB - Ultrasound has been shown to deliver small compounds, macromolecules, and DNA into cells, which suggests potential applications in drug and gene delivery. However, the effect of molecular size on intracellular uptake has not been quantified. This study measured the effect of molecule size (calcein, 623 Da; bovine serum albumin, 66 kDa; and two dextrans, 42 and 464 kDa) on molecular uptake and cell viability in DU145 prostate cancer cells exposed to 500 kHz ultrasound. Molecular uptake in viable cells was shown to be very similar for small molecules and macromolecules and found to correlate with acoustic energy exposure. Molecular uptake was seen to be heterogeneous among viable cells exposed to the same ultrasound conditions; this heterogeneity also correlated with acoustic energy exposure. In a fraction of these cells, molecular uptake reached thermodynamic equilibrium with the extracellular solution for the small molecule and all three macromolecules. The results demonstrate that ultrasound provides a means to load viable cells with large numbers of macromolecules, which may be of use for laboratory and possible clinical drug delivery applications. PMID- 12115833 TI - Nasal administration of low molecular weight heparin. AB - The main objective of this study was to determine if the systemic absorption of therapeutic amounts of heparin was possible following nasal administration. Sprague-Dawley rats received nosedrops containing a low molecular weight heparin (LMWH) or unfractionated heparin (UFH) formulated with or without tetradecylmaltoside (TDM). TDM is a nonionic surfactant that has been previously shown to be a potent absorption enhancer in studies with peptide drugs. LMWH/UFH absorption was determined by measuring plasma anti-Factor Xa activity. The inclusion of 0.25% TDM in nasal formulations containing LMWH resulted in a significant increase in the C(max) and area under the curve (AUC) of anti-Factor Xa activity when compared to LMWH formulated in saline alone. The addition of TDM to a nasal formulation containing UFH resulted in a much smaller increase in the C(max) and the AUC of anti-Factor Xa activity. The absolute bioavailability of LMWH was increased from 4.0 +/- 0.4% in the absence of TDM to 19 +/- 0.3% in the presence of TDM. The reversibility of the absorption enhancing effect of TDM was studied by applying LMWH nasally 60 or 120 min after the enhancer. The effect of TDM on the nasal epithelia appeared to be rapidly reversible. In conclusion, nasal delivery of LMWH, but not UFH, was successful when an absorption enhancer was included to increase nasal permeability. PMID- 12115834 TI - Interaction of methylparaben preservative with selected sugars and sugar alcohols. AB - The interaction of methylparaben preservative with selected sugars (glucose, fructose, sucrose, lactose, maltose, cellobiose) and sugar alcohols (lactitol, maltitol) were demonstrated in this study. It was observed that the formation of transesterification reaction products between methylparaben and the selected sugars occurred only under mild reaction conditions (e.g., pH 7.4 at 50 degrees C ), which were confirmed by HPLC-UV studies and mass spectrometry. On the other hand, under alkaline conditions and high temperature, degradation of the sugars predominated. Because sugars could easily undergo many possible degradation reactions and isomerization including on-column anomerization, the chromatograms of the reaction products were more complicated than those obtained from sugar alcohols. Sucrose, a nonreducing sugar, was much more stable than other selected sugars. The chromatogram of the transesterification reaction products of methylparaben with sucrose clearly showed eight peaks, which were likely to correspond to the same number of hydroxyl groups of sucrose. To compare the rate of the transesterification reaction of methylparaben with sucrose to that with sorbitol, kinetic studies were carried out. Similar rate constants were observed: 5.4 x 10(-7) L mol(-1) s(-1) and 4.9 x 10(-7) L mol(-1) s(-1) for sucrose and sorbitol, respectively. PMID- 12115835 TI - Kit formulation for the preparation of radioactive blue liposomes for sentinel node lymphoscintigraphy. AB - A radioactively labeled blue liposome formulation was developed for use in presurgical lymphoscintigraphy and intraoperative sentinel node localization to avoid the differing injection site clearance kinetics of the conjunctive use of separate formulations of low molecular weight blue dye and radioactively labeled macromolecular sulfur colloid. Blue liposomes containing glutathione in the internal aqueous phase were prepared from blue dyed lipids obtained by covalently binding Reactive Blue II to phosphatidylethanolamine (PE) fractions of phospholipid extracts. Four phospholipid extracts with differing PE fractions and a centrifugation technique were evaluated with the goal of maximizing the blue color intensity of the formulation. Stability of the formulations was evaluated by studying radiolabeling efficiency (using a membrane permeable lipophilic carrier of the commonly used diagnostic radionuclide, technetium-99m) and particle-size distribution over 30-60-day periods. Blue color was not altered by varying the PE content, while centrifugation was an effective and convenient method to maximize the blue color intensity of the final preparation. The particle size distribution of the prepared liposomes ranged from 200-300 nm (considered ideal for lymphoscintigraphy studies) and did not change significantly, while radiolabeling efficiency exceeded 80% for up to 1 month. The described kit formulation for the preparation of radiolabeled blue liposomes is suitable for commercial production allowing widespread clinical use. The combination of a means of visual identification and tracking of the liposomes through the lymphatic channels along with the ability to trace the preparation using standard radiation detection instrumentation provides the surgeon with an improved radiolabeled compound for lymphoscintigraphy and intraoperative sentinel lymph node identification. PMID- 12115836 TI - Solution equilibria of deferoxamine amides. AB - The physico-chemical solution properties of deferoxamine were modified by acylating the terminal amino group with short-chain aliphatic, succinic, and methylsulphonic moieties. The analog iron(III)-binding constants and stabilities under physiological conditions were determined to confirm that the iron binding ability of the parent molecule was retained following modification. The proton dissociation constants of the lipophilic deferoxamine analogs were determined by potentiometric titration and nonlinear least-squares analysis. However, because the iron(III) binding complex is fully formed below pH 2, the metal-ligand equilibria could not be studied using potentiometric methods. The iron binding constants of the deferoxamine analogs were determined by spectrophotometrically following the proton-dependent exchange of iron with EDTA in the pH range of 4.0 to 6.5 and solving mass balance equations. The proton-dissociation constants and the iron binding constants of the lipophilic deferoxamine analogs were comparable to those of deferoxamine. However, at physiological conditions, the iron-binding complex of the most lipophilic butylamide derivative was slightly less stable and the succinamide derivative complex was slightly more stable. Like deferoxamine, the hydroxamate groups of the analogs were unhindered and free to form a 1:1 coordination complex with iron(III). Consequently, changes in aqueous solvation, conformation, and steric interference, imparted by the modifications at the terminal amino group of deferoxamine, may have affected the stabilities of the iron(III) complex and the efficiency of iron binding. PMID- 12115837 TI - Interconversion pharmacokinetics of eplerenone, a selective aldosterone blocker, and its lactone-ring open form. AB - The interconversion pharmacokinetics of eplerenone and its lactone-ring open form, SC-70303, were examined in dogs using a stable isotope method. [13CD3]EP and SC-70303 were coadministered orally (10 mg/kg) and intravenously (5 mg/kg) as aqueous solutions under fasted conditions. After I.V. administration of [13CD3]EP, the mean AUC of [13CD3]EP was 16.0 h. microg/mL, while the C(max), T(max), and AUC for [13CD3]SC-70303 acid were 0.744 microg/mL, 0.5 h, and 3.49 h. microg/mL, respectively. After I.V. administration of SC-70303, the AUC for SC 70303 acid was 6.36 h. microg/mL, while the C(max), T(max), and AUC for EP were 2.26 microg/mL, 0.5 h, and 9.48 h. microg/mL, respectively. After oral administration of [13CD3]EP, the C(max), T(max), and AUC for [13CD3]EP were 6.01 microg/mL, 0.5 h, and 27.7 h. microg/mL, respectively, and the corresponding values for [13CD3]SC-70303 acid were 0.972 microg/mL, 0.75 h, and 5.52 h. microg/mL, respectively. After oral administration of SC-70303, the C(max), T(max), and AUC for EP were 1.38 microg/mL, 0.83 h, and 9.29 h. microg/mL, respectively, and the corresponding values for SC-70303 acid were 0.330 microg/mL, 0.67 h, and 2.19 h. microg/mL, respectively. The systemic availability was 90% for [13CD3]EP and 17.5% for SC-70303 acid. EP and SC-70303 acid were rapidly interconvertible in the dog. The percentage of dose converted to [13CD3]SC-70303 acid following I.V. administration of [13CD3]EP was 55.0%, while the percentage of dose converted to EP following I.V. administration of SC-70303 was 60.2%. PMID- 12115838 TI - Formulation and food effects on the oral absorption of a poorly water soluble, highly permeable antiretroviral agent. AB - DPC 961 is a low-solubility, high-permeability, second-generation non-nucleoside reverse transcriptase inhibitor. The purpose of these studies was to evaluate the effects of drug substance and formulation variables on DPC 961 oral absorption, and to compare fed and fasted state oral absorption. To accomplish this, groups of four to six dogs were dosed with various formulations of DPC 961 under fasted or fed conditions, and DPC 961 pharmacokinetics were examined. Absolute oral bioavailability, based on i.v. AUC in the same dogs, was 24% after a suspension dose in fasted dogs and was 51% in fed dogs. Bioavailability with an unoptimized tablet formulation was 30% in fasted dogs and 86% in fed dogs. DPC 961 oral absorption was shown to be dependent on drug substance particle size in fasted dogs, after dosing with a tablet formulation where only the drug substance particle size was varied, but there was no difference in fed dogs. AUC and C(max) increased in proportion with increases in tablet strength from 100 to 400 mg, using tablets manufactured from a common granulation. Tablets made with 50 and 66% drug loadings showed similar relative oral bioavailabilities. Tablets prepared with two different polymorphic forms of DPC 961 were also compared, and these were found to be equivalent. These studies provided a useful component of the formulation development process, to help identify and control the variables affecting oral absorption of this potential new therapeutic agent. PMID- 12115839 TI - Formulation pH modulates the interaction of insulin with chitosan nanoparticles. AB - Previous studies on chitosan-insulin nanoparticles have reported diverse encapsulation efficiency and insulin release profiles despite similar formulation and preparation procedures. This study examined the efficiency and mechanism of association of insulin with chitosan nanoparticles in the pH range of 2.3 to 6.3. Nanoparticles of 237 to 235 nm were prepared by ionotropic gelation of chitosan with tripolyphosphate counterions. Insulin was quantified by an RP-HPLC method. The insulin association efficiency (AE) spanned a broad range from 2 to 85%, and was highly sensitive to formulation pH. Highest AE was measured at insulin loading concentrations >/= 4.28 U/mL and pH 6.1, close to the pI of native insulin and the pK(a) of chitosan. This association, attributed to physical adsorption of insulin through hydrophobic interactions with chitosan, was labile, and the associated insulin rapidly and completely released by dilution of the nanoparticles in aqueous media of pH 2 to 7.4. AE obtained at pH 5.3 was less than half that measured at pH 6.1 at corresponding insulin concentration, but the association at pH 5.3 appeared to be based on stronger interactions, because the release of insulin was pH-dependent and recovery was less than 25% even upon disintegration of the chitosan matrix. Interaction of insulin with the chitosan nanoparticles rendered the protein more susceptible to acid and enzymatic hydrolyses, the effects being more predominant in nanoparticles prepared at pH 5.3 than at pH 6.1. PMID- 12115840 TI - VLDL-resembling phospholipid-submicron emulsion for cholesterol-based drug targeting. AB - The objective of the current study was to develop and evaluate VLDL-resembling phospholipid-submicron emulsion (PSME) as a carrier system for new cholesterol based compounds for targeted delivery to cancer cells. BCH, a boronated cholesterol compound, was originally developed in our laboratory to mimic the cholesterol esters present in the LDL and to follow a similar pathway of cholesterol transport into the rapidly dividing cancer cells. The VLDL-resembling system was then designed to solubilize BCH, facilitate the interaction with LDL, and thus assist the BCH delivery to cancer cells. BCH-containing PSME was prepared by sonication. Chemical compositions and particle sizes of different PSME fractions were determined. The lipid structure of PSME and location of BCH in the formulation were assessed based on experimental results. Density gradient ultracentrifugation fractionated the emulsion into three particle-size populations with structures and compositions resembling native VLDL. In vitro interaction between PSME and LDL was evident by agarose electrophoresis, as both formed a single band with an intermediate mobility. The transfer of BCH from PSME to LDL was also observed in the presence of other serum components including serum proteins. Cell culture data showed sufficient uptake of BCH in rat 9L glioma cells (> 50 microg boron/g cells). In conclusion, this system has the capability to incorporate the cholesterol-based compound, interact with native LDL, and assist the delivery of this compound into cancer cells in vitro. PMID- 12115841 TI - Characterization of the interaction of 2-hydroxypropyl-beta-cyclodextrin with itraconazole at pH 2, 4, and 7. AB - Phase-solubility techniques were used to assess the effect of pH on itraconazole complexation with 2-hydroxypropyl-beta-cyclodextrin (HPbetaCD). In addition, molecular modeling using beta-cyclodextrin as a surrogate for HPbetaCD was completed. Data suggested A(p)-type solubility relationships, indicating higher order complexation at higher HPbetaCD concentrations. Stability constants were derived from the solubility isotherms using a simplex optimization procedure. At pH 2 (2 units below the pK(a4)), a 1:2 complex formation was observed, whereas at pH 4 (i.e., the pK(a4) for itraconazole) and at pH 7, 1:3 complexation occurred. The lower order of complexation observed at lower pH may be related to substructure protonation which reduced HPbetaCD interaction. Molecular mechanics also suggest 1:3 complex formation for the neutral species, indicating that possible interaction sites may include (in order of binding) triazole > 1,4 diaminophenyl > 2-butyl approximate, equals piperazine. PMID- 12115842 TI - Dehydration kinetics of neotame monohydrate. AB - The dehydration of neotame monohydrate was monitored at various temperatures by differential scanning calorimetry (DSC), thermogravimetry (TGA), hot-stage microscopy (HSM), powder X-ray diffractometry (PXRD), and (13)C solid-state nuclear magnetic resonance (SSNMR) spectroscopy. This work emphasizes kinetic analysis of isothermal TGA data by fitting to various solid-state reaction models and by model-free kinetic treatment. The dehydration of neotame monohydrate follows the kinetics of a two-dimensional phase boundary reaction (R2) at 40-50 degrees C with an activation energy of 75 +/- 9 kJ/mol, agreeing well with 60-80 kJ/mol from model-free kinetics. At a low heating rate in DSC and TGA, neotame monohydrate undergoes dehydration to produce anhydrate Form E, which then converts to anhydrate Form A, followed by the melting of A. Neotame monohydrate under dry nitrogen purge at 50 mL/min undergoes partial isothermal dehydration at 50 degrees C to produce neotame anhydrate Form A. When neotame monohydrate is heated very slowly from 50 to 65-70 degrees C over 24 h, pure Form A is obtained. PMID- 12115843 TI - Face specific surface properties of pharmaceutical crystals. AB - A variety of surface specific techniques have been used to determine the face specific structure, chemistry, and wettability of model pharmaceutical crystals, i.e., N,n-octyl-d-gluconamide and sulfathiazole (polymorphic forms I and III). The surface energetics of individual crystal faces were investigated by studying their wetting characteristics and interaction with chemically modified silica spheres using colloid probe atomic force microscopy (AFM). Contact angles (dynamic and static), interaction forces, and adhesion properties have been shown to correlate strongly with the face specific surface chemistry. This, in turn, is controlled by the molecular arrangement at the specific crystal face, which has been characterized by time-of-flight secondary-ion mass spectrometry (ToF SIMS) and inferred from molecular models. Of specific note, the magnitude of the adhesion force between a crystal face and a hydrophobic colloid probe is related linearly to the face-specific equilibrium contact angle. These studies further our understanding of the face-specific properties of pharmaceutical crystals and have implications when considering processing, formulation and delivery. PMID- 12115844 TI - Solubility of E2050 at various pH: a case in which apparent solubility is affected by the amount of excess solid. AB - The solubility of E2050, supplied as a dihydrochloride salt, in aqueous solutions at different pHs was studied. Two pK(a)s controlling the equilibrium between the various protonated species were determined. The solubility-pH profile of E2050 is expected to be high in acidic solutions because protonated species are formed and to be low in alkaline conditions due to the formation of hydrophobic free base. The solubility is also affected by chloride ion, a common ion for this drug substance. Two solubility products (K(sp)) were determined corresponding to the solubility of di-HCl salt and mono-HCl salt. Based on the pK(a)s (3.10 and 7.71), the solubility products with chloride (2.92 and 3.77 as corresponding pK(sp)), and the solubility of free base (2 x 10(-5) M), the solubility in solutions with different pH and different levels of chloride ion can be predicted. The prediction of the solubility change during the dilution of E2050 parenteral formulations by saline was also demonstrated. Furthermore, the present study presents an interesting example in which an apparent solubility can be different if varying (excess) amounts of salt are added to the solution. In this case, excess chloride ion suppresses the solubility in the pH region where mono-HCl salt controls the solubility. PMID- 12115845 TI - Synthetic long-chain alkyl maltosides and alkyl sucrose esters as enhancers of nasal insulin absorption. AB - A series of new glycosides with extended alkyl side-chains (C(13-16)) linked to maltose or sucrose were synthesized and tested for their efficacy in enhancing nasal insulin absorption in anesthetized rats. The new reagents were compared to previously tested alkylglycosides with shorter alkyl side chains (C(8-12)). Dose response studies revealed that within the family of alkylmaltoside derivatives, (C(8-16)), maximal increases in insulin absorption took place when tetradecylmaltoside (C(14)) was added to the formulation. Pentadecylmaltoside (C(15)) and hexadecylmaltoside (C(16)) were less potent at increasing insulin absorption, although both reagents achieved maximal effects when used at higher concentrations. Within the family of alkanoylsucrose derivatives, tridecanoylsucrose (C(13)) and tetradecanoylsucrose (C(14)) were most potent at increasing insulin absorption. Cross-comparisons between alkylmaltoses and alkanoylsucroses showed that the alkyl chain length had a greater impact than the glycoside moiety in determining the potency of a potential insulin-absorption enhancing agent. When tetradecylmaltoside was applied to the nasal mucosa 15 min before insulin was applied, the enhanced insulin absorption was still observed. PMID- 12115846 TI - Preparation, characterization, and drug release behaviors of drug nimodipine loaded poly(epsilon-caprolactone)-poly(ethylene oxide)-poly(epsilon-caprolactone) amphiphilic triblock copolymer micelles. AB - Amphiphilic triblock copolymers, poly(epsilon-caprolactone)-poly(ethylene oxide) poly(epsilon-caprolactone) (PCL-PEO-PCL), were synthesized by ring opening polymerization of epsilon-caprolactone initiated with the hydroxyl functional groups of poly(ethylene glycol) at both ends of the chain. The micelles composed of this type of copolymer had such a structure that both ends of the PEO chain were anchored to the micelle. The critical micelle concentration of the block copolymer in distilled water was determined by a fluorescence probe technique using pyrene. As the hydrophobic components of the block copolymer increased, the critical micelle concentration value decreased. To estimate the feasibility as novel drug carriers, the block copolymer micelles were prepared by precipitation of polymer from acetone solution into water. From the observation of transmission electron microscopy, the micelles exhibited a spherical shape. Nimodipine was incorporated into the hydrophobic inner core of micelles as a lipophilic model drug to investigate the drug release behavior. The PEO/PCL ratio of copolymer was a main factor in controlling micelle size, drug-loading content, and drug release behavior. As PCL weight ratio increased, the micelle size and drug-loading content increased, and the drug release rate decreased. PMID- 12115847 TI - Surface adsorption of recombinant human interferon-gamma in lyophilized and spray lyophilized formulations. AB - Recombinant human interferon-gamma (rhIFN-gamma) was lyophilized or spray lyophilized in 9.5% trehalose, +/- 0.12% polysorbate 20 in 10 mM potassium phosphate, pH 7.5. We measured recovery of soluble protein after spraying, freeze thawing, and drying and reconstitution. Infrared spectroscopy showed rhIFN-gamma secondary structure to be native-like in all dried powders. Powders were characterized using electron spectroscopy for chemical analysis, time-of-flight secondary ion mass spectroscopy, X-ray diffraction, and gas adsorption isotherms. rhIFN-gamma adsorbed at air/liquid interfaces during spraying, and to ice/liquid interfaces during lyophilization. The concentration of rhIFN-gamma at ice/liquid interfaces was approximately one-fourth that adsorbed at air/liquid interfaces. Addition of 0.12% polysorbate 20 reduced the concentration of rhIFN-gamma at both interfaces. Time-of-flight secondary ion mass spectroscopy detected polysorbate 20 on surfaces of lyophilized powders. Lyophilized samples dried more slowly but reconstituted more quickly than spray-lyophilized samples. rhIFN-gamma aggregated after nebulization, but aggregation decreased in 0.12% polysorbate 20. Addition of 0.12% polysorbate 20 reduced protein surface adsorption and decreased but did not completely prevent aggregation. Insignificant aggregation occurred after exposure to ice/liquid interfaces, but subsequent drying and reconstitution caused aggregation. The majority of the aggregation is due to adsorption at air liquid and solid-air interfaces formed during spray-lyophilization or lyophilization. PMID- 12115848 TI - Doxorubicin pharmacokinetics: Macromolecule binding, metabolism, and excretion in the context of a physiologic model. AB - The studies described herein were designed to determine whether doxorubicin (DOX) pharmacokinetics (PKs) could be described by a physiologically based PK model that incorporated macromolecule-specific binding and organ-specific metabolism and excretion. Model parameters were determined experimentally, or were gathered from the literature, in a species-specific manner, and were incorporated into a physiologically based description of DOX blood and tissue distribution for mice, dogs, and humans. The resulting model simulation data were compared with experimentally determined data using PK parameters calculated using compartmental or noncompartmental analysis to assess the predictability of the models. The resulting physiologically based PK model that was developed could accurately predict blood and tissue PKs of DOX in mice. When this model was interspecies extrapolated to predict DOX levels in dogs and humans undergoing treatment for cancer, predictions in dog plasma or human serum were also consistent with the actual clinical data. This model has potential utility for predicting the magnitude of PK interactions of DOX with other drugs, and for predicting changes in DOX PKs in any number of clinical situations. PMID- 12115849 TI - Hydrophobic ion pairing of isoniazid using a prodrug approach. AB - Inhalation therapy for infectious lung diseases, such as tuberculosis, is currently being explored, with microspheres being used to target alveolar macrophages. One method of drug encapsulation into polymeric microspheres to form hydrophobic ion-paired (HIP) complexes, and then coprecipitate the complex and polymer using supercritical fluid methodology. For the potent antituberculosis drug, isoniazid (isonicotinic acid hydrazide, INH), to be used in this fashion, it was modified into an ionizable form suitable for HIP. The charged prodrug, sodium isoniazid methanesulfonate (Na-INHMS), was then ion paired with hydrophobic cations, such as alkyltrimethylammonium or tetraalkylammonium. The logarithms of the apparent partition coefficients (log P') of various HIP complexes of INHMS display a roughly linear relationship with the numbers of carbon atoms in the organic counterions. The water solubility of the tetraheptylammonium-INHMS complex is about 220-fold lower than that of Na-INHMS, while the solubility in dichloromethane exceeds 10 mg/mL, which is sufficient for microencapsulation of the drug into poly(lactide) microspheres. The actual logarithm of the dichloromethane/water partition coefficient (log P) for tetraheptylammonium-INHMS is 1.55, compared to a value of - 1.8 for the sodium salt of INHMS. The dissolution kinetics of the tetraheptylammonium-INHMS complex in 0.9% aqueous solutions of NaCl was also investigated. Dissolution of tetraheptylammonium-INHMS exhibited a first-order time constant of about 0.28 min(-1), followed by a slower reverse ion exchange process to form Na-INHMS. The half-life of this HIP complex is on the order of 30 min, making the enhanced transport of the drug across biological barriers possible. This work represents the first use of a prodrug approach to introduce functionality that would allow HIP complex formation for a neutral molecule. PMID- 12115850 TI - Preparation and characterization of a customized cellulose acetate butyrate dispersion for controlled drug delivery. AB - The purpose of the present experiment was to prepare and characterize the aqueous based pseudolatex system of cellulose acetate butyrate (CAB) for controlled drug delivery. Aqueous pseudolatex systems are advantageous over organic-based coating systems because these systems are devoid of criteria pollutants such as carbon monoxide, nitrogen oxides, nonmethane volatile organic compounds, and sulfur dioxide. Pseudolatex was prepared with CAB and polyvinyl alcohol (stabilizer) by a polymer emulsification technique. The stability of pseudolatex was evaluated. Particle size was measured and rheological experiments were conducted. The glass transition temperature, microscopic free volume, permeation coefficient, and mechanical properties of plasticized pseudolatex films were estimated. Surface roughness of coating on inert Nu-Pareil beads (Ingredient Technology Corp., Mahwah, NJ) was measured as a function of coating weight gain. The CAB Pseudolatex was found to be stabilized by steric forces. From intrinsic viscosity, the thickness of the stabilization layer was estimated. An increase in polymeric particles proportionately decreased the thickness of the stabilization layer. All the essential properties of a coating membrane such as microscopic free-volume fraction, permeability coefficient, mechanical properties, and glass transition temperature were fairly controllable as a function of plasticizer concentration. The pseudolatex dispersion of CAB was stable with negligible sedimentation volume and a particle size of 300 nm. Because CAB is water insoluble and non-ionizable, this pseudolatex can be used for pH-independent coating. The films obtained were strong and flexible for controlled drug delivery applications. Coating with the CAB dispersion reduced the surface roughness of beads but it remained stable as a function of increase in coating weight gain. PMID- 12115851 TI - Influx and efflux of amphetamine and N-acetylamphetamine in keratinocytes, pigmented melanocytes, and nonpigmented melanocytes. AB - To establish an in vitro model of drug incorporation into hair and to elucidate the potential roles of hair cell selectivity and hair color in the incorporation of certain drugs into hair, the basic drug amphetamine and its nonbasic analog N acetylamphetamine (N-AcAp) were analyzed for influx and efflux into and out of keratinocytes, pigmented melanocytes (PM), and nonpigmented melanocytes (NPM) as a model for incorporation and efflux of these drugs from hair cells. NPM were of the same melan-a cell line as PM, but cultured in the presence of the tyrosinase inhibitor phenylthiocarbamide. Results show that PM take up large amounts of the basic drug amphetamine (levels of uptake dependent on melanin content), whereas keratinocytes and NPM take up only small amounts of amphetamine. None of the cells take up N-AcAp above background levels. Interestingly, whereas keratinocytes and NPM quickly efflux most of the influxed drug, PM are slow to efflux and only efflux approximately 65% of influxed drug, if efflux media is not refreshed. (If efflux media is periodically refreshed, PM will eventually redistribute essentially all influxed drug back into the media.) These results demonstrate that pigmented cells take up greater amounts of the basic drug amphetamine, and efflux it more slowly than nonpigmented cells. Also, these results are consistent with previous data for in vivo incorporation of amphetamine in animal hair. In combination with previous data, an overall comparison of the amphetamine and N-AcAp incorporation data support a non diffusion mediated model for drug incorporation into hair cells. PMID- 12115852 TI - Utility of nuclear magnetic resonance spectroscopy to characterize the structure of dexamethasone sodium phosphate inclusion complexes with cyclodextrins in solution and to analyze potential competitive effects. AB - The interaction between dexamethasone sodium phosphate (DSP) and four cyclodextrin (CyD) derivatives [2,6-di-O-beta-cyclodextrin (DIMEB), gamma cyclodextrin (gamma-CyD), and hydroxypropyl-beta-cyclodextrin with either 2.7 or 4.6 degrees of substitution (HPbetaCyD 2.7 and HPbetaCyD 4.6, respectively)] was investigated by proton nuclear magnetic resonance spectroscopy (1H NMR). The data suggested the formation of inclusion complexes in solution in which B and C rings of the molecule are located inside the cavity. Nevertheless, the structure, in terms of depth within CyD, depends on the derivative considered. Molecular mechanics calculations of DSP complexes with DIMEB and gamma-CyD support the NMR results. The potential displacement of DSP from the CyD cavity by usual ophthalmic drugs (e.g., polymyxin B, trimethoprim, and benzalkonium chloride) was determined by NMR. The technique has been found useful to analyze this problem in pharmaceutical preparations. PMID- 12115853 TI - Molecular mobility study of amorphous and crystalline phases of a pharmaceutical product by thermally stimulated current spectrometry. AB - Two crystalline forms and the amorphous state of irbesartan, a pharmaceutical drug chosen as a model, were analyzed by Thermally Stimulated Current (TSC) spectroscopy, a powerful technique currently used in polymer science to investigate the molecular dynamics of heterogeneous and complex materials. Whereas no specific dielectric response was noted for the B crystalline form, the A form of irbesartan exhibited molecular motions localized inside its channel structure. The dynamics involved in the dielectric glass transition of amorphous samples followed a compensation law characteristic of highly cooperative relaxation processes. Concerning the amorphous content in physical mixtures, a calibration curve and a limit of detection (2.5%) were established. The limit of detection could be improved by optimizing the TSC experimental parameters. The amorphous sample recrystallized at a single temperature was interpreted by the "idealized one-state model" defined here to describe systems composed of identical semicrystalline particles in which amorphous and crystalline phases are independent of each other (i.e., no chemical and physical interaction between the two phases). Therefore, the idealized one-state model may be simulated by a two state model, which is representative of the two-phase model. Other samples recrystallized through a complex annealing stage were explained by the classical one-state model in agreement with the three-phase model used to describe bulk semicrystalline systems. These results demonstrate that, as for polymers, the semicrystalline state of pharmaceutical drugs should not be considered as a single state but as a more complex system that can be described as an idealized one-state model or a one-state model depending on the applied thermal treatment. These results give a new view that should be taken into account in the development of amorphous pharmaceutical drugs and formulations. PMID- 12115854 TI - Transport and local metabolism of budesonide and fluticasone propionate in a human bronchial epithelial cell line (Calu-3). AB - Calu-3 cells grown on microporous filters at an air interface for 16-18 days were incubated with the glucocorticosteroid (GCS) budesonide (BUD). Apparent permeability (P(app)) of BUD across Calu-3 cell monolayers was determined. Amount of BUD transported across Calu-3 cells was clearly concentration dependent, and no polarised transport was detected. When cells were loaded with BUD by incubation with drug solution (30 microM) for 2 h, 9.35 +/- 0.53% of the initial dose of BUD, corresponding to 5.5 microg BUD/mg cell protein was retained intracellularly, and released over a time period of 10 h. Apical release of BUD was significantly higher than release to the basolateral side of the monolayer. In comparison, when Calu-3 cells were loaded with fluticasone propionate (FP) solution (0.8 microM), about 20% of the initial dose of FP, corresponding to 0.3 microg FP/mg total cell protein content was detected intracellularly and released immediately (45 min). There was no difference in FP release between the apical and basolateral side of the cell layer. Mass spectrometry of cell extracts indicated the presence of fatty acid conjugates of BUD. We conclude that Calu-3 cells are able to store BUD by intracellular conjugation to fatty acids. We, therefore, suggest the use of the Calu-3 cell model as a tool for examination of local pharmacokinetics and metabolism of GCS at the bronchial epithelium. PMID- 12115855 TI - Partition of N-monodemethylated phenothiazine drugs to phosphatidylcholine bilayer vesicles studied by second-derivative spectrophotometry. AB - The partition coefficients (Kps) of three N-monodemethylated metabolites of phenothiazines (chlorpromazine (CPZ), triflupromazine, and promazine) between phosphatidylcholine (PC) vesicles (SUV) and water were determined to evaluate their affinity to biomembranes by second-derivative spectrophotometry without any separation procedures. The second derivative spectra showed distinct derivative isosbestic points confirming the entire elimination of the residual background signal effects of the SUV, which were observed in the absorption spectra. From the relationship between PC SUV concentration and the derivative intensity change (DeltaD) of each metabolite induced by its interaction with the PC bilayer, the Kp values were calculated and could be obtained with the R.S.D. of below 10% (n = 5) proving an accuracy of the derivative method. The obtained Kp values were similar to those of the parent drugs (the relative differences were within 15%), although the partition coefficient of N-monodemethylated CPZ measured in an octanol/buffer system was reported to be about 1/18 of CPZ. The results obtained from the PC liposome/buffer system do not contradict with psychoactivity and brain accumulation of N-monodemethylated metabolites of phenothiazines, in contrast to the results of the octanol/buffer system. PMID- 12115858 TI - An introduction to stem cells. AB - 1998 saw the publication of two papers describing the growth in vitro of human embryonic stem (ES) cells derived either from the inner cell mass (ICM) of the early blastocyst or the primitive gonadal regions of early aborted fetuses. Work on murine ES cells over many years had already established the amazing flexibility of ES cells, essentially able to differentiate into almost all cells that arise from the three germ layers. The realization of such pluripotentiality (see below) has, of course, resulted in the field of stem cell research going into overdrive, the establishment of many new biotechnology companies (http://www.stemcellresearchnew.com/catalog1677.html), and a genuine belief that stem cell research will deliver a revolution in terms of how we treat cardiovascular disease, neurodegenerative disease, cancer, diabetes, and the like. However, many people believe that early human embryos should be accorded the same status as any sentient being and thus their 'harvesting' for stem cells is morally unjustifiable. With this in mind, other sources of malleable stem cells have been sought. In the adult, organ formation and regeneration was thought to occur through the action of organ- or tissue-restricted stem cells (i.e. haematopoietic stem cells giving rise to all the cells of the blood, neural stem cells making neurons, astrocytes, and oligodendrocytes). However, it is now believed that stem cells from one organ system, for example the haematopoietic compartment can develop into the differentiated cells within another organ system, such as the liver, brain or kidney. Thus, certain adult stem cells may turn out be as malleable as ES cells and so also be useful in regenerative medicine. This brief overview summarizes the important attributes of tissue-based stem cells and clarifies the terms used. PMID- 12115859 TI - Embryonic stem cells. AB - The capacity of embryonic stem cells for virtually unlimited self-renewal and differentiation capacity has opened up the prospect of widespread applications in biomedical research and regenerative medicine. For the latter, the cells provide hope that it will be possible to overcome the problems of donor tissue shortage and also, by making the cells immunocompatible with the recipient, implant rejection. Four years after the first derivation of human pluripotent cell lines from pre-implantation embryos, a great deal has been learnt about their biology and how differentiation can be encouraged towards particular cell lineages. However, considerable research is needed, not least into means to enrich and purify derivative cell lineages, before clinical trials can be considered. PMID- 12115860 TI - Haematopoietic stem cells. AB - Considerable efforts have been made in recent years in determining the composition of the cell types that constitute the human haematopoietic stem cell (HSC) compartment. These studies have emphasized the heterogeneity of the human HSC in terms of proliferative and self-renewal capacities. Recent studies have indicated that CD34 is not the universal marker of all human HSCs. New markers for purifying HSCs have been described. A number of genes that regulate the formation, self-renewal, or differentiation of HSCs has been identified. The elucidation of the molecular phenotype of the HSC has just begun. Finally, an unexpected degree of developmental or differentiation plasticity of HSC has emerged. This review summarizes all the recent advances made in the human HSC field and examines the impacts that these discoveries may have both clinically and in understanding the organization of the human haematopoietic system. PMID- 12115861 TI - Adult stem cell plasticity. AB - Observations made in the last few years support the existence of pathways, in adult humans and rodents, that allow adult stem cells to be surprisingly flexible in their differentiation repertoires. Termed plasticity, this property allows adult stem cells, assumed, until now, to be committed to generating a fixed range of progeny, to switch, when they have been relocated, to make other specialized sets of cells appropriate to their new niche. Reprogramming of some adult stem cells can occur in vivo; the stem cells normally resident in bone marrow appear particularly flexible and are able to contribute usefully to multiple recipient organs. This process produces cells with specialized structural and metabolic adaptations commensurate with their new locations. In a few examples, the degree of support is sufficient to assist or even rescue recipient mice from genetic defects. Some studies provide evidence for the expansion of the reprogrammed cells locally, but in most it remains possible that cells arrive and redifferentiate, but are no longer stem cells. Nevertheless, the fact that appropriately differentiated cells are delivered deep within organs simply by injection of bone marrow cells should make us think differently about the way that organs regenerate and repair. Migratory pathways for stem cells in adult organisms may exist that could be exploited to effect repairs using an individual's own stem cells, perhaps after gene therapy. Logical extensions of this concept are that a transplanted organ would become affected by the genetic susceptibilities of the recipient, alleles that re-express themselves via marrow derived stem cells, and that plasticity after bone marrow transplantation would also transfer different phenotypes, affecting important parameters such as susceptibility to long-term complications of diabetes, or the ability to metabolize drugs in the liver. This article reviews some of the evidence for stem cell plasticity in rodents and man. PMID- 12115862 TI - Muscle stem cells. AB - Since its discovery four decades ago, the satellite cell of skeletal muscle has been implicated as the major source of myogenic cells involved in growth and repair of muscle fibres. This review not only looks at the role of the satellite cell in these processes but discusses how cells derived from other sources and tissues have recently been implicated in muscle formation and regeneration. Muscle itself also yields cells that contribute to other cell lineages although it is currently debated as to whether these cells originate within muscle or have migrated there from other tissues. The reality of using cells from muscle or other tissues to repair diseased muscle fibres is also addressed. PMID- 12115863 TI - Cardiac stem cells. AB - Augmentation of myocardial performance in experimental models of therapeutic infarction and heart failure has been achieved by the transplantation of exogenous cells into damaged myocardium, a procedure known as cellular cardiomyoplasty (CCM). Historically, a wide range of cell types have been used for CCM, including rat and human fetal ventricular myocytes, but the availability of human fetal donor cells for clinical purposes is limited. The quest for suitable alternative donor cells has prompted research into the use of both embryonic stem (ES) cells and adult somatic stem cells, but the optimal choice of donor cell source is not yet known. Recently, there has been a growing body of evidence that multipotent somatic stem cells in adult bone marrow exhibit tremendous functional plasticity and can reprogramme in a new environmental tissue niche to give rise to cell lineages specific for the new organ site. This phenomenon has made a huge impact on myocardial biology and has captured the imagination of scientists who have recently discovered that multipotent adult bone marrow haematopoeitic stem cells and mesenchymal stem cells can repopulate infarcted rodent myocardium and differentiate into both cardiomyocytes and new blood vessels. These data, coupled with the identification of a putative primitive cardiac stem cell population in the adult human heart, may pave the way for novel therapeutic modalities for enhancing myocardial performance and treating end-stage cardiac disease. PMID- 12115864 TI - Epidermal stem cells. AB - The clinical implications of understanding epidermal stem cell biology abound. Thousands of burns victims across the world have benefited from early research into the proliferation of epidermal keratinocytes in vitro. Advances now indicate there are a number of stem cell repositories within the epidermis, two of which, the interfollicular epidermis and the bulge region of the hair follicle, may supply each other when damaged. This review details the progress made in the identification and characterisation of stem cells within the epidermis and discusses the molecules involved in the epidermal stem cell's choice of fate. Finally, the skin, like bone marrow, could be a readily accessible source of stem cells for therapeutic intervention and evidence of skin stem cell plasticity is highlighted. PMID- 12115865 TI - Gastrointestinal stem cells. AB - Turnover of the epithelial cell lineages within the gastrointestinal tract is a constant process, occurring every 2-7 days under normal homeostasis and increasing after damage. This process is regulated by multipotent stem cells, which give rise to all gastrointestinal epithelial cell lineages and can regenerate whole intestinal crypts and gastric glands. The stem cells of the gastrointestinal tract are as yet undefined, although it is generally agreed that they are located within a 'niche' in the intestinal crypts and gastric glands. Studies of allophenic tetraparental chimeric mice and targeted stem cell mutations suggest that a single stem cell undergoes asymmetrical division to produce an identical daughter cell, and thus replicate itself, and a committed progenitor cell which further differentiates into an adult epithelial cell type. The discovery of stem cell plasticity in many tissues, including the ability of transplanted bone marrow to transdifferentiate into intestinal subepithelial myofibroblasts, provides a potential use of bone marrow cells to deliver therapeutic genes to damaged tissues, for example, in treatment of mesenchymal diseases in the gastrointestinal tract, such as fibrosis and Crohn's disease. Studies are beginning to identify the molecular pathways that regulate stem cell proliferation and differentiation into adult gastrointestinal cell lineages, such as the Wnt and Notch/Delta signalling pathways, and the importance of mesenchymal epithelial interactions in normal gastrointestinal epithelium and in development and disease. PMID- 12115866 TI - Hepatic stem cells. AB - The liver in an adult healthy body maintains a balance between cell gain and cell loss. Though normally proliferatively quiescent, hepatocyte loss such as that caused by partial hepatectomy, uncomplicated by virus infection or inflammation, invokes a rapid regenerative response to restore liver mass. This restoration of moderate cell loss and 'wear and tear' renewal is largely achieved by hepatocyte self-replication. Furthermore, hepatocyte transplants in animals have shown that a certain proportion of hepatocytes can undergo significant clonal expansion, suggesting that hepatocytes themselves are the functional stem cells of the liver. More severe liver injury can activate a potential stem cell compartment located within the intrahepatic biliary tree, giving rise to cords of bipotential so-called oval cells within the lobules that can differentiate into hepatocytes and biliary epithelial cells. A third population of stem cells with hepatic potential resides in the bone marrow; these haematopoietic stem cells can contribute to the albeit low renewal rate of hepatocytes, make a more significant contribution to regeneration, and even completely restore normal function in a murine model of hereditary tyrosinaemia. How these three stem cell populations integrate together to achieve a homeostatic balance is not known. This review focuses on two major aspects of liver stem cell biology: firstly, the identity of the liver stem cells, and secondly, their potential value in the treatment of major liver disease. PMID- 12115867 TI - Pancreatic stem cells. AB - beta-cell replacement therapy via islet transplantation has had renewed interest, due to the recent improved success. In order to make such a therapy available to more than a few of the thousands of patients with diabetes, new sources of insulin-producing cells must be readily available. The recent conceptual revolution of the presence of adult pluripotent stem cells in bone marrow and in most, if not all, organs suggests that adult stem cells may be a potential source of insulin-producing cells. Pancreatic stem/progenitor cells or markers for these cells have been sought in both islets and ducts. There is considerable evidence that such cells exist and several candidate cells have been reported. However, no clearly identifiable adult pancreatic stem cell has been found as yet. The putative pancreatic stem cells will be the focus of this review. PMID- 12115868 TI - Lung epithelial stem cells. AB - This review concentrates on recent evidence about lung stem cell origins and plasticity. The range of potential cells which can repopulate the injured lung, classically the basal and mucous secretory cells of the trachea, the Clara cells of the bronchiole, and the type II pneumocyte of the alveolus, has been extended to include the mucus-gland duct cells of the trachea and bronchus. Some evidence suggests that there are variant Clara cells that lack cytochrome P-450 and so are spared toxic activation of xenobiotics, and may aid bronchiolar repopulation after injury, such as with naphthalene. There may even be involvement of the neuroepithelial bodies or cells in this, though the evidence is not yet conclusive. The search for a resident pulmonary multipotent cell for repopulating any lung epithelium has not yet been successful. The picture remains similar to earlier conclusions, in that the local stem or precursor cell is the most likely to contribute to local needs in times of tissue damage. There remains a major challenge for lung cancer treatment, where high-dose chemo- or radio-therapy may be hoped to promote the seeding and repair of lung parenchyma by circulating bone marrow stem cells, as seen in liver models. Patient survival rates do not yet suggest that this occurs to any great extent, but this remains to be shown formally. The effects of prior fibrosis and tumour necrosis are probably confounding factors in this lack of rescue. PMID- 12115869 TI - Neural stem cells. AB - Neural stem cells (NSCs) have the ability to self-renew, and are capable of differentiating into neurones, astrocytes and oligodendrocytes. Such cells have been isolated from the developing brain and more recently from the adult central nervous system. This review aims to provide an overview of the current research in this evolving area. There is now increasing knowledge of the factors controlling the division and differentiation of NSCs during normal brain development. In addition, the cues for differentiation in vitro, and the possibility of transdifferentiation are reviewed. The discovery of these cells in the adult brain has encouraged research into their role during neurogenesis in the normal mature brain and after injury. Lastly other sources of neural precursors are discussed, and the potential for stem cells to be used in cell replacement therapy for brain injury or degenerative brain diseases with a particular emphasis on cerebral ischaemia and Parkinson's disease. PMID- 12115870 TI - Prostatic stem cells. AB - Multipotent cells within the epithelial compartment, together with phenotypically 'plastic' mesenchyma cells (stromal stem cells), provide a repository of protected genetic information from which the structure, stability and functionality of the prostate gland can be maintained. However, mere preservation of cells in a non-dividing state is insufficient to provide the necessary reservoir of information from which the structure and function of the prostate gland can be retained or recreated. Rather, there is a constant dynamic interaction, at the level of information exchange, between stem cells (whether epithelial or mesenchymal) and their surrounding environment (both humoral and physical). Thus, with respect to epithelial stem cells, these reside within environmental 'niches' which allow their controlled and limited proliferation while preserving genomic integrity. Similar 'mesenchymal niches' are also predicted to occur, although not yet identified, thus providing the multipotent source from which the full spectrum of stromal phenotypes might be regenerated. Recent data from studies of the haematopoietic and hepato-biliary systems indicate that the potential scope of stem cells far exceeds the immediate phenotypic complement of those tissues within which they originate, being dependent upon their precise environment as well as their genomic integrity. PMID- 12115871 TI - Development and pathology: the Pax gene. AB - During development, many genes support different functions at different times, when they are expressed in different tissues, or in response to different transcription factors. Pathologists should be cautious about ascribing functions to such genes in pathological processes. PMID- 12115872 TI - Mast cells in the pathogenesis of chronic back pain: a hypothesis. AB - The pathophysiology of chronic low back pain is poorly understood, mainly because it is difficult to study experimentally or objectively. Recently it has been found that there is a relationship between neovascularization and innervation of the usually avascular and aneural intervertebral disc at the sites of discogenic pain. These data, together with the recognized involvement of mast cells in tissue repair, in the induction of angiogenesis, and in the production of and response to neurotrophic stimuli such as nerve growth factor, has suggested the hypothesis that mast cells may have a causative role in chronic low back pain. If so, the mast cell may represent an attractive therapeutic target. PMID- 12115873 TI - Nerve growth factor expression and innervation of the painful intervertebral disc. AB - Following a previous description of nociceptive nerve fibre growth into usually aneural inner parts of painful intervertebral disc (IVD), this study has investigated whether nociceptive nerve ingrowth into painful IVD is stimulated by local production of neurotrophins. Immunohistochemistry and in situ hybridization have been used to investigate expression of the candidate neurotrophin, nerve growth factor (NGF), and its high- and low-affinity receptors trk-A and p75, respectively, in painful IVD excised for the management of low back pain. IVD from patients with back pain were of two types: those that when examined by discography reproduced the patient symptoms (pain level IVD) and those that did not (non-pain level IVD). Microvascular blood vessels accompanied nerve fibres growing into pain level IVD and these expressed NGF. The adjacent nerves expressed the high-affinity NGF receptor trk-A. These vessels entered the normally avascular IVD through the discal end plates. NGF expression was not identified in non-pain level or control IVD. Some non-pain level IVD had vessels within them, which entered through the annulus fibrosus. These did not express NGF nor did nerves accompany them. These findings show that nociceptive nerve ingrowth into painful IVD is causally linked with NGF production by blood vessels growing into the IVD, from adjacent vertebral bodies. PMID- 12115874 TI - Progressive loss of PAX9 expression correlates with increasing malignancy of dysplastic and cancerous epithelium of the human oesophagus. AB - Pax genes encode a family of transcription factors that play key roles in embryonic development. Whereas the functions of Pax genes in the adult organism are largely unknown, upregulated Pax gene expression has been implicated in tumourigenesis. In this study, PAX9-specific monoclonal antibodies have been generated and it has been shown that PAX9 protein is expressed in the normal epithelium of the adult human oesophagus. PAX9 expression was either lost or significantly reduced in the majority of invasive carcinomas and epithelial dysplasias, the latter representing precancerous lesions. Notably, the percentage of PAX9-positive cells within the epithelium decreased with increasing malignancy of the epithelial lesion. These results identify PAX9 as a sensitive marker for deregulated differentiation of oesophageal keratinocytes and indicate a role for PAX9 in the normal differentiation process of internal stratified squamous epithelia. These data suggest that upregulated PAX9 expression is not required for the formation of the majority of squamous cell carcinomas of the human oesophagus. PMID- 12115875 TI - Loss of heterozygosity in the clonal evolution of flat colorectal neoplasms. AB - In contrast to invasive colorectal carcinomas that develop in typical exophytic adenoma-carcinoma sequences, some invasive cancers may evolve from flat mucosal dysplastic lesions. Despite their relatively small size, these flat colorectal lesions are often associated with high-grade dysplasia and may show an aggressive clinical course. To delineate the genetic pathways in the clonal evolution of these tumors, multiple foci were microdissected from 13 cases and the allelic deletions of 15 chromosomal arms were analysed. Loss of heterozygosity (LOH) was detected most frequently on 17p (77%), followed by 18q (69%), and 5q (54%). In five cases with concomitant low-grade adenomas, only one case showed LOH in low grade adenoma foci. In high-grade dysplasia with/without submucosal invasion, early and homogeneous LOH of one to several chromosomal arms was detected. Overall, homogeneous and thus early LOH were most frequently detected on 17p (seven of 10 cases with 17p LOH), followed by 3p (two of three cases with 3p LOH), and 5q (four of seven cases with 5q LOH). In addition to homogeneous LOH, the LOH patterns observed in different portions of dysplasias and invasive cancers in individual cases identified several different genetic patterns of tumour progression, either with linear or branching (divergent) trees. Positive immunostaining for p53 was detected in 10 of the 13 cases; of these, five cases were concomitant with 17p LOH in all of the microdissected foci, four cases were concomitant with 17p LOH in a majority of foci and, one case showed retention of 17p. Except for the flat configuration and early 17p LOH, genetic heterogeneity in the flat high-grade dysplastic foci was found to be similar to genetic chaos in the late dysplastic and preinvasive stages of exophytic adenoma. These findings suggest a potentially aggressive course for these neoplasms. PMID- 12115876 TI - Enhanced mRNA expression of tissue inhibitor of metalloproteinase-1 (TIMP-1) in breast carcinomas is correlated with adverse prognosis. AB - Tissue inhibitor of metalloproteinase-1 (TIMP-1) has emerged as a multifunctional protein with the contrasting activities of inhibiting tissue-degrading enzymes and promoting cellular growth. In an attempt to elucidate the clinical significance of TIMP-1 in breast cancer, the expression of TIMP-1 mRNA was evaluated in 117 invasive breast carcinomas by mRNA in situ hybridization, in correlation with clinicopathological parameters, immunohistochemical prognostic factors (Ki-67, c-erb-B-2, bcl-2) and clinical outcome. TIMP-1 was detected in stromal cells in areas within the tumours and at the tumour margin. High TIMP-1 mRNA expression in the marginal portion of the tumours was significantly correlated with lymph node metastasis (p<0.05) and c-erbB-2 expression (p<0.05). On the other hand, increased TIMP-1 mRNA expression within the tumours showed a statistically significant correlation with ER detection (p<0.01). Multivariate analysis revealed worse survival for patients with high TIMP-1 mRNA expression in the marginal portion of the tumours; the subgroup of these patients co-expressing high levels of TIMP-1 mRNA within the tumours as well had even worse survival (p=0.042). In conclusion, our data support the multifunctional role of TIMP-1, particularly its growth-promoting activity, on the basis of its significant correlation with lymph node metastasis and adverse prognosis. In addition to the latter property, a probable association of TIMP-1 with tumour cell differentiation is suggested by its topographical correlation with ER detection. PMID- 12115877 TI - Expression of carbonic anhydrase IX in breast is associated with malignant tissues and is related to overexpression of c-erbB2. AB - CA IX is a tumour-associated carbonic anhydrase with proposed roles in pH modulation and intercellular communication. Its distribution was examined in normal, benign and malignant breast tissues and compared with expression of breast tumour markers including oestrogen receptor, c-erbB2, c-erbB3 and CD44. Tissue specimens were analysed using immunohistochemistry and/or reverse transcriptase-polymerase chain reaction (RT-PCR). CA IX was detected by IHC in 12/26 (46%) malignant tissues, 4/36 (11%) benign lesions, but not in 10 normal breasts. Staining was mostly confined to plasma membranes of abnormal epithelial cells, but in five cases was found in adjacent stroma. Semi-quantitative RT-PCR detected CA9 mRNA in 25/39 (64%) malignant tumours, 11/33 (33%) benign lesions, but in none of three normal breasts. Comparative RT-PCR analysis of malignant tissues revealed a relationship between CA9 positivity and c-erbB2 overexpression (p=0.05). Moreover, CA9-positive specimens displayed a significantly higher median level of c-erbB2 than CA9-negative ones (p=0.02). No significant association was found with the other markers. The results of this study support the possible importance of CA IX for breast carcinogenesis and suggest its potential use as a breast tumour marker. PMID- 12115878 TI - Evidence for the early involvement of interleukin 17 in human and experimental renal allograft rejection. AB - Inflammatory processes can stimulate renal epithelial cells to release cytokines, chemoattractants and matrix proteins into the interstitium, thus contributing to interstitial injury during acute allograft rejection. To test the role of interleukin 17 (IL-17) in this process, cultured human renal epithelial cells (hRECs) were first established and treated with or without human IL-17 (hIL-17) for 2, 4, 8 and 10 h in vitro. Significant elevations of IL-6 and IL-8 levels were noted in the supernatants in a dose-dependent and time-dependent manner, as also for IL-6 mRNA expression. Secondly, using a rat acute allograft rejection model, the correlation between IL-17 expression and histopathological changes was serially studied. The results demonstrated that increased expression of IL-17 protein on infiltrating mononuclear cells (MNCs) was detectable on day 2. This corresponds to the borderline change of acute rejection according to the Banff classification, and it increased progressively to day 5. Serial study of IL-6, IL 8 and IL-17 mRNA expression of the renal allograft confirmed IL-17 mRNA expression in the allograft early on post-transplant day 2, whereas IL-6 and IL-8 expression started on day 3. Thirdly, IL-17 expression was observed in human renal allograft and urinary sediment. IL-17 protein expression was found in human subclinical (borderline) rejection renal allograft biopsy tissue and none in biopsy tissue not showing any evidence of rejection. There was also a 100% detectable rate of IL-17 mRNA expression in the MNCs of urinary sediment of patients with subclinical borderline rejection. These results demonstrate that hRECs exposed to IL-17 can produce inflammatory mediators with the potential to stimulate early alloimmune responses, which may also serve to give warning of acute renal allograft rejection. PMID- 12115879 TI - Extranodal marginal zone B cell lymphomas of the uvea: an analysis of 13 cases. AB - The majority of primary lymphoproliferative lesions of the uvea represent low grade B cell lymphomas and often display a prominent plasmacellular differentiation. The purpose of the current study was to classify the uveal lymphoproliferations according to the REAL classification; examine the immune profile of the plasmacellular differentiated tumour cells using the plasma cell related antigens multiple myeloma oncogene-1-protein (MUM1), Vs38c, CD38 and CD138; and to compare this profile with that of mature reactive plasma cells. Following fixation, 13 lymphoproliferative lesions of the uvea were categorized on the basis of their morphology and immunophenotype according to the REAL classification. Included in the immunohistochemistry were B cell-specific activator protein (BSAP), MUM1, Vs38c, CD38 and CD138. Nested polymerase chain reaction (PCR) was also performed on DNA extracted from paraffin sections for the detection of gene rearrangements of the heavy immunoglobulin chain (IgH). All of the 13 uveal tract lymphoproliferative lesions represented malignant lymphoma of B cell non-Hodgkin type and could be diagnosed as 'extranodal marginal zone B cell lymphomas' (EMZL). The degree of plasmacellular differentiation varied between the tumours. In contrast to their non-plasmacytoid counterparts, the 'plasmacytoid' EMZL tumour cells were negative for the B cell markers CD20 and BSAP, and demonstrated heterogeneous positivity for the markers MUM1, Vs38c, CD38 and CD138. The most consistent marker was MUM1, being observed in all tumours. Co expression of all plasma cell markers was observed in four (31%) uveal EMZL. Loss of CD138 expression was observed in six (46%) tumours, of Vs38c expression in five (38%) and of CD38 in one (7%) tumour. Although the diagnosis of malignant lymphoma was unequivocally based on morphological and immunophenotypical features, the molecular analysis was able to demonstrate clonal B cell populations in only one uveal EMZL. All uveal lymphoid proliferations investigated represented EMZL, with the corresponding morphology and immunophenotype as seen in EMZL in other extranodal locations. MUM1, followed by CD38 expression, were the most constant plasma cell antigens in the plasmacytoid EMZL tumour cells, with both Vs38c and CD138 positivity being lost in many tumours. Aberrant immune profiles of plasma cell-related antigens may be of help in the establishment of malignancy in uveal lymphoproliferative lesions, particularly where interpretation of light chain expression and/or PCR results is difficult. PMID- 12115880 TI - Immunohistochemical detection of CD79a expression in precursor T cell lymphoblastic lymphoma/leukaemias. AB - Twenty-three cases of precursor T cell lymphoid malignancies were examined with respect to CD79a expression. Immunohistochemical staining was performed on frozen tissue sections using a broad panel of antibodies and Southern blot analysis was undertaken using DNA probes encoding immunoglobulin heavy chain (IgH) gene and T cell receptor (TCR) beta, gamma and delta genes. Twelve (52%) of the 23 cases examined demonstrated CD79a expression. IgH and TCRbeta, gamma and delta gene rearrangements were found in 5, 9, 12 and 20 cases, respectively. CD79a-positive neoplastic cells exhibited a phenotype and genotype characteristic of an early stage of T cell differentiation. Immunohistochemical staining was also performed on human thymus and thymoma to investigate the normality of CD79a expression, to discover that low-level expression of CD79a is common in thymocytes and thymoma associated lymphocytes. These results suggest that CD79a is expressed weakly and transiently in immature T-lineage cells. PMID- 12115881 TI - Clonal T cell receptor gamma-chain gene rearrangement by PCR-based GeneScan analysis in the skin and blood of patients with parapsoriasis and early-stage mycosis fungoides. AB - Cutaneous T cell lymphoma (CTCL) and reactive T cell skin diseases represent opposite ends of a spectrum of diseases ranging from overtly malignant to persistently benign. Within this spectrum, the parapsoriasis group is not clearly defined regarding malignant potential. In contrast to consistent findings in advanced-stage CTCL, clonality analysis of parapsoriasis has produced conflicting results in previous studies. As T cell receptor gamma-chain polymerase chain reaction GeneScan analysis (TCR-gamma-PCR-GSA) stands out by its sensitivity, its accuracy in size determination of PCR products, its capacity to identify false positives by repeated analysis and its easy applicability, this approach was used to analyse the clonality status of 41 patients with borderline T cell lymphoproliferative skin diseases, including parapsoriasis (n=27) and early-stage mycosis fungoides (MF) (n=14). A monoclonal T cell infiltrate was demonstrated by repeated TCR-gamma-PCR-GSA in lesional skin specimens in 19.2% of parapsoriasis patients and in 66.6% of early-stage MF cases (p=0.013). In peripheral blood, a monoclonal T cell population was found in a similar percentage of parapsoriasis and of early-stage MF patients (26.7% versus 12.5%; p=0.611). A detailed analysis of parapsoriasis subentities, namely small and large plaque parapsoriasis, and parapsoriasis lichenoides, revealed monoclonality in 2(6)/2(5), 3(14)/2(8) and 0(6)/0/(3) of the skin and peripheral blood specimens, respectively. The high detection rate of false positive cases by repeated analysis (20-37.5%) provides a corrected perspective for the high rates of dominant T cell clones found by others in the peripheral blood of such patients. From the results obtained, three major conclusions can be drawn: firstly, CTCL is clearly associated with detection of monoclonality, even in its early stages; secondly, monoclonality is not a prerequisite for potential CTCL precursor entities; and thirdly, recirculating malignant T cells identical to the skin clone are not readily detected in parapsoriasis or early-stage MF, but may rather indicate disease progression. PMID- 12115882 TI - Vascularization of cutaneous melanoma involves vessel co-option and has clinical significance. AB - This study was undertaken to determine the role and the fate of the peritumoural vascular plexus during the vascularization of human malignant melanoma (hMM) and in an appropriate murine melanoma model system. The prognostic significance of the vascularity of different tumour areas was also evaluated. Despite morphometry revealing several-fold higher microvessel densities (MVDs) in the peritumoural tissue than at the centre of the tumour, the development of visceral metastases of hMM was exclusively correlated with the MVD of the tumour centre. Furthermore, the 5-year survival of the patient group with low tumour centre MVD (<30/mm(2), n=29) was 100%, compared to 1/16 patients alive with high tumour centre MVD (>30/mm(2), n=16). Morphometric analysis and three-dimensional reconstruction of vessel networks of both human and murine melanomas showed clearly that the peritumoural vascular plexus present at the melanoma base is continuously being incorporated into the growing tumour mass. Once vessels become incorporated, sprouting ceases and the proliferating endothelial cells (EC) take part only in vessel dilatation. Moreover, the immunohistochemical and ultrastructural characterization of microvessels demonstrated that the pericyte coverage of endothelial tubes was complete in all of the investigated areas, in both human and murine melanomas. PMID- 12115883 TI - Molecular cytogenetic differences between histological subtypes of malignant mesotheliomas: DNA cytometry and comparative genomic hybridization of 90 cases. AB - It is established that subtypes of human malignant mesotheliomas (MM) are associated with different survival times. Ninety cases of MM were examined using DNA cytometry and comparative genomic hybridization (CGH), with emphasis on the main histological subtypes; epithelioid, sarcomatoid and biphasic. A comparison by DNA cytometry revealed moderate differences, with the rare subgroup of mesodermomas having the highest and the sarcomatoid group the lowest rate of aneuploidy. Using CGH, 6.2 chromosomal imbalances per case on average could be detected. Losses (4.1/case) were more common than gains of chromosomal material (2.1/case). MM show no single, specific defect, but a typical pattern of genomic defects can be attributed to this tumour entity. Common losses are clustered at the chromosomal regions 9p21 (34%), 22q (32%), 4q31-32 (29%), 4p12-13 (25%), 14q12-24 (23%), 1p21 (21%), 13q13-14 (19%), 3p21, 6q22, 10p13-pter and 17p12-pter (16% each). Common gains are located on 8q22-23 (18%), 1q23/1q32 (16%), 7p14-15 and 15q22-25 (14% each). While differences in the frequencies of the defects between epithelioid and sarcomatoid MM are not as pronounced as are seen with the pleomorphic mesodermomas, several chromosomal locations (3p, 7q, 15q, 17p) show significant variations. The most pronounced distinguishing feature of sarcomatoid MM is a more than fourfold higher number of amplicons. These data indicate that MM has a distinctive tumour biology with a broad spectrum of heterogeneity, as reflected in morphology and also, more subtly, in the patterns of chromosomal imbalances of the subtypes. PMID- 12115884 TI - iNOS gene upregulation is associated with the early proliferative response of human lung fibroblasts to cytokine stimulation. AB - Increased release of oxidants has been implicated in the pathogenesis of pulmonary fibrosis. Previous work in the rat showed that formation of the early fibrotic lesion is associated with increased expression of inducible nitric oxide synthase (iNOS) in pulmonary fibroblasts. The aim of this study was to test the hypothesis that NO is involved in the activation of pulmonary fibroblasts. The effects of endogenous and exogenous NO on proliferation of human pulmonary fibroblasts were investigated by administration of cytomix or SNAP, respectively. At low concentrations, both treatments increased cell numbers, an effect attenuated by iNOS inhibitor or NO scavenger. Induction of iNOS was confirmed by measurement of nitrate/nitrite production and by immunodetection. Quantitative RT PCR showed an increase in iNOS mRNA as early as 3 h after stimulation. These results support the hypothesis and show that upregulation of the iNOS gene is an early event in the proliferative response of human lung fibroblasts to inflammatory stimuli. PMID- 12115885 TI - Changes in myosin heavy chain and its localization in rat heart in association with hypobaric hypoxia-induced pulmonary hypertension. AB - Experimental pulmonary hypertension induced in a hypobaric hypoxic environment (HHE) is characterized by structural remodelling of the heart. In rat cardiac ventricles, pressure and volume overload are well known to be associated with changes in cardiac myosin heavy chain (MHC) isoforms. To study the effects of HHE on the MHC profile in the ventricles, 83 male Wistar rats were housed in a chamber at the equivalent of 5500 m altitude for 1-8 weeks. Pulmonary arterial pressure, right ventricular free wall (RVFW) weight, the ratio of RVFW weight over body weight (BW), the ratio of left ventricular free wall (LVFW) weight over BW, and myocyte diameter in both ventricles showed significant increases after 1 week, 2 weeks, 1 week, 6 weeks, and 4 weeks of HHE, respectively. Semi quantitative reverse transcriptase-polymerase chain reaction revealed that beta MHC mRNA expression was increased significantly in both ventricles at 6 and 8 weeks of HHE, whereas alpha-MHC mRNA expression was decreased significantly at 6 and 8 weeks of HHE in the right ventricle (RV) and at 6 weeks of HHE in the left ventricle (LV). The percentage of myosin containing the beta-MHC isoform was increased significantly at 4-8 weeks of HHE in RV and at 6 weeks of HHE in LV. In situ hybridization showed that the area of strong staining for beta-MHC mRNA was increased in both ventricles at 8 weeks of HHE, and showed a decrease from RVFW to cardiac septum, and from cardiac septum to LVFW. These results suggest that HHE has a significant effect on the expression of both MHC mRNA and protein in the heart, particularly in RV. These changes may reflect a role for cardiac MHC in the response to pulmonary hypertension in HHE. PMID- 12115886 TI - Tenascin-C is a useful marker for disease activity in myocarditis. AB - Tenascin-C (TNC) is an extracellular matrix protein which appears at active sites of tissue remodelling during embryogenesis or cancer invasion. In normal heart, TNC is only present during the early stages of development but reappears in pathological states. This study examined the diagnostic value of TNC for assessing disease activity of myocarditis. Expression of TNC was examined in myosin-induced autoimmune myocarditis mouse models. Sequential changes in amount, localization and the producing cells were analysed by reverse transcriptase polymerase chain reaction, western blotting, immunohistochemistry and in situ hybridization and compared with the histological picture. The expression of TNC was upregulated at a very early stage of myocarditis. Immunostaining was detectable before cell infiltration and myocytolysis became histologically apparent, remained during the active stage while cell infiltration and necrosis continued, and disappeared in scar tissue with healing. TNC immunostaining was always observed at the periphery of necrotic or degenerating cardiomyocytes in foci of inflammation, the expression level correlating with histological evidence of inflammatory activity. Interstitial fibroblasts were the major source of TNC, expressing the large isoform containing alternative splicing sites. These data demonstrate that TNC is a useful marker for evaluation of disease activity in myocarditis. PMID- 12115887 TI - Herpesviruses in brain and Alzheimer's disease. AB - It has been established, using polymerase chain reaction (PCR), that herpes simplex virus type 1 (HSV1) is present in a high proportion of brains of elderly normal subjects and Alzheimer's disease (AD) patients. It was subsequently discovered that the virus confers a strong risk of AD when in brain of carriers of the type 4 allele of the apolipoprotein E gene (apoE-epsilon4). This study has now sought, using PCR, the presence of three other herpesviruses in brain: human herpesvirus 6 (HHV6)-types A and B, herpes simplex virus type 2 (HSV2) and cytomegalovirus (CMV). HHV6 is present in a much higher proportion of the AD than of age-matched normal brains (70% vs. 40%, p=0.003) and there is extensive overlap with the presence of HSV1 in AD brains, but HHV6, unlike HSV1, is not directly associated in AD with apoE-epsilon4. In 59% of the AD patients' brains harbouring HHV6, type B is present while 38% harbour both type A and type B, and 3% type A. HSV2 is present at relatively low frequency in brains of both AD patients and normals (13% and 20%), and CMV at rather higher frequencies in the two groups (36% and 35%); in neither case is the difference between the groups statistically significant. It is suggested that the striking difference in the proportion of elderly brains harbouring HSV1 and HSV2 might reflect the lower proportion of people infected with the latter, or the difference in susceptibility of the frontotemporal regions to the two viruses. In the case of HHV6, it is not possible to exclude its presence as an opportunist, but alternatively, it might enhance the damage caused by HSV1 and apoE-epsilon4 in AD; in some viral diseases it is associated with characteristic brain lesions and it also augments the damage caused by certain viruses in cell culture and in animals. PMID- 12115888 TI - Expert system support using a Bayesian belief network for the classification of endometrial hyperplasia. AB - Accurate morphological classification of endometrial hyperplasia is crucial as treatments vary widely between the different categories of hyperplasia and are dependent, in part, on the histological diagnosis. However, previous studies have shown considerable inter-observer variation in the classification of endometrial hyperplasias. The aim of this study was to develop a decision support system (DSS) for the classification of endometrial hyperplasias. The system used a Bayesian belief network to distinguish proliferative endometrium, simple hyperplasia, complex hyperplasia, atypical hyperplasia and grade 1 endometrioid adenocarcinoma. These diagnostic outcomes were held in the decision node. Four morphological features were selected as diagnostic clues used routinely in the discrimination of endometrial hyperplasias. These represented the evidence nodes and were linked to the decision node by conditional probability matrices. The system was designed with a computer user interface (CytoInform) where reference images for a given clue were displayed to assist the pathologist in entering evidence into the network. Reproducibility of diagnostic classification was tested on 50 cases chosen by a gynaecological pathologist. These comprised ten cases each of proliferative endometrium, simple hyperplasia, complex hyperplasia, atypical hyperplasia and grade 1 endometrioid adenocarcinoma. The DSS was tested by two consultant pathologists, two junior pathologists and two medical students. Intra- and inter-observer agreement was calculated following conventional histological examination of the slides on two occasions by the consultants and junior pathologists without the use of the DSS. All six participants then assessed the slides using the expert system on two occasions, enabling inter- and intra-observer agreement to be calculated. Using unaided conventional diagnosis, weighted kappa values for intra-observer agreement ranged from 0.645 to 0.901. Using the DSS, the results for the four pathologists ranged from 0.650 to 0.845. Both consultant pathologists had slightly worse weighted kappa values using the DSS, while both junior pathologists achieved slightly better values using the system. The grading of morphological features and the cumulative probability curve provided a quantitative record of the decision route for each case. This allowed a more precise comparison of individuals and identified why discordant diagnoses were made. Taking the original diagnoses of the consultant gynaecological pathologist as the 'gold standard', there was excellent or moderate to good inter-observer agreement between the 'gold standard' and the results obtained by the four pathologists using the expert system, with weighted kappa values of 0.586-0.872. The two medical students using the expert system achieved weighted kappa values of 0.771 (excellent) and 0.560 (moderate to good) compared to the 'gold standard'. This study illustrates the potential of expert systems in the classification of endometrial hyperplasias. PMID- 12115889 TI - Recovery of song preferences after excitotoxic HVC lesion in female canaries. AB - The courtship solicitation display (CSD) of the female canary is a model to study estrogen dependent auditory preferences for male songs. The forebrain auditory vocal nucleus, HVC, is part of the circuit that determines such preferences. To further develop this model we show that bilateral excitotoxic lesions of the medial part of HVC involving between 18-60% of the bilateral nucleus are behaviorally effective while complete unilateral lesions are not. Further, we show that animals recover their song preferences over a period of several months after the lesion. This functional recovery does not involve anatomical recovery of the HVC. Even 9 months after the lesion, the HVC size of these females was similar to that of females sacrificed 2 days after the lesion and thus was 40 +/- 8% smaller compared to normal females. Further, ipsilaterally, the lesion procedure transiently disturbed the neurochemistry, such as GAD-mRNA expression, in the part of HVC that did not undergo cell death. These results suggest that the integrity of the lateral part of at least one HVC is required to perform CSD in response to relevant auditory stimuli. PMID- 12115890 TI - Distributions of estrogen receptors alpha and beta in sympathetic neurons of female rats: enriched expression by uterine innervation. AB - Estrogen modulates many features of the sympathetic nervous system, including cell numbers and ganglion synapses, and can induce uterine sympathetic nerve degeneration. However, distributions of estrogen receptors alpha and beta within sympathetic neurons have not been described, and their regulation by target tissue or estrogen levels has not been explored. We used immunofluorescence and retrograde tracing to define estrogen receptor expression in sympathetic neurons at large in pre- and paravertebral ganglia and in those projecting to the uterine horns. Estrogen receptor alpha immunoreactivity was present in 29 +/- 1%, while estrogen receptor beta was expressed by 92 +/- 1% of sympathetic neurons at large. The proportions of neurons expressing these receptors were comparable in the superior cervical and thoraco-lumbar paravertebral ganglia from T11 through L5, and in the suprarenal, celiac, and superior mesenteric prevertebral ganglia. Injections of FluoroGold into the uterine horns resulted in labeled neurons, with peak occurrences in T13, L1, and the suprarenal ganglion. Uterine-projecting neurons showed small but significantly greater incidence of estrogen receptor beta expression relative to the neuronal population at large, whereas the proportion of uterine-projecting neurons with estrogen receptor alpha immunoreactivity was nearly threefold greater. Numbers of estrogen receptor expressing neurons were not altered by acute estrogen administration. We conclude that the vast majority of sympathetic neurons express estrogen receptor beta immunoreactive protein, whereas a smaller, presumably overlapping subset expresses the estrogen receptor alpha. Expression of the latter apparently can be enhanced by target-mediated mechanisms. PMID- 12115891 TI - Regulation of neuronal excitability in Drosophila by constitutively active CaMKII. AB - The ability of calcium/calmodulin-dependent protein kinase II (CaMKII) to become calcium independent after autophosphorylation makes this enzyme a temporal marker of neuronal activity. Here we show that the calcium-independent form of CaMKII has unique effects on larval viability, locomotion, and neuronal excitability in Drosophila. Expression of constitutively active T287D, but not calcium-dependent T287A, mutant CaMKII in Drosophila neurons resulted in decreased viability, behavioral defects, and failure of action potential propagation. The actions of T287D may be mediated, at least in part, by increased potassium conductances. Expression of T287D CaMKII also stimulated an increase in the number of boutons at the larval neuromuscular junction, but did not affect the mechanics of release. This study defines a role for autophosphorylation of CaMKII in the regulation of multiple neuronal functions including the intrinsic properties of neurons. PMID- 12115892 TI - The hippocampus and caudomedial neostriatum show selective responsiveness to conspecific song in the female zebra finch. AB - The perception of song is vital to the reproductive success of both male and female songbirds. Several neural structures underlying this perception have been identified by examining expression of immediate early genes (IEGs) following the presentation of conspecific or heterospecific song. In the few avian species investigated, areas outside of the circuit for song production contain neurons that are active following song presentation, specifically the caudal hyperstriatum ventrale (cHV) and caudomedial neostriatum (NCM). While studied in detail in the male zebra finch, IEG responses in these neural substrates involved in song perception have not been quantified in females. Therefore, adult female zebra finches were presented with zebra finch song, nonzebra finch song, randomly generated tones, or silence for 30 min. One hour later they were sacrificed, and their brains removed, sectioned, and immunocytochemically processed for FOS expression. Animals exposed to zebra finch song had a significantly higher density of FOS-immunoreactive cells in the NCM than those presented with other songs, tones, or silence. Neuronal activation in the cHV was equivalent in birds that heard zebra finch and non-zebra finch song, expression that was higher than that observed in the groups that heard no song. Interestingly, the hippocampus (HP) and adjacent parahippocampal area (AHP) were activated in a manner comparable to the NCM. These results suggest a general role for the cHV in song perception and a more specific role for the NCM and HP/AHP in facilitating recognition of and responsiveness to species-specific song in female zebra finches. PMID- 12115893 TI - Activin and bone morphogenetic proteins are present in perinatal sensory neuron target tissues that induce neuropeptides. AB - Previous studies have shown that sensory target tissues induce neuropeptides in naive sensory neurons, and that activin and bone morphogenetic proteins (BMPs) are capable of inducing neuropeptides associated with nociception in embryonic sensory neurons in vitro. The goal of the present study was to learn if these ligands were available in native sensory neuron target tissues at correct developmental periods to play this inductive role in vivo. Sensory neurons initially contact their peripheral target tissues and begin to express neuropeptides during late embryogenesis, and we demonstrate that activin and BMPs are present in the embryo and neonate to regulate sensory neuron differentiation. Native embryonic and neonatal target tissues were analyzed by immunoblot and immunohistochemical studies using ligand-specific antibodies. Although activin was easily solubilized, BMPs were detected only after high salt extraction, suggesting that BMPs were bound to extracellular moieties and were capable of acting only locally in native tissues. One inhibitor, noggin, was present in both embryonic skin and muscle. In combination, these data suggest that neuronal differentiation is unlikely to be regulated by simple expression of ligand, but that the functional availability of ligand is a critical component confering biological activity. PMID- 12115894 TI - Altered odor-induced expression of c-fos and arg 3.1 immediate early genes in the olfactory system after familiarization with an odor. AB - In adult rats, repeated exposure to an odorant, in absence of any experimentally delivered reinforcement, leads to a drastic decrease in mitral/tufted (M/T) cell responsiveness, not only for the familiar odor but also for other novel odors. In the present study, using two different and complementary in situ hybridization methods, we analyzed the effect of familiarization with an odorant on c-fos and arg 3.1 mRNA expression levels, and we examined the odor specificity of this effect. Odor exposure induces a specific increase in c-fos and arg 3.1 expression in some particular olfactory bulb quadrants. Previous familiarization with the test odor results in a decreased expression of both IEGs in these quadrants, leading to the alteration of the odor-specific pattern of c-fos and arg 3.1 expression. In contrast, this odor-specific pattern is not affected when different odors are used for familiarization and test. Similarly, an odor specific familiarization effect leading to a reduced c-fos and arg 3.1 expression was also detected in the cingulate cortex and in the anterior piriform cortex. These results support our hypothesis that the decrease in M/T cell responsiveness following a preceding familiarization with an odorant may be related to a particular form of synaptic plasticity involving changes at the genomic level, and reveals further insight in olfactory information processing and the cellular mechanisms underlying familiarization in the olfactory system. PMID- 12115895 TI - Serotonergic sensory-motor neurons mediate a behavioral response to hypoxia in pond snail embryos. AB - Oxygen (O(2)) is one of the most important environmental factors that affects both physiological processes and development of aerobic animals, yet little is known about the neural mechanism of O(2) sensing and adaptive responses to low O(2) (hypoxia) during development. In the pond snail, Helisoma trivolvis, the first embryonic neurons (ENC1s) to develop are a pair of serotonergic sensory motor cells that regulate a cilia-driven rotational behavior. Here, we report that the ENC1-ciliary cell circuit mediates an adaptive behavioral response to hypoxia. Exposure of egg masses to hypoxia elicited a dose-dependent and reversible acceleration of embryonic rotation that mixed capsular fluid, thereby facilitating O(2) diffusion to the embryo. The O(2) partial pressures (Po(2)) for threshold, half-maximal, and maximal rotational response were 60, 28, and 13 mm Hg, respectively. During hypoxia, embryos relocated to the periphery of the egg masses where higher Po(2) levels occurred. Furthermore, intermittent hypoxia treatments induced a sensitization of the rotational response. In isolated ciliary cells, ciliary beating was unaffected by hypoxia, suggesting that in the embryo, O(2) sensing occurs upstream of the motile cilia. The rotational response of embryos to hypoxia was attenuated by application of the serotonin receptor antagonist, mianserin, correlated to the development of ENC1-ciliary cell circuit, and abolished by laser-ablation of ENC1s. Together, these data suggest that ENC1s are unique oxygen sensors that may provide a good single cell model for the examination of mechanistic, developmental, and evolutionary aspects of O(2) sensing. PMID- 12115897 TI - Fish gill morphology: inside out. AB - In this short review of fish gill morphology we cover some basic gross anatomy as well as in some more detail the microscopic anatomy of the branchial epithelia from representatives of the major extant groups of fishes (Agnathans, Elasmobranchs, and Teleosts). The agnathan hagfishes have primitive gill pouches, while the lampreys have arch-like gills similar to the higher fishes. In the lampreys and elasmobranchs, the gill filaments are supported by a complete interbranchial septum and water exits via external branchial slits or pores. In contrast, the teleost interbranchial septum is much reduced, leaving the ends of the filaments unattached, and the multiple gill openings are replaced by the single caudal opening of the operculum. The basic functional unit of the gill is the filament, which supports rows of plate-like lamellae. The lamellae are designed for gas exchange with a large surface area and a thin epithelium surrounding a well-vascularized core of pillar cell capillaries. The lamellae are positioned for the blood flow to be counter-current to the water flow over the gills. Despite marked differences in the gross anatomy of the gill among the various groups, the cellular constituents of the epithelium are remarkably similar. The lamellar gas-exchange surface is covered by squamous pavement cells, while large, mitochondria-rich, ionocytes and mucocytes are found in greatest frequency in the filament epithelium. Demands for ionoregulation can often upset this balance. There has been much study of the structure and function of the branchial mitochondria-rich cells. These cells are generally characterized by a high mitochondrial density and an amplification of the basolateral membrane through folding or the presence of an intracellular tubular system. Morphological subtypes of MRCs as well as some methods of MRC detection are discussed. PMID- 12115898 TI - Vascular anatomy of the fish gill. AB - The fish gill is the most physiologically diversified vertebrate organ, and its vasculature the most intricate. Application of vascular corrosion techniques that couple high-fidelity resins, such as methyl methacrylate, with scanning electron microscopy yields three-dimensional replicas of the microcirculation that have fostered a better appreciate gill perfusion pathways. This is the focus of the present review. Three vascular networks can be identified within the gill filament. The arterioarterial (respiratory) pathway consists of the lamellae and afferent and efferent segments of the branchial and filamental arteries and lamellar arterioles. The body of the filament contains two post-lamellar pathways: the interlamellar and nutrient. The interlamellar system is an extensive ladder-like network of thin-walled, highly distensible vessels that traverses the filament between, and parallel to, the lamellae and continues around the afferent and efferent borders of the filament. Interlamellar vessels are supplied by short, narrow-bore feeder vessels from the medial wall of the efferent filamental artery. A myriad of narrow-bore, tortuous arterioles arise from the basal efferent filamental artery and efferent branchial artery and anastomose to form the nutrient circulation of the arch and filament. In the filament body, nutrient capillaries and interlamellar vessels are often closely associated, and the former may ultimately drain into the latter. Many of the anatomical characteristics of interlamellar vessels are strikingly similar to those of mammalian lymphatic capillaries, with the exception that interlamellar vessels are directly fed by arteriovenous-like anastomoses. It is likely that gill interlamellar and mammalian lymphatics are physiologically, if not embryologically, equivalent. PMID- 12115899 TI - Branchial innervation. AB - Inspection of the dorsal end of fish gills reveals an impressive set of nerve trunks, connecting the gills to the brain. These trunks are branches of cranial nerves VII (the facial) and especially IX (the glossopharyngeal) and X (the vagus). The nerve trunks carry a variety of nervous pathways to and from the gills. A substantial fraction of the nerves running in the branchial trunks carry afferent (sensory) information from receptors within the gills. There are also efferent (motor) pathways, which control muscles within the gills, blood flow patterns and possibly secretory functions. Undertaking a more careful survey of the gills, it becomes evident that the arrangement of the microanatomy (particularly the blood vessels) and its innervation are strikingly complex. The complexity not only reflects the many functions of the gills but also illustrates that the control of blood flow patterns in the gills is of crucial importance in modifying the efficiency of its chief functions: gas transfer and salt balance. The "respiratory-osmoregulatory compromise" is maintained by minimizing the blood/water exchange (functional surface area of the gills) to a level where excessive water loss (marine teleosts) or gain (freshwater teleosts) is kept low while ensuring sufficient gas exchange. This review describes the arrangement and mechanisms of known nervous pathways, both afferent and efferent, of fish (notably teleosts) gills. Emphasis is placed primarily on the autonomic nervous system and mechanisms of blood flow control, together with an outline of the afferent (sensory) pathways of the gill arches. PMID- 12115900 TI - Sensing and transfer of respiratory gases at the fish gill. AB - The gill is both a site of gas transfer and an important location of chemoreception or gas sensing in fish. While often considered separately, these two processes are clearly intricately related because the gases that are transferred between the ventilatory water and blood at the gill are simultaneously sensed by chemoreceptors on, and within, the gill. Modulation of chemoreceptor discharge in response to changes in O(2) and CO(2) levels, in turn, is believed to initiate a series of coordinated cardiorespiratory reflexes aimed at optimising branchial gas transfer. The past decade has yielded numerous advances in terms of our understanding of gas transfer and gas sensing at the fish gill, particularly concerning the transfer and sensing of carbon dioxide. In addition, recent research has moved from striving to construct a single model that covers all fish species, to recognition of the considerable inter-specific variation that exists with respect to the mechanics of gas transfer and the cardiorespiratory responses of fish to changes in O(2) and CO(2) levels. The following review attempts to integrate gas transfer and gas sensing at the fish gill by exploring recent advances in these areas. PMID- 12115901 TI - Na(+), Cl(-), Ca(2+) and Zn(2+) transport by fish gills: retrospective review and prospective synthesis. AB - The secondary active Cl(-) secretion in seawater (SW) teleost fish gills and elasmobranch rectal gland involves basolateral Na(+),K(+)-ATPase and NKCC, apical membrane CFTR anion channels, and a paracellular Na(+)-selective conductance. In freshwater (FW) teleost gill, the mechanism of NaCl uptake is more controversial and involves apical V-type H(+)-ATPase linked to an apical Na(+) channel, apical Cl(-)-HCO-3 exchange and basolateral Na(+),K(+)-ATPase. Ca(2+) uptake (in FW and SW) is via Ca(2+) channels in the apical membrane and Ca(2+)-ATPase in the basolateral membrane. Mainly this transport occurs in mitochondria rich (MR) chloride cells, but there is a role for the pavement cells also. Future research will likely expand in two major directions, molded by methodology: first in physiological genomics of all the transporters, including their expression, trafficking, operation, and regulation at the molecular level, and second in biotelemetry to examine multivariable components in behavioral physiological ecology, thus widening the integration of physiology from the molecular to the environmental levels while deepening understanding at all levels. PMID- 12115902 TI - Ammonia excretion and urea handling by fish gills: present understanding and future research challenges. AB - In fresh water fishes, ammonia is excreted across the branchial epithelium via passive NH(3) diffusion. This NH(3) is subsequently trapped as NH(4)(+) in an acidic unstirred boundary layer lying next to the gill, which maintains the blood to-gill water NH(3) partial pressure gradient. Whole animal, in situ, ultrastructural and molecular approaches suggest that boundary layer acidification results from the hydration of CO(2) in the expired gill water, and to a lesser extent H(+) excretion mediated by apical H(+)-ATPases. Boundary layer acidification is insignificant in highly buffered sea water, where ammonia excretion proceeds via NH(3) diffusion, as well as passive NH(4)(+) diffusion due to the greater ionic permeability of marine fish gills. Although Na(+)/H(+) exchangers (NHE) have been isolated in marine fish gills, possible Na(+)/NH(4)(+) exchange via these proteins awaits evaluation using modern electrophysiological and molecular techniques. Although urea excretion (J(Urea)) was thought to be via passive diffusion, it is now clear that branchial urea handling requires specialized urea transporters. Four urea transporters have been cloned in fishes, including the shark kidney urea transporter (shUT), which is a facilitated urea transporter similar to the mammalian renal UT-A2 transporter. Another urea transporter, characterized but not yet cloned, is the basolateral, Na(+) dependent urea antiporter of the dogfish gill, which is essential for urea retention in ureosmotic elasmobranchs. In ureotelic teleosts such as the Lake Magadi tilapia and the gulf toadfish, the cloned mtUT and tUT are facilitated urea transporters involved in J(Urea). A basolateral urea transporter recently cloned from the gill of the Japanese eel (eUT) may actually be important for urea retention during salt water acclimation. A multi-faceted approach, incorporating whole animal, histological, biochemical, pharmacological, and molecular techniques is required to learn more about the location, mechanism of action, and functional significance of urea transporters in fishes. PMID- 12115903 TI - Acid-base regulation in fishes: cellular and molecular mechanisms. AB - The mechanisms underlying acid-base transfers across the branchial epithelium of fishes have been studied for more than 70 years. These animals are able to compensate for changes to internal pH following a wide range of acid-base challenges, and the gill epithelium is the primary site of acid-base transfers to the water. This paper reviews recent molecular, immunohistochemical, and functional studies that have begun to define the protein transporters involved in the acid-base relevant ion transfers. Both Na(+)/H(+) exchange (NHE) and vacuolar type H(+)-ATPase transport H(+) from the fish to the environment. While NHEs have been thought to carry out this function mainly in seawater-adapted animals, these proteins have now been localized to mitochondrial-rich cells in the gill epithelium of both fresh and saltwater-adapted fishes. NHEs have been found in the gill epithelium of elasmobranchs, teleosts, and an agnathan. In several species, apical isoforms (NHE2 and NHE3) appear to be up-regulated following acidosis. In freshwater teleosts, H(+)-ATPase drives H(+) excretion and is indirectly coupled to Na(+) uptake (via Na(+) channels). It has been localized to respiratory pavement cells and chloride cells of the gill epithelium. In the marine elasmobranch, both branchial NHE and H(+)-ATPase have been identified, suggesting that a combination of these mechanisms may be utilized by marine elasmobranchs for acid-base regulation. An apically located Cl(-)/HCO(3)(-) anion exchanger in chloride cells may be responsible for base excretion in fresh and seawater-adapted fishes. While only a few species have been examined to date, new molecular approaches applied to a wider range of fishes will continue to improve our understanding of the roles of the various gill membrane transport processes in acid-base balance. PMID- 12115904 TI - Gill circulation: regulation of perfusion distribution and metabolism of regulatory molecules. AB - The fish gill is the primary regulatory interface between internal and external milieu and a variety of neurocrine, endocrine, paracrine, and autocrine signals coordinate and control gill functions. Many of these messengers also affect gill vascular resistance, and they, in turn, may be inactivated (or activated) by branchial vessels. Few studies have critically addressed how flow is distributed within the gill filament, the physiological consequences thereof, or the impact of gill hormone metabolism on gill and systemic homeostasis. In most fish, the entire cardiac output perfuses the arterioarterial pathway, and this network probably accounts for the majority of passive- and stimulus-induced changes in vascular resistance. The in-series arrangement of the extensive gill microcirculation with systemic vessels is also indicative of a high capacity for metabolism of plasma-borne messengers as well as xenobiotics. Adenosine, arginine vasotocin (AVT), and endothelin (ET) are the most potent gill constrictors identified to date, and all decrease lamellar perfusion. Perhaps not surprising, they are also inactivated by gill vessels. Acetylcholine favors perfusion of the alamellar filamental vasculature, although the physiological relevance of acetylcholine-mediated responses remains unclear. Angiotensin, bradykinin, urotensin, natriuretic peptides, prostaglandins, and nitric oxide are vasoactive to varying degrees, but their effects on intrafilamental blood flow are unknown. If form befits function, then the complex vascular anatomy of the gill suggests a level of regulatory sophistication unparalleled in other vertebrate organs. Resolution of these issues will be technically challenging but unquestionably rewarding. PMID- 12115905 TI - Cell signaling and ion transport across the fish gill epithelium. AB - A large array of circulating and local signaling agents modulate transport of ions across the gill epithelium of fishes by either affecting transport directly or by altering the size and distribution of transporting cells in the epithelium. In some cases, these transport effects are in addition to cardiovascular effects of the same agents, which may affect the perfusion pathways in the gill vasculature and, in turn, affect epithelial transport indirectly. Prolactin is generally considered to function in freshwater, because it is the only agent that allows survival of some hypophysectomized fish species in freshwater. It appears to function by either reducing branchial permeability, Na,K-activated ATPase activity, or reducing the density of chloride cells. Cortisol was initially considered to produce virtually opposite effects (e.g., stimulation of Na,K activated ATPase and of chloride cell size and density), but more recent studies have found that this steroid stimulates ionic uptake in freshwater fishes, as well as the activity of H-ATPase, an enzyme thought to be central to ionic uptake. Thus, cortisol may function in both high and low salinities. Growth hormone and insulin-like growth factor appear to act synergistically to affect ion regulation in seawater fishes, stimulating both Na,K-activated ATPase and Na K-2Cl co-transporter activity, and chloride cell size, independent of their effects on growth. Some of the effects of the GH-IGF axis may be via stimulation of the number of cortisol receptors. Thyroid hormones appear to affect seawater ion regulation indirectly, by stimulating the GH-IGF axis. Natriuretic peptides were initially thought to stimulate gill ionic extrusion, but recent studies have not corroborated this finding, so it appears that the major mode of action of these peptides may be reduction of salt loading by inhibition of oral ingestion and intestinal ionic uptake. Receptors for both arginine vasotocin and angiotensin have been described in the gill epithelium, but their respective roles and importance in fish ion regulation remains unknown. The gill epithelium may be affected by both circulating and local adrenergic agents, and a variety of studies have demonstrated that stimulation of alpha-adrenergic versus beta adrenergic receptors produces inhibition or stimulation of active salt extrusion, respectively. Local effectors, such as prostaglandins, nitric oxide, and endothelin, may affect active salt extrusion as well as gill perfusion. Recent studies have suggested that the endothelin inhibition of salt extrusion is actually mediated by the release of both NO and prostaglandins. It is hoped that modern molecular techniques, combined with physiological measurements, will allow the dissection of the relative roles in ion transport across the fish gill epithelium of this surprisingly large array of putative signaling agents. PMID- 12115906 TI - Identification of alpha1-adrenergic receptors and their involvement in phosphoinositide hydrolysis in the frog heart. AB - The aim of this study was to characterize alpha(1)-adrenergic receptors in frog heart and to examine their related signal transduction pathway. alpha(1) Adrenergic binding sites were studied in purified heart membranes using the specific alpha(1)-adrenergic antagonist [(3)H]prazosin. Analysis of the binding data indicated one class of binding sites displaying a K(d) of 4.19 +/- 0.56 nM and a B(max) of 14.66 +/- 1.61 fmol/mg original wet weight. Adrenaline, noradrenaline, or phenylephrine, in the presence of propranolol, competed with [(3)H]prazosin binding with a similar potency and a K(i) value of about 10 microM. The kinetics of adrenaline binding was closely related to its biological effect. Adrenaline concentration dependently increased the production of inositol phosphates in the heart in the presence or absence of propranolol. Maximal stimulation was about 8.5-fold, and the half-maximum effective concentration was 30 and 21 microM in the absence and presence of propranolol, respectively. These data clearly show that alpha(1)-adrenergic receptors are coupled to the phosphoinositide hydrolysis in frog heart. To our knowledge, this is the first direct evidence supporting the presence of functional alpha(1)-adrenergic receptors in the frog heart. PMID- 12115907 TI - Dynamics of Na(+),K(+),2Cl(-) cotransporter and Na(+),K(+)-ATPase expression in the branchial epithelium of brown trout (Salmo trutta) and Atlantic salmon (Salmo salar). AB - The dynamics of branchial Na(+),K(+),2Cl(-) cotransporter (NKCC) and Na(+),K(+) ATPase (NKA) expression were investigated in brown trout and Atlantic salmon during salinity shifts and the parr-smolt transformation, respectively. In the brown trout, Western blotting revealed that NKCC and NKA abundance increased gradually and in parallel (30- and ten-fold, respectively) after transfer to seawater (SW). The NKA hydrolytic activity increased ten-fold after SW-transfer. Following back-transfer to fresh water (FW), the levels of both proteins and NKA activity decreased. The NKCC immunostaining in the gill of SW-acclimated trout was strong, and mainly localized in large cells in the filament and around the bases of the lamellae. In FW-acclimated trout, immunostaining was less intense and more diffuse. Partial cDNAs of the secretory NKCC1 isoform were cloned and sequenced from both brown trout and Atlantic salmon gills. Two differently sized transcripts were detected by Northern blotting in the gill but not in other osmoregulatory tissues (kidney, pyloric caeca, intestine). The abundance in the gill of these transcripts and of the associated NKCC protein increased four- and 30-fold, respectively, during parr-smolt transformation. The abundance of NKA alpha-subunit protein also increased in the gill during parr-smolt transformation though to a lesser extent than enzymatic activity (2.5- and eight-fold, respectively). In separate series of in vitro experiments, cortisol directly stimulated the expression of NKCC mRNA in gill tissue of both salmonids. The study demonstrates the coordinated regulation of NKCC and NKA proteins in the gill during salinity shifts and parr-smolt transformation of salmonids. PMID- 12115908 TI - Leptin, ghrelin, and energy metabolism of the spawning burbot (Lota lota, L.). AB - The aim of this study was to investigate the energy metabolism of the burbot (Lota lota, n=38) before, during, and after spawning, which represents the greatest annual metabolic demand for the species. A decrease in body mass, relative weight of the livers, and glycogen concentration of the livers was observed toward the end of spawning. The prespawning period was characterized by high rates of liver glycogenolysis and lipid mobilization. Also, plasma triiodothyronine and sex steroid levels were high before reproduction. During spawning, liver lipolysis was reduced and muscle glycogenolysis stimulated. The levels of triiodothyronine and sex steroids decreased. After reproduction, liver glycogenolysis was suppressed and the rate of gluconeogenesis increased. Thyroid hormone levels were elevated after spawning. Leptin protein and a ghrelin immunoreactive peptide were detected in burbot plasma. Their concentrations were relatively low before and during reproduction but increased after spawning. The functions of leptin and the ghrelin-immunoreactive peptide in the physiology of the burbot are not consistent with the models of their function in mammals. PMID- 12115909 TI - Phenotype plasticity in postural muscles of the crayfish Orconectes limosus Raf.: correlation of myofibrillar ATPase-based fiber typing with electrophysiological fiber properties and the effect of chronic nerve stimulation. AB - The characteristics of the medial and lateral superficial extensor muscles (sem and sel) in the crayfish Orconectes limosus abdomen and their developmental and activity-dependent plasticity were studied. It was shown that both muscles are innervated by at least five excitatory and one inhibitory motor neuron in a nonuniform pattern. The muscles are composed of at least three different mATPase histochemistry-based fiber types that are all different from a fourth type in the uniform deep extensor muscles. sem and sel are composed of different ratios of these fiber types but do not show a constant fiber type pattern between segments and even between hemisegments. The three histochemically defined superficial extensor-fiber types have characteristic electrophysiological properties. The fiber types were shown to develop successively during the first postembryonic stages of development without a change in the number of muscle fibers. Based on histochemical ATPase staining after 21 days of chronic stimulation by means of an implantable, double-hook electrode, we show preliminary evidence that the fiber composition in the sem can switch from the presumably fast fiber type III to an intermediate type II. Repeated axotomy up to 53 days had no effect on the fiber type composition of the muscles. PMID- 12115910 TI - The reptilian oviduct: a review of structure and function and directions for future research. AB - The reptilian oviduct is a complex organ with a variety of functions (albumen production, eggshell production, placentation, oviposition or parturition, and sperm storage), depending on the parity mode of the species in question. These functions are under complex physiological control, the details of which are far from understood. The aims of this review are to summarise the information available concerning the structure and functions of the reptilian oviduct and to highlight areas in particular need of further research. PMID- 12115911 TI - Inbreeding in Littledale's whistling rat Parotomys littledalei. AB - Despite its rarity in nature, inbreeding is sometimes evident in species occupying ephemeral, unpredictable habitats, and which occur at low densities. One such species is Littledale's whistling rat, Parotomys littledalei, a murid rodent endemic to the south-west arid region of South Africa. Using a captive population of P. littledalei, I studied mate choice for kin and nonkin, and the reproductive performance of inbred and outbred pairs. In choice tests, estrous females presented with either odors or actual males showed a preference for siblings or half-siblings to unrelated males. Males did not discriminate between the odor of estrous kin and nonkin. In breeding studies, inbred (mother-son; brother-sister) and outbred (proven female and an unrelated young male and nonsiblings) pairs had a similar reproductive output, although the sex ratio favored males in inbred litters. The development of inbred young was indistinguishable from outbred young. The results indicate that female P. littledalei prefer to inbreed, but there are no apparent advantages to inbreeding over outbreeding. PMID- 12115913 TI - Cyclopamine induces digit loss in regenerating axolotl limbs. AB - Axolotls, with their extensive ability to regenerate as adults, provide a useful model for studying the mechanisms of regeneration in a vertebrate, in hopes of understanding why other vertebrates cannot regenerate. Although the expression of many genes has been described in regeneration, techniques for gain and loss of function analyses have been limited. We demonstrated in a previous study that gain of function for secreted proteins was possible in the axolotl using the vaccinia virus to drive expression of the transgene. In this study, we used a pharmacological approach made possible by the existence of a naturally occurring compound that specifically blocks shh signaling, cyclopamine. The treatment of axolotls with cyclopamine during the process of limb regeneration caused a loss of digits similar to that described for the shh knockout mouse. Our results further demonstrate that shh signaling and function are conserved during limb regeneration in urodeles as in limb development in other vertebrates. PMID- 12115912 TI - Ped gene deletion polymorphism frequency in wild mice. AB - The Ped gene influences the rate of cleavage of preimplantation embryos and their subsequent survival. Embryos that express the product of the Ped gene, Qa-2 protein, cleave at a faster rate than embryos with an absence of Qa-2 protein. In addition, the Ped gene has pleiotropic effects on reproduction. Thus, there is a reproductive advantage to those mouse strains that are Qa-2 positive. The presence or absence of Qa-2 is reflected at the DNA level by the presence or absence (deletion polymorphism) of the gene(s) encoding Qa-2 protein. Many inbred and wild-derived mouse strains have been characterized as Qa-2 positive or negative, but no previous studies have looked at the distribution of the Ped gene in a population of free-living wild mice. The purpose of this study was to determine the Ped gene deletion polymorphism frequency in a sample of free-living wild mice. Twenty-nine mice were collected and identified as Mus musculus. Genomic DNA extraction was performed on tail tips, and PCR was used to amplify a region from the Ped gene. Known Qa-2 positive and negative mice were used as controls. Results showed that all 29 wild mice were positive for the Ped gene. Since the Ped gene is dominant and provides a reproductive advantage, it is not surprising that all of the wild mice were Qa-2 positive. However, our assay could not distinguish homozygous from heterozygous mice. It is possible that the Qa-2 deletion polymorphism is segregating in the population, and a larger sample size would identify some Qa-2 negative mice. PMID- 12115915 TI - Branchial chloride cells in sea bass (Dicentrarchus labrax) adapted to fresh water, seawater, and doubly concentrated seawater. AB - Branchial chloride cells (CC) were studied in sea bass (Dicentrarchus labrax) maintained in seawater (SW: 35 per thousand) or gradually adapted to and subsequently maintained in fresh water (0.2 per thousand) or doubly concentrated seawater (DSW: 70 per thousand). Changes were observed in the location, number, and structure of CCs, that were discriminated by light, scanning, and transmission electron microscopy, as well as by immunofluorescence on the basis of their high Na(+)/K(+)-ATPase antigen content. The number of CCs increased in both fresh water and doubly concentrated seawater compared to control fish maintained in SW. In both experimental conditions, these cells were found on the gill filament (as in control fish) and even on the lamellae, especially in hypersaline conditions. Structural changes concerned the shapes and sizes of CCs and their apical outcrops and particularly the structures of their functional complexes (mitochondria, tubular system, and endoplasmic reticulum), which developed significantly in DSW adapted fish. The changes in the expression of the Na(+)/K(+)-ATPase were evaluated by assessing the enzyme's density at the ultrastructural level following immunogold labeling. This parameter was significantly higher in doubly concentrated seawater. The adaptative significance of the quantitative and morphofunctional changes in branchial chloride cells is discussed in relation to the original osmoregulatory strategy of this marine euryhaline teleost. PMID- 12115914 TI - Hg(2+) affects the intracellular free Ca(2+) oscillatory pattern and the correlated membrane conductance changes in Mg(2+)-stimulated oocytes of the prawn Palaemon serratus. AB - The impact of mercuric ions (Hg(2+)) on prawn oocytes was examined. Prawn oocytes constitute an unusual system in that they are activated at spawning by seawater Mg(2+), which mediates correlated dynamic changes in intracellular free calcium concentration [(Ca(2+))(i)] and membrane conductance associated with the meiosis resumption. Using a voltage clamp technique and intracellular calcium measurements, we observed that treatment with Hg(2+) (5, 10, and 20 microM) resulted in simultaneous impairments of both (Ca(2+))(i) and membrane current responses to external Mg(2+). Treatment with Hg(2+) also resulted in a gradual dose-dependent slow increase in the baseline level of both (Ca(2+))(i) and membrane conductance, independent of stimulation with external Mg(2+). The effect of Hg(2+) on (Ca(2+))(i) and membrane conductance changes resulted from a block of the signal transduction pathway at some point before the InsP(3) receptor channel involved in Ca(2+) release from the endoplasmic reticulum (ER) stocks. The Hg(2+)-dependent gradual increase in both (Ca(2+))(i) and membrane conductance baseline levels may potentially result from a slow permeabilization of the ER membrane, resulting in Ca(2+) leaking into the cytosol. Indeed, this effect could be blocked with the cell permeable Hg(2+) competitor dithiothreitol, which was able to displace Hg(2+) from its intracellular target regardless of whether external Ca(2+) was present or not. PMID- 12115916 TI - Immunocytochemical analysis of beta keratins in the epidermis of chelonians, lepidosaurians, and archosaurians. AB - Beta (beta) keratins are present only in the avian and reptilian epidermises. Although much is known about the biochemistry and molecular biology of the beta keratins in birds, little is known for reptiles. In this study we have examined the distribution of beta keratins in the adult epidermis of turtle, lizard, snake, tuatara, and alligator using light and electron immunocytochemistry with a well-characterized antiserum (anti-beta(1) antiserum) made against a known avian scale type beta keratin. In lizard, snake, and tuatara epidermis this antiserum reacts strongly with the beta-layer, more weakly with the oberhautchen before it merges with the beta-layer, and least intensely with the mesos layer. In addition, the anti-beta(1) antiserum reacts specifically with the setae of climbing pads in gekos, the plastron and carapace of turtles, and the stratum corneum of alligator epidermis. Electron microscopic studies confirm that the reaction of the anti-beta(1) antiserum is exclusively with characteristic bundles of the 3-nm beta keratin filaments in the cells of the forming beta-layer, and with the densely packed electron-lucent areas of beta keratin in the mature bet- layer. These immunocytochemical results suggest that the 3-nm beta keratin filaments of the reptilian integument are phylogenetically related to those found in avian epidermal appendages. PMID- 12115917 TI - Elevated levels of trimethylamine oxide in deep-sea fish: evidence for synthesis and intertissue physiological importance. AB - Tissue levels of trimethylamine oxide (TMAO) were compared for seven teleost and two elasmobranch species captured from three depth ranges: shallow (<150 m), moderate (500-700 m), and deep (1,000-1,500 m). Within the teleosts, the deep caught species had significantly greater TMAO content than shallow- or moderate caught species. In all teleosts, muscle had substantially more TMAO than all other tissues. Kidney or, in some cases, liver had elevated trimethylamine (TMA) content, 2.20-9.65 mmol/kg, along with appreciable trimethylamine oxidase (TMAoxi) activity, suggesting active TMAO synthesis. No correlation was found between TMAoxi activity and TMAO content. The elasmobranchs in this study, Squalus acanthias and Centroscyllium fabricii from shallow and deep water, respectively, were both squaliform sharks. The deep-caught species had significantly more TMAO in all tissues than the shallow species. Furthermore, urea was significantly less in the deep species in all tissues except liver, while the urea:TMAO ratio was significantly less in all tissues. As with teleosts, the TMAO content of muscle was substantially higher for both elasmobranchs than in all other tissues. TMAoxi was below levels of detection in both elasmobranch species, suggesting that TMAO is obtained solely from the diet. This study expands the trend of increased muscle TMAO in deep-sea fish to a variety of other tissues. The accumulation of TMAO in various tissues in deep-sea teleosts and the accumulation of TMAO and concurrent urea decrease in a deep-sea elasmobranch in comparison to a shallow water species strongly support the contention that TMAO is of physiological importance in deep-sea fish. PMID- 12115918 TI - Transfection of an inducible trout anion exchanger (AE1) into HEK-EcR cells. AB - A permanent cell line with inducible expression of the trout anion exchanger protein (trAE1) was constructed in a derivative of human embryonic kidney cells (HEK-293). In the absence of the inducer, muristerone A, the new cell line had no detectable trAE1 protein by Western analysis, biotinylation, and (36)Cl(-) flux. The amount of trAE1 protein increased with increasing dose and incubation time with muristerone A. Anion exchange inhibitors significantly inhibited the inducible flux of anions (i.e., (36)chloride and (35)sulfate) and taurine in isotonic media. The transfected cells had the characteristics of trAE1-mediated transport in intact trout erythrocytes: (1) inhibition by anion transport inhibitors, (2) pH independence over the pH range of 6.5-7.5, and (3) activation of (35)sulfate efflux by external anions in the selective order of Cl > Br > I > or = F. These cells, in contrast to trout erythrocytes, were not sensitive to the anion exchange inhibitor, 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS), suggesting some difference in the properties of the transfected AE1. These results demonstrate the inducible expression of new anion transport membrane protein in HEK-293 cells. This is the first expression of trAE1 in a mammalian system. PMID- 12115919 TI - Experimental manipulation of steroid concentrations in circulation and in egg yolks of turtles. AB - Steroid hormones in egg yolks are increasingly recognized as an important component of maternal and offspring fitness in oviparous vertebrates. Yet, except for in birds, the mechanism by which females allocate these resources is poorly understood. We manipulated systemic levels of hormones in reproductively mature female red-eared slider turtles (Trachemys scripta elegans) with silastic implants to test the hypothesis that hormones are allocated to developing follicles as a quantitative function of circulating levels in the females. Turtles exhibited similar amounts (<1 ng/ml) of circulating steroids (dihydrotestosterone, estradiol-17 beta, or testosterone) in early September immediately prior to experimental manipulation. After treatment with silastic implants, circulating levels of steroids increased markedly. By the following April after hibernation, circulating levels of dihydrotestosterone had returned to preimplantation levels, but circulating levels of estradiol-17 beta and testosterone in estradiol-17 beta- and testosterone-treated turtles, respectively, remained substantially elevated through April. Focusing on testosterone, we detected nearly six-fold higher concentrations in yolk from mature follicles from testosterone-treated turtles than in yolk from mature follicles from control turtles. Our results provide support for the hypothesis that concentrations of steroids in egg yolks of turtles reflect circulating concentrations of steroids during follicular development rather than the hypothesis that females selectively allocate specific amounts of steroid hormones to each egg separately. Our findings also highlight an unambiguous physiological mechanism by which nongenetic maternal effects in oviparous species can directly influence the nutritional milieu experienced by developing embryos. PMID- 12115920 TI - Maternally derived yolk hormones vary in follicles of the painted turtle, Chrysemys picta. AB - The transfer of hormones from a female to her offspring is known to occur in egg laying vertebrates, and the potential for these early, maternally derived hormones to influence sex determination in reptiles with temperature-dependent sex determination is intriguing. In the present study, we examine variation in the concentrations of progesterone, testosterone, and estradiol among three follicle size classes within a female painted turtle (Chrysemys picta) and among females across four periods that span the pre- to post-nesting season. Females were collected, and both follicles and shelled eggs (when present) were harvested for hormone analysis. Progesterone levels did not vary seasonally. However, the concentration of progesterone did vary among and within follicle classes, and was primarily dependent upon ovulatory state: Recently ovulated follicles (as yolks within shelled eggs) contained significantly more progesterone than unovulated follicles. Concentrations of testosterone were low and did not vary either among size classes or across the season. Estradiol levels decreased with increasing follicle size and were higher later in the nesting season. Thus, hormone concentrations varied among follicle sizes and states but in patterns that differed among hormones. This variation has the potential to influence sex determination. PMID- 12115921 TI - Pituitary adenylate cyclase-activating polypeptide induces testicular testosterone synthesis through PGE(2) mediation in crested newt, Triturus carnifex. AB - The aim of the present work was to study the possible role of adenylate cyclase activating polypeptide (PACAP) 38 in the testicular intracellular mechanism regulating steroidogenesis of crested newt, Triturus carnifex. Gonads were incubated in vitro with PACAP 38 and prostaglandin (PG) E(2) alone or with inhibitors of cyclooxygenase (COX), adenylate cyclase (AC), and phospholipase C (PLC) for 30 min and 60 min. PGE(2), PGF(2 alpha), testosterone, and estradiol-17 beta were measured in the culture medium; aromatase (AR) activity and cAMP were assessed in the tissue. PACAP 38 increased PGE(2) (30 min and 60 min), estradiol 17 beta (60 min), cAMP (60 min), and AR (60 min) but decreased testosterone (60 min). PGE(2) increased estradiol-17 beta, cAMP, and AR and decreased testosterone at 30 and 60 min.PLC inhibitor counteracted the effects of PACAP 38, while AC inhibitor counteracted these effects except for PGE(2) increase. AC inhibitor counteracted the effects of PGE(2), while PLC did not. COX inhibitor decreased PGF(2 alpha) (30 min and 60 min), PGE(2) (30 min and 60 min), estradiol-17 beta (60 min), cAMP (60 min), and AR (60 min), but increased testosterone (60 min). These in vitro results suggest that, in newt testis, PACAP 38 acts on PLC, inducing the increase of PGE(2) which, in turn, acting on AC, increases AR activity with the consequent estradiol-17 beta increase and testosterone decrease. PMID- 12115923 TI - Chloride-ATPase dephosphorylation in Aplysia gut. AB - The present study was done primarily to compare cation-ATPase dephosphorylation kinetics with a Cl(-)-ATPase's dephosphorylation kinetics because of the paucity of information in this area. Utilizing a proteoliposomal preparation containing Cl(-)-ATPase from Aplysia gut, it was demonstrated that dephosphorylation of this P-type ATPase was absolutely dependent upon Cl(-). Adenosine triphosphate (ATP) concentrations directly stimulated dephosphorylation of Cl(-)-ATPase in the presence of increasing concentrations of Cl(-). It was also shown that the calculated rate constant for E(1)-P disintegration was 20/sec. This rate constant value approximated E(1)-P rate constant disintegration values for other electrogenic, uniport P-type ATPases. Therefore, it was concluded from these results that the Cl(-)-ATPase dephosphorylation kinetics did not differ greatly from cation-ATPase dephosphorylation kinetics. PMID- 12115922 TI - An experimental test of the relationship between temporal variability of feeding opportunities and baseline levels of corticosterone in a shorebird. AB - In this study, we tested the hypothesis that baseline corticosterone levels increase with a change from constant to variable feeding schedules. Captive red knots, Calidris canutus, were presented with food that was either available during the same time each day (constant) or starting at variable times during the day. Food intake rates, frequency of aggressive interactions, and baseline levels of corticosterone were measured. In the majority of cases, red knots showed higher plasma corticosterone concentrations during feeding schedules that were irregular than when food was available at consistent times. These findings are supported by a previous study that showed that red knots take a long time to adjust to the newly offered, predictable conditions of their aviary environment. The frequency of conflicts in the different groups and (size-corrected) body mass were not correlated with average corticosterone level. The results are examined in the light of literature showing that increases in corticosterone in response to acute, unpredictable events mediate behavioral responses such as increased explorative behavior and memory. For red knots that have to find their food on the temperate-zone mudflats in Western Europe, an increased circulating corticosterone level may be adaptive during periods when the patchily distribution of buried bivalves and the burying behavior of such prey presents them with a variable and unpredictable food supply. PMID- 12115924 TI - Acceleration of Ambystoma tigrinum metamorphosis by corticotropin-releasing hormone. AB - Previous work of others and ours has shown that corticotropin-releasing hormone (CRH) is a positive stimulus for thyroid and interrenal hormone secretion in amphibian larvae and that activation of CRH neurons may mediate environmental effects on the timing of metamorphosis. These studies have investigated CRH actions in anurans (frogs and toads), whereas there is currently no information regarding the actions of CRH on metamorphosis of urodeles (salamanders and newts). We tested the hypothesis that CRH can accelerate metamorphosis of tiger salamander (Ambystoma tigrinum) larvae. We injected tiger salamander larvae with ovine CRH (oCRH; 1 microg/day; i.p.) and monitored effects on metamorphosis by measuring the rate of gill resorption. oCRH-injected larvae completed metamorphosis earlier than saline-injected larvae. There was no significant difference between uninjected and saline-injected larvae. Mean time to reach 50% reduction in initial gill length was 6.9 days for oCRH-injected animals, 11.9 days for saline-injected animals, and 14.1 days for uninjected controls. At the conclusion of the experiment (day 15), all oCRH-injected animals had completed metamorphosis, whereas by day 15, only 50% of saline-injected animals and 33% of uninjected animals had metamorphosed. Our results show that exogenous oCRH can accelerate metamorphosis in urodele larvae as it does in anurans. These findings suggest that the neuroendocrine mechanisms controlling metamorphosis are evolutionarily conserved across amphibian taxa. PMID- 12115925 TI - Differential induction of branchial carbonic anhydrase and NA(+)/K(+) ATPase activity in the euryhaline crab, Carcinus maenas, in response to low salinity exposure. AB - The time course of induction of activity of carbonic anhydrase (CA) and Na/K ATPase, two enzymes that are central to osmotic and ionic regulation in the eyryhaline green crab, Carcinus maenas, was measured in response to a transfer from 32 to 10 ppt salinity. CA activity was low in all gills in crabs acclimated to high salinity. Activity was induced in the posterior three gills (G6-G9) starting at 96 hr following transfer to low salinity, with activity peaking at seven post-transfer. Na/K ATPase activity in posterior gills was already high in crabs acclimated to 32 ppt salinity, and it did not increase as a result of transfer to 10 ppt. Acclimation of crabs to hypersaline (40 ppt) conditions resulted in uniformly low levels of Na/K ATPase activity, and transfer from 40 ppt to 10 ppt stimulated a four-fold induction of activity in the posterior gills that was evident by seven days of low salinity exposure. Low salinity stimulates the activity of both enzymes, but a different degree of salinity change appears to be necessary to cause the induction of each enzyme. The Na/K ATPase activity is already high at a salinity (32 ppt) at which the crab is still an osmotic and ionic conformer. CA activity, however, even when expressed in low levels, is still present in excess of what is needed to supply counterions at a rate adequate to match the rate of active ion transport. It is possible that two strategies exist for the regulation of these two enzymes that coincide with the crab's intertidal and estuarine lifestyle: short-term modulation of activity of highly expressed enzyme (Na/K ATPase) and long-term modulation of enzyme concentration by changes in gene expression (CA). For all ranges of low salinity exposure, crabs undergo hemodilution, cell swelling, and subsequent cell volume readjustment as evidenced by the increase in concentration of TNPS in the hemolymph. This response takes place before the induction of enzyme activity, and it could serve as the initial signal in the induction pathway. PMID- 12115926 TI - Transport physiology of the urinary bladder in teleosts: a suitable model for renal urea handling? AB - The transport physiology of the urinary bladder of both the freshwater rainbow trout (Oncorhychus mykiss) and the marine gulf toadfish (Opsanus beta) was characterized with respect to urea, and the suitability of the urinary bladder as a model for renal urea handling was investigated. Through the use of the in vitro urinary bladder sac preparation urea handling was characterized under control conditions and in the presence of pharmacological agents traditionally used to characterize urea transport such as urea analogues (thiourea, acetamide), urea transport blockers (phloretin, amiloride), and hormonal stimulation (arginine vasotocin; AVT). Na(+)-dependence and temperature sensitivity were also investigated. Under control conditions, the in vitro trout bladder behaved as in vivo, demonstrating significant net reabsorption of Na(+), Cl(-), water, glucose, and urea. Bladder urea reabsorption was not affected by pharmacological agents and, in contrast to renal urea reabsorption, was not correlated to Na(+). However, the trout bladder showed a threefold greater urea permeability compared to artificial lipid bilayers, a prolonged phase transition with a lowered E(a) between 5 degrees C and 14 degrees C, and differential handling of urea and analogues, all suggesting the presence of a urea transport mechanism. The in vitro toadfish bladder did not behave as in vivo, showing significant net reabsorption of Na(+) but not of Cl(-), urea, or water. As in the trout bladder, pharmacological agents were ineffective. The toadfish bladder showed no differential transport of urea and analogues, consistent with a low permeability storage organ and intermittent urination. Our results, therefore, suggest the possibility of a urea transport mechanism in the urinary bladder of the rainbow trout but not the gulf toadfish. While the bladders may not be suitable models for renal urea handling, the habit of intermittent urination by ureotelic tetrapods and toadfish seems to have selected for a low permeability storage function in the urinary bladder. PMID- 12115927 TI - Ubiquitin and actin expression in claw muscles of land crab, Gecarcinus lateralis, and American lobster, Homarus americanus: differential expression of ubiquitin in two slow muscle fiber types during molt-induced atrophy. AB - The closer muscle of large-clawed decapod crustaceans undergoes a proecdysial (premolt) atrophy to facilitate withdrawal of the appendage at ecdysis. This atrophy involves the activation of both calcium-dependent (calpains) and ubiquitin (Ub)/proteasome-dependent proteolytic systems that break down proteins to reduce muscle mass. Moreover, the large slow-twitch (S(1)) fibers undergo a greater atrophy than the small slow-tonic (S(2)) fibers. Both polyUb mRNA and Ub protein conjugates increase during claw muscle atrophy. In this study in situ hybridization and RT-PCR were used to determine the temporal and spatial expression of polyUb and alpha-actin. A cDNA encoding the complete sequence of lobster muscle alpha-actin was characterized; a probe synthesized from the cDNA provided a positive control for optimizing RT-PCR and in situ hybridization. PolyUb was expressed at low levels in claw closer muscle from anecdysial (intermolt) land crab. By early proecdysis (premolt; stage D(0)), polyUb mRNA levels increased in medial fibers that insert along the midline of the apodeme, with greater expression in S(1) than S(2), while levels remained low in peripheral fibers. By late proecdysis, polyUb mRNA decreased in central fibers, while mRNA increased in peripheral S(1) fibers. In contrast, alpha-actin was expressed in lobster claw muscles at relatively constant levels during the intermolt cycle. These results suggest that Ub/proteasome-dependent proteolysis contributes to enhanced turnover of myofibrillar proteins during claw closer muscle atrophy. Furthermore, atrophy is not synchronous within the muscle; it begins in medial fibers and then progresses peripherally. PMID- 12115928 TI - Temporal and spatial localization of prothymosin alpha transcript in the Harderian gland of the frog, Rana esculenta. AB - The Harderian gland (hg) is the only orbital gland of the frog Rana esculenta, and it has the essential function of lubricating the eyes. The hg secretory activity is seasonal, showing the highest value in summer. There is, at present, no data on gene expression of the frog hg. This study reports, for the first time, on the temporal and spatial expression of a cDNA clone encoding for the prothymosin alpha (Prot-alpha), a highly acidic nuclear protein present in virtually all mammalian cells. Northern blot analysis revealed a single 1.7 kb transcript detected in the frog hg throughout the year, with a lowest expression in September in concomitance with the minimum secretory activity. In situ hybridization indicated that hg secretory cells express Prot-alpha transcript, and the hybridization signal was less intense in the September gland. The constant expression of the frog Prot-alpha mRNA during the whole year suggests a constitutive role for this molecule in the hg. In addition, taking into account that, in mammals, many immunomodulatory functions have been attributed to this protein, it is suggested that frog Prot-alpha might contribute to the hg immunity processes, probably acting as a protective agent against infections of the eyeball. Interestingly, although the presence of Prot-alpha gene in animals other than mammals has been considered to be highly unlikely, the present paper confirms the presence of Prot-alpha transcript in a nonmammalian vertebrate, the frog R. esculenta. PMID- 12115929 TI - Stress inhibits seasonal and FSH-induced ovarian recrudescence in the lizard,Mabuya carinata. AB - Stressors (handling, chasing, and noise) applied randomly five times per day for one month to lizards during the recrudescence phase of the ovarian cycle caused a significant reduction in mean number of oocytes and primordial follicles when compared to those of controls. Further, vitellogenic follicles were absent in the ovary of lizards subjected to stressors. Administration of bovine FSH during post breeding regression phase of the ovarian cycle induced ovarian recrudescence as shown by significant increases in the mean number of oogonia, oocytes, and primordial follicles compared to controls, as well as vitellogenic growth of follicles. However, lizards treated with FSH and exposed to stressors did not exhibit ovarian recrudescence. Furthermore, FSH administration during the post breeding regression phase caused a significant increase in serum levels of estradiol compared to controls, which was accompanied by significant increases in the relative weight of the liver and oviduct, as well as vitellogenic growth of follicles. Despite administration of FSH to lizards subjected to stressors, there was neither any increase in serum levels of estradiol and weight of the liver nor vitellogenic growth of follicles. The results indicate that repeated application of stressors inhibits vitellogenic growth of follicles by suppression of steroidogenic activity in M. carinata. This is the first report revealing that the ovary does not respond to gonadotrophin treatment under stressful conditions in reptiles. PMID- 12115930 TI - Influence of incubation temperature on hatching success, energy expenditure for embryonic development, and size and morphology of hatchlings in the oriental garden lizard, Calotes versicolor (Agamidae). AB - We incubated eggs of Calotes versicolor at four constant temperatures ranging from 24 degrees C to 33 degrees C to assess the effects of incubation temperature on hatching success, embryonic use of energy, and hatchling phenotypes that are likely to affect fitness. All viable eggs increased in mass throughout incubation due to absorption of water, and mass gain during incubation was dependent on initial egg mass and incubation temperature. The average duration of incubation at 24 degrees C, 27 degrees C, 30 degrees C, and 33 degrees C was 82.1 days, 60.5 days, 51.4 days, and 50.3 days, respectively. Incubation temperature affected hatching success, energy expenditure for embryonic development, and several hatchling traits examined, but it did not affect the sex ratio of hatchlings. Hatching success was lowest (3.4%) at 33 degrees C, but a higher incidence of deformed embryos was recorded from eggs incubated at this temperature compared to eggs incubated at lower temperatures. Most of the deformed embryos died at the last stage of incubation. Energy expenditure for embryonic development was, however, higher in eggs incubated at 33 degrees C than those similarly incubated at lower temperatures. A prolonged exposure of eggs of C. versicolor at 33 degrees C appears to have an adverse and presumably lethal effect on embryonic development. Hatching success at 24 degrees C was also low (43.3%), but hatchlings incubated at 24 degrees C did not differ in any of the examined traits from those incubated at two intermediate temperatures (27 degrees C and 30 degrees C). Hatchlings incubated at 33 degrees C were smaller (snout-vent length, SVL) than those incubated at lower incubation temperatures and had larger mass residuals (from the regression on SVL) as well as shorter head length, hindlimb length, tympanum diameter, and eye diameter relative to SVL. Hatchlings from 33 degrees C had significantly lower scores on the first axis of a principal component analysis representing mainly SVL-free head size (length and width) and fore- and hindlimb lengths, but they had significantly higher scores on the second axis mainly representing SVL-free wet body mass. Variation in the level of fluctuating asymmetry in eye diameter associated with incubation temperatures was quite high, and it was clearly consistent with the prediction that environmental stress associated with the highest incubation temperatures might produce the highest level of asymmetry. Newly emerged hatchlings exhibited sexual dimorphism in head width, with male hatchlings having larger head width than females. PMID- 12115932 TI - Influence of simulated microgravity on avian primordial germ cell migration and reproductive capacity. AB - Fertilized eggs of chicken and quail were incubated under the simulated microgravity condition provided by a clinostat. The number of Primordial Germ Cells (PGCs) was counted in early embryogenesis, and the reproductive capacity of quail hatched following the simulated microgravity was investigated. Simulated microgravity caused significant decline of PGCs in the blood of early chicken embryos and in the gonads. The numbers of spermatogonia in the hatchling testis were also fewer than those in the control groups. Therefore, simulated microgravity may retard gonadial development and reduce the reproductive capacity. PMID- 12115931 TI - Effects of inorganic and organic phosphate exposure on aspects of reproduction in the common sea urchin Lytechinus variegatus (Echinodermata: Echinoidea). AB - The common nearshore sea urchin Lytechinus variegatus is capable of surviving exposure to inorganic phosphate concentrations as high as 3.2 mg/l(-1) and organic phosphate concentrations of 1,000 mg/l(-1). However, chronic exposure to low, medium, or high sublethal concentrations of these phosphates inhibits gonadal tissue indices and spawning activity while altering biochemical composition of gonads, reducing size frequencies of oocyte diameters, and changing gonadal volume fractions. Gonad indices declined significantly in individuals maintained in all phosphate concentrations after both one- and two month exposures, while percentages of ripe individuals (oozing gametes upon dissection) were reduced after a two-month exposure in individuals maintained in medium and high organic phosphate concentrations. Levels of carbohydrates and lipids were lower in gonads of individuals maintained in all concentrations of both phosphates. Size frequency distributions of oocyte diameters revealed a dramatic decrease in oocyte size with increasing concentrations of both phosphates. Gonadal volume fractions of developing male and female gametes decreased with exposure to increasing phosphate levels. Volume fractions of nutritive phagocytes declined in testes of individuals held in the highest concentration of organic phosphate but displayed no significant change in ovaries. Volume fractions of mature gametes also decreased in gonads of individuals exposed to increasing concentrations of inorganic phosphate, but they remained constant in individuals exposed to all concentrations of organic phosphate. These findings indicate that shallow-water populations of L. variegatus subjected to inorganic and organic phosphate pollutants will exhibit stress that may impair reproductive output, gametogenesis, and spawning in the natural environment. PMID- 12115933 TI - Molecular sexing of monomorphic endangered Ara birds. AB - Survival of most endangered birds may depend on breeding programs where sex identification plays an important role. Molecular sexing has shown to be a rapid and safe procedure. In this work we established sex identification of monomorphic endangered Ara birds using a chromosome W-linked DNA marker, the Chromo-helicase DNA-Binding 1 (CHD) gene. Most birds have two CHD sex-linked genes, one W-linked (CHD-W) and one Z-linked (CHD-Z). These markers were characterized from Ara militaris and gender sex was determined by PCR and restriction analyzes. The procedure here reported was successfully applied to five different species of the genus Ara and confirmed the validity of the technique. To our knowledge, this is the first report of molecular sexing of the Ara species. This molecular sexing is currently been used in breeding programs of Ara birds. PMID- 12115934 TI - Preparation of metal ion buffers for biological experimentation: a methods approach with emphasis on iron and zinc. AB - Transition metal ions are a challenge to study in physiology because of problems associated with solubility, oxidation, binding, and attaining appropriate free activities in solution. This review discusses these problems and potential ways of accommodating them. Special attention is given to iron and zinc ions, but many of the concepts can be applied for studying other transition metals. Selection of reagents appropriate for metal work (including water, salts, noncomplexing pH buffers) is briefly discussed. Calculation of the solubility product (K(sp)) for common iron and zinc precipitates is covered, as well as techniques used to solubilize Fe(3+) with organic chelates. Factors that affect Fe(2+) oxidation are mentioned, and the use of ascorbate as a reducing agent is considered. Measurement of the rate of Fe(2+) oxidation (or Fe(3+) reduction) with the Fe(2+) chromophores ferrozine and BPS is also discussed. Generation of a free metal ion activity through use of metal buffers (chelators) is discussed. Theoretical problems associated with this technique are explored, and selected shareware metal ion buffer calculators are described. Finally, techniques for measuring and minimizing nonspecific binding of iron and zinc ions to biological membranes are considered. PMID- 12115935 TI - Pheromonal discriminations of sex, reproductive condition, and species by the lacertid lizard Podarcis hispanica. AB - In some vertebrate taxa, pheromones provide important information about species, sex, reproductive condition, kinship, and even individual identity. Because they possess highly developed nasal chemosensory systems, lizards are capable of many chemical discriminations, but many aspects of their pheromonal communication remain poorly understood even in major families. We report that males of a lacertid lizard, Podarcis muralis, are capable of differential response to surface chemical cues from conspecific males and females, from gravid and nongravid females, and from conspecific females and females of the closely related sympatric congener, P. bocagei carbonelli. In 60 sec trials in which stimuli from the femoral, cloacal, lateral, and upper body surfaces were presented to males on cotton swabs, males tongue-flicked at significantly higher rates to stimuli from conspecific females than males, from conspecific nongravid than gravid females, and from conspecific than heterospecific females. Responses to stimuli from conspecific males did not differ from those to distilled water. Together with previous findings that males can distinguish between chemical cues from familiar and unfamiliar males, these findings suggest that pheromones provide male P. hispanica important information regarding the presence of sexual rivals and the reproductive condition of potential mates without visually encountering other lizards. A growing body of literature indicates that lacertids are capable of sophisticated pheromonal discriminations that may play important roles in their social behavior. PMID- 12115936 TI - Regeneration in planarians and other worms: New findings, new tools, and new perspectives. AB - Molecular biology, recombinant DNA techniques, and new methods of cell lineage have reignited the interest of planarians and other worms (mainly annelids and nemerteans) as invertebrate model systems of regeneration. Here, the mean results produced in the last five years are reviewed, an update of the genes and molecules involved in planarian regeneration is provided, and a new morphallactic epimorphic model of pattern formation is suggested. Moreover, and most importantly, we highlight the new strides brought upon by genomic/proteomic analyses, RNA interference (RNAi) to inactivate gene function, and Bromodeoxyuridine (BrdU) cell labelling. The raising hope to obtain transformed neoblasts and transgenic planarians is also stressed. Altogether, such approaches will eventually lead to solve the long-standing open questions on regeneration which still baffles us. Finally, we warn against overlooking the evident links between regeneration processes and those controlling the daily wear and tear of tissues and cells. Both processes act, at least in planarians, upon a unique stem cell endowed with an unrivaled developmental potential in the animal kingdom-the neoblast. This cell could be considered the forebear and a model system for stem cell analysis. PMID- 12115937 TI - Expression and biological effect of urodele fibroblast growth factor 1: relationship to limb regeneration. AB - Fibroblast growth factors (FGFs) have been previously implicated in urodele limb regeneration. Here, we examined expression of FGF-1 by blastema cells and neurons and investigated its involvement in wound epithelial formation and function and in the trophic effect of nerves. Neurons innervating the limb and blastema cells in vivo and in vitro expressed the FGF-1 gene. The peptide was present in blastemas in vivo. Wound epithelium thickened when recombinant newt FGF-1 was provided on heparin-coated beads, demonstrating that the FGF-1 was biologically active and that the wound epithelium is a possible target tissue of FGF. FGF-1 did not stimulate accessory limb formation. FGF-1 was as effective as 10% fetal bovine serum in maintaining proliferative activity of blastema cells in vitro but was unable to maintain growth of denervated, nerve-dependent stage blastemas when provided on beads or by injection. FGF-1 had a strong stimulating effect on blastema cell accumulation and proliferation of limbs inserted into the body cavity that were devoid of an apical epithelial cap (AEC). These results show that FGF-1 can signal wound epithelium cap formation and/or function and can stimulate mesenchyme accumulation/proliferation in the absence of the AEC but that FGF-1 is not directly involved in the neural effect on blastema growth. PMID- 12115938 TI - Physiological changes in the spawning gilthead seabream, Sparus aurata, succeeding the removal of males. AB - The physiological effects triggered in females by the removal of males from a group of spawning fish were examined in the multiple batch spawner, the gilthead seabream, Sparus aurata. One week after the removal of males, a large portion of the oocytes underwent atresia, and sporadic release of low quality eggs continued at low frequency over a period of seven weeks. The transcript levels of the three native gonadotropin releasing hormone (GnRH) forms, salmon (s)GnRH, seabream (sb)GnRH, and chicken (c)GnRH-II, and the two beta GtH subunits were measured. Brain mRNA levels for all three GnRHs and pituitary beta LH mRNA levels significantly declined in the females as a result of removing the males compared to females that were maintained with males. Pituitary beta FSH mRNA levels showed the opposite trend and were significantly higher in females that were separated from males. Circulating levels of LH, testosterone, estradiol, 17 alpha, 20 beta dihydroxy-4-pregnen-3-one, and 17 alpha, 20 beta,21-trihydroxy-4-pregnen-3-one all declined in the group of females without males. These results imply the existence of an endocrine response to socio-sexual stimuli during the reproductive process in the gilthead seabream. PMID- 12115939 TI - High performance liquid chromatographic separation of steroids from ovarian follicles of fresh water perch Anabas testudineus: identification and characterization of the maturation-inducing hormone. AB - Accurate separation and identification of steroids from the postvitellogenic ovarian follicles of Indian climbing perch Anabas testudineus was performed by high-performance liquid chromatography (HPLC) and specific radioimmunoassay (RIA). The steroids from such follicles, incubated in Cortland's saline with or without homologous fish pituitary extract (FPE), were extracted with dichloromethane and separated on a micro Bondapak C(18) column. Identification of the HPLC fractions was further confirmed by thin layer chromatography. As HPLC peaks for 17 alpha, 20 beta-dihydroxy-4-pregnen-3-one (DHP) and testosterone (T) were close, clear separation of these steroids and accurate measurement of their quantities was achieved by RIA of HPLC fractions using specific antibodies. Altogether, nine eluted fractions in the FPE-untreated and ten in FPE-treated samples were obtained. Of these, six were identified as: 5 beta-pregnan-3 alpha,17 alpha,20 beta-triol (5 beta-3 alpha,17 alpha,20 beta-P); DHP; T; 17 alpha-hydroxyprogesterone (17 alpha-P(4)); progesterone (P(4)); and androstenedione (AD). Three fractions from untreated and four from FPE-treated samples, however, remained unidentified. Of all the HPLC fractions examined for their relative maturational inducing (MI) potency on full grown (postvitellogenic) ovarian follicles of perch, the fraction identified as DHP was found to be the most effective inducer of germinal vesicle breakdown (GVBD) both at low and high concentrations. Fractions identified as 5 beta-3 alpha, 17 alpha, 20 beta-P and 17alpha-P(4) could induce only 32% and 20% GVBD at their highest concentration, while none of the unidentified fractions showed any MI activity. FPE caused increased production of DHP, testosterone, and 5 beta-3 alpha, 17 alpha, 20 beta-P. The qualitative differences between the fractions obtained from FPE-treated samples and those from FPE-untreated samples were only the appearance of a new polar metabolite of unknown function. The present study showed that, as a single steroid, DHP was the most potent MIH for A. testudineus. PMID- 12115940 TI - Temperature sex determination in the European sea bass, Dicentrarchus labrax (L., 1758) (Teleostei, Perciformes, Moronidae): critical sensitive ontogenetic phase. AB - The temperature sex determination (TSD) mechanism in the European sea bass (Dicentrarchus labrax L.) was studied in respect to: a) the TSD sensitivity during the different developmental stages; and b) the intrapopulation correlation of sex determination with the growth rate up to the end of the TSD-sensitive period. At the stage of half-epiboly, eggs from the same batch were divided into four groups and subjected to different thermal treatments: a) 15 degrees C (G15 group) and b) 20 degrees C (G20 group) up to the middle of metamorphosis stage; c) 15 degrees C up to the end of yolk-sac larval stage and subsequently to 20 degrees C (G15-5 group); and d) 15 degrees C up to the end of the preflexion stage and then to 20 degrees C (G15-10 group). At the end of the treatments, size grading was applied and four additional populations were established from the upper (L) and lower (S) size portions of the G15 and G20 populations: G15L, G15S, G20L, and G20S. During the following growing phase, all populations were subjected to common rearing conditions. The sex ratios of each population were macroscopically determined at 190-210 mm mean total length. Female incidence was significantly affected (P < 0.05) by the different thermal treatments: 66.1% in the G15, 47.1% in the G15-10, 37.6% in the G15-5, and 18.1% in the G20 group. In addition, sex ratio was correlated with the growth rate of the fish up to the end of the TSD-sensitive period, with the larger fish presenting a significantly higher (P < 0.01) female incidence than the smaller fish in both thermal regimes tested: 73.1% in G15L vs. 57% in G15S, and 36.6% in G20L vs. 22.5% in G20S group. Results provide, for the first time, clear evidence that the sea bass is sensitive to TSD during all different ontogenetic stages up to metamorphosis, and that sex ratio is correlated with the growth rate of the fish well before the differentiation and maturation of the gonads. PMID- 12115942 TI - Role of cyclic AMP-dependent protein kinase in oocyte maturation of the catfish, Clarias batrachus. AB - The results of the present study demonstrate the probable involvement of adenosine 3',5'-cyclic monophosphate (cAMP)-dependent protein kinase (PKA) in the regulation of oocyte maturation in the catfish, Clarias batrachus. A decrease in total PKA activity with a concomitant increase in the percentage of germinal vesicle breakdown (GVBD) was found in oocytes treated with different doses of N (2-[p-bromocinnamylamino]ethyl)-5-isoquinoline sulfonamide (H-89), a selective, potent inhibitor of PKA and 17 alpha, 20 beta-dihydroxy-4-pregnen-3-one (17 alpha, 20 beta-DP), the natural maturation-inducing steroid of this catfish. Evaluation of time-course of response to H-89 and 17 alpha, 20 beta-DP revealed that PKA activity decreased, and incidence of GVBD increased at all the time points when compared with their respective controls. The data further indicate that the decrease in PKA activity in H-89-treated oocytes was more prominent, but the induction of maturation was slower than that induced by 17 alpha, 20 beta-DP. Moreover, cyanoketone (CK), an inhibitor of steroidogenesis that blocks the salmon gonadotropin (SG-G100)-induced GVBD, failed to abolish the maturational effect of H-89, suggesting that H-89 directly promotes GVBD by reducing PKA activity in oocytes. Taken together, these results indicate that inhibition of PKA activity in the oocyte of C. batrachus is directly involved in the mechanism leading to oocyte maturation. PMID- 12115941 TI - Development of embryos from in vitro ovulated and fertilized oocytes of the quail (Coturnix coturnix japonica). AB - The development of quail embryos obtained after in vitro fertilization of oocytes ovulated in vitro was investigated. About 40% of the specimens, after 18-20 hr of incubation, had undergone cleavage to reach stages IV-VI when viewed under a stereo microscope. However, only 36% of these embryos contained normal, DAPI stained nuclei when observed under a fluorescent microscope; the other 64% showing a morphologically normal cleavage pattern did not contain nuclei. Control unfertilized oocytes, ovulated in vitro and cultured for the same time, also sometimes attained the morphologically correct stages IV-VI but their "blastomeres" were always devoid of nuclei. Therefore, it is advisable to monitor early avian embryos for the presence of nuclei when assessing development in culture. The results demonstrate, for the first time, that cytoplasmic segmentation can occur in the absence of nuclear divisions in the germinal disc of the quail and show the existence and significance of ooplasmic maternal information in birds. This phenomenon is also known for sea urchin and frogs. It is indicative of the role of maternal information in early development. The in vitro method presented here links the steps of ovulation and fertilization with the early cleavage stages under in vitro conditions and may be useful in studying mechanisms of fertilization and differentiation in birds as well as in obtaining transgenic birds by DNA injection or application of foreign, DNA-carrying sperm. PMID- 12115943 TI - "Crisis" in myasthenia gravis: an historical perspective. PMID- 12115945 TI - Brainstem reflexes: electrodiagnostic techniques, physiology, normative data, and clinical applications. AB - An overview is provided on the physiological aspects of the brainstem reflexes as they can be examined by use of clinically applicable neurophysiological tests. Brainstem reflex studies provide important information about the afferent and efferent pathways and are excellent physiological tools for the assessment of cranial nerve nuclei and the functional integrity of suprasegmental structures. In this review, the blink reflex after trigeminal and nontrigeminal inputs, corneal reflex, levator palpebrae inhibitory reflex, jaw jerk, masseter inhibitory reflex, and corneomandibular reflex are discussed. Following description of the recording technique, physiology, central pathways, and normative data of these reflexes, including an account of the recording of recovery curves, the application of these reflexes is reviewed in patients with various neurological abnormalities, including trigeminal pain and neuralgia, facial neuropathy, and brainstem and hemispherical lesions. Finally, simultaneous electromyographic recording from the orbicularis oculi and the levator palpebrae muscles is discussed briefly in different eyelid movement disorders. PMID- 12115944 TI - Spinal muscular atrophy: recent advances and future prospects. AB - Spinal muscular atrophies (SMA) are characterized by degeneration of lower motor neurons associated with muscle paralysis and atrophy. Childhood SMA is a frequent recessive autosomal disorder and represents one of the most common genetic causes of death in childhood. Mutations of the SMN1 gene are responsible for SMA. The knowledge of the genetic basis of SMA, a better understanding of SMN function, and the recent generation of SMA mouse models represent major advances in the field of SMA. These are starting points towards understanding the pathophysiology of SMA and developing therapeutic strategies for this devastating neurodegenerative disease, for which no curative treatment is known so far. PMID- 12115946 TI - Use of intravenous pulsed cyclophosphamide in severe, generalized myasthenia gravis. AB - Twenty-three myasthenia gravis (MG) subjects, mean (SD) age 41.6 years (14), showing poor disease control or steroid-related side effects, received treatment for 12 months with intravenous cyclophosphamide (CP; n = 12) or placebo (PL; n = 11) in a randomized, double-blind trial. Pulses were given monthly at an initial dose of 500 mg/m(2) of body surface, and titrated according to changes of peripheral muscle strength or side effects. Changes of muscle strength, steroid and pyridostigmine requirements, and development of ventilatory failure or swallowing impairment were evaluated at 0, 3, 6, and 12 months. No differences were observed between groups at baseline. Statistically significant reductions of methylprednisone doses were noted in both groups but were more pronounced in subjects receiving CP than PL at 6 months (P < 0.05) and at 12 months (P < 0.03). At 12 months, five subjects on CP had tapered off their steroids whereas no subject on PL achieved further reductions (P < 0.03). Four CP subjects were not receiving steroids 36 months after completing the study and three other CP subjects had stopped pyridostigmine. CP improved muscle strength at 3 and 6 months, and this reached statistical significance compared to PL at 12 months mainly in the bulbar and masticatory (P < 0.009) and extraocular muscles (P < 0.03). Ventilatory failure was noted in one subject on CP (due to bronchopneumonia) and two on PL (due to muscle weakness). No significant increases of CP-related side effects were observed. Thus, this study suggests that intravenous pulses of CP allow reductions of systemic steroids usage without muscle strength deterioration or CP-related side effects. PMID- 12115947 TI - New evidence for a presynaptic action of prednisolone at neuromuscular junctions. AB - The action of prednisolone at the neuromuscular junction was studied in mouse isolated phrenic nerve-diaphragm and rat external popliteal/sciatic nerve tibialis anterior muscle preparations. Prednisolone (0.03 mM and 0.3 mM) did not alter the twitch-tension in phrenic nerve-diaphragm preparations after 120 min, but increased the frequency (170 +/- 4%) and amplitude (200 +/- 13%) of miniature end-plate potentials. Quantal content was not influenced by the glucocorticoid treatment. Prednisolone (400 microg/kg) did not change the twitch-tension in rat external popliteal/sciatic nerve-tibialis anterior muscle preparations. However, this steroid (0.3 mM) prevented the neuromuscular blockade by d-tubocurarine (1.45 microM) in mouse preparations by 70 +/- 10% (P < 0.05). A similar effect (82 +/- 6% protection, P < 0.05) occurred in rats treated with prednisolone (400 microg/kg) before d-tubocurarine (225 microg/kg). In phrenic nerve-diaphragm preparations, prednisolone (0.3 mM) increased (13 +/- 4%, p < 0.05) the twitch tension in the presence of beta-bungarotoxin (1 microM), and prevented the blockade produced by this toxin (0.15 microM) in its third phase of action. This presynaptic facilitatory effect may contribute to the usefulness of prednisolone in myasthenia gravis. PMID- 12115948 TI - Modality-related scalp responses after electrical stimulation of cutaneous and muscular upper limb afferents in humans. AB - To elucidate whether the selective electrical stimulation of muscle as well as cutaneous afferents evokes modality-specific responses in somatosensory evoked potentials (SEPs) recorded on the scalp of humans, we compared scalp SEPs to electrical stimuli applied to the median nerve and to the abductor pollicis brevis (APB) motor point. In three subjects, we also recorded SEPs after stimulation of the distal phalanx of the thumb, which selectively involved cutaneous afferents. Motor point and median nerve SEPs showed the same scalp distribution; moreover, very similar dipole models, showing the same dipolar time courses, explained well the SEPs after both types of stimulation. Since the non natural stimulation of muscle afferents evokes responses also in areas specifically devoted to cutaneous input processing, it is conceivable that, in physiological conditions, muscle afferents are differentially gated in somatosensory cortex. The frontocentral N30 response was absent after purely cutaneous stimulation; by contrast, it was relatively more represented in motor point rather than in mixed nerve SEPs. These data suggest that the N30 response is specifically evoked by proprioceptive inputs. PMID- 12115949 TI - IGF-I, IGF-II, and IGF-receptor-1 transcript and IGF-II protein expression in myostatin knockout mice tissues. AB - Semiquantitative reverse transcriptase-polymerase chain reaction (RT-PCR) and immunohistochemistry were performed to demonstrate whether a correlation exists between insulin-like growth factors (IGFs)-positive regulators of growth-and myostatin, a negative regulator of muscle growth. IGF-I, -II, and IGF-receptor-1 (IGF-R1) mRNA and IGF-II protein expressions were determined in control and myostatin knockout mice tissues. IGF-I gene expressions were similar between control and knockout mice tissues, whereas IGF-II mRNA levels were significantly higher in myostatin knockout mice kidney and soleus muscles than those of control mice (P <.01). IGF-R1 mRNA levels from control mice heart (P <.05) and kidney (P <.01) were significantly higher than in myostatin knockout mice, whereas levels were lower in pectoralis muscle of control mice than knockout mice (P <.01). The strongly IGF-II-positive cells in soleus muscle were more common in myostatin knockout mice and were seen in a few foci in control mice. IGF-II immunoreactivity in both control and myostatin knockout mice kidneys was localized to the epithelium of renal tubules and collecting ducts. Reciprocal changes in the expression of myostatin and IGF-II and IGF-R1 may underlie normal growth of skeletal muscle and other organs in mammals, and the changes in these tissues associated with disease. PMID- 12115950 TI - Reliability of hand-held dynamometry in spinal muscular atrophy. AB - We have assessed the reliability of hand-held myometry in 33 patients with spinal muscular atrophy (SMA), testing elbow flexion, handgrip, three-point pinch, knee flexion, knee extension, and foot dorsiflexion, and determining intraclass correlation coefficients (ICC). Interrater reliability was high for upper limbs, with an ICC of 0.92 for three-point pinch and 0.98 for elbow flexion and grip. For lower limbs interrater reliability was good with ICC >0.85 for all measures except foot dorsiflexion. Test-retest results were excellent with ICC >0.91 in all instances. Hand-held myometry is easily performed in SMA patients of various ages and muscle strengths, is a reliable measure of limb muscle strength, and can be used in longitudinal studies and clinical trials. PMID- 12115951 TI - Improvement of dy/dy dystrophic diaphragm by 3,4-diaminopyridine. AB - Concerns have been raised that inotropic agents may worsen function of dystrophic muscle due to structural fragility. Studies tested the hypothesis that force increments elicited by potassium (K(+)) channel blockade can be maintained during the course of repetitive stimulation. In vitro twitch force of dy/dy dystrophic mouse diaphragm was significantly lower than normal (796 versus 1271 g/cm(2)). 3,4-Diaminopyridine (DAP) increased twitch force of dystrophic diaphragm by 111 +/- 12% (P <.0001) and increased force at stimulation frequencies of 5-50 Hz by 41-77%. During fatigue-inducing stimulation, force augmentation by DAP was well maintained in dystrophic muscle throughout 25 Hz (P =.0047) and 50 Hz (P =.0059) stimulation. These findings indicate that the K(+) channel blocker DAP augments the force of dystrophic muscle to values close to that of normal muscle over a range of stimulation frequencies. Furthermore, these functional increments can be achieved without causing force to eventually deteriorate below that of untreated dystrophic muscle during fatiguing stimulation. It is possible that DAP may be useful for the clinical management of a variety of disorders causing muscle weakness. PMID- 12115952 TI - Macrophage apoptosis in rat skeletal muscle treated with bupivacaine hydrochloride: possible role of MCP-1. AB - The fate of macrophages infiltrating damaged rat skeletal muscle fibers after intramuscular injection of the anesthetic bupivacaine hydrochloride (BPVC) and the possible roles of monocyte chemoattractant protein-1 (MCP-1) were investigated. The number of macrophages reached a maximum level at 2 days after the injection and then gradually decreased. The number of apoptotic cells detected by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) assay was elevated at 2-4 days and decreased thereafter. In serial sections, TUNEL-positive cells were also immunopositive for RM-4, an antibody specific for identification of macrophages. Electron microscopy further confirmed that it was the macrophages themselves that were undergoing apoptosis. As assessed by reverse transcriptase-polymerase chain reaction (RT-PCR), high levels of MCP-1 mRNA in BPVC-treated muscles were observed and positively correlated with maximum macrophage infiltration. However, the levels of MCP-1 mRNA returned to normal low values coincident with decrease of inflammation and healing of damaged muscle fiber. Local apoptosis of macrophages, under the control of MCP-1, may be involved in healing of BPVC-treated muscles. PMID- 12115953 TI - Influence of insulin-like growth factor-I (IGF-I) on nerve autografts and tissue engineered nerve grafts. AB - To overcome the problems of limited donor nerves for nerve reconstruction, we established nerve grafts made from cultured Schwann cells and basal lamina from acellular muscle and used them to bridge a 2-cm defect of the rat sciatic nerve. Due to their basal lamina and to viable Schwann cells, these grafts allow regeneration that is comparable to autologous nerve grafts. In order to enhance regeneration, insulin-like growth factor (IGF-I) was locally applied via osmotic pumps. Autologous nerve grafts with and without IGF-I served as controls. Muscle weight ratio was significantly increased in the autograft group treated with IGF I compared to the group with no treatment; no effect was evident in the tissue engineered grafts. Autografts with IGF-I application revealed a significantly increased axon count and an improved g-ratio as indicator for "maturity" of axons compared to autografts without IGF-I. IGF-I application to the engineered grafts resulted in a decreased axon count compared to grafts without IGF-I. The g-ratio, however, revealed no significant difference between the groups. Local administration of IGF-I improves axonal regeneration in regular nerve grafts, but not in tissue-engineered grafts. Seemingly, in these grafts the interactive feedback mechanisms of neuron, glial cell, and extracellular matrix are not established, and IGF-I cannot exert its action as a pleiotrophic signal. PMID- 12115954 TI - Mechanisms of force failure during repetitive maximal efforts in a human upper airway muscle. AB - The upper airway respiratory muscles play an important role in the regulation of airway resistance, but surprisingly little is known about their contractile properties and endurance performance. We developed a technique that allows measurement of force and the electromyogram (EMG) of human nasal dilator muscles (NDMs). Endurance performance was quantified by measuring NDM "flaring" force and EMG activity as healthy human subjects performed 10 s maximal voluntary contractions (MVCs), separated by 10 s rest, until the area under the force curve fell to 50% MVC (the time limit of the fatigue task, Tlim), which was reached in 34.2 +/- 3.1 contractions (685.0 +/- 62.3 s). EMG activity was unchanged except at Tlim, where it averaged 78.7 +/- 3.6% of pretest activity (P < 0.01). M-wave amplitude did not change, suggesting that neuromuscular propagation was not impaired. MVC force increased to 80% of the pretest level within 10 min of recovery but twitch force failed to recover, suggesting low-frequency fatigue. The data suggest that a failure of the nervous system to excite muscle could explain at most only a small fraction of the NDM force loss during an intermittent fatigue task, and then only at Tlim. Thus, the majority of the force failure during this task is due to impairment of mechanisms that reside within the muscle fibers. PMID- 12115955 TI - Relative distribution of three major lactate transporters in frozen human tissues and their localization in unfixed skeletal muscle. AB - We have prepared affinity-purified rabbit polyclonal antibodies to the near-C terminal peptides of human monocarboxylate transporters (MCTs) 1, 2, and 4 coupled to keyhole limpet hemocyanin. Each antiserum reacted only with its specific peptide antigen and gave a distinct molecular weight band (blocked by preincubation with antigen) after chemiluminescence reaction on Western blots from sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) of tissue membrane proteins. Densitometry showed distinctive expression patterns for each MCT in a panel of 15 frozen human tissues, with the distribution of MCT1 >>MCT2>MCT4. Fluorescence microscopy of unfixed skeletal muscle using fluorescein conjugated secondary antibody was correlated with reverse adenosine triphosphatase (ATPase) stained sequential sections to identify fiber-type localization. MCT1 expression was high in the sarcolemma of type 1 fibers, modest to low in type 2a fibers, and almost absent in type 2b fibers. In contrast, MCT4 expression was low to absent in the membrane of most type 1 fibers, but high in most 2a and in all 2b fibers, favoring the view that their high lactate levels during work may be channeled in part to neighboring type 1 (and perhaps 2a) fibers for oxidation, thereby delaying fatigue. MCT2 expression was limited to the sarcolemma of a type 1 fiber subset, which varied from <5 to 40%, depending on the specific muscle under study. Quantitative chemiluminescent densitometry of 10 muscle biopsies for their MCT2 and MCT4 content, each normalized to MCT1, confirmed the unique variation of MCT2 expression with biopsy site. The application of these antibodies should add to the understanding of motor unit physiology, and may contribute to the muscle-biopsy assessment of low-level denervation. PMID- 12115956 TI - Effect of sympathetic muscle vasoconstrictor activity on capsaicin-induced muscle pain. AB - Neuropathic pain syndromes with sympathetically maintained pain are often associated with a deep somatic pain component. An adrenergic interaction between sympathetic vasoconstrictor neurons and cutaneous afferents has been demonstrated. To determine whether a sympathetic-afferent interaction exists in deep somatic tissues, we investigated the effect of sympathetic muscle vasoconstrictor activity on experimentally induced pain. In 12 healthy volunteers, capsaicin was infused into the anterior tibial muscle. Intensity and quality of muscle and referred pain were assessed. The analyses were performed during the presence of low sympathetic muscle vasoconstrictor activity induced by breathing 100% O(2) gas (normocapnia), and during high activity induced by inspiration of 95% O(2) and 5% CO(2) (hypercapnia). The degree of sympathetic muscle vasoconstrictor discharge was monitored indirectly by measuring systemic blood pressure and end-expiratory CO(2) and by performing duplex sonography of muscle resistance vessels. The intensity, quality, and spatial distribution of muscle and referred pain were not significantly different during resting and increased sympathetic muscle vasoconstrictor discharge, indicating that such activity does not influence pain after intramuscular infusion of capsaicin. PMID- 12115958 TI - Partial conduction blocks in N-hexane neuropathy. AB - Nerve conduction blocks, defined by a significant reduction in amplitude or area of the compound muscle action potential at proximal compared with distal sites of stimulation, have been described in glue-sniffers and in workers with industrial exposure at an early stage of n-hexane neuropathy. The frequency with which this focal conduction anomaly appears is described and discussed in the case of a very homogeneous group of 10 young workers diagnosed with n-hexane polyneuropathy. Partial conduction blocks occurred in only two workers and may have been related to the intensity and duration of toxic exposure. PMID- 12115957 TI - Differential fatigue of paralyzed thenar muscles by stimuli of different intensities. AB - Muscles paralyzed by injury or disease fatigue excessively when stimulated. This study examined whether the first few paralyzed thenar motor units recruited by electrical stimulation of the median nerve were more fatigue resistant than the total thenar motor unit population. The paralyzed thenar muscles of four subjects with chronic cervical spinal cord injury were fatigued by a 2-min intermittent 40 HZ protocol on 2 days. One experiment involved submaximal stimulation, the other supramaximal stimulation. These stimuli resulted in activation of part and all of the thenar muscles, respectively. Relative force loss, force-time integral decline, and slowing of half-relaxation time were always significantly less when only part rather than all of the muscles was fatigued. The part of the paralyzed muscles that was activated was also relatively fatigue resistant compared with control single thenar motor units. Thus, a reversal of recruitment order from fatigable to fatigue-resistant units cannot explain the extreme fatigability of paralyzed muscles. Use of submaximal stimulation during functional electrical stimulation may therefore help to reduce muscle fatigue because it recruits the more fatigue-resistant units. PMID- 12115959 TI - High efficiency transfection of primary skeletal muscle cells with lipid-based reagents. AB - Lipofection is a convenient method for gene transfer into muscle cells but reportedly is inefficient. We tested the efficacy of commercially available lipid based and polyamine transfection reagents. Primary rat skeletal muscle cell cultures were transfected at three stages of development and assayed after fusion. Efficiency reached 30% during the proliferation stage and up to 23% when most myoblasts had fused into myotubes. Optimization of transfection conditions with three different vectors yielded efficiencies exceeding 50%. Thus, lipid based transfection into primary skeletal muscle cells can be several times more efficient than previously reported. PMID- 12115960 TI - Hypothyroid myopathy with a strikingly elevated serum creatine kinase level. AB - Although serum creatine kinase (CK) levels are frequently modestly elevated in patients with hypothyroid myopathy, elevations in serum CK to the levels usually seen in inflammatory myopathies or dystrophies are rare. We report a patient with progressive proximal weakness and a serum CK level of over 29,000 IU/L, in whom subsequent laboratory evaluation identified profound hypothyroidism. Thyroid hormone replacement therapy resulted in resolution of clinical symptoms and a marked reduction in the serum CK level. Such a high serum CK level in a patient with hypothyroidism underscores the importance of assessing thyroid function in patients with weakness, regardless of serum CK levels, even when systemic symptoms and signs of hypothyroidism are minimal or absent. PMID- 12115961 TI - Recovery from distal ulnar motor conduction block injury: serial EMG studies. AB - Acute conduction block injuries often result from nerve compression or trauma. The temporal pattern of clinical, electrophysiologic, and histopathologic changes following these injuries has been extensively studied in experimental animal models but not in humans. Our recent evaluation of a young man with an injury to the deep motor branch of the ulnar nerve following nerve compression from weightlifting exercises provided the opportunity to follow the course and recovery of a severe conduction block injury with sequential nerve conduction studies. The conduction block slowly and completely resolved, as did the clinical deficit, over a 14-week period. The reduction in conduction block occurred at a linear rate of -6.1% per week. PMID- 12115963 TI - Activation order of motor axons in electrically evoked contractions. PMID- 12115964 TI - How selective is the reinnervation of skeletal muscle fibers? PMID- 12115965 TI - Neuromuscular diseases and disorders of the alimentary system. AB - This review outlines the relationship and interaction between neuromuscular diseases and disorders of the alimentary system. Neuromuscular manifestations of gastrointestinal and hepatobiliary diseases are first considered. Such diseases may cause neuromuscular disorders by leading to nutritional deficiency or by more direct mechanisms. The pathogenesis, clinical features, and treatment of these various neuromuscular manifestations are discussed. The impact of disorders of nerve, neuromuscular transmission, and muscle on the alimentary system is then reviewed. The main sequelae are impaired deglutition and gastrointestinal dysmotility. The management of these complications is considered. PMID- 12115966 TI - Neuromuscular fatigue and aging: central and peripheral factors. AB - A limited number of studies have investigated the effect of old age on neuromuscular fatigue, yet a variety of protocols have been used to compare the fatigability of old and young humans. These include voluntary isometric and isokinetic contraction protocols at maximal and submaximal intensities, and electrical stimulation protocols of continuous or intermittent stimulation at a variety of stimulation frequencies. The results of these studies are summarized in this review. Although it seems reasonable to suggest that age-related changes in muscle morphology and motor unit remodeling, as well as the associated loss of strength and slowed contractile properties, may improve the resistance to neuromuscular fatigue in old humans, the collective results suggest that it is not possible to make this generalization. In fact, it cannot be generalized that the muscles of old humans are either more or less fatigable than young adults because the extent of the difference in fatigability relies strongly on the fatigue task performed (task-dependency). Age-related changes that occur within the neuromuscular system may result in some candidate fatigue sites increasing or decreasing their susceptibility to failure under specific task conditions. These candidate fatigue sites include central drive, muscle membrane excitability, excitation-contraction coupling mechanisms, and metabolic capacities. The effect of old age on these various central and peripheral sites is discussed with respect to their relative contribution during different fatigue tasks. Moreover, the impact of the possible confounding effects of subject habituation, physical activity status, and sex on the fatigability comparison is addressed. PMID- 12115967 TI - Motor unit activation order during electrically evoked contractions of paralyzed or partially paralyzed muscles. AB - The activation order of motor units during electrically evoked contractions of paralyzed or partially paralyzed thenar muscles was determined in seven subjects with chronic cervical spinal cord injury. The median nerve was stimulated percutaneously with pulses of graded intensity to produce increments in the compound electromyogram (EMG) and force. Each increment corresponded to the activation of another unit. The evoked unit EMG and force was obtained by digital subtraction. The thenar muscles had between 15 and 83 units (26 +/- 19) that produced 114.3 +/- 127.1 mN force (n = 290). In six subjects, a significant positive correlation was found between activation order and unit force indicating that weaker units were excited before stronger units. These data are contrary to the notion that a reversal of unit activation order occurs during evoked versus voluntary contractions. PMID- 12115968 TI - Regional distribution of slow-twitch muscle fibers after reinnervation in adult rat hindlimb muscles. AB - In adult rats, the sciatic nerve was unilaterally sectioned and reunited above the knee. Following a survival time of 21 weeks, five muscles were removed from both lower hindlimbs after determining their intra-limb positions. In each muscle, cryostat sections from seven equidistant proximo-distal levels were stained for myofibrillar ATPase. Intramuscular positions were determined for all slow-twitch type I fibers. Within each muscle, type I fibers were heterogeneously distributed, and the direction of type I fiber accumulation was, on average, almost identical in reinnervated muscles and contralateral controls. Furthermore, as in controls, a proximo-distal decline of type I fiber density was found in reinnervated muscles. Compared to contralateral controls, reinnervated muscles consistently showed a very high number of type I fibers at close interfiber distances, indicating respecification of muscle fiber types by the ingrowing nerve fibers. The results suggest that slow-twitch motor axons preferentially grew back toward the original slow-twitch muscle regions. PMID- 12115969 TI - Effect of reference electrode position on the compound muscle action potential (CMAP) onset latency. AB - Compound muscle action potential (CMAP) onset latency is interpreted to reflect the arrival time at the muscle of impulses in the fastest-conducting motor nerve fiber. However, we have observed that the position of the reference or indifferent electrode (E2) affects CMAP onset latency. Motor nerve conduction studies (NCS) of the median, ulnar, and deep ulnar motor (DUM) nerves on 20 normal hands were performed using both traditional bipolar and experimental monopolar (referenced to the contralateral hand) montages. As the position of E2 was altered, the CMAP onset latency varied 0.1-0.5 ms for the median NCS, 0.1-0.3 ms for the ulnar NCS, and 0.1-1.5 ms for the DUM NCS. This study demonstrates that E2 recorded potentials are significant and vary with positioning, affecting motor onset latency. This has implications both for reference values and the physiologic interpretation of the CMAP waveform. PMID- 12115970 TI - Impact of initial muscle length on force deficit following lengthening contractions in mammalian skeletal muscle. AB - The purpose of this study was to determine whether initial muscle length influenced the extent of isometric force deficit following 20 in vitro lengthening contractions of the soleus muscle from Fischer 344 rats. Force deficit was evaluated following one of five protocols: (1) lengthening contractions from optimal muscle length (Lo) to 120% Lo; (2) lengthening contractions from 80% Lo to Lo; (3) lengthening contractions from Lo to 120% Lo but with a stimulation frequency that elicited the same force as protocol 2; (4) 20 isometric contractions at Lo; (5) 20 stretches +/- 20% Lo in inactive muscle. Following lengthening contractions, extent of force deficit significantly differed between protocols 1, 2, and 3 (P < 0.05). Maximal isometric force (Po) was decreased by approximately 32%, approximately 8%, and approximately 15% in protocols 1, 2, and 3, respectively. In contrast, neither isometric contractions nor passive stretching (protocols 4 and 5) resulted in any reduction in Po. Irrespective of muscle length, the extent of force deficit was highly correlated (R = -0.774, P < 0.001) with peak force during active lengthening. Thus, the magnitude of isometric force deficit following lengthening contractions is influenced by both initial muscle length and peak force development. These findings have important practical implications for both exercise conditioning and rehabilitation, which are discussed. PMID- 12115971 TI - Severe sensory ataxia and demyelinating polyneuropathy with IgM anti-GM2 and GalNAc-GD1A antibodies. AB - Several polyneuropathy syndromes have been described with polyclonal serum immunoglobulin G (IgG) or immunoglobulin M (IgM) binding to gangliosides GM2 and GalNAc-GD1a that contain the terminal trisaccharide moiety GalNAc(beta1 4)Gal(alpha2-3)NeuAc. We describe the clinical and electrodiagnostic features in two patients with serum IgM monoclonal anti-GM2 and anti-GalNAc-GD1a antibodies. These patients had slowly progressive, panmodal sensory loss with severe sensory ataxia. Electrodiagnostic testing showed demyelinating features. Prominent improvement in gait ataxia occurred after treatment with human immune globulin but not after other immunomodulating therapies. Enzyme-linked immunoabsorbent assay and thin-layer chromatography demonstrate that the patient's serum monoclonal IgM bound to gangliosides GM2 and GalNac-GD1a but not to gangliosides without the GalNAc(beta1-4)Gal(alpha2-3)NeuAc moiety. This neuropathy differs from previously reported neuropathy syndromes associated with polyclonal GM2 and GalNAc-GD1a antibodies and from other chronic demyelinating polyneuropathies. We conclude that a distinct syndrome of chronic demyelinating neuropathy with sensory ataxia, unresponsive to corticosteroids, is associated with monoclonal IgM binding to gangliosides with a terminal GalNAc(beta1-4)Gal(alpha2-3)NeuAc trisaccharide moiety. Diagnosis of this syndrome is important to direct appropriate treatment. PMID- 12115972 TI - CD11b+ neutrophils predominate over RAM11+ macrophages in stretch-injured muscle. AB - The purpose of this study was to determine whether both neutrophils and macrophages infiltrate the hematoma site of stretch-injured rabbit tibialis anterior muscle. The Mab.198 antibody was used to detect CD11b(+) neutrophils or macrophages. Neutrophils were identified specifically by using the RPN3/57 antibody. The RAM11 antibody was used to detect macrophages. The histological characteristics of the hematoma site, torn fibers or inflammatory cells, were present primarily at 4 and 24 h, but not at 48 and 72 h after injury. A difference in the Mab.198(+) cellular concentration was detected over time between uninjured and injured muscles (P = 0.03). The injured-uninjured difference in the RPN3/57(+) or RAM11(+) cellular concentrations approached significance (P = 0.07) or else was deemed insignificant (P = 0.13), respectively. Therefore, neutrophils may predominate over RAM11(+) macrophages in stretch-injured muscle. These findings may influence the antiinflammatory strategies used to treat stretch injuries. PMID- 12115973 TI - Lateral spread response elicited by double stimulation in patients with hemifacial spasm. AB - In patients with hemifacial spasm (HFS), a lateral spread response (or abnormal muscle response) is recorded from facial muscles after facial nerve stimulation. The origin of this response is not completely understood. We studied the lateral spread responses elicited by double stimulation in 12 patients with HFS during microvascular decompression. The response was recorded from the mentalis muscle by electrical stimulation of the temporal branch of the facial nerve or from the orbicularis oculi muscles by stimulation of the marginal mandibular branch. The interstimulus intervals (ISIs) of double stimulation ranged from 0.5 to 7.0 ms. R1 was defined as the response elicited by the first stimulus, and R2 as the response elicited by the second stimulus. R1 had a constant latency and amplitude regardless of the ISI, whereas R2 appeared after a fixed refractory period without facilitation or depression in a recovery curve of latency and amplitude. From these findings, we consider that the lateral spread response is due to cross transmission of facial nerve fibers at the site of vascular compression rather than arising from facial nerve motor neurons. PMID- 12115974 TI - Effects of age and gene dose on skeletal muscle sodium channel gating in mice deficient in myotonic dystrophy protein kinase. AB - Myotonic muscular dystrophy (DM) is characterized by abnormal skeletal muscle Na channel gating and reduced levels of myotonic dystrophy protein kinase (DMPK). Electrophysiological measurements show that mice deficient in Dmpk have reduced Na currents in muscle. We now find that the Na channel expression level is normal in mouse muscle partially or completely deficient in Dmpk. Reduced current amplitudes are not changed by age or gene dose, and the reduction is not due to changes in macroscopic or microscopic gating kinetics. The mechanism of abnormal membrane excitability in DM may in part be silencing of muscle Na channels due to Dmpk deficiency. PMID- 12115975 TI - Contractile muscle volume and agonist-antagonist coactivation account for differences in torque between young and older women. AB - It is controversial whether specific tension (the ratio between muscle strength and size) declines with aging. Therefore, contractile muscle volume was estimated separately from the intramuscular noncontractile tissue by magnetic resonance imaging, and maximum isometric torque was measured in the knee extensors and flexors of 10 young (22.8 +/- 5.7 years) and 10 older (69.5 +/- 2.4 years) healthy active women. Specific tension was lower in the older women both in the extensors (93.1 +/- 20.1 kN x m(-2) vs. 112.1 +/- 12.3 kN x m(-2); P < 0.05) and in the flexors (100 +/- 31 kN x m(-2) vs. 142.7 +/- 23.9 kN x m(-2); P < 0.01). This was accompanied by an increase in the percentage coactivation of the knee flexors during knee extension. These data suggest that the lower level of muscle torque in the older women can be explained not only by smaller contractile muscle mass but also by increased coactivation of the antagonist muscles during knee extension. PMID- 12115976 TI - Central fatigue and motor cortical excitability during repeated shortening and lengthening actions. AB - A decline in voluntary muscle activation and adaptations in motor cortical excitability contribute to the progressive decline in voluntary force during sustained isometric contractions. However, the neuronal control of muscle activation differs between isometric and dynamic contractions. This study was designed to investigate voluntary activation, motor cortex excitability, and intracortical inhibition during fatiguing concentric and eccentric actions. Eight subjects performed 143 torque motor-controlled, repeated shortening and lengthening actions of the elbow flexor muscles. Transcranial magnetic stimulation (TMS) was applied three times every 20 cycles. Magnetic evoked motor potentials (MEP), duration of the silent period (SP), and the torque increase due to TMS were analyzed. TMS resulted in a small torque increase in unfatigued actions. With repeated actions, voluntary torque dropped rapidly and the amplitude of the TMS-induced twitches increased, especially during repeated lengthening actions. MEP area of biceps brachii and brachioradialis muscles increased during repeated actions to a similar extent during lengthening and shortening fatigue. The duration of biceps and brachioradialis SP did not change with fatigue. Thus, voluntary activation became suboptimal during fatiguing dynamic actions and motor cortex excitability increased without any changes in intracortical inhibition. The apparent dissociation of voluntary activation and motor cortex excitability suggests that the central fatigue observed, especially during lengthening actions, did not result from changes in motor cortex excitability. PMID- 12115977 TI - Impaired performance of skeletal muscle in alpha-glucosidase knockout mice. AB - Glycogen storage disease type II (GSD II) is an inherited progressive muscle disease in which lack of functional acid alpha-glucosidase (AGLU) results in lysosomal accumulation of glycogen. We report on the impact of a null mutation of the acid alpha-glucosidase gene (AGLU(-/-)) in mice on the force production capabilities, contractile mass, oxidative capacity, energy status, morphology, and desmin content of skeletal muscle. Muscle function was assessed in halothane anesthetized animals, using a recently designed murine isometric dynamometer. Maximal torque production during single tetanic contraction was 50% lower in the knockout mice than in wild type. Loss of developed torque was found to be disproportionate to the 20% loss in muscle mass. During a series of supramaximal contraction, fatigue, expressed as percentile decline of developed torque, did not differ between AGLU(-/-) mice and age-matched controls. Muscle oxidative capacity, energy status, and protein content (normalized to either dry or wet weight) were not changed in knockout mice compared to control. Alterations in muscle cell morphology were clearly visible. Desmin content was increased, whereas alpha-actinin was not. As the decline in muscle mass is insufficient to explain the degree in decline of mechanical performance, we hypothesize that the large clusters of noncontractile material present in the cytoplasm hamper longitudinal force transmission, and hence muscle contractile function. The increase in muscular desmin content is most likely reflecting adaptations to altered intracellular force transmission. PMID- 12115978 TI - Neurophysiologic parameters and symptoms in chronic renal failure. AB - We studied: (1) the sensitivity of various neurophysiologic parameters in the diagnosis of uremic polyneuropathy, (2) the relationship between subjective symptoms and neurophysiologic parameters, and (3) the effect of a single hemodialysis on the neurophysiologic parameters in 21 patients undergoing hemodialysis. The following parameters were studied: sensory and motor nerve conduction, including F-wave parameters; vibration detection thresholds; and thermal thresholds. The clinical findings and subjective symptoms were studied using a standardized questionnaire. The most sensitive parameters in the diagnosis of uremic neuropathy were F-wave parameters from lower limbs, vibration detection thresholds from the feet, and the sural nerve sensory action potential amplitude. The nerves from the upper extremities on the side of the fistula should not be used in the diagnosis of uremic polyneuropathy due to numerous mild local nerve lesions. The positive neuropathic symptoms correlated with quantitative vibratory detection thresholds and sensory nerve conduction studies, especially the amplitude of the sensory nerve action potential in the sural nerve. We found no significant change in any of the neurophysiologic parameters following a single hemodialysis session. PMID- 12115979 TI - Reinnervation of muscles after transection of the sciatic nerve in adult rats. AB - Functional recovery after transection of the sciatic nerve in adult rats is poor, probably because of abnormalities in reinnervation. Denervation and reinnervation patterns were studied morphologically in the lateral gastrocnemius (LGC), tibialis anterior (TA), and soleus (SOL) muscles for 21 weeks after nerve transection (motor endplates by acetylcholinesterase staining; nerves by silver impregnation). Motor endplates in the TA showed improving morphology with age, and, at 21 weeks, three-quarters of these were normal. Poorest recovery was observed in the SOL, as, at 21 weeks, only one-third of the motor endplates had a normal morphology. Polyneuronal innervation initially was more pronounced in the SOL, but, at 21 weeks, 10% of the motor endplates in all three muscles were still polyneuronally innervated. Our results indicate important differences in the reinnervation of these three hindleg muscles, and, even at 5 months, abnormalities were still present. These factors may in part explain the abnormal locomotion in rats as well as the limited recovery of function observed clinically in humans after nerve transection. PMID- 12115980 TI - CDNA microarray analysis of gene expression in fibroblasts of patients with X linked Emery-Dreifuss muscular dystrophy. AB - To clarify the molecular nature of the pathogenesis in X-linked Emery-Dreifuss muscular dystrophy (EDMD), we monitored the expression of 2400 genes in control and EDMD fibroblasts by using complementary DNA (cDNA) microarray techniques. A total of 60 genes whose expression was altered in EDMD fibroblasts when compared with control fibroblasts were identified. Twenty-eight genes whose expression was altered with the emerin deficiency were rescued by infection with a recombinant adenovirus expressing emerin. The altered expression in five genes, including the lamin A/C gene, was confirmed by reverse transcription-polymerase chain reaction. Our preliminary results suggest a correlation between disease similarity and gene expression. We conclude that the cDNA microarray is a very efficient tool to clarify genetic and pathological features of diseases. PMID- 12115981 TI - Attenuated HSP70 response in skeletal muscle of aged rats following contractile activity. AB - The aim of this study was to investigate the production of HSP70 in gastrocnemius muscles from adult (6-month-old) and aged (28-month-old) rats following contractile activity. At 24 h following a period of repeated isometric contractions, muscles from adult rats contained significantly elevated levels of HSP70 compared with nonexercised muscle. This was not evident in muscles from aged rats. This attenuated response may play a major role in development of the age-related functional deficit that occurs in skeletal muscle. PMID- 12115982 TI - Effect of triggering potential on calculations of jitter in single-fiber EMG. AB - Jitter values are calculated in reference to a triggering potential during single fiber electromyography (SFEMG) performed during voluntary contraction. When there are more than two single-fiber action potentials (SFAP) and the selected triggering potential is from an abnormal end-plate, all the jitters calculated from the same trace will be affected. In this study, the effect of triggering potential on calculated jitter was investigated in myasthenic and healthy volunteers by switching the triggering potential and recalculating the jitter off line. Selecting a triggering potential from an abnormal end-plate increased the number of abnormal individual jitters as well as the mean jitter. Therefore, if the equipment software has the capacity to change the trigger potential, the triggering potential should not be from an abnormal end-plate if all possible jitter values are to be calculated for traces having three or more single-fiber potentials. Otherwise, only one jitter value should be included from any one trace to prevent false-positive results. PMID- 12115984 TI - Serial studies of carpal tunnel syndrome during and after pregnancy. AB - Carpal tunnel syndrome (CTS) is a frequent and underdiagnosed complication of pregnancy. Conservative therapies are common initial measures, but data on the course of improvement are limited. We report a case of pregnancy-associated CTS with unusually detailed serial electrophysiologic studies before and after wrist splinting. Physiologic measures reached a nadir and then rapidly improved following conservative therapy, paralleling clinical improvement. Responses took between 6 and 20 months postpartum to approach baseline values. PMID- 12115983 TI - Severe exacerbation of hepatitis C-associated vasculitic neuropathy following treatment with interferon alpha: a case report and literature review. AB - Vasculitic neuropathy may occur in association with chronic hepatitis C infection. Interferon alpha (IFN(alpha)), an effective treatment for chronic hepatitis C, can precipitate or worsen autoimmune diseases. We report a patient with chronic hepatitis C and mild indolent vasculitic sensorimotor peripheral polyneuropathy, who developed severe mononeuropathy multiplex soon after IFN(alpha) was initiated, and review the literature on worsening vasculitic neuropathy after IFN(alpha) treatment for chronic hepatitis C. Care should be taken after starting patient with chronic hepatitis C-associated vasculitic neuropathy on IFN(alpha), as there is evidence that IFN(alpha) may exacerbate the neuropathy. PMID- 12115985 TI - Practice parameter for electrodiagnostic studies in carpal tunnel syndrome: summary statement. PMID- 12115986 TI - Muscle endurance after limb immobilization. PMID- 12115987 TI - Advanced care planning in ALS. PMID- 12115990 TI - Hepatitis A virus prevalence in plasma donations. PMID- 12115991 TI - Trend of hepatitis B virus infection in freshmen classes at two high schools in Hualien, Taiwan from 1991 to 1999. AB - Taiwan is an endemic area of hepatitis B virus (HBV) infection. A nationwide mass vaccination program to prevent HBV infection was started in 1985. Perinatal and horizontal transmission of HBV decreased substantially after the launching of this program. However, the influence of this program on children born before 1985 has not been studied. From 1991 to 1999, annual surveys of hepatitis B virus surface antigen (HBsAg) and antibody (anti-HBs) were carried out in freshmen at two high schools in Hualien, Taiwan. The average age was 16 years old. Although these students were born 2-10 years after the start of the national HBV vaccination program, there is a significant trend of decreasing HBsAg carrier rate (from 21.0% to 10.5% in males and 14.3% to 4.7% in females) and increasing anti-HBs rate (from 56.6% to 67.8% in males and 70.3% to 75.9% in females) over the 9 years. With yearly comparison, the carrier rate of HBsAg started to show significant decrease since 1994, while the anti-HBs began to rise significantly after 1996, especially in male students. The HBsAg carrier rate in male students was significantly higher, while the anti-HBs rate was significantly lower, than that in female students in most of the years. It is concluded that the effect of HBV vaccination also reduced horizontal transmission of HBV to children born up to 7 years before the start of the program. PMID- 12115992 TI - Hepatitis B virus genotypes in Uzbekistan and validity of two different systems for genotyping. AB - Hepatitis B virus (HBV) has been classified into seven genotypes, designated A-G. The HBV genotype has a characteristic geographical distribution. The Republic of Uzbekistan is located in the heart of Asia and has been considered to be a region with high endemicity of hepatitis viruses. However, the present distribution of hepatitis virus infection in this region is unknown. The aim of this study was to investigate the distribution of HBV genotypes and to elucidate the validity of two genotyping systems in Uzbekistan. Fifty-four patients with hepatitis B surface antigen were investigated. HBV genotypes were determined by two methods: one based on restriction fragment length polymorphism (RFLP) targeting to S region, and another on enzyme-linked immunosorbent assay (ELISA), using monoclonal antibodies to pre-S2 region. Seven (13%) and 47 (87%) of the 54 subjects were classified into genotypes A and D, respectively. Dual infection of two viral populations of the same genotype was observed in one subject. No significant difference of ALT level (203.3 +/- 244.7 vs. 190.6 +/- 39.5) and HBeAg (42.9% vs. 42.6%) were found between genotypes A and D. In this study, the validity of the genotyping systems in this region was confirmed. PMID- 12115993 TI - Preponderance of hepatitis B virus genotype B contributes to a better prognosis of chronic HBV infection in Okinawa, Japan. AB - The present study was designed to examine the distribution of hepatitis B virus (HBV) genotypes among patients at various stages of chronic liver disease type B in Okinawa Prefecture, Japan, where the prevalence of hepatitis B surface antigen is the highest in Japan despite the lowest mortality rate from primary liver cancer. Serum samples from 227 HBV carriers were determined for HBV genotype by polymerase chain reaction (PCR)-restriction fragment length polymorphism. Five of 227 sera were negative for HBV DNA by nested PCR and were excluded from the genotype analysis. Genotype B was predominant in asymptomatic carriers (45/67, 67%), whereas genotype C was predominant in chronic liver disease: 49% (50/103) in patients with chronic hepatitis, 63% (20/32) in patients with cirrhosis, and 60% (12/20) in patients with hepatocellular carcinoma. The distribution of genotype B decreased with increasing liver disease severity. However, this tendency was seen among patients aged less than 50 years old, whereas the prevalence of genotype B was similar among carriers with various liver diseases who were older than age 50. In conclusion, HBV genotype B was prevalent and less frequent among patients with advanced liver disease, particularly in patients aged less than 50 years. These findings suggest that the preponderance of genotype B is responsible for the low mortality rate of primary liver cancer associated with HBV seen in Okinawa Prefecture, despite having the highest HBV carrier rate in Japanese. PMID- 12115994 TI - Variant analysis and immunogenicity prediction of envelope gene of HCV strains from China. AB - The putative envelope protein E2 of hepatitis C virus (HCV) is the most variable antigenic fragment in the whole viral genome and is responsible mainly for the large inter- and intra-individual heterogeneity of the infecting virus. To analyze variation and immunogenicity of HCV envelope genes of HCV strains from China, the serum samples from 12 Chinese patients with chronic hepatitis C from 10 cities of China were amplified, sequenced directly, and compared with international strains reported previously. Variant analysis and immunogenicity prediction were then carried out with computer programs. The results suggested that HCV envelope genes show diversity in the north and the south of China and that they vary among different genotypes. Therefore, the diversity among different genotypes and derived areas should be calculated for HCV vaccine design; simultaneously, domains characterized by the hydrophilicity and the conserved immunogenicity could be targeted, for example, the domain of aa434-444, aa408-414, aa449-462 in the envelope region and aa399-406 in HVR1. These data may hold the key for future development of HCV vaccine in China. PMID- 12115995 TI - High TT virus load as an independent factor associated with the occurrence of hepatocellular carcinoma among patients with hepatitis C virus-related chronic liver disease. AB - The TT virus (TTV) load was estimated in sera obtained from 237 patients with hepatitis C virus (HCV)-related chronic liver disease including 42 patients with hepatocellular carcinoma (HCC), by real-time detection PCR using primers and a probe derived from the well-conserved untranslated region of the TTV genome, which can detect all known TTV genotypes. Of the 237 patients studied, 18 (8%) were negative for TTV DNA, 87 (37%) had low TTV viremia (1.3 x 10(2)-9.9 x 10(3) copies/ml), and 132 (56%) had high TTV viremia (1.0 x 10(4)-2.1 x 10(6) copies/ml). Various features were compared between the patients with high TTV load (n = 132) and those with no TTV viremia or low viral load (n = 105). High TTV viremia (> or =10(4) copies/ml) was significantly associated with higher age (P < 0.05), past history of blood transfusion (P < 0.001), complication of cirrhosis (P < 0.05) or HCC (P < 0.0005), lower HCV RNA titer (P < 0.05), and lower platelet count (P < 0.01). On multivariate logistic regression analysis, high TTV viral load was a significant risk factor for HCC (P < 0.05), independent from known risk factors such as complication of liver cirrhosis (P < 0.0001) and high age (> or =65 years, P < 0.05), among all 237 patients. Furthermore, high TTV viral load was an independent risk factor for HCC among the 90 cirrhotic patients (P < 0.05). These results suggest that a high TTV viral load is associated independently with the complication of HCC and may have prognostic significance in patients with HCV-related chronic liver disease, although whether high TTV viremia mediates the progression of HCV-related chronic liver disease remains to be defined. PMID- 12115996 TI - Hepatitis C virus-Epstein-Barr virus interaction in patients with AIDS. AB - Immortalization of B cells by Epstein-Barr virus (EBV) and their subsequent proliferation leads to B-cell non-Hodgkin's lymphoma in immunocompromised patients. The role of hepatitis C virus (HCV) in B-cell non-Hodgkin's lymphoma has recently been raised, and an interaction between HCV and EBV is supported by recent in vitro experiments. The aim of this study was to investigate in vivo interactions between HCV and EBV in patients with AIDS, i.e., patients exposed to the risk of EBV-related B-cell non-Hodgkin's lymphoma. A total of 135 patients were prospectively studied. Serological and molecular markers of HCV, EBV, and human immunodeficiency virus (HIV) infection were sought. All the patients harbored latent EBV infection, and 20% had detectable HCV RNA in serum. No significant relationship was found between HIV, HCV, and EBV viral load in peripheral blood mononuclear cells or plasma. There was no difference between anti-HCV-positive and -negative patients or between HCV RNA-positive and negative patients with regard to the prevalence of EBV markers, especially EBV replication markers. The presence of EBV replication markers was not related to HCV RNA seropositivity or to HCV viral load. Five patients subsequently developed B-cell non-Hodgkin's lymphoma, none of whom had markers of EBV or HCV replication. These results argue against an in vivo interaction between HCV and EBV in patients with AIDS, and against a role of HCV infection in the occurrence of B-cell non-Hodgkin's lymphoma in these patients. PMID- 12115997 TI - Prevalence, isolation, and partial sequence analysis of hepatitis E virus from domestic animals in China. AB - Evidence that hepatitis E is zoonotic is accumulating. Serum samples were collected from pigs, cattle, and goats from various regions of China to determine whether they had been infected with hepatitis E virus (HEV). An in-house enzyme immunoassay (EIA) and reverse transcriptase-polymerase chain reaction (RT-PCR) with primers from open reading frame (ORF) 2 were used to detect anti-HEV antibodies and HEV RNA. The mean positivity rates of anti-HEV antibody for pigs and cattle were 78.8% and 6.3% but none of the goat sera were positive. Pigs may be more susceptible to infection with HEV than cattle or goats. Five of 263 pig sera were positive for HEV RNA and four of these five were also positive for anti HEV. The PCR products (nt 6007-6354) were cloned and sequenced and compared to other HEV sequences in the nucleotide databases. The five sequences shared 83-93% identity to each other at the nucleotide level and 74-79%, 73-74%, 73-78%, and 83 99% identity to HEV genotypes 1, 2, 3, and 4, respectively. They were closely related to human isolates of HEV genotype 4. Phylogenetic analyses also place these swine sequences in HEV genotype 4, resembling most closely viruses isolated from Chinese patients with acute hepatitis. These data support the hypothesis that sporadic hepatitis E in China is zoonotic. PMID- 12115998 TI - Sero-epidemiological patterns of Epstein-Barr and herpes simplex (HSV-1 and HSV 2) viruses in England and Wales. AB - The aim was to carry out a population-based sero-prevalence survey of Epstein Barr virus (EBV) across a wide age range in England and Wales and to identify any associations between EBV and herpes simplex virus types one and two (HSV-1 and 2). Sera from an age-stratified sample of 2,893 individuals, submitted for diagnostic purposes to 15 public health laboratories in England and Wales in 1994, were tested for immunoglobulin G (IgG) antibody to EBV. The samples had been tested previously for IgG antibody to HSV-1 and HSV-2. The serological profile of EBV was consistent with an endemic infection with peaks in transmission in those less than 5 years old and in young adults. An age adjusted analysis found a significant association between EBV and HSV-1 seropositivity that is most likely explained by similarities in their mode of transmission. The very low seroprevalence of HSV-2 in this sample complicated the comparisons of EBV and HSV-1 with HSV-2. Any associations were most likely explained by chance. Given the association between EBV and HSV-1, it is likely that recently documented epidemiological changes in HSV-1 also apply to EBV. Continuing surveillance of these herpes viruses is necessary as the predicted changes could have a significant public health impact, especially in the young adult population. PMID- 12116000 TI - Serological examination of human herpesvirus 6 and 7 in patients with coronary artery disease. AB - Fifty-three (96%) of 55 patients with coronary artery stenosis were positive for serum anti-HHV-6 IgG, and 50 (91%) of these patients had anti-HHV-7 IgG. The number of cases sero-positive for HHV-6 and -7 in the 54 age matched control volunteers was 52 (96%) and 53 (98%), respectively. No statistical difference in the sero-prevalence of the viruses existed between the patients and the control group. The mean geometric titer (log10) of anti-HHV-6 IgG in both the patients and controls were the same (1.4) (P = 0.845), whereas anti-HHV-7 titers were 1.4 and 1.5, respectively (P = 0.161). Ten (18%) of the 55 patients had anti-HHV-6 IgM; eight (15%) of the 54 control volunteers were also positive (P = 0.636). Three (5%) of the 55 patients had anti-HHV-7 IgM, whereas 3 (6%) of the 54 control volunteers had detectable serum antibody (P = 0.691). Forty-seven of the 55 patients were examined by follow-up angiographic evaluation to clarify the association between viral infection and restenosis following balloon angioplasty. Fifteen of these patients developed restenosis, as determined by angiography. The mean geometric titer (log10) of anti-HHV-6 IgG were 1.3 and 1.4 in patients with restenosis and those without restenosis, respectively (P = 0.724). The mean geometric titer (log10) of anti-HHV-7 IgG in patients with restenosis was not significantly higher (1.5) than in patients without restenosis (1.3) (P = 0.099). Three (20%) of the 15 patients affected by restenosis had anti-HHV-6 IgM; five (16%) of the 32 control patients also had the antibody (P = 0.965). One (7%) of the 15 patients with restenosis and 2 (6%) of the 32 patients without restenosis had anti-HHV-7 IgM (P = 0.558). The present study demonstrates that HHV-6 and -7 reactivation is not associated with the establishment of coronary artery stenosis and restenosis following balloon angioplasty. PMID- 12115999 TI - HHV-6 infects human aortic and heart microvascular endothelial cells, increasing their ability to secrete proinflammatory chemokines. AB - Endothelial cells are important targets for herpesvirus infection. To evaluate the biological effects of human herpesvirus-6 (HHV-6) infection, adult heart microvascular and aortic endothelial cells were examined for in vitro susceptibility to HHV-6 and for the alterations induced by viral infection on the production of monocyte chemoattractant protein-1 (MCP-1) and interleukin-8 (IL 8). Analysis by reverse transcription-polymerase chain reaction and by in situ polymerase chain reaction showed that HHV-6 replicates in endothelium in the absence of cytopathic effects, and that viral sequences were present in 20% umbilical vein and in 10% aortic and 1% microvascular endothelium. HHV-6 infection upregulated the production of MCP-1 and IL-8, with differences observed between aortic and microvascular endothelium. These findings demonstrate that endothelial cells represent a potential reservoir for HHV-6 infection, and the altered pattern of chemokine production can lead to attraction of immunocompetent cells and to the development of inflammatory processes. PMID- 12116001 TI - Neuroinvasion during delayed primary HHV-7 infection in an immunocompetent adult with encephalitis and flaccid paralysis. AB - Antibody avidity tests have been used to detect primary human herpesvirus-7 (HHV 7) infection in an immunocompetent 19-year-old man with encephalitis and flaccid paralysis for which all other suspected causes had been excluded. The finding of the viral DNA in the cerebrospinal fluid (CSF) but not in serum samples suggests that primary HHV-7 infection with invasion of the central nervous system and consequential disease had occurred. As almost all adults are infected with HHV-7 in early childhood, the present case of delayed primary infection with serious symptoms must be exceptionally rare and no cases of such late acquisition of the virus have been documented in the literature. This report of HHV-7 DNA in the CSF of an immunocompetent adult is also unique. PMID- 12116002 TI - High prevalence of human herpesvirus 8 (HHV-8) infection in south Texas children. AB - Human herpesvirus 8 (HHV-8), also known as Kaposi's sarcoma-associated herpesvirus, is etiologically associated with Kaposi's sarcoma and other rare malignancies. HHV-8 infection is common in certain areas of Africa and Italy, but occurs in only 0-15% of populations in North America and Europe. The epidemiology and prevalence of HHV-8 infection among children in the United States has not been determined, but is assumed to be low based on limited studies. The objective of this study was to determine the seroprevalence and possible risk factors of HHV-8 infection in children living in south Texas. Questionnaire data were collected and HHV-8 serologic tests were performed from a consecutive, non probability sample of 123 healthy children (ages 4-13 years) attending general pediatric clinics in south Texas. Serum was tested for HHV-8 antibodies by latent immunofluorescence assay and ORF65 enzyme-linked immunosorbent assay confirmed by immunoblot. HHV-8 prevalence and 95 percent confidence intervals were calculated using standard epidemiologic methods. Logistic regression was used to assess independent risk factors associated with HHV-8 seropositivity. The overall prevalence of HHV-8 infection was 26%. No statistically significant associations were exhibited between HHV-8 prevalence and the variables under study. The prevalence of HHV-8 infection among children in south Texas, particularly among those under the age of 12 years, indicates that non-sexual transmission of this virus is likely to occur among this population. Future investigations of larger study samples will be necessary to develop an understanding of specific routes and risk factors of HHV-8 transmission among children in south Texas. PMID- 12116003 TI - Impaired fibrinolysis in the pathogenesis of dengue hemorrhagic fever. AB - The mechanisms contributing to bleeding complications in dengue hemorrhagic fever were studied by investigating the pattern of activation of the coagulation and fibrinolytic systems in 50 children with severe dengue hemorrhagic fever. Thirteen patients (26%) died, and activation of coagulation was most pronounced in the deceased group. Fibrinolysis was also activated, but this activation was relatively weak compared with that of coagulation as a result of persistently high plasminogen activator inhibitor levels. Plasminogen activator inhibitor also prevented a switch from the procoagulant to the profibrinolytic state in lethal dengue hemorrhagic fever, which was further enhanced by an acquired protein C deficiency. The present study is the first to demonstrate such a mechanism in a viral infection. This imbalance between coagulation and fibrinolysis may be used as a prognostic marker, but it may also be a target for future therapeutic intervention. PMID- 12116004 TI - Development and evaluation of a 'real-time' RT-PCR for the detection of enterovirus and parechovirus RNA in CSF and throat swab samples. AB - A two-step reverse transcriptase TaqMantrade mark duplex PCR (RT-PCR) assay was developed using the ABI 7700 Sequence Detection System for the detection of enterovirus (EV) and parechovirus type 1 and 2 (PEV) RNA from samples of cerebrospinal fluid (CSF) and throat swabs. Using sequence-specific fluorescent dye labeled probes and continuous 'real-time' monitoring, PCR amplified product accumulation was measured. Based on limiting dilutions, the TaqMantrade mark enterovirus and parechovirus RT-PCR showed an increase of two orders of magnitude compared to cell culture with sensitivity of 100% (7/7) when assessed using enterovirus cell culture positive samples (CSF, TS). The assays were specific for enterovirus and parechovirus and did not amplify a wide selection of virus and bacterial isolates. RNA was amplified from 22 enterovirus serotypes: coxsackie A7, A9, A21; coxsackie B2, B3, B4, B5; echovirus 2, 4, 6, 7, 9, 11, 13, 17, 18, 19, 30, 31; poliovirus types 1, 2, and 3, and parechovirus types 1 and 2. The assay was used to assess the incidence of enterovirus and parechovirus RNA in cell culture negative CSF and throat swab samples (n = 200). An additional 33 (15.9%) enterovirus and 2 (1%) parechovirus were identified as positive by RT PCR. Also, of 100 CSF samples from suspected cases of meningococcal meningitis submitted for meningococcal PCR testing, 59 (59%) were enterovirus and 2 (2%) parechovirus 1 and 2 were positive by RT-PCR. The TaqMantrade mark duplex assay offers a more rapid and sensitive alternative to conventional cell culture for the diagnosis of enterovirus and parechovirus infection. Closed tube real-time detection using the ABI Sequence Detection System obviates the need for post-PCR manipulation, which reduces hands on time and eliminates the risk of contamination from amplified PCR product. PMID- 12116005 TI - Detection and characterisation of human astroviruses in children with acute gastroenteritis in Blantyre, Malawi. AB - In a 2-year hospital-based study of paediatric gastroenteritis in Blantyre, Malawi, astroviruses were detected by enzyme immunoassay in 15 (1.9%) of 786 inpatients and in 9 (2.3%) of 400 outpatients. Greater disease severity was noted in children coinfected with human immunodeficiency virus (HIV). Six human astrovirus (HAstV) genotypes were identified, including HAstV-1 (25%), HAstV-2 (21%), HAstV-3 (25%), HAstV-4 (13%), HAstV-5 (4%), and HAstV-8 (13%). Although astroviruses are not major causes of gastroenteritis among children admitted to hospital in Blantyre, concomitant HIV infection appears to be a risk factor for increased severity of disease. PMID- 12116006 TI - Molecular detection and sequence analysis of human caliciviruses from acute gastroenteritis outbreaks in Hungary. AB - Three viral gastroenteritis (VGE) outbreaks that occurred in 1998-1999, in Hungary were investigated for the presence of human caliciviruses (HuCVs). HuCVs in stool specimens were detected by reverse transcription-polymerase chain reaction (RT-PCR) using primer pair 289/290, which was designed based on the RNA dependent RNA polymerase (RdRp) sequence. RT-PCR results were confirmed by sequencing showing that all three outbreak strains belonged to genogroup II of "Norwalk-like viruses" (NLVs). Two strains had high sequence identity with strains in known genetic clusters (Hawaii and Lordsdale clusters). The third strain (MOH) had distinct RdRp sequence, sharing 77/86% (nt/aa) identity with Snow Mountain virus (SMV), the closest genogroup II virus. To characterize MOH further, we cloned, sequenced, and expressed in baculovirus its capsid gene. It had 75/79% (nt/aa) identity with SMV, but 97/98% (nt/aa) identity with NLV/Hillingdon/90/UK, a recently identified genetic cluster of HuCVs. The recombinant MOH (rMOH) capsid protein self-assembled into virus-like particles (VLPs), which is antigenically distinct from other recombinant HuCV capsid antigens available in our laboratory. Further study of this VLP will have important applications in antigenic characterization and diagnosis of HuCVs. PMID- 12116007 TI - HPV16 and HPV18 in genital tumors: Significantly different levels of viral integration and correlation to tumor invasiveness. AB - The integration of the high-risk HPV16 and HPV18 types into the cell genome is considered an important step in malignant transformation. The relationship between the physical status of the virus and clinical/pathological parameters was studied by type-specific and multiplex PCR for E6, E2, and E1 sequences in 86 genital tumors from different sites, consisting of 69 invasive carcinomas (including 5 microinvasive carcinomas), 9 carcinomas in situ, 6 severe dysplasias, and 2 moderate dysplasias. Forty tumors contained HPV16 (46.6%), 7 HPV18 (8.1%), and 39 both viruses (45.3%). HPV16 DNA was found either as pure integrant (35.4%), or pure episome (36.7%), or a mixture of both (27.8%). Conversely, all 46 lesions containing HPV18 showed pure integrated forms. The physical status of both types was not related to the tumor site, the tumor/node/metastasis stage, or the histological differentiation grade of the invasive carcinomas. HPV16 integration was significantly associated with invasiveness. Interestingly, in double infections when HPV16 coexisted with HPV18, its genome was found more frequently in episomal form than in single infections where, conversely, it was mostly integrated (P < 0.0001), suggesting a sort of competition for cell integration sites. The complete HPV18 integration, even in pre-neoplastic lesions, indicates a different behavior in genital transformation compared with HPV16 and may reflect a major aggressiveness of this viral type. In conclusion, virus typing in conjunction with the evaluation of the integration status may provide a better prognostic evaluation together with an improved diagnosis. PMID- 12116008 TI - Prevalence and genotype distribution of cervical human papillomavirus infection: Comparison between pregnant women and non-pregnant controls. AB - Controversies exist on the effect of pregnancy on human papillomavirus (HPV) infection. A cross-sectional section study was conducted to compare the prevalence and genotype distribution of cervical HPV infection between pregnant and non-pregnant women in Hong Kong. Cervical samples were collected from 308 pregnant women and from the same number of age-matched controls recruited from a cervical cancer screening center located at the same hospital. HPV was detected by the polymerase chain reaction, followed by genotype identification by restriction fragment length polymorphism and direct sequencing analyses. The prevalence of HPV for pregnant women was 10.1%, without significant variation with age, gestation, gravidity and parity. The prevalence of HPV for non-pregnant group was 11.4% and did not show significant difference when compared to the pregnant group either by overall or age-stratified subgroup analyses. When the analysis was stratified according to the risk-type of HPV infection, still no significant difference between pregnant and non-pregnant groups was observed (all types: 10.1 vs. 11.4%, P = 0.602; high-risk types: 5.8 vs. 7.8%, P = 0.338; low risk types: 1.0 vs. 2.9%, P = 0.080; unknown-risk types: 3.2% vs. 1.3%, P = 0.105). The results of this study show no evidence for an influence of pregnancy on HPV prevalence, and a majority of HPV-infected pregnant women had normal cervical cytology. HPV positive results in pregnant women per se should be managed conservatively. PMID- 12116009 TI - Divergent evolution of hemagglutinin and neuraminidase genes in recent influenza A:H3N2 viruses isolated in Canada. AB - Limited information is available concerning the molecular drift of the influenza neuraminidase (NA) genes. We report on the genetic variability of the NA gene from 31 influenza A:H3N2 viruses isolated in the Province of Quebec (Canada) during the last three flu seasons (1997-2000). Amino acid substitutions within the NA protein were observed at rates of 1.01% and 0.45% between strains of the 1997-1998 and 1998-1999 seasons and between those of the 1998-1999 and 1999-2000 seasons, respectively. In most strains (28/31), amino acid changes occurred within at least one of four codons (197, 339, 370, and 401) previously implicated as antigenic sites. The 8 functional and 10 framework residues that compose the catalytic site of the NA enzyme were completely conserved over the study period. All isolates contained the seven conserved asparagine-linked glycosylation sites found in the NA of the progenitor A/Hong Kong/8/68 strain. In addition, most strains (30/31) had an eighth potential glycosylation site at position 329, whereas a ninth one was found at position 93 in 16 strains. The NA of all strains in this study was related to that of the vaccine strain A/Beijing/353/89, whereas the HA genes of these strains were related to the A/Beijing/32/92 vaccine strain. Thus, it appears that recent influenza strains continue to evolve from a reassortant produced during the cocirculation of the two above variants. Interestingly, some strains whose HA genes were closely related showed significant differences in their NA genes and conversely. This study confirms the importance of analyzing both the NA and the HA genes to assess the evolution of recent influenza epidemic strains. PMID- 12116011 TI - Foodborne outbreak caused by a Norwalk-like virus in India. AB - An outbreak of acute gastroenteritis occurred in the nurses' hostel of a civil hospital in Delhi, after a farewell party involving 130 nurses and some of the housekeeping staff. All affected persons had eaten salad sandwiches at the party. Stool samples were collected from six of these patients on the second day of infection. All six samples, when tested for the presence of common bacteria, parasites, and rotavirus, were found to be negative. The clinical features of this outbreak matched the criteria set for outbreaks caused by Norwalk-like viruses (NLVs). Reverse transcription-polymerase chain reaction (RT-PCR) was carried out on these six samples, using primers from the RNA-dependent RNA polymerase (RdRp) gene of NLVs. Immunoelectron microscopy was carried out on two of the samples, using convalescent phase serum. All six samples were positive for genogroup (GG) II NLVs by RT-nested PCR. Aggregates of 32-nm viral particles were visualized by immunoelectron microscopy in one of the two samples. Sequencing of the RdRp gene was done on amplicons from three samples; phylogenetic analysis placed the isolates NDV/1999 in a Toronto virus cluster of GG II NLVs. This is the first report of a food-borne outbreak attributable to NLVs from India. PMID- 12116010 TI - Detection of measles virus RNA in whole blood stored on filter paper. AB - The purpose of this study was to evaluate the use of dried blood spots stored on filter paper as a means to provide specimens for virologic surveillance for measles virus (MV) in situations when the reverse cold chain is not available. Two single-step RT-PCR assays were evaluated for sensitivity of detection of MV nucleoprotein gene RNA. The more sensitive assay was then used to assess the stability of MV RNA in dried whole blood stored on filter paper. MV RNA was found to be stable in dried blood spots for up to 2 months at room temperature or 1 month at 37 degrees C. As few as 100 infected human peripheral blood mononuclear cells (PBMC) per blood spot could be detected using a single-step RT-PCR reaction and ethidium bromide detection. MV RNA was also detected in dried blood spots obtained from rhesus macaques after challenge with wild-type MV. In the rhesus samples, the single-step RT-PCR reaction could detect approximately 10(3) TCID(50) per blood spot, while nested PCR detected 3 TCID(50) per blood spot. The results of this laboratory-based study suggest that the use of dried blood spots stored on filter has the potential to improve virologic surveillance for MV in some areas, and they emphasize the need for continued testing under field conditions. PMID- 12116013 TI - Prolonged shedding of rotavirus in a geriatric inpatient. AB - This study concerns a nosocomial rotaviral infection of a geriatric patient with clinical symptoms of acute gastroenteritis. The virological diagnosis was based on the detection of rotaviral antigens using a Rota kit, viral genome RNA by reverse transcription-polymerase chain reaction method, and viral particles by electron microscopy in the stool samples. Prolonged rotaviral shedding was suggested to be due to impaired natural killer cell activity, possibly together with deficiency of specific local immune response of the patient. PMID- 12116012 TI - Circulation of the novel G9 and G8 rotavirus strains in Nigeria in 1998/1999. AB - An epidemiological survey investigating the prevalence of rotavirus infection in infants and young children with acute diarrhoea was undertaken in Jos State, Nigeria, between January 1998 and April 1999. In total, 672 faecal specimens were collected from children aged between 1 and 60 months with acute infantile gastroenteritis. The 10-20% stool suspensions were examined by an ELISA for the presence of group A rotavirus antigen (Rotavirus IDEIA, Dako, UK). Only 116 specimens (17.3%) were positive for the group A rotavirus antigen detected by this ELISA. The rotavirus-positive specimens were analysed with monoclonal antibodies specific for rotavirus VP6 subgroup I and II, and for VP7 serotypes G1 G4, G8, and G9. Of the rotavirus strains that could be subgrouped, VP6 subgroup I and II strains circulated at similar levels. Amongst the strains that could be serotyped, VP7 G9 strains predominated occurring in 17 cases, with G3 (n = 10) and G1 (n = 9) strains occurring in lower numbers. Four G8 strains were detected and only one G2 and no G4 strains were identified. This report extends the description of the global distribution of G9 rotavirus strains. PMID- 12116014 TI - Prevalence of respiratory syncytial virus IgG antibodies in infants living in a rural area of Mozambique. AB - A case control study was carried out in Manhica (Mozambique). Serum samples were collected from infants < 1 year of age in hospital to assess the effect of serum antibodies on the incidence of respiratory syncytial virus (RSV) infection. Sera were collected from a total of 31 cases of RSV infection and paired uninfected controls matched for age and sex. Anti-RSV antibodies were assessed by a membrane fluorescent antibody test (MFAT) for immunoglobulin G (IgG) antibodies and by a neutralizing antibody test. IgG RSV antibodies were of higher prevalence and at higher levels in the control group when compared to the infected case group (P < 0.001), indicating an important role for IgG antibodies in protection. To assess infection before recruitment, IgA RSV antibodies were also measured by MFAT. IgA RSV antibody prevalence was very low in patients and controls (0/31 and 4/31 respectively), suggesting that most of the detected IgG RSV antibody in both groups was of maternal origin. Re-analysis of data from the subset of 27 matched, IgA RSV antibody negative infant pairs mirrored the full analysis indicating that maternal antibody has an important role in RSV protection. Similar results were obtained when neutralizing antibodies were measured and when the measurement was done against subgroup A virus strain A2, subgroup B virus strain 8/60 and a contemporary subgroup A isolate, Moz00. No significant differences in the reactivity of maternal antibodies with the three virus strains were observed. The data described below represent the first analysis of the role of maternal antibodies in reducing the risk of pediatric infection in developing countries. The results reinforce the concept of maternal vaccination for the control of RSV in very young children in whom the risk and severity of infection are the highest. PMID- 12116015 TI - SEN virus infection does not affect the progression of non-A to -E liver disease. AB - SEN virus (SEN V) was discovered recently as a potential causative agent of non A, non-B, non-C, and non-E (non-A to -E) hepatitis. The aim of this study was to obtain information about the prevalence of this virus in Japan and its association with non-A to -E liver disease. Sixty-seven patients hospitalized for non-A to -E liver disease, including hepatocellular carcinoma (19 patients), cirrhosis (7 patients), chronic hepatitis (18 patients), and acute hepatitis (23 patients), were tested, along with 49 blood donors. The patients were admitted to Nihon University Hospital between 1991 and 1998. SEN V DNA was detected by a nested polymerase chain reaction, targeting the 5' untranslated region. SEN V DNA was detected in 14 of 49 (28.6%) blood donors and in 33 of 67 (49.3%) patients with non-A to -E liver disease. The prevalence of SEN V DNA was similar among patients with various liver diseases, including hepatocellular carcinoma (42.1%), cirrhosis (57.1%), chronic hepatitis (55.6%) and acute hepatitis (47.8%) and among blood donors (28.6%). There were no significant differences in the clinical profiles of patients with SEN V DNA-positive or -negative chronic hepatitis, cirrhosis, and hepatocellular carcinoma. Similarly, there were no significant differences in the clinical profiles between patients with SEN V DNA-positive and -negative acute hepatitis. In conclusion, SEN V infection is present among many blood donors and is common in patients with non-A to -E liver disease. There are insufficient data to prove a causal role for SEN virus infection in non-A to -E liver disease. PMID- 12116016 TI - High genetic diversity revealed by the study of TLMV infection in French hemodialysis patients. AB - TT virus-like minivirus (TLMV) was recently discovered as a human circovirus. Little is known about its natural history and molecular epidemiology. A study of TLMV infection is described in a population of French hemodialysis patients. TLMV DNA was tested by seminested PCR system located in the noncoding region in 81 patients divided into seven groups according to the origin of their renal disease. Quantitation of TLMV DNA in serum was carried out. Sequences from 28 patients were compared with 40 sequences retrieved from databases and 53 TLMV sequences cloned from the serum of a single patient. The prevalence of TLMV DNA in hemodialysis patients was 95.1%. In this study, 24 samples (29.6%) presented viral loads of > 125 equivalents of plasmid (Ep)/ml, and only 6 (7.8%) had viral loads of > 125 x 10(2) Ep/ml. A significant correlation (P < 0.029) was found between viral loads of > 125 x 10(2) Ep/ml and the neoplastic origin of end-stage renal disease. Analysis of 53 sequences cloned from a single individual demonstrated high sequence variability, as shown by the genetic distance of 40.2%. This genetic distance is comparable to that between the most divergent sequences of TLMV reported to date (43.5%). These data suggest that TLMV viral load is possibly related to the level of immunocompetence of hemodialysis patients; the genetic diversity of TLMV is extremely high; and co-infection by different strains is possible. PMID- 12116018 TI - Safety of live mumps virus vaccines. PMID- 12116019 TI - In vitro susceptibility to infection with SIVcpz and HIV-1 is lower in chimpanzee than in human peripheral blood mononuclear cells. AB - This study was undertaken to evaluate and compare the susceptibility of chimpanzee versus human peripheral blood mononuclear cells (PBMCs) to infection with SIVcpz and HIV-1 non-syncitium inducing primary isolates. The results demonstrate clearly that chimpanzee PBMCs have a lower capacity to support viral replication as compared to human PBMCs. There was no experimental evidence that this difference was due to a lower availability of target cells for viral infection (PBMCs positive for CD4 and CCR5 molecules) or to a differential susceptibility to apoptosis (PBMCs positive for CD4 and CD95 molecules). A lower capacity of chimpanzee PBMCs to support SIVcpz and HIV-1 replication in vitro is related to a post-entry barrier to virus replication. PMID- 12116020 TI - Longitudinal use of phenotypic resistance testing to HIV-1 protease inhibitors in patients developing HAART failure. AB - An "in-house" recombinant virus protease inhibitor susceptibility assay was carried out (median of 3 per patient) retrospectively in 26 patients failing HIV protease inhibitor based therapy at regular intervals to the initiation of the first protease inhibitor. Patients were treated with either indinavir (N = 6), ritonavir (N = 10), or saquinavir (N = 10) and two nucleoside analogues. Second line therapy was based on single or dual protease inhibitor regimens occasionally containing nelfinavir. Clinically relevant resistance cut-offs associated with a poorer virological outcome from 6 months on and the clinical outcome from 3 months on were determined tentatively as 4- to 8-fold resistance for indinavir and ritonavir and 2.5- to 8-fold to saquinavir. In addition, the degree of cross resistance at the time of the change of protease inhibitor was associated with the response in viral load at 6 months to the second line therapy (P = 0.018). Cross-resistance (> or = 8-fold) between ritonavir and indinavir was common (78 and 100%). Cross-resistance between indinavir or ritonavir and saquinavir was less frequent (75 and 60% respectively) than the opposite (100%, P = 0.004). Cross-resistance to nelfinavir was encountered more frequently (> 70%) than to amprenavir (9%). The magnitudes of resistance were correlated between each other. In summary, the protease inhibitor susceptibility carried out longitudinally appears to be an earlier prognostic marker than viral load in a context of cross resistance. The magnitude of resistance, as a marker of cross-resistance, should be useful to guide second line therapy. PMID- 12116021 TI - Epstein-Barr virus and human immunodeficiency virus serological responses and viral burdens in HIV-infected patients treated with HAART. AB - Epstein-Barr virus (EBV) associated non-Hodgkin lymphoma is recognized as a complication of human immunodeficiency virus (HIV) infection. Little is known regarding the influence of highly active antiretroviral therapy (HAART) on the biology of EBV in this population. To characterize the EBV- and HIV-specific serological responses together with EBV DNA levels in a cohort of HIV-infected adults treated with HAART, a study was conducted to compare EBV and HIV serologies and EBV DNA copy number (DNAemia) over a 12-month period after the commencement of HAART. All patients were seropositive for EBV at baseline. Approximately 50% of patients had detectable EBV DNA at baseline, and 27/30 had detectable EBV DNA at some point over the follow-up period of 1 year. Changes in EBV DNA copy number over time for any individual were unpredictable. Significant increases in the levels of Epstein-Barr nuclear antigen (EBNA) and Epstein-Barr early antigen (EA) antibodies were demonstrated in the 17 patients who had a good response to HAART. Of 29 patients with paired samples tested, four-fold or greater increases in titers were detected for EA in 12/29 (41%), for EBNA in 7/29 (24%), for VCA-IgG in 4/29 (14%); four-fold decreases in titers were detected in 2/29 (7%) for EA and 12/29 (41%) for EBNA. A significant decline in the titer of anti-HIV antibodies was also demonstrated. It was concluded that patients with advanced HIV infection who respond to HAART have an increase in their EBV specific antibodies and a decrease in their HIV-specific antibodies. For the cohort overall, there was a transient increase in EBV DNA levels that had declined by 12 months. PMID- 12116022 TI - Field evaluation of the efficacy and immunogenicity of recombinant hepatitis B vaccine without HBIG in newborn Vietnamese infants. AB - A study involving more than 2,000 infants was conducted in Vietnam to assess the field effectiveness and immunogenicity of recombinant hepatitis B vaccine given at birth, 1 month, 2 months, without concomitant hepatitis B immune globulin (HBIG). All received a 5 microg dose of H-B-VAX II at birth. Infants born to non carrier mothers (Group 1; N = 1798) then received 2.5 microg doses at 1 and 2 months of age, while infants of HBeAg-negative (Group 2; N = 125) or HBeAg positive (Group 3; N = 88) carrier mothers received 5 microg doses. No Group 1 or 2 vaccinees were infected. In Group 3, 12 (14.6%) of 82 infants did become infected (estimated efficacy 84%). 98.0-98.6% of uninfected infants who were tested for anti-HBs developed a seroprotective concentration > or = 10 IU/L. In hyperendemic Vietnam, where routine maternal screening and passive-active prophylaxis of high-risk infants with vaccine plus HBIG is not feasible, administration of vaccine alone to all newborns may control effectively HBV infection. PMID- 12116023 TI - Comparison of the efficacy of lamivudine and famciclovir in Asian patients with chronic hepatitis B: results of 24 weeks of therapy. AB - Lamivudine therapy improves hepatic necro-inflammatory activity, decreases progression of fibrosis, and suppresses hepatitis B virus (HBV) replication. Famciclovir has also been shown to have some effect in the suppression of HBV replication. The aim of the study was to compare the effect of treatment with lamivudine and famciclovir on serum HBV DNA levels in patients with chronic hepatitis B and to assess safety. A prospective randomised clinical study was carried out on 100 patients with chronic hepatitis B infection (50 patients received lamivudine 100 mg daily and 50 patients received famciclovir 500 mg three times a day for 12 weeks. From the twelfth week onwards, patients were offered lamivudine 100 mg daily up to 48 weeks). Significantly more patients treated by lamivudine than by famciclovir had undetectable HBV DNA levels after 12 weeks of therapy (P < 0.001). The median HBV DNA levels were significantly lower in the lamivudine-treated patients from the second week of treatment onwards (P < 0.001 for all time points up to 12 weeks). At week 16, 4 weeks after the famciclovir treated patients were put on lamivudine, there was no longer any difference in HBV DNA levels between the two groups of patients. Both treatments were well tolerated and no serious adverse events were reported. It was concluded that in Chinese patients with chronic hepatitis B infection, lamivudine achieved effective suppression of HBV DNA levels within 4 weeks of therapy whereas famciclovir had a significantly weaker action. PMID- 12116024 TI - Endemic hepatitis C virus infection in a Sicilian town: further evidence for iatrogenic transmission. AB - The prevalence of and risk factors for HCV and HBV infections in the general population and the predictive value of ALT screening in identifying anti-HCV positive subjects have been evaluated in a small Sicilian town. A random 1:4 sampling from the census of the general population was performed. Anti-HCV, HCV RNA, HCV genotype, HBsAg, and anti-HBc were tested. The linkage between HCV infection and potential risk factors was evaluated by multiple logistic regression analysis. Among 721 subjects studied, 75 (10.4%) were anti-HCV positive. The HCV infection rate increased from 0.4% in subjects 10-29 years of age to 34% in those > 60 years of age. Among the 75 anti-HCV positive subjects, 66.7% were HCV-RNA positive and 36% had abnormal ALT, in contrast abnormal ALT levels were found in 4.3% of the 646 anti-HCV negative subjects (P < 0.01). HCV genotype 1b infected the majority (88.0%) of viremic subjects. Exposure to HBV infection (anti-HBc positivity) was found in 11.2% of subjects; HBsAg positivity was 0.7%. At multivariate analysis, two variables were associated with HCV infection: age > or = 45 years (OR 27.8; CI 95% = 11.0-70.2) and previous hospitalization (OR 2.5; CI 95% = 1.3-4.7). ALT testing had low positive predictive value (PPV = 49.1%) for HCV infection. The positive predictive value was good (88%) in people > or = 60 years of age, but minimal (16.7%) in those below 60. These findings indicate that HCV infection is common in the elderly, perhaps as a result of past iatrogenic transmission. The present low rate of HCV infection among the younger generations coupled with the low progression of the viral related liver damage does not support the projection of a future increasing incidence in the next decades of the burden of HCV-related chronic disease. HBV infection, formerly common in this area, is already in sharp decline. In an area of high HCV endemicity, screening of the general population by ALT cannot be used a surrogate marker to detect HCV infection in those susceptible to treatment. PMID- 12116025 TI - Seroprevalence and determinants of herpes simplex type 2 infection in an STD clinic in Milan, Italy. AB - A number of studies have shown that the seroprevalence of herpes simplex virus type 2 (HSV-2) is higher among persons attending clinics for sexually transmitted diseases (STD) than among the general population. The HSV-2 seroprevalence among STD patients, however, varies greatly among studies, possibly reflecting differences in the baseline prevalence of the infection among different general populations or in the distribution of risk factors. A cross-sectional study was carried out to determine the seroprevalence of and the risk factors for HSV-2 infection among 776 HIV-negative persons attending an STD clinic in Milan, Italy. All samples were tested with a commercial HSV type-2 specific gG ELISA test. The HSV-2 seroprevalence was 29.5% (95% CI: 26.3-32.7%). The seroprevalence increased with age, yet it did not differ by gender. Among persons with a current STD, the seroprevalence was 44.3%. At the multivariate analysis, older age was independently associated with HSV-2 infection. A self-reported history of genital herpes was predictive of HSV-2 infection. The agreement between history of genital herpes and HSV-2 seroprevalence was poor, however, stressing that in clinical practice, caution should be used in interpreting the presence or absence of a history of genital herpes as an indicator of the presence or absence of HSV 2 infection. Our data show that HSV-2 seroprevalence among persons attending an STD clinic in Italy is high; thus serological screening for HSV-2 might be advisable for STD patients. PMID- 12116026 TI - Quantitation of viral load in neonatal herpes simplex virus infection and comparison between type 1 and type 2. AB - Neonatal herpes simplex virus (HSV) infection is a severe disease with high mortality and morbidity in spite of the development of effective anti-viral therapies. The viral load in neonatal herpes simplex virus (HSV) infection was measured retrospectively in 37 patients. HSV DNA copy numbers in serum and cerebrospinal fluid (CSF) were quantified using a real-time PCR assay. Patients with disseminated infection had a higher viral load in their sera. whereas patients with central nervous system (CNS) infection exhibited a higher viral load in the CSF. The viral load was significantly higher in the serum of patients who died later. Interestingly, patients with HSV type-2 infection exhibited more CNS involvement and neurological impairment, together with a high viral load in the CSF, than did HSV type-1 patients. These results suggest that quantitation of HSV viral load may be useful for assessing the prognosis, and may provide additional information for the management of neonatal HSV infection. PMID- 12116027 TI - Reactivation of human herpesvirus 6 and 7 in pregnant women. AB - To elucidate the roles of human herpesvirus (HHV)-6 and -7 in pregnant women, peripheral blood samples and genital tract secretions were collected serially from pregnant women, and both serological testing and polymerase chain reaction (PCR) were carried out to detect viral DNA in the secretions. HHV-6 or HHV-7 Immunoglobulin(Ig)M antibodies were not detected in 432 plasma samples collected from pregnant women and cord blood, but IgG antibodies against both viruses were detected in all plasma samples. Significant increases in HHV-6 and HHV-7 IgG antibodies were observed in two (1.6%) and three (2.4%) pregnant women respectively of a total of 123 cases. HHV-6 DNA was detected in the genital tract in three (3.7%) of 82 pregnant women at the first trimester, and in 10 (12.2%) of the same women in the third trimester. The detection rate in the third trimester was significantly higher than that in the first trimester (P = 0.043). Although HHV-7 DNA was detected in the genital tract of two (2.7%) and seven (9.6%) pregnant women of a total of 73 during the first and third trimesters respectively, there was no statistical difference in the detection rate of the viral DNA between the trimesters. Because a significant increase in HHV-6 IgG antibodies was detected in only two pregnant women, it was not possible to carry out statistical analysis to determine the relationship between HHV-6 infection and associated clinical features. Although there was a significant increase in HHV-7 antibody titers in three pregnant women, a positive correlation between the virus infection and the clinical features was not demonstrated. There was no statistical association between virus shedding in the genital tract and the clinical features examined in this study. PMID- 12116028 TI - Frequency of Epstein-Barr virus-specific cytotoxic T lymphocytes in the blood of Southern Chinese blood donors and nasopharyngeal carcinoma patients. AB - Undifferentiated nasopharyngeal carcinoma is very common among Southern Chinese. While most patients have the disease detected and treated early, those who are diagnosed with advanced stages face a poor prognosis. Nasopharyngeal carcinoma is associated with latent Epstein-Barr virus (EBV); it was suggested previously that a cytotoxic T-lymphocyte (CTL)-based therapy targeting EBV proteins may offer a possible new form of treatment for this disease. The most likely target of this treatment is latent membrane protein 2 (LMP2). To define further the preexisting level of anti-EBV immunity in Chinese subjects, the frequency of peripheral blood mononuclear cells (PBMCs) responding to peptide epitopes was determined using an ELISPOT assay in 50 healthy control blood donors and in 26 patients newly diagnosed with nasopharyngeal carcinoma. A total of 7 LMP2, 2 LMP1, 1 EBNA3A, and 1 EBNA3B epitopes were used in a HLA-restricted manner. As reported previously for healthy virus carriers in western countries, it was found that in both groups the strongest responses were to epitopes in the EBNA proteins with weaker responses to the LMP epitopes. It was found that LMP2 epitopes were recognized in a greater percentage of both groups than previously reported, due most likely to the greater sensitivity of the ELISPOT method. However, patients with nasopharyngeal carcinoma demonstrated a weaker response than that displayed by healthy control subjects to several epitopes. The results demonstrate that LMP2 epitopes are recognized widely in an HLA-restricted manner in patients with nasopharyngeal carcinoma and that immunotherapy to boost preexisting immunity to these epitopes may offer a viable method to treat such patients or to protect against recurrence. PMID- 12116029 TI - Strong interaction between human herpesvirus 6 and peripheral blood monocytes/macrophages during acute infection. AB - Human herpesvirus 6 (HHV-6) encodes a viral chemokine and chemokine receptors that may modify the functions of monocytes/macrophages (MO/M phi) during productive HHV-6 infection. The interactions between HHV-6 and MO/M phi during acute infection, however, remain poorly understood. In this study, we investigated the tropism of HHV-6 in peripheral blood mononuclear cells (PBMCs) during acute infection. We detected 637 +/- 273 copies of viral DNA in 10(4) MO/M phi. in contrast, in 10(4) CD4+ T cells, which have been reported to be viral carriers during the acute infection of HHV-6, we found only 115 +/- 42 copies of viral DNA. Consistent with these data, virus was isolated from MO/M phi an order of magnitude more frequently than from CD4+ T cells. Viral mRNA U79/80, which indicates viral replication, was detectable in the MO/M phi. In addition, the mRNAs that encode viral chemokine receptors U12 and U51, which may modify the function of MO/M phi, were expressed in the cells. Therefore, productively infected MO/M phi may be the dominant cell population that is responsible for HHV 6 viremia during acute HHV-6 infection. The strong interaction of HHV-6 with MO/M phi may be partly responsible for the pathogenesis of this virus. PMID- 12116030 TI - Combined detection and genotyping of Chikungunya virus by a specific reverse transcription-polymerase chain reaction. AB - A reverse transcription-polymerase chain reaction (RT-PCR) was developed for the detection of Chikungunya virus infection. Based on the nonstructural protein 1 (nsP1) and glycoprotein E1 (E1) genes of Chikungunya, two primer sets were designed. Total RNA were extracted from the cell culture fluid of Aedes albopictus C6/36 cells inoculated with the S27 prototype virus, isolated in Tanzania in 1953, and the Malaysian strains (MALh0198, MALh0298, and MALh0398), isolated in Malaysia in 1998. For both sets of RNA samples, the expected 354- and 294-base pair (bp) cDNA fragments were amplified effectively from the nsP1 and E1 genes, respectively. Phylogenetic analysis was conducted for the Malaysian strain and other virus strains isolated from different regions in the world endemic for Chikungunya, using partial E1 gene sequence data. The Malaysian strains isolated during the epidemics of 1998 fell into a cluster with other members of the Asian genotype. PMID- 12116031 TI - Detection of parvovirus B19 NS1-specific antibodies by ELISA and western blotting employing recombinant NS1 protein as antigen. AB - Human parvovirus B19 (B19) encodes a number of nonstructural proteins, including the major protein, NS1, and two structural proteins, VP1 and VP2. The use of denatured NS1 in enzyme-linked immunosorbent assay (ELISA) and Western blot (WB) assay has provided an opportunity to study some of the immunologic properties of NS1, but the results have been equivocal and the diagnostic sensitivity poor, probably because of the absence of conformational epitopes. Various viral isolates and baculovirus vectors were employed to produce recombinant B19 NS1 under nondenaturing conditions for the first time. To assess the antigenicity of purified B19 NS1, the reaction patterns of 252 samples were compared by B19 NS1 and VP2 ELISA. In sera from individuals with past infection (VP2 IgG-positive), the use of this new antigen increased significantly the sensitivity of ELISA compared with WB (78% vs. 33%, P = 0.001), contradicting perpetuated claims that B19 NS1 IgG is detected primarily in patients with arthralgia or chronic infection. Previous reports of the absence of NS1 IgG during the initial phase of infection (< 6 weeks) were proved incorrect by the detection of NS1 IgG in 60% of samples from patients recently infected by B19. Including conformational epitopes in the ELISA increases the diagnostic sensitivity, although immunologically, a temporal (years) attenuation of NS1 antibodies appears to take place. This novel diagnostic tool may be useful as a supplement in case of borderline results by VP2 ELISA and for monitoring the efficacy of future capsid-based B19 vaccines. PMID- 12116032 TI - Role for mucosal immune responses and cell-mediated immune functions in protection from airborne challenge with Venezuelan equine encephalitis virus. AB - Venezuelan equine encephalitis virus (VEEV) replicates in lymphoid tissues following peripheral inoculation and a high titre viraemia develops. Encephalitis develops after the virus enters the central nervous system from the blood, with the earliest neuronal involvement being via the olfactory nerve. Following aerosol challenge with virulent VEEV, the virus is thought to replicate in the nasal mucosa and there could be direct entry into the olfactory nerve via infected neuroepithelial cells. Protection from VEEV infection is believed to be primarily mediated by virus specific antibody. The correlation between protection and neutralising serum antibody titres is, however, inconsistent when the virulent virus is administered by the airborne route. This study demonstrates a link between antibody in serum and the nasal mucosa and protection by means of passive immunisation studies. Intra-nasal administration of antibody increased protection against airborne virus in Balb/c mice. Vaccination of mu MT strain mice that do not have functional B cells and cannot produce antibody revealed normal proliferation of spleen cells in vitro and robust cytokine production. Aerosol challenge of mu MT mice demonstrated that complete protection was only achieved when passive immunisation with antibody was supplemented with active immunisation with the TC-83 vaccine strain of the virus. This implies that cell mediated immune functions are required for protection against airborne challenge with virulent VEEV. PMID- 12116033 TI - Demographic, clinical, and virological characteristics of hepatocellular carcinoma in Asia: survey of 414 patients from four countries. AB - Demographic, etiological, clinical characteristics and treatment of hepatocellular carcinoma (HCC) were surveyed in 414 patients in Asia, including 107 from China, 15 from India, 101 from Indonesia and 191 from Japan. Males predominated in all countries, accounting for up to 75%. The mean +/- SD age at the development of HCC was about 10 years older for the patients from Japan (63.8 +/- 9.5, P < 0.001) and India (63.1 +/- 11.2, P < 0.05) than those from China (54.0 +/- 13.7) and Indonesia (53.7 +/- 14.2). Hepatitis B surface antigen (HBsAg) in serum was detected in 67% of patients from China who were tested, 27% from India, 21% from Indonesia and 18% from Japan, whereas antibody to hepatitis C virus was detected in 4%, 53%, 40% and 70%, respectively; co-occurrence of hepatitis B and C infections was seen only in 7%, 0%, 2%, and 1%, leaving an etiology other than hepatitis viruses in 22%, 20%, 36% and 11%. HCC was diagnosed primarily by ultrasonography in China (43%) and Japan (52%), and on physical examination in India (60%) and Indonesia (52%). The size of the largest tumor exceeded 5.0 cm in diameter only in 24% of the patients from Japan, much less often than in 67%, 87%, and 71%, respectively, of those from China, India and Indonesia (P < 0.001). The most favored treatment was chemolipidolization in China (81%) and Japan (81%), whereas it was transarterial embolization in India (13%) and Indonesia (26%). These results highlight common as well as distinct characteristics of HCC in Asia, and warrant the need for close cooperation toward early diagnosis and effective treatment of HCC. PMID- 12116034 TI - Vaginal dysplastic lesions in women with hysterectomy and receiving radiotherapy are linked to high-risk human papillomavirus. AB - Patients undergoing radiotherapy for advanced cervical and endometrial cancer bear a considerable risk of developing vaginal preneoplastic lesions. Radiotherapy itself has been considered to have a role in the pathogenesis of vaginal dysplasia, although human papillomavirus (HPV) involvement has also been suggested. A series of 88 patients who underwent hysterectomy and were irradiated for gynecological cancer, including 43 with postradiation vaginal dysplasia at colposcopy and 45 without vaginal lesions, were included in this study. Detection and genotyping of HPV DNA in vaginal scraping were carried out by a PCR-based method and compared with colposcopic and cytological findings and with other clinical and laboratory data. Forty-two (97.7%) colposcopy-positive subjects and 6 (13.3%) colposcopically-negative patients were PCR-positive for high-risk HPV DNA (P < 0.000001). Twenty-two out of the 43 patients with colposcopic lesions showed an abnormal Papanicolau (PAP) test. Cytologic examination was negative in all colposcopically negative women. Type 16 HPV DNA was more frequent in patients with high-grade squamous intraepithelial lesions and in patients treated with external radiotherapy, whereas other types of high-risk HPV were more common in patients with low-grade lesions and in those treated with brachytherapy. When considering colposcopy as the standard for diagnosing vaginal dysplasia, HPV DNA testing was more sensitive than the PAP test. However, the specificity of the PAP test was higher with no false-positive case. In conclusion, vaginal preneoplastic changes in women post-hysterectomy and receiving radiotherapy for cervical, endometrial, and vaginal cancer represent an HPV-related nosologic entity. Whereas colposcopic examination can detect these preneoplastic lesions, HPV genotyping is a sensitive, inexpensive, and noninvasive method that may complement colposcopy and the PAP test. PMID- 12116035 TI - Prophylaxis and treatment of influenza encephalitis in an experimental mouse model. AB - A mouse model study using mouse brain-adapted influenza A virus was performed to establish the prophylaxis and treatment of influenza encephalitis and encephalopathy. All mice died after intranasal inoculation of the brain-adapted influenza A virus (H7N3), and the pathological findings indicated the presence of significant encephalitis. Viral antigen was also detected in the brain, both pathologically and virologically. By contrast, infected mice immunized with inactivated vaccine of the same strain did not lose weight, which is an indicator of the overall condition of the mice, and all of them survived. Similarly, antiserum treatment in the early period (0-1 day post-infection) resulted in 100% survival, and no pathological findings were observed in the brain. However, mice treated with antiserum 3 days post-infection showed encephalitis with viral antigens in both glial cells and neurocytes. Although amantadine treatment for 4 days delayed weight loss, it did not prevent death from encephalitis. These results show vaccination and early antiserum treatment to be highly effective, whereas 4-day treatment of amantadine was not very effective in treating or preventing influenza encephalitis. The life-prolonging effect of amantadine, however, suggests that use of amantadine together with other treatments may inhibit the progression of encephalitis. PMID- 12116036 TI - Neonatal rotavirus infection in Belem, northern Brazil: nosocomial transmission of a P[6] G2 strain. AB - A total of 614 fecal specimens were obtained during a survey for rotavirus infection conducted between May 1996 and May 1998 among 437 newborns admitted to special care nurseries at a public hospital in the urban area of Belem, Brazil. Routine stool samples were taken weekly from all babies up to the age of 28 days. Overall, 51 (11.7%) of the neonates excreted rotaviruses while in hospital, of whom 42 (82.3%) developed asymptomatic nosocomial infection; nosocomial infection was also proved in five of the nine patients with diarrhea. Three distinct RNA profiles were detected, of which one short electropherotyping pattern was far more frequent ( approximately 90% of the strains). Using monoclonal antibody based enzyme immunoassays, 32 (62.7%) of the rotavirus-positive strains were classified as G2, and 1 (1.9%) as mixed G1 and G2. A G serotype could not be assigned to 18 (35.3%) of the isolates. A reverse transcription-polymerase chain reaction was used for determining the VP4 type-specificity of a subset of 28 rotavirus-positive samples. Characterization of the VP7-genotype specificity was also sought for 18 of these latter strains. Overall, P[6] and G2 genotypes were identified in 93% and 94% of tested samples respectively, with results being further confirmed by Southern hybridization. Although surveillance was conducted during a 25-month period, 50 (98%) of 51 rotavirus isolates clustered between January and December 1997. The earliest [P6]G2 rotavirus infections were detected by late January 1997, involving two (13- and 14-day-old) babies admitted with acute diarrhea. Thereafter, strains bearing these genotype specificities were identified among five infants with hospital-acquired gastroenteritis, followed by 16 others who were infected asymptomatically. This is the first report from Brazil describing nosocomial transmission of P[6]G2 rotavirus strains among neonates. PMID- 12116037 TI - Transforming growth factor-beta enhances human T-cell leukemia virus type I infection. AB - Human T-cell leukemia virus type I (HTLV-I), the causative agent of adult T-cell leukemia/lymphoma, is transmitted vertically by breast milk and sexually by semen. The transforming growth factor-beta (TGF-beta), a pleiotropic cytokine that is abundant in breast milk and semen, facilitates replication of HTLV-I in lymphocytes derived from asymptomatic HTLV-I carriers and transmission to cord blood lymphocytes in vitro. Transient expression assays revealed that TGF-beta can transactivate HTLV-I long terminal repeat promoter. These results suggest that TGF-beta may play a role in replication and transmission of HTLV-I. PMID- 12116038 TI - Hepatitis B in Italy: where we are 10 years since the introduction of mass vaccination. Rome, Italy, September 20-21, 2001. Proceedings. PMID- 12116039 TI - Progress in prevention and control of hepatitis B: a congress reviewing the first ten years since the introduction of mass vaccination in Italy. PMID- 12116040 TI - Hepatitis B control in Europe by universal vaccination programmes: the situation in 2001. AB - In the nine years since the Global Advisory Group of the Expanded Programme on Immunisation (WHO) set 1997 as the target for integrating hepatitis B vaccination into national immunisation programmes worldwide, 129 countries have included hepatitis B vaccine as part of their routine infant or adolescent immunisation programmes (June 2001). By the end of 2002, 41 out of the 51 countries of the WHO European Region will be implementing universal hepatitis B immunisation. The rewards of effective implementation of the programmes in countries that started 10 years ago are becoming apparent; and their success offers an exemplary model for other countries. Some other countries, however, have difficulties to incorporate hepatitis B vaccine into universal childhood immunisation programmes, because of major economic constrains and the inability to procure a constant vaccine supply. The next decade will be characterised by expanded use of hepatitis B vaccines and the increasing efforts to sustain vaccine programmes and make the vaccine available to those countries and regions that cannot afford it. In Europe, as well as in the rest of the world, work still remains to be done to support and implement interventions that will bring us closer to the WHO goal and to eradicate hepatitis B. PMID- 12116041 TI - Hepatitis B in Italy: where we are ten years after the introduction of mass vaccination. AB - In Italy, a program of vaccination against hepatitis B targeted at the immunisation of persons at high risk began in 1983. In 1991, vaccination became mandatory for all newborns and adolescents. Since then, the vaccine has been given to more than 10 million children, with an outstanding record of safety and efficacy. The coverage rate is globally around 94%, with differences between the Northern and Southern regions, with the latter having the lower acceptance rate. According to the National Surveillance System (SEIEVA), the incidence of acute hepatitis B per 10(5) inhabitants declined from 5.4 in 1990 to 2 in 2000. The reduction was even greater among 15-24-year-old individuals, where the incidence rate per 10(5) decreased from 17.3 to 2 in the same period. In parallel with the decline of hepatitis B, hepatitis delta has also declined significantly. Catch-up immunisation of unvaccinated adolescents, as well as an effort to improve the vaccination coverage rate in high-risk groups, are required to ameliorate the efficacy of the vaccination campaign. Routine administration of booster doses of vaccine is not considered necessary to sustain immunity in immunocompetent persons. PMID- 12116042 TI - Hepatitis B immunization and hepatocellular carcinoma: The Gambia Hepatitis Intervention Study. AB - The Gambia Hepatitis Intervention Study (GHIS) was initiated by the International Agency for Research on Cancer (IARC) in 1986. It consisted of a randomized trial in the Gambian population, aiming to evaluate the protection provided by HBV vaccination administered during the first year of life against primary infection, the development of chronic carriage, and primary liver cancer. The results, reported at 9 years from the introduction of the vaccine as conferring a high degree of protection (83% against primary infection and 95% against chronic carriage), are briefly discussed. It is expected that HBV vaccination will result in a reduction of mortality from hepatocellular carcinoma, the most frequent cancer in males in sub-Sahara Africa. PMID- 12116043 TI - Prevention and control of hepatitis B in China. AB - About 170 million Chinese are infected chronically with HBV and 10% suffer from chronic hepatitis. Around half a million Chinese die from hepatitis B caused hepatocellular carcinoma and endstage cirrhosis each year. From 1983 to the present, a controlled clinical trial involving 80,000 children on a universal hepatitis B vaccination programme to prevent chronic hepatitis, hepatocellular carcinoma, and endstage cirrhosis was implemented in Qidong, China. A pilot study demonstrated that the HBsAg rate reached the adult level before the fifth year of age, and neonatal vaccination with either plasma-derived or recombinant hepatitis B vaccines provided a similar 75% protective efficacy against HBV infection. The high rate of follow-up and blood tests coverage of the cohorts provided data to show 75% protection at the tenth to eleventh years of age against serum HBsAg and also against prolonged hepatic dysfunction. The strategy of controlling hepatitis B nationwide was based on the universal immunisation of newborns, beginning in cities and then the rural areas. The large-scale vaccine source was provided by domestic plants through technology transfer, first providing plasma-derived vaccine replaced completely by recombinant DNA vaccine in 1997. An official survey in 1999 using a cluster sampling of 25,878 children from 31 provinces reported an average coverage rate of three dose of hepatitis B vaccination of 70.7%, being higher in urban areas. The Ministry of Public Health of China has planned to integrate hepatitis B vaccination into the nationwide EPI program with Government-provided vaccines starting January 1, 2002. PMID- 12116044 TI - Hepatitis B vaccination: risk-benefit profile and the role of systematic reviews in the assessment of causality of adverse events following immunisation. AB - Since the early 1990s, several cases of demyelinating diseases were reported in France in association with the vaccination against hepatitis B. A large scientific, regulatory, and public debate took place to reassure the growing concern of the population. The objective of this paper is to examine the decision process undertaken both in France and in Italy; to outline the main findings of the studies conducted before and after the French decision to suspend the vaccination campaign among adolescents; and to describe the contribution of systematic review and causality criteria in the evaluation of the risk-benefit profile of vaccines. Even on the basis of the early findings, which appeared to be compatible with a low increase in the risk associated with the vaccination, it was apparent that the risk-benefit profile was unchanged for newborns, and was essentially unchanged for adolescents and for high-risk adults. The availability of subsequent negative association studies provided further reassurance. It is essential to rely on well-conducted systematic reviews to produce valid and reliable estimates of the risk-benefit profile. PMID- 12116045 TI - Viral determinants and host immune responses in the pathogenesis of HBV infection. AB - Hepatitis B virus (HBV) is a virus that infects about 350,000,000 people worldwide with a clinical spectrum of acute hepatitis, the healthy carrier state, cirrhosis and hepatocellular carcinoma (HCC). The outcome of HBV infection is the result of complicated viral-host interactions. As in other infections with non cythopatic viruses, the immune response is thought to play a crucial role in disease pathogenesis but there is increasing evidence that a variety of viral mechanisms, some depending on the function of virally encoded proteins, have a profound impact on the infected hepatocytes, the liver microenvironment, and host anti-viral responses. Indeed, the virus has evolved multiple mechanisms to ensure its success in infecting a susceptible host. The essential aspects of the life cycle of HBV and the host immune response are reviewed and recent new developments in the molecular virology of HBV, including experimental animal models, in the role of accessory viral proteins in disease pathogenesis and HCC development and in the characterisation of the T cell response in the control of HBV infection, are highlighted. PMID- 12116046 TI - Recent progress and new trends in the treatment of hepatitis B. AB - The annual rate of progression to cirrhosis in patients with chronic HBV is 0.4 to 14.2% and that of death 4 to 10%. HCC risk increases in parallel with the severity and duration of infection, with an annual incidence less than 0.5% in carriers and 6% in patients with cirrhosis. The main aim of antiviral therapy for chronic "wild-type" HBV infection is to suppress viral replication before cirrhosis and HCC develop. Two drugs are approved: IFN alpha and lamivudine. IFN alpha is costly, has a narrow range of efficacy, safety, and tolerability. Lamivudine is active, cheaper, and better tolerated but has limited efficacy, being associated with increasing resistance and loss of clinical response in the long term. IFN may be the first choice treatment in HBeAg-positive patients with a favourable profile and compensated liver disease. Patients with HBeAg-negative active disease can benefit from 12-24 months IFN treatment if early response is observed. Lamivudine should be started only after considering the uncertainties about duration of therapy and risks of stopping it. In patients with slowly progressive liver disease, treatment is better postponed until effective combination regimens are available. Lamivudine is of paramount importance in end stage chronic liver disease to suppress HBV replication and allow successful transplantation. The role of interferon in preventing HCC is controversial. In two studies comparing the incidence of HCC in patients with HBeAg-negative chronic hepatitis treated with IFN, HCC developed less frequently in sustained responders than in non-responders in Greece (2 vs. 10%, P = 0.045), but not in Milan (7 vs. 10%, P = ns). PMID- 12116047 TI - Progress in the prevention and control of viral hepatitis type B: closing remarks. AB - Hepatitis type B remains an important disease globally. Rapid advances have been made during the last two decades in the understanding of the biology of HBV, the complex interaction between the virus and the host, and the pathology and the clinical aspects of infection. Mass vaccination has been shown to be a safe and highly effective means for the control and prevention of hepatitis B. The presence of a large worldwide reservoir of carriers requires a continuous effort to develop new drugs for the treatment of persistent HBV infection. Efforts to sustain immunization programs and to make vaccines and drugs available to countries with limited resources, are the main challenges to the eradication of hepatitis B in the next few decades. PMID- 12116048 TI - Pharmacoepidemiologic detection of calcium channel blocker-induced change on digoxin clearance using multiple trough screen analysis. AB - Nonlinear mixed effects modeling was used to estimate the effects of digoxin calcium channel blockers (verapamil, diltiazem, nifedipine) in 336 serum levels gathered from 172 patients (104 male and 68 female) receiving oral digoxin as hospital in-patients. The final model describing digoxin clearance (CL) was CL (l/day)=(87.9+2.71C(cr))0.872(CCB)0.880(CHF)0.937(SPI)0.854(DFAC), where C(cr) is the estimated creatinine clearance (ml/min); CCB=1 for concomitant administration of calcium channel blockers and CCB=zero otherwise; CHF=1 for the patients with congestive heart failure and CHF=zero otherwise; SPI=1 for concomitant administration of spironolactone and SPI=zero otherwise; DFAC=1 for administration of a half-tablet of digoxin and DFAC=zero otherwise. Concomitant administration of calcium channel blockers resulted in a 13% decrease in digoxin relative clearance. PMID- 12116049 TI - Evaluation of Supermix as an in vitro model of human liver microsomal drug metabolism. AB - SUPERMIX is a commercially available formulation of insect cell-expressed human drug-metabolizing cytochrome P450 (CYP) isoforms, mixed in proportions that are optimized to parallel their relative activities in human liver microsomes. We have evaluated the apparent functional affinity and capacity of individual CYP isoforms in SUPERMIX in comparison with microsomes from a panel of 12 human livers, using enzyme kinetic studies of isoform-selective index reactions. In addition, we have measured the concentration of NADPH cytochrome P450 oxidoreductase (OR) in SUPERMIX and compared it with the concentrations of this accessory electron transfer protein in human liver microsomes. No important differences were evident in the catalytic activities of CYPs 1A2, 2C8, 2C9, 2C19, 2D6 and 3A4 between SUPERMIX and human liver microsomes. However, SUPERMIX lacks CYP2B6 activity and did not hydroxylate the antidepressant bupropion, a clinically relevant substrate of this enzyme. In addition, the concentration of OR in SUPERMIX (1198 pmol mg protein(-1)) is 17-fold higher than the mean value in human liver microsomes (70 pmol mg protein(-1)). In conclusion, SUPERMIX lacks CYP2B6 activity and contains supraphysiological concentrations of the accessory electron transfer protein OR. These factors should be considered when this formulation is used as an in vitro model in human liver microsomal drug metabolism studies. PMID- 12116050 TI - Investigation of the effects of concomitant caffeine administration on the metabolic disposition of pyrazinamide in rats. AB - The utility of pyrazinamide (PZA) in the short-course antituberculous treatment is well established. All available data support the idea that the PZA metabolite pyrazinoic acid (PA) is the active compound against M. tuberculosis. This situation warranted a deeper investigation of possible interactions with respect to its metabolic disposition. Caffeine, which is widely used as a drug and is a common constituent of most diets, shares with PZA the same metabolic enzyme, xanthine oxidase (XO). This study investigated if, and in what manner, concomitant administration of caffeine affects PZA metabolism. PZA and caffeine, in various doses (PZA=50 or 100 mg kg(-1) and caffeine= 0, 50, 100, and 150 mg kg(-1)), were administered to female Sprague-Dawley rats. PZA and its three main metabolites were quantified in 24 h urine samples by reversed phase-HPLC Concomitant administration of 100 mg kg(-1) caffeine and 50 mg kg(-1) PZA increased from the excretion (p<0.05) of the most water-soluble and the least toxic PZA metabolite 5-hydroxypyrazinoic acid (5-OH-PA) from 66.18+/-10.87 to 94.56+/-8.65 micromol/24 h. This effect was more pronounced when 100 mg kg(-1) of PZA was administered increasing excretion of 5-OH-PA from 113.28+/-70 to 173.23+/ 17.82 micromol/24 h. These results show that the metabolic disposition of PZA is affected by concomitant caffeine intake. PMID- 12116051 TI - Bioequivalence evaluation of two brands of gliclazide 80 mg tablets (Glyzide & Diamicron)--in healthy human volunteers. AB - A randomized, two-way, crossover, bioequivalence study in 24 fasting, healthy, male volunteers was conducted to compare two brands of gliclazide 80 mg tablets, Glyzide (Julphar, UAE) as test and Diamicron (Servier Industries, France) as reference product. The study was performed at the International Pharmaceutical Research Centre (IPRC), in joint venture with Speciality Hospital, Amman, Jordan. The drug was administered with 240 ml of 20% glucose solution after a 10 h overnight fasting. After dosing, serial blood samples were collected for a period of 48 h. Plasma harvested from blood was analyzed for gliclazide by validated HPLC method. Various pharmacokinetic parameters including AUC(0-t), AUC(0- proportional, variant), C(max), T(max), T(1/2), and elimination rate constant were determined from plasma concentrations of both formulations. Statistical modules (ANOVA and 90% confidence intervals) were applied to AUC(0-t), AUC(0- proportional, variant), and C(max) for bioequivalence evaluation of the two brands which revealed no significant difference between them, and 90% CI fell within US FDA accepted bioequivalence range of 80-125%. Based on these statistical inferences, Glyzide was judged bioequivalent to Diamicron. PMID- 12116052 TI - Pharmacokinetics and metabolism of a novel antifibrotic drug pirfenidone, in mice following intravenous administration. AB - The present study describes the pharmacokinetics and metabolism of pirfenidone (PD), a compound which has been shown to have significant antifibrotic effects in rodent models of pulmonary and cardiac fibrosis. Despite the fact that this compound is currently in phase II clinical trials, little data are available on the metabolism and disposition of this agent in rodents or humans. Radioactive PD [benzene ring (14)C(U)] was administered i.v. to mice at 40 mg PD/kg body weight, and animals were killed at varying times for determination of parent compound and metabolites in various tissues. The disappearance of parent compound from the plasma followed apparent 2-compartment elimination kinetics with a terminal elimination half-life of 8.6 min. Cl (0.10 ml/min/g) and V(d(ss)) (0.67 ml/g) indicated that PD was rapidly distributed in body water. This is consistent with the finding that peak tissue radioactivity occurred within 5 min following the i.v. administration of [(14)C]-PD and that well-perfused tissues, kidney>liver>lung have much higher levels of parent compound and metabolites than did fat. Two peaks isolated from plasma samples by HPLC yielded mass spectra that were consistent with initial oxidation to the alcohol followed by further metabolism to the carboxylic acid. The radioactivity recovered in the 24 h urine samples averaged 97% of the administered dose and none of that was associated with the parent compound. The short plasma half-life of parent compound in mice supports the need for additional studies in humans where the compound has been shown to have clinical benefits. PMID- 12116053 TI - Is epinephrine administration by sublingual tablet feasible for the first-aid treatment of anaphylaxis? A proof-of-concept study. AB - PURPOSE: In order to explore the feasibility of sublingual administration of epinephrine tablets as a non-invasive first-aid treatment for anaphylaxis, we studied epinephrine absorption from this dosage form in an animal model. METHODS: In a prospective, randomized, four-way crossover study, six rabbits received epinephrine 2.5 or 10 mg as a sublingual tablet, epinephrine 0.03 mg (0.3 ml) by intramuscular (IM) injection (positive control), and 0.9% NaCl (0.3 ml) IM (negative control). Pre- and post-dose blood samples were obtained for measurement of plasma epinephrine concentrations by HPLC-EC. RESULTS: After administration of epinephrine 2.5 mg as a sublingual tablet, the mean (+/-SEM) C(max) was 2369+/-392 pg/ml, and the t(max) was 20.8+/-5.7 min. After administration of epinephrine 10 mg sublingually, the C(max) was 10836+/-2234 pg/ml, and the t(max) was 21.7+/-5.4 min. After IM epinephrine, the C(max) was 6445+/-4233 pg/ml, and the t(max) was 15.8+/-4.7 min. After IM 0.9% NaCl, the C(max) (endogenous epinephrine) was 518+/-142 pg/ml. The t(max) after both of the sublingual epinephrine tablet doses did not differ significantly from the t(max) after IM epinephrine, and the C(max) after the 10 mg sublingual epinephrine tablet dose did not differ significantly from the C(max) after IM epinephrine. CONCLUSIONS: In this proof-of-concept study, administration of epinephrine as a sublingual tablet formulation resulted in rapid achievement of peak plasma epinephrine concentrations. Absorption studies in humans are needed. DEFINITIONS: HPLC-high performance liquid chromatography; EC - electrochemical detection; C(max) - maximum plasma epinephrine concentration after dosing; t(max) - time of maximum plasma epinephrine concentration. PMID- 12116054 TI - Evolving concepts of management of febrile neutropenia in children with cancer. AB - BACKGROUND: Recent investigations of febrile neutropenia in pediatric cancer patients have identified subsets of low-risk patients who can be managed with less antibiotic therapy than previously recommended standards. METHODS AND MATERIALS: PubMed and Medline were searched for prospective trials and reviews of febrile neutropenia in children. Magnitude and duration of fever and neutropenia, comorbidities, and therapeutic strategies were examined. RESULTS: Twenty-seven prospective trials and five reviews were identified. The child with cancer and low-risk febrile neutropenia is clinically well and afebrile within 24-96 hr of antibiotic therapy and has evidence of marrow recovery with a rising phagocyte count. Disqualifying comorbidities include leukemia at diagnosis or in relapse, uncontrolled cancer, age under 1 year, medical condition(s) that would otherwise require hospitalization and social or economic conditions that may potentially compromise access to care or compliance. Therapeutic strategies include parenteral or oral antibiotics in the hospital with early discharge or parenteral antibiotics in the outpatient setting followed by oral or parenteral therapy and daily reassessment. Although as many as 25% of low-risk patients require modification of therapy and/or hospitalization, life-threatening or fatal infection is exceptional. CONCLUSION: One-third to one-half the children with febrile neutropenia are at low-risk of serious infection. In the context of clinic trials, they can be safely managed with inpatient or outpatient strategies that maintain close follow-up and reduce the burden of antibiotic therapy. Adoption of these alternative strategies as the standard of care should proceed with caution guided by written protocols. PMID- 12116055 TI - Cardiac status in bone tumor survivors up to nearly 19 years after treatment with doxorubicin: a longitudinal study. AB - BACKGROUND: Longitudinal assessment of cardiac toxicity in anthracycline-treated long-term bone tumor survivors. PROCEDURES: Cardiac status was assessed in 29 patients 14.1 (range 7-18.7) years after treatment with doxorubicin (DOXO) 360 mg/m(2) (median 225-550). The median age of the patients at the time of the study was 32.5 years (range 19.7-52). The evaluation consisted of an electrocardiogram (ECG), 24-hr ambulatory ECG with analysis of heart rate variability (HRV) and echocardiography. The results were compared to those of a study of the same patients that was performed 5 years earlier 8.9 years (range 2.3-14.1) after treatment. [Postma et al.: Med Pediatr Oncol 26:230-237, 1996] RESULTS: We found no progression of ECG abnormalities, arrhythmias, or echocardiographic abnormalities. Females were at risk for reduced contractility (P = 0.006). HRV was significantly reduced compared to age- and sex-matched controls and compared to the previous results. CONCLUSIONS: Anthracycline-related late echocardiographic abnormalities and arrhythmias detected 8.9 years after treatment, showed no further deterioration with ongoing follow-up. However, there was a significant reduction of HRV. This suggests that HRV might be a sensitive test for detection of anthracycline-induced cardiac toxicity. PMID- 12116056 TI - Acute arrhythmogenicity of first-dose chemotherapeutic agents in children. AB - BACKGROUND: Chemotherapeutic agents have been reported to cause severe arrhythmias and sudden death in the first 24 hr after administration. In this prospective study, we determined the magnitude of acute arrhythmogenicity of those agents in children. PROCEDURE: Thirty-three patients with diverse malignancies (leukemia n = 16, Wilms tumor n = 3, brain tumor n = 3, lymphoma n = 3, others n = 8) were studied with Holter monitors 24 hr before, during, and in the first 24 hr following the first-dose therapy. RESULTS: Two patients experienced conduction disturbances (phases of 2nd degree sinuatrial and atrioventricular blocks) during a 4-hr period corresponding to a 30 mg/m(2) daunorubicin infusion. Eight patients experienced supraventricular extrasystole (SE), ventricular extrasystole (VE), and/or short salvos of supraventricular (SVT) and/or ventricular tachycardia (VT). Six had leukemia (therapy: daunorubicin + vincristine), one had a lymphoma (therapy: vincristine + cyclophosphamide), and the last one a brain tumor (therapy: carboplatin + procarbazine). Three patients with leukemia had pretreatment arrhythmias (1 VT, 2 SVT). One of them and the five other patients had arrhythmias during and after the first-dose therapy (2 VE, 2 SVT, 1 SVT + VE, 1 VE + SE + SVT). No patient had life-threatening arrhythmias and no prognostic value of those disturbances could be demonstrated. CONCLUSIONS: Conduction disturbances and arrhythmias are common in cancer children at the beginning of the therapy, but no acute or long-term adverse consequences are related to their appearance. PMID- 12116057 TI - Radiotherapy for medulloblastoma in children: a perspective on current international clinical research efforts. AB - BACKGROUND: The North America and four European pediatric cooperative groups have undertaken prospective studies for medulloblastoma continuously since the 1970s. In this article, we will review the results of these studies with respect specifically to the use of radiotherapy, and trace the developments that have led up to the present trials for patients with this tumor. PROCEDURE: Published and unpublished data from the North American CCG and POG and now COG studies, from the UKCCG and SIOP groups, as well as from the French and German groups were reviewed. Issues of especial interest included radiotherapy dose and dose fractionation schedules, scheduling of chemotherapy and radiotherapy, and technical aspects of treatment with radiotherapy that might impact on outcome. RESULTS AND CONCLUSIONS: Much progress has been made in the management of medulloblastoma in childhood as a consequence of the studies undertaken sequentially by these groups over the past two decades. It now seems clear that chemotherapy plays an important role for all patients. In patients with average risk disease, the use of chemotherapy has allowed a reduction in the dose of radiotherapy to the craniospinal axis and the combination of chemotherapy with radiotherapy appears to have brought about a significant improvement in disease free and overall survival in this patient population. Patients with high-risk disease fare better now than in the past as a consequence of the routine use of aggressive chemotherapy and preliminary data suggest that the use of higher doses of radiation as in the POG studies is associated with a particularly favorable outcome. Accurate delivery of radiotherapy is essential for optimal results. The avail-ability of better tools at the treating centres and quality control as an integral part of cooperative studies are likely to bring about further improvements in outcome in the future. PMID- 12116058 TI - Malignant vascular tumors in children and adolescents: a report from the Italian and German Soft Tissue Sarcoma Cooperative Group. AB - BACKGROUND: Malignant vascular tumors are extremely rare in childhood and few data on their clinical management are available. We report on a series of 18 children who had malignant vascular tumors, treated from 1980 to 2000 by the Italian and German Soft Tissue Sarcoma Cooperative Group. PROCEDURE: Twelve patients had angiosarcoma, four had malignant hemangioendothelioma, and two had Kaposi's sarcoma. Surgical resection was completed in six cases; radiotherapy was administered to 6 children, and chemotherapy to 14. RESULTS: After a median follow-up of 208 months, the 5-year survival and event-free survival rates were 30.9 and 20.8%. Six patients were alive, four in first remission (three had tumor < 5 cm, grossly completely resected), and two in second remission. Response to chemotherapy was evaluable in nine cases and was: six no response, two partial remission, one complete remission. CONCLUSIONS: Angiosarcoma and related malignant vascular tumors are aggressive neoplasms with a poor prognosis; their behavior in children seems no different from their adult counterparts. Complete surgical resection remains the mainstay of treatment, but is probably sufficient in only a minority of cases. Postoperative radiotherapy may have a role and could be added to improve local control. The role of chemotherapy is uncertain, but the high rate of metastatic spread prompts investigation into new chemotherapeutic approaches. PMID- 12116059 TI - Interferon-alpha therapy in children with malignant diseases: clinical experience in twenty-four patients treated in a single pediatric oncology unit. PMID- 12116060 TI - Retroperitoneal paraganglioma. PMID- 12116061 TI - Treatment of Czech children with acute lymphoblastic leukemia: a report of the Czech Working Group for Pediatric Hematology. PMID- 12116062 TI - Malignant rhabdoid tumor of adrenal gland. PMID- 12116063 TI - Microsatellite instability in three cases of embryonal rhabdomyosarcoma of the orbit. PMID- 12116064 TI - Cellular neurothekeoma of the chest wall. PMID- 12116065 TI - Myelodysplasia as masquerader: A woman with hypereosinophilic syndrome and twelve years of "chronic ITP". PMID- 12116066 TI - Aggressive cutaneous aspergillus infection: control by local irrigation with amphotericin B. PMID- 12116067 TI - Reversible vincristine-related flaccid paralysis in a child with acute lymphoblastic leukemia. PMID- 12116068 TI - Simultaneous occurrence of Wilms tumor and rhabdomyosarcoma in two patients. PMID- 12116069 TI - Reply to Langer et al. Expression of vascular endothelial growth factor (VEGF) and VEGF receptors in human neuroblastomas. PMID- 12116070 TI - Immunosuppression in childhood acute lymphoblastic leukemia after remission induction therapy concerns B not T lymphocytes. PMID- 12116072 TI - Multi-component behavioral intervention to promote health protective behaviors in childhood cancer survivors: the protect study. AB - BACKGROUND: Improved cure rates for childhood cancer have produced a growing population of survivors at risk for late toxicities of chemotherapy and radiation therapy. Healthy behaviors can modify these risks. We initiated a controlled prospective trial to determine if a multi-component behavioral intervention could induce change in childhood cancer survivors' health knowledge, health perceptions, and practice of health-protective behaviors. PROCEDURE: Adolescent cancer survivors attending a long-term follow-up clinic were randomized to receive standard follow-up care or standard care plus the educational intervention. Baseline measures were obtained at randomization (T(0)) and repeated 1 year (T(1)) later during the survivors' annual check-up. RESULTS: Of 272 patients enrolled and randomized, 251 are evaluable at both time points. Treatment and control groups were similar in regards to diagnosis, gender, race, and age. The change in outcome measures over the year (T(1)-T(0)) was not significantly different between the two groups as assessed by a two-sample pooled t test. However, additional exploratory analyses indicated a significant gender difference in knowledge with female survivors in the intervention group having higher scores. In addition, patients who choose certain individual health goals, such as breast/testicular self-examination, showed improved practice of the health behavior. In addition, in a very exploratory analysis, a gender difference in response to the intervention was noted, with females exhibiting a greater improvement in knowledge scores than did males. CONCLUSIONS: Although the multi behavioral educational intervention did not induce change in health knowledge, perceptions, and behaviors of childhood cancer survivors for the treatment group as a whole, gender differences and specific health goal differences were found. These findings suggest that future interventions should be tailored to reflect gender differences and the nature of the health goal under assessment. PMID- 12116073 TI - Multiplex RT-PCR assay for the detection of major fusion transcripts in Taiwanese children with B-lineage acute lymphoblastic leukemia. AB - BACKGROUND: The classification of B-lineage acute lymphoblastic leukemia (ALL) by specific chromosomal translocations may have prognostic implications. Reverse transcriptase-polymerase chain reaction (RT-PCR) assay is a useful tool for the detection of fusion transcript resulting from specific chromosomal translocation of the leukemic cells. In general, fusion transcripts are determined individually, a process which is labor intensive in order to detect all major fusion transcripts. PROCEDURE: We use a multiplex RT-PCR assay to detect both the CML- and ALL-type BCR-ABL transcripts of the t(9;22), all described variants of the E2A-PBX1 transcripts of t(1;19), the MLL-AF4 transcripts of t(4;11), and all described variants of TEL-AML1 (also termed ETV6-CBFA2) of the cryptic t(12;21) in 165 leukemic samples at diagnosis. RESULTS: The study yielded a completely concordant result with those obtained by the individual RT-PCR assay. In this cohort of Taiwan children, the relative frequencies of the four translocations of B-lineage ALL were as following: 6% with ALL-type t(9;22)/BCR-ABL, 7% t(1;19)/E2A PBX1, 3% t(4;11)/MLL-AF4, and 18% t(12;21)/TEL-AML1, comparable to those in the Western countries. CONCLUSION: Multiplex RT-PCR assay is an efficient, sensitive, accurate, and cost-effective diagnostic tool, which will likely improve our ability in accurately and rapidly risk-stratifying children with ALL. PMID- 12116074 TI - Multicentre analysis of anthracycline-induced cardiotoxicity in children following treatment according to the nephroblastoma studies SIOP No.9/GPOH and SIOP 93-01/GPOH. AB - BACKGROUND: To study cardiac function and the incidence of anthracycline-induced cardiotoxicity in children following treatment according to the nephroblastoma studies SIOP No.9/GPOH and SIOP 93-01/GPOH. PROCEDURE: Analysis of clinical status, echocardiography, and ECG findings prior to administration of anthracyclines (median cumulative doxorubicin dose: 250 mg/m(2) [range: 90-411 mg/m(2)] and after a median posttherapeutic interval of 2.9 years [range: 0-10.2 years]. Data on cardiac function before and/or after therapy could be obtained of 186 patients. RESULTS: Posttherapy left ventricular fractional shortening was reduced in 4/157 (2.5%) patients. Out of the 4 children, 2 had clinically reduced tolerance to exercise and received anticongestive therapy. Abnormal ECG findings that were not detectable prior to therapy were found in 7/124 (5.6%) children. CONCLUSIONS: The incidence of abnormal findings is low in our study group in comparison to data from the literature and might be due to the comparably short posttherapeutic interval. PMID- 12116075 TI - Clinical and social factors that affect the time to diagnosis of Mexican children with cancer. AB - BACKGROUND: There are few studies on the factors that influence the time to diagnosis (TD) in childhood cancer. The object of the present study was to determine the influence of some clinical and social factors associated to TD in children with cancer seen at Mexico City (MC) hospitals. PROCEDURE: A retrospective study was performed. A total of 4,940 clinical records of children with cancer were reviewed. Cases of cancer were grouped, according to the International Classification of Childhood Cancer. The median (med) TD was calculated for each group (type) of cancer. The association between delayed TD (longer than 1 month) and type, age at diagnosis, parental educational level, medical institution, and place of residence was analyzed, for which the odds ratio (OR) and 95% confidence intervals (CI) were obtained. RESULTS: Leukemias had the shortest TD (med = 1 month), while Hodgkin disease (HD) and retinoblastoma had the longest TD (med = 5 months). The highest risk for delayed TD was in children with HD (OR = 7.0; 95% CI 5.3-9.3), in the 10-14 age group (OR = 1.8; 95% CI 1.4-2.3), with low maternal educational level (OR = 1.5; 95% CI 1.2 2.1), in the population with no access to social security (OR = 1.3; 95% CI 1.1 1.4), and whose place of residence is far from MC (OR = 1.5; 95% CI 1.2-2.1). CONCLUSIONS: In Mexican children with cancer, age at diagnosis, and societal characteristics are important factors affecting timely diagnosis. PMID- 12116076 TI - Symptomatic hypoglycemia in children receiving oral purine analogues for treatment of childhood acute lymphoblastic leukemia. AB - BACKGROUND: Antimetabolite-based continuation therapy is commonly used for childhood acute lymphoblastic leukemia (ALL) and hypoglycemia after prolonged fasting has been recently reported. We have found that spontaneous, symptomatic hypoglycemia (SH) may also occur in such patients. PROCEDURE: Between 1995 and 1999, patients treated according to the AIEOP-ALL-95 study received BFM-type intensive chemotherapy; mercaptopurine (6-MP) was given (60 mg/m(2)/days, orally for 14 days) during the second part of induction and during consolidation therapy (25 mg/m(2)/day, orally for 8 weeks); thioguanine (6-TG) was given during reinduction therapy with protocol II (60 mg/m(2)/day, orally for 14 days); continuation therapy consisted of a combination of 6-MP (50 mg/m(2)/day orally) and methotrexate (MTX, 20 mg/m(2)/weekly, i.m.). We reviewed the charts of all patients treated for childhood ALL at our two centers. This was done to assess the incidence and the characteristics of all episodes of SH: sweating, pallor, nausea, abdominal pain with or without transient alterations of alertness, in the presence of blood glucose level of under 60 mg/dl. RESULTS: Six of 86 patients (6.9%) developed 18 episodes of SH. Five were male, none was older than 5 years, and four were only 3 years old. SH episodes occurred during consolidation (n = 2), reinduction (n = 7), and continuation (n = 9) phases. CONCLUSIONS: SH is a rare complication associated with administration of the purine analogues, mercaptopurine and thioguanine to children with reduced fat storage and young age. PMID- 12116077 TI - Need for a cooperative study: Pulmonary Langerhans cell histiocytosis and its management in adults. AB - BACKGROUND: Pulmonary involvement with Langerhans cell histiocytosis (LCH, formerly known as histiocytosis-X) presents as an interstitial process in children and adults either with or without symptoms. In contrast to other manifestations of LCH, most patients with pulmonary disease are adults. PROCEDURES: We reviewed the literature on pulmonary LCH to determine what were the clinical presentations, prognostic variables, and treatment options for this disease. RESULTS: Although there are spontaneous remissions, a large number of patients have progressive pulmonary deficiency and experience significant morbidity if not mortality from the disease. The efficacy of steroid versus chemotherapy in halting the process remains controversial, even if smoking is taken into consideration. CONCLUSIONS: A multicenter study of therapy for pulmonary LCH is the obvious answer to this dilemma. We propose that interested centers organize via the Histiocyte Society to plan and execute such a trial. PMID- 12116078 TI - Study of the distribution of 289 non-Hodgkin lymphomas using the WHO classification among children and adolescents in India. PMID- 12116079 TI - Osteopenia in children with acute lymphoblastic leukemia: a pilot study of amelioration with Pamidronate. PMID- 12116080 TI - Aseptic osteonecrosis in a child with nephroblastoma healed by hyperbaric oxygen therapy. PMID- 12116081 TI - Mucoepidermoid carcinoma of the bronchus in a 15-year-old girl with complex cytogenetic rearrangement involving 11q and over-expression of cyclin D1. PMID- 12116082 TI - Pancreatoblastoma is associated with chromosome 11p loss of heterozygosity and IGF2 overexpression. PMID- 12116083 TI - Response to chemotherapy in a child with primary bronchopulmonary leiomyosarcoma. PMID- 12116084 TI - Hepatitis C infection during treatment for childhood cancer: pitfalls in diagnosis and management. PMID- 12116085 TI - Radiotherapy for metastases to the brain in children. PMID- 12116086 TI - Variant translocations of 11q23 in infant acute lymphoblastic leukemia (ALL): do outcomes differ from t(4;11)? PMID- 12116087 TI - Duchenne muscular dystrophy-rhabdomyosarcoma, ichthyosis vulgaris/acute monoblastic leukemia: association of rare genetic disorders and childhood malignant diseases. PMID- 12116088 TI - Acute tumor lysis syndrome in Hodgkin disease. PMID- 12116089 TI - Increased factor VIII activity and dural sinus thrombosis. PMID- 12116090 TI - Asparaginase-induced acute parotitis: an uncommon and self-limiting complication. PMID- 12116091 TI - Choroid plexus carcinoma. PMID- 12116093 TI - Sonographic evaluation of mammographically detected microcalcifications without a mass prior to stereotactic core needle biopsy. AB - PURPOSE: The purpose of this prospective study was to evaluate the clinical usefulness of sonographically re-evaluating areas of microcalcification found mammographically before undertaking stereotactic core needle biopsy (SCNB). METHODS: Patients with nonpalpable breast lesions appearing as microcalcifications on mammograms and who had been referred to us for SCNB were re-evaluated sonographically before the procedure. None of the breast lesions had been associated with a density on the mammograms, and the initial sonographic evaluations had been negative. Using the mammograms for correlation, we meticulously re-evaluated the areas of microcalcifications sonographically using a high-frequency linear-array transducer. The sonographic and histopathologic results were then reviewed and correlated. The sonographic findings and visibility of the mammographically detected microcalcifications were analyzed by the 2-tailed Fisher's exact test and the chi-square test. RESULTS: Sixty-six patients, who had 68 cases of microcalcifications, were enrolled. Thirteen of the 66 patients underwent surgery, and 9 of the 13 were found to have breast carcinoma. In the sonographic re-evaluation before SCNB in these 9 patients, an associated soft tissue mass was demonstrated in 5 patients but not in the other 4. Sonographic re-evaluation also revealed abnormalities in 24 of 68 cases (35.3%), in contrast to the negative findings on the initial sonography. Using the chi-square test to identify a trend, we found that the percentage of cases that were sonographically visible was highest for clustered benign microcalcifications and lowest for segmental benign microcalcifications (p < 0.0001). CONCLUSIONS: In breast lesions that appear as microcalcifications without an associated mass on mammograms, pre-SCNB sonographic re-evaluation with a high-frequency transducer can depict microcalcifications, particularly the clustered ones, and can detect small associated masses. Although the absence of a sonographically detectable mass in areas of mammographically detected microcalcifications does not guarantee the absence of cancer, the presence of an associated mass on sonography should warrant close follow-up in the case of negative results to avoid a delay in the diagnosis of breast carcinoma. PMID- 12116094 TI - Observer experience improves reproducibility of color Doppler sonography of orbital blood vessels. AB - PURPOSE: The study investigated the reproducibility of orbital blood flow measurements with color Doppler imaging (CDI) at different stages of observer experience. METHODS: The subjects were 31 healthy volunteers and 2 sequential groups of 25 glaucoma patients each. Repeated blood flow measurements (usually 3 sets) in orbital vessels (ophthalmic artery, short posterior ciliary arteries, central retinal artery, and central retinal vein) were performed by the same observer in a single session in each subject. RESULTS: The parameters with the best reproducibility were the resistance index (mean coefficient of variation [COV], 3.3-8.8%), the peak systolic velocity (mean COV, 6.9-13.7%), the time averaged velocity (mean COV, 7.2-16.0%), and the systolic acceleration time (mean COV, 8.8-12.3%). The mean COV was greater (9.9-20.3%) for the other arterial flow parameters (end-diastolic velocity and systolic acceleration) and for the venous flow velocities (maximum and minimum). The COVs of the parameters were improved by 20-40% as the observer became more experienced in ophthalmic CDI. CONCLUSIONS: We confirm the general reliability of CDI measurements in orbital vessels and show that observer experience improves reproducibility. It appears, however, that observer performance in these measurements is vessel specific. PMID- 12116095 TI - Diagnostic criteria for locating acquired arteriovenous fistulas with color Doppler sonography. AB - PURPOSE: The purpose of this prospective study was to evaluate and determine criteria for locating acquired arteriovenous fistulas using color Doppler sonography. METHODS: We performed color Doppler sonography on 12 consecutive patients with acquired arteriovenous fistulas. We evaluated the morphologic and hemodynamic changes in the involved vessels to help locate the fistulas (10 in the extremities, 1 in the neck, and 1 in the abdomen). RESULTS: In all cases, turbulent high-velocity flow spectrum and flow signals were present at the fistula sites, and arterialized waveforms from the draining veins were detected. In the 10 cases of acquired arteriovenous fistulas in the extremities, the resistance indices in the arteries proximal to the fistulas were all less than 1.00 (mean, 0.65), whereas the resistance indices in the arteries distal to the fistulas were all 1.00 or greater (mean, 1.17). In 70% of the cases, the diameter of the artery proximal to the fistula was at least 1.2 mm larger than that distal to the fistula. The fistula site was inferred by the point of maximal venous dilatation in 70% of the cases and by the focal perivascular color artifact in 82% of the cases. The fistula site was identified on gray-scale sonography and color flow imaging in 33% and 75% of the cases, respectively. CONCLUSIONS: Fistula sites can be located effectively and quickly by a combination of major and minor diagnostic criteria. The major diagnostic criteria are (1) junction of low- and high-resistance flow in the supplying artery, (2) a high-velocity arterialized waveform in the draining vein, and (3) a turbulent, high-velocity flow spectrum at the junction of the artery and the vein. The minor diagnostic criteria are (1) direct communication between the involved artery and vein, (2) significant change in the diameter of the supplying artery, (3) a focal point of venous dilatation, and (4) a focal perivascular color artifact. PMID- 12116096 TI - Sonographically guided hydrostatic reduction of intussusception in children. AB - PURPOSE: The aim of this study was to assess the efficacy of sonographically guided hydrostatic enema in therapeutic reduction of intussusception in children and to determine whether certain factors may predict the outcome of this technique. METHODS: We retrospectively reviewed the medical records and sonographic examinations of 83 consecutive children sonographically diagnosed with 101 cases of intussusception over a 40-month period. In 99 cases, sonographically guided hydrostatic reduction was attempted. The presence of free peritoneal fluid, the presence of fluid inside the intussusception, and the initial location of the intussusception, as confirmed by sonography, along with the level of experience of the radiologist who performed the reduction were statistically analyzed to determine their effect on outcome. A p value less than 0.05 was considered significant. RESULTS: In 88 (89%) of the 99 cases, hydrostatic reduction was successful. No complications during or after hydrostatic enema were noted. The success rate was significantly lower among patients whose intussusception was located in the left side of the abdomen (p < 0.01) or contained entrapped fluid (p < 0.02) or those in whom hydrostatic reduction was not performed by an experienced sonologist (p < 0.01). The presence of free peritoneal fluid was not a predictor of outcome (p > 0.1). No complications during or after hydrostatic enema were noted. CONCLUSIONS: Sonographically guided hydrostatic reduction of intussusception is safe and effective. We recommend that this method be attempted before surgery is considered, even in cases in which the intussusception contains entrapped fluid or is located in the left side of the abdomen. The level of experience of the radiologist who performs the reduction significantly affects the results of this procedure and should be carefully considered, particularly in cases in which initial sonography reveals the presence of risk factors. PMID- 12116097 TI - Intraurethral sonography and the test-retest reliability of urethral sphincter measurements in women. AB - PURPOSE: The purpose of this prospective study was to determine the test-retest reliability of urethral sphincter morphologic measurements obtained with intraurethral sonography. METHODS: The cross-sectional urethral sphincter anatomy of 29 asymptomatic nulliparous women was studied in a blinded fashion. Each patient returned for a repeat examination on a different day. At the point of maximal rhabdosphincter thickness, the urethral diameter and circumference and the longitudinal smooth muscle and rhabdosphincter thickness, diameter, circumference, and area were measured using the ultrasound scanner's integrated software. For each measured variable, the reliability between patients was assessed with a paired t test. Intraclass correlation coefficients were calculated to assess the reliability of each intraurethral sonographic measurement obtained from the same patient. RESULTS: On test-retest analysis, the differences for each measured variable between patients were not statistically significant (p > 0.05). Of the measurements obtained from the same patient, however, longitudinal smooth muscle thickness (rho = 0.44; p = 0.006), diameter (rho = 0.49, p = 0.003), circumference (rho = 0.49, p = 0.003), and area (rho = 0.43; p = 0.009) were significantly correlated. CONCLUSIONS: The urethral longitudinal smooth muscle layer is the only structure that can be measured reliably using sonography for diagnostic use. Sonographic measurements of the rhabdosphincter may not be reliable because the outer portion of that structure lies outside the depth of penetration of a 12.5-MHz transducer. PMID- 12116098 TI - Sonography of soft tissue masses of the neck. AB - In many clinical conditions, high-resolution sonography and color (power) Doppler sonography can be used as the first-line modality for evaluating cervical soft tissue masses. Cervical cysts, lipomas, paragangliomas, neurogenic tumors, hemangiomas, and lymphangiomas often exhibit characteristic sonographic appearances. Sonography can be used for lymph node assessment, and most salivary gland diseases can be diagnosed sonographically. Sonography can be used to guide needle biopsy of soft tissue neoplasms and lymph nodes. In addition, the relationship between a cervical mass and the great vessels can be evaluated. PMID- 12116099 TI - Metastasis of primitive neuroectodermal tumor to the breast. AB - We performed mammography and sonography on a 49-year-old woman who had a mass in her left axilla that had been present for 1 month and who had undergone excision of a primitive neuroectodermal tumor (PNET) on her back 1 year before. Mammography revealed 2 adjacent, dense nodules in the left breast and enlarged lymph nodes, without a fatty hilum or internal microcalcifications, in the left axilla. Sonography showed 2 round to oval, markedly hypoechoic nodules in the left breast and enlarged, markedly hypoechoic lymph nodes, without microcalcifications, in the left axilla. We then performed sonographically guided core biopsy. Histopathologic analysis of the specimens confirmed the presence of PNET in the left breast and axillary lymph nodes. The patient was then treated with chemotherapy. To our knowledge, this report is the first to describe radiologic findings of metastasis of PNET to the breast. PMID- 12116100 TI - Prenatal sonographic diagnosis of dilated cavum vergae. AB - Several cases of enlarged cavum vergae have been reported, but prenatal diagnosis of this condition is very rare. We report 3 cases of dilated cavum vergae diagnosed prenatally using sonography. In 1 of the 3 fetuses, ventriculomegaly and lumbar meningomyelocele were additional sonographic findings. In 1 of the 3 infants, a stereotactic cyst-peritoneal shunt was placed at 6 months of age to relieve intracranial hypertension due to progressive enlargement of the cavum vergae. The infant who had a meningomyelocele required surgical repair of this defect shortly after birth; in the third infant, the dilated cavum vergae remained asymptomatic, and no surgery was necessary. When interhemispheric cystic lesions are identified prenatally, physicians must distinguish them from pathologic cysts and determine whether associated malformations are present. Sonography is useful for both the differential diagnosis and identification of associated anomalies. PMID- 12116101 TI - Role of Doppler sonography in the diagnosis of cystic lymphangioma of the scrotum. AB - Cystic lymphangioma is a congenital lymphatic malformation that is a rare cause of extratesticular scrotal masses in children; it is frequently misdiagnosed preoperatively. Complete excision is curative, but recurrence may result from incomplete excision. We report a case of cystic lymphangioma of the scrotum in a 3-year-old boy, which had been previously diagnosed as a hydrocele. Gray-scale sonography showed a multicystic extratesticular lesion; color Doppler sonography further characterized the lesion by showing blood flow within the septa. CT scanning ruled out extrascrotal involvement. The cystic mass was surgically resected. The appearance of the lesion both macroscopically and microscopically was consistent with a diagnosis of cystic lymphangioma. The child recovered uneventfully and was discharged on the third day after surgery; no evidence of recurrence was found in 6 months of follow-up. In such cases of scrotal masses in children, gray-scale and color Doppler sonography, followed by CT or MRI, are useful in diagnosing cystic lymphangioma, differentiating it from other lesions, and defining its extent, thus allowing proper surgical planning. PMID- 12116102 TI - Color duplex sonography of occlusion of the common carotid artery with reversed flow in the extracranial internal carotid artery. AB - In a small percentage of cases with an occluded common carotid artery (CCA), the patency of the arteries beyond the carotid bulb is preserved. Color duplex sonography is useful for assessing blood flow in these vessels. We present a case of retrograde flow in an internal carotid artery (ICA) with an occluded ipsilateral CCA identified using color duplex sonography in a 70-year-old man with diabetes and known atherosclerotic disease. Color duplex sonography revealed that the right CCA was totally occluded near its origin and that flow was re established at the bulb. Flow in the right ICA was directed cephalad, with a low frequency, damped waveform; flow in the right external carotid artery (ECA) was bidirectional, with increased reversed diastolic flow. Extensive atherosclerotic lesions were also found in the left side. Endarterectomy of the left carotid bifurcation was performed. Follow-up color duplex sonography 3 months later revealed a small increase of stenosis in the left CCA and mild stenosis in the left ICA. The right CCA remained occluded, but reversed flow with a high resistance flow pattern was seen in the ipsilateral ICA that supplied the ECA, which had cephalad-directed flow. PMID- 12116104 TI - Doppler sonography of normal fetal vertebral and internal carotid arteries during pregnancy. AB - PURPOSE: The aim of this study was to determine the resistance index (RI) in the fetus in both the vertebral artery and the internal carotid artery and to evaluate the relationship of those RIs with that of the umbilical artery. METHODS: In this prospective study, color Doppler examinations of the vertebral, internal carotid, and umbilical arteries were performed in fetuses with normal growth between 17 and 41 weeks' gestational age. For every week, the 10th, 50th, and 90th percentiles of the RIs in the 3 arteries plus the ratios of the RIs of the vertebral and umbilical arteries and of the internal carotid and umbilical arteries were calculated. RESULTS: In 225 examinations in 114 women, the vertebral and internal carotid arteries showed similar RI patterns, with higher RIs at mid gestation. The only difference was that the RIs in the internal carotid artery reached maximum values at a slightly later gestational age (26-28 weeks) than did those in the vertebral artery (24-25 weeks). Also, the RIs in the vertebral artery were slightly higher than were those in the internal carotid artery. RI values in the umbilical artery decreased progressively throughout gestation. Conversely, the ratios of the RIs in the vertebral and umbilical arteries and those of the RIs in the internal carotid and umbilical arteries increased slightly until the end of pregnancy, with the former ratios always higher than the latter ones. CONCLUSIONS: In normal fetuses, the pattern of blood flow resistance in the vertebral artery resembles that in the internal carotid artery. However, compared with the internal carotid artery, the vertebral artery shows higher absolute RIs with maximum values appearing earlier in the course of pregnancy. PMID- 12116103 TI - Color Doppler sonographic finding of retrograde jugular venous flow as a sign of innominate vein occlusion. AB - Occlusion or stenosis of the superior vena cava, the innominate vein, or both is an important clinical problem that requires prompt diagnosis. To confirm a suspected occlusion, imaging studies revealing the obstruction and the presence of collateral venous routes are needed. Color Doppler sonography (CDUS) is widely used to evaluate suspected venous thrombosis and collateral pathways. We present the CDUS findings in 2 cases of innominate vein occlusion. In case 1, CDUS of the neck and left upper arm, which harbored a permanent hemodialysis access, showed engorged veins in the upper arm, a patent dialysis access, and some collateral veins in the axilla. The subclavian and internal jugular veins were patent, but the flow in the left internal jugular vein was reversed. The left innominate vein was occluded. In case 2, CDUS of the upper arms showed that the veins, the dialysis access in the left upper arm, and the subclavian and jugular veins were patent, but the flow in the left internal jugular vein and in the right subclavian vein was reversed. Collateral veins were seen in the right axillary region. Both innominate veins were occluded. The resulting collateral pathways, ie, retrograde flow in the ipsilateral jugular vein crossing to the contralateral jugular vein through dural sinuses, were confirmed by venography in both cases. PMID- 12116105 TI - Role of color Doppler imaging in diagnosing and managing pregnancies complicated by placental chorioangioma. AB - PURPOSE: The purpose of this study was to evaluate the role of color Doppler imaging in the diagnosis and management of placental chorioangioma. METHODS: The medical records, sonographic reports, and sonograms of all pregnant women who had placental masses diagnosed in our sonography unit during the years 1992 through 2000 and had been evaluated using both gray-scale and color Doppler sonography were included in this study. Subjective evaluation of the amount and distribution of intralesional vascularity by color Doppler imaging was made in all cases. Cases of chorioangioma of the placenta were compared with cases of placental hemorrhage or subchorionic hematoma. The outcomes of the pregnancies were also recorded. RESULTS: Fifteen cases of placental masses were evaluated; 8 of them were identified as placental hemorrhage or subchorionic hematoma on the basis of the sonographic findings. The other 7 cases were identified prenatally as placental chorioangioma, at a mean menstrual age of 23 weeks and a mean maternal age of 29 years. The mean size of the tumor was 6.5 cm (range, 4-13 cm). All cases of chorioangioma showed either substantial internal vascularity or a large feeding vessel within the tumor. Three infants were delivered at term with favorable outcome; 2 of them demonstrated reduction of the intratumoral blood flow during follow-up. The other 4 cases were delivered at or before 32 weeks' menstrual age (1 intrauterine fetal death, 2 terminated pregnancies, and 1 normal infant). No case of placental hematoma demonstrated blood flow within the lesion or was associated with complications of the pregnancy. CONCLUSIONS: Color Doppler imaging helps differentiate placental chorioangioma from other placental lesions and may be useful in the prenatal follow-up of chorioangioma. PMID- 12116106 TI - Role of sonography in the diagnosis of gallbladder perforation. AB - PURPOSE: Gallbladder perforation is a dreaded complication of acute cholecystitis that is associated with a high mortality rate. Early detection of gallbladder perforation reduces the associated mortality and morbidity rates. The purpose of this study was to highlight the role of sonography in the diagnosis of gallbladder perforation and to compare the diagnostic accuracy of sonography with that of CT. METHODS: We retrospectively evaluated the sonographic and CT findings in surgically proven cases of gallbladder perforation. RESULTS: In 18 of 23 cases, both sonography and CT had been performed; in the other 5 cases, only sonography had been performed. Sonography helped to diagnose the defect in the gallbladder wall and gallbladder perforation in 16 (70%) of 23 patients. In the 18 cases in which both sonography and CT had been performed, sonography showed the wall defect in 11 cases (61%), whereas CT was diagnostic in 14 cases (78%). The difference between sonography and CT in the ability to visualize a defect in the gallbladder wall was not statistically significant. CONCLUSIONS: Sonography is useful for diagnosing gallbladder perforation and detecting the defect in the gallbladder wall. We believe that sonography should be the first-line imaging modality for evaluating the patients in these cases. PMID- 12116107 TI - Duplex sonographic criteria for measuring carotid stenoses. AB - PURPOSE: The aim of this retrospective study was to determine optimal duplex sonographic criteria for use in our institution for diagnosing severe carotid stenoses and to correlate those findings with angiographic measurements obtained by the European Carotid Surgery Trial (ECST), North American Symptomatic Carotid Endarterectomy Trial (NASCET), and Common Carotid (CC) methods of grading carotid stenoses. METHODS: We analyzed the angiographic data using the ECST, NASCET, and CC methods and compared the results with the duplex sonographic findings. We then calculated the sensitivity, specificity, positive and negative predictive values, and accuracy of the duplex sonographic method. Taking these parameters into account, the optimal intrastenotic peak systolic velocity (PSV) and end diastolic velocity (EDV) were derived for diagnosing severe stenoses according to the 3 angiographic methods. RESULTS: Optimal PSV and EDV values for diagnosing a 70% or greater stenosis in our laboratory were as follows: with the NASCET method of angiographic grading of stenoses, PSV 220 cm/second or greater and EDV 80 cm/second or greater, and with the ECST and CC methods, PSV 190 cm/second or greater, and EDV 65 cm/second or greater. The optimal PSV and EDV for diagnosing a stenosis of 80% or greater with the ECST grading method were 215 cm/second or greater and 90 cm/second or greater, respectively. CONCLUSIONS: Duplex sonography is a sensitive and accurate tool for evaluating severe carotid stenoses. Optimal PSVs and EDVs vary according to the angiographic method used to grade the stenosis. They are similar for stenoses 70% or greater with the NASCET method and for stenoses 80% or greater with the ECST method. PMID- 12116108 TI - Incidence and importance of reflux in testicular veins of healthy men evaluated with color duplex sonography. AB - PURPOSE: Reflux in the testicular veins plays a crucial role in the diagnosis of a varicocele. The aim of this study was to evaluate the incidence and the sonographic features-duration and velocity-of reflux in testicular veins of healthy men using color duplex sonography (CDS). METHODS: Healthy male volunteers, 18-45 years old, whose physical examinations and semen analyses were normal, were recruited for this study. The maximum diameters of testicular veins during both normal respiration and Valsalva's maneuver were measured by CDS using a 7.5-MHz linear-array transducer. Veins greater than 2 mm in diameter were considered to be a varicocele, and the subjects in these cases were excluded from the analysis. In cases in which reflux was present, the velocity and duration of reflux in the testicular veins during Valsalva's maneuver were measured. RESULTS: Seventy men, whose mean (+/- standard deviation) age was 27 +/- 7 years, were enrolled in this study. Fourteen of the 70 patients had a left varicocele and thus were excluded from the analysis. Of the 112 hemiscrotums in the remaining 56 patients, 61 (54%) had reflux induced by Valsalva's maneuver and 51 (46%) did not. Twenty-two (39%) of refluxes were on the right side, with a mean duration of 1.1 +/- 0.5 seconds, and a mean velocity of 4.2 +/- 2.1 cm/second; 39 (70%) of the refluxes were on the left side, with a mean duration of 1.1 +/- 0.5 seconds and a mean velocity of 4.9 +/- 2.3 cm/second. The incidence of reflux was significantly higher on the left side (p = 0.003). The duration and velocity of the reflux did not differ significantly between the right and left sides. The difference in the testicular vein diameters between the right (1.3 +/- 0.2 mm; n = 56) and left (1.6 +/- 0.2 mm; n = 56) sides was statistically significant (p < 0.001). CONCLUSIONS: Normal-sized testicular veins in healthy subjects had a remarkably high incidence of reflux induced by Valsalva's maneuver. The presence of reflux in subfertile men with normal testicular vein diameters is a diagnostic criterion, but it is necessary to quantify the reflux to prevent misdiagnosis of a varicocele and unnecessary surgery. The measurement of the duration and velocity limits of reflux in a large series of subjects may provide a reliable indicator for the diagnosis of varicocele. PMID- 12116109 TI - Complications of orthopedic implants: sonographic evaluation. PMID- 12116111 TI - Prenatal sonographic diagnosis of Majewski syndrome. AB - Majewski syndrome is an autosomal recessive disorder characterized by short ribs, polydactyly, short limbs, and a cleft lip. A 26-year-old woman with no family history of genetic diseases presented at 31 weeks' menstrual age with preterm labor and underwent prenatal sonography to screen for fetal anomalies. Sonography revealed a small thorax, markedly short ribs, micromelia, polydactyly, a cleft lip, dolichocephaly, and severe oligohydramnios. The other structures appeared normal. The most likely diagnosis was Majewski syndrome. Vaginal breech delivery was allowed to proceed. The 1,850-g male newborn died of respiratory failure shortly after birth. The postnatal appearance and radiographs confirmed the prenatal diagnosis. PMID- 12116110 TI - Sonographic diagnosis of a giant aneurysm of the common hepatic artery. AB - Hepatic artery aneurysms are rare vascular lesions sometimes found incidentally during abdominal imaging. We present the case of a 61-year-old man whose initial symptoms were tenderness in the right upper quadrant of the abdomen and epigastric pain. Gray-scale sonography revealed ascites and an 8.1-cm mass in the region of the porta hepatis; color Doppler sonography revealed a turbulent arterial waveform with high peak systolic velocity. We diagnosed a giant aneurysm of the common hepatic artery. Three-dimensional CT angiography confirmed this diagnosis and also revealed hemoperitoneum. The patient underwent aneurysmectomy and recovered well. This case shows that the use of both sonography and CT angiography offers a promising alternative to conventional angiography for the diagnosis of and treatment planning for hepatic artery aneurysms. PMID- 12116112 TI - Color Doppler sonography for preoperative diagnosis of an aneurysm of the ileal branch of the superior mesenteric artery. AB - Aneurysms of the superior mesenteric artery are uncommon, and aneurysms of its branches occur even less frequently. We report the case of a 60-year-old man with an aneurysm of the ileal branch of the superior mesenteric artery whose initial symptom was abdominal pain. Gray-scale and color Doppler sonography provided noninvasive, accurate preoperative identification of the aneurysm despite its uncommon location, a small branch of the superior mesenteric artery. CT and conventional and magnetic resonance angiography confirmed the sonographic diagnosis. The aneurysm was resected successfully, and the patient's postoperative course was uneventful. Although angiography is required for a definitive diagnosis and for surgical planning, sonography is a useful tool for preoperative evaluation and diagnosis of such aneurysms. PMID- 12116113 TI - Postextrasystolic potentiation of vessel compression in myocardial bridging: detection by intravascular sonography. AB - We present the case of 67-year-old man with exertional chest pain. Coronary angiography revealed myocardial bridging of the left anterior descending coronary artery (LAD) with mild systolic vessel compression. An intracoronary Doppler sonographic examination of the LAD showed a fingertip-shaped antegrade flow pattern with a steep acceleration of coronary flow velocity during early diastole, followed by a steep deceleration and then a plateau in the coronary flow velocity during mid and late diastole. An intravascular sonographic examination using a 30-MHz transducer catheter showed eccentric vessel compression during systole and substantial augmentation of vessel compression after an incidental extrasystole. Both intracoronary Doppler sonography and intravascular sonography provided important information that helped us to identify myocardial bridging in this patient. PMID- 12116114 TI - Sonographic appearance of a free-floating atheromatous plaque in a patient with acute stroke. AB - We report the case of a neurologically unstable acute stroke patient in whom carotid sonography identified the need for emergency carotid surgery because of an increased risk of distal embolization from a free-floating atheromatous plaque. Longitudinal and transverse sonograms revealed a 6-mm free-floating thrombus loosely attached to the wall of the left common carotid artery. During emergency surgery, a free-floating atheromatous plaque was seen at this location, along with extensive atheromatous lesions. The diseased segment of the common carotid artery was resected and repaired with an interposition graft, and the patient recovered uneventfully, with no recurrence of stroke. PMID- 12116120 TI - Theory of system zones in capillary zone electrophoresis. AB - In the last years, it has been shown that the formation and migration of system zones is an inherent feature of capillary zone electrophoresis (CZE) and that it depends predominantly on the composition of an actual background electrolyte (BGE). In most of the currently used BGEs, the SZs are invisible by the UV absorbance detection system, however, the comigration of SZs with the zones of analytes deteriorates the analytical performance of CZE and may be fatal for its utilization. Therefore, the theoretical predictions of the existence and migration of SZs is of key importance for the expediency of CZE. This is a review of the theoretical treatments of SZs which reveals the origin and the properties of SZs and shows how to cope with them. Also, a table of some typical BGEs is presented where the existence and mobilities of SZs are given. PMID- 12116119 TI - Characterization of dendrimer properties by capillary electrophoresis and their use as pseudostationary phases. AB - The general properties of dendrimers and in particular their electrolytic characteristics that are relevant in electrokinetic separations, are described. In order to confirm theoretical considerations on commercial dendrimer charge and hydrodynamic radius, several capillary zone electrophoresis (CZE) experiments were performed. Electrophoretic mobilities measured at different pH values indicated a sensible increase of dendrimer hydrodynamic radius at pH values lower than 2.5. This was probably due to the Coulombic repulsion of charged amine groups of the inner dendrimer shells. The principal reasons that should address the use of dendrimers as pseudostationary phases in micellar electrokinetic chromatography (MEKC) are discussed. Moreover, a survey of different separations performed utilizing dendrimers in MEKC as well as of several future plausible uses of various classes of dendrimers is presented. PMID- 12116121 TI - Quantitative structure-mobility relationship modelling of electrokinetic chromatography of metal complexes: approaches and limitations. AB - A charged surfactant or an ionic polymer added to the capillary electrolyte introduces micellar solubilization and ion-exchange interactions, respectively, as a supplementary separation principle for metal complexes among a great many other analytes. Acting as a pseudostationary phase, such electrolyte additives make the separation mechanism fairly different from that based only on differences in electrophoretic mobility. A range of quantitative structure mobility relationships were developed to explain the migration behavior of metal complexes in micellar and ion-exchange electrokinetic chromatographic systems as a function of their primary structural parameters. The validity of migration models tested using a comprehensive selection of experimental data available in the literature was shown to depend significantly on the judicious choosing of the analyte structural descriptors and on the dominance of either of the electrophoretic mobility of the analyte, partitioning into the micelle, and the ion-exchange interaction with the polymer. In the systems where the separation mechanism is dominated by neither chromatography nor electrophoresis, their relative contributions were accounted for by the degree of correlation between experimental and calculated mobilities and by variations in the absolute value and sign of regression coefficients at the most influencing parameters. PMID- 12116122 TI - Capillary electrophoretic study of interactions of metal ions with crown ethers, a sulfated beta-cyclodextrin, and zwitterionic buffers present as additives in the background electrolyte. AB - Stability constants of K, Na, Ca, and Ba with 18-crown-6, K, Na, Li with sulfated beta-cyclodextrin and K, Li, Ca, Mg, Sr, and Ba ions with ([2-hydroxy-1,1 bis(hydroxymethyl) ethyl]-amino)-1-propanesulfonic acid (TAPS) were determined by capillary electrophoresis and computed using a general least squares minimizing program CELET. The results for 18-crown-6 agreed well with those evaluated by graphical methods or reported in the literature. Previously unknown stability constants of sulfated beta-cyclodextrins and TAPS determined for alkali and alkaline earth metals show that sulfated beta-cyclodextrin interacts with monovalent metals allowing to manipulate their effective mobility. It interacts stronger with divalent metal cations. TAPS, as zwitterionic buffer widely used in various analytical, biochemical and other applications, forms complexes with alkali and alkaline earth cations, and although the stability constants are rather low, the equilibria should be taken into account when TAPS is used and metal cations are present in solution at the same time. PMID- 12116123 TI - High-resolution computer simulation of the dynamics of isoelectric focusing using carrier ampholytes: the post-separation stabilizing phase revisited. AB - A dynamic electrophoresis simulator that accepts 150 components and voltage gradients employed in the laboratory was used to provide a detailed description of the stabilizing phase in isoelectric focusing under conditions that were hitherto inaccessible. High-resolution focusing data are presented for pH gradients spanning 7 units (pH 3-10 and pH 4-11 with 20 carrier ampholytes/pH unit) and 3.5 units (pH 7-10.5 and pH 5-8.5 with 40 carrier ampholytes/pH unit). Stabilizing phase behavior for configurations (i) with the focusing column ends only permeable to OH(-) and H(+) at cathode and anode, respectively, and (ii) with the focusing column being sandwiched between NaOH (catholyte) and phosphoric acid (anolyte) are described. Simulation data reveal the stabilizing phase to be diffusion-controlled and characterized by changes that progress from the column ends towards neutrality (i.e., towards the center in case of pH gradients bracketing neutrality). Transient states are characterized by moving concentration valleys of carrier ampholytes that significantly alter the distributions of pH and conductivity. Nonlinear pH gradients are produced. The magnitude of the changes occuring is dependent on the span of the pH gradient. Gradients that encompass greater extremes of pH show more pronounced stabilizing phases. For all systems subjected to a constant 300 V/cm, the initial separation and subsequent stabilization require less than 10 min and more than 7000 min, respectively. The presence of electrolytes at the column ends disrupts the stabilizing phase, with the degree of disruption dependent on the concentrations of the acid and base employed as electrode solutions. The data not only indicate that a true steady state is never attained in the average laboratory experiment, they also suggest that a true steady state in absence of immobilized pH gradients cannot be achieved experimentally at all. PMID- 12116124 TI - Artificial neural networks for modeling electrophoretic mobilities of inorganic cations and organic cationic oximes used as antidote contra nerve paralytic chemical weapons. AB - Electrophoretic mobility of various analytes can be modeled and thus also predicted using artificial neural networks (ANNs) evaluating experiments done according to a suitable experimental design. In contrast to response surfaces modeling which can be used to predict optimal separation conditions, ANNs combined with experimental design were shown to be efficient for modeling and prediction of optimal separation conditions, while no explicit model and any knowledge of the physicochemical constants is needed. Methodology has been developed and demonstrated on separation of inorganic cations and organic oximes while various additives (methanol, complexation agent), pH or buffer concentration were followed. In our approach proposed the number of experiments necessary to find optimal separation conditions can be reduced significantly. PMID- 12116125 TI - Electrophoresis in the presence of gradients: I. Viscosity gradients. AB - In many cases, the resolution provided by capillary electrophoresis systems approaches that predicted for diffusion-limited separations. Once all device related sources of band broadening have been eliminated or minimized, only thermal diffusion remains. In principle, peaks can be sharpened using gradients of various system characteristics such as gel concentration, buffer viscosity and electric field. However, it is not clear whether this can actually increase the resolution of the system. In this article, we focus our attention on viscosity gradients and we examine both continuous and step-like variations. Our results indicate that the performance of electrophoretic systems cannot be improved by viscosity gradients. They may provide extra stacking, and thus improve the resolution, when the injection width is non-negligible. However, for the systems considered here, the best resolution is obtained when the viscosity is uniform and the stacking is entirely performed at injection. We conclude by discussing the link between these results, the fundamental laws of thermodynamics, the nature of the detection process and the importance of having nonlinear effects in nonuniform systems. PMID- 12116126 TI - Quantitation of trace analytes in capillary zone electrophoresis with UV absorbance detection: a critical study of approaches based on peak height. AB - This contribution is aimed at providing a survey of the limitations of the peak height technique. It is shown that the shape and the slope of calibration curves give a precise insight into the separation mechanism and can warn the analyst against improper selection of background electrolyte or sample dilution. This work investigates three typical situations where the sample contains (i) the minor analyte only, (ii) the minor analyte and a nonstacking bulk component, (iii) the minor analyte and a stacking bulk component, and various modes of measurement of the calibration curves differing in the medium used for the dilution of the original sample (water, background electrolyte, solution of the bulk component). Based on simple theoretical models, the shapes of the calibration curves are derived and elucidated for all modes, clearly demonstrating that only a few modes can provide useful results while most of them seem useless for quantitation. The knowledge of regularities affecting the character of calibration curves can be used for optimization of the analysis so that reliable results and highest attainable sensitivity can be reached. The outcome of this study brings a clear instruction how to successfully quantitate trace analytes even in samples with a complex and variable matrix*. PMID- 12116127 TI - Modelling, optimisation and control of selectivity in the separation of aromatic bases by electrokinetic chromatography using a neutral cyclodextrin as a pseudostationary phase. AB - A simple mathematical model describing the separation of a series of aromatic bases by electrokinetic chromatography using beta-cyclodextrin (beta-CD) as a pseudostationary phase is described. The model takes into account changes in electrolyte pH and the different formation constants between the neutral and charged forms of the analytes with the CD. Constants in the model were obtained within the two-dimensional experimental space defined by pH and [beta-CD] with nonlinear regression using only five experimental points. These constants agreed with expected trends in analyte-CD interactions and predicted much higher formation constants for the neutral analyte-CD complex than for the charged analyte-CD complex. Correlation between predicted and observed mobilities using additional 20 points within the experimental space gave r(2) = 0.995. Optimisation of the pH and [beta-CD] was performed using both the normalised resolution product and minimum resolution product criteria and provided two optimum separations which exhibited different selectivities. Differences between predicted and observed migration times at these optima were less than 2.5 and 5% for the normalised resolution product and the minimum resolution criteria, respectively. In both cases the correct migration order was predicted. The model was also applied successfully to the optimisation of conditions for the separation of a specific mixture of analytes or for conditions under which particular analytes migrated in a desired order. PMID- 12116128 TI - Characteristics of the electroosmotic flow of electrolyte systems for nonaqueous capillary electrophoresis. AB - Nonaqueous capillary electrophoresis (NACE) is a powerful tool for the analysis of surface-active substances, which represent a broad class of analytes containing cationic and anionic species, such as surfactants, phosphoric acid esters, and amines. In order to conduct an efficient method development in NACE, the influence of the electrolyte composition on the electroosmotic flow (EOF) of organic separation systems was systematically investigated. Background electrolytes and background chromophores appropriate for direct and indirect UV detection were considered, as the majority of surface-active substances do not absorb UV-light. It was found that theoretical models developed to describe the EOF in aqueous electrolyte systems are insufficient for organic electrolyte systems. Experimental data on electroosmosis in a variety of organic solvents and mixtures of methanol and acetonitrile applying different background chromophores and basic or acidic additives are given. Differences between them are discussed with relation to the physicochemical properties of the organic solvents. PMID- 12116129 TI - Electroosmotic pumping in microchips with nonhomogeneous distribution of electrolytes. AB - A general equation to calculate the node pressure at a junction in a microfluidic network is presented. The node pressure is generated from both the hydrodynamic flow due to the external applied hydraulic pressures and the electrokinetic flow resulted from the applied electric field. Pure electroosmotic flow has a plug flow profile and pressure flow has a parabolic flow profile. In a first order approximation, these two flows can be treated separately, and the total flow is the sum of the two. An externally applied pressure simply creates a constant offset in the node pressure as long as the flow resistances remain the same. In a nonhomogeneous microfluidic network, where the electrical resistivity or the electroosmotic mobility is not constant everywhere, the differences in electroosmotic flow in various sections of the network will create an electroosmotically induced pressure at the internal nodes. Our theoretical approach can easily be extended to networks with more than one internal node. One prediction of this theory is that any variation in electroosmotic mobility or solution resistivity in different network branches will generate a pressure, and can thus be used as a pump. As an example, we demonstrate electroosmotic pumping in a high-low buffer system. PMID- 12116131 TI - Capillary electrophoretic detection in apolipoprotein E genotyping. AB - We describe the application of capillary electrophoresis to detect DNA fragments, obtained after amplifying a part of the apolipoprotein E (apoE) gene with polymerase chain reaction (PCR). Compared to conventional agarose slab gel electrophoresis (AGE), CE appears the method of choice with regard to resolution and sensitivity, to detect DNA fragments in the range of 20-100 base pairs. Especially discrimination between apoE2/E2 and apoE2/E3 genotypes is more reliable with CE than with AGE, this being of great clinical value in the diagnosis of familiary dysbetalipoproteinemia. PMID- 12116130 TI - Separation of reboxetine enantiomers by means of capillary electrophoresis. AB - The novel antidepressant reboxetine, a selective norepinephrine reuptake inhibitor, is increasingly used in the treatment of different forms of major depression. Reboxetine is a chiral compound, and is marketed as a racemic mixture of (R,R)- and (S,S)-reboxetine; however, the pharmacokinetic and toxicological profiles of the two enantiomers are rather different. For this reason, a simple capillary electrophoretic method for the separation of reboxetine enantiomers has been developed. Sulfobutyl ether-beta-cyclodextrin was chosen as the chiral selector, and several parameters, such as cyclodextrin and buffer concentration, buffer pH and capillary temperature were investigated in order to obtain good separation and acceptable run times. Using an uncoated, fused-silica capillary (internal diameter 50 microm, total length 48.5 cm, effective length 40.0 cm) and a background electrolyte consisting of a pH 3.0, 100 mM phosphate buffer containing 1.25 mM cyclodextrin, reboxetine enantiomers were baseline separated (resolution > 4) with a voltage of 20 kV in less than 16 min. Since pure enantiomers of reboxetine were not available, they were obtained from the racemic powder by means of direct-phase, high-performance liquid chromatography and their identity confirmed by circular dichroism spectra. PMID- 12116133 TI - Electrolyte and additive effects on enantiomer separation of peptides by nonaqueous ion-pair capillary electrophoresis using tert.-butylcarbamoylquinine as chiral counterion. AB - Nonaqueous ion-pair capillary electrophoresis separations of N-protected (all R)/(all-S) alanine peptide enantiomers with up to six amino acid residues using tert.-butylcarbamoylquinine as selector and employing the partial filling technique are presented. The effects of various conditional parameters on separation were studied, namely chemical nature of the capillary wall, solvent composition of the background electrolyte (BGE), acid-base-ratio (equivalent to apparent pH), ionic strength and selector concentration. The influence of the solvent composition (methanol-ethanol ratios) on resolution turned out to be rather complex. The separation of the peptide enantiomers was strongly altered by small changes in pH and ionic strength. An increase of the selector concentration was found to offer an easy way for enhancing enantioselectivity, although some drawbacks, e.g., elongation of run times, have to be considered. A method was developed that allowed the separation of N-3,5-dinitrobenzoyl oligoalanine enantiomers containing 1-6 amino acid residues in one run. Like in a recent high performance liquid chromatography (HPLC) study, separation selectivity thereby decreased from 1.541 (Ala), 1.340 (Ala(2)), 1.054 (Ala(3)), 1.029 (Ala(4)), 1.024 (Ala(5)) to 1.020 (Ala(6)). In addition, all four stereoisomers of N-2,4 dinitrophenyl- and N-3,5-dinitrobenzyloxycarbonyl-protected alanylalanine could be baseline-resolved. PMID- 12116132 TI - The effect of sodium dodecyl sulfate and Pluronic F127 on the electrophoretic separation of protein and polypeptide test mixtures at acid pH. AB - Using a test mixture consisting of standard proteins (cytochrome c, chymotrypsinogen A, hen egg albumin, bovine serum albumin, aldolase, catalase and ferritin) and synthetic polypeptides (polylysine, polyaspartic, polyglutamic acid and polyproline) it was revealed that using sodium dodecyl sulfate (SDS) as background electrolyte modifier at acid pH (2.5) allows selective separation of highly positively charged polypeptides (polylysine) provided that their relative molecular mass is sufficiently low (3300 Da). The altered elution sequence of standard proteins as compared to a separation done without SDS may help their identification. Addition of Pluronic F127 offers clear-cut separations of standard proteins up to a relative molecular mass of 5 x 10(4) Da and allows to reveal protein/polypeptide microheterogeneity where applicable. None of the systems tested is suitable for the separation of acidic polypeptides and polyproline. PMID- 12116134 TI - Enhanced analysis of purine and pyrimidine bases by the use of double-strand polyaniline coatings in micellar electrokinetic capillary chromatography. AB - A double-strand polymeric complex, which suppresses electroosmotic flow relative to fused-silica, is described. The polymeric complex contains a strand polyaniline (PAN) with the second strand containing polyacrylic acid (PAA) and methacrylate (MA) groups. The complex is referred to as PAN:P(AAMA). This polymeric complex has pH-controlled electroactive and hydrophobic characteristics and can be easily coated onto fused-silica. Enhanced separations of theophylline, theobromine, caffeine and adenine, thymine, uracil and cytosine were obtained by the use of the coated capillary in the micellar electrokinetic capillary electrophoresis (MEKC) system. The purine and pyrimdine bases were separated on the coated capillary with a 20 mM, pH 7 phosphate buffer which contains 0.05 M sodium dodecyl sulfate (SDS) as an additive. PMID- 12116135 TI - Enantioseparation of glutethimide and its 5-OH-metabolite in capillary electrophoresis and study of selector-selectand interactions using one dimensional rotating frame nuclear Overhauser and exchange spectroscopy. AB - Enantioseparation of glutethimide (GT) and its 5-hydroxy metabolite (5-OH-GT) has been studied with several charged cyclodextrin (CD) derivatives. The emphasis was made on the enantiomer migration order of GT and simultaneous enantioseparation of GT and 5-OH-GT. The possible structural differences of GT complexes with three different single isomer charged CD derivatives were studied using one-dimensional rotating frame nuclear Overhauser and exchange spectroscopy (1-D ROESY). PMID- 12116136 TI - Univalent salts as modifiers in micellar capillary electrophoresis. AB - The influence of three univalent salts (LiCl, NaCl and RbCl) on the separation of amino acids labelled with 3-(4-carboxybenzoyl)-quinoline-2-carboxaldehyde (CBQCA) in micellar capillary electrophoresis has been studied. Capacity factors for a series of eight CBQCA-labelled amino acids in a sodium dodecyl sulfate (SDS) micellar system containing different concentrations of salt were measured and were found to be related to both the hydrodynamic radius of the salt counter-ion (Li(+), Na(+), Rb(+)) and the relative hydrophobicity of the amino acid. Affinities of the analytes for the micelles were generally observed to decrease as the salt concentration in the background electrolyte was increased from 10 to 50 mM. This decrease in affinity was greatest in the presence of the salt counter ion with the smallest hydrodynamic radius and is primarily due to an increased resistance to mass transfer. Furthermore, interaction of hydrophobic analytes with the micelles is greater than that of hydrophilic analytes at all salt concentrations due to the greater strength of the hydrophobic interactions and this effect is also enhanced in the presence of a smaller counter-ion. No negative effects due to Joule heating or electromigrative dispersion were observed for low to moderate concentrations of salt, which suggests that the use of simple univalent salts to modify analyte/micelle affinities can be a practical method for improving the separation of complex mixtures. PMID- 12116137 TI - Application of nonaqueous capillary electrophoresis to the simultaneous analysis of anionic surfactants. AB - In aqueous capillary electrophoresis selectivity between different alkyl chain lengths within one anionic surfactant-group markedly exceeds selectivity between different functionalities at a given chain length. Peak identification and quantitative analysis in complex mixtures is almost impossible, especially, if the sample contains ethoxylated surfactants as well. Applying nonaqueous capillary electrophoresis (NACE), significant differences in the mobilities of the various functionalities can be generated to exceed at satisfying separation. In this paper, method development of NACE systems is described and the application of these systems to anionic surfactant analysis in real sample matrices is documented. PMID- 12116138 TI - Determination of nitrite and nitrate in a proposed certified reference material for nutrients in seawater by capillary zone electrophoresis with artificial seawater as the background electrolyte using transient isotachophoresis. AB - We describe a combination of selected ions as a terminating ion which is useful for transient isotachophoresis (ITP) in capillary zone electrophoresis (CZE) for the determination of nitrite and nitrate in seawater. In addition to 150 mM sulfate as the principal terminating ion, 10 mM bromate was added to a sample solution as the additional terminating ion. Artificial seawater containing 3 mM cetyltrimethylammonium chloride (CTAC) was adopted as a background electrolyte (BGE). The limits of detection (LODs) for nitrite and nitrate were 2.2 and 1.0 microg/L (as nitrogen), respectively. The LODs were obtained at a signal to noise ratio (S/N) of 3. The values of the relative standard deviation (RSD) of peak area for these ions were 1.9 and 1.4%. The RSDs of peak height were 1.7 and 1.9%, the RSDs of migration time 0.11%. The proposed method was applied to the determination of nitrite and nitrate in a proposed certified reference material for nutrients in seawater, MOOS-1, distributed by the National Research Council of Canada (NRC). The results almost agreed with the assigned tolerance interval. PMID- 12116139 TI - The free solution mobility of DNA in Tris-acetate-EDTA buffers of different concentrations, with and without added NaCl. AB - The free solution mobility of a high-molecular-weight DNA, linear pUC19, and a 20 bp oligomer called dsA5 have been studied as a function of Tris-acetate-EDTA (TAE) buffer concentration, with and without added NaCl. The two DNAs migrate as separate peaks during capillary electrophoresis, because the mobility of linear pUC19 is higher than that of the 20-bp oligomer. In TAE buffers ranging from 10 400 mM in concentration, the migration times and peak areas of the two DNAs are independent of whether they are electrophoresed separately or in mixtures, indicating that DNA-DNA and DNA-buffer interactions are absent in these solutions. The migration times of the two DNAs vary and the peak areas are not additive when the TAE buffer concentration is reduced to 5 mM or below, indicating that DNA-DNA and DNA-buffer interactions are occurring at very low TAE buffer concentrations. The mobilities of linear pUC19 and dsA5 decrease slowly with increasing conductivity or ionic strength when the conductivity is increased by increasing the TAE buffer concentration. When the Tris buffer concentration is held constant and the conductivity is increased by adding various concentrations of NaCl to the solution, the mobilities of linear pUC19 and dsA5 first increase slightly, then become independent of solution conductivity (or ionic strength), and finally decrease when the NaCl concentration is increased above approximately 50 mM. The mobility variations observed in the various TAE and TAE-NaCl solutions are described qualitatively by Manning's theory, although quantitative agreement is not achieved. The free solution mobilities of single-stranded pUC19 and two 20 base oligonucleotides have also been measured. The free solution mobility of single-stranded pUC19 is approximately 15% lower than that of native pUC19, in agreement with other results in the literature. Somewhat surprisingly, the mobilities of the single- and double-stranded 20-mers are equal to each other in TAE buffers with and without added NaCl. PMID- 12116140 TI - Multivalent weak electrolytes - risky background electrolytes for capillary zone electrophoresis. AB - Multivalent weak acids and bases are useful components of buffers in electrophoresis. The use of such buffers as background electrolytes (BGEs) in capillary zone electrophoresis (CZE) is, however, risky due to the existence of unsafe regions in the analytical window of the separation. This contribution discusses the problems and shows that multivalent weak species in BGEs bring about the same effects as mixtures of two independent co-ions, i.e., the presence of two centers of symmetry in the electropherograms and the existence of a migrating system zone with a mobility in between these two centers of symmetry. The system zone deteriorates the analytical separation and detection of the analytes in its neighborhood. Illustrative experimental examples for both cationic and anionic CZE are shown and related discussion is given. Finally, some basic rules are formulated to avoid the preparation of risky BGEs. PMID- 12116141 TI - Multiple effect of surfactants used as additives in background electrolytes in capillary zone electrophoresis: cetyltrimethylammonium bromide as example of model surfactant. AB - Surfactants are frequently used in the preparation of background electrolytes (BGEs) in capillary zone elcetrophoresis (CZE) in order to affect and to optimize both the electroosmotic flow (EOF) and the separation process. Their effects are, however, always multiple, the resulting situation may be very complex and the separation process may even be destroyed. We use the surfactant cetyltrimethylammonium bromide (CTAB) as a model example and bring experimental results and related discussion which elucidate the multiple effect of surfactants in an integrated way. It is shown that even at concentration levels lower than 10(-4) M CTAB strongly reduces the cathodic EOF in bare fused-silica capillaries and converts it into anodic EOF. The magnitude and polarity of the EOF depends not only on the concentration of CTAB but also on the composition of BGEs used. The interactions of CTA cations with the bare capillary wall reduce sorption of cationic analytes and enables their analysis. CTA cations at levels below their critical micelles concentration (CMC) already interact with anionic analytes and reduce their mobilities. This association is strong with highly charged anions and by this, the reversal of the EOF, applying BGEs with highly charged anions is less effective. These interactions are competitive and also depend on the composition of the BGE used. At levels above its CMC, CTAB forms micelles and enables the application of the micellar electrokinetic capillary chromatography (MEKC) mode and the analysis of, e.g., neutral components. Simultaneously, it is shown that the presence of CTAB may increase the number of potentially formed system zones. PMID- 12116145 TI - Capillary array electrophoresis. PMID- 12116146 TI - Recent progress in DNA analysis by capillary electrophoresis. AB - A number of recent developments in DNA analysis by capillary electrophoresis are here reviewed. They include capillary arrays for fast, parallel DNA sequencing as well as microfabricated capillary arrays. Microfluidic chips for DNA sizing and quantitation are also covered, as well as microdevices containing arrays of regular obstacles acting as size-separators during DNA migration. Screening of DNA point mutations by two much improved techniques is also reported: in one case, such mutations are detected (but only on relative short, ca. 60-70 base long fragments) by free electrophoresis in rather acidic (pH ca. 3) buffers; in the case of single-strand chain polymorphism, an improved technique is described based on near-neutral pH buffers with mixtures of Tris/MES cations/zwitterions. When studying the behavior of inorganic and organic cations in the Debye-Huckel layer of DNA, it was found that the latter (especially a large number of Good's buffers and other zwitterions, such as His) would bind to the DNA filament not only via charge interaction, but also via additional bonds, notably hydrogen bonds, thus altering the electrophoretic (and possibly the biological) behavior of DNA molecules. However, whether or not borate ions would bind to DNA remains still an unsettled question. Finally, capillary electrophoresis was found to be instrumental in measuring fine physicochemical parameters pertaining to DNA polyelectrolytes, such as their free mobility and their translational diffusion coefficients. PMID- 12116147 TI - Technical challenges in applying capillary electrophoresis-single strand conformation polymorphism for routine genetic analysis. AB - Recent and future advances in population genetics will have a significant impact on health care practices and the economics of health care provision only if a spectrum of patient-tailored, effective methods of DNA screening for sequence alterations has been developed. Genetic screening by capillary electrophoresis single strand conformation polymorphism (CE-SSCP), which is based upon the differences in electrophoretic mobilities of wild-type and mutant DNA species, offers an important complement to other presently available techniques such as Sanger sequencing and DNA hybridization arrays due to its simplicity, versatility, and low cost of analysis. A two-part review of CE-SSCP that discusses its advantages and limitations is presented. Emphasis is placed on technological aspects of CE-SSCP (including such rarely addressed issues as sample preparation protocols and the nature of the polymeric DNA separation matrix) as well as on the potential of CE-SSCP for routine genetic analysis. An attempt is made to organize and present the information in sufficient detail to allow the use of SSCP for routine genetic screening even by those inexperienced in CE. Some discussion of CE-based heteroduplex analysis (HA) is also presented. PMID- 12116148 TI - Trends in DNA forensic analysis. AB - The paper gives a historical perspective of forensic DNA analysis and overviews existing technologies implemented in forensic laboratories for DNA profiling. Short tandem repeat analysis, mitochondrial DNA and Y-chromosome analysis are described. The review also focuses on emerging new technologies, which represent an interest for the DNA forensic community. Short tandem repeat analysis, by microelectronic chip device, electrophoretic microdevice and matrix assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry are reviewed in this context. PMID- 12116149 TI - Microchannel DNA sequencing matrices with switchable viscosities. AB - We review the variety of thermo-responsive and shear-responsive polymer solutions with "switchable" viscosities that have been proposed for application as DNA sequencing matrices for capillary and microfluidic chip electrophoresis. Generally, highly entangled polymer solutions of high-molar mass polymers are necessary for the attainment of long DNA sequencing read lengths (> 500 bases) with short analysis times (< 3 h). However, these entangled polymer matrices create practical difficulties for microchannel electrophoresis with their extremely high viscosities, necessitating high-pressure loading into capillaries or chips. Shear-responsive (shear-thinning) polymer matrices exhibit a rapid drop in viscosity as the applied shear force is increased, but still require a high initial pressure to initiate flow of the solution into a microchannel. Polymer matrices designed to have thermo-responsive properties display either a lowered (thermo-thinning) or raised (thermo-thickening) viscosity as the temperature of the solution is elevated. These properties are generally designed into the polymers by the incorporation of moderately hydrophobic groups in some part of the polymer structure, which either phase-separate or hydrophobically aggregate at higher temperatures. In their low-viscosity states, these matrices that allow rapid loading of capillary or chip microchannels under low applied pressure. The primary goal of work in this area is to design polymer matrices that exhibit this responsive behavior and hence easy microchannel loading, without a reduction in DNA separation performance compared to conventional matrices. While good progress has been made, thermo-responsive matrices have yet to offer sequencing performance as good as nonthermo-responsive networks. The challenge remains to accomplish this goal through the innovative design of novel polymer structures. PMID- 12116150 TI - A theoretical study of an empirical function for the mobility of DNA fragments in sieving matrices. AB - The separation of DNA fragments by gel electrophoresis has been studied extensively over the last two decades. More recently, similar studies have been carried out to characterize the separation achieved by the current capillary array electrophoresis systems and their sieving polymer solutions. In all cases, at least three different mobility regimes have been shown to exist: the Ogston regime when the radius of gyration of the DNA fragment is smaller than the pore size, the reptation regime when the DNA is larger than the pore size but remains in a random coil conformation, and finally the reptation-with-orientation regime where the DNA orients in the field direction and essentially all resolution is lost. Unfortunately, although theory helps us understand the different regimes and how to properly exploit them, we still have no theory-based general equations that would apply to all regimes. Such equations would be especially useful to analyze data, optimize separation systems and interpolate mobilities to estimate unknown molecular sizes. Recently, van Winkle, Beheshti and Rill (Electrophoresis 2002, 23, 15-19) proposed an intriguing empirical formula that seems to adequately fit the mobility of dsDNA fragments across all three regimes. In this paper, I investigate the relation between this empirical formula and the known theories of gel electrophoresis, and I study the dependence of its fitting parameters upon the experimental conditions. Finally, I examine how this equation may need to be modified to capture the more subtle details predicted by fundamental theories of DNA gel electrophoresis. PMID- 12116151 TI - Theory of sequence effects on DNA translocation through proteins and nanopores. AB - A general formula for the sequence effects on the average translocation time of a polymer passing through a narrow hole under a chemical potential gradient is provided. The utility of the general expression is illustrated by considering the diblock sequence of a polymer with two kinds of monomers. The experimental conditions required for sufficient discrimination of various sequences are addressed. PMID- 12116152 TI - DNA sequencing with hydrophilic and hydrophobic polymers at elevated column temperatures. AB - Read length in DNA sequencing by capillary electrophoresis at elevated temperatures is shown to be greatly affected by the extent of hydrophobicity of the polymer separation matrix. At column temperatures of up to 80 degrees C, hydrophilic linear polyacrylamide (LPA) provides superior read length and separation speed compared to poly(N,N-dimethylacrylamide) (PDMA) and a 70:30 copolymer of N,N-dimethylacrylamide and N,N-diethylacrylamide (PDEA30). DNA polymer and polymer intramolecular interactions are presumed to be a major cause of band broadening and the subsequent loss of separation efficiency with the more hydrophobic polymers at higher column temperatures. With LPA, these interactions were reduced, and a read length of 1000 bases at an optimum temperature of 70 degrees -75 degrees C was achieved in less than 59 min. By comparison, PDMA produced a read length of roughly 800 bases at 50 degrees C, which was close to the read length attained in LPA at the same temperature; however, the migration time was approximately 20% longer, mainly because of the higher polymer concentration required. At 60 degrees C, the maximum read length was 850 bases for PDMA, while at higher temperatures, read lengths for this polymer were substantially lower. With the copolymer DEA30, read length was 650 bases at the optimum temperature of 50 degrees C. Molecular masses of these polymers were determined by tandem gel permeation chromatography-multiangle laser light scattering method (GPC-MALLS). The results indicate that for long read, rapid DNA sequencing and analysis, hydrophilic polymers such as LPA provide the best overall performance. PMID- 12116154 TI - Comb-like copolymers as self-coating, low-viscosity and high-resolution matrices for DNA sequencing. AB - Comb-like copolymers with a polyacrylamide backbone and poly(N,N dimethylacrylamide) grafts were prepared, as a way to combine the superior sieving properties of polyacrylamide with the self-coating properties of polydimethylacrylamide. These matrices function well in the absence of a capillary coating, and achieve separation performances for single-stranded DNA that are comparable to those of state-of-the-art long-chain linear polyacrylamide. Structural parameters such as the grafting density and the polymer molecular mass were varied, and good performance appears to be achieved with a relatively large range of parameters. Surprisingly, excellent separation is achieved even with matrices that have a viscosity as low as 200 mPa/s. A discussion of the physics underlying this behavior is provided. PMID- 12116153 TI - Poly-N-hydroxyethylacrylamide (polyDuramide): a novel, hydrophilic, self-coating polymer matrix for DNA sequencing by capillary electrophoresis. AB - A replaceable polymer matrix, based on the novel monomer N-hydroxyethylacrylamide (HEA), has been synthesized for application in DNA separation by microchannel electrophoresis. The monomer was found by micellar electrokinetic chromatography analysis of monomer partitioning between water and 1-octanol to be more hydrophilic than acrylamide and N,N-dimethylacrylamide. Polymers were synthesized by free radical polymerization in aqueous solution. The weight-average molar mass of purified polymer was characterized by tandem gel permeation chromatography multiangle laser light scattering. The steady-shear rheological behavior of the novel DNA sequencing matrix was also characterized, and it was found that the viscosity of the novel matrix decreases by more than 2 orders of magnitude as the shear rate is increased from 0.1 to 1000 s(-1). Moreover, in the shear-thinning region, the rate of change of matrix viscosity with shear rate increases with increasing polymer concentration. Poly-N-hydroxyethylacrylamide (PHEA) exhibits good capillary-coating ability, via adsorption from aqueous solution, efficiently suppressing electroosmotic flow (EOF) in a manner comparable to that of poly-N,N dimethylacrylamide. Under DNA sequencing conditions, adsorptive PHEA coatings proved to be stable and to maintain negligible EOF for over 600 h of electrophoresis. Resolution of DNA sequencing fragments, particularly fragments > 500 bases, in PHEA matrices generally improves with increasing polymer concentration and decreasing electric field strength. When PHEA is used both as a separation matrix and as a dynamic coating in bare silica capillaries, the matrix can resolve over 620 bases of contiguous DNA sequence within 3 h. These results demonstrate the good potential of PHEA matrices for high-throughput DNA analysis by microchannel electrophoresis. PMID- 12116155 TI - Cross-linked polyacrylamide gel electrophoresis of single-stranded DNA for microfabricated genomic analysis systems. AB - Microfabricated devices are poised to offer inexpensive self-contained alternatives to conventional benchtop-scale laboratory equipment for performing a variety of important DNA analysis assays. In order to realize the dramatic cost savings possible through photolithographic fabrication techniques, these devices must occupy an extremely compact footprint on the silicon wafer. This requirement implies that electrophoretic separations must be performed over ultrashort distances. Employing cross-linked polyacrylamide gels in place of conventional uncross-linked sieving media offers a convenient strategy to achieve this goal. In this paper, we show how the increased resolving power offered by cross-linked polyacrylamide gels, along with improved sample injection techniques, can be exploited to enhance separation performance in microscale systems. We use these techniques to perform high-resolution gel electrophoresis of single-stranded DNA fragments in microfabricated devices over separation distances of 1.5 cm or less. The results presented here are in agreement with theoretical predictions and suggest that it is possible to perform DNA sequencing on compact microchips. More importantly, the separation performance demonstrated in this work is already more than adequate to perform a number of important genomic assays imposing less stringent resolution requirements than sequencing. Successfully adapting even a few of these assays to the microdevice format has the potential to provide a new generation of inexpensive and portable devices suitable for direct end-user applications. PMID- 12116156 TI - Separation of double-stranded DNA fragments by capillary electrophoresis using polyvinylpyrrolidone and poly(N,N-dimethylacrylamide) transient interpenetrating network. AB - A noncross-linked interpenetrating polymer network (IPN), consisting of poly(N,N dimethylacrylamide) (PDMA) and polyvinylpyrrolidone (PVP, weight-average molecular weight M(w) = 1 x 10(6) g/mol) was synthesized by polymerizing N,N dimethylacrylamide (DMA) monomers directly in PVP buffer solution and tested as a separation medium for double-stranded (ds)DNA analysis without further purification. Due to the incompatibility of PVP and PDMA, a simple solution mixture could incur a microphase separation and showed poor performance on dsDNA separation. However, a dramatic improvement was achieved by the formation of an IPN. We attributed the high sieving ability of IPN as due to an increase in the number of entanglements by the more extended polymer chains. Apparent viscosity studies showed that the IPN had a much higher viscosity than the simple mixture containing the same amount of PDMA and PVP. In 1 x Tris-borate-EDTA (TBE) buffer, the concentration ratio of PDMA and PVP had a great effect on the DNA separation. At optimal conditions, the 22 fragments in pBR322/HaeIII DNA were successfully separated within 15 min, with a resolution of better than 1.0 for 123/124 bp. PMID- 12116157 TI - Multiplex high-throughput solid-phase minisequencing by capillary electrophoresis and liquid core waveguide fluorescence detection. AB - Minisequencing, solid-phase single-nucleotide primer extension reaction, is a robust method for performing multiplex single-nucleotide polymorphism (SNP) analysis. We have combined this technology with capillary gel electrophoresis in a multicapillary format, using liquid core waveguide (LCW) fluorescence detection. Polymerase chain reaction (PCR) amplification of multiple DNA targets is performed with one primer for each target biotinylated. Separation of the complementary strands, minisequencing and washing steps are carried out using streptavidin-coated magnetic beads. Dideoxynucleotides analogues labelled with different fluorophores are used for the extension of the minisequencing primers. The extended oligonucleotides, the length of which defines the position on the target and the color the identity of the polymorphism, are then separated in a gel-filled array of capillaries, coated on the outside with a layer of a fluoropolymer to provide the liquid core waveguide characteristics. The technology has a potential for extremely high throughputs when a combination of multiplex PCR-minisequencing is used together with a large array of capillaries, four-color detection and high-speed separation. PMID- 12116158 TI - Mono- and bis-intercalating dyes for multiplex fluorescence lifetime detection of DNA restriction fragments in capillary electrophoresis. AB - The use of novel intercalating dyes as labels in DNA restriction fragment analysis by capillary electrophoresis with frequency-domain fluorescence lifetime detection is described. The dyes, including one mono-intercalating dye with three positive charges and three bis-intercalating, homodimeric dyes with four positive charges, were excited by the 488 nm line of an argon ion laser and exhibited lifetimes in the range of 1-3 ns. The separations were performed using a gel containing 1% high-molecular-weight (HMW) hydroxyethylcellulose (HEC) (90,000 105,000) and 0.3% low-molecular-weight (LMW) HEC (24,000-27,000) in Tris-borate EDTA buffer (TBE). Multiplex lifetime detection of mixtures of dye-labeled DNA restriction fragment digests and size standard fragments was achieved. Compared to previous results obtained with several mono-intercalating dyes of lesser charge (McIntosh, S. L., Nunnally, B. K., Nesbit, A. R., Deligeorgiev, T. G., Gadjev, N. I., McGown, L. B., Anal. Chem. 2000, 72, 5444-5449), the present dyes provided a wider range of lifetimes and better lifetime discrimination in multiplex detection. There was no evidence of dye exchange during the capillary electrophoresis experiment. PMID- 12116159 TI - Optimization of sequencing conditions using near-infrared lifetime identification methods in capillary gel electrophoresis. AB - We have investigated the sample preparation and electrophoresis conditions necessary to prepare DNA sequencing samples appropriate for use with near infrared (IR) fluorescent labels with dye identification accomplished via lifetime techniques. It was found that several sample preparation protocols required attention to maximize the fluorescence yields of the labeling dyes, such as thermal cycling conditions, choice of counter ion used for the ethanol precipitation step and also, dye-primer versus dye-terminator chemistries. In addition, several different sieving matrices were investigated for their effects on both the fluorescence properties of the labeling dyes and electrophoretic resolution. Extended times used for the high temperature denaturing of duplexed DNA fragments during cycle sequencing produced cleavage products, in which the covalently attached dye to the sequencing primer was released through attack by dithiothreitol (DTT). Even under optimized thermal cycling conditions, free dye was generated that masked readable data from the sequencing traces. Ethanol precipitation was necessary to remove this free dye with the proper choice of counter ion (sodium). The results using different sieving matrices indicated that linear polyacrylamides (LPAs) were appropriate for any fluorescence measurement, since they could readily be replaced between runs minimizing deleterious memory effects associated with cross-linked polyacrylamide gels. After investigation of several different sieving LPAs, the commercially available POP6 was found to be particularly attractive, since it produced good electrophoretic resolution, single exponential behavior for the near-IR dye series investigated herein, and also, discernible lifetime differences within the dye set. Finally, dye terminator chemistry was also found to minimize bleeding in the gel matrix produced by large amounts of unextended dye-primer within the gel lane. PMID- 12116160 TI - Two-color excitation system for fluorescence detection in DNA sequencing by capillary array electrophoresis. AB - Two computer-controlled galvanometer scanners are adapted for two-dimensional step scanning across a 96-capillary array for laser-induced fluorescence detection. 488 nm and 514 nm laser lines from the same Ar(+) laser were alternately coupled for two-color excitation in each capillary. The signal at a single photomultiplier tube is temporally sorted to distinguish among the capillaries and the excitation wavelengths. Based on the differences in absorption spectra for the dyes, the peak-height ratios in the 488 nm and 514 nm excitation electropherograms were used for peak identification for multiplexed capillary electrophoresis. Successful base calling for 24-capillary DNA sequencing was achieved to 450 bp with 99% accuracy. Advantages include the efficient utilization of light due to the high duty-cycle of step scan, good detection performance due to the reduction of stray light, ruggedness due to the small mass of the galvanometer mirror, low cost due to the simplicity of components and flexibility due to the independent paths for excitation and emission. PMID- 12116162 TI - Direct haplotype detection of adjacent polymorphic sites in the regulatory region of the dopamine D4 receptor (DRD4) gene. AB - A novel method of haplotype identification is discussed allowing simultaneous detection of two adjacent polymorphic sites, a single nucleotide polymorphism (SNP) and a length polymorphism within 1-2 kilobase distance. The method combines allele specific-amplification with high-throughput, automated ultrathin-layer gel electrophoresis analysis of fragment size polymorphism. A typical application is shown for genotyping the -521 C/T single nucleotide polymorphism and the 120 bp duplication in the 5"-upstream region of the dopamine D4 receptor (DRD4) gene. We have also demonstrated that the haplotypes of double heterozygotes for -521 C/T and for the 120 bp duplication can be clearly distinguished, that has only been possible previously by extensive pedigree analysis. PMID- 12116161 TI - Integrated platform for detection of DNA sequence variants using capillary array electrophoresis. AB - We have developed a highly versatile platform that performs temperature gradient capillary electrophoresis (TGCE) for mutation/single-nucleotide polymorphism (SNP) detection, sequencing and mutation/SNP genotyping for identification of sequence variants on an automated 24-, 96- or 192-capillary array instrument. In the first mode, multiple DNA samples consisting of homoduplexes and heteroduplexes are separated by CE, during which a temperature gradient is applied that covers all possible temperatures of 50% melting equilibrium (Tms) for the samples. The differences in Tms result in separation of homoduplexes from heteroduplexes, thereby identifying the presence of DNA variants. The sequencing mode is then used to determine the exact location of the mutation/SNPs in the DNA variants. The first two modes allow the rapid identification of variants from the screening of a large number of samples. Only the variants need to be sequenced. The third mode utilizes multiplexed single-base extensions (SBEs) to survey mutations and SNPs at the known sites of DNA sequence. The TGCE approach combined with sequencing and SBE is fast and cost-effective for high-throughput mutation/SNP detection. PMID- 12116163 TI - Single-strand conformation polymorphism for p53 mutation by a combination of neutral pH buffer and temperature gradient in capillary electrophoresis. AB - A large number of point mutations in the p53 gene have been detected by capillary zone electrophoresis via single-strand conformation polymorphism (SSCP) analysis. A much improved detection sensitivity was obtained via the following modifications in running conditions: use of low-viscosity 3% hydroxyethylcellulose (HEC), a neutral pH (pH 6.8) buffer, in which the standard Tris moiety was substituted with a 2-(N-morpholino)ethanesulfonic acid (MES)/Tris mixture, use of SYBR Green II for improved fluorescent signal at the lower pH adopted; and, finally, the use of a temperature gradient in the 15-25 degrees C interval, for favoring the conformational transitions in the mutated samples. The typical temperature gradient activated had a slope of 2 degrees C/min and were induced externally. A total of 24 samples from affected patients, both in the homo- and heterozygous state, were analyzed. All the mutations could be detected by this improved protocol, raising the sensitivity from the standard ca. 80% of conventional SSCP to essentially 100% with the present methodology. All the mutations were confirmed by sequence analysis of the affected samples. PMID- 12116164 TI - Membrane-mediated ultrafast restriction digestion and subsequent rapid gel microchip electrophoresis of DNA. AB - Ultrafast, membrane-mediated restriction digestion of DNA molecules followed by rapid gel microchip electrophoresis of the resulting fragments is described. Combination of restriction endonuclease digestion on small pore-size microfibrous membranes with sample loading and electrophoresis analysis in a multilane (up to 96) format resulted in very fast restriction digest based microscale DNA analysis. Complete digestion of several nanogram target DNA was accomplished on the microporous membrane at room temperature just in a few minutes with a single or a combination of various restriction enzymes, using only submicroliter quantities of samples and reagents. The reaction mixture containing membrane also served as sample loading device for the subsequent gel microchip electrophoresis based analysis. This work establishes methods for high-speed, high-throughput DNA analysis, featuring extremely low sample and reagent consumption, and fast restriction digestion in combination with sample loading and rapid gel microchip analysis of the resulting fragments. The entire restriction digestion, sample loading and electrophoresis analysis process required less than 20 min. PMID- 12116165 TI - A nanoliter rotary device for polymerase chain reaction. AB - Polymerase chain reaction (PCR) has revolutionized a variety of assays in biotechnology. The ability to implement PCR in disposable and reliable microfluidic chips will facilitate its use in applications such as rapid medical diagnostics, food control testing, and biological weapons detection. We fabricated a microfluidic chip with integrated heaters and plumbing in which various forms of PCR have been successfully demonstrated. The device uses only 12 nL of sample, one of the smallest sample volumes demonstrated to date. Minimizing the sample volume allows low power consumption, reduced reagent costs, and ultimately more rapid thermal cycling. PMID- 12116166 TI - Analysis of DNA polymorphisms on the human Y-chromosome by microchip electrophoresis. AB - Validation of microchip electrophoresis in DNA analysis has been carried out using an Agilent 2100 Bioanalyzer. With a DNA 500 Assay Kit, the reproducibility and accuracy of fragment sizing of a 10 bp DNA ladder have been shown to be satisfactory with the relative standard deviation and the relative error mostly below 1.0 and 5.0% (n = 12), respectively. Both intraday and interday validations of fragment sizing and quantitation have also been performed with a 7500 Assay Kit (n = 48). Although the results of quantitation are not as good as that of sizing, due to the manual introduction of samples and markers into the chip wells, they are still sufficient to carry out further analyses of practical samples. Based on such reliable results, fast analysis of DNA polymorphisms on the human Y-chromosome has been realized with microchip electrophoresis. The total analysis times of three genomic polymorphisms on the Y-chromosome, Y Alu polymorphism, 47z/StuI, and 12f2, are all within 100 s, and the relative standard deviation and relative error of fragment sizes are below 3.5 and 3.7%, respectively. In addition, a mixture of nine DNA markers on the human Y chromosome related to examine the cause of spermatogenic failure have been separated successfully with the smallest fragment size difference of 7 bp. Our results demonstrate the potential of microchip electrophoresis in polymorphism analysis with the advantages of high speed, good reproducibility, high precision, and high resolution. PMID- 12116167 TI - Microelectronic array devices and techniques for electric field enhanced DNA hybridization in low-conductance buffers. AB - A variety of electronic DNA array devices and techniques have been developed that allow electric field enhanced hybridization to be carried out under special low conductance conditions. These devices include both planar microelectronic DNA array/chip devices as well as electronic microtiter plate-like devices. Such "active" electronic devices are able to provide controlled electric (electrophoretic) fields that serve as a driving force to move and concentrate nucleic acid molecules (DNA/RNA) to selected microlocation test-sites on the device. In addition to ionic strength, pH, temperature and other agents, the electric field provides another controllable parameter that can affect and enhance DNA hybridization. With regard to the planar microelectronic array devices, special low-conductance buffers were developed in order to maintain rapid transport of DNA molecules and to facilitate hybridization within the constrained low current and voltage ranges for this type of device. With regard to electronic microtiter plate type devices (which do not have the low current/voltage constraints), the use of mixed buffers (low conductance upper chamber/high conductance lower chamber) can be used in a unique fashion to create favorable hybridization conditions in a microzone within the test site location. Both types of devices allow DNA molecules to be rapidly and selectively hybridized at the array test sites under conditions where the DNA in the bulk solution can remain substantially denatured. PMID- 12116168 TI - Kinetics of heterogeneous hybridization on indium tin oxide surfaces with and without an applied potential. AB - The rate of hybridization of oligonucleotide target sequences to chemically immobilized oligonucleotide probes has been studied both with and without an electrical field. The probe size was 20-24 nucleotides (nt) while the target size ranged from 157 to 864 nt. In agreement with previous studies, complete hybridization under normal conditions required 10-30 hours, depending on target size. The kinetics were characterized by a characteristic lag time followed by an asymptotic rise to the final value. In contrast, with an applied electrical field, all but the largest target hybridized in about 10 min while the longest hybridized within 1 h. Deleterious electrode reactions were avoided by close spacing of the anode and cathode and application of very small voltages. Our results suggest that probes and targets orient flat on the surface. A model is suggested to explain the kinetics observed that involves a series of surface states between initial target arrival and final hybridized state. Our results show that the electric field accelerated hybrid capture of solution-phase targets by surface-bound probes. This approach may have implications for enhancing array based hybrid capture for mutation detection, copy number determination and/or gene expression profiling. PMID- 12116170 TI - Genetics of diabetes mellitus. PMID- 12116171 TI - Interplay between heredity and environment in the recent explosion of type 1 childhood diabetes mellitus. AB - The fast increase in the incidence of childhood type 1 diabetes mellitus (T1DM) that cannot be explained by changes in the genetic susceptibility, led us to look for environmental causes. To test the hypothesis that the initiation of the autoimmune process of childhood T1DM in genetically susceptible subjects begins in the perinatal period by a viral infection, we studied the seasonal variations in the month of birth of several cohorts of patients compared to the general population. Population groups with high or low T1DM incidence were analyzed separately by t-test and the Cosinor methods. In areas with populations with a high incidence (Israeli Jews, Sicily, Sardinia, Slovenia, Germany) we found that the children (in Sicily also young adults) who subsequently developed T1DM, have a higher incidence of births in the summer months than in other seasons of the year, a mirror image of the seasonality of the clinical onset of disease. This pattern differed significantly from the seasonality of the total live births in the same populations. In populations with a low T1DM incidence, (China, Japan and Cuba) no seasonality of month of birth was found. Similar findings have been reported, from five counties in the U.K. and the Netherlands. It is hypothesized that mothers who become pregnant during the period of yearly viral epidemics transmit to the fetus, either a virus or antiviral antibodies, which determine whether an autoimmune process against the pancreatic beta is initiated or whether the fetus is protected against that process. PMID- 12116172 TI - Impact of genetic and non-genetic factors in type 1 diabetes. AB - Type 1 insulin-dependent diabetes is due to destruction of the insulin secreting cells of the islets of Langerhans. The disease is caused by non-genetic, probably environmental, factors operating in a genetically susceptible host to initiate a destructive immune process. These unknown environmental factors may operate over a limited period either in early or later and to a variable degree, playing a particularly substantial role in adults. The environment then induces an immune process associated with destruction of the islet beta cell that can be detected in early life and persists up to disease onset. Apart from an association with the insulin gene there is no evidence that genes associated with type 1 diabetes, including HLA and CTLA4 influence the targeting of the immune response to the insulin-secreting cells. The critical period of immune activation is probably short and the process leading to diabetes probably has a long prodrome but of variable duration that determines the age at presentation with clinical disease. The amplification both of this immune response and the destructive process is in part genetically determined, involving HLA genes. The clinical spectrum of the disease process associated with type 1 diabetes is wide, encompassing insulin dependence, non-insulin dependence and even transient impaired glucose tolerance. Type 1 diabetes presenting in adults, in contrast to children, is predominantly determined by non-genetic factors with a reduced role for protective and susceptibility HLA alleles. Thus, the evidence is that genes involved in genetic susceptibility to type 1 diabetes operate predominantly in children not adults and in both amplify the immune response and the rate of disease progression. PMID- 12116173 TI - Environmental factors in the etiology of type 1 diabetes. AB - Type 1 diabetes is considered to be an autoimmune disease in which T lymphocytes infiltrate the islets of pancreas and destroy the insulin producing beta cell population. Besides antigen specificity, the quality of immune reactivity against islet cell antigen(s) is an important determinant of the beta cell destruction. Much evidence indicates that the function of the gut immune system is central in the pathogenesis, as the regulation of the gut immune system may be aberrant in type 1 diabetes. The role of virus infections in the pathogenesis of type 1 diabetes has been supported by substantial new evidence suggesting that one virus group, enteroviruses, may trigger the beta-cell damaging process in a considerable proportion of patients. The latest evidence comes from studies indicating the presence of viral genome in diabetic patients and from prospective studies confirming epidemiological risk effect. If this association holds still true in ongoing large-scale studies, intervention trials should be considered to confirm causality. Of the dietary putative etiological factors, cow's milk proteins have received the main attention. Many studies indicate an association between early exposure to dietary cow's milk proteins and an increased risk of type 1 diabetes. The question will be answered by a large scale, prospective, randomized, international intervention trial. Another dietary factor in need of more studies is the deficiency of vitamin D. Among toxins, N-nitroso compounds are the main candidates. An interaction of genetic and environmental factors is important in evaluating the possible role of a certain environmental factor in the etiology of type 1 diabetes. PMID- 12116174 TI - Estimation of genetic risk for type 1 diabetes. AB - The most important gene loci defining risk of type 1 diabetes mellitus (T1DM) are located within the HLA gene region. HLA-DQ molecules are of primary importance but HLA-DR gene products modify the risk conferred by HLA-DQ. The risk associated with an HLA genotype is defined by the particular combination of susceptible and protective alleles. The highest risk is associated with a combination of two different risk haplotypes (7% risk to develop T1DM in Finland) whereas protective genotypes covering 69% of population have a risk of less than 0.2%). The complicated analysis of HLA genotypes is simplified by strong linkage disequilibrium between HLA-DRB1, -DQA1 and -DQB1 loci. In many cases one can deduce the alleles of other loci based on determination of the alleles in one locus. Differences between various populations in the frequency of marker alleles and in the linkages between them has to be taken into account. We have developed PCR based typing methods that utilize blood spot samples, microtiter plate format and lanthanide labeled oligonucleotide probes to define HLA-DQ and -DR alleles relevant for T1DM risk. Typing is run stepwise so that after initial HLA-DQB1 typing only those samples will be further analyzed in which -DQA1 or -DRB1 typing is informative and expected to contribute to the risk estimation. This method has been used to screen more than 50,000 newborn infants in Finland over a time period of 6 years, and it has been able to identify most children who have developed T1D during the follow-up period. The efficiency of the procedure has also been tested in Finnish and Greek populations. PMID- 12116175 TI - Molecular properties of HLA-DQ alleles conferring susceptibility to or protection from insulin-dependent diabetes mellitus: keys to the fate of islet beta-cells. AB - The major histocompatibility complex Class II alleles, HLA-DQ, and the related HLA-DR, are the chief genetic elements of human type 1 diabetes. These genes code for polymorphic heterodimeric proteins, whose chief function is to trap peptide antigens in the endosome and present them on the surface of antigen-presenting cells (dendritic cells, B lymphocytes, monocytes/macrophages) to CD4(+) T helper cells. A systematic investigation of the molecular properties of HLA-DQ alleles linked to susceptibility or resistance to type 1 diabetes has shown that these properties segregate along lines of susceptibility or resistance. A correlation of these features with the function of each particular segment of the HLA-DQ molecule yields interesting insights into the possible pathways leading to type 1 diabetes. There remain, however, areas to be clarified, including mechanisms by which dominant protection is conferred by certain alleles, the interplay between HLA-DQ and the related locus HLA-DR, that also shows autoantigen-specific reactivity, and the cross-Class help delivered to CD8(+) T cells, the final effectors in pancreatic beta-cell destruction. Clarification of these issues may lead to ways to prevent diabetes in predisposed individuals already exhibiting the genetic and immunological characteristics, and perhaps a cure in those with the disease, by means of transplantation, and measures for prevention of disease recurrence. PMID- 12116176 TI - Humoral beta-cell autoimmunity in relation to HLA-defined disease susceptibility in preclinical and clinical type 1 diabetes. AB - We have studied the relationship between diabetes-associated autoantibodies and various HLA genotypes and alleles in patients with newly diagnosed type 1 diabetes, in non-diabetic siblings and in children representing the general population to test the hypothesis that specific HLA genes regulate the humoral immune response to various autoantigens. Among the newly diagnosed patients we observed that those carrying the DR4-DQB1*0302 haplotype had increased levels of insulin autoantibodies (IAA) and IA-2 antibodies (IA-2A), but low levels of GAD antibodies (GADA). In contrast, those with the DR3-DQB1*02 haplotype had increased GADA titers but low IAA and IA-2 levels. DQB1*02-homozygous patients had a conspicuously low frequency and low levels of IA-2A. Among the siblings there was an apparent association between genetic risk and the prevalence of islet cell antibodies (ICA), IAA, GADA, IA-2A and multiple autoantibodies, the latter being detectable at a frequency of 24.1% in high risk siblings and at a frequency of only 0.9% in those with genotypes conferring disease protection. Among 7-16-year-old Finnish schoolchildren there was an association between GADA and the DQB1*02/*0302 genotype and the DQB1*0302 allele. Among young children identified from the general population based on high (DQB1*02/0302 heterozygosity) or moderate (DQB1*0302/x; x not equal *02, *0301, *0602, *0603) genetic risk for type 1 diabetes the high risk children seroconverted more often to positivity for ICA, GADA and IA-2A over their first 2 years of life. Among older sibs of these children we observed an obvious relationship between the degree of genetic risk and the frequency of ICA, IAA, GADA, IA-2A, and multiple antibodies. Taken together these observations suggest that HLA genes have a strong impact on the appearance of diabetes-associated autoantibodies both in first-degree relatives of affected children and in the general population. In patients with newly diagnosed disease IA-2A may be a more specific marker of beta cell damage, whereas GADA might reflect a propensity to autoimmunity in general. PMID- 12116177 TI - Combined genome and proteome approach to identify new susceptibility genes. AB - Type 1 diabetes mellitus (T1DM) is a multifactorial disorder characterized by a specific destruction of the insulin-producing beta cells in the islets of Langerhans. Cells from the immune system infiltrate the islet during the pathogenesis, releasing a mixture of cytokines demonstrated to be specifically toxic to the beta cells within the islets. The goal is to understand the molecular mechanisms responsible for this specific beta-cell toxicity, which will allow the design of novel intervention strategies for T1DM. The proteome approach provides a detailed picture of the beta-cell proteins changing expression pattern during cytokine-mediated beta-cell destruction. Combining the information from this proteome approach with genetic studies makes us believe that it is possible to reach this goal. PMID- 12116178 TI - Wolfram syndrome: identification of a phenotypic and genotypic variant from Jordan. AB - Wolfram syndrome is an autosomal recessive disorder with probable locus heterogeneity. Only insulin-dependent diabetes mellitus and progressive optic nerve atrophy are necessary to make the diagnosis, but associated findings include diabetes insipidus, sensorineural hearing loss, ataxia, peripheral neuropathy, urinary-tract atony, and psychiatric illnesses. We performed clinical and molecular studies on four consanguineous families with 16 affected individuals. We point out a new phenotypic variant with absent diabetes insipidus, presence of peptic ulcer disease and bleeding tendency secondary to a platelet aggregation defect. The same phenotypic variant turned out to be a genotypic variant with linkage to a second Wolfram syndrome locus (WFS2) on chromosome 4q22-24. PMID- 12116179 TI - Mitochondrial diabetes: pathophysiology, clinical presentation, and genetic analysis. AB - This study provides a compact overview on the most common form of the maternally inherited diabetes and deafness syndrome (MIDD) that associates with an A-G mutation in mitochondrial DNA at position 3243 in the tRNA(Leu,UUR) gene. The pathobiochemistry and pathophysiology is discussed. The mutation leads predominantly to a reduced insulin secretion by beta cells in response to glucose stimulation, however, without marked involvement of autoimmune processes as seen in type 1 diabetes mellitus. The underlying biochemical mechanism leading to beta cell dysfunction is discussed. Furthermore, the clinical presentation of the disease is summarized as are the methods to detect the A3243G mutation, particular in view of the often low levels of heteroplasm of the A3243G mutation. PMID- 12116180 TI - Possible association of a cholecystokinin promoter variant to schizophrenia. AB - Several lines of research indicate a cholecystokinin (CCK) deficit in schizophrenia patients. A C to T substitution was found in the promoter region of the CCK gene. We investigated this promoter variant in patients with schizophrenia and geographically-matchedcontrols. The T allele was detected in 24% of the 85 schizophrenics and 16% of the 247 controls. No significant difference in the T allele frequency was found between patients and controls (chi(2) = 2.77, P > 0.1). The schizophrenia sample was analyzed further along the dimensions of positive and negative symptoms. The patients with prominent negative symptoms presented a statistically significant association to the T allele (chi(2) = 4.13, P < 0.04). However, the significance disappeared after the Bonferroni correction (P > 0.15). Since the case-control analysis may present incorrect ethnic match between cases and controls, we applied the family-based tests to verify the above findings. Both transmission disequilibrium test (TDT; chi(2) = 5.33, P < 0.025 in 12 trios) and haplotype relative risk (HRR; chi(2) = 3.844, P < 0.05 in 60 trios) indicated a significantly high transmission of T allele to schizophrenia offspring probands from their parents. While our family based tests seem to support the CCK involvement in schizophrenia, no definite conclusion can be drawn based on such a small sample size. This preliminary finding is subjected to future investigations. PMID- 12116181 TI - Heritability and number of quantitative trait loci of neurocognitive functions in families with schizophrenia. AB - Despite evidence for several chromosomal loci linked to schizophrenia, no susceptibility genes have been identified for the disorder. Using quantitative measures of phenotypic affection in place of clinical diagnostic categories or dichotomous classification of the affection status may be more effective in searching for susceptibility genes. Neurocognitive traits have been suggested as putative quantitative endophenotypes of the disorder, but their heritability estimates are not well known. We investigated the heritability of working memory, verbal declarative memory and its different components, and both verbal and visual ability functions in schizophrenia families with a well-ascertained pedigree structure. We also estimated the number of quantitative trait loci (QTLs) contributing to these neurocognitive functions. Additive genetic heritability of the neurocognitive functions was estimated in a sample of schizophrenia patients and their first-degree relatives (N = 264) from an isolated geographical subregion in Finland. The number of QTLs was analyzed using Markov chain Monte Carlo segregation analysis. Significant heritabilities were found in working memory and ability functions. Furthermore, the working memory functions revealed the most restricted number of QTLs. The mean numbers of loci for verbal and visual working memory were 1.2 and 1.0, respectively, with corresponding posterior probabilities of 73% and 70% for at least one locus. In declarative memory variables, the number of loci was more dispersed. Our results suggest that neurocognitive measures, particularly working memory, may provide valid quantitative phenotypic traits for linkage analyses searching predisposing genes for schizophrenia. PMID- 12116182 TI - Schizophrenia and functional polymorphisms in the MAOA and COMT genes: no evidence for association or epistasis. AB - Several lines of evidence suggest that psychosis is associated with altered dopaminergic neurotransmission. Dopamine is catabolized by monoamine oxidase (MAO) and catechol-O-methyl transferase (COMT). We hypothesized that the genes encoding MAOA and COMT might contain genetic variation conferring increased risk to schizophrenia. In order to test this hypothesis, we genotyped the 941T > G and the promoter VNTR polymorphisms in the MAOA gene and the V158M COMT polymorphism in 346 DSMIV schizophrenics and 334 controls. We also genotyped the-287A > G COMT promoter polymorphism in 177 schizophrenics and 173 controls. No significant differences were found in allele or genotype frequencies between affecteds and controls for any of the polymorphisms. As both genes are involved in degrading catecholamines, we also sought evidence for additive and epistatic effects but none was observed. Our data, therefore, do not support the hypothesis that genetic variation in MAOA and COMT is involved individually or in combination in the etiology of schizophrenia. PMID- 12116183 TI - Genome-wide scan for linkage to schizophrenia in a Spanish-origin cohort from Costa Rica. AB - Genetic isolates have been useful cohorts in which to search for genes underlying disorders of unknown pathology. One such cohort is thought to exist in the Central Valley of Costa Rica surrounding the city of San Jose. Previous investigators identified a rare dominant gene for hereditary deafness in this population, and a suggestive linkage of severe bipolar psychosis has been reported in another study. Ninety-nine families with at least one pair of siblings affected with schizophrenia or a schizophrenia-spectrum diagnosis had clinical evaluations and DNA collected for genotyping. The Marshfield Medical Research Foundation (NHLBI) Mammalian Genotyping Service performed all genotyping using 404 short-tandem repeat polymorphic markers (STRPs) spaced on average 10 cM apart. Data were analyzed using the nonparametric program, GeneHunterPlus. The population structure was investigated using the STRUCT program. No region was found with genome-wide significance for linkage. Using a phenotype of schizophrenia plus schizoaffective disorder, the highest maximum likelihood score (MLS) observed was 1.78 (P < 0.004) at 176.6 cM from pter on chromosome 5q, an area previously implicated by some other groups. In addition, five regions on chromosomes 1p, 2p, 2q, 14p, and 8p had MLSs above 1.0. All other regions produced scores below 1.0. Population genetic analysis reveals no evidence for population substructure, for admixture with other populations, such as Amerindians, or for inbreeding in the parental generation. The latter casts some doubt on this population being an isolate, although there was evidence of inbreeding among the offspring. PMID- 12116184 TI - No evidence for phenotypic variation between probands in case-control versus family-based association studies of schizophrenia. AB - Traditional case-control genetic association studies utilizing unrelated probands are often used interchangeably with family-based designs to detect genes for complex psychiatric disorders. This strategy may be limited, however, if significant phenotypic variation exists between probands enrolled in these two types of studies. The present study compared 37 probands enrolled in a case control study of schizophrenia with 37 age-, sex-, and ethnically matched probands enrolled in a family-based study of schizophrenia. Age of onset of illness was compared as well as performance on a battery of cognitive tests assessing attention, working memory, executive function, and verbal memory. Results revealed no significant differences in age of onset between the two groups or on any measure of cognitive performance. These data do not support reports of significant phenotypic variation between probands in case-control and family-based studies, and suggest that studies utilizing family-based approaches may be used to replicate reports of association made with case-control designs in schizophrenia. PMID- 12116185 TI - Lateral gene transfer of foreign DNA: the missing link between cannabis psychosis and schizophrenia. PMID- 12116186 TI - 1q21-q22 locus is associated with susceptibility to the reality-distortion syndrome of schizophrenia spectrum disorders. PMID- 12116187 TI - Association study between two variants in the DOPA decarboxylase gene in bipolar and unipolar affective disorder. AB - Irregularities of dopaminergic and serotonergic neurotransmission have been implicated in a variety of neuropsychiatric disorders. DOPA decarboxylase (DDC), also known as aromatic L-amino acid decarboxylase, is an enzyme involved directly in the synthesis of dopamine and serotonin and indirectly in the synthesis of noradrenaline. Therefore, the DDC gene can be considered as a candidate gene for affective disorders. Recently, two novel variants were reported in the DDC gene: a 1-bp deletion in the promoter and a 4-bp deletion in the untranslated exon 1. Subsequently, an association case-control study including 112 English patients and 80 Danish patients with bipolar affective disorder (BPAD) revealed a significant association with the 1-bp deletion. This finding prompted us to analyze whether this effect was also present in a larger and ethnically homogeneous sample of 228 unrelated German patients with BPAD (208 patients with BP I disorder, 20 patients with BP II disorder), 183 unrelated patients with unipolar affective disorder (UPAD), and 234 healthy control subjects. For both BPAD and UPAD we could not detect a genetic association with either variant. Thus, our results do not support an involvement of the 1-bp or 4-bp deletion within the DDC gene in the etiology of affective disorders. PMID- 12116188 TI - Association study of the short tandem repeat in the 5' upstream region of the cholecystokinin gene with mood disorders in the Japanese population. AB - We have recently identified a novel polymorphic short tandem repeat (STR) in the 5' upstream region of the cholecystokinin (CCK) gene and reported its association with panic disorder. A linkage study of affective disorder showed a modest linkage signal on the short arm of chromosome 3, the location of the CCK gene. Furthermore, clinical comorbidity of depression and anxiety disorders have been documented. In the present study, we examined a possible association of the CCK STR with mood disorders. We genotyped 165 subjects with mood disorders consisting of unipolar and bipolar disorders and 253 control samples. However, no significant allelic associations were detected between the STR and either the combined mood disorders (P = 0.885), the unipolar group (P = 0.296), or the bipolar group (P = 0.605). These data suggest that the CCK promoter STR is unlikely to have a major genetic effect on the development of mood disorders in the Japanese population. PMID- 12116189 TI - Association of the muscarinic cholinergic 2 receptor (CHRM2) gene with major depression in women. AB - Cholinergic neurons have been implicated in depression and in the disorders of REM sleep in depression. We examined a common A-> T 1890 polymorphism in the 3' UTR of the cholinergic muscarinic receptor 2 (CHRM2) gene. There was a significant increase in the frequency of 11 homozygotes in 126 women with major depression (43.7%) compared to 304 women without major depression (25.7%), P =.001. There was no increase in the frequency of 11 homozygotes in 52 men with depression (26.9%) compared to 278 men without depression (27.7%). Regression analysis, scoring subjects with the 11 genotype as 1, and those with other genotypes as 0, showed that in women r(2) =.030, F = 13.37, P =.0003. By contrast, in men r(2) =.00001, F = 0.002, P =.96. These results are consistent with a gender-specific role of the CHRM2 gene in depression in women. PMID- 12116190 TI - Association analysis of a polymorphism in the G-protein stimulatory alpha subunit in patients with major depression. AB - Growing evidence suggests that G-proteins may be involved in pathogenesis and treatment of affective disorders. Several studies have reported altered levels and/or activities of stimulatory G-proteins in depression. The aim of this study was to investigate whether a polymorphism in the stimulatory alpha subunit of G proteins (T/C point mutation in exon 5; ATT --> ATC at codon 131) is associated with major depression or response to antidepressant treatment. Therefore, we performed a case-control association study with 212 depressive patients and 137 healthy, unrelated controls. There was no evidence for an association between the investigated polymorphism in the G(alpha)(s) gene and major depression, as well as to treatment response. The results of our study are in concordance with recently published findings which do not support the hypothesis that the gene for the stimulatory alpha subunit of G-proteins is a major susceptibility factor in the pathophysiology of major depression. PMID- 12116191 TI - Meta-analysis of the association between tryptophan hydroxylase and suicidal behavior. AB - Tryptophan hydroxylase (TPH) has been the candidate gene of focus in many of the association studies of suicidal behavior in recent years. Initial positive findings with respect to an association between the TPH gene and suicidal behavior have been replicated, but not consistently. Typically, individual studies have investigated small samples, and thus they repeatedly had insufficient statistical power to detect a positive association. Meta-analysis is one approach that can be used to achieve greater statistical power and may be helpful in providing a more conclusive understanding. We used meta-analytic techniques to investigate the association between an intron 7 polymorphism in the TPH gene and suicidal behavior. A total of 39 publications were identified and reviewed, 17 of which were selected for inclusion in this study. We performed two meta-analyses. One compared suicide attempters or completers (N = 1,290) with healthy controls (N = 2,295); the other compared suicide attempters (N = 625) with nonattempters (N = 1,475). None of these studies provided evidence for association (odds ratio (OR) = 1.14, 95% confidence interval (CI) = 0.97-1.34 for the former and OR = 0.96, 95% CI = 0.77-1.20 for the latter). The combined results from comparisons within both groups showed no overall association between suicidal behavior and an intron 7 polymorphism of the TPH gene. PMID- 12116192 TI - Genome-wide linkage analysis of families with obsessive-compulsive disorder ascertained through pediatric probands. AB - The goal of this study was to identify chromosomal regions likely to contain susceptibility alleles for early-onset obsessive-compulsive disorder (OCD). A genome scan was done in 56 individuals from seven families ascertained through pediatric OCD probands; 27 of the 56 subjects had a lifetime diagnosis of definite OCD. Denser mapping of regions on chromosomes 2, 9, and 16 was subsequently done with those subjects and ten additional subjects from the largest family in the study. Direct interviews were completed with 65 of the 66 genotyped individuals. Relatives were interviewed blind to proband status. Of the 65 interviewed individuals, 32 had a lifetime diagnosis of definite OCD. Three of the seven probands had a history of Tourette disorder. Two of the 25 relatives with OCD had a tic history, whereas none of the 33 relatives without OCD had tics. The genome scan consisted of 349 microsatellite markers with an average between-marker distance of 11.3 centiMorgan (cM). Fine mapping was done with 24 additional markers at an average spacing of 1.6 cM. Parametric and nonparametric linkage analyses were conducted using GENEHUNTER(+). The maximum multipoint LOD score with a dominant model was 2.25 on 9p. However, with fine mapping and additional subjects, that LOD score decreased to 1.97. The maximum multipoint nonparametric LOD* score was 1.73 on 19q. The maximum multipoint LOD score with a recessive model was 1.40 on 6p. The results provide suggestive evidence for linkage on 9p and identify regions requiring further study with much larger samples. PMID- 12116193 TI - A dimensional impulsive-aggressive phenotype is associated with the A218C polymorphism of the tryptophan hydroxylase gene: a pilot study in well characterized impulsive inpatients. AB - Tryptophan hydroxylase (TPH) is the rate-limiting enzyme in the biosynthesis of serotonin, and association and linkage studies of its variants in suicidal and impulsive-aggressive behavior have brought conflicting results. This pilot study was designed to investigate whether TPH A218C genotypes could be associated with impulsive behavioral tendencies (IBTs) in consecutively admitted nonpsychotic nonorganic inpatients. Patients (20 females and 34 males; age, 38.8 +/- 11.8) did not differ from healthy nonimpulsive controls (16 females and 11 males; age, 35.2 +/- 10.2) regarding TPH genotypes, but in the patients, the number of IBT was related to the presence of the 218C allele. It was concluded that impulsive aggressive behavior may be associated with the TPH genotype in well-characterized impulsive patients and that the present results stress the importance of considering impulsiveness-aggressiveness in studies investigating the relationship between suicidal behavior and TPH genotypes. PMID- 12116194 TI - A risk-factor model of epistatic interaction, focusing on autism. AB - Research to date on the genetics of autism has not uncovered a major susceptibility locus and indications are that a number of genes, perhaps as many as 15-20, may play detectable but minor roles in the etiology of the condition. To cope with this situation, a risk-factor model based on standard epidemiologic designs is proposed. The model supposes that adding a factor to a fixed set of existing factors always increases the total risk. Thus, according to the model genetic contributions cumulate but are not necessarily additive. A threshold, hence, epistasis is required. The model is applied to several conditions in which the risk of autism is elevated, some genetic (fragile X, tuberous sclerosis) and some exogenous (rubella and thalidomide embryopathies). Male gender is discussed as a risk factor. This approach is contrasted primarily with Gillberg and Coleman's view of autism as "a syndrome or series of syndromes caused by many different separate individual diseases." The principal point of difference is whether the effects of different causes cumulate or do not cumulate. In the present approach they do, in Gillberg and Coleman's they do not. PMID- 12116195 TI - Evaluation of FOXP2 as an autism susceptibility gene. AB - A mutation in the gene FOXP2 was recently identified as being responsible for a complicated speech and language phenotype in a single large extended pedigree. This gene is of interest to autism because it lies in one of the most consistently linked autism chromosomal regions of interest. We therefore tested this gene for its involvement in autism in a large sample of autism families. We completely sequenced the exon containing the mutation, screened the remaining coding sequence using SSCP technology, and identified and genotyped two novel intronic tetranucleotide repeat polymorphisms that were then analyzed for evidence of linkage and linkage disequilibrium (LD). We identified two families in which heterozygous deletions of a small number of glutamines in a long poly glutamine stretch were found in one parent and the autistic probands; no other non-conservative coding sequence changes were identified. Linkage and LD analyses were performed in 75 affected sibling pair families and in two subgroups of this sample defined by the presence/absence of severe language impairment. One allele appeared to have an opposite pattern of transmission in the language based subgroups, but otherwise the linkage and LD analyses were negative. We conclude that FOXP2 is unlikely to contribute significantly to autism susceptibility. PMID- 12116196 TI - Effect of the APOE-491A/T promoter polymorphism on apolipoprotein E levels and risk of Alzheimer disease: The Rotterdam Study. AB - The apolipoprotein E (APOE) gene is involved in lipid transport. A common polymorphism in this gene with the APOE*2, APOE*3, and APOE*4 alleles influences plasma levels of apolipoprotein E and cholesterol. Besides its role in lipid transport, the APOE*4 allele is a genetic risk factor for Alzheimer disease (AD). Recently, a polymorphism in the APOE promoter region was found to be involved in plasma apolipoprotein E levels and was found associated with AD. We studied the effect of this -491A/T promoter polymorphism on plasma apolipoprotein E levels and risk for AD in a population-based case-control study. We found that there was a modest but statistically significant effect of the -491A/T polymorphism on plasma apolipoprotein E levels independent of the APOE genotype. The lowest plasma levels were measured for the AA genotype, highest levels for the TT genotype, and intermediate levels for the heterozygotes. There was a small effect of the -491 AA genotype on AD risk that disappeared after adjusting for APOE genotypes. Our data suggest that the -491A/T polymorphism has an APOE genotype independent effect on plasma apolipoprotein E levels but no APOE-independent effect on AD risk. PMID- 12116197 TI - Association between the TNFalpha-308 A/G polymorphism and the onset-age of Alzheimer disease. AB - Local inflammatory processes associated with amyloid plaques would contribute to the progression of late-onset Alzheimer disease (LOAD). Tumor necrosis factors alpha (TNF(alpha)) and beta (LT(alpha)) are inflammatory cytokines involved in the local immune response occurring in the central nervous system of LOAD patients. Genetic variation at these genes could contribute to the risk of developing AD or influence the age at the onset of the disease. We genotyped 315 LOAD patients and 400 healthy controls for DNA-polymorphisms in the genes encoding TNF(alpha) (-308 G/A, -238G/A) and LT(alpha) (Asn26Thr). Carriers of 308A showed a mean age at onset 3 years younger than noncarriers of this allele (P = 0.019). Our data suggest an effect of the TNF(alpha)-308 polymorphism on the age at onset of late AD. This represents additional evidence of the importance of genetic variation at the proinflammatory components in the origin and progression of this common neurodegenerative disease. PMID- 12116198 TI - Difference in disease-free survival curve and regional distribution according to subtype of spinocerebellar ataxia: a study of 1,286 Japanese patients. AB - Expansions of trinucleotide repeats have been discovered in spinocerebellar ataxia (SCA) types 1, 2, 6, 7, 12, and 17, Machado-Joseph disease (MJD/SCA3), and dentatorubropallidoluysian atrophy (DRPLA). However, the frequency of familial SCA in Japan remains unclear. The number of trinucleotide repeats was determined for 1,286 patients. Three hundred and thirty families (523 cases) were autosomal dominant group (A), and 165 families were positive for family history but not autosomal dominant group (B), while the remaining 598 cases were the sporadic group (C). The frequency of SCA subtypes in autosomal dominant group was: 1) 5.5% for SCA1; 2) 2.4% for SCA2; 3) 27.6% for MJD/SCA3; 4) 25.5% for SCA6; 5) 0.3% for SCA17; and 6) 7.3% for DRPLA. Abnormal expansion of SCA12 was not detected. Another 31.5% of the patients in the autosomal dominant group had unknown genetic abnormalities. Within group B, SCA6 was the most prominent and within the sporadic group MJD/SCA3 and SCA6 were the most common subtypes observed. The disease-free survival curve of SCA6 was different from that of other SCAs and the mean age at onset for SCA6 was found to be later than that of the other types. Regional differences were observed in the relative rate of SCA subtypes. MJD/SCA3 appears more common in the Kanto and Kyushu districts of Japan, whereas SCA6 is most common in the Chugoku district. In order to establish an effective social welfare system for SCA patients, clinical course and regional differences in the prevalence of SCA subtypes must be taken into consideration. PMID- 12116199 TI - Complex relationship between Parkin mutations and Parkinson disease. AB - Mutations in the Parkin gene cause juvenile and early onset Parkinsonism. While Parkin-related disease is presumed to be an autosomal-recessive disorder, cases have been reported where only a single Parkin allele is mutated and raise the possibility of a dominant effect. In this report, we re-evaluate twenty heterozygous cases and extend the mutation screening to include the promoter and intron/exon boundaries. Novel deletion, point and intronic splice site mutations are described, along with promoter variation. These data, coupled with a complete review of published Parkin mutations, confirms that not only is recessive loss of Parkin a risk factor for juvenile and early onset Parkinsonism but that Parkin haplo-insufficiency may be sufficient for disease in some cases. PMID- 12116201 TI - Genetic risk assessment in carrier testing for spinal muscular atrophy. AB - As evidenced by the complete absence of a functionally critical sequence in exon 7, approximately 94% of individuals with clinically typical spinal muscular atrophy (SMA) lack both copies of the SMN1 gene at 5q13. Hence most carriers have only one copy of SMN1. Combining linkage and dosage analyses for SMN1, we observed unaffected individuals who have two copies of SMN1 on one chromosome 5 and zero copies of SMN1 on the other chromosome 5. By dosage analysis alone, such individuals, as well as carriers of non-deletion disease alleles, are indistinguishable from non-carrier individuals. We report that approximately 7% of unaffected individuals without a family history of SMA have three or four copies of SMN1, implying a higher frequency of chromosomes with two copies of SMN1 than previously reported. We present updated calculations for disease and non-disease allele frequencies and we describe how these frequencies can be used for genetic risk assessment in carrier testing for SMA. PMID- 12116202 TI - Genetic heterogeneity of syndromic X-linked recessive microphthalmia anophthalmia: is Lenz microphthalmia a single disorder? AB - Nonsyndromic congenital microphthalmia or anophthalmia is a heterogeneous malformation with autosomal dominant, autosomal recessive, and X-linked modes of inheritance. Lenz microphthalmia syndrome comprises microphthalmia with mental retardation, malformed ears, skeletal anomalies, and is inherited in an X-linked recessive pattern. Prior studies have shown linkage of both isolated (or nonsyndromic) anophthalmos (ANOP1, [MIM 301590]) and Lenz syndrome [MIM 309800] to Xq27-q28. Nonsyndromic colobomatous microphthalmia [MIM 300345] has been linked to Xp11.4-Xq11.1. We describe a five-generation African-American family with microphthalmia or anophthalmia, mental retardation, and urogenital anomalies, in an X-linked recessive inheritance pattern, consistent with Lenz syndrome. Initial linkage analysis with microsatellite markers excluded the region in Xq27-q28 previously reported as a candidate region for ANOP1 [MIM 301590]. An X-chromosome scan revealed linkage to a 10-cM region between markers DXS228 and DXS992 in Xp11.4-p21.2. Multipoint analysis gave a maximum LOD score of 2.46 at marker DXS993. These data show that X-linked recessive syndromic microphthalmia exhibits genetic heterogeneity. In addition, it suggests that Lenz microphthalmia syndrome, previously thought to be a single disorder, may represent an amalgam of two distinct disorders. PMID- 12116203 TI - Investigations of micro-organic brain damage (MOBD) in heterozygotes of metachromatic leukodystrophy. AB - Potential damage of central and peripheral nervous system expressed as micro organic brain damage (MOBD) was investigated in 27 unrelated heterozygotes with metachromatic leukodystrophy (MLD). Arylsulfatase A (ARSA) was determined in peripheral blood leukocytes and sulfatide excretion was estimated in 24-hour urine collections. Genomic DNA was analyzed for the ARSA pseudodeficiency (PD) allele by a PCR method. Clinical investigations included examination of hyper reflexia, Babinski reflex, Wechsler Adult Intelligence Scale, Benton test, evoked potentials, and nerve conduction velocity (NCV). In our study, a higher incidence of evident or possible micro-organic brain damage was observed in true MLD/PD and MLD heterozygotes (NO/MLD, where NO means the wild allele) than in controls. On the basis of the Benton test, MOBD was suggested or indicated in 67% of MLD heterozygotes, 50% of MLD/PD heterozygotes, and 26% of controls. In our small group of carriers with MLD and PD mutations, persons NO/MLD(PD) with one wild type allele did not show MOBD and displayed higher ARSA/beta-galactosidase ratios, unlike true MLD/PD compound heterozygotes who carry MLD-causing mutation in one allele and the ARSA-PD polymorphism in the second. Theoretically, this is a shift from autosomal recessive to autosomal dominant-like inheritance, especially when one cannot exclude the influence of polymorphisms (like ARSA-PD) in the wild allele. Since all psychological tests were age-matched, it can be assumed that the MOBD observed in MLD carriers does not have a progressive character unlike in MLD patients. However, it should be mentioned that MOBD appears to have no overt clinical consequences. PMID- 12116204 TI - Developmental field defects: coming together of associations and sequences during blastogenesis. AB - We report on two patients with an unusual combination of multiple congenital anomalies including holoprosencephaly, encephalocele, and additional defects commonly observed in the VACTERL and schisis "associations." One of the infants had a chromosome abnormality characterized by partial duplication and deletion of chromosome 18. VACTERL association was characterized recently as a primary developmental field defect (DFD) [Martinez-Frias et al., 1998: Am J Med Genet 76:291-296]. In some cases, sequences may also represent uncomplicated DFDs. We suggest that findings in both of these cases represent abnormalities of blastogenesis involving the primary field resulting in holoprosencephaly and VACTERL and schisis anomalies, and show that similar primary DFDs are causally heterogeneous. PMID- 12116205 TI - Personal experiences of cystic fibrosis (CF) carrier couples prospectively identified in CF families. AB - This qualitative study explores the experiences of cystic fibrosis (CF) carrier couples, prospectively identified in CF families, and the impact of the resulting genetic risk on reproductive behavior. Of the 12 couples identified until 1997, seven couples participated in semistructured interviews and two couples filled in a questionnaire, two to eight years after receipt of the test-results. After receiving the results, most couples reported that they were shocked, because they did not expect to both be carriers. More anxiety was expressed by those who were pregnant (n = 4) at the time of testing. There were reported difficulties in disclosing the results to family members, and the reactions of family members were not always supportive. After testing, some couples had problems with reproductive decision-making. All viable pregnancies (17 in 8 couples) were monitored by prenatal diagnosis; all affected pregnancies were terminated (6 in 4 couples). Couples who have live-born children after testing may subsequently have concerns during infancy about the correctness of the results of prenatal diagnosis and how to inform their children. Most couples did not regret the testing and, in general, the counseling was experienced positively, although some dissatisfaction was reported with regard to the psychological support received during pregnancy. Couples supported the idea of carrier screening in the general population, although various concerns were expressed. The results indicate a preference for testing before pregnancy. These findings may be useful in investigating possible dilemmas caused by the introduction of population carrier screening. Observations reported here might also apply to other recessively inherited disorders. PMID- 12116206 TI - Ring chromosome 21 in a boy and a derivative chromosome 21 in the mother: implication for ring chromosome formation. AB - We report on r(21) chromosome in a boy and a der(21) chromosome in his mother. Cytogenetic studies revealed a mosaic 45,XY[4]/46,XY,r(21)[96] karyotype in the boy and a 46,XX,der(21)[100] karyotype in the mother. Fluorescence in situ hybridization analysis for D21Z1 at the centromere, AML1 at 21q22.1, LSI21 at 21q22.2, and 21qtel at the telomere region showed that the r(21) chromosome retained single copies of D21Z1, AML1, and LSI21 and lacked the 21qtel, whereas the der(21) chromosome had two copies of the 21qtel on both of its ends and single copies of D21Z1, AML1, and LSI21, with a paracentric inversion of AML1 and LSI21 (21qtel-D21Z1-LSI21-AML1-21qtel). Microsatellite analysis for nine loci on 21q22.3 indicated that the r(21) chromosome was missing a distal 21q22.3 region involving D21S1890, D21S1411, and D21S1903 with no maternally derived alleles, and that the der(21) chromosome was associated with duplication of a distal 21q22.3 region encompassing D21S1890 and D21S1446. The results suggest that a U type exchange occurred between the homologous distal 21q regions duplicated on the der(21) chromosome, leading to the r(21) formation. This is a novel mechanism put forward for the formation of a monocentric ring chromosome. PMID- 12116207 TI - Childhood-onset ataxia: testing for large CAG-repeats in SCA2 and SCA7. AB - Infantile- and juvenile-onset spinal cerebellar ataxia (SCA) is associated with expansion of 130 to more than 200 CAG-repeats in the SCA2 and SCA7 genes. Routine clinical assays for SCA2 and SCA7, which use polymerase chain reaction (PCR) and denaturing PAGE (polyacrylamide gel electrophoresis), will not reliably detect such large expansions. An assay based on separation of PCR products on an agarose gel, blotting, and hybridization with a (CAG)6 oligonucleotide probe was used to test DNA from individuals more than 10 years of age who had a possible diagnosis of SCA. Among 25 cases, the PCR-blot assay confirmed the presence of SCA2 expansions between 230 and 500 repeats in four unrelated individuals, but did not detect any cases of extreme expansion in the SCA7 gene. The PCR-blot assay provides reliable detection of extreme expansion mutations. Routine incorporation of this assay in clinical laboratories may reveal that infantile-juvenile forms of SCA2 and SCA7 are more prevalent than previously recognized. PMID- 12116208 TI - Refinement of the autosomal recessive polycystic kidney disease (PKHD1) interval and exclusion of an EF hand-containing gene as a PKHD1 candidate gene. AB - Autosomal recessive polycystic kidney disease (ARPKD) is an often devastating form of polycystic kidney disease that presents primarily in infancy. The locus, PKHD1 (polycystic kidney and hepatic disease 1), on chromosome 6p21.1-p12, has been linked to all classical forms of this disorder. In previous studies, we cloned the PKHD1 interval in a set of overlapping YACs, converted this YAC-based framework into a BAC/PAC contig, and delimited the critical interval to a region flanked by the markers D6S1714 and D6S1024. We now have refined the genetic interval using new polymorphic markers developed from our BAC/PAC resources. In addition, we have evaluated a recently identified, EF hand-containing gene that maps to the interval of interest, established its transcript sequence, defined its genomic organization, and excluded this new gene as a PKHD1 candidate. Therefore, this study has narrowed the PKHD1 interval and excluded a potentially relevant gene as a PKHD1 candidate gene. This further refinement of the PKHD1 interval will facilitate efforts to identify the PKHD1 gene by positional cloning. These data also provide additional, highly polymorphic markers for haplotype-based diagnostic testing for ARPKD. PMID- 12116209 TI - Unbalanced translocation (3;5)(q26.1;p14): a clinical report. AB - A patient with a multiple congenital anomalies/mental retardation (MCA/MR) syndrome had an unbalanced translocation (3;5)(q26.1;p14), causing partial 5p monosomy and partial 3q trisomy. The phenotype observed in this patient results from the combination of those described in the isolated dup(3q) and del(5p) syndromes. Some clinical features of this patient are shared by the Smith-Lemli Opitz syndrome (SLOS), a well-known MCA/MR syndrome due to the deficiency of 7 dehydrocholesterol reductase (DHCR7). We review the previously reported cases of chromosomal anomalies with clinical features suggesting SLOS. PMID- 12116210 TI - Auditory canal atresia, humeroscapular synostosis, and other skeletal abnormalities: confirmation of the autosomal recessive "SAMS" syndrome. AB - A second girl with the unique combination of auditory canal atresia and scapulohumeral synostosis is reported. This patient also had bilateral clubfeet and genital abnormalities. The other patient reported with this syndrome and the presently reported child both had consanguineous parents. Mental development was normal in both children. The acronym SAMS (Short stature, Auditory canal atresia, Mandibular hypoplasia, and Skeletal abnormalities) was suggested to describe the main manifestations in this syndrome. PMID- 12116211 TI - Novel mutation of TBX3 in a Japanese family with ulnar-mammary syndrome: implication for impaired sex development. AB - We report on a Japanese family (two brothers and their mother) with ulnar-mammary syndrome (UMS). Clinical features included hypoplasia or aplasia of upper limbs on the ulnar side in the three affected individuals, micropenis with or without cryptorchidism, and hypoplastic nipples in the brothers; and hypoplastic mammary glands and nipples, poor perspiration, and bicornuate uterus in the mother. Endocrine studies performed for the underdeveloped external genitalia when the brothers were 11 6/12 and 7 2/12 years old, respectively, indicated low to low normal responses of luteinizing hormone (LH) and follicle stimulating hormone (FSH) to gonadotropin releasing hormone stimulation tests (elder brother: LH = < 0.2 --> 2.2 IU/L, FSH = 0.6 --> 2.2 IU/L; younger brother: LH = < 0.2 --> 3.3 IU/L, FSH = 0.7 --> 4.4 IU/L) and normal testosterone responses to human gonadotropin stimulation tests (elder brother: < 0.5 --> 8.8 nmol/L; younger brother: < 0.5 --> 6.3 nmol/L). Testosterone enanthate therapy (25 mg/dose IM twice) was effective in the brothers, with penile length increase being similar between the brothers (approximately 5 mm/dose) and 23 age-matched boys with idiopathic micropenis (mean 4.4 mm/dose, range 2.5-7.5 mm/dose). Sequence analysis of the TBX3 gene showed a novel heterozygous nonsense mutation (A817T, K273X) in exon 4 of the three patients. The results are consistent with the previous finding that UMS is caused by haploinsufficiency of TBX3, and imply that mild gonadotropin deficiency may be the primary cause for underdeveloped external genitalia in males with UMS. PMID- 12116212 TI - Probable identity of Goltz syndrome and Van Allen-Myhre syndrome: evidence from phenotypic evolution. AB - We describe a girl who was diagnosed with split foot-split hand anomaly prenatally, in whom at birth the diagnosis of Van Allen-Myhre syndrome was made, and who at 8 months of age was recognized to have Goltz syndrome. Based on the evolution of clinical features in this infant, we suggest that our case, as well as that reported by Van Allen and Myhre, is an example of unusually severe Goltz syndrome. PMID- 12116213 TI - Possible third case of Lin-Gettig syndrome. AB - We report a patient with craniosynostosis, severe mental retardation, absence of the corpus callosum, camptodactyly, hypogonadism, and ventricular septal defect. We propose that he has Lin-Gettig syndrome and that he is the third reported patient with this entity. Our patient also had additional phenotypic features, including palatal cleft and absent rapid eye movement (REM) sleep that were not present in the two previously described patients with this syndrome. High resolution karyotype and subtelomeric fluorescence in situ hybridization (FISH) for cryptic telomeric rearrangement were normal. The existence of an unrelated patient with Lin-Gettig syndrome supports that this is a separate and distinct clinical entity. PMID- 12116215 TI - Craniofrontonasal syndrome and diaphragmatic hernia. PMID- 12116214 TI - Kabuki syndrome: report of six Thai children and further phenotypic and genetic delineation. AB - We describe six Thai children with the Kabuki syndrome. Monozygotic twin boys discordant for the syndrome were encountered in a family. The affected twin had all five cardinal features of the syndrome, whereas the unaffected twin had none of them. The presence of monozygotic twins discordant for the syndrome argues against a single gene origin of the disorder, but by no means excludes it. In another family, a mother had a facial appearance similar to her affected son. Lower lip pits with or without symmetrical lower lip nodules were present in three of the six children, and pilonidal sinus was seen in five children. These clinical manifestations were much more common than previously described. Other inconsistent findings included early eruption of the lower central incisors, a skin defect of the head, and transient hyperthyrotropinemia in infancy. PMID- 12116217 TI - Structural and mutational analysis of antiquitin as a candidate gene for Meniere disease. PMID- 12116216 TI - Multicolor karyotypic interpretation of a heterochromatin-associated marker chromosome in a dysmorphic girl with developmental delay. PMID- 12116218 TI - Bilateral hyperplasia of the mandibular coronoid processes in patients with nevoid basal cell carcinoma syndrome: an undescribed sign. PMID- 12116219 TI - MTHFR is not a risk factor in the development of isolated nonsyndromic cleft lip and palate. PMID- 12116220 TI - Evidence for a cleft palate only locus on chromosome 4 near MSX1. PMID- 12116221 TI - Resolution of non-immune hydrops in Noonan syndrome with favorable outcome. PMID- 12116222 TI - Localization of a gene for nonspecific X-linked mental retardation (MRX 76) to Xp22.3-Xp21.3. PMID- 12116223 TI - Mosaic trisomy of a small r(1) with an abnormal phenotype. PMID- 12116225 TI - Testing for genetic associations with the PAX gene family in a spina bifida population. AB - Neural tube defects (NTDs) are among the most common severely disabling birth defects in the United States, affecting approximately 1-2 of every 1,000 live births. The etiology of NTDs is multifactorial, involving the combined action of both genetic and environmental factors. A nonparametric linkage method, the transmission disequilibrium test (TDT), was utilized to determine if the genes in the PAX family play a role in the formation of NTDs. DNA from 459 spina bifida (SB) patients and their parents (430 mothers and 239 fathers, for a total population of 1,128 subjects) was tested for linkage and association utilizing polymorphic markers from within or very close to the PAX genes of interest. Significant findings were obtained for the following markers: marker locus D20S101 flanking the PAX1 gene (P = 0.019), marker locus D1S228 within the PAX7 gene (P = 0.011), and marker locus D2S110 within the PAX8 gene (P = 0.013). Even though our findings are only mildly significant, given the known expression patterns of the PAX genes in development and the availability of their sequences, we elected to follow up these results by testing these genes directly for mutations utilizing single-strand conformational analysis (SSCA) and direct sequencing. Multiple variations were detected in each of the PAX genes with significant TDT results; however, these variations were not passed from parent to child in phase with the positively transmitted allele. Therefore, it is unlikely that these variations contribute to susceptibility for SB, but rather are previously unreported polymorphisms. PMID- 12116226 TI - Testing for genetic associations in a spina bifida population: analysis of the HOX gene family and human candidate gene regions implicated by mouse models of neural tube defects. AB - Neural tube defects (NTDs) are among the most common severely disabling birth defects in the United States, affecting approximately 1-2 of every 1,000 live births. The etiology of NTDs is multifactorial, involving the combined action of both genetic and environmental factors. HOX genes play a central role in establishing the initial body plan by providing positional information along the anterior-posterior body and limb axis and have been implicated in neural tube closure. There are many mouse models that exhibit both naturally occurring NTDs in various mouse strains as well as NTDs that have been created by "knocking out" various genes. A nonparametric linkage method, the transmission disequilibrium test (TDT), was utilized to test the HOX gene family and human equivalents of genes (when known) or the syntenic region in humans to those in mouse models which could play a role in the formation of NTDs. DNA from 459 spina bifida (SB) affected individuals and their parents was tested for linkage and association utilizing polymorphic markers from within or very close to the HOXA, HOXB, HOXC, and HOXD genes as well as from within the genes/gene regions of eight mouse models that exhibit NTDs. No significant findings were obtained for the tested markers. PMID- 12116227 TI - SNPs in the CpG island of NAP1L2: a possible link between DNA methylation and neural tube defects? AB - Deletion of the murine X-linked Nap1l2 gene causes lethality from midgestation onwards. The affected embryos exhibit neural tube defects (NTDs) closely resembling spina bifida and anencephaly in humans. X-linked familial and spontaneous cases of NTD were analyzed for sequence alterations in the human NAP1L2. No differences were found in the familial cases. However, a number of single nucleotide polymorphisms (SNPs) within the 5' region of NAP1L2 were identified both in cases of spontaneous NTD and in normal controls. Most of these SNPs lead to the replacement of guanidines or cytosines within a CpG island that is conserved between the human and the mouse promoter regions. Demethylation in vitro activates Nap1l2 transcriptional activity, suggesting the importance of the CpG island in regulating the activity of the Nap1l2/NAP1L2 genes, and the potential importance of the polymorphisms in modifying their transcriptional activity. NAP1L2/Nap1l2 expression may therefore depend on the genetic environmental factors that are frequently associated with NTDs. PMID- 12116228 TI - T locus shows no evidence for linkage disequilibrium or mutation in American Caucasian neural tube defect families. AB - We investigated the T locus as a candidate gene in a series of patients and families with lumbosacral myelomeningocele. Single-strand conformation polymorphism (SSCP) analysis was used to identify sequence variation in all 8 exons and in intron 7 of this locus. We found evidence of substantial polymorphism within this locus, as previously reported [Papapetrou et al., 1999, J Med Genet 36:208-213], and moderately significant evidence of linkage disequilibrium with the CacI polymorphism of exon 8. However, when the locus was considered as a whole, with all single nucleotide polymorphisms (SNPs) integrated into a haplotype, there was no evidence for linkage disequilibrium. In addition, we did not identify any new sequence variants. Thus, we conclude that the T locus is not a major locus for human NTDs in this sample. PMID- 12116229 TI - Heterogeneity for congenital generalized lipodystrophy in seventeen patients from Oman. AB - Seventeen children with congenital generalized lipodystrophy or Berardinelli-Seip Congenital Lipodystrophy (BSCL) from 12 consanguineous sibships were observed in Oman. All children had widespread absence of adipose tissue from infancy together with apparent muscle hypertrophy and hepatomegaly. They did not appear to represent a single homogenous entity, and it was possible to subclassify the cases into two distinct groups. In the first group of seven cases, the features were similar to other published cases with acanthosis nigricans, raised insulin levels, and insulin resistance. In this group, there was an association between the degree of acanthosis nigricans and the severity of the disorder. Molecular analysis of these cases showed homozygosity at the BSCL2 locus on chromosome 11q13 in four of the seven cases. In the second group of ten cases, there were striking abnormalities in both skeletal and nonskeletal muscle. Reduced exercise tolerance and percussion myoxedema were observed in skeletal muscle, while infantile hypertrophic pyloric stenosis, prominent veins (phlebomegaly), disturbance of cardiac rhythm, and cardiomyopathy were observed in nonskeletal muscle. There was evidence against homozygosity in some cases for the known loci for BSCL, and this group may represent a new clinical syndrome with lipodystrophy at a different genetic location. PMID- 12116230 TI - Prevalence of the fragile X syndrome in African-Americans. AB - Since the development of a molecular diagnosis for the fragile X syndrome in the early 1990s, several population-based studies in Caucasians of mostly northern European descent have established that the prevalence is probably between one in 6,000 to one in 4,000 males in the general population. Reports of increased or decreased prevalence of the fragile X syndrome exist for a few other world populations; however, many of these are small and not population-based. We present here the final results of a 4-year study in the metropolitan area of Atlanta, Georgia, establishing the prevalence of the fragile X syndrome and the frequency of CGG repeat variants in a large Caucasian and African-American population. Results demonstrate that one-quarter to one-third of the children identified with the fragile X syndrome attending Atlanta public schools are not diagnosed before the age of 10 years. Also, a revised prevalence for the syndrome revealed a higher point estimate for African-American males (1/2,545; 95% CI: 1/5,208-1/1,289) than reported previously, although confidence intervals include the prevalence estimated for Caucasians from this (1/3,717; 95% CI: 1/7,692 1/1,869) and other studies. Further population-based studies in diverse populations are necessary to explore the possibility that the prevalence of the fragile X syndrome differs among world populations. PMID- 12116231 TI - Candidate genes involved in cardiovascular risk factors by a family-based association study on the island of Kosrae, Federated States of Micronesia. AB - Altered plasma levels of lipids and lipoproteins, obesity, hypertension, and diabetes are major risk factors for atherosclerotic cardiovascular disease. To identify genes that affect these traits and disorders, we looked for association between markers in candidate genes (apolipoprotein AII (apo AII), apolipoprotein AI-CIII-AIV gene cluster (apo AI-CIII-AIV), apolipoprotein E (apo E), cholesteryl ester transfer protein (CETP), cholesterol 7alpha-hydroxylase (CYP7a), hepatic lipase (HL), and microsomal triglyceride transfer protein (MTP)) and known risk factors (triglycerides (Tg), total cholesterol (TC), apolipoprotein AI (apo AI), apolipoprotein AII (apo AII), apolipoprotein B (apo B), body mass index (BMI), blood pressure (BP), leptin, and fasting blood sugar (FBS) levels.) A total of 1,102 individuals from the Pacific island of Kosrae were genotyped for the following markers: Apo AII/MspI, Apo CIII/SstI, Apo AI/XmnI, Apo E/HhaI, CETP/TaqIB, CYP7a/BsaI, HL/DraI, and MTP/HhpI. After testing for population stratification, family-based association analysis was carried out. Novel associations found were: 1) the apo AII/MspI with apo AI and BP levels, 2) the CYP7a/BsaI with apo AI and BMI levels. We also confirmed the following associations: 1) the apo AII/MspI with Tg level; 2) the apo CIII/SstI with Tg, TC, and apo B levels; 3) the Apo E/HhaI E2, E3, and E4 alleles with TC, apo AI, and apo B levels; and 4) the CETP/TaqIB with apo AI level. We further confirmed the connection between the apo AII gene and Tg level by a nonparametric linkage analysis. We therefore conclude that many of these candidate genes may play a significant role in susceptibility to heart disease. PMID- 12116232 TI - Expanding phenotype of XNP mutations: mild to moderate mental retardation. AB - Mutations in the XNP gene have been reported in alpha thalassemia/mental retardation (MR) syndrome (ATR-X) and other severe X-linked MR conditions with facial dysmorphisms. In this report, we describe a missense mutation in exon 18 in a family with borderline to moderate MR. Like other disorders associated with an XNP mutation, skewed X-inactivation was found in all carrier females in this family. Only retrospective examination revealed childhood facial hypotonia and HbH inclusions in some of the affected males. These results expand the spectrum of clinical phenotypes known to be due to mutations in the XNP gene, and indicate that XNP mutation analysis should not be restricted to patients with severe MR and characteristic facial features. PMID- 12116233 TI - Mutations in the XPD gene in xeroderma pigmentosum group D cell strains: confirmation of genotype-phenotype correlation. AB - Xeroderma pigmentosum (XP) is a sun-sensitive and cancer-prone genetic disorder consisting of seven genetically distinct complementation groups (groups A-G). XP group D (XP-D) is a heterogeneous group. Mutations in the XPD gene (XPD) can exhibit three distinct clinical phenotypes: XP, trichothiodystrophy (TTD), or XP combined with Cockayne syndrome. XPD protein is required for both nucleotide excision repair (NER) and basal transcription. Therefore, different mutations in XPD may affect NER and transcription activities to various degrees and result in such diverse phenotypes. In this study, we identified six causative mutations, two of which have not been described, in five XP-D cell strains tested. The cell strains were all compound heterozygotes with different mutations. In all cell strains, one allele was thought to be functionally null and the other was a less severe allele with R683W, R683Q, and R666W substitutions. The second allele in each strain was specific to the XP phenotype. The findings are consistent with the hypothesis that the site of mutation of the XPD gene determines the clinical phenotype, XP or TTD. PMID- 12116234 TI - Familial dysautonomia: detection of the IKBKAP IVS20(+6T --> C) and R696P mutations and frequencies among Ashkenazi Jews. AB - Familial dysautonomia (FD) is an autosomal recessive congenital neuropathy that occurs almost exclusively in the Ashkenazi Jewish (AJ) population. Mutations in the IkappaB kinase complex-associated protein (IKBKAP) gene cause FD. Two IKBKAP mutations, IVS20(+6T --> C) and R696P, have been identified in FD patients of AJ descent. The splice site mutation IVS20(+6T --> C) is responsible for > 99.5% of known AJ patients with FD, and haplotype analyses were consistent with a common founder. In contrast, the R696P mutation has been identified in only a few AJ patients. To facilitate carrier detection, a single PCR and allele-specific oligonucleotide (ASO) hybridization assay was developed to facilitate the detection of both the IVS20(+6T --> C) and R696P mutations. Screening of 2,518 anonymous AJ individuals from the New York metropolitan area revealed a carrier frequency for IVS20(+6T --> C) of 1 in 32 (3.2%; 95% CI, 2.5-3.9%), similar to the previously estimated carrier frequency (3.3%) based on disease incidence. No carrier was identified for the R696P lesion, indicating that the mutation was rare in this population (< 1 in 2,500). This sensitive and specific assay should facilitate carrier screening for FD mutations in the AJ community, as well as postnatal diagnostic testing. PMID- 12116235 TI - Chromosome 13q neocentromeres: molecular cytogenetic characterization of three additional cases and clinical spectrum. AB - We report three new cases of chromosome 13 derived marker chromosomes, found in unrelated patients with dysmorphisms and/or developmental delay. Molecular cytogenetic analysis was performed using fluorescence in situ hybridization (FISH) with chromosome-specific painting probes, alpha satellite probes, and physically mapped probes from chromosome 13q, as well as comparative genomic hybridization (CGH). This analysis demonstrated that these markers consisted of inversion duplications of distal portions of chromosome 13q that have separated from the endogenous chromosome 13 centromere and contain no detectable alpha satellite DNA. The presence of a functional neocentromere on these marker chromosomes was confirmed by immunofluorescence with antibodies to centromere protein-C (CENP-C). The cytogenetic location of a neocentromere in band 13q32 was confirmed by simultaneous FISH with physically mapped YACs from 13q32 and immunofluorescence with anti-CENP-C. The addition of these three new cases brings the total number of described inv dup 13q neocentic chromosomes to 11, representing 21% (11/52) of the current overall total of 52 described cases of human neocentric chromosomes. This higher than expected frequency suggests that chromosome 13q may have an increased propensity for neocentromere formation. The clinical spectrum of all 11 cases is presented, representing a unique collection of polysomy for different portions of chromosome 13q without aneuploidies for additional chromosomal regions. The complexity and variability of the phenotypes seen in these patients does not support a simple reductionist view of phenotype/genotype correlation with polysomy for certain chromosomal regions. PMID- 12116236 TI - Paternal UPD14 is responsible for a distinctive malformation complex. AB - We present a boy and two girls with paternal uniparental disomy of chromosome 14q (patUPD14). One girl had a Robertsonian translocation, whereas two a normal karyotype. Based on the manifestations of these patients and four previously reported patients who all had translocated chromosome 14, The patUPD14 was thought to constitute a distinctive syndrome. The hallmarks included abdominal muscular defects, skeletal anomalies, and characteristic facies. The phenotype of patUPD14 was consistent with that of a previously reported mouse model, i.e., mouse embryos with paternal uniparental disomy of chromosome 12 that has a region orthologous to that of human chromosome 14. Dose effects of newly recognized imprinted genes on human chromosome 14q32, DLK1 and GTL2, could play an important role in the pathogenic mechanism of the distinctive malformation complex. PMID- 12116237 TI - Mosaic variegated aneuploidy with growth hormone deficiency and congenital heart defects. AB - We describe a 12-year-old boy with mosaic variegated aneuploidy (MVA), subnormal response to growth hormone (GH) stimulation testing, and short stature. In addition to features more commonly described in MVA such as microcephaly, cognitive deficits, and certain facial features, he also has features not commonly reported in MVA, including short limb segments, epidermoid cysts, ventricular septal defect, and subaortic stenosis. Chromosomal analysis revealed hyperdiploid chromosome numbers ranging from 47 to 70; modal number 50, in 24% of the metaphases. This case demonstrates that although the phenotype of MVA almost always includes growth failure, microcephaly, and mental retardation, additional features may vary greatly across individuals. His clinical features and course suggest that in addition to GH deficiency, he may have an intrinsic inability of the growth plate to respond to growth hormone. PMID- 12116238 TI - Clinical, cytogenetic, and molecular findings in 45,X/47,XX,+18 mosaicism: clinical report and review of the literature. AB - We report cytogenetic and molecular findings performed in a patient with double mosaic aneuploidy. Chromosome analysis of amniotic fluid cells from a 17-week-old fetus was performed because of advanced maternal age. Two karyotypes were detected: 45,X and 47,XX,+18 (50:50%). The same cell lines were determined in uncultured and cultured amniocytes of a second amniotic fluid sample, in fetal lymphocytes, and in uncultured and cultured cells of achilles tendon by conventional cytogenetics and fluorescence in situ hybridization (FISH). In the different investigated tissues, the percentage of cells with 45,X karyotype ranged from 20-99% and the percentage of cells with 47,XX,+18 ranged from 1-80%. The pregnancy was terminated at 22 + 0 weeks because of a severe cardiac malformation. Pathologic examination showed a fetus with aspects typical for manifestation of trisomy 18 and monosomy X, especially in the internal organs. The parent and cell stage of origin was determined by short tandem repeat typing and revealed a maternal meiotic division error that led to trisomy 18, as well as a somatic loss of a paternal sex chromosome. Only two other patients with the same mosaicism have been reported so far. Genetic counseling and prognosis remains challenging. PMID- 12116239 TI - Report of a child with aortic aneurysm, orofacial clefting, hemangioma, upper sternal defect, and marfanoid features: possible PHACE syndrome. AB - We report a female patient who had a scalp hemangioma, a cleft uvula, an upper sternal defect, pectus excavatum, arachnodactyly, pes planus, and joint hypermobility. She had rupture of an aortic aneurysm after minor trauma at 11 years of age. At 17 years of age, elective repair of a dilated, ectatic aorta was complicated by cerebral ischemia. Other vascular abnormalities in the proband included an aneurysm of the left subclavian artery, atresia of the right carotid artery, and calcified cerebral aneurysms. We believe that the proband's physical anomalies are best described by the PHACE (posterior fossa brain malformations, hemangiomas, arterial anomalies, coarctation of the aorta and cardiac defects, and eye abnormalities) phenotypic spectrum. This spectrum of physical anomalies also includes sternal clefting and hemagiomas as part of the sternal malformation/vascular dysplasia (SM/VD) association, as found in our patient, and the acronym PHACES has also been used. We consider that the PHACE phenotypic spectrum is likely to be broader than previously recognized and includes orofacial clefting and aortic dilatation and rupture. Our patient also had skeletal anomalies that lead to consideration of Marfan syndrome as a diagnosis. It should be recognized that there is clinical overlap between PHACE syndrome and Marfan syndrome when aortic dilatation is present. We would also like to emphasize the minor nature of the cutaneous findings in our patient despite her severe vascular complications. This is in contrast to previous reports of large or multiple hemangiomas in PHACE syndrome. PMID- 12116240 TI - Novel de novo mutation of MADH4/SMAD4 in a patient with juvenile polyposis. PMID- 12116241 TI - Transposition of the great arteries in asplenia and polysplenia phenotypes. PMID- 12116242 TI - No threat to board certified geneticists and genetic counselors. PMID- 12116244 TI - Higher tPA levels are associated with Apo E. PMID- 12116245 TI - Abnormal sterol metabolism in a patient with Antley-Bixler syndrome and ambiguous genitalia. AB - Antley-Bixler syndrome (ABS) is a rare multiple anomaly syndrome comprising radiohumeral synostosis, bowed femora, fractures of the long bones, premature fusion of the calvarial sutures, severe midface hypoplasia, proptosis, choanal atresia, and, in some, ambiguous genitalia. Of fewer than 40 patients described to date, most have been sporadic, although reports of parental consanguinity and affected sibs of both sexes suggests autosomal recessive inheritance in some families. Known genetic causes among sporadic cases of ABS or ABS-like syndromes are missense mutations in the IgII and IgIII regions of FGFR2, although the assignment of the diagnosis of ABS to such children has been disputed. A third cause of an ABS-like phenotype is early in utero exposure to fluconazole, an inhibitor of lanosterol 14-alpha-demethylase. The fourth proposed cause of ABS is digenic inheritance combining heterozygosity or homozygosity for steroid 21 hydroxylase deficiency with effects from a second gene at an unknown locus. Because fluconazole is a strong inhibitor of lanosterol 14-alpha-demethylase (CYP51), we evaluated sterol metabolism in lymphoblast cell lines from an ABS patient without a known FGFR2 mutation and from a patient with an FGFR2 mutation and ABS-like manifestations. When grown in the absence of cholesterol to stimulate cholesterol biosynthesis, the cells from the ABS patient with ambiguous genitalia but without an FGFR2 mutation accumulated markedly increased levels of lanosterol and dihydrolanosterol. Although the abnormal sterol profile suggested a deficiency of lanosterol 14-alpha-demethylase, mutational analysis of its gene, CYP51, disclosed no obvious pathogenic mutation in any of its 10 exons or exon intron boundaries. Sterol metabolism in lymphoblasts from the phenotypically unaffected mother was normal. Our results suggest that ABS can occur in a patient with an intrinsic defect of cholesterol biosynthesis at the level of lanosterol 14-alpha-demethylase, although the genetic nature of the deficiency remains to be determined. PMID- 12116246 TI - Novel mutation in the Delta-sterol reductase gene in three Lebanese sibs with Smith-Lemli-Opitz (RSH) syndrome. AB - The Smith-Lemli-Opitz syndrome (SLOS), or RSH syndrome, is a well-characterized multiple congenital anomalies/mental retardation syndrome. The phenotype has been redefined to include mildly affected individuals with minor anomalies and developmental delay, and severe malformations with pre- and perinatal mortality. The condition is due to the deficient activity of the enzyme 7-dehydrocholesterol (7-DHC) reductase [Shefer et al., 1995: J Clin Invest 96:1779-1785], and the gene has been mapped to chromosome 11q13 [Moebius et al., 1998: Proc Natl Acad Sci USA 95:1899-1902]. We describe here a consanguineous family of Syrian-Lebanese ancestry with three sibs affected with SLOS: two with a mild variant, while the other had severe disease and died in the first year of life. Mutation analysis demonstrated a novel mutation in the DHCR7 gene, present in homozygous form in the two affected individuals available for testing, and heterozygous in the parents. The wide intrafamilial variation of clinical severity in these three sibs is an important finding in SLOS. PMID- 12116247 TI - Predicting the risk of cystic fibrosis with abnormal ultrasound signs of fetal bowel: results of a French molecular collaborative study based on 641 prospective cases. AB - Hyperechogenic fetal bowel is prenatally detected by ultrasound during the second trimester of pregnancy in 0.1-1.8% of fetuses. It has been described as a normal variant but has often been associated with severe diseases, notably cystic fibrosis (CF). The aim of our study was to determine the risk of CF in a prospective study of 641 fetuses with ultrasonographically abnormal fetal bowel and the residual risk when only one mutation is detected in the fetus. Fetal cells and/or parental blood cells were screened for CFTR mutations. Two screening steps were used, the first covering the mutations most frequently observed in French CF patients (mutation detection rate of 70-90%) and, when a CF mutation was detected, a DGGE-sequencing strategy. We observed a 3.1% risk of CF when a digestive tract anomaly was prenatally observed at routine ultrasound examination. The risk was higher when hyperechogenicity was associated with bowel dilatation (5/29; 17%) or with the absence of gall bladder (2/8; 25%). The residual risk of CF was 11% when only one CF mutation was detected by the first screening step, thereby justifying in-depth screening. Mutations associated with severe CF (DeltaF508 mutation) were more frequently observed in these ultrasonographically and prenatally detected CF cases. However, the frequency of heterozygous cases was that observed in the normal population, which demonstrates that heterozygous carriers of CF mutations are not at increased risk for hyperechogenic bowel. In conclusion, fetal bowel anomalies indicate a risk of severe cystic fibrosis and justify careful CFTR molecular analysis. PMID- 12116249 TI - Anorectal anomalies associated with or as part of other anomalies. AB - Anorectal anomalies occurring with other anomalies or as part of syndromes were analyzed to determine how their epidemiological characteristics differed from those of isolated anal anomalies. Almost 15% of cases were chromosomal, monogenic or teratogenic syndromes, whereas the rest were of unknown cause including sequences (9.3%), VACTERL associations (15.4%) and multiple congenital anomalies (MCA) (60.2%). Almost half of babies with MCA had one or two VACTERL anomalies with distribution frequencies that did not differ significantly from those in babies with the full VACTERL association. There were considerable differences in the frequency of the VACTERL association among babies with different types of anorectal anomaly. Babies with anal anomalies occurring with sequences, VACTERL or MCA showed the same sex differences as babies with isolated anal anomalies, namely male predominance in anal atresia without fistula or cloaca, no sex difference in anal atresia with fistula, and female predominance in ectopic anus and congenital anal fistula. These anomalies, however, were associated with significantly lower mean gestational lengths and birth weights, and higher frequencies of fetal death and pregnancy termination than babies with isolated anal anomalies. Twins were more frequent in sequences, VACTERL and MCA than in isolated anomalies, monogenic syndromes or chromosome anomalies. Five cases were conjoined twins, representing 15% of all cases of twin pregnancies with an anal anomaly. Indeterminate sex was more frequent in babies with anal atresias without fistula than in those with fistula. Anal anomalies are defects of blastogenesis attributable to disorders in expression of pattern determining genes. The differential sex involvement in different types of anal anomaly may be manifestations of expression of the HY/SRY genes during blastogenesis or of X linkage. PMID- 12116248 TI - HOXD13 polyalanine tract expansion in classical synpolydactyly type Vordingborg. AB - In 1927, Oluf Thomsen, in a classic paper, described a seven-generation family with autosomal dominant axial synpolydactyly (SPD)--the Vordingborgtyp of axis duplication and dysostosis. Expansion of a polyalanine tract in the HOXD13 gene is known to cause synpolydactyly. We have rediscovered part of the family described by Thomsen, and detected a 9 triplet polyalanine expansion within HOXD13segregating with the disorder. The phenotypic spectrum in mutation carriers ranged from severe to inapparent bone malformations. In the latter case, only dermatoglyphics revealed the genetic status. PMID- 12116250 TI - Leukocyte adhesion deficiency (LAD) type II/carbohydrate deficient glycoprotein (CDG) IIc founder effect and genotype/phenotype correlation. AB - Leukocyte adhesion deficiency (LAD) type II is a rare autosomal recessive syndrome characterized by recurrent infections, typical dysmorphic features, the Bombay blood phenotype and severe growth and psychomotor retardation. It is attributed to a general absence of fucosylated glycans on the cell surface. Three Arab Israeli patients and one Turkish child have been reported so far. The primary defect in a specific GDP-L-fucose transporter of the Golgi apparatus has been disclosed recently. All three children reported by us are homozygous for one single founder mutation, different from that reported in the Turkish child. The amount of mRNA of the GDP-L-fucose transporter in cells from Arab patients and their parents are comparable to controls. Genotype/phenotype correlation studies show that the two different mutations are distinguished by differences in response to fucose supplementation and in the clinical phenotypes. PMID- 12116251 TI - Genetic analysis of patients with the Saethre-Chotzen phenotype. AB - Saethre-Chotzen syndrome is a common craniosynostosis syndrome characterized by craniofacial and limb anomalies. Intragenic mutations of the TWIST gene within 7p21 have been identified as a cause of this disorder. There is phenotypic overlap with other craniosynostosis syndromes, and intragenic mutations in FGFR2 (fibroblast growth factor receptor 2) and FGFR3 (fibroblast growth factor receptor 3) have been demonstrated in the other conditions. Furthermore, complete gene deletions of TWIST have also been found in a significant proportion of patients with Saethre-Chotzen syndrome. We investigated 11 patients clinically identified as having the Saethre-Chotzen phenotype and 4 patients with craniosynostosis but without a clear diagnosis. Of the patients with the Saethre Chotzen phenotype, four were found to carry the FGFR3 P250R mutation, three were found to be heterozygous for three different novel mutations in the coding region of TWIST, and two were found to have a deletion of one copy of the entire TWIST gene. Developmental delay was a distinguishing feature of the patients with deletions, compared to patients with intragenic mutations of TWIST, in agreement with the results of Johnson et al. [1998: Am J Hum Genet 63:1282-1293]. No mutations were found for the four patients with craniosynostosis without a clear diagnosis. Therefore, 9 of our 11 patients (82%) with the Saethre-Chotzen phenotype had detectable genetic changes in FGFR3 or TWIST. We propose that initial screening for the FGFR3 P250R mutation, followed by sequencing of TWIST and then fluorescence in situ hybridization (FISH) for deletion detection of TWIST, is sufficient to detect mutations in > 80% of patients with the Saethre Chotzen phenotype. PMID- 12116252 TI - A 52-kb deletion in the SOST-MEOX1 intergenic region on 17q12-q21 is associated with van Buchem disease in the Dutch population. AB - Van Buchem disease is an autosomal recessive sclerosing bone dysplasia characterized by skeletal hyperostosis, overgrowth of the mandible, and a liability to entrapment of the seventh and eighth cranial nerves. The genetic determinant maps to chromosome 17q12-q21. We refined the critical interval to the < 1-Mb region between D17S2250 and D17S2253 in 15 affected individuals, all of whom shared a common disease haplotype. Furthermore, we report here the identification of a 52-kb deletion located within the interval and encompassing D17S1789 that is 100% concordant with the disorder. Although the deletion itself does not appear to disrupt the coding region of any known or novel gene(s), the closest flanking genes are MEOX1 on the proximal side, and SOST on the distal side of the deletion. MEOX1 is known to be important for the development of the axial skeleton, whereas the SOST gene is the determinant of sclerosteosis, a disorder that shares many features with van Buchem disease, thus raising the possibility that van Buchem disease results from dysregulation of the expression of one or both of these genes. PMID- 12116253 TI - Pseudodominant inheritance of Langer mesomelic dysplasia caused by a SHOX homeobox missense mutation. AB - We report the clinical and molecular analysis in a consanguineous family in which the skeletal dysplasias Leri-Weill dyschondrosteosis (LWD) and Langer mesomelic dysplasia (LMD) both segregate. A newborn male and his mother, both with Langer mesomelic dysplasia, are described. A homozygous SHOX homeobox point mutation, C517T, was identified by direct sequencing in the proband and his mother. The same mutation was present in the heterozygous state in the proband's father and in the maternal grandmother, both of whom had features of LWD. This C to T transition is predicted to cause an arginine to cysteine amino acid change in a highly conserved region of the recognition helix of the homeodomain, which may reduce the stability of the interaction between the SHOX protein and its target DNA. In addition, the mutation may disrupt a nuclear localization signal in SHOX. This is the first SHOX point mutation identified in a case of LMD, and the first case in which parent to child transmission of LMD has been described. PMID- 12116255 TI - Bilateral nephroblastoma in familial Hay-Wells syndrome associated with familial reticulate pigmentation of the skin. AB - We report on a girl with maxillary hypoplasia, prominent ears, dry sparse hair, palmar and plantar keratoderma, dystrophic nails, patchy pigmented skin lesions in hands and feet and bilateral Wilms tumor. She was born with bilateral ankyloblepharon. The mother and maternal grandmother presented similar ectodermal defects. Skin biopsies of the patient and her mother proved to contain cells overexpressing p63 by immunohistochemistry. Karyotypes of the patient and her mother, and FISH studies on lymphocytes and tumor cells of the girl demonstrated a mosaic 11p15.5 deletion. These findings suggest a relationship between familial ankyloblepharon, ectodermal defects and cleft lip and palate (AEC) syndrome (Hay Wells syndrome) and familial reticulate pigmentation of the skin. In addition the development of Wilms tumor and 11p15.5 region involvement expand the genetic relationship between these conditions and the enlarging group of genetic entities related to nephroblastoma. PMID- 12116254 TI - Complete SHOX deficiency causes Langer mesomelic dysplasia. AB - The SHOX (short-stature homeobox-containing) gene encodes isoforms of a homeodomain transcription factor important in human limb development. SHOX haploinsufficiency has been implicated in three human growth disorders: Turner syndrome, idiopathic short stature, and Leri-Weill dyschondrosteosis. Langer mesomelic dysplasia is thought to be the homozygous form of dyschondrosteosis. However, complete SHOX deficiency has not been demonstrated for any postnatal patient with the classic Langer phenotype. We studied four adults and one child with Langer mesomelic dysplasia. SHOX abnormalities were detected in all five probands. One was a homozygote or hemizygote and two were compound heterozygotes. The homozygous or hemizygous mutation was in exon 6a, implying that the SHOXa isoform is essential for normal skeletal development. These findings confirm clinical inferences that Langer mesomelic dysplasia is the homozygous form of Leri-Weill dyschondrosteosis and add to our understanding of genotype/phenotype relationships in SHOX deficiency disorders. PMID- 12116257 TI - "Apple-peel" intestinal atresia, ocular anomalies, and microcephaly syndrome: brain magnetic resonance imaging study. AB - We report on a male child with "apple-peel" atresia associated with microcephaly and ocular anomalies. To date, no magnetic resonance imagings have been published. We report on the fourth reported case with this phenotype, but the first to be studied by brain magnetic resonance imaging. PMID- 12116256 TI - Giant platelets in a case of deletion 11q24-qter confirmed by fluorescence in situ hybridization. AB - Here we report the association of giant platelets and an increase in platelet volume in a 19-month-old black female with de novo del 11q24-qter. The deletion, which was visible on karyotype, was further confirmed and more precisely localized by fluorescence in situ hybridization studies (FISH) that showed the deletion to lie distal to the MLL gene region (11q23). Clinically, the case presented less severe symptoms than Jacobsen syndrome-the well known partial deletion of the distal end of chromosome 11. Platelet glycoproteins CD 41, CD 42a, C 42b, CD 61, and PAC-1 were also assayed and found to be normally expressed. To our knowledge, giant platelets are described for the first time in the relevant deleted region. PMID- 12116258 TI - Association of external auditory canal atresia, vertical talus, and hypertelorism: confirmation of Rasmussen syndrome. AB - In 1979, Rasmussen et al. reported six members of a family with congenital, bilateral, symmetrical, and isolated subtotal atresia of the external auditory canal, bilateral foot abnormalities, and increased interocular distance. The family history suggested autosomal dominant inheritance of the syndrome. We report a 3-year-old girl whose symptoms are compatible with this diagnosis. Therefore, we suggest confirmation of the description by Rasmussen et al. as a distinct entity and suggest the term Rasmussen syndrome for this condition. PMID- 12116259 TI - Phenotypic discordance in a family with monozygotic twins and nonsyndromic cleft lip and palate: follow-up. PMID- 12116260 TI - Ectrodactyly and Germany's eugenics law of 14 July 1933. AB - The family reported herein serves as a genetically and historically important vignette on the issues of nonpenetrance (versus germinal mosaicism) in nonsyndromic autosomal dominant ectrodactyly and the Eugenics Law of Germany of 14 July 1933, which was used to coerce the sterilization of the propositus despite infertility in his first marriage. In a sibship of seven children (with normal parents), three boys were affected. The propositus (adoptive grandfather of the author) was the patient of Paul Leopold Friedrich and Georg Perthes, who published their observations on the propositus. Except for an adopted daughter, the propositus was childless. His two affected brothers each had an affected child, and the father- to son transmission confirmed the hypothesis of autosomal dominant inheritance. The issue of nonpenetrance versus germinal mosaicism in ectrodactyly was debated by Auerbach [1956:Ann Hum Genet 20:266-269] and Vogel [1958:Ann Hum Genet 22:132-137], and remains unresolved. PMID- 12116261 TI - Response to "genetic polymorphisms of methylenetetrahydrofolate reductase (MTHFR) and methionine synthase reductase (MTRR) in ethnic populations in Texas; a report of a novel MTHFR polymorphic site, G1793A". PMID- 12116264 TI - Genetic counseling for sex chromosome abnormalities. AB - Sex chromosome abnormalities (SCAs) are the most frequently occurring chromosomal abnormalities encountered at both prenatal diagnosis and at birth. Approximately 1/400 newborns has an SCA, and incidence at prenatal diagnosis is even greater, 1/250 to 1/300. Physicians and health providers from various specialties are encountering diagnoses of SCAs with increased frequency as more individuals are becoming identified, both prenatally and postnatally. Because these conditions generally have relatively few serious physical implications and because they are extremely variable, genetic counseling is often more complex and challenging than that occurring with an autosomal abnormality. It is imperative that health professionals have the knowledge of content and methodology to provide appropriate counseling to such individuals and their families. During the period from 1964 to 1975, seven international groups (including the Denver group) screened a total of 199,898 consecutive births and identified 307 individuals with SCA. The Denver group has followed more than 40 such individuals from birth to adulthood. In addition, the Denver group has experience in counseling over 1,000 families with a prenatal diagnosis of SCA. Based on these studies and contacts, guidelines for the counseling of individuals and families with SCA are provided. Accurate information must be presented and the variability and imprecise prognosis recognized. Successful counseling strategies include interfamily contact, viewing photographs, and utilizing support groups. Issues of disclosure, follow-up, and anticipatory guidance should be addressed. PMID- 12116265 TI - Fifty-one prenatally diagnosed children and adolescents with sex chromosome abnormalities. AB - Children with sex chromosome abnormalities (SCA) are known to be at risk for developmental delays. These risks were identified 2 decades ago by seven international research groups who prospectively followed children ascertained after birth. Subsequently, some of these investigators suggested the course of prenatally identified children with SCA may be different from children in earlier studies. The first such evidence was published by Robinson et al. [1992: Am J Med Genet 44:365-368], who compared 20 prenatally diagnosed children to the original postnatally diagnosed cohort. The following report presents an update and expansion of that research and includes 51 children and adolescents prenatally diagnosed with SCA, now 7-18 years of age. Results confirm that this cohort of prenatally diagnosed children has a milder developmental course than children ascertained postnatally. The study provides new information to health professionals counseling families faced with prenatal diagnosis of SCA. PMID- 12116266 TI - Significant impact of the highly informative (CA)n repeat polymorphism of the APOA-II gene on the plasma APOA-II concentrations and HDL subfractions: The ECTIM study. AB - High density lipoproteins (HDL) are heterogeneous in their apolipoprotein composition and the role of apolipoprotein A-II (APOA-II) in HDL structure and metabolism is poorly understood. Yet, studies of naturally occurring variations of APOA-II in mice and experiments in transgenic mice overexpressing the APOA-II gene (APOA-II) have shown that APOA-II expression influences APOA-II plasma levels and HDL size and composition. In humans, two RFLPs (BstNI and MspI) have been described in the APOA-II gene. These RFLPs, however, have been inconstantly associated with variations in APOA-II plasma levels. In particular, the large multicentric ECTIM Study did not show any significant effect of the two RFLPs. Other polymorphisms consisting of repetitive sequences have been proposed as more informative markers than RFLPs. Thus, data from the ECTIM Study were reconsidered by integrating the additional information obtained from a highly informative multiallelic (CA)(n)-repeat polymorphism located in the second intron of the gene. The population study was composed of 763 non-treated male controls and 594 cases of myocardial infarction. In controls, the (CA)(19) allele was associated with significantly decreased APOA-II (P < 0.0009) and LpA-II:A-I (P < 0.02) plasma levels. Although the APOA-I plasma levels were not affected by the polymorphism, the (CA)(19) allele was associated with an increased LpA-I/LpA-II:A I ratio (P < 0.004). No effect, however, could be detected on myocardial infarction. Study of the linkage disequilibrium and the estimation of haplotype frequencies indicated that the impact of the APOA-II locus could hardly be detected by using the BstNI and MspI RFLPs. These data revive interest in evaluating the role of the APOA-II locus in the control of APOA-II plasma levels and HDL composition. PMID- 12116267 TI - C7 complement deficiency in an Israeli Arab village. AB - Deficiencies of terminal complement components, particularly the latter ones, are often detected because of increased susceptibility to Neisserial infections. Herein we document the first report of C7 deficiency among a highly inbred Arab population living in the lower Galilee region of Israel. Both biochemical and molecular analysis were performed on samples from infected survivors and parents of children who succumbed to Neisserial infections in a 4-year period. Only the index case who suffered recurrent infections and a sibling who had not suffered an infection during the outbreak were found to be C7-deficient. The mutation was found to be the one previously described to be prevalent among Israeli Jews of Moroccan ancestry (mutation G1135C). The implications of this finding are discussed in the context of family pedigree, the protective effect of complement deficiency, and the clinical outcome. PMID- 12116268 TI - Lack of association of the two polymorphisms in alpha-2 macroglobulin with Alzheimer disease. AB - Alzheimer disease is a complex neurodegenerative disorder that is characterized by cognitive decline and distinct neuropathology. The gene for alpha-2 macroglobulin (A2M) on chromosome 12 has two polymorphisms that, in some cases, are associated with Alzheimer disease. We examined these two polymorphisms in our families in the DNA Bank (a collection of DNA samples from families with Alzheimer disease in Texas). Using both association studies and sib transmission/disequilibrium tests, we found no association of the two polymorphisms with the disease in our collection. In addition, we did not find an association of the disease with the two polymorphisms in A2M in a small subset of National Institute of Mental Health (NIMH) families. Thus, our studies do not support the A2M polymorphisms as a risk factor for Alzheimer disease. PMID- 12116269 TI - Carrier testing in fragile X syndrome: when to tell and test. AB - This study explored age preferences about when to learn at-risk status and have carrier testing in women who were undergoing testing for fragile X. Forty-two women (20 carriers and 22 noncarriers) completed a structured interview prior to carrier testing and after learning their result. The majority favored learning at risk status and carrier status at < 18 years for themselves and their children. Preferred ages fell into four developmental categories: early childhood (0-9), preteen (10-13), teen (14-17), and adult (>or= 18). Although no significant mean changes in age responses were found between interviews or between carrier and noncarrier responses, a difference in the pattern of responses related to age categories was suggested. There appeared to be an increase in the number of responses in the 0-9 category at time 2. Also, the mean ages for testing were older than they were for telling at time 1, but not at time 2. For women indicating ages 0-9, the most frequent reason was to provide children with time to adjust. Those reporting ages 10-13 felt that the onset of puberty as well as a child's ability to understand, adjust, and cope were key determinants. Those preferring the teen years felt the possibility of sexual activity and planning for the future were important considerations. The developmental focus for adults was serious relationships. This study, through its unique longitudinal perspective of transition from uncertainty to certainty, builds on prior knowledge, has implications for genetic counseling, and suggests that the developmental stage of the child is important in determining when to tell and test. PMID- 12116270 TI - Relationship of the delta-opioid receptor gene to heroin abuse in a large Chinese case/control sample. AB - Pharmacological and electrophysiological evidence has shown that opioid receptors are involved in the mechanism of heroin dependence. Thus, opioid receptors are appropriate candidate genes for case-control association studies of heroin dependence. Previously, two single nucleotide polymorphisms (SNPs), OPRD1 921T > C and 80G > T, of the human delta opioid receptor gene were used in population based studies of heroin dependence. One study in a German population found that OPRD1 921T > C was associated with heroin dependence. This finding, however, was not replicated in a different German sample. To test the hypothesis that OPRD1 or a closely linked gene is associated with heroin dependence, we used 5' nuclease assays to genotype both OPRD1 SNPs in 450 Chinese heroin dependent patients and 304 unaffected controls from the same population. In addition, five SNPs distributed in four other genes: ADH2, ALDH2, OPRM1, and DRD1, were used as genomic control loci to test the case and control populations for stratification bias. Genotype and allele frequencies at OPRD1 921T > C were not significantly different, and the OPRD1 80G was absent from both Chinese opioid dependence patients and controls. Based on the genotype and allele frequencies of the genomic control loci, there was no evidence for stratification bias capable of masking an association of OPRD1 to heroin dependence in this large and homogenous Chinese sample. Therefore, these data do not support an association between the OPRD1 gene and heroin dependence in the Chinese population. PMID- 12116271 TI - Additional patient with del(12)(q21.2q22): further evidence for a candidate region for cardio-facio-cutaneous syndrome? AB - Cardio-facio-cutaneous (CFC) syndrome is characterized by a distinct facial appearance, cardiac defects, ectodermal anomalies and developmental delay. Recently, we reported a 19-month-old girl with phenotypic manifestations consistent with the CFC syndrome who had an interstitial deletion of the long arm of chromosome 12, del(12)(q21.2q22), implicating a possible locus for CFC syndrome. Here, we report an additional patient with a cytogenetically identical interstitial deletion: 47,XYY,del(12)(q21.2q22). To further characterize this deletion we used microarray-based comparative genomic hybridization (array CGH). Array CGH confirmed both the deletion and the second Y chromosome. The deletion on chromosome 12q spanned at least 14 Mb as indicated by the positions on the genome sequence of the 4 BAC clones included in the deletion. While the proband did not have the classic features of CFC, he had some dysmorphic craniofacial characteristics, ectodermal anomalies and moderate developmental delay which were suggestive of CFC syndrome; however, this patient did not have classical CFC. The phenotypic differences between the two del(12)(q21.2q22) patients may be due to variability in the expression of the syndrome, or this deletion may present as a syndrome with overlapping features. Alternatively, the phenotypic differences may result from discordance at the molecular level, which may yield a critical minimal region of deletion for CFC. The region 12q21.2 --> q22 remains a possible candidate region for CFC syndrome. Additional characterization of these and other CFC patients may confirm and further refine this candidate region. PMID- 12116273 TI - Anticipation in migraine with affective psychosis. AB - Anticipation is the term given to the apparent occurrence of an inherited disorder at a progressively earlier age of onset in successive generations. A family of a mother and two children, a 17-year-old girl and a 13-year-old boy, all experienced migraine with affective psychosis. There is a strong genetic predisposition to the psychiatric disorder of affective psychosis along with a dominant pattern of migraine in the family, which suggests a genetic connection between migraine and affective psychosis. PMID- 12116272 TI - Pearson marrow-pancreas syndrome with worsening cardiac function caused by pleiotropic rearrangement of mitochondrial DNA. AB - Pearson marrow-pancreas syndrome is a usually fatal disorder that involves the hematopoietic system, exocrine pancreas, liver, kidneys, and often presents clinically with failure to thrive. We report a 5-year-old patient who developed, in addition to the typical features of Pearson syndrome, worsening cardiac function, mainly affecting the left ventricle. The latter finding is particularly interesting because cardiac involvement has not yet been regarded as a major feature of Pearson syndrome. The diagnosis was proved by the finding of so far undescribed pleioplasmatic rearrangement of mitochondrial (mt)DNA (loss of 5,630 bp, 70% deleted and duplicated mtDNA) in blood cells. Our report demonstrates that patients with Pearson syndrome may also have impaired cardiac function. Thus, Pearson syndrome should be considered in the differential diagnosis of patients with left ventricular dysfunction of unknown origin and other clinical findings suggestive of a mitochondrial disease. PMID- 12116274 TI - Further delineation of the chromosome 14q terminal deletion syndrome. AB - A patient with hypotonia, blepharophimosis, ptosis, a bulbous nose, a long philtrum, upturned corners of the mouth, and mild developmental delay was found to have a small subtelomeric deletion of the long arm of chromosome 14 (q32.31 qter). In comparing her phenotype with previously reported patients with similar 14q deletions, due to either a linear deletion or to a ring chromosome 14, a clinically recognizable terminal 14q microdeletion syndrome was evident. Due to the limited number of cases reported, it was not possible to assign specific features to specific regions of terminal 14q. The comparison of features in cases with a linear deletion of 14qter (n = 19) to those in cases with a deletion due to a ring chromosome 14 (n = 23), both with the same breakpoint in 14q, showed that seizures and retinitis pigmentosa have been found only in patients with ring chromosomes. Several hypotheses are put forward to explain this difference: mitotic instability of ring chromosomes; a telomere position effect in ring chromosomes in which the 14p telomere silences nearby gene(s) on the q-arm; and dose-dependent gene(s) involved in seizures and retinitis pigmentosa located on the short arm of chromosome 14. In our opinion, only seizures may be explained by the mitotic instability of ring chromosomes, while both seizures and retinitis pigmentosa may be explained by silencing of gene(s) on 14q by the 14p telomere; the third hypothesis seems unlikely to explain either symptom. PMID- 12116275 TI - Terminal deletion of the chromosome 7(q36-qter) in an infant with sacral agenesis and anterior myelomeningocele. AB - We report on a new patient with a 7q terminal deletion. The 18-month-old boy had metal retardation, microcephaly, a distinctive face, bilateral coloboma, cafe-au lait spot on the abdomen, and sacral agenesis. The high resolution GTG bands (550 850 bands), currently used in our laboratory, showed a 7q terminal deletion, which was also confirmed with fluorescence in situ hybridization (FISH). The homeobox HLXB9 gene, localized at 7q36 has been demonstrated to be involved in sacral agenesis; in fact patients with 7q terminal deletions frequently have this malformation. We could not perform molecular studies in this patient to confirm the HLXB9 haploinsufficiency, but we postulate that he carried it. PMID- 12116276 TI - Complete overlap of PHACE syndrome and sternal malformation--vascular dysplasia association. AB - PHACE syndrome is the term applied to the association of posterior fossa brain abnormalities, hemangiomas, arterial anomalies in the cranial vasculature, coarctation of the aorta/cardiac defects, and eye abnormalities. An overlap with the sternal malformation/vascular dysplasia association has been described. We report an adult patient with complete manifestations of both conditions. As an adult she has demonstrated resolution of the hemangiomas and only mild intellectual difficulties. PMID- 12116277 TI - A novel germline point mutation, c.2304 G-->T, in codon 768 of the RET proto oncogene in a patient with medullary thyroid carcinoma. PMID- 12116278 TI - Festschrift for Erik V. Stalberg, MD, PhD. PMID- 12116280 TI - Early struggles with single-fiber electromyography. AB - The development of single-fiber electromyography is described. This method, introduced in 1963, was met with doubts, and there were strong arguments that subunits, 10-30 synchronized muscle fibers, were in fact being recorded. The dispute ended in 1971, and single-fiber electromyography is now generally accepted. PMID- 12116281 TI - Clinical impact of single-fiber electromyography. AB - The major clinical impact of single-fiber electromyography has been from its role in confirming, or excluding, the diagnosis of myasthenia gravis (MG). Jitter measurements also have a clinical role in demonstrating changes in disease severity in patients with MG and Lambert-Eaton myasthenic syndrome, in demonstrating subtle changes in motor unit architecture and physiology in patients with nerve and muscle diseases, and in demonstrating the remote effects of locally injected botulinum toxin. In addition to these clinical roles, the ability to identify the activity from single muscle fibers makes it possible to mark the discharges of single motor units. This, along with information gained by jitter and fiber-density measurements, has uniquely increased our understanding of motor unit organization and function in normal and disease states. PMID- 12116282 TI - Safety margin at single neuromuscular junctions. AB - Jitter measurement with axonal microstimulation was used to study synaptic function at 115 neuromuscular junctions (NMJs) of normal subjects at various stimulation rates. Jitter was lowest at 0.5 Hz; it increased slightly at 1, 2, and 5 Hz and remained at that level at 10 Hz (a light work load) and 20 Hz (a heavy work load); and it increased further at 50 Hz (an extreme load). This pattern was seen for the majority of the NMJs, suggesting a high safety factor of neuromuscular transmission maintained rather uniformly over a wide range of discharge rates. A proportion of the normal NMJs had relatively large jitter; these tended to show prominent facilitation as the rate was raised from 5 or 10 to 20 Hz. Similar but more dramatic facilitation improving the safety factor was seen at most NMJs in myasthenia, which was studied for comparison. Such facilitation was not found at normal NMJs with low jitter. PMID- 12116283 TI - Microelectrode recordings from human peripheral nerves (microneurography). AB - The development of microneurography, which began in the laboratory of clinical neurophysiology in Uppsala, is described. PMID- 12116284 TI - Macroelectromyography: a review of the technique and its value in the investigation of neuromuscular disorders. AB - The background for the macroelectromyography technique, which was developed by Erik Stalberg to measure the size of motor unit potentials in human muscles, is reviewed. The method employs a modified single-fiber electrode with a large nonselective recording surface capable of recording the activity from all the fibers of a motor unit. The findings in normal subjects and its application in the study of motor unit recruitment are described. The value of the technique in the investigation, diagnosis, and monitoring of various neuromuscular diseases, including primary myopathies and neurogenic conditions, is discussed. PMID- 12116285 TI - Models and simulations in electromyography. AB - In electromyography, one assesses the pathophysiology on the basis of the waveform characteristics of the recorded signal. This requires detailed knowledge of the relationship between the waveform generators and the waveform measurements. Models and computer simulations can be used to explore this relationship in an efficient manner. Combining models with experimental methods will allow us to define new measurements and new rules of interpretation. This is discussed with some of the models developed for electromyography signal analysis. PMID- 12116286 TI - Isaacs' syndrome as a potassium channelopathy of the nerve. AB - Isaacs' syndrome (acquired neuromyotonia) is an antibody-mediated potassium channel disorder (channelopathy). The target channel proteins of the antigens are voltage-gated potassium channels (VGKCs), especially dendrotoxin-sensitive fast potassium channels. The suppression of voltage-gated outward K(+) current by antibodies induces hyperexcitability of the peripheral nerve. Patch clamp studies show that antibodies may not directly block the kinetics of VGKCs but may decrease channel density. Electrophysiological, pharmacological, and immunological findings indicate that the site of origin of spontaneous discharges is principally in the distal portion of the motor nerve and/or within the terminal arborization. The spectrum of potassium channelopathies is expanding. The existence of antibodies against VGKCs should be considered in patients who present with generalized nerve hyperexcitability of undetermined etiology. PMID- 12116287 TI - Neurography: late responses. AB - The meaningfulness of routine conduction velocity studies can be increased when so-called late responses (F waves, H reflex, and intermediate late responses) are considered. The techniques to elicit different types of late responses are described as well as their occurrence in physiological and pathological conditions. Late responses are muscle action potentials of different origins and different configurations, and they have different clinical implications. F waves are recurrent discharges of alpha-motor neurons that have diagnostic value in patients with demyelinating neuropathies and proximal lesions of peripheral nerves. H reflexes are similar to the muscle stretch reflex. H reflexes can be elicited in only a few nerves, so they are routinely used only in patients suffering from sacral plexopathies, S1 radiculopathy, and polyneuropathies. Intermediate late discharges are of various origins and clinical significance. True axon reflexes occur seldom in routine neurography and are usually caused by submaximal stimulation. However, A waves with a constant shape, latency, and configuration are often found in patients with polyneuropathies and may be early signs of acute inflammatory demyelinating neuropathy (Guillain-Barre syndrome). PMID- 12116288 TI - Scanning electromyography. AB - A special electromyography (EMG) method, scanning EMG, was introduced by Stalberg and Antoni in 1980 to study the electrophysiological cross sections and sizes of motor units. Scanning EMG gives a new approach for the evaluation of the electrical properties of motor units, providing new data on the normal anatomical distribution of muscle fibers and its changes in different pathologies of the muscle. The description of scanning EMG recordings required the introduction of new parameters (lengths of motor unit cross sections, fractions of motor units, and silent areas), in addition to those used with conventional EMG recordings, and the traditional parameters (duration, amplitude, etc.) acquired new and more accurate explanations. Normal scanning EMG recordings are available for biceps brachii, anterior tibial, and masseter muscles. The findings in normal muscles agree with the nonrandom distribution of muscle fibers in motor units and confirm the suggestion that muscle fibers within motor units tend to be arranged in clusters. In muscular dystrophies, the sizes of motor unit territories do not differ significantly from the normal values. However, the configuration of motor units changes considerably. Abrupt changes in amplitude and duration, segments of short and long duration, increased numbers of fractions, and silent areas have been revealed, showing that dystrophic motor units are definitely fragmented. Scanning EMG supports the assumption that there is clustering of muscle fibers within the dystrophic motor unit, with local grouping of muscle fibers. In neurogenic lesions, the length of motor units is normal or only slightly increased. Reinnervated motor units are restricted to the fascicles in which they are originally found. Reinnervation does not result in an increase in the number of fractions, but the amplitude of the potentials, the length of polyphasic sections, and the duration increase. The increase in the number and length of polyphasic sections can differentiate normal motor units from abnormal ones. However, other features (amplitude, duration, number of fractions, and presence of silent areas) are also necessary to distinguish neurogenic processes from myogenic ones. PMID- 12116289 TI - Defining extensible markup language standards for electromyography data transmission across the world-wide web. AB - Different electromyography (EMG) machines store their data in different formats that vary from manufacturer to manufacturer and even between different EMG machines from the same manufacturer. As advanced as these machines are today, it is necessary in most cases to use faxes, scanners, or a common interface such as Adobe's file format to exchange data between them. A possible solution to this problem is to translate the EMG machine's data output into a common format that can be read (and displayed) by commonly available programs or, even better, by a Web browser. We describe an EMG-extensible markup language (XML) translation program that takes proprietary data from an EMG machine and translates the data into XML, a common language format. Predefined extensible stylesheet language (XSL) style sheets allow the display of the data in user-defined formats for EMG machines, personal digital assistants such as Palm Pilots, and even cellular phones. PMID- 12116290 TI - Spontaneous electromyographic activity in spinal cord lesions. AB - Different types of spontaneous activity may be found during electromyographic examinations in patients with spinal cord diseases. Syringomyelia and intramedullary tumor patients may show continuous motor unit activity, synchronous motor unit potentials, myokymic discharges, segmental and propriospinal myoclonus, and respiratory synkinesis. These types of discharges are less commonly encountered in other types of spinal cord lesions. It is suggested that the derangement of inhibitory mechanisms by a central spinal cord lesion may favor the appearance of abnormal spontaneous activity. An increase in the excitability of spinal motor neurons is probably the basic underlying mechanism. PMID- 12116291 TI - Assessment of spinal cord pathology following trauma using early changes in the spinal cord evoked potentials: a pharmacological and morphological study in the rat. AB - The possibility that spinal cord pathology following trauma can be assessed with early changes in the spinal cord evoked potentials (SCEPs) was examined in a rat model. Spinal cord injury (SCI) was produced in Equithesin-anesthetized (3 ml/kg, i.p.) rats through a longitudinal incision into the right dorsal horn at the T10 11 segments. The SCEPs were recorded with epidural electrodes placed over the T9 (rostral) segment of the cord. The SCEPs consisted of a small positive amplitude and a broad and high negative amplitude (NA). SCI resulted in an instant depression of the rostral NA that lasted for 1 h. However, the latency of NA continued to increase over time. At 5 h, spinal cord blood flow declined by 30% in the T9 segment, whereas the spinal cord water content and the permeability of the blood-spinal cord barrier (BSCB) were markedly increased. Damage to the nerve cells, glial cells, and myelin was quite common in the spinal cord, as seen by light and electron microscopy. Pretreatment with p-chlorophenylalanine, indomethacin, ibuprofen, and nimodipine attenuated the SCEP changes immediately after trauma and resulted in a marked reduction in edema formation, BSCB permeability, and blood flow changes at 5 h. However, pretreatment with cyproheptadine, dexamethasone, phentolamine, and propranolol failed to attenuate the SCEP changes after SCI and did not reduce the cord pathology. These observations suggest that early changes in SCEP reflect secondary injury-induced alterations in the cord microenvironment. Obviously, these changes are crucial in determining the ultimate magnitude and severity of cord pathology. PMID- 12116292 TI - What do we learn from motor unit action potentials in surface electromyography? AB - This article gives an overview of what multichannel surface electromyography can teach us about a motor unit. Background information is given about the generation of surface electromyography in general and surface motor unit potentials in particular. Furthermore, we describe how surface motor unit potentials are related to several motor unit characteristics, such as size, location, neuromuscular junction position, fiber length, fiber type, and metabolic fiber properties. In addition, we show how the spatial characteristics of multichannel surface electromyography can be used to obtain single-surface motor unit potentials. The possibilities, challenges, and problems are discussed. Finally, several examples of surface motor unit potential analyses are given. PMID- 12116293 TI - New attempts to quantify concentric needle electromyography. AB - Quantitative motor unit potential (MUP) analysis, which is a leading method of quantitative evaluation of concentric needle electromyography, has several inherent limitations. First, the most essential features of neurogenic or myogenic changes manifest as recruitment abnormalities, rather than as changes in MUP morphology. Second, two factors related to MUP sampling, focusing and level of contraction, greatly influence the parameters of sampled MUPs. Third, the MUP duration, considered to be the cardinal parameter in MUP analysis, has several drawbacks, including low stability and low discriminant sensitivity. We developed a new MUP parameter, the size index (SI), which is calculated from the MUP amplitude and area/amplitude ratio (thickness). The SI remained almost constant during electrode movements, as demonstrated by manual scanning of MUPs. It is a stable and robust parameter and achieved an extremely high ability to discriminate between normal and large neurogenic MUPs. It identifies features related to the sound produced by the MUP on the audio monitor, which is often used by trained electromyographers for qualitative assessments of MUPs. PMID- 12116294 TI - Rippling muscle disease: a review. AB - Rippling muscle disease (RMD) is a benign myopathy with symptoms and signs of muscular hyperexcitability. The typical finding is electrically silent muscle contractions provoked by mechanical stimuli and stretch. After the first description in 1975, there have been several publications on this disorder. Although RMD most often is reported with autosomal dominant inheritance, some sporadic cases are found, and an association with other diseases such as myasthenia gravis has also been reported. The pathophysiological mechanism is still not clarified. Abnormalities in calcium homeostasis in the sarcoplasmic reticulum have been proposed as the most probable causes. However, recent genetic studies make a primary channelopathy unlikely. In this article, a review of this curious disease is presented. PMID- 12116295 TI - What have I learned from single-fiber electromyography? AB - The author recounts the lessons taught by single-fiber electromyography that reached him through the medium of low-frequency attenuation and a small concentric needle electrode. He concludes that it is difficult to imagine practicing electromyography today without the benefits that have flowed from the work of Erik Stalberg and his colleagues. PMID- 12116296 TI - Clinical neurophysiology in Uppsala, 1967-2001. AB - This is a review of the activities at the Department and Institution of Clinical Neurophysiology at Uppsala University Hospital from 1967 to 2001. The most important routine clinical methods are briefly described, and a summary of some of the research projects is given. PMID- 12116297 TI - Small bits to big bites. AB - The Department of Clinical Neurophysiology in Uppsala, Sweden, has reached a high degree of computerization. Patient booking, administration, recording equipment, reporting, and telemedicine are linked components forming an integrated laboratory. Today's configuration is a result of the continuous development and implementation of new technologies. During the 1960s and 1970s, the focus was set on the development of signal analysis procedures. The introduction of personal computers and a local network was the main interest during the 1980s. The 1990s were devoted to the Internet and the development of Keypoint electromyography/evoked potential equipment. PMID- 12116298 TI - Story of electromyography equipment. AB - It all started in 1950 with the introduction of the first commercially available electromyography (EMG) system. From 1950 to 1973 was the era of the analog EMG systems: EMG signals were recorded, and subsequent analyses were carried out manually on film or paper. From 1973 to 1982, the first modular digital EMG systems were introduced. Dedicated analysis modules were introduced, but detailed analysis was still done on paper. In 1982, the first system controlled by a microprocessor was introduced. From 1982 to 1993, many new ways of analyzing EMG signals and basic reporting features were implemented in the EMG systems. Since 1993, personal computer technology has been used in EMG systems. Standard software and hardware components are used to record, analyze, and document EMG examinations. Since 1950, many people have influenced the development of new features in commercial EMG systems. However, within the last 3 decades, Erik Stalberg has always been in the forefront and has shown ways of implementing new methods for analyzing EMG activity or nerve signals. The development of new commercial EMG systems has been dependent on the technology introduced to the market at that particular period of time. This article only refers to systems that have been sold or are now being sold worldwide. PMID- 12116299 TI - What does the neurologist expect from clinical neurophysiology? AB - The future role of clinical neurophysiology is considered in the light of its achievements. It is argued that there is a need to develop methods for specific diagnosis, especially in neuropathies. There is also an unmet requirement for the development of techniques for the prediction of treatment outcomes and for the measurement of changes during the natural history of neuromuscular disorders and their treatment. These issues are not addressed by currently available clinical test methods. PMID- 12116300 TI - What did it give, what does it take? PMID- 12116301 TI - Invasive testing for the karyotyping of mid-trimester intrauterine fetal death (IUFD): a pilot study. AB - INTRODUCTION: Aneuploidy remains a common cause of fetal loss after the first trimester. Conventional karyotyping from fetal solid tissues post-delivery unfortunately has a poor success rate particularly where the fetus is macerated. To overcome this we obtained amniocentesis and/or chorionic villus samples from mid-trimester intrauterine fetal deaths (IUFDs) prior to medical termination of pregnancy. SUBJECTS: Ten women with diagnosed IUFD between 12 and 24 weeks' gestation underwent amniocentesis and/or CVS performed after counselling. RESULTS: Successful karyotypes were obtained in all pregnancies. Five of the ten pregnancies were complicated by aneuploidy (two with trisomy 21, two with trisomy 18, and one with trisomy 13). CONCLUSION: The high rate of aneuploidy (50%) in this small cohort emphasises the need for karyotyping. A successful karyotype in all ten pregnancies demonstrates the value of offering these procedures before a termination of pregnancy. We would recommend the adoption of this approach in the management of IUFD occurring after the first trimester. PMID- 12116302 TI - Improving the prenatal diagnosis of citrullinemia using citrulline/ornithine+arginine ratio in amniotic fluid. AB - Prenatal diagnosis of citrullinemia is performed using a direct argininosuccinate synthetase (ASS) assay on chorionic villi (CV) and citrulline concentration measurement in early amniotic fluid (AF). Here we report the results of 40 prenatal diagnoses performed using this method, discuss the difficulties encountered in interpreting the results, and propose the use of the citrulline/ornithine+arginine ratio (which is more discriminatory than citrulline concentration alone) when performing prenatal diagnosis of citrullinemia. PMID- 12116303 TI - Pilot study for the neonatal screening of fragile X syndrome. AB - Fragile X syndrome (SFX) is the commonest form of inherited mental retardation. Due to the highly variable phenotype clinical diagnosis is complicated. In nearly all cases, the disorder is caused by expansion of a CGG-repeat in the 5' untranslated region of the FMR1 (fragile X mental retardation-1) gene. We have evaluated the feasibility, efficiency and costs of two methodologies in order to develop a simple test to screen large populations: PCR and fragile X mental retardation-1 protein (FMRP) immunodetection. We studied 100 newborn males using PCR and immunodetection (26.91 Euro). All but one amplified the CGG repeat of the FMR1 gene within the normal size range. The sample that failed to amplify showed only 28% of FMRP expression by immunodetection study; both results indicated an affected male. A further 100 males were studied only by polymerase chain reaction (PCR) (7.8 Euro); all of them amplified within the normal size range. Both methodologies, PCR and immunodetection, are feasible for screening large populations, PCR being the most suitable, economical and less time-consuming. However, it is advisable to keep slides for immunodetection when PCR fails or the external control shows no amplification. Early detection of SFX-affected individuals would represent a great benefit for their maximum social integration, due to appropriate treatment and early stimulation and would permit a cascade screening in their pedigree. PMID- 12116305 TI - Pallister-Killian syndrome: difficulties of prenatal diagnosis. AB - The first prenatal diagnosis of Pallister-Killian syndrome (PKS) was reported by Gilgenkrantz et al. in1985. Since this report, about 60 prenatal cases have been reported but both sonographic and cytogenetic diagnoses remain difficult. Although ultrasound anomalies such as congenital diaphragmatic hernia, polyhydramnios and rhizomelic micromelia in association with fetal overgrowth are very suggestive of the syndrome, they are inconstant and they may even be absent. The mosaic distribution of the supernumerary isochromosome 12p greatly increases these difficulties. No prenatal cytogenetic technique is sensitive enough to ensure prenatal diagnosis and false-negative results have been described on fetal blood, chorionic villi and amniocentesis. We report here two prenatal cases of PKS which illustrate the great variability of the fetal phenotype. In reviewing the 63 reported cases, we attempt to determine ultrasound indicators of the syndrome and to define a cytogenetic strategy. In cases where ultrasound indicators are present, our proposal is first to perform chorionic villus or placental sampling and then amniocentesis when the first cytogenetic result is normal. Fetal blood sampling is the least indicated method because of the low frequency of the isochromosome in lymphocytes. In this cytogenetic strategy, fluorescent in situ hybridization (FISH) and especially interphase FISH on non cultured cells increases the probability or identifying the isochromosome. A misdiagnosis remains possible when ultrasound is not contributory; the identification of new discriminating ultrasound indicators would be very helpful in this context. PMID- 12116304 TI - Expandability of haemopoietic progenitors in first trimester fetal and maternal blood: implications for non-invasive prenatal diagnosis. AB - OBJECTIVES: Selective amplification of rare fetal cells in maternal blood is a potential strategy for non-invasive prenatal diagnosis. We assessed the proliferative potential of first trimester fetal progenitors compared to maternal ones. METHODS: Fetal and maternal haemopoietic progenitors were cultured separately and in two model mixtures: (i) co-cultures of male fetal nucleated cells mixed with maternal nucleated cells and (ii) co-cultures of malefetal CD34+ cells with maternal CD34+ cells. Cell origin was detected by X-Y fluorescence in situ hybridisation (FISH) RESULTS: The frequency of haemopoietic progenitors in first trimester fetal blood (predominantly CFU-GEMM) differed from those in peripheral blood from pregnant women (predominantly BFU-e). First trimester haemopoietic progenitors formed larger colonies (p=0.0001) and their haemoglobinisation was accelerated compared to those of maternal origin (p<0.001). CD34+ fetal haemopoietic progenitor cells could be expanded four times more than their maternal counterparts (median 235.8-fold, range 174.0-968.0 vs 71.9-fold, range 41.1-192.0; p=0.003). While selective expansion of fetal cells was not observed in the mononuclear cell model, the CD34+ cell rare event mixtures produced a 463.2-fold (range 128.0-2915.0) expansion of fetal cells. CONCLUSION: Selective expansion of first trimester fetal haemopoietic progenitors may be useful for amplifying fetal cells from maternal blood. PMID- 12116306 TI - The role of hyperglycosylated hCG in trophoblast invasion and the prediction of subsequent pre-eclampsia. AB - OBJECTIVE: Hyperglycosylated hCG (HhCG) is the predominant form of chorionic gonadotrophin in states characterized by aggressive trophoblast invasion such as early pregnancy or choriocarcinoma. Pre-eclampsia may be the result of failed or inadequate trophoblast invasion. We investigated whether low levels of maternal urine HhCG levels would predict subsequent pre-eclampsia. STUDY DESIGN: Mid trimester urine (14-21 weeks) was collected and frozen from non-hypertensive women undergoing genetic amniocentesis. Inclusion criteria were: normal singleton pregnancies without a prior history of pre-eclampsia, hypertension, diabetes or other vascular disorders. The specimens were subsequently thawed, and HhCG levels standardized to urine creatinine were measured. Maternal charts were reviewed after delivery to determine the development of pre-eclampsia. There were a total of 568 study subjects. RESULTS: Pre-eclampsia developed in 26 (4.6%) women. There was a significant correlation between low urine HhCG and subsequent pre-eclampsia (Mantel-Haenszel test of linear association: Chi-square 10.52, p=0.001). The mean HhCG level (ng/mg creatinine) was significantly greater in normals than in those destined to develop pre-eclampsia: 42.7 versus 20.3, p=0.002 (Mann-Whitney U test). There was a progressive increase in the risk of subsequent pre-eclampsia as HhCG levels fell: HhCG < or =0.9 MoM RR (95% CI)=1.51 (1.15-1.98) compared with < or =0.1 MoM 10.42 (2.0-54.3). CONCLUSION: Low maternal mid-trimester urine HhCG predicted subsequent pre-eclampsia. This appears to support the view that pre-eclampsia results at least in part from poor trophoblast invasion. Thus, HhCG may play a role in trophoblast invasion and measurement of this in urine identifies women at high risk for developing pre-eclampsia. PMID- 12116307 TI - An increase in cost-effectiveness of first trimester maternal screening programmes for fetal chromosome anomalies is obtained by contingent testing. AB - We assessed the discriminatory efficiency and cost-effectiveness of a novel way of organising first trimester screening for Down syndrome (DS), contingent testing, where a serological test (PAPP-A and beta-hCG: the double test) is made in early first trimester and followed by nuchal translucency testing (NT) only in women with an intermediate risk, e.g. <1:65 and >1:1000, and not in all women as in normal first trimester screening (NFTS). Using Monte Carlo simulation contingent testing had a detection rate (DR) of 78.9% and a false-positive rate (FPR) of 4.0% for DS with 19.4% of women offered NT testing. The DR of NFTS was 85.5% and the FPR 4.4%. The decrease in NT screening was associated with an increase from 23% to 29% in the proportion of DS cases born. The cost of the contingent testing programme was pound 53,000 per DS case not born and pound 91,000 in NFTS. The number of aborted fetuses per DS case were 0.35 and 0.36, respectively. Thus, contingent testing is an organisation of first trimester screening where costs can be reduced with a marginal decrease in performance. Contingent testing is attractive in areas where NT screening is the bottleneck preventing the introduction of first trimester screening. PMID- 12116308 TI - First prenatal diagnosis of partial trisomy 10 and partial monosomy 15 derived from a maternal translocation (10;15)(q11;q13). PMID- 12116310 TI - Brain anomalies associated with 47,XYY karyotypes detected on a prenatal scan. PMID- 12116309 TI - In utero first trimester exposure to low-dose methotrexate with increased fetal nuchal translucency and associated malformations. PMID- 12116311 TI - Trisomy 18 in a fetus with normal NT and abnormal maternal serum biochemistry. PMID- 12116312 TI - Mucopolysaccharidosis VII (Sly disease) as a cause of increased nuchal translucency and non-immune fetal hydrops: study of a family and technical approach to prenatal diagnosis in early and late pregnancy. PMID- 12116313 TI - Varix of the portal vein: prenatal diagnosis in a fetus with mosaic trisomy 9 syndrome. PMID- 12116314 TI - Two more possible pitfalls of rapid prenatal diagnostics using interphase nuclei. PMID- 12116315 TI - Application of monopolar thermocoagulation in an acardiac fetus. PMID- 12116316 TI - Non-disclosing preimplantation genetic diagnosis for Huntington disease. AB - OBJECTIVES: Individuals at risk for Huntington disease face difficult decisions regarding their reproductive options. Most do not wish to pass on the gene for Huntington disease to their children, but may not be prepared themselves to undergo presymptomatic testing and learn their genetic status. For these reasons, many at-risk individuals with a family history of HD would choose a method of genetic diagnosis that would assure them that they can have children unaffected with HD without revealing their own genetic status (non-disclosing). We have shown that, with a carefully designed and executed programme of non-disclosing preimplantation genetic testing, one can successfully assist at-risk couples to have their own biological children who are free from Huntington disease, without forcing parents to confront knowledge of their own genetic status. METHODS: Couples where one partner was at 50% risk for Huntington disease underwent in vitro fertilization with preimplantation embryo biopsy and molecular analysis for Huntington disease where appropriate. RESULTS: After extensive counselling and informed consent, 10 couples underwent 13 in vitro fertilization and two frozen embryo transfer cycles in a programme for non-disclosing preimplantation genetic diagnosis for Huntington disease. In 11 cycles, embryos determined to be free of Huntington disease were transferred, resulting in five clinical pregnancies. One set of twins and three singleton pregnancies have delivered. One pregnancy resulted in a first-trimester loss. CONCLUSIONS: The option of non-disclosing preimplantation genetic diagnosis should be reviewed, along with other relevant medical options, when counselling at-risk Huntington disease families. PMID- 12116317 TI - Attitude of at-risk subjects towards preimplantation genetic diagnosis of alpha- and beta-thalassaemias in Hong Kong. AB - OBJECTIVES: The aim of the study was to assess whether preimplantation genetic diagnosis (PGD) was an acceptable alternative to prenatal diagnosis in couples at risk of giving birth to a child with alpha- or beta-thalassaemia in an Asian population. METHODS: An information leaflet was distributed to the women at risk. They were asked to complete a questionnaire after having an interview with a designated investigator. RESULTS: A total of 141 valid questionnaires were analysed; 82.3% of the women considered PGD either the same or better than conventional prenatal diagnosis. Women with an affected child or a subfertility problem were more willing to accept PGD and to undergo this procedure in their future pregnancies. Their main concern about PGD was damage to the embryo during the PGD procedure. The most important perceived advantage of PGD was avoidance of termination of an affected pregnancy. CONCLUSIONS: PGD is an acceptable alternative to conventional prenatal diagnosis in women at risk of giving birth to a child with alpha- or beta-thalassaemia in an Asian population. This is particularly true in women with a subfertility problem and in women who already have an affected child. PMID- 12116318 TI - Preimplantation genetic diagnosis for aneuploidy screening in early human embryos: a review. AB - Embryonic aneuploidies may be responsible for pregnancy failure in many IVF patients. In recent years, fluorescent in situ hybridisation (FISH) for multiple chromosomes has been used to document a high frequency of chromosomal errors and aneuploidy in human preimplantation embryos and, after embryo biopsy, to select embryos that are more likely to implant. Such studies suggest that women with recurrent miscarriage and advanced maternal age may benefit most from preimplantation genetic diagnosis with aneuploidy screening (PGD-AS). The success of PGD-AS is likely to be enhanced by new technologies, such as comparative genomic hybridisation, which enable full karyotyping of single cells. PMID- 12116319 TI - First application of preimplantation genetic diagnosis to neurofibromatosis type 2 (NF2). AB - Neurofibromatosis type 2 (NF2) is a dominantly inherited cancer predisposition syndrome that is caused bymutations in the NF2 gene. We report here the first clinical preimplantation genetic diagnosis (PGD) forNF2. A protocol was developed to simultaneously amplify the mutation and a single nucleotide polymorphism (SNP) located within the gene. The mutation and polymorphism were analysed by simultaneous fluorescent single-strand conformation polymorphism (SSCP) on an automated DNA sequencer. The mutation, carried by the male partner, was a single base pair substitution affecting a splice site in intron 4 of the gene. The female partner was infertile due to polycystic ovary syndrome and would require IVF to conceive. The couple was found to be informative at a linked intragenic SNP situated in the 5' untranslated region of the gene. The SNP was included in the assay to reduce the risk of misdiagnosis due to allele dropout (ADO). The couple underwent three cycles of treatment during which a total of 43 blastomeres were biopsied from 31 embryos. Amplification at both loci was obtained in 35 cells (81%). A total of five embryos were transferred, two in the first cycle, two in the second and one in the third. No pregnancy ensued. The results of the diagnoses indicated that, in this couple, the inheritance of the mutation may be non-Mendelian. Out of a total of 32 embryos tested only four were found not to carry the mutation. The reasons for this apparent skew remain unknown. PMID- 12116321 TI - Current awareness. PMID- 12116320 TI - Preimplantation genetic diagnosis for single gene disorders: experience with five single gene disorders. AB - We report our experience of 14 preimplantation genetic diagnosis (PGD) cycles in eight couples carrying five different single gene disorders, during the last 18 months. Diagnoses were performed for myotonic dystrophy (DM), cystic fibrosis (CF) [Delta F508 and exon 4 (621+1 G>T)], fragile X and CF simultaneously, and two disorders for which PGD had not been previously attempted, namely neurofibromatosis type 2 (NF2) and Crouzon syndrome. Diagnoses for single gene disorders were carried out on ideally two blastomeres biopsied from Day 3 embryos. A highly polymorphic marker was included in each diagnosis to control against contamination. For the dominant disorders, where possible, linked polymorphisms provided an additional means of determining the genotype of the embryo hence reducing the risk of misdiagnosis due to allele dropout (ADO). Multiplex fluorescent polymerase chain reaction (F-PCR) was used in all cases, followed by fragment analysis and/or single-stranded conformation polymorphism (SSCP) for genotyping. Embryo transfer was performed in 13 cycles resulting in one biochemical pregnancy for CF, three normal deliveries (a twin and a singleton) and one early miscarriage for DM and a singleton for Crouzon syndrome. In each case the untransferred embryos were used to confirm the diagnoses performed on the biopsied cells. The results were concordant in all cases. The inclusion of a polymorphic marker allowed the detection of extraneous DNA contamination in two cells from one case. Knowing the genotype of the contaminating DNA allowed its origin to be traced. All five pregnancies were obtained from embryos in which two blastomeres were biopsied for the diagnosis. Our data demonstrate the successful strategy of using multiplex PCR to simultaneously amplify the mutation site and a polymorphic locus, fluorescent PCR technology to achieve greater sensitivity, and two-cell biopsy to increase the efficiency and success of diagnoses. PMID- 12116324 TI - Cavity preparation using a superpulsed 9.6-microm CO2 laser--a histological investigation. AB - BACKGROUND AND OBJECTIVES: The superpulsed 9.6-microm CO(2) laser is an effective laser for ablating dental tissues and decay. This histological study compares laser class V preparations with conventional treatment to evaluate the resulting formation at the cavity walls. STUDY DESIGN/MATERIALS AND METHODS: Four class V preparations (one made with a diamond drill and three with the CO(2) laser (9.6 microm, 60 microseconds pulse width, 40 mJ pulse energy, 100 Hz, integrated scanner system, water cooling) were performed on ten extracted teeth. The cavities were filled with a composite resin partly including enamel and dentine conditioning. RESULTS: After laser preparation, no cracks or signs of carbonisation were detected. The results were comparable to those attained with conventional treatment. Following cavity filling without prior conditioning, gaps were noted at the cavosurface indicating a lack of adhesion. Dentinal bonding decreased gap formation significantly. CONCLUSION: The 9.6-microm CO(2) laser is an effective tool for cavity preparation. PMID- 12116325 TI - Calcium solubility of dental enamel following sub-ablative Er:YAG and Er:YSGG laser irradiation in vitro. AB - BACKGROUND AND OBJECTIVE: The objective of the present study was to investigate the effect of sub-ablative Er:YAG (lambda = 2.94 microm) and Er:YSGG (lambda = 2.79 microm) laser radiation on the acid solubility of dental enamel. The influence of fluoride application prior to laser irradiation was additionally evaluated. STUDY DESIGN/MATERIALS AND METHODS: To this end, 294 enamel specimens were prepared from bovine teeth and divided into 14 groups of 21 specimens each. The enamel samples were irradiated in their groups with the Er:YAG and the Er:YSGG laser, using energy densities of 4, 6, and 8 J/cm(2) in each case. Irradiation was additionally repeated in the same way on specimens, which had previously been immersed in 1% sodium fluoride solution for 15 minutes. One group was left untreated and served as a control group. A further group was not irradiated, but only immersed in the 1% fluoride solution for 15 minutes. The enamel specimens were demineralised for 24 hours in an acetate buffer solution. The calcium content in the demineralisation solution was subsequently determined with the aid of atomic absorption spectrometry. RESULTS: The results indicate a decline in calcium solubility after laser irradiation. Compared to the control group, a 20% lower calcium content was detected in the demineralisation solution after irradiation with the Er:YSGG laser at 8 J/cm(2). The difference between the laser-irradiated groups and the untreated control group was, however, not statistically significant. A significantly lower calcium content was found in the demineralisation solution after fluoridation of the specimens. Additional laser radiation had no further effect on this result. CONCLUSIONS: In summary, it can be stated that, although the erbium laser wavelengths apparently have the potential to increase acid resistance, their application solely for caries prevention would not appear to be sensible under the prevailing conditions. PMID- 12116326 TI - Selective ablation of surface enamel caries with a pulsed Nd:YAG dental laser. AB - BACKGROUND AND OBJECTIVE: High intensity infrared light from the pulsed Nd:YAG dental laser is absorbed by carious enamel and not absorbed by healthy enamel. Consequently, this system has potential for selective removal of surface enamel caries. Safety and efficacy of the clinical procedure was evaluated in two sets of clinical trials at three dental schools. Selective ablation was evaluated with FTIR spectroscopy. STUDY DESIGN/MATERIALS AND METHODS: Carious lesions were randomized to drill or laser treatment. Pulp diagnosis, enamel surface condition, preparations, and restorations were evaluated by blinded evaluators. In Study I, surface caries were removed from 104 third molars scheduled for extraction. One week post-treatment teeth were evaluated clinically, extracted, and the pulp was examined histologically. In Study II, 90 patients with 462 lesions on 374 teeth were randomized to laser or drill and followed for 6 months. RESULTS: Pulsed Nd:YAG laser removal of surface enamel caries was demonstrated to be both safe and effective. Caries were removed in all conditions. There were no adverse events and both clinical and histological evaluations of pulp vitality showed no abnormalities. A significantly greater number of preparations in the drill groups vs. laser groups entered dentin (drill = 11, laser = 1, P = 0.007). CONCLUSION: The more conservative laser treatment removed the caries but not the sound enamel below the lesion. The pulsed Nd:YAG dental laser was found to be both safe and effective for surface caries removal. PMID- 12116327 TI - Comparison of the bond strength of composite resin to Er:YAG laser irradiated human enamel pre-treated with various methods in vitro. AB - BACKGROUND AND OBJECTIVE: We have reported that the application of ultrasonic scalers on the Er:YAG laser-irradiated enamel surface achieved bond strengths of approximately 16 MPa. However, it is reported that a bond strength of 20 MPa is needed to prevent the marginal gaps, so the tensile bond strength (TBS) of composite resin to the Er:YAG laser-irradiated enamel is still insufficient. STUDY DESIGN/MATERIALS AND METHODS: Various methods (phosphoric acid etching, air scaling, ultrasonic scaling, air powder polishing, combination with phosphoric acid etching, and prior mechanical removal methods) were tested as a technique to increase TBS of composite resins to the Er:YAG irradiated enamel. RESULTS: Both the methods of mechanical removal following Er:YAG laser irradiation and the phosphoric acid etching following irradiation and mechanical removal had significant effects on TBS. TBS in the group with acid etching following air powder polishing showed highest value, 22.0 MPa. CONCLUSION: The application of phosphoric acid etching following mechanical removal of Er:YAG laser irradiation was effective to achieve high bond strength. PMID- 12116328 TI - The Er:YAG laser in endodontics: results of an in vitro study. AB - BACKGROUND AND OBJECTIVE: Until recently, the main field of Er:YAG laser application was the removal of dental hard substances within the scope of cavity preparation. Nowadays, several new delivery-systems are available, permitting the application of the Er:YAG laser in endodontics. The aim of the present study was to assess the effects of Er:YAG laser irradiation on root canals in vitro. STUDY DESIGN/MATERIALS AND METHODS: For this purpose, 220 extracted human teeth were endodontically processed and subsequently irradiated at different settings using an Er:YAG laser imitating in vivo irradiation procedures. The teeth were then subdivided into three groups and subjected to bacteriological evaluations, scanning electron microscopy, and temperature measurements. RESULTS: The bacteriological evaluation revealed a decisive bactericidal effect of the Er:YAG laser in the root canal. The bactericidal effect was dependent on the applied output power and specific for the different species of bacteria investigated. Scanning electron microscopy showed discrete removal of dentine from the root canal walls. The temperature rise during irradiation was moderate when standardized power settings were used. CONCLUSION: The investigations indicate that the Er:YAG laser is a suitable tool for the elimination of bacteria in root canals under in vitro conditions. PMID- 12116329 TI - Low level 809-nm diode laser-induced in vitro stimulation of the proliferation of human gingival fibroblasts. AB - BACKGROUND AND OBJECTIVE: The authors investigated the effects of low level laser irradiation on the proliferation rate of human gingival fibroblasts (HGF) in vitro. STUDY DESIGN/MATERIALS AND METHODS: HGF were obtained from gingival connective tissue explants and cultured under standard conditions. 110 cell cultures in their logarithmic growth phase were spread on 96-well tissue culture plates and were irradiated at energy fluences of 1.96-7.84 J/cm(2). Another 110 cultures served as control. An 809-nm semiconductor laser operated at a power output of 10 mW in the cw-mode was used. The time of exposure varied between 75 and 300 seconds. Laser treatment was performed alternatively once, twice, and three times at a 24-hour interval. After lasing, incubation was continued for 24 hours. The proliferation rate was determined by means of fluorescence activity of a redox indicator added to the cell culture. Proliferation was determined 24, 48, and 72 hours after irradiation and expressed in relative fluorescence units (RFU). RESULTS: The irradiated cells revealed a considerably higher proliferation activity. The differences were highly significant 24 hour after irradiation (Mann Whitney U-test, P < 0.05) but decreased in an energy-dependent manner after 48 and 72 hour after irradiation. CONCLUSIONS: A cellular effect of the soft laser irradiation on HGF is evident. Its duration, however, seems to be limited. These findings might be clinically relevant, indicating that repeated treatments are necessary to achieve a positive laser effect in clinical applications. PMID- 12116330 TI - Scanning laser system for photodynamic therapy of choroidal neovascularization. AB - BACKGROUND AND OBJECTIVES: In order to improve selectivity of photodynamic therapy (PDT) to choroidal neovascularization (CNV) associated with age-related macular degeneration, a laser scanning technique was applied to perform focal laser irradiation to the retina, and the occlusion effects of a new device to the choriocapillaris were evaluated in primate eyes. STUDY DESIGN/MATERIALS AND METHODS: The device contains lasers for fundus observation of 785 nm and for PDT of 670 nm, matching the absorption peak of a photosensitizer, ATX-S10(Na). The laser irradiated the shape on the retina specified before treatment and shut off automatically when the predetermined treatment was achieved. The occlusion of the choriocapillaris after PDT was documented by fluorescein and indocyanine green angiography and histology. RESULTS: The area designated for PDT was easily drawn on the touch-screen monitor, and occlusion of the choriocapillaris was achieved precisely in the area pre-selected for treatment with 5 J/cm(2) or more of radiance following administration of 8 mg/kg ATX-S10(Na). CONCLUSIONS: This device is useful for irradiating CNV of any shape, sparing the surrounding retina. Since our previous studies suggested that selective occlusion of CNV would decrease not only the functional disturbance caused by PDT, but also the recurrence of CNV, the present device may allow more effective PDT than the slit lamp system presently used. PMID- 12116331 TI - Acute electrophysiologic effect of pulsed gallium-arsenide low energy laser irradiation on configuration of compound nerve action potential and nerve excitability. AB - BACKGROUND AND OBJECTIVES: We evaluated the acute electrophysiologic effects of low-energy pulsed laser irradiation, measured by extracellular recording technique on compound action potential configuration and nerve excitability in the isolated frog sciatic nerve STUDY DESIGN/MATERIALS AND METHODS: A pulsed gallium-arsenide (GaAs) laser (wavelength, 904 nm; pulse duration, 220 nanoseconds; peak power per pulse, 27 W; spot size, 0.28 cm(2); total applied energy density, 0.005-2.5 J/cm(2)) was used for the experiment. Sixty isolated nerves were divided into six groups (n = 10), each of which received a different repetition frequency. In each group, action potentials were recorded, before laser irradiation, which served as the control data. The extracellular action potentials were recorded for each combination of 1, 3, 5, 7, 10, 13, and 15 minutes of irradiation time and 4, 8, 16, 32, 64, 128 repetition frequency by using a BIOPAC MP 100 Acquisition System Version 3.5.7 (Santa Barbara USA). Action potential latency, duration of depolarization and repolarization, and the stimulating voltage were measured. Statistical evaluation was performed using linear correlation analysis by SPSS 9.05. RESULTS: Although there was no correlation between applied energy density and action potential latency, the duration of depolarization and repolarization phases (P > 0.05), there was a weak correlation between applied energy density and stimulating voltage. CONCLUSIONS: The study showed that low-energy GaAs irradiation at 42 different energy density between 0.005 and 2.5 J/cm(2) generates no effect on action potential configuration and nerve excitability. PMID- 12116332 TI - Scatter-limited phototherapy: a model for laser treatment of skin. AB - BACKGROUND AND OBJECTIVES: To effectively deliver laser light into the skin for non-ablative resurfacing, hair removal, and other applications, one must account for scatter, absorption, and thermal diffusion. A novel method to control the penetration of laser light into tissue is proposed. This method uses the incident beam diameter, the pulse duration, and the intrinsic light scattering of tissues such as skin to limit the laser light penetration and thermal damage. We term this innovative laser delivery concept 'scatter-limited phototherapy'. STUDY DESIGN/MATERIALS AND METHODS: This study demonstrates how the delivery mechanism simplifies the non-ablative treatment of sun-damaged skin. Calculations are based upon Gaussian scatter of light and isotropic thermal diffusion. Calculations of light distribution and profiles of temperature increase are created. RESULTS: Using an optical fiber delivery system with the appropriate diameter, coupled with skin surface protection, one can create thermal damage at a given depth with preservation of the epidermis. CONCLUSIONS: Scatter-limited phototherapy is a predictive model, allowing one to design better laser delivery systems. Scatter limited phototherapy should also be applicable to other fields of dermatologic surgery, such as hair removal and the treatment of vascular lesions. Additionally, other medical specialties will be able to use the concept of scatter-limited phototherapy to predict and better understand laser-tissue interactions. PMID- 12116333 TI - Laser-tattoo removal--a study of the mechanism and the optimal treatment strategy via computer simulations. AB - BACKGROUND AND OBJECTIVE: The physical mechanisms for laser-tattoo interactions and the tattoo particle breakup process are not well understood. This study investigates whether the mechanism of the breakup process can be identified via computer simulations and proposes a treatment strategy that can potentially minimize the collateral damage to the surrounding tissues. Note that the "removal" of tattoo particles is defined here as breakup of particles into smaller ones with sizes approaching or smaller than the visible wavelength of light so that they become less visible. STUDY DESIGN/MATERIALS AND METHODS: The radiation-hydrodynamics code LATIS is used for the modeling. We first identify the magnitude of the tensile stress generated inside graphite tattoo particles as functions of laser pulse length and particle size. We then calculate the relationship between the surface laser fluence (defined as the time integrated energy flux) and the tensile strength of the tattoo particle at a given depth. RESULTS: If the laser pulse length is sufficiently short, strong acoustic waves with tensile strengths exceeding the fracture thresholds for graphite are generated. The strength of the wave decreases with particle size and increases as the laser pulse length decreases. Simulation results are in general agreement with clinical studies. Although temperatures of the tattoo particles never reach the melting point, a cavitation bubble around the particle can be formed. The steam generated can get into the cracked particles and induce steam-carbon reactions. Laser energy density decreases rapidly with the skin depth. Therefore, the minimum surface laser fluence, for a given pulse length, required for breaking up tattoo particles at a given skin depth, increases with particle depth. CONCLUSIONS: Computer simulations confirm that the breakup of tattoo particles is photoacoustic. For the same amount of laser energy, a shorter pulse is more efficient. The optimal pulse length is approximately 10-100 picosecond to minimize the laser fluence and the collateral damage. It is more difficult to break up the smallest tattoo particles that have diameters smaller than 10 nm; however, smaller particles are less important because they are less visible. Tissue surrounding the tattoo particles can be damaged by cavitation bubbles. These bubbles could be the cause of the empty vacuoles in the ash-white lesions throughout the dermis seen after treatment. Steam-carbon reactions can be induced. Particles then become grossly transparent because of this reaction. Different laser intensity should be used for pigments at different depths in order to minimize the collateral damage to the dermis. PMID- 12116334 TI - Scar sarcoidosis--treatment with the Q-switched ruby laser. AB - BACKGROUND AND OBJECTIVE: Scar sarcoidosis is a circumscribed form of cutaneous sarcoidosis, which is often very difficult to treat. To date, therapeutic approaches have yielded little success and often been accompanied by adverse effects, some of which are severe. Laser treatment is one alternative, which has proven to be effective in a dramatically increasing number of cases over the past years. STUDY DESIGN/MATERIALS AND METHODS: We report about a 50-year-old female patient who presented with histologically confirmed scar sarcoidosis on the right elbow and both knees; the sarcoidosis had spontaneously developed on three sites which had traumatic tattoos from abrasions. Because of the reddish-brown livid discolorations, we treated the granulomas with the pulsed dye laser for three sessions, although without success. Treatment with the Q-switched ruby laser was commenced to remove the traumatic tattoos. RESULTS: Not only had the areas lightened fully after four sessions, the sarcoidosis foci had resolved completely, and the patient has been recurrence-free for over 3 years. CONCLUSION: The Q-switched ruby laser appears to be a rapid and effective means of treating scar sarcoidosis with traumatic tattoos without incurring adverse effects. The definitive mode of action is still not fully known, however. PMID- 12116335 TI - Long-pulsed ruby laser for permanent hair reduction: histological analysis after 3, 4 1/2, and 6 months. AB - BACKGROUND AND OBJECTIVES: The histology of hair follicles in both animal and human skin treated with ruby lasers has been evaluated to a limited extent in previous studies. We have previously looked at such follicles up to 2 months after treatment. This study examines the longer-term effects at a microscopic level and attempts to further elucidate the mechanism of ruby laser hair reduction. STUDY DESIGN/MATERIALS AND METHODS: Thirty-six patients underwent 1, 2, or 3 treatments of their axillary or bikini area skin with a 3 milliseconds ruby laser at 10, 20, 30, or 40 J/cm(2). Biopsies were taken 3, 4(1/2), or 6 months after the last treatment and examined histologically. Nine control biopsies were taken from comparable bikini areas of untreated patients and similarly evaluated histologically. RESULTS: There was a significant increase in telogen compared to anagen follicles in treated skin, which was slightly increased by multiple compared to single treatments, but unaffected by different time intervals since the last treatment. There was also a significant increase in miniaturized compared to terminal hairs in treated compared to control skin, a finding that was further increased with higher energies used. Multiple treatments and time after treatment had a slight, but not statistically significant effect on follicle size. CONCLUSIONS: Induction of telogen in terminal follicles followed by miniaturization appears to be the main mechanism of ruby laser hair reduction. PMID- 12116336 TI - Clinical applications of laser scanning cytometry. AB - This study reviews existing and potential clinical applications of laser scanning cytometry (LSC) and outlines possible future developments. LSC provides a technology for solid phase cytometry. Fluorochrome-labeled specimens are immobilized on microscopic slides that are placed on a conventional epifluorescence microscope and analyzed by one or two lasers. Data comparable to flow cytometry are generated. In addition, the position of each event is recorded, a feature that allows relocalization and visualization of each measured event. The major advantage of LSC compared with other cytometric methods is the combination of two features: (a) the minimal clinical sample volume needed and (b) the connection of fluorescence data and morphological information for the measured event. Since the introduction of LSC, numerous methods have been established for the analysis of cells, cellular compartments, and tissues. Although most cytometric methods use only two or three colors, the characterization of specimens with up to five fluorochromes is possible. Most clinical applications have been designed to determine ploidy and immunophenotype; other applications include analyses of tissue biopsies and sections, fluorescence in situ hybridization, and the combination of vital and nonvital information on a single-cell basis. With the currently available assays, LSC has proven its wide spectrum of clinical applicability in slide-based cytometry and can be introduced as a standard technology in multiple clinical settings. PMID- 12116337 TI - Double-fluorescence image microscopy for quantitation of prostate-specific antigen in histologic sections of the prostate. AB - BACKGROUND: Although the assessment of serum prostate-specific antigen (PSA) has become a powerful instrument in the diagnosis and for prognosis of prostate carcinoma, there are few quantitative studies of PSA in tissue sections. METHODS: We developed a technique using double-fluorescence image microscopy for quantifying immunohistochemical reactions in tissue sections. PSA was stained by Texas Red and the cellular DNA was counterstained with 4,6-diamidino-2 phenylindole, dihydrochloride (DAPI). The fluorescence of Texas Red and DAPI was quantified separately after subtraction of background and shading correction. The amount of PSA related to the amount of DNA in identical tissue parts was studied in archival specimens from patients with hyperplasia and prostate carcinoma. RESULTS: The amount of tissue PSA decreased with the increase in tumor grade, Gleason score, and the change from diploid to aneuploid. CONCLUSION: Double fluorescence image microscopy is a valuable technique for obtaining quantitative information of cellular constituents. For standardization of immunochemical reactions in tissue sections, cellular DNA seems to be most appropriate. PMID- 12116338 TI - Incidence of numerical chromosome aberrations in meningioma tumors as revealed by fluorescence in situ hybridization using 10 chromosome-specific probes. AB - OBJECTIVE: Although information on the cytogenetic characteristics of meningioma tumors has accumulated progressively over the past few decades, information on the genetic heterogeneity of meningiomas is still scanty. The aim of the present study was to analyze by interphase fluorescence in situ hybridization (FISH) the incidence of numerical abnormalities for chromosomes 1, 9, 10, 11, 14, 15, 17, 22, X, and Y in a group of 70 consecutive meningioma tumors. Another goal was to establish the potential associations among the altered chromosomes, as a way to assess both intertumoral and intratumoral heterogeneity. METHODS: For the purpose of the study, 70 patients diagnosed with meningioma were analyzed. Interphase FISH for the detection of numerical abnormalities for chromosomes 1, 9, 10, 11, 14, 15, 17, 22, X, and Y was applied to fresh tumor samples from each of the patients studied. RESULTS: The overall incidence of numerical abnormalities was 76%. Chromosome Y in males and chromosome 22 in the whole series were the most common abnormalities (46% and 61%, respectively). Despite the finding that monosomy of chromosome 22/22q(-) deletions are the most frequent individual abnormality (53%), we have observed that chromosome gains are significantly more common than chromosome losses (60% versus 40%). Chromosome gains corresponded to abnormalities of chromosomes 1 (27%), 9 (25%), 10 (23%), 11 (22%), 14 (33%), 15 (22%), 17 (23%), and X in females (35%) and males (23%) whereas chromosome losses apart from chromosome 22 frequently involved chromosomes 14 (19%), X in males (23%), and Y in males (32%). Although an association was found among most gained chromosomes on one side and chromosome losses on the other side, different association patterns were observed. Furthermore, in the latter group, monosomy 22/22q(-) was associated with monosomy X in females and monosomy 14/14q(-) was associated with nulisomy Y in males. In addition, chromosome losses usually involved a large proportion of the tumor cells whereas chromosome gains were restricted to small tumor cell clones, including tetraploid cells. CONCLUSIONS: Our results show that meningiomas are genetically heterogeneous tumors that display different patterns of numerical chromosome changes, as assessed by interphase FISH. PMID- 12116339 TI - Monitoring tumour cells in the peripheral blood of small cell lung cancer patients. AB - BACKGROUND: Flow cytometry was used to enumerate tumour cells in longitudinal studies of peripheral blood from small cell lung cancer (SCLC) patients, together with magnetic bead selection to isolate and identify these cells. As part of a trial, 11 patients received either standard (four weekly) chemotherapy with ifosfamide, carboplatin, and etoposide (ICE) or accelerated (two weekly) ICE with filgrastim (granulocyte colony-stimulating factor [G-CSF]) and autologous stem cell support. METHODS: Fresh venous blood was taken throughout treatment and follow-up. Aliquots were stained with a "tumour-specific" antibody against epithelial tissue (Ber EP4), verified as a good marker of SCLC cells by immunohistochemistry. Matched samples labelled with Ber EP4 were separated magnetically by adding a secondary bead-antibody conjugate for confirmation of tumour cell identity. RESULTS: Circulating tumour cells were detected and monitored throughout treatment periods. An initial rise in circulating cells after the first cycle was followed by a fall in both treatment arms to baseline levels set by normal controls. This was achieved by week 12 in the accelerated treatment arm and by week 24 in the standard arm. CONCLUSIONS: Flow cytometry and magnetic bead isolation can be used to identify changes in numbers of circulating tumour cells in patients undergoing chemotherapy for SCLC and thereafter during follow-up periods. Absence of tumour cells may indicate a more favourable patient group who would benefit from a more intense course of treatment. PMID- 12116340 TI - Increase of sensitivity of sputum cytology using high-resolution image cytometry: field study results. AB - Lung cancer remains the leading cause of cancer deaths in the developed world. There is no widely accepted method to screen for this cancer. The most commonly used method remains conventional sputum cytology, but this method is hampered by low sensitivity. We tested the hypothesis that sensitivity of sputum cytology for early lung cancer can be greatly improved by using image analysis of sputum cells, at a modest reduction of specificity. The study was double-blinded and used sputum samples from subjects with well-characterized clinical diagnoses. There were 177 cancers, 98 dysplasias, and 558 normals. The study samples were separated into two independent sets: training set and test set. Sputum samples were collected prospectively from subjects with a high probability of having lung cancer. Seven institutions from five countries participated in the study. All subjects had complete clinical diagnoses which included, as a minimum, negative chest x-rays for all negative cancers, while all cancers had confirmed tissue pathology. Samples were prepared according to the Saccomanno method. For conventional cytology, slides were stained using Papanicolaou stain. For image analysis, slides were stained using a DNA-specific (Feulgen-Thionin) stain. An automated, high-resolution image cytometer was used for measurements. At 90% specificity, sensitivity of 60% can be achieved for adenocarcinoma, compared to only 14% sensitivity of conventional cytology (at 99% specificity). Similarly, 45% sensitivity at 90% specificity can be reached for stages 0 and I lung cancer, compared to only 14% (at 99% specificity) using conventional cytology.Cytometry combined with conventional cytology shows an increase in sensitivity to early stage cancer and to adenocarcinomas compared to conventional cytology alone. While the results are encouraging, the sensitivity to detect early lung cancer should be further improved to 70-80% at 90-95% specificity before this test can be considered for screening of high-risk individuals for lung cancer. Cytometry (Clin. Cytometry) 50:168-176, 2002. PMID- 12116341 TI - An approach to diagnosis of T-cell lymphoproliferative disorders by flow cytometry. AB - T-cell lymphoproliferative disorders are among the most challenging diagnoses in hematopathology. Unlike the more common B-cell disorders, in which clonality is often readily discernible by surface immunoglobulin light chain restriction, there is no specific immunophenotypic signature that is diagnostic of a clonal T cell population. Immunophenotypic criteria that are helpful in the diagnosis of T cell neoplasms include T-cell subset antigen restriction, anomalous T-cell subset antigen expression, deletion or diminution of one of the pan T-cell antigens, a precursor T-cell phenotype, and expression of additional markers (e.g., CD30, CD20, major myeloid antigens, and TCRgammadelta). Analysis of the inherent forward and orthogonal light scatter properties of the cell can also provide important diagnostic clues. None of these features is 100% specific, however, for aberrant expression of pan-T antigens may be seen in viral infections, B-cell malignancies, or in reactive changes following administration of certain medications. An increased CD4:CD8 ratio is often observed in Hodgkin's lymphoma. Based on the analysis of 87 neoplastic and 80 control cases, we conclude that flow cytometric features that are most suspicious for malignancy include the loss or markedly dim expression of CD45; complete loss of one or more pan-T antigens; diminished expression of more than two pan-T antigens in conjunction with altered light scatter properties; and CD4/CD8 dual-positive or dual-negative expression (except thymic lesions). PMID- 12116342 TI - Comparative analysis of different flow cytometry-based immunophenotypic methods for the analysis of CD59 and CD55 expression on major peripheral blood cell subsets. AB - BACKGROUND: Flow cytometry-based immunophenotypic techniques for the analysis of CD55 and CD59 expression on the major cell populations present in blood are the preferred method for the diagnostic screening of paroxysmal nocturnal hemoglobinuria (PNH). METHODS: In the present study, we comparatively analyze the effects of stain-lyse-and-then-wash techniques and lyse-wash-and-then-stain procedures on the detection of both CD55 and CD59 expression on the major peripheral blood (PB) leucocyte subsets, as analyzed by flow cytometry. Our major goal was to establish the minimum amounts of anti-CD55 and anti-CD59 reagents required to be added to a minimum volume of blood, which would allow an optimal staining for both antigens on red cells, platelets, and leucocytes present in a single tube. RESULTS: Our results show that upon comparing stain-lyse-and-then wash techniques with lyse-wash-and-then-stain protocols, the presence of important amounts of red cells at the time peripheral blood leucocytes are stained for CD55 and CD59 is associated with a significantly (P < 0.01) lower and more heterogeneous pattern of antigen expression on almost all major PB leucocyte subsets, supporting the need to use red cell lysing procedures prior to the staining of leucocytes. Identical, optimal patterns of antigen staining for CD55 and CD59 were obtained upon incubating 3 microL of blood with 10 microL of each of these monoclonal antibody (mAb) reagents (protein concentration of 0.05 microg/microL and 0.2 microg/microL respectively) for 30 min (room temperature [RT]) using a non-lyse-non-wash sample preparation procedure. This latter procedure allowed for the simultaneous analysis of CD55 and CD59 expression on red cells, platelets, neutrophils, monocytes, and lymphocytes present in the sample through the combined staining of CD55 and CD59 with CD64-fluorescein isothiocyante (FITC) plus CD61-peridinin chlorophyll protein (PerCP) and CD45 PerCP. CONCLUSIONS: In summary, our results show that the sample preparation protocol has a significant impact on the quality of the staining obtained for the CD55 and CD59 antigens on the major PB leucocyte subsets; additionally, we propose a simple and reliable stain-non-lyse-non-wash method for the simultaneous analysis of CD55 and CD59 expression on PB red cells, platelets, neutrophils, monocytes, and lymphocytes, which could be reached through the use of two triple stainings. PMID- 12116344 TI - T-cell subset counting and the fight against AIDS: reflections over a 20-year struggle. AB - The story of T-lymphocyte subset immunophenotyping technology is reviewed on the occasion of the 20th anniversary of CD4 T-cell enumeration. Over time, immunophenotyping has evolved into precise, reliable, but complicated and expensive technology requiring fresh blood samples. The gating technologies that were universally adapted for clinical flow cytometry for the past decade relied on rapidly deteriorating morphological scatter characteristics of leukocytes. This special issue dedicated to CD4 T-cell enumeration features most of the available new options that will have a significant impact on how this technology will be implemented within the first decade of the 21st century. In a series of original publications, including the new NIH guideline for T-cell subset enumeration, contemporary gating protocols that use immunologically logical parameters are presented as part of the more reliable and affordable immunophenotyping alternative. Some of the improvements addressed here include the costs of the assays and the capacity to monitor interlaboratory and intralaboratory performances. It is clear that an effective attack on the human immunodeficiency virus (HIV) epidemic has to embrace resource-poor regions. Reducing the cost of the assay while improving reliability and durability is a move in the right direction. PMID- 12116345 TI - Use of CD45 gating in three and four-color flow cytometric immunophenotyping: guideline from the National Institute of Allergy and Infectious Diseases, Division of AIDS. PMID- 12116346 TI - Selection of lymphocyte gating protocol has an impact on the level of reliability of T-cell subsets in aging specimens. AB - BACKGROUND: In the past decade, human immunodeficiency virus (HIV) lymphocyte immunophenotyping has evolved significantly. New fluorochromes, new multicolor reagents, enhanced instruments, and the capacity to provide absolute cell counts using the single-platform technique have all contributed to the reliability of T cell subset measurements. In this study, four gating protocols were evaluated to select the most robust method for T-cell subset enumeration. METHODS: Peripheral blood specimens from 21 HIV(+) and 20 HIV(-) individuals were monitored up to 96 h. Aliquots of specimens were stored at room temperature and analyzed at 6 (baseline), 48, 72, and 96 h. Aliquots were stained with CD45-fluorescein isothiocyanate (FITC)/CD3PC5/CD4RD1/CD8ECD. Data analysis was performed with all four gating protocols. RESULTS: Only with fresh blood did all protocols provide similar results. From samples that were 48 h old, the choice of gating strategy had a dramatic impact on immunophenotyping results. The largest deviations from baseline values occurred at 96 h and gating protocols that included dual light scatter gates provided the greatest shift of T-cell subset values over time. The gating protocols that were based exclusively on cell lineage-specific gates gave the most robust T-cell values up to 96 h. CONCLUSION: By selecting the appropriate gating protocol, the temporal integrity of specimens can be extended up to 4 days. PMID- 12116347 TI - Evaluation of a universal template for single-platform absolute T-lymphocyte subset enumeration. AB - The single-platform absolute T-lymphocyte subset analysis was evaluated utilizing a universal protocol in a Canadian multicenter study with the collaboration of the members of the Canadian HIV Trials Network (CTN). Participants used flow cytometers and reagents of their choice for labeling and lysing whole blood. Over a 2-year period, CTN laboratories performed single-platform absolute T-lymphocyte subset enumerations on fresh and commercial stabilized blood products using commercially available microfluorospheres TruCount and Flow-Count. This multicenter evaluation demonstrated that the application of a universal template for single-platform analysis provides a generic approach that embraces a wide array of immunophenotyping settings available in clinical laboratory. PMID- 12116348 TI - CD45-assisted PanLeucogating for accurate, cost-effective dual-platform CD4+ T cell enumeration. AB - BACKGROUND: North American and European guidelines for dual-platform (DP) flow cytometry recommend absolute CD4 T-cell counts to be calculated from two parameters: the absolute lymphocyte counts obtained on a hematology analyzer and the percentages of CD4+ cells among lymphocytes (CD4%/lympho) obtained by flow cytometry. Nevertheless, the identification of lymphocytes is error-prone: a poor match between these common denominators in the two systems is the main source of inaccuracy. In contrast, total leucocyte counts (white cell counts [WCC]) and CD4% among the gated CD45+ leucocytes (CD4%/leuco) can be determined with greater accuracy. METHODS: We introduced "PanLeucogating," i.e., we used total leucocytes as the common denominator for improving the precision of DP absolute CD4 counting. Correlations and Bland-Altman tests were used for statistical analysis. RESULTS: First, 22 stabilized blood product samples were provided by U.K. National External Quality Assessment Scheme (NEQAS) and a higher accuracy and precision of CD4 counts were documented using PanLeucogating compared with lymphocyte gating. Next, 183 fresh and 112 fixed (TransFix) whole blood samples were used to compare DP methods and single-platform (SP) methodology, including both volumetric and bead-based techniques. A particularly high correlation and comparable precision of absolute CD4 counts were observed between the SP volumetric method and DP PanLeucogating (R(2) = 0.990; bias 6 +/- SD 17%). The SP volumetric method showed lower levels of agreement with the DP lymphocyte gating (R(2) = 0.758; bias 14 +/- SD 51%) and with the SP bead-based method (R(2) = 0.923; bias 4 +/-SD 31%). CONCLUSIONS: These observations show that DP leucocyte counts (WCC) should replace lymphocyte counts as the "common denominator" although CD4%/lympho values can, as an extra step, be also provided readily if requested. When coupled with quality control for WCC on hematology analyzers, the DP method with CD45 PanLeucogating represents a robust CD4 T-cell assay that is as accurate as the SP volumetric technique. This DP method uses only two, CD45 and CD4, antibody reagents and can be run on any pair of hematological analyzer plus flow cytometer. PMID- 12116349 TI - Precise CD4 T-cell counting using red diode laser excitation: for richer, for poorer. AB - BACKGROUND: Measuring CD4 T-cell counts at low cost is relevant in dealing with the human immunodeficiency virus (HIV) epidemic throughout the developing world. The recently introduced novel concepts in gating strategies and sample stabilization facilitate affordable immunophenotyping by flow cytometry. However, the impact of these developments is still limited by the high cost of currently available flow cytometers. METHODS: Diode lasers emitting 10-15 mW at 635 nm are one-tenth the size and cost and require one thousandth the power of an equivalent 488-nm argon ion laser. We used the available 635-nm diode-based flow cytometers, including PA-II, Luminex 100, SuperMot, and FACSCalibur, to investigate whether these instruments can generate reliable CD4 counts when used with allophycocyanin (APC) and cyanin-5 (Cy5)-labeled CD4 antibodies. RESULTS: We document the feasibility of obtaining leucocyte differential counts using orthogonal side scatter (SSC) without the need for forward scatter (FSC). Accurate CD4% values among lymphocytes and leucocytes can be obtained by primary CD4 gating using a single CD4 monoclonal antibody conjugated to APC or Cy5. Double immunofluorescence (IF) staining with CD4-APC (FL1) and CD45-APC-Cy7 (FL2) introduces pan-leucogating for a convenient assessment of absolute CD4 counts on double platforms. We demonstrate that small flow cytometers with laser diodes are capable of delivering absolute CD4 T-cell counts with a precision similar to the performance of the current state-of-the-art single-platform instruments (e.g., the CytoronAbsolute; R(2) = 0.961). In this respect, they appear to be superior to the nonflow CD4 counting techniques. CONCLUSIONS: Accurate CD4 counts can be generated at minimal cost on red diode laser-operated flow cytometers, retaining the potential for high throughput capacity without compromising precision. With further improvements in volumetric technology and clinical software, these cytometers may develop into a new generation of inexpensive battery-operated laboratory hardware that combines cellular phenotyping with bead-based multiplexing immunoassays for (HIV) serology. PMID- 12116350 TI - Evaluation of stabilized blood cell products as candidate preparations for quality assessment programs for CD4 T-cell counting. AB - BACKGROUND: Exceptionally robust cell preparations are needed for quality assessment programs (QAPs) such as the International Program for Quality Assessment and Standardization for Immunological Measures (QASI) relevant to HIV/AIDS. A suitable product must withstand environmental stress related to transportation for a minimum of 6 days. The two objectives of this study are (1) to evaluate the performance of various commercial preparations with multicenter participation and (2) to evaluate the robustness of stabilized blood cell products. METHODS: Phase 1: The performance of stabilized blood cell products was evaluated in a multicenter QAP utilizing various staining procedures and flow cytometers. Absolute cell enumeration was achieved using single-platform T-cell subset methodology. Phase 2: The robustness of stabilized blood cell products was evaluated by monitoring T-cell subset values from samples stored at 4 degrees C, 22 degrees C, and 37 degrees C for up to 10 days. RESULTS: The largest interlaboratory variation in both absolute and relative T-cell values was 16% in samples with CD4 levels > or =400 cells per microliter and 21% in samples with CD4 levels <400 cells per microliter. Six preparations retained their phenotypic expression for 7 days at 4 degrees C and 22 degrees C. However, only two preparations remained stable for 4 days at 37 degrees C. CONCLUSION: Some stabilized cell preparations are more robust and therefore more suitable for quality assessment purposes. PMID- 12116351 TI - Reduction of variation in T-cell subset enumeration among 55 laboratories using single-platform, three or four-color flow cytometry based on CD45 and SSC-based gating of lymphocytes. AB - BACKGROUND: Enumeration of CD4(+) and CD8(+) T-cell subsets provides relevant information for diagnosis and monitoring of patients with cellular immunodeficiencies. As a result, an external quality assurance scheme was implemented in Belgium, The Netherlands, and Luxembourg in 1995. A workshop was held to train the participants in state-of-the art technology for assessment of absolute T-cell subset counts (i.e., a three or four-color, single-platform assay with lymphocyte gating based on CD45 and sideward light scatter) with the aim to achieve between-site coefficients of variation (CVs) <10% and within-site CVs <5% for > or =75% of the participants. METHODS: Three send-outs of stabilized blood from a healthy donor were distributed to 55 laboratories, each with the request to perform the standard assay on three occasions. For comparison, each laboratory performed its local technique in parallel. RESULTS: With the standard technique, between-site CVs of approximately 8% (CD3+ T cells), approximately 9% (CD4+ T cells), and approximately 10% (CD8+ T cells) were achieved. Within-site CVs were <5% for 82% (CD3+ T cells) and approximately 70% (CD4+ and CD8+ subsets) of the participants. Local techniques yielded between-site CVs of 13%-17% for CD3+, CD4+, and CD8+ T cells. CONCLUSIONS: The state-of-the-art technology for T-cell subset enumeration was implemented successfully among 55 Belgian-Dutch laboratories and resulted in significant reductions of between-site variation of absolute CD3+, CD4+, and CD8+ T-cell counts. PMID- 12116352 TI - Quality control of CD4+ T-lymphocyte enumeration: results from the last 9 years of the United Kingdom National External Quality Assessment Scheme for Immune Monitoring (1993-2001). AB - The human immunodeficiency virus (HIV) global epidemic has necessitated the routine enumeration of T-lymphocyte subsets, which has created a need for external quality assurance (EQA). The United Kingdom National External Quality Assessment Scheme (UK NEQAS) for Immune Monitoring provides EQA for 296 laboratories in 40 countries. In 1993, UK NEQAS developed and incorporated into its program stabilized whole blood that enables the accurate monitoring of laboratory performance. Overall, the mean interlaboratory coefficient of variation (CV) for percentage CD4(+) T-lymphocyte subset enumeration has fallen from 15% to less than 5%, as a direct result of the increased use of CD45/ side scatter (SSC) gating. Laboratories using alternative gating strategies (i.e., CD45/CD14 or forward scatter [FSC]/SSC) were about 7.4 times more likely to fail an EQA exercise. Furthermore, the adoption of single-platform technology resulted in a reduction of the overall mean interlaboratory CV for absolute CD4(+) T lymphocytes from 56% (prior to the widespread use of single-platform technology) to 9.7%. Individual laboratory deficiencies were also identified using a performance monitoring system and, through re-education by collaboration with the coordinating center, satisfactorily resolved. In conclusion, during the last 9 years, the UK NEQAS for Immune Monitoring program has highlighted the significant technological advances made by laboratories worldwide that undertake lymphocyte subset enumeration. PMID- 12116353 TI - Impact of the international program for Quality Assessment and Standardization for Immunological Measures Relevant to HIV/AIDS: QASI. AB - Measurements of CD4 T-cell levels are essential for the assessment of human immunodeficiency virus (HIV) disease course, clinical staging, epidemiological studies, and decisions regarding prophylactic therapies against opportunistic infection. Until now, only in the industrialized countries was T-cell subset monitoring considered a practical option to assess disease progression. The Quality Assessment and Standardization for Immunological Measures Relevant to HIV/AIDS (QASI) program was established in 1997 to meet performance assessment for immunophenotyping laboratories in countries where such service is not available. The QASI program is provided at no cost to any laboratory in a resource-poor setting that wishes to participate. This report describes the beneficial impact of participation in the QASI program. Carefully selected commercial stabilized whole blood preparations were sent regularly to participating laboratories. Participants reported the T-cell subset values they obtained by flow cytometry. Once the aggregate mean values for the T-cell subsets were established for the shipment, a comprehensive and confidential report was sent to each laboratory. The results from five consecutive shipments were analyzed. The coefficient of variation decreased from 7.2% to 4.7% and from 14.2% to 8.8% for percent and absolute CD4 T-cell counts, respectively. With the implementation of the QASI program using commercial stabilized whole blood specimens, it is possible to reduce interlaboratory error. This study illustrates that a quality assessment program can improve the overall performance of laboratories. Reducing interlaboratory variation can enhance significantly the effectiveness of multicenter HIV vaccine or drug trial evaluation. PMID- 12116354 TI - Grading of laboratories on CD4+ T-lymphocyte evaluations based on acceptable data boundaries defined by the measurement error. AB - BACKGROUND: We addressed the definition of limits of error of %CD4+ and CD4+ counts (AbsCD4+) typical of laboratories of excellence, as well as the grading of laboratories based on the decision to take these limits as boundaries of unacceptable data. METHODS: We studied the 99.9% confidence intervals of the means of 24 human immunodeficiency virus (HIV)+ and HIV- blood samples analyzed by 18 laboratories of the Liguria Region Quality Assessment Program (Liguria Region QALI). Regression equations of lower (L1) and upper (L2) confidence limits over the means of data cleared of unusual results were used to interpolate limits of error for mean values in the tested range. RESULTS: L1 and L2 were symmetric around the mean and a single absolute difference (Abs Res) between the limits and the mean was found. Abs Res significantly increased over mean values (P = 0.0005 for %CD4+, P < 0.0001 for AbsCD4+). Limits were compatible with errors shown with blind replicates. Unacceptable results, outside the limits, accounted for 25% and 30% of %CD4+ and for 18% and 35% AbsCD4+ in the Liguria Region QALI and in the Piemonte Region QA Program, respectively. Limits interpolated over the median showed a similar grading. A comparable fraction of unacceptable data was also found with the method used in the U.K. National External Quality Assessment Scheme (NEQAS) immune monitoring scheme. CONCLUSIONS: We propose the general use of these regression equations to determine bounds for unacceptable data in proficiency testing and to identify laboratories of excellence. PMID- 12116355 TI - Beckman Coulter and CD4+ T cells. PMID- 12116356 TI - BD Biosciences contributions in CD4 counting and immune status for HIV/AIDS. AB - BD Biosciences is a leader in the use of flow cytometry for determining immune system status and for counting CD4 cells in patients with human immunodeficiency virus (HIV) infection. The company has gained this position through many years of basic research and product development in immunology and cell biology, dye chemistry, immunoassays, instrumentation, and software. Some of the highlights of these developments and their historical perspective are described in this review. PMID- 12116357 TI - Near-infrared dyes for six-color immunophenotyping by laser scanning cytometry. AB - BACKGROUND: To adequately analyze the complexity of the immune system and reduce the required sample volume for immunophenotyping in general, more measurable colors for the discrimination of leukocyte subsets are necessary. Immunophenotyping by the laser scanning cytometer (LSC), a slide-based cytometric technology, combines cell detection based on multiple colors with their subsequent visualization without the need for physical cell sorting. In the present study, the filter setting of the LSC was adapted for the measurement of the far-red emitting dye cyanine 7 (Cy7), thereby increasing the number of measurable commercially available fluorochromes. METHODS: The optical filters of the LSC were replaced-photomultiplier (PMT) 3/allophycocyanin (APC): 740-nm dichroic long pass, and 670-/55-nm bandpass; PMT 4/Cy7: 810-/90-nm bandpass. Peripheral blood leukocytes were stained directly by fluorochrome-labeled antibodies or by indirect staining. The tandem dyes of Cy7 (phycoerythrin [PE] Cy7, APC-Cy7) and the fluorochromes fluorescein isothiocyanate (FITC), PE, PE Cy5, and APC were tested alone and in different combinations. RESULTS: With the new filter combination and tandem fluorochromes, Cy7 was measurable at 488-nm (argon laser) or 633-nm (helium-neon laser) excitation. Resolution was in the range of FITC for PE-Cy7 but approximately 30% lower for APC-Cy7; spillover into the respective donor fluorochrome channel for both tandem dyes was prominent. A six-color panel for leukocyte subtyping was designed. CONCLUSIONS: With this adaptation, it is possible to measure the tandem conjugates PE-Cy7 and APC-Cy7. This new setup opens the way for six-color immunophenotyping by LSC. PMID- 12116358 TI - Long wavelength fluorophores and cell-by-cell correction for autofluorescence significantly improves the accuracy of flow cytometric energy transfer measurements on a dual-laser benchtop flow cytometer. AB - BACKGROUND: Flow cytometric fluorescence resonance energy transfer (FCET) is an efficient method to map associations between biomolecules because of its high sensitivity to changes in molecular distances in the range of 1-10 nm. However, the requirement for a dual-laser instrument and the need for a relatively high signal-to-noise system (i.e., high expression level of the molecules) pose limitations to a wide application of the method. METHODS: Antibodies conjugated to cyanines 3 and 5 (Cy3 and Cy5) were used to label membrane proteins on the cell surface. FCET measurements were made on a widely used benchtop dual-laser flow cytometer, the FACSCalibur, by using cell-by-cell analysis of energy transfer efficiency.ResultsTo increase the accuracy of FCET measurements, we applied a long wavelength donor-acceptor pair, Cy3 and Cy5, which beneficially affected the signal-to-noise ratio in comparison with the classic pair of fluorescein and rhodamine. A new algorithm for cell-by-cell correction of autofluorescence further improved the sensitivity of the technique; cell subpopulations with only slightly different FCET efficiencies could be identified. The new FCET technique was tested on various direct and indirect immunofluorescent labeling strategies. The highest FCET values could be measured when applying direct labeling on both (donor and acceptor) sides. Upon increasing the complexity of the labeling scheme by introducing secondary antibodies, we detected a decrease in the energy transfer efficiency. CONCLUSIONS: We developed a new FCET protocol by applying long wavelength excitation and detection of fluorescence and by refining autofluorescence correction. The increased accuracy of the new method makes cells with low receptor expression amenable to FCET investigation, and the new approach can be implemented easily on a commercially available dual-laser flow cytometer, such as a FACSCalibur. PMID- 12116359 TI - Development and characterization of Ni-NTA-bearing microspheres. AB - BACKGROUND: For ease of purification, proteins are often expressed with a short affinity sequence of five or six adjacent histidine residues (His-tag). This His tag binds to the metal of metal chelator complexes such as Ni(2+) nitrilotriacetic acid (Ni-NTA) or -iminodiacetic acid (Ni-IDA). Chromatography resins bearing covalently attached metal chelator complexes are used widely for the easy affinity purification of His-tagged proteins or peptides. Because Ni-NTA microspheres were not commercially available at the beginning of our studies, we prepared and characterized such microspheres to immobilize His-tagged proteins and study their interactions. Our microspheres are of three types: (a) metal chelator complexes bound covalently to polystyrene microspheres, (b) metal chelator complexes bound covalently to silica microspheres, and (c) lipid-linked metal chelator complexes adsorbed to silica microspheres forming self-assembled bilayer membranes where the metal chelators have lateral mobility. METHODS: The microspheres bearing covalently attached Ni-chelator were synthesized by reacting a primary amine-bearing Ni-NTA ligand with carboxy-functionalized microspheres and then loading with Ni(2+). Microspheres with laterally mobile metal chelator were made by incubating glass microspheres with liposomes containing phosphatidylcholine (PC) and the metal chelating lipid 1,2-dioleoyl-sn-glycero-3 [(N (5-amino-1-carboxypentyl)iminodiacetic acid)succinyl]. Binding of a His tagged enhanced green fluorescent protein (EGFP) was used to characterize these microspheres by flow cytometry for their specificity, sensitivity, capacity and stability. RESULTS: While all micospheres specifically bind His-tagged proteins, the conditions to achieve this are different for the polystyrene- and silica based spheres. All three types of microspheres bind His-EGFP with saturation occurring at 30-50 nM and an apparent avidity (concentration of half-maximal binding) of approximately 1 to 2 x 10(-8) M at pH 7.4. Binding of His-EGFP is inhibited by imidazole or ethylene-diaminetetraacetic acid (EDTA). Polystyrene Ni NTA microspheres showed significant nonspecific binding as measured by binding in the presence of imidazole or EDTA or by binding of fluorescent proteins lacking a His-tag. This nonspecific binding of proteins to and aggregation of polystyrene spheres could only be prevented by the inclusion of low concentrations of Tween 20, but not by including bovine serum albumin (BSA), polyethylene glycols, or polyvinylpyrrolidones as blocking agents. In contrast, silica-based microspheres with covalently attached Ni-NTA or silica microspheres bearing adsorbed bilayers that contain Ni-NTA-lipid showed little nonspecific binding in the presence of BSA. Our results on the stability of immobilization indicate that washing destabilizes the binding of His-tagged proteins to Ni-NTA microspheres. This binding consists of two interactions of different affinities. We also demonstrate that limited multiplexed analysis with differently sized silica microspheres bearing the Ni-NTA-lipid is feasible. CONCLUSIONS: The microspheres described are well suited to selectively immobilize His-tagged proteins to analyze their interactions by flow cytometry. The affinity and kinetic stability of the interaction of His-tagged proteins with Ni-NTA are insufficient to use Ni-NTA microspheres in multiplexed analysis formats where different His-tagged proteins are bound to distinct microspheres. Improvements towards this end (improved chelators and/or improved affinity tags) are critical for extending the use of this method. We are currently working on novel chelators to strengthen the stability of immobilization of His-tagged proteins to surfaces. Such improvements would greatly enhance the analysis of interactions of immobilized His-tagged proteins and could make the development of microsphere-based arrays with His tagged protein/antibody possible. PMID- 12116360 TI - Five-color flow cytometric analysis of swine lymphocytes for detection of proliferation, apoptosis, viability, and phenotype. AB - BACKGROUND: The objective of this study was to develop a method to simultaneously examine phenotype, proliferation, apoptosis, and death of antigen-stimulated porcine lymphocytes. METHODS: Peripheral blood mononuclear cells (PBMCs) were isolated from pigs vaccinated with a Brachyspira hyodysenteriae bacterin. RESULTS: Once isolated, PBMCs were stained with the fluorescent membrane intercalating dye, PKH67, and cultured with or without B. hyodysenteriae whole cell sonicate antigen. Serial samples of nonstimulated and B. hyodysenteriae stimulated PBMCs were harvested for flow cytometric analysis. Fluorochrome excitation was performed with spatially separated air-cooled argon and red helium neon laser beams. Five-color analysis included signal detection of PKH67 (proliferation), phycoerythrin (cell surface antigen), Texas Red phycoerythrin tandem (cell surface antigen), allophycocyanin (annexin V), and 7-amino actinomysin D (7AAD; viability). For analysis, gates were set on live (annexin V( ), 7AAD(-)), intact apoptotic (annexin V(+), 7AAD(dim)), and live plus intact apoptotic (annexin V(+/-), 7AAD(dim/-)) cells, and the phenotypes of PBMCs within these populations were determined during the course of the in vitro response. Dead cells (i.e., 7AAD(bright)) were excluded from the analysis. CONCLUSION: Application of this method for the determination of porcine lymphocyte subset proliferation is presented. PMID- 12116361 TI - Optical compartmentation of vegetating algae species as a basis for their growth specific characterization. AB - BACKGROUND: The number of microalgal strains known to date is enormous and continuously growing, and their characterization accordingly requires quick and reliable methodologies. METHODS: Asynchronously growing logarithmic (3- and 6-day cultures) and stationary (9-day cultures) phase cell populations of two algae species that are difficult to distinguish microscopically (one Chlorella sp., C. vulgaris [c-27], and another that might belong to the same genus, SA-3 algae exsymbiotic from Paramecium bursaria) were characterized by means of flow cytometry (FCM). Forward light scatter (FSC) of algae was monitored in association with their 90 degrees side light scatter (SSC) and fluorescence of endogenous chlorophyll (FL3-height). RESULTS: Two-parameter FSC versus SSC and FSC versus FL3-height plots distinctly showed growth-specific compartmentation of algae into discrete cell subpopulations staying at a particular stage of the life cycle, and numbers of cells constituting these subpopulations could be quantitated. The growth pattern of C. vulgaris (c-27) differed substantially from that of SA-3 algae, particularly in the late-logarithmic (6-day) cultures. At this phase of growth, C. vulgaris (c-27) cells compartmentalized into three subpopulations, whereas SA-3 cells compartmentalized into two subpopulations. Different compartmentations of optical signals from late-logarithmic phase SA-3 algae and C. vulgaris (c-27) likely were caused by the differences in timing of the life cycle stages of these types of cells. CONCLUSIONS: Growth-specific compartmentation of vegetating microalgae by FCM provides a good basis for characterization of morphologically similar algae species. Because algae are also present in symbiotic relationships with other organisms, this tool might be of potential interest for the study of symbiosis mechanisms. PMID- 12116362 TI - Measurement of phagosomal pH of normal and CGD-like human neutrophils by dual fluorescence flow cytometry. AB - BACKGROUND: Phagosomal pH is thought to play an important role in the antimicrobial activity of polymorphonuclear leukocytes (PMNs). In this study, we set up a method for a rapid and accurate measurement of phagosomal pH in PMNs with the use of Candida albicans doubly labeled with a pH-insensitive and a pH sensitive probe and flow cytometry. METHODS: Heat-killed, serum-opsonized C. albicans were doubly labeled with fluorescein, a pH-sensitive probe, and rhodamine, a pH-insensitive probe, and incubated with human PMNs. Flow cytometric readings of PMN-associated Candida were then taken, and the intraphagosomal pH was calculated on the basis of the ratio of fluorescein:rhodamine fluorescence by using a calibration curve obtained after equilibration of phagosomal pH with different external pH values after addition of digitonin. RESULTS: A rapid rise in phagosomal pH, which reached pH 7.8, was observed 2 min after initiation of phagocytosis and progressively declined to pH 6.9 after 15 min. Such a rise was not observed in PMNs with defective microbicidal activity (deficient in nicotinamide adenine dinucleotide phosphate oxidase), where phagosomal pH dropped to pH 6.6, 2 min after phagocytosis. The abnormal initial acidification in PMNs deficient in nicotinamide adenine dinucleotide phosphate oxidase was prevented by using lysosomotropic weak bases or the vacuolar-type H(+) pump inhibitor concanamycin A. CONCLUSIONS: Phagosomal pH of PMNs can be easily and accurately measured by dual fluorescence flow cytometry. The method can be applied to assess phagosomal pH in PMNs with defective microbicidal activity and to monitor the outcome of pharmacologic interventions aimed at correcting its abnormalities. PMID- 12116363 TI - Heterogeneity of freshly isolated human tonsil dendritic cells demonstrated by intracellular markers, phagocytosis, and membrane dye transfer. AB - BACKGROUND: Heterogeneity within human dendritic cells (DCs) has been described but its functional relationships to cells of macrophage lineage and its role in human immunodeficiency virus (HIV) infection in vivo remain unclear. METHODS: Tonsil macrophages and DCs were isolated from low-density cells by negative selection and DCs were sorted into myeloid and plasmacytoid populations using antibodies to CD11c or CD123. Phagocytosis of latex beads and uptake of dye labeled target cells were compared by flow cytometry and CD68 and S-100 by immunofluorescence on cytospins of sorted cells. RESULTS: Bead uptake and membrane dye transfer were found in both blood and tonsil CD11c(+) DCs and in CD14(+) cells particularly from blood monocytes. CD11c(-) DCs were poorly phagocytic but took up fluorescent dye from intact, necrotic or apoptotic cells. Tonsil DCs and macrophages expressed both CD68 and S-100 but CD11c(-) DCs expressed CD68 only. CONCLUSIONS: Freshly isolated CD11c(+) tonsil DCs are similar to CD14(+) macrophages in phagocytic function but the poorly phagocytic CD11c(-) DCs can also take up membrane from target cells. The intracellular markers commonly used to identify DCs and macrophages in situ do not identify accurately the CD11c(-) DC subset nor do they distinguish tonsil macrophages from DCs. PMID- 12116366 TI - Flow cytometric S-phase fraction measurement in breast carcinoma: Influence of software and histogram resolution. AB - BACKGROUND: S-phase fraction (SPF) measurement by flow cytometry is a clinically useful prognostic factor in patients with breast carcinoma. Standardized SPF determination is essential. As part of a multicenter study, we evaluated the influence of the choice of software and histogram resolution (256, 512, or 1,024 channels) on SPF quantification. METHODS: One hundred thirty-three DNA histograms were analyzed in three laboratories with Modfit 5.2, Modfit LT, and Multicycle AV software. Strict rules for histogram interpretation and software management were applied. The following five options were compared: MF 5.2 1024, MF 5.2 256, MF LT 256, MC AV 256, and MC AV 512. RESULTS: In the DNA diploid and aneuploid groups, SPF distributions were not statistically different among the five options. Excellent quantitative correlations were obtained between pairs of options. When using tertiles as cutpoints for SPF classification, concordance rates ranged from 79.7% to 93.2% for DNA diploid samples and from 87.8% to 95.9% for DNA aneuploid samples, the best results being obtained with software working with a similar histogram resolution. CONCLUSIONS: Standardized use of commercially available software, including the choice of histogram resolution, provides comparable SPF results. PMID- 12116365 TI - Flow cytometric analysis of breast cancer resistance protein expression and function. AB - BACKGROUND: The breast cancer resistance protein (BCRP) is an ATP-binding cassette (ABC) half-transporter that mediates energy-dependent drug efflux. Assessing the clinical relevance of the BCRP will require sensitive and specific methods for detecting its expression and function that allow high-volume specimen throughput and employ widely available instrumentation. METHODS: The BXP-34 and BXP-21 monoclonal antibodies were evaluated for flow cytometric detection of BCRP expression. The modulation of efflux of rhodamine-123, 3,3' diethyloxacarbocyanine iodide, doxorubicin, and mitoxantrone by fumitremorgin C was studied as an assay for BCRP function in BCRP-overexpressing cell lines and controls. RESULTS: BXP-34 and BXP-21 allowed detection of BCRP expression by flow cytometry in all BCRP-expressing cell lines. Mitoxantrone was the only substrate transported by BCRP in all lines, and with mitoxantrone at a 3-microM concentration, light emission (>670 nm) caused by excitation at 488 nm was sufficiently intense to allow detection of differences in retention associated with low levels of BCRP expression. CONCLUSIONS: Immunophenotyping with BXP-21 or BXP-34 and fumitremorgin C modulation of mitoxantrone retention allow detection of BCRP expression and function by flow cytometry with standard instrumentation. These assays will facilitate determination of the role of BCRP in clinical drug resistance. PMID- 12116367 TI - Measurement of nuclear factor-kappa B translocation on lipopolysaccharide activated human dendritic cells by confocal microscopy and flow cytometry. AB - BACKGROUND: Nuclear factor kappa B (NF-kappaB) is a ubiquitously expressed transcription factor that regulates cytokine and immunoglobulin (Ig) gene expression. In most cell types, the inactive p50/p65 NF-kappaB heterodimer is located in the cytoplasm, complexed to its IkappaB inhibitory unit. Stimulation of cells by various reagents such as bacterial endotoxin or cytokines leads to a dissociation of NF-kappaB from IkappaB and a rapid translocation of free NF kappaB to the nucleus. The aim of this article is to define optimal conditions for the measurement of NF-kappaB translocation by both confocal microscopy and flow cytometry. METHODS: Four commercial anti-NF-kappaB antibodies were evaluated by confocal microscopy, after using two methods of fixation and permeabilization of the cells. These antibodies were examined further by flow cytometry on purified nuclei. RESULTS: Paraformaldehyde-methanol treatment of dendritic cells is a good combination to visualize NF-kappaB translocation by confocal microscopy. Three of the four antibodies tested gave good results on nonactivated and on lipopolysaccharide (LPS)-activated dendritic cells. The measurement of NF kappaB translocation by flow cytometry on purified nuclei is a quick and sensitive method. Only one of the four evaluated antibodies showed a significant difference between nonactivated and activated cells. CONCLUSIONS; Microscopy and flow cytometry are quick and reproducible methods to measure NF-kappaB translocation and can be adapted to identify new molecules that activate dendritic cells. PMID- 12116368 TI - Flow cytometric detection of activated mouse integrin alphaIIbbeta3 with a novel monoclonal antibody. AB - BACKGROUND: Integrin alphaIIbbeta3 mediates platelet adhesion and aggregation and plays a crucial role in thrombosis and hemostasis. alphaIIbbeta3 is expressed in a low affinity state on resting platelets. Upon platelet activation, alphaIIbbeta3 shifts to a high affinity conformation that efficiently binds its ligands. On human platelets, the high affinity conformation of alphaIIbbeta3 is detected by the monoclonal antibody (mAb), PAC-1. However, a reagent with binding specificity to high affinity mouse alphaIIbbeta3 has not been described so far. METHODS: A novel rat mAb directed against mouse alphaIIbbeta3 (JON/A) was generated and characterized. JON/A was conjugated with fluorescein isothiocyanate (JON/A(FITC)) or with R-phycoerythrin (JON/A(PE)) and used for flow cytometric analysis of mouse platelets. RESULTS: Although JON/A(FITC) bound to resting and activated platelets, virtually no binding of the larger JON/A(PE) to resting platelets was detectable. However, strong binding of JON/A(PE) occurred on platelet activation in a dose-dependent manner. Binding of JON/A(PE) required extracellular free calcium and was irreversible, thereby stabilizing the high affinity conformation of alphaIIbbeta3. CONCLUSION: JON/A(PE) is the first tool for direct assessment of integrin alphaIIbbeta3 activation in mice. Furthermore, JON/A(FITC) and JON/A(PE) provide the first examples of fluorescent antibody derivatives with identical antigenic specificity that allow the discrimination between the resting and the activated state of an integrin. PMID- 12116369 TI - Detection and relocation of cord blood nucleated red blood cells by laser scanning cytometry. AB - BACKGROUND: Fetal nucleated red blood cells (NRBC) present in the peripheral blood of pregnant women at low frequency are a potential target for noninvasive prenatal diagnostics. METHODS: CD71-enriched cells from male cord blood (CB) were stained for the gamma chain of HbF (Hb-gamma) and cytocentrifuged. Fluorescence in situ hybridization (FISH) was done for the Y chromosome. Following staining of the nucleus with TO-PRO-3, laser scanning cytometry was performed. Artificial mixtures of small volumes of male CB and blood drawn from nonpregnant females were analyzed. RESULTS: In CB, 59% of events double positive for Hb-gamma and TO PRO-3 were identified as CB-NRBC. In contamination studies, male fetal CB-NRBC were identified perfectly on the basis of morphologic characteristics and FISH reactivity following relocation and visual assessment. Mean recovery was 8.7%. CONCLUSIONS: Laser scanning cytometry of preenriched fetal NRBC may offer a promising way for noninvasive prenatal diagnostics. This is because it provides a virtual enrichment step and the position on the slides of cells visually confirmed to correspond to fetal NRBC is known. Further experimental procedures on well-defined and located target cells may be feasible. PMID- 12116370 TI - Use of SYTOX green dye in the flow cytometric analysis of bacterial phagocytosis. AB - BACKGROUND: Fluorescein isothiocyanate (FITC) is used widely to label the targets used in flow cytometric phagocytosis assays. Unfortunately, the fluorescence intensity of phagocytosed FITC-labeled targets is influenced by changes in intracellular pH level, making quantitative measurements with this fluorophore problematic. We describe the use of SYTOX green nucleic acid stain to measure phagocytosis by flow cytometry. METHODS: Suspensions of isopropyl alcohol permeabilized Escherichia coli DH5alpha were stained with the SYTOX green dye and then incubated with resident peritoneal macrophages. The samples were analyzed by flow cytometry and phagocytosis was determined by gating the cells. RESULTS: Results are expressed as percentage of phagocyte-associated green fluorescent cells. The validity of the method was shown by the effects of a phagocytosis inhibitor (incubation at 4 degrees C) or enhancer (gamma interferon [IFN- gamma] treatment) being accurately assessed with this assay. CONCLUSIONS: The method described was reproducible and provides an advantageous alternative to the use of FITC to label bacteria for the flow cytometric measurement of target uptake by phagocytic cells. PMID- 12116371 TI - Use of phycoerythrin and allophycocyanin for fluorescence resonance energy transfer analyzed by flow cytometry: advantages and limitations. AB - BACKGROUND: This study validates the use of phycoerythrin (PE) and allophycocyanin (APC) for fluorescence energy transfer (FRET) analyzed by flow cytometry. METHODS: FRET was detected when a pair of antibody conjugates directed against two noncompetitive epitopes on the same CD8alpha chain was used. FRET was also detected between antibody conjugate pairs specific for the two chains of the heterodimeric alpha (4)beta(1) integrin. Similarly, the association of T-cell receptor (TCR) with a soluble antigen ligand was detected by FRET when anti-TCR antibody and MHC class I/peptide complexes (<>) were used. RESULTS: FRET efficiency was always less than 10%, probably because of steric effects associated with the size and structure of PE and APC. Some suggestions are given to take into account this and other effects (e.g., donor and acceptor concentrations) for a better interpretation of FRET results obtained with this pair of fluorochromes. CONCLUSIONS: We conclude that FRET assays can be carried out easily with commercially available antibodies and flow cytometers to study arrays of multimolecular complexes. PMID- 12116372 TI - Stable expression of Anthozoa fluorescent proteins in mammalian cells. AB - BACKGROUND: Fluorescent proteins have become invaluable reporters in many areas of cellular and developmental biology. An enhanced version of the Aequorea victoria green fluorescent protein (AvEGFP) is the most widely used fluorescent protein. For a variety of reasons, it is useful to have alternative fluorescent proteins to AvEGFP. METHODS: The cDNA sequences for enhanced variants of the Anemonia cyan fluorescent protein (AmCyan1), as well as the Zoanthus green (ZsGreen1) and yellow (ZsYellow1) fluorescent proteins, were cloned downstream of a constitutive cytomegalovirus (CMV) promoter within a retroviral expression vector. NIH3T3, HEK293, SW620, and WM35 cells were transduced with recombinant retroviruses at a low multiplicity of infection (MOI) to bias for single-copy integration. Both unselected and stably selected cells transduced with the retroviral expression constructs were characterized. Expression of each fluorescent protein in cells was detected using flow cytometry and fluorescence microscopy with filter sets typically used for AvEGFP/fluorescein isothiocyanate (FITC) detection and was compared with the expression of AvEGFP. In addition, a fluorescence plate reader with several excitation and emission filter sets was used for detection. RESULTS: Expression of each protein was observable by fluorescence microscopy. Under given conditions of flow cytometry, the ZsGreen1 mean fluorescence was approximately 3-fold, 10-fold, and 50-fold greater than that of AvEGFP, ZsYellow1, and AmCyan1, respectively. AmCyan1, ZsGreen1, and AvEGFP were detected by a fluorescence plate reader. CONCLUSION: We determined that fluorescent proteins from Anthozoa species are detectable using a standard flow cytometer and fluorescence microscope. All of the mammalian cell lines tested expressed detectable levels of fluorescent proteins from stable integrated provirus. In cell lines where the AvEGFP protein is toxic or poorly expressed, these Anthozoa fluorescent proteins may serve as alternative fluorescent reporters. PMID- 12116374 TI - A novel flow cytometric technique for drug cytotoxicity gives results comparable to colony-forming assays. AB - BACKGROUND: Drug sensitivity is commonly determined by assays that utilize colony formation to discriminate between surviving and lethally treated cells. These assays require cells with high plating efficiency that form discernible colonies, are time-consuming and laborious, and require manual counting of large numbers of colonies. To overcome these drawbacks, we developed a flow cytometric technique that assays survival of proliferative capacity in cultured cells. METHODS: Labeling with bromodeoxyuridine for 72 h followed by bivariate Hoechst 33258/ethidium bromide flow cytometry allows discrimination of nonproliferating cells from those that have undergone one to three divisions. Addition of an internal standard, chicken erythrocyte nuclei, permits determination of total cell number. To validate our assay, we used flow and colony-forming assays to determine the sensitivity of cell lines derived from Werner syndrome patients and unaffected individuals to 4-nitroquinoline-1-oxide (4NQO) and camptothecin. RESULTS: The flow and colony-forming assays yielded comparable sensitivity for each drug and essentially identical increases in drug sensitivity exhibited by Werner syndrome cells. CONCLUSION: Our results indicate that the flow assay is a less laborious surrogate for colony-forming assays. The flow technique will also facilitate the analysis of drug sensitivity in cells that are not amenable to colony-forming assays. PMID- 12116375 TI - G(1)/S but not G(0)/G(1)cell fraction is related to 5-fluorouracil cytotoxicity. AB - BACKGROUND: Bromodeoxyuridine (BrdU) cell cycle analysis using flow cytometry is of clinical interest for making treatment decisions or for predicting response and survival, through proliferation rate (labeling index or S-phase fraction) assessment or T(pot) calculation. Thymidylate synthase expression was tested in vitro, in vivo, and clinically as a prognostic factor for 5-fluorouracil (5FU) sensitivity. However, results were still controversial. Moreover, we had reported that 5FU sensitivity was related to the labeling index of untreated cell cultures. METHODS: We used six human cancer cell lines that exhibited a wide range of 5FU sensitivity. Cell cycle analysis was performed using flow cytometry monovariate propidium iodide (PI) analysis and bivariate distributions of BrdU incorporation versus DNA content. 5FU sensitivity was assayed using a 3-(4,5 dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) colorimetric assay. RESULTS: In all cell lines, 5FU exposure resulted in a statistically significant G(1)/S accumulation. No statistically significant relationship was seen between G(0)/G(1) delay determined by monovariate analysis and 5FU sensitivity. However, 5FU sensitivity was statistically correlated to the labeling index and G(1)/S subpopulation assessed with bivariate analysis using BrdU incorporation versus DNA content. CONCLUSIONS: Cellular proliferation parameters using BrdU incorporation are more informative than PI for in vitro 5FU sensitivity. Because BrdU incorporation could be assessed clinically, it could also be informative for 5FU clinical response prediction. PMID- 12116376 TI - Analysis of radiation-induced apoptosis in human lymphocytes: flow cytometry using Annexin V and propidium iodide versus the neutral comet assay. AB - BACKGROUND: The neutral comet assay was devised to measure double-stranded DNA breaks, but it has also been used to measure apoptosis based on its characteristic DNA fragmentation patterns. There is still uncertainty about the reliability of this method. By comparing the comet assay with a flow cytometry method that uses Annexin V binding to apoptotic cells, we have provided further evidence for evaluating the usefulness of the comet assay for detecting apoptosis. METHODS: Apoptosis was induced in human peripheral blood mononuclear cells (PBMC) by ionizing radiation and measured using the comet assay and a flow cytometry method that measures Annexin V and propidium iodide (PI) staining. RESULTS: The Annexin V flow cytometry assay distinguished among early apoptosis, late apoptosis, and an apoptotic or necrotic phase in which the cells were labeled with both Annexin V and PI. The comet assay detected only the latter two phases of apoptosis. CONCLUSIONS: The comet assay is a useful tool for measuring the late stages of apoptosis whereas the Annexin V assay measures higher amounts of apoptosis because it can detect cells in an earlier stage of the apoptotic pathway. PMID- 12116377 TI - Flow cytometry measurement of the DNA contents of G0/G1 diploid cells from three different teleost fish species. AB - BACKGROUND: Although there is a lot information in the literature about genome size in fish, a high variability among data for the same species is reported, being mainly related to methodological aspects. Flow cytometry-based fluorescence measurements of intercalating dyes is the most attractive approach due to its precision, objectivity, high speed, and relative simplicity. METHODS: We analyze the DNA content of G0/G1 diploid nuclei of three teleost species (Carassius auratus, Tinca tinca, and Danio rerio) using flow cytometry. Forty-three animals were used and up to 50,000 retinal cells were analyzed per sample. Propidium iodide-associated fluorescence was assessed using a FACSCalibur flow cytometer. Standard human leukocytes were used as a reference. RESULTS: Our results show that C. auratus (3.584 +/- 0.058 pg per nucleus) and D. rerio (3.357 +/- 0.074 pg per nucleus) showed similar DNA contents per cell, whereas it was significantly lower (2.398 +/- 0.038 pg per nucleus) in T. tinca. Interestingly, a low intraspecies variability was observed, the coefficient of variation being 1.608%, 2.198%, and 1.573% for C. auratus, D. rerio, and T. tinca, respectively. CONCLUSIONS: The methodology used in this study provides an accurate and easy measurement of the genome size of a species. PMID- 12116378 TI - Nonparametric discriminant analysis of phytoplankton species using data from analytical flow cytometry. AB - BACKGROUND: Analytical flow cytometry (AFC) provides rapid and accurate measurement of particles from heterogeneous populations. AFC has been used to classify and identify phytoplankton species, but most methods of discriminant analysis of resulting data have depended on normality assumptions and outcomes have been disappointing. METHODS AND RESULTS: In this study, we consider nonparametric methods based on density estimation. In addition to the familiar kernel method, methods based on wavelets are also implemented. Full five dimensional wavelet estimation proves to be computationally prohibitive with current workstation power, so we employ projection pursuit for reduction of dimensionality. AFC typically produces very large samples, so we also investigate data simplification through binning. Further modifications to the discrimination strategy are suggested by specific features of phytoplankton data, namely, a hierarchical group structure, the possible presence of many groups, and the likelihood of encountering an aberrant group in a test sample. CONCLUSIONS: We apply all the resultant procedures to appropriate subsets of a very large data set, demonstrate their efficacy, and compare their error rates with those of more conventional methods. We further show that incorporation of the specific features of phytoplankton data into the analysis leads to improved results and provides a general framework for analysis of such data. PMID- 12116379 TI - Comparing telomere length of sister chromatids in human lymphocytes using three dimensional confocal microscopy. AB - BACKGROUND: The length of the terminal sequences of linear chromosomes changes dynamically during cellular proliferation. A crucial element in the study of telomere-related regulation mechanisms is the ability to measure telomere lengths of individual chromosomes. Individual telomere lengths can be measured using digital imaging fluorescence microscopy-based techniques. We extended this method using confocal microscopy for the acquisition of three-dimensional (3D) images. Consequently, variations in measured signal intensities due to erroneous focusing are avoided. METHODS: We employed our 3D telomere sizing method to compare telomere lengths of sister chromatids within metaphase preparations from human lymphocytes. The samples were treated following a quantitative fluorescence in situ hybridization (Q-FISH) protocol using fluorescein isothiocyanate (FITC) labeled telomeric peptidic nucleic acid (PNA) probes and propidium iodide (PI) counterstain. RESULTS: We demonstrated that the telomere lengths of two sister chromatids are not necessarily equal in human lymphocytes. Profound statistical analysis demonstrated significant differences in the distribution of the sister chromatid telomere lengths, but we were not able to prove a discrete distribution of telomere sister ratios. These telomere length differences were more apparent in older individuals. CONCLUSION: Whereas the majority of sister telomere pairs have equal lengths, surprisingly, a minority was significantly different in each individual studied. We are convinced that these observations are not linked to the methodology or the protocol applied. We suggest that a biological phenomenon might be involved. PMID- 12116380 TI - Flow cytometric discrimination of various phycobilin-containing phytoplankton groups in a hypertrophic reservoir. AB - BACKGROUND: Knowledge of phytoplankton structure is important information in water quality control. Lake restoration and sanitation measures in particular must be evaluated on the organismic level to valuate biological effects and assess the risk of potentially toxic Cyanobacteria blooms. We used and comparatively tested three independent methods for phytoplankton analysis in a hypertrophic reservoir under restoration. METHODS: Nine unialgal cultures and outdoor samples were examined by high-performance liquid chromatography pigment analysis, microscopical cell counting, and flow cytometric (FCM) light scatter and fluorescence analysis to measure the percentage contribution of the major algal groups to chlorophyll a and biovolume. The FCM instrument settings and identification criteria were developed using a single excitation wavelength at 514 nm to differentiate nine algal species representing the major groups of algae. Fluorescence was detected at 585, 620, 650, and 680 nm. RESULTS: The results show that FCM is the only method for determining changes in the phytoplankton composition on both a chlorophyll a and biovolume basis. CONCLUSIONS: Each of the three methods has specific advantages and disadvantages, and should be chosen depending on the experimental problem. FCM sorting allows the combination of all three and offers further new perspectives. PMID- 12116381 TI - Efficiency of the multicanonical simulation method as applied to peptides of increasing size: the heptapeptide deltorphin. AB - The advantage of the multicanonical (MUCA) simulation method of Berg and coworkers over the conventional Metropolis method is in its ability to move a system effectively across energy barriers thereby providing results for a wide range of temperatures. However, a MUCA simulation is based on weights (related to the density of states) that should be determined prior to a production run and their calculation is not straightforward. To overcome this difficulty a procedure has been developed by Berg that calculates the MUCA weights automatically. In a previous article (Yasar et al. J Comput Chem 2000, 14, 1251-1261) we extended this procedure to continuous systems and applied it successfully to the small pentapeptide Leu-enkephalin. To investigate the performance of the automated MUCA procedure for larger peptides, we apply it here to deltorphin, a linear heptapeptide with bulky side chains (H-Tyr(1)-D-Met(2)-Phe(3)-His(4)-Leu(5) Met(6)-Asp(7)-NH(2)). As for Leu-enkephalin, deltorphin is modeled in vacuum by the potential energy function ECEPP. MUCA is found to perform well. A weak second peak is seen for the specific heat, which is given a special attention. By minimizing the energy of structures along the trajectory it is found that MUCA provides a good conformational coverage of the low energy region of the molecule. These latter results are compared with conformational coverage obtained by the Monte Carlo minimization method of Li and Scheraga. PMID- 12116382 TI - Applicability of MNDO techniques AM1 and PM3 to ring-structured polymers. AB - Semiempirical Hartree-Fock techniques are widely used to study properties of long ring-structured chains, although these types of systems were not included in the original parametrization ensembles. These techniques are very useful for an ample class of studies, and their predictive power should be tested. We present here a study of the applicability of some techniques from the NDDO family (MNDO, AM1, and PM3) to the calculation of the ground state geometries of a specific set of molecules with the ring-structure characteristic. For this we have chosen to compare results against ab initio Restricted Hartree-Fock 6-31G(d,p) calculations, extended to Moller-Plesset 2 perturbation theory for special cases. The systems investigated comprise the orthobenzoquinone (O(2)C(6)H(4)) molecule and dimers (O(2)C(6)H(4))(2), as well as trimers of polyaniline, which present characteristics that extend to several systems of interest in the field of conducting polymers, such as ring structure and heterosubstitution. We focus on the torsion between rings, because this angle is known to affect strongly the electronic and optical properties of conjugated polymers. We find that AM1 is always in qualitative agreement with the ab initio results, and is thus indicated for further studies of longer, more complicated chains. PMID- 12116384 TI - An efficient parallel algorithm for the calculation of canonical MP2 energies. AB - We present the parallel version of a previous serial algorithm for the efficient calculation of canonical MP2 energies (Pulay, P.; Saebo, S.; Wolinski, K. Chem Phys Lett 2001, 344, 543). It is based on the Saebo-Almlof direct-integral transformation, coupled with an efficient prescreening of the AO integrals. The parallel algorithm avoids synchronization delays by spawning a second set of slaves during the bin-sort prior to the second half-transformation. Results are presented for systems with up to 2000 basis functions. MP2 energies for molecules with 400-500 basis functions can be routinely calculated to microhartree accuracy on a small number of processors (6-8) in a matter of minutes with modern PC-based parallel computers. PMID- 12116383 TI - Can the calculation of ligand binding free energies be improved with continuum solvent electrostatics and an ideal-gas entropy correction? AB - The prediction of a ligand binding constant requires generating three-dimensional structures of the complex concerned and reliably scoring these structures. Here, the scoring problem is investigated by examining benzamidine-like inhibitors of trypsin, a system for which errors in the structures are small. Precise and consistent binding free energies for the inhibitors are determined experimentally for this test system. To examine possible improvement of scoring methods, we test the suitability of continuum electrostatics to account for solvation effects and use an ideal-gas entropy correction to account for the changes in the degrees of freedom of the ligand. The small observed root-mean-square deviation of 0.55 kcal/mol of the calculated relative to the experimental values indicates that the essentials of the binding process have been captured. Even though all six ligands make the same salt bridge and H-bonds to the protein, the electrostatic contribution varies among the ligands by as much as 2 kcal/mol. Moreover, although the ligands are rigid and similar in size, the entropic terms also significantly affect the relative binding affinities (by up to 2.7 kcal/mol). The present approach to solvation and entropy may allow the ranking of the ligands to be considerably improved at a cost that makes the method applicable to the optimization of lead compounds or to the screening of small collections of ligands. PMID- 12116385 TI - Reducing CAS-SDCI space. Using selected spaces in configuration interaction calculations in an efficient way. AB - A new method is presented, which allows an important reduction of the size of some Configuration Interaction (CI) matrices. Starting from a Complete Active Space (CAS), the numerous configurations that have a small weight in the CAS wave function are eliminated. When excited configurations (e.g., singly and doubly excited) are added to the reference space, the resulting MR-SDCI space is reduced in the same proportion as compared with the full CAS-SDCI. A set of active orbitals is chosen, but some selection of the most relevant excitations is performed because not all the possible excitations act as SDCI generators. Thanks to a new addressing technique, the computational time is drastically reduced, because the new addressing of the selected active space is as efficient as the addressing of the CAS. The presentation of the method is followed by two test calculations on the N(2) and HCCH molecules. For the N(2) the FCI results are taken as a benchmark reference. The outer valence ionization potentials of HCCH are compared to the experimental values. Both examples allow to test the accuracy of the MR-SDCI compared to that of the corresponding CAS-SDCI, despite the noticeable reduction of the CI space. The algorithm is suitable for the dressing techniques that allow for the correction of the size-extensivity error. The corrected results are also shown and discussed. PMID- 12116386 TI - Quasi-degenerate perturbation theory with general multiconfiguration self consistent field reference functions. AB - The quasi-degenerate perturbation theory (QDPT) with complete active space (CAS) self-consistent field (SCF) reference functions is extended to the general multiconfiguration (MC) SCF references functions case. A computational scheme that utilizes both diagrammatic and sum-over-states approaches is presented. The second-order effective Hamiltonian is computed for the external intermediate configurations (including virtual or/and core orbitals) by the diagrammatic approach and for internal intermediate configurations (including only active orbitals) by the configuration interaction matrix-based sum-over-states approach. The method is tested on the calculations of excitation energies of H(2)O, potential energy curves of LiF, and valence excitation energies of H(2)CO. The results show that the present method yields very close results to the corresponding CAS-SCF reference QDPT results and the available experimental values. The deviations from CAS-SCF reference QDPT values are less than 0.1 eV on the average for the excitation energies of H(2)O and less than 1 kcal/mol for the potential energy curves of LiF. In the calculation of the valence excited energies of H(2)CO, the maximum deviation from available experimental values is 0.28 eV. PMID- 12116387 TI - Pattern recognition strategies for molecular surfaces. I. Pattern generation using fuzzy set theory. AB - A new method for the characterization of molecules based on the model approach of molecular surfaces is presented. We use the topographical properties of the surface as well as the electrostatic potential, the local lipophilicity/hydrophilicity, and the hydrogen bond density on the surface for characterization. The definition and the calculation method for these properties are reviewed shortly. The surface is segmented into overlapping patches with similar molecular properties. These patches can be used to represent the characteristic local features of the molecule in a way that is beyond the atomistic resolution but can nevertheless be applied for the analysis of partial similarities of different molecules as well as for the identification of molecular complementarity in a very general sense. The patch representation can be used for different applications, which will be demonstrated in subsequent articles. PMID- 12116388 TI - Pattern recognition strategies for molecular surfaces. II. Surface complementarity. AB - Fuzzy logic based algorithms for the quantitative treatment of complementarity of molecular surfaces are presented. Therein, the overlapping surface patches defined in article I1 of this series are used. The identification of complementary surface patches can be considered as a first step for the solution of molecular docking problems. Standard technologies can then be used for further optimization of the resulting complex structures. The algorithms are applied to 33 biomolecular complexes. After the optimization with a downhill simplex method, for all these complexes one structure was found, which is in very good agreement with the experimental results. PMID- 12116389 TI - Atomic charges, dipole moments, and Fukui functions using the Hirshfeld partitioning of the electron density. AB - In the Hirshfeld partitioning of the electron density, the molecular electron density is decomposed in atomic contributions, proportional to the weight of the isolated atom density in the promolecule density, constructed by superimposing the isolated atom electron densities placed on the positions the atoms have in the molecule. A maximal conservation of the information of the isolated atoms in the atoms-in-molecules is thereby secured. Atomic charges, atomic dipole moments, and Fukui functions resulting from the Hirshfeld partitioning of the electron density are computed for a large series of molecules. In a representative set of organic and hypervalent molecules, they are compared with other commonly used population analysis methods. The expected bond polarities are recovered, but the charges are much smaller compared to other methods. Condensed Fukui functions for a large number of molecules, undergoing an electrophilic or a nucleophilic attack, are computed and compared with the HOMO and LUMO densities, integrated over the Hirshfeld atoms in molecules. PMID- 12116390 TI - Theoretical study on the mechanism of the (3)CH(2) + NO(2) reaction. AB - The complex doublet potential energy surface of the CH(2)NO(2) system is investigated at the B3LYP/6-31G(d,p) and QCISD(T)/6-311G(d,p) (single-point) levels to explore the possible reaction mechanism of the triplet CH(2) radical with NO(2). Forty minimum isomers and 92 transition states are located. For the most relevant reaction pathways, the high-level QCISD(T)/6-311 + G(2df,2p) calculations are performed at the B3LYP/6-31G(d,p) geometries to accurately determine the energetics. It is found that the top attack of the (3)CH(2) radical at the N-atom of NO(2) first forms the branched open-chain H(2)CNO(2) a with no barrier followed by ring closure to give the three-membered ring isomer cC(H(2))ON-O b that will almost barrierlessly dissociate to product P(1) H(2)CO + NO. The lesser followed competitive channel is the 1,3-H-shift of a to isomer HCN(O)OH c, which will take subsequent cis-trans conversion and dissociation to P(2) OH + HCNO. The direct O-extrusion of a to product P(3) (3)O + H(2)CNO is even much less feasible. Because the intermediates and transition states involved in the above three channels are all lower than the reactants in energy, the title reaction is expected to be rapid, as is consistent with the measured large rate constant at room temperature. Formation of the other very low-lying dissociation products such as NH(2) + CO(2), OH + HNCO and H(2)O + NCO seems unlikely due to kinetic hindrance. Moreover, the (3)CH(2) attack at the end-O of NO(2) is a barrier-consumed process, and thus may only be of significance at very high temperatures. The reaction of the singlet CH(2) with NO(2) is also briefly discussed. Our calculated results may assist in future laboratory identification of the products of the title reaction. PMID- 12116391 TI - Modern protein force fields behave comparably in molecular dynamics simulations. AB - Several molecular dynamics simulations were performed on three proteins--bovine apo-calbindin D9K, human interleukin-4 R88Q mutant, and domain IIA of bacillus subtilis glucose permease--with each of the AMBER94, CHARMM22, and OPLS-AA force fields as implemented in CHARMM. Structural and dynamic properties such as solvent-accessible surface area, radius of gyration, deviation from their respective experimental structures, secondary structure, and backbone order parameters are obtained from each of the 2-ns simulations for the purpose of comparing the protein portions of these force fields. For one of the proteins, the interleukin-4 mutant, two independent simulations were performed using the CHARMM22 force field to gauge the sensitivity of some of these properties to the specific trajectory. In general, the force fields tested performed remarkably similarly with differences on the order of those found for the two independent trajectories of interleukin-4 with CHARMM22. When all three proteins are considered together, no force field showed any consistent trend in variations for most of the properties monitored in the study. PMID- 12116392 TI - Quantum chemical geometry optimizations in proteins using crystallographic raw data. AB - A method is developed for the combination of quantum chemical geometry optimizations and crystallographic structure refinement. The method is implemented by integrating the quantum chemical software Turbomole with the crystallographic software Crystallography and NMR System (CNS), using three small procedures transferring information between the two programs. The program (COMQUM X)is used to study the binding of the inhibitor N-methylmesoporphyrin to ferrochelatase, and we show that the method behaves properly and leads to an improvement of the structure of the inhibitor. It allows us to directly quantify in energy terms how much the protein distort the structure of the bound inhibitor compared to the optimum vacuum structure (4-6 kJ/mol). The approach improves the standard combined quantum chemical and molecular mechanics (QC/MM) approach by guaranteeing that the final structure is in accordance with experimental data (the reflections) and avoiding the risk of propagating errors in the crystal coordinates. The program can also be seen as an improvement of standard crystallographic refinement, providing an accurate empirical potential function for any group of interest. The results can be directly interpreted in standard crystallographic terms (e.g., R factors or electron density maps). The method can be used to interpret crystal structures (e.g., the protonation status of metal bound water molecules) and even to locally improve them. PMID- 12116393 TI - Electronic state of small and large cavities for methane hydrate. AB - It is well known that methane hydrate is aggregates of small and large hydrogen bonded water cavities (composed of 12 pentagonal faces of 20 water molecules, and 12 pentagonal and two hexagonal faces of 24 water molecules, respectively) where one methane molecule is encaged. We calculated the methane molecule in vacuum, the small and large cavities by ab initio MO method to clarify the electronic state. The proton of methane in the cavities is shown to form the weak hydrogen bond (O...H[bond]C) between methane and four water molecules, and the H-bond lengths and energies in the small and large cavities were estimated as (0.293 nm, 6.8 kJ/mol) and (0.309 nm, 5.2 kJ/mol), respectively. The calculated values of symmetric C[bond]H stretching frequencies and (13)C-NMR chemical shieldings of the methane in the two cluster cavities show good agreement with the experimental ones observed by Sum et al. and Ripmeester and coworker, respectively. PMID- 12116394 TI - Ab initio study of the mechanism of singlet-dioxygen addition to hydroxyaromatic compounds: negative evidence for the involvement of peroxa and endoperoxide intermediates. AB - In this article we report our study of two possible mechanisms of photooxidation of hydroxyaromatic compounds, involving the intermediacy of zwitterionic peroxa intermediates or 1,4-endoperoxides. To study the pathway of the first of them, as yet unexplored by theoretical methods, a simpler system composed of 1,3-butadiene 1-ol and singlet ((1)Delta(g)) dioxygen was considered first, for which calculations were carried out at the CASSCF/MCQDPT2 ab initio level, mostly with the 6-31G* basis set. The cumulative activation barrier to this reaction was found to be 20 kcal/mol and corresponded to a proton transfer (from the hydroxy oxygen atom to the attached oxygen molecule) in the cyclic zwitterionic peroxacyclopenta-3-ene-2-ol intermediate. This intermediate and the proton transfer transition state were found to have a closed-shell character, which enabled us to estimate the corresponding activation barrier for the phenol dioxygen system by carrying out optimization at the RHF level and single-point calculations at the MP2, CASSCF, and MCQDPT2 levels of theory. The energy barrier to the reaction was estimated to at least about 40 kcal/mol, rendering this mechanism for the phenol-oxygen system unlikely for nonpolar solvents. Similarly, calculations of the barrier to proton transfer from the 1,4-endoperoxide of phenol to its hydroperoxide were found to exceed 60 kcal/mol, eliminating such a mechanism too, which leaves only the earlier postulated mechanisms involving an initial charge or hydrogen-atom transfer to dioxygen as probable. PMID- 12116395 TI - Distance and exposure dependent effective dielectric function. AB - In an effort to develop a dielectric screening function for molecular dynamics simulations of biomolecules in implicit solvent, effective dielectric constants (D(eff)) for a large number of atom pairs in a typical globular protein are calculated by continuum electrostatics. Plots of D(eff) versus the intercharge distance are in general sigmoidal with the characteristics of the curve depending on the distance of the two charges from the dielectric boundary and, secondarily, on the extent to which the area surrounding each charge is occupied by solvent (the "exposure"). The D(eff) values were fitted to an empirical, analytical function of these parameters that reproduces the data reasonably well, although considerable scatter exists in the range of D(eff) from 30 to 80. In the system used for parameterization, the mean square deviation of electrostatic interaction energies with this function is 0.48 kcal/mol, compared to 1.45 for an analytical Generalized Born model and 1.52 for the linear distance-dependent dielectric model. When tested in other proteins of varying size and compactness, the present function is superior to both of the above models, except for a fully unfolded polypeptide chain, where the Generalized Born model is superior. PMID- 12116396 TI - Gravitational smoothing as a global optimization strategy. AB - An optimization scheme for atomic cluster structures, based on exaggerating the importance of the gravitational force, is introduced. Results are presented for calculations on Lennard-Jones clusters of 13, 38, and 55 atoms, and the 13-atom Morse cluster. PMID- 12116397 TI - FA(I):A(+) and FA(II):Cu(+) laser activity and photographic sensitization at the low coordinated surfaces of AgBr ab initio calculations. AB - The twofold potentials of F(A)(I):Au(+) and F(A)(II)Cu(+) color centers at the low coordinated surfaces of AgBr thin films in providing tunable laser activity and photographic sensitization were investigated using ab initio methods of molecular electronic structure calculations. Clusters of variable size were embedded in simulated Coulomb fields that closely approximated the Madelung fields of the host surfaces, and the nearest neighbor ions to the F(A) defect site were allowed to relax to equilibrium in each case. Based on the calculated Stokes shifted optical transition bands and horizontal shifts along the configuration coordinate diagrams, both F(A)(I):Au(+) and F(A)(II):Cu(+) color centers were found to be laser active. The laser activity faded quickly as the bromide ion coordination decreased from 5 (flat) to 4 (edge) to 3 (corner) and as the size of the impurity cation increased from Cu(+) to Au(+). The latter relation was explainable in terms of the axial perturbation of the impurity cation. The smallest calculated Stokes-shift at the corner surface suggested that emission had the same oscillator strength as absorption. All relaxed excited states RESs of the defect containing surfaces were deep below the lower edges of the conduction bands of the defect free ground state surfaces, indicating that F(A)(I):Au(+) and F(A)(II):Cu(+) are suitable laser defects. The probability of orientational destruction of the two centers attributed to the assumed RES saddle point ion configurations along the <110> axis was found to be directly proportional to the size of the impurity cation, with activation energy barriers of about 0.655-3.294 eV for Cu(+), and about 1.887-3.404 eV for Au(+). The possibility of exciton (energy) transfer from the sites of higher coordination to those of lower coordination is demonstrated. The more laser active F(A)(II):Cu(+) center was more easily formed than the less laser active F(A)(I):Au(+) center. The Glasner-Tompkins empirical relation was generalized to include F(A) centers at the low coordinated surfaces of silver bromide thin film. As far as color photographic sensitization is concerned, the lowest unoccupied molecular orbitals of the selected dye molecules in the excited states were high enough for electron injection. F(A) defect formation and rotational diffusion of silver clusters reduced the energy gaps between the excited dye molecules and the lower edges of the conduction bands and allowed for hole injection. About 54-60% of the reduction of silver ions at the flat surface of AgBr was attributed to the host anions and F(A) defect formation, leaving about 40-46% for the reduction of photoelectrons as well as the electrons of the developer or dye molecules. The unrelaxed rotational diffusions of the central Ag(4) by 90 degrees decreased the latter percentage, but were severely hindered by activation energy barriers. PMID- 12116398 TI - Reducing I/O costs for the eigenvalue procedure in large-scale configuration interaction calculations. AB - Several aspects of the matrix diagonalization method used for CI calculations in the COLUMBUS Program System are discussed, including a linear basis-contraction algorithm and the use of a nonorthogonal expansion basis. Both of these features significantly reduce the I/O requirements during the iterations. PMID- 12116399 TI - Use of exchange maximization to generate starting vectors for self-consistent field calculations on metal cluster/adsorbate systems. AB - Localized molecular orbitals (LMOs) derived from exchange maximization with respect to all atom-centered basis functions in the basis set are shown to generate a good starting electronic field for self-consistent field calculations on extended systems such as metal clusters, for which well-defined chemical bonds are not present. Examples studied are a cluster of 20 Ni atoms and the Pt(97)CO, Ag(43)/H(3)CNON, Ag(91)/H(2)CO, and vinylidene/Ni metal cluster plus adsorbate systems. It is also shown that improved starting vectors can be obtained by remixing a subset of the LMOs with the largest exchange eigenvalues through diagonalization of the Fock matrix computed with a null electronic field. Employing only a subset of the exchange-maximized LMOs in the first iterations, and then gradually expanding the space in which the diagonalizations are carried out in succeeding cycles, is shown to be an effective means of guiding the SCF procedure to the converged full-basis solution. PMID- 12116400 TI - Ab initio study on the photochemical behavior of styrene. AB - Ab initio complete active space self-consistent field (CASSCF) and the second order multireference Moller-Plesset calculations have been performed to examine the photochemical behavior of styrene upon the strong S(0)-S(2) electronic excitation in the low-lying excited states. The optimized structure at the S(2)/S(1) conical intersection (CIX) is characterized by a quinoid structure. The transition state (TS) in S(1) is in the vicinity of the S(2)/S(1)-CIX. At the S(1)-TS, two reaction paths branch. One is the relaxation into the stable structure in S(1) and then emission into S(0). The other is the radiationless decay through the S(1)/S(0)-CIX. PMID- 12116401 TI - Intruder state avoidance multireference Moller-Plesset perturbation theory. AB - A new perturbation approach is proposed that enhances the low-order, perturbative convergence by modifying the zeroth-order Hamiltonian in a manner that enlarges any small-energy denominators that may otherwise appear in the perturbative expansion. This intruder state avoidance (ISA) method can be used in conjunction with any perturbative approach, but is most applicable to cases where small energy denominators arise from orthogonal-space states-so-called intruder states that should, under normal circumstances, make a negligible contribution to the target state of interests. This ISA method is used with multireference Moller Plesset (MRMP) perturbation theory on potential energy curves that are otherwise plagued by singularities when treated with (conventional) MRMP; calculation are performed on the 1(3)Sigma(-)(u) state of O(2); and the 2(1)Delta, 3(1)Delta, 2(3)Delta, and 3(3)Delta states of AgH. This approach is also applied to other calculations where MRMP is influenced by intruder states; calculations are performed on the (3)Pi(u) state of N(2), the (3)Pi state of CO, and the 2(1)A' state of formamide. A number of calculations are also performed to illustrate that this approach has little or no effect on MRMP when intruder states are not present in perturbative calculations; vertical excitation energies are computed for the low-lying states of N(2), C(2), CO, formamide, and benzene; the adiabatic (1)A(1)-(3)B(1) energy separation in CH(2), and the spectroscopic parameters of O(2) are also calculated. Vertical excitation energies are also performed on the Q and B bands states of free-base, chlorin, and zinc-chlorin porphyrin, where somewhat larger couplings exists, and-as anticipated-a larger deviation is found between MRMP and ISA-MRMP. PMID- 12116402 TI - Ab Initio calculations of the stabilization energies of the conformational and the structural isomers of C(3)H(7)X where X = F, Cl, and Br. AB - HF, MP2, and B3LYP calculations with different basis sets have been used in the computation of the stabilization energies of C(3)H(7)X isomers, where X is F, Cl, and Br. The experimental stabilization energies of the structural isomers of C(3)H(7)Cl and C(3)H(7)Br have been reproduced via B3LYP calculations. However, the calculated stabilization energies of fluoropropane isomers from their reported enthalpies of formation have been reproduced in all methods of calculations in present work. The experimental relative stabilities of the gauche conformers of 1-fluoro-, 1-chloro-, and 1-bromopropanes have been also reproduced via some of the used calculations in the present work. The effect of the geminal interactions on X atomic charges and on the C-X and C-C bond lengths in halopropane isomers are also discussed. PMID- 12116404 TI - Intensity-carrying modes important for vibrational polarizabilities and hyperpolarizabilities of molecules: derivation from the algebraic properties of formulas and applications. AB - The intensity-carrying mode (ICM) theory is developed for analyzing the vibrational motions that mainly contribute to vibrational polarizabilities and hyperpolarizabilities, which are important for describing intermolecular electrostatic interactions and nonlinear optical properties of molecules. The ICMs are derived from dipole derivatives, polarizability derivatives, and first hyperpolarizability derivatives by using algebraic properties of intensity formulas. The way to obtain explicit forms of ICMs, including the optimization method of the basis of the ICM vector space, is discussed in detail. One- and two dimensional models are constructed on the basis of the ICMs. The theory is applied to three molecules (a push-pull type polyene, a streptocyanine dye cation, and a symmetric neutral polyene) taken as typical examples. It is shown that the ICM theory provides a reasonable picture on the vibrational polarization properties of these molecules. On the basis of this result, the validity of the valence-bond charge transfer (VB-CT) model, which is a one-dimensional model and is widely used to describe the electronic and vibrational properties of dye molecules, is also discussed. PMID- 12116403 TI - Parameterization of OPLS-AA force field for the conformational analysis of macrocyclic polyketides. AB - The parameters for the OPLS-AA potential energy function have been extended to include some functional groups that are present in macrocyclic polyketides. Existing OPLS-AA torsional parameters for alkanes, alcohols, ethers, hemiacetals, esters, and ketoamides were improved based on MP2/aug-cc-pVTZ and MP2/aug-cc-pVDZ calculations. Nonbonded parameters for the sp(3) carbon and oxygen atoms were refined using Monte Carlo simulations of bulk liquids. The resulting force field predicts conformer energies and torsional barriers of alkanes, alcohols, ethers, and hemiacetals with an overall RMS deviation of 0.40 kcal/mol as compared to reference data. Densities of 19 bulk liquids are predicted with an average error of 1.1%, and heats of vaporization are reproduced within 2.4% of experimental values. The force field was used to perform conformational analysis of smaller analogs of the macrocyclic polyketide drug FK506. Structures that adopted low energy conformations similar to that of bound FK506 were identified. The results show that a linker of four ketide units constitutes the shortest effector domain that allows binding of the ketide drugs to FKBP proteins. It is proposed that the exact chemical makeup of the effector domain has little influence on the conformational preference of tetraketides. PMID- 12116405 TI - Exact Gaussian expansions of Slater-type atomic orbitals. AB - Three exact Slater-type function (STO) integral transforms are presented. The STO NG basis set can then be developed using either only 1s Gaussian functions, the same Gaussian exponents for each shell, or using the first Gaussian of each symmetry. The use of any of these three alternatives depends only on appropriate numerical integration techniques. PMID- 12116406 TI - Can we understand the different coordinations and structures of closed-shell metal cation-water clusters? AB - We present a model potential for studying M(q+)(H(2)O)(n=1,9) clusters where M stands for either Na(+), Cs(+), Ca(2+), Ba(2+), or La(3+). The potential energy surfaces (PES) are explored by the Monte Carlo growth method. The results for the most significant equilibrium structures of the PES as well as for energetics are favorably compared to the best ab initio calculations found in the literature and to experimental results. Most of these complexes have a different coordination number in cluster compared to experimental results in solution or solid phase. An interpretation of the coordination number in clusters is given. In order to well describe the transition between the first hydration sphere and the second one we show that an autocoherent treatment of the electric field is necessary to correctly deal with polarization effects. We also explore the influence of the cation properties (charge, size, and polarizability) on both structures and coordination number in clusters, as well as the meaning of the second hydration sphere. Such an approach shows that the leading term in the interaction energy for a molecule in the second hydration sphere is an electrostatic attraction to the cation and not a hydrogen bond with the water molecules in the first hydration sphere. PMID- 12116407 TI - Molecular dynamics simulation of thymine glycol-lesioned DNA reveals specific hydration at the lesion. AB - One nanosecond molecular dynamics (MD) simulation of a thymine glycol (TG) lesioned part of human lymphoblast AG9387 was performed to determine structural changes in DNA molecule caused by the presence of a lesion. These changes can be significant for proper recognition of lesions by a repair enzyme. Thymine glycol is the DNA oxidative lesion formed by addition of OH radicals to C5 and C6 atoms of the thymine base. This lesion is known as causing Cockayne Syndrome-inherited genetic disorder. Distribution of water molecules in a hydration shell around the DNA molecule was analyzed for its contribution to the recognition of the TG lesion by the repair enzyme. The results of MD simulation show there is a specific DNA structural configuration formed at the lesion. After 500 ps the DNA is bent in a kink at the TG site. This change dislocates the glycosyl bond at C5' to a position closer to the DNA surface, and thus its atoms are more exposed to the surrounding water shell. The increased number of water molecules that are close to the TG site indicates that the glycosyl bond may be easily contacted by the repair enzyme. In addition, the higher number of water molecules at the TG site substantiates the importance of water-mediated hydrogen bonds created between the repair enzyme and the DNA upon formation of the complex. Copyright 2001 John Wiley & Sons, Inc. J Comput Chem 22: 1723-1731, 2001 PMID- 12116408 TI - Direct hydride transfer in the reaction mechanism of quinoprotein alcohol dehydrogenases: a quantum mechanical investigation. AB - Oxidation of alcohols by direct hydride transfer to the pyrroloquinoline quinone (PQQ) cofactor of quinoprotein alcohol dehydrogenases has been studied using ab initio quantum mechanical methods. Energies and geometries were calculated at the 6-31G(d,p) level of theory. Comparison of the results obtained for PQQ and several derivatives with available structural and spectroscopic data served to judge the feasibility of the calculations. The role of calcium in the enzymatic reaction mechanism has been investigated. Transition state searches have been conducted at the semiempirical and STO-3G(d) level of theory. It is concluded that hydride transfer from the Calpha-position of the substrate alcohol (or aldehyde) directly to the C(5) carbon of PQQ is energetically feasible. Copyright 2001 John Wiley & Sons, Inc. J Comput Chem 22: 1732-1749, 2001 PMID- 12116409 TI - EUDOC: a computer program for identification of drug interaction sites in macromolecules and drug leads from chemical databases. AB - The completion of the Human Genome Project, the growing effort on proteomics, and the Structural Genomics Initiative have recently intensified the attention being paid to reliable computer docking programs able to identify molecules that can affect the function of a macromolecule through molecular complexation. We report herein an automated computer docking program, EUDOC, for prediction of ligand receptor complexes from 3D receptor structures, including metalloproteins, and for identification of a subset enriched in drug leads from chemical databases. This program was evaluated from the standpoints of force field and sampling issues using 154 experimentally determined ligand-receptor complexes and four "real-life" applications of the EUDOC program. The results provide evidence for the reliability and accuracy of the EUDOC program. In addition, key principles underlying molecular recognition, and the effects of structural water molecules in the active site and different atomic charge models on docking results are discussed. Copyright 2001 John Wiley & Sons, Inc. J Comput Chem 22: 1750-1771, 2001 PMID- 12116410 TI - Viabilty of atomistic potentials for thermodynamic properties of carbon dioxide at low temperatures. AB - Investigation into volumetric and energetic properties of several atomistic models mimicking carbon dioxide geometry and quadrupole momentum covered the liquid-vapor coexistence curve. Thermodynamic integration over a polynomial and an exponential-polynomial path was used to calculate free energy. Computational results showed that model using GROMOS Lennard-Jones parameters was unsuitable for bulk CO(2) simulations. On the other hand, model with potential fitted to reproduce only correct density-pressure relationship in the supercritical region proved to yield correct enthalpy of vaporization and free energy of liquid CO(2) in the low-temperature region. Except for molar volume at the upper part of the vapor-liquid equilibrium line, the bulk properties of exp-6-1 parametrization of ab initio CO(2) potential were in a close agreement with the experimental results. Copyright 2001 John Wiley & Sons, Inc. J Comput Chem 22: 1772-1781, 2001 PMID- 12116411 TI - Derivation of class II force fields. VIII. Derivation of a general quantum mechanical force field for organic compounds. AB - A class II valence force field covering a broad range of organic molecules has been derived employing ab initio quantum mechanical "observables." The procedure includes selecting representative molecules and molecular structures, and systematically sampling their energy surfaces as described by energies and energy first and second derivatives with respect to molecular deformations. In this article the procedure for fitting the force field parameters to these energies and energy derivatives is briefly reviewed. The application of the methodology to the derivation of a class II quantum mechanical force field (QMFF) for 32 organic functional groups is then described. A training set of 400 molecules spanning the 32 functional groups was used to parameterize the force field. The molecular families comprising the functional groups and, within each family, the torsional angles used to sample different conformers, are described. The number of stationary points (equilibria and transition states) for these molecules is given for each functional group. This set contains 1324 stationary structures, with 718 minimum energy structures and 606 transition states. The quality of the fit to the quantum data is gauged based on the deviations between the ab initio and force field energies and energy derivatives. The accuracy with which the QMFF reproduces the ab initio molecular bond lengths, bond angles, torsional angles, vibrational frequencies, and conformational energies is then given for each functional group. Consistently good accuracy is found for these computed properties for the various types of molecules. This demonstrates that the methodology is broadly applicable for the derivation of force field parameters across widely differing types of molecular structures. Copyright 2001 John Wiley & Sons, Inc. J Comput Chem 22: 1782-1800, 2001 PMID- 12116412 TI - EQUIPATH-an equilibrial path tracing routine for the use with the program package GAUSSIAN94. AB - The present article deals with the numerical details important for a successful implementation of the equilibrial path tracing procedure EQUIPATH. The results of an application to the RHF/6-31G(d) potential energy surface of butane are reported and discussed. Sixteen saddle points and several valley-ridge inflection points have been located. For the fragmentation C(4)H(10)--> C(2)H(6)+C(2)H(4) two different reaction paths have been found. Copyright 2001 John Wiley & Sons, Inc. J Comput Chem 22: 1801-1816, 2001 PMID- 12116413 TI - Conformational analysis of the HIV-1 virus reverse transcriptase nonnucleoside inhibitors: TIBO and nevirapine. AB - TIBO (Tetrahydro-imidazo[4,5,1-jk][1,4]-benzodiazepin-2-one) and nevirapine (11 cyclopropyl-5,11-dihydro-4-methyl-6H-dipyrido[3,2-b:2',3'-e][1,4]diazepin-6-one) are models for two classes of nonnucleoside inhibitors of the HIV-1 virus reverse transcriptase (NNRTI). This work presents the parameterization of compounds belonging to these two classes in the Cornell et al. force field through ab initio and semiempirical methods. The new parameters were used in the conformational analysis for TIBO R82913, TIBO R79882, and nevirapine. Various conformational search protocols were tested and the pseudosystematic method SUMM led to the best results. A better understanding of the distribution of conformers was obtained through clustering techniques in the data reduction stages. It was possible to reproduce various experimental data such as the crystallographic structures of the isolated or reverse transcriptase-complexed (RT) molecules. The proton-proton coupling constants (?documentclass{article}?pagestyle{empty}?begin{document}$?,J^{3}_{?mathrm{HH}}$? nd{document}) obtained for TIBO through NMR were also reproduced. Cremer and Pople puckering parameters enabled a precise description of both the conformation of the seven-membered rings and the relative position of the substituents on them. These parameters also demonstrated the efficiency and precision of the two stage clustering method. Copyright 2001 John Wiley & Sons, Inc. J Comput Chem 22: 1817-1829, 2001 PMID- 12116414 TI - Molecular mechanics and dynamics of biomolecules using a solvent continuum model. AB - An easy implementation of molecular mechanics and molecular dynamics simulation using a continuum solvent model is presented that is particularly suitable for biomolecular simulations. The computation of solvation forces is made using the linear Poisson-Boltzmann equation (polar contribution) and the solvent-accessible surface area approach (nonpolar contribution). The feasibility of the methodology is demonstrated on a small protein and a small DNA hairpin. Although the parameters employed in this model must be refined to gain reliability, the performance of the method, with a standard choice of parameters, is comparable with results obtained by explicit water simulations. Copyright 2001 John Wiley & Sons, Inc. J Comput Chem 22: 1830-1842, 2001 PMID- 12116415 TI - A new program for optimizing periodic boundary models of solvated biomolecules (PBCAID). AB - Simulations of solvated macromolecules often use periodic lattices to account for long-range electrostatics and to approximate the surface effects of bulk solvent. The large percentage of solvent molecules in such models (compared to macromolecular atoms) makes these procedures computationally expensive. The cost can be reduced by using periodic cells containing an optimized number of solvent molecules (subject to a minimal distance between the solute and the periodic images). We introduce an easy-to-use program "PBCAID" to initialize and optimize a periodic lattice specified as one of several known space-filling polyhedra. PBCAID reduces the volume of the periodic cell by finding the solute rotation that yields the smallest periodic cell dimensions. The algorithm examines rotations by using only a subset of surface atoms to measure solute/image distances, and by optimizing the distance between the solute and the periodic cell surface. Once the cell dimension is optimized, PBCAID incorporates a procedure for solvating the domain with water by filling the cell with a water lattice derived from an ice structure scaled to the bulk density of water. Results show that PBCAID can optimize system volumes by 20 to 70% and lead to computational savings in the nonbonded computations from reduced solvent sizes. Copyright 2001 John Wiley & Sons, Inc. J Comput Chem 22: 1843-1850, 2001 PMID- 12116416 TI - Prediction of infinite dilution activity coefficients of chlorinated organic compounds in aqueous solution from quantum-chemical descriptors. AB - A quantitative structure-property relationship (QSPR) model is developed to correlate the natural logarithm of infinite dilution activity coefficients, ln (gamma(infinity)), of 45 chlorinated organic compounds in aqueous solution from quantum-chemical descriptors. The best correlation equation contains five theoretical molecular descriptors. All descriptors were obtained from the chemical structure of the compounds and have definite physical meaning corresponding to different intermolecular interactions. The model predicts ln (gamma(infinity)) with a correlation coefficients of 0.949 and a standard error of 0.442 ln units. The obtained QSPR equation may be applied to the prediction of gamma(infinity) of other chlorinated organic compounds not present in the data set used for the development of the present model. Copyright 2001 John Wiley & Sons, Inc. J Comput Chem 22: 1851-1856, 2001 PMID- 12116417 TI - Effective atom volumes for implicit solvent models: comparison between Voronoi volumes and minimum fluctuation volumes. AB - An essential element of implicit solvent models, such as the generalized Born method, is a knowledge of the volume associated with the individual atoms of the solute. Two approaches for determining atomic volumes for the generalized Born model are described; one is based on Voronoi polyhedra and the other, on minimizing the fluctuations in the overall volume of the solute. Volumes to be used with various parameter sets for protein and nucleic acids in the CHARMM force field are determined from a large set of known structures. The volumes resulting from the two different approaches are compared with respect to various parameters, including the size and solvent accessibility of the structures from which they are determined. The question of whether to include hydrogens in the atomic representation of the solute volume is examined. Copyright 2001 John Wiley & Sons, Inc. J Comput Chem 22: 1857-1879, 2001 PMID- 12116418 TI - Seabird and louse coevolution: complex histories revealed by 12S rRNA sequences and reconciliation analyses. AB - We investigated the coevolutionary history of seabirds (orders Procellariiformes and Sphenisciformes) and their lice (order Phthiraptera). Independent trees were produced for the seabirds (tree derived from 12S ribosomal RNA, isoenzyme, and behavioral data) and their lice (trees derived from 12S rRNA data). Brook's parsimony analysis (BPA) supported a general history of cospeciation (consistency index = 0.84, retention index = 0.81). We inferred that the homoplasy in the BPA was caused by one intrahost speciation, one potential host-switching, and eight or nine sorting events. Using reconciliation analysis, we quantified the cost of fitting the louse tree onto the seabird tree. The reconciled trees postulated one host-switching, nine cospeciation, three or four intrahost speciation, and 11 to 14 sorting events. The number of cospeciation events was significantly more than would be expected from chance alone (P < 0.01). The sequence data were used to test for rate heterogeneity for both seabirds and lice. Neither data set displayed significant rate heterogeneity. An examination of the codivergent nodes revealed that seabirds and lice have cospeciated synchronously and that lice have evolved at approximately 5.5 times the rate of seabirds. The degree of sequence divergence supported some of the postulated intrahost speciation events (e.g., Halipeurus predated the evolution of their present hosts). The sequence data also supported some of the postulated host-switching events. These results demonstrate the value of sequence data and reconciliation analyses in unraveling complex histories between hosts and their parasites. PMID- 12116419 TI - Divergence and reticulation among montane populations of a jumping spider (Habronattus pugillis Griswold). AB - Populations of the jumping spider Habronattus pugillis Griswold isolated on nearby mountain ranges in southern Arizona are differentiated in many features of the males (color, shape, and orientation of setae on face; shape of carapace; markings of palpi and legs; motions during courtship behavior). These features are (mostly) consistent within a range and different between ranges. The concentration of differences in male courtship behavior and body parts exposed to the female during courtship and correlations between form and courtship behavior suggest sexual selection was involved in the differentiation. A phylogenetic analysis of the populations yields a tree that for the most part groups geographic neighbors, but the history of H. pugillis populations may not be adequately described by a tree. Geographic proximity of apparent convergences suggests that populations from at least some of the mountain ranges acquired characteristics through introgression. Lowering of the woodland habitat during the last glacial period probably brought some populations into contact, but it is not clear whether the interrange woodlands would have provided corridors for extensive mixing. PMID- 12116420 TI - Testing hybridization hypotheses based on incongruent gene trees. AB - Hybridization is an important evolutionary mechanism in plants and has been increasingly documented in animals. Difficulty in reconstruction of reticulate evolution, however, has been a long-standing problem in phylogenetics. Consequently, hybrid speciation may play a major role in causing topological incongruence between gene trees. The incongruence, in turn, offers an opportunity to detect hybrid speciation. Here we characterized certain distinctions between hybridization and other biological processes, including lineage sorting, paralogy, and lateral gene transfer, that are responsible for topological incongruence between gene trees. Consider two incongruent gene trees with three taxa, A, B, and C, where B is a sister group of A on gene tree 1 but a sister group of C on gene tree 2. With a theoretical model based on the molecular clock, we demonstrate that time of divergence of each gene between taxa A and C is nearly equal in the case of hybridization (B is a hybrid) or lateral gene transfer, but differs significantly in the case of lineage sorting or paralogy. After developing a bootstrap test to test these alternative hypotheses, we extended the model and test to account for incongruent gene trees with numerous taxa. Computer simulation studies supported the validity of the theoretical model and bootstrap test when each gene evolved at a constant rate. The computer simulation also suggested that the model remained valid as long as the rate heterogeneity was occurring proportionally in the same taxa for both genes. Although the model could not test hypotheses of hybridization versus lateral gene transfer as the cause of incongruence, these two processes may be distinguished by comparing phylogenies of multiple unlinked genes. PMID- 12116421 TI - The phylogenetic basis of sexual size dimorphism in orb-weaving spiders (Araneae, Orbiculariae). AB - Extreme sexual body size dimorphism (SSD), in which males are only a small fraction of the size of the females, occurs only in a few, mostly marine, taxonomic groups. Spiders are the only terrestrial group in which small males are relatively common, particularly among orb-weavers (especially in the families Tetragnathidae and Araneidae) and crab spiders (Thomisidae). We used a taxonomic sample of 80 genera to study the phylogenetic patterns (origins and reversals) of SSD in orb-weaving spiders (Orbiculariae). We collected and compiled male and female size data (adult body length) for 536 species. Size data were treated as a continuous character, and ancestral sizes, for males and females separately, were reconstructed by using Wagner parsimony on a cladogram for the 80 genera used in this study. Of these 80 genera, 24 were female-biased dimorphic (twice or more the body length of the male); the remaining 56 genera were monomorphic. Under parsimony only four independent origins of dimorphism are required: in the theridiid genus Tidarren, in the distal nephilines, in the "argiopoid clade," and in the araneid genus Kaira. Dimorphism has reversed to monomorphism at least seven times, all of them within the large "argiopoid clade." The four independent origins of dimorphism represent two separate instances of an increase in female size coupled with a decrease of male size (involving only two genera), and two separate instances of an increase in female size with male size either remaining the same or increasing, but not as much as females (involving 30 genera). In orb weaving spiders, far more taxa are sexually dimorphic as a result of female size increase (22 genera) than as a result of male size decrease (two genera). SSD in orb-weaving spiders encompasses several independent evolutionary histories that together suggest a variety of evolutionary pathways. This multiplicity strongly refutes all efforts thus far to find a general explanation for either the origin or maintenance (or both) of SSD, because the different pathways very likely will require distinctly different, possibly unique, explanations. Each pattern must be understood historically before its origin and maintenance can be explained in ecological and evolutionary terms. The most frequently cited example of male dwarfism in spiders, the golden orb-weaving spider genus Nephila (Tetragnathidae), is in fact a case of female giantism, not male dwarfism. PMID- 12116422 TI - Age rank/clade rank metrics--sampling, taxonomy, and the meaning of "stratigraphic consistency". AB - Paleontologists frequently contrast clade rank (i.e., nodal or patristic distance from the base of a cladogram) with age rank (i.e., relative first known appearances of the analyzed taxa) to measure the degree of congruence between the estimated phylogeny and the fossil record. Although some potential biases of these methods have been examined (e.g., the effect of tree imbalance), other properties of age rank/clade rank (ARCR) comparisons have not been studied in detail. A basic premise of ARCR metrics is that outgroup taxa diverged earlier than ingroups and thus should first appear in older strata. For example, given phylogeny (A,(B,C)), then taxon A should be sampled before either taxon B or taxon C. We examine this premise in the context of (1) phylogenetic theory, (2) taxonomic practice, (3) sampling intensity (R), and (4) factors other than sampling intensity (including cladogram accuracy). Simulations combining clade evolution and sampling over time indicate a poor relationship between ARCR metrics and R when all taxa are apomorphy-based monophyletic groups. However, a good relationship exists when taxa are either stem-based monophyletic groups or if workers include taxa without a priori decisions about monophyly or paraphyly. These results are not surprising because cladograms predict the order in which lineages diverged (which applies to stem-based monophyletic taxa) and the order in which morphologic grades appeared (which applies to paraphyletic taxa relative to derived monophyletic groups). Other factors that increase ARCR metrics when the average R stays the same include high temporal variation in R, budding instead of bifurcating speciation patterns, low extinction rates, cladogram inaccuracy, and (to a much lesser extent) large clade size. These results suggest several plausible explanations for patterned differences in ARCR metrics among clades, thereby compromising their validity as measures of the quality of the fossil record. PMID- 12116423 TI - Taxic homology and three-taxon statement analysis. AB - Three-taxon statement analysis (3TA) and standard cladistic analysis (SCA) were evaluated relative to propositions of taxic homology. There are definite distinctions between complement relation homologs and paired homologs. The complement relation is discussed, relative to rooting, parsimony, and taxic propositions of homology. The complement relation, as implemented in SCA, makes sense only because SCA is a simple evolutionary model of character-state transformation. 3TA is a method for implementing complement relation data from a taxic perspective. The standard approach to cladistic analysis distinguishes taxa by rooting a tree, which means that that approach is incompatible with taxic propositions of homology, because a taxic homology is a hypothesis of relationship between taxa that possess a homolog relative to taxa that lack a homolog. It is not necessary to treat paired homologs from a transformational perspective to distinguish informative from uninformative data. 3TA yields results markedly different from those of SCA. SCA, which seeks to minimize tree length, may not maximize the relation of homology (congruence) relative to a tree. PMID- 12116424 TI - c-myc gene sequences and the phylogeny of bats and other eutherian mammals. AB - The complete protein-coding sequences of the c-myc proto-oncogene were determined for five species of four new orders of eutherian (placental) mammals. These newly obtained sequences were aligned to each other and to other available orthologs for the phylogenetic estimation of eutherian interordinal relationships. Several measures of sequence difference and base composition were first calculated to assess the major evolutionary properties of the three codon positions and two protein-coding exons of the gene. On the basis of these calculations, different parsimony, distance, and maximum likelihood approaches were adopted, with the most sophisticated involving the separate, then combined, likelihood analyses of the third codon positions of exon 2 versus all other sites. These phylogenetic approaches provided clear support for the grouping of Chiroptera (bats) with Artiodactyla (ruminants, camels, and pigs) and Carnivora (cats, dogs, and their allies), an interordinal arrangement that receives strong corroboration from other lines of evidence including complete mitochondrial DNA sequences. In contrast, these analyses failed to provide strong to reasonable support for any other interordinal group. This study concludes with specific recommendations about sampling and other strategies for maximizing the phylogenetic contributions of the c-myc gene to the continued resolution of the eutherian ordinal tree. PMID- 12116425 TI - Hidden morphological support for the phylogenetic placement of Pseudoryx nghetinhensis with bovine bovids: a combined analysis of gross anatomical evidence and DNA sequences from five genes. AB - The saola, Pseudoryx nghetinhensis, was unknown to science until its formal description in 1993. This endangered species is a member of the ruminant artiodactyl family Bovidae (cattle, sheep, goats, and antelopes). However, given its puzzling combination of morphological traits, the specific affinities of Pseudoryx within Bovidae are controversial. A preliminary genetic investigation suggested that Pseudoryx should be placed in the subfamily Bovinae (cattle, buffaloes, spiral-horned antelopes, and nilgai), but a recent cladistic analysis of skeletal and dental characters allied Pseudoryx with caprine bovids (sheep, goats, musk oxen, goat antelopes, and Pantholops). The morphological and molecular hypotheses differ in assigning the saola to either of the two most divergent clades of Bovidae. In this report, phylogenetic analyses of DNA sequences from five genes are used to test these alternatives. Protein coding regions, introns, and ribosomal DNAs from the nuclear and mitochondrial genomes discount the hypothesis that Pseudoryx is a close relative of Caprinae. Instead, combined analyses of the DNA data and published morphological evidence place Pseudoryx with Bovini (cattle and buffaloes), a subclade of Bovinae. In a separate analysis, the matrix of morphological characters links Pseudoryx with caprine bovids, but in the context of the molecular data, the gross anatomical evidence strongly supports a grouping of Pseudoryx with Bovinae. Surprisingly, the morphological partition provides the most character support in the combined analysis. This striking result is obscured by separate analyses of the individual data sets and the taxonomic congruence approach. PMID- 12116426 TI - Triploblastic relationships with emphasis on the acoelomates and the position of Gnathostomulida, Cycliophora, Plathelminthes, and Chaetognatha: a combined approach of 18S rDNA sequences and morphology. AB - Triploblastic relationships were examined in the light of molecular and morphological evidence. Representatives for all triploblastic "phyla" (except Loricifera) were represented by both sources of phylogenetic data. The 18S ribosomal (rDNA) sequence data for 145 terminal taxa and 276 morphological characters coded for 36 supraspecific taxa were combined in a total evidence regime to determine the most consistent picture of triploblastic relationships for these data. Only triploblastic taxa are used to avoid rooting with distant outgroups, which seems to happen because of the extreme distance that separates diploblastic from triploblastic taxa according to the 18S rDNA data. Multiple phylogenetic analyses performed with variable analysis parameters yield largely inconsistent results for certain groups such as Chaetognatha, Acoela, and Nemertodermatida. A normalized incongruence length metric is used to assay the relative merit of the multiple analyses. The combined analysis having the least character incongruence yields the following scheme of relationships of four main clades: (1) Deuterostomia [((Echinodermata + Enteropneusta) (Cephalochordata (Urochordata + Vertebrata)))]; (2) Ecdysozoa [(((Priapulida + Kinorhyncha) (Nematoda + Nematomorpha)) ((Onychophora + Tardigrada) Arthropoda))]; (3) Trochozoa [((Phoronida + Brachiopoda) (Entoprocta (Nemertea (Sipuncula (Mollusca (Pogonophora (Echiura + Annelida)))))))]; and (4) Platyzoa [((Gnathostomulida (Cycliophora + Syndermata)) (Gastrotricha + Plathelminthes))]. Chaetognatha, Nemertodermatida, and Bryozoa cannot be assigned to any one of these four groups. For the first time, a data analysis recognizes a clade of acoelomates, the Platyzoa (sensu Cavalier-Smith, Biol. Rev. 73:203-266, 1998). Other relationships that corroborate some morphological analyses are the existence of a clade that groups Gnathostomulida + Syndermata (= Gnathifera), which is expanded to include the enigmatic phylum Cycliophora, as sister group to Syndermata. PMID- 12116427 TI - Variation in mandible shape in Thrichomys apereoides (Mammalia: Rodentia): geometric analysis of a complex morphological structure. AB - The model of development and evolution of complex morphological structures conceived by Atchley and Hall in 1991 (Biol. Rev. 66:101-157), which establishes that changes at the macroscopic, morphogenetic level can be statistically detected as variation in skeletal units at distinct scales, was applied in combination with the formalism of geometric morphometrics to study variation in mandible shape among populations of the rodent species Thrichomys apereoides. The thin-plate spline technique produced geometric descriptors of shape derived from anatomical landmarks in the mandible, which we used with graphical and inferential approaches to partition the contribution of global and localized components to the observed differentiation in mandible shape. A major pattern of morphological differentiation in T. apereoides is attributable to localized components of shape at smaller geometric scales associated with specific morphogenetic units of the mandible. On the other hand, a clinical trend of variation is associated primarily with localized components of shape at larger geometric scales. Morphogenetic mechanisms assumed to be operating to produce the observed differentiation in the specific units of the mandible include mesenchymal condensation differentiation, muscle hypertrophy, and tooth growth. Perspectives for the application of models of morphological evolution and geometric morphometrics to morphologically based systematic biology are considered. PMID- 12116428 TI - Timing transantarctic disjunctions in the Atherospermataceae (Laurales): evidence from coding and noncoding chloroplast sequences. AB - Previous studies of the small Southern Hemisphere family Atherospermataceae have drawn contradictory conclusions regarding the number of transantarctic disjunctions and role of transoceanic dispersal in its evolution. Clarification of intergeneric relationships is critical to resolving (1) whether the two Chilean species, Laurelia sempervirens and Laureliopsis philippiana, are related to different Austral-Pacific species, implying two transantarctic disjunctions as suggested by morphology; (2) where the group is likely to have originated; and (3) whether observed disjunctions reflect the breakup of Gondwana. We analyzed chloroplast DNA sequences from six regions (the rbcL gene, the rpl16 intron, and the trnL-trnF, trnT-trnL, psbA-trnH, and atpB-rbcL spacer regions; for all six regions, 4,372 bp) for all genera and most species of Atherospermataceae, using parsimony and maximum likelihood (ML). The family's sister group, the Chilean endemic Gomortega nitida (Gomortegaceae), was used to root the tree. Parsimony and ML yielded identical single best trees that contain three well-supported clades (> or = 75% bootstrap): Daphnandra and Doryphora from south-eastern Australia; Atherosperma and Nemuaron from Australia-Tasmania and New Caledonia, respectively; and Laurelia novac-zelandiac and Laureliopsis philippiana from New Zealand and Chile, respectively. The second Chilean species, Laurelia sempervirens, is sister to this last clade. Likelihood ratio testing did not reject the molecular clock assumption for the rbcL data, which can therefore be used for divergence time estimates. The atherosperm fossil record, which goes back to the Upper Cretaceous, includes pollen, wood, and leaf fossils from Europe, Africa, South America, Antarctica, New Zealand, and Tasmania. Calibration of rbcL substitution rates with the fossils suggests an initial diversification of the family at 100-140 million years ago (MYA), probably in West Gondwana, early entry into Antarctica, and long-distance dispersal to New Zealand and New Caledonia at 50-30 MYA by the ancestors of L. novae-zelandiae and Nemuaron. PMID- 12116430 TI - Quantification of homoplasy for nucleotide transitions and transversions and a reexamination of assumptions in weighted phylogenetic analysis. AB - Nucleotide transitions are frequently down-weighted relative to transversions in phylogenetic analysis. This is based on the assumption that transitions, by virtue of their greater evolutionary rate, exhibit relatively more homoplasy and are therefore less reliable phylogenetic characters. Relative amounts of homoplastic and consistent transition and transversion changes in mitochondrial protein coding genes were determined from character-state reconstructions on a highly corroborated phylogeny of mammals. We found that although homoplasy was related to evolutionary rates and was greater for transitions, the absolute number of consistent transitions greatly exceeded the number of consistent transversions. Consequently, transitions provided substantially more useful phylogenetic information than transversions. These results suggest that down weighting transitions may be unwarranted in many cases. This conclusion was supported by the fact that a range of transition: transversion weighting schemes applied to various mitochondrial genes and genomic partitions rarely provided improvement in phylogenetic estimates relative to equal weighting, and in some cases weighting transitions more heavily than transversions was most effective. PMID- 12116429 TI - Phylogenetic relationships among the phrynosomatid sand lizards inferred from mitochondrial DNA sequences generated by heterogeneous evolutionary processes. AB - Nucleotide sequences of the mitochondrial protein coding cytochrome b (cyt b; 650 bp) and small-subunit 12S ribosomal RNA (approximately 350 bp) genes were used in analyses of phylogenetic relationships among extant phrynosomatid sand lizards, including an examination of competing hypotheses regarding the evolution of "earlessness." Sequences were obtained from all currently recognized species of sand lizards as well as representatives of the first and second outgroups and analyzed using both parsimony and likelihood methods. The cyt b data offer strong support for relationships that correspond with relatively recent divergences and moderate to low support for relationships reflecting more ancient divergences within the clade. These data support monophyly of the "earless" taxa, the placement of Uma as the sister taxon to the other sand lizards, and monophyly of all four taxa traditionally ranked as genera. All well-supported relationships in the 12S phylogeny are completely congruent with well-supported relationships in the cyt b phylogeny; however, the 12S data alone provide very little support for deeper divergences. Phylogenetic relationships within species are concordant with geography and suggest patterns of phylogeographic differentiation, including the conclusion that at least one currently recognized species (Holbrookia maculata) actually consists of more than one species. By independently optimizing likelihood model parameters for various subsets of the data, we found that nucleotide substitution processes vary widely between genes and among the structural and functional regions or classes of sites within each gene. Therefore, we compared competing phylogenetic hypotheses, using parameter estimates specific to those subsets, analyzing the subsets separately and in various combinations. The hypothesis supported by the cyt b data was favored over rival hypotheses in all but one of the five comparisons made with the entire data set, including the set of partitions that best explained the data, although we were unable to confidently reject (P < 0.05) alternative hypotheses. Our results highlight the importance of optimizing models and parameter estimates for different genes or parts thereof--a strategy that takes advantages of the strengths of both combining and partitioning data. PMID- 12116431 TI - Integrating ambiguously aligned regions of DNA sequences in phylogenetic analyses without violating positional homology. AB - Phylogenetic analyses of non-protein-coding nucleotide sequences such as ribosomal RNA genes, internal transcribed spacers, and introns are often impeded by regions of the alignments that are ambiguously aligned. These regions are characterized by the presence of gaps and their uncertain positions, no matter which optimization criteria are used. This problem is particularly acute in large scale phylogenetic studies and when aligning highly diverged sequences. Accommodating these regions, where positional homology is likely to be violated, in phylogenetic analyses has been dealt with very differently by molecular systematists and evolutionists, ranging from the total exclusion of these regions to the inclusion of every position regardless of ambiguity in the alignment. We present a new method that allows the inclusion of ambiguously aligned regions without violating homology. In this three-step procedure, first homologous regions of the alignment containing ambiguously aligned sequences are delimited. Second, each ambiguously aligned region is unequivocally coded as a new character, replacing its respective ambiguous region. Third, each of the coded characters is subjected to a specific step matrix to account for the differential number of changes (summing substitutions and indels) needed to transform one sequence to another. The optimal number of steps included in the step matrix is the one derived from the pairwise alignment with the greatest similarity and the least number of steps. In addition to potentially enhancing phylogenetic resolution and support, by integrating previously nonaccessible characters without violating positional homology, this new approach can improve branch length estimations when using parsimony. PMID- 12116432 TI - Likelihood-based tests of topologies in phylogenetics. AB - Likelihood-based statistical tests of competing evolutionary hypotheses (tree topologies) have been available for approximately a decade. By far the most commonly used is the Kishino-Hasegawa test. However, the assumptions that have to be made to ensure the validity of the Kishino-Hasegawa test place important restrictions on its applicability. In particular, it is only valid when the topologies being compared are specified a priori. Unfortunately, this means that the Kishino-Hasegawa test may be severely biased in many cases in which it is now commonly used: for example, in any case in which one of the competing topologies has been selected for testing because it is the maximum likelihood topology for the data set at hand. We review the theory of the Kishino-Hasegawa test and contend that for the majority of popular applications this test should not be used. Previously published results from invalid applications of the Kishino Hasegawa test should be treated extremely cautiously, and future applications should use appropriate alternative tests instead. We review such alternative tests, both nonparametric and parametric, and give two examples which illustrate the importance of our contentions. PMID- 12116433 TI - Improved bootstrap confidence limits in large-scale phylogenies, with an example from Neo-Astragalus (Leguminosae). AB - Phylogenetic analyses of large data sets pose special challenges, including the apparent tendency for the bootstrap support for a clade to decline with increased taxon sampling of that clade. We document this decline in data sets with increasing numbers of taxa in Astragalus, the most species-rich angiosperm genus. Support for one subclade, Neo-Astragalus, declined monotonically with increased sampling of taxa inside Neo-Astragalus, irrespective of whether parsimony or neighbor-joining methods were used or of which particular heuristic search algorithm was used (although more stringent algorithms tended to yield higher support). Three possible explanations for this decline were examined, including (1) mistaken assignment of the most recent common ancestor of the taxon sample (and its bootstrap support) with the most recent common ancestor of the clade from which it was sampled; (2) computational limitations of heuristic search strategies; and (3) statistical bias in bootstrap proportions, especially that from random homoplasy distributed among taxa. The best explanation appears to be (3), although computational shortcomings (2) may explain some of the problem. The bootstrap proportion, as currently used in phylogenetic analysis, does not accurately capture the classical notion of confidence assessments on the null hypothesis of nonmonophyly, especially in large data sets. More accurate assessments of confidence as type I error levels (relying on iterated bootstrap methods) remove most of the monotonic decline in confidence with increasing numbers of taxa. PMID- 12116434 TI - Ability of geometric morphometric methods to estimate a known covariance matrix. AB - Landmark-based morphometric methods must estimate the amounts of translation, rotation, and scaling (or, nuisance) parameters to remove nonshape variation from a set of digitized figures. Errors in estimates of these nuisance variables will be reflected in the covariance structure of the coordinates, such as the residuals from a superimposition, or any linear combination of the coordinates, such as the partial warp and standard uniform scores. A simulation experiment was used to compare the ability of the generalized resistant fit (GRF) and a relative warp analysis (RWA) to estimate known covariance matrices with various correlations and variance structures. Random covariance matrices were perturbed so as to vary the magnitude of the average correlation among coordinates, the number of landmarks with excessive variance, and the magnitude of the excessive variance. The covariance structure was applied to random figures with between 6 and 20 landmarks. The results show the expected performance of GRF and RWA across a broad spectrum of conditions. The performance of both GRF and RWA depended most strongly on the number of landmarks. RWA performance decreased slightly when one or a few landmarks had excessive variance. GRF performance peaked when approximately 25% of the landmarks had excessive variance. In general, both RWA and GRF performed better at estimating the direction of the first principal axis of the covariance matrix than the structure of the entire covariance matrix. RWA tended to outperform GRF when > approximately 75% of the coordinates had excessive variance. When < 75% of the coordinates had excessive variance, the relative performance of RWA and GRF depended on the magnitude of the excessive variance; when the landmarks with excessive variance had standard deviations (sigma) > or = 4 sigma minimum, GRF regularly outperformed RWA. PMID- 12116435 TI - A phylogenetic perspective on habitat shifts and diversity in the North American Enallagma damselflies. AB - Community ecologists are increasingly aware that the regional history of taxon diversification can have an important influence on community structure. Likewise, systematists recognize that ecological context can have an important influence on the processes of speciation and extinction that create patterns of descent. We present a phylogenetic analysis of 33 species of a North American radiation of damselflies (Zygoptera: Coenagrionidae: Enallagma Selys), which have been well studied ecologically, to elucidate the evolutionary mechanisms that have contributed to differences in diversity between larval habitats (lakes with and without fish predators). Analysis of molecular variation in 842 bp of the mitochondrial cytochrome oxidase I and II subunit and of the intervening Leu-tRNA and 37 morphological characters resulted in three well-defined clades that are only partially congruent with previous phylogenetic hypotheses. Molecular and morphological data partitions were significantly incongruent (p < .01). Lack of haplotype monophyly within species and small amounts of sequence divergence (< 1%) between related species in three of the four clades suggest that recent, and parallel, speciation has been an important source of community diversity. Reconstruction of habitat preference over the phylogeny suggests that the greater species diversity in fish-containing lake habitats reflects the recency of shifts into the fishless lake habit, although a difference in speciation or extinction rates between the two habitats is difficult to exclude as an additional mechanism. PMID- 12116436 TI - Multiple data sets, congruence, and hypothesis testing for the phylogeny of basal groups of the lizard genus Sceloporus (Squamata, Phrynosomatidae). AB - Several data partitions, including nuclear and mitochondrial gene sequences, chromosomes, isoenzymes, and morphological characters, were used to propose a new phylogeny and to test previously published hypotheses about the phylogenetic positions of basal clades of the lizard genus Sceloporus and the relationship of Sceloporus to the former genus "Sator". In accord with earlier studies, our results grouped "Sator" as internal to Sceloporus, and both support a hypothesis of transgulfian vicariance for the origin of the former genus "Sator" on islands in the Sea of Cortez. Robustness of support for internal nodes in our best tree was established though widely used indices (bootstrap proportions, decay values) but also through congruence among independent data partitions. Several deep nodes in the tree recovered by several methods, including equally weighted and differentially weighted parsimony and maximum likelihood models, are only weakly supported by the traditional indices. This methodological concordance is taken as evidence for insensitivity of the deep structure of the topology to alternative assumptions. PMID- 12116437 TI - Use of two-block partial least-squares to study covariation in shape. AB - The relatively new two-block partial least-squares method for analyzing the covariance between two sets of variables is described and contrasted with the well-known method of canonical correlation analysis. Their statistical properties, type of answers, and visualization techniques are discussed. Examples are given to show its usefulness in comparing two sets of variables--especially when one or both of the sets of variables are shape variables from a geometric morphometric study. PMID- 12116438 TI - A chain is no stronger than its weakest link: double decay analysis of phylogenetic hypotheses. AB - In decay analyses the support for a particular split in most-parsimonious trees is its decay index, that is, the extra steps required of the shortest trees that do not include the split. By focusing solely on the support for splits, traditional decay analysis may provide an incomplete and potentially misleading summary of the support for phylogenetic relationships common to the most parsimonious tree or trees. Here, we introduce double decay analysis, a new approach to assessing support for phylogenetic relationships. Double decay analysis is the determination of the decay indices of all n-taxon statements/partitions common to the most-parsimonious tree. The results of double decay analyses are presented in a partition table, but various approaches to graphical representation of the results, including the use of reduced consensus support trees, are also discussed. Double decay analysis provides a more comprehensive summary and facilitates a better understanding of the strengths and weaknesses of complex phylogenetic hypotheses than does traditional decay analysis. The limitations of traditional decay analyses and the utility of double decay analyses are illustrated with both contrived data and real data for sauropod dinosaurs. PMID- 12116439 TI - Molecular phylogenetics and biogeography of galaxiid fishes (Osteichthyes: Galaxiidae): dispersal, vicariance, and the position of Lepidogalaxias salamandroides. AB - The galaxiid fishes exhibit a gondwanan distribution. We use mitochondrial DNA sequences to test conflicting vicariant and dispersal biogeographic hypotheses regarding the Southern Hemisphere range of this freshwater group. Although phylogenetic resolution of cytochrome b and 16S rRNA sequences is largely limited to more recent divergences, our data indicate that the radiation can be interpreted as several relatively recent dispersal events superimposed on an ancient gondwanan radiation. Genetic relationships contradict the findings of recent morphological analyses of galaxioid fishes. In particular, we examine several hypotheses regarding phylogenetic placement of the enigmatic Lepidogalaxias. Although most workers consider Lepidogalaxias to be an unusual scaled member of the Southern Hemisphere galaxioids, it has also been suggested that this species is related to the Northern Hemisphere esocoids. Our data strongly suggest that this species is not a galaxiid, and the alternative hypothesized esocoid relationship cannot be rejected. The species-rich genus Galaxias is shown to be polyphyletic and the generic taxonomy of the Galaxiinae is reassessed in the light of phylogenetic relationships. Juvenile saltwater tolerance is phylogenetically distributed throughout the Galaxiinae, and the loss of this migratory phase may be a major cause of speciation. PMID- 12116440 TI - Why morphometrics is special: the problem with using partial warps as characters for phylogenetic inference. PMID- 12116441 TI - Linked branch support and tree stability. PMID- 12116442 TI - SINE evolution, missing data, and the origin of whales. PMID- 12116443 TI - Parametric phylogenetics? PMID- 12116444 TI - Tree robustness and clade significance. PMID- 12116445 TI - [Do corticosteroids have a place in the treatment of chronic obstructive pulmonary disease?]. PMID- 12116446 TI - [Drug therapy of Crohn disease]. PMID- 12116447 TI - [Can the postoperative relapse of Crohn disease be influenced?]. PMID- 12116449 TI - [Halitosis]. PMID- 12116448 TI - [Itching legs and stiff neck]. PMID- 12116450 TI - [Altarpiece on the pilgrimage theme]. PMID- 12116451 TI - [Are external forces needed when preparing text books, and who owns the pictures?]. PMID- 12116452 TI - [Paresthesia and paresis symptoms of lower extremities]. PMID- 12116454 TI - [Eating disorders and traumatic experiences]. PMID- 12116453 TI - [Eating disorders--an increasing problem in our culture?]. PMID- 12116455 TI - [Viewpoints about prevalence and risk factors of anorexia and bulimia nervosa]. PMID- 12116456 TI - [An eating disorder or not?]. PMID- 12116457 TI - [Treatment of bulimia]. PMID- 12116458 TI - [Treatment of anorexia nervosa]. PMID- 12116459 TI - [Guidelines for the treatment of lung cancer]. PMID- 12116460 TI - [Use of clodronate in the adjuvant treatment of breast cancer is questionable for the time being]. PMID- 12116461 TI - [Use of immunoglobulin in the therapy of neurological illnesses]. PMID- 12116462 TI - [Results from telemedicine projects]. PMID- 12116463 TI - [Lymphomatoid papulosis--a sheep in wolves' clothes]. PMID- 12116464 TI - [Use of blood-patch in the treatment of pneumothorax]. PMID- 12116465 TI - [Viruses as helpful instruments in medicine]. PMID- 12116466 TI - [Outi Heiskanen: The doctor arrives]. PMID- 12116467 TI - [Acute anuria in a patient with an urinary catheter]. PMID- 12116468 TI - [Weapons of our health care system to react when children and adolescents have problems]. PMID- 12116469 TI - [Children of our times--sleepless but tired during the daytime]. PMID- 12116470 TI - [Preventing infections in the day-care centers]. PMID- 12116471 TI - [Has pertussis been tamed?]. PMID- 12116472 TI - [Why does a child have stomach ache?]. PMID- 12116473 TI - [Infant depression]. PMID- 12116474 TI - Is there a justification for differential a priori weighting in coding sequences? A case study from rbcL and Apocynaceae s.l. AB - Functional constraints are often assumed to influence the performance of nucleotide characters in phylogenetic analysis: First and second codon positions and sites of structural importance are considered to show less homoplasy. We investigate the performance of rbcL characters with differential functional constraints in a cladistic analysis of the plant family Apocynaceae s.l. (Sennblad and Bremer, in prep.). Performance is measured as rescaled consistency indices (rc). We show there is no significant difference in performance between parsimony-informative sites constrained by function in the enzyme, and sites that are not. Furthermore, the substitutions in third-codon position performed significantly better than those in first and second. The variation of rc within the different classes was high, however. Consequently, there is no support for routinely applied a priori differential weighting, neither of codon positions, nor of different functional classes from the present analysis of rbcL data in the Apocynaceae s.l. PMID- 12116475 TI - Mitochondrial phylogeny of notothenioids: a molecular approach to Antarctic fish evolution and biogeography. AB - Antarctic waters represent a unique marine environment delimited by an oceanographic barrier, the Polar Front Zone, and characterized by constant subzero temperatures and presence of sea ice. A group of teleost fish, the Notothenioidei, have adapted to these challenging environmental conditions, undergoing a remarkable diversification. In the present study a total of 798 base pairs, generated from partial sequencing of 16S and 12S mitochondrial ribosomal RNA genes, were examined in 33 notothenioid species representative of all families included in the suborder Notothenioidei. Phylogenetic trees, reconstructed on the basis of sequence data by different methods, indicate that traditional hypotheses on notothenioid systematics and biogeography might be in need of reexamination. Molecular evidence suggests that vicariant speciation could be invoked to explain the early divergence of Eleginops maclovinus, a species previously included in the family Nototheniidae, which is now proposed as the closest sister group to all the rest of notothenioids apart from bovichtids. On the other hand, repeated, independent dispersal through the Polar Front is proposed for the divergence of other subantarctic notothenioid species. Likewise, multiple, independent transitions from benthic to pelagic habit are inferred from molecular data, at variance with the more conservative hypothesis based on cladograms reconstructed from morphological data. PMID- 12116476 TI - Early evolution of the bilateria. AB - The phylogeny of the Bilateria and especially the early steps in the evolution of the bilaterian bauplan are still a controversial topic. In this context the relationships of the platyhelminths and the nematodes play a crucial role. Previous molecular studies of the relationships of these groups, which were based on 18S ribosomal DNA sequences, yielded conflicting results. In the present study a new framework is developed for the phylogenetic analysis of bilaterian relationships, using concatenated amino acid sequences of several nuclear genes. In this analysis, the rhabditophoran platyhelminths are probably the sister group of all other analyzed Bilateria, the Eubilateria, which are characterized by a one-way intestine with an anus. The Eubilateria are split into the nematode lineage and the coelomates. The phylogenetic results of the present study indicate that genetic features found in the model organisms Caenorhabditis and Drosophila might be found in all Eubilateria. Estimations of the divergence times show that the major bilaterian phyla did not originate in an explosive radiation during the Cambrian but rather that the Bilateria have a several hundred million years long Precambrian history. PMID- 12116477 TI - War of the Iguanas: conflicting molecular and morphological phylogenies and long branch attraction in iguanid lizards. AB - Recent studies based on different types of data (i.e., morphology, molecules) have found strongly conflicting phylogenies for the genera of iguanid lizards but have been unable to explain the basis for this incongruence. We reanalyze published data from morphology and from the mitochondrial ND4, cytochrome b, 12S, and 16S genes to explore the sources of incongruence and resolve these conflicts. Much of the incongruence centers on the genus Cyclura, which is the sister taxon of Iguana, according to parsimony analyses of the morphology and the ribosomal genes, but is the sister taxon of all other Iguanini, according to the protein coding genes. Maximum likelihood analyses show that there has been an increase in the rate of nucleotide substitution in Cyclura in the two protein-coding genes (ND4 and cytochrome b), although this increase is not as clear when parsimony is used to estimate branch lengths. Parametric simulations suggest that Cyclura may be misplaced by the protein-coding genes as a result of long-branch attraction; even when Cyclura and Iguana are sister taxa in a simulated phylogeny, Cyclura is still placed as the basal member of the Iguanini by parsimony analysis in 55% of the replicates. A similar long-branch attraction problem may also exist in the morphological data with regard to the placement of Sauromalus with the Galapagos iguanas (Amblyrhynchus and Conolophus). The results have many implications for the analysis of diverse data sets, the impact of long branches on parsimony and likelihood methods, and the use of certain protein-coding genes in phylogeny reconstruction. PMID- 12116478 TI - Comparison of three methods for estimating internal support on phylogenetic trees. PMID- 12116479 TI - A simulation study of reduced tree-search effort in bootstrap resampling analysis. PMID- 12116480 TI - Transformationalism, taxism, and developmental biology in systematics. AB - Issues concerning transformational and taxic comparisons are central to understanding the impact of the recent proliferation of molecular developmental data on evolutionary biology. More importantly, an understanding of taxism and transformationalism in comparative biology is critical to assessing the impact of the recent developmental data on systematic theory and practice. We examine the philosophical and practical aspects of the transformational approach and the relevance of this approach to recent molecular-based developmental data. We also examine the theoretical basis of the taxic approach to molecular developmental data and suggest that developmental data are perfectly amenable to the taxic approach. Two recent examples from the molecular developmental biology literature -the evolution of insect wings and the evolution of dorsal ventral inversion in vertebrates and invertebrates--are used to compare the taxic and transformational approaches. We conclude that the transformational approach is entirely appropriate for ontogenetic studies and furthermore can serve as an excellent source of hypotheses about the evolution of characters. However, the taxic approach is the ultimate arbiter of these hypotheses. PMID- 12116481 TI - Developmental regulatory genes and echinoderm evolution. AB - Modified interactions among developmental regulatory genes and changes in their expression domains are likely to be an important part of the developmental basis for evolutionary changes in morphology. Although developmental regulatory genes are now being studied in an increasing number of taxa, there has been little attempt to analyze the resulting data within an explicit phylogenetic context. Here we present comparative analyses of expression data from regulatory genes in the phylum Echinodermata, considering the implications for understanding both echinoderm evolution as well as the evolution of regulatory genes in general. Reconstructing the independent evolutionary histories of regulatory genes, their expression domains, their developmental roles, and the structures in which they are expressed reveals a number of distinct evolutionary patterns. A few of these patterns correspond to interpretations common in the literature, whereas others have received little prior mention. Together, the analyses indicate that the evolution of echinoderms involved: (1) the appearance of many apomorphic developmental roles and expression domains, some of which have plesiomorphic bilateral symmetry and others of which have apomorphic radial symmetry or left right asymmetry; (2) the loss of some developmental roles and expression domains thought to be plesiomorphic for Bilateria; and (3) the retention of some developmental roles thought to be plesiomorphic for Bilateria, although with modification in expression domains. Some of the modifications within the Echinodermata concern adult structures; others, transient larval structures. Some changes apparently appeared early in echinoderm evolution (> 450 Ma), whereas others probably happened more recently (< 50 Ma). Cases of likely convergence in expression domains suggest caution when using developmental regulatory genes to make inferences about homology among morphological structures of distantly related taxa. PMID- 12116482 TI - Developmental variation, homology, and the pharyngula stage. AB - Understanding how development varies both inter- and intraspecifically can be important for systematic and evolutionary studies. This review will explore three different ways such understanding can be applied to evolutionary analyses. First, developmental data can be useful for homology determination. Interspecific variation in development has been thought to make developmental data poor candidates for determining homology. However, an updated developmental criterion that is more broadly comparative and mechanistic augments the available criteria used in homology determination. Second, modern cell and molecular biology are providing a better understanding of the many developmental processes involved in a structure's formation and will augment the number of characters available for phylogenetic analyses. Recent work has revealed that what had been thought to be a highly conserved developmental stage, the pharyngula (the phylotypic and zootypic stage of craniates) is highly variable. This variation can be seen in the development of such tissues as neural crest and placodes. These tissues are particularly interesting from a phylogenetic standpoint because they and the structures they form contribute to key synapomorphies of craniates. Finally, understanding developmental processes and how they form the variety of morphologies seen in nature will help in constructing the transformations that occurred during evolution. One such example involves descriptions of how lateral line development is affected in different mutant lines of zebrafish. The many species of teleost fishes express great variation in the patterns of their lateral lines, and this is often an important systematic character. Understanding the genetic basis of lateral line development would help not only in hypothesizing possible transformational series but also in determining how many genes may have been required for these transformations. PMID- 12116484 TI - The quality of the fossil record and the accuracy of phylogenetic inferences about sampling and diversity. AB - Because phylogenies can be estimated without stratigraphic data and because estimated phylogenies also infer gaps in sampling, some workers have used phylogeny estimates as templates for evaluating sampling from the fossil record and for "correcting" historical diversity patterns. However, it is not known how sampling intensity (the probability of sampling taxa per unit time) and completeness (the proportion of taxa sampled) affect the accuracy of phylogenetic inferences, nor how phylogenetically inferred estimates of sampling and diversity respond to inaccurate estimates of phylogeny. Both issues are addressed with a series of simulations using simple models of character evolution, varying speciation patterns, and various rates of speciation, extinction, character change, and preservation. Parsimony estimates of simulated phylogenies become less accurate as sampling decreases, and inaccurate trees chronically underestimate sampling. Biotic factors such as rates of morphologic change and extinction both affect the accuracy of phylogenetic estimates and thus affect estimated gaps in sampling, indicating that differences in implied sampling need not reflect actual differences in sampling. Errors in inferred diversity are concentrated early in the history of a clade. This, coupled with failure to account for true extinction times (i.e., the Signor-Lipps effect), inflates relative diversity levels early in clade histories. Because factors other than differences in sampling predict differences in the numbers of gaps implied by phylogeny estimates, inferred phylogenies can be misleading templates for evaluating sampling or historical diversity patterns. PMID- 12116483 TI - Urochordates are monophyletic within the deuterostomes. AB - Understanding the phylogenetic relationships of the three major urochordate groups within the deuterostomes is central to understanding the evolution of the chordates. We have prepared a detailed phylogenetic analysis of urochordates based on comparisons of 10 new urochordate 18S ribosomal DNA sequences with other urochordate sequences in GenBank. Maximum parsimony, neighbor-joining, minimum evolution, and maximum likelihood analyses of this large urochordate data set are consistent with a topology in which the urochordates are monophyletic within the deuterostomes and there are four separate clades of urochordates. These four distinct clades--styelid + pyurid ascidians, molgulid ascidians, phlebobranch ascidians + thaliaceans, and larvaceans--are mostly consistent with traditional morphological hypotheses and classifications. However, we find that the ascidians may not be a monophyletic group (as they have been considered traditionally) but instead appear paraphyletic. Another disparity with traditional classification is that the thaliaceans do not form a separate urochordate clade but rather cluster with the phlebobranch ascidians. Larvaceans have long branch lengths, which can be problematic for molecular phylogenetic methods, and their position within the urochordates cannot be unequivocally determined with 18S rDNA. This is important because the tadpole morphology of larvacean and ascidian larvae is the key trait of interest that distinguishes urochordates as chordates. Nevertheless, the present data set resolves at least three clades of urochordates and suggests strongly that urochordates form a monophyletic clade within the deuterostomes. PMID- 12116485 TI - Phylogenetic relationship among horseshoe crab species: effect of substitution models on phylogenetic analyses. AB - The horseshoe crabs, known as living fossils, have maintained their morphology almost unchanged for the past 150 million years. The little morphological differentiation among horseshoe crab lineages has resulted in substantial controversy concerning the phylogenetic relationship among the extant species of horseshoe crabs, especially among the three species in the Indo-Pacific region. Previous studies suggest that the three species constitute a phylogenetically unresolvable trichotomy, the result of a cladogenetic process leading to the formation of all three Indo-Pacific species in a short geological time. Data from two mitochondrial genes (for 16S ribosomal rRNA and cytochrome oxidase subunit I) and one nuclear gene (for coagulogen) in the four species of horseshoe crabs and outgroup species were used in a phylogenetic analysis with various substitution models. All three genes yield the same tree topology, with Tachypleus-gigas and Carcinoscorpius-rotundicauda grouped together as a monophyletic taxon. This topology is significantly better than all the alternatives when evaluated with the RELL (resampling estimated log-likelihood) method. PMID- 12116486 TI - [Post-coital contraception is changing]. PMID- 12116487 TI - [New light therapies for dermatitis]. PMID- 12116488 TI - [Illnesses and stigmatization]. PMID- 12116490 TI - [Why did a young man get a recurrent pulmonary embolism?]. PMID- 12116489 TI - [Insidious drug eruption]. PMID- 12116491 TI - [A red eye]. PMID- 12116492 TI - [Changes in the liver]. PMID- 12116493 TI - [Suitable exercise for suitable persons!]. PMID- 12116494 TI - [Exercise as an addiction]. PMID- 12116495 TI - [Exercise-associated sudden deaths in young persons]. PMID- 12116496 TI - [Research of athletic injuries is useful for everybody]. PMID- 12116497 TI - [Exercise and diet as targeted treatments of metabolic syndrome]. PMID- 12116498 TI - [Exercise as the prevention and treatment of coronary artery disease]. PMID- 12116499 TI - [Exercise and lungs]. PMID- 12116500 TI - [Problems of a young, growing athlete]. PMID- 12116501 TI - [Exercise preserves working ability and the elderly's functional capacity]. PMID- 12116502 TI - [Common traits in overtrained athletes and in persons with professional burn out]. PMID- 12116503 TI - Synthetic surgical gloves. AB - Surgical gloves are used by healthcare workers to protect them against bloodborne pathogens and other potential infectants and to prevent wound contamination in patients. In response to the increasing prevalence of allergies to natural rubber latex (NRL) among patients and medical staff, the trend toward purchasing gloves made of synthetic materials is on the rise. However, latex continues to dominate the market, and some people still perceive synthetic gloves as providing less protection and being less comfortable than NRL gloves. For this Update Evaluation, we present our findings for three newly evaluated glove models from three manufacturers and summarize our findings for the seven previously evaluated models that are still on the market. (Our earlier Evaluation was published in the February-March 2000 Health Devices.) As in the previous Evaluation, our ratings are based on the gloves' barrier effectiveness--that is, their resistance to viral penetration and their durability--and comfort. We also compared these characteristics of the synthetic gloves to those of NRL gloves. We found that all the evaluated gloves offer adequate barrier protection but that their level of comfort varies considerably. We rate three models Preferred, five models Acceptable, and two models Not Recommended. PMID- 12116504 TI - Medical telemetry is on the move. Is it time for you to change frequencies? AB - Medical telemetry in the United States is in the midst of a transformation. The VHF and UHF transmission bands traditionally used for most medical telemetry have become increasingly crowded. In response, the U.S. Federal Communications Commission (FCC) is making new frequency bands available exclusively for medical telemetry. Many users of the traditional bands will need to migrate out of them- but how soon will this become necessary? And what's involved in making the switch? In our September 2000 article "New Frequencies for Medical Telemetry: FCC's Plan Is Final--Now What?" we discussed some of the changes affecting telemetry. In the article that follows, we review those changes and explain what your facility needs to do in response. PMID- 12116505 TI - An introduction to FMEA. Using failure mode and effects analysis to meet JCAHO's proactive risk assessment requirement. Failure Modes and Effect Analysis. PMID- 12116506 TI - Connecting or disconnecting display during use can disrupt performance of Puritan Bennett 7200 ventilator. PMID- 12116508 TI - Is the customer always right? AB - What we have to offer as dental professionals cannot be weighed in mere physical balances. While what we do is by definition tangible and physical, what makes the physical valuable is our care, skill, and judgment. If we relegate care and judgment to the observation deck and skill is left to fend for itself, we risk no longer being professionals but rather sales clerks and technicians of treatment for individual teeth. Patients who are looking for a sales clerk or technician rather than a professional unfortunately will find one ... but it should be in another office. Let's be dentists by choice and of choice. Only then can we hold out for what is best for our patients and thereby help them to be people of choice as well. PMID- 12116509 TI - Interdisciplinary treatment for esthetic restorative dentistry. AB - Many of our patients have dental needs that general dentists cannot satisfy and may not even know about. Using an interdisciplinary team will expand the knowledge of all of the team members and can only enhance the restorative result, including function, long-term prognosis, and esthetics. Suggestions have been offered to help develop these teams, which may include general dentists, periodontists, orthodontists, oral surgeons, and endodontists. Team members are chosen based on individual skills needed for a particular case but all must be able to motivate and educate patients while providing excellent dentistry. PMID- 12116507 TI - New reports on dental analgesics. NSAIDs and cardiovascular effects, celecoxib for dental pain, and a new analgesic--tramadol with acetaminophen. PMID- 12116510 TI - The effect of air polishing on contemporary esthetic restorative materials. AB - Air polishing devices are designed primarily to remove soft deposits and stains from tooth surfaces. While improved strength and durability of esthetic restorative materials have resulted in increased usage, the effect of air polishing on these improved materials has not been determined. The purpose of this study was to examine the effect of air polishing on contemporary esthetic restorative materials. Four materials were tested: ceramic, hybrid composite resin, microfilled composite resin, and glass ionomer cement. Ceramics and hybrid composites exhibited the least change in surface roughness, followed by microfilled composites. Glass ionomers showed the greatest change in surface roughness. Results from this study suggest the surface roughness of all of the materials tested increased after exposure to air polishing instrumentation. Practitioners utilizing air polishing devices for prophylaxis procedures should exercise caution in the area of esthetic restorations. PMID- 12116511 TI - Clinical trials on the use of whitening strips in children and adolescents. AB - Vital tooth bleaching is becoming increasingly popular. Both professionally supervised and over-the-counter tooth whitening systems are being utilized. Recently, a tooth whitening system became available where a gel containing hydrogen peroxide was delivered via a flexible polythene strip. Three clinical trials were conducted using whitening strips to treat tooth discoloration on 132 children and adolescents. Results indicated the strip bleaching system to be an effective means for tooth whitening. Minimal sensitivity was associated with this whitening system, with most of the reported sensitivity being noted as mild. All sensitivity was relieved upon discontinuance of the whitening agent. PMID- 12116512 TI - Effect of bleaching gels on the surface roughness, hardness, and micromorphology of composites. AB - This study sought to evaluate the effect of three bleaching gels on the surface roughness, hardness, and morphology of resin-based composites basically designed for posterior and anterior use. The application of the bleaching agents tested had no statistically significant effect on the surface roughness or hardness of the composites tested. With both unbleached and bleached samples, the posterior composite was significantly harder than the anterior composite while the roughness was not significantly different (p > 0.06). PMID- 12116513 TI - Modern endodontic therapy for an incompletely developed tooth. AB - An important goal of modern dentistry is to preserve a functional, esthetic, and pain-free dentition. If irreversible pulp pathology occurs in an immature permanent tooth from traumatic or carious exposure, normal apical maturation may be interrupted. Conventional endodontic obturation techniques, if necessary, will be difficult and may not be adequate in such a tooth. The endodontic techniques of apexogenesis and apexification have been successful in achieving adequate root development and apical construction. These conventional techniques and materials are reviewed along with a promising material called mineral trioxide aggregate. PMID- 12116514 TI - Management of the dental patient with congestive heart failure. AB - This article presents the basic pathophysiology, clinical findings, medical management, and dental management for patients with congestive heart failure. The emphasis is on the role of the dentist in detecting these patients based on history and clinical findings, referring the patient for medical diagnosis and management, and working closely with the physician to develop a dental management plan that will be effective and safe for the patient. PMID- 12116515 TI - Evidence-based dentistry for the 21st century. AB - Evidence-based clinical dental practice relies upon the best available research evidence to formulate novel or improved modes of intervention specific to the needs of each patient. Evidence-based dentistry is a complex field. It is essential that its elements are clearly defined and understood for it to serve adequately and appropriately in clinical decision making. This paper discusses salient issues in evidence-based dentistry and examines the modality by which evidence-based dentistry will become the instrument for the integration of certain cutting-edge areas of science in dentistry in the new millennium. PMID- 12116516 TI - Dentofacial trauma in sport accidents. AB - Trauma due to sports shows a tremendous impact to a site of the body which is not coverable. The purpose of this study was to document the overall rates of dentofacial injuries sustained in 42 different sports and to determine the distribution of main injury types among sports and the most common injury type in frequent kinds of sport. The results indicate that sport accidents were responsible for six times as many facial injuries as work accidents and accounted for three times more injuries than violence or traffic accidents. Sports with high speed and high impact resulted more in facial bone fractures, whereas sports with low speed and low impact ended more in dental injuries. PMID- 12116517 TI - Pyogenic granuloma: case report in a 9-year-old girl. AB - Based on clinical features alone, pyogenic granuloma can be difficult to differentiate from a peripheral giant cell granuloma, a more aggressive oral lesion that could have consequences such as teeth displacement and alveolar bone resorption. A thorough clinical and radiographic examination is important to determine whether teeth and/or bone are involved. Furthermore, the early onset of puberty in females may increase the prevalence of pyogenic granuloma at a young age. PMID- 12116518 TI - The risks of oral-genital contact: a case report. AB - Our practice of dentistry allows us the opportunity to educate our patients about the many health risks that can affect the oral cavity. Certain behaviors have the potential for serious risk to dentists and staff as well as patients and their partners. This case report details the diagnosis, treatment, and discussion of oral gonococcal infection and the need for heightened awareness among the dental population. PMID- 12116519 TI - Oral diagnosis. Anterior mandibular radiolucency. PMID- 12116521 TI - U.S. Supreme Court decides seniority under ADA Title I. PMID- 12116520 TI - Oral diagnosis. Multifocal mixed radiolucencies and radiopacities. PMID- 12116522 TI - 2001 employment decisions under the ADA Title I--survey update. PMID- 12116524 TI - Chroniccaresolutions.com: a unique Web site for the chronically ill and disabled. PMID- 12116525 TI - www. Information resources for stroke. AB - The World Wide Web (www), or the Internet, is an essential educational tool for stroke survivors, caregivers and family members, rehabilitation nurses, and other healthcare providers. An immense amount of information about stroke is available on the Web, from its prevention to dealing with its effects. Anyone can publish anything about stroke on the Web. This article presents criteria for evaluating Internet resources and identifies numerous reliable Web resources related to stroke, along with suggestions about how the information from these Web sites can be helpful to both public and professional consumers. PMID- 12116526 TI - The use of computers and the Internet as a source of health information for people with disabilities. AB - It becomes increasingly obvious that computers and the Internet will play a prominent role in healthcare in the 21st century in America. The use of these tools in telecommunications technology to inform and educate has resulted in the emerging field of interactive health communication (IHC). The value of this new field is heightened by its potential to make health-related information and services more accessible to vulnerable populations such as people with disabilities. The assessment of user needs is one of the first activities necessary in the development of IHC applications. This article reports our findings from a survey conducted as the first step in developing a program of interactive health applications for people with disabilities resulting from brain and spinal cord injuries. A rehabilitation center conducted a 3-year retrospective survey of patients who had completed rehabilitation from brain or spinal cord injury. The survey found that 73% of respondents surveyed had access to and used computers, and 68% had access to the Internet. Our findings show that people with disabilities are using computers and the Internet, suggesting these tools' potential as a medium for the dissemination of health-related information and services for this underserved population. Further findings and key issues related to IHC design and application are discussed. PMID- 12116527 TI - You've got mail: rehabilitation nurses on the RehabNurse-L LISTSERV. AB - Rehabilitation nursing is a specialty with its own values, beliefs, practice, and language. Nurses are socialized to a specialty through their work interactions, professional readings, and professional affiliations. In this new millennium, nurses can communicate beyond their immediate environment with peers around the world through e-mail discussion groups (e.g., LISTSERV). The Association of Rehabilitation Nurses (ARN) created the RehabNurse-L LISTSERV in May 1999 to provide a vehicle for nurses to network with colleagues worldwide. In this study, we reviewed the communications posted on that LISTSERV in an 18-month period to identify the discussion topics and the demographics of the 475 people who became members of the LISTSERV simply by logging onto the site. Of that number, 318 posted some type of communication--whether it be a comment or a question or a response to a previously-asked question. Members were from Australia, Canada, Finland, Ireland, Portugal, Slovenia, the United Kingdom, and the United States. Topics discussed included professional and clinical issues, disability and impairment, administration and regulation, cultural and international issues, and LISTSERV housekeeping issues. LISTSERV provides quick access to peers, an avenue for information sharing, and a unique support system--all of which offer many potential benefits to professional nurses in their daily practice. PMID- 12116528 TI - Geriatric rehabilitation: the influence of efficacy beliefs and motivation. AB - The theory of self-efficacy is that the more one believes in the efficacy of a specific activity, the more likely (motivated) one is to perform that activity. Most of the research into self-efficacy beliefs among older adults has been quantitative and has consistently supported the influence of those beliefs on behavior. However, it has not been established how efficacy beliefs actually influence motivation in older adults, or what sources of efficacy-enhancing information help strengthen those beliefs. The purpose of this study was to better understand the factors that influence the efficacy beliefs that motivate older adults in a rehabilitation program, and to uncover the relation between efficacy beliefs, motivation, and behavior (i.e., participation in rehabilitation activities). Seventy-seven older adults, 55 women and 22 men, in an inpatient geriatric rehabilitation program were interviewed. Through content analysis, 11 major themes were identified as factors that influence efficacy beliefs and motivate people to participate in rehabilitation: personal expectations, personality, role models, verbal encouragement, progress, past experiences, spirituality, physical sensations, individualized care, social supports, and goals. The findings support the theory of self-efficacy, and are best explained within a social cognitive theory framework. PMID- 12116529 TI - Local injection of botulinum toxin type A for hemifacial spasm. AB - The preliminary experience of botulinum toxin treatment for hemifacial spasm is reported in this study. Five patients were treated with 10 injections of botulinum toxin in total. Botulinum toxin had a good to excellent effect in all cases. Improvement was observed 2 weeks to 1 month after the injection. The duration of improvement was 0-9 months (mean 4.2 months). The peak rank tended to decrease and the duration of improvement increased after several treatments. Hemifacial spasm caused by the anterior inferior cerebellar artery tended to subside easily. In contrast, compression by the vertebral artery was more refractory. Continuous facial spasm caused by operative trauma subsided after the injection, but paroxysmal spasm still occurred when eating or laughing. Spasm caused by trauma disappeared 4.5 months after the injection. The complications, which were facial nerve paresis in two cases (3 injections, 30%) and diplopia in one case (1 injection, 10%), were transient and subsided in 2 weeks. PMID- 12116530 TI - Anterior cerebral artery dissections manifesting as cerebral hemorrhage and infarction, and presenting as dynamic angiographical changes--case report. AB - A 65-year-old woman presented with multiple dissecting aneurysms of the anterior cerebral artery (ACA) manifesting as hemiparesis on the right with dominance in the lower extremity. Computed tomography revealed hematoma in the left frontal lobe, corresponding to the area perfused by the callosomarginal artery. Initial angiography showed string sign and occlusion in the distal portion of the left callosomarginal artery and abnormal feeding suggesting double lumen of the A2 portion of the left ACA. The patient was treated conservatively under a diagnosis of multiple spontaneous dissecting aneurysms of the left ACA. Repeat angiography on Day 8 showed improvement of the string sign and occlusion in the left callosomarginal artery, and change of the double lumen of the A2 portion into string sign. Further angiography on Day 36 showed normalization of the left callosomarginal artery and improvement of the string sign in the A2 portion. Multiple spontaneous dissecting aneurysms of the ACA are extremely rare. Serial angiography beginning in the early stage will be important for correct diagnosis. PMID- 12116531 TI - Subarachnoid hemorrhage caused by dural arteriovenous fistula of the sphenobasal sinus--case report. AB - A 59-year-old woman presented with a rare middle fossa dural arteriovenous fistula (AVF) unrelated to the cavernous sinus manifesting only as subarachnoid hemorrhage. Angiography revealed shunts between the meningeal branches of both the internal and external carotid arteries and the sphenobasal sinus. The AVF drained into the superficial middle cerebral vein (SMCV) which had a varix and an anastomosis to a superior cerebral vein. The arterial supply vessels were eliminated surgically and the sinus was excised. Bleeding did not recur and there was no venous infarction. Dural AVF of the sphenoparietal sinus is associated with pulsatile exophthalmos and dural AVF of the sphenopetrosal sinus with tinnitus, but dural AVF of the sphenobasal sinus has no obvious symptom. Simple interruption of the SMCV at the penetration of the arachnoid membrane was possible because of the absence of a draining vessel to preserve AVF patency, but the arteries were eliminated in this patient to prevent formation of another AVF. PMID- 12116532 TI - Intramedullary spindle cell hemangioendothelioma of the thoracic spinal cord- case report. AB - A 28-year-old Malay man presented with progressive paraparesis over a period of 6 months. Magnetic resonance imaging of the spine revealed a thoracic intramedullary spinal cord tumor at the T-7 level with homogeneous enhancement following intravenous gadolinium administration. Laminectomy and partial decompression of the tumor was performed. Histological examination of the tumor revealed features of spindle cell hemangioendothelioma. The patient was managed with limited field radiotherapy followed by systemic interferon therapy. Good neurological improvement was seen subsequently. The patient has survived 48 months with growth restraint at the primary site, although residual neurological deficit persists. Immunotherapy should be considered as a treatment modality for intramedullary hemangioendothelioma of the spinal cord after surgery and radiotherapy. PMID- 12116533 TI - Calcified metastatic brain tumor--two case reports. AB - A 29-year-old man and a 46-year-old woman presented with calcified brain metastasis from pulmonary adenocarcinoma. Both patients had a relatively benign clinical course and prolonged survival after total removal of the tumors. The male patient was still alive 2 years 3 months after surgical resection of the metastatic and primary tumors. The female patient had had brain metastasis 8 years after surgical resection of lung cancer, and survived for 3 years 3 months after removal of the brain metastasis. The MIB-1 indexes of the two cases were relatively low compared with other cases of brain metastases. Calcification and low MIB-1 index may indicate longer survival in patients with metastatic tumors if the primary tumor is controlled. PMID- 12116534 TI - Multiple intracranial seeding of craniopharyngioma after repeated surgery--case report. AB - A 17-year-old woman presented with a rare case of intracranial seeding of craniopharyngioma after repeated surgery. She initially presented with secondary amenorrhea and visual impairment. Magnetic resonance imaging revealed a suprasellar mass. Subtotal removal of the tumor was performed. The diagnosis was adamantinomatous craniopharyngioma. Seven months later, the patient underwent a second operation for recurrence of the craniopharyngioma. Subsequently, ventriculoperitoneal (VP) shunting and gamma knife surgery were performed. Twenty seven months after the first operation, multiple cystic lesions were found in the right frontal and temporal lobes. Positive tumor cytology was observed in the cerebrospinal fluid obtained from the VP shunt chamber. These tumors were subtotally resected. However, the patient died from consecutive tumor recurrence 4 years after the initial diagnosis. PMID- 12116535 TI - Anterior transpetrosal approach for pontine cavernous angioma--case report. AB - A 58-year-old male patient presented with headache and unsteady gait. Magnetic resonance imaging revealed hemorrhage from a pontine cavernous angioma. The patient experienced stepwise aggravation of symptoms due to repeated hemorrhages. We decided to surgically remove the pontine cavernous angioma through an anterior transpetrosal approach, since the angioma and hematoma were located near the ventrolateral surface of the pons. The brain stem was incised at a site caudal to the trigeminal nerve and the hematoma and angioma were totally removed. No additional neurological deficits were observed following surgery. Brain stem cavernous angiomas are usually removed via a trans-fourth ventricle or lateral suboccipital approach. However, these approaches may not be appropriate if the angioma is located ventrally to the pons. We propose that the anterior transpetrosal approach is the method of choice for ventrally located pontine cavernous angioma. PMID- 12116536 TI - Integration of ultrasonography and endoscopy into transsphenoidal surgery with a "picture-in-picture" viewing system--technical note. AB - A technique to integrate ultrasonography and endoscopy is described for transsphenoidal surgery to prevent intraoperative internal carotid artery (ICA) related, life-threatening complications such as aneurysmal formation and carotid cavernous fistula. The ultrasound unit helps avoid direct injury to the ICA. The technical advantage of this system is the miniature 1-mm diameter microvascular probe, which does not disturb the operative field. An arterial or venous flow source of even an invisible vessel can be detected easily, noninvasively, and reproducibly. Real-time information with a 100% detection rate for the ICA is helpful for predicting localization even in the intracavernous portion, where the ICA is invisible. The endoscope unit can visualize the dead angle areas of the operating microscope by varying the endoscopic gateways and display on a "picture in-picture" system. The advantage of both devices is the integration with a video processor, so that the real-time information from each unit can be switched intraoperatively onto the display as required. This method is of particular help for removing lesions with intracavernous invasion or encasement of the ICA. PMID- 12116537 TI - Conflict, compromise formation, and structural theory. AB - Evidence is put forth to support the previously presented view (Brenner 1994, 1998) that present knowledge of mental conflict and compromise formation renders invalid the widely accepted theory of mind as functionally separable structures called id, ego, and superego. The nature, origins, and timing of conflict and compromise formation in mental development are discussed, as well as their relation to psychic trauma. PMID- 12116538 TI - Some hazards to neutrality in the psychoanalysis of candidates. AB - The author looks at the challenges confronting psychoanalysts as they attempt to approach a desirable goal of neutrality in the treatment of psychoanalytic candidates and others who will become or are professional colleagues. Many such hazards are embedded in the interlocking relationships that inevitably exist between the analytic institute and its personnel, its training analysts, and its candidate-analysands. The author considers difficulties in the crucial analysis of aggression in training analyses. The analysis of aggression plays a determinative role in the personal and professional lives of analysands, and vitally affects the health and creativity of analytic institutions and the future of psychoanalysis. PMID- 12116539 TI - Why don't our institutes teach the methodology of clinical psychoanalytic evidence? AB - One of the most pressing theoretical, therapeutic, and educational problems confronting the advance of psychoanalysis is our confusion about the nature of clinical evidence. There is considerable disagreement among psychoanalysts about how to define evidence. Recently, there has also been a growing tendency toward evidential nihilism, justified by some of the more radical constructivist narrative views of postmodern epistemology. Two surveys, conducted ten years apart, of all the psychoanalytic institutes recognized by the American Psychoanalytic Association, indicate a continuing and disturbing neglect of this topic in the curricula of our institutes. This paper reports the views of a number of psychoanalytic educators who participated in these surveys and is intended to stimulate discussion about the causes of this critical problem. PMID- 12116540 TI - Superego transformations through the analyst's capacity for reverie. AB - The author discusses the application of certain Kleinian and Bionian principles to psychoanalytic work with patients who suffer from symptoms arising out of pathological superego functioning. Clinical vignettes are presented to demonstrate how the analyst's willingness to employ reverie, and to move from conviction-based interpretations to more open and tentative ones, can help such patients to change maladaptive behavioral patterns that may have stemmed from early interactions with an unavailable or nonreceptive Other. PMID- 12116541 TI - Where models intersect: a metapsychological approach. AB - Current scientific interest in how the mind works creates a major challenge for psychoanalysis. The author proposes metapsychology as a bridging concept for an interdisciplinary dialogue. She presents a new framework on a microstructural level, within which different psychic representations are hierarchically organized. This framework permits a detailed comparison with Alexander Luria's (1973) neuropsychological model of the working brain (including recent theories of affect), and makes it possible to delineate the similarities as well as the differences between the psychoanalytic model of the mind and the neuropsychological model of the brain. PMID- 12116542 TI - Boesky examines our failure to teach. PMID- 12116543 TI - On psychic bisexuality. PMID- 12116544 TI - Lou shoe's lament. PMID- 12116545 TI - [Vaccination program against measles. 2 vaccinations closes the gap]. PMID- 12116546 TI - [New hope for heart failure patients. Fatigued hearts need medication and electrical aids]. PMID- 12116547 TI - [Until a donor organ becomes available. New life force for the liver]. PMID- 12116548 TI - [Tumors, headaches and concentration problems... How dangerous are mobile telephones really? (interview by Waltraud Paukstadt)]. PMID- 12116549 TI - [Dietary fats, salt and alcohol drive up blood pressure. Educate hypertensive patient on avoidance]. AB - General non-drug measures today form the accepted basis for every antihypertensive treatment. WHO identifies the most important of these as weight reduction, salt restriction and moderation in the use of alcohol. These measures can effectively lower raised blood pressure and normalize grade I hypertension, especially in borderline cases. The target of these efforts is a weight reduction of at least 5%, restriction of salt to 5-6 g/day and alcohol consumption not exceeding 20 g/day for women and 25 g/day for men. No negative effects are to be expected from these restrictions, which are really nothing more than a return to "normal portions" with regard to eating and drinking habits. PMID- 12116550 TI - [An exercise program for the hypertensive patient. Leaving hypertension behind]. AB - Today, regular exercise is considered an important therapeutic principle in arterial hypertension. Apart from weight reduction, the positive effects of sports (moderately practiced) include a reduction in peripheral resistance, improvement of endothelial dysfunction, and an increase in insulin sensitivity of the skeletal muscles. Training programs should be tailored to the physical and mental status of the patient, in particular to the severity of the hypertension, where indicated, with consideration being given to the hypertensive drugs used. In the case of marked hypertension or already existing secondary complications, training should be started only when the patient's blood pressure has been brought under pharmacological control. Ideally, training should be applied as 20 60 minute sessions at least twice weekly, with exercise keeping within aerobic capacity. PMID- 12116551 TI - [Essential hypertension and stress. When do yoga, psychotherapy and autogenic training help?]. AB - Psychosocial factors play an important role in the development and course of essential hypertension, although "stress" can account for only 10% of blood pressure variance. A variety of psychotherapeutic interventions, such as relaxation techniques (autogenic training or progressive muscular relaxation), behavioral therapy or biofeedback techniques, can lower elevated blood pressure by an average of 10 mmHg (systolic) and 5 mmHg (diastolic). As a "secondary effect", such measures may also prompt the hypertensive to adopt a more health conscious lifestyle. PMID- 12116552 TI - [The treatment options broaden. New chances for libido]. PMID- 12116553 TI - [Emergencies in general practice. Acute poisoning with medicinal drugs]. PMID- 12116554 TI - [Illness--a "subjective definition". Illness determined not just by the physician]. PMID- 12116555 TI - [Comparative study with 2 L-thyroxine-iodide combinations. Iodine deficient goiter can be decreased also with less thyroxine]. PMID- 12116556 TI - [Vertigo of unknown origin. A pacemaker cannot help here]. PMID- 12116557 TI - [Impotence as a relationship problem: what is the attitude of women?]. PMID- 12116558 TI - [Secondary prevention after stroke. 2 platelet inhibitors are better than one]. PMID- 12116559 TI - [Hypertensive patient with concomitant illnesses. Killing 2 flies with 1 stroke]. PMID- 12116560 TI - [Therapy of hypertension. AT1 blockers soon to be the first choice?]. PMID- 12116561 TI - [When nucleoside analogs cannot be tolerated. HIV therapy with booster]. PMID- 12116562 TI - [Fascinating film about physiological processes. Discovery trip through the body]. PMID- 12116563 TI - [Hypoglycemia risk. Why elderly patients are especially at risk]. PMID- 12116565 TI - [Slender patient with type 2 diabetes. That could be a diagnostic error!]. PMID- 12116564 TI - [New treatment strategies. Will diabetes be curable?]. PMID- 12116566 TI - [Study: our students are becoming ever more slack. Does movement deficit result in dumbness? (interview by Michael Koczorek)]. PMID- 12116567 TI - [Early diagnosis of Lyme borreliosis. Do not look only for erythema migrans]. AB - Although the typical erythema migrans is a relatively easy-to-recognize manifestation of Lyme borreliosis, it is by no means the sole cutaneous manifestation of infection with borreliae. In particular the early stages of the disease, with their paucity of symptoms or wide variability of the clinical symptomatology, often present a diagnostic challenge. This means that in the event of unclear unspecific cutaneous lesions indicating, for example, erysipelas or urticaria, Lyme borreliosis should also receive differential diagnostic consideration. Culture of the organism, DNA-based confirmation of B. burgdorferi or, where indicated, a serological investigation can confirm the diagnosis at an early stage. The treatment of choice compromises an antibiotic administered for an adequate duration and at an adequate dose, e.g. doxycycline 2 x 100 mg over three weeks). If treatment is initiated in a local or disseminated early stage, healing rates of more than 85% can be achieved. PMID- 12116568 TI - [Patient with questionable Lyme borreliosis. Which laboratory parameters validate your clinical suspicion?]. AB - Lyme borreliosis is the most common of the infectious diseases transmitted by ticks in the Northern hemisphere. In Europe, Lyme borreliosis can be caused by three species of Borrelia: B. burgdorferi sensu stricto, B. garinii and B. afzelii. The microbiological diagnosis is established primarily on the basis of antibody detection, and secondarily by detection of the pathogen itself. Suitable material for the detection of the latter are various body fluids (CSF, joint fluid and biopsy material, in particular in the skin). In the case of antibodies, the substrate is usually serum. If neuroborreliosis is suspected, CSF should always be investigated (CSF/serum, pairwise, same day!). The microbiological findings must be interpreted in conjunction with the clinical presentation. A negative serological result does not exclude an early manifestation, a positive finding is no proof of a clinically manifest infection--it might be a titer from an earlier infection. PMID- 12116570 TI - [Neck pain. Diagnosis and therapy of acute cervical syndrome]. PMID- 12116569 TI - [Diagnosis and therapy of neuroborreliosis. On the hunt for the "great imitator"]. AB - Neurological manifestations are characteristic of stage 2 and stage 3 borreliosis. In stage 2, some 15% of the patients have neurological symptoms expressed as a triad of aseptic meningitis, cranial neuritis and radiculitis. Stage 3--chronic neuroborreliosis affects some 5% of untreated patients. The condition has its onset at the earliest 6 months after the infection, and is characterized by encephalopathic symptoms, such as fatigue, sleep and memory disturbances, and depressive states. Further manifestations of this stage may be Lyme polyneuropathy, in rare cases also progressive borrelia encephalomyelitis and cerebrovascular neuroborreliosis. The treatment of choice is intravenous administration of cephalosporins over 2-4 weeks. The success of treatment should be assessed on the basis of the clinical course rather than on laboratory results. Patience is required in the treatment of the post-Lyme syndrome, characterized by residual symptoms, recurrences or a relapsing course. PMID- 12116571 TI - [Suspected cardiac neurosis? On to heart catheterization?]. PMID- 12116572 TI - [The coronary heart disease factors obesity, faulty nutrition, smoking, inactivity. Give your patient the deciding motivation]. AB - Primary prevention of cardiovascular disease means recognizing risk factors and- as far as possible--eliminating them. Risk factors that can be influenced include smoking, a high-fat diet and lack of exercise which, by damaging the endothelium and causing atherosclerosis, can lead to myocardial infarction and stroke. It is estimated that 70-80% of all cardiovascular events could be avoided by adopting an optimized lifestyle. The rewards of implementing a more health-conscious way of life are improved quality of life and an increase in life expectancy of five to six years. The aim of the Germany-wide "CHD Scoring Week" lasting from 3 June to 7 June, organized in cooperation with the German Cardiological Society, is to promote a greater awareness of the need for primary prevention among both physicians and patients. PMID- 12116573 TI - [Arterial compression syndrome. First the fingers tingle, then thrombosis of the brachial artery]. AB - The thoracic outlet syndrome and the entrapment syndrome involving the popliteal artery are two major examples of arterial compression syndrome. Since not only the arteries, but also the entire neurovascular bundle is compressed, the initial symptoms are often neurological: paresthesias, tingling, numbness, etc., in particular when certain movements are carried out. In such a case, bilateral blood pressure measurements, provocative tests and Doppler (duplex) examination are indicated. A point that is, perhaps, less well-known is the fact that compression can lead to serious vascular lesions ranging from post-stenotic aneurysms to complete thrombotic occlusion and peripheral emboli. When the diagnosis has been confirmed, early surgery is indicated. PMID- 12116574 TI - [Stabilizing affective symptoms without adverse cardiac effects. Myocardial infarct risk after depression--and vice versa]. PMID- 12116575 TI - [Heart patients with depression. Pay special attention!]. PMID- 12116576 TI - The emergence of Systematic Biology. PMID- 12116577 TI - Philosophy and phylogenetic inference: a comparison of likelihood and parsimony methods in the context of Karl Popper's writings on corroboration. AB - Advocates of cladistic parsimony methods have invoked the philosophy of Karl Popper in an attempt to argue for the superiority of those methods over phylogenetic methods based on Ronald Fisher's statistical principle of likelihood. We argue that the concept of likelihood in general, and its application to problems of phylogenetic inference in particular, are highly compatible with Popper's philosophy. Examination of Popper's writings reveals that his concept of corroboration is, in fact, based on likelihood. Moreover, because probabilistic assumptions are necessary for calculating the probabilities that define Popper's corroboration, likelihood methods of phylogenetic inference- with their explicit probabilistic basis--are easily reconciled with his concept. In contrast, cladistic parsimony methods, at least as described by certain advocates of those methods, are less easily reconciled with Popper's concept of corroboration. If those methods are interpreted as lacking probabilistic assumptions, then they are incompatible with corroboration. Conversely, if parsimony methods are to be considered compatible with corroboration, then they must be interpreted as carrying implicit probabilistic assumptions. Thus, the non probabilistic interpretation of cladistic parsimony favored by some advocates of those methods is contradicted by an attempt by the same authors to justify parsimony methods in terms of Popper's concept of corroboration. In addition to being compatible with Popperian corroboration, the likelihood approach to phylogenetic inference permits researchers to test the assumptions of their analytical methods (models) in a way that is consistent with Popper's ideas about the provisional nature of background knowledge. PMID- 12116578 TI - Philosophical conjectures and their refutation. AB - Sir Karl Popper is well known for explicating science in falsificationist terms, for which his degree of corroboration formalism, C(h,e,b), has become little more than a symbol. For example, de Queiroz and Poe in this issue argue that C(h,e,b) reduces to a single relative (conditional) probability, p(e,hb), the likelihood of evidence e, given both hypothesis h and background knowledge b, and in reaching that conclusion, without stating or expressing it, they render Popper a verificationist. The contradiction they impose is easily explained--de Queiroz and Poe fail to take account of the fact that Popper derived C(h,e,b) from absolute (logical) probability and severity of test, S(e,h,b), where critical evidence, p(e,b), is fundamental. Thus, de Queiroz and Poe's conjecture that p(e,hb) = C(h,e,b) is refuted. Falsificationism, not verificationism, remains a fair description of the parsimony method of inference used in phylogenetic systematics, not withstanding de Queiroz and Poe's mistaken understanding that "statistical" probability justifies that method. Although de Queiroz and Poe assert that maximum likelihood has the power "to explain data", they do not successfully demonstrate how causal explanation is achieved or what it is that is being explained. This is not surprising, bearing in mind that what is assumed about character evolution in the accompanying likelihood model M cannot then be explained by the results of a maximum likelihood analysis. PMID- 12116579 TI - Towards an inclusive philosophy for phylogenetic inference. AB - We defend and expand on our earlier proposal for an inclusive philosophical framework for phylogenetics, based on an interpretation of Popperian corroboration that is decoupled from the popular falsificationist interpretation of Popperian philosophy. Any phylogenetic inference method can provide Popperian "evidence" or "test statements" based on the method's goodness-of-fit values for different tree hypotheses. Corroboration, or the severity of that test, requires that the evidence is improbable without the hypothesis, given only background knowledge that includes elements of chance. This framework contrasts with attempted Popperian justifications for cladistic parsimony--in which evidence is the data, background knowledge is restricted to descent with modification, and "corroboration," as a by-product of nonfalsification, is to be measured by cladistic parsimony. Recognition that cladistic "corroboration" reflects only goodness-of-fit, not corroboration/severity, makes it clear that standard cladistic prohibitions, such as restrictions on the evolutionary models to be included in "background knowledge," have no philosophical status. The capacity to assess Popperian corroboration neither justifies nor excludes any phylogenetic method, but it does provide a framework in phylogenetics for learning from errors -cases where apparent good evidence is probable even without the hypothesis. We explore these issues in the context of corroboration assessments applied to likelihood methods and to a new form of parsimony. These different forms of evidence and corroboration assessment point also to a new way to combine evidence -not at the level of overall fit, but at the level of overall corroboration/severity. We conclude that progress in an inclusive phylogenetics will be well served by the rejection of cladistic philosophy. PMID- 12116580 TI - Empirical and hierarchical Bayesian estimation of ancestral states. AB - Several methods have been proposed to infer the states at the ancestral nodes on a phylogeny. These methods assume a specific tree and set of branch lengths when estimating the ancestral character state. Inferences of the ancestral states, then, are conditioned on the tree and branch lengths being true. We develop a hierarchical Bayes method for inferring the ancestral states on a tree. The method integrates over uncertainty in the tree, branch lengths, and substitution model parameters by using Markov chain Monte Carlo. We compare the hierarchical Bayes inferences of ancestral states with inferences of ancestral states made under the assumption that a specific tree is correct. We find that the methods are correlated, but that accommodating uncertainty in parameters of the phylogenetic model can make inferences of ancestral states even more uncertain than they would be in an empirical Bayes analysis. PMID- 12116581 TI - Mining the mammalian genome for artiodactyl systematics. AB - A total of 7,806 nucleotide positions derived from one mitochondrial and eight nuclear DNA segments were used to provide a robust phylogeny for members of the order Artiodactyla. Twenty-four artiodactyl and two cetacean species were included, and the horse (order Perissodactyla) was used as the outgroup. Limited rate heterogeneity was observed among the nuclear genes. The partition homogeneity tests indicated no conflicting signal among the nuclear genes fragments, so the sequence data were analyzed together and as separate loci. Analyses based on the individual nuclear DNA fragments and on 34 unique indels all produced phylogenies largely congruent with the topology from the combined data set. In sharp contrast to the nuclear DNA data, the mtDNA cytochrome b sequence data showed high levels of homoplasy, failed to produce a robust phylogeny, and were remarkably sensitive to taxon sampling. The nuclear DNA data clearly support the paraphyletic nature of the Artiodactyla. Additionally, the family Suidae is diphyletic, and the nonruminating pigs and peccaries (Suiformes) were the most basal cetartiodactyl group. The morphologically derived Ruminantia was always monophyletic; within this group, all taxa with paired bony structures on their skulls clustered together. The nuclear DNA data suggest that the Antilocaprinae account for a unique evolutionary lineage, the Cervidae and Bovidae are sister taxa, and the Giraffidae are more primitive. PMID- 12116582 TI - The importance of time/space in diagnosing the causality of phylogenetic events: towards a "chronobiogeographical" paradigm? AB - A shift from a traditional biogeographical paradigm in cladistic biogeography to a chronobiogeographical paradigm is proposed. The chronobiogeographical paradigm aims to utilize temporal data in conjunction with spatial data in the detection of discrete historical events, such as vicariance and vicariant speciation, in cladograms. The concepts of primary and secondary congruency are introduced in relation to the distinction between repeated area relationships (primary congruency) and common extrinsic causality (secondary congruency). Simple hypothetical examples demonstrate that area cladograms cannot be safely interpreted purely as representing the sequence of area fragmentation; rather, they reflect recency of biotic interaction. Temporal data are shown to have a direct and potentially profound influence on the results of traditional cladistic biogeographical analyses, indicating the necessity of developing a chronobiogeographical approach. The implementation of the paradigm is considered first from a theoretical viewpoint and then in the context of the type of empirical data usually available. An as yet undevised "time/space algorithm" is deemed necessary for the latter, and guidelines are presented for the development of such an algorithm. Finally, we argue that the most rigorous and philosophically justified approach to the detection of phylogenetic causal events can be found only when temporal and spatial data are considered simultaneously. Consequently, the chronobiogeographical paradigm is seen as a logical elaboration of, not a replacement for, the biogeographical paradigm. PMID- 12116583 TI - Failure of the ILD to determine data combinability for slow loris phylogeny. AB - Tests for incongruence as an indicator of among-data partition conflict have played an important role in conditional data combination. When such tests reveal significant incongruence, this has been interpreted as a rationale for not combining data into a single phylogenetic analysis. In this study of lorisiform phylogeny, we use the incongruence length difference (ILD) test to assess conflict among three independent data sets. A large morphological data set and two unlinked molecular data sets--the mitochondrial cytochrome b gene and the nuclear interphotoreceptor retinoid binding protein (exon 1)--are analyzed with various optimality criteria and weighting mechanisms to determine the phylogenetic relationships among slow lorises (Primates, Loridae). When analyzed separately, the morphological data show impressive statistical support for a monophyletic Loridae. Both molecular data sets resolve the Loridae as paraphyletic, though with different branching orders depending on the optimality criterion or character weighting used. When the three data partitions are analyzed in various combinations, an inverse relationship between congruence and phylogenetic accuracy is observed. Nearly all combined analyses that recover monophyly indicate strong data partition incongruence (P = 0.00005 in the most extreme case), whereas all analyses that recover paraphyly indicate lack of significant incongruence. Numerous lines of evidence verify that monophyly is the accurate phylogenetic result. Therefore, this study contributes to a growing body of information affirming that measures of incongruence should not be used as indicators of data set combinability. PMID- 12116584 TI - A conditional probability of reconstruction measure for internal cladogram branches. AB - The conditional probability of reconstruction is a measure of the robustness of cladogram internodes and, unlike Bremer support and bootstrapping values, directly gauges probability. The new method compares the three putative branch lengths (the optimal and two alternatives) obtained through branch recalculation after nearest neighbor interchange and recalculation under constraint. With rooted trees, one switches the two free lineages attached at the distal end of an internal branch with the basal lineage. Probabilistic reconstruction of a branch for small data sets (e.g., morphological) is defined as having no contrary support for the two alternative branches and, when sufficient data are available (e.g., molecular studies), as meeting a selected confidence limit in chi-squared analysis. The exact probability that the internal branch is reconstructed is the same as the preselected confidence level met with chi-squared analysis; alternatively, it is a simple calculation of the length of the optimal branch divided by the sum of the lengths of all three putative branches. This new measure of robustness allows calculation of summary probabilities of subclade and tree reconstruction. The measure is conditional on a particular data set and optimization method but may also compare support from conflicting gene trees. Examples are provided by a morphological data set (the bryophyte Didymodon) and a molecular data set (primates). PMID- 12116585 TI - Popper and likelihood versus "Popper". PMID- 12116586 TI - Are the fossil data really at odds with the molecular data? Morphological evidence for cetartiodactyla phylogeny reexamined. PMID- 12116587 TI - Incorporation, relative homoplasy, and effect of gap characters in sequence-based phylogenetic analyses. PMID- 12116589 TI - Taxon sampling and the phylogenetic position of Passeriformes: evidence from 916 avian cytochrome b sequences. PMID- 12116588 TI - A partitioned likelihood analysis of swallowtail butterfly phylogeny (Lepidoptera:Papilionidae). AB - Although it is widely agreed that data from multiple sources are necessary to confidently resolve phylogenetic relationships, procedures for accommodating and incorporating heterogeneity in such data remain underdeveloped. We explored the use of partitioned, model-based analyses of heterogeneous molecular data in the context of a phylogenetic study of swallowtail butterflies (Lepidoptera: Papilionidae). Despite substantial basic and applied study, phylogenetic relationships among the major lineages of this prominent group remain contentious. We sequenced 3.3 kb of mitochondrial and nuclear DNA (2.3 kb of cytochrome oxidase I and II and 1.0 kb of elongation factor-1 alpha, respectively) from 22 swallowtails, including representatives of Baroniinae, Parnassiinae, and Papilioninae, and from several moth and butterfly outgroups. Using parsimony, we encountered considerable difficulty in resolving the deepest splits among these taxa. We therefore chose two outgroups with undisputed relationships to each other and to Papilionidae and undertook detailed likelihood analyses of alternative topologies. Following from previous studies that have demonstrated substantial heterogeneity in the evolutionary dynamics among process partitions of these genes, we estimated evolutionary parameters separately for gene-based and codon-based partitions. These values were then used as the basis for examining the likelihoods of possible resolutions and rootings under several partitioned and unpartitioned likelihood models. Partitioned models gave markedly better fits to the data than did unpartitioned models and supported different topologies. However, the most likely topology varied from model to model. The most likely ingroup topology under the best-fitting, six-partition GTR + gamma model favors a paraphyletic Parnassiinae. However, when examining the likelihoods of alternative rootings of this tree relative to rootings of the classical hypothesis, two rootings of the latter emerge as most likely. Of these two, the most likely rooting is within the Papilioninae, although a rooting between Baronia and the remaining Papilionidae is only nonsignificantly less likely. PMID- 12116590 TI - Useful characters in gastropod phylogeny: soft information or hard facts? PMID- 12116591 TI - Landmark coordinates aligned by procrustes analysis do not lie in Kendall's shape space. PMID- 12116592 TI - Molecular and morphological phylogenetic analysis of an insular radiation in Pacific black flies (Simulium). AB - Ecological adaptation within islands may have figured prominently in the insular radiation of black flies (subgenus Inseliellum) in the Society Islands, French Polynesia. To aid in understanding the sequence of ecological shifts in this group, we have constructed a phylogeny by using morphology, the cytochrome oxidase I (COI) gene, and the small ribosomal subunit (12S) gene. The strong influence of COI on the combined analysis tree was evident from its contribution to the partitioned Bremer support (62%). The net effect of including 12S was to reduce overall tree support. Different character sets resolved different portions of the combined analysis tree, with COI resolving recent lineages, 12S resolving basal relationships, and morphology supporting the monophyly of taxa having smaller larval feeding fans (oviceps group). The Partition Homogeneity and Kashino-Hasegawa tests indicated significant incongruence between morphological and mitochondrial data. The Templeton test revealed that morphology and the combined (COI + 12S) mitochondrial data were incongruent. This conflict stems primarily from disagreement over the monophyly of taxa having much smaller larval feeding fans. Either convergence in a subset of morphological characters, low phylogenetic signal among mitochondrial sequences, or lineage-sorting causing the mitochondrial data to track an incorrect evolutionary history may be responsible for these results. PMID- 12116593 TI - Incongruence of mitochondrial and nuclear gene trees in the Carabid beetles Ohomopterus. AB - We studied the molecular phylogeny of the carabid subgenus Ohomopterus (genus Carabus), using two mitochondrial (mt) DNA regions (16SrRNA and NADH dehydrogenase subunit 5) and three nuclear DNA regions (wingless, phosphoenolpyruvate carboxykinase, and an anonymous locus). We revisited the previously reported incongruence between the distribution of mtDNA markers and morphologically defined species (Su et al., 1996; J. Mol. Evol. 43:662-671), which those authors attributed to "type switching", a concerted change in many morphological characters that results in the repeated evolution of a particular morphological type. Our mtDNA gene tree obtained from 44 individuals representing all 15 currently recognized species of Ohomopterus revealed that haplotypes isolated from individuals of a single "species" were frequently separated into distant clades, confirming the previous report. The three nuclear markers generally conformed better-with the morphologically defined species than did the mitochondrial markers. The phylogenetic signal in mtDNA and nuclear DNA data differed strongly, and these two partitions were significantly incongruent with each other according to the incongruence length difference test of Farris et al. (1994; Cladistics 10:315-320), although the three nuclear partitions were not homogeneous either. Our results did not support the type-switching hypothesis that had been proposed to fit the morphological data to the mitochondrial gene tree: The incongruence of the mtDNA tree with other nuclear markers indicates that the mtDNA-based tree does not reflect species history any better than the morphological data do. Incongruence of gene trees in Ohomopterus may have been promoted by the complex processes of geographic isolation and hybridization in the Japanese Archipelago that have led to occasional gene flow and recombination between separated entities. The occurrence of reticulate patterns in this group is intriguing, because species of Ohomopterus exhibit extremely divergent genitalic structures that represent a highly efficient reproductive isolation mechanism. PMID- 12116594 TI - Changing the landscape: a new strategy for estimating large phylogenies. AB - In this paper we describe a new heuristic strategy designed to find optimal (parsimonious) trees for data sets with large numbers of taxa and characters. This new strategy uses an iterative searching process of branch swapping with equally weighted characters, followed by swapping with reweighted characters. This process increases the efficiency of the search because, after each round of swapping with reweighted characters, the subsequent swapping with equal weights will start from a different group (island) of trees that are only slightly, if at all, less optimal. In contrast, conventional heuristic searching with constant equal weighting can become trapped on islands of suboptimal trees. We test the new strategy against a conventional strategy and a modified conventional strategy and show that, within a given time, the new strategy finds trees that are markedly more parsimonious. We also compare our new strategy with a recent, independently developed strategy known as the Parsimony Ratchet. PMID- 12116595 TI - Exploring among-site rate variation models in a maximum likelihood framework using empirical data: effects of model assumptions on estimates of topology, branch lengths, and bootstrap support. AB - We have investigated the effects of different among-site rate variation models on the estimation of substitution model parameters, branch lengths, topology, and bootstrap proportions under minimum evolution (ME) and maximum likelihood (ML). Specifically, we examined equal rates, invariable sites, gamma-distributed rates, and site-specific rates (SSR) models, using mitochondrial DNA sequence data from three protein-coding genes and one tRNA gene from species of the New Zealand cicada genus Maoricicada. Estimates of topology were relatively insensitive to the substitution model used; however, estimates of bootstrap support, branch lengths, and R-matrices (underlying relative substitution rate matrix) were strongly influenced by the assumptions of the substitution model. We identified one situation where ME and ML tree building became inaccurate when implemented with an inappropriate among-site rate variation model. Despite the fact the SSR models often have a better fit to the data than do invariable sites and gamma rates models, SSR models have some serious weaknesses. First, SSR rate parameters are not comparable across data sets, unlike the proportion of invariable sites or the alpha shape parameter of the gamma distribution. Second, the extreme among site rate variation within codon positions is problematic for SSR models, which explicitly assume rate homogeneity within each rate class. Third, the SSR models appear to give severe underestimates of R-matrices and branch lengths relative to invariable sites and gamma rates models in this example. We recommend performing phylogenetic analyses under a range of substitution models to test the effects of model assumptions not only on estimates of topology but also on estimates of branch length and nodal support. PMID- 12116596 TI - Stochastic search strategy for estimation of maximum likelihood phylogenetic trees. AB - The maximum likelihood (ML) method of phylogenetic tree construction is not as widely used as other tree construction methods (e.g., parsimony, neighbor joining) because of the prohibitive amount of time required to find the ML tree when the number of sequences under consideration is large. To overcome this difficulty, we propose a stochastic search strategy for estimation of the ML tree that is based on a simulated annealing algorithm. The algorithm works by moving through tree space by way of a "local rearrangement" strategy so that topologies that improve the likelihood are always accepted, whereas those that decrease the likelihood are accepted with a probability that is related to the proportionate decrease in likelihood. Besides greatly reducing the time required to estimate the ML tree, the stochastic search strategy is less likely to become trapped in local optima than are existing algorithms for ML tree estimation. We demonstrate the success of the modified simulated annealing algorithm by comparing it with two existing algorithms (Swofford's PAUP* and Felsenstein's DNAMLK) for several theoretical and real data examples. PMID- 12116597 TI - Phylogenetic utility of different types of molecular data used to infer evolutionary relationships among stalk-eyed flies (Diopsidae). AB - A phylogenetic hypothesis of relationships among 33 species of stalk-eyed flies was generated from a molecular data set comprising three mitochondrial and three nuclear gene regions. A combined analysis of all the data equally weighted produced a single most-parsimonious cladogram with relatively strong support at the majority of nodes. The phylogenetic utility of different classes of molecular data was also examined. In particular, using a number of different measures of utility in both a combined and separate analysis framework, we focused on the distinction between mitochondrial and nuclear genes and between faster-evolving characters and slower-evolving characters. For the first comparison, by nearly any measure of utility, the nuclear genes are substantially more informative for resolving diopsid relationships than are the mitochondrial genes. The nuclear genes exhibit less homoplasy, are less incongruent with one another and with the combined data, and contribute more support to the combined analysis topology than do the mitochondrial genes. Results from the second comparison, however, provide little evidence of a clear difference in utility. Despite indications of rapid divergence and saturation, faster-evolving characters in both the nuclear and mitochondrial data sets still provide substantial phylogenetic signal. In general, inclusion of the more rapidly evolving data consistently improves the congruence among partitions. PMID- 12116598 TI - Gender differences in the proarrhythmic potential of QT-prolonging drugs. AB - Female gender is associated with an increased risk of torsades de pointes (TDP) in the setting of drugs that can prolong the QT interval. These drugs are all potassium channel blocking agents and include many frequently used antiarrhythmic drugs, as well as a variety of noncardiac medications. The mechanism of this gender difference is unknown. Some women at risk are silent carriers of the long QT syndrome. The concept of "repolarization reserve" can help predict which individuals are at risk for developing drug-induced TDP. Generic formulations of QT-prolonging drugs have the potential to increase proarrhythmic risk to women. Female subjects need to be well represented in the testing of new potassium channel blocking drugs. PMID- 12116599 TI - Assessment of women with suspected myocardial ischemia: review of findings of the Women's Ischemia Syndrome Evaluation (WISE) Study. AB - Coronary artery disease (CAD) is the most common cause of death in women. In general, noninvasive testing in women is less reliable compared with testing in men, and most major clinical trials in CAD have included only a minority of female subjects. The Women's Ischemia Syndrome Evaluation (WISE) Study--sponsored by the National Heart, Lung, and Blood Institute--was initiated to improve diagnostic testing in women. The study consisted of four centers that tested existing methodologies as well as innovative techniques to improve diagnostic testing in women. The WISE study also aimed to clarify physiologic determinants of myocardial ischemia and determine whether angiographically normal coronary arteries are associated with myocardial ischemia. The following discussion provides an overview of the WISE study and reviews the reported and published data from the study. PMID- 12116600 TI - Gender differences in the presentation and symptoms of coronary artery disease. AB - Chest pain is a typical feature of obstructive coronary disease, but unless carefully evaluated, may not be a reliable predictor in women. The use of standardized questionnaires and evaluation tools has been developed and validated in men, but only partially in women. If women over the age of 65 are evaluated, typical features of angina are much more reliable in representing coronary disease than in younger women, who may have risk factors, but are less likely to have significant coronary disease. Many studies have shown that chest pain is the most common presenting symptom for both men and women with unstable coronary syndromes or myocardial infarction. Other associated features, such as nausea, shortness of breath, and back pain, may be more common in women, while diaphoresis is more common in men. Since men and women at risk for coronary disease should be evaluated when any potential symptoms emerge, it is useful to employ a standardized assessment of the characteristics of the symptoms as well as a uniform approach to further evaluation. PMID- 12116601 TI - Digital mammography. AB - Digital mammography, using novel detector technology, has been shown to be at least as sensitive as screen-film mammography for detecting early breast cancer. However, there are barriers to overcome, particularly in soft-copy display, workstation design, and equipment cost. These problems are actively being addressed and are solvable, eventually allowing digital mammography to assume a "prime time" role. PMID- 12116602 TI - Breast cancer risks: some clinically useful approaches. AB - Information about breast cancer risk is often confusing and may even be misleading when presented as a comparison of one risk versus another. Frequently used comparison formats include relative risks, odds ratios, and proportional risk reductions. This paper discusses five areas of breast cancer risk--average risk, prognosis following a breast cancer diagnosis, risk associated with use of hormone replacement therapy at menopause, use of tamoxifen as prevention, and risks associated with BRCA mutations--to show the clarity and clinical usefulness that are obtained when risks are presented not as comparisons, but in absolute terms with a time frame. PMID- 12116603 TI - Adjuvant systemic therapy for lymph node-negative breast cancer less than or equal to 1 cm. AB - Routine screening mammography has increased the incidence of stage I breast cancers. Many more women are being diagnosed with lymph node-negative tumors that are less than or equal to 1 cm in greatest diameter. The National Surgical Adjuvant Breast and Bowel Project recently performed a retrospective analysis of 10,302 women participating in one of five clinical trials, including women with lymph node-negative breast cancer. Of these women, 1259 had tumors less than or equal to 1 cm. The analysis of the women with tumors less than or equal to 1 cm revealed an improved relapse-free survival (RFS) if tamoxifen was given after surgery, compared with surgery alone, for women with estrogen receptor (ER) positive tumors; and for women with ER-negative tumors, RFS was improved by delivering chemotherapy after surgery. The authors suggested that adjuvant systemic therapy should be considered for anyone with an invasive breast cancer, regardless of the size of the tumor. This paper reviews the data presented in that important, historic article, and discusses their conclusions. Also reviewed are the most recent recommendations for treatment of primary breast cancer from the International Consensus Panel that convened at the Seventh International Conference on Adjuvant Therapy of Primary Breast Cancer in St. Gallen, Switzerland. That panel also addressed the issue of adjuvant systemic therapy in women considered to have a minimal or low risk of developing recurrent disease. PMID- 12116605 TI - Breast health--introduction and overview. PMID- 12116604 TI - Understanding and intervening in breast cancer's emotional and sexual side effects. AB - There is an evolving body of research and clinical literature that illuminates the variety of psychological and sexual sequelae of a breast cancer diagnosis and treatment, and the interventions that may aid in recovery. This article presents findings that highlight the impact on both patients and their family members. Recommendations are offered regarding the helpful role that physicians may play in mitigating the effects of some of the consequences of diagnosis and treatment. PMID- 12116606 TI - The effects of hormones on arrhythmias in women. AB - The reproductive hormones play a significant role in modulating the presentation and behavior of numerous types of arrhythmias. Differences exist between men and women in normal electrophysiology; along with heart rate, the QT interval is a basic parameter that is affected by the presence or absence of certain reproductive hormones. Women have a lower prevalence of atrial fibrillation, yet they have a higher mortality once atrial fibrillation is established. There are clear-cut gender differences in the prevalence of other re-entrant supraventricular tachycardias as well. Women are at lower risk of sudden cardiac death overall, but they have a higher risk of acquired long QT syndrome from antiarrhythmic drugs. It is possible that differences in myocardial repolarization modulated by sex hormones are a major factor in the differential presentation of these arrhythmias in men and women. PMID- 12116607 TI - The controversy over estrogen replacement therapy: an update on clinical trials. AB - This is a review and presentation of recent clinical trials designed to ascertain the effects of estrogen or estrogen plus progesterone on the risks of heart disease. The framework of the epidemiologic evidence that estrogen is cardioprotective is reviewed and the impact of these data on apparent findings from clinical trials discussed. The Heart and Estrogen/Progestin Replacement Study is examined in detail, and the most frequent criticisms of its findings are presented. Findings from other clinical trials are presented and the clinical implications from the data discussed in relation to the larger body of literature pertaining to hormone replacement therapy and heart disease. PMID- 12116609 TI - The hypnosis community can be compared, in many ways, with a large choral society. PMID- 12116608 TI - Anticoagulation during pregnancy. AB - Anticoagulation of a pregnant woman is a complex issue for both the treating physician and the patient. In patients with mechanical prosthetic valves, long term anticoagulation is mandatory to prevent thromboembolic complications; and in those with thrombophilic disorders and history of thromboembolism, anticoagulation is strongly indicated. With an increase in the number of patients with prosthetic heart valves, as well as the increase in maternal age, the issue of anticoagulation has become a very important one. Despite the widespread use of warfarin and unfractionated heparin during pregnancy, the optimal use of anticoagulants during pregnancy remains controversial because of a lack of appropriate prospective randomized clinical trials. In fact, even retrospective data on heparin provide miserably inadequate information for those making a decision on the correct dosing regimen. More recently, low molecular weight heparin has been proposed as a safer method of anticoagulation. This review summarizes current data and recommendations on anticoagulation during pregnancy. PMID- 12116610 TI - The role of hypnosis in the detection of psychogenic seizures. AB - In this preliminary clinical investigation, hypnosis was used in the differential diagnosis of epileptic versus psychogenic seizures (PS). Eight patients with a clinical profile suggesting the presence of PS were given a hypnotic suggestion in which they had to go back in time to the exact moment of their last seizure. They were then asked to concentrate their attention on any unusual feeling or bodily sensation. All 8 patients presented a PS during the age regression protocol. In 6 cases, independent testimony from family members corroborated the morphological similarity of the induced attack and the ones presented in their natural environment. Also, the seizures ended abruptly after a command was given to stop them. A control group of 5 epileptic subjects did not present any signs of discomfort or seizure behavior during the hypnotic protocol. It is argued that a simple procedure as the one described in this investigation can be useful as a diagnostic tool in the differentiation of epileptic from PS attacks. PMID- 12116611 TI - No link between hypnotizability and the Self-Monitoring Scale. AB - Socio-cognitive theorists have often claimed that hypnotizability is in part a function of social role-playing. We thus expected to find an association between a measure of hypnotizability (SHSS:C) and the Self-Monitoring Scale-Revised, a purported measure of sensitivity to social cues. The data failed to reveal any significant correlations, and therefore cannot be said to provide any support for the socio-cognitive position. Nevertheless, as hypothesized, we found that the hypnotist tended to consider subjects who were high but not low on the Other Directedness subscale to be more deeply hypnotized than the subjects themselves felt. PMID- 12116612 TI - The search for Bridey Murphy: implications for modern hypnosis. AB - The 1956 publication of The Search for Bridey Murphy was a noteworthy event for the field of hypnosis. This internationally best selling book, written for lay readers, described several recorded sessions of alleged time-regression to a prior life nearly two centuries before 1956. While subsequent investigations disproved that claim, there were a number of important implications for the science and practice of hypnosis. Although it was concluded that the Bridey Murphy interviews were products of cryptomnesia, the book was a significant factor associated with a resurgence of public and professional interest in the modality. PMID- 12116613 TI - Hypnotherapy and refractory irritable bowel syndrome: a single case study. AB - The current study describes the successful administration of hypnotherapy with a subject suffering from refractory Irritable Bowel Syndrome (IBS) and Generalized Anxiety Disorder (GAD). The subject had suffered from IBS for 30 years and had unsuccessfully pursued multiple psychological treatments, both traditional and non-traditional. He was referred to the Center for Stress and Anxiety Disorders and commenced hypnotherapy directed primarily at the IBS symptoms. After 6 treatment sessions, his IBS symptomatology had improved 53%. He stopped treatment at that point and continued autohypnosis with the aid of treatment audiotapes provided by his therapist. Follow-up at 6 months indicated continued improvement (70%). A 2-year follow-up revealed an improvement of 38% in IBS symptomatology. Concurrent levels of depression and anxiety had also substantially decreased. Hypnotherapy is shown to be a viable, palatable, and enduring treatment option for an individual who had been refractory to many previous therapies. PMID- 12116614 TI - Medical hypnosis and quadruplets: a case report. AB - This case report presents a new association reaction and a new treatment for quadruplet pregnancies. The hypnotic interventions can increase clinical management of quadruplet pregnancy. It illustrates new insights into the treatment of quadruplet pregnancies, and it suggests useful future research. PMID- 12116615 TI - Hypnosis provoked pseudoseizures: a case report and literature review. AB - Only a few studies have been reported in which suggestion was used to provoke pseudoseizures (PS). In these studies PS were video EEG monitored, and saline injections were administered as placebo. This method may be somewhat unethical and carries a low success rate. The authors, two child psychiatrists (GZ and DS) and a neurologist (NG), applied hypnosis to provoke PS which were monitored by video-EEG. Pre-, intra- and post-ictal serum prolactin levels were determined. The first hypnotic session was diagnostic and for this reason featured controlled attempts to determine whether there might be childhood trauma material. The following sessions easily provoked PS during which the EEG was normal and pre-, intra- and post-ictal serum prolactin levels were identical and within normal values. We conclude that hypnosis (with informed consent) for PS monitored by video-EEG telemetry, seems to be an ethical, cheap and quite easy way to demonstrate PS. PMID- 12116616 TI - What does your practice do to make patient encounters more efficient? PMID- 12116617 TI - Subspecialize or not? Look at what fits your practice. Yea/highly focused physicians provide superior patient care. PMID- 12116618 TI - Subspecialize or not? Look at what fits your practice. Nay/subspecializing submits a practice to inflexibility, lost skills. PMID- 12116619 TI - Medical liability reform bill introduced. PMID- 12116620 TI - CMS proposes two changes to the transaction standard and finalizes the employer ID. PMID- 12116621 TI - Electronic medical risks. EMR and management responsibility. PMID- 12116622 TI - Be safe, not sorry. Government regulations require safety devices in physicians' offices. PMID- 12116623 TI - Shall we dance? Physician/administrator teams can step in harmony. PMID- 12116624 TI - Let your fingers do the walking. Handheld technology puts formularies at your fingertips. PMID- 12116625 TI - Demand strong for many subspecialists. Compensation, benefits increase for hard to-find doctors. PMID- 12116626 TI - Medical merge. Alternative medicine case study. PMID- 12116628 TI - Ducks in a row. Aligning incentives to improve profitability. PMID- 12116627 TI - Compliance--what's good about a necessary evil? PMID- 12116629 TI - Show me the money. One practice's experience with employee bonuses. PMID- 12116630 TI - Children. The real victims of our nation's insurance crisis. PMID- 12116631 TI - Six months to improved cash flow and productivity. A physician practice case study. PMID- 12116632 TI - A wish list for systematic biology. PMID- 12116633 TI - Exploring data interaction and nucleotide alignment in a multiple gene analysis of Ips (Coleoptera: Scolytinae). AB - The possibility of gene tree incongruence in a species-level phylogenetic analysis of the genus Ips (Coleoptera: Scolytidae) was investigated based on mitochondrial 16S rRNA (16S) and nuclear elongation factor-1 alpha (EF-1 alpha) sequences, and existing cytochrome oxidase I (COI) and nonmolecular data sets. Separate cladistic analyses of the data partitions resulted in partially discordant most-parsimonious trees but revealed only low conflict of the phylogenetic signal. Interactions among data partitions, which differed in the extent of sequence divergence (COI > 16S > EF-1 alpha), base composition, and homoplasy, revealed that much of the branch support emerges only in the simultaneous analysis, particularly for deeper nodes in the tree, which are almost entirely supported through "hidden support" (sensu Gatesy et al., Cladistics 15:271-313, 1999). Apparent incongruence between data partitions is in part due to suboptimal alignments and bias of character transformations, but little evidence supports invoking incongruent phylogenetic histories of genetic loci. There is also no justification for eliminating or downweighting gene partitions on the basis of their apparent homoplasy or incongruence with other partitions, because the signal emerges only in the interaction of all data. In comparison with traditional taxonomy, the pini, plastographus, and perturbatus groups are polyphyletic, whereas the grandicollis group is monophyletic except for inclusion of the (monophyletic) calligraphus group. The latidens group and some European species are distantly related and closer to other genera within Ipini. Our robust cladogram was used to revise the classification of Ips. We provide new diagnoses for Ips and four subgeneric taxa. PMID- 12116634 TI - Phylogeny of Trichoptera (caddisflies): characterization of signal and noise within multiple datasets. AB - Trichoptera are holometabolous insects with aquatic larvae that, together with the Lepidoptera, make up the Amphiesmenoptera. Despite extensive previous morphological work, little phylogenetic agreement has been reached about the relationship among the three suborders--Annulipalpia, Spicipalpia, and Integripalpia--or about the monophyly of Spicipalpia. In an effort to resolve this conflict, we sequenced fragments of the large and small subunit nuclear ribosomal RNAs (1078 nt; D1, D3, V4-5), the nuclear elongation factor 1 alpha gene (EF-1 alpha; 1098 nt), and a fragment of mitochondrial cytochrome oxidase I (COI; 411 nt). Seventy adult and larval morphological characters were reanalyzed and added to molecular data in a combined analysis. We evaluated signal and homoplasy in each of the molecular datasets and attempted to rank the particular datasets according to how appropriate they were for inferring relationships among suborders. This evaluation included testing for conflict among datasets, comparing tree lengths among alternative hypotheses, measuring the left-skew of tree-length distributions from maximally divergent sets of taxa, evaluating the recovery of expected clades, visualizing whether or not substitutions were accumulating with time, and estimating nucleotide compositional bias. Although all these measures cast doubt on the reliability of the deep-level signal coming from the nucleotides of the COI and EF-1 alpha genes, these data could still be included in combined analyses without overturning the results from the most conservative marker, the rRNA. The different datasets were found to be evolving under extremely different rates. A site-specific likelihood method for dealing with combined data with nonoverlapping parameters was proposed, and a similar weighting scheme under parsimony was evaluated. Among our phylogenetic conclusions, we found Annulipalpia to be the most basal of the three suborders, with Spicipalpia and Integripalpia forming a clade. Monophyly of Annulipalpia and Integripalpia was confirmed, but the relationships among spicipalpians remain equivocal. PMID- 12116636 TI - Molecular data indicate the protostome affinity of brachiopods. AB - Although the phylogenetic position of brachiopods has always been subject to debate, many authors place them as a sister group to deuterostomes on the basis of morphological and developmental characters. However, molecular phylogeny consistently places them among protostomes. More precisely, brachiopods are predicted to branch inside the lophotrochozoan assemblage, together with annelids, molluscs, nemerteans, flatworms, and others. That result has been criticized on the basis of (1) prior knowledge of brachiopod morphology and (2) the known limitations of molecular phylogenies. Here I review recent data of molecular origin, particularly those displaying qualitative properties close to those of morphological characters. The complement of Hox genes present in all metazoa tested to date has proved to be a powerful tool for broad phylogenetic reconstruction. The mitochondrial genome also provides qualitative characters, showing discrete events of gene rearrangements. After discussing the data and the way they should be interpreted in the perspective of several hypotheses for metazoan phylogeny, I conclude that they argue strongly in favor of the protostome (and lophotrochozoan) affinity of the brachiopods. There is therefore a need for a reinterpretation of brachiopod morphological and developmental characters. I also identify some research axes on brachiopod morphology. PMID- 12116635 TI - Phylogeny of Saxifragales (angiosperms, eudicots): analysis of a rapid, ancient radiation. AB - Rapid, ancient radiations pose one of the most difficult challenges for phylogenetic estimation. We used DNA sequence data of 9,006 aligned base pairs from five genes (chloroplast atpB, matK, rbcL, and 18S and 26S nrDNA) to elucidate relationships among major lineages of Saxifragales (angiosperms, eudicots). These relationships were poorly supported in previous studies, apparently because the lineages originated in rapid succession. Using an array of methods that explicitly incorporate assumptions about evolutionary process (weighted maximum parsimony, maximum likelihood, LogDet/paralinear transformed distances), we show that the initial diversification of Saxifragales was indeed rapid. We suggest that the poor resolution of our best phylogenetic estimate is not due to violations of assumptions or to combining data partitions having conflicting histories or processes. We show that estimated branch lengths during the initial diversification are exceedingly short, and we estimate that acquiring sufficient sequence data to resolve these relationships would require an extraordinary effort (approximately 10(7) bp), assuming a linear increase in branch support with branch length. However, our simulation of much larger data sets containing a distribution of phylogenetic signal similar to that of the five sampled gene sequences suggests a limit to achievable branch support. Using statistical tests of differences in the likelihoods of topologies, we evaluated whether the initial radiation of Saxifragales involved the simultaneous origin of major lineages. Our results are consistent with predictions that resolving the branching order of rapid, ancient radiations requires sampling characters that evolved rapidly at the time of the radiation but have since experienced a slower evolutionary rate. PMID- 12116638 TI - War and peace in phylogenetics: a rejoinder on total evidence and consensus. AB - For more than 10 years, systematists have been debating the superiority of character or taxonomic congruence in phylogenetic analysis. In this paper, we demonstrate that the competing approaches can converge to the same solution when a consensus method that accounts for branch lengths is selected. Thus, we propose to use both methods in combination, as a way to corroborate the results of combined and separate analyses. This so-called "global congruence" approach is tested with a wide variety of examples sampled from the literature, and the results are compared with those obtained by standard consensus methods. Our analyses show that when the total evidence and consensus trees differ topologically, collapsing weakly supported nodes with low bootstrap support usually improves "global congruence". PMID- 12116637 TI - Combining and comparing morphometric shape descriptors with a molecular phylogeny: the case of fruit type evolution in Bornean Lithocarpus (Fagaceae). AB - Fruit type in the genus Lithocarpus (Fagaceae) includes both classic oak acorns and novel modifications. Bornean taxa with modified fruits can be separated into two sections (Synaedrys and Lithocarpus) based on subtle shape differences. By following strict criteria for homology and representation, this variation in shape can be captured and the sections distinguished by using elliptic Fourier or eigenshape analysis. Phenograms of fruit shape, constructed by using restricted maximum likelihood techniques and these morphometric descriptors, were incorporated into combined and comparative analyses with molecular sequence data from the internal transcribed spacer (ITS) region of the nuclear rDNA, using branch-weighted matrix representation. The combined analysis strongly suggested independent derivation of the novel fruit type in the two sections from different acornlike ancestors, while the comparative analysis indicated frequent decoupling between the molecular and morphological changes as inferred at well-supported nodes. The acorn fruit type has undergone little modification between ingroup and outgroup, despite large molecular distance. Greater morphological than molecular change was inferred at critical transitions between acorn and novel fruit types, particularly for section Lithocarpus. The combination of these two different types of data improved our understanding of the macroevolution of fruit type in this difficult group, and the comparative analysis highlighted the significant incongruities in evolutionary pattern between the two datasets. PMID- 12116640 TI - A likelihood approach to estimating phylogeny from discrete morphological character data. AB - Evolutionary biologists have adopted simple likelihood models for purposes of estimating ancestral states and evaluating character independence on specified phylogenies; however, for purposes of estimating phylogenies by using discrete morphological data, maximum parsimony remains the only option. This paper explores the possibility of using standard, well-behaved Markov models for estimating morphological phylogenies (including branch lengths) under the likelihood criterion. An important modification of standard Markov models involves making the likelihood conditional on characters being variable, because constant characters are absent in morphological data sets. Without this modification, branch lengths are often overestimated, resulting in potentially serious biases in tree topology selection. Several new avenues of research are opened by an explicitly model-based approach to phylogenetic analysis of discrete morphological data, including combined-data likelihood analyses (morphology + sequence data), likelihood ratio tests, and Bayesian analyses. PMID- 12116639 TI - On areas of endemism, with an example from the African Restionaceae. AB - Areas of endemism are central to cladistic biogeography. The concept has been much debated in the past, and from this has emerged the generally accepted definition as an area to which at least two species are endemic. Protocols for locating areas of endemism have been neglected, and to date no attempt has been made to develop optimality criteria against which to evaluate competing hypotheses of areas of endemism. Here various protocols for finding areas of endemism are evaluated--protocols based on both phonetic and parsimony analyses, on both unweighted data and data weighted by various criteria. The optimality criteria used to compare the performance of the methods include the number of species included in the areas of endemism, the number of areas delimited, and the degree of distributional congruency of the species restricted to each area of endemism. These methods are applied to the African Restionaceae in the Cape Floristic Region. Parsimony methods using weighted data are shown to perform best on the combination of all three optimality criteria. By varying the weighting parameters, the size of the areas of endemism can be varied. This provides a very useful tool for locating areas of endemism that satisfy prespecified scale criteria. PMID- 12116641 TI - Unraveling the evolutionary radiation of the families of the Zingiberales using morphological and molecular evidence. AB - The Zingiberales are a tropical group of monocotyledons that includes bananas, gingers, and their relatives. The phylogenetic relationships among the eight families currently recognized are investigated here by using parsimony and maximum likelihood analyses of four character sets: morphological features (1), and sequence data of the (2) chloroplast rbcL gene, (3) chloroplast atpB gene, and (4) nuclear 18S rDNA gene. Outgroups for the analyses include the closely related Commelinaceae + Philydraceae + Haemodoraceae + Pontederiaceae + Hanguanaceae as well as seven more distantly related monocots and paleoherbs. Only slightly different estimates of evolutionary relationships result from the analysis of each character set. The morphological data yield a single fully resolved most-parsimonious tree. None of the molecular datasets alone completely resolves interfamilial relationships. The analyses of the combined molecular dataset provide more resolution than do those of individual genes, and the addition of the morphological data provides a well-supported estimate of phylogenetic relationships: (Musaceae ((Strelitziaceae, Lowiaceae) (Heliconiaceae ((Zingiberaceae, Costaceae) (Cannaceae, Marantaceae))))). Evidence from branch lengths in the parsimony analyses and from the fossil record suggests that the Zingiberales originated in the Early Cretaceous and underwent a rapid radiation in the mid-Cretaceous, by which time most extant family lineages had diverged. PMID- 12116642 TI - Sequence alignment of 18S ribosomal RNA and the basal relationships of Adephagan beetles: evidence for monophyly of aquatic families and the placement of Trachypachidae. AB - Current hypotheses regarding family relationships in the suborder Adephaga (Coleoptera) are conflicting. Here we report full-length 18S ribosomal RNA sequences of 39 adephagans and 13 outgroup taxa. Data analysis focused on the impact of sequence alignment on tree topology, using two principally different approaches. Tree alignments, which seek to minimize indels and substitutions on the tree in a single step, as implemented in an approximate procedure by the computer program POY, were contrasted with a more traditional procedure based on alignments followed by phylogenetic inference based on parsimony, likelihood, and distance analyses. Despite substantial differences between the procedures, phylogenetic conclusions regarding basal relationships within Adephaga and relationships between the four suborders of Coleoptera were broadly similar. The analysis weakly supports monophyly of Adephaga, with Polyphaga usually as its sister, and the two small suborders Myxophaga and Archostemata basal to them. In some analyses, however, Polyphaga was reconstructed as having arisen from within Hydradephaga. Adephaga generally split into two monophyletic groups, corresponding to the terrestrial Geadephaga and the aquatic Hydradephaga, as initially proposed by Crowson in 1955, consistent with a single colonization of the aquatic environment by adephagan ancestors and contradicting the recent proposition of three independent invasions. A monophyletic Hydradephaga is consistently, though not strongly, supported under most analyses, and a parametric bootstrapping test significantly rejects an hypothesis of nonmonophyly. The enigmatic Trachypachidae, which exhibit many similarities to aquatic forms but whose species are entirely terrestrial, were usually recovered as a basal lineage within Geadephaga. Strong evidence opposes the view that terrestrial trachypachids are related to the dytiscoid water beetles. PMID- 12116643 TI - Complexity of the likelihood surface for a large DNA dataset. PMID- 12116644 TI - Difficulties in detecting hybridization. PMID- 12116645 TI - The troubled growth of statistical phylogenetics. PMID- 12116646 TI - The evolutionary biology of ... everything. PMID- 12116647 TI - Environmental diversity of bacteria and archaea. AB - The microbial way of life spans at least 3.8 billion years of evolution. Microbial organisms are pervasive, ubiquitous, and essential components of all ecosystems. The geochemical composition of Earth's biosphere has been molded largely by microbial activities. Yet, despite the predominance of microbes during the course of life's history, general principles and theory of microbial evolution and ecology are not well developed. Until recently, investigators had no idea how accurately cultivated microorganisms represented overall microbial diversity. The development of molecular phylogenetics has recently enabled characterization of naturally occurring microbial biota without cultivation. Free from the biases of culture-based studies, molecular phylogenetic surveys have revealed a vast array of new microbial groups. Many of these new microbes are widespread and abundant among contemporary microbiota and fall within novel divisions that branch deep within the tree of life. The breadth and extent of extant microbial diversity has become much clearer. A remaining challenge for microbial biologists is to better characterize the biological properties of these newly described microbial taxa. This more comprehensive picture will provide much better perspective on the natural history, ecology, and evolution of extant microbial life. PMID- 12116648 TI - Catalyzing bacterial speciation: correlating lateral transfer with genetic headroom. AB - Unlike crown eukaryotic species, microbial species are created by continual processes of gene loss and acquisition promoted by horizontal genetic transfer. The amounts of foreign DNA in bacterial genomes, and the rate at which this is acquired, are consistent with gene transfer as the primary catalyst for microbial differentiation. However, the rate of successful gene transfer varies among bacterial lineages. The heterogeneity in foreign DNA content is directly correlated with amount of genetic headroom intrinsic to a bacterial species. Genetic headroom reflects the amount of potentially dispensable information- reflected in codon usage bias and codon context bias--that can be transiently sacrificed to allow experimentation with functions introduced by gene transfer. In this way, genetic headroom offers a potential metric for assessing the propensity of a lineage to speciate. PMID- 12116649 TI - Genomic and phylogenetic perspectives on the evolution of prokaryotes. AB - Prokaryotes have been at the forefront of the genome sequencing revolution. Many genomes have been completely sequenced, revealing much about bacterial and archaeal genome content and organization. Yet, a meaningful evolutionary picture of prokaryotes still eludes us. Much of the problem lies in understanding the mode and tempo of genome evolution. Here phenylalanyl-tRNA synthetase is used as an example of the complex interplay among lateral gene transfer, operon recombination, and gene recruitment in the evolution of some prokaryotic genes. Promising new approaches to genomic analyses, which could add to our understanding prokaryotic evolution and help in their classification, are discussed. PMID- 12116650 TI - Bacterial species and speciation. AB - Bacteria are profoundly different from eukaryotes in their patterns of genetic exchange. Nevertheless, ecological diversity is organized in the same way across all of life: individual organisms fall into more less discrete clusters on the basis of their phenotypic, ecological, and DNA sequence characteristics. Each sequence cluster in the bacterial world appears to correspond to an "ecotype," defined as a population of cells in the same ecological niche, which would all be out-competed by any adaptive mutant coming from the population. Ecotypes, so defined, share many of the dynamic properties attributed to eukaryotic species: genetic diversity within an ecotype is limited by a force of cohesion (in this case, periodic selection); different ecotypes are free to diverge without constraint from one another; and ecotypes are ecologically distinct. Also, ecotypes can be discovered and classified as DNA sequence clusters, even when we are ignorant of their ecology. Owing to the rarity and promiscuity of bacterial genetic exchange, speciation in the bacterial world is expected to be much less constrained than in the world of animals and plants. PMID- 12116651 TI - Bias in phylogenetic estimation and its relevance to the choice between parsimony and likelihood methods. PMID- 12116652 TI - Toward understanding Anophelinae (Diptera, Culicidae) phylogeny: insights from nuclear single-copy genes and the weight of evidence. AB - A phylogeny of the mosquito subfamily Anophelinae was inferred from fragments of two protein-coding nuclear genes, G6pd (462 bp) and white (801 bp), and from a combined data set (2,136 bp) that included a portion of the mitochondrial gene ND5 and the D2 region of the ribosomal 28S gene. Sixteen species from all three anopheline genera and six Anopheles subgenera were sampled, along with six species of other mosquitoes used as an outgroup. Each of four genes analyzed individually recovered the same well-supported clades; topological incongruence was limited to unsupported or poorly supported nodes. As assessed by the incongruence length difference test, most of the conflicting signal was contributed by third codon positions. Strong structural constraints, as observed in white and G6pd, apparently had little impact on phylogenetic inference. Compared with the other genes, white provided a superior source of phylogenetic information. However, white appears to have experienced accelerated rates of evolution in few lineages, the affinities of which are therefore suspect. In combined analyses, most of the inferred relationship were well-supported and in agreement with previous studies: monophyly of Anophelinae, basal position of Chagasia, monophyly of Anopheles subgenera, and subgenera Nyssorhynchus + Kerteszia as sister taxa. The results suggested also monophyletic origin of subgenera Cellia + Anopheles, and the white gene analysis supported genus Bironella as a sister taxon to Anopheles. The present data and other available evidence suggest a South American origin of Anophelinae, probably in the Mesozoic; a rapid diversification of Bironella and basal subgeneric lineages of Anopheles, potentially associated with the breakup of Gondwanaland; and a relatively recent and rapid dispersion of subgenus Anopheles. PMID- 12116653 TI - Ancestral state estimation and taxon sampling density. AB - A set of experiments based on simulation and analysis found that using the parsimony algorithm for ancestral state estimation can benefit from increased sampling of terminal taxa. Estimation at the base of small clades showed strong sensitivity to tree topology and number of descendent tips. These effects were largely driven by the creation and negation of ambiguity across a topology. Root state and internal state estimation showed similar behavior. We conclude that increased taxon sampling density is generally advisable, and attention to topological effects may be advisable in evaluating the confidence placed in state estimation. We also explore the factors affecting ancestral state estimation and conjecture that as taxa are added to a tree, the total amount of information for root state estimation depends on the tree topology and distance to root state of added taxa. For a pure-birth model tree, we conjecture that the addition of N taxa increases root state information in proportion to log(N). PMID- 12116654 TI - Assessment of the accuracy of matrix representation with parsimony analysis supertree construction. AB - Despite the growing popularity of supertree construction for combining phylogenetic information to produce more inclusive phylogenies, large-scale performance testing of this method has not been done. Through simulation, we tested the accuracy of the most widely used supertree method, matrix representation with parsimony analysis (MRP), with respect to a (maximum parsimony) total evidence solution and a known model tree. When source trees overlap completely, MRP provided a reasonable approximation of the total evidence tree; agreement was usually > 85%. Performance improved slightly when using smaller, more numerous, or more congruent source trees, and especially when elements were weighted in proportion to the bootstrap frequencies of the nodes they represented on each source tree ("weighted MRP"). Although total evidence always estimated the model tree slightly better than nonweighted MRP methods, weighted MRP in turn usually out-performed total evidence slightly. When source studies were even moderately nonoverlapping (i.e., sharing only three-quarters of the taxa), the high proportion of missing data caused a loss in resolution that severely degraded the performance for all methods, including total evidence. In such cases, even combining more trees, which had positive effects elsewhere, did not improve accuracy. Instead, "seeding" the supertree or total evidence analyses with a single largely complete study improved performance substantially. This finding could be an important strategy for any studies that seek to combine phylogenetic information. Overall, our results suggest that MRP supertree construction provides a reasonable approximation of a total evidence solution and that weighted MRP should be used whenever possible. PMID- 12116655 TI - Selecting the best-fit model of nucleotide substitution. AB - Despite the relevant role of models of nucleotide substitution in phylogenetics, choosing among different models remains a problem. Several statistical methods for selecting the model that best fits the data at hand have been proposed, but their absolute and relative performance has not yet been characterized. In this study, we compare under various conditions the performance of different hierarchical and dynamic likelihood ratio tests, and of Akaike and Bayesian information methods, for selecting best-fit models of nucleotide substitution. We specifically examine the role of the topology used to estimate the likelihood of the different models and the importance of the order in which hypotheses are tested. We do this by simulating DNA sequences under a known model of nucleotide substitution and recording how often this true model is recovered by the different methods. Our results suggest that model selection is reasonably accurate and indicate that some likelihood ratio test methods perform overall better than the Akaike or Bayesian information criteria. The tree used to estimate the likelihood scores does not influence model selection unless it is a randomly chosen tree. The order in which hypotheses are tested, and the complexity of the initial model in the sequence of tests, influence model selection in some cases. Model fitting in phylogenetics has been suggested for many years, yet many authors still arbitrarily choose their models, often using the default models implemented in standard computer programs for phylogenetic estimation. We show here that a best-fit model can be readily identified. Consequently, given the relevance of models, model fitting should be routine in any phylogenetic analysis that uses models of evolution. PMID- 12116656 TI - Finding fault with vicariance: a critique of Heads (1998). PMID- 12116658 TI - Misleading results from the use of ambiguity coding to score polymorphisms in higher-level taxa. PMID- 12116657 TI - Efficiency of strict consensus trees. PMID- 12116659 TI - Monitoring doctors' clinical competence: a Queensland focus. AB - The medical profession is held accountable in numerous ways, many of which are concerned with clinical competence. However, while Australia's State medical boards are statutorily charged with protecting the public from incompetent practice, they have never instituted programs aimed at maintaining the standards of all practitioners. The article describes recent legislative changes and developments in undergraduate medical education, which aim to increase physicians' accountability in relation to competence, and compares developments in Queensland and other States with those in the United Kingdom, Quebec and New Zealand. The investigation of clinical incompetence as currently undertaken by the Medical Board of Queensland should, and will, be adopted in other States. However, responding to incompetence is necessarily piecemeal, and the article further argues that the States should develop inclusive revalidation strategies. Both the community and leading voices within the profession are demanding greater commitment to a self-regulation culture that is more transparent and has sharper teeth. PMID- 12116660 TI - Liability of health professionals for a breach of the abortion law of New Zealand. AB - The abortion law of New Zealand appears to have been interpreted very liberally over recent years by sectors of the medical profession. Indeed the interpretation of the law appears to have been so liberal that it raises questions as to the lawfulness of many of the abortions carried out in New Zealand. The current practice and application of the abortion law is such that it may expose some medical consultants certifying and performing abortions to criminal proceedings and civil claims. PMID- 12116661 TI - Access to infertility treatments and single women: what is the state of play? PMID- 12116662 TI - Human cloning offences: the gene Technology Act 2000 (Cth). PMID- 12116663 TI - Suicide and the media. PMID- 12116664 TI - Therapeutic drugs and death. PMID- 12116665 TI - Re A (children) (conjoined twins: surgical separation) [2001] 2 WLR 480. PMID- 12116666 TI - Rosenberg v Percival (2001) 75 ALJR 734; [2001] HCA 18. PMID- 12116667 TI - Rajan v General Medical Council [2000] Lloyd's Rep Med 153; [2000] UKPC 1. PMID- 12116668 TI - Drake v Pontefract Health Authority [1998] Lloyd's Rep Med 425. PMID- 12116669 TI - Prenatal diagnosis, genetics and reproductive decision-making. AB - Recent developments in genetic science will potentially have a significant impact on reproductive decision-making by adding to the list of conditions which can be diagnosed through prenatal diagnosis. This article analyses the jurisdictional variations that exist in Australian abortion laws and examines the extent to which Australian abortion laws specifically provide for termination of pregnancy on the grounds of fetal disability. The article also examines the potential impact of pre-implantation genetic diagnosis on reproductive decision-making and considers the meaning of reproductive autonomy in the context of the new genetics. PMID- 12116670 TI - The regulation of nursing in Australia: a comparative analysis. AB - This article uses the International Council of Nurses framework for nursing regulation to examine Nurses Acts in Australian States and Territories. It measures their compliance with the standards contained in the framework and exposes the anomalies among jurisdictions in spite of mutual recognition legislation now in force in all jurisdictions, including the Commonwealth. It also provides examples of difficulties encountered with cross-border practice and concludes that there is an urgent need for national nursing legislation. PMID- 12116671 TI - Rogers v Whitaker reconsidered. PMID- 12116672 TI - Exorcising excision: medico-legal issues arising from male and female genital surgery in Australia. AB - Genital surgery is one of the most controversial and contested practices, yet it is frequently described and referred to with little or no attention to cultural and social context. This article examines the practice, performed on both men and women, and the extent to which it clashes with issues of consent and capacity, as well as multicultural concepts of toleration for minority group practices. It then questions why female genital surgery, unlike male genital surgery, is legally prohibited in Australia. It argues that such legal gender bias stems from a liberal conception of "tolerance" and the limits of consent in Australia, placing female genital surgery in an "unacceptable" category and male genital surgery in an "acceptable" category. PMID- 12116673 TI - Males, medical mutilation and the law: some recent developments. AB - Two recent decisions of the English Court of Appeal--Re J (Specific Issue Orders: Child's Religious Upbringing and Circumcision) [2000] 1 FLR 571 and Re A (Male Sterilisation) [2000] 1 FLR 549--raise serious issues relating to controversial matters involving law and medicine and deserve wider consideration than in their jurisdiction of origin. This article discusses the practical and policy implications of these cases and places them in the appropriate context. PMID- 12116674 TI - Ethical considerations underpinning the donation of live, non-regenerative organs. AB - The recent donation of a kidney to one of Australia's most prominent citizens by a long-time friend and employee has brought to attention the problems of access facing patients who require renal transplantation as a life-saving measure. The lack of availability of cadaver organs, the improved techniques available to minimise tissue rejection and the potential to genetically engineer tissue compatible individuals for future organ donation have generated an interest in the ethical and legal considerations that underlie live organ donation. PMID- 12116675 TI - Consent, commercialisation and benefit-sharing. AB - This article considers two issues in relation to the increasing commercial exploitation of biological materials: first, whether the consent of the individual who is the source of biological material is needed for the material to be used commercially; and secondly, whether the source has any right to share in the profits. Whilst the legal requirement for consent to commercial use is uncertain, the overwhelming view from ethical statements suggests that such consent is appropriate. With regard to profit-sharing, source individuals have no legal right to share in the profits of commercial exploitation of their sample. However, there is support in some ethical statements for benefit-sharing with source individuals and their communities. PMID- 12116676 TI - Doctors as fiduciaries--revisiting the past with an eye on the future. AB - A substantial proportion of the body of literature dealing with the question of whether or not a doctor stands in a fiduciary relationship with a patient in Australia assumes or asserts that this should be the case, despite strong indications to the contrary in Australian case law. Three key bases for making such assertions, the internationalist, revisionist and remedialist approaches, are identified and critiqued. It is argued that each of these approaches to justifying the characterisation of the doctor-patient relationship as a fiduciary one is flawed and unlikely to meet with success in future litigation. Additionally, there are issues of economic and resource allocation conflict in doctor-patient relationships. The implications for these conflicts in the doctor patient fiduciary debate are briefly considered. It is concluded that, contrary to the dominant assertion in the extant literature on the subject, in Australia at least, the scales tip against, rather than towards, the characterisation of the doctor-patient relationship as a fiduciary one. PMID- 12116677 TI - [Focus on physicians' clinical skills]. PMID- 12116678 TI - [Early statin treatment in acute coronary syndrome. Is this evidence-based?]. AB - Lipid lowering treatment with 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitor (statins) may reduce morbidity and mortality in patients with stable ischaemic heart disease. In trials where a statin was used as secondary prevention after an acute coronary syndrome (ACS), the treatment was initiated 3 6 months after the event. A high rate of serious complications occurs in the unstable clinical state after an ACS, including myocardial infarction and death. The incidence rate of serious complications declines after 1 month and then gradually diminishes. It has not been determined whether initiation of treatment with a statin soon after an ACS reduces the occurrence of early coronary events. Statins have effects on the vessel wall other than the lipid lowering effect, and these may partly account for the benefit of statin therapy. Whether these mechanisms are associated with a prognostic effect soon after an ACS has not been clarified. PMID- 12116679 TI - [Brain metastases]. AB - The incidence of symptomatic brain metastases in Denmark is about 3500. In the present review, the aetiology, symptomatology, and diagnostic procedures are described. The main topic is a review of current treatments and the evidence for their efficacy. Treatment of brain metastases rarely cures the patient, the goal is rather to improve the quality of life and prolong survival. Without treatment, the median survival following diagnosis of brain metastases is about one month, with steroid treatment two months, with whole brain irradiation four to six months, and after surgery or stereotactic radiosurgery 10-12 months. A relatively simple treatment scheme based on the number of brain metastases and the overall condition of the patient is provided. PMID- 12116680 TI - [Osteoclast function is regulated by neighbouring osteoblasts. Osteoprotegerin, RAND and RANK ligand constitute a unique regulatory system for bone resorption with important pathophysiological and therapeutic aspects]. AB - Maturation of macrophages to osteoclasts requires the presence of marrow stromal cells or osteoblasts. Most calcitropic hormones act indirectly on osteoclasts through receptors on neighbouring osteoblasts. The discovery of osteoprotegerin (OPG), the receptor activator of nuclear factor-kappa b ligand (RANKL), and its receptor (RANK) has elucidated these phenomena. It appears that osteoclast differentiation, activity, and survival are regulated by the proportion of inhibiting OPG to stimulating RANKL. OPG and RANKL are produced by osteoblasts, whereas RANK is located to the osteoclasts. Treatment with OPG inhibits bone resorption in postmenopausal women. Mutations in the system may be responsible for focal skeletal disorders. The discovery opens up for new treatment opportunities in postmenopausal and steroid-induced osteoporosis, Paget's disease, hypercalcaemia, and rheumatoid arthritis. PMID- 12116681 TI - [Expected practical clinical skills and newly qualified physicians]. AB - INTRODUCTION: We compared and contrasted the learned and intended practical clinical skills of the three Danish medical schools. MATERIAL AND METHODS: An anonymous questionnaire listing 210 practical clinical skills was posted to 226 newly qualified Danish physicians. They were asked if they could meet the minimum level for each of the skills, which had earlier been identified by a Delphi study. RESULTS: The response rate was 80%. None of the responders met the minimum of all the 210 skills. Only 8% (14) met the minimum level for at least 90% (189) of the skills. On average, the responders met the minimum for 74% (155) of the skills. More than 90% of the responders could take medical histories and carry out examinations. The responders did not meet 28 medical emergency procedures. DISCUSSION: We found that the clinical skills learned constituted 75% of those intended. Those responsible for pre- and postgraduate medical training should be aware of the discrepancy between expected and learned skills. We discuss the role of experts in the process of defining the core curriculum. PMID- 12116682 TI - [Secondary prophylaxis in general practice for patients with ischemic heart disease]. AB - INTRODUCTION: We examined the extent to which the Danish College of General Practitioners' guidelines of 1998, "Prevention of ischaemic heart disease in general practice--with special focus on dyslipidaemia", are complied with in the secondary preventive care of patients with ischaemic heart disease. MATERIAL AND METHOD: Twenty-six general practitioners from Ringkjobing County registered all patients with ischaemic heart disease consecutively over three weeks in May 1999. One hundred and ten patients had suffered acute myocardial infarction, 141 had ischaemic heart disease without acute myocardial infarction. Blood pressure, total cholesterol, and use of cardiovascular drugs were registered. RESULTS: In patients with previous acute myocardial infarction, 66% had had their serum cholesterol measured within the past year; 76% were being treated with acetylsalicylic acid, 46% with beta-blockers, and 27% with lipid-lowering drugs. Of the patients with previous myocardial infarction and a serum cholesterol level > or = 5.0 mmol/l, 33% took lipid-lowering drugs. DISCUSSION: The present study demonstrates that in some areas actual treatment practice does not always comply with the given recommendations. In particular the frequency of cholesterol measuring and treatment with beta-blockers and lipid-lowering drugs show that continued, targeted quality development is required. PMID- 12116683 TI - [Use of dipsticks and microscope in the diagnosis of hematuria in general practice]. AB - INTRODUCTION: Haematuria is a serious symptom and is often detected by dipstick analysis. We examined the use of dipsticks in general practice. MATERIALS AND METHODS: An anonymous questionnaire was sent to all GPs in a county in Denmark in April 2000. RESULTS: The response rate was 92%. All GPs used dipsticks for the detection of haematuria. Answers to the questionnaire were compared with the guidelines of the dipstick manufacturers, and we found that 80% handled the dipstick incorrectly. A microscope was available in 65% of practices, and 44% performed urine microscopy in the case of dipstick haematuria. DISCUSSION: The validity of dipsticks in the detection of haematuria is discussed, and we conclude that dipsticks must be handled strictly in accordance with the instructions of the manufacturer, so as to maintain a high rate of specificity and sensitivity. A positive dipstick should be followed up immediately with microscopy of the same urine sample. PMID- 12116684 TI - [Osteoporosis prevention in patients with low energy hip fractures hospitalized in the orthopedic department]. AB - INTRODUCTION: The aim of this study was to investigate the following in patients treated for low energy hip fractures: 1) whether they were using osteoporosis preventative therapy before admission to hospital; 2) whether therapy was initiated before discharge from hospital; 3) the therapy used; and 4) whether they were referred for follow-up treatment for osteoporosis after discharge. MATERIAL AND METHODS: We carried out a retrospective study of medical journals on patients with a hip fracture, brought into the Department of Orthopaedic Surgery in 1997-1998. RESULTS: One hundred and forty-six patients, 80% women and 20% men with an average age of 80.7 years, were hospitalised during this period. Twenty patients had specific risk factors for developing osteoporosis, five patients of whom were taking osteoporosis preventative therapy. On admission, eight patients were in treatment. One patient started treatment during the hospital stay. Forty percent of the patients on discharge were referred for treatment by their GP or at the Department of Internal Medicine. DISCUSSION: Of the patients brought into the Department of Orthopaedic Surgery with low energy hip fractures, 6.2% were given the relevant medical treatment for osteoporosis on discharge from hospital, and only one of these patients started osteoporosis preventative therapy during the stay. As the study shows that 14% of the patients had specific risk factors for developing osteoporosis, only 25% of whom started treatment before admission to hospital, we find a need for osteoporosis preventative therapy in this group of patients. PMID- 12116685 TI - [Bilateral necrosis of the femoral head after use of working harness]. AB - The case of a 42-year old man, who developed bilateral necrosis of the femoral head from traumatic causes is described. He hung in stirrups, working 4-6 hours a day decorating for three weeks, during which he complained of numbness, fatigue and radiating pain in both legs. X-rays and a bone scan were performed and bilateral femoral head necrosis was diagnosed, probably because of repeated temporary hypoxia of the femoral heads caused by the stirrups compressing the blood vessels. PMID- 12116686 TI - [Picture of the month: encrusted VIVA plug]. PMID- 12116688 TI - [Equal dialogue between the physician and the patient--or is the physician a God?]. PMID- 12116687 TI - [Can we rely on the results of urine microscopy?]. PMID- 12116689 TI - Establishing a gradient of risk in patients with acute coronary syndromes using troponin I measurements. AB - OBJECTIVE: To evaluate the role of serum troponin I (Tnl) estimations in the early risk stratification of patients with acute coronary syndromes (ACS) subsequently diagnosed as acute myocardial infarction (AMI) or unstable angina (UA). SUBJECTS AND METHODS: Blood samples were collected from 86 patients admitted to the Coronary Care Unit of the Mubarak Al-Kabeer Hospital, Kuwait, with a diagnosis of ACS on admission (Tnl-1) and after 8 h (Tnl-2) and 16 h (Tnl 3). Blood was also collected from 38 age-matched healthy controls for comparison. Serum Tnl was measured by paramagnetic particle chemiluminescent immunoassay. RESULTS: Serum Tnl of < 0.05 ng/ml, corresponding to the 99th percentile, was established for healthy subjects. Patients diagnosed as UA had a 99th percentile Tnl-1 value of about 0.30 ng/ml. The best specificity and sensitivity for ACS was obtained for Tnl-2; indeed, Tnl-2 > 0.3 ng/ml gave a > 80% certainty of diagnosis of AMI. Also, Tnl-2 < 0.3 ng/ml in ACS patients was approximately 80% sensitive for the diagnosis of UA but relatively nonspecific (approximately 40%). Specificity for Tnl-2 for the diagnosis of UA improved to about 90% by narrowing the diagnostic range to 0.05-0.3 ng/ml. Tnl values in UA increased by < 100% at 8 h, while in AMI, this increase was up to 1,000%. CONCLUSION: In the evaluation of ACS, admission and 8-hour serum Tnl < 0.05 ng/ml is probably not cardiac in origin; serum Tnl > 0.3 ng/ml on admission and increasing rapidly by 8 h is likely AMI, and serum Tnl > 0.05 and < 0.3 ng/ml on admission with a mild increase by 8 h is likely due to UA. PMID- 12116690 TI - Irritable bowel syndrome: update on pathogenesis and management. AB - Irritable bowel syndrome (IBS) comprises a major proportion of gastrointestinal and primary care practice worldwide. The past several years have seen the rapid evolution of a new and comprehensive model of IBS based on alterations in brain gut interactions. Alterations in the bidirectional communication between the enteric nervous system and the central nervous system are implicated in the pathogenesis of IBS. 5-Hydroxytryptamine (5-HT; serotonin), a major neurotransmitter in the gastrointestinal tract, and its receptors 5-HT3 and 5-HT4 are involved in the control of gastrointestinal function. A number of abnormal motor and sensory patterns have been reported in patients with IBS. However, it is not known whether these abnormalities are related to symptoms or have a role in establishing a diagnosis of functional gastrointestinal disorders. Visceral hyperalgesia in IBS patients can be secondary to altered receptor sensitivity at the viscus itself and altered central modulation of sensation involving psychological influences in the interpretation of these sensations. The development of diagnostic criteria for IBS helps to avoid unnecessary and costly investigations. A detailed history allows us to diagnose IBS and search for another cause if warning symptoms are present. The Rome criteria are presently used to define IBS and are currently the most widely applied criteria used in clinical diagnosis and research purposes. Abdominal pain or discomfort associated with chronic altered bowel habits are the mainstay in diagnosis, while the supportive criteria may be used to further classify IBS patients into diarrhea predominant or constipation-predominant subgroups. Minimal diagnostic tests have been advocated in the initial diagnostic approach to patients with suspected IBS, depending on the predominant symptom. The therapeutic goals in IBS must focus on the overall well-being of the patient, including abdominal symptoms and the accompanying nonbowel symptoms and affective disorders. It is important to establish an effective physician-patient relationship and to reassure the patient once the diagnosis of IBS is made. Dietary modification may be of value in some patients with IBS. Dietary fiber is frequently recommended for patients with constipation-predominant IBS. Two novel serotonin agonists are currently under development for constipated IBS patients, tegaserod and prucalopride. Antidiarrheal agents, including loperamide and diphenoxylate, may help patients with diarrhea-predominant IBS. 5-HT3 receptor antagonists may play a role in the management of such patients in the future. Psychological treatment and antidepressants should be considered when IBS symptoms are severe or refractory or associated with psychological distress and impaired quality of life. PMID- 12116691 TI - Hospital-acquired Clostridium difficile infection amongst ICU and burn patients in Kuwait. AB - OBJECTIVES: To prospectively study the prevalence of nosocomially acquired Clostridium difficile, a major cause of diarrhoea in hospitalized patients, in the intensive care units (ICUs) and burn unit (BUs) of three teaching hospitals in Kuwait. METHODS: During a 1-year prospective study, stool/rectal swabs were obtained from 344 patients admitted into the ICUs of Mubarak Hospital (ICU-1), the Kuwait Cancer Control Centre (ICU-2), and the BU of Ibn Sina Hospital. The presence of C. difficile and/or its toxin was detected by serially culturing the specimens on differential, selective and enriched media and the use of TOX-A/B test, on admission and at subsequent 1-weekly interval until discharge. RESULTS: Out of the 344 patients admitted into these units, over a study period of 1 year, only 263 (77%) were evaluable. All of them had negative stool culture/toxin on admission. Overall, 25 (9.5%) of these 263 patients acquired C. difficile during their hospitalization. Thirteen (7%) of 187 patients acquired C. difficile in ICU 1, 9 (36%) of 25 on ICU-2 and 3 (5.9%) of 51 patients in BU. Eight (32%) developed diarrhoea attributable only to C. difficile and/or toxin, and the remaining 17 (68%) were asymptomatic: none had pseudomembranous colitis. The diarrhoea in these patients was associated with antibiotic use, the main trigger antibiotics being the third-generation cephalosporins. Acquisition occurred within 4-53 days of admission, with the majority occurring in the first 15 days. CONCLUSION: Overall, the prevalence of hospital-acquired C. difficile infection/colonization was less than 10%. The use of third-generation cephalosporins was high and was related to the development of diarrhoea. Once acquired, diarrhoea developed in about one third of C. difficile-positive cases, an indication that C. difficile infection/colonization endemic in the hospital ICUs studied is usually transmitted among the hospitalized patients. PMID- 12116693 TI - The relationship between end tidal carbon dioxide and arterial carbon dioxide during controlled hypotensive anaesthesia. AB - OBJECTIVES: To prospectively assess the magnitude of changes in the arterial-to end tidal carbon dioxide gradient [P(a-ET)CO2] as well as in the ratio of physiological dead space to tidal volume (Vdphys/Vt) during controlled hypotensive anaesthesia, and to evaluate whether or not ventilatory requirements remain unaltered during this procedure. SUBJECTS AND METHODS: Twelve adult patients with American Society of Anesthesiologists' physical status I and II undergoing middle ear surgery were selected. A standard anaesthetic procedure was followed for all cases, using thiopental sodium, succinylcholine, fentanyl, atracurium and 60% N2O in 40% oxygen supplemented with isoflurane. Mean arterial blood pressure (MAP) was reduced to 60 +/- 5 mm Hg in all patients using a sodium nitroprusside infusion. The end tidal (ET) CO2, PaCO2, MAP, peak airway pressure, plateau pressure and expiratory minute volume were recorded during a period of normal arterial blood pressure (time 1) and during hypotension (time 2). RESULTS: A significant decrease in PaCO2 (7%) and ETCO2 (17%) from time 1 to time 2 (p < 0.01) was noted, as was a significant increase in P(a-ET)CO2 (48%) and in the Vdphys/Vt ratio (41.17%) (p < 0.01) during the same period. CONCLUSION: The decrease in ETCO2 does not reflect the changes in PaCO2. The larger decrease in ETCO2 is mainly due to the increase in the Vdphys/Vt ratio. During anaesthesia, once normocapnia is achieved with normal arterial blood pressure, there is hardly any need to change the ventilation after initiation of controlled hypotension. PMID- 12116692 TI - Inferior long-term outcome of renal transplantation in patients with diabetes mellitus. AB - OBJECTIVE: To retrospectively review the long-term outcome of renal transplant in diabetics at Mubarak Al-Kabeer Hospital and Hamad Al-Essa Organ Transplant Center, Kuwait from 1983 to 1998. METHODS: There were 631 renal transplant patients, comprising 79 (12.5%) patients with pretransplant diabetes mellitus (pre-TDM), 117 (18.5%) patients with post-transplant diabetes mellitus and 435 (69%) nondiabetics (ND). Subjects with post-transplant diabetes mellitus were excluded from the comparative analysis. Distribution of sex, source of donors and mode of immunosuppression were similar in pre-TDM and ND groups. RESULTS: Fifty three (67%) recipients in pre-TDM and 90 (20.5%) in the ND group (p < 0.01) were above 45 years of age. However, 26 (33.3%) pre-TDM and 345 (79.5%) ND were below age 45. Among those who died, coronary artery disease led to death in 36% of pre TDM and 27% in ND. Hyperlipidemia requiring drug therapy was observed in 37% pre TDM and 6% ND. The incidence of severe infections was nearly twice in pre-TDM over ND recipients (1.9 vs. 1.0 per patient, p < 0.001). Acute rejection episodes were more frequently seen in pre-TDM (43%) than ND (33%), however the difference was not statistically significant. The patient survivals at 1, 5, 10, 14 years were significantly lower in pre-TDM (84, 65, 58 and 58%, respectively) than in ND (97, 93, 86 and 82%, respectively). The major contributory factors were severe infections and coronary artery disease. The cumulative graft survival showed a similar pattern (52% in pre-TDM, 73% in ND at 10 years). However, when death is excluded, the 10-year pure graft survival probability was similar for the pre-TDM and ND groups (76% vs. 80%). CONCLUSION: Our study indicates poor patient survival in pre-TDM due to coronary artery disease and infections, whereas the pure long-term graft survival was equally good in pre-TDM and ND transplant recipients. PMID- 12116694 TI - Hemoglobin electrophoresis and hemoglobinopathies in Kuwait. AB - OBJECTIVES: To analyze the results of hemoglobin electrophoresis (HE) in the routine laboratory of a tertiary hospital in Kuwait and to review the common types of hemoglobinopathies prevalent in the country. METHODS: This was a prospective study of HE performed on 2,386 samples in Mubarak Al-Kabeer Hospital, which serves more than 30% of the population of Kuwait, from June 1997 to May 1998. RESULTS: Of the 2,386 HE tests, only 561 (23.5%) had abnormal hemoglobin genotypes. The most commonly identified hemoglobinopathies were beta-thalassemia minor (14%), sickle cell trait (6%), sickle cell anemia (0.9%), S beta zero thal (0.8%) and S beta + thal (0.8%). Two rare hemoglobin variants, Hb DPunjab and Hb E, were encountered. CONCLUSION: HE yielded only 23.5% abnormal results, thus indicating the need to streamline requests for the test. The test should be limited to patients with hematological and clinical features suggestive of hemoglobinopathies or to individuals with a positive family history. PMID- 12116695 TI - A prospective evaluation of preoperative screening laboratory tests in general surgery patients. AB - OBJECTIVE: To assess the value of routine biochemical and hematological screening of otherwise healthy patients prior to elective general surgery. MATERIALS AND METHODS: Prospective laboratory screening tests were done for 1,000 consecutive patients undergoing elective general surgery at Mubarak Al-Kabeer Hospital, Kuwait, from January to August 1999. Patients with abnormal laboratory results were interviewed and examined preoperatively as part of the present study protocol, to identify a possible cause for the abnormal laboratory result. The perioperative course of these patients was also monitored. RESULTS: Approximately 14% of the preoperative tests were abnormal, 9.2% of which was expected while 4.9% was unexpected; there was no change in the preoperative care of patients with unexpected abnormalities, nor was there surgical delay or related postoperative complication. CONCLUSION: The results indicate that laboratory tests should be selectively used when the patient has appropriate risk factors. Instead, greater emphasis should be placed on history and physical examination. Application of such policy will result in substantial financial savings that could be utilized to improve the health care system. PMID- 12116696 TI - Leigh's disease in 3 sibs of a Kuwaiti family. AB - OBJECTIVE AND IMPORTANCE: To describe Leigh's disease in 3 sibs of a Kuwaiti family. CASE PRESENTATION: Two brothers presented in early infancy with progressive neurological symptoms of hypotonia, delayed milestones and brisk reflexes. Investigations revealed metabolic acidosis, high serum and cerebrospinal fluid lactate. Magnetic resonance imaging (MRI) showed characteristic changes of Leigh's disease. The 3rd brother, who was asymptomatic initially, was investigated because of his family history and was found to have similar changes. INTERVENTION: All children developed progressive neurological deterioration and persistent metabolic lactic acidosis, which was treated with sodium bicarbonate, and the 1st patient needed renal dialysis to control the acidosis. The 2nd child was placed on vitamins and carnitine. CONCLUSION: The neurological deterioration was progressive in all 3 sibs, and they eventually died of respiratory failure despite ventilatory support. Since MRI changes are characteristic, MRI should be done to confirm the diagnosis. PMID- 12116698 TI - Successful implantation of a permanent pacemaker through a persistent left superior vena cava. AB - We present a case of successful implantation of a permanent pacemaker through an unusual course of a persistent left superior vena cava (PLSVC). A young male patient presented with symptomatic bradycardia and a heart rate of 35 beats per minute. The pacing lead was introduced through the standard left subclavian vein approach and was found to pass through an unusual course that was suspected to be a PLSVC. The lead was further advanced to the right atrium and positioned successfully in the right atrial appendage with excellent short-term and long term pacing parameters. PLSVC should be suspected whenever the catheter takes an unusual position during central venous catheterization. Most right heart catheterization procedures, including permanent pacemaker implantation, can be safely completed in spite of this anomaly. PMID- 12116697 TI - Unexpected prolonged neuromuscular block after mivacurium: a case report. AB - OBJECTIVE: To present a case of unexpected prolonged apnoea following the administration of mivacurium, a short-acting muscle relaxant and to identify the factors for early diagnosis and management. CLINICAL PRESENTATION AND INTERVENTION: A 19-year-old physically fit lady without personal or family history suggestive of anaesthetic problems had an excision of fibro-adenoma from the breast. She did not recover as quickly as was expected from the anaesthetic, which included the administration of mivacurium. She had prolonged neuromuscular blockade. She was kept ventilated and sedated. Five hours after the last dose of mivacurium, she showed signs of spontaneous respiration and at 6 h she was extubated and fully recovered. It was shown later that the patient had a pseudocholinesterase deficiency. CONCLUSION: Pseudocholinesterase deficiency is an uncommon occurrence but should be highly suspected in cases of prolonged paralysis following the administration of a short-acting muscle relaxant. The use of a nerve stimulator is recommended whenever muscle relaxants are used. Muscle relaxants should be used only when facilities for prolonged ventilation are available. PMID- 12116700 TI - High and low luminance letters, acuity reserve, and font effects on reading speed. AB - We compared reading speed with two fonts, Dutch (serif) and Swiss (sans serif). Text was displayed on a computer monitor, white letters on black, with the RSVP method. Luminance of the letters was either 146.0 or 0.146 cd m-2. Lower-case x height of the fonts was approximately 5.5 times as large as letter acuity. At the high luminance, there was no difference between reading rates. There was a significant advantage for the Swiss font at the low luminance. The acuity reserve for Swiss was higher than for Dutch at the low luminance, which may account for the difference in reading speeds. PMID- 12116699 TI - gamma-aminobutyric acid transporter-mediated current from bipolar cells in tiger salamander retinal slices. AB - About 10% of bipolar cells in salamander retina synthesize and take up gamma aminobutyric acid (GABA), and may use GABA as a neurotransmitter. As GABA uptake is electrogenic, bipolar cells expressing GABA transporters (GATs) should give transport current (IGAT) to extracellular GABA. Using whole-cell patch recording, 28 bipolar cells responded to 30-200 microM GABA puffed to the axon terminals with a picrotoxin (PTX)-sensitive chloride current (ICI) only. Another three bipolar cells had, in addition to ICI, a PTX-resistant, sodium-dependent current that was completely and reversibly blocked by NO-711, an IGAT inhibitor, indicating that this component was an IGAT. This finding provides further support for a subset of GABAergic bipolar cells in the salamander retina. PMID- 12116701 TI - Sensitivity to disparity corrugations in peripheral vision. AB - Disparity discrimination thresholds are known to increase with both retinal eccentricity and distance from the horopter. However, little is known about how the detectability of cyclopean gratings varies with retinal position. Thresholds for disparity corrugations were measured as a function of corrugation frequency for different visual eccentricities. Subjects viewed annular displays of random dot stereograms, and judged in which of two intervals a circumferential disparity modulation was present. For any given eccentricity, visual sensitivity to disparity corrugations was bandpass. As eccentricity increased from 3.5 to 21.0 degrees, peak-to-trough thresholds were found to increase, the optimal corrugation frequency for detection decreased, and the upper cutoff corrugation frequency also decreased. The M-Scaling functions of Rovamo and Virsu were used to replot the data in terms of cycles per unit cortical distance. Peak detection frequency was constant at 0.8 cycles per mm of cortex after this rescaling, demonstrating that acuity for disparity modulations is approximately M-scaled beyond the fovea. PMID- 12116702 TI - Functional architecture of primate cone and rod axons. AB - The cone axon is nearly four times thicker than the rod axon (1.6 vs 0.45 microns diameter). To assess how signal transfer and integration at the terminal depend on cable dimensions, a transducer (cone = ohmic conductance, rod = current source) coupled via passive cable to a sphere with a chloride conductance (representing GABAA receptor) was modelled. For a small signal in peripheral cone with a short axon, steady photosignal transfers independently of axon diameter despite a significant chloride conductance at the cone terminal. Temporally varying photosignal also transfers independently of axon diameter up to 20 Hz and is attenuated only 20% at 50 Hz. Thus, to accomplish the basic electrical functions of a peripheral cone, a thin axon would suffice. For a foveal cone with a long axon steady photosignal transfers independently of axon diameter, but temporally varying photosignal is attenuated 5-fold at 50 Hz for a thick axon and 10-fold for a thin axon. This might contribute to the lower sensitivity of central retina to high temporal frequencies. The cone axon contains 14-fold more microtubules than the rod axon, and its terminal contains at least 20-fold more ribbon synapses than the rod's. Since ribbon synapses sustain high rates of exocytosis, the additional microtubules (which require a thicker axon) may be needed to support a greater flux of synaptic vesicle components. PMID- 12116703 TI - Modeling the physiological responses of anuran R3 ganglion cells. AB - Teeters and Arbib (Bio Cybernet 1991;64:197-207) presented a model of the anuran retina which qualitatively accounts for some of the characteristic response properties used to distinguish ganglion cell type in anurans. Teeters et al. (Vis Res 1993;33:2361-2379) tested the model's ability to reproduce data of Ewert and Hock (Exp Brain Res 1972;16:41-59) relating toad R2, R3 and R4 ganglion cell responses to moving worm, antiworm and square-shaped stimuli of various edge lengths for stimulus shape and size dependency. In this paper we provide an exhaustive analysis of the performance of the modeled R3 cells with respect to most of the known qualitative and quantitative physiological properties of natural R3 ganglion cells. We also introduce several relevant predictions of the model relating different responses of R3 cells under the effect of changes in different model components. In some cases the predictions have been tested in neurophysiological experiments. PMID- 12116704 TI - The spatial arrangement of the L and M cones in the central fovea of the living human eye. AB - Experiments designed to estimate the placement of L and M cones in fovea centralis of the living human eye are presented. Hyperacuity performances for two observers were measured for the full and the separate L and M cone submosaics using 2-dot chromatic stimuli on cone-selective adapting backgrounds. Simulated performances, based on an ideal observer model, were generated for all possible mosaics by varying L and M cone relative numerosity and spatial configuration. The best match between the simulated and measured performances determined the solution mosaic. Each observer's solution mosaic contained more L than M cones, randomly arrayed as assessed by statistical tests. PMID- 12116705 TI - Long-lasting, long-range detection facilitation. AB - We examined the time course of threshold reduction in the Gabor lateral masking paradigm. Contrast detection thresholds were measured (2AFC) for a briefly presented (36 ms) foveal Gabor signal (GS), preceded by a presentation (90 ms) of two high-contrast GS flanked masks, with stimulus onset asynchrony (SOA) varying from 0 to 16,290 ms. Using target-to-mask separations of 3 lambda and 12 lambda (lambda = 0.15 degree, GS wavelength), the 3 lambda separated GS masks enhanced target threshold by 0.25 log units at SOA = 0 and by 0.17 log units at 2700 ms. At 12 lambda separation, threshold was enhanced by 0.11 log units at SOA = 0 and by 0.14 log units at 2700 ms. Long-range (12 lambda) and short-range (3 lambda) enhancements persisted for over 16 s. Delayed and simultaneous enhancement depended on the stimulus configuration (maximal for collinear target and masks), local parameters (orientation, spatial frequency and phase), and the presented eye (dichoptic versus monoptic). The results suggest that spatial filters in early vision retain an input trace far beyond the perceptual integration range. This trace may subserve the consolidation of filter activity into long-term memory. PMID- 12116706 TI - Spatial frequency discrimination and detection characteristics for gratings defined by orientation texture. AB - We describe evidence consistent with the proposal that the visual system contains a parallel array of size-tuned mechanisms sensitive to orientation texture defined (OTD) form, and propose that the relative activity of these mechanisms determines spatial frequency discrimination threshold for OTD gratings. Using a pattern of short lines we measured spatial frequency discrimination thresholds for OTD gratings and luminance-defined (LD) gratings. For OTD gratings, the orientation of texture lines varied sinusoidally across the bars of the gratings, but line luminance was constant. For LD gratings, line orientation was constant, but line luminance varied sinusoidally across the bars of the grating. When the number of texture lines (i.e. spatial samples) per grating cycle was below about six, spatial sampling strongly affected both the spatial frequency discrimination and grating detection thresholds for OTD and LD gratings. However, when the number of spatial samples per grating cycle exceeded about six, plots of both discrimination threshold and detection threshold were different for OTD and LD gratings. For an OTD grating of any given spatial frequency, spatial frequency discrimination threshold fell as the number of samples per grating cycle was increased while holding texture line length constant: the lower limit was reached at six to ten samples per cycle. When we progressively increased the viewing distance (keeping the cycles per degree (cpd) constant), spatial frequency discrimination threshold reached a lower limit and increased thereafter. We propose that this minimum threshold represents a balance between opposing effects of the number of samples per grating cycle and the length of texture lines, and approaches the absolute physiological lower limit for OTD gratings. Spatial frequency discrimination was possible up to at least 7 cpd. Grating acuity for an OTD grating was considerably lower than the physiological limit for LD gratings, presumably because detectors of OTD form include a spatial integration stage following the processing of individual lines. For an LD grating, discrimination threshold fell as the number of samples per grating cycle was increased and asymptoted at six to ten samples per cycle. Spatial frequency discrimination thresholds for OTD and LD gratings were similar at low spatial frequencies (up to 3-4 cpd), but increased more steeply for OTD gratings at high spatial frequencies. For both OTD and LD gratings, discrimination threshold fell steeply as the number of grating cycles was increased from 0.5 to ca. 2.5 cycles, and thereafter decreased more slowly or not at all suggesting that, for both OTD and LD gratings, spatial frequency discrimination can be regarded as a special case of line interval or bar width discrimination. As orientation contrast was progressively increased, discrimination threshold for an OTD grating fell steeply up to about four to five times grating detection threshold, then saturated. This parallels the effect of luminance contrast on discrimination threshold for an LD grating. PMID- 12116707 TI - Tracing the timing of human analysis of motion and chromatic signals from occipital to temporo-parieto-occipital cortex: a transcranial magnetic stimulation study. AB - In human visual analysis, the initial processing of motion and chromatic signals may be mediated by feed-forward pathways from striate cortex to segregated areas of extrastriate cortex. The time-course of occipital to temporo-parieto-occipital motion processing was unknown, as was the selectivity of the effect of transcranial magnetic stimulation (TMS) on motion processing. TMS delivered over occipital cortex degraded the discrimination of motion-defined form (MDF) in a discrete time window beginning 100-120 ms from the onset of the visual stimulus. Bilateral focal TMS delivered over the temporo-parieto-occipital junction (TPO) disrupted the discrimination of MDF in a time window beginning 20-40 ms later than the effect of TMS delivered over occipital cortex. Bilateral focal TMS delivered over TPO also degraded the discrimination of CDF, motion direction, and color. PMID- 12116708 TI - Assessment of visual acuity and contrast sensitivity in the chick using an optokinetic nystagmus paradigm. AB - While the chick is one of the widely used animal models for eye growth studies very little is known about its visual spatial resolution performance. Using optokinetic nystagmus responses as an indicator of stimulus visibility, we estimated the visual acuity of young chicks to be between 6.0 and 7.7 cycles deg 1 at 2 and 4 days of age and slightly higher, between 7.7 and 8.6 cycle deg-1, at 8 days. Contrast sensitivity measured using the same experimental paradigm was greatest at around 1.2 cycle deg-1, for which the contrast threshold lay between 4% and 11%. Sensitivity became progressively poorer for frequencies both higher and lower than this. These data suggest that the visual performance of the chick is slightly poorer than that of the pigeon which has a similar eye size and exhibits similar foraging behaviour. PMID- 12116709 TI - Ideal observer perturbation analysis reveals human strategies for inferring surface orientation from texture. AB - Optical texture patterns contain three quasi-independent cues to planar surface orientation: perspective scaling, projective foreshortening and density. The purpose of this work was to estimate the perceptual weights assigned to these texture cues for discriminating surface orientation and to measure the visual system's reliance on an isotropy assumption in interpreting foreshortening information. A novel analytical technique is introduced which takes advantage of the natural cue perturbations inherent in stochastic texture stimuli to estimate cue weights and measure the influence of an isotropy assumption. Ideal observers were derived which compute the exact information content of the different texture cues in the stimuli used in the experiments and which either did or did not rely on an assumption of surface texture isotropy. Simulations of the ideal observers using the same stimuli shown to subjects in a slant discrimination task provided trial-by-trial estimates of the natural cue perturbations which were inherent in the stimuli. By back-correlating subjects' judgments with the different ideal observer estimates, we were able to estimate both the weights given to each cue by subjects and the strength of subjects' prior assumptions of isotropy. In all of the conditions tested, we found that subjects relied primarily on the foreshortening cue. A small, but significant weight was given to scaling information and no significant weight was given to density information. In conditions in which the surface textures deviated from isotropy by random amounts from stimulus to stimulus, subject judgements correlated well with the estimates of an ideal observer which incorrectly assumed surface texture isotropy. This correlation was not complete, however, suggesting that a soft form of the isotropy constraint was used. Moreover, the correlation was significantly lower for textures containing higher-order information about surface orientation (skew of rectangular texture elements). The results of the analysis clearly implicate texture foreshortening as a primary cue for perceiving surface slant from texture and suggest that the visual system incorporates a strong, though not complete, bias to interpret surface textures as isotropic in its inference of surface slant from texture. They further suggest that local texture skew, when available in an image, contributes significantly to perceptual estimates of surface orientation. PMID- 12116710 TI - Texture segregation, surface representation and figure-ground separation. AB - A widespread view is that most texture segregation can be accounted for by differences in the spatial frequency content of texture regions. Evidence from both psychophysical and physiological studies indicate, however, that beyond these early filtering stages, there are stages of 3-D boundary segmentation and surface representation that are used to segregate textures. Chromatic segregation of element-arrangement patterns--as studied by Beck and colleagues--cannot be completely explained by the filtering mechanisms previously employed to account for achromatic segregation. An element arrangement pattern is composed of two types of elements that are arranged differently in different image regions (e.g. vertically on top and diagonally on the bottom). FACADE theory mechanisms that have previously been used to explain data about 3-D vision and figure-ground separation are here used to simulate chromatic texture segregation data, including data with equiluminant elements on dark or light homogeneous backgrounds, or backgrounds composed of vertical and horizontal dark or light stripes, or horizontal notched stripes. These data include the fact that segregation of patterns composed of red and blue squares decreases with increasing luminance of the interspaces. Asymmetric segregation properties under 3-D viewing conditions with the equiluminant elements close or far are also simulated. Two key model properties are a spatial impenetrability property that inhibits boundary grouping across regions with non-collinear texture elements and a boundary-surface consistency property that uses feedback between boundary and surface representations to eliminate spurious boundary groupings and separate figures from their backgrounds. PMID- 12116711 TI - Entoptic image quality of the retinal vasculature. AB - Spatial details of entoptically visible retinal vessels were investigated using transcleral and Maxwellian-view stimulators. Nine normal subjects provided detailed drawings of the entoptic images which were digitized and superimposed onto digitized fundus photographs and fluorescein angiograms from the same eyes. Subjects also used a tracing method to locate visible entoptic features. The trans-scleral method provided images similar in detail to standard fundus photography (lacking capillary detail, but capturing larger arteries, veins, arterioles and venules) in the macula and around the disk. The Maxwellian-view method illuminated the fovea (7.7 degree field) and provided foveola capillary detail (capillaries traversing the foveola, the capillary arcade forming the FAZ) as well as the larger foveal vessels supplying the foveola, and often contained more foveal detail that available with fluorescein angiography. PMID- 12116712 TI - [Diabetes of adult type is a polymorphic disease]. PMID- 12116713 TI - [Nummular eczema]. PMID- 12116714 TI - [Autologous stem cell transplantations in non-Hodgkin lymphomas]. PMID- 12116715 TI - [Two faces of gonadotroph adenoma]. PMID- 12116716 TI - [Treatment of intestinal ischemia with balloon dilatation and stent]. PMID- 12116717 TI - [Imaging diagnostics of appendiceal mucocele]. PMID- 12116718 TI - [Treatment of genital herpes]. PMID- 12116719 TI - [Pediatric Research Foundation's research grants and efficiency of their use]. PMID- 12116720 TI - [A physician in his place?]. PMID- 12116721 TI - [The physician's work--skills or art?]. PMID- 12116722 TI - [Psychomotor slowness of a mentally retarded woman]. PMID- 12116723 TI - Effectiveness of a powered toothbrush compared with a manual toothbrush for orthodontic patients with fixed appliances. AB - Orthodontic patients with fixed appliances have an increased risk for caries and gingivitis. Therefore, the use of special toothbrushes and additional cleaning tools is recommended. The aim of this longitudinal study was to compare the plaque removal efficacy and reduction of gingivitis from using a powered toothbrush compared to a manual toothbrush in patients with fixed orthodontic appliances. Eighty subjects were included in the study with a mean age of 13.53 years. After a baseline examination, patients were randomly assigned to two groups and their teeth were professionally cleaned. The patients were assessed at baseline, two and four weeks using the QHI and SBI. The results showed a significantly superior plaque removal effect (p = 0.0001) and reduction of gingival inflammation (p < 0.05) in those patients using the powered versus the manual toothbrush. The findings were for both whole mouth and partial mouth assessments. In accordance with previous studies, it was demonstrated that the use of a powered toothbrush can be recommended for orthodontic patients with fixed appliances. PMID- 12116724 TI - Applications of polymers in dentifrices and mouthrinses. AB - This review highlights the specific applications of polymers in oral hygiene products. Specific examples are provided to show how polymers can be used to improve retention and release of active agents for therapeutic effects in an oral environment where the residence time of the actives is low, owing to the continuous flow of saliva which washes them away. Polymers are uniquely suited as active therapy for intraoral applications and as replacement agents for augmenting the macromolecule's deficiency (saliva) caused in the aging mouth. It is expected that the next generation of products and delivery systems will be based on polymers derived from natural sources and will have functions such as enamel replacement agents for caries control, adhesion macromolecules for gingival tissue attachment to teeth, saliva mucin replacement, and the delivery of new active agents using biopolymers. PMID- 12116725 TI - Triclosan/pyrophosphate dentifrice: dental plaque and gingivitis effects in a 6 month randomized controlled clinical study. AB - A double-blind, parallel, randomized and controlled clinical trial was conducted on 186 subjects over six months to assess the effects of a 0.28% triclosan/5% pyrophosphate (with NaF/silica) dentifrice on dental plaque and gingivitis as compared to a NaF/silica negative control dentifrice. An initial examination was performed to assess the health of the oral soft and hard tissues and to measure plaque (by Turesky modified Quigley-Hein Plaque Index), gingivitis (by Loe Silness Gingival and Ainamo and Bay Gingival Bleeding [GBI] indices). Only those subjects with a GBI score > or = 5 were accepted into the study. Each enrolled subject received an oral prophylaxis and was requested to brush and floss twice per day with the negative control NaF/silica dentifrice. After one month, the subjects were recalled and a baseline examination was performed for each of the previously described parameters. Following the baseline examination, the subjects received another oral prophylaxis. The subjects were then separated by gender and by baseline GBI scores of < or = 7 or > 7 and arrayed by the changes in GBI bleeding sites from initial to baseline. Within strata, subjects were randomly assigned to brush twice per day with either the triclosan/pyrophosphate dentifrice or the negative control dentifrice. The subjects were subsequently examined for all of the above-described parameters following use of the test dentifrices for five weeks, three and six months. The data generated in this trial were analyzed using an analysis of covariance on all indices for all subjects completing the examinations. The results from this study demonstrated that the use of the triclosan/pyrophosphate dentifrice resulted in statistically significant reductions of dental plaque compared to the control by 10% (p < 0.05), 15.4% (p < 0.01) and 13.9% (p < 0.01) at five weeks, three and six months, respectively. However, there were no statistically significant differences between the test dentifrices for any of the gingivitis or gingival bleeding evaluations throughout the study. Based on 1) the fact that subjects possessed plaque-induced gingivitis in this clinical study, 2) the similarity in the magnitude of the plaque reductions observed from the triclosan/pyrophosphate dentifrice relative to those reported for other triclosan-containing dentifrices, 3) the similarity in the dose of triclosan relative to other triclosan dentifrices, and 4) the reported magnitude of gingivitis reductions from other triclosan-containing dentifrices, these findings were unexpected. Possible explanations of these results are that the triclosan/pyrophosphate dentifrice may be uniquely different from other triclosan dentifrices relative to its effects on gingivitis, or alternatively, the clinical design utilized here may not be optimized for triclosan/pyrophosphate dentifrice. PMID- 12116726 TI - Experimental gingivitis studies: effects of triclosan and triclosan-containing dentifrices on dental plaque and gingivitis in three-week randomized controlled clinical trials. AB - A recently reported six-month gingivitis study demonstrated that in subjects with gingivitis, a triclosan/pyrophosphate dentifrice provided supragingival plaque control. The level of plaque reduction was comparable with that reported for other triclosan-containing dentifrices; however, no reductions in gingivitis were observed for triclosan/pyrophosphate relative to the negative control. One possible explanation of this result is that the Hawthorne effect in the study was too great to allow the detection of a treatment benefit for the triclosan product. In order to further explore the relevance of these results, three independent clinical studies were undertaken utilizing designs based on a 21-day experimental gingivitis model in which Hawthorne effects are minimized, in part due to the absence of toothbrushing. In each model, a pre-study prophylaxis was followed by a three-week period of oral hygiene instruction to establish optimum baseline gingival health in study participants. The studies varied in enrollment; 120, 33 and 32 subjects completed treatment on studies 1, 2, and 3, respectively. In study 1, test articles were dentifrice products (0.28% triclosan/5% pyrophosphate/0.145% sodium fluoride, 0.2% triclosan/0.5% zinc citrate/0.112% sodium fluoride, 0.145% sodium fluoride and 0.15% sodium monofluorophosphate) applied neat and undiluted via a performed tooth shield (that prevents mechanical tooth-brushing at the test sites in the oral cavity) in a partial mouth design. In study 2, test articles were also dentifrice products (0.28% triclosan/5% pyrophosphate/0.243% sodium fluoride, 0.3% triclosan/2% Gantrez copolymer/0.24% sodium fluoride and 0.243% sodium fluoride) but administered to subjects in the form of 1:3 aqueous slurry rinses. Lastly, in study 3, test articles were all mouthrinses (0.12% chlorhexidine, 0.045% triclosan in ethanol plus respective vehicle placebos). Clinical assessments to quantify the test articles' effects on the development of plaque and gingivitis were conducted at baseline (studies 1, 2 and 3), day 7 (studies 2 and 3), day 14 (studies 2 and 3) and day 21 (studies 1, 2 and 3). In study 1, no statistically significant treatment effects were observed between the test articles and controls for plaque or gingivitis development. In study 2, no statistically significant treatment effects were observed at any time point between test products for the development of gingivitis. At days 7 and 14, there were no significant differences between test products and control for plaque development as well. At day 21, the group rinsing with the triclosan/pyrophosphate/sodium fluoride slurry had significantly less plaque accumulation than the group rinsing with the triclosan/copolymer/sodium fluoride slurry (p < 0.05); however, neither of the groups using test products containing triclosan was significantly different for plaque development from the group using the sodium fluoride control test article. In addition, aspartate aminotransferase activity in gingival crevicular fluid was assayed at days 0 and 21; no between-group differences were found at either of these time points, though day 21 AST activities were higher than those at baseline. In study 3, statistically significant treatment differences in plaque regrowth and gingivitis were observed at day 21 for the chlorhexidine rinse versus all other rinses (p < 0.05). No other statistically significant treatment effects were observed between test compounds at any other time points. The results benchmark the anti-plaque and anti-gingivitis benefit for a range of triclosan-based product forms against positive and negative controls in a three different experimental gingivitis models, a design considered predictive of clinical efficacy in longer-term investigations. It is concluded that dentifrice products containing triclosan do not possess sufficient antimicrobial activity to suppress plaque and gingivitis development in the absence of normal oral hygiene, and that relative to chlorhexidine, triclosan itself offers only modest efficacy for the prevention of plaque accumulation and therefore the delayed onset of gingivitis. PMID- 12116727 TI - Effects of triclosan/copolymer dentifrice on dental plaque and gingivitis in a 3 month randomized controlled clinical trial: influence of baseline gingivitis on observed efficacy. AB - Triclosan is a broad-spectrum antimicrobial agent widely used in oral care products. In recent studies, a triclosan/pyrophosphate dentifrice has shown efficacy against dental plaque but not gingivitis. Further, experimental gingivitis studies on triclosan itself and in combination with other dentifrice ingredients demonstrate only moderate antimicrobial activity. In contrast, there are a number of other studies in the literature reporting the antigingivitis efficacy of triclosan/copolymer and triclosan/zinc citrate dentifrices. These dentifrices possess similar effects on dental plaque to triclosan/pyrophosphate, thus the lack of effect on gingivitis from triclosan/pyrophosphate was unexpected since comparable effects on dental plaque from similar formulations may infer similar antimicrobial activity and duration of action within the oral cavity. Therefore, the objective of this research was to understand the clinical variables important to observe the reported effects for triclosan dentifrice so this clinical model could in turn be used to test the effects of the triclosan/pyrophosphate dentifrice on gingivitis. To achieve this objective, it was determined that a suitable approach was to duplicate the methodology of previous successful clinical trials on triclosan/copolymer dentifrice in order to better understand the study design used to demonstrate the antigingivitis efficacy of triclosan-containing dentifrices. To this end, a prospective trial was conducted employing the same active product, clinical site, investigator and gingivitis/plaque examiner previously used. The study was a randomized, blinded, placebo-controlled, parallel group 3-month trial, which recruited subjects with Loe-Silness Gingival Index (GI) scores > or = 1.0 and Turesky Plaque Index (PI) scores of > or = 1.5. The study population consisted of 160 adults who brushed twice daily with either triclosan/copolymer or placebo control (containing copolymer) following a prophylaxis. Gingivitis and plaque were measured using the GI and PI, respectively, and scores were analyzed using a one-way analysis of covariance. There was no evidence that 3-month gingivitis or plaque scores (whole mouth or severity index) for the triclosan/copolymer group were different from the placebo group. Additional analyses were conducted on study population subgroups with increasing intervals of baseline GI bleeding sites to gauge the effect of baseline bleeding on the treatment effect. For subjects in the > or = 40 bleeding sites subset, triclosan/copolymer demonstrated a 4.2% GI and 15% gingivitis severity index reduction versus placebo at 3 months (0.10 < p < 0.20). These findings suggest that a study design which includes subjects with greater numbers of gingival bleeding sites at baseline may have the required sensitivity to demonstrate treatment benefits for triclosan/copolymer. However, additional experimental parameters remain to be fully articulated in order to replicate previously successful trials. The overall results of this trial are consistent with the experimental gingivitis results, and indicate that, even when formulated with the copolymer, triclosan as an oral antimicrobial agent possesses limited activity as an antigingivitis ingredient. PMID- 12116729 TI - Biological evaluation as part of risk assessment. AB - This overview of how to conduct a risk assessment highlights the use of one of the newest standards in this area ISO 14971, which covers the application of risk management, as well as ISO 10993, which covers biological evaluation of medical devices. A hypothetical risk evaluation of a device is also described. PMID- 12116728 TI - Caries prevention in Chinese children with sodium fluoride dentifrice delivered through a kindergarten-based oral health program in China. AB - Caries in China appears to be a significant problem, especially among preschool children. The effect of caries prevention in the primary teeth of preschool children through the use of fluoridated dentifrices and prevention programs has not been widely addressed. The purpose of this study was to examine the caries preventive effects of an 1100 ppm sodium fluoride dentifrice used in the context of a kindergarten-based oral health program compared to a matched placebo dentifrice in the absence of a kindergarten-based oral health program. This was a randomized, placebo-controlled, examiner-blind, two-year caries study. A population of 1,334 preschool school children, three years of age, was recruited from 24 school kindergartens in Huairoi and Miyun counties, located approximately 60 kilometers northeast of Beijing, China. Classrooms were stratified based on mean baseline dmfs scores derived from the visual-tactile baseline examination, and randomly assigned to one of the two dentifrice treatment groups: 0.243% sodium fluoride (1100 ppm fluoride ion) or placebo (0 ppm fluoride ion). Children attending the schools participating in the program brushed twice a day (morning and afternoon) at school under the supervision of classroom teachers during the school week. The children randomized to schools receiving the placebo dentifrice were supplied with toothbrushes and dentifrice for ad libitum use at home and did not participate in the school program or supervised classroom brushing. For the primary examiner, the two-year caries increment data demonstrated evidence of anticaries efficacy for the sodium fluoride dentifrice/school program group as compared to the placebo/no school program group. In the evaluable subset, the 1100 ppm fluoride treatment group had a 20.7% reduction in dmfs compared to the placebo treatment group, that was statistically significant (p = 0.004). The secondary examiner observed a similar overall treatment effect, as the 1100 ppm fluoride treatment group had an 22.1% reduction in dmfs compared to the placebo treatment group that was statistically significant (p = 0.014). In contrast to the primary examiner, there was a county-by-treatment interaction for the secondary examiner's results necessitating that the counties be examined independently. In Miyun county, the sodium fluoride/school program group had a 39.9% reduction in caries compared to the placebo/no program group that was statistically significant (p = 0.001). In Huairou county, the sodium fluoride/school program group had a 6.8% reduction in caries compared to the placebo/no program group that was not statistically significant (p > 0.05). These results demonstrate that fluoride in conjunction with increased dental awareness can deliver important reductions in caries in preschool children. PMID- 12116730 TI - The mechanics of removing organisms by filtration: the importance of theory. AB - Very often, the concept of filtration is only considered in terms of sieve retention, which can sometimes be misleading. In this review of the adsorptive particle-capture mechanism, the authors aim to familiarize the filtration practitioner with the possible optimisation of particle retention by the manipulation of factors such as ionic strength, osmolarity, temperature and the use of polymeric filters. PMID- 12116731 TI - Trends in medical filtration. AB - Advances in materials, mould tooling and control systems are offering the industry greater design choices in filtration as well as the potential to reduce manufacturing costs. This article describes what is possible. PMID- 12116732 TI - The technological edge. AB - The development of a real-time 2D-3D duplex ultrasonic scanner offers faster diagnostic imaging. PMID- 12116733 TI - Implementation of the medical device directives in Italy. AB - Medical device manufacturers marketing their products in Italy, or planning to do so, need to be aware of any national requirements such as those concerning the registration of manufacturers and devices and language requirements that may apply to their products. This article provides an update on the implementation of the European Directives for medical devices in Italy. PMID- 12116734 TI - Mirandola: Italy's biomedical valley. AB - Revenues of the vibrant medical device sector in Mirandola grew by 28% between 1997 and 2000 and figures are expected to show an increase of 10.5% for 2001. This article explores the dynamics of the region. PMID- 12116735 TI - Filters and open-heart surgery. PMID- 12116736 TI - Laser marking: flexible and fast. PMID- 12116737 TI - Reassessing bioactive surfaces. PMID- 12116738 TI - Optimal values of flow velocity on transcranial Doppler in grading middle cerebral artery stenosis in comparison with magnetic resonance angiography. AB - BACKGROUND AND PURPOSE: To investigate the optimal values of flow velocity on transcranial Doppler (TCD) in grading the severity of middle cerebral artery (MCA) stenosis in comparison with magnetic resonance angiography (MRA). METHODS: Both TCD and MRA examinations were performed on 148 asymptomatic patients. The peak flow velocities of each MCA were recorded. Severity of MCA stenosis on MRA was classified as normal-mild (< 50% lumen diameter reduction), moderate (50% 75%), and severe-void (> 75% and void of flow signal). RESULTS: Among 296 MCAs evaluated, normal-mild stenosis was found in 75 (25%), moderate stenosis in 112 (38%), and severe stenosis in 109 (37%). The mean of systolic velocity (Vs) of MCA differed significantly among these three groups: mean Vs = 121.83 +/- 22.52 cm/s in the normal-mild group; 155.96 +/- 21.62 cm/s for the moderate group; and 199.39 +/- 43.86 cm/s for the severe group (P < .001). The optimal cutoff velocity for detection of MCA (> 50%) stenosis was found at Vs > 140 cm/s on TCD (area under the ROC curve is 0.87, P < 0.001). The best cutoff points for grading severity of on TCD were 140 cm/s and 180 cm/s. CONCLUSION: TCD enables grading of the severity of MCA stenosis according to the flow velocity. This method provides a noninvasive and reliable method for grading MCA stenosis and allows longitudinal monitoring of the relationship between clinical outcome and hemodynamic change. PMID- 12116739 TI - Echocardiographic findings of patients with retinal ischemia or embolism. AB - BACKGROUND AND PURPOSE: A potential source of emboli is not detected in more than 50% of patients with retinal arterial occlusive events. Echocardiographic studies are not always included in the diagnostic workup of these patients. The authors studied the diagnostic yield of transthoracic (TTE) and/or transesophageal (TEE) echocardiography in identifying potential sources of emboli in patients with retinal ischemia or embolism. METHODS: In a prospective study, 73 consecutive patients with clinically diagnosed retinal ischemia or embolism received a standardized diagnostic workup including retinal photography, echocardiography, and imaging studies of the internal carotid arteries. TTE was performed in 83.6% of patients, TEE was performed in 5.5% of patients, and both TTE and TEE were performed in 11.0% of patients. Ophthalmological diagnoses consisted of amaurosis fugax (n = 28), asymptomatic cholesterol embolism to the retina (n = 34), and branch or central retinal artery occlusion (n = 11). RESULTS: Echocardiography identified a potential cardiac or proximal aortic source for embolism in 16 of 73 (21.9%) patients, including 8 who also had either atrial fibrillation or internal carotid artery stenosis of more than 50% on the side of interest. Thus, 8 of 73 (11.0%) patients had lesions detected only by echocardiography. The most commonly identified lesions were proximal aortic plaque of more than 4 mm thickness (n = 7, 9.6%) and left ventricular ejection fraction of less than 30% (n = 6, 8.2%). TEE was particularly helpful in identifying prominent aortic plaques. CONCLUSION: Echocardiography frequently identifies lesions of the heart or aortic arch that can act as potential sources for retinal ischemia or embolism. Further studies are needed to evaluate the prognostic and therapeutic relevance of these findings. PMID- 12116740 TI - Focal cortical dysplasia: improving diagnosis and localization with magnetic resonance imaging multiplanar and curvilinear reconstruction. AB - OBJECTIVE: To establish the contribution of multiplanar reconstruction (MPR) and curvilinear reformatting (CR) to the MRI investigation of focal cortical dysplasia (FCD). METHODS: From a group of patients with intractable frontal lobe epilepsy, we selected patients with neuroimaging diagnosis of FCD. The diagnosis of FCD was based on the neuroimaging findings after a three step evaluation, always in the same order: (a) plain MRI films, (b) MPR, and (c) CR. After the selection of patients, the process of reviewing all the images in the three stages described above was performed by one of us, who did not take part on the selection of patients nor on the initial evaluation, and who was blind to the clinical and EEG findings of the patients. For data analysis, we first assessed the contribution of the additional findings of MPR analysis compared to the results of the evaluation using only plain MRI films, as is usually done in routine practice. Second, we assessed the contribution of CR to the findings of plain MRI films plus MPR. After completing the multistep evaluation, we all went back to review the plain MRI films with knowledge of lesion topography, in order to identify possible subtle features associated with FCD. RESULTS: Seventeen patients met the inclusion criteria. Twelve had imaging diagnosis of FCD and were included in the second step of this project. Plain films of high resolution MRI showed the lesion in 6 (50%) of the 12 patients. By adding MPR to the plain MRI films, we identified lesions in all 12 patients. Furthermore, we found that MPR provided a better lesion localization and ascertainment of its relationship to other cerebral structures in 5 of 6 (83%) patients who had a lesion identified on plain films. By adding CR to the plain MRI films plus MPR analysis, we observed that (a) CR also allowed the identification of the dysplastic lesion in all patients, (b) CR improved lesion localization in one patient, and (c) CR provided a better visualization of the lesion extent in 4 patients (33%), showed a larger lesion in 3, and demonstrated that part of the area suspected as abnormal was more likely volume averaging in 2. CONCLUSION: MPR and CR analysis add to the neuroimaging evaluation of FCD by improving the lesion diagnosis and localization. CR helps to establish the extent of the lesion more precisely, allowing the visualization of some areas not shown on high resolution MRI and MPR. These techniques are complementary and do not replace the conventional wisdom of MRI analysis. PMID- 12116741 TI - Criteria for normalcy of cavities observed within the adult hippocampus: high resolution magnetic resonance imaging study on a 3.0-T system. AB - BACKGROUND AND PURPOSE: Cavities occasionally found within the hippocampus during routine clinical magnetic resonance imaging (MRI) studies are believed to be a normal variant reflecting fluid collection within the vestigial hippocampal sulcus. However, the lack of systematic studies defining objective criteria for such cavities has hampered further assessment of potential abnormalities within the hippocampus. This study assessed the detailed characteristics of hippocampal cavities in normal subjects using a new high-resolution MRI technique in an attempt to define objective criteria of normalcy. METHODS: A new high-resolution imaging technique, T2-reversed MRI on a high-field (3.0-T) system was used to image the hippocampus in 74 normal volunteers in 3 age groups (28 young, 24 middle, and 22 senior). RESULTS: Residual cavities in the vestigial hippocampal sulcus were resolved as single, crescent-shaped structures along the deep aspect of the vestigial hippocampal sulcus. The size of the cavities did not vary with age and never exceeded 1 mm in width and 3 mm in length. The frequency of detection, however, increased with age (21.4% [5/28] in the young, 25.0% [6/24] in the middle, and 36.4 [8/22] in the senior age group). CONCLUSION: The study established clear objective criteria for normal cavities within the hippocampus. These cavities likely represent physiological fluid collection within the vestigial hippocampal sulcus. Any cavity that does not meet these defined criteria should be considered potentially abnormal. PMID- 12116742 TI - Insonation method and diagnostic flow signatures for transcranial power motion (M mode) Doppler. AB - BACKGROUND AND PURPOSE: Power motion mode Doppler (PMD) simultaneously displays flow signal intensity and direction over several centimeters of intracranial space. Insonation protocol for PMD and spectral transcranial Doppler (TCD) with typical PMD flow signatures is described in serial patients with acute stroke symptoms examined via conventional windows with a PMD/TCD unit. RESULTS: Thirty five patients were studied within 12 hours after stroke onset (age 64 +/- 15 years; 8 received intravenous and 3 intra-arterial thrombolysis). One patient had no temporal window, and 3 patients had suboptimal windows. In 90% of patients, PMD showed more than 1 ipsilateral temporal windows. In 63% of patients (n = 22), PMD simultaneously displayed the entire M1 (65-45 mm) and proximal M2 (45-30 mm) flows, leading to spectral TCD examination of the proximal M2 middle cerebral artery (MCA) in 28 of 35 patients (80%). All patients had sufficient foraminal (depth display = 60-110 mm) and orbital (depth display = 30-80 mm) windows. PMD displayed the entire basilar artery stem (75-100+ mm) in 69% (n = 24) of patients, and the distal basilar flow was detected in all patients by both PMD and TCD. TCD results were normal (12), proximal intracranial stenosis (5), large vessel occlusion (17), and cerebral circulatory arrest (1). Compared to spectral TCD, PMD signatures of similar diagnostic significance were low resistance (vessel identification and recanalization), high resistance (ophthalmic artery identification and distal obstruction), collateral (communicating arteries and leptomeningeal flow), reverberating (circulatory arrest), and branch embolization. CONCLUSIONS: PMD is a window-finding tool and a guide for spectral TCD gate placement. PMD facilitates flow detection in the M2 branches and the distal basilar artery. PMD can demonstrate recanalization of the entire MCA main stem and proximal branches, increase the yield of embolus detection and procedure monitoring, and facilitate abnormal flow pattern recognition. PMID- 12116744 TI - Transcranial Doppler study of the cerebral hemodynamic changes during breath holding and hyperventilation tests. AB - BACKGROUND AND PURPOSE: The aim of the present study was to assess the time course of hyperventilation (HV) and breath-holding (BH) tests in healthy volunteers. SUBJECTS AND METHODS: Young healthy volunteers (n = 29) underwent continuous registration of the middle cerebral artery mean blood flow velocity (MCAV) during and after 30 seconds of BH and 60 seconds of HV. Absolute values as well as percentage changes of the MCAV are reported. In 13 subjects, determination of capillary blood gas parameters (pH, pCO2, pO2, and O2 saturation) was performed before tests, after BH and after HV. RESULTS: MCAV during 30 seconds of breath-holding starts to increase after 10 seconds and reaches its highest level at 30 seconds. After breathing normally, MCAV normalizes within 30 seconds. Hyper-ventilation results in a decrease in MCAV, which reaches a plateau at 20 to 30 seconds after starting to hyperventilate, and blood flow velocity did not change significantly any further until the end of the procedure. The normalization of the MCAV is incomplete at 30 seconds after finishing hyperventilation. None of the capillary blood gases changed significantly after breath-holding, whereas capillary pH, pO2, and oxygen saturation increased and pCO2 decreased after hyperventilation. No relationship was found between capillary blood gas parameters and MCAV values. CONCLUSIONS: The authors concluded that breath-holding and hyperventilation tests seem to be a practical alternative to acetazolamide and the CO2 inhalation method in the assessment of cerebral hemodynamics. PMID- 12116743 TI - Volumetric assessment of plaque progression with 3-dimensional ultrasonography under statin therapy. AB - BACKGROUND AND PURPOSE: Lowering of serum cholesterol levels with HMG-CoA reductase inhibitors (statins) slowed the progression of atherosclerosis in the carotid arteries in several clinical trials using carotid artery intima media thickness as primary outcome measure. Whereas conventional ultrasonography is limited to thin 2-dimensional image planes, 3-dimensional (3D) ultrasonography provides quantitative measurement of the entire carotid artery plaque volume. This study aims to assess the feasibility of 3D ultrasonography to monitor plaque progression in hypercholesterolemic patients. METHODS: The authors prospectively assessed the progression of 31 carotid artery plaques over 15.1 +/- 4.5 months in a study of 23 patients (6 women, 17 men; mean age = 61.7 +/- 7.5 years) with hypercholesterolemia under therapy with HMG-CoA reductase inhibitors. All patients were maintained on a lipid-lowering diet. Sixteen patients were additionally treated with statins. Quantitative measurements of carotid artery plaque volumes were performed after 3D reconstruction of exactly parallel transverse duplex ultrasound scans (slice distance = 0.1 mm) into volumetric 3D data sets and segmentation of voxels representing the carotid artery plaque. RESULTS: Within the treatment group, plaques were significantly less frequently progressive if they had a hypoechoic echogenicity (11%, n = 9 vs 64%, n = 14; P = .016) or if baseline serum cholesterol levels were above 8.0 mmol/L (9%, n = 11 vs 75%, n = 12; P = .002). CONCLUSION: Three-dimensional ultrasonography extends the measurement of the arterial wall thickness to the 3D volume of an entire atherosclerotic plaque including analysis of its morphology and configuration. However, further clinical trials with an adequate sample size to achieve sufficient statistical power are necessary to assess the effect of statin therapy on plaque progression. PMID- 12116745 TI - Transcranial Doppler characteristics of different embolic materials during in vivo testing. AB - PURPOSE: The authors investigated whether ultrasonic characteristics of embolic signals could be used to differentiate embolic composition. MATERIALS AND METHODS: The authors analyzed high-intensity transient signals (HITS) from 3 patients with patent foramen ovale during the bubble contrast test and during total joint replacement surgery. In 3 anesthetized dogs, latex microspheres, fat particles, and air bubbles were injected into the internal carotid artery and HITS were identified in the cerebral circulation. The area under the receiver operating characteristic curve quantified the usefulness of each measure to distinguish embolic composition. RESULTS: In humans, HITS intensity (area: 0.80) and frequency (area: 0.73) but not duration (area: 0.32) were useful to distinguish air bubbles from presumed solid emboli. In animals, intensity distinguished microspheres from air (area: 0.94) and microspheres from fat (area: 0.94) but was less useful for fat and air (area: 0.64). The duration (area: 0.54 0.76) and frequency (area: 0.54-0.63) were poor discriminators. CONCLUSION: The HITS intensity best distinguished embolic composition. Particle size should be taken into account in future research. PMID- 12116746 TI - Diffusion-weighted magnetic resonance imaging in superior sagittal sinus thrombosis. AB - BACKGROUND AND PURPOSE: Diffusion-weighted imaging (DWI) has shown high sensitivity in the diagnosis of acute arterial strokes. The pathophysiology of cerebral venous thrombosis with associated venous stroke appears to differ from that of arterial strokes. The purpose of this study was to describe DWI findings in venous strokes. METHODS: The authors reviewed 3 adults with superior sagittal sinus thrombosis who underwent DWI and magnetic resonance imaging within 24 hours of symptom onset. DWI was obtained at 1.5 T using the multishot echo planar technique (TR = 8000, TE = 97, field of view = 30 x 19 cm, slice thickness = 6.0 mm, interslice gap = 0.5 mm, matrix 128 x 128, NEX = 1). The diffusion gradients were applied in 3 orthogonal directions with 3 increasing b values (0-1000 s/mm2) to create average (trace) DWI images. Apparent diffusion coefficient (ADC) values were calculated on a pixel-by-pixel basis and displayed as ADC maps. RESULTS: DWI showed hyperintensities in patients 1 and 2 and hypointensity in patient 3 in corresponding to parenchymal lesions on conventional images. As compared to the homologous uninvolved location in the contralateral hemisphere, ADC values were decreased (0.53 x 10(-3) mm2/s [patient 1] and 0.68 x 10(-3) mm2/s [patient 2]) and increased (1.1 x 10(-3) mm2/s [patient 3]). The ADC ratio of the lesion in the involved to uninvolved side was 88% (patient 1), 81% (patient 2), and 120% (patient 3). CONCLUSION: Acute cerebral venous strokes may contain cytotoxic edema and/or vasogenic edema on DWI scans. DWI may be helpful in diagnosing cerebral venous thrombosis in cases with cryptic presentations. PMID- 12116747 TI - Transient focal leptomeningeal enhancement in Sturge-Weber syndrome. AB - The authors describe a 36-year-old man with Sturge-Weber syndrome who presented with focal seizures and subsequently developed a temporary post-ictal hemianopia. Magnetic resonance imaging of the brain demonstrated focal leptomeningeal enhancement, which subsequently resolved. PMID- 12116748 TI - A case of adult influenza A virus-associated encephalitis: magnetic resonance imaging findings. AB - A 27-year-old man presented with fever, convulsive seizure, and sudden impairment of consciousness. Magnetic resonance imaging (MRI) abnormalities were found in the bilateral thalami, including the brain stem and white matter. The possibility of a previous influenza A virus infection was considered, and cerebrospinal fluid cells and interleukin-6 were elevated. The MRI findings closely resembled those found in cases of childhood acute necrotizing encephalopathy (ANE). The present case suggests that adult influenza A virus-associated encephalitis/encephalopathy or ANE can occur during winter influenza epidemics. PMID- 12116750 TI - Results on coregistration of mediotemporal 18F-fluoro-2-deoxy-D-glucose-positron emission tomography (FDG-PET) hyperactivity and 3D magnetic resonance imaging hyperintense lesions in limbic encephalitis. PMID- 12116749 TI - Reorganization of motor representation in a patient with epilepsia partialis continua as shown by [O15]-labeled butanol positron emission tomography and functional magnetic resonance imaging. AB - The authors investigated a 38-year-old patient with focal cortical dysplasia in the right precentral cortex using positron emission tomography and functional magnetic resonance imaging to localize the hand and finger motor representations. The patient presented clinically with epilepsia partialis continua, supposed to originate from the perirolandic area harboring the cortical malformation. Both methods revealed an abnormal bilateral activation of motor cortex during left hand finger movements. The results suggest that the so-called eloquent but nevertheless pathological dysplastic cortex accommodates motor representations. PMID- 12116751 TI - Cost effectiveness of wound management in long-term care. AB - Today's healthcare environment -- and the economic environment in general -- necessitate efforts to lower healthcare costs, both for patients and for providers. This need is especially relevant to long-term care facilities, where residents' healthcare problems are a daily and expensive certainty. Both pressure ulcers and venous ulcers are common in long-term facilities, and the aging of our citizens means managing these facilities will be an increasingly significant concern for providers. PMID- 12116752 TI - It's tough to swallow: a practical approach to nutritional care of dysphagia. AB - Have you ever swallowed a food or liquid only to have it "go down the wrong way"? If you have, then you have an idea of what it might be like to have a swallowing problem. Now multiply that experience times the more than 600 times a day that we normally swallow! Imagine how difficult it would be to deal with that on a daily basis. Approximately 6-15 million Americans are affected by dysplagia (chewing and swallowing problems). About 53-74% of nursing home residents, 14% of hospital patients and 33% of rehabilitation center patients have some form of dysphagia, which can have a dramatic impact on nutritional status. PMID- 12116754 TI - Integrative care--controlling inflammatory conditions with diet. PMID- 12116753 TI - New treatment options for overactive bladder and incontinence. PMID- 12116755 TI - [Theory of space-time organization of biological systems]. PMID- 12116756 TI - [Pathological systems in CNS performance]. AB - The occurrence of new pathological integrations from the altered nervous structures in the central nervous system (CNS) is a highly important mechanism of nervous disorders. At the level of systemic relations, such an integration represents a new pathodynamic arrangement from the primarily and secondary altered formations of the CNS. The results of its performance are of a biologically negative, pathogenic value for the body. So such a pathodynamic arrangement is determined by the author as a pathological system (PS). PS substantially and basically differs from the physiological system in the mechanisms of occurrence, clinical manifestations, and performance results. PS is formed and fixed due to the plastic processes intrinsic to the nervous system, under the influence of the pathological determinant the hyperactive CNS formation that can also determine the nature of PS activity. The pathological systems are a pathobiological basis and a pathophysiological mechanism of neuropathological syndromes that are a clinical manifestation of PS performance. Each neuropathological syndrome has its PS. The pathological systems may consist of different formations, levels, and spheres of CNS performance, which determines the content of neuropathological syndromes, including those applied to higher nervous performance, behavior, and the mental sphere. PMID- 12116757 TI - [Evolutionary aspects of a stress reaction]. AB - Interconnections between the integrative systems, such as nervous, endocrine, and immune ones, are clearly seen during a stress response. The sources of such a cooperation should be sought at the earliest stages of evolution of multicellular organisms. The paper deals with the main points of the formation of a stress response and the development of the systems involved in this process. Based on the data available in the literature and their own findings, the authors conclude that just primitive multicellular organisms, such as coelenterates, have hemolymph cells (chemocytes) that combines the properties of all regulatory systems of more highly organized species. However, the need for such cells no longer arises afterwards. Glucocorticoids, one of the major stress hormones, show up during the development of a stress response relatively late--in fish. While in fish, the production of ACTH and glucocorticoids in the pituitary cells is under urotensin control, this function belongs to corticotropin-releasing hormone of pituitary origin in more advanced animals. The predominant value of corticotropin releasing hormone simultaneously occurs with the development of the pituitary portal vascular system. Such transformations lead to the formation of the hierarchically arranged neuroendocrine axes that are responsible for showing a stress response, on the one hand, and to the development of qualitatively new interactions between the nervous, endocrine, and immune systems, on the other. PMID- 12116758 TI - [Ultrastructure of human cerebral cortical synapses: age-specific aspects]. AB - The paper deals with electron microscopic studies of the cerebral cortex in human beings aged 70-80 years. The synapses show a decrease in the number of synaptic vesicles, their impaired distribution in the presynaptic process. There are changes in the mechanisms of interaction of synaptic vesicles and a presynaptic membrane. In the latter, the vesicles loose their discreteness, some of them are replaced by a fine-granular material. The joining of the vesicles and membrane is impaired. A part of active synaptic areas is transformed to desmose-like contacts. The findings are indicative of synaptic dysfunction. PMID- 12116759 TI - [Structural bases of individual variability in the human brain]. AB - The paper shows considerable differences in the cytoarchitectonical organization of cortical fields 44, 45, and 4, 6 in the brain of different persons. The profile fields of pyramidal neurons in layers III and V substantially differ from each other for the cortical fields under investigation. The specific features of the total fractions of neurons and glia in the cortical fields in different brains are outlined. The density of neuronal location is an important criterion for individual variability. The magnitude of individual variability manifests itself the right and left hemispheres by different ways. PMID- 12116761 TI - [Rules of interactions and functional continuum of neuropeptides (on the way to the common conception)]. AB - The biological activity of neuropeptides (NP) is extremely wide and, moreover, there are thousands of peptides. Due to the pronounced multifunctionality of each peptide and to the overlapping reduplication of bioactivity spectra of each peptide, a functionally persistent aggregate (i.e. the functional continuum of NP) is formed. The method of vector representation of NP effects is outlined, which makes it possible to classify the multitude of NP according to functional characteristics, to reveal the evolutionary rules of development of the peptide system for regulation of biological parameters, and to indicate the constellations of regulatory peptides that are the most promising to be studied from the medical point of view. PMID- 12116760 TI - [Involvement of the neuropeptide cholecystokinin in the mechanisms of regulation of emotions and craving]. AB - The paper presents an idea on the organization and functions of the cholecystokinin (CCK) system. Particular emphasis is laid on the modulating influence of CCK on dopamine neuromediation in the mesolymbic structures of the brain, which are linked with the regulation of emotions and craving. Experimental and clinical studies have indicated that CCK and its preparations arrest the major manifestation of the withdrawal syndrome, including pathological craving for alcohol, and anxiety. In the post-withdrawal period, these drugs also effectively suppress alcohol craving. The designed new tetrapeptide, a CCK analogue, selectively inhibits anxiety in animals with this emotional disorder, which is followed by normalization of dopamine exchange and GABA-system functions. It is concluded that by interacting with the dopamine-system, CCK takes a direct part in the regulation of emotions and craving. PMID- 12116763 TI - [Scientific school of Anohin-Sudakov]. PMID- 12116762 TI - [Neurotrophic factors and autoantibodies to them as molecular predictors of cerebral dysfunctions]. AB - A complex of clinical and immunochemical studies was made in patients with chronic brain ischemia and ischemic stroke and in neonatal infants with CNS dysfunctions and retarded intrauterine development. Enzyme immunoassay was used to measure the levels of brain proteins with trophic properties--S100b, the major protein myelin, lectins CSL, R1, and the levels of primary and antiidiotypic antibodies to these proteins in the biological fluids of the patients. The findings suggest that the study brain proteins and autoimmune processes against these factors are involved in the mechanisms of the pathogenesis of the diseases in question and they enable changes and variations in the levels of neurotropic factors and their autoantibodies to be considered as predictors of brain ischemia and perinatal cerebral lesions. PMID- 12116764 TI - [Regulation of the human cardiovascular system under microgravitation]. AB - The paper presents the results of studies of the cardiovascular system and analyzes the mechanisms of regulation and adaptation of this system during long term space flights. Special attention is given to the analysis of the homeostatic mechanisms for maintaining a number of integrative parameters of the cardiovascular system at the pre- and near-flight level and to the mechanisms of development of orthostatic detraining character after flights. How a new level of circulatory function in the absence of microgravitation is formed is shown. PMID- 12116765 TI - [Features of informational hemopoiesis-regulating signalization]. AB - The paper presents the currently available data on hemopoiesis-regulating signalization, on the reception of hemopoietic hormones, on the role of nerve signalization and extracellular matrix in the development of a hemopoietic response adequate to environmental influence on the body. PMID- 12116766 TI - [Time course of changes in cardiac rhythms in successful and unsuccessful solution of visual and motor tasks under the conditions of binocular and monocular visions]. AB - The P. K. Anokhin theory of the general functional system considers integral behaviour as realization of processes aimed at reaching a useful result. Electrograms were recorded in students who were solving visual and motor puzzle tasks. Their results were estimated. There were two groups of subjects: 1) those with normal binocular vision and 2) those with monocular vision. Some specific features were found in cardiac rhythm power in each group. The findings suggest that there are relationships between the results of task solution, the type of visual perception, and heart rate parameters. PMID- 12116767 TI - [Informational facets of vital activity]. AB - The paper considers informational processes occurring within and between the body's functional systems. Informational demand-supply relations are constructed by the holographic principle. The apparatuses of outcome acceptors act as holographic informational screens in the functional systems. A concept of the non specific informational syndrome of disintegration of different functional systems upon extreme exposures of human beings is formulated. Principles in non-drug rehabilitation of broken inter- and intrasystemic informational bonds are outlined. PMID- 12116768 TI - The intrinsic force control of functional treatment. PMID- 12116769 TI - Accelerated orthodontics--adults demand it! PMID- 12116770 TI - Bond failure analysis. PMID- 12116771 TI - Refractive patients: TMJ splint therapy combined with open joint surgery 60 patients; 117 joints. PMID- 12116772 TI - [Treating congestive heart failure with pacemaker]. PMID- 12116773 TI - [Law about the embryo research comes into force]. PMID- 12116774 TI - [Tinnitus]. PMID- 12116776 TI - [Challenges in the psychic development of an adopted child]. PMID- 12116775 TI - [Treatment of aortic aneurysms with endovascular stent prostheses]. PMID- 12116777 TI - [Williams syndrome--developmental syndrome's common cardiovascular disorders]. PMID- 12116778 TI - [Nuchal translucency in the fetus and congenital heart defects]. PMID- 12116779 TI - [Alcoholism and gait dysfunction]. PMID- 12116780 TI - [The skier's thumb]. PMID- 12116781 TI - [Inflammation around the insulin injection sites]. PMID- 12116782 TI - [How does the analysis of flowering plant Arabidopsis thaliana genes benefit medicine?]. PMID- 12116783 TI - [Year 2001 Matti Ayrapaa award to Riitta Hari]. PMID- 12116784 TI - [Magnetoencephalography as an instrument for the scientist in the field of brain research]. PMID- 12116785 TI - [Thrombocytopenia and pregnancy]. PMID- 12116786 TI - [Endometriosis]. PMID- 12116787 TI - [Stenotic heart valves can be reopened with balloon dilatation]. PMID- 12116789 TI - [Abdominal pain caused by mesenteric panniculitis]. PMID- 12116788 TI - [Actinomycosis--a rare cause of purulent peritonitis in a woman in fertile age]. PMID- 12116790 TI - [Child and recurrent infections]. PMID- 12116791 TI - [Deadly tired]. PMID- 12116792 TI - [What was behind the brain infarctions?]. PMID- 12116793 TI - [Does radiotherapy help against painful arthrosis?]. PMID- 12116795 TI - [Correction to the article about the diagnostics of milk allergy]. PMID- 12116796 TI - [Spinal cord injury]. PMID- 12116798 TI - The role of endomyocardial biopsy in the diagnosis of cardiomyopathies. AB - Though many publications in the field of myocarditis and cardiomyopathies have renewed interest in the value of endomyocardial biopsy, its role in the work-up of patients with cardiomyopathies, idiopathic arrhythmias and even ischemic heart disease, is still debated. Since its introduction in 1963 the technique has been developed with current routine use of a biventricular and even atrial approach. At the same time immunohistochemistry and molecular biology studies have greatly enhanced the information obtainable from myocardial samples. The authors report their experience and briefly review the pertinent literature on the current use of endomyocardial biopsy in the diagnosis and treatment of dilated, hypertrophic and restrictive cardiomyopathy, of idiopathic arrhythmias and in ischemic heart disease. PMID- 12116797 TI - Life expectancy and quality of life in adult patients with congenital heart disease. AB - The survival and quality of life of patients with congenital heart disease have significantly improved in the last 20 years. This is due to more effective medical and surgical care. The new community of grown-up congenital heart patients consists of a few natural survivors with trivial congenital lesions or very rare complex cardiac abnormalities which are naturally compensated, and of more than 75% of patients who had been submitted to cardiac surgery during infancy or childhood. Clinical follow-up is however mandatory for many of them with scheduled times and types of exams to control the effects of sequelae and late complications, and to prevent deterioration and premature death because cardiac surgery may not have resulted in normality. Moreover, these patients have many needs and even more, many questions. Not giving a correct answer to each specific question reduces the entity of surgical success. PMID- 12116799 TI - Transesophageal electrical cardioversion of persistent atrial fibrillation: a new approach for an old technology. AB - BACKGROUND: Low energy intracardiac cardioversion may be considered the elective, alternative method for the acute restoration of sinus rhythm when direct current cardioversion fails or is contraindicated. Transesophageal cardioversion is a further alternative method for the recovery of sinus rhythm and obviates the potential complications of the low energy intracardiac cardioversion venous approach. METHODS: The present prospective study including 30 patients (21 males, 9 females, mean age 65.1 years, range 52-76 years), with persistent atrial fibrillation (mean duration 4.3 months), was undertaken in order to further evaluate, with regard to transesophageal cardioversion: 1) the acute efficacy, 2) the patient acceptance of the procedure, 3) the preferable choice among direct current cardioversion, low energy intracardiac cardioversion and transesophageal cardioversion, 4) the time required to perform the procedure, 5) the incidence of complications, and 6) the persistence of sinus rhythm after 1 month. RESULTS: Sinus rhythm was acutely restored in 29 patients (96.7%). Discomfort induced by the electrical shock was minimal or mild in most patients (75.8%). Transesophageal cardioversion was usually preferred by patients who had been previously submitted to direct current cardioversion or low energy intracardiac cardioversion. The mean total time required to perform the procedure was 107.9 min. No complications related to the procedure occurred. In spite of adequate pharmacological prophylaxis of atrial fibrillation only 41.4% of patients were in sinus rhythm 1 month after successful transesophageal cardioversion. CONCLUSIONS: Transesophageal cardioversion may be considered a very effective, well accepted and non-time consuming procedure for the short-term restoration of sinus rhythm. The incidence of complications is low. PMID- 12116800 TI - Usefulness of three-dimensional non-fluoroscopic mapping in the ablation of typical atrial flutter. AB - BACKGROUND: Catheter ablation of the cavo-tricuspid isthmus is rapidly becoming the first line of treatment in the management of atrial flutter. The standard procedure is usually performed under fluoroscopy guidance and relays on multisite endocardial recordings to assess the completeness of the isthmus conduction block. Despite the high rate of success there is, at follow-up, a considerable number of recurrences which could reflect the limitations of conventional assessment of conduction block across the isthmus. This new non-fluoroscopic mapping system allowing high density mapping along the entire length of the ablation line, could provide a more accurate way of verifying complete conduction block. The aim of the present study was to describe our overall results and long term follow-up using a three-dimensional mapping system to guide radiofrequency ablation of typical atrial flutter. METHODS: A multipoint three-dimensional map of the cavo-tricuspid isthmus, septal and lateral atrial wall was performed in 87 patients prior to and following ablation for typical atrial flutter. Evidence of persisting gaps in the line of block was identified by visual inspection of the color-coded activation maps and these sites were re-ablated. The conduction sequence was also assessed with conventional bidirectional pacing and recording. The assess the reduction in fluoroscopy time, two groups of patients were compared: group A (14 patients) in whom the entire mapping-ablation procedure was guided by the three-dimensional system (Carto, Biosense-Webster, Diamond Bar, CA, USA) and group B (32 patients) in whom the same protocol was used but the procedure was guided by standard fluoroscopic imaging. RESULTS: Acute success was achieved in every patient. During bilateral isthmus pacing, the mean local activation time increased from 20.3 +/- 13.3 ms pre-ablation to 148.3 +/- 53.2 ms (p < 0.01) post-ablation with a mean difference of 120 +/- 31 ms. In 11 patients (9.2%) there was evidence of persisting conduction across the line of block despite evidence of reverse activation of the cavo-tricuspid isthmus by conventional pacing. A gap in the ablation line was identified and re-ablated. At a mean follow-up of 16.3 +/- 2.2 months, there were 5 (4.2%) recurrences of atrial flutter and 12 (10%) recurrences of isolated atrial fibrillation. Four of the 5 recurrences occurred in patients in whom ablation was guided by conventional fluoroscopy (group B). The fluoroscopy time was 4.2 +/- 1.5 min in group A and 27.2 +/- 8.2 min in group B (p < 0.001). CONCLUSIONS: Multipoint mapping of the ablation line following radiofrequency ablation of typical atrial flutter performed using the Carto system allows a more accurate assessment of the isthmus conduction block. This has the potential to reduce the recurrence rate to the level observed for other supraventricular tachycardias. PMID- 12116801 TI - Endoluminal stent grafting of the descending thoracic aorta. AB - BACKGROUND: The timing and optimal therapy for descending thoracic aortic diseases still remain a challenging problem for surgeons. Nowadays endovascular treatment is becoming more and more popular both for acute as well as chronic cases. This technique is more respectful of the tissue integrity and avoids major and demolitive surgery for the patient. METHODS: In 1 year 32 patients presenting with descending thoracic aorta dissection (n = 25) or with descending thoracic aorta aneurysms (n = 7) were submitted to an endovascular procedure using covered stents. Ten of them were operated upon in general anesthesia whereas in 22 spinal anesthesia was administered. In neither group did anesthesia-related complications occur. RESULTS: In all cases in which endovascular treatment was possible, an endovascular stent was used for the treatment of the descending thoracic aorta disease. Only 1 patient had a major complication, which was a retrograde dissection of the ascending aorta surgically treated in an emergency setting. Our policy is to treat uncomplicated type B dissections in the subacute phase after 1 week of antihypertensive pharmacological treatment, but within 1 month of onset. Our mid-term follow-up shows very good results with no mortality and no stent-related complications. CONCLUSIONS: Stent grafting is replacing conventional surgery for descending thoracic aorta aneurysms and dissections. Our results suggest that in case of dissections, endovascular treatment should be delayed until the subacute phase, in the absence of complications. The risks and mortality are decreased. PMID- 12116802 TI - Surgical restoration of the left ventricle for postinfarction aneurysm. AB - BACKGROUND: Surgical left ventricular reduction is under investigation as an alternative to, or a bridge for, heart transplantation in patients with a left ventricular aneurysm. In fact, acute myocardial infarction can result in the development of a dyskinetic or akinetic left ventricular aneurysm which may in turn cause congestive heart failure, ventricular arrhythmias, and the formation of mural thrombi. The aim of this study was to evaluate the current operative risk of surgical restoration of the left ventricle and the early and late clinical results. METHODS: From January 1997 to December 2001, 94 patients (84 males and 10 females) presenting with a postinfarction aneurysm were submitted to surgical restoration of the left ventricle. All patients presented with symptoms of heart failure and/or angina. The preoperative NYHA functional class was: I in 6 patients, II in 22 patients, and III in 66 patients. No patient was in NYHA class IV at the time of surgery. The preoperative ejection fraction was 30 +/- 7.9%. In 25 patients mural thrombi were identified and surgically removed. In patients with preoperative evidence of ventricular arrhythmias the Harken procedure was performed intraoperatively. The ventricular preoperative and postoperative performances were also studied in 10 patients using P-V loops obtained through a conductance catheter. RESULTS: The in-hospital mortality was 3.2%. The mean length of hospitalization was 7 +/- 2.9 days. At follow-up (mean 26 +/- 14.8 months) we observed an early improvement in the ejection fraction (30 +/- 7.9 vs 48 +/- 8.0%) and a decrease in the end-diastolic and end-systolic volumes and mean pulmonary pressure (139 +/- 37 vs 84 +/- 17 ml/m2, 105 +/- 39 vs 52 +/- 20 ml/m2, 35 +/- 8.4 vs 23 +/- 4.3 mmHg). CONCLUSIONS: These results suggest that ventricular restoration is indicated in all patients with a postinfarction dyskinetic or akinetic aneurysm. The operation, if performed appropriately, is associated with a low in-hospital mortality and morbidity. A postoperative improvement in the early and long-term cardiac functions was demonstrated. An improvement in symptoms and quality of life was documented, increasing our expectations of an increased long-term survival. PMID- 12116803 TI - A rare cause of cardiogenic shock: catecholamine cardiomyopathy of pheochromocytoma. AB - Pheochromocytoma is a rare catecholamine secreting tumor that accounts for about 0.04% of cases of hypertension. Other less common cardiovascular manifestations such as arrhythmias, angina pectoris, acute myocardial infarction, dilated cardiomyopathy, acute heart failure, and cardiogenic shock have occasionally been reported. We describe the case of a 32-year-old previously healthy male patient who died of cardiogenic shock within 10 hours of admission. Postmortem examination showed a catecholamine cardiomyopathy and a pheochromocytoma of the right adrenal gland. Pheochromocytoma with predominant epinephrine or dopamine secretion may take a hypotensive course. Sudden excessive catecholamine release can, as in the described case, cause cardiogenic shock. PMID- 12116804 TI - Mustard procedure in atrial situs inversus: three-dimensional ablation of intra atrial reentry tachycardia on a surgically created inversus cavo-tricuspid isthmus. AB - It has been demonstrated that the Biosense Carto system can improve the success rate of ablation in case of an intra-atrial reentry tachycardia in patients submitted to the Mustard repair. This system was used to map an intra-atrial reentry tachycardia in a young patient who had been submitted to the Mustard procedure for atrial situs inversus. A line of block was created connecting the right sided tricuspid valve to the left sided inferior vena cava. This terminated the arrhythmia and prevented its re-initiation. This case confirmed the notion that the cavo-tricuspid isthmus is often critical to the maintenance of an intra atrial reentry tachycardia after the Mustard procedure even if its location is in the inversus side. PMID- 12116805 TI - Dilated cardiomyopathy and celiac disease. PMID- 12116806 TI - [Contemporary university: a betrayed tradition?]. AB - The current evolution of University is examined in parallel with the evolution of society. Society of uncertainty, crisis of the social actor, selective mechanisms of the elite, fundamental role of technical predominance and of economical processes are analyzed based on the "tradition" concept. The possibility of an authoritative comeback of the role of University is found in an active and articulated intervention of integration and cultural mediation, substantiating the bridge of tradition. PMID- 12116807 TI - [Fibril-forming proteins: the amyloidosis. New hopes for a disease that cardiologists must know]. AB - Several proteins share the property of conforming as antiparallel beta-sheets, and forming insoluble amyloid fibrils that deposit in the interstitium of organs/tissues and cause systemic amyloidosis. Cardiac involvement is frequent and constitutes a major predictor of poor outcome. Its typical phenotype is that of restrictive cardiomyopathy. The biochemical classification of the amyloidogenic proteins provides the bases for innovative therapeutic approaches. Primary systemic amyloidosis (AL) is a protein conformation disorder in which monoclonal immunoglobulin light chains (kappa or lambda) produced by clonal plasma cells, are deposited as amyloid in kidneys, heart, liver, and other organs. The recent evidence that chemotherapy reduces or even eradicates the amyloidogenic clone with consequent functional improvement of the affected organs raises new hopes for a treatment, whose key of success is early diagnosis. Heart transplantation can be proposed in patients < 60 years of age in association with autologous stem cell transplantation. In serum amyloid A amyloidosis, fibrils are constituted of the acute phase serum amyloid A protein that is produced in excess in chronic inflammatory diseases such as familial mediterranean fever, autoimmune disorders and chronic infections. The strategy is to treat the underlying inflammatory disease, but new molecules inhibiting amyloid formation and promoting amyloid resorption are facing the clinical scenario and trials are in progress. In transthyretin (TTR) amyloidosis, the non-senile forms are autosomal dominant diseases caused by defective proteins synthesized by mutated TTR genes (more than 70 known mutations with different genotype-phenotype correlations). The treatment is based on transplantation of the TTR-producing liver; exceptionally, liver plus heart or kidney are transplanted. Apolipoprotein A1 amyloidosis is an inherited autosomal dominant disease that benefits from the transplantation of the most impaired organs, usually heart, liver or kidney, either single or combined. The diagnosis of apolipoprotein A1 and TTR amyloidosis relies on positive family history, immunocharacterization of the amyloid fibrils in a tissue biopsy, gene defect detection and absence of light chains in serum and urines. Vice versa, non-familial primary amyloidoses are diagnosed when kappa or lambda light chains are identified with immunofixation in serum or urines. Tissue studies provide the gold standard for the diagnosis and immunocharacterization of amyloid protein. Heart involvement is diagnosed with a multiparametric approach that includes clinical, electrocardiographic and echocardiographic evaluation. The fine-needle biopsy of the periumbilical fat is the preferral procedure for amyloid detection and immunocharacterization of amyloid protein. This approach excludes, with a few exceptions, the need of endomyocardial biopsy. PMID- 12116808 TI - [The appropriateness of diagnostic angiography in cardiology]. AB - Coronary angiography is the most frequently performed invasive diagnostic test in the western world, but regional differences are common and have been outlined in both observational and randomized studies. Appropriateness evaluation is hence fundamental, as the use of invasive cardiac procedures is strongly associated with the population-based availability of catheterization facilities. A procedure is judged appropriate if the expected health benefit exceeds the possible negative consequences by a sufficiently wide margin; it is necessary when not performing it could result in harm for the patient. In the first period (1980 1995) researchers seemed to try to find an explanation for geographic variations regarding overuse: the appropriateness remained at the same level through time (75%), while a trend towards a reduction in the number of inappropriate procedures (< or = 20%) and an increase in that of the uncertain ones was evident. The different opinions of the expert panels constituted the major cause of variability. The factors mainly affecting the appropriateness were advanced age, angina class, intensity of medical therapy, exercise test results and income. Canadians and Europeans seemed to request a higher standard of scientific evidence as compared to US doctors; surgeons tended to give higher scores than cardiologists and internists. Inappropriate indications were similar in high- and low-use hospitals. More recently, important data emerged on the lower than necessary use of this procedure and this was more evident in hospitals without on site catheterization facilities and in patients without fee-for-service insurance. Patient selection was suboptimal and coronary angiography was more frequently performed in low-risk populations. This phenomenon is of concern, because the lower than necessary use of indicated procedures can bear on the patients' outcome. In fact, an inverse relationship between mortality and coronary angiography use has been observed, especially in patients in whom it has been judged necessary. No lower than necessary use of differences in appropriateness have been found in females, but this is possible in ethnic minorities. If the appropriateness is to be improved, specific actions have to be directed to increase the know-how of doctors, patients, and administrators, to promote research in the fields where knowledge is still missing and to implement simplified guidelines and appropriateness criteria, in order to favor a more extensive use. It is mandatory to assess the necessity of coronary angiography procedures and to grant access to those patients who meet the necessity criteria. PMID- 12116810 TI - [Evaluation of the appropriateness of prescribing echocardiography]. AB - BACKGROUND: We evaluated the appropriateness of indications to echocardiography for ambulatory patients performed during 4 weeks in 21 laboratories in Tuscany and Umbria, Italy. METHODS: We collected the following data: the appropriateness of the prescription (according to the guidelines of the Italian Federation of Cardiology), the prescribing physician (cardiologist vs non-cardiologist), the synthetic result (normal vs abnormal) and the clinical utility (useful vs useless) of each exam. RESULTS: We evaluated 2848 prescriptions (patients: 1450 males, 1398 females; mean age 62 years, range 15-90 years). The indications to test were of class I (appropriate) in 43.6%, of class II (of doubtfully appropriateness) in 36.8% and of class III (inappropriate) in 19.6% of the cases. In 60.8% of the cases the exam was considered abnormal. In particular, an abnormal result was found in 83.8% of class I, in 56.6% of class II and in 17.8% of class III exams (p < 0.05). The exam was considered useful in 51.1% of the cases. In particular, a useful result was found in 78.9% of class I, in 39% of class II and in 12.1% of class III exams (p < 0.05). Cardiologists prescribed 856/2848 tests (30%). Their indications were of class I in 58.8%, of class II in 29.8% and of class III in 11.4% of the cases vs 37, 39.9 and 23.1% of non cardiologists' prescriptions (p < 0.05). Abnormal findings were found in 74.3% of cardiologist- vs 55% of non-cardiologist-prescribed examinations (odds ratio 2.45, 95% confidence interval 2.04-2.92; p < 0.05); similarly, clinically useful information could be derived from 63.1% of cardiologist- vs 46% of non cardiologist-prescribed examinations (odds ratio 2.07, 95% confidence interval 1.75-2.45; p < 0.05). CONCLUSIONS: In Tuscany and Umbria, Italy, about half of the prescriptions for echocardiography can be considered inappropriate; appropriately prescribed exams more often provide abnormal and useful results; cardiologist-prescribed exams are significantly more appropriate, abnormal and useful. PMID- 12116811 TI - [Evaluation of the appropriateness of prescribing Holter dynamic electrocardiography]. AB - BACKGROUND: We evaluated the appropriateness of indications to Holter monitoring performed on ambulatory patients during 4 weeks in 21 laboratories in Tuscany and Umbria, Italy. METHODS: We collected the following data: the appropriateness of the prescription (according to the guidelines of the Italian Federation of Cardiology), the prescribing physician (cardiologist vs non-cardiologist), the synthetic result (normal vs abnormal) and the clinical utility (useful vs useless) of each exam. RESULTS: We evaluated 863 prescriptions (population: 435 males, 428 females; mean age 64 years, range 15-90 years). The indications to the test were of class I (appropriate) in 59.6%, of class II (doubtfully appropriate) in 11.7%, and of class III (inappropriate) in 28.7% of the cases. In 33% of the cases the exam was considered abnormal. In particular, an abnormal result was found in 37.9% of class I, in 36.7% of class II, and in 24.5% of class III exams (p < 0.05). The exam was considered useful in 46.7% of the cases. In particular, a useful result was found in 59.2% of class I, in 45.5% of class II, and in 21% of class III exams (p < 0.05). Cardiologists prescribed 373/863 tests (43.2%). Their indications were of class I in 67.6%, of class II in 12% and of class III in 24% of the cases vs 53.7, 11.4 and 34.9% of non-cardiologists' prescriptions (p < 0.05). Abnormal findings were found in 40% of cardiologist- vs 27.6% of non cardiologist-prescribed examinations (odds ratio 1.74, 95% confidence interval 1.31-2.32; p < 0.05); similarly, clinically useful information could be derived from 59.8% of cardiologist- vs 36.7% of non-cardiologist-prescribed examinations (odds ratio 2.56, 95% confidence interval 1.94-3.37; p < 0.05). CONCLUSIONS: In Tuscany and Umbria, Italy, about 40% of Holter exams are inappropriate; appropriately prescribed exams are more often abnormal and useful; cardiologist prescribed exams are significantly more appropriate, abnormal and useful. PMID- 12116812 TI - [Evaluation of the appropriateness of prescribing exercise tests]. AB - BACKGROUND: We evaluated the appropriateness of the indications to exercise testing for ambulatory patients performed during 4 weeks in 21 laboratories in Tuscany and Umbria, Italy. METHODS: We collected the following data: the appropriateness of the prescription (according to the guidelines of the Italian Federation of Cardiology), the prescribing physician (cardiologist vs non cardiologist), the synthetic result (normal vs abnormal) and the clinical utility (useful vs useless) of each exam. RESULTS: We evaluated 1158 prescriptions (population: 822 males, 336 females; mean age 60 years, range 16-82 years). Prescriptions were of class I (appropriate) in 38.9%, of class II (of doubtful appropriateness) in 52.5% and of class III (inappropriate) in 8.6% of the cases. In 14.2% of the cases the exam was abnormal: it was abnormal in 35.5% of class I, in 26.6% of class II and in 23% of class III exams (p < 0.05). The exam was useful in 51.6% of the cases; it was useful in 62.4% of class I, in 50.2% of class II and in 13% of class III exams (p < 0.05). Cardiologists required 596/1158 tests (51.5%). Their indications were included in class I in 45.6%, in class II in 49.7% and in class III in 4.7% of the cases vs 31.7, 55.5 e 12.8% of non-cardiologists' prescriptions (p < 0.05). The test was abnormal in 35.7% of cardiologist vs 23.5% of non-cardiologist-prescribed examinations (odds ratio 1.81, 95% confidence interval 1.4-2.34; p < 0.05); the test was useful in 64.4% of cardiologist vs 38.2% of non-cardiologist-prescribed exams (odds ratio 2.92, 95% confidence interval 2.3-3.71; p < 0.05). CONCLUSIONS: In Tuscany and Umbria, Italy, less than half of exercise testing procedures are appropriate; appropriately-prescribed exams are more often abnormal and useful; cardiologist prescribed exams are significantly more appropriate, abnormal and useful. PMID- 12116813 TI - [Ad-hoc coronary angioplasty: organizational model, clinical results and costs]. AB - BACKGROUND: Our center routinely employs the strategy of ad hoc percutaneous coronary intervention (PCI) after diagnostic catheterization in previously informed and prepared patients with anatomical and clinical indications for some years. The aim of this study was to evaluate clinical results and resource consumption of the ad hoc PCI strategy in our center. METHODS: We evaluated the results and resource consumption of 783 PCIs performed between January 1, 1999 and June 30, 2001, divided into 642 (82%) ad hoc and 141 (18%) deferred PCIs. We analyzed the patients' in-hospital clinical and procedural characteristics, the 1 and 6-month outcomes and resource consumption (costs of materials, quantity of contrast medium, fluoroscopic time and duration of procedures) in the two groups. RESULTS: Patients in the ad hoc group had more frequently previous PCI, hypertension, diabetes, acute coronary syndrome, single vessel disease, single lesion and single vessel PCI, stent use and direct stenting, use of glycoprotein IIb/IIIa inhibitors and hemostatic devices; those in the deferred PCI group had more frequently previous myocardial infarction, stable angina, elective programmed hospital admission for PCI and multilesion single vessel PCI. The clinical results were good: clinical success in 97% of cases, in-hospital major adverse clinical events occurred in 2%, non-Q wave myocardial infarction in 3.4% (creatine-kinase-MB > 3 times higher than the upper normal limit in serial routine controls), major vascular complications in 0.4%, 1-month and 6-month major adverse clinical events in 4 and 9% respectively, without any difference between the two groups. Ad hoc PCI resulted in less contrast medium use, a shorter procedure duration, lower costs and shorter fluoroscopy times with respect to deferred PCI plus diagnostic catheterization, although not statistically significant. CONCLUSIONS: In our experience, ad hoc PCI was safe and effective. Costs were lower and less resources were required. Patients were satisfactorily assisted and the logistics and organization of the procedure were optimal. PMID- 12116809 TI - [Evaluation of the appropriateness of prescribing non-invasive cardiologic tests]. AB - BACKGROUND: We evaluated the appropriateness of the prescription of echocardiography, exercise testing, Holter monitoring and vascular sonography for ambulatory patients, performed during 4 weeks in 21 outpatient laboratories in Tuscany and Umbria, Italy. METHODS: We collected the following data: the appropriateness of the prescription (according to the guidelines of the Italian Federation of Cardiology), the prescribing physician (cardiologist vs noncardiologist), the synthetic result (normal vs abnormal) and the clinical utility (useful vs useless) of each exam. RESULTS: We evaluated 5614 prescriptions (patients: 3027 males, 2587 females; mean age 63 years, range 14-96 years). The indication to the test was of class I (appropriate) in 45.3%, of class II (doubtfully appropriate) in 34.8% and of class III (inappropriate) in 19.9% of the cases. The test was abnormal in 58.3% of class I exams vs 17% of class III exams (p < 0.05). The test was useful in 72.4% of class I exams vs 17.1% of class III exams (p < 0.05). The test was prescribed by a cardiologist in 1882 cases (33.5%). Cardiologist-prescribed exams were of class I in 57.3%, of class II in 32.4% and of class III in 10.3% of the cases vs 39.2, 36.1 and 24.7% of non-cardiologist-prescribed exams (p < 0.05). Cardiologist-prescribed exams were abnormal in 53.4% of the cases vs 39% of those of non-cardiologists' (odds ratio 1.76, 95% confidence interval 1.58-1.97; p < 0.05). Cardiologist-prescribed exams were useful in 64.7% of the cases vs 44.4% of those of non-cardiologists' (odds ratio 2.26, 95% confidence interval 2.02-2.53; p < 0.05). CONCLUSIONS: In Tuscany and Umbria, Italy, less than half of the prescriptions for non-invasive diagnostic tests are appropriate: appropriately prescribed exams more often provide abnormal and useful results; cardiologist-prescribed exams are more often appropriate, abnormal and useful. PMID- 12116814 TI - [Electrical cardioversion for atrial fibrillation: advantages of the transthoracic biphasic method]. AB - BACKGROUND: The purpose of this study was to evaluate efficacy and safety of biphasic shock for atrial fibrillation cardioversion to sinus rhythm. A second endpoint was to evaluate myocardial damage by means of cardiac troponin I dosage. METHODS: We studied 164 patients, with drug-resistant atrial fibrillation (208 episodes). Group A patients underwent biphasic shock normalized with respect to weight: 50 J (weight < 60 kg), 70 J (weight 61-84 kg) and 100 J (weight > 84 kg; the second and third shocks were 2 and 3 times higher than the first. Group B underwent sequential monophasic shock of 200, 300 and 360 J. Troponin I was evaluated at baseline, and 6, 12 and 24 hours after cardioversion. RESULTS: Total efficacy was 92% for biphasic shock and 89% for monophasic shock. First-shock efficacy with biphasic waveform (57.3%) was significantly greater than with first monophasic waveform (21.5%) (p = 0.000). Cardiac troponin I increased from 0.4 +/ 1.1 to 0.8 +/- 2.2 compared to a normal value of 2 ng/ml. CONCLUSIONS: For transthoracic cardioversion of atrial fibrillation, biphasic shock has a greater efficacy requiring less energy compared to monophasic shock. Normal mean values of cardiac troponin I proved the absence of myocardial damage. PMID- 12116815 TI - [Inflammatory and fibrinolytic activation after coronary artery bypass with extracorporeal circulation]. AB - BACKGROUND: The aim of this study was to determine the course of the main inflammatory and fibrinolytic markers in patients undergoing primary elective coronary artery bypass graft with extracorporeal circulation. METHODS: One hundred and thirteen patients (105 males, 8 females) undergoing primary isolated coronary artery bypass with normo- (37 degrees C) or hypothermic (26 degrees C) systemic perfusion were prospectively studied. The clinical course of the patients was recorded and inflammatory and fibrinolytic markers (C-reactive protein, fibrinogen, interleukin-6, plasminogen activator inhibitor-1, prothrombin time, activated partial thromboplastin time, platelets and white blood cell counts) were determined before surgery, 24, 48 and 72 hours thereafter, and at hospital discharge. RESULTS: Two patients died (mortality 1.7%) and 6 had a major complication (event free survival > 94%). Interleukin-6, lymphocyte, neutrophil and monocyte levels increased after surgery but returned to normal at hospital discharge. C-reactive protein levels increased after 24 hours and remained high at hospital discharge. Plasminogen activator inhibitor-1, prothrombin time, and activated partial thromboplastin time increased from few hours postoperatively and returned to normal before discharge. Platelets decreased immediately after surgery and normalized only at hospital discharge. Fibrinogen decreased in the first 24 postoperative hours, raised later and remained elevated at hospital discharge. CONCLUSIONS: Cardiopulmonary bypass activates inflammatory response and hemostatic/fibrinolytic balance in patients undergoing primary isolated coronary artery bypass. PMID- 12116816 TI - [Life style and adherence to the recommended treatments after cardiac transplantation]. AB - BACKGROUND: All the previous studies showed an increase in survival after cardiac transplantation. Nevertheless, a certain number of patients persist, in the short and long-term period, in leading an incorrect lifestyle. Owing to this high-risk behavioral profile clinical, psychological and social problems could arise. METHODS: We have analyzed the quality of life and adherence to the recommended treatments in 107 patients at least 6 months (range 6-132 months) after they had been submitted to heart transplant and returned to their preoperative social and family environments. RESULTS: Data analysis has shown, in these patients, the return to their previous high-risk lifestyle habits: 18.4% of them did not adhere to the suggested diet, 13.2% did not perform physical activity, and 36.7% of the smokers took up the habit again after the transplant. Perceptions of emotional distress were still there: anxiety in 10.3% of cases and depression in 13.1%. An unsatisfactory sexuality was reported by 27.1% and only 27% returned to their previous job. Their outlook towards their self-efficacy seems to be predictive of the quality of life as perceived by the patient after the heart transplant: the better their outlook towards their self-efficacy, the better the quality of life. CONCLUSIONS: These data show that following heart transplant, before implementing postoperative preparation and rehabilitation programs one must carefully study those problem areas that unfortunately still exist. PMID- 12116817 TI - [Association of non-compacted myocardium and patent ductus arteriosus in a young asymptomatic male patient: findings in screening for contact sport eligibility]. AB - Spongy or noncompacted myocardium is a rare congenital cardiomyopathy characterized by parietal myocardial structural alterations secondary to intrauterine arrest of myocardial fiber compaction. It can be associated with other congenital cardiovascular malformations or manifest as an isolated disease. It is characterized by a wide variety of clinical presentations that range from complete absence of symptoms to thromboembolic manifestations, ventricular arrhythmias and congestive heart failure; the short- and mid-term prognosis is quite severe. We here describe a casual finding of an advanced form of noncompacted myocardium associated with patent ductus arteriosus in a young asymptomatic athlete during a routine visit for agonistic sports eligibility. Although the patient had a normal functional capacity, transthoracic echocardiography showed left ventricular dilation with a moderately depressed contractile function and the presence of numerous prominent trabeculae and deep intertrabecular recesses which communicated with the left ventricular cavity. These findings which were confirmed by further echocardiographic examinations, and the absence of other cardiovascular disorders associated with this disease, led us to the diagnosis of noncompacted myocardium. The patient was not allowed to perform sports activities and underwent percutaneous closure of the patient ductus arteriosus. He was asymptomatic and because in the recent literature there are no data regarding pharmacological therapy in these patients, the subject was submitted to routine clinical follow-up evaluation with no medical therapy. PMID- 12116819 TI - ["Fast medicine" and "slow medicine"]. PMID- 12116818 TI - [Non-invasive coronary angiography by multislice computed tomography: a new diagnostic method?]. AB - A significant improvement in the noninvasive evaluation of coronary anatomy has been obtained after the introduction of the new high-speed multislice computed tomography systems. Images are reconstructed using retrospective ECG-gated protocol along with contrast analysis and three-dimensional display algorithms. The 8 detectors and the reduced tube rotation time of last-generation scanners allow the coverage of the entire heart during a single breath-hold following an intravenous bolus of 120 ml of nonionic contrast. Faster computer software offers submillimeter resolution reconstructions and increased post-processing capabilities, such as quantitative angiography, virtual angioscopy, and calcium score evaluation. At least in this phase of technical development the visualization of side branches of coronary vessels seems to advantage multislice computed tomography with respect to nuclear magnetic resonance and electron-beam computed tomography in the challenge for clinical noninvasive evaluation of coronary microcirculation. PMID- 12116821 TI - Regulation of endothelial function in coronary microcirculation by HMG-CoA reductase drugs. PMID- 12116820 TI - Coated stents: a novel approach to prevent in-stent restenosis. PMID- 12116823 TI - Robotic-assisted off-pump coronary surgery. PMID- 12116824 TI - The role of synergy between cardiology and cardiac surgery in the choice of the treatment of dilated cardiomyopathy. PMID- 12116822 TI - Pathophysiology of ischemia-reperfusion injury: experimental data. PMID- 12116825 TI - Ischemic heart disease: the platelet paradox. PMID- 12116827 TI - [Tendon transfer for radial paralysis]. PMID- 12116826 TI - Modern strategies to prevent coronary restenosis. PMID- 12116828 TI - [Osteosynthesis for scaphoid fractures through a dorsal approach: one or two Herbert screws?]. AB - The aim of this work is to show the merits of retrograde scaphoid fixation through a dorsal approach using two Herbert screws in both recent and old fractures, without the need for bone grafting. Fifty three scaphoid fractures were operated on by the author between 1992 and 2000, using a dorsal approach (mean follow-up 4 years). The time between trauma and surgery, the type of fracture, the mode of bone fixation, the time of immobilisation, and the clinical and radiological results were all been evaluated. All fractures but one united, in a mean time of 75 days. The time needed to achieve bone union increases with the delay in treatment, as does the time for clinical healing which may be as long as one year. The technique of retrograde scaphoid fixation with two Herbert screws is described, evaluated from the biomechanical point of view, and compared to other methods of fixation. In contrast to a single screw, two Herbert screws give complete stability of fixation, and guarantee a good clinical result provided the correct indications have been selected. This technique has advantages as an alternative procedure, especially in fractures that present after a significant delay, and enables one to avoid having to bone graft. PMID- 12116829 TI - [Elastic stable intramedullary nailing of hand bones. ECMES even with the hand]. AB - AIM: Apply the principles of elastic intra-medullary nailing developed by Metaizeau for children, to bone fixation of bones of the hand. METHOD: Retrospective review of 43 patients treated between 1995 and 1999 for fracture of one or more bones of the hand. Thirty five concerned the right hand, 11 the left; 54 involved metacarpal fractures and 2 fractures of the phalanx. Only those patients whose lesions were considered mended were taken into account in this study. RESULTS: In 32 cases, bone fixation was completed with the use of a single pre-stressed pin, in 24 cases 2 prestressed pins forming a double arc were used. The delay in removal of material varied from between 35 days to 4 1/2 months. In three cases it required a dynamic splint treatment. Functional re-education was prescribed for two patients. Eight patients were lost for follow up. Fourty eight were followed with complete recovery of joint range of motion. We deplored one necrosis of the skin due to the contusion at the time of the initial trauma, one delayed healing after an animal bite required keeping the material in place for 4 months, one case of infection; in another case insufficient stabilization by the ECMES resulted in a 10 degrees mal-union of the metacarpal diaphysis. Three times the site of a fracture had to be opened in order to perform bone fixation. CONCLUSION: The ECMES appears perfectly suitable for bone fixation of the diaphysis of bones of the hand and certain metaphyseal and epiphyseal fractures. Small incisions help to avoid unbecoming scars and adhesions and foster a rapid recovery of function. PMID- 12116830 TI - Single injection digital block: comparison between three techniques. AB - INTRODUCTION: Regional anesthesia of a single finger is commonly achieved by the traditional ring block. The major drawback of this technique is the need for at least two painful injections in the digit. Single injection techniques have been described. A comparison of their results could help health professionals select the most appropriate technique. MATERIAL AND METHODS: A prospective randomized study was designed to compare three techniques in term of patient tolerance, distribution of anesthesia and efficiency: the modified transthecal digital block, the subcutaneous digital block and a combination of the two. Digits were randomized in three groups (n = 30). Blocks were performed by a single investigator. A visual analogic scale was used to evaluate pain associated with the injection. Prick-testing was used to evaluate anesthesia at the volar and dorsal aspects of the phalanxes. Statistical analysis of the results was performed. RESULTS: All techniques allowed surgery to be performed without complementary injection most of the time (25/30). The dorsum of the proximal phalanx, however, was unpredictably included in the anesthetized territory. The highest rate of full digital block was achieved with the combined technique. DISCUSSION: The least invasive of equally effective techniques should be considered as the first choice. The subcutaneous single injection digital block is safe, efficient and easy to perform. It allows treatment of all conditions on the volar aspect of the finger and on the dorsal aspect of the distal and middle phalanxes. For surgery on the dorsal aspect of the proximal phalanx, a supplementary dorsal block should be used. PMID- 12116831 TI - [The square pronator: an "aspirin" strip in antalgic surgery for painful neuromas of the wrist]. AB - From June 96 to January 2001, 25 patients have been operated on for painful neuromas localised at the wrist. They were between 10 and 52 years of age. These neuromas were located in at the median nerve 17 cases, at the ulnar nerve in seven cases and at the radial nerve in one case. In all cases the pain was not spontaneous but triggered off by external stimulus. All these neuromas were in continuity except 1 case at the median nerve where a total severing was present. After external neurolysis, the quadratus pronatus muscle is wrapped around the neuromas in 24 cases and around epineurial suture in one case. No internal neurolysis was done. This flap was vascularized by the anterior interosseous artery in 23 cases and by the posterior interosseous artery in two cases of distal ulnar nerve neuroma. All patients have been improved by using this method. In 21 cases (84%) (17 median, 4 ulnar nerves) the pain has completely disappeared and in four cases (3 ulnar, 1 radial), the pain has considerably decreased but without total disappearance. In the cases of median nerve neuromas the dysaesthesia has greatly decreased. No complications and no sequelae on the donor site are reported. PMID- 12116832 TI - Clinical results and thoughts on sensory nerve repair by autologous vein graft in emergency hand reconstruction. AB - Lesions of the digital and other sensory nerves in the hand are common. Based on experimental studies on vein graft as a support for peripheral nerve regeneration, the Authors have been using a simple vein graft to bridge sensory nerve gaps when treating acute hand injuries. This is a retrospective study on the results of 22 sensory nerves repaired using vein grafts in cases in which primary suture was not feasible, in emergency hand reconstruction. Patients were informed that a secondary nerve graft could possibly be necessary in the future. Patients were reviewed by two independent observers at least one year after repair and evaluated using the Highest scale as modified by MacKinnon & Dellon. Evaluation chart included influence of repair on rehabilitation program and presence of painful neuromas and scars as well as patient satisfaction. Results were classified according to Sakellarides and 20/22 were classified as very good or good. Cases classified as poor were satisfied and no secondary nerve grafting has been carried out. Rehabilitation of the associated lesions (tendon lacerations or bone and soft tissue damage) was not influenced by the nerve repair and no painful neuroma was reported in the series. In conclusion, since the literature shows unsatisfactory results in repair of digital nerves with nerve grafts, since it's been demonstrated that an unrepaired sensory nerve leads to painful scar and painful neuroma and since we are reluctant to use nerve grafts in emergency procedures, we recommend this simple method because it is easy, low-cost and effective. PMID- 12116833 TI - [Rupture of the radial collateral ligament of the fifth metacarpopharyngeal joint. A case report with Stener effect]. AB - We report a case of complete rupture of the radial collateral ligament of the fifth metacarpophalangeal joint. At surgical exploration, a Stener like lesion was identified in which the ruptured and of the ligament was trapped by the proximal portion of the extensor hood and sagittal band; thus reattachment to its original site (the base of proximal phalanx) was performed. Postoperative care consisted of protected active motion exercises which were begun immediately. An early functional recovery was obtained with full range of motion, normal joint stability and complete pain relief. The purpose of this study was to present an uncommon injury and to analyse the literature. PMID- 12116834 TI - Cavernous haemangioma in the hand mimicking subacute tenosynovitis. AB - A case of cavernous haemangioma arising from the superficial palmar arch is described. The initial symptoms were those of a subacute tenosynovitis. Surgical exploration showed that the tumor was not affecting the flexor tendons. It was completely resected and the patient had full recovery of hand function. PMID- 12116835 TI - [Cancer of the prostate]. PMID- 12116837 TI - [Localized cancer of the prostate. What to tell the patient?]. AB - Answering the question from patients with adenocarcinoma of the prostate is always difficult; choice of radiation therapy (versus prostatectomy), indications of brachytherapy frequency and severity of acute and late effects as rate of survival, local control, are among the most frequent inquiries. A part of the answers are not evident of due lack, consensus and the physicians need to speak honestly and give the most appropriate responses without improving anxiety and fears of the patient. PMID- 12116836 TI - [What systematic work-up should be required for the patient with localized adenocarcinoma of the prostate?]. AB - The systematic work-up for patient with clinically localised prostate cancer must allow a reconciliation of all the necessary elements to propose one or several therapeutical options. The evaluation must take into account the life's expectancy, and must precise the cancer progression and prognostic factors. The life expectancy depends out age, performance status and comorbidities. Digital rectal exam, PSA and Gleason score analysis are systematic. These tests allow the specialist to decide whether or not to complete the progression work-up by bone scan and pelvic CT. This first work-up should define if a patient will benefit from some curative local treatment. In case of palliative treatment or deferred treatment, the work-up highlights the specialist to decide the medical supervision. Some exams, systematic or not, are very specific from each therapeutic modality considered; they allow to precise individually the efficacy and toxicity prognostic factors for each therapeutic strategy. The final therapeutic decision is based at first on this objective data even if they are probabilistic, and finally on informed patients preferences. PMID- 12116838 TI - [Prostate cancer: has local radiation treatment had an impact on survival?]. AB - Local control is an important goal in the treatment of prostate cancer. Firstly, it avoids the morbidity due to locoregional evolution (urethral obstruction, vascular compression, rectal or vesical involvement). Moreover, local control of the disease may decrease the mortality due to metastases disseminated from local relapse. Local control evaluation remains difficult: neither rectal examination nor imaging or prostate biopsies have an absolute value in diagnostic of local relapse. PSA increase does not permit to differentiate local from distant relapses. Recent developments in radiotherapy techniques allow dose escalation without major toxicity. Retrospective studies and one randomized study have shown that an increase from 70 to 80 Gy or more, improve biological relapse-free survival. In one randomized study comparing 70 to 78 Gy, the biochemical disease free survival was improved from 69 to 79% at five years. Such an improvement can only be explained by an improvement of local control. The benefit in term of overall survival is not yet demonstrated and needs a longer follow-up and other studies. Another approach to improve local control is the association of a local radiotherapy with hormonal adjuvant therapy. Four randomized studies have been published for locally advanced prostate cancer. These studies have all demonstrated an improvement of local control, and a decrease of metastatic risk. The benefit in term of overall survival, observed in one of this trial, may be explained by the improvement of either local or distant control or both. Such therapeutic progress, associated with the development of prostate cancer screening should lead to a decrease of prostate cancer mortality for the next ten years. PMID- 12116839 TI - [Conformal radiotherapy in prostate cancer: for whom and how?]. AB - External radiotherapy is one of the modalities used to cure localized prostate carcinoma. Most of localized prostate carcinomas, specially those of the intermediate prognostic group, may benefit from escalated dose above 70 Gy at least as regard biochemical and clinical relapse free survival. 3D-CRT allows a reduction of the dose received by organs at risk and an increase of prostate dose over 70 Gy. It is on the way to become a standard. Intensity modulated radiation therapy increases dose homogeneity and reduces rectal dose. These methods necessitate rigorous procedures in reproducibility, delineation of volumes, dosimetry, daily treatment. They need also technological and human means. It is clear that localized prostate cancer is a good example for evaluation of these new radiotherapy modalities. PMID- 12116840 TI - [Indications for curietherapy of the prostate using permanent implants]. AB - In the last decade, brachytherapy emerged as a particularly appealing new way ot treating localized prostate cancer. Recently published 10-12 years biochemical control results appear to be superimposable to the best percentages achieved by surgery or conformal radiotherapy, with a small percentage of complications. This applied to severely patients. Only patients with T1/T2, PSA < 10 ng/mL, and Gleason score < 7 should be proposed such a treatment. The potential benefit of exploring patients with a endorectal coil MRI is being evaluated. The number of positive biopsies is also a parameter which should probably be considered in the therapeutic choice. Moreover, a prostate volume > 60 g, hip mobility limitations, a urinary obstructive syndrome and previous transurethral resection lead to difficulties in technical implantation and therefore must be taken into account when discussing brachytherapy. In conclusion, for adequately selected patients, brachytherapy offers a particularly applied alternative to surgery and external radiotherapy, with satisfactory long term biochemical control rates and limited complications. PMID- 12116841 TI - [Clinical and biological surveillance after radiotherapy for localized prostate cancer]. AB - Serum PSA is an excellent marker of disease status after external beam radiotherapy or brachytherapy for patients with prostate carcinoma. A low PSA nadir < or = 1 even < or = 0.5 ng/mL has been shown to be as a surrogate end point for disease control. Three successive increases of this marker after achieving the nadir defines recurrence as recommended by the American Society for Therapeutic Radiology and Oncology. The biochemical relapse or PSA failure after treatment precedes clinical disease relapse by several months. PSA profile or kinetics may have implications for patterns of failure and prognosis. Prostate post-radiotherapy biopsies should not be part of routine follow-up as its interpretation is frequently problematic. Other exams should not be performed unless clinical symptoms are present. Post-radiotherapy relapse treatment has generally no curative intent. PMID- 12116843 TI - [Hormono-radiotherapy of non-metastatic prostate cancers]. AB - Patients presenting with non-metastatic cancer of prostate have a high probability of relapse if they are treated by either surgery alone or irradiation alone, when poor prognosis factors are present. Clinical stage (> or = T3a), Gleason score, and PSA level (> 20 ng/mL) are the more significant factors. It is likely that many patients can draw benefit of combined androgenic suppression and radiotherapy. However, despite results of European and American trials published the last decade, a number of questions remain without a clear response, especially on the modalities of treatment according to the characteristics of the disease. PMID- 12116842 TI - [Post-prostatectomy radiotherapy: for which patients and when?]. AB - Biochemical relapse after radical prostatectomy is not exceptional, ranging from 10 to 40% in the literature. To prevent this biochemical failure, adjuvant radiotherapy was proposed to patients with a high risk of relapse. No phase III trial has actually validated this attitude. Best indications for adjuvant irradiation seem to be patients with an extensive extracapsular extension or multiple positive margins. Historical comparisons seems to confer, in these case, a benefit in biochemical control for adjuvant irradiation versus observation. Others authors prefer immediate post-operative irradiation, a delayed treatment, when biochemical relapse has occurred. This attitude has spared some patients irradiation useless. This salvage irradiation lowered the PSA level in 40 to 70% of the cases, but long-term efficiency is obtained only in the case of a low value of the PSA before irradiation. Delayed radiotherapy is, therefore, justified only if a close follow-up is performed, with repeated dosage of PSA. Whatever the case, it is important to differentiate between local and distant relapse: patients with positive nodes at the time of surgery are most likely at risk of distant relapse. It seems that patients with seminal vesicles involvement are also at high risk for distant relapse, but this must be confirmed. PMID- 12116844 TI - [Does one have a sexual life after prostate cancer treatment? ]. AB - The precise assessment of sexual dysfunction after treatment of prostatic cancer cannot be avoided in 2002. These iatrogenic complications may significantly alter the quality of life of the patients. In addition, sexual toxicity is progressively becoming a cardinal parameter for the treatment choice, both for the patient and the physician. Significant efforts allowed to reduce sexual toxicity after therapy in the recent years. As an example, nerve-sparing surgical techniques have been proposed, whenever reasonable. However, in spite of these surgical advances, data suggest that overall, the new irradiation techniques (conformal radiotherapy and brachytherapy) are responsible for less alteration of sexual life than surgery. Another potential advantage is that sildenafil (Viagra) is able to restore potency in a majority of cases after radiotherapy, while it is usually poorly effective after surgery. PMID- 12116846 TI - The human parvovirus B19 and its interactions in vivo. PMID- 12116845 TI - [Metabolic radiotherapy: what role will it have in 2001?]. AB - Metabolic radiotherapy is a new therapy for management of bone pain in patients with bone metastatic prostate carcinoma. Strontium-89 and Samarium-153 concentrate in bone metastases and radiate them. A pain decrease is obtained in 60-70% of cases. Side effects are a significant hematological depression without great clinical consequences if good therapeutic indications are respected. Our multidisciplinary experience of these radionuclides in 54 performed treatments shows a rate of good responders of 66% with a rate of excellent results (total decrease of pain) in 47%. The therapeutic effectiveness is correlated with pain intensity measured by Visual Analogic Scale (VAS) and equivalent dose of morphine. Radionuclide therapy should be applied to patients as early as possible after establishment of bone metastases. PMID- 12116847 TI - Current molecular epidemiology and human parvovirus B19 infection. AB - Viruses evolve gradually through replication. Therefore, isolates of a virus species can have different genome sequences, albeit slightly, if isolates are epidemiologically unrelated. The difference in virus genome involves difference in virus functions and clinical manifestations of virus infection. Molecular epidemiology of virus infection is a relatively new field directed at infection in humans but not other animals. Analyses are based on genomic differences between virus strains with advances in methodology related to DNA analyses, progress is being made. Classification of virus strains, tracing of transmission of a strain, analyses of outbreaks (including nosocomial infection), and analyses of pathogenesis of virus infection in humans (a natural host) are given attention in molecular epidemiological studies. Human parvovirus B19 is a common human pathogen associated with a wide variety of diseases, including erythema infectiosum, aplastic crisis, hydrops fetalis, and arthritis. B19 is not propagatable in conventional cell lines, hence, molecular cloning of B19 DNA directly from clinical materials has to be done. Events concerning B19 infection were analyzed based on the concept of molecular epidemiology and studies proved to be productive to better understand the pathogenesis of B19 infection. PMID- 12116848 TI - Recombinant human parvovirus B19 vectors. AB - In an attempt to exploit the remarkable tissue-tropism of the human parvovirus B19 to target human hematopoietic cells of the erythroid lineage, recombinant human adeno-associated virus 2 genomes were encapsidated in parvovirus B19 capsids. Although efficient transduction of primary human hematopoietic cells in the erythroid lineage occurred, a low-level of transgene expression in non erythroid cells was also detected. These studies suggest that cell surface expression of P antigen, the primary receptor for parvovirus B19, is necessary but not sufficient for parvovirus B19 vector-mediated transduction of human hematopoietic cells. These studies also suggest the existence of a putative cell surface co-receptor, which is required for successful infection of human hematopoietic cells by parvovirus B19. PMID- 12116849 TI - Persistence of human parvovirus B19 in human tissues. AB - Human parvovirus B19 infection causes various clinical symptoms, such as rash, arthropathy, anemias and fetal death, but it can also remain asymptomatic. The arthropathies and anemias can become chronic for several years, not infrequently resembling autoimmune syndromes. B19 replicates only in red blood cell precursors of bone marrow or fetal liver, resulting in high-titred short-lived viremia, but viral DNA is detectable also in cells of several other types. Recently B19 DNA has been found, by very sensitive amplification tests, in certain tissues not only of symptomatic but also of healthy individuals for several years or decades after B19 infection. The mere presence of B19 DNA in these tissues of a symptomatic patient (e.g. joints in chronic arthritis or skin in dermatomyositis) thereby does not prove that the present disease is caused by B19. The diagnosis has to be verified by other innovative means. How and why viral DNA persists in the tissues of healthy individuals is under investigation. PMID- 12116850 TI - T lymphocyte responses against human parvovirus B19: small virus, big response. AB - Parvovirus B19 elicits both humoral and cellular immune responses. Recently some advances have been made in determining the frequencies, peptide targets and function of virus-specific CD8+ T lymphocyte responses. A single HLA B35 restricted epitope derived from the NS1 protein has been studied so far, but others clearly exist. Surprisingly large, persistent responses have been detected in healthy seropositive individuals, using interferon-gamma ELISpot assays and HLA class I peptide tetramers. Similar techniques are available for exploration of the CD4+ T cell epitopes, although less detail is currently available. Mapping of cellular immune responses against the entire B19 genome (the parvovirus "immunome") is now possible and if similarly large populations are found consistently, this could yield important insight into normal immunological control and abnormalities in B19-related disease. PMID- 12116851 TI - Antibody responses in parvovirus B19 infected patients. AB - Parvovirus B19 is the causative agent of erythema infectiosum. In addition, the infection may be associated with other disease manifestations: anemia and aplastic crisis, thrombo- or granulocytopenies; spontaneous abortion or hydrops fetalis in pregnant women; acute and chronic arthritis in adults and children, myocarditis and hepatitis. Both acute and persistent courses of B19-infections have been reported. All patients develop IgG against the capsid proteins VP1 and VP2, the majority of virus neutralizing antibodies that offer life-long protection against reinfections are directed against the VP1-unique region. IgM is mainly directed against VP2-specific epitopes. These antibodies may be present for only a rather short period of two to ten weeks after acute infection. IgG antibodies against the nonstructural protein NS1 are preferentially found in patients which are unable to eliminate the virus and develop persisting viremia or virus persistence in distinct organs, e.g. synovial fluid, liver, bone marrow. PMID- 12116852 TI - Diagnostic procedures in B19 infection. AB - In immunologic normal hosts, both children and adults, B19 can cause acute, generally self-limiting diseases. The infection leads to a viremia that can be present, at high titre, for about one week, then the onset of a specific immune response controls the infection. B19 infection in pregnancy can be associated with non-immunologic foetal hydrops or foetal death. In immunocompromised hosts, B19 can persist over several months and sometimes years. Persistent or recurrent B19 infections can be associated with chronic clinical manifestations or with transient clinical syndromes, generally related to the recrudescence of viral replication. Since the infection has been associated with a wide variety of clinical manifestations and some clinical features of B19 infection, such as anemia, artropathy and rash, can be common to other pathogens, a laboratory diagnosis of B19 infection is required. A diagnostic protocol must consider both the type of pathology and the type of patient. In immunocompetent individuals serological and virological testing is complementary, while in immunocompromised patients viral detection is the diagnosis of choice. Viral detection methods are generally based, nowadays, on the direct detection of B19 genome in clinical specimens. B19 DNA is mainly detected by hybridizations assays and by the most sensitive PCR assays. Serological diagnosis of B19 infection is generally achieved by detection of IgM and IgG antibodies to the B19 structural proteins VP1 and VP2. IgM detection is most often performed by capture assays, both in EIA and RIA formats, IgG are mainly detected by indirect EIA and immunofluorescence tests. PMID- 12116854 TI - Parvovirus B19 and erythroid cells. AB - Parvovirus B19 is a human erythrovirus, i.e. which induces the death of erythroid progenitors. In such cells, until now only ubiquitous transcription factors have been described to regulate promoter driven gene expression. Their possible interactions with erythroid specific transcription factors merit further investigations. Effectively, the high level of replication of B19 in erythroid cells is not well understood. In addition to apoptosis, necrosis or inhibition of cell growth, the death of B19 infected erythroid progenitors has been never clearly reported as the result of immunological attack: this mecanism will merit further investigations. The interactions with other cell types in vitro remain at present not well defined but many obstacles have been mentioned which counteract B19 expression. PMID- 12116853 TI - Parvoviruses and blood products. AB - Erythrovirus B19 (B19), a small isocahedral, non-enveloped virus (18-26 nm), is a ubiquitous infection agent in industrialised countries. Depending on the infected host, B19 has a wide range of disease manifestations from asymptomatic (the majority) to severe, including persistent infection. The risk of B19 transmission by blood products is enhanced by a high virus titre in the infected donor, by pooling of a large number of donations, and by the virus's resistance to effective inactivation methods such as heat and solvent-detergent treatments. B19 DNA has been detected in single donations, in manufacture plasma pools and in plasma derivatives (clotting factors, albumin, antithrombin III and immunoglobulins) produced by different processes. B19 transmission is mostly found in patients treated with clotting factors, as shown by a higher seroprevalence in treated haemophiliacs, by the presence of B19 DNA, and by active seroconversion. Chronic B19 infection can successfully be treated with polyvalent intravenous immunoglobulins. The key role of neutralising anti-B19 antibodies and of the virus titre has been demonstrated by B19 transmission after infusion of several B19-positive plasma batches treated with solvent-detergent. Two strategies can be followed to reduce the B19 risk: (1) reducing the viral load in the manufacture plasma pool by discarding B19-DNA-positive donations; (2) developing new strong virus inactivation methods. The physico-resistant properties of B19 make it a good model for new emergent viruses capable of infecting blood products. PMID- 12116855 TI - Onchocerciasis in West Africa after 2002: a challenge to take up. AB - Initially planned for a 20 year life time, the Onchocerciasis Control Programme in West Africa (OCP) will have finally continued its activities for nearly three decades (vector control alone from 1975 to 1989, then vector control and/or therapeutic treatment until 2002). Although onchocerciasis is no longer a problem of public health importance nor an obstacle to socio-economic development in the OCP area, the control of this filariasis is not over because OCP never aimed at eradication, neither of the parasite (Onchocerca volvulus), nor of its vector (Simulium damnosum s.l.). In 2003, the eleven Participating countries of OCP will take over the responsibility of carrying out the residual activities of monitoring and the control of this disease. This mission is of great importance because any recrudescence of the transmission could lead in the long run to the reappearance of the clinical signs of onchocerciasis, if not its most serious manifestations. For epidemiological and operational reasons, and given the disparity in national health policies and infrastructures, the capacities of the countries to take over the residual activities of monitoring and control of onchocerciasis are very unequal. Indeed, the interventions to be carried out are very different from one country to another and the process of integrating the residual activities into the national health systems is not taking place at the same pace. This inequality among the countries vis-a-vis the challenges to be met does not, however, prejudge the epidemiological situation after 2002 whose evolution will also depend on the effectiveness of the provisions made before that date by OCP, then after 2002, by the Regional Office for Africa of the World Health Organization which is currently setting up a sub-regional multidisease surveillance centre. PMID- 12116856 TI - Unusual snail species involved in the transmission of Fasciola hepatica in watercress beds in central France. AB - Four freshwater pulmonate species (Lymnaea ovata, L. stagnalis, Physa acuta, Planorbis leucostoma) were living in several watercress beds known for their relationships with human cases of fasciolosis, whereas L. truncatula was never found. The aims of these studies were to determine the prevalence of natural infections with Fasciola hepatica in snails and to verify if these species might ensure the full larval development of this trematode (with cercarial shedding) when they were experimentally subjected to F. hepatica only, or to co-infections with an other trematode species. Investigations were so carried out in six snail populations living in watercress beds (including three for P. acuta) and in four others originating from three brooks or a pond (as controls). Snails naturally infected with F. hepatica were found in two watercress beds inhabited by L. ovata (prevalence of infection: 1.4%) and P. leucostoma (0.1%), respectively. The L. ovata from the watercress bed could be infected at a higher size than those from the control population and the prevalence of this infection was greater in the bed population. Similar findings were noted for L. stagnalis. Despite single or dual infections, the results obtained with the four populations of P. acuta were unsuccessful. In contrast, the co-infections of young P. leucostoma with Paramphistomum daubneyi and F. hepatica resulted in the shedding of some F. hepatica cercariae. According to the authors, the occurrence of fasciolosis in these watercress beds would be the consequence of frequent natural encounters between parasite and snails (L. ovata, L. stagnalis), or of co-infections with P. daubneyi and F. hepatica (P. leucostoma). In watercress beds only colonized by P. acuta, a lymnaeid species would have ensured the larval development of F. hepatica but it would have been eliminated by P. acuta, as this last species was known to be invasive and could colonize open drainage ditches on siliceous soil. PMID- 12116857 TI - Anthobothrium galeorhini n. sp. (Eucestoda: Tetraphyllidea) a parasite of Galeorhinus galeus (Triakidae) from the Argentine coast. AB - Anthobothrium galeorhini n. sp. (Eucestoda: Phyllobothriidae) collected from the spiral intestine of Galeorhinus galeus (Linnaeus, 1758) at the Puerto Madryn (Atlantic coastal region) is described. The new species differs from A. cornucopia as described by Euzet (1959) by the size of body, bothridia, gravid proglottids and eggs; by the neck length and by the different number of testes and proglottids. The size of bothridia and the gravid proglottids, the cirrus length and the number of testes allow differentiating the species here proposed from A. laciniatum Linton, 1890 as described by Euzel (1959). The body and cirrus sac size, the bothridia shape and the number of testes separate the species here described from A. parvum Stossich, 1895 as described by Yamaguti (1952). The anatomic differences between the cestode described by Arandas Rego (1977) and located laxonomycally under the name Anthobothrium laciniatum are listed and the systematic position of the cestode is discussed. Species parasitic of skates listed in scientific literature must be taxonomically reaccommodated, since their characteristics are not consistent with the diagnosis of Anthobothrium. PMID- 12116858 TI - Trichostrongylina (Nematoda) from Malagasy muridae. II--Description of two new species of Heligmonina (Heligmonellidae) in Nesomys rufus and Eliurus tanala. AB - Two new species of Heligmonellidae from Madagascar are described, Heligmonina madagascariensis n. sp. in Nesomys rufus and Heligmonina tanala n. sp. in Eliurus tanala. Both species belong to the Heligmonina species with a pattern of type 1-3 1 for the right lobe of the caudal bursa and 1-4 for the left lobe. In H. madagascariensis, H. dupuisi (Desset, 1964) and H. praomyos Baylis, 1928, left ray 6 arises before ray 3 from the common trunk to rays 3 to 6 while in H. tanala and the other species, it arises at the same level. H. madagascariensis is differentiated from H. dupuisi and H. praomyos by the symmetry of the branches of the dorsal ray. H. tanala is differentiated from H. malacomys Sakka & Durette Desset, 1988, the closely related species by a different pattern of the cuticular ridges at mid-body, by the sharpness of the tips of the spicules and by the ratio of the length of the spicules on the length of the body (6.9, 8.8% versus 25 27.8%). Heligmonina chippauxi (Desset, 1964) a parasite of Oenomys hypoxanthus from the Republic of Central Africa is considered a valid species. PMID- 12116859 TI - The effect of substrate on survival and development of two species of desert fleas (Siphonaptera: Pulicidae). AB - Fleas Xenopsylla conformis mycerini and Xenopsylla ramesis replace each other on the same rodent host (Meriones crassus) in two habitats that differ in substrate texture (sand and loess-like sediments, respectively). We hypothesized that the substrate is an important factor determining flea distribution and studied survival of larvae, pupae and newly emerged adults as well as the rate of pre imaginal development of these flea species in sand and loess rearing medium (= substrate). Texture of rearing medium did not affect survival and development rate of eggs in either X. c. mycerini or X. ramesis. Larval survival and the rate of development were both affected by the factor of substrate. Survival of X. c. mycerini larvae was significantly higher in sand than in loess substrate, whereas survival of X. ramesis larvae did not differ in different substrates. Larvae of both species developed faster in sand substrate than in loess substrate. Maximal survival time of X. c. mycerini larvae that died before pupation did not depend on substrate, whereas X. ramesis larvae survived significantly longer in loess than in sand substrate. Most pupae of both species survived successfully on both substrates, but the duration of pupal stages in sand substrate was longer than that in loess substrate in both species. Newly emerged adults of both species survived similar time in both sand and loess substrate. Irrespective of substrate, adult X. c. mycerini survived for a shorter time than did adult X. ramesis. No between-sex within-species differences in survival time of newly emerged adults in sand versus loess substrate were found in X. c. mycerini. Survival time of males and females of X. ramesis differed in sand substrate but not in loess substrate. PMID- 12116860 TI - Schistosoma mansoni: kinetics of glomerulonephritis in Mongolian gerbils and its correlation with intensity and duration of infection. AB - The frequent occurrence of glomerular lesions in schistosomiasis patients has been reported, although appropriate animal models for the study of schistosomal glomerulonephritis have not been developed. To analyze the relationship between glomerulonephritis and Schistosoma mansoni infection, gerbils, Meriones unguiculatus, were infected with different number of cercariae and sacrificed at different weeks of post infection. Fifty cercariae were the optimum dose to produce the disease, glomerulonephritis, without early death of the animal. Infected gerbils showed heterogeneous types of glomerular lesions with increased serum creatinine level. Immune complex deposition was not detected at glomeruli of infected gerbils even by means of immunuofluorescence and also by transmission electron microscopy. However, infiltration of mononuclear cells in and around some of the altered glomeruli was observed. Immunohistochemical staining, using monoclonal antibody (HUSM-M.g. 15) specific to gerbil's T-cells, revealed significant infiltration of T-cells. These findings suggest that T-cells might be involved in the development of glomerulonephritis. Gerbil could be a useful model to clarify the role of T-cells in the development of glomerulonephritis of schistosomiasis. PMID- 12116861 TI - Comparison of cholinesterase activities in the excretion-secretion products of Trichinella pseudospiralis and Trichinella spiralis muscle larvae. AB - The presence of cholinesterases (ChE) is reported in T. pseudospiralis excretion secretion products (ESP) by spectrophotometric method, using acetylthiocholine (ATCI) and butyrilthiocholine (BTCI) as substrates. By inhibition assays, we found that T. pseudospiralis release both acetyl- and butiryl-cholinesterases (AchE and BchE, respectively). The sedimentation coefficientes of these enzymes were determined by sucrose density gradient. We studied the in vivo ChE secretion by immunoblot assays using AchE from Electrophorus (electric eel) and sera from normal or infected mice with T. pseudospiralis or T. spiralis. The presence of anti-AchE antibodies was only demonstrated in the sera from T. pseudospiralis infected mice. Moreover the in vivo secretion was corroborated by the high difference determinate between the ChE activity of the immuno complexes from T. pseudospiralis infected sera and the immunocomplexes from T. spiralis infected sera as well as normal sera. Finally, we analyzed the effect of the organophosphate Neguvon (metrifonate) on the ChE activity from the T. pseudospiralis ESP. The drug inhibits in part this activity. Moreover Neguvon (metrifonate) showed a high activity against the T. pseudospiralis viability. PMID- 12116862 TI - Ultrastructural study of the development of Sarcocystis singaporensis sarcocysts in the muscles of its rat host. AB - Laboratory rats fed sporocysts of Sarcocystis singaporensis (Zaman & Colley, 1975) Zaman & Colley, 1976 originating from Singapore were euthanized 22, 23, 33 and 80 days later. Sporocysts were extracted from feces of either naturally or laboratory-infected Python reticulatus. Electron microscopically examined longue and esophageal muscles yielded images of successive developing stages of sarcocysts. The primary wall evolved from a continuous thin layer into folds and later, into villar protrusions. At all stages the wall was interrupted by pinocytotic-like indentations. Young sarcocysts contained only metrocytes, they divided by endodyogeny into daughter metrocytes. The first bradyzoites appeared only 33 d.p.i. Sarcocysts by 80 d.p.i. were enclosed in a fully differentiated primary wall and contained almost entirely bradyzoites. PMID- 12116863 TI - Intestinal parasitoses among Wayampi Indians from French Guiana. AB - Intestinal parasitism and its epidemiological characteristics were studied in an isolated Amerindian population from Upper Oyapock (French Guiana) that has retained its traditional social and cultural specificities. This population consisted of 138 Wayampi Indians, 68 adults and 70 children (below the age of 15 years), with a sex ratio (M/F) of 0.86, spread over the four villages of the community of Trois Sauts, corresponding to more than two thirds of the population recorded as inhabiting the sector in the last census (375 inhabitants). Fecal examination combined the direct examination of fresh feces with the quantitative techniques of Kato-Katz method, Baermann and MIF staining. Overall, 92% of the subjects were found to have intestinal parasites, 85% if only direct examination of fresh stools was taken into account. Fourteen species of human parasite were identified: seven protozoa and seven helminths. We observed in particular 1) a high frequency of hookworm infection due to Necator americanus. Over 50% of subjects were affected, with a range of 25% to 75% according to the village, but with only moderate parasite loads; 2) a high level of parasitism by E. histolytica/E. dispar (17%), Stongyloides stercoralis (16%) and Hymenolepis nana (18%); 3) a lower level of parasitism by Ascaris lumbricoides and very low levels (almost absent) of Trichiuris trichiura; 4) the absence of Schistosoma and fluke eggs. With the exception of H. nana, which was more frequent in children than in adults, there was no significant difference in the level of parasitism according to sex and age. Although the Wayampi of French Guiana are French citizens and consequently have quite high incomes and ready access to clinics and medicines, intestinal parasites are far from under control in this population. A lack of fecal hygiene and the habit of walking barefoot are widespread in the unchanging Amazonian environment and contribute to this phenomenon. PMID- 12116864 TI - Alternative treatment for Anacanthorus penilabiatus (Monogenea: Dactylogyridae) infection in cultivated pacu, Piaractus mesopotamicus (Osteichthyes: Characidae) in Brazil and its haematological effects. AB - The present work, studied the effect of 0, 1,000, 1,500 and 2,000 mg of garlic powder/kg dry ration for Piaractus mesopotamicus (Osteichthyes: Characidae), weighting 73.6 +/- 39.4 g and measuring 15.0 +/- 2.7 cm, fed for a period of 15, 30 and 45 days. Fifteen days after treatment with 1,000 and 2,000 mg of garlic/kg dry ration, significant reduction of Anacanthorus penilabiatus (Monogenea: Dactylogyridae) in the gills was related. Nevertheless, the addition of garlic to the ration caused significant increase in the erythrocyte number and in the thrombocyte percentage in the circulating blood. However, a decrease in the lymphocyte percentage was also observed. After 45 days, fish fed with garlic showed significant increase in the erythrocyte number, leucocyte, haemoglobin rate, hematocrit and thrombocyte. PMID- 12116865 TI - Stichosome ultrastructure of the fish nematode Capillaria pterophylli Heinze, 1933. AB - The stichosome (posterior glandular esophagus) of Capillaria pterophylli Heinze, 1933 consists of large gland cells (stichocytes) and lumenal epithelium with cuticular lining. Both structures are enclosed in a reticulum of muscle cells. The stichocyte cytoplasm contains small cisternae of rough endoplasmic reticulum, Golgi complexes, one kind of electron dense secretory granules, mitochondria and a branching system of intracellular collecting ducts without filament bundles around them. PMID- 12116867 TI - Human dirofilariasis in Costa Rica. A report of three new cases of Dirofilaria immitis infection. PMID- 12116868 TI - Human pulmonary dirofilariasis in El Salvador. PMID- 12116866 TI - Molecular characterisation of airport malaria: four cases in France during summer 1999. AB - Four airport malaria cases have been observed in the vicinity of the Roissy Charles-de-Gaulle International Airport, Paris, France. These cases were geographically very close to each other and clustered in a short period of time during the summer of 1999. The phenotype and genotype of the Plasmodium falciparum isolates obtained from these patients were determined in order to know whether a single mosquito could have infected more than one subject. The genomic characterisation of isolates was performed using the polymorphic markers merozoite surface protein 1 (Msp 1) and merozoite surface protein 2 (Msp 2) genes, the kappa and omega repeats domains of cg2 and the dihydrofolate reductase (DHFR) genotypes. Results showed identical genotypes for isolates 1, 2 and 4 whereas the genotype of isolate 3 differed at one locus. The molecular analysis was consistent with the hypothesis that all patients could have been bitten by the same mosquito and that patient 3, may have received a different clone and an additional species. In vitro susceptibility data did not confirm or rule out this hypothesis because isolates had the same profile of susceptibility to the tested drugs. PMID- 12116869 TI - New 3-[4-(aryl)piperazin-1-yl]-1-(benzo[b]thiophen-2-yl)propane derivatives with dual action at 5-HT1A serotonin receptors and serotonin transporter as a new class of antidepressants. AB - Some benzo[b]thiophene derivatives with different substituents in positions 3 and 5 have been synthesized in order to obtain new dual antidepressant drugs. Compounds derived from 2-acetyl-3-methylbenzo[b]thiophene or 2-acetyl-3,5 dimethylbenzo[b]thiophene were prepared with two different phenylpiperazines (2 methoxy and 2-hydroxyphenylpiperazine) and evaluated for in vitro 5-HT1A receptor affinity and serotonin reuptake inhibition by radioligand assays. Compound 1-(3,5 dimethylbenzo[b]thiophen-2-yl)-3-[4-(2-methoxyphenyl)piperazin-1- yl]propan-1-ol (II.2.a) shows good values (nM) for both activities: Ki = 85 for 5-HT1A receptor and Ki = 120 for serotonin transporter. PMID- 12116870 TI - Modified berberine and protoberberines from Enantia chlorantha as potential inhibitors of Trypanosoma brucei. AB - Phytochemical study of the stem bark of Enantia chlorantha resulted in the isolation of two protoberberines 1 and 2. These alkaloids as well as commercially available berberine were modified chemically and tested in vitro against Typanosoma brucei proliferation as well as on three targeted glycolytic enzymes. The inhibitory activities observed were in the range of 20 microM (ED50 values). PMID- 12116871 TI - [Pyridazino(3,4-c)quinolines and pyridazino(4,5-c)quinolines--synthesis and investigation of lipoxygenase inhibition]. AB - The cyclic hemiketone acetal 6 reacts with hydrazine in tert-butanol to yield the 1-amino-2,3-dihydro-2-hydroxy-pyrrole 7, while in acetic acid a mixture of the 1 amino-pyrrole 8 and the 1,4-dihydropyridazine 9 is obtained. The X-ray crystal structure of 9 shows a boat conformation flattened about N-1 with respect to nifedipine. Removing the boc-group of 9 gives the tautomeric 2,5 dihydropyridazine 13. The lactams 15 and 17 and the cyclic hydroxamic acids 16 and 18, respectively, are synthesized from 13 or from its oxidation product 14 using reductive conditions. The cyclic hemiacetal 21 reacts with hydrazine in a different manner from 6. In acetic acid the 1-aminopyrrole 22 is formed, while ethanol yields the 1,4-dihydropyridazine 23. The pyridazine 24, obtained by dehydrogenation of 23, gives the lactam 25 and the hydroxamic acid 26, respectively, when the nitro-group is reduced. The dihydropyridazines 9, 11 and 23 are transformed photochemically to give the nitrosophenyl-pyridazines 19, 20 and 27. The chloro-substituted hydroxamic acid 28 is isolated after treatment of 27 with hydrochloric acid. The stability of the partially saturated pyridazine compounds is discussed on the basis of half wave potentials measured by anodic oxidation by means of differential pulse voltammetry. The tricyclic hydroxamic acids 18 and 28 show only a weak inhibition of 5-lipoxygenase (5-LOX). PMID- 12116872 TI - Atractyligenine chemistry, Part VI: Synthesis and biological activities of atractyligenine derivatives. AB - Derivatives of atractyligenine, the aglycone of atractyloside, were tested in vitro for their antimicrobial and antiproliferative activity against K-562 (human chronic myelogeneous leukemia), HL-60 (human leukemia) and MCF-7 (human breast adenocarcinoma) cell lines. The most active compound showed IC50 values in the range 0.8-6.9 microM. PMID- 12116873 TI - Synthesis, antiinflammatory and analgesic activity of new hexahydropyrimidine derivatives. AB - A number of potentially active hexahydropyrimidine derivatives of pharmaceutical interest have been synthesized. Various diSchiff's bases prepared by reacting different aromatic aldehydes with 1,3-diaminopropane were suitably reduced to give their tetrahydro derivatives which were then condensed with appropriate aldehydes to give a series of hitherto unreported hexahydropyrimidines. The resulting products were evaluated by oral route for their antiinflammatory activity. The activity of compounds 11, 23 and 4 was excellent and comparable to indomethacin. In addition to oral route of administration, the antiinflammatory activity of hexahydropyrimidine derivatives was also studied topically through transdermal gels. Compounds 11, 23, 4 and 22 produced significant inhibition in edema and showed good antiinflammatory activity comparable to diclofenac sodium gel (Relaxyl gel). All these compounds were also tested for their analgesic activity and their LD50 determined. Compounds 11, 20 and 23 showed a comparable activity with aspirin. The MTD for all the compounds was found to be > 1800 mg/kg. PMID- 12116874 TI - Synthesis and properties of N-substituted saccharin derivatives. AB - Four different routes for the synthesis of saccharin containing peptides were studied. While reactions between saccharin sodium and alpha-halogeno acids were limited to two examples, reactions of sulfobenzoic anhydride (4) or saccharin-N carboxylate (6) with amino acid esters yielded the ring opened products 5 and 7. Finally, we found, that the reaction between the benzoxathiol derivative 8 and amino acid derivatives represents a versatile route to the peptidic compounds 9 and 11. Hydrolysis and hydrogenolysis were studied, and by combination of the different routes the "saccharin tripeptides" 18 were obtained. Structures and stereochemistry were elucidated by spectroscopic and chromatographic methods. Selected compounds were tested as sweeteners or as inhibitors of elastase, but no exiting results were found. PMID- 12116875 TI - 1,3-Dibromo-5,5-dimethylhydantoin (DBH) as oxidant and precipitant for drug identification according to PH. EUR Analytical methods of pharmacopoeias with DBH in respect of environmental and economical concern, Part 15(1). AB - Elemental bromine serves as oxidant for the identification of propylthiouracil, 2 thiouracil and sulphur according to PH. EUR. 2002. Phenol is identified according to PH. EUR. 2002 with bromine water as the sparingly water-soluble 2,4,4,6 tetrabromo-2,5-cyclohexadiene-1-one. These tests can be performed better using 1,3-dibromo-5,5-dimethylhydantoin (DBH). PMID- 12116876 TI - Glass classification by linear discriminant analysis of LA-ICP-MS data. AB - The classification of pharmaceutical and common glasses (ampoules, infusion bottles, cylinders, lead crystal and bottle glass) has been performed means by of chemometric methods. For this purpose intensity data of glass examinations received by laser ablation inductively coupled plasma mass spectrometry (LA-ICP MS) were applied. A Nd:YAG laser with 10 Hz repetition rate in the Q-switch mode at its 4th harmonic (266 nm) was used to get mass spectrometric data of the glasses. 13 isotopes (= variables) were used for measurements (7Li, 11B, 23Na, 24Mg, 27Al, 28Si, 39K, 42Ca, 47Ti, 57Fe, 123Sb, 137Ba, 207Pb). The glass samples represented four types of glasses: 1. borosilicate glass (type "Duran") 2. borosilicate glass (type "Fiolax") 3. soda-lime glass 4. lead glass. Calculations were made by using the SAS statistical software. All results were obtained on the assumption that the underlying data are normally distributed. By applying linear discriminant data analysis a classification of all types of the glasses was possible (13 variables included). The use of the method of "stepwise discriminant analysis" reduced the minimal required number of variables to eight. All types of glasses could be distinguished faultlessly. Besides a graphic system consisting of the isotopes 24Mg-47Ti-137Ba was developed. By means of this system optical distinction of glass classes was possible. The developed method allows to distinguish several types of pharmaceutical and common glasses on the basis of intensity data of mass spectrometric measurement without calibration experiments and linear regression. PMID- 12116878 TI - Isolation and identification of the photodegradation products of the photosensitizing antidepressant drug clomipramine. Phototoxicity studies on erythrocytes. AB - The isolation and identification of the photodegradation products of clomipramine (CIP) in phosphate buffered saline (PBS pH 7.4 and 6.0) solution and methanol under aerobic conditions were studied. Six compounds were identified and four of them were isolated and characterized by spectroscopic methods. A radical mechanism with the participation of the solvent is proposed for the photodegradation of CIP which undergoes homolytic cleavage of the carbon-chlorine bond and also photooxidation of the amine group. CIP was able to induce photohemolysis when it was irradiated in PBS pH 7.4 and in PBS pH 6.0 containing a suspension of human red blood cells (RBCs). The photohemolysis experiments in the presence of additives DABCO and GSH showed nearly total inhibition of drug induced photohemolysis. The efficient inhibition of photohemolysis by the radical scavenger GSH compared with the inhibition show by DABCO suggests a moderate effect by singlet oxygen. Clomipramine-N-oxide was the unique photoproduct able to induce hemolysis and photohemolysis when it was incubated and irradiated with RBCs for 1 h. A mechanism involving singlet oxygen, radicals and photoproducts is suggested for the reported phototoxicity. PMID- 12116877 TI - Studies on the photostability and phototoxicity of aloe-emodin, emodin and rhein. AB - Aloe-emodin (1), emodin (2) and rhein (3) were found to be photolabile by visible (390-500 nm) light under aerobic conditions. The drugs 1, 2 and 3 were phototoxic in vitro when examined by the photohemolysis test under both oxygen and argon atmospheres, although the photohemolysis rate was markedly lower under anaerobic conditions. The experiments were also carried out in the presence of butylated hydroxyanisole (BHA), reduced glutathione (GSH), sodium azide (NaN3) and superoxide dismutase (SOD). Based on the inhibition of this process on addition of BHA, GSH, SOD and NaN3, there would seem to be involvement of free radicals (type I mechanism) and singlet oxygen in the process (type II mechanism). The in vitro phototoxicity of this anthraquinone series was also verified in a lipid photoperoxidation test with linoleic acid. In summary, this anthraquinone series is phototoxic in vitro. This behavior can be explained through the involvement of singlet oxygen and stable photoproducts. PMID- 12116879 TI - Synergistic and potentiating effects of ranitidine and two new anti-ulcer compounds from Enantia chlorantha and Voacanga africana in experimental animal models. AB - TN, an alkaloid from the fruit of Voacanga africana and a protoberberine-type alkaloid (7,8-dihydro-8-hydroxypalmatine) (1), obtained from the bark of Enantia chlorantha were tested for ulcer preventive and antisecretory activity in combination with ranitidine. When tested alone (50 and 100 mg/kg, p.o.), TN and 1 achieved their anti-ulcer actions through reduced gastric secretion and improved mucus production. 1:1 combinations of 1 and the antisecretory agents (25/25 and 50/50 mg/kg) resulted in significant reduction of ulceration under highly acidic conditions (50-70 mEq/l), suggesting potentiating effects. A combination of TN and ranitidine led to synergistic antisecretory effects. PMID- 12116880 TI - Antimalarial activity of neurolenin B and derivates of Eupatorium inulaefolium (Asteraceae). AB - Dried stems and leaves of Eupatorium inulaefolium (Austroeupatorium inulaefolium) (Asteraceae) were used to obtain four crude extracts (hexane, dichloromethane, methanol and ethanol). Two fractions were obtained from the hexane extract (S1 and S2) and three compounds (neurolenin B, lobatin A and lobatin B) from the dichloromethane extract. The ethanol, hexane, dichloromethane and methanol extracts, two fractions from the hexane extract (S1 and S2), and neurolenin B were evaluated in vitro against Plasmodium falciparum, FCB-2 strain. Two extracts (dichloromethane and methanol), the S2 fraction and neurolenin B showed statistically significant antiplasmodial activity. PMID- 12116881 TI - Screening of Basidiomycete mushroom extracts for antigenotoxic and bio antimutagenic activity. AB - In this study we screened crude methanol:water extracts of 89 different mushroom species for their antigenotoxic and bio-antimutagenic activity. The screening was performed with the SOS/umu test and we monitored the ability of extracts to inhibit UV induced expression of umuC gene in Salmonella typhimurium TA1535/pSK1002. Seventeen extracts inhibited umuC expression by more than 50%. These extracts were further evaluated for the ability to inhibit UV induced mutations in Escherichia coli WP2. Five extracts (Cortinarius evernius, Rozites caperatus, Lactarius vellereus, Russula integra and Pleurotus cornucopiae) inhibited also UV induced mutations. The study showed that certain mushrooms contain substances with bio-antimutagenic potential. Particularly interesting for further investigations are Pleurotus cornucopiae (Lentinaceae), which was the most effective and species of Russulaceae and Cortinaceae families, which might contain common family specific bio-antimutagenic substance(s). PMID- 12116882 TI - In vitro vascular smooth muscle contractile activity of Aspilia africana extract on rat aortic preparations. AB - Aspilia africana is widely used in ethnomedical practice in Africa for its ability to stop bleeding, even from a severed artery, as well as promote rapid healing of wounds and sores, and for the management of problems related to cardiovascular diseases. In the present paper, the methylene chloride/methanol extract of A. africana leaves was tested for its contractile activity in vitro. Rings of rat aorta, with or without an intact endothelium, were mounted in tissue baths, contracted with norepinephrine, and then exposed to the plant extract. The effect of the extract was also assessed on the baseline tension of aortic rings in normal and calcium-free PSS. At the lower doses, A. africana slowly re inforced contractions induced by norepinephrine and relaxed precontracted tension at the highest concentration. The relaxant activity of the extract was endothelium-independent and was not modified by pre-treatment with Nw-nitro-L arginine methyl ester or indomethacin, suggesting that its effect was not mediated by either nitric oxide or prostacyclin. A. africana extract induced slow and progressive increase in the basal vascular tone which was partially endothelium-dependent. In calcium-free PSS, a high proportion of the contractile activity was inhibited (77%), suggesting that A. africana contractile activity in vascular tissue depends, in part, on extracellular calcium. PMID- 12116883 TI - Antimicrobial activity of Polyscias filicifolia cell biomass extracts. AB - Antibacterial activity of extracts of Polyscias filicifolia biomass from bioreactor and callus was determined using the agar disc-diffusion method. The microorganisms Staphylococcus aureus (three strains) showed the highest sensitivity to extracts of P. filicifolia biomass from a bioreactor. The values were comparable with nitrofurantoine used as a standard. Micrococcus flavus, Sreptococcus pyogenes and S. agalatiae were less sensitive. The effect of P. filicifolia callus extract on the above bacteria was less pronounced than that of extracts of biomass from a bioreactor. PMID- 12116884 TI - Norditerpenoid alkaloids from the roots of Aconitum leave Royle. AB - From the roots of Aconitum leave Royle growing wild in Northern Pakistan, Swat district, three new alkaloids, 8-methyllycaconitine (1), 14-demethyllycaconitine (2), N-deethyllycaconitine-N-aldehyde (3) have been isolated along with four known compounds, lappaconitine (4), lycaconitine (5), lapaconidine (6) and lycoctonine (7). PMID- 12116885 TI - 4-Nitro-1,8-naphthalimides exhibit antinociceptive properties. PMID- 12116886 TI - Bioactive compounds from Leycesteria formosa. PMID- 12116887 TI - FDA states policy on pharmacy compounding. Agency follows up on supreme court ruling. PMID- 12116889 TI - More hospitals report medication errors, but USP finds few changes. PMID- 12116888 TI - CDC guidelines address treatment of HIV, STD infections. PMID- 12116890 TI - Pharmacogenetics and cardiovascular disease: impact on drug response and applications to disease management. AB - The genetic polymorphisms that may affect individual responses to cardiovascular agents are reviewed, and the application of pharmacogenetics to cardiovascular disease management is discussed. Pharmacogenetics is the search for genetic polymorphisms that affect responses to drug therapy. Investigators have found many associations between genetic polymorphisms and responses to cardiovascular drugs. Some of these relationships have been demonstrated in large patient populations, such as patients with ischemic heart disease receiving statins. Study data consistently show a greater response to statins in ischemic heart disease patients with genotypes associated with worse prognoses. Studies of other polymorphisms, such as those in the genes encoding anglotensin-converting enzyme and beta 1-adrenergic receptors, have less consistently found relationships between these variations and cardiovascular drug responses. For gene-drug response associations for which the data are inconsistent, the interaction of multiple polymorphisms in multiple genes coding for proteins affected by drug therapy or influencing drug metabolism may prove to have a greater influence on drug responses than any one polymorphism. Once the polymorphisms that best determine the response to a particular drug are known and tests to rapidly identify these variations are available, individual patients may be screened for genetic polymorphisms before drug therapy is begun and the information used to choose agents with the greatest potential for efficacy and least potential for toxicity. Pharmacogenetics has many possible applications in the drug therapy of cardiovascular diseases. Much more must be learned, however, before pharmacogenetic factors can be routinely incorporated into therapeutic decisions. PMID- 12116891 TI - Investigation of the interaction between argatroban and acetaminophen, lidocaine, or digoxin. AB - The potential pharmacokinetic interactions between argatroban and acetaminophen, lidocaine, or digoxin were examined. Three randomized crossover studies were conducted. In the first study, 11 subjects completed three sessions (with a five day washout period between sessions), receiving two 500-mg acetaminophen caplets at 0, 6, 12, 18, and 24 hours; i.v. argatroban 1.5 micrograms/kg/min from hours 12 to 30; or a combination of both. In the second study, 12 subjects completed three sessions (with a five-day washout period between sessions), receiving lidocaine hydrochloride injection 2 mg/kg/hr for 16 hours (after receiving a loading dose of 1.5 mg/kg over 10 minutes), i.v. argatroban 1.5 micrograms/kg/min for 16 hours, or a combination of both. In the third study, 12 subjects completed two sessions (with a seven-day washout period between sessions), receiving oral digoxin 0.375 mg/day for 15 days and either i.v. placebo or argatroban 2 micrograms/kg/min on days 11 through 15. Primary pharmacokinetic values in each study included area under the drug concentration versus time curve and steady state concentrations of argatroban and the concomitantly administered drug. Lack of a pharmacokinetic interaction (individually defined for each study) was demonstrated in each study. Argatroban, regardless of acetaminophen or lidocaine administration, prolonged activated partial thromboplastin time values approximately 1.6-1.8 times the baseline values. No deaths, unexpected adverse events, or clinically significant changes in safety laboratory values occurred. No pharmacokinetic interaction was detected between argatroban and acetaminophen, lidocaine, or digoxin. Argatroban is well tolerated during coadministration with these drugs. In practice, argatroban coadministered with these frequently prescribed drugs should require no dosage adjustments. PMID- 12116892 TI - Stability of valganciclovir in an extemporaneously compounded oral liquid. PMID- 12116893 TI - Infliximab in graft-versus-host disease. AB - Straightforward reports of unusual drug experiences are included in this section. While selected references may be cited, the purpose of a Drug Experience report is not to present an extensive review of the literature. A related section, Grand Rounds, includes papers that are well-documented patient case reports with a thorough review of the important literature to help put the case in perspective. Authors should report serious adverse drug reactions to the FDA medical products reporting program (MedWatch). Such reporting will not jeopardize the chances that AJHP will publish manuscripts on the same drug reactions. PMID- 12116894 TI - Using medications purchased in Mexico: position statement of the Texas Society of Health-System Pharmacists. PMID- 12116895 TI - Impact of pharmacist home visits on drug therapy. PMID- 12116896 TI - Underutilization of aspirin for secondary prophylaxis of cardiovascular disease in a long-term-care facility. PMID- 12116897 TI - Responsibility for sincalide injection obtained from compounding pharmacies. PMID- 12116898 TI - [Non-occupational dust exposure of the population living in the vicinity of amphibolite mining and processing plant]. AB - Amphibolite bed working in a strip mine, material processing, storage and waste disposal are associated with dust emission to the environment. The aim of our study was to assess non-occupational exposure to dust of people living in a village located about 200 m away from the mine. The fallout dust and the sediment obtained from water samples collected from the stream were found to contain tremolite. Respirable mineral fibers were detected in dust samples collected by filtration of the atmospheric air. Mean 24-h concentrations of the respirable dust at the place of measurements (school) varied considerably, from 11 micrograms/m3 to 20,059 micrograms/m3. The highest dust concentrations were found during the third shift. As fibrous contaminants were detected in many mines of the Lower Silesia region, it is necessary to assess the non-occupational exposure of populations living close to all mines in this area. PMID- 12116899 TI - [Assessment of workers' exposure to airborne bacteria at a small wastewater treatment plant]. AB - A study of workers' exposure to airborne culturable bacteria was performed at the wastewater treatment plant in Myszkow. A six-stage Andersen impactor was used to carry out measurements. The concentrations of total bacterial aerosols ranged from 10(2) to 10(3) CFU/m3 and were lower than the proposed standards. The results of the study show that Gram-negative bacteria contributed to about 35% of the total bacterial aerosol. The largest part of the airborne Gram-negative bacteria constituted Enterobacteriaceae. Pathogenic bacteria found in the air: were as follows: Aeromanas hydrophila, Enterobacter agglomerans, Yersinia enterocolitica, Escherichia coli and Streptococcus faecalis. The microbiological contamination of the air was highest near the aeration basin. Exposure to airborne culturable bacteria may cause respiratory and gastrointestinal symptoms among sewage workers. It is necessary to reduce the exposure to airborne bacteria at this phase of the process of wastewater treatment. PMID- 12116900 TI - [Exposure to selected aldehydes among municipal transport bus drivers]. AB - The objective of the study was to determine occupational exposure of municipal transport bus drovers to selected aldehydes: formaldehyde, acetaldehyde and acrolein. Ten drivers serving 5 bus lines were selected for the study. Air samples were collected in the driver's breathing zone and in bus cabins. The test air (about 10 l) was drawn through columns filled with silicagel coated by dinitrophenylhydrazine (DNPH). The products of DNPH reaction with aldehydes were extracted with acetonitrile and analyzed by HPLC with a spectrophotometric detector (lambda = 360 nm). The analytical procedure enabled to determine the selected aldehydes in the concentration range equivalent to 0.1-2.0 of the Polish maximum allowable concentrations (MAC). In the quantitative analysis, the concentrations of selected aldehydes were found to be much lower than MAC values, regardless of the bus type. The concentrations of formaldehyde and acrolein ranged from 0.025 to 0.090 mg/m3 and from 0.010 to 0.035 mg/m3, respectively. In all samples, acetaldehyde concentrations were well below the limit of detection of the analytical method. The combined exposure to aldehydes was also below the limit value for mixtures. PMID- 12116901 TI - [The usefulness of troponin I in the diagnosis of cardiac damage in acute carbon monoxide poisoning]. AB - Tissue hypoxia in carbon monoxide poisoning is often responsible for cardiac damage that is not always registered in ECG recordings. It is therefore necessary to look for biochemical markers of this damage. Troponin I, the protein not detected in serum of healthy people meets these criteria as its presence proves the cardiac damage. The aim of this paper was to evaluate the usefulness of troponin I determination in diagnosing cardiac damage in acute carbon monoxide poisoning. The study was performed in a group of 44 patients treated because of carbon monoxide poisoning. In addition, other biochemical markers of cardiac damage were determined. The results of the study showed that of the 44 cases of carbon monoxide poisoning, only in 16 patients troponin I was not found in serum. The level of troponin I was increased in all cases with ECG changes, but also in 13 patients with normal ECG. PMID- 12116902 TI - [Evaluation of the computerized program in stress testing in workplace]. AB - The aim of the study was to design a computerized version of two paper-pencil questionnaires: "Objective Work Characteristics" and "Subjective Work Characteristics". The methods allowed to objectively assess the level of occupational stress related to a given work post and to obtain its subjective assessment as perceived by the employees. The computerized version of the questionnaires contributed to fast identification of psychological factors present at workplace and their influence on the level of stress experienced by individual employees. The computer programming simplified scoring, as well as proved to be helpful in a detailed analysis of psychosocial characteristics of the work post and developing periodical reports in any time interval from 0.5 to 5 years. The efficiency of the program was verified on the basis of the study carried out in five sanitary and epidemiological stations located in the city of Lodz. A group of experts evaluated the level of stress at six work posts and 140 workers provided the self-assessment of occupational stress experienced at work. The data were collected and calculated in two databases of the statistical program SPSS+ and the originally designed application. The comparisons of the results obtained in these two programs showed no differences which proved the proper functioning of the program. PMID- 12116903 TI - [The activity of angiotensin converting enzyme in vascular mesenteric bed of rats poisoned with lead and cadmium]. AB - Heavy metals can increase the arterial blood pressure by influencing the endothelial renin-angiotensin system. The aim of this study was to determine the effect of single and combined exposure to lead and cadmium at hypertensive doses on the tissue renin-angiotensin system in rats. An attempt was made to assay the induced synthesis of angiotensin II (AII) in the isolated mesenteric rat bed, as well as the reactivity of vessels to AII and norepinephrine used before and after infusion of angiotensin receptor blocker. Angiotensin converting enzyme (ACE), used in infusion, induced slight increase in perfusion pressure only in mesenteric vessels of cadmium poisoned rats. In this group, AII given during ACE infusion induced slightly stronger vasoconstriction than in other groups of rats, and losartan, used in infusion totally eliminated the pressor effect of AII. Therefore, the share of endothelial renin-angiotensin system in the regulation of vessel resistance was greater than in mesenteric bed of lead-poisoned rats or controls. It is likely that the effect of ACE inhibitors and blockers of the AII receptors may be slightly stronger in persons with arterial hypertension exposed to cadmium than in non-exposed patients. PMID- 12116904 TI - [Cancerogenic effect of occupational exposure to man-made mineral fibers: results from the epidemiological study]. AB - The size and technology of the man-made mineral fibers production in Poland is presented. The results of the case-referent and cohort studies aimed at assessing risk for or mortality from respiratory system cancers in populations occupationally exposed to dust containing artificial mineral fibers are also analyzed. The majority of studies focused on the exposure to glass fibers, however, their results revealed no increased risk for cancers in those exposed. As to the increased risk for respiratory system cancers induced by exposure to glass and slag wool fibers, the same proportion of positive and negative results was found in the literature. Most of the studies showed no association between the risk level and the exposure period or cumulative dose. The association with latency period of more than 20 years was only emphasized. This together with the fact that in the majority of studies neither smoking histories nor exposures to other respiratory carcinogens were considered significantly undermine the cause effect inference. Bearing this mind, it should be concluded that there is an urgent need to undertake further studies with the aim to assess the health risk of exposure to man-made mineral fibers, but only such studies in which besides exposure to fibers, it will be possible to analyze exposure to other coexisting carcinogenic agents. PMID- 12116905 TI - [Risk assessment of known and new chemicals]. AB - It has become obvious that exposure to chemicals entails risk. Their hazardous and threatening effects may be not only direct but also indirect affecting microorganisms, animals and plants. This aspect of hazard and risk assessment is almost identical to that of toxicity. There are various categories of chemicals and for each of them risk assessment regulations have been developed. There are two major groups of chemicals: those which are already known and new ones. There are also some specific categories of chemical compounds, such as pesticides, biocides, medicinal products, cosmetics, food additives, feed additives, as well as radioactive substances and others. At present, the binding national and international regulations regarding different categories of chemical compounds are the subject of discussion in the member states of the European Union. In Poland, and attempt has been made to find appropriate examples of legal regulations that could be followed in our country, and this problem remains still open. PMID- 12116906 TI - [Berylliosis in the work environment: etiology and medical treatment]. AB - Beryllium is a metal responsible for the incidence of chronic beryllium disease- an illness affecting from 2 to 5% of workers exposed to this metal and its compounds. There is a growing evidence provided by epidemiological and toxicological studies that exposure limits for workers dealing with beryllium should be revised. This paper gathers epidemiological and pathological data particularly on chronic beryllium toxicity and carcinogenesis. It also reviews the most important aspects of beryllium toxicology and explains the mechanisms of its effect on humans. In addition, the paper presents suggestions on the diagnosis and treatment of berylliosis. The content of the paper is based on medical data, as well as on reference materials of national and international organizations and governmental agencies. PMID- 12116907 TI - [Occupational health problems in dental practice]. AB - The overload of the osteoarticular system resulting from standing and stooping position of the body is the main health problem of dentists. This may cause vertebral pain, symptoms of sciatica and foot valgo-planus. Symptoms of carpal tunnel syndrome are induced by repeated carpus movements. Frequent numbness associated with the peripheral nerves changes result from using drills by dentists. Menstruation disturbances observed in dental assistants could be related to the increased levels of mercury in serum and urine. Allergy is also a frequent medical problem, particularly allergy to latex. manifested by contact eczema or allergic urticaria, asthma and shock. There also is an increased risk for infectious diseases through the contact with biological material, mostly saliva and blood. PMID- 12116908 TI - [Anthrax as biological warfare weapon]. AB - Anthrax-disease of herbivorous animals, occasionally affecting humans, is regarded as an ideal biowarfare weapon. This was witnessed during a tragic accident in Swierdlowsk some time ago and nowadays it has been observed among post office workers in the USA. Depending on the way the pathogen enters the body, several forms of disease develops: skin anthrax specific for the man (with mortality up to 20%), intestinal anthrax resulting from the consumption of infected food or flash of infected animals (with mortality up to 60-80%) and inhalation pulmonary anthrax (with mortality up to 90%). The lethal effects of disease are due to a toxin composed of three agents. It is recommended that the exposed persons be given antibiotics to prevent them from the development of acute forms of the disease. Bearing in mind that there are B. anthracis strains resistant to Penicillin and Doxycyclin, Ciprofloxacin should be given to the patients, including children, pregnant women (for 60 days until determining antibiotic resistance) and immuno-compromised persons. In single cases- intravenous, and in a large number of cases--oral treatment is recommended. PMID- 12116909 TI - [Commentary on the letter to the editor "Is the May 5 2001 order of the Minister of Health a mistake?"]. PMID- 12116911 TI - [On miracles of our clinical language]. PMID- 12116910 TI - [On investigating sexual abuse of a child]. PMID- 12116913 TI - [What kind of a brain is a spongiform brain?]. PMID- 12116914 TI - [Headache attacks]. PMID- 12116915 TI - [Alternatives of stroke prevention increasing]. PMID- 12116917 TI - [How does the completion of human genome project affect our possibilities to clarify pathogenesis of diseases?]. PMID- 12116916 TI - [Malaria]. PMID- 12116918 TI - [Medicine and genetics--searching for the basis of medicine]. PMID- 12116919 TI - [Nerve root entrapment through a tear in dura mater--a treatable cause of postoperative ischias syndrome]. PMID- 12116920 TI - [Treatment of a renal artery stab wound with selective embolization]. PMID- 12116921 TI - [Can you trust on a biopsy from oral mucosa? The lesion can be cancer, after all]. PMID- 12116922 TI - [Factors that increase the risk of oral cancer and cancer-resembling changes of oral mucosa]. PMID- 12116923 TI - [Investigation and treatment of strabismus]. PMID- 12116924 TI - [Career of a surgeon is searching the balance]. PMID- 12116925 TI - SSZ 1970-1989: a view of the years of conflict. PMID- 12116926 TI - Generalized frequency coding: a method of preparing polymorphic multistate characters for phylogenetic analysis. AB - A new method of coding polymorphic multiistate characters for phylogenetic analysis is presented. By dividing such characters into subcharacters, their frequency distributions can be represented with discrete states. Differential weighting is used to counter the effect of representing one character with multiple characters. The new method, generalized frequency coding (GFC), is potentially superior to previously used methods in that it incorporates more information and is applicable to both qualitative and quantitative characters. When applied to a previously published data set that includes both types of polymorphic multistate characters, the method performed well, as assessed with g1 and nonparametric bootstrap statistics and giving results congruent with those of other studies. The data set was also used to compare GFC with both gap-weighting and Manhattan distance step matrix coding. On these grounds and for philosophical reasons, we consider GFC to be a better estimator of phylogeny. PMID- 12116927 TI - The phylogenetic trunk: maximal inclusion of taxa with missing data in an analysis of the lepospondyli (Vertebrata, Tetrapoda). AB - The importance of fossils to phylogenetic reconstruction is well established. However, analyses of fossil data sets are confounded by problems related to the less complete nature of the specimens. Taxa that are incompletely known are problematic because of the uncertainty of their placement within a tree, leading to a proliferation of most-parsimonious solutions and "wild card" behavior. Problematic taxa are commonly deleted based on a priori criteria of completeness. Paradoxically, a taxon's problematic behavior is tree dependent, and levels of completeness are not directly associated with problematic behavior. Exclusion of taxa on the basis of completeness eliminates real character conflict and, by not allowing incomplete taxa to determine tree topology, diminishes the phylogenetic hypothesis. Here, the phylogenetic trunk approach is proposed to allow optimization of taxonomic inclusion and tree stability. The use of this method in an analysis of the Paleozoic Lepospondyli finds a single most-parsimonious tree, or trunk, after the removal of one taxon identified as being problematic. Moreover, the 38 trees found at one additional step from this primary trunk were reduced to 2 by removal of one additional taxon. These trunks are compared with the trees that were found by excluding taxa with various degrees of completeness, and the effects of incomplete taxa are explored with regard to use of the trunk. Correlated characters associated with limblessness are discussed regarding the assumption of character independence; however, inclusion of intermediate taxa is found to be the single best method for breaking down long branches. PMID- 12116928 TI - Multiple molecular data sets suggest independent origins of highly eusocial behavior in bees (Hymenoptera:Apinae). AB - Different views of the pattern of social evolution among the highly eusocial bees have arisen as a result of discordance between past molecular and morphology based phylogenies. Here we present new data and taxa for four molecular data sets and reassess the morphological characters available to date. We show there is no significant character incongruence between four molecular data sets (two nuclear and two mitochondrial), but highly significant character incongruence leads to topological incongruence between the molecular and morphological data. We investigate the effects of using different outgroup combinations to root the estimated tree. We also consider various ways in which biases in the sequence data could be misleading, using several maximum likelihood models, LogDet corrections, and spectral analyses. Ultimately, we concede there is strong discordance between the molecular and morphological data partitions and appropriately apply the conditional combination approach in this case. We also find two equally well supported placements of the root for the molecular trees, one supported by 16S and 28S sequences, the other supported by cytochrome b and opsin. The strength of the evidence leads us to accept two equally well supported hypotheses based on analyses of the molecular data sets. These are the most rigorously supported hypotheses of corbiculate bee relationships at this time, and frame our argument that highly eusocial behavior within the corbiculate bees evolved twice independently. PMID- 12116930 TI - Phylogenetic relationships of fig wasps pollinating functionally dioecious Ficus based on mitochondrial DNA sequences and morphology. AB - The obligate mutualism between pollinating fig wasps in the family Agaonidae (Hymenoptera: Chalcidoidea) and Ficus species (Moraceae) is often regarded as an example of co-evolution but little is known about the history of the interaction, and understanding the origin of functionally dioecious fig pollination has been especially difficult. The phylogenetic relationships of fig wasps pollinating functionally dioecious Ficus were inferred from mitochondrial cytochrome oxidase gene sequences (mtDNA) and morphology. Separate and combined analyses indicated that the pollinators of functionally dioecious figs are not monophyletic. However, pollinator relationships were generally congruent with host phylogeny and support a revised classification of Ficus. Ancestral changes in pollinator ovipositor length also correlated with changes in fig breeding systems. In particular, the relative elongation of the ovipositor was associated with the repeated loss of functionally dioecious pollination. The concerted evolution of interacting morphologies may bias estimates of phylogeny based on female head characters, but homoplasy is not so strong in other morphological traits. The lesser phylogenetic utility of morphology than of mtDNA is not due to rampant convergence in morphology but rather to the greater number of potentially informative characters in DNA sequence data; patterns of nucleotide substitution also limit the utility of mtDNA findings. Nonetheless, inferring the ancestral associations of fig pollinators from the best-supported phylogeny provided strong evidence of host conservatism in this highly specialized mutualism. PMID- 12116931 TI - Australian Lasioglossum + Homalictus form a monophyletic group: resolving the "Australian enigma". AB - The bee genus Lasioglossum includes > 1,000 species of bees distributed on all continents except Antarctica. Lasioglossum is a major component of the bee fauna in the Holarctic, Ethiopian, and Asian regions and is an important group for investigating the evolution of social behavior in bees. Given its cosmopolitan distribution, the historical biogeography of the genus is of considerable interest. We reconstructed phylogenetic relationships among the subgenera and species within Lasioglossum s.s., using DNA sequence data from a slowly evolving nuclear gene, elongation factor-1 alpha. The entire data set includes > 1,604 aligned nucleotide sites (including three exons plus two introns) for 89 species (17 outgroups plus 72 ingroups). Parsimony and maximum likelihood analyses provide strong evidence that the primarily Indoaustralian subgenera (Homalictus, Chilalictus, Parasphecodes) form a monophyletic group. Bootstrap support for the Australian clade ranged from 73% to 77%, depending on the method of analysis. Monophyly of the Australian Lasioglossum suggests that a single colonization event (by way of Southeast Asia and New Guinea) gave rise to a lineage of > 350 native Indoaustralian bees. We discuss the implications of Australian monophyly for resolving the "Australian enigma"--the similarity in social behavior among the Australian halictine bees relative to that of Holarctic groups. PMID- 12116929 TI - Analysis of character correlations among wood decay mechanisms, mating systems, and substrate ranges in homobasidiomycetes. AB - Homobasidiomycetes include the majority of wood-decaying fungi. Two basic forms of wood decay are known in homobasidiomycetes: white rot, in which lignin and cellulose are degraded, and brown rot, in which lignin is not appreciably degraded. An apparent correlation has been noted between production of a brown rot, decay of conifer substrates, and possession of a bipolar mating system (which has a single mating-type locus, in contrast to tetrapolar systems, which have two mating-type loci). The goals of this study were to infer the historical pattern of transformations in decay mode, mating type, and substrate range characters, and to determine if a causal relationship exists among them. Using nuclear and mitochondrial rDNA sequences, we performed a phylogenetic analysis of 130 species of homobasidiomycetes and performed ancestral state reconstructions by using parsimony on a range of trees, with various loss:gain cost ratios. We evaluated pairwise character correlations by using the concentrated changes test (CCT) of Maddison and the maximum likelihood (ML) method of Pagel. White rot, tetrapolar mating systems, and the ability to decay conifers and hardwoods appear to be plesiomorphic in homobasidiomycetes, whereas brown rot, bipolar mating systems, and exclusive decay of conifers appear to have evolved repeatedly. The only significant correlation among characters was that between brown rot (as the independent character) and exclusive decay of conifer substrates (P < 0.03). This correlation was supported by the CCT on a range of plausible trees, although not with every reconstruction of ancestral states, and by the ML test. Our findings suggest that the evolution of brown rot has promoted repeated shifts to specialization for confier substrates. PMID- 12116932 TI - Does the T-PTP test tell us anything we want to know? PMID- 12116933 TI - Species or supraspecific taxa as terminals in cladistic analysis? Groundplans versus exemplars revisited. PMID- 12116934 TI - Strengths and limitations of the minimum evolution principle. PMID- 12116935 TI - Random sampling of constrained phylogenies: conducting phylogenetic analyses when the phylogeny is partially known. AB - Statistical randomization tests in evolutionary biology often require a set of random, computer-generated trees. For example, earlier studies have shown how large numbers of computer-generated trees can be used to conduct phylogenetic comparative analyses even when the phylogeny is uncertain or unknown. These methods were limited, however, in that (in the absence of molecular sequence or other data) they allowed users to assume that no phylogenetic information was available or that all possible trees were known. Intermediate situations where only a taxonomy or other limited phylogenetic information (e.g., polytomies) are available are technically more difficult. The current study describes a procedure for generating random samples of phylogenies while incorporating limited phylogenetic information (e.g., four taxa belong together in a subclade). The procedure can be used to conduct comparative analyses when the phylogeny is only partially resolved or can be used in other randomization tests in which large numbers of possible phylogenies are needed. PMID- 12116936 TI - Comparing ontogenetic trajectories using growth process data. AB - Ontogenetic trajectories are commonly quantified by characterizing changes in the sizes and shapes of organisms over the course of development. This formulation of ontogenetic transformations can be misleading in that it ignores critical aspects of the biological processes responsible for constructing morphology. Hypothetical examples are used to illustrate some of the shortcomings of methods that rely exclusively on size and shape data for ontogenetic analyses. By characterizing growth as a vector field, and representing growth vectors as complex numbers, one can simultaneously analyze size, shape, and growth processes. The utility of such an approach is demonstrated in a study of shape and growth process variation in turtle shells. PMID- 12116937 TI - Inferring and testing hypotheses of cladistic character dependence by using character compatibility. AB - The notion that two characters evolve independently is of interest for two reasons. First, theories of biological integration often predict that change in one character requires complementary change in another. Second, character independence is a basic assumption of most phylogenetic inference methods, and dependent characters might confound attempts at phylogenetic inference. Previously proposed tests of correlated character evolution require a model phylogeny and therefore assume that nonphylogenetic correlation has a negligible effect on initial tree construction. This paper develops "tree-free" methods for testing the independence of cladistic characters. These methods can test the character independence model as a hypothesis before phylogeny reconstruction, or can be used simply to test for correlated evolution. We first develop an approach for visualizing suites of correlated characters by using character compatibility. Two characters are compatible if they can be used to construct a tree without homoplasy. The approach is based on the examination of mutual compatibilities between characters. The number of times two characters i and j share compatibility with a third character is calculated, and a pairwise shared compatibility matrix is constructed. From this matrix, an association matrix analogous to a dissimilarity matrix is derived. Eigenvector analyses of this association matrix reveal suites of characters with similar compatibility patterns. A priori character subsets can be tested for significant correlation on these axes. Monte Carlo tests are performed to determine the expected distribution of mutual compatibilities, given various criteria from the original data set. These simulated distributions are then used to test whether the observed amounts of nonphylogenetic correlation in character suites can be attributed to chance alone. We have applied these methods to published morphological data for caecilian amphibians. The analyses corroborate instances of dependent evolution hypothesized by previous workers and also identify novel partitions. Phylogenetic analysis is performed after reducing correlated suites to single characters. The resulting cladogram has greater topological resolution and implies appreciably less change among the remaining characters than does a tree derived from the raw data matrix. PMID- 12116938 TI - mtDNA ribosomal gene phylogeny of sea hares in the genus Aplysia (Gastropoda, Opisthobranchia, Anaspidea): implications for comparative neurobiology. AB - Sea hares within the genus Aplysia are important neurobiological model organisms; as more studies based on different Aplysia species are appearing in the literature, a phylogenetic framework has become essential. We present a phylogenetic hypothesis for this genus, based on portions of two mitochondrial genes (12S and 16S). In addition, we reconstruct the evolution of several behavioral characters of interest to neurobiologists to illustrate the potential benefits of a phylogeny for the genus Aplysia. These benefits include determination of ancestral traits, direction and timing of evolution of characters, prediction of the distribution of traits, and identification of cases of independent acquisition of traits within lineages. This last benefit may prove especially useful in understanding the linkage between behaviors and their underlying neurological bases. PMID- 12116939 TI - Character analysis in morphological phylogenetics: problems and solutions. AB - Many aspects of morphological phylogenetics are controversial in the theoretical systematics literature and yet are often poorly explained and justified in empirical studies. In this paper, I argue that most morphological characters describe variation that is fundamentally quantitative, regardless of whether they are coded qualitatively or quantitatively by systematists. Given this view, three fundamental problems in morphological character analysis (definition, delimitation, and ordering of character states) may have a common solution: coding morphological characters as continuous quantitative traits. A new parsimony method (step-matrix gap-weighting, a modification of Thiele's approach) is proposed that allows quantitative traits to be analyzed as continuous variables. The problem of scaling or weighting quantitative characters relative to qualitative characters (and to each other) is reviewed, and three possible solutions are described. The new coding method is applied to data from hoplocercid lizards, and the results show the sensitivity of phylogenetic conclusions to different scaling methods. Although some authors reject the use of continuous, overlapping, quantitative characters in phylogenetic analysis, quantitative data from hoplocercid lizards that are coded using the new approach contain significant phylogenetic structure and exhibit levels of homoplasy similar to those seen in data that are coded qualitatively. PMID- 12116940 TI - Repeated evolution of dioecy from monoecy in Siparunaceae (Laurales). AB - Siparunaceae comprise Glossocalyx with one species in West Africa and Siparuna with 65 species in the neotropics; all have unisexual flowers, and 15 species are monoecious, 50 dioecious. Parsimony and maximum likelihood analyses of combined nuclear ribosomal ITS and chloroplast trnL-trnF intergenic spacer sequences yielded almost identical topologies, which were used to trace the evolution of the two sexual systems. The African species, which is dioecious, was sister to all neotropical species, and the monoecious species formed a grade basal to a large dioecious Andean clade. Dioecy evolved a second time within the monoecious grade. Geographical mapping of 6,496 herbarium collections from all species sorted by sexual system showed that monoecy is confined to low-lying areas (altitude < 700 m) in the Amazon basin and southern Central America. The only morphological trait with a strong phylogenetic signal is leaf margin shape (entire or toothed), although this character also correlates with altitude, probably reflecting selection on leaf shapes by temperature and rainfall regimes. The data do not reject the molecular clock, and branch lengths suggest that the shift to dioecy in the lowlands occurred many million years after the shift to dioecy in the ancestor of the Andean clade. PMID- 12116941 TI - Maximum likelihood estimation of phylogenetic trees is consistent when substitution rates vary according to the invariable sites plus gamma distribution. AB - Maximum likelihood estimation of phylogenetic trees from nucleotide sequences is completely consistent when nucleotide substitution is governed by the general time reversible (GTR) model with rates that vary over sites according to the invariable sites plus gamma (I + gamma) distribution. PMID- 12116943 TI - Bilaterian phylogeny and uncritical recycling of morphological data sets. PMID- 12116942 TI - Should we use model-based methods for phylogenetic inference when we know that assumptions about among-site rate variation and nucleotide substitution pattern are violated? PMID- 12116944 TI - Improving interpretation of the decay index for DNA sequence data. PMID- 12116945 TI - [Cracking sounds in the jaw joint]. PMID- 12116946 TI - [Musical surgery]. PMID- 12116948 TI - [The value of clinical (retrospective) studies]. PMID- 12116947 TI - [Reaction to the illustration in, "Mention of tooth discoloration by drugs," Ned Tijdschr Tandheelkd 1996; 103: 3-4]. PMID- 12116949 TI - [Dental hygienists and local anesthesia: temporarily not on their own authority]. PMID- 12116950 TI - [The authority of the dental hygienist: which are able and which are not?]. PMID- 12116952 TI - Proceedings of the Fifth Annual Meeting of the International Forum Tuohilampi. Essen, Germany. 2001. PMID- 12116951 TI - [Charles Rob (1913-2001)]. PMID- 12116953 TI - [New scenarios for medical science]. PMID- 12116954 TI - [IBAT 2001, Naples]. PMID- 12116955 TI - [Work schedule for residents]. PMID- 12116956 TI - [ACGME. Accreditation Council for Graduate Medical Education. Recommendations on the schedule for medical residents and their training environment]. PMID- 12116958 TI - Proposed General ACGME Requirements for Resident Duty Hours and Supervision. PMID- 12116957 TI - [Comments on the report of the Accreditation Council for Graduate Medical Education on the hours on duty for residents and the learning environment]. PMID- 12116959 TI - [Alphabetical list of the members of the Spanish Neurosurgery Society]. PMID- 12116960 TI - Autoimmune hemolytic anemia in patients infected with human immunodeficiency virus-1. AB - Four men were diagnosed with human immunodeficiency virus infection (AIDS) and autoimmune hemolytic anemia (HIV-AIHA) during the years 1997-2000 at Cook County Hospital, Chicago. All patients presented with the acute onset of severe hemolytic anemia, fever, and splenomegaly. The direct and indirect antiglobulin tests were positive in all, and three patients had mixed warm and cold autoantibody hemolytic anemia. Two patients responded to prednisone therapy and remain in remission from AIHA for 15 and 30 months, respectively. PMID- 12116962 TI - 2002 Resource guide. Company product listing. PMID- 12116961 TI - 2002 Resource guide. Company alphabetical listing. PMID- 12116963 TI - Paracetamol recall: a natural experiment influencing analgesic poisoning. PMID- 12116964 TI - The right number of doctors. A collaborative endeavour based on Delphi-type surveys carried out in 2000-2001 in five countries. PMID- 12116965 TI - Proceedings of the First International Conference on Rural Aging: a Global Challenge. Charleston, West Virginia, USA. June 2000. PMID- 12116966 TI - Involvement of SipA in modulating actin dynamics during Salmonella invasion into cultured epithelial cells. AB - Salmonella entry into epithelial host cells results from the host actin cytoskeleton reorganization that is induced by a group of bacterial proteins delivered to the host cells by the Salmonella type III secretion system. SopE, SopE2 and SopB activate CDC42 and Rac1 to intercept the signal transduction pathways involved in actin cytoskeleton rearrangements. SipA and SipC directly bind actin to modulate the actin dynamics facilitating bacterial entry. Biochemical studies have indicated that SipA decreases the critical concentration for actin polymerization and may be involved in promoting the initial actin polymerization in Salmonella-induced actin reorganization. In this report, we conducted experiments to analyze the in vivo function(s) of SipA during Salmonella invasion. SipA was found to be preferentially associated with peripheral cortical actin filaments but not stress fibres using permeabilized epithelial cells. When polarized Caco-2 cells were infected with Salmonella, actin cytoskeleton rearrangements induced by the wild-type strain had many filopodia structures that were intimately associated with the bacteria. In contrast, ruffles induced by the sipA null mutant were smoother and distant from the bacteria. We also found that the F-actin content in cells infected with the sipA mutant decreased nearly 80% as compared to uninfected cells or those infected with the wild-type Salmonella strain. Furthermore, expression of either the full-length or the SipA(459-684) actin-binding fragment induced prominent punctuate actin assembly in the cortical region of COS-1 cells. These results indicate that SipA is involved in modulating actin dynamics in cultured epithelial cells during Salmonella invasion. PMID- 12116967 TI - Prostate cancer incidence in the world. PMID- 12116968 TI - AIDS and privacy groups criticize proposed changes to federal privacy rule. AB - Privacy advocates blast the Bush administration's proposed changes to the 1996 Health Insurance Portability and Accountability Act and say the changes undermine some of the privacy rule's most important strengths, including the consent requirement prior to disclosures about private medical information. The American Hospital Association counters that the proposed changes will save money, cut bureaucracy and paperwork, and still provide adequate privacy protections. PMID- 12116969 TI - HHS seeks to 'fix' aspects of privacy rule. AB - While critics of proposed changes to the federal health care privacy rule are most concerned about the elimination of a signed consent, they also say some of the other proposals will seriously undermine patients' right to privacy. PMID- 12116970 TI - Minority patients ignored in HIV drug trials. AB - Using a snapshot representing all HIV patients in the United States, researchers found that African-Americans and Latinos had substantially less access to clinical trials than were whites. 'This was true even when they had comparable health insurance, socioeconomic status, and education to whites,' says Allen L. Gifford, MD, assistant professor of medicine at the VA Healthcare System in San Diego. The analysis was conducted as part of the RAND-led HIV Cost and Services Utilization Study Consortium, which is sponsored by the Agency for Healthcare Research and Quality in Rockville, MD. PMID- 12116971 TI - Acute HIV infection transmits greater risk. AB - It's long been hypothesized that semen viral load is the explanation for why there appears to be greater HIV transmission associated with acute infection. Now a study confirms this perception. People who have sex with a partner during an acute HIV infection phase may be at a 20-fold greater risk per exposure than are partners of individuals who are at a virologic set point, according to a recent study. PMID- 12116973 TI - Future spread of HIV will be drug-resistant strains. AB - in a collaboration that includes researchers from California and the United Kingdom, a recent study predicts that people who are newly infected with drug resistant virus will be the major source of new resistant infections. 'We wished to explore the underlying causes of drug resistance, and made use of retrospective clinical data,' says Andrew Leigh Brown, PhD, a visiting professor at the University of California - San Diego and chief author of the study. Brown also is a professor at the University of Edinburgh in Scotland. PMID- 12116972 TI - Separate lineages of HIV super-fast progressors. AB - Researchers at the University of Washington in Seattle made an interesting discovery during their routine study of a cohort of HIV-infected patients: Two men in the cohort were discovered to have two divergent subtype B sequences, indicating they had two independent viruses from independent people. PMID- 12116974 TI - New HIV guidelines offer adverse effects data. AB - The 'Guidelines for Using Antiretroviral Agents Among HIV-Infected Adults and Adolescents,' which are recommendations of the Panel on Clinical Practices for Treatment of HIV, have been updated with additional information linking the use of antiretroviral therapy to chronic diseases and adverse events. PMID- 12116975 TI - FDA notification. Shortage of test kits for detecting HIV-1 antibodies. PMID- 12116976 TI - [Applications and advantages of multimedia systems in the autopsy practice]. PMID- 12116977 TI - WORLD's 11th anniversary: a focus on living. PMID- 12116978 TI - The GFATM (Global Fund to Fight AIDS, Tuberculosis, and Malaria): which countries owe, and how much? PMID- 12116979 TI - I-Med Exchange. Bridging the digital divide. PMID- 12116980 TI - WHO draft guidelines for antiretroviral therapy in resource limited settings. PMID- 12116981 TI - New CDC STD treatment guidelines. PMID- 12116983 TI - Peripheral neuropathy and HIV. PMID- 12116982 TI - Acute neuromyopathy syndrome. PMID- 12116984 TI - Substance abuse and HIV. PMID- 12116985 TI - Formation of [Ar*Ge(CH2C(Me)C(Me)CH2)CH2C(Me)=]2 (Ar* = C6H3-2,6-Trip2; Trip = C6H2-2,4,6-i-Pr3) via reaction of Ar*GeGeAr* with 2,3-dimethyl-1,3-butadiene: evidence for the existence of a germanium analogue of an alkyne. AB - The reduction of Ar*GeCl (Ar* = C6H3-2,6-Trip2; Trip = C6H2-2,4,6-i-Pr3) with one equivalent of potassium leads to the formation of a germanium analogue of an alkyne Ar*GeGeAr* 1; reaction of 1 with 2,3-dimethyl-1,3-butadiene yields [Ar*Ge(CH2C(Me)C(Me)CH2)CH2C(Me)=]2 2, which was structurally characterized. PMID- 12116986 TI - Turner syndrome associated with acquired von Willebrand disease, primary biliary cirrhosis, and inflammatory bowel disease. AB - We report a unique case of Turner syndrome associated with acquired von Willebrand disease (AvWD), primary biliary cirrhosis (PBC), and inflammatory bowel disease (IBD). During 7 years of close follow-up, the patient presented with multiple major episodes of upper and lower gastrointestinal bleeding caused by different pathogenic mechanisms, such as IBD, AvWD, gastric varices, and thrombocytopenia. AvWD mimicking familial vWD type III on laboratory testing was most probably triggered by autoimmune mechanism associated with PBC. Therapy of PBC with ursodeoxycholic acid (UDCA) resulted in significant decrease of liver enzymes followed by normalization of vWF and FVIII levels. Portosystemic shunt placement with ligation of gastric varices improved hypersplenism and severe thrombocytopenia and led to clinical stability for more than 24 months. The clinicopathological features of these disorders and of the recurrent bleeding episodes are discussed in the text along with a review of the literature. PMID- 12116987 TI - Hypertension guidelines in elderly patients: is anybody listening? AB - PURPOSE: Previous guidelines for the management of uncomplicated hypertension in elderly patients have recommended diuretic agents and then beta-blockers. We examined trends in the initial treatment choice for elderly people with hypertension and the effects of a government-sponsored program to publish and disseminate a simplified version of the national guidelines for the treatment of hypertension to all physicians in Ontario, Canada. SUBJECTS AND METHODS: Linked administrative databases containing information on the more than 1.2 million elderly residents in Ontario were used. Time series analysis was performed to determine prescribing trends from 1993 to 1998 for patients who began antihypertensive medication for the treatment of hypertension and to examine the effects of the simplified guidelines that were distributed in July of 1995. RESULTS: Diuretic agents were the most commonly prescribed medications, with annual rates from 35% to 38% (P = 0.59) throughout the study. Beta-blocker prescribing rose from 12% to 16% (P <0.01), whereas angiotensin-converting enzyme (ACE) inhibitor prescribing rose from 27% to 32% (P <0.01). Prescriptions for calcium channel blockers dropped significantly, from 23% to 14% (P <0.01). However, the publication and dissemination of the Ontario hypertension guidelines had no statistically significant effects on the proportion of patients who began treatment with a diuretic agent (P = 0.55), beta-blocker, (P = 0.32), ACE inhibitor (P = 0.09), or calcium channel blocker (P = 0.07). CONCLUSION: The dissemination of simplified practice guidelines for hypertension did not have notable effects on prescribing patterns in Ontario. PMID- 12116988 TI - Cases from the Osler Medical Service at Johns Hopkins University. PMID- 12116990 TI - Depth perception from pairs of overlapping cues in pictorial displays. AB - The experiments reported herein probe the visual cortical mechanisms that control near-far percepts in response to two-dimensional stimuli. Figural contrast is found to be a principal factor for the emergence of percepts of near versus far in pictorial stimuli, especially when stimulus duration is brief. Pictorial factors such as interposition (Experiment 1) and partial occlusion (Experiments 2 and 3) may cooperate, as generally predicted by cue combination models, or compete with contrast factors in the manner predicted by the FACADE model. In particular, if the geometrical configuration of an image favors activation of cortical bipole grouping cells, as at the top of a T-junction, then this advantage can cooperate with the contrast of the configuration to facilitate a near-far percept at a lower contrast than at an X-junction. Varying the exposure duration of the stimuli shows that the more balanced bipole competition in the X junction case takes longer exposures to resolve than the bipole competition in the T-junction case (Experiment 3). PMID- 12116991 TI - Color constancy: the role of low-level mechanisms. AB - Color constancy (CC) is an important psychophysical phenomenon, which has been studied extensively. However, it is not clearly understood. This study presents a novel biological model for the contribution of low level mechanisms to CC. The model is based on two chromatic adaptation mechanisms in the color-coded retinal ganglion cells, 'local' and 'remote', which cause a 'curve-shifting' effect at each receptive field subregion. Simulations are employed for calculating the perceived image and measuring the degree of CC using both 'human-perception' and 'machine-vision' indices. The results indicate that the contribution of adaptations to CC is significant, robust and in agreement with experimental findings. The model is successful in performing CC under multiple chromatic illumination sources, a condition which mimics common natural environments. PMID- 12116989 TI - Clinicopathological features of hepatitis C virus infection in dialysis and renal transplantation. AB - BACKGROUND: Liver biopsy (LB) gives an accurate picture of the severity of hepatitis C virus (HCV) infection in end-stage renal disease. The aim of this study was to find out whether clinical and histopathological course of HCV infection in renal transplant (RT) patients (pts) is different from dialysis (Dx) pts. METHODS: Forty Dx and 46 RT pts underwent LB. Clinical and biochemical data were retrospectively collected from medical charts. ALT level above the upper limit was described as elevated. LB was done regardless of the ALT level. LB specimens were examined using a semiquantitative scoring system locally modified from Scheuer. Histological activity (grade) and fibrosis (stage) were scored separately. RESULTS: ALT was elevated in 65% of Dx pts. At the time of LB 30% of Dx pts had elevated ALT and 95% were viremic. Normal/minimal inflammation was detected in 25% of LBs, chronic hepatitis in 72.5%, cirrhosis in 2.5%. Stage and grade were respectively 1.08 +/- 1.02 and 4.30 +/- 2.98. Normal/minimal inflammation was detected in 9% of the 46 RT pts, chronic hepatitis in 84%, cirrhosis in 7%. Stage and grade were respectively 1.74 +/- 1.1 and 5.39 +/- 2.21. Although there was no significant difference in the histological grade between Dx and RT pts, histological stage was significantly higher in RT pts than Dx. The frequency of cirrhosis, hepatitis and normal inflammation was similar in the two groups. CONCLUSION: Histopathological liver injury due to HCV infection seems to be more severe in RT than Dx pts but this does not seem to be clear at the clinical and biochemical level. Sequential histopathological assessment and longer follow-up will be required to clarify this issue. PMID- 12116992 TI - Properties of some variants of adaptive staircases with fixed step sizes. AB - Because the estimation of thresholds is daily practice in sensory psychophysics, efficient methods must be used to reduce experimental cost and burden. A large number of such methods are available, and each one further has a multitude of variants. All methods presumably provide a threshold estimate that is the stimulus level at which repeated testing would result in a specific percentage of correct responses on a forced-choice task, a percentage that varies across methods and variants thereof. A recent study (Garcia-Perez, 1998) showed that the most popular method (up-down staircases with fixed step sizes) yields threshold estimates that do not correspond to the presumed percent-correct points. Two modifications of this type of staircase have recently been proposed. In one (Zwislocki and Relkin, 2001), the up-down rule does not require correct responses to occur consecutively. In the other (Kaernbach, 1999), subjects are allowed to respond 'don't know' instead of guessing at random when unsure. Although the statistical basis of either modification were described in general, only a few of their many variants were subjected to evaluation under a limited set of conditions. This paper provides an extensive evaluation of a reasonable number of variants of either modification under a broad set of conditions. The results show that they are generally unfit for threshold estimation because in most cases the percent-correct point that is targeted varies greatly with the relative size of the steps with respect to the spread of the psychometric function. Dependable conditions for the use of these modified staircases are also determined. PMID- 12116993 TI - Two-dimensional motion perception without feature tracking. AB - Feature-tracking explanations of 2D motion perception are fundamentally distinct from motion-energy, correlation, and gradient explanations, all of which can be implemented by applying spatiotemporal filters to raw image data. Filter-based explanations usually suffer from the aperture problem, but 2D motion predictions for moving plaids have been derived from the intersection of constraints (IOC) imposed by the outputs of such filters, and from the vector sum of signals generated by such filters. In most previous experiments, feature-tracking and IOC predictions are indistinguishable. By constructing plaids in apparent motion from missing-fundamental gratings, we set feature-tracking predictions in opposition to both IOC and vector-sum predictions. The perceived directions that result are inconsistent with feature tracking. Furthermore, we show that increasing size and spatial frequency in Type 2 missing-fundamental plaids drives perceived direction from vector-sum toward IOC directions. This reproduces results that have been used to support feature-tracking, but under experimental conditions that rule it out. We discuss our data in the context of a Bayesian model with a gradient-based likelihood and a prior favoring slow speeds. We conclude that filter-based explanations alone can explain both veridical and non-veridical 2D motion perception in such stimuli. PMID- 12116994 TI - Effect of exposure duration, contrast and base blur on coding and discrimination of edges. AB - We extend a neural network model, developed to examine neural correlates for the dynamic synthesis of edges from luminance gradients (Ogmen, 1993), to account for the effects of exposure duration, base blur and contrast on the perceived sharpness of edges. This model of REtino-COrtical Dynamics (RECOD) predicts that (i) a decrease in exposure duration causes an increase in the perceived blur and the blur discrimination threshold for edges, (ii) this increase in perceived blur is more pronounced for sharper edges than for blurred edges, (iii) perceived blur is independent of contrast while the blur discrimination threshold decreases with contrast, (iv) perceived blur increases with increasing base blur while the blur discrimination threshold has a nonmonotonic U-shaped dependence on base blur, (v) the perceived location of an edge shifts progressively towards the low-luminance side of the edge with increasing contrast, and (vi) perceived contrast of suprathreshold stimuli is essentially independent of spatial frequency over a wide range of contrast values. These predictions are shown to be in quantitative agreement with existing psychophysical data from the literature and with data collected on three observers to quantify the effect of exposure duration on perceived blur. PMID- 12116995 TI - Fixating on the wallpaper illusion: a commentary on 'The role of vergence in the perception of distance: a fair test of Bishop Berkeley's claim' by Logvinenko et al. (2001). PMID- 12116996 TI - Bone status in pregnancy and lactation. PMID- 12116997 TI - Outcomes in young adulthood for very-low-birth-weight infants. PMID- 12116998 TI - Outcomes in young adulthood for very-low-birth-weight infants. PMID- 12116999 TI - Outcomes in young adulthood for very-low-birth-weight infants. PMID- 12117000 TI - Oral and topical corticosteroids in bullous pemphigoid. PMID- 12117001 TI - Oral and topical corticosteroids in bullous pemphigoid. PMID- 12117002 TI - Oral and topical corticosteroids in bullous pemphigoid. PMID- 12117003 TI - Case 8-2002: pleural effusion. PMID- 12117005 TI - Taxonomic bias in conservation research. PMID- 12117004 TI - Healthy People 2010 criteria for data suppression. PMID- 12117006 TI - Igniting nanotubes with a flash. PMID- 12117007 TI - Being human. PMID- 12117008 TI - Mother knows best. PMID- 12117009 TI - The Medicare appeals process. PMID- 12117010 TI - Place and practice in field biology. PMID- 12117011 TI - Comparative study of the introduction of modern botany in Japan and China. AB - Prior to the eighteenth-century, a similar approach towards the vegetable kingdom, mainly influenced by the tradition of the Chinese pharmacopoeias, could be observed in China and Japan. During the eighteenth-century, the interest for "Dutch learning" led some Japanese physicians and interpreters to be more and more interested in Western knowledge about medicinal plants. At the beginning of the nineteenth-century, a few scholars, through direct contact with foreigners or with foreign books, realised that there was a specific scientific field called "botany" and began to introduce the Japanese scholarly community to this new science which became one of the subjects taught at the "University of Tokyo" in 1877. In China, up to the middle of the nineteenth-century, no trace of modern botany can be found in any published document. In the second half of the century, a few botanical treatises were published, all being adaptations or translations of Western books, done by foreign-Chinese teams of translators. This situation began to change when Chinese students had the opportunity to go and study abroad, mainly to Japan, at the beginning of the twentieth-century, and, actually, it is between 20 and 30 years later that botany became a real scientific practice in China. We will analyse these two processes, their specificities and their interactions. PMID- 12117012 TI - [Music and pharmacy]. PMID- 12117013 TI - [Pharmacy in Canton Vaud during the period of radical domination: 1845-1945]. AB - Between 1845 and 1945, major changes took place both in society and in pharmacy. These two dates represent important phases of mutation for the pharmacy profession. In 1845, university studies were about to become compulsory. Towards 1945, the prodigious development of patent medicines had been disconcerting the pharmaceutical community. In this time period, pharmacy had taken a more modern face: School of pharmacy was founded (1873), professional associations were being formed up and the first hospital pharmacy was established in 1882. Politically, this period of time was marked by the hegemony of the Radical party. Society was at a turning point. The expanding railway network and the development of industry in certain regions were influencing the implantation of pharmacies and their unequal distribution in the Canton. Based on archive documents, the different aspects of pharmacy in Canton Vaud at this epoch have been examined and analyzed. In particular, evolution of legislation and of professional training have been studied with reference to previous time periods or to the situation in other cantons of countries.The creation and development of professional associations has also been treated. One chapter is devoted to a brief description of the careers of more than one hundred pharmacists. The activities of the Cantonal Hospital pharmacy are also outlined, as well as the increase in the number of pharmacies within the context of economical growth in different regions of the Canton. Tables and addenda are used to support the discussions in the different chapters. This study should allow the present generation of pharmacists to have a better knowledge of the history of their profession and thus understand its evolution. PMID- 12117015 TI - Risk factors for hepatitis B surface antigen positivity among pregnant women. AB - A case-control study of risk factors for hepatitis B surface antigen (HBsAg) positivity was carried out among 130 pregnant women who were HBsAg positive (the case group) and 284 pregnant women who were HBsAg negative (the control group). Data obtained from the interviews and medical records including socio-demographic factors, personal history and behavioral factors related to HBV infection of the case and control groups were analyzed by using Odds ratio (OR), 95 per cent confidence interval of OR and chi2-test. The results revealed that the significant risk factors for HBsAg positivity were (a) a history of jaundice, OR=3.83 (p=0.0044), (b) tattooing, OR=3.98 (p=0.0411), (c) a history of jaundice in husbands, OR=7.93 (p<0.0001), (d) sharing articles with their husbands, such as a toothbrush, a spoon or a drinking glass, OR=5.90 (p<0.0001), (e) duration of marriage more than 4 years, OR=1.58 (p=0.0446) and (f) average sexual relations > or = 2 times per week, OR=2.12 (p=0.0007). The HBV preventive program should emphasize not only HBV vaccination in spouses of HBV carriers or pre-marital couples, but also health education for improving personal hygiene and sexual behavior in these target groups. PMID- 12117014 TI - Antibody response to hepatitis B immunization in infants born to HIV-infected mothers. AB - OBJECTIVES: To determine the antibody response of hepatitis B immunization begun at birth in HIV-1 exposed infants. DESIGN: Prospective, clinical trial. SITE: King Chulalongkorn Memorial Hospital, Bangkok, Thailand. MATERIAL AND METHOD: Seventy six infants born to HIV-1 seropositive mothers, who were not hepatitis B carriers, received three 10 microgram doses of recombinant DNA hepatitis B vaccine (Engerix B, Smith Kline, Belgium) in a 0, 1 and 6 month schedule. The first dose was given at birth. Serum hepatitis B surface antibody (Anti -HBs) was measured at age 3, 9 and 12 months. Anti-HBs levels were determined by enzyme linked immunoassay using the commercial kits (AUSAB EIA diagnostic kits, Abbott Laboratories, Chicago, USA) Antibody titer > or = 10 mIU/ml was defined as seroconversion. HIV infection was diagnosed by a positive test of HIV antibody at age > or = 18 months and/or by positive test of HIV polymerase chain reaction at age > or = 3 months. RESULTS: There were 14 HIV-1 infected (group 1) and 62 HIV-1 non infected (group 2) infants enrolled in this study. Anti-HBs titers of group 1 infants were significantly lower than those of groups 2 infants at both 3 and 6 months after the 3rd dose of vaccine, (Mann Whitney U test, p=0.019 and 0.001 respectively). Ten infants in group 1 and 57 infants in group 2 had anti-HBs titer > or = 10 mIU/ml. Their peak antibody titers were also noted at both 3 and 6 months after the 3rd dose of vaccine. Seroconversion rates were 71.4 per cent and 91.9 per cent in group 1 and 2 infants respectively, (p<0.05). Among the infants who had blood tests performed at age 12 months or 6 months after the 3rd dose of vaccine, anti-HBs titers declined in approximately 50 per cent of both groups of infants. There was a significantly higher percentage of seroconverters in group 1 who lost their protective titers than those in group 2, (p<0.001). CONCLUSION: The results in this study suggested that HIV-1 infected infants have poor antibody response to hepatitis B immunization and the protection was less durable. A fourth dose of vaccine at 6 months after the 3rd dose may be necessary. PMID- 12117016 TI - Associated genitourinary tract anomalies in anorectal malformations: a thirteen year review. AB - Congenital anomalies in the genitourinary tract are the leading associated anomalies in infants with anorectal malformations (ARM). Certain anomalies such as vesicoureteric reflux (VUR) may cause permanent renal damage. OBJECTIVE: To review associated genitourinary tract anomalies in-cases of anorectal malformations and evaluate the efficacy of ultrasonography in detecting VUR. MATERIAL AND METHOD: Retrospective review of 183 patients with ARM undergoing anoplasty between 1988-2001. RESULTS: Genital anomalies were found in 14 per cent (26 cases). Urologic anomalies were detected in 25.6 per cent (47 cases), with a higher incidence in supralevator anomalies. The most common upper tract anomaly was hydronephrosis, which resolved spontaneously in most of them. VUR was found in 16 cases (21 refluxing units) or 20 per cent of patients to whom voiding cystourethrogram (VCUG) was done. Sonography detected hydroureter and/or hydronephrosis in 3 of 21 refluxing units, despite 17 of them being grade three or more. Half of the cases with reflux had urinary tract infection at least once in the follow-up period despite normal initial urinalysis. Parenchymal scar was positive in four cases with VUR. CONCLUSION: Thorough evaluation of the urinary tract is necessary in infants with anorectal malformations. Ultrasound is an accurate tool in the examination of the upper tract, but not sensitive enough to detect lower tract anomalies, especially VUR. PMID- 12117018 TI - The ability of primary health care physicians to detect mental disorders in a university hospital setting. AB - OBJECTIVE: To determine how primary health care physicians differ in their ability and the factors underlying the differences between them in the ability to recognize mental disorders. METHOD: The group studied consisted of 15 primary health care physicians detecting mental disorders in 750 randomly selected adult patients of the general practice clinic in Thammasat University Hospital. The GHQ 28-Thai version was used as the reference method in the identification of psychiatric cases compared with the physicians' own assessment. Univariate and multivariate statistical analysis were used. RESULTS: There was a great variation in the ability of the physicians to detect mental disorders. The recognition ability was associated with the medical school from which the physician graduated. The discrimination ability was not associated with any factors. CONCLUSION: The medical school plays a role in determining the ability to detect mental disorders. This information would usually inform future developments in psychiatry teaching at both undergraduate and postgraduate levels. PMID- 12117017 TI - Efficacy and safety of atorvastatin 10 mg every other day in hypercholesterolemia. AB - OBJECTIVES: The authors sought to evaluate the safety and efficacy of atorvastatin administered every other day in patients with hypercholesterolemia. BACKGROUND: Statins have efficacy in lowering cholesterol and reducing cardiovascular events but their cost is a major disadvantage. Atorvastatin is the most potent statin and has a long half-life. Therefore, atorvastatin given on alternate days may be reasonable and cost effective, particularly in hypercholesterolemia patients. METHOD AND RESULT: Sixty patients with hypercholesterolemia despite diet therapy were enrolled into the study. They received atorvastatin 10 mg every other day before bedtime. Duration of treatment was 8 weeks. A lipid profile was determined as baseline, at 4 weeks and again at 8 weeks. Atorvastatin every other day significantly reduced total cholesterol (TC), triglyceride (TG), and LDL-c versus baseline. The TC, TG, and LDL-c levels were lower by 23 per cent, 8 per cent, and 30 per cent. Increase in HDL-c level was not statistically significant. Three patients had drug side effects. One patient had increased serum transaminase and one patient had increased serum muscle enzyme. The other one had somnolence. CONCLUSIONS: In hypercholesterolemia patients, atorvastatin 10 mg every other day is safe and effective in lowering TC, TG, with LDL-c and a slight increase in HDL-c. PMID- 12117019 TI - Self-assessment of sexual maturation in Thai children by Tanner photograph. AB - Assessment of sexual maturation in adolescence is crucially important in clinical practice, because the possibility of concealed disease during the earlier period of child development needs to be detected. However, to undress children sometimes is problematic and unethical in several countries. Therefore, in this study the authors evaluated the concordance between children's self-assessment by Tanner photograph with a written description, and examination by pediatricians. One hundred and ninety four children (100 girls, 94 boys), aged 7-15 years, were recruited in this study. The outcome demonstrated that the two processes of investigation were in good concord: the weight kappa of 0.76 and 0.79 for the breast stage (B) and pubic hair stage (PH), respectively, in girls. In boys, the weight kappa were 0.59 and 0.73 for genital stage (G) and pubic hair (PH) stage, respectively. However, the weight kappa for genital stage was improved after the subjects were provided more time to examine themselves before choosing the photograph. In girls, the mean chronological age (CA) for B stage II, III, IV was 10.4+/-1.3, 12.5+/-1.3, 13.6+/-0.7 years and for PH stage II, III, IV was 11.9+/ 1.4, 12.9+/-1.2, 13.5+/-0.9 years. The normal CA for the onset of puberty in girls was between 7.8-13.0 years. In boys, the mean CA for G stage II, III, IV was 11.3+/-1.7, 12.4+/-1.2, 13.2+/-1.2 years, and for PH II, III, IV was 12.2+/ 1.3, 13.1+/-1.0, 13.9+/-1.7 years. The normal CA for the onset of puberty in boys was 7.9-14.7 years. In addition, the authors constructed the normal value for penile length in Thai boys, aged between 9-15 years, to be used as a reference. Therefore, this study demonstrated a good concordance between the self-assessment by Tanner photograph and examination by pediatricians. This can be applied for use in filed research, school screening and clinical practice. PMID- 12117020 TI - A comparative study of diagnostic tests for tuberculous lymphadenitis: polymerase chain reaction vs histopathology and clinical diagnosis. AB - To compare diagnostic tests for tuberculous lymphadenitis by polymerase chain reaction (PCR) with histopathology and clinical diagnosis in sensitivity, specificity and predictive value. This retrospective analytic, single blind study was done at King Chulalongkorn Memorial Hospital. Paraffin-embedded specimens were classified into 2 groups. The study group contained 30 proved AFB positive paraffin-embedded specimens from patients who also had clinical diagnosis of tuberculosis and improved by antituberculous treatment. The control group contained 30 formalin-fixed, paraffin-embedded specimens of lymph node hyperplasia proved by histopathological and clinical review. All 60 specimens were slided, and systematically labeled and sent to PCR lab. Polymerase Chain Reaction method had sensitivity = 43.33 per cent, specificity = 100 per cent, positive predictive value = 100 per cent and negative predictive value = 63.83 per cent. The present findings revealed that the PCR results were related to the age of the paraffin-embedded tissues. No positive results were obtained from tissues kept since 1996. Positive results were obtained in 3/7 cases (42.86%), 2/3 (66.67%) and 8/10 cases (80%) from tissue of 1997, 1998 and 1999 respectively. CONCLUSION: Polymerase chain reaction has sufficient reliability best as a confirmatory diagnostic test for tuberculous lymphadenitis; however, it is not appropriate as a screening test. PMID- 12117021 TI - Association between estrogen receptor concentration in breast cancer tissue and bone mineral density. AB - Both bone and the breast are major target tissues of estrogen actions. The biological actions of estrogen depend on the interaction between estrogen and estrogen receptors (ER) in the target tissues. Therefore, ER concentration in tissues such as breast cancer might be associated with the amount of bone mass. The present study was aimed to examine whether there is a relationship between ER concentration in breast cancer tissue (ER-BCA) and bone mineral density (BMD). Forty-seven pre-menopausal and 34 post-menopausal women with newly diagnosed breast cancer were studied. The ER-BCA ranged from 0 to 339 fmol/mg cytosol protein (mean +/- SD = 68.6 +/- 97.0). Pearson's correlation analyses showed that ER-BCA negatively correlated to BMD of the spine (r = -0.251, p = 0.024), forearm (r = -0.341, p = 0.002), hip (r = -0.373, p = 0.001) and total body (r = -0.317, p = 0.004) in all 81 women. In 47 pre-menopausal women, the ER-BCA negatively correlated to the hip (r = -0.455, p = 0.001) and total body (r = -0.395, p = 0.006) but not to the spine and forearm BMD. Whereas, in 34 post-menopausal women, the ER-BCA negatively correlated to forearm BMD (r = -0.399, p = 0.019). Stepwise multiple regression analyses showed that the ER-BCA independently correlated to hip BMD in all 81 women (r = -0.373, p < 0.01) and in pre menopausal women (r = -0.486, p < 0.001) and independently correlated to forearm BMD in post-menopausal women (r = -0.399, p < 0.05). The results of this study suggest that the presence of high estrogen receptor concentration in breast cancer tissue might induce a deleterious effect on bone mass particularly in pre menopausal women. PMID- 12117022 TI - Outcome of Bell's palsy in children. AB - Acute idiopathic facial nerve paralysis (Bell's palsy) is a non life-threatening disorder but may cause important impact. In Thailand there has been no report of the outcome of Bell's palsy in children. Eighty four children with facial palsy were treated at Prasat Neurological Institute from January 1996 to July 2001. The etiology was found in 9 children (10.7%). Seventy five children were Bell's palsy. Twenty eight children were excluded, twenty two of these were loss to follow-up after the first visit and in six the onset were more than 30 days before presentation. Forty seven children remained for study. The mean age was 8.8 years (range from 2 years to 15 years 8 months). The male to female ratio was 1:1.1. The ratio of left to right side involvement was 1.3:1. Two children had recurrent facial palsy (4.3%). The duration from onset to recurrence was 6 months to 3 years 5 months. Oral prednisolone was given in 39 children. Complete recovery was observed in 29 children (61.7%) and almost complete recovery in 18 children (38.2%). All children recovered within 7 months. The mean duration of recovery was 6.61 weeks (range from 9 days to 28 weeks). The outcome of children aged under and over six years was not statistically different. The outcome of early and late steroid treatment could not be compared in this study. PMID- 12117023 TI - Is there a difference between the management of grade 2b and 3 corrosive gastric injuries? AB - OBJECTIVE: To evaluate the differences between treatment of patients with grade 2b or grade 3 gastric injuries. SETTING: A University Hospital. DESIGN: Retrospective review. PATIENTS: Thirty patients who ingested caustic agents over a 5 year period were examined by endoscope within 48 hours of injury. RESULT: Twenty one patients ingested strong acid or alkali. Among these patients, five had grade 2b, and two had grade 3 injuries. In both cases of grade 3 injuries, extensive surgical approach was initially performed, then delayed jejunal and colonic interpositions were done. On the other hand, one 2b patient had exploratory laparotomy, while others were treated conservatively. All 2b patients had satisfactory conditions during the initial follow-ups. Three patients were healthy during the 11, 16, and 44 months follow-up, one developed chronic gastritis at 5 months and one patient failed to follow-up. CONCLUSION: Early and aggressive extensive removal of necrotic tissue is necessary and can certainly increase the survival. There is on going controversy in the management of injuries less than grade 3. Grade 2b gastric injury patients can be managed conservatively. PMID- 12117024 TI - Comparative study between multiple and single rubber band ligation in one session for bleeding internal, hemorrhoids: a prospective study. AB - OBJECTIVE: The aim of this study was to compare the cessation of bleeding and the complications between multiple and single ligation using high ligation technique. MATERIAL AND METHOD: All first-visit patients with bleeding internal hemorrhoids were studied and randomly divided into multiple and single ligation groups. High ligation technique was used. Patients visited the clinic in the second week and were invited to visit the clinic or completed questionnaires after one year. RESULTS: 109 patients were included in the study. 61 patients had multiple ligation and 48 patients had single ligation. The cessation of bleeding in one week occurred in 96.7 per cent of patients in the multiple group and 79 per cent of patients in the single group (p = 0.004). There were no differences between the multiple group and single group concerning postligation pain and tenesmus (6.5% vs 2%, p = 0.532), urinary hesitancy and frequency (6.5% vs 4%, p = 0.904), and rebleeding in one year (27.9% vs 34%, p = 0.710). No major complications such as massive bleeding and pelvic sepsis were noted. CONCLUSIONS: Multiple ligation of bleeding internal hemorrhoids in one session can stop bleeding better than single ligation with no more complications. PMID- 12117025 TI - Ureterocalicostomy for reconstruction of complicated ureteropelvic junction obstruction. AB - OBJECTIVE: To review our experience with ureterocalicostomy using the treatment of complicated ureteropelvic junction (UPJ) obstruction. MATERIAL AND METHOD: Medical records of all patients with complicated ureteropelvic junction obstruction treated by ureterocalicostomy from 1985 to 2000 were reviewed. Causes of UPJ obstruction, surgical techniques, peri-operative course and outcome were noted. RESULTS: Fifteen patients were enrolled in this study (6 males and 9 females) with the mean age of 39 years old (28-45). Twelve patients were after stone surgery, 2 were after pyeloplasty for congenital UPJ obstruction and one patient was after blunt abdominal trauma. All the procedures were done by flank incision. After excision of the lower pole, the ureter was anastomosed to the lower caliceal mucosa without tension over an internal stent. Nephrostomy tubes were used in all of the patients. The mean hospital stay was 14 days (10-20). Twelve cases (80%) were found to be successful and are still doing well with the mean follow-up time of 2.5 years (0.5-12). Three patients (20%) were found to have failed, and subsequently nephrectomy was done in one case and permanent nephrostomy was used in 2 cases due to a solitary kidney. CONCLUSION: Ureterocalicostomy is one of the options for treatment of complicated UPJ obstruction that can provide good drainage as well as excellent long term results. PMID- 12117026 TI - Benign prostatic hyperplasia in elderly Thai men in an urban community: the prevalence, natural history and health related behavior. AB - PROBLEM: Although benign prostatic hyperplasia (BPH) is a common disease in elderly Thai men the prevalence in the community and its natural history is unknown. OBJECTIVES: To determine the prevalence of symptomatic BPH and its natural history. To determine the health related behaviors which have an impact on the voiding symptoms. MATERIAL AND METHOD: 879 elderly men aged > or = 60 years from communities around Siriraj Hospital were studied. The International Prostate Symptom Scores (IPSS) and Quality of Life (QOL) scores were evaluated in the participants at the beginning of the study and 1 year later. The overall assessment, complications and health related behaviors were also evaluated. RESULT: The prevalence of symptomatic BPH in the community was 41.3 per cent. In terms of overall assessment at 1 year follow-up, symptomatic BPH patients (IPSS 8 35), the rate of "improved", "same" and "worse" was 10.6, 70.2 and 19.2 per cent respectively. The complication rate was about 10 per cent. Three quarters of the elderly men had health related behavior at risk. CONCLUSION: The prevalence of symptomatic BPH was high. Its natural history was unpredictable and some BPH symptoms may be influenced by their behaviors. PMID- 12117027 TI - Good doctor: what the good and bad attributes are. AB - This study aimed to find the good and bad attributes of a doctor from people besides doctors themselves. MATERIAL AND METHOD: One thousand, one hundred people were asked to complete a 120 item questionnaire on the good and bad attributes of doctors in Maharaj Nakorn Chiang Mai Hospital. The sample groups comprised of patients, patients' relatives, and hospital personnel. RESULTS: Forty-four per cent of them responded. The top three good attributes were; to have good knowledge, to have a good rapport and to deal with a patient carefully. The top three bad attributes were; to be immodest in handling a female patients, being deceptive, and neurotic or having a psychiatric problem. Some good and bad attributes were different among different groups. DISCUSSION: Comparison with other studies was discussed including the limitation and application of this study. PMID- 12117029 TI - Intra-abdominal abscess in Crohn's disease: a case report. AB - Inflammatory bowel disease is uncommon in Thailand. The authors report a case of Crohn's disease in a 47-year-old Thai female. The patient presented with a three week history suggestive of an appendiceal abscess requiring an operation. The intra-operative findings of a lesion involving the terminal ileum and cecum, necessitated a right hemicolectomy to rule out reliably the presence of malignancy. Pathologic examination of the specimen suggested Crohn's disease. PMID- 12117028 TI - Iron bioavailability in Thai diets. AB - Dietary low iron bioavailability intake is an important causation factor of iron deficiency anemia in Asian countries including Thailand. The aim of this study was to estimate the iron bioavailability in the Thai diet by a calculation method that is based on dependent factors, dietary components and physiological iron store. Based on the the latest national nutrition survey of the Thai diet, 1995, the data of nutrient intake per capita per day by region were used for calculating the iron bioavailability at physiological iron store levels; 0, 250, 500 and 1,000 mg of iron. The results showed that the diets consumed by the populations in the Central, North, Northeast and South of Thailand were classified under the calculation method as being of moderate nonheme iron availability. The per cent iron bioavailability values of the Thai diets were within the range 3.7-12.4 per cent of total iron, depending on physiological iron store. The values of all region Thai diets at each iron store level were similar. By the same method, the dietary iron bioavailability of the total Thai diet at any iron store level was markedly lower than the general US diet, which was classified as high nonheme iron availability. When comparison of the iron bioavailability among other different diets was carried out, the values of the total Thai diet were slightly lower than Utah, but higher than US vegetarian and Regional Latin American diets. PMID- 12117030 TI - Correction of truncus arteriosus using a fresh autologous pericardial trileaflet valve conduit. AB - A fresh autologous pericardial trileaflet valve conduit was used in a 5-month-old infant for correction of truncus arteriosus. The patient recovered from the operation with satisfactory hemodynamics and post-operative echocardiogram at 3 months showed only mild to moderate pulmonary regurgitation. This technique is a useful alternative for correction in an infant with congenital heart disease who needs a tissue valve conduit. It may be more suitable than an aortic homograft by reason of unavailability of small homograft and limitation of organ donation. PMID- 12117031 TI - Streptococcus agalactiae meningitis in adults: report of two cases. AB - Two cases of Streptococcus agalactiae meningitis in adults are reported. The first patient was a 40-year-old man who presented with acute fever, headache, stiffness of the neck and confusion. During treatment, he developed left hemiparesis from cerebral infarction and bilateral deafness. The other was an 80 year-old man who presented with acute confusion and stiffness of the neck. During treatment, he developed septic shock and generalized tonic-clonic convulsions. Diagnosis was established by latex agglutination of streptococcus B-antigen and confirmed by cerebrospinal fluid-culture later on. The first patient survived but continued to have deafness whilst the other died from septic shock. PMID- 12117032 TI - Salmonella neck abscess. AB - Salmonella neck abscesses have rarely been reported in the world literature. Two patients with underlying diabetes mellitus developed deep neck abscesses which did not respond to empirical antimicrobials. Diagnosis of salmonella infection was made by culture of the discharge. Successful treatment was obtained by prescribing appropriate antibiotics and proper drainage. PMID- 12117033 TI - Botulinum toxin injection for objective tinnitus from palatal myoclonus: a case report. AB - Objective tinnitus may be caused by many etiologies-palatal myoclonus being one of them. We report one patient of voluntary palatal myoclonus presenting with objective tinnitus treated with botulinum toxin injection. Five units of botulinum toxin A were injected into each side of the soft palate at the palatal muscles (levator veli palatini and tensor veli palatini muscle). The tinnitus disappeared within two days of injection and no side effect was observed. PMID- 12117034 TI - Quality of life and functional parameters in patients with chronic obstructive pulmonary disease (COPD): an update. PMID- 12117035 TI - Survey of patients' views of domiciliary nebuliser treatment for chronic lung disease. AB - There is some controversy amongst respiratory physicians over the value of domiciliary nebuliser use for chronic lung conditions. Most recommendations for assessment of suitability for this form of treatment rely upon response to lung function tests and reported improvements in exercise ability. Relatively little emphasis has been placed upon the patient view of this therapy. This survey examined the subjective views of patients receiving domiciliary nebulisers regarding this treatment. A postal questionnaire was sent to 82 patients using home nebuliser treatment provided by the respiratory clinic at Whipps Cross University Hospital, London. It consisted of 29 structured questions covering topics of well-being and symptom control, self-confidence, dependency, time and technical issues, as well as side effects and compliance. Most patients surveyed had chronic obstructive lung disease. For almost all sections of the questionnaire patients reported overwhelmingly that the benefits of using a nebuliser outweighed potential disadvantages. The main perceived advantages werethe ability for patients themselves to control symptoms and to be less dependent on General Practitioners, hospitals and carers. Compliance was generally excellent, and the reported side effects were minor and relatively infrequent. The results strongly support the view that nebulisers are helpful in managing chronic lung disease in the community with benefit to patient well-being and potential health cost savings. PMID- 12117036 TI - Indoor women jobs and pulmonary risks in rural areas of Isfahan, Iran, 2000. AB - The prevalence rate of chronic airway's diseases in women and associated risk factors in developing countries are not well clarified. We evaluated the role of indoor duties in the prevalence of asthma, chronic bronchitis, and related symptoms among females in Isfahan suburbs. In three randomly selected villages, 561 responder females (response rate=95.25%) were evaluated through medical interview and physical examinations. Symptoms, signs, occupational and smoking histories, indoor and farming duties, possible outdoor jobs, housing and farming conditions were assessed. The observed respiratory morbidities were: current asthma (11.2%), history of asthma (1.3%), using asthma medications in life (15.2%), chronic bronchitis (3.4%), exercise-induced dyspnea and/or cough (16.2%), and frequent night coughs and/or dyspnea (15.3%). Age, childhood pulmonary infection, bread baking, carpet weaving and using biomass fuels were significant risk factors for all the pulmonary morbidities (P<0.05 to <0.001). Poultry feeding, using kerosene and gas fuels, were less strong risk factors for asthma and chronic bronchitis, respectively Only seven women were current or ex smokers. Indoor respirable particulate matters were two to four folds more concentrated than outdoors. Women doing indoor jobs in Iran are potential risk factors for development of chronic obstructive pulmonary disease. PMID- 12117037 TI - Lung deposition of budesonide from the novel dry powder inhaler Airmax. AB - The deposition of budesonide at fast (60 l min(-1)) and slow (301 min(-1)) inspiratory flow rates from Airmax, a new multi-dose dry powder inhaler, was compared with that from Turbuhaler and a standard pressurized metered dose inhaler (pMDI). Twelve patients with mild to moderate asthma took part in a five way randomized crossover study, and inhaled a single nominal dose of 200 microg budesonide, labelled with 99mTc, on each study day. Deposition was determined by gamma scintigraphy. At the fast flow rate, Airmax and Turbuhaler deposited 25.8+/ 6.5% (mean +/- sD) and 29.8+/-6.9%, respectively of the delivered dose in the whole lung (P = 0.080). At the slow flow rate, Airmax deposited 28.3+/-5.6%, Turbuhaler 22.7+/-5.6% and pMDI 12.1+/-3.4%. Using data on emitted doses determined in vitro, it was estimated that Airmax deposited 53.1+/-13.3 microg and 43.6+/-8-6 microg budesonide in the lungs at 60 l min(-1) and 30 l min(-1) respectively whilst Turbuhaler deposited 48.3+/-11.2 microg at 60 l min -and 24.2+/-6.0 microg at 30 l min(-1). In conclusion, lung deposition of budesonide from Airmax was comparable to that of Turbuhaler at a high flow rate but was markedly superior to Turbuhaler and pMDI at a lower flow rate. Unlike Turbuhaler, Airmax performs with relative flow-rate independence. PMID- 12117038 TI - Delivery of formoterol from a novel multi-dose inhaler Airmax. AB - Using a proprietary technology known as the X-ACT system--Active-metering, Cyclone-separator Technology, a novel multi-dose inhaler (Airmax) was developed to provide accurate and consistent dosing and a high-fine particle fraction ofthe drug. Formoterol, present as a blend with lactose monohydrate was delivered from Airmax to obtain a nominal formoterol dose of 6 or 12 microg. The devices were tested using a five-stage liquid impinger and a unit dose sampling apparatus, operated under conditions specified in European Pharmacopoeia (2000). Fine particle dose (FPD) was defined as the dose of the aerosolized drug particles with an aerodynamic diameter < 5 microm and fine particle fraction (FPF) was the ratio of FPD to the total recovered dose. Dose per actuation was found to be 97.0+/-11.5% label claim (LC) or 5.8+/-0.7 microg (n = 140), and 100+/-9.4% LC or 12+/-1.1 microg (n=440), for the 6 and 12 microg strengths, respectively. The mass median aerodynamic diameter was 2.4+/-0.1 microm (n = 14), the geometric standard deviation 2.1+/-0.1 (n = 14), and FPF 44.4+/-24% (n= 14) for both strengths. Thus, the combination of active metering and cyclone separator produces highly consistent doses of formoterol that have a large respirable fraction. PMID- 12117039 TI - Delivery of salbutamol and of budesonide from a novel multi-dose inhaler Airmax. AB - A novel multi-dose inhaler has been developed to closely approach the characteristics of an "ideal" inhaler. The new device, Airmax, uses proprietary technologies known as the X-ACT system to provide accurate and consistent dosing and excellent lung deposition--even at low inspiratory flow rates--combined with ease of use by the patient. Dose delivery was close to label claim, with relative standard deviation of typically around 5% for through-life emitted mass and around 10% for dose per actuation. At a flow rate (60-70 l/min), which corresponds to 4 kPa pressure drop across the device, the mean fine particle (<5 microm) dose (FPD) from 100, 200 and 400 microg strength budesonide Airmax was around 46, 98 and 244 microg, respectively. The mean FPD from 100 microg strength salbutamol Airmax was approximately 50 microg at the same flow rate. At 30 l/min, the delivered dose from Airmax is over 85% label claim with fine particle fraction of over 35%. Performance was unaffected by shaking or orientation, provided the device was not used completely upside down, and priming was not required. There was no change in dose content uniformity and aerodynamic particle size distribution after the devices have been stored unwrapped at 30 degrees C/60% RH up to 24 months. Airmax is robust, portable and intuitive to use. PMID- 12117041 TI - Up-regulation of heme oxygenase-I in alveolar macrophages of newly diagnosed asthmatics. AB - Exhaled carbon monoxide (CO), which has been found to be elevated in asthma, is generated primarily by heme oxygenase I (HO-I), an enzyme induced by oxidant stress and cytokines. The aim of this study was to assess the distribution and expression of HO-I in various human lung cells in acute and stable asthma. Normal lung tissue biopsies (from 6 non-smoking subjects operated on for a lung tumour) and macrophages from induced sputum (from 5 healthy controls, 5 untreated asthmatics, 7 stable treated asthmatics and 5 asthmatics recovering from exacerbation and being on systemic steroids) were investigated for HO-I by immunohistochemistry. The time response of HO-I induction was examined in cultured monocytes, which are known to maturate into monocyte-derived macrophages in culture. Lung biopsies showed prominent HO-I immunoreactivity only in alveolar macrophages. Macrophages in the induced sputum of healthy controls showed no HO-I immunoreactivity, with the exception of one case. Moderate or intense HO-I immunoreactivity could be observed in alveolar macrophages in 4/5 cases with recent asthma, and 2/7 with stable asthma, but in none ofthe patients treated with systemic corticosteroids for acute exacerbation. Experiments with cultured cells revealed that HO-I was induced by oxidants within the first 24 h, but the induction was reversed during the next 48 h. HO-I is mainly expressed in alveolar macrophages of human lung. Macrophages of induced sputum show prominent but transient HO-I immunoreactivity, in untreated asthmatics, but not in asthmatics treated with corticosteroids. PMID- 12117040 TI - Relative therapeutic index between inhaled formoterol and salbutamol in asthma patients. AB - A double-blind, randomized crossover study in 28 asthmatic patients assessed the relative therapeutic index for inhaled formoterol and salbutamol. Pre-drug administration FEV1 (mean 2.08 l) was 49-93% of predicted and reversibility 16 82% after inhalation of salbutamol. Patients inhaled single doses of formoterol (Oxis) (4.5,18 and 54 microg, delivered doses) via Turbuhaler, salbutamol (Ventolin) (200 and 1800 microg) via pressurized metered dose inhaler (pMDI) and placebo at intervals of 48 h or more. Individual maximum FEV1 and minimum S-K+ were calculated. Relative local (maximum FEV1) and systemic (minimum S-K+) dose potencies, and their ratio, the relative therapeutic index, were estimated using a non-linear mixed effect model. The drug effects were well tolerated and dose dependent. A log-linear approximation was used to describe the bronchodilatory effect, whereas a sigmoid approximation was more apt to describe the decrease in serum potassium concentration. A bivariate dose-response model based on these principles was fitted simultaneously to all data. The mean relative therapeutic index between formoterol 4.5-54 microg given via Turbuhaler and salbutamol 200 1800 microg given via pMDI was estimated to be 2.5 in favour of formoterol; this trend was not statistically significant. PMID- 12117042 TI - Cost-benefit and cost-effectiveness analysis of self-management in patients with COPD--a 1-year follow-up randomized, controlled trial. AB - The aims were to explore the effects and health economic consequences of patient education in patients with COPD in a 12-month follow-up. Sixty-two patients with mild-to-moderate chronic obstructive pulmonary disease (COPD) were at our outpatient clinic randomly allocated to an intervention group or a control group. The intervention group participated in a 4-h schooling, followed by one-to-two individual nurse and physiotherapist consultations. Self-management was emphasized following a stepwise treatment plan. Effectiveness was expressed in terms of proportions in need of general practitioner (GP) consultations, patient satisfaction and utilization of rescue medication. Doctor visits, days off work, dispensed pharmaceuticals, hospital admissions, travel costs, educational and time costs were recorded. The control and intervention groups induced mean total costs of NOK 19900 and 10600 per patient, respectively. The results were robustto realistic changes in the assumptions upon which they were based. For every NOK put into patient education, there was a saving of 4.8. The NNE to make one patient independent of their GP was 1.7 (95% CI: 1.3--2.8) and associated with a concomitant saving of NOK 15 800. The corresponding NNE to make one person satisfied with their GP was 4.5 (95% CI: 2.9--10) and NOK 41900, respectively. A reduced need of 100 DDD of rescue medication was associated with a concomitant saving of NOK5600. We conclude that patient education of patients with COPD in a 12-month follow-up improved patient outcomes and reduced costs. PMID- 12117043 TI - Effect of long-term treatment with inhaled budesonide or theophylline on lung function, airway reactivity and asthma symptoms. AB - Asthma is characterized by inflammation of the airways and long-term treatment with inhaled glucocorticosteroids improve clinical control in patients previously treated with inhaled rescue beta-2 agonist. We investigated whether the dose of inhaled glucocorticosteroid was related to outcome compared with oral theophylline. Budesonide 800 microg bd, budesonide 200 microg bd, or theophylline (Theo-Dur 300 mg bd was given double-blind, double-dummy and randomized, in a parallel group design for 9 months; when therapy was stopped patients were followed for an additional 3 months. Forced expiratory volume in 1 sec (FEV1), bronchial reactivity and asthma symptom scores were assessed before entering the study and after 1, 2, 3, 5, 7, and 9 months of treatment and monthly after treatment was stopped. Eighty-five patients (38 females and 47 males) were enrolled in the study during 1 1/2 year. Withdrawal from the study due to exacerbations during the treatment period was significantly increased (P <0.01) in the theophylline group. After treatment was stopped more patients withdrew in the budesonide group. In the budesonide 800 microg bd group, FEV1 improved significantly after 1 months treatment (P <0.01) and persisted throughout the study period. In the budesonide 200 microg bd group, FEV1 improved slightly and reached significance (P=0.05) after 5 months of treatment. In the theophylline group, FEV1 was unchanged during the 9 months of treatment. In both budesonide groups, FEV1 deteriorated significantly (P<0.01 and P<0.02, respectively) after termination of study medication and reached pretreatment values during the first month. In the budesonide 800 microg bd group, the concentration of histamine causing a 20% fall in FEV1 (PC20) increased significantly (P<0.01) after 1 months treatment and increased further after 9 months (P<0.0001), equivalent to two doubling dilutions. In the budesonide 200 microg bd, group PC20 histamine significantly increased (P <0.005) after 2 months of treatment and remained constant; theophylline was unchanged. After treatment with budesonide 800 microg bd and 200 microg bd were stopped, PC20 decreased significantly (P<0.002 and P=0.05, respectively) within the first month. PC20 remained unchanged after theophylline was stopped. After budesonide 800 microg bd and 200 microg bd treatment, symptom severity decreased in a dose-related and highly significant manner (P < 0.00001 and P < 0.0001, respectively). With theophylline, asthma symptoms decreased slightly after 1 and 2 months treatment (P < 0.01 and P < 0.02, respectively) and when treatment was stopped no increase in asthma symptoms was evident. Oral theophylline slightly reduced airways symptoms and had no influence on FEV1 and PC20 histamine. Maintenance treatment with inhaled budesonide gave a dose-related reduction in airways obstruction, bronchial reactivity and asthma symptom severity. The efficacy of inhaled corticosteroid was superior to oral theophylline. PMID- 12117044 TI - Addition of an extra dose of salmeterol Diskus to conventional dose of salmeterol Diskus in patients with COPD. AB - Patients experiencing dyspnoea can request an additional dose of salmeterol during the dose interval for the control of their symptoms, although under treatment with salmeterol. In this study we have explored the effects on respiratory function of an additive dose of salmeterol Diskus in 15 chronic obstructive pulmonary disease (COPD) patients in regular treatment with a conventional dose of 50 microg salmeterol. On two different days, patients inhaled 50 microg Diskus. After 240 min, they inhaled additional 50 microg salmeterol Diskus (salmeterol arm) or placebo Diskus (placebo arm). Lung function was controlled before first drug administration and 0.5, 1, 2, 3, 4, 4.5, 6, 8, 10, and 12 h thereafter. The mean (95% CI) peak increase in FEV1 from baseline was reached after 4 h in the salmeterol arm (0.174 L; 0.144-0204) and after 5 h (0.141 L; 0.115-0.168) inthe placebo arm; after 12 h, the mean (95% Cl) increase in FEV1 from basal values was still 0.149 L (0.119-0.179) in salmeterol arm, but only 0.041 L (0.017-0.064) in placebo arm. The mean (95% CI) FEV1 AUC0-12h for all patients were 2.01 (1.72-2.30) L when salmeterol was added and 1.30 (1.03 1.58) L when placebo was inhaled. The difference (mean; 95% CI) between the FEV1 AUC0-12h of the two arms (0.71 L; 0.47-0.95) was statistically significant (P<0.0001), although the difference (mean; 95% CI) between the FEV1 AUC0-4h of the two treatments (0.08 L; -0.02-0.18) was notstatistically significant (P=0.126). The addition of an extra dose of salmeterol did not significantly increase the heart rate or decrease the SpO2. This study suggests that the addition of an extra dose of salmeterol does not give room for further increase in peak FEV1, but the effect of adding salmeterol to salmeterol is largely additive when considering the duration of action and safe. PMID- 12117046 TI - Relationship between patient and disease characteristics, and health-related quality of life in adults with asthma. AB - The purpose of this study was to examine relationships between patient- and disease-related variables and health-related quality of life (HQL). This cross sectional study surveyed adults with asthma enrolled in a managed care organization (MCO). Data were obtained from a mailed questionnaire and the MCO's patient and claims databases. The Asthma Quality of Life Questionnaire (AQLQ) and the SF-36 instruments were used. The behavioral Model of Health Services Utilization was used to characterize independent variables and their relationships to HQL. Independent variables included predisposing (age, gender, education, race, number of comorbidities, years with asthma, social support, health-belief questions); enabling (income, number of metered dose inhaler (MDI) instructors, perceived inconvenience of accessing the physician); and illness level (perceived and symptom-derived asthma severity). Multivariate linear regression models were developed to examine the relationships between the independent variables and the domain and summary scores of the AQLQ and the SF 36. The survey response rate was 63% (n=603). for the AQLQ, symptom-derived severity perceived severity education level, and the health-belief factor Barriers were significant in all five models. Symptom-derived severity had consistently higher standardized regression coefficients than perceived severity Barriers had the highest coefficient in all but the Symptoms domain model. Number of Comorbidities was significant in all eight domain and two summary score SF-36 models. Symptom-derived and/or perceived severity were also significant in all but the Mental Health domain model. Other frequently significant variables included the health-belief factor Barriers and Yearly Household Income. When assessing HQL of a population, such as this group of patients with asthma, one must consider patient and disease variables that may influence the results. PMID- 12117045 TI - The FEV1/FEV6 predicts lung function decline in adult smokers. AB - The use of FEV1/FEV6 in place of the traditional FEV1/FVC to detect airways obstruction during spirometry testing performed by primary care providers would reduce time and patient effort. We hypothesized that the FEV1/FEV6, would predict the subsequent decline in FEV1 in adult cigarette smokers who enrolled in the multicenter Lung Health Study. Ten clinical centers in the U.S. and Canada recruited 5887 male and female smokers, aged 35-60 years, with borderline to mild airways obstruction by spirometry. Those who successfully stopped smoking during the 5-yr study (usually as the result of the smoking cessation intervention) were excluded from this analysis. In those continuing to smoke, the relative strength of spirometric predictors of the change in FEV1 during 5 years of follow-up (DFEV1) was determined using a linear regression model. The following covariates were significant independent predictors of DFEV1: the baseline degree of airways obstruction, age, gender, cigarettes per day, years of education, and bronchial hyperresponsiveness. The FEV1/FEV6 was nearly as strong an independent predictor as was the FEV1/FVC (a traditional index of airways obstruction). The degree of airways obstruction, as determined by the FEV1/FEV6 from spirometry, is an independent predictor of subsequent decline in lung function; and therefore, may be used to detect smokers at higher risk of developing COPD. PMID- 12117047 TI - Cardiac output increases prior to development of pulmonary edema after re expansion of spontaneous pneumothorax. AB - Pulmonary edema following reexpansion of spontaneous pneumothorax is an uncommon complication. The underlying mechanism of this condition is unclear. We report the hemodynamic characteristics in a series of 7 male patients with spontaneous large (>50%) pneumothoraces of > or = 24 h and correlate the changes with reexpansion pulmonary edema (REPE). A pulmonary artery floatation catheter was inserted and hemodynamic data were obtained before therapeutic chest tube insertion, 1 h after chest tube insertion and the following day. Four (57%) patients developed REPE. There was a tendency for larger pneumothorax to develop REPE. Capillary wedge pressure did not change significantly 1 h after the insertion of chest tube in all our patients. Cardiac output increased significantly in patients who developed REPE compared to those who did not (+ 1.06 l/min vs -0.27 l/min; P = 0.03) 1 h after insertion of chest tube. One patient did not develop pulmonary edema despite having a large (> 80%) pneumothorax. His cardiac output did not rise 1 h after chest tube insertion. REPE is not an uncommon complication following chest tube drainage in patients with large and long-standing pneumothorax. The increase in cardiac output after chest tube insertion may be associated with subsequent development of REPE. PMID- 12117048 TI - Investigative bronchoscopy and endobronchial biopsy is well tolerated in hyperreactive asthma patients. AB - The tolerability of 57 non-smoking asthma patients inhaling salbutamol as needed (ATS, 18--60 years, 60% < or = FEV1 < or =100%, PD15FEV1 <0.4 mg histamine) to fibreoptic bronchoscopy (FOB) and endobronchial biopsy was studied. The FOB was done in local Lignocaine anaesthesia, and from five to eight biopsy specimens were taken from the bronchial mucosa of the right lung. The tolerability was measured as cough/bronchospasm during the procedure (from 0 = normal to 3 = interrupted procedure), success of the procedure, and untoward occurrences. Twenty-seven of the 57 patients (48%) had no cough or bronchospasm during the FOB (score 0). Few coughs of no importance (score 1) were documented in 23 patients (40%). Seven patients (12%) had cough and/or bronchospasm interfering with the FOB procedure (score 2). The FOB procedure was not interrupted because of cough and/or bronchospasm (score 3) in any patient. Scores of cough and/or bronchospasm diminished progressively with the increase of PD15FEV1 histamine. The success of the procedure was 100%. Two patients had untoward medical occurrences requiring additional rescue medication (3.5%). In conclusion, we found that hyperreactivity predicts cough and/or bronchospasm during the FOB. Cough and/or bronchospasm are frequently observed during the bronchial procedure, but they are mild and of minor clinical importance. An investigational endobronchial procedure can be successfully performed in mildly or moderately obstructive asthmatic patients, even in cases with severe bronchial hyperreactivity. PMID- 12117049 TI - Treatment of asthma patients with herbal medicine TJ-96: a randomized controlled trial. AB - Alternative medicine use has increased at a remarkable pace all over the world in recent years. Although herbal medicine for the treatment of asthma is becoming the focus of public attention, randomized studies had not been performed, even in Eastern countries including Japan. This study was designed to investigate whether one of the Japanese government approved herbal complexes Saiboku-to (TJ-96) is effective for the treatment of atopic asthma, and to investigate whether this protective activity is associated with a reduction in eosinophilic inflammation. A double-blind, randomized, crossover design was used. Subjects received 2.5 g of TJ-96 or placebo orally 3 times daily for 4 weeks and then, after a washout period of at least 4 weeks, crossed over to receive the alternative treatment. We assessed the effects of pretreatment with TJ-96 on bronchoconstriction precipitated by inhalation of methacholine. Furthermore, eosinophil counts and measurement of eosinophilic cationic protein (ECP) were performed. After 4 weeks of treatment with TJ-96, values of PC20 -methacholine significantly improved in the treatment with TJ-96. Also, patients' symptoms, blood eosinophils, serum ECP, sputum eosinophils, and sputum ECP were significantly decreased. Our results suggest that TJ-96 has an antiinflammatory effect on bronchial eosinophilic infiltration. This study raises further interesting therapeutic possibilities and argues for further trials of new approaches to the treatment of asthma. PMID- 12117050 TI - Environmental assessment of polycyclic aromatic hydrocarbons (PAHs) in surface sediments of the Santander Bay, Northern Spain. AB - Samples of intertidal surface sediments (0-2 cm) were collected in 17 stations of the Santander Bay, Cantabric Sea, Northern Spain. The concentrations of polycyclic aromatic hydrocarbons (PAHs), 16, were analysed by HPLC and MS detection. Surface sediments show a good linear correlation among the parameters of the experimental organic matter evaluation, where total carbon (TC) and loss on ignition (LOI) are approximately 2.5 and 5 times total organic carbon (TOC). A wide range of TOC from 0.08% to 4.1%, and a broad distribution of the sum of sigma16PAHs, from 0.02 to 344.6 microg/g d.w., which can be correlated by an exponential equation to the TOC, has been identified. A qualitative relationship may be established between the industrial input along the rivers and the concentration of sigma6PAHs in the sediments of the estuaries: Boo estuary (8404 4631 microg/g OC), Solia-San Salvador estuaries (305-113 microg/g OC) and Cubas estuary (31-32 microg/g OC). This work shows a dramatic change in the spatial distribution in the concentration of PAHs of intertidal surface sediments. The left edge of the Bay has the main traffic around the city and the major source of PAHs is from combustion processes and estuarine inputs, leading to medium values of PAHs in the sediments; the right edge of the Bay has much lesser anthropogenic activities leading to lower values of PAHs in sediments. The distribution of individual PAHs in sediments varies widely depending on their structure and molecular weight; the 4-6 ring aromatics predominate in polluted sediments due to their higher persistence. The isomer ratio does not allow any clear identification of the PAHs origin. Environmental evaluation according to Dutch guidelines and consensus sediment quality guidelines based on ecotoxicological data leads to the same conclusion, sediments in the Santander Bay show a very different environmental quality depending on the spatial position from heavily polluted/medium effects to non-polluted/below threshold effects. These results indicate that local sources of PAHs, especially estuary discharges, lead to very different qualities of sediments in coastal zones, where traffic and industrial activities take place. PMID- 12117051 TI - Levels and congener distributions of PCDDs, PCDFs and non-ortho PCBs in Belgian foodstuffs--assessment of dietary intake. AB - Congener-specific analyses of 7 polychlorinated dibenzo-p-dioxins (PCDDs), 10 polychlorinated dibenzofurans (PCDFs) and 4 non-ortho (coplanar) polychlorinated biphenyls (cPCBs) were performed on 197 foodstuffs samples of animal origin from Belgium during years 2000 and 2001. All investigated matrices (except horse) present background levels lower than the Belgian non-commercialization value of 5 pg TEQ/g fat. Pork was the meat containing the lowest concentration of both PCDD/Fs and cPCBs. The mean background concentration of 2,3,7,8-TCDD toxicity equivalent in milk was 1.1 pg/g of fat, with a congener distribution typical of non-contaminated milk. The relative contribution of 2,3,7,8-TCDD, 2,3,7,8-TCDF, 1,2,3,7,8-PeCDD and 2,3,4,7,8-PeCDF to the PCDD/Fs TEQ was 85+/-7.9% for all investigated matrices. The cPCBs contribution to the total TEQ was 47+/-19.0% for products of terrestrial species and 69+/-20.0% for aquatic species. Once the contribution of cPCBs was added to the TEQ, few foodstuffs such as horse, sheep, beef, eggs and cheese presented levels above the future European guidelines that currently only include PCDD/Fs but will be re-evaluated later in order to include 'dioxin-like' PCBs. Based on levels measured in the samples, the estimation of the dietary intake was 65.3 pg WHO-TEQ/day for PCDD/Fs only (1.00 pg WHO-TEQ/kg bw/day, for a 65 kg person) and 132.9 pg WHO-TEQ/day if cPCBs were included (2.04 pg WHO-TEQ/kg bw/day, for a 65 kg person). Meat (mainly beef), dairy products, and fish each account for roughly one third of the intake. PMID- 12117052 TI - Multipoint moss passive samplers assessment of urban airborne polycyclic aromatic hydrocarbons: concentrations profile and distribution along Warsaw main streets. AB - Polycyclic aromatic hydrocarbons (PAHs) distribution along 28 km of Warsaw main street have been surveyed in July 2000 using moss passive samplers as a simple and economic surrogate of direct air sampling. Altogether 74 samplers at 39 crossroads with traffic lights were placed on the lamp post approximately/=3.5 m above ground. PAHs levels determined in samplers are in range from 828 to 3573 ng/g moss dry weight. The spatial spread of pollution within this range is statistically close to normal distribution with mean value of 2332 ng/g. Variability within and between study areas are rationalized in terms of urban environmental factors. PAHs concentrations profiles across the town have appeared uniform. The dominant compounds are phenantrene, fluoranthene and pyrene. Their contribution is 49-68% of total PAHs burden. PMID- 12117053 TI - Environmental impact of a new hazardous waste incinerator in Catalonia, Spain: PCDD/PCDF levels in herbage samples. AB - In April 1996 and 1998, the concentrations of polychlorinated dibenzo-p-dioxins (PCDD) and polychlorinated dibenzofurans (PCDF) were determined in 40 herbage samples collected in the neighborhood of a hazardous waste incinerator (HWI) under construction in Constanti (Catalonia, Spain). In April 2000, 20 months after the HWI began operating, herbage samples were again collected at the same sampling points in which samples had been taken in the previous surveys. PCDD/F concentrations ranged between 0.13 and 0.65 ng I-TEQ/kg (dry matter), with a median and mean values of 0.29 and 0.32 ng I-TEQ/kg (dry matter), respectively. The results were compared with those obtained in the 1996 (median, 0.53 ng I TEQ/kg; mean, 0.61 ng I-TEQ/kg) and the 1998 (median, 0.23 ng I-TEQ/kg; mean, 0.31 ng I-TEQ/kg) surveys. While in the period 1996-1998 a significant decrease (49%, P < 0.001) in the mean PCDD/F levels was noted, in the period 1998-2000 an increase of 3% (P > 0.05) was found in the concentrations of PCDD/Fs. The analysis of the results suggests two potential hypotheses: either the emissions of PCDD/Fs from the HWI are not negligible, or the current PCDD/F emissions from other sources near the HWI remained at similar levels to those reached in 1998. Anyhow, an exhaustive evaluation of the present data shows an absence of notable PCDD/F contamination by the HWI in the area under its direct influence. It seems also probable that the decline in the atmospheric levels of PCDD/Fs due other emission sources of PCDD/Fs in this area is currently stopped. PMID- 12117054 TI - Uptake of injected PCBs from the yolk by the developing chicken embryo. AB - In this study, PCB uptake by the developing chicken embryo was measured after injection of two different doses of Aroclor 1254 before incubation. It was shown that 2% of the injected PCBs was absorbed on day 13, and this increased exponentially to 18% at day 19. This exponential increase could be described by a similar model for both low and high injection doses. Differences in injection dose resulted in corresponding differences in concentration in the embryos. Lipid corrected concentrations in the embryo were stable through development from day 13 up to day 19 and could be predicted from injection doses by using a conversion factor of 0.15 g(-1). PMID- 12117055 TI - Degradation characteristics of a dibenzofuran-degrader Terrabacter sp. strain DBF63 toward chlorinated dioxins in soil. AB - To obtain basic information towards applying a dibenzofuran (DF)-degrader Terrabacter sp. strain DBF63 to bioremediate dioxin-contaminated soil, we investigated the degradative potential of strain DBF63 for either chlorinated or polychlorinated dibenzo-p-dioxins and dibenzofurans (ClxDD/ClxDF) in soil. In the soil slurry system with a soil to water ratio of 1:5 (w/v), the DF-grown DBF63 cells degraded 90% of 1 ppm 2,8-Cl2DF, whereas only 40% of 1 ppm 2,3-Cl2DD during the 7-day incubation. The degradation rates of 2-CIDF, 2-ClDD, 2,8-Cl2DF and 2,3 Cl2DF by strain DBF63 in the soil slurry system (5-day incubation) were approximately 89%, 65%, 78% and 32%, respectively. These results suggest that strain DBF63 was able to degrade mono- to dichlorinated dibenzofurans more effectively than mono- to dichlorinated dibenzo-p-dioxins. Using the same soil slurry system, we performed a preliminary bioremediation experiment using the actual dioxin-contaminated soil at an incineration site. We found that approximately 10% of tetra- to hexa-chlorinated congeners was decreased by a single inoculation with DBF63 cells within a 7-day incubation. PMID- 12117056 TI - PCDD/F and metal concentrations in soil and herbage samples collected in the vicinity of a cement plant. AB - In May 2000, the levels of a number of metals (As, Cd, Pb, Hg, Zn, Co, Cu, Mn, Sn, Tl, Cr, Ni and V) were determined in 16 soil and herbage samples collected in the vicinity of a cement plant from Sta. Margarida i els Monjos (Catalonia, Spain). Metal concentrations were also analyzed in air filters from three sampling stations placed nearthe facility. For most metals, concentrations were similar or even lower than previously reported values for other areas from Catalonia. On the other hand, the levels of polychlorinated dibenzo-p-dioxins (PCDD) and dibenzofurans (PCDF) were also determined in four soil and 16 herbage samples. Mean values were 0.37 and 0.16 ng I-TEQ/kg for soils and herbage, respectively, values which in comparison with data from other surveys are rather low. No significant differences between metal and PCDD/F concentrations in samples collected at distances lower or greater than 3.5 km of the facility were noted. The current results show that the cement plant has a low impact on the metal and PCDD/F levels in the environment under direct influence of the facility. These results should be of interest to assess future temporal variations in the levels of metals and PCDD/Fs in this area. PMID- 12117057 TI - Degradation of organophosphoric esters in leachate from a sea-based solid waste disposal site. AB - Degradation of organophosphoric esters (OPEs) in leachate from a sea-based solid waste disposal site was investigated by laboratory experiment. Aryl-phosphates, tricresyl phosphate and triphenyl phosphate, in leachate rapidly decreased to less than the detection limit within 20 days under aerobic condition, suggesting high biodegradability. These phosphates also decreased in sterilized leachate, which suggested a contribution to degradation by reactions (adsorption and chemical degradation) with chemicals in the leachate. Concerning alkyl phosphates, tributyl phosphate decreased rapidly after one week, which is considered to have been caused by biodegradation. Tris-2-ethylhexyl phosphate and tris-2-butoxyethyl phosphate decreased slowly in all samples but that of sterilized distilled water. This however, suggested contribution of biodegradation because the velocity of decrease in the leachate was higher than in control samples. Among chloro alkylphosphates, decrease of tris-2-chloroethyl phosphate and tris-dichloropropyl phosphate were observed though it was not obvious whether by biodegradation or not. Decrease of tris-2-chloropropyl phosphate (TCPP) was not observed for 80 days suggesting that TCPP remains in the leachate over a long period of time. Except for aryl-phosphates decrease of OPEs was not observed under anaerobic condition. It was considered that the composition ratio and the behavior of OPEs in leachate in the field reflects the biological and chemical degradation as well as the chemical properties of OPEs. PMID- 12117058 TI - Electronic elasticity-toxicity relationships for polychlorinated dibenzo-p-dioxin congeners. AB - SCF-MO computations have been performed on tetra- to octa-chlorinated dibenzo-p dioxin congeners (PCDD) using an MNDO-PM3 Hamiltonian. Qualitative relationships were developed between empirical, international-toxic equivalence factors for PCDD congeners and their relative (specific) polarizabilities and mean values of second hyperpolarizabilities estimated using finite-field theory. PMID- 12117059 TI - Distribution and fate of organochlorine pesticide residues in sediments from the selected rivers in Taiwan. AB - The contamination of organochlorine pesticides (OCPs) in sediments from selected rivers in Taiwan was investigated to evaluate the pollution potentials and hazard in river sediments. Da-han River and Erh-jen River were selected as the target rivers due to their serious pollution. A total of 40 surface sediment samples were collected at five sampling stations along the rivers. Results showed that the concentrations of various pesticides in sediments were in the range of 0.57 14.1 ng/g for sigmaHCH, 0.05-0.15 ng/g for aldrin, 0.12-5.8 ng/g for dieldrin, 0.22-0.64 for endrin, 0.24-6.37 ng/g for endosulfan and 0.21-8.81 ng/g for EDDT (p,p'-DDD, p,p'-DDE, p,p'-DDT). Among the OCPs, sigmaHCH, endosulfan and sigmaDDT were the most dominant compounds in the river sediments. Endosulfan sulfate was the most frequent detected compound in the sediments from the selected rivers. Also, sigmaDDT, dieldrin and beta-HCH were in abundance. Different contamination patterns between the selected river sediments were also observed. Da-han River was mainly contaminated with endosulfan sulfate and sigmaDDT. Whereas the main pesticides in Erh-jen River were beta-HCH and sigmaDDT. Among the cyclodiene compounds, dieldrin was in abundance in most of the sediments. Moreover, the frequencies of detection of the metabolites were higher than those of parent compounds, depicting that the sediments have contaminated for a long time. The results obtained in this study showed that there still exist a variety of OCP residues in the river sediments in Taiwan. PMID- 12117060 TI - Levels and tissue-dependent distribution of dioxin in Japanese domestic leafy vegetables--from the 1999 national investigation. AB - In 1999, Japanese domestic leafy vegetables were successively investigated for levels of dioxins, including 17 dibenzo-p-dioxins/dibenzofurans (PCDD/Fs), four non-ortho co-planar PCBs (co-PCBs) and eight mono-ortho co-PCBs, all of which had been assigned toxic equivalency factors (TEFs) by WHO in 1997. The mean levels of dioxins in the edible portions were 0.07 (0.09) pg TEQ/g in spinach, 0.13 (0.14) pgTEQ/g in garland chrysanthemum, 0.01 (0.04) pg TEQ/g in mitsuba (marsh parsley) and 0.01 (0.03) pg TEQ/g in chingentsuai (Brassica Campestris var. chinesis), when non-detects were set to zero (and set to half the limit of detection). In order to understand the dioxin pollution of leafy vegetables in detail, a further investigation of dioxin levels in the tissues of spinach was conducted. As a result, the dioxin levels in the leaves were found to be higher than those in the stem and red collar, but they were much lower than those found in the primary and secondary roots, which are considerably affected by the soil, which is recognized as a sink of airborne dioxins. The dioxin levels in edible portions (leaves, stem and red collar) were obviously lower than those in non-edible portions (primary and secondary roots). In addition, from the finding that several lower chlorinated PCDD/Fs and co-PCBs, namely 2,3,7,8-TCDD/F, 1,2,3,7,8-PeCDD, 1,2,3,7,8/2,3,4,7,8-PeCDFs, 1,2,3,7,8,9-HxCDD, 1,2,3,4,7,8-/1,2,3,6,7,8-HxCDFs, 3,3',4,4'-TCB, 2,3,3',4,4'-/2,3',4,4',5-PeCBs, and 2,3',4,4',5,5'-HxCB, were more highly represented in the dioxins in the leaves than in those in the secondary roots, it was suggested that in leafy vegetables the deposition of gaseous, presumably moderately volatile dioxins in leaf wax is another pollution pathway in addition to the adhesion of dioxin-contaminated particles including soil. PMID- 12117061 TI - Alcohol consumption pattern among women in a rural Yoruba community in Nigeria. AB - Indigenous Nigerian societies discourage alcohol consumption among women, yet international trends show alcohol consumption increasing in populations of developing countries, especially among women. This research implemented in 1994, examined the pattern of alcohol consumption among women in the rural town of Igbo Ora, located in the southwestern state of Oyo in Nigeria. A majority of the 300 respondents (64%) were found to have tasted alcoholic beverages, and over half of these reported current alcohol use. Current drinkers reported consuming an average of 1.3 bottles (60 cl per bottle) of alcoholic beverage in the week preceding the survey. Current drinking status was associated with religion. Only 9% of the respondents with indigenous beliefs reported using alcohol, compared to 40% of Christian and 30% of Moslem respondents. Those who never drank were, on average, 5 years older than current or previous drinkers. Single, separated, or divorced women were more likely to drink than married or widowed women. Special uses of alcohol for women were identified, including easing the pains of childbirth. Furthermore, the respondents identified problems associated with drinking alcohol that women confronted: accidents, fighting, illnesses, mental problems, children learning to drink, child neglect, rape, and tarnishing of one's image. With less than one-third of women reporting that they are current drinkers, and among those weekly consumption being low, one could say that there is little evidence of alcohol misuse among women in Igbo-Ora. At the same time, the fact that current drinkers are younger implies that consumption rates may increase in the future. This information about women's beliefs, practices, and preferences will be of value in designing health education programs to prevent future alcohol-associated problems. PMID- 12117062 TI - Drug use and associated factors among rural adolescents in Costa Rica. AB - The objectives of this study, carried out in 1995, were to assess both licit and illicit substance use among rural male and female Costa Rican adolescents, and associated health, psychological, and psychosocial problems. A sample of 304 students from rural schools was randomly selected. The mean age for females was 14.7 years (S.D. = 1.71), and for males was 14.4 years (S.D. = 1.62). The data were collected using the Latin-American version of Drug Use Screening Inventory (DUSI). Results showed a high prevalence of past-year alcohol use for both males and females (56.6% and 47.4%, respectively), and a lower prevalence of past-year tobacco use (44.0% and 7.7%). There results also showed a low level of use of solvent inhalants and benzodiazepines. In terms of illicit drugs, males preferred cocaine and marijuana, while females only reported amphetamine use. An analysis of adolescent functioning showed differences among alcohol users and nonusers in behavior patterns and peer relationships. However, no significant differences were found regarding rebellion, depression, and social isolation. The implications of these results are discussed, along with the importance of enhancing prevention, as well as early detection and intervention. PMID- 12117063 TI - Prevalence of cigarette smoking among rural adolescents in the United States. AB - Results are reported from a national U.S. study of cigarette smoking carried out from 1996 to 2000 involving 68,270 adolescents. Hierarchical linear modeling was used to model smoking as a function of grade, gender, region, and community size (rurality). Significant effects were found for rurality, region, grade, and gender. The highest levels of smoking were found for rural adolescents, and adolescents living in the South. Males smoked more than females in all regions except the West, where the reverse was true. Given that rural adolescents smoke more "heavily" than do their nonrural peers, researchers must devote more attention to understanding the factors that underlie smoking initiation in rural youth. PMID- 12117064 TI - Drug-related offenses and the structure of communities in rural Australia. AB - This article examines the relationship of drug use with the social and economic characteristics of rural communities in New South Wales (NSW), Australia. Data is derived from the 1996 Australian Census of Population and Housing, and data on drug-related offenses from the NSW police between 1995 and 1999. Arrest rates for breaking and entering, assault, and vandalism showed statistically significant associations across types of rural communities, but drug-related arrests varied considerably less. The widespread, relatively-even distribution of drug arrests in rural NSW suggests that the underlying causes of drug-related violations are unique when compared to other types of crime. PMID- 12117065 TI - Review of drinking patterns of rural Arab and Jewish youth in the north of Israel. AB - This article reviews four studies addressing alcohol drinking patterns among rural Arab and Jewish youth. Three religions, Moslem, Druze, and Christianity, were represented among the Arab population studied. The Arab adolescents come from villages, Arab towns, and mixed Arab-Jewish towns, while the Jewish youth come from kibbutzim and developing towns in the northern district of Israel. The first epidemiological study among rural adolescents was implemented in 1990. This study focused on frequency of drinking during the previous month, and amounts of alcohol consumed on a drinking occasion. The 1992 study focused on preferred sources of support after acquiring a drinking problem, reasons for drinking, and the social context of drinking in the previous year. The 1994 study focused on reasons for not drinking, preferred places of drinking, and ways of obtaining alcoholic beverages. The 1996 study dealt with frequency of drinking in the last year, and amounts of alcohol consumed on a drinking occasion. This review also includes urban-rural comparisons. Urban adolescents were drawn from Haifa, the largest city northern Israel. PMID- 12117066 TI - Attitudes and values of Peruvian coca growers. AB - This study uses semi-structured interviews to examine the attitudes and values of Peruvian coca growers toward coca leaf and cocaine basic paste (CBP) consumption and its distribution. The subjects of the study were 186 coca growers from Peruvian jungle valleys who are involved in illegal commercialization of coca leaf and cocaine paste production. Data collected in 1994 reveal that growers consider coca leaf to be a most profitable product and a unique opportunity to improve their quality of life. Although growers acknowledge that a problem exists among local users, they do not assume any responsibility for CBP consumption and dissemination in rural areas. This leads to the conclusion that awareness of a CBP consumption problem is not enough for growers to stop drug production; they need consistent training in social values, as well as support in legal and economic alternatives. PMID- 12117067 TI - Social cohesion, cultural identity, and drug use in Mexican rural communities. AB - The objective of this study was to explore drug use in Mexican rural communities and its relationship to social cohesion, cultural identity, migration, and transculturation. Community models typification was used, considering cohesion as the central point of analysis. The research was conducted during 15-day periods in each of nine communities during 1991. Both documentary and ethnographic techniques were used to gather information. Results indicated that rural communities where there was little or no drug use among its members show more social cohesion, cultural identity, and community links consolidation, and more capacity for integrating change. This pattern is most apparent among young community members who have had more contact with the outer world (drug trafficking, North American culture, and Mexican urban culture). PMID- 12117068 TI - History, culture, and substance use in a rural Scottish community. AB - This paper provides a detailed discussion of substance use and misuse in a rural community in the Western Highlands of Scotland, United Kingdom. Attention is focused on the way in which patterns of substance use arise from a complex interplay of historical, cultural, social, and personal events. The discussion illustrates how large changes in patterns of intoxication in rural communities can be rendered intelligible through an understanding of the impact of economic, religious, and social changes. The analysis is based on an historical and ethnographic account, carried out between 1987 and 1990 with adolescents, of patterns of use which range from pagan ceremonies in the 16th century to contemporary "soft drug" use. PMID- 12117069 TI - Drug use and AIDS: estimating injection prevalence in a rural state. AB - This paper presents approaches used in one rural U.S. state to describe the level of injecting drug use and to estimate the number of injectors not receiving drug user treatment. The focus is on two broad areas of estimation that were used to present the prevalence of injecting drug use in Kentucky. The first estimation approach uses available data from secondary data sources. The second approach involves three small community studies. PMID- 12117070 TI - The community epidemiology work group approach. AB - "Drug abuse" provides many unique challenges to the research community. Some of these involve fundamental epidemiologic issues, such as measuring the extent of the problem, identifying and assessing changes in patterns and trends, detecting emerging "drugs of abuse", characterizing vulnerable populations and determining health and social consequences. A number of research methods are employed to address these issues. This paper describes one of these--a model in which ongoing surveillance of "drug abuse" is maintained through a network of community-based researchers, local officials, academics, and other interested and qualified members of the community. Timely, accurate, and cost-effective data can be generated through systematic collection and analysis of indirect indicators of "drug abuse" that are often routinely produced by a variety of community sources. This information, in turn, can be used to make informed public health policy decisions. The community-based network model has been implemented at the city, state, national, regional, and international levels, and a case is made that this type of program could be useful, as well, in understanding the dynamics of "drug abuse" in rural areas of the country. PMID- 12117071 TI - Utilizing technology: the challenges and opportunities facing "substance abuse" professionals in rural communities. AB - Many rural communities are actively pursuing technology as a resource for solving education, health care, and economic development issues. These communities are establishing a technology and telecommunication infrastructure that makes them appealing to individuals and companies from urban communities. But this has created a challenge and an opportunity for the mental health industry in general, and more specifically, "substance abuse" professionals. The opportunity for the "substance abuse" profession is to design and use new services using the exact same technologies that may precipitate the need for the services. PMID- 12117072 TI - Introduction: substance use in rural communities around the world. PMID- 12117073 TI - The impact of hormonal treatments on quality of life of patients with metastatic breast cancer. AB - BACKGROUND: The concept of quality of life (QOL) increasingly has been used to assess health-related outcomes associated with a specific disease or its treatment, especially in patients with incurable tumors, such as metastatic breast cancer (MBC). Hormonal therapy (HT) is often used to treat hormone receptor-positive MBC, with the primary treatment goal of reducing both disease burden and patient suffering. OBJECTIVE: This article reviews the instruments used to assess QOL in patients with breast cancer, the adverse effects of HTs, and the clinical trials that assess QOL in patients with MBC receiving various HTs. METHODS: Articles were identified for inclusion in this manuscript through the following searches, limited to English-language publications: MEDLINE (mid 1960s to January 2002), American Society of Oncology abstracts (1997-2001), and San Antonio Breast Cancer Symposium abstacts (2001 and 2002). The following search terms were used: quality of life, breast cancer, hormonal therapies, tamoxifen, toremifene, letrozole, anastrozole, exemestane, and megestrol acetate. CONCLUSIONS: QOL assessment following MBC treatment has become an important indicator of treatment effectiveness, and numerous clinical trials have studied the impact of HT on QOL. In general, the older HTs, such as the progestins and selective estrogen receptor modulators (SERMs), produce more adverse effects than do the newer HTs, such as the aromatase inhibitors (AIs) and estrogen receptor (ER) antagonists. QOL data regarding tamoxifen, a SERM associated with a high incidence of vasomotor symptoms and vaginal discharge, are limited, although tamoxifen has not been associated with significant psychological distress when administered as a chemopreventive or adjuvant MBC therapy in clinical trials. QOL studies comparing the third-generation AIs with tamoxifen or megestrol acetate show that the AIs produce a more favorable QOL, probably because these agents target the aromatase enzyme, which results in a lower incidence of thromboembolism and vaginal bleeding. Although QOL studies of the ER antagonist fulvestrant have not been conducted, several attributes of this new HT may contribute to the retention of a good QOL in patients with MBC. A variety of QOL assessment tools to measure the impact of HTs on patients with MBC are available. Clinical trial data regarding QOL in patients with MBC receiving HT will be useful for both clinicians and patients in evaluating treatment options and developing treatment strategies. PMID- 12117074 TI - Evolving uses of hormonal agents for breast cancer therapy. AB - BACKGROUND: During the past decade, a number of new hormonal therapies (HTs) have been developed, including the selective estrogen receptor modulators (SERMs), aromatase inhibitors (AIs), and estrogen receptor (ER) antagonists. Their uses in breast cancer are continually evolving as new clinical trial results become available. Although tamoxifen, the most widely used HT for breast cancer, was originally approved for and used in the treatment of metastatic breast cancer (MBC), its effectiveness as MBC therapy led to its subsequent assessment and use as adjuvant and risk-reduction therapy for breast cancer. However, tamoxifen is not universally effective in these settings and is associated with infrequent known toxicities such as increased risk of thromboembolism and endometrial cancer; therefore, a search for more effective and more tolerable HTs has evolved. OBJECTIVE: This article reviews the data supporting the use of newer HTs as initial treatment of MBC and their potential use as adjuvant, neoadjuvant, and chemopreventive therapies. METHODS: Articles for inclusion in this manuscript were identified through the following searches, limited to English-language publications: MEDLINE (mid 1960s to January 2002), American Society of Oncology abstracts (1997-2001), and San Antonio Breast Cancer Symposium abstracts (2001 and 2002). The following search terms were used: breast cancer, breast cancer guidelines, hormonal therapies, tamoxifen, toremifine, letrozole, anastrozole, exemestane, megestrol acetate, fulvestrant, and ICI 182,780. RESULTS: Recent studies have focused on newer agents as initial and subsequent treatment of MBC, adjuvant or neoadjuvant treatments of breast cancer, and chemopreventive agents in both healthy women and women with a history of ductal carcinoma in situ (DCIS). Results of clinical trials comparing AIs with tamoxifen as first-line MBC treatment show that AIs are as effective as, or more effective than, tamoxifen and are associated with fewer serious adverse events. Tamoxifen remains the gold standard for adjuvant therapy. However, preliminary results of ongoing clinical trials comparing tamoxifen with anastrozole suggest that anastrozole may be the superior agent. Both tamoxifen and the AIs have been shown to be active in the neoadjuvant treatment of breast cancer. Trial results have shown that tamoxifen is effective for breast cancer prevention in patients at high risk of developing breast cancer but who are otherwise healthy, patients with a history of DCIS, and patients with lobular carcinoma in situ. CONCLUSIONS: Although tamoxifen has been the gold standard of HT for breast cancer, results of ongoing trials assessing the newer HTs as initial, neoadjuvant, adjuvant, and chemopreventive therapies may substantially change our current clinical practice patterns. PMID- 12117075 TI - Sequencing of hormonal therapy in postmenopausal women with metastatic breast cancer. AB - BACKGROUND: Hormonal therapy (HT) is an important consideration in the management of postmenopausal women with metastatic breast cancer. Despite the fact that the advanced-stage disease is virtually incurable, HTs can offer patients disease control equivalent to that of chemotherapy, but with improved quality of life (QOL). Knowledge of the estrogen and progesterone receptor status, as well as other clinical factors, allows for selection of patients who are most likely to benefit from HT. Disease that becomes refractory to an initial HT may respond to another agent or class of HTs. Thus, HTs are generally administered sequentially, delaying the need for cytotoxic chemotherapy, which often reduces QOL. Optimal sequencing is thus one of the more important facets of HT. Prior to the release of a number of newer agents, tamoxifen had been considered as initial HT. At present, more agents exist, including the aromatase inhibitors, progestins, and the estrogen receptor antagonist fulvestrant. OBJECTIVE: This article reviews key trials evaluating the use of sequential HTs. METHODS: Articles were identified for inclusion in this manuscript through the following searches, limited to English-language publications: MEDLINE (mid 1960s to January 2002), American Society of Oncology abstracts (1997-2001), and San Antonio Breast Cancer Symposium abstracts (2001 and 2002). The following search terms were used: breast cancer, hormonal therapies, tamoxifen, toremifene, letrozole, anastrozole, exemestane, megestrol acetate, fulvestrant, and ICI 182,780. RESULTS: Results of Phase III studies have shown many of these agents to be equivalent or superior to tamoxifen and can be used initially to treat patients who either have failed tamoxifen therapy or may be unable to tolerate some of the toxicities associated with tamoxifen. For example, the aromatase inhibitors have been shown to be highly active and tolerable in postmenopausal women with breast cancer who have failed tamoxifen therapy or who are naive to HT. Other clinical trials have demonstrated the efficacy of fulvestrant in patients with metastatic breast cancer who are tamoxifen-resistant, and have shown fulvestrant to be at least as effective as anastrozole in tamoxifen-resistant patients. CONCLUSIONS: Although the optimum sequence of HTs remains controversial, using the newer agents as initial or subsequent therapy should improve QOL and may improve overall survival. PMID- 12117077 TI - Current issues on resistance, treatment guidelines, and the appropriate use of fluoroquinolones for respiratory tract infections. AB - BACKGROUND: Community-acquired respiratory tract infections comprise a large percentage of diseases treated by primary care physicians, and rates of antimicrobial use for respiratory tract infections are increasing. The fluoroquinolones comprise a drug class with broad-spectrum activity against many of the key pathogens associated with community-acquired respiratory tract infections, including Streptococcus pneumoniae, and other significant pathogens, such as Staphylococcus aureus and Pseudomonas aeruginosa. While fluoroquinolones have gained popularity, the settings for their appropriate use in treating respiratory tract infections remain unclear. OBJECTIVE: In this article, the mechanisms of fluoroquinolone resistance in S. pneumoniae, treatment guidelines, and the mode of spread of resistance are reviewed. METHODS: The authors conducted a MEDLINE search for articles published from 1990 to the present. Search terms included Streptococcus pneumoniae, fluoroquinolones, and resistance. Articles were selected for inclusion based on their relevance to the objective of this review. RESULTS: Although 3 sets of treatment guidelines for community-acquired pneumonia (CAP) currently exist in the United States, a consensus for the role of fluoroquinolones in the outpatient management of CAP has not been achieved. Factors mitigating for restraint in the outpatient use of fluoroquinolones include concern for the spread of resistance to "innocent-bystander" organisms, such as S. aureus and P. aeruginosa, as well as possible inappropriate "trickle down" use for other, less severe respiratory syndromes, such as bronchitis. CONCLUSION: Although the fluoroquinolones are potent agents against respiratory pathogens and have a clearly defined role in the treatment of hospitalized patients with CAP, their optimal role in the outpatient management of respiratory tract infections remains controversial. PMID- 12117076 TI - Statistical comparison of consumer drug expenditures and discretionary purchases to assess drug affordability. AB - BACKGROUND: Affordability may be defined as the absence of economic barriers to a good or service. There are 2 frequently observed measures of affordability: a consumer's ability to pay and his or her physical access to a good or service. Thus, most programs designed to subsidize consumers' health care costs, especially state programs that address prescription drug expenditures for people aged > or =65 years, base eligibility on measures of income as a proxy for a consumer's ability to pay. These measures do not explicitly include a consumer's willingness to pay for medications. For example, it is possible that some Medicare beneficiaries may be resistant to paying for medication because other major health care expenditures are typically covered by insurance. This resistance could be exacerbated by the keen awareness among the general population of the rising costs of medications. Because medications are considered a necessity, expenditure levels are usually compared with expenditures for other necessities, such as housing and medical services. OBJECTIVE: In an attempt to assess consumers' potential willingness to pay for medications, this article draws on data from the US Bureau of Labor Statistics' Consumer Expenditure Surveys to compare pharmaceutical expenditures with out-of-pocket expenditures for discretionary purchases, such as dining outside the home. RESULTS: Personal out-of-pocket expenditures for medications have ranged from 0.8% to 1.0% of consumer unit income since 1985. These expenditures are relatively small compared with those for necessities, such as housing (33%) and food (13.5%). They are also less than the share of income dedicated to many nonessentials. CONCLUSION: Assessing inability versus unwillingness to pay for medication remains a problem for both researchers and health care policy makers attempting to determine the affordability of medications. PMID- 12117078 TI - A modern rationale for the use of phenoxybenzamine in urinary tract disorders and other conditions. AB - BACKGROUND: Phenoxybenzamine (PBZ) is a nonselective, irreversible alpha adrenergic receptor antagonist that is approved for the treatment of diaphoresis and hypertension associated with pheochromocytoma. It may also be useful in several chronic conditions whose pathogenesis is mediated or affected by alpha adrenergic stimulation, such as lower urinary tract symptoms associated with benign prostatic hyperplasia (BPH) and neurogenic bladder (eg, secondary to myelomeningocele and in sphincter dyssynergia and autonomic dysreflexia); in an adjunctive role after urogenital surgery or brachytherapy by relieving symptoms associated with increased alpha-adrenergic tone; and in the treatment of complex regional pain syndrome (CRPS) and prostatitis. However, carcinogenic concerns may have limited its potential application. OBJECTIVE: The purpose of this article is to reassess the usefulness and contemporary application of PBZ for the control of urinary tract symptoms associated with BPH and neurogenic bladder, after urogenital surgery and brachytherapy, and in certain other conditions (eg, CRPS, prostatitis). METHODS: A search of literature published from 1966 to 2002 was performed on MEDLINE using the search terms phenoxybenzamine, alpha-adrenergic blockers, benign prostatic hyperplasia, neurogenic bladder, urinary retention, and complex regional pain RESULTS: Despite concerns about possible carcinogenicity, no reports of drug-related tumors have been made since PBZ's introduction in 1953. Investigators have used PBZ in off-label trials to alleviate symptoms of a variety of conditions that cause urinary retention. In adult male patients with retention due to inguinal hernioplasty and female patients with retention caused by vaginal repair, as well as in pediatric patients with myelomeningocele, treatment with PBZ improved bladder function and, in the patients with myelomeningocele, was associated with reduced incidence of urinary tract infection. Larger tri- als of PBZ in men with BPH produced significant urinary symptom relief (P < 0.05 in 2 studies). Moreover, studies suggest that PBZ may be useful in alleviating pain due to trauma and CRPS. The most common adverse events appear to be dizziness, impotence and ejaculatory dysfunction, and nasal stuffiness. CONCLUSIONS: No drug-related tumors in humans have been reported after -50 years of clinical experience with PBZ. Clinical trials have demonstrated that it can relieve symptoms in patients with BPH and other urologic and pain-related conditions. PMID- 12117079 TI - Donepezil and rivastigmine in the treatment of Alzheimer's disease: a best evidence synthesis of the published data on their efficacy and cost effectiveness. AB - BACKGROUND: Various drugs have been approved for the treatment of Alzheimer's disease (AD) in the United States and Canada, including donepezil and rivastigmine, although questions remain as to their efficacy, effectiveness, and long-term benefits. OBJECTIVE: The goal of this study was to conduct a best evidence synthesis of data on the efficacy and cost-effectiveness of donepezil and rivastigmine in the treatment of AD. METHODS: Relevant published randomized controlled trials (RCTs) and Phase IV open-label extension studies (excluding abstracts) were identified through searches of MEDLINE, HealthSTAR, and PsycINFO for the period January 1984 to October 2001. The bibliographies of retrieved articles were searched for additional publications. For inclusion in the best evidence synthesis, clinical trials had to pass a blinded quality assessment (score > or =5 on the Jadad scale) and use National Institute of Neurological and Communicative Disease and Stroke-Alzheimer's Disease and Related Disorders Association diagnostic criteria. Economic studies were selected using National Health Service Centre for Reviews and Dissemination criteria for reporting critical summaries of economic evaluations. RESULTS: Nine RCTs of donepezil and 2 of rivastigmine were identified and met inclusion criteria for the best-evidence synthesis. Eight donepezil trials and both rivastigmine trials included patients with mild AD (Mini-Mental State Examination [MMSE] score, 15-27) or moderate AD (MMSE score, 8-14); 1 donepezil trial included patients with moderate or severe AD (MMSE score, 0-7). In the RCTs of donepezil, the mean decrease in scores on the Alzheimer's Disease Assessment Scale-cognitive subscale (ADAS-cog) was greater with active treatment than with placebo (lower scores indicate less cognitive deterioration). In the RCTs of rivastigmine, ADAS-cog scores decreased over the follow-up period with both active treatment and placebo; however, scores decreased more with active treatment. Three Phase IV studies of donepezil and I Phase IV study of rivastigmine were identified. Their results were consistent with those of the RCTs. Ten economic studies (7 donepezil, 3 rivastigmine) were identified and reviewed. In 4 of the donepezil studies and all 3 rivastigmine studies, use of the drug cost less than a no-drug strategy. CONCLUSIONS: The efficacy data indicate that both donepezil and rivastigmine can delay cognitive impairment and deterioration in global health for at least 6 months in patients with mild to moderate AD. Patients receiving active treatment will have more favorable ADAS-cog scores for at least 6 months, after which their scores will begin to converge with those of patients receiving placebo. Differences in methodology, types of direct or indirect costs included, and sources of cost data made it difficult to compare and synthesize findings of the economic studies; therefore, the cost-effectiveness data are inconclusive. PMID- 12117080 TI - Efficacy of budesonide in moderate to severe asthma. AB - BACKGROUND: The worldwide prevalence of asthma is increasing by approximately 50% per decade. Budesonide is one of several inhaled corticosteroids available for the treatment of asthma and has been extensively evaluated in clinical trials. OBJECTIVE: This article reviews the published literature on the efficacy of budesonide in the management of adult and pediatric patients with moderate to severe asthma and compares budesonide with other inhaled corticosteroids and nonsteroidal treatment options. METHODS: All controlled, randomized studies in patients with moderate or severe asthma were considered for inclusion. Relevant studies were identified through a MEDLINE search of the period from 1980 to 2000 using the terms budesonide plus efficacy, with or without the termsfluticasone, mometasone, and beclomethasone. The manufacturer's reference database was used to identify additional publications. RESULTS: Budesonide is associated with a dose response effect in adults and children with moderate to severe asthma. The data on budesonide are in line with the current recommendation for a high starting dose of inhaled corticosteroid (800 microg/d), followed by downward titration to the minimal effective dose. Budesonide administered by Turbuhaler (AstraZeneca Pharmaceuticals LP, Wilmington, Del) dry-powder inhaler (DPI) was effective at a significantly lower dose than beclomethasone dipropionate (BDP) administered by pressurized metered-dose inhaler (pMDI) (P = 0.009), whereas its efficacy was similar to that of BDP delivered by hydrofluoroalkane pMDI and that of fluticasone propionate administered by DPI. Inhaled budesonide therapy was shown to be oral corticosteroid sparing in patients with severe asthma, thus reducing the total corticosteroid dose and the risk of systemic side effects. Pulmicort Respules (AstraZeneca), a nebulized formulation, was effective in the treatment of moderate to severe asthma in patients aged > or =12 months. CONCLUSIONS: Once- or twice-daily administration of budesonide delivered via the Turbuhaler and Pulmicort Respules systems has been shown to be well tolerated and efficacious in populations with moderate to severe asthma. PMID- 12117081 TI - Efficacy and tolerability of gatifloxacin in community treatment of acute exacerbations of chronic bronchitis. AB - BACKGROUND: Recognizing acute exacerbations of chronic bronchitis (AECB) and selecting appropriate antibiotic treatment for patients who would benefit most is a challenge for community-based physicians. OBJECTIVE: The Tequin Clinical Experience Study, an open-label, noncomparative, postmarketing trial, assessed the efficacy and tolerability of gatifloxacin, an 8-methoxy fluoroquinolone, in the treatment of AECB in the community-practice setting. METHODS: Consecutive patients with respiratory tract infections in community-based settings were eligible for participation. Treated patients (N = 2512) included 1107 men (44.1%) and 1405 women (55.9%) aged > or =18 years with a clinical diagnosis of chronic bronchitis. All participants received oral gatifloxacin 400 mg once daily for 7 to 10 days. Clinical response was determined via telephone contact conducted by the investigator or study coordinator using case-report forms or during an office visit after the last dose. The investigator or coordinator collected expectorated or induced sputum specimens that were then smeared on a microscope slide, stored in a tube, and transported to a central reference laboratory for Gram-staining and culture. Of 1388 pretreatment sputum specimens submitted, pathogens were isolated from 424. RESULTS: The most frequently detected pathogens were Haemophilus influenzae, Moraxella catarrhalis, and Streptococcus pneumoniae. All H. influenzae and 99% of S. pneumoniae isolates tested were susceptible to gatifloxacin. Of the 2267 patients with a determinable clinical response, 2084 (91.9% [95% CI, 90.8%-93.0%]) were cured (all acute symptoms improved or returned to baseline level, no new symptoms present, no additional antibiotic required). The 95.8% cure rate in 166 patients with H. influenzae included 100% of those with beta-lactamase-positive strains. Overall, 89.2% of 111 patients with M. catarrhalis were cured; rates were similar regardless of beta-lactamase production. The clinical cure rate in 74 patients with S. pneumoniae was 98.6% and was independent of the degree of penicillin resistance (minimum inhibitory concentration > or =2.0 microg/ mL). All 6 patients infected with S. pneumoniae fully resistant to penicillin were cured. Gatifloxacin was generally well tolerated, and the majority of adverse events were mild to moderate; only 11 drug related adverse events in 10 patients (0.4%) were serious. Drug-related nausea (3.0%), dizziness (1.5%), diarrhea (1.2%), and vomiting (0.9%) were the most common adverse events. CONCLUSIONS: The high clinical cure rate and favorable tolerability support gatifloxacin as a rational choice for the treatment of AECB in patients such as those in this community-based study. PMID- 12117083 TI - A retrospective electronic chart review of blood pressure changes in elderly patients treated with amlodipine or an angiotensin-converting enzyme inhibitor or angiotensin II receptor blocker. AB - BACKGROUND: Despite the high costs of managing hypertension, pharmacologic intervention is cost-effective, particularly in patients at highest risk for cardiovascular events. The prevalence of hypertension in the elderly and the age associated risks of coronary artery disease and stroke suggest that early identification and aggressive treatment should be priorities in this population. OBJECTIVE: The aim of this study was to compare the effect of amlodipine and angiotensin-converting enzyme (ACE) inhibitors and angiotensin II receptor blockers (ARBs) in the treatment of essential hypertension in elderly patients (>60 years) in an actual practice setting. METHODS: This was a retrospective cohort analysis using electronic medical records stored in the Physicians Data Corporation cardiology database. Patients aged >60 years who received care from a cardiologist and who had a recorded diagnosis of hypertension during 1997 or 1998 were identified. For inclusion, patients had to have received an initial prescription for amlodipine, an ACE inhibitor, or an ARB at the index visit. Systolic blood pressure (SBP) and diastolic blood pressure (DBP) readings from the index visit and > or =1 subsequent visit (<180 days after the index visit) were assessed. RESULTS: A total of 192 patients (56.3% male; mean age, 71.9 years) met the inclusion criteria. Amlodipine-treated patients experienced a mean decrease in SBP of 26.7 mm Hg, compared with 18.8 mm Hg in patients receiving an ARB and 15.8 mm Hg for patients receiving an ACE inhibitor (P = 0.008, amlodipine vs ACE inhibitor). DBP decreased 8.8 mm Hg with amlodipine, 8.7 mm Hg with an ARB, and 6.2 mm Hg with an ACE inhibitor. After adjusting for age, sex, and disease severity, amlodipine-treated patients were -4 times as likely to move to a better blood pressure stage than patients treated with an ARB or an ACE inhibitor (odds ratio, ARB vs amlodipine: 0.245; 95% CI, 0.080-0.753; odds ratio, ACE inhibitor vs amlodipine: 0.234; 95% CI, 0.072-0.761). CONCLUSION: Results of this study indicate that in patients aged >60 years, amlodipine may be an effective therapy for hypertension. PMID- 12117082 TI - Comparison of the clinical efficacy and tolerability of olopatadine hydrochloride 0.1% ophthalmic solution and loteprednol etabonate 0.2% ophthalmic suspension in the conjunctival allergen challenge model. AB - BACKGROUND: Olopatadine hydrochloride 0.1% ophthalmic solution and loteprednol etabonate 0.2% ophthalmic suspension are topical antiallergic agents indicated for treatment of the signs and symptoms of allergic conjunctivitis and seasonal allergic conjunctivitis (SAC), respectively. OBJECTIVE: The purpose of this study was to compare the efficacy and tolerability of olopatadine, loteprednol, and placebo in inhibiting the early-phase allergic reaction (within 30 minutes) after conjunctival allergen challenge (CAC). METHODS: This was a single-center, randomized, double-masked, parallel-controlled CAC study. It consisted of 3 visits, with CAC performed at visit 1, confirmation and randomization at visit 2, and evaluation of the treatments at visit 3. Subjects with a history of allergic conjunctivitis were randomized to receive olopatadine, loteprednol, or placebo in a 2:2:1 ratio. Because loteprednol requires a loading period to achieve maximum efficacy, subjects assigned to this treatment received loteprednol QID bilaterally for a 14-day period; the olopatadine and placebo groups received placebo QID bilaterally during this period. At the evaluation visit, subjects received 1 drop of the assigned treatment in each eye. Fifteen minutes later, they were challenged with allergen. Subjects evaluated itching at 3, 5, and 10 minutes after challenge using a standardized 5-point scale; the investigator evaluated redness at 10, 15, and 20 minutes after challenge. Intraocular pressure (IOP) was measured at baseline and after the 14-day loading period. Nonparametric analyses were performed on the change from visit 2 to visit 3 in mean itching and redness scores for each time point, and on the change in mean IOP from visit 1 to visit 3. RESULTS: Fifty subjects (86% white; 42% male, 58% female; age range, 21 71 years) were enrolled and completed the study (20 olopatadine, 20 loteprednol, 10 placebo). The allergens to which subjects reacted were ragweed pollen (40%), cat hair or dander (30%), grass pollen (24%), and tree pollen (6%). The difference in inhibition of itching and redness was clinically significant (> or =1 unit difference) and statistically significant (P < 0.05) in favor of olopatadine compared with loteprednol at all 3 time points. The loteprednol group had a statistically significant increase in IOP after 2 weeks of treatment (P < 0.001). CONCLUSION: In the population studied, olopatadine was more efficacious than loteprednol in reducing the acute signs and symptoms of SAC during the early phase of the ocular allergic reaction and appeared to be better tolerated. PMID- 12117084 TI - Efficacy and patient satisfaction with cromolyn sodium nasal solution in the treatment of seasonal allergic rhinitis: a placebo-controlled study. AB - BACKGROUND: Because the symptoms of seasonal allergic rhinitis are readily recognizable, many individuals self-medicate with nonprescription agents. The mast-cell stabilizer cromolyn sodium is available for over-the-counter (OTC) use in the prevention and treatment of allergic rhinitis. OBJECTIVE: The goal of the study was to evaluate the efficacy of cromolyn sodium 4% nasal solution for the treatment of allergic rhinitis in self-selected patients in a nonprescription setting. A secondary objective was to determine whether the instructions and warnings in the OTC labeling provide adequate information for safe and proper OTC use. METHODS: This 2-week, multicenter study had a randomized, double-blind, placebo-controlled, parallel-group design. Advertisements were used to identify allergy sufferers who used OTC products but had not used OTC allergy medications within 48 hours of study entry and did not require prescription medications to control allergy symptoms. Enrolled patients were randomized to receive either cromolyn sodium 4% nasal solution in a metered-spray bottle containing 200 doses or an identical-appearing placebo. They were instructed to follow the directions for use on the label: that is, to administer 1 spray in each nostril every 4 to 6 hours, no more than 6 times per day, for the prevention or relief of symptoms. Daily use of study drug and concomitant medications was recorded in patient diaries. Patients also rated symptom severity, symptom relief, medication efficacy, and helpfulness of the label instructions. Relief of overall and individual rhinitis symptoms was assessed at the end of weeks 1 and 2. RESULTS: The intent-to-treat population consisted of 1150 patients (580 cromolyn sodium, 570 placebo). Cromolyn sodium provided greater relief than placebo on all efficacy measures and was statistically significantly more effective than placebo in controlling allergy symptoms (P = 0.02), providing overall symptom relief (P = 0.02), and relieving sneezing (P = 0.01) and nasal congestion (P = 0.03). The instructions for use were rated helpful to extremely helpful by >92% of patients, although only approximately 48% of patients used the drug according to the label instructions. The most common adverse events in both groups were headache and rhinitis, and there was no significant difference in the rates of such events between groups. CONCLUSION: In this study, cromolyn sodium 4% nasal solution was well tolerated and effective, suggesting that it is suitable for OTC use in the treatment of seasonal allergic rhinitis. PMID- 12117086 TI - Physician-reported management of edema and destabilized blood pressure in cyclooxygenase-2-specific inhibitor users with osteoarthritis and treated hypertension. AB - BACKGROUND: The addition of a nonsteroidal anti-inflammatory drug to the regimen of a patient with treated hypertension can cause a destabilization of blood pressure. OBJECTIVE: The aim of this study was to describe physician-reported management of clinically significant edema and/or destabilized blood pressure in patients with osteoarthritis (OA) and hypertension when initiating therapy with rofecoxib or celecoxib. METHODS: A cross-sectional survey was administered to physicians who attended one of several arthritis consultant programs sponsored by Pharmacia Corporation, with attendees selected by local sales representatives. Each program included a clinical presentation by a physician concerning the cardiorenal safety of celecoxib, followed by a consultative presentation and session led by a Pharmacia Clinical Education Manager. RESULTS: A total of 828 physicians in the following specialties completed the survey: family practice (33.0%), internal medicine (25.0%), orthopedics (15.2%), and rheumatology (11.4%). Responding physicians expected that the majority of patients who experienced edema would contact them (68.4%). They reported that they schedule follow-up visits for blood pressure monitoring 65.6% of the time after initiating a cyclooxygenase-2 (COX-2)-specific inhibitor, with family practitioners and internists most likely to indicate that they would do so and orthopedists least likely. Responding physicians indicated that the presence of edema and destabilized blood pressure generally led to discontinuation of the COX-2 specific inhibitor (58%-82% of the time). Internists and family practitioners were most likely to report that they treat edema by initiating or modifying diuretic therapy (33%-51% of the time). For destabilized blood pressure, an antihypertensive drug was reported to be initiated or modified 40% to 55% of the time by family practitioners and internists, whereas orthopedists indicated that they referred patients to the primary care provider. The COX-2-specific inhibitor prescribed resulted in management differences: physicians indicated that they were more likely to switch from rofecoxib to celecoxib in the event of edema or destabilized blood pressure, whereas they were more likely to adjust the celecoxib dose than the rofecoxib dose. Because the data were captured from convenience samples of physicians attending sponsored meetings, it is possible that respondents provided the answers they thought the sponsor would want. Because this was a cross-sectional survey, reported behavior was not compared with actual behavior. CONCLUSIONS: A significant percentage of physicians reported that they monitor patients with OA and hypertension for the occurrence of destabilized blood pressure and edema after initiation of a COX-2-specific inhibitor. Physicians indicated that they would nearly always intervene when either event is identified. PMID- 12117087 TI - Evaluation of the efficacy and tolerability of 0.5% fluorouracil cream and 5% fluorouracil cream applied to each side of the face in patients with actinic keratosis. AB - BACKGROUND: A new 0.5% fluorouracil cream has been developed that provides an alternative to the more highly concentrated topical formulations of fluorouracil that are currently available. OBJECTIVE: This was a comparison of the tolerability and efficacy of the 0.5% and 5% fluorouracil creams in the treatment of actinic keratosis (AK). METHODS: During this single-blind, randomized study, patients with > or =6 AK lesions were treated for 4 weeks with the 0.5% (once daily) and 5% (twice daily) fluorouracil creams applied to opposite sides of the face. After the end of treatment, patients were followed for an additional 4 weeks. Efficacy variables included absolute and percent reductions in AK lesions from baseline and total clearance of AK lesions. A questionnaire was used to evaluate patients' treatment preferences. Tolerability was evaluated through continuous monitoring of adverse events. RESULTS: Treatment with 0.5% fluorouracil cream reduced the number of AK lesions from 11.3 at baseline to 2.5 at the end of the 4-week follow-up phase, compared with a reduction from 10.3 to 4.2 lesions after treatment with 5% fluorouracil cream. The reduction was significantly greater with the 0.5% cream compared with the 5% cream (P = 0.044). The 0.5% cream was as effective as the 5% cream in terms of the percent reduction in AK lesions from baseline (67% and 47%, respectively) and in achieving total clearance of AK lesions (both treatments, approximately 43% of patients). Both treatments were associated with similar degrees of investigator-rated irritation; however, patients preferred the 0.5% cream because they felt it was more tolerable (P = 0.003), easier to apply, and had a once-daily application schedule. Although all patients experienced facial irritation in association with both creams, fewer patients treated with the 0.5% cream reported symptoms of facial irritation. CONCLUSIONS: In this study, 0.5% fluorouracil cream once daily was at least as effective as 5% fluorouracil cream twice daily in terms of the percent reduction in AK lesions and total clearance of AK lesions; it was more effective than the 5% cream in reducing the absolute number of AK lesions from baseline. Patients preferred the 0.5% cream to the 5% cream. PMID- 12117088 TI - Despite the remarkable advances in oral health care, pain and anxiety continue to be significant deterrents for seeking dental services. PMID- 12117085 TI - A double-blind, single-dose comparison of the analgesic efficacy of tramadol/acetaminophen combination tablets, hydrocodone/acetaminophen combination tablets, and placebo after oral surgery. AB - BACKGROUND: Improved clinical outcomes have been documented with combinations of oral analgesic agents, particularly those with complementary activities. However, because not all combinations or dose ratios lead to enhanced analgesia or reduced adverse events (AEs), each combination and dose ratio must be evaluated individually in carefully designed preclinical and clinical trials. OBJECTIVE: The goal of the study was to compare the efficacy and safety of 37.5 mg tramadol/325 mg acetaminophen tablets (T/APAP), 10 mg hydrocodone bitartrate/650 mg acetaminophen tablets (HC/APAP), and placebo in the treatment of postoperative dental pain. METHODS: This was a single-center, double-blind, parallel-group, placebo- and active-controlled study in adults with at least moderate pain (score > or =50 on a 100-mm pain visual analog scale) after extraction of > or =2 impacted third molars. Patients were randomized to receive 1 or 2 T/APAP tablets, 1 HC/APAP tablet, or placebo. Scores for hourly pain relief (PAR), pain intensity difference (PID), and combined PAR and PID (PRID) were based on reported pain at 30 minutes and each successive hour for 8 hours. Primary efficacy measures were summary pain intensity and pain relief scores (total pain relief [TOTPAR], sum of pain intensity differences [SPID], and sum of pain relief and pain intensity differences [SPRIDI) for 0 to 4 hours, 4 to 8 hours, and 0 to 8 hours. Secondary efficacy measures were hourly PAR, PID, and PRID scores; onset and duration of pain relief; time to remedication with a supplemental analgesic agent; and patients' overall assessment of medication. RESULTS: Two hundred adults took part in the study (50 per treatment group) and were included in the efficacy and safety analyses. T/APAP 75/650 mg and HC/APAP were statistically superior to placebo on the primary efficacy measures of TOTPAR, SPID, and SPRID (P < or = 0.024), as well as on hourly PAR, PID, and PRID over 6 hours (P < or = 0.045). All active treatments were statistically superior to placebo in terms of onset of pain relief (P < or = 0.001), duration of pain relief (P < or = 0.024), time to remedication (P < 0.001), and patients' overall assessment of medication (P < 0.001). A statistically significant dose response with T/APAP (2 tablets > 1 tablet > placebo) was seen for TOTPAR, SPID, and SPRID (all, P < or = 0.018). The median time to onset of pain relief was approximately 34.0 minutes with 2 T/APAP tablets and 25.4 minutes with HC/APAP. Although the median time to onset of pain relief was shorter with HC/APAP, two T/APAP tablets had comparable efficacy to HC/APAP. The median time to remedication with a supplemental analgesic agent was 169.0 minutes in the T/APAP 75/650 mg group and 204.0 minutes in the HC/APAP group. However, the duration of pain relief, as defined by time to remedication, was not significantly different between these 2 groups. The overall incidence of AEs was lower with T/APAP (0% treatment-related AEs) than with HC/APAP (4%) or placebo (10%). The incidence of nausea (18% T/APAP, 36% HC/APAP) and vomiting (12% T/APAP, 30% HC/APAP) was approximately 50% lower with 2 T/APAP tablets than with HC/APAP (P < 0.05). CONCLUSIONS: T/APAP tablets provided effective, rapid (< or = 34 minutes), dose-dependent analgesia for the treatment of postoperative dental pain. Two T/APAP tablets provided analgesia comparable to that provided by HC/APAP with better tolerability. PMID- 12117089 TI - A method for comparison of biomedical publication quality across ISI discipline categories. AB - The purpose of this paper is to offer a method to help to objectively compare quality of publication in biomedical journals in different disciplines with varying ISI Impact Factors (IF). Three methods--the number of journals per ISI Journal Citation Report discipline category/10, the IF/10, and the log (IF+1)/10- were used to calculate an article score. The distribution of article scores were compared across three defined ISI discipline categories: two clinical categories, dentistry (ISI category--dentistry, oral surgery, and medicine, forty-five journals) and medicine (ISI category--medicine, internal and general, 110 journals), and one basic science category, physiology (ISI category--physiology, seventy-four journals). The use of article scores per discipline category enables a reasonable, relative comparison of the quality of biomedical publications of individuals across disciplines for the purpose of promotion or awarding of research grants. PMID- 12117090 TI - The use of radiation dose-reduction techniques in the practices of dental faculty members. AB - X-ray exposure to dental patients has been significantly reduced by the introduction of speed group E intraoral film, rectangular beam limitation, long position indicating devices (PIDs), and rare-earth intensifying screens for extraoral radiography. Research indicates that many dentists do not use these techniques. However, schools of dentistry have implemented them to varying degrees for many years, so this investigation was conducted to determine the extent to which dental school faculty members use these materials and techniques in their own practices. Comparisons were made between full- and part-time instructors, those in practice for fifteen years or less and those in practice for more than fifteen years, and those with postgraduate education versus those with no formal education beyond dental school. The significance of differences was measured with chi-square analysis. The results indicate that dentists with faculty appointments utilize dose-reducing techniques to degrees that are comparable to or greater than reported usage by non-dental faculty practitioners. Faculty dentists in practice fifteen years or less are more likely than their older colleagues to use E-speed film (p = 0.001), whereas those in practice more than fifteen years are more likely to use longer PIDs (p = 0.049). Greater acceptance of these practices by faculty may lead to reinforcement of their use in the clinical education of dental students. PMID- 12117091 TI - Mentoring future dental educators through an apprentice teaching experience. AB - To address concerns about the growing shortage of dental educators, the UCLA School of Dentistry initiated an elective course to introduce fourth-year students to issues in academic dentistry and to provide an apprentice teaching experience. Participants in the elective (referred to as student teachers) developed a microcourse entitled "Welcome to Dental Anatomy," presented to incoming first-year students during orientation week. Under the guidance of faculty mentors, the student teachers were responsible for development of course content, teaching aids, and evaluation methodology. Two cycles of the elective have been completed reaching a total of twenty-one fourth-year students to date. The positive impact on student teachers and incoming first-year students indicates that this approach has great potential for encouraging more graduates to pursue careers in academic dentistry. In addition, the program has the potential to be expanded by adaptation to other foundational courses in the dental and dental hygiene curricula. PMID- 12117092 TI - Reflections on clinical practice by first-year dental students: a qualitative study. AB - During the first and second year of the dental curriculum, students have little time to process and learn the didactic material in a meaningful way because of the large number of required courses in the curriculum. If an outcome of dental education is to promote critical thinking, however, methods need to be explored to encourage this process in dental students throughout the curriculum. Reflecting on experience is the way learners "make meaning" out of the information they acquire, and "making meaning" is an integral component in the development of critical thinking. The purpose of this pilot study was to explore how reflection on clinical experiences may facilitate the development of critical thinking in first-year dental students as well as assist them in integrating their didactic coursework with clinical care. I used Luborsky's thematic analysis to analyze semi-structured interviews, clinical observations, and written reflection papers from dental students. The major themes identified from the data were: 1) connections between didactic material and the clinical experience, 2) the students' vision of their future role as dentists, and 3) the nature of the dentist-patient interaction. The data further suggest a process of reflective thinking that begins with students' questioning assumptions about their prior experience and knowledge that leads them to look at things in a new way and ultimately to recognize the need to take some action to provide care to the patient. The findings suggest that encouraging students to keep a clinic journal or write reflection papers about their experience may be a way to enhance student learning and is an area that deserves further research. PMID- 12117093 TI - Predicting student performance in preclinical technique courses using the theory of ability determinants of skilled performance. AB - The purpose of this research was to validate Ackerman's theory of ability determinants of skilled performance using sub-test scores of the Dental Admission Test (DAT) in predicting student performance in preclinical technique courses. The Ackerman theory is a valid, reliable schema in the applied psychology literature used to predict complex skill acquisition. Inconsistent stimulus response skill acquisition depends primarily on determinants of cognitive ability. The cognitive phase of skill acquisition is associated with demands on general abilities. Task accomplishment requires attentional resources, and inconsistent tasks do not improve with practice. It is theorized that the Perceptual Ability Test (PAT) is a valid cognitive determinant for spatial ability in this schema. Each new procedure learned in the preclinical technique courses is novel, includes a spatial relations component, and reflects inconsistent skill acquisition. The PAT scores of four classes were compared to the final grades in eight preclinical technique courses. Results showed that PAT scores account for a significantly high proportion, approximately 25 percent, of the variance of the final grades in the preclinical technique courses. Based on this strong correlation, dental school admissions committees may consider re evaluating the weight of the DAT scores in the admissions process. PMID- 12117094 TI - Introduction to section: Dentistry and primary care--an evaluation of postgraduate general dentistry training. PMID- 12117095 TI - Critical issues for dentistry: PGD program directors respond. AB - Discussion of critical issues facing postgraduate education in general dentistry (PGD) and dental education in general has been intense in the past decade. This study reports on critical issues raised by directors of PGD programs that may help direct future research and action within dental education and the larger profession. The analysis reports responses to an open-ended question sent to all U.S. PGD program directors regarding critical issues facing their training programs. Of 212 surveys, 169 program directors submitted written responses regarding critical issues. Twelve unique themes were identified: lack of postdoctoral applicants (two subthemes were high student debt and students' preference for private practice); student quality; professionalism and attitudes; number of postdoctoral positions; lack of funding; quality of facilities; special patient care; program curriculum; educator issues; mandatory or encouraged PGD year; value of dental program; and dentist shortage. Significant differences between AEGD and GPR directors were observed for two of the twelve areas: high student debt and value of dental program. The study provided insight into the thoughts of a large proportion of the U.S. PGD program directors "in the trenches." Some consideration of allowable expenses may be needed to align federal training support to best address program director needs. PMID- 12117097 TI - Curriculum emphasis and resident preparation in postgraduate general dentistry programs. AB - In 1999 HRSA contracted with the UCLA School of Dentistry to evaluate the impact of federal funding on postgraduate general dentistry programs. Part of that evaluation analyzed curriculum emphasis and preparation of incoming residents in advanced general dentistry programs over a five-year period. Directors of 208 civilian AEGD and GPR programs were surveyed about the curriculum content of their programs, increased or decreased emphasis in thirty subject areas, and resident preparation and quality (GPA and National Board scores). Results indicate that curriculum changes in AEGD and GPR programs over the time period have been responsive to the changing nature of general practice. At least half of all program directors reported that their residents were less than adequately prepared in fourteen curriculum areas. Sub-analyses were conducted for AEGD/GPR programs and HRSA-funded versus nonfunded programs. Multivariate regression identified lower student quality as the most important program variable in predicting a perceived need for resident remediation. Logistic regression showed that programs with higher resident GPA and National Board Part I scores had less difficulty filling resident positions. PMID- 12117096 TI - Military and VA general dentistry training: a national resource. AB - In 1999, HRSA contracted with the UCLA School of Dentistry to evaluate the postgraduate general dentistry (PDG) training programs. The purpose of this article is to compare the program characteristics of the PGD training programs sponsored by the Armed Services (military) and VA. Surveys mailed to sixty-six VA and forty-two military program directors in fall 2000 sought information regarding the infrastructure of the program, the program emphasis, resident preparation prior to entering the program, and a description of patients served and types of services provided. Of the eighty-one returned surveys (75 percent response rate), thirty were received from military program directors and fifty one were received from VA program directors. AEGDs reported treating a higher proportion of children patients and GPRs more medically intensive, disadvantaged and HIV/AIDS patients. Over half of the directors reported increases in curriculum emphasis in implantology. The program directors reported a high level of inadequate preparation among incoming dental residents. Having a higher ratio of residents to total number of faculty predicted inadequate preparation (p=.022) although the model was weak. Although HRSA doesn't financially support federally sponsored programs, their goal of improved dental training to care for medically compromised individuals is facilitated through these programs, thus making military and VA general dentistry programs a national resource. PMID- 12117098 TI - Characteristics of civilian postdoctoral general dentistry programs. AB - U.S. civilian (non-VA/non-military) Advanced Education in General Dentistry (AEGD) and General Practice Residency (GPR) programs were identified (n=208) and surveyed. The assessment evaluated infrastructure support, populations served, services provided, and trainee stipends. One hundred thirty-one programs responded (thirty-two AEGD, 64 percent/ninety-nine GPR, 63 percent). Sixty-nine programs were HRSA-funded (53 percent), and sixty-three (47 percent) were nonfunded. One hundred and five responses identified hospital/medical center resources; fifty-six indicated dental school support. Mean faculty support was similar regardless of program type or HRSA funding. Mean first-year positions in AEGDs were greater than GPRs. Mean first-year GPR positions were greater in funded than in nonfunded programs. A comparison of AEGD and GPR programs showed that residents in GPRs treated more children, medically intensive, economically/socially disadvantaged, and in-patient/same-day surgery patients (p<0.05). Residents in AEGDs treated more healthy adults (p<0.05). GPRs treated more lower fee (no pay, Medicaid, welfare/general relief, Medicare, and capitation/HMO) patients. AEGDs treated more insurance/private pay patients (p=.0001). No differences existed in comprehensive care and emergency visits between AEGDs and GPRs. GPRs treated more hospital-based patients. The mean stipends for GPRs ($32,055) and AEGDs ($22,403) were different. PMID- 12117099 TI - Dental and skeletal stem cells: potential cellular therapeutics for craniofacial regeneration. AB - The study of stem cells has received considerable attention since the discovery that adult stem cells have the capacity to form many different tissue types. Technical advances have helped identify potential stem cells, and their capacity for regenerating tissues is being studied in transplantation models. Further study of the isolation, nature, and differentiation potential of stem cells will likely have a positive impact on our understanding of human development and regenerative medicine. This review highlights the difference between embryonic and adult stem cells and discusses the potential use of these cells for cellular therapeutics for craniofacial regeneration. PMID- 12117100 TI - Marbofloxacin in equine medicine: have we got the doses right? PMID- 12117101 TI - Carbohydrate digestion by the horse: is it a limiting factor? PMID- 12117102 TI - The use of the Tridan system of identifying equine teeth is gaining credence in the veterinary literature. PMID- 12117103 TI - Evaluation of nebulised hay dust suspensions (HDS) for the diagnosis and investigation of heaves. 1: Preparation and composition of HDS. AB - Currently, heaves is investigated by exposing susceptible horses to dusty hay. Consequently, the response will be dependent on the organic dust content and composition of the hay. It was hypothesised that the use of a hay dust suspension (HDS) would reduce the variability of the challenge and therefore standardise experimental protocols. Furthermore, analysis of HDS would also permit further investigation of the organic dust components responsible for the response. Three hay dust suspensions (HDS-1, 2 and 3) were prepared for use in the diagnosis and investigation of heaves. HDS were produced from fine dust particles, comprising mostly fungal spores, collected from 3 batches of dusty hay. HDS-1 and 3 were analysed for endotoxin, beta-D-glucan and protein concentrations, general protease activity and enumeration and size distribution of particulates. Protease activity was mainly attributable to a 28 kDa serine protease and to 85 kDa and 160 kDa metalloproteases. The particulate and soluble components of HDS could be aerosolised by jet nebulisation. We therefore conclude that detailed analysis of HDS is possible, that such a challenge system provides a method of standardising experimental protocols and that all components of HDS (both soluble and particulate) can be delivered to the lung using standard nebulisation techniques. For the above reasons, nebulised HDS offers considerable advantages over conventional hay/straw challenge for the diagnosis and investigation of heaves. PMID- 12117104 TI - Evaluation of nebulised hay dust suspensions (HDS) for the diagnosis and investigation of heaves. 2: Effects of inhaled HDS on control and heaves horses. AB - To evaluate inhaled hay dust suspensions (HDS) as a tool for the diagnosis and investigation of heaves, the pulmonary inflammatory and functional consequences of inhalation challenge with 3 different HDS were determined in 6 control and 7 asymptomatic heaves horses. Heaves horses given HDS challenge developed the characteristic features of heaves, including airway neutrophilia, obstructive airway dysfunction and mucus hypersecretion. While HDS challenge induced a mild airway neutrophilia in controls, the no-response threshold for controls was greater than that of heaves horses, and there was no overlap in BALF neutrophil ratio of controls and heaves horses. Furthermore, HDS challenge did not induce airway dysfunction or mucus hypersecretion in controls. Therefore, HDS challenges enabled differentiation of control and heaves horses. Interestingly, in both groups, the airway neutrophilia was a dose-dependent response rather than an 'all or nothing' response. This study suggests that HDS challenges are of value in the diagnosis and investigation of heaves. PMID- 12117105 TI - Evaluation of nebulised hay dust suspensions (HDS) for the diagnosis and investigation of heaves. 3: Effect of fractionation of HDS. AB - To investigate the relative importance of inhaled particulates and soluble components in the response to inhaled hay dust suspension (HDS), heaves (previously termed chronic obstructive pulmonary disease; n = 7) and control (n = 6) horses were given inhalation challenges with whole and fractionated HDS. Fractionation was achieved by centrifugation to yield supernatant (SUP) and particulate debris. The particulates were then washed repeatedly in saline to produce a washed particulate (WP) fraction which comprised mainly fungal spores, and a wash fraction (WF) which comprised saline and soluble components washed from the surface of the particulates. Inhalation of HDS and SUP induced a significant airway neutrophilia in both groups, with the magnitude of the response being significantly greater in heaves horses. SUP induced significantly less airway neutrophilia than HDS in both groups, despite the endotoxin and protease content of HDS and SUP being comparable. WP and WF induced only a slight airway neutrophilia in heaves horses. However, a combined challenge with SUP and WP induced a neutrophilic response approaching the magnitude of that following HDS challenge, indicating that dust particulates contribute to the pulmonary recruitment of neutrophils in heaves. Consequently, inhalation challenge with HDS, which contains both particulates and soluble dust components, may be a more useful tool for the diagnosis and investigation of heaves than aqueous dust extracts, which contain only soluble components. PMID- 12117106 TI - Molecular characterisation of carbohydrate digestion and absorption in equine small intestine. AB - Dietary carbohydrates, when digested and absorbed in the small intestine of the horse, provide a substantial fraction of metabolisable energy. However, if levels in diets exceed the capacity of the equine small intestine to digest and absorb them, they reach the hindgut, cause alterations in microbial populations and the metabolite products and predispose the horse to gastrointestinal diseases. We set out to determine, at the molecular level, the mechanisms, properties and the site of expression of carbohydrate digestive and absorptive functions of the equine small intestinal brush-border membrane. We have demonstrated that the disaccharidases sucrase, lactase and maltase are expressed diversely along the length of the intestine and D-glucose is transported across the equine intestinal brush-border membrane by a high affinity, low capacity, Na+/glucose cotransporter type 1 isoform (SGLT1). The highest rate of transport is in duodenum > jejunum > ileum. We have cloned and sequenced the cDNA encoding equine SGLT1 and alignment with SGLT1 of other species indicates 85-89% homology at the nucleotide and 84 87% identity at the amino acid levels. We have shown that there is a good correlation between levels of functional SGLT1 protein and SGLT1 mRNA abundance along the length of the small intestine. This indicates that the major site of glucose absorption in horses maintained on conventional grass-based diets is in the proximal intestine, and the expression of equine intestinal SGLT1 along the proximal to distal axis of the intestine is regulated at the level of mRNA abundance. The data presented in this paper are the first to provide information on the capacity of the equine intestine to digest and absorb soluble carbohydrates and has implications for a better feed management, pharmaceutical intervention and for dietary supplementation in horses following intestinal resection. PMID- 12117107 TI - Pharmacokinetics of marbofloxacin in mature horses after single intravenous and intramuscular administration. AB - The pharmacokinetic behaviour of marbofloxacin, a new fluoroquinolone antimicrobial agent developed exclusively for veterinary use, was studied in mature horses (n = 5) after single-dose i.v. and i.m. administrations of 2 mg/kg bwt. Drug concentrations in plasma were determined by high performance liquid chromatography (HPLC) and data obtained were subjected to compartmental and noncompartmental kinetic analysis. This compound presents a relatively high volume of distribution (V(SS) = 1.17 +/- 0.18 l/kg), which suggests good tissue penetration, and a total body clearance (Cl) of 0.19 +/- 0.042 l/kgh, which is related to a long elimination half-life (t(1/2beta) = 4.74 +/- 0.8 h and 5.47 +/- 1.33 h i.v. and i.m. respectively). Marbofloxacin was rapidly absorbed after i.m. administration (MAT = 33.8 +/- 14.2 min) and presented high bioavailability (F = 87.9 +/- 6.0%). Pharmacokinetic parameters are not significantly different between both routes of administration (P>0.05). After marbofloxacin i.m. administration, no adverse reactions at the site of injection were observed. Serum CK activity levels 12 h after administration increased over 8-fold (range 3 15) compared with pre-injection levels, but this activity decreased to 3-fold during the 24 h follow-up period. Based on the value of surrogate markers to predict clinical success, Cmax/MIC ratio or AUC/MIC ratio, single daily marbofloxacin dose of 2 mg/kg bwt may not be effective in treating infections in horses caused by pathogens with an MIC > or = 0.25 microg/ml. However, if we use a classical antimicrobial efficacy criteria, marbofloxacin can reach a high plasma peak concentration and maintain concentrations higher than MICs determined for marbofloxacin against most gram-negative veterinary pathogens throughout the administration period. Taking into account the fact that fluoroquinolones are considered to have a concentration-dependent effect and a long postantibiotic effect against gram-negative bacteria, a dose of 2 mg/kg bwt every 24 h could be adequate for marbofloxacin in horses. PMID- 12117109 TI - Characterisation of reconstituted equine cartilage formed in vitro. AB - Lesions in cartilage of equine weightbearing joints commonly result in lameness. Cell-based resurfacing techniques are currently being developed for human and veterinary applications. Biopsies of stifle joint cartilage (1 g) were harvested aseptically and chondrocytes were isolated by sequential enzyme digestion. The cells were grown in vitro on filter inserts. Analysis of cultures 8 weeks later showed that the cells had accumulated extracellular matrix and formed a continuous layer of cartilagenous tissue as determined histologically. The cells maintained their phenotype as they synthesised type II collagen and proteoglycans similar in size to those synthesised by chondrocytes in native cartilage, but this reconstituted tissue had more sulphated glycosaminoglycan and lower collagen content than native cartilage. This experiment tests the feasibility of growing equine cartilagenous tissue in vitro. This tissue may be useful in the management of chondral injuries in the horse in a scenario where the patient donates cells, the cells are propagated under laboratory conditions and the resulting tissue becomes the therapeutic agent. PMID- 12117108 TI - Pharmacokinetics of marbofloxacin in horses. AB - Marbofloxacin is a fluoroquinolone antibiotic expected to be effective in the treatment of infections involving gram-negative and some gram-positive bacteria in horses. In order to design a rational dosage regimen for the substance in horses, the pharmacokinetic properties of marbofloxacin were investigated in 6 horses after i.v., subcutaneous and oral administration of a single dose of 2 mg/kg bwt and the minimal inhibitory concentrations (MIC) assessed for bacteria isolated from equine infectious pathologies. The clearance of marbofloxacin was mean +/- s.d. 0.25 +/- 0.05 l/kg/h and the terminal half-life 756 +/- 1.99 h. The marbofloxacin absolute bioavailabilities after subcutaneous and oral administration were 98 +/- 11% and 62 +/- 8%, respectively. The MIC required to inhibit 90% of isolates (MIC90) was 0.027 microg/ml for enterobacteriaceae and 0.21 microg/ml for Staphylococcus aureus. The values of surrogate markers of antimicrobial efficacy (AUIC, Cmax/MIC ratio, time above MIC90) were calculated and the marbofloxacin concentration profiles simulated for repeated administrations. These data were used to determine rational dosage regimens for target bacteria. Considering the breakpoint values of efficacy indices for fluoroquinolones, a marbofloxacin dosage regimen of 2 mg/kg bwt/24 h by i.v., subcutaneous or oral routes was more appropriate for enterobacteriaceae than for S. aureus. PMID- 12117110 TI - Equine oocyte maturation with epidermal growth factor. AB - Epidermal growth factor (EGF) has been shown to have a positive effect during oocyte in vitro maturation in several species. This study was performed to establish the capacity of equine oocytes to undergo nuclear maturation in the presence of EGF and to localise its receptor in the equine ovary by immunohistochemical methods. Oocytes were obtained by aspiration and subsequent scraping from equine follicles (15-25 mm diameter) and cultured in 3 different treatment groups for 36 h: control Group (modified TCM 199 with 0.003% BSA), EGF Group (TCM-199 supplemented with 50 ng/ml EGF) and EMS Group (TCM 199 supplemented with 10% v/v oestrous mare serum). Each group was divided further into 3 treatments with tyrphostin A-47, a specific tyrosine kinase inhibitor, at 0, 10(-4) and 10(-6) mmol/l. Maturation was determined as the percentage of oocytes reaching metaphase II stage at the end of the culture period. Immunohistochemical detection of EGF-receptor (EGFR) was performed using a streptoavidin-biotin method. The recovery rate and oocyte retrieval were 84.6% (recovered oocytes/follicles aspirated) and 6.55 (oocytes/mare), respectively. Treatment with EGF significantly (P<0.05) increased the incidence of metaphase II stage compared with the control group (69.4 vs. 26.9% in controls, respectively). The specific-tyrosine kinase inhibitor A-47 was effective in suppressing EGF effect on EGF-cultured oocytes; no significant differences were observed in EMS supplemented oocytes when cultured with A-47. EGF-receptor was localised in follicles, with localisation being more prominent in the cumulus than in mural granulosa cells. This finding, together with the increase of oocyte nuclear maturation rate when using EGF in culture media and the inhibition of maturation by tyrphostin A-47, suggests a physiological role for EGF in the regulation of equine oocyte maturation. The results should help successful development of assisted reproductive technology in the horse. PMID- 12117111 TI - Changes in physiological parameters in overtrained Standardbred racehorses. AB - Various changes in physiological parameters are associated with overtraining, which can be a serious problem for human and equine athletes. A 34 week longitudinal study was conducted to investigate the effects of an acute training overload on physiological parameters in 10 Standardbred racehorses. After 24 weeks of training, horses received 8 weeks of increased workload, followed by 2 weeks recovery. Horses performed a 2400 m time trial and a progressive submaximal exercise test on alternate weeks. By the end of the heavy training period, the average time for the final 1200 m of the time trial increased by 4.0% (95% probable range of true value 1.7-5.8) and peak velocity decreased by 6.9% (4.7 8.9), indicating that overtraining had occurred. Acute overtraining coincided with an increase in blood lactate concentration after the time trial and submaximal test. There were also substantial decreases in bodyweight, plasma cortisol concentration and packed cell volume after the time trial, and in the velocity at a heart rate of 200/min (V200). Parameters that showed no clear-cut change with overtraining included maximal and recovery heart rate, basal plasma cortisol, plasma and red cell volume, and markers of skeletal damage (plasma concentrations of creatine kinase and aspartate aminotransferase). Bodyweight, V200, postexercise blood lactate and plasma cortisol concentrations may all be useful for detecting acute overtraining in equine athletes. PMID- 12117112 TI - Interleukin-4 and interferon-gamma gene expression in summer pasture-associated obstructive pulmonary disease affected horses. AB - We hypothesised that horses affected with summer pasture-associated obstructive pulmonary disease (SPAOPD) react to an allergen or allergens in their summer environment that is either absent or present at lower levels in their winter environment; and that such allergens stimulate SPAOPD-affected horses to produce a different T helper lymphocyte cytokine profile from that of control horses. The primary objective of this study was to determine the cytokine mRNA profile of T helper lymphocytes obtained from summer pasture-associated obstructive pulmonary disease (SPAOPD) affected horses when 1) the horses were showing signs of disease (summer) and 2) they were in clinical remission (winter). A further objective was to determine the differences between cytokine mRNA T helper lymphocyte profiles of control and affected horses in the summer and winter seasons. Interleukin 4 (IL-4) and interferon-gamma (IFN-gamma) mRNA expression levels were increased in bronchoalveolar lavage fluid (BALF) and peripheral blood mononuclear cell (PBMC) samples of affected horses during disease expression. No significant amounts of IL-5 mRNA were detected in any of the samples. These results suggest that there is an allergic component to SPAOPD of horses and that appropriate manipulation of the immune system could offer hope for treatment and prevention of the disease in the future. Further research studies will be needed to determine the most appropriate treatments to use to alter the antigen-stimulated cytokine profile being expressed by SPAOPD-affected horses or to alter the effects that these cytokines produce. PMID- 12117113 TI - Effect of tapered normal and interval training on performance of Standardbred pacers. AB - Human athletes taper or reduce their training load before a race to enhance performance, apparently because recovery from the effects of fatigue occurs faster than the loss of fitness from the reduced training. However, there appear to be no previous studies of tapering of equine athletes. Our aim in the present study was, therefore, to investigate the efficacy of tapering with Standardbred pacers. We determined the effect of repeated cycles of tapered training on performance of Standardbred pacers. After 8 weeks of jogging and 3 x 2 week cycles of pace work, 19 horses were randomised to a taper and a control group. The taper group completed 5 consecutive 2 week cycles, each incorporating a 7 day taper; some cycles included high-intensity interval training. The control group continued with 5 more cycles of pace work. All horses completed a 2400 m individual time trial after each cycle. Peak and mean speed of the taper group were faster than those of the control group in all cycles; the differences were clear-cut in all cycles for peak speed (overall 4.4%, 95% confidence interval 1.7 to 7.1%), but only in one of the interval-training cycles for mean speed (2.4%, 0.3 to 4.7%). Four horses in the taper group were injured during interval training. Repeated tapering produces a worthwhile enhancement of performance in Standardbreds, but the addition of interval training appears to increase the risk of injury. PMID- 12117114 TI - Detection of cold-adapted vaccine-strain influenza virus using two commercial assays. AB - Because of the contagious nature of influenza virus it is necessary to identify infected individuals after the virus is introduced into a population. The aim of this study was to characterise influenza virus detection with commercially available assays after intranasal vaccinating horses with cold-adapted influenza virus. Seven horses were vaccinated and placed with 3 unvaccinated horses. Nasal secretion samples were evaluated using 2 antigen detection assays. All 10 horses were positive in the Flu OIA assay during the study period, but only one horse was positive on one sample using the Directigen Flu A assay. Horses were most likely to be positive during the first 3 days following vaccination, and several horses were intermittently positive for several days after this. Obtaining positive test results from nonvaccinated, incontact horses suggests they became infected with vaccine-strain virus that was shed by vaccinated horses. These results are important for the correct interpretation of influenza antigen detection tests in situations when this modified-live intranasal vaccine has been used. PMID- 12117115 TI - Intradermal skin testing in Icelandic horses in Austria. AB - Icelandic horses in Austria are commonly affected by an allergic inflammatory skin disease recurring during the summer seasons, which shares characteristic features with Culicoides hypersensitivity. However, the causative agents have not yet been identified. Therefore, intradermal skin testing (IDST) with a standardised extract of Culicoides variipennis and 21 other allergens relevant within Austria was performed in 81 Icelandic horses. All horses included into the study were treated regularly with ivermectin and had no history of administration of anti-inflammatory drugs. Forty-three of these horses were affected by summer seasonal recurrent dermatitis (SSRD). No history or signs of any other disease were evident in any horse. Pruritic dermatitis due to ectoparasites, bacteria and dermatophytes were ruled out by means of fungal culture, skin scraping and biopsy. Culicoides variipennis antigens evoked a positive cutaneous reaction in 1 of 38 normal and 3 of 43 SSRD horses at the proposed dilution of 1:50,000 or 1:25,000, and in 24 of 38 normal and 13 of 43 SSRD horses at a dilution of 1:10,000. Furthermore, no significant differences in onset or intensity of skin reactions to the 21 other allergens, including pollens, moulds, mites and insects, except deerfly and horsefly, were obvious between the 2 groups. Efficiency (percentage of correct results) for the used antigens in the skin test was 0.47-0.60. Maximal sensitivity was 0.51. Altogether, 38 of 43 SSRD horses and 28 of 38 normal horses were positive 4 h after allergen administration. The divergence between IDST results and manifestation of clinical signs found in this study underlines the difficulties associated with establishing a skin test protocol in horses within a geographic area. Whether the outcome of this study would have been influenced significantly by using Culicoides spp. present in Austria has to be clarified in future research. PMID- 12117116 TI - Density and binding characteristics of beta-adrenoceptors in the normal and failing equine myocardium. AB - Beta-adrenoceptors are important regulators of cardiac function and their characteristics are known to change in human and canine diseased myocardium. This study aimed to determine the density and subtypes of beta-adrenoceptors in the normal and failing equine ventricular myocardium. Membrane preparations of the left papillary muscles were incubated with increasing concentrations of the nonselective beta-adrenoceptor antagonist [3H]-CGP12177. Saturable and reversible binding of [3H]-CGP12177 to myocardial membranes was demonstrated with Kd values (+/- s.d.) of 0.49 +/- 0.40 and 0.43 +/- 0.22 nmol/l and Bmax values of 93.4 +/- 20.5 and 110.0 +/- 21.2 and fmol/mg protein for normal (n = 19) and heart failure (n = 10) tissues, respectively. Heart failure had no significant effect on the density of ventricular beta-adrenoceptors. The cardiac beta-adrenoceptors were further characterised by studying displacement of [3H]-CGP12177 (0.6 nmol/l) with the beta1-selective antagonists CGP20712A and the beta2-selective antagonist ICI118.551. In normal ventricular muscle, CGP20712A was 26 times more potent than ICI118.551 (Ki values 30.4 +/- 24.8 and 814.1 +/- 485.2 nmol/l, respectively). In heart failure cases, CGP 20712A curves were monophasic with a Ki value of 45.6 +/ 39.7 nmol/l. ICI 118.551 curves were biphasic in 5 horses where 11-31% of the cardiac beta-adrenoceptors had a high affinity for ICI 118.551. These data suggest that the normal equine ventricular myocardium possesses predominately beta1-adrenoceptors, with no evidence for co-existence of a significant population of beta2-adrenoceptors. The density of beta-adrenoceptors did not appear to change in heart failure, but the appearance of receptors with a high affinity for ICI118.551 may suggest that, in some cases, heart failure increases the expression of beta2-adrenoceptors in equine ventricular myocardium. This study provides an insight into the role of the adrenergic system in heart disease in the horse. Further studies in this area are warranted. PMID- 12117117 TI - Deslorelin acetate (Ovuplant) therapy in cycling mares: effect of implant removal on FSH secretion and ovarian function. AB - Following induction of ovulation with deslorelin acetate (Ovuplant), gonadotrophin concentrations are reduced in the subsequent cycle, leading to increased interovulatory intervals in some mares. This study determined whether implant removal after 2 days prevented the decrease in gonadotrophin concentrations and follicular growth during the ensuing cycle. Twenty-four mares were randomised equally into 3 groups. Group 1 ovulated spontaneously, Groups 2 and 3 received the deslorelin implant to induce ovulation. Two days after treatment, the implant was removed from Group 3. On Day 10 postovulation, FSH was lower (P = 0.009) in Group 2, but not different between Groups 1 and 3. Follicular diameter on Day 14 was less (P<0.05) in Group 2 (19.0 +/- 2.1 mm) than in Groups 1 and 3 (36.6 +/- 2.5 and 30.5 +/- 2.0 mm, respectively). Interovulatory interval was longer (P<0.05) for Group 2 (25.8 +/- 2.9 days) compared to Groups 1 and 3 (18.5 +/- 0.7 and 19.4 +/- 0.3 days, respectively). Removal of the deslorelin implant eliminated the decreased FSH secretion and the increased interovulatory interval associated with implant administration. Therefore, it is recommended that the implant be removed after ovulation is detected to prevent the occurrence of a prolonged interovulatory interval. PMID- 12117118 TI - The arthroscopic approach and intra-articular anatomy of the equine temporomandibular joint. PMID- 12117119 TI - Right diagnosis, wrong prescription. PMID- 12117120 TI - Are Americans closer than we think to national health insurance? PMID- 12117121 TI - The most expensive medical conditions in America. AB - This study uses a nationally representative survey to identify the most expensive conditions in the United States and to examine the association between spending and disability. The most expensive conditions at a population level were ischemic heart disease and motor vehicle accidents; at the per capita level they were respiratory malignancies. There was not a significant association between rank order of treatment costs and disability; the conditions with the greatest disability relative to expenditures were mood disorders, chronic obstructive pulmonary disease, and arthropathies. We use the findings to discuss the role for cost-of-illness and burden-of-disease estimates in setting priorities. PMID- 12117122 TI - Health care spending during 1991-1998: a fifty-state review. AB - Health care spending varies considerably across states. Spending per person ranged from $2,731 in Utah to $4,810 in Massachusetts in 1998, with Medicaid's share of total health care spending rangingfrom 9.1 percent in Nevada to 31.5 percent in New York. Research has suggested many reasons for such differences, including socioeconomic and demographic factors, market forces, and diversity in practice patterns. By using consistent methodologies among states, these 1991 1998 estimates, last produced for 1991 alone, will further the understanding of these differences. PMID- 12117123 TI - Integrated delivery networks: a detour on the road to integrated health care? AB - This paper reviews the rationales and evidence for horizontal and vertical integration involving hospitals. We find a disjunction between the integration rationales espoused by providers and those cited in the academic literature. We also generally find that integration fails to improve hospitals' economic performance. We offer seven lessons from hospitals' efforts to integrate and then suggest four alternative models for achieving integrated delivery of health care services. PMID- 12117124 TI - Why is there a quality chasm? AB - Medical care seems to obtain less value from the resources it uses than other industries do, a phenomenon not limited to the United States. I explore several reasons for this, including consumers' ignorance, the rate of technological change, the widespread use of administered pricing, the difficulty of appraising a given provider's quality, and the role of the public sector with objectives other than efficiency. Although these causes suggest that the performance of medical care may always lag behind that of other industries, greater use of information technology and improved financial incentives will help to reduce the size of the quality chasm. PMID- 12117125 TI - A longitudinal perspective on health plan-provider risk contracting. AB - During the past decade many health plans adopted risk-contracting arrangements that transferred substantial financial risk and care management responsibility to physician groups and hospital-sponsored integrated delivery systems. Risk transfer arrangements are now believed to be in steep decline, but there is little empirical evidence on this topic, particularly at the local-market level. Data from the Community Tracking Study were used to examine changes in risk contracting from 1996 to 2000. A decline in reliance on risk contracting is evident in nearly all markets. However, retrenchment in risk contracting has followed different patterns ranging from refinements in the scope of risk transfer to reduced use of risk arrangements to total rejection of risk-sharing arrangements. Modified risk-transfer agreements remain viable in several markets, but continued refinement in the nature and scope of risk sharing will be necessary. PMID- 12117126 TI - Pharmacogenetic challenges for the health care system. AB - Pharmacogenetics--the effect of genotype on drug response--holds the promise of safer and more effective drug therapy. Genetic tests would be routinely given to patients prior to prescription of a drug, with therapeutic decisions based on the patient's drug-response profile. This paper examines the operational changes and the ethical, legal, and policy challenges that pharmacogenetic medicine poses for key actors in the health care system. Adaptation by drug companies, regulatory agencies, physicians, patients, insurers, and public funding agencies will be necessary to integrate pharmacogenetic medicine into health care. PMID- 12117127 TI - Between strangers: the practice of medicine online. AB - An emerging consensus supports online communication between patients and physicians in an existing relationship to improve the quality, timeliness, and efficiency of medical care. Patients are also seeking medical care online from physicians they have never met, ranging from one-time interactions for a second opinion to psychotherapy. These practices call for a new regulatory paradigm to ensure accountability, establish acceptable parameters for online medical practice, and distinguish online health care delivery from online health information. The new patient-physician encounters also challenge the medical profession and society to reexamine core assumptions that define medical practice and the patient-physician relationship. PMID- 12117128 TI - Computerized physician order entry systems in hospitals: mandates and incentives. AB - Concerns with health care quality and medical errors are evident in media reports and research studies. A number of studies have demonstrated that computerized physician order entry (CPOE) can reduce medication error rates. In response, the California government and the Leapfrog Group have called for hospitals to implement CPOE for medications. However, few hospitals now use CPOE. Barriers include the large investment needed and the state of commercial CPOE systems. We argue that government, employers, and insurers should share the costs of CPOE and should fund further research into its benefits and means of implementation. PMID- 12117129 TI - Patients in conflict with managed care: a profile of appeals in two HMOs. AB - Despite speculation about the nature of disputes between managed care enrollees and their health plans over benefit denials, little empirical information exists about the details of such disputes and how they are actually handled. In this study we profile more than 11,000 appeals lodged between 1998 and 2000 by enrollees at two of the nation's largest health maintenance organizations (HMOs), to shed some preliminary light on the vast terrain of enrollee appeals. As many as half of appeals involved requests for reimbursement for costs of services already obtained ("retrospective" appeals), as opposed to services sought ("prospective appeals"). Enrollees won 36 percent of prospective appeals at Plan 1 and 70 percent at Plan 2, compared with 89 percent and 78 percent, respectively, of retrospective appeals. The success rate among retrospective appeals involving emergency room services--95 percent at both plans--was particularly striking. PMID- 12117130 TI - Transmission of financial incentives to physicians by intermediary organizations in California. AB - Many U.S. physicians participate in provider-sponsored organizations that act as their intermediaries in contracting with managed care plans, particularly where capitation contracts are used. Examining a survey of 153 intermediary entities in California, we trace the cascade of financial incentives from health plans through physician organizations to primary care physicians. Although the physician organizations received the vast majority (84 percent) of their revenues through capitation contracts, most of the financial risk related to utilization and costs was retained at the group level. Capitation of primary care physicians was common in independent practice associations (IPAs), but payments typically were restricted to primary care services. Thirteen percent of medical groups and 19 percent of IPAs provided bonuses or withholds based on utilization or cost performance, which averaged 10 percent of base compensation. PMID- 12117131 TI - National trends in use of medications in office-based practice, 1985-1999. AB - Increases in physician office visits involving the use or prescribing of a drug were observed between 1985 and 1999 using data from the National Ambulatory Medical Care Survey. The prescription rate increased from 109 to 146 prescriptions per 100 visits. Growth in drug mention rates for specific therapeutic classes varied by patients' age. The rate of multiple prescriptions per visit rose 39 percent. Similar-size increases were observed after differences in patients' age, number of comorbidities, source of payment, and physician specialty were controlled for. PMID- 12117132 TI - The effect of physician-owned surgicenters on hospital outpatient surgery. AB - Hospitals increasingly find themselves subject to competition from freestanding outpatient treatment facilities such as diagnostic imaging centers and ambulatory surgery centers. That competition causes hospitals particularly intense concern when the freestanding facility is owned by physicians who are on the hospital's medical staff. We find some basis for that concern. Further, this particular form of rivalry raises competitive complications that differentiate it from the standard antitrust analysis of new competitive entry. PMID- 12117134 TI - The rest is silence. PMID- 12117133 TI - Challenged to care: informal caregivers in a changing health system. AB - This report is from a 1998 national survey of 1,002 informal caregivers. Each year 23 percent of Americans provide unpaid assistance to ill, disabled, or elderly persons. Most caregivers (71 percent) do not live with care recipients. Primary caregivers provide more care of all types. Nonprimary caregivers also provide substantial care and services. Caregivers perform complex medical tasks, including medication administration, and errors can result. Few receive assistance from paid professionals or aides because of quality or financial concerns. In many areas, support and instruction could lighten caregivers' burdens and help to ensure high-quality care at home. PMID- 12117135 TI - Life but no limb: the aftermath of medical error. PMID- 12117136 TI - The curtain. What happens when doctors reach out to bereaved family members. PMID- 12117137 TI - Health insurance expansions for working families: a comparison of targeting strategies. AB - We compare three eligibility criteria for targeting health insurance expansions in working families: poverty, hourly wages, and employment in a small firm. Making pairwise comparisons among these, we find that targeting by poverty is the most effective and efficient. A poverty-based method is also the most effective way to target those lacking access to employer-sponsored insurance and those with low take-up of such coverage. When we examine the effectiveness of targeting by family type, we find that marital status and number of workers in the family make little difference once we control for the presence of children and for poverty level. PMID- 12117138 TI - Tracking Medicaid managed care in rural communities: a fifty-state follow-up. AB - This study updates a 1997 study examining implementation of rural Medicaid managed care programs. Most states operate Medicaid managed care programs for their beneficiaries, but the types of programs vary across urban and rural settings. Over the past four years the number of rural counties covered by Medicaid managed care, including fully capitated programs, has grown, although primary care case management (PCCM) remains the predominant program type in rural areas. Health plan withdrawals from rural areas have led some states with rural capitated programs to provide financial incentives or develop alternative approaches, such as enhanced PCCM programs. PMID- 12117140 TI - Public support for policies that would help people with chronic conditions. AB - According to a national public opinion survey, Americans strongly support government solutions to help people with chronic conditions and their caregivers. Such solutions include a Medicare prescription drug benefit, a tax credit for caregivers, a tax break for people who purchase private long-term care insurance, and public long-term care insurance. A majority is willing to support several of these initiatives by paying higher taxes, but our survey suggests that other initiatives may not have sufficient support to offset their incremental costs. In addition, support for these proposals varies systematically by individual characteristics, which suggests that there are different constituencies for each proposal. PMID- 12117139 TI - Preventing errors in the outpatient setting: a tale of three states. AB - Although error in medicine has received sustained policy attention recently, the problem of error in the outpatient setting has been relatively neglected. In this paper we review what is known about the incidence and nature of error-related adverse events in physicians' offices, ambulatory care facilities, and surgicenters. We then analyze policies to improve outpatient safety in New Jersey, New York, and Florida, three states that took very different paths toward this goal. Their experience suggests that accreditation, combined with particular attention to ensuring anesthesia safety, can improve quality of care for outpatients. These actions are best accomplished through proactive legislation and the development of regulations, rather than reactive responses to adverse events. PMID- 12117141 TI - Foundation approaches to U.S.-Mexico border and binational health funding. PMID- 12117142 TI - Integration and its discontents: substance abuse treatment in the Oregon Health Plan. AB - With the creation of the Oregon Health Plan (OHP) in 1994, Oregon placed its Medicaid program under a managed care system. This paper examines the managed care practices of seven health plans serving OHP enrollees between 1996 and 1998. Results indicated that the original vision of integrating substance abuse treatment services with physical care for OHP enrollees evolved into a multilayered, carved-out approach. Factors working against integration included changes in the administration and management of the chemical dependency benefit, financial losses by health plans, and lack of training and incentives for physicians to refer clients to substance abuse treatment. PMID- 12117143 TI - Recommendations on quality. PMID- 12117145 TI - A plug for quality initiatives. PMID- 12117144 TI - The quality chasm. PMID- 12117146 TI - Concern about drug reps. PMID- 12117147 TI - Drug reps are marketers, period. PMID- 12117148 TI - Influence at a higher level. PMID- 12117149 TI - Acting on health promotion. PMID- 12117150 TI - Researchers from Mars, policymakers from Venus. PMID- 12117151 TI - Health and wealth. PMID- 12117152 TI - Achieving and sustaining improved quality: lessons from New York State and cardiac surgery. AB - Since 1989 the New York State Department of Health has published annual data on risk-adjusted mortality following coronary artery bypass graft surgery by hospital and surgeon. It was the first such program in the nation and is now the most long-lived. Many hospitals were prompted by the data to improve their cardiac surgery programs, and statewide mortality fell substantially as a result. This paper examines what physicians and hospitals did in response to the data, how the market reacted, and whether this approach to quality measurement and improvement could be used more widely. PMID- 12117153 TI - Improving quality through public disclosure of performance information. AB - Despite a growing consensus that serious quality problems afflict U.S. health care, state and federal governments have done little to improve the quality of care. Proposed health insurance reforms, including a Medicare prescription drug benefit and the use of tax credits for insurance expansion, could create a mechanism for stimulating and then monitoring improvements in quality. We propose legislative requirements that any new expenditure of federal funds for health benefits be accompanied by public disclosure of performance information regarding quality, effectiveness, and safety. Such disclosure could yield diverse public and institutional benefits. PMID- 12117154 TI - HMO plan performance update: an analysis of the literature, 1997-2001. AB - This paper synthesizes results from peer-reviewed literature published from 1997 to mid-2001, on various dimensions of health maintenance organization (HMO) plan performance. Results from seventy-nine studies suggest that both types of plans provide roughly comparable quality of care, while HMOs lower use of hospital and other expensive resources somewhat. At the same time, HMO enrollees report worse results on many measures of access to care and lower levels of satisfaction, compared with non-HMO enrollees. Quality-of-care results in particular are heterogeneous, which suggests that quality is not uniform--that it varies widely among providers, plans (HMO and non-HMO), and geographic areas. PMID- 12117155 TI - Paying for national health insurance--and not getting it. AB - The threat of steep tax hikes has torpedoed the debate over national health insurance. Yet according to our calculations, the current tax-financed share of health spending is far higher than most people think: 59.8 percent. This figure (which is about fifteen percentage points higher than the official Centers for Medicare and Medicaid Services [CMS] estimate) includes health care-related tax subsidies and public employees' health benefits, neither of which are classified as public expenditures in the CMS accounting framework. U.S. tax-financed health spending is now the highest in the world. Indeed, our tax-financed costs exceed total costs in every nation except Switzerland. But the sub rosa character of much tax-financed health spending in the United States obscures its regressivity. Public spending for care of the poor, elderly, and disabled is hotly debated and intensely scrutinized. But tax subsidies that accrue mostly to the affluent and health benefits for middle-class government workers are mostly below the radar screen. National health insurance would require smaller tax increases than most people imagine and would make government's role in financing care more visible and explicit. PMID- 12117156 TI - Does U.S. tax-financed health spending really incur waste? PMID- 12117157 TI - Statesmanship in the health service. PMID- 12117158 TI - A re-appraisal of the burden of infectious disease in New Zealand: aggregate estimates of morbidity and mortality. AB - AIM: To assess the aggregate burden of infectious disease in New Zealand in terms of mortality and hospital admissions. METHODS: New Zealand mortality records for the years 1980-1998, and hospital discharges for the period 1988-2000, were re analysed using a recoding of ICD-9 codes to estimate the aggregate burden of infectious disease. The recoding scheme was modified, as in an earlier analysis, from that developed by Centers for Disease Control and Prevention. RESULTS: Following recoding, the proportion of deaths attributable to infectious disease increased from 0.7% of deaths to 6.6% of deaths. Likewise recoding of hospital discharges showed an increase in the proportion due to infectious disease from 2.2% to 12.6%, second only to "complications of pregnancy, childbirth and the puerperium". Over the study period infectious disease mortality rates have showed little decline, and there has been a nearly 60% increase in infectious disease hospital discharge rates. CONCLUSIONS: The findings confirm and extend those of an earlier study, indicating the substantial burden of disease that is still attributable to infectious disease in New Zealand. The burden remains inequitable. PMID- 12117159 TI - Venous thromboembolism in cancer patients in Christchurch, 1995-1999. AB - AIMS: To establish the incidence of venous thromboembolism (VTE) in oncology patients, describe risk factors, and assess outcome. METHODS: The clinical records of Christchurch Hospital were searched for all patients with a history of deep venous thrombosis (DVT) and/or pulmonary embolism (PE) between January 1995 and December 1999, who were registered with the Oncology Service. Follow up closed in December 2000. RESULTS: Of 7987 patients referred to the Oncology Service, 106 patients had 122 episodes of DVT and/or PE. The overall incidence rates per 1000 for VTE, PE and DVT were 13.1, 5.5 and 7.6 respectively. The recurrence rate was 226 per 1000, 50% occurring within four months. 70% of patients with VTE had one or more risk factors in addition to their malignancy: previous VTE (8%), tumour compression (7%), hospitalisation at the time of VTE diagnosis (23%), chemotherapy (25%), radiotherapy (21 %), hormonal therapy (10%), surgery (8%). 26% were smokers. 36 patients were not anticoagulated after their initial VTE because of contraindications, including brain metastasis, terminal illness or recent bleeding. 34 of these 36 patients died, 23 within three months of initial VTE, including ten of PE. The median survival time was 5.2 months for DVT and three months for PE. Survival plateaued at 22% for DVT and 16% for PE. CONCLUSIONS: VTE is a relatively common problem in cancer patients. The high recurrence rate and mortality within one year emphasise the need for better understanding of the role of predisposing factors and better guidelines for prophylaxis. PMID- 12117160 TI - Gastrointestinal decontamination in paediatric exploratory ingestions. AB - AIM: To review the effect of treatment changes in paediatric exploratory ingestion at Christchurch Hospital. METHODS: We carried out a retrospective review of paediatric patients presenting with potentially toxic ingestion during six month periods of 1994, 1996 and 1999. RESULTS: All three groups were comparable in respect to age and gender. There were minor changes in the range and proportion of substances ingested - with those in the 1999 group more likely to have taken paracetamol. In 1994, 36% of children were treated with syrup of ipecac. By 1996, only 9% were given ipecac, with 49% treated with activated charcoal. By 1999, 12% were treated with activated charcoal, while 88% received no decontamination. There was a lower admission rate in the 1999 group with no overall change in outcome. CONCLUSIONS: It is rare for paediatric exploratory ingestions to result in significant toxicity. Gastrointestinal decontamination should not be routinely used in these patients as the risk of the procedure may outweigh the risk of the poison exposure. PMID- 12117161 TI - Review of developments in colorectal surgery in New Zealand. PMID- 12117162 TI - Appendiceal diverticulitis and actinomycosis. PMID- 12117163 TI - "Let's talk of graves, of worms and epitaphs". PMID- 12117164 TI - Retention of body parts: reflections from anatomy. PMID- 12117165 TI - Assessment of snorers in primary care: straight path to treatment. AB - Habitual snoring needs to be taken seriously, both as a symptom of other sleep disorders and as a condition in its own right. GPs approached by patients with problematic snoring face a dilemma regarding whether (and to which service) those patients should be referred for a specialist opinion. Using the Sparks Chart, snoring patients can be grouped according to the two symptomatic dimensions of excessive daytime sleepiness and nocturnal hypoxaemia. We believe that the approach outlined in this article offers GPs a coherent and pragmatic guideline for referring and/or managing problematic snoring by using a simple questionnaire and pulse oximetry. The method has the potential to improve primary and secondary liaison. Most importantly, it offers patients a straight path to treatment. PMID- 12117166 TI - Capitation funding of primary care services: principles and prospects. PMID- 12117167 TI - PHARMAC and availability of pharmaceuticals. PMID- 12117168 TI - Anticoagulation and non-traumatic splenic rupture. PMID- 12117169 TI - Supervision of junior doctors. PMID- 12117170 TI - Depression in patients in general practice. PMID- 12117171 TI - What price accountability? PMID- 12117172 TI - The future of New Zealand's public health system. From top-down to bottom-up? PMID- 12117173 TI - Improved clinical management of retinoblastoma through gene testing. AB - AIMS: To investigate the relative benefits of retinoblastoma gene testing over conventional ophthalmological screening methods in a New Zealand setting, and to determine the importance of tumour material in resolving germline status. METHODS: Three cases of gene testing are described to illustrate the clinical advantages over conventional ophthalmological screening. To determine the role of tumour material in resolving germline status, 24 New Zealand families were tested, of which tumour material was available for eight. RESULTS: In the three cases reported, we found genetic testing of the RB1 gene resulted in clinically significant benefits and cost savings. When fresh tumour was available for high molecular weight DNA extraction, germline status was resolved in 8/8 (100%) cases. In these cases tumour mutations were not present in the corresponding peripheral blood DNA, indicating that the tumours were sporadic. In the absence of tumour DNA, mutations were identified in only 8/13 (62%) heritable cases. Germline status remains unresolved in all of the three cases of unilateral tumour without a family history or tumour DNA. CONCLUSIONS: Our experience indicates that retinoblastoma gene testing has significant benefits to the affected individuals and their families in New Zealand. Moreover, DNA extracted from fresh tumour allows retinoblastoma germline status in most cases to be defined. Without rumour material, the germline status of potentially sporadic cases will remain undetermined since the absence of detectable RB1 coding region mutations does not exclude all possible mutations in the RB1 gene, which is too large for DNA analysis. A lack of conclusive results will mean that infants will be subjected to the unnecessary inconvenience of surveillance under general anaesthesia. PMID- 12117174 TI - Acute gastroenteritis associated with seafood privately imported from the Pacific Islands. AB - AIMS: To investigate a potential link between consumption of food privately imported from the Pacific Islands and presentation with acute gastroenteritis to Middlemore Hospital Emergency Department. METHODS: This was a three month prospective observational case study that included patients aged greater than fifteen years presenting with acute gastroenteritis and a history of food privately imported from the Pacific Islands. Data included case demographics, symptoms, island of food origin and food type. Stool and blood samples were collected and analysed. RESULTS: Of 358 patients who presented to Middlemore Emergency Department during the study period with gastroenteritis, 34 (9.4%) had a history of consumption of food privately imported from the Pacific Islands. The seafood came from Tonga (23 cases), Samoa (10 cases) or Niue (1 case). The implicated seafood was shellfish (28 cases), jellyfish (2 cases), fish intestine (2 cases), seaweed or seaslug (1 case each). Fourteen patients (41%) provided stool samples; all were culture positive for Vibrio parahaemolyticus (VPH). CONCLUSIONS: This case series confirms a link between acute VPH gastroenteritis and consumption of seafood privately imported from the Pacific Islands. A number of public health initiatives to reduce the burden of VPH gastroenteritis among Auckland's Pacific Islanders have commenced. The Ministries of Health, Agriculture and Forestry are considering tighter controls or banning food privately imported from the Pacific Islands. PMID- 12117175 TI - Could laboratory-based notification improve the control of foodborne illness in New Zealand? AB - AIMS: To estimate the completeness and timeliness of notifications of seven potentially foodborne diseases in Auckland. METHODS: The diseases audited were shigellosis, salmonellosis, campylobacteriosis, yersiniosis, listeriosis, hepatitis A and verocytotoxigenic (VTEC) E. coli infections. Hospital and community laboratory-confirmed cases for the calendar year 2000 were audited against those notified to the Auckland Regional Public Health Service. Cases were matched on disease, name, date of birth, gender and National Health Index number. RESULTS: There were 3182 laboratory-confirmed cases of the seven diseases identified of which 77% had been notified to the Auckland Regional Public Health Service. The proportion of laboratory-confirmed cases notified ranged from a 65% for hepatitis A to 100% for VTEC infection. The median delay between laboratory confirmation and practitioner notification was two days. Notification of all laboratory-confirmed cases would have resulted in an estimated 145 additional investigations in the year 2000. CONCLUSION: A change to laboratory-based notification could improve public health investigation and control of foodborne disease in New Zealand. PMID- 12117176 TI - Infant bed-sharing among Pacific families in New Zealand. AB - AIM: To describe infant bed-sharing among Pacific families in New Zealand. METHODS: The data were gathered as part of the Pacific Island Families: First Two Years of Life (PIF) Study in which 1376 mothers were interviewed when their infants were six-weeks-old. Maternal reports of infant bed-sharing practices were assessed by questions about infant sleep location and the number of people who usually shared a mattress with the infant. RESULTS: Over half of the mothers (54.9%) reported that their infants shared a mattress with other people, 44.2% sharing with one other person, the remainder sharing with two or more people. Of the bed-sharing infants, 4.7% slept on a mattress on top of the bed, and 4.7% only slept part of the night in the shared bed. CONCLUSIONS: Together with effective information delivery, the educational and housing issues that many Pacific families in New Zealand face need to be addressed so that parents can make informed decisions about infant care practices. PMID- 12117177 TI - Auckland paediatric liver transplant experience 1990-2000. AB - AIMS: New Zealand is establishing its own Paediatric Liver Transplant Service. However there have been no readily available data on the experience of New Zealand paediatric transplant recipients to date. The aim of our study was to determine numbers and indications for transplant at present, current outcomes and to estimate the likely demand for the service in the future. METHODS: A retrospective search of computerised records was performed on children cared for at Starship Hospital from 1990 to 2000. RESULTS: Seventeen children received eighteen transplants. The indication for transplantation was biliary atresia in the majority of patients (11/17, 65%). A higher proportion of Maori and Pacific Island children received transplants than would be expected from their proportion in the population (59 vs 29%, p<0.01). Significant and often multiple complications occurred post transplantation in the majority of children, but overall outcomes were good. CONCLUSIONS: A New Zealand Paediatric Liver Transplant Program is likely to perform about six transplants per year. PMID- 12117178 TI - Can cancer centres in New Zealand help the cancer registry generate survival data? A pilot study in prostate cancer. AB - AIMS: One of the current limitations of reports issued by the New Zealand Cancer Registry (NZCR) is that the only measure of the success of treatment is provided by the mortality ratio. A pilot study was therefore carried out to see if collaboration between cancer centres and the NZCR might allow the generation of more meaningful survival data that could be used for the audit of treatment outcome. METHODS: Clinical details of patients seen at the Wellington Cancer Centre (WCC), in whom a diagnosis of prostate cancer was made in 1997, were provided to the NZCR. These details were matched with registration and mortality data held by the NZCR. RESULTS: WCC records identified 82 patients who were diagnosed with prostate cancer in 1997. Of these, the NZCR registered 60 (73%) in 1997, 3 (4%) prior to 1997, and 14 (17%) after 1997. Five patients (6%) were not registered at all. In the cohort of 82 patients, 17 (21%) had subsequently died. Of these, 11 (65%) had been treated with palliative intent, and six (35%) with radical intent. Of those patients treated radically, three had died of prostate cancer and three of other causes. CONCLUSIONS: Cooperation between Cancer Centres and the NZCR would allow the NZCR to generate useful survival data. This could help evaluate the impact on survival of specific treatments and interventions, such as screening programs. Regional variations in outcome could be detected. The exercise is feasible, without compromising patient confidentiality. PMID- 12117179 TI - Bioterrorism in the Northern Hemisphere and potential impact on New Zealand. AB - Given the historical evidence, and the characteristics of biological weapons, it appears unlikely that terrorists will use these weapons to produce mass casualties. Yet this terrorist threat will continue to be a concern while many countries still have bioweapon programmes, with advances in genetic engineering, and while determinants of terrorism persist around the world (eg, unresolved conflicts, poverty, inequality and environmental degradation). Terrorist use of smallpox, pneumonic plague and genetically engineered pathogens in the Northern Hemisphere could lead to imported cases reaching New Zealand and some risk of ongoing disease outbreaks. However, a range of disease control measures are available that could substantially limit the size of any resulting outbreaks. The risk of terrorist use of bioweapons needs to be considered in the context of the more important risk of pandemic influenza on New Zealand, the many thousands of preventable deaths in each year in this country (eg, from smoking and physical inactivity), and the current epidemic of meningococcal disease. Nevertheless, attention needs to be given to the primary prevention of terrorism and to preparatory measures that improve the country's public health infrastructure. PMID- 12117180 TI - Enduring powers of attorney. PMID- 12117181 TI - And now Auckland. PMID- 12117182 TI - Push enteroscopy: introduced where? PMID- 12117183 TI - Screening mammography: proven benefit, continued controversy. AB - Screening mammography, despite its limitations, remains the best means for diagnosing breast cancer in asymptomatic women. Regarding the continuing controversies concerning the age at which screening should start, evidence supports beginning regular screening at age 40 in women at average risk . Similarly, evidence suggests that the screening interval should be yearly, especially in younger women. Rather than an arbitrary age at which screening should stop, the decision on screening elderly women should be made on an individual basis, taking into account level of health and life expectancy. More work needs to be done on determining the optimum screening strategies for high risk women. As to the interpretation of screening mammography, a certain level of observer variability and of false-negative and false-positive readings are inherent in the process. These should be kept to a minimum through efforts by the interpreting radiologist to improve performance through auditing of individual results and continuing education. The impact of double reading and computer-aided detection in the interpretation of screening mammograms warrants further evaluation in terms of efficacy and cost-effectiveness. Despite these continuing controversies, mortality from breast cancer in the United States has been decreasing steadily for the past 25 years. The magnitude of the decrease has been reported to range from 8% to 25%. Although some of this decrease may be attributable to improvements in the treatment of breast cancer, early detection through screening mammography has undoubtedly played a role in this mortality reduction. The controversies that surround the issue of screening should not detract from the fact that screening mammography has proved to save lives. PMID- 12117184 TI - Breast imaging reporting and data system (BI-RADS). AB - The Breast Imaging Reporting and Data System (BI-RADS) lexicon was developed by the American College of Radiology to standardize mammographic reporting. The BI RADS lexicon defines terms to describe abnormalities on mammograms, and it defines final assessment categories that are predictive of the likelihood of malignancy. Although the lexicon is clinically useful and facilitates communication and research, there is still substantial interobserver variability in its application. Lexicons for breast sonography and breast MRI are in progress. PMID- 12117185 TI - Ultrasound for breast cancer screening and staging. AB - The question then arises whether and for whom BWBS should be recommended. As yet there are no scientific criteria on which to base an answer, and the examination should not be considered the standard of care until its benefits can be established prospectively. We know that mass screening mammography will detect occult cancers in two to seven of every 1000 women screened, depending on patient age and whether the screens are prevalence or incidence examinations. Should we expect a similar yield for survey US? Kopans commented that Kolb's cancer detection rate was lower than would be expected from a mammographic prevalence screen. This was not a reasonable comparison. These women all had negative findings on screening mammography and would normally be told to have repeat screening mammography 1 year later. Kolb's cancer detection rate using US was comparable to a mammographic incidence screen, so the cancer diagnoses of these fortunate women were advanced by 1 year. To maximize the yield, it is obvious that US has little to offer over mammography in women with fatty breasts because mammography is less likely to be falsely negative. The group of patients in whom incidental cancers would be expected to be found more commonly are those with dense breasts who also are at higher-than-average risk either because of a previous personal history of breast cancer (Fig. 2) or a significant family history. Because it would be impractical to consider BWBS for all women with radiographically dense breasts, it would be useful to know what its potential yield would be in the relatively smaller group of high-risk patients. Annual mammography remains the standard of care for breast cancer screening. However, in our practice in Vancouver, I suggest that high-risk women undergo mammography and US annually, recognizing that this goes beyond the standard of care. Instead of having both examinations simultaneously, I recommend that they alternate the two modalities at 6-month intervals. Theoretically, this could increase lead-time in the detection of occult cancers. The usefulness of this approach remains to be determined. BWBS for staging in women known to have breast cancer has tremendous promise and should be considered for any breast cancer patient with dense breast tissue in whom the finding of additional unsuspected foci would change the planned management. The cost of implementation would be substantial but considerably less than staging MRI. A large-scale study comparing these two modalities is needed, including assessment of the impact of identifying additional mammographically occult lesions on breast cancer mortality. PMID- 12117186 TI - Breast cancer imaging with MRI. AB - Breast MRI is an emerging technology that may revolutionize our management of women with known or suspected breast cancer. MRI examinations should be interpreted with an awareness of the pitfalls and artifacts that can affect on image evaluation. Development of an MRI lexicon will assist by providing standardized terminology that may improve our understanding of the positive predictive value of different MRI features. To date, breast MRI has proven most useful in patients with proven breast cancer to assess for multifocal/multicentric disease, chest wall involvement, chemotherapy response, or tumor recurrence or to identify the primary site in patients with occult breast cancer. Further work is necessary to assess the utility of breast MRI in other settings, such as screening of women at high risk for breast cancer. PMID- 12117187 TI - New modalities in breast imaging: digital mammography, positron emission tomography, and sestamibi scintimammography. AB - Digital mammography, PET, and sestamibi scintimammography are three new modalities in breast imaging. DM has advantages over film-screen mammography in image storage, retrieval, and processing and may lower the recall rate. Computer aided detection may increase the sensitivity of mammographic screening without a substantial reduction in specificity. Whereas PET and sestambi scintimammography are not useful in breast cancer screening, PET may play a role in detecting nodal metastases and monitoring treatment response, and sestamibi scintimammography in selected cases may serve as an adjunct to conventional imaging. The cost effectiveness of these new modalities remains to be evaluated, but all have the potential to significantly advance the diagnosis and management of women with breast cancer. PMID- 12117188 TI - Percutaneous image-guided core breast biopsy. AB - Percutaneous image-guided core biopsy is an accurate, fast, minimally invasive, and less expensive alternative to surgery for the diagnosis of breast lesions. Percutaneous core biopsy is usually performed under stereotactic or ultrasound guidance, using an automated needle or vacuum-assisted biopsy probe. Use of percutaneous core biopsy spares the need for surgery in most women with benign disease and expedites treatment in women with breast cancer. This article reviews advantages, limitations, controversies, and future directions in percutaneous image-guided core breast biopsy. PMID- 12117189 TI - Breast imaging and the conservative treatment of breast cancer. AB - Breast conservation, where appropriate, offers effective treatment for breast cancer while preserving the breast. The increased use of mammographic screening has led to increased detection of small, curable breast cancers that are amenable to breast-conserving surgery. Mammography and other imaging modalities, such as sonography and MRI, assist in the determination of the appropriateness of breast conservation and in the differentiation of recurrence from benign sequelae of treatment. PMID- 12117190 TI - Breast imaging: a breast surgeon's perspective. AB - Many changes have occurred in the past decade in the imaging of the breast. These improvements have led to more sensitive and specific breast imaging and to the widespread use of minimally invasive biopsy techniques. They have also facilitated a closer working relationship between breast imager and surgeon and have contributed greatly to the surgeon's ability to optimally diagnose and treat breast cancer. PMID- 12117191 TI - What do we expect from imaging? AB - The objectives of imaging in gynecologic cancer include tumor detection, tumor diagnosis, staging, and follow-up. In addition, both monitoring response to treatment and differentiating tumor recurrence from post-treatment changes are important indications for imaging. In 2001 it was estimated that there would be 38,300 cases of endometrial cancer, 23,400 cases of ovarian cancer, and 12,900 cases of cervical cancer. This article reviews what information is required by the practicing gynecologist or gynecologic oncologist prior to surgery and briefly summarizes state-of-the-art imaging in answering clinically pertinent questions. PMID- 12117192 TI - Postmenopausal bleeding: value of imaging. AB - Endovaginal sonography in combination with HSG is an effective screening tool in evaluating patients with postmenopausal bleeding. Endovaginal sonography is highly sensitive for detecting endometrial carcinoma and can identify patients at low risk for endometrial disease obviating the need for endometrial sampling in this subgroup of patients. In patients with abnormal findings at sonography, a detailed morphologic analysis can be used to determine which patients can undergo blind endometrial sampling successfully versus those who would benefit from hysteroscopic guidance. In patients in whom endovaginal sonography and HSG are inadequate, MRI may provide additional information on the appearance of the endometrium, particularly in patients in whom endometrial sampling is difficult (eg, patients with cervical stenosis). PMID- 12117193 TI - Imaging of cancer of the endometrium. AB - Transvaginal US is often the initial imaging examination for women with dysfunctional (postmenopausal or intermenstrual) uterine bleeding. However, once the diagnosis of endometrial cancer has been made, contrast-enhanced MRI should be performed in patients who require multifactorial assessment (eg, depth of myometrial invasion, cervical involvement, lymph node metastasis). The results of contrast-enhanced MRI help distinguish patients who need more aggressive therapy and referral to a gynecologic oncologist from those who will do well treated by a community gynecologist. PMID- 12117194 TI - Imaging of cancer of the cervix. AB - Cancer of the endometrium is the most common invasive gynecologic malignancy in North America. Although transvaginal sonography is often the initial imaging examination in women with dysfunctional uterine bleeding, MRI offers multifactorial assessment once the diagnosis of endometrial cancer has been established. Specifically, preoperative contrast-enhanced MRI alters the likelihood ratios for myometrial invasion, which in turn affects type and extent of surgery performed. This information also helps identify patients who would most benefit from referral to a tertiary care center for treatment by a gynecologic oncologist. PMID- 12117195 TI - Detection and characterization of adnexal masses. AB - The main challenge to the radiologist is to differentiate benign from malignant adnexal masses. Both US and MRI perform well for prediction of benignity. There is less specificity for diagnosis of malignancy but features, such as papillary projections, thickened septations, and internal vascularity within nodules, aid in this differentiation. The combination of morphology and Doppler characteristics provide the most accurate US diagnosis. For sonographically indeterminate masses, MRI is useful for additional lesion characterization. Analysis of T1- and T2-weighted signal intensities for benign-appearing lesions with the addition of fat saturation for high signal on T1-weighted sequences may lead to an exact diagnosis or a narrow differential. For cases considered suspicious by TVUS, more specific diagnosis by MRI may obviate the need for surgery or otherwise change management by identification of benign etiology. PMID- 12117196 TI - Staging ovarian cancer: role of imaging. AB - Ovarian cancer is relatively common, and often presents at an advanced stage with widespread intraperitoneal metastases. The constellation of complex pelvic masses, ascites, omental cake, and other peritoneal implants is virtually diagnostic. All patients are potential surgical candidates, since suspected early stage disease is treated by a comprehensive staging laparotomy including total abdominal hysterectomy, bilateral salpingo-oophorectomy, and omentectomy. Operable advanced disease is treated by surgical debulking and adjuvant combination chemotherapy. The role of imaging is to detect and characterize adnexal masses as likely malignant, recognize unusual findings that may suggest atypical pathology, demonstrate metastases in order to prevent under-staging, and detect specific sites of disease that may be unresectable. These aims are directly related to clinical management; characterization of an adnexal mass as malignant guides appropriate surgical referral, recognition of atypical pathology such as malignant granulosa cell tumor in a young woman may be an indication for fertility-preserving surgery. Demonstration of metastatic site-assists surgical planning, and detection of unresectable disease may be an indication for neoadjuvant (ie, preoperative) chemotherapy with interval debulking rather than primary debulking with adjuvan (postoperative) chemotherapy. PMID- 12117197 TI - Imaging of the vagina and vulva. AB - The imaging evaluation of female lower genital tract cancers has undergone dramatic changes in the last two decades. Technical improvements and increased availability of cross-sectional modalities (US, CT, MR) have increased their use to such an extent that they have largely replaced more conventional imaging techniques. US is of limited value in the staging of vaginal and vulvar malignancies. CT is most useful for staging more advanced disease of the vagina and vulva. It is widely available and provides quick imaging time. CT is used in the detection and biopsy of suspected lymph nodes and metastases. MRI provides the best soft tissue contrast and is the most useful imaging modality available to evaluate carcinomas of the vagina and vulva. Future advancements in the imaging evaluation of vaginal and vulvar cancers will likely focus on functional imaging. PMID- 12117198 TI - Postsurgical pelvis: treatment follow-up. AB - Imaging for recurrence and complications of gynecologic malignancies following treatment with radical hysterectomy, chemotherapy, and radiation therapy has become an important determinant for treatment options available to patients. MR imaging and computed tomography can be used to provide evidence of limited local disease recurrence and thereby identify disease that is still potentially curable with adjuvant treatments. This article examines the imaging modalities currently used to detect recurrence and assist in making treatment changes for gynecologic malignancies and presents specific patient findings following definitive primary treatment of uterine cancer and ovarian cancer with radical hysterectomy, radiation therapy, or chemotherapy. PMID- 12117199 TI - Oral health in patients with hepatitis C virus infection: an underestimated problem? PMID- 12117200 TI - Oral health of patients with hepatitis C virus infection: a pilot study. AB - OBJECTIVES: This study examined the oral health of a cohort of hepatitis C virus (HCV) patients. In particular, the prevalence of lichen planus and xerostomia were determined. Experiences of discrimination against HCV-infected patients by their dentists were also recorded. METHODS: Forty patients infected with HCV, who were not undergoing anti-viral treatment, were examined. Patient information collected included demographic details together with patients' perception of their oral health and access to dental care since being diagnosed with hepatitis C. Both extra-oral and intra-oral examinations were conducted. Teeth present and visible caries were recorded, periodontal condition was measured using a Community Periodontal Index of Treatment Need (CPITN) probe and denture fit and hygiene were assessed where appropriate. The soft tissues were examined and lichen planus diagnosed clinically. Salivary flow rates were estimated by the Salivette system. RESULTS: The oral health of this cohort was poor. Eight patients had clinical evidence of oral lichen planus (OLP), although this was not confirmed histologically. The salivary flow rates were significantly lower (P < 0.001) than in previously reported healthy controls. Of the 15 (37.5%) regular dental attenders, two had encountered problems accessing dental care. CONCLUSIONS: Chronic hepatitis C patients have significant oral health needs. More effective oral health education is required for both HCV-infected patients and their carers, including dental practitioners. PMID- 12117202 TI - Molded bone augmentation by a combination of barrier membrane and recombinant human bone morphogenetic protein-2. AB - OBJECTIVES: To provide the histological background to a new method of local bone augmentation, we examined the events occurring beneath a barrier membrane applied with recombinant human bone morphogenetic protein-2 (rhBMP-2). MATERIALS AND METHODS: The effects on bone augmentation of rhBMP-2, applied with a membrane mold (BMP-Memb), over surgically-induced bone defects in rat calvaria were examined histologically, and the results compared with those from application of rhBMP-2 (BMP) alone, or of a molded membrane (Memb) alone. RESULTS: At postoperative week 2, the BMP group showed the most marked bone formation. However, the bone diminished in size by week 8. The Memb group showed slow but continuous bone formation by week 8. In the BMP-Memb group, bone filled the space in the mold at week 2, and this was maintained until week 8. Moreover, the soft tissue that had intervened between newly formed bone and the membrane in the Memb group was not evident in the BMP-Memb group, in which bone had formed directly on the membrane. CONCLUSIONS: The results suggest that the combination of rhBMP-2 and barrier membrane has advantages in producing and maintaining bone in the intended shape by inducing osteoblasts directly on the inner surface of the membrane. PMID- 12117203 TI - Apoptosis of periodontal ligament cells induced by mechanical stress during tooth movement. AB - The mechanical force generated during tooth movement creates compressed and cell free areas in the periodontal membrane. The way in which periodontal ligament cells disappear at the compressed area during tooth movement remains unclear. In the present study we examined whether periodontal ligament cells undergo apoptosis by mechanical stress during tooth movement using the terminal deoxynucleotidyl transferase (TdT)-mediated dUTP-biotin nick-end-labelling method (TUNEL). TUNEL-positive stainings of periodontal ligament cells began to appear at the compressed areas 12 h after tooth movement, and the number of those cells reached maximum at 24 h after tooth movement. Thereafter TUNEL-positive cells disappeared at 48 h, and direct and undermining bone resorption began at the same area 72 h after tooth movement. These results showed that compressed periodontal ligament cells were eliminated by apoptosis in the early phase of tooth movement. PMID- 12117201 TI - The clinical and microbiological effects of a novel acidified sodium chlorite mouthrinse on oral bacterial mucosal infections. AB - Acidified sodium chlorite mouthrinses have been shown to have equivalent anti plaque activity to those containing chlorhexidine, the current 'gold standard'. In this study, sodium chlorite mouthrinses (ASC) acidified with either malic or gluconic acids were compared to each other and with a chlorhexidine rinse and sterile water for their effect on salivary bacterial counts. Sixteen subjects participated in the study, which had a cross-over Latin square design. In a second study, a sodium chlorite/gluconic acid rinse was compared with chlorhexidine for its clinical and microbiological effects in 36 patients with oral mucosal infections. The sodium chlorite rinses acidified with malic and gluconic acids and the chlorhexidine rinse caused significant reductions in salivary bacterial counts up to 7 h after a single rinse compared with water. There were no significant differences between the three active treatments. In the mucosal infection study, there was a significant reduction in erythema in the chlorhexidine group compared with the ASC group. Patients who received the ASC rinse reported significantly less discomfort following treatment than those receiving the chlorhexidine rinse. Staphylococcus aureus counts were significantly reduced in the group who received the sodium chlorite rinse. There were no other significant differences between the treatments. ASC appears to be an effective alternative to chlorhexidine mouthrinse. PMID- 12117204 TI - Clinical study of disseminated intravascular coagulation in oral and maxillofacial regions--predictors of onset and prognosis. AB - OBJECTIVES: To examine disseminated intravascular coagulation (DIC) cases in detail to identify the predictors of onset and prognosis. STUDY DESIGN: The clinical records of eight patients with DIC were examined with regard to the patient's background, underlying disease, direct inducers, duration, contents of treatments and outcome. The factors which controlled the onset and prognosis of DIC were examined in cases receiving surgery for malignant neoplasm compared with non-DIC cases. Wilcoxon test was used for statistical analysis. RESULTS: There were seven males and one female, age range from 25 to 93 years. The underlying diseases were six malignant neoplasms, one severe infection, and one case of multiple injuries of the mandible. Seven patients had pretherapeutic complications, and six patients recovered. Reduction in platelet counts immediately after the operation revealed high values up to 36%, and patients with more than two pretherapeutic complications were more susceptible to DIC when major postoperative complications occurred. The operation time and blood loss may also have affected the outcome. CONCLUSION: Change of platelet counts, number of pretherapeutic complications, operation time and blood loss are important for predicting the onset and prognosis of DIC in the oral and maxillofacial regions. PMID- 12117205 TI - Immunohistochemical study of oral lichen planus associated with hepatitis C virus infection, oral lichenoid contact sensitivity reaction and idiopathic oral lichen planus. AB - OBJECTIVES: Oral lichen planus (OLP) is a common mucocutaneous disorder and might be associated to a possible pathogenic relationship with hepatitis C virus (HCV) infection or hypersensitivity to dental alloy. We examined the clinical and immunohistochemical features of OLP associated with HCV infection (OLP-HCV), oral lichenoid contact sensitivity reaction (OLCSR), and idiopathic oral lichen planus (iOLP). The immunohistochemical expressions of CD4, CD8, B cells, Class II major histocompatibility complex antigen (HLA-DR), S-100, HSP60, Proliferating cell nuclear antigen (PCNA) and Ki-67 were compared to study the pathogenic differences of the three OLP groups. MATERIALS AND METHODS: Three groups of OLP patients, (I) OLP-HCV patients (n = 17), (2) OLCSR patients (n = 10) and (3) iOLP patients (n = 14) were retrieved from clinical records and tissues examined immunohistochemically by the avidin-biotin-complex technique. RESULTS: The patients with OLP-HCV showed widespread lesions. The proportion of CD8+ cells was found to be significantly higher in the lamina propria of the OLP-HCV patients and a significantly lower proportion of CD8+ cells of the OLCSR patients was noticed in the epithelium or the connective tissue papillae than in the iOLP patients. There were no significant differences in either the number of CD4+ cells or B cells between the three OLP groups. No significant differences in the number of HLA-DR+ cells were found between the three OLP groups and some OLP-HCV patients showed a significant increase of S-100+ cells in the epithelium compared with iOLP patients. There were no significant differences in either the number of PCNA+ or Ki-67+ cells between the groups. The patients showed similar weak expressions of HSP60 in the three OLP groups. CONCLUSION: The different distributions of the CD8+ cells that could have functionally different roles might be related to the distinct pathogenic mechanisms in the three OLP groups. PMID- 12117206 TI - Facial swelling and gingival enlargement in a patient with sickle cell disease. AB - Sickle cell anemia is a frequent hemoglobinopathy in the Caribbean. While vaso occlusion induced tissue injury in sickle cell anemia is common in various organs, orofacial lesions are rare. A 14-year-old Afro-Trinidadian boy suffering from sickle cell anemia developed an acute facial swelling, mimicking facial cellulitis of dental origin, which was caused by sickle cell-related hemorrhage. He also exhibited gingival enlargement, considered to be an outcome of repeated hemorrhagic episodes and fibrous repair. A new finding is the presence of erythrocyte-filled intraepithelial blood vessels in the gingival epithelium. We hypothesize this phenomena is a tissue response to hypoxia that occurs in sickle cell disease. PMID- 12117207 TI - Bannayan-Riley-Ruvalcaba syndrome: report of a family. AB - Bannayan-Riley-Ruvalcaba (BRR) syndrome is a rare inherited condition. We describe the protean orofacial manifestations of this syndrome in one family and consider their management. The dental surgeon should be aware of this entity, its orofacial connotations and the possible association with Cowden's syndrome. PMID- 12117208 TI - How to manage oral inflammatory myofibroblastic tumor (inflammatory pseudotumor)? PMID- 12117209 TI - Sentinel node biopsy, lymphatic pattern and selective neck dissection in oral cancer. PMID- 12117210 TI - Surname analysis of the Corsican population reveals an agreement with geographical and linguistic structure. AB - The surname is a cultural trait that is extremely useful for historical and linguistic studies and can effectively be used as a genetic marker. In many human populations the surname is inherited in the paternal lineage, and can therefore be considered a marker for the Y chromosome. In this study, surnames were recorded from the white pages of telephone directories in current use in Corsica in 1993. All surnames present in thirteen villages scattered over the whole island and covering the main historical regions were transcribed. Surname variability was found to be higher in coastal villages, and lower in more isolated communities. The isonymy detected among the thirteen villages allowed the calculation of kinship values, visualized in a tree showing two main clusters, one referring to the northern villages and one encompassing the villages of the south. The pattern reflects the administrative division of the island, with the exception of Vico, which belongs to the southern administrative region but is geographically close to the northern villages, and Ghisoni, which belongs to the northern district but is more similar to the village of Bastelica in the southern district. The data presented here show a structure in the surname distribution that is in substantial agreement with the geographical patterns. The kinship values are consistent with a moderated gene flow among villages producing a surname structure according to the geographic features of the territory. PMID- 12117211 TI - Induced abortion in Sri Lanka: who goes to providers for pregnancy termination? AB - The sociodemographic characteristics of abortion seekers and the reasons they give for procuring termination were studied in 356 clients selected from two abortion clinics in the city of Colombo. Nearly 80% were Buddhists and about 10% were Christians. Almost all had some formal education but only 20% were employed outside the home. Over 95% were currently married and at the peak of their childbearing age. More than one-half were aged 30 years or over, while adolescents only constituted about 3%. Fourteen per cent were nulliparous and about two-thirds had one or two living children at the time of obtaining the abortion. A significantly high proportion also had a very young child. In total, the 356 women had had 1130 pregnancies, and the mean rate of abortion was 42 per 100 pregnancies. Over one-quarter had had more than one abortion and about 10% had had three or more. Almost all abortions were performed within the first trimester with a mean gestation period of 6 weeks. About one-third of the clients were using some method of contraception at the time they became pregnant. The most common reasons cited for the present abortion were 'pregnancy too soon after previous delivery', 'no more children desired' or 'curtailment of opportunity for foreign employment'. Unmarried women constitute a special group of abortion seekers who have different needs and behave differently from married women. Their needs are not currently being met by reproductive health programmes in Sri Lanka, and it is important that they should be given special attention in the future. An interesting finding is that a significant minority of the abortion seekers answered negatively to the question regarding providing medical facilities for abortions without difficulty. This underscores the ambivalence many people have to abortion. PMID- 12117213 TI - Son preference and fertility in China. AB - This paper examines the effect of son preference on the hazards of having a second and a third birth. With data from the Two-per-thousand National Sample Survey on Fertility and Contraception conducted in 1988 by the State Family Planning Commission of China, the hazard of having a second birth among 62+ thousand married women who have had a first birth, and the hazard of having a third birth among 43+ thousand married women who have had two births was examined. These two hazards (i.e. the hazard of moving from the first to the second birth, and the hazard of moving from the second to the third birth) were analysed by estimating Cox proportional hazard models. The major covariate in the first analysis is whether or not the first-born was a daughter. In the second analysis the main covariate is whether both of the first two children were girls. In both models seven covariates known to have independent effects on the transition to a second (or third) birth are controlled for, namely, whether the woman is a Han, whether she is a farmer, her age at the birth of the first (or second) child, whether she had her first (or second) birth prior to the initiation in 1979 of the one-child policy, and three dummy variables reflecting her level of education. The results show the important influence of son preference on the hazard of having another birth. PMID- 12117212 TI - The hide-and-seek game: men's perspectives on abortion and contraceptive use within marriage in a rural community in Zimbabwe. AB - This paper is based on a study aimed at understanding the perceptions of men to induced abortion and contraceptive use within marriage in rural Zimbabwe. Two qualitative methods were combined. Men were found to view abortion not as a reproductive health problem for women. Instead, they described abortion as a sign of illicit sexual activity and contraceptive use as a strategy married women use to conceal their involvement in extramarital sexual activity. Men felt anxious and vulnerable for lack of control over women. In the absence of verbal communication on sexual matters, women and men resort to what are called here 'hide-and-seek' strategies, where women acquire and use contraceptives secretly while men search for evidence of such use. It is concluded that promoting women's sexual and reproductive health requires both short- and long-term strategies. The short-term strategy would entail providing women with reproductive technology they can use without risking violence. The long-term strategy would entail understanding men's concerns and the way these are manifested. In turn this requires the use of methodologies that encourage dialogue with research participants, in order to capture their deep meanings and experiences. PMID- 12117214 TI - Trends and differentials in menarcheal age in China. AB - This study examines trends in menarcheal age of women born in China between 1950 and 1973, and explores the impact of relevant social background characteristics on the timing of first menarche. Data on recalled ages of menarche collected in the 1988 Chinese Two-per-Thousand Fertility Survey are used in a linear regression model where the covariates are transformed with the help of an Alternating Conditional Expectation (ACE) algorithm. The results indicate that a trend towards early menarche has evolved in China during recent decades. The pattern of early menarche is especially pronounced among women residing in urban areas, and those who are better educated. PMID- 12117215 TI - The effects of type of female circumcision on infertility and fertility in Sudan. AB - This study explores the association between type of female circumcision and infertility and fertility in Sudan using the 1989-90 Demographic and Health Survey. It is hypothesized that women with either Pharaonic or Intermediate circumcision would have higher infertility and lower fertility compared with women with Sunna circumcision, and that uncircumcised women would have the lowest infertility and highest fertility of the three groups. This hypothesis, a widely held assumption, proved to be largely incorrect. Though women with Pharaonic or Intermediate circumcision did have a higher prevalence of primary infertility than uncircumcised women, women with Sunna circumcision had even lower rates of primary infertility compared with uncircumcised women. This pattern prevailed in multivariate models controlling for confounding variables, where women with Pharaonic or Intermediate circumcision had significantly higher primary infertility. Moreover, though women with Pharaonic or Intermediate circumcision also had the highest prevalence of secondary infertility, once confounding covariates were controlled in multivariate models, there was no significant difference among the three groups of women. With respect to fertility, the total fertility rate was 7.6 for women with Pharaonic or Intermediate circumcision, 8.1 for women with Sunna circumcision and 8.3 for uncircumcised women. Differences in fertility were found to be insignificant when covariates were controlled. The multivariate models were estimated using logistic regression. In conclusion, Pharaonic or Intermediate circumcision may be associated with higher primary infertility while there was no evidence suggesting that either secondary infertility or fertility was associated with a woman's circumcision status. PMID- 12117216 TI - The provision of family planning services in the Caribbean. AB - This article examines the provision of family planning services in selected countries in the Caribbean. The potential impact of the funding shortfall resulting from the phasing out of funding by the International Planned Parenthood Federation (IPPF), and the strategies being adopted by the selected countries to cope with this, are considered. Stratified random sampling methods were employed to select eight Caribbean countries and a pre-designed questionnaire was administered to the agency responsible for family planning services in each country. The sample was stratified geographically to include countries from different parts of the Caribbean. The questionnaire was designed to collect information on the services provided, the name of the agency responsible for the provision of services and, where possible, the number of users of each type of service in 1998 and 1997. Vast disparities were found in the provision of family planning services in different Caribbean countries, in terms of the groups involved, the services available in each country, as well as methods of data collection and compilation. Anguilla and Bermuda were found to provide only limited family planning services, while Barbados, Jamaica and Grenada provide much more sophisticated services. A salient finding was the innovative approaches that various countries in the region have adopted to fund family planning programmes in anticipation of the phasing out of IPPF funding. The standpoint taken in the study is that countries such as Anguilla and Bermuda must strive to improve their provision of family planning services, and that they could learn from Barbados, Grenada and Jamaica, which provide much more comprehensive services. It is also concluded that, unless alternative funding sources are identified and accessed, the provision of family planning services in the Caribbean is likely to decline in the future. PMID- 12117217 TI - Estimation of inbreeding from ecclesiastical dispensations: application of three procedures to a Spanish case. AB - The inbreeding coefficient of a population, estimated from ecclesiastical Roman Catholic dispensations, results from the relative contribution of different degrees of relationships (uncle-niece, first cousin, etc.). The interpopulation comparisons of consanguinity patterns may be obscured by the fact that in 1918 the Roman Catholic Church norm regulating the closest marriageable kinship was modified, limiting the application for an ecclesiastical dispensation to relatives of third degree (second cousins) or closer. Depending on the length of the period before or after the change of regulation, coefficients and rates may differ. Deviation of frequencies for multiple marriages may also occur. The aim of the present paper is to determine how the chosen procedure based on ecclesiastical dispensations may affect results, regarding the inbreeding coefficient, the consanguinity rate, the structure of consanguinity and the close/remote kinship ratio. As a sample case, information from the Gredos mountain range (central Spain) has been used. PMID- 12117218 TI - Does early age at marriage influence gynaecological morbidities among Pakistani women? AB - This paper presents the prevalence of and investigates predictors for specific perceived gynaecological morbidities in Pakistani women. A total of 717 women were identified from eight squatter settlements in Karachi, Pakistan. Detailed information on demographics, contraceptive use and gynaecological morbidities was elicited. The perceived prevalence of uterine prolapse was 19.1% and that of pelvic inflammatory disease 12.8%. The prevalence of uterine prolapse (adjusted odds ratio 1.8; 95% confidence interval 1.0-3.0) was significantly higher among women who married at younger ages (< or = 16 years), independent of education, socioeconomic status and parity. That of pelvic inflammatory disease was significantly higher among those under 21 years of age (adjusted odds ratio 2.3; 95% confidence interval 1.1-4.8), independent of education, socioeconomic status and parity. Young Pakistani women report an immense burden of reproductive ill health, especially those who began sexual activity at an early age. PMID- 12117219 TI - Universal recommendations for some weaning practices do make sense. PMID- 12117220 TI - Validity of hair mineral testing. AB - The variance of testing was compared between the College of American Pathologists clinical survey and that of a recent review about hair mineral testing. The review suggested that the accuracy of hair mineral testing was unreliable. In general, there was a greater range of variance in the College of American Pathologists testing results. These latter results are based on laboratory testing and are used as a "yardstick" to determine if a laboratory passes or fails that analyte and are considered a "gold standard." An extract, which resulted from a method that avoided the washing step, was compared among five laboratories. Very good precision resulted, indicating that the varied washing steps used by the laboratories in a recent review were probably the source of much variance. Analysis of hair analysis seemed to yield important information in several historical or forensic cases involving Ludwig von Beethoven, Napoleon Bonaparte, ex-US-presidents Zachary Taylor and Andrew Jackson, and Charles Hall, an Arctic explorer. Several elements that were reviewed, including arsenic, cadmium, cobalt, germanium, lead, lithium, manganese, mercury, nickel, and thallium, showed relationships between body burden, dosage, and exposure or toxicity. Evidence of toxicity could not be found by measuring hair aluminum or vanadium. Chromium, selenium, and zinc seemed to have nutritional value. Ratios of hair elements with clinical importance could not be found. PMID- 12117221 TI - Effects of dietary chromium and ascorbic acid supplementation on digestion of nutrients, serum antioxidant status, and mineral concentrations in laying hens reared at a low ambient temperature. AB - This experiment was conducted to evaluate the effects of chromium (chromium picolinate, CrPic) and vitamin C (L-ascorbic acid) supplementation on the digestion of nutrients and serum concentration of some antioxidant vitamins and minerals of laying hens (Hy-Line) reared at a low ambient temperature (6.8 degrees C). One hundred twenty laying hens (32 wk old) were divided into 4 groups, 30 hens per group. The laying hens were fed either a basal diet or the basal diet supplemented with either 400 microg of Cr/kg diet, 250 mg of L ascorbic acid/kg diet, or 400 microg of Cr plus 250 mg L-ascorbic acid/kg diet. The digestibility of nutrients (DM, OM, CP, and EE) increased by the supplementation of chromium and vitamin C (p < 0.05). Supplemental chromium and vitamin C also increased serum vitamin C and E but decreased malondialdehyde concentrations (p < 0.05). Additionally, supplemental chromium and vitamin C caused an increase in the serum concentrations of Fe, Zn, Mn, and Cr (p < 0.05) but a decrease in Cu concentration. The results of the present study showed that each dietary supplement influenced most of the parameters measured in a similar way. Also, a combination of the two supplements resulted in an additive effect, and supplementing a combination of vitamin C (250 mg/kg of diet) and chromium (400 microg Cr/kg diet) may offer a potential protective management practice in preventing cold-stress-related depression in the performance of laying hens. PMID- 12117222 TI - Effects of zinc supplementation to ration on some hematological parameters in broiler chicks. AB - The aim of the study was to examine the effects of zinc supplementation on some hematological parameters. Sixty newborn male broiler chicks were utilized in the study. Zinc (Zn) was added into drinking water at levels of 0, 125, 500, and 1000 mg/kg. In the study, there was no significant difference between control and Zn supplemented groups in erythrocyte count, hemoglobin amount, hematocrit levels, total leukocyte count, and differential leukocyte % levels, but the alpha naphthyl acetate esterase ANAE(+) lymphocyte rate significantly (p < 0.05) increased in the 125-ppm Zn-supplemented group compared with the control group. In conclusion, the data obtained may be beneficial in demonstrating the effects of zinc on, at least, these parameters. PMID- 12117223 TI - An interaction between dietary silicon and arginine affects immune function indicated by con-A-induced DNA synthesis of rat splenic T-lymphocytes. AB - Sporadic reports have appeared that suggest silicon plays a functional role in immune function by affecting lymphocyte proliferation. In addition, there is also considerable interest in supplemental arginine as a modulator of immune function. Therefore, the purpose of this animal experiment was to determine the effect of supplemental compared to adequate arginine on immune function as measured by splenic T-lymphocyte proliferation in the presence of adequate or inadequate dietary silicon. The independent variables were, per gram of fresh diet, silicon supplements of 0 or 35 microg and arginine supplements of 0 or 5 mg. The basal diet contained 2.3 microg silicon/g and 7.82 mg L-arginine/g. After feeding the male rats (nine per treatment group) for 8 wk, spleen lymphoid cells were isolated and cultured with methyl-3[H]thymidine. Supplemental arginine significantly decreased Con-A-induced DNA synthesis of splenic T-lymphocytes, but the response to arginine was influenced by dietary silicon. The decreased DNA synthesis was more marked when rats were fed adequate silicon than when fed inadequate silicon. Also, when arginine was not supplemented, DNA synthesis was higher in lymphocytes from rats fed an adequate silicon diet than rats fed the inadequate silicon diet. These findings support the huypothesis that an interaction between silicon and arginine affects immune function and that inadequate dietary silicon impairs splenic lymphocyte proliferation in response to an immune challenge. PMID- 12117224 TI - Manganese accumulates in iron-deficient rat brain regions in a heterogeneous fashion and is associated with neurochemical alterations. AB - Previous studies have shown that iron deficiency (ID) increases brain manganese (Mn), but specific regional changes have not been addressed. Weanling rats were fed one of three semipurified diets: control (CN), iron deficient (ID), or iron deficient/manganese fortified (IDMn+). Seven brain regions were analyzed for Mn concentration and amino acid (glutamate, glutamine, taurine, gamma-aminobutyric acid) concentrations. Both ID and IDMn+ diets caused significant (p<0.05) increases in Mn concentration across brain regions compared to CN. The hippocampus was the only brain region in which the IDMn+ group accumulated significantly more Mn than both the CN and ID groups. ID significantly decreased GABA concentration in hippocampus, caudate putamen, and globus pallidus compared to CN rats. Taurine was significantly increased in the substantia nigra of the IDMn+ group compared to both ID and CN. ID also altered glutamate and glutamine concentrations in cortex, caudate putamen, and thalamus compared to CN. In the substantia nigra, Mn concentration positively correlated with increased taurine concentration, whereas in caudate putamen, Mn concentration negatively correlated with decreased GABA. These data show that ID is a significant risk factor for central nervous system Mn accumulation and that some of the neurochemical alterations associated with ID are specifically attributable to Mn accumulation. PMID- 12117225 TI - Body zinc distribution profile during N-methyl-N-nitrosourea-induced mammary tumorigenesis in rats at various levels of dietary zinc intake. AB - Zinc distribution is apparently altered in breast cancer patients. It is unclear if this apparent zinc redistribution is a consequence of altered zinc nutrition or tissue-specific response to breast cancer. Our objectives were to assess effects of N-methyl-N-nitrosourea-treatment and N-methyl-N-nitrosourea-induced mammary tumorigenesis on body zinc-distribution profile in rats and to assess effects of dietary zinc intake on the body zinc-distribution profile during N methyl-N-nitrosourea treatment and N-methyl-N-nitrosourea-induced mammary tumorigenesis in rats. Female Sprague-Dawley rats were assigned to zinc-deficient (3 mg/kg diet) or zinc-adequate (31 mg/kg diet) ad libitum or pair-fed group. Rats were sham treated or N-methyl-N-nitrosourea treated (50 mg/kg body weight; Experiment 1 or 40 mg/kg body weight; Experiment 2) (n = 6). In both experiments, the zinc concentration was significantly higher (6-19 times) in mammary tumor than in mammary gland. Tissue zinc concentration was essentially unaffected by N methyl-N-nitrosourea treatment and tumor bearing, but was reduced by zinc deficiency in the bone, kidney, and liver. Overall, higher mammary tumor zinc concentration and absence of zinc redistribution during N-methyl-N-nitrosourea treatment and N-methyl-N-nitrosourea-induced mammary tumorigenesis, regardless of zinc intakes, indicates zinc accumulation in mammary tumors. Because zinc is essential for growth and cancer is characterized by uncontrolled growth, this zinc accumulation suggests an involvement of zinc in N-methyl-N-nitrosourea induced rat mammary tumorigenesis. PMID- 12117226 TI - Cardiac functions and taurine's actions at different extracellular calcium concentrations in forced swimming stress-loaded rats. AB - Modulation of the sinus rate and contractile force by taurine at different extracellular Ca2+ concentrations ([Ca2+]o) was examined using rat right atria loaded with forced swimming stress. Serum concentration of corticosterone profoundly increased in stress-loaded rats as compared with native rats. The taurine level in serum also increased in stress-loaded rats, but was not changed in the different heart tissues and aorta. Heat-shock protein (HSP72) was detectable in cardiac muscles and in the lumen of cardiac blood vessels of stress loaded rats using a monoclonal antibody. Increasing [Ca2+]o (from 0.9 to 3.6 mM) enhanced the sinus rate and contractile force in a [Ca2+]o-dependent fashion in native rats, but not in stress-loaded rats. Taurine (1-20 mM) caused a negative chronotropic and inotropic effect in a concentration-dependent manner. At 1.8 mM [Ca2+]o, the negative chronotropic effect of taurine (10-20 mM) was attenuated in stress-loaded rats as compared with native rats. These results indicate that swimming stress causes a release of taurine into the serum and reduces the sensitivity to [Ca2+]o. Taurine administration might, in part, exhibit the protective actions on acute stress-induced responses. PMID- 12117227 TI - Association of different zinc concentrations combined with a fixed caffeine dose on plasma and tissue caffeine and zinc levels in the rat. AB - Because caffeine and tissue levels of Zn are closely related, the objectives of this study were to determine the changes in plasma caffeine levels over a period of 5 h when different concentrations of Zn combined with a fixed concentration of caffeine were injected into the femoral vein of rats and to determine the relationship between tissue levels of caffeine and Zn at 5 h postinjection. Rats were divided into three groups: group 1, 220 microg caffeine; group 2, 220 microg caffeine + 8 microg Zn/g body weight (BW); group 3, 220 microg caffeine + 16 microg Zn/g BW. Blood from groups 1 and 3 was collected at 3 min, 30 min, 1 h, 3 h, and 5 h to determine the pharmacokinetics of caffeine. All groups were killed at 5 h. Caffeine and Zn concentrations of the brain, kidney, heart, and liver of all groups were determined. The plasma-caffeine curve in group 3 showed a lower concentration at 3 min and a slower caffeine-elimination rate during the first 3 h. Brain and kidney caffeine levels remained constant in all groups, whereas caffeine levels were increased in the heart in group 2 and in the liver in group 3. Zn concentrations in the brain and kidney were lower in group 2 compared with groups 1 and 3 and higher in group 3 compared to groups 1 and 2. Zn concentration in the heart was the same among the three groups but was increased in the liver in group 3 compared to groups 1 and 2. Therefore, we concluded that caffeine combined with Zn affects caffeine pharmacokinetics. With caffeine intake, levels of Zn (16 microg/g BW) that are slightly higher than the daily requirements (12 microg/g BW) may prevent a reduction of Zn in tissue. In addition, caffeine's effects on Zn concentration among organs are different. PMID- 12117228 TI - Study on the mechanism of cerium nitrate effects on germination of aged rice seed. AB - Attempts were made to promote germination of naturally and artificially aged rice seeds by treating them with cerium nitrate. The germination rate, germination index, and vigor index of aged rice seed were significantly increased by cerium. It was because the treatments of aged rice seed with cerium nitrate enhanced respiratory rate and activities of superoxide dismutase, catalase, and peroxidase, and decreased superoxide O2- and malondialdehyde contents that plasma membrane permeability was reduced. It was suggested that cerium be used for the seed treatment before sowing. PMID- 12117229 TI - Changes of element ratios of cultured cells from dune reed under adverse environmental conditions. AB - Dune reed, as an ecotype of reed plant (Phragmites communis Trin.), is an ideal material for studies on the adaptations of plant to environmental conditions. Scanning electron microscope, energy-dispersive X-ray analysis, and plant tissue culture techniques were used to investigate the effect of extreme temperature, salt, and polyethylene glycol-induced osmotic stress on the intracellular elements K, Na, Ca, and Cl in cultured cells from dune reed and swamp reed (as control). The results indicated that the percentages of the studied elements in dune reed cells exposed to various stresses increased or decreased obviously compared to the swamp reed cells. It has been found that a pattern of absorbing K and discharging Na exists in dune reed cells, which did not exist in swamp reed cells. The pattern is thought to be a significant physiological mechanism of the dune reed response to adverse environmental factors. In addition, the percentages of Ca and Cl in dune reed cells were also shown to increase at high temperature. The growth of cells along with their surface features under different stress conditions were observed and the results are discussed. PMID- 12117230 TI - Simultaneous analysis for multiple heavy metals in contaminated biological samples. AB - In the present study, the conditions of analysis by inductively coupled plasma atomic emission spectrometry (ICP-AES) were investigated. Twenty-six elements (Mg: 25 ppm; Sc: 10 ppm; Ti: 50 ppm; others: 100 ppm) were used as the elements interfering with selected 24 wavelengths. Consequently, the background values in 19 elements were subjected to some influences. However, all of these effects disappeared at low concentrations--less than 1 ppm of interfering elements. Next, the values from the ordinary calibration method were compared with those from the standard addition method using several biological samples. There was a discrepancy in the results obtained from both methods because of the sample, and three patterns were observed. However, no discrepancy was observed in the values for the standard reference materials using both methods. There was no significant difference between the certified values of the standard reference materials and the obtained ones by ICP. Therefore, the analytical wavelengths and the methods in the present study were suggested to be useful for ICP-AES analysis for environmental and/or biological samples. PMID- 12117231 TI - Sequential thoracentesis: minor change of zinc compared to other selected essential trace elements in the serum. AB - This study first indicates that the serum trace element Zn tends to decrease in the course of sequential thoracenteses. Other selected essential elements such as copper (Cu), manganese (Mn), molybdenum (Mo), and cobalt (Co) do not reveal loss changes in their serum levels. Therefore, Zn should be monitored in patients who undergo repeated thoracentesis. To measure the magnitude of changes in serum trace elements and the clinical relevance of potential imbalances, concentrations of the essential elements are analyzed in 57 serum/effusion pairs obtained from 5 patients (4 male, 1 female; age 28-78 yr) who underwent repeated thoracenteses as a result of recurrent pleural effusion. All patients declined other therapeutic options such as chemical pleurodesis and/or chest tube placement. The total volumes of fluid removed ranged from 2.3 to 19.3 L and the frequency of thoracentesis ranged from 6 to 15 within a period of 102-174 days. Two patients had benign pleural disease and three had malignancies. Three patients suffered from pleural effusions resulting from exudates (total protein content > 3.0 g/dL, LDH > 200 U/L), and two resulting from transudates (total protein < 3.0 g/dL, LDH < 200 U/L). All trace elements were simultaneously determined by inductively coupled argon plasma-mass spectrometry. In addition, the concentrations of the following clinically relevant parameters were analyzed by standard methods: total protein, pH, leukocyte count, lactate dehydrogenase, and glucose. PMID- 12117232 TI - Altered elemental profile as indicator of homeostatic imbalance in pathogenesis of oral submucous fibrosis. AB - Oral submucous fibrosis (OSF) is a potential precancerous condition of the oral cavity and oropharynx. The etiopathogenesis of this complex precancerous condition is still obscure. In addition to deleterious oral habits, malnutrition, and possible genetic predisposition, altered bioelemental status is also likely to play an important role in its pathogenesis. The present study analyzed 68 elements by inductively coupled plasma-mass spectroscopy in oral mucosa of normal and OSF individuals and some interesting alterations in elemental profile in the diseased tissue have been noted, indicating a homeostatic imbalance. These bioelemental alterations leading to homeostatic imbalance might be considered as an important biological event in the pathogenesis of OSF. PMID- 12117233 TI - Differences in calcium accumulation between human plantar and palmar aponeuroses. AB - To elucidate the characteristics of calcium accumulation of human plantar and palmar aponeuroses, the authors determined the calcium content of human plantar and palmar aponeuroses by atomic absorption flame emission spectrophotometry. The subjects consisted of 9 men and 14 women, ranging in age from 61 to 93 yr. In the plantar aponeurosis, the calcium content was significantly higher in the anterior and posterior parts than in the middle part. It is known that pressure distribution under the sole of a foot is higher in the anterior and posterior parts than in the middle part. The present study suggests that the accumulation of calcium in the plantar aponeurosis is related with the pressure distribution under the sole of a foot. The calcium content increased progressively with aging in the anterior part of the plantar aponeurosis, but not in the middle and posterior parts. Regarding the palmar aponeurosis, the calcium content was significantly higher in the anterior and posterior parts in comparison with the middle part. It was found that the calcium content increased progressively with aging in the posterior part of the palmar aponeurosis, whereas it did not increase significantly with aging in the anterior and middle parts. Regarding the relationship between the calcium content of the aponeuroses and the bone mineral density, a significant correlation was found between the calcium content in the anterior part of the palmar aponeurosis and the bone mineral density of the scaphoid bone. PMID- 12117234 TI - High accumulation of calcium and phosphorus in the coronary artery of the Thai in comparison with the Japanese. AB - To examine whether there were differences between races in regard to age-related changes of mineral contents and the relationships among element contents in the arteries, the authors investigated the coronary arteries of Thai and Japanese. The Thai subjects consisted of 13 men and 3 women, ranging in age from 39 to 84 yr, whereas the Japanese subjects consisted of 17 men and 9 women, ranging in age from 55 to 92 yr. After the ordinary dissections at Chiang Mai University and Nara Medical University were finished, the coronary arteries were resected and the element contents were determined by inductively coupled plasma-atomic emission spectrometry. In the Thai, an accumulation of calcium and phosphorus began to occur in the forties and increased markedly in the fifties, whereas in the Japanese, an accumulation of calcium and phosphorus began to occur in the seventies and increased markedly in the nineties. The result revealed that an accumulation of calcium and phosphorus occurred earlier in the Thai than in the Japanese. Regarding the relationships among element contents, extremely significant correlations were found between calcium and phosphorus contents, between calcium and magnesium contents, and between phosphorus and magnesium contents in both the coronary arteries of the Thai and the Japanese. As far as the coronary arteries with a very high accumulation of calcium and phosphorus, the mass ratios of magnesium to calcium and phosphorus were lower in the coronary arteries of the Thai in comparison with the Japanese. PMID- 12117235 TI - Compositional changes of the aortic valve similar to the artery with aging. AB - To elucidate compositional changes of the cardiac valves with aging, the authors investigated the relationships among element contents in the aortic valves, in which the accumulation of calcium and phosphorus was the highest. The subjects consisted of 10 men and 14 women, ranging in age from 65 to 102 yr. Extremely significant direct correlations were found among the contents of calcium, phosphorus, magnesium, sodium, and zinc in the aortic valves. In contrast, significant inverse correlations were found between the contents of sulfur and the other elements, such as calcium, phosphorus, magnesium, and sodium. It should be noted that these correlations present in the aortic valves were very similar to those in the arteries, especially those in the thoracic aortas. The changes of the mass ratios of Mg/Ca and Mg/P in the aortic valves were similar to those in the thoracic aortas. As calcium and phosphorus increased in the aortic valve, the mass ratios Mg/Ca and Mg/P decreased reversely in the aortic valve. These results suggest that the compositional change of the aortic valve is very similar to that of the thoracic aorta. PMID- 12117236 TI - Concentrations of cadmium, zinc, copper, iron, and metallothionein in liver and kidney of nonhuman primates. AB - To evaluate the species specificity of Cd accumulation and the relationship of Cd with other essential metals and metallothionein (MT), the concentrations of Cd, Zn, Cu, and Fe in the liver and kidney and the MT concentrations in the soluble fractions of the liver and kidney were determined in Cd-uncontaminated nonhuman primates (11 species, 26 individuals) kept in a zoo and two wild-caught Japanese macaques. The compositions of metal-binding proteins in the soluble fractions were also investigated by high-performance liquid chromatography (HPLC). The hepatic Cd concentration was 0.03-14.0 microg/g and the renal Cd concentration was 0.35-99.0 microg/g, both varying greatly and being higher in nonhuman primates, which were more closely related to man. The hepatic Zn concentration was 24.0-176 microg/g and the renal Zn concentration was 13.5-138 microg/g, showing 7- to 10-fold differences, and a correlation (r=0.558, p<0.01) was found between renal Zn and renal Cd concentrations. It was proved that in the liver, MT is more closely correlated with Zn (r=0.795, p<0.001) than with Cd (r=0.492, p<0.01) and that in the kidney MT is correlated with both Cd (r=0.784, p<0.001) and Zn (r=0.742, p<0.001). HPLC analysis of metals bound to MT-like protein in chimpanzees, de Brazza's monkeys, and Bolivian squirrel monkeys showed that more than 90% of Cd in both the liver and kidney, approx 40% of Zn in liver and 28-69% of Zn in kidney were bound to MT-like protein. The higher percentage Zn was bound to high-molecular protein. PMID- 12117237 TI - Survival differences of Escherichia coli O157:H7 strains in apples of three varieties stored at various temperatures. AB - Differences in survival and growth among five different Escherichia coli O157:H7 strains in three apple varieties were determined at various temperatures. Jonathan, Golden Delicious, and Red Delicious apples were wounded and inoculated with E coli O157:H7 strains C7929 (apple cider isolate), 301C (chicken isolate), 204P (pork isolate), 933 (beef isolate), and 43890 (human isolate) at an initial level of 6 to 7 log CFU/g. The inoculated apples were stored at a constant temperature of 37, 25, 8, or 4 degrees C or at 37 degrees C for 24 h and then at 4 degrees C, and bacterial counts were determined every week for 28 days. By day 28, for Jonathan apples at 25 degrees C, the apple isolate counts were significantly higher than the chicken and human isolate counts. At 4 degrees C for 28 days, the human isolate inoculated into Jonathan, Golden Delicious, and Red Delicious apples was present in significantly smaller numbers than the other strains. The apple isolate survived significantly better at 4 degrees C, yielding the highest number of viable cells. By days 21 and 28, for apples stored at 37 degrees C for the first 24 h and then at 4 degrees C, the counts of viable E. coli O157:H7 apple and human isolates were 6.8 and 5.8 log CFU/g at the site of the wound, whereas for apples kept at 4 degrees C for the duration of storage, the respective counts were 5.6 and 1.5 log CFU/g. Our study shows that E. coli O157:H7 strains responded differentially to their ability to survive in these three apple varieties at 25 or 4 degrees C and produced higher viable counts when apples were temperature abused at 37 degrees C for 24 h and then stored at 4 degrees C for 27 days. PMID- 12117238 TI - Inactivation of Salmonella Typhimurium in orange juice containing antimicrobial agents by pulsed electric field. AB - Combinations of different hurdles, including moderately high temperatures (<60 degrees C), antimicrobial compounds, and pulsed electric field (PEF) treatment, to reduce Salmonella in pasteurized and freshly squeezed orange juices (with and without pulp) were explored. Populations of Salmonella Typhimurium were found to decrease with an increase in pulse number and treatment temperature. At a field strength of 90 kV/cm, a pulse number of 20, and a temperature of 45 degrees C, PEF treatment did not have a notable effect on cell viability or injury. At and above 46 degrees C, however, cell death and injury were greatly increased. Salmonella numbers were reduced by 5.9 log cycles in freshly squeezed orange juice (without pulp) treated at 90 kV/cm, 50 pulses, and 55 degrees C. When PEF treatment was carried out in the presence of nisin (100 U/ml of orange juice), lysozyme (2,400 U/ml), or a mixture of nisin (27.5 U/ml) and lysozyme (690 U/ml), cell viability loss was increased by an additional 0.04 to 2.75 log cycles. The combination of nisin and lysozyme had a more pronounced bactericidal effect than did either nisin or lysozyme alone. An additional Salmonella count reduction of at least 1.37 log cycles was achieved when the two antimicrobial agents were used in combination. No significant difference (P > 0.05) in cell death was attained by lowering the pH value; only cell injury increased. Inactivation by PEF was significantly more extensive (P < 0.05) in pasteurized orange juice than in freshly squeezed orange juice under the same treatment conditions. This increase might be due to the effect of the chemical composition of the juices. PMID- 12117239 TI - Inhibition of Salmonella Typhimurium and Listeria monocytogenes in mung bean sprouts by chemical treatment. AB - This study was undertaken to compare the efficacies of chlorous acid (268 ppm), sodium hypochlorite (200 ppm), and lactic acid (2%) in eliminating total mesophilic microorganisms, Salmonella Typhimurium, and Listeria monocytogenes on commercial mung bean sprouts immediately after treatment and during posttreatment refrigerated storage. Treatment with sodium hypochlorite for 10 min did not reduce the total aerobic count. However, treatment with lactic acid and chlorous acid for 10 min initially reduced the total aerobic count by 0.6 and 0.8 log CFU/g, respectively, and maintained the same level or a lower level of the total aerobic count during the storage time. Treatment with chlorous acid reduced Salmonella Typhimurium from 5.0 log to undetectable levels (<0.48 log CFU/g), and the pathogen remained undetectable over a 9-day storage period. Treatment with lactic acid resulted in an initial 3-log reduction and further reduced the number of Salmonella Typhimurium cells to undetectable levels after 3 days. For L. monocytogenes, treatment with chlorous acid resulted in an initial 5-log reduction, and treatment with lactic acid resulted in a 2-log reduction at the beginning and undetectable levels after 9 days. When chemically injured cells were investigated by the selective overlay method, no statistical difference was observed (P < 0.05) between the number of injured cells recovered following treatment with chlorous acid and the number of bacteria counted on selective media, whereas sodium hypochlorite generated more injured cells than the other treatments did. These data suggest that treatment with chlorous acid may be useful in reducing total mesophilic microorganisms, Salmonella Typhimurium, and L. monocytogenes in commercial mung bean sprouts. PMID- 12117240 TI - Relationship of cell surface charge and hydrophobicity to strength of attachment of bacteria to cantaloupe rind. AB - The cantaloupe melon has been associated with outbreaks of Salmonella infections. It is suspected that bacterial surface charge and hydrophobicity may affect bacterial attachment and complicate bacterial detachment from cantaloupe surfaces. The surface charge and hydrophobicity of strains of Salmonella, Escherichia coli (O157:H7 and non-O157:H7), and Listeria monocytogenes were determined by electrostatic and hydrophobic interaction chromatography, respectively. Initial bacterial attachment to cantaloupe surfaces and the ability of bacteria to resist removal by washing with water were compared with surface charge and hydrophobicity. Whole cantaloupes were submerged in inocula containing individual strains or in cocktails containing Salmonella, E. coli, and L. monocytogenes, either as a mixture of strains containing all three genera or as a mixture of strains belonging to a single genus, for 10 min. Inoculated cantaloupes were dried for 1 h in a biosafety cabinet and then stored for up to 7 days at 4 degrees C. Inoculated melons were washed with water, and bacteria still attached to the melon surface, as well as those in the wash water, were enumerated. Initial bacterial attachment was highest for individual strains of E. coli and lowest for L. monocytogenes, but Salmonella exhibited the strongest attachment on days 0, 3, and 7. When mixed-genus cocktails were used, the relative degrees of attachment of the three genera ware altered. The attachment of Salmonella strains was the strongest. but the attachment of E. coli was more extensive than that of L. monocytogenes on days 0, 3, and 7. There was a linear correlation between bacterial cell surface hydrophobicity (r2 = 0.767), negative charge (r2 = 0.738), and positive charge (r2 = 0.724) and the strength of bacterial attachment to cantaloupe surfaces. PMID- 12117241 TI - Molecular characterization of Salmonella spp. isolated from bulk tank milk and cull dairy cow fecal samples. AB - The consumption of meat from cull dairy cows and of raw milk has been associated with foodborne salmonellosis. This survey was conducted to establish the prevalence of Salmonella in cull dairy cow fecal samples and bulk tank milk and to determine the proportion of Salmonella-positive dairy farms (n = 30) in east Tennessee. Food and Drug Administration bacteriological analytical protocols were generally used for Salmonella isolation. Primary enrichment was performed with lactose broth, and secondary enrichment was conducted with tetrathionate broth. Eosin methylene blue, hektoen enteric, xylose lysine desoxycholate, bismuth sulfite, and brilliant green (BG) were used as isolation agars. BG agars supplemented with individual antibiotics and/or sulfur compounds were also evaluated. Six of 268 (2.24%) bulk tank milk samples and 9 of 415 (2.17%) fecal samples from 7 of 30 (25.3%) dairy farms were Salmonella-positive. Most isolates (11 of 15) were obtained between September and December. Salmonella isolates were further characterized using polyvalent somatic O Salmonella antiserum, o nitrophenyl-beta-D-galactopyranoside (ONPG), and Analytical Profile Index (API) 20E strips for Enterobacteriaceae. Serological evaluation of presumptive positive Salmonella isolates resulted in substantial numbers of false positives (41.2%). ONPG and API 20E tests enabled further biochemical distinction of the majority of Salmonella spp. from Salmonella Arizonae and closely related members of Enterobacteriaceae like Citrobacter youngae. Pulsed-field gel electrophoresis of SpeI-digested Salmonella DNA was used to subtype isolates. The isolates grouped into four clusters. The baseline information generated in this survey is being used to develop preharvest pathogen reduction programs on selected farms. PMID- 12117242 TI - Fate of field-isolated Escherichia coli O157 in ground beef at different storage temperatures. AB - The survival of six Escherichia coli O157 strains, including five strains recently isolated from beef carcasses and strain ATCC 43895, was evaluated at 0, 1, 7, and 14 days in ground beef held at -20, 1, 4, and 7 degrees C. Only small losses in cell numbers occurred at -20 and 1 degree C; in general, cell numbers decreased during the first day of storage and then remained unchanged through day 14. At -20 degrees C, statistically significant reductions in cell numbers were observed only for strains 55AC1 and 299AB3 due to greater losses in the first day. At 1 degree C, strain 131AC1 did not decrease in cell numbers during the first day of storage, but both this strain and strain 55AC1 experienced statistically significant reductions in viable cell numbers by day 14, primarily due to losses after day 7. At 4 degrees C, after an initial loss of cell numbers for four strains, minor increases were observed for all six strains by day 14. The differences were statistically significant for strains 114AC1, 299AB3, and ATCC 43895, but were small enough to question whether they refect actual growth. When the inoculated ground beef was stored at 7 degrees C for 14 days, growth of all six strains was statistically significant, with populations increasing between 0.9 and 1.5 log10 CFU/g. This study demonstrates that there are small differences in the abilities of various E. coli O157 strains to survive and sometimes grow in fresh ground beef at cold storage temperatures, but overall these differences do not appear to be meaningful. The differences cannot be attributed to recency of isolation, since strain ATCC 43895 behaved similarly to recently isolated strains. Storage temperatures of 4 degrees C or below limited growth of E. coli O157 isolates, but did not have a noteworthy effect on survival. PMID- 12117243 TI - Survival of Campylobacter jejuni in biofilms isolated from chicken houses. AB - Campylobacter jejuni is a thermophilic and microaerophilic enteric pathogen associated with poultry. Biofilms may be a source of C. jejuni in poultry house water systems since they can protect constituent microorganisms from environmental stress. In this study, the viability of C. jejuni in biofilms of gram-positive chicken house isolates (P1, Y1, and W1) and a Pseudomonas sp. was determined using a cultural method (modified brucella agar) and direct viable count (DVC). Two-day biofilms grown on polyvinyl chloride (PVC) coupons in R2A broth at 12 and 23 degrees C were incubated with C. jejuni for a 6-h attachment period. Media were then refreshed every 24 h for 7 days to allow biofilm growth. Two-day biofilms of P1, Y1, and Pseudomonas spp. enhanced attachment (P < 0.01) of C. jejuni (4.74, 4.62, and 4.78 log cells/cm2, respectively) compared to W1 and controls without preexisting biofilm (4.31 and 4.22 log cells/cm2, respectively). On day 7, isolates P1 and Y1 and Pseudomonas biofilms covered 5.4, 7.0, and 21.5% of the surface, respectively, compared to 4.9% by W1. Viable C. jejuni on the surface decreased (P < 0.05) with time, with the greatest reduction occurring on surfaces without a preexisting biofilm. The number of viable C. jejuni determined by DVC was greater than that determined by the cultural method, indicating that C. jejuni may form a viable but nonculturable state within the biofilm. Both DVC and the cultural method indicate that biofilms enhance (P < 0.01) the survival of C. jejuni during incubation at 12 and 23 degrees C over a 7 day period. PMID- 12117244 TI - Effectiveness of chemical sanitizers against Campylobacter jejuni-containing biofilms. AB - Survival of Campylobacter jejuni in mixed-culture biofilms was determined after treatment with chemical sanitizers including chlorine, quaternary ammonia, peracetic acid (PAA), and a PAA/peroctanoic acid mixture (PAA/POA). Biofilm producing bacteria (gram-positive rods, Y1 and W1) were isolated from chicken house nipple drinkers. A meat plant isolate (Pseudomonas sp.) was also included as a biofilm producer. Two-day-old biofilms grown on polyvinyl chloride (PVC) plastic coupons in R2A broth at 12 degrees C were incubated with 10(6) CFU/ml C jejuni for 6 h to allow attachment. The coupons were then rinsed and incubated in fresh media for an additional 24 h. C. jejuni-containing biofilms were detached by vortexing with glass beads in modified brucella broth, which was then enumerated for C. jejuni on selective/differential media. The presence of biofilm enhanced (P < 0.01) the attachment and survival of C. jejuni After the 24-h incubation, only 20 CFU/cm2 of C. jejuni were recovered from the control without biofilms compared to 2,500 to 5,000 CFU/cm2 in samples with preexisting biofilms. The presence of biofilm microflora decreased (P < 0.01) the effectiveness of sanitizers against C. jejuni. Chlorine was the most effective sanitizer since it completely inactivated C. jejuni in the biofilms after treatment at 50 ppm for 45 s. C. jejuni in biofilms was susceptible to all sanitizers tested but was not completely inactivated by treatment with quaternary ammonia, PAA, or PAA/POA mixture at 50 and 200 ppm for 45 s. PMID- 12117246 TI - Transfer of persistent Listeria monocytogenes contamination between food processing plants associated with a dicing machine. AB - The possibility of the transfer of persistent Listeria monocytogenes contamination from one plant to another with a dicing machine was evaluated, and possible reasons for persistent contamination were analyzed. A dicing machine that diced cooked meat products was transferred from plant A to plant B and then to plant C. After the transfer of the dicing machine, L. monocytogenes PFGE type I, originally found in plant A, was soon also found in plants B and C. This L. monocytogenes PFGE type I caused persistent contamination of the dicing lines in plants B and C. The persistent L. monocytogenes strain and three nonpersistent L. monocytogenes strains found in the dicing line of plant C were tested for adherence to stainless steel surfaces and minimal inhibitory concentrations of a quaternary ammonium compound and sodium hypochlorite, disinfectants widely used in the dicing lines. The persistent strain showed significantly higher adherence to stainless steel surfaces than did the nonpersistent strains. The minimal inhibitory concentrations of sodium hypochlorite were similar for all strains, and the minimal inhibitory concentrations of the quaternary ammonium compound for three of the L. monocytogenes PFGE types, including the persistent PFGE type, were high. All persistent L. monocytogenes PFGE type I isolates were found in an area with high hygienic standards, with the dicing machine being the first point of contamination. These observations show that the dicing machine sustained the contamination and suggest that the dicing machine transferred the persistent L. monocytogenes PFGE type from one plant to another. PMID- 12117245 TI - Development of a technique to quantify the effectiveness of enrichment regimes in recovering "stressed" Listeria cells. AB - A rapid, reliable microwell plate method based on the most probable number (MPN) technique was used to determine the effectiveness of five enrichment regimes in the recovery and enumeration of Listeria spp. cells from five seafood products. The products tested were chosen to reflect conditions under which cells were exposed to the "stresses" associated with a variety of food-processing techniques, such as treatments involving an ethanol-based marinade, lowered pH (acetic acid), heat, sugar and salt brine (Gravilax), or frozen storage. Either Listeria monocytogenes and Listeria innocua were present in food samples as natural contaminants or L monocytogenes was added in the laboratory. Listeria repair broth (LRB), buffered Listeria enrichment broth, Listeria enrichment broth (LEB), Fraser broth, and University of Vermont modified Listeria enrichment broth were used to recover Listeria cells. The effectiveness of these enrichment regimes was found to be dependent on the type of stresses the cells had been exposed to. After exposure to ethanol, recovery of L monocytogenes cells was inhibited in enrichment regimes involving a nonselective period of resuscitation. On exposure to acetic acid, there were no significant differences (P < 0.05) between any of the regimes used. With heat-stressed cells, LRB recovered significantly fewer (P < 0.05) cells than did any other medium. On exposure to osmotic stress (elevated sugar and salt concentrations), LEB recovered the fewest cells. The largest number of cells was recovered from frozen fish (Hoki [Macruronus novazelandiae]) fillets with LRB. No single enrichment regime was consistently the most effective. PMID- 12117247 TI - Immobilized bacterial spores for use as bioindicators in the validation of thermal sterilization processes. AB - Spores of Bacillus subtilis ATCC 6051 and Bacillus stearothermophilus NCTC 10003 were immobilized in monodisperse alginate beads (diameter, 550 microm +/- 5%), and the capacity of the immobilized bioindicators to provide accurate and reliable F-values for sterilization processes was studied. The resistance of the beads to abrasion and heat was strong enough to ensure total retention of the bioindicators in the beads in a sterilization cycle. D- and z-values for free spores were identical to those for immobilized spores, which shows that immobilization does not modify the thermal resistance of the bioindicators. A D(100 degrees C) value of 1.5 min was found for free and immobilized B. subtilis spores heated in demineralized water, skimmed milk, and milk containing 4% fat, suggesting that a lipid concentration as low as 4% does not alter the thermal resistance of B. subtilis spores. Providing that the pH range is kept between 3.4 to 10 and that sufficiently low concentrations of Ca2+ competitors or complexants are present in the medium, immobilized bioindicators may serve as an efficient, accurate, and reliable tool with which to validate the efficiency of any sterilization process. The environmental factors (pH, media composition) affecting the thermoresistance of native contaminants are intrinsically reflected in the F-value, allowing for a sharper adjustment of the sterilization process. Immobilized spores of B. stearothermophilus were successfully used to validate a resonance and interference microwave system that is believed to offer a convenient alternative for the sterilization of temperature-sensitive products and medical wastes. PMID- 12117248 TI - Effects of UV irradiation on selected pathogens in peptone water and on stainless steel and chicken meat. AB - Effects of intensity and processing time of 254 nm UV irradiation on Listeria monocytogenes, Escherichia coli O157:H7, and Salmonella Typhimurium were investigated. Intensities measured at 5.08, 10.1, 15.2, and 20.3 cm from the light source were 1.000, 500, 250, and 150 microW/cm2, respectively. Intensities of 250 or 500 microW/cm2 reduced all suspended pathogen cells in peptone water about 5 log cycles after 2 min and completely inactivated L. monocytogenes and E. coli O157:H7 after 3 min by reductions of 8.39 and 8.64 log cycles, respectively. Intensities of 250 or 500 microW/cm2 also reduced (P < or = 0.05) the tested pathogens inoculated on stainless steel (SS) chips, and E. coli O157:H7 was completely destroyed at 500 microW/cm2 for 3 min. After UV treatment for 3 min at 500 microW/cm2, all selected pathogens on chicken meat with or without skin showed reduction ranges from 0.36 to 1.28 log cycles. Results demonstrated that UV irradiation could effectively decrease pathogens in peptone water and on SS but that it was less effective on chicken meat. PMID- 12117249 TI - Influence of catfish skin mucus on trisodium phosphate inactivation of attached Salmonella Typhimurium, Edwardsiella tarda, and Listeria monocytogenes. AB - This study examined the antimicrobial effectiveness of trisodium phosphate (TSP) on Edwardsiella tarda, Listeria monocytogenes, and Salmonella Typhimurium attached to catfish skin with and without mucus. Salmonella Typhimurium and E. tarda attached more readily to catfish skin than did L monocytogenes. At high inoculum levels (10(7) CFU/ml), TSP treatments (at 2 to 6%) for 10 min reduced bacterial counts of E. tarda by >2.5 to >3.3 log10 CFU per skin sample for firmly attached cells and by 3.5 to 3.6 log10 CFU per skin sample for loosely attached cells. Counts of L. monocytogenes declined by 0.6 to >1.8 log10 CFU per skin sample for firmly attached cells and by 1.2 to 2.2 log10 CFU per skin sample for loosely attached cells. Counts of Salmonella Typhimurium were reduced by 3.6 to >3.8 log10 CFU per skin sample for firmly attached cells and by 3.5 to >3.8 log10 CFU per skin sample for loosely attached cells. Overall, counts of firmly attached bacteria on TSP-treated skins with mucus were higher than counts on skin without mucus. Firmly attached L. monocytogenes was more resistant to TSP than was firmly attached Salmonella Typhimurium or E. tarda. The presence of mucus on skins slightly decreased the antimicrobial effect of TSP Significant (P < 0.05) reduction in the numbers of all three bacteria can be achieved by treatment with 6% TSP for 10 min. PMID- 12117250 TI - Amino acid decarboxylase activity and other chemical characteristics as related to freshness loss in iced cod (Gadus morhua). AB - Biogenic amine levels and other biochemical indicators were measured to study the safety of and the loss of freshness in iced Atlantic cod. Biogenic amine content exhibited high variability during iced storage of Atlantic cod. Ornithine and lysine decarboxylase activity apparently increased at the end of the storage period. Amino acid activity was probably generated by endogenous amino acid decarboxylases of raw fish. No statistical differences were observed in the total volatile base fraction or in the ammonia or monomethylamine contents during iced storage. However, trimethylamine contents showed a significant exponential relationship with time and sensory score. Cod formed inosine as the major metabolite of IMP. The H and G indices showed a linear relationship with time and sensory score and served as good indicators of cod freshness quality. However, the K, Ki, and P indices showed a logarithmic relationship with time and sensory score. IMP, K, Ki, and P served as indicators of freshness lost during the early stages of chilled storage of cod. PMID- 12117251 TI - Real-time polymerase chain reaction detection of bovine DNA in meat and bone meal samples. AB - We describe a real-time polymerase chain reaction (PCR) assay for the detection of bovine DNA extracted from meat and bone meal (MBM) samples. PCR primers were used to amplify a 271-bp region of the mitochondrial ATPase 8-ATPase 6 gene, and a fluorogenic probe (BOV1) labeled with a 5' FAM reporter and a 3' TAMRA quencher was designed to specifically detect bovine PCR product. The specificity of the BOV1 probe for the detection of the bovine PCR product was confirmed by Southern blot hybridization analysis of the probe with PCR products generated from ovine, porcine, and bovine genomic DNA extracted from blood and with PCR products generated from genomic DNA extracted from single-species laboratory scale rendered MBM samples. The specificity of the BOV1 probe was also evaluated in real-time PCR reactions including these genomic targets. Both methods demonstrated that the BOV1 probe was specific for the detection of bovine PCR product. The BOV1 probe had a detection limit of 0.0001% bovine material by Southern blot DNA probe hybridization analysis and a detection limit of 0.001% bovine material in the real-time PCR assay. Application of the real-time PCR assay to six industrial samples that had previously tested positive for the presence of bovine material with a conventional PCR assay yielded positive results with the real-time PCR assay for four samples. PMID- 12117252 TI - Factors affecting lead leaching from microwavable plastic ware made with lead containing pigments. AB - Although food contact polymers do not normally contain lead, it is suspected that lead may be leached from some microwavable plastic ware items made in Thailand with lead-containing pigments. The purpose of this study was to examine relationships with regard to lead leached from microwavable plastic ware. Four factors were studied: pH, heat level, extraction time, and number of repeated extractions. A total of 243 samples of microwavable plastic ware items locally manufactured in Thailand were used. This study used three pH values (3.5, 4.5, and 6.5) and three heat levels (levels 3, 6, and 9 [170, 500, and 850 W, respectively]). Acetic acid was used both as the extracting agent and for adjusting the pH. Samples were collected at each level at 1, 3, and 5 min, and the amount of leached lead was measured with an atomic adsorption spectrometer. The results of this study show that pH, heat, and extraction time affected the amount of lead leaching from microwavable plastic ware. The amount of lead leaching increased with decreasing pH but increased with increasing heat level and extraction time. On the basis of these three factors, the results of this study indicate that the pH of the extractant (r = -0.592, P < 0.01), the heat level of extraction (r = 0.293, P < 0.01), the extraction time (r = 0.226, P < 0.01), and the number of extractions (r = -0.153, P < 0.01) are related to lead leaching from microwavable plastic ware. The relationship between the pH of the extractant, the heat level of extraction, and the extraction time significantly moderated lead leaching from microwavable plastic ware (R2 = 0.511, P < 0.001). For all factors, the amount of lead leaching was lower than the permissible level of 1 mg/liter specified by the Minister of Public Health. In conclusion, a combination of high acid, prolonged heating, and extraction time accelerated the amount of lead leaching from microwavable plastic ware, but the incidence of lead leaching was negligible. PMID- 12117253 TI - Pulsed-field gel electrophoresis characterization of Shiga toxin-producing Escherichia coli O157 from hides of cattle at slaughter. AB - Contamination of the brisket areas of the hides of healthy adult cattle with Shiga toxin-producing Escherichia coli O157 at slaughter in England was studied. In total, 73 cattle consignments comprising 584 animals delivered to one abattoir over 3 days during 1 week in July 2001 were studied: 26 cattle consignments arriving on Monday, 32 consignments arriving on Wednesday, and 15 consignments arriving on Friday. Consignment sizes ranged from 1 to 23 animals, with a mean consignment size of 8. The hide of the first animal to be slaughtered in each consignment was sampled by using a sponge swab moistened with 0.85% saline to rub an unmeasured brisket (ventral) area (ca. 30 by 30 cm). The process of isolating E. coli O157 from the swabs consisted of enrichment, screening with immunoprecipitation assay kits, and immunomagnetic separation. E. coli O157 was found on 24 of 73 (32.9%) cattle hides examined, and 21 of these 24 isolates produced Shiga toxins. The 24 E. coli O157 isolates produced six different pulsed field gel electrophoresis profiles, and 18 (75%) of the isolates were of one prevalent clone. The high prevalence of one E. coli O157 clone on the hides of cattle at slaughter could be due to a high prevalence of that clone on the 18 farms involved (not investigated in the current study), in the postfarm transport or lairage environments, or both. Since the lairage environment, but not the farm of origin or the postfarm transport vehicle, was a factor common to all 18 cattle consignments, it could have played an important role in spreading the prevalent E. coli O157 clone to the cattle hides. Lairage pen floors and the stunning box floor were identified as the most probable sites along the unloading-to-slaughter route at which the brisket areas of cattle hides could become contaminated. PMID- 12117254 TI - A nucleic acid sequence-based amplification method to detect Salmonella enterica serotype enteritidis strain PT4 in liquid whole egg. AB - Nucleic acid sequence-based amplification (NASBA) was applied to the detection of Salmonella enterica cells in liquid whole egg. Samples (25 g) of liquid whole egg inoculated with Salmonella cells were enriched for 16 h in buffered peptone water, and the NASBA procedure was effective in detecting the presence of Salmonella in samples inoculated with 10 to 100 CFU prior to enrichment. PMID- 12117255 TI - Degradation of natural phosphorylated compounds and added polyphosphates in milk by Pseudomonas fluorescens CECT378, Lactococcus lactis CECT539, and Kluyveromyces marxianus CECT10584. AB - The degradation of natural phosphorylated compounds (galactose-1-phosphate, N acetyl-glucosamine-1-phosphate, glycerophosphoethanolamine, and glycerophosphocholine) and added phosphorylated compounds (diphosphate) in milk was investigated by phosphorus 31 nuclear magnetic resonance on the incubation of a sterile milk with Pseudomonas fluorescens CECT381, Lactococcus lactis CECT539, and Kluyveromyces marxianus CECT10584. This preliminary study showed that the degradation of these compounds was dependent on the compound, microorganism, and temperature of incubation. K. marxianus CECT10584 did not show any capability to degrade these compounds, and L. lactis CECT539 was only able to degrade diphosphate at its optimum growth temperature. P. fluorescens CECT381 was the most active strain and possessed more hydrolytic capabilities at 10 degrees C than at its optimum growth temperature. It is suggested that cold-induced enzymes are involved in the ability of P. fluorescens CECT381 to hydrolyze the natural phosphorylated compounds in milk. Consequent potential alterations of dairy products are discussed. PMID- 12117256 TI - A superantigen bioassay to detect staphylococcal enterotoxin A. AB - Current detection methods for enterotoxins of Staphylococcus aureus are labor intensive and limited in sensitivity. Furthermore, these immunochemical protocols fail to adequately detect heat-treated enterotoxins. Staphylococcal enterotoxins cause severe gastrointestinal illness at relatively low concentrations and retain toxigenicity even after heat treatment. Presented here is a novel method to detect staphylococcal enterotoxin A (SEA). This method is a bioassay that exploits SEA's activity as a superantigen in that it induces in cytotoxic T lymphocytes a cytotoxic response against SEA-bound Raji cells. Target cell death is assayed colorimetrically with the CytoTox 96 cell lysis detection kit. In the experiments presented here, this bioassay was also able to detect heat-treated SEA, albeit with a slight compromise in sensitivity. This system detected SEA at picomolar concentrations. Because of the sensitivity of this assay, it is conceivable that it could be incorporated into current detection methods as a confirmatory test. PMID- 12117257 TI - Antifungal activity of sodium acetate and Lactobacillus rhamnosus. AB - The inhibition of molds by sodium acetate in deMan Rogosa Sharpe (MRS) medium, along with the antifungal activity of Lactobacillus rhamnosus VT1, was studied by the slope agar plate method. MRS agar prepared with and without sodium acetate was used as the agar substrate. A total of 42 strains of Aspergillus, Penicillium, Fusarium, Alternaria, Cladosporium, and Rhizopus were used to compare sensitivities to the inhibitory activity of sodium acetate and L. rhamnosus VT1. It was found that sodium acetate in MRS medium affected the growth of 33 of the 42 mold strains tested to various degrees. The highest sensitivity to sodium acetate was shown by strains of Fusarium, followed by strains of Penicillium, Aspergillus, and Rhizopus. L. rhamnosus VT1 also inhibited mold growth. A significant finding was that sodium acetate and L. rhamnosus VT1 in combination exhibited a possible synergistic action. Thirty-nine of the 42 mold strains tested were completely inhibited by the presence of both antifungal agents. This finding confirms that sodium acetate, a basic component of commercial MRS medium, has strong antifungal properties, and this must be taken into consideration when evaluating the antifungal activity of Lactobacillus cultures grown in MRS broth. PMID- 12117258 TI - Changes in the antigenic and immunoglobulin E-binding properties of hen's egg albumin with the combination of heat and gamma irradiation treatment. AB - This study was carried out to evaluate the changes in the allergenic and antigenic properties of hen's egg albumin (ovalbumin [OVA]) with the combination of heat and gamma irradiation treatment. OVA solution samples were treated by (i) heating (sample 1), (ii) irradiation after heating (sample 2), and (iii) heating after irradiation (sample 3). Samples were isothermally heated and irradiated at the absorption dose of 10 kGy. Competitive indirect enzyme-linked immunosorbent assays (ELISAs) were performed with blood serum to test the ability of treated OVA to bind to immunoglobulin E (IgE) and mouse murine monoclonal antibody (IgG). OVA's ability to bind to mouse IgG changed upon heating at 75 degrees C, and its ability to bind to egg-allergic IgE changed upon heating at 80 degrees C. The ELISAs showed that egg-allergic IgE did not recognize OVA very well when heated at > or = 80 degrees C, while mouse IgG retained better activity under these conditions. Egg-allergic IgE binding was low both for OVA samples treated by heating and for samples treated by irradiation followed by heating. These results show that allergies induced by OVA could be effectively reduced by the combination of heat and gamma irradiation treatment. PMID- 12117260 TI - A review of aerobic and psychrotrophic plate count procedures for fresh meat and poultry products. AB - This is a review of reports that employed aerobic plate counts on fresh meat and poultry products since 1985; it lists synopses of 100 applications. A total of 15 different plating media were used, with 48 (48%) being either plate count agar (PCA) or tryptone glucose yeast extract agar. The temperature-time relations ranged from a low temperature of 20 degrees C for 120 h to 37 degrees C for 24 h. Some 29 different temperature-time combinations were used among the total of 109, with 21 (19.3%) being 35 degrees C/48 h, followed by 12 (11.0%) at 32 degrees C/48 h, 11 (10.1%) at 25 degrees C/48 h, and 9 (8.3%) at 25 degrees C/72 h. Fifty four (49.5%) plate count applications employed incubation temperatures of 30 degrees C and below. From the 26 reports that employed psychrotrophic counts, 16 (61.5%) used PCA; 18 different temperature-time combinations were used, with 7 degrees C/10 d employed by only four. Twenty-one (80.8%) employed an incubation temperature at or <10 degrees C, and five employed an incubation temperature >10 degrees C. There is a serious need for some consensus on methodologies for aerobic and psychrotrophic counts on fresh meat and poultry products. PMID- 12117259 TI - Allergenicity of hen's egg ovomucoid gamma irradiated and heated under different pH conditions. AB - This study was conducted to evaluate the effect of a treatment combining gamma radiation and heating on the allergenic properties of hen's egg ovomucoid (OM) under basic pH conditions. OM solutions of 2.0 mg/ml with pHs of 7.0, 9.0, and 10.0 were gamma irradiated at 10 kGy, heated at 100 degrees C for 15 min, or both. Half of the treated pH 10.0 sample solution was restored to pH 7.4 by dialysis. OM solutions were tested by a competitive direct enzyme-linked immunosorbent assay formatted with immunoglobulin E from egg-hypersensitive patients. An equation was obtained for quantifying intact OM from the standard curve, and the detected concentration of intact OM was calculated. The concentration of intact OM decreased with irradiation or heating, and the rate of the decrease was higher for a basic pH condition than for the physiological condition. The combination of irradiation and heating was very effective in reducing the amount of intact OM regardless of the pH condition. After treatment, the restoration of the pH to 7.4 did not affect the concentration of OM. The results of this study indicate that a combination of irradiation and heating might be an effective method for reducing egg hypersensitivity resulting from OM. PMID- 12117261 TI - Prolactin and zinc in dialysis patients. AB - The objectives of the present study were to investigate the frequencies of hyperprolactinemia and hypozincemia in patients undergoing hemodialysis (HD) or continuous ambulatory peritoneal dialysis (CAPD), the associations between blood levels of zinc (Zn2+) and hormones, and dietary zinc intake amount and its relation to zincemia. We studied 28 patients (14 HD and 14 CAPD) who had their blood levels of Zn2+, prolactin (PRL), parathyroid hormone (PTH), and gonadotropins (LH, FSH) evaluated. Thirteen patients had dietary nutrient amounts evaluated from a 3-d nutritional record. Hyperprolactinemia occurred in 29% patients (HD = CAPD), hypozincemia in 62% (20% HD and 42% CAPD), and low dietary Zn2+ intake in 90% of patients. No correlation among blood concentration of Zn2+ and PRL, PTH, LH, and FSH were observed in the two modalities of dialysis or between zincemia and Zn2+ ingestion. We concluded that the occurrence of hyperprolactinemia and hypozincemia were not related to dialysis modality and that zincemia did not reflect the observed low dietary intake of Zn2+. PMID- 12117262 TI - Concentration of selenium in the whole blood and the thyroid tissue of patients with various thyroid diseases. AB - We investigated the possible differences between the concentrations of selenium in the whole blood and thyroid tissue of patients with thyroid disease. The study comprises 41 women with nodular goiter, 19 women and 2 men with thyroid cancer, 18 women with Graves' disease, and 7 women with thyroiditis. The concentration of selenium was determined by the TRXRF method. The lowest mean selenium level was achieved in the whole blood of women with Graves' disease and the highest in the whole blood of healthy people. In the thyroid cancer tissue, we found the lowest concentration of selenium and the highest in the thyroid gland of women with nodular goiter and Graves' disease. The low selenium levels in the thyroid tissue may increase thyroid cancer risk. PMID- 12117263 TI - Smoking habits and cadmium intake in Turkey. AB - Tobacco was estimated to account over 4 million annual deaths in 1998 and deaths attributable to tobacco usage will rise to 8.4 million in 2020 (http://tobacco.who.int/en/advocacy/wntdzoola.html.) In Turkey, 74% of the males and 29% of the females are smoking, and each year, 150,000 deaths are associated with cigaret smoking. There are 4700 chemical compounds in cigaret smoke, including 43 carcinogens. Cadmium (Cd) is only one of these harmful chemicals in the cigaret. The level of cadmium in whole blood is a reliable index of extent of recent metal uptake. In this study, cadmium concentrations in fasting whole-blood samples, from 119 healthy subjects (58 males and 61 females), ranging in age from 17 to 77 yr, who were not occupationally exposed to cadmium were measured by graphite furnace atomic absorption spectrometry, which is the most widely used technique for the measurement of cadmium concentrations in the whole blood. The blood cadmium concentration of nonsmokers, ex-smokers, and smokers were compared. The blood cadmium concentration of female smokers were found to be highest (mean: 2.62 +/- 0.72; median: 0.90 ng/mL Cd) and that of nonsmokers lowest (mean: 0.67 +/- 0.57; median: 0.44 ng/mL Cd). For smokers, an analysis of smoking history exhibited significant correlations between the number of daily cigarets smoked and the blood cadmium concentration (r = 0.54, p = 0.001) and years of smoking and blood cadmium concentration (r = 0.51, p < 0.001). Cadmium intake from cigaret smoking was found to be higher than the intake from air and diet. PMID- 12117264 TI - Effect of low level zinc pretreatment on zinc-mediated toxicity in different lung cell lines. AB - Reduced toxicity of high zinc exposure was observed after pretreatment of various lung cells with nonlethal zinc concentrations. This effect became significant when various parameters of cytotoxicity were assessed (e.g., inhibition of protein synthesis, depletion of reduced glutathione [GSH], increase of oxidized glutathione [GSSG], release of lactate dehydrogenase [LDH]). Similar protective effects by zinc have already been shown by several investigators for a variety of toxicity studies dealing with cadmium, in vitro and in vivo. Zinc-induced toxicity has been linked to glutathione metabolism and cellular GSH contents. Activity of glutathione reductase (GR) and rates of glutathione synthesis were identified as determinants of zinc (cyto)toxicity. However, these variables were virtually unaffected in our adapted cells. Consequently, another variable appears to be crucial for modulating cellular suscepticibility in zinc pretreated cells. Protection in our cells was achieved by pretreatment with 80-120 micromol/L zinc chloride for 24-72 h, roughly 10-fold more zinc in the medium than is normally found in human plasma. Protection was not observed when the cells were concomitantly exposed to cycloheximide, an inhibitor of protein synthesis, or actinomycin D, an inhibitor of RNA synthesis, but it was found in the presence of amanitin, an inhibitor of mRNA synthesis. It is therefore concluded that the altered zinc tolerance of pretreated cells is not attributable to the induction of metallothionein. PMID- 12117265 TI - Influence of age on lead-induced oxidative stress in rat. AB - Influence of age on lead-induced oxidative stress was investigated in young, adult, and old rats maintained on 0.2% lead acetate (2000 ppm lead) in drinking water for 3 mo. The lead-induced depletion of blood and liver reduced glutathione was about equal in young and adult but not in old rats. The increases in blood, liver, and brain oxidized glutathione and blood and liver superoxide dismutase levels were related to the accumulation of lead in these tissues and followed the order young > adult > old. The lead-induced inhibition of blood delta aminolevulinic acid dehydratase activity, lowering in hemoglobin, and enhanced urinary excretion of delta-aminolevulinic acid were independent of variation in age. The results indicate that young rats may be most sensitive, whereas old rats may be most resistant to some of the oxidative effects of lead examined, which may be related to the accumulation of lead. PMID- 12117266 TI - Influence of antioxidants on metallothionein-mediated protection in cadmium-fed rats. AB - Reduced glutathione (GSH) and alpha-tocopherol offer significant protection against cadmium toxicity in rats. They mobilize cadmium from soft tissues like the liver and kidney, but not from the testis. They diminish oxidative stress by raising GSH and reducing lipid peroxidation. However, no further induction of metallothionein (MT) was observed on their treatment to cadmium-fed rats, with the exception of the kidney in the cadmium- and GSH-treated group. Involvement of regulatory domains (viz. metal responsive elements and metal transcription factor) has also been discussed. It is concluded that MT and antioxidants employ different mechanisms of protection. Genes maintaining the intracellular level of GSH seem to control the entire process of protection by these antioxidants. PMID- 12117267 TI - Effects of dietary iron overload on glutathione peroxidase knockout mice. AB - Excess iron (Fe) intake has been associated with an increased risk of cardiovascular disease in humans, presumably the result of increased oxidative stress. Previous work by us has shown that feeding a high-Fe diet to selenium (Se)-deficient weanling mice for 4 wk resulted in elevated plasma cholesterol and triglycerides and increased hepatic thiobarbituric acid reactive substances (TBARS). Here, we report the effect of Fe overload in mice lacking cellular glutathione peroxidase (GPX1 knockout [KO] mice), the selenoenzyme thought to account for much of the antioxidant action of Se. Four groups of 9-13 weanling wild-type (WT) or GPX1 KO mice were randomly assigned, then fed either an Fe adequate (35 ppm Fe) or high-Fe (1100 ppm Fe) casein-based diet for 4 wk. Iron was added as ferric citrate. Both diets also contained 0.2 ppm Se added as sodium selenite. As expected, liver GPX1 activity was essentially absent in the KO mice. Another Se parameter measured (hepatic thioredoxin reductase activity) did not vary across groups. Although liver Fe was elevated in mice fed the high-Fe diet, liver TBARS was largely unaffected either by mouse genotype or diet fed. Moreover, plasma lipids were not elevated in the Fe-over-loaded GPX1 KO mice. Thus, decreased GPX1 activity cannot account for the pro-oxidant hyperlipidemic effects observed earlier in mice fed the high-Fe Se-deficient diet. This suggests that impairment of Se functions other than GPX1 activity may be responsible for the elevated plasma lipids and hepatic TBARS seen in the Fe-overloaded Se deficient mice. PMID- 12117268 TI - Short communication serum copper, zinc, and calcium concentrations in lice infested sheep. AB - The purpose of this study was to investigate changes in serum concentration of copper, zinc, and calcium in sheep naturally infested with lice (Bovicola caprae, Linognathus africanus, Linognatus ovillus, and Linognattus pedalis). Twenty sheep naturally infested with lice and 20 healthy sheep were used as subjects. Blood samples were collected from the sheep before and 8 and 15 d after treatment with Avermectin, a veterinary antiparasitic drug. The samples were analyzed for their serum copper, zinc, and calcium concentrations by atomic absorption spectrometry. The concentrations of these elements in the infested animals were lower than in the healthy controls, mainly because the general condition of the affected sheep was poor. When the infested animals were treated with an ectoparasitic drug, the serum levels of the studied elements rose to normal ranges while the health of the animals improved. PMID- 12117269 TI - Trace element changes in sclerotic heart valves from patients undergoing aortic valve surgery. AB - Several trace elements are essential nutrients for an optimal functioning of organs and tissues, including the immune system and the heart. The pathogenesis of some heart diseases has been associated with changes in the balance of certain trace elements. The etiology of nonrheumatic aortic valve sclerosis is unknown, however. A prospective study was performed on trace element changes in the sclerotic valves of 46 patients undergoing surgical aortic valve replacement because of aortic stenosis. Valves from 15 individual forensic cases without known cardiac disease served as controls. The contents of 15 trace elements (Al, As, Cd, Ca, Co, Cu, Fe, Pb, Mg, Mn, Hg, Se, Ag, V, and Zn) were measured by inductively coupled plasma - mass spectrometry (ICP-MS) of aortic valve tissue from both patients and forensic autopsy controls. Some trace elements showed similar concentrations in sclerotic and control valves (Al, Ag, Hg, Mn), whereas a few were moderately changed in the sclerotic as compared with the control valves, including an increase in Cd by 52% (p < 0.05) and decreases in Se by 14% (p < 0.05), in V by 42% (p < 0,001), and in Cu by 45% (p < 0.001). However, there were pronounced increases (p < 0.001) in the concentrations of As (5-fold), Ca (70-fold), Co(10-fold), Fe (20-fold), Pb (8-fold), Mg (20-fold), and Zn (10-fold) in the sclerotic valves. Thus, sclerotic aortic valve disease is associated with a pronounced imbalance in several trace elements of well-known importance for cardiovascular and immune function as well as in trace elements with hitherto unknown significance. PMID- 12117270 TI - A unique early gastric tubular adenocarcinoma arising from a pre-existent carcinoid tumor in a patient with a more than 20-year history of type A gastritis: an immunohistochemical and ultrastructural study. AB - A unique early gastric tubular adenocarcinoma developed from a pre-existent carcinoid tumor in a patient with a more than 20-year history of type A gastritis, multiple endocrine cell micronests, hypergastrinemia, and a high level of serum antiparietal cell autoantibody. The patient was a 60-year-old Japanese man. The background gastric mucosa around the tumor showed marked atrophy with intestinal metaplasia, in which endocrine cell micronests were frequently observed, and was consistent with type A gastritis. The mass was composed of both adenocarcinoma and carcinoid tumor. The adenocarcinoma was restricted to the lamina mucosa and submucosal area, and constituted a minor component of the tumor mass. The carcinoid tumor was the dominant constituent of the tumor, that invaded continuously the subserosa and muscularis propria. Based on this examination together with the detailed immunohistochemical and ultrastructural studies, the adenocarcinoma was presumed to have developed from the pre-existent carcinoid tumor. Ultrastructurally there were no amphicrine cells in the tumor, containing both endocrine granules and mucin droplets. PMID- 12117271 TI - An ex vivo model to study the monocyte-endothelial cell interaction in the prelesional stage of experimentally-induced atherogenesis in hamster. AB - We imagined an ex vivo atherogenic model consisting in hypercholesterolemic versus normocholesterolemic aortic arch rings or sigmoid valves incubated in cell culture conditions with human monocytes. Normal tissues did not show any attached monocytes. On the atherogenic aortic arch three different aspects were observed: a) there were no monocytes attached on a normal zone; b) many monocytes adhered to the endothelium on a thickened area, with intimal smooth muscle cells infiltration, and c) the greatest number of attached monocytes was seen in mechanically injured zones of the aortic arch, where the subendothelial area was totally exposed. Immunohisto- and immunocytochemistry for LFA-1 revealed the presence of this leukocyte integrin on monocyte plasma membrane. The labelled monocyte had an activated shape, with pseudopodes extended over the endothelial cells and the anti-LFA-1 antibody coupled with colloidal gold decorating areas apposing to a morphologically modified endothelium. In conclusion, the ex vivo model reproduced the in vivo situations where the monocytes adhere to the modified endothelium covering the thickened areas of hypercholesterolemic aortic wall; they express at least one of the adhesion integrins, namely LFA-1. This study intended to contribute, at least in part, to the understanding of some mechanisms governing the monocyte-endothelial cell interactions in hypercholesterolemia. PMID- 12117272 TI - Comparative ultracytochemical study of the acrosome in four different Veneroida species from the Turkish coast. AB - Cytoplasmic acidic phosphoproteins and complex polysaccharides were stained with ammoniacal silver nitrate-formalin and phosphotungstic acid-chromic acid, respectively. In Cerastoderma glaucum (Cardiacea), acrosomal vesicle contents are differentiated into an apical intermediate-dense component and a basal dense region. PTA stained the apical component and silver stained the basal region and the apex of the acrosome. In Spisula subtruncata (Mactracea) the acrosome showed a PTA-stained apical component and a silver-positive basal dense region. In the Veneracea, Chamelea gallina and Pitar rudis show a tripartite acrosomal vesicle, with apical light, outer dense and inner intermediate-dense regions. In both species, the apical and inner components were stained by PTA, whereas silver stained all regions of the acrosomal vesicle in C. gallina and the apical and outer regions in P. rudis. In midpiece, only C. glaucum showed a positive silver reaction at the centriolar fossa. PMID- 12117273 TI - Correlative microscopy of cerebellar Bergmann glial cells. AB - Double fluorescent labelling of rat cerebellar cortex using antibody to glial fibrillary acidic protein (GFAP) and Alexa fluor conjugates for secondary detection for confocal laser scanning microscope (CLSM), field emission scanning electron microscopy (FESEM) of Rhesus monkey cerebellar cortex, ultrathin sectioning and freeze-etching replica method for transmission electron microscopy of mouse cerebellar cortex have been examined in an attempt to obtain a new and more accurate view of three-dimensional image of Bergmann glial cells (BGC) and their topographic relations in the molecular layer. Intense immunopositive GFAP green staining was observed in the BGC and glial limiting layer. Secondary antibody conjugated with Alexa fluor 488 and Alexa fluor 668-1B4 stained in red capillary endothelial cells and microglial cells. BGC morphology revealed the existence of several cell types or subpopulations of BGC. Bergmann glial fibers, in palisade arrangement, branch and rebranch forming a complex glial network in the molecular layer. Field emission SEM and freeze-fracture SEM method show the SE-I image of high mass dense Bergmann glial cytoplasm ensheathing like a veil the Purkinje cell (PC) soma and dendritric arborization. Bergmann glial fibers appeared completely surrounding individual parallel fibers or parallel fiber bundles, terminal climbing fiber collaterals, basket and stellate cells and capillaries. Freeze-etching direct replicas showed the typical orthogonal arrangement of intramembrane particles, corresponding to the large repertoire of BGC receptors. The study reveals three-dimensional Bergmann glial cells heterogeneity and the complex network formed by Bergmann glial cells in the molecular layer. PMID- 12117274 TI - Eggs of the stink bug Acrosternum (Chinavia) marginatum (Hemiptera: Pentatomidae): a scanning electron microscopy study. AB - The morphology of the outer face of the eggshell, its appearance in cross fractures and the surface morphology of the inner face are described in the green stink bug Acrosternum (Chinavia) marginatum (Hemiptera: Pentatomidae) using scanning electron microscopy. Deposited eggs are barrel-shaped and the surface consists of hexagonally arranged, funnel-shaped pits that possess transverse diaphragms in their slender, basal portions. Only minor differences are detectable between the central portion of the anterior plate (the operculum) and the lateral wall together with the basal portion of the eggshell. The rim at the anterior pole of the egg carries processes with a bulbous end, the aero micropylar processes. A narrow band of the anterior plate bordering these processes shows hexagonally arranged elements. These differ from those of the operculum and the lateral wall, in that cylindrical pits are abundant. During hatching, the operculum is lifted precisely at the rim of this area. Inspection of the periphery of the inner face of the operculum reveals slender, radially arranged grooves. Their arrangement and spacing indicate that they bear a spatial relationship with the aero-micropylar processes. Their role may lie in guiding the sperm to the site where fertilization occurs and in facilitating the gas exchange of the embryo. The remainder of the eggshell carries shallow elevations at its inner face. Exterior and interior features of the eggshell, as well as its appearance in cross-fractures, are discussed. PMID- 12117276 TI - The ultrastructure of the pre-meiotic and meiotic stages of spermatogenesis in Plagioscion squamosissimus (Teleostei, Perciformes, Sciaenidae). AB - Spermatogenesis of 'corvina' P. squamosissimus starts from a stem cell that gives rise to germ cells. These cells are enveloped by Sertoli cells, forming cysts. The germ cells in the cysts are all at the same stage of development and are interconnected by cytoplasmic bridges. Spermatogonia are the largest germ cells. In the cysts, these cells differentiate into primary spermatogonia and secondary spermatogonia. The primary spermatogonia are isolated in the cyst and give rise to the secondary spermatogonia. After several mitotic divisions, they produce spermatocytes I, which can be identified by synaptonemal complexes in the nucleus. The spermatocytes I enter the first phase of meiosis to produce the spermatocytes II. These are not very frequently seen because they rapidly undergo a second phase of meiosis to produce spermatids. PMID- 12117275 TI - Nuclear localization of non-specific acid phosphatase(s) activity in rat vaginal epithelium. AB - The nuclear localization of non-specific acid phosphatase(s) in rat vaginal epithelial cells (VEC) by histochemical and biochemical techniques has been reported in the present work. By different histochemical methods, the enzyme activity was predominantly localized in certain nuclei but not in all of them, however, within the nucleus nucleoli are free from the enzyme activity. This enzyme is diffused in the nucleoplasm and perhaps tightly associated with the nuclear proteins. Very little activity was seen in the cytoplasm in the selective population of VEC. After 12 h treatment of estradiol 17beta in vivo the VEC lose the enzyme activity in the basal and intermediate layers, and the activity remains within the nuclei of luminal cells. Enzyme assay in low and high salt extracts of the isolated nuclei of the VEC with or without the enzyme inhibitors indicates that this enzyme may be present in isoforms in the VEC. No activity was observed in these cells at alkaline pH. PMID- 12117277 TI - Comparative analysis of senescent exorbital lacrimal glands in male and female albino rats. AB - In our previous study we described a bilateral-macroscopic and structural dimorphism of young rat exorbital lacrimal gland (Loewenthal's gland), which was the probable cause of the bibliographic discrepancies in the entity and the onset of its sexual dimorphism. Relevant literature also reported sex-dependent alterations in gland structure during senescence. The present study aims to carry out a comparative analysis on age-dependent changes in glands of both sides from male and female rats, using histological, histochemical and transmission electron microscopy, to evaluate whether the gland bilateral-macroscopic and structural dimorphism might influence the kind of alterations which occur in senescence. Our findings indicate that the macroscopic and structural side-specific dimorphism is not so evident in comparison with young rats. The side-specific dimorphism is evident only in male rats, in which the roundish gland appears to be more Sudan positive in comparison with the ellipsoidal gland. The gland bilateral macroscopic and structural dimorphism, although more evident in comparison with young animals, does not seem to influence these kinds of alterations due to senescence, a time-window in which we still observed some sexual differences also in more aged rats. PMID- 12117279 TI - Periosteal donor site regeneration in rats. AB - Periosteal regeneration was investigated in two periosteal donor sites of the femur. The periosteum was taken from the femur epiphyses and diaphyses of 32 rats. The animals were sacrificed 1, 2, 3, 4 and 8 weeks after periosteal stripping. Intense cell proliferation occurred in the first week. After two weeks, a thick tissue layer formed by osteoblasts and undifferentiated cells was seen at the two donor sites. Eight weeks after, the periosteum had the same aspect as that from the right femur, which was used as control. Histomorphometric analysis showed that periosteal regeneration was significantly different between epiphyses and diaphyses. Periosteal regeneration at donor site located in epiphyses presented greater proliferation and better osteogenic activity than that observed in diaphyses. PMID- 12117278 TI - Intranuclear virus-like particles of a Drosophila hybrid. AB - Intranuclear virus-like particles (VLPs) have been observed in different cell lines and adult tissues of Drosophila. In the present study, intranuclear VLPs have been found in larval tissues (salivary glands, midgut, fat body) as well as in adult tissues (midgut, genitals, fat body) of a rare interspecific hybrid (D. mauritiana x D. melanogaster) called 'mame'. The intranuclear VLPs were round or slightly elliptical with a diameter of 30 nm, and they were found mainly in highly organised clusters, forming large crystalline arrays, near the nucleolus and the polycene chromosomes. These particles were never observed in the cytoplasm of any mame's tissue. A few VLPs were also seen in the corresponding tissues of D. melanogaster, but they were never observed in any tissue of D. mauritiana. There is the intriguing possibility that these VLPs are related to transposable elements and probably contribute to the speciation process, in an unknown, so far, manner. PMID- 12117280 TI - Virus-like particles in an antarctic Aggregata sp.: I. Sporogonial stages. AB - An unknown Aggregata sp. was found in the renal organ of an antarctic Benthoctopus sp., when it was inspected for the presence of dicyemid mesozoans. Merozoites invaded the renal epithelium, whereas sporogonial stages resided in the submucosal connective tissue. Interestingly, individuals of all developmental stages were found to be infected with hitherto unknown virus-like particles. These virus-like particles, spherical in shape, nonenveloped and measuring approximately 30 nm in diameter, could be observed in the nuclei as well as within the cytoplasm. Greater amounts of virus-like particles tended to be arranged in paracrystalline arrays. Although the infection was extensive, it obviously had no pathogenic effect on the parasites themselves. On the basis of the host specificity, size, morphology and histochemical analysis, which suggested the putative viral genome as RNA, a relationship with Totoviridae is assumed. PMID- 12117281 TI - The ultrastructural aspects of vitellogenesis or oocyte secondary growth in Serrasalmus spilopleura (Teleostei, Characiformes, Serrasalminae). AB - Oocyte secondary growth in S. spiloleura corresponds to the period in which different vesicular structures are formed, including the cortical alveoli and the yolk granules. The oocytes with cortical alveolus formation show vesicular structures with filamentous content in the cortical cytoplasmic region, which are the cortical alveolus precursors. In these oocytes, electron-dense vesicles of heterogenous content are dispersed in the inner cytoplasmic region and their nuclei are irregular, showing many nucleoli of different sizes. The oocytes in vitellogenesis are filled with many vesicles. The cortical alveolus precursors are in the peripheral region, and electron-dense granules are seen near to the nucleus. These fuse and form yolk granules. The oocytes in vitellogenesis show a very irregular nucleus that has nucleoli of different sizes. In the oocytes in final vitellogenesis, the yolk granules are scattered throughout the cytoplasm, displacing the cortical alveoli toward cell periphery. The nucleus is similar to the other stages. PMID- 12117282 TI - Ultrastructural cytochemical observations of the cell coat of two Trichomonas vaginalis isolates with different degree of virulence. AB - The cell coat of two Trichomonas vaginalis isolates with different degree of virulence isolated in Mexico from symptomatic women was studied by cytochemical assays. The use of carbohydrate cell surface markers allowed us to visualize greater electron-dense deposits in the highly virulent T. vaginalis isolate than in the less virulent one. On the contrary, parasites treated with concanavalin A showed a heavy uniform electron-dense deposit on the cell surface that was similar in both isolates. When parasites were treated with cationized ferritin the amount of bounded particles on the cell surface was greater in the higher virulent isolate. PMID- 12117283 TI - Immunocytochemical localization of antigens recognised by tropical pulmonary eosinophilia and individuals with intestinal helminths antisera in microfilaria of Wuchereria bancrofti. AB - Ultrathin sections of microfilaria of W. bancrofti embedded in the hydrophilic resin L.R. White were incubated with sera from patients with a typical picture of filarial tropical pulmonary eosinophilia (TPE) and sera from patients of a non endemic region for filariasis regarding intestinal helminths. Both groups had a similar pattern of labelling, except that the labelling intensity was higher with the sera of patients with filarial TPE. The present study indicates relevant epitopes recognised by sera from TPE-patients and also individuals with intestinal helminths in all tissues of microfilaria of W. bancrofti, instead of being localised in a specific nematode region. These findings suggest that sera from people from an area not endemic for filaria, harbouring intestinal helminths, also share antifilarial antibodies that recognise antigens of microfilaria of W. bancrofti. PMID- 12117284 TI - Cytochemical localisation of lysosomal enzymes and acidic mucopolysaccharides in the salivary glands of Aplysia depilans (Opisthobranchia). AB - Three types of secretory cells were reported in the salivary glands of Aplysia depilans: granular cells, vacuolated cells and mucocytes. To improve the characterisation of these cells, cytochemical methods for the detection of lysosomal enzymes and acidic mucopolysaccharides were applied. In granular cells, acid phosphatase and arylsulphatase were present in small lysosomes and in some secretory granules. The secretory granules could have received these enzymes after fusion with the small lysosomes that were frequently found very close to them. These cells were not stained with colloidal iron because they do not contain acidic mucopolysaccharides. In vacuolated cells, acid phosphatase and arylsulphatase were detected in lysosomes but not in the secretory vacuoles. Colloidal iron staining revealed the presence of acidic mucopolysaccharides in the vacuoles and in the Golgi apparatus of these cells. In mucocytes, lysosomes were very rare, but the secretion of these cells was very rich in acidic mucopolysaccharides. The filamentous network within the secretory vesicles was completely covered with iron particles, but practically no particles were observed over the granular masses attached to the membrane of the vesicles. Iron particles were also found in the trans-face cisternae of the U-shaped Golgi stacks, but were not seen in the cis-face cisternae or in the rough endoplasmic reticulum. PMID- 12117285 TI - Distribution of COX-negative mitochondria in myofibers of rats intoxicated with Senna occidentalis seeds. AB - We have described that administration of seeds or parts of the seed of Senna occidentalis (coffee senna) for long periods, induces histochemical changes in the skeletal muscles of hens and rats that are characteristic of a mitochondrial myopathy--as decrease of SDH and COX activity, with some COX negative fibers. In this experimental model of mitochondrial myopathy, as in many human mitochondrial diseases, there is a random distribution of COX negative fibers. Some fibers are completely COX negative while others are partially negative and others are completely positive. In the present work we have studied the distribution of COX negative mitochondria at transmission electron microscopy in skeletal muscle of rats in this experimental myopathy. In myofibers of intoxicated animals the expression of COX was heterogeneous. The histochemical reaction was observed in the internal membrane (more evident in mitochondrial cristae) of all mitochondria of some myofibers, while it was almost absent in other myofibers. In these myofibers the great part of the mitochondria were negative for COX reaction while other ones had a weak expression of this enzyme (dot or focal expression of COX). Our results indicated that the COX mitochondrial activity is heterogeneously impaired in myofibers of rats intoxicated with S. occidentalis. These abnormalities remember those observed in some types of human mitochondrial myopathies. PMID- 12117286 TI - Arterial hypotension in patients on peritoneal dialysis. PMID- 12117287 TI - Therapeutic use of the dialysis prescription for improvement in blood pressure control. PMID- 12117288 TI - The current status of therapeutic apheresis devices in the United States. AB - Over the last 40 years, plasmapheresis technology and its indications for use have been continually evolving. With the growing incidence for autoimmune diseases, unsatisfactory therapeutic options, side effects of drug therapy, and economic relevance, apheresis clinicians have been leaning toward more selective plasmapheresis techniques through the use of plasma fractionators and immunoadsorption columns. Plasma fractionators are mostly used in Asia, and rarely utilized in the U.S. The majority of plasma filters approved by the Food and Drug Administration (FDA) are primary membrane plasma separators, which still require replacement fluid. The secondary plasma fractionators available in the U.S. are limited in used and mostly investigational. Immunoadsorption columns, mostly used in Europe, are gaining popularity in the U.S. Some FDA-approved immunoadsorption columns include the Prosorba protein-A silica column, Immunosorba tryptophan and phenylalanine columns, Immunosorba protein-A sepharose column, and Liposorber dextran sulfate column. In addition, the heparin-induce extracorporeal lipoprotein precipitation (HELP) system is also FDA-approved. However, there are other immunoadsorption technologies used in Europe, such as direct adsorption lipoprotein LDL-hemoperfusion, not yet available in the U.S. While each method of apheresis carries its own risks and benefits, it is the opinion of the authors that these additional apheresis techniques be available to U.S. researchers and clinicians as a therapeutic option. The authors believe that the U.S. need to aggressively investigate more specific plasmapheresis modalities and establish appropriate dialogue with regulatory and reimbursement officials for FDA-approval of these devices. PMID- 12117289 TI - Recent advances in the management of hepatitis C in the dialysis population. PMID- 12117291 TI - The effect of heparin rinse on the biocompatibility of continuous veno-venous hemodiafiltration. AB - The aims of our cross-over randomized study were (1) to assess hemostasis in patients with acute renal failure (ARF) and (2) to determine whether or not the generally recommended heparin rinse of the extracorporeal circuit (ECC) prior to the procedure affects thrombogenicity, complement activation, and leukocyte count in blood during continuous venovenous hemodiafiltration (CVVHDF). Eleven critically ill ARF patients were treated, in random order, using CVVHDF in postdilution setup following ECC rinse with saline (A) with heparin at a concentration of 2,000 IU/L (10 procedures), (B) with heparin at a concentration of 10,000 IU/L (7 procedures), and (C) without heparin (9 procedures). Except for the rinse, anticoagulation therapy did not differ in individual patients during the procedures. Blood was withdrawn before, and at minutes 15, 60, and 360 invariably at diafilter inlet and outlet. Compared with healthy individuals, patients showed lower blood thrombocyte counts (153 vs 233*10(9)/L, p<0.01, arithmetic means, Student's t test), longer aPTT (44 vs 36 s, p<0.05), higher plasma levels of heparin (0.1 vs 0.0 U/mL, p<0.05), D-dimer (1129 vs 36 ng/mL, p<0.001) and beta-thromboglobulin (BTG) (159 vs 37 U/mL, p<0.001) prior to CVVHDF. The comparison of procedures with different rinsing technique did not reveal any significant difference in their effects on blood thrombocyte and leukocyte counts, aPTT, plasma levels of heparin, BTG, thrombin-antithrombin III complexes, D-dimer, or the C5a complement component. CONCLUSIONS: (1) Patients indicated for CVVHDF show impaired hemostasis involving thrombocytes, coagulation, and fibrinolysis, (2) no beneficial effect of heparin rinse on CVVHDF ECC thrombogenicity, complement activation or blood leukocyte counts was demonstrated. PMID- 12117290 TI - Continuous veno-venous hemodiafiltration or hemofiltration: impact on calcium, phosphate and magnesium concentrations. AB - BACKGROUND AND OBJECTIVES: Different techniques of continuous renal replacement therapy (CRRT) might have different effects on calcium, phosphate and magnesium concentrations. Accordingly, we tested whether continuous veno-venous hemodia filtration (CVVHDF) or continuous venovenous hemofiltration (CVVH) would achieve better control of these electrolytes. DESIGN: Retrospective controlled study SETTING: Two tertiary Intensive Care Units PATIENTS: Critically ill patients with acute renal failure (ARF) treated with CVVHDF (n=49) or CVVH (n=50) INTERVENTIONS: Retrieval of daily morning ionized calcium, phosphate and magnesium before and after the initiation of CRRT for up to 2 weeks of treatment. MEASUREMENTS AND RESULTS: Before treatment, both groups had a high incidence of abnormal ionized calcium concentrations (57.2% for CVVHDF vs 46.0% for CVVH; NS). After treatment, both groups showed a significant increase in serum calcium concentration over the first 48 h (p=0.041 vs p=0.0048) but hypercalcemia was more common during CVVHDF (15.3% vs 0.4%; p<0.0001). However, in both groups, hypocalcemia remained common (30.9% vs 36.7%; NS). Before treatment, abnormal serum phosphate concentrations were also common (65.1% for CVVHDF vs 78.1% for CVVH; NS). After treatment, both groups achieved a significant reduction of serum phosphate within 48 hours (p<0.0001 in both groups). There was no difference in the prevalence of abnormal phosphate levels during treatment (45.5% vs 42.4%; NS). Before treatment, both groups had a high incidence of abnormal magnesium concentrations (50.0% for CVVHDF vs 51.2% for CVVH; NS). During treatment, there was no significant change in serum magnesium concentrations during the first 48 hours or in the prevalence of abnormal magnesium concentrations (56.3% vs 63.4%; p=0.13). However CVVHDF was associated with a higher prevalence of hypomagnesemia (8.1% vs 0.4%; p<0.0001) and a lower incidence of hypermagnesemia (48.2% vs. 63.0%; p=0.0014). CONCLUSIONS: In critically ill patients with ARF, calcium, phosphate and magnesium were commonly abnormal and they were only partly corrected by CRRT. CVVH and CVVHDF had a different effect on serum magnesium concentrations. PMID- 12117292 TI - Cell activation and cellular-cellular interactions during hemodialysis: effect of dialyzer membrane. AB - During hemodialysis (HD), circulating blood cells can be activated and also engage in dynamic interplay. These phenomena may be important factors behind dialysis membrane bio(in)compatibility. In the present prospective cross-over study, we have used flow cytometry to evaluate the influence of different dialysis membranes on the activation of circulating blood cells (leukocytes, platelets) and their dynamic interactions (formation of circulating platelet leukocyte and platelet-erythrocyte aggregates) during in vivo HD. Each patient (n = 10) was treated with dialyzers containing membranes of cellulose diacetate, polysulfone and ethylenevinylalcohol (EVAL) in a randomized order. Upregulation of adhesion receptor expression (CD15s, CD11b/CD18) occurred mainly with the cellulosic membrane, though an increase in CD11b/CD18 circulating on neutrophils was also found with both synthetic membranes. Circulating activated platelets (P selectin/CD63-positive platelets) increased during HD sessions with cellulose diacetate and polysulfone. An increased formation of platelet-neutrophil aggregates was found at 15 and 30 min during dialysis with cellulose diacetate and polysulfone but not with EVAL. Platelet-erythrocyte aggregates also increased with cellulose diacetate and at 15 min with polysulfone as well. Generally in concomitance with the increase in platelet-neutrophil coaggregates, there was an increased hydrogen peroxide production by neutrophils. The results of this study indicate that cellular mechanisms can be activated during HD largely depending on the membrane material, EVAL causing less reactivity than the other two membranes. It appears that each dialysis membrane has multiple and different characteristics that may contribute to interactions with blood components. Our results also indicate that derivatizing cellulose (cellulose diacetate) may be a useful way to improve the biocompatibility of the cellulose polymer and that there may be great variability in the biocompatibility profile of synthetic membranes, dialysis with polysulfone being in general associated with a higher degree of cell activation than EVAL membrane. PMID- 12117293 TI - Digoxin poisoning and anuric acute renal failure: efficiency of the treatment associating digoxin-specific antibodies (Fab) and plasma exchanges. AB - Digoxin-specific antibodies (Fab) are currently the treatment of choice for digoxin intoxication. These fragments bind to digoxin, leading to Fab-digoxin complexes, and promote the release of receptor-bound digoxin. These complexes are renally excreted. In the case of anuria, they could be dissociated and lead to renewed intoxication. In this case plasma exchanges are proposed. We report the case of an anuric patient with digoxin intoxication, treated with a Fab injection, followed by a plasma exchange 16 hours later, a second Fab injection was given followed by two plasma exchanges, 38 and 86 hours later. The disappearance of cardiac abnormalities showed the efficiency of the Fab, the drop in serum digoxin concentration and the high digoxin concentration in the exchanged plasma indicate effective elimination. The association of Fab and plasma exchanges could be proposed in the case of digoxin intoxication in the anuric patient. PMID- 12117294 TI - A method to assess biochemical activity of liver cells during clinical application of extracorporeal hybrid liver support. AB - Biochemical activity of a hybrid liver support system based on porcine liver cells was investigated in patients suffering from acute liver failure, coma stage III-IV Patient plasma was drawn systemically and after circulation through the bioreactor at four hour intervals. A method is used that takes into account the rate of plasma flow and the differences in plasma concentration systemically and after circulation through the liver support system to determine the net release or uptake of metabolites such as ammonia, urea and glucose. Urea release (mean 2.28+/-0.37 micromol/h/g cells) and ammonia uptake (mean 0.17+/-0.11 micromol/h/g cells) was seen during treatment, an active role of the system in glucose metabolism was observed. All patients were bridged successfully to liver transplantation. PMID- 12117295 TI - In vitro comparison of blood pump induced platelet microaggregates between a centrifugal and roller pump during cardiopulmonary bypass. AB - Platelets are consumed during cardiopulmonary bypass (CPB) and mechanical ventricular assistance, at least partly as a result of the formation of platelet microaggregates in the blood pump. There is no commonly accepted method currently available to detect platelet microaggregates during the use of CPB or left ventricular assist devices (LVAD). The purpose of this study was to develop a flow cytometric method for the quantification of platelet microaggregates generated in blood pumps, and to evaluate the effect of cellular fragments from hemolyzed erythrocytes on the perioperative assessment of platelet counts during CPB. METHOD: Fresh human anticoagulated blood (1IU heparin/mL, activated clotting time 250+/-24 sec.) was circulated for 120 minutes in an artificial circulatory system, containing either a centrifugal pump (CP) or roller pump (RP). Whole blood was used to quantify platelet consumption and to detect circulating platelet microaggregates in a flow cytometer. Platelet consumption was additionally analyzed using an automated "Coulter" blood cell counter. Hemolysis was analyzed by measurement of plasma free hemoglobin (fHb), as well a by flow cytometric detection of red blood cell (RBC) fragments. RESULTS: Flow cytometric analysis demonstrated significantly more circulating platelet aggregates and platelet consumption in the RP than in the CP (p<0.01). Quantification of RBC fragments and plasma free hemoglobin (fHb) levels also indicated significantly increased hemolysis in the RP than in the CP (p<0.01). In contrast, the Coulter count data indicated less platelet consumption in the PP compared to the CP. CONCLUSION: Fragments from hemolyzed erythrocytes have the same size distribution as intact platelets and the number of RBC fragments correlates with the extent of pump-induced hemolysis during CPB. Our data suggest that assessment of platelets by "Coulter counting" cannot distinguish platelets from RBC fragments and may underestimate platelet consumption in the presence of hemolysis during CPB. We conclude that flow cytometry is more accurate in the perioperative assessment of platelet count and platelet aggregation during CPB and LVAD support. PMID- 12117296 TI - Application of the moving-actuator type pump as a ventricular assist device: in vitro and in vivo studies. AB - A moving actuator type pump has been developed as a multifunctional Korean artificial heart (AnyHeart). The pump consists of a moving actuator as an energy converter, right and left sacs, polymer (or mechanical) valves, and a rigid polyurethane housing. The actuator containing a brushless DC motor moves back and forth on an epicyclical gear train to produce a pendular motion, which compresses both sacs alternately. Of its versatile functions of ventricular assist device and total artificial heart use, we have evaluated the system performance as a single or biventricular assist device through in vitro and in vivo experiments. Pump performance and anatomical feasibility were tested using various animals of different sizes. In the case of single ventricular assist device (VAD) use, one of the sacs remained empty and a mini-compliance chamber was attached to either an outflow or inflow port of the unused sac. The in vitro and in vivo studies show acceptable performance and pump behavior. Further extensive study is required to proceed to human application. PMID- 12117297 TI - Carcinogenic effects of polychlorinated biphenyls. AB - As part of its mandate, the Agency for Toxic Substances and Disease Registry (ATSDR) prepares toxicological profiles on hazardous chemicals found at Comprehensive Environmental Response, Compensation, and Liability Act (CERCLA) National Priorities List (NPL) sites that have the greatest public health impact. These profiles comprehensively summarize toxicological and environmental information. This article constitutes the release of an important section of the Toxicological profile for polychlorinated biphenyls [ATSDR. 2000: Toxicological profile for polychlorinated biphenyls. Atlanta, GA: US Department of Health and Human Services, Agency for Toxic Substances and Disease Registry.] into the scientific literature. This article focuses on the carcinogenic effects of this group of synthetic organic chemicals (polychlorinated biphenyls) in humans and animals. Information on other health effects, toxicokinetics, mechanisms of toxicity, biomarkers, interactions, chemical and physical properties, potential for human exposure, and regulations and advisories is detailed in the profile. PMID- 12117298 TI - Effects of polychlorinated biphenyls on development and reproduction. AB - As part of its mandate, the Agency for Toxic Substances and Disease Registry (ATSDR) prepares toxicological profiles on hazardous chemicals found at Comprehensive Environmental Response, Compensation, and Liability Act (CERCLA) National Priorities List (NPL) sites that have the greatest public health impact. These profiles comprehensively summarize toxicological and environmental information. This article, which constitutes the release of an important section of the Toxicological Profile for Polychlorinated Biphenyls (ATSDR 2000) into the scientific literature, focuses on the developmental and reproductive effects of this group of synthetic organic chemicals (PCBs) in humans and animals. Information on other health effects, toxicokinetics, mechanisms of toxicity, biomarkers, interactions, chemical and physical properties, potential for human exposure, and regulations and advisories is detailed in the profile. Interested readers are encouraged to consult the original toxicological profile for more information. Profiles can be requested from ATSDR's Information Center by telephone (1-888-42-ATSDR [1-888-422-8737] or E-mail: (atsdric@cdc.gov). PMID- 12117299 TI - Adenosine participates in regulation of smooth muscle relaxation in aortas from rats with experimental hypothyroidism. AB - The contribution of adenosine receptors was evaluated in vascular relaxation in experimental hypothyroidism. Hypothyroid aortic rings contracted less than normal controls with noradrenaline, phenylephrine, and KCl; the difference was maintained after incubation with 1,3-dipropyl-8-p-sulfophenylxanthine (an A1 and A2 adenosine receptor blocker). The vascular relaxation induced by acetylcholine or carbachol was similar in normal and hypothyroid aortic rings. However, adenosine, N6-cyclopentyladenosine (an A1 adenosine receptor analogue), and 5'-N ethylcarboxamidoadenosine (an A2 and A3 adenosine analogue) induced vasodilation that was larger in hypothyroid than in normal aortas. Nomega-nitro-L-arginine methyl ester shifted the dose-response curves of adenosine, N6 cyclopentyladenosine, or 5'-N-ethylcarboxamidoadenosine to the right in both normal and hypothyroid vessels. The blocker 1,3-dipropyl-8-p-sulfophenylxanthine significantly reduced adenosine-induced relaxation in the hypothyroid but not in the normal aortic vessels. These results suggest that in hypothyroid aortas, a larger adenosine-mediated vasodilation is observed probably due to an increase in receptor number or sensitivity. PMID- 12117300 TI - Differentiation-dependent expression of cathepsin D and importance of lysosomal proteolysis in the degradation of UCP1 in brown adipocytes. AB - The lysosomal protease cathepsin D increased markedly in brown adipocytes during differentiation in primary cultures. Differentiated cells had 20 times the amount of immunoreactive cathepsin D found in preadipocytes. Cathepsin D mRNA, as estimated by relative RT-PCR, was also present in higher amounts in differentiated brown fat cells. Cathepsin D expression was not influenced by repeated exposures of brown adipocytes to norepinephrine (NE). Cathepsin D levels were also unchanged when NE was withdrawn for 48 h after cells had been exposed to NE for 7 days. In contrast, exposure of the cells to NE for 7 days increased their UCP1 content by more than twofold, which returned to basal levels within 48 h of withholding NE. The half-life of UCP1 under basal conditions and in cells chronically exposed to NE was estimated from reductions in [35S]methionine labelled immunoprecipitable UCP1 over 72 h. UCP1 t1/2 under basal conditions was 3.7+/-0.4 days, which was similar to the half-lives of labelled mitochondrial translation products (3.6+/-0.8 days). The turnover rates of both UCP1 and mitochondrial translation products were reduced by NE. The turnover rate of UCP1 in the presence or absence of NE cannot account solely for the rapid loss of UCP1 from brown adipocytes upon withdrawal of NE. This loss was reduced when cells were incubated with inhibitors of phosphatidylinositol 3-kinases (PI 3-kinase), previously shown to block formation of autophagic vacuoles. Thus, brown adipocytes acquire a large capacity for both uncoupled metabolism and for lysosomal proteolysis during differentiation. Withdrawal of NE, as often occurs in vivo from suppression of sympathetic nervous system activity, would not only terminate thermogenesis but also favor formation of autophagic vacuoles to rapidly reduce the cell content of UCP1-containing mitochondria. PMID- 12117301 TI - Hyperhydration induced by glycerol ingestion: hormonal and renal responses. AB - The simultaneous time courses of hydromineral hormones (renin-aldosterone system, arginine vasopressin, and atrial natriuretic peptide) and renal responses were examined during and after the completion of hyperhydration induced by glycerol and fluid ingestion. Eight healthy young male Caucasian subjects participated in two separate trials, each including three consecutive phases in a thermoneutral environment. Phases 1 and 3 involved a 90-min period at rest, while phase 2 involved a 120-min period at rest designed to provide either (i) euhydration (control trial) or (ii) hyperhydration induced by ingestion of glycerol (1.1 g/kg body mass) and fluid (21.4 mL/kg body mass). During the 2-h time period of glycerol and fluid ingestion, urine flow, urine osmolality, and plasma levels of hydromineral hormones remained at basal values. In contrast, after hyperhydration completion during phase 3, the diuresis increased markedly together with a dilution of the urine (p < 0.05) while hormonal responses did not change. These results indicate significant differences in renal responses during and after hyperhydration completion and suggest that these changes are independent of fluid regulating hormonal responses. PMID- 12117302 TI - Characterization of cyclic AMP-regulated chloride conductance in the pigmented rabbit conjunctival epithelial cells. AB - We have previously reported that the pigmented rabbit conjunctiva is a Cl- secreting tissue, subject to cAMP, Ca2+, and PKC modulation. The present study was conducted to characterize, at the cellular and molecular levels, cAMP regulated Cl- channels in rabbit conjunctival epithelial cells. cAMP-inducible Cl channel properties were evaluated by monitoring the whole-cell currents using patch clamp techniques. Results showed that 10 microM forskolin significantly stimulated a glibenclamide-inhibitable whole-cell conductance by approximately five-fold. Furthermore, reduction of the Cl- concentration in the bathing solution through partial substitution of NaCl with Na-isethionate resulted in a rightward shift of the reversal potential for both baseline and forskolin stimulated whole-cell currents from 0 to values close to the theoretical Cl- reversal potential predicted by the Nernst equation. Western blot analysis with a monoclonal antibody recognizing the epitope in the C-terminus of the cystic fibrosis transmembrane conductance regulator (CFTR) showed a positive band at its molecular weight, approximately 170 kD. Immunostaining under confocal microscopy revealed a CFTR specific signal in the apical sections of primary conjunctival epithelial cells. In addition, RT-PCR detection amplified a cDNA fragment 100% identical to the predicted portion of the cloned rabbit CFTR message. The stage is thus set for determining the extent of CFTR contribution to cAMP-regulated Cl- conductance in pigmented rabbit conjunctival epithelial cells. PMID- 12117303 TI - Control of proteolysis by norepinephrine and insulin in brown adipocytes: role of ATP, phosphatidylinositol 3-kinase, and p70 S6K. AB - The objective of this study was to evaluate some of the mechanisms by which norepinephrine (NE) and insulin may influence protein degradation in mouse brown adipocytes differentiated in cultures. The effects of NE and insulin, alone or in combination, on three factors known to influence proteolysis (maintenance of cell ATP and 1-phosphatidylinositol 3-kinase (PI 3-kinase) and p70 ribosomal S6-kinase (p70 S6K) activities) were examined. It was proposed that NE affects proteolysis indirectly by decreasing cell ATP from activation of uncoupling protein-1 (UCP1) dependent mitochondrial respiration. This was tested by comparing the effects of NE and fatty acids (which directly activate UCP1) on proteolysis in brown adipocytes, as well as in pre-adipocytes and 3T3-L1 adipocytes, which do not express UCP1. An inhibitory effect of insulin on proteolysis is observed in both pre-adipocytes and differentiated cells, whereas NE and exogenously added fatty acids inhibit proteolysis only in brown adipocytes. There is a linear relationship between reductions in cell ATP and proteolysis in response to increasing concentrations of NE or fatty acids. PI 3-kinase activity is required for proteolysis, because two selective inhibitors (wortmannin and LY294002) reduce proteolysis in both pre-adipocytes and differentiated cells. This effect is not additive to that of NE, which suggests they affect the same proteolytic pathway. In contrast to NE, insulin increases PI 3-kinase activity and phosphorylation of p70 S6K. Rapamycin, which prevented insulin-dependent increase in phosphorylation of p70 S6K, increases proteolysis in brown adipocytes and antagonizes the inhibitory effect of insulin on proteolysis, but not the inhibitory effect of NE. Thus, insulin inhibits proteolysis via rapamycin sensitive activation of p70 S6K, whereas the effect of NE appears largely to be a function of decreasing cell ATP content. PMID- 12117304 TI - Magnesium supplementation and deoxycorticosterone acetate--salt hypertension: effect on arterial mechanical properties and on activity of endothelin-1. AB - The aim of this study was to show whether the decrease in blood pressure induced by Mg supplementation in deoxycorticosterone acetate - salt (DOCA-salt) hypertensive rats is associated with mechanical modifications of blood vessels and (or) changes in tissular production and (or) vasoconstrictor activity to endothelin-1. DOCA-salt treatment increased blood pressure, media thickness, cross-sectional area, and lumen diameter of carotid arteries. Distensibility and incremental elastic modulus versus stress were not altered in carotid arteries, suggesting that the DOCA-salt vessel wall adapts structurally to preserve its blood pressure buffering capacity. Magnesium supplementation attenuated DOCA-salt hypertension. In comparison with normotensive rats, systolic, mean, and pulse pressures were higher whereas diastolic pressure was not different in Mg supplemented DOCA-salt rats. Magnesium supplementation did not significantly modify the elastic parameters of carotid arteries. In resistance mesenteric arteries, DOCA-salt hypertension induces an inward hypertrophic remodeling. Magnesium supplementation attenuates wall hypertrophy and increases lumen diameter to the normotensive diameter, suggesting a decrease in peripheral resistance. Magnesium supplementation normalizes the altered vasoconstrictor activity of endothelin-1 in mesenteric arteries and attenuates endothelin-1 overproduction in kidney, left ventricle, and aorta of DOCA-salt rats. These findings suggest that Mg supplementation prevents blood pressure elevation by attenuating peripheral resistance and by decreasing hypertrophic effect of endothelin-1 via inhibition of endothelin-1 production. PMID- 12117305 TI - Effect of purinergic agonists and antagonists on insulin secretion from INS-1 cells (insulinoma cell line) and rat pancreatic islets. AB - The effects of purinergic agonists on insulin release are controversial in the literature. In our studies (mainly using INS-1 cells, but also using rat pancreatic islets), ATP had a dual effect on insulin release depending on the ATP concentration: increasing insulin release (EC50 approximately/= 0.0032 microM) and inhibiting insulin release (EC50 approximately/= 0.32 microM) at both 5.6 and 8.3 mM glucose. This is compatible with the view that either two different receptors are involved, or the cells desensitize and (or) the effect of an inhibitory degradation product such as adenosine (ectonucleotidase effect) emerges. The same dual effects of ATP on insulin release were obtained using rat pancreatic islets instead of INS-1 cells. ADPbetaS, which is less degradable than ATP and rather specific for P2Y1 receptors, had a dual effect on insulin release at 8.3 mM glucose: stimulatory (EC50 approximately/= 0.02 microM) and inhibitory (EC50 approximately/= 0.32 microM). The effectiveness of this compound indicates the possible involvement of a P2Y1 receptor. 2-Methylthio-ATP exhibited an insulinotropic effect at very high concentrations (EC50 approximately/= 15 microM at 8.3 mM glucose). This indicated that distinct P2X or the P2Y1 receptor may be involved in these insulin-secreting cells. UTP increased insulin release (EC50 approximately/= 2 microM) very weakly, indicating that a P2U receptor (P2X3 or possibly a P2Y2 or P2Y4) are not likely to be involved. Suramin (50 microM) antagonized the insulinotropic effect of ATP (0.01 microM) and UTP (0.32 microM). Since suramin is not selective, the data indicated that various P2X and P2Y receptors may be involved. PPADS (100 microM), a P2X and P2Y1,4,6 receptor antagonist, was ineffective using either low or high concentrations of ATP and ADPbetaS, which combined with the suramin data hints at a P2Y receptor effect of the compounds. Adenosine inhibited insulin release in a concentration-dependent manner. DPCPX (100 microM), an adenosine (A1) receptor antagonist, inhibited the inhibitory effects of both adenosine and of high concentrations of ATP. Adenosine deaminase (1 U/mL) abolished the inhibitory effect of high ATP concentrations, indicating the involvement of the degradation product adenosine. Repetitive addition of ATP did not desensitize the stimulatory effect of ATP. U-73122 (2 microM), a PLC inhibitor, abolished the ATP effect at low concentrations. The data indicate that ATP at low concentrations is effective via P2Y receptors and the PLC-system and not via P2X receptors; it inhibits insulin release at high concentrations by being metabolized to adenosine. PMID- 12117306 TI - The inotropic effect of nitric oxide on mammalian papillary muscle is dependent on the level of beta1-adrenergic stimulation. AB - We tested the hypothesis that nitric oxide has a positive inotropic effect on mammalian cardiac muscle contractility and that this effect sums with the positive inotropic effect of beta1-adrenergic agonists when both are present. Feline right ventricular papillary muscles were stimulated to contract isometrically at 0.2 Hz in Krebs-Henseleit bicarbonate buffer (KREBS) gassed with 95% O2 and 5% CO2 (26 degrees C; pH 7.34). The nitric oxide (NO) donor, S-nitroso N-acetylpenicillamine (SNAP, 10(-5) M), and the membrane permeable cGMP analog 8 bromoguanosine-3',5'-cyclophosphate sodium (Br-cGMP, 10(-5) M), significantly increased developed force by 13.3+/-1.5% (n = 11) and 7.8+/-2.8% (n = 7), respectively. SNAP, at 10(-5) M, significantly increased the force developed by papillary muscle treated with 10(-11) M or 10(-9) M dobutamine hydrochloride (a beta1-adrenergic agonist) (n = 25, 11.3+/-2.9% and 10.0+/-3.6%, respectively) when compared with the addition of KREBS (n = 27, 2.6+/-0.9% and 5.5+/-0.9%), but the increase was less than predicted by the sum of inotropic effects of SNAP and dobutamine. SNAP at 10(-5) M did not change developed force in muscles treated with 10(-7) M dobutamine but it significantly decreased developed force in muscles challenged with 10(-5) M dobutamine (n = 18, 29.3+/-5.0%) when compared with KREBS (n = 10, 41.5+/-6.8%). Similarly, 10(-4) M 8-bromo-adenosine cyclic 3',5'-hydrogen phosphate monosodium (a membrane permeable cAMP analog) increased developed force 14.9+/-3.3% and the addition of 10(-5) M Br-cGMP to those muscles significantly reduced developed force by 3.5%+/-1.1% (n = 7). Thus, the positive inotropic effect of NO decreased and ultimately became an attenuation as the level of beta1-adrenergic stimulation increased due at least in part, to an interaction between the cAMP and cGMP second messenger pathways. PMID- 12117307 TI - Functional and autoradiographic characterization of dopamine D2-like receptors in the guinea pig heart. AB - Dopamine receptors include the D1- (D1 and D5 subtypes) and D2-like (D2, D3, and D4 subtypes) families. D1-like receptors are positively and D2-like receptors negatively coupled to the adenylyl cyclase. Dopamine D2-like (D4 subtype) receptors have been identified in human and rat hearts. However the presence of D2 and D3 receptor subtypes is unclear. Furthermore, their role in cardiac functions is unknown. By autoradiographic studies of guinea pig hearts, we identified D3 and D4 receptors, using the selective radioligands [3H]-7-OH-DPAT and [3H]emonapride (YM-09151-2 plus raclopride). Western blot analysis confirmed D3 and D4 receptors in the right and left ventricle of the same species. Selective agonists of D3 and D4 receptors (+/-)-7-OH-DPAT and PD 168 077 (10(-9) to 10(-5) M, respectively) induced a significant negative chronotropic and inotropic effect in the isolated guinea pig heart preparation. Negative inotropic effect induced by PD 168 077 was associated with an inhibition in cyclase activity. No changes in cyclase activity were found with (+/-)-7-OH-DPAT. The aim of this study is to support the presence of D3 and D4 receptors in the heart. Although our results suggest that D3 and D4 receptors are functionally active in the heart, we need additional information with an antagonist and an agonist of improved potency and selectivity to understand the respective roles of D3 and D4 receptors in the cardiac functions. PMID- 12117308 TI - Characterization of Ca2+ release from heterogeneous Ca2+ stores in sarcoplasmic reticulum isolated from arterial and gastric smooth muscle. AB - Ca2+ transport was investigated in vesicles of sarcoplasmic reticulum subfractionated from bovine main pulmonary artery and porcine gastric antrum using digitonin binding and zonal density gradient centrifugation. Gradient fractions recovered at 15-33% sucrose were studied as the sarcoplasmic reticulum component using Fluo-3 fluorescence or 45Ca2+ Millipore filtration. Thapsigargin blocked active Ca2+ uptake and induced a slow Ca2+ release from actively loaded vesicles. Unidirectional 45Ca2+ efflux from passively loaded vesicles showed multicompartmental kinetics. The time course of an initial fast component could not be quantitatively measured with the sampling method. The slow release had a half-time of several minutes. Both components were inhibited by 20 microM ruthenium red and 10 mM Mg2+. Caffeine, inositol 1,4,5-trisphosphate, ATP, and diltiazem accelerated the slow component. A Ca2+ release component activated by ryanodine or cyclic adenosine diphosphate ribose was resolved with Fluo-3. Comparison of tissue responses showed that the fast Ca2+ release was significantly smaller and more sensitive to inhibition by Mg2+ and ruthenium red in arterial vesicles. They released more Ca2+ in response to inositol 1,4,5 trisphosphate and were more sensitive to activation by cyclic adenosine diphosphate ribose. Ryanodine and caffeine, in contrast, were more effective in gastric antrum. In each tissue, the fraction of the Ca2+ store released by sequential application of caffeine and inositol 1,4,5-trisphosphate depended on the order applied and was additive. The results indicate that sarcoplasmic reticulum purified from arterial and gastric smooth muscle represents vesicle subpopulations that retain functional Ca2+ channels that reflect tissue-specific pharmacological modulation. The relationship of these differences to physiological responses has not been determined. PMID- 12117309 TI - Phosphate absorption by Aplysia californica gut: thyroid hormone stimulation. AB - Mucosal membranes of foregut epithelia of Aplysia californica contain a sodium/phosphate symporter. Triiodothyronine stimulated the absorptive activity of the sodium/phosphate symporter, whereas reverse triiodothyronine had no effect on the sodium/phosphate symporter. It appears that thyroid hormone or its molluscan equivalent plays a role in the overall regulation of phosphate homeostasis by the Aplysia californica gut. PMID- 12117310 TI - Electrical stimulation for the treatment of bladder dysfunction: current status and future possibilities. AB - Electrical stimulation of peripheral nerves can be used to cause muscle contraction, to activate reflexes, and to modulate some functions of the central nervous system (neuromodulation). If applied to the spinal cord or nerves controlling the lower urinary tract, electrical stimulation can produce bladder or sphincter contraction, produce micturition, and can be applied as a medical treatment in cases of incontinence and urinary retention. This article first reviews the history of electrical stimulation applied for treatment of bladder dysfunction and then focuses on the implantable Finetech-Brindley stimulator to produce bladder emptying, and on external and implantable neuromodulation systems for treatment of incontinence. We conclude by summarizing some recent research efforts including: (a) combined sacral posterior and anterior sacral root stimulator implant (SPARSI), (b) selective stimulation of nerve fibers for selective detrusor activation by sacral ventral root stimulation, (c) microstimulation of the spinal cord, and (d) a newly proposed closed-loop bladder neuroprosthesis to treat incontinence caused by bladder overactivity. PMID- 12117311 TI - An overview of the state of the art of noninvasive FES for independent ambulation by thoracic level paraplegics. AB - This paper is an overview of the status of transcutaneous noninvasive (unbraced) functional electrical stimulation (FES) for independent standing and for independent ambulation by traumatic spinal-cord injured (SCI) paraplegics with complete spinal cord lesions at the thoracic level. The paper discusses aspects of patient selection, patient training, system performance, ambulation range, medical benefits and psychological benefits. It also considers problems relating to system adoption and long term system use. Furthermore, the paper discusses the various aspects of transcutaneous noninvasive FES as compared with implanted FES systems for ambulation by thoracic level SCI patients. PMID- 12117312 TI - Neuroprostheses for grasping. AB - In recent years a number of neuroprostheses have been developed and used to assist stroke and spinal cord injured subjects to restore or improve grasping function. These neuroprostheses clearly demonstrated that the targeted group of subjects can significantly benefit from this technology and that functional electrical stimulation (FES) is a viable method for restoring or improving grasping function. In this article the FES technology is briefly explained and some of the better known neuroprostheses for grasping are discussed. Furthermore, a typical population of subjects that can benefit from this technology is indicated as well as the methodology to select and train these subjects to apply the neuroprosthesis in daily living activities. This article also provides a brief summary of the achieved results with the existing neuroprostheses for grasping and discusses some of the challenges this technology is currently facing. PMID- 12117313 TI - Motor recovery after stroke: lessons from functional brain imaging. AB - Several theories have been proposed to explain recovery from stroke. Functional brain imaging offers an opportunity to evaluate these theories and visualize recovery after stroke. Functional brain imaging has proven to be an effective tool to map brain areas activated during a specific task. This paradigm can extend our understanding of the mechanisms of motor recovery after stroke. Functional brain imaging tools such as functional MRI, PET, transcranial Doppler ultrasonography, and transcranial magnetic stimulation can be used to evaluate motor activation after stroke. Functional imaging is proving useful in identifying areas, pathways and mechanisms involved in motor recovery after stroke. Studies have shown changes in motor organization with rehabilitation. Functional brain imaging may assist in the selection of rehabilitation methods that best foster recovery. PMID- 12117314 TI - Somatosensory evoked magnetic fields in hemimegalencephaly. AB - Somatosensory maps were determined in three patients with hemimegalencephaly using magnetic resonance imaging (MRI) and magnetoencephalography (MEG). MRIs were characterized by thickened gray matter with clearly aberrant lamination patterns. Somatosensory Evoked Fields (SEFs), as measured by MEG, were absent from the affected hemisphere in the two patients with severe cortical lamination defects. The third patient presented with relatively preserved cortical lamination in the frontal lobe and clear cortical SEFs in this region, indicating somatotopical reorganization. These findings suggest that the presence and location of MEG-derived somatosensory maps reflect the severity of the cortical lamination defects in hemimegalencephaly. PMID- 12117315 TI - Callosal anomalies in patients with spinal dysraphism: correlation of clinical and neuroimaging features with hemispheric abnormalities. AB - Dysgenesis of the corpus callosum can occur in association with spinal dysraphic lesions. Clinical and neuroimaging features were reviewed in 23 patients (12 male, 11 female; mean age 11.3 years) with caudal spinal dysraphism (myeloschisis in eight, meningomyelocele in 10, and lumbosacral lipoma in five) to characterize types and degrees of callosal and other cerebral anomalies. T1- and T2-weighted magnetic resonance images were obtained, and the total midsagittal cross sectional area of the corpus callosum was determined. The corpus callosum appeared normal in nine patients and was abnormal in 14. In five patients the corpus callosum was narrow, with all regions present; the cerebral hemispheres were hypoplastic. Two patients with dysgenesis of frontal, parietal, and occipital lobes had a small, partly agenetic corpus callosum. In the remaining seven patients the posterior third of the corpus callosum was absent or hypoplastic; six of them had ventriculomegaly that selectively affected the occipital horns (colpocephaly). All callosal anomalies were accompanied by hemispheric ones. This supports a disordered developmental relationship between the corpus callosum and the hemispheres as a cause. Spinal dysraphism can no longer be considered a single developmental abnormality, given the frequent association of other defects. PMID- 12117316 TI - Trophic effect of olmesartan, a novel AT1R antagonist, on spinal motor neurons in vitro and in vivo. AB - Olmesartan is a novel compound which has been shown to exhibit various neuropharmacological effects. For the purpose of clarifying the effect of Olmesartan on spinal motor neurons, we studied the following tests. We studied the effect in vitro of Olmesartan on neurite outgrowth and choline acetyltransferase (ChAT) activity in primary explant cultures of ventral spinal cord (VSCC) of fetal rats. Olmesartan-treated VSCC, compared with control VSCC, had a significant neurite outgrowth and increased activity of ChAT. The effect was dose-related in neurite outgrowth. However, there was no relationship between activity of ChAT andgiven doses of Olmesartan. We examined in vivo the effect of Olmesartan on axotomized spinal motor neuron death in the rat spinal cord. After post-natal unilateral section of sciatic nerve, there was approximately a 50% survival of motor neurons in the fourth lumbar segment. In comparison with vehicle, intraperitoneal injection of Olmesartan for consecutive 14 days reduced spinal motor neuron death. There was no relationship between number of surviving neurons and doses of Olmesartan. These in vitro and in vivo studies showed that Olmesartan has a neurotrophic effect on spinal motor neurons. Our data suggest a potential therapeutic use of Olmesartan in treating diseases that involve degeneration and death of motor neurons, such as motor neuropathy and amyotrophic lateral sclerosis. PMID- 12117318 TI - The tissue level of dexamethasone in human brain tumors is about 1000 times lower than the cytotoxic concentration in cell culture. AB - In glioblastoma patients dexamethasone is routinely administered as an antiedematous drug. In contrast to its empirically proven effect, the biochemical way of action remains poorly understood. In order to assess whether a direct cytotoxic effect is present in vivo we compared dexamethasone levels in brain tumor specimens with its cytotoxic concentrations in cell culture. Biopsy specimens were taken during microsurgical tumor removal, homogenized and dexamethasone levels were measured by high pressure liquid chromatography. In cell culture we tested different concentrations of dexamethasone on A172, U87, U373 cells and on eleven primary glioblastoma cell lines. Furthermore a pilocytic astrocytoma I, an astrocytoma II and an oligodendroglioma III and a meningioma were examined. Cell viability was assessed using the Alamar Blue assay and the concentrations resulting in loss of 50% of the cell population were calculated (LD50). The average brain tumor tissue concentration of dexamethasone was 225 nanogram g(-1). The mean LD50 in cell culture ranged at 222 microgram ml(-1). We conclude that a direct cytotoxic effect of dexamethasone on brain tumor cells is not present in vivo because the tissue levels of the drug are about 1000 times lower than the LD50 in cell culture. PMID- 12117317 TI - The effects of (-)clausenamide on functional recovery in transient focal cerebral ischemia. AB - The effects of (-)clausenamide (clau) on spatial cognitive functions and hippocampal long-term potentiation (LTP) after transient focal cerebral ischemia in rats were investigated. Four weeks after middle cerebral artery occlusion, Morris water maze tasks demonstrated that 2 h of transient forebrain ischemia resulted in a significant decrease in spatial discrimination performance. The escape latency at 4 and 5 days of acquisition trial was lower in the ischemic rats than in sham-operated rats (33.8+/-6.7 sec and 26.8+/-5 sec versus 12.2+/ 4.0 sec and 10.4+/-3.6 sec), chronic treatment with clau (10 mg kg(-1) p.o. once daily) significantly improved the impairment (12.4+/-4.1 sec and 15.2+/-3.1 sec). After Morris water maze, the changes in population spike (PS) amplitude were recorded as an index of LTP in the perforant path-dentate gyrus synapses. There was no difference in PS amplitude between the sham-operated and vehicle-treated animals, whereas the fractional increase of PS 20-50 min after tetanus was significantly larger in clau-treated group. Histopathological analysis revealed that clau could protect against neuron loss in the regions of cortex and striatum. In conclusion, these data indicate a beneficial effect of clau for synaptic plasticity and cognitive function impaired by transient focal cerebral ischemia. PMID- 12117319 TI - Burr hole cover for ventriculoperitoneal shunts and ventriculostomy: technical note. AB - In patients who have undergone intracranial procedures, bone gaps or burr holes often result in small but undesirable scalp or skin depressions. The authors designed a burr hole cover for hydrocephalus shunt system or external ventricular drainage, which is shaped to alleviate the deformity of the burr hole by filling the bone defect and allowing the passage of the ventricular catheter. The specifications of this device and its clinical application are described. PMID- 12117320 TI - Feasibility and limitations of the rat model by C6 gliomas implanted at the subcutaneous region. AB - Although rat models implanted with C6 glioma cells have been widely utilized for the assessment of new therapeutic modalities, a convenient in vivo model is still required. We implanted C6 glioma cells into the brains and the abdominal subcutaneous regions of Wistar rats, and evaluated the volumes of the growing tumors. The cultures of 5 x 10(6) cells had successfully formed tumors at 100% (52/52) of sites in the subcutaneous regions on day 5 after implantation. It was easy to measure the visible tumors with a sliding caliper. The tumor volume at the subcutaneous region reached a maximum volume (950 mm3+/-167 SE) on day 15, after which it diminished. In contrast, tumors implanted in the intracerebral region showed a maximum volume on day 20 after the implantation of 1 x 10(5) cultured cells. Pathological examinations of the subcutaneous and intracranial tumors on day 15 showed similar findings exhibiting high nuclear cell ratio, mitosis and pseudopalisading with small populations of GFAP positive cells. The results suggested that rat C6 glioma models with implantation into the subcutaneous abdomen are available from day 5 to day 15 as a convenient model for the assessment of anti-proliferative drugs. The usefulness and the limitations of the rat C6 glioma model are discussed in this manuscript. PMID- 12117321 TI - Advanced atherosclerosis of the aortic arch is uncommon in ischemic stroke: an autopsy study. AB - Aortic and carotid atherosclerosis are known risk factors for stroke. The aim of the study was to determine the frequency of atherosclerotic lesions in the aorta and carotid arteries in subjects dying of ischemic and hemorrhagic stroke and to determine whether aortic atherosclerosis was associated with any specific ischemic stroke subtype. Autopsies were performed in 207 patients who died during hospitalization for stroke from 1993 to 1997. Subjects ranged in age from 37 to 98 years, mean 74.45 years (SD +/- 11.84). There were 132 women and 75 men. Stroke was hemorrhagic in 66 and ischemic in 141. Advanced atherosclerotic lesions were less frequent in the aortic arch (1.9%) than in the thoracic aorta (51.7%), abdominal aorta (60.4%), or carotid arteries (23.7%). Moderate atherosclerotic lesions in the aortic arch were observed more frequently in ischemic (75.2%) than hemorrhagic stroke (56.1%, p=0.026). Advanced or moderate atherosclerotic lesions in any part of the aorta did not predict ischemic stroke subtype. Advanced atherosclerosis of the carotids was more common in ischemic stroke (28.4%) than hemorrhagic stroke (13.6%, p < 0.05). Advanced atherosclerosis of the carotids was more common in stroke due to atherothrombosis (51.4%) than in stroke due to cardiac embolism (22.1%) or stroke of unknown etiology (5.6%). The low frequency of advanced atherosclerotic lesions of the aortic arch suggests that this disease process is not a common mechanism of stroke. PMID- 12117322 TI - Relationship of nidal vessel radius and wall thickness to brain arteriovenous malformation hemorrhage. AB - Cerebral (brain) arteriovenous malformations (BAVMs) are a tangle of disorganized vessels that are a rare cause of hemorrhagic stroke in the general population. Although clinical presentation of hemorrhage may be related to the structure of BAVM vessels, there has been no systematic quantitative analysis of BAVM vessel morphology. Histological sections of excised BAVM lesions were prepared from patients who presented with hemorrhage (n = 14) and from patients with no history of hemorrhage (n = 22). Mean values of radius and wall thickness in each section were determined. BAVM radii were 422+/-136 microm (mean +/- SD), minimum wall thickness (thinnest portion of the wall) was 54+/-14 microm; and the minimum thickness/radius ratio was 0.23+/-0.07. Greater vessel wall thickness was associated with hemorrhagic presentation (OR= 1.1; p = 0.046) after adjusting for feeding artery pressure. Because BAVM vessels from patients presenting with hemorrhage had thicker vessel walls, the search for structural properties predisposing BAVM rupture should be expanded beyond the morphological properties analyzed here. PMID- 12117323 TI - Klippel-Feil syndrome associated with posterior fossa dermoid cyst. Case report. AB - A patient with association of Klippel-Feil syndrome and posterior fossa dermoid cyst is presented. The patient, a 36-year-old man, presented with an acute obstructive hydrocephalus due to the cyst and exhibited the typical triad of the Klippel-Feil abnormality with short neck, low hairline implantation and limited neck motion along with a complex cervical vertebrae fusion. The anatomical and clinical features as well as the pathophysiology of this rare association are discussed after a review of the literature. PMID- 12117324 TI - Classification of medullary venous malformations in the temporal lobe: according to location and drainage pathway. AB - Medullary venous malformation (MVM) is rare in the temporal lobe, and the radiologic characteristics of temporal MVM have not yet been clarified. In 12 previously reported cases with satisfactory angiographic or magnetic resonance information as well as two newly reported here, we analyzed the specific location and hemodynamics of temporal lobe MVMs, particularly with respect to venous drainage. Temporal lobe MVM typically were seen in the superior lateral portion of the temporal lobe near either the atrium or the inferior horn of the lateral ventricle. Venous drainage was classified into two main patterns: deep (three cases) and superficial (11 cases). Superficial drainage could be divided into two subtypes: lateral and anterior. Dilated deep medullary veins converged toward either the lateral wall of the atrium or the inferior horn of the lateral ventricle. In the deep-drainage type, medullary veins drained into subependymal veins such as the inferior ventricular vein and the lateral atrial vein, and then emptied into the basal vein of Rosenthal. The anastomotic lateral mesencephalic vein was involved in one case as a variant of the basal vein. When the subependymal veins and/or the basal vein of Rosenthal or transverse sinus were hypoplasic, the medullary veins drained into either the Sylvian veins (anterior superficial type) or the vein of Labbe (lateral superficial type) through a characteristic large transcerebral vein. Drainage of temporal lobe MVM can be classified as deep, lateral superficial, or anterior superficial. PMID- 12117325 TI - Mouse model of subarachnoid hemorrhage associated cerebral vasospasm: methodological analysis. AB - The transgenic mouse has been used to study subarachnoid hemorrhage (SAH) induced delayed cerebral vasospasm (DCV). Methodological parameters have not been analyzed to validate this model and associated neurological deficits have not been described. We introduce a technique to quantify DCV and associated neurological deficits. C57BL/6J mice were subjected to SAH or sham surgery. Seventy-two hours later, the vasculature was cast in situ with India ink/gelatin at perfusion pressures of 40-60, 60-80, or 100-120 mmHg. Mice were perfused with and without microfiltration. Additional mice underwent grading of SAH size, measurement of vascular diameters, and neurological examination (score range 5 27; 27= normal). When cast at 60-80 mmHg, SAH was associated with an intraluminal cross-sectional diameter reduction in 3 of 7 ipsilateral vascular segments. At 40 60 mmHg, the diameter of only one segment was reduced. No changes were observed at 100-120 mmHg. Emboli prevented adequate perfusion of vascular segments in the absence of microfiltration. Median (interquartile range) neurologic score was reduced after SAH (sham, 27(27); SAH 11(7-17)). Deficits correlated with middle cerebral artery (MCA) diameter and SAH grade. MCA diameter also correlated with SAH grade. Only when utilizing microfiltration, controlling for hemorrhage size, and casting at perfusion pressures of 60-80 mmHg does India ink/gelatin vascular casting demonstrate consistent DCV that correspnds to neurological deficits. This allows measurement of both anatomical and clinical DCV in the mouse. PMID- 12117326 TI - Proliferating cell nuclear antigen positive cells in the hippocampal subgranular zone decline after irradiation in a rodent model. AB - Four-week-old ICR mice were systemically exposed to 18Gy X-rays. Afterwards, the expression of proliferating cell nuclear antigen (PCNA) in the cerebrum was observed with time using Western blot analysis and immunohistochemical staining. As a result, PCNA-positive cells were observed in the subgranular zone (SGZ) of the hippocampus and subventricular zone (SVZ) of the lateral ventricles in unirradiated mice. The number of PCNA-positive cells decreased with time in all zones after irradiation, but the decrease was more marked in the hippocampal SGZ. We think that PCNA-positive cells are stem cells. The selective vulnerability to radiation in the hippocampus is considered to be attributed to the fact that stem cells in the SGZ selectively undergo radiation-induced apoptosis. PMID- 12117327 TI - Classification and terminology of neck dissection. PMID- 12117328 TI - Neck dissection classification update: revisions proposed by the American Head and Neck Society and the American Academy of Otolaryngology-Head and Neck Surgery. PMID- 12117329 TI - Practice of pediatric otolaryngology: results of the future of pediatric education II project. AB - OBJECTIVES: To define the practice of pediatric otolaryngology compared with general otolaryngology and to estimate pediatric otolaryngology workforce utilization and needs. METHODS: Survey of members of the American Academy of Pediatrics Section on Otolaryngology and Bronchoesophagology and the American Society of Pediatric Otolaryngology and of a random sample of the membership of the American Academy of Otolaryngology-Head and Neck Surgery. RESULTS: Pediatric otolaryngologists were more likely to practice in urban and/or academic settings than were general otolaryngologists. Children (age <18 years) comprised over 88% of the patients of pediatric otolaryngologists and 30% to 35% of the patients of general otolaryngologists. Pediatric otolaryngologists were more likely to see children with complicated diseases such as airway disorders or congenital anomalies than were general otolaryngologists. Pediatric otolaryngologists, unlike general otolaryngologists, reported an increasing volume of pediatric referrals, as well as increased complexity in the patients referred. The surveyed physicians estimated the present number of pediatric otolaryngologists in their communities as approximately 0.2 to 0.3 per 100 000 people. CONCLUSIONS: Most children receiving otolaryngologic care in the United States receive such care from general otolaryngologists. The patient profile and practice setting of the subspecialty of pediatric otolaryngology differ from those of general otolaryngology. The demand for pediatric otolaryngologists appears to be increasing, but many general otolaryngologists do not believe there is an increased need. PMID- 12117330 TI - Pediatric otolaryngology: too much specialization? PMID- 12117331 TI - Asymmetric tonsil size in children. AB - OBJECTIVE: To assess the clinical implications of asymmetrically enlarged tonsils in children. DESIGN: A prospective controlled study of asymmetric tonsil size in children scheduled for tonsillectomy with or without adenoidectomy. Data were recorded on tonsil size and position, tonsillar fossa depth, degree of asymmetry, and pathological findings. Control patients were matched for age, sex, race, diagnosis, and surgical procedure. PATIENTS: A total of 258 children, aged 2 to 18 years, scheduled for tonsillectomy with or without adenoidectomy during a 27 month period. SETTING: A tertiary care academic medical center. RESULTS: Forty seven children (18.2%) were determined to have asymmetric tonsils. There were 43 matched controls with symmetric tonsils. Three-dimensional quantitative measurements of the resected tonsils revealed little or no actual asymmetry in tonsil size even though preoperative intraoral observations gave the impression that one tonsil was larger than the other. Statistically, tonsillar asymmetry was more apparent than real. When measured by volume, there was asymmetry in both groups. However, there was no statistical difference in the degree of asymmetry between the groups (P =.50). A difference in the depth of the tonsil fossa contributed to the putative asymmetry (P<.001). No malignant neoplasms were identified on microscopic examination in either group. CONCLUSIONS: Tonsillar asymmetry in children may often be an illusion secondary to a difference in the depth of the tonsillar fossa. Tonsillar asymmetry in children in the absence of other findings such as ipsilateral cervical adenopathy or other constitutional symptoms may not indicate a malignancy. PMID- 12117332 TI - Child behavior and quality of life before and after tonsillectomy and adenoidectomy. AB - OBJECTIVE: To determine the relationship between child behavior and quality of life before and after tonsillectomy and adenoidectomy by means of a standardized assessment of child behavior, the Child Behavior Checklist (CBCL), and a validated quality-of-life survey of pediatric obstructive sleep apnea, the OSA 18. DESIGN: Before-after study. SETTING: Hospital-based pediatric otolaryngology practice in a metropolitan area. PARTICIPANTS: Sixty-four children (mean [SD] age, 5.8 [3.1] years; 36 boys, 28 girls) who underwent tonsillectomy and adenoidectomy for treatment of sleep-disordered breathing or recurrent tonsillitis. INTERVENTION: Parents or caretakers completed the OSA-18 and the CBCL for ages 2 to 3 years or 4 to 18 years before surgery and 3 months postoperatively. MAIN OUTCOME MEASURES: The OSA-18 mean survey scores and change scores, and the CBCL total problem T scores and change in total problem T scores. RESULTS: The mean (SD) preoperative OSA-18 score was 3.9 (1.5) and change score was 2.3 (95% confidence interval, 1.9-2.7). The mean total problem score was 7.3 points lower after surgery (95% confidence interval, 4.9-9.7), indicating a significant decrease (P<.001, matched t test). The preoperative CBCL total problem score was consistent with abnormal behavior for 16 children (25%), but only 5 children (8%) scored in the abnormal range postoperatively (P =.03, log likelihood ratio test). The OSA-18 preoperative mean survey score had fair to good correlation with the preoperative CBCL total problem T score (r = 0.50, P<.001, Pearson correlation), and the OSA-18 change score had fair to good correlation with the change in CBCL total problem T score (r = 0.54, P<.001, Pearson correlation). CONCLUSIONS: Behavioral and emotional difficulties are found in children with sleep-disordered breathing before treatment and improve after intervention. Scores on a standardized measure of assessment of behavior demonstrate significant correlation with scores on a validated quality-of-life instrument. PMID- 12117334 TI - Can mumps vaccine induce remission in recurrent respiratory papilloma? AB - OBJECTIVE: To describe our experience using laser excision and locally injected mumps vaccine to induce remission in patients with recurrent respiratory papilloma (RRP). SETTING: Tertiary care regional medical center. PARTICIPANTS: Initially, 11 children with RRP treated in a pilot study with laser excision at regular intervals for at least a year without adjuvant therapy; later, a series of 18 children and 20 adults with RRP, some of whom had used various adjuvant therapy with interval laser excision. INTERVENTIONS: Both patient groups continued their same interval laser excision with the same or similar laser, same clinical setting, and same surgeon. Locally injected mumps vaccine was then administered into the excision site after each laser removal of papilloma. OUTCOME MEASURES: Larynx and trachea were microphotographed with each treatment. Two consecutive disease-free intervals and a follow-up of at least 1 year were required criteria for remission. RESULTS: In the pilot study, remission was induced in 9 (82%) of 11 patients by 1 to 10 injections, with follow-up of 5 to 19 years. In the subsequent series, remission was induced in 29 (76%) of 38 patients by 4 to 26 injections, and follow-up was 2 to 5 years. CONCLUSIONS: Combined with serial laser excision, mumps vaccine positively influences induction of remission in children with RRP. The mechanisms of this effect are unclear, but the treatment is readily available, inexpensive, and has a low risk of adverse effects. PMID- 12117333 TI - Quality-of-life outcomes after surgical intervention for otitis media. AB - OBJECTIVE: To assess the change in disease-specific quality of life in children with recurrent acute otitis media and/or chronic otitis media with effusion treated with surgical intervention. DESIGN: Prospective questionnaire-based outcome study. SETTING: An academic pediatric otolaryngology practice. PARTICIPANTS: Consecutive series of 123 children referred for surgical treatment of recurrent acute otitis media and/or chronic otitis media with effusion. INTERVENTION AND METHODS: Surgery included bilateral myringotomy and tympanostomy tube placement either alone or with adenoidectomy. An otitis media disease specific questionnaire was administered before and after surgical intervention. MAIN OUTCOME MEASURES: Comparison of the mean percentage change in total ear symptom score between presurgery and postsurgery scores at 1 and 6 months after surgery. RESULTS: The mean percentage change in total ear symptom score was a 74.5% improvement (P<.001) at the 1-month follow-up and a 59.8% improvement (P<.001) at the 6-month follow-up. Parental worry related to the child's ear problems was also significantly decreased, with a mean otitis media disease specific questionnaire score of 3.43 (P<.001) at 1 month and 2.64 (P<.001) at 6 months after surgery. When caregivers were asked if they would have their child undergo tympanostomy tube placement if they had to make the decision again, 91% and 84% responded yes at the 1- and 6-month follow-up, respectively. CONCLUSIONS: The disease-specific quality of life of children with recurrent acute otitis media and/or chronic otitis media with effusion with appropriate surgical indications significantly improved after surgical intervention. The amount of parental worry concerning their children's ear problems also significantly improved following surgery, and most caregivers would opt again for tube placement. PMID- 12117335 TI - Another potential adjuvant therapy for recurrent respiratory papillomatosis. PMID- 12117336 TI - Obstructive adenoid tissue: an indication for powered-shaver adenoidectomy. AB - OBJECTIVES: To quantify the incidence of intranasal extension of adenoid tissue and residual adenoidal obstruction of the posterior choanae following traditional curette adenoidectomy to determine the efficiency of adenoid curettage and the usefulness of intraoperative endoscopic examination and powered-shaver adenoidectomy in achieving better postnasal patency. DESIGN: Prospective intraoperative endoscopic evaluation of the posterior choanae and nasopharynx of a case series of 130 patients before and after curette and powered-shaver adenoidectomy. SETTING: Tertiary referral center. PATIENTS: One hundred thirty consecutive pediatric patients with obstructive adenoidal hypertrophy undergoing adenoidectomy. MAIN OUTCOME MEASURES: The degree of residual postnasal obstruction due to adenoid tissue was assessed endoscopically (grades 0-3) after curette and adjuvant powered-shaver adenoidectomy. The presence of intranasal adenoid tissue was also recorded. RESULTS: Following traditional curette adenoidectomy, 51 (39%) of 130 patients had residual obstructive adenoid with 42 patients (32%) having occlusive intranasal adenoid tissue. Having determined the presence of remaining obstructive tissue with intraoperative nasal endoscopy in these 51 patients, complete airway patency was achieved with powered-shaver adenoidectomy. CONCLUSION: The presence of intranasal extension of adenoids obstructing the posterior choanae is common in children with adenoid hypertrophy. Traditional adenoidectomy is ineffective in removing this tissue and may also leave obstructive tissue high in the nasopharynx. Intraoperative nasal endoscopy allows assessment of the completeness of surgery. Powered-shaver adenoidectomy enables complete removal of obstructive adenoid tissue thereby ensuring postnasal patency. PMID- 12117337 TI - Powered partial adenoidectomy. AB - OBJECTIVE: To confirm our clinical impression that the powered microdebrider is superior to curettes for performing partial adenoidectomy (removal of the superior one half to three fourths of the adenoid pad). DESIGN: Observational study of 100 children undergoing partial adenoidectomy with the powered microdebrider compared with 40 children undergoing conventional partial adenoidectomy with curettes. SETTING: Private and public tertiary care centers. PATIENTS: All patients younger than 20 years undergoing partial adenoidectomy at the respective institutions during the study period. INTERVENTIONS: Partial adenoidectomy as indicated for chronic otitis media, airway obstruction, or chronic or recurrent tonsillitis with either the powered microdebrider or curettes. MAIN OUTCOMES MEASURES: Operative time (with specific quantification of the time required for tissue removal and hemostasis), blood loss, complications, and subjective ease of use. RESULTS: Operative time was 59% shorter for the microdebrider group (mean, 3 minutes 22 seconds; range, 1 minute 6 seconds to 12 minutes 45 seconds) than for the conventional group (mean, 8 minutes 8 seconds; range, 1 minute 2 seconds to 22 minutes 0 seconds) (P<.001). Blood loss was comparable for both groups (powered group: mean, 2.0 mL/kg; range, 0.4 to 9.4 mL/kg; conventional group: mean, 2.0 mL/kg; range, 0.3 to 6.7 mL/kg; P=.34). There were no intraoperative or postoperative complications in either group. Surgeon satisfaction with the microdebrider was high. CONCLUSIONS: The powered microdebrider for partial adenoidectomy is quicker and is not associated with blood loss or complications above that of conventional partial adenoidectomy. The degree of control afforded by the microdebrider technique is of utmost value in preventing complications such as velopharyngeal insufficiency, and this is now our procedure of choice. PMID- 12117338 TI - Computer-assisted navigation system in pediatric intranasal surgery. AB - OBJECTIVES: To introduce a computer-assisted navigation system and to evaluate its application in pediatric sinusonasal surgery. METHODS: A commercially available wireless passive marker system that allows the calibration and tracking of virtually any instrument was adapted to children and used during pediatric endoscopic sinusonasal surgery. RESULTS: The headset localizer that was initially used in computed tomographic scanning was not well accepted by children. Correlation of the preoperative computed tomographic scan to the actual patient was made possible by a laser device. Setup time was able to be decreased from an initial 20 minutes to 3 minutes. The average recording accuracy was 1.1 mm. The advantages of the system became apparent as experience increased in cases involving sinus polyposis, choanal atresia, nasopharyngeal fibroma removal, tumor biopsy, and minimally invasive maxillary, frontal, and sphenoidal surgery. CONCLUSIONS: The computer-assisted navigation system was used first as a control system and then, as experience increased, as a true surgical guide. Indications for its use also increased. Pediatric intranasal surgery was performed using 2 complementary guides: an endoscopic view and a computed tomographic view of the instrument's position. PMID- 12117339 TI - Bilateral submandibular gland excision with parotid duct ligation for treatment of sialorrhea in children: long-term results. AB - BACKGROUND: Multiple procedures have been advocated for the surgical control of chronic sialorrhea in children. However, some of them are associated with significant complications or only short-term success. OBJECTIVES: To evaluate the safety of bilateral submandibular gland excision (SGE) with parotid duct ligation (PDL) and to assess its long-term complications and efficacy in the treatment of chronic sialorrhea in children. DESIGN: Case series. Telephone interview of patients' families. SETTING: Tertiary care children's hospital. PATIENTS: Ninety three patients with chronic sialorrhea who underwent bilateral SGE with PDL from 1988 to 1997. MAIN OUTCOME MEASURES: Operative and postoperative complications, length of postoperative hospitalization, postoperative drooling, care requirements, xerostomia, dental caries, and overall satisfaction. RESULTS: The mean postoperative stay was 2.4 days. There were 3 postoperative complications. Seventy-two families were interviewed (follow-up time, 1-10 years): 62 (87%) reported no further drooling or significant improvement; 7 reported the occurrence of dry mouth; and 2 reported an increase in dental caries. CONCLUSION: Bilateral SGE with PDL is a safe and consistently efficient procedure for the treatment of chronic sialorrhea in children. PMID- 12117340 TI - The natural history of congenital cholesteatoma. AB - OBJECTIVES: To describe the natural history of congenital cholesteatoma (CC) and to determine whether such a description provides clues about the origins and end points of these lesions. DESIGN: A retrospective qualitative analysis of intraoperative illustrations of 34 consecutive patients with 35 CCs (1 bilateral). SETTING: Two tertiary care children's hospitals. PATIENTS: Thirty four children with CC, mean age, 5.6 years (range, 2-13 years). RESULTS: Congenital cholesteatoma originates generally, but not universally, in the anterior superior quadrant. The progression of growth is toward the posterior superior quadrant and attic and then into the mastoid. Contact with the ossicular chain generally results in loss of ossicular continuity and in conductive hearing loss. CONCLUSIONS: Congenital cholesteatoma appears to have a predictable trajectory of growth, starting as a small pearl in the middle ear, eventually growing to involve the ossicles and mastoid, and causing varying degrees of destruction and functional impairment. The clinical picture of a young child with otorrhea, conductive hearing loss, tympanic membrane perforation in a nontraditional location, and a mastoid filled with cholesteatoma may represent the end point in the natural history of CC, despite the fact that this type of lesion is outside the accepted definition of CC. PMID- 12117341 TI - Congenital cholesteatoma: classification, management, and outcome. AB - OBJECTIVES: To assess whether a classification system for congenital cholesteatoma (CC) can be derived from analysis of a large clinical sample of cases and to assess whether such a classification system is a reliable guide for surgical intervention, reexploration, and hearing outcome. DESIGN: A retrospective review of clinical and surgical records of 119 patients with CC. SETTING: Four tertiary care children's hospitals. PATIENTS: One hundred nineteen children with CC (age range, 2-14 years). RESULTS: Congenital cholesteatomas in the anterior mesotympanum were treated successfully with exploratory tympanotomy. Congenital cholesteatomas involving the posterior superior quadrant and the attic usually had concurrent involvement of the incus and stapes and often required a canal wall up tympanomastoidectomy and a second look for its control. Congenital cholesteatoma involving the mastoid usually involved all of the ossicles, was inconsistently controlled with canal wall up tympanomastoidectomy, and had a poor prognosis for restoration of conductive hearing loss. The mean +/- SD age of children with CC was 5.6 +/- 2.8 years, while that of children with acquired cholesteatoma was 9.7 +/- 3.3 years. CONCLUSIONS: The sequence of spread of CC, involving 3 sites, suggests a natural classification system. The CC usually originates in the anterior superior quadrant, but does not consistently remain there, and may variably occupy the middle ear and mastoid and result in ossicular destruction and conductive hearing loss. The location of CC and the involvement of the ossicles is an accurate predictor of the type of surgery necessary for its control and for the success of hearing restoration. PMID- 12117342 TI - The familial aggregation of pediatric obstructive sleep apnea syndrome. AB - OBJECTIVE: To determine the role of genetic mechanisms in the development of pediatric obstructive sleep apnea syndrome (OSAS). DESIGN: Genetic-epidemiologic survey of families of index children with laboratory-confirmed OSAS. SETTING: Tertiary care academic medical center. PARTICIPANTS: Six-hundred nap polysomnograms performed in our institution's pediatric sleep laboratory over a 6 year period (1994-2000) were reviewed, and the 497 children who tested positive for OSAS were selected. A caretaker of 200 of these index patients was contacted, and 115 were enrolled in the study. INTERVENTION AND MAIN OUTCOME MEASURE: Questionnaire-type telephone interviews were conducted with the current caretakers of the index patients to assess the distribution of sleep-disordered breathing in the first-degree relatives. RESULTS: Data were collected for 445 first-degree relatives (256 adults and 189 children) of the 115 index patients. Habitual snoring was found in 194 (43.6%) of the family members, while symptoms highly suggestive of OSAS (nighttime "gasping for air" or "cessation of breathing") were found in 91(20.4%). Sixty-eight (26.6%) of the adult first degree relatives and 23 (12.2%) of the pediatric first-degree relatives had symptoms highly suggestive of OSAS. Of the 115 index children, 50 (43.5%) had at least 1 relative with symptoms highly suggestive of OSAS; 6 (1.3%) of the first degree relatives had sleep study results positive for OSAS, 4 (0.9%) were using nasal continuous positive airway pressure, and 21 (4.7%) had prior surgery for the treatment of OSAS. CONCLUSION: Considering the established prevalence of OSAS in the general population (2%-4%), the results of this study support a familial basis for this disorder. PMID- 12117343 TI - Validation of the snore outcomes survey for patients with sleep-disordered breathing. AB - OBJECTIVE: To develop and validate a self-reported outcomes measure for patients with sleep-disordered breathing-the Snore Outcomes Survey. DESIGN: Item areas of the SOS were developed by an expert panel. Consecutive patients were enrolled into the study in a prospective manner. Patients received the SOS, the Epworth Sleepiness Scale, the Pittsburgh Sleep Quality Index, the Medical Outcomes Study 36-Item Short-Form Health Survey, and standard overnight polysomnography at baseline and after 4 months of continuous positive airway pressure therapy. SETTING: A tertiary care, academic otolaryngology and sleep disorders referral center. PATIENTS: One hundred fifty-six adult patients presenting with a chief complaint of snoring or sleep-disordered breathing to the Massachusetts Eye and Ear Infirmary, Boston. MAIN OUTCOME MEASURES: Test-retest reliability, intrasurvey reliability, internal consistency, validity, and standardized response means of the SOS. RESULTS: Overall, reliability of the SOS was excellent (test-retest reliability r = 0.86; P<.001; Cronbach alpha coefficient, 0.85). The SOS index significantly correlated with the Epworth Sleepiness Scale (r = - 0.42; P<.001) and the global Pittsburgh Sleep Quality Index score (r = - 0.38; P<.001), as well as with the number of recorded arterial oxygen saturation levels below 85% (r = - 0.46; P =.02). The SOS index was sensitive to clinical changes after intervention (standardized response mean, 0.57). CONCLUSION: The SOS is a reliable and valid instrument for assessing sleep-related health status for patients with snoring and sleep-disordered breathing and for measuring change in health status following therapy. PMID- 12117344 TI - Impact of nodal metastases on prognosis in patients with well-differentiated thyroid cancer. AB - OBJECTIVES: To study the clinical and pathological variables predicting lymph node metastases in patients with well-differentiated thyroid carcinoma and to examine the impact of these metastases on recurrence and survival. DESIGN: Cohort study. Median follow-up, 56 months. SETTING: Tertiary referral university teaching hospital. PATIENTS: The study included 522 consecutive patients with well-differentiated thyroid carcinoma treated between 1964 and 1999. Data were collected on age, sex, family history of thyroid disease, prior radiation exposure, stage of disease, pathological diagnosis, size of tumor, multifocality of disease, recurrence, and survival. INTERVENTION: Total thyroidectomy and postoperative iodine 131 ablation. MAIN OUTCOME MEASURES: Disease-free and overall survival. RESULTS: A total of 347 patients with stage I disease and 118 with stage II disease were identified. The median age of patients with neck disease was 3 years younger than those without neck disease and most had papillary carcinoma. Patients with multifocal disease were more likely to have neck disease (P =.02). On univariate analysis, disease-free and overall survival rates were significantly lower in patients who presented with neck node metastases (P<.001 and P =.005); this difference in survival remained highly significant on multivariate analysis for disease-free survival (P =.001), with a relative hazard of 6.27. CONCLUSIONS: When treated with total thyroidectomy and routine postoperative iodine 131 ablation, patients with well-differentiated thyroid carcinoma who present with neck node metastases outside the central compartment of the neck have an approximately 6-fold risk of developing recurrences, most of which occur in the neck. PMID- 12117345 TI - Acute pain management following laryngectomy. AB - OBJECTIVES: To evaluate the adequacy of as-needed (prn) dosing of narcotics during the acute postoperative period following laryngectomy and to evaluate the role of nurses' interpretation and implementation of narcotic orders in postoperative pain management. STUDY DESIGN: A retrospective review of the medical records of 37 patients who underwent laryngectomy at the University of Oklahoma. The postoperative care was standardized through a clinical pathway to provide a uniform level of care. METHODS: The parameters reviewed include (1) the type and dose of analgesic prescribed, (2) the quantity and frequency of analgesic administered to each patient, and (3) the adequacy of the initial pain control prescription. RESULTS: All physician orders for narcotics were at or above the minimum dosing guidelines; 68% met a recommended adequate postoperative prescription for moderate pain. However, none of the patients actually received the intended dose during a 24-hour period while hospitalized. Physicians were contacted about 13 patients (35%) because of inadequate pain relief, but only 8 patients (22%) had their narcotic dose increased appropriately. Patients were dosed below the minimum prescribed dose 19 times (2.8%), and in 24 instances (3.6%) the backup analgesic, designated as "prn breakthrough pain," was given as the primary analgesic. CONCLUSIONS: As-needed dosing of analgesia resulted in suboptimal pain control for at least 35% of patients undergoing laryngectomy. Inadequate prescription and variable implementation of prn orders contributed to this. PMID- 12117346 TI - Cancer of the external auditory canal. AB - OBJECTIVE: To evaluate the outcome of surgery for cancer of the external auditory canal and relate this to the Pittsburgh staging system used both on squamous cell carcinoma and non-squamous cell carcinoma. DESIGN: Retrospective case series of all patients who had surgery between 1979 and 2000. Median follow-up was 47 months (range, 2-148 months). Data on age, sex, symptoms, TNM status, histopathological diagnosis, surgery, adjunctive therapy, sequelae, recurrence, and status at follow-up were obtained. SETTING: An ear, nose, and throat department in an ambulatory and hospitalized care center. PATIENTS: Ten women and 10 men with previously untreated primary cancer. Median age at diagnosis was 67 years (range, 31-87 years). Survival data included 18 patients with at least 2 years of follow-up or recurrence. INTERVENTION: Local canal resection or partial temporal bone resection. MAIN OUTCOME MEASURE: Recurrence rate. RESULTS: Half of the patients had squamous cell carcinoma. Thirteen of the patients had stage I tumor (65%), 2 had stage II (10%), 2 had stage III (10%), and 3 had stage IV tumor (15%). Twelve patients were cured. All patients with stage I or II cancers were cured except 1 with adenoid cystic carcinoma. No patients with stage III or IV cancer were cured. All recurrences developed in patients with incompletely resected tumors. CONCLUSIONS: The outcome was related to the stage of disease, suggesting that the Pittsburgh staging system is useful also in patients with non squamous cell carcinoma. Patients with early cancer benefited from a less aggressive surgical approach, while survival was poor in patients with advanced cancer with incompletely resected tumors despite adjuvant radiotherapy. PMID- 12117347 TI - Downsizing of voice prosthesis diameter in patients with laryngectomy: an in vitro study. AB - In patients with laryngectomy, voice prostheses inserted into a tracheoesophageal fistula (TEF) are widely used for vocal rehabilitation. Gradual dilation of the TEF may cause bothersome leakage around voice prostheses. Prosthesis-related weight and mechanical trauma possibly exacerbate TEF dilation. If prosthesis size were to be decreased, with a concomitant decrease in prosthesis weight and diameter, dilation of the TEF would probably lessen. We performed in vitro tests to study the effects on aerodynamic prosthesis function when the prosthesis size in particular, the inner diameter-was decreased. The effects on airflow and pressure were specifically studied in the airflow range of patients with laryngectomy. A 1-mm decrease of the regular inner prosthesis diameter from 5 mm to 4 mm showed no significant aerodynamic consequences at the average laryngectomized airflow point. Also, such a 1-mm decrease in diameter involved a prosthesis weight reduction of 18%. In view of these findings, downsizing the standard prosthetic diameter should be considered in future voice prosthesis development. PMID- 12117349 TI - Scopolamine and the murder of King Hamlet. PMID- 12117348 TI - Management of the keel nose and associated valve collapse. AB - OBJECTIVES: To analyze the anatomical abnormality of the keel nose and correlate the findings with etiologic maneuvers of a routine rhinoplasty procedure; to identify the contributing factors and offer suggestions to avoid or decrease the severity of these surgical complications; and to present an effective revisional procedure to correct the functional and cosmetic consequences of this deformity. PATIENTS AND METHODS: A total of 47 patients (31 women and 16 men; age range, 18 71 years) with a keel-appearing nose presented for revision rhinoplasty. All had undergone at least 1 rhinoplasty procedure, and 39 had undergone 2 or more previous nasal procedures. All patients had bilateral lateral nasal wall collapse and an associated severely compromised internal nasal valve. All patients underwent reconstruction with a conchal cartilage overlay graft. RESULTS: All patients had a moderate to excellent cosmetic improvement; the subjective improvement in nasal airway was more dramatic. Since patients with a keel nose have an associated internal valve collapse, both abnormalities are addressed simultaneously with the conchal cartilage overlay repair, which results in minimal morbidity with no major complications. CONCLUSIONS: Conchal cartilage overlay repair uses a cartilage graft from the auricle with a recommended external rhinoplasty for placement. Appropriate sizing and fashioning precede the precise placement and suture fixation. This technique addresses both functional and cosmetic abnormalities. PMID- 12117351 TI - Pathology quiz case. Disseminated blastomycosis. PMID- 12117350 TI - Dysphagia due to a large schwannoma of the oropharynx: case report and review of the literature. AB - Schwannoma is a benign, encapsulated tumor that is derived from Schwann cells. Approximately 25% to 45% of schwannomas occur in the head and neck. The most common site is the parapharyngeal space of the neck; intrapharyngeal occurrence is extremely rare. To our knowledge, this is the first report of a pedunculated schwannoma in the supraglottic oropharynx. Because of the location and mass of the tumor, the main symptom was dysphagia. The tumor was excised via direct microlaryngoscopy, and no recurrence was seen after 2 years of follow-up. When schwannomas are located in the pharynx, they may cause dyspnea and dysphagia or impair phonation. Therefore, when dysphagia is present, a thorough diagnostic procedure should be performed to evaluate the morphology and function of the upper aerodigestive tract. PMID- 12117352 TI - Radiology quiz case. Papillary neoplasm of the endolymphatic sac in a patient with von Hippel-Lindau disease. PMID- 12117353 TI - Cardiac surgery and magnetic resonance imaging of the brain. PMID- 12117354 TI - The role of hypothermia in the management of severe brain injury: a meta analysis. AB - CONTEXT: Hypothermia is utilized in the management of severe traumatic brain injury despite the lack of unequivocal evidence supporting its use. Because of its widespread use, the effects of hypothermia are a concern. OBJECTIVE: To determine the effectiveness of hypothermia in the management of severe brain injury. DATA SOURCES: Two investigators working independently abstracted data in a blinded fashion from studies identified using multiple literature databases, including MEDLINE, Ovid, PubMed, the Cochrane Database of Systematic Reviews, EMBASE, and the abstract center for the American Association of Neurological Surgery and the Congress of Neurological Surgery, as well as the bibliographies of these articles. Additionally, experts in the field of hypothermia and neurotrauma provided additional references. STUDY SELECTION: Seven studies met predetermined inclusion criteria: (1) the study was a randomized clinical trial comparing the efficacy of hypothermia vs normothermia in patients with posttraumatic head injury, (2) only subjects aged 10 years or older were included in the study, and (3) relative risks (odds ratios [ORs], cumulative incidence, or incidence density measures) and 95% confidence intervals (CIs) or weighted mean differences and 95% CIs could be calculated from the data presented in the article. These criteria were applied in a blinded fashion by 2 independent investigators. DATA EXTRACTION: No single outcome variable was evaluated in all studies. The following outcome variables were assessed: intracranial pressure, Glasgow Outcome Scale score, pneumonia, cardiac arrhythmia, prothrombin time, and partial thromboplastin time. Either ORs or weighted mean differences (when the data provided did not permit calculation of an OR) comparing the effects of hypothermia vs normothermia were calculated from the data provided. DATA SYNTHESIS: The weighted mean difference (hypothermia - normothermia) for intracranial pressure was -2.98 mm Hg (95% CI, -7.58 to 1.61; P =.2). The OR (hypothermia vs normothermia) for Glasgow Outcome Scale score was 0.61 (95% CI, 0.26-1.46; P =.3). The OR for pneumonia was 2.05 (95% CI, 0.79-5.32; P =.14). The OR for cardiac arrhythmia was 1.27 (95% CI, 0.38-4.25; P =.7). The weighted mean difference for prothrombin time was 0.02 seconds (95% CI, -0.07 to 0.10; P =.7). The weighted mean difference for partial thromboplastin time was 2.22 seconds (95% CI, 1.73-2.71; P<.001). CONCLUSIONS: This meta-analysis of randomized controlled trials suggests that hypothermia is not beneficial in the management of severe head injury. However, because hypothermia continues to be used to treat these injuries, additional studies are justified and urgently needed. PMID- 12117355 TI - Mechanisms of action of the 5-HT1B/1D receptor agonists. AB - Recent studies of the pathophysiology of migraine provide evidence that the headache phase is associated with multiple physiologic actions. These actions include the release of vasoactive neuropeptides by the trigeminovascular system, vasodilation of intracranial extracerebral vessels, and increased nociceptive neurotransmission within the central trigeminocervical complex. The 5-HT(1B/1D) receptor agonists, collectively known as triptans, are a major advance in the treatment of migraine. The beneficial effects of the triptans in patients with migraine are related to their multiple mechanisms of action at sites implicated in the pathophysiology of migraine. These mechanisms are mediated by 5-HT(1B/1D) receptors and include vasoconstriction of painfully dilated cerebral blood vessels, inhibition of the release of vasoactive neuropeptides by trigeminal nerves, and inhibition of nociceptive neurotransmission. The high affinity of the triptans for 5-HT(1B/1D) receptors and their favorable pharmacologic properties contribute to the beneficial effects of these drugs, including rapid onset of action, effective relief of headache and associated symptoms, and low incidence of adverse effects. PMID- 12117356 TI - Brain damage after coronary artery bypass grafting. AB - BACKGROUND: Coronary artery bypass grafting (CABG) is associated with a risk for focal neurological deficits and neuropsychological impairment postoperatively. OBJECTIVES: To examine the brain damage after CABG using diffusion-weighted magnetic resonance imaging and (1)H-magnetic resonance spectroscopy (MRS) and to correlate the results with neurological and neuropsychological findings. PATIENTS AND METHODS: Thirty-five consecutive patients undergoing elective CABG were included. Patients underwent a neurological and neuropsychological examination before and after CABG. The magnetic resonance protocol was applied before and after (mean, 3 days) surgery and included a diffusion-weighted sequence and single-voxel MRS measurements in the frontal lobes. RESULTS: None of the patients revealed a new focal neurological deficit after surgery. Diffusion-weighted magnetic resonance imaging demonstrated new ischemic lesions in 9 (26%) of the patients. The presence of an ischemic lesion was not related to impaired postoperative test performance (P>.50). The apparent diffusion coefficient values in the cerebellum and the centrum semiovale exhibited an increase after surgery (P<.01), consistent with vasogenic edema. Following surgery, MRS revealed a significant decrease in the metabolite ratio of N-acetylaspartate-creatine (mean +/- SD, 1.69 +/- 0.20 vs 1.52 +/- 0.19; P<.001). The extent of deterioration in neuropsychological test performance after surgery was closely related to the degree of the N-acetylaspartate-creatine ratio decrease (P<.01). A follow-up MRS scan revealed a normalization of the N-acetylaspartate-creatine ratio, which accompanied the recovery in psychological test performance. CONCLUSIONS: Postoperative impairment in neuropsychological test performance is associated with a transient metabolic neuronal disturbance. Focal ischemic lesions after CABG are more frequent than the apparent neurological complication rate; however, they are not related to the diffuse postoperative encephalopathy. PMID- 12117357 TI - Clinical and neuropathological characteristics of hippocampal sclerosis: a community-based study. AB - BACKGROUND: Hippocampal sclerosis (HS) is a neuropathologic finding characterized by neuronal loss and gliosis in the CA-1 and subiculum of the hippocampus. Previous studies of HS have shown that this is a common postmortem finding in elderly subjects with dementia. However, these studies were from selected samples and therefore are not necessarily representative of patients seen in the general medical community. OBJECTIVES: To examine the clinical and pathologic characteristics of HS in a community-based case series of dementia and to compare these characteristics with those observed in subjects with Alzheimer disease (AD) from the same study sample. METHODS: One hundred thirty-four autopsy cases were available from a community-based registry of dementia. Sixteen cases (12%) had a postmortem diagnosis of HS. Thirty-two comparison control cases with a neuropathologic diagnosis of AD were selected from the same files. Each case of HS was reviewed for HS neuropathologic features, including severity, distribution, and additional pathologic processes. Blinded review of clinical characteristics for the HS and control groups was performed to assess risk factors. RESULTS: There was a wide range of severity and distribution of HS lesions between cases and substantial variability in lesion severity and age within individual cases. Serial neuropsychologic and behavioral assessments revealed similar clinical features and rates of dementia progression between HS and AD groups. Of all neuropsychologic tests performed at enrollment, only enhanced performance on Trails A differentiated the HS from the AD group (64 seconds, 0 errors vs 114 seconds, 0.6 errors; P< or = .05). The number of AD cases with at least 1 apolipoprotein epsilon 4 allele was significantly greater than the HS cases (61% vs 31%; chi(2) = 3.81, P< or = .05). Although medical record review indicated higher frequencies of clinical stroke and neuroradiologic white matter abnormalities in the HS group, risk factors for vascular disease and neuropathologic evidence of cerebrovascular disease did not differ between the groups. CONCLUSIONS: Our results suggest that HS is a frequent pathologic finding in community-based dementia. Individuals with HS have similar initial symptoms and rates of dementia progression to those with AD and therefore are frequently misclassified as having AD. Our clinical and pathologic findings suggest that HS has characteristics of a progressive disorder although the underlying cause remains elusive. PMID- 12117358 TI - Prevalence of dementia and dementing diseases in Japan: the Tajiri project. AB - BACKGROUND: Vascular dementia (VaD) has been considered to be more prevalent than Alzheimer disease in Japan. However, this might be the result of overdiagnosis stemming from some problematic diagnosis of VaD or of the frequent use of magnetic resonance imaging to detect cerebrovascular disease in older adults. OBJECTIVES: We investigated the prevalence of dementia and the ratios of dementing diseases. The effects of different criteria for VaD (DSM-IV, Alzheimer's Disease Diagnostic and Treatment Centers [ADDTC], and National Institute of Neurological Disorders and Stroke and the Association Internationale pour la Recherche et l'Enseignement en Neurosciences [NINDS-AIREN]) were considered. Hippocampal atrophy and vascular contribution to dementia were evaluated using magnetic resonance imaging findings. METHODS: We targeted all residents 65 years and older (n = 3207) in Tajiri, Japan, and examined 1654 (participant group 1). Of these, 564 (participant group 2) were randomly selected, and 497 underwent magnetic resonance imaging and diagnosis of dementing diseases. RESULTS: We found the overall prevalence of dementia to be 8.5% (141/1654) in participant group 1. Of these, 21 (14.9%) had a history of stroke. Of the 113 participants who had a history of stroke independent of dementia, 18.6% (21/113) were demented. For participant group 2 (n = 497), 32 were demented. The ratio among the dementia for probable VaD based on the NINDS-AIREN criteria was 18.8% (6/32), whereas that for ischemic vascular dementia was 31.3% (10/32) according to the ADDTC criteria. CONCLUSION: We confirmed the overall prevalence of dementia in adults 65 years and older to be 8.5%. We found that VaD was not a common disorder according to the NINDS-AIREN criteria. Rather, the condition of possible Alzheimer disease with cerebrovascular disease was more common. PMID- 12117359 TI - Down-regulation of survivin expression in T lymphocytes after interferon beta-1a treatment in patients with multiple sclerosis. AB - BACKGROUND: Treatment with interferon beta reduces clinical exacerbations in multiple sclerosis (MS) through several immunomodulatory mechanisms that involve the augmentation of programmed cell death (apoptosis) of peripheral T lymphocytes. The expression of survivin, a cell cycle-regulated antiapoptosis protein, is up-regulated in mitogen-stimulated T lymphocytes from patients with MS, and this expression correlates with MS disease activity. OBJECTIVE: To evaluate the effect of interferon beta on the expression of survivin and other apoptosis regulatory molecules in peripheral T lymphocytes from patients with MS. PATIENTS AND METHODS: In a prospective, combined clinical and immunologic study, we evaluated the expression of survivin, Bcl-2 protein, and the death receptor Fas in mitogen-stimulated T lymphocytes from 26 patients with MS, before and serially after treatment with interferon beta-1a. We also investigated the long term effects of interferon beta-1a on cellular expression of these proteins and T lymphocyte apoptosis in a cross-sectional study of 19 patients with MS receiving long-term interferon beta-1a therapy. RESULTS: Treatment with interferon beta-1a reduced the expression of survivin in in vitro stimulated T lymphocytes. This reduced expression correlated with augmented T-cell susceptibility to apoptosis and with clinical response to treatment. In contrast, interferon beta-1a therapy did not significantly alter cellular expression of Bcl-2 protein or Fas. This down-regulatory effect of interferon beta-1a on cellular expression of survivin was maintained after long-term therapy. CONCLUSIONS: Our observations suggest that interferon beta exerts a regulatory effect on peripheral T lymphocytes through an antiapoptosis mechanism that involves the down-regulation of cellular survivin expression. PMID- 12117360 TI - Vitamin E and cognitive decline in older persons. AB - BACKGROUND: Previous studies raise the possibility that antioxidants protect against neurodegenerative diseases. OBJECTIVE: To examine whether intake of antioxidant nutrients, including vitamin E, vitamin C, and carotene, is associated with reduced cognitive decline with age. DESIGN: Longitudinal population-based study conducted from September 17, 1993, to November 20, 2000, with an average follow-up of 3.2 years. PATIENTS: The patients were 2889 community residents, aged 65 to 102 years, who completed a food frequency questionnaire, on average 18 months after baseline. MAIN OUTCOME MEASURE: Cognitive change as measured by 4 tests (the East Boston Memory Test, which tests immediate and delayed recall; the Mini-Mental State Examination; and the Symbol Digit Modalities Test) at baseline and 3 years for all participants, and at 6 months for 288 randomly selected participants. RESULTS: We used random-effects models to estimate nutrient effects on individual change in the average score of the 4 cognitive tests. The cognitive score declined on average by 5.0 x 10(-2) standardized units per year. There was a 36% reduction in the rate of decline among persons in the highest quintile of total vitamin E intake (-4.3 x 10(-2) standardized units per year) compared with those in the lowest quintile (-6.7 x 10(-2) standardized units per year) (P =.05), in a model adjusted for age, race, sex, educational level, current smoking, alcohol consumption, total calorie (energy) intake, and total intakes of vitamin C, carotene, and vitamin A. We also observed a reduced decline with higher vitamin E intake from foods (P =.03 for trend). There was little evidence of association with vitamin C or carotene intake. CONCLUSION: Vitamin E intake, from foods or supplements, is associated with less cognitive decline with age. PMID- 12117361 TI - Down syndrome and Alzheimer disease: response to donepezil. AB - BACKGROUND: Individuals with Down syndrome who develop Alzheimer disease may show an improvement in cognitive functioning after treatment with acetylcholinesterase inhibitors. OBJECTIVE: To determine whether individuals with Down syndrome and Alzheimer disease will show improvement after institution of donepezil treatment. DESIGN: A nonrandomized controlled trial using donepezil in a pilot study format. SETTING: Academic medical center. PATIENTS: Convenience sample of 6 treated patients with Down syndrome and 9 closely matched historical control subjects. INTERVENTION: Oral administration of donepezil for a 5-month period. PRIMARY OUTCOME MEASURE: The Down Syndrome Dementia Scale. RESULTS: Significant improvement in dementia scores for the treated group during a 3- to 5-month period (P =.03). CONCLUSIONS: Acetylcholinesterase inhibitors may be helpful in reversing the symptoms of dementia during early and middle stages of cognitive decline. These findings support the rationale for a more extensive study of the efficacy of acetylcholinesterase inhibitors in Down syndrome dementia. PMID- 12117362 TI - A splice-site mutation in GABRG2 associated with childhood absence epilepsy and febrile convulsions. AB - CONTEXT: Missense mutations in the GABRG2 gene, which encodes the gamma 2 subunit of central nervous gamma-aminobutyric acid (GABA)(A) receptors, have recently been described in 2 families with idiopathic epilepsy. In one of these families, the affected individuals predominantly exhibited childhood absence epilepsy and febrile convulsions. OBJECTIVE: To assess the role of GABRG2 in the genetic predisposition to idiopathic absence epilepsies. DESIGN: The GABRG2 gene was screened by single-strand conformation analysis for mutations. Furthermore, a population-based association study assessing a common exon 5 polymorphism (C588T) was carried out. PATIENTS: The sample was composed of 135 patients with idiopathic absence epilepsy and 154 unrelated and ethnically matched controls. RESULTS: A point mutation (IVS6 + 2T-->G) leading to a splice-donor site mutation in intron 6 was found. The mutation, which is predicted to lead to a nonfunctional protein, cosegregates with the disease status in a family with childhood absence epilepsy and febrile convulsions. The association study did not find any significant differences in the allele and genotype frequencies of the common exon 5 polymorphism (C588T) between patients with idiopathic absence epilepsy and controls (P>.35). CONCLUSIONS: Our study identified a splice-donor site mutation that was probably causing a nonfunctional GABRG2 subunit. This mutation occurred in heterozygosity in the affected members of a single nuclear family, exhibiting a phenotypic spectrum of childhood absence epilepsy and febrile convulsions. The GABRG2 gene seems to confer a rare rather than a frequent major susceptibility effect to common idiopathic absence epilepsy syndromes. PMID- 12117363 TI - Interrelationship of genetics and prenatal injury in the genesis of malformations of cortical development. AB - CONTEXT: Although the causes of some malformations of cortical development (MCD) have been established, others remain unclear. There are several lines of evidence supporting the theory of a complex mechanism that involves genetic and environmental factors. OBJECTIVE: To investigate the interrelationship of genetics and prenatal injury in the genesis of MCD. PATIENTS AND DESIGN: A series of 76 consecutive patients with MCD and their families were systematically questioned about their family histories of epilepsy or other neurological impairment and the occurrence of prenatal events. Whenever possible, magnetic resonance imaging was performed in other family members if MCD was suspected or in the presence of any neurological impairment. Patients were divided into 3 groups according to the type of MCD. Patients in group 1 had focal cortical dysplasia, group 2 had heterotopias (periventricular or subcortical) or agyria pachygyria, and group 3 had polymicrogyria or schizencephaly. These findings were also compared with a disease-control group of 40 consecutive patients with epilepsy but without MCD. SETTING: Neurology clinic of a university hospital. RESULTS: Of the 76 patients with MCD, 21 (28%) had focal cortical dysplasia, 19 (25%) had heterotopias or agyria-pachygyria, and 36 (47%) had polymicrogyria or schizencephaly. There were 39 men and 37 women, aged 2 to 52 years (mean age, 13 years). In group 2, 6 patients (32%) had a family history of MCD, mental retardation, or miscarriages, suggesting a genetic predisposition. In group 3, family history of MCD was present in 5 patients (14%). Prenatal events occurred in 28 patients with MCD (37%) and 2 controls (5%) and were more frequent in patients with heterotopia or agyria-pachygyria and polymicrogyria (P<.001). Conversely, epilepsy occurred in all patients in group 1, in 17 patients (89%) in group 2, and in 17 patients (47%) in group 3. In group 3, epilepsy was less frequent (P<.001) and also more easily controlled (P =.005) than in other forms of MCD. CONCLUSIONS: Our findings support the idea of a spectrum among the different types of MCD. Focal cortical dysplasia (group 1) is associated with more frequent and severe epilepsy and less important genetic and prenatal events, heterotopias and agyria-pachygyria (group 2) are frequently associated with genetic predisposition, and polymicrogyria and schizencephaly (group 3) are less frequently associated with epilepsy but have a stronger association with genetic and detectable prenatal events. PMID- 12117364 TI - The apolipoprotein E epsilon 4 allele and decline in different cognitive systems during a 6-year period. AB - CONTEXT: Impairment of episodic memory is an early and defining feature of Alzheimer disease (AD). The apolipoprotein E (APOE) epsilon 4 allele is known to influence risk of AD but it has been difficult to establish whether it affects episodic memory differently from other cognitive functions. OBJECTIVE: To examine the association of epsilon 4 with decline in different cognitive systems. DESIGN: Longitudinal cohort study. SETTING: More than 40 groups of Catholic clergy from across the United States. PARTICIPANTS: Older Catholic clergy members without clinical evidence of dementia at baseline underwent annual clinical evaluations for up to 6 years. Of 624 persons eligible for follow-up, 611 (98%) participated, of whom 161 (26%) had at least 1 epsilon 4 allele. They completed an average of 5.5 evaluations (range, 2-7). MAIN OUTCOME MEASURES: Incident AD and annual rates of change in episodic memory, semantic memory, working memory, perceptual speed, and visuospatial ability. RESULTS: The presence of epsilon 4 was associated with risk of developing AD on follow-up (relative risk, 1.92; 95% confidence interval, 1.27-2.89). In a series of random effects models, epsilon 4 was associated with impaired baseline function in episodic memory and visuospatial ability and with more rapid decline in all domains. The effect of epsilon 4 on annual decline in episodic memory (>3-fold increase) was significantly stronger than its effect on decline in other cognitive systems (P<.01), and at baseline, its effect on episodic memory was marginally stronger than its effect on other cognitive domains (P =.06). CONCLUSION: The results suggest that the APOE epsilon 4 allele influences risk of AD by a relatively selective effect on episodic memory. PMID- 12117365 TI - Long-term risk of stroke and other vascular events in patients with asymptomatic carotid artery stenosis. AB - CONTEXT: The annual risk of ischemic stroke in patients with asymptomatic carotid artery stenosis is about 2% during the short-term (2-3 years), but the long-term risks of stroke and other vascular events are unknown, although they may affect surgical decision making. OBJECTIVE: To evaluate the long-term risk of stroke and other vascular events in patients with asymptomatic carotid artery stenosis. DESIGN: Cohort study with a median follow-up of 10 years (range, 5-18 years). SETTING: The teaching hospital of the University of Toronto, Toronto, Ontario. PATIENTS: From the initial cohort of 500 patients, 106 patients with asymptomatic carotid artery stenosis were selected because they had completed at least 5 years of follow-up. MAIN OUTCOME MEASURES: Ipsilateral stroke, myocardial infarction, and nonstroke vascular death. RESULTS: The 10- and 15-year actuarial risks of ipsilateral stroke were 5.7% (95% confidence interval [CI], 0%-12%) and 8.7% (95% CI, 1%-17%), respectively, in patients with 0% to 49% internal carotid artery stenosis, and 9.3% (95% CI, 1%-18%) and 16.6% (95% CI, 1%-32%) in patients with 50% to 99% internal carotid artery stenosis. The 10- and 15-year risks of myocardial infarction and nonstroke vascular death were 10.1% (95% CI, 4%-16%) and 24.0% (95% CI, 14%-34%). Age (P =.02), diabetes mellitus (P =.02), and internal carotid artery stenosis of 50% or more (P =.04) were predictive of increased risks of myocardial infarction and nonstroke vascular death. Internal carotid artery stenosis of 50% or more did predict the risk of ipsilateral stroke (P =.003) when all 181 asymptomatic carotid arteries were included. CONCLUSIONS: The annual stroke risk in patients with asymptomatic carotid artery stenosis was low and remained stable during long-term follow-up. Any benefit from carotid surgery is therefore unlikely to increase significantly with long-term follow-up. The high long-term risks of myocardial infarction and nonstroke vascular death suggest that prevention strategies should concentrate on coronary risk more than stroke risk. PMID- 12117366 TI - A functional magnetic resonance imaging study of left hemisphere language dominance in children. AB - BACKGROUND: Functional magnetic resonance imaging is a noninvasive method of assessing language dominance in a pediatric population. OBJECTIVE: To determine the pattern of receptive language lateralization in healthy children. DESIGN: We used functional magnetic resonance imaging to assess an auditory language task in 11 children (7 girls, 4 boys; mean age, 8.5 years). Participants alternately rested and listened to descriptors of nouns presented auditorily, naming the object described silently. Asymmetry indices ([(left - right)/(left + right)]) were calculated for a priori-determined regions of interest. RESULTS: The results showed strong activation bilaterally, with greater activation on the left in the superior and middle temporal gyri. Other areas of activation included the cuneus, the left inferior temporal gyrus, the prefrontal area, and the left fusiform and lingual gyri. Regions of interest analysis of individual scans showed additional activation in the left frontal lobe. Asymmetry indices showed strong left lateralization of the inferior frontal gyrus, middle frontal gyrus, and the Wernicke region. CONCLUSIONS: Hemispheric lateralization was clearly demonstrated in 8 children. As in adults, left hemisphere lateralization of receptive language is present at age 8 years. PMID- 12117367 TI - Atypical ganglion cell tumor of the sciatic nerve. AB - CONTEXT: Although herniation of a lumbosacral intervertebral disk is a major cause of sciatic distribution pain, relentlessly progressive symptoms or signs should alert one to the possibility of a tumor involving the nerve. OBJECTIVE: To describe the clinical, neurophysiological, and histological features of a pathologically unique tumor involving the sciatic nerve. SETTING: Tertiary referral university hospital. PATIENT: A 36-year-old woman was seen with a 6-year history of increasingly severe symptoms in the distribution of the left sciatic nerve. RESULTS: Electromyography indicated a sciatic nerve lesion in the region of the greater sciatic notch. Magnetic resonance imaging demonstrated a tumor involving the left sciatic nerve in this area. Light microscopy, electron microscopy, and immunohistochemistry results confirmed the presence of an atypical ganglion cell tumor of the sciatic nerve that exhibited prognostically conflicting clinical and histological features. CONCLUSIONS: To our knowledge, this is the first report of an atypical ganglion cell tumor affecting the sciatic nerve, and illustrates the value of detailed neurophysiological examination in localizing the site of peripheral nerve injury to facilitate focused neuroimaging when standard investigations are uninformative. Longer follow-up is required to determine the true biologic potential of this lesion. PMID- 12117368 TI - Enhanced magnetic resonance angiography of a spinal dural arteriovenous fistula. PMID- 12117369 TI - Brachial plexus. PMID- 12117371 TI - Standardizing frontotemporal dementia: the problem of polysemy. PMID- 12117373 TI - A piece of my mind. A survivor's way. PMID- 12117370 TI - Nervous system disorders: a global epidemic. AB - Major new epidemiological analyses are focusing attention on disorders of the nervous system as important causes of death and disability around the world. One in every 9 individuals dies of a disorder of the nervous system. Stroke outweighs all other neurological disorders combined as a cause of mortality. Most disorders of the nervous system occur in developing countries. Developmental disability due to malnutrition, and cognitive dysfunction associated with parasitic infections are the most common neurological disorders. As the world's population ages and the effects of infectious disease decline, the relative effects of many disorders of the nervous system, including stroke and dementia, are increasing. The disorders of the nervous system causing the highest rates of death and disability are preventable and treatable. Increased awareness of the global effects of neurological disorders should help health care planners and the neurological community set appropriate priorities in research, prevention, and management of these conditions. PMID- 12117377 TI - Improved care for neglected population must be "rule rather than exception". PMID- 12117378 TI - Can massive prevention efforts avert 29 million new cases of HIV by 2010? PMID- 12117379 TI - WHO declares the individual's right to be safe. PMID- 12117385 TI - Brain imaging to assess the effects of dopamine agonists on progression of Parkinson disease. PMID- 12117386 TI - Brain imaging to assess the effects of dopamine agonists on progression of Parkinson disease. PMID- 12117387 TI - Brain imaging to assess the effects of dopamine agonists on progression of Parkinson disease. PMID- 12117389 TI - Effects of leptin on platelet function in obese patients. PMID- 12117393 TI - Transfusion-related acute lung injury. PMID- 12117391 TI - Imaging vs biochemical testing for pheochromocytoma. PMID- 12117394 TI - Transfusion-related acute lung injury. PMID- 12117396 TI - Characteristics of health-related web sites identified by common internet portals. PMID- 12117398 TI - Menopausal hormone replacement therapy and risk of ovarian cancer. AB - CONTEXT: The association between menopausal hormone replacement therapy and ovarian cancer is unclear. OBJECTIVE: To determine whether hormone replacement therapy using estrogen only, estrogen-progestin only, or both estrogen only and estrogen-progestin increases ovarian cancer risk. DESIGN: A 1979-1998 cohort study of former participants in the Breast Cancer Detection Demonstration Project, a nationwide breast cancer screening program. SETTING: Twenty-nine US clinical centers. PARTICIPANTS: A total of 44 241 postmenopausal women (mean age at start of follow-up, 56.6 years). MAIN OUTCOME MEASURE: Incident ovarian cancer. RESULTS: We identified 329 women who developed ovarian cancer during follow-up. In time-dependent analyses adjusted for age, menopause type, and oral contraceptive use, ever use of estrogen only was significantly associated with ovarian cancer (rate ratio [RR], 1.6; 95% confidence interval [CI], 1.2-2.0). Increasing duration of estrogen-only use was significantly associated with ovarian cancer: RRs for 10 to 19 years and 20 or more years were 1.8 (95% CI, 1.1 3.0) and 3.2 (95% CI, 1.7-5.7), respectively (P value for trend <.001), and we observed a 7% (95% CI, 2%-13%) increase in RR per year of use. We observed significantly elevated RRs with increasing duration of estrogen-only use across all strata of other ovarian cancer risk factors, including women with hysterectomy. The RR for estrogen-progestin use after prior estrogen-only use was 1.5 (95% CI, 0.91-2.4), but the RR for estrogen-progestin-only use was 1.1 (95% CI, 0.64-1.7). The RRs for less than 2 years and 2 or more years of estrogen progestin-only use were 1.6 (95% CI, 0.78-3.3) and 0.80 (95% CI, 0.35-1.8), respectively, and there was no evidence of a duration response (P value for trend =.30). CONCLUSION: Women who used estrogen-only replacement therapy, particularly for 10 or more years, were at significantly increased risk of ovarian cancer in this study. Women who used short-term estrogen-progestin-only replacement therapy were not at increased risk, but risk associated with short-term and longer-term estrogen-progestin replacement therapy warrants further investigation. PMID- 12117397 TI - Risks and benefits of estrogen plus progestin in healthy postmenopausal women: principal results From the Women's Health Initiative randomized controlled trial. AB - CONTEXT: Despite decades of accumulated observational evidence, the balance of risks and benefits for hormone use in healthy postmenopausal women remains uncertain. OBJECTIVE: To assess the major health benefits and risks of the most commonly used combined hormone preparation in the United States. DESIGN: Estrogen plus progestin component of the Women's Health Initiative, a randomized controlled primary prevention trial (planned duration, 8.5 years) in which 16608 postmenopausal women aged 50-79 years with an intact uterus at baseline were recruited by 40 US clinical centers in 1993-1998. INTERVENTIONS: Participants received conjugated equine estrogens, 0.625 mg/d, plus medroxyprogesterone acetate, 2.5 mg/d, in 1 tablet (n = 8506) or placebo (n = 8102). MAIN OUTCOMES MEASURES: The primary outcome was coronary heart disease (CHD) (nonfatal myocardial infarction and CHD death), with invasive breast cancer as the primary adverse outcome. A global index summarizing the balance of risks and benefits included the 2 primary outcomes plus stroke, pulmonary embolism (PE), endometrial cancer, colorectal cancer, hip fracture, and death due to other causes. RESULTS: On May 31, 2002, after a mean of 5.2 years of follow-up, the data and safety monitoring board recommended stopping the trial of estrogen plus progestin vs placebo because the test statistic for invasive breast cancer exceeded the stopping boundary for this adverse effect and the global index statistic supported risks exceeding benefits. This report includes data on the major clinical outcomes through April 30, 2002. Estimated hazard ratios (HRs) (nominal 95% confidence intervals [CIs]) were as follows: CHD, 1.29 (1.02-1.63) with 286 cases; breast cancer, 1.26 (1.00-1.59) with 290 cases; stroke, 1.41 (1.07-1.85) with 212 cases; PE, 2.13 (1.39-3.25) with 101 cases; colorectal cancer, 0.63 (0.43-0.92) with 112 cases; endometrial cancer, 0.83 (0.47-1.47) with 47 cases; hip fracture, 0.66 (0.45-0.98) with 106 cases; and death due to other causes, 0.92 (0.74-1.14) with 331 cases. Corresponding HRs (nominal 95% CIs) for composite outcomes were 1.22 (1.09-1.36) for total cardiovascular disease (arterial and venous disease), 1.03 (0.90-1.17) for total cancer, 0.76 (0.69 0.85) for combined fractures, 0.98 (0.82-1.18) for total mortality, and 1.15 (1.03-1.28) for the global index. Absolute excess risks per 10 000 person-years attributable to estrogen plus progestin were 7 more CHD events, 8 more strokes, 8 more PEs, and 8 more invasive breast cancers, while absolute risk reductions per 10 000 person-years were 6 fewer colorectal cancers and 5 fewer hip fractures. The absolute excess risk of events included in the global index was 19 per 10 000 person-years. CONCLUSIONS: Overall health risks exceeded benefits from use of combined estrogen plus progestin for an average 5.2-year follow-up among healthy postmenopausal US women. All-cause mortality was not affected during the trial. The risk-benefit profile found in this trial is not consistent with the requirements for a viable intervention for primary prevention of chronic diseases, and the results indicate that this regimen should not be initiated or continued for primary prevention of CHD. PMID- 12117399 TI - Impact of a clinical decision rule on hospital triage of patients with suspected acute cardiac ischemia in the emergency department. AB - CONTEXT: Emergency department (ED) physicians often are uncertain about where in the hospital to triage patients with suspected acute cardiac ischemia. Many patients are triaged unnecessarily to intensive or intermediate cardiac care units. OBJECTIVE: To determine whether use of a clinical decision rule improves physicians' hospital triage decisions for patients with suspected acute cardiac ischemia. DESIGN AND SETTING: Prospective before-after impact analysis conducted at a large, urban, US public hospital. PARTICIPANTS: Consecutive patients admitted from the ED with suspected acute cardiac ischemia during 2 periods: preintervention group (n = 207 patients enrolled in March 1997) and intervention group (n = 1008 patients enrolled in August-November 1999). INTERVENTION: An adaptation of a previously validated clinical decision rule was adopted as the standard of care in the ED after a 3-month period of pilot testing and training. The rule predicts major cardiac complications within 72 hours after evaluation in the ED and stratifies patients' risk of major complications into 4 groups--high, moderate, low, and very low--according to electrocardiographic findings and presence or absence of 3 clinical predictors in the ED. MAIN OUTCOME MEASURES: Safety of physicians' triage decisions, defined as the proportion of patients with major cardiac complications who were admitted to inpatient cardiac care beds (coronary care unit or inpatient telemetry unit); efficiency of decisions, defined as the proportion of patients without major complications who were triaged to an ED observation unit or an unmonitored ward. RESULTS: By intention to-treat analysis, efficiency was higher in the intervention group (36%) than the preintervention group (21%) (difference, 15%; 95% confidence interval [CI], 8% 21%; P<.001). Safety was not significantly different (94% in the intervention group vs 89%; difference, 5%; 95% CI, -11% to 39%; P =.57). Subgroup analysis of intervention-group patients showed higher efficiency when physicians actually used the decision rule (38% vs 27%; difference, 11%; 95% CI, 3%-18%; P =.01). Improved efficiency was explained solely by different triage decisions for very low-risk patients. Most surveyed physicians (16/19 [84%]) believed that the decision rule improved patient care. CONCLUSIONS: Use of the clinical decision rule had a favorable impact on physicians' hospital triage decisions. Efficiency improved without compromising safety. PMID- 12117400 TI - Beta-blocker therapy and symptoms of depression, fatigue, and sexual dysfunction. AB - CONTEXT: beta-Blocker therapy remains substantially underused in cardiac patients despite its proven mortality benefits. Reluctance to prescribe these agents may derive from concerns about their association with symptoms of depression, fatigue, and sexual dysfunction. OBJECTIVE: To determine the association of beta blockers with depressive symptoms, fatigue, and sexual dysfunction by performing a quantitative review of randomized trials that tested beta-blockers in myocardial infarction, heart failure, and hypertension. DATA SOURCES: Randomized trials of beta-blockers used in the treatment of myocardial infarction, heart failure, or hypertension were identified by searching the MEDLINE database for English-language articles (1966-2001). In addition, we searched the reference lists of previously published trials and reviews of beta-blockers for additional studies. STUDY SELECTION: Criteria for inclusion of trials in the review were: random allocation of study treatments, placebo control, noncrossover design, enrollment of at least 100 patients, and a minimum of 6 months of follow-up. The initial search produced 475 articles, 42 of which met these criteria. Fifteen of these trials reported on depressive symptoms, fatigue, or sexual dysfunction and were selected for inclusion. DATA EXTRACTION: For each trial, 1 author abstracted the frequency of adverse events in the beta-blocker and placebo groups and the numbers of patients randomized to the treatment groups. Two other authors verified the counts of events, and all authors adjudicated any discrepancies. Two different types of information on adverse events were abstracted: patient reported symptoms and withdrawal of therapy due to a specified symptom. We categorized the tested beta-blockers by generation (early vs late) and lipid solubility (high vs low to moderate). DATA SYNTHESIS: The 15 trials involved more than 35,000 subjects. beta-Blocker therapy was not associated with a significant absolute annual increase in risk of reported depressive symptoms (6 per 1000 patients; 95% confidence interval [CI], -7 to 19). beta-Blockers were associated with a small significant annual increase in risk of reported fatigue (18 per 1000 patients; 95% CI, 5-30), equivalent to 1 additional report of fatigue for every 57 patients treated per year with beta-blockers. beta-Blockers were also associated with a small, significant annual increase in risk of reported sexual dysfunction (5 per 1000 patients; 95% CI, 2-8), equivalent to one additional report for every 199 patients treated per year. None of the risks of adverse effects differed significantly by degree of beta-blocker lipid solubility. The risk associated with reported fatigue was significantly higher for early generation than for late-generation beta-blockers (P =.04). CONCLUSION: The conventional wisdom that beta-blocker therapy is associated with substantial risks of depressive symptoms, fatigue, and sexual dysfunction is not supported by data from clinical trials. There is no significant increased risk of depressive symptoms and only small increased risks of fatigue and sexual dysfunction. The risks of these adverse effects should be put in the context of the documented benefits of these medications. PMID- 12117401 TI - The continuing unethical conduct of underpowered clinical trials. AB - Despite long-standing critiques of the conduct of underpowered clinical trials, the practice not only remains widespread, but also has garnered increasing support. Patients and healthy volunteers continue to participate in research that may be of limited clinical value, and authors recently have offered 2 related arguments to support the validity and value of underpowered clinical trials: that meta-analysis may "save" small studies by providing a means to combine the results with those of other similar studies to enable estimates of an intervention's efficacy, and that although small studies may not provide a good basis for testing hypotheses, they may provide valuable estimates of treatment effects using confidence intervals. In this article, we examine these arguments in light of the distinctive moral issues associated with the conduct of underpowered trials, the disclosures that are owed to potential participants in underpowered trials so they may make autonomous enrollment decisions, and the circumstances in which the prospects for future meta-analyses may justify individually underpowered trials. We conclude that underpowered trials are ethical in only 2 situations: small trials of interventions for rare diseases in which investigators document explicit plans for including their results with those of similar trials in a prospective meta-analysis, and early-phase trials in the development of drugs or devices, provided they are adequately powered for defined purposes other than randomized treatment comparisons. In both cases, investigators must inform prospective subjects that their participation may only indirectly contribute to future health care benefits. PMID- 12117402 TI - Informing clinical trial participants about study results. PMID- 12117403 TI - Failure of estrogen plus progestin therapy for prevention. PMID- 12117404 TI - Estrogen replacement therapy and risk of ovarian cancer. PMID- 12117405 TI - Commercial filming of patient care activities in hospitals. AB - Commercial filming of patient care activities is common in hospital settings. This article reviews common circumstances in which patients are commercially filmed, explores the potential positive and negative aspects of filming, and considers the ethical and legal issues associated with commercial filming of patients in hospital settings. We examine the competing goals of commercial filming and the duties of journalists vs the rights of patients to privacy. Current standards and recommendations for commercial filming of patient care activities are reviewed and additional recommendations are offered. PMID- 12117406 TI - Residents' prescription writing for nonpatients. AB - CONTEXT: Writing prescriptions is one of the most tangible new responsibilities that residents acquire after graduating from medical school. During their regular duties, house officers' prescription writing is carefully monitored. Little is known, however, about residents' patterns of prescription writing outside of supervision or about residents' knowledge of the ethical and legal guidelines that regulate prescription writing. OBJECTIVE: To study what factors influence residents' decision to write prescriptions for nonpatients. DESIGN, SETTING, AND PARTICIPANTS: Survey distributed in December 1997 to 92 internal medicine and family practice residents at a US community-based teaching hospital. Eighty percent responded. MAIN OUTCOME MEASURES: Self-reported prescribing activities for nonpatients and for individuals in 12 hypothetical vignettes. RESULTS: Eighty five percent of respondents reported having written prescriptions for nonpatients. Based on their responses to the vignettes, under certain circumstances, up to 95% of residents would write a prescription for an individual who is not their patient (eg, a sibling). Thirteen percent of residents believed that some ethical guidelines on prescription-writing activity existed. Only 4% of residents reported being aware of federal or state laws addressing the appropriateness of physician prescription writing for nonpatients. None of the residents were able to describe the circumstances that make prescription writing for nonpatients illegal or unethical based on legal statutes or ethical guidelines, respectively. CONCLUSIONS: In a sample of community-based internal medicine and family practice residents, unsupervised prescription writing by residents for individuals who are not their patients is a common occurrence. Since residency training is a time when practice habits are established, it is important that all residents learn about the ethical, legal, and liability implications of writing prescriptions for nonpatients. PMID- 12117412 TI - Urokinase upregulates matrix metalloproteinase-9 expression in THP-1 monocytes via gene transcription and protein synthesis. AB - Urokinase-type plasminogen activator (uPA) is suggested to exert its proliferatory, migratory and invasive action through binding with its membrane receptor, promoting pericellular proteolysis and mediating cell signal transduction. One of the possible actions of urokinase can be related to the regulation of activity and/or the expression of proteolytic enzymes participating in extracellular matrix degradation. In the present study, the role of uPA in regulating matrix metalloproteinase (MMP) expression and release by the monocyte cell line THP-1 was investigated. Recombinant uPA induced the release of MMP9/gelatinase B, as detected by zymography and Western blotting, and this release was abolished by actinomycin D and cycloheximide (inhibitors of DNA transcription and protein synthesis) and partially suppressed by monensin (an inhibitor of secretion). Proteolytically inactive urokinase with substitution of His(204) for Gln was able to reproduce about 70% of the effect induced by the wild-type recombinant uPA. The reverse transcription-PCR and Northern blot data indicated that the action of r-uPA on THP-1 cells resulted in formation of MMP9 mRNA, which depended on time, within 6-48 h, of the cell incubation with r-uPA. These results suggest that urokinase upregulates MMP9 expression in monocytes via MMP9 gene transcription and protein biosynthesis. PMID- 12117413 TI - A novel cycling assay for nicotinic acid-adenine dinucleotide phosphate with nanomolar sensitivity. AB - Nicotinic acid-adenine dinucleotide phosphate (NAADP) is a novel nucleotide derived from NADP that has now been shown to be active in releasing Ca(2+) from intracellular stores in a wide variety of cells ranging from plant to human. Despite the obvious importance of monitoring its cellular levels under various physiological conditions, no assay has been reported for NAADP to date. In the present study, a widely applicable assay for NAADP with high sensitivity is described. NAADP was first dephosphorylated to nicotinic acid-adenine dinucleotide by treatment with alkaline phosphatase. The conversion was shown to be stoichiometric. NMN-adenylyltransferase was then used to convert nicotinic acid-adenine dinucleotide into NAD in the presence of high concentrations of NMN. The resultant NAD was amplified by a cycling assay involving alcohol dehydrogenase and diaphorase. Each time NAD cycled through these coupled reactions, a molecule of highly fluorescent resorufin was generated. The reaction could be performed for hours, resulting in more than a 1000-fold amplification. Concentrations of NAADP over the 10-20 nM range could be routinely measured. This novel cycling assay was combined with an enzymic treatment to provide the necessary specificity for the assay. NAADP was found to be resistant to NADase and apyrase. Pretreatment of samples with a combination of the hydrolytic enzymes completely eliminated the interference from common nucleotides. The versatility of the cycling assay can also be extended to measure nicotinic acid, which is a substrate in the synthesis of NAADP catalysed by ADP-ribosyl cyclase, over the micromolar range. All the necessary reagents for the cycling assay are widely available and it can be performed using a multi-well fluorescence plate reader, providing a high-throughput method. This is the first assay reported for NAADP and nicotinic acid, which should be valuable in elucidating the messenger functions of NAADP. PMID- 12117414 TI - Differential effects of reduced glycoprotein VI levels on activation of murine platelets by glycoprotein VI ligands. AB - We have investigated the effects of decreased levels of the complex between glycoprotein VI (GPVI) and the Fc receptor gamma-chain (FcRgamma) on responses to collagen and GPVI-specific ligands in murine platelets. We show that levels of GPVI-FcRgamma of the order of 50% and 20% of wild-type levels caused 2- and 5 fold shifts to the right respectively in the dose-response curve for aggregation in response to collagen, the snake toxin convulxin and the monoclonal antibody JAQ1. In addition, there is a delay in the onset of aggregation in response to collagen. In contrast, the stimulation of protein tyrosine phosphorylation by collagen (as measured after 150 s) and adhesion to a collagen-coated surface under static conditions were unaffected in platelets with 50% and 20% of wild type levels of GPVI. In contrast, responses to a collagen-related peptide (CRP), made up of repeat glycine-proline-hydroxyproline motifs, were markedly inhibited and abolished in platelets expressing 50% and 20% of wild-type levels of GPVI respectively. We suggest that the marked effect of a reduction in GPVI levels on the CRP-induced activation of platelets is due to the multivalent nature of CRP and the fact that GPVI is its sole receptor on platelets. Thus it appears that the interaction of CRP with GPVI is determined by a combination of affinity and avidity. The observation that collagen does not behave like CRP in platelets expressing reduced levels of GPVI, even in the combined presence of blocking antibodies against integrin alpha2beta1 and GPV, suggests that collagen has a greater affinity than CRP for GPVI, and/or that other receptors are involved in its binding to platelets. The clinical significance of these results is discussed. PMID- 12117415 TI - Correlations between the functional integrity of the endoplasmic reticulum and polarized Ca2+ signalling in mouse lacrimal acinar cells: a role for inositol 1,3,4,5-tetrakisphosphate. AB - Ca(2+) signalling in exocrine acinar cells has been shown to be both polarized and pulsatile in all cell types tested, except acutely isolated mouse lacrimal acinar cells. Lacrimal cells are also unusual in that they display a very low sensitivity to Ins(1,4,5) P (3) (Ins P (3)) that may be enhanced by placing the cells in primary culture for 12-72 h or by intracellular infusion of a low concentration of Ins(1,3,4,5) P (4) (Ins P (4)). We have proposed previously that this atypical behaviour stemmed from vesiculation of the endoplasmic reticulum (ER) incurred during isolation of the cells and, furthermore, that time in culture or Ins P (4) increased sensitivity to Ins P (3) by increasing ER integrity [Smith, Harmer, Letcher and Irvine (2000) Biochem. J. 347, 77-82]. We have measured the half time for fluorescence recovery after photobleaching (FRAP) of a fluorescent marker (Mag-fluo 4) loaded into the ER lumen in order to determine directly the functional integrity of the ER in lacrimal cells. The half time for FRAP was increased (indicating a reduction in the functional integrity of the ER) following exposure to anti-microtubule agents (taxol and nocodazole) known to perturb ER structure and decreased (indicating an increase in the functional integrity of the ER) by time in culture and exposure to Ins P (4). The action of Ins P (4) was particularly pronounced because it occurred under patch clamp whole-cell conditions that were themselves found to reduce ER functional integrity. These data show that ER remodelling could be a physiological regulator of Ca(2+) signalling and indicate a role for Ins P (4) in control of this process. PMID- 12117417 TI - Transport of a neurotoxicant by molecular mimicry: the methylmercury-L-cysteine complex is a substrate for human L-type large neutral amino acid transporter (LAT) 1 and LAT2. AB - Methylmercury (MeHg) readily crosses cell membrane barriers to reach its target tissue, the brain. Although it is generally assumed that this rapid transport is due to simple diffusion, recent studies have demonstrated that MeHg is transported as a hydrophilic complex, and possibly as an L-cysteine complex on the ubiquitous L-type large neutral amino acid transporters (LATs). To test this hypothesis, studies were carried out in Xenopus laevis oocytes expressing two of the major L-type carriers in humans, LAT1-4F2 heavy chain (4F2hc) and LAT2-4F2hc. Oocytes expressing LAT1-4F2hc or LAT2-4F2hc demonstrated enhanced uptake of [(14)C]MeHg when administered as the L-cysteine or D,L-homocysteine complexes, but not when administered as the D-cysteine, N -acetyl-L-cysteine, penicillamine or GSH complexes. Kinetic analysis of transport indicated that the apparent affinities ( K (m)) of MeHg-L-cysteine uptake by LAT1 and LAT2 (98+/-8 and 64+/-8 microM respectively) were comparable with those for methionine (99+/-9 and 161+/ 11 microM), whereas the V (max) values were higher for MeHg-L-cysteine, indicating that it may be a better substrate than the endogenous amino acid. Uptake and efflux of [(3)H]methionine and [(14)C]MeHg-L-cysteine were trans stimulated by leucine and phenylalanine, but not by glutamate, indicating that MeHg-L-cysteine is both a cis - and trans -substrate. In addition, [(3)H]methionine efflux was trans -stimulated by leucine and phenylalanine even in the presence of an inwardly directed methionine gradient, demonstrating concentrative transport by both LAT1 and LAT2. The present results describe a major molecular mechanism by which MeHg is transported across cell membranes and indicate that metal complexes may form a novel class of substrates for amino acid carriers. These transport proteins may therefore participate in metal ion homoeostasis and toxicity. PMID- 12117416 TI - Control of eukaryotic protein synthesis by upstream open reading frames in the 5' untranslated region of an mRNA. AB - Control of gene expression is achieved at various levels. Translational control becomes crucial in the absence of transcription, such as occurs in early developmental stages. One of the initiating events in translation is that the 40 S subunit of the ribosome binds the mRNA at the 5'-cap structure and scans the 5' untranslated region (5'-UTR) for AUG initiation codons. AUG codons upstream of the main open reading frame can induce formation of a translation-competent ribosome that may translate and (i) terminate and re-initiate, (ii) terminate and leave the mRNA, resulting in down-regulation of translation of the main open reading frame, or (iii) synthesize an N-terminally extended protein. In the present review we discuss how upstream AUGs can control the expression of the main open reading frame, and a comparison is made with other elements in the 5' UTR that control mRNA translation, such as hairpins and internal ribosome entry sites. Recent data indicate the flexibility of controlling translation initiation, and how the mode of ribosome entry on the mRNA as well as the elements in the 5'-UTR can accurately regulate the amount of protein synthesized from a specific mRNA. PMID- 12117419 TI - Predicting residents' performance: a prospective study. AB - BACKGROUND: Objective criteria for predicting residents' performance do not exist. The purpose of this study was to test the hypothesis that global assessment by an intern selection committee (ISC) would correlate with the future performance of residents. METHODS: A prospective study of 277 residents between 1992 and 1999. Global assessment at the time of interview was compared to subsequent clinical (assessed by chief residents) and cognitive performance (assessed by the American Board of Pediatrics in-service training examination). RESULTS: ISC ratings correlated significantly with clinical performance at 24 and 36 months of training (r = 0.58, P <.001; and r = 0.60, P <.001 respectively). ISC ratings also correlated significantly with in-service exam scores in the 1st, 2nd, and 3rd years of training (r = 0.35, P =.0016; r = 0.39, P = 0.0003; r = 0.50, P = 0.005 respectively). CONCLUSIONS: Global assessment by an ISC predicted residents' clinical and cognitive performances. PMID- 12117418 TI - Structural and functional characterization of tissue factor pathway inhibitor following degradation by matrix metalloproteinase-8. AB - Vascular injury results in the activation of coagulation and the release of proteolytic enzymes from neutrophils and connective- tissue cells. High concentrations of these inflammatory proteinases may destroy blood coagulation proteins, contributing to coagulation and bleeding disorders associated with severe inflammation. Matrix metalloproteinase-8 (MMP-8) is released from neutrophils at sites of inflammation and vascular disease. We have investigated the effect of MMP-8 degradation on the anticoagulant function of tissue factor pathway inhibitor (TFPI) as a potential pathological mechanism contributing to coagulation disorders. MMP-8 cleaves TFPI following Ser(174) within the connecting region between the second and third Kunitz domains ( k (cat)/ K (m) approximately 75 M(-1).s(-1)) as well as following Lys(20) within the NH(2) terminal region. MMP-8 cleavage of TFPI decreases the anticoagulant activity of TFPI in factor Xa initiated clotting assays as well as the ability of TFPI to inhibit factor Xa in amidolytic assays. Yet, MMP-8 cleavage does not alter the ability of TFPI to inhibit trypsin. Since the inhibition of both factor Xa and trypsin is mediated by binding to the second Kunitz domain, these results suggest that regions of TFPI other than the second Kunitz domain may directly interact with factor Xa. (125)I-factor Xa ligand blots of TFPI fragments generated following MMP-8 degradation were used for probing binding interactions between factor Xa and regions of TFPI, other than the second Kunitz domain. In experiments performed under reducing conditions that disrupt the Kunitz domain structure, (125)I-factor Xa binds to the C-terminal fragment of MMP-8-degraded TFPI. This fragment contains portions of TFPI distal to Ser(174), which include the third Kunitz domain and the basic C-terminal region. An altered form of TFPI lacking the third Kunitz domain, but containing the C-terminal region, was used to demonstrate that the C-terminal region directly interacts with factor Xa. PMID- 12117420 TI - Separate urinary bladder and prostate neurons in the central nervous system of the rat: simultaneous labeling with two immunohistochemically distinguishable pseudorabies viruses. AB - BACKGROUND: This work examines the central nervous system distribution of virus labeled neurons from the rat urinary bladder and the prostate simultaneously within the same tissue sections. Two immunohistochemically distinct pseudorabies virus strains were simultaneously injected into male Sprague Dawley rats (approximately 280 gm). One virus was injected into the bladder and the other into the prostate. After incubation intervals of 2.25, 2.5, 2.75, 3 and 4 days, sections from the spinal cord and brain were processed immunohistochemically to detect cells, within a single section, which were labeled separately by each virus or were labeled by both viruses. RESULTS: Each strain of virus labeled a separate population of neurons and some neurons were labeled by both strains. The majority of neurons labeled by virus from the urinary bladder were found in the L6-S1 spinal cord segments within the dorsal gray commissure, the intermediolateral area and the superficial dorsal horn. Neurons labeled by virus from the prostate were mainly found in the L1-L2 spinal cord segments in the dorsal gray commissure and the intermediolateral areas. Double-labeled interneurons in L1-L2 were mainly located in the intermediolateral area. In L6-S1 they were divided between the dorsal gray commissure and the intermediolateral area. CONCLUSIONS: Spinal neurons innervating the bladder are clearly separate and different from those innervating the prostate. This difference also persists in the brain. In disagreement with previous reports, no direct anatomical evidence of parasympathetic innervation of the prostate was observed. PMID- 12117422 TI - Parenteral iron therapy in obstetrics: 8 years experience with iron-sucrose complex. AB - Fe is an essential component of haem in myoglobin and accounts for 70 % of haemoglobin. The balance of Fe, unlike that of other metals such as Na or Ca, is regulated solely by gastrointestinal absorption, which itself depends on the bioavailability of Fe in food, i.e. the chemical Fe species. Factors that maintain Fe homeostasis by modulating Fe transfer through the intestinal mucosa are found at the luminal, mucosal and systemic levels. Fe deficiency and its consequence, Fe-deficiency anaemia, form the commonest nutritional pathology in pregnant women. The current gold standard to detect Fe deficiency remains the serum ferritin value. Previously there was general consensus against parenteral Fe administration, i.e. parenteral Fe was only recommended for special conditions such as unresponsiveness to oral Fe, intolerance to oral Fe, severe anaemia, lack of time for therapy etc. However, especially in hospital settings, clinicians regularly face these conditions but are still worried about reactions that were described using Fe preparations such as Fe-dextrans. A widely used and safe alternative is the Fe-sucrose complex, which has become of major interest to prevent functional Fe deficiency after use of recombinant erythropoietin Numerous reports show the effectiveness and safety of the Fe-sucrose complex. Good tolerance to this Fe formulation is partly due to the low allergenic effect of the sucrose complex, partly due to slow release of elementary Fe from the complex. Accumulation of Fe-sucrose in parenchyma of organs is low compared with Fe-dextrans or Fe-gluconate, while incorporation into the bone marrow for erythropoiesis is considerably faster. Oral Fe is only started if haemoglobin levels are below 110 g/l. If levels fall below 100 g/l or are below 100 g/l at time of diagnosis, parenteral Fe-sucrose is used primarily. In cases of severe anaemia (haemoglobin <90 g/l) or non-response to parenteral Fe after 2 weeks, recombinant erythropoietin is considered in combination. By using parenteral Fe sucrose in cases of severe Fe deficiency, anaemia during pregnancy is treated efficiently and safely according to our results and rate of blood transfusion could be reduced considerably to below 1 % of patients per year. PMID- 12117423 TI - Dietary n-3 and n-6 fatty acids alter avian metabolism: metabolism and abdominal fat deposition. AB - The effects of dietary saturated fatty acids and polyunsaturated fatty acids (PUFA) of the n-3 and n-6 series on weight gain, body composition and substrate oxidation were investigated in broiler chickens. At 3 weeks of age three groups of chickens (n 30; ten birds per group) were fed the fat-enriched experimental diets for 5 weeks. These diets were isonitrogenous, isoenergetic and contained 208 g protein/kg and 80 g edible tallow, fish oil or sunflower oil/kg; the dietary fatty acid profiles were thus dominated by saturated fatty acids, n-3 PUFA or n-6 PUFA respectively. Resting RQ was measured in five birds from each treatment group during weeks 4 and 5 of the experiment. There were no significant differences between treatments in total feed intake or final body mass. Birds fed the PUFA diets had lower RQ and significantly reduced abdominal fat pad weights (P<0.01) compared with those fed tallow. The dietary lipid profile changes resulted in significantly greater partitioning of energy into lean tissue than into fat tissue (calculated as breast lean tissue weight:abdominal fat mass) in the PUFA groups compared with the saturated fat group (P<0.01; with no difference between the n-3 and n-6 PUFA groups). In addition, the PUFA-rich diets lowered plasma concentrations of serum triacylglycerols and cholesterol. The findings indicate that dietary fatty acid profile influences nutrient partitioning in broiler chickens. PMID- 12117424 TI - Dietary n-3 and n-6 fatty acids alter avian metabolism: molecular-species composition of breast-muscle phospholipids. AB - The effects of diets high in n-3 polyunsaturated fatty acids (PUFA; provided by fish oil), n-6 PUFA (sunflower oil) or in more-saturated fatty acids (tallow) on the distribution of subclasses of choline phospholipids (PC) and ethanolamine phospholipids (PE) from the breast muscle of broiler chickens were examined. Supplementation with the different fatty acids had no effect on the distribution of phospholipid subclasses. Feeding sunflower oil or tallow gave a molecular species profile similar in both fatty acid subtype and proportion. In the diacyl PC phospholipids, 16 : 0-18 : 1n-9 and 16 : 0-18 : 2n-6 accounted for approximately 60 % of the total molecular species, whereas for the alkylenyl PC the predominant species were 16 : 0-18 : 1n-9 and 16 : 0-20 : 4n-6. Of the diacyl PE the dominant species was 18 : 0-20 : 4n-6 which accounted for 50 % of the molecular species, and of the alkylenyl PE the dominant species were 16 : 0-18 : 1n-9, 16 : 0-20 : 4n-6 and 18 : 0-20 : 4n-6. Supplementation with fish oil significantly increased levels of both eicosapentaenoic acid (20 : 5n-3) and docosahexaenoic acid (22 : 6n-3) in PC and PE when compared with either sunflower oil or tallow supplementation. The increase in the n-3 PUFA incorporation was associated with a corresponding decrease in the proportion of arachidonic acid (20 : 4n-6) in both PC and PE. Different dietary fats induce different patterns of fatty acid incorporation and substitution in the sn-2 position of the diacyl and alkylenyl PC and PE of avian breast muscle, and this finding is indicative of selective acyl remodelling in these two phospholipids. PMID- 12117425 TI - Variation in fat, lactose and protein in human milk over 24 h and throughout the first year of lactation. AB - Fat in human milk is extremely variable and can represent up to 50 % of infant energy intake. To accurately determine milk composition and infant intake at 1 (n 17), 2 (n 17), 4 (n 17), 6 (n 15), 9 (n 6) and 12 (n 5) months of lactation, samples of fore- and hind-milk were collected from each breast at each feed over 24 h periods from an initial group of seventeen women. The content of fat in milk varied over 24 h, with a mean CV of 47.6 (se 2.1) % (n 76) and 46.7 (se 1.7) % (n 76) for left and right breasts respectively. The 24 h amounts of fat, lactose and protein in milk differed between women (P=0.0001), but were consistent between left and right breasts. Daily milk production differed between breasts (P=0.0001) and women (P=0.0001). Accordingly, amounts of fat (P=0.0008), lactose (P=0.0385) and protein (P=0.0173) delivered to the infant over 24 h also differed between breasts and women (P=0.0001). The energy content of milk and the amount of energy delivered to the infant over 24 h were the same between breasts, but differed between women (P=0.0001). The growth rate of a group of only six infants in the present study was not related to either the concentrations or amounts of fat, lactose, protein and energy in milk over the first 6 months of life. These results show the individuality of milk composition and suggest that only a rigorous sampling routine that takes into account all levels of variation will allow the accurate determination of infant intake of fat, lactose, protein and energy. PMID- 12117426 TI - The effect of dietary protein content and feeding level on the rate of protein deposition and energy utilization in growing Iberian pigs from 15 to 50 kg body weight. AB - The effects of dietary protein content and feeding level on the utilization of metabolizable energy (ME) and on the rates of gain, protein and fat deposition have been studied in seventy-two Iberian pigs growing from 15 to 50 kg body weight (BW) by means of comparative slaughter experiments. The animals were fed on six diets providing 223, 192, 175, 156, 129 and 101 g crude ideal protein ( CP)/kg DM and 14.64, 14.14, 14.37, 14.80, 15.36 and 15.53 MJ ME/kg DM respectively. Each diet was offered at three levels of feeding: 0.60, 0.80 and 0.95xad libitum intake. Protein deposition (PD) increased significantly (P<0.01) with each decrease in dietary CP content and reached a maximum value (74.0 g) when the diet providing 129 g CP/kg DM (6.86 g digestible ideal protein/MJ ME) was offered at the highest feeding level. This feeding regimen resulted in average values for live-weight gain and retained energy (RE) of 559 g/d and 10.9 MJ/d respectively. RE increased significantly (P<0.001) from 480 to 626 kJ/kg BW0.75 with each decrease in dietary CP content from 192 to 129 g/kg DM. Raising the level of feed intake led to significant linear increases in PD and RE irrespective of the diet fed (P<0.001). When diets approaching an adequate supply of CP were given, the net efficiency of use of ME for growth (kw) and the maintenance energy requirements were 58.2 % and 422 kJ/kg BW0.75 per d respectively. PMID- 12117427 TI - Divalent metal inhibition of non-haem iron uptake across the rat duodenal brush border membrane. AB - Duodenal Fe2+ uptake is essential to body Fe2+ homeostasis, but the interaction of metals with the uptake process remains unclear. The present study compared the effects of four essential trace metals (Mn2+, Zn2+, Co2+ and Ni2+) with two toxic metals (Pb2+ and Cd2+) on Fe2+ uptake across the brush border membrane of villus attached duodenal enterocytes. Everted rat duodenum was exposed to buffer containing 0.2 mm-59Fe2+-ascorbate with or without the competing metal (2 mm) and the tissue was then processed for autoradiography allowing Fe2+ uptake to be determined at specific crypt-villus regions. The quantification method ensured that uptake by cells, rather than Fe2+ binding to the tissue surface, was measured. Fe2+ uptake was significantly inhibited by Cd2+ in upper villus enterocytes only and Pb2+ was without effect on Fe2+ uptake. The inhibition by Cd2+ was not due to general cell damage as judged by the release of lactate dehydrogenase from tissue into incubation fluid. Essential divalent trace metals reduced uptake significantly along the whole length of the crypt-villus axis. Cd2+ uptake, measured separately, took place at all regions of the villus-crypt axis, highest uptake being into crypt enterocytes. The very different uptake profiles for Cd2+ and Fe2+ suggests that the divalent metal transporter 1 is not the principal transporter of Cd2+. The addition of Fe2+ to incubation buffer inhibited Cd2+ uptake by both crypt and villus enterocytes. The possibility that the inhibitory actions of Fe2+ and Cd2+ on the uptakes of Cd2+ and Fe2+ respectively can be explained by a non-competitive action or the involvement of an additional metal transporter is discussed. PMID- 12117428 TI - The intake of fried virgin olive or sunflower oils differentially induces oxidative stress in rat liver microsomes. AB - The effects of non-fried and fried virgin olive and sunflower oils on rat liver microsomal compositional features have been investigated. In addition, plasma antioxidants (alpha-tocopherol and ubiquinone 9) were investigated as well as the possible oxidative modifications suffered by virgin olive and sunflower oils during the frying process. The frying process decreased the content of alpha tocopherol and phenolics in the oils and increased total polar materials. Sunflower oil was affected to a greater extent than olive oil. In rats, the intake of fried oil led to higher levels of lipid peroxidation and a lower concentration of plasma antioxidants. Microsomal fatty acid and antioxidant profiles were also altered. It seems that a strong relationship exists between the loss of antioxidants and the production of toxic compounds in the oils after frying and the extent of the peroxidative events in microsomes, which were also different depending on the fat source. The highly unsaturated sunflower oil was less resistant to the oxidative stress produced by frying and led to a higher degree of lipid peroxidation in liver microsomes in vivo than virgin olive oil. PMID- 12117429 TI - Differences in iron requirements by concanavalin A-treated and anti-CD3-treated murine splenic lymphocytes. AB - Fe availability is critical for optimal lymphocyte proliferation; however, the minimum required levels are unknown. Such information is valuable when assessing in vitro immune responses in Fe-deficient subjects, because serum (Fe) added to the culture medium may replete lymphocytes. To address this issue, splenic lymphocytes obtained from seventeen 3-month-old C57BL/6 mice were incubated without and with 1 mg/l concanavalin A or 50 microg/l anti-CD3 antibody in media that contained between 0.113 and 9.74 micromol Fe/l. Fe was provided by either fetal calf serum (FCS, 0-100 ml/l), newborn calf serum (NBCS, 0-100 ml/l), or NBCS (10 ml/l) plus ferric ammonium citrate. As expected, the rate of DNA synthesis increased with Fe levels (P<0.01). Maximum DNA synthesis was obtained with 2.26 micromol Fe/l (50 ml FCS/l) for concanavalin A and 0.895 micromol/l (20 ml FCS/l) for anti-CD3-treated cells. In serum-free media (0.113 micromol Fe/l), the proliferative responses to concanavalin A were below the background, while they rose 5.5-fold in anti-CD3-treated cells (P<0.05). In apotransferrin supplemented media (0.13 micromol Fe/l), the proliferative responses to concanavalin A and anti-CD3 antibody were 18.6 and 71 %, respectively, of that obtained with 4.66 micromol Fe/l (100 ml FCS/l). Interleukin 2 secretion also followed the same trend as lymphocyte proliferation. Since differences between both mitogens persisted after FCS was substituted with NBCS, we can rule out an effect on ribonucleotide reductase activity, or by other serum growth factors. We speculate an Fe effect at an early step of T-cell activation. Data suggest that the minimum Fe concentration required for lymphocyte proliferation varies with the mitogen. PMID- 12117430 TI - Influence of the pattern of peptide supply on microbial activity in the rumen simulating fermenter (RUSITEC). AB - The source and pattern of N supply was varied in the rumen simulation technique (RUSITEC) in order to determine if continuous, rather than transient, availability of peptides was required for optimum ruminal fermentation. The energy source was fibre prepared from sugar-beet pulp. N was added as NH3 continuously infused (AC) or peptides (Bacto(R) Casitone, a pancreatic hydrolysate of casein; Difco Laboratories, Detroit, MI, USA) continuously infused (PC) or added as a single dose at the time of feeding (PS). Free peptides were detected in the fermenter liquid for 4 h after feeding in the AC treatment, for 10 h in the PS treatment, and at all times with the PC treatment. Treatments had no effect on DM degradation. Approximately 40 % of the degradation occurred during the time no peptides were detected in the PS treatment. Microbial N flow tended to be higher with the peptide additions (P<0.061), with no significant difference between the two peptides treatments. The production of liquid associated micro-organisms (LAM) was higher in the PC treatment (P<0.05) and the proportion of LAM derived from NH3 lower (P<0.05). However, LAM only accounted for 20-30 % total microbial population. Our main conclusion was that peptides had a small stimulatory effect on the fermentation, but there was no indication that synchrony of supply of energy and amino acid-N in the fermenter promoted a more efficient fermentation than non-synchronous supply. This conclusion must be qualified, however, because some N remained in the fibre and may have become available progressively as the fibre was digested by the micro-organisms. PMID- 12117431 TI - The effect of consumption of foods that differ in energy density and/or sodium bicarbonate supplementation on subsequent diet selection in sheep. AB - The short-term consumption of foods that differed in energy density (ED) and/or NaHCO3 supplementation, on subsequent food intake and diet selection in sheep were measured. Thirty sheep weighing 35.9 (sd 2.89) kg were used. Two foods were formulated: H had 11 and L had 8 MJ metabolizable energy/kg fresh matter. Four further foods were formulated by adding either 40 g NaHCO3/kg or 16.5 g NaCl/kg to foods H and L. NaCl was added to give the same Na concentration as with 40 g NaHCO3/kg to control for any effects of Na per se. In a preliminary test, it was found that a 2 h consumption of food H supplemented with NaHCO3 could buffer potential impact on the rumen environment of subsequent consumption of food H alone (as judged by rumen pH and acid-buffering capacity); however, it was not as effective as the consumption of food L alone in doing so. Each food treatment was offered to one of six groups (n 5) for 2 h following 16 h of food deprivation. Sheep were then offered a choice between H and L for a further 6 h. Supplementing H or L with either NaHCO3 or NaCl had no significant effect on either intake or diet selection. ED significantly (P<0.01) affected intake during the 2 h single feeding period, with sheep offered H or L consuming 540 and 663 (sed 37) g respectively, but had no effect on subsequent intake during the choice period. During the choice period all sheep showed a preference for food H, but sheep previously offered L selected significantly more H (0.873 g/g) than sheep previously offered H (0.544 (sed 0.028) g/g; P<0.001). It is concluded that short term consumption of foods that differ in ED, and hence in their potential impact on the rumen environment, significantly affects subsequent diet selection. This is in agreement with the hypothesis that ruminant animals select a diet to help maintain the rumen environment within a certain physiological range. Food H with 40 g NaHCO3 added/kg may not have been sufficient to affect subsequent diet selection. It is suggested that larger, rather than smaller, changes in the rumen environment achieved through previous feeding should be expected to alter subsequent diet selection. PMID- 12117432 TI - Comparison of dietary and non-dietary risk factors in overweight and normal weight Chinese adults. AB - The aim of the present study was to compare the differences in dietary and non dietary factors contributing to normal weight and overweight among urban Chinese adults. Two cross-sectional population surveys were carried out in Tianjin, one of the largest cities in China. A total of 2631 subjects aged 25-64 years were selected by random stratified cluster sampling; 398 men and 490 women were overweight, and 886 men and 857 women were of normal weight. The diet was assessed by food weighing plus consecutive individual 3 d food records. Health related behaviours and anthropometry were assessed. The overweight group had significantly higher mean daily intakes of energy and carbohydrate than the normal-weight group. Overweight men also had significantly higher mean daily intakes of protein and fat than normal-weight men. Age, daily intakes of energy and carbohydrate, and marital status were positively associated with overweight, while occupational and commuting physical activity, as well as smoking, were inversely associated with overweight among both genders. Daily intakes of protein, fat and alcohol were positively related to the incidence of being overweight among men. People with 7-12 years education were more likely to be overweight compared with those with less than 6 years of education. High intakes of energy and carbohydrate among both genders, as well as high intakes of protein and fat among men, and lower levels of occupational and commuting physical activity, being a non-smoker, and partly higher socio-economic status were related to a greater incidence of being overweight in this population. PMID- 12117433 TI - UK Food Standards Agency cis-monounsaturated fatty acid workshop report. AB - The UK Food Standards Agency convened a group of expert scientists to review current research investigating the optimal dietary intake for n-9 cis monounsaturated fatty acids (MUFA). The aim was to review the mechanisms underlying the reported beneficial effects of MUFA on CHD risk, and to establish priorities for future research. The issue of optimal MUFA intake is contingent upon optimal total fat intake; however, there is no consensus of opinion on what the optimal total fat intake should be. Thus, it was recommended that a large multi-centre study should look at the effects on CHD risk of MUFA replacement of saturated fatty acids in relation to varying total fat intakes; this study should be of sufficient size to take account of genetic variation, sex, physical activity and stage of life factors, as well as being of sufficient duration to account for adaptation to diets. Recommendations for studies investigating the mechanistic effects of MUFA were also made. Methods of manipulating the food chain to increase MUFA at the expense of saturated fatty acids were also discussed. PMID- 12117435 TI - Laboratory analyses for poisoned patients. AB - Guidelines on the appropriate availability and use of laboratory analysis have been produced by the National Poisons Information Service and the Association of Clinical Biochemists and are published in this issue. These are intended to support the consistent and high quality investigation of the poisoned patient. PMID- 12117436 TI - Laboratory analyses for poisoned patients: joint position paper. AB - To enable consistency of investigation and the establishment of best practice standards, consensus guidelines have been formulated jointly by the UK National Poisons Information Service (NPIS) and the Association of Clinical Biochemists (ACB). The types of laboratory investigation required for poisoned patients were categorized as either (a) essential common laboratory investigations or (b) specific toxicological assays, and also as either (i) common or (ii) specialist or infrequent. Tests in categories (a) and (bi) are expected to be available 24 h per day, with a maximum turnaround time of 2 h. For the specialist assays, i.e. category (bii), availability and turnaround times have been specified individually. The basis for selection of these times has been clinical utility. The adoption of these guidelines, along with the use of the NPIS online poisons information resource TOXBASE (www.spib.axl.co.uk), will enable the poisoned patient to receive appropriate, 'best practice' investigations according to their clinical needs and will avoid the use of unnecessary investigations. PMID- 12117437 TI - Quarts out of pint pots -- expanding the pint. Clinical budgeting in practice. AB - Clinical budgeting is the process whereby clinical users are charged for the resources they use. A system for recharging users for the costs of tests was introduced at the Northern General Hospital, Sheffield, in 1995, and has been in operation since. The system has allowed pathology to maintain budgetary balance, has automatically compensated for workload increases, has allowed the introduction of new tests, and has encouraged clinical users to include pathology costs in their bids for funding for clinical developments. The system works according to rules agreed between pathology and its users at the outset, but once set up takes a minimal amount of work to operate and maintain. PMID- 12117438 TI - Using Medline to achieve an evidence-based approach to diagnostic clinical biochemistry. AB - Medline is the US National Library of Medicine database that is used for searching the medical biochemistry literature. The database is structured using medical subject subheadings (MeSH terms) to classify the content of references; indexing is done manually using MeSH terms as key words. Searching the database effectively means finding the maximum number of relevant references together with the minimum number of irrelevant ones. This article is aimed at explaining the limitations of Medline and suggesting some solutions to key problems. The goal is that users can improve their literature search technique by employing a structured approach. As usual, asking relevant questions before starting a search is essential. PMID- 12117439 TI - Screening for colorectal cancer. AB - Colorectal cancer (CRC) causes 20 000 deaths per annum in the UK alone. Screening has been shown to reduce mortality but debate exists as to which approach to use. Direct visualization of the colorectum has the advantage that it detects lesions most effectively and is required at less frequent intervals, but the procedure is invasive and at present too costly for screening purposes. Faecal occult blood measurement, despite its limitations, is currently the recommended screening method, with follow-up of positive tests by colonoscopy or other visualization techniques. This strategy has been shown to reduce mortality from CRC by about 20% and screening trials directed towards individuals in the over 50 years age group are underway in the UK and elsewhere. Future developments in CRC screening include colorectal visualization by computed colonography -- a less-invasive alternative to colonoscopy. Developments in stool analysis are also occurring. Examination of faecal samples for cellular products derived from neoplasms (e.g. calprotectin) may prove more sensitive and specific than faecal occult blood measurements. In addition, detection of altered DNA in faeces is being investigated by molecular biology techniques. Using a multi-target assay panel to detect point mutations and other neoplasia-associated DNA abnormalities may be an effective strategy for CRC screening in the future. PMID- 12117440 TI - Interference in immunoassay is an underestimated problem. AB - The presence of antibodies in some patients' serum has long been known to be a potential source of interference in immunoassays, as shown by numerous case reports. These often appear after the introduction of a new analyte (e.g. troponin) and then decrease in number as the topic becomes exhausted. This highlights the persistent and intrinsic nature of this problem, despite attempts by the manufacturers to compensate for this source of error. However, an explanation of the immunoanalytical basis underpinning these assays could be more effective in raising awareness than intermittent case reports. In this review we have outlined the use of antibodies as reagents, the factors determining how they bind to antigen(s), and the nature of the immune response in order to explain the insidious and unpredictable nature of this form of interference. Studies on the prevalence of interference have yielded values ranging from 0.05 to more than 2%. However, these figures are analyte- and assay-specific, influenced by the study design, and are not therefore generally applicable. It is also highly likely that figures on prevalence and incidence will worsen in the future because of the wider use of monoclonal antibodies as diagnostic and therapeutic tools. Clinical laboratories should be alert to assay interference from antibodies irrespective of its nature, as immunoassays will remain an indispensable analytical tool, unlikely to be replaced in the foreseeable future by a practical alternative. PMID- 12117441 TI - Are there any clinical indications for measuring IgG subclasses? AB - It is questionable as to whether a low serum concentration of one of the IgG subclasses identifies a disease state. A low IgG(1) concentration is found in primary or secondary immunodeficiency states but does not occur in isolation. Low IgG(2) concentration is associated with an increased risk of bacterial infections but only in some individuals and not in others. Isolated IgG(3) and IgG(4) deficiency have not been convincingly demonstrated. Therefore, the isolated finding of low concentrations of one or more IgG subclass does not identify individuals at risk. In contrast, the finding of low serum concentrations of antibodies to specific bacterial antigens (Haemophilus influenzae type B, pneumococcus, tetanus and diphtheria) does identify individuals at risk and these measurements should be used in preference to IgG subclass measurement. PMID- 12117442 TI - Analysis of carbon monoxide. AB - The degree of exposure to carbon monoxide is most often assessed by measuring the blood carboxyhaemoglobin saturation. This measurement is relevant to investigations of acute accidental or deliberate poisoning and of chronic exposure in a domestic or work place environment. Simple spectrophotometric methods based on differential protein precipitation or dithionite reduction are prone to interference from other haemoglobin pigments and are imprecise for low level estimations. Automated spectrophotometric devices (CO-oximeters) that estimate simultaneously total haemoglobin, percentage oxyhaemoglobin and percentage carboxyhaemoglobin have acceptable accuracy for carboxyhaemoglobin saturation levels of > 5% and are recommended for most clinical purposes. For the investigation of low-level exposure and the detection of increased haemolysis in neonates, more sensitive methods involving the release of carbon monoxide and its measurement by gas chromatography are required. Gas chromatographic methods are also appropriate when examining post-mortem blood samples where putrefaction or heat stress has resulted in a significant change in haemoglobin composition. PMID- 12117443 TI - Biochemical detection of minor myocardial injury after elective, uncomplicated, successful percutaneous coronary intervention in patients with stable angina: clinical outcome. AB - BACKGROUND: Minor elevations of creatine kinase MB isoform (CK-MB) identified a population with a worse long-term prognosis after successful coronary intervention. Recent studies provide evidence that cardiac troponin I (cTnI) is more sensitive than CK-MB for the detection of minor myocardial injury after coronary intervention. The purpose of the study was to determine the prognostic value of cTnI elevation after elective uncomplicated successful percutaneous coronary intervention (PCI). METHODS: cTnI was measured in 96 patients with stable angina before and 24 h after elective uncomplicated successful percutaneous transluminal coronary angioplasty (PTCA) with or without stenting. Patients were followed up for adverse cardiac events (recurrent angina, non-fatal myocardial infarction, cardiac death, repeat PCI or coronary bypass surgery). Procedure success was achieved in all cases. RESULTS: Cardiac events were best predicted by cTnI when a cut-off value of 2.0 microg/L was used. Abnormal cTnI values at 24 h after PCI were observed in 26 patients (27%). Over a follow-up period of 24 months with no significant difference in the medication used, the incidence of recurrent angina, repeat PCI, coronary bypass surgery and cardiac death was 54%, 46%, 4% and 4%, respectively, in the cTnI-positive patients versus 27%, 16%, 4% and 0% in the cTnI-negative patients. Kaplan-Meier survival analysis showed that cTnI elevation was a significant correlate of cardiac events (P = 0.0198, by log rank analysis). CONCLUSIONS: Elevation of cTnI is not uncommon after elective uncomplicated successful PCI in patients with stable angina and this elevation might be a marker of adverse long-term outcome. PMID- 12117444 TI - Limitations of Pneumovax as a detection antigen in the measurement of serotype specific antibodies by enzyme-linked immunosorbent assay. AB - BACKGROUND: Measurement of antibody responses to polysaccharide antigens is regarded as an important assessment of an individual's ability to respond to carbohydrate antigens. The currently used assays for the measurement of pneumococcal-specific antibody use the multi-serotype vaccine Pneumovax as the detection antigen. METHODS: An equal potency enzyme-linked immunosorbent assay (ELISA) system was used to compare the measurement of serotype-specific antibody with the multi-serotype assay. RESULTS: Our results show that the concentration of specific antibody to Pneumovax is not related to the concentration of antibody to the individual serotypes. Neither is any correlation found between the antibody concentrations to any of the three single serotypes investigated, to the mixture of the three serotypes or to Pneumovax. CONCLUSION: We conclude that the measurement of the concentration of the specific antibody to the mixed serotypes present in Pneumovax has serious limitations when used to evaluate the protection acquired from Pneumovax immunization against any specific serotype. PMID- 12117445 TI - Evaluation of a sandwich enzyme-linked immunosorbent assay for the measurement of serum heart fatty acid-binding protein. AB - BACKGROUND: We evaluated the sandwich enzyme-linked immunosorbent assay (ELISA) MARKIT-M for the determination of heart fatty-acid-binding protein (H-FABP). RESULTS AND CONCLUSIONS: The between-run coefficient of variation of this assay was <3.9 and it showed good correlation with a previously established ELISA method. The upper reference limit in 30 healthy individuals was 6.1 microg/L. Admission serum H-FABP was evaluated against myoglobin in 41 patients with suspected myocardial infarction (onset of symptoms < or = 5 h). H-FABP showed the same diagnostic efficiency as myoglobin [area (standard error) under the receiver operating characteristic curve: 0.798 (0.079) for H-FABP, 0.771 (0.085) for myoglobin, P = 0.55]. However, using the upper reference limit as decision cut off, the sensitivity for H-FABP [91%; 95% confidence interval (CI): 76-98%] was significantly (P = 0.019) higher than that of myoglobin (65%; 95% CI: 47-80%). PMID- 12117446 TI - Free cortisol index as a surrogate marker for serum free cortisol. AB - BACKGROUND: The biologically active component of a hormone is the unbound or free fraction. Changes in cortisol-binding protein could give misleading results if only total cortisol is measured for the interpretation of dynamic function tests. METHODS: This study aimed to measure serum free cortisol using a steady-state gel filtration method and then to evaluate the correlation between the serum free cortisol and the free cortisol index (FCI), defined as serum total cortisol/cortisol-binding globulin (CBG). RESULTS: Forty-eight serum samples from healthy volunteers undergoing a short Synacthen test were analysed for total cortisol, free cortisol and CBG. The FCI correlated well with a previously established, but more complex, calculation of serum free cortisol (R = 0.98, P <0.001) and with measured serum free cortisol (R = 0.90, P < 0.001). CONCLUSION: Free cortisol index is a reliable and user-friendly measure of serum free cortisol. PMID- 12117447 TI - Severe hypertriglyceridaemia associated with human immunodeficiency virus and highly active antiretroviral therapy. AB - We report two cases of severe hypertriglyceridaemia associated with human immunodeficiency virus infection and highly active antiretroviral therapy (HAART). The first patient, a 39-year-old man, developed moderate hypertriglyceridaemia (5.88 mmol/L) and hypercholesterolaemia (7.0 mmol/L) after 8 months of HAART. When his therapy was altered, triglyceride and cholesterol concentrations increased further to 15.9 and 10.9 mmol/L, respectively, after 6 weeks. The second patient, a 31-year-old man, presented with triglyceride and cholesterol concentrations of 16.2 and 5.7 mmol/L, respectively, following an 8 year history of HAART. Therapy was changed, but 1 month later the triglyceride concentration had increased to 39.4 mmol/L and the cholesterol concentration to 12.1 mmol/L. Both patients were managed by a change in HAART and the introduction of a fibric acid derivative. Although neither patient displayed any clinical symptoms associated with hypertriglyceridaemia, it is important to recognize such cases because of the associated risk of pancreatitis and coronary disease. PMID- 12117449 TI - Survival after very high blood alcohol concentrations. PMID- 12117452 TI - Cyclo-oxygenase-2-specific inhibitors and cardiovascular morbidity. PMID- 12117453 TI - Putting LIFE into hypertension. PMID- 12117454 TI - Is fixed combination therapy appropriate for initial hypertension treatment? AB - Recent clinical trials in hypertension prove how seldom single drug therapy achieves target blood pressure (BP) and reduces cardiovascular morbidity and mortality. A natural response is the testing and marketing of fixed-dose combination products for hypertension, of which 14 have been approved in the United States since 1993. Currently, only five products are indicated by the Food and Drug Administration for initial therapy of hypertension; all include a diuretic. To achieve such an indication, studies must show not only safety and efficacy of the combination, but also BP lowering that is at least additive compared with the two agents given separately, as well as a "synergy" not present when each agent is given alone. Some advantages to initial combination therapy include greater BP reduction, improved adherence to pill taking, fewer side effects, and lower cost. The most likely candidates for initial combination therapy are patients with initial BP higher than 160/100 mm Hg, or those with a BP goal lower than the customary 140/90 mm Hg. These include patients with target organ damage, clinical cardiovascular disease, proteinuria, renal impairment, or diabetes mellitus. In many of these circumstances, an angiotensin converting enzyme inhibitor or angiotensin II receptor antagonist is frequently recommended; adding a diuretic or calcium antagonist to it is much more likely to result in achievement of the BP goal. More research is being done to explore the combination of not only two representatives from classes of conventional agents, but also other drugs that may help address the multiple manifestations of the "metabolic syndrome" that often accompanies hypertension. PMID- 12117455 TI - Are angiotensin converting enzyme inhibitors and angiotensin receptor blockers becoming the treatment of choice in African-Americans? AB - African-American patients constitute a significant and important group who are at high risk for developing hypertension-related complications. The proportion of African-American patients succumbing to or suffering from cardiovascular, renal, and neurologic sequelae is unacceptably high. Therefore, it is extremely crucial to develop appropriate therapeutic strategies for this vital subset of our society. The renin-angiotensin system may play a role in the pathophysiology of hypertension-related diseases, and therefore drugs that block this system, ie, angiotensin converting enzyme inhibitors and angiotensin receptor blockers, may have a special indication for African-American patients. Although these drugs may not be the most efficacious agents in terms of blood pressure reduction, they have a major benefit in offering target organ protection and arresting disease progression in the African-Americans. Hence, contrary to the old notions, drugs blocking the renin-angiotension system have an important place in the management of hypertension and related disorders in African-American patients. PMID- 12117456 TI - Are clinical endpoint benefits of angiotensin converting enzyme inhibitors independent of their blood pressure effects? AB - Both basic and experimental data indicate that the renin-angiotensin system through angiotensin II mediates its classic hemodynamic role, but also has a significant deleterious role in a number of cardiac, vascular, and renal disorders. Indeed, evidence indicates that angiotensin II negatively impacts endothelial function, cardiac remodeling, vessel wall hypertrophy, atherosclerosis, and progressive renal disease. Newer data point to a significant role for angiotensin II in inflammation and in inducing plasminogen activator inhibitor. This widespread negative effect can be countered by newer antihypertensive drugs, angiotensin converting enzyme inhibitors, and angiotensin receptor blockers. Both small and large clinical trials suggest a large benefit of such drugs on not only organ-specific endpoints such as renal disease or proteinuria, but on global cardiovascular events. It does appear that when blood pressure is significantly elevated, lowering blood pressure does indeed provide protection for larger endpoints such as stroke. However, at lower blood pressure levels, a hemodynamically independent effect is likely to be contributing to the positive effects. We should embrace these effects and champion them for our patients. PMID- 12117457 TI - Fixed low-dose combination therapy for hypertension. AB - Despite the availability of various classes of antihypertensive agents that lower blood pressure by different primary actions, the treatment of hypertension remains a difficult task. Essential hypertension has a highly heterogeneous character, so that monotherapies are often not sufficient to normalize blood pressure. This is especially true since the goal of treatment is currently to normalize both systolic and diastolic blood pressure. By combining medications acting by different mechanisms, it is possible to gain considerably in terms of antihypertensive efficacy because of synergistic impacts on the cardiovascular system. Furthermore, low doses of antihypertensive agents are generally sufficient when used in combination, which accounts for the excellent tolerability of combination products. Fixed low-dose combinations are very useful tools for treating hypertensive patients. Because of their simplicity of use, and the fact that they improve the blood pressure response rate while minimizing the incidence of adverse effects, such combinations are increasingly being considered as suitable for both second-line and first-line therapy. PMID- 12117458 TI - Contemporary treatment of heart failure: is there adequate evidence to support a unique strategy for African-Americans? Pro position. AB - Clinical trials provide average effects of interventions but are not powered to identify differences in subgroup responses. African-Americans have been under represented in most previous trials in patients with heart failure. Furthermore, physiologic differences have been demonstrated between African-Americans and whites in the mechanisms and response to therapy in hypertension. Review of previous heart failure trials reveals that African-Americans exhibit less benefit than whites from ACE inhibitors, angiotensin receptor blockers, and at least one b-blocker. In contrast, black patients experienced a greater benefit than white patients from the combination of nitrate and hydralazine. These data emphasize the need for trials in black patients to identify effective therapy. The first such trial, African-American Heart Failure Trial, is currently underway. PMID- 12117459 TI - Contemporary treatment of heart failure: is there adequate evidence to support a unique strategy for African-Americans? Con position. AB - Heart failure is a substantial cause of increased morbidity and mortality in the African-American population, with poorer prognosis versus white patients. Systolic heart failure is predominantly caused by poorly controlled hypertension in African-Americans. Overall, African-Americans remain underrepresented in morbidity and mortality heart failure trials, and further data are needed to confirm the potential benefit of present therapies and newer approaches to heart failure in African-Americans. Intensive blood pressure control and control of other risk factors, along with the appropriate application of evidence-based therapies including angiotensin converting enzyme (ACE) inhibitors and approved beta-blockers, are required to decrease racial disparities. Although some data suggest that contemporary treatment with ACE inhibitors and beta-blockers may be less effective in African-Americans in terms of reducing heart failure morbidity and mortality, there is not adequate evidence to support a unique strategy for this population. The use of evidence-based therapies should be equally applied to African-Americans as well as to other ethnic groups while awaiting further studies. PMID- 12117460 TI - Cardiovascular morbidity and mortality in patients with diabetes in the Losartan Intervention For Endpoint reduction in hypertension study (LIFE): a randomised trial against atenolol. PMID- 12117461 TI - Renal protection by antihypertensive therapy. AB - The attainment of adequate renal protection requires strict blood pressure control and a diminution of proteinuria or microalbuminuria to values as near from normalcy as possible. It has been considered that by getting the first, the second could be attained at the same price. Recent data have confirmed that renal protection in hypertensive patients, diabetics or not, requires combination therapy that has to include an angiotensin converting enzyme inhibitor or an angiotensin receptor blocker. A calcium channel blocker can be added to this without renal compromise. A diuretic will also be needed in most cases. Proteinuria will diminish with this combination in particular if up-titration of the drug blocking the effects of angiotensin II is performed. The control of other associated risk factors is also required, in particular smoking and lipids. PMID- 12117462 TI - Low sodium diet after DASH: has the situation changed? Dietary Approaches to Stop Hypertension. AB - The Dietary Approaches to Stop Hypertension (DASH) trial adds to the large body of evidence indicating a direct association of dietary sodium with blood pressure, and showing that rigorous reduction of dietary sodium can reduce blood pressure over a 30-day period by statistically significant amounts. DASH, however, neither addressed nor answered whether a reduction of dietary sodium reduces morbidity and mortality. Data linking baseline sodium to mortality and morbidity outcomes are sparse, with only six known studies. Of these, two showed no association, two showed an inverse association, and two showed a direct association only in obese subsets. No studies have examined outcomes after sodium reduction, and no studies have linked sodium to outcomes or even a blood pressure benefit among treated hypertensives. Universal recommendations for sodium reduction or dietary sodium goals should await evidence that such interventions are both safe and effective as measured by morbidity and mortality outcomes. PMID- 12117463 TI - Toxicity of hexavalent chromium to Daphnia magna: influence of reduction reaction by ferrous iron. AB - The reaction kinetics of hexavalent chromium with ferrous ions were studied to determine the influence of reduction on the toxicity of chromium to aquatic organisms. The changes in chemical forms of the chromate in the presence of ferrous ions were examined in a bioassay system using Daphnia magna as a test organism. This study demonstrated that the reaction kinetics of chromate with ferrous ions showed a significant decrease of chromate concentration with the second-order rate coefficient (k) for the reduction of Cr(VI) being determined as 55.2M(-1)s(-1). The concentration of Cr(VI) remaining in the solution decreased as the ratio of ferrous ion to chromate increased, revealing a non-stoichiometric reaction due to oxygenation and the moderately alkaline pH of the solutions. The toxicity test indicated that the bioavailability of chromate to D. magna was reduced in the presence of Fe(II) and that it decreased further with increasing Fe(II) concentrations. However, the toxic effect of chromate to aquatic organisms was not controlled kinetically in the presence of ferrous ions. It was also found that LC(50) of chromate to D. magna decreased about 1.5-fold as the test period increased from 24 to 48h in the presence of Fe(II). PMID- 12117464 TI - Stabilization of phosphogypsum using class C fly ash and lime: assessment of the potential for marine applications. AB - Phosphogypsum (PG, CaSO(4).H(2)O), a solid byproduct of phosphoric acid manufacturing, contains low levels of radium ((266)Ra), resulting in stackpiling as the only currently allowable disposal/storage method. PG can be stabilized with class C fly ash and lime for potential use in marine environments. An augmented simplex centroid design with pseudo-components was used to select 10 PG:class C fly ash:lime compositions. The 43cm(3) blocks were fabricated and subjected to a field submergence test and 28 days saltwater dynamic leaching study. The dynamic leaching study yielded effective calcium diffusion coefficients (D(e)) ranging from 1.15 x 10(-13) to 3.14 x 10(-13)m(2)s(-1) and effective diffusion depths (X(c)) ranging from 14.7 to 4.3mm for 30 years life. The control composites exhibited diametrical expansions ranging from 2.3 to 17.1%, providing evidence of the extent of the rupture development due to ettringite formation. Scanning electron microscopy (SEM), microprobe analysis showed that the formation of a CaCO(3) on the composite surface could not protect the composites from saltwater intrusion because the ruptures developed throughout the composites were too great. When the PG:class C fly ash:lime composites were submerged, saltwater was able to intrude throughout the entire composite and dissolve the PG. The dissolution of the PG increased the concentration of sulfate ions that could react with calcium aluminum oxides in class C fly ash forming additional ettringite that accelerated rupture development. Effective diffusion coefficients and effective diffusion depths alone are not necessarily good indicators of the long-term survivability of PG:class C fly ash:lime composites. Development of the ruptures in the composites must be considered when the composites are used for aquatic applications. PMID- 12117465 TI - Performance study of cementitious systems containing zeolite and silica fume: effects of four metal nitrates on the setting time, strength and leaching characteristics. AB - The aim of this study is to investigate the effect of four metal nitrate contaminants, namely chromium, manganese, lead and zinc on the mechanical and leaching characteristics of cement-based materials. For this purpose, three different matrices made of: (i) Portland cement, (ii) Portland cement and silica fume, and (iii) Portland cement and natural zeolite were studied. The effects of metals on the stabilised/solidified (S/S) product characteristics were monitored by measuring: (i) setting time, (ii) compressive strength, (iii) acid neutralisation capacity (ANC), and (iv) solubility of the metal contaminants as a function of pH. The results of both mechanical and leaching tests showed the importance of the contaminant/matrix couple considered. Setting time was accelerated in presence of chromium, while in presence of manganese, lead and zinc it was delayed. However, for the last two contaminants, a 10% replacement of cement by silica fume and zeolite, markedly accelerated the setting time compared to the cement-only matrix. Although the early strength development was adversely affected in presence of all four contaminants, the long-term strength was less affected compared to the control materials. Although the ANC of the materials was not markedly affected by the presence of contaminants, the nature of the matrix did modify the ANC behaviour of the solidified materials. The increased strength and reduced ANC observed in the presence of silica fume are both due to pozzolanic reaction. The type of matrix used for solidification did not affect the solubility of the four metal contaminants. Overall, the results showed that the use of blended cements must be carried out with care and the performance assessment of waste-containing cement-based materials must take into consideration both the mechanical and leaching characteristics of the systems. PMID- 12117466 TI - Characteristics of slag produced from incinerated hospital waste. AB - Ash produced from a hospital waste incinerator was treated using a high temperature melting process at 1200 degrees C. The quality of the produced slag was characterized by X-ray diffraction (XRD), X-ray fluorescence (XRF), leaching tests and sequential chemical extraction of metals. The slag contained large amounts of SiO(2,) CaO, Al(2)O(3), Sn, Ni, Cu, Ba and B. XRD analysis revealed a moderate crystal structure for the melted slag and identified the main crystals as quartz (SiO(2)), kaolinite (Al(2)Si(2)O(5)(OH)(4)), albite (NaAlSi(3)O(8)) and gibbsite (Al(OH)(3)). The observed crystal structure assists in preventing the leaching of heavy metals from the slag. Furthermore, the leaching results found the produced slag to comply with disposal limits set by the US EPA. Results from sequential chemical extraction analysis showed that metals in the slag exhibited the strongest preference to be bound to the residual fraction (stable fraction), which is known to have very low leaching characteristics. Melting was found to stabilize heavy metals in hospital waste successfully and therefore it can be an acceptable method for disposal. PMID- 12117467 TI - Low-molecular-weight carboxylic acids produced from hydrothermal treatment of organic wastes. AB - This article reports production of low-molecular-weight carboxylic acids from the hydrothermal treatment of representative organic wastes and compounds (i.e. domestic sludge, proteinaceous, cellulosic and plastic wastes) with or without oxidant (H(2)O(2)). Organic acids such as acetic, formic, propionic, succinic and lactic acids were obtained in significant amounts. At 623 K (16.5 MPa), acetic acid of about 26 mg/g dry waste fish entrails was obtained. This increased to 42 mg/g dry waste fish entrails in the presence of H(2)O(2). Experiments on glucose to represent cellulosic wastes were also carried out, getting acetic acid of about 29 mg/g glucose. The study was extended to terephthalic acid and glyceraldehyde, reaction intermediates of hydrothermal treatment of polyethylene terephthalate (PET) plastic wastes and glucose, respectively. In addition, production of lactic acid, one of the interesting low-molecular-weight carboxylic acids, was discussed on the viewpoint of resources recovery. Studies on temperature dependence of formation of organic acids showed thermal stability of acetic acid, whereas, formic acid decomposed readily under hydrothermal conditions. In general, results demonstrated that the presence of oxidants favored formation of organic acids with acetic acid being the major product. PMID- 12117468 TI - Chloride influence on the incorporation of Cr(2)O(3) and NiO in clinker: a laboratory evaluation. AB - Co-incinerating wastes in cement rotary kilns may introduce elements not usually found in the raw material into the process or even increase the quantity of some minor elements. Chromium and nickel are present in some electroplating sludges. These wastes are hazardous and must be treated and disposed of in a controlled landfill. The objective of this work was to study the incorporation of chromium and nickel oxides when they were added to clinker raw meal. The clinker raw meal used in this study was prepared by mixing pure compounds in the ratio that made it possible to obtain the same phases of an industrial Portland clinker at the end of the heating process. Twelve samples were prepared by adding 0.05, 0.1, 0.3, 0.5, 0.8 and 1.0wt.% of Cr(2)O(3) and 0.05, 0.1, 0.3, 0.5, 0.8 and 1.0wt.% of NiO to the base charge. Also, four more samples were prepared with additions of 0.6% Cr(2)O(3) and 0.1% NaCl, 0.7% Cr(2)O(3) and 0.1% KCl, 0.8% NiO and 0.1% NaCl and 0.8% NiO and 0.1% KCl. In order to study the Cr and Ni incorporation, thermogravimetric (TG) tests were performed, as well as tests in an assembled experimental device. The products were characterized by X-ray fluorescence chemical analysis, microprobe analyses by energy disperse scanning (EDS) and leaching tests. The results showed that the chromium and nickel added were incorporated into the clinker, even when chlorides were present. PMID- 12117469 TI - Equilibrium and kinetic modeling of adsorption of reactive dye on cross-linked chitosan beads. AB - The adsorption of reactive dye (Reactive Red 189) from aqueous solutions on cross linked chitosan beads was studied in a batch system. The equilibrium isotherms at different particle sizes (2.3-2.5, 2.5-2.7 and 3.5-3.8mm) and the kinetics of adsorption with respect to the initial dye concentration (4320, 5760 and 7286 g/m(3)), temperature (30, 40 and 50 degrees C), pH (1.0, 3.0, 6.0 and 9.0), and cross-linking ratio (cross-linking agent/chitosan weight ratio: 0.2, 0.5, 0.7 and 1.0) were investigated. Langmuir and Freundlich adsorption models were applied to describe the experimental isotherms and isotherm constants. Equilibrium data fitted very well to the Langmuir model in the entire saturation concentration range (0-1800 g/m(3)). The maximum monolayer adsorption capacities obtained from the Langmuir model are very large, which are 1936, 1686 and 1642 g/kg for small, mediumand large particle sizes, respectively, at pH 3.0, 30 degrees C, and the cross-linking ratio of 0.2. The pseudo first- and second-order kinetic models were used to describe the kinetic data, and the rate constants were evaluated. The experimental data fitted well to the second-order kinetic model, which indicates that the chemical sorption is the rate-limiting step, instead of mass transfer. The initial dye concentration and the solution pH both significantly affect the adsorption capacity, but the temperature and the cross-linking ratio are relatively minor factors. An increase in initial dye concentration results in the increase of adsorption capacity, which also increases with decreasing pH. The activation energy is 43.0 kJ/mol for the adsorption of the dye on the cross linked chitosan beads at pH 3.0 and initial dye concentration 3768 g/m(3). PMID- 12117476 TI - An expanding view for the molecular basis of familial periodic paralysis. AB - The periodic paralyses are rare disorders of skeletal muscle characterized by episodic attacks of weakness due to intermittent failure of electrical excitability. Familial forms of periodic paralysis are all caused by mutations in genes coding for voltage-gated ion channels. New discoveries in the past 2 years have broadened our views on the diversity of phenotypes produced by mutations of a single channel gene and have led to the identification of potassium channel mutations, in addition to those previously found in sodium and calcium channels. This review focuses on the clinical features, molecular genetic defects, and pathophysiologic mechanisms that underlie familial periodic paralysis. PMID- 12117477 TI - Welander distal myopathy outside the Swedish population: phenotype and genotype. AB - Welander distal myopathy is a late onset disorder that is mainly seen in Sweden. It is linked to chromosome 2p13 and all Swedish patients show a common shared haplotype, indicating a founder mutation. Here we report the clinical manifestations, magnetic resonance imaging, pathophysiology and haplotype analysis of Welander patients in the Finnish population. The clinical examination of patients from 12 different families showed a distal myopathy with onset in the long extensor muscles of the hands and fingers, also seen in Swedish Welander patients. Muscle biopsies showed characteristic myopathic changes. Haplotype analysis with the five polymorphic markers that make up the common core haplotype, seen in the Swedish patients, revealed that this haplotype is also co segregating in the Finnish patients and a common ancestry is therefore further supported for patients with Welander distal myopathy. PMID- 12117478 TI - Congenital myasthenic syndrome associated with episodic apnea and sudden infant death. AB - The sudden infant death syndrome has multiple etiologies. Some congenital myasthenic syndromes can cause sudden infant death syndrome by apnea, but the frequency of this etiology is unknown. We report here a young patient with sudden respiratory crises culminating in apnea followed by recovery, against a background of no or variable myasthenic symptoms without dyspnea. One sib without myasthenic symptoms and one sib who only had mild ptosis died previously during febrile episodes. Studies reported by us elsewhere traced the proband's illness to mutations in choline acetyltransferase. Here, we describe in detail the morphologic investigations and electrophysiologic findings, which point to a presynaptic defect in acetylcholine resynthesis or vesicular filling, in the proband. Analysis of DNA from a sib who previously died of sudden infant death syndrome revealed the same choline acetyltransferase mutation. Thus, mutations in choline acetyltransferase may be a cause of sudden infant death syndrome as, theoretically, could other presynaptic myasthenic disorders. PMID- 12117479 TI - Facioscapulohumeral (FSHD1) and other forms of muscular dystrophy in the same family: is there more in muscular dystrophy than meets the eye? AB - We report on two unrelated Brazilian families with members affected by two different forms of muscular dystrophy. In the first one, the 35-year-old male proband has limb-girdle muscular dystrophy with proximal weakness, elevated creatine kinase and a myopathic muscle biopsy. All the proteins known to be associated with limb-girdle muscular dystrophy were normal. Two of his sisters also complained of muscle weakness. The oldest sister showed clinical signs consistent with facioscapulohumeral muscular dystrophy, confirmed through molecular analysis. She presented a 30 kb EcoRI/BlnI fragment which was found in another six relatives, but surprisingly not in the affected proband or the other sister. In the second family, a 57-year-old male with a typical facioscapulohumeral muscular dystrophy phenotype has a 17 kb EcoRI/BlnI fragment, which was also present in other affected relatives. However in a 14-year-old severely affected male cousin, confined to a wheelchair since age 12, but without facial weakness, the small fragment was absent. These families illustrate the importance of testing all affected individuals in a family. PMID- 12117480 TI - A G468-T AMPD1 mutant allele contributes to the high incidence of myoadenylate deaminase deficiency in the Caucasian population. AB - Myoadenylate deaminase deficiency is the most common metabolic disorder of skeletal muscle in the Caucasian population, affecting approximately 2% of all individuals. Although most deficient subjects are asymptomatic, some suffer from exercise-induced myalgia suggesting a causal relationship between a lack of enzyme activity and muscle function. In addition, carriers of this derangement in purine nucleotide catabolism may have an adaptive advantage related to clinical outcome in heart disease. The molecular basis of myoadenylate deaminase deficiency in Caucasians has been attributed to a single mutant allele characterized by double C to T transitions at nucleotides +34 and +143 in mRNA encoded by the AMPD1 gene. Polymerase chain reaction-based strategies have been developed to specifically identify this common mutant allele and are considered highly sensitive. Consequently, some laboratories preferentially use this technique over other available diagnostic tests for myoadenylate deaminase deficiency. We previously identified a G468-T mutation in one symptomatic patient who was only heterozygous for the common AMPD1 mutant allele. In this report, nine additional individuals with this compound heterozygous genotype are revealed in a survey of 48 patients with documented deficiency of skeletal muscle adenosine monophosphate deaminase and exercise-induced myalgia. Western blot analysis of leftover biopsy material from one of these individuals does not detect any immunoreactive myoadenylate deaminase polypeptide. Baculoviral expression of the G468-T mutant allele produces a Q156H substitution enzyme exhibiting labile catalytic activity. These combined results demonstrate that the G468-T transversion is dysfunctional and further indicate that AMPD1 alleles harboring this mutation contribute to the high incidence of partial and complete myoadenylate deaminase deficiency in the Caucasian population. Consequently, genetic tests for abnormal AMPD1 expression designed to diagnose patients with metabolic myopathy, and to evaluate genetic markers for clinical outcome in heart disease should not be based solely on the detection of a single mutant allele. PMID- 12117481 TI - Multifocal motor neuropathy and asymptomatic Hashimoto's thyroiditis: first report of an association. AB - Motor neuropathy with multifocal conduction blocks represents a recently identified autoimmune disorder of the peripheral nerve myelin. Association of motor neuropathies or neuronopathies with thyroid disorders, such as hyperthyroidism, hypothyroidism or thyroid neoplasms has been rarely described. We studied a 61-year-old man with a 2-year-history of slowly progressive weakness of the left limbs with atrophy and fasciculations. Nerve conduction velocity studies revealed multifocal motor conduction blocks. Serum IgM titer of antibodies against GM1 was elevated (1:1280; n.v. up to 1:640). Thyroid studies were compatible with Hashimoto's thyroiditis. Therapy with high dose intravenous immunoglobulins was followed by a prompt clinical recovery. Then the disease assumed an intravenous immunoglobulins dependent course with a full clinical, but transient, recovery. This is the first observation of an association of multifocal motor neuropathy with high titers of GM1 and Hashimoto's thyroiditis and reinforces the multifocal motor neuropathy autoimmune origin as well as the repeated clinical recoveries after intravenous immunoglobulins. This case also suggests to deeply investigate the thyroid function in patients with multifocal motor neuropathy. PMID- 12117482 TI - Diaphragm kinetics during pneumatic belt respiratory assistance: a sonographic study in Duchenne muscular dystrophy. AB - The principal aim of this study was to demonstrate the usefulness of M-mode sonography as a noninvasive technique to evaluate diaphragm excursion. The secondary aim was to assess the efficacy of pneumatic abdomino-diaphragmatic belt ventilation in patients with Duchenne muscular dystrophy. Using M-mode sonography, we measured the amplitude of diaphragm excursion in seven patients with Duchenne muscular dystrophy in various positions (0 degrees, 45 degrees, 75 degrees ) with and without pneumatic abdomino-diaphragmatic belt respiratory assistance. The belt significantly increased mean amplitude of diaphragm excursion by 62% at 45 degrees and by 55% at 75 degrees, and increased mean tidal volume by 43.5% at 45 degrees and by 49% at 75 degrees. Two patients were unable to tolerate the horizontal position (0 degrees ) During quiet breathing without the belt, amplitude of diaphragm excursion and tidal volume were positively correlated at 45 degrees (r=0.81; P=0.027) and 75 degrees (r=0.75; P=0.05). There was a significant intra-individual correlation between these two parameters during belt use but no inter-individual correlation. Without the belt, thoracic posture had no significant effect on amplitude of diaphragm excursion, either in quiet or deep breathing. After overnight respiratory assistance, arterial oxygen pressure and arterial oxygen saturation increased significantly, and arterial carbon dioxide pressure decreased from 52+/-6.4 to 46.4+/-4 mmHg. The pneumatic abdomino-diaphragmatic belt significantly improved gas exchanges and ventilation by increasing diaphragm excursion, as was clearly shown by noninvasive M-mode sonography. Indeed, M-mode sonography may be helpful in pneumatic abdomino diaphragmatic belt pressure adjustment. PMID- 12117483 TI - Dose-dependent effect of individualized respiratory muscle training in children with Duchenne muscular dystrophy. AB - The aim of this study was to evaluate the effects of low intensity, home inspiratory muscle training on respiratory muscle endurance in children with Duchenne muscular dystrophy, using a double-blind protocol. The originality aspect of this study is the use of a reproducible method of endurance and of the same method for evaluation and training. We studied eight trained children (mean age 14.7+/-4.5 years) and eight control children (mean age, 12.6+/-1.8 years). For 6 weeks, children breathed twice a day for 10 min through a valve with either 30% (training group) or less than 5% (control group) of their maximum inspiratory pressure (P(imax)). The results showed (1) a 46% improvement in the time limit after training in the training group and no change in the control group and (2) a significant correlation between the total time of respiratory muscle training and the percentage of endurance improvement in the training group. We conclude that specific training improves respiratory muscle endurance in Duchenne muscular dystrophy and the effectiveness of training appears to be dependent on the quantity of training. PMID- 12117484 TI - Transected myofibres may remain permanently divided in two parts. AB - During regeneration of transected myofibres a scar is formed between their stumps. Myofibres restore their tendon-muscle-tendon continuity and contractile function by attaching to the scar with new myotendinous junctions. The scar contracts with time, and thereby the stumps are pulled close to each other. During early regeneration, myoblasts and myotubes can fuse with the surviving parts of the transected myofibres. However, it is not known whether it is possible that the opposite stumps could eventually fuse to reunite the divided parts of the transected fibres. In this study, we show in rat that even after 12 months the stumps remain attached to the separating scar by myotendinous junctions without showing definite fusion of the stumps. We conclude that transected myofibres probably remain permanently divided in two consecutive tendon-muscle-tendon units. PMID- 12117485 TI - Workshop report of the 89th ENMC International Workshop: Central Core Disease, 19th-20th January 2001, Hilversum, The Netherlands. PMID- 12117486 TI - 99th ENMC international workshop: myotonic dystrophy: present management, future therapy. 9-11 November 2001, Naarden, The Netherlands. PMID- 12117487 TI - A curious experiment: the paradigm switch from observation and speculation to experimentation, in the understanding of neuromuscular function and disease. AB - The four-link chain of the motor unit represents the contemporary end-point of some two millennia of evolving knowledge in neuroscience. The paradigm shift in neuromuscular epistemology occurred in the mid-17th century. In 1666, the newly graduated Dutch doctor, Jan Swammerdam (1637-1680) published his former investigations of dissected nerve-muscle preparations. These experiments comprised the quantum leap from observation and speculation, to that of experimentation in the field of neuroanatomy and neurophysiology. In what he termed 'A Curious Experiment' he also described the phenomenon of intrinsic muscle excitability - "I cannot observe that the muscle in the living animal ever absolutely ceases from all motion". Eighty years later (1752), von Haller demonstrated experimentally that irritability (contractility) was an intrinsic property of all muscular tissue; and distinguished between the sensibility of nerve impulses and the irritability of muscular contraction. This experimental progression from Swammerdam to von Haller culminated in 1850, when Claude Bernard's studies in experimental pharmacology confirmed that muscle was a functional unit, independent of any electrical innervation via its supplying nerve. This account comprises an audit of Swammerdam's work in the perspective of neuromuscular knowledge. PMID- 12117488 TI - Comment on Connolly AM, et al. Three mouse models of muscular dystrophy: the natural history of strength and fatigue in dystrophin-, dystrophin/utrophin-, and laminin alpha2-deficient mice (Neuromuscul Disord 2001;11:703-712). PMID- 12117490 TI - Natural and induced dyskinetoplastic trypanosomatids: how to live without mitochondrial DNA. AB - Salivarian trypanosomes are the causative agents of several diseases of major social and economic impact. The most infamous parasites of this group are the African subspecies of the Trypanosoma brucei group, which cause sleeping sickness in humans and nagana in cattle. In terms of geographical distribution, however, Trypanosoma equiperdum and Trypanosoma evansi have been far more successful, causing disease in livestock in Africa, Asia, and South America. In these latter forms the mitochondrial DNA network, the kinetoplast, is altered or even completely lost. These natural dyskinetoplastic forms can be mimicked in bloodstream form T. brucei by inducing the loss of kinetoplast DNA (kDNA) with intercalating dyes. Dyskinetoplastic T. brucei are incapable of completing their usual developmental cycle in the insect vector, due to their inability to perform oxidative phosphorylation. Nevertheless, they are usually as virulent for their mammalian hosts as parasites with intact kDNA, thus questioning the therapeutic value of attempts to target mitochondrial gene expression with specific drugs. Recent experiments, however, have challenged this view. This review summarises the data available on dyskinetoplasty in trypanosomes and revisits the roles the mitochondrion and its genome play during the life cycle of T. brucei. PMID- 12117491 TI - Effect of pH and temperature on protein kinase release by Leishmania donovani. AB - During their life cycle Leishmania are exposed to environments that differ markedly in pH and temperature. The effect of these factors on protein kinase release into the surrounding environment by Leishmania donovani promastigotes was examined. Promastigotes release protein kinase activity both constitutively and following induction by incubation with an exogenous substrate, phosvitin. The substrate specificity of the constitutive and induced activities was similar, unlike that previously described for Leishmania major promastigotes. The Leishmania donovani enzymes phosphorylate phosvitin, but not casein, mixed histones or protamine sulphate, and both activities are shed over a wide pH range from 6 to 9. Transfer of promastigotes from pH 7.4/30 degrees C to pH 5.0-5.5/37 degrees C, conditions that mimic those encountered by parasites following transmission from sandflies to a mammalian host and uptake by macrophages, inhibited release of the constitutive activity. Identical conditions had only a minor effect on induced protein kinase release. Both types of protein kinase activities released at pH 7.4 were still active when assayed at pH 5.0. Characterisation of the constitutive and induced promastigote protein kinases showed that casein kinase 1- and casein kinase 2-like activities are released by Leishmania donovani. Constitutive enzyme release decreased over time, however, the addition of phosvitin to these "casein kinase-depleted" promastigotes induced elevated casein kinase 1 and casein kinase 2 shedding. These results suggest that shed protein kinase might play a role in parasite survival and adaptation to host environments. PMID- 12117492 TI - Localisation of Globodera pallida FMRFamide-related peptide encoding genes using in situ hybridisation. AB - The present study employed an in situ hybridisation technique to detect the expression of a number of FMRFamide-like peptide encoding (flp) genes, previously identified from Globodera pallida, in whole-mount preparations of the J(2) stage of this worm. gpflp-1, encoding the FMRFamide-related peptide (FaRP) KSAYMRFamide, was expressed in neurones associated with the circumpharyngeal nerve ring and specifically in a number of cell bodies in the lumbar ganglia of the perianal nerve ring. The lumbar ganglia and pre-anal ganglia along with the BDU neurones and a number of cells in the retrovesicular ganglion were observed to express gpflp-2, encoding KNKFEFIRFamide. gpflp-3 (encoding KHEYLRFamide) expression was localised to the anterior ganglion and a number of paired cells posterior to the circumpharyngeal nerve ring whilst expression of gpflp-4, encoding a number of -P(G/Q)VLRFamides, was localised to the retrovesicular ganglion. No expression of gpflp-5 was observed. Identification of the reactive cells has implicated distinct roles for the FaRPs encoded on these genes in regulation of both dorsal and ventral body wall muscles, the musculature of the vulva and in the function of a number of sensory structures in both the head and tail of G. pallida. Comparison with the expression patterns of analogous genes in Caenorhabditis elegans suggests that, whilst some of the encoded peptides are conserved between nematode species, their functions therein are distinct. Furthermore, the expression of some of these genes in a number of interneurones supports the idea that FaRPs fulfil neuromodulatory as well as neurotransmitter roles. PMID- 12117493 TI - Molecular cloning and expression of the catalytic subunit of protein kinase A from Trypanosoma cruzi. AB - The activation of protein kinase A (cyclic adenosine monophosphate-dependent protein kinase) by cyclic adenosine monophosphate is believed to play an important role in regulating the growth and differentiation of Trypanosoma cruzi. A PCR using degenerate oligonucleotide primers against conserved motifs in the VIb and VIII subdomains of the ACG family of serine/threonine protein kinases was utilised to amplify regions corresponding to the parasite homologue of the protein kinase A catalytic subunit. This putative protein kinase A fragment was used to isolate the entire gene from T. cruzi genomic libraries. The deduced 329 amino acid sequence of this gene contained all of the signature motifs of known protein kinase A catalytic subunit proteins. The recombinant protein expressed in Escherichia coli was shown to phosphorylate Kemptide, a synthetic peptide substrate of protein kinase A, in a protein kinase inhibitor (PKI)-inhibitory manner. Immunoprecipitation with polyclonal antisera raised against recombinant protein of this gene was able to pull-down PKI-inhibitory phosphotransferase activity from epimastigote lysates. Immunoblot and Northern blot analyses, in combination with enzyme activity assays, revealed that this gene was a stage regulated enzyme in T. cruzi with higher levels and activity being present in epimastigotes compared with amastigotes or trypomastigotes. Overall these studies indicate that the cloned gene encodes an authentic protein kinase A catalytic subunit from T. cruzi and are the first demonstration of PKI-inhibitory phosphotransferase activity in an expressed protozoan protein kinase A catalytic subunit. PMID- 12117494 TI - Competition for minerals between Acanthocephalus lucii and its definitive host perch (Perca fluviatilis). AB - Concentrations of various essential and toxic elements (Ba, Ca, Co, Cu, Fe, Mg, Mn, Sr, Zn and Al, Ag, Cd, Cr, Ni, Pb, Tl) were analysed by inductively coupled plasma mass spectrometry in the acanthocephalan Acanthocephalus lucii and in different tissues of its host Perca fluviatilis. Nearly all the elements were found in significantly higher concentrations in the acanthocephalan than in the host tissues. Spearman correlation analysis revealed several strong inter-element associations within the organs of perch and within the parasites. Furthermore, statistical analysis revealed different competitive interactions. The concentrations of several essential elements (Ba, Ca, Fe, Mn, Sr and Zn) within the parasites decreased with an increasing number or weight of worms inside the intestine of the host. Additionally, the levels of some elements in the perch liver were negatively correlated with the weight of A. lucii in the intestine. Thus, it emerged that not only is there competition for essential elements between acanthocephalans inside the gut but there is also competition for these elements between the host and its parasites. PMID- 12117495 TI - ITS 2 sequences heterogeneity in Phlebotomus sergenti and Phlebotomus similis (Diptera, Psychodidae): possible consequences in their ability to transmit Leishmania tropica. AB - An intraspecific study on Phlebotomus sergenti, the main and only proven vector of Leishmania tropica among the members of the subgenus Paraphlebotomus was performed. The internal transcribed spacer 2 (ITS2) sequences of 12 populations from 10 countries (Cyprus, Egypt, Italy, Lebanon, Morocco, Pakistan, Portugal, Spain, Syria, and Turkey) were compared. Samples also included three species closely related to P. sergenti: Phlebotomus similis (three populations from Greece and Malta), Phlebotomus jacusieli and Phlebotomus kazeruni. Our results confirm the validity of the taxa morphologically characterised, and imply the revision of their distribution areas, which are explained through biogeographical events. At the Miocene time, a migration route, north of the Paratethys sea would have been followed by P. similis to colonise the north of the Caucasus, Crimea, Balkans including Greece and its islands, and western Turkey. Phlebotomus sergenti would have followed an Asiatic dispersion as well as a western migration route south of the Tethys sea to colonise North Africa and western Europe. This hypothesis seems to be well supported by high degree of variation observed in the present study, which is not related to colonisation or to intra-populational variation. Two groups can be individualised, one oriental and one western in connection with ecology, host preferences and distribution of L. tropica. We hypothesise that they could be correlated with differences in vectorial capacities. PMID- 12117496 TI - Apoptosis in the malaria protozoan, Plasmodium berghei: a possible mechanism for limiting intensity of infection in the mosquito. AB - Death by apoptosis regulates cell numbers in metazoan tissues and it is mediated by activation of caspases and results in characteristic morphological and biochemical changes. We report here that the malaria protozoan, Plasmodium berghei, exhibits features typical of metazoan apoptotic cells including condensation of chromatin, fragmentation of the nuclear DNA and movement of phosphatidylserine from the inner to the outer lamellae of the cell membrane. In addition, proteins with caspase-like activity were identified in the cytoplasm of the ookinete suggesting that the cellular mechanism of cell death may be similar to that of multicellular eukaryotes. Our data show that more than 50% of the mosquito midgut stages of the parasite die naturally by apoptosis before gut invasion. Cell death was prevented by a caspase inhibitor, treatment resulting in a doubling of parasite intensity. All these features also occur in vitro. Cell suicide thus plays a major and hitherto unrecognised role in controlling parasite populations and could be a novel target for malaria control strategies. PMID- 12117497 TI - Diplostomum spathaceum cercariae respond to a unique profile of cues during recognition of their fish host. AB - During its normal life cycle, Diplostomum spathaceum cercariae attach to and invade fish intermediate hosts. They are also known to attach to various other aquatic animals in response to water currents, touch and carbon dioxide. The purpose of this study was to identify the specific stimuli used by D. spathaceum cercariae to recognise the appropriate fish host. We characterised the host cues which stimulate them to remain on the host (enduring contact) and to penetrate the skin. Cercariae were exposed to animal skin tissues and fish skin surface mucus, their extracts and chemical modifications integrated into agar or offered via membrane filters. Enduring contact was stimulated by hydrophilic extracts Mr<3kDa, which were sensitive to oxidation of carbohydrates. The stimulating cues are probably small molecular carbohydrates, as monosaccharides stimulated enduring contacts, but amino acids, urea, electrolytes and peptides did not. Penetration was stimulated by hydrophilic macromolecules, Mr>30kDa, and by lipids. The hydrophilic stimuli were protease resistant and precipitable with Alcian blue and they were sensitive to alkaline cleavage, to digestion with lysozyme and neuraminidase as well as to oxidation of sialic acids. They were considered to be glycoproteins with O-glycosidically linked carbohydrate chains and bound sialic acids as signal structures. The lipophilic penetration stimuli were contained exclusively in the fatty acid fractions, and the stimulating characteristics of these fatty acids resembled the stimulating penetrations in other cercarial species. Diplostomum spathaceum cercariae respond to a unique profile of cues in their sequence of host-recognition phases. These cues differ from those used in other fish parasites studied to date and underline the diversity of fish recognition strategies. PMID- 12117498 TI - Excretory/secretory products from plerocercoids of Spirometra erinaceieuropaei suppress the TNF-alpha gene expression by reducing phosphorylation of ERK1/2 and p38 MAPK in macrophages. AB - We previously reported that excretory/secretory products from plerocercoids of Spirometra erinaceieuropaei suppress gene expression and production of tumour necrosis factor-alpha in murine macrophages stimulated with lipopolysaccharide. The present study investigated the suppressive mechanisms of tumour necrosis factor-alpha mRNA by excretory/secretory products in lipopolysaccharide stimulated murine macrophages. Electrophoretic mobility shift assay and supershift assay revealed that neither nuclear translocation of nuclear factor kappa B nor conformation of the p50/p65 nuclear factor-kappa B subunits was affected by the treatment of excretory/secretory products in lipopolysaccharide stimulated macrophages. Inhibition of extracellular signal-regulated protein kinase 1/2 with PD98059 or p38 mitogen-activated protein kinase with SB203580 partially reduced tumour necrosis factor-alpha mRNA expression, and a combination of the two inhibitors additionally suppressed the level of tumour necrosis factor alpha mRNA, revealing that both pathways are crucial for full induction of the gene. Northern blot analysis showed that excretory/secretory products additionally suppressed tumour necrosis factor-alpha mRNA expression in cells treated with PD98059 or SB208530 and, in turn, we found that excretory/secretory products reduced phosphorylation of extracellular signal-regulated protein kinase 1/2 and p38 mitogen-activated protein kinase in lipopolysaccharide-stimulated macrophages by Western blot analysis. This is the first report demonstrating that excretory/secretory products from parasites suppress tumour necrosis factor-alpha mRNA expression by reducing phosphorylation of extracellular signal-regulated protein kinase 1/2 and p38 mitogen-activated protein kinase without any effect on nuclear factor-kappa B activity in macrophages stimulated with lipopolysaccharide. We hypothesise that excretory/secretory products may enable this parasite to survive within the host. PMID- 12117499 TI - Reverse transcriptase activity and untranslated region sharing of a new RTE-like, non-long terminal repeat retrotransposon from the human blood fluke, Schistosoma japonicum. AB - A new RTE-like, non-long terminal repeat retrotransposon, termed SjR2, from the human blood fluke, Schistosoma japonicum, is described. SjR2 is approximately 3.9 kb in length and is constituted of a single open reading frame encoding a polyprotein with apurinic/apyrimidinic endonuclease and reverse transcriptase domains. The open reading frame is bounded by 5'- and 3'-terminal untranslated regions and, at its 3'-terminus, SjR2 bears a short (TGAC)(3) repeat. Phylogenetic analyses based on conserved domains of reverse transcriptase or endonuclease revealed that SjR2 belonged to the RTE clade of non-long terminal repeat retrotransposons. Further, SjR2 was homologous, but probably not orthologous, to SR2 from the African blood fluke, Schistosoma mansoni; this RTE like family of non-long terminal repeat retrotransposons appears to have arisen before the divergence of the extant schistosome species. Hybridisation analyses indicated that approximately 10,000 copies of SjR2 were dispersed throughout the S. japonicum chromosomes, accounting for up to 14% of the nuclear genome. Messenger RNAs encoding the reverse transcriptase and endonuclease domains of SjR2 were detected in several developmental stages of the schistosome, indicating that the retrotransposon was actively replicating within the genome of the parasite. Exploration of the coding and non-coding regions of SjR2 revealed two notable characteristics. First, the recombinant reverse transcriptase domain of SjR2 expressed in insect cells primed reverse transcription of SjR2 mRNA in vitro. By contrast, recombinant SjR2-endonuclease did not appear to cleave schistosome or plasmid DNA. Second, the 5'-untranslated region of SjR2 was >80% identical to the 3'-untranslated region of a schistosome heat shock protein-70 gene (hsp-70) in the antisense orientation, indicating that SjR2-like elements were probably inserted into the non-coding regions of ancestral S. japonicum HSP 70, probably after the species diverged from S. mansoni. PMID- 12117500 TI - Cloning, production and characterisation of a recombinant Cu/Zn superoxide dismutase from Taenia solium. AB - A full-length complementary DNA clone encoding a cytosolic Cu/Zn superoxide dismutase with a M(r) of 15,588 Da was isolated from a Taenia solium larvae complementary DNA library. Comparison analysis of its deduced amino acid sequence revealed a 71% identity with Schistosoma mansoni, 57.2-59.8% with mammalian and less than 54% with other helminth cytosolic Cu/Zn superoxide dismutase. The characteristic motifs and the amino acid residues involved in coordinating copper and zinc enzymatic function are conserved. The T. solium Cu/Zn superoxide dismutase was expressed in the pRSET vector. Enzymatic and filtration chromatographic analysis showed a recombinant enzyme with an activity of 2,941 U/mg protein and a native M(r) of 37 kDa. Inhibition assays using KCN, H(2)O(2), NaN(3) and SDS indicated that Cu/Zn is the metallic cofactor in the enzyme. Thiabendazole (500 microM) and albendazole (300 microM) completely inhibited the activity of T. solium Cu/Zn superoxide dismutase. Thiabendazole had no effect on bovine Cu/Zn superoxide dismutase; in contrast, albendazole had a moderate effect on it at same concentrations. Antibodies against T. solium Cu/Zn superoxide dismutase did not affect the enzymatic function; nevertheless, it cross reacts with several Taenia species, but not with trematodes, nematodes, pig, human and bovine Cu/Zn superoxide dismutase enzymes. Western blot analysis indicated the enzyme was expressed in all stages. These results indicate that T. solium possesses a Cu/Zn superoxide dismutase enzyme that can protect him from oxidant damage caused by the superoxide anion. PMID- 12117502 TI - Prevalence of Toxoplasma gondii in commercial meat products as monitored by polymerase chain reaction--food for thought? AB - DNA was extracted from 71 meat samples obtained from UK retail outlets. All of these DNA preparations gave the expected polymerase chain reaction products when amplified with primers specific for the species from which the meat originated. A second polymerase chain reaction analysis, using primers specific for the Toxoplasma gondii SAG2 locus, revealed the presence of this parasite in 27 of the meat samples. Restriction analysis and DNA sequencing showed that 21 of the contaminated meats contained parasites genotyped as type I at the SAG2 locus, whilst six of the samples contained parasites of both types I and II. Toxoplasma- positive samples were subjected to further polymerase chain reaction analysis to determine whether any carried an allele of the dihydropteroate synthase gene that has recently been shown to be causally associated with sulfonamide resistance in T. gondii. In all cases, this analysis confirmed that parasites were present in the samples and, additionally, revealed that none of them carried the drug resistant form of dihydropteroate synthase. These results suggest that a significant proportion of meats commercially available in the UK are contaminated with T. gondii. Although none of the parasites detected in this study carried the sulfonamide-resistance mutation, a simplified procedure for monitoring this situation merits development. PMID- 12117501 TI - The symbiont Capsaspora owczarzaki, nov. gen. nov. sp., isolated from three strains of the pulmonate snail Biomphalaria glabrata is related to members of the Mesomycetozoea. AB - While investigating the resistance of some strains of Biomphalaria glabrata to infection with Schistosoma mansoni, a unicellular eukaryotic symbiont was noted in the snail haemolymph. It was similar in appearance to Nuclearia sp. reported from B. glabrata. Sequences comprising the 18S, ITS1, 5.8S, ITS2 and the beginning of the 28S rDNA gene regions were obtained from symbionts isolated from three strains of B. glabrata, and compared with the same sequences obtained from a culture of Nuclearia sp. 18S rDNA sequences were identical for all four isolates. 18S rDNA sequences were used in a phylogenetic analysis to produce minimum evolution, maximum parsimony, maximum likelihood and Bayesian trees. All four analyses indicated that the B. glabrata symbiont is not closely related to Nuclearia but instead to the Mesomycetozoea, a recently recognised clade of symbiotic eukaryotes. Based on phylogenetic analysis, life history and morphological differences, the symbiont is described as a new genus and species, Capsaspora owczarzaki. Distinguishing characters are the presence of life cycle stage(s) that occur within snail haemolymph; ability to kill and ingest digenetic trematode larvae; ability to undergo asexual fission to produce daughter cells; absence of flagella, a mucous sheath and membranes containing chitin, elastin, or collagen; and presence of long unbranching pseudopodia and a penetration process. Using both polymerase chain reaction (PCR) and culturing techniques, the S. mansoni-resistant Salvador and 13-16-R1 strains were found to be significantly more likely to harbour the symbiont than the susceptible M line strain. Small but consistent sequence differences were noted among symbiont isolates from different snail strains, raising the possibility that the symbiont has diverged in different snail lineages. This suggests further that the symbiont is not restricted to albino lab-reared snails. A role, if any, of the symbiont in resistance awaits further study. PMID- 12117503 TI - Unusual prevalence of the Giardia intestinalis A-II subtype amongst isolates from humans and domestic animals in Mexico. PMID- 12117504 TI - Serotonin in the inferior colliculus. AB - It has been recognized for some time that serotonin fibers originating in raphe nuclei are present in the inferior colliculi of all mammalian species studied. More recently, serotonin has been found to modulate the responses of single inferior colliculus neurons to many types of auditory stimuli, ranging from simple tone bursts to complex species-specific vocalizations. The effects of serotonin are often quite strong, and for some neurons are also highly specific. A dramatic illustration of this is that serotonin can change the selectivity of some neurons for sounds, including species-specific vocalizations. These results are discussed in light of several theories on the function of serotonin in the IC, and of outstanding issues that remain to be addressed. PMID- 12117505 TI - Unbiased stereological estimates of neuron number in subcortical auditory nuclei of the rat. AB - The mammalian auditory system consists of a large number of cell groups, each containing its own complement of neuronal cell types. In recent years, much effort has been devoted to the quantitation of auditory neurons with common morphological, connectional, pharmacological or functional features. However, it is difficult to place these data into the proper quantitative perspective due to our lack of knowledge of the number of neurons contained within each auditory nucleus. To this end, we have employed unbiased stereological methods to estimate neuron number in the cochlear nuclei, superior olivary complex, lateral lemniscus, inferior colliculus and medial geniculate body. Additionally, we generated a three-dimensional model of the superior olivary complex. The utility of unbiased stereological estimates of auditory nuclei is discussed in the context of various experimental paradigms. PMID- 12117506 TI - AMPA and NMDA receptors mediate synaptic excitation in the rat's inferior colliculus. AB - The synaptic mechanisms underlying excitation in the rat's central nucleus of the inferior colliculus (ICC) were examined by making whole-cell patch clamp recordings in brain slice preparations of the auditory midbrain. Responses were elicited by current pulse stimulation of the lateral lemniscus and recordings were made in ICC using either current clamp or voltage clamp methods. The excitatory postsynaptic responses in either current or voltage clamp mode consisted of two distinct components, an early component that could be blocked by bath application of the alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor antagonists, 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) or 1,2,3,4-tetrahydro-6-nitro-2,3-dioxo-benzo[f]quinoxaline-7-sulfonamide (NBQX), and a later component that could be blocked by application of the N-methyl-D aspartate (NMDA) receptor antagonists, (+/-)-2-amino-5-phosphonovaleric acid (APV) or (+/-)-3-(2-carboxypiperazin-4-yl)-propyl-1-phosphonic acid (CPP). Both AMPA and NMDA receptor-mediated responses were present at resting potential and could be isolated pharmacologically by application of receptor antagonists. Voltage clamp experiments revealed that the NMDA receptor-mediated current was voltage-dependent and increased in magnitude as the cell membrane was depolarized. This NMDA receptor-mediated response was enhanced at resting potential when Mg(2+) was eliminated from the bath solution. The ratio of response amplitudes associated with the late and early components, an estimate of the relative contribution of NMDA and AMPA receptor types, changed with age. There was a progressive decline in the ratio between 9 and 13 days of age, but no further reduction between days 13 and 16. The data show that both AMPA and NMDA receptors are important for determining excitatory responses in the ICC and that both receptor types probably play a role in auditory processing after the onset of hearing. PMID- 12117507 TI - Contribution of AMPA and NMDA receptors to excitatory responses in the inferior colliculus. AB - Brain slice studies of neurons in the central nucleus of the inferior colliculus (ICC) indicate that excitatory responses evoked by electrical stimulation of the lateral lemniscus consist of two components, an early, rapid response mediated by alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors and a later, a slower one mediated by N-methyl-D-aspartate (NMDA) receptors. The early response can be selectively blocked by AMPA receptor antagonists (1,2,3,4 tetrahydro-6-nitro-2,3-dioxo-benzo[f]quinoxaline-7-sulfonamide disodium [NBQX]; or 6-cyano-7-nitroquinoxaline-2,3-dione) [CNQX], and the later one by NMDA receptor antagonists ((+/-)-3-(2-carboxypiperazin-4-yl)-propyl-1-phosphonic acid [CPP]; or (+/-)-2-amino-5-phosphonovaleric acid) [APV]. Both AMPA and NMDA receptor-mediated responses can be elicited at resting potential, although the NMDA response is voltage dependent and makes a greater contribution when the cell membrane is depolarized. In vivo studies indicate that both AMPA and NMDA receptors contribute to sound-evoked responses. Both AMPA and NMDA receptor antagonists reduce the firing rate of single neurons in the ICC to contralaterally presented tones. Both classes of antagonist lower evoked activity over a wide range of sound intensities from threshold to maximum sound pressure levels. Thus, both NMDA and AMPA receptors contribute to responses over the full dynamic range of auditory sensitivity. The AMPA receptor antagonist, NBQX, is more effective than the NMDA receptor antagonist, CPP, in blocking responses of onset cells. Furthermore, NBQX and CPP have preferential effects in blocking the early or late responses of neurons that exhibited sustain activity to a 100 ms tone. Excitatory responses to sinusoidally amplitude-modulated stimuli are also reduced by application of either AMPA or NMDA antagonists. However, the synchrony of firing of action potentials to the modulation period (vector strength) is largely unaffected. The data suggest that the synchrony of firing of neurons in the inferior colliculus is determined primarily by the pattern of activity at lower levels of the auditory pathway and/or the local intrinsic properties of the cells. PMID- 12117508 TI - Synaptic modification in neurons of the central nucleus of the inferior colliculus. AB - Whole-cell patch clamp recordings were made from neurons in the central nucleus of the inferior colliculus (ICC) in brain slices from rat (8-13 days old). ICC neurons were classified by their discharge pattern in response to depolarizing and hyperpolarizing current injection. Excitatory postsynaptic currents (EPSCs) were elicited by stimulation of synaptic inputs under the condition that the synaptic inhibition was suppressed by strychnine and picrotoxin. EPSCs in all tested types of ICC neurons showed posttetanic, long-term potentiation (LTP) and long-term depression with tetanic stimulation. The potentiated EPSCs consisted of alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor and NMDA receptor mediated components. The magnitude of LTP was larger when the intracellular concentration of the calcium buffer ethylene glycol-bis (beta aminoethyl ether)-N,N,N',N'-tetracetic acid (EGTA) was lower and stimulation frequency was higher in cells with rebound firing patterns. Blocking N-methyl-D aspartate (NMDA) receptors in rebound cells prevented generation of LTP. These results suggest that excitatory synaptic transmission in ICC neurons can be modified. LTP in the auditory midbrain may be important for activity-dependent, adaptive changes in response to normal and pathological stimulus conditions. PMID- 12117509 TI - Peripheral auditory processing, the precedence effect and responses of single units in the inferior colliculus. AB - The purpose of this paper is to illustrate how interactions occurring within the auditory periphery are relevant to the interpretation of neurophysiological data obtained in recent studies seeking to find physiological correlates of 'the precedence effect'. Similar information was presented orally at the recent International Symposium on the Central Auditory System held in Salamanca, Spain. The physiological data of interest are responses from single neural units in the inferior colliculus recorded following stimulation by successive pairs of binaural clicks. We show how peripheral, monaural, within-filter interactions of such successive clicks can produce internal values of interaural temporal differences (ITDs) and interaural intensitive differences (IIDs) that can differ greatly from those present in the external stimulus. These interactions can produce unintended internal ITDs and IIDs that are well 'outside the tuning range' of the single unit being studied. When this occurs, the responses of the single units to the pairs of clicks would be expected to diminish. We also discuss how hair cell-related adaptation and compression can also lead to diminished responses. It is suggested that effects resulting from peripheral interactions should be taken into account or evaluated quantitatively before other factors, including central inhibition, are invoked. PMID- 12117511 TI - Transformations in processing interaural time differences between the superior olivary complex and inferior colliculus: beyond the Jeffress model. AB - Interaural time differences (ITDs) are used to localize sounds and improve signal detection in noise. Encoding ITDs in neurons depends on specialized mechanisms for comparing inputs from the two ears. Most studies have emphasized how the responses of ITD-sensitive neurons are consistent with the tenets of the Jeffress model. The Jeffress model uses neuronal coincidence detectors that compare inputs from both sides and delay lines so that different neurons achieve coincidence at different ITDs. Although Jeffress-type models are successful at predicting sensitivity to ITDs in humans, in many respects they are a limited representation of the responses seen in neurons. In the superior olivary complex (SOC), ITD sensitive neurons are distributed across both the medial (MSO) and lateral (LSO) superior olives. Similar response types are found in neurons sensitive to ITDs in two signal types: low-frequency sounds and envelopes of high-frequency sounds. Excitatory-excitatory interactions in the MSO are associated with peak-type responses, and excitatory-inhibitory interactions in the LSO are associated with trough-type responses. There are also neurons with responses intermediate between peak- and trough-type. In the inferior colliculus (IC), the same basic types remain, presumably due to inputs arising from the MSO and LSO. Using recordings from the SOC and IC, we describe how the response types can be described within a continuum that extends to very large values of ITD, and compare the functional organization at the two levels. PMID- 12117510 TI - Roles of inhibition for transforming binaural properties in the brainstem auditory system. AB - This review is concerned with the operation of circuits in the central auditory system, how they transform response features and what functional significance may be attributed to those transformations. We focus on the role that GABAergic inhibition plays in processing interaural intensity disparities (IIDs), the principal cues for localizing high frequencies, and the transformations of IID coding that occur between the superior olivary complex and the inferior colliculus (IC). IIDs are coded by excitatory-inhibitory (EI) cells, so called because they are excited by one ear and inhibited by the other. EI neurons are first created in the lateral superior olive (LSO), but they also dominate the dorsal nucleus of the lateral lemniscus (DNLL) and regions of the IC. The three nuclei are intimately linked through a complex arrangement of excitatory and inhibitory connections. One of these is a crossed excitatory projection from the LSO to both the DNLL and IC. The binaural properties of EI neurons in LSO, DNLL and IC appear strikingly similar, suggesting that the EI properties created in the LSO are simply imposed on the DNLL and IC through the crossed excitatory projections. Recent studies support the idea that EI properties created in lower centers are imposed on some IC cells. However, other studies show that the circuitry linking LSO, DNLL and IC generates a number of response transformations in many IC cells. These transformations include marked changes in EI properties with stimulus duration, the generation of highly focused spatial receptive fields, shifts in sensitivity to IIDs, and the de novo creation of the EI response property. All of these transformations are produced by inhibitory innervation of the IC. An additional emergent property is also imposed on IC cells that receive GABAergic innervation from the DNLL. That property is a change in the binaural features of the IC cell, a change produced by the reception of an earlier sound whose IID is strongly excitatory to the IC cell. We illustrate each of these transformations, propose circuitry that could account for the observed properties and suggest some functional relevance for each. In the final section, we discuss some of the inherent uncertainties associated with attributing functional consequences to response features and then consider whether the transformations found in some mammals are species-specific or are universal features of all mammals. PMID- 12117512 TI - Monaural and binaural processing in the ventral nucleus of the lateral lemniscus: a major source of inhibition to the inferior colliculus. AB - The ventral nucleus of the lateral lemniscus (VNLL) is a major source of input to the inferior colliculus. This paper reviews recent studies of neural responses in the VNLL of the unanesthetized rabbit. The VNLL has generally been viewed as a monaural nucleus, with its neurons responding primarily to stimulation of the contralateral ear. In the rabbit, the VNLL is divided into a medial division (VNLLm) comprising neurons intercalated in the medial limb of the lemniscus, a compact lateral division (VNLLl), and a dorsal division. The VNLLm contains an abundance of neurons sensitive to interaural temporal disparities (ITDs), one of the major binaural cues for sound localization. These neurons respond only at the onset of tones, and therefore appear to encode the ITDs of transients. Even in the VNLLl, many neurons are sensitive to binaural stimulation. The VNLLl contains a variety of neurons with different discharge patterns, the two most common of which are sustained and onset. The discharge patterns, frequency-tuning and dynamic ranges of VNLLl neurons indicate that this division is able to supply the inferior colliculus with a variety of inputs, each serving a different function in the analysis of sound. PMID- 12117513 TI - Auditory gating processes and binaural inhibition in the inferior colliculus. AB - Physiological/behavioral/perceptual responses to an auditory stimulus can be inhibited by another leading auditory stimulus at certain stimulus intervals, and have been considered useful models of auditory gating processes. Two typical examples of auditory gating are prepulse inhibition of the startle reflex and the precedence effect (echo suppression). This review summarizes studies of these two auditory gating processes with regard to their biological significance, cognitive modulation, binaural properties, and underlying neural mechanisms. Both prepulse inhibition and the precedence effect have gating functions of reducing the disruptive influence of the lagging sound, but prepulse inhibition has a much longer temporal window than the precedence effect. Attentional processes can modulate prepulse inhibition, and the listener's previous experience can modulate the precedence effect. Compared to monaural hearing, binaural hearing reduces prepulse inhibition but enhances the precedence effect. The inferior colliculus, the major structure of the auditory midbrain, plays an important role in mediating these two auditory gating processes, and inhibitory neural transmissions within the inferior colliculus may account for binaural inhibition observed in prepulse inhibition and lag suppression recorded in the inferior colliculus. The neural mechanisms underlying binaural inhibition in the inferior colliculus are also discussed. PMID- 12117514 TI - Temporal and spatial coding of periodicity information in the inferior colliculus of awake chinchilla (Chinchilla laniger). AB - Amplitude modulation responses and onset latencies of multi-unit recordings and evoked potentials were investigated in the central nucleus of inferior colliculus (ICC) in the awake chinchilla. Nine hundred and one recording sites with best frequencies between 60 and 30 kHz showed either phasic (18%), tonic (25%), or phasic-tonic (57%) responses. Of 554 sites tested for responses to modulation frequencies 73% were responsive and 57% showed clear preference for a narrow range of modulation frequencies. Well defined bandpass characteristics were found for 32% of rate modulation transfer functions (rate-MTFs) and 36% of synchronization MTFs (sync-MTFs). The highest best modulation frequency (BMF) of a bandpass rate-MTF was 600 Hz. Neurons with phasic responses to best-frequency tones showed strong phase coupling to modulation frequencies and were dominated by bandpass rate-MTFs and sync-MTFs. Most neurons with tonic responses showed bandpass tuning only for sync-MTFs. Both BMFs and onset latencies changed systematically across frequency-band laminae of the ICC. Low BMFs and long latencies were located medially and high BMFs and short latencies laterally. Latency distributions obtained with evoked potentials to clicks showed a similar gradient to the multi-unit data. These findings are in line with previous findings in different animals including humans and support the hypothesis that temporal processing results in a topographic arrangement orthogonal to the spectral processing axis, thus forming a second neural axis of the auditory system. PMID- 12117515 TI - Excitatory and facilitatory frequency response areas in the inferior colliculus of the mustached bat. AB - In the mustached bat's central nucleus of the inferior colliculus (ICC), many neurons display facilitatory or inhibitory responses when presented with two tones of distinctly different frequencies. Our previous studies have focused on spectral interactions between specific frequency bands contained in the bat's sonar vocalization. In this study, we describe excitatory and facilitatory frequency response areas across all frequencies in the mustached bat's audible range. We show that many neurons in the ICC have more extensive frequency interactions than previously documented. We recorded responses of 96 single units to single tones and combinations of two tones. Best frequencies of the units ranged from 59-15 kHz. Forty-one units had a single, excitatory frequency response area. The rest of the units had more complex frequency tuning that included multiple excitatory frequency response areas and facilitatory frequency response areas. Some of the facilitatory frequency interactions were between one sound with energy in a sonar frequency band and a second sound with energy in a non-sonar frequency band. We also found that neurons could be facilitated by more than one additional frequency band. Our findings of extensive frequency interactions in the ICC of the mustached bat suggest that some neurons may be well suited for the analysis of complex sounds, possibly including social communication sounds. PMID- 12117516 TI - The effect of two-tone stimulation on responses of two simultaneously recorded neurons in the inferior colliculus of the big brown bat, Eptesicus fuscus. AB - This study examined auditory responses of two simultaneously recorded neurons in the central nucleus of bat inferior colliculus (IC) under two-tone stimulation conditions. We specifically examined how a sound within the excitatory frequency tuning curve (FTC) of one IC neuron might affect responses of the other IC neuron in amplitude and frequency domains. Under this specific two-tone stimulation condition, responses of 82% neurons were suppressed and their excitatory FTCs sharpened. Responses of the other 18% neurons were facilitated and their excitatory FTCs broadened. Two-tone suppression was greater at low than at high stimulus amplitudes. Two-tone suppression also decreased with increasing recording depth and best frequency (BF) difference between each pair of neurons. The suppressive or facilitatory FTC of a neuron plotted under two-tone stimulation conditions was always within the excitatory FTC of the other neuron. Two-tone suppression or two-tone facilitation was weak near the BF but became increasingly strong with frequencies away from the BF. Biological significance of these findings is discussed. PMID- 12117517 TI - Responses of inferior colliculus neurons to harmonic and mistuned complex tones. AB - Responses of inferior colliculus neurons to simplified stimuli that may engage mechanisms that contribute to auditory scene analysis were obtained. The stimuli were harmonic complex tones, which are heard by human listeners as single sounds, and the same tones with one component 'mistuned', which are heard as two separate sounds. The temporal discharge pattern elicited by a harmonic complex tone usually resembled the same neuron's response to a pure tone. In contrast, tones with a mistuned component elicited responses with distinctive, stereotypical temporal patterns that were not obviously related to the stimulus waveform. For a particular stimulus configuration, the discharge pattern was similar across neurons with different pure-tone frequency selectivity. A computational model that compared response envelopes across multiple narrow bands successfully reproduced the stereotypical response patterns elicited by different stimulus configurations. The results suggest that mistuning created a temporally synchronous distributed representation of the mistuned component that could be identified by higher auditory centers in the presence of the ongoing response produced by the remaining components; this kind of representation might facilitate the identification of individual sound sources in complex acoustic environments. PMID- 12117518 TI - GABAergic inputs shape responses to amplitude modulated stimuli in the inferior colliculus. AB - The inferior colliculus (IC) is an important auditory processing center receiving inputs from lower brainstem nuclei, higher auditory and nonauditory structures, and contralateral IC. The IC, along with other auditory structures, is involved in coding information about the envelope of complex signals. Biologically relevant acoustic signals, including animal vocalizations and speech, are spectrally and temporally complex and display amplitude and frequency variations over time. Certain IC neurons respond selectively over a narrow range of modulation frequencies to sinusoidally amplitude modulated (SAM) stimuli. Responses to SAM stimuli can be measured in terms of discharge rate, with rate plotted against the modulation frequency to generate rate modulation transfer functions (rMTF). A role for the inhibitory neurotransmitter, gamma-aminobutyric acid (GABA), in shaping selective responses to SAM stimuli has been suggested. The present study examined the role of GABA in shaping responses to SAM stimuli in the IC of anesthetized chinchilla. Responses from 94 IC neurons were obtained before, during and after iontophoretic application of the GABA(A) receptor antagonist bicuculline methiodide. Complete responses to SAM stimuli were obtained from 55 extensively tested neurons, displaying band-pass (38) and low pass rMTFs (17). For neurons showing band-pass rMTFs, GABA(A) receptor blockade selectively increased discharge rate at low modulation frequencies for 14 units, increased discharge near the best modulation frequency for 12 units. For neurons showing low-pass rMTFs, GABA(A) receptor blockade selectively increased discharge rate at low modulation frequencies for nine units. GABA(A) receptor blockade consistently reduced peak modulation gain, producing low-pass gain functions in a subset of IC neurons. In support of previous findings suggesting that selective temporal responses to SAM stimuli are coded in lower brainstem nuclei, temporal responses to SAM stimuli were relatively unaffected by GABA(A) receptor blockade. These findings support a role for GABA in shaping selective rate responses to SAM stimuli for a subset of chinchilla IC neurons. PMID- 12117519 TI - Effects of excitatory amino acid antagonists on the activity of inferior colliculus neurons during sleep and wakefulness. AB - The contribution of N-methyl-D-aspartate to the response to sound of guinea pig inferior colliculus neurons was analyzed by recording single-unit activity before and after iontophoretic injection of a receptor specific antagonist, 2-amino-5 phosphonovaleric acid (AP5), during the sleep-waking cycle. The AP5 produced a significant firing decrease in most of the units recorded, while some neurons exhibited a particular decrease in the later part of the response. A latency reduction in one out of three units in paradoxical sleep was observed. A low proportion of them exhibited a significant firing increase. These actions were observed in wakefulness (W) as well as during sleep phases. We compared the action of kynurenic acid (Kyn) and the electrical stimulation of the auditory cortex on the same inferior colliculus neuron in anesthetized animals and during W. Both Kyn iontophoresis and cortical stimulation evoked similar changes, decreased firing rate in most inferior colliculus units, whereas a low proportion of them increased their discharge, in anesthetized guinea pigs and in W. Ascending as well as descending - efferent - glutamatergic fibers impinging on inferior colliculus neurons contribute to sound-evoked responses. The enhanced unitary activity observed in some neurons with after glutamatergic receptor blocking may indicate that polysynaptic pathways involving inhibitory neurons decreased their activity. These effects were observed in anesthetized and in behaving animals. PMID- 12117520 TI - Descending projections to the inferior colliculus from the posterior thalamus and the auditory cortex in rat, cat, and monkey. AB - Projections from the posterior thalamus and medial geniculate body were labeled retrogradely with wheat germ agglutinin conjugated to horseradish peroxidase injected into the rat, cat, and squirrel monkey inferior colliculus. Neurons were found ipsilaterally in the (1) medial division of the medial geniculate body, (2) central gray, (3) posterior limitans nucleus, and the (4) reticular part of the substantia nigra. Bilateral projections involved the (5) peripeduncular/suprapeduncular nucleus, (6) subparafascicular and posterior intralaminar nuclei, (7) nucleus of the brachium of the inferior colliculus, (8) lateral tegmental/lateral mesencephalic areas, and (9) deep layers of the superior colliculus. The medial geniculate projection was concentrated in the caudal one-third of the thalamus; in contrast, the labeling in the subparafascicular nucleus, substantia nigra, and central gray continued much further rostrally. Robust anterograde labeling corresponded to known patterns of tectothalamic projection. Biotinylated dextran amine deposits in the rat inferior colliculus revealed that (1) many thalamotectal cells were elongated multipolar neurons with long, sparsely branched dendrites, resembling neurons in the posterior intralaminar system, and that other labeled cells were more typical of thalamic relay neurons; (2) some cells have reciprocal projections. Similar results were seen in the cat and squirrel monkey. The widespread origins of descending thalamic influences on the inferior colliculus may represent a phylogenetically ancient feedback system onto the acoustic tectum, one that predates the corticocollicular system and modulates nonauditory centers and brainstem autonomic nuclei. Besides their role in normal hearing such pathways may influence behaviors ranging from the startle reflex to the genesis of sound induced seizures. PMID- 12117521 TI - Brief and short-term corticofugal modulation of acoustic signal processing in the bat midbrain. AB - This article reviews our recent studies of brief and short-term corticofugal modulation of signal processing in the central nucleus of the inferior colliculus (ICc) by electrical stimulation in the primary auditory cortex (AC). When cortical electrical stimulation was synchronized with an acoustic stimulus, auditory responses of ICc neurons were either inhibited or facilitated and the modulative effect typically vanished within 5-10 s after the stimulation. When cortical electrical stimulation synchronized with an acoustic stimulus was repetitively delivered for 30 min, corticofugal modulation of collicular responses typically persisted up to 40 min after the stimulation. In the frequency domain, cortical electrical stimulation decreased the excitatory frequency tuning curves (FTCs) and asymmetrically increased the lateral inhibitory FTCs of corticofugally inhibited ICc neurons but produced the opposite effect on corticofugally facilitated ICc neurons. Cortical electrical stimulation facilitated auditory responses of neurons in the external nucleus of the inferior colliculus (ICx) while electrical stimulation in the ICx decreased auditory responses of ICc neurons. Auditory responses of simultaneously recorded ICx and ICc neurons varied in opposite ways during cortical electrical stimulation or drug application to recorded ICx neurons. In the amplitude domain, cortical electrical stimulation compressed rate-amplitude functions so as to increase the slope of rate-amplitude functions of ICc neurons. This modulative effect decreased with increasing stimulus amplitude. The possible biological relevance of these findings is discussed. PMID- 12117522 TI - A critical review on the participation of inferior colliculus in acoustic-motor and acoustic-limbic networks involved in the expression of acute and kindled audiogenic seizures. AB - The main goal of this article is to review the key role that the inferior colliculus plays in the expression of acoustic-motor and acoustic-limbic integration involved, respectively, in acute and chronic audiogenic seizures. In order to put this in context, we will review the behavioral characterization of acute and chronic audiogenic seizures, neuroanatomical substrates, neurochemistry, neuropharmacology, electrophysiology, as well as the cellular and molecular mechanisms involved in their expression. Secondly, we will also correlate our results, collected from audiogenic seizures susceptible rats, before and after the genetic selection of our own audiogenic susceptible strain, and from those sensitized by lesions or drug microinjections, with those pertinent from the international literature. In brief, genetic or sensitized animals express acute audiogenic seizures as a wild running behavior preceding the onset of tonic-clonic seizures. The latter can have several presentations including opistotonus and fore- and hindlimb tonic hyperextensions, followed by clonic convulsions of fore- and hindlimbs. Chronic (kindled) audiogenic seizures change this behavioral expression, with similar patterns such as those present in temporal lobe epileptic seizures, intermingled with the original audiogenic seizure pattern, which is known to be dependent on brainstem networks. PMID- 12117523 TI - Role of GABA abnormalities in the inferior colliculus pathophysiology - audiogenic seizures. AB - gamma-Aminobutyric acid (GABA), acting at GABA(A) receptors, mediates inhibition in inferior colliculus (IC) central nucleus (ICc) neurons and plays a prominent role in mediating acoustically evoked non-monotonicity, offset inhibition, and binaural inhibition, and is also important in tonic inhibition. The IC plays an important role in a number of pathophysiological conditions that involve hearing, including tinnitus, age-related hearing loss, and audiogenic seizures (AGS). AGS are a major form of rodent neurological disorder that can be genetically mediated and can also be readily induced in both young and mature animals. A deficit in GABA-mediated inhibition in IC neurons has been shown to be a critical mechanism in genetic and induced forms of AGS. Thus, both endogenously evoked GABA-mediated inhibition and exogenously applied GABA are reduced in efficacy in IC neurons of rats that are susceptible to AGS. GABA-mediated inhibition in IC neurons is significantly more easily blocked by a GABA(A) antagonist in genetic and induced forms of AGS in vivo and in vitro. AGS can be induced in normal animals by treatments that reduce the effectiveness of GABA in the IC. Glutamate-mediated excitation is a critical element of neurotransmission in IC neurons, and excessive activation of glutamate receptors in the IC is also strongly implicated as the other major mechanism in the pathophysiology of AGS. These neurotransmitter abnormalities result in excessive firing of ICc neurons that acts as the critical initiation mechanism for triggering seizures in response to intense acoustic stimuli. PMID- 12117524 TI - Functional reorganization in chinchilla inferior colliculus associated with chronic and acute cochlear damage. AB - This paper describes some of the unexpected functional changes that occur in the inferior colliculus (IC) following noise- and drug-induced cochlear pathology. A striking example of this is the compensation that is seen in IC responsiveness after drug-induced selective inner hair cell (IHC) loss. Despite a massive reduction in the compound action potential (CAP) caused by partial IHC loss, the evoked potential amplitude from the IC shows little or no reduction. Acoustic trauma, which impairs cochlear sensitivity and tuning, also reduces the CAP amplitude. Despite this reduced neural input, IC amplitude sometimes increases at a faster than normal rate and the response amplitude is enhanced at frequencies below the hearing loss. Single unit recordings suggest the IC enhancement phenomenon may be due to the loss of lateral inhibition. After an acute traumatizing exposure to a tone located above the characteristic frequency (CF), approximately 50% of IC neurons show a significant increase in their spike rate, a significant expansion of the low frequency tail of the tuning curve and a significant improvement in sensitivity in the tail of the tuning curve. These changes suggest that IC neurons receive inhibition from a high frequency side band and that this inhibition is diminished by acoustic trauma above CF. To determine if side band inhibition was locally mediated, specific antagonist(s) to inhibitory neurotransmitters were applied and found to produce effects similar to acoustic trauma. The results suggest that lesioned-induced central auditory plasticity could contribute to several symptoms associated with sensorineural hearing loss such as loudness recruitment, tinnitus and poor speech discrimination in noise. PMID- 12117525 TI - Air versus bone conduction: an equal loudness investigation. AB - Air conduction (AC) versus bone conduction (BC) loudness balance testing was conducted at frequencies of 0.25, 0.5, 0.75, 1, 2, and 4 kHz for two groups: 23 normal hearing subjects and eight subjects with a mild to moderate pure sensorineural hearing loss. Narrow-band noise was presented interchangeably between earphones and a bone transducer fitted to the subjects. Loudness matching was carried out at each frequency and at the levels 30-80 dB hearing level (HL) (10 dB steps) in the following manner: the sound pressure from the earphones was fixed and the subject adjusted the output level of the bone transducer for equal loudness by bracketing the standard. The results revealed somewhat different loudness functions for AC and BC sound with a 6-10 dB difference in the AC and BC loudness functions for the normal hearing group over the dynamic range 30-80 dB HL at the frequencies 250-750 Hz. At the higher frequencies, 1-4 kHz, the difference was only 4-5 dB over the same dynamic range. Similar results were obtained for the sensorineural hearing-impaired group. The difference between the AC and the BC loudness functions may originate from changes with level of the AC sound path, e.g. contraction of the stapedius muscle, but also distortion from the bone transducer and tactile stimulation could have contributed to the results seen. PMID- 12117526 TI - Estimating auditory neuronal dynamic range using a fitted function. AB - To obtain the dynamic range of an auditory afferent, the neuron's firing rate is plotted versus stimulus level, and the dynamic range is taken as the difference between the threshold for evoked firing, and the level at which firing rate saturates. Those dynamic range endpoints are typically defined in terms of the neuron's spontaneous firing rate and its maximum firing rate, according to a plurality of schemes, each of which depends on user-chosen sets of numerical criteria. The dynamic ranges predicted by some of these schemes are compared for the first time, and the resulting estimates can differ by a factor of 2. A step can be taken towards standardizing the measurement of neuronal dynamic range, if dynamic range is incorporated into a rate-level function as a parameter. To build this function, it is first assumed that the neuron's rate-level response reaches half its maximum at a level half-way between the threshold and the level at saturation, i.e. at threshold plus half the dynamic range. Then the firing rates at threshold and at threshold plus dynamic range are defined according to the most popular of the endpoint schemes. The resulting equation produces credible estimates of neuronal properties when fitted, and correctly predicts the behavior of the slope of the empirical rate-level plot [McGee, 1983. M.S. thesis, Creighton University; Ohlemiller et al., 1991. J. Acoust. Soc. Am. 90, 274-287]. Thus, despite not being deterministic, the new equation has remarkable predictive power. When two of the rate-level functions are added and weighted, the resulting equation fits sloping-saturating data better than any functions presently employed. PMID- 12117527 TI - DPOAEs in young normal-hearing subjects with histories of otitis media: evidence of sub-clinical impairments. AB - To investigate the origin of the susceptibility to noise in subjects with histories of otitis media (OM), we assessed early sub-clinical impairments in normally hearing subjects with a history of OM using distortion product otoacoustic emissions (DPOAEs). DPOAEs of 213 normal-hearing subjects aged 18-24 years were obtained and comparisons of DPOAE levels in several groups as a function of OM past infections were tested by ANOVA. A main finding was that young normal-hearing subjects with a history of OM had significantly lower DPOAEs over all octaves tested compared to normal-hearing subjects without antecedent of OM. The mean difference was 3.5+/-1.1 dB in the 2-4 kHz zone, which was especially marked in subjects (n=21) that had undergone a myringotomy (6.6+/-1.5 dB) in the 4 kHz zone. The level of impairment seemed to depend on the severity of the past infection as characterised by the importance and the duration of the infectious effusions. PMID- 12117528 TI - The middle ear of gekkonoid lizards: interspecific variation of structure in relation to body size and to auditory sensitivity. AB - Wishing to assess the effects of the dimensions of the middle ear on the auditory sensitivity of gekkonoid lizards, we measured middle ear components in preserved geckos, which in life had yielded 'cochlear microphonics' audiograms. We examined two to seven specimens of 14 species. The measures of middle ear elements varied relative to head or body length similarly within species and among species. The areas of the external ear opening, tympanic membrane, and columellar footplate, and the ratio between the last two ('hydraulic lever'), were correlated with animal length. The hydraulic and mechanical (extracolumellar) lever ratios appeared to complement each other, the former being emphasized in large animals, the latter in small animals. The apparent auditory sensitivity correlated with the sizes of the animal, head and external ear opening, and negatively (insignificantly) correlated with the mechanical lever ratio. The correlation of sensitivity with the hydraulic lever was insignificant, perhaps due to a 'tympanic membrane lever' (catenary effect). The most sensitive frequency negatively correlated with the area of the external ear opening, the area of the tympanic membrane, and with the level of greatest sensitivity. It was positively correlated with the relative length of the cartilaginous portion of the ossicular chain. However, the number of hair cells in the basilar papilla, too, is known to correlate with animal size. Moreover, the least sensitive species were not only the smallest species, they were also the species known to lack a zone of unidirectional hair cells in the basilar papilla. Hence the apparent sensitivity hypothetically depends on both middle ear dimensions and summation of inner ear output. This hypothesis requires verification by other methods. PMID- 12117529 TI - Magnitudes and phases of human distortion-product otoacoustic emissions at 2f(1) f(2) against f(2)/f(1): effects of an audiometric notch. AB - The influence of the frequency ratio f(2)/f(1) of two pure-tone stimuli on the distortion-product otoacoustic emissions (DPOAEs) at 2f(1)-f(2) was assessed in 14 hearing-impaired human ears exhibiting a narrow audiometric notch at 4 kHz, whereas 11 normally hearing ears served as controls. A fixed-f(2) paradigm was used, with f(2) values varying from 2 to 8 kHz in 1-kHz steps. The intensities of the two stimuli were either 60 or 70 dB SPL. The magnitudes and phases of DPOAEs were extracted so that the bandpass filter-like profile of DPOAE magnitude against f(2)/f(1) (f(2)/f(1) magnitude function) could be characterized by the presence and position of its maximum, and DPOAE group delays were derived from the phase gradient of the DPOAE when f(1) varied. The main difference between normal and impaired ears occurred at 4 kHz in that, in contrast with normal ears, nine impaired ears out of 14 did not present any peak in their f(2)/f(1) magnitude function, while the remaining five ears only differed from normal ones by a slightly shifted maximum toward larger f(2)/f(1)s. Group delays were significantly shortened in all impaired ears, with a tendency to be shorter in the subset of ears with flat DPOAE magnitude profiles with f(2)/f(1). No clear effect of notch depth was observed, and with the chosen stimulus characteristics, no abnormal DPOAE result was observed whenever f(2) fell outside the audiometric notch. It is concluded that DPOAE group delays apparently provide useful objective clues of cochlear dysfunction, more sensitive than DPOAE magnitudes inasmuch as many of them remained near the normal range. Although a majority of f(2)/f(1) magnitude functions were clearly flattened when f(2) was impaired, this feature was less systematic. PMID- 12117530 TI - Differential thresholds of local field potentials and unit discharges in rat auditory cortex. AB - Thresholds for responses to tone bursts in the primary auditory field of pentobarbital-anesthetized rat are significantly lower for local field potentials (5.60 dB+/-1.76 S.E.M.) than for multiple unit discharges (17.80 dB+/-3.17), recorded simultaneously from the same microelectrode. The characteristic frequencies (CFs) of local field potentials provide a good 'predictive' estimate of unit CFs at their higher thresholds. The findings are consistent with the view that local field potentials in the auditory cortex reflect summed synaptic potentials rather than cellular discharges. PMID- 12117531 TI - Intra-cochlear administration of dexamethasone attenuates aminoglycoside ototoxicity in the guinea pig. AB - This study demonstrates the attenuation of aminoglycoside ototoxicity by cochlear infusion of dexamethasone (Dex) using a microcannulation-osmotic pump delivery system. The results indicate that treating the cochlea with Dex both before and after kanamycin administration was more effective in preventing ototoxicity than Dex treatment only after kanamycin administration. A concentration of 1 ng/ml Dex showed the greatest protective effect on both kanamycin-induced threshold shift of the auditory brainstem response and outer hair cell survival. These results show that the Dex treatment attenuates both functional and structural damage of the inner ear from aminoglycoside toxicity. PMID- 12117532 TI - Hypoplasia of spiral and Scarpa's ganglion cells in GABA(A) receptor beta(3) subunit knockout mice. AB - This study documents morphologic alterations in the spiral ganglion and Scarpa's ganglion from gamma-aminobutyric acid A (GABA(A)) receptor beta(3) subunit null mutant mice. The ganglion cells of the mutant mice were hypoplastic in hematoylin&eosin-stained sections. Hypoplasia was observed at every location of the spiral ganglion and Scarpa's ganglion except the apical cochlear turn. Calretinin immunostaining demonstrated a selective hypoplasia of calretinin negative cells at every location of spiral and Scarpa's ganglion cells, while the soma area of calretinin-positive cells was not affected by the gene deletion. Meanwhile, in the spiral ganglion of both wild type and knockout mice, there were apical to basal gradients in the soma size and the proportion of calretinin positive cells. The absence of statistically significant hypoplasia in hematoylin&eosin sections through the apical turn of the cochlea can be explained by the relatively higher proportion of calretinin-positive ganglion cells, which were unaffected by the gene deletion. These findings suggest that GABA(A) receptor isoforms containing the beta(3) subunit may play an important role in the development and differentiation of non-calyceal terminals of Scarpa's ganglion cells and type II and smaller type I spiral ganglion cells. PMID- 12117533 TI - Origin of the binaural interaction component in wave P4 of the short-latency auditory evoked potentials in the cat: evaluation of serial depth recordings from the brainstem. AB - There is no general agreement on the origin of the binaural interaction (BI) component in auditory brainstem responses (ABRs). To study this issue the ABRs to monaural and binaural clicks with various interaural time differences (ITDs) were simultaneously recorded from the vertex and from a recording electrode aiming at the superior olive (SO) in cats. Electrode path was along the fibers of the lateral lemniscus (LL). Binaural difference potentials (BDPs), which were computed by subtracting the sum of the two monaural responses from the binaural response, were obtained at systematic depths and across a range of ITD values. It was observed that only a specific BDP deflection recorded at the level at which lemniscal fibers terminate in the nuclei of LL coincided in time with the most prominent BDP in the cat's vertex-recorded ABRs, the BDP in their wave P4. As ITD was increased, the latency shifts and amplitude decrements of the scalp-recorded far-field BDP wave exactly followed those recorded at this lemniscal near-field BDP locus. The data support our hypothesis that the BI component in wave P4 results from a binaural reduction in dischargings of axons ascending in the LL, with this reduction due to contralateral inhibition of the discharge activity of the inhibitory-excitatory units in the lateral nucleus of the SO. Furthermore, at the level of the SO, the BDP in the responses to contra-leading binaural clicks always had larger magnitudes than those evoked by ipsi-leading ones. This bilateral asymmetry is consistent with the view that the BDP in scalp-recorded ABRs is related to the function of sound lateralization. PMID- 12117534 TI - OCP2 immunoreactivity in the human fetal cochlea at weeks 11, 17, 20, and 28, and the human adult cochlea. AB - The two most abundant proteins of the organ of Corti, OCP1 and OCP2, are acidic, cytosolic, low molecular weight proteins diffusely distributed within the cytoplasm of supporting cells. A recent study by Henzl et al. (2001) found first, that these two proteins co-localize with connexin 26 along the epithelial gap junction system and second, that OCP2 could participate with OCP1 in an organ of Corti-specific SCF complex (Skp1, cul1in, and Fbp), a ubiquitin ligase complex. Previous study has also implicated OCP2 in the recycling and regulation of intracellular K(+) efflux as well as pH homeostatic mechanisms. In the present study, we document the emergence and distribution features of OCP2 through various stages (weeks 11-28) of gestation in human fetal cochleae. Four fetal cochleae, the cochleae of a normal hearing human adult and a mature rat for positive control were fixed in 4% formalin within 2 h post mortem. Immunohistochemical studies were performed using a rabbit polyclonal antibody raised against a synthetic peptide corresponding to amino acids 3-16. Specimens were mounted in paraffin sections. Results show that OCP2 immunoreactivity is evident at a prenatal age of 11 weeks, peaks in expression at the onset of cochlear function at 20 weeks and achieves adult-like patterns of distribution just prior to histological maturation at 28 weeks. Though this protein could be associated with the development, maturation, and electrochemical maintenance of the cochlear gap junction system, the nature of this protein's function in the developing and mature human cochlea remains unclear. PMID- 12117535 TI - Interaction of spiral ganglion neuron processes with alloplastic materials in vitro(1). AB - The cochlear implant (CI) involves the introduction of alloplastic materials into the cochlea. While current implants interact with cochlear neurons at a distance, direct interactions between spiral ganglion (SG) neurites and implants could be fostered by appropriate treatment with neurotrophic factors. The interactions of fibroblasts and osteoblasts with alloplastic materials have been well studied in vitro and in vivo. However, interactions of inner ear neurons with such alloplastic materials have yet to be described. To investigate survival and growth behavior of SG neurons on different materials, SG explants from post-natal day 5 rat SG were cultured for 72 h in the presence of neurotrophin-3 (10 ng/ml) on titanium, gold, stainless steel, platinum, silicone and plastic surfaces that had been coated with laminin and poly-L-lysine. Neurite outgrowth was investigated after immunohistological staining for neurofilament, by image analysis to determine neurite extension and directional changes. Neurite morphology and adhesion to the alloplastic material were also evaluated by scanning electron microscopy (SEM). On titanium, SG neurites reached the highest extent of outgrowth, with an average length of 662 microm and a mean of 31 neurites per explant, compared to 568 microm and 21 neurites on gold, 574 microm and 24 neurites on stainless steel, 509 microm and 16 neurites on platinum, 281 microm and 12 neurites on silicone and 483 microm and 31 neurites on plastic. SEM revealed details of adhesion of neurites and interaction with non-neuronal cells. The results of this study indicate that the growth of SG neurons in vitro is strongly influenced by alloplastic materials, with titanium exhibiting the highest degree of biocompatibility with respect to neurite extension. The knowledge of neurite interaction with different alloplastic materials is of clinical interest, as development in CI technology leads to closer contact of implanted electrodes with surviving inner ear neurons. PMID- 12117536 TI - The alpha9alpha10 nicotinic acetylcholine receptor is permeable to and is modulated by divalent cations. AB - The native cholinergic receptor that mediates synaptic transmission between olivocochlear fibers and outer hair cells of the cochlea is permeable to Ca(2+) and is thought to be composed of both the alpha 9 and the alpha 10 cholinergic nicotinic subunits. The aim of the present work was to study the permeability of the recombinant alpha 9 alpha 10 nicotinic acetylcholine receptor to Ca(2+), Ba(2+) and Mg(2+) and its modulation by these divalent cations. Experiments were performed, by the two-electrode voltage-clamp technique, in Xenopus laevis oocytes injected with alpha 9 and alpha 10 cRNA. The relative divalent to monovalent cation permeability was high ( approximately 10) for Ca(2+), Ba(2+) and Mg(2+). Currents evoked by acetylcholine (ACh) were potentiated by either Ca(2+) or Ba(2+) up to 500 microM but were blocked by higher concentrations of these cations. Potentiation by Ca(2+) was voltage-independent, whereas blockage was stronger at hyperpolarized than at depolarized potentials. Mg(2+) did not potentiate but it blocked ACh-evoked currents (IC(50)=0.38 mM). In the absence of Ca(2+), the EC(50) for ACh was higher (48 microM) than that obtained with 1.8 mM Ca(2+) (14.3 microM), suggesting that potentiation by Ca(2+) involves changes in the apparent affinity of the alpha 9 alpha 10 receptor for ACh. PMID- 12117537 TI - Localization of efferent neurotransmitters in the inner ear of the homozygous Bronx waltzer mutant mouse. AB - Naturally occurring mutant mice provide an excellent model for the study of genetic malformations of the inner ear. Mice homozygous for the Bronx waltzer (bv/bv) mutation are severely hearing impaired or deaf and exhibit a 'waltzing' gait. Functional aspects of cochlear and vestibular efferents in the bv/bv mutant mouse are not well known. The present study was designed to evaluate several candidates of efferent neurotransmitters or neuromodulators including choline acetyltransferase (ChAT), gamma-aminobutyric acid (GABA), and calcitonin gene related peptide (CGRP) in the inner ear of the bv/bv mutant mouse. Ultrastructural investigations at both light and electron microscopic level were performed. Ultrastructural morphologic evaluations of the cochlea and the vestibular end-organs were also undertaken. It is demonstrated that ChAT, GABA and CGRP immunoreactivities are present in the cochlea and in vestibular end organs of bv/bv mutant mice. In the organ of Corti, immunoreactivity of ChAT, GABA and CGRP is confined to the inner spiral fibers, tunnel-crossing fibers, and the vesiculated nerve endings synapsing with outer hair cells. Interestingly, immunoreactivity was detectable even where inner hair cells appeared missing. Results also revealed malformations of the outer hair cells with synaptic contacts to efferent nerve endings consistently intact. In the neurosensory epithelia of the vestibular end-organs, the presence of ChAT, GABA, and CGRP immunoreactivity was localized at the vestibular efferents, with the exception of the macula of saccule. In one 8-month-old macula of utricle where the depletion of hair cells appeared highest, ChAT immunostaining was still discernible. Ultrastructural investigation demonstrated that vesiculated efferent nerve endings make synaptic contact with the outer hair cells in the organ of Corti and with type II hair cells in the vestibular end-organs. The present study provides further support that the efferent system in the bv/bv mutant inner ear is morphologically as well as functionally mature. These findings also demonstrate that if and when the onset of efferent degeneration in the bv/bv mutant inner ear occurs, it transpires subsequent to pathological conditions in the hair cells. The present findings give further indication that the efferent systems of the bv/bv mutant inner ear are independent of the afferent systems in many aspects including development, maturation as well as degeneration. PMID- 12117538 TI - Behavioral examinations of the level of auditory processing of speech context effects. AB - One of the central findings of speech perception is that identical acoustic signals can be perceived as different speech sounds depending on adjacent speech context. Although these phonetic context effects are ubiquitous in speech perception, their neural mechanisms remain largely unknown. The present work presents a review of recent data suggesting that spectral content of speech mediates phonetic context effects and argues that these effects are likely to be governed by general auditory processes. A descriptive framework known as spectral contrast is presented as a means of interpreting these findings. Finally, and most centrally, four behavioral experiments that begin to delineate the level of the auditory system at which interactions among stimulus components occur are described. Two of these experiments investigate the influence of diotic versus dichotic presentation upon two phonetic context effects. Results indicate that context effects remain even when context is presented to the ear contralateral to that of the target syllable. The other two experiments examine the time course of phonetic context effects by manipulating the silent interval between context and target syllables. These studies reveal that phonetic context effects persist for hundreds of milliseconds. Results are interpreted in terms of auditory mechanism with particular attention to the putative link between auditory enhancement and phonetic context effects. PMID- 12117539 TI - Female MRL.MpJ-Fas(lpr) autoimmune mice have greater hearing loss than males. AB - Although women make up approximately 60-70% of the patients with autoimmune hearing loss, little is known about the impact of gender on this cochlear disease. To explore this relationship of gender and autoimmune inner ear disease, an evaluation was made of cochlear function in male and female autoimmune MRL.MpJ Fas(lpr) mice. Autoimmune disease and hearing loss onset occur at 3-4 months of age, so mice were tested with auditory brainstem response audiometry at 3, 6, and 9 months of age to identify potential gender differences in thresholds. Sera also were analyzed for differences in the autoimmune factors of immune complexes, anti nuclear antibodies, and hematocrits. By 9 months of age the surviving mice showed a dramatic gender difference. Female mice had thresholds 25-45 dB higher than males at 4, 8, and 16 kHz, although male thresholds at 32 kHz had risen sufficiently to be statistically similar to those for females. No gender differences were seen in any of the systemic autoimmune factors. These findings of worse hearing in female autoimmune mice parallel a reported female preponderance in clinical immune hearing disorders. This potential gender influence in autoimmune inner ear disease must be better understood for effective evaluation and treatment of this disorder. PMID- 12117540 TI - A model for perilymphatic fistula induced hearing loss in the guinea pig cochlea. AB - In cases of sudden, reversible hearing loss where perilymphatic fistulas are thought to be the cause, a multitude of causes have been postulated. These include perilymphatic pressure changes, Simmons' double-break theory, perilymphatic hemorrhage, pneumolabyrinth, and others. This study was proposed to explore the role pneumolabyrinth may have in this pathology. Guinea pigs were fitted with cochlear perfusion pumps pumping artificial perilymph into the left scala tympani. One group of animals received a bubble ( approximately 1.5-2 microl) introduced into the scala tympani while the other group of animals received continuous infusion of artificial perilymph. The animals' cochlear function was monitored using distortion product otoacoustic emissions (DPOAEs) while the animals' behavior was assessed to evaluate for vestibular dysfunction. While unaffected by pump surgery, animals that received air into the scala tympani had their DPOAEs eliminated by day 2 after pump placement. On day 6, positional changes in the DPOAEs were observed where the left lateral decubitus position showed a sharp demarcation at 4 kHz, where lower DPOAE frequencies were abolished and higher ones minimally affected, with DPOAEs normal in the prone position. By days 8 and 20, DPOAEs recovered to normal amplitudes. Vestibular dysfunction was never detected in any animal that received a pneumolabyrinth. All control animals receiving a continuous perilymph infusion exhibited no loss of cochlear function throughout the testing period. The reversible nature of pneumolabyrinth induced hearing loss and the pneumolabyrinth's sole presence (without other possible causes of hearing loss, such as pressure differentials or round window membrane perforations) are powerful indicators of the role of air in the pathology of perilymphatic fistulas. In addition, the fluctuation of the hearing loss with positional changes supports the use of positional audiometry when evaluating perilymphatic fistulas. PMID- 12117541 TI - Detection of intracochlear and intracranial pressure changes with otoacoustic emissions: a gerbil model. AB - Increased intracranial pressure (ICP) is known to affect the phases and levels of lower-frequency distortion-product otoacoustic emissions (DPOAE) in a characteristic manner suggestive of an increase in the stiffness of the stapes system, likely in relation to an attendant increased intracochlear pressure (ICoP). DPOAEs may thus provide an easy non-invasive means of gaining access to the otherwise elusive ICoP. However, the mechanisms by which DPOAEs actually relate to ICoP are unclear and may involve changes in the stiffness of the annular ligament, stapedius muscle and even some indirect contributions of other parts of the middle ear such as the tensor tympani. A systematic study of the role of each middle-ear element on ICoP-to-DPOAE relationships as a function of frequency was undertaken in gerbils under direct control of ICP via an intracranial catheter (from 0 to 500 daPa). After the bulla was widely opened, the tendons of the stapedius and tensor tympani muscles were severed in turn. A standard electroacoustic analog model of the middle ear was used for predicting the forward and reverse middle-ear transfer-functions changes under different experimental manipulations and their consequences on DPOAEs. The observed DPOAE changes chiefly consisted in a phase-lead peaking around 2.15 kHz in closed bulla, and 1.2 kHz in open-bulla conditions. It was proportional to ICP increase provided ICP exceeded a threshold of about 50 daPa. The profiles of DPOAE shifts matched those derived from the premise that ICoP mainly induced a change in the stiffness of the stapes system. The possible involvement of the stapedius muscle was ruled out by the absence of any effect of cutting its tendon so that the intrinsically non-linear stiffness of the annular ligament must have been the main factor. A relatively minor contribution from the tensor tympani was observed, possibly in relation to the detection of ICoP-induced displacement of the ossicular chain by neuromuscular spindles. PMID- 12117542 TI - Central auditory onset responses, and temporal asymmetries in auditory perception. AB - Historically, central auditory responses have been studied for their sensitivity to various parameters of tone and noise burst stimulation, with response rate plotted as a function of the stimulus variable. The responses themselves are often quite brief, and locked in time to stimulus onset. In the stimulus amplitude domain, it has recently become clear that these responses are actually driven by properties of the stimulus' onset transient, and this has had important implications for how we interpret responses to manipulations of tone (or noise) burst plateau level. That finding was important in its own right, but a more general scrutiny of the available neurophysiological and psychophysical evidence reveals that there is a significant asymmetry in the neurophysiological and perceptual processing of stimulus onsets and offsets: sound onsets have a more elaborate neurophysiological representation, and receive a greater perceptual weighting. Hypotheses about origins of the asymmetries, derived independently from psychophysics and from neurophysiology, have in common a response threshold mechanism which adaptively tracks the ongoing level of stimulation. PMID- 12117543 TI - Inputs from the cochlea and the inferior colliculus converge on olivocochlear neurones. AB - Medial olivocochlear (MOC) neurones, located in the superior olivary complex, can suppress cochlear gain by their action on the cochlear outer hair cells. Inputs from the contralateral cochlea and the inferior colliculus (IC) have been separately shown to increase activity of MOC neurones. In this study we have investigated in guinea-pigs under barbiturate anaesthesia the interactions between these two inputs by combining electrical stimulation of the IC with acoustic stimulation of the contralateral cochlea. Electrical stimulation of the IC resulted in a significant suppression of the amplitude of the compound action potential (CAP) of the auditory nerve to test tones. This suppression was equivalent to an average decrease in sound intensity of 5.7 dB and 3.7 dB for contralateral and ipsilateral stimulation, respectively. Acoustic stimulation of the contralateral cochlea with broadband noise produced no detectable change in the amplitude of the CAP in the test cochlea in all but one animal. However, simultaneous electrical stimulation of the IC and acoustic stimulation of the contralateral cochlea resulted in a reduction in CAP amplitude that was markedly larger than that produced by IC stimulation alone. The suppression with the addition of contralateral noise was equivalent to a mean reduction in sound intensity of 8.7 dB with contralateral and 5.7 dB with ipsilateral IC stimulation. We hypothesise that excitatory input from the contralateral cochlea converges with excitatory input from the IC on the MOC neurones and in this way augments the activity of these neurones, resulting in a larger peripheral effect. PMID- 12117544 TI - Alterations of basilar membrane response phase and velocity after acoustic overstimulation. AB - To investigate the physiology of noise-induced hearing loss, the sound-induced vibrations of the basilar membrane (BM) of the inner ear were measured in living anesthetized guinea pigs before and after intense sound exposure. The vibrations were measured using a laser Doppler velocimeter after placing reflective glass beads on the BM. Pseudo-random noise waveforms containing frequencies between 4 and 24 kHz were used to generate velocity tuning curves. Before overstimulation, sharp response peaks were seen at stimulus frequencies between 15 and 17 kHz, consistent with the expected best frequency of the recording location. The response to low level stimuli lagged the high level ones by up to 90 degrees at the characteristic frequency. Following exposure to loud sound, the BM vibrations showed a pronounced reduction in amplitude, primarily at low stimulus levels, and the best frequency moved to approximately 12 kHz. At higher levels, the reduction was either absent or much smaller. In addition to the amplitude changes, increased phase lags were seen at frequencies near the characteristic frequency. In animals with more severe exposures, response phases were altered also at frequencies showing no change of the amplitude. The phase was independent of stimulus level after severe exposures. PMID- 12117545 TI - Ontogeny of kainate receptor gene expression in the developing rat midbrain and striatum. AB - Kainate (KA) receptors are a family of ionotropic glutamate receptors, which mediate the excitatory synaptic transmission in various areas of the mammalian CNS. We have studied the expression pattern of the genes encoding for KA receptor subunits (Glur5-1, Glur5-2, Glur6, Glur7, KA1 and KA2) in rat prenatal (E), postnatal and adult ventral mesencephalon (MES) and striatum (STR) and in fetal midbrain primary cultures. Each receptor subunit shows a unique area- and temporal-expression pattern. In MES the onset of both Glur5 subunits is delayed when compared to the other subunits. In addition, most of the transcripts for KA subunits gradually increase during embryonic development and show a slight decrease during the first postnatal week. Differently, Glur6 and KA2 mRNAs show a sharp increase at E14.5 and decrease thereafter, reaching the lowest levels during late embryonic and postnatal development. In the STR, the gene expression of all KA subunit mRNAs is higher during embryonic development than after birth, except KA1 transcripts, that show a peak at P5. In embryonic MES primary cultures, Glur5-2, Glur6 and KA2 mRNAs are higher at the beginning of the culture when compared to older cultures, while the other subunit mRNAs do not show significant variation throughout the days in vitro. Thus, all the KA receptor subunit transcripts appear independently regulated during MES and STR development, probably contributing to the establishment of the fine tuning of the excitatory circuits reciprocally established between these CNS areas. PMID- 12117546 TI - Changes in rat frontal cortex gene expression following chronic cocaine. AB - Alterations in gene expression caused by repeated cocaine administration have been implicated in the long-term behavioral aspects of cocaine abuse. The frontal cortex mediates reinforcement, sensory, associative, and executive functions and plays an important role in the mesocortical dopamine reinforcement system. Repeated cocaine administration causes changes in frontal cortex gene expression that may lead to changes in the behaviors subserved by this brain region. Rats treated non-contingently with a binge model of cocaine (45 mg/kg/day, i.p.) for 14 days were screened for changes in relative mRNA abundance in the frontal cortex by cDNA hybridization arrays. To confirm changes, immunoreactive protein was measured (via protein-specific immunoblots) in a second group of identically treated animals. Protein levels of protein tyrosine kinase 2 (PYK2), activity regulated cytoskeletal protein (ARC), as well as an antigen related to nerve growth factor I-B (NGFI-B-RA) were shown to be significantly induced after cocaine administration. Levels of NGFI-B mRNA were confirmed by real-time RT-PCR to be increased with cocaine administration. These observations are similar to previously reported cocaine-responsive changes in gene expression but novel to the frontal cortex. This study also validates the use of hybridization arrays for screening of neuronal gene expression changes and the utility of relative protein quantification as a post-hoc confirmation tool. PMID- 12117547 TI - Hypoxia-induced cell death and changes in hypoxia-inducible factor-1 activity in PC12 cells upon exposure to nerve growth factor. AB - The transcription factor hypoxia-inducible factor-1 (HIF-1) strongly contributes to the expression of adaptive genes under hypoxic conditions. In addition, HIF-1 has been implicated in the regulation of delayed neuronal cell death. Suspension grown and adherent PC12 cells treated with NGF were used as an experimental model for studying the relationship between hypoxia-induced cell death and activation of HIF-1. Cell damage was assessed by flow cytometry of double-stained (Annexin V and propidiumiodide) cells, and by analysis of the overall death parameters LDH and mitochondrial dehydrogenase. In parallel, cells were transfected with a control and a three-hypoxia-responsive-elements (HRE)-containing vector and HIF-1 driven luciferase activity was determined. Exposure of NGF-treated PC12 cells to hypoxia resulted in a higher cell death rate when compared to untreated controls. PC12 cells exposed for 2 days to NGF exhibited a decrease of HIF-1 activity up to a factor of ten. This decrease may contribute to the enhanced hypoxia-induced cell death via reduced expression of HIF-1alpha-regulated genes responsible for adaptation to hypoxia, like those for glucose transport proteins and enzymes of the glycolytic chain. The decrease in HIF-1 activity and the increase in hypoxia sensitivity may suggest that NGF act as an hierarchically organized signaling molecule. PMID- 12117548 TI - c-fos mRNA expression associated with PGF(2alpha)-induced nest-building behaviour in female pigs. AB - Domestic pigs, Sus scrofa, build a maternal nest on the day before parturition. A model for nest building has been established in pigs, in which exogenously administered prostaglandin (PG) F(2alpha) may be used to elicit nesting behaviour in cyclic, pseudopregnant and pregnant pigs. The central mechanisms mediating this response are unknown. The present study determined regional brain activity using semi-quantitative analysis of c-fos mRNA, after induction of nest-building behaviour by PGF(2alpha) in Large White pseudopregnant pigs. Oestradiol valerate injections (5 mg/day) were given on days 11-15 of the oestrous cycle to induce pseudopregnancy. The pigs were housed individually in pens (2.8 x 1.7 m) containing straw. On the test day (day 46 or 47 of pseudopregnancy) animals were injected with 3 ml saline (n=5) or 15 mg of PGF(2alpha) (Lutalyse, Upjohn; n=6) intramuscularly. Pigs treated with PGF(2alpha), but not saline, displayed bouts of rooting, pawing and gathering straw, which we interpret as nest building behaviour. The pigs were killed 65 min after treatment, which was 30 min after peak nest building activity, and the brain, uterus and ovaries removed for processing using in situ hybridisation. Saline-treated pigs had elevated levels of c-fos mRNA, compared to background, in the pituitary, corpus luteum and uterus, and a lower, but elevated, level of expression in cerebellum, cortex, hippocampus and olfactory bulb. PGF(2alpha)-treated pigs had significantly higher levels of c-fos mRNA expression than saline-treated pigs in the parvocellular and magnocellular regions of the hypothalamic paraventricular nucleus, the supraoptic nucleus (including the pars dorso-medialis), the neural lobe of the pituitary gland and the cerebellum. PGF(2alpha)-treated pigs also had significantly higher c-fos induction in corpus luteum. These data show that the pattern of c-fos mRNA expression in specific brain areas is different between pigs that show PGF(2alpha)-induced nest building and saline-injected controls. PMID- 12117549 TI - LRP and senile plaques in Alzheimer's disease: colocalization with apolipoprotein E and with activated astrocytes. AB - The low density lipoprotein receptor-related protein (LRP) is a multifunctional receptor which is present on senile plaques in Alzheimer's disease (AD). It is suggested to play an important role in the balance between amyloid beta (Abeta) synthesis and clearance mechanisms. One of its ligands, apolipoprotein E (apoE), is also present on senile plaques and has been implicated as a risk factor for AD, potentially affecting the deposition, fibrillogenesis and clearance of Abeta. Using immunohistochemistry we show that LRP was present only on cored, apoE containing senile plaques, in both PDAPP transgenic mice and human AD brains. We detected strong LRP staining in neurons and in reactive astrocytes, and immunostaining of membrane-bound LRP showed colocalization with fine astrocytic processes surrounding senile plaques. LRP was not present in plaques in young transgenic mice or in plaques of APOE-knockout mice. As LRP ligands associated with Abeta deposits in AD brain may play an important role in inducing levels of LRP in both neurons and astrocytes, our findings support the idea that apoE might be involved in upregulation of LRP (present in fine astrocytic processes) and act as a local scaffolding protein for LRP and Abeta. The upregulation of LRP would allow increased clearance of LRP ligands as well as clearance of Abeta/ApoE complexes. PMID- 12117550 TI - Colocalization of estrogen receptor alpha and NMDA-2D mRNAs in amygdaloid and hypothalamic nuclei of the mouse brain. AB - Interactions between gonadal steroid hormones and glutamatergic neurons participate in limbic and hypothalamic functions. Glutamate receptors are divided into metabotropic and ionotropic receptors. Among ionotropic receptors, N-methyl D-aspartate (NMDA) is involved in a variety of neurophysiological processes. In turn, NMDA receptors are composed of subunits from two families: NR1 and NR2. Recently, molecular studies have shown that the expression of NMDA-2D receptor is regulated by estrogen. Although the expression patterns of NMDA-2D and ERalpha in the rodent brain appear to overlap, it remained to be determined whether or not these two receptors co-exist, in vivo, at the level of single neurons. To test the hypothesis that NMDA-2D and ERalpha messenger ribonucleic acid (mRNA) are co expressed in the same neurons of the adult mouse brain, we used a dual-label in situ hybridization technique. Neuronal populations were identified with digoxigenin-tagged complementary RNA probes for NMDA-2D and 35S-labeled cRNA probes for ERalpha. Our results demonstrate that a majority of the ERalpha positive neurons also express NMDA-2D mRNA. Quantitative examination of the cellular expression in the ventromedial and arcuate nuclei of the hypothalamus (VMH and Arc) showed that 52.5% and 61.5%, respectively, of the neurons endowed with ERalpha mRNA also contain NMDA-2D mRNA. In the amygdala, 51% of ERalpha positive cells also contain NMDA-2D mRNA. These findings provide the first anatomical evidence that ER and NMDA-2D receptors can be found in the same hypothalamic and amygdaloid neurons. Co-expression of ERalpha and NMDA-2D receptors supports the hypothesis of the interactions between glutamate receptors and estrogens in brain regions where estrogens control female reproductive behaviors and neuroendocrine functions. PMID- 12117551 TI - Pycnogenol protects neurons from amyloid-beta peptide-induced apoptosis. AB - Neuronal apoptosis is one of the pathological features of Alzheimer's disease (AD). Morphological pathology reveals that neuronal apoptosis is associated with senile plaques containing amyloid-beta peptide (Abeta) in AD brains. Reactive oxygen species (ROS) has been proposed to be involved in the apoptotic mechanism of Abeta-mediated neurotoxicity. In the present study, using a rat pheochromocytoma (PC12) cell line, we investigated the effect of Pycnogenol (PYC), a potent antioxidant and ROS scavenger, on Abeta(25-35)-induced apoptosis and ROS generation. We used vitamin E, a known antioxidant agent, to verify the effect of PYC. Abeta(25-35)-induced apoptosis in PC12 cells was demonstrated by: (1) a dose-dependent loss of cell viability; (2) a time- and dose-dependent increase in the apoptotic cells; (3) an induction of DNA fragmentation; and (4) an increase in caspase-3 activity and cleavage of poly (ADP-ribose) polymerase (PARP). Our data showed that a significant increase in ROS formation preceded apoptotic events after PC12 cells were exposed to Abeta(25-35). We further found that PYC not only suppressed the generation of ROS but also attenuated caspase-3 activation, DNA fragmentation, PARP cleavage, and eventually protected against Abeta-induced apoptosis. Vitamin E also suppressed cell death and caspase-3 activation induced by Abeta(25-35). Taken together, these results suggest that ROS may be involved in Abeta-induced apoptosis in PC12 cells. They further suggest that PYC can reduce apoptosis, possibly by decreasing free radical generation in PC12 cells. PMID- 12117552 TI - Altered expression and phosphorylation of N-methyl-D-aspartate receptors in piglet striatum after hypoxia-ischemia. AB - The mechanisms for the profound degeneration of striatal neurons after hypoxia ischemia in newborns are not understood. We hypothesized that this striatal neurodegeneration is related to N-methyl-D-aspartate (NMDA) receptor-mediated excitotoxicity. Using a 1-week-old piglet model of hypoxia-ischemia, we evaluated whether the expression and phosphorylation of NMDA receptor subunits in striatum are modified with severity of evolving neuronal injury after hypoxia-ischemia. Protein levels of NR1, phosphorylated NR1 897serine, NR2A and NR2B in striatum were measured by immunoblotting after piglets underwent hypoxic-asphyxic cardiac arrest, cardiopulmonary resuscitation, and recovery for 3, 6, 12 or 24 h. In membrane fractions isolated from total striatum, mean NR1 and NR2A levels did not change significantly with time after hypoxia-ischemia compared to control; however, the levels of both NR1 and phosphorylated NR1 897serine correlated with neuronal injury in putamen, with higher levels associated with greater neuronal injury in individual animals. NR2B levels were increased at 24 h after hypoxia ischemia. Astrocyte expression of NR2B was prominent after hypoxia-ischemia. We conclude that NMDA receptors are changed in striatum after neonatal hypoxia ischemia and that abnormal NMDA receptor potentiation through increased NR1 phosphorylation may participate in the mechanisms of striatal neuron degeneration after hypoxia-ischemia. PMID- 12117553 TI - CART promoter CRE site binds phosphorylated CREB. AB - It has been shown previously that: CART (cocaine- and amphetamine-regulated transcript) mRNA is tightly regulated in brain; protein kinase A (PKA) is involved in CART expression in GH3 cells; and a cyclic AMP-responsive element (CRE) site is present in the proximal promoter region of the CART gene. Thus, the goal of this study was to test if CRE binding protein (CREB) can bind to the consensus CRE site and if phosphorylation of CREB occurs in GH3 cells under conditions of enhanced CART gene expression. Electromobility shift assays showed that a 27-bp oligonucleotide containing the CART CRE site was indeed bound by nuclear factors. Western blotting showed that incubation of GH3 cells with forskolin, which enhances CART mRNA expression, caused an increase in phosphorylated CREB (P-CREB) levels. Supershift analyses indicated that the CART CRE oligo/protein complex interacted with a P-CREB antibody. Taken together, these data indicate that P-CREB is a likely regulator of CART expression in GH3 cells. PMID- 12117554 TI - Erythropoietin induces changes in gene expression in PC-12 cells. AB - Erythropoietin (EPO) is the primary modulator of red blood cell production. Recently EPO has received considerable attention for its functions outside of hematopoiesis, including its effects in the nervous system where it has been shown to act as a neuroprotectant. To understand the function of EPO in the nervous system and to determine if EPO functions through the same signaling pathways identified in hematopoietic cells, we used cDNA array hybridization and RT-PCR to investigate the changes in gene expression induced by EPO in the neuronal-like PC-12 cell line. PC-12 cells cultured in the presence of EPO (10 U/ml) showed significant changes in gene expression by 3 h with a return to basal expression levels for the vast majority of genes by 24 h. The genes influenced by EPO included genes with known functions in cell proliferation, differentiation and apoptosis. Semi-quantitative RT-PCR confirmed that 24 h pre-treatment with EPO (10 pM) resulted in a 2.5-fold increase in the expression of the anti apoptotic gene bcl(XL) and a 4-fold decrease in the expression of the pro apoptotic gene bak. In addition to supporting the current models of EPO function these results suggest previously unidentified mechanisms by which EPO may function in neurons. PMID- 12117555 TI - Differential regulation of aquaporin-5 and -9 expression in astrocytes by protein kinase A. AB - Aquaporins (AQPs) transport water through the membranes of numerous tissues, but the molecular mechanisms for regulating water balance in brain are unknown. In this study, we investigated the effects of a protein kinase A (PKA) activator on the expression of AQP4, 5 and 9 in cultured rat astrocytes. Treatment of the cells with dbcAMP caused decreases in AQP5 mRNA and protein and increases in AQP9 mRNA and protein in time- and concentration-dependent manners. However, AQP4 mRNA and protein were not changed by treatment with dbcAMP. The dbcAMP-induced effects on AQP5 and AQP9 mRNAs were inhibited by PKA inhibitors. In addition, pretreating the cells with an inhibitor of protein synthesis, cycloheximide, inhibited the increase in AQP9 mRNA induced by dbcAMP, but not the decrease in AQP5 mRNA. These results suggest that signal transduction via PKA may play important roles in regulating the expression of AQP5 and AQP9, and the effect on AQP9 may be mediated by some factors induced by dbcAMP. PMID- 12117556 TI - Psychological stress, but not physical stress, causes increase in diazepam binding inhibitor (DBI) mRNA expression in mouse brains. AB - Effects of conditioned emotional stimuli (CES), which induce psychological stress, on the expression of cerebral diazepam binding inhibitor (DBI) mRNA in mouse were examined using a communication box. Cerebral DBI mRNA expression significantly increased in a time-dependent manner after the application of CES. The maximal enhancement of DBI mRNA expression was observed 2 days after the application of CES, and this increase faded out over 7 days after the treatment. Flunitrazepam (1 mg/kg), an agonist for central benzodiazepine (BZD) receptors, completely abolished the CES-induced elevation of cerebral DBI contents and its mRNA expressions. These results indicate that cerebral DBI is enhanced by psychological stress, which is regulated by central BZD receptors. PMID- 12117557 TI - Role of hepatic lipase and scavenger receptor BI in clearing phospholipid/free cholesterol-rich lipoproteins in PLTP-deficient mice. AB - Phospholipid transfer protein knock-out (PLTP0) mice have defective transfer of phospholipids (PL) from triglyceride-rich lipoproteins (TRL) into high-density lipoproteins (HDL). In this study, we examined the role of diet, hepatic lipase (HL) and scavenger receptor BI (SRBI) in determining the accumulation of excess PL and free cholesterol (FC, "surface remnants") in plasma of PLTP0 mice. PL and FC accumulated in the very low-density lipoprotein (VLDL)-LDL region of PLTP0 mice on a highly saturated, coconut oil-based diet, but not on chow or milk-fat based Western diets. Accumulation of PL and FC was dramatically increased in PLTP0/HL0 mice, compared to PLTP0 mice, but only on the coconut oil diet. Turnover studies indicated that the coconut oil diet was associated with delayed catabolism of PL of PL/FC-rich particles. Incubation of these particles with primary hepatocytes in the presence of SRBI neutralizing antibody indicated that SRBI was primarily responsible for removal of FC and PL on the Western diet. In hepatocytes of coconut oil-fed mice, removal of FC and PL from these particles by SRBI was markedly reduced, even though SRBI protein expression levels were unchanged. These studies indicate that HL and SRBI both have major role in the clearance of PL and FC of surface remnants in PLTP0 mice. SRBI appears to be dysfunctional in coconut oil diet-fed animals, possibly related to changes in hepatocyte membrane fatty acid composition. PMID- 12117558 TI - Susceptibility to murine cholesterol gallstone formation is not affected by partial disruption of the HDL receptor SR-BI. AB - High density lipoprotein (HDL) promotes reverse cholesterol transport from peripheral tissues to the liver where its cholesterol is secreted preferentially into bile. The scavenger receptor class B type I (SR-BI) is believed to play a pivotal role in unloading HDL cholesterol and its ester to hepatocytes. Here, using male SR-BI "att" mice with a dysfunctional mutation in the Sr-b1 promoter, we studied whether approximately 50% of normal SR-BI expression influences gallstone susceptibility in these mice fed a lithogenic diet containing 1% cholesterol, 0.5% cholic acid and 15% butterfat. Our results showed that the disruption of SR-BI expression reduced cholesterol secretion by 37% in the chow fed state and 10% on the lithogenic diet, and while delaying incidence slightly, did not influence cumulative susceptibility to cholesterol gallstones. The lithogenic diet induced marked increases in biliary cholesterol and phospholipid secretion rates but not of bile salts. Basal expression of hepatic SR-BI protein was dissimilar in both wild-type and SR-BI mice, and remained unaltered in response to the lithogenic diet. By two independent dual isotope methods, intestinal cholesterol absorption was unimpaired by attenuation of the SR-BI which also displays low-density expression on small intestinal enterocytes. We conclude that although HDL cholesterol is a principal source of biliary cholesterol in the basal state, uptake of cholesterol from chylomicron remnants appears to be the major contributor to biliary cholesterol hypersecretion during diet-induced cholelithogenesis in the mouse. PMID- 12117559 TI - Use of vitamin D(4) analogs to investigate differences in hepatic and target cell metabolism of vitamins D(2) and D(3). AB - In this study, we used molecules with either of the structural differences in the side chains of vitamin D(2) and vitamin D(3) to investigate which feature is responsible for the significant differences in their respective metabolism, pharmacokinetics and toxicity. We used two cell model systems-HepG2 and HPK1A-ras to study hepatic and target cell metabolism, respectively. Studies with HepG2 revealed that the pattern of 24- and 26-hydroxylation of the side chain reported for 1alpha-hydroxyvitamin D(2) (1alpha-OH-D(2)) but not for 1alpha-OH-D(3) is also observed in both 1alpha-OH-D(4) and Delta(22)-1alpha-OH-D(3) metabolism. This suggests that the structural feature responsible for targeting the enzyme to the C24 or C26 site could be either the C24 methyl group or the 22-23 double bond. In HPK1A-ras cells, the pattern of metabolism observed for the 24 methylated derivative, 1alpha,25-(OH)(2)D(4), was the same pattern of multiple hydroxylations at C24, C26 and C28 seen for vitamin D(2) compounds without evidence of side chain cleavage observed for vitamin D(3) derivatives, suggesting that the C24 methyl group plays a major role in this difference in target cell metabolism of D(2) and D(3) compounds. Novel vitamin D(4) compounds were tested and found to be active in a variety of in vitro biological assays. We conclude that vitamin D(4) analogs and their metabolites offer valuable insights into vitamin D analog design, metabolic enzymes and maybe useful clinically. PMID- 12117560 TI - Pro-oxidant and antioxidant potential of catecholestrogens against ferrylmyoglobin-induced oxidative stress. AB - Ferryl heme proteins may play a major role in vivo under certain pathological conditions. Catecholestrogens, the estradiol-derived metabolites, can act either as antioxidants or pro-oxidants in iron-dependent systems. The aim of the present work was (1) to determine the effects of ferrylmyoglobin on hepatocyte cytotoxicity, and (2) to assess the pro/antioxidant potential of a series of estrogens (phenolic, catecholic and stilbene-derived) against ferrylmyoglobin induced lipid peroxidation in rat hepatocytes. Cells were exposed to metmyoglobin plus hydrogen peroxide to form ferrylmyoglobin in the presence of the transition metal chelator diethylentriaminepentaacetic acid. Results showed that ferrylmyoglobin induced an initial oxidative stress, mainly reflected in an early lipid peroxidation and further decrease in GSH and ATP. However, cells gradually adapted to this situation, by recovering the endogenous ATP and GSH levels at longer incubation times. Phenolic and stilbene-derived estrogens inhibited ferrylmyoglobin-induced lipid peroxidation to different degrees: diethylstilbestrol>estradiol>resveratrol. Catecholestrogens at concentrations higher than 1 microM also inhibited lipid peroxidation with similar efficacy. The ability of estrogens to reduce ferrylmyoglobin to metmyoglobin may account for their antioxidant activity. In contrast, physiological concentrations (100 pM-100 nM) of the catecholestrogens exerted pro-oxidant activities, 4-hydroxyestradiol being more potent than 2-hydroxyestradiol. The implications of these interactions should be considered in situations where local myoglobin or hemoglobin microbleeding takes place. PMID- 12117561 TI - On-line EPR study of free radicals induced by peroxidase/H(2)O(2) in human low density lipoprotein. AB - The aim of this study was to use direct electron paramagnetic resonance (EPR) spectroscopy at 37 degrees C and spin trapping techniques to study radical species formed during horseradish peroxidase/H(2)O(2)-initiated low-density lipoprotein (LDL) oxidation. Using direct EPR, we obtained evidence for the formation not only of the alpha-tocopheroxyl radical but also of a protein radical(s), assigned to a tyrosyl radical(s) of apolipoprotein B-100 (apo B-100). Spin trapping with 2-methyl-2-nitrosopropane revealed (i) the formation of a mobile adduct with beta-hydrogen coupling assigned to a lipid radical and (ii) a partially immobilised adduct detected in LDL as well as in apo B-100, assigned after proteolytic digestion to the trapping of a radical centred on a tertiary carbon atom of an aromatic residue, probably tyrosine. Our results support the hypothesis that radicals are initiators of the oxidative process, and show that their formation is an early event in peroxidase-mediated oxidation. We also tested the effects of resveratrol (RSV), a polyphenolic antioxidant present in red wine. Our data indicate that 1-10 microM RSV is able to accelerate alpha tocopherol consumption, conjugated dienes formation and the decay kinetics of LDL centred radicals. Since phenols are substrates for peroxidases, this result may be ascribed to a RSV-mediated catalysis of peroxidase activity. PMID- 12117562 TI - Phospholipase C inhibitors and prostaglandins differentially regulate phosphatidylcholine synthesis in rat renal papilla. Evidence of compartmental regulation of CTP:phosphocholine cytidylyltransferase and CDP-choline:1,2 diacylglycerol cholinephosphotransferase. AB - Phosphatidylcholine (PC) is the most abundant phospholipid in mammalian cell membranes. Several lines of evidence support that PC homeostasis is preserved by the equilibrium between PC biosynthetic enzymes and phospholipases catabolic activities. We have previously shown that papillary synthesis of PC depends on prostaglandins (PGs) that modulate biosynthetic enzymes. In papillary tissue, under bradikynin stimulus, arachidonic acid (AA) mobilization (the substrate for PG synthesis) requires a previous phospholipase C (PLC) activation. Thus, in the present work, we study the possible involvement of PLC in PC biosynthesis and its relationship with PG biosynthetic pathway on the maintenance of phospholipid renewal in papillary membranes; we also evaluated the relevance of CDP-choline pathway enzymes compartmentalization. To this end, neomycin, U-73122 and dibutiryl cyclic AMP, reported as PLC inhibitors, were used to study PC synthesis in rat renal papilla. All the PLC inhibitors assayed impaired PC synthesis. PG synthesis was also blocked by PLC inhibitors without affecting cyclooxygenase activity, indicating a metabolic connection between both pathways. However, we found that PC biosynthesis decrease in the presence of PLC inhibitors was not a consequence of PG decreased synthesis, suggesting that basal PLC activity and PGs exert their effect on different targets of PC biosynthetic pathway. The study of PC biosynthetic enzymes showed that PLC inhibitors affect CTP:phosphocholine cytidylyltransferase (CCT) activity while PGD(2) operates on CDP-choline:1,2 diacylglycerol cholinephosphotransferase (CPT), both activities associated to papillary enriched-nuclei fraction. The present results suggest that renal papillary PC synthesis is a highly regulated process under basal conditions. Such regulation might occur at least at two different levels of the CDP-choline pathway: on the one hand, PLC operates on CCT activity; on the other, while PGs regulate CPT activity. PMID- 12117563 TI - Partitioning of polyunsaturated fatty acid oxidation between mitochondria and peroxisomes in isolated rat hepatocytes studied by HPLC separation of oxidation products. AB - The extent of mitochondrial and peroxisomal contribution to beta-oxidation of 18 , 20- and 24-carbon n-3 and n-6 polyunsaturated fatty acids (PUFAs) in intact rat hepatocytes is not fully clear. In this study, we analyzed radiolabeled acid soluble oxidation products by HPLC to identify mitochondrial and peroxisomal oxidation of 24:5n-3, 18- and 20-carbon n-3 and n-6 PUFAs. Mitochondrial fatty acid oxidation produced high levels of ketone bodies, tricarboxylic acid cycle intermediates and CO(2), while peroxisomal beta-oxidation released acetate. Inhibition of mitochondrial fatty acid oxidation with 2-tetradecylglycidic acid (TDGA), high amounts of [14C]acetate from oxidation of 24:5n-3, 18- and 20-carbon PUFAs were observed. In the absence of TDGA, high amounts of [14C]-labeled mitochondrial oxidation products were formed from oxidation of 24:5n-3, 18- and 20-carbon PUFAs. With 18:1n-9, high amounts of mitochondrial oxidation products were formed in the absence of TDGA, and TDGA strongly suppressed the oxidation of this fatty acid. Data of this study indicated that a shift in the partitioning from mitochondrial to peroxisomal oxidation differed for each individual fatty acid and is a specific property of 24:5n-3, 18- and 20-carbon n-3 and n-6 PUFAs.[14C]22:6n-3 was detected with [3-14C]24:5n-3, but not with [1-14C]24:5n-3 as the substrate, while [14C]16:0 was detected with [1-14C]24:5n-3, but not with [3-14C]24:5n-3 as the substrate. Furthermore, the amounts of 14CO(2) were similar when cells were incubated with [3-14C]24:5n-3 versus [1-14C]24:5n-3. These findings indicated that the proportion of 24:5n-3 oxidized in mitochondria was high, and that 24:5n-3 and 24:6n-3 were mostly beta-oxidized only one cycle in peroxisomes. PMID- 12117564 TI - Expression and characterization of Ca(2+)-independent lipase from Bacillus pumilus B26. AB - A lipase-producing Bacillus pumilus strain (B26) was isolated from a soil sample collected in Korea. The cloned gene showed that the lipase B26 composed of a 34 amino-acid signal sequence and a 181-amino-acid mature part corresponding to a molecular mass (M(r)) of 19,225. Based on the M(r) and the protein sequence, the lipase B26 belongs to the lipase family I.4. The optimum temperature and pH of the purified enzyme were 35 degrees C and 8.5, respectively. The lipase B26 showed a 'Ca(2+)-independent thermostability and catalytic activity'. These are novel properties observed for the first time in lipase B26 among all bacterial lipases and correspond with the suggestion that this enzyme had no Ca(2+)-binding motif around the catalytic His156 residue. This enzyme seems to be a true lipase based on the experimental results that it could hydrolyze various long-chain triglycerides (C(14)-C(18)) and triolein (C(18:1)) and that it showed a typical interfacial activation mechanism toward both tripropionin and p-nitrophenyl butyrate. PMID- 12117565 TI - Different interactions of egg-yolk phosphatidylcholine and sphingomyelin with detergent bile salts. AB - To examine physical-chemical aspects of bile salt-phospholipid interactions that could contribute to preferential phosphatidylcholine (PC) secretion into bile, we have compared transitions between vesicles and micelles in model systems containing taurocholate (TC) and either egg-yolk PC (EYPC), egg-yolk sphingomyelin (EYSM), buttermilk SM (BMSM) or dipalmitoyl PC (DPPC). Phase transitions from micelles to vesicles were observed at 4-fold dilution of serially diluted EYPC/TC systems, but not earlier than at 16-fold dilution of SM/TC or DPPC/TC systems, indicating lower concentrations of the detergent required for micellization in the case of SM or DPPC. Cryo-transmission electron microscopy of phase transitions initiated by addition of TC to phospholipid vesicles revealed extremely long SM-containing intermediate structures, but shorter EYPC-containing intermediate structures. Again, larger amounts of bile salt were required to induce phase transitions in the case of EYPC compared to SM. Sizes of TC-phospholipid micelles increased progressively upon increasing phospholipid contents in the rank order: DPPC-TCTC/TUDC 70%/30%>TC 100%>TC/TDC 70%/30%>TDC 100%) and decreased with increasing phospholipid contents, lower TLCo and lower cholesterol saturation index (CSI). In contrast, in the presence of vesicles (three-phase zone), crystallization decreased at increasing bile salt hydrophilicity, with concomitant increased vesicular cholesterol solubilization. CONCLUSIONS: Presence of vesicular phases is a prerequisite for inhibition of cholesterol crystallization by tauroursodeoxycholate. PMID- 12117567 TI - The atypical interaction of peroxisome proliferator-activated receptor alpha with liver X receptor alpha antagonizes the stimulatory effect of their respective ligands on the murine cholesterol 7alpha-hydroxylase gene promoter. AB - Cholesterol 7alpha-hydroxylase (cyp7a) mediates cholesterol elimination in the liver by catalyzing the first and rate-limiting step in the conversion of cholesterol into bile acids. Peroxisome proliferator-activated receptor alpha (PPARalpha; NR1C1) and liver X receptor alpha (LXRalpha; NR1H3) are two nuclear receptors that stimulate the murine Cyp7a1 gene. Here we report that co expression of PPARalpha and LXRalpha in hepatoma cells abolishes the stimulation of Cyp7a1 gene promoter in response to their respective agonists. PPARalpha and LXRalpha form an atypical heterodimer that binds to two directly adjacent hexameric sequences localized within overlapping PPARalpha and LXRalpha response elements (termed Site I), antagonizing the interaction of PPARalpha:retinoid X receptor alpha (RXRalpha) or RXRalpha:LXRalpha with the Cyp7a1 gene promoter. Mutations within either hexameric sequences that specifically abolished LXRalpha:PPARalpha heterodimer binding to the murine Cyp7a1 Site I also relieved promoter inhibition. The LXRalpha:PPARalpha heterodimer may be important in coordinating the expression of genes that encode proteins involved in metabolism of fats and cholesterol. PMID- 12117568 TI - Characterization of a new endogenous vitamin A metabolite. AB - Here, we describe the discovery of a new major endogenous vitamin A metabolite with particularly high hepatic concentrations. This metabolite was isolated from mouse livers and was characterized as 9-cis-4-oxo-13,14-dihydro-retinoic acid (RA) based on mass spectral, ultraviolet, and nuclear magnetic resonance analyses. It was also detected in one human liver. To gain further insight into endogenous retinoid metabolism, mice were fed over a period of 14 days ad libitum with diets enriched with different amounts of retinyl palmitate [15,000, 45,000 or 150,000 international units (IU)/kg diet]. Higher retinyl palmitate amounts in the diet resulted surprisingly in a dose-dependent decrease in all-trans-RA levels in serum, kidney, and brain, whereas levels of 9-cis-4-oxo-13,14-dihydro RA, retinol, and retinyl esters were dose-dependently elevated in serum, kidney, and liver. 13-cis-RA levels could be detected in serum, liver, and kidney, but were unaffected by the dietary vitamin A status. 9-cis-RA levels were below the detection limit of 0.2 ng/ml serum or 0.4 ng/g tissue. This study indicates that the oxidation at C4 of the cyclohexenyl ring, isomerization of the C9/C10 double bond, and reduction of the C13/C14 double bond are major endogenous metabolic pathways of vitamin A. PMID- 12117571 TI - Of human bondage: food craving, obsession, compulsion, and addiction. AB - Is it more than a linguistic accident that the same term, craving, is used to describe intense desires for both foods and for a variety of drugs of abuse? There is strong evidence for common pathways that are affected by most addictive drugs. As the other contributors to this volume will indicate, a strong case can also be made for some shared substrates for food and drug rewards in animals. There has been less explicit work on this topic in humans but many lines of evidence support the common mechanism view: Opioid peptides seem to influence food palatability for humans. There is mounting evidence for comorbidity between drug/alcohol abuse and excessive craving or liking for sweets. Anecdotally, elderly individuals tend to 'age-out' of drug abuse, and the elderly also experience markedly fewer food cravings with age. If we focus on the compulsive aspects of food and drug cravings, there is also evidence for overlap: for example, activity in the orbitofrontal cortex is associated with cocaine and alcohol craving. This area is also implicated in the pathology of obsessive compulsive disorder. Although there is no direct evidence of orbitofrontal involvement in food cravings, there is indirect evidence such as higher than expected co-occurrence of obsessive-compulsive behavior and eating disorders. As a result of bringing together evidence for common substrates for food and for drug rewards, we hope to be able to advance fundamental knowledge of motivational processes and to promote the development of better treatments for drug addiction and for eating disorders. PMID- 12117572 TI - Augmentation of drug reward by chronic food restriction: behavioral evidence and underlying mechanisms. AB - Chronic food restriction and maintenance of low body weight have long been known to increase the self-administration and motor-activating effects of abused drugs. Using a lateral hypothalamic self-stimulation (LHSS) rate-frequency method, it is shown that chronic food restriction augments the rewarding (i.e., threshold lowering) effect of diverse drugs of abuse. Further, the effect is attributed to increased sensitivity of a neural substrate, rather than a change in drug bioavailability or pharmacokinetics, because it is preserved when drugs are injected directly into the lateral cerebral ventricle (intracerebroventricularly). The food restriction regimen that augments drug reward also increases the induction of c-fos, by intracerebroventricular amphetamine, in limbic forebrain dopamine (DA) terminal areas. The possibility of increased DA receptor function is suggested by findings that rewarding and motor activating effects of direct DA receptor agonists are augmented by food restriction, and the augmented behavioral effects of amphetamine are reversed by an otherwise subthreshold dose of D-1 antagonist. Initial studies of DA receptor mediated signal transduction, that are focused on the D-2 receptor, suggest increased functional coupling between receptor and G-protein (i.e., quinpirole stimulated [(35)S]GTPgammaS binding) in dorsal striatum. Unlike behavioral sensitization induced by intermittent stress or psychostimulant treatment, which persist indefinitely following induction, the augmenting effect of food restriction abates within 1 week of restored ad libitum feeding and weight gain. The possible involvement of endocrine hormones and/or 'feeding-related' neuropeptides, whose levels change dynamically with depletion and repletion of adipose stores, is therefore under investigation. Initial tests have been limited to acute treatments aimed at attenuating the effects of hypoinsulinemia, hypoleptinemia and elevated corticosterone levels in food-restricted rats. None of these treatments has attenuated the behavioral effect of food restriction. While a melanocortin receptor agonist has been found to enhance drug reward, melanocortin receptors do not seem to mediate the augmenting effect of food restriction. Continuing investigations of endocrine adiposity signals, 'feeding related' neuropeptides and dopaminergic signal transduction may further elucidate the way in which drugs of abuse exploit mechanisms that mediate survival-related behavior, and help explain the high comorbidity of drug abuse and eating disorders. PMID- 12117573 TI - Opioid modulation of taste hedonics within the ventral striatum. AB - There is a long-standing interest in the role of endogenous opioid peptides in feeding behavior and, in particular, in the modulation of food reward and palatability. Since drugs such as heroin, morphine, alcohol, and cannabinoids, interact with this system, there may be important common neural substrates between food and drug reward with regard to the brain's opioid systems. In this paper, we review the proposed functional role of opioid neurotransmission and mu opiate receptors within the nucleus accumbens and surrounding ventral striatum. Opioid compounds, particularly those selective for the mu receptor, induce a potent increase in food intake, sucrose, salt, saccharin, and ethanol intake. We have explored this phenomenon with regard to macronutrient selection, regional specificity, role of output structures, Fos mapping, analysis of motivational state, and enkephalin gene expression. We hypothesize that opioid-mediated mechanisms within ventral striatal medium spiny neurons mediate the affective or hedonic response to food ('liking' or food 'pleasure'). A further refinement of this hypothesis is that activation of ventral striatal opioids specifically encodes positive affect induced by tasty and/or calorically dense foods (such as sugar and fat), and promotes behaviors associated with this enhanced palatability. It is proposed that this brain mechanism was beneficial in evolutionary development for ensuring the consumption of relatively scarce, high energy food sources. However, in modern times, with unlimited supplies of high calorie food, it has contributed to the present epidemic of obesity. PMID- 12117574 TI - Nucleus accumbens cell firing during goal-directed behaviors for cocaine vs. 'natural' reinforcement. AB - Numerous investigations indicate that the nucleus accumbens (Acb) is an important neural substrate mediating the reinforcing properties of 'natural' rewards (food or water) as well as abused substances. Here, our electrophysiological studies that examined Acb cell firing within seconds of lever press responding for intravenous cocaine vs. water or food reinforcement in rats are reviewed. Initial investigations revealed that a subset of Acb neurons exhibits four types of firing patterns within seconds of the reinforced response for intravenous cocaine during self-administration sessions. Three of those four cell types were also observed during water reinforcement sessions. In a subsequent study, the activity of the same Acb neurons was examined in rats responding on multiple schedules for either two distinct 'natural' reinforcers (water and food), or one of those 'natural' reinforcers and the intravenous self-administration of cocaine. The results showed that the majority of neurons tested exhibited similar, overlapping neuronal firing patterns across the two 'natural' reinforcer conditions. In contrast, the majority of neurons examined displayed differential, nonoverlapping firing patterns relative to operant responding for water (or food) vs. cocaine reinforcement. Additional studies that examined the role of associative factors on Acb cell firing during cocaine self-administration sessions are reviewed. Collectively, these findings illustrate the dynamic nature of Acb cell firing in behaving animals, and provide insight into how Acb neurons process information about goal-directed behaviors for 'natural' reinforcers vs. abused substances. PMID- 12117575 TI - Like drugs for chocolate: separate rewards modulated by common mechanisms? PMID- 12117576 TI - Metabolic sensors: viewing glucosensing neurons from a broader perspective. AB - Glucose is a critical substrate for brain and organ function. Specialized glucosensing neurons, which are involved in the control of energy homeostasis and neuroendocrine function, are located in specific anatomic locations in the brain. Glucose-excited neurons increase their firing rate when ambient glucose levels rise. This glucosensing capacity appears to be regulated by a combination of glucokinase and an ATP-sensitive K(+) (K(ATP)) channel whose activity is regulated by ATP derived from glucose metabolism. Glucose inhibited neurons decrease their firing rate when glucose levels rise, although it is unclear what mechanism is used to control this function. Neuropeptide Y and proopiomelanocortin neurons in the hypothalamic arcuate nucleus are examples of neurons that are capable of sensing both glucose and a host of other peripheral metabolic signals, possibly by their actions on the K(ATP) channel. These metabolic sensing neurons are intimately involved in energy homeostasis, and it is postulated that glucose is only one of several peripheral metabolic signals involved in this process under physiologic conditions. However, when glucose supply is severely limited, glucose appears to assume primacy as a stimulant of glucosensing in order to activate the counterregulatory and ingestive processes necessary to restore the vital supply of glucose. Thus, the role of glucosensing is postulated to be a relative one that is dependent upon the supply of peripheral glucose. PMID- 12117577 TI - Glucose-sensing neurons: are they physiologically relevant? AB - Glucose homeostasis is of paramount concern to the brain since glucose is its primary fuel. Thus, the brain has evolved mechanisms to sense and respond to changes in glucose levels. The efferent aspects of the central nervous system response to hypoglycemia are relatively well understood. In addition, it is accepted that the brain regulates food intake and energy balance. Obesity and diabetes both result from and cause alterations in the central nervous system function. Thus, it is reasonable to hypothesize that the brain also regulates daily glucose homeostasis and energy balance. However, little is known about how the brain actually senses and responds to changes in extracellular glucose. While there are neurons in the brain that change their action potential frequency in response to changes in extracellular glucose, most studies of these neurons have been performed using glucose levels that are outside the physiologic range of extracellular brain glucose. Thus, the physiologic relevance of these glucose sensing neurons is uncertain. However, recent studies show that glucose-sensing neurons do respond to physiologic changes in extracellular glucose. This review will first investigate the data regarding physiologic glucose levels in the brain. The various subtypes of physiologically relevant glucose-sensing neurons will then be discussed. Based on the relative glucose sensitivity of these subtypes of glucose-sensing neurons, possible roles in the regulation of glucose homeostasis are hypothesized. Finally, the question of whether these neurons are only glucose sensors or whether they play a more integrated role in the regulation of energy balance will be considered. PMID- 12117578 TI - Early osmoregulatory signals in the control of water intake and neurohypophyseal hormone secretion. AB - Dehydrated dogs inhibit secretion of vasopressin (VP) within minutes after drinking water, before plasma osmolality (pOsm) diminishes. In recent studies, we found that water ingestion by rats similarly inhibits VP and oxytocin (OT) secretion rapidly, before pOsm is diluted. Adult male rats were infused with 1 M NaCl (2 ml/h iv) for 240 min to stimulate VP and OT secretion. After 220 min of infusion, rats were given water or isotonic saline (IS) to drink for 5 min, and blood samples were taken 5 and 15 min later. Plasma levels of VP (pVP) and OT (pOT) were much lower when rats ingested water instead of IS, even though rats drank comparable amounts of both fluids ( approximately 5.5 ml) and pOsm was not significantly affected in either case. In another study, rats were infused with 1 M NaCl (2 ml/h iv) for 120 min before receiving 4-ml gastric loads of either 0.5 M NaCl (HS) or IS; blood samples taken 25 min later showed that pVP and pOT were much higher when rats were given gastric loads of HS instead of IS, even though pOsm was not significantly altered. Comparable results were obtained when gastric loads of HS or IS were given to rats that had been deprived of drinking water overnight. Other dehydrated rats treated similarly but given access to drinking water consumed much more when they had been given gastric loads of HS instead of IS. Collectively, these and other findings suggest the importance of early signals, perhaps from hepatoportal osmoreceptors or Na(+) receptors, in the control of VP and OT secretion and water intake in rats. PMID- 12117579 TI - Effects of mouth dryness on drinking behavior and beverage acceptability. AB - In humans, the association between mouth dryness and thirst has been examined in a variety of contexts. Typically, drinking behavior produces a concomitant reduction in unpleasant dry mouth sensations. Evidence is reviewed for a mechanism that influences the termination of drinking behavior by metering this change. Drinking behavior causes a progressive increase in parotid saliva flow. Thus, one possibility is that satiety results from a decrease in the reward associated with mouth wetting during a drinking episode. Beverages can differ in their satiating ability. This variability may be related to their mouth-wetting characteristic, and may be reflected in a shift in their acceptability when the mouth becomes dry. Physically drying the mouth appears to increase the acceptability of beverages that are either cold or acidic. It may be significant that two important determinants of mouth wetting are temperature and acidity. Cold or acidic beverages are also likely to be regarded as 'thirst-quenching.' Thus, shifts in acceptability, 'thirst quenching' and satiety may all be related to the mouth-wetting properties of a beverage. The extent to which this coincidence is meaningful warrants further investigation. However, if a common underlying process exists, then this may help to elucidate reasons for voluntary dehydration and aberrant drinking behavior in the elderly. PMID- 12117580 TI - The dorsomedial hypothalamic nucleus and its role in ingestive behavior and body weight regulation: lessons learned from lesioning studies. AB - This review article discusses the well-established role of the dorsomedial hypothalamic nucleus (DMN) in feeding, drinking and body weight (BW) regulation. DMN lesions (L) in both weanling and mature rats of both sexes produce hypophagia, hypodipsia and reduced ponderal and linear growth in the presence of normal body composition. The growth reduction is not due to a deficient secretion of growth hormone, insulin-like growth factor-1, thyroxine, triiodothyronine or insulin. DMNL rats actively defend their lower BW (BW settling point) by becoming either hyper- or hypophagic, depending on the experimental manipulation, thereby defending both lean and fat mass. They also regulate their 24-h caloric intake, but they may overeat during the first hour of refeeding following a fast, possibly due to a reduced ability to monitor blood glucose or to respond to cholecystokinin (CCK). 2-Deoxy-D-glucose (2DG) increases c-fos expression in orexin-A neurons in the DMN, and DMNL eliminated the orexigenic effect of 2DG. DMNL rats on high-fat diets do not get as obese as controls, which may be due to a reduction of DMN neuropeptide Y (NPY). Rats lacking DMN CCK-A receptors are obese and have increased expression of NPY in the DMN, supporting earlier data that CCK may act at the DMN to suppress food intake. Excitotoxin studies showed that loss of DMN cell somata, and not fibers of passage, is important in the development of the DMNL syndrome. The DMN is a site where opioids increase food intake and knife-cut studies have shown that fibers traveling to/from the DMN are important in this response. An interaction of glucose and opioids in DMN may also be involved in the control of food intake. DMN knife cuts interrupting fibers in the posterior and ventral directions additively produce the hypophagia and reduced linear and ponderal growth observed after DMNL. Ventral cuts may interrupt important connections with the arcuate nucleus. Lateral and posterior DMN cuts additively produce the hypodipsic effect seen after DMNL, but DMNL rats respond normally to all water-regulatory challenges, i.e., the hypophagia is not due to a primary hypodipsia. The DMN has been shown to be involved in the rat's feeding response to an imbalanced amino acid diet. These data show the DMN has an important role in many processes that control both food intake and BW regulation. PMID- 12117581 TI - Metabotropic glutamate receptors. PMID- 12117582 TI - Neuronal and glial mGluR5 modulation prevents stretch-induced enhancement of NMDA receptor current. AB - Neuronal stretching in culture has been used to model diffuse axonal injury caused by head trauma, and activation of N-methyl-D-aspartate receptors (NMDARs) has been implicated in the pathophysiology of such injury. Here we report the effects of modulating injury severity and the metabotropic glutamate receptor subtype 5 (mGluR5) on NMDAR activity after stretch injury. Following mild stretch, cortical neurons plated upon a confluent layer of astrocytes (NG) exhibited both increased maximal current (I(NMDA)) and reduction in the voltage dependent Mg2+ block. In contrast, neurons grown without an astrocyte monolayer (PN) only exhibited increased I(NMDA). In NG, surprisingly, pretreatment with either the mGluR5 agonist CHPG or the mGluR5 antagonist MPEP decreased the enhancement of I(NMDA). In contrast, in PN, MPEP similarly limited I(NMDA) changes, but CHPG was without effect. In both culture conditions, MPEP, but not CHPG, limited the stretch-reduced Mg2+ block. Severe stretch had no effect on I(NMDA) or the Mg2+ block in either culture condition, despite a correlation between injury severity and the release of lactose dehydrogenase measured postinjury. Neither CHPG nor MPEP had any direct effects upon the NMDA receptor. We conclude that mGluR5 regulates NMDAR activity during mild stretch injury, but not severe injury, by modulating both the Mg2+ block and I(NMDA). PMID- 12117583 TI - NMDA-induced phosphorylation and regulation of mGluR5. AB - Glutamate regulates neuronal function by acting on ionotropic receptors such as the N-methyl-D-aspartate (NMDA) receptor and metabotropic receptors (mGluRs). We have previously shown that low concentrations of NMDA are able to significantly potentiate mGluR5 responses via activation of a protein phosphatase and reversal of phosphorylation-induced desensitization. While low concentrations of NMDA are able to potentiate mGluR5 responses, higher concentrations of NMDA are actually inhibitory. In this report, we show that NMDA receptors and mGluR5 are highly colocalized in cortical regions. We also show that in voltage-clamp recordings obtained from Xenopus oocytes expressing mGluR5 and NMDA receptors, high concentrations of NMDA (50-100 microM) that elicited large currents (>400 nA) caused an inhibition of mGluR5 currents. Additionally, agonist-induced phosphoinositide hydrolysis presumably mediated by activation of mGluR5, is inhibited by NMDA (30 microM and above). Additional data presented in this report suggest that the inhibitory effect of NMDA is caused by phosphorylation of mGluR5 at protein kinase C (PKC) sites since NMDA induces phosphorylation of the receptor as measured in a back phosphorylation assay. PMID- 12117584 TI - Metabotropic glutamate receptors control gating of spike transmission in the hippocampus area CA1. AB - Signal transmission in the brain is regulated by a number of filters and modulatory systems. In particular, theta rhythm modulates local inhibition of networks and facilitates the induction of synaptic plastic processes. Additionally, the transmission of spikes in the network is controlled by pulse facilitation as a noise filter. Metabotropic glutamate receptors (mGluRs) that are found on interneurons in area CA1 of the hippocampus play a role in the fine tuning of inhibitory circuits and in the transmission of spikes through the network. It was found that the mGluR agonist 1S,3S-1-amino-cyclo-pentyl-1,3 dicarboxylic acid (1S,3S-ACPD) blocked the induction of long-term potentiation (LTP) by high-frequency stimulation (HFS). In addition, theta-patterned stimulation was blocked by the drug. However, learning of spatial tasks in the water maze or radial arm maze was not inhibited by 1S,3S-ACPD. Yet, when stimulating with short bursts phase-locked with theta rhythm at the low inhibition phase, 1S,3S-ACPD did not inhibit the development of LTP. This suggests that burst transmission is not blocked in the network, while high frequency trains are reduced to prevent overexcitation and the transmission of nonphysiologic stimuli patterns. PMID- 12117585 TI - Modulation of DOI-induced increases in cortical BDNF expression by group II mGlu receptors. AB - Previous studies have shown that 5-hydroxytryptamine(2A) (5-HT(2A)) receptor activation induces changes in the pattern of brain-derived neurotrophic factor (BDNF) mRNA expression in the neocortex and hippocampus, and that 5-HT(2A) receptor blockade interferes with the induction of BDNF mRNA by stress. Recent studies have also shown that activation of metabotropic glutamate group II (mGlu2/3) receptors suppresses 5-HT(2A) receptor-stimulated excitatory postsynaptic potentials/currents (EPSP/Cs) in pyramidal neurons in medial prefrontal cortex. Conversely, blockade of mGlu2/3 receptors enhances 5-HT induced EPSCs. The current study examined the effects of the highly selective mGlu2/3 agonist (1S,2S,5R,6S)-2-aminobicyclo[3.1.0]hexane-2,6-dicarboxylate monohydrate (LY354740) and the mGlu2/3 antagonist 2S-2-amino-2-(1S,2S-2 carboxycycloprop-1-yl)-3(xanthy-9-yl)propanoic acid (LY341495) on BDNF mRNA expression in medial prefrontal cortex induced by the hallucinogen and 5 HT(2A/2B/2C) agonist 1-(2,5-dimethoxy-4-iodophenethyl)-2-aminopropane (DOI). LY354740 (0.1-10 mg/kg) dose-dependently suppressed DOI-induced BDNF mRNA levels in medial prefrontal cortex. In contrast, the mGlu2/3 antagonist LY341495 (1 mg/kg) enhanced DOI-induced BDNF mRNA levels. BDNF mRNA expression was not altered by administration of the mGlu agonist or the antagonist alone. These results are discussed with respect to a potential role for group II mGlu agonists in the treatment of depression and schizophrenia. PMID- 12117586 TI - Group II mGlu receptor agonists inhibit behavioural and electrophysiological effects of DOI in mice. AB - It has been suggested that metabotropic glutamate (mGlu) receptor agonists selective for Group II mGlu receptors may have antipsychotic action. Therefore, we studied whether the effects, which could be related to psychotomimetic action of hallucinogenic drugs, are inhibited by Group II mGlu receptor agonists. The selective mGlu2/3 agonists LY354740 and LY379268 inhibited (+/-)1-(2,5-dimethoxy 4-iodophenyl)-2-aminopropane (DOI)-induced head twitches in mice in a dose dependent manner. Furthermore, LY379268 suppressed an increase in the frequency of spontaneous excitatory synaptic potentials induced by bath-applied DOI in layer V pyramidal cells recorded in the murine medial frontal cortex. The data indicate that Group II mGlu receptor agonists may counteract the effects of hallucinogenic drugs. PMID- 12117587 TI - The mGlu2/3 receptor agonist LY379268 blocks the expression of locomotor sensitization by amphetamine. AB - The present experiments assessed the effect of the Group II-specific metabotropic glutamate receptor (mGluR) agonist, LY379268, on the expression of the locomotor sensitization observed following repeated exposure to amphetamine (AMPH). Rats in different groups were administered five injections of AMPH (1 mg/kg ip), one injection every 2-3 days. Two weeks after the last injection, rats were challenged with either AMPH (1 mg/kg ip) or AMPH coinjected with LY379268 (1 mg/kg ip). As expected, AMPH produced levels of locomotion that increased progressively from the first to the fifth injection. This locomotor sensitization was still evident 2 weeks later in rats challenged with AMPH. Rats challenged on this test with AMPH+LY379268, however, showed levels of locomotion similar to those observed following the first AMPH injection. These results indicate that Group II mGluRs can play an important role in the expression of locomotor sensitization by AMPH. The ability of Group II mGluR activation to block the expression of sensitization indicates that it can be targeted as a possible molecular candidate for the development of therapeutic drugs directed at drugs of abuse. PMID- 12117588 TI - Effects of the mGlu2/3 receptor agonist LY379268 on motor activity in phencyclidine-sensitized rats. AB - Previous work has shown that mGlu2/3 receptor agonists such as LY379268 inhibit motor responses to acutely administered phencyclidine (PCP) in rats. However, it has not been determined whether mGlu2/3 receptor agonists will reverse the enhanced effects of repeatedly administered PCP (so called PCP sensitization). In these studies, rats were administered daily PCP and monitored for the number of ambulations, fine movements, time at rest and rears using an automated activity system. At Day 10, when compared the first (Day 1) response, PCP-treated animals showed enhanced responses to all measures tested. Augmentations of these PCP induced behaviors generally peaked between the third and tenth day after PCP administration had begun. Acute administration of LY379268 effectively suppressed PCP-evoked motor behaviors in rats sensitized to PCP. However, daily administrations of LY379268 (for 9 days), along with PCP, did not prevent the expression of the enhanced PCP response on Day 10. Thus, LY379268 administration can suppress PCP responses after either acute or chronic exposure to PCP. However, the underlying plasticity that leads to PCP sensitization was not affected by this treatment. PMID- 12117589 TI - D1 and D2 dopamine receptors contribute to the locomotor response induced by Group II mGluRs activation in the rat nucleus accumbens. AB - Whereas the involvement of ionotropic glutamate receptors (iGluRs) in the functional interaction between glutamate and dopamine (DA) systems in the nucleus accumbens (N. Acc.) is well established, the role of metabotropic glutamate receptors (mGluRs) is less clear. This study was thus aimed to investigate the mechanisms involving DA and glutamate systems via mGluRs in the generation of motor activity in rats. Intra-accumbens infusion of the Group II agonist (2S,3S,4S)-2-(carboxycyclopropyl)glycine (L-CCG-I; 25, 50, 100 nmol) increased locomotor activity, whereas the Group I agonist (S)-3,5-dihydroxyphenylglycine (S 3,5-DHPG) at the same doses had no effect. The effects of L-CCG-I were blocked by a selective Group II mGluRs antagonist (2S,3S,4S)-2-methyl-2 (carboxypropyl)glycine (MCCG; 50 nmol). The locomotor stimulant effect induced by L-CCG-I might be partly DA mediated, as it is abolished by a pretreatment with the DA receptor antagonist haloperidol (0.1 mg/kg ip) and potentiated by D amphetamine systemic injection (0.5 mg/kg sc). Furthermore, selective D1 (SCH 23390; 0.005, 0.01 and 0.02 mg/kg) or D2 (raclopride; 0.05, 0.1 and 0.2 mg/kg) antagonists injected systemically were also effective in decreasing L-CCG-I induced hyperactivity. Taken together, these results demonstrate that stimulation of Group II but not Group I mGluRs contributes to the regulation of motor behavior in the N. Acc. and that this increased activity requires the activation of both D1 and D2 DA receptors. PMID- 12117590 TI - Anxiolytic-like activity of the mGluR5 antagonist MPEP: a comparison with diazepam and buspirone. AB - The selective and systemically active antagonist for the metabotropic glutamate receptor subtype 5 (mGluR5), 2-methyl-6-(phenylethynyl)pyridine (MPEP) was shown to display anxiolytic-like activity in a number of unconditioned assays of stress and anxiety (elevated plus maze, shock probe burying, marble burying, social interaction, and stress-induced hyperthermia) in rodents. In this report, we extend these observations found using unconditioned models of anxiety to include three models of conditioned anxiety, comparing the activity of MPEP to the clinically used anxiolytics, diazepam, and buspirone. MPEP and diazepam, but not buspirone, showed anxiolytic-like activity in the fear-potentiated startle (FPS) model. In a conditioned ultrasonic vocalization (USV) procedure, MPEP, diazepam, and buspirone reduced vocalizations to a similar degree. In the modified Geller Seifter procedure, MPEP, diazepam, and buspirone displayed statistically significant anxiolytic-like activity, increasing the number of punished responses. Thus, these findings confirm and extend previous reports that MPEP exhibits anxiolytic-like activity in rats, and suggests that development of mGluR5 antagonists may provide a novel approach to treating anxiety disorders. PMID- 12117591 TI - The anxiolytic action of mGlu2/3 receptor agonist, LY354740, in the fear potentiated startle model in rats is mechanistically distinct from diazepam. AB - The fear-potentiated startle paradigm has been characterized for drugs that act via ionotropic (NMDA and AMPA/kainate receptor) glutamate receptor mechanisms. Previous studies have shown that the potent systemically active mGlu2/3 receptor agonist, LY354740, effectively reduced the expression of fear-potentiated startle responses in rats. The present study examined the effects of LY354740 in a pre- versus post-fear conditioning paradigm and compared the effects to diazepam. Diazepam (0.3, 0.6, and 1.0 mg/kg ip) attenuated both pre- and post-fear conditioning startle responses in a dose-related manner. In contrast, LY354740 (0.03, 0.3, and 3.0 mg/kg ip) did not disrupt preconditioning startle responses at doses that attenuated post-fear conditioning responses. The benzodiazepine antagonist, flumazenil, at a dose (2 mg/kg sc) that did not alter fear potentiated startle per se, selectively reversed suppression of fear responses to diazepam (0.6 mg/kg ip) while not affecting fear suppression induced by LY354740 (0.3 mg/kg ip). At a dose of 1 mg/kg ip, the mGlu2/3 receptor antagonist, LY341495, did not disrupt fear-enhanced startle per se, but completely reversed the postconditioning anxiolytic effects of LY354740 in this model. This dose of LY341495 had no effect on fear suppression by diazepam. These results demonstrate that fear suppression by diazepam and LY354740 involves different neuronal mechanisms. While diazepam acts via the facilitation of GABAergic transmission, LY354740 induces its actions via the glutamatergic system, specifically mGlu2/3 receptor activation. Furthermore, in contrast to disruption of fear conditioning as well as fear suppression by diazepam, LY354740 had selective effects on fear expression, suggesting anxiolytic actions without the associated memory impairment. PMID- 12117592 TI - Inhibition of mGluR5 blocks hippocampal LTP in vivo and spatial learning in rats. AB - Particular subtypes of metabotropic glutamate receptors (mGluRs) have been shown to be specifically involved in certain types of long-term synaptic plasticity and learning. We examined whether inhibition of mGluR5 by the specific noncompetitive antagonist 2-methyl-6-(phenylethynyl)-pyridine (MPEP) has any functional consequences on long-term potentiation in the dentate gyrus in vivo and on learning of a spatial alternation task. Intracerebroventricular application of 13.8 microg MPEP 30 min before tetanization resulted in a rapid decline of potentiation during the first 7 min and a significantly lower potentiation of the MPEP group as compared to controls. The same dose of the antagonist given 30 min before training of a Y-maze spatial alternation task caused a marked impairment of retention tested 24 h later. In contrast, MPEP had virtually no effects on retention if injected immediately after the training session. Our findings suggest an important function of mGluR5 during the initiation of synaptic plasticity and memory formation. PMID- 12117593 TI - Differential effects of mGluR1 and mGlur5 antagonism on spatial learning in rats. AB - The effects of selective mGluR1 and mGluR5 antagonists on long-term acquisition were tested in a spatial three-choice reward-finding test. Bilateral prelimbic injections of the mGluR1 antagonist, (S)-4-carboxyphenylglycine (4-CPG), before training sessions blocked acquisition of correct performance between sessions. Similar injections given after full training of a control group significantly impaired correct performance without causing a complete block. Pretraining injections (intraperitoneal or intravenous) of the systemically active mGluR5 antagonist, 2-methyl-6-(phenylethynyl)pyridine (MPEP), had no effect on long-term acquisition in the reward-finding task. In an open-field test, bilateral prelimbic pretest application of 4-CPG prevented normal adaptation of spontaneous exploration as seen in control animals. MPEP, on the other hand, had no effect. In conclusion, the results confirmed that mGluR1 is involved in spatial long-term acquisition and suggested an additional role in recall of acquired skills. Furthermore, it was concluded that antagonism of mGluR1 or mGluR5 had different effects both in the appetitive spatial task and in the open-field test. PMID- 12117594 TI - Impairment of contextual fear conditioning in rats by Group I mGluRs: reversal by the nootropic nefiracetam. AB - The blockade of Group I metabotropic glutamate receptors (mGluRs) may be a potential strategy for prevention therapy of neurotoxicity. We here confirm previous reports that systemic application of the Group I antagonist, 1 aminoindan-1,5-dicarboxylic acid (AIDA), causes amnesia in a contextual fear conditioning paradigm in rats. This deficit was fully reversed by long-term pretreatment with the nootropic nefiracetam, which in fact obtained supranormal performance. Our data suggest that application of Group I antagonists to prevent neurotoxicity, combined with nootropic treatment to prevent cognitive deficits, may be a therapeutic strategy for the development of novel antineurotoxic treatments. PMID- 12117595 TI - Antisense oligonucleotide knockdown of mGluR1 alleviates hyperalgesia and allodynia associated with chronic inflammation. AB - Chronic inflammation induced by injection of complete Freund's adjuvant (CFA) into one hindpaw elicits thermal hyperalgesia and mechanical allodynia in the injected paw. Metabotropic glutamate receptors (mGluRs) have been implicated in dorsal horn neuronal nociceptive responses and pain associated with short-term inflammation. The goal of the present study was to assess the role of mGluR1 in the hyperalgesia and allodynia associated with the CFA model of chronic inflammation. Here we show that antisense (AS) oligonucleotide knockdown of spinal mGluR1 attenuates thermal hyperalgesia and mechanical allodynia in rats injected with CFA in one hindpaw. When intrathecal infusion of mGluR1 AS oligonucleotide (50 microg/day) began prior to CFA injection, mechanical allodynia was attenuated from Days 1 to 8 following CFA injection, whereas heat hyperalgesia was attenuated on Day 1 and then from Days 4 to 8. When intrathecal infusion of mGluR1 AS oligonucleotide was begun 2 days after CFA injection, both mechanical allodynia and heat hyperalgesia were attenuated at all time points following the oligonucleotide infusion. Thus, the present data suggest a role for mGluR1 in persistent inflammatory nociception. PMID- 12117596 TI - Antinociceptive effects following intrathecal pretreatment with selective metabotropic glutamate receptor compounds in a rat model of neuropathic pain. AB - In the present study, we examined the effects of intrathecal pretreatment (twice daily injections on postoperative (PO) days 0-3 with the selective Group I (mGluR1a) mGluR antagonist, (RS)-1-aminoindan-1,5-dicarboxylic acid ((RS)-AIDA), the selective Group I (mGluR5a) antagonist, 2-methyl-6-(phenylethynyl)-pyridine (MPEP), the selective Group II mGluR agonist, (2R,4R)-4-aminopyrrolidine-2,4 dicarboxylate ((2R,4R)-APDC) or the selective Group III mGluR agonist, L-2-amino 4-phosphonobutyrate (L-AP4), on mechanical and cold hypersensitivity associated with chronic constriction injury (CCI) of the sciatic nerve in rats. Mechanical and cold sensitivity was assessed prior to surgery (baseline) and then at 4, 8 and 12 days following CCI. Pretreatment with all of the mGluR agents produced reductions in the development of mechanical hypersensitivity. In addition, all the mGluR agents, except MPEP, were effective in reducing the development of cold hypersensitivity. This study demonstrates that spinal Group I mGluR antagonism, and Group II or III mGluR agonism, can effectively decrease the development of mechanical and cold hypersensitivity associated with CCI in rats. In addition, the results can be interpreted to suggest that activation of spinal Group I mGluRs contributes to spinal plasticity leading to the development of neuropathic pain, and that this effect is offset by activation of groups II and III mGluRs. PMID- 12117597 TI - Group II mGluR receptor agonists are effective in persistent and neuropathic pain models in rats. AB - The involvement of Group II metabotropic receptors in acute and persistent pain states was evaluated in several in vivo models of pain with selective and potent Group II metabotropic glutamate (mGlu) 2,3 agonists. LY354740, LY379268 and LY389795 attenuated late-phase paw-licking pain behavior in a dose-dependent manner in the formalin model of persistent pain. Effects occurred in the absence of overt neuromuscular deficits as measured by performance in the rotorod test for ataxia. The effects of LY354740 and LY379268 were also stereoselective. The order of potency of the agonists was LY389795>LY379268>LY354740. The attenuation of licking behavior by LY379268 (3 mg/kg) in the formalin model was reversed by a potent and selective mGlu2,3 receptor antagonist, LY341495 (1 mg/kg). In the L5/L6 spinal nerve ligation model of neuropathic pain in rats, LY379268 significantly reversed mechanical allodynia behavior in a dose-related manner. In contrast, LY379268 had no significant effects on the tail flick test or paw withdrawal test of acute thermal nociceptive function. These results support the involvement of Group II mGlu2,3 receptors in persistent pain mechanisms and suggest the potential utility of selective Group II mGlu agonists for the treatment of persistent pain. PMID- 12117598 TI - Activation of brainstem metabotropic glutamate receptors inhibits spinal nociception in adult rats. AB - In this study, we provide evidence that focal electrical stimulation applied to the rostroventral medulla (RVM) at a high frequency (100 Hz) produced inhibition of the spinal nociceptive tail-flick (TF) reflex in lightly anesthetized adult rats. Chemical activation of metabotropic glutamate (mGlu) receptors by local injection of (+/-)-1-aminocyclopentane-trans-1,3-dicarboxylic acid (tACPD), the mGlu receptor agonist, produced a dose-related inhibition of the TF reflex. Injection of the Group II mGlu receptor agonist (2S,2'R,3'R)-2-(2',3' dicarboxycyclopropyl) glycine (DCGIV) produced strong inhibition, while injection of the Group III mGlu receptor agonist L(+)-2-amino-4-phosphonobutyric acid (L AP4) did not produce any effect. (RS)-alpha-Methyl-4-carboxyphenylglycine (MCPG), a selective mGlu receptor antagonist, but not naloxone reversed the inhibitory effects of DCGIV. Our results provide physiological evidence in vivo that activation of Group II mGlu receptors in the brainstem is antinociceptive and drugs targetting these receptors may help to control pain in humans. PMID- 12117599 TI - Activation of mGlu1 but not mGlu5 metabotropic glutamate receptors contributes to postischemic neuronal injury in vitro and in vivo. AB - In order to investigate the involvement of mGlu1 and mGlu5 metabotropic glutamate receptors in the development of postischemic neuronal death, we examined the effects of selective agonists and antagonists in models of cerebral ischemia in vitro and in vivo. In murine cortical cell cultures and rat organotypic hippocampal slices exposed to oxygen and glucose deprivation (OGD), the mGlu1 antagonists 1-aminoindan-1,5-dicarboxylic acid (AIDA; 300 microM), (S)-(+)-2-(3' carboxybicyclo[1.1.1]pentyl)-glycine (CBPG; 300 microM), 7 hydroxyiminocyclopropan[b]chromen-1a-carboxylic acid ethyl ester (CPCCOEt; 10-30 microM) and (+)-2-methyl-4-carboxyphenylglycine (LY367385; 30-100 microM) reduced neuronal loss when added to the medium during OGD and the subsequent 24-h recovery period. On the contrary, the potent and selective mGlu5 antagonist methyl-6-(phenylethynyl)-pyridine (MPEP; 0.1-1 microM) did not exhibit neuroprotection in any of these in vitro models. Incubation with the nonselective mGlu1 and mGlu5 agonist 3,5-dihydroxyphenylglycine (3,5-DHPG; 300 microM) but not with the mGlu5 agonist (RS)-2-chloro-5-hydroxyphenylglycine (CHPG; 1 mM) enhanced the severity of OGD-induced neuronal damage. In gerbils subjected to global ischemia, intracerebroventricular administration of AIDA (100 nmol two times) or CBPG (300 nmol, two times) afforded consistent protection against CA1 pyramidal cell death, whereas MPEP (10 pmol i.c.v two times and 10 mg/kg i.p two times) failed to reduce postischemic hippocampal damage. Our results suggest that activation of mGlu1 but not mGlu5 receptor contributes to postischemic neuronal injury. PMID- 12117600 TI - Spinal cord mGlu1a receptors: possible target for amyotrophic lateral sclerosis therapy. AB - Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disorder characterized by the progressive loss of motor neurons, whose pathogenesis, probably multifactorial, is thought to involve AMPA/kainate receptor-mediated Ca2+ influx and excitotoxicity. We evaluated the possible involvement of Group I metabotropic glutamate (mGlu) receptors in the control of motor neuron viability. mGlu1a receptor distribution was analyzed in rat and human spinal cord by immunohistochemistry. In both species, the expression of mGlu1a receptor was developmentally regulated and showed a general trend to increase during foetal and postnatal maturation, reaching the maximum level of expression in the dorsal laminae I-II and in motor neurons in adult life. Exposure of spinal cord slices from adult rats to 300 microM kainate for 30 min induced motor neuron death, which was prevented by the Group I mGlu receptor agonist 3-hydroxyphenylglycine (3-HPG; 100 microM). Since motor neurons do not express mGlu5 receptors, mGlu1a receptor activation might be responsible for the observed neuroprotection. mGlu1a immunohistochemistry was conducted on spinal cord autoptic specimens from ALS and control subjects. Surviving motor neurons from ALS spinal cord still revealed the presence of mGlu1a at levels comparable to that from controls. We suggest that mGlu1a receptors may act as suitable targets for ALS experimental therapies. PMID- 12117601 TI - Evaluation of the mGluR2/3 agonist LY379268 in rodent models of Parkinson's disease. AB - The aim of the present studies was to examine the ability of a potent, systemically active, selective Group II mGlu receptor (mGluR2/3) agonist, 1R,4R,5S,6R-2-oxa-4-minobicyclo[3.1.0.]hexane-4,6-dicarboxylate (LY379268) to provide both functional relief and neuroprotection in rodent models of Parkinson's disease (PD). In functional studies, intracerebroventricular administration of LY379268 (1, 5, 10, 20 nmol/2 microl) produced a dose-dependent increase in locomotor activity in the reserpine (5 mg/kg ip)-treated rat. In contrast, systemic administration of LY379268 (0.1, 1, 10 mg/kg ip) did not reverse reserpine-induced akinesia and failed to effect rotational behaviour 1 month after unilateral lesioning of the nigrostriatal tract by 6-hydroxydopamine (6-OHDA; 4 microg infused into the substantia nigra (SN)). In neuroprotective studies, animals were treated with LY379268 (10 mg/kg/day ip) either for 7 days following 6-OHDA injection into the SN (4 microg) or for 21 days following 6-OHDA injection into the striatum (10 microg) before measurement of tyrosine hydroxylase immunoreactivity in the striatum and/or SN as an index of neuroprotection. LY379268 provided some protection against nigral infusion of 6 OHDA and also some functional improvement and correction of dopamine turnover was observed. The compound also provided significant protection in the striatum and some protection in the SN against striatal infusion of 6-OHDA. These data suggest that activation of Group II mGlu receptors can provide some protection in models of PD, while their role in providing functional improvement is less clear. PMID- 12117602 TI - The selective group mGlu2/3 receptor agonist LY379268 suppresses REM sleep and fast EEG in the rat. AB - Studies of ionotropic receptors indicate that glutamate (Glu) neurotransmission plays a role in sleep. Here, we show for the first time that metabotropic 2/3 Glu (mGlu2/3) receptors play an active or permissive role in the control of REM sleep. The potent, selective, and systemically active mGlu2/3 receptor agonist LY379268 was administered systemically in doses of 1.0 and 0.25 mg/kg sc. The drug produced a dose-dependent suppression of rapid eye movement (REM) sleep and fast (10-50 Hz) EEG in non-rapid eye movement (NREM) sleep. The 1.0-mg/kg effect on REM sleep was remarkably powerful: REM sleep was totally suppressed in the 6-h postinjection and reduced by 80% in the next 6 h. NREM duration was unchanged during the REM suppression in spite of the strong and unusual depression of EEG power in fast NREM frequencies. These sleep and EEG effects were unaccompanied by motor or behavioral abnormalities. We hypothesize that the REM and the fast EEG suppression were both caused by a depression of brain arousal levels by LY379268. If correct, depressing arousal by reducing excitatory neurotransmission with an mGlu2/3 receptor agonist produces electrophysiological effects that differ drastically from those produced by depressing arousal by enhancing neural inhibition with GABAergic drugs. This different approach to modifying the excitation/inhibition balance in the brain might yield novel therapeutic actions. PMID- 12117603 TI - Metabotropic and NMDA glutamate receptor interactions with osmotic stimuli in supraoptic neurons. AB - During increases in plasma osmolality, the actions of extrinsic glutamatergic synaptic inputs on magnocellular neuroendocrine cells (MNCs) are thought to combine with intrinsic osmosensitivity of these cells to promote the release of vasopressin (VP). In the present study, changes in intracellular calcium were used as an endpoint to examine putative interactions between osmotic stimuli and NMDA and metabotropic glutamate receptors (mGluRs). Exposure of MNCs to hyperosmotic solutions resulted in a very small, gradual increase in intracellular calcium. NMDA (100-300 microM) combined with osmotic stimulation gave rise to a synergistic increase in intracellular calcium. The broad spectrum mGluR agonist, (1S,3R)-1-aminocyclopentane-1,3-dicarboxylic acid (1S,3R-ACPD) and the type I mGluR agonist, (RS)-3,5-dihydroxyphenylglycine (DHPG) evoked an acute calcium rise followed by a sustained increase. However, when combined with the hyperosmotic stimulus, the calcium responses to 1S,3R-ACPD and DHPG were suppressed. The type II agonist, (2S,2'R,3'R)-2-(2',3' dicarboxycyclopropyl)glycine (DCG-IV), and the type III agonist, (+)-2-amino-4 phosphonobutyric acid (AP-4), facilitated the loss of calcium from the MNCs and were largely unaffected by the osmotic stimulus. These osmotic interactions with NMDA and mGluR function not only help to explain the mechanisms that underlie osmotically mediated changes in MNC function, but also have implications for the impact of hyperosmotic stress in various pathological conditions. PMID- 12117604 TI - MRI measurement of blood-brain barrier permeability following spontaneous reperfusion in the starch microsphere model of ischemia. AB - Quantification of the acute increases in blood-brain barrier (BBB) permeability that occur subsequent to experimental ischemic injury has been limited to single time-point, invasive methodologies. Although permeability can be qualitatively assessed to visualise regional changes during sequential studies on the same animal using contrast-enhanced magnetic resonance imaging (MRI), quantitative information on the magnitude of change is required to compare barrier function during sequential studies on the same animal or between different animals. Recently, improvements in MRI tracer kinetic models and in MR hardware design mean that an estimate of permeability in vivo can now be obtained with acceptable accuracy and precision. We report here the use of such methods to study acute changes following spontaneous reperfusion in an animal model of ischemia. We have obtained estimates of BBB permeability following spontaneous reperfusion, subsequent to forebrain ischemia by unilateral carotid injection of starch microspheres in the rat. T2*-weighted and diffusion-trace imaging were used to monitor the initial reduction in CBF and the time-course of ischemia, respectively. Following reperfusion, an intraveneous bolus of dimeglumine gadopentetate (Gd-DTPA) and horseradish peroxidase (HRP) was given during a continuous acquisition of T1 maps with a 48 s temporal resolution. Permeability maps were constructed using a 4-compartment model; K(trans), the permeability surface area product of the capillary walls was estimated to be 9.2 +/- 0.6 x 10( 4) min(-1) in the cortex. Visualisation of the regional extent of HRP extravasation on histological sections following termination of the experiment demonstrated very little correspondence to the region of Gd-DTPA leakage. Quantitative MRI assessment of BBB permeability following ischemia-reperfusion is consistent with published values obtained by invasive methods. Differences between Gd-DTPA-enhancement and HRP may reflect differences in the molecular size of the tracers. PMID- 12117605 TI - q-Space high b value diffusion MRI of hemi-crush in rat spinal cord: evidence for spontaneous regeneration. AB - The development of the damage following hemi-crush trauma in rat spinal cord was studied ex vivo using high b value (bmax = 1 x 10(7) s cm(-2)) q-space diffusion weighted MRI (DWI) at five days, ten days and six weeks post-trauma. Rat spinal cord trauma, produced by hemi-crush of 15s and 60s duration, was studied. The water signal decay in these diffusion experiments was found to be non mono exponential and was analyzed using the q-space approach. The q-space MRI parameters were compared with T1 and T2 MR images, behavioral tests and histopathological osmium staining. A very good anatomical correlation was found between the q-space MRI parameters and the osmium staining. Interestingly, we found that in the 15s hemi-crush model significant recovery was observed in both the q-space MR images and the osmium staining six weeks post-trauma. However, in the 60s hemi-crush trauma model very little recovery was observed. These results paralleled those obtained from behavioral tests demonstrating that partial spontaneous recovery seems to occur in the 15s hemi-crush spinal cord model, which should be taken in consideration when using it to evaluate new therapies. PMID- 12117606 TI - In vivo diffusion tensor imaging of rat spinal cord at 7 T. AB - In vivo diffusion tensor imaging of normal rat spinal cord was performed using a multi-segmented, blipped EPI sequence at 7 T field strength. At high diffusion weighting, the signal exhibited a non-monoexponential decay that was fitted to a biexponential function, associated with the fast and slow components of diffusion in the cord tissue, using a nonlinear regression analysis along with a constrained optimization procedure. From the measured tensors, the eigenvalues and the maps of invariant scalar measures (fractional anisotropy, relative anisotropy, volume ratio, and trace) were calculated and analyzed statistically. The results were combined to quantitatively characterize the anisotropic properties of the fast and slow diffusions in white- and gray matter of live spinal cords. PMID- 12117607 TI - Intraventricular dispersion and temporal delay of early left ventricular filling after acute myocardial infarction. Assessment by magnetic resonance velocity mapping. AB - This article aims to describe early left ventricular diastolic inflow using magnetic resonance velocity mapping in patients with recent acute myocardial infarction and in normal volunteers. Magnetic resonance velocity mapping was performed in a long axis plane through the hearts of 46 patients with recent, first time acute myocardial infarction and 43 age-matched normal volunteers. The peak velocities at six levels of the early diastolic inflow stream were recorded. A velocity index was calculated as the peak velocity in each position relative to the peak velocity at the mitral leaflet tips. Also, the temporal delay of velocity propagation was computed. Velocity index 4 cm downstream of mitral leaflet tips was lower in the acute myocardial infarction group (0.42 (0.17)) (mean (SD)) compared to controls (0.59 (0.25)) (p < 0.001). Temporal delay in the same position was longer in the acute myocardial infarction group (62 (67) ms) than in controls (32 (39) ms) (p < 0.02). Blood flow patterns in patients after acute myocardial infarction were characterized by increased dispersion of velocities and increased temporal delay of velocity propagation, probably reflecting impaired active left ventricular relaxation. Intraventricular flow measurements constitute a promising new technique for non-invasive assessment of left ventricular diastolic function. PMID- 12117608 TI - The value of single-shot black-blood MR imaging for mapping of the coronary arteries: a comparison of four different orientations during breath-holding and free breathing. AB - The value of ECG-gated single-shot black-blood MR imaging for rapid visualization of the origin and course of the coronary arteries was investigated. The study population included 28 patients with known or suspected cardiac disease. ECG gated single-shot black-blood MR acquisitions were acquired in the transverse, coronal, sagittal and LAO orientations, during free breathing and breath-holding. The origin of the left coronary artery was most frequently visualized in the coronal and LAO orientations and the origin of the right coronary artery was most frequently visualized in the LAO orientation. Overall, no significant difference was found for the visualization of the coronary artery segments and the overall image quality among acquisitions during breath-holding and free breathing. ECG gated single-shot black-blood MR imaging (HASTE) appears to be a time-efficient and robust method for mapping of the entire coronary artery tree, without the need for breath-holding. The LAO orientation provides the most consistent visualization of the origins and major coronary artery segments. PMID- 12117609 TI - In vivo fate of superparamagnetic iron oxides during sepsis. AB - The enzymatic generation of nitric oxide (NO) in vivo has been reported to be modulated by ions, such as copper and iron. Superparamagnetic iron oxide (SPIO) or ferumoxides is a liver-specific magnetic resonance contrast agent that is taken up by the Kupffer cells, where NO is generated by inducible nitric oxide synthase (iNOS). Thus, it is important to evaluate SPIO in vivo under conditions, such as infectious disease, where significant amounts of NO are generated by iNOS. In this study, we monitored the pharmacokinetics of SPIO in the liver of septic-shock mice and rats. A significant decrease in the ferric iron EPR signal was observed during NO generation in septic-shock mice compared with control mice doped with only SPIO. These results were also confirmed in a model reaction system consisting of SPIO and the NO donor, S-nitroso-N-acetyl DL penicillamine (SNAP). We compared NO generation quantitatively in the liver of the septic-shock rats, either in the presence or absence of SPIO, and found that the presence of SPIO did not affect the NO-generating activity of NOS expressed in the liver. T2 weighted MR images of an agarose gel phantom containing different SPIO to NO donor (SNAP) ratios clearly demonstrated that the contrast enhancement by SPIO decreased with increasing NO at constant SPIO levels. The reduced contrast is most probably due to the reduction of ferric to ferrous irons, resulting in a decrease in paramagnetic relaxation of water protons. These results show that SPIO can be a versatile NO-sensitive indicator, especially employing MRI as a powerful tool to 'visualize' sites of NO generation. PMID- 12117610 TI - In vivo temporal EPR imaging of the brain of rats by using two types of blood brain barrier-permeable nitroxide radicals. AB - In vivo temporal EPR imaging was conducted on the brain of rats that received one of two kinds of blood-brain barrier-permeable nitroxide radicals via the tail vein-one is a water-soluble 3-hydroxymethyl-2,2,5,5-tetramethylpyrrolidine-1-oxyl (hydroxymethyl-PROXYL); and the other is a non-water-soluble 3-methoxycarbonyl 2,2,5,5-tetramethylpyrrolidine-1-oxyl (PCAM). From temporal EPR imaging data, temporal changes in the distribution of the nitroxide radical in the cerebral cortex, striatum, and hippocampus in the brain were investigated. It was found that the half-lives of the three parts in the brain of hydroxymethyl-PROXYL are longer and their EPR signal intensities are greater than those of PCAM. PMID- 12117611 TI - Equivalent cross-relaxation rate imaging in the synthetic copolymer gels and invasive ductal carcinomas of the breast. AB - The values of equivalent cross-relaxation rate (ECR) correlated well with [i] water conditions in various copolymer gels and [ii] nature of malignant cells with regard to nuclear dysplasia and mitotic potential in breast carcinomas. The synthetic copolymer gels composed of any two or three monomers among 2 hydroxyethyl methacrylate (HEMA), glycidyl methacrylate (GMA), N-vinyl-2 pyrrolidinone (N-VP), methyl methacrylate (MMA) and benzyl methacrylate (BMA). The ECR measurement was performed by using an off-resonance saturation pulse under conventional field-echo imaging at frequency within +/- 75 ppm apart from the water resonance frequency. The ECR values were readily to determine and non time consuming parameter for cross relaxation rate. The ECR values at the frequency offset by 7-ppm (ECR-7) were divided the sample gels two classes, which must correspond to hydrophilic or hydrophobic ones. The sensitivity in the gels was nearly equivalent to the cross-relaxation rate itself. In the breast carcinomas, the ECR-7 correlates with the nature of malignant cells with regard to nuclear dysplasia and mitotic potential. The ECR-7 is better or more accurate than the STR-7 because the SDNRs between carcinoma and glandular tissue increased by approximately 50% on the ECR-7 compared with the STR-7. Thus the ECR values could be a new parameter for malignancy and cell proliferative activity of the breast carcinomas with non-invasive modalities by magnetic resonance imaging. PMID- 12117612 TI - Real time MRI-ultrasound image guided stereotactic prostate biopsy. AB - To report a technique for target directed transperineal ultrasound guided biopsy using high resolution endorectal MRI images Ultrasound fusion. Two patients presented after external beam irradiation for prostate cancer with a rising PSA. An Endorectal MRI using a 1.5 Tesla scanner was obtained. Subsequently a Transrectal Ultrasound guided biopsy was performed. The Ultrasound probe was fixed to a stepper-stabilizer to provide a reference coordinate system for stereotaxic needle biopsy needle placement. The MRI image set was fused to the Ultrasound images in real time. Abnormal areas determined in the MR images were targeted for biopsy. Recurrent prostate carcinoma was detected pathologically in 3 of 4 stereotactic biopsies. Abnormal areas suspicious for cancer detected on T1 weighted images obtained in a strong field Endorectal MRI scan can be targeted for stereotactic biopsy using Transrectal Ultrasound. This image guide technique may be very useful in directing biopsies. PMID- 12117613 TI - A decidualized endometrial cyst in a pregnant woman: a case observed with a steady-state free precession imaging sequence. AB - Decidual changes of the ectopic endometrial stroma during pregnancy are well known among pathologists and obstetricians. However, they appear very similar to endometrial cysts with malignant transformation when imaged. Balanced fast field echo (BFFE) is a steady-state free precession imaging sequence and its contrast is decided by the T1/T2 ratio. The authors report a case of a decidualized endometrial cyst in which mural nodules were isointense with the nomotopic decidualized endometrium on T1- and T2-weighted images and BFFE. Isointensity with the nomotopic endometrium is an MR characteristic that can differentiate a decidualized endometrial cyst from malignant transformation. BFFE is a good alternative sequence during pregnancy because of its shorter acquisition time and lower radiofrequency absorption. PMID- 12117614 TI - Estrogenic effects of 7alpha-methyl-17alpha-ethynylestradiol: a newly discovered tibolone metabolite. AB - Tibolone is a synthetic steroid that is prescribed to postmenopausal women for relief of climacteric symptoms and prevention of osteoporosis. It has been reported to be metabolized in a tissue-selective manner to three steroids that collectively have weak estrogenic, progestogenic, and androgenic activities. Recently, a new tibolone metabolite, 7alpha-methyl-17alpha-ethynyl-17beta estradiol (7alpha-Me-EE2), was identified in women. In this report, we describe the pre-clinical estrogenic activities of this metabolite and compare these effects to those obtained with 17alpha-ethynyl-17beta-estradiol (EE2) and 17beta estradiol (E2). In an in vitro ligand-binding assay, 7alpha-Me-EE2 bound to both human estrogen receptor (ER)-alpha and -beta with IC(50)'s of 1.2 and 3.0 nM, respectively. Using MCF-7 human breast cancer cells that express high levels of ER-alpha, 7alpha-Me-EE2 transactivated an estrogen response element (ERE)-tk luciferase reporter gene construct with an EC(50) of 0.021 nM. Likewise, 7alpha Me-EE2 stimulated MCF-7 breast cancer cell proliferation with an EC(50) of 0.002 nM. In immature female rats, subcutaneous (s.c.) administration of 7alpha-Me-EE2 stimulated uterine wet weight gain with an ED(50) of 0.2 microg/kg. Moreover, 7alpha-Me-EE2 induced uterine complement component C3 gene expression, an estrogenic marker of epithelial cell stimulation, with an ED(50) of 0.5 microg/kg. When compared to EE2 and E2, 7alpha-Me-EE2 exhibited equivalent or greater potencies and efficacies in these assays. In summary, these results indicate that 7alpha-Me-EE2 is a very potent estrogen. This steroid appears to be the most potent estrogenic metabolite of tibolone identified to date, and additional studies are, therefore, warranted regarding the role of this metabolite in the biological actions of the drug. PMID- 12117615 TI - Chemoselective construction of novel steroid derivatives. AB - Alpha-halo-alpha-heteroarylalkyllithiums, generated by deprotonation of the corresponding halides, when added promptly to steroids with C=O or C=NR groups, lead to epoxides and aziridines. The reactions are regio- and stereoselective; in fact, in the presence of more than one C=O group, the oxido or aziridino functions are formed uniquely at the C=O of C-17 (or C-20 depending on its position in the starting molecule), and the C-20(R) stereoisomer is often the only product isolated. Protection of the hydroxyl group present on several considered steroids was required, and it was accomplished through derivatization in acetyl, ether, or lactone. PMID- 12117616 TI - Influence of polyhydroxysteroids on [Ca(2+)](i). AB - Recently, we have shown that two biologically active, disulfated polyhydroxysteroids from the Pacific brittle star Ophiopholis aculeata stimulate Ca(2+) influx into different cell types. In the present study, 45Ca(2+) and two fluorescent calcium probes, quin-2/AM and fura-2/AM, were employed to investigate the course and amplitude of calcium signals induced in different mouse cells using an radio-isotope, spectrofluorimetry, and microcytofluorimetry techniques. The cytotoxic and hemolytic effects were not observed for both steroids at the wide range of concentrations. Steroids did not influence [3H]-uridine incorporation in a variety of cells. The investigated steroids stimulated a rapid increase in cytosolic Ca(2+) content in Ehrlich mouse carcinoma cells, mouse spleen lymphocytes, and mouse peritoneal macrophages in the concentration range 1 100 microg/ml on a dose-dependent basis. Blockers of L-type calcium channels, such as verapamil, diltiazem, nifedipine (1 x 10(-7)M), and 1mM EGTA, inhibited this process and reduced the response of cells to steroid application. The stimulatory effect of steroids on human fibroblast proliferation and mouse macrophage lysosome activity was observed also. It is suggested that the investigated compounds may act as Ca(2+)-agonists and increase Ca(2+)-transport across cell membranes. PMID- 12117617 TI - A comparative study of the conversion of 7-hydroxycholesterol in rabbit, guinea pig, rat, hamster, and chicken. AB - The metabolism of epimeric 7-hydroxycholesterol was studied in vitro. 7Alpha hydroxycholesterol or 7beta-hydroxycholesterol were incubated with rabbit, guinea pig, rat, hamster, and chicken microsomal suspensions and then extracted and analyzed using high-performance liquid chromatography (HPLC). 7Alpha-hydroxy-4 cholesten-3-one was the main product from 7alpha-hydroxycholesterol in the rabbit, guinea pig, and rat. A considerable amount of 7-ketocholesterol was also produced in the hamster and chicken. In all vertebrates, 7beta-hydroxycholesterol was converted only to 7-ketocholesterol in all vertebrates. 7Beta-hydroxy-4 cholesten-3-one was not detected. Reduction of 7-ketocholesterol was also studied in the rat and hamster. Whereas 7-ketocholesterol was converted to 7beta hydroxycholesterol in the rat, it was converted to both 7alpha- and 7beta hydroxycholesterol in the hamster. These results suggest that 7alpha hydroxycholesterol is converted not only to 7alpha-hydroxy-4-cholesten-3-one but also to 7-ketocholesterol in the hamster and chicken. 7Beta-hydroxycholesterol was converted to 7-ketocholesterol in all vertebrates tested. The interconversion between 7alpha- and 7beta-hydroxycholesterol via 7-ketocholesterol was observed in the hamster in this in vitro study. PMID- 12117619 TI - Effects of glucocorticoids on generation of reactive oxygen species in platelets. AB - Since prednisolone and dexamethasone are known as potent anti-inflammatory agents, the effects of prednisolone and dexamethasone on production of intracellular reactive oxygen species (ROS) were investigated in human platelets. Platelet ROS were measured using the intracellular fluorescent dye dichlorofluorescein diacetate after activation of protein kinase C by phorbol-12 myristate-13-acetate (PMA) or 1-oleoyl-2-acetyl-sn-glycerol (OAG). NAD(P)H oxidase activity was measured photometrically. PMA and OAG significantly increased ROS in platelets (P<0.001). Prednisolone or dexamethasone concentration dependently reduced the PMA-induced ROS production. The PMA-induced ROS increase was significantly reduced in the presence of 10 micromol/l prednisolone to 9+/-1% (n=31; P<0.001) or in the presence of 10 micromol/l dexamethasone to 9+/-1% (n=24; P<0.001). The inhibitory effect of prednisolone or dexamethasone could also be observed in the presence of the glucocorticoid receptor inhibitor, mifepristone (RU486). Administration of testosterone or aldosterone did not significantly reduce PMA-induced ROS increase. Prednisolone had no effect on platelet NAD(P)H oxidase activity. The inhibition of oxidative phosphorylation by sodium azide reduced platelets ROS to 8+/-1% (n=35). It is concluded that glucocorticoids, prednisolone and dexamethasone, directly inhibit production of intracellular ROS. This effect may contribute to the anti-inflammatory actions of these agents. PMID- 12117618 TI - Microwave-assisted Stille-coupling of steroidal substrates. AB - Steroidal dienes were synthesised by Stille-coupling of the corresponding alkenyl iodides with vinyltributyltin under microwave irradiation in a domestic microwave oven in drastically reduced reaction times. Rate acceleration was observed also in the one-pot Stille-coupling-Diels-Alder reaction of 17-iodo-5alpha-androst-16 ene. Stereoselectivity of cycloaddition was slightly improved with diethyl maleate as the dienophile, compared to that achieved with thermal heating. PMID- 12117620 TI - Inhibition of 3beta-hydroxysteroid dehydrogenase-isomerase in mouse adrenal cells: a direct effect of testosterone. AB - Gonadal steroids modulate adrenal gland size and function in a variety of species, and our previous studies demonstrate that circulating androgens suppress 3beta-hydroxysteroid dehydrogenase-isomerase (3betaHSD) activity in the adrenal cortex of male mice. The present study tests the hypothesis that androgens have a direct, receptor-mediated inhibitory effect on adrenal 3betaHSD. Treatment of cultured adrenal cells from C57BL/6J and C3H/HeJ mice with 0.02-2.0 microM testosterone for 7 days significantly reduces 3betaHSD activity in cells from both strains. However, treatment for 3 days reduces 3betaHSD activity in the adrenal cells from C3H/HeJ, but not C57BL/6J mice. The decreases in 3betaHSD activity in response to testosterone treatment is reflected in decreases in the amount of 3betaHSD immunoreactive protein, such that extended treatment decreases 3betaHSD immunoreactive protein in adrenal cells from both strains, but short term treatment only decreases 3betaHSD immunoreactive protein in adrenal cells from C3H/HeJ mice. Thus, there appears to be a temporal difference between strains in the effect of the testosterone on 3betaHSD activity and immunoreactive protein. Treatment of the adrenal cells with androgen agonists and an antagonist indicate that the effect of testosterone is androgen receptor mediated. The effect of testosterone appears to be specific for 3betaHSD, since none of the treatments alter P450(scc) in cells from either strain. Testosterone treatment also causes a decrease in the amount of 3betaHSD mRNA. However, in contrast to the effect on activity and immunoreactive protein, there is no strain-related temporal difference because testosterone decreases 3betaHSD mRNA within 24h in adrenal cells from both strains. These results indicate that testosterone can act directly on the adrenal gland to decrease 3betaHSD activity, immunoreactive protein, and mRNA content in mouse adrenal glands, and thus contribute to the sex difference in adrenal function observed in many species. PMID- 12117621 TI - Single-chain Fv fragments derived from an anti-11-deoxycortisol antibody. Affinity, specificity, and idiotype analysis. AB - Single-chain Fv fragments (scFvs) against a corticosteroid, 11-deoxycortisol (11 DC), have been generated as a template antibody fragment from which a comprehensive mutated antibody library containing various anti-steroid antibodies could be constructed. The cDNAs encoding variable heavy (V(H)) and light (V(L)) domains of a mouse anti-11-DC antibody (CET-M8), were amplified by RT-PCR, combined via a common linker to construct the sequence of 5'-V(H)-(Gly(4)Ser)(3) V(L)-3', and cloned into a phagemid vector, pEXmide 5. The phage clones exhibiting binding activity to 11-DC were isolated after single panning against a hapten-immobilizing immunotube. The scFv gene in one of these clones was reamplified to introduce the ochre codons, and then expressed in the bacterial periplasm as the soluble antibody fragment. Two different scFvs (#6 and #12) were cloned, whose binding characteristics were examined by a radioimmunoassay using a tritium-labeled 11-DC. Both of them showed high affinity (K(a)=1.3x10(10)M(-1)) and practical specificity (cross-reactivity: cortisol, <0.2%; cortisone, <0.3%) to 11-DC, and furthermore, strong reactivity with an anti-idiotype antibody which recognizes the paratope of CET-M8. These results suggest that the present scFvs retain the three-dimensional structure of the paratope of the original monoclonal antibody. PMID- 12117622 TI - The search for determinants of chronic depression: a review of six factors. AB - While strides have been made in the classification, assessment and identification of chronic depression, there remains a limited understanding of the factors underlying chronicity. This review focuses on six putative determinants of chronic depression: developmental factors, personality and personality disorders, psychosocial stressors, comorbid disorders, biological factors and cognitive factors. The strongest support was found for the role of developmental factors in the chronicity of depression. Some support was found for the role of chronic stressors and certain personality features such as stress reactivity. Few other factors found support. The determinants of chronic depression do not differ qualitatively from acute depression. Rather, the development of chronic depression may involve increased levels of childhood adversity, protracted environmental stress and heightened stress reactivity. However, it is difficult to determine to what extent these putative determinants might reflect retrospective bias in data collection, or even parental reaction to children with subthreshold depressive traits. Detailed etiological models await further research attention to understudied areas and improved research designs. Suggestions for future research include greater specification of criteria for chronicity, use of more appropriate comparison groups and longer term prospective follow-up studies. PMID- 12117623 TI - Parental attitudes towards early intervention in children at high risk for affective disorders. AB - BACKGROUND: Parents volunteered to complete surveys on attitudes toward treatment intervention in children at a theoretically high (20-30%) or very high (70%) risk for affective disorders because of an assumed uni-lineal or bi-lineal family history of bipolar illness. METHODS: Questions focused on examining at what ages and stage of symptom and syndrome evolution parents would wish their child to begin treatment with different types of therapeutic approaches and clinical trial designs. Sixty percent of the respondents had a personal history of unipolar or bipolar affective disorders. RESULTS: In 156 completed surveys, 83% of parents favored acute medication intervention and 67% favored long-term medication treatment for those children at very high risk at or before the development of severe symptoms (i.e. even prior to meeting full diagnostic thresholds). On the average, parents indicated that they would enter their child in a trial of: two types of psychotherapy at a point in illness evolution between moderate and severe symptoms, two types of open medications between severe symptoms and a definite diagnosis, two blind medications at a definite diagnosis, and a blind trial of placebo and medication after a definite diagnosis but before multiple recurrences of the illness. Parents, primarily on the basis of perceived safety, would allow their children to use medications that have been found to be effective in adults. LIMITATIONS: In addition to a number of methodological limitations, responders to the survey were self-selected. CONCLUSIONS: The results indicate a willingness on the part of most parents to treat a child at high risk for affective illness early in the course of symptom evolution, even prior to a full syndromic illness or diagnosis. This and other parental views of the risk-benefit and ethical dimensions of early intervention may be helpful in the initiation and design of studies aimed at assessing the efficacy of early interventions in childhood-onset bipolar illness and its prodromes. PMID- 12117624 TI - Prevalence and predictors of premenstrual dysphoric disorder (PMDD) in older premenopausal women. The Harvard Study of Moods and Cycles. AB - BACKGROUND: Consistent data on the prevalence and predictors of premenstrual dysphoric disorder (PMDD) in the general population are lacking. METHODS: Premenstrual symptoms of 4164 women (aged 36-44 years) were retrospectively assessed by the Moos Premenstrual Inventory. From this original sample, 976 subjects were further interviewed at which time demographic and lifestyle characteristics, gynecologic history, and medical conditions were examined. The Structured Clinical Interview for DSM-IV Axis I disorders (SCID) was used to assess past and current psychiatric morbidity. Additionally, 513 of these women completed a prospective evaluation of premenstrual symptoms by using the Daily Rating of Severity of Problems Form (DRSP). RESULTS: The diagnosis of PMDD was confirmed in 33 of 513 women (6.4%) who completed the prospective evaluation with daily records. Fourteen subjects (2.7%) met criteria for PMDD without a previous history of depression. PMDD was associated with lower education (odds ratio [OR]=2.3, confidence interval [CI]=1.1-4.9), a history of major depression (OR=3.6, CI=1.7-7.4), and current cigarette smoking (OR=4.1, CI=1.5-11.1). In addition, women not working outside the home were significantly less likely to meet criteria for PMDD (OR=0.2, CI=0.1-0.9). LIMITATIONS: Only 513 of 976 women agreed to have their symptoms documented prospectively. Symptoms were assessed over the course of one menstrual cycle. CONCLUSIONS: This study indicates a significant prevalence of PMDD in a large community-based sample. Given the associated impairment in social and occupational functioning seen in PMDD, these prevalence data provide a strong rationale for enhanced recognition and treatment of the disorder. PMID- 12117625 TI - The long-term outcome of a benzodiazepine discontinuation programme in depressed outpatients. AB - OBJECTIVE: To assess longitudinally the prescription of psychotropic drugs in depressed patients after they participated in a benzodiazepine discontinuation programme. METHODS: Two hundred and thirty depressed patients on chronic benzodiazepine therapy took part in a discontinuation programme conducted in 36 general practices. After 2.3 years (S.D.=0.65, range 0.1-3.6) medical records were reviewed. RESULTS: Follow-up was achieved for 207 (90%) patients. Twenty five (12%) patients remained benzodiazepine free during the full follow-up period. The majority (n=181, 87%) was prescribed benzodiazepines at an average of 13 (+/-14) mg of diazepam equivalents for 537 (+/-375) days. Fifty-five (74% of 74) of the successfully discontinued patients restarted benzodiazepine therapy. Sixty-eight (33%) patients were prescribed benzodiazepines during the whole follow-up period. Successful taper predicted no or lower subsequent benzodiazepine prescription rates (OR=7.3; 95% CI: 2-16). No influence of GP policy towards benzodiazepine prescription could be detected (P=0.275). Antidepressants were prescribed in 115 (55%) patients for an average duration of 476 (+/-360) days. There was no difference in benzodiazepine prescription (dosage, duration) between patients who had or had not been prescribed an antidepressant. LIMITATIONS: Patients were not been diagnosed systematically during the follow-up period. CONCLUSIONS: If measured longitudinally, the rate of benzodiazepine prescription after discontinuation is much higher than reported in previous studies that have measured this cross-sectionally. Successful discontinuation is a strong predictor of modest or no future benzodiazepine prescription. Two-thirds of patients altered their benzodiazepine usage after taking part in a discontinuation programme. Treatment with antidepressants does not seem to influence benzodiazepine prescription. Patients' request (not GPs'policy) seems to be an important factor in continuing or resuming benzodiazepine prescription. PMID- 12117626 TI - Social adjustment in depression: the impact of depression severity, personality, and clinic versus community sampling. AB - BACKGROUND: Impaired social adjustment in depression has been associated with clinical variables, although results have been inconsistent. Previous research has not examined how social adjustment in depression varies by source of sample (clinical vs. community) or the associations with personality. METHODS: A total of 260 depressed outpatients from two samples completed the Social Adjustment Scale and were assessed on a number of clinical variables including depression severity, age, duration of depression, and personality symptoms. RESULTS: Overall social adjustment scores in our clinical samples were similar to those of an overseas clinical sample although there were some differences by subscales. In addition, the social adjustment scores of our clinical sample were significantly more impaired than those of individuals identified as depressed from a local epidemiological sample. Finally, in multiple regression equations, social adjustment scores were predicted by the clinical variables, severity of depression, younger age and personality symptoms. However, these only explained a small amount of variance in scores. CONCLUSION: Social adjustment in depression does not vary across different countries but varies according to the sample source (clinic vs. community). Some clinical variables are emerging as consistently associated with impaired social adjustment in depression. The associations with personality suggest this must be considered when assessing social adjustment in depressed patients. PMID- 12117627 TI - Does early intervention increase latency to relapse in major depressive disorder? Re-evaluation with cognitive behavior therapy. AB - BACKGROUND: Major depressive disorder (MDD) has been studied in relation to its propensity for remission and likelihood of relapse. While the general clinical lore suggests that early intervention benefits treatment outcome, the empirical validation of this assumption is inconclusive. Specifically, no studies have been conducted concerning Time to Treatment Entry and long-term clinical course for MDD. METHODS: For the current study, 53 participants received 16-weeks of cognitive behavioral therapy (CBT). Participants who remitted (n=41) from their depression were then inducted into a longitudinal follow-up protocol. RESULTS: Longer Time to Treatment Entry was predictive of longer time to relapse. A greater number of previous depressive episodes was associated with decreased Time to Treatment Entry. LIMITATIONS: A more elaborate protocol could be designed in order to explore the nature of treatment effects and Time to Treatment Entry within one study. CONCLUSIONS: CBT may be the most effective for patients who have delayed seeking treatment. Although the present study adds to the developmental neurobiological assumptions of Post [Severe depressive disorders (1994) 23-65] concerning affective 'kindling,' it also challenges the kindling theory's assumptions concerning early intervention. PMID- 12117628 TI - A comparison of characteristics of depressed patients and efficacy of sertraline and amitriptyline between Japan and the West. AB - BACKGROUND: This study was conducted to investigate the differences and similarities of the profile of depressed patients and the efficacy of the antidepressants, sertraline and amitriptyline, between Japan and the West (United States, Europe), using the Hamilton Depression Rating Scale (HAM-D) score of the individual patients. METHODS: Using common selection criteria, 680 patients from three regional clinical studies were chosen for this investigation. Factor analysis of the HAM-D scores for each regional group was carried out to compare the profile of depressed patients. Analysis of covariance was used to compare the efficacy of sertraline and amitriptyline between the regions. RESULTS: Factor analyses clearly showed three main factors (major depressive symptoms, anxiety and sleep disturbance) to be common across all three geographic regions. Higher HAM-D component scores of "Work and interests" and "Retardation" and lower ones of "Depressed mood" and "Feeling of guilt" were observed for the Japanese patients compared to the Western patients. Improvement of anxiety symptoms was marked for the Japanese amitriptyline treated patients. LIMITATIONS: Although the patient data used in these analyses were restricted by using identical selection criteria, there still remains some methodological shortcomings due to the original study design differences. CONCLUSIONS: Overall, the three main factors of depression and their magnitudes were similar between Japan and the West. The presentation of major depressive symptoms in Japanese patients differed from Western patients; this could be related to social, cultural and religious differences. Marked sedative effect for Japanese patients appeared to be due to the factor of anxiety being the strongest of the three main factors in Japanese depressed patients. PMID- 12117629 TI - Seasonal variations in children's calls to a help-line: implications for preventive services. AB - BACKGROUND: To investigate seasonal variations in telephone calls to a help-line for children, and their correlation to changes in length of day, latitude and age. METHOD: 691,787 calls to the Red Cross Help-Line in Norway for children in the three-year period 1996-1998 were included. RESULTS: Monthly frequencies of calls deviated significantly from an expected even distribution (chi2=9446.34, df=11, P<0.0001). The frequency curve for calls peaked in April and October and had its lowest level in July and December. Variation was pronounced: the mean number of daily calls varied between 436 in July and 886 in April. There was a strong and positive correlation between the number of calls and the change in length of day across the year (r(s)=0.76, N=12, P<0.01). Increasing latitude correlated positively with the range of the monthly observed/expected ratios of calls (r(s)=0.79, N=7, P<0.05). The frequency of calls was largest among children 12 to 14 years of age. Age correlated negatively with the range of monthly observed/expected ratios of calls (r=-0.94, N=12, P<0.001). LIMITATIONS: Social and cultural factors could not be explored in the design used in the present study. CONCLUSIONS: The frequency of calls from children correlates positively with change in length of day (i.e., maximal in spring and fall), and the magnitude of the seasonal variation correlates positively with latitude and negatively with age. Knowledge of seasonal variation in requests for help may have value in planning services for children. PMID- 12117630 TI - Atypical cognitive profile in patients with depression after myocardial infarction. AB - BACKGROUND: We evaluated the cognitive profile of 48 patients with major depression following their first myocardial infarction (MI). METHODS: The cognitive performance of the patients was compared with the performance of 48 non depressed MI patients and 48 healthy controls. RESULTS: Depressed MI patients performed slower on a simple cognitive speed related measure compared with non depressed MI patients and healthy controls. Attention and speed-related aspects of cognitive functioning were not affected. Surprisingly, (depressed) MI patients showed even better performances with respect to memory function. LIMITATION: No patients with non-MI-related depression were included. CONCLUSIONS: The cognitive profile of major depression after MI differs from that of non-cardiac-related depressive disorder, as described in the literature. This may reflect a different etiology of post MI depression from non-cardiac-related depression. PMID- 12117631 TI - Clinical databases of patients receiving antidepressants. The missing link between research and practice? AB - BACKGROUND: In the last 10 years the use of antidepressants has increased drastically. Unfortunately, the epidemiology of these compounds has shown significant gaps between recommendations derived from randomised controlled trials and current clinical practice. METHODS: We argue for the need to develop and maintain clinical databases of patients receiving antidepressants as a way of bridging this situation. RESULTS: In addition to experimental data generated in selected patients and settings, observational databases of large cohorts of typical patients, followed in typical settings, should be developed and maintained. Clinical databases could collect information on patient social and demographic characteristics, clinical symptoms, diagnosis and pharmacological and non-pharmacological treatments. In addition, they can provide accurate estimates of probabilities of different outcomes and on factors that affect outcome. CONCLUSION: Clinical databases should not be seen as another expensive administrative task for busy doctors. Clinical databases should be developed, organised and utilised only by clinicians who are interested in monitoring their clinical practice and want to provide patients, relatives and the public with information on prognosis and outcome in their specific context of care. Maintaining clinical databases is a routine process, nested in everyday clinical activity, which aims at constituting a permanent link between research and practice. PMID- 12117632 TI - Is occupation relevant in suicide? AB - BACKGROUND: The seasonality of suicide rates and methods of suicide may be related to changes in weather and conditions of employment. Particularly the amount of occupational outdoor exposure could show differences in the distribution of suicides by season and the selection of suicide method, in addition to age at the time of death. METHODS: The data consisted of all death certificates (n=1359) of completed suicides in the province of Oulu, Finland, during the years 1988-1999. For male subjects included in this study, four occupational groups were identified according to decreasing occupational outdoor exposure. The mean ages, the distribution of suicide methods and the seasonal variation in suicides for each occupational group were analyzed. RESULTS: Farmers were significantly older at the time of suicide than construction or indoor workers, and farmers employed significantly more violent methods than the other occupational groups. In the spring, farmers had a significant peak in the rate of suicides. In the winter, forest workers had a significant trough in the rate of suicides. In the summer, indoor workers had a significant peak in the rate of suicides. LIMITATIONS: The analyses were restricted to males due to the low number of females in the study population. CONCLUSIONS: The novel finding in this study was that the seasonality of violent suicides was most strongly seen as a spring peak and a winter trough among outdoor workers. The recognition of typical risk factors of different occupations, such as outdoor exposure, and occupational related susceptibility towards certain suicide methods could benefit in the prevention of suicides. PMID- 12117633 TI - Depression among hospitalised medically ill patients: a two-stage screening study. AB - BACKGROUND: There is wide variability in prevalence of depression in the physically ill. Moreover, data from the developing world is scarce. METHODS: Consecutive patients between the age 20-60 years and excluding those with malignancies and neurological illnesses, admitted to medical units (N=176), were screened with Beck's Depression Inventory (BDI). Those scoring more than 10 on BDI (N=55) were assessed with the Schedule for Clinical Assessment in Neuropsychiatry (SCAN). Age/sex matched healthy relatives served as controls. RESULTS: Thirty two percent (32%) of patients scored more than 10 on the BDI. None among controls scored more than the cut-off on BDI. Second-stage screening with SCAN on 33 subjects yielded a figure of 17% for depressive syndrome among the physically ill. Depression was mostly of the mild to moderate variety. CONCLUSIONS: High rates of depression were detected among physically ill hospitalised patients. Perhaps, differences in instruments used and population screened accounted for this. CLINICAL IMPLICATIONS: Need for increasing awareness as well as improving methods of detecting depression in physically ill is underscored. LIMITATIONS: a significant number of patients who screened positive on the BDI could not be interviewed with the SCAN for various reasons. Thus figures obtained after second stage screening are likely to be an underestimate. PMID- 12117634 TI - The impact of comorbid anxiety disorders on the course of dysthymic disorder: a 5 year prospective longitudinal study. AB - BACKGROUND: Few studies have examined the impact of comorbid anxiety disorder on the course of dysthymic disorder despite the high rate of comorbidity between these disorders. This research prospectively examines the naturalistic course of dysthymic disorder in patients with and without a comorbid anxiety disorder over a 5-year period. METHODS: Thirty-two comorbid patients and 54 non-comorbid patients with dysthymic disorder were assessed at three different time points (baseline, 30 months, and 60 months). Follow-up assessments included the Longitudinal Interval Follow-Up Evaluation and Hamilton Rating Scale for Depression (HRSD). RESULTS: The rate of recovery from dysthymic disorder was significantly lower in patients with (31.3%) than without (61.1%) comorbid anxiety disorders and, at all three time points, patients with comorbid anxiety had significantly higher HRSD scores. The estimated recovery rate from anxiety disorders was 53.8%. Only five of the comorbid patients recovered from both dysthymic disorder and all anxiety disorders during follow-up. Including new onsets, 72.1% of patients experienced an episode of an anxiety disorder during the 5 years. LIMITATIONS: There was no pure anxiety disorder group and patients were asked to report on relatively lengthy follow-up intervals. CONCLUSIONS: While the course of dysthymic disorder is debilitating, these results suggest that the prognosis for patients with a comorbid anxiety disorder is even poorer. PMID- 12117635 TI - The temperament and character inventory in major depression. AB - Patients admitted for pharmacological treatment of a non-bipolar major depressive episode completed the Temperament and Character Inventory (TCI) prior to and after at least 6 weeks of treatment. Treatment with various antidepressants resulted in a 43% reduction of symptomatology. Scores on the harm avoidance dimension before and after treatment appeared to be significantly higher as compared to Dutch normative data. TCI scores did not predict response to treatment or show a change during treatment. It is concluded that, in this group of patients, the personality dimension harm avoidance is a trait factor without predictive value for antidepressant responsiveness. PMID- 12117636 TI - Determinants of serum polychlorinated biphenyls and organochlorine pesticides measured in women from the child health and development study cohort, 1963-1967. AB - We examined predictors of organochlorine concentrations in serum specimens from women who were pregnant in the 1960s and participated in the Child Health and Development Study in the San Francisco Bay Area of California. That study enrolled pregnant women at the Kaiser-Permanente Medical Facilities, conducted interviews, and drew blood specimens; these specimens were centrifuged and the resulting serum specimens were frozen and placed in long-term storage. For the current investigation, organochlorines were measured by dual-column GC-electron capture detection in specimens collected in 1963-1967 from 399 pregnant women during the second and third trimesters. Using multiple linear regression models adjusted for serum lipids, we evaluated factors predicting concentrations of 11 polychlorinated biphenyl (PCB) congeners, their sum, and several pesticides and metabolites. Variables evaluated were age, race, place of birth, date of blood draw, body mass index, occupation, past residence on a farm, parity, and duration of pregnancy at blood draw. Concentrations of highly chlorinated PCBs and the sum of the PCBs increased with age. Concentrations of certain PCB congeners, as well as the sum, were significantly higher among nonwhites and increased with calendar date of blood draw. (italic)p,p(/italic) -DDT and (italic)p,p(/italic) -DDE concentrations were about 50% higher for nonwhites compared with whites and for those born in California or the southeastern United States versus elsewhere in the United States. Higher body mass index was associated with lower concentrations of several PCBs and (italic)p,p(/italic) -DDE but with higher heptachlor epoxide and DDT levels. The increase in use of PCBs during the 1960s is apparently detectable as increasing concentrations in maternal sera between 1963 and 1967. Marked racial and regional differences in serum pesticide levels were likely caused by geographic variation in previous agricultural and vector control uses. The relationship to body mass index appears to be complex. PMID- 12117637 TI - Serum dioxin concentrations and breast cancer risk in the Seveso Women's Health Study. AB - 2,3,7,8-Tetrachlorodibenzo-(italic)p(/italic)-dioxin (TCDD or dioxin), a widespread environmental contaminant, has been shown to disrupt multiple endocrine pathways. The International Agency for Research on Cancer classified TCDD as a known human carcinogen, primarily based on occupational studies of increased mortality from all cancers combined. Using data from the Seveso Women's Health Study (SWHS), we examined the association between individual serum TCDD levels and breast cancer risk in women residing around Seveso, Italy, in 1976, at the time of an industrial explosion that resulted in the highest known population exposure to TCDD. The SWHS cohort comprises 981 women who were infants to 40 years old in 1976, resided in the most contaminated areas at the time of the explosion, and had archived sera that was collected soon after the explosion. For each woman, serum TCDD exposure was measured by high-resolution mass spectrometry. Cancer cases were identified during interview and confirmed by medical record. At interview, 15 women (1.5%) had been diagnosed with breast cancer and serum TCDD levels for cases ranged from 13 to 1,960 ppt. Cox proportional hazards modeling showed that the hazard ratio for breast cancer associated with a 10-fold increase in serum TCDD levels (log(subscript)10(/subscript) TCDD) was significantly increased to 2.1 (95% confidence interval, 1.0-4.6). Covariate-adjusted results were not different. Individual serum TCDD is significantly related with breast cancer incidence among women in the SWHS cohort. Continued follow-up of the cohort will help shed light on the possible role of TCDD in the pathogenesis of breast cancer. PMID- 12117639 TI - Increasing habitat suitability in the United States for the tick that transmits Lyme disease: a remote sensing approach. AB - The warnings about the spread of (italic)Ixodes scapularis(/italic), one of the vectors of Lyme disease, into the United States are based on reports about regional distribution and increasing local abundance. In a modeling approach, I used the recorded, current distribution of this tick and remotely sensed bioclimatic factors over the United States to establish the changes of habitat for this tick since 1982 and to detect the areas with factors adequate to support tick colonization. Results indicate the geographic expansion of areas with adequate habitat suitability in the period 1982-2000. A discriminant analysis of counties with different degrees of habitat suitability shows that the increase in winter temperatures and in vegetation vitality (as a direct consequence of higher rainfall) is key to habitat switch from unsuitable to suitable. PMID- 12117638 TI - Serum dioxin concentrations and endometriosis: a cohort study in Seveso, Italy. AB - Dioxin, a ubiquitous contaminant of industrial combustion processes including medical waste incineration, has been implicated in the etiology of endometriosis in animals. We sought to determine whether dioxin exposure is associated with endometriosis in humans. We conducted a population-based historical cohort study 20 years after the 1976 factory explosion in Seveso, Italy, which resulted in the highest known population exposure to 2,3,7,8-tetrachlorodibenzo (italic)p(/italic)-dioxin (TCDD). Participants were 601 female residents of the Seveso area who were (3/4) 30 years old in 1976 and had adequate stored sera. Endometriosis disease status was defined by pelvic surgery, current transvaginal ultrasound, pelvic examination, and interview (for history of infertility and pelvic pain). "Cases" were women who had surgically confirmed disease or an ultrasound consistent with endometriosis. "Nondiseased" women had surgery with no evidence of endometriosis or no signs or symptoms. Other women had uncertain status. To assess TCDD exposure, individual levels of TCDD were measured in stored sera collected soon after the accident. We identified 19 women with endometriosis and 277 nondiseased women. The relative risk ratios (RRRs) for women with serum TCDD levels of 20.1-100 ppt and >100 ppt were 1.2 [90% confidence interval (CI) = 0.3-4.5] and 2.1 (90% CI = 0.5-8.0), respectively, relative to women with TCDD levels (3/4) 20 ppt. Tests for trend using the above exposure categories and continuous log TCDD were nonsignificant. In conclusion, we report a doubled, nonsignificant risk for endometriosis among women with serum TCDD levels of 100 ppt or higher, but no clear dose response. Unavoidable disease misclassification in a population-based study may have led to an underestimate of the true risk of endometriosis. PMID- 12117640 TI - Mitogen-activated protein kinase activation by oxidative and bacterial stress in an amphibian cell culture model. AB - The decline of many amphibian species could be caused by their susceptibility to environmental pollutants that cause cellular stress and cell death. A variety of intracellular signal transduction pathways are activated by environmental stress factors, which result in cell death. Mitogen-activated protein kinases are intracellular signaling molecules that include the extracellular signal-regulated kinases (ERK-1 and ERK-2). We used cultured (italic)Xenopus(/italic) tadpole cells (XTC-2 cells) to investigate the activation of ERK by oxidative or bacterial stress, two environmental factors that could contribute to pollution in aquatic systems. We exposed XTC-2 cell monolayers to hydrogen peroxide or bacterial lipopolysaccharide and measured ERK activation by Western blotting using antibodies raised against phosphorylated ERK-1 and ERK-2. Only ERK-2 was detected in XTC-2 cells. Both hydrogen peroxide and lipopolysaccharide caused ERK 2 phosphorylation in a time- and concentration-dependent manner. Hydrogen peroxide caused a 20- to 30-fold increase in ERK-2 activation that peaked 30 min after treatment, and lipopolysaccharide induced a 5- to 10-fold increase in ERK-2 activation that peaked 60 min after treatment. PD98059, an inhibitor of the ERK pathway, reduced the cytotoxic response of XTC-2 cells to hydrogen peroxide or lipopolysaccharide. These data suggest that ERK-2 is an intracellular target of oxidative and bacterial stress in amphibians that mediates, at least in part, the cytotoxic response to hydrogen peroxide or lipopolysaccharide. Moreover, the (italic)Xenopus(/italic) (XTC-2) cell culture system could serve as a useful model to identify agents that might threaten amphibian populations and human health. PMID- 12117641 TI - Indoor exposure to molds and allergic sensitization. AB - Evidence that indoor dampness and mold growth are associated with respiratory health has been accumulating, but few studies have been able to examine health risks in relation to measured levels of indoor mold exposure. In particular, little is known about the contribution of indoor molds to the development of allergic sensitization. As a part of an ongoing study examining the effects of ambient air pollutants on respiratory health and atopic diseases in German school children, we examined the relation between viable mold levels indoors and allergic sensitization in 272 children. We examined whether allergic sensitization in children is associated with higher fungal spore count in settled house dust sampled from living room floors. Adjusting for age, sex, parental education, region of residency, and parental history of atopy, we found that mold spore counts for Cladosporium and Aspergillus were associated with an increased risk of allergic sensitization. Sensitized children exposed to high levels of mold spores (> 90th percentile) were more likely to suffer from symptoms of rhinoconjunctivitis. We conclude that elevated indoor concentrations of molds in wintertime might play a role in increasing the risk of developing atopic symptoms and allergic sensitization not only to molds but also to other common, inhaled allergens. These effects were strongest in the group of children who had lived in the same home since birth. PMID- 12117644 TI - The importance of weight-normalized exposure data when issuing fish advisories for protection of public health. AB - Public health protection from environmental contaminants requires an understanding of the extent of contamination and of the extent of exposure to the contamination. My argument here is that weight-normalized, species-specific, individual-consumption pattern data are vital for determining exposure levels used to ascertain health protection measures and impacts from consuming contaminated fish. This study demonstrates the importance of adequate consumption pattern data for determining exposure distributions used for public health protection by examining three populations exposed to methylmercury through fish consumption: one recreational angler population and two Native-American populations. I compared exposure distributions derived from empirically derived species-specific, individual-consumption data from the three populations and exposure distributions derived, in part, from summary statistics for populations. In so doing, I conducted sensitivity analyses and population-specific probabilistic assessments of exposure. Although the goals of present-day accepted practices--using exposure distributions derived partly from point-estimate-based consumption and body-weight values--are laudable, results presented here indicate that weight-adjusted intake values for a population of concern are warranted when determining exposure distributions and should not be neglected in a health assessment instigated by available data on contaminant concentrations. If individual intake data are unobtainable, raw data from similar populations or tabulated values providing contaminant intake normalized for body weight may be viable alternatives to default values, and can be used to adequately protect public health. Without weight-normalized consumption pattern data to determine exposure, health assessment conclusions can mislead the public and have diminishing protective value. PMID- 12117642 TI - Apartment residents' and day care workers' exposures to tetrachloroethylene and deficits in visual contrast sensitivity. AB - Tetrachloroethylene (also called perchloroethylene, or perc), a volatile organic compound, has been the predominant solvent used by the dry-cleaning industry for many years. The U.S. Environmental Protection Agency (EPA) classified perc as a hazardous air pollutant because of its potential adverse impact on human health. Several occupational studies have indicated that chronic, airborne perc exposure adversely affects neurobehavioral functions in workers, particularly visual color discrimination and tasks dependent on rapid visual-information processing. A 1995 study by Altmann and colleagues extended these findings, indicating that environmental perc exposure at a mean level of 4,980 microg/m(3) (median=1,360 microg/m(3)) alters neurobehavioral functions in residents living near dry cleaning facilities. Although the U.S. EPA has not yet set a reference concentration guideline level for environmental exposure to airborne perc, the New York State Department of Health set an air quality guideline of 100 microg/m(3). In the current residential study, we investigated the potential for perc exposure and neurologic effects, using a battery of visual-system function tests, among healthy members of six families living in two apartment buildings in New York City that contained dry-cleaning facilities on the ground floors. In addition, a day care investigation assessed the potential for perc exposure and effects among workers at a day care center located in the same one-story building as a dry-cleaning facility. Results from the residential study showed a mean exposure level of 778 microg/m(3) perc in indoor air for a mean of 5.8 years, and that perc levels in breath, blood, and urine were 1-2 orders of magnitude in excess of background values. Group-mean visual contrast sensitivity (VCS), a measure of the ability to detect visual patterns, was significantly reduced in the 17 exposed study participants relative to unexposed matched-control participants. The groups did not differ in visual acuity, suggesting that the VCS deficit was of neurologic origin. Healthy workers in the day care investigation were chronically exposed to airborne perc at a mean of 2,150 microg/m(3) for a mean of 4.0 years. Again, group-mean VCS, measured 6 weeks after exposure cessation, was significantly reduced in the nine exposed workers relative to matched controls, and the groups did not differ significantly in visual acuity. These results suggested that chronic, environmental exposure to airborne perc adversely affects neurobehavioral function in healthy individuals. Further research is needed to assess the susceptibility of the young and elderly to perc induced effects, to determine whether persistent solvent-induced VCS deficits are a risk factor for the development of neurologic disease, and to identify the no observable adverse effect level for chronic, environmental, perc exposure in humans. PMID- 12117643 TI - Synergism between rhinovirus infection and oxidant pollutant exposure enhances airway epithelial cell cytokine production. AB - Of the several factors believed to exacerbate asthmatic symptoms, air pollution and viral infections are considered to be particularly important. Although evidence indicates that each of these respiratory insults individually can increase asthma severity in susceptible individuals, we know little about the extent to which exposure to environmental oxidant pollutants can influence the course of respiratory viral infection and its associated inflammation. To investigate the interaction of these two stimuli within their common epithelial cell targets in the upper and lower respiratory tracks, we infected primary human nasal epithelial cells and cells of the BEAS-2B line grown at the air-liquid interface with human rhinovirus type 16 (RV16) and exposed them to NO2 (2.0 ppm) or O3 (0.2 ppm) for 3 hr. Independently, RV16, NO2, and O3 rapidly increased release of the inflammatory cytokine interleukin-8 through oxidant-dependent mechanisms. The combined effect of RV16 and oxidant ranged from 42% to 250% greater than additive for NO2 and from 41% to 67% for O3. We abrogated these effects by treating the cells with the antioxidant N-acetylcysteine. Surface expression of intercellular adhesion molecule 1 (ICAM-1) underwent additive enhancement in response to combined stimulation. These data indicate that oxidant pollutants can amplify the generation of proinflammatory cytokines by RV16 infected cells and suggest that virus-induced inflammation in upper and lower airways may be exacerbated by concurrent exposure to ambient levels of oxidants commonly encountered the indoor and outdoor environments. PMID- 12117645 TI - Trichloroacetic acid as a biomarker of exposure to disinfection by-products in drinking water: a human exposure trial in Adelaide, Australia. AB - We addressed the need for a biomarker of ingestion exposure to drinking water disinfection by-products by performing a human exposure trial. We evaluated urinary excretion of trichloroacetic acid (TCAA) as an exposure biomarker using 10 volunteers who normally consume their domestic tap water. We recruited the volunteers at a water quality research laboratory in Adelaide, Australia. Participants maintained a detailed consumption and exposure diary over the 5-week study. We also analyzed tap water and first morning urine (FMU) samples for TCAA, and tap water for chloral hydrate (CH). We documented both interindividual and intraindividual variability in TCAA ingestion and urinary excretion, and both were substantial. With a TCAA-free bottled water intervention, we used creatinine adjusted urinary TCAA levels to estimate urinary TCAA excretion half-lives for three of the participants. We observed correspondence over time between estimated TCAA excretion, calculated from TCAA + CH ingestion levels, and measured TCAA urinary excretion. This study demonstrates the merits and feasibility of using TCAA in FMU as an exposure biomarker, and reveals remaining concerns about possible alternate sources of TCAA exposure for individuals with low drinking water ingestion exposure. PMID- 12117646 TI - Volatile organic compounds as breath biomarkers for active and passive smoking. AB - We used real-time breath measurement technology to investigate the suitability of some volatile organic compounds (VOCs) as breath biomarkers for active and passive smoking and to measure actual exposures and resulting breath concentrations for persons exposed to tobacco smoke. Experiments were conducted with five smoker/nonsmoker pairs. The target VOCs included benzene, 1,3 butadiene, and the cigarette smoke biomarker 2,5-dimethylfuran. This study includes what we believe to be the first measurements of 1,3-butadiene in smokers' and nonsmokers' breath. The 1,3-butadiene and 2,5-dimethylfuran peak levels in the smokers' breath were similar (360 and 376 microg/m(3), respectively); the average benzene peak level was 522 microg/m(3). We found higher peak values of the target chemicals and shorter residence times in the body than previously reported, probably because of the improved time resolution made possible by the continuous breath measurement method. The real-time breath analyzer also showed the presence of the chemicals after exposure in the breath of the nonsmokers, but at greatly reduced levels. Single breath samples collected in evacuated canisters and analyzed independently with gas chromatography/mass spectrometry confirmed the presence of the target compounds in the postexposure breath of the nonsmokers but indicated that there was some contamination of the breath analyzer measurements. This was likely caused by desorption of organics from condensed tar in the analyzer tubing and on the quartz fiber filter used to remove particles. We used the decay data from the smokers to estimate residence times for the target chemicals. A two-compartment exponential model generally gave a better fit to the experimental decay data from the smokers than a single compartment model. Residence times for benzene, 1,3-butadiene, and 2,5 dimethylfuran ranged from 0.5 (1,3-butadiene) to 0.9 min (benzene) for tau1 and were essentially constant (14 min) for tau2. These findings will be useful in models of environmental tobacco smoke exposure and risk. PMID- 12117647 TI - Cadmium and lead in blood in relation to low bone mineral density and tubular proteinuria. AB - Long-term exposure to cadmium may cause kidney and bone damage. Urinary cadmium is commonly used as the dose estimate for the body burden of cadmium. However, elevated levels of cadmium in the urine may reflect not only high levels of cadmium dose but also renal dysfunction. In this study we used blood cadmium as the dose estimate. In addition, we analyzed blood lead. We examined 479 men and 542 women, ages 16-81 years, who were environmentally or occupationally exposed to cadmium and lead. We used urinary protein alpha 1-microglobulin as a marker for tubular proteinuria and measured forearm bone mineral density using dual energy X-ray absorptiometry. The relationship between blood cadmium and tubular proteinuria was strong, even when we excluded occupationally exposed participants. The subgroup with the highest blood cadmium levels had a 4-fold risk of tubular proteinuria compared to the subgroup with the lowest blood cadmium levels. In the older age group (age > 60), the risk of low bone mineral density (z-score < -1) for the subgroup with the highest blood cadmium levels was almost 3-fold compared to the group with lowest blood cadmium levels. We found no similar associations for lead. The observed effects may be caused by higher cadmium exposure in the past. This study strengthens previous evidence that cadmium exposure may affect both bone mineral density and kidney function. PMID- 12117648 TI - Characterization of the dust/smoke aerosol that settled east of the World Trade Center (WTC) in lower Manhattan after the collapse of the WTC 11 September 2001. AB - The explosion and collapse of the World Trade Center (WTC) was a catastrophic event that produced an aerosol plume impacting many workers, residents, and commuters during the first few days after 11 September 2001. Three bulk samples of the total settled dust and smoke were collected at weather-protected locations east of the WTC on 16 and 17 September 2001; these samples are representative of the generated material that settled immediately after the explosion and fire and the concurrent collapse of the two structures. We analyzed each sample, not differentiated by particle size, for inorganic and organic composition. In the inorganic analyses, we identified metals, radionuclides, ionic species, asbestos, and inorganic species. In the organic analyses, we identified polycyclic aromatic hydrocarbons (PAHs), polychlorinated biphenyls, polychlorinated dibenzodioxins, polychlorinated dibenzofurans, pesticides, phthalate esters, brominated diphenyl ethers, and other hydrocarbons. Each sample had a basic pH. Asbestos levels ranged from 0.8% to 3.0% of the mass, the PAHs were > 0.1% of the mass, and lead ranged from 101 to 625 microg/g. The content and distribution of material was indicative of a complex mixture of building debris and combustion products in the resulting plume. These three samples were composed primarily of construction materials, soot, paint (leaded and unleaded), and glass fibers (mineral wool and fiberglass). Levels of hydrocarbons indicated unburned or partially burned jet fuel, plastic, cellulose, and other materials that were ignited by the fire. In morphologic analyses we found that a majority of the mass was fibrous and composed of many types of fibers (e.g., mineral wool, fiberglass, asbestos, wood, paper, and cotton). The particles were separated into size classifications by gravimetric and aerodynamic methods. Material < 2.5 microm in aerodynamic diameter was 0.88-1.98% of the total mass. The largest mass concentrations were > 53 microm in diameter. The results obtained from these samples can be used to understand the contact and types of exposures to this unprecedented complex mixture experienced by the surviving residents, commuters, and rescue workers directly affected by the plume from 11 to 12 September and the evaluations of any acute or long-term health effects from resuspendable dust and smoke to the residents, commuters, and local workers, as well as from the materials released after 11 September until the fires were extinguished. Further, these results support the need to have the interior of residences, buildings, and their respective HVAC systems professionally cleaned to reduce long-term residential risks before rehabitation. PMID- 12117649 TI - Biologic effects induced in vitro by PM10 from three different zones of Mexico City. AB - Exposure to urban airborne particles is associated with an increase in morbidity and mortality. There is little experimental evidence of the mechanisms involved and the role of particle composition. We assessed cytotoxicity (crystal violet assay), apoptosis [terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) or annexin V assay], DNA breakage (comet assay), and production of proinflammatory mediators [tumor necrosis factor Alpha (TNF-Alpha), interleukin 6 (IL-6), prostaglandin E2 (PGE2)] (enzyme-linked immunosorbent assay), and E-selectin (flow cytometry) in cell lines exposed to particulate matter < 10 microm in size (PM10) obtained from the northern, central, and southern zones of Mexico City. Particle concentrations ranged from 2.5 to 160 microg/cm(2). We used epithelial, endothelial, fibroblastic, and monocytic cells and assessed DNA damage in Balb-c cells, TNF-Alpha and IL-6 production in mouse monocytes, and PGE2 in rat lung fibroblasts. We determined the expression of E selectin in human endothelial cells and evaluated the cytotoxic potential of the PM10 samples in all cell types. PM10 from all three zones of Mexico City caused cell death, DNA breakage, and apoptosis, with particles from the north and central zones being the most toxic. All of these PM10 samples induced secretion of proinflammatory molecules, and particles from the central zone were the most potent. Endothelial cells exposed to PM10 from the three zones expressed similar E-selectin levels. Mexico City PM10 induced biologic effects dependent on the zone of origin, which could be caused by differences in the mixture or size distribution within particle samples. Our data suggest that particle composition as well as particle size should be considered in assessing the adverse effects of airborne particulate pollution. PMID- 12117651 TI - Family correlations of arsenic methylation patterns in children and parents exposed to high concentrations of arsenic in drinking water. AB - We investigated the evidence of a familial contribution to urinary methylation patterns in families ingesting arsenic in drinking water. Arsenic methylation can be assessed by measuring urinary levels of inorganic arsenic (InAs) and its methylated metabolites, monomethylarsonate (MMA), and dimethylarsinate (DMA). Methylation activity is reflected in the ratios: InAs/methylated arsenic (InAs/metAs) and MMA/DMA. Eleven families from Chile were selected because of their long-term exposure to very high levels of arsenic in drinking water (735 762 microg/L). Each family consisted of a father, a mother, and two children. We measured urinary arsenic and its methylated metabolites for each participant (n = 44). The intraclass correlation coefficients showed that 13-52% of the variations in the methylation patterns were from being a member of a specific family. Family correlations were calculated for father-mother, parent-child, and sibling-sibling pairs. Methylation patterns correlated strongly between siblings [r = 0.78 for InAs/metAs, 95% confidence interval (CI), 0.34-0.94; r = 0.82 for MMA/DMA, 95%CI, 0.43-0.95] compared to lower correlations in father-mother pairs (r = 0.18, r = 0.01, respectively), after adjustment for total urinary arsenic, age, and sex. Family correlations were not notably altered when adjustments were made for specific blood micronutrients (methionine, homocysteine, folate, vitamin B6, selenium, and vitamin B12 potentially related to methylation. We also report on a family pedigree with high prevalence of arsenic-induced effects. Participants from this family had low InAs/metAs values, which is consistent with increased toxicity of trivalent methylated arsenic species. Despite our small sample size, we observed that methylation patterns aggregate in families and are correlated in siblings, providing evidence of a genetic basis for the variation in arsenic methylation. Larger studies with more extensive pedigrees will need to be conducted to confirm these findings. PMID- 12117650 TI - Environmental pollutants and disease in American children: estimates of morbidity, mortality, and costs for lead poisoning, asthma, cancer, and developmental disabilities. AB - In this study, we aimed to estimate the contribution of environmental pollutants to the incidence, prevalence, mortality, and costs of pediatric disease in American children. We examined four categories of illness: lead poisoning, asthma, cancer, and neurobehavioral disorders. To estimate the proportion of each attributable to toxins in the environment, we used an environmentally attributable fraction (EAF) model. EAFs for lead poisoning, asthma, and cancer were developed by panels of experts through a Delphi process, whereas that for neurobehavioral disorders was based on data from the National Academy of Sciences. We define environmental pollutants as toxic chemicals of human origin in air, food, water, and communities. To develop estimates of costs, we relied on data from the U.S. Environmental Protection Agency, Centers for Disease Control and Prevention, National Center for Health Statistics, the Bureau of Labor Statistics, the Health Care Financing Agency, and the Practice Management Information Corporation. EAFs were judged to be 100% for lead poisoning, 30% for asthma (range, 10-35%), 5% for cancer (range, 2-10%), and 10% for neurobehavioral disorders (range, 5-20%). Total annual costs are estimated to be $54.9 billion (range $48.8-64.8 billion): $43.4 billion for lead poisoning, $2.0 billion for asthma, $0.3 billion for childhood cancer, and $9.2 billion for neurobehavioral disorders. This sum amounts to 2.8 percent of total U.S. health care costs. This estimate is likely low because it considers only four categories of illness, incorporates conservative assumptions, ignores costs of pain and suffering, and does not include late complications for which etiologic associations are poorly quantified. The costs of pediatric environmental disease are high, in contrast with the limited resources directed to research, tracking, and prevention. PMID- 12117652 TI - Cockatiel-induced hypersensitivity pneumonitis. AB - Diagnosing an environmental or occupationally related pulmonary disorder often involves a process of elimination. Unlike commonly diagnosed conditions in other specialties, a cause-and-effect relationship may be implied, yet other factors such as temporality and biologic plausibility are lacking. Our patient was referred with a suspected work-related pulmonary disorder. For several years, she had suffered with dyspnea on exertion and repeated flulike illnesses. She worked at an automobile repair garage that performed a large number of emission tests, and there was concern that her workplace exposures were the cause of her symptoms. After a careful review of her history, physical examination, and laboratory testing, we came to the conclusion that she had hypersensitivity pneumonitis related to pet cockatiels in her home. Clinical points of emphasis include the importance of a complete environmental history and careful auscultation of the chest when performing the physical examination. In addition, we encountered an interesting physical diagnostic clue, a respiratory sound that assisted with the eventual diagnosis. PMID- 12117653 TI - Mathematical modeling in environmental health. PMID- 12117654 TI - Estrogenicity of styrene oligomers and assessment of estrogen receptor binding assays. PMID- 12117655 TI - Up a chemical creek. PMID- 12117656 TI - Decoding the cryptic origins of colon cancer. PMID- 12117657 TI - Antibiotic resistance in livestock: more at stake than steak. PMID- 12117658 TI - Compensating for cold war cancers. AB - Although the Cold War has ended, thousands of workers involved in nuclear weapons production are still living with the adverse health effects of working with radioactive materials, beryllium, and silica. After a series of court battles, the U.S. government passed the Energy Employees Occupational Illness Act in October 2000 to financially assist workers whose health has been compromised by these occupational exposures. Now work is underway to set out guidelines for determining which workers will be compensated. The National Institute for Occupational Safety and Health has been assigned the task of developing a model that can scientifically make these determinations, a heavy task considering the controversies that lie in estimating low-level radiation risks and the inadequate worker exposure records kept at many of the plants. PMID- 12117659 TI - A BETR way to track toxic pollutants. AB - Why are chemicals that have been banned for over a decade still being detected in the environment? How is it that chemicals and other pollutants such as particulate matter turn up hundreds or even thousands of miles from their sources? These are just two of the questions that a team of American and Canadian scientists hope will be answered using a model they have developed to track pollutants across North America. In the short term, the model's developers are concentrating on using it to assess rates of exposure from food sources, but they hope to expand the model to include global data, a step they hope will help policy makers better understand the consequences of pollution. PMID- 12117660 TI - Fluticasone for the treatment of symptomatic bronchial asthma in children treated with sodium cromogylate--a prospective, randomised trial. AB - BACKGROUND: Children with persistent mild to moderate bronchial asthma require anti-inflammatory therapy. According to current treatment guidelines both sodium cromoglycate (SCG) and inhaled corticosteroids can be used. If children remain symptomatic despite regular SCG therapy, corticosteroids are the next therapeutic option. AIMS: To determine whether combined SCG and fluticasone (inhaled corticosteroid) therapy is of additional benefit in children who are symptomatic on SCG compared with simply switching to fluticasone. PATIENTS AND METHODS: Children with mild or moderate persistent asthma aged 6 to 16 years who had been treated with inhaled SCG for at least 3 months prior to the study received either 2 mg SCG four times daily from a metered dose inhaler plus fluticasone propionate powder 50 microg b.i.d. from the Diskus inhaler (group FS) or fluticasone 50 microg b.i.d. only (group F). The randomised, controlled, parallel-group study had a 2 week run-in phase and an 8 week treatment period. Morning and evening peak expiratory flow rates (PEFR) were measured daily by patients and recorded in diaries. Asthma symptoms, use of rescue medication and spirometry were also documented. RESULTS: Paediatricians from 21 study centres recruited 124 children with asthma, of whom 104 fulfilled randomisation criteria and were allocated to study medication. Morning PEFR increased by 47 l/min and by 45 l/min after 8 weeks of treatment in groups F and FS, respectively. The adjusted difference between groups was 0.84% of predicted (95% CI, -7.3 to 5.6, p=0.80). Asthma symptoms and lung function also improved with no significant differences between treatment groups (p>0.24). Frequency and severity of adverse events was similar in both groups. CONCLUSION: In children who are symptomatic while taking sodium cromoglycate four times daily, the combination of inhaled fluticasone and SCG is not superior to fluticasone alone. SCG can safely be withdrawn when commencing fluticasone, thus facilitating asthma treatment. PMID- 12117661 TI - Coronary artery stenting in diabetes mellitus--unfavourable clinical outcome due to increased rate of myocardial ischemia and percutaneous interventions. AB - Diabetes mellitus is one of the major risk factors for cardiovascular diseases. The aim of this study was to analyze if diabetic patients, compared to nondiabetic patients, have a worse angiographic and clinical success rate and a reduced clinical and angiographic outcome at three-months follow-up after coronary artery stenting according to postprocedural complications, recurrent angina, myocardial ischemia, restenosis and revascularization. A total of 307 unselected patients with coronary artery disease and myocardial ischemia who underwent intracoronary stenting were included in this study. Diabetes was present in 49 patients. Morphological criteria, angiographic results and clinical in-hospital outcome did not differ significantly between both groups. At follow up diabetics presented significantly more often recurrent angina and myocardial ischemia. The rate of restenosis and target lesion revascularization was not different. Among diabetics, the rate of percutaneous coronary interventions because of different lesions was significantly increased. Diabetes does not reduce the angiographic result initially and at follow-up after coronary artery stenting. Diabetes mellitus limits the clinical outcome because of recurrent angina, myocardial ischemia and the need of coronary interventions. PMID- 12117662 TI - Thyroid hypoechogenicity in patients with chronic hepatitis receiving interferon alpha therapy: evaluation by standardized grey scale ultrasonography. AB - BACKGROUND: Recombinant human interferon-a therapy of chronic hepatitis B and C is associated with the induction of thyroid dysfunction in up to 15%. Little is known about morphological changes of the thyroid gland, especially about its tissue echogenicity under this immunomodulative drug treatment. METHODS: 116 patients with chronic hepatitis B or C were consecutively investigated. 53 patients qualified for treatment with interferon-a alone or in combination with ribavirin. Patients with normal serum aminotransferase levels, advanced liver cirrhosis, hepatocellular carcinoma, active intravenous drug or alcohol abusus and with focal thyroid lesions (nodes, cysts, calcifications) were excluded. Thyroid function was determined by measurements of FT4, TSH and thyroidal autoantibodies. Ultrasonography was performed before, during and after interferon a therapy including volumetry and standardized grey scale analysis. The data were compared with values of 100 euthyroid volunteers as control group. RESULTS: After six months of therapy patients differed from controls by significant TSH elevation (12.8 +/- 9.34 vs. 2.8 +/- 1.1 microU/ml, p<0.02). Six patients (11%) developed overt hypothyroidism with detectable thyroidal autoantibodies. Thyroid volume in patients was similar (13.0 +/- 4.1 ml) to that in the control group (12.6 +/- 4.7 ml). However, thyroid echogenicity of the patients was significantly lower after 6 months of therapy (21.9 +/- 2.5 grey scales) compared to the status before (25.6 +/- 2.3 grey scales) and compared to the values of controls (25.4 +/- 2.1 grey scales, p <0.002). CONCLUSION: Beside of functional disorders interferon-alpha leads to thyroid hypoechogenicity suggesting relevant morphological changes of the organ. PMID- 12117663 TI - Nosocomial infections in internal medicine, University of Frankfurt, Germany--a prospective surveillance study. AB - Nosocomial infections (NI) are serious complications associated with high morbidity and mortality. In the present study, NI were analyzed prospectively at the Center of Internal Medicine (CIM) (300 beds) of the J. W. Goethe-University a 1380-bed major tertiary care teaching hospital- during a study period of six month. During the study period a single physician evaluated all patients with signs and symptoms of an infection during his daily rounds. NI was defined as body temperature >38 degrees C and evidence of an infection not before the third day of admission to the hospital. NI was diagnosed in 127 patients (3.5%) of the 3605 patients studied. The data of 126 patients with NI could be collected and analyzed completely. Of the 126 patients 34 patients died. The mean length of hospitalization before the diagnosis of NI was 12.0 days (standard-deviation: +/ 13.1 days; median: 7 days). Compared to all patients with NI significantly more patients of the Internal Intensive care unit (11.3%), of the HIV-ward (10.3%), and of the hematology / oncology ward (5.8%) acquired a NI (p<0.05). With respect to other groups of patients the frequency of NI ranged from 0.5 to 4.6 per 100 admitted inpatients. The lower rate was in patients admitted for invasive diagnostic procedures who were hospitalized only for 3 days or less. PMID- 12117664 TI - Determination of vitamin A palmitate in buccal mucosal cells: a pilot study. AB - We report in our present pilot study on the uptake of retinyl palmitate and its formation to retinol in buccal mucosal cells. Retinoids are able to change metaplastic lesions in several tissues. Prior to any clinical evaluation it is necessary to measure its uptake in target tissues such as the buccal mucosal epithelium. In this pilot study 12 volunteers creamed the inside of their cheeks lightly with a 0.1% retinyl palmitate containing toothpaste for 10 days followed by a wash out phase. On day 0, 3, 7, 10, 17 and 21, buccal mucosal cell samples were taken from volunteers and the uptake of retinol and retinyl palmitate was determined by HPLC analysis. An uptake of vitamin A was demonstrated in all volunteers. Comparing day 0 to day 3 a significant uptake of retinyl palmitate (p<0.05) was detected and comparing day 3 to 17 and 21 a significant decrease (p<0.05) during growth and differentiation of the mucosal epithelium could be seen. The subsequently formed retinol showed an increase but no statistical significance was determined. - The uptake of retinyl palmitate and the formation to retinol led to an enrichment of vitamin A in buccal mucosal cells. In this way metaplastic alterations of the buccal mucosal epithelium might be prevented or reversed by the application of topical retinyl palmitate. PMID- 12117665 TI - HHV-8 DNA in blood and the development of HIV-associated Kaposi's sarcoma in the era of HAART--a prospective evaluation. AB - OBJECTIVE: To explore the significance of HHV-8 viremia in HIV-positive individuals for the risk of developing Kaposi's sarcoma (KS) in the era of highly active antiretroviral therapy. METHODS: 237 HIV-positive patients were included in this prospective evaluation and followed over an average duration of 34 months. HHV-8 DNA in peripheral blood mononuclear cells (PBMCs) and CD4 lymphocytes were determined. In addition AIDS-defining conditions and antiretroviral therapy were documented of all participating subjects. RESULTS: HHV-8 DNA was detectable in PBMCs of 12.6% out of all individuals. 53.3% of these patients initially complained about KS, although 9.2% of patients without HHV-8 DNA in PBMCs were found on KS as well. Furthermore, four patients in total were observed with newly developed KS during follow up visits. None of these patients were noted with detectable HHV-8 DNA at their initial evaluation. CONCLUSIONS: Prevalence of HHV-8 DNA in PBMCs of subjects in this investigation was quite similar to former investigations. However, new diagnosed KS occurred less frequently than demonstrated in previous studies. All of those observed patients with new KS manifestations were negative for HHV-8 DNA in PBMCs at study entry. This observation differs from earlier studies which have postulated the detection of HHV-8 DNA in PBMCs as a predictive value for development of KS. Due to results as presented, a single HHV-8 DNA test in blood has no predictive value in support of predictability of KS development. With respect toto costs and to a less complicated performance antibody assays should be preferred. PMID- 12117666 TI - Mycobacterium tuberculosis gene-amplification in breath condensate of patients with lung tuberculosis. AB - The applicability of Mycobacterium tuberculosis-gene-amplification test (NAT) in breath condensate (BC) was examined in lung tuberculosis (TB). Ten patients with bacteriologically confirmed lung TB have been examined by ligase chain reaction (LCx). In BC the NAT were negative in all patients as well as the examination of acid fast smears and of solid phase or liquid media cultures were. This indicates that the use of NAT in BC can not replace or complement sputum or materials obtained invasively in patients with lung TB. PMID- 12117667 TI - Clinical management and evidence based treatment of chronic hepatitis C--12th European Congress of Clinical Microbiology and Infectious Diseases, Milan, Italy, 21-24 April 2002. AB - As in totally 1% of the worldwide population is afflicted by hepatitis C virus with most of the 200 Mio carriers not diagnosed or treated, the clinical management of chronic hepatitis C remains an urgent demand for global health. The 12th European Congress of Clinical Microbiology and Infectious Diseases (12th ECCMID) held in Milan, Italy, from 21 to 24 April 2002, focussed on viral hepatitis in a keynote lecture as well as a session on evidence based treatment of chronic infection and various poster presentations. PMID- 12117668 TI - A profibrotic polymorphism (of TGFbeta1) in systemic sclerosis. PMID- 12117669 TI - Muscle, exercise and arthritis. PMID- 12117670 TI - Recurrent fevers in the presence of multiple autoimmune diseases and antibody deficiency. PMID- 12117671 TI - Analysis of transforming growth factor beta1 gene polymorphisms in patients with systemic sclerosis. AB - OBJECTIVES: To determine the distribution of transforming growth factor beta1 (TGFbeta1) genotypes at codon 10 (+869 polymorphism) and codon 25 (+915 polymorphism) in patients with scleroderma (SSc). Differences between diffuse and limited SSc (dSSc and lSSc) were also investigated. METHODS: Patients with lSSc (n=89) and dSSc (n=63) were compared with 147 controls. DNA was isolated from peripheral blood and polymorphisms at codons 10 (C/T) and 25 (G/C) of the TGFbeta1 gene analysed by polymerase chain reaction and sequence specific oligonucleotide probing. RESULTS: Significantly more patients with SSc than controls carried allele C at codon 10 (controls v SSc, 38% v 48%, chi(2)=8.2, 1df, p=0.004), OR=1.95 (95% CI 1.16 to 3.27). The difference remained when patients with SSc were split into those with limited or diffuse disease, (controls v dSSc, chi(2)=5, 1df, p=0.02 and controls v lSSc, chi(2)=6, 1df, p=0.013). The patients with SSc had significantly more subjects heterozygous at codon 10 (controls v SSc, chi(2)=45, 1df, p<0.0001). Possession of allele C at codon 10 gave an OR=4.8 (95% CI 2.8 to 8.4). No difference in allele frequency was seen between patients with SSc and controls at codon 25. More patients with SSc than controls carried the GG genotype (controls v SSc, 80% v 88%, chi(2)=7, 2df, p=0.027). Possession of allele G gave an OR=1.7 (95% CI 0.5 to 5.9). There was no difference between diffuse and limited disease at either codon. CONCLUSIONS: These results suggest that patients with SSc are genetically predisposed to high TGFbeta1 production. These polymorphisms do not, however, explain the difference in the clinical phenotypes of limited and diffuse SSc. PMID- 12117672 TI - Renal functional reserve is impaired in patients with systemic sclerosis without clinical signs of kidney involvement. AB - OBJECTIVE: To evaluate the functional response of the kidney to an amino acid challenge (the so called renal functional reserve (RFR)) in patients with systemic sclerosis (SSc) with no clinical sign of renal involvement. METHODS: Before and after an intravenous amino acid load (Freamine III Baxter, 8.5% solution, 4.16 ml/min for two hours), glomerular filtration rate (GFR, as creatinine clearance), effective renal plasma flow (ERPF, as para-aminohyppurate clearance), and calculated total renal vascular resistance (TRVR) were measured in 21 patients with SSc with apparently normal renal function and 10 normal controls. RESULTS: In basal conditions, patients had lower ERPF (403.5 (SD 43.8) v 496.4 (SD 71.3) ml/min, p<0.0002) and higher TRVR (10 822 (SD 2044) v 8874 (SD 1639) dyne/sxcm(-5), p<0.014) than controls. The RFR, evaluated as the percentage increase of GFR after the amino acid load, was significantly reduced in patients with SSc (SSc +1.9 (SD18.6)%, controls +34.8 (SD 13.9)%; p<0.0002). However, the response of patients was not uniform. Multiple regression analysis showed that the RFR was inversely dependent on the patients' mean arterial pressure at admission and basal GFR (R(2)=65%, p<0.0001). CONCLUSIONS: Most patients with SSc cannot increase renal filtration under the challenge of a protein overload. This defective renal response to the amino acid load test sustains the concept of the prevalence of vasoconstrictor over vasodilating factors in the kidney of these patients. PMID- 12117673 TI - Investigation of hip abductor activation in subjects with clinical unilateral hip osteoarthritis. AB - OBJECTIVES: (a) To compare the magnitude of gluteus medius and tensor fascia lata activation between a group of subjects with clinical unilateral hip osteoarthritis and a group of healthy older adults. (b) To compare the magnitude of activation of the gluteus medius and tensor fascia lata between sides in a group of subjects with clinical unilateral hip osteoarthritis and a group of healthy older adults. METHODS: 19 subjects with clinical unilateral hip osteoarthritis and 19 healthy controls were investigated. The subjects performed a stepping task during which recordings were obtained using surface electromyograms from the hip abductors, and kinetic data were obtained from a dual force platform. RESULTS: Subjects with clinical hip osteoarthritis had higher gluteus medius activation than the healthy older adults (p=0.037). In addition, there were no differences in the magnitude of gluteus medius activation between the sides (p=0.733). There was no difference in the force platform data between the groups (p=0.078). CONCLUSIONS: The increased magnitude of gluteus medius activation in the group with hip osteoarthritis is evidence of a muscular dysfunction associated with hip disease. This has implications for the progressive nature of the disease and for its conservative management. PMID- 12117674 TI - Employment perspectives of patients with ankylosing spondylitis. AB - OBJECTIVES: To assess the labour market position of patients with ankylosing spondylitis (AS) in relation to disease duration and to identify potential factors in relation to withdrawal from the labour force. METHODS: A cross sectional mail survey was conducted among 658 patients with AS. Participation in the labour force was defined as having a paid job. The independent effect of duration of disease was examined by an indirect method of standardisation. A broad variety of risk factors were examined separately and in a combined analysis, including sociodemographic factors, disease related variables, coping styles, and work related factors. Attributable and preventable fractions were calculated from the combined analyses to assess the relative importance of the contributing factors. RESULTS: Probability of participation in the labour force was similarly reduced in patients with AS with different durations of disease. Pacing to cope with limitations was the most relevant factor in increasing the risk of withdrawal from the labour force, accounting for 73% of withdrawals. Coping with limitations by often seeking creative solutions, high disease activity, increased age, and insufficient support from colleagues or management were also positively associated with withdrawal from the labour force. Technical or ergonomic adjustments of the workplace, working in large companies, and coping with dependency style through frequent acceptance were negatively associated. Of these factors, technical or ergonomic adjustment was the most relevant in terms of reducing the risk. CONCLUSION: Sociodemographic factors, disease related factors, coping styles, and work related factors contribute simultaneously to withdrawal from the labour force. PMID- 12117675 TI - One year outcome of undifferentiated polyarthritis. AB - OBJECTIVE: To identify variables that can predict a progressive outcome after one year of follow up in patients presenting with undifferentiated polyarthritis (UPA) at an early arthritis clinic. METHODS: New patients with arthritis in two or more joints of less than three years' duration were categorised at entry as UPA or as rheumatoid arthritis (RA) based on the clinical diagnosis of the rheumatologist. Outcome variables after one year were radiographic damage (Sharp/van der Heijde score) and functional status (Health Assessment Questionnaire: HAQ score). A progressive disease at one year was defined as radiographic progression > or =4, or one year radiographic damage > or =10, or HAQ score > or =1. The baseline variables of patients with UPA with a progressive or mild outcome were compared. RESULTS: 280 patients (70% women; median age 56 years (range 18-90), median duration of symptoms 3.5 months) were included. 203 (72%) patients were clinically diagnosed as having RA and 77 (27%) as having UPA. The group of patients with progressive UPA (n=32 (42%)) had a significantly higher mean age, prevalence of arthritis of the hands, and disease activity (DAS28) at the first visit compared with the patients of the mild UPA group (n=45 (58%)). The RA group had significantly more frequent serum IgM-RF positivity, higher mean disease activity (DAS28) and mean C reactive protein concentration, more frequent symmetric arthritis, and arthritis in more than three joint groups than the progressive UPA group. Six (19%) of the progressive UPA group versus eight (4%) of the RA group did not receive disease modifying antirheumatic drugs during the first year. CONCLUSIONS: After one year of follow up, 32 (42%) of the patients with UPA had a progressive disease. A progressive outcome was associated with older age, higher disease activity, and arthritis of the hands at baseline. To avoid undertreatment of patients with UPA, treatment should be based on severity rather than on diagnosis. PMID- 12117676 TI - Interleukin 17 induces cartilage collagen breakdown: novel synergistic effects in combination with proinflammatory cytokines. AB - OBJECTIVE: To investigate whether interleukin 17 (IL17), derived specifically from T cells, can promote type II collagen release from cartilage. The ability of IL17 to synergise with other proinflammatory mediators to induce collagen release from cartilage, and what effect anti-inflammatory agents had on this process, was also assessed. METHODS: IL17 alone, or in combination with IL1, IL6, oncostatin M (OSM), or tumour necrosis factor alpha (TNFalpha), was added to bovine nasal cartilage explant cultures. Proteoglycan and collagen release were determined. Collagenolytic activity was determined by bioassay. Chondroprotective effects of IL4, IL13, transforming growth factor beta1 (TGFbeta1) and insulin-like growth factor-1 (IGF1) were assessed by inclusion in the explant cultures. RESULTS: IL17 alone stimulated a dose dependent release of proteoglycan and type II collagen from bovine nasal cartilage explants. Suboptimal doses of IL17 synergised potently with TNFalpha, IL1, OSM, and IL6 to promote collagen degradation. This collagen release was completely inhibited by tissue inhibitor of metalloproteinase-1 and BB-94 (a synthetic metalloproteinase inhibitor), and was significantly reduced by IL4, IL13, TGFbeta1, and IGF1. In IL17 treated chondrocytes, mRNA expression for matrix metalloproteinase (MMP)-1, MMP-3, and MMP-13 was detected. Moreover, a synergistic induction of these MMPs was seen when IL17 was combined with other proinflammatory cytokines. CONCLUSIONS: IL17 can, alone and synergistically in combination with other proinflammatory cytokines, promote chondrocyte mediated MMP dependent type II collagen release from cartilage. Because levels of all these proinflammatory cytokines are raised in rheumatoid synovial fluids, this study suggests that IL17 may act as a potent upstream mediator of cartilage collagen breakdown in inflammatory joint diseases. PMID- 12117677 TI - Effect of HLA-B and HLA-DR genes on susceptibility to and severity of spondyloarthropathies in Mexican patients. AB - OBJECTIVE: To investigate the role of HLA-B and HLA-DR genes as contributors to genetic susceptibility and clinical expression of the spondyloarthropathies (SpA) in the Mexican population. METHODS: The study included 172 patients with SpA (undifferentiated SpA 83, ankylosing spondylitis (AS) 64, and reactive arthritis 25) and 99 healthy controls. The HLA-B and HLA-DR alleles were detected by the polymerase chain reaction with sequence-specific primers technique. Patient assessment included demographic data, diagnostic categories, and disease patterns. Statistical methods included the Mantel-Haenzel chi(2) test, Fisher's exact test, and Woolf method for odds ratio (OR). Differences of continuous variables between HLA allele groups were calculated by Student's t test. RESULTS: Increased frequencies of HLA-B27 (pCh10(-3), OR=28.7), HLA-DR1 (pC=0.045, OR=2.77), and HLA-B15 (p=0.034, pC=NS, OR=2.04) alleles in the whole group were found. HLA-B27 strength of association (OR) was 41.4 in AS; 20.9 in undifferentiated SpA; 27.2 in reactive arthritis. HLA-DR1 and HLA-B15 were increased in undifferentiated SpA (pC=0.045, OR=2.98 and p=0.004, pC=NS, OR=2.75). By analysing 58 HLA-B27 negative patients it was found that HLA-B15 and HLA-DR1 associations with SpA were independent of HLA-B27; increased frequencies of HLA-B15 were found in the whole SpA group and in patients with undifferentiated SpA (pC=0.03, OR=3.09 and pCh0.01, OR=3.77) and of HLA-DR1 in the latter (p=0.04, pC=NS, OR=3.15). HLA-B27 positive patients were younger than HLA-B27 negative patients at onset (p=0.03), but HLA-DR1 positive patients were older than HLA-DR1 negative patients (p=0.03). Bath indices for disease activity and functioning were higher in HLA-B27 positive patients (p=0.006 and p=0.004 v HLA-B27 negative patients). In contrast, neither HLA-DR1 nor HLA-B15 influenced these indices. CONCLUSION: Apart from HLA-B27, there is a significant association of HLA-DR1 and HLA-B15 with SpA in Mexicans which is independent of B27. HLA-B27 is associated with younger age at onset and increased disease severity and HLA DR1 with older age at onset. The strength of HLA-B15, HLA-B27, and HLA-DR1 associations varied in different forms of SpA. PMID- 12117678 TI - Standardised nomenclature for glucocorticoid dosages and glucocorticoid treatment regimens: current questions and tentative answers in rheumatology. AB - In rheumatology and other medical specialties there is a discrepancy between the widespread use and the imprecise designation of glucocorticoid treatment regimens. Verbal descriptions of glucocorticoid treatment regimens used in various phases of diseases vary between countries and institutions. Given this background, a workshop under the auspices of the EULAR Standing Committee on International Clinical Studies including Therapeutic Trials was held to discuss this issue and to seek a consensus on nomenclature for glucocorticoid treatment. This report summarises the panel's discussion and recognises that answers derived from consensus conferences are not definitive. Nevertheless, recommendations on glucocorticoid treatment are presented that (1) reflect current and best knowledge available and (2) take into account current clinical practice. A question-answer rationale presentation style has been chosen to convey the messages, to summarise the meeting in a readable format, and to avoid dogmatism. PMID- 12117680 TI - Anti-tumour necrosis factor (TNF) alpha treatment of rheumatoid arthritis (infliximab) selectively down regulates the production of interleukin (IL) 18 but not of IL12 and IL13. AB - OBJECTIVE: To measure interleukin (IL)18 serum concentrations in patients with rheumatoid arthritis (RA) undergoing infliximab treatment (tumour necrosis factor (TNF) alpha blockade) and to evaluate the concomitant modification of IL12 and IL13 serum concentrations, two cytokines belonging to the Th1 and Th2 profile respectively and biologically related to IL18. METHODS: Ten patients with RA not responding to disease modifying antirheumatic drugs (DMARDs) received intravenous infliximab at a dose of 3 mg/kg at baseline and after two and six weeks. Serum samples were collected from all patients before each infusion and assayed for IL18, IL12, and IL13 by enzyme linked immunosorbent assay (ELISA); IL18 was also measured eight weeks after the last infusion. RESULTS: Serum concentrations of IL18 in all patients were already markedly reduced from baseline after two weeks (p<0.005). Serum IL18 was also decreased in a stable manner after six (p<0.01) and 14 weeks (p<0.01) compared with baseline concentrations. No significant modifications were found in serum concentrations of IL12 and IL13 at any time point. CONCLUSION: There was a rapid and persistent decrease in serum concentrations of IL18 in all the patients studied. This result provides evidence of an in vivo regulation of IL18 by TNFalpha and suggests that anti-TNFalpha therapy is likely to interrupt the synergistic effect between these two cytokines. PMID- 12117681 TI - Serum interleukin 18 and interleukin 18 binding protein in rheumatoid arthritis. AB - OBJECTIVE: To measure serum interleukin 18 (IL18) and IL18 binding protein (IL18BP) levels in patients with inflammatory arthropathies, and to identify associations with disease status and the response to treatment. METHODS: Serum samples were obtained before and after methotrexate treatment from patients with rheumatoid arthritis (RA) and psoriatic arthritis (PsA) attending an early arthritis clinic. IL18 and IL18BP were measured by enzyme linked immunosorbent assay (ELISA). RESULTS: Sixty patients with RA and 13 with PsA were evaluated. Serum IL18 levels were significantly higher in RA than in PsA (p<0.001). After six months' treatment with methotrexate, IL18 levels were reduced, but the differences were not significant (p=0.052). In cross sectional analyses, no correlations between IL18 levels and measures of disease activity or structural damage in RA were found. In longitudinal analyses, no correlations between IL18 levels and the response to treatment or the degree of progressive joint damage were found. Similarly, IL18BP levels were raised in RA, and were not associated with measures of the clinical status or the response to treatment. CONCLUSION: Raised serum levels of IL18 are consistent with a pathophysiological role in RA. However, in this study measurement of circulating IL18 and IL18BP did not correlate significantly with clinical measures of disease activity or the response to treatment in patients with early RA. PMID- 12117683 TI - An eight year prospective study of outcome prediction by antiperinuclear factor and antikeratin antibodies at onset of rheumatoid arthritis. AB - OBJECTIVES: Antikeratin antibodies (AKA) and antiperinuclear factor (APF) are specific antibodies found very early in rheumatoid arthritis (RA). The objective of this eight year follow up study was to assess the value of early AKA and APF assays in predicting functional disability and cartilage damage. METHOD: In 2000, 64 patients tested for AKA and APF (antifilaggrin antibodies) between 1990 and 1993 during evaluation of early symmetric oligoarthritis or polyarthritis (suspected RA) were invited to participate in this non-concurrent cohort study. The Health Assessment Questionnaire (HAQ) score, disease activity score (DAS), and Larsen radiographic score were the primary evaluation criteria. RESULTS: Twenty nine patients were re-evaluated. Clinical and laboratory data obtained in 1993 were similar in this group and in the 35 other patients. Twenty five patients had received a diagnosis of RA. Nine (31%) were rheumatoid factor (RF) positive, nine (31%) were AKA positive, and six (21%) were APF positive during the first year of the disease. APF was correlated with the Larsen score (p=0.011) and DAS (p=0.035) evaluated after a mean disease duration of 8.55 years. All APF positive patients had erosive disease. AKA was correlated with the DAS (p=0.035). CONCLUSION: The presence of AKA or APF early in the course of RA was associated with the DAS or Larsen score eight years later. The number of patients was small, but the findings confirmed those of studies with shorter follow ups. Antifilaggrin antibodies should be included in the initial investigation of patients with RA and, when positive, should alert to a high risk of poor outcomes. PMID- 12117682 TI - Addition of interleukin 1 (IL1) and IL17 soluble receptors to a tumour necrosis factor alpha soluble receptor more effectively reduces the production of IL6 and macrophage inhibitory protein-3alpha and increases that of collagen in an in vitro model of rheumatoid synoviocyte activation. AB - OBJECTIVES: To evaluate the usefulness of combination treatment with cytokine inhibitors. METHODS: A simplified model was set up to evaluate the effect of tumour necrosis factor alpha (TNFalpha) soluble receptors (sTNFR) used alone and in combination with soluble interleukin 1 receptor (sIL1R) and sIL17R on the production of markers of inflammation (IL6), of migration of dendritic cells (macrophage inhibitory protein-3alpha (MIP-3alpha)), and of matrix synthesis (C propeptide of type 1 collagen (P1CP)). Synoviocytes were stimulated with supernatants of activated peripheral blood mononuclear cells (PBMC) from patients with rheumatoid arthritis (RA). Soluble receptors (sR) were preincubated at 1 gammag/ml alone or in combination with the supernatants before addition to RA synoviocytes. IL6, MIP-3alpha, and P1CP production was measured by enzyme linked immunosorbent assay (ELISA) in 48 hour synoviocyte supernatants. RESULTS: IL6 production decreased by 16% with sTNFR alone compared with no sTNFR (p<0.001) and by 41% with the combination of the three sR (p<0.001). MIP-3alpha production decreased by 77% with sTNFR alone compared with no sTNFR (p<0.001) and by 98% with the combination of the three sR (p<0.001). In the presence of sTNFR alone, P1CP production increased by 25% compared with no sR (p<0.01). The combination of the three sR increased P1CP production by 48% (p<0.01). CONCLUSION: The effect of sTNFR on IL6, MIP-3alpha, and P1CP production by RA synoviocytes stimulated by activated PBMC supernatants was further enhanced when combined with sIL1R and sIL17R. PMID- 12117684 TI - Histamine stimulates matrix metalloproteinase-3 and -13 production by human articular chondrocytes in vitro. AB - OBJECTIVES: To determine the effects of histamine on matrix metalloproteinase (MMP) production by human articular chondrocytes (HAC) in vitro. METHODS: Conditioned culture medium from HAC cultures incubated with and without 20 microM histamine was assayed by enzymne linked immunosorbent assay (ELISA) for MMP-1, MMP-8, MMP-13 (collagenases 1, 2, and 3, respectively) and MMP-3 (stromelysin). Monolayer cultures of HAC were also immunostained for MMP-13 and MMP-3. RESULTS: The HAC cultures showed a significant increase in MMP-13 and MMP-3 production (2.2- and 1.9-fold, respectively) after treatment with 20 microM histamine for 24 hours, but MMP-1 and MMP-8 were unaffected. All cultures showed MMP-13 and MMP-3 detectable by immunolocalisation. MMP-3 was the more prominent enzyme as shown by both ELISA and immunolocalisation techniques. CONCLUSIONS: Histamine exposure increased both MMP-13 and MMP-3 production by HAC in vitro, thereby suggesting a pathophysiological role in the chondrocytic phenotype associated with degenerative changes in osteoarthritis. PMID- 12117685 TI - Expression of ferritin, transferrin receptor, and non-specific resistance associated macrophage proteins 1 and 2 (Nramp1 and Nramp2) in the human rheumatoid synovium. AB - OBJECTIVE: To gain a better understanding of how iron accumulates in human rheumatoid synovium. METHODS: The distribution of ferritin, transferrin receptor, and non-specific resistance associated macrophage proteins 1 and 2 (Nramp1 and Nramp2) in the human rheumatoid synovium was investigated by immunocytochemistry and reverse transcriptase-polymerase chain reaction (RT-PCR). RESULTS: Both heavy and light ferritin subunit types were detected in the lining layer and the subinitimal zone of rheumatoid synovium, heavy ferritin generally being more abundant than light. Both heavy and light ferritin were detected in isolated synovial macrophages and fibroblasts. Transferrin receptor expression was largely confined to fibroblasts of the synovial lining layer. Nramp2 was detected by PCR in both isolated synovial macrophages and fibroblasts, whereas Nramp1 was detected by PCR and immunocytochemistry in macrophages and neutrophils in the lining and subinitimal zone, and in inflammatory infiltrates, but was absent from fibroblasts. CONCLUSION: A complex chain of events, perhaps initiated by proinflammatory cytokines, may culminate in a toxic build up of iron in the rheumatoid joint. PMID- 12117686 TI - Genetic testing for haemochromatosis in patients with chondrocalcinosis. AB - Hereditary haemochromatosis (HH) is the most common lethal monogenic human disease, affecting roughly 1 in 300 white northern Europeans. Homozygosity for the C282Y polymorphism within the HFE gene causes more than 80% of cases, with compound heterozygosity of the C282Y and H63D polymorphism also increasing susceptibility to disease. The aim of this study was to determine the frequency of the C282Y and H63D polymorphisms in the disease, and to assess the risk of HH in heterozygotes for the C282Y polymorphism. 128 patients were recruited because of either radiographic chondrocalcinosis (at least bicompartmental knee disease or joints other than the knee involved) or CPPD pseudogout. Genotyping of the HFE C282Y and H63D mutations was performed using PCR/SSP and genotypes for the C282Y polymorphism confirmed by PCR/RFLP. Historical white European control data were used for comparison. Two previously undiagnosed C282Y homozygotes (1.6%), and 16 C282Y heterozygotes (12.5%), including four (3.1%) C282Y/H63D compound heterozygotes were identified. This represents a significant overrepresentation of C282Y homozygotes (relative risk 3.4, p=0.037), but the number of heterozygotes was not significantly increased. At a cost per test of pound1 for each subject, screening all patients with chondrocalcinosis using the above ascertainment criteria costs only pound64 for each case of haemochromatosis identified, clearly a highly cost effective test given the early mortality associated with untreated haemochromatosis. Routine screening for haemochromatosis in patients with appreciable chondrocalcinosis is recommended. PMID- 12117687 TI - Patients with systemic lupus erythematosus show a normal responsive search score in exploratory eye movement analysis: comparison with schizophrenia. AB - OBJECTIVE: To assess whether a difference in psychiatric vulnerability exists between patients with systemic lupus erythematosus (SLE) and those with schizophrenia. METHODS: Twenty women with SLE underwent exploratory eye movement analysis, and a responsive search score (RSS) was obtained, two months after the onset of the disease. Fifteen women with schizophrenia in remission also underwent this analysis. Exploratory eye movement was recorded by an eye mark recorder, which detects corneal reflection of infrared light. The number of eye fixations (instance of more than 0.2 seconds of eye fixation time) was recorded, and the RSS was calculated from eye fixation analysis. RESULTS: Mean (SD) RSS differed significantly between patients with SLE and those with schizophrenia (9.85 (1.87) v 7.27 (1.58) points, respectively, p<0.0001), whereas no difference in mean RSS was found between patients with SLE and 19 normal women. No difference in mean RSS was found between patients with active SLE and those with inactive SLE (9.51 (1.87) v 10.0 (1.77) points). CONCLUSION: The psychiatric vulnerability in patients with SLE, measured by the RSS, differs from that in patients with schizophrenia. PMID- 12117688 TI - Necrotising myositis in Behcet's disease: characteristic features on magnetic resonance imaging and a review of the literature. PMID- 12117689 TI - Systemic lupus erythematosus with haemophagocytosis and severe liver disorder. PMID- 12117690 TI - Ultrastructural study of the muscle coat of the gastric wall in a case of systemic sclerosis. PMID- 12117691 TI - Relapse of rheumatoid arthritis after substitution of oral for parenteral administration of methotrexate. PMID- 12117692 TI - Spontaneous spondylodiscitis caused by Klebsiella oxytoca. PMID- 12117693 TI - Elderly onset isolated B27 associated dactylitis. PMID- 12117694 TI - Poor predictive value of antinucleosome and antineutrophil cytoplasmic antibodies in a 270 inception cohort of patients with early naked arthritis of less than one year's duration. PMID- 12117695 TI - Cogan's syndrome with antineutrophil cytoplasmic autoantibody. PMID- 12117696 TI - Haemochromatosis arthropathy and repetitive trauma. PMID- 12117697 TI - Role of HLA genes in familial spondyloarthropathy. PMID- 12117698 TI - Pooled analysis and meta-analysis of glutathione S-transferase M1 and bladder cancer: a HuGE review. AB - Smoking is a known risk factor for bladder cancer. The product of the GSTM1 gene, glutathione S-transferase M1 (GSTM1), is involved in the detoxification of polycyclic aromatic hydrocarbons found in tobacco smoke; a homozygous deletion of this gene in approximately 50% of Caucasians and Asians results in a lack of GSTM1 enzyme activity. Most studies examining the relation between bladder cancer and GSTM1 have reported an increased risk associated with a lack of GSTM1 activity. The authors performed meta- and pooled analyses of published and unpublished, case-control, genotype-based studies that examined this association (17 studies, 2,149 cases, 3,646 controls) and excluded studies conducted in populations with a high prevalence of exposure to known bladder cancer risk factors other than tobacco smoke. Using random effects models in the meta analysis, the authors obtained a summary odds ratio of 1.44 (95% confidence interval (CI): 1.23, 1.68) for GSTM1 null status with all studies included. Results from studies with at least 100 cases and 100 controls produced a summary odds ratio of 1.42 (95% CI: 1.26, 1.60). Pooled analyses using original data sets from 10 studies (1,496 cases and 1,444 controls) and adjusting for age, sex, and race produced similar results. There was no evidence of multiplicative interaction between the GSTM1 null genotype and ever smoking in relation to bladder cancer, although there was a suggestion of additive interaction (additive interaction = 0.45, 95% CI: -0.03, 0.93). These results indicate that, among populations studied to date, GSTM1 null status is associated with a modest increase in the risk of bladder cancer. PMID- 12117699 TI - Associations of body composition with physical performance and self-reported functional limitation in elderly men and women. AB - To understand how body composition relates to functional impairment, the authors examined cross-sectional associations of absolute and relative measures of fat and lean mass with physical performance and self-reported functional limitation. The sample consisted of a community-based cohort of 1,655 older women and men from Sonoma, California, who had complete baseline data in 1993-1994 on body composition, physical performance, and functional limitation. Physical performance was assessed by walking speed and grip strength, while global functional limitation, across several domains, was assessed by self-report using standard questions. Lean mass and fat mass were estimated from bioelectric impedance using population-specific prediction equations derived from dual x-ray energy absorptiometry. Higher fat mass was associated with slower walking speed and greater likelihood of functional limitation, while higher lean mass was generally associated only with increased grip strength. A higher lean mass-to-fat mass ratio, a relative measure of body composition, was associated with faster walking speed and less limitation. These findings suggest that fat mass negatively impacts some domains of physical performance and overall functioning, while lean mass is less significant in absolute terms but is important relative to amount of body fat. PMID- 12117701 TI - Invited commentary: body composition in studies of aging: new opportunities to better understand health risks associated with weight. AB - The expected increase in the proportion of older persons over the next century underscores the need to identify modifiable risk factors for disease and disability in this population. One such risk factor is weight, which plays a role in many of the diseases common in old age and contributes to risk of disability and death. However, there is confusion and controversy regarding health risks associated with weight in old age. The emergence of new technologies to assess body composition should allow opportunities to better understand health risks associated with weight in old age, as suggested by the new report in this issue of the Journal. While application of these technologies to population studies will still require careful attention to methodological caveats important in studies of weight, the ability to separately examine lean mass, bone, and fat should shed light on the underlying biologic processes pertinent to risk. PMID- 12117703 TI - Baseline cataract type and 10-year mortality in the Italian-American Case-Control Study of age-related cataract. AB - Age-related cataract is reported to be associated with increased risk of death. The authors investigated the association of presence and type of cataract with mortality in the participants of the Italian-American Case-Control Cataract Study (Parma, Italy, 1987-1989), which included 1,008 persons aged 45-79 years who had age-related cataract and 469 who had clear lenses. Slit-lamp and retroillumination lens photographs were taken at baseline and graded with the Lens Opacities Classification System II. During 10 years of follow-up (range, 8.9 11.8 years; 11,318 person-years), the authors collected information on 1,429 participants and documented 339 deaths. After adjustment for age, sex, and other mortality risk factors, mixed cataracts with a nuclear/posterior subcapsular component were significantly associated with higher risk of death by Cox proportional hazards regression analyses. Hazard ratios were 2.26 (95% confidence interval (CI): 1.07, 4.76) for nuclear/posterior subcapsular and 1.62 (95% CI: 1.01, 2.61) for cortical/nuclear/posterior subcapsular opacities. In multivariate analysis, mixed types of opacity (any) were associated with increased mortality for malignancy (hazard ratio = 1.81, 95% CI: 1.04, 3.15) and "other" causes (hazard ratio = 2.29, 95% CI: 1.07, 4.92). These findings are compatible with the hypothesis that mixed types of cataract with a nuclear/posterior subcapsular component are indicators of accelerated aging. PMID- 12117704 TI - Body mass index, weight change, and death in older adults: the systolic hypertension in the elderly program. AB - The relation between relative weight and health differs between young and old. In older populations, weight change may cloud the association between a single relative weight and health outcomes. To determine whether weight or weight change is a more important determinant of mortality in a population of older adults, the authors analyzed data from the Systolic Hypertension in the Elderly Program (1984 1990), a randomized clinical trial testing the efficacy of antihypertensive drug treatment to reduce the risk of stroke in older adults (aged 60 years or more) with isolated systolic hypertension. After adjustment for covariates, an average annualized weight loss of at least 1.6 kg/year (odds ratio = 4.9), a weight loss between 1.6 and 0.7 kg/year (odds ratio = 1.7), a weight gain of more than 0.5 kg/year (odds ratio = 2.4), and a baseline body mass index of less than 23.6 (odds ratio = 1.4) all had a significant (p < 0.05) association with all-cause mortality compared with a referent group that was weight stable and of intermediate body mass index (23.6 to <28.0 kg/m(2)) and weight change (-0.7 to <0.5 kg/year). The authors conclude that, in older adults, dynamic measures (e.g., annualized weight change) of weight change predict mortality better than do static weight measures (e.g., baseline body mass index). Even in those with high or low baseline body mass index, weight stability is associated with a lower mortality risk. PMID- 12117705 TI - Physical inactivity is associated with lower forced expiratory volume in 1 second : European Prospective Investigation into Cancer-Norfolk Prospective Population Study. AB - Forced expiratory volume in 1 second (FEV(1)) is a strong risk factor for cardiovascular disease, stroke, lung cancer, and all-cause mortality. One possible explanation for this association is that FEV(1) is a marker of other determinants of mortality risk, such as obesity and physical inactivity. In a population-based cohort study of 12,283 men and women aged 45-74 years from the European Prospective Investigation into Cancer-Norfolk Study recruited in 1993 1997, the cross-sectional association between physical activity and FEV(1) and that between physical activity and change in FEV(1) were analyzed. Indices of physical activity, including participation in vigorous recreational activity, stair climbing, and television viewing, were assessed with a validated questionnaire designed to assess activity in the previous year. Television viewing was negatively associated with FEV(1) in men and women (p < 0.001), whereas stair climbing and participation in vigorous leisure time activities were positively associated with FEV(1) in men and women (p < 0.001). The associations remained after adjustment for known confounders, including age, height, vitamin C, and smoking. Climbing more stairs and participating in vigorous leisure-time activity predicted a slower rate in annual percent decline in FEV(1) (p < 0.004 and p < 0.002, respectively). In conclusion, physical activity is associated with higher levels of FEV(1), whereas television viewing is associated with lower levels. PMID- 12117706 TI - Association of dairy products, lactose, and calcium with the risk of ovarian cancer. AB - Epidemiologic findings have been inconsistent regarding the association of dietary fat, dairy products, and lactose with risk of ovarian cancer. The authors conducted a case-control study in Hawaii and Los Angeles, California, to examine several dietary hypotheses regarding the etiology of ovarian cancer in a population with a broad range of dietary intakes. A total of 558 patients with ovarian cancer diagnosed in 1993-1999 and 607 controls were interviewed regarding their diet. Consumption of all dairy products, all types of milk, and low-fat milk, but not consumption of whole milk, was significantly inversely related to the odds of ovarian cancer. Similar inverse gradients in the odds ratios were obtained for intakes of lactose and calcium, although these nutrients were highly correlated (r = 0.77). The odds ratio for ovarian cancer was 0.46 (95% confidence interval: 0.27, 0.76) among women in the highest quartile of dietary calcium intake versus the lowest (p for trend = 0.0006). The significant dietary association was limited to dairy sources of calcium (p for trend = 0.003), although a nonsignificant inverse gradient in risk was also found in relation to calcium supplement intake. These results suggest that intake of low-fat milk, calcium, or lactose may reduce the risk of ovarian cancer. PMID- 12117707 TI - Human papillomavirus and the long-term risk of cervical neoplasia. AB - The risk of cervical neoplasia for women with normal Papanicolaou smears was calculated for those whose smears were human papillomavirus (HPV) positive and those whose smears were HPV negative. Data on 347 cases and controls were analyzed in a population-based, nested case-control study. Cases (n = 77) were women who participated in the Utrecht screening program (1976-1984) in the Netherlands and who developed cervical intraepithelial neoplasia 3 or microinvasive or invasive squamous cervical cancer after having a negative smear (1980-1986). Controls (n = 270) were matched on age (+/-5 years) and follow-up period. DNA was isolated from the Papanicolaou smears and was tested for the presence of HPV DNA by using the ultrasensitive broad-spectrum, general short fragment polymerase chain reaction. HPV was found in 55 (71%) of the baseline smears of the 77 cases and in 31 (11%) of those of the 270 controls. The age adjusted odds ratios for developing cervical intraepithelial neoplasia or microinvasive or invasive cervical cancer were 19.2 (95 percent confidence interval (CI): 10.3, 35.7) for HPV positivity in general, 5.4 (95% CI: 1.5, 19.5) for infection with low-risk HPV genotypes, 24.0 (95% CI: 12.4, 46.4) for high risk HPV genotypes, and 104.8 (95% CI: 29.5, 372.7) for HPV type 16. PMID- 12117708 TI - Delayed diagnosis of US females with cystic fibrosis. AB - This study was conducted to examine a patient's age and condition at the time of diagnosis as one potential factor contributing to the "gender gap" in cystic fibrosis. The study population consisted of 11,275 US patients diagnosed during 1986-1998 and reported to the Cystic Fibrosis Foundation Registry in the same or the following calendar year. Parallel analyses were performed for Wisconsin patients identified prospectively during 1985-1994 to obtain more detailed information on their condition at diagnosis. Analyses of the registry data showed that females identified because of symptoms other than meconium ileus were diagnosed at significantly older ages (median, 12.7 months) than were males (median, 8.7 months) (p < 0.001). The delay in diagnosis for females was most evident among patients presenting with respiratory symptoms only (median, 40.7 vs. 22.3 months; p < 0.001). Analyses of Wisconsin patients demonstrated no significant gender differences in cough and wheezing experiences or in chest radiographic severity scores between males and females during their first 10 years of life, although a disproportionately high number of males were referred for diagnostic sweat testing. A delay in diagnosis of females with cystic fibrosis was discovered, suggesting either differential recognition of respiratory symptoms or a gender bias. PMID- 12117709 TI - Increased fibrinogen levels among South Asians versus Whites in the United Kingdom are not explained by common polymorphisms. AB - Determinants of fibrinogen level among South Asians are not established. In 1997 1999, plasma fibrinogen levels and prevalences of the fibrinogen polymorphisms A alpha Thr312Ala, beta-445G/A, and B beta Arg448Lys and correlates were compared among 100 apparently healthy United Kingdom South Asians and 100 age- and sex matched Whites. Mean fibrinogen levels were higher in South Asians (3.33 g/liter, 95% confidence interval (CI): 3.16, 3.51) than in Whites (2.84 g/liter, 95% CI: 2.72, 2.96) (p < 0.0001), but genotype distributions were similar. B beta Arg448Lys was related to fibrinogen in South Asians (RR (n = 67): 3.22 g/liter, 95% CI: 3.03, 3.43; RK (n = 26): 3.72 g/liter, 95% CI: 3.65, 4.11; KK (n = 7): 3.07 g/liter, 95% CI: 2.53, 3.72) (p = 0.04) and Whites (p = 0.06). beta-455G/A was related to fibrinogen in Whites (GG (n = 56): 2.68 g/liter, 95% CI: 2.56, 2.86; GA (n = 37): 2.97 g/liter, 95% CI: 2.79, 3.17; AA (n = 5): 3.22 g/liter, 95% CI: 2.85, 3.65) (p = 0.02) and South Asians (p = 0.07). After adjustment for age, gender, body mass index, hypertension, cholesterol, triglycerides, smoking, A alpha Thr312Ala, and beta-455G/A, fibrinogen levels remained significantly higher in South Asians (3.56 g/liter, 95% CI: 3.35, 3.77) than in Whites (3.03 g/liter, 95% CI: 2.85, 3.22) (p < 0.0001). These findings suggest that increased fibrinogen levels among South Asians versus Whites are not due to differences in the prevalence of genetic polymorphisms that encode for fibrinogen. PMID- 12117710 TI - Familial aggregation of fetal growth restriction in a French cohort of 7,822 term births between 1971 and 1985. AB - An association between fetal growth restriction and increased rates of metabolic and cardiovascular diseases in adulthood has been reported. This study evaluated familial aggregation of fetal growth restriction in term births. The population consisted of 3,505 sibships comprised of 7,822 full-term singleton infants born between 1971 and 1985 in Haguenau, France, and selected from a regional register of births. Sib-sib odds ratios were estimated for being born small for gestational age (SGA), defined as having a birth weight below the 10th percentile of the sex-specific curve of birth weight by week of gestation. SGA births were further stratified according to ponderal index (birth weight/length(3)). After adjustment for maternal factors, the sib-sib odds ratios were 4.8 (95% confidence interval (CI): 3.7, 6.3) for all SGA births, 7.7 (95% CI: 4.1, 14.7) for SGA births with a low ponderal index (<10th percentile), and 4.4 (95% CI: 2.3, 8.2) for SGA births with a normal ponderal index (25th-75th percentile). None of the maternal factors investigated significantly influenced the magnitude of these odds ratios. This strong residual sib-sib aggregation suggests a role for genetic and/or shared environmental factors in the etiology of fetal growth restriction, especially when associated with a low ponderal index. PMID- 12117712 TI - Genetic association studies. PMID- 12117713 TI - Association studies of vascular phenotypes: how and why? PMID- 12117714 TI - SNP judgments and freedom of association. PMID- 12117715 TI - Effects of inflammation on high-density lipoproteins. PMID- 12117716 TI - ATVB in focus: noninvasive assessment of atherosclerosis--from structure to function. PMID- 12117718 TI - MRI and characterization of atherosclerotic plaque: emerging applications and molecular imaging. AB - Noninvasive high-resolution magnetic resonance has the potential to image atherosclerotic plaque and to determine its composition and microanatomy. This review summarizes the rationale for plaque imaging and describes the characteristics of plaque by use of existing MRI techniques. The use of MRI in human disease and in animal models, particularly in rabbits and mice, is presented. Present and future applications of MRI, including real-time vascular intervention, new contrast agents, and molecular imaging, are also discussed. PMID- 12117719 TI - Aspirin inhibits Chlamydia pneumoniae-induced nuclear factor-kappa B activation, cytokine expression, and bacterial development in human endothelial cells. AB - OBJECTIVE: Chlamydia pneumoniae has been associated with atherosclerosis. Infection of vascular endothelial cells with C pneumoniae increases the expression of proatherogenic cytokines mediated by nuclear factor (NF)-kappaB, a transcription factor. The present study was designed to test the effect of aspirin on C pneumoniae-induced NF-kappaB activation, interleukin expression, and bacterial development in cultured human endothelial cells. METHODS AND RESULTS: Aspirin, its metabolite salicylic acid, and 2 other unrelated NF-kappaB inhibitors showed a strong concentration-dependent inhibitory effect on chlamydial growth, indicated by the reduction of bacterial inclusions and the titer of infectious progeny. Involvement of the transcription factor NF-kappaB was confirmed by electrophoretic mobility shift assay and by transfection experiments with appropriate decoy oligodeoxynucleotides. Attenuation of the C pneumoniae-induced activation of NF-kappaB by aspirin also reduced the secretion of interleukin-6 and interleukin-8, indicating efficient inhibition of NF-kappaB gene expression. Reduction of chlamydial growth was not caused by apoptosis of the host cell, as determined by monitoring characteristic chromatin condensation. CONCLUSIONS: These data provide evidence that NF-kappaB-mediated gene activation represents a crucial step in the developmental cycle of C pneumoniae. Aspirin exerts an anti-chlamydial effect that is due to the inhibition of C pneumoniae induced NF-kappaB activation, which might account for some of the cardioprotective activity of aspirin. PMID- 12117720 TI - Role of SRC homology 2-containing tyrosine phosphatase 2 on proliferation of rat smooth muscle cells. AB - OBJECTIVE: Src homology 2-containing phosphotyrosine phosphatase 2 (SHP2) is ubiquitously expressed and believed to function as part of a positive signaling pathway mediating growth factor-induced protein tyrosine phosphorylation. Proliferation of aortic vascular smooth muscle cells (SMCs) is an important contributor to atherosclerosis. We examined the effect of SHP2 expression on SMC proliferative activity. METHODS AND RESULTS: SHP2 was abundant in cultured aortic SMCs, and SHP2 staining was markedly increased in the thickened aortic intima in rats with balloon-induced injury. We obtained several SMC clones by using geneticin screening. Endogenous SHP2 expression varied among individual clones. Significant positive relationships were observed between SHP2 expression and bromodeoxyuridine uptake in SMCs stimulated by FBS, platelet-derived growth factor, or insulin-like growth factor-1. In SMCs transiently transfected with SHP2, FBS stimulation significantly increased bromodeoxyuridine uptake beyond the uptake by control SMCs. CONCLUSIONS: Increased SHP2 expression in SMCs may accelerate aortic atherosclerosis by increasing cell growth. PMID- 12117721 TI - NO attenuates insulin signaling and motility in aortic smooth muscle cells via protein tyrosine phosphatase 1B-mediated mechanism. AB - OBJECTIVE: Hyperinsulinemia is a significant risk factor for the pathogenesis of vascular disease. Protein tyrosine phosphatase 1B (PTP1B) has been recognized as a modulator of insulin signaling in nonvascular cells, and we have recently reported that NO increases the activity of PTP1B in rat vascular smooth muscle cells. In the present study, we tested the hypothesis that NO attenuates insulin stimulated cell motility via a PTP1B-mediated mechanism involving downregulation of insulin signal transduction. METHODS AND RESULTS: Treatment of primary aortic smooth muscle cells from newborn rats with the NO donor S-nitroso-N acetylpenicillamine reduced cell motility, tyrosine phosphorylation levels of insulin receptor beta subunit and insulin receptor substrate-1, and extracellular signal-regulated kinase activity. Overexpression of wild-type PTP1B via an adenoviral vector blocked the capacity of insulin to stimulate cell motility and insulin receptor phosphorylation, whereas expression of a dominant-negative mutant of PTP1B attenuated the capacity of NO to decrease cell motility. CONCLUSIONS: Our findings indicate that activation of PTP1B is necessary and sufficient to account for the capacity of NO to decrease insulin-stimulated signal transduction and cell motility in cultured aortic smooth muscle cells. The results could explain the capacity of NO to oppose neointima formation in states of hyperinsulinemia. PMID- 12117723 TI - Flow-dependent remodeling in the carotid artery of fibroblast growth factor-2 knockout mice. AB - OBJECTIVE: Fibroblast growth factor-2 (FGF2) has been implicated as a mediator in the structural remodeling of arteries. Chronic changes in blood flow are known to cause reorganization of the vessel wall, resulting in permanent changes in artery size (flow-dependent remodeling). Using FGF2 knockout (Fgf2(-/-)) mice, we tested the hypothesis that FGF2 is required during flow-dependent remodeling of the carotid arteries. METHODS AND RESULTS: All branches originating from the left common carotid artery (LCCA), except for the left thyroid artery, were ligated to reduce flow in the LCCA and increase flow in the contralateral right common carotid artery (RCCA). Age- and sex-matched control animals did not undergo ligation of the LCCA branches. Morphometric analysis showed that by day 7, vessel diameter was significantly greater in the high-flow RCCA of FGF2 wild-type (Fgf2(+/+)) and Fgf2(-/-) mice versus the respective control RCCA, demonstrating outward remodeling. In contrast, vessel diameter was decreased by day 7 in the low-flow LCCA of both genotypes compared with the control LCCA, showing inward remodeling. No differences were observed between Fgf2(+/+) and Fgf2(-/-) mice in either high-flow or low-flow remodeling. CONCLUSIONS: Given these results, we demonstrate that FGF2 is not essential for flow-dependent remodeling of the carotid arteries. PMID- 12117722 TI - Heterogeneity of smooth muscle cell populations cultured from pig coronary artery. AB - OBJECTIVE: Heterogeneous smooth muscle cell (SMC) populations have been described in the arteries of several species. We have investigated whether SMC heterogeneity is present in the porcine coronary artery, which is widely used as a model of restenosis. METHODS AND RESULTS: By using 2 isolation methods, distinct medial populations were identified: spindle-shaped SMCs (S-SMCs) after enzymatic digestion, with a "hill-and-valley" growth pattern, and rhomboid SMCs (R-SMCs) after explantation, which grow as a monolayer. Moreover, the intimal thickening that was induced after stent implantation yielded a large proportion of R-SMCs. R-SMCs exhibited high proliferative and migratory activities and high urokinase activity and were poorly differentiated compared with S-SMCs. Heparin and transforming growth factor-beta2 inhibited proliferation and increased differentiation in both populations, whereas fibroblast growth factor-2 and platelet-derived growth factor-BB had the opposite effect. In addition, S-SMCs treated with fibroblast growth factor-2 or platelet-derived growth factor-BB or placed in coculture with coronary artery endothelial cells acquired a rhomboid phenotype. This change was reversible and was also observed with S-SMC clones, suggesting that it depends on phenotypic modulation rather than on selection. CONCLUSIONS: Our results show that 2 distinct SMC subpopulations can be recovered from the pig coronary artery media. The study of these subpopulations will be useful for understanding the mechanisms of restenosis. PMID- 12117724 TI - Opposing functions of the Ets factors NERF and ELF-1 during chicken blood vessel development. AB - OBJECTIVE: The purpose of this study was to evaluate the role of the Ets factor NERF in the regulation of the Tie1 and Tie2 genes during chicken blood vessel development. METHODS AND RESULTS: We have isolated the full-length cDNA for the chicken homologue of the human Ets factor NERF2 (cNERF2). Northern blot analysis and in situ hybridization demonstrate that cNERF2 is enriched in the developing blood vessels of the chicken chorioallantoic membrane. Interestingly, cNERF2 functions as a competitive inhibitor of a highly related Ets factor cELF-1, which we have previously shown to be enriched in chicken blood vessel development. Although in vitro-translated cELF-1 and cNERF2 can bind equally well to conserved Ets binding sites in the promoters of the Tie1 and Tie2 genes, cELF-1 preferentially binds to the Ets sites in these promoters during early stages of chicken blood vessel development, suggesting that cNERF may bind during later stages of blood vessel development and vascular remodeling. CONCLUSIONS: cNERF2 is enriched during embryonic and extraembryonic blood vessel development in the chicken and facilitates tight control of Tie1 and Tie2 gene regulation. PMID- 12117725 TI - Angiotensin II type 1 and 2 receptors in conduit arteries of normal developing microswine. AB - OBJECTIVE: To identify vascular cells capable of responding to angiotensin II (Ang II) generated in conduit arteries, we examined the Ang II type 1 receptor (AT1R) and Ang II type 2 receptor (AT2R) in the thoracic aorta (TA) and abdominal aorta (AA) and branches in 90-day fetal, 3-week postnatal, and 6-month adult microswine. METHODS AND RESULTS: By autoradiography ((125)I-[Sar(1)Ile(8)]-Ang II with or without AT1R- or AT2R-selective analogues or (125)I-CGP 42112), there were striking rostrocaudal differences in (1) AT2R binding at all ages (prominent in AA wall and branches, sparse in TA wall and branches) and (2) a non-AT2R binding site for CGP 42112 (consistently evident in postnatal TA and branches but absent in AA and branches). Furthermore, patterns of AT2R distribution in infradiaphragmatic arteries were developmentally distinct. In fetal AAs, high density AT2Rs occupied the inner 60% of the medial-endothelial wall. In postnatal AAs, AT2Rs were sparse in the medial-endothelial wall but prominent in a circumferential smooth muscle alpha-actin-negative cell layer at the medial adventitial border, occupying approximately 20% to 25% of the AA cross-sectional area. AT1R density in the TA and AA medial-endothelial wall increased with age, whereas AT2R density decreased after birth. CONCLUSIONS: A novel AT2R-positive cell layer confined to postnatal infradiaphragmatic arteries physically links adventitial and medial layers, appears optimally positioned to transduce AT2R dependent functions of local Ang II, and suggests that adventitial Ang II may elicit regionally distinct vascular responses. PMID- 12117726 TI - Saphenous vein endothelin system expression and activity in African American patients. AB - OBJECTIVE: Plasma endothelin (ET)-1 levels are significantly higher in African American hypertensive patients than in white hypertensive patients. However, whether the molecular components of vascular ET-1 biosynthesis and function are altered in this population remains to be established. Accordingly, the overall goal of this study was to investigate the effects of race on vascular mRNA and protein levels of ET-converting enzyme (ECE)-1 subisoforms, ET-1, and ET receptor profiles in hypertension. METHODS AND RESULTS: Saphenous vein samples were obtained from African American (n=13) and white (n=15) patients undergoing coronary artery grafting surgery. The expression of preproET-1 and of ECE-1a was upregulated approximately 2- and 3-fold, respectively, in African Americans. In endothelium-intact vessels, the ET(A) expression was higher in whites. In endothelium-denuded vessels, the ET(B) mRNA was 3-fold higher in African Americans, suggesting that vasoconstriction-promoting ET(B) receptors are upregulated in this population. Vascular tissue ET-1 levels and ECE-1 activity were also augmented in African American patients. CONCLUSIONS: This study demonstrated that the biosynthetic pathway of ET-1 is activated to a higher degree and that the ET(B) receptor subtype expression is altered in the peripheral vasculature of African American hypertensive patients. The augmented synthesis and altered expression of ET(B) receptors may both contribute to the increased incidence of hypertension and related complications in this patient population. PMID- 12117727 TI - Mechanism of accumulation of cholesterol and cholestanol in tendons and the role of sterol 27-hydroxylase (CYP27A1). AB - OBJECTIVE: Tendon xanthomas are deposits of lipids and connective tissue commonly found in hypercholesterolemic patients. Macrophages are likely to be responsible for the lipid accumulation. Normolipidemic patients with the rare disease cerebrotendinous xanthomatosis, lacking the enzyme sterol 27-hydroxylase (CYP27A1), develop prominent xanthomas in tendons and brain containing both cholestanol and cholesterol, with a cholestanol:cholesterol ratio higher than that in the circulation. Because of its ability to convert cholesterol into polar metabolites that leave the cells faster, CYP27A1 has been suggested to be an antiatherogenic enzyme. The hypothesis was tested that tendons contain CYP27A1 that may be of importance for the normal efflux of both steroids. METHODS AND RESULTS: Western blotting and combined gas chromatography-mass spectrometry showed that human tendons contain significant amounts of CYP27A1 and its product, 27-hydroxycholesterol. Immunohistochemistry showed that CYP27A1 is present in macrophages and tenocytes. The tendons also contained cholestanol, with a cholestanol:cholesterol ratio slightly higher than that in the circulation. Recombinant human CYP27A1, and cultured human macrophages containing this enzyme, had similar activity toward cholesterol and cholestanol. After loading of macrophages with labeled cholesterol and cholestanol, there was an efflux of these steroids in both unmetabolized and 27-oxygenated form, resulting in a significant cellular accumulation of cholestanol compared with cholesterol. CONCLUSION: The results are consistent with the possibility that CYP27A1 is of importance for the efflux of both cholesterol and cholestanol from tendons. PMID- 12117728 TI - Prospective study of effect of androgens on serum inflammatory markers in men. AB - OBJECTIVE: Because male sex is an independent risk factor for the severity of atherosclerosis, it is possible that androgens may be proatherogenic. There is evidence that sex hormones, particularly estrogens, regulate (or modulate) inflammation, a process integral to atherogenesis. Because levels of serum inflammatory markers predict cardiovascular outcomes, we prospectively assessed the effects of androgen therapy on these markers in older men. METHODS AND RESULTS: Levels of high-sensitivity C-reactive protein (CRP), soluble intracellular adhesion molecule-1 (sICAM-1), and soluble vascular cell adhesion molecule-1 (sVCAM-1) were measured from sera collected at baseline and at the end of 2 randomized double-blind placebo-controlled trials evaluating the effects of 3 months of androgen treatment with either dihydrotestosterone (DHT) or recombinant human chorionic gonadotropin (rhCG) in healthy men aged >60 years with partial androgen deficiency (serum testosterone levels <15 nmol/L). For the DHT study (70 mg transdermally daily), 33 men completed 3 months of treatment (16 men were treated with DHT, and there were 17 controls). For the rhCG (250 microg twice weekly) study, 20 men were treated with rhCG, and there were 20 controls. In both studies, groups were well matched for age and vascular risk factors. Androgen levels (DHT and testosterone) were consistently maintained at eugonadal levels throughout the trials, with estradiol markedly increased by rhCG but not DHT. Baseline CRP levels were 0.74 to 1.49 mg/L, sVCAM-1 levels were 847 to 950 ng/mL, and sICAM-1 levels were 256 to 292 ng/mL in all groups. Neither DHT nor rhCG resulted in significant changes in CRP, sVCAM-1, or sICAM-1 compared with placebo (P>0.3 in both studies). CONCLUSIONS: Exogenous androgen therapy with or without increased estradiol levels does not alter serum inflammatory markers in older men; this finding is in contrast to the effects of estrogens on inflammatory markers that have been found in postmenopausal women. These data provide a measure of reassurance concerning potential adverse cardiovascular effects of androgen therapy in older men. PMID- 12117729 TI - Genetic and environmental influences on lipids, lipoproteins, and apolipoproteins: effects of menopause. AB - OBJECTIVE: Levels of lipids and (apo)lipoproteins are known to increase after menopause, but it is unknown whether the genetic and environmental variability alters or whether lipids and (apo)lipoproteins are influenced by different genes before and after menopause. METHODS AND RESULTS: We studied 453 monozygotic and 1280 dizygotic pairs of female white twins recruited from the St. Thomas' UK Adult Twin Registry and measured total cholesterol, low density lipoprotein (LDL), high density lipoprotein (HDL), triglycerides, lipoprotein(a) [Lp(a)], apolipoprotein A1 (apoA1), and apolipoprotein B (apoB). Variance components software was used to estimate genetic and environmental influences on serum lipid levels in premenopausal and postmenopausal women. Total variance was higher for triglycerides, HDL, and apoB after menopause. Postmenopausal women showed larger genetic variance for most lipids, apart from apoB and Lp(a). In premenopausal females, total cholesterol, LDL, HDL, apoA1, and apoB all showed an influence of the shared environment (22% to 34%), which, after menopause, decreased in HDL and completely disappeared in total cholesterol, LDL, and apoA1. Only for Lp(a), with a high heritability of 87%, did the same model fit premenopausal and postmenopausal women. Generally, there was no indication that different genes influence lipids before and after menopause. CONCLUSIONS: These findings imply that genetic studies of lipids can pool results from premenopausal and postmenopausal women and that family-based interventions, such as changes in diet, are more likely to succeed in younger women, in whom the environmental influences are greater. PMID- 12117730 TI - Cholesteryl ester transfer protein TaqI B2B2 genotype is associated with higher HDL cholesterol levels and lower risk of coronary heart disease end points in men with HDL deficiency: Veterans Affairs HDL Cholesterol Intervention Trial. AB - OBJECTIVE: We have previously reported that genetic variation at the cholesteryl ester transfer protein (CETP) TaqIB locus is correlated with plasma lipid levels and coronary heart disease (CHD) risk in the Framingham Offspring Study (FOS). In FOS, the B2 allele was associated with increased levels of high density lipoprotein (HDL) cholesterol (HDL-C), decreased CETP activity, and reduced CHD risk for men having the B2B2 genotype. The present study was undertaken to further define the relationship between this polymorphism and CHD risk at the population level. METHODS AND RESULTS: We tested for associations between the CETP TaqIB genotype and plasma lipoprotein levels, response to gemfibrozil therapy, and CHD end points in 852 men participating in the Veterans Affairs HDL C Intervention Trial (VA-HIT), a study designed to explore the potential benefits of raising HDL levels in men having established CHD with low HDL-C (< or =40 mg/dL) as their primary lipid abnormality. In VA-HIT, 13.9% of the men had the B2B2 genotype relative to 19.1% of the men in FOS (-27%, P<0.03), whereas more men in VA-HIT had the B1B1 genotype (15%, P<0.05). Similar to our finding in FOS, B2B2 men in VA-HIT had the highest mean level of HDL-C (32.6+/-4.8 mg/dL), followed by B1B2 men (32.0+/-5.3 mg/dL), and, last, by B1B1 men (30.9+/-4.9 mg/dL). Interestingly, B1B1 men, who had the least favorable plasma lipid profile at baseline, had the greatest triglyceride-lowering response to gemfibrozil ( 34%, P=0.006). CETP TaqIB genotype was also associated with the risk of CHD end points in VA-HIT, with an adjusted risk ratio of 0.52 for B2B2 men (P=0.08). CONCLUSIONS: Our data demonstrate that in men with CHD and HDL deficiency, the CETP TaqI B2B2 genotype is (1) significantly reduced and (2) associated with higher levels of plasma HDL-C and lower CHD risk. Together with our earlier report, these results support the concept that increased HDL-C levels, resulting from reduced CETP activity, are associated with decreased CHD risk. PMID- 12117731 TI - Value of HDL cholesterol, apolipoprotein A-I, lipoprotein A-I, and lipoprotein A I/A-II in prediction of coronary heart disease: the PRIME Study. Prospective Epidemiological Study of Myocardial Infarction. AB - OBJECTIVE: We have examined the association between the incidence of coronary heart disease (CHD) and plasma high density lipoprotein (HDL) cholesterol, apolipoprotein A-I (apoA-I), and 2 HDL fractions, lipoprotein A-I and lipoprotein A-I:A-II. METHODS AND RESULTS: These parameters were measured in subjects recruited in France and in Northern Ireland in the Prospective Epidemiological Study of Myocardial Infarction (PRIME) Study, a prospective cohort study. Among the subjects free of CHD on entry, 176 in France and 113 in Northern Ireland suffered an ischemic attack (CHD patients) during the 5-year follow-up, whereas 6612 French and 2172 Northern Irish men showed no CHD symptoms (CHD-free subjects). All 4 HDL parameter levels were lower in CHD patients than in CHD-free subjects. After the cohort was divided into quintiles based on the distribution of HDL parameter levels, a significant (P<0.0001) linear increase in relative risk was observed for each HDL parameter level. However, regression logistic analyses showed that apoA-I was the strongest predictor (more powerful than HDL cholesterol) and that lipoprotein A-I and lipoprotein A-I:A-II did not supplement apoA-I in predicting CHD. CONCLUSIONS: Among the parameters related to HDL, apoA I appears to be the strongest independent risk factor. PMID- 12117732 TI - Circulating oxidized LDL is associated with subclinical atherosclerosis development and inflammatory cytokines (AIR Study). AB - OBJECTIVE: Circulating oxidized LDL (Ox-LDL) is associated with clinical manifestations of atherosclerosis. However, no previous study has examined the relationship between subclinical atherosclerosis and Ox-LDL. The aims of the present study were to investigate the relationship between clinically silent ultrasound-assessed atherosclerotic changes in the carotid and femoral arteries and Ox-LDL and to explore the relationship between Ox-LDL, C-reactive protein, and the inflammatory cytokines interleukin-6 and tumor necrosis factor-alpha. METHODS AND RESULTS: The study group (n=391) consisted of clinically healthy, 58 year-old men recruited from the general population. Ox-LDL was measured by using a specific monoclonal antibody, mAb-4E6. The results showed that Ox-LDL was related to intima-media thickness and plaque occurrence in the carotid and femoral arteries. In addition, Ox-LDL was associated with tumor necrosis factor alpha and C-reactive protein. Circulating Ox-LDL was also associated with LDL cholesterol but not with blood pressure or smoking. When adjusting for other risk factors, both LDL cholesterol and Ox-LDL seemed to be independent predictors of plaque occurrence in the carotid and femoral arteries (odds ratios for quintile 5 versus quintile 1 were 2.17, P=0.049 and 2.25, P=0.050, for LDL cholesterol and Ox-LDL, respectively). CONCLUSIONS: Ox-LDL was associated with both subclinical atherosclerosis and inflammatory variables, supporting the concept that oxidatively modified LDL may play a major role in atherosclerosis development, although no causality can be shown in this cross-sectional study. PMID- 12117733 TI - Transvascular low-density lipoprotein transport in patients with diabetes mellitus (type 2): a noninvasive in vivo isotope technique. AB - OBJECTIVE: The increased risk of atherosclerosis associated with diabetes cannot be explained by conventional cardiovascular risk factors alone. We hypothesized that transvascular lipoprotein transport may be increased in patients with diabetes, possibly explaining increased intimal lipoprotein accumulation and, thus, atherosclerosis. METHODS AND RESULTS: We developed an in vivo method for measurement of transvascular transport of low density lipoprotein (LDL) and applied it in 16 patients with maturity-onset diabetes (type 2) and 29 healthy control subjects. Autologous 131I-labeled LDL was reinjected intravenously in addition to 125I-labeled albumin, and the 1-hour fractional escape rates were taken as indices of transvascular transport. Both parameters were normally distributed, and they were tightly correlated (R2=0.69, P<0.0001). Transvascular LDL transport was 5.4+/-2.9%/h and 4.1+/-1.5%/h in patients with diabetes and control subjects, respectively (P<0.05); equivalent values for albumin were 6.5+/ 2.5%/h and 5.3+/-1.6%/h (P<0.05). This difference most likely was not caused by altered hepatic LDL receptor expression, glycosylation of LDL, small LDL size, nephropathy, statin use, or different plasma insulin levels in diabetic patients. CONCLUSIONS: Transvascular LDL transport may be increased in patients with type 2 diabetes. This suggests that lipoprotein flux into the arterial wall is increased in people with diabetes, possibly explaining the accelerated development of atherosclerosis. PMID- 12117734 TI - Nuclear magnetic resonance spectroscopy of lipoproteins and risk of coronary heart disease in the cardiovascular health study. AB - OBJECTIVES: Relationships between incident cardiovascular disease and lipoprotein subclass measurements by nuclear magnetic resonance spectroscopy were evaluated in the Cardiovascular Health Study (CHS) in a nested case-cohort analysis. METHODS AND RESULTS: The case group consisted of 434 participants with incident myocardial infarction (MI) and angina diagnosed after entry to the study (1990 to 1995) and the comparison group, 249 "healthy" participants with no prevalent clinical or subclinical disease. By univariate analysis, the median levels for healthy participants versus participants with incident MI and angina were 0 versus 7 mg% for small low density lipoprotein (LDL), 1501 versus 1680 nmol/L for the number of LDL particles, and 21.6 versus 21.3 for LDL size, and these values were significantly different between "healthy" participants and those with incident MI and angina for women but not men. The levels of less dense LDL, which is most of the total LDL cholesterol among women, was not related to incident MI and angina. For women, large high density lipoprotein cholesterol (HDLc), but not small HDLc, levels were significantly higher for healthy participants compared with levels for participants with MI and angina. For men and women, levels of total and very low density lipoprotein triglycerides were higher for the case group than for the healthy group. In multivariate models for women that included triglycerides and HDLc, the number of LDL particles (but not LDL size) remained significantly related to MI and angina. CONCLUSIONS: Small LDL, the size of LDL particles, and the greater number of LDL particles are related to incident coronary heart disease among older women. PMID- 12117735 TI - Characterization of LDL particle size among carriers of a defective or a null mutation in the lipoprotein lipase gene: the Quebec LIPD Study. AB - OBJECTIVE: The objective of the present study was to compare the impact of the null P207L and defective D9N mutations in the LPL gene on LDL particle size among heterozygous carriers. METHODS AND RESULTS: LDL particle size was measured on whole plasma by 2% to 16% non-denaturing polyacrylamide gradient gel electrophoresis in a cohort of 206 heterozygous carriers of either the P207L or the D9N mutation. The P207L carriers (N=88) presented with a more atherogenic lipoprotein-lipid profile compared with the D9N carriers (N=118). Accordingly, LDL particle size was smaller in the P207L carriers than in the D9N subjects (248.8+/- 1.0 vs 254.5+/-1.0 A, P< 0.001), and the difference remained significant after adjustment for plasma triglyceride (TG) levels. The difference in LDL diameter between the P207L and the D9N carriers was 3-fold greater in individuals with plasma TG levels >3.5 mmol/L than in subjects with TG < or =3.5 mmol/L. The factors that statistically contributed to LDL particle size variation in multivariate analyses were plasma TG levels (11.6%) and age (6.4%) in subjects with TG levels < or =3.5 mmol/L and HDL cholesterol levels (15.5%) and the LPL gene mutation (null versus defective, 7.0%) in patients with TG levels >3.5 mmol/L. CONCLUSIONS: These results suggest that the null P207L mutation in the LPL gene has a greater impact on LDL particle size than the defective D9N mutation and that this mutation-specific effect is amplified at greater plasma TG concentrations. PMID- 12117736 TI - Monocyte and neutrophil adhesion molecule expression during acute hyperglycemia and after antioxidant treatment in type 2 diabetes and control patients. AB - OBJECTIVE: We hypothesized that acute hyperglycemia (an independent cardiovascular risk factor) increases the expression of proatherogenic leukocyte adhesion molecule in type 2 diabetes and controls and that the expression of these adhesion molecules would be antioxidant sensitive. METHODS AND RESULTS: Twenty-three type 2 diabetes patients and 13 control patients underwent two oral glucose tolerance tests 14 days apart and took placebo or 800 IU daily of oral alpha tocopherol between tests. Monocyte and neutrophil expression of adhesion molecules Mac-1, LFA-1 and 3, ICAM-1, and VLA-4 were measured at 0, 120, and 240 minutes by using laser flow cytometry. Baseline adhesion molecule expression did not differ between groups, but there was a rapid, highly significant increase (P<0.0001) in the intensity of monocyte Mac-1 expression after a glucose load in both groups. Alpha-tocopherol supplementation reduced only Mac-1 expression in the diabetes group (P=0.03). CONCLUSIONS: Acute glycemic excursions of any degree cause highly significant, rapid increases in monocyte Mac-1 expression in type 2 diabetes patients and controls. Mac-1 mediates leukocyte vascular infiltration and is prothrombotic. These data suggest a mechanism for the link between glycemic excursions and increased vascular event rates. PMID- 12117737 TI - Simvastatin reduces expression of cytokines interleukin-6, interleukin-8, and monocyte chemoattractant protein-1 in circulating monocytes from hypercholesterolemic patients. AB - OBJECTIVE: A number of studies have shown that statins decrease morbidity and mortality in patients with cardiovascular diseases. The anti-inflammatory effects of statins have recently been implicated in the clinical benefit that can be obtained in the treatment of atherosclerosis. Little is known about the mechanisms by which statins counteract inflammation. METHODS AND RESULTS: In this study, we asked whether simvastatin can influence in vitro and in vivo production of the proinflammatory cytokines interleukin (IL)-6, IL-8, and monocyte chemoattractant protein-1. A total of 107 hypercholesterolemic patients were treated with simvastatin. As measured by ELISA, serum levels of cytokines significantly decreased after 6 weeks of treatment (P<0.05). Furthermore, simvastatin decreased the expression of IL-6, IL-8, and monocyte chemoattractant protein-1 mRNA in peripheral blood mononuclear cells. Similar results were obtained in vitro by using cultured human umbilical vein endothelial cells and peripheral blood mononuclear cells from healthy normolipemic donors. Exposure to simvastatin, atorvastatin, or cerivastatin caused downregulation of the expression of cytokine mRNA in a time- and dose-dependent manner. Furthermore, all statins tested were able to reduce the concentrations of cytokines in cellular and extracellular fractions of human umbilical vein endothelial cells (P<0.05). CONCLUSIONS: Our data show that simvastatin is anti-inflammatory through the downregulation of cytokines in the endothelium and leukocytes. These effects may explain some of the clinical benefits of these drugs in the treatment of atherosclerosis. PMID- 12117738 TI - Extracellular matrix metalloproteinase inducer (EMMPRIN) is induced upon monocyte differentiation and is expressed in human atheroma. AB - OBJECTIVE: Because extracellular matrix metalloproteinase inducer (EMMPRIN), a tumor cell-derived protein, induces matrix metalloproteinases (MMPs) in fibroblasts and because MMPs are important in atheroma formation, we investigated if EMMPRIN was expressed in granulocyte/macrophage-colony stimulating factor (GM CSF)-differentiated human peripheral blood monocytes (HPBM) and macrophage foam cells. In addition, EMMPRIN was studied for its expression in human atheroma. METHODS AND RESULTS: After 10 days of GM-CSF-induced monocyte differentiation, EMMPRIN mRNA increased 5- to 8-fold relative to undifferentiated monocytes. GM CSF treatment of HPBM revealed that both EMMPRIN mRNA and protein were upregulated by day 2 over undifferentiated monocytes. GM-CSF-differentiated HPBM showed characteristic macrophage phenotype by showing increases in pancake-like morphology and increases in biochemical markers such as apolipoprotein E, MMP-9, and cholesterol ester (CE). While acetylated LDL treatment of the 10-day GM-CSF differentiated HPBM increased CE mass 13- to 321-fold, EMMPRIN expression was unchanged relative to nonlipid-loaded macrophages. In human coronary atherosclerotic samples, EMMPRIN was observed in CD68(+) macrophage-rich areas as well as areas of MMP-9 expressions. CONCLUSIONS: Based on these data, we conclude that monocyte differentiation induces EMMPRIN expression, CE enrichment of foam cells has no further effect on EMMPRIN expression, and EMMPRIN is present in human atheroma. Therefore, EMMPRIN may play a role in atherosclerosis development. PMID- 12117739 TI - Angiotensin II type 1 receptor antagonism improves hypercholesterolemia associated endothelial dysfunction. AB - OBJECTIVE: Hypercholesterolemia-induced angiotensin II type 1 (AT1) receptor overexpression is thought to be a key event in the development of endothelial dysfunction. METHODS AND RESULTS: The effect of a 6-week treatment with the AT1 receptor antagonist candesartan (16 mg/d) on endothelial function and serum inflammation markers was compared with the effect of treatment with placebo or the calcium channel antagonist felodipine (5 mg/d) in 47 hypercholesterolemic patients (low density lipoprotein cholesterol >160 mg/dL). Endothelial function was assessed by measurement of forearm blood flow (FBF) by venous occlusion plethysmography. FBF during reactive hyperemia was significantly improved by candesartan, whereas felodipine and placebo exerted no effect. Nitroglycerin induced vasorelaxation and basal FBF were not altered significantly. Blood pressure and cholesterol levels were not affected significantly by any drug. Serum concentrations of 8-isoprostane, monocyte chemoattractant protein-1, and soluble intercellular adhesion molecule-1 were significantly reduced by candesartan treatment but not by placebo or felodipine (ELISA assays). Levels of high-sensitivity C-reactive protein and tumor necrosis factor-alpha were not altered significantly by any treatment. CONCLUSIONS: These data suggest that AT1 receptor antagonism improves endothelial function during hypercholesterolemia and that this applies not only to endothelium-dependent vasodilatation but also to oxidative stress and events involved in monocyte attraction and adhesion. AT1 receptor blockade may potentially represent a novel approach for the prevention of vascular dysfunction associated with hypercholesterolemia that is independent of lipid-lowering and blood pressure-lowering interventions. PMID- 12117740 TI - Human secretory phospholipase A2 mediates decreased plasma levels of HDL cholesterol and apoA-I in response to inflammation in human apoA-I transgenic mice. AB - OBJECTIVE: Plasma levels of high density lipoprotein (HDL) cholesterol and apolipoprotein (apo)A-I are decreased in inflammatory states. Secretory phospholipase A2 (sPLA2), an acute-phase protein, may play a key role in the pathophysiology of this phenomenon. METHODS AND RESULTS: To investigate the effects of sPLA2 on human-like HDL particles in vivo, we generated transgenic mice overexpressing human apoA-I and human sPLA2 (apoA-I/sPLA2 mice). Compared with apoA-I mice, apoA-I/sPLA2 mice had significantly lower plasma levels of phospholipids, HDL cholesterol, and apoA-I (each P<0.01). HDL from apoA-I/sPLA2 mice was significantly depleted in phospholipids and cholesteryl esters (each P<0.001) but was enriched in protein and triglycerides (each P<0.001). As assessed by gel filtration and nondenaturing gel electrophoresis, sPLA2 overexpression in apoA-I mice resulted in a dramatic shift of the HDL particle size toward smaller particles. Furthermore, virtually all plasma sPLA2 in apoA I/sPLA2 mice was found in association with the HDL fraction. The acute-phase response was induced in apoA-I/sPLA2 double-transgenic and apoA-I single transgenic mice by intraperitoneal lipopolysaccharide (LPS) injection. Plasma sPLA2 was significantly increased after LPS injection in apoA-I/sPLA2 mice. Twelve hours after LPS administration, plasma total cholesterol, HDL cholesterol, apoA-I, and phospholipids were unchanged in apoA-I transgenic control mice but had decreased significantly in the apoA-I/sPLA2 mice (-57%, -62%, and -54%, -61%, respectively; each P<0.001). Both groups of mice had increased plasma levels of serum amyloid A (SAA) in response to LPS. To test the hypothesis that SAA may be an in vivo activator of sPLA2, we specifically overexpressed SAA in apoA-I/sPLA2 mice by means of liver-directed gene transfer. Despite high plasma levels of SAA, plasma lipid and lipoprotein profiles were not different than those in control mice. CONCLUSIONS: These results in a mouse model of human-like HDL indicate that sPLA2 expression significantly influences HDL particle size and composition and demonstrate that an induction of sPLA2 is required for the decrease in plasma HDL cholesterol in response to inflammatory stimuli in mice and that this effect is independent of SAA. PMID- 12117741 TI - Chronic exercise improves endothelial calcium signaling and vasodilatation in hypercholesterolemic rabbit femoral artery. AB - OBJECTIVE: This study was to investigate the effects of chronic exercise on vasodilatation and endothelial intracellular calcium (EC [Ca2+]i) signaling in atherosclerotic animals. METHODS AND RESULTS: For 8 weeks, male New Zealand White rabbits were fed rabbit chow with or without the addition of 2% cholesterol. They were further divided into control and exercise groups. Animals in the exercise groups ran on a leveled treadmill at 0.88 km/h for 10 to 60 minutes gradually for 5 days per week for a total of 8 weeks. At the end of experiments, femoral arteries were dissected, loaded with fura 2-AM, and mounted in a tissue flow chamber. PE-precontracted vessel specimens were exposed to acetylcholine (ACh). The EC [Ca2+]i elevation and vasorelaxation were determined simultaneously under an epifluorescence microscope equipped with a ratio-imaging capability. Our results showed the following: (1) high cholesterol diet feeding caused lipid deposition on vascular surface, reduced the ACh-evoked EC [Ca2+]i elevation, and impaired endothelium-dependent and endothelium-independent vascular responses, but chronic exercise had the opposite effects; (2) ACh-induced vasorelaxation was associated with EC [Ca2+]i elevation in all groups; and (3) vasorelaxation at high levels of EC [Ca2+]i elevation decreased in hypercholesterolemia. CONCLUSIONS: Our data suggest that hypercholesterolemia induces vascular structural changes and impairs EC [Ca2+]i signaling and vasodilatation, whereas chronic exercise partially reverses these adverse effects. PMID- 12117742 TI - Curcuma longa extract supplementation reduces oxidative stress and attenuates aortic fatty streak development in rabbits. AB - OBJECTIVE: This study evaluates the effect of a Curcuma longa extract on the development of experimental atherosclerosis (fatty streak) in rabbits and its interaction with other plasmatic antioxidants. METHODS AND RESULTS: Two experimental groups of male New Zealand White rabbits, a control group and a curcuma-extract (CU) group, were fed an atherogenic diet. Additionally, the CU group received an oral curcuma hydroalcoholic extract. Six animals from each experimental group were killed after 10, 20, and 30 days. Compared with the CU group, the control group showed significantly higher plasma lipid peroxide at all experimental times (10, 20, and 30 days) and significantly lower alpha-tocopherol and coenzyme Q levels at 20 and 30 days. Histological results for the fatty streak lesions revealed damage in the thoracic and abdominal aorta that was significantly lower in the CU group than in the control group at 30 days. CONCLUSIONS: Supplementation with Curcuma longa reduces oxidative stress and attenuates the development of fatty streaks in rabbits fed a high cholesterol diet. PMID- 12117744 TI - Physiological variations of isoprostanes: a step forward? PMID- 12117743 TI - Lp(a) particles mold fibrin-binding properties of apo(a) in size-dependent manner: a study with different-length recombinant apo(a), native Lp(a), and monoclonal antibody. AB - OBJECTIVE: Small-sized apolipoprotein(a) [apo(a)] isoforms with high antifibrinolytic activity are frequently found in cardiovascular diseases, suggesting a role for apo(a) size in atherothrombosis. To test this hypothesis, we sought to characterize the lysine (fibrin)-binding function of isolated apo(a) of variable sizes. METHODS AND RESULTS: Recombinant apo(a) [r-apo(a)] preparations consisting of 10 to 34 kringles and a monoclonal antibody that neutralizes the lysine-binding function were produced and used in parallel with lipoprotein(a) [Lp(a)] particles isolated from plasma in fibrin-binding studies. All r-apo(a) preparations displayed similar affinity and specificity for lysine residues on fibrin regardless of size (K(d) 3.6+/-0.3 nmol/L) and inhibited the binding of plasminogen with a similar intensity (IC50 16.8+/-5.4 nmol/L). In contrast, native Lp(a) particles displayed fibrin affinities that were in inverse relationship with the apo(a) kringle number. Thus, a 15-kringle apo(a) separated from Lp(a) and a 34-kringle r-apo(a) displayed an affinity for fibrin that was higher than that in the corresponding particles (K(d) 2.5 versus 10.5 nmol/L and K(d) 3.8 versus 541 nmol/L, respectively). However, fibrin-binding specificity of the r-apo(a) preparations and the Lp(a) particles was efficiently neutralized (IC50 0.07 and 4 nmol/L) by a monoclonal antibody directed against the lysine binding function of kringle IV-10. CONCLUSIONS: Our data indicate that fibrin binding is an intrinsic property of apo(a) modulated by the composite structure of the Lp(a) particle. PMID- 12117745 TI - Exercise-induced suppression of postprandial lipemia: a possible mechanism of endothelial protection? PMID- 12117746 TI - MMP inhibition and lumen loss after balloon angioplasty or stenting. PMID- 12117748 TI - Herman P. Schwan: a scientist and pioneer in biomedical engineering. PMID- 12117747 TI - Effect of dexrazoxane on homocysteine-induced endothelial dysfunction in normal subjects. AB - OBJECTIVE: Dexrazoxane is an antioxidant prodrug that on hydrolysis is converted into an intracellular iron chelator. We hypothesized that the antioxidant effects of dexrazoxane would prevent homocysteine-induced endothelial dysfunction in the brachial artery of normal human subjects. METHODS AND RESULTS: Ten healthy volunteers completed a randomized, double-blind, crossover study. Plasma homocysteine levels and brachial artery endothelium-dependent (flow-mediated dilation [FMD]) and endothelium-independent (sublingual nitroglycerin) responses were measured before and 4 hours after ingestion of L-methionine (100 mg/kg), preceded by intravenous administration of dexrazoxane (500 mg/m2) or placebo over 30 minutes. After placebo, oral methionine increased plasma homocysteine (from 5.1+/-0.4 micromol/L at baseline to 14.2+/-1.3 micromol/L at 4 hours, P<0.001) and decreased FMD (from 3.8+/-0.7% at baseline to 1.2+/-0.5% at 4 hours, P=0.02). Dexrazoxane did not change homocysteine concentrations after methionine administration (14.9+/-1.1 micromol/L at 4 hours, P=0.29 versus placebo) but did completely abrogate the homocysteine-induced reduction in FMD (from 3.5+/-0.5% at baseline to 5.9+/-1.1% at 4 hours, P<0.01 versus placebo). Endothelium independent responses to sublingual nitroglycerin did not differ after the administration of placebo and dexrazoxane. CONCLUSIONS: Administration of the novel antioxidant agent dexrazoxane prevents homocysteine-induced impairment of vascular endothelial function in the brachial artery of healthy subjects. PMID- 12117749 TI - Roles for learning sciences and learning technologies in biomedical engineering education: a review of recent advances. AB - Education in biomedical engineering offers a number of challenges to all constituents of the educational process-faculty, students, and employers of graduates. Although biomedical engineering educational systems have been under development for 40 years, interest in and the pace of development of these programs has accelerated in recent years. New advances in the learning sciences have provided a framework for the reexamination of instructional paradigms in biomedical engineering. This work shows that learning environments should be learner centered, knowledge centered, assessment centered, and community centered. In addition, learning technologies offer the potential to achieve this environment with efficiency. Biomedical engineering educators are in a position to design and implement new learning systems that can take advantage of advances in learning science, learning technology, and reform in engineering education. PMID- 12117750 TI - Spine ergonomics. AB - Occupational low back pain (LBP) is an immense burden for both industry and medicine. Ergonomic and personal risk factors result in LBP, but psychosocial factors can influence LBP disability. Epidemiologic studies clearly indicate the role of mechanical loads on the etiology of occupational LBP. Occupational exposures such as lifting, particularly in awkward postures; heavy lifting; or repetitive lifting are related to LBP. Fixed postures and prolonged seating are also risk factors. LBP is found in both sedentary occupations and in drivers as well as those involved in manual materials handling. Any prolonged posture will lead to static loading of the soft tissues and cause discomfort. Standing and sitting have specific advantages and disadvantages for mobility, exertion of force, energy consumption, circulatory demands, coordination, and motion control. The seated posture leads to inactivity causing an accumulation of metabolites, accelerating disk degeneration and leading to disk herniation. Driver's postures can also lead to musculoskeletal problems. Workers in a driving environment are often subjected to postural stress leading to back, neck, and upper extremity pain. This exacerbates the problems due to the vibration. Prevention is by far the treatment of choice. Improved muscle function can be preventative. Poor coordination and motor control systems are as important as endurance and strength. Fixed postures should be avoided. Seats offering good lumbar support should be used in the office. A suspension seat should be used in vehicles whenever possible. Heavy and awkward lifting should be avoided and lifting aids should be made available. Workers should report LBP as early as possible and seek medical advice if they think occupational exposure is harming them. The combined effects of the medical community, labor, and management are required to cause some impact on this problem. PMID- 12117751 TI - Three-dimensional confocal microscopy of the living human eye. AB - Three-dimensional confocal microscopy of the living eye is a major development in instrumentation for biomicroscopy of the eye. This noninvasive optical technology has its roots in the application of optics to reflected light imaging of the eye. These instrument developments began with Leeuwenhoek's use of his single lens microscope to investigate the structure of the eye. There followed a series of connected instruments: the ophthalmoscope, the slit lamp, the specular microscope, and the clinical confocal microscope. In vivo confocal microscopy produces high contrast, reflected light images or optical sections through the depth of living ocular tissue. Stacks of registered optical sections can be transformed by computer visualization techniques into three-dimensional volume images of ocular tissues: cornea, ocular lens, retina, and optic nerve. The clinical confocal microscope has resulted in new diagnostic techniques and new cellular descriptions of ocular disorders and pathology. PMID- 12117752 TI - Bioengineering of therapeutic aerosols. AB - The new field of therapeutic aerosol bioengineering (TAB), driven primarily by the medical need for inhaled insulin, is now expanding to address medical needs ranging from respiratory to systemic diseases, including asthma, growth deficiency, and pain. Bioengineering of therapeutic aerosols involves a level of aerosol particle design absent in traditional therapeutic aerosols, which are created by conventionally spraying a liquid solution or suspension of drug or milling and mixing a dry drug form into respirable particles. Bioengineered particles may be created in liquid form from devices specially designed to create an unusually fine size distribution, possibly with special purity properties, or solid particles that possess a mixture of drug and excipient, with designed shape, size, porosity, and drug release characteristics. Such aerosols have enabled several high-visibility clinical programs of inhaled insulin, as well as earlier-stage programs involving inhaled morphine, growth hormone, beta interferon, alpha-1-antitrypsin, and several asthma drugs. The design of these aerosols, limited by partial knowledge of the lungs' physiological environment, and driven largely at this stage by market forces, relies on a mixture of new and old science, pharmaceutical science intuition, and a degree of biological-impact empiricism that speaks to the importance of an increased level of academic involvement. PMID- 12117753 TI - Denaturation of collagen via heating: an irreversible rate process. AB - Heating therapies are increasingly used in cardiology, dermatology, gynecology, neurosurgery, oncology, ophthalmology, orthopedics, and urology, among other medical specialties. This widespread use of heating is driven primarily by the availability of new technology, not by a detailed understanding of the biothermomechanics. Without basic quantification of the underlying physical and chemical processes in terms of parameters that can be controlled clinically, identification of preferred interventions will continue to be based primarily on trial and error, thus necessitating large clinical studies and years of accumulative experience. Perusal of the literature reveals that much has been learned over the past century about the response of cells, proteins, and tissues to supra-physiologic temperatures; yet, the associated findings are reported in diverse journals and the underlying basic processes remain unidentified. In this review, we seek to contrast various findings on the kinetics of the thermal denaturation of collagen and to encourage investigators to consider the many open problems in part via a synthesis of results from the diverse literatures. PMID- 12117754 TI - DNA microarray technology: devices, systems, and applications. AB - In this review, recent advances in DNA microarray technology and their applications are examined. The many varieties of DNA microarray or DNA chip devices and systems are described along with their methods for fabrication and their use. This includes both high-density microarrays for high-throughput screening applications and lower-density microarrays for various diagnostic applications. The methods for microarray fabrication that are reviewed include various inkjet and microjet deposition or spotting technologies and processes, in situ or on-chip photolithographic oligonucleotide synthesis processes, and electronic DNA probe addressing processes. The DNA microarray hybridization applications reviewed include the important areas of gene expression analysis and genotyping for point mutations, single nucleotide polymorphisms (SNPs), and short tandem repeats (STRs). In addition to the many molecular biological and genomic research uses, this review covers applications of microarray devices and systems for pharmacogenomic research and drug discovery, infectious and genetic disease and cancer diagnostics, and forensic and genetic identification purposes. Additionally, microarray technology being developed and applied to new areas of proteomic and cellular analysis are reviewed. PMID- 12117755 TI - Peptide aggregation in neurodegenerative disease. AB - In the not-so-distant past, insoluble aggregated protein was considered as uninteresting and bothersome as yesterday's trash. More recently, protein aggregates have enjoyed considerable scientific interest, as it has become clear that these aggregates play key roles in many diseases. In this review, we focus attention on three polypeptides: beta-amyloid, prion, and huntingtin, which are linked to three feared neurodegenerative diseases: Alzheimer's, "mad cow," and Huntington's disease, respectively. These proteins lack any significant primary sequence homology, yet their aggregates possess very similar features, specifically, high beta-sheet content, fibrillar morphology, relative insolubility, and protease resistance. Because the aggregates are noncrystalline, secrets of their structure at nanometer resolution are only slowly yielding to X ray diffraction, solid-state NMR, and other techniques. Besides structure, the aggregates may possess similar pathways of assembly. Two alternative assembly pathways have been proposed: the nucleation-elongation and the template-assisted mode. These two modes may be complementary, not mutually exclusive. Strategies for interfering with aggregation, which may provide novel therapeutic approaches, are under development. The structural similarities between protein aggregates of dissimilar origin suggest that therapeutic strategies successful against one disease may have broad utility in others. PMID- 12117756 TI - Mechano-electrochemical properties of articular cartilage: their inhomogeneities and anisotropies. AB - In this chapter, the recent advances in cartilage biomechanics and electromechanics are reviewed and summarized. Our emphasis is on the new experimental techniques in cartilage mechanical testing, new experimental and theoretical findings in cartilage biomechanics and electromechanics, and emerging theories and computational modeling of articular cartilage. The charged nature and depth-dependent inhomogeneity in mechano-electrochemical properties of articular cartilage are examined, and their importance in the normal and/or pathological structure-function relationships with cartilage is discussed, along with their pathophysiological implications. Developments in theoretical and computational models of articular cartilage are summarized, and their application in cartilage biomechanics and biology is reviewed. Future directions in cartilage biomechanics and mechano-biology research are proposed. PMID- 12117757 TI - Electromagnetic fields: human safety issues. AB - Most of the recently revised safety standards worldwide are set in terms of internal rates of electromagnetic energy deposition (specific absorption rates or SAR) at radio frequencies (RF) and microwave frequencies, and of induced electric fields or current densities at lower frequencies up to 10 MHz. Numerical methods have been developed that use millimeter resolution anatomically based models of the human body to determine SAR or the induced electric fields and current densities for real-life EM exposure conditions. A popular method for use at RF and microwave frequencies is the finite-difference time-domain method. This method is described and illustrated for SAR distributions due to cellular telephones for head models based on human anatomy. A method often used for calculations of induced electric fields and current densities at low frequencies is the impedance method. Use of this method is illustrated by an example of an electronic article surveillance (EAS) system for anatomic models of an adult and 10- and 5-year-old children. Experimental phantoms using a fluid to simulate the dielectric properties of the brain may be used for determination of peak 1- or 10 g SAR needed for compliance with the various safety standards. PMID- 12117758 TI - Advances in in vivo bioluminescence imaging of gene expression. AB - To advance our understanding of biological processes as they occur in living animals, imaging strategies have been developed and refined that reveal cellular and molecular features of biology and disease in real time. One rapid and accessible technology for in vivo analysis employs internal biological sources of light emitted from luminescent enzymes, luciferases, to label genes and cells. Combining this reporter system with the new generation of charge coupled device (CCD) cameras that detect the light transmitted through the animal's tissues has opened the door to sensitive in vivo measurements of mammalian gene expression in living animals. Here, we review the development and application of this imaging strategy, in vivo bioluminescence imaging (BLI), together with in vivo fluorescence imaging methods, which has enabled the real-time study of immune cell trafficking, of various genetic regulatory elements in transgenic mice, and of in vivo gene transfer. BLI has been combined with fluorescence methods that together offer access to in vivo measurements that were not previously available. Such studies will greatly facilitate the functional analysis of a wide range of genes for their roles in health and disease. PMID- 12117759 TI - Physics and applications of microfluidics in biology. AB - Fluid flow at the microscale exhibits unique phenomena that can be leveraged to fabricate devices and components capable of performing functions useful for biological studies. The physics of importance to microfluidics are reviewed. Common methods of fabricating microfluidic devices and systems are described. Components, including valves, mixers, and pumps, capable of controlling fluid flow by utilizing the physics of the microscale are presented. Techniques for sensing flow characteristics are described and examples of devices and systems that perform bioanalysis are presented. The focus of this review is microscale phenomena and the use of the physics of the scale to create devices and systems that provide functionality useful to the life sciences. PMID- 12117760 TI - Telerehabilitation research: emerging opportunities. AB - The field of clinical rehabilitation is rooted in the premise that carefully planned and delivered therapeutic intervention enhances patient outcomes. Underlying this statement is a deeper scientific reality: The field exists because biosystems (e.g., tissues, cells, organs, persons) are inherently adaptive and can dynamically change as a function of a sequence of inputs (e.g., exercise, pharmaceuticals). The tools of telerehabilitation help minimize the barrier of distance, both of patients to rehabilitative services and of researchers to subject populations. This enhanced access opens up new possibilities for discovering and implementing optimized intervention strategies across the continuum of care. Telecommunications technologies are reviewed from the perspective of systems models of the telerehabilitation process, with a focus on human-technology interface design and a special emphasis on emerging home and mobile technologies. Approaches for providing clinical rehabilitation services through telerehabilitation are addressed, including innovative consumer-centered approaches. Finally, telerehabilitation is proposed as a tool for reinvigorating the rehabilitative bioengineering research enterprise. PMID- 12117761 TI - Biomechanical dynamics of the heart with MRI. AB - Magnetic resonance imaging (MRI) provides a noninvasive way to evaluate the biomechanical dynamics of the heart. MRI can provide spatially registered tomographic images of the heart in different phases of the cardiac cycle, which can be used to assess global cardiac function and regional endocardial surface motion. In addition, MRI can provide detailed information on the patterns of motion within the heart wall, permitting calculation of the evolution of regional strain and related motion variables within the wall. These show consistent patterns of spatial and temporal variation in normal subjects, which are affected by alterations of function due to disease. Although still an evolving technique, MRI shows promise as a new method for research and clinical evaluation of cardiac dynamics. PMID- 12117762 TI - Advances in proteomic technologies. AB - Proteomics is a rapidly emerging set of key technologies that are being used to identify proteins and map their interactions in a cellular context. With the sequencing of the human genome, the scope of proteomics has shifted from protein identification and characterization to include protein structure, function and protein-protein interactions. Technologies used in proteomic research include two dimensional gel electrophoresis, mass spectrometry, yeast two-hybrids screens, and computational prediction programs. While some of these technologies have been in use for a long time, they are currently being applied to study physiology and cellular processes in high-throughput formats. It is the high-throughput approach that defines and characterizes modern proteomics. In this review, we discuss the current status of these experimental and computational technologies relevant to the three major aspects of proteomics-characterization of proteomes, identification of proteins, and determination of protein function. We also briefly discuss the development of new proteomic technologies that are based on recent advances in analytical and biochemical techniques, engineering, microfabrication, and computational prowess. The integration of these advances with established technologies is invaluable for the drive toward a comprehensive understanding of protein structure and function in the cellular milieu. PMID- 12117763 TI - On the metrics and euler-lagrange equations of computational anatomy. AB - This paper reviews literature, current concepts and approaches in computational anatomy (CA). The model of CA is a Grenander deformable template, an orbit generated from a template under groups of diffeomorphisms. The metric space of all anatomical images is constructed from the geodesic connecting one anatomical structure to another in the orbit. The variational problems specifying these metrics are reviewed along with their associated Euler-Lagrange equations. The Euler equations of motion derived by Arnold for the geodesics in the group of divergence-free volume-preserving diffeomorphisms of incompressible fluids are generalized for the larger group of diffeomorphisms used in CA with nonconstant Jacobians. Metrics that accommodate photometric variation are described extending the anatomical model to incorporate the construction of neoplasm. Metrics on landmarked shapes are reviewed as well as Joshi's diffeomorphism metrics, Bookstein's thin-plate spline approximate-metrics, and Kendall's affine invariant metrics. We conclude by showing recent experimental results from the Toga & Thompson group in growth, the Van Essen group in macaque and human cortex mapping, and the Csernansky group in hippocampus mapping for neuropsychiatric studies in aging and schizophrenia. PMID- 12117764 TI - Selective electrical interfaces with the nervous system. AB - To achieve selective electrical interfacing to the neural system it is necessary to approach neuronal elements on a scale of micrometers. This necessitates microtechnology fabrication and introduces the interdisciplinary field of neurotechnology, lying at the juncture of neuroscience with microtechnology. The neuroelectronic interface occurs where the membrane of a cell soma or axon meets a metal microelectrode surface. The seal between these may be narrow or may be leaky. In the latter case the surrounding volume conductor becomes part of the interface. Electrode design for successful interfacing, either for stimulation or recording, requires good understanding of membrane phenomena, natural and evoked action potential generation, volume conduction, and electrode behavior. Penetrating multimicroelectrodes have been produced as one-, two-, and three dimensional arrays, mainly in silicon, glass, and metal microtechnology. Cuff electrodes circumvent a nerve; their selectivity aims at fascicles more than at nerve fibers. Other types of electrodes are regenerating sieves and cone-ingrowth electrodes. The latter may play a role in brain-computer interfaces. Planar substrate-embedded electrode arrays with cultured neural cells on top are used to study the activity and plasticity of developing neural networks. They also serve as substrates for future so-called cultured probes. PMID- 12117765 TI - The initiation of breast and prostate cancer. AB - The agents responsible for the initiation of human mammary and prostatic cancers remain unidentified. Population migration studies on breast and prostate cancer risk have revealed that incidence rates in migrants from low-risk to high-risk "Westernized" countries rise over time to match those of the host populations. The parallels suggest that the two diseases may share a common aetiology, with changes in diet, rather than in environment, being responsible for the migration related increases in cancer incidence. Genotoxins, such as polycyclic aromatic hydrocarbons and heterocyclic aromatic amines, are formed when foodstuffs are cooked at elevated temperatures and can be extracted with solvents: other genotoxins may only be released from cooked proteins when digestion occurs in the gastrointestinal tract. Human mammary and prostatic epithelial cells are known to be capable of metabolically activating members of different classes of chemical carcinogens to DNA-reactive species and, in rodents, five out of six mammary carcinogens can also induce prostatic neoplasms. Genotoxins have been detected in some 40% of breast lipid and milk samples donated by UK-resident women but the agents, currently thought to be of dietary origin, have not been characterized or identified as yet. Reduction mammoplasty and lactation both reduce breast cancer risk and the reduction is proportional either to the amount of tissue removed or to the total duration of lactation. As DNA damage has been detected in otherwise untreated mammary epithelial cells isolated both from breast tissue and from breast milk, we have proposed that reduction mammoplasty and lactation reduce risk through a common mechanism, i.e. the loss of pre-malignant cells. Further research, perhaps aimed particularly at the characterization of all the carcinogens formed when different dietary components are cooked in different ways, should succeed in identifying the agents that initiate breast and prostate cancer. PMID- 12117766 TI - The impact of chromatin in human cancer: linking DNA methylation to gene silencing. AB - For decades, chromatin was considered to be an inert structure whose only role was the compacting and confining of DNA inside the eukaryotic nucleus. However, tremendous progress in this field over the last 10 years has dramatically elevated chromatin to a key position in the control of gene activity. Its role in mediating the transformation of a normal cell into a malignant state is particularly interesting. On one side of this story there is the discovery that aberrant methylation patterns in an increasing number of tumour suppressor and DNA repair genes determine carcinogenetic transformation; while on the other side, there is the existence of a series of methyl-DNA binding activities that recruit co-repressor complexes and modify the structure of the chromatin to produce a transcriptionally silenced state. Although this field has seen rapid progress in recent years, detailed mechanisms by which this machinery modifies chromatin structure to its appropriate state and the specific targeting of repressor complexes have yet to be resolved. In this review we present the models of how repressor complexes may modify chromatin structure and mediate silencing of tumour suppressor and DNA repair genes. PMID- 12117767 TI - DNA damage in lung epithelial cells isolated from rats exposed to quartz: role of surface reactivity and neutrophilic inflammation. AB - Respirable quartz has been classified as a human lung carcinogen (IARC, 1997). However, the mechanisms involved in quartz-induced carcinogenesis remain unclear. The aim of the present study was to investigate acute DNA damage in epithelial lung cells from rats exposed to quartz. Since surface reactivity is considered to play a crucial role in the toxicity of quartz, the effect of surface modifying agents polyvinylpyridine-N-oxide (PVNO) and aluminium lactate (AL) was evaluated. Therefore, rats were instilled with quartz (DQ12, 2 mg/rat) or quartz treated with PVNO or AL. After 3 days animals were killed and brochoalveolar lavage (BAL) was performed to evaluate inflammatory cell influx. BAL-fluid levels of lactate dehydrogenase (LDH), alkaline phosphatase (AP) and total protein were used as lung damage markers. Neutrophil activation was assessed by myeloperoxidase (MPO) measurement, and total antioxidant capacity of the BAL-fluid was determined using the TEAC (trolox equivalent antioxidant capacity) assay. Lung epithelial cells were isolated and DNA strand breakage was determined by single cell gel electrophoresis (comet assay). DNA damage was significantly increased in epithelial cells from rats instilled with DQ12, whereas no enhanced DNA strand breakage was observed when quartz was treated with PVNO or AL. Total protein, LDH and TEAC were increased in rats treated with native quartz, and this was inhibited by both coatings. A significant correlation between neutrophil numbers and MPO levels was observed, indicating neutrophil activation. Inhibition of DNA damage by both coatings was paralleled by a reduction of neutrophil influx as well as MPO activity. In this study we provide evidence that modification of the particle surface prevents DNA strand breakage in epithelial lung cells from quartz-exposed rats. Furthermore, the present data show the feasibility of our in vivo model to evaluate the role of inflammation, antioxidant status, and cytotoxicity in particle-induced DNA damage. PMID- 12117768 TI - Attenuation of the formation of DNA-repair foci containing RAD51 in Fanconi anaemia. AB - The role of the Fanconi anaemia genes in DNA repair was examined by a quantitative analysis of nuclear DNA repair foci in FA primary fibroblasts after ionising irradiation using antibodies directed against RAD51, MRE11 and BRCA1 for visualisation. IR induced foci detected with anti-RAD51, but not those detected with anti-MRE11, are reduced in fibroblasts of all eight FA complementation groups in comparison to control cells. Correction of FA-A, FA-C and FA-G cells by retroviral cDNA transfer specifically corrected the RAD51-foci response but did not affect formation of foci containing BRCA1 or MRE11. Since all FA cells, except FA-D1, lack the monoubiquitinated FANCD2-L protein, this isoform is likely to be involved in the formation of nuclear foci containing RAD51 in diploid FA cells. FA-D1 cells show the same attenuation in RAD51 foci formation, suggesting that the unknown FANCD1 protein is similarly involved in RAD51 foci formation, either independently or as a subsequent step in the FANCD2 pathway. These findings indicate that Fanconi anaemia cells have an impairment in the RAD51 dependent homologous recombination pathway for DNA repair, explaining their chromosomal instability and extreme sensitivity to DNA cross-linking agents. PMID- 12117769 TI - Alterations of the p16(INK4) locus in human malignant mesothelial tumors. AB - The INK4 locus has two promoters and encodes two unique proteins that share exons in different reading frames, p16(INK4a) and p14(ARF). The p16(INK4a) protein, by inhibiting cyclin-dependent kinase, down regulates Rb-E2F and leads to cell cycle arrest in the G1 phase. The p14(ARF) protein interacts with the MDM2 protein, neutralizing MDM2-mediated degradation of p53. Since p53/Rb genes are not altered in malignant mesothelioma, additional components of these pathways, such as p16(INK4a) and p14(ARF), are candidates for inactivation. In this study, we have examined p16(INK4a) and p14(ARF) alterations (gene deletion, mutation and promoter methylation) in 45 primary malignant mesothelioma specimens. Fourteen patients (31%) had altered p16; four tumors had a methylated promoter region (8.8%), 10 tumors showed p16 to be deleted (22.2%), and one tumor had a point mutation (2%). We did not find any instances of methylation in the p14(ARF) 5' CpG island. Patients whose tumors had p16 deletion were significantly younger than those with methylation, and, in the patients whose lungs were studied for the prevalence of asbestos fibers, those with any p16 alteration had lower fiber counts than those with no p16 alteration. Hence, p16 gene alteration is relatively common in malignant mesothelioma, while p14(ARF) is rarely, if ever, methylated. Our data suggest that deletion of p16 occurs in a relatively susceptible subset of the population. PMID- 12117770 TI - The colonic response to genotoxic carcinogens in the rat: regulation by dietary fibre. AB - The apoptotic response to DNA damage appears to be an innate biological mechanism for protection against tumourigenesis. It is possible that agents that protect against colorectal cancer act by enhancing the apoptotic deletion of cells suffering DNA damage, with consequent removal of those with tumourigenic mutations. We examined the acute apoptotic response to genotoxic carcinogens ("AARGC") in colonic epithelium and the possibility that dietary fibres of different fermentability might regulate AARGC. To fully define the time-course and nature of AARGC in response to the carcinogen azoxymethane (AOM), a single injection of AOM (10 mg/kg) was given to rats and apoptosis monitored in the colon by light microscopy and terminal deoxynucleotidyl transferase-mediated dUTP biotin nick end labelling staining over a 72 h period. Having defined the site and time of maximum response, two groups of eight rats were fed diets containing 10% wheat bran fibre (WB; fermentable) or 10% methylcellulose (MC; poorly fermentable) for 4 weeks. Colonic AARGC was compared by light microscopy; lumenal short chain fatty acids (SCFAs) and pH were measured as indicators of the fermentative environment. AOM-induced AARGC was maximal at 8 h and greater in distal compared with proximal colon. Apoptotic cells were situated predominantly in the lower half of the crypt, with the median at position 9 indicating involvement of daughter as well as stem cells. There was no "second wave" of apoptosis within 72 h as follows irradiation in small intestine. Distal colonic AARGC in rats fed WB was twice that in rats fed MC (P < 0.01). Compared with MC, WB significantly lowered lumenal pH and increased all SCFAs including butyrate, while proliferation did not differ between the fibres. Certainly, dietary fibres can regulate AARGC and further studies are warranted to determine if this biological effect is the way in which dietary factors regulate tumourigenesis. Lumenal generation of butyrate may enhance AARGC as butyrate is proapoptotic in vitro. PMID- 12117772 TI - Association of chromosome 19q13.2-3 haplotypes with basal cell carcinoma: tentative delineation of an involved region using data for single nucleotide polymorphisms in two cohorts. AB - We have previously used single nucleotide polymorphisms to detect an association of basal cell carcinoma (BCC) in Caucasian Americans and Danes with the genome region 19q13.2-3, which contains several genes involved in the nucleotide excision repair of DNA. In this exploratory paper we have extended the data and used them in a chromosomal scan. The results indicate the presence of a gene variation modulating the risk of developing BSS in a submegabase region including and surrounding the gene RAI. Specifically, persons that are homozygous for the haplotype RAI intron 1(A) RAI exon 6(A) appear at increased risk for BCC. In addition, we have looked for possible synergisms between all pairs of markers. We find that a marker in GLTSCR1, presumably separated from RAI by several million bases, supplements the most significant marker in RAI in separating cases from controls, which may suggest the presence of an independent, risk-modulating variation in this second gene region. PMID- 12117771 TI - Protocadherin LKC, a new candidate for a tumor suppressor of colon and liver cancers, its association with contact inhibition of cell proliferation. AB - Protocadherins are a major subfamily of the cadherin superfamily, but little is known about their functions and intracellular signal transduction. We cloned a novel human protocadherin gene, containing seven EC domains, and identified functional aspects of this gene. The gene was predominantly expressed in liver, kidney and colon tissues, and was thus designated Protocadherin LKC. The expression of Protocadherin LKC is markedly reduced in cancers arising from these tissues at both transcriptional and protein levels. To investigate the effects of Protocadherin LKC expression in colon cancer, we introduced the gene into colon cancer cell line HCT116, which does not express this gene. Significantly, Protocadherin LKC expression induced contact inhibition of cell proliferation although it did not affect growth rate. When grown to post-confluence in monolayer cells cultures, Protocadherin LKC-expressing HCT116 no longer formed multiple cell layers and showed the typical paving stone morphology of normal epithelial cells. Furthermore, expression of Protocadherin LKC suppressed tumor formation of HCT116 cells in a nude mouse model. In addition, we identified a protein, hMAST205 (microtubule-associated serine/threonine kinase-205 kDa), which interacted with Protocadherin LKC; the interaction occurring between the PDZ domain of hMAST205 and C-terminal tail of Protocadherin LKC. Our results suggest that Protocadherin LKC, which directly binds PDZ protein, is a molecular switch for contact inhibition of epithelial cells in the liver, kidney and colon tissues. PMID- 12117773 TI - Chemoprotective effects of garden cress (Lepidium sativum) and its constituents towards 2-amino-3-methyl-imidazo[4,5-f]quinoline (IQ)-induced genotoxic effects and colonic preneoplastic lesions. AB - The chemoprotective effect of garden cress (GC, Lepidium sativum) and its constituents, glucotropaeolin (GT) and benzylisothiocyanate (BITC), a breakdown product of GT, towards 2-amino-3-methyl-imidazo [4,5-f] quinoline (IQ)-induced genotoxic effects and colonic preneoplastic lesions was investigated in single cell gel electrophoresis (SCGE) assays and in aberrant crypt foci (ACF) experiments, respectively. Pretreatment of F344 rats with either fresh GC juice (0.8 ml), GT (150 mg/kg) or BITC (70 mg/kg) for three consecutive days caused a significant (P < 0.05) reduction in IQ (90 mg/kg, 0.2 ml corn oil/animal)-induced DNA damage in colon and liver cells in the range of 75-92%. Chemical analysis of GC juice showed that BITC does not account for the effects of the juice as its concentration in the juice was found to be 1000-fold lower than the dose required to cause a chemoprotective effect. Parallel to the chemoprotection experiments, the modulation of the activities of cytochrome P4501A2, glutathione-S-transferase (GST) and UDP glucuronosyltransferase (UDPGT) by GC juice, GT and BITC was studied. Whereas GT and BITC did not affect the activity of any of the enzymes significantly, GC juice caused a significant (P < 0.05) increase in the activity of hepatic UDPGT-2. In the ACF assay, IQ was administered by gavage on 10 alternating days in corn oil (dose 100 mg/kg). Five days before and during IQ treatment, subgroups received drinking water which contained 5% cress juice. The total number of IQ-induced aberrant crypts and ACF as well as ACF with crypt multiplicity of > or =4 were reduced significantly (P < 0.05) in the group that received IQ plus GC juice compared with the group that was fed with IQ only. However, crypt multiplicity was not significantly different in these two groups when all ACF with all classes of crypt multiplicity were considered in the analysis. This is the first report on the inhibition of HA-induced DNA damage and preneoplastic lesions by a cruciferous plant. Our findings suggest that the chemoprotective effect of GC is mediated through enhancement of detoxification of IQ by UDPGT. PMID- 12117774 TI - The roles of ERK1/2 and p38 MAP kinases in the preventive mechanisms of mushroom Phellinus linteus against the inhibition of gap junctional intercellular communication by hydrogen peroxide. AB - Modulation of gap junctional intercellular communication (GJIC) is a known cellular event associated with tumor promotion. The present study was undertaken to test the potential preventive effect of mushroom Phellinus linteus extract (PL) on the inhibition of GJIC, induced by hydrogen peroxide (H(2)O(2)), in WB F344 rat liver epithelial cells (WB cells). Cells were pre-incubated with PL (5 and 25 microg/ml) for 24 h and this was followed by co-treatment with PL and H(2)O(2) (500 microM) for 1 h. PL (at 5 and 25 microg/ml) prevented the inhibition of GJIC and blocked the hyper-phosphorylation of connexin 43 by H(2)O(2). Moreover, H(2)O(2) activated p38 kinase, extracellular signal-regulated protein kinases (ERK)1/2 and c-Jun N-terminal kinase (JNK) in WB cells. The present study indicates that PL is able to inactivate both ERK1/2 and p38 MAP kinases. However, PL did not affect the JNK pathway. For this reason, to elucidate the relation between MAP kinases and GJIC, we treated cells with PD98059 (an MEK inhibitor) and SB202190 (a p38 kinase inhibitor). These inhibitors were also found to prevent the inhibition of GJIC induced by H(2)O(2), which suggests that PL may act as a natural anticancer product by preventing the inhibition of GJIC through the inactivation of ERK1/2 and p38 MAP kinases. In addition, our results indicate that the p38 kinase signaling pathway may be closely related functionally to the gap junction in rat liver epithelial cells. PMID- 12117775 TI - A molecular dosimetry approach to assess human exposure to environmental tobacco smoke in pubs. AB - Although the involvement of environmental tobacco smoke (ETS) in human lung cancer is no longer a matter of dispute, the magnitude of its impact still is. This is mainly due to the inefficiency of methodology to assess exposure to ETS especially in public places. Setting a real life exposure condition (3 h stay in local pubs) and using a matched-control study design, we quantified smoke-related DNA adducts in induced sputum and peripheral blood lymphocytes (PBL) of healthy non-smokers (n = 15) before and after a single pub visit by means of the (32)P post-labeling assay. For verification, we also measured a spectrum of polycyclic aromatic hydrocarbons (PAH) in the ambient air of the pubs by personal air monitors, and determined the plasma concentrations of nicotine and cotinine by gas chromatography/mass spectrometry. The ambient air concentrations of all PAH were several orders of magnitude higher than those already reported for other indoor environments. The plasma concentrations of both nicotine and cotinine increased significantly after the pub visit (P = 0.001 and P = 0.0007, respectively). Accordingly, the overall DNA adduct profile in induced sputum, but not in PBL, changed quantitatively and qualitatively after the pub visit. Of most significance was the formation of a distinct DNA adduct in induced sputum of three individuals consequent to ETS exposure. This adduct co-migrated with the standard (+/-)-anti-benzo[a]pyrene diol epoxide-DNA adduct, which is known to form at lung cancer mutational hotspots. We conclude that real life exposure to ETS can give rise to pro-mutagenic lesions in the lower airway, and this can be best investigated in a relevant surrogate matrix such as induced sputum. PMID- 12117776 TI - Localization of a human lung adenocarcinoma susceptibility locus, possibly syntenic to the mouse Pas1 locus, in the vicinity of the D12S1034 locus on chromosome 12p11.2-p12.1. AB - Pulmonary adenoma susceptibility 1 (Pas1) is a major locus affecting inherited predisposition to the development of lung adenocarcinoma in mice, and is mapped to chromosome 6q near the Kras2 gene. However, it is still unclear whether the PAS1 locus on human chromosome 12p11.2-p12.1, the region showing synteny to the mouse Pas1 region, is involved in susceptibility to human lung adenocarcinoma development. Thus, we conducted a case-control study of 100 lung adenocarcinoma cases and 100 controls using 20 highly polymorphic microsatellite markers dispersed in a 13 cM region covering a putative PAS1 locus. The differences in the allele and genotype distributions were observed at several loci, and the difference was at a maximum at the D12S1034 locus (P = 0.034 and P = 0.036, respectively). The differences in the allele and genotype distributions at D12S1034 remained significant in the analysis in which 239 lung adenocarcinoma cases and 63 controls were added to the 100 cases and 100 controls used for the initial screening (P = 0.031 and P = 0.027, respectively). The D12S1034 locus was located 800-1350 kb proximal to the KRAS2 locus, and in the region syntenic to the core Pas1 region of approximately 1.5 Mb in size where a single haplotype is shared by several mouse-inbred strains susceptible to lung adenocarcinoma development. These results indicate that the PAS1 locus is located in the vicinity of D12S1034 and a genetic variation(s) at this locus is involved in susceptibility to human lung adenocarcinoma. PMID- 12117778 TI - Anomalous elevation of glutathione S-transferase P-form (GST-P) in the elementary process of epigenetic initiation of chemical hepatocarcinogenesis in rats. AB - The molecular mechanism of the specific expression of glutathione S-transferase P form (GST-P) in the rat hepatic preneoplastic foci and "GST-P-positive" single cells requires elucidation. Immunochemical and stereological analyses revealed that the enzyme level in preneoplastic foci was 150-250-fold (6.7 +/- 2.4 mg/g liver and 0.29 +/- 0.1 mM subunits) higher than in normal cells. GST-P content in the single cells was higher than in preneoplastic foci, as determined by densitometry. In addition, the single cells were larger in cell diameter and area, corresponding to 2-3-fold increase in cell volume, relative to normal cells, but showed a significant shrinkage of their nuclei. Prior to the induction of single cells in the liver by diethylnitrosamine (DEN), microsomes were severely damaged as reflected by the low yield (approximately 60% that of untreated controls) after 2 h of DEN injection. Considering that GST-P is mainly a binding protein for GSH conjugates of endogenous carcinogens, together with our findings of morphological expansion, low viability of single cells and microsomal damage, our results suggest anomalous elevation of the ligand counterparts to lethal levels in preneoplastic cells, especially in single cells. We propose that the epigenetic mechanism rather than the genetic mechanism could account for GST P induction in hepatocytes. PMID- 12117777 TI - Prevention of mouse lung tumors by budesonide and its modulation of biomarkers. AB - Chemopreventive drugs have the potential to decrease the morbidity and mortality of lung cancer. The development of these drugs could be expedited by the application of surrogate end-point biomarkers that demonstrate chemopreventive efficacy. In this study, the ability of budesonide to prevent lung tumors in mice was characterized further and its effects on biomarkers were determined. Lung tumors were induced in female strain A mice by vinyl carbamate (16 mg/kg) administered once weekly for 2 consecutive weeks. Four weeks later the mice started to receive 0.6, 1.2 or 2.4 mg/kg budesonide continually in the diet until killed at week 20. Budesonide caused a dose-dependent decrease in the multiplicity of lung tumors of 25, 58 and 82%, respectively. Budesonide (2.4 mg/kg diet) administered starting at weeks 4, 10 or 16, decreased tumor multiplicity by 82, 66 and 30% at week 20. Administering 2.4 mg/kg budesonide at weeks 4-20 or 20-35 and killing the mice at week 35 did not significantly decrease the yield of tumors, although both treatment regimens did decrease the size of the tumors and the progression of adenomas to carcinomas. Thus, budesonide delayed the appearance of lung tumors and decreased their growth and progression to carcinomas. To determine the effect of limited exposure to budesonide on biomarkers, it was administered for only 7 days prior to death at week 35. Budesonide decreased the proliferating cell nuclear antigen labeling in lung adenomas, carcinomas, parenchyma and bronchial airways by 87.6, 59.0, 41.1 and 25.4%, respectively. Budesonide treatment also increased the protein level of the p21 and p27 genes and increased the mRNA level of p21. Thus, short-term treatment with budesonide modulated biological and molecular end-points in lung tumors that might be developed further as biomarkers for its clinical chemopreventive efficacy in the lung. PMID- 12117779 TI - Arylhydrocarbon receptor-dependent induction of liver and lung cytochromes P450 1A1, 1A2, and 1B1 by polycyclic aromatic hydrocarbons and polychlorinated biphenyls in genetically engineered C57BL/6J mice. AB - Arylhydrocarbon receptor knock-out, AhR(-/-), mice have recently been shown to be rather resistant to benzo[a]pyrene (B[a]P)-induced tumor formation, probably reflecting the inability of these mice to express significant levels of cytochrome P450 (P450 or CYP) 1A1 that activates B[a]P to reactive metabolites (Y. Shimizu, Y. Nakatsuru, M. Ichinose, Y. Takahashi, H. Kume, J. Mimura, Y. Fujii-Kuriyama and T. Ishikawa (2000) PROC: Natl Acad. Sci. USA, 97, 779-782). However, it is not precisely determined whether CYP1B1, another enzyme that is also active in activating B[a]P, plays a role in the B[a]P carcinogenesis in mice. To understand the basis of roles of CYP1A1 and CYP1B1 in the activation of chemical carcinogens, we compared levels of induction of liver and lung CYP1A1, 1A2, and 1B1 by various polycyclic aromatic hydrocarbons (PAHs) and polychlorinated biphenyls in AhR(+/+) and AhR(-/-) mice. Liver and lung CYP1A1 and 1B1 mRNAs were highly induced in AhR(+/+) mice by a single intraperitoneal injection of each of the carcinogenic PAHs, such as B[a]P, 7,12 dimethylbenz[a]anthracene, dibenz[a,l]pyrene, 3-methylcholanthrene, 1,2,5,6 dibenzanthracene, benzo[b]fluoranthene, and benzo[a]anthracene and by a co-planar PCB congener 3,4,3',4'-tetrachlorobiphenyl. We also found that 6-aminochrysene, chrysene, benzo[e]pyrene, and 1-nitropyrene weakly induced the mRNA expression of CYP1A1 and 1B1, whereas anthracene, pyrene, and fluoranthene that have been reported to be non-carcinogenic in rodents, were very low or inactive in inducing these P450s. The extents of induction of liver CYP1A2 by these chemicals were less than those of CYP1A1 and 1B1 in AhR(+/-/+/-) mice. In AhR(-/-) mice, there was no induction of these P450s by PAHs and polychlorinated biphenyls. Liver microsomal activities of 7-ethoxyresorufin and 7-ethoxycoumarin O-deethylations and of mutagenic activation of (+/-)-trans-7,8-dihydroxy-7,8-dihydro-B[a]P to DNA damaging products were found to correlate with levels of CYP1A1 and 1B1 mRNAs in the liver. Our results suggest that carcinogenicity potencies of PAHs may relate to the potencies of these compounds to induce CYP1A1 and 1B1 through AhR dependent manner and that these induced P450s participate in the activation of B[a]P and related carcinogens causing initiation of cancers in mice. PMID- 12117780 TI - Inhibitory effects of 17beta-estradiol and 4-n-octylphenol on 7,12 dimethylbenz[a]anthracene-induced mammary tumor development in human c-Ha-ras proto-oncogene transgenic rats. AB - Experiments were conducted to determine whether the natural estrogen and an environmental compound with estrogenic action, 4-n-octylphenol (4nOP), could modify tumor development in human c-Ha-ras proto-oncogene transgenic (Tg) rats which are highly susceptible to mammary and skin carcinogens. Female and male Tg and non-transgenic (non-Tg) rats were given a single oral dose of 7,12 dimethylbenz[a]anthracene (DMBA) (25 mg/kg body weight) at 50 days of age and thereafter subcutaneously implanted with cholesterol pellets containing 0.01, 0.1 or 1.0 mg beta-estradiol 3-benzoate (E2) per rat or received diets containing 1000 or 100 p.p.m. 4nOP for 12 weeks in females or for 20 weeks in males. E2 reduced the mammary tumor incidence and multiplicity in a dose dependent manner, especially in female Tg rats. In contrast, E2 increased mammary tumor incidence and multiplicity at the lowest dose (0.01 mg), however it reduced skin tumor induction in male Tg rats. 4nOP at a dose of 100 p.p.m. decreased mammary tumor multiplicity in female Tg rats (P < 0.001). No effects were observed in males. In separate in vitro studies, E2 at low doses (10(-11)-10(-8) M) enhanced the growth of both MCF-7 and T47D cells and this was similarly the case for 4nOP at high doses (10(-7)-10(-5) M) in T47D cells. The finding that E2 and 4nOP at high doses caused reduction in mammary tumor development in female Tg and possibly non-Tg rats, may indicate that excess estrogen can exert a paradoxical inhibitory influence. E2 also appears to have bipotential effects in males, promoting mammary, but inhibiting skin carcinogenesis. These contrasting observations may be caused by differences in background physiological estrogen levels. In addition, the results suggest that Tg rats can be used in medium-term bioassay models to test for the modifying effects of estrogenic environmental compounds on mammary tumor development. PMID- 12117781 TI - Altered expression of G1/S regulatory genes occurs early and frequently in lung carcinogenesis in transforming growth factor-beta1 heterozygous mice. AB - We developed the AJBL6 transforming growth factor-beta 1 (TGF-beta1) heterozygous (HT) mouse by mating A/J mice with C57BL/6 TGF-beta1 HT mice that shows increased carcinogen-induced lung lesions with decreased latency to examine progressive events in lung tumorigenesis. Mouse cDNA macroarrays were used to identify cell cycle genes that are differentially regulated in ethyl carbamate-induced lung adenocarcinomas compared with normal lung tissue in AJBL6 TGF-beta1 HT mice using probes that were generated from tissues isolated using laser capture microdissection. While expression of the genes for cyclin D1, CDK4, and E2F1 increased in lung adenocarcinomas relative to normal lung, expression of p15(Ink4b), p16(Ink4a), p21(Cip1), p27(Kip1), p57(Kip2), and pRb genes decreased in comparison. Competitive RT-PCR showed that the levels of cyclin D1 and CDK4 mRNAs were 2- and 3-fold higher, respectively, in lung adenocarcinomas than in normal lung, while the mRNAs for p15(Ink4b), p16(Ink4a), p21(Cip1), p27(Kip1), and pRb were 3- to 4-fold lower in adenocarcinomas than in normal lung, thus validating the macroarray findings. Competitive RT-PCR of microdissected lesions also showed that the levels of cyclin D1 and CDK4 mRNAs increased significantly, while the mRNAs for p15(Ink4b) and p27(Kip1) decreased significantly as lung tumorigenesis progressed. Immunohistochemical staining for cyclin D1 and CDK4 showed staining in >80% of nuclei in adenocarcinomas compared with fewer than 20% of nuclei staining positively in normal lung. In contrast, while >60% of normal lung cells showed immunostaining for p15(Ink4b), p16(Ink4a), p21(Cip1), p27(Kip1), and pRb, staining for these proteins decreased in hyperplasias, adenomas, and adenocarcinomas. These data show that multiple components of the cyclin D1/CDK4/p16(Ink4a)/pRb signaling pathway are frequently altered early in lung lesions of AJBL6 TGF-beta1 HT mice that are induced by ethyl carbamate as a function of progressive lung carcinogenesis, suggesting that components of this pathway may be potential targets for gene therapy. PMID- 12117782 TI - The human OGG1 DNA repair enzyme and its association with orolaryngeal cancer risk. AB - The human OGG1 (hOGG1) gene encodes a DNA glycosylase that is involved in the excision repair of 8-hydroxy-2'-deoxyguanine (8-OH-dG) from oxidatively-damaged DNA. To determine whether hOGG1 plays a role in risk for orolaryngeal cancer, we screened normal orolaryngeal tissue specimens for hOGG1 expression and assessed the role of the hOGG1 Ser326Cys polymorphism in risk for orolaryngeal cancer. hOGG1 expression was determined by reverse transcription-polymerase chain reaction of total RNA from aerodigestive tract tissues, and hOGG1 genotyping was performed by polymerase chain reaction-restriction fragment length polymorphism analysis of buccal cell DNA isolated from 169 Caucasian orolaryngeal cancer cases and 338 race-, sex- and age-matched controls. hOGG1 mRNA was detected in all aerodigestive tract tissues tested including tonsil, tongue, floor of mouth, larynx and esophagus. Significantly increased risk for orolaryngeal cancer was observed for both the hOGG1 326(Ser)/326(Cys) (odds ratio [OR] = 1.6, 95% confidence interval [CI] = 1.04-2.6) and hOGG1 326(Cys)/326(Cys) (OR = 4.1, 95% CI = 1.3-13) genotypes. Although no significant difference in risk for orolaryngeal cancer was observed for hOGG1 genotypes in never-smokers, increased risk for orolaryngeal cancer was observed for subjects with the homozygous polymorphic hOGG1 326(Cys)/326(Cys) genotype in smokers (>100 cigarettes lifetime; OR = 4.8, 95% CI = 1.3-18). Similarly, although no association was observed in never drinkers of alcohol, significantly increased risk was observed for the hOGG1 326(Cys)/326(Cys) genotype in alcohol drinkers (>1 shot/week; OR = 6.9, 95% CI = 1.6-29). These results suggest that hOGG1 may play an important role in the repair of 8-OH-dG adducts in the aerodigestive tract and that the hOGG1 Ser326Cys polymorphism plays an important role in risk for smoking- and alcohol-related orolaryngeal cancer. PMID- 12117783 TI - Hemizygous mice for the angiotensin II type 2 receptor gene have attenuated susceptibility to azoxymethane-induced colon tumorigenesis. AB - Evidence suggests that the use of angiotensin-converting enzyme inhibitors potentially reduces the risk of cancer, though the mechanism is unclear. To clarify a potential involvement of angiotensin II (Ang II) signaling in cancer risk, we have examined the effect of Ang II receptor deficiency on azoxymethane (AOM)-induced colon tumorigenesis. Male Ang II type 2 receptor gene-disrupted (AT(2)-null) mice with a 129/Ola and C57BL/6J genetic background, AT(2)-null mice with an SWR/J genetic background, and their corresponding control wild type mice were treated once a week with AOM (10 mg/kg, i.p., 4 consecutive weeks) or saline vehicle. All mice were killed 23-26 weeks after the initial injection of AOM, and tumor burdens were examined. AOM treatment caused the development of colon tumors in all wild type control mice regardless of genetic background (100% tumor prevalence), but only one tumor was present in AT(2)-null mice with a 129/Ola and C57BL/6J genetic background (11.1% tumor prevalence). Although the introduction of the AOMsusceptible SWR/J genetic background induced AOM susceptibility in AT(2) null mice, the tumor multiplicity (6.3) and tumor size (19.8 +/- 3.0 mm(3)) were significantly smaller than those in wild type mice (multiplicity, 12.0 and size, 36.8 +/- 3.2 mm(3)). AOM efficiently downregulated cytochrome P450 2E1 (CYP2E1) in the liver of wild type mice significantly more than in AT(2)-null mice. The levels of DNA methyl adducts formed in wild type mouse colon epithelium by AOM treatment were also significantly higher than in AT(2)-null mice. These results imply that the AT(2) receptor functions to augment AOM-induced downregulation of CYP2E1 expression in the liver, and thus increases AOM-induced tumorigenesis in the colon. The AT(2) receptor function in the liver may be a potential determinant of tumor susceptibility in chemical carcinogen-induced colon tumorigenesis. PMID- 12117784 TI - Decreased gap junctional intercellular communication in hexachlorobenzene-induced gender-specific hepatic tumor formation in the rat. AB - Hexachlorobenzene (HCB), an epigenetic carcinogen, HCB induces the formation of liver tumors in female rats, whereas only a small percentage of males are responsive. Intercellular communication via gap junctions is decreased in carcinogenesis. Gap junctions are composed of proteins termed connexins (Cxs). The objectives of this study were (i) to determine if HCB-induced tumor development is associated with a loss of gap junctional communication; (ii) to assess if HCB causes a gender-specific decrease in the expression of Cx32 and Cx26; and (iii) to establish if these effects result from gender differences in the constitutive expression of these Cxs. Rats were given HCB by gavage for five consecutive days. In the first experiment, control and HCB-treated female rats were sampled on day 100. Intercellular communication was significantly decreased in HCB-treated females compared to controls. To investigate if changes in Cx levels occur prior to day 100, experiments done using male and female rats sampled on day 50. Hepatic mRNA levels for Cx26 and Cx32 were significantly lower only in HCB-treated females as compared to controls. Cx26 mRNA levels were 3-fold higher and Cx32 mRNA levels were 8-fold lower in females compared with males. In a third experiment, ovariectomy abolished any differences between male and female controls for both Cxs, while estradiol had a partial role in the regulation of Cx32. This suggests that the sexual dimorphism in hepatic Cx levels is determined by the ovarian hormones. However, the HCB-induced decrease in Cx32 and Cx26 mRNA levels was maintained in ovariectomized rats, suggesting that the HCB effects are not mediated via an ovary-dependent pathway. Overall results show that HCB exposure induces gender-specific long-term alterations in intercellular gap junctional communication in female rat liver. This effect appears to be a critical mechanism of HCB-induced liver carcinogenesis and tumor promotion. PMID- 12117786 TI - Bioinformatics in structural genomics. PMID- 12117785 TI - Role of Raf-1 and FAK in cell density-dependent regulation of integrin-dependent activation of MAP kinase. AB - MAP kinase can be activated by integrin-dependent adhesion in a FAK-dependent manner. Cell-cell contact inhibition is continuously active in controlling cell growth and the loss of cell-cell contact inhibition is correlated with the malignant characteristics of cancer cells. In this study we showed that cell adhesion to fibronectin for 1 h activated MAP kinase phosphorylation. However, when non-tumorigenic HSG cells, MCF-10A cells, or 293 cells were plated on fibronectin-coated substrates for 1 h at high cell density (which favors cell cell contact), MAP kinase phosphorylation was not enhanced. Tumorigenic breast cancer cells, BT474, Cama, MCF-7, MDA-MB-231 and SKBR3, did not show inhibition of MAP kinase phosphorylation but rather enhanced MAP kinase phosphorylation when cultured at high density on fibronectin-coated substrates. Adhesion of HSG cells to fibronectin also increased FAK phosphorylation and this FAK phosphorylation was partially inhibited when cells were cultured at high density. Expression of Raf-1 catalytic domain-GFP in HSG cells could overcome the cell density-dependent inhibition of MAP kinase phosphorylation and FAK phosphorylation. The expression of Raf-1-catalytic domain-GFP also upregulated the expression of alphav integrin and promoted cell-cell adhesion in HSG cells. These results suggest that the active form of Raf-1 may interrupt cell-cell contact inhibition by promoting alphav integrin expression, which has been implicated in cell aggregation. PMID- 12117787 TI - Selecting targets for structural determination by navigating in a graph of protein families. AB - MOTIVATION: A major goal in structural genomics is to enrich the catalogue of proteins whose 3D structures are known. In an attempt to address this problem we mapped over 10 000 proteins with solved structures onto a graph of all Swissprot protein sequences (release 36, approximately 73 000 proteins) provided by ProtoMap, with the goal of sorting proteins according to their likelihood of belonging to new superfamilies. We hypothesized that proteins within neighbouring clusters tend to share common structural superfamilies or folds. If true, the likelihood of finding new superfamilies increases in clusters that are distal from other solved structures within the graph. RESULTS: We defined an order relation between unsolved proteins according to their 'distance' from solved structures in the graph, and sorted approximately 48 000 proteins. Our list can be partitioned into three groups: approximately 35 000 proteins sharing a cluster with at least one known structure; approximately 6500 proteins in clusters with no solved structure but with neighbouring clusters containing known structures; and a third group contains the rest of the proteins, approximately 6100 (in 1274 clusters). We tested the quality of the order relation using thousands of recently solved structures that were not included when the order was defined. The tests show that our order is significantly better (P-value approximately 10(5)) than a random order. More interestingly, the order within the union of the second and third groups, and the order within the third group alone, perform better than random (P-values: 0.0008 and 0.15, respectively) and are better than alternative orders created using PSI-BLAST. Herein, we present a method for selecting targets to be used in structural genomics projects. AVAILABILITY: List of proteins to be used for targets selection combined with a set of biological filters for narrowing down potential targets is in http://www.protarget.cs.huji.ac.il. PMID- 12117788 TI - Clustering of proximal sequence space for the identification of protein families. AB - MOTIVATION: The study of sequence space, and the deciphering of the structure of protein families and subfamilies, has up to now been required for work in comparative genomics and for the prediction of protein function. With the emergence of structural proteomics projects, it is becoming increasingly important to be able to select protein targets for structural studies that will appropriately cover the space of protein sequences, functions and genomic distribution. These problems are the motivation for the development of methods for clustering protein sequences and building families of potentially orthologous sequences, such as those proposed here. RESULTS: First we developed a clustering strategy (Ncut algorithm) capable of forming groups of related sequences by assessing their pairwise relationships. The results presented for the ras super family of proteins are similar to those produced by other clustering methods, but without the need for clustering the full sequence space. The Ncut clusters are then used as the input to a process of reconstruction of groups with equilibrated genomic composition formed by closely-related sequences. The results of applying this technique to the data set used in the construction of the COG database are very similar to those derived by the human experts responsible for this database. AVAILABILITY: The analysis of different systems, including the COG equivalent 21 genomes are available at http://www.pdg.cnb.uam.es/GenoClustering.html. PMID- 12117789 TI - Target space for structural genomics revisited. AB - MOTIVATION: Structural genomics eventually aims at determining structures for all proteins. However, in the beginning experimentalists are likely to focus on globular proteins to achieve a rapid basic coverage of protein sequence space. How many proteins will structural genomics have to target? How many proteins will be excluded since we already have structural information for these or since they are not globular? We have to answer these questions in the context of our target selection for the North-East Structural Genomics Consortium (NESG). RESULTS: We estimated that structural information is available for about 6-38% of all proteins; 6% if we require high accuracy in comparative modelling, 38% if we are satisfied with having a rough idea about the fold. Excluding all regions that are not globular, we found that structural genomics may have to target about 48% of all proteins. This corresponded to a similar percentage of residues of the entire proteomes (52%). We explored a number of different strategies to cluster protein space in order to find the number of families representing these 48% of structurally unknown proteins. For the subset of all entirely sequenced eukaryotes, we found over 18 000 fragment clusters each of which may be a suitable target for structural genomics. AVAILABILITY: All data are available from the authors, most results are summarized at: http://cubic.bioc.columbia.edu/genomes/RES/2002_bioinformatics/ PMID- 12117790 TI - About the use of protein models. AB - Protein models can be of great assistance in functional genomics, as they provide the structural insights often necessary to understand protein function. Although comparative modelling is far from yielding perfect structures, this is still the most reliable method and the quality of the predictions is now well understood. Models can be classified according to their correctness and accuracy, which will impact their applicability and usefulness in functional genomics and a variety of situations. PMID- 12117791 TI - Ligand-induced changes in the binding sites of proteins. AB - Classical molecular interaction potentials, in conjunction with other theoretical techniques, are used to analyze the dependence of the binding sites of representative proteins on the bound ligand. It is found that the ligand bound introduces in general small structural perturbations at the binding site of the protein. However, such small structural changes can lead to important alterations in the recognition pattern of the protein. The impact of these findings in docking procedures is discussed. PMID- 12117792 TI - A bioinformatic strategy to rapidly characterize cDNA libraries. AB - MOTIVATION: Complementary DNA libraries can define the genetic constituents of specific cells and/or tissues. Their sequencing will illuminate the transcriptome but it is a monumental task requiring considerable resources. RESULTS: We have employed a computational search in conjunction with a microarray-based strategy to alleviate the impediments of deriving a consensus of records that describe testis gene expression. This strategy identified 5681 unique testes-expressed genes of which 3265 were previously portrayed in the UniGene database. Interestingly, a total of 2416 novel testes-expressed genes were identified. This clearly demonstrates that microarrays can be used to rapidly discover a large number of new transcripts. PMID- 12117793 TI - Quantitative assessment of filter-based cDNA microarrays: gene expression profiles of human T-lymphoma cell lines. AB - MOTIVATION: While the use of cDNA microarrays for functional genomic analysis has become commonplace, relatively little attention has been placed on false positives, i.e. the likelihood that a change in measured radioactive or fluorescence intensity may reflect a change in gene expression when, in fact, there is none. Since cDNA arrays are being increasingly used to rapidly distinguish biomarkers for disease detection and subsequent assay development (Wellman et al., Blood, 96, 398-404, 2000), the impact of false positives can be significant. For the use of this technology, it is necessary to develop quantitative criteria for reduction of false positives with radioactively-labeled cDNA arrays. RESULTS: We used a single source of RNA (HuT78 T lymphoma cells) to eliminate sample variation and quantitatively examined intensity ratios using radioactively labeled cDNA microarrays. Variation in intensity ratios was reduced by processing microarrays in side-by-side (parallel mode) rather than by using the same microarray for two hybridizations (sequential mode). Based on statistical independence, calculation of the expected number of false positives as a function of threshold showed that a detection limit of [log(2)R] >0.65 with agreement from three replicates could be used to identify up- or down-modulated genes. Using this quantitative criteria, gene expression differences between two related T lymphoma cell lines, HuT78 and H9, were identified. The relevance of these findings to the known functional differences between these cell types is discussed. PMID- 12117794 TI - Deriving quantitative conclusions from microarray expression data. AB - MOTIVATION: The last few years have seen the development of DNA microarray technology that allows simultaneous measurement of the expression levels of thousands of genes. While many methods have been developed to analyze such data, most have been visualization-based. Methods that yield quantitative conclusions have been diverse and complex. RESULTS: We present two straightforward methods for identifying specific genes whose expression is linked with a phenotype or outcome variable as well as for systematically predicting sample class membership: (1) a conservative, permutation-based approach to identifying differentially expressed genes; (2) an augmentation of K-nearest-neighbor pattern classification. Our analyses replicate the quantitative conclusions of Golub et al. (1999; Science, 286, 531-537) on leukemia data, with better classification results, using far simpler methods. With the breast tumor data of Perou et al. (2000; Nature, 406, 747-752), the methods lend rigorous quantitative support to the conclusions of the original paper. In the case of the lymphoma data in Alizadeh et al. (2000; Nature, 403, 503-511), our analyses only partially support the conclusions of the original authors. AVAILABILITY: The software and supplementary information are available freely to researchers at academic and non profit institutions at http://cc.ucsf.edu/jain/public PMID- 12117795 TI - Comparative genomics study of inverted repeats in bacteria. AB - MOTIVATION: Comparative genomics provides a powerful way to investigate regularities and differences observed at DNA level across species. Here we study the number and location of inverted repeats occurring in complete genomes of bacteria. Inverted repeats are compatible with the formation of hairpin structures in the messenger RNA. Some of these structures are known to be rho independent intrinsic terminators. RESULTS: We investigate the number of inverted repeats observed in 37 complete genomes of bacteria. The number of inverted repeats observed is much higher than expected using Markovian models of DNA sequences in most of the eubacteria. By using the information annotated in the genomes we discover that in most of the eubacteria the inverted repeats of stem length longer than 8 nucleotides preferentially locate near the 3' end of the nearest coding regions. We also show that IRs characterized by large values of the stem length locate preferentially in short non-coding regions bounded by two 3' ends of convergent genes. By using the program TransTerm recently introduced to predict transcription terminators in bacterial genomes, we conclude that only a part of the observed inverted repeats fullfils the model requirements characterizing rho-independent termination in several genomes. AVAILABILITY: http://lagash.dft.unipa.it/IR.html PMID- 12117796 TI - CX, an algorithm that identifies protruding atoms in proteins. AB - MOTIVATION: A simple and fast algorithm is described that calculates a measure of protrusion (cx) for atoms in protein structures, directly useable with the common molecular graphics programs. RESULTS: A sphere of predetermined radius is centered around each non-hydrogen atom, and the volume occupied by the protein and the free volume within the sphere (internal and external volumes, respectively) are calculated. Atoms in protruding regions have a high ratio (cx) between the external and the internal volume. The program reads a PDB file, and writes the output in the same format, with cx values in the B factor field. Output structure files can be directly displayed with standard molecular graphics programs like RASMOL, MOLMOL, Swiss-PDB Viewer and colored according to cx values. We show the potential use of this program in the analysis of two protein protein complexes and in the prediction of limited proteolysis sites in native proteins. AVAILABILITY: The algorithm is implemented in a standalone program written in C and its source is freely available at ftp.icgeb.trieste.it/pub/CX or on request from the authors. PMID- 12117798 TI - PATIKA: an integrated visual environment for collaborative construction and analysis of cellular pathways. AB - MOTIVATION: Availability of the sequences of entire genomes shifts the scientific curiosity towards the identification of function of the genomes in large scale as in genome studies. In the near future, data produced about cellular processes at molecular level will accumulate with an accelerating rate as a result of proteomics studies. In this regard, it is essential to develop tools for storing, integrating, accessing, and analyzing this data effectively. RESULTS: We define an ontology for a comprehensive representation of cellular events. The ontology presented here enables integration of fragmented or incomplete pathway information and supports manipulation and incorporation of the stored data, as well as multiple levels of abstraction. Based on this ontology, we present the architecture of an integrated environment named Patika (Pathway Analysis Tool for Integration and Knowledge Acquisition). Patika is composed of a server-side, scalable, object-oriented database and client-side editors to provide an integrated, multi-user environment for visualizing and manipulating network of cellular events. This tool features automated pathway layout, functional computation support, advanced querying and a user-friendly graphical interface. We expect that Patika will be a valuable tool for rapid knowledge acquisition, microarray generated large-scale data interpretation, disease gene identification, and drug development. AVAILABILITY: A prototype of Patika is available upon request from the authors. PMID- 12117797 TI - Calculations of protein volumes: sensitivity analysis and parameter database. AB - MOTIVATION: The precise sizes of protein atoms in terms of occupied packing volume are of great importance. We have previously presented standard volumes for protein residues based on calculations with Voronoi-like polyhedra. To understand the applicability and limitations of our set, we investigated, in detail, the sensitivity of the volume calculations to a number of factors: (i) the van der Waals radii set, (ii) the criteria for including buried atoms in the calculations or atom selection, (iii) the method of positioning the dividing plane in polyhedra construction, and (iv) the set of structures used in the averaging. RESULTS: We find that different radii sets have only moderate affects to the distribution and mean of volumes. Atom selection and dividing plane methods cause larger changes in protein atoms volumes. More significantly, we show how the variation in volumes appears to be clearly related to the quality of the structures analyzed, with higher quality structures giving consistently smaller average volumes with less variance. PMID- 12117799 TI - Determining a unique defining DNA sequence for yeast species using hashing techniques. AB - MOTIVATION: Yeasts are often still identified with physiological growth tests, which are both time consuming and unsuitable for detection of a mixture of organisms. Hence, there is a need for molecular methods to identify yeast species. RESULTS: A hashing technique has been developed to search for unique DNA sequences in 702 26S rRNA genes. A unique DNA sequence has been found for almost every yeast species described to date. The locations of the unique defining sequences are in accordance with the variability map of large subunit ribosomal RNA and provide detail of the evolution of the D1/D2 region. This approach will be applicable to the rapid identification of unique sequences in other DNA sequence sets. AVAILABILITY: Freely available upon request from the authors. SUPPLEMENTARY INFORMATION: Results are available at http://www.sys.uea.ac.uk/~jjw/project/paper PMID- 12117800 TI - TangleSolve: topological analysis of site-specific recombination. AB - TangleSolve is a program for analysing site-specific recombination using the tangle model. The program offers an easy-to-use graphical user interface and a visualization tool. Biologists working in topological enzymology can use this program to compute and visualize site-specific recombination mechanisms that accommodate their experimental data. TangleSolve can also prove useful as a teaching aid for mathematical biology and computational molecular biology courses. AVAILABILITY: http://bio.math.berkeley.edu/TangleSolve/ PMID- 12117801 TI - CaGE: cardiac gene expression knowledgebase. AB - CaGE is a Cardiac Gene Expression knowledgebase we have developed to facilitate the analysis of genes important to human cardiac function. CaGE integrates the functionality of the LocusLink database with data from several human cardiac expression libraries, phenotypic data from OMIM and data from large-scale microarray gene expression studies to create a knowledgebase of gene expression in human cardiac tissue. The knowledgebase is fully searchable via the web using several intuitive query interfaces. Results can be displayed in several concise easy to navigate formats. AVAILABILITY: CaGE is located at http://www.cage.wbmei.jhu.edu PMID- 12117802 TI - CS-PSeq-Gen: simulating the evolution of protein sequence under constraints. AB - CS-PSeq-Gen is a program derived from PSeq-Gen, designed to perform simulations of the evolution of protein sequences under the constraints of a reconstructed phylogeny. It also provides a basis for the investigation of the correlated evolution of sites. AVAILABILITY: http://condor.urbb.jussieu.fr/CS-PSeq-Gen.html PMID- 12117803 TI - SNAPper: gene order predicts gene function. AB - SNAPper is a network service for predicting gene function based on the conservation of gene order. AVAILABILITY: The SNAPper server is available at http://pedant.gsf.de/snapper. SNAPper-based functional predictions will soon be offered as part of the PEDANT genome analysis server http://pedant.gsf.de. PMID- 12117805 TI - Engrailed homeoprotein secretion is a regulated process. AB - Chicken Engrailed 2 homeoprotein is transported between cells in culture. This intercellular transfer is based on unconventional secretion and internalisation mechanisms: Engrailed 2 has access to vesicles but lacks a signal sequence for secretion and is internalised by a non-endocytic process. We show that phosphorylation of a serine-rich domain within Engrailed 2 by the protein kinase CK2 specifically inhibits Engrailed 2 secretion. The availability of the serine rich domain to CK2 is highly increased when it is displaced from its normal position to the C terminus of Engrailed 2, leading to a constitutive blockage of Engrailed 2 intercellular transfer. This indicates that intercellular transfer of Engrailed 2 is a highly regulated process. PMID- 12117804 TI - PLC zeta: a sperm-specific trigger of Ca(2+) oscillations in eggs and embryo development. AB - Upon fertilisation by sperm, mammalian eggs are activated by a series of intracellular Ca(2+) oscillations that are essential for embryo development. The mechanism by which sperm induces this complex signalling phenomenon is unknown. One proposal is that the sperm introduces an exclusive cytosolic factor into the egg that elicits serial Ca(2+) release. The 'sperm factor' hypothesis has not been ratified because a sperm-specific protein that generates repetitive Ca(2+) transients and egg activation has not been found. We identify a novel, sperm specific phospholipase C, PLC zeta, that triggers Ca(2+) oscillations in mouse eggs indistinguishable from those at fertilisation. PLC zeta removal from sperm extracts abolishes Ca(2+) release in eggs. Moreover, the PLC zeta content of a single sperm was sufficient to produce Ca(2+) oscillations as well as normal embryo development to blastocyst. Our results are consistent with sperm PLC zeta as the molecular trigger for development of a fertilised egg into an embryo. PMID- 12117806 TI - Evolving role of Antennapedia protein in arthropod limb patterning. AB - Evolutional changes in homeotic gene functions have contributed to segmental diversification of arthropodan limbs, but crucial molecular changes have not been identified to date. The first leg of the crustacean Daphnia lacks a prominent ventral branch found in the second to fourth legs. We show here that this phenotype correlates with the loss of Distal-less and concomitant expression of Antennapedia in the limb primordium. Unlike its Drosophila counterpart, Daphnia Antennapedia represses Distal-less in Drosophila assays, and the protein region conferring this activity was mapped to the N terminal region of the protein. The results imply that Dapnia Antennapedia specifies leg morphology by repressing Distal-less, and this activity was acquired through a change in protein structure after separation of crustaceans and insects. PMID- 12117807 TI - Embryonic retinoic acid synthesis is required for forelimb growth and anteroposterior patterning in the mouse. AB - Numerous studies, often performed on avian embryos, have implicated retinoic acid (RA) in the control of limb bud growth and patterning. Here we have investigated whether the lack of endogenous RA synthesis affects limb morphogenesis in mutant mouse embryos deficient for the retinaldehyde dehydrogenase 2 (Raldh2/Aldh1a2). These mutants, which have no detectable embryonic RA except in the developing retina, die at E9.5-E10 without any evidence of limb bud formation, but maternal RA supplementation through oral gavage from E7.5 can extend their survival. Such survivors exhibit highly reduced forelimb rudiments, but apparently normal hindlimbs. By providing RA within maternal food, we found both a stage- and dose dependency for rescue of forelimb growth and patterning. Following RA supplementation from E7.5 to 8.5, mutant forelimbs are markedly hypoplastic and lack anteroposterior (AP) patterning, with a single medial cartilage and 1-2 digit rudiments. RA provided until E9.5 significantly rescues forelimb growth, but cannot restore normal AP patterning. Increasing the RA dose rescues the hypodactyly, but leads to lack of asymmetry of the digit pattern, with abnormally long first digit or symmetrical polydactyly. Mutant forelimb buds are characterized by lack of expression or abnormal distal distribution of Sonic hedgehog (Shh) transcripts, sometimes with highest expression anteriorly. Downregulation or ectopic anterior expression of Fgf4 is also seen. As a result, genes such as Bmp2 or Hoxd genes are expressed symmetrically along the AP axis of the forelimb buds, and/or later, of the autopod. We suggest that RA signaling cooperates with a posteriorly restricted factor such as dHand, to generate a functional zone of polarizing activity (ZPA). PMID- 12117808 TI - The role of TGF beta signaling in the formation of the dorsal nervous system is conserved between Drosophila and chordates. AB - Transforming growth factor beta signaling mediated by Decapentaplegic and Screw is known to be involved in defining the border of the ventral neurogenic region in the fruitfly. A second phase of Decapentaplegic signaling occurs in a broad dorsal ectodermal region. Here, we show that the dorsolateral peripheral nervous system forms within the region where this second phase of signaling occurs. Decapentaplegic activity is required for development of many of the dorsal and lateral peripheral nervous system neurons. Double mutant analysis of the Decapentaplegic signaling mediator Schnurri and the inhibitor Brinker indicates that formation of these neurons requires Decapentaplegic signaling, and their absence in the mutant is mediated by a counteracting repression by Brinker. Interestingly, the ventral peripheral neurons that form outside the Decapentaplegic signaling domain depend on Brinker to develop. The role of Decapentaplegic signaling on dorsal and lateral peripheral neurons is strikingly similar to the known role of Transforming growth factor beta signaling in specifying dorsal cell fates of the lateral (later dorsal) nervous system in chordates (Halocythia, zebrafish, Xenopus, chicken and mouse). It points to an evolutionarily conserved mechanism specifying dorsal cell fates in the nervous system of both protostomes and deuterostomes. PMID- 12117809 TI - Elbow and Noc define a family of zinc finger proteins controlling morphogenesis of specific tracheal branches. AB - The elbow (elB) gene encodes a conserved nuclear protein with a single zinc finger. Expression of ElB is restricted to a specific subset of tracheal cells, namely the dorsal branch and the lateral trunks. Stalled or aberrant migration of these branches is observed in elB mutant embryos. Conversely, ElB misexpression in the trachea gave rise to absence of the visceral branch and an increase in the number of cells forming the dorsal branch. These results imply that the restricted expression of ElB contributes to the specification of distinct branch fates, as reflected in their stereotypic pattern of migration. As elB loss-of function tracheal phenotypes are reminiscent of defects in Dpp signaling, the relationship between ElB and the Dpp pathway was examined. By using pMad antibodies that detect the activation pattern of the Dpp pathway, we show that Dpp signaling in the trachea is not impaired in elB mutants. In addition, expression of the Dpp target gene kni was unaltered. The opposite is true as well, because expression of elB is independent of Dpp signaling. ElB thus defines a parallel input, which determines the identity of the lateral trunk and dorsal branch cells. No ocelli (Noc) is the Drosophila protein most similar to ElB. Mutations in noc give rise to a similar tracheal phenotype. Noc is capable of associating with ElB, suggesting that they can function as a heterodimer. ElB also associates with the Groucho protein, indicating that the complex has the capacity to repress transcription of target genes. Indeed, in elB or noc mutants, expanded expression of tracheal branch-specific genes was observed. PMID- 12117810 TI - Mixl1 is required for axial mesendoderm morphogenesis and patterning in the murine embryo. AB - In Xenopus, the Mix/Bix family of homeobox genes has been implicated in mesendoderm development. Mixl1 is the only known murine member of this family. To examine the role of Mixl1 in murine embryogenesis, we used gene targeting to create mice bearing a null mutation of Mixl1. Homozygous Mixl1 mutant embryos can be distinguished from their littermates by a marked thickening of the primitive streak. By the early somite stage, embryonic development is arrested, with the formation of abnormal head folds, foreshortened body axis, absence of heart tube and gut, deficient paraxial mesoderm, and an enlarged midline tissue mass that replaces the notochord. Development of extra-embryonic structures is generally normal except that the allantois is often disproportionately large for the size of the mutant embryo. In chimeras, Mixl1(-/-) mutant cells can contribute to all embryonic structures, with the exception of the hindgut, suggesting that Mixl1 activity is most crucial for endodermal differentiation. Mixl1 is therefore required for the morphogenesis of axial mesoderm, the heart and the gut during embryogenesis. PMID- 12117811 TI - Cell coherence during production of the presomitic mesoderm and somitogenesis in the mouse embryo. AB - In this study, we investigated (in the early mouse embryo) the clonal properties of precursor cells which contribute to the segmented myotome, a structure derived from the somites. We used the laacZ method of single cell-labelling to visualise clones born before segmentation and bilateralisation. We found that clones which contribute to several segments both unilateral and bilateral were regionalised along the mediolateral axis and that their mediolateral position was maintained in successive adjacent segments. Furthermore, clones contributed to all segments, from their most anterior to their most posterior borders. Therefore, it appears that mediolateral regionalisation of myotomal precursor cells is a property established before bilateralisation of the presomitic mesoderm and that coherent clonal growth accompanies cell dispersion along both the mediolateral and anteroposterior axes. These findings in the mouse correlate well with what is known in the chick, suggesting conservation of the mode of production and distribution of the cells of the presomitic mesoderm. However, in addition, we also found that the mediolateral contribution of a clone is already determined in the pool of self-renewing cells that produces the myotomal precursor cells and thus that this pool is itself regionalised. Finally, we found that bilateral clones exhibit symmetry in right and left sides in the embryo at all levels of the mediolateral axis of the myotome. All these properties indicate synchrony and symmetry of formation of the presomitic mesoderm on both sides of the embryo leading to formation of a static embryonic structure with few cell movements. We suggest that sequential production of groups of cells with an identical clonal origin for both sides of the embryo from a single pool of self-renewing cells, coupled with acquisition of static cell behaviour, could play a role in colinearity of expression of Hox genes and in the segmentation system of higher vertebrates. PMID- 12117812 TI - Ephrin-B ligands play a dual role in the control of neural crest cell migration. AB - Little is known about the mechanisms that direct neural crest cells to the appropriate migratory pathways. Our aim was to determine how neural crest cells that are specified as neurons and glial cells only migrate ventrally and are prevented from migrating dorsolaterally into the skin, whereas neural crest cells specified as melanoblasts are directed into the dorsolateral pathway. Eph receptors and their ephrin ligands have been shown to be essential for migration of many cell types during embryonic development. Consequently, we asked if ephrin B proteins participate in the guidance of melanoblasts along the dorsolateral pathway, and prevent early migratory neural crest cells from invading the dorsolateral pathway. Using Fc fusion proteins, we detected the expression of ephrin-B ligands in the dorsolateral pathway at the stage when neural crest cells are migrating ventrally. Furthermore, we show that ephrins block dorsolateral migration of early-migrating neural crest cells because when we disrupt the Eph ephrin interactions by addition of soluble ephrin-B ligand to trunk explants, early neural crest cells migrate inappropriately into the dorsolateral pathway. Surprisingly, we discovered the ephrin-B ligands continue to be expressed along the dorsolateral pathway during melanoblast migration. RT-PCR analysis, in situ hybridisation, and cell surface-labelling of neural crest cell cultures demonstrate that melanoblasts express several EphB receptors. In adhesion assays, engagement of ephrin-B ligands to EphB receptors increases melanoblast attachment to fibronectin. Cell migration assays demonstrate that ephrin-B ligands stimulate the migration of melanoblasts. Furthermore, when Eph signalling is disrupted in vivo, melanoblasts are prevented from migrating dorsolaterally, suggesting ephrin B ligands promote the dorsolateral migration of melanoblasts. Thus, transmembrane ephrins act as bifunctional guidance cues: they first repel early migratory neural crest cells from the dorsolateral path, and then later stimulate the migration of melanoblasts into this pathway. The mechanisms by which ephrins regulate repulsion or attraction in neural crest cells are unknown. One possibility is that the cellular response involves signalling to the actin cytoskeleton, potentially involving the activation of Cdc42/Rac family of GTPases. In support of this hypothesis, we show that adhesion of early migratory cells to an ephrin-B-derivatized substratum results in cell rounding and disruption of the actin cytoskeleton, whereas plating of melanoblasts on an ephrin-B substratum induces the formation of microspikes filled with F-actin. PMID- 12117813 TI - Morphological boundary forms by a novel inductive event mediated by Lunatic fringe and Notch during somitic segmentation. AB - Boundary formation plays a central role in differentiating the flanking regions that give rise to discrete tissues and organs during early development. We have studied mechanisms by which a morphological boundary and tissue separation are regulated by examining chicken somite segmentation as a model system. By transplanting a small group of cells taken from a presumptive border into a non segmentation site, we have found a novel inductive event where posteriorly juxtaposed cells to the next-forming border instruct the anterior cells to become separated and epithelialized. We have further studied the molecular mechanisms underlying these interactions by focusing on Lunatic fringe, a modulator of Notch signaling, which is expressed in the region of the presumptive boundary. By combining DNA in ovo electroporation and embryonic transplantation techniques we have ectopically made a sharp boundary of Lunatic fringe activity in the unsegmented paraxial mesoderm and observed a fissure formed at the interface. In addition, a constitutive active form of Notch mimics this instructive phenomenon. These suggest that the boundary-forming signals emanating from the posterior border cells are mediated by Notch, the action of which is confined to the border region by Lunatic fringe within the area where mRNAs of Notch and its ligand are broadly expressed in the presomitic mesoderm. PMID- 12117814 TI - Drumstick is a zinc finger protein that antagonizes Lines to control patterning and morphogenesis of the Drosophila hindgut. AB - Elongation of the Drosophila embryonic hindgut epithelium occurs by a process of oriented cell rearrangement requiring the genes drumstick (drm) and lines (lin). The elongating hindgut becomes subdivided into domains -- small intestine, large intestine and rectum -- each characterized by a specific pattern of gene expression dependent upon normal drm and lin function. We show that drm encodes an 81 amino acid (10 kDa) zinc finger protein that is a member of the Odd-skipped family. drm expression is localized to the developing midgut-hindgut junction and is required to establish the small intestine, while lin is broadly expressed throughout the gut primordium and represses small intestine fate. lin is epistatic to drm, suggesting a model in which localized expression of drm blocks lin activity, thereby allowing small intestine fate to be established. Further supporting this model, ectopic expression of Drm throughout the hindgut produces a lin phenotype. Biochemical and genetic data indicate that the first conserved zinc finger of Drm is essential for its function. We have thus defined a pathway in which a spatially localized zinc finger protein antagonizes a globally expressed protein, thereby leading to specification of a domain (the small intestine) necessary for oriented cell rearrangement. PMID- 12117815 TI - The different components of a multisubunit cell number-counting factor have both unique and overlapping functions. AB - Dictyostelium aggregation streams break up into groups of 10(3) to 2 x 10(4) cells. The cells sense the number of cells in a stream or group by the level of a secreted counting factor (CF). CF is a complex of at least 5 polypeptides. When the gene encoding countin (one of the CF polypeptides) was disrupted, the cells could not sense each other's presence, resulting in non-breaking streams that coalesced into abnormally large groups. To understand the function of the components of CF, we have isolated cDNA sequences encoding a second component of CF, CF50. CF50 is 30% identical to lysozyme (but has very little lysozyme activity) and contains distinctive serine-glycine motifs. Transformants with a disrupted cf50 gene, like countin(-) cells, form abnormally large groups. Addition of recombinant CF50 protein to developing cf50(-) cells rescues their phenotype by decreasing group size. Abnormalities seen in aggregating countin(-) cells (such as high cell-cell adhesion and low motility) are also observed in the cf50(-) cells. Western blot analysis of conditioned medium sieve column fractions showed that the CF50 protein is present in the same fraction as the 450 kDa CF complex. In the absence of CF50, secreted countin is degraded, suggesting that one function of CF50 may be to protect countin from degradation. However, unlike countin(-) cells, cf50(-) cells differentiate into an abnormally high percentage of cells expressing SP70 (a marker expressed in a subset of prespore cells), and this difference can be rescued by exposing cells to recombinant CF50. These observations indicate that unlike other known multisubunit factors, CF contains subunits with both overlapping and unique properties. PMID- 12117816 TI - Translational repression by MSY4 inhibits spermatid differentiation in mice. AB - In developing male germ cells, newly synthesized protamine mRNAs are stored for up to 7 days before translational activation. Translational repression of protamine 1 (Prm1) mRNA requires sequences present in its 3' untranslated region (UTR) and substantial evidence suggests a role for the murine Y-box protein MSY4 in this process. To determine if MSY4 can mediate translational repression in vivo, we generated transgenic mice in which the temporal window of MSY4 expression was extended during spermatogenesis. Expression of MSY4 disrupted the normal completion of spermatogenesis and caused dominant sterility. Immunocytochemical analysis of several markers, including the protamines, indicated that MSY4 prevented normal activation of translation. mRNAs whose translation was inhibited contained at least one MSY4 RNA recognition site, suggesting sequence-dependent translational repression. Altered translational activation resulted in defective processing of protamine 2 and severe defects in sperm morphogenesis. These results suggest that MSY4 plays an active role in translational repression of several mRNAs in differentiating spermatids. PMID- 12117817 TI - Expression and function of an even-skipped homolog in the leech Helobdella robusta. AB - We have identified homologs of the Drosophila pair-rule gene even-skipped in the glossiphoniid leeches Helobdella robusta and Theromyzon trizonare. In leech embryos, segments arise sequentially from five pairs of embryonic stem cells (teloblasts) that undergo iterated divisions to generate columns (bandlets) of segmental founder cells (primary blast cells), which in turn generate segmentally iterated sets of definitive progeny. In situ hybridization revealed that Hro-eve is expressed in the teloblasts and primary blast cells, and that these transcripts appear to be associated with mitotic chromatin. In more advanced embryos, Hro-eve is expressed in segmentally iterated sets of cells in the ventral nerve cord. Lineage analysis revealed that neurons expressing Hro-eve arise from the N teloblast. To assess the function of Hro-eve, we examined embryos in which selected blastomeres had been injected with antisense Hro-eve morpholino oligonucleotide (AS-Hro-eve MO), concentrating on the primary neurogenic (N teloblast) lineage. Injection of AS-Hro-eve MO perturbed the normal patterns of teloblast and blast cell divisions and disrupted gangliogenesis. These results suggest that Hro-eve is important in regulating early cell divisions through early segmentation, and that it also plays a role in neuronal differentiation. PMID- 12117818 TI - Two linked hairy/Enhancer of split-related zebrafish genes, her1 and her7, function together to refine alternating somite boundaries. AB - The formation of somites, reiterated structures that will give rise to vertebrae and muscles, is thought to be dependent upon a molecular oscillator that may involve the Notch pathway. hairy/Enhancer of split related [E(spl)]-related (her or hes) genes, potential targets of Notch signaling, have been implicated as an output of the molecular oscillator. We have isolated a zebrafish deficiency, b567, that deletes two linked her genes, her1 and her7. Homozygous b567 mutants have defective somites along the entire embryonic axis. Injection of a combination of her1 and her7 (her1+7) morpholino modified antisense oligonucleotides (MOs) phenocopies the b567 mutant somitic phenotype, indicating that her1 and her7 are necessary for normal somite formation and that defective somitogenesis in b567 mutant embryos is due to deletion of her1 and her7. Analysis at the cellular level indicates that somites in her1+7-deficient embryos are enlarged in the anterior-posterior dimension. Weak somite boundaries are often found within these enlarged somites which are delineated by stronger, but imperfect, boundaries. In addition, the anterior-posterior polarity of these enlarged somites is disorganized. Analysis of her1 MO-injected embryos and her7 MO-injected embryos indicates that although these genes have partially redundant functions in most of the trunk region, her1 is necessary for proper formation of the anteriormost somites and her7 is necessary for proper formation of somites posterior to somite 11. By following somite development over time, we demonstrate that her genes are necessary for the formation of alternating strong somite boundaries. Thus, even though two potential downstream components of Notch signaling are lacking in her1+7-deficient embryos, somite boundaries form, but do so with a one and a half to two segment periodicity. PMID- 12117820 TI - The transformer gene in Ceratitis capitata provides a genetic basis for selecting and remembering the sexual fate. AB - The medfly Ceratitis capitata contains a gene (Cctra) with structural and functional homology to the Drosophila melanogaster sex-determining gene transformer (tra). Similar to tra in Drosophila, Cctra is regulated by alternative splicing such that only females can encode a full-length protein. In contrast to Drosophila, however, where tra is a subordinate target of Sex-lethal (Sxl), Cctra seems to initiate an autoregulatory mechanism in XX embryos that provides continuous tra female-specific function and act as a cellular memory maintaining the female pathway. Indeed, a transient interference with Cctra expression in XX embryos by RNAi treatment can cause complete sexual transformation of both germline and soma in adult flies, resulting in a fertile male XX phenotype. The male pathway seems to result when Cctra autoregulation is prevented and instead splice variants with truncated open reading frames are produced. We propose that this repression is achieved by the Y-linked male determining factor (M). PMID- 12117819 TI - Oskar anchoring restricts pole plasm formation to the posterior of the Drosophila oocyte. AB - Localization of the maternal determinant Oskar at the posterior pole of Drosophila melanogaster oocyte provides the positional information for pole plasm formation. Spatial control of Oskar expression is achieved through the tight coupling of mRNA localization to translational control, such that only posterior localized oskar mRNA is translated, producing the two Oskar isoforms Long Osk and Short Osk. We present evidence that this coupling is not sufficient to restrict Oskar to the posterior pole of the oocyte. We show that Long Osk anchors both oskar mRNA and Short Osk, the isoform active in pole plasm assembly, at the posterior pole. In the absence of anchoring by Long Osk, Short Osk disperses into the bulk cytoplasm during late oogenesis, impairing pole cell formation in the embryo. In addition, the pool of untethered Short Osk causes anteroposterior patterning defects, owing to the dispersion of pole plasm and its abdomen inducing activity throughout the oocyte. We show that the N-terminal extension of Long Osk is necessary but not sufficient for posterior anchoring, arguing for multiple docking elements in Oskar. This study reveals cortical anchoring of the posterior determinant Oskar as a crucial step in pole plasm assembly and restriction, required for proper development of Drosophila melanogaster. PMID- 12117821 TI - Bmp signaling is required for development of primary lens fiber cells. AB - We have investigated the role of Bmp signaling in development of the mouse lens using three experimental strategies. First, we have shown that the Bmp ligand inhibitor noggin can suppress the differentiation of primary lens fiber cells in explant culture. Second, we have expressed a dominant-negative form of the type 1 Bmp family receptor Alk6 (Bmpr1b -- Mouse Genome Informatics) in the lens in transgenic mice and shown that an inhibition of primary fiber cell differentiation can be detected at E13.5. Interestingly, the observed inhibition of primary fiber cell development was asymmetrical and appeared only on the nasal side of the lens in the ventral half. Expression of the inhibitory form of Alk6 was driven either by the alpha A-cystallin promoter or the ectoderm enhancer from the Pax6 gene in two different transgenes. These expression units drive transgene expression in distinct patterns that overlap in the equatorial cells of the lens vesicle at E12.5. Despite the distinctions between the transgenes, they caused primary fiber cell differentiation defects that were essentially identical, which implied that the equatorial lens vesicle cells were responding to Bmp signals in permitting primary fiber cells to develop. Importantly, E12.5 equatorial lens vesicle cells showed cell-surface immunoreactivity for bone-morphogenetic protein receptor type 2 and nuclear immunoreactivity for the active, phosphorylated form of the Bmp responsive Smads. This indicated that these cells had the machinery for Bmp signaling and were responding to Bmp signals. We conclude that Bmp signaling is required for primary lens fiber cell differentiation and, given the asymmetry of the differentiation inhibition, that distinct differentiation stimuli may be active in different quadrants of the eye. PMID- 12117823 TI - Pax2.1 is required for the development of thyroid follicles in zebrafish. AB - The thyroid gland is an organ primarily composed of endoderm-derived follicular cells. Although disturbed embryonic development of the thyroid gland leads to congenital hypothyroidism in humans and mammals, the underlying principles of thyroid organogenesis are largely unknown. In this study, we introduce zebrafish as a model to investigate the molecular and genetic mechanisms that control thyroid development. Marker gene expression suggests that the molecular pathways of early thyroid development are essentially conserved between fish and mammals. However during larval stages, we find both conserved and divergent features of development compared with mammals. A major difference is that in fish, we find evidence for hormone production not only in thyroid follicular cells, but also in an anterior non-follicular group of cells. We show that pax2.1 and pax8, members of the zebrafish pax2/5/8 paralogue group, are expressed in the thyroid primordium. Whereas in mice, only Pax8 has a function during thyroid development, analysis of the zebrafish pax2.1 mutant no isthmus (noi(-/-)) demonstrates that pax2.1 has a role comparable with mouse Pax8 in differentiation of the thyroid follicular cells. Early steps of thyroid development are normal in noi(-/-), but later expression of molecular markers is lost and the formation of follicles fails. Interestingly, the anterior non-follicular site of thyroid hormone production is not affected in noi(-/-). Thus, in zebrafish, some remaining thyroid hormone synthesis takes place independent of the pathway leading to thyroid follicle formation. We suggest that the noi(-/-) mutant serves as a new zebrafish model for hypothyroidism. PMID- 12117822 TI - Diverse dependencies of developing Merkel innervation on the trkA and both full length and truncated isoforms of trkC. AB - This study demonstrates that innervation dependent on two different neurotrophin tyrosine kinase (trk) receptors can form the same types of sensory endings (Merkel endings) in the same target (Merkel cells of vibrissa follicles). Some endings transiently express trkA during their initial development, whereas others express trkC throughout their development. Consequently, elimination of kinase domains of either trkA or trkC each result in a partial loss of Merkel endings, whereas absence of kinase domains of both receptors results in a total loss. At the onset of Merkel ending development, at least one kinase-lacking trkC isoform is transiently expressed on all the follicle cells, while neurotrophin 3 is transiently expressed only in the cells at the middle third of the follicle where the Merkel endings and cells develop. This transient non-neuronal expression of truncated trkC is essential for development of any Merkel endings, whereas some Merkel endings and cells still begin to develop in the absence of neurotrophin 3. Therefore, truncated trkC plays a more important role in the development of this innervation than kinase forms of trkA or trkC or of NT3, the only known ligand for trkC receptors. PMID- 12117824 TI - Survivors of coarctation repair: fixed but not cured. AB - While often considered to be cured, patients with repaired coarctation of the aorta frequently have premature morbidity and even mortality. PMID- 12117825 TI - The erythrocyte: a new player in atheromatous core formation. AB - The membrane of the red blood cell hides constituents that are lipid rich and can bind to macrophage scavenger receptors, thus posing the challenging hypothesis that erythrocytes contribute to atheroma formation in coronary arteries. PMID- 12117826 TI - Physician administered sedation for DC cardioversion. AB - Providing anaesthetic cover for DC cardioversion can sometimes prove a challenge for the cardiologist, with potentially disastrous consequences for the patient PMID- 12117827 TI - Stamps in cardiology: Quinine. PMID- 12117828 TI - Trends in mortality from cardiovascular and cerebrovascular diseases in Europe and other areas of the world. AB - OBJECTIVE: To analyse trends in mortality from coronary heart disease (CHD) and cerebrovascular disease (CVD) over the period 1965 to 1998 in the European Union, other European countries, the USA, and Japan. METHODS AND RESULTS: Data were derived from the World Health Organization database. In the European Union, CHD mortality in men rose from 146/100 000 in 1965-9 to 163/100 000 in 1975-9 and declined thereafter to 99/100 000 in 1995-8 (-39%). In women, the fall was from 70 to 45/100 000 (-36%). A > 55% decline in CVD was registered in both sexes. In eastern Europe, mortality from both CHD and CVD rose up to the early 1990s but has declined over the past few years in Poland and the Czech Republic. In the Russian Federation during 1995-8, mortality rates from CHD reached 330/100 000 men and 154/100 000 women and mortality rates from CVD were 203/100 000 men and 150/100 000 women-that is, they were among the highest rates worldwide. In the USA and Japan, long term trends were favourable for both CHD and CVD. CONCLUSIONS: Trends in mortality from CHD and CVD were favourable in several developed areas of the world, but there were major geographical differences. In a few eastern European countries, mortality from CHD and CVD remains exceedingly high. PMID- 12117829 TI - Prevalence of myocardial viability assessed by single photon emission computed tomography in patients with chronic ischaemic left ventricular dysfunction. AB - OBJECTIVE: To assess the prevalence of myocardial viability by technetium-99m (Tc 99m)-tetrofosmin/fluorine-18-fluorodeoxyglucose (FDG) single photon emission computed tomography (SPECT) in patients with ischaemic cardiomyopathy. DESIGN: A retrospective observational study. SETTING: Thoraxcenter Rotterdam (a tertiary referral centre). PATIENTS: 104 patients with chronic coronary artery disease and severely depressed left ventricular function presenting with heart failure symptoms. MAIN OUTCOME MEASURES: Prevalence of myocardial viability as evaluated by Tc-99m-tetrofosmin/FDG SPECT imaging. Two strategies for assessing viability in dysfunctional myocardium were used: perfusion imaging alone, and the combination of perfusion and metabolic imaging. RESULTS: On perfusion imaging alone, 56 patients (54%) had a significant amount of viable myocardium, whereas 48 patients (46%) did not. Among the 48 patients with no significant viability by perfusion imaging alone, seven additional patients (15%) had significantly viable myocardium on combined perfusion and metabolic imaging. Thus with a combination of perfusion and metabolic imaging, 63 patients (61%) had viable myocardium and 41 (39%) did not. CONCLUSIONS: On the basis of the presence of viable dysfunctional myocardium, 61% of patients with chronic coronary artery disease and depressed left ventricular ejection fraction presenting with heart failure symptoms may be considered for coronary revascularisation. The combination of perfusion and metabolic imaging identified more patients with significant viability than myocardial perfusion imaging alone. PMID- 12117830 TI - Images in cardiology: Spontaneous right coronary artery dissection. PMID- 12117833 TI - Images in cardiology: Imaginary aortic leaflets. PMID- 12117832 TI - Images in cardiology: Cyanosis, cor triatriatum, and primum atrial septal defect in an adult. PMID- 12117831 TI - Relation between early mitral regurgitation and left ventricular thrombus formation after acute myocardial infarction: results of the GISSI-3 echo substudy. AB - OBJECTIVE: To evaluate the prevalence and correlates of left ventricular thrombosis in patients with acute myocardial infarction, and whether the occurrence of early mitral regurgitation has a protective effect against the formation of left ventricular thrombus. DESIGN AND SETTING: Multicentre clinical trial carried out in 47 Italian coronary care units. PATIENTS AND METHODS: 757 patients from the GISSI-3 echo substudy population with their first acute myocardial infarct were studied by echocardiography at 24-48 hours from symptom onset (S1), at discharge (S2), at six weeks (S3), and at six months (S4). The diagnosis of left ventricular thrombosis was based on the detection of an echo dense mass with defined margins visible throughout the cardiac cycle in at least two orthogonal views. RESULTS: In 64 patients (8%), left ventricular thrombosis was detected in one or more examinations. Compared with the remaining 693 patients, subjects with left ventricular thrombosis were older (mean (SD) age: 64.6 (13.0) v 59.8 (11.7) years, p < 0.005), and had larger infarcts (extent of wall motion asynergy: 40.9 (11.5)% v 24.9 (14)%, p < 0.001), greater depression of left ventricular ejection fraction at S1 (43.3 (6.9)% v 48.1 (6.8)%, p < 0.001), and greater left ventricular volumes at S1 (end diastolic volume: 87 (22) v 78 (18) ml/m(2), p < 0.001; end systolic volume: 50 (17) v 41 (14) ml/m(2), p < 0.001). The prevalence of moderate to severe mitral regurgitation on colour Doppler at S1 was greater in patients who had left ventricular thrombosis at any time (10.2% v 4.2%, p < 0.05). On stepwise multiple logistic regression analysis the only independent variables related to the presence of left ventricular thrombosis were the extent of wall motion asynergy and anterior site of infarction. CONCLUSIONS: Left ventricular thrombosis is not reduced, and may even be increased, by early moderate to severe mitral regurgitation after acute myocardial infarction. The only independent determinant of left ventricular thrombosis is the extent of the akinetic-dyskinetic area detected on echocardiography between 24-48 hours from symptom onset. PMID- 12117835 TI - Images in cardiology: A rare congenital coronary anomaly: anomalous origin of the right coronary artery from the pulmonary artery. PMID- 12117836 TI - Images in cardiology: Effect of coronary angioplasty on "stunned" myocardium. PMID- 12117834 TI - Can coronary flow velocity reserve determined by transthoracic Doppler echocardiography predict the recovery of regional left ventricular function in patients with acute myocardial infarction? AB - OBJECTIVE: To determine whether the early assessment of coronary flow velocity reserve (CFVR) by transthoracic Doppler echocardiography (TTDE) can predict myocardial viability after revascularisation in patients with acute myocardial infarction. METHODS: 29 patients with anterior acute myocardial infarction who were successfully treated by coronary angioplasty were studied. TTDE was used to record coronary flow velocities in the distal left anterior descending artery at rest and during hyperaemia induced by intravenous infusion of adenosine triphosphate. CFVR was calculated immediately and 24 hours after revascularisation and at discharge. Regional wall motion was analysed to calculate the anterior wall motion score index (A-WMSI) by two dimensional echocardiography before revascularisation and at discharge. RESULTS: CFVR immediately and 24 hours after revascularisation correlated significantly with A WMSI at discharge (r = -0.58, p < 0.001 and r = -0.80, p < 0.0001, respectively). CFVR 24 hours after revascularisation was a better predictor of recovery of regional left ventricular function than CFVR immediately after revascularisation. The optimal cut off ratio for predicting viable myocardium was 1.5 for CFVR 24 hours after revascularisation (sensitivity = 94%, specificity = 91%). CONCLUSIONS: CFVR by TTDE was useful for predicting the recovery of left ventricular function after revascularisation in patients with acute myocardial infarction PMID- 12117837 TI - Images in cardiology: Common atrium in a child with Ellis-Van Creveld syndrome. PMID- 12117838 TI - Quantification of glyceryl trinitrate effect through analysis of the synthesised ascending aortic pressure waveform. AB - OBJECTIVE: To establish through analysis of the radial pressure pulse waveform the dose dependent effects of glyceryl trinitrate (GTN) on properties of different blood vessels. DESIGN: Radial pulse waveform was measured in randomised order before, during a five hour application of a GTN patch delivering 0.104 0.625 mg/h, and for two hours after patch removal. The radial pressure waveform (Millar applanation tonometer) was convolved into an ascending aortic wave using a generalised transfer function (SphygmoCor process) enabling measurement of aortic systolic, diastolic, pulse, mean, and augmented pressure and left ventricular ejection duration in addition to standard brachial cuff pressures. SETTING: Fu Wai and Ren Ming hospitals in Beijing, China. PATIENTS: 46 recumbent hospitalised patients aged 56 (9) years, awaiting electrophysiological or other diagnostic studies, fasting, and with other treatments suspended. MAJOR OUTCOME MEASURES: Conventional brachial pressure measures and data from the synthesised aortic pulse. RESULTS: There was no consistent change in heart rate or brachial pressures except for a decrease in systolic and pulse pressures (p < 0.01) at dose > 0.416 mg/h. In contrast, there were substantial and significant (p < 0.0001) decreases in aortic systolic, pulse, and augmented pressures at all doses, mean pressure (p < 0.001) at doses > 0.416 mg/h, and ejection duration (p < 0.001) at doses > 0.208 mg/h. CONCLUSIONS: Pulse waveform analysis exposes dose dependent effects of GTN on the aortic waveform, suggesting muscular conduit arterial dilatation with reduced wave reflection at the lowest dose, arteriolar dilatation and decreased peripheral resistance at the highest dose, and venous dilatation at the intermediate dose. PMID- 12117839 TI - Constrictive pericarditis caused by infiltration of non-Hodgkin's lymphoma. PMID- 12117840 TI - Physiological changes in ventricular filling alter cardiac electrophysiology in patients with abnormal ventricular function. AB - OBJECTIVE: To explore the hypothesis that patients with abnormal ventricular function have an altered electrophysiological response to physiological changes in ventricular filling which is not evident in people with normal ventricles. DESIGN: The influence of an acute alteration in ventricular filling on dispersion of repolarisation, measured as QT dispersion, was examined in subjects with normal (n = 9) and abnormal ventricles (n = 9). A physiological reduction in ventricular filling was achieved using dual chamber atrioventricular (AV) pacing in two different modes-AV pacing: atrial activation 120 ms before ventricular activation such that atrial contraction occurred normally in late diastole; and VA (ventriculoatrial) pacing: atrial activation 50 ms after ventricular activation, such that atrial contraction occurred after closure of the AV valves. The absence of effective atrial contraction was confirmed by echocardiography. Ventricular cycle length and sequence of excitation through the ventricle was constant throughout both VA and AV sequences within each patient. RESULTS: During AV pacing (normal ventricular filling) there was no significant difference in QT dispersion between the two groups. In contrast during VA pacing, when the atrial component to ventricular filling was abolished, there was an immediate and consistent increase in QT dispersion compared with baseline in subjects with abnormal ventricular function (p < 0.001) but not in those with normal ventricles. CONCLUSIONS: An abrupt change in ventricular filling, within the physiological range, increased QT dispersion in subjects with abnormal ventricular function but not in subjects with normal ventricles. The findings suggest an altered electrophysiological response to ventricular load in patients with abnormal ventricular function. PMID- 12117842 TI - Relation between QT duration and maximal wall thickness in familial hypertrophic cardiomyopathy. AB - BACKGROUND: QT abnormalities have been reported in left ventricular hypertrophy and hypertrophic cardiomyopathy. OBJECTIVE: To determine the relation between left ventricular hypertrophy and increased QT interval in familial hypertrophic cardiomyopathy. METHODS: The QT interval was measured in 206 genotyped adult subjects with familial hypertrophic cardiomyopathy from 15 unrelated families carrying mutations in the beta myosin heavy chain (beta-MHC) gene (five families, n = 68) or the cardiac myosin binding protein C (MyBPC) gene (10 families, n = 138). Subjects were classified as genetically unaffected (controls, n = 112), affected with left ventricular hypertrophy (penetrants, n = 58), or affected without left ventricular hypertrophy (non-penetrants, n = 36). RESULTS: There was a significant increase in QTmax and QTmin from controls to non-penetrants and penetrants for both the MyBPC group (p < or = 0.001 and p < or = 0.001, respectively) and the beta-MHC group (p < or = 0.001 and p < or = 0.001, respectively). In the MyBPC group, the increase in the QT interval could be explained by increased left ventricular hypertrophy. In the beta-MHC group, non penetrants had a significantly longer QTmax than controls despite the absence of left ventricular hypertrophy, and a similar QT interval to penetrants despite a lesser degree of left ventricular hypertrophy. CONCLUSIONS: In familial hypertrophic cardiomyopathy, genetically affected subjects without left ventricular hypertrophy may have a prolonged QT duration, which depends not only on the degree of left ventricular hypertrophy, when present, but also on the causative mutation. PMID- 12117843 TI - Images in cardiology: Autocapture stimulus simulating atrioventricular sequential pacing. PMID- 12117844 TI - Images in cardiology: Angiographic evidence of aberrant right subclavian artery associated with common carotid trunk. PMID- 12117845 TI - Use of secondary preventive drugs in patients with acute coronary syndromes treated medically or with coronary angioplasty: results from the nationwide French PREVENIR survey. AB - BACKGROUND: There is evidence that several classes of drugs are beneficial for secondary prevention in patients with coronary artery disease. OBJECTIVE: To compare the use of secondary preventive drugs in patients with acute coronary syndromes given conservative treatment or percutaneous coronary interventions. DESIGN: The PREVENIR survey was designed to assess the management of patients with acute coronary syndromes admitted to hospital in France in January 1998. Drugs prescribed at hospital discharge were recorded retrospectively from the hospital records, and treatment at six months was assessed prospectively. SETTING: University hospitals, general hospitals, and private clinics throughout the country. RESULTS: Of 1394 patients participating in the survey, 668 underwent coronary angioplasty during the initial hospital stay and 706 had medical treatment only. At hospital discharge, aspirin, beta blockers, and statins were prescribed significantly more often in patients undergoing angioplasty. Using multivariate logistic regression, coronary angioplasty was an independent predictor of treatment with aspirin (odds ratio 3.55), statins (1.92), and beta blockers (1.41). Compared with treatment at discharge, only statin use differed at six months, with a significant increase both in patients treated medically and in those who had undergone angioplasty. Increased use of statins, aspirin, and beta blockers was significantly correlated with coronary angioplasty during the initial hospital stay. CONCLUSIONS: In this national French survey, patients treated with percutaneous coronary interventions were more likely to receive secondary preventive drugs than patients receiving medical treatment alone. PMID- 12117846 TI - Prevalence of hypertension in children after early repair of coarctation of the aorta: a cohort study using casual and 24 hour blood pressure measurement. AB - OBJECTIVE: To study the prevalence of hypertension in a cohort of patients using the current strategy of repair in early childhood. PATIENTS: The cohort of patients with coarctation of the aorta born between 1983 and 1992. INTERVENTION: Casual (mean of three resting readings) and 24 hour blood pressure were measured in 119 children and compared with data from 1034 normal controls. The arch repair and left ventricular parameters were assessed using Doppler echocardiography. RESULTS: Median ages at first intervention and at blood pressure measurement were 0.2 years (interquartile range 0.04-2.0) and 12.0 years (9.0-14.5), respectively. Doppler velocity in the descending aorta was significantly associated with blood pressure (r = 0.28, p = 0.002 for casual systolic blood pressure (SBP); r = 0.26, p = 0.005 for mean 24 hour SBP). Patients were classified as having "no" (n = 70) or "mild" (n = 49) arch obstruction. Casual SBP was > 95th centile in 28% (34 of 119) overall and in 21% (15 of 70) of the no arch obstruction group. Mean 24 hour SBP was > 95th centile in 30% (36 of 119) overall and in 19% (13 of 70) of the no obstruction group. The sensitivity and specificity of casual SBP in detecting increased 24 hour SBP were 66% and 88%, respectively. CONCLUSIONS: This unique study of a large cohort of patients treated for coarctation in early childhood showed that a disappointingly high prevalence of hypertension is already apparent in children aged 7-16 years in the absence of significant arch obstruction, whether assessed by 24 hour or by casual blood pressure measurement. PMID- 12117847 TI - Coil occlusion of systemic venous collaterals in hypoplastic left heart syndrome. AB - OBJECTIVE: To assess the frequency of systemic venous collaterals to the atria, which may cause desaturation, after stage II reconstructive surgery for hypoplastic left heart syndrome (HLHS) and to determine whether coil occlusion prevents the need for surgical ligation. DESIGN: Prospective interventional study. SETTING: Tertiary referral centre. PATIENTS: 27 children with HLHS undergoing cardiac catheterisation between October 1996 and February 2001. INTERVENTIONS: 19 children were catheterised prestage II, 1 poststage II, and 17 prestage III. Aortic oxygen saturation (SaAo) and pulmonary artery pressure (pPA) were recorded. Angiography was performed into the left internal jugular vein to look for venous collaterals. If present, they were occluded with Cook MReye coils. Angiography was repeated to confirm occlusion, and SaAo and pPA were remeasured. RESULTS: Collaterals were found in 7 of 27 children: 1 poststage II and 6 prestage III. These were occluded with 1-3 coils without complication. Mean (SE) SaAo before occlusion was 80.2 (2.1)% in those with collaterals compared with 88.7 (1.0)% in those without (p = 0.007). There was no difference in mean pPA between the two groups. After coil occlusion mean SaAo rose to 83.8 (1.8)% (p = 0.007) and mean pPA rose from 12.5 (1.5) to 14.5 (1.8) mm Hg (p = 0.02). None required surgical ligation. CONCLUSION: Angiography should be performed at catheterisation before stage II and III surgery for HLHS to exclude systemic venous collaterals. If present, they may be safely and effectively occluded with coils to improve saturation and prevent the need for subsequent surgical ligation. PMID- 12117849 TI - Images in cardiology: Computed tomography of the aortic valve. PMID- 12117848 TI - Preload-adjusted maximal power: a novel index of left ventricular contractility in atrial fibrillation. AB - BACKGROUND: Left ventricular contractility in atrial fibrillation is known to change in a beat to beat fashion, but there is no gold standard for contractility indices in atrial fibrillation, especially those measured non-invasively. OBJECTIVE: To determine whether the non-invasive index of contractility "preload adjusted PWR(max)" (maximal ventricular power divided by the square of end diastolic volume) can accurately measure left ventricular contractility in a beat to beat fashion in atrial fibrillation. METHODS: Atrial fibrillation was induced experimentally using 60 Hz stimulation of the atrium and maintained in 12 sheep; four received diltiazem, four digoxin, and four no drugs (control). Aortic flow, left ventricular volume, and left ventricular pressure were monitored simultaneously. Preload-adjusted PWR(max), the slope of the end systolic pressure volume relation (E(max)), and the maximum rate of change of left ventricular pressure (dP/dt(max)) were calculated in a beat to beat fashion. RESULTS: Preload adjusted PWR(max) correlated linearly with load independent E(max) (p < 0.0001) and curvilinearly with load dependent dP/dt(max) (p < 0.0001), which suggested the load independence of preload-adjusted PWR(max). After five minutes of diltiazem administration, preload-adjusted PWR(max), dP/dt(max), and E(max) fell significantly (p < 0.0001) to 62%, 64%, and 61% of baseline, respectively. Changes were not significant after five minutes of digoxin (103%, 98%, and 102%) or in controls (97%, 96%, and 95%). CONCLUSIONS: Preload-adjusted PWR(max) correlates linearly with E(max) and is a useful measure of contractility even in atrial fibrillation. Non-invasive application of this method, in combination with echocardiography and tonometry, may yield important information for optimising the treatment of patients with atrial fibrillation. PMID- 12117850 TI - Plaque composition in plexogenic and thromboembolic pulmonary hypertension: the critical role of thrombotic material in pultaceous core formation. AB - BACKGROUND: Patients with pulmonary hypertension develop intimal plaques in large pulmonary arteries. OBJECTIVE: To test the hypothesis that the composition of such plaques differs depending on whether the aetiology of the disease is thromboembolic or hypertensive. DESIGN: Chronic thromboembolic and plexogenic pulmonary hypertension (primary and secondary (Eisenmenger syndrome)) were investigated. These are spontaneous human models and were used to examine the independent role of thrombus and hypertension in plaque composition. SETTING: A national tertiary referral centre for lung transplantation and pulmonary thromboendoarterectomy. PATIENTS: Thirty nine patients with chronic thromboembolic pulmonary hypertension who had undergone thromboendoarterectomy (n = 32) or lung transplantation (n = 7), 28 with plexogenic diseases (nine primary and 19 Eisenmenger), and three with Eisenmenger syndrome complicated by thromboembolic events. INTERVENTIONS: The lung and thromboendoarterectomy samples were sectioned, stained with Movat pentachrome, and immunostained with antibodies for fibrin, platelets, inflammatory cells, smooth muscle cells, and erythrocyte membrane glycophorin A. MAIN OUTCOME MEASURE: Composition of the plaques affecting large pulmonary arteries. RESULTS: Two types of intimal lesion were distinguished in chronic thromboembolic pulmonary hypertension: fibrous plaques with angioneogenesis; and core-rich atherosclerotic plaques with pultaceous cores largely consisting of glycophorin immunoreactive material, with cholesterol clefts (61.5%), CD68 positive macrophages (84.6%), T lymphocytes (87%), and calcification (46.1%). The samples from the patients with Eisenmenger syndrome and thromboembolic complications had similar characteristics, whereas those from patients with uncomplicated primary pulmonary hypertension had core-free fibrous plaques, spotted with macrophages and T lymphocytes. CONCLUSIONS: Chronic thromboembolic pulmonary hypertension is associated with atherosclerotic plaques with glycophorin-rich pultaceous cores, and plexogenic pulmonary hypertension with fibrous plaques. Thromboembolic material thus plays a critical role in the formation of pultaceous cores, of which erythrocyte membrane derived glycophorin is a major component. PMID- 12117851 TI - Reversible left ventricular dysfunction simulating a myocardial infarction after pericardiectomy. AB - A 39 year old man with postoperative constrictive pericarditis after pericardiectomy developed major left ventricular systolic dysfunction with an anterior wall infarct pattern on ECG but no regional wall motion abnormalities by echocardiography or serum enzymatic evidence of a myocardial infarction. The left ventricular dysfunction resolved over two weeks with supportive treatment. It is postulated that this patient's transient left ventricular dysfunction and ECG changes were caused by myocardial inflammation and oedema induced by operative trauma during pericardiectomy. PMID- 12117852 TI - Effects of combined treatment with enalapril and losartan on myocardial function in heart failure. PMID- 12117853 TI - Images in cardiology: Preoperative treatment with phenoxybenzamine restores ECG to normal in a woman with pheochromocytoma. PMID- 12117854 TI - Dying from heart failure in hospital: palliative decision making analysis. PMID- 12117855 TI - Congenital junctional ectopic tachycardia in children and adolescents: a 20 year experience based study. PMID- 12117856 TI - Images in cardiology: Visualisation of activation and repolarisation in congenital long QT syndrome. PMID- 12117857 TI - Plasma concentrations of N-terminal atrial natriuretic peptide are raised in asymptomatic relatives of dilated cardiomyopathy patients with left ventricular enlargement. PMID- 12117858 TI - Images in cardiology: Narrowing of the thoraco-abdominal aorta. PMID- 12117859 TI - Arteriosclerotic renal artery stenosis: conservative versus interventional management. PMID- 12117861 TI - Effect of partial compliance on cardiovascular medication effectiveness. PMID- 12117860 TI - Heart failure in the young. PMID- 12117864 TI - Use of the transseptal puncture in transcatheter closure of long tunnel-type patent foramen ovale. AB - Two patients with long tunnel-type patent foramen ovale presented for elective transcatheter closure following transient ischaemic attack and stroke. Right to left shunting was confirmed on transthoracic and transoesophageal echocardiography. A new technique that used a transseptal procedure was devised to enable closure of the tunnel-type patent foramen ovale using the CardioSEAL transseptal occluder to avoid "bunching up" of the device and residual transatrial shunting. PMID- 12117865 TI - Endovascular stent repair for a dissecting thoracoabdominal aneurysm is feasible in the setting of a district general hospital: a multidisciplinary approach. AB - A patient presented to a district general hospital with a type B dissection of the aorta. He was deemed too unwell for surgical intervention. An endovascular stent repair was successfully carried out. The case shows that such a procedure can be safely performed by a multidisciplinary team within a district general hospital. PMID- 12117866 TI - TPMT in the treatment of Crohn's disease with azathioprine. AB - Azathioprine induced profound myelosuppression linked to TPMT deficiency has now been documented in many patient groups, including those with Crohn's disease. At the start of azathioprine or mercaptopurine therapy, measurement of TPMT activity has a role in identifying the 1 in 300 patients who are at risk of severe myelosuppression when treated with standard thiopurine dosages. During the initial months of azathioprine therapy a knowledge of TPMT status warns of early bone marrow toxicity. In patients established on azathioprine these is no clear evidence to suggest that TPMT is predictive of clinical response or drug toxicity, indicating a role for TPMT in the prediction of early events rather than long term control. In patients with Crohn's disease on long term azathioprine therapy, it is clear that myelosuppression, particularly leucopenia, is caused by other factors in addition to variable TPMT activity and therefore monitoring of blood cell counts throughout treatment is essential. PMID- 12117867 TI - Obesity, gender, and colon cancer. PMID- 12117868 TI - Extragastric MALT lymphoma. PMID- 12117869 TI - Where do IgA plasma cells in the gut come from? PMID- 12117870 TI - Protagonist: Crohn's disease recurrence can be prevented after ileal resection. PMID- 12117871 TI - Antagonist: Crohn's disease recurrence can be prevented after ileal resection. PMID- 12117872 TI - Defining the roles of perforin, Fas/FasL, and tumour necrosis factor alpha in T cell induced mucosal damage in the mouse intestine. AB - BACKGROUND AND AIMS: Mucosal flattening and epithelial cell apoptosis are typical features of T cell induced inflammatory diseases of the bowel, such as coeliac disease and graft versus host disease. Mice injected with a T cell activating anti-CD3 antibody develop a severe diarrhoeal illness. We describe the histological features of this enteropathy and define the effector mechanisms involved in T cell induced mucosal injury in this in vivo model. METHODS: Wild type and genetically modified mice were injected with the anti-CD3 antibody 3C11 (50 microg). Changes in the murine intestine were characterised by light microscopy analysis and terminal uridine nick-end labelling (TUNEL) assay. The role of perforin, Fas/Fas ligand (FasL), tumour necrosis factor alpha (TNF alpha), and interferon gamma (IFN-gamma) in T cell induced mucosal damage was assessed using selected immunodeficient mouse strains. RESULTS: T cell activation caused severe damage, including small intestinal mucosal flattening and apoptosis of crypt epithelial cells. Mucosal damage was unaltered in anti-CD3 treated mice lacking IFN-gamma, Fas, or TNF-alpha receptors. In mice lacking TNF-alpha receptors and Fas (TNF-R1xR2 lpr/lpr strain), enterocyte apoptosis was diminished but there was no significant reduction in tissue damage. Apoptosis and mucosal injury were significantly reduced in perforin knockout mice. Abrogation of both FasL and perforin (perforin KOxgld mice) further significantly reduced tissue damage and apoptotic bodies. CONCLUSIONS: T cell induced mucosal injury is mediated by the combined effect of multiple pathways but predominantly by perforin. The redundancy of the mechanisms of tissue damage will have significant impact on therapeutic strategies aimed at specific and targeted inhibition of inflammatory processes. PMID- 12117873 TI - Fat composition may be a clue to explain the primary therapeutic effect of enteral nutrition in Crohn's disease: results of a double blind randomised multicentre European trial. AB - BACKGROUND: Dietary fat has been suggested to determine the therapeutic effect of enteral diets in Crohn's disease. AIM: To assess the efficacy of two whole protein based diets with different fat compositions (n6 polyunsaturated fatty acids v monounsaturated fatty acids) in inducing clinical remission in active Crohn's disease compared with steroids. METHODS: Sixty two patients with active Crohn's disease were randomised to receive, for not more than 4 weeks: (a) a polymeric enteral diet containing 35 g of lipids per 1000 kcal, high in oleate (79%) and low in linoleate (6.5%) (PEN1), (b) an identical enteral diet except for the type of fat which was high in linoleate (45%) and low in oleate (28%) (PEN2), or (c) oral prednisone (1 mg/kg/day). Diets were double blindly administered. The steroid group received a conventional ward diet. Treatment failure was considered when remission was not achieved at week 4. Clinical activity and biological and nutritional parameters were monitored. Independent predictors of remission were identified by stepwise logistic regression analysis. RESULTS: Overall remission rates (by intention to treat) were 20% (4/20) for PEN1, 52% (12/23) for PEN2, and 79% (15/19) for steroids (overall p=0.001; p<0.0005 steroids v PEN1, and p=0.056 PEN2 v PEN1). After excluding those patients who were non-compliant during the first week (per protocol analysis), remission rates were 27%, 63%, and 79%, respectively (p=0.008, steroids and PEN2 v PEN1). After adjusting for confounding variables, PEN1 remained significantly associated with a poor response. CONCLUSION: The type of dietary fat may be of importance for the primary therapeutic effect of enteral nutrition in active Crohn's disease. PMID- 12117874 TI - Distribution and partial characterisation of IgG Fc binding protein in various mucin producing cells and body fluids. AB - BACKGROUND AND AIMS: Mucus released from goblet cells is important in intestinal mucosal defence, and mucin glycoproteins are thought to be major components of mucus. Recently, we identified and cloned another component of human colonic mucus, IgG Fc binding protein (Fc gamma BP). Fc gamma BP is immunologically distinct from known Fc gamma receptors and its structure contains repeated cysteine rich unit sequences resembling those present in mucins. In this work, we assessed the tissue distribution of Fc gamma BP, its binding activity in various body fluids, and its ability to inhibit complement mediated haemolysis. METHODS: Immunohistochemical localisation of Fc gamma BP, using monoclonal antibodies against Fc gamma BP (K9 or K17) and labelled IgG, was conducted in various mucin producing tissues: colon, small intestine, stomach, gall bladder, cystic duct, choledochus, bronchus, submandibular gland, conjunctiva, and cervix uteri. The binding activity of Fc gamma BP in mucus extracted from colon, gastric juice, bile, nasal discharges, saliva, sputum, and tears was also examined by immunodotblot and immunoprecipitation using these monoclonal antibodies. Inhibition of complement mediated haemolysis by Fc gamma BP was investigated using sheep red blood cells (SRBC) and anti-SRBC IgG. RESULTS: The immunohistochemical study revealed that mucin secreting cells in the colon, small intestine, gall bladder, cystic duct, choledochus, bronchus, submandibular gland, and cervix uteri contained Fc gamma BP, and immunodotblot and immunoprecipitation analysis using IgG and monoclonal antibodies demonstrated that the fluids secreted by these cells were capable of binding IgG. Mucin producing cells of the conjunctiva did not express Fc gamma BP molecules or bind to IgG. The surface mucus cells in the stomach were variably positive for Fc gamma BP. Perhaps because of proteolytic degradation, Fc gamma BP in gut lavage fluid did not have IgG binding activity, although this activity was present in the mucus covering the colon. Fc gamma BP suppressed complement mediated haemolysis of SRBC. CONCLUSIONS: Fc gamma BP is widely expressed on mucosal surfaces and in external secretions. It is functionally intact in several fluids. These findings lend support to the concept that Fc gamma BP is an important component of mucosal immunological defences. PMID- 12117875 TI - Anti-tissue transglutaminase antibodies from coeliac patients inhibit transglutaminase activity both in vitro and in situ. AB - BACKGROUND AND AIMS: Coeliac disease (CD) is a multifactorial disorder which has an autoimmune component characterised by the occurrence of disease specific autoreactive antibodies against the enzyme tissue transglutaminase (tTG). The aim of this study was to investigate whether binding of antibodies to the enzyme influences tTG activity. METHODS: tTG activity was assayed in the presence of immunoglobulin A (IgA) and immunoglobulin G (IgG) purified from the serum of coeliac patients, CUB 7402 (an anti-tTG mouse monoclonal antibody), and human anti-tTG monoclonal antibodies derived from both intestinal lymphocytes from three patients with CD and from peripheral blood lymphocytes from healthy subjects. For our studies we used calcium treated and untreated recombinant human tTG. Furthermore, the effects of antibodies were determined by immunohistochemical detection of tTG activity in sections of human umbilical cord. RESULTS: IgG and IgA from CD patients inhibited tTG activity in vitro in a dose dependent manner, with a different rate of inhibition among patients. The monoclonal antibody CUB 7402 and human monoclonal antibodies displayed a dose dependent inhibitory effect towards the catalytic activity of the enzyme, both in vitro and in situ. Preincubation of tTG with CaCl(2) caused loss of the inhibitory effect due to CUB 7402 but not that caused by human monoclonal antibodies. CONCLUSIONS: Purified CD IgA, IgG, as well as human anti-tTG monoclonal antibodies inhibited the enzymatic activity of human tTG both in vitro and in situ. PMID- 12117876 TI - New steroids for IBD: progress report. PMID- 12117878 TI - Obesity and colorectal cancer risk in women. AB - BACKGROUND: Several large studies of obesity and colorectal cancer risk have found no association among women but a reasonably consistent positive association among men. In women, a positive association that is stronger among, or limited to, those who are premenopausal has been suggested by studies that stratified analyses by age, although no previous study has examined the association by menopausal status. METHODS: We used proportional hazards analyses to estimate hazard ratios relating obesity to colorectal cancer risk among 89,835 women aged 40-59 years at recruitment into the Canadian National Breast Screening Study, a multicentre randomised controlled trial of mammography screening for breast cancer. During an average 10.6 years of follow up (936,433 person years), a total of 527 women were diagnosed with incident colorectal cancer (363 colon and 164 rectal). RESULTS: We found that obesity (body mass index > or = 30 kg/m(2)) was associated with an approximately twofold increased risk of colorectal cancer among women who were premenopausal at baseline (hazard ratio 1.88, 95% confidence interval 1.24-2.86). There was no association among postmenopausal women (p for interaction=0.01), and there was only a weak positive association in the entire cohort. CONCLUSIONS: Our data suggest that obesity is associated with a twofold increased risk of colorectal cancer in premenopausal women but is not associated with altered risk in postmenopausal women. Effect modification by menopausal status may better explain the inconsistent or weak findings in previous studies than the presumed lack of an association among women. PMID- 12117877 TI - Expression of CDX2 in normal and neoplastic human colon tissue and during differentiation of an in vitro model system. AB - BACKGROUND: The Cdx genes are expressed in the colorectal epithelium and are frequently downregulated during tumorigenesis. Overexpression of Cdx genes has been shown previously to result in cellular differentiation. AIM: To study expression of CDX2 in normal and neoplastic human colon using a newly isolated monoclonal antibody. To define expression of CDX1 and CDX2 in an in vitro model system of colorectal tumour progression and to ascertain whether these are subject to regulation during differentiation. METHODS: Normal and neoplastic human colon was immunostained for CDX2. CDX1 and CDX2 expression was assayed in cell lines derived from premalignant colonic adenomas by western blotting. Differentiation was induced by sodium butyrate treatment or post confluent growth, and changes in CDX expression compared with carcinoma cell lines with low levels of CDX expression. RESULTS: CDX2 protein displayed no gradient of expression within the colonic crypt. Cell lines derived from adenomas, with high levels of CDX1 and CDX2, showed no regulation of these proteins when induced to differentiate by butyrate or confluency. CDX expression in these cell lines was independent of their APC or Ras status. CDX1 and CDX2 were expressed at very low levels in some carcinoma cell lines and were modestly upregulated on differentiation but were not restored to levels seen in adenoma cells. CONCLUSION: The lack of significant regulation on cellular differentiation and the absence of a detectable gradient in the crypt implies that CDX2 may confer tissue specificity but may not play the previously suggested role in crypt patterning. PMID- 12117879 TI - Effect of folate supplementation on mucosal cell proliferation in high risk patients for colon cancer. AB - AIMS: Intracellular folate deficiency has been implicated in colonic carcinogenesis in epidemiological studies and animal and human cancer models. Our aim was to determine the effect of folate supplementation on patients with recurrent adenomatous polyps using rectal mucosal cell proliferation as a biomarker. PATIENTS AND METHODS: Eleven patients with recurrent adenomatous polyps of the colon were randomised into a treatment group (n=6) receiving a dietary supplement of 2 mg folic acid per day for three months and a control group (n=5) receiving a placebo. Rectal biopsies where taken at 10 cm from the anal verge prior to supplementation and repeated at four, 12, and 18 weeks from the start of the supplementation. Each biopsy was immediately incubated in culture medium enriched with bromodeoxyuridine (BrdU). The S phase cells which incorporated BrdU into their DNA were identified following immunohistochemical staining. Twenty five orientated crypts were identified for each time point and the number and position of BrdU positive and BrdU negative cells were counted. BrdU labelling indices (LIs) were calculated for the entire crypt and for each of five equal compartments running consequently from the base to the luminal surface. RESULTS: The LI of the treatment group (9.1 (6.7, 12.3)) and the control group (9.3 (7.8, 10.3)) were comparable at the start. Over the duration of the supplementation period, LI in the control group did not alter significantly (9.3 (7.8, 10.3) v 9.6 (8.9, 10.4)). However, LI of the folate treated group was lowered after 12 weeks of supplementation (9.1 (6.7, 12.3) v 7.4 (5.3, 9.6)). Analysis of the LI for compartments within the crypt showed that the most significant drop in number of proliferating cells was in the upper most regions of the crypt. CONCLUSION: These data indicate that (a) folate supplementation decreases colonic mucosal cell proliferation in a high risk group for colon cancer and (b) the most significant reduction takes place at the luminal aspect of the crypt. PMID- 12117880 TI - Molecular characteristics of serrated adenomas of the colorectum. AB - BACKGROUND: Serrated adenomas (SAs) of the colorectum combine architectural features of hyperplastic polyps and cytological features of classical adenomas. Molecular studies comparing SAs and classical adenomas suggest that each may be a distinct entity; in particular, it has been proposed that microsatellite instability (MSI) distinguishes SAs from classical adenomas and that SAs and the colorectal cancers arising from them develop along a pathway driven by low level microsatellite instability (MSI-L). AIMS: To define the molecular characteristics of SAs of the colorectum. MATERIALS AND METHODS: We analysed 39 SAs from 27 patients, including eight SAs from patients with familial adenomatous polyposis (FAP). We screened these polyps for selected molecular changes, including loss of heterozygosity (LOH) close to APC (5q21) and CRAC1 (15q13-q22), MSI, and mutations of K-ras, APC, p53, and beta-catenin. Expression patterns of beta catenin, p53, MLH1, MSH2, E-cadherin, and O(6)-methylguanine DNA methyltransferase (MGMT) were assessed by immunohistochemistry. Comparative genomic hybridisation was performed on several polyps. RESULTS: MSI was rare (<5% cases) and there was no loss of expression of mismatch repair proteins. Wnt pathway abnormalities (APC mutation/LOH, beta-catenin mutation/nuclear expression) occurred in 11 SAs, including 6/31 (19%) non-FAP tumours. CRAC1 LOH occurred in 23% of tumours. K-ras mutations and p53 mutations/overexpression were found in 15% and 8% of SAs, respectively. Loss of MGMT expression occurred in 18% of polyps and showed a borderline association with K-ras mutations. Aberrant E cadherin expression was found in seven polyps. Comparative genomic hybridisation detected no gains or deletions of chromosomal material. CONCLUSIONS: The serrated pathway of colorectal tumorigenesis appears to be heterogeneous. In common with classical adenomas, some SAs develop along pathways involving changes in APC/beta catenin. SAs rarely show MSI or any evidence of chromosomal-scale genetic instability. K-ras mutations may however be less common in SAs than in classical adenomas. Some SAs may harbour changes in the CRAC1 gene. Changes in known genes do not account for the growth of the majority of SAs. PMID- 12117881 TI - Performance of multidetector computed tomography colonography compared with conventional colonoscopy. AB - BACKGROUND AND AIMS: This was a prospective blinded study to compare computed tomography (CT) colonography, performed with multidetector arrays CT scan (MDCT), with conventional colonoscopy for the detection of colorectal neoplasia. METHODS: Fifty patients were examined by MDCT after standard bowel preparation and rectal air insufflation in the supine and prone positions. Data sets were examined by one radiologist and one gastroenterologist blinded to the patient's history and colonoscopy results. Patients subsequently underwent colonoscopy on the same day, which served as the gold standard. RESULTS: Nine of 11 lesions >10 mm (82%), 5/15 lesions of 6-9 mm (33%), and 1/42 polyps <5 mm (3%) were detected by MDCT colonography. One false positive result for a structure larger than 10 mm was described. Nineteen of 21 patients who had no lesions during conventional colonoscopy were considered free of lesions by MDCT colonography, yielding a per patient specificity of 90%. CONCLUSION: MDCT colonography provides good data quality and has good sensitivity and specificity for the detection of colonic lesions of 10 mm or more. PMID- 12117882 TI - Role of nitric oxide in gastric motor and sensory functions in healthy subjects. AB - BACKGROUND AND AIMS: Impaired accommodation and hypersensitivity to distension of the proximal stomach are considered to be important factors in the pathogenesis of dyspeptic complaints. As fundus relaxing agents may be effective in the treatment of these symptoms, insight into the mediators involved in fundic accommodation and associated perceptual responses is important. Therefore, we studied the effect of nitric oxide (NO) synthase inhibition by N(G)-monomethyl-L arginine (L-NMMA) on fundic tone, postprandial sensations, and gastric perception in healthy volunteers. SUBJECTS AND METHODS: Eighteen healthy volunteers participated in a double blind, placebo controlled, randomised study. They underwent a gastric barostat study to evaluate the effect of L-NMMA on meal and distension induced sensations and on fundic relaxation in response to oral meal intake, intraduodenal lipid, and glucagon administration. RESULTS: Compared with placebo, L-NMMA decreased fundic volume after oral meal intake (438 (55) v 304 (67) ml; n=8; p<0.05) and during intraduodenal lipid infusion (384 (37) v 257 (43) ml; n=10; p<0.05) but not after glucagon injection (570 (62) v 540 (52) ml; n=4; p=0.4). In addition, basal fundic volume was significantly reduced by L NMMA. Scores for nausea and satiation were decreased by L-NMMA after oral meal intake but not during intraduodenal lipid infusion. Perception scores to gastric distension were not altered by L-NMMA. CONCLUSIONS: NO is involved in maintaining basal fundic tone and in meal induced fundic relaxation in humans, but not in visceral perception. PMID- 12117883 TI - Role of nitric oxide in the gastric accommodation reflex and in meal induced satiety in humans. AB - AIMS: In humans, impaired gastric accommodation is associated with early satiety and weight loss. In animals, accommodation involves activation of gastric nitrergic neurones. Our aim was to study involvement of nitric oxide in gastric accommodation and in meal induced satiety in humans. METHODS: The effect of N(G) monomethyl-L-arginine (L-NMMA) 4 mg/kg/h and 8 mg/kg/h on gastric compliance, on sensitivity to distension, and on gastric accommodation was studied with a barostat in double blind, randomised, placebo controlled studies. The effect of L NMMA 8 mg/kg/h on meal induced satiety was studied using a drinking test. RESULTS: L-NMMA had no significant effect on fasting compliance and sensitivity. Ingestion of a meal induced a relaxation of 274 (15) ml which was significantly smaller after L-NMMA 4 mg/kg/h (132 (45) ml; p=0.03) or L-NMMA 8 mg/kg/h (82 (72) ml; p=0.03). L-NMMA 8 mg/kg/h significantly decreased the amount of food ingested at maximum satiety from 1058 (67) to 892 (73) kcal (p<0.01). CONCLUSION: In humans, fasting gastric tone and sensitivity to distension are not influenced by nitric oxide synthase inhibition, but the gastric accommodation reflex involves activation of nitrergic neurones. Inhibition of nitric oxide synthase impairs accommodation and enhances meal induced satiety. PMID- 12117884 TI - Injectable silicone biomaterial for faecal incontinence due to internal anal sphincter dysfunction. AB - BACKGROUND: A weak or disrupted internal anal sphincter can cause passive faecal incontinence. Conservative measures may help some patients but there is no simple surgical solution for those who fail conservative treatment. A successful technique using trans-sphincteric injection of a bulking agent to augment the internal anal sphincter was developed in a previous pilot study. AIM: To determine the clinical results and underlying physiological effects of biomaterial injection. PATIENTS: Six patients (four males, median age 53 years (range 36-65)) with faecal incontinence to solid or liquid stool related to poor internal anal sphincter function, of varied aetiology, were recruited. METHODS: Silicone based biomaterial injections were performed, under local anaesthesia, with antibiotic cover. Three injections were placed circumferentially, trans sphincterically, entering away from the anal margin and injecting at or just above the dentate line. Anorectal physiological studies, endoanal ultrasound, a bowel symptom diary, a validated incontinence score, and quality of life questionnaires were completed before treatment and on completion of follow up. RESULTS: At a median follow up of 18 months (range 15-19), five of six patients had marked symptom improvement. Faecal incontinence scores improved from a median of 14/24 (range 11-20) before to 8/24 (6-15) after injection. Short form-36 quality of life physical and social function scores improved from a median of 26/100 (5-33) to 79/100 (25-100) and from 10/100 (5-37) to 100/100 (50-100), respectively. There was a corresponding physiological increase in maximum anal resting and squeeze pressures. Ultrasound showed the Bioplastique to be retained in the correct position in the improved patients without migration. There were no complications. CONCLUSION: Trans-sphincteric injection of silicone biomaterial can provide a marked improvement in faecal incontinence related to a weak or disrupted internal anal sphincter. This is associated with improved sphincter function and quality of life. PMID- 12117886 TI - Alcohol abuse and the risk of pancreatic cancer. AB - BACKGROUND: Although most epidemiological studies do not support a role for alcohol in the aetiology of pancreatic cancer, an increased risk among heavy drinkers cannot be excluded. METHODS: In a retrospective cohort based on the Swedish Inpatient Register, we analysed the risk of pancreatic cancer among patients admitted to hospital for alcoholism (n=178 688), alcoholic chronic pancreatitis (n=3500), non-alcoholic chronic pancreatitis (n=4952), alcoholic liver cirrhosis (n=13 553), or non-alcoholic liver cirrhosis (n=7057) from 1965 to 1994. Follow up through to 1995 was accomplished by linkage to nationwide registers. Standardised incidence ratios (SIRs) express the relative risks by taking the general Swedish population as reference. To minimise the possible influence of selection bias, we excluded the first year observations. RESULTS: Alcoholics had only a modest 40% excess risk of pancreatic cancer (SIR 1.4, 95% confidence interval (CI) 1.2-1.5). Overrepresented smokers among alcoholics might confound a true SIR of unity among alcoholics to approximately 1.4. SIR among alcoholic chronic pancreatitis patients (2.2, 95% CI 0.9-4.5) was considerably lower than that among non-alcoholic chronic pancreatitis patients (8.7, 95% CI 6.8-10.9), and decreased with increasing duration of follow up in both groups, indicating that most of the excess might be explained by reversed causation from undiagnosed cancers. Among patients with alcoholic liver cirrhosis, the increased risk of pancreatic cancer was also moderate (SIR 1.9, 95% CI 1.3-2.8) while no significant excess risk was found among non-alcoholic liver cirrhosis patients (SIR 1.2, 95% CI 0.6-2.2). CONCLUSIONS: The excess risk for pancreatic cancer among alcoholics is small and could conceivably be attributed to confounding by smoking. PMID- 12117885 TI - Serum levels of procarboxypeptidase B and its activation peptide in patients with acute pancreatitis and non-pancreatic diseases. AB - BACKGROUND: Carboxypeptidase B from the pancreatic gland may exist in three different molecular and immunoreactive forms: the proenzyme, the active enzyme, and the activation peptide. AIMS: To investigate levels of procarboxypeptidase B (proCAPB) and its activation peptide in serum in acute pancreatitis to test the accuracy of these two variables as markers for the diagnosis of acute pancreatitis and for prediction of pancreatic necrosis. To elucidate whether leakage of proenzymes and activation of proenzymes reflect two different pathophysiological events in acute pancreatitis. METHODS: Sera from patients with acute pancreatitis (n=85) and acute abdominal pain of non-pancreatic origin (n=53) were analysed for proCAPB and its activation peptide. Patients with pancreatitis were divided into necrotising (n=33) and oedematous attacks (n=52) using contrast enhanced computed tomography. Accuracy was determined using receiver operating characteristic curve analysis. RESULTS: Immunoreactive carboxypeptidase B activation peptide (ir-CAPAP) concentration in serum on admission was 0.7 nmol/l (0-18.1) in patients with oedematous pancreatitis compared with 5.8 nmol/l (1.9-34) in patients with later development of pancreatic necrosis. Elevated levels of the activation peptide on admission correlated with an accuracy of 92% to later development of pancreatic necrosis. Ir-proCAPB concentration in serum on admission was 16.0 nmol/l (1.4-50.5) in all patients with acute pancreatitis versus 0.3 nmol/l (0-3.6) in patients with non pancreatic acute abdominal disorders. Cases with oedematous pancreatitis had ir proCAPB levels of 15.4 nmol/l (1.4-50.5) versus 19.1 nmol/l (2.7-36.1) in cases with later development of pancreatic necrosis. Measurement of the proenzyme can thus be useful for the diagnosis of acute pancreatitis (accuracy 99%) but levels did not correlate with later development of pancreatic necrosis (accuracy 56%). CONCLUSION: Leakage of proenzymes occurs in acute pancreatitis, irrespective of severity, while development of pancreatic necrosis occurs only when there is activation of the proenzymes. PMID- 12117887 TI - Endoscopic transpapillary biopsies and intraductal ultrasonography in the diagnostics of bile duct strictures: a prospective study. AB - BACKGROUND: In bile duct strictures, examination of wall layers by intraductal ultrasonography (IDUS) performed during endoscopic retrograde cholangiopancreatography (ERCP) may be diagnostically useful. METHODS: In the present study 60 patients with bile duct strictures of unknown aetiology were examined preoperatively by ERCP, including transpapillary biopsies and IDUS. Histopathological correlation was available for all patients undergoing these procedures. RESULTS: Postoperative diagnosis revealed 30 pancreatic carcinomas, 17 bile duct cancers, three gall bladder cancers, and 10 benign bile duct strictures. Using endoscopic transpapillary forceps biopsies (ETP), a correct preoperative diagnosis was achieved in 36 of 60 patients (60% of cases). Among the 50 malignant tumours, preoperative diagnosis by ETP revealed a sensitivity of 52% and a specificity of 100%. ERCP supplemented by IDUS allowed for correct preoperative diagnosis in 83% of cases (50 of 60 patients), which was significantly higher than the accuracy of ETP (p=0.008). By combining ETP with IDUS, a correct preoperative diagnosis was made in 59 of 60 patients resulting in an accuracy rate of 98%. CONCLUSIONS: Because of its low accuracy, exclusive use of ETP is not a reliable diagnostic tool for a definitive preoperative diagnosis of bile duct strictures. By combining IDUS and ETP with ERCP however, preoperative diagnostic accuracy can be improved substantially. PMID- 12117888 TI - A randomised controlled trial of endoscopic sphincterotomy in acute cholangitis without common bile duct stones. AB - BACKGROUND: Biliary decompression with endoscopic sphincterotomy (EPT) is beneficial in patients with biliary obstruction due to common bile duct (CBD) stones. However, it is not known whether EPT with decompression of the bile duct is beneficial in patients with acute cholangitis and gall bladder stones but without evidence of CBD stones. AIM: A randomised controlled study to assess the effect of EPT on the outcome of patients suffering from acute cholangitis with gall bladder stones but with no CBD stones on initial endoscopic retrograde cholangiopancreatography. PATIENTS: A total of 111 patients were recruited into the study. METHODS AND RESULTS: Fifty patients were randomised to receive EPT while 61 patients received no endoscopic intervention. There was a significant difference in the duration of fever in the EPT and non-EPT groups (mean (SD): 3.2 (2.2) days v 4.3 (2.1) days; p<0.001). Duration of hospital stay was also shorter in the EPT group than in the non-EPT group (mean (SD): 8.1 (3.0) v 9.1 (3.2) days; p=0.04). Patients were followed up for a mean (SD) of 42.4 (11.1) months. Twenty three patients (20.3%) developed recurrent acute cholangitis (RAC): 14 patients (12.6%) in the EPT group and nine patients (8.1%) in the non-EPT group (p=0.09). CONCLUSION: EPT in patients with acute cholangitis without CBD stones decreased the duration of acute cholangitis and reduced hospital stay but it did not decrease the incidence of RAC. PMID- 12117889 TI - Advancing donor liver age and rapid fibrosis progression following transplantation for hepatitis C. AB - BACKGROUND AND AIM: Cirrhosis with liver failure due to hepatitis C virus (HCV) infection is the most common indication for liver transplantation (LT). Reinfection of the transplanted liver by HCV is inevitable, and aggressive hepatitis with accelerated progression to graft cirrhosis may be observed. Of concern, recent reports suggest that the outcome of LT for HCV may have deteriorated in recent years. Determinants of rate of progression to cirrhosis in the immunocompetent non-transplant patient are well defined, and the most powerful determinant is patient age at the time of infection. Following LT for HCV, recipient age does not affect outcome of HCV reinfection. However, the impact of donor age on graft fibrosis progression rate following LT has not been examined. METHODS: We have examined post-transplant biopsies to assess histological activity, including fibrosis stage (scored 0-6 units, 6 representing established cirrhosis), and to calculate fibrosis progression rates in 101 post transplant specimens from 56 HCV infected LT patients. Univariate and multivariate analyses examined the impact of parameters including recipient and donor age and sex on fibrosis progression rate, and on predicted time to cirrhosis. RESULTS: For the cohort, median fibrosis progression rate was 0.78 units/year, and median interval from transplantation to development of cirrhosis was 7.7 years. In multivariate analysis, donor age (not recipient age) was a powerful determinant (p=0.02) of fibrosis progression rate. When the liver donor was younger than 40 years, median progression rate was 0.6 units/year and interval to cirrhosis was 10 years. When the donor was aged 50 years or more, median progression rate was 2.7 units/year and interval to cirrhosis only 2.2 years. During the observation period there has been a significant increase in donor age (p=0.01) but date of transplantation per se is not a determinant of progression rate when included in multivariate analyses. CONCLUSIONS: Donor age has a major influence on graft outcome following transplantation for HCV. The changing organ donor profile will affect the long term results of LT for HCV. These observations have important implications for donor liver allocation. PMID- 12117890 TI - INK4a-ARF alterations in liver cell adenoma. AB - BACKGROUND: The INK4a-ARF (CDKN2A) locus on chromosome 9p21 encodes two tumour suppressor proteins, p16(INK4a) and p14(ARF), whose functions are inactivated in many human cancers. AIMS: To evaluate p14(ARF) and p16(INK4a) alterations in liver cell adenoma. METHODS: After microdissection, DNA from 25 liver cell adenomas and corresponding normal liver tissue were analysed for INK4-ARF inactivation by DNA sequence analysis, methylation specific polymerase chain reaction, restriction enzyme related-polymerase chain reaction (RE-PCR), mRNA expression, microsatellite analysis, and immunohistochemistry. In addition, microdeletion of p14(ARF) and p16(INK4a) were assessed by differential PCR. RESULTS: Methylation of p14(ARF) was found in 3/25 cases (12%) and alterations in p16(INK4a) occurred in 6/25 liver cell adenomas (24%) which correlated with loss of mRNA transcription. We failed to detect microdeletions or specific mutations of both exons. p16(INK4a) methylation appeared in the context of an unmethylated p14(ARF) promoter in six cases. In normal liver tissue, p14(ARF) or p16(INK4a) alterations were not observed. CONCLUSIONS: Our data suggest that p14(ARF) methylation occurs independently of p16(INK4a) alterations in liver cell adenomas. Furthermore, methylation of p14(ARF) and p16(INK4a) may be a result of cell cycle deregulation and does not seem to be a prerequisite of malignancy. PMID- 12117891 TI - Establishment of standardised SLA/LP immunoassays: specificity for autoimmune hepatitis, worldwide occurrence, and clinical characteristics. AB - BACKGROUND: Antibodies to soluble liver antigen/liver pancreas (SLA/LP) are specific markers of autoimmune hepatitis. Their target antigen has recently been cloned. AIMS: To establish standardised immunoassays using the recombinant antigen, and to assess the frequency and significance of seropositivity in patients from different countries. METHODS: An enzyme linked immunoassay was developed using purified recombinant antigen and validated by testing sera from 200 healthy blood donors and 1026 patients with various liver and non-liver diseases. The assay was then applied to 454 sera from 419 patients with autoimmune hepatitis from different countries. All sera were also tested by inhibition immunoassay and western blot. RESULTS: Antibodies were reliably detected by the recombinant immunoassay and occurred exclusively in patients with autoimmune liver disease. Twenty three of 149 patients from the USA (15%), 23/132 from Brazil (17%), 21/108 from Germany (19%), and 2/30 from Japan (7%) were seropositive. Clinical features at presentation were similar between seropositive and seronegative patients. However, relapse after corticosteroid withdrawal or during maintenance therapy occurred more commonly in seropositive patients. CONCLUSIONS: Antibodies to SLA/LP can be reliably detected by these standardised immunoassays based on recombinant antigen. Antibodies to SLA/LP occur with similar frequencies in different geographical regions, races, and age groups, and are of exquisite diagnostic specificity. Whether SLA/LP positive patients represent a clinically distinct subgroup remains to be determined; relapse during treatment reduction appeared to be more common in the SLA/LP group. PMID- 12117892 TI - Hyperlipidaemic state and cardiovascular risk in primary biliary cirrhosis. AB - BACKGROUND: Primary biliary cirrhosis (PBC), a chronic cholestatic liver disease, is frequently associated with severe hypercholesterolaemia but the clinical significance of this finding is unclear. AIMS: To characterise changes in serum lipid profile over time and to assess the risk of cardiovascular disease in PBC. SUBJECTS AND METHODS: We studied a cohort of 400 PBC patients for 6.2 years (range 4 months to 24 years) by serial determinations of serum lipid levels and registration of all cardiovascular events. Subjects included in an Italian prospective population based study served as controls. RESULTS: At presentation, 76% of patients had serum cholesterol levels >5.2 mmol/l. Hyperbilirubinaemic patients had higher total cholesterol and lower high density lipoprotein (HDL) cholesterol levels (p<0.001). With time, disease progression was associated with a reduction in total (p<0.001) and HDL (p<0.05) cholesterol. The incidence of cardiovascular events was similar to that of the general population (cerebrovascular events: standardised ratio 1.4; 95% confidence interval 0.5-3.7; coronary events: 2.2; 0.9-4.3). Hypertension was associated with an increased risk of cardiovascular events (3.8; 1.6-8.9). Association with moderate hypercholesterolaemia was of borderline significance (3.8; 0.9-17) whereas severe hypercholesterolaemia was not associated with increased risk (2.4, 0.5-11). CONCLUSIONS: In PBC, serum cholesterol levels markedly increase with worsening of cholestasis, and decrease in the late disease stages, despite a severe reduction in biliary secretion. Marked hypercholesterolaemia, typical of severe longstanding cholestasis, is not associated with an excess risk of cardiovascular disease while less advanced patients with moderate hypercholesterolaemia are exposed to an increased cardiovascular risk. Putative protective factors in PBC patients with severe hypercholesterolaemia should be assessed. PMID- 12117893 TI - The role of the transjugular intrahepatic portosystemic stent shunt (TIPSS) in the management of bleeding gastric varices: clinical and haemodynamic correlations. AB - BACKGROUND: The transjugular intrahepatic portosystemic stent shunt (TIPSS) is effective in the management of both oesophageal and gastric variceal bleeding. Although it has been reported that gastric varices can bleed at pressures of < or = 12 mm Hg, this phenomenon has been little studied in the clinical setting. AIMS: To assess the efficacy of TIPSS on rebleeding and mortality following gastric and oesophageal variceal bleeding, and the importance of portal pressure in both groups. METHODS: Forty eligible patients who had bled from gastric varices and 232 from oesophageal varices were studied. Patients were also subdivided into those whose portal pressure gradients (PPG) prior to TIPSS were < or = 12 mm Hg (group 1) and >12 mm Hg (group 2). RESULTS: There was no difference in Child-Pugh score, age, sex, or alcohol related disease between patients bleeding from gastric or oesophageal varices. Patients who bled from gastric varices had a lower PPG pre-TIPSS (15.8 (0.8) v 21.44 (0.4) mm Hg; p<0.001). There was no difference in the rebleeding rate (20.0% v 14.7%; NS). There was a significant difference (p<0.05) in favour of the gastric varices group in the one year mortality (30.7% v 38.7%) and five year mortality (49.5% v 74.9%), particularly in those patients in group 2. Gastric variceal bleeding accounted for significantly more cases in group 1 than in group 2 (36.8% v 10.2%; p<0.001). Most patients in group 2 who rebled had a PPG post-TIPSS of >7 mm Hg. CONCLUSIONS: TIPSS is equally effective in the prevention of rebleeding following gastric and oesophageal variceal bleeding. A significant proportion of gastric varices bleed at a PPG < or = 12 mm Hg. The improved mortality in patients with gastric variceal bleeding is seen only in those that bleed at a PPG >12 mm Hg, and warrants further study. PMID- 12117894 TI - Idiopathic non-cirrhotic intrahepatic portal hypertension in the West: a re evaluation in 28 patients. AB - BACKGROUND: Non-cirrhotic portal hypertension of unknown cause is a poorly understood condition attributed to obstructive portal venopathy. AIM: To reassess the manifestations, course, and causes, with special attention to thrombosis. METHODS: Analysis of a cohort of 28 patients. RESULTS: Gastrointestinal bleeding occurred in 11 patients. Liver failure developed at the time of concurrent disease in eight patients, including all four patients who died. Portal vein thrombosis developed in 13 patients. A prothrombotic disorder was found in 12 of 23 fully investigated patients. Hepatoportal sclerosis was observed in 11 patients (with associated perisinusoidal fibrosis and/or nodular regenerative hyperplasia in six); periportal fibrosis, perisinusoidal fibrosis, nodular regenerative hyperplasia, or a combination thereof were observed in other patients. A morphometric evaluation showed an increased number of portal vessels in patients with hepatoportal sclerosis. There was no relation between pathological results and haemodynamic findings or prothrombotic disorders. CONCLUSIONS: Outcome was related to associated conditions. Overlap in pathological, haemodynamic, and causal features suggests a single entity, with prothrombotic disorders as major causal factors, and injury to sinusoids as well as to portal venules as the primary mechanism. Activated coagulation could mediate vascular injury in the absence of thrombosis. Anticoagulation should be considered. PMID- 12117895 TI - A case of primary low grade mucosa associated lymphoid tissue (MALT) lymphoma of the oesophagus. AB - We report a very rare case of primary low grade mucosa associated lymphoid tissue (MALT) lymphoma of the oesophagus. An 83 year old woman was referred to our hospital in June 1999 for further examination and treatment of oesophageal tumour. Although a physical examination and laboratory data showed no significant abnormalities, endoscopic observation revealed two slightly elevated submucosal tumour-like lesions of the oesophagus. Tissue specimens were obtained by endoscopic mucosal resection of the oesophagus using a cap fitted panendoscope. The lesions were composed of diffuse small atypical lymphoid cells--that is, centrocyte-like cells--which were stained with CD20, L26, BCL-2, and kappa, but not with CD3, CD5, CD10, or cyclin D1. Monoclonality was detected by polymerase chain reaction analysis using the primer for CDR-3 of immunoglobulin H and diagnosed as low grade MALT lymphoma of the oesophagus. The tumours were considered to be completely resected and therefore additional treatment was not administered. The patient is alive and well 22 months after treatment and diagnosis. PMID- 12117896 TI - Emergency colonoscopy for distal intestinal obstruction syndrome in cystic fibrosis patients. PMID- 12117897 TI - Evaluating ERCP is important but difficult. AB - ERCP is a valuable technique now practised widely throughout the world. It revolutionised the diagnosis and management of benign and malignant biliary and pancreatic diseases in the 1970s and 1980s. However, recent developments have highlighted the need for detailed evaluation of current ERCP practice. This review is based on a presentation to a recent NIH "state of the science" conference on ERCP, and refers to an article which appears in this issue of Gut from researchers in Hong Kong who report on a randomised controlled trial of endoscopic sphincterotomy in acute cholangitis. PMID- 12117899 TI - Current guidelines fail young patients with oesophagogastric cancer. PMID- 12117898 TI - Molecular pathogenesis of iron overload. AB - Our current understanding of iron absorption under normal conditions is presented, together with an overview of the clinical disorders of iron overload and the molecular processes that contribute to increased iron deposition in iron overload. Recently, a number of new genes involved in iron metabolism have been identified which is allowing the molecular mechanisms of iron absorption to be elucidated. PMID- 12117900 TI - Immunosuppression, IBD, and risk of lymphoma. PMID- 12117901 TI - GRP and stimulation of acid secretion. PMID- 12117902 TI - Acute appendicitis in Japanese soldiers in Burma: support for the "fibre" theory. PMID- 12117903 TI - Bacteriophage control of bacterial virulence. PMID- 12117904 TI - Leishmania priming of human dendritic cells for CD40 ligand-induced interleukin 12p70 secretion is strain and species dependent. AB - A major question in the study of leishmaniasis is what dictates clinical disease expression produced by different Leishmania species, i.e., cutaneous versus systemic and healing versus nonhealing. Animal models using a Leishmania species associated with self-limiting cutaneous disease (L. major) have revealed that protective immunity requires CD40/CD40 ligand (CD40L)-dependent, interleukin-12 (IL-12)-driven Th1 responses. We recently showed that L. major can prime human dendritic cells (DCs) for CD40L-triggered IL-12p70 secretion and that these cells can drive a Th1 response in autologous T cells from sensitized individuals. Here we show that in contrast to L. major, Leishmania species responsible for visceral disease (L. donovani), as well as species associated with persistent, cutaneous lesions and occasional systemic disease (L. tropica), did not induce CD40L dependent IL-12p70 production, despite comparable levels of uptake by DCs. Up regulated surface expression of CD40 did not correlate with IL-12p70 production, and appreciable CD40L-induced IL-12p40 secretion was observed in uninfected as well as infected DCs, regardless of species. Reverse transcription-PCR analysis confirmed that the production of heterodimeric IL-12 was limited by expression of IL-12p35 mRNA, which was dependent on both a microbial priming signal and CD40 engagement for its high-level induction. The intrinsic differences in the ability of Leishmania species to prime DCs for CD40L-dependent IL-12p70 secretion may account, at least in part, for the evolution of healing and nonhealing forms of leishmanial disease. PMID- 12117905 TI - Up-regulation of Fas (CD95) and induction of apoptosis in intestinal epithelial cells by nematode-derived molecules. AB - Infection by the intestinal nematode Nippostrongylus brasiliensis induces acceleration of apoptosis in the small intestinal villus epithelial cells in vivo. In the present study, we examined whether worm extract or excretory secretory product induces apoptosis in the rat intestinal epithelial cell line IEC-6 in vitro. In the presence of worm extract or excretory-secretory product (> or =6 microg/ml), IEC-6 cell growth was significantly suppressed, and there was a concomitant increase in the number of detached cells in culture dishes. Detached cells showed nuclear fragmentation, activation of caspase-3, and specific cleavage of poly(ADP-ribose) polymerase, suggesting that apoptosis was induced in these cells. Semiquantitative reverse transcription-PCR showed that expression of Fas (CD95) mRNA was up-regulated as early as 6 h after addition of excretory secretory product, while Fas ligand expression and p53 expression were not up regulated. Fluorescence-activated cell sorter analyses revealed a significant increase in Fas expression and a slight increase in FasL expression in IEC-6 cells cultured in the presence of excretory-secretory product, while control IEC 6 cells expressed neither Fas or FasL. These results indicated that N. brasiliensis worms produce and secrete biologically active molecules that trigger apoptosis in intestinal epithelial cells together with up-regulation of Fas expression, although the mechanism of induction of apoptosis remains to be elucidated. PMID- 12117906 TI - Salmonella enterica serovar Typhi live vector vaccines delivered intranasally elicit regional and systemic specific CD8+ major histocompatibility class I restricted cytotoxic T lymphocytes. AB - We investigated the ability of live attenuated Salmonella enterica serovar Typhi strains delivered to mice intranasally to induce specific cytotoxic T-lymphocyte (CTL) responses at regional and systemic levels. Mice immunized with two doses (28 days apart) of Salmonella serovar Typhi strain Ty21a, the licensed oral typhoid vaccine, and genetically attenuated mutants CVD 908 (DeltaaroC DeltaaroD), CVD 915 (DeltaguaBA), and CVD 908-htrA (DeltaaroC DeltaaroD DeltahtrA) induced CTL specific for Salmonella serovar Typhi-infected cells in spleens and cervical lymph nodes. CTL were detected in effector T cells that had been expanded in vitro for 7 days in the presence of Salmonella-infected syngeneic splenocytes. A second round of stimulation further enhanced the levels of specific cytotoxicity. CTL activity was observed in sorted alphabeta+ CD8+ T cells, which were remarkably increased after expansion, but not in CD4+ T cells. CTL from both cervical lymph nodes and spleens failed to recognize Salmonella infected major histocompatibility complex (MHC)-mismatched cells, indicating that the responses were MHC restricted. Studies in which MHC blocking antibodies were used showed that H-2L(d) was the restriction element. This is the first demonstration that Salmonella serovar Typhi vaccines delivered intranasally elicit CD8+ MHC class I-restricted CTL. The results further support the usefulness of the murine intranasal model for evaluating the immunogenicity of typhoid vaccine candidates at the preclinical level. PMID- 12117907 TI - Phosphoantigen presentation by macrophages to mycobacterium tuberculosis- reactive Vgamma9Vdelta2+ T cells: modulation by chloroquine. AB - Vgamma9Vdelta2+ T cells (gammadelta T cells) are activated by Mycobacterium tuberculosis and recognize mycobacterial nonpeptide phosphoantigens. The role of antigen-presenting cells in the processing and presentation of phosphoantigens to Vgamma9Vdelta2+ T cells is not understood. We analyzed the role of macrophages for activation of gammadelta T cells by a new synthetic phosphoantigen bromohydrin pyrophosphate (BrHPP) and M. tuberculosis. Macrophages greatly increased gammadelta T-cell activation by both BrHPP and M. tuberculosis. Fixation of macrophages before infection demonstrated that uptake of M. tuberculosis was required for presentation to gammadelta T cells. Antigens of M. tuberculosis remained stably associated with macrophage surface and were not removed by paraformaldehyde fixation or washing. Macrophages processed M. tuberculosis for gammadelta T cells through a brefeldin A-insensitive pathway, suggesting that transport through the endoplasmic reticulum and Golgi complex of a putative presenting molecule is not important in the early processing of M. tuberculosis antigens for gammadelta T cells. Processing of M. tuberculosis was not eliminated by chloroquine, indicating that processing of gammadelta antigens is not dependent on acidic pH in the lysosomes. Chloroquine treatment of BrHPP pulsed macrophages increased activation of gammadelta T cells. Ammonium chloride treatment of macrophages did not increase reactivity of gammadelta T cells to BrHPP, indicating that the effect of chloroquine was independent of pH changes in endosomes. Chloroquine, by inhibiting membrane traffic, may increase association and retention of phosphoantigens with cell surface membrane molecules on macrophages. PMID- 12117908 TI - Immunization with the recombinant PorB outer membrane protein induces a bactericidal immune response against Neisseria meningitidis. AB - Infections with Neisseria meningitidis are characterized by life-threatening meningitis and septicemia. The meningococcal porin proteins from serogroup B meningococci have been identified as candidates for inclusion in vaccines to prevent such infections. In this study, we investigated the vaccine potential of the PorB porin protein free of other meningococcal components. The porB gene from a strain of Neisseria meningitidis expressing the class 3 outer membrane porin protein (PorB3) was cloned into the pRSETB vector, and the protein was expressed at high levels in a heterologous host Escherichia coli. The recombinant protein was purified to homogeneity by affinity chromatography and used for immunization after incorporation into liposomes and into micelles composed either of zwitterionic detergent or nondetergent sulfobetaine. The immunogenicity of these preparations was compared to recombinant PorB protein adsorbed to Al(OH)(3) adjuvant as a control. Although sera raised against the protein adsorbed to Al(OH)(3) reacted with the purified recombinant protein, sera raised against liposomes and micelles showed greater activity with native protein, as measured by enzyme immunoassay with outer membranes and by whole-cell immunofluorescence. Reactivity with native protein was considerably enhanced by incorporation of the adjuvant monophosphoryl lipid A into the liposome or micelle preparations. Recognition of the native protein was in a serotype-specific manner and was associated with the ability of the antisera to promote high levels of serotype specific complement-mediated killing of meningococci. These results demonstrate that the PorB protein should be considered as a component of a vaccine designed to prevent serogroup B meningococcal infection. PMID- 12117909 TI - Interaction of Neisseria meningitidis with human meningeal cells induces the secretion of a distinct group of chemotactic, proinflammatory, and growth-factor cytokines. AB - The interactions of Neisseria meningitidis with cells of the leptomeninges are pivotal events in the progression of bacterial leptomeningitis. An in vitro model based on the culture of human meningioma cells was used to investigate the role of the leptomeninges in the inflammatory response. Following challenge with meningococci, meningioma cells secreted specifically the proinflammatory cytokine interleukin-6 (IL-6), the CXC chemokine IL-8, the CC chemokines monocyte chemoattractant protein 1 (MCP-1) and regulated-upon-activation, normal-T-cell expressed and secreted protein (RANTES), and the cytokine growth factor granulocyte-macrophage colony-stimulating factor (GM-CSF). A temporal pattern of cytokine production was observed, with early secretion of IL-6, IL-8, and MCP-1 followed by later increases in RANTES and GM-CSF levels. IL-6 was induced equally by the interactions of piliated and nonpiliated meningococci, whereas lipopolysaccharide (LPS) had a minimal effect, suggesting that other, possibly secreted, bacterial components were responsible. Induction of IL-8 and MCP-1 also did not require adherence of bacteria to meningeal cells, but LPS was implicated. In contrast, efficient stimulation of RANTES by intact meningococci required pilus-mediated adherence, which served to deliver increased local concentrations of LPS onto the surface of meningeal cells. Secretion of GM-CSF was induced by pilus-mediated interactions but did not involve LPS. In addition, capsule expression had a specific inhibitory effect on GM-CSF secretion, which was not observed with IL-6, IL-8, MCP-1, or RANTES. Thus, the data demonstrate that cells of the leptomeninges are not inert but are active participants in the innate host response during leptomeningitis and that there is a complex relationship between expression of meningococcal components and cytokine induction. PMID- 12117910 TI - Rickettsia rickettsii infection of cultured human endothelial cells induces heme oxygenase 1 expression. AB - Existing evidence suggests that oxidative insults and antioxidant defense mechanisms play a critical role in the host cell response during infection of endothelial cells by Rickettsia rickettsii, the causative agent of Rocky Mountain spotted fever. Heme oxygenase (HO), a rate-limiting enzyme in the pathway for heme catabolism, protects against oxidant damage in a variety of stress situations. Here, we report on the expression of the inducible and constitutive HO isozymes, HO-1 and HO-2, during R. rickettsii infection of endothelial cells. Steady-state levels for HO-1 mRNA were increased two- to threefold, as early as 4 h postinfection, whereas HO-2 mRNA was not affected. Induction of HO-1 mRNA was dependent on the dose of infection and occurred in a time-dependent manner, reaching maximal levels at 4 to 7 h. The increase in HO-1 mRNA occurred at the level of trancription as it was blocked by the transcriptional inhibitors, actinomycin D and alpha-amanitin. The eukaryotic protein synthesis inhibitor, cycloheximide, caused a >50% reduction in the infection-induced increase in HO-1 mRNA level, suggesting its dependence on de novo protein synthesis of host cell. The uptake of viable organisms appeared to be necessary, since inactivation of R. rickettsii by heat or formalin fixation, or incubation of cells with cytochalasin B to prevent entry resulted in marked inhibition of HO-1 response. N-Acetyl-L cysteine, a known oxidant scavenger, inhibited the HO-1 induction by R. rickettsii. Finally, Western analysis with a specific monoclonal antibody revealed higher levels of HO-1 protein ( approximately 32 kDa), confirming that changes in HO-1 mRNA levels were followed by increases in the levels of protein. The findings indicate that R. rickettsii infection induces HO-1 expression in host endothelial cells and suggest an important role for this enzyme in cellular response to infection, possibly by serving a protective function against oxidative injury. PMID- 12117911 TI - Proteasomal degradation of cytotoxic necrotizing factor 1-activated rac. AB - The cytotoxic necrotizing factor 1 (CNF1) from Escherichia coli has been shown to activate members of the Rho family by deamidation of glutamine 63. This amino acid is essential for hydrolysis of GTP, and any substitution results in a constitutively active Rho. Activation of Rho induces the formation of stress fibers, filopodia, and membrane ruffles due to activation of RhoA, Cdc42, and Rac, respectively. Here we show that the level of endogenous Rac decreased in CNF1-treated HEK293 and HeLa cells. The amount of mRNA remained unaffected, leaving the possibility that Rac is subject to proteolytic degradation. Treatment of cells with lactacystin, an inhibitor of the 26S proteasome, protected Rac from degradation. We have previously shown that CNF1 activates the c-Jun N-terminal kinase (JNK) only transiently in HeLa cells (M. Lerm, J. Selzer, A. Hoffmeyer, U. R. Rapp, K. Aktories, and G. Schmidt, Infect. Immun. 67:496-503, 1998). Here we show that CNF1-induced JNK activation is stabilized in the presence of lactacystin. The data indicate that Rac is degraded by a proteasome-dependent pathway in CNF1-treated cells. PMID- 12117912 TI - Microarray-based identification of htrA, a Streptococcus pneumoniae gene that is regulated by the CiaRH two-component system and contributes to nasopharyngeal colonization. AB - Nasopharyngeal carriage is the reservoir from which most disease with Streptococcus pneumoniae arises. Survival as a commensal in this environment is likely to require a set of adaptations distinct from those needed to cause disease, some of which may be mediated by two-component signal transduction systems (TCSTS). We examined the contributions of nine pneumococcal TCSTS to the process of nasopharyngeal colonization by using an infant rat model. Whereas deletions in all but one of these systems have been associated previously with a high degree of attenuation in a murine model of pneumonia, only the CiaRH system was necessary for efficient carriage. Transcriptional analysis by using microarray hybridization identified a locus consisting of two adjacent genes, htrA and spoJ, that was specifically and strongly downregulated in a DeltaciaRH null mutant. A S. pneumoniae strain lacking the htrA gene encoding a putative serine protease, but not one lacking spoJ, showed decreased fitness in a competitive model of colonization, a finding consistent with this gene mediating a portion of the carriage deficit observed with the DeltaciaRH strain. PMID- 12117913 TI - Lipopolysaccharide-binding protein- and CD14-dependent activation of mitogen activated protein kinase p38 by lipopolysaccharide in human neutrophils is associated with priming of respiratory burst. AB - Neutrophil (PMN) functions can be primed for greatly increased oxidative radical release by exposure to certain agents such as lipopolysaccharide (LPS). Although a variety of signaling pathways involving both tyrosine kinases and mitogen activated protein (MAP) kinases may be operative, the mechanisms of PMN priming are still not understood. We found that PMN priming was not achieved by treatment of cells with a very low concentration (5 ng/ml) of LPS unless additional "helper" factors were present in plasma (5%). Under these conditions, LPS induced tyrosine phosphorylation of a 38-kDa protein, which was coincident with the MAP kinase p38 action in this situation. LPS-mediated activation of p38 in human PMNs was dependent on the presence of LPS binding protein from plasma and CD14 on the surfaces of the cells. Phosphorylation of p38 was highly correlated with LPS priming of a formyl-methionyl-leucyl-phenylalanine (fMLP)-stimulated PMN respiratory burst. Treatment of PMN with the p38-specific inhibitor SB203580 significantly attenuated the respiratory burst in cells primed by LPS and stimulated by fMLP. These results suggest that the LPS signaling pathway leading to p38 activation may be an important mechanism in regulation of PMN priming. The mediator(s) linking CD14 to p38 involves proteins that are functionally sensitive to genistein but insensitive to tyrphostin AG126 and to Src- and Syk-family kinase, protein kinase C, and phosphatidylinositol 3-kinase inhibitors. Elucidating this pathway will provide insight into possible regulation of PMN priming by LPS. PMID- 12117914 TI - Bacterium-induced CXCL10 secretion by osteoblasts can be mediated in part through toll-like receptor 4. AB - Two common pathogens known to cause bone infection, Salmonella and Staphylococcus aureus, were investigated to determine their abilities to induce chemokine expression in cultured mouse and human osteoblasts. While these cells are responsible for bone formation, we were surprised to find that they could respond to bacterial infection by upregulating expression of the chemokine CXCL10 (IP 10). However, there were significant differences in the abilities of the gram negative bacterium Salmonella and the gram-positive bacterium S. aureus to induce expression of CXCL10. Reverse transcription-PCR and enzyme-linked immunosorbent assay analyses showed high levels of Salmonella-induced CXCL10 mRNA and protein expression, respectively, whereas the osteoblast response to S. aureus was significantly less. Consistent with these findings, Salmonella-derived lipopolysaccharide (LPS), but not S. aureus-derived peptidoglycan, could induce expression of CXCL10. An antibody against toll-like receptor 4 (TLR4) could block the LPS-induced CXCL10 production, demonstrating the functional expression of TLR4 by osteoblasts. Despite the inducible nature of TLR2 mRNA expression by bacterium-infected osteoblasts, peptidoglycan failed to stimulate CXCL10 secretion. Immunofluorescent staining of bacterium-infected calvaria (i.e., skull bone) demonstrated the presence of CXCL10 in osteoblasts. The fact that osteoblasts did not express CXCR3 mRNA, whereas T lymphocytes can express high levels of this receptor, suggests that osteoblast-derived CXCL10 may recruit T lymphocytes to the sites of bone infections. PMID- 12117915 TI - Recurrent variable region gene usage and somatic mutation in the human antibody response to the capsular polysaccharide of Streptococcus pneumoniae type 23F. AB - Combinatorial cloning and expression library analysis were used to isolate human antibody Fab fragments specific for the capsular polysaccharide of Streptococcus pneumoniae serotype 23F. Thirty 23F-specific Fabs were isolated from seven vaccinated donors, and the sequences of the heavy (H)- and light (L)-chain variable regions were determined. All individuals utilized either the Vkappa A23 L chain, the Vkappa L6 L chain, or both chains in forming the 23F-specific combining site. Vkappa A23 L chains paired primarily with VH3-23 H chains. Vkappa L6 L chains were more promiscuous in heavy-chain usage between individuals. Both H and L chains were mutated, primarily in the complementarity-determining regions, compared to their closest germ line counterpart, suggesting a recall response that has undergone affinity maturation. H-chain isotypes were reflective of those found in the serum. Shared somatic modifications demonstrated that immunoglobulin G2 (IgG2) and IgA antibodies arose from the same somatically matured B cell. Our results indicate that the response to the serotype 23F pneumococcal capsular polysaccharide is oligoclonal within the individual, with one or two paratope families accounting for the majority of expressed antibody. We also determined that, in spite of the combinatorial diversity available to the immune system, the 23F-specific response is highly restricted at the population level, with the same two L-chain-determined paratope families recurring in all individuals. Lastly, analysis of the isolated Fabs indicate all have undergone extensive somatic mutation, as well as class switch, maturational events that presumably require the participation of T cells. PMID- 12117916 TI - Modification of the structure and activity of lipid A in Yersinia pestis lipopolysaccharide by growth temperature. AB - Yersinia pestis strain Yreka was grown at 27 or 37 degrees C, and the lipid A structures (lipid A-27 degrees C and lipid A-37 degrees C) of the respective lipopolysaccharides (LPS) were investigated by matrix-assisted laser desorption ionization-time-of-flight (MALDI-TOF) mass spectrometry. Lipid A-27 degrees C consisted of a mixture of tri-acyl, tetra-acyl, penta-acyl, and hexa-acyl lipid A's, of which tetra-acyl lipid A was most abundant. Lipid A-37 degrees C consisted predominantly of tri- and tetra-acylated molecules, with only small amounts of penta-acyl lipid A; no hexa-acyl lipid A was detected. Furthermore, the amount of 4-amino-arabinose was substantially higher in lipid A-27 degrees C than in lipid A-37 degrees C. By use of mouse and human macrophage cell lines, the biological activities of the LPS and lipid A preparations were measured via their abilities to induce production of tumor necrosis factor alpha (TNF-alpha). In both cell lines the LPS and the lipid A from bacteria grown at 27 degrees C were stronger inducers of TNF-alpha than those from bacteria grown at 37 degrees C. However, the difference in activity was more prominent in human macrophage cells. These results suggest that in order to reduce the activation of human macrophages, it may be more advantageous for Y. pestis to produce less-acylated lipid A at 37 degrees C. PMID- 12117917 TI - LuxS-mediated quorum sensing in Borrelia burgdorferi, the lyme disease spirochete. AB - The establishment of Borrelia burgdorferi infection involves numerous interactions between the bacteria and a variety of vertebrate host and arthropod vector tissues. This complex process requires regulated synthesis of many bacterial proteins. We now demonstrate that these spirochetes utilize a LuxS/autoinducer-2 (AI-2)-based quorum-sensing mechanism to regulate protein expression, the first system of cell-cell communication to be described in a spirochete. The luxS gene of B. burgdorferi was identified and demonstrated to encode a functional enzyme by complementation of an Escherichia coli luxS mutant. Cultured B. burgdorferi responded to AI-2 by altering the expression levels of a large number of proteins, including the complement regulator factor H-binding Erp proteins. Through this mechanism, a population of Lyme disease spirochetes may synchronize production of specific proteins needed for infection processes. PMID- 12117918 TI - Novel 33-kilodalton lipoprotein from Mycobacterium leprae. AB - A novel Mycobacterium leprae lipoprotein LpK (accession no. ML0603) was identified from the genomic database. The 1,116-bp open reading frame encodes a 371-amino-acid precursor protein with an N-terminal signal sequence and a consensus motif for lipid conjugation. Expression of the protein, LpK, in Escherichia coli revealed a 33-kDa protein, and metabolic labeling experiments and globomycin treatment proved that the protein was lipidated. Fractionation of M. leprae demonstrated that this lipoprotein was a membrane protein of M. leprae. The purified lipoprotein was found to induce production of interleukin-12 in human peripheral blood monocytes. The studies imply that M. leprae LpK is involved in protective immunity against leprosy and may be a candidate for vaccine design. PMID- 12117919 TI - Plasma membrane cholesterol modulates cellular vacuolation induced by the Helicobacter pylori vacuolating cytotoxin. AB - The Helicobacter pylori vacuolating cytotoxin (VacA) induces the degenerative vacuolation of mammalian cells both in vitro and in vivo. Here, we demonstrate that plasma membrane cholesterol is essential for vacuolation of mammalian cells by VacA. Vacuole biogenesis in multiple cell lines was completely blocked when cholesterol was extracted selectively from the plasma membrane by using beta cyclodextrins. Moreover, increasing plasma membrane cholesterol levels strongly potentiated VacA-induced vacuolation. In contrast, inhibiting de novo biosynthesis of cholesterol with lovastatin or compactin had no detectable effect on vacuolation. While depletion of plasma membrane cholesterol has been shown to interfere with both clathrin-mediated endocytosis and caveola-dependent endocytosis, neither of these two internalization pathways was found to be essential for vacuolation of cells by VacA. Depleting plasma membrane cholesterol attenuated the entry of VacA into HeLa cells. In addition, beta-cyclodextrin reagents blocked vacuolation of cells that were either preloaded with VacA or had VacA directly expressed within the cytosol. Collectively, our results suggest that plasma membrane cholesterol is important for both the intoxication mechanism of VacA and subsequent vacuole biogenesis. PMID- 12117920 TI - Disruption of the gene homologous to mammalian Nramp1 in Mycobacterium tuberculosis does not affect virulence in mice. AB - Natural-resistance-associated macrophage protein 1 (Nramp1) is a divalent cation transporter belonging to a family of transporter proteins highly conserved in eukaryotes and prokaryotes. Mammalian and bacterial transporters may compete for essential metal ions during mycobacterial infections. The mycobacterial Nramp homolog may therefore be involved in Mycobacterium tuberculosis virulence. Here, we investigated this possibility by inactivating the M. tuberculosis Nramp1 gene (Mramp) by allelic exchange mutagenesis. Disruption of Mramp did not affect the extracellular growth of bacteria under standard conditions. However, the Mramp mutation was associated with growth impairment under conditions of limited iron availability. The Mramp mutant displayed no impairment of growth or survival in macrophages derived from mouse bone marrow or in Nramp1(+/+) and Nramp1(-/-) congenic murine macrophage cell lines. Following intravenous challenge in BALB/c mice, counts of parental and Mramp mutant strains were similar in the lungs and spleens of the animals at all time points studied. These results indicate that Mramp does not contribute to the virulence of M. tuberculosis in mice. PMID- 12117921 TI - Roles of p38 mitogen-activated protein kinase, NF-kappaB, and protein kinase C in proinflammatory cytokine mRNA expression by human peripheral blood leukocytes, monocytes, and neutrophils in response to Anaplasma phagocytophila. AB - Anaplasma phagocytophila, an obligately intracellular bacterium of granulocytes, causes human granulocytic ehrlichiosis. Within 2 h after addition of A. phagocytophila, interleukin-1beta (IL-1beta), tumor necrosis factor alpha (TNF alpha), and IL-6 mRNAs are induced in human peripheral blood leukocytes (PBLs) or monocytes in vitro. However, neutrophils generate only IL-1beta mRNA. In the present study, signaling pathways for induction of these three cytokines were examined. TNF-alpha and IL-6 mRNA expression by PBLs was inhibited with SB 203580 (a p38 mitogen-activated protein kinase [MAPK] inhibitor), MG-132 (a proteasome inhibitor), and SN-50 (an NF-kappaB inhibitor). Activation of p38 MAPK and NF kappaB mRNAs in monocytes was detectable within 15 to 30 min after addition of A. phagocytophila. Expression of these two cytokine mRNAs in PBLs and monocytes was also dependent on protein kinase C (PKC), protein kinase A (PKA), and protein tyrosine kinase (PTK). IL-1beta mRNA expression by neutrophils was not dependent on p38 MAPK, and p38 MAPK was not activated in neutrophils incubated with A. phagocytophila. IL-1beta mRNA induction by PBLs, monocytes, and neutrophils was dependent on PKC and PKA. Neutrophil expression of IL-1beta mRNA was dependent on transglutaminase, phospholipase C, and PTK, all of which are also required for internalization of A. phagocytophila. However, monocyte expression of IL-1beta mRNA was less dependent on these enzymes. These results suggest that A. phagocytophila transduces different signals between its host neutrophils and monocytes for proinflammatory cytokine generation. PMID- 12117922 TI - Eighty-kilodalton N-terminal moiety of Bordetella pertussis filamentous hemagglutinin: adherence, immunogenicity, and protective role. AB - Bordetella pertussis, the etiological agent of whooping cough, produces a number of factors, such as toxins and adhesins, that are required for full expression of virulence. Filamentous hemagglutinin (FHA) is the major adhesin of B. pertussis. It is a protein of approximately 220 kDa, found both associated at the bacterial cell surface and secreted into the extracellular milieu. Despite its importance in B. pertussis pathogenesis and its inclusion in most acellular pertussis vaccines, little is known about the functional importance of individual domains in infection and in the induction of protective immunity. In this study, we analyzed the role of the approximately 80-kDa N-terminal domain of FHA, designated Fha44, in B. pertussis adherence, colonization, and immunogenicity. Although Fha44 contains the complete heparan sulfate-binding domain, it is not sufficient for adherence to epithelial cells or macrophages. It also cannot replace FHA during colonization of the mouse respiratory tract. Infection with a B. pertussis strain producing Fha44 instead of FHA does not induce anti-FHA antibodies, whereas such antibodies can readily be induced by intranasal administration of purified Fha44. In addition, mice immunized with purified Fha44 were protected against challenge with wild-type B. pertussis, indicating that Fha44 contains protective epitopes. Compared to FHA, Fha44 is much smaller and much more soluble and is therefore easier to purify and to store. These advantages may perhaps warrant considering Fha44 for inclusion in acellular pertussis vaccines. PMID- 12117923 TI - Mycobacterium bovis BCG-infected mice are more susceptible to staphylococcal enterotoxin B-mediated toxic shock than uninfected mice despite reduced in vitro splenocyte responses to superantigens. AB - Type 1 T-cell responses against intracellular pathogens play a crucial role in mediating protection. We examined whether the induction of a strong type 1 T-cell response during a chronic bacterial infection influences responses to superantigens capable of inducing acute shock. Intravenous infection of mice with Mycobacterium bovis BCG appeared to induce a progressive anergy towards staphylococcal enterotoxin B (SEB) and towards antigen preparation of BCG (BCG Ag) itself, based on diminished gamma interferon (IFN-gamma) production by SEB- and BCG-Ag-stimulated splenocytes from infected mice. In contrast to these in vitro results, injection of SEB into BCG-infected mice led to a dramatic increase in the serum IFN-gamma levels and the death of infected but not of control mice. In vitro hyporesponsiveness towards SEB and BCG-Ag occurred only with unfractionated splenocyte cultures, as purified T cells from infected mice produced higher levels of IFN-gamma. Hyporesponsiveness towards SEB and BCG-Ag in unfractionated splenocyte cultures was not due to suppressive antigen-presenting cells (APCs), as APCs from infected mice stimulated higher levels of IFN-gamma from purified T cells. The diminished IFN-gamma levels observed with bulk splenocytes appear to be due to changes in the T-cell-to-APC ratio that result in a decreased proportion of T cells, coupled to reduced proliferative responses and an increased susceptibility of effector T cells to activation-induced cell death in vitro. Our results indicate that the reported phenomena of T-cell anergy during mycobacterial infection may be an in vitro consequence of the development of a strong type 1 response in vivo. PMID- 12117924 TI - Generation and characterization of human monoclonal scFv antibodies against Helicobacter pylori antigens. AB - Infection with Helicobacter pylori is chronic despite a vigorous cellular and humoral immune response and causes severe pathology in some patients. In this study, phage display was used as a new approach in order to investigate the role of the host's humoral immune response in the pathogenesis of H. pylori gastritis. Human monoclonal single-chain Fv (scFv) antibody fragments against H. pylori cell lysate and the H. pylori urease were isolated from an immune phage display library, constructed from peripheral blood lymphocytes of an H. pylori-infected patient. After affinity selection, 23% of the clones tested showed binding activity against a lysate of the H. pylori Sydney strain in enzyme-linked immunosorbent assay (ELISA) and 9% bound the H. pylori urease. Further characterization by PCR-fingerprint analysis and sequencing revealed that two closely related H. pylori binders and one antiurease scFv could be isolated. The selected scFvs were highly specific as analyzed by ELISA and immunoblots using various bacterial lysates and recombinant proteins. Analysis of the humoral immune response following H. pylori infection using human monoclonal antibodies might contribute to a better understanding of the pathogenesis of the disease. Moreover, using immune phage display libraries, it might be possible for relevant epitopes of H. pylori antigens to be determined, which might be of use for vaccine development. PMID- 12117925 TI - Role of Yops and adhesins in resistance of Yersinia enterocolitica to phagocytosis. AB - Yersinia enterocolitica is a pathogen endowed with two adhesins, Inv and YadA, and with the Ysc type III secretion system, which allows extracellular adherent bacteria to inject Yop effectors into the cytosol of animal target cells. We tested the influence of all of these virulence determinants on opsonic and nonopsonic phagocytosis by PU5-1.8 and J774 mouse macrophages, as well as by human polymorphonuclear leukocytes (PMNs). The adhesins contributed to phagocytosis in the absence of opsonins but not in the presence of opsonins. In agreement with previous results, YadA counteracted opsonization. In every instance, the Ysc-Yop system conferred a significant level of resistance to phagocytosis. Nonopsonized single-mutant bacteria lacking either YopE, -H, -T, or -O were phagocytosed significantly more by J774 cells and by PMNs. Opsonized bacteria were phagocytosed more than nonopsonized bacteria, and mutant bacteria lacking either YopH, -T, or -O were phagocytosed significantly more by J774 cells and by PMNs than were wild-type (WT) bacteria. Opsonized mutants lacking only YopE were phagocytosed significantly more than were WT bacteria by PMNs but not by J774 cells. Thus, YopH, -T, and -O were involved in all of the phagocytic processes studied here but YopE did not play a clear role in guarding against opsonic phagocytosis by J774. Mutants lacking YopP and YopM were, in every instance, as resistant as WT bacteria. Overexpression of YopE, -H, -T, or -O alone did not confer resistance to phagocytosis, although it affected the cytoskeleton. These results show that YopH, YopT, YopO, and, in some instances, YopE act synergistically to increase the resistance of Y. enterocolitica to phagocytosis by macrophages and PMNs. PMID- 12117926 TI - Leishmania promastigotes release a granulocyte chemotactic factor and induce interleukin-8 release but inhibit gamma interferon-inducible protein 10 production by neutrophil granulocytes. AB - Recent data from our laboratory suggest that neutrophil granulocytes (polymorphonuclear leukocytes [PMN]) can serve as host cells for Leishmania major in the early phase of infection. In line with these findings, an early influx of PMN to the infected tissues was shown by others to be associated with susceptibility to infection with L. major. The mechanisms underlying the initial PMN recruitment to the site of infection is poorly understood. In the present study we investigated whether Leishmania can influence PMN migration. Supernatants of Leishmania promastigotes were tested for their chemotactic activity using an in vitro chemotaxis assay. All Leishmania species tested (L. major, L. aethiopica, and L. donovani) displayed a marked chemotactic effect on human PMN. However, no effect on the migration of macrophages and NK cells was observed. Checkerboard analysis revealed that the observed PMN migration was due to chemotaxis rather than chemokinesis. Most of the chemotactic activity was found in fractions containing molecules with sizes between 10 and 50 kDa. Pretreatment of PMN with N-formyl-methionyl-leucyl-phenylalanine blocked the chemotactic activity of Leishmania supernatants up to 75%. In addition, we found that leishmanial contact induced the release of interleukin-8 (IL-8) and inhibited the production of gamma interferon-inducible protein 10 (IP-10) by PMN. These data suggest that infection with Leishmania promastigotes leads to PMN accumulation via the production of a chemotactic factor by the parasites, and this effect is amplified by the induction of IL-8 production in PMN. On the other hand, the inhibition of IP-10 production can lead to prevention of NK cell activation. PMID- 12117928 TI - Evidence that the variable regions of the central domain of VlsE are antigenic during infection with lyme disease spirochetes. AB - It has been postulated that the vls system of the Lyme disease spirochetes contributes to immune evasion through antigenic variation. While it is clear that vlsE undergoes sequence change within its variable regions at a high frequency during the early stages of infection, a definitive role in immune evasion has not been demonstrated. In this report we assessed the murine and human humoral immune response to recombinant (r)-VlsE variants that originally arose during infection in mice. Immunoblot analyses of r-VlsE variants were conducted by using serum samples collected from mice infected with Borrelia burgdorferi clones that carried different vlsE variants. All of the r-VlsE variants were recognized by infection sera regardless of the identity of the infecting clone or isolate. In addition, all variants were immunoreactive with a panel of human Lyme disease patient serum samples. It is evident from these analyses that the infection induced VlsE variants share common epitopes that reside within conserved segments of these proteins. However, preabsorption experiments revealed that the variable regions of the central domain of VlsE, which undergo rapid mutation during infection, also influence the antigenic properties of the protein. A subset of the antibodies elicited against vlsE variants that differ in the sequences of their variable regions were found to be variant specific. Hence, in spite of a robust antibody response to conserved segments of VlsE, infection-induced sequence changes within the variable regions alter the antigenicity of VlsE. These results provide the first direct evidence of antigenic variation in the VlsE protein. PMID- 12117927 TI - Phenotypes of a naturally defective recB allele in Neisseria meningitidis clinical isolates. AB - Neisseria meningitidis strains belonging to the hypervirulent lineage ET-37 and several unrelated strains are extremely UV sensitive. The phenotype is consequent to the presence of a nonfunctional recB(ET-37) allele carrying multiple missense mutations. Phenotypic analysis has been performed with congenic meningococcal strains harboring either the wild-type recB allele or the recB(ET-37) allele. Congenic recB(ET-37) meningococci, in addition to being sensitive to UV, were defective both in repair of DNA lesions induced by UV treatment and, partially, in recombination-mediated transformation. Consistently, the wild-type, but not the recB(ET-37), allele was able to complement the Escherichia coli recB21 mutation to UV resistance and proficiency in recombination. recB(ET-37) meningococci did not exhibit higher frequencies of spontaneous mutation to rifampin resistance than recB-proficient strains. However, mutation rates were enhanced following UV treatment, a phenomenon not observed in the recB-proficient counterpart. Interestingly, the results of PCR-based assays demonstrated that the presence of the recB(ET-37) allele considerably increased the frequency of recombination at the pilin loci. The main conclusion that can be drawn is that the presence of the defective recB(ET-37) allele in N. meningitidis isolates causes an increase in genetic diversity, due to an ineffective RecBCD-dependent DNA repair and recombination pathway, and an increase in pilin antigenic variation. PMID- 12117929 TI - Yersiniabactin production requires the thioesterase domain of HMWP2 and YbtD, a putative phosphopantetheinylate transferase. AB - One requirement for the pathogenesis of Yersinia pestis, the causative agent of bubonic plague, is the yersiniabactin (Ybt) siderophore-dependent iron transport system that is encoded within a high-pathogenicity island (HPI) within the pgm locus of the Y. pestis chromosome. Nine gene products within the HPI have demonstrated functions in the nonribosomal peptide synthesis (NRPS)/polyketide (PK) synthesis or transport of Ybt. NRPS/PK synthetase or synthase enzymes are generally activated by phosphopantetheinylation. However, no products with similarities to known phosphopantetheinyl (P-pant) transferases were found within the pgm locus. We have identified a gene, ybtD, encoded outside the HPI and pgm locus, that is necessary for function of the Ybt system and has similarities to other P-pant transferases such as EntD of Escherichia coli. A deletion within ybtD yielded a strain (KIM6-2085+) defective in siderophore production. This strain was unable to grow on iron-deficient media at 37 degrees C but could be cross-fed by culture supernatants from Ybt-producing strains of Y. pestis. The promoter region of ybtD was fused to lacZ; beta-galactosidase expression from this reporter was not regulated by the iron status of the bacterial cells or by YbtA, a positive regulator of other genes of the ybt system. The ybtD mutant failed to express indicator Ybt proteins (high-molecular-weight protein 1 [HMWP1], HMWP2, and Psn), a pattern similar to those seen with several other ybt biosynthetic mutants. In contrast, cells containing a single amino acid substitution (S2908A) in the terminal thioesterase domain of HMWP2 failed to exhibit any ybt regulatory defects but did not elaborate extracellular Ybt under iron-deficient conditions. PMID- 12117931 TI - Pathogenesis of pneumococcal pneumonia in cyclophosphamide-induced leukopenia in mice. AB - Streptococcus pneumoniae pneumonia frequently occurs in leukopenic hosts, and most patients subsequently develop lung injury and septicemia. However, few correlations have been made so far between microbial growth, inflammation, and histopathology of pneumonia in specific leukopenic states. In the present study, the pathogenesis of pneumococcal pneumonia was investigated in mice rendered leukopenic by the immunosuppressor antineoplastic drug cyclophosphamide. Compared to the immunocompetent state, cyclophosphamide-induced leukopenia did not hamper interleukin-1 (IL-1), IL-6, macrophage inflammatory protein-1 (MIP-1), MIP-2, and monocyte chemotactic protein-1 secretion in infected lungs. Leukopenia did not facilitate bacterial dissemination into the bloodstream despite enhanced bacterial proliferation into lung tissues. Pulmonary capillary permeability and edema as well as lung injury were enhanced in leukopenic mice despite the absence of neutrophilic and monocytic infiltration into their lungs, suggesting an important role for bacterial virulence factors and making obvious the fact that neutrophils are ultimately not required for lung injury in this model. Scanning and transmission electron microscopy revealed extensive disruption of alveolar epithelium and a defect in surfactant production, which were associated with alveolar collapse, hemorrhage, and fibrin deposits in alveoli. These results contrast with those observed in immunocompetent animals and indicate that leukopenic hosts suffering from pneumococcal pneumonia are at a higher risk of developing diffuse alveolar damage. PMID- 12117930 TI - Immunization with a polyprotein vaccine consisting of the T-Cell antigens thiol specific antioxidant, Leishmania major stress-inducible protein 1, and Leishmania elongation initiation factor protects against leishmaniasis. AB - Development of an effective vaccine against Leishmania infection is a priority of tropical disease research. We have recently demonstrated protection against Leishmania major in the murine and nonhuman primate models with individual or combinations of purified leishmanial recombinant antigens delivered as plasmid DNA constructs or formulated with recombinant interleukin-12 (IL-12) as adjuvant. In the present study, we immunized BALB/c mice with a recombinant polyprotein comprising a tandem fusion of the leishmanial antigens thiol-specific antioxidant, L. major stress-inducible protein 1 (LmSTI1), and Leishmania elongation initiation factor (LeIF) delivered with adjuvants suitable for human use. Aspects of the safety, immunogenicity, and vaccine efficacy of formulations with each individual component, as well as the polyprotein referred to as Leish 111f, were assessed by using the L. major challenge model with BALB/c mice. No adverse reactions were observed when three subcutaneous injections of the Leish 111f polyprotein formulated with either MPL-squalene (SE) or Ribi 529-SE were given to BALB/c mice. A predominant Th1 immune response characterized by in vitro lymphocyte proliferation, gamma interferon production, and immunoglobulin G2A antibodies was observed with little, if any, IL-4. Moreover, Leish-111f formulated with MPL-SE conferred immunity to leishmaniasis for at least 3 months. These data demonstrate success at designing and developing a prophylactic leishmaniasis vaccine that proved effective in a preclinical model using multiple leishmanial antigens produced as a single protein delivered with a powerful Th1 adjuvant suitable for human use. PMID- 12117932 TI - Variation in extracellular protease production among clinical isolates of Staphylococcus aureus due to different levels of expression of the protease repressor sarA. AB - Staphylococcus aureus produces four major extracellular proteases: staphylococcal serine protease (V8 protease; SspA), cysteine protease (SspB), metalloprotease (aureolysin; Aur), and staphopain (Scp). Several in vitro studies have suggested that these enzymes are important virulence factors. Here we analyzed the protease production of 92 S. aureus strains from infected human soft tissue. Twenty-one strains produced variable zones of proteolysis on casein agar plates, while the remaining 71 strains appeared to be protease negative. The major protease genes were present in all protease-positive (n = 5) and protease-negative (n = 12) strains analyzed. Northern blotting showed that transcription of the protease genes was suppressed due to increased sigma factor B (SigB)-dependent expression of the protease repressor SarA. Other SigB-dependent traits such as pigmentation and expression of asp 23 were also increased in protease-negative compared to protease-positive strains. Inactivation of sarA in three protease-negative strains resulted in increased transcription of all protease genes and increased protease production, while overexpression of sarA in a strain producing protease at high levels repressed protease production. Our results suggest that the protease genes are conserved among clinical S. aureus strains and that the level of SigB-dependent expression of the protease repressor sarA determines the level of protease production in each strain. PMID- 12117933 TI - Leukotriene B(4) induces nitric oxide synthesis in Trypanosoma cruzi-infected murine macrophages and mediates resistance to infection. AB - The production of nitric oxide (NO) by gamma interferon (IFN-gamma)-activated macrophages is a major effector mechanism during experimental Trypanosoma cruzi infection. In addition to IFN-gamma, chemoattractant molecules, such as platelet activating factor (PAF) and CC chemokines, may also activate macrophages to induce NO and mediate the killing of T. cruzi in an NO-dependent manner. Here we investigated the ability of leukotriene B(4) (LTB(4)) to induce the production of NO by macrophages infected with T. cruzi in vitro and whether NO mediated LTB(4) induced parasite killing. The activation of T. cruzi-infected but not naive murine peritoneal macrophages with LTB(4) induced the time- and concentration dependent production of NO. In addition, low concentrations of LTB(4) acted in synergy with IFN-gamma to induce NO production. The NO produced mediated LTB(4) induced microbicidal activity in macrophages, as demonstrated by the inhibitory effects of an inducible NO synthase inhibitor. LTB(4)-induced NO production and parasite killing were LTB(4) receptor dependent and were partially blocked by a PAF receptor antagonist. LTB(4) also induced significant tumor necrosis factor alpha (TNF-alpha) production, and blockade of TNF-alpha suppressed LTB(4)-induced NO release and parasite killing. A blockade of LTB(4) or PAF receptors partially inhibited IFN-gamma-induced NO and TNF-alpha production but not parasite killing. Finally, daily treatment of infected mice with CP-105,696 was accompanied by a significantly higher level of blood parasitemia, but not lethality, than that seen in vehicle-treated animals. In conclusion, our results suggest a role for LTB(4) during experimental T. cruzi infection. Chemoattractant molecules such as LTB(4) not only may play a major role in leukocyte migration into sites of inflammation in vivo but also, in the event of an infection, may play a relevant role in the activation of recruited leukocytes to kill the invading microorganism in an NO-dependent manner. PMID- 12117934 TI - Induction of cell-mediated immunity to Staphylococcus aureus in the mouse mammary gland by local immunization with a live attenuated mutant. AB - The efficacy of intramammary (Ima) immunization with a live attenuated (la) Staphylococcus aureus mutant to protect the mouse mammary gland from infection has previously been established. The present study was aimed at evaluating whether Ima immunization with la-S. aureus can induce cell-mediated immune responses to the pathogen within the mammary gland. Mice were immunized by Ima route with la-S. aureus, and regional lymph node mononuclear cells were obtained thereafter. A higher expression of the interleukin-2 receptor was found on B and T cells from immunized mice when they were compared with control mice. Immunization with la-S. aureus induced strong proliferative responses to S. aureus. Moreover, significantly increased levels of gamma interferon (IFN-gamma) were produced by CD4+ T cells when lymphocytes from immunized mice, but not from control mice, were cultured in the presence of staphylococcal antigens. Moreover, a significant increase in the percentage of IFN-gamma-producing CD4+ and CD8+ T cells was observed after S. aureus Ima challenge in immunized mice compared to challenged control mice. Our results demonstrated that Ima immunization with la S. aureus induced primed lymphocyte populations capable of responding against staphylococcal antigens during in vitro stimulation, as well as during in vivo infection by S. aureus. CD4+ and CD8+ T cells appear to be the main lymphocyte subpopulations involved in this response. It is suggested that IFN-gamma production induced by Ima immunization may play a pivotal role in the eradication of intracellular staphylococci. PMID- 12117935 TI - Organization of the plasmid cpe Locus in Clostridium perfringens type A isolates. AB - Clostridium perfringens type A isolates causing food poisoning have a chromosomal enterotoxin gene (cpe), while C. perfringens type A isolates responsible for non food-borne human gastrointestinal diseases carry a plasmid cpe gene. In the present study, the plasmid cpe locus of the type A non-food-borne-disease isolate F4969 was sequenced to design primers and probes for comparative PCR and Southern blot studies of the cpe locus in other type A isolates. Those analyses determined that the region upstream of the plasmid cpe gene is highly conserved among type A isolates carrying a cpe plasmid. The organization of the type A plasmid cpe locus was also found to be unique, as it contains IS1469 sequences located similarly to those in the chromosomal cpe locus but lacks the IS1470 sequences found upstream of IS1469 in the chromosomal cpe locus. Instead of those upstream IS1470 sequences, a partial open reading frame potentially encoding cytosine methylase (dcm) was identified upstream of IS1469 in the plasmid cpe locus of all type A isolates tested. Similar dcm sequences were also detected in several cpe-negative C. perfringens isolates carrying plasmids but not in type A isolates carrying a chromosomal cpe gene. Contrary to previous reports, sequences homologous to IS1470, rather than IS1151, were found downstream of the plasmid cpe gene in most type A isolates tested. Those IS1470-like sequences reside in about the same position but are oppositely oriented and defective relative to the IS1470 sequences found downstream of the chromosomal cpe gene. Collectively, these and previous results suggest that the cpe plasmid of many type A isolates originated from integration of a cpe-containing genetic element near the dcm sequences of a C. perfringens plasmid. The similarity of the plasmid cpe locus in many type A isolates is consistent with horizontal transfer of a common cpe plasmid among C. perfringens type A strains. PMID- 12117936 TI - Fimbriated Salmonella enterica serovar typhimurium abates initial inflammatory responses by macrophages. AB - Oral immunization of mice with a Salmonella vaccine expressing colonization factor antigen I (CFA/I) from enterotoxigenic Escherichia coli results in the rapid onset of interleukin-4 (IL-4) and IL-5 production, which explains the observed elevations in mucosal immunoglobulin A (IgA) and serum IgG1 antibodies. In contrast, oral immunization with the Salmonella vector does not result in the production of Th2-type cytokines. To begin to assess why such differences exist between the two strains, it should be noted that in vitro infection of RAW 264.7 macrophages resulted in the absence of nitric oxide (NO) production in cells infected with the Salmonella-CFA/I vaccine. This observation suggests differential proinflammatory cytokine production by these isogenic Salmonella strains. Upon measurement of proinflammatory cytokines, minimal to no tumor necrosis factor alpha (TNF-alpha), IL-1alpha, IL-1beta, or IL-6 was produced by Salmonella-CFA/I-infected RAW 264.7 or peritoneal macrophages, but production was greatly induced in Salmonella vector-infected macrophages. Only minute levels of IL-12 p70 were induced by Salmonella vector-infected macrophages, and none was induced by Salmonella-CFA/I-infected macrophages. The absence of IL-12 was not due to overt increases in production of either IL-12 p40 or IL-10. CFU measurements taken at 8 h postinfection showed no differences in colonization in RAW 264.7 cells infected with either Salmonella construct, but there were differences in peritoneal macrophages. However, after 24 h, the Salmonella vector strain colonized to a greater extent in RAW 264.7 cells than in peritoneal macrophages. Infection of RAW 264.7 cells or peritoneal macrophages with either Salmonella construct showed no difference in macrophage viabilities. This evidence shows that the expression of CFA/I fimbriae alters how macrophages recognize or process salmonellae and prevents the rapid onset of proinflammatory cytokines which is typical during Salmonella infections. PMID- 12117938 TI - Comparison of alteration of cell surface carbohydrates of the chinchilla tubotympanum and colonial opacity phenotype of Streptococcus pneumoniae during experimental pneumococcal otitis media with or without an antecedent influenza A virus infection. AB - Experimental and clinical studies suggest that influenza A virus promotes Streptococcus pneumoniae-induced otitis media; however, the mechanism underlying this synergistic interaction has not been completely defined. In this study, glycoconjugate expression patterns were evaluated on the cell surface in the chinchilla eustachian tube (ET) lumen of a cohort challenged intranasally (i.n.) with S. pneumoniae type 6A, which is predominantly transparent and a cohort with an antecedent influenza A virus infection, followed by i.n. inoculation with S. pneumoniae. The labeling patterns obtained with six lectin probes revealed that the binding of Bandeiraea simplicifolia lectin II, succinylated wheat germ agglutinin, and peanut agglutinin were significantly increased in the lumenal surface of the ET in the cohort infected with both pathogens compared to the cohort inoculated with only S. pneumoniae, which indicated that N acetylglucosamine (GlcNAc) and D-galactose residues were exposed. A significant decreased labeling with Sambucus nigra agglutinin in the combined influenza A virus and pneumococcus infection cohort suggested that there were few sialic acid residues remaining in the ET epithelium. In addition, the colonial opacity of S. pneumoniae during the disease course was examined. The opaque phenotype was predominant among the pneumococcus isolates from the middle-ear fluid in the cohort infected with the both pathogens. Together, these data suggest that the synergic effect of influenza A virus and S. pneumoniae on the changes of the carbohydrate moieties in the ET epithelium and that the selection of the opaque variant may facilitate the pneumococcal invasion of the middle ear. PMID- 12117937 TI - One of two copies of the gene for the activatable shiga toxin type 2d in Escherichia coli O91:H21 strain B2F1 is associated with an inducible bacteriophage. AB - Shiga toxin (Stx) types 1 and 2 are encoded within intact or defective temperate bacteriophages in Stx-producing Escherichia coli (STEC), and expression of these toxins is linked to bacteriophage induction. Among Stx2 variants, only stx(2e) from one human STEC isolate has been reported to be carried within a toxin converting phage. In this study, we examined the O91:H21 STEC isolate B2F1, which carries two functional alleles for the potent activatable Stx2 variant toxin, Stx2d, for the presence of Stx2d-converting bacteriophages. We first constructed mutants of B2F1 that produced one or the other Stx2d toxin and found that the mutant that produced only Stx2d1 made less toxin than the Stx2d2-producing mutant. Consistent with that result, the Stx2d1-producing mutant was attenuated in a streptomycin-treated mouse model of STEC infection. When the mutants were treated with mitomycin C to promote bacteriophage induction, Vero cell cytotoxicity was elevated only in extracts of the Stx2d1-producing mutant. Additionally, when mice were treated with ciprofloxacin, an antibiotic that induces the O157:H7 Stx2-converting phage, the animals were more susceptible to the Stx2d1-producing mutant. Moreover, an stx(2d1)-containing lysogen was isolated from plaques on strain DH5alpha that had been exposed to lysates of the mutant that produced Stx2d1 only, and supernatants from that lysogen transformed with a plasmid encoding RecA were cytotoxic when the lysogen was induced with mitomycin C. Finally, electron-microscopic examination of extracts from the Stx2d1-producing mutant showed hexagonal particles that resemble the prototypic Stx2-converting phage 933W. Together these observations provide strong evidence that expression of Stx2d1 is bacteriophage associated. We conclude that despite the sequence similarity of the stx(2d1)- and stx(2d2)-flanking regions in B2F1, Stx2d1 expression is repressed within the context of its toxin-converting phage while Stx2d2 expression is independent of phage induction. PMID- 12117939 TI - Identification of an aggregative adhesion fimbria (AAF) type III-encoding operon in enteroaggregative Escherichia coli as a sensitive probe for detecting the AAF encoding operon family. AB - Enteroaggregative Escherichia coli (EAEC) is recognized as an emerging cause of diarrhea in children and adults worldwide, and recent studies have implicated EAEC in persistent diarrhea in patients infected with human immunodeficiency virus (HIV). In this study, we identified aggregative adhesion fimbria type III (AAF-III) in isolate 55989, a typical EAEC strain. Analysis of the sequence of the plasmid-borne agg-3 gene cluster encoding AAF-III showed this cluster to be closely related to the agg and aaf operons and to the afa operons carried by diffusely adherent pathogenic E. coli. We investigated the adhesion properties of a collection of 25 EAEC strains isolated from HIV-infected patients presenting with persistent diarrhea. We found that a minority of strains (36%) carried sequences similar to those of the agg and aaf operons, which encode AAF-I and AAF II, respectively. We developed PCR assays specific for the agg-3 operon. In our collection, the frequency of AAF-III strains was similar (12%) to that of AAF-I strains (16%) but higher than that of AAF-II isolates (0%). Differences between EAEC strains in terms of the virulence factors present render detection of these strains difficult with the available DNA probes. Based on comparison of the agg, aaf, and agg-3 operons, we defined an AAF probe internal to the adhesion gene clusters and demonstrated that it was efficient for the identification of EAEC strains. We investigated 32 EAEC isolates, of which only 34.4% were detected with the classical CVD432 probe (detecting pAA virulence plasmids) whereas 65.6% were detected with the AAF probe. PMID- 12117941 TI - Contribution of phospholipomannan to the surface expression of beta-1,2 oligomannosides in Candida albicans and its presence in cell wall extracts. AB - beta-1,2-Oligomannosides (beta-1,2-Man) derived from Candida albicans mannan have been shown to act as adhesins and to induce protective antibodies. We used monoclonal antibodies specific for beta-1,2-Man in electron, confocal, and fluorescence microscopy to study the surface expression of beta-1,2-Man epitopes. These monoclonal antibodies were also used for Western blotting of cell surface extracts to study the nature of the molecules expressing the beta-Man epitopes. Evidence was obtained for the contribution of a glycolipid, phospholipomannan (PLM), to the complex expression of beta-1,2-Man epitopes at the cell wall surfaces of yeasts grown on solid media. PLM was present in intercellular matrixes of colonies grown on agar and was detected as a contaminant in mannan batches prepared by conventional methods. PMID- 12117940 TI - Streptococcus suis interactions with the murine macrophage cell line J774: adhesion and cytotoxicity. AB - Streptococcus suis capsular type 2 is an important etiological agent of swine meningitis, and it is also a zoonotic agent. Since one hypothesis of the pathogenesis of S. suis infection is that bacteria enter the bloodstream and invade the meninges and other tissues in close association with mononuclear phagocytes, the objective of the present study was to evaluate the capacity of S. suis type 2 to adhere to macrophages. An enzyme-linked immunosorbent assay technique was standardized to simply and accurately measure the rate of bacterial attachment to phagocytic cells. Results were confirmed by plate counting. Adhesion was dependent on bacterial concentration and incubation time and was not affected by cytochalasin pretreatment of macrophages. Inhibition studies showed that the sialic acid moiety of the S. suis capsule would be, at least in part, responsible for bacterial recognition by macrophages. Serum preopsonization of bacteria increased adhesion levels. Complement would be partially implicated in the serum-enhanced binding of S. suis to cells. Adhesion varied among different S. suis type 2 isolates. However, high bacterial concentrations of several isolates were cytotoxic for cells, and these cytotoxic effects correlated with suilysin production. Indeed, hemolytic strain supernatants, as well as purified suilysin, reproduced cytotoxic effects observed with live bacteria, and these effects were inhibited by cholesterol pretreatment. Bacterial adhesion and cytotoxicity were confirmed by scanning and transmission electron microscopy. We hypothesize that attachment of bacteria to phagocytes could play an important role in the pathogenesis of S. suis infection by allowing bacterial dissemination and causing a bacteremia and/or septicemia. This interaction could also be related to the activation of the host inflammatory response observed during meningitis. PMID- 12117942 TI - Protection of rhesus macaques against lethal Plasmodium knowlesi malaria by a heterologous DNA priming and poxvirus boosting immunization regimen. AB - We tested a cytokine-enhanced, multiantigen, DNA priming and poxvirus boosting vaccine regimen for prevention of malaria in the Plasmodium knowlesi-rhesus macaque model system. Animals were primed with a mixture of DNA plasmids encoding two preerythrocytic-stage proteins and two erythrocytic-stage proteins from P. knowlesi and combinations of the cytokines granulocyte-macrophage colony stimulating factor, interleukin-4, and tumor necrosis factor alpha and were boosted with a mixture of four recombinant, attenuated vaccinia virus strains encoding the four P. knowlesi antigens. Two weeks after boosting, the geometric mean immunofluorescence titers in the immunized groups against sporozoites and infected erythrocytes ranged from 160 to 8,096 and from 1,810 to 5,120, respectively. The geometric mean anti-P. knowlesi circumsporozoite protein (PkCSP) titers ranged from 1,761 to 24,242. Peripheral blood mononuclear cells (PBMC) from the immunized monkeys produced gamma interferon (IFN-gamma) in response to incubation with pooled peptides from the PkCSP at frequencies of 10 to 571 spot-forming cells/10(6) PBMC. Following challenge with 100 infectious P. knowlesi sporozoites, 2 of 11 immunized monkeys were sterilely protected, and 7 of the 9 infected monkeys resolved their parasitemias spontaneously. In contrast, all four controls became infected and required treatment for overwhelming parasitemia. Early protection was strongly associated with IFN-gamma responses against a pool of peptides from the preerythrocytic-stage antigen, PkCSP. These findings demonstrate that a multistage, multiantigen, DNA priming and poxvirus boosting vaccine regimen can protect nonhuman primates from an otherwise lethal malaria sporozoite challenge. PMID- 12117943 TI - Inflammatory cytokines enhance the interaction of Mannheimia haemolytica leukotoxin with bovine peripheral blood neutrophils in vitro. AB - Mannheimia (Pasteurella) haemolytica A1 produces several virulence factors that play an important role in the pathogenesis of bovine pneumonic pasteurellosis. Foremost among these is a leukotoxin (LKT) that specifically kills ruminant leukocytes. Recent evidence suggests that M. haemolytica LKT binding to bovine leukocytes is mediated by the beta(2)-integrin CD11a/CD18 (lymphocyte function associated antigen 1 [LFA-1]), which subsequently induces activation and cytolysis of these cells. Inflammatory cytokines, which are released during viral and bacterial infection, are reported to increase LFA-1 expression and conformational activation. We investigated the effects of the inflammatory cytokines interleukin-1beta (IL-1beta), tumor necrosis factor alpha (TNF-alpha), and gamma interferon (IFN-gamma) on the interaction of M. haemolytica LKT with bovine peripheral blood neutrophils (PMNs). In this study we demonstrated, by flow cytometry, that bovine PMNs increased their binding to an anti-bovine LFA-1 monoclonal antibody (BAT75A) following in vitro incubation with IL-1beta, TNF alpha, or IFN-gamma. Incubation with cytokines also increased CD18 expression, as assessed by real-time PCR and by Western blotting. Increased LFA-1 expression by PMNs exposed to cytokines was associated with increased LKT binding and cytotoxicity. The latter represented, at least in part, enhanced PMN apoptosis, as assessed by propidium iodine staining and caspase-3 activation. The results of this study suggest that inflammatory cytokines may play an important role in enhancing the biological response of bovine PMNs to M. haemolytica LKT. PMID- 12117944 TI - Differential expression of virulence-related genes in Enterococcus faecalis in response to biological cues in serum and urine. AB - Enterococci rank among leading causes of nosocomial bacteremia and urinary tract infection and are also a leading cause of community acquired subacute endocarditis. Limited evidence suggests that biological cues in serum and urine may play an important role in modulating enterococcal virulence at sites of infection. To determine the extent to which biological cues affect enterococcal virulence-associated gene expression, we used quantitative real-time PCR to compare mRNA levels in Enterococcus faecalis cultures grown in serum or urine to that achieved in laboratory medium. Both environment- and growth phase-specific variations were observed, demonstrating the occurrence of as-yet-uncharacterized mechanisms for control of gene expression in E. faecalis that may play an important role in vivo. PMID- 12117946 TI - Tissue tropism of enteropathogenic Escherichia coli strains belonging to the O55 serogroup. AB - Four enteropathogenic Escherichia coli (EPEC) strains belonging to the O55 serogroup (G21 and G30 [both O55:H6], G35 [O55:H-], and G58 [O55:H7]) were tested for their tissue tropism by using human intestinal in vitro organ culture. Strains showed restricted adhesion with attaching-and-effacing activity to follicle-associated epithelium of Peyer's patches, with no apparent adhesion to duodenum or colon. G35 and G58 express intimin gamma and show a similar tropism to intimin gamma-expressing enterohemorrhagic E. coli (EHEC) O157:H7. However, strains G21 and G30 were unusual because they expressed intimin alpha and had a restricted tissue tropism of intimin gamma phenotype. The amino acid sequence of the carboxy-terminal 280 amino acids of intimin from G21 was determined. Comparison with the prototype intimin alpha from strain E2348/69 (O127:H6) showed a single amino acid difference (corresponding to Val907 and Ala907 in the whole intimins). This mutation was reproduced by site-directed mutagenesis in an intimin alpha plasmid template, pCVD438, with the hypothesis that it may induce a change in tropism. However, when the mutated plasmid was placed in both EPEC and EHEC backgrounds, duodenal adhesion in a manner similar to strain E2348/69 was evident upon in vitro organ culture. Thus, additional factor(s) unrelated to intimin exist in the O55:H6 genome that influence human intestinal tissue tropism. PMID- 12117945 TI - Clostridium septicum alpha-toxin is active against the parasitic protozoan Toxoplasma gondii and targets members of the SAG family of glycosylphosphatidylinositol-anchored surface proteins. AB - As is the case with many other protozoan parasites, glycosylphosphatidylinositol (GPI)-anchored proteins dominate the surface of Toxoplasma gondii tachyzoites. The mechanisms by which T. gondii GPI-anchored proteins are synthesized and transported through the unusual triple-membrane structure of the parasite pellicle to the plasma membrane remain largely unknown. As a first step in developing tools to study these processes, we show here that Clostridium septicum alpha-toxin, a pore-forming toxin that targets GPI-anchored protein receptors on the surface of mammalian cells, is active against T. gondii tachyzoites (50% effective concentration, 0.2 nM). Ultrastructural studies reveal that a tight physical connection between the plasma membrane and the underlying membranes of the inner membrane complex is locally disrupted by toxin treatment, resulting in a massive outward extension of the plasma membrane and ultimately lysis of the parasite. Toxin treatment also causes swelling of the parasite endoplasmic reticulum, providing the first direct evidence that alpha-toxin is a vacuolating toxin. Alpha-toxin binds to several parasite GPI-anchored proteins, including surface antigen 3 (SAG3) and SAG1. Interestingly, differences in the toxin binding profiles between the virulent RH and avirulent P strain were observed. Alpha-toxin may prove to be a powerful experimental tool for molecular genetic analysis of GPI anchor biosynthesis and GPI-anchored protein trafficking in T. gondii and other susceptible protozoa. PMID- 12117947 TI - Effect of acid adaptation on the fate of Listeria monocytogenes in THP-1 human macrophages activated by gamma interferon. AB - In Listeria monocytogenes the acid tolerance response (ATR) takes place through a programmed molecular response which ensures cell survival under unfavorable conditions. Much evidence links ATR with virulence, but the molecular determinants involved in the reactivity to low pHs and the behavior of acid exposed bacteria within host cells are still poorly understood. We have investigated the effect of acid adaptation on the fate of L. monocytogenes in human macrophages. Expression of genes encoding determinants for cell invasion and intracellular survival was tested for acid-exposed bacteria, and invasive behavior in the human myelomonocytic cell line THP-1 activated with gamma interferon was assessed. Functional approaches demonstrated that preexposure to an acidic pH enhances the survival of L. monocytogenes in activated human macrophages and that this effect is associated with an altered pattern of expression of genes involved in acid resistance and cell invasion. Significantly decreased transcription of the plcA gene, encoding a phospholipase C involved in vacuolar escape and cell-to-cell spread, was observed in acid-adapted bacteria. This effect was due to a reduction in the quantity of the bicistronic plcA-prfA transcript, concomitant with an increase in the level(s) of the monocistronic prfA mRNA(s). The transcriptional shift from distal to proximal prfA promoters resulted in equal levels of the prfA transcript (and, as a consequence, of the inlA, hly, and actA transcripts) under neutral and acidic conditions. In contrast, the sodC and gad genes, encoding a cytoplasmic superoxide dismutase and the glutamate-based acid resistance system, respectively, were positively regulated at a low pH. Morphological approaches confirmed the increased intracellular survival and growth of acid-adapted L. monocytogenes cells both in vacuoles and in the cytoplasm of interferon gamma-activated THP-1 macrophages. Our data indicate that preexposure to a low pH has a positive impact on subsequent challenge of L. monocytogenes with macrophagic cells. PMID- 12117948 TI - Identification of chromosomal Shigella flexneri genes induced by the eukaryotic intracellular environment. AB - Upon entry into the eukaryotic cytosol, the facultative intracellular bacterium Shigella flexneri is exposed to an environment that may necessitate the expression of particular genes for it to survive and grow intracellularly. To identify genes that are induced in response to the intracellular environment, we screened a library containing fragments of the S. flexneri chromosome fused to a promoterless green fluorescent protein gene (gfp). Bacteria containing promoter fusions that had a higher level of gfp expression when S. flexneri was intracellular (in Henle cells) than when S. flexneri was extracellular (in Luria Bertani broth) were isolated by using fluorescence-activated cell sorting. Nine different genes with increased expression in Henle cells were identified. Several genes (uhpT, bioA, and lysA) were involved in metabolic processes. The uhpT gene, which encoded a sugar phosphate transporter, was the most frequently isolated gene and was induced by glucose-6-phosphate in vitro. Two of the intracellularly induced genes (pstS and phoA) encode proteins involved in phosphate acquisition and were induced by phosphate limitation in vitro. Additionally, three iron regulated genes (sufA, sitA, and fhuA) were identified. The sufA promoter was derepressed in iron-limiting media and was also induced by oxidative stress. To determine whether intracellularly induced genes are required for survival or growth in the intracellular environment, we constructed mutations in the S. flexneri uhpT and pstS genes by allelic exchange. The uhpT mutant could not use glucose-6-phosphate as a sole carbon source in vitro but exhibited normal plaque formation on Henle cell monolayers. The pstS mutant had no apparent growth defect in low-phosphate media in vitro but formed smaller plaques on Henle cell monolayers than the parent strain. Both mutants were as effective as the parent strain in inducing apoptosis in a macrophage cell line. PMID- 12117949 TI - Characterization of pit, a Streptococcus pneumoniae iron uptake ABC transporter. AB - Bacteria frequently have multiple mechanisms for acquiring iron, an essential micronutrient, from the environment. We have identified a four-gene Streptococcus pneumoniae operon, named pit, encoding proteins with similarity to components of a putative Brachyspira hyodysenteriae iron uptake ABC transporter, Bit. An S. pneumoniae strain containing a defined mutation in pit has impaired growth in medium containing the iron chelator ethylenediamine di-o-hydroxyphenylacetic acid, reduced sensitivity to the iron-dependent antibiotic streptonigrin, and impaired virulence in a mouse model of S. pneumoniae systemic infection. Furthermore, addition of a mutation in pit to a strain containing mutations in the two previously described S. pneumoniae iron uptake ABC transporters, piu and pia, resulted in a strain with impaired growth in two types of iron-deficient medium, a high degree of resistance to streptonigrin, and a reduced rate of iron uptake. Comparison of the susceptibilities to streptonigrin of the individual pit, piu, and pia mutant strains and comparison of the growth in iron-deficient medium and virulence of single and double mutant strains suggest that pia is the dominant iron transporter during in vitro and in vivo growth. PMID- 12117950 TI - Flavohemoglobin Hmp protects Salmonella enterica serovar typhimurium from nitric oxide-related killing by human macrophages. AB - Survival of macrophage microbicidal activity is a prerequisite for invasive disease caused by the enteric pathogen Salmonella enterica serovar Typhimurium. Flavohemoglobins, such as those of Escherichia coli, Salmonella, and yeast, play vital roles in protection of these microorganisms in vitro from nitric oxide (NO) and nitrosative stress. A Salmonella hmp mutant defective in flavohemoglobin (Hmp) synthesis exhibits growth that is hypersensitive to nitrosating agents. We found that respiration of this mutant exhibited increased inhibition by NO, whereas wild-type cells pregrown with sodium nitroprusside or S nitrosoglutathione showed enhanced tolerance of NO. Most significantly, hmp mutants internalized by primary human peripheral monocyte-derived macrophages survived phagocytosis relatively poorly compared with similarly bound and internalized wild-type cells. That the enhanced sensitivity to macrophage microbicidal activity is due primarily to the failure of Salmonella to detoxify NO was suggested by the ability of L-N(G)-monomethyl arginine-an inhibitor of NO synthase-to eliminate the difference in killing between wild-type and hmp mutant Salmonella cells. These observations suggest that Salmonella Hmp contributes to protection from NO-mediated inhibition by human macrophages. PMID- 12117951 TI - Loss of regulatory protein RfaH attenuates virulence of uropathogenic Escherichia coli. AB - RfaH is a regulatory protein in Escherichia coli and Salmonella enterica serovar Typhimurium. Although it enhances expression of different factors that are proposed to play a role in bacterial virulence, a direct effect of RfaH on virulence has not been investigated so far. We report that inactivation of rfaH dramatically decreases the virulence of uropathogenic E. coli strain 536 in an ascending mouse model of urinary tract infection. The mortality rate caused by the wild-type strain in this assay is 100%, whereas that of its isogenic rfaH mutant does not exceed 18%. In the case of coinfection, the wild-type strain 536 shows higher potential to colonize the urinary tract even when it is outnumbered 100-fold by its rfaH mutant in the inoculum. In contrast to the wild-type strain, serum resistance of strain 536rfaH::cat is fully abolished. Furthermore, we give evidence that, besides a major decrease in the amount of hemin receptor ChuA (G. Nagy, U. Dobrindt, M. Kupfer, L. Emody, H. Karch, and J. Hacker, Infect. Immun. 69:1924-1928, 2001), loss of the RfaH protein results in an altered lipopolysaccharide phenotype as well as decreased expression of K15 capsule and alpha-hemolysin, whereas levels of other pathogenicity factors such as siderophores, flagella, Prf, and S fimbriae appear to be unaltered in strain 536rfaH::cat in comparison to the wild-type strain. trans complementation of the mutant strain with the rfaH gene restores wild-type levels of the affected virulence factors and consequently restitutes virulence in the mouse model of ascending urinary tract infection. PMID- 12117952 TI - Molecular cloning and characterization of genes for Shigella sonnei form I O polysaccharide: proposed biosynthetic pathway and stable expression in a live salmonella vaccine vector. AB - The gene region for biosynthesis of Shigella sonnei form I O polysaccharide (O Ps) and flanking sequences, totaling >18 kb, was characterized by deletion analysis to define a minimal construct for development of Salmonella-based live vaccine vector strains. Lipopolysaccharide (LPS) expression and DNA sequence studies of plasmid deletion derivatives indicated form I O-Ps expression from a 12.3-kb region containing a putative promoter and 10 contiguous open reading frames (ORFs), one of which is the transposase of IS630. A detailed biosynthetic pathway, consistent with the predicted functions of eight of the nine essential ORFs and the form I O-Ps structure, is proposed. Further sequencing identified partial IS elements (i.e., IS91 and IS630) and wzz upstream of the form I coding region and a fragment of aqpZ and additional full or partial IS elements (i.e., IS629, IS91, and IS911) downstream of this region. The stability of plasmid-based form I O-Ps expression was greater from low-copy vectors than from high-copy vectors and was enhanced by deletion of the downstream IS91 from plasmid inserts. Both core-linked (i.e., LPS) and non-core-linked (i.e., capsule-like) surface expression of form I O-Ps were detected by Western blotting and silver staining of polyacrylamide gel electrophoresis-separated Shigella and Escherichia coli extracts. However, salmonellae, which have a core that is chemically dissimilar to that of shigellae, expressed only non-core-linked surface-associated form I O Ps. Finally, attenuated Salmonella enterica serovar Typhi live vaccine vector candidates, containing minimal-sized form I operon constructs, elicited immune protection in mice against virulent S. sonnei challenge, thereby supporting the promise of live, oral vaccines for the prevention of shigellosis. PMID- 12117953 TI - ADP and other metabolites released from Acanthamoeba castellanii lead to human monocytic cell death through apoptosis and stimulate the secretion of proinflammatory cytokines. AB - Monocytes/macrophages are thought to be involved in Acanthamoeba infections. The aim of this work was to study whether soluble metabolites (ADP and other compounds) released by Acanthamoeba castellanii trophozoites could induce morphological and biochemical changes in human monocytic cells in vitro. We demonstrate here that ADP constitutively released in the medium by A. castellanii, interacting with specific P2y(2) purinoceptors expressed on the monocytic cell membrane, caused a biphasic rise in [Ca(2+)](i), morphological changes characteristics of cells undergoing apoptosis, caspase-3 activation, and secretion of tumor necrosis factor alpha (TNF-alpha). The same results were found in monocytes exposed to purified ADP. Cell damage and TNF-alpha release induced by amoebic ADP were blocked by the P2y(2) inhibitor suramin. Other metabolites contained in amoebic cell-free supernatants, with molecular masses of, respectively, >30 kDa and between 30 and 10 kDa, also caused morphological modifications and activation of intracellular caspase-3, characteristics of programmed cell death. Nevertheless, mechanisms by which these molecules trigger cell damage appeared to differ from that of ADP. In addition, other amoebic thermolable metabolites with molecular masses of <10 kDa caused the secretion of interleukin-1beta. These findings suggest that pathogenic free-living A. castellanii by release of ADP and other metabolites lead to human monocytic cell death through apoptosis and stimulate the secretion of proinflammatory cytokines. PMID- 12117954 TI - Immunochemical properties of the staphylococcal poly-N-acetylglucosamine surface polysaccharide. AB - Staphylococcus aureus and Staphylococcus epidermidis often elaborate adherent biofilms, which contain the capsular polysaccharide-adhesin (PS/A) that mediates the initial cell adherence to biomaterials. Biofilm cells produce another antigen, termed polysaccharide intercellular adhesin (PIA), which is composed of a approximately 28 kDa soluble linear beta(1-6)-linked N-acetylglucosamine. We developed a new method to purify PS/A from S. aureus MN8m, a strain hyperproducing PS/A. Using multiple analytical techniques, we determined that the chemical structure of PS/A is also beta(1-6)-N-acetylglucosamine (PNAG). We were unable to find N-succinylglucosamine residues in any of our preparations in contrast to previously reported findings (D. McKenney, K. Pouliot, Y. Wang, V. Murthy, M. Ulrich, G. Doring, J. C. Lee, D. A Goldmann, and G. B. Pier, Science 284:1523-1527, 1999). PNAG was produced with a wide range of molecular masses that could be divided into three major fractions with average molecular masses of 460 kDa (PNAG-I), 100 kDa (PNAG-II), and 21 kDa (PNAG-III). The purified antigens were not soluble at neutral pH unless first dissolved in 5 M HCl and then neutralized with 5 M NaOH. PNAG-I was very immunogenic in rabbits, but the responses of individual animals were variable. Immunization of mice with various doses (100, 50, or 10 microg) of PNAG-I, -II, and -III demonstrated that only PNAG-I was able to elicit an immunoglobulin G (IgG) immune response with the highest titers obtained with 100-microg dose. When we purified a small fraction of PNAG with a molecular mass of approximately 780 kDa (PNAG-780) from PNAG-I, significantly higher IgG titers than those in mice immunized with the same doses of PNAG-I were obtained, suggesting the importance of the molecular mass of PNAG in the antibody response. These results further clarify the chemical structure of PS/A and help to differentiate it from PIA on the basis of immunogenicity, molecular size, and solubility. PMID- 12117955 TI - Interleukin-10 controls the onset of irreversible septic shock. AB - Lethality from sepsis is believed to be mediated by a proinflammatory cytokine cascade, yet blocking the proinflammatory cytokines tumor necrosis factor alpha (TNF-alpha) and interleukin-1 (IL-1) fails to prevent mortality in human disease and a mouse model of sepsis induced by cecal ligation and puncture (CLP). The role of the antiinflammatory cytokine IL-10 in the CLP model of sepsis is unclear, with either protective or harmful effects demonstrated, depending upon the time of intervention. We therefore hypothesize that IL-10 functions as a temporal regulator of the transition from early reversible sepsis to the late phase of irreversible shock. Transition from reversible sepsis to irreversible shock in the CLP model was defined as the time when removal of the necrotic cecum by rescue surgery is no longer effective. We subjected IL-10-deficient (IL-10(-/ )) and wild-type (IL-10(+/+)) mice to CLP and monitored the progression of sepsis, the onset of irreversible shock, and mortality. Onset of lethality in IL 10(-/-) mice occurred significantly earlier than in IL-10(+/+) mice and was associated with 15-fold-higher serum levels of TNF-alpha and IL-6. Consistent with these findings, the efficacy of rescue surgery after lethal CLP is lost 10 h earlier in IL-10(-/-) mice than in IL-10(+/+) mice. Treatment with recombinant human IL-10 5 h after CLP significantly improved survival and lengthened the therapeutic window for rescue surgery in both strains of mice. These results demonstrate that IL-10 controls the onset of irreversible septic shock after CLP. PMID- 12117956 TI - Autotransported serine protease A of Neisseria meningitidis: an immunogenic, surface-exposed outer membrane, and secreted protein. AB - Several autotransporter proteins have previously been identified in Neisseria meningitidis. Using molecular features common to most members of the autotransporter family of proteins, we have identified an additional novel ca. 112-kDa autotransporter protein in the meningococcal genomic sequence data. This protein, designated autotransported serine protease A (AspA), has significant N terminal homology to the secreted serine proteases (subtilases) from several organisms and contains a serine protease catalytic triad. The amino acid sequence of AspA is well-conserved in serogroup A, B, and C meningococci. In Neisseria gonorrhoeae, the AspA homologue appears to be a pseudogene. The gene encoding AspA was cloned and expressed from meningococcal strain MC58 (B15:P1.16b). Anti AspA antibodies were detected in patients' convalescent-phase sera, suggesting that AspA is expressed in vivo during infection and is immunogenic and cross reactive. Rabbit polyclonal monospecific anti-AspA serum was used to probe whole cell proteins from a panel of wild-type meningococcal strains and two AspA mutant strains. Expression of the ca. 112-kDa precursor polypeptide was detected in 12 of 20 wild-type meningococcal strains examined, suggesting that AspA expression is phase variable. Immunogold electron microscopy and cellular fractionation studies showed that the AspA precursor is transported to the outer membrane and remains surface exposed. Western blot experiments confirmed that smaller, ca. 68- or 70-kDa components of AspA (AspA68 and AspA70, respectively) are then secreted into the meningococcal culture supernatant. Site-directed mutagenesis of S426 abolished secretion of both rAspA68 and rAspA70 in Escherichia coli, confirming that AspA is an autocleaved autotransporter protein. In conclusion, we characterized a novel, surface-exposed and secreted, immunogenic, meningococcal autotransporter protein. PMID- 12117957 TI - Defective Hyphal induction of a Candida albicans phosphatidylinositol 3-phosphate 5-kinase null mutant on solid media does not lead to decreased virulence. AB - A phosphatidylinositol 3-phosphate [PI(3)P] 5-kinase gene (CaFAB1) of the most important human pathogenic yeast, Candida albicans, was cloned and sequenced. An open reading frame was detected which encodes a 2,369-amino-acid protein with a calculated molecular mass of 268 kDa and a relative isoelectric point of 6.76. This protein exhibits 38% overall amino acid sequence identity with Saccharomyces cerevisiae Fab1p. We localized the CaFAB1 gene on chromosome R. To determine the influence of the PI(3)P 5-kinase CaFab1p on processes involved in C. albicans morphogenesis and pathogenicity, we sequentially disrupted both copies of the gene. Homozygous deletion of C. albicans CaFAB1 resulted in a mutant strain which exhibited defects in morphogenesis. A Cafab1 null mutant had enlarged vacuoles, an acidification defect, and increased generation times and was unable to form hyphae on different solid media. The sensitivities to hyperosmotic and high temperature stresses, adherence, and virulence compared to those of wild-type strain SC5314 were not affected. PMID- 12117958 TI - High-level expression of the malaria blood-stage vaccine candidate Plasmodium falciparum apical membrane antigen 1 and induction of antibodies that inhibit erythrocyte invasion. AB - Apical membrane antigen 1 (AMA-1) is a highly promising malaria blood-stage vaccine candidate that has induced protection in rodent and nonhuman primate models of malaria. Authentic conformation of the protein appears to be essential for the induction of parasite-inhibitory antibody responses. Here we have developed a synthetic gene with adapted codon usage to allow expression of Plasmodium falciparum FVO strain AMA-1 (PfAMA-1) in Pichia pastoris. In addition, potential N-glycosylation sites were changed, exploiting the lack of conservation of these sites in Plasmodium, to obtain high-level secretion of a homogeneous product, suitable for scale-up according to current good manufacturing procedures. Purified PfAMA-1 displayed authentic antigenic properties, indicating that the amino acid changes had no deleterious effect on the conformation of the protein. High-titer antibodies, raised in rabbits, reacted strongly with homologous and heterologous P. falciparum by immunofluorescence. In addition, purified immunoglobulin G from immunized animals strongly inhibited invasion of red blood cells by homologous and, to a somewhat lesser extent, heterologous P. falciparum. PMID- 12117959 TI - Identification of amino acid residues of anthrax protective antigen involved in binding with lethal factor. AB - Protective antigen (PA) and lethal factor (LF) are the two components of anthrax lethal toxin. PA is responsible for the translocation of LF to the cytosol. The binding of LF to cell surface receptor-bound PA is a prerequisite for the formation of lethal toxin. It has been hypothesized that hydrophobic residues P184, L187, F202, L203, P205, I207, I210, W226, and F236 of domain 1b of PA play an important role in the binding of PA to LF. These residues are normally buried in the 83-kDA version of PA, PA83, as determined by the crystal structure of PA. However, they become exposed due to the conformational change brought about by the cleavage of PA83 to PA63 by a cell surface protease. Mutation of the above mentioned residues to alanine resulted in mutant proteins that were able to bind to the cell surface receptors and also to be specifically cleaved by the cellular proteases. All the mutant proteins except the F202A, L203A, P205A, and I207A mutants were able to bind to LF and were also toxic to macrophage cells in combination with LF. It was concluded that residues 202, 203, 205, and 207 of PA are essential for the binding of LF to PA. PMID- 12117960 TI - Structure-function analysis of decay-accelerating factor: identification of residues important for binding of the Escherichia coli Dr adhesin and complement regulation. AB - Decay-accelerating factor (DAF), a complement regulatory protein, also serves as a receptor for Dr adhesin-bearing Escherichia coli. The repeat three of DAF was shown to be important in Dr adhesin binding and complement regulation. However, Dr adhesins do not bind to red blood cells with the rare polymorphism of DAF, designated Dr(a(-)); these cells contain a point mutation (Ser165-Leu) in DAF repeat three. In addition, monoclonal antibody IH4 specific against repeat three was shown to block both Dr adhesin binding and complement regulatory functions of DAF. Therefore, to identify residues important in binding of Dr adhesin and IH4 and in regulating complement, we mutated 11 amino acids-predominantly those in close proximity to Ser165 to alanine-and expressed these mutations in Chinese hamster ovary cells. To map the mutations, we built a homology model of repeat three based on the poxvirus complement inhibitory protein, using the EXDIS, DIAMOD, and FANTOM programs. We show that perhaps Ser155, and not Ser165, is the key amino acid that interacts with the Dr adhesin and amino acids Gly159, Tyr160, and Leu162 and also aids in binding Dr adhesin. The IH4 binding epitope contains residues Phe148, Ser155, and L171. Residues Phe123 and Phe148 at the interface of repeat 2-3, and also Phe154 in the repeat three cavity, were important for complement regulation. Our results show that residues affecting the tested functions are located on the same loop (148 to 171), at the same surface of repeat three, and that the Dr adhesin-binding and complement regulatory epitopes of DAF appear to be distinct and are approximately 20 A apart. PMID- 12117961 TI - Identification and characterization of a novel heme-associated cell surface protein made by Streptococcus pyogenes. AB - Analysis of the genome sequence of a serotype M1 group A Streptococcus (GAS) strain identified a gene encoding a previously undescribed putative cell surface protein. The gene was cloned from a serotype M1 strain, and the recombinant protein was overexpressed in Escherichia coli and purified to homogeneity. The purified protein was associated with heme in a 1:1 stoichiometry. This streptococcal heme-associated protein, designated Shp, was produced in vitro by GAS, located on the bacterial cell surface, and accessible to specific antibody raised against the purified recombinant protein. Mice inoculated subcutaneously with GAS and humans with invasive infections and pharyngitis caused by GAS seroconverted to Shp, indicating that Shp was produced in vivo. The blood of mice actively immunized with Shp had significantly higher bactericidal activity than the blood of unimmunized mice. The shp gene was cotranscribed with eight contiguous genes, including homologues of an ABC transporter involved in iron uptake in gram-negative bacteria. Our results indicate that Shp is a novel cell surface heme-associated protein. PMID- 12117962 TI - Dissemination of Mycobacterium tuberculosis is influenced by host factors and precedes the initiation of T-cell immunity. AB - We report that dissemination of Mycobacterium tuberculosis in the mouse is under host control and precedes the initiation of T-cell immunity. Nine to eleven days after aerosol inoculation, M. tuberculosis disseminates to the pulmonary lymph nodes (LN), where M. tuberculosis-specific T cells are detected 2 to 3 days thereafter. This indicates that the initial spread of bacteria occurs via lymphatic drainage and that the acquired T-cell immune response is generated in the draining LN. Dissemination to peripheral sites, such as the spleen and the liver, occurs 11 to 14 days postinfection and is followed by the appearance of M. tuberculosis-specific T cells in the lung and the spleen. In all cases studied, dissemination to the LN or the spleen preceded activation of M. tuberculosis specific T cells in that organ. Interestingly, bacteria disseminate earlier from the lungs of resistant C57BL/6 mice than from the lungs of susceptible C3H mice, and consequently, C57BL/6 mice generate an immune response to M. tuberculosis sooner than C3H mice generate an immune response. Thus, instead of spreading infection, early dissemination of M. tuberculosis may aid in the initiation of an appropriate and timely immune response. We hypothesize that this early initiation of immunity following inoculation with M. tuberculosis may contribute to the superior resistance of C57BL/6 mice. PMID- 12117963 TI - Genes for glycosylphosphatidylinositol toxin biosynthesis in Plasmodium falciparum. AB - About 2.5 million people die of Plasmodium falciparum malaria every year. Fatalities are associated with systemic and organ-specific inflammation initiated by a parasite toxin. Recent studies show that glycosylphosphatidylinositol (GPI) functions as the dominant parasite toxin in the context of infection. GPIs also serve as membrane anchors for several of the most important surface antigens of parasite invasive stages. GPI anchoring is a complex posttranslational modification produced through the coordinated action of a multicomponent biosynthetic pathway. Here we present eight new genes of P. falciparum selected for encoding homologs of proteins essential for GPI synthesis: PIG-A, PIG-B, PIG M, PIG-O, GPI1, GPI8, GAA-1, and DPM1. We describe the experimentally verified mRNA and predicted amino acid sequences and in situ localization of the gene products to the parasite endoplasmic reticulum. Moreover, we show preliminary evidence for the PIG-L and PIG-C genes. The biosynthetic pathway of the malaria parasite GPI offers potential targets for drug development and may be useful for studying parasite cell biology and the molecular basis for the pathophysiology of parasitic diseases. PMID- 12117964 TI - A hag mutant of Moraxella catarrhalis strain O35E is deficient in hemagglutination, autoagglutination, and immunoglobulin D-binding activities. AB - Previous studies correlated the presence of a 200-kDa protein on the surface of Moraxella catarrhalis with the ability of this organism to agglutinate human erythrocytes (M. Fitzgerald, R. Mulcahy, S. Murphy, C. Keane, D. Coakley, and T. Scott, FEMS Immunol. Med. Microbiol. 18:209-216, 1997). In the present study, the gene encoding the 200-kDa protein (designated Hag) of M. catarrhalis strain O35E was subjected to nucleotide sequence analysis and then was inactivated by insertional mutagenesis. The isogenic hag mutant was unable to agglutinate human erythrocytes and lost its ability to autoagglutinate but was still attached at wild-type levels to several human epithelial cell lines. The hag mutation also eliminated the ability of this mutant strain to bind human immunoglobulin D. The presence of the Hag protein on the M. catarrhalis cell surface, as well as that of the UspA1 and UspA2 proteins (C. Aebi, I. Maciver, J. L. Latimer, L. D. Cope, M. K. Stevens, S. E. Thomas, G. H. McCracken, Jr., and E. J. Hansen, Infect. Immun. 65:4367-4377, 1997), was investigated by transmission electron and cryoimmunoelectron microscopy. Wild-type M. catarrhalis strain O35E possessed a dense layer of surface projections, whereas an isogenic uspA1 uspA2 hag triple mutant version of this strain did not possess any detectable surface projections. Examination of a uspA1 uspA2 double mutant that expressed the Hag protein revealed the presence of a relatively sparse layer of surface projections, similar to those seen on a uspA2 hag mutant that expressed UspA1. In contrast, a uspA1 hag mutant that expressed UspA2 formed a very dense layer of relatively short surface projections. These results indicate that the surface-exposed Hag protein and UspA1 and UspA2 have the potential to interact both with each other and directly with host defense systems. PMID- 12117965 TI - Sequential expression of the neuropeptides substance P and somatostatin in granulomas associated with murine cysticercosis. AB - Neurocysticercosis, a parasitic infection of the human central nervous system caused by Taenia solium, is a leading cause of seizures. Seizures associated with neurocysticercosis are caused mainly by the host inflammatory responses to dying parasites in the brain parenchyma. We previously demonstrated sequential expression of Th1 cytokines in early-stage granulomas, followed by expression of Th2 cytokines in later-stage granulomas in murine cysticercosis. However, the mechanism leading to this shift in cytokine response in the granulomas is unknown. Neuropeptides modulate cytokine responses and granuloma formation in murine schistosomiasis. Substance P (SP) induces Th1 cytokine expression and granuloma formation, whereas somatostatin inhibits the granulomatous response. We hypothesized that neuropeptides might play a role in regulation of the granulomatous response in cysticercosis. To test this hypothesis, we compared expression of SP and expression of somatostatin in murine cysticercal granulomas by using in situ hybridization and immunohistochemistry. We also compared expression with granuloma stage. Expression of SP mRNA was more frequent in the early-stage granulomas than in the late-stage granulomas (34 of 35 early-stage granulomas versus 1 of 13 late-stage granulomas). By contrast, somatostatin was expressed primarily in later-stage granulomas (13 of 14 late-stage granulomas versus 2 of 35 early-stage granulomas). The median light microscope grade of SP mRNA expression in the early-stage granulomas was significantly higher than that in the late-stage granulomas (P = 0.008, as determined by the Wilcoxon signed rank test). By contrast, somatostatin mRNA expression was higher at later stages (P = 0.008, as determined by the Wilcoxon signed rank test). SP and somatostatin are therefore temporally expressed in granulomas associated with murine cysticercosis, which may be related to differential expression of Th1 and Th2 cytokines. PMID- 12117966 TI - Sat, the secreted autotransporter toxin of uropathogenic Escherichia coli, is a vacuolating cytotoxin for bladder and kidney epithelial cells. AB - The secreted autotransporter toxin (Sat) of uropathogenic Escherichia coli exhibits cytopathic activity upon incubation with HEp-2 cells. We further investigated the effects of Sat on cell lines more relevant to the urinary tract, namely, those derived from bladder and kidney epithelium. Sat elicited elongation of cells and apparent loosening of cellular junctions upon incubation with Vero kidney cells. Additionally, incubation with Sat triggered significant vacuolation within the cytoplasm of both human bladder (CRL-1749) and kidney (CRL-1573) cell lines. This activity has been associated with only a few other known toxins. Following transurethral infection of CBA mice with a sat mutant, no reduction of CFU in urine, bladder, or kidney tissue was seen compared to that in mice infected with wild-type E. coli CFT073. However, significant histological changes were observed within the kidneys of mice infected with wild-type E. coli CFT073, including dissolution of the glomerular membrane and vacuolation of proximal tubule cells. Such damage was not observed in kidney sections of mice infected with a Sat-deficient mutant. These results indicate that Sat, a vacuolating cytotoxin expressed by uropathogenic E. coli CFT073, elicits defined damage to kidney epithelium during upper urinary tract infection and thus contributes to pathogenesis of urinary tract infection. PMID- 12117967 TI - Patterns of epithelial cell invasion by different species of the Burkholderia cepacia complex in well-differentiated human airway epithelia. AB - Burkholderia cepacia has emerged as a serious respiratory pathogen in cystic fibrosis (CF) patients. The clinical course of B. cepacia infections is variable, but approximately 20% of patients eventually succumb to the cepacia syndrome, which is characterized as a fatal necrotizing pneumonia with bacteremia. The mechanisms that permit B. cepacia to cause bacteremia are not yet known but probably involve sequential penetration of airway barriers. This study evaluated the abilities of different species of the B. cepacia complex, including a strain from the ET12 lineage (BC-7, genomovar III, cblA(+)), which is associated with most cepacia syndrome fatalities among CF populations, a genomovar IV strain (HI2258), and a genomovar II strain (J-1) to penetrate polarized, well differentiated human airway epithelial cell cultures. As revealed by light and electron microscopy, all three B. cepacia strains tested circumvented the mechanical barriers of mucus and ciliary transport to penetrate the airway epithelium but they used different routes. The BC-7 strain (genomovar III) formed biofilms in close proximity to the apical cell surface, followed by invasion and destruction of epithelial cells. This process involved disruption of the glycocalyx and rearrangements of the actin cytoskeleton. The HI2258 strain (genomovar IV) did not form biofilms, and the majority of bacteria that penetrated the epithelium were located between epithelial cells, suggesting paracytosis. Strain J-1 penetrated the epithelium both by cell destruction and paracytosis. These studies suggest that the distinct invasion pathways employed by B. cepacia may account for differences in virulence between B. cepacia genomovars. PMID- 12117968 TI - Identification of Escherichia coli outer membrane protein A receptor on human brain microvascular endothelial cells. AB - Neonatal Escherichia coli meningitis continues to be a diagnostic and treatment challenge despite the availability of active antibiotics. Our earlier studies have shown that outer membrane protein A (OmpA) is one of the major factors responsible for Escherichia coli traversal across the blood-brain barrier that constitutes a lining of brain microvascular endothelial cells (BMEC). In this study we showed that OmpA binds to a 95-kDa human BMEC (HBMEC) glycoprotein (Ecgp) for E. coli invasion. Ecgp was partially purified by wheat germ agglutinin and Maackia amurensis lectin (MAL) affinity chromatography. The MAL affinity purified HBMEC proteins bound to OmpA(+) E. coli but not to OmpA(-) E. coli. In addition, the deglycosylated MAL-bound proteins still interact with OmpA(+) E. coli, indicating the role of protein backbone in mediating the OmpA binding to HBMEC. Interestingly, the MAL affinity-bound fraction showed one more protein, a 65-kDa protein that bound to OmpA(+) E. coli in addition to Ecgp. Further, the 65 kDa protein was shown to be a cleavage product of Ecgp. Immunocytochemistry of HBMEC infected with OmpA(+) E. coli by using anti-Ecgp antibody suggests that Ecgp clusters at the E. coli entry site. Anti-Ecgp antibody also reacted to microvascular endothelium on human brain tissue sections, indicating the biological relevance of Ecgp in E. coli meningitis. Partial N-terminal amino acid sequence of Ecgp suggested that it has 87% sequence homology to gp96, an endoplasmic reticulum-resident molecular chaperone that is often expressed on the cell surface. In contrast, the 65-kDa protein, which could be the internal portion of Ecgp, showed 70% sequence homology to an S-fimbria-binding sialoglycoprotein reported earlier. These results suggest that OmpA interacts with Ecgp via the carbohydrate epitope, as well as with the protein portion for invading HBMEC. PMID- 12117969 TI - Induction of a potential paracrine angiogenic loop between human THP-1 macrophages and human microvascular endothelial cells during Bartonella henselae infection. AB - Bartonella henselae is responsible for various disease syndromes that loosely correlate with the immune status of the host. In the immunocompromised individual, B. henselae-induced angiogenesis, or bacillary angiomatosis, is characterized by vascular proliferative lesions similar to those in Kaposi's sarcoma. We hypothesize that B. henselae-mediated interaction with immune cells, namely, macrophages, induces potential angiogenic growth factors and cytokines which contribute in a paracrine manner to the proliferation of endothelial cells. Vascular endothelial growth factor (VEGF), a direct inducer of angiogenesis, and interleukin-1beta (IL-1beta), a potentiator of VEGF, were detected within 12 and 6 h, respectively, in supernatants from phorbol 12-myristate 13-acetate differentiated human THP-1 macrophages exposed to live B. henselae. Pretreatment of macrophages with cytochalasin D, a phagocytosis inhibitor, yielded comparable results, suggesting that bacterium-cell attachment is sufficient for VEGF and IL 1beta induction. IL-8, an angiogenic cytokine with chemotactic properties, was induced in human microvascular endothelial cells (HMEC-1) within 6 h of infection, whereas no IL-8 induction was observed in infected THP-1 cells. In addition, conditioned medium from infected macrophages induced the proliferation of HMEC-1, thus demonstrating angiogenic potential. These data suggest that Bartonella modulation of host or target cell cytokines and growth factors, rather than a direct role of the bacterium as an endothelial cell mitogen, is the predominant mechanism responsible for angiogenesis. B. henselae induction of VEGF, IL-1beta, and IL-8 outlines a broader potential paracrine angiogenic loop whereby macrophages play the predominant role as the effector cell and endothelial cells are the final target cell, resulting in their proliferation. PMID- 12117972 TI - Tumor necrosis factor alpha stimulates killing of Mycobacterium tuberculosis by human neutrophils. AB - The ability of human neutrophils to aid in defense against pulmonary infection with Mycobacterium tuberculosis is controversial. In this study, we have shown that neutrophils respond to and phagocytose M. tuberculosis in human lesions. Neutrophils from healthy individuals were able to kill significant fractions of an inoculum of M. tuberculosis within 1 h of phagocytosis, and this ability was enhanced by tumor necrosis factor alpha but not by gamma interferon. The mycobactericidal mechanism was nonoxidative, as inhibitors of reactive oxygen or reactive nitrogen intermediates did not interfere with killing. However, the mycobactericidal mechanism was associated with increased exposure of intracellular M. tuberculosis to neutrophil defensins. In vitro, human neutrophil peptides 1 to 3 were not able to kill the bacilli even at much higher levels. These studies support the concept that human neutrophils are directly involved in defense against infection with M. tuberculosis. PMID- 12117970 TI - Opsonization modulates Rac-1 activation during cell entry by Leishmania amazonensis. AB - Lesions caused by Leishmania amazonensis normally heal, but relapses occur due to parasite persistence in host tissues. It has been proposed that infection of fibroblasts plays an important role in this process by providing the parasites with a safe haven in which to replicate. However, most previous studies have focused on the entry of Leishmania into macrophages, a process mediated by serum opsonins. To gain insight into a possible role of nonopsonic entry in the intracellular persistence of amastigotes, we examined the invasion of Chinese hamster ovary (CHO) cells. Amastigotes entered CHO cells by a cytochalasin D, genistein, wortmannin, and 2,3-butanedione monoxime-sensitive pathway and replicated within phagolysosomes. However, unlike most phagocytic processes described to date, amastigote internalization in CHO cells involved activation of the GTPases Rho and Cdc42 but not Rac-1. When uptake was mediated by fibronectin or when amastigotes were opsonized with immunoglobulin G and internalized by Fc receptor-expressing CHO cells, Rac-1 activation was restored and found to be required for parasite internalization. Given the essential role of Rac in assembly of the respiratory burst oxidase, invasion through this nonopsonic, Rac 1-independent pathway may play a central role in the intracellular survival of Leishmania in immune hosts. PMID- 12117971 TI - Helicobacter pylori-induced activation of human endothelial cells. AB - Helicobacter pylori infection causes active chronic inflammation with a continuous recruitment of neutrophils to the inflamed gastric mucosa. To evaluate the role of endothelial cells in this process, we have examined adhesion molecule expression and chemokine and cytokine production from human umbilical vein endothelial cells stimulated with well-characterized H. pylori strains as well as purified proteins. Our results indicate that endothelial cells actively contribute to neutrophil recruitment, since stimulation with H. pylori bacteria induced upregulation of the adhesion molecules VCAM-1, ICAM-1, and E-selectin as well as the chemokines interleukin 8 (IL-8) and growth-related oncogene alpha (GRO-alpha) and the cytokine IL-6. However, there were large variations in the ability of the different H. pylori strains to stimulate endothelial cells. These interstrain variations were seen irrespective of whether the strains had been isolated from patients with duodenal ulcer disease or asymptomatic carriers and were not solely related to the expression of known virulence factors, such as the cytotoxin-associated gene pathogenicity island, vacuolating toxin A, and Lewis blood group antigens. In addition, one or several unidentified proteins which act via NF-kappaB activation seem to induce endothelial cell activation. In conclusion, human endothelial cells produce neutrophil-recruiting factors and show increased adhesion molecule expression after stimulation with certain H. pylori strains. These effects probably contribute to the continuous recruitment of neutrophils to H. pylori-infected gastric mucosa and may also contribute to tissue damage and ulcer formation. PMID- 12117973 TI - Quantitative priming with inactivated pertussis toxoid vaccine in the aerosol challenge model. AB - Serum antibodies to pertussis toxin (PT) have been shown to be protective against severe pertussis disease, although a specific level of anti-PT antibody that correlates with protection has not been demonstrated. Current animal models such as the intracerebral challenge model have significant limitations in correlating protection to a specific level of anti-PT antibody. This study examines the protective effects of priming with tetranitromethane-inactivated pertussis toxoid (PTx) vaccine in the aerosol challenge model and whether a measurable response to a priming dose of PTx is enough to initiate a protective secondary response when challenged with infection. The correlation of priming with markers of illness such as leukocytosis, weight loss, bacterial proliferation, and mortality after established infection with Bordetella pertussis was explored. BALB/c mice were immunized with PTx vaccine on day 6 of life and then challenged with B. pertussis using the aerosol challenge model. Data were analyzed according to the primary immunologic response, differentiating responders (anti-PT immunoglobulin G [IgG] > or =1 microg/ml) from nonresponders (anti-PT IgG <1 microg/ml). Mice that showed evidence of priming on the day of aerosol challenge were able to mount a secondary response to the challenge with a > or =2-fold rise in anti-PT IgG antibody by day 7 and a > or =10-fold rise by day 14 post-aerosol challenge. These primed mice were significantly better protected against leukocytosis, weight loss, and proliferation of B. pertussis in the lungs following aerosol challenge than the nonprimed group. This protection correlated with levels of anti-PT antibody in serum present on the day of aerosol challenge. PMID- 12117974 TI - Fusobacterium necrophorum leukotoxin induces activation and apoptosis of bovine leukocytes. AB - Fusobacterium necrophorum, a gram-negative, rod-shaped, anaerobic bacterium, is a primary or secondary etiological agent in a variety of necrotic, purulent infections in humans and animals. Its major virulence factor is leukotoxin, a high-molecular-weight secreted protein, primarily toxic to ruminant leukocytes. In this study, bovine peripheral blood leukocytes were exposed to various concentrations of immunoaffinity-purified leukotoxin and the cytotoxicity was analyzed by flow cytometry and scanning and transmission electron microscopy. At very low toxin concentrations, polymorphonuclear leukocytes (PMNs) showed activation, as indicated by translocation of primary and secondary granules to the periphery of the cytoplasm. Furthermore, these cells showed changes characteristic of apoptosis, including decreased cell size, organelle condensation, cytoplasmic membrane blebbing (zeiosis), and chromatin condensation and margination, and decrease in cellular DNA content. At moderately high concentrations of leukotoxin, bovine mononuclear cells were also induced to undergo programmed cell death. At very high concentrations, leukotoxin caused necrotic cell death of bovine peripheral leukocytes. The ability of F. necrophorum leukotoxin to modulate the host immune system by its toxicity, including cellular activation of PMNs and apoptosis-mediated killing of phagocytes and immune effector cells, represents a potentially important mechanism of its pathogenesis. PMID- 12117975 TI - Effects of oral vaccination and immunomodulation by cholera toxin on experimental Helicobacter pylori infection, reinfection, and gastritis. AB - Therapeutic vaccination is an attractive strategy to control infection and disease caused by Helicobacter pylori. In mice infected with H. pylori we have studied the protective effect of oral immunization with an H. pylori lysate preparation given together with the mucosal adjuvant cholera toxin (CT), both against the initial infection and against a later reinfection challenge. We have also examined the effects of treatment with the CT adjuvant alone on H. pylori infection and reinfection. Specific immunization with lysate was found to result in a sixfold reduction of the extent (bacterial load) of the primary infection and also to provide similar levels of protection against reinfection. However, these effects were associated with severe postimmunization gastritis. In contrast, oral treatment with CT alone at the time of initial infection, while unable to suppress the initial infection, gave rise to a 20-fold reduction in bacterial load upon reinfection without causing any associated gastric inflammation. Both the infected animals that were specifically immunized and those that were treated with CT only displayed increased in vitro proliferative responses of mononuclear cells to H. pylori antigens. Antibody levels in response to H. pylori were on the other hand only marginally increased after treatment with CT, whereas they were markedly elevated after immunization with lysate plus CT, with a rise in both (Th2-driven) immunoglobulin G1 (IgG1) and, especially, (Th1-driven) IgG2a antibodies. The results illustrate the complex balance between protection and harmful inflammation after postinfection vaccination against H. pylori as studied in a mouse model. PMID- 12117977 TI - Expression of inducible nitric oxide synthase in skin lesions of patients with american cutaneous leishmaniasis. AB - Cytokine-inducible (or type 2) nitric oxide synthase (iNOS) is indispensable for the resolution of Leishmania major or Leishmania donovani infections in mice. In contrast, little is known about the expression and function of iNOS in human leishmaniasis. Here, we show by immunohistological analysis of skin biopsies from Mexican patients with local (LCL) or diffuse (DCL) cutaneous leishmaniasis that the expression of iNOS was most prominent in LCL lesions with small numbers of parasites whereas lesions with a high parasite burden (LCL or DCL) contained considerably fewer iNOS-positive cells. This is the first study to suggest an antileishmanial function of iNOS in human Leishmania infections in vivo. PMID- 12117978 TI - High expression of a C protein beta antigen gene among invasive strains from certain clonally related groups of type Ia and Ib group B streptococci. AB - Serotyped strains of group B streptococci can be divided into subtypes based on restriction endonuclease digestion patterns (RDP). Profiles of cell-bound proteins were compared among RDP types. Proteins that showed a remarkable difference in the degree of expression were found among strains of RDP Ia-3, which has been considered potentially virulent, as well as of RDP Ib-1. For RDP Ia-3 strains, the protein was predominant in strains from cerebrospinal fluid (CSF) but was mostly a minor component in other strains. For RDP Ib-1 strains, the protein was predominant in strains from CSF, showed diversity in strains from blood, and was mostly a minor component in other strains. By N-terminal sequencing analysis, the protein was identified as a C protein beta antigen. The level of bound immunoglobulin A (IgA) or anti-beta antigen monoclonal antibody correlated with the level of expressed beta antigen, and invasive strains showed remarkably high levels of binding; the exception was a CSF-derived strain of RDP Ib-1 which produced a large amount of beta antigen and showed a high level of binding of anti-beta antigen monoclonal antibody but no IgA binding. PCR-based amplification revealed that the beta antigen gene was detected in all RDP Ia-3 and Ib-1 strains but was not found in any strains of other RDP types. Competitive reverse transcriptase PCR demonstrated that the difference in the amount of protein produced was due to the difference in the level of expression of the beta antigen mRNA. Our findings imply that differences in gene expression for a protein may contribute to the invasiveness of RDP Ia-3 and Ib-1 strains for the host. PMID- 12117979 TI - Mobilization of protein kinase C in macrophages induced by Listeria monocytogenes affects its internalization and escape from the phagosome. AB - Listeriolysin O (LLO) and a phosphatidylinositol-specific phospholipase C (PI PLC) are known virulence factors of Listeria monocytogenes in both tissue cultures and the murine model of infection. LLO is a member of a family of pore forming cholesterol-dependent cytotoxins and is known to play an essential role in escape from the primary phagocytic vacuole of macrophages. PI-PLC plays an accessory role, in that PI-PLC mutants are partially defective in escape. We have shown that both of these molecules are essential for initiating rapid increases in the calcium level in the J774 murine macrophage cell line (S. J. Wadsworth and H. Goldfine, Infect. Immun. 67:1770-1778, 1999). Here we show that both LLO and PI-PLC are required for translocation of protein kinase C delta (PKC delta) to the periphery of J774 cells and for translocation of PKC beta II to early endosomes beginning within the first minute after addition of bacteria to the culture medium. Treatment with the calcium channel blocker SK&F 96365 inhibited translocation of PKC beta II but not PKC delta. Our findings lead us to propose a host signaling pathway requiring LLO and the formation of diacylglycerol by PI PLC in which calcium-independent PKC delta is responsible for the initial calcium signal and the subsequent PKC beta II translocation. LLO-dependent translocation of PKC beta I to early endosomes also occurs between 1 and 4 min after infection, but this occurs in the absence of PI-PLC. All of these signals were observed in cells that had not internalized bacteria. Blocking PKC beta translocation with hispidin resulted in more rapid uptake of wild-type bacteria and greatly reduced escape from the primary phagocytic vacuoles of J774 cells. PMID- 12117976 TI - Old mice express a transient early resistance to pulmonary tuberculosis that is mediated by CD8 T cells. AB - During the natural aging process the immune system undergoes many alterations. In particular, both the CD4 and CD8 T-cell compartments become compromised, and these changes have serious implications for the capacity of the elderly to control infection. As a result, the elderly are more susceptible to many infectious diseases, including primary infection and reactivation of latent infections. In this study we addressed the capacity of old mice to control an infection with Mycobacterium tuberculosis and to characterize the mechanism by which old mice, paradoxically, can express a transient early resistance to infection. This resistance was shown to be associated with the presence of CD8 T cells within the lungs that were capable of secreting gamma interferon, as illustrated by the demonstration that early resistance was lost in aged CD8 gene disrupted mice. These studies therefore show that, despite a documented decline in general CD8 T-cell responsiveness in the elderly, a subset of CD8 T cells is an important early mediator of protection in the lungs of old mice that have been infected with M. tuberculosis. PMID- 12117980 TI - Acylation of the lipooligosaccharide of Haemophilus influenzae and colonization: an htrB mutation diminishes the colonization of human airway epithelial cells. AB - Haemophilus influenzae is a commensal and opportunistic pathogen of the human airways. A number of surface molecules contribute to colonization of the airways by H. influenzae, such as adhesins, including structures found in the lipooligosaccharide (LOS). A human bronchiolar xenograft model was employed to investigate the host-bacterial interactions involved in the colonization of the airway by H. influenzae. Differential display was used to identify H. influenzae mRNA that reflect genes which were preferentially expressed in the xenograft compared to growth. Eleven mRNA fragments had consistent increased expression when the bacteria grew in xenografts. On sequencing these fragments, eight open reading frames were identified. Three of these had no match in the NCBI or the TIGR database, while an additional three were homologous to genes involved in heme or iron acquisition and utilization: two of the mRNAs encoded proteins homologous to enzymes involved in LOS biosynthesis: a heptosyl transferase (rfaF) involved in the synthesis of the LOS core and a ketodeoxyoctonate phosphate dependent acyltransferase (htrB) that performs one of the late acylation reactions in lipid A synthesis. Inoculation of human bronchiolar xenografts revealed a significant reduction in colonization capacity by htrB mutants. In vitro, htrB mutants elicited lesser degrees of cytoskeletal rearrangement and less stimulation of host cell signaling with 16HBE14o(-) cells and decreased intracellular survival. These results implicate acylation of H. influenzae lipid A as playing a key role in the organisms' colonization of the normal airway. PMID- 12117982 TI - The Enterococcus faecalis fsrB gene, a key component of the fsr quorum-sensing system, is associated with virulence in the rabbit endophthalmitis model. AB - We used a rabbit endophthalmitis model to explore the role of fsrB, a gene required for the function of the fsr quorum-sensing system of Enterococcus faecalis, in pathogenicity. A nonpolar deletion mutant of fsrB had significantly reduced virulence compared to wild type. Complementation of mutation restored virulence. These data corroborate the role of fsrB in E. faecalis pathogenesis and suggest that the rabbit endophthalmitis model can be used to study the in vivo role of quorum sensing. PMID- 12117981 TI - Escherichia coli shiga-like toxins induce apoptosis and cleavage of poly(ADP ribose) polymerase via in vitro activation of caspases. AB - Shiga-like toxin-producing Escherichia coli causes hemorrhagic colitis and hemolytic-uremic syndrome in association with the production of Shiga-like toxins, which induce cell death via either necrosis or apoptosis. However, the abilities of different Shiga-like toxins to trigger apoptosis and the sequence of intracellular signaling events mediating the death of epithelial cells have not been completely defined. Fluorescent dye staining with acridine orange and ethidium bromide showed that Shiga-like toxin 1 (Stx1) induced apoptosis of HEp-2 cells in a dose- and time-dependent manner. Stx2 also induced apoptosis in a dose dependent manner. Apoptosis induced by Stx1 (200 ng/ml) and apoptosis induced by Stx2 (200 ng/ml) were maximal following incubation with cells for 24 h (94.3% +/- 1.8% and 81.7% +/- 5.2% of the cells, respectively). Toxin-treated cells showed characteristic features of apoptosis, including membrane blebbing, DNA fragmentation, chromatin condensation, cell shrinkage, and the formation of apoptotic bodies, as assessed by transmission electron microscopy. Stx2c induced apoptosis weakly even at a high dose (1,000 ng/ml for 24 h; 26.7% +/- 1.3% of the cells), whereas Stx2e did not induce apoptosis of HEp-2 cells. Thin-layer chromatography confirmed that HEp-2 cells express the Stx1-Stx2-Stx2c receptor, globotriaosylceramide (Gb3), but not the Stx2e receptor, globotetraosylceramide (Gb4). Western blot analysis of poly(ADP-ribose) polymerase (PARP), a DNA repair enzyme, demonstrated that incubation with Stx1 and Stx2 induced cleavage, whereas incubation with Stx2e did not result in cleavage of PARP. A pan-caspase inhibitor (Z-VAD-FMK) and a caspase-8-specific inhibitor (Z-IETD-FMK) eliminated, in a dose dependent fashion, the cleavage of PARP induced by Shiga-like toxins. Caspase-8 activation was confirmed by detection of cleavage of this enzyme by immunoblotting. Cleavage of caspase-9 and the proapoptotic member of the Bcl-2 family BID was also induced by Stx1, as determined by immunoblot analyses. We conclude that different Shiga-like toxins induce different degrees of apoptosis that correlates with toxin binding to the glycolipid receptor Gb3 and that caspases play an integral role in the signal transduction cascade leading to toxin-mediated programmed cell death. PMID- 12117983 TI - Colony-stimulating factor 1-dependent cells protect against systemic infection with Listeria monocytogenes but facilitate neuroinvasion. AB - By using mice genomically lacking the mononuclear phagocytic growth factor colony stimulating factor 1 and thereby deficient in macrophage and dendritic cell populations, we show that these cells play a dual role: they constitute a major defense against systemic infection but also facilitate cerebral bacterial invasion by Listeria monocytogenes. PMID- 12117985 TI - Orientia tsutsugamushi inhibits apoptosis of macrophages by retarding intracellular calcium release. AB - Orientia tsutsugamushi shows both pro- and antiapoptotic activities in infected vertebrate cells. Apoptosis of THP-1 cells induced by beauvericin was inhibited by O. tsutsugamushi infection. Beauvericin-induced calcium redistribution was significantly reduced and retarded in cells infected with O. tsutsugamushi. Antiapoptotic activities of O. tsutsugamushi in infected cells are most probably due to inhibition of the increase in the cytosolic calcium concentration. PMID- 12117984 TI - cag+ Helicobacter pylori induces homotypic aggregation of macrophage-like cells by up-regulation and recruitment of intracellular adhesion molecule 1 to the cell surface. AB - Infection with cag+ but not cag-negative Helicobacter pylori leads to the formation of large homotypic aggregates of macrophage-like cells. Intracellular adhesion molecule 1 is up-regulated and recruited to the cell surface of infected cells and mediates the aggregation via lymphocyte function-associated molecule 1. This signaling may regulate cell-cell interactions and inflammatory responses. PMID- 12117986 TI - Internalization of Staphylococcus aureus by human corneal epithelial cells: role of bacterial fibronectin-binding protein and host cell factors. AB - Wild-type Staphylococcus aureus was observed to be capable of invading human corneal epithelial cells (HCEC) in vitro. Internalization of S. aureus required expression of fibronectin-binding proteins (FnBPs); the capacity of an FnBP deficient isogenic strain to invade HCEC was reduced by more than 99%. The binding of S. aureus to HCEC did not require viable bacteria, since UV-killed cells were observed to adhere efficiently. Invasion of HCEC by S. aureus involved active host cell mechanisms; uptake was nearly completely eliminated by cytochalasin D and genistein. These data suggest that FnBPs play a key role in host-parasite interactions and may serve as an important adhesin or invasin in ulcerative keratitis caused by S. aureus. PMID- 12117987 TI - The omp-1 major outer membrane multigene family of Ehrlichia chaffeensis is differentially expressed in canine and tick hosts. AB - Sixteen of 22 omp-1 paralogs encoding 28-kDa-range immunodominant outer membrane proteins of Ehrlichia chaffeensis were transcribed in blood monocytes of dogs throughout a 56-day infection period. Only one paralog was transcribed by E. chaffeensis in three developmental stages of Amblyomma americanum ticks before or after E. chaffeensis transmission to naive dogs. PMID- 12117988 TI - Diverse bacteria are pathogens of Caenorhabditis elegans. AB - Practically and ethically attractive as model systems, invertebrate organisms are increasingly recognized as relevant for the study of bacterial pathogenesis. We show here that the nematode Caenorhabditis elegans is susceptible to a surprisingly broad range of bacteria and may constitute a useful model for the study of both pathogens and symbionts. PMID- 12117989 TI - Role of polyphosphate kinase in biofilm formation by Porphyromonas gingivalis. AB - In order to assess the role of polyphosphate kinase (PPK) in the physiology of Porphyromonas gingivalis, a ppk gene mutant, CW120, was constructed and characterized. P. gingivalis was demonstrated to synthesize short-chain polyphosphate (polyP) but not long-chain polyP. CW120 failed to survive in the stationary phase as well as the parental cell did, and it was attenuated in biofilm formation on polyvinylchloride and glass surfaces. Furthermore, the complementation by insertion of an intact copy of the ppk gene into the mutant CW120 restored its biofilm formation and stationary-phase survival. These results suggest that PPK may be important for incorporation of these organisms into subgingival plaque in the human oral cavity. PMID- 12117990 TI - CD40 ligand (CD154) does not contribute to lymphocyte-mediated inhibition of virulent Mycobacterium tuberculosis within human monocytes. AB - Human monocytes displayed increased expression of CD40 following infection with virulent Mycobacterium tuberculosis. Nevertheless, soluble CD40 ligand (CD40L; also designated CD154) had no effect on the intracellular growth of the organism. Restriction of the intracellular growth of M. tuberculosis by peripheral blood lymphocytes and antigen-specific CD4+ T-cell lines likewise was not reduced by blocking anti-CD40L monoclonal antibody 5c8. PMID- 12117991 TI - Role of the high-affinity zinc uptake znuABC system in Salmonella enterica serovar typhimurium virulence. AB - The Salmonella enterica serovar Typhimurium znuABC genes encoding a high-affinity zinc uptake system and its regulatory zur gene have been cloned. Salmonella serovar Typhimurium zur and znuC knockout mutants have been constructed by marker exchange. The 50% lethal dose of the znuC mutant increased when either orally or intraperitoneally inoculated in BALB/c mice, while virulence of the zur mutant decreased only when mice were intraperitoneally challenged. PMID- 12117992 TI - Alternative splicing of LYT1 transcripts in Trypanosoma cruzi. AB - As a result of alternative trans splicing, three distinct LYT1 mRNAs are produced in Trypanosoma cruzi, two encoding the full-length LYT1 protein and the third encoding a truncated LYT1 protein lacking a possible signal sequence. Analysis of the three mRNAs in different developmental forms of the parasite revealed that the alternative processing events were regulated differently during the parasite life cycle. PMID- 12117993 TI - Identification of Klebsiella pneumoniae genes involved in intestinal colonization and adhesion using signature-tagged mutagenesis. AB - Klebsiella pneumoniae is an opportunistic pathogen responsible for nosocomial infections that initially colonize the intestinal tract of patients. Signature tagged mutagenesis was used to identify genes required for this function. A library of 2,200 mutants was analyzed for the inability of the mutants to survive in a murine model of intestinal colonization and to adhere to human intestinal cells (Int-407) in vitro. Twenty-nine attenuated mutants were selected for further analyses after competition assays against the wild-type strain. Whatever the screening model, most of the transposon insertions occurred in genes involved in metabolic pathways, membrane transport, DNA metabolism, transcriptional regulation, and unknown functions. Only one mutant was attenuated in both the murine colonization and the in vitro adhesion models, and the sequence disrupted by the transposon had homology to adhesin-encoding genes of Haemophilus sp. PMID- 12117994 TI - Vibrio pathogenicity island and cholera toxin genetic element-associated virulence genes and their expression in non-O1 non-O139 strains of Vibrio cholerae. AB - A non-O1 non-O139 Vibrio cholerae strain, 10259, belonging to the serogroup O53 was shown to harbor genes related to the vibrio pathogenicity island (VPI) and a cholera toxin (CT) genetic element called CTX. While the nucleotide sequence of the strain 10259 tcpA gene differed significantly (26 and 28%) from those of O1 classical and El Tor biotype strains, respectively, partial sequence analysis data of certain other VPI-associated genes (aldA, tagA, tcpP/H, toxT, acfB/C, and int) and intergenic regions (tcpF to toxT and tcpH to tcpA) of the strain showed only minor variations (0.4 to 4.8%) from corresponding sequences in O1 strains. Strain 10259 also contained CTX element-associated toxin genes with sequences almost identical to those of O1 strains. Growth of the organism in Luria broth (LB) under ToxR inducing conditions (30 degrees C and pH 6.5) led to transcriptional activation of tcpP/H, toxR, toxT, and tcpA genes, but not of ctxA, as determined by reverse transcription-PCR (RT-PCR). Subsequent analysis revealed that strain 10259 possessed only two copies (instead of three or more copies found in epidemic-causing O1 or O139 strains) of the heptanucleotide (TTTTGAT) repeats in the intergenic region upstream of ctxAB. Therefore, a strain 10259 mutant was generated by replacement of this region with a homologous region (1.4 kb) derived from a V. cholerae O1 classical biotype strain (O395) that contained seven such repeats. The resultant recombinant strain (10259R) was found to be capable of coordinately regulated expression of toxT, ctxA, and tcpA when grown under the ToxR inducing conditions. Serological studies also demonstrated that the recombinant strain produced TcpA and a significantly ( approximately 1,000-fold) higher level of CT in vitro compared to that of the parent strain. Virulence gene expression in two other non-O1 non-O139 strains (serogroup O37) containing VPI and the CTX element was studied by RT-PCR and serological assay. One strain (S7, which was involved in an epidemic in Sudan in 1968) showed coordinately regulated expression of virulence genes leading to the production of both CT and TcpA in LB medium. However, the other strain, V2, produced RT-PCR detectable transcripts of toxT, ctxA, or tcpA genes in the early phase (6 h), but not in the late phase (16 h) of growth in LB medium. These results are consistent with the low levels of production of CT and TcpA by the strain that were serologically detectable. The significance of these results is discussed in relation to the role of virulence genes and their expression to the pathogenic potential of V. cholerae strains belonging to non-O1 serogroups. PMID- 12117995 TI - Interleukin-8 secretion from Mycobacterium tuberculosis-infected monocytes is regulated by protein tyrosine kinases but not by ERK1/2 or p38 mitogen-activated protein kinases. AB - Mycobacterium tuberculosis upregulates NF-kappaB binding and interleukin-8 (IL-8) gene expression and secretion in primary human monocytes. Inhibition of tyrosine protein kinases but not of ERK1/2 or p38 mitogen-activated protein kinases downregulates tuberculosis-induced IL-8 secretion. The inhibitor genistein decreased NF-kappaB nuclear translocation and IL-8 gene transcription in addition to acting on posttranscriptional processing. PMID- 12117996 TI - Phase variation analysis of Coxiella burnetii during serial passage in cell culture by use of monoclonal antibodies. AB - Antigenic changes in Coxiella burnetii Nine Mile strain phase I during serial passages in cell culture were analyzed with three groups of monoclonal antibodies (MAbs) against lipopolysaccharide. The MAbs of group 1 did not react with organisms that were passaged over five times, and the MAbs of group 2 did not react with organisms that were passaged over eight times. The MAbs of group 3 reacted with organisms passaged up to 15 times but did not react with phase II cells. These results suggest that C. burnetii could be differentiated into four phase states during phase variation. PMID- 12117997 TI - Mice that are congenic for the char2 locus are susceptible to malaria. AB - A major advance has been made towards the positional cloning of char2 (a quantitative trait locus encoding resistance to Plasmodium chabaudi malaria). Mice congenic for the locus have been used to fine map the gene and to prove that char2 plays a significant role in the outcome of malarial infection, independently of other resistance loci. PMID- 12117999 TI - Hsp90 interactions and acylation target the G protein Galpha 12 but not Galpha 13 to lipid rafts. AB - The heterotrimeric G proteins, G(12) and G(13), are closely related in their sequences, signaling partners, and cellular effects such as oncogenic transformation and cytoskeletal reorganization. Yet G(12) and G(13) can act through different pathways, bind different proteins, and show opposing actions on some effectors. We investigated the compartmentalization of G(12) and G(13) at the membrane because other G proteins reside in lipid rafts, membrane microdomains enriched in cholesterol and sphingolipids. Lipid rafts were isolated after cold, nonionic detergent extraction of cells and gradient centrifugation. Galpha(12) was in the lipid raft fractions, whereas Galpha(13) was not associated with lipid rafts. Mutation of Cys-11 on Galpha(12), which prevents its palmitoylation, partially shifted Galpha(12) from the lipid rafts. Geldanamycin treatment, which specifically inhibits Hsp90, caused a partial loss of wild-type Galpha(12) and a complete loss of the Cys-11 mutant from the lipid rafts and the appearance of a higher molecular weight form of Galpha(12) in the soluble fractions. These results indicate that acylation and Hsp90 interactions localized Galpha(12) to lipid rafts. Hsp90 may act as both a scaffold and chaperone to maintain a functional Galpha(12) only in discrete membrane domains and thereby explain some of the nonoverlapping functions of G(12) and G(13) and control of these potent cell regulators. PMID- 12118000 TI - Peroxisome proliferator-activated receptor alpha (PPARalpha ) turnover by the ubiquitin-proteasome system controls the ligand-induced expression level of its target genes. AB - Peroxisome proliferator activated-receptor alpha (PPARalpha) is a ligand activated transcription factor belonging to the nuclear receptor family. PPARalpha is implicated in the regulation of lipid and glucose metabolism and in the control of inflammatory response. Recently, it has been demonstrated that a number of nuclear receptors are degraded by the ubiquitin-proteasome pathway. Since PPARalpha exhibits a circadian expression rhythm and since PPARalpha is rapidly regulated under certain pathophysiological conditions such as the acute phase inflammatory response, we hypothesized that PPARalpha protein levels must be under tight control. Here, we studied the mechanisms controlling PPARalpha protein levels and their consequences on the transcriptional control of PPARalpha target genes. Using pulse-chase experiments, it is shown that PPARalpha is a short-lived protein and that addition of its ligands stabilizes this nuclear receptor. By transient cotransfection experiments using expression vectors for PPARalpha and hemagglutinin-tagged ubiquitin, it is demonstrated that PPARalpha protein is ubiquitinated and that its ligands decrease the ubiquitination of this nuclear receptor, thus providing a mechanism for the ligand-dependent stabilization observed in pulse-chase experiments. In addition, treatment with MG132, a selective proteasome inhibitor, increases the level of ubiquitinated PPARalpha and inhibits its degradation in transfected cells. Furthermore, MG132 treatment enhances the level of endogenous PPARalpha in HepG2 cells. Finally, transient transfection and quantitative reverse transcription-PCR show that inhibition of PPARalpha degradation increases its transcriptional activation and expression of target genes such as apoA-II and fatty acid transport protein (FATP). Taken together, these data demonstrate that PPARalpha is degraded by the ubiquitin-proteasome system in a ligand-dependent manner. Regulation of its degradation provides a novel regulatory mechanism of transcriptional activity of this nuclear receptor. PMID- 12118001 TI - Evidence for a stabilizer element in the untranslated regions of Drosophila glutathione S-transferase D1 mRNA. AB - The neighboring genes gstD1 and gstD21 share 70% sequence identity. gstD1 encodes a 1,1,1-trichloro-2,2-bis-(P-chlorophenyl)ethane dehydrochlorinase; gstD21, a ligandin. Both of their mRNAs are inducible by pentobarbital but otherwise behave very differently. Intact gstD21 mRNA is intrinsically labile, but becomes stabilized when separated from its native untranslated region (UTR). In contrast, whereas gstD1 mRNA is very stable in its entirety, without its native UTRs it becomes even more labile than that of gstD21. Decay patterns from four chimeric D1-D21 mRNAs, designed to reveal the individual importance of each molecular region to stability, strongly indicate the presence of destabilizing elements in the coding region of gstD1 mRNA. Thus, the UTRs of this molecule must contain a dominant stabilizer element that overrides the destabilizing influence of the coding region and confers overall stability to the entire molecule. The suspected presence of such a stabilizer element in gstD1 mRNA extends a concept from mRNA metabolism in yeast and cultured mammalian cells to include a multicellular organism, Drosophila melanogaster. The complementary presence of destabilizing and stabilizer elements on the same mRNA reveals a regulatory mechanism by which an abundant mRNA can be further induced by a chemical stimulus, or otherwise be returned to normal levels during recovery. PMID- 12118002 TI - Ribonuclease L proteolysis in peripheral blood mononuclear cells of chronic fatigue syndrome patients. AB - A 37-kDa binding polypeptide accumulates in peripheral blood mononuclear cell (PBMC) extracts from chronic fatigue syndrome (CFS) patients and is being considered as a potential diagnostic marker (De Meirleir, K., Bisbal, C., Campine, I., De Becker, P., Salehzada, T., Demettre, E., and Lebleu, B. (2000) Am. J. Med. 108, 99-105). We establish here that this low molecular weight 2-5A binding polypeptide is a truncated form of the native 2-5A-dependent ribonuclease L (RNase L), generated by an increased proteolytic activity in CFS PBMC extracts. RNase L proteolysis in CFS PBMC extracts can be mimicked in a model system in which recombinant RNase L is treated with human leukocyte elastase. RNase L proteolysis leads to the accumulation of two major fragments with molecular masses of 37 and 30 kDa. The 37-kDa fragment includes the 2-5A binding site and the N-terminal end of native RNase L. The 30-kDa fragment includes the catalytic site in the C-terminal part of RNase L. Interestingly, RNase L remains active and 2-5A-dependent when degraded into its 30- and 37-kDa fragments by proteases of CFS PBMC extract or by purified human leukocyte elastase. The 2-5A-dependent nuclease activity of the truncated RNase L could result from the association of these digestion products, as suggested in pull down experiments. PMID- 12118003 TI - The functionally distinct hemoglobins of the Arctic spotted wolffish Anarhichas minor. AB - The Arctic fish Anarhichas minor, a benthic sedentary species, displays high hemoglobin multiplicity. The three major hemoglobins (Hb 1, Hb 2, and Hb 3) show important functional differences in pH and organophosphate regulation, subunit cooperativity, and response of oxygen binding to temperature. Hb 1 and Hb 2 display a low, effector-enhanced Bohr effect and no Root effect. In contrast, Hb 3 displays pronounced Bohr and Root effects, accompanied by strong organophosphate regulation. Hb 1 has the beta (beta(1)) chain in common with Hb 2; Hb 3 and Hb 2 share the alpha (alpha(2)) chain. The amino acid sequences have been established. Several substitutions in crucial positions were observed, such as Cys in place of C-terminal His in the beta(1) chain of Hb 1 and Hb 2. In Hb 3, Val E11 of the beta(2) chain is replaced by Ile. Homology modeling revealed an unusual structure of the Hb 3 binding site of inositol hexakisphoshate. Phylogenetic analysis indicated that only Hb 2 displays higher overall similarity with the major Antarctic hemoglobins. The oxygen transport system of A. minor differs remarkably from those of Antarctic Notothenioidei, indicating distinct evolutionary pathways in the regulatory mechanisms of the fish respiratory system in the two polar environments. PMID- 12118004 TI - Downstream codons in the retinoic acid receptor beta -2 and beta -4 mRNAs initiate translation of a protein isoform that disrupts retinoid-activated transcription. AB - Retinoic acid receptors (RARs) are essential for the differentiation and maintenance of normal epithelium. In studies of RARs in breast cancer, there are striking differences in the expression of certain protein isoforms of the RARbeta gene between cells derived from normal human mammary glands and those derived from breast tumors. While the protein isoforms RARbeta2 and RARbeta4 consist of the longest open reading frames of the RARbeta2 and RARbeta4 mRNAs, respectively, we find that a fraction of scanning ribosomes bypass these upstream RARbeta2 and RARbeta4 protein start codons and initiate translation downstream. This downstream translation initiation site is identical in the RARbeta2 and RARbeta4 transcripts and generates a third RARbeta protein isoform, here termed RARbeta' (formerly human RARbeta4). RARbeta' lacks protein domains found in the N terminus of RARbeta2 and RARbeta4, including one of two zinc fingers required for DNA binding. However, RARbeta' retains the ability to heterodimerize with RXRalpha and interact with transcription cofactors. In reporter gene assays, RARbeta' repressed retinoic acid-activated transcription of co-transfected RARbeta2, RARbeta4, and RARalpha. This repression required the presence of acidic amino acids within the AF2 domain. These findings demonstrate an antagonistic role for RARbeta' in signaling by retinoic acid. PMID- 12118005 TI - Post-transcriptional control of chloroplast gene expression. Accumulation of stable psaC mRNA is due to downstream RNA cleavages in the ndhD gene. AB - Intergenic cleavages, intron splicing, and editing of primary transcripts of the plastid ndhH-D operon produce multiple overlapping RNAs, of which the most abundant by far is the monocistronic 400-nucleotide mRNA of psaC (encoding the PsaC protein of photosystem I), in contrast with the low level of transcripts of the six ndh genes. Like other plastid operons containing genes for functionally unrelated proteins, the contrasting accumulation of ndh and psaC transcripts provides a model to investigate the mechanisms of the post-transcriptional control of gene expression, a feature of chloroplast genetic machinery, with a minimum of interference by transcriptional control. In leek (Allium porrum L), the ndhD transcript (which follows the psaC gene and ends the ndhH-D operon) requires C --> U editing to restore its start codon and may be used as a marker for the processing of psaC and ndhD transcripts. By determining the editing state and 5' end sequences of specific transcripts, we demonstrated that stable monocistronic psaC mRNA results from downstream cleavages in the ndhD sequence, which renders non-functional ndhD transcripts as by-products. Alternative psaC ndhD intergenic cleavages produce complete mRNAs for both genes, but only take place in precursors containing editing-restored ndhD start codons. Hence, post transcriptional control acts by promoting the ndhD cleavage alternative, which allows the accumulation of psaC mRNA at the expense of ndhD mRNA levels. PMID- 12118006 TI - Epigallocatechin gallate, a constituent of green tea, represses hepatic glucose production. AB - Herbs have been used for medicinal purposes, including the treatment of diabetes, for centuries. Plants containing flavonoids are used to treat diabetes in Indian medicine and the green tea flavonoid, epigallocatechin gallate (EGCG), is reported to have glucose-lowering effects in animals. We show here that the regulation of hepatic glucose production is decreased by EGCG. Furthermore, like insulin, EGCG increases tyrosine phosphorylation of the insulin receptor and insulin receptor substrate-1 (IRS-1), and it reduces phosphoenolpyruvate carboxykinase gene expression in a phosphoinositide 3-kinase-dependent manner. EGCG also mimics insulin by increasing phosphoinositide 3-kinase, mitogen activated protein kinase, and p70(s6k) activity. EGCG differs from insulin, however, in that it affects several insulin-activated kinases with slower kinetics. Furthermore, EGCG regulates genes that encode gluconeogenic enzymes and protein-tyrosine phosphorylation by modulating the redox state of the cell. These results demonstrate that changes in the redox state may have beneficial effects for the treatment of diabetes and suggest a potential role for EGCG, or derivatives, as an antidiabetic agent. PMID- 12118008 TI - Macrophage foam cell formation with native low density lipoprotein. AB - This investigation has elucidated a mechanism for development of macrophage foam cells when macrophages are incubated with native low density lipoprotein (LDL). LDL is believed to be the main source of cholesterol that accumulates in monocyte derived macrophages within atherosclerotic plaques, but native LDL has not previously been shown to cause substantial cholesterol accumulation when incubated with macrophages. We have found that activation of human monocyte derived macrophages with phorbol 12-myristate 13-acetate (PMA) stimulates LDL uptake and degradation and acyl-CoA:cholesterol acyltransferase-mediated esterification of LDL-derived cholesterol, resulting in massive macrophage cholesterol accumulation that could exceed 400 nmol/mg of cell protein. Cholesterol accumulation showed a biphasic linear LDL concentration dependence with LDL levels as high as 4 mg/ml, similar to LDL levels in artery intima. Protein kinase C mediated the PMA-stimulated macrophage uptake of LDL because the protein kinase C inhibitors, Go6983 and GF109203X, inhibited cholesterol accumulation. LDL receptors did not mediate macrophage cholesterol accumulation because accumulation occurred with reductively methylated LDL and in the presence of an anti-LDL receptor-blocking monoclonal antibody. LDL-induced cholesterol accumulation was not inhibited by antioxidants, was not accompanied by increased LDL binding to macrophages, did not depend on the apoB component of LDL, and was not down-regulated by prior cholesterol enrichment of macrophages. We have shown that the mechanism of LDL uptake by macrophages was PMA-stimulated endocytosis of LDL taken up as part of the bulk phase fluid (i.e. fluid phase endocytosis). The amount of LDL taken up with the bulk phase fluid was measured with [(3)H]sucrose and accounted for a minimum of 83% of the LDL cholesterol delivery and accumulation in PMA-activated macrophages. This novel mechanism of macrophage cholesterol accumulation shows that modification of LDL is not necessary for foam cell formation to occur. In addition, the findings direct attention to macrophage fluid phase endocytosis as a relevant pathway to target for modulating macrophage cholesterol accumulation in atherosclerosis. PMID- 12118007 TI - Mutation of tyrosine 332 to phenylalanine converts dopa decarboxylase into a decarboxylation-dependent oxidative deaminase. AB - A flexible loop (residues 328-339), presumably covering the active site upon substrate binding, has been revealed in 3,4-dihydroxyphenylalanine decarboxylase by means of kinetic and structural studies. The function of tyrosine 332 has been investigated by substituting it with phenylalanine. Y332F displays coenzyme content and spectroscopic features identical to those of the wild type. Unlike wild type, during reactions with l-aromatic amino acids under both aerobic and anaerobic conditions, Y332F does not catalyze the formation of aromatic amines. However, analysis of the products shows that in aerobiosis, l-aromatic amino acids are converted into the corresponding aromatic aldehydes, ammonia, and CO(2) with concomitant O(2) consumption. Therefore, substitution of Tyr-332 with phenylalanine results in the suppression of the original activity and in the generation of a decarboxylation-dependent oxidative deaminase activity. In anaerobiosis, Y332F catalyzes exclusively a decarboxylation-dependent transamination of l-aromatic amino acids. A role of Tyr-332 in the Calpha protonation step that catalyzes the formation of physiological products has been proposed. Furthermore, Y332F catalyzes oxidative deamination of aromatic amines and half-transamination of d-aromatic amino acids with k(cat) values comparable with those of the wild type. However, for all the mutant-catalyzed reactions, an increase in K(m) values is observed, suggesting that Y --> F replacement also affects substrate binding. PMID- 12118009 TI - Identification of rare partially unfolded states in equilibrium with the native conformation in an all beta-barrel protein. AB - Human acidic fibroblast growth factor 1 (hFGF-1) is an all beta-barrel protein, and the secondary structural elements in the protein include 12 antiparallel beta strands arranged into a beta-trefoil fold. In the present study, we investigate the stability of hFGF-1 by hydrogen-deuterium exchange as a function of urea concentration. Urea-induced equilibrium unfolding of hFGF-1 monitored by fluorescence and CD spectroscopy suggests that the protein unfolds by a two-state (native to denatured) mechanism. Hydrogen exchange in hFGF-1, under the experimental conditions used, occurs by the EX2 mechanism. In contrast to the equilibrium unfolding events monitored by optical probes, native state hydrogen exchange data show that the beta-trefoil architecture of hFGF-1 does not behave as a single cooperative unit. There are at least two structurally independent units with differing stabilities in hFGF-1. Beta-strands I, II, III, VI, VII, X, XI, and XII fit into the global unfolding isotherm. By contrast, residues in beta strands IV, V, VIII, and IX exchange by the subfolding isotherm and could be responsible for the occurrence of high-energy partially unfolded state(s) in hFGF 1. There appears to be a broad continuum of stabilities among the four beta strands (beta-strands IV, V, VIII, and IX) constituting the subglobal folding unit. The slow exchanging residues in hFGF-1 do not represent the folding nucleus of the protein. PMID- 12118010 TI - The role of electrostatic interactions in human serum albumin binding and stabilization by halothane. AB - Electrostatic interactions have been proposed as a potentially important force for anesthetics and protein binding but have not yet been tested directly. In the present study, we used wild-type human serum albumin (HSA) and specific site directed mutants as a native protein model to investigate the role of electrostatic interactions in halothane binding. Structural geometry analysis of the HSA-halothane complex predicted an absence of significant electrostatic interactions, and direct binding (tryptophan fluorescence and zonal elution chromatography) and stability experiments (hydrogen exchange) confirmed that loss of charge in the binding sites, by charged to uncharged mutations and by changing ionic strength of the buffer, generally increased both regional (tryptophan region) and global halothane/HSA affinity. The results indicate that electrostatic interactions (full charges) either do not contribute or diminish halothane binding to HSA, leaving only the more general hydrophobic and van der Waals forces as the major contributors to the binding interaction. PMID- 12118011 TI - Identification of a neuropeptide modified with bromine as an endogenous ligand for GPR7. AB - We isolated a novel gene in a search of the Celera data base and found that it encoded a peptidic ligand for a G protein-coupled receptor, GPR7 (O'Dowd, B. F., Scheideler, M. A., Nguyen, T., Cheng, R., Rasmussen, J. S., Marchese, A., Zastawny, R., Heng, H. H., Tsui, L. C., Shi, X., Asa, S., Puy, L., and George, S. R. (1995) Genomics 28, 84-91; Lee, D. K., Nguyen, T., Porter, C. A., Cheng, R., George, S. R., and O'Dowd, B. F. (1999) Mol. Brain Res. 71, 96-103). The expression of this gene was detected in various tissues in rats, including the lymphoid organs, central nervous system, mammary glands, and uterus. GPR7 mRNA was mainly detected in the central nervous system and uterus. In situ hybridization showed that the gene encoding the GPR7 ligand was expressed in the hypothalamus and hippocampus of rats. To determine the molecular structure of the endogenous GPR7 ligand, we purified it from bovine hypothalamic tissue extracts on the basis of cAMP production-inhibitory activity to cells expressing GPR7. Through structural analyses, we found that the purified endogenous ligand was a peptide with 29 amino acid residues and that it was uniquely modified with bromine. We subsequently determined that the C-6 position of the indole moiety in the N-terminal Trp was brominated. We believe this is the first report on a neuropeptide modified with bromine and have hence named it neuropeptide B. In in vitro assays, bromination did not influence the binding of neuropeptide B to the receptor. PMID- 12118012 TI - Trafficking of ganglioside GD3 to mitochondria by tumor necrosis factor-alpha. AB - The interaction of mitochondria with proapoptotic proteins activates apoptosis pathways. Previous findings have identified ganglioside GD3 (GD3) as an emerging apoptotic lipid intermediate that targets mitochondria in response to death signals. Using immunoelectron and laser scanning confocal microscopy, we characterize the trafficking of GD3 to mitochondria in response to tumor necrosis factor-alpha (TNF-alpha) in rat hepatocytes. In control hepatocytes, GD3 is present predominantly at the plasma membrane as well as in the endosomal/Golgi network, as verified by its colocalization with the asialoglycoprotein receptor. Following TNF-alpha exposure, GD3 undergoes a rapid cellular redistribution with a gradual loss from the plasma membrane before its colocalization with mitochondria. This process is mimicked by acidic sphingomyelinase and ionizing radiation but not by neutral sphingomyelinase or staurosporin. TNF-alpha stimulated the colocalization of GD3 with early and late endosomal markers, Rab 5 and Rab 7, whereas perturbation of plasma membrane cholesterol or actin cytoskeleton or inhibition of glucosylceramide synthase prevented the trafficking of GD3 to mitochondria. Finally, prevention of the TNF-alpha-stimulated neosynthesis of GD3, cyclosporin A, and latrunculin A or filipin protected sensitized hepatocytes from TNF-alpha-mediated cell death. Thus, the intracellular redistribution and mitochondrial targeting of GD3 during TNF-alpha signaling occurs through actin cytoskeleton vesicular trafficking and contributes to TNF-alpha-mediated hepatocellular cell death. PMID- 12118013 TI - Differential sensitivity of inward rectifier K+ channels to metabolic inhibitors. AB - Inhibition of inward rectifier K(+) channels under ischemic conditions may contribute to electrophysiological consequences of ischemia such as cardiac arrhythmia. Ischemia causes metabolic inhibition, and the use of metabolic inhibitors is one experimental method of simulating ischemia. The effects of metabolic inhibitors on the activity of inward rectifier K(+) channels K(ir)2.1, K(ir)2.2, and K(ir)2.3 were studied by heterologous expression in Xenopus oocytes and two-electrode voltage clamp. 10 microm carbonyl cyanide p trifluoromethoxyphenylhydrazone (FCCP) inhibited K(ir)2.2 and K(ir)2.3 currents but was without effect on K(ir)2.1 currents. The rate of decline of current in FCCP was faster for K(ir)2.3 than for K(ir)2.2. K(ir)2.3 was inhibited by 3 mm sodium azide (NaN(3)), whereas K(ir)2.1 and K(ir)2.2 were not. K(ir)2.2 was inhibited by 10 mm NaN(3). All three of these inward rectifiers were inhibited by lowering the pH of the solution perfusing inside-out membrane patches. K(ir)2.3 was most sensitive to pH (pK = 6.9), whereas K(ir)2.1 was least sensitive (pK = 5.9). For K(ir)2.2 the pK was 6.2. These results demonstrate the differential sensitivity of these inward rectifiers to metabolic inhibition and internal pH. The electrophysiological response of a particular cell type to ischemia may depend on the relative expression levels of different inward rectifier genes. PMID- 12118014 TI - The molecular determinants of ionic regulatory differences between brain and kidney Na+/Ca2+ exchanger (NCX1) isoforms. AB - The Na(+)/Ca(2+) exchanger gene NCX1 undergoes alternative splicing leading to several isoforms that differ in a small portion of the large cytoplasmic loop. This loop is involved in many regulatory processes of NCX1, including ionic regulation by the transported substrates Na(+) and Ca(2+). High intracellular Ca(2+) can alleviate intracellular Na(+)-dependent inactivation in exon A (NCX1.4)-containing isoforms but not in those containing the mutually exclusive exon B (NCX1.3). Giant excised patches from Xenopus oocytes expressing various NCX1 constructs were used to examine the specific amino acids responsible for these observed regulatory differences. Using a chimeric approach, the region responsible was narrowed down to the small central part of exon A (IDDEEYEKNKTF). Replacing the second aspartic acid of this sequence with arginine (the corresponding amino acid in exon B) in an exon A background completely prevented the effect of Ca(2+) on intracellular Na(+)-dependent inactivation. Mutating the second lysine to cysteine (exon B) had a similar, but only partial, effect. The converse double mutant, but neither single mutation alone, introduced into an exon B background (arginine to aspartic acid and cysteine to lysine) was able to restore the NCX1.4 regulatory phenotype. These data demonstrate that aspartic acid 610 and lysine 617 (using the rat NCX1.4 numbering scheme) are critical molecular determinants of the unique Ca(2+) regulatory properties of NCX1.4. PMID- 12118015 TI - The early rad catches the tumor? PMID- 12118016 TI - Radiotherapy and organ preservation in bladder cancer: are we ignoring the evidence? PMID- 12118017 TI - When quality of life is the major challenge. PMID- 12118018 TI - Phase III study of concurrent versus sequential thoracic radiotherapy in combination with cisplatin and etoposide for limited-stage small-cell lung cancer: results of the Japan Clinical Oncology Group Study 9104. AB - PURPOSE: To evaluate the optimal timing for thoracic radiotherapy (TRT) in limited-stage small-cell lung cancer (LS-SCLC), the Lung Cancer Study Group of the Japan Clinical Oncology Group conducted a phase III study in which patients were randomized to sequential TRT or concurrent TRT. PATIENTS AND METHODS: We treated 231 patients with LS-SCLC. TRT consisted of 45 Gy over 3 weeks (1.5 Gy twice daily), and the patients were randomly assigned to receive either sequential or concurrent TRT. All patients received four cycles of cisplatin plus etoposide every 3 weeks (sequential arm) or 4 weeks (concurrent arm). TRT was begun on day 2 of the first cycle of chemotherapy in the concurrent arm and after the fourth cycle in the sequential arm. RESULTS: Concurrent radiotherapy yielded better survival than sequential radiotherapy (P =.097 by log-rank test). The median survival time was 19.7 months in the sequential arm versus 27.2 months in the concurrent arm. The 2-, 3-, and 5-year survival rates for patients who received sequential radiotherapy were 35.1%, 20.2%, and 18.3%, respectively, as opposed to 54.4%, 29.8% and 23.7%, respectively, for the patients who received concurrent radiotherapy. Hematologic toxicity was more severe in the concurrent arm. However, severe esophagitis was infrequent in both arms, occurring in 9% of the patients in the concurrent arm and 4% in the sequential arm. CONCLUSION: This study strongly suggests that cisplatin plus etoposide and concurrent radiotherapy is more effective for the treatment of LS-SCLC than cisplatin plus etoposide and sequential radiotherapy. PMID- 12118019 TI - Combined-modality treatment and selective organ preservation in invasive bladder cancer: long-term results. AB - PURPOSE: To evaluate our long-term experience with combined modality treatment and selective bladder preservation and to identify factors that may predict treatment response, risk of relapse, and survival. PATIENTS AND METHODS: Between 1982 and 2000, 415 patients with bladder cancer (high-risk T1, n = 89; T2 to T4, n = 326) were treated with radiotherapy (RT; n = 126) or radiochemotherapy (RCT; n = 289) after transurethral resection (TUR) of the tumor. Six weeks after RT/RCT, response was evaluated by restaging-TUR. In case of complete response (CR), patients were observed at regular intervals. In case of persistent or recurrent invasive tumor, salvage-cystectomy was recommended. Median follow-up was 60 months (range, 6 to 199 months). RESULTS: CR was achieved in 72% of patients. Local control after CR without muscle-invasive relapse was maintained in 64% of patients at 10 years. Distant metastases were diagnosed in 98 patients with an actuarial rate of 35% at 10 years. Ten-year disease-specific survival was 42%, and more than 80% of survivors preserved their bladder. Early tumor stage and a complete TUR were the most important factors predicting CR and survival. RCT was more effective than RT alone in terms of CR and survival. Salvage cystectomy for local failure was associated with a 45% disease-specific survival rate at 10 years. Cystectomy because of a contracted bladder was restricted to 2% of patients. CONCLUSION: TUR with RCT is a reasonable option for patients seeking an alternative to radical cystectomy. Ideal candidates are those with early-stage and unifocal tumors, in whom a complete TUR is accomplished. PMID- 12118021 TI - VAMP and low-dose, involved-field radiation for children and adolescents with favorable, early-stage Hodgkin's disease: results of a prospective clinical trial. AB - PURPOSE: To evaluate outcome and assess toxicity of children and adolescents with early-stage, favorable Hodgkin's disease treated with vinblastine, doxorubicin, methotrexate, and prednisone (VAMP) and low-dose, involved-field radiation. PATIENTS AND METHODS: One hundred ten patients with clinical stages I and II, favorable (nonbulky) Hodgkin's disease were treated with four cycles of VAMP chemotherapy and 15 Gy involved-field radiation for those who achieved a complete response, or 25.5 Gy for those who achieved a partial response to two cycles of VAMP. RESULTS: With a median follow-up of 5.6 years (range, 1.1 to 10.4 years), the 5-year survival and event-free survival were 99% (lower confidence limit [CL], 97.4%) and 93% (lower CL, 88.6%), respectively. Factors associated with event-free survival of 100% were complete response to two cycles of VAMP and histology other than nodular sclerosing Hodgkin's disease (NSHD). No serious early or late toxicity has been observed. Patients presenting with clinical stages I and IIA, nonbulky disease involving fewer than three nodal sites have a projected survival and event-free survival of 100% and 97% (lower CL, 93%), respectively, at 5 years. CONCLUSION: Risk-adapted, combined-modality therapy using only four cycles of VAMP chemotherapy with 15 to 25.5 Gy of involved-field radiation for patients with early-stage/favorable Hodgkin's disease is highly effective and without demonstrable late effects. These results indicate that pediatric patients with stages I and II favorable Hodgkin's disease can be cured with limited therapy that does not include an alkylating agent, bleomycin, etoposide, or high-dose, extended-field radiation therapy. PMID- 12118020 TI - Results of a phase II study with doxorubicin, etoposide, and cisplatin in patients with fully characterized small-cell carcinoma of the prostate. AB - PURPOSE: To determine the activity and toxicity of doxorubicin in combination with cisplatin and etoposide in patients with small-cell prostate carcinoma (SCPCa) and to characterize the clinicopathologic features of SCPCa. PATIENTS AND METHODS: Patients with SCPCa (pure or mixed), measurable disease, good organ function, and no prior treatment with doxorubicin, etoposide, or cisplatin were treated every 4 weeks with doxorubicin 50 mg/m(2) as a 24-hour intravenous (IV) infusion followed by etoposide 120 mg/m(2)/d and cisplatin 25 mg/m(2)/d IV on days 2 to 4. RESULTS: Thirty-eight patients (36 assessable for response) were treated for a median of four cycles. Twenty-nine (81%) of 36 patients had prior hormonal therapy. Study patients had visceral metastases, lytic bone disease, and relatively low serum prostate-specific antigen (PSA). We observed 22 partial responses (response rate, 61% in an intent-to-treat analysis); toxicity was severe (grade 3 or 4 neutropenia 100%, thrombocytopenia 66%, mucositis 21%, and infection 68%). Three patients died of toxicity. Median time to progression and overall survival time were 5.8 months and 10.5 months, respectively. Performance status, serum albumin, and number of organs involved (but not PSA, carcinoembryonic antigen, or neuroendocrine markers) were predictors of survival. CONCLUSION: SCPCa presents unique clinicopathologic features. Addition of doxorubicin to the etoposide/cisplatin regimen caused higher toxicity in this patient population and failed to improve outcome. Given these results, we do not recommend further development of this regimen for patients with SCPCa. Improvement in therapy will come from understanding the biology of SCPCa progression and integrating new targeted therapies into the treatment of SCPCa. PMID- 12118022 TI - Treatment of unfavorable childhood Hodgkin's disease with VEPA and low-dose, involved-field radiation. AB - PURPOSE: Between January 1990 and April 1993, 56 pediatric patients with Hodgkin's disease were treated on a single-arm trial at three institutions with a regimen designed to maintain high cure rates while minimizing the potential late effects of treatment, such as infertility, second malignant neoplasms, and cardiopulmonary injury. PATIENTS AND METHODS: The regimen used combined-modality therapy with six cycles of vinblastine, etoposide, prednisone, and doxorubicin (VEPA) chemotherapy and low-dose, involved-field radiation. Unfavorable features comprised bulky presentations of localized (stage I or II) disease or advanced (stage III or IV) Hodgkin's disease. RESULTS: Of 56 patients enrolled, 26 (46%) had unfavorable presentations of stage I/II disease and 30 (54%) had advanced (stage III/IV) disease. Seventy-nine percent of the patients are alive without disease at a median follow-up time of 8.9 years from diagnosis. Nineteen patients had events at a median of 1.5 years (range, 0.4 to 7.9 years) from diagnosis; 17 patients relapsed, one died of cardiomyopathy, and one died of accidental injuries. Survival and event-free survival (EFS) estimates at 5 years for the entire cohort were 81.9% (SE, 5.2%) and 67.8% (SE, 6.3%), respectively. Five-year EFS by stage was 100% for stage I, 79.2% (SE, 8.3%) for stage II, 70% (SE, 14.5%) for stage III, and 49.5% (SE, 11.3%) for stage IV patients. CONCLUSION: Combined modality therapy with VEPA chemotherapy and low-dose, involved-field radiation is adequate for disease control of early-stage patients with unfavorable features, but it is inferior to other standard regimens for advanced-stage patients. PMID- 12118023 TI - Evaluation of HER-2/neu gene amplification and overexpression: comparison of frequently used assay methods in a molecularly characterized cohort of breast cancer specimens. AB - PURPOSE: To compare and evaluate HER-2/neu clinical assay methods. MATERIALS AND METHODS: One hundred seventeen breast cancer specimens with known HER-2/neu amplification and overexpression status were assayed with four different immunohistochemical assays and two different fluorescence in situ hybridization (FISH) assays. RESULTS: The accuracy of the FISH assays for HER-2/neu gene amplification was high, 97.4% for the Vysis PathVision assay (Vysis, Inc, Downers Grove, IL) and 95.7% for the the Ventana INFORM assay (Ventana, Medical Systems, Inc, Tucson, AZ). The immunohistochemical assay with the highest accuracy for HER 2/neu overexpression was obtained with R60 polyclonal antibody (96.6%), followed by immunohistochemical assays performed with 10H8 monoclonal antibody (95.7%), the Ventana CB11 monoclonal antibody (89.7%), and the DAKO HercepTest (88.9%; Dako, Corp, Carpinteria, CA). Only the sensitivities, and therefore, overall accuracy, of the DAKO Herceptest and Ventana CB11 immunohistochemical assays were significantly different from the more sensitive FISH assay. CONCLUSION: Based on these findings, the FISH assays were highly accurate, with immunohistochemical assays performed with R60 and 10H8 nearly as accurate. The DAKO HercepTest and the Ventana CB11 immunohistochemical assay were statistically significantly different from the Vysis FISH assay in evaluating these previously molecularly characterized breast cancer specimens. PMID- 12118024 TI - Effects on quality of life of combined trastuzumab and chemotherapy in women with metastatic breast cancer. AB - PURPOSE: The study was designed to compare the effects of treatment with a combination of trastuzumab (Herceptin; Genentech, Inc, South San Francisco, CA) and chemotherapy versus chemotherapy alone on health-related quality of life (HRQL) in patients with HER-2/neu overexpressing, metastatic breast cancer. PATIENTS AND METHODS: A sample of 400 patients, not previously treated for metastatic disease and randomized to receive either trastuzumab plus chemotherapy (208 patients) or chemotherapy alone (192 patients), completed the European Organization for Research and Treatment Care Quality of Life Questionnaire at baseline and on at least one subsequent occasion at 8, 20, 32, 44, and 56 weeks. HRQL improvement or worsening was defined as a >or= 10-point change (range, 0 to 100 points) in the scores of six preselected domains (global quality of life [QOL], physical, role, social, and emotional functioning, and fatigue). Stable HRQL was defined as a change of less than 10. A Bonferroni correction was applied for multiple testing. RESULTS: After completion of chemotherapy, patients treated with trastuzumab and chemotherapy reported significant improvement in fatigue (P <.05) as compared with their baseline scores. Higher proportions of patients receiving the combined therapy achieved improvement in global QOL (P <.05) than did patients treated with chemotherapy alone. Higher proportions of the combined therapy group also achieved improvement in physical and role functioning and in fatigue as compared with the chemotherapy group, but the differences were not statistically significant. There were no differences in the proportions of patients in the two groups that reported worsening. CONCLUSION: Statistically significantly higher proportions of patients treated with a combination of trastuzumab and chemotherapy reported improved global QOL than did patients treated by chemotherapy alone. PMID- 12118025 TI - Doxorubicin and paclitaxel versus doxorubicin and cyclophosphamide as first-line chemotherapy in metastatic breast cancer: The European Organization for Research and Treatment of Cancer 10961 Multicenter Phase III Trial. AB - PURPOSE: To compare the efficacy and tolerability of the combination of doxorubicin and paclitaxel (AT) with a standard doxorubicin and cyclophosphamide (AC) regimen as first-line chemotherapy for metastatic breast cancer. PATIENTS AND METHODS: Eligible patients were anthracycline-naive and had bidimensionally measurable metastatic breast cancer. Two hundred seventy-five patients were randomly assigned to be treated with AT (doxorubicin 60 mg/m(2) as an intravenous bolus plus paclitaxel 175 mg/m(2) as a 3-hour infusion) or AC (doxorubicin 60 mg/m(2) plus cyclophosphamide 600 mg/m(2)) every 3 weeks for a maximum of six cycles. A paclitaxel (200 mg/m(2)) and cyclophosphamide (750 mg/m(2)) dose escalation was planned at cycle 2 if no grade >or= 3 neutropenia occurred in cycle 1. The primary efficacy end point was progression-free survival (PFS). Secondary end points were response rate (RR), safety, overall survival (OS), and quality of life. RESULTS: A median number of six cycles were delivered in the two treatment arms. The relative dose-intensity and delivered cumulative dose of doxorubicin were lower in the AT arm. Dose escalation was only possible in 17% and 20% of the AT and AC patients, respectively. Median PFS was 6 months in the two treatments arms. RR was 58% versus 54%, and median OS was 20.6 versus 20.5 months in the AT and AC arms, respectively. The AT regimen was characterized by a higher incidence of febrile neutropenia, 32% versus 9% in the AC arm. CONCLUSION: No differences in the efficacy study end points were observed between the two treatment arms. Treatment-related toxicity compromised doxorubicin-delivered dose intensity in the paclitaxel-based regimen PMID- 12118027 TI - Multicenter randomized phase III trial comparing protracted venous infusion (PVI) fluorouracil (5-FU) with PVI 5-FU plus mitomycin in inoperable pancreatic cancer. AB - PURPOSE: To compare protracted venous infusion (PVI) fluorouracil (5-FU) with PVI 5-FU plus mitomycin (MMC) in patients with advanced pancreatic cancer in a multicenter, prospectively randomized study. PATIENTS AND METHODS: Two hundred eight patients were randomized to PVI 5-FU (300 mg/m(2)/d for a maximum of 24 weeks) or PVI 5-FU plus MMC (7 mg/m(2) every 6 weeks for four courses). The major end points were tumor response, survival, toxicity, and quality of life (QOL). RESULTS: The two treatment groups were balanced for baseline demographic factors, and 62% had metastatic disease. The overall response rate was 8.4% (95% confidence interval [CI]) 3.2% to 13.7% for patients treated with PVI 5-FU alone compared with 17.6%; 95% CI 10.3% to 25.1% for PVI 5-FU plus MMC (P =.04). Median failure-free survival was 2.8 months for PVI 5-FU and 3.8 months for PVI 5-FU plus MMC (P =.14). Median survival was 5.1 months for PVI 5-FU and 6.5 months for PVI 5-FU plus MMC (P =.34). Toxicities in both arms were mild. There was an increased incidence of neutropenia in the 5-FU plus MMC arm (P <.01), although no differences in infection were seen. No patients developed hemolytic uremic syndrome. Global QOL improved significantly after 24 weeks of treatment compared with baseline for patients receiving 5-FU plus MMC, although there was no statistically significant difference in QOL between arms. CONCLUSION: PVI 5-FU plus MMC resulted in a superior response rate in comparison with PVI 5-FU alone in advanced pancreatic cancer, but this did not translate into a survival advantage. These results emphasize the importance of chemotherapy in this setting and the continuing value of the fluoropyrimidines in pancreatic cancer. PMID- 12118026 TI - Modulation of irinotecan metabolism by ketoconazole. AB - PURPOSE: Irinotecan (CPT-11) is a prodrug of SN-38 and has been registered for the treatment of advanced colorectal cancer. It is converted by the cytochrome P450 3A4 isozyme (CYP3A4) into several inactive metabolites, including 7-ethyl-10 [4-N-(5-aminopentanoic acid)-1-piperidino]-carbonyloxycamptothecin (APC). To investigate the role of CYP3A4 in irinotecan pharmacology, we evaluated the consequences of simultaneous treatment of irinotecan with a potent enzyme inhibitor, ketoconazole, in a group of cancer patients. PATIENTS AND METHODS: A total of seven assessable patients was treated in a randomized, cross-over design with irinotecan (350 mg/m(2) intravenously for 90 minutes) given alone and followed 3 weeks later by irinotecan (100 mg/m(2)) in combination with ketoconazole (200 mg orally for 2 days) or vice versa. Serial plasma, urine, and feces samples were obtained up to 500 hours after dosing and analyzed for irinotecan, metabolites (7-ethyl-10-hydroxycamptothecin [SN-38], SN-38 glucuronide [SN-38G], and APC), and ketoconazole by high-performance liquid chromatography. RESULTS: With ketoconazole coadministration, the relative formation of APC was reduced by 87% (P =.002), whereas the relative exposure to the carboxylesterase-mediated SN-38 as expected on the basis of dose (area under the plasma concentration-time curve normalized to dose) was increased by 109% (P =.004). These metabolic alterations occurred without substantial changes in irinotecan clearance (P =.90) and formation of SN-38G (P =.93). CONCLUSION: Inhibition of CYP3A4 in cancer patients treated with irinotecan leads to significantly increased formation of SN-38. Simultaneous administration of various commonly prescribed inhibitors of CYP3A4 can potentially result in fatal outcomes, and up to four-fold reductions in irinotecan dose are indicated. PMID- 12118028 TI - Anxiety disorders in cancer patients: their nature, associations, and relation to quality of life. AB - PURPOSE: We aimed to estimate the prevalence and types of anxiety disorders diagnosed according to standardized criteria in cancer patients, to compare screening tools in detecting them, and to examine their demographic, oncologic, and psychosocial associations. METHODS: In this cross-sectional observational study of 178 subjects with lymphoma, renal cell carcinoma, malignant melanoma, or plasma cell dyscrasia, we related responses to questionnaires (administered by computer touch-screen) measuring psychological symptoms, quality of life (QOL), and social support to standardized psychiatric interviews and cancer management. RESULTS: Forty-eight percent of subjects reported sufficient anxiety for anxiety disorder to be considered. At subsequent diagnostic interview, 18% fulfilled International Classification of Disorders, 10th Revision criteria for anxiety disorder, including 6% of patients who reported low levels of anxiety by questionnaire. When subjects reported anxiety by questionnaire, if disruptive somatic anxiety was present, this increased the probability of diagnosable anxiety disorder from.31 to.7. The most accurate screening questionnaires were the trait scale of the State-Trait Anxiety Inventory and the Hospital Anxiety and Depression scale. Female sex and negative aspects of social support were associated with anxiety disorder in multivariate analyses. Anxiety disorder was independently associated with a deficit in QOL, particularly insomnia. CONCLUSION: Anxiety symptoms are common in cancer patients. Screening by questionnaire seems to assess anxiety symptoms adequately but discriminates abnormal anxiety inadequately. To improve this, we may need to use criteria such as disruption from anxiety, as illustrated by the impact of anxiety disorders on QOL. There seem to be few oncologic variables that could target screening for anxiety disorders. PMID- 12118029 TI - Survival of patients with newly diagnosed glioblastoma multiforme treated with RSR13 and radiotherapy: results of a phase II new approaches to brain tumor therapy CNS consortium safety and efficacy study. AB - PURPOSE: The objectives of this phase II study were to determine survival, safety, pharmacokinetics (PK), and pharmacodynamics (PD) of 2,4-[[(3,5 dimethylanilino)carbonyl]methyl]phenoxy]-2-methylpropionic acid (RSR13, efaproxiral) 100 mg/kg per day administered with standard cranial radiotherapy (RT) for the treatment of glioblastoma multiforme (GBM). RSR13, a synthetic allosteric modifier of hemoglobin, is a radiation-enhancing agent that noncovalently binds to hemoglobin, reduces oxygen-binding affinity, and increases oxygen unloading to hypoxic tissue. PATIENTS AND METHODS: Fifty patients with newly diagnosed GBM (Karnofsky performance status >or= 60) were enrolled onto this multicenter phase II study. Patients received daily RSR13 100 mg/kg intravenously infused for 30 minutes immediately before cranial RT (60 Gy in 30 fractions). Supplemental oxygen was given during RSR13 infusion and continued until after the RT treatment was completed. RT was given within 30 minutes of the end of RSR13 infusion. PK and PD determinations were performed. RESULTS: The median survival for the RSR13-treated patients was 12.3 months with 1-year and 18 month survival rates of 54% and 24%, respectively. Twenty-four percent of patients had greater than grade 2 toxicity, which was generally transient and self-limited. A significant PD effect on hemoglobin-oxygen binding affinity was demonstrated for most patients. CONCLUSION: RSR13 (100 mg/kg) administered immediately before cranial RT is well tolerated and is pharmacodynamically active. Median survival in excess of 1 year is favorable. PMID- 12118030 TI - Response to neoadjuvant chemotherapy combined with regional hyperthermia predicts long-term survival for adult patients with retroperitoneal and visceral high-risk soft tissue sarcomas. AB - PURPOSE: To determine the efficacy of neoadjuvant chemotherapy combined with regional hyperthermia (RHT) for local tumor control and overall survival (OS) in adult patients with retroperitoneal or visceral (RP/V) high-risk soft tissue sarcomas (HR-STS). PATIENTS AND METHODS: From 1991 to 1997, 58 patients with HR STS at RP/V sites were prospectively treated with four cycles of etoposide, ifosfamide, and doxorubicin combined with RHT followed by surgery, adjuvant chemotherapy, and radiation. RESULTS: Objective response rate assessable in 40 patients was 13% (five partial responses). Including minor responses (n = 8), the radiographic response rate was 33%. The pathologic response rate assessable in 26 patients after surgical resection was 42%. Median OS was 31 months. At a median observation time of 74 months, 5-year probability of local failure-free survival (LFFS), distant metastasis-free survival, event-free survival, and OS were 25%, 51%, 20%, and 32%, respectively. Averaged minimum temperatures (T(min)) and time averaged temperatures achieved in 50% (T(50)) and 90% (T(90)) of all measured tumor sites differed significantly between responders and nonresponders (T(min), 39.3 degrees C v 38.0 degrees C; P =.002; T(50), 40.9 degrees C v 40.3 degrees C; P =.038; T(90), 40.1 degrees C v 39.3 degrees C; P =.017). At 5-year follow-up, probability of LFFS (59% v 0%; P <.001) and OS (60% v 10%; P <.001) was significantly in favor of patients responding to neoadjuvant thermochemotherapy. CONCLUSION: Response to neoadjuvant chemotherapy combined with RHT is predictive for an improved local tumor control resulting in a long-term survival benefit for patients with HR-STS at unfavorable RP/V sites; however, the impact of RHT has to be defined in a randomized phase III trial. PMID- 12118031 TI - Microarrays as cancer keys: an array of possibilities. AB - Malignant transformation results from accumulation of genetic and epigenetic events. Functional studies of cancer will be crucial to our understanding of its complexity and polymorphism. There is no doubt that emerging genomic and proteomic technologies will facilitate such investigations. Microarray technology is a new and efficient approach to extract data of biomedical relevance for a wide range of applications. In cancer research, it will provide high-throughput and valuable insights into differences in an individual's tumor as compared with constitutional DNA, mRNA expression, and protein expression and activity. Across individuals, comparisons could provide tissue-specific disease signatures that provide diagnosis based on hundreds of informative genes. The resulting product should be a wealth of tumor-associated and tumor-specific biomarkers, which may help in cancer etiology, diagnosis, and therapy and ultimately lead to "molecular nosology" of cancers. This review highlights the recent developments in microarray technologies in cancer research, focuses on the results obtained so far, and describes the eventual use of microarray technology for clinical applications. PMID- 12118032 TI - Phase I trial of adoptive immunotherapy with cytolytic T lymphocytes immunized against a tyrosinase epitope. PMID- 12118033 TI - Missense mismatch repair gene alterations, microsatellite instability, and hereditary nonpolyposis colorectal cancer. PMID- 12118034 TI - Treatment of newly diagnosed glioblastoma multiforme. PMID- 12118035 TI - Promising survival and concomitant radiation plus temozolomide followed by adjuvant temozolomide. PMID- 12118036 TI - More concern about transfusion requirement when evaluating quality of life in anemic patients. PMID- 12118037 TI - Carbohydrate Metabolism: Glycogen Phosphorylase and the Work of Carl F. and Gerty T.Cori. 1928-1943. PMID- 12118038 TI - Fatty acid homeostasis and induction of lipid regulatory genes in skeletal muscles of peroxisome proliferator-activated receptor (PPAR) alpha knock-out mice. Evidence for compensatory regulation by PPAR delta. AB - Ablation of peroxisome proliferator activated receptor (PPAR) alpha, a lipid activated transcription factor that regulates expression of beta-oxidative genes, results in profound metabolic abnormalities in liver and heart. In the present study we used PPAR alpha knockout (KO) mice to determine whether this transcription factor is essential for regulating fuel metabolism in skeletal muscle. When animals were challenged with exhaustive exercise or starvation, KO mice exhibited lower serum levels of glucose, lactate, and ketones and higher nonesterified fatty acids than wild type (WT) littermates. During exercise, KO mice exhausted earlier than WT and exhibited greater rates of glycogen depletion in liver but not skeletal muscle. Fatty acid oxidative capacity was similar between muscles of WT and KO when animals were fed and only 28% lower in KO muscles when animals were starved. Exercise-induced regulation and starvation induced regulation of pyruvate-dehydrogenase kinase 4 and uncoupling protein 3, two classical and robustly responsive PPAR alpha target genes, were similar between WT and KO in skeletal muscle but markedly different between genotypes in heart. Real time quantitative PCR analyses showed that unlike in liver and heart, in mouse skeletal muscle PPAR delta is severalfold more abundant than either PPAR alpha or PPAR gamma. In both human and rodent myocytes, the highly selective PPAR delta agonist GW742 increased fatty acid oxidation about 2-fold and induced expression of several lipid regulatory genes, including pyruvate-dehydrogenase kinase 4 and uncoupling protein 3, responses that were similar to those elicited by the PPAR alpha agonist GW647. These results show redundancy in the functions of PPARs alpha and delta as transcriptional regulators of fatty acid homeostasis and suggest that in skeletal muscle high levels of the delta-subtype can compensate for deficiency of PPAR alpha. PMID- 12118039 TI - A point mutation of the AF2 transactivation domain of the glucocorticoid receptor disrupts its interaction with steroid receptor coactivator 1. AB - Glucocorticoids cause a 10-fold increase in hepatic phosphoenolpyruvate carboxykinase (PEPCK) gene transcription through two low affinity glucocorticoid receptor (GR) binding sites and a complex array of accessory factor DNA elements and associated proteins. To analyze how co-activators interact with the GR in this context, we took advantage of the C656G GR mutant that binds ligand with very high affinity. This GR activates PEPCK gene transcription at a 500-fold lower dexamethasone concentration than does wild type GR. Transfected C656G GR containing additional mutations or deletions was tested on PEPCK gene expression in H4IIE hepatoma cells. We found that the AF2 domain is the only one of the three defined transactivation domains in GR that is required for PEPCK gene expression and that mutation of this domain disrupts the direct interaction of GR with steroid receptor coactivator 1 (SRC-1). These data help define the functional interaction between GR and SRC-1 and further define the role of the GR in glucocorticoid-mediated expression of the PEPCK gene. PMID- 12118044 TI - An evaluation of clinical governance in the public health departments of the West Midlands Region. AB - STUDY OBJECTIVES: (1) To evaluate the development of clinical governance within public health departments. (2) To assess two models for examining clinical governance in public health departments. DESIGN: Semi-structured interviews carried out during the annual visits of the regional director of public health to the health authority public health departments. SETTING: West Midland Region, England. PARTICIPANTS: Directors of public health plus other members of public health departments. MAIN RESULTS: These visits demonstrated that there is already a substantial amount of clinical governance activity taking place in the region's public health departments. There was also a need to reclassify many routinely occurring activities and include them under the clinical governance heading. CONCLUSIONS: The two models both proved useful for examining clinical governance in public health departments, however combining them into a matrix provided the best results. This matrix will still be useful after the reorganisation of the NHS and could be used to assess any public health department in the world. The West Midland public health departments find the visits valuable as they provide a source of external peer review of their activities. The public health departments have ownership of the process. PMID- 12118045 TI - Employment security and health. AB - OBJECTIVE: To study the relation of contractual and perceived employment security to employee health. DESIGN: Cross sectional survey. SETTING: Municipal sector employees in eight Finnish towns. PARTICIPANTS: 5981 employees with a permanent contract and 2786 employees with a non-permanent contract (2194 fixed term contract, 682 government subsidised contract). OUTCOME MEASURES: Poor self rated health, chronic disease, and psychological distress. RESULTS: Compared with permanent employees, fixed term men and women had better self rated health (men odds ratio 0.70; 95% confidence intervals 0.50 to 0.98, women 0.70 (0.60 to 0.82) and less chronic disease (men 0.69; 0.52 to 0.91; women 0.89; 0.79 to 1.02), but women had more psychological distress (1.26; 1.09 to 1.45). The only difference between subsidised employees and permanent employees was the high level of psychological distress in women (1.35; 1.09 to 1.68). Low perceived employment security was associated with poor health across all three indicators. The association of low perceived security with psychological distress was significantly stronger in permanent employees than among fixed term and subsidised employees, indicating that perceived security is more important for mental health among employees with a permanent contract. CONCLUSIONS: Contractual security and perceived security of employment are differently associated with health. It is therefore important to distinguish between these aspects of employment security in studies of labour market status and health. Such studies will also need to control for health selection, which is unlikely to operate in the same way among permanent and non-permanent employees. PMID- 12118046 TI - Unexplained illness among injecting drug users in Dublin: a case-control study. PMID- 12118047 TI - Time of day variation in rates of obstetric intervention to assist in vaginal delivery. PMID- 12118048 TI - Transitions to informal care in Great Britain during the 1990s. AB - OBJECTIVES: To estimate annual changes and trends in the population of informal carers and to investigate transitions to caregiving by age, gender, locus of care, and level of involvement. DESIGN: Longitudinal analysis of data from the British household panel survey, 1991 to 1998, an annual prospective survey of a nationally representative sample of more than 5000 private households in England, Scotland, and Wales. SUBJECTS: Over 9000 adults over 16 years interviewed personally in successive waves of the survey, including around 1300 informal carers each year. RESULTS: One third of co-resident carers and 40% of extra resident carers start caregiving each year and similar proportions cease to provide care. Five year period rates are at least 75% higher than the one year prevalence estimates. Almost everyone is involved in caregiving at one time or another and over half are likely to provide 20 hours or more care per week at some point in their lives. Recent trends indicate that more adults are becoming heavily involved in providing longer episodes of care. Although the onset of caregiving peaks in late middle and early older age, above average incidences span three decades or more of adult life. Age variations in the start of caring relationships are driven by the changing demands for care within and between generations over the life course. There is no firm evidence that carers increase their involvement in caring activities over the first three years of a caring episode. CONCLUSIONS: The population of carers is constantly changing as some people stop providing care and others take on a caring role or vary their level of involvement. Policy measures responsive to the diversity of caring roles, and geared around key transitions, are likely to be most effective in supporting carers through changing circumstances. Recognition and support for carers who are heavily involved in caring activities from the outset should be a priority. PMID- 12118049 TI - A glossary for multilevel analysis. AB - Multilevel analysis has recently emerged as a useful analytical technique in several fields, including public health and epidemiology. This glossary defines key concepts and terms used in multilevel analysis. PMID- 12118050 TI - Assessing socioeconomic status in adolescents: the validity of a home affluence scale. AB - STUDY OBJECTIVE: To examine the completion rate, internal reliability, and external validity of a home affluence scale based on adolescents' reports of material circumstances in the home as a measure of family socioeconomic status. DESIGN: Cross sectional survey. SETTING: Data were collected from a school based study in seven schools in the north of England Cheshire over a five month period from September 1999 to January 2000. PARTICIPANTS: 1824 students (1248 girls, 567 boys) aged 13-15 years who were attending normal classes in Years 9 and 10 in 7 schools on the days of data collection. MAIN RESULTS: Comparatively poor completion rates were found for questions on parental education and occupation while material deprivation items had much higher completion rates. There was evidence that students with poorer material circumstances were less able to report parental education and occupation whereas material based questions showed less bias. A home affluence scale composed of material items was found to have adequate internal reliability and good external validity. CONCLUSIONS: A home affluence scale based on material markers provides a useful alternative in assessing family affluence in adolescents. Additionally, it prevents exclusion of those less materially well off adolescents who fail to complete conventional socioeconomic status items. PMID- 12118051 TI - Impact numbers in health policy decisions. AB - OBJECTIVE: To outline the major methodological issues appropriate to the use of the population impact number (PIN) and the disease impact number (DIN) in health policy decision making. DESIGN: Review of literature and calculation of PIN and DIN statistics in different settings. SETTING: Previously proposed extensions to the number needed to treat (NNT): the DIN and the PIN, which give a population perspective to this measure. MAIN RESULTS: The PIN and DIN allow us to compare the population impact of different interventions either within the same disease or in different diseases or conditions. The primary studies used for relative risk estimates should have outcomes, time periods and comparison groups that are congruent and relevant to the local setting. These need to be combined with local data on disease rates and population size. Depending on the particular problem, the target may be disease incidence or prevalence and the effects of interest may be either the incremental impact or the total impact of each intervention. For practical application, it will be important to use sensitivity analyses to determine plausible intervals for the impact numbers. CONCLUSIONS: Attention to various methodological issues will permit the DIN and PIN to be used to assist health policy makers assign a population perspective to measures of risk. PMID- 12118052 TI - Impact numbers: measures of risk factor impact on the whole population from case control and cohort studies. AB - OBJECTIVE: To describe new measures of risk from case-control and cohort studies, which are simple to understand and relate to numbers of the population at risk. DESIGN: Theoretical development of new measures of risk. SETTING: Review of literature and previously described measures. MAIN RESULTS: The new measures are: (1) the population impact number (PIN), the number of those in the whole population among whom one case is attributable to the exposure or risk factor (this is equivalent to the reciprocal of the population attributable risk); (2) the case impact number (CIN) the number of people with the disease or outcome for whom one case will be attributable to the exposure or risk factor (this is equivalent to the reciprocal of the population attributable fraction); (3) the exposure impact number (EIN) the number of people with the exposure among whom one excess case is attributable to the exposure (this is equivalent to the reciprocal of the attributable risk); (4) the exposed cases impact number (ECIN) the number of exposed cases among whom one case is attributable to the exposure (this is equivalent to the reciprocal of the aetiological fraction). The impact number reflects the number of people in each population (the whole population, the cases, all those exposed, and the exposed cases) among whom one case is attributable to the particular risk factor. CONCLUSIONS: These new measures should help communicate the impact on a population, of estimates of risk derived from cohort or case-control studies. PMID- 12118053 TI - A health impact assessment model for environmental changes attributable to development projects. AB - STUDY OBJECTIVE: European Union legislation requires large industrial and civil development projects to undergo environmental impact assessment. The study objective was to identify environmental health risk estimates for these developments from the epidemiological literature and to develop, and apply these within, a mathematical health impact assessment model. DESIGN AND RESULTS: In the UK, good practice guidelines have set out environmental issues to be considered in development projects, but little attention is given to direct health effects. Broad quantifiable risks were identified for four-air, chemicals, noise, and road traffic-of 14 standard environmental effects. A mathematical model was constructed that is based on people moving between different health states over their lifetime. Age related hazard functions are applied to cause specific measures of mortality and morbidity. A hypothetical example for a development creating air and chemical pollutants is given. CONCLUSIONS: A mathematical model applying epidemiological risks to an exposed population can provide quantification of environmental health effects. The approach may in future find application during project development, and by public health regulatory authorities for environmental health impact assessment. PMID- 12118054 TI - Climate variability and Ross River virus transmission. AB - OBJECTIVES: (1) To examine the feasibility to link climate data with monthly incidence of Ross River virus (RRv). (2) To assess the impact of climate variability on the RRv transmission. DESIGN: An ecological time series analysis was performed on the data collected between 1985 to 1996 in Queensland, Australia. METHODS: Information on the notified RRv cases was obtained from the Queensland Department of Health. Climate and population data were supplied by the Australian Bureau of Meteorology and the Australian Bureau of Statistics, respectively. Spearman's rank correlation analyses were performed to examine the relation between climate variability and the monthly incidence of notified RRv infections. The autoregressive integrated moving average (ARIMA) model was used to perform a time series analysis. As maximum and minimum temperatures were highly correlated with each other (r(s)=0.75), two separate models were developed. RESULTS: For the eight major cities in Queensland, the climate-RRv correlation coefficients were in the range of 0.12 to 0.52 for maximum and minimum temperatures, -0.10 to 0.46 for rainfall, and 0.11 to 0.52 for relative humidity and high tide. For the whole State, rainfall (partial regression coefficient: 0.017 (95% confidence intervals 0.009 to 0.025) in Model I and 0.018 (0.010 to 0.026) in Model II), and high tidal level (0.030 (0.006 to 0.054) in Model I and 0.029 (0.005 to 0.053) in Model II) seemed to have played significant parts in the transmission of RRv in Queensland. Maximum temperature was also marginally significantly associated with the incidence of RRv infection. CONCLUSION: Rainfall, temperature, and tidal levels may be important environmental determinants in the transmission cycles of RRv disease. PMID- 12118055 TI - Changing sex ratio in Iran, 1976-2000. PMID- 12118056 TI - Early unemployment can contribute to adult health problems: results from a longitudinal study of school leavers. AB - STUDY OBJECTIVE: To investigate the long term effects of early unemployment (a total of more than half a year of unemployment between the ages of 16 and 21) on health behaviour and psychological and somatic symptoms. DESIGN: A 14 year follow up of a cohort of school leavers was conducted from 1981 to 1995. Information was collected by questionnaires. SETTING: An industrial town in northern Sweden. PARTICIPANTS: The original cohort was defined as all pupils in a middle sized municipality in the last year of compulsory school at age 16 (n=1083). The participants were followed up between the ages of 16 and 30. The analysis included 96% of the original sample, 547 men and 497 women MAIN RESULTS: After controlling for initial health behaviour and symptoms as well as for working class background and late unemployment, early unemployment among young men and women showed a significant explanatory effect on smoking, psychological symptoms and-among men only-somatic symptoms after a follow up of 14 years. No correlation was found between early unemployment and late excess alcohol consumption. CONCLUSIONS: Early unemployment can contribute to adult health problems. Thus, youth unemployment constitutes a significant public health problem, which to a certain extent remains in adult age. PMID- 12118057 TI - Depression and falls among community dwelling elderly people: a search for common risk factors. AB - STUDY OBJECTIVE: s: Depression and falls are two common conditions that impair the health of older people. Both are relatively underdiagnosed and undertreated problems in primary care. The study objective was to investigate whether there was a common set of risk factors that could predict an increased risk of both falls and depression. DESIGN: This was a cohort study drawn from a primary care clinic, with a one year follow up. Dependent measures included: reporting two or more falls in the past year and a score of 7 or over on the S-GDS (Short Geriatric Depression Scale). A parsimonious set of risk factors was selected that predicted both outcomes based on a series of discriminant function analyses. PARTICIPANTS AND SETTING: The setting was a primary care clinic serving a mixed socioeconomic population, in Beer Sheva, Israel. The sample included 283 General Sick Fund members, aged 60 and over, who completed both baseline assessments and one year follow up interviews. MAIN RESULTS: At the one year follow up, 12% of the sample reported frequent falls in the past year and 25.5% of the sample screened positive for depressive symptoms. A set of five risk factors that included: poor self rated health, poor cognitive status, impaired ADL, two or more clinic visits in the past month, and slow walking speed (g10 seconds over five metres) was successful at discriminating between fallers and non-fallers (86% discrimination) and between those with and without depressive symptoms (76%). For every risk factor added, there was a significant increase in the proportion of respondents who had depressive symptoms. A similar result was found for falls. CONCLUSIONS: These results show that there is a common set of risk factors that increase the risk of two common outcomes in geriatric medicine, falls and depression. For a general practitioner or a geriatric physician, it might be easier to detect these risk factors than to diagnose depression or high risk for falls. When these risk factors are detected in patients the physician can then be more active in direct probing about depression and falls. PMID- 12118059 TI - mRNA surveillance: the perfect persist. AB - In eukaryotes, an elaborate set of mechanisms has evolved to ensure that the multistep process of gene expression is accurately executed and adapted to cellular needs. The mRNA surveillance pathway works in this context by assessing the quality of mRNAs to ensure that they are suitable for translation. mRNA surveillance facilitates the detection and destruction of mRNAs that contain premature termination codons by a process called nonsense-mediated decay. Moreover, recent studies have shown that a distinct mRNA surveillance process, called nonstop decay, is responsible for depleting mRNAs that lack in-frame termination codons. mRNA surveillance thereby prevents the synthesis of truncated and otherwise aberrant proteins, which can have dominant-negative and other deleterious effects. PMID- 12118058 TI - Socioeconomic status of very small areas and stroke incidence in the Netherlands. AB - OBJECTIVE: To examine whether characteristics of very small living areas can be used to predict disease incidence and to use these characteristics to assess socioeconomic differences in stroke incidence in the Netherlands. DESIGN: Characteristics of postcode areas of stroke patients are compared with characteristics of postcode areas of all individual people in the study region, using Poisson regression analysis. SETTING: Six provinces of the Netherlands, covering about half of the country. PATIENTS: 760 patients who in 1991 or 1992 were consecutively admitted because of stroke to 23 Dutch hospitals. MAIN RESULTS: Stroke incidence is significantly higher among people living in postcode areas with below average socioeconomic status (relative risk=1.27; 95% confidence intervals 1.08 to 1.51) and among people living in postcode areas with predominantly older inhabitants (RR=3.17; 95% CI=2.29 to 4.39). It is also significantly increased in more urbanised areas compared with the countryside, the highest incidence being found in the large cities (RR=1.78; 95% CI=1.31 to 2.44). CONCLUSIONS: A clear socioeconomic gradient in stroke incidence in the Netherlands is observed, with people living in detailed postcode areas with below average socioeconomic status experiencing a significantly higher risk of stroke. The analysis also confirms that characteristics of detailed postcode areas can effectively be used to differentiate between areas with and areas without stroke patients. PMID- 12118060 TI - Beyond calcium: new signaling pathways for Tec family kinases. AB - The Tec kinases represent the second largest family of mammalian non-receptor tyrosine kinases and are distinguished by the presence of distinct proline-rich regions and pleckstrin homology domains that are required for proper regulation and activation. Best studied in lymphocyte and mast cells, these kinases are critical for the full activation of phospholipase-C gamma (PLC-gamma) and Ca(2+) mobilization downstream of antigen receptors. However, it has become increasingly clear that these kinases are activated downstream of many cell-surface receptors, including receptor tyrosine kinases, cytokine receptors, integrins and G-protein coupled receptors. Evidence suggests that the Tec kinases influence a wide range of signaling pathways controlling activation of MAP kinases, actin reorganization, transcriptional regulation, cell survival and cellular transformation. Their impact on cellular physiology suggests that the Tec kinases help regulate multiple cellular processes beyond Ca(2+) mobilization. PMID- 12118061 TI - Host cells: mobilizable lipid resources for the intracellular parasite Toxoplasma gondii. AB - Successful replication of the intracellular parasite Toxoplasma gondii within its parasitophorous vacuole necessitates a substantial increase in membrane mass. The possible diversion and metabolism of host cell lipids and lipid precursors by Toxoplasma was therefore investigated using radioisotopic and fluorophore conjugated compounds. Confocal microscopic analyses demonstrated that Toxoplasma is selective with regards to both the acquisition and compartmentalization of host cell lipids. Lipids were compartmentalized into parasite endomembranes and, in some cases, were apparently integrated into the surrounding vacuolar membrane. Additionally, some labels became concentrated in discrete lipid bodies that were biochemically and morphologically distinct from the parasite apical secretory organelles. Thin layer chromatography established that parasites readily scavenged long-chain fatty acids as well as cholesterol, and in certain cases modified the host-derived lipids. When provided with radiolabeled phospholipid precursors, including polar head groups, phosphatidic acid and small fatty acids, intracellular parasites preferentially accrued phosphatidylcholine (PtdCho) over other phospholipids. Moreover, Toxoplasma was found to be competent to synthesize PtdCho from radiolabeled precursors obtained from its environment. Together, these studies underscore the ability of Toxoplasma gondii to divert and use lipid resources from its host, a process that may contribute to the biogenesis of parasite membranes. PMID- 12118063 TI - Ephrin-B1 transduces signals to activate integrin-mediated migration, attachment and angiogenesis. AB - Ephrin-B/EphB family proteins are implicated in bidirectional signaling and were initially defined through the function of their ectodomain sequences in activating EphB receptor tyrosine kinases. Ephrin-B1-3 are transmembrane proteins sharing highly conserved C-terminal cytoplasmic sequences. Here we use a soluble EphB1 ectodomain fusion protein (EphB1/Fc) to demonstrate that ephrin-B1 transduces signals that regulate cell attachment and migration. EphB1/Fc induced endothelial ephrin-B1 tyrosine phosphorylation, migration and integrin-mediated (alpha(v)beta(3) and alpha(5)beta(1)) attachment and promoted neovascularization, in vivo, in a mouse corneal micropocket assay. Activation of ephrin-B1 by EphB1/Fc induced phosphorylation of p46 JNK but not ERK-1/2 or p38 MAPkinases. By contrast, mutant ephrin-B1s bearing either a cytoplasmic deletion (ephrin B1DeltaCy) or a deletion of four C-terminal amino acids (ephrin-B1DeltaPDZbd) fail to activate p46 JNK. Transient expression of intact ephin-B1 conferred EphB1/Fc migration responses on CHO cells, whereas the ephrin-B1DeltaCy and ephrin-B1DeltaPDZbd mutants were inactive. Thus ephrin-B1 transduces 'outside-in' signals through C-terminal protein interactions that affect integrin-mediated attachment and migration. PMID- 12118062 TI - Mitosis in primary cultures of Drosophila melanogaster larval neuroblasts. AB - Although Drosophila larval neuroblasts are routinely used to define mutations affecting mitosis, the dynamics of karyokinesis in this system remain to be described. Here we outline a simple method for the short-term culturing of neuroblasts, from Drosophila third instar larvae, that allows mitosis to be followed by high-resolution multi-mode light microscopy. At 24 degrees C, spindle formation takes 7+/-0.5 minutes. Analysis of neuroblasts containing various GFP tagged proteins (e.g. histone, fizzy, fizzy-related and alpha-tubulin) reveals that attaching kinetochores exhibit sudden, rapid pole-directed motions and that congressing and metaphase chromosomes do not undergo oscillations. By metaphase, the arms of longer chromosomes can be resolved as two chromatids, and they often extend towards a pole. Anaphase A and B occur concurrently, and during anaphase A chromatids move poleward at 3.2+/-0.1 microm/minute, whereas during anaphase B the spindle poles separate at 1.6+/-01 microm/minute. In larger neuroblasts, the spindle undergoes a sudden shift in position during midanaphase, after which the centrally located centrosome preferentially generates a robust aster and stops moving, even while the spindle continues to elongate. Together these two processes contribute to an asymmetric positioning of the spindle midzone, which, in turn, results in an asymmetric cytokinesis. Bipolar spindles form predominately (83%) in association with the separating centrosomes. However, in 17% of the cells, secondary spindles form around chromosomes without respect to centrosome position: in most cases these spindles coalesce with the primary spindle by anaphase, but in a few they remain separate and define additional ectopic poles. PMID- 12118065 TI - MEK/ERK pathway mediates cell-shape-dependent plasminogen activator inhibitor type 1 gene expression upon drug-induced disruption of the microfilament and microtubule networks. AB - Changes in cellular morphology induced as a consequence of direct perturbation of cytoskeletal structure with network-specific targeting agents (i.e. microfilament or microtubule-disrupting drugs) results in the stimulated expression of a specific subset of genes. Transcription of c-fos, collagenase, transforming growth factor-beta, actin, urokinase plasminogen activator and its type-1 inhibitor (PAI-1) appears to be particularly responsive to shape-activated signaling pathways. Cytochalasin D (CD) or colchicine treatment of contact inhibited and serum-deprived vascular smooth muscle (R22) cells was used, therefore, as a model system to evaluate morphology-associated controls on PAI-1 gene regulation in the absence of added growth factors. PAI-1 transcript levels in quiescent R22 cells increased rapidly and in a CD-concentration-dependent fashion, with kinetics of expression paralleling the morphological changes. Colchicine concentrations that effectively disrupted microtubule structure and reduced the cellular 'footprint' area (to approximately that of CD treatment) also stimulated PAI-1 synthesis. Shape-related increases in PAI-1 mRNA synthesis were ablated by prior exposure to actinomycin D. Unlike the mechanism of induction in growth-factor-stimulated cells, CD- and colchicine-induced PAI-1 expression required on-going protein synthesis (i.e. it was a secondary response). Although PAI-1 is a TGF-beta-regulated gene and TGF-beta expression is also shape dependent, an autocrine TGF-beta loop was not a factor in CD-initiated PAI-1 transcription. Since CD exposure resulted in actin microfilament disruption and subsequent morphological changes, with uncertain effects on interactions between signaling intermediates or 'scaffold' structures, a pharmacological approach was selected to probe the pathways involved. Signaling events leading to PAI-1 induction were compared with colchicine-treated cells. CD- as well as colchicine-stimulated PAI-1 expression was effectively and dose dependently attenuated by the MEK inhibitor PD98059 (in the 10 to 25 microM concentration range), consistent with the known MAP kinase dependency of PAI-1 synthesis in growth-factor-stimulated cells. Reduced PAI-1 mRNA levels upon exposure to genistein prior to CD addition correlated with inhibition of ERK1/2 activity, implicating a tyrosine kinase in shape-dependent MEK activation. Src-family kinases, moreover, appeared to be specific upstream elements in the CD- and colchicine-dependent pathways of PAI-1 transcription since both agents effectively activated pp60(c-src) kinase activity in quiescent R22 cells. The restrictive (src-family) kinase inhibitor PP1 completely inhibited induced, as well as basal, ERK activity in a coupled immunoprecipitation myelin-basic-protein phosphorylation assay and ablated shape-initiated PAI-1 mRNA expression. These data suggest that PP1-sensitive tyrosine kinases are upstream intermediates in cell-shape-associated signaling pathways resulting in ERK1/2 activation and subsequent PAI-1 transcription. In contrast to the rapid and transient kinetics of ERK activity typical of serum-stimulated cells, the ERK1/2 response to CD and colchicine is both delayed and relatively sustained. Collectively, these data support a model in which MEK is a focal point for the convergence of shape initiated signaling events leading to induced PAI-1 transcription. PMID- 12118064 TI - Suppression of Synaptotagmin II restrains phorbolester-induced downregulation of protein kinase Calpha by diverting the kinase from a degradative pathway to the recycling endocytic compartment. AB - Downregulation of protein kinase Calpha (PKCalpha) following long-term exposure to phorbol esters such as TPA is traffic dependent and involves delivery of the active, membrane-associated PKCalpha to endosomes. In this study, we show that synaptotagmin II (Syt II), a member of the Syt family of proteins, is required for TPA-induced degradation of PKCalpha. Thus, whereas the kinase half-life in TPA-treated cultured mast cells (the mast cell line rat basophilic leukemia RBL 2H3) is 2 hours, it is doubled in RBL-Syt II(-) cells, in which the cellular level of Syt II is reduced by >95% by transfection with Syt II antisense cDNA. We demonstrate that in TPA-treated RBL cells, PKCalpha travels from the cytosol to the plasma membrane, where it is delivered to early endosomes on its route to degradation. By contrast, in TPA-treated RBL-Syt II(-) cells, PKCalpha is diverted to recycling endosomes and remains distributed between the plasma membrane and the perinuclear recycling endocytic compartment. Notably, in both RBL and RBL-Syt II(-) cells, a fraction of PKCalpha is delivered and maintained in the secretory granules (SG). These results implicate Syt II as a critical factor for the delivery of internalized cargo for degradation. As shown here, one consequence of Syt II suppression is a delay in PKCalpha downregulation, resulting in its prolonged signaling. PMID- 12118066 TI - The PtdIns3P phosphatase myotubularin is a cytoplasmic protein that also localizes to Rac1-inducible plasma membrane ruffles. AB - Myotubularin, the phosphatase mutated in X-linked myotubular myopathy, was shown to dephosphorylate phosphatidylinositol 3-monophosphate (PtdIns3P) and was also reported to interact with nuclear transcriptional regulators from the trithorax family. We have characterized a panel of specific antibodies and investigated the subcellular localization of myotubularin. Myotubularin is not detected in the nucleus, and localizes mostly as a dense cytoplasmic network. Overexpression of myotubularin does not detectably affect vesicle trafficking in the mammalian cells investigated, in contrast to previous observations in yeast models. Both mutation of a key aspartate residue of myotubularin and dominant activation of Rac1 GTPase lead to the recruitment of myotubularin to specific plasma membrane domains. Localization to Rac1-induced ruffles is dependent on the presence of a domain highly conserved in the myotubularin family (that we named RID). We thus propose that myotubularin may dephosphorylate a subpool of PtdIns3P (or another related substrate) at the plasma membrane. PMID- 12118067 TI - Distance-dependent cellular palmitoylation of de-novo-designed sequences and their translocation to plasma membrane subdomains. AB - Using recursive PCR, we created an artificial protein sequence that consists of a consensus myristoylation motif (MGCTLS) followed by the triplet AGS repeated nine times and fused to the GFP reporter. This linker-GFP sequence was utilized as a base to produce multiple mutants that were used to transfect COS-7 cells. Constructs where a 'palmitoylable' cysteine residue was progressively moved apart from the myristoylation site to positions 3, 9, 15 and 21 of the protein sequence were made, and these mutants were used to investigate the effect of protein myristoylation on subsequent palmitoylation, subcellular localization, membrane association and caveolin-1 colocalization. In all cases, dual acylation of the GFP chimeras correlated with translocation to Triton X-100-insoluble cholesterol/sphingomyelin-enriched subdomains. Whereas a strong Golgi labeling was observed in all the myristoylated chimeras, association with the plasma membrane was only observed in the dually acylated constructs. Taking into account the conflicting data regarding the existence and specificity of cellular palmitoyl-transferases, our results provide evidence that de-novo-designed sequences can be efficiently S-acylated with palmitic acid in vivo, strongly supporting the hypothesis that non-enzymatic protein palmitoylation can occur within mammalian cells. Additionally, this palmitoylation results in the translocation of the recombinant construct to low-fluidity domains in a myristate palmitate distance-dependent manner. PMID- 12118068 TI - Selective impairment of p53-mediated cell death in fibroblasts from sporadic Alzheimer's disease patients. AB - In this study, we evaluated the response of different human skin fibroblast cultures obtained from eight probable Alzheimer's disease patients and eight non Alzheimer's disease subjects to an acute oxidative injury elicited by H(2)O(2). This treatment generates reactive oxygen species, which are responsible for DNA damage and apoptosis. To compare the sensitivity of fibroblasts from Alzheimer's disease or non-Alzheimer's disease patients to H(2)O(2) exposure, we evaluated different parameters, including cell viability, the extension of DNA damage and the ability of the cells to arrest proliferation and to activate an apoptotic program. We found that fibroblasts from Alzheimer's disease patients were more resistant that those from control subjects to H(2)O(2) treatment, although the extent of DNA damage induced by the oxidative injury was similar in both experimental groups. The protective mechanism of Alzheimer's disease fibroblasts was related to an impairment of H(2)O(2)-induced cell cycle arrest and characterized by an accelerated re-entry into the cell cycle and a diminished induction of apoptosis. Fibroblasts from Alzheimer's disease patients also have a profound impairment in the H(2)O(2)-activated, p53-dependent pathway, which results in a lack of activation of p53 or p53-target genes, including p21, GADD45 and bax. This study demonstrates a specific alteration of an intracellular pathway involved in sensing and repairing DNA damage in peripheral cells from Alzheimer's disease patients. PMID- 12118069 TI - Rho5p downregulates the yeast cell integrity pathway. AB - The Rho family of proteins and their effectors are key regulators involved in many eukaryotic cell functions. In Saccharomyces cerevisiae the family consists of six members, Rho1p to Rho5p and Cdc42p. With the exception of Rho5p, these enzymes have been assigned different biological functions, including the regulation of polar growth, morphogenesis, actin cytoskeleton, budding and secretion. Here we show that a rho5 deletion results in an increased activity of the protein kinase C (Pkc1p)-dependent signal transduction pathway. Accordingly, the deletion shows an increased resistance to drugs such as caffeine, Calcofluor white and Congo red, which indicates activation of the pathway. In contrast, overexpression of an activated RHO5Q91H mutant renders cells more sensitive to these drugs. We conclude that Rho5p acts as an off-switch for the MAP-kinase cascade, which differentiates between MAP-kinase-dependent and -independent functions of Pkc1p. Kinetics of actin depolarisation and repolarisation after heat treatment of rho5 deletions as well as strains overexpressing the activated RHO5Q91H allele provide further evidence for such a function. PMID- 12118070 TI - S100A13 and S100A6 exhibit distinct translocation pathways in endothelial cells. AB - S100 proteins have attracted great interest in recent years because of their cell and tissue-specific expression and association with various human pathologies. Most S100 proteins are small acidic proteins with calcium-binding domains - the EF hands. It is thought that this group of proteins carry out their cellular functions by interacting with specific target proteins, an interaction that is mainly dependent on exposure of hydrophobic patches, which result from calcium binding. S100A13, one of the most recently identified members of the S100 family, is expressed in various tissues. Interestingly, hydrophobic exposure was not observed upon calcium binding to S100A13 even though the dimeric form displays two high- and two low- affinity sites for calcium. Here, we followed the translocation of S100A13 in response to an increase in intracellular calcium levels, as protein translocation has been implicated in assembly of signaling complexes and signaling cascades, and several other S100 proteins are involved in such events. Translocation of S100A13 was observed in endothelial cells in response to angiotensin II, and the process was dependent on the classic Golgi-ER pathway. By contrast, S100A6 translocation was found to be distinct and dependent on actin-stress fibers. These experiments suggest that different S100 proteins utilize distinct translocation pathways, which might lead them to certain subcellular compartments in order to perform their physiological tasks in the same cellular environment. PMID- 12118071 TI - Focus-formation of replication protein A, activation of checkpoint system and DNA repair synthesis induced by DNA double-strand breaks in Xenopus egg extract. AB - The response to DNA damage was analyzed using a cell-free system consisting of Xenopus egg extract and demembranated sperm nuclei. In the absence of DNA damaging agents, detergent-resistant accumulation of replication protein A appeared in nuclei after a 30 minute incubation, and a considerable portion of the replication protein A signals disappeared during a further 30 minute incubation. Similar replication protein A accumulation was observed in the nuclei after a 30 minute incubation in the extract containing camptothecin, whereas a further 30 minute incubation generated discrete replication protein A foci. The addition of camptothecin also induced formation of gamma-H2AX foci, which have been previously shown to localize at sites of DSBs. Analysis of the time course of DNA replication and results obtained using geminin, an inhibitor of licensing for DNA replication, suggest that the discrete replication protein A foci formed in response to camptothecin-induced DNA damage occur in a DNA-replication dependent manner. When the nuclei were incubated in the extract containing EcoRI, discrete replication protein A foci were observed at 30 minutes as well as at 60 and 90 minutes after incubation, and the focus-formation of replication protein A was not sensitive to geminin. DNA replication was almost completely inhibited in the presence of EcoRI and the inhibition was sensitive to caffeine, an inhibitor of ataxia telangiectasia mutated protein (ATM) and ATM- and Rad3-related protein (ATR). However, the focus-formation of replication protein A in the presence of EcoRI was not influenced by caffeine treatment. EcoRI-induced incorporation of biotin-dUTP into chromatin was observed following geminin-mediated inhibition of DNA replication, suggesting that the incorporation was the result of DNA repair. The biotin-dUTP signal co-localized with replication protein A foci and was not significantly suppressed or stimulated by the addition of caffeine. PMID- 12118072 TI - Occludin TM4(-): an isoform of the tight junction protein present in primates lacking the fourth transmembrane domain. AB - The tight junction protein occludin possesses four transmembrane domains, two extracellular loops, and cytoplasmic N- and C-termini. Reverse transcription-PCR analysis of human tissues, embryos and cells using primers spanning the fourth transmembrane domain (TM4) and adjacent C-terminal region revealed two products. The larger and predominant product corresponded in sequence to canonical occludin (TM4(+)), while the smaller product exhibited a 162 bp deletion encoding the entire TM4 and immediate C-terminal flanking region (TM4(-)). Examination of the genomic occludin sequence identified that the 162 bp sequence deleted in TM4(-) coincided precisely with occludin exon 4, strongly suggesting that TM4(-) is an alternative splice isoform generated by skipping of exon 4. Indeed, the reading frame of downstream exons is not affected by exclusion of exon 4. The presence of both TM4(+) and TM4(-) occludin isoforms was also identified in monkey epithelial cells but TM4(-) was undetected in murine and canine tissue and cells, indicating a late evolutionary origin for this alternative splicing event. Conceptual translation of TM4(-) isoform predicts extracellular localisation of the C terminus. Immunocytochemical processing of living human Caco-2 cells using a C terminal occludin antibody revealed weak, discontinuous staining restricted to the periphery of subconfluent islands of cells, or islands generated by wounding confluent layers. In occludin immunoblots, a weak band at approximately 58 kDa, smaller than the predominant band at 65 kDa and corresponding to the predicted mass of TM4(-) isoform, is evident and upregulated in subconfluent cells. These data suggest that the TM4(-) isoform may be translated at low levels in specific conditions and may contribute to regulation of occludin function. PMID- 12118073 TI - Unbound E2F modulates TGF-beta1-induced apoptosis in HuH-7 cells. AB - E2F is an important target of the retinoblastoma protein (pRb) and plays a critical role in G(1)/S progression through the cell cycle. TGF-beta1 arrests HuH 7 cells in G(1) by suppressing phosphorylation of pRb and induces apoptosis by inhibiting its expression. In this study, we examined the downstream effects of TGF-beta1-induced apoptosis and the potential roles for pRb and E2F. The results indicated that greater than 90% of the TGF-beta1-induced preapoptotic cells were arrested in G(1) phase of the cell cycle. This was associated with a significant increase in both E2F-DNA-binding activity and transcription of E2F-responsive reporter constructs. In contrast, no significant changes were observed in E2F mRNA and protein levels, and the overexpression of pRb partially inhibited E2F activation. Gel-shift assays identified more than four E2F complexes from preapoptotic and synchronized G(1) HuH-7 cells, each exhibiting different patterns of E2F-associated proteins. The increased E2F activity did not affect the association patterns with pRb, p107 and p130, but altered the formation of an E2F-DP-1 complex. In contrast, E2F-DP-2 exhibited little change in the preapoptotic cells. Moreover, TGF-beta1 induced apoptosis at G(1) and inhibited entry into S phase irrespective of the increased E2F activity. The release of preapoptotic cells from TGF-beta1 resulted in rapid S phase entry and subsequent apoptosis in 33% of cells over a 72 hour period. In conclusion, the results demonstrate that TGF-beta1-induced apoptosis in HuH-7 cells is associated with a marked increase in activity of transcription factor E2F that is partially inhibited by overexpression of pRb. Preapoptotic changes are, in part, reversible upon removal of TGF-beta1 and the majority of cells re-enter the normal cell cycle. Finally, TGF-beta1-induced apoptosis with the associated increase in E2F activity can occur in both the G(1) and S phases of the cell cycle. PMID- 12118074 TI - p38 mitogen-activated protein kinase is required for TGFbeta-mediated fibroblastic transdifferentiation and cell migration. AB - Transforming growth factor beta (TGFbeta) contributes to tumor progression by inducing an epithelial to mesenchymal transdifferentiation (EMT) and cell migration. We found that TGFbeta-induced EMT was blocked by inhibiting activation of p38 mitogen-activated protein kinase (MAPK) with H-7, a protein kinase C inhibitor, and with SB202190, a direct inhibitor of p38MAPK. Inhibition of the p38MAPK pathway affected TGFbeta-mediated phosphorylation of ATF2, but did not inhibit phosphorylation of Smad2. SB202190 impaired TGFbeta-mediated changes in cell shape and reorganization of the actin cytoskeleton. Forced expression of dominant-negative (DN) MAPK kinase 3 (MKK3) inhibited TGFbeta-mediated activation of p38MAPK and EMT. Expression of DN-p38alpha impaired TGFbeta-induced EMT. Inhibition of p38MAPK blocked TGFbeta-induced migration of non-tumor and tumor mammary epithelial cells. TGFbeta induced activation of the p38MAPK pathway within 15 minutes. Expression of TGFbeta type II (TbetaRII) and type I (TbetaRI/Alk5) kinase-inactive receptors blocked EMT and activation of p38MAPK, whereas expression of constitutively active Alk5-T204D resulted in EMT and phosphorylation of MKK3/6 and p38MAPK. Finally, dominant-negative Rac1N17 blocked TGFbeta-induced activation of the p38MAPK pathway and EMT, suggesting that Rac1 mediates activation of the p38MAPK pathway. These studies suggest that the p38MAPK pathway is required for TGFbeta-mediated EMT and cell migration. PMID- 12118075 TI - NUANCE, a giant protein connecting the nucleus and actin cytoskeleton. AB - NUANCE (NUcleus and ActiN Connecting Element) was identified as a novel protein with an alpha-actinin-like actin-binding domain. A human 21.8 kb cDNA of NUANCE spreads over 373 kb on chromosome 14q22.1-q22.3. The cDNA sequence predicts a 796 kDa protein with an N-terminal actin-binding domain, a central coiled-coil rod domain and a predicted C-terminal transmembrane domain. High levels of NUANCE mRNA were detected in the kidney, liver, stomach, placenta, spleen, lymphatic nodes and peripheral blood lymphocytes. At the subcellular level NUANCE is present predominantly at the outer nuclear membrane and in the nucleoplasm. Domain analysis shows that the actin-binding domain binds to Factin in vitro and colocalizes with the actin cytoskeleton in vivo as a GFP-fusion protein. The C terminal transmembrane domain is responsible for the targeting the nuclear envelope. Thus, NUANCE is the first alpha-actinin-related protein that has the potential to link the microfilament system with the nucleus. PMID- 12118076 TI - Protein interactions: two methods for assessment of the reliability of high throughput observations. AB - High throughput methods for detecting protein interactions require assessment of their accuracy. We present two forms of computational assessment. The first method is the expression profile reliability (EPR) index. The EPR index estimates the biologically relevant fraction of protein interactions detected in a high throughput screen. It does so by comparing the RNA expression profiles for the proteins whose interactions are found in the screen with expression profiles for known interacting and non-interacting pairs of proteins. The second form of assessment is the paralogous verification method (PVM). This method judges an interaction likely if the putatively interacting pair has paralogs that also interact. In contrast to the EPR index, which evaluates datasets of interactions, PVM scores individual interactions. On a test set, PVM identifies correctly 40% of true interactions with a false positive rate of approximately 1%. EPR and PVM were applied to the Database of Interacting Proteins (DIP), a large and diverse collection of protein-protein interactions that contains over 8000 Saccharomyces cerevisiae pairwise protein interactions. Using these two methods, we estimate that approximately 50% of them are reliable, and with the aid of PVM we identify confidently 3003 of them. Web servers for both the PVM and EPR methods are available on the DIP website (dip.doe-mbi.ucla.edu/Services.cgi). PMID- 12118077 TI - Mass measurements of C-terminally truncated alpha-crystallins from two dimensional gels identify Lp82 as a major endopeptidase in rat lens. AB - Molecular chaperone activity of lens alpha-crystallins is reduced by loss of the C terminus. The purpose of this experiment was to 1) determine the cleavage sites produced in vitro by ubiquitous m-calpain and lens-specific Lp82 on alpha crystallins, 2) identify alpha-crystallin cleavage sites produced in vivo during maturation and cataract formation in rat lens, and 3) estimate the relative activities of Lp82 and m-calpain by appearance of protease-specific cleavage products in vivo. Total soluble protein from young rat lens was incubated with recombinant m-calpain or Lp82 and 2 mM Ca2+. Resulting fragmented alpha crystallins were separated by two-dimensional gel electrophoresis. Eluted alpha crystallin spots were analyzed by mass spectrometry. Cleavage sites on insoluble alpha-crystallins were determined similarly in mature rat lens nucleus and in cataractous rat lens nucleus induced by selenite. In vitro proteolysis of alphaA crystallin by Lp82 and m-calpain produced unique cleavage sites by removing 5 and 11 residues, respectively, from the C terminus. In vivo, the protease-specific truncations removing 5 and 11 residues from alphaA were both found in maturing lens, whereas only the truncation removing 5 residues was found in cataractous lens. Other truncation sites, common to both calpain isoforms, resulted from the removal of 8, 10, 16, 17, and 22 residues from the C terminus of alphaA. Using uniquely truncated alphaA-crystallins as in vivo markers, Lp82 and m-calpain were both found to be active during normal maturation of rat lens, whereas Lp82 seemed especially active during selenite cataract formation. These C-terminal truncations decrease chaperone activity of alpha-crystallins, possibly leading to the observed increases in insoluble proteins during aging and cataract. The methodology that allowed accurate mass measurements of proteins eluted from 2D gels should be useful to examine rapidly other post-translational modifications. PMID- 12118078 TI - Alterations in the mouse and human proteome caused by Huntington's disease. AB - Huntington's disease is an autosomal dominantly inherited disease that usually starts in midlife and inevitably leads to death. In our effort to identify proteins involved in processes upstream or downstream of the disease-causing huntingtin, we studied the proteome of a well established mouse model by large gel two-dimensional electrophoresis. We could demonstrate for the first time at the protein level that alpha1-antitrypsin and alphaB-crystalline both decrease in expression over the course of disease. Importantly, the alpha1-antitrypsin decrease in the brain precedes that in liver and testes in mice. Reduced expression of the serine protease inhibitors alpha1-antitrypsin and contraspin was found in liver, heart, and testes close to terminal disease. Decreased expression of the chaperone alphaB-crystallin was found exclusively in the brain. In three brain regions obtained post-mortem from Huntington's disease patients, alpha1-antitrypsin expression was also altered. Reduced expression of the major urinary proteins not found in the brain was seen in the liver of affected mice, demonstrating that the disease exerts its influence outside the brain of transgenic mice at the protein level. Maintaining alpha1-antitrypsin and alphaB crystallin availability during the course of Huntington's disease might prevent neuronal cell death and therefore could be useful in delaying the disease progression. PMID- 12118079 TI - Stable isotope labeling by amino acids in cell culture, SILAC, as a simple and accurate approach to expression proteomics. AB - Quantitative proteomics has traditionally been performed by two-dimensional gel electrophoresis, but recently, mass spectrometric methods based on stable isotope quantitation have shown great promise for the simultaneous and automated identification and quantitation of complex protein mixtures. Here we describe a method, termed SILAC, for stable isotope labeling by amino acids in cell culture, for the in vivo incorporation of specific amino acids into all mammalian proteins. Mammalian cell lines are grown in media lacking a standard essential amino acid but supplemented with a non-radioactive, isotopically labeled form of that amino acid, in this case deuterated leucine (Leu-d3). We find that growth of cells maintained in these media is no different from growth in normal media as evidenced by cell morphology, doubling time, and ability to differentiate. Complete incorporation of Leu-d3 occurred after five doublings in the cell lines and proteins studied. Protein populations from experimental and control samples are mixed directly after harvesting, and mass spectrometric identification is straightforward as every leucine-containing peptide incorporates either all normal leucine or all Leu-d3. We have applied this technique to the relative quantitation of changes in protein expression during the process of muscle cell differentiation. Proteins that were found to be up-regulated during this process include glyceraldehyde-3-phosphate dehydrogenase, fibronectin, and pyruvate kinase M2. SILAC is a simple, inexpensive, and accurate procedure that can be used as a quantitative proteomic approach in any cell culture system. PMID- 12118080 TI - Plasma from cancer patients featuring a characteristic protein composition mediates protection against apoptosis. AB - By comparative proteome analysis we searched for characteristic alterations of human plasma accompanying neoplastic disease. We identified protein alterations in plasma of prostate-, lung-, and breast-cancer patients in comparison to controls, comprising elevated levels of fibrinogen gamma-chain dimer, degradation products of antiplasmin and laminin gamma-chain, and elevated levels of acute phase proteins. The latter proteins and laminin fragments have been described as anti-apoptotic factors. We raised the question whether these alterations may have any relevance for the regulation of apoptosis. In contrast to plasma derived from healthy donors, samples from prostate-, lung-, and breast-cancer patients selectively inhibited Fas- and staurosporine-induced apoptosis in Jurkat cells but remained ineffective upon UV light-induced apoptosis. These data suggested that inhibition occurred by extracellular interference with apoptosis induction. Supporting this hypothesis, we found that formation of the CD95 death-inducing signal complex was strongly inhibited in the presence of plasma from cancer patients. PMID- 12118081 TI - A multidomain TIGR/olfactomedin protein family with conserved structural similarity in the N-terminal region and conserved motifs in the C-terminal region. AB - Based on the similarity between the TIGR (trabecular-meshwork inducible glucocorticoid response) (also known as myocilin) and olfactomedin protein families identified throughout the length of the TIGR protein, we have identified more distantly related proteins to determine the elements essential to the function/structure of the TIGR and olfactomedin proteins. Using a sequence walk method and the Shotgun program, we have identified a family including 31 olfactomedin domain-containing sequences. Multiple sequence alignments and secondary structure analyses were used to identify conserved sequence elements. Pairwise identity in the olfactomedin domain ranges from 8 to 64%, with an average pairwise identity of 24%. The N-terminal regions of the proteins fall into two subgroups, one including the TIGR and olfactomedin families and another group of apparently unrelated domains. The TIGR and olfactomedin sequences display conserved motifs including a residual leucine zipper region and maintain a similar secondary structure throughout the N-terminal region. The correlation between conserved elements and disease-associated mutations and apparent polymorphisms in human TIGR was also examined to evaluate the apparent importance of conserved residues to the function/structure of TIGR. Several residues have been identified as essential to the function and/or structure of the human TIGR protein based on their degree of conservation across the family and their implication in the pathogenesis of primary open-angle glaucoma. Additionally, we have identified a group of chitinase sequences containing several of the highly conserved motifs present in the C-terminal region of the olfactomedin domain containing sequences. PMID- 12118083 TI - Long-chain L-3-hydroxyacyl-coenzyme a dehydrogenase deficiency: a molecular and biochemical review. AB - Since the first report of long-chain L-3-hydroxyacyl-coenzyme A dehydrogenase deficiency a little more than a decade ago, its phenotypic and genotypic heterogeneity in individuals homozygous for the enzyme defect has become more and more evident. Even more interesting is its association with pregnancy-specific disorders, including preeclampsia, HELLP syndrome (hemolysis, elevated liver enzymes, low platelets), hyperemesis gravidarum, acute fatty liver of pregnancy, and maternal floor infarct of the placenta. In this review we discuss the biochemical and molecular basis, clinical features, diagnosis, and management of long-chain L-3-hydroxyacyl-coenzyme A dehydrogenase deficiency. PMID- 12118084 TI - Nitric oxide-releasing aspirin decreases vascular injury by reducing inflammation and promoting apoptosis. AB - Endothelial dysfunction, defined as a deficit in the bioavailability of nitric oxide (NO), occurs as sequelae of many vascular diseases; however, the utility of supplementing NO to obviate the extent of disease is understudied. Here, we examined if prolonged treatment with an NO-releasing form of aspirin (NO-ASA) can influence neointimal remodeling of femoral arteries of hypercholesterolemic ApoE (-/-) mice. Treatment of ApoE (-/-) mice with NO-ASA, but not aspirin (ASA), improved neointimal remodeling post-injury. NO-ASA treatment increased lumen diameters and reduced intimal-to-medial ratios of injured femoral arteries compared with ASA- or vehicle-treated mice. The reduction in lumen diameter in NO ASA-treated mice was associated with a marked reduction in CD45-positive inflammatory cells and an increased number of TUNEL-positive cells. Thus, NO-ASA, by virtue of releasing NO, can reduce vascular inflammation and promote apoptosis during vascular remodeling associated with neointimal thickening. PMID- 12118085 TI - The diabetes-prone NZO/HlLt strain. I. Immunophenotypic comparison to the related NZB/BlNJ and NZW/LacJ strains. AB - New Zealand Obese (NZO)/HlLt male mice exhibit a polygenic obesity and approximately 50% develop type 2 diabetes. This strain is known to produce a variety of autoantibodies, including autoantibodies to the insulin receptor. Because of their relatedness to the autoimmune-predisposed New Zealand Black (NZB) and New Zealand White (NZW) inbred strains, we compared NZO to its two related strains for shared hematologic and immunologic characteristics. Comparison of the three strains by serotyping and genotyping methods indicated that NZO shared with NZW the rare (recombinant) H2(z) haplotype at the major histocompatibility complex. Similar to the NZB and NZW strains, spleens from NZO mice contained increased numbers of CD19(+)CD43(+) IgM(+) B-1 B cells, a phenotype associated with natural autoantibody production. NZO mice developed a progressive microcytic anemia that was distinguished from NZB hemolytic anemia by absence of demonstrable antierythrocyte antibodies in the former. Outcross of NZO females with NZB males accelerated development of obesity and diabetes in F1 males. NZO males made B-lymphocyte-deficient by a disrupted immunoglobulin heavy chain gene did not become diabetic. These results suggest that NZO mice should be useful to investigators interested in studying the genetic contributions to autoimmunity made by the related NZW and NZB strains. Further, these results, combined with the pancreatic histopathology contained in the companion manuscript, suggest that B lymphocytes may be important contributors to diabetes pathogenesis in the NZO mouse. PMID- 12118086 TI - The diabetes-prone NZO/Hl strain. II. Pancreatic immunopathology. AB - We report the first combined light and electron microscopic analysis of the pancreas during the development of type 2 diabetes in the New Zealand Obese (NZO) mouse. As in most other polygenic rodent models of type 2 diabetes, hyperglycemia associated with beta cell destruction is male sex-limited. Increasing degrees of hyperinsulinemia and transition to diabetes were clearly reflected by the islet volume fraction, by the beta cell granulation state, and by ultrastructural changes, primarily of the endoplasmic reticulum. One of the unusual histopathologic features of NZO mice of both sexes was the presence of B lymphocyte enriched leukocytic aggregates in the pancreas. Immunocytochemical analysis of the pancreas of 52-week-old diabetic males indicated enrichment for CD19(+) B lymphocytes. Staining of adjacent sections for CD3 and CD5 indicated CD5 coexpression on some of the CD19(+) cells, suggesting the presence of the B1 B subset associated with generation of natural autoantibodies in other autoimmune prone New Zealand mouse strains. In addition, plasma cells in peri-insular leukocytic infiltrates were identified by electron microscopy. Hence, although autoimmunity has previously proven to be a secondary manifestation of beta cell destruction in most rodent models of type 2 diabetes, the present observations suggest that B lymphocyte function, in association with male gender, may contribute to the development of insulin resistance and chronic hyperglycemia in the NZO model. PMID- 12118087 TI - PDGF enhancement of IL-1 receptor levels in smooth muscle cells involves induction of an attachment-regulated, heparan sulfate binding site (IL-1RIII). AB - This study shows that increase in IL-1 receptor levels by platelet derived growth factor (PDGF) involves an enhancement of a matrix-dependent, low-affinity receptor that constitutes a heparan sulfate. Fibronectin attachment caused pronounced alterations in IL-1 receptor function in smooth muscle cells, involving a pronounced increase in cell surface binding from an average of 2,000 up to approximately 8,000 receptors/cell and an increase in affinity (K(a)) of the type I receptor from 1.8 +/- 0.9 x 10(9) to 3.7 +/- 0.5 x 10(9) M(-1). PDGF stimulation similarly enhanced the level of cell surface binding by between 30% and 100%, with, in general, less effect on cells plated on fibronectin. Further, PDGF had a pronounced effect on the type I receptor affinity in the absence of matrix attachment, increasing the K(a) from 1.77 +/- 0.93 x 10(9) to 5.1 +/- 2.1 x 10(9) M(-1). Scatchard analyses revealed that PDGF, similarly to fibronectin attachment, caused enhancement of a second low-affinity binding site. Antibody blocking showed that approximately 50% of the attachment-induced increase was independent of type I receptor binding. Further, a similar fraction of the cell surface interaction was blocked by soluble heparan sulfate and dependent on cell binding to the heparan binding site. Cross-linking demonstrated that, in addition to the type I receptor, IL-1 bound to a second high molecular weight complex of 300 kd, induced by fibronectin attachment as well as by PDGF in the absence of matrix. Biochemical analyses demonstrated that this second site constitutes a heparan sulfate, which directly interacted with the type I receptor after recruitment to the complex, and which bound up to 50% and 25% of the ligand after fibronectin attachment and PDGF stimulation, respectively. The data show that PDGF induces an attachment-regulated low-affinity IL-1 binding site in smooth muscle cells, constituting a heparan sulfate. Correlation of the recruitment of this component to the IL-1 receptor complex with structural regulation of receptor function and enhancement of IL-1-mediated responses suggests that this is a significant mechanism in PDGF augmentation of local inflammatory responses during vessel wall pathogenesis. PMID- 12118088 TI - Photocoagulation-induced retinal gliosis is inhibited by systemically expressed soluble TGF-beta receptor type II via adenovirus mediated gene transfer. AB - Retinal gliosis is one of the major causes of visual dysfunction due to the loss of the retinal regular structure and function in various diseases, including diabetic retinopathy, retinal detachment, and glaucoma. Transforming growth factor-beta (TGF-beta) is assumed to play an important role in this disease process. In the present study, we determined whether the systemically expressed extracellular domain of the TGF-beta type II receptor by adenovirus-mediated gene delivery could inhibit experimental retinal gliosis both in vitro and in vivo. Cultured bovine retinal glial cells, Muller cells, were stimulated by recombinant TGF-beta and the expression of the glial marker, glial fibrillary acidic protein (GFAP), was evaluated by immunohistochemistry, semiquantitative RT-PCR, and Western blotting. In cultured Muller cells, TGF-beta stimulated the GFAP expression in a dose-dependent fashion, and the conditioned medium from 293 cells transfected with adenovirus encoding for a soluble form TGF-beta type II receptor (AdT beta-ExR) inhibited the expression of GFAP stimulated by exogenous TGF-beta (p < 0.05). In this process, Smad4 protein, which plays a key role in intracellular signaling after cell surface receptors, actually translocated from cytosol to nucleus with TGF-beta stimulation. The conditioned medium from AdT beta-ExR also inhibited the cytosol-nuclear translocation of Smad4. For in vivo studies, AdT beta-ExR was injected into the femoral muscles of Brown Norway rats and retinal photocoagulation was subsequently carried out. Immunohistochemical studies revealed that GFAP was strongly expressed around the photocoagulation spots after 12 days and these phenomena were inhibited by AdT beta-ExR. Western blotting of total retinal extract demonstrated the same results as those observed after immunohistochemistry. Our results suggest that TGF-beta plays a pivotal role in the pathologic processes in retinal gliosis, and that the systemically expressed soluble TGF receptor by gene delivery may thus have a potential therapeutic value by inhibiting excessive retinal gliosis in various ocular diseases. PMID- 12118089 TI - Suppressed angiogenesis in kininogen-deficiencies. AB - We investigated whether the kinin-generating system enhanced angiogenesis in chronic and proliferative granuloma and in tumor-surrounding stroma. In rat sponge implants, angiogenesis was gradually developed in normal Brown Norway Kitasato rats (BN-Ki). The development of angiogenesis was significantly suppressed in kininogen-deficient Brown Norway Katholiek rats (BN-Ka). The angiogenesis enhanced by basic fibroblast growth factor was also significantly less marked in BN-Ka than in BN-Ki. Naturally occurring angiogenesis was significantly suppressed by B(1) or B(2) antagonist. mRNA of vascular endothelial growth factor was more highly expressed in the granulation tissues in BN-Ki than in BN-Ka. Daily topical injections of aprotinin, but not of soy bean trypsin inhibitor, suppressed angiogenesis. Daily topical injections of low-molecular weight kininogen enhanced angiogenesis in BN-Ka. Topical injections of serum from BN-Ki, but not from BN-Ka, also facilitated angiogenesis in BN-Ka. FR190997, a nonpeptide mimic of bradykinin, promoted angiogenesis markedly, with concomitant increases in vascular endothelial growth factor mRNA. Angiogenesis in the granulation tissues around the implanted Millipore chambers containing Walker-256 cells was markedly more suppressed in BN-Ka than in BN-Ki. Our results suggest that endogenous kinin generated from the tissue kallikrein-kinin system enhances angiogenesis in chronic and proliferative granuloma and in the stroma surrounding a tumor. Thus, the agents for the kinin-generating system and/or kinin receptor signaling may become useful tools for controlling angiogenesis. PMID- 12118090 TI - Role of the aspartyl-asparaginyl-beta-hydroxylase gene in neuroblastoma cell motility. AB - Aspartyl (asparaginyl) beta-hydroxylase (AAH) is overexpressed in various malignant neoplasms, and high levels of immunoreactivity mainly occur in infiltrating or metastasized tumors. In addition, AAH is abundantly expressed in normally invasive placental trophoblastic cells. These observations led to the hypothesis that AAH may have a role in motility and aggressive behavior of tumor cells. The present study demonstrates that AAH is overexpressed in primary human malignant neuroectodermal tumors, including medulloblastomas and neuroblastomas, and that AAH expression is at a low level or undetectable in the normal mature brain. In the Sy5y neuroblastoma cell line, endogenous expression of the approximately 86-kd AAH protein was demonstrated by Western blot analysis, and immunoreactivity predominantly localized to the cell surface by immunocytochemical staining and FACS analysis. Sy5y cells that were stably transfected with the human AAH cDNA had increased levels of proliferating cell nuclear antigen and Bcl-2, and reduced levels of p21/Waf1 and p16. In addition, increased AAH expression enhanced Sy5y cell motility, whereas antisense oligodeoxynucleotide inhibition of AAH significantly reduced Sy5y cell motility and increased the levels of p21/Waf1 and p16. The findings suggest that AAH overexpression contributes to the malignant phenotype of neuroectodermal tumor cells by increasing motility and enhancing proliferation, survival, and cell cycle progression. Because AAH expression is at a low level or undetectable in normal brain, the AAH gene may be a target for treating primitive neuroectodermal tumors. PMID- 12118091 TI - High expression of methionine aminopeptidase type 2 in germinal center B cells and their neoplastic counterparts. AB - Methionine aminopeptidase type 2 (MetAP2) is a bifunctional protein that plays critical roles in the regulation of protein synthesis and post-translational processing by (a) protecting the alpha subunit of eukaryotic initiation factor 2 from inhibitory phosphorylation by eukaryotic initiation factor 2 kinases and (b) removing the amino-terminal methionine residue from nascent protein. MetAP2 is also known as the molecular target of the angiogenesis inhibitor TNP-470. In addition, it has been recently suggested that MetAP2 has an antiapoptotic function in mesothelioma. To know the pattern of expression of MetAP2 in normal and neoplastic tissues, we raised two specific rabbit polyclonal Abs and examined the pattern of MetAP2 expression in various normal and pathologic specimens. Unexpectedly, we found a very high and selective expression of MetAP2 in germinal center B cells. In the germinal center, dark zone B cells tended to express more MetAP2 than light zone B cells. When 200 malignant lymphomas of various subtypes were studied, a high level of MetAP2 expression, equivalent to that observed in germinal center B cells, was noted exclusively on B-cell lymphoma subtypes that are currently regarded as the neoplastic counterparts of germinal center B cells. The expression of MetAP2 in diffuse large B-cell lymphomas correlated well with that of BCL6 (p < 0.05) but not with that of either CD10 or BCL2. These data suggest that MetAP2 has specific function(s) in germinal center B cells and that the function is shared by neoplastic counterparts of germinal center B cells. PMID- 12118092 TI - Hypoxia and an angiogenic response in the partially obstructed rat bladder. AB - Previous molecular and blood flow studies performed on animal models of partial bladder outlet obstruction (PBOO) caused us to propose that bladder hypoxia/ischemia was a significant effector of the cellular and functional changes that occur in the bladder as a result of this condition. To confirm the occurrence of hypoxia in the partially obstructed bladder, we obtained rat bladders at increasing intervals following PBOO and measured biomarkers of hypoxia (intracellular formation of hypoxyprobe-1 adducts and expression of hypoxia inducible factor-1 alpha [HIF-1 alpha] protein) and whether such hypoxia might elicit an angiogenic response in the tissue. Rats receiving PBOO or controls were treated with hypoxyprobe-1 at increasing intervals subsequent to surgery and their bladders were sectioned and immunostained using an antibody that detects hypoxyprobe-1 adducts. Control rat bladders were unstained, whereas intense, but regionally restricted, hypoxyprobe-1 immunostaining was detected in all obstructed bladders in a unique pattern that changed over time. Proteins were extracted from bladders removed from similarly treated rats and were analyzed for the expression of the HIF-1 alpha protein as well as for expression of angiogenic regulatory factors (vascular endothelial growth factor, angiopoietin-1, and endostatin) using Western blotting techniques. HIF-1 alpha protein was not expressed in control bladders, however, the protein was highly up-regulated over the 2-week period after PBOO. Likewise, the expression of vascular endothelial growth factor (a downstream target of HIF-1 alpha action) and angiopoietin-1 was also up-regulated in obstructed bladders confirming an angiogenic response to this hypoxia. Enigmatically, however, expression of the antiangiogenic molecule endostatin was also up-regulated by chronic PBOO. These results further support the concept that hypoxia is involved in the cellular remodeling as well as in the progressive functional impairment exhibited by the urinary bladder after PBOO. PMID- 12118093 TI - Functional significance of erythropoietin receptor expression in breast cancer. AB - Erythropoietin (EPO) is the principal hematopoietic cytokine that regulates mammalian erythropoiesis by binding to its transmembrane receptor EpoR. Recent experimental evidence suggests that the biologic effects of EPO are not limited to the regulation of erythropoiesis. In studies focusing on nonhematopoietic effects of EpoR signaling, we found high levels of EpoR protein expression in human breast cancer cells. The purpose of the present study was to evaluate clinical breast cancer specimens for EPO and EpoR expression, characterize the relationship between EPO expression and tumor hypoxia in biopsies prelabeled with hypoxia marker pimonidazole, analyze breast cancer cell lines for EpoR expression, and study the functional significance of EpoR expression in breast cancer cells in vivo. Immunohistochemical analysis for EPO, EpoR expression, and pimonidazole adducts was performed on 26 tumor biopsies with contiguous sections from 10 patients with breast cancer. High levels of EpoR expression were found in cancer cells in 90% of tumors. EPO expression was found in 60% of tumors and EPO and EpoR colocalization in tumor cells was present in many cases. The expression pattern of EPO with respect to tumor hypoxia was variable, without consistent colocalization of EPO and hypoxia in tumor cells. Human and rat breast cancer tissue culture cells express EpoR mRNA and protein. To study the in vivo function of EpoR expression in breast cancer cells, we used rat syngeneic R3230Ac mammary adenocarcinoma cells in a tumor Z-chamber model (dual porous plexiglass chambers containing fibrin gel, cancer cells, and a putative anti-tumor compound implanted into the subcutaneous tissue of rats). Local, one-time administration of a neutralizing anti-EPO antibody, soluble EPO receptor, or an inhibitor of Jak2, a cytoplasmic tyrosine kinase essential for EPO-mediated mitogenesis, resulted in a delay in tumor growth with 45% reduction in maximal tumor depth in tumor Z chambers in a dose-dependent manner. These studies demonstrate the expression of functional receptors for EPO in breast cancer cells. PMID- 12118094 TI - Both constitutive and inducible prostaglandin H synthase affect dermal wound healing in mice. AB - In an attempt to define the roles of prostaglandin H synthase 1 (PGHS-1, cyclooxygenase-1, COX-1) and prostaglandin H synthase 2 (PGHS-2, cyclooxygenase 2, COX-2) in wound healing, we investigated the healing of incisional dermal wounds in wild-type, PGHS-1 null, and PGHS-2 null mice. We measured tensile strength of the wounds, levels of PGHS-1 and PGHS-2 mRNA in the wound site, and histologic markers for the inflammatory, proliferative, and remodeling phases of wound healing. Although no gross visible differences were noted among healed wounds of the different mouse types, measurement of tensile strength showed that both PGHS-1 and PGHS-2 null wounds were weaker (75% and 70%, respectively) than wild-type wounds at 12 days after incision. At Day 8 the endothelial staining was 70% greater in the wounds of PGHS-2 null mice compared with their wild-type counterparts. In contrast at Day 12, staining for macrophages and myofibroblasts was less in PGHS-1 null wounds compared with wild-type and PGHS-2 null tissue. Compensatory expression of the alternate PGHS mRNA could be demonstrated by RT PCR in the wounds of PGHS null mice on Days 1 and 4. We conclude that both PGHS-1 and PGHS-2 genes play distinct roles in the process of dermal wound healing. PMID- 12118095 TI - CCR3-blocking antibody inhibits allergen-induced eosinophil recruitment in human skin xenografts from allergic patients. AB - Eosinophil, basophil, and T helper 2 (TH2) cell recruitment into tissues is a characteristic feature of allergic diseases. These cells have in common the expression of the chemokine receptor CCR3, which may represent a specific pathway for their accumulation in vivo. Although animal models of allergic reactions are available, findings cannot always be extrapolated to man. To overcome these limitations, we have developed a humanized mouse model of allergic cutaneous reaction using severe combined immunodeficiency mice engrafted with skin and autologous peripheral blood mononuclear cells from allergic donors. Intradermal injection of the relevant allergen into human skin xenografts from allergic individuals induced a significant recruitment of human CD4(+) T cells, basophils, and TH2-type cytokine mRNA-expressing cells, as well as murine eosinophils. Human skin xenografts, atopic status, and autologous peripheral blood mononuclear cell reconstitution were all mandatory to induce the allergic reaction. Next, we addressed the role of CCR3 in the endogenous mechanisms involved in the inflammatory cell recruitment in this experimental model of allergic cutaneous reaction. In vivo administration of an anti-human CCR3-blocking antibody selectively reduced accumulation of eosinophils but not that of CD4(+) cells, basophils, or cells expressing mRNA for TH2-type cytokines. These findings establish a new in vivo model of humanized allergic reaction and suggest that eosinophil migration is mediated mainly through CCR3. Finally, these results suggest that this model might be useful to test human-specific antiallergic modulators. PMID- 12118096 TI - Failure of BCL-2 up-regulation in proximal tubular epithelial cells of donor kidney biopsy specimens is associated with apoptosis and delayed graft function. AB - SUMMARY: In renal transplantation, postischemic acute renal failure (ARF) develops in more than 20% of patients. We investigated whether tubular epithelial cells obtained from donor kidneys without subsequent ARF express a different pattern of survival genes, compared with cells from kidneys exhibiting ARF. Donor kidney biopsy specimens were obtained before transplantation from eight recipients of cadaveric kidneys with primary graft function (CAD-PF), eight patients with biopsy-proven ARF without rejection (CAD-ARF), and eight recipients of living donor kidneys with primary graft function (LIV). One thousand proximal tubular epithelial cells per biopsy specimen were isolated by laser capture microdissection. Quantitative analysis of apoptosis and the apoptosis regulatory genes Bcl-2, Bcl-xL, and Bax were performed by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-digoxigenin nick-end labeling staining and real-time PCR, respectively. Primary cultures of human proximal tubular epithelial cells served as calibrator. The number of apoptotic cells was significantly higher in CAD-ARF compared with LIV and CAD-PF (1.5 +/- 1.1% [p < 0.05] vs. 0.3 +/- 0.2% vs. 0.4 +/- 0.2%; mean +/- SD). The apoptosis inhibitors Bcl-2 and Bcl-xL were significantly up-regulated in renal tubular cells of recipients without ARF compared with CAD-ARF. The ratios of Bcl-2/GAPDH normalized to calibrator were as follows: LIV 48 +/- 30, CAD-PF 38 +/- 55, and CAD-ARF 5 +/- 7 (p < 0.05). The corresponding ratios for Bcl-xL were as follows: LIV 6 +/- 6, CAD-PF 5 +/- 3, and CAD-ARF 1 +/- 1 (p < 0.05). No difference in the expression of the proapoptotic Bax could be observed. These data suggest that failure of proximal tubular cells to respond to injury by up-regulation of survival factors from the Bcl-2 family contributes to postischemic ARF in patients after cadaveric renal transplantation. PMID- 12118097 TI - Cyclic strain stimulates early growth response gene product 1-mediated expression of membrane type 1 matrix metalloproteinase in endothelium. AB - SUMMARY: Matrix metalloproteinases (MMPs) are hypothesized to be involved in the processes of endothelial cell (EC) migration and matrix remodeling during angiogenesis. Although hemodynamic forces (such as blood pressure, wall tension, and shear stress) are considered to be strong stimuli for angiogenesis, the role of hemodynamic forces on the regulation of MMPs including membrane type 1 matrix metalloproteinase (MT1-MMP) has not been fully elucidated. To study this, rat microvascular EC were exposed to 60 cycles/minute of 24% maximum strain for up to 24 hours. MT1-MMP mRNA and protein increased in a time-dependent manner through 24 hours of exposure to cyclic strain. Cyclic strain induced early growth response gene product (Egr-1) mRNA and protein within 1 hour. A specific nucleoprotein complex was formed when an oligonucleotide containing binding sites for Sp1 and Egr-1 was incubated with nuclear extracts from EC exposed to 1 hour of cyclic strain. Antibodies to Egr-1 completely supershifted this complex. Increased binding of Egr-1 by cyclic strain to the MT1-MMP promoter correlated with enhanced transcriptional activity. These results suggest that cyclic strain up-regulates the Egr-1-mediated expression of MT1-MMP in rat microvascular EC, emphasizing the importance of hemodynamic forces in the regulation of MT1-MMP in vivo. PMID- 12118098 TI - Epstein-Barr virus nuclear antigen (EBNA)-1 carboxy-terminal and EBNA-4 sequence polymorphisms in nasal natural killer/T-cell lymphoma in the United States. AB - Epstein-Barr virus (EBV) polymorphisms were examined in 12 cases of nasal natural killer (NK)/T-cell lymphoma diagnosed in the United States (U.S.-NL) with respect to the EBV-associated nuclear antigen (EBNA)-1 carboxy (C)-terminal region and the EBNA-4 region. A single dominant EBV strain was found in all cases. EBNA-1 sequences were remarkably homogeneous, showing either a P-ala (2/12) or P-ala variant (9/12) sequence. Other EBNA-1 subtypes known to be common in U.S. reactive samples, such as P-thr or V-leu, were not identified. The final case had a base deletion with frame shift and premature stop codon. EBNA-1 C-terminal amino acid substitutions were common at codons 499 (10/12 cases), 502 (7/12), 524 (9/12), and 528 (6/12), all previously reported "hot spots." However, unlike previous reports of other EBV-associated neoplastic and reactive tissues, mutations were absent at residues 487 and 492. Mutations within HLA-A11 restricted immunogenic EBNA-4 epitopes 399-408 and 416-424 occurred in 3 of 12 cases but were not associated with HLA-A11 status. In summary, the exclusive finding of P-ala variant or P-ala EBNA-1 sequences in U.S.-NL cases differs from that reported in U.S.-reactive and non-U.S.-NL cases. Although the significance of this difference is not known for certain, it may be related to geographic and/or site-specific variations, rather than oncogenicity per se. PMID- 12118099 TI - Separating the wheat from the chaff: focus on "in silico data filtering to identify new angiogenesis targets from a large in vitro gene profile data set". PMID- 12118100 TI - QTL associated with blood pressure, heart rate, and heart weight in CBA/CaJ and BALB/cJ mice. AB - To better understand the genetic basis of essential hypertension, we conducted a quantitative trait locus (QTL) analysis of a population of 207 (BALB/cJ x CBA/CaJ) F(2) male mice to identify genomic regions that regulate blood pressure, heart rate, and heart weight. We identified two loci, Bpq6 (blood pressure quantitative locus 6) on chromosome 15 (Chr 15; peak, 16 cM; 95% confidence interval, 0-25 cM) and Bpq7 on Chr 7 (peak, 42 cM; 95% confidence interval, 35-50 cM) that were significantly associated with blood pressure. We also identified two loci, Hrq1 (heart rate quantitative locus 1) and Hrq2, on D2Mit304 (peak, 72 cM; 95% confidence interval 60-80 cM) and D15Mit184 (peak, 25 cM; 95% confidence interval 20-35 cM), respectively, that were significantly associated with heart rate. A significant gene-gene interaction for heart rate was found between Hrq1 and D1Mit10 (peak, 57 cM; 95% confidence interval, 45-75 cM); the latter QTL was named Hrq3. We identified a significant locus for heart weight, Hwq1 (heart weight quantitative locus 1), at D14Mit67 (peak, 38 cM; 95% confidence interval, 20-43 cM). Identification of the genes for these QTL should lead to a better understanding of the causes of essential hypertension. PMID- 12118101 TI - In silico data filtering to identify new angiogenesis targets from a large in vitro gene profiling data set. AB - The objective of this study was to use gene expression data from well-defined cell culture models, in combination with expression data from diagnostic samples of human diseased tissues, to identify potential therapeutic targets and markers of disease. Using Affymetrix oligonucleotide array technology, we identified a common profile of genes upregulated during endothelial morphogenesis into tubelike structures in three in vitro models of angiogenesis. Rigorous data selection criteria were used to identify a list of over 1,000 genes whose expression was increased more than twofold over baseline at either 4, 8, 24, 40 or 50 h. To further refine and prioritize this list, we used standard bioinformatic algorithms to identify potential transmembrane and secreted proteins. We then overlapped this gene set with genes upregulated in colon tumors vs. normal colon, resulting in a subset of 128 genes in common with our endothelial list. We removed from this list those genes expressed in 6 different colon tumor lines, resulting in a list of 24 putative, vascular-specific angiogenesis-associated genes. Three genes, gp34, stanniocalcin-1 (STC-1), and GA733-1, were expressed at levels 10-fold or more in colon tumors compared with normal mucosa. We validated the vascular-specific expression of one of these genes, STC-1, by in situ hybridization. The ability to combine in vitro and in vivo data sets should permit one to identify putative angiogenesis target genes in various tumors, chronic inflammation, and other disorders where therapeutic manipulation of angiogenesis is a desirable treatment modality. PMID- 12118102 TI - Pleiotropy of quantitative trait loci for organ weights and limb bone lengths in mice. AB - We investigated the genetic basis of several limb bone lengths and weights of organs in mice produced from a cross of the F1 between CAST/Ei (wild strain) and M16i (selected for rapid growth rate) back to M16i. From previous correlation studies, we hypothesized that quantitative trait loci (QTLs) would exhibit greater pleiotropy within than between the limb length and organ weight character sets. Using interval mapping procedures and significance testing at the chromosome-wise level, we discovered 14 putative QTLs affecting weight of the liver, spleen, heart, and/or kidney, 9 of which affected more than one organ; and 12 QTLs for limb lengths, all of which affected the length of two or more of the limb bones in these mice. As was hypothesized, most QTLs affected either organ weights or limb lengths independently of each other, although five QTLs were found that affected both sets of characters. The direction of the effect of these QTLs was almost always consistent within and between characters, with little evidence for antagonistic pleiotropy. PMID- 12118103 TI - Microarray gene expression profiles in dilated and hypertrophic cardiomyopathic end-stage heart failure. AB - Despite similar clinical endpoints, heart failure resulting from dilated cardiomyopathy (DCM) or hypertrophic cardiomyopathy (HCM) appears to develop through different remodeling and molecular pathways. Current understanding of heart failure has been facilitated by microarray technology. We constructed an in house spotted cDNA microarray using 10,272 unique clones from various cardiovascular cDNA libraries sequenced and annotated in our laboratory. RNA samples were obtained from left ventricular tissues of precardiac transplantation DCM and HCM patients and were hybridized against normal adult heart reference RNA. After filtering, differentially expressed genes were determined using novel analyzing software. We demonstrated that normalization for cDNA microarray data is slide-dependent and nonlinear. The feasibility of this model was validated by quantitative real-time reverse transcription-PCR, and the accuracy rate depended on the fold change and statistical significance level. Our results showed that 192 genes were highly expressed in both DCM and HCM (e.g., atrial natriuretic peptide, CD59, decorin, elongation factor 2, and heat shock protein 90), and 51 genes were downregulated in both conditions (e.g., elastin, sarcoplasmic/endoplasmic reticulum Ca2+-ATPase). We also identified several genes differentially expressed between DCM and HCM (e.g., alphaB-crystallin, antagonizer of myc transcriptional activity, beta-dystrobrevin, calsequestrin, lipocortin, and lumican). Microarray technology provides us with a genomic approach to explore the genetic markers and molecular mechanisms leading to heart failure. PMID- 12118104 TI - C-kit expression distinguishes salivary gland adenoid cystic carcinoma from polymorphous low-grade adenocarcinoma. AB - Adenoid cystic carcinoma (ACC) is characterized by persistent, relentless growth and a high rate of eventual metastasis. In contrast, polymorphous low-grade adenocarcinoma (PLGA) has a much lower risk of recurrence and rarely metastasizes. The histologic patterns of these two neoplasms can be similar. Expression of c-kit, a transmembrane receptor tyrosine kinase, has recently been reported to be expressed in ACC but not PLGA. Expression of galectin-3, a nonintegrin beta-galactosidase-binding lectin, has been reported to be significant in PLGA and decreased in ACC.Formalin-fixed paraffin-embedded tissue from 9 ACC and 14 PLGA were immunostained for c-kit and galectin-3. Cases were scored as 1+ (5-25% positive), 2+ (26-50% positive), or 3+ (>50% positive). C-kit was expressed by 100% of ACC (3+: 7 cases; 2+: 1 case; 1+: 1 case) and by 57% of PLGA (2+: 2 cases; 1+: 6 cases). In all but one ACC, c-kit expression was confined to the inner cell layer. C-kit expression was also noted in the intercalated duct epithelium of the salivary glands and the acinar cells of the lacrimal gland. Galectin-3 was expressed in 8 of 9 cases of ACC and 14 of 14 cases of PLGA. The results of this, the first study to compare c-kit and galectin 3 expression in ACC and PLGA, suggest that c-kit expression characterizes ACC, but not PLGA. Galectin-3 immunohistochemistry does not have a role in the differentiation of ACC and PLGA. C-kit immunostaining may be a valuable adjunctive tool for this differential diagnosis, particularly in the setting of a limited biopsy. Our finding of different patterns of c-kit expression in tubular and solid variants of ACC supports the concept of solid variant ACC as a high grade tumor, with progression toward an entirely "inner cell" phenotype. PMID- 12118105 TI - Complex genetic alterations in gastrointestinal stromal tumors with autonomic nerve differentiation. AB - Gastrointestinal stromal tumors (GISTs) with neurogenic differentiation, also referred to as "gastrointestinal autonomic nerve tumors (GANTs)," form an ultrastructurally distinctive subgroup of mesenchymal neoplasms of gastrointestinal tract. Cytogenetic and molecular data of these tumors are limited. In the current study, c-KIT gene sequenc-ing analysis, comparative genomic hybridization (CGH), and interphase fluorescence in situ hybrid- ization (FISH) analysis, utilizing chromosome 14- and 22-specific probes, were performed on five primary ultrastructurally confirmed GANTs. FISH and CGH analysis revealed loss of a whole or part of chromosome 14q in two tumors and of chromo- some 22q, with the common overlapping area of loss at q13, in all five tumors evaluated. c KIT mu- tations were found in all cases; three tumors carried point mutation and/or deletions of exon 11, and in two tumors, insertion in exon 9 was found. These findings suggest that accumulated genetic changes contribute to the pathogenesis of GANTs and that 22q13 loss may be a characteristic feature of these tumors. PMID- 12118106 TI - Kupffer cell aggregation and perivenular distribution in steatohepatitis. AB - Cytokine release from inflammatory cells, endotoxin, lipid peroxidation, and generation of reactive oxygen species are among the factors currently thought to be important in the pathogenesis of alcoholic and nonalcoholic steatohepatitis (SH). To more fully evaluate the role of mononuclear inflammatory cells in SH, 11 needle liver biopsies showing SH were selected for immunohistochemical staining to analyze the type and distribution of mononuclear inflammatory cells, including T and B lymphocytes and Kupffer cells (using immunostains for CD3, CD4, CD8; CD20; and CD68, respectively). An additional seven biopsies showing normal or fatty liver were also selected for CD68 immunostaining. Immunohistochemistry showed mild to moderate (1+ to 2+) numbers of T cells, with equal representation of CD4 and CD8 cells. T cells were found in portal tracts and in regions of SH. B cells were only rarely present. CD68 staining of simple fatty liver and normal liver showed elongated, spindle-shaped Kupffer cells diffusely distributed along the sinusoids throughout the lobules. In contrast, in cases of SH, there was prominent enlargement and aggregation of Kupffer cells in perivenular regions. Scattered large vacuoles of fat that had appeared to be within hepatocytes on routine stain were found actually to be within Kupffer cells. These results support the concept that hepatic Kupffer cells are a major immune effector cell in the pathogenesis of steatohepatitis. A potential direct Kupffer cell role in hepatic lipid processing is also postulated. PMID- 12118107 TI - Monoclonal antibody specific for histone H1 phosphorylated by cyclin-dependent kinases: a novel immunohistochemical probe of proliferation and neoplasia. AB - Monoclonal antibody 12D11 (MAb 12D11) has been shown to bind histone H1 isolated from human placenta and other tissues but not histone H1 that has been digested with bacterial alkaline phosphatase. We show here that phosphorylation of phosphatase-treated histone H1 with cyclin dependent-kinase (CDK) restores binding by MAb 12D11. We conclude that MAb 12D11 selectively binds histone H1 that has been phosphorylated by CDKs, and we have investigated the use of MAb 12D11 as an immunohistochemical probe of CDK activity in situ. Previous immunofluorescence studies have revealed strong nuclear staining by MAb 12D11 in proliferating cultured cells and the absence of staining in terminally differentiated cells. Immunohistochemical staining of frozen and formalin-fixed, paraffin-embedded sections of benign tissues with MAb 12D11 was nuclear and confined to recognized foci of cell proliferation. In lymphoid germinal centers, MAb 12D11 preferentially stained large lymphoid cells with a relative lack of staining in small cleaved cells, contrasting with a lack of cell size discrimination observed with the monoclonal antibody proliferation probe, MIB-1. Tumor tissues displayed strong albeit heterogeneous staining of malignant cells by MAb 12D11, with little or no staining observed in surrounding nonneoplastic stromal cells. Differential staining by MAb 12D11 of invasive and in situ carcinoma suggest applications in prognostication. MAb 12D11 may also be useful in identification of tumors more likely to respond to therapeutic CDK inhibitors. PMID- 12118108 TI - Cytokeratins 7 and 20 immunoreactivity in chromophobe renal cell carcinomas and renal oncocytomas. AB - Chromophobe renal cell carcinomas and renal oncocytomas share morphologic similarities and may present a diagnostic challenge on routine hematoxylin-eosin staining. Currently recommended additional studies of Hale's colloidal iron staining and electron microscopy are often difficult to interpret and technically challenging and may not be readily available. Previous studies have reported conflicting results with regard to the cytokeratin 7 staining pattern in chromophobe renal cell carcinomas and renal oncocytomas. Cytokeratin 20 expression in chromophobe renal cell carcinomas has not previously been studied. Formalin-fixed paraffin-embedded tissue of 11 chromophobe renal cell carcinomas and 21 renal oncocytomas were retrieved from the archived files (1984-2000) of four teaching hospitals. Of the 11 chromophobe renal cell carcinomas, eight stained positive (73%) for cytokeratin 7, one stained focally positive (9%), and two cases (18%) were completely negative. Cytokeratin 7 staining of the 21 oncocytomas revealed 4 positive (19%), 7 focally positive (33%), and 10 negative cases (48%). Cytokeratin 20 was uniformly negative on all 11 cases of chromophobe renal cell carcinomas and all 21 cases of oncocytomas. Cytokeratin 7 does not appear to show the consistent immunoreactivity in chromophobe renal cell carcinomas and renal oncocytomas, as has been previously suggested. Cytokeratin 20 immunostaining in chromophobe renal cell carcinomas and renal oncocytomas is uniformly negative. Despite the technical and interpretive challenges of Hale's colloidal iron, it is still the most useful stain in differentiating chromophobe renal cell carcinomas from renal oncocytomas. PMID- 12118109 TI - Sporadic fundic gland polyposis: a clinical, histological, and molecular analysis. AB - Sporadic fundic gland polyposis (SFGP) is defined as multiple fundic gland polyps in patients without familial adenomatous polyposis syndrome (FAP). Although little is known about the genetic changes in SFGP, mutations in the Wnt signaling pathway have been recently linked to fundic gland polyps in other settings: sporadic polyps are linked to activating beta-catenin mutations, whereas FAP associated fundic gland polyps are caused by second somatic hits in the adenomatous polyposis coli gene. The relationship between SFGP, single sporadic fundic gland polyps, and FAP-associated polyps remains unclear, and SFGP remain poorly characterized at the clinical, histological, and molecular levels. A retrospective study was undertaken of eight patients with SFGP who had >/=10 polyps with at least five endoscopic biopsy specimens available for study. One additional patient with attenuated FAP who underwent partial gastrectomy was included as a control. The medical records and biopsy specimens were reviewed. Mutations of the beta-catenin gene were evaluated in each fundic gland as well as in control nonpolypoid tissue by direct sequencing of a mutational hot spot in exon 3 of the beta-catenin gene, which encodes the GSK-3beta phosphorylation sites, and a HinfI endonuclease digestion assay. The four men and four women in the study were an average of 57 years of age at biopsy. All patients were on acid suppression therapy, 5/8 with proton-pump inhibitors (PPI) and 3/8 with Zantac. Sixty-two polyps were studied, and all were <10 mm, with most between 2 and 7 mm. The polyps were histologically identical to single sporadic fundic gland polyps. No dysplasia was seen. Forty-seven of 62 polyps (76%) had detectable beta-catenin mutations. Mutations were found in all eight of the patients. All were point mutations in codons 32, 33, 34, and 37 and are either phosphorylation sites or immediately adjacent to phosphorylation sites, findings identical to that seen in single sporadic fundic gland polyps. Each polyp had a single mutation, and each patient had more than one unique mutation (median = 4), indicating a multifocal origin for the polyps. No mutations were found in nonpolypoid control tissue and in polyps from the attenuated FAP patient. The patients with SFGP in this series were all between 40 and 70 years of age and had histories of acid-suppressive therapy. The fundic gland polyps were histologically and genetically identical to single sporadic fundic gland polyps and demonstrated frequent somatic activating mutations in exon 3 of the beta-catenin gene. PMID- 12118110 TI - Hybrid carcinomas of the salivary glands: report of nine cases with a clinicopathologic, immunohistochemical, and p53 gene alteration analysis. AB - Hybrid carcinomas of the salivary gland are a recently defined and rare tumor entity, consisting of two histologically distinct types of carcinoma within the same topographic area. In this study, we examined nine such cases, which mainly arose in the parotid gland (seven cases), with an additional one each from submandibular and lacrimal glands, and analyzed their clinicopathologic profiles, including immunohistochemical features and p53 gene alterations. The prevalence of hybrid carcinomas was 0.4% among the 1863 cases of parotid gland tumors in our series. The nine patients comprised five men and four women, ranging in age from 40 to 81 years (mean, 62 y). Tumor size ranged from 2 to 10 cm (mean, 4.2 cm). Of the seven patients who were followed up, two were alive with disease and five were alive with no evidence of disease, although the follow-up period was short. Three cases had cervical lymph nodal metastases. The combinations of carcinoma components in our hybrid carcinomas were as follows: epithelial-myoepithelial carcinoma and basal cell adenocarcinoma in two cases, epithelial-myoepithelial carcinoma and squamous cell carcinoma in one case, salivary duct carcinoma and adenoid cystic carcinoma in two cases, myoepithelial carcinoma and salivary duct carcinoma in one, acinic cell carcinoma and salivary duct carcinoma in one, and squamous cell carcinoma and salivary duct carcinoma in two. Although the proportion of each carcinoma component in a tumor mass varied from case to case, the minor component always represented >or= 10% of the area. Differences in cellular composition were studied by immunohistochemistry and electron microscopy. The Ki-67-labeling index apparently differed between the two carcinoma elements in five cases. Diffusely positive p53 immunoreactivity was observed in four cases, restricted to the more aggressive component in each pair. Furthermore, p53 gene alteration analysis of these p53-positive cases revealed that all and three cases demonstrated loss of heterozygosity at p53 microsatellite loci and p53 gene point mutations, respectively, which were detected only in the p53-immunoreactive carcinoma component. Therefore, there is the possibility that such molecular-genetic events take an integral part for inducing the transformation from histologically lower to higher grade tumor during the hybrid carcinoma genesis of the salivary glands. PMID- 12118111 TI - Increased p53 protein expression in malignant mammary phyllodes tumors. AB - The authors reviewed 143 cases (87 benign, 37 borderline, and 19 malignant) of mammary phyllodes tumors (PTs) and used immunohistochemistry to detect p53 protein product semi-quantitatively as negative, weak, moderate and strong (scored 0 to 3). For all PTs, an increasing trend of tumor size and malignancy was detected with increasing age. For p53 staining, 60 cases (42%) were negative, 55 (38%) stained weakly, 28 (13%) stained moderately, and 10 (7%) stained strongly. Of the 87 benign PTs, 41 (47%) were negative, 37 (43%) stained weakly, and 9 (10%) stained moderately. For the 37 borderline PTs, 16 (43%) were negative, 14 (38%) stained weakly, 6 (16%) stained moderately, and 1 (3%) stained strongly. Of the 19 malignant PTs, 3 (16%) were negative, 4 (21%) stained weakly, 3 (16%) stained moderately, and 9 (47%) stained strongly. The mean intensity score for p53 staining increased progressively from benign to borderline to malignant PT, with established statistical significance (P <.0001). This is significantly correlated with mitotic count but not stromal cellularity, pleomorphism, margin, and stromal overgrowth. When considering strong staining alone (score, 3), 47% of malignant, 3% of borderline, and none of the benign PTs were positive. The use of strong positive staining for diagnosing malignant PT gave positive and negative predictive values, specificity, and sensitivity of 90%, 92.5%, 99%, and 47%, respectively. Thus diffuse strong p53 protein staining can be used as a soft sign in assisting the diagnosis of malignant PT. Conversely, negative or weak staining of p53 protein in PT is of little discriminatory value. The role of p53 gene mutation in the malignant transformation of PT is unclear; but this may not be the sole mechanism as many malignant PT were p53 protein negative. PMID- 12118113 TI - Centroblastic and centroblastic/centrocytic lymphoma associated with a prominent epithelioid granulomatous response: a clinicopathologic study of 50 cases. AB - A minority of centroblastic and centroblastic/centrocytic cell lymphomas are accompanied by a prominent epithelioid cell response and were suggested to be a distinct variant of B-cell lymphoma of germinal center cell origin. To confirm the clinicopathologic significance of these mainly large B-cell lymphomas with an epithelioid cell response (LBCL-ER), we reviewed 50 patients with LBCL-ER and compared the results with those of 167 other diffuse large B-cell lymphomas (DLBCL) and 94 follicular lymphomas (FL) without epithelioid response. The patients with LBCL-ER showed a higher age distribution (median 71, P =.03), a female predominance (M:F = 18:32, P =.001) and less frequent involvement of extranodal sites >1 (P =.004) compared with those with DLBCL, and presented with a bulky mass of the affected lymph nodes in 54% of cases. They were also older (P =.0006) and more associated with the aggressive clinical factors such as serum LDH level and International Prognostic Index score than those with FL. Histologically, nine cases (18%) partially showed a follicular growth pattern, and the others (82%) were occupied by a diffuse growth pattern. The epithelioid cells were accumulated in large demarcated masses, partially imparting a lymphoepithelioid (Lennert) lymphoma-like appearance to some portions of the lesions in every case. Immunohistochemically, LBCR-ER was positive for CD20 in every case, CD10 in 43% of the cases, and BCL-2 in 56%. None of the tumor cells in the 40 cases tested expressed CD5 antigen. Immunostaining also often highlighted the remnants of the follicular dendritic cell network. The BCL-2 gene rearrangement was detected in only 19% of the cases examined. The survival curve of the cases of LBCL-ER was almost identical with that of DLBCL and was significantly inferior to that of FL. The centroblastic and centroblastic/centrocytic lymphoma with an epithelioid cell response may be regarded as the morphologic variant of DLBCL preferentially arising in the aged population and reflecting the disease progression of FL. PMID- 12118112 TI - Immunohistochemical pattern of MLH1/MSH2 expression is related to clinical and pathological features in colorectal adenocarcinomas with microsatellite instability. AB - Detection of colorectal carcinomas with high-frequency microsatellite instability (MSI-H) is clinically important for several reasons. Recent studies suggested that immunohistochemical analysis of MLH1 and MSH2 expression is a rapid and accurate method for identifying large bowel tumors of the MSI-H phenotype. In this study, we evaluated by immunohistochemistry MLH1 and MSH2 protein expression in 132 MSI-H, 23 MSI-L (low-frequency MSI), and 150 microsatellite stable (MSS) colorectal adenocarcinomas. Loss of MLH1 or MSH2 expression was detected in 120 (90.9%) MSI-H carcinomas, whereas all MSI-L and MSS tumors showed normal expression of both proteins. Lack of MLH1 nuclear staining was observed much more often than absence of MSH2 nuclear staining (106 and 14 cases, respectively). Among MSI-H carcinomas, MLH1/MSH2 pattern of expression was significantly related to several clinical and pathological variables. In particular, MSI-H MLH1/MSH2 positive carcinomas were more often located in the distal colon, were more frequently classified as ordinary adenocarcinomas, and were more likely to be well or moderately differentiated, p53 positive, and <7 cm in diameter than were MLH1-negative and MSH2-negative carcinomas. In addition, MLH1-negative carcinomas were less common among patients with hereditary nonpolyposis colorectal cancer (HNPCC) or suspected HNPCC and in the group of patients aged <50 years. Patients with MLH1-negative carcinomas more frequently died of disease than did patients with MLH1/MSH2-positive and MSH2-negative MSI-H tumors, but the difference was not statistically significant. The results of the present investigation strongly indicate that immunohistochemical analysis of MLH1 and MSH2 expression is a practical and reliable method for the routine detection of the vast majority of MSI-H large bowel adenocarcinomas. Our data also point out that MSI-H MLH1/MSH2 positive colorectal carcinomas are characterized by distinctive pathological features. PMID- 12118114 TI - Detection of Epstein-Barr virus in rapidly growing fibroadenomas of the breast in immunosuppressed hosts. AB - Fibroadenomas are the most common benign tumors of the female breast and are associated with a slight increase in the risk of subsequent breast cancer. Multiple fibroadenomas have been described in patients after renal transplantation and are thought to be secondary to drug-related growth stimulation. Epstein-Barr virus (EBV) has been detected in many neoplasms, including breast cancer. We set out to investigate whether EBV plays a role in the development of rapidly growing fibroadenomas in immunocompromised patients. We studied 19 fibroadenomas and one invasive ductal carcinoma that developed after organ transplantation or treatment for lupus erythematosus. As a control group we included 11 fibroadenomas from non-immunocompromised patients. DNA was amplified using polymerase chain reaction (PCR) of the EBV-encoded small RNA (EBER-2) DNA sequence. EBV latent membrane protein 1 (LMP-1) transcripts were amplified using reverse transcription (RT) PCR. Immunohistochemical (IHC) staining for LMP-1 protein was performed. A total of 9 out of 20 tumors (45%) were concordantly positive by PCR and IHC. IHC stained exclusively the epithelial cells. All the fibroadenomas in non-immunocompromised patients were negative for LMP-1 (Fisher's exact test P =.0006). These data suggest that EBV is associated with fibroadenomas in this immunosuppressed population and that the infection is specifically localized to epithelial cells. This is the first study suggesting a role for EBV in the pathogenesis of fibroadenomas. PMID- 12118115 TI - Symptomatic nephrogenic metaplasia of ureter: a morphologic and immunohistochemical study of four cases. AB - Nephrogenic metaplasia of the bladder and urethra has been the subject of extensive studies in recent years. However, information about ureteral involvement is still limited because of the rarity of the lesion. We described four cases of nephrogenic metaplasia of the ureter. They occurred in two men and two women whose ages ranged from 46 to 69 years. Three patients had stones, and one had multiple episodes of cystitis and chronic pyelonephritis. The lesions led to ureteral obstruction that in two patients was radiographically suspicious for carcinoma. Microscopically, three lesions were composed of tiny mucin-containing microcysts and medium-sized tubular structures lined by cuboidal cells that showed cytologic atypia characterized by enlarged vesicular nuclei and prominent nucleoli. However, there were no mitotic figures. Two lesions invaded the full thickness of the wall of the ureter and exhibited an infiltrative growth pattern highlighted by cytokeratin stains. The remaining two lesions were confined to the lamina propria. The cells of nephrogenic metaplasia were immunoreactive to cytokeratin 7 and AE1-AE3. They lacked reactivity for monoclonal and polyclonal CEA and p53. The MIB-1-labeling index was <5%. The cytologic atypia and infiltrative growth pattern of ureteral nephrogenic metaplasia should not be misinterpreted as evidence of malignancy. All four patients are alive and symptom free 8 months to 7 years after diagnosis. PMID- 12118116 TI - The distinction between Burkitt lymphoma and diffuse large B-Cell lymphoma with c myc rearrangement. AB - To compare immunophenotypic and molecular features between Burkitt lymphoma (BL) and diffuse large B-cell lymphoma (DLBCL) with c-myc rearrangements (c-mycR DLBCL), we analyzed 18 cases of B-cell non-Hodgkin's lymphoma with c-mycR that were confirmed by chromosomal and/or Southern blotting analyses. The cases were histologically classified into 10 BLs and five DLBCLs. The remaining three cases could not be classified because of suboptimal quality of the surgical materials. BLs were from five adults and five children, whereas all DLBCLs were from adults. BLs were positive for CD20 (10/10 cases examined), CD10 (9/10), Bcl-2 (1/9), and Bcl-6 (10/10), whereas they were negative for CD3 (0/10) and EBV (0/8), by Epstein-Barr virus (EBV) EBER-1 RNA in situ hybridization. c-MycR DLBCLs were positive for CD20 (5/5), CD10 (2/5), Bcl-2 (3/4), and Bcl-6 (4/4), whereas none of them were positive for CD3 and EBV. A mean of MIB-1 index (MIB-1+ cells/neoplastic cells, %) of BLs (98.1%) was higher than that of c-mycR DLBCLs (66.3%; P <.0001). Somatic mutation of immunoglobulin heavy-chain gene variable region (VH gene) in BLs (four cases) ranged from 0.7 to 4.9% with an average value of 2.3%, whereas those in DLBCLs (three cases) from 8.2 to 32.0% with an average value of 17.0%. It is, therefore, concluded that a growth fraction of nearly 100%, as well as a monotonous proliferation of medium-sized cells and c myc(R), should be of value in the diagnosis of BL, which is probably different from c-myc(R) DLBCL. In addition, CD10+, Bcl-2-, and low frequency of mutation of the VH gene could be helpful for the histologic distinction of BL from (c-mycR) DLBCL. PMID- 12118117 TI - Cartilaginous differentiation in peritoneal tissues: a report of two cases and a review of the literature. AB - Two cases of cartilaginous differentiation of the peritoneum not associated with an intraabdominal malignancy are described. This is the first detailed report of cartilaginous metaplasia of the peritoneum. The patients were female, ages 53 (Patient 1) and 77 years (Patient 2). Prior medical histories were significant for a culdotomy (to drain pelvic abscesses associated with pelvic inflammatory disease) in Patient 1 and for an open abdominal surgery in Patient 2. The peritoneal lesions were incidental findings in both cases. In Patient 1, surgery was performed for a septated ovarian cyst; the other patient underwent surgery to relieve obstructive bowel symptoms. In Patient 1, multiple firm, white lesions ranging from 2.0 to 7.0 mm were present on the serosal surfaces and the mesenteries of the small and large bowel. In Patient 2, a single firm, white lesion measuring 2 cm in maximum dimension was removed from the mesentery of the ileum. Microscopically, the lesions consisted of small nodules of mature hyaline cartilage surrounded by nondescript fibrous tissue and covered by mesothelium. There was no foreign body giant cell reaction, inflammation, or other reactive changes in the surrounding adipose tissue. These may represent metaplastic lesions of the secondary mullerian system, or a unique peritoneal response to previous surgical manipulation. Alternatively, these may represent benign neoplastic lesions (chondroma) of the submesothelium. PMID- 12118118 TI - Synergism between trematode infection and pesticide exposure: a link to amphibian limb deformities in nature? AB - The apparently rapid increase in the prevalence of amphibian limb deformities has led to substantial interest from ecologists and public health professionals. Hypotheses proposed to explain the deformities fall into two broad categories: chemical contaminants and trematode infection. Although there are convincing experimental demonstrations that certain factors can lead to some deformities, the causes for recent increases in amphibian malformation remain controversial. Moreover, no experimental studies on amphibian deformities have been conducted in the field, and no studies have attempted to examine the synergistic effects of trematode infection and exposure to chemical contaminants. Here, I present the results of field and laboratory experiments that link increased trematode infection, and increased limb deformities, to pesticide exposure. Field experiments conclusively demonstrated that exposure to trematode infection was required for the development of limb deformities in wood frogs, Rana sylvatica. However, deformities were more common at sites adjacent to agricultural runoff. Laboratory experiments corroborated the association between pesticide exposure and increased infection with pesticide-mediated immunocompetency as the apparent mechanism. Given the conservative contaminant exposure levels used [Environmental Protection Agency (EPA) drinking water standards] and the widespread use of many pesticides, these negative impacts may help to explain pathogen-mediated amphibian declines in many regions. PMID- 12118119 TI - Vascular endothelial growth factor stimulates bone repair by promoting angiogenesis and bone turnover. AB - Several growth factors are expressed in distinct temporal and spatial patterns during fracture repair. Of these, vascular endothelial growth factor, VEGF, is of particular interest because of its ability to induce neovascularization (angiogenesis). To determine whether VEGF is required for bone repair, we inhibited VEGF activity during secondary bone healing via a cartilage intermediate (endochondral ossification) and during direct bone repair (intramembranous ossification) in a novel mouse model. Treatment of mice with a soluble, neutralizing VEGF receptor decreased angiogenesis, bone formation, and callus mineralization in femoral fractures. Inhibition of VEGF also dramatically inhibited healing of a tibial cortical bone defect, consistent with our discovery of a direct autocrine role for VEGF in osteoblast differentiation. In separate experiments, exogenous VEGF enhanced blood vessel formation, ossification, and new bone (callus) maturation in mouse femur fractures, and promoted bony bridging of a rabbit radius segmental gap defect. Our results at specific time points during the course of healing underscore the role of VEGF in endochondral vs. intramembranous ossification, as well as skeletal development vs. bone repair. The responses to exogenous VEGF observed in two distinct model systems and species indicate that a slow-release formulation of VEGF, applied locally at the site of bone damage, may prove to be an effective therapy to promote human bone repair. PMID- 12118120 TI - A small-molecule inhibitor of the ribonucleolytic activity of human angiogenin that possesses antitumor activity. AB - The results of previous preclinical and clinical studies have identified angiogenin (ANG) as a potentially important target for anticancer therapy. Here we report the design and implementation of a high-throughput screening assay to identify small molecules that bind to the ribonucleolytic active site of ANG, which is critically involved in the induction of angiogenesis by this protein. Screening of 18,310 compounds from the National Cancer Institute (NCI) Diversity Set and ChemBridge DIVERSet yielded 15 hits that inhibit the enzymatic activity of ANG with K(i) values <100 microM. One of these, NCI compound 65828 [8-amino-5 (4'-hydroxybiphenyl-4-ylazo)naphthalene-2-sulfonate; K(i) = 81 microM], was selected for more detailed studies. Minor changes in ANG or ligand structure markedly reduced potency, demonstrating that inhibition reflects active-site rather than nonspecific binding; these observations are consistent with a computationally generated model of the ANG.65828 complex. Local treatment with modest doses of 65828 significantly delayed the formation of s.c. tumors from two distinct human cancer cell types in athymic mice. ANG is the likely target involved because (i) a 65828 analogue with much lower potency against the enzymatic activity of ANG failed to exert any antitumor effect, (ii) tumors from 65828-treated mice had fewer interior blood vessels than those from control mice, and (iii) 65828 appears to have no direct effect on the tumor cells. Our findings provide considerable support for the targeting of the enzymatic active site of ANG as a strategy for developing new anticancer drugs. PMID- 12118123 TI - Role of femoral ring allograft in anterior interbody fusion of the spine. AB - A review was carried out on 59 patients (10 males and 49 females) who had anterior interbody fusion performed with femoral ring allograft packed with autograft bone chips with a minimum follow up of 2 years. The average age at the time of surgery was 49.1 year old (26 to 75). The total number of levels grafted was 141. The diagnosis consisted of multiple degenerative disease in 6, degenerative change below the long segment of fusion for scoliosis in 9, osteoporosis with collapsed fracture in 3, pseudarthrosis after posterior laminectomy and fusion in 35, congenital scoliosis in 3, scoliosis in 2 and paralytic scoliosis due to multiple sclerosis in one. The distribution of levels fused was T12-L1 in 6, L1-2 in 12, L2-3 in 17, L3-4 in 22, L4-5 in 35 and L5-S1 in 39. The remaining 10 levels were in the lower thoracic areas (T7-T12). The operations were performed as anterior fusion alone in 13 patients, one-stage anterior and posterior fusion in 26 patients and two-stage surgery in 20 patients. Anterior instrumentation was used in all 141 levels. At average follow up (33.7 months) there was no significant change in allograft angles (average = 1.6 degrees ). Fusion of the allograft was classified by Bridwell's grading system. At 24 months of the follow up, 97 % of the allografts were in grade I (fully incorporated) and 3% were in grade II (partially incorporated). Compared to 12 months follow-up only 76.2% of the grafts were in grade I, 28 % were in grade II and 0.8% were in grade III. Two patients had deep posterior infections which required further surgery (without resorption of the allograft anteriorly). One patient had a screw migration anteriorly which required removal. Three patients had persistence of radiolucent line at one of the vertebral end plates - graft interfaces but no subsidence of the graft or pain. In conclusion, the femoral ring allograft appeared to benefit the anterior interbody fusion in complex spinal surgery. PMID- 12118121 TI - Bcl-X(L) affects Ca(2+) homeostasis by altering expression of inositol 1,4,5 trisphosphate receptors. AB - An oligonucleotide-based microarray analysis of 9,500 genes and expressed sequence tags (ESTs) demonstrated that the type 1 inositol 1,4,5-trisphosphate receptor (IP(3)R) was significantly down-regulated in Bcl-X(L)-expressing as compared with control cells. This result was confirmed at the mRNA and protein levels by Northern and Western blot analyses of two independent hematopoietic cell lines and murine primary T cells. Bcl-X(L) expression resulted in a dose dependent decrease in IP(3)R protein. IP(3)R expression is regulated as part of a mitochondrion-to-nucleus stress-responsive pathway. The uncoupling of mitochondrial oxidative phosphorylation resulted in induction of binding of the transcription factor NFATc2 to the IP(3)R promoter and transcriptional activation of IP(3)R. Expression of Bcl-X(L) led to a decreased induction of both NFATc2 DNA binding to the IP(3)R promoter and IP(3)R expression in response to the inhibition of mitochondrial oxidative phosphorylation. The Bcl-X(L)-dependent decrease in IP(3)R expression also correlated with a reduced T cell antigen receptor ligation-induced Ca(2+) flux in Bcl-X(L) transgenic murine T cells, and microsomal vesicles prepared from Bcl-X(L)-overexpressing cells exhibited lower IP(3)-mediated Ca(2+) release capacity. Furthermore, reintroducing IP(3)R into Bcl-X(L)-transfected cells partially reversed Bcl-X(L)-dependent anti-apoptotic activity. These results suggest that even under non-apoptotic conditions, expression of Bcl-2-family proteins influences a signaling network that links changes in mitochondrial metabolism to alterations in nuclear gene expression. PMID- 12118122 TI - Emergency response to a smallpox attack: the case for mass vaccination. AB - In the event of a smallpox bioterrorist attack in a large U.S. city, the interim response policy is to isolate symptomatic cases, trace and vaccinate their contacts, quarantine febrile contacts, but vaccinate more broadly if the outbreak cannot be contained by these measures. We embed this traced vaccination policy in a smallpox disease transmission model to estimate the number of cases and deaths that would result from an attack in a large urban area. Comparing the results to mass vaccination from the moment an attack is recognized, we find that mass vaccination results in both far fewer deaths and much faster epidemic eradication over a wide range of disease and intervention policy parameters, including those believed most likely, and that mass vaccination similarly outperforms the existing policy of starting with traced vaccination and switching to mass vaccination only if required. PMID- 12118124 TI - The timing of spinal fusion in adolescent idiopathic scoliosis. AB - 102 cases of idiopathic adolescent scoliosis seen over a period of 5 years were studied. 59 patients who were treated surgically and followed up for a minimum of 48 months, fell into one of two groups: Group I - those operated on within 3 years following the adolescent growth spurt, and Group II - those who were operated on at or after skeletal maturity. 35 patients were treated by Harrington instrumentation and posterior fusion and 24 by Harrington instrumentation, segmental sublaminar wiring and posterior fusion. In 7 patients anterior release was performed initially. In Group I, the extent of deformity correction and elimination of the rib hump were better, and complications such as neurological deficit, hook dislodgement and implant breakage were encountered less frequently. Harrington instrumentation, segmental sublaminar wiring and posterior fusion gave better results than instrumentation and fusion. Our results suggest that surgical correction should be done within 3 years following growth spurt, i.e. 14 to 16 years of age. PMID- 12118125 TI - Interlaminar discectomy and selective foraminotomy in lumbar disc herniation. AB - Our objective was to assess the clinical outcome of interlaminar discectomy in patients suffering with degenerated lumbar disc lesions. We made a prospective study of 50 consecutive patients who underwent limited lumbar discectomy. The clinico-radiological parameters, type of surgery performed and the post-operative follow up were assessed. We found that interlaminar discectomy without laminotomy was adequate in 33 cases (66%). Most patients requiring laminotomy (17 cases 34%) for discectomy had associated lumbar canal stenosis, herniation at proximal levels (L3-4) and/or sacralization of L5 vertebra. Selective foraminotomy in addition to discectomy was performed in 28 cases (56%). The post-operative results were good in 43 (86%fair in 6 (12%) and poor subjective in 1 case (2%). No patient was classified as poor objective. In conclusion, interlaminar discectomy without laminotomy is a safe, effective and reliable surgical technique for treating properly selected patients with herniated lumbar disc at L4-5 and L5-S1 levels. PMID- 12118126 TI - Enhanced apoptosis of soft tissue sarcoma cells with chemotherapy: A potential new approach using TRAIL. AB - Soft tissue sarcomas are less responsive to conventional chemotherapy when compared to bone sarcomas. We investigated the possibility of enhancing the efficacy of chemotherapy by utilising the recently identified cytokine, tumour necrosis factor-related apoptosis-inducing ligand (TRAIL/Apo2L) in combination with standard chemotherapeutic agents. Fresh human soft tissue sarcomas (rhabdomyosarcoma, fibrosarcoma, malignant fibrous histiocytoma) were obtained at biopsy and dispersed tumour cells were incubated in cell culture with standard cytotoxic agents, either as single agents or in combination with TRAIL. The chemotherapeutic agents were, at best, moderately effective, in terms of induction of cellular apoptosis, although the fibrosarcoma was completely unresponsive to all single agents. TRAIL alone had no effect on any sarcoma cell culture. In contrast, the addition of TRAIL and drug together produced a significant increase in sarcoma cell apoptosis, with TRAIL and doxorubicin the most effective combination. PMID- 12118127 TI - Polyethylene particles from a hip simulator cause (45)Ca release from cultured bone. AB - Periprosthetic osteolysis is a dominant factor in the success or failure of total hip prostheses. Polyethylene wear debris has been implicated in the process of bone resorption and subsequent implant loosening. The present study is the first to examine the effect of ultra high molecular weight polyethylene (UHMWPE) wear debris produced by a hip simulator on calvarial bone resorption in vitro. (45)Ca release was measured in cultured mouse calvarial bone samples. Although short term exposure to UHMWPE particles (2 h) decreased (45)Ca release, longer-term exposure for 1-2 days increased release in a dose-dependent manner. After one-day exposure to 7.5 x 10(6) particles per mL, 18% more (45)Ca was released from cultured calvarial bone than from control samples. It was concluded that UHMWPE wear particles either directly or indirectly stimulated osteoclasts to activate bone resorption. Polyethylene wear debris contributes to the osteolytic process at the bone-implant interface. PMID- 12118128 TI - Knee stability after repair of isolated midsubstance tears of the posterior cruciate ligament. AB - This study was made to compare the results after operative versus non-operative management for tears in the substance of posterior cruciate ligament. The subjects were 40 patients, half of whom were managed operatively and the other half non-operatively. End-to-end suture was performed on the operated group. The procedure was followed by cast immobilization for 4 weeks. Non-operative management consisted of immobilization in a cast for 4 weeks after arthroscopy. The average follow-up periods were 6 years 7 months and 4 years 3 months. The operated knees were significantly more stable than those of the non-operated group on stress radiographs (p<0.0001), but not to the extent of an age-matched control group (p<0.0001). The knee rating scores did not indicate any improvement of the functional outcome in the operated group. Surgery resulted in better posterior laxity than non-operative management; the achieved stability, however, was clearly less than in controls. PMID- 12118129 TI - A study of vascular injuries in pediatric supracondylar humeral fractures. AB - 194 children with supracondylar fractures of the humerus were reviewed. Of the 49 children with Gartland grade III displacement, signs of vascular compromise were clinically suspected in 5 cases. Immediate open reduction, internal fixation and exploration were performed. Four children had a satisfactory outcome. One child required amputation. A careful clinical evaluation for vascular injury and an aggressive surgical approach is suggested, when indicated. PMID- 12118130 TI - Septic arthritis of the hip joint in cervical cancer patients after radiotherapy: Three case reports. AB - Cervical cancer patients may experience hip problems related to the cancer itself or therapeutic management for the cancer. Septic arthritis should be one of the possibilities but there have been no reports on this. Here we present three patients who developed hip problems more than two years after radiotherapy with or without a radical operation. One patient was managed as septic arthritis because of significant inflammatory signs around the affected hip joint even though the causative organism was not confirmed. Succeeding total hip arthroplasty functioned well and had no recurrence of infection. The hip problems of the other two patients were diagnosed as radiation osteonecrosis of the femoral head initially. However, Bacteroides fragilis infection was found several months after total hip arthroplasties. Radiotherapy to the pelvis may damage the hip joint and compromise host-defense mechanisms of the pelvic region. Both factors may increase the possibility of infection of hip joints. Further clinical evidence is needed to understand whether subacute or chronic anaerobic infection could also be one of the causes leading to progressive destruction of the femoral head. PMID- 12118131 TI - The Hoffa fracture: Three case reports. AB - Unicondylar fractures of the lower end of the femur are uncommon injuries that usually occur in the saggital plane 2. Coronal (tangential) plane fractures, first described by Hoffa in 1904, are unusual.(1,2, 3,4) We report our experience with three cases and the mechanism of injury is discussed. Open reduction and internal fixation is mandatory for good long-term results. PMID- 12118132 TI - Acromial hyperplasia, the sequel of deltoid contracture: A case report. AB - The major complications of deltoid contracture frequently encountered are abduction contracture, winging of the scapula, humeral head flattening and shoulder dislocation (Bhattacharyya 1966; Chatterjee and Gupta 1983). To our knowledge acromium hyperplasia has not been a reported complication of deltoid contracture in the English-language literature. We encountered a patient who had bilateral acromium hyperplasia that appeared to be secondary to deltoid muscle contracture due to large volumes of fluid injected intramuscularly as a child. PMID- 12118134 TI - Intratendinous ganglion in the extensor tendon of a finger: A case report. PMID- 12118133 TI - Intradural spinal metastasis from renal cell carcinoma: A case report. AB - Intradural spinal metastasis is rare. This is the third case ever reported on the finding of intradural spinal metastasis from a renal cell carcinoma that had been removed surgically. The patient had a history of epidural metastasis for which excision and anterior stabilization were done 3 years before the new presentation with cauda equina lesion. Seeding from the involved osseous structure to the cerebrospinal fluid through the dura was believed to be the course that tumour had taken to reach the intradural space. PMID- 12118135 TI - Peroneal compartment syndrome of non-traumatic origin: A case report. AB - A patient with acute peroneal compartment syndrome is presented. This case is unusual because the pathology was localised to the peroneal compartment only and because trauma was not an aetiological factor. Acute and chronic compartment syndromes are discussed and differentiated, and the importance of a high index of suspicion in all cases is emphasised. PMID- 12118136 TI - Qualitative analysis of polyethylene wear in a bipolar femoral prosthesis: A case report. AB - We researched the qualitative changes in the polyethylene of a bipolar head retrieved at revision occurring in vivo using optical microscopy and Fourier transform infrared spectroscopy. The bearing surface of the outer head was smooth with no definable scratches delamination. However, the evidence of delamination of the beveled throat region and cracking at the base of the leaf was noticed. The degree of oxidation was different in each area of the polyethylene in the bearing component and that of the rim was shown to be higher than that of the bearing surface. Comparing the rate of ketone and ester, ketone binding was frequently found around the prosthetic rim rather that at the bearing surface. However, the ratio of ester at the surface was higher than that at the deep area of the same rim. We considered these results showed local environmental effects in vivo play a significant role in dictating the response. PMID- 12118137 TI - Review Article: The future of knee ligament surgery. PMID- 12118138 TI - Review Article: The rationale for posterior cruciate substituting total knee arthroplasty. PMID- 12118139 TI - Bioabsorbable laminated membranes for guided bone regeneration. PMID- 12118140 TI - Biomaterials in orthopaedic surgery and traumatology. PMID- 12118141 TI - Critical issues in bioartificial liver development. AB - End-stage liver disease accounts for over 30,000 deaths annually in the United States. Orthotopic liver transplantation is the only clinically proven treatment for patients with end-stage liver failure. A limitation of this therapy is a shortage of donor organs available. This donor organ shortage is exacerbated by the fact that the number of patients listed for transplantation has continued to increase. As a result, there has been a continuing increase in the number of patients who die waiting for a donor liver. Extracorporeal bioartificial liver devices consisting of viable hepatocytes have the potential to provide temporary support for patients with fulminant hepatic failure, thereby serving as a "bridge" to transplantation. In some patients, this temporary support would allow the native liver to regenerate function, eliminating the need for transplantation and the resulting life-long immunosuppressive therapy, all of which translates into a cost savings to the health care system. Although the bioartificial liver device is a promising technology for the treatment of liver failure, significant technical challenges remain in order to develop systems with sufficient processing capacity and of manageable size. An overview of the critical issues in the development of bioartificial liver devices is discussed. PMID- 12118142 TI - Fundamentals of tissue engineering: carrier materials and an application. PMID- 12118143 TI - Fundamentals of tissue engineering: tissues and applications. PMID- 12118144 TI - Novel polypeptide-comprising biopolymer systems. AB - Covalent bioconjugation between anionic polyelecrolytes and polypeptide antigens chemically synthesized by solid-phase chemistry, were studied in hydrated reversed micelle systems. The epitops of Foot-and-Mouse disease virus VP1 protein (40--60 and 135--160 residues) were used as polypeptide antigens. The polypeptide comprising Biopolymer Systems were obtained by two methods: 1) Inclusion of peptides into electrostatic polyelectrolyte complexes of polycations with proteins, 2) Inclusion of peptides into Cu(+2)-induced polyelectrolyte complexes of polyanions with similarly (anionic) charged proteins. The immunogenetic properties of polymer-peptide conjugates and peptide-comprising Biopolymer Systems were investigated and the specificity of antibodies produced was analyzed. Polymer-Peptide conjugates, as well as peptide-comprising Biopolymer Systems appeared to possess a high peptide specific immunogenecity even without the addition of traditional adjuvants. It was found that PE-BSA.FMD.Ag conjugates conferred effective immunoprotection against Foot-and-Mouth disease virus. PMID- 12118146 TI - A new method for the estimation for the absorption time of bioabsorbable polymers in the body. AB - A new method was developed to estimate the in vivo time of absorption of alpha hydroxy polyester type polymers. This method is based on the pH Stat titration of the polymer as a function of time which can be related to the absorption time of the polymer in the body. PMID- 12118145 TI - Current approaches to analgesic drug delivery for chronic pain. AB - A growing number of governmental and professional guidelines internationally have supported aggressive treatment of acute (e.g., postsurgical), cancer, and noncancer pain. The basis for such support is awareness that aggressive control of acute pain reduces postoperative complications and speeds recovery. Chronic noncancer pain (e.g., back pain, headache...) exacts enormous financial costs in each developed nation. Patients' quality of life and possibly even duration of survival as well as associated caregiver burden are enhanced by adequate pain control in patients with chronic pain due to cancer and noncancer causes. Because humanitarian benefits of pain control are supplemented by economic savings, a variety of techniques have been introduced to improve the temporal or spatial profiles of analgesic drug delivery. This brief survey describes the physiological basis for considering pain itself as a disease, the principal drugs and delivery approaches for treatment of severe pain, and the future of "combination analgesic chemotherapy". PMID- 12118147 TI - Prehension and perception of size in left visual neglect. AB - Right hemisphere damaged patients with and without left visual neglect, and age matched controls had objects of various sizes presented within left or right body hemispace. Subjects were asked to estimate the objects' sizes or to reach out and grasp them, in order to assess visual size processing in perceptual-experiential and action-based contexts respectively. No impairments of size processing were detected in the prehension performance of the neglect patients but a generalised slowing of movement was observed, associated with an extended deceleration phase. Additionally both patient groups reached maximum grip aperture relatively later in the movement than did controls. For the estimation task it was predicted that the left visual neglect group would systematically underestimate the sizes of objects presented within left hemispace but no such abnormalities were observed. Possible reasons for this unexpected null finding are discussed. PMID- 12118148 TI - Is grasping impaired in hemispatial neglect? AB - Patients with right unilateral cerebral stroke, four of which showed acute hemispatial neglect, and healthy aged-matched controls were tested for their ability to grasp objects located in either right or left space at near or far distances. Reaches were performed either in free vision or without visual feedback from the hand or target object. It was found that the patient group showed normal grasp kinematics with respect to maximum grip aperture, grip orientation, and the time taken to reach the maximum grip aperture. Analysis of hand path curvature showed that control subjects produced straighter right hand reaches when vision was available compared to when it was not. The right hemisphere lesioned patients, however, showed similar levels of curvature in each of these conditions. No behavioural differences, though, could be found between right hemisphere lesioned patients with or without hemispatial neglect on either grasp parameters, path deviation or temporal kinematics. PMID- 12118149 TI - Deficits of motor intention following parietal lesions. AB - Patients with lesions to the right parietal lobe were tested on their ability to reach to targets, or to respond verbally to targets. The targets occurred at the same two spatial locations--to the left and right of the patient--with the task being cued by the color of the target. Patients were able to perform both tasks separately rapidly and without error. However, when the two tasks were interleaved, they had difficulty making a response in the left (contralesional) field when this was different to a response that they had just made. These results suggest that lesions to the parietal cortex may cause a deficit in the coding for motor intention, as well as attention in the contralesional field. PMID- 12118150 TI - Impaired initiation but not execution of contralesional saccades in hemispatial neglect. AB - Patients with unilateral neglect are impaired at making saccades to contralesional targets. Whether this problem arises from a deficit in perception, in planning the saccade or in executing the eye movement or some combination thereof remains unclear. We measured several variables related to the initiation and execution of saccades in an experiment which crossed two factors: target side (left, right) and direction of saccade (leftwards, rightwards). Relative to control subjects, patients with left-sided neglect were impaired in planning but not executing the contralesional saccade; while the latency to move their eyes following the onset of the target was increased, the duration and velocity to reach the target were normal. In addition, there were also no directional differences for saccades that were hypometric or inaccurate in the patients, further ruling out an execution impairment. Interestingly, this directional initiation deficit was exaggerated for leftward saccades to left targets, compared with all other conditions. We suggest that the disadvantage for contralesional saccades in neglect patients is attributable to a deficit not only in perceiving contralateral targets but also in planning leftward saccades. Once the saccade is initiated, however, execution apparently proceeds unimpaired. PMID- 12118151 TI - Lexical processes and eye movements in neglect dyslexia. AB - Neglect dyslexia is a disturbance in the allocation of spatial attention over a letter string following unilateral brain damage. Patients with this condition may fail to read letters on the contralesional side of an orthographic string. In some of these cases, reading is better with words than with non-words. This word superiority effect has received a variety of explanations that differ, among other things, with regard to the spatial distribution of attention across the letter string during reading. The primary goal of the present study was to explore the interaction between attention and lexical processes by recording eye movements in a patient (F.C.) with severe left neglect dyslexia who was required to read isolated word and non-word stimuli of various length. F.C.'s ocular exploration of orthographic stimuli was highly sensitive to the lexical status of the letter string. We found that: (1) the location to which F.C. directed his initial saccade (obtained approximately 230 ms post-stimulus onset) differed between word and non-word stimuli; (2) the patient spent a greater amount of time fixating the contralesional side of word than non-word strings. Moreover, we also found that F.C. failed to identify the left letters of a string despite having fixated them, thus showing a clear dissociation between eye movement responses and conscious access to orthographic stimuli. Our data suggest the existence of multiple interactions between lexical, attentional and eye movement systems that occur from very initial stages of visual word recognition. PMID- 12118152 TI - Knowing what you need but not what you want: affordances and action-defined templates in neglect. AB - We examined search for target objects in a patient, MP, showing symptoms of left unilateral neglect. The conditions varied how the target was defined, the numbers of targets and distractors, and whether search was for multiple or single targets. We found that search was substantially improved when the target was defined by a description of its action rather than its name. This advantage for action-defined targets increased with larger display sizes. For both action and name-defined targets, there were also larger effects of the number of distractors when search was for multiple rather than single targets, even when the numbers of distractors were kept constant. However, these effects were increased for name defined targets. The differences between action- and name-defined targets decreased when objects were replaced with words. The data suggest that search could be based on action-defined templates of targets, activated by affordances from objects. The action-defined templates facilitated detection and reduced MP's tendency to re-search displays. PMID- 12118155 TI - Peroxisome proliferator-activated receptors (PPARs) and peroxisomes in rat cortical and cerebellar astrocytes. AB - Astrocytes are the most versatile cells of the neural tissue. Numerous astrocytic functions--such as protection from oxidative damage, catabolism of neuroactive D amino acids acting as neuromodulators, synthesis and catabolism of some lipid molecules, and, possibly, gluconeogenesis--reside in peroxisomes. The expression of several peroxisomal enzymes, particularly those of the acyl-CoA beta-oxidation pathway, is regulated by a class of ligand-activated transcription factors, known as peroxisome proliferator-activated receptors (PPARs), acting on their target genes as heterodimers with the retinoid X receptors (RXRs). In this work, primary and secondary cultures of astrocytes from the cerebral cortices and cerebella of neonatal rats (2 and 7 days of postnatal age) were utilized to investigate the expression of peroxisomal enzymes, PPAR and RXR isotypes (alpha, beta and gamma), by both biochemical and immunological methods. The results obtained demonstrate that astrocytes in vitro express peroxisomal enzymes, PPARs, and RXRs and that differences dependent on brain area, animal age, and culture time are reminiscent of the in vivo situation. Therefore, primary cultures of astrocytes and, particularly, high purified subcultures may constitute a useful model for further studies aimed to gain further insights into the roles of peroxisomes and PPARs related to lipid and glucose metabolism in these cells. PMID- 12118154 TI - Increased number of nitric oxide synthase immunoreactive Purkinje cells and dentate nucleus neurons in schizophrenia. AB - There is growing interest in the cerebellum as a site of neuropathological changes in schizophrenia. Reports showing that schizophrenics have higher nitric oxide synthase (NOS) activity and MAPKinase levels in the vermis, point to possible aberrations in the cerebellar signal transduction of schizophrenics. It has been speculated that Ca(2+)-dependent extracellular to intracellular signal transduction may be disrupted in the cerebellum of schizophrenics. We decided to test this hypothesis by studying the nitrergic system and markers of the Ca(2+) triggered signal cascade in the cerebellum of schizophrenics, depressives and controls. The cellular distribution of two calcium sensor proteins (VILIP-1 and VILIP-3) and of neuronal NOS immunoreactivity was studied morphometrically in the flocculonodulus, the inferior vermis and the dentate nucleus of 9 schizophrenics, 7 depressive patients and 9 matched controls. In comparison to controls and depressed patients there were fewer Nissl-stained neurons in the dentate nucleus of schizophrenics. The number of NOS-expressing Purkinje neurons was however strongly increased. In the flocculonodulus and the vermis no differences between the groups were found with regard to the density of Nissl-stained Purkinje cells. The number of NOS-expressing Purkinje neurons was increased in schizophrenics, however. No differences between schizophrenics, depressives and controls were found in the number of VILIP-1 immunoreactive dentate nucleus neurons and VILIP-3 immunoreactive vermal and flocculonodular Purkinje cells. Our data provide further histochemical evidence in favor of structural abnormalities in discrete cerebellar regions of schizophrenics. They confirm and extend earlier reports of increased cerebellar NOS immunoreactivity in schizophrenia and point to possible neurodevelopmental disturbances. Our failure to show an altered expression of two calcium sensor proteins possibly points to a less important role of calcium signaling in cerebellar pathology of the disease. PMID- 12118156 TI - Herpes simplex virus in the vestibular ganglion and the geniculate ganglion-role of loose myelin. AB - This study presents the first direct evidence for herpes simplex virus type 1 (HSV-1) infection in the neurons of the vestibular ganglion. Although many investigators have reported electron microscopic evidence of HSV-1 infection in sensory ganglia, HSV-1 infection in the vestibular ganglion has not been described. Vestibular ganglion neurons have a unique structure, with a loose myelin sheath instead of the satellite cell sheath that is seen in other ganglia. This loose myelin is slightly different from compact myelin which is known as too tight for HSV-1 to penetrate. The role of loose myelin in terms of HSV-1 infection is completely unknown. Therefore, in an attempt to evaluate the role of loose myelin in HSV-1 infection, we looked for HSV-1 particles, or any effects mediated by HSV-1, in the vestibular ganglion as compared with the geniculate ganglion. At the light microscopic level, some neurons with vacuolar changes were observed, mainly in the distal portion of the vestibular ganglion where the communicating branch from the geniculate ganglion enters. At the electron microscopic level, vacuoles, dilated rough endoplasmic reticulum and Golgi vesicles occupied by virus were observed in both ganglia neurons. In contrast, viral infections in Schwann and satellite cells were observed only in the geniculate ganglion, but not in the vestibular ganglion. These results suggest that loose myelin is an important barrier to HSV-1 infection, and it must play an important role in the prevention of viral spread from infected neurons to other cells. PMID- 12118157 TI - Lamination of spinal cells projecting to the zona incerta of rats. AB - In this study, the lamination patterns of spinal cells projecting to the zona incerta (ZI), intralaminar nuclei and ventral posterior nucleus of the thalamus have been explored. Injections of cholera toxin subunit B or latex beads were made into the ZI, intralaminar and ventral posterior nuclei of Sprague Dawley rats. The brain and spinal cord were then aldehyde fixed and processed using standard methods. Our results show two major findings. First, after injections into the ZI, there is a distinct pattern of lamination of labelled cells in the spinal cord, a pattern that changes across the different levels. At cervical levels, labelled cells are located within the medial region of the deep dorsal horn, while at lumbar and sacral levels, they are found in the intermediate grey matter. These results are similar to those seen after injections into the intralaminar or ventral posterior nuclei, except that in the latter cases, more labelled cells are located in the superficial laminae of the dorsal horn, particularly from the ventral posterior nucleus. Second, the ZI is not associated uniformly with all spinal levels; labelling is heaviest at cervical and lightest at thoracic levels. From each thalamic injection site, labelling is noted on both sides of the spinal cord, with a clear contralateral predominance. In conclusion, the results indicate that the ZI receives a distinct set of spinal projections principally from the cervical level. The particular pattern of lamination of spinal cells projecting to the ZI suggests that the type of information relayed is from deep somatic and/or visceral structures, and probably nociceptive in nature. PMID- 12118158 TI - Immunogold study of interendothelial junction-associated and glucose transporter proteins during postnatal maturation of the mouse blood-brain barrier. AB - The distribution of glucose transporter (GLUT-1) and of interendothelial junction associated proteins--zonula occludens protein (ZO-1), occludin, and beta-catenin- was studied using quantitative immunogold procedure. Lowicryl K4M-embedded samples of the cerebral cortex of 1-, 7-, and 14-day-, and 6-week-old (young adult) mice were used. Ultrathin sections were exposed to specific rabbit polyclonal antibodies followed by colloidal gold-labelled secondary antibodies. We found that the density of immunosignals for GLUT-1 in both luminal and abluminal plasma membranes of the endothelial cells, and those closely related to the interendothelial junctions was low in blood microvessels from newborn mice, dropped slightly at the 7th day, and increased through the 14th day to the level of mature blood-brain barrier (BBB) observed in 6-week-old mice. The expression of ZO-1 was high in newborn mice and increased at the 7th day to the level similar to that found in 14-day- and 6-week-old mice. The expression of occludin was less intense than that of ZO-1 and increased from birth, reaching at the 14th day the level typical for mature BBB found in young-adult animals. The immunosignals for occludin were sparsely distributed inside the junctional clefts. Such a distribution indicates that the tight junctional characteristics are limited to a few short segments of the entire interendothelial cleft. The density of immunosignals for beta-catenin was lowest, and it had the tendency to a gradual, although inconsiderable, drop in the time course of BBB maturation. These findings suggest that the relatively high concentration of GLUT-1 in the interendothelial junctions results from the participation of abluminal plasma membranes of adjacent endothelial cells in the formation of the junctional complexes. The interendothelial junctions of newborn mice are equipped already with the main components of the tight junctions, and the concentration of these components (ZO-1, occludin) reaches the level of the mature BBB at the 14th day of postnatal life. PMID- 12118159 TI - Brain phagocytes may empty tissue debris into capillaries. AB - The possibility that brain phagocytes may empty remnants of degenerated neurons into capillaries has been studied in frogs. Degeneration of nerve fibers was brought about by transectioning the optic tract, the tectothalamic and tectoisthmic tracts, the postoptic commissure or the radial nerve. To help identification of phagocytozed degenerated neuronal elements, the transected fibers were filled either with horseradish peroxidase (HRP) or cobaltous-lysine complex. The survival times were 3, 4, 7, 27, 47 and 70 days after the application of the markers. The HRP-labeled structures were identified in 60 microm thick sections using diaminobenzidine as chromogen, while cobalt was precipitated in the form of cobaltous sulfide. Small pieces of these sections were further processed for electron microscopy. In each area of the brain and spinal cord investigated, microglial cells and astrocytic processes containing fragments of degenerated neuronal elements could be seen close to capillaries. In some cases a microglial or astrocytic process pierced the capillary basal lamina and seemingly delivered inclusion bodies into the cytoplasm of capillary endothelial cells and pericytes. In the inclusion bodies, which were usually large vesicles, fragments of HRP or cobalt-labeled or unlabeled membranes with a foamy appearance, or condensed myelin lamellae could be observed. These vesicles protruded the luminal membrane of the endothelial cell that was disrupted in some cases suggesting that the content of the inclusion body was discharged into the lumen of the capillary. These results give support to Penfield's hypothesis (1925) that glial cells may empty phagocytozed materials into capillaries. PMID- 12118161 TI - Information generated by the application of molecular genetic techniques. General introduction. PMID- 12118162 TI - Neurochemical anatomy of the zebrafish retina as determined by immunocytochemistry. AB - The zebrafish retina is rapidly becoming a major preparation for the study of molecular genetic mechanisms underlying neural development and visual behavior. Studies utilizing retinal mutants would benefit by the availability of a data base on the distribution of neurotransmitter systems in the wild-type fish. To this end, the neurochemical anatomy of the zebrafish retina was surveyed by light microscopic immunocytochemistry. An extensive series of 60 separate antibodies were used to describe the distribution of major transmitter systems and a variety of neuron-associated membrane channels and proteins. These include markers (i.e., antibodies against enzymes, receptors, transporters) for transmitters: GABA, glycine, glutamate, biogenic amines, acetylcholine, cannabinoids and neuropeptides; as well as a sample of voltage-gated channels and synapse associated membrane proteins. Discussion of the comparative localization of these antibodies is restricted to other teleost fishes, particularly goldfish. Overall, there was great similarity in the distribution of the various markers, as might be expected. However, there were some notable differences, including several antibodies that did not label zebrafish at all, even though goldfish retinas that were processed in parallel, labeled beautifully. This survey is extensive, but not exhaustive, and hopefully will serve as a valuable resource for future studies of the zebrafish retina. PMID- 12118164 TI - Anterior uveitis: current concepts of pathogenesis and interactions with the spondyloarthropathies. AB - Anterior uveitis describes inflammation that involves the iris or ciliary body. Anterior uveitis may be part of a systemic illness such as a spondyloarthropathy. It may also arise from an infection such as herpes simplex; be part of an ocular syndrome, such as Fuchs' heterochromic iridocyclitis; be part of trauma, as in cataract surgery; or result from an idiopathic eye disease with a presumed immune pathogenesis. During 2001, progress has been made understanding uveitis in general, as well as specifically, in association with spondyloarthropathy. Here, we review recent insights into anterior uveitis with regard to clinical presentation, immune mechanisms, genetics, and anti-tumor necrosis factor therapy. PMID- 12118163 TI - A molecular phenotype atlas of the zebrafish retina. AB - The rasborine cyprinid Danio rerio (the zebrafish) has become a popular model of retinal function and development. Its value depends, in part, on validation of homologies with retinal cell populations of cyprinine cyprinids. This atlas provides raw and interpreted molecular phenotype data derived from computationally classified sets of small molecule signals from different cell types in the zebrafish retina: L-alanine, L-aspartate, L-glutamine, L-glutamate, glutathione, glycine, taurine and gamma-aminobutyrate. This basis set yields an 8 dimensional signature for every retinal cell and formally establishes molecular signature homologies with retinal neurons, glia, epithelia and endothelia of other cyprinids. Zebrafish photoreceptor classes have been characterized previously: we now show their metabolic profiles to be identical to those of the corresponding photoreceptors in goldfish. The inner nuclear layer is partitioned into precise horizontal, bipolar and amacrine cell layers. The horizontal cell layer contains at least three and perhaps all four known classes of cyprinine horizontal cells. Homologues of cyprinid glutamatergic ON-center and OFF-center mixed rod-cone bipolar cells are present and it appears likely that all five classes are present in zebrafish. The cone bipolar cells defy simple analysis but comprise the largest fraction of bipolar cells, as in all cyprinids. Signature analysis reveals six molecular phenotypes in the bipolar cell cohort: most are superclasses. The amacrine cell layer is composed of approximately equal 64% GABA+ and 35% glycine+ amacrine cells, with the remainder being sparse dopaminergic interplexiform cells and other rare unidentified neurons. These different amacrine cell types are completely distinct in the dark adapted retina, but light adapted retinas display weak leakage of GABA signals into many glycinergic amacrine cells, suggesting widespread heterocellular coupling. The composition of the zebrafish ganglion cell layer is metabolically indistinguishable from that in other cyprinids, and the signatures of glial and non-neuronal cells display strong homologies with those in mammals. As in most vertebrates, zebrafish Muller cells possess a high glutamine, low glutamate signature and contain the dominant pool of glutathione in the neural retina. The retinal pigmented epithelium shows a general mammalian signature but also has exceptional glutathione content (5-10 mM), perhaps required by the unusually high oxygen tensions of teleost retinas. The optic nerve and the marginal zone of the retina reveal characteristic metabolic specializations. The marginal zone is strongly laminated and its nascent neurons display their characteristic signatures before taking their place in the retina proper. PMID- 12118165 TI - Immune linkages between inflammatory bowel disease and spondyloarthropathies. AB - Gut involvement is a prominent feature of spondyloarthropathy (SpA). Analysis of immune alterations of the gut in SpA have shown two distinct aspects. On the one hand, gut inflammation in SpA seems closely related with gut inflammation seen in Crohn disease. On the other hand, gut inflammation in SpA is associated with peripheral joint inflammation. Recent studies have provided new insights into this gut-synovium axis. First, there is little new evidence to support the concept of viable microbial pathogens recirculating to the joint. In contrast, it seems likely that both bacterial antigens and reactive T cell clones home to the joint, and that adhesion molecules such as the beta7 integrins and VAP1 play an important role in this process. Second, there is increasing evidence that the different disease localizations in SpA are characterized by alterations of the innate immune system, which contribute to a breakdown of the immune tolerance and the creation of an inflammation-prone environment. Mediators of the innate immune system, such as scavenger receptors, interleukin-10 (IL-10), and tumor necrosis factor alpha (TNF-alpha), may therefore be interesting targets for therapeutic intervention, as illustrated by the effect of TNF-alpha blockade in SpA. PMID- 12118166 TI - Immune features of seronegative and seropositive arthritis in early synovitis studies. AB - Synovitis of recent onset is a challenging problem, both from a diagnostic and a mechanistic point of view. The role of the immune system in mediating the systemic and synovial inflammatory response remains an area of active investigation. Studies in early synovitis cohorts have confirmed the relatively specific association of rheumatoid factor positive polyarthritis with a number of autoantibodies, particularly anticyclical citrullinated peptide (CCP) antibodies, antifilaggarin antibodies (AFA), and anti-Sa antibodies. Immunopathologic studies of synovial tissue samples from patients with early synovitis have generally suggested quantitative rather than qualitative differences between various forms of synovitis. In particular, Th1 cytokines appear to predominate in rheumatoid arthritis and psoriatic synovitis, while Th2 cytokines are more often detectable in the synovium of reactive arthritis patients. This latter observation is consistent with an immune response profile that favors persistence of intracellular organisms. PMID- 12118167 TI - Genetic aspects of susceptibility, severity, and clinical expression in ankylosing spondylitis. AB - While twin studies have previously demonstrated high heritability of susceptibility to ankylosing spondylitis (AS), it is only recently that the involvement of genetic factors in determining the severity of the disease has been demonstrated. The genes involved in determining the rate of ankylosis in AS are likely to be different from those involved in the underlying immunologic events, and represent important potential targets for treatment of AS. This article will describe the progress that has been made in the genetic epidemiology of AS, and in identifying the genes involved. PMID- 12118168 TI - Current concepts in psoriatic arthritis. AB - This review summarizes articles written about psoriatic arthritis in the past year. It concentrates on clinical and epidemiologic issues, pathogenesis and treatment, and updates the reader regarding new concepts in psoriatic arthritis. PMID- 12118169 TI - HLA-B27 and pathogenesis of spondyloarthropathies. AB - Although the influence of HLA-B27 on the development of spondyloarthropathies is undisputed, its role in pathogenesis remains unclear. New ideas have focused on abnormal characteristics of HLA-B27 resulting from aberrant folding, disulfide bond formation, or both, rather than a predilection for selecting arthritogenic peptides. This reflects, in part, unanswered questions about whether immunologic recognition of HLA-B27 is required for disease. Recent studies suggest that CD4+ T cells, immunomodulatory killer cell Ig receptors, and Ig-like transcript receptors may recognize aberrant forms of HLA-B27. Other reports suggest that HLA B27 expression can alter cytokine production from monocytes and T cells-effects that appear unrelated to antigen presentation. Novel bioinformatics approaches have led to the identification of HLA-B27-restricted pathogen-derived peptides and may prove useful in determining whether HLA-B27 presents arthritogenic peptides. Elucidating the role of HLA-B27 in the pathogenesis of these conditions will require an integration of information from animal models, genome-wide screens for susceptibility alleles, and translational studies using human samples. PMID- 12118171 TI - Lyme arthritis and post-Lyme disease syndrome. AB - In the United States, intermittent or chronic mono- or oligoarthritis, particularly affecting the knee, is the most common manifestation of late Lyme disease (LD). Lyme arthritis (LA) can usually be prevented by early treatment of acute LD. However, the erythema migrans rash may go undetected in children and in the dark skin of African Americans, leading to delayed treatment and a relatively increased incidence in LA. Virtually all untreated patients with LA have high levels of serum immunoglobulin G antibodies, and sometimes low levels of immunoglobulin M antibodies, to Borrelia burgdorferi (Bb) by ELISA and Western blot. These responses may persist for many years after antibiotic treatment, and therefore, serologic results do not accurately distinguish between active or past infection. Most patients with LA respond well to standard courses of antibiotic treatment, but a small percentage have persistent knee synovitis, in some cases possibly related to the triggering of intrasynovial autoimmunity. Other patients develop a syndrome of diffuse arthralgia, myalgia, fatigue, and subjective cognitive difficulty during or soon after LD, which persists despite antibiotic treatment. Patients with this post-treatment, post-LD syndrome were recently studied in a placebo-controlled double-blind antibiotic trial. There was no evidence of Borrelial infection in these patients by culture or detection of Bb DNA in blood or spinal fluid. Furthermore, there was no difference in responsiveness of these patients to a 3-month course of antibiotic compared with placebo treatment. Thus, LA caused by active Bb infection, post-treatment LA with persistent knee synovitis and post-LD syndrome are distinct and distinguishable clinical entities. PMID- 12118170 TI - Innate immunity in host-microbial interactions: beyond B27 in the spondyloarthropathies. AB - The spondyloarthropathies are diseases influenced by genetic predisposition and, to a varying extent, infectious triggers. A causal role for bacterial infections is most clear for reactive arthritis. Recent insights into arthritogenic components of bacteria may set the stage for a better understanding of disease pathogenesis, the role of heat shock proteins in antigen processing and immune activation, and the adjuvant effect of CpG-DNA. Recent developments in the area of innate immunity broaden current concepts of genetically defined factors in host-pathogen interactions. In particular, the biology of toll-like receptors as important elements in the innate immune response to pathogens is being defined. These factors in innate immunity may have important implications for sequelae of infections, such as reactive arthritis. PMID- 12118172 TI - Development of autoimmunity in Lyme arthritis. AB - Treatment-resistant Lyme arthritis (TRLA) develops in 10% of Lyme arthritis patients and is characterized by continuous joint inflammation that does not resolve with antibiotic therapy. TRLA is associated with HLA-DRB1*0401 and related alleles, as well as with an immune response to the Borrelia burgdorferi (Bb) outer surface protein A (OspA). The immunodominant epitope of OspA in the context of HLA-DRB1*0401 corresponds to amino acids 165-173 (OspA165-173). The human Lymphocyte Function Antigen-1 (hLFA1alpha) contains a peptide with homology to OspA165-173. Treatment-resistant Lyme arthritis patients' T cells, cloned based on their ability to bind OspA165-173-loaded HLA-DRB1*0401 tetramers, respond to OspA and hLFA1alpha with a different cytokine profile, suggesting that hLFA1alpha acts as a partial agonist with a potential role in the perpetuation of joint inflammation. PMID- 12118173 TI - Borrelia burgdorferi and its tropisms for adhesion molecules in the joint. AB - Borrelia burgdorferi, the spirochete that causes Lyme disease, has evolved elegant strategies for interacting with its mammalian hosts. Among them are several distinct mechanisms of adhesion to cells and extracellular matrix components. The mammalian receptors for B. burgdorferi that have been most thoroughly studied, and for which candidate bacterial ligands have been identified, are decorin, fibronectin, glycosaminoglycans, and beta3-chain integrins. This diversity of adhesion mechanisms allows B. burgdorferi to infect multiple tissues, including the synovial tissues of the joints. PMID- 12118174 TI - Host-pathogen interactions and the pathogenesis of murine Lyme disease. AB - Lyme disease results from persistent infection with the spirochete Borrelia burgdorferi. A combination of bacterial factors and host factors contributes to the development of inflammatory disease. Studies from the past year have provided insight into both sides of this host-pathogen interplay. We now have a better appreciation of the bacterial genes and products that are involved in pathology and the components of the host response that participate in disease development. PMID- 12118176 TI - Pathogenesis of bone erosions in rheumatoid arthritis. AB - Focal marginal joint erosions represent the radiographic hallmark of rheumatoid arthritis (RA). These bone changes are characteristically localized to the joint margins, but in addition, regions of focal bone resorption can be detected in the subchondral bone adjacent to the bone marrow space into which the synovial inflammatory tissues have extended. Because progressive destruction of the periarticular bone contributes significantly to joint dysfunction and disability in patients with RA, there is considerable interest in developing a better understanding of the pathologic mechanisms involved in this process and in developing therapies that can arrest these events. Previous analysis of joint tissues from patients with RA have provided morphologic evidence that osteoclasts are the cell types that mediate the focal bone resorption associated with the rheumatoid synovial lesion. Additional recent data from animal models have helped to further implicate these cells in the pathogenesis of focal bone erosions. Furthermore, analysis of RA synovium and joint tissues from animal models of inflammatory arthritis, as well as cell and tissues culture studies, have helped to define the cytokines and inflammatory mediators that are involved in the recruitment and activation of bone resorbing cells associated with focal bone erosions. These findings provide a rational framework for developing targeted therapies that can specifically inhibit or slow the progressive focal bone destruction associated with the rheumatoid synovial lesion. PMID- 12118177 TI - Recent developments in the epidemiology of osteoporosis. AB - Osteoporosis is a widespread condition, often unrecognised in clinical practice, which may have devastating health consequences through its association with fragility fractures. Osteoporotic fractures represent an enormous public health burden. The total number of fractures, and hence the cost to society, will increase dramatically over the next 50 years as a result of demographic changes in the number of elderly people. This article reviews the latest advances in our knowledge of epidemiologic aspects of osteoporosis, including the epidemiology of fragility fracture, the determinants of fracture risk, and genetic, intrauterine, and environmental risk factors for osteoporosis. Novel relationships between breast cancer and bone mineral density will also be discussed. PMID- 12118178 TI - Current controversies in bone densitometry. AB - In the face of increasing use of all types of bone densitometry in the diagnosis and management of osteoporosis, the limitations of the World Health Organization criteria for the diagnosis of osteoporosis based on bone density measurements have become apparent. Controversy has arisen about whether these criteria should be used for the diagnosis of osteoporosis. Using densitometry to monitor changes in bone density as a measure of therapeutic efficacy has been criticized. It has been suggested that changes in bone density are not surrogates for reduction in fracture risk and that regression to the mean invalidates serial testing. There is both truth and fallacy in these controversies. The resolutions are critical to the role of densitometry in clinical practice. PMID- 12118179 TI - Osteoporosis in men: pathophysiology, evaluation, and therapy. AB - Osteoporosis is a serious health problem for men. An advance in our understanding of the pathophysiology and treatment of this disorder has resulted in the possibility of a gender-specific approach to screening, diagnosis, and treatment. Here we review the data on osteoporosis in men, discuss controversies regarding whom to screen, whom to treat, and how to treat. Recent treatment data as they relate to men are reviewed, and a clinical treatment algorithm is presented. PMID- 12118180 TI - Raloxifene: recent information on skeletal and non-skeletal effects. AB - Raloxifene, a selective estrogen receptor modulator, is currently used for both prevention and treatment of postmenopausal osteoporosis. Recent data from the MORE (Multiple Outcomes of Raloxifene Evaluation) trial have evaluated health related quality of life, 1-year clinical vertebral fracture reduction, and the correlation of bone mineral density and biochemical markers of bone turnover with vertebral fracture reduction. In addition, new information on the effects of raloxifene in breast cancer prevention, cardiovascular disease in high-risk women, and uterine disorders is reviewed. PMID- 12118182 TI - Nonmedical management of osteoporosis. AB - Nonmedical therapies are playing an increasing role in the management of osteoporosis and its complications. They serve as useful adjuncts to medical treatment. Several areas of nonmedical options for the management of osteoporosis include bracing, exercise, and vertebroplasty and kyphoplasty. PMID- 12118181 TI - New anabolic therapies in osteoporosis. AB - While antiresorptive drugs have been the cornerstone of osteoporosis therapy, anabolic drugs are an important new advance in the treatment of osteoporosis. By directly stimulating bone formation, anabolic agents might have greater potential than the antiresorptives to increase bone mass and to decrease fractures. It is also possible that the combination of an antiresorptive agent with an anabolic agent could be more potent than either agent alone. Potential anabolic therapies for osteoporosis, including fluoride, growth hormone, insulin-like growth factor I, strontium, and parathyroid hormone, are reviewed here. Of these, parathyroid hormone has clearly emerged as the most promising treatment at this time. PMID- 12118183 TI - Novel therapeutic options for osteoporosis. AB - Osteoporosis remains a significant clinical problem despite effective therapies. Many patients cannot or will not take currently available therapies. For this reason, research continues in search of more effective and more tolerable agents. Arzoxifene and TSE-424 are investigational selective estrogen receptor modulators that have been shown to be effective in animal studies and are now in clinical studies. Tibolone is a tissue-specific steroid that is currently used in Europe for the prevention and treatment of osteoporosis. Multiple studies have shown efficacy in improving bone mineral density, but no fracture studies have been conducted to date. Although studies of the effect of isoflavones on bone mineral density have been encouraging, a large multicenter study in Europe recently showed no effect of isoflavones on fractures. The investigational bisphosphonates ibandronate and zoledronic acid may offer the advantage of less frequent dosing. The newly described agent osteoprotegerin has been shown in early studies to inhibit bone turnover. Finally, the issue of efficacy of statins in bone continues to be debated with no prospective, randomized studies yet to confirm the suggestion of benefit seen in epidemiologic studies. PMID- 12118184 TI - Practice pattern variation in the prevention and treatment of osteoporosis. AB - Prevention and treatment of osteoporosis will be of increasing importance as the mean age of the world population rapidly increases in the coming decades. Nearly half of postmenopausal women not already diagnosed with osteoporosis have significantly low bone mineral density and are at increased risk of fracture, yet this risk is often unrecognized and untreated. Postmenopausal women who have experienced fractures of the hip, vertebra, and wrist and patients using glucocorticoids are at highest risk of fractures but often remain untested and untreated for osteoporosis. Because of the rising societal costs of osteoporosis anticipated in coming years, an improved understanding of the predictors of practice pattern variations and interventions designed to improve underutilization of appropriate care are important to clinicians, health services researchers, and policy makers. PMID- 12118185 TI - Ligands of the peroxisome proliferator-activated receptor-PPAR-a reduce myocardial infarct size. AB - BACKGROUND: This study was designed to investigate the effects of two, chemically distinct activators of PPAR-a (clofibrate and WY14643) in a rat model of acute myocardial infarction. MATERIAL/METHODS: Male Wistar rats were anesthetized with sodium thiopentone (120 mg/kg i.v.) and subjected to regional myocardial ischemia (for 25 min) and reperfusion (for 2 h). Area at risk was determined by injection of Evans Blue dye and infarct size after staining with nitroblue tetrazolium. RESULTS: Pre-treatment of rats with the PPAR-a agonists clofibrate (0.3 mg/kg) caused a significant reduction in myocardial infarct size of 43%. Similarly, pre treatment of rats with the PPAR-a agonists WY14643 (1 mg/kg) caused a significant reduction in myocardial infarct size of 43%. CONCLUSIONS: Taken together, these results show that ligands of PPAR-a reduce the tissue necrosis associated with acute myocardial infarction. We propose that fibrates and other PPAR-a agonists may be useful in conditions associated with ischemia-reperfusion of the heart and other organs. PMID- 12118187 TI - Betulinic acid, a potent inhibitor of eukaryotic topoisomerase I: identification of the inhibitory step, the major functional group responsible and development of more potent derivatives. AB - BACKGROUND: Betulinic acid, a naturally abundant, plant derived, pentacyclic triterpenoid possesses anti-HIV, anti-malarial and anti-inflammatory properties and has recently emerged as a potent anti-tumor compound. This study explores the mode of action of betulinic acid on eukaryotic topoisomerase I and identifies the major functional group responsible along with more potent derivatives. MATERIAL/METHODS: Topoisomerase I relaxation activity was electrophoretically measured by the decreased mobility of the relaxed monomers followed by ethidium bromide staining. DNA cleavage was studied by electrophoretic separation of the nicked monomers from the relaxed and supercoiled monomers in presence of ethidium bromide. In-vivo DNA cleavage was studied in blasted mouse splenocytes by the SDS K+ trapping of 3H-DNA-topoisomerase I-camptothecin ternary complex. RESULTS: Betulinic acid exerts its inhibitory effect by preventing topoisomerase I-DNA interaction as a result of which the 'cleavable complex' is not formed. In consequence, it also acts as an antagonist to camptothecin-mediated cleavage. A series of analogues modified at C-3, C-17 and C-20 positions of betulinic acid were subsequently assayed for inhibition of topoisomerase I catalytic activity. Replacement of the 17-carboxylic group reduces the inhibitory effect and decarboxylation leads to the complete loss of inhibitory effect. CONCLUSIONS: This study is the first detail report of betulinic acid as a very potent inhibitior of eukaryotic topoisomerase I and highlights the necessity of the carboxylic functional group. Dihydro betulinic acid is the most potent (IC50=0.5 mM) pentacyclic triterpenoid to inhibit eukaryotic topoisomerase I till date and can be exploited as a strong candidate for anti-tumor drug designing. PMID- 12118186 TI - Biochemical characterization of the arginine degrading enzymes arginase and arginine deiminase and their effect on nitric oxide production. AB - BACKGROUND: Nitric oxide (NO) is a biomediator believed to be synthesized primarily from extracellular arginine. Various methodologies have been used to inhibit NO synthesis so as to elucidate its physiological and pathophysiological functions. Several investigators have utilized various argin ine degrading enzymes as a means of lowering extracellular arginine. Arginase, most commonly derived from mammalian sources, has been most often used. However, arginase has failed to inhibit NO synthesis. Therefore, a systematic biochemical characterization of arginase and arginine deiminase (ADI) derived from M. Hominus was undertaken. MATERIAL/METHODS: The murine macrophage cell line N-9 was treated with either arginase or arginine deiminase to determine the effect on intracellular and extracellular arginine and nitric oxide production. RESULTS: Arginase was found to have an alkaline pH optima(approximately 9.5) with little enzyme activity at physiological pH. In contrast, the pH optima of ADI was approximately 6.5, retaining >70% of its activity at physiological pH. ADI had more than 1000 fold higher affinity for arginine (Km approximately 30 KM for ADI vs approximately 45 mM for arginase), and was able to lower arginine levels to a much greater extent than arginase. ADI, unlike arginase, was effective in lowering extracellular arginine in tissue culture media and inhibit NO production by the murine macrophage cell line N-9 in response to gamma interferon and LPS stimulation. CONCLUSIONS: These data suggest that ADI may be useful for delineating the role of NO in a variety of biological systems as well as determining the role of extracellular arginine in its synthesis. PMID- 12118188 TI - Biophysical characterization of albumin preparations from blood serum of healthy donors and patients with renal diseases. Part I: spectrofluorometric analysis. AB - BACKGROUND: Serum albumin (SA) is a major proteinacous component of the blood plasma. In the present study we report on the determination and structural characterization of the modified SA in the plasma of glomerulonephritis (GN) patients. Fluorescence analysis of albumin-enriched fractions (AEF) isolated from serum of GN patients leads to the discovery of a proteinacous component with essential blue shift of intrinsic fluorescence. MATERIAL/METHODS: AEF were isolated from serum blood of 35 GN patients, 30 GN patients with chronic renal failure (CRF) corrected by hemodialysis (HD) and 40 healthy donors. Their fluorescence characteristics have been compared using variations in the value of parameter A = I320/I365 that characterizes fluorescence spectrum position and shape. RESULTS: The fluorescence spectra of AEF from GN and GN with CRF patients were blue- and red-shifted, respectively, in comparison with spectra of donors' serum. The value of parameter A was 1.27I0.05 for protein preparations from healthy donors, while this characteristic lies within range of 1.3-2.1 or 0.77 1.29 for GN and GN with CRF patients, respectively. Moreover, for GN patients, the magnitude of spectral parameter A was approaching control values during the treatment of disease. CONCLUSIONS: hSA enriched fractions isolated from GN patients or GN patients with CRF corrected by HD differ by their spectrofluorometric properties from those of healthy donors. This technique allows the detection of renal diseases and the efficiency of their cure. PMID- 12118189 TI - Biophysical characterization of albumin preparations from blood serum of healthy donors and patients with renal diseases. Part II: evidence for the enhancement of the haptoglobin. Level at the pathological conditions. AB - BACKGROUND: Human serum albumin (hSA), the major component in blood plasma, fulfills a fundamental biological role as a universal carrier and reservoir in blood plasma, tissues, and secretions. Because of essential diversity in the bound ligands, hSA was shown to possess significant structural divergence. In the first paper of this series we reported on that albumin-enriched fractions isolated from serum of glomerulonephritis (GN) patients possesses essential blue shift of intrinsic fluorescence. Identification of this component was the major goal of this study. MATERIAL/METHODS: Partially purified SA preparations were isolated from blood plasma of 19 GN patients and 8 healthy donors. These preparations have been analyzed by combination of chromatographic (gel-filtration and hydrophobic sorption) and spectrofluorometric techniques. RESULTS: At least six different albumin enriched fractions were isolated from blood of GN patients. High molecular weight protein fraction with blue shifted intrinsic fluorescence spectrum was segregated. Higher concentration of this fraction may be responsible for higher parameter A values, which is characteristic of protein preparations from GN patients. Amino acid sequence showed that b-haptoglobin is one of the components of this fraction. CONCLUSIONS: Combination of chromatographic methods with monitoring intrinsic fluorescence spectra of albumin-enriched fractions from blood plasma of healthy donors and GN patients represents a useful tool for the description of hSA heterogeneity and for the detection of the changes in the level of disease-related compounds. This approach is also appropriate for the isolation of individual components, different in their parameter A values, from the blood plasma. PMID- 12118191 TI - Chromosome alterations reflect clonal evolution in squamous cell carcinoma of the larynx. AB - BACKGROUND: The mechanism of multistage carcinogenesis initiated by environmental carcinogens produces clonal evolution of initiated cells to the stage of metastasis. However, the simultaneous co-existence of various cell clones is not excluded. The analysis of chromosome alterations appears to be the best method to assess the clonal composition of a tumor. MATERIAL/METHODS: Laryngeal tumor specimens and their corresponding metastases to the adjacent lymph nodes (20 pairs) were analysed by comparative genome hybridization (CGH). RESULTS: The profile of gains and losses of DNA copy number was found to be fairly similar in the primary tumor location and in its metastasis. In this study the most frequent losses were found on 3p, 5q, 9p, 13p, and 13q, while gains occurred in 1q, 3q, 5p and 11q. The gains and losses were found more frequently in metastasis than in primary tumor, except for the loss of 3p and the gain of 5p. The biological function of the latter chromosome alterations seems to be limited to tumor growth, rather than metastasis formation. CONCLUSIONS: A comparison of the profiles of chromosome alterations in primary tumor locations and metastases indicate that the progression of laryngeal cancer is connected with the clonal evolution of tumor cells. PMID- 12118190 TI - Effect of hydroxyethyl starch (HAES) on degree and kinetics of erythrocyte aggregation studied with dielectric spectroscopy method. AB - BACKGROUND: Colloidal fluids, e.g. hydroxyethyl starch (HAES), used as blood plasma substitutes during acute controlled normovolemic hemodilution may cause erythrocyte aggregation. The aim of the presented study was to assess the degree and rate of erythrocyte aggregation in the media containing different concentrations of HAES. MATERIAL/METHODS: The experiments were carried out on heparinized venous blood and on erythrocyte suspension in 0.9% NaCl. RESULTS: Under stationary conditions, an increase of blood and erythrocyte suspension relative electric permitivity (e), with an increase of HAES concentration in the sample, was observed after HAES supplementation. Sample flow through a measuring chamber resulted in decrease of the eL value (determined for 100 kHz). Stopping the flow led to non-linear increase of eL values in the function of time, where two components - characterized by rapid and slow variability, respectively, could be distinguished. The increase was characterized by t1 and t2 relaxation times, which can be related respectively to the process of erythrocyte rouleaux formation and to the formation of 3-D network aggregates. CONCLUSIONS: 1) The values of eL are differentiated by the degree of erythrocyte aggregation due to the presence of HAES, both in whole blood and in erythrocyte suspension in 0.9% NaCl solution, 2) The degree of erythrocyte disaggregation due to flow increases with the increase of flow intensity and correlates with decreasing eL value, 3) t1 and t2, which characterize the kinetics of erythrocyte aggregation, indicate a difference between aggregation processes taking place in whole blood and after hemodilution with HAES. PMID- 12118192 TI - Plasma soluble L-selectin following cardiopulmonary bypass (CPB) in children: is it a marker of the postoperative course? AB - BACKGROUND: There is increasing evidence that cytokine-inducible leukocyte endothelial adhesion molecules are instrumental in the postoperative inflammatory response following cardiopulmonary bypass (CPB). L-selectin was shown to be one of those neutrophil-endothelial cell adhesion molecules. This study aimed to investigate the relationship of the soluble adhesion molecule, sL-selectin, and the postoperative course in children undergoing CPB. MATERIAL/METHODS: To determine the time course of sL-selectin after CPB, serial blood samples of 9 children undergoing CPB were collected from the arterial line or from the bypass circuits preoperatively, on initiation of CPB and 1, 6, 12, 18, 24, and 48 hours postoperatively. Plasma was recovered immediately, aliquoted and frozen at -70 degrees C until use. Circulating sL-selectin molecules were measured with a sandwich enzyme-linked immunoabsorbent assay (ELISA) technique. There were significant changes in plasma levels of sL-selectin in patients following CPB, and these levels were associated with patient characteristics, operative variables and postoperative course. Low values of sL-selectin significantly correlated with inotropic support, low PRISM score, postoperative hypotension and fever. There was a significant association between the development of postoperative sepsis and low sL-selectin levels. No correlation was found between sL-selectin values and lactate concentration or neutrophil count. CONCLUSIONS: Our results suggest a relation between CPB-induced mediators and both early and late clinical effects. Although the mechanism for the changes of sL-selectin remains undetermined, the down-regulation of sL-selectin indicates neutrophil activation and supports the possibility that anti-adhesion therapies might participate in the prevention and treatment of the inflammatory response associated with CPB. PMID- 12118193 TI - Effect of renin-angiotensin system activation by dietary sodium restriction and upright position on plasma leptin concentration in patients with essential hypertension. AB - BACKGROUND: Both leptin and the renin-angiotensin system (RAS) are involved in the regulation of arterial blood pressure. This study was undertaken to assess the relationship between RAS and plasma leptin concentration in hypertensive patients under conditions of normal and restricted sodium supply and upright position. MATERIAL/METHODS: In 31 patients with essential hypertension (EHP - 14 F, 17 M, age 44I14 years, BMI 29.3I6.4 kg/m2) and 8 healthy subjects (NHS - 4 F, 4 M, age 37(17 years, BMI 25.3I6.6 kg/m2) plasma leptin concentration, plasma renin activity (PRA), and 24-hour urinary sodium excretion (UNa) were evaluated twice: first on a diet containing 100-120 mmol sodium per day and after 8 hours overnight bed rest, and a second time after 3 days of dietary sodium restriction (10-20 mmol daily) and 3 hours in upright position. RESULTS: Dietary sodium restriction and upright position was followed by a significant increase in PRA and decrease of UNa. By contrast, plasma leptin concentration showed a moderate decrease both in EHP and NHS. No significant correlation was found between PRA and plasma leptin concentrations in either of the groups examined. CONCLUSIONS: From the results obtained in this study we may conclude that dietary sodium restriction and upright position exerts only a moderate effect on plasma leptin concentration, in contrast to PRA, in both hypertensive and normotensive subjects. PMID- 12118195 TI - Assessment of the prevalence of iron deficiency anemia, by serum ferritin, in pregnant women of Southern Iran. AB - BACKGROUND: Iron-deficiency anemia (IDA) is a public health problem in the developing and even industrialized countries. Pregnant women and children under 5 years of age are among the high-risk population. Our main objectives in this study were to obtain the prevalence of IDA in pregnant women by routine methods and by serum ferritin. MATERIAL/METHODS: We analysed the blood of 270 healthy pregnant mothers, 16 weeks of gestational age. A series of determinations were conducted to determine haemoglobin concentration (Hb); red blood cells count (RBC); serum ferritin and other indexes. Then a questionnaire for epidemiological data, type of diet, level of education, laboratory data, etc. was filled. RESULTS: The mean values (SD) of haematological indexes were as follows: Hb 12.07I1.5 g/dl; serum ferritin 24.87I19.32 ng/ml; mean corpuscular haemoglobin concentration (MCHC) 31.9I1.4 g/dl; mean corpuscular volume (MCV) 82.2I9 fl and mean corpuscular haemoglobin (MCH) 26.4I3.2 pg. 28.5 % of the subjects were anaemic at the time of the study according to serum ferritin (SF < 12 ng/ml) and 16.7% of the mothers had low serum Haemoglobin (Hb<11 g/dl) (P=0.005). There was a positive correlation (r=0.76; P=0.01) between Hb concentration and serum ferritin levels. CONCLUSIONS: The prevalence of IDA was 28.5%, which is the same as the prevalence found in other developing countries (25-35%). This shows that in southern Iran we are still far behind the health status in the industrialized countries (5-8%). PMID- 12118194 TI - Phenotypic characterization of infiltrates in dilated cardiomyopathy - diagnostic significance of T-lymphocytes and macrophages in inflammatory cardiomyopathy. AB - BACKGROUND: Dilated cardiomyopathy (DCM) is linked to inflammatory cardiomyopathy (InfCM). To gain a profound insight into the underlying mechanisms, we phenotypically characterized pan leukocytes (CD18), naive T-lymphocytes (CD3, CD2, CD4, CD8), activated lymphocytes (LFA-1, LFA-3, VLA-4, ICAM-1, CD69, CD45R0), macrophages (Mac-1, 27E10), B-lymphocytes (CD19) and NK-cells (CD57) in DCM and control hearts. MATERIAL/METHODS: Biopsies from DCM patients (n=164, LVEF<40%) and specimens from non-cardiac death causes (controls; n=17) were immunostained. Biopsies exceeding >2.0 CD3+ lymphocytes per high power field/hpf and/or >1.5 CD3+ lymphocytes/hpf with numerous foci and HLA class I/DR abundance were evaluated positive for InfCM. RESULTS: InfCM+ biopsies (n=102; 63%) demonstrated significantly increased infiltrates with respect to all studied phenotypes except for CD19 and CD57 when compared with both DCM biopsies negative for InfCM (n=62) and the controls, whereas the latter two groups did not differ (Tukey-Kramer analysis). Virtually all phenotypes correlated with one another in multivariate analysis (except for B-lymphocytes and NK cells). Whereas HLA class I/DR abundance was present in 14% of the controls and 26% of the DCM biopsies not yielding InfCM, InfCM+ biopsies demonstrated significantly (<0.001) higher frequencies of HLA abundance (76%). CONCLUSIONS: The inflammatory process in InfCM comprises T-lymphocytes and macrophages, whereas B-lymphocytes and NK-cells are not significantly increased. InfCM is associated with HLA induction. CD69+, CD45R0+ and adhesion molecule bearing infiltrates indicate the activated state of lymphocytes, and 27E10 of macrophages in InfCM, respectively. Our data are in accordance with the hypothesis of a 'chronic active inflammatory process' involved in DCM. PMID- 12118197 TI - Spectrum and proteinase production of yeasts causing vaginitis in Saudi Arabian women. AB - BACKGROUND: Yeast vaginitis is a common infection. Data on the identity and prevalence of the etiological species would serve both epidemiological and therapeutic ends. Proteinase secretion by the invading yeast has been implicated in facilitating infection. Hence a prospective study was conducted to determine the species causing vaginitis in women from Saudi Arabia and the yeast's ability to produce proteinase. MATERIAL/METHODS: The subjects were patients with clinical signs of vaginitis. A high vaginal swab specimen from each patient was cultured, and only women yielding heavy yeast growth were enrolled. The isolated yeasts were identified by routine procedures, then evaluated for proteinase production in a medium containing bovine serum albumin. RESULTS: A total of 500 patients were studied. Candida albicans was the major species to cause vaginitis (59%), followed by C. glabrata (31%), C. tropicalis (4%), C. krusei (3.2%), Saccharomyces cerevisiae (1.6%), C. parapsilosis (0.6%), and C. kefyr (0.6%). All isolates of C. albicans and C. parapsilosis and 95% of C. tropicalis produced proteinase, while isolates from the remaining species did not. CONCLUSIONS: These results indicate that 59.6% of yeast vaginitis in women from Saudi Arabia is caused by C. albicans, and 31.6% by C. glabrata. Of relatively low prevalence were C. tropicalis and C. krusei. The study also revealed that both proteinase producers and non-producers were involved in causing vaginitis, indicating that proteinase secretion is not an essential factor in the pathogenesis of yeast vaginitis. PMID- 12118196 TI - CT- guided transthoracic fine needle aspiration of pulmonary lesions: accuracy and complications in 294 patients. AB - BACKGROUND: The purpose of this study was to prospectively evaluate the effectiveness of CT-guided transthoracic fine needle aspiration in the diagnosis of pulmonary lesions and to determine the complication rate of this procedure. MATERIAL/METHODS: A prospective review was undertaken of 316 patients who underwent CT-guided transthoracic fine needle aspiration performed at our center between October 2000 and April 2001. Twenty-two patients were excluded because no final diagnosis was achieved. The present study included 294 patients. All fine needle aspirations were performed with a 22-gauge aspirating needle under CT guidance. RESULTS: An accurate diagnosis was made in 228 of 259 malignant lung lesions (88%). A specific diagnosis was obtained in 34.3% of the benign lesions. The sensitivity for malignancy and specificity for benign lesions were 88% and 100%, respectively. Positive and negative predictive values were 100% and 53%, respectively. There was no false-positive diagnosis for malignancy. Sensitivity was 87% for centrally located lesions and 89.3% for peripherally located lesions (p>0.05). Complications included pneumothorax, 24 of 294 cases (8.2%); chest tube, 3 cases (12.5%); minor hemoptysis, 5 cases (1.7%); and pulmonary hemorrhage, 4 cases (1.4%). CONCLUSIONS: Our results suggest that this procedure has high diagnostic accuracy and an acceptable rate of complications. PMID- 12118198 TI - Changes of arterial pressure in patients with hyperthyroidism during therapy. AB - BACKGROUND: Hyperthyroidism affects the circulatory system, producing an increase in cardiac output and an enhanced vascular bed volume. These phenomena are important in the regulation of blood pressure. The present study was designed to evaluate the effects of short- and long-term treatment of hyperthyroidism on arterial pressure, taking into account the indices of cardiac function and peripheral vessel resistance. MATERIAL/METHODS: Fifty-one patients with hyperthyroidism and 30 healthy controls were investigated. The patients were examined before treatment, two weeks after initiation of therapy, and after attainment of a euthyroid state. Thiamazole was used as the antithyroid agent. Blood pressure and serum thyroid hormones were determined and ultrasonographic examination was performed on all the subjects, after the application of a diet containing 120 mmol of sodium and 70 mmol of potassium for three days. RESULTS: Patients with hyperthyroidism had higher systolic blood pressure and lower diastolic blood pressure than the healthy controls. After short-term treatment, systolic blood pressure returned to normal, while diastolic blood pressure was normalized only after long-term treatment. CONCLUSIONS: Regulation of blood pressure in patients with hyperthyroidism is a complex process. Systolic blood pressure is mainly related to cardiac output and returns to normal range shortly after the initiation of therapy, while diastolic blood pressure is related to peripheral vascular resistance and is normalized after long-term treatment. PMID- 12118199 TI - Left atrial function in patients with renal transplantation. AB - BACKGROUND: Chronic renal failure leads to structural changes and cardiac functional abnormalities known as uremic cardiomyopathy. Eliminating excess water and improving the composition of the inner environment leads to at least partial cardiac function improvement, which is reflected primarily in favorable changes in left ventricular indices. The aim of our study was to evaluate left atrial function in patients after renal transplantation and compare them with clinically healthy subjects. MATERIAL/METHODS: 10 renal transplant patients (2 women, 8 men; mean age 47.8 +/- 6.4 years), treated before transplantation with repeated hemodialysis, were subjected to standard transthoracic echocardiographic examination. Atrial function was evaluated in M-mode and 2D projections and by cross-sectional Doppler echocardiography. The results obtained were compared with 16 healthy controls (9 women, 7 men; mean age 39.7 +/- 9.4 years). RESULTS: Maximal left atrial dimension (LAmax), left atrial dimension obtained in M-mode of the long axis in parasternal projection (LAa), ejection time (ETlp) and pre ejection period (PEPlp) were significantly higher in renal transplant patients than in healthy subjects. In both investigated groups there were no differences in minimal left atrial dimensions (LAmin), PEPlp/Etlp ratio, P wave time (P), left atrial fiber shortening fraction (FS%lp), passive evacuate fraction (FBOlp), total left atrial fraction (FClp), or IElp ratio. CONCLUSIONS: Abnormal function of the left atrium in the course of uremia treated with repeated hemodialysis is not fully corrected after renal transplantation despite the elimination of many cardiovascular complications observed in chronic renal disease. PMID- 12118200 TI - Sulfonylurea receptor gene 16-3 polymorphism - association with sulfonylurea or insulin treatment in type 2 diabetic subjects. AB - BACKGROUND: The presence of a complex phenotype of type 2 diabetes results from impaired insulin secretion and action, whereas the mechanism of action of sulfonylurea derivatives, most commonly used in the treatment of type 2 diabetes, is based on their ability to directly inhibit the ATP-sensitive potassium channel (KATP), which leads to b-cell depolarization, subsequent influx of calcium and then insulin exocytosis. It has recently been demonstrated in healthy subjects that molecular variants of the gene encoding for the KATP subunit - sulfonylurea receptor gene (SUR1) are associated with a decreased response of insulin secretion to intravenous injection of tolbutamide, a sulfonylurea derivative. In this study we tested whether a molecular variant of the SUR1 gene, 16-3t, has a different distribution in type 2 diabetic patients with early failure of sulfonylurea therapy, compared to patients treatable with sulfonylurea despite long diabetes duration. MATERIAL/METHODS: The SUR1 polymorphism was genotyped in 68 type 2 diabetic patients who required insulin treatment and had known diabetes duration L 5 years, compared to 99 patients receiving oral agents (sulfonylurea alone or in combination with metformin or acarbose) with known diabetes duration of at least 15 years. RESULTS: We observed no significant differences in SUR1 16 3t genotype distributions or allele frequencies between the two examined groups. CONCLUSIONS: Our study provides evidence against a major impact of the SUR1 c16 3t polymorphism on the long-term effectiveness of therapy with sulfonylurea derivatives in type 2 diabetic patients. PMID- 12118201 TI - Hepatitis B virus serologic markers and anti-hepatitis B vaccination in patients with diabetes. AB - BACKGROUND: In Poland HBV infections are mainly the result of nosocomial infections or risky behavior. Active prevention plays a crucial role in limiting such infections. Patients suffering from insulin-dependent diabetes (type 1) incur high risk of infection with hepatotropic viruses because of frequent hospitalization and blood tests. The aim of our study was to determine the frequency of occurrence of serological markers of HBV infection among patients with diabetes type 1, to evaluate the humoral response to HBV vaccine depending on dosage, and to evaluate the influence of past HBV infections on vaccination response. MATERIAL/METHODS: 299 diabetic patients were vaccinated against hepatitis B with Engerix B in two doses (20 mg or 40 mg), on a 0-1-6 month schedule. The humoral response to the vaccination was evaluated in the 5th and 8th months after the first dose of vaccine according to sex, dose and previous detection of anti-HBc in serum. Serologic markers of HBV infection were detected with the immunoenzymatic-fluorescent ELFA method, with bioMerieux tests using the VIDAS system. RESULTS: 98.7% of patients achieved protective anti-HBs titer after vaccination. Women responded better than men. There were no differences in mean anti-HBs concentration by vaccine dose. Patients with previous serum anti-HBc achieved anti-HBs titers (p<0.00001). CONCLUSIONS: The frequency of anti-HBc among patients with diabetes confirms the widespread risk of HBV infection in the Polish population. Diabetics should be vaccinated against hepatitis B with standard doses. Diabetics with anti-HBc in serum can be vaccinated with a single dose. PMID- 12118202 TI - Ozone therapy and the activity of selected lysosomal enzymes in blood serum of patients with lower limb ischaemia associated with obliterative atheromatosis. AB - BACKGROUND: The paper compares the effects of ozone therapy and conventional balneological methods on health condition of patients with obliterative atheromatosis and on serum activity of three lysosomal enzymes. MATERIAL/METHODS: Sixty-four patients with lower limb ischaemia in the course of obliterative atheromatosis (without diabetes) were enrolled in the study. Thirty-two patients were treated with ozone administered by intravenous infusions and 30-minute aerosol oxygen-ozone baths. A comparative group was formed of 32 patients treated with traditional balneology. There was also a control group made up of 30 healthy subjects. Ozone therapy as well as traditional balneology were administered daily for the period of 10 days, excluding Saturdays and Sundays. Blood for biochemical analysis was collected from elbow vein in the following time intervals: 24 hours before ozone therapy or classical balneology, one hour after therapy and on the 10th day of treatment. The activity of cathepsin D, acid phosphatase and arylsulphatase as well as the levels of a-1-antitrypsin (protease inhibitor) were determined in blood serum of patients with obliterative atheromatosis. RESULTS: In patients who received ozone therapy the activity of analysed lysosomal hydrolases returned to the values typical for healthy subjects. Patients' general condition also improved. The use of traditional balneological methods did not result in any significant change either in the activity of lysosomal hydrolases, the level of a-1-antitrypsin or general condition of patients. CONCLUSIONS: Ozone therapy administered by intravenous infusions and aerosol oxygen-ozone baths of lower extremities yields much better therapeutic results in comparison with classical balneology. PMID- 12118203 TI - TH1/TH2 balance in the subretinal fluid of patients with rhegmatogenous retinal detachment. AB - BACKGROUND: The pathogenesis of rhegmatogenous retinal detachments remains unclear, but some data suggest immune system involvement. Cytokines are substances secreted by cells in order to modulate the function of other cells. They exert primarily local effects. Because the TH1/TH2 balance is crucial in the immune response, the predominance of one cytokine profile in subretinal fluid may be of particular importance in understanding its pathogenesis. MATERIAL/METHODS: We measured the titres of IL-10 and tIL-12 in the subretinal fluid of 36 rhegmatogenous retinal detachment patients using ELISA assays. RESULTS: The mean level of IL-10 and tIL-12 in the subretinal fluid was 4.75 pg/ml and 106.5 pg/ml, respectively. CONCLUSIONS: It would appear that IL-10 and IL-12 are produced by cells in subretinal fluid, and their presence may be evidence for an existing inflammatory process. Despite the tendency to decreased cytokine concentration over time and the tendency to higher concentrations of IL-10 and tIL-12 when the detachment of the retina is more extensive, no predominance of TH1- or TH2-type response was observed. PMID- 12118204 TI - Frequency, consequences and pharmacological treatment of gastroesophageal reflux in children with cystic fibrosis. AB - BACKGROUND: Gastroesophageal reflux (GER) may aggravate chronic bronchopulmonary diseases. The study evaluated GER frequency and characteristics in children with Cystic Fibrosis (CF) as well as its consequences and pharmacological treatment. MATERIAL/METHODS: 40 CF children aged 1.3 to 20 years were examined. The study methodology involved: medical files analysis, anamnesis, physical examination, growth status estimation, esophageal pH-metry and upper gastrointestinal tract endoscopy with histological examination of esophageal biopsies. RESULTS: Based on pH-metry results, the diagnosis of GER was established in 22 children (55%). Mild GER (Index Reflux - IR 5-10%) was found in 12 children (54.5%), moderate GER (IR 10-20%) in 7 (31.8%) and severe GER (IR>20%) in 3 (13.6%). Ten patients with moderate or severe GER underwent endoscopy, which revealed GER-related esophagitis in 8 cases. There was no statistical difference of GER frequency and degree according to: age, sex, growth status, presence of type DF508 mutation in CFTR genome and typical GERD symptoms. According to the ESPGAN proposition, cisapride or cisapride with ranitidine medication was instituted. Treatment analysis was performed in 19 cases after successful follow-up examinations carried out three months later, indicating a significant decrease in reflux index, the longest episode duration and the number of episodes longer than 5 minutes. Improvement of endoscopic picture was noticed after treatment. CONCLUSIONS: High frequency of gastroesophageal reflux and its consequences among children with cystic fibrosis, as well as the possibility of well-tolerated and efficient treatment of GER, indicate that diagnostics of GER among children with CF should be obligatory. PMID- 12118205 TI - Concentration of b2-microglobulin and percentage of CD4 lymphocytes in peripheral blood in patients with chronic HCV infection during IFN-a therapy. AB - BACKGROUND: We evaluated the serum b2-MG concentration and CD4 lymphocytes in the blood of HCV infected patients during IFN-a therapy, searching for a correlation between b2-MG concentration, CD4 lymphocytes, and therapy effectiveness, as well as morphological changes in the liver. MATERIAL/METHODS: 24 patients with chronic HCV infection were treated with IFN-a2a. The serum b2-MG concentration was measured with the use of a method based on the fluorescent modification of the immunoenzymatic technique. The percentage of T CD4 lymphocytes in the blood was measured by direct immunofluorescence using monoclonal antibodies. RESULTS: The number of CD4 lymphocytes in the blood was lower in the HCV infected patients (864/ml; preferred values from 1,300 to 2,100/ml), the b2-MG concentration was elevated (2.37 mg/dl) in comparison to the preferred values (1.52 mg/dl; p<0.05). IFN-a therapy caused an increase in the b2-MG. The highest increase was observed among patients who did not eliminate the virus (from 2.39 to 4.10 mg/dl). In the initial period of interferon therapy an increase was observed (from 729 to 1082/ml) in the number of CD4 lymphocytes among those patients who eliminated the virus and a decrease (from 947 to 853/ml) in the patients who were not treated successfully. CONCLUSIONS: A significant increase in b2-MG during interferon therapy in patients with chronic HCV infection is a predictor of poor outcome. An increase in the number of CD4 lymphocytes in the initial phase of treatment suggests a positive outcome. PMID- 12118206 TI - Cheyne-Stokes respiration during sleep: a possible effect of body position. AB - BACKGROUND: Cheyne-Stokes Respiration (CSR) is a common finding in Chronic Heart Failure and Stroke patients. The body position effect during sleep on obstructive breathing abnormalities is well known. However, the effect of body position during sleep on breathing abnormalities of central type like CSR has not been well documented. MATERIAL/METHODS: Six sleep studies (two complete Polysomnographic (PSG) evaluations and four Pulse Oximetry recordings (PO)), were carried out in a 57-year-old female patient with a recent Cerebro Vascular Accident (CVA who had both Obstructive Sleep Apnea (OSA) and CSR. RESULTS: The first PSG was carried out two months post-stroke and revealed a severe, continuous CSR pattern during Non Rapid Eye Movements (NREM) sleep (mainly with central apneas), and Obstructive Sleep Apnea (OSA) during Rapid Eye Movements (REM) sleep, independent of body position: Supine Respiratory Disturbance Index (SRDI) = 85.2 and Lateral RDI (LRDI) = 95.4. A second PSG was performed three months later after an overall clinical improvement and showed a complete disappearance of CSR during NREM sleep and OSA during REM sleep in her lateral posture (LRDI = 0), while the RDI in the supine posture was only slightly improved (SRDI = 73.2). The CSR pattern was less severe and was characterized mainly by central hypopneas. Two PO recordings between the PSG studies showed similar improvement trends. Two additional PO recordings, two and three weeks after the last PSG (the first one with the patient lying supine and the second one with the patient lying on her side throughout the night), revealed a further significant improvement in the supine posture (SRDI = 37.5). CONCLUSIONS: The results of this study suggest that body posture may play a role not only in the prevalence and severity of obstructive breathing disorders, but also in CSR, a central type of breathing abnormalities during sleep. PMID- 12118207 TI - Psychiatric symptoms in a patient with the clinical features of MELAS. AB - BACKGROUND: This article presents a case study of a 32-year-old Polish male patient clinically diagnosed with the MELAS syndrome, the second such patient reported in this country. CASE REPORT: The patient presents with a long history of neurological episodes, consisting of acute neurological deficits suggestive of CVA with no evidence of focal lesion and very rapid remission of symptoms. These episodes began when the patient was 19 years old and have recurred since that time at intervals of 1-5 years. A recent psychiatric episode with unusual visual hallucinations brought the case to the attention of the present authors, who initiated further diagnostic testing. A muscle biopsy revealed ragged red fibers. CONCLUSIONS: The features in this case meet the diagnostic criteria for a clinical diagnosis of MELAS, pending confirmation of a pathogenic mutation. PMID- 12118209 TI - Robotically-assisted coronary artery surgery with and without cardiopulmonary bypass - from first clinical use to endoscopic operation. AB - BACKGROUND: Recently, the ZEUS(tm) Robotic Surgical System has been introduced to increase the precision of endoscopic cardiac surgery. This study investigated its clinical use for endoscopic coronary artery bypass grafting. MATERIAL/METHODS: Between 1998 and 2001, 41 patients with single and multivessel disease were operated on using the ZEUS(tm) system. The robotic system was introduced step by step into clinical practice. Initially, the system was used only for endoscopic internal mammary artery (IMA) harvest (n=12), later for coronary anastomoses on the arrested (n=13) or beating heart after median sternotomy (n=6), and finally for endoscopic coronary bypass grafting on either the arrested (n=2) or beating heart (n=8). RESULTS: Endoscopic IMA harvest ranged from 48 to 110 min and was completed in all cases. In the sternotomy group, the robotic anastomosis time averaged 21 min on the arrested and 25 min on the beating heart, respectively (n.s.). In the endoscopic cases, the average time for endoscopic anastomosis was 41 min on the arrested and 36.5 min on the beating heart (n.s.), with an overall duration of surgery between 4.0 and 8.0 hours. One endoscopic case was intraoperatively converted to a MIDCAB procedure with manual anastomosis. The total patency rate of all graft anastomoses, confirmed by early postoperative angiographic control, was 97%. One patient underwent reoperation with an uneventful postoperative course. CONCLUSIONS: The present study demonstrates the feasibility of endoscopic coronary revascularization using a computer-assisted surgical robotic system on the arrested and beating heart in selected patients. PMID- 12118208 TI - Thermal effect of intravascular MR imaging using an MR imaging-guidewire: an in vivo laboratory and histopathological evaluation. AB - BACKGROUND: Intravascular magnetic resonance (MR) imaging to guide interventional procedures is a rapidly growing field. A primary concern with these new techniques is their thermal safety. The purpose of this study was to evaluate, in vivo, the thermal effect of an MR imaging-guidewire (MRIG) for intravascular MR imaging (IVMRI). MATERIAL/METHODS: Two indications of potentially adverse local heating were investigated: blood coagulation disorders and pathologic changes in target vessels. Experiments were performed on ten rabbits with a 1.5 T MR scanner. Using a 0.64-mm MRIG as the RF receiver, we imaged the target aorta using a fast spin-echo pulse sequence with an average specific absorption rate (SAR) of 0.6 W/kg. The total MR imaging time was approximately 70 minutes. RESULTS: There were no abnormal value changes of the coagulation factors between pre- and post-IVMRI, no clinical manifestations of blood coagulation disorders, and, histopathologically, no thermal damage in target vessels. CONCLUSIONS: This study demonstrates, from a pathophysiological point of view, the potential safe use of the MR imaging-guidewire for intravascular MR imaging. Further study is required to precisely define the boundaries of these safe operating parameters. PMID- 12118210 TI - Towards a semantic medical Web: HealthCyberMap's tool for building an RDF metadata base of health information resources based on the Qualified Dublin Core Metadata Set. AB - BACKGROUND: HealthCyberMap (http://healthcybermap.semanticweb.org/) aims at mapping Internet health information resources in novel ways for enhanced retrieval and navigation. This is achieved by collecting appropriate resource metadata in an unambiguous form that preserves semantics. MATERIAL/METHODS: We modelled a qualified Dublin Core (DC) metadata set ontology with extra elements for resource quality and geographical provenance in Prot g -2000. A metadata collection form helps acquiring resource instance data within Prot g . The DC subject field is populated with UMLS terms directly imported from UMLS Knowledge Source Server using UMLS tab, a Prot g -2000 plug-in. The project is saved in RDFS/RDF. RESULTS: The ontology and associated form serve as a free tool for building and maintaining an RDF medical resource metadata base. The UMLS tab enables browsing and searching for concepts that best describe a resource, and importing them to DC subject fields. The resultant metadata base can be used with a search and inference engine, and have textual and/or visual navigation interface(s) applied to it, to ultimately build a medical Semantic Web portal. Different ways of exploiting Prot g -2000 RDF output are discussed. CONCLUSIONS: By making the context and semantics of resources, not merely their raw text and formatting, amenable to computer 'understanding,' we can build a Semantic Web that is more useful to humans than the current Web. This requires proper use of metadata and ontologies. Clinical codes can reliably describe the subjects of medical resources, establish the semantic relationships (as defined by underlying coding scheme) between related resources, and automate their topical categorisation. PMID- 12118211 TI - Microcirculation in the diabetic foot as measured by a multichannel laser Doppler instrument. AB - BACKGROUND: The purpose of this study was to investigate microvascular perfusion in insulin-dependent diabetic patients at various locations on the foot, and to determine which part of the foot is most sensitive to microangiopathic changes. All the parameters of postocclusive reactive hyperemia calculated from multichannel laser Doppler recordings were also evaluated to find the most valuable measure for diabetes examination. MATERIAL/METHODS: Our study involved 65 subjects divided into four subgroups: male and female controls, and male and female IDDM patients without overt complications. The measurements were performed with a multichannel laser Doppler perfusion monitor using surface probes located in the distal parts of the lower limbs. The occlusion test was performed using a cuff located on the limb above the knee. Multivariate discriminatory analysis was used to evaluate the data. RESULTS: The most valuable data were obtained by recordings from the laser-Doppler probes located on the hallex and the base of the little toe. The maximum hyperemic response for both sex subgroups was significantly lower in the diabetic patients. The time to peak flow was higher in male diabetics. The half-time for hyperemia was significantly longer in the male diabetic patients. CONCLUSIONS: The females showed smaller changes in foot perfusion than the males, probably due to protection by estrogens. The best locations for perfusion measurement are the most distal, especially the hallex and the base of the little toe. The most valuable parameters of postocclusive hyperemia were maximum response, time to peak flow, and half-time of hyperemia. PMID- 12118212 TI - Mechanisms of knowledge learning and acquisition. AB - The mechanism by which knowledge enters into memory has been a source of debate for some time. Theorists have proposed several models that aim at explaining the sequence of events from the perception of a stimulus, to its entrance into long term storage. Much of this work was prompted by early research into the nuances of classical conditioning where it was first firmly established that organisms are capable of detecting covariations of stimuli within their environment. Subsequent work in the field has shown that these covariations form the basis for the mental representation of our surroundings, as well as the basis of learning. Work within the field of classical conditioning, along with the advance of computer technology and neuroscience has made these architectural models even more complex. Furthermore, experiments designed to support some of these proposed models have revealed that there are several conditions that can either aid or inhibit the transition of information into permanent storage. In this review we explore a number of these models, along with some classic critiques that have been levied against them. We also provide some history into the form of knowledge, termed 'implicit knowledge', as well as some of the proposed mechanisms of implicit knowledge acquisition. We conclude by exploring the newly proposed theoretical framework within which implicit learning theory operates. PMID- 12118213 TI - The Fragile Histidine Triad gene and breast cancer. AB - The Fragile Histidine Triad (FHIT) Gene, encompassing the FRA3B fragile site at chromosome 3p14.2, is a tumor suppressor gene involved in different tumor types. Abnormalities of the FHIT locus were found in many established cancer cell lines and tumors including breast cancer. In sporadic breast cancer, loss of heterozygosity within the FHIT gene has been observed at different frequencies and has been correlated with tumor progression. Aberrant FHIT transcripts have been reported in breast cancer. Furthermore, Fhit protein loss is correlated with prognosis. We summarized the research advances of FHIT genetic, epigenetic change and aberrant Fhit protein expression in breast cancer. Fhit protein may be a novel prognostic factor for breast cancer. FHIT gene therapy may potentially be clinically useful for treatment of breast cancer, suggesting further research involving FHIT introduction should be performed in breast cancer. PMID- 12118214 TI - TGF-beta (transforming growth factor-beta) in chronic inflammatory conditions - a new diagnostic and prognostic marker? AB - TGF-beta is a cytokine with varied properties and pleiotropic activity. It is released in an inactive form. To exhibit its biological activity, it requires binding to extracellular matrix proteins and, after that, proteolytic elimination of LAP (Latent Associated Protein) and LTBP (Latent TGF-beta Binding Protein). The process involves, among others, tissue transglutaminase, thrombin and plasmin. By stimulation of specific receptors, it influences transcription of some genes and translation of formed mRNA. Locally, it demonstrates proinflammatory properties whereas systemically, it has primarily a potent immunosuppressive effect. TGF-beta, by affecting proliferation, differentiation and migration of cells, as well as stimulation of extracellular matrix protein production, plays an important role in tissue regeneration and remodeling, but also in fibrosis. TGF-beta is also indispensable to maintain immune homeostasis of the organism. Reduced TGF-beta activity is considered to be responsible for development of autoimmune disorders in the course of several pathologic conditions. This cytokine plays an important role in the pathogenesis of chronic inflammatory processes taking place, among others, in inflammatory bowel diseases (IBD) and chronic hepatitis B and C. The paper presents a review of literature concerning diagnostic and prognostic value of TGF-beta level determinations in blood and tissue bioptates of patients with chronic non-specific enteritis and chronic hepatitis B and C. PMID- 12118217 TI - Use of spiral computerized tomography angiography in patients with cerebral aneurysm. Our experience. AB - BACKGROUND: The purpose is to highlight the usefulness of CT angiography (CTA) in the diagnosis and surgical treatment of cerebral aneurysms. METHODS: Thirty-one patients with subarachnoid haemorrhages were subjected to CT angiography and in those cases where this test did not reveal the aneurysm or did not supply sufficient information relating to it, subsequently a digital subtraction angiography was also performed. Each aneurysm-positive CTA was re-processed using the 3-D techniques, with the neuro-radiologist and the neuro-surgeon working in close co-operation. RESULTS: In 27 cases the CTA diagnosed an aneurysm, and in the 4 cases where no vascular malformations were revealed, also the traditional angiography did not show any pathology. In 17 out of 18 cases operated on in order to clip the aneurysm, the CTA supplied all the information needed for the surgery and it was possibile to reconstruct images similar to those of the surgical field. This led to improvement in the programming of the surgical intervention; in 1 case only was it also necessary to perform the DSA before the operation. CONCLUSIONS: CT angiography, because it is non-invasive, easy to perform, diagnostically reliable, and because the 3-D re-constructions offer the chance to create images of the possible operating field, is the first-choice test to be adopted in the treatment of subarachnoid haemorrhages, even though in some cases the use of the traditional angiography is still necessary and should be carried out whenever the CTA does not reveal vascular malformations. PMID- 12118218 TI - Hypoflow and hyperflow in diffuse axonal injury. Prognostic and therapeutic implications of transcranial Doppler sonography evaluation. AB - BACKGROUND: In the present report we describe the results of a study aimed at evaluating the cerebral haemodynamics and the neuroradiological findings observ ed in 7 consecutive patients, 4 adults and 3 children (6, 8 and 10 years old), affected by diffuse axonal injury (DAI). METHODS: All the patients were admitted to the Paediatric or Adult Intensive Care Unit with GCS scores less than 8 after a severe brain injury. Serial head CT scan and trans-cranial Doppler sonography (TCD) examinations were carried out in all patients; MRI was carried out in the paediatric patients only. TCD of the middle cerebral arteries was performed through the temporal bone window. In 6 cases (2 paediatric) diuretic osmotic therapy was immediately administered and in 6 cases (3 paediatric) barbiturates and hyperventilation were also used. RESULTS: Hyperflow, variably responsive to barbiturate therapy of vasoparalysis, was observed in all paediatric patients and in 3 adult subjects (85.7%: 6 out of 7 pa-tients) by means of TCD. CONCLUSIONS: Observation of these phenomena allowed us to modify the pharmacological treatment and/or perform external cerebrospinal fluid (CSF) drainage (4 cases). Compartimental hyperflow TCD pattern was evident in 1 patient. Although the limited number of patients in our series does not allow definitive conclusions, we strongly believe that TCD monitoring is an useful tool in planning surgical strategy in patients with DAI. PMID- 12118219 TI - Spinal intradural extramedullary tumors. Personal experience. AB - BACKGROUND: Spinal intradural extramedullary tumors account for 2/3 of all intraspinal neoplasms and are mainly represented by meningiomas and schwannomas, with the former accounting for the 25-46% of all primary intraspinal tumors. Technical advances in imaging technique, magnetic resonance imaging (MRI) and surgical procedures have brought about significant better clinical results in the last 2 decades. Neverthless a small percentage of patients still present poor postoperative outcome mainly related to the duration of clinical history, the severity of preoperative neurological deficits and to some specific anatomo surgical aspects. METHODS: In an effort to clarify the influence of these factors on patient's outcome, the authors analyze the clinical, surgical and prognostic data of 41 patients with intradural extramedullary spinal tumor surgically treated between January 1990 and December 1999. The follow-up period ranged from 1 to 9 years. The clinical history until admission and treatment was 3-48 months for meningiomas and 1-72 months for schwannomas. RESULTS: Morbidity and mortality rate was 5 and 0% for meningiomas and 0 and 6% for schwannomas. Almost all the patients experienced a significant neurological improvement after surgery, with a percentage of Nurick's grade 1 and 2 of 68% among patients with meningiomas and 66% among patients with schwannomas. Removal of the tumor was complete in 90% of meningiomas and 94% of schwannomas. CONCLUSIONS: The authors address radical surgery as the ideal goal in these neoplasms, as it gives the patients the best long-terms results, pointing out the importance of systematic early identification of any main radiculomedullary artery during debulking and dissection of the tumor to avoid any risk of severe postoperative neurological worsening of the patients. PMID- 12118220 TI - Penetrating brain injury with nasal entry by a plastic stick. Case report. AB - A case of a 52-year-old male presented with an unusual penetrating brain injury with nasal entry. At admission he had erythema of periorbital soft tissue in the left eye and epistaxis. His neurological condition was lethargic (Glasgow Coma Scale of 13) with nonfluent aphasia. Computed tomography scan revealed intracranial contusion hematoma in the left frontal lobe and fracture of the left frontal base, which were treated surgically. At the 6-month follow-up he still showed nonfluent aphasia. Disturbances, mostly cognitive, were noted on his psychological tests. A survey of the literature reveals a few cases of this nature in penetrating brain injury with nasal entry. A penetrating brain injury with nasal entry which causes nonfluent aphasia is discussing. PMID- 12118221 TI - "Matrioska head". Case report of calcified chronic subdural hematoma. AB - We report an asymptomatic case of a large calcified chronic subdural hematoma (CCSH). Skull X-ray examination, computer tomography (CT) and magnetic resonance imaging (MRI), showed an adhering calcification extending to the whole cerebral cortex, as if the skull had another concentric skull inside it. These radiological findings recall the famous Russian doll named "Matrioska"; for this reason we defined this case "Matrioska head". The patient was absolutely asymptomatic and we discuss the causes that may give the calcification of chronic subdural hematoma (CSH). PMID- 12118222 TI - Trigeminal palsy caused by triventricular hydrocephalus. A case report. AB - A case of left trigeminal palsy caused by triventricular hydrocephalus and completely recovered after ventriculo peritoneal shunt is reported. The case was studied by MR before and after the operation. Preoperative MR showed triventricular hydrocephalus with marked reduction of the left Meckel's cave. Following ventriculo peritoneal shunt the patient showed a complete recovery of the left 5th nerve palsy and NR showed the expansion of the left Meckel's cave. The authors suggest that the trigeminal palsy was due to intracranial hypertension probably associated to a weakness of the lateral wall of the left Meckel's cave. No similar case have been previously reported in the literature. PMID- 12118223 TI - Cerebellar mature teratoma in adulthood. AB - Mature teratoma of the posterior cranial fossa in adults is extremely rare. We report a particularly rare case of medio-lateral cerebellar mature teratoma that became symptomatic in a middle-aged man. The CT revealed the lesion of heterogeneous density with calcifications in the solid medial portion. Only the MRI could reliably define the borders of the cystic component extending into the left cerebellar lobe. Histologically the presence of fully matured representative tissues of the 3 germ layers ensured the diagnosis of mature teratoma. We suggest that the cyst formation from progressive latent hemorrhage and/or secretion from the gland cells of the tumor, may be responsible for the clinical decompensation even in adulthood. PMID- 12118224 TI - An unusual presentation of occult spinal dysraphism. AB - Occult spinal dysraphism can lead to irreversible neurological complications, early diagnosis and treatment are necessary. It can be suspected from the presence of any cutaneous abnormality. We report a case with bony spur formation on the top of the 5th lumbar vertebra spinose process covered with skin mimicking a meningocel sac. By the help of this bony spur tethered cord could be diagnosed before any neurological deficit. PMID- 12118225 TI - Management Principles. PMID- 12118226 TI - Biologic Approaches to Managing Advanced Breast Cancer. PMID- 12118228 TI - First-Line Chemotherapy for Advanced Breast Cancer. PMID- 12118227 TI - New Hormones for Advanced Breast Cancer. PMID- 12118229 TI - Salvage Chemotherapy for Metastatic Breast Cancer. PMID- 12118230 TI - Management of Bone Metastases in Advanced Breast Cancer. PMID- 12118231 TI - New Perspectives in Acute Pain Management. PMID- 12118233 TI - The Role of Non-opioid Analgesics for the Management of Postoperative Pain. PMID- 12118232 TI - The Efficacy of Nonsteroidal Anti-inflammatory Drugs for Acute Pain. PMID- 12118234 TI - Nonnarcotic Analgesics in Short-term Pain: Musculoskeletal Disorders. PMID- 12118235 TI - The Safety of NSAIDs and Related Drugs for the Management of Acute Pain: Maximizing Benefits and Minimizing Risks. PMID- 12118236 TI - Clinical Implications of Cyclooxygenase Enzymes: COX-1/COX-2 Role of the New NSAIDs. PMID- 12118237 TI - Nonnarcotic Analgesic Use in Acute and Cancer Pain: Results of Selected Meta analyses. PMID- 12118238 TI - Legal and Clinical Issues in Prescribing Controlled Substances. PMID- 12118239 TI - Lens epithelial cell death and reduction of anti-apoptotic protein Bcl-2 in human anterior polar cataracts. AB - PURPOSE: In light of the growing body of data implicating apoptosis in cataractogenesis, and in particular, the reported detection of apoptosis in posterior capsular opacification, the purported etiology of which, like that of anterior polar cataracts, is an aberrant transdifferentiation of lens epithelial cells into myofibroblastic cells, we hypothesized that apoptosis could also occur in anterior polar cataracts. Here we sought to examine whether apoptotic cell death occurs in lens epithelial cells from patients with anterior polar cataracts. METHODS: Cell death of lens epithelial cells from anterior polar cataracts, nuclear cataracts, and non-cataractous clear lenses was measured by TUNEL assay and DNA fragmentation assay. The expression of Bcl-2 and Bax was examined using reverse transcription-polymerase chain reaction and Western blot analysis. RESULTS: Cell death was detected in specimen from patients with anterior polar cataracts by TUNEL assay. DNA fragmentation assay showed the characteristic laddering pattern from the genomic DNA from anterior polar cataracts. The expression of Bcl-2 mRNA and its protein was markedly decreased in lens epithelial cells from patients with anterior polar cataracts. CONCLUSIONS: This study suggests that apoptotic cell death might occur in lens epithelial cells from anterior polar cataracts and decreased expression of Bcl-2 might play a role in the pathologic cellular mechanism of anterior polar cataracts. PMID- 12118240 TI - Translocation of macromolecules into whole rat lenses in culture. AB - PURPOSE: Little is known about the endocytosis and transcytosis of macromolecules into lens epithelium and fiber cells. The objective of this study was to determine if proteins (alpha-crystallins, beta-crystallins, and gamma crystallins), carbohydrate (dextran), and plasmid DNA translocate from culture medium into these parts of the lens, with and without prior encapsulation into liposomes. METHODS: alpha-Crystallins, beta-crystallins, gamma-crystallins, and dextran were coupled with the fluorochrome Texas red, and plasmid DNA was labeled with propidium iodide. Adult rat lenses were incubated in medium containing one of these components with and without prior encapsulation of the macromolecule in commercially available liposomes (BioPORTER for alpha-crystallins, beta crystallins, gamma-crystallins, and dextran; GenePORTER for plasmid DNA). Translocation of fluorescent macromolecule from the medium into the lens capsule, epithelium and fiber cells was monitored by confocal microscopy. RESULTS: alpha Crystallins, beta-crystallins, gamma-crystallins, and dextran were present in the capsule, epithelium, and fiber cells after 5 h of incubation. Translocation of fluorescent protein macromolecules into the epithelium was greatly facilitated by encapsulation in BioPORTER liposomes. These macromolecules were localized within the cytoplasm of epithelium and fiber cells. Plasmid DNA was localized to the epithelium, but not the fiber cells. Prior encapsulation of plasmid DNA into GenePORTER liposomes did not increase the intensity of fluorescence localized in epithelium. Without encapsulation, plasmid DNA preferentially localized to the nuclei of epithelial cells, while after encapsulation, plasmid DNA preferentially localized to the cytoplasm. CONCLUSIONS: After incubation with cultured lenses, large macromolecules comprised of proteins and carbohydrates were localized within the cytoplasm of epithelial cells and fiber cells. Prior encapsulation of protein macromolecules into BioPORTER liposomes facilitated the translocation of macromolecules into the cytoplasm of epithelium. Incubation of lenses with plasmid DNA resulted in localization to the epithelium, but not fiber cells. Localization of plasmid DNA was not facilitated by prior encapsulation in GenePORTER. Encapsulated DNA preferentially localized to the cytoplasm of epithelial cells, while without encapsulation, plasmid DNA localizes to the nuclei of epithelial cells. Together, these studies demonstrate that macromolecules of potential biological importance can readily pass through the lens capsule into epithelial cells and in some cases transcytose through the epithelium into fiber cells of the cortex. Furthermore, these studies suggest that prior encapsulation of protein macromolecules may be a possible therapeutic delivery system of physiologically important macromolecules into the epithelium and/or fiber cells of the intact lens. PMID- 12118241 TI - Monitoring post-translational modification of proteins with allosteric ribozymes. AB - An allosteric hammerhead ribozyme activated specifically by the unphosphorylated form of the protein kinase ERK2 was created through a rational design strategy that relies on molecular recognition of ERK2 to decrease the formation of an alternate, inactive ribozyme conformer. Neither closely related mitogen-activated protein kinases (MAPKs) nor the phosphorylated form of ERK2 induced ribozyme activity. The ribozyme quantitatively detected ERK2 added to mammalian cell lysates and also functioned quantitatively in a multiplexed solution-phase assay. This same strategy was used to construct a second ribozyme selectively activated by the phosphorylated (active) form of ERK2. This approach is generally applicable to the development of ribozymes capable of monitoring post translational modification of specific proteins. PMID- 12118242 TI - Three-dimensional structure of a bacterial oxalate transporter. AB - The major facilitator superfamily (MFS) represents one of the largest classes of evolutionarily related membrane transporter proteins. Here we present the three dimensional structure at 6.5 A resolution of a bacterial member of this superfamily, OxlT. The structure, derived from an electron crystallographic analysis of two-dimensional crystals, reveals that the 12 helices in the OxlT molecule are arranged around a central cavity, which is widest at the center of the membrane. The helices divide naturally into three groups: a peripheral set comprising helices 3, 6, 9 and 12; a second set comprising helices 2, 5, 8 and 11 that faces the central substrate transport pathway across most of the length of the membrane; and a third set comprising helices 1, 4, 7 and 10 that participate in the pathway either on the cytoplasmic side (4 and 10) or on the periplasmic side (1 and 7). Overall, the architecture of the protein is remarkably symmetric, providing a compelling molecular explanation for the ability of such transporters to carry out bi-directional substrate transport. PMID- 12118243 TI - Crystal structure of a laccase from Melanocarpus albomyces with an intact trinuclear copper site. AB - We have crystallized the ascomycete laccase from Melanocarpus albomyces with all four coppers present and determined the crystal structure at 2.4 A resolution. The enzyme is heavily glycosylated and consists of three cupredoxin-like domains, similar to those found in the Cu-depleted basidiomycete laccase from Coprinus cinereus. However, there are significant differences in the loops forming the substrate-binding pocket. In addition, the crystal structure of the M. albomyces laccase revealed elongated electron density between all three coppers in the trinuclear copper site, suggesting that an oxygen molecule binds with a novel geometry. This oxygen, required in the reaction, may enter the trinuclear site through the tunnel, which is open in the structure of the C. cinereus laccase. In contrast, the C-terminus on the M. albomyces laccase forms a plug that blocks this access. PMID- 12118244 TI - Gene-expression profiles predict survival of patients with lung adenocarcinoma. AB - Histopathology is insufficient to predict disease progression and clinical outcome in lung adenocarcinoma. Here we show that gene-expression profiles based on microarray analysis can be used to predict patient survival in early-stage lung adenocarcinomas. Genes most related to survival were identified with univariate Cox analysis. Using either two equivalent but independent training and testing sets, or 'leave-one-out' cross-validation analysis with all tumors, a risk index based on the top 50 genes identified low-risk and high-risk stage I lung adenocarcinomas, which differed significantly with respect to survival. This risk index was then validated using an independent sample of lung adenocarcinomas that predicted high- and low-risk groups. This index included genes not previously associated with survival. The identification of a set of genes that predict survival in early-stage lung adenocarcinoma allows delineation of a high risk group that may benefit from adjuvant therapy. PMID- 12118245 TI - Smac agonists sensitize for Apo2L/TRAIL- or anticancer drug-induced apoptosis and induce regression of malignant glioma in vivo. AB - A major concern in cancer therapy is resistance of tumors such as glioblastoma to current treatment protocols. Here, we report that transfer of the gene encoding second mitochondria-derived activator of caspase (Smac) or Smac peptides sensitized various tumor cells in vitro and malignant glioma cells in vivo for apoptosis induced by death-receptor ligation or cytotoxic drugs. Expression of a cytosolic active form of Smac or cell-permeable Smac peptides bypassed the Bcl-2 block, which prevented the release of Smac from mitochondria, and also sensitized resistant neuroblastoma or melanoma cells and patient-derived primary neuroblastoma cells ex vivo. Most importantly, Smac peptides strongly enhanced the antitumor activity of Apo-2L/tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) in an intracranial malignant glioma xenograft model in vivo. Complete eradication of established tumors and survival of mice was only achieved upon combined treatment with Smac peptides and Apo2L/TRAIL without detectable toxicity to normal brain tissue. Thus, Smac agonists are promising candidates for cancer therapy by potentiating cytotoxic therapies. PMID- 12118246 TI - Dystrophin deficiency markedly increases enterovirus-induced cardiomyopathy: a genetic predisposition to viral heart disease. AB - Both enteroviral infection of the heart and mutations in the dystrophin gene can cause cardiomyopathy. Little is known, however, about the interaction between genetic and acquired forms of cardiomyopathy. We previously demonstrated that the enteroviral protease 2A cleaves dystrophin; therefore, we hypothesized that dystrophin deficiency would predispose to enterovirus-induced cardiomyopathy. We observed more severe cardiomyopathy, worsening over time, and greater viral replication in dystrophin-deficient mice infected with enterovirus than in infected wild-type mice. This difference appears to be a result of more efficient release of the virus from dystrophin-deficient myocytes. In addition, we found that expression of wild-type dystrophin in cultured cells decreased the cytopathic effect of enteroviral infection and the release of virus from the cell. We also found that expression of a cleavage-resistant mutant dystrophin further inhibited the virally mediated cytopathic effect and viral release. These results indicate that viral infection can influence the severity and penetrance of the cardiomyopathy that occurs in the hearts of dystrophin-deficient individuals. PMID- 12118247 TI - Antidepressant, anxiolytic and anorectic effects of a melanin-concentrating hormone-1 receptor antagonist. AB - Melanin concentrating hormone (MCH) is an orexigenic hypothalamic neuropeptide, which plays an important role in the complex regulation of energy balance and body weight. Here we show that SNAP-7941, a selective, high-affinity MCH1 receptor (MCH1-R) antagonist, inhibited food intake stimulated by central administration of MCH, reduced consumption of palatable food, and, after chronic administration to rats with diet-induced obesity, resulted in a marked, sustained decrease in body weight. In addition, after mapping the binding sites for [(3)H]SNAP-7941 in rat brain, we evaluated its effects in a series of behavioral models. SNAP-7941 produced effects similar to clinically used antidepressants and anxiolytics in three animal models of depression/anxiety: the rat forced-swim test, rat social interaction and guinea pig maternal-separation vocalization tests. Given these observations, an MCH1-R antagonist may be useful not only in the management of obesity but also as a treatment for depression and/or anxiety. PMID- 12118248 TI - Structure of human CD1b with bound ligands at 2.3 A, a maze for alkyl chains. AB - The human genome encodes five nonpolymorphic major histocompatibility complex class I-like glycoproteins, CD1a to CD1e, that present lipid antigens for specific recognition by T lymphocytes. Using single alkyl chain detergents, we developed a protocol to generate recombinant human CD1b-lipid complexes. We present here the crystal structures of CD1b in complex with either phosphatidylinositol or ganglioside GM2 at 2.3 A and 2.8 A resolutions, respectively. The antigen-binding groove houses four interlinked hydrophobic channels that are occupied by the alkyl chains of the glycolipid plus two detergent molecules. A distinct exit beneath the alpha 2 helix further contributes to the plasticity of the binding groove. These structures reveal the mechanism by which two alkyl chain lipids bind to CD1b, and how CD1b can adapt to ligands of different alkyl chain length. They also suggest how very long alkyl chains, such as those of mycolic acid, could be fully contained within the binding groove. These results extend the spectrum of potential CD1b ligands by revealing that, in addition to two alkyl chain lipids, mono-alkyl and triple alkyl chain lipids can be accommodated in the binding groove. PMID- 12118249 TI - PKC-beta controls I kappa B kinase lipid raft recruitment and activation in response to BCR signaling. AB - NF-kappa B signaling is required for the maintenance of normal B lymphocytes, whereas dysregulated NF-kappa B activation contributes to B cell lymphomas. The events that regulate NF-kappa B signaling in B lymphocytes are poorly defined. Here, we demonstrate that PKC-beta is specifically required for B cell receptor (BCR)-mediated NF-kappa B activation. B cells from protein kinase C-beta (PKC beta)-deficient mice failed to recruit the I kappa B kinase (IKK) complex into lipid rafts, activate IKK, degrade I kappa B or up-regulate NF-kappa B-dependent survival signals. Inhibition of PKC-beta promoted cell death in B lymphomas characterized by exaggerated NF-kappa B activity. Together, these data define an essential role for PKC-beta in BCR survival signaling and highlight PKC-beta as a key therapeutic target for B-lineage malignancies. PMID- 12118250 TI - Mutations in the gene encoding the lamin B receptor produce an altered nuclear morphology in granulocytes (Pelger-Huet anomaly). AB - Pelger-Huet anomaly (PHA; OMIM *169400) is an autosomal dominant disorder characterized by abnormal nuclear shape and chromatin organization in blood granulocytes. Affected individuals show hypolobulated neutrophil nuclei with coarse chromatin. Presumed homozygous individuals have ovoid neutrophil nuclei, as well as varying degrees of developmental delay, epilepsy and skeletal abnormalities. Homozygous offspring in an extinct rabbit lineage showed severe chondrodystrophy, developmental anomalies and increased pre- and postnatal mortality. Here we show, by carrying out a genome-wide linkage scan, that PHA is linked to chromosome 1q41-43. We identified four splice-site, two frameshift and two nonsense mutations in LBR, encoding the lamin B receptor. The lamin B receptor (LBR), a member of the sterol reductase family, is evolutionarily conserved and integral to the inner nuclear membrane; it targets heterochromatin and lamins to the nuclear membrane. Lymphoblastoid cells from heterozygous individuals affected with PHA show reduced expression of the lamin B receptor, and cells homozygous with respect to PHA contain only trace amounts of it. We found that expression of the lamin B receptor affects neutrophil nuclear shape and chromatin distribution in a dose-dependent manner. Our findings have implications for understanding nuclear envelope-heterochromatin interactions, the pathogenesis of Pelger-like conditions in leukemia, infection and toxic drug reactions, and the evolution of neutrophil nuclear shape. PMID- 12118251 TI - Digenic inheritance of severe insulin resistance in a human pedigree. AB - Impaired insulin action is a key feature of type 2 diabetes and is also found, to a more extreme degree, in familial syndromes of insulin resistance. Although inherited susceptibility to insulin resistance may involve the interplay of several genetic loci, no clear examples of interactions among genes have yet been reported. Here we describe a family in which five individuals with severe insulin resistance, but no unaffected family members, were doubly [corrected] heterozygous with respect to frameshift/premature stop mutations in two unlinked genes, PPARG and PPP1R3A these encode peroxisome proliferator activated receptor gamma, which is highly expressed in adipocytes, and protein phosphatase 1, regulatory subunit 3, the muscle-specific regulatory subunit of protein phosphatase 1, which are centrally involved in the regulation of carbohydrate and lipid metabolism, respectively. That mutant molecules primarily involved in either carbohydrate or lipid metabolism can combine to produce a phenotype of extreme insulin resistance provides a model of interactions among genes that may underlie common human metabolic disorders such as type 2 diabetes. PMID- 12118252 TI - Cblb is a major susceptibility gene for rat type 1 diabetes mellitus. AB - The autoimmune disease type 1 diabetes mellitus (insulin-dependent diabetes mellitus, IDDM) has a multifactorial etiology. So far, the major histocompatibility complex (MHC) is the only major susceptibility locus that has been identified for this disease and its animal models. The Komeda diabetes-prone (KDP) rat is a spontaneous animal model of human type 1 diabetes in which the major susceptibility locus Iddm/kdp1 accounts, in combination with MHC, for most of the genetic predisposition to diabetes. Here we report the positional cloning of Iddm/kdp1 and identify a nonsense mutation in Cblb, a member of the Cbl/Sli family of ubiquitin-protein ligases. Lymphocytes of the KDP rat infiltrate into pancreatic islets and several tissues including thyroid gland and kidney, indicating autoimmunity. Similar findings in Cblb-deficient mice are caused by enhanced T-cell activation. Transgenic complementation with wildtype Cblb significantly suppresses development of the KDP phenotype. Thus, Cblb functions as a negative regulator of autoimmunity and Cblb is a major susceptibility gene for type 1 diabetes in the rat. Impairment of the Cblb signaling pathway may contribute to human autoimmune diseases, including type 1 diabetes. PMID- 12118253 TI - Paxillin binds schwannomin and regulates its density-dependent localization and effect on cell morphology. AB - Neurofibromatosis type 2 is an autosomal dominant disorder characterized by tumors, predominantly schwannomas, in the nervous system. It is caused by mutations in the gene NF2, encoding the growth regulator schwannomin (also known as merlin). Mutations occur throughout the 17-exon gene, with most resulting in protein truncation and undetectable amounts of schwannomin protein. Pathogenic mutations that result in production of defective schwannomin include in-frame deletions of exon 2 and three independent missense mutations within this same exon. Mice with conditional deletion of exon 2 in Schwann cells develop schwannomas, which confirms the crucial nature of exon 2 for growth control. Here we report that the molecular adaptor paxillin binds directly to schwannomin at residues 50-70, which are encoded by exon 2. This interaction mediates the membrane localization of schwannomin to the plasma membrane, where it associates with beta 1 integrin and erbB2. It defines a pathogenic mechanism for the development of NF2 in humans with mutations in exon 2 of NF2. PMID- 12118254 TI - Genome-wide scan for familial nasopharyngeal carcinoma reveals evidence of linkage to chromosome 4. AB - Nasopharyngeal carcinoma (NPC) occurs with high frequency in Asian populations, especially among people of Cantonese ancestry. In areas with high incidence, NPC clusters in families, which suggests that both geography and genetics may influence disease risk. Although the HLA-Bw46 locus is associated with increased risk of NPC, no predisposing genes have been identified so far. Here we report the results of a genome-wide search carried out in families at high risk of NPC from Guangdong Province, China. Parametric analyses provide evidence of linkage to the D4S405 marker on chromosome 4 with a logarithm of odds for linkage (lod) score of 3.06 and a heterogeneity-adjusted lod (hlod) score of 3.21. Fine mapping with additional markers flanking D4S405 resulted in a lod score of 3.54 and hlod score of 3.67 for the region 4p15.1-q12. Multipoint nonparametric linkage analysis gives lod scores of 3.54 at D4S405 (P = 5.4 x 10(-5)) and 4.2 at D4S3002 (P = 1.1 x 10(-5)), which is positioned 4.5 cM away from D4S405. When Epstein Barr virus antibody titer was included as a covariate, the lod scores reached 4.70 (P = 2.0 x 10(-5)) and 5.36 (P = 4.36 x 10(-6)) for D4S405 and D4S3002, respectively. Our findings provide evidence of a major susceptibility locus for NPC on chromosome 4 in a subset of families. PMID- 12118255 TI - Identification of the gene (BBS1) most commonly involved in Bardet-Biedl syndrome, a complex human obesity syndrome. AB - Bardet-Biedl syndrome (BBS, OMIM 209900) is a genetic disorder with the primary features of obesity, pigmentary retinopathy, polydactyly, renal malformations, mental retardation and hypogenitalism. Individuals with BBS are also at increased risk for diabetes mellitus, hypertension and congenital heart disease. What was once thought to be a homogeneous autosomal recessive disorder is now known to map to at least six loci: 11q13 (BBS1), 16q21 (BBS2), 3p13 p12 (BBS3), 15q22.3 q23 (BBS4), 2q31 (BBS5) and 20p12 (BBS6). There has been considerable interest in identifying the genes that underlie BBS, because some components of the phenotype are common. Cases of BBS mapping ro BBS6 are caused by mutations in MKKS; mutations in this gene also cause McKusick-Kaufman syndrome (hydrometrocolpos, post-axial polydactyly and congenital heart defects). In addition, we recently used positional cloning to identify the genes underlying BBS2 (ref. 16) and BBS4 (ref. 17). The BBS6 protein has similarity to a Thermoplasma acidophilum chaperonin, whereas BBS2 and BBS4 have no significant similarity to chaperonins. It has recently been suggested that three mutated alleles (two at one locus, and a third at a second locus) may be required for manifestation of BBS (triallelic inheritance). Here we report the identification of the gene BBS1 and show that a missense mutation of this gene is a frequent cause of BBS. In addition, we provide data showing that this common mutation is not involved in triallelic inheritance. PMID- 12118256 TI - Single granule cells reliably discharge targets in the hippocampal CA3 network in vivo. AB - Processing of neuronal information depends on interactions between the anatomical connectivity and cellular properties of single cells. We examined how these computational building blocks work together in the intact rat hippocampus. Single spikes in dentate granule cells, controlled intracellularly, generally failed to discharge either interneurons or CA3 pyramidal cells. In contrast, trains of spikes effectively discharged both CA3 cell types. Increasing the discharge rate of the granule cell increased the discharge probability of its target neuron and decreased the delay between the onset of a granule cell train and evoked firing in postsynaptic targets. Thus, we conclude that the granule cell to CA3 synapses are 'conditional detonators,' dependent on granule cell firing pattern. In addition, we suggest that information in single granule cells is converted into a temporal delay code in target CA3 pyramidal cells and interneurons. These data demonstrate how a neural circuit of the CNS may process information. PMID- 12118257 TI - Hormonally regulated alpha(4)beta(2)delta GABA(A) receptors are a target for alcohol. AB - Here we report that low concentrations of alcohol (1-3 mM) increased Cl(-) currents gated by a recombinant GABA(A) receptor, alpha(4)beta(2)delta, by 40-50% in Xenopus laevis oocytes. We also found greater hippocampal expression of receptors containing alpha(4) and delta subunits, using a rat model of premenstrual syndrome (PMS) in which 1-3 mM alcohol preferentially enhanced GABA gated currents, and low doses of alcohol attenuated anxiety and behavioral reactivity. The alcohol sensitivity of delta-containing receptors may underlie the reinforcing effects of alcohol during PMS, when eye saccade responses to low doses of alcohol are increased. PMID- 12118258 TI - Control of photoreceptor axon target choice by transcriptional repression of Runt. AB - Drosophila photoreceptor neurons (R cells) project their axons to one of two layers in the optic lobe, the lamina or the medulla. The transcription factor Runt (Run) is normally expressed in the two inner R cells (R7 and R8) that project their axons to the medulla. Here we examine the relationship between Run and the ubiquitously expressed nuclear protein Brakeless (Bks), which has previously been shown to be important for axon termination in the lamina. We report that Bks represses Run in two of the outer R cells: R2 and R5. Expression of Run in R2 and R5 causes axonal mistargeting of all six outer R cells (R1-R6) to the inappropriate layer, without altering expression of cell-specific developmental markers. PMID- 12118259 TI - Pharmacological upregulation of h-channels reduces the excitability of pyramidal neuron dendrites. AB - The dendrites of pyramidal neurons have markedly different electrical properties from those of the soma, owing to the non-uniform distribution of voltage-gated ion channels in dendrites. It is thus possible that drugs acting on ion channels might preferentially alter dendritic, but not somatic, excitability. Using dendritic and somatic whole-cell and cell-attached recordings in rat hippocampal slices, we found that the anticonvulsant lamotrigine selectively reduced action potential firing from dendritic depolarization, while minimally affecting firing at the soma. This regional and input-specific effect resulted from an increase in the hyperpolarization-activated cation current (I(h)), a voltage-gated current present predominantly in dendrites. These results demonstrate that neuronal excitability can be altered by drugs acting selectively on dendrites, and suggest an important role for I(h) in controlling dendritic excitability and epileptogenesis. PMID- 12118261 TI - Toxicology and carcinogenesis studies of p-nitrotoluene (CAS no. 99-99-0) in F344/N rats and B6C3F(1) mice (feed studies). AB - p-Nitrotoluene is used to synthesize agricultural and rubber chemicals, azo and sulfur dyes, and dyes for cotton, wool, silk, leather, and paper. p-Nitrotoluene was nominated by the National Institute for Occupational Safety and Health and the NTP for study based on its considerable human exposure as well as the absence of long-term studies of its carcinogenicity in rodents. Male and female F344/N rats and B6C3F1 mice were exposed to p-nitrotoluene (greater than 99% pure) in feed for 2 years. Genetic toxicology studies were conducted in Salmonella typhimurium, L5178Y mouse lymphoma cells, cultured Chinese hamster ovary cells, and rat and mouse bone marrow cells. 2-YEAR STUDY IN RATS: Groups of 50 male and 50 female rats were fed diets containing 0, 1,250, 2,500, or 5,000 ppm p nitrotoluene (equivalent to average daily doses of approximately 55, 110, or 240 mg p-nitrotoluene/kg body weight to males and 60, 125, or 265 mg/kg to females) for 105 or 106 weeks. Survival, Body Weights, and Feed Consumption: Survival of all exposed groups of rats was similar to that of the control groups. Mean body weights of 5,000 ppm male and 2,500 and 5,000 ppm female rats were less than those of the controls during most of the study; mean body weights of 1,250 ppm females were less during the second year of the study. Feed consumption by 5,000 ppm females was less than that by the controls during year 2 of the study. Biomarkers of Exposure: Two urinary metabolites were followed during the study as biomarkers of exposure. The ratios of p-nitrobenzoic acid to creatinine and of p acetamidobenzoic acid to creatinine determined at 2 weeks and at 3, 12, and 18 months were linearly related to exposure concentration in males and females. Pathology Findings: The incidence of clitoral gland adenoma or carcinoma (combined) was significantly greater in 2,500 ppm females than that in the controls and exceeded the historical control ranges. The incidence of clitoral gland neoplasms was not increased in 5,000 ppm females, possibly because of the lower body weights in this group. The incidences of subcutaneous fibroma and of subcutaneous fibroma or fibrosarcoma (combined) in 2,500 ppm male rats were significantly increased and exceeded the historical control ranges. The incidences of several nonneoplastic kidney lesions were significantly increased in exposed groups of rats, and the severities of these lesions generally increased with increasing exposure concentration. In the spleen, incidences of hematopoietic cell proliferation and pigmentation were significantly increased in the 2,500 and 5,000 ppm groups. Significantly increased incidences of various types of altered cell foci in the liver of males and females were associated with exposure. Incidences of germinal epithelial atrophy of the testis in 5,000 ppm males and endometrial cystic hyperplasia of the uterus in 2,500 and 5,000 ppm females were significantly increased. The incidences of mononuclear cell leukemia were significantly decreased in all exposed groups except 1,250 ppm females. The incidence of interstitial cell adenoma of the testis in 5,000 ppm males was significantly decreased. 2-YEAR STUDY IN MICE: Groups of 50 male and 50 female mice were fed diets containing 0, 1,250, 2,500, or 5,000 ppm p-nitrotoluene (equivalent to average daily doses of approximately 170, 345, or 690 mg/kg to males and 155, 315, or 660 mg/kg to females) for 105 or 106 weeks. Survival, Body Weights, and Feed Consumption: Survival of all exposed groups of male and female mice was similar to that of the control groups. Mean body weights of 5,000 ppm males and females were less than those of the control groups during most of the study. Mean body weights of 2,500 ppm males were less than those of the controls after week 92. Feed consumption by all exposed groups of mice was similar to that by the control groups. Pathology Findings: The incidence of alveolar/bronchiolar adenoma or carcinoma (combined) was significantly greater in 5,000 ppm male mice than in the controls, as was the incidence of alveolar epithelial hyperplasia in this group. The incidences of alveolar epithelial bronchiolization were significantly increased in all exposed groups of males and females. GENETIC TOXICOLOGY: p-Nitrotoluene was not mutagenic in any of several strains of S. typhimurium, with or without metabolic activation enzymes (S9). A positive response to p-nitrotoluene was observed in the L5178Y mouse lymphoma cell assay in trials with S9. Significantly increased sister chromatid exchange frequencies were observed in cultured Chinese hamster ovary cells treated with p-nitrotoluene with and without S9. Chromosomal aberrations were also induced in Chinese hamster ovary cells treated with p-nitrotoluene in the presence of S9; no increased aberrations were seen without S9. p-Nitrotoluene did not induce a significant reproducible increase in the frequency of micronuclei in bone marrow polychromatic erythrocytes of male rats or male mice when administered by intraperitoneal injection. CONCLUSIONS: Under the conditions of these 2-year feed studies there was equivocal evidence of carcinogenic activity* of p-nitrotoluene in male F344/N rats based on increased incidences of subcutaneous skin neoplasms. There was some evidence of carcinogenic activity of p-nitrotoluene in female F344/N rats based on increased incidences of clitoral gland neoplasms. There was equivocal evidence those of the control groups during most of the study. Mean body weights of 2,500 ppm males were less than those of the controls after week 92. Feed consumption by all exposed groups of mice was similar to that by the control groups. Pathology Findings: The incidence of alveolar/bronchiolar adenoma or carcinoma (combined) was significantly greater in 5,000 ppm male mice than in the controls, as was the incidence of alveolar epithelial hyperplasia in this group. The incidences of alveolar epithelial bronchiolization were significantly increased in all exposed groups of males and females. GENETIC TOXICOLOGY: p Nitrotoluene was not mutagenic in any of several strains of S. typhimurium, with or without metabolic activation enzymes (S9). A positive response to p nitrotoluene was observed in the L5178Y mouse lymphoma cell assay in trials with S9. Significantly increased sister chromatid exchange frequencies were observed in cultured Chinese hamster ovary cells treated with p-nitrotoluene with and without S9. Chromosomal aberrations were also induced in Chinese hamster ovary cells treated with p-nitrotoluene in the presence of S9; no increased aberrations were seen without S9. p-Nitrotoluene did not induce a significant reproducible increase in the frequency of micronuclei in bone marrow polychromatic erythrocytes of male rats or male mice when administered by intraperitoneal injection. CONCLUSIONS: Under the conditions of these 2-year feed studies there was equivocal evidence of carcinogenic activity of p-nitrotoluene in male F344/N rats based on increased incidences of subcutaneous skin neoplasms. There was some evidence of carcinogenic activity of p-nitrotoluene in female F344/N rats based on increased incidences of clitoral gland neoplasms. There was equivocal evidence in male and female rats, testis in male rats, and lung in male and female mice. Decreased incidences of mononuclear cell leukemia in male and female rats and testicular interstitial cell adenoma in male rats were attributed to exposure to p-nitrotoluene. PMID- 12118260 TI - Conversion of cerebral cortex into basal ganglia in Emx2(-/-) Pax6(Sey/Sey) double-mutant mice. AB - The molecular mechanisms that activate morphogenesis of cerebral cortex are currently the subject of intensive experimental analysis. Transcription factor genes of the homeobox, basic helix-loop-helix (bHLH) and zinc-finger families have recently been shown to have essential roles in this process. However, the actual selector genes activating corticogenesis have not yet been identified. Here we show that high-level expression of at least one functional allele of either of the homeobox genes Emx2 or Pax6 in the dorsal telencephalon is necessary and sufficient to stably activate morphogenesis of cerebral cortex and to repress that of adjacent structures, such as striatum. PMID- 12118262 TI - NTP Technical Report on the toxicity studies of trans-1,2-dichloroethylene (CAS no. 156-60-5) administered in microcapsules in feed to F344/N rats and B6C3F(1) mice. AB - 1,2-Dichloroethylene exists in two isomeric states: trans-1,2-dichloroethylene and cis-1,2-dichloroethylene. The trans isomer is used more widely in industry than the cis isomer. trans-1,2-Dichloroethylene is used as a solvent for waxes, resins, and acetylcellulose. It is also used in the extraction of rubber, as a refrigerant, and in the manufacture of pharmaceuticals and artificial pearls. F344/N rats and B6C3F1 mice were administered trans-1,2-dichloroethylene in microcapsules in feed for 14 weeks. Animals were evaluated for clinical pathology, reproductive system effects, and histopathology. Genetic toxicity studies were conducted in vitro in Salmonella typhimurium and Chinese hamster ovary (CHO) cells, and in vivo in mouse bone marrow cells and peripheral blood erythrocytes. In the 14-week feed studies, groups of 10 male and 10 female rats and mice were fed diets containing microcapsules with a chemical load of 45% trans-1,2-dichloroethylene. Dietary concentrations of 3,125, 6,250, 12,500, 25,000, and 50,000 ppm microencapsulated trans-1,2-dichloroethylene resulted in average daily doses of 190, 380, 770, 1,540, and 3,210 mg/kg for male rats; 190, 395, 780, 1,580, and 3,245 mg/kg for female rats; 480, 920, 1,900, 3,850, and 8,065 mg/kg for male mice; and 450, 915, 1,830, 3,760, and 7,925 mg/kg for female mice. Additional groups of 10 male and 10 female rats and mice served as untreated and vehicle controls. There were no exposure-related deaths of rats or mice. Mean body weights of male rats and male and female mice in the 50,000 ppm groups were significantly less than those of the vehicle controls. The mean body weight gains of female mice in the 12,500 and 25,000 ppm groups were also significantly less than that of the vehicle controls. On day 21 and at week 14, there were mild decreases in hematocrit values, hemoglobin concentrations, and erythrocyte counts in groups of male and female rats in the 25,000 and 50,000 ppm groups. At week 14, these effects were seen in male rats exposed to 6,250 and 12,500 ppm. There were no exposure-related alterations in clinical chemistry parameters in rats or mice. The liver weights of female rats exposed to 6,250 ppm or greater were significantly greater than those of the vehicle controls. The absolute kidney weights of male rats exposed to 25,000 or 50,000 ppm were significantly decreased. No gross or microscopic lesions were observed in rats or mice that could be attributed to trans-1,2-dichloroethylene exposure. Neither cis , trans-, nor cis,trans-1,2-dichloroethylene was mutagenic in S. typhimurium strain TA97 (cis isomer only), TA98, TA100, TA1535, or TA1537, with or without S9 metabolic activation enzymes. In CHO cells in vitro, cis- 1,2-dichloroethylene induced sister chromatid exchanges (SCEs) in the absence of S9; with S9, the single trial that was performed yielded equivocal results. The cis,trans isomer induced significant increases in SCEs in cultured CHO cells with and without S9. In contrast to these positive results, trans-1,2-dichloroethylene gave negative results in the SCE test, with and without S9. Neither cis-, trans-, nor cis,trans 1,2-dichloroethylene induced chromosomal aberrations (Abs) in cultured CHO cells, with or without S9. In vivo, no induction of SCEs or Abs was noted in bone marrow cells of male mice administered cis- or trans-1,2-dichloroethylene by intraperitoneal injection once, with sampling performed 23 hours (for SCE analyses) or 17 hours (for Abs analyses) after injection. In addition, negative results were obtained in a peripheral blood micronucleus test in male and female mice administered trans- 1,2-dichloroethylene in microcapsules in feed for 14 weeks. Very little toxicity was associated with ingestion of microencapsulated trans-1-2-dichloroethylene. Histopathology and clinical chemistry data, combined with body and organ weight data, revealed that the maximum tolerated dose was not reached in these studies. PMID- 12118263 TI - NTP Comparative Toxicity and Carcinogenicity Studies of o-Nitrotoluene and o Toluidine Hydrochloride (CAS Nos. 88-72-2 and 636-21-5) Administered in Feed to Male F344/N Rats. AB - o-Nitrotoluene and o-toluidine hydrochloride are structurally related chemicals that are suspected and demonstrated animal carcinogens, respectively. The metabolic potential of the gastrointestinal flora is considered an important factor in o-nitrotoluene-induced toxicity and involves the reduction of the nitro group to the corresponding amine (forming o-toluidine). These studies were designed to 1) compare the target organ toxicities of o-nitrotoluene and o toluidine hydrochloride administered in feed at approximately equimolar doses (5,000 ppm) to male F344/N rats for 13 or 26 weeks, 2) determine the potential progression or reversibility of toxic or proliferative lesions following chemical withdrawal (stop-exposure) for 13 weeks after 13 weeks of exposure, and 3) examine the effect of antibiotic-altered gastrointestinal flora on the toxicity and/or carcinogenicity of o-nitrotoluene. o-Nitrotoluene and o-toluidine hydrochloride caused mesothelial hyperplasia and mesothelioma in male rats after 13 or 26 weeks of dietary exposure. The incidence of mesothelioma was greater and the latency was less in rats administered o-nitrotoluene than in rats administered o-toluidine hydrochloride. Additionally, o-nitrotoluene caused testicular degeneration in rats. Effects of o-nitrotoluene administration in the liver included progressive, irreversible increases in liver weight and irreversible increases in the incidences of cytoplasmic vacuolization and oval cell hyperplasia. Placental glutathione S-transferase (PGST)-positive foci of cellular alteration occurred in the liver after 13 weeks of o-nitrotoluene exposure, and the number and size (as reflected by the volume fraction) of foci were increased after 26 weeks of continuous exposure. During the recovery period, the number of PGST-positive foci in rats in the stop-exposure group decreased slightly, but the size of the foci continued to increase. After 26 weeks, cholangiocarcinoma occurred in 2 of 20 rats in the stop-exposure group and 1 of 20 rats in the continuous-exposure group administered o-nitrotoluene. In contrast, liver effects in rats administered o-toluidine hydrochloride consisted of minimal hemosiderin accumulation in Kupffer cells; the incidence of this lesion in the stop-exposure group decreased during the recovery period. o Toluidine hydrochloride caused fewer and much smaller PGST-positive foci than those caused by o-nitrotoluene. o-Nitrotoluene caused an accumulation of hyaline droplets in the renal tubule epithelium; this accumulation did not increase in severity with continued exposure and completely regressed during the recovery period. Exposure to o-toluidine hydrochloride did not cause hyaline droplet accumulation but did cause an accumulation of hemosiderin pigment in renal tubule epithelium. This change progressed in severity during the 26-week continuous exposure study but decreased in severity in the stop-exposure study during the recovery period. Exposure to o-nitrotoluene or o-toluidine hydrochloride caused increased incidences of hematopoiesis, hemosiderin accumulation, and capsular fibrosis in the spleen. In rats administered o-toluidine, spleen effects were much more prominent and were also reflected by congestion and markedly increased spleen weights. During the recovery period for the stop-exposure groups administered either compound, incidences of hemosiderin accumulation and hematopoiesis were decreased, but the capsular fibrosis did not resolve. Hyperplasia of the transitional epithelium in the urinary bladder was observed only in rats administered o-toluidine hydrochloride; this lesion did not increase in severity after 26 weeks of continuous exposure and completely regressed in the stop-exposure group during the recovery period. Alteration of the gastrointestinal flora by daily gavage administration of antibiotics did not affect the pattern or severity of toxicity at any site or the development of mesothelioma in rats exposed to o-nitrotoluene, although cholangiocarcinomas that occurred in three rats with the normal flora did not occur in groups with the altered intestinal flora. Subsequent studies determined that the antibiotic regimen used was effective only in reducing the gut population of aerobic microorganisms and had little effect on obligate anaerobes, which are thought to play a mayor role in nitro group reduction. In summary, these studies confirmed the target organs and compared the relative toxicity for o-nitrotoluene and o toluidine hydrochloride administered to male F344/N rats at equimolar concentrations in feed. With the exception of the spleen toxicity observed with each chemical, but more prominently with o-toluidine hydrochloride, morphologic effects of exposure to each of these chemicals in the testis/epididymis, liver, kidney, and urinary bladder were different. The results of these studies demonstrate the somewhat greater relative carcinogenic potential of o nitrotoluene compared to o-toluidine hydrochloride after 13 or 26 weeks of administration based on the occurrence of mesothelioma and cholangiocarcinoma. The apparently lower potency of o-toluidine hydrochloride relative to o-o nitrotoluene in the induction of mesothelioma suggests that simple intestinal reduction of the nitro group may not be sufficient for carcinogenic activity in the mesothelium. PMID- 12118264 TI - Preoperative serum levels of CA 72-4, CEA, CA 19-9, and alpha-fetoprotein in patients with gastric cancer. AB - INTRODUCTION: The clinical importance of preoperative serum levels of CA 72-4, carcinoembryonic antigen (CEA), CA 19-9, and alpha-fetoprotein (AFP) was prospectively evaluated in 44 patients with gastric cancer. METHOD: The serum tumor marker levels were determined by commercial radioimmunoassay kits. Positivity for CA 72-4 (>4 U/mL), CEA (>5 ng/mL), CA 19-9 (>37 U/mL), and AFP (>10 ng/mL) were correlated according to the stage, histology, and lymph node metastasis. RESULTS AND DISCUSSION: CA 72-4 showed a higher positivity rate for gastric cancer (47.7%) than CEA (25%), CA 19-9 (25%), and AFP (0%). The combination of CA 72-4 with CEA and CA 19-9 increased the sensitivity to 61.4%. The positivity rates of CA 72-4 in patients at stages I and II (initial disease) and in patients at stages III and IV (advanced disease) were 9% and 60.6%, respectively (P < 0.005). No correlation was found between CEA and CA 19-9 levels and the stage of gastric cancer. There was a tendency of positivity for CA 72-4 to suggest lymph node involvement, but it was not significant (P = 0.075). Serum levels of tumor markers did not show a correlation with the histological types of gastric cancer. CONCLUSION: Preoperative serum levels of CA 72-4 provided a predictive value in indicating advanced gastric cancer. PMID- 12118265 TI - Tissue expander complications in plastic surgery: a 10-year experience. AB - INTRODUCTION: Tissue expanders have been of great value in plastic surgery. Tissue expansion was developed for a specific indication; however, within a very short time, the concept of tissue expansion found wide applicability. From 1990 to 1999, 315 expanders in 164 patients were utilized. A retrospective analysis of complications and prognostic factors for complications were done. METHODS: The indications for tissue expansion were burns (50%), trauma (32%), and sequelae of previous surgery (8.8%). The expanders were inserted most frequently in the scalp, trunk and neck. RESULTS: There were 22.2% of complications and the most common were expander exposure (50%), infection (24%) and bad function of the expander (12.8%). The present study revealed an increased rate of minor complications in the group of 0 to 10 years of age and an increased rate of major complications for face and neck expansions compared to trunk expansion. There were no increased complication rates for the other age and anatomic site groups, previous expansion, concomitant expansion and type of expander used. CONCLUSIONS: The outcomes from tissue expansion procedures done in our hospital are similar to those reported in the literature. Tissue expansion is a good and safe technique. PMID- 12118266 TI - Evaluation of efficacy, reliability, and tolerability of sibutramine in obese patients, with an echocardiographic study. AB - This is a double-blind, placebo-controlled study of the efficacy, safety, and tolerability of sibutramine in the management of obese patients for a 6-month period. METHOD: Sixty-one obese patients (BMI >30, <40 kg/m2), aged 18-65 years were evaluated. In the first phase of the study (30 days), the patients were given a placebo. We monitored compliance with a low-calorie diet (1200 kcal/day) and to the placebo. In the next stage, the double-blind phase (6 months), we compared placebo and sibutramine (10 mg/day). The criteria for evaluating efficacy were weight loss, reduction in body mass index (BMI), and abdominal and hip circumferences. Tolerability was assessed based on reported side effects, variation in arterial blood pressure and heart rate, metabolic profile (fasting glucose, total cholesterol and its fractions, and triglycerides), laboratory tests (renal and hepatic functions), and flow Doppler echocardiogram. RESULTS: We observed a greater weight loss (7.3 kg, 8% vs 2.6 kg, 2.8%) and a reduction in body mass index (7.4% vs 2.1%) in the sibutramine group than in the placebo group. Classifying the patients into 4 subgroups according to weight loss (weight gain, loss <5%, loss of 5% to 9.9%, and loss >10%), we observed a weight loss of >5% in 40% of the patients on sibutramine compared with 12.9% in the placebo group. We also detected weight gain in 45.2% of the placebo group compared to 20% in the sibutramine group. The sibutramine group showed improvement in HDL- cholesterol values (increased by 17%) and triglyceride values (decreased by 12.8%). This group also showed an increase in systolic blood pressure (6.7%, 5 mmHg). There were no changes in echocardiograms comparing the beginning and end of follow-up, and side effects did not lead to discontinuation of treatment. DISCUSSION: Sibutramine proved to be effective for weight loss providing an 8% loss of the initial weight. Compliance to prolonged treatment was good, and side effects did not result in discontinuation of treatment. These data confirmed the good efficacy, tolerability, and safety profiles of sibutramine for treatment of obesity. PMID- 12118267 TI - Appraisal of surgical treatment of 47 cases of patellofemoral instability. AB - INTRODUCTION: Patellofemoral instability is a common knee disease. Its etiology is complex and variable, with many components making different contributions in each individual, resulting in several distinct clinical presentations. Our goal was to analyze the results of surgical treatment in our hospital over a period of 10 years. PATIENTS AND METHODS: We analyzed 55 knees of 47 patients who underwent surgery for patellofemoral instability and were classified into 2 main groups: proximal realignment and combined proximal and distal realignment. Three other groups were analyzed according to the duration of preoperative symptoms: less than 1 year (group I); 1 to 10 years (group II); and more than 10 years (group III). RESULTS: There were 62% good results overall, with 78% good results in groups I and II. Group III had 81% bad results, showing that a late diagnosis of advanced disease results in a poor prognosis. In addition to late diagnosis, bad results were usually associated with incorrect diagnosis or choice of surgical technique. There was no significant difference between isolated proximal realignment and combined proximal and distal realignment in groups I or II, but in group III, the combined technique yielded better results. DISCUSSION: Our results indicate that patellofemoral instability should be addressed in its early stages. Patients with long-lasting symptoms or more severe disease seem to achieve better results with combined techniques. CONCLUSION: Proximal and distal realignments produce better results than isolated proximal realignment in patients with joint degeneration or with greater duration of disease. The realignment surgery does not produce good results in patients with advanced disease. PMID- 12118268 TI - Complications of tracheobronchial foreign body aspiration in children: report of 5 cases and review of the literature. AB - Foreign body aspiration (FBA) is one of leading causes of death in children, especially among those younger than 3 years of age. The inhalation of a foreign body may cause a wide variety of symptoms, and early diagnosis is highly associated with the successful removal of the inhaled foreign material. Despite the great advances in endoscopic procedures and anesthesia, a large number of difficulties and complications still result from foreign body aspiration. We describe 5 cases of serious acute complications following aspiration of foreign bodies that became lodged in the tracheobronchial tree, including pneumomediastinum, pneumothorax, total atelectasis, foreign body dislodgment, and need for thoracotomy in children admitted into our intensive care unit in 1999 and 2000; these were all situations that could have been prevented with early recognition and prompt therapeutic intervention. PMID- 12118269 TI - Hypothyroidism following struma ovarii tumor resection: a case report. AB - Struma ovarii is an infrequent ovarian tumor, and there are only few reports with detailed data of thyroid function. In several cases, malignant struma ovarii have been shown to produce hyperthyroidism, but there is no reported case of hypothyroidism following struma ovarii tumor resection. A 62-year-old white woman underwent right ovary resection that had a pathologic diagnosis of struma ovarii. After 6 days, she developed weakness, myalgia, somnolence, nausea, and arterial hypotension. Laboratory tests showed a high level of thyroid-stimulating hormone (TSH) and a decreased level thyroxin. Thyroxin replacement therapy was initiated, and the patient became completely asymptomatic. This is the first reported case of a previously asymptomatic woman who developed a definite clinical hypothyroidism after resection of a struma ovarii tumor. PMID- 12118270 TI - Pediatric cardiac postoperative care. AB - The Heart Institute of the University of Sao Paulo, Medical School is a referral center for the treatment of congenital heart diseases of neonates and infants. In the recent years, the excellent surgical results obtained in our institution may be in part due to modern anesthetic care and to postoperative care based on well structured protocols. The purpose of this article is to review unique aspects of neonate cardiovascular physiology, the impact of extracorporeal circulation on postoperative evolution, and the prescription for pharmacological support of acute cardiac dysfunction based on our cardiac unit protocols. The main causes of low cardiac output after surgical correction of heart congenital disease are reviewed, and methods of treatment and support are proposed as derived from the relevant literature and our protocols. PMID- 12118271 TI - Hypertonic/hyperoncotic solution in hypovolemic patients: experience in the emergency room. AB - Hypertonic solutions have been studied extensively in the treatment of hypovolemic shock, both in experimental and clinical models. Safety, efficacy, and long-term effects on animals and patients have been evaluated. The present article reviews indications, safety, mortality rates, and outcome in patients with hemorrhagic hypovolemic shock who were treated after admission with a hypertonic/hyperoncotic solution under strict observation in the emergency room. PMID- 12118273 TI - Ecology of sand flies (Diptera: psychodidae: phlebotominae) in the north of the state of Mato Grosso, Brazil. AB - Peixoto de Azevedo is located in the north of State of Mato Grosso, where environmental alterations led to an outbreak of American cutaneous leishmaniasis in the 80s. The parasite from patients was characterized as Leishmania (V.) braziliensis. The aim of this study is to contribute to the sand fly ecology of Central-West Brazil. Captures were carried out monthly using CDC light traps. Twenty-six species of sand fly were characterized; among which Lutzomyia (Lutzomyia) spathotrichia, L. runoides and L. (Psychodopygus) llanosmartinsi were recorded in the State of Mato Grosso for the first time. L. (Nyssomyia) whitmani, L. (N.) antunesi, L. (L.) spathotrichia, L. (P.) c. carrerai, L. (P.) complexa, L. (P.) lainsoni and L. (N.) umbratilis constituted 92.4% of the local fauna, among which L. (N.) whitmani and L. (N.) antunesi, accounting for about 53% of the fauna at the stations of capture. On the vertical distribution of sand flies on the Beira-Rio Farm, L. (N.) whitmani and L. (N.) antunesi prevailed at ground level and in the canopy, respectively, whereas on the BR-080, L. (P.) llanosmartinsi was prevalent on the ground and L. (P.) c. carrerai, in the canopy. It is suggested that L. (N.) umbratilis is the local vector. PMID- 12118272 TI - Some aspects of protozoan infections in immunocompromised patients- a review. AB - Protozoa are among the most important pathogens that can cause infections in immunocompromised hosts. These microorganisms particularly infect individuals with impaired cellular immunity, such as those with hematological neoplasias, renal or heart transplant patients, patients using high doses of corticosteroids, and patients with acquired immunodeficiency syndrome. The protozoa that most frequently cause disease in immunocompromised patients are Toxoplasma gondii, Trypanosoma cruzi, different Leishmania species, and Cryptosporidium parvum; the first two species cause severe acute meningoencephalitis and acute myocarditis, Leishmania sp. causes mucocutaneous or visceral disease, and Cryptosporidium can lead to chronic diarrhea with hepatobiliary involvement. Various serological, parasitological, histological and molecular methods for the diagnosis of these infections are currently available and early institution of specific therapy for each of these organisms is a basic measure to reduce the morbidity and mortality associated with these infections. PMID- 12118274 TI - Longitudinal Study on the Natural Infection of Biomphalaria straminea and B. glabrata by Schistosoma mansoni in an Endemic Area of Schistosomiasis in Pernambuco, Brazil. AB - The abundance of snail hosts and the rates of infection with Schistosoma mansoni were monitored monthly for four years in two representative localities subjected to repeated chemotherapy of infected persons. Snail abundance varied from 1.0 to 4.4 collected per person/minute/station for Biomphalaria straminea and from 0.1 to 7.0 for B. glabrata. Infection rates of snails in nature varied from 0% to 15% for the former and from 0% to 70% for the latter species. Human infection increased from 35.5% to 61.9% in the locality occupied by B. straminea, and decreased from 40.3% to 20.8% in that occupied by B. glabrata. No relationship could be detected between human infection and the snail variables. Despite seasonal variations, natural infection persisted throughout the monitoring period in both snail species. It reached remarkably high levels in B. straminea when compared to those obtained by other authors probably because of differences in methodology. It is recommended that longitudinal studies should be carried out focally and periodically to avoid underestimating the prevalence of schistosome infection in snails. PMID- 12118275 TI - Risk zones of human Leishmaniases in the Western Mediterranean basin: correlations between vector sand flies, bioclimatology and phytosociology. AB - Correspondence analysis was applied to sand fly sampling in 865 stations from the Western Mediterranean basin. The position of each of 24 species was determined with respect to the bioclimatic belts. Thus, the multidimensional analyses manifest clear correlations between bioclimatic belts and their expression in the area, the phytosociological groupings, and vector species of visceral and cutaneous leishmaniases. The transfer of these data to usual maps allows to delimit the geographical distribution of these diseases in the Western Mediterranean basin and contributes to the determination, in a rational manner, of the high risk zones. PMID- 12118277 TI - Chagas disease in dogs from endemic areas of Costa Rica. AB - Dogs with the presumptive diagnosis of Chagas disease are commonly sent to our School of Veterinary Medicine by independent veterinarians. This prompted us to evaluate the prevalence of canine trypanosomiasis in some villages of the Central Valley of Costa Rica. A total of 54 dogs (21 males and 33 females) from five rural villages, with ages between 3 months and 10 years old, were bled and submitted to three serological tests: indirect immunofluorescence, indirect hemagglutination and ELISA. Among all animals, 15 (27.7%) revealed antibodies (6 pure bred and 9 mongrels) and in 3 of them the parasite was also demonstrated by xenodiagnosis. All positive animals except 1, and 9 negative animals (control group) were examined by X-rays and electrocardiography, revealing different degrees of cardiomegaly and ECG alteration, consistent with Chagas disease pathology in one dog (SA-11) of the infected ones. Examination of 50 inhabitants living in the houses where dogs and Triatoma dimidiata were found, yielded negative serological reactions. This was assumed to support the hypothesis that dogs are commonly infected by the oral route, a more effective means of infection compared with the vector transmission mechanism that occurs in humans. PMID- 12118276 TI - IgM-immunofluorescence test as a diagnostic tool for epidemiologic studies of Schistosomiasis in low endemic areas. AB - The high sensitivity and the ability to diagnose schistosomiasis in a very early phase after infection have indicated the detection of IgM antibodies to Schistosoma mansoni gut antigens by the immunofluorescence test (IgM-IFT) as a useful serological test for epidemiological studies in low endemic areas. When applied in a follow-up study for two years, higher rates of seroconversion from IFT negative to positive were observed during the summer months, suggesting seasonal transmission of schistosomiasis in the rural area of the municipality of Itariri (Sao Paulo, Brazil). In each survey, blood samples from about 600 schoolchildren were collected on filter paper and submitted to IgM-IFT. When the blood samples were classified for the IgM antibody levels, according to the intensity of fluorescent reaction observed at fluorescence microscopy, and correlated to the egg counts in the Kato-Katz positive patients, no association was observed. This observation might suggest that the intensity of fluorescence observed in the IgM-IFT, as an indicator of IgM antibody levels, could not be an useful seroepidemiological marker for classifying areas of low endemicity according to degrees of infection. PMID- 12118278 TI - [Redescription of Litomosoides brasiliensis Almeida 1936 (Nematoda: Filariiidae) from a new host Anoura caudifera (Chiroptera: Phyllostomidae)]. AB - The study of the surface topography added details regarding the disposition of male caudal papillae, spicules and area rugosa apart from vulva and oral aperture. The occurrence of this nematode in the state of Amapa represents a new geographical distribution. PMID- 12118279 TI - Description of Pintomyia (Pifanomyia) Falcaorum sp. n. (Diptera: Psychodidae: Phlebotominae), a fossil sand fly from dominican amber. AB - A new species of sand fly, Pintomyia (Pifanomyia) falcaorum is described from an amber originated from the northern mountain range of Dominican Republic. The male sand fly specimen is well preserved and most features used in Phlebotominae taxonomy are seen with remarkable clarity. PMID- 12118280 TI - Plesiophysa dolichomastix sp. n. (Gastropoda: Planorbidae). AB - A new species of planorbid mollusc, Plesiophysa dolichomastix (Greek dolichos = long, mastix = flagellum), collected from Lagoa da Pedra, municipality of Santa Rosa, state of Goias, Brazil (15 degrees 01'S, 47 degrees 13'W) is described. It is indistinguishable by the shell characters from the five congeneric species described so far: P. striata (Orbigny, 1841), P. granulata ("Shuttleworth" Sowerby, 1873), P. guadeloupensis ("Fischer" Maze, 1883), P. ornata (Haas, 1938) and P. hubendicki Richards & Ferguson, 1962. It differs from the anatomically studied species in the following characters: about 50 ovotestis diverticula, against 12 in granulata, 100 in ornata, unstated in hubendicki; and length of flagella - about as long as the penial complex -, against about 1/3 to 1/6 in the other three. PMID- 12118281 TI - Capillariidae eggs found in the urine of a free ranging maned wolf from Argentina. AB - The first finding of a Capillariid in the urinary tract of a free ranging maned wolf (Chrysocyon brachyurus) is described. The individual was an adult male attacked by dogs in the locality of Cayastacito (Santa Fe, Argentina, 31 degrees 05' S, 60 degrees 34' W). Eggs found in urine measured 64.6-66.9 micrometer (mean 65.4 micrometer) x 26.9-31 micrometer (mean 29 micrometer). Further studies are needed to determine whether this finding corresponds to a new Capillariid species, related to C. brachyurus, or it is an already described species that has been introduced by domestic dogs. PMID- 12118282 TI - The Zymovars of Vibrio cholerae: multilocus enzyme electrophoresis of Vibrio cholerae. AB - Zymovars analysis also known as multilocus enzyme electrophoresis is applied here to investigate the genetic variation of Vibrio cholerae strains and characterise strains or group of strains of medical and epidemiological interest. Fourteen loci were analyzed in 171 strains of non-O1 non-O139, 32 classical and 61 El Tor from America, Africa, Europe and Asia. The mean genetic diversity was 0.339. It is shown that the same O antigen (both O1 and non-O1) may be present in several genetically diverse (different zymovars) strains. Conversely the same zymovar may contain more than one serogroup. It is confirmed that the South American epidemic strain differs from the 7th pandemic El Tor strain in locus LAP (leucyl leucyl aminopeptidase). Here it is shown that this rare allele is present in 1 V. mimicus and 4 non-O1 V. cholerae. Non toxigenic O1 strains from South India epidemic share zymovar 14A with the epidemic El Tor from the 7th pandemic, while another group have diverse zymovars. The sucrose negative epidemic strains isolated in French Guiana and Brazil have the same zymovar of the current American epidemic V. cholerae. PMID- 12118283 TI - Anionic sites, fucose residues and class I human leukocyte antigen fate during interaction of Toxoplasma gondii with endothelial cells. AB - Toxoplasma gondii invades and proliferates in human umbilical vein endothelial cells where it resides in a parasitophorous vacuole. In order to analyze which components of the endothelial cell plasma membrane are internalized and become part of the parasitophorous vacuole membrane, the culture of endothelial cells was labeled with cationized ferritin or UEA I lectin or anti Class I human leukocyte antigen (HLA) before or after infection with T. gondii. The results showed no cationized ferritin and UEA I lectin in any parasitophorous vacuole membrane, however, the Class I HLA molecule labeling was observed in some endocytic vacuoles containing parasite until 1 h of interaction with T. gondii. After 24 h parasite-host cell interaction, the labeling was absent on the vacuolar membrane, but presents only in small vesicles near parasitophorous vacuole. These results suggest the anionic site and fucose residues are excluded at the time of parasitophorous vacuole formation while Class I HLA molecules are present only on a minority of Toxoplasma-containing vacuoles. PMID- 12118284 TI - Polymorphism of the Human Immunodeficiency Virus Type 1 in Brazil: genetic characterization of the nef gene and implications for vaccine design. AB - Most of the Brazilian HIV-1 samples have been characterized based on the structural genes (env, gag and pol) and no data concerning the variability of the accessory genes such as nef have been available so far. Considering the role of the nef on virus biology and the inclusion of this region in some HIV/AIDS vaccine products under testing, the purpose of this study was to document the genetic diversity of the nef gene in third-four HIV-1 Brazilian samples previously subtyped based on the env C2-V3 region. Although only few non-subtype B samples have already been analyzed so far, the cytotoxic Tlymphocyte epitopes encoded in this region were relatively conserved among the subtypes, with some amino acid signatures mainly in the subtype C samples. Considering the increasing of the non-B HIV-1 subtypes worldwide, in special the subtype C, more data should be generated concerning the genetic and antigenic variability of these subtypes, as well as the study of the impact of such polymorphism in HIV/AIDS vaccine design and testing. PMID- 12118285 TI - Microsporidiosis of Tachinaephagus zealandicus Ashmead (Hymenoptera: Encyrtidae). AB - An undescribed microsporidium was found infecting Tachinaephagus zealandicus, a gregarious parasitoid that attacks third instar larvae of muscoid flies. Spores were present in all body regions and in all stages of development. Infected adults contained an average of 3.75 x 10(5) spores, and the pathogen was vertically transmitted to progeny. Infected female adults were fed either rifampicin or albendazole mixed with honey to determine the effectiveness of these drugs in preventing vertical transmission. After eight days of feeding on rifampicin the parasitoids produced progeny of which only 37% were infected. In contrast, albendazole-treated and untreated females produced progeny that were 97% and 100% infected, respectively. Healthy and infected colonies were established and studies were conducted to determine the mechanisms of transmission. It was observed that the efficiency of vertical (maternal) transmission was 96.3%. Uninfected parasitoid immatures also became infected when they shared superparasitized hosts with infected immatures. The method of transmission within superparasitized hosts is not known. PMID- 12118286 TI - Predation potential of the water bugs Sphaerodema rusticum on the sewage snails Physa acuta. AB - The sewage snail Physa acuta is a serious threat to certain economic plants and to the purification plant of sewage works by rendering the biofilters ineffective. Various attempts are being made to control it. The efficacy of the predacious water bugs Sphaerodema rusticum was judged experimentally, in the laboratory in the potential control of P. acuta. It is revealed that, when supplied separately, the first, second and third instar and the adult S. rusticum did not attack P. acuta belonging to 3.1-8 mm, 5.1-8 mm, 7.1-8 mm and 7.1; these extracts exhibited no cytotoxicity. Six extracts (13%) representing 4 plant species (40%) showed cytotoxic activity. The highest cytotoxicity was found in the dichloromethane extract obtained from E. cotinifolia leaves and the CC50 values for the most susceptible cell lines, HEp-2 and CHO, were 35.1 and 18.1 microgram/ml, respectively. PMID- 12118289 TI - An adenovirus vector containing the suicide gene thymidine kinase for a broad application in cancer gene therapy. AB - Treatment of cancer using gene therapy is based on adding a property to the cell leading to its elimination. One possibility is the use of suicide genes that code for enzymes that transform a pro-drug into a cytotoxic product. The most extensively used is the herpes simplex virus thymidine kinase (TK) gene, followed by administration of the antiviral drug ganciclovir (GCV). The choice of the promoter to drive the transcription of a transgene is one of the determinants of a given transfer vector usefulness, as different promoters show different efficiencies depending on the target cell type. In the experiments presented here, we report the construction of a recombinant adenovirus carrying TK gene (Ad TK) driven by three strong promoters (P CMV IE, SV40 and EN1) and its effectiveness in two cell types. Human HeLa and mouse CCR2 tumor cells were transduced with Ad-TK and efficiently killed after addition of GCV. We could detect two sizes of transcripts of TK gene, one derived from the close together P CMV IE/SV40 promoters and the other from the 1.5 Kb downstream EN1 promoter. The relative amounts of these transcripts were different in each cell type thus indicating a higher flexibility of this system. PMID- 12118290 TI - In vitro and in vivo anti-Trypanosoma cruzi activity of a novel nitro-derivative. AB - Nitroarylidenemalononitriles and their cyanoacetamide derivatives with remarkable anti-epimastigote properties, were synthesized attempting to obtain new 3,5 diamino-4-(5'-nitroarylidene)-4H-thiadiazine 1,1-dioxide derivatives, which in previous reports had shown anti-Trypanosoma cruzi activity. Tests to evaluate the cytotoxicity of compounds were performed on J774 macrophages. 5-nitro-2-thienyl malononitrile (5NO2TM), was the only product which maintained a high anti epimastigote activity at concentrations in which it was no longer cytotoxic, thus it was assayed against intracellular amastigotes. Its anti-amastigote activity was similar to that of nifurtimox. Afterwards in vivo toxicity and anti-chagasic activity were determined. A reduction in parasitemia was observed. PMID- 12118291 TI - Assessment of therapeutic response of Plasmodium vivax and Plasmodium falciparum to chloroquine in a Malaria transmission free area in Colombia. AB - In order to determine the frequency of therapeutic failures to chloroquine (CQ) in patients with malaria due to either Plasmodium falciparum or P. vivax, and to explore the usefulness of a malaria-free city as a sentinel site to monitor the emergence of drug resistance, 53 patients (44 infected with P. vivax and 9 with P. falciparum) were evaluated at the Laboratory of Parasitology, Universidad del Valle in Cali, Colombia. Patients received 25 mg/kg of CQ divided in three doses over 48 h; they were followed during 28 days according to WHO/PAHO protocols. While therapeutic failures to CQ in the P. vivax group were not detected, the proportion of therapeutic failures in the P. falciparum group was high (78%) and consistent with the reports from endemic areas in Colombia. The diverse origin of cases presenting therapeutic failure confirmed that P. falciparum resistant to CQ is widespread in Colombia, and further supports the change in the national antimalarial drug scheme. Monitoring of drug resistance in malaria free areas would be useful to identify sites requiring efficacy evaluation, and in some situations could be the most appropriate alternative to collect information from endemic areas where therapeutic efficacy studies are not feasible. PMID- 12118293 TI - Aggregation behaviour and interspecific responses in Rhodnius prolixus stal. AB - The response to intra- and interspecific faecal assembling signals was tested in Rhodnius prolixus. Papers impregnated with excrement of R. prolixus induced the aggregation of larvae of this species, but also of those of Triatoma infestans. However, faeces belonging to T. infestans were not able to assemble larvae of R. prolixus. On the other hand, there was no response of R. prolixus to putative chemical factors from their cuticle (footprints), in contrast to T. infestans. Results are discussed as related to the ecology of both species. PMID- 12118292 TI - Lack of correlation between seminal and plasma HIV-1 viral loads is associated with CD4 T cell depletion in therapy-naive HIV-1+ patients. AB - The present study was conducted to investigate a possible correlation between plasma (PVL) and seminal viral load (SVL) on treatment-naive HIV-1-infected patients in Vitoria, ES, Brazil. We also evaluated whether the progressive immunosuppression associated with HIV disease (as evidenced by declining CD4 T cell counts) has any impact on the correlation between PVL and SVL HIV-1. Viral load on paired blood and semen samples from 56 consecutive treatment-naive patients were evaluated and compared to CD4 cell counts. Viral load and T cell counts (cells/microl) were determined by NASBA and by flow cytometry, respectively. Overall, a strong positive correlation between PVL and SVL (rho = 0.438, p = 0.001) was observed. However, when patients were grouped according to their CD4 counts, this correlation was only significant among patients with CD4 counts > 200 cells/microl. Results presented here demonstrate the existence of a strong correlation between PVL and SVL on patients with CD4 cell counts > 200 cells/microl, suggesting that this association may correlate with disease progression. PMID- 12118294 TI - Experimental infection of Aedes albopictus (Diptera: Culicidae) Larvae with the Xiphidiocercariae of a hematolechid. AB - Aedes albopictus larvae were exposed, either individually or in groups, to different concentrations of xiphidiocercariae of Haematoloechus sp. for parasitological studies. It was observed the acute lethal effect and some aspects of the host-parasite relationship, such as delay or progress in the host life cycle, the number and location of the metacercariae in the host, adult host malformations and the amount of metacercariae required to cause death. A delay in the cycle and a high mortality rate was in general observed. Inside the larvae, the metacercariae were found predominantly in the thorax, abdominal segments and in the head, along with a reduced number in the anal lobe and cervix. It was shown that in addition to the quantity of metacercariae present, their location in the larvae was also relevant in the determination of mortality and anomalies. Malformed adults developed from larvae containing from one to three metacercariae. PMID- 12118295 TI - Effect of sequential cold shocks on survival and molting rate in Triatoma infestans klug. AB - The survival and molting incidence in Triatoma infestans, a vector of Chagas disease, were investigated following sequential shocks at 0 degrees C in fifth instar nymphs under moderate fasting and full nutritional conditions. The shocks were separated by intervals of 8 h and 24 h at 30 degrees C. The results indicated that in terms of insect survival, T. infestans is tolerant to a single cold shock at 0 degrees C even for 12 h, or to sequential cold shocks, regardless of the nutritional state of the specimens. In terms of molting rate, fasting enhanced the tolerance to sequential cold shocks, but did not exceed the tolerance acquired by fully-nourished specimens, except when cold shocks were separated by an 8 h interval at 30 degreesC. The protective action elicited by fasting was assumed to be additive to that induced by a single mild cold shock or sequential cold shocks. The cold-tolerance response of T. infestans may have favoured its survival in areas of South America with low temperatures, even considering that this species is predominantly associated with human habitats. PMID- 12118296 TI - Biology of Triatoma klugi Carcavallo, Jurberg, Lent & Galvao 2001 (Heteroptera: Reduviidae) under laboratory conditions: effects of distinct blood sources and susceptibility to Trypanosoma cruzi and Trypanosoma rangeli. AB - The life cycle of Triatoma klugi Carcavallo, Jurberg, Lent & Galvao 2001 was compared under laboratory conditions using two groups of the F1 generation obtained from field-collected bugs. Among the 100 nymphs weekly fed on mice (Group A) or chicken (Group B), 77% of Group A and 67% of Group B reached the adult stage, and the mean time from the first nymphal stage to adult was 190.08 +/- 28.31 days and 221.23 +/- 40.50, respectively. The average span in days for each stage per group and the number of blood meals required for each stage were also evaluated. The overall mortality rate was 23% and 33% for Groups A and B, respectively. The mean number of eggs laid per month in a three-month period was of 56.20, 51.70 and 73.20 for Group A, and 64.50, 53.50 and 38.71 for Group B. Despite the blood source, comparative analysis revealed no statistically significant differences in the life cycle of T. klugi under laboratory conditions. Infection rates over 60% were observed for both Trypanosoma cruzi strains tested. Even revealing high infection rates of the hemolymph by T. rangeli strains, T. klugi revealed no salivary gland infections and was not able to transmit the parasite. PMID- 12118297 TI - Anopheles albitarsis embryogenesis: morphological identification of major events. AB - Anopheles albitarsis embryogenesis was analyzed through confocal microscopy of clarified eggs. Using Drosophila melanogaster as reference system, the major morphogenetic events (blastoderm, gastrulation, germ band extension, germ band retraction, dorsal closure) were identified. The kinetics of early events is proportionally similar in both systems, but late movements (from germ band retraction on) progress slower in An. albitarsis. Major differences in An. albitarsis related to D. melanogaster were: (1) pole cells do not protrude from the blastoderm; (2) the mosquito embryo undergoes a 180 degrees rotation movement, along its longitudinal axis; (3) the head remains individualized throughout embryogenesis; (4) extraembryonary membranes surround the whole embryo. A novel kind of malaria control is under development and is based on the use of genetically modified mosquitoes. Phenotypic analysis of the embryonic development of mutants will be imposed as part of the evaluation of effectiveness and risk of employment of this strategy in the field. In order to accomplish this, knowledge of the wild type embryo is a prerequisite. Morphological studies will also serve as basis for subsequent development biology approaches. PMID- 12118298 TI - Pelecitus helicinus Railliet & Henry, 1910 (Filarioidea, Dirofilariinae) and other nematode parasites of Brazilian birds. AB - We report Pelecitus helicinus Railliet & Henry, 1910 from 13 species of birds of 2 orders and 7 families, collected from the states of Sao Paulo and Mato Grosso, Brazil. All 13 constitute new host records for this nematode. In addition, we report the first record of Aprocta golvani Diaz-Ungria, 1963 from Brazil and Monasa nigrifrons (Bucconidae), as well as a number of other nematode records from Neotropical birds. PMID- 12118299 TI - Experimental neuroschistosomiasis: inadequacy of the murine model. AB - Neuroschistosomiasis is rarely observed in human pathology, but it is of considerable importance. To investigate its pathogenesis, consequences and response to treatment, an experimental model would be desirable, but is not yet available, in spite of a few indications of a suitable mouse model in the literature. Severe, recent and late Schistosoma mansoni infections in outbred and inbred strains of mice revealed widespread distribution of parasite eggs in several organs, but only exceptionally did eggs reach the encephalus, thus revealing the inadequacy of the mouse as an experimental model for neuroschistosomiasis. PMID- 12118300 TI - [Health systems reform and equity in Latin America and the Caribbean]. PMID- 12118301 TI - [Health systems reform and equity in Latin America and the Caribbean: lessons from the 1980s and 1990s]. AB - This essay proposes a review of the issues of equity and reform in Latin America and the Caribbean in the context of changes in recent decades, emphasizing the discussion of health systems reform. The economic, political, and social context prevailing in the critical 1970s extensively favored budget cuts for public expenditures, cost containment, changes in the health sector power structure, and health services reorganization from an 'economicist', pragmatic, and restrictive perspective. An inventory of the Latin American economic and social situation is markedly negative, and efforts to recover from the damage done in the 1980s were largely unsuccessful in the 1990s. The reforms implemented in some paradigmatic countries (Chile, Colombia, Costa Rica, Argentina, and Brazil), in light of their specific characteristics, demonstrate the dissemination of a common agenda, adapted to the various national conditions. Some positive results of these processes were diluted in new problems caused by the reforms themselves, especially in countries with more radical adherence to the new reformist model; meanwhile, in the country where the public, universal system based on solidarity was most consolidated, the management changes have obtained the best results. However, overcoming inequalities is still a distant goal. PMID- 12118303 TI - [Equity and geographic distribution of financial resources in health systems]. AB - This study focuses on equity in health and specifically the geographic distribution of financial resources. The author reviews the main contemporary theories of social justice and discusses the concept of equity in general and specifically in the health field. Based on the discussion of selected international experiences (United Kingdom, Spain, and Italy), the Resource Allocation Working Party (RAWP) formula used in the United Kingdom is identified as the most adequate distributive methodology, sizing the relative needs based on the population's demographic and epidemiological profiles. Finally, the results are presented from a simulation performed for the Brazilian case, showing that a more equitable geographic distribution of financial resources would require a redistribution favoring the States of the North and Northeast. The article concludes by highlighting that a comparison of actual fund outlays by the Ministry of Health in 1994 and the results of the simulation with the RAWP methodology for the Brazilian case show that the principles written into Brazilian legislation were absent from the geographic distribution of financial resources. PMID- 12118302 TI - [Equity and health systems reform in Latin America]. AB - The aim of any health care system is to help improve the people's health, and to do so as efficiently as possible. In order to improve the efficiency and equity of health services provision, many countries around the world have implemented reforms, including several Latin American nations. However similar the objectives may appear, the various ways societies implement such reforms reflect different values and concepts. This article analyzes the egalitarian and neoliberal values underlying different concepts of equity in health care. The authors develop criteria to interpret selected health services funding and provision strategies in Latin American health system reforms. These criteria are then applied to health care financing and delivery policies under the reforms currently being implemented in Colombia and Costa Rica. PMID- 12118304 TI - [Health inequality indicators: a discussion of some methodological approaches as applied to neonatal mortality in the Municipality of Rio de Janeiro, 2000]. AB - Epidemiology has investigated the relationship between health status and different social and economic factors ever since the field emerged. Studies have consistently shown that the population's health status bears a strong social gradient, invariably unfavorable to the less privileged groups. Increasing interest in understanding and characterizing health inequalities has broadened the discussion in the recent literature on appropriate concepts and methodological procedures for measuring differences in health status according to socioeconomic level. This study presents a critical assessment of health inequality indicators, focusing on the following: the redistribution principle and its application to health status; the influence of income inequality; epidemiological and statistical approaches to the problem; and evaluation of health system performance in reducing health inequalities. As an example, inequalities in the neonatal mortality rate are analyzed in the city of Rio de Janeiro, Brazil, 2000, according to the mother's level of schooling, reviewing the minimum requisites for defining an adequate health inequality indicator. PMID- 12118306 TI - [Health reform, equity and the right to health in Colombia]. AB - The author develops a long-term perspective to assess advances in equity and the right to health in the Colombian health system reform. In a restricted political system, actors in the field of health in Colombia have chosen individualistic alternatives to legalize inequities in individual purchasing power for services. Despite the complex regulations established in the General System for Social Security in Health, there is a trend towards consolidating traditional inequities and to further restrict opportunities for achieving the right to health with full, equitable, universal guarantees. PMID- 12118305 TI - [Theory and practice of the health systems reforms: the cases of Brazil and Mexico]. AB - This study focuses on the role of public health experts in the contemporary health sector reform process. The authors discuss the issue based on the case of Brazil and Mexico, where a group of public health specialists have oriented their participation to influence the conflict concerning health policy reform in the respective countries. One approach has been to develop a new cognitive framework for technical health sector reform projects viewed as policy proposals with technical content. The purpose is to demonstrate how these specialists have managed to influence the national debate over health sector reform when the technical and scientific discussion leaves the academic sphere and reaches the social and political realm. The authors contend that this occurs because such technical and scientific knowledge has been postulated (independently of its intrinsic value) as a political and ideological alternative platform for sustaining a health sector reform proposal which, once transformed into a policy project, has served to aggregate certain political and social forces. PMID- 12118308 TI - [National pharmaceutical policy in Colombia and social security reform: access and rational use of medicines]. AB - Based on the new social security system in Colombia (1993), which establishes equity and mandatory care as the basis for public health care provision, the authors analyze whether the formulation and implementation of pharmaceutical policy promote accessibility, availability, and rational use of medicines, thereby contributing to equity in health. Two approaches were used: a macro approach centered on the legal framework and various actors in the reform process and a micro approach related to the processes and results in the drug supply system. The authors studied the legal instruments backing the country's pharmaceutical policy and evaluated their application, using indicators and a specific disease (diabetes mellitus) as a marker. Although there is a legal framework providing the people's right to access health care services and essential medicines, the country lacks a comprehensive pharmaceuticals policy. Most of the institutions experience problems in distributing the medicines listed under the Mandatory Health Plan, a low percentage of medicines is dispensed at zero cost, and a major portion of patients purchase medicines through associations of diabetics or rely on alternative medicine. The study unveiled several obstacles to equity in health care coverage and access to essential medicines. PMID- 12118307 TI - [The impact of social security system reform on health services equity in Colombia]. AB - To evaluate the impact on access to, and use of, health services in Colombia's new national health insurance system, the authors compared two cross sections of the population: before (1993) and after (1997), with the approval of Act 100, creating the General System for Social Security in Health (SGSSS). Two equity indicators were assessed: concentration curves (CC) and concentration indices (CI), summarizing the distribution of access to health care and utilization of health care services provided by the SGSSS according to income deciles. Between 1993 and 1997, the CI for access to insurance halved from 0.34 to 0.17; simultaneously, coverage increased from 23% to 57%, especially among the poorest segments of the population, where it increased from 3.7% to 43.7% as a result of subsidies provided by local governments. The CI for utilization of health care services did not vary significantly. Increased disease prevalence and utilization of services among the insured, due to biased selection of risks and moral hazards, were also documented. These findings suggest a positive impact by the Reform on inequalities in access to health care insurance; however, a similar effect on inequities in utilization of health services is not clear. PMID- 12118309 TI - [Neoliberal reinvention of inequality in Chile: the case of the health sector]. AB - This paper examines the main changes in the Chilean health system from the 1920s to the present, identifying contradictions, limits, and successes in terms of policy outcomes. Taking equity as the central theme, the analysis focuses on decisions made from the mid-1920s until 1973 in moving toward socialized medicine, with the sudden interruption of this process by the military coup in 1973. The author then discusses the set of regressive measures taken by authoritarian neoliberalism, with the help of Pinochet, to insert health into the market economy. Finally, the article analyzes efforts made by democratic administrations since 1990 to redress the legacy of socioeconomic inequalities, focusing on the commitment to rebuild the health care system with a new basis in equity, solidarity, and people's participation. PMID- 12118310 TI - [Inequalities in public health care provision in Chile]. AB - From 1997 to 1999, the Chilean Ministry of Health conducted studies on the health care networks in each of the country's 13 regions in order to help plan regional health sector development and define investment projects. Health insurance coverage displayed major geographic, age, and gender variations. Out-patient and in-patient medical care in the public sector showed substantial geographic variations. According to patient discharge records from national referral hospitals, only some 20% of total health care capability is used to treat 60% of the Chilean population living in regions outside the Greater Metropolitan area. Analysis of primary care funding shows that municipalities allocating the highest per capita funds are not the ones with the greatest health care needs. New reform proposals must address the issue of complementarity between the public and private health sectors and strengthen the Ministry of Health's leadership role in order for the health system to improve its overall response to the population's health care needs. PMID- 12118311 TI - [Equity issues in health care reform in Argentina]. AB - This article analyzes the historical and contemporary development of the Argentine health care system from the viewpoint of equity, a principle which is not explicitly mentioned in the system's founding documents. However, other values can be identified such as universal care, accessibility, and solidarity, which are closely related to equity. Nevertheless, the political dynamics characterizing the development of the country's health care system led to the suppression of more universalistic approaches, with group solidarity the only remaining principle providing structure to the system. The 1980s financial crisis highlighted the relative value of this principle as the basis for an equitable system. The authors illustrate the current situation with data on coverage under the medical social security system. PMID- 12118312 TI - [Deregulation and equity: the Obras Sociales reconversion process in Argentina]. AB - The health care services managed by trade unions and known as "Obras Sociales" form the groundwork for Argentina's Social Security and Health system. However, far from taking an equitable approach, these institutions highlight the country's prevailing income disparities, which in turn lead to major differences in access to care. The main focus of this study was the reformulation of social security health policies within the framework of deregulation from 1998 to 2000, analyzing the effects on availability of health care services from an equity perspective. The methodology used two related analytical levels: (1) a macro level viewing the process from the various players' strategies and (2) a micro level featuring the changes within a well-known trade union social security organization during its reconversion process, emphasizing its institutional scope and the opinions of its membership. The results thus pointed to the slow implementation of reforms initiated by the public sector, hindered by constant negotiations among the main corporate actors seeking to serve their particular interests, along with increased inequity and fragmentation due to the limited opening of free choice by members. PMID- 12118313 TI - [Equity in the health sector: evaluation of public policy in Belo Horizonte, Minas Gerais State, Brazil, 1993-1997]. AB - This article evaluates government measures to reduce inequity in the health sector in Belo Horizonte from 1993 to 1997. Our hypothesis is that a municipal administration committed to equity can reduce disparities in health with the support of the Unified National Health System (SUS). The methodology used an urban quality of life index in Belo Horizonte to detect social inequalities in living conditions, as well as differences between the component indices in the infant mortality rate. Other municipal measures were assessed according to the investment resulting from the implementation of a participatory local budget and open planning process. The urban quality of life index appeared to be an appropriate measure for orienting municipal administration. The infant mortality rate proved to be a good indicator for measuring inequality in health. There was a reduction in IMR and mortality reducing gaps in the districts studied. We observed greater investment of physical and financial resources in the districts with the lowest urban quality of life index, and it can thus be stated that the municipal administration reduced the prevailing inequalities. PMID- 12118314 TI - [Proposals for health reform and equity in Uruguay: a redefinition of the Welfare State?]. AB - This article reviews and analyzes health sector reform proposals in Uruguay and the possible effects of such reforms in terms of equity, the health sector's institutional structure, and the power relationship between the various actors in the process. The authors contend that a highly structured yet simultaneously fragmented system has conspired against any attempt to introduce major reforms into the system. Thus the only possibility for reform resides neither in the consolidation of the so-called Institutions for Collective Medical Care (IAMCs) nor in the move towards a residual model. Rather, Uruguay is witnessing the system's passive restructuring (i.e., reform by default). In this context and given the system's built-in inequities, the current trend is towards an even more regressive distribution of goods and services. The authors use qualitative and quantitative techniques to show that inequities in expenditure, access, and quality have resulted from long-term developments and adaptive movements of an IAMC system in fiscal stress and the public system's declining quality. Thus, in the absence of changes in state policy that redefine the actors' power or in the absence of system collapse, the country should expect this same regressive trend to deepen. PMID- 12118315 TI - [Health sector reform and pharmaceutical policy in Peru]. AB - This article analyzes the Shared Pharmaceutical Management Program (PACFARM) and its relationship to pharmaceutical policy in Peru within the scope of health sector reform. Implementation of various programs for essential medicines has involved an on-going effort towards improving the supply of essential drugs to the community. However, the corresponding legal framework includes random and disconnected regulations which hinder the feasibility of a consistent national drug policy. PACFARM is a decentralized system for the provision of essential medicines on a care-level basis, self-supported by revolving funds. While expanded coverage and decreased economic barriers to access to essential medicines provided the basis for a pharmaceutical policy and traits of supply management efficiency kept pace with administrative modernization as part of the reform, other aspects hindered the program's implementation and limited its effects, including deregulation and the very processes of change in the sector. The study's methodology included qualitative and quantitative techniques, prioritizing an analysis of the program's implementation. PMID- 12118316 TI - Normal ras genes: their onco-suppressor and pro-apoptotic functions (review). AB - The ras family of oncogenes has been extensively studied for its implication in several types of human malignancies. Activation of ras genes involves mutations that alter the catalytic activity of the protein enhancing the downstream signals mostly towards cell proliferation and malignant transformation. Ras genes are also involved in induction of senescence or apoptosis, suggesting activation of alternative pathways that may be anti-oncogenic. Early experiments showed that transfection of wild-type ras in transformed cells reversed the oncogenic phenotype suggesting that wild-type ras has onco-suppressive properties. Indeed, expression of wild-type ras genes in several human malignancies is associated with good prognosis. In tumors carrying mutant ras genes the levels of expression of the wild-type allele never exceeded the mutant counterpart, indicating that the wild-type protein suppresses the effect of the mutant one. Recent development of the Kras2 deficient mice provided the tool to study the role of wild-type ras genes in tumorigenesis. PMID- 12118317 TI - Association of p53 mutations, microvessel density and neoangiogenesis in pairs of colorectal cancers and corresponding liver metastases. AB - p53 suppressor gene mutations are a well known step which occurs in the late stages of the complex tumourigenesis of colorectal cancer. A deregulation of p53 protein function may be associated with increased neovascularization and aggressive tumour growth. In vitro studies have shown that these genetic alterations cause a loss of wild-type p53-induced anti-angiogenetic control and could possibly induce expression of the neoangiogenic vascular endothelial growth factor (VEGF). Therefore, this in vivo study was performed to assess p53 mutations, i.e. hot spots in exons 4-9, in primary colorectal cancers and in corresponding liver metastases in order to test whether there is an association between p53 mutated tumours with increased microvessel density (MVD) and VEGF overexpression. Twenty-two tissue samples taken from primary colorectal cancers and the corresponding liver metastases were immediately snap-frozen in liquid nitrogen and fixed in formaldehyde. After DNA extraction exons 4-9 were amplified and directly sequenced. Cryostat sections were stained immunohistochemically using antibodies against VEGF, CD34, and p53 protein. A modified semiquantitative Weidner score and interactive computerized image analysis was used to assess MVD. Overexpression of immunohistochemically detected p53 protein was found in 7 of the 11 primary tumours and liver metastases (64%). Sequencing showed 3 out of 11 primary tumours (27%) and 5 out of 11 liver metastases (46%) to have p53 point or frameshift mutations; these samples tested immunohistochemically positive for p53 protein. Two p53 mutations in samples of liver metastases were not detectable in the corresponding primaries. We detected one frameshift mutation in exon 4 that has not yet been described in the literature. Tumour samples with p53 mutations and increased VEGF immunoreactivity were associated with higher MVD (p<0.01 and p<0.05, respectively). However, there was no association detected immunohistochemically between p53 and MVD as well as p53 mutations and VEGF overexpression. Our data demonstrate specific genetic alterations in the coding regions of p53 suppressor gene in both primary colorectal cancers and corresponding liver metastases, these alterations are associated with an increase in MVD, but not in VEGF overexpression. In addition, a novel frameshift mutation in both colorectal cancer and metastasis is described. PMID- 12118318 TI - TNF-alpha gene and proton radiotherapy in an orthotopic brain tumor model. AB - The major goal of this study was to evaluate the safety and efficacy of TNF-alpha gene therapy (pGL1-TNF-alpha) in combination with proton radiation in an orthotopic brain tumor model. C6 glioma cells were implanted into the left hemibrain of athymic rats (day 0). On day 5, pGL1-TNF-alpha (19 microg/10 microl) was injected into the same site; appropriate control groups were included. Proton irradiation (10 Gy, single fraction) was performed 18-20 h thereafter and, on day 10, a portion of animals from each group was assayed. Nearly all tumor-bearing groups had lower body mass compared to those without tumor; brain mass was somewhat increased with plasmid (pGL1-TNF-alpha or pWS4) injection (p<0.05). Histopathological analysis of brain sections revealed that rats receiving pGL1 TNF-alpha/proton irradiation had the smallest tumors and lowest number of mitotic tumor cells, although survival time for animals kept long-term was not significantly prolonged. A decline in leukocyte populations was noted with combination treatment compared to controls (p<0.05), but no differences were found compared to groups receiving each modality alone. Based on DNA synthesis, the pGL1-TNF-alpha/proton irradiated group had the highest levels of leukocyte activation. The highest percentage of lymphocytes expressing the CD71 activation marker occurred with pGL1-TNF-alpha, whereas the proton-irradiated group had the highest percentage of activated NK cells (NK1.1+/CD71+). No significant differences were found in erythrocyte and thrombocyte numbers, hemoglobin, and hematocrit. Overall, the data indicate that pGL1-TNF-alpha/proton treatment results in a measurable antitumor effect and is safe under the conditions used. PMID- 12118319 TI - Tissue-specific regulation of Fas/APO-1/CD95 expression by p53. AB - The regulation of Fas/APO-1(CD95), an important member of the tumor necrosis factor (TNF) superfamily involved in membrane-mediated apoptosis, has been a subject of recent research. Ligation of Fas by Fas ligand or an anti-Fas cross linking antibody triggers receptor trimerization followed by recruitment of FADD to the cytoplasmic domain of the receptor and the activation of the caspase cascade. The tumor suppressor p53 has been shown to upregulate Fas expression under numerous pro-apoptotic stimuli in vitro. Using the p53 knockout mouse model, we demonstrate by Western blot analysis, immunohistochemistry, and semi quantitative RT-PCR that Fas expression is reduced in spleen and liver from p53-/ mice compared to p53+/+ controls, while similar expression levels were observed in brain, heart, kidney, lung, skin, testis, and thymus between the two groups. While Fas protein was abundant in brain, heart, liver, and spleen, low levels of endogenous expression was observed in other tissues from the p53+/+ and p53-/- mice. These data indicate that p53 regulates Fas expression in a tissue-specific manner. PMID- 12118320 TI - Expression of telomerase genes in thyroid carcinoma. AB - Telomerase (T) is a ribonucleoprotein complex that includes the telomerase RNA component (hTR), telomerase associated protein (TP1) and the telomerase catalytic subunit (hTERT). Telomerase has been shown in stem cells and found to be activated in tumor tissues and immortalized cells. We wanted to test whether the expression of the telomerase complex subunits correlate with the enzyme activity in human thyroid tissue. Hence, we determined the expression of hTERT, hTR and TP1 mRNA by RT-PCR and compared the results to telomerase activity as detected by the telomeric repeat amplification protocol (TRAP) assay. Fifteen benign goiters (G), 11 follicular carcinomas (FTC) including 2 oncocytic follicular carcinomas (also called Hurthle cell carcinoma, oFTC), 12 papillary carcinomas (PTC) including 3 microcarcinomas (mPTC), and 12 undifferentiated anaplastic thyroid carcinomas (UTC) were investigated. Experienced pathologists performed histological and pTNM classification in each specimen. RT-PCR analysis revealed that TP1 was ubiquitously expressed in all G and carcinomas. hTR was expressed in 4 out of 15 G, in 2 out of 3 mPTC, in 5 out of 9 PTC, in 5 out of 9 FTC, in all oFTC and in 9 out of 12 UTC samples. Regarding all carcinomas, no statistically significant correlation was observed between hTR-expression and tumor stage, lymph node or distant metastasis. hTERT-expression was associated with malignancy and tumor stage. All mPTC and 13 out of 15 G did not express hTERT, whereas all samples of pT3-4 tumor stage of FTC, PTC, UTC and all oFTC were positive for hTERT. No telomerase activity could be detected in G. Telomerase activity in carcinoma was only measurable in tissues that expressed the catalytic subunit hTERT. Our data indicate that telomerase activity is up-regulated in neoplastic cells. In contrast to TP1 and hTR, hTERT and telomerase activity may be of help in identifying invasive tumors and may be additional markers for classification of benign goiter and malignant thyroid carcinoma. PMID- 12118321 TI - Expression of the potential novel gene E6DG1 downregulated by the E6 protein of human papillomavirus type 16 is correlated with anchorage-independent growth. AB - A novel gene transcript that is downregulated by HPV16 E6 protein was identified in mouse cells using differential hybridization and designated E6DG1. The cloned cDNA of E6DG1 was 1.3 kb in length and contained a small ORF potentially encoding a polypeptide of 45 amino acids. In vitro transcription and translation of E6DG1 cDNA resulted in a product of approximately 7 kDa and Western blot analysis using antibodies for E6DG1 peptide detected 7 and 14 kDa proteins. Downregulation of E6DG1 mRNA levels in cells expressing HPV16 E6 protein was observed at subconfluent cell densities, but not in confluent cells. Repression of E6DG1 protein enhanced the anchorage-independent growth and weakened the cell adhesion in Panc1 cells. Immunofluorescence analysis revealed the localization of E6DG1 within the nucleus. These results indicate that the E6DG1 protein may function in a signaling pathway related to anchorage-independent growth and adhesion control. PMID- 12118322 TI - Expression of platelet-derived endothelial cell growth factor/thymidine phosphorylase in cervical intraepithelial neoplasia. AB - Angiogenesis contributes to the growth and secondary spreading of solid tumors. Platelet-derived endothelial cell growth factor (PD-ECGF)/thymidine phosphorylase (TP) has been identified as such an angiogenic factor. In this study, the expression of PD-ECGF/TP and VEGF was evaluated by immunohistochemical staining of tumor specimens from 40 patients with cervical intraepithelial neoplasia (10 with moderate dysplasia; 10 with severe dysplasia; 10 with carcinoma in situ; 10 with invasive carcinoma). The microvessel density was assessed by immunostaining for factor VIII-related antigen in the most highly neovascularized area. In both the nucleus and cytoplasm, the intensity of PD-ECGF/TP expression in carcinoma in situ and invasive carcinoma was significantly stronger than that in moderate dysplasia. However, the intensity of VEGF expression was not significantly different in the various specimens. The microvessel density in mild dysplasia was significantly different from that in carcinoma in situ (p<0.05), and that in invasive carcinoma (p<0.05). There was no significant relationship between the microvessel density and the expression of PD-ECGF/TP or that of VEGF. These results show that the expression of PD-ECGF/TP appears to be involved in the promotion of angiogenesis in cervical intraepithelial neoplasia. PMID- 12118323 TI - Differential changes in purine nucleotides after Doxorubicin treatment of human cancer cells in vitro. AB - The present investigation was performed to elucidate the role of purine nucleotides as potential indicators of chemosensitivity of malignant tumors. Drug sensitive (s) and -resistant (r) tumor cell lines grown as monolayers (s: T47D, MCF-7 wild-type; r: NCI/ADR-RES, MCF-7/MDR) or as multicellular spheroids (T47D; NCI/ADR-RES) were exposed to 0.1, 1.0, and 10.0 microM Doxorubicin for up to 24 h. Purine nucleotides were assayed using HPLC and with some selected spheroids using imaging bioluminescence. The data show that in the time frame of the experiments reproducible and statistically significant changes in the nucleotides only occur at the highest drug concentration investigated. Under these conditions and using monolayer cultures, Doxorubicin caused a significant increase in ATP and GTP in sensitive but not in resistant cancer cells. Consequently, this differential change may be exploited for drug sensitivity testing in vitro. Doxorubicin exposure to spheroids was associated with significant increases in ATP and GTP in both sensitive and resistant variants. However, the kinetic of the changes in GTP was largely different between T47D and NCI/ADR-RES spheroids with a long-lasting, almost 3-fold elevation and a smaller, relatively short transient increase in GTP, respectively. Supplementing experiments with Doxorubicin treatment under inhibition of oxidative phosphorylation with Oligomycin abolished the drug-induced ATP and GTP peaks at persistent increases in ADP and AMP. Assuming that the spheroids may represent the in vivo situation to a better degree than monolayer cultures, experimental in vivo studies should clarify whether kinetic changes in GTP could be used as differential markers for the chemosensitivity of solid tumors. The experiments using Oligomycin support the hypothesis that purine nucleotides may be recycled from DNA fragments that result from the interaction of the drug with the DNA strands. PMID- 12118325 TI - Efficacy and safety of FdUMP[10] in treatment of HT-29 human colon cancer xenografts. AB - Thymidylate synthase (TS) is the molecular target of fluoropyrimidine (FP) chemotherapy, and novel anticancer drugs effective against TS-overexpressing tumors are required to treat patients with FP-refractory solid tumors. We have evaluated the inhibition of cell proliferation in vitro and antitumor activity in vivo of FdUMP[10], an oligodeoxynucleotide 10mer in which 5-fluorouracil (5-FU) is the only nucleobase. FdUMP[10] is a pro-drug of FdUMP, the TS inhibitory metabolite of FPs. FdUMP[10] was 338-fold more potent than 5-FU at inhibiting cell proliferation in the NCI 60 cell line screen. The antitumor activity of FdUMP[10] was compared to 5-FU using H-T29 xenografts in female CD-1 athymic (nu+/nu+) mice. Treatment with FdUMP[10] as a single agent (40 mg/kg/daily x 5, i.v.) delayed tumor growth and resulted in a smaller mean tumor size (T/C value = 51%, p<0.001 compared with the control group). Treatment with 5-FU (25 mg/kg/daily x 5, i.p.) had similar results as single agent FdUMP[10] (T/C value = 65%, p=0.238 compared with the FdUMP[10] treated group. Simultaneous treatment of tumor-bearing mice with both drugs (FdUMP[10] plus 5-FU) further delayed tumor growth (T/C value = 36%; p=0.003 relative to 5-FU). The results from the combined treatment group were not, however, statistically significant relative to the group receiving single agent FdUMP[10] treatment (p=0.059). Histological examination revealed systemic damage was limited to the colonic epithelium in all treatment groups and was least extensive with single agent FdUMP[10] compared to the other treatment groups. The data support the concept that FdUMP[10] is a useful prototype of a novel type of FP that is likely to be more efficacious than FPs in clinical use. PMID- 12118324 TI - Prevalence of microsatellite instability, inactivation of mismatch repair genes, p53 mutation, and human papillomavirus infection in Korean oral cancer patients. AB - To determine the etiologic factors of human oral cancer, we examined the prevalence of microsatellite instability (MSI), the inactivation of mismatch repair (MMR) genes, p53 mutation, and human papillomavirus (HPV) infection (HPV 16, -18, and -33) in 86 Korean oral cancer specimens, including 76 squamous cell carcinomas and 10 salivary gland tumors. MSI was observed in 3 of the 76 squamous cell carcinomas (4%) and 2 of 10 salivary gland tumors (20%). As MSI is a hallmark of the inactivation of the MMR genes, the genetic status of hMSH2 and hMLH1, and hypermethylation of the hMLH1 promoter region were investigated in oral cancers displaying MSI. Inactivation of the hMLH1 gene by either mutation or hypermethylation was observed 4 of the 5 MSI oral cancers. Mutation of the p53 gene was found in 11 of 76 squamous cell carcinomas (14.5%) but not in the salivary gland tumors. PCR assay revealed the presence of HPV DNA in 11 of the 76 squamous cell carcinomas (14.5%) and 4 of the 10 salivary gland tumors (40%). Type 18 HPV DNA was predominant in 11 of the HPV-infected squamous cell carcinomas (72.7%) and 4 of the HPV-infected salivary gland tumors (50%). Two squamous cell carcinoma tissues were found both to be HPV-infected and to harbor the p53 mutation. Our results suggest: i) that MSI plays a role in the pathogenesis of Korean oral cancers, squamous cell carcinomas (4%) and salivary gland tumors (20%); ii) that genetic alteration or hypermethylation of the hMLH1 gene may be the principal inactivating mechanism in Korean oral cancer with MSI; and iii) that inactivation of the p53 gene by either mutation or HPV infection is frequent in Korean squamous cell carcinomas (26%) and salivary gland tumors (40%). PMID- 12118326 TI - Role of retinoblastoma protein and E2F-1 transcription factor in the acquisition of 5-fluorouracil resistance by colon cancer cells. AB - 5-fluorouracil (5-FU) is an important antineoplastic agent that has proven to be effective in the treatment of colorectal cancer. However, one of the main obstacles to the clinical use of 5-FU is the acquisition of resistance to the drug by cancer cells. In vitro studies have demonstrated that the resistance to 5 FU is correlated with increased activity of thymidylate synthase (TS), whose gene has a E2F binding site in its promoter region. To understand the mechanisms through which cancer cells acquire resistance to 5-FU, human colon cancer-derived cell line DLD-1 and its subcloned cell line DLD-1/5-FU, which has acquired resistance to 5-FU, were compared by assessing their phosphorylation of retinoblastoma protein (pRb) and E2F-1 transcriptional activity. The level of pRb phosphorylation in the DLD-1/5-FU cells was higher than in the parental DLD 1cells. In parallel with the increased phosphorylation of pRb, E2F-1 transcriptional activity, which has been shown to be a result of E2F-1 dissociation from hyperphosphorylated pRb, was increased in the DLD-1/5-FU cells. Examination of the effect of E2F-1 decoy oligodeoxynucleotides (ODNs) on the proliferation of DLD-1/5-FU cells in the presence of 5-FU to confirm the importance of E2F-1 in the mechanisms of the acquisition of 5-FU resistance showed that DLD-1/5-FU cells transfected with E2F-1 decoy ODNs recovered their sensitivity to 5-FU. These results suggested that pRb and E2F-1 play important roles in the acquisition of 5-FU resistance by cancer cells and that cancer therapy targeting transcription factor E2F might be effective. PMID- 12118328 TI - Fusion of RDC1 with HMGA2 in lipomas as the result of chromosome aberrations involving 2q35-37 and 12q13-15. AB - Rearrangements of chromosome bands 12q13-15 are frequent in various benign mesenchymal and epithelial tumors, and the gene HMGA2 seems to be the most common target within this chromosome region. In the majority of cases, the rearrangements result in a fusion of the first three exons of HMGA2 with different translocation partners. Despite the large number of HMGA2 mutations that have been reported, very little is known about the fusion partners. In this study, we have characterized a recurrent fusion of the first three exons of HMGA2 5' to the G protein-coupled receptor gene (RDC1) in lipomas with rearrangements involving chromosome bands 2q35-37 and 12q13-15, one of several recurrent chromosomal rearrangements in lipomas. The functional impact of the fusion is truncation of HMGA2, because the RDC1 part contributes with a stop codon one amino acid downstream of the breakpoint. The breakpoint within RDC1 was localized in a previously uncharacterized exon of the gene, and our data suggest that RDC1 is subject to alternative splicing. PMID- 12118327 TI - Survivin expression and its correlation with cell proliferation and prognosis in epithelial ovarian tumors. AB - Survivin is a new member of the inhibitors of apoptosis proteins (IAP) family, selectively overexpressed in common human cancers but not in normal adult tissues, and associated with aggressiveness of the disease and unfavorable outcomes. Recent study also found that survivin expression is associated with cell proliferation. In order to gain insight into the role of survivin in ovarian tumors, we investigated the expression of survivin in a group of epithelial ovarian tumors, and examined the relationship of its expression with cell proliferation and clinical outcome. Immunohistochemical analysis was performed in 103 cases of epithelial ovarian tumors. Twenty-six of the 103 cases were evaluated by Western blot analysis. The results showed that survivin overexpression was detected in 21.2% (7 of 33) of benign tumors, 47.8% (11 of 23) of borderline tumors, and 51.1% (24 of 47) of ovarian carcinomas. The positive ratio was significantly higher in malignant or borderline tumors than in benign tumors, and the overexpression of survivin was significantly correlated with the size of residual disease. A positive correlation between survivin expression and proliferative activity of tumor cell measured by PCNA index was found. Kaplan Meier analysis demonstrated that the patients with survivin overexpression have a short overall survival. These findings suggest that survivin overexpression may play a pivotal role in the progression of ovarian tumors and may provide an important prognostic implication for epithelial ovarian carcinomas. PMID- 12118330 TI - Radiation-induced effects in unirradiated cells: a review and implications in cancer. AB - A long-held central dogma of radiation biology has been that the carcinogenic effects of ionizing radiation (IR) are induced by the direct and radiolytic actions of IR on nuclear DNA. Numerous investigations, however, have revealed that several cancer relevant effects of IR can occur in cells that have received only cytoplasmic or plasmalemmal membrane exposure to IR. Further, mounting evidence now indicates that many effects that have been attributed to IR-induced damage to nuclear DNA or that occur following irradiation of the cytoplasmic compartment of cells can also occur in cells that have received no direct exposure to IR per se. These so-called , i.e., radiation induced effects in unirradiated cells, include cell killing, increases in intracellular reactive oxygen species, the induction of mutations, enhanced cell growth, the induction of apoptosis, the induction of genomic instability and neoplastic transformation. In this report, we summarize the evidence that demonstrates IR can cause this array of effects in non-irradiated cells, and we discuss recent findings concerning the potential mechanisms that may underlie IR induced effects in unirradiated, or cells. Additionally, we discuss IR-induced bystander effects and their possible relationship to some in vivo observations, how bystander effects may pertain to carcinogenesis the treatment of tumors with radiotherapy, and how effects in bystander cells contribute to uncertainties in assessing cancer risks associated with exposure to IR. PMID- 12118331 TI - 7-hydroxystaurosporine (UCN-01) and ionizing radiation combine to inhibit the growth of Bcl-2-overexpressing U937 leukemia cells through a non-apoptotic mechanism. AB - A clinically relevant dose (2.0 Gy) of ionizing radiation (IR) was employed to determine if subsequent exposure to the protein kinase C (PKC) and Chk 1 inhibitor UCN-01 for 24 h could abrogate IR-induced G2/M arrest and promote apoptosis in U937 leukemic cells ectopically expressing Bcl-2 (U937/Bcl-2). To this end, empty-vector control (U937/pCEP4) and U937/Bcl-2 cells were exposed to two UCN-01 concentrations following IR: i) a 50 nM concentration, which by itself was minimally toxic to both cell lines, and ii) a 150 nM concentration, which modestly induced apoptosis (e.g., ~19%) in control cells after 24 h. The effects of UCN-01 on IR responses were examined in relation to apoptosis induction, suspension culture growth inhibition, loss of clonogenic survival, and cell cycle perturbations. IR (2 Gy) alone minimally induced apoptosis in both U937 transfectant cell lines (e.g., <5% at 24 h in each case). Although UCN-01 failed to potentiate IR-mediated apoptosis at either early (e.g., 24 h) or late (e.g., 72 h) intervals, exposure to 50 or 150 nM UCN-01 resulted in a significant, albeit modest, reduction in proliferation and colony formation in irradiated U937/pCEP4 and U937/Bcl-2 cells. Despite failing to enhance apoptosis, UCN-01 treatment abrogated IR-induced G2/M arrest in both cell lines, an event associated with enhanced activation of cyclin-dependent kinase 1 (cdk1), promotion of G0/G1 arrest, and dephosphorylation of the retinoblastoma protein (pRb). Together, these findings indicate that exposure of U937 cells ectopically expressing Bcl-2 to the combination of UCN-01 + IR leads to a further reduction in cell proliferation, and that this phenomenon appears to involve a non apoptotic mechanism. PMID- 12118332 TI - Pemetrexed disodium combined with oxaliplatin, SN38, or 5-fluorouracil, based on the quantitation of drug interactions in human HT29 colon cancer cells. AB - Premetrexed disodium (MTA) is a novel multitargeted antifolate that inhibits thymidylate synthase, dihydrofolate reductase, and glycinamide ribonucleotide formyltransferase. It exhibits a broad spectrum of activity against several human tumor types including colorectal cancer. Therefore, we evaluated the anti proliferative potential of MTA combined with drugs known to exert therapeutic activity against colon cancer, including 5-fluorouracil, oxaliplatin, and SN38, the active metabolite of irinotecan. The effects of MTA, alone or combined with one of theses 3 drugs, were investigated in parental human HT29 colon cancer cells and in 5-fluorouracil-resistant counterparts HT29-5FU cells. These drugs were administered either simultaneously or sequentially. Functional interactions between MTA, 5-fluorouracil, oxaliplatin, and SN38 were evaluated using median effect plot analysis. The drug combination and sequence with optimal effects were evaluated in athymic mice bearing human HT29 tumor cell xenografts. Combinations of MTA with 5-fluorouracil required high concentrations to achieved additive and/or synergistic effects. Simultaneous exposure to MTA and oxaliplatin led to synergistic activity in both parental and 5-fluorouracil-resistant human HT29 colon cancer cells, leading to additive antitumor effects and minimal toxicity in athymic mice bearing HT29 cell tumors. Synergism between MTA and SN38 was also observed in both parental and 5-fluorouracil-resistant HT29 cells. These results argue in favor of clinical trials of chemotherapy combining MTA with oxaliplatin or irinotecan (CPT-11), for the treatment of patients with colon cancer. PMID- 12118333 TI - Consensus statements from the Second International Lung Cancer Molecular Biomarkers Workshop: a European strategy for developing lung cancer molecular diagnostics in high risk populations. AB - The Second Molecular Biomarkers Workshop was held at the Roy Castle International Centre for Lung Cancer Research in Liverpool, in June 2001 and it brought together experts in the clinical, epidemiological and molecular-pathology of lung cancer from Europe and the USA, to address issues surrounding the development of a European strategy for early lung cancer detection. The 2001 Workshop Breakout Groups concentrated on the current challenges in the early detection of lung cancer which need to be addressed in the light of the recent surge in interest in many countries for mounting new clinical trials to evaluate the utility of Spiral CT in early lung cancer detection. If population-based trials of CT screening are mounted it will also be a favorable clinical environment in which to evaluate efficiently recent advances in molecular screening and genotyping. The Workshop focused specifically on: a) clinical and molecular biomarkers, b) sputum as an early detection and diagnostic tool, c) validation of molecular markers prior to their use in early detection trials and d) ethical issues that have to be considered in early lung cancer detection trials. A distillation of the Workshop discussions is given in this article. PMID- 12118329 TI - IGFBP-3 mediates p53-induced apoptosis during serum starvation. AB - Insulin-like growth factor binding protein (IGFBP)-3, a p53-response gene, can induce apoptosis in an IGF-independent manner. Here we demonstrate that IGFBP-3 mediates p53-induced apoptosis during serum starvation using two foil neoplastic cell models: one which introduces p53 activity and one which eliminates it. We created a doxycycline-inducible p53 model from the p53-negative PC-3 prostate cancer cell line. Doxycycline treatment increased both p53 and IGFBP-3 levels. It also augmented apoptosis, but not during insulin-like growth factor-I co treatment. In a second model, lung carcinoma H460 cells expressing fully functional p53 were stably transfected with E6, which targets p53 for degradation. H460-E6 cells contained less p53 and IGFBP-3 than control neo transfected cells, and proteasome blockade restored both. In serum deprivation, H460-E6 cells had enhanced growth and less apoptosis than did H460-neo cells. Reductions in H460-neo apoptosis, comparable in magnitude to H460-E6, were achieved by adding anti-IGFBP-3-antibody or IGFBP-3 antisense oligomers, but not non-specific immunoglobulin or IGFBP-3 sense oligomers. In summary, turning p53 in two foil neoplastic cell models induced IGFBP-3 expression and increased apoptosis during serum starvation, an effect inhibited by insulin-like growth factor-I treatment and specific IGFBP-3 blockade. This is the first demonstration of inhibition of p53 action by antagonizing IGFBP-3. PMID- 12118334 TI - Oxytocin modulates estrogen receptor alpha expression and function in MCF7 human breast cancer cells. AB - Oxytocin (OT) inhibits the proliferation of MCF7 estrogen-dependent human breast cancer cells, via specific OT receptors (OTR). Besides this effect, we report that OT modulates the expression of estrogen receptor alpha (ERalpha) in MCF7 cells, both at mRNA and protein level. Since the first 24 h of OT treatment the ERalpha mRNA levels are down-regulated; in contrast, ERalpha protein expression decreases at a later time. The reduced number of ERalpha goes in parallel to a temporary increase in the binding affinity of these receptors as well as to a significant increase in their estradiol (E2)-induced transcription activity. The increase in both binding affinity and transcriptional activity of ERalpha likely balances the reduction in the number of ERalpha binding sites, ruling out the hypothesis that part of the OT contrasting effect on E2-induced cell proliferation could depend on the reduced E2 binding to MCF7 cells and supporting the hypothesis of an exclusively direct OT-antimitogenic effect. This is the first evidence that OT modulates the expression of ERalpha receptors in human neoplastic cells. PMID- 12118335 TI - Curcumin inhibits cell cycle progression of immortalized human umbilical vein endothelial (ECV304) cells by up-regulating cyclin-dependent kinase inhibitor, p21WAF1/CIP1, p27KIP1 and p53. AB - To elucidate possible mechanisms of anti-angiogenic activity by curcumin, we performed cDNA microarray and found that curcumin modulated cell cycle related gene expression. For further confirmation, DNA contents and expression levels of cyclins, cyclin-dependent kinases (CDKs), and CDK inhibitors (CDKIs) were examined by FACS analysis and Western blotting, respectively. Curcumin was found to induce G0/G1 and/or G2/M phase cell cycle arrest, up-regulate CDKIs, p21WAF1/CIP1, p27KIP1, and p53, and slightly down-regulate cyclin B1 and cdc2 in ECV304 cells. However, expression level of other cyclins and CDKs were not changed by curcumin. We, therefore, conclude that the up-regulation of CDKIs by curcumin plays a critical role in the regulation of cell cycle distribution in these cells, which may have a major role in anti-angiogenic activity of curcumin. PMID- 12118336 TI - Resistance of MCF-7 cells to dimethylbenz(a)anthracene-induced apoptosis is due to reduced CYP1A1 expression. AB - We have developed a series of aryl hydrocarbon (AH)-resistant cell lines derived from MCF-7 human breast epithelial cancer cells by continuous exposure to the AH benzo[a]pyrene. These cell lines display cross-resistance to the mammary carcinogen dimethylbenz[a]anthracene (DMBA). Apoptosis induced by exposure to DMBA is greatly decreased in the resistant cell lines compared to the wild-type, in proportion to the level of resistance. Apoptosis induced by DMBA could be blocked by inhibitors of DMBA metabolism such as alpha-naphthoflavone and diosmetin. We therefore examined the resistant cell lines for their ability to metabolize DMBA and for the formation of DMBA-DNA adducts, and found that both parameters were decreased compared to wild-type cells in proportion to the level of resistance. When exposed to DMBA or 2,3,7,8-tetrachlorodibenzo-p-dioxin, the resistant cell lines have a diminished capacity to carry out ethoxyresorufin-O deethylation, indicating that the induction of cytochrome P450 1A1 (CYP1A1) enzyme is impaired. We therefore examined the expression of the CYP1A1 gene, and found reduced levels of both CYP1A1 mRNA and CYP1A1-promoter controlled transcription in resistant cells compared to the wild-type. The deleterious effects of AHs are believed to be mediated by the aryl hydrocarbon receptor (AhR), a ligand-activated transcription factor which regulates CYP1A1 expression. Resistant cell lines had a reduced expression of the AhR, as measured at the mRNA and protein levels. These data demonstrate that AH resistance in these cells is mediated by changes in the signal transduction pathway which regulates CYP1A1 expression. PMID- 12118337 TI - Involvement of reactive oxygen species in aclarubicin-induced differentiation and invasiveness of HL-60 leukemia cells. AB - Aclarubicin (ACLA), which belongs to the anthracycline class of antineoplastic agents, has been demonstrated as a differentiating agent of human leukemia, including HL-60 cells. We report here on the incidence of ACLA-induced differentiation on matrix metalloproteinase (MMP) expression and cell invasiveness. The aim of this study was to investigate the involvement of reactive oxygen species (ROS) as mediators of ACLA-induced effects. By using a fluorescent probe, we showed that subtoxic (i.e. differentiating) concentration of ACLA generate reactive oxygen species in HL-60 cells. ACLA-differentiated cells exhibited an increased proMMP-9 secretion which has been observed by gelatin zymography and immunoassay. Antioxidants were able to inhibit ACLA induced differentiation and proMMP-9 secretion. Furthermore, RT-PCR showed that ACLA increased MMP-9 and tissue inhibitor of MMP (TIMP-1) expression in a ROS dependent manner. In addition, the migration and invasion capacities of HL-60 cells were enhanced by ACLA treatment, but only partially reversed by antioxidants. Altogether, these results evidenced ROS as messengers of ACLA induced differentiation and MMP-9 expression. PMID- 12118338 TI - Mutational status of overexpressed p16 in head and neck cancer: evidence for germline mutation of p16/p14ARF. AB - Inactivation of the p16 tumor suppressor gene is a common phenomenon in squamous cell carcinoma of the head and neck (SCCHN). Less commonly described is the observation of p16 overexpression in SCCHN. Since overexpression of p16 is a potent predictor of outcome in other cancers, we were interested in determining the level of expression of p16 in our SCCHN specimens as a prerequisite to later prognostic studies. We were also interested in determining the mutational status of p16 in these tumors, in order to determine whether the combination of overexpression and gene alteration may predict a different clinical outcome from overexpression alone. A total of 84 specimens of SCCHN were selected for study. These specimens were obtained from all major sites within the oral cavity, oropharynx, pharynx and larynx. The level of expression of p16 in SCCHN specimens was measured by semi-quantitative RT-PCR. In 35 cases, RNA was also isolated from matched normal tissue obtained from a negative tumor margin. In the other 49 cases, the expression level was compared with the level of expression measured in pooled normal RNA obtained from 10 specimens of normal epithelial tissue. Overexpression of p16 was documented when the level of expression in the tumor specimen was 2-fold or greater above the level of expression found in normal tissue. A total of 46 specimens demonstrated overexpression of p16 (55%). All specimens demonstrating overexpression were then subject to sequence analysis. Thirty specimens (65%) showed p16-specific gene alterations, ranging from intragenic deletions to single point mutations, and 15 of these cases concomitantly affect p14ARF. A single specimen demonstrated a silent point mutation within the p16 reading frame. This mutation produces a stop codon at residue 85 in the context of the p14ARF reading frame, predicting premature termination of p14ARF within a previously determined nucleolar localization signal. This observation suggests that in some cases at least, p14ARF may be a selective target for alteration, independently of p16. Analysis of a normal tissue specimen obtained from a negative tumor margin, and a blood sample obtained approximately five years after surgery indicate that this p14ARF specific alteration may represent a germline mutation. PMID- 12118339 TI - P53 mutations in colorectal cancer assessed in both genomic DNA and cDNA as compared to the presence of p53 LOH. AB - Mutations in p53 is a most common genetic alteration in human cancer and has been related to outcome, also in colorectal cancer. However, published results are not unanimous on this point without clear-cut explanations. Different analytical methods have been applied which could explain some discrepancies. Another factor of importance may be the source of nucleic acids used for identification of sequence alterations. Therefore we compared mutations in exons 7 and 8 of p53 found in genomic DNA, with mutations in cDNA from the same patients and evaluated to what extent LOH and mutations in p53 occurred simultaneously, which are prerequisites for a complete loss of p53 function according to the classic concept. cDNA and genomic DNA from tumor tissue from 123 patients were used. Thirty-four mutations were found in both tumor cDNA and genomic DNA. Twenty missense mutations were the same in both cDNA and genomic DNA. Two missense mutations, one 1 bp deletion and one nonsense mutation were only found in genomic DNA, while nine missense mutations and one 9 bp deletion were found in cDNA only. Statistical analysis showed no significant difference among mutations in cDNA and genomic DNA (p=1.00). Only 2 patients (5%) had both LOH and mutations in exons 2 11 of p53. One patient had LOH without p53 mutation, while 18 patients (44%) showed p53 mutation without LOH and 20 patients (49%) had wt p53 without LOH. Forty-four patients (52%) were non-informative for LOH of the entire cohort consisting of 85 patients. Our findings suggest that different results reported on the role mutated p53 may have in colorectal cancer are probably not explained by the source of nucleic acids (RNA vs DNA) for sequence determinations. Rather, unknown information about simultaneous but different presence of p53 LOH in previously published reports is more likely. However, an unexpectedly infrequent occurrence (5-10%) of complete and isolated p53 ablation (LOH + mutation) would demand considerably larger patient populations (1500-2000 patients) than ever published for relating p53 function to survival in patients with colorectal cancer. Therefore, the role of p53 dysfunction in progression of colorectal cancer remains uncertain. PMID- 12118340 TI - In vitro pharmacological characterizations of the anti-angiogenic and anti-tumor cell migration properties mediated by microtubule-affecting drugs, with special emphasis on the organization of the actin cytoskeleton. AB - The aim of the present work is to investigate whether microtubule-affecting drugs including vincristine, vinblastine, vindesine and vinorelbine are able to produce an anti-angiogenic effect at non-cytotoxic doses in the same way of taxol. The cytotoxic effects were determined by means of the colorimetric MTT assay, and the anti-angiogenic effects on HUVEC cells growing on Matrigel and forming capillary networks. Sixteen additional drugs (camptothecin, SN38, topothecan, adriamycin, daunomycin, etoposide, bleomycin, melphalan, mitomycin C, TNP-470, cisplatin, carboplatin, 5-fluorouracil, methotrexate, suramin and batimastat) were used as control in order to test the specificity of the microtubule-affecting drug effects. We also investigated by means of videomicroscopy whether microtubule affecting drugs could produce anti-migratory effects at non-cytotoxic doses on tumor cells. Finally, we used computer-assisted fluorescence microscopy to characterize the influence of microtubule-affecting drugs on the polymerization/depolymerization dynamics of the actin cytoskeleton in tumor cells. Our results show that taxol, vincristine and vindesine behave similarly in their ability to reduce the capillary network formation by HUVEC cells cultured on Matrigel. These anti-angiogenic effects appear at non-cytotoxic concentrations. In contrast, vinblastine and vinorelbine produce apparent anti angiogenic effects by direct cytotoxicity. Microtubule-affecting agents are also able to significantly reduce the level of migration of tumor cells at non cytotoxic concentrations, some of these effects may occur via modifications to the actin cytoskeleton organization. Several types of microtubule-affecting agents could be used as anti-angiogenic agents by administering them at non cytotoxic concentrations, and some microtubule-affecting agents abandoned in pharmacological assays could turn out to be potent anti-migratory drugs acting on tumor cells, though without being too cytotoxic. PMID- 12118342 TI - Misrepair of radiation-induced DNA double-strand breaks and its relevance for tumorigenesis and cancer treatment (review). AB - The faithful repair of DNA double-strand breaks (DSBs) is probably one of the most critical tasks for a cell in order to maintain its genomic integrity since these lesions may lead to chromosome breaks or rearrangements, mutations, cell death or cancer. DSBs can arise spontaneously during normal cellular DNA metabolism or may be induced by exogenous agents such as ionizing radiation. To overcome the danger that emanates from these lesions, eukaryotic cells have evolved specific pathways for processing DSBs by either homology-dependent or non homologous repair mechanisms. This review focuses on the formation of genomic rearrangements that arise by joining incorrect break ends and on the factors that influence repair fidelity. Recent studies indicate that the probability for a break to be incorrectly rejoined is fairly low when DSBs are spatially separated but increases drastically when multiple breaks coincide. The formation of genomic rearrangements in situations of multiple breaks is mediated by non-homologous end joining, the predominant DSB repair pathway in mammalian cells. Interestingly, the same pathway is required for efficiently preserving chromosomal integrity in situations of separated breaks. Furthermore, the probability for a DSB to be faithfully repaired depends on its genomic location and on the cell cycle position. Methods for assaying DSB repair are discussed, again with emphasis on experimental systems that allow to determine whether a DSB is correctly or incorrectly rejoined. PMID- 12118341 TI - Long-term outcomes of transcatheter arterial chemoembolization with autologous blood clot for unresectable hepatocellular carcinoma. AB - Ninety-two consecutive patients with unresectable hepatocellular carcinoma (HCC) greater than 2 cm and less than 5 cm in diameter were treated by transcatheter arterial chemoembolization using autologous blood clot as an embolizing agent (Short-TAE; S-TAE). Survival, patency of hepatic arteries and side effects were retrospectively analyzed. The median follow-up interval was 42 months. The median size of the main tumor was 3.4 cm in diameter. All tumors were determined to be inoperable because of intrahepatic spread of tumors and/or poor hepatic functional reserves. S-TAE was performed by injecting a mixture of iodized oil and anticancer drugs followed by embolization of hepatic arteries with autologous blood clot. Embolization with clots maintained patency of hepatic arteries in all the patients. The overall survival rates for 92 HCC patients at 1, 3, 5, 7 and 8 years were estimated by the Kaplan-Meier method to be 100, 52, 34, 12 and 6%, respectively, which were better compared with prior records for conventional method with gelfoam. S-TAE was successfully performed in 16 patients with Childs class C liver cirrhosis presenting icterus and ascitis. The survival rates for Childs class C patients at 1, 2 and 3 years were 100, 66 and 32%, respectively. The Cox proportional hazard model was used to assess influence of each pretreatment parameter (age, gender, virus marker, stage of cirrhosis, number of tumor) on survival. The univariate analysis demonstrated that both the stage of cirrhosis and the multiplicity of tumor were significant factors predicting survival (p=0.003 and 0.046, respectively). The multivariate analysis showed the stage of cirrhosis to be the sole factor which significantly affected prognosis of HCC patients (p=0.005). In conclusion, chemoembolization with autologous blood clot is an effective and safe therapeutic option for unresectable HCC smaller than 5 cm. The hepatic functional reserves play a key role in determining prognosis of HCC patients. PMID- 12118343 TI - CYFRA 21-1 is released in TNF-alpha-induced apoptosis in the hepatocellular carcinoma cell line HuH-7. AB - Many types of cancer cells widely express cytokeratin 19 (CK19). Clinical investigations have suggested that serum CYFRA 21-1, a fragment of CK19, is one of the most useful tumor markers. In the present study, we hypothesized that released CYFRA 21-1 is closely associated with cellular apoptosis during tumor growth. Apoptosis was induced by tumor necrosis factor-alpha (TNF-alpha) in the HuH-7 hepatocellular carcinoma cell line. Both TNF-alpha-treated and non-treated cells of HuH-7 were simultaneously examined by immunoradiometric assay, annexin-V apoptosis analysis, immunohistochemical staining and colorimetric protease measurement. Levels of CYFRA 21-1 increased significantly in TNF-alpha-treated cells displaying a high percentage of apoptosis, granular-like aggregation of CK19, and elevated activity of caspase-3 in contrast to non-treated cells. Levels of CYFRA 21-1 decreased significantly after caspase-3 was inhibited in TNF-alpha treated cells. Thus, the release of CYFRA 21-1 may suggest cellular apoptosis in the process of tumor growth. PMID- 12118344 TI - DNA content and cell number determination in microdissected samples of breast carcinoma in situ. AB - Paraffin-embedded tissue (PET) is the specimen of choice for the histopathological diagnosis of cancer. PET has become a valuable resource for correlating cellular phenotype and genotype in microdissected lesions. A definitive improvement in this field has been the development of infra-red laser capture microdissection (LCM), which yields homogeneous populations of cells for DNA extraction and in vitro amplification by polymerase chain reaction (PCR). We report here a photographic and fluorescent technique for determining the number of nuclei and concentration of DNA obtained respectively by laser capture microdissection from paraffin-embedded breast cancer tissue. Breast biopsies containing carcinoma in situ were serially sectioned, mounted on plain glass slides and tumor cells were microdissected using a laser capture microscope. The DNA was extracted in digestion buffer and used directly as a template for PCR. In our protocols, each capture contained 21+/-5.4 nuclei with a DNA concentration of 115+/-5.3 pg. Also, a linear relationship was found between number of captures and DNA content (R2=0.9995). These results represent a novel contribution for a more precise correlation between phenotypic and genotypic diversity in cancer cells studied from microdissected paraffin-embedded tissue. PMID- 12118345 TI - Duplex ultrasound criteria for defining the severity of carotid stenosis. AB - Duplex ultrasound scan (DUS) criteria for grading >50% carotid artery stenosis is typically divided into broad categories such as 50-79% stenosis, 80-99% stenosis, and occlusion. The purpose of this study is to validate DUS criteria for stratifying 50 to 100% carotid stenosis into 10% intervals using digital substraction cerebral angiography (DSCA) as the standard of comparison. Between 1996 and 2001, 163 patients were evaluated with duplex ultrasound and angiography. A total of 326 carotid arteries were studied using DUS in an accredited ICAVL vascular laboratory. Threshold velocity criteria for determining the degree of carotid stenosis was defined according to seven categories: <50%, 50-59%, 60-69%, 70-79%, 80-89%, 90-99%, and occlusion. Treatment decisions were based on the angiographic findings. In cases where the degree of stenosis as defined by duplex velocity criteria did not correlate with angiographically defined stenosis, each record was reviewed to determine whether the angiographic findings altered the surgeon's treatment decision. The sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) for DUS defined degree of stenosis as compared to angiographically defined stenosis were determined. There was a high correlation (R = 0.96) between duplex scan and angiography in 93% (302/326) of the cases. Clinical management was altered in only 3% (10/326) of the cases because of the results of angiography. The DUS velocity criteria to grade the severity of carotid disease in 10% intervals is reliable and accurate. Clinical management of patients with carotid stenosis can be based solely on carotid DUS in 97% of patients considered for treatment of carotid artery disease. PMID- 12118346 TI - Bilateral subclavian arterial aneurysm and ruptured abdominal aorta pseudoaneurysm in Behcet's disease. AB - Behcet's disease is characterized by recurrent ulcers of the mouth and genitalia and relapsing iritis. It is recognized as a chronic multisystem disease affecting the skin, mucous membranes, eye, joints, central nervous system, and blood vessels. About 8% of the patients with Behcet's disease have severe vascular complications such as arterial aneurysm and occlusion. In our patient, there was a massive, painful, pulsatile mass on the clavicle on the right side of neck. A left subclavian artery aneurysm mass was observed on the left apex on a chest X ray. Through angiography, a lobular giant saccular aneurysm on the proximal side of the right subclavian artery, giant aneurysm on the left subclavian artery, and occlusion on the left subclavian-axillary artery were observed. We treated first the right and then the left subclavian arterial aneurysm with a two-stage operation. The aneurysms were resected and polytetrafluoroethylene (PTFE) graft interposition was performed. Control angiography was performed 6 months postoperatively. Both grafts were open and there was no anastomotic aneurysm. The patient was reoperated on for a ruptured abdominal aorta pseudoaneurysm 13 months after the first operation. The aortic defect was repaired using a Dacron patch. PMID- 12118347 TI - Free flap coverage of pyoderma gangrenosum leg ulcers. AB - BACKGROUND: Pyoderma gangrenosum is a necrotizing skin disorder of unknown cause. Treatment of pyoderma gangrenosum usually entails medical treatments. Surgical treatment is generally not successful. OBJECTIVE: Two patients with pyoderma gangrenosum of their lower extremities are presented. The application and utility of microvascular free flap coverage of their ulcers is discussed. RESULTS: Both patients achieved successful healing with microvascular free flaps. CONCLUSION: Microvascular free flap coverage may be a surgical treatment option for selected patients with pyoderma gangrenosum. PMID- 12118348 TI - Experimental keloid scar models: a review of methodological issues. AB - BACKGROUND: Keloid scars are benign fibrous proliferations in the dermis that arise after dermal trauma. The scars are raised in appearance and extend beyond the boundaries of the original wound. Scarring in predisposed individuals is out of proportion to the severity of the inciting wound. Current treatments sometimes yield early benefit but scars often resume exuberant growth. The pathophysiology of keloid scars is still poorly understood. In order for new treatments to be developed, the mechanisms leading to the formation of keloid scars must be further elucidated. The search for improved experimental models is of critical importance because such models have an important role to play in both the study of keloid formation and in the development of new therapies. OBJECTIVE: The objective of this article is to introduce the reader to the experimental models available for studying keloid scars and to outline the advantages and limitations of animal and tissue culture models. CONCLUSION: Both models may help to elucidate the pathways of keloid formation and promote development and testing of therapies. Tissue culture is better suited to studies of pathogenesis, whereas the animal models are more suitable for therapeutic testing. PMID- 12118363 TI - Nonsteroidal antiinflammatory drug-induced pseudoporphyria: a case series. AB - BACKGROUND: Pseudoporphyria is a diagnosis that is used when porphyria-like clinical lesions arise in the setting of normal porphyrin levels. This condition was first described in the 1960s and was initially related to the use of certain antibiotic drugs. In 1985, pseudoporphyria was first attributed to the use of nonsteroidal antiinflammatory drugs (NSAIDs). Subsequently, a host of NSAIDs and other drugs have been found to elicit the same clinical entity. The exact mechanism by which certain drugs create clinical lesions resembling porphyria cutanea tarda or erythropoietic protoporphyria is still unknown. A phototoxic mechanism is hypothesized. OBJECTIVE: We describe six patients diagnosed with pseudoporphyria and detail the diagnostic tests leading to the eventual diagnosis. RESULTS: The patients ranged in age from 27 to 59 years and had a female:male predominance of 2:1. The offending NSAID was DayPro (oxaprozin) for three of the patients, Relafen (nabumetone) for two of the patients, and Aleve (naproxen) for one patient. For each patient, histology and immunofluorescence was either consistent with the diagnosis of porphyria cutanea tarda or nonspecific, while serum, stool, and urine porphyrins did not support that diagnosis. Withdrawal of the offending agent provided relief from the clinical symptoms for each patient. None of our patients were rechallenged with the putative offending drug. However, prolonged avoidance has provided a sustained remission from symptoms in all six patients. CONCLUSIONS: Pseudoporphyria is a relatively rarely reported condition. Clinical suspicion with appropriate laboratory and histopathologic findings help to make this diagnosis, and exclude true porphyrias. Rechallenge with the offending drug to produce symptom relapse has been proposed to be helpful in confirming this diagnosis of exclusion. Since all 6 patients with drug-induced pseudoporphyria experienced resolution of their symptoms after discontinuing the offending agent, we propose that this clinical correlation alone is sufficient to confirm this diagnosis. Our observation of six new cases of NSAID-induced pseudoporphyria over a two-year interval suggests that this is not a rare entity. PMID- 12118364 TI - Effects of stimulus level on speech perception with cochlear prostheses. AB - This study is one of a series that examines stimulus features important for cochlear implant function. Here, we examine effects of stimulus level. In subjects with cochlear implants, a number of psychophysical tests of temporal discrimination (pulse rate discrimination, gap detection, etc.) show marked improvement as a function of stimulus level through most or all of the dynamic range, while electrode-place discrimination can improve or degrade as a function of level. In this study, effects of these combined potential influences were studied by examining the effects of stimulus level on syllable identification. We tested two hypotheses: that syllable identification varies as a function of stimulus level and that level and electrode configuration interact in affecting syllable identification. We examined vowel and consonant identification as a function of stimulus level for bipolar and monopolar electrode configurations. We used experimental processor maps where upper and lower stimulation limits of each electrode pair were equated to eliminate confounding effects of dynamic range, which varies across subjects and electrodes. For each channel, stimulation amplitude was set to a fixed percentage of its dynamic range. Eight adult subjects with Nucleus CI24M implants were tested using the SPEAK processing strategy. With each electrode configuration, stimulus levels were tested from 0% to 90% of the dynamic range in nine steps. The effects on consonant and vowel identification were similar. Phoneme identification was usually better for monopolar than for bipolar stimulation. In the lower half of the dynamic range, syllable identification usually increased as a function of stimulus level. In the upper half of the dynamic range, syllable identification continued to increase as a function of level to 90% of the dynamic range for some subjects, while for others there was no appreciable change or a decrease as a function of level. Decreases in performance at high levels were more common with monopolar than bipolar stimulation. These results suggest that if speech processors are programmed to optimize level for each individual, speech perception performance could be improved. PMID- 12118365 TI - SV40 large T-antigen disturbs the formation of nuclear DNA-repair foci containing MRE11. AB - The accumulation of DNA repair proteins at the sites of DNA damage can be visualized in mutagenized cells at the single cell level as discrete nuclear foci by immunofluorescent staining. Formation of nuclear foci in irradiated human fibroblasts, as detected by antibodies directed against the DNA repair protein MRE11, is significantly disturbed by the presence of the viral oncogene, SV40 large T-antigen. The attenuation of foci formation was found in both T-antigen immortalized cells and in cells transiently expressing T-antigen, indicating that it is not attributable to secondary mutations but to T-antigen expression itself. ATM-mediated nibrin phosphorylation was not altered, thus the disturbance of MRE11 foci formation by T-antigen is independent of this event. The decrease in MRE11 foci was particularly pronounced in T-antigen immortalized cells from the Fanconi anaemia complementation group FA-D2. FA-D2 cells produce essentially no MRE11 DNA repair foci after ionizing irradiation and have a significantly increased cellular radiosensitivity at low radiation doses. The gene mutated in FA-D2 cells, FANCD2, codes for a protein which also locates to nuclear foci and may, therefore, be involved in MRE11 foci formation, at least in T-antigen immortalized cells. This finding possibly links Fanconi anaemia proteins to the frequently reported increased sensitivity of Fanconi anaemia cells to transformation by SV40. From a practical stand point these findings are particularly relevant to the many studies on DNA repair which exploit the advantages of SV40 immortalized cell lines. The interference of T-antigen with DNA repair processes, as demonstrated here, should be borne in mind when interpreting such studies. PMID- 12118366 TI - Yes-associated protein (YAP65) interacts with Smad7 and potentiates its inhibitory activity against TGF-beta/Smad signaling. AB - Members of the TGF-beta family of growth factors signal from the cell surface through serine/threonine kinase receptors. Intracellular propagation of the signal occurs by phosphorylation of intracellular proteins of the Smad family. Smad7 belongs to the subclass of inhibitory Smads that function as antagonists of TGF-beta signaling. A yeast two-hybrid screen of a human placental cDNA expression library using full-length mouse Smad7 as bait identified Yes Associated Protein (YAP65) as a novel Smad7-interacting protein. The association of Smad7 with YAP65 was confirmed using co-expressed tagged proteins in COS-7 cells. Deletion of the PY motif of Smad7 reduced but did not abolish YAP65-Smad7 association, suggesting the existence of several interacting domains. We demonstrate that YAP65 potentiates the inhibitory activity of Smad7 against TGF beta-induced, Smad3/4-dependent, gene transactivation. Furthermore, YAP65 augments the association of Smad7 to activated TGF-beta receptor type I (TbetaRI), whereas YAP65(1-301), which exerts a dominant-negative effect against Smad7-driven inhibition of TGF-beta signaling, reduces these interactions. Together, these data provide the first evidence that YAP65 is a Smad7 partner that facilitates the recruitment of the latter to activated TbetaRI, and enhances the inhibitory activity of Smad7 against TGF-beta signaling. PMID- 12118368 TI - Microsatellite instability at tetranucleotide repeats in skin and bladder cancer. AB - Recently, a novel form of MSI has been described that occurs only at tetranucleotide repeat markers. This has been termed elevated microsatellite instability at selected tetranucleotide repeats (EMAST). EMAST has been related to alterations of the p53 gene, and to the nature of the repeat sequence. We initially tested whether loss of heterozygosity (LOH) at the p53 and the patched (ptch) genes was related to EMAST in a series of 61 non-melanoma skin cancer (NMSC) tumors. We then analysed a series of 57 primary bladder cancers for the presence of EMAST, testing whether this was related to mutation or expression of the p53 gene. In both NMSC and bladder tumors we found a high prevalence of EMAST (75.4 and 43.9%). In NMSC the prevalence of EMAST was higher in tumors that had either p53 or ptch LOH, although the difference was not statistically significant. There was a significant association of extensive EMAST (three or more loci) with mutations in p53 among the bladder cancer tumors, but no indication of elevated EMAST in tumors with abnormal p53 staining without mutation. The association of EMAST with p53 mutation was confined to non-invasive disease. Hence, EMAST likely reflects a particular pattern of somatic events that are interactive with p53 mutation, particularly common in skin cancer and limited to non-invasive disease in bladder cancer. PMID- 12118367 TI - K252a inhibits the oncogenic properties of Met, the HGF receptor. AB - The ATP analog K252a is a potent inhibitor for receptor tyrosine kinases of the Trk family. Here we show that nanomolar concentrations of K252a prevent HGF mediated scattering in MLP-29 cells (30 nM), reduce Met-driven proliferation in GTL-16 gastric carcinoma cells (100 nM), and cause reversion in NIH3T3 fibroblasts transformed by the oncogenic form of the receptor, Tpr-Met (75 nM). K252a inhibits Met autophosphorylation in cultured cells and in immunoprecipitates and prevents activation of its downstream effectors MAPKinase and Akt. Interestingly, K252a seems to be more effective at inhibiting the mutated form of Met (M1268T) found in papillary carcinoma of the kidney than the wild type receptor. Pretreatment of both Tpr-Met-transformed NIH3T3 fibroblasts and of GTL-16 gastric carcinoma cells with K252a results in loss of their ability to form lung metastases in nude mice upon injection into the caudal vein. These observations suggest that K252a derivatives, which are active in vivo as anti cancer drugs in models of Trk-driven malignancies, should also be effective for treatment of Met-mediated tumors. PMID- 12118369 TI - The breast proto-oncogene, HRGalpha regulates epithelial proliferation and lobuloalveolar development in the mouse mammary gland. AB - Members of the EGF family of growth factors play critical roles during normal and neoplastic breast development. EGF family member HRGalpha is the only HRG1 isoform expressed in the mouse mammary gland and our previous experiments suggest that HRG1 has a unique role in mammary development. To determine the function of HRGalpha activity during mouse mammary gland development, we generated a HRGalpha deficient mouse strain. Unlike mice with HRG1 or isoform specific HRGbeta gene deletions, HRGalpha-null mice survive to adulthood. HRGalpha-deficient mice display pronounced defects in mammary gland lobuloalveolar development at 17 days of pregnancy and 3 days post-partum. Terminal and lateral ductal alveoli were condensed and alveolar outgrowth during pregnancy was severely impaired. A dramatic reduction in beta-casein expression accompanied defective alveolar development in the HRGalpha-null mice, as determined by in situ hybridization and Northern blot analysis of HRGalpha-deficient mammary glands at 3 days post partum. Expression of the milk-protein genes WAP and alpha-lactalbumin was not adversely affected. In situ incorporation of BrdU demonstrated that epithelial proliferation was significantly curtailed in mammary glands of HRGalpha-deficient mice at 17 days post-coitus and 3 days post-partum. These results demonstrate that HRGalpha is an important mammary gland mitogen regulating alveolar development and lactogenesis. PMID- 12118370 TI - Loss of B cell identity correlates with loss of B cell-specific transcription factors in Hodgkin/Reed-Sternberg cells of classical Hodgkin lymphoma. AB - In classical Hodgkin lymphoma the malignant Hodgkin/Reed-Sternberg (HRS) cells characteristically constitute only a small minority of the tumour load. Their origin has been debated for decades, but on the basis of rearrangement and somatic hypermutations of their immunoglubulin (Ig) genes, HRS cells are now ascribed to the B-cell lineage. Nevertheless, phenotypically HRS cells have lost their B cell identity: they usually lack common B cell-specific surface markers such as CD19 and CD79a as well as Ig gene transcripts. Here we demonstrate that Ig promoters as well as both intronic and 3' enhancer sequences are transcriptionally inactive in HRS cell lines. This inactivity correlates with either reduced levels or even a complete lack of several B cell-specific transcription factors required for their expression: Oct-2, OBF-1, PU.1, E47/E12, PAX-5 and EBF. Moreover, we demonstrate that PU.1 and PAX-5 are significantly down-regulated in HRS cells in pathological specimens from primary tumour tissues. However, forced expression of these transcription factors can activate regulatory sequences of silenced B cell marker genes, and in one instance also transcription from a silenced endogenous locus. Thus, HRS cells are dedifferentiated B cells with global down-regulation of B cell-specific genes. PMID- 12118371 TI - Phosphorylation of human estrogen receptor alpha at serine 118 by two distinct signal transduction pathways revealed by phosphorylation-specific antisera. AB - Estrogen receptor alpha (ERalpha) is a transcription factor that regulates expression of target genes in a ligand-dependent manner. Activation of gene expression is mediated by two transcription activation functions AF-1 and AF-2, which act in a promoter- and cell-specific manner. Whilst AF-2 activity is regulated by estrogen (E2) binding, the activity of AF-1 is additionally modulated by phosphorylation at several sites. One of these phosphorylation sites, serine 118 (S118) is of particular interest as its mutation significantly reduces ERalpha activity. Previous studies have shown that S118 can be phosphorylated by the ERK1/2 mitogen activated protein kinases (MAPK) and by the cyclin-dependent protein kinase Cdk7. In this study we use antisera that specifically recognize ERalpha phosphorylated at S118 to demonstrate that MAPK phosphorylates S118 in a ligand-independent manner, whereas Cdk7 mediates E2 induced phosphorylation of S118. E2 stimulation of S118 phosphorylation was observed within 10 min of its addition and was maximal at 10(-7) M E2. S118 phosphorylation was maximal at 30 min but then declined, such that by 180 min following E2 addition little S118 phosphorylation was evident. S118 phosphorylation was also induced by the partial estrogen antagonist 4 hydroxytamoxifen, but not by the complete antagonist ICI 182, 780. S118 phosphorylation upon addition of the MAPK inducers EGF or PMA followed the expected time courses. Finally, we show that ERalpha is phosphorylated at S118 in vivo using immunoblotting of extracts prepared from a series of ERalpha-positive breast tumours. PMID- 12118372 TI - Molecular cloning and characterization of a novel gene which is highly expressed in hepatocellular carcinoma. AB - To gain new insight into the molecular mechanism underlying the pathogenesis of human primary hepatocellular carcinoma (HCC), we searched for HCC-specific molecules through screening genes that are differentially expressed between cancerous and noncancerous counterparts of liver and identified a novel HCC associated gene, HCCA1 encoding a approximately 80 kDa cytoplasmic protein that contains several proline-rich motifs likely for SH3-binding. HCCA1 transcript, albeit present in some adult tissues, is up-regulated selectively in HCC but not in other tumor cells. High expression of HCCA1 occurs as a late event frequently (89.2%) in HCCs and correlated significantly with the degree of tumor progression. When treated with antisense oligonucleotides to HCCA1, HCCA1 expression in HCC cells (HuH-7) was effectively suppressed and cell growth was down-regulated in a time- and dose-dependent manner. Furthermore, HuH-7 cells harboring the HCCA1 antisense expression clone displayed a remarkably reduced efficiency in colony formation. Together, these data strongly suggest that HCCA1 is a positive effector in cell proliferation and contributes to HCC carcinogenesis and progression. We believe that this protein will serve as a novel useful marker for HCC and is a potential target for pharmaceutical intervention of this malignant disease. PMID- 12118373 TI - Activation of mitochondrial voltage-dependent anion channel by apro-apoptotic BH3 only protein Bim. AB - Bcl-2 family of proteins regulates apoptosis by controlling mitochondrial membrane permeability. We have previously shown that the voltage-dependent anion channel (VDAC) plays a crucial role in apoptotic changes of the mitochondria and its activity is directly regulated by some Bcl-2 family members, including Bcl 2/Bcl-x(L) and Bax/Bak but not Bid. Here, we showed that in isolated mitochondria, Bim induced loss of membrane potential and cytochrome c release like Bax/Bak, with these changes being inhibited by an anti-VDAC antibody. In addition, microinjection of the anti-VDAC antibody significantly reduced Bim induced apoptosis. Study using purified proteins indicated that Bim directly interacts with the VDAC. Immunoprecipitation analysis revealed that Bim interacts with the VDAC and the interaction is remarkably enhanced during apoptosis. An experiment using liposomes indicated that Bim enhanced VDAC activity, as did Bax/Bak. Furthermore, Bim (but not tBid) was able to induce apoptotic changes of yeast mitochondria in a VDAC-dependent manner, and also induced the lysis of red blood cells, with this effect being inhibited by the anti-VDAC antibody. These results indicate that Bim has an ability to activate directly the VDAC, which plays an important role in apoptosis of mammalian cells. PMID- 12118375 TI - Inhibition of Rel/Nuclear Factor-kappaB signaling in skin results in defective DNA damage-induced cell cycle arrest and Ha-ras- and p53-independent tumor development. AB - In recent years a growth inhibitory role in skin for the Rel/NF-kappaB transcription factors has been established, and the block of Rel/NF-kappaB signaling results in rapid development of spontaneous skin cancer. The molecular mechanism underlying tumor development is however unknown. In the present study, we show that inhibition of NF-kappaB signaling in mouse skin by targeted expression of degradation resistant IkappaB-alpha generates transgenic keratinocytes unable to arrest the cell cycle in response to DNA damage induced by gamma-radiation. The results indicate that transgenic keratinocytes have a defect at the G1-S checkpoint whereas the G2-M checkpoint response was found to be intact. However, transgenic keratinocytes still respond by induction of the cyclin dependent kinase inhibitor p21(Cip1/Waf) after exposure to gamma radiation. In the spontaneous skin tumors that develop in transgenic mice no mutations were found in the Ha-ras or p53 gene, suggesting that inhibition of NF kappaB signaling in skin can induce cancer development independently of initiating mutations in the Ha-ras gene or additional mutations in the p53 gene. These findings demonstrate an involvement of NF-kappaB signaling in the DNA damage response and cell cycle checkpoint control in the skin. PMID- 12118374 TI - The protective effect of phorbol esters on Fas-mediated apoptosis in T cells. Transcriptional and postranscriptional regulation. AB - Phorbol esters are tumor promoters that bind and activate both conventional and new Protein kinase C (PKC) isoforms. In various circumstances, PKC-dependent signaling pathways can promote cell survival and protect against cell death. This was first analysed in Jurkat T cells where Phorbol Myristate Acetate (PMA) was found to inhibit Fas-mediated apoptosis as judged by DiOC6(3) staining, caspase activation and DNA fragmentation, indicating that PMA exerts its protective effect upstream or at the mitochondrial level in these cells. PMA activated most of the main kinase pathways in T cells such as PKCs, p42/44MAPK, p38MAPK and p90Rsk but not JNK and Akt. A pharmacological approach allowed us to identify that nPKCs are both necessary and likely sufficient to promote T cell survival. Besides this post-transcriptional regulation, nPKCs may also regulate apoptosis at the transcriptional level. cDNA arrays were used to identify a set of genes whose expression was modulated in death versus survival conditions. Following PMA treatment, expression of Mcl-1 and Bcl-x increased while that of c-Myc was significantly reduced. Moreover, survivin expression decreased upon CH11 or PMA treatment. c-Myc, survivin and Bcl-x modulation seems to be regulated at the transcriptional level while decrease in Mcl-1 protein in CH11-treated cells resulted especially from a caspase-dependent proteolysis. Taken together, our data demonstrate that PMA-mediated inhibition of apoptosis is a complex process that is integrated at both the transcriptional and post-transcriptional level and point out to the potential role of Mcl-1, Bcl-x, c-Myc and survivin in this process. PMID- 12118376 TI - Loss of neurofibromatosis-1 and p19(ARF) cooperate to induce a multiple tumor phenotype. AB - Inactivation of the neurofibromatosis-1 (NF1) gene de-regulates RAS and cooperates with mutation or loss of the p53 tumor suppressor to induce tumorigenesis. p19(ARF) acts upstream of p53 in an oncogene checkpoint to induce apoptosis in response to activated RAS and other factors that stimulate proliferation. Therefore, we bred p19(ARF-/-) to NF1(+/-) mice to determine if loss of these genes collaborates in tumorigenesis. As expected from the embryonic lethality of NF1 null mice, no mice lacking both p19(ARF) and NF1 were born. Unexpectedly, the loss of one allele of NF1 did not greatly shorten the time to tumor formation in a p19(ARF) null background. The tumor types observed were characteristic of p19(ARF) null animals, not those associated with neurofibromatosis or those observed with NF1(+/-)/p53(+/-) mice. However, seven out of 12 animals developed multiple tumors, some with metastases. This multiple tumor phenotype was not previously observed with p19(ARF)-null mice and suggests a distinct form of cooperation between the loss of these tumor suppressors. PMID- 12118377 TI - A novel chromosomal translocation t(1;14)(q25;q32) in pre-B acute lymphoblastic leukemia involves the LIM homeodomain protein gene, Lhx4. AB - Chromosome 1q21-25 is one of the hotspots of chromosomal abnormalities including translocations and duplications in hematological malignancies. This would suggest that oncogene(s) reside in this region. We have cloned the junctional sequence of t(1;14)(q25;q32) in pre-B acute lymphoblastic leukemia cells by an inverse PCR method. A novel sequence was fused to the joining region of the immunoglobulin heavy chain gene. We confirmed this rearrangement by Southern blot analysis, genomic PCR and fluorescence in situ hybridization. We found a coding sequence which is homologous to the mouse Lhx4 cDNA sequence 17 kb from the breakpoint. The human Lhx4 gene encodes 390 amino-acids, including one tandem pair of LIM domains and one homeodomain. The human Lhx4 gene consists of six exons. Lhx4 protein is very homologous to human Lhx3 protein except in the N-terminal region. The transcripts of the Lhx4 gene were not detected in adult multiple tissues analysed by Northern blotting, but were detected in the leukemic cells carrying t(1;14)(q25;q32) by reverse-transcription PCR. The protein expression of Lhx4 in these leukemic cells was confirmed by Western blot analysis. Lhx4 activated the reporter gene carrying the mouse alpha-glycoprotein subunit promoter region, which is regulated by Lhx3. LIM protein and homeodomain protein genes are frequently involved in translocations of hematological malignancies. The Lhx4 gene is deregulated in the leukemic cells and Lhx4 protein may play an important role, possibly as an activator, in leukemogenesis. PMID- 12118378 TI - Connexin 43, but not connexin 32, is mutated at advanced stages of human sporadic colon cancer. AB - The membrane-spanning connexin proteins form microscopic intercellular channels that directly connect the cytoplasms of adjacent cells and as such have been implicated in maintenance of tissue homeostasis. They are considered to act as tumor suppressors since their function or expression is frequently aberrant in tumor cells. Several mechanisms appear to be involved in this, but irreversible mutational alterations have not yet been proved to be among them. In this study we have demonstrated for the first time that connexin 43 but not connexin 32 is specifically and quite frequently mutated in human colon sporadic adenocarcinomas. All tumor-associated mutations led to a shift of reading frame and were located in the multifunctional carboxyl-terminal domain of the protein. Expression of mutated connexin 43 protein was restricted to invasive structures of tumors. These findings suggest that mutational alterations of connexin 43 are involved in advanced stages of progression of human colon cancer towards malignancy. PMID- 12118379 TI - Gamma interferon down-regulates Fer and induces its association with inactive Stat3 in colon carcinoma cells. AB - Gamma interferon (IFN-gamma) is a regulator of cell growth, which suppresses the proliferation of HT-29 colon carcinoma cells. Here we show that in HT-29 cells IFN-gamma transiently increased the cellular level of the tyrosine kinase Fer, whose functioning was found to be essential for the proliferation of malignant cell-lines. The transient elevation in the level of Fer, was followed by its down regulation, an effect which was most prominent after 6-8 h of IFN-gamma treatment. Up- and down-regulation of Fer was paralleled by the activation and subsequent deactivation of Stat3, which is a potent oncogene and a putative substrate of the tyrosine kinase Fer. Moreover, IFN-gamma induced the association of Fer and Stat3 and the newly formed complex was most stable at the down regulated states of the two proteins. Formation of the Fer/Stat3 complex was accompanied by an attenuation in cell-cycle progression and accumulation of cells in the G1 phase. Thus, Fer and Stat3 are two proliferation-promoting factors whose down-regulation could contribute to the cytostatic activity of IFN-gamma in colon carcinoma cells. PMID- 12118380 TI - Impaired DNA damage-induced nuclear Rad51 foci formation uniquely characterizes Fanconi anemia group D1. AB - Fanconi anemia is a hereditary cancer susceptibility disorder characterized at the cellular level by spontaneous chromosomal instability and specific hypersensitivity to DNA cross-linking agents such as mitomycin C. This phenotype suggests a possible role for the Fanconi anemia proteins in the repair of DNA lesions induced by these agents, but the molecular mechanism underlying the defect in this disorder has not yet been identified. Here, we show that amongst eight so far identified complementation groups of Fanconi anemia, only fibroblasts derived from group D1 are defective in the formation of nuclear Rad51 foci after X-ray irradiation or mitomycin C treatment. This indicates that the FANCD1 gene product is uniquely involved in the assembly and/or stabilization of the Rad51 complex. Since DNA damage-induced Rad51 nuclear foci are thought to reflect repair of DNA double-strand breaks by homologous recombination, our results suggest that FANCD1 is likely to be involved in homologous recombination dependent repair. PMID- 12118381 TI - Cloning and characterization of a novel 90 kDa 'companion' auto-antigen of p62 overexpressed in cancer. AB - Recently our laboratory identified a cytoplasmic RNA-binding protein p62 which binds to and regulates the expression of IGF II mRNA. p62 was initially shown to be recognized by auto-antibodies in hepatocellular carcinoma (HCC) but now anti p62 has been described in diverse malignancies. p62 is uniformly expressed in fetal liver and prominently in 33% of HCC nodules, but not detectable in adult liver or normal tissue adjacent to HCC nodules. In this study, a 90 kDa protein (p90), auto-antibodies to which were found associated with anti-p62 responses in the same HCC patient group, was identified by cDNA expression cloning. Indirect immunofluorescence showed that, like p62, p90 localized to the cytoplasm in cultured cells and mouse fetal, but not adult liver. Among 11 human gastric cancer tissues examined, p90 was overexpressed in six (55%). Together with other cancer associated auto-antibodies such as anti-p53, anti-p62, anti-Koc, and anti CENP-F, auto-antibodies to p90 represent a new marker for tumors such as HCC and gastric cancer. Our data support the working hypothesis that auto-antibody production in cancer may be directly linked to aberrant auto-antigen expression. PMID- 12118382 TI - Delineation and candidate gene mutation screening of the 18q22 minimal region of deletion in head and neck squamous cell carcinoma. AB - The 18q chromosome arm is frequently lost in advanced head and neck squamous cell carcinoma. Twenty-four microsatellite markers located on chromosome 18q were genotyped in 145 primary tumors and 10 cell lines in order to identify putative tumor suppressor genes implicated in tumor progression. Two different minimal common regions of loss (MCRL) were identified at 18q22 and 18q23 respectively. To refine and delineate boundaries of an homozygous deletion found in one cell line, 44 extra markers located at 18q22 were analysed and the homozygous deletion was precisely defined within a critical region of 4.9 Mb. Four known genes (CDH7, CDH19, DNAM-1, FLJ23594) located in this critical region and two EST clusters (Hs.96900, Hs.98628) were selected for further investigations. For these six genes, genomic structures were established, somatic mutations were screened in 20 HNSCC and 10 cell lines and transcription levels were determined in eight cell lines. No somatic mutations were found in any of the candidate genes analysed (57 coding exons). However, differential transcription levels were observed for CDH19 and Hs.96900 in head and neck cancer cell lines supporting their putative involvement through down regulation mechanisms in head and neck cancer progression. PMID- 12118383 TI - Isolation and characterization of a novel gene, hRFI, preferentially expressed in esophageal cancer. AB - hTID1, a human homologue of Drosophila tumor suppressor, I(2)tid regulates the release of cytochrome c from mitochondria and subsequent alteration of caspase-3 activity on apoptosis induced by exogenous stimuli, such as tumor necrosis factor alpha and mitomycin C. To search for an interacting molecule with hTid1, we applied two-hybrid yeast screening and isolated a novel gene, which encodes a 46 kDa protein of 373 residues. Within the deduced amino acid sequence, a region showing homology to the Ring Finger domain of X-linked inhibitor of apoptosis protein was identified and the gene was designated as hRFI, standing for human Ring Finger homologous to IAP type. A 2.0 kb hRFI transcript was ubiquitously expressed in all human tissues as well as several cancer cell lines examined. Northern blot analysis showed that in 70% (14 out of 20) of esophageal cancer patients, expression of hRFI in cancerous regions was two or more times higher than in the corresponding normal tissues. HeLa cells transfected with hRFI construct exhibited a tendency to resist TNF-alpha induced apoptosis, suggesting an anti-apoptotic function of the hRFI product. Finally, hRFI protein was shown to be cleaved within the DEDD sequence spanning residues 230-233 by caspase-3 during the apoptotic induction. PMID- 12118384 TI - Intensity-modulated radiation therapy: a clinical perspective. Introduction. PMID- 12118385 TI - Dose-volume specification: new challenges with intensity-modulated radiation therapy. AB - It has long been recognized that the specification of volumes and doses is an important issue for radiation oncology. Although in any individual center, policies and procedures of treatment delivery may be well understood by staff, reporting of treatment techniques in the archival literature in an unambiguous manner has been found to be less than desirable in many instances. For clinical studies utilizing three-dimensional conformal radiation therapy (3D-CRT), and even more so, intensity-modulated radiation therapy (IMRT), the situation has become even more complex. 3D-CRT and IMRT are now recognized to be more sensitive to geometric uncertainties than conventional radiation therapy because of their ability to create sharper dose gradients around target volumes and organs at risk (OARs). This article reviews the current status of specifying target volumes and doses for 3D-CRT and IMRT, and discusses some of the pertinent issues regarding the use of recommendations in Reports 50 and 62 of the International Commission on Radiation Units and Measurements (ICRU) in this task. It is imperative that physician and physicist fully appreciate the need to account for clinical and spatial uncertainties in the planning and delivery of cancer patients' treatment, paying even more attention to these issues for those cases in which 3D-CRT and/or IMRT is used. A brief review of the reporting requirements for Radiation Therapy Oncology Group (RTOG) 3D-CRT and IMRT protocols is also presented. PMID- 12118386 TI - Issues in optimization for planning of intensity-modulated radiation therapy. AB - The clinical use of intensity-modulated radiation therapy (IMRT) is expanding rapidly in academic and, more recently, in community-based radiotherapy centers due to a high level of clinician interest, improving reimbursement patterns, and the availability of the tools required to plan and deliver IMRT plans. These tools include inverse planning optimization algorithms and linear accelerator control systems with automated, multifield delivery capabilities. The hazards of this new technology are due primarily to the nonintuitive nature of the inverse planning process and the highly complex methods of delivery required for IMRT dose delivery. Important efforts are being made to define the required quality assurance for these computer-optimized IMRT plans and to find ways to reduce their complexity without reducing the quality of the resulting plans. By minimizing the complexity of these dose plans, one also minimizes the treatment time and the probability of dose delivery errors. Methods of optimization and evaluation of dose plans and practical considerations in inverse planning are discussed. In addition, this article points out the potential hazards of inverse planned IMRT and discusses methods by which the complexity of these plans might be reduced. PMID- 12118387 TI - Quality assurance of intensity-modulated radiotherapy. AB - Intensity-modulated radiation therapy (IMRT) requires the use of inverse treatment planning and nonuniform fluence beams delivered by a series of complex radiation portals. The quality assurance procedures for conventional three dimensional conformal radiation therapy (3D-CRT) have been developed and are in worldwide clinical use, but the more complex nature of IMRT limits the application of much of the quality assurance (QA) procedures developed for IMRT. Although consensus has not yet been reached regarding which procedures will eventually become recommended by official organizations, the field is rapidly coming to agreement on a basic set of procedures. This manuscript describes some of the novel techniques recently developed for IMRT QA, both for the validation of the calculated dose distribution and for assuring that the dose distribution reaches its intended targets. PMID- 12118388 TI - Intensity-modulated radiation therapy for prostate cancer. AB - Intensity-modulated radiation therapy (IMRT) represents a new paradigm in radiation treatment planning and delivery for treatment of prostate cancer with enormous potential. Preliminary data indicate that this highly conformal treatment technique can effectively reduce acute and late-occurring toxicities, improving the quality of life of the treated patient and serving as the optimal dose escalation tool. IMRT produces radiation distributions capable of delivering different dose prescriptions to multiple target sites, providing a new opportunity for differential dose painting to increase the dose selectively to specific, image-defined regions within the prostate. Clinical trials will be necessary to define more clearly the true extent of improved tumor control and reduction in normal tissue complications with IMRT in the treatment of prostate cancer. PMID- 12118389 TI - Intensity-modulated radiation therapy for head and neck cancer: emphasis on the selection and delineation of the targets. AB - The head and neck contain many critical, noninvolved structures in close vicinity to the targets. The tightly conformal doses produced by intensity-modulated radiation therapy (IMRT), and the lack of internal organ motion in the head and neck, provide the potential for organ sparing and improved tumor irradiation. Many studies of treatment planning for head and neck cancer have demonstrated the dosimetric superiority of IMRT over conventional techniques in these respects. The initial results of clinical studies demonstrate reduced xerostomia. They suggest an improvement in tumor control, which needs to be verified in larger studies and longer follow-up. Critical issues for successful outcome of head and neck IMRT are accurate selection of the neck lymph nodes that require adjuvant treatment, and accurate delineation on the planning computed tomography (CT) of the lymph-node bearing areas and subclinical disease adjoining the gross tumor. This review emphasizes these topics and provides some guidelines. PMID- 12118390 TI - Clinical aspects of intensity-modulated radiotherapy in the treatment of breast cancer. AB - In recent years, interest has grown throughout the radiotherapy community in investigation and clinical application of intensity-modulated radiation therapy (IMRT) for adjuvant treatment of breast cancer. IMRT removes the usual reliance on flat (or uniform-intensity) radiation fields, and instead replaces that simple paradigm with a variable-intensity pattern that is usually determined with the aid of a computerized optimization algorithm. The main goal of much IMRT and optimization work is the delivery of more conformal plans to the patient. Thus, IMRT has the potential to improve target coverage and reduce inhomogeneities observed within the breast (and regional lymph nodes) that are obtained with standard plans. Furthermore, IMRT may be able to reduce doses delivered to the heart and lungs, and may potentially minimize further the probability of complications from radiotherapy. PMID- 12118391 TI - Clinical application of intensity-modulated radiotherapy for locally advanced cervical cancer. AB - Intensity-modulated radiotherapy (IMRT) offers technical advantages over conventional external beam radiotherapy (CXRT) that might prove clinically advantageous in the management of gynecologic malignancies. Especially in the case of locally advanced cervical cancer, IMRT provides an opportunity to improve the therapeutic ratio by allowing a selective combination of normal tissue dose reduction and/or concomitant integrated boost dose to the tumor. The clinical and biologic rationale for IMRT in this setting is presented here, and pertinent technical considerations such as the delineation of relevant clinical and planning target volumes are discussed. The capacity for IMRT-mediated normal tissue sparing is illustrated by example and review of the literature. Furthermore, for a small cohort of patients with locally advanced or recurrent cervical cancer treated with concomitant integrated boost IMRT and concurrent chemotherapy, preliminary clinical observations of toxicity and tumor response are presented. Concomitant integrated boost IMRT appears clinically tolerable and efficacious in this setting, and formal clinical investigation is warranted as a means of exploiting the fraction-size dependence of radiosensitizers in common clinical use. PMID- 12118392 TI - Intensity-modulated radiation therapy: the inverse, the converse, and the perverse. AB - Intensity-modulated radiation therapy (IMRT) is a refinement of current radiotherapy techniques rather than a major breakthrough. The term IMRT includes several different techniques that all share with classical arc therapy the principle of using multiple fields to reduce the dose to normal tissues, but integrating to a higher dose throughout the tumor volume itself. This paper reviews not only the putative upside but also the downside of the development of IMRT. Theoretical, practical, and cost considerations, both positive and negative, are discussed. There are several issues to be considered, but the most important perversely predict a significant increase in radiation-induced neoplasms, resulting not only from larger volumes of tissue exposed to more modest but still mutagenic doses, but also from a significant increase in total body dose from leakage, because the beam is typically on for a considerably longer period of time than is conventional. A plea is made for radiation oncologists to maintain a strong biologic and cellular orientation as oncology rapidly becomes more molecular in its orientation. PMID- 12118394 TI - Hepatorenal syndrome. AB - The hepatorenal syndrome is defined as functional renal failure in advanced chronic or acute liver disease with portal hypertension. Morphologic abnormalities of the kidneys are frequently absent and tubular function is preserved. Patients with the hepatorenal syndrome are characterized by progressive splanchnic and systemic vasodilation and decreased effective arterial blood volume. Compensatory activation of vasoconstrictory systems maintains systemic hemodynamic stability but causes progressive afferent renal vasoconstriction, leading to reduction of glomerular filtration rate. Renal failure may be rapidly progressive (type I hepatorenal syndrome, frequently associated with spontaneous bacterial peritonitis) or may develop more slowly (type II). Orthotopic liver transplantation is the best current treatment and leads to a gradual recovery of renal function in the vast majority of patients. Because mortality of type I hepatorenal syndrome is excessive, supportive treatment by vasoconstrictor drugs, transjugular intrahepatic portosystemic shunt, and renal replacement therapy has been investigated to achieve stability until transplantation. The definite role of these promising developments, however, is still uncertain, emphasizing the need for large prospective multicentric investigations. PMID- 12118395 TI - Beyond hepatorenal syndrome: glomerulonephritis in patients with liver disease. AB - Liver disease is frequently associated with renal abnormalities. In liver cirrhosis, impaired hepatic clearance of immune complexes leads to their trapping in the kidney, causing the lesions of hepatic immunoglobulin A (IgA) nephropathy and hepatic glomerulosclerosis. Chronic hepatitis C virus (HCV) infection can induce cryoglobulinemia type II with membranoproliferative glomerulonephritis, whereas chronic hepatitis B virus (HBV) infection may cause membranous nephropathy, or, more rarely, polyarteritis nodosa. Treatment aims at eliminating the viral infection, in HCV infection with interferon alfa and ribavirin and in HBV infection with interferon alfa or lamivudine. Short-term immunosuppressive treatment may be indicated in patients with severe inflammation. In alpha1 antitrypsin deficiency with liver disease a membranoproliferative type of glomerulonephritis can occur. In addition, partial or complete deficiency is frequently observed in patients with c-ANCA-positive systemic vasculitis. PMID- 12118396 TI - Kidney and liver involvement in cryoglobulinemia. AB - Cryoglobulinemia is a pathological condition characterized by the presence in the blood of a group of proteins (cryoglobulins) showing the common property of precipitating from cooled serum. Mixed cryoglobulins (MCs) are composed of a polyclonal immunoglobulin G (IgG) bound to another immunoglobulin that acts as an anti-IgG rheumatoid factor; 2 types of MCs can be identified. The origin is not clear in 30% of all MCs, and this subset of cryoglobulinemias is called "essential." The great majority (up to 90%) of patients with essential MC (EMC) of both types has been related to HCV infection. The renal involvement in essential MC has been observed in 2% to 50% of patients in reported series. A well-characterized pattern of glomerular disease termed "cryoglobulinemic glomerulonephritis" is present in individuals with type-II EMCs in serum and IgMk RF being the most frequent monoclonal component. Aspecific and infrequent glomerular lesions occur in EMC patients with type III cryoglobulins. The most frequent histologic picture of cryoglobulinemic GN is that of membranoproliferative GN (MPGN) with subendothelial deposits. However, cryoglobulinemic GN may have some distinctive features that differentiate it from idiopathic type-I MPGN. Evidence of liver involvement has been found in 60% to 80% of EMC patients. Recent data support the notion that the stage of liver disease in patients with type II or III MC has association with the prevalence of cryoglobulinemia. Few controlled trials have been published on the treatment of HCV-related MC; in addition, the majority of the patients enrolled in these trials had an unclear renal involvement. Interferon (IFN) is an effective drug for the treatment of patients with HCV-associated MC and cryoglobulinemic GN. In the presence of acute cryoglobulinemic GN, IFN does not prevent progression of renal damage; combination therapy with cytotoxic ad anti-inflammatory drugs, and sometimes plasma exchange, is recommended. Prospective controlled trials are underway on this purpose. PMID- 12118397 TI - Kidney and liver involvement in monoclonal light chain disorders. AB - Monoclonal light chains (LCs) are responsible for a wide spectrum of renal and hepatic diseases, that above all include amyloid light-chain (AL) amyloidosis and light chain deposition disease (LCDD). Amyloid deposits stain for Congo red on light microscopy and have fibrillar aspect on electron microscopy, whereas deposits in LCDD are positive using monotypic LCs on immunofluorescence and have a granular aspect on electron microscopy. Sometimes fibrillar and granular deposits are observed in the same organ or in different organs of the same patient. Kidney and liver involvement is a frequent finding, both in primary amyloidosis (AL amyloidosis) or in LCDD. Renal manifestations include proteinuria, nephrotic syndrome, and progressive renal failure. End-stage renal disease requiring dialysis is observed in about 20% of patients with AL amyloidosis and in 70% of patients with LCDD. The mean survival time is about 12 to 18 months in AL amyloidosis and 34 months in LCDD. The most important prognostic factor is severe cardiac involvement, which reduces the mean survival to only 6 months. Hepatic manifestations include hepatomegaly, portal hypertension, ascites, intrahepatic cholostatic jaundice, and hepatic insufficiency. The mean survival of patients with liver damage is 14 months, but it is reduced to 5 months in patients with cholostatic jaundice. Contemporary kidney and liver involvement is usually observed on histologic examination, less frequently as clinical manifestation. No specific treatment exists for AL amyloidosis and LCDD, and the prognosis remains severe. The aim of treatment is to suppress proliferation of the abnormal clone of plasma cells and remove tissue deposits. The regimens, including melphalan-prednisone (MP) or vincristine doxorubicin-dexamethasone (VAD), are used both in AL amyloidosis or in LCDD with some effectiveness. New approaches, especially the use of 4'-iodo 4'deoxydoxorubicin, could achieve better results. Dialysis seems to not worsen the outcome in both diseases because survival of patients on dialysis is not different from that of patients not reaching uremia. Also, kidney and liver transplantation is effective, though amyloidosis or LCDD may occur in transplanted organs. The most interesting therapeutic approach is autologous blood stem-cell transplantation, which may produce a complete remission of the plasma-cell dyscrasia and a substantial improvement of clinical manifestations related to LC deposits. PMID- 12118398 TI - HCV infection and hemodialysis. AB - Hepatitis C virus (HCV) infections are frequent in hemodialyzed patients and are mainly related to transfusions and nosocomial contamination. HCV-related infection may result in cirrhosis in 10% of dialysis patients and is worsened by transplantation because of the immunosuppressive therapy for prevention of graft rejection. Because there is a risk for significant liver disease and because cirrhosis contraindicates a renal transplantation, a liver biopsy should be performed early in HCV-RNA positive hemodialysis patients to evaluate histologic impact of the liver disease. A combined liver-kidney transplantation should be discussed in dialysis patients with cirrhosis. Standard alpha-interferon is the only treatment for HCV in dialysis patients because ribavirin is contraindicated by a high risk for hemolytic anemia. It leads to an overall 30% rate of sustained viral eradication. It is indicated in dialysis patients with acute hepatitis C, significant liver disease (fibrosis score > or =2), or symptomatic cryoglobulinemia, and to candidates for renal transplantation, whatever the severity of the liver disease. Indeed, alpha-interferon is contraindicated in kidney recipients given the risk for rejection. Preventive treatment for HCV is only respect for universal hygiene rules in the dialysis setting because there is no available vaccine. PMID- 12118399 TI - Hepatitis B virus infection in hemodialysis patients. AB - Patients with chronic renal failure on hemodialysis suffer from impaired immune defense mechanisms. A defect in costimulatory signalling from antigen-presenting cells (APCs) for the antigen-specific activation of T cells leads to immune incompetence, particularly toward viral infections. The epidemiologic situation of dialysis patients who are treated in centers together with other immunocompromised patients and who need blood access 3 times weekly places them at a high risk for viral hepatitis B. Clinically, the infection shows a mild and subclinical, often anicteric, course, leading to chronic infection in the majority of cases. Typical consequences such as cirrhosis and hepatocellular carcinoma may occur, however, seem to be less frequent than in hepatitis B infected persons with normal renal function. Protection by vaccination is also hampered by the immune defect. Nevertheless, a high percentage of vaccinated patients is still desirable for infection control. Treatment of hepatitis B in dialysis patients is difficult because interferon-alfa is less effective and frequently leads to side effects. Lamivudine may become a future alternative, however, current experience is limited. PMID- 12118400 TI - Hepatitis B and C in peritoneal dialysis patients. AB - In most countries the prevalence of hepatitis B virus (HBV) and hepatitis C virus (HCV) infection in peritoneal dialysis (PD) patients is lower than in hemodialysis (HD) patients. Besides a history of blood transfusions, previous HD is an important risk factor for developing HCV infection in PD patients. Many HCV positive patients already are anti-HCV-positive before initiation of PD. Seroconversion to HCV during PD treatment is, therefore, a rare event. HCV RNA in serum is positive in 53% to 84% of anti-HCV-positive patients. Routine screening for HBV and HCV by using a second- or third-generation enzyme-linked immunosorbent assay (ELISA) should be performed in PD patients every 6 months. Asymptomatic HBV and HCV infection may be detected by elevation of transaminases, but lower cut-off levels should be preferred in PD patients. Prophylactic strategies include hygienic measures and HBV vaccination. The staff should be aware of the infectiousity of the PD effluent, especially in hepatitis B surface antigen (HBsAg)-positive patients. Because of the smaller number of required blood transfusions and the increased use of home therapy, which reduces the risk for environmental contamination, PD is considered to be an important strategy for prevention of hepatitis in end-stage renal disease patients. PMID- 12118401 TI - Hepatitis C virus-positive patients on the waiting list for transplantation. AB - Hepatitis C virus (HCV) infection is a common problem in renal transplant patients, associated with an increase in morbidity and mortality. HCV infection is associated with a lower graft and patient survival. The problem of HCV infection is the increase in viral load and liver transaminases after renal transplantation secondary to immunosuppressive therapy. After renal transplantation, interferon therapy is not recommended because of the risk for acute rejection and acute nephritis. In this context, it is absolutely necessary to consider the evaluation and treatment of HCV infection during the dialysis period. Several studies have defined the benefits of interferon therapy in dialysis patients, with rates of maintenance response significantly higher than in the general population. The difference in the pharmacokinetic profile of interferon in dialysis patients could justify its higher efficacy. PMID- 12118402 TI - Hepatitis C virus infection and kidney transplantation. AB - Hepatitis C virus (HCV) infection is an important problem in the patient with end stage renal disease. After transplantation, liver disease is more frequent in HCV positive patients than in HCV-negative patients. In the long run, this leads to important liver complications. The patients have a higher risk for developing proteinuria and infections. Long-term patient and graft survival rates are lower in HCV-positive patients than in HCV-negative graft recipients. Mortality is higher, mainly as a result of liver disease and infections. Despite this, transplantation is the best option for the HCV-positive patient with end-stage renal disease. Transplantation of HCV-positive kidneys should be offered to HCV positive recipients in whom HCV RNA is detected in the serum. Finally, several measures after transplantation minimize the consequences of HCV infection. Adjustment of immunosuppression and careful follow-up in the outpatient clinic for early detection of proteinuria, infection, or worsening of liver disease are mandatory. PMID- 12118403 TI - Chronic hepatitis B virus infection in renal transplant recipients. AB - Although in Western Europe and North America the prevalence of chronic hepatitis B virus (HBV) infection has declined in patients awaiting renal transplantation, it remains a relevant clinical problem, mainly in patients with a long history of renal replacement therapy (RRT) who may have been infected many years ago. At the same time, a significant proportion of renal transplant recipients (RTR) is at risk for HBV infection in areas with endemic HBV. HBV infection may increase morbidity and mortality in RTR. The majority of long-term studies reported reduced patient survival compared with hepatitis B surface antigen (HBsAg) negative RTR. The risk for morbidity and mortality of HBsAg-positive RTR transplantation is probably related to the extent of pretransplant liver disease. A thorough evaluation, including liver biopsy in many patients, is required to assess the individual HBsAg-positive patient's risk-benefit ratio. The influence of immunosuppressive therapy on HBV replication and HBV-associated complications is not well established and for clinical practice individually tailored immunosuppression is recommended in HBsAg-positive RTR. Careful screening for HBV reactivation in HBsAg-positive RTR and a regular clinical follow-up after renal transplantation (RTX) including liver sonography is required for early detection of HBV-associated complications. With the availability of new antiviral drugs, new options for pre- and posttransplant therapy might improve the prognosis of RTR with chronic HBV infection. PMID- 12118404 TI - A new method to localize and test the significance of incongruence: detecting domain shuffling in the nuclear receptor superfamily. AB - When a data set is partitioned, the resulting subsets may contain phylogenetically conflicting signals if they have different evolutionary histories. In a data set with many taxa, a single taxon that contains multiple phylogenetic histories may result in global incongruence, but no methods are available in a parsimony framework to localize incongruence to specific clades in a phylogeny or to test the significance of incongruence on a local scale. Here we present a new method to quantify the conflict between data partitions for any clade in a phylogeny and to test the statistical significance of that conflict by using a metric called the local incongruence length difference. We apply this method to the evolutionary history of the nuclear receptor superfamily, a large group of transcriptional regulators that play essential roles in metazoan development and physiology. All nuclear receptors are composed of several discrete domains, including one that binds to DNA response elements on specific target genes and another that binds to the appropriate ligand. We have performed combined and separate phylogenetic analyses of these two domains and have tested the hypothesis that nuclear receptors evolved by a simple process of lineage splitting and divergence, without domain shuffling or other forms of sequence transfer between proteins. Our analysis indicates that significant conflict exists between the partitined domains at a few nodes on the tree, suggesting that several groups of receptors are "hybrid proteins" formed by domain shuffling or other forms of sequence transfer between more ancient nuclear receptors. PMID- 12118405 TI - More taxa or more characters revisited: combining data from nuclear protein encoding genes for phylogenetic analyses of Noctuoidea (Insecta: Lepidoptera). AB - A central question concerning data collection strategy for molecular phylogenies has been, is it better to increase the number of characters or the number of taxa sampled to improve the robustness of a phylogeny estimate? A recent simulation study concluded that increasing the number of taxa sampled is preferable to increasing the number of nucleotide characters, if taxa are chosen specifically to break up long branches. We explore this hypothesis by using empirical data from noctuoid moths, one of the largest superfamilies of insects. Separate studies of two nuclear genes, elongation factor-1 alpha (EF-1 alpha) and dopa decarboxylase (DDC), have yielded similar gene trees and high concordance with morphological groupings for 49 exemplar species. However, support levels were quite low for nodes deeper than the subfamily level. We tested the effects on phylogenetic signal of (1) increasing the taxon sampling by nearly 60%, to 77 species, and (2) combining data from the two genes in a single analysis. Surprisingly, the increased taxon sampling, although designed to break up long branches, generated greater disagreement between the two gene data sets and decreased support levels for deeper nodes. We appear to have inadvertently introduced new long branches, and breaking these up may require a yet larger taxon sample. Sampling additional characters (combining data) greatly increased the phylogenetic signal. To contrast the potential effect of combining data from independent genes with collection of the same total number of characters from a single gene, we simulated the latter by bootstrap augmentation of the single-gene data sets. Support levels for combined data were at least as high as those for the bootstrap-augmented data set for DDC and were much higher than those for the augmented EF-1 alpha data set. This supports the view that in obtaining additional sequence data to solve a refractory systematic problem, it is prudent to take them from an independent gene. PMID- 12118407 TI - Evaluating trans-tethys migration: an example using acrodont lizard phylogenetics. AB - A phylogenetic tree for acrodont lizards (Chamaeleonidae and Agamidae) is established based on 1434 bases (1041 informative) of aligned DNA positions from a 1685-1778 base pair region of the mitochondrial genome. Sequences from three protein-coding genes (ND1, ND2, and COI) are combined with sequences from eight intervening tRNA genes for samples of 70 acrodont taxa and two outgroups. Parsimony analysis of nucleotide sequences identifies eight major clades in the Acrodonta. Most agamid lizards are placed into three distinct clades. One clade is composed of all taxa occurring in Australia and New Guinea; Physignathus cocincinus from Southeast Asia is the sister taxon to the Australia-New Guinea clade. A second clade is composed of taxa occurring from Tibet and the Indian Subcontinent east through South and East Asia. A third clade is composed of taxa occurring from Africa east through Arabia and West Asia to Tibet and the Indian Subcontinent. These three clades contain all agamid lizards except Uromastyx, Leiolepis, and Hydrosaurus, which represent three additional clades of the Agamidae. The Chamaeleonidae forms another clade weakly supported as the sister taxon to the Agamidae. All eight clades of the Acrodonta contain members occurring on land masses derived from Gondwanaland. A hypothesis of agamid lizards rafting with Gondwanan plates is examined statistically. This hypothesis suggests that the African/West Asian clade is of African or Indian origin, and the South Asian clade is either of Indian or Southeast Asian origin. The shortest tree suggests a possible African origin for the former and an Indian origin for the latter, but this result is not statistically robust. The Australia-New Guinea clade rafted with the Australia-New Guinea plate and forms the sister group to a Southeast Asian taxon that occurs on plates that broke from northern Australia New Guinea. Other acrodont taxa are inferred to be associated with the plates of Afro-Arabia and Madagascar (Chameleonidae), India (Uromastyx), or southeast Asia (Hydrosaurus and Leiolepis). Introduction of different biotic elements to Asia by way of separate Gondwanan plates may be a major theme of Asian biogeography. Three historical events may be responsible for the sharp faunal barrier between Southeast Asia and Australia-New Guinea, known as Wallace's line: (1) primary vicariance caused by plate separations; (2) secondary contact of Southeast Asian plates with Eurasia, leading to dispersal from Eurasia into Southeast Asia, and (3) dispersal of the Indian fauna (after collision of that subcontinent) to Southeast Asia. Acrodont lizards show the first and third of these biogeographic patterns and anguid lizards exhibit the second pattern. Modern faunal diversity may be influenced primarily by historical events such as tectonic collisions and land bridge connections, which are expected to promote episodic turnover of continental faunas by introducing new faunal elements into an area. Repeated tectonic collisions may be one of the most important phenomena promoting continental biodiversity. Phylogenetics is a powerful method for investigating these processes. PMID- 12118406 TI - Invariable sites models and their use in phylogeny reconstruction. AB - Phylogenetic inference is well known to be problematic if both long and short branches occur together in the underlying tree. With biological data, correcting for this problem may require simultaneous consideration for both substitution biases and rate heterogeneity between lineages and across sequence positions. A particular form of the latter is the presence of invariable sites, which are well known to mislead estimation of genetic divergences. Here we describe a capture recapture method to estimate the proportion of invariable sites in an alignment of amino acids or nucleotides. We use it to investigate phylogenetic signals in 18S ribosomal DNA sequences from Holometabolus insects. Our results suggest that, as taxa diverged, their 18S rDNA sequences have altered in both their distribution of sites that can vary as well as in their base compositions. PMID- 12118408 TI - Evolution and phylogenetic information content of mitochondrial genomic structural features illustrated with acrodont lizards. AB - DNA sequences from 195 squamate reptiles indicate that mitochondrial gene order is the most reliable phylogenetic character establishing monophyly of acrodont lizards and of the snake families Boidae, Colubridae, and Viperidae. Gene order shows no evidence of evolutionary parallelisms or reversals in these taxa. Derived secondary structures of mitochondrial tRNAs also prove to be useful phylogenetic characters showing no reversals. Parallelisms for secondary structures of tRNAs are restricted to deep lineages that are separated by at least 200 million years of independent evolution. Presence of a stem-and-loop structure between the genes encoding tRNA(Asn) and tRNA(Cys), where the replication origin for light-strand synthesis is typically located in vertebrate mitochondrial genomes, is found to undergo at least three and possibly as many as seven evolutionary shifts, most likely parallel losses. This character is therefore a less desirable phylogenetic marker than the other structural changes examined. Sequencing regions that contain multiple genes, including tRNA genes, may be preferable to the common practice of obtaining single-gene fragments for phylogenetic inference because it permits observation of major structural changes in the mitochondrial genome. Such characters may occasionally provide phylogenetic information on relatively short internal branches for which base substitutional changes are expected to be relatively uninformative. PMID- 12118409 TI - Phylogenetic relationships of agaric fungi based on nuclear large subunit ribosomal DNA sequences. AB - Phylogenetic relationships of mushrooms and their relatives within the order Agaricales were addressed by using nuclear large subunit ribosomal DNA sequences. Approximately 900 bases of the 5' end of the nucleus-encoded large subunit RNA gene were sequenced for 154 selected taxa representing most families within the Agaricales. Several phylogenetic methods were used, including weighted and equally weighted parsimony (MP), maximum likelihood (ML), and distance methods (NJ). The starting tree for branch swapping in the ML analyses was the tree with the highest ML score among previously produced MP and NJ trees. A high degree of consensus was observed between phylogenetic estimates obtained through MP and ML. NJ trees differed according to the distance model that was used; however, all NJ trees still supported most of the same terminal groupings as the MP and ML trees did. NJ trees were always significantly suboptimal when evaluated against the best MP and ML trees, by both parsimony and likelihood tests. Our analyses suggest that weighted MP and ML provide the best estimates of Agaricales phylogeny. Similar support was observed between bootstrapping and jackknifing methods for evaluation of tree robustness. Phylogenetic analyses revealed many groups of agaricoid fungi that are supported by moderate to high bootstrap or jackknife values or are consistent with morphology-based classification schemes. Analyses also support separate placement of the boletes and russules, which are basal to the main core group of gilled mushrooms (the Agaricineae of Singer). Examples of monophyletic groups include the families Amanitaceae, Coprinaceae (excluding Coprinus comatus and subfamily Panaeolideae), Agaricaceae (excluding the Cystodermateae), and Strophariaceae pro parte (Stropharia, Pholiota, and Hypholoma); the mycorrhizal species of Tricholoma (including Leucopaxillus, also mycorrhizal); Mycena and Resinomycena; Termitomyces, Podabrella, and Lyophyllum; and Pleurotus with Hohenbuehelia. Several groups revealed by these data to be nonmonophyletic include the families Tricholomataceae, Cortinariaceae, and Hygrophoraceae and the genera Clitocybe, Omphalina, and Marasmius. This study provides a framework for future systematics studies in the Agaricales and suggestions for analyzing large molecular data sets. PMID- 12118410 TI - Phylogenetics of flowering plants based on combined analysis of plastid atpB and rbcL gene sequences. AB - Following (1) the large-scale molecular phylogeny of seed plants based on plastid rbcL gene sequences (published in 1993 by Chase et al., Ann. Missouri Bot. Gard. 80:528-580) and (2) the 18S nuclear phylogeny of flowering plants (published in 1997 by Soltis et al., Ann. Missouri Bot. Gard. 84:1-49), we present a phylogenetic analysis of flowering plants based on a second plastid gene, atpB, analyzed separately and in combination with rbcL sequences for 357 taxa. Despite some discrepancies, the atpB-based phylogenetic trees were highly congruent with those derived from the analysis of rbcL and 18S rDNA, and the combination of atpB and rbcL DNA sequences (comprising approximately 3000 base pairs) produced increased bootstrap support for many major sets of taxa. The angiosperms are divided into two major groups: noneudicots with inaperturate or uniaperturate pollen (monocots plus Laurales, Magnoliales, Piperales, Ceratophyllales, and Amborellaceae-Nymphaeaceae-Illiciaceae) and the eudicots with triaperturate pollen (particularly asterids and rosids). Based on rbcL alone and atpB/rbcL combined, the noneudicots (excluding Ceratophyllum) are monophyletic, whereas in the atpB trees they form a grade. Ceratophyllum is sister to the rest of angiosperms with rbcL alone and in the combined atpB/rbcL analysis, whereas with atpB alone, Amborellaceae, Nymphaeaceae, and Illiciaceae/Schisandraceae form a grade at the base of the angiosperms. The phylogenetic information at each codon position and the different types of substitutions (observed transitions and transversions in the trees vs. pairwise comparisons) were examined; taking into account their respective consistency and retention indices, we demonstrate that third-codon positions and transitions are the most useful characters in these phylogenetic reconstructions. This study further demonstrates that phylogenetic analysis of large matrices is feasible. PMID- 12118411 TI - Simple but fundamental limitations on supertree and consensus tree methods. PMID- 12118412 TI - Gaps as characters in sequence-based phylogenetic analyses. PMID- 12118413 TI - [The dental examination not by one's own dentist]. PMID- 12118414 TI - [Dental examination of course by one's own dentist]. PMID- 12118415 TI - [Obligation to provide information in dentistry: a problem?]. PMID- 12118416 TI - [Is bruxism the cause of complaints of TMD?]. PMID- 12118418 TI - Population data on X chromosome short tandem repeat loci HPRTB and AR in Taiwan. AB - This report contains the results of population studies on the X chromosome STR HPRTB and AR carried out in Taiwan. The numbers of unrelated individuals were 428: female 143 and male 285 for HPRTB locus, and 416: female 142 and male 274 for AR locus. PMID- 12118419 TI - Distribution of alleles of 15 STR loci of the Powerplex 16 Multiplex system in four predominant population groups of South India. AB - Allele distribution of the 15 STR loci of Powerplex 16 Multiplex System were studied in four predominant population groups of South India for evaluating their significance in human identification and population study: Iyengar Brahmin (65), Gowda (59), Lingayat (98) and Muslim (45) from the state of Karnataka. The loci analyzed are--D3S1358, THOI, D2ISI I, D18S51, D5S818, Penta E, D13S317, D7S820, D16S539, CSFIPO, Penta D, vWA, D8S179, TPOX and FGA. Out of 15 STR loci Penta D and D18S51 were found highly polymorphic in the studied populations. PMID- 12118420 TI - Allele frequencies for STR loci of the Powerplex 16 multiplex system in five endogamous populations of India. AB - Allele frequencies for the 15 STR locus of PowerPlex 16 were analyzed in 95 healthy unrelated individuals belonging to five important population groups inhabiting different part of India. Fifteen loci studied are Penta E, D18S51, D21S11, THO1,D3S1358, FGA, TPOX, D8S1179, vWA, Amelogenin, Penta D, CSF1PO, D16S539, D7S820, D13S317 and D5S818. In addition of all tetra nucleotide loci, two penta nucleotide loci Penta D and Penta E of the studied system are also found highly polymorphic in all the five studied populations of India. These loci are found highly informative in solving paternity cases and other forensic testing in studied population. PMID- 12118422 TI - [Mortality of Swiss dentists]. AB - In this study, the survival of Swiss dentists is investigated using a cohort of 5749 members of the Swiss Dental Association (SSO). The study shows that the male dentists of this cohort have at any age a lower mortality than the Swiss male population in general. However, a similar statement cannot be made for the Swiss female dentists as there is uncertainty about their reasons for leaving the cohort. In addition, the death causes of 446 deceased dentists were studied with respect to a connection of death causes and exposure to amalgam. There was no evidence of higher mortality by causes commonly associated with amalgam exposure, in spite of the fact that these dentists had mostly practiced in times of fairly high amalgam exposure. By its nature, however, this study cannot provide information on non-lethal diseases possibly caused by amalgam exposure. On the whole, Swiss dentists present themselves as rather healthy and long-living professionals. PMID- 12118423 TI - [Reconstruction of the anterior floor of the mouth with nasolabial flaps. Report of 10 years' experience]. AB - The aim of this study was to analyse, with particular consideration to functional aspects, the results of treatment following the reconstruction of intraoral soft tissue using a pedicled nasolabial flap. Over a period of 10 years (1.1.1990 31.12.1999) the intraoral repair of the defect was carried out in 20 patients in the region of the anterior floor of the mouth, using a unilateral nasolabial flap in 13 cases and a bilateral nasolabial flap in 7 cases. The reasons for the defects were resection of squamous cell carcinomas (T1-T2) of the intraoral mucosa in 19 cases and osteoradionecrosis with soft tissue defect in one case. Local wound healing disturbance was observed in two cases. In 18 patients the prosthetic rehabilitation was successful and allowed a return to masticatory function. In another three cases a secondary operation for flap remodelling was needed. In one case three ITI-Implants were inserted between the foramina 12 months following the placement of the flap. A magnetically retained full denture has been functioning normally for more than four years. The nasolabial flap for the replacement of soft tissue in the mouth appears to represent a functionally and aesthetically satisfactory alternative to reconstruction by microsurgery, in cases of small to medium-sized defects, especially in older, medically compromised patients with an enhanced surgical risk PMID- 12118424 TI - Topical tacrolimus ointment in ulcerative lichen planus: an alternative therapeutic approach. PMID- 12118425 TI - Comparison of UVA protection afforded by sunscreens with a high sun protection factor. PMID- 12118426 TI - Sunscreens with high SPF values are not equivalent in protection from UVA induced polymorphous light eruption. PMID- 12118427 TI - Evaluation of the capacity of sunscreens to photoprotect lupus erythematosus patients by employing the photoprovocation test. PMID- 12118428 TI - High protective effect of a broad-spectrum sunscreen against tetracycline phototoxicity. PMID- 12118429 TI - Prevention of solar-induced immunosuppression by a new highly protective broadspectrum sunscreen. PMID- 12118430 TI - Baseline levels of plasma endothelin-1 (ET-1) and changes during transfusion in thalassemic patients. AB - We studied levels of plasma endothelin-1 (ET-1) in 25 beta-thalassemia/Hb E patients before and after blood transfusion. Baseline ET-1 levels in these patients were significantly higher than in normal controls (10.17 +/- 2.1 pg/mL vs. 8.9 +/- 2.0 pg/mL, P < 0.05). After blood transfusion, levels of plasma ET-1 tend to slightly increase during the first 24 hr but significantly decline at the 7th day to levels (8.01 +/- 1.7 pg/mL) which do not differ from those of controls. This study highlights a different alteration of plasma ET-1 in patients with beta-thalassemia compared to that of patients with sickle-cell anemia. PMID- 12118431 TI - Cytomegalovirus infection in patients undergoing autologous peripheral blood stem cell transplantation. PMID- 12118432 TI - Partial recovery from rabies in a six-year-old girl. PMID- 12118433 TI - Profound neutropenia and atypical lymphocytosis in a traveller to Ghana. PMID- 12118434 TI - Pleural effusion, liver abscesses, and thrombocytosis. PMID- 12118436 TI - Bioinformatic dreams and village realities. PMID- 12118437 TI - The frequency of pre-core gene mutations in chronic hepatitis B infection: a study of Malaysian subjects. AB - A retrospective study was carried out to determine the frequency of the pre-core stop codon mutant virus in a group of chronic hepatitis B carriers: 81 cases were considered [33 hepatits B e antigen (HBe) positive and 48 HBe negative]. All of the HBe positive cases had detectable viral DNA by hybridization analysis; in the case of the HBe negative cases, one third had detectable viral DNA by hybridization analysis and two thirds had HBV DNA detectable by polymerase chain reaction (PCR) amplification. Pre-core stop codon mutant detection was carried out on all specimens using allele-specific oligonucleotide hybridization following PCR amplification of the target sequence. The pre-core mutant was detected in 13/33 (39.4%) of HBe positive cases and in 32/48 (66.7%) of HBe negative cases. Sequence analysis was carried out on 8 of the 16 HBe negative specimens that did not carry the pre-core mutant virus to determine the molecular basis for the HBe minus phenotype in these cases: the 1762/1764 TA paired mutation in the second AT rich region of the core promoter was detected in five cases; a start codon mutation was detected in one case. The predominant mutation resulting in the HBe minus phenotype in our isolates was the 1896A pre-core ("pre core stop codon") mutation; other mutations responsible for the phenotype included the core promoter paired mutation and pre-core start codon mutation. In view of the high frequency of the pre-core mutant virus, sequence analysis was performed to determine the virus genotype on the basis of the nucleotide sequence of codon 15. The sequences of 21 wild type virus (14 HBe positive and 7 HBe negative cases) were examined: 15 were found to be codon 15 CCT variants (71.4%); the frequency in the HBe positive group was 12/14 (85.7%), while that in the HBe negative group was 3/7 (42.9%). The high frequency of the codon 15 CCT variant in association with the frequent occurrence of the pre-core mutant in our isolates concurs with the results of other studies. PMID- 12118438 TI - Chronic carriers of hepatitis B virus in Bangladesh: a comparative analysis of HBV-DNA, HBeAg/anti-HBe, and liver function tests. AB - Serological markers of hepatitis B virus (HBV), liver function tests and quantitative estimation of HBV-DNA are important in the assessment of the state of infection and prognosis following treatment for hepatitis B. This study aimed to determine whether low-cost assays, eg hepatitis B e antigen (HBeAg) and liver function tests, could be used for the assessment of infectivity as an alternative to HBV-DNA estimation. We tested 125 hepatitis B carriers for HBeAg, antibody to HBeAg (anti-HBe), and serum HBV-DNA; we also carried out a range of standard liver function tests. Seventy-three subjects were positive and 52 were negative for HBeAg. Of the HBeAg positive cases, 3 were also positive for anti-HBe; of the HBeAg negative cases, 5 were also negative for anti-HBe. Of these 8 cases, 7 had no detectable HBV-DNA. Most of the HBeAg positive but anti-HBe negative subjects were positive for HBV-DNA (74.3%; 52/ 70) whereas most of the HBeAg negative and anti-HBe positive subjects (93.6%; 44/47) were also negative for HBV-DNA. Of 56 HBV-DNA positive individuals, alanine transaminase (ALT) was found to be raised in 69.6% (p=0.066) and aspartate transaminase (AST) was raised in 66.1% (p=0.011), while 67.9% had normal alkaline phosphatase (ALP) (p=0.054). HBeAg (p=0.018) and raised ALT (p=0.008) were found to be independent predictors for HBV-DNA positivity among HBV carriers. This study suggests that HBeAg positive and anti-HBe negative hepatitis B carriers with raised ALT and AST are likely to be positive for HBV-DNA; the combination of routine serology and biochemical tests may be considered as an alternative to HBV-DNA in evaluating the state of chronic HBV infection. However, HBV-DNA should be specifically assessed if discordance is observed between seromarkers and transaminases. PMID- 12118439 TI - Cost-benefit analysis of hepatitis a vaccination in Thailand. AB - We constructed a decision model to simulate costs and benefits for persons in the context of hepatitis A prevention. Three strategies were compared: i) no intervention; ii) vaccination against hepatitis A without screening; iii) vaccination against hepatitis A for those susceptible after screening for anti HAV. We divided the population into 3 age groups : 3-11 years, 12-18 years and 19 40 years. Data regarding the cost of treatment and vaccination were obtained from the King Chulalongkorn Memorial Hospital. Relevant probabilities were obtained from published literature and expert opinion. At the present incidence of hepatitis A infection, in all age groups examined, the net benefits of a universal no-intervention strategy were higher than those of either vaccination (intervention) strategy. The cost of vaccination without screening in the 3-11 year and 12-18-year groups would equal the benefit if the incidence rates amounted to approximately 138 and 212 infected individuals per 100,000, respectively, that of vaccination with screening at incidence rates of about 200 and 260 infected persons per 100,000, respectively. In the 19-40-year group, the cost incurred by vaccination either with or without screening would equal the benefit at an incidence rate above 450 infected individual per 100,000. For the benefits to outweigh the estimated vaccination costs at present the vaccine is still too expensive. The cost of vaccination without screening in the 3-11-year group would equal the benefit if the cost of vaccine was about 586 baht/2 doses (293 baht/dose), and about 500 baht/2 doses (250 baht/dose) in the 12-18-year group. Likewise, because of the cost of vaccine, it would not be cost-beneficial in the 19-40-year group both with and without screening, and neither would it be in the 3-11-year and 12-18-year groups including screening. According to current standards, under the conditions of the present study the benefit of hepatitis A vaccination administered to the general population between the age of 3 and 40 years in Thailand does not justify the expenses incurred. Major changes in hepatitis A incidence, anti-HAV seroprevalence, vaccine cost or the treatment outcome would be required to potentially render either intervention strategy cost beneficial. PMID- 12118440 TI - Evaluation of sputum staining by modified cold method and comparison with Ziehl Neelsen and fluorochrome methods for the primary diagnosis of tuberculosis. AB - An improved acid-fast staining technique for sputum examination for the primary diagnosis of tuberculosis is described. The technique was modified and simplified by the elimination of heating and by combining the stages of counterstaining: making the technique easier and safer, with less risk of phenol aerosols. The efficiency of this method was evaluated by comparison with two conventional methods, Ziehl-Neelsen (ZN) staining and fluorochrome staining; culture was deemed the gold standard for tuberculosis diagnosis. Of the 392 sputum samples examined, 22.7%, 19.4% and 22.9% were positive by the ZN, fluorochrome and modified cold (MC) staining methods respectively. In comparison with culture results, the sensitivities of ZN, fluorochrome and MC methods were 68.9%, 59.7% and 70.6% respectively; the specificities were 97.4%, 98.2% and 97.8% respectively and the efficiencies were 88.8%, 86.5% and 89.5% respectively. The fluorochrome method was statistically less sensitive than the ZN and MC (p < 0.05), but no significant differences between the ZN and MC were found (p > 0.05). The results of the MC and ZN methods were in close agreement (97.2%); the slides stained by these techniques could be stored for a long time and the staining reagents were stable for several weeks. In conclusion, the MC method proved to be a valuable alternative to ZN staining for the primary diagnosis of tuberculosis. PMID- 12118441 TI - Comparison of microplate hybridization with gel electrophoresis and dot blot hybridization for the rapid detection of Mycobacterium tuberculosis PCR products. AB - A microplate ELISA hybridization assay has been developed for the detection of the IS6110 PCR products of M. tuberculosis from sputum specimens. In this study, its efficacy was evaluated by comparison with agarose gel electrophoresis (AGE) and dot blot hybridization (DBH), with culture results as the 'gold standard'. The assay was used with 190 sputum samples: the PCR results detected by ELISA and AGE showed close agreement, with sensitivity, specificity and accuracy of 90%, 100% and 96% respectively. The same values for DBH were 92%, 98% and 96% respectively. The validities of these methods were not statistically significantly different (p>0.05). The agreement rates of PCR product detection by AGE comparing with DBH and ELISA were 0.964 and 0.964 respectively, while that of DBH and ELISA was 1.0 by Kappa analysis. The overall agreement was not statistically significantly different (p>0.05). Use of DBH or ELISA hybridization increased the sensitivity of detection by AGE 10-fold from 10 pg to 1 pg of purified DNA per reaction; ie from about 30 to about 3 organisms. The amount of PCR product detected by ELISA was only one half of that detected by the other methods; the total assay time of ELISA following the PCR was 4 hours. In conclusion, the microplate hybridization assay may replace AGE and DBH for the detection of the PCR products of M. tuberculosis because of its sensitivity, specificity and accuracy. Additional advantages of the microplate assay over AGE and DBH include rapidity, ease of use, greater safety, cost effectiveness and greater objectivity in the reading of results; the technique is suitable for use in epidemiological studies for the analysis of a large number of samples. PMID- 12118442 TI - Clinical aspects of tonsillar tuberculosis. AB - A clinical analysis of 6 patients with pathologically confirmed tonsillar tuberculosis was carried out retrospectively. The subjects comprised three men and three women, ranging in age from 20 to 74 years. All of the patients presented with a sore throat and 5 had lymphadenopathy. Ulcerations, masses and white patches characterized the tonsillar lesions; the pathological findings included caseous granuloma with positive acid-fast bacilli (AFB) in 5 patients and chronic granulomatous inflammation with negative AFB in one patient. Four of the six patients had pulmonary tuberculosis. The three patients who received complete treatment responded well. The presenting symptoms and abnormal tonsillar findings associated with tonsillar tuberculosis are similar to those of malignant tumors and therefore it is difficult to differentiate the two pathologies; moreover, tonsillar tuberculosis often occurs with pulmonary tuberculosis and AIDS and therefore, a chest X-ray and HIV-screening are recommended for all patients with tonsillar tuberculosis. PMID- 12118443 TI - Modified antimicrobial disc susceptibility testing for nutritionally-variant streptococci. AB - Streptococci that were dependent for their growth upon staphylococci were isolated from a patient with sub-acute bacterial endocarditis and subsequently identified as nutritionally-variant streptococci (NVS). Failure of the isolate to grow on agar media supplemented with pyridoxal hydrochloride or L-cysteine, the known supporting growth factors for NVS, made conventional antimicrobial disc diffusion assay impossible. We modified the assay by co-inoculating Staphylococcus aureus resistant to the drugs being tested as a helper to support the growth of the NVS. Streaking S. aureus closely to the antibiotic discs that were placed above NVS resulted in the growth of satellite colonies of NVS that orbited the S. aureus and that produced a pattern of interrupted zones of growth inhibition. Using an alternative method--adding staphylococcal secreting factor(s) to a 10% staphylococcal cell-free culture supernatant and adding this to an antibiotic susceptibility testing medium,--we found that the NVS formed colonies that formed clear zones of growth inhibition around the disc. When the sizes of the growth inhibition zones produced by both these methods were compared with those recommened by the NCCLS, the NVS were found to be susceptible to penicillin, vancomycin, erythromycin, chloramphenicol, cefoperazone, cefamandole and ofloxacin and resistant to co-trimoxazole, gentamicin and tetracycline. Based on these findings, vancomycin was selected for treatment and the patient was cured of endocarditis. The correlation between the in vitro drug susceptibility testing and the in vivo clinical response indicated that the modified antibiotic susceptibility test is an appropriate method for establishing antibiotic regimens. PMID- 12118444 TI - Leptospirosis in northeastern Thailand: hypotension and complications. AB - During an outbreak of leptospirosis in northeastern Thailand, 148 patients with serologically diagnosed leptospirosis were seen in Loei Hospital. The clinical features were consistent with those described for the classic manifestation of the disease. However, hypotension was a common finding: noted in 94 patients (64%) upon admission or early in the course of the disease. Of these hypotensive patients, 64 (68%) had impaired renal function: 30 patients (32%) had prerenal azotemia and 34 (36%) were in renal failure. Pulmonary complications, including pulmonary edema, hemorrhage, ARDS, and interstitial pneumonitis, occurred in 22% of patients and were often associated with renal failure. A clear association existed between hypotension and renal failure and pulmonary complications. The overall mortality rate was 3.4%. The causes of death were pulmonary complications, renal failure, and sepsis. The death rate among patients with complications was 11.6%. Blood exchange, in addition to conventional treatment, was beneficial in severe leptospirosis with complications and hyperbilirubinemia. PMID- 12118445 TI - Endocarditis and pericarditis caused by Salmonella paratyphi A: two case reports and review of the literature. AB - We report on two children with paratyphoid fever and rare cardiac complications (endocarditis and pericarditis) during an outbreak of Salmonella paratyphi A infection in Bangkok, Thailand, in 1996. Both of the patients had underlying congenital heart disease. Two cases in the literatures of endocarditis and five cases of pericarditis caused by Salmonella paratyphi were reviewed. These rare cardiac complications should be considered among persons who reside in an endemic area of enteric fever or during disease outbreaks, especially in children with underlying heart diseases. PMID- 12118446 TI - Bronchial adenoma presenting with chronic asthma and obstructive pneumonia: a case report. AB - A rare case of primary pulmonary neoplasm is reported. The patient was a 38-year old woman presenting with obstructive pneumonia. Fiberoptic bronchoscopy revealed an endobronchial mass obstructing the left main bronchus: a reddish polypoid mass which bled on contract that was suggestive of bronchial adenoma. The patient also had a long-standing history of bronchial asthma and hemoptysis and the delay in establishing the eventured diagnosis was caused by the minor symptoms mimicking those of asthma. A persistent restrictive lung and the presentation of obstructive pneumonia were important clues which warranted further investigation by computed tomography (CT) scan and bronchoscopy. The patient underwent rigid bronchoscopy with CO2-laser ablation under general anesthesia. Histopathology confirmed a bronchial adenoma. The clinical response was excellent. Bronchial adenoma should be considered in young patients presenting with asthma, hemoptysis and obstructive pneumonia. Bronchoscopic CO2-laser ablation is an effective treatment and provides an alternative to aggressive thoracotomy. PMID- 12118447 TI - Krait bite requiring high dose antivenom: a case report. AB - Anti snake venom (ASV) is the most specific therapy available for treatment of snakebite envenomation. The ASV available in Nepal are polyvalent ASV produced in India and are effective against envenomation by cobra and krait, the two most common species found in Eastern Nepal. Neurotoxic signs respond slowly and unconvincingly and continuous absorption of venom may cause recurrent neurotoxicity. Therefore, close observation and continuous administration of ASV is essential to save the victim. We report a case of neurotoxic envenomation due to bite by common krait (Bangarus caeruleus). The victim required very high dose of polyvalent ASV for reversal of neurological manifestations. PMID- 12118448 TI - Magnesium and zinc status in survivors of sudden unexplained death syndrome in northeast Thailand. AB - Sudden Unexplained Death Syndrome (SUDS) is a major health problem in rural residents of Northeast Thailand. The cause of death in SUDS is suspected to be cardiovascular abnormalities. As magnesium (Mg) and zinc (Zn) deficiency contribute significantly to several cardiovascular diseases, we investigated the Mg- and Zn-status of patients with sudden respiratory distress and cardiac arrest who had survived resuscitation attempts or a near-SUDS episode (N-SUDS). The following subjects were enrolled: 12 N-SUDS inhabitants of rural Northeast Thailand (rural group 1, R1), 13 rural villagers with no past history of N-SUDS (rural group 2, R2), 15 urban Northeasterners (urban group 1, U1); 13 Bangkokians (urban group 2, U2). All subjects were free of structural heart disease. Magnesium and zinc were assessed by atomic absorption spectrophotometry of samples of plasma, red blood cells (RBC), white blood cells (WBC), and 24-hour urine. The mean levels of magnesium in the RBC, WBC, and 24-hour urine of N-SUDS patients (R1) were significantly lower than those of the urban groups (U1 and U2), while the plasma levels did not show any differences. When comparing the Zn status of R1 with that of the urban groups (U1 and U2), the plasma, RBC, and WBC levels were found to be significantly lower in R1 (except for the RBC-Zn of the U1 group), while the 24-hour urine levels was higher. Although the magnesium and zinc parameters were not significantly different between the rural groups R1 and R2, the prevalence of hypomagnesuria (<2.2 mmol/day), hypozincemia (<9.7 micromol/l), and hyperzincuria (>10.7 micromol/day) was higher in the R1 group. These findings suggest that the homeostasis of both magnesium and zinc is altered in N-SUDS patients. Similar alterations, to a lesser degree, were observed in those people living in the same rural environment (R2). PMID- 12118449 TI - Intestinal parasitic infection among five interior communities at upper Rejang River, Sarawak, Malaysia. AB - Intestinal parasitic infection among five interior communities at Bakun Valley, upper Rejang River, Sarawak, Malaysia, was investigated as part of a public health impact assessment of the proposed US$ 3 billion Bakun Hydroelectric Project. Coproparasitological examination of 355 stool samples from 7 of 16 villages representing 5 of 7 tribes in the area revealed infection rate of 41%. A higher infection rate was found among the settled Kayans (56%) than the seminomadic Penans (29%). Infection rate was high (68%) among children less than 14 years old. Trichuris trichiura accounted for more than 90% of all infections; less common were Ascaris lumbricoides, hookworms and Strongyloides stercoralis. Polyparasitism was found in 8% of the individuals surveyed with dual infection due to T. trichiura and A. lumbricoides being more common than dual infection with T. trichiura and hookworm. Women had higher infection rates (57%) than men (33%). PMID- 12118450 TI - Non-occupational determinants of cadmium and lead in blood and urine among a general population in Thailand. AB - In this study the levels of cadmium and lead in blood and urine were measured by the method of graphite furnace atomic absorption spectrometry in 356 healthy, non occupationally exposed individuals and the factors affecting the metal concentrations were investigated. The geometric means for cadmium in blood and urine were 0.98 microg/l (Cd-B) and 0.87 microg/gCr (Cd-U). The lead levels were 32.5 microg/l for blood (Pb-B) and 2.54 microg/gCr for urine (Pb-U). Men had significantly higher blood cadmium and lead levels than women whereas the urinary excretion rates of both metals were higher in women than men. Cigarette smoking was found to affect the levels of Cd-B, Cd-U, and Pb-B. Other factors like alcohol intake and place of residence also related to blood lead levels. Both blood and urine levels of cadmium and lead in this study group were within acceptable ranges for non-occupationally exposed populations and were decreased compared with the past. It is important to continue monitoring levels of these metals in order to prevent adverse health effects in the Thai population. PMID- 12118451 TI - The diagnosis and reporting of occupational diseases: the performance of physicians in Thailand. AB - The diagnosis and reporting of occupational diseases are important components of any occupational disease surveillance system. These two factors were assessed in 222 Thai physicians by using a self-administered questionnaire. Study results show that a proper diagnosis of occupational disease is hampered by the following: lack of knowledge about occupational medicine; a shortage of environmental data; a lack of consultation services and laboratory facilities. Concern about possible legal implications also prevents physicians from making a diagnosis of occupational disease. Evidence shows that financial incentive seems to play a crucial role in physicians' compliance with the reporting system. A number of remedial approaches are proposed, including the improvement of professional training, the development of standard practice guidelines, and novel financial measures for healthcare providers. Improvement calls for the collaborative effort of all responsible agencies and warrants further research that will guide policy and practice. PMID- 12118452 TI - Non-fatal injuries sustained in road traffic accidents: a pilot study in provincial hospitals in Chon Buri, Thailand. AB - The characteristics of patients with non-fatal road traffic injuries who received care from non-referral hospitals are described; an assessment of the difference between the characteristics of patients who received care at a referral hospital and those of patients who were treated at non-referral hospitals is made. A retrospective study, conducted in Chon Buri Province, Thailand, reviewed information from two sources: 324 records from eight non-referral hospitals and the 1999 Injury Surveillance Report of Chon Buri Hospital, a referral hospital. A data collection tool was designed to retrieve information from the non-referral hospitals. Data were analysed descriptively and analytically. The majority of the patients of the non-referral hospitals were male (71.1%) motorcyclists (84.2%), and received ambulatory care (83.9%). Young patients had a higher risk of being admitted to these hospitals. Non-motorcyclists, pedestrians, and nonlocal persons were more likely to receive care from the referral hospital. The results were similar to those of previous studies. The different characteristics of patients who receive care at referral--and non-referral hospitals need to be taken into account when designing traffic accident reduction programs. PMID- 12118453 TI - A sieving method for collecting the metacercariae of trematode parasites from freshwater fish. AB - This study describes a sieving method for the collection of metacercariae from frozen (-20 degrees C) freshwater fish. Digested fish tissue is filtered through a series of sieves; the crude filtrate is then centrifuged. Centrifugation produces a sediment from which metacercariae can be removed. Half of the metracercariae that were obtained from the fish meat that had been frozen for 10 days (-20 degrees C) were dead; the other half were alive and some larvae were moving slowly. PMID- 12118454 TI - Visceral leishmaniasis in two cases of leukemia. AB - Two cases of visceral leishmaniasis (VL), one in a 51-year-old man with accelerated-phase chronic myeloid leukemia and another in a 35-year-old woman with acute myeloblastic leukemia, are reported. Incidental finding of Leishman Donovan (LD) bodies in patients with leukemia highlights VL as a potent opportunistic infection in immunosuppressed patients. PMID- 12118455 TI - Eye lesions and onchocerciasis in a rural farm settlement in Delta state, Nigeria. AB - A study of 326 volunteers from a rural farm settlement in Aniocha North, Delta State, Nigeria, was conducted between December 1999 and September 2000. Volunteers were screened for eye lesions and onchocerciasis. Microfilariae were found in the skin tissues of 134 (41.1%) inhabitants; of these, males had greater microfiladermia (49.9%) than females (33.3%). The percentage prevalence of the clinical signs of onchocerciasis in the farm settlement were: hanging groin 14.1%, onchocercal nodules 25.1%, onchocercal dermatitis 27.6%, scrotal enlargement 3.7%. The percentage prevalence of the eye lesions among those with onchocerciasis were: cataract 24 (7.4%), glaucoma 12 (3.7%), optic atrophy 2 (0.6%) and uveitis 18 (5.5%). Impaired vision was found in 80 (24.5%) inhabitants and blindness was suffered by 2(0.6%) inhabitants. PMID- 12118456 TI - Sandwich ELISA detection of excretory-secretory antigens of Toxocara canis larvae using a specific monoclonal antibody. AB - We produced a new monoclonal antibody (mAb) to the excretory-secretory (ES) antigens of Toxocara canis larvae. The mAb (IgG1) reacts specifically with the 120 kDa protein of many ES molecules and does not have any cross-reactivity with adult T. canis antigens. Sandwich ELISA to detect the ES antigens was performed using the mAb and rabbit polyclonal antiserum. The lower limit for the detection of ES antigen was 4 ng/ml; assay was proportional within a concentration range of 4 ng/ml to 1 microg/ml of ES antigen. This assay system may prove valuable when seeking to quantify parasite burden early in infection and when determining the efficacy of anthelmintic treatment. PMID- 12118457 TI - In vitro effect of antifungal drugs on pathogenic Naegleria spp. AB - An ameba of the genus Naegleria causing fatal meningoencephalitis in human subjects was investigated for its sensitivity to antifungal drugs: amphotericin B, ketoconazole, fluconazole and itraconazole. The efficacy of these antifungal drugs for pathogenic Naegleria spp was investigated in three strains isolated from patients who had died of primary amebic meningoencephalitis infection at Siriraj Hospital (1986), Ramathibodi Hospital (1987) and Chachoengsao Hospital (1987). All of the isolates were maintained in axenic culture in the Department of Parasitology, Faculty of Medicine, Siriraj Hospital, Mahidol University, Thailand. The sensitivities of the antifungal drugs (MIC50) were: amphotericin B (0.05-0.5 microg/ml), ketoconazole (0.125 microg/ml), fluconazole (0.5-2.0 mg/ml), and itraconazole (10 mg/ml) (p < 0.05). It is important to explain that ketoconazole is slightly more effective than amphotericin B because its action is directed of the permeability of the amebic membrane. The amebae were more resistant ot fluconazole and itraconazole due to the action of the cytochrome P450 multienzyme (in the case of fluconazole) and the direct effect on heme-iron, blocking cytochrome P450-dependent chitin synthesis (in the case of itraconzole). We conclude that amphotericin B and ketoconazole remain the main drugs with proven activity against pathogenic Naegleria spp. PMID- 12118458 TI - Malaria species and Southeast Asian ovalocytosis defined by a 27-bp deletion in the erythrocyte band 3 gene. AB - To evaluate the resistance of SAO against species specific malaria infection, relationships between parasite species and the 27-bp deletion in the band 3 gene were studied in malaria endemic Sumba Island, eastern Indonesia. Thick blood films were prepared from patients with malaria symptoms (n=129) and healthy controls (n=231). Species of Plasmodium was identified by microscopic observation. The 27-bp deletion was screened by the PCR method. Among 231 healthy controls, 29 (12.6%) had the 27-bp deletion, whereas 14 (10.9%) among 129 patients confirmed with malaria infection harbored the 27-bp deletion. No significant difference was observed in the prevalence of the 27-bp deletion between controls and patients (p>0.8). There was no significant difference in the frequency of the 27-bp deletion between P. vivax and P. falciparum infected subjects at 5% level by Fisher's exact test. The present result showing no correlation between the presence of the 27-bp deletion and infected parasite species is consistent with the post-invasion resistance hypothesis that may involve not a single malaria species. PMID- 12118460 TI - Dengue hemorrhagic fever in infants. AB - A report of 19 cases of serologically-proven dengue hemorrhagic fever (DHF) in infants aged 3-12 months who were admitted to the Department of Pediatrics, Chon Buri Regional Hospital, Thailand, during 1995 to 1998. Subjects were 8 males and 11 females, with the peak age of 8 months. Four cases (21%) had DHF and other common co-infections ie pneumonia (2 cases), Staphylococcus aureus sepsis (1 case) and Haemophilus influenzae meningitis (1 case). The clinical manifestations of the 15 DHF cases were high fever (100%), coryza (93.3%), hepatomegaly (80%), drowsiness (53.3 %), and vomiting (46.7%); rash was observed in only 27%; one fifth developed febrile convulsions. Sites of bleeding were the skin (petechiae) 58%, gastrointestinal system (melena) 16%, and mucous membrane (epistaxis) 5%; thrombocytopenia and increased hematocrit (> or =20%) were noted in 95% and 84% respectively. The majority of the patients (18 cases, 95%) had primary infection; only one (5%) had secondary infection. The clinical severity of the DHF was Grade I, II, and III (dengue shock syndrome) in 21%, 47% and 32% of cases respectively. After appropriate and effective management, all the infants recovered fully. PMID- 12118459 TI - Predominance of the DEN-3 genotype during the recent dengue outbreak in Bangladesh. AB - A recent outbreak of dengue in Bangladesh was marked by many fatal complications. As clinical virulence varies among the genotypes of dengue virus, a study was conducted to investigate the molecular genotypes of dengue in Bangladesh. Reverse transcription polymerase chain reaction was used to determine viral genotypes using oligonucleotide generic primers that produce a 511 bp product. The resulting product was typed by nested PCR with strain-specific primers, yielding 482 (DEN-1), 119 (DEN-2), 290 (DEN-3) and 392 (DEN-4), visualized on UV transilluminator after electrophoresis on 2% agarose gel stained with ethidium bromide. Of 45 clinically diagnosed dengue patients (mean age 28 years; male/female 30/15), 19 (42.2%) had detectable viral RNA in their blood. However, during the first 5 days of fever in 30 patients, the frequency was 60% (18/30), implying that the sooner serum is drawn after the fever, the greater the chances of detecting viral RNA. DEN-3 was detected in all except 2 patients who were infected with DEN-2. DEN-2 (two cases) and DEN-4 (one case) were present as co infections with DEN-3. All of the patients presented with fever, anorexia and vomiting; many had headache and general body ache; a few had a rash. About a quarter had suffered episodes of bleeding, while ascites, pleural effusion and CNS symptoms were found in a few patients Patients positive for viral RNA were also positive for anti-dengue IgM (p=0.007) in subsequent sampling. The study suggests the predominance of DEN-3 infection with occasional co-infection with other types, during the recent outbreak of dengue in Bangladesh. PMID- 12118461 TI - Dengue hemorrhagic fever in Thai society. AB - Dengue hemorrhagic fever (DHF) is one of the most important infectious diseases in Thailand for many decades. Knowledge of DHF is vital to its control. Like other tropical countries, Thailand is facing this resurgent disease. The Thai National Dengue Prevention and Control Plan has been recently implemented to prevent and reduce the problems resulting from the spread of DHF. In this paper, a three-pronged strategy is offered that will create social mobilization at family, community, and national level and that will, therefore, reduce the socioeconomic and health impacts of DHF. PMID- 12118462 TI - An ecological survey of dengue vector mosquitos in central Lao PDR. AB - An ecological survey of dengue vector mosquitos was carried out in June 2000 in central Lao PDR. Two areas in Khammouane Province, Nongbok and Thakhek, were selected for the survey. Of the 7 mosquito species identified, Aedes aegypti was dominant in both study areas. The container index for Ae. aegypti in Nongbok was 51.8% and was significantly higher than that of Thakhek (40.2%); moreover, significant differences between the study areas were found with records to containers and to the conditions surrounding the houses. The key containers in Nongbok were water jars, whereas drums or small or discarded containers had the highest occurrence rate of Ae. aegypti in Thakhek. Mesocyclops aspericornis was found in large water jars and cement water tanks; no Aedes larvae were found at these sites. Strategy to control dengue vectors in the study areas was discussed. PMID- 12118463 TI - Dengue virus infection of the central nervous system (CNS): a case report from Brazil. AB - Dengue infection that is accompanied by unusual complications has been described in Brazil. We report on the presence of dengue virus in the central nervous system (CNS) of a patient who died in 1998 in Rio Grande do Norte, northeast Brazil. DEN-2 viruses were isolated from the brain liver, and lymphnode tissue of a 67-year-old man whose signs and symptoms were those of dengue infection and a secondary immune response. A postmortem revealed nose bleeds a liver that was brownish with yellow areas, and pulmonary and cerebrae congestion. Immunoperoxidase staining showed a dengue antigen-specific positive reaction in the gray matter cells of the cerebrall cortex; a granular citoplasmatic reaction was seen in the neurons. Dengue infection should always be considered as a cause encephalitis in tropical countries, especially in those where the disease is endemic. PMID- 12118464 TI - Relationship between male hydrocele and infection prevalences in clustered communities with uncertain transmission of Wuchereria bancrofti on the Thailand Myanmar border. AB - A cross-sectional community-based study was conducted in three clustered communities, belonging to a single small village in Mae Chan subdistrict, Umphang district, Tak Province, close to the Thailand-Myanmar border, where regular night blood survey have been discontinued since 1997 and no epidemiological study had been conducted. In order to understand prevalences of distribution of male hydrocele and infection in clinically diagnostic and epidemiologic implications in uncertain transmission of Wuchereria bancrofti, we analyzed the relationship between male hydrocele and community infection prevalences in 219 (90.5% coverage) subjects aged > or =1 year old, including 54.8% migratory and 45.2% local Karen inhabitants. Migratory inhabitants tended to have high prevalence of antigenemia (p < 0.05) and hydrocele. Overall rates of 23.7% antigenemia, 3.7% microfilaremia, and 4.6% male hydrocele were observed. Male hydrocele prevalence was significantly correlated (r = 0.348, p < 0.0001) with antigenemia prevalence, but not with microfilaremia prevalence (r = 0.065, p = 0.493). However, high antigenemia prevalence in local inhabitants was evident, particularly antigenemia prevalence in children suggesting that transmission in the village may have occurred in recent years. PMID- 12118465 TI - A novel molecular method for HIV-1 proviral DNA detection: non-radioactively- reversed probe hybridization and nested PCR. AB - A novel molecular method for HIV-1 proviral DNA detection comprising two main techniques: nested PCR, amplifying a target sequence of the ENV-gene of HIV-1, and nonradioactively-reversed probe hybridization for the detection of the amplified target sequence. The dual amplification of inserted HIV-1 proviral DNA in each DNA sample to be tested was performed by nested PCR in two steps: firstly with two outer primers covering the target sequence of the ENV-gene of HIV-1; secondly with two 5'-biotinylated primers specific to the target sequence. The biotinylated PCR product could be visualized as a single band of 141bps in length on agarose gel stained with ethidium bromide. For the confirmation of the primary result, a method of reversed probe hybridization, using a nylon membrane immobilized with the oligonucleotide probe specific to the target sequence, was established. The oligonucleotide probe was given a homopolymer tail with terminal deoxyribonucleotidyl-transferase; the tail was spotted onto a nylon membrane and bound covalently by UV irradiation. Owing to its length, the tail bound to the nylon, leaving the oligonucleotide probe free to hybridize. Hybridization of the amplified target sequence to the immobilized probe was accomplished by a simple colorimetric reaction involving the enzymatic oxidation of a colorless chromogen that yielded a purple color wherever hybridization occurred. PMID- 12118466 TI - Prevalence of HIV-1 polymerase gene mutations in pre-treated patients in Thailand. AB - To determine the prevalence of drug resistance-conferring mutations in human immunodeficiency virus type 1 (HIV-1), 83 HIV-1 infected Thai patients who had been treated with any antiretroviral drug were studied. HIV-1 RNA was reverse transcribed and amplified by RT-PCR. The direct sequencing of HIV-1 reverse transcriptase (RT) and protease was then performed. Changes in nucleotide and amino acid sequences were determined by comparison with a pNL4-3 reference sequence. Data on mutations associated with resistance to antiretroviral drugs were obtained from literature. The mutations associated with lamivudine resistance (M184V/I) were found most often (in 45.7% of individuals). Zidovudine resistant mutants: T215Y/F (36%), M41L (28%) and K70R (25.3%) were common; but mutations linked to didanosine (L74V) and multinucleoside-resistant genotypes (Q151M) were rarely recognized (2.4% and 3.6%, respectively). The stavudine resistant mutant (V75T) and T69 insertions were not found. All subjects who had a significant exposure to antiretroviral drugs and current virological failure in the past carried drug-resistant genotypes. Genotypic resistance to zidovudine, lamivudine, zalcitabine, indinavir and ritonavir appeared in more than one third of the samples, which suggested that the prevalence of the HIV-1 resistance conferring genotype resisting reverse transcriptase inhibitors and/or protease inhibitors was high in treatment experienced patients. PMID- 12118467 TI - Necropsy in HIV-infected patients. AB - Human immunodeficiency virus (HIV)infection is usually followed by opportunistic infections, especially in the full-blown acquired immunodeficiency syndrome (AIDS). This study details the histopathological changes of different organs in relation to HIV infection, with particular emphasis on the opportunistic infections. Various organs from seventeen HIV-infected patients were collected by necropsy and analyzed for histopathological changes. The major histopathological changes included cytomegalovirus infection, cryptococcosis, penicilliosis, bacterial pneumonia, cryptosporidiosis, pneumocystosis, candidiasis, tuberculosis, granulomatosis of unknown etiology, early cirrhosis and chronic active hepatitis. General organ changes from seventeen cases of HIV-infected patients were described and discussed. PMID- 12118468 TI - A preliminary study of the influence of HIV infection in the transmission of tuberculosis. AB - The human immunodeficiency virus (HIV) epidemic has had a profound influence on the epidemiology of tuberculosis (TB). The potential for HIV-associated TB cases to transmit M. tuberculosis and to produce a secondary increase in TB morbidity is unknown. A cross-sectional study was carried out to compare the prevalence of M. tuberculosis infection among the household contacts of HIV-positive and HIV negative pulmonary tuberculosis (PTB) patients. Records of tuberculin (Mantoux) tests administered during routine contact investigations at the Chest Clinic, Hospital Kota Bharu, from 1999 to 2000 were reviewed. The HIV status of the patients was based on the results of ELISA tests while information on household contacts was gathered during visits to their houses. Ninety-four contacts of 39 HIV-negative patients and 44 contacts of 17 cases of HIV-positive patients were included in this preliminary study. 30% (12/40) of the contacts of HIV-positive PTB had a positive tuberculin compared with 52.8% (47/ 94) of the contacts of HIV negative patients [OR = 0.41, 95% Confidence interval (CI) 0.17 - 0.97; p = 0.016]. The difference was still significant after performing multivariate logistic regression analysis to adjust for variables associated with infectiousness of TB (adjusted OR = 0.24, 95% CI 0.07 - 0.87; p = 0.03). This study has shown that HIV-infected PTB patients are less infectious to their contacts than HIV-negative patients. The presence of MV in the community may not necessitate a change of the current policy of the management of contacts. PMID- 12118469 TI - The effects of antiretroviral dose modification on the re-emergence of HIV-1 wild type strains. AB - Four human immunodeficiency virus type 1 (HIV-1) treatment-naive Thai patients began antiretrovival therapy with a triple drug regimen -zidovudine plus lamivudine plus indinavir; this regimen was modified at week 20 of therapy because of drug toxicity. The virus in all patients was suppressed to lower than 400 copies/ml while they were taking the triple antiretroviral drug regimen. However, suppression was lost after changing the antiretroviral regimen. A comparison of HIV-1 DNA sequences taken from the baseline (day 0) and week 24 showed no significant overgrowth in HIV-1 drug-resistant strains. There was no difference in the protease and reverse transcriptase (RT) mutation profiles. Resistant variants did not emerge, even after sub-therapeutic levels of antiretroviral drugs had been introduced to these patients for 4 weeks. These findings may have clinical implications for long-term treatment strategies. PMID- 12118470 TI - Mitochondrial disorders and exertional intolerance: controversy continues. PMID- 12118471 TI - Mitochondrial disorders and exertional intolerance: controversy continues. PMID- 12118472 TI - When doctors don't know best. PMID- 12118473 TI - Vaccine shortages: how to cope with a new back-to-school hassle. PMID- 12118474 TI - Regulation of dietary supplements: FDA's strategic plan. PMID- 12118475 TI - Lost food and liability: the Good Samaritan food donation law story. PMID- 12118476 TI - Ethics of AIDS clinical trials in developing countries: a review. PMID- 12118477 TI - FDA has substantial and sufficient authority to regulate dietary supplements. PMID- 12118478 TI - The right to a jury trial under the Waxman-Hatch Act--the question revisited and resolved. PMID- 12118479 TI - Maintaining a level playing field: the need for a uniform standard to evaluate health claims for foods and dietary supplements. PMID- 12118480 TI - General principles in the regulatory requirements for health claims. PMID- 12118481 TI - Regulation of natural health products in Canada. PMID- 12118482 TI - Considering changes to CMS's national coverage decision process: applying lessons learned from FDA as a regulator of access to healthcare technology. PMID- 12118483 TI - The insurance aftermath of September 11: myriad claims, multiple lines, arguments over occurrence counting, war risk exclusions, the future of terrorism coverage, and new issues of government role. PMID- 12118484 TI - Constitutional protection of scientific and educational activities from tort liability: the first amendment as a defense to personal injury litigation. PMID- 12118485 TI - Tranexamic acid in patients with hemoptysis. AB - Hemoptysis is a common respiratory symptom leading to admission to hospital. The main management of hemoptysis depends on treating the underlying cause. The use of tranexamic acid is recommended by many doctors without much information available. MATERIAL AND METHOD: This study was a randomized double blinded placebo controlled trial in using tranexamic acid (Transamine) in hemoptysis patients. The study period was one week. Patients with hemoptysis were separated into 3 groups depending on the amount of blood. Group 1 consisted of patients with blood streak sputum. Group 2 coughed up less than 20 ml of frank blood. Patients in Group 3 were those who coughed up 20-500 ml of blood per day. A record of the amount of bleeding and drug side effects was done. RESULTS: From June 1994 to May 1997, 46 patients with hemoptysis completed the study. There were 21 in the tranexamic acid group and 25 in the placebo group. The placebo group had a tendency not to have underlying lung disease and more patients who had a normal chest X-ray. The benefit of tranexamic acid in shortening the days of hemoptysis is not shown in this study. There was a low incidence of side effects of tranexamic acid in this study. CONCLUSION: This randomized double blinded placebo controlled trial could not demonstrate the benefit of tranexamic acid in shortening the days of hemoptysis and confirm the low incidence of side effects of this drug. PMID- 12118486 TI - Efficacy and safety of percutaneous metallic mitral valvuloplasty in rheumatic mitral stenosis at Siriraj Hospital. AB - Mitral stenosis is an important problem that leads to heart failure and stroke in Thailand. The options of treatment at present are either surgical or balloon mitral commissurotomy. However, the cost of balloon is very expensive. To reduce the expense of the procedure, the authors prospectively did a study using a new device called the metallic valvulotome in symptomatic severe mitral stenosis to assess the safety, feasibility and immediate outcomes. Fifty-seven patients were included in the study. The successful outcome achieved by the metallic valvulotome was 96.2 per cent in patients in whom the procedure was actually performed. The mean transmitral gradient, left atrial pressure and pulmonary artery pressure were significantly decreased and the mitral valve area was also significantly increased. Three cases failed the procedure due to inappropriate position of the septal puncture. No death occurred in the study and complications of the procedure included only two cases of hemopericardium. In the future, it is believed that this new innovative device will provide improvement and reduce the cost of the procedure in patients with severe mitral stenosis. PMID- 12118487 TI - Misoprostal use for therapeutic abortion in Siriraj Hospital: the year 2000. AB - OBJECTIVE: To evaluate the efficacy and the adverse effects of misoprostal usage for therapeutic abortion in Siriraj Hospital. STUDY DESIGN: Cross-sectional, descriptive study. MATERIAL AND METHOD: A consecutive series of 101 therapeutic abortions was performed in the Department of Obstetrics & Gynecology, Siriraj Hospital in the year 2000. The patients were interviewed for general information and registered, as well as in-patient data. Any adverse events were recorded and collected from the inpatient record file. All data were analyzed statistically. RESULTS: 42 therapeutic abortions were conducted in association with misoprostal usage. Misoprostal was used for cervical ripening in 8 patients with a good outcome. 31 therapeutic abortions were induced by misoprostal alone regimen. A higher success rate (74.1%) was correlated with higher gestational age. 27 out of 31 cases were second trimester abortion. The induction to abortion interval was 18.0+/-10.5 hours (range 5-48). No factor, including age, weight, total dose of misoprostal use, nulliparity and viability of the fetus, could be demonstrated to affect the misoprostal activity defined by induction to abortion interval. There was no serious adverse event, except for severe abdominal cramping (26.2%) and fever (14.3%). CONCLUSION: Misoprostal alone can be used with caution for abortion induction especially in second trimester abortion. PMID- 12118488 TI - Paclitaxel and carboplatin plus megestrol acetate in the treatment of advanced non-small cell lung cancer. AB - The present study evaluated the efficacy and toxicity of paclitaxel and carboplatin with megestrol acetate for patients with stage IIIb and IV non-small cell lung cancer (NSCLC). Forty patients with no prior chemotherapy and Karnofsky performance status of > or = 60 were enrolled in the study. There were 18 males and 22 females with a median age of 57.5 years, and the median performance status was 70 per cent. Eleven cases were stage IIIb and 29 cases were stage IV. Twenty five cases were adenoCA, 12 were squamous cell, 2 were large cell and one was undifferentiated NSCLC. These patients received paclitaxel 135 mg/m2 by intravenous infusion over 24 hours before carboplatin was given at AUC=6 by 2 hours infusion. Megestrol acetate 160 mg/day was given to all patients from day 2 to 14. This treatment produced partial remission in 12 of 39 evaluable patients (30.76%). Toxicity caused mild nausea, vomiting, myalgia, neuropathy, 20.95 per cent grade 3 neutropenia and 4.15 per cent grade 4 neutropenia. Grade 3 thrombocytopenia was 5.4 per cent, without grade 4. There were no statistically significant changes in weight, serum albumin, and quality of life throughout the cycle 1-6. CONCLUSION: The addition of megestrol acetate to chemotherapy benefitted these patients by minimizing constitute symptoms throughout the treatment period especially in the quality of life, weight loss and stabilized serum albumin. PMID- 12118489 TI - Ureteral calculi during pregnancy: review of the management at Ramathibodi Hospital. AB - OBJECTIVE: To review our experiences with diagnosis and management of symptomatic ureteral calculi complicating pregnancy. MATERIAL AND METHOD: Medical records of all pregnant patients documented with symptomatic ureteral calculi treated at the Division of Urology, Department of Surgery, Faculty of Medicine, Ramathibodi Hospital from 1990 to 2000 were reviewed. Presenting symptoms, diagnostic studies and management of ureteral stone were evaluated. RESULTS: Twenty patients were found in this study with the mean age of 27.5 years (18-36). The mean gestational age at presentation was 18.5 weeks (12-33). Severe flank pain was the common presenting symptom (100%), 60 per cent were on the right side and 40 per cent were on the left side, 20 per cent had associated fever and 20 per cent had irritative voiding symptoms. All of the cases had micro or macroscopic hematuria. Ultrasonography was the initial test confirming the diagnosis and visualized stones were obtained in 60 per cent of the cases. Plain KUB film was done in 6 cases and stones could be seen in 5 cases (83%). Limited IVP was done in 3 cases and the diagnosis could be done in all of them (100%). Spontaneous passing of stones was noted in 14 cases (70%) and double J stents were placed in 6 cases. Ureterolithotomy was done in 2 cases and percutaneous nephrostomy with subsequent definite stone treatment in the post partum period was done in 2 cases. No abortion and no congenital anomalies of the infant were noted. Four cases had premature labor but there was no correlation with the procedures performed for treating the ureteral stone. CONCLUSION: This study provides evidence for effectiveness of diagnosis and treatment of ureteral stone during pregnancy. The appropriate management may be helpful to reduce morbidity of urinary calculi during pregnancy. PMID- 12118490 TI - Dose-expanded study in the reinforcement of efficacy of simvastatin. AB - Two hundred and twenty two hyperlipidemic patients were recruited for a 12-week prospective, multicenter, open-label, titrate-to-goal study to evaluate the efficacy and safety of 20 to 40 mg per day of simvastatin in a Thai population. The efficacy on lipid lowering was evaluated at 4 weeks and 8 weeks after medication. Based on NCEP ATP II guideline and ADA position statement, subjects were categorized into three groups according to LDL-C goals; group I: patients without CHD and with < 2 CHD risk factors, group II: patients without CHD and with > or = 2 CHD risk factors and group III: CHD patients or diabetic patients with > or = 1 risk factors. Significant changes of all lipid parameters from baselines were noted at 4 weeks after medication except for HDL-C levels. Reduction of serum LDL-C, TC and TG by 40 per cent, 29 per cent and 16 per cent respectively and increase of serum HDL-C by 5 per cent were observed at 8 weeks of therapy (p<0.05). At 4 weeks after taking simvastatin 20 mg/day, 78.9 per cent of patients in group I, 67.4 per cent in group II and 40.9 per cent in group III achieved LDL-C goals. Seventeen per cent of the patients who were evaluated at 8 weeks increased the simvastatin dosage to 40 mg per day in the second month of treatment. At 8 weeks of therapy with simvastatin 20-40 mg/day, 90.1 per cent of patients in group I, 77.4 per cent in group II and 66.7 per cent in group III achieved LDL-C goals. Adverse symptoms during therapy, mostly mild, developed in 6.3 per cent of the 222 patients. CONCLUSION: Simvastatin 20-40 mg/day was effective and well tolerated in managing lipid parameters in Thai patients similar to other ethnic populations. PMID- 12118491 TI - Measuring functional status in Thai children with disabilities. AB - OBJECTIVE: The purpose of this study was to compare normal children with age appropriate functional abilities and children with identified disabilities in Thailand. SUBJECTS AND METHOD: Data were collected for 157 nondisabled children and 80 children with cerebral palsy. Their ages ranged from 6 to 100 months. The Functional Independence Measure for children (WeeFIM) is an instrument used to assess independence in self-care, sphincter control, transfers, locomotion, communication, and social cognition. RESULTS: The WeeFIM of the disabilities scored consistently lower in all areas than those of the nondisabled children (p<0.05). Total score, motor score, and cognitive subscores increased with age. When data from Thai children was compared with that from American and Japanese children, total WeeFIM mean scores for each age group and Pearson's correlation coefficients between each age group and total WeeFIM scores showed similar trends. CONCLUSION: WeeFIM can be used as a disability-measuring instrument for Thai children. PMID- 12118492 TI - Comparison among Op-site, polyvinyl chloride film and tulle gauze in the treatment of skin graft donor sitet. AB - A prospective analytic study was performed at the Division of Plastic and Reconstructive Surgery, Department of Surgery, King Chulalongkorn Memorial University Hospital and the Department of Surgery, Chiang Mai University Hospital to compare among Polyvinyl chloride film (PVC film), Op-site and tulle gauze in the treatment of skin graft donor site. From October 1998 to January 2000, 81 donor sites in the same number of patients were treated by three different methods; tulle gauze (26 patients), Op-site (27 patients) and PVC film (28 patients). Each wound was followed until it was completely healed and visual analogue scale was used for pain evaluation. Donor site dressed with PVC film had a healing time of 10.44 days which was not different from Op-site (10.54 days) but significantly faster (p<0.001) than tulle gauze (17.84 days). Pain as measured with visual analogue scale in the group of PVC film (1.48) was not different from Op-site (1.34) but significantly less than (p<0.001) tulle gauze (5.45). There was no difference in the rate of infection between each group. In conclusion, the authors found no difference between Op-site and PVC film in healing time and pain. Both of them were better than tulle gauze. The results demonstrate the usefulness of PVC film as a donor site dressing as it promises relatively rapid healing, less pain and is inexpensive. PMID- 12118493 TI - The role of carbamylated hemoglobin in identifying acute and chronic renal failure. AB - Carbamylated hemoglobin (CarbHb) levels expressed as valipe hydantoin (VH) were measured by high performance liquid chromatography (HPLC) in patients with acute renal failure (ARF, n=35) and chronic renal failure (CRF, n=39). CarbHb levels in CRF patients were approximately 2.5 times of those in ARF ones (121.2 +/- 8 vs 54.8 +/- 6 microgVH/gHb, p<0.01). CarbHb levels of 80 microgVH/gHb provided the best statistical values (sensitivity of 89% and specificity of 82%). CarbHb/BUN and Carb/Cr ratios were also effective determinants in differentiation between ARF and CRF. CarbHb/BUN ratio of 1.5 and CarbHb/Cr ratio of 20 were the best statistical cut off points. As such, measurement of CarbHb levels could be a reliable non-invasive method in identifying ARF from CRF patients. PMID- 12118494 TI - A placebo-controlled, randomized trial of droperidol versus metoclopramide for outpatients undergoing gynecological laparoscopy under conscious sedation. AB - This study compared the prophylactic antiemetic efficacy and the adverse effects of 0.5 mg droperidol, 5.0 mg metoclopramide, and placebo for outpatients undergoing gynecological laparoscopy under conscious sedation. One hundred and fifty outpatients were randomly allocated, in a randomized double-blind manner, into three groups to receive intravenous normal saline, 0.5 mg droperidol, and 5.0 mg metoclopramide before operation. Conscious sedation using intravenous pethidine, midazolam and local infiltration were given to each patient during the operation. Emetic symptoms were graded twice by the patients, at discharge time and the 24th post-operative hour. The difference of antiemetic effect of both study drugs failed to reach statistical significance. There was also no statistical difference of intra-operative hypoxemia, sedation score, and discharge time among the groups. Therefore, using 0.5 mg droperidol or 5.0 mg metoclopramide is not effective in providing antiemetic prophylaxis for outpatients undergoing gynecological laparoscopy under conscious sedation. PMID- 12118495 TI - Peritonitis in acute peritoneal dialysis in a university hospital. AB - OBJECTIVE: Although acute peritoneal dialysis is a useful procedure, peritonitis is often a complication. When the patient is mainly at risk of peritonitis is controversial. The purpose of this study was to find the incidence time of peritonitis, the infecting microorganism, and risk factors. DESIGN: A retrospective study. PATIENTS: 118 cases of acute peritoneal dialysis in 93 patients were included in this study. METHOD: Data were collected from medical records. RESULTS: Overall, the peritonitis rate was 36.45 per cent. The peritonitis rate rose following the duration of dialysis from 11 per cent on the first day to 21 per cent on the third day, although the difference was not statistically significant. Gram-negative bacilli were predominant, at 81.6 per cent. Acinetobacter baumanii and Enterobacter cloacae were the two most common organisms (23.7 and 21.1% respectively). There was a significantly higher male to female ratio in the peritonitis group than the no-peritonitis group (3.33:1 and 1.2:1 respectively, p=0.028). CONCLUSION: There was a high peritonitis rate in acute peritoneal dialysis. The most common microorganisms were gram-negative bacilli, Acinetobacter baumanii and Enterobacter cloacae. The risk factor was male sex. Duration of dialysis of more than 2 days tended to increase the risk of peritonitis. PMID- 12118496 TI - Therapeutic efficacy and safety of loratadine syrup in childhood atopic dermatitis treated with mometasone furoate 0.1 per cent cream. AB - Atopic dermatitis is a common skin disease in Thai children. The treatment of atopic dermatitis requires topical corticosteroids, emollients, systemic antihistamine as well as avoidance of the precipitating factors. A double blind multicenter placebo controlled study was conducted to assess the therapeutic efficacy of topical mometasone furoate 0.1 per cent cream in combination with loratadine syrup. Forty-eight patients, 23 boys and 25 girls, mean age 73.67 months, with atopic dermatitis were included in the study. The severity of the disease was measured by using the SCORAD index including the degree of erythema, dryness, edema/papulation, oozing/crusting, lichenification, and excoriation. Total area involved was measured and a target area of dermatitis was selected for specific evaluation. The degree of clinical signs and pruritic symptom was graded. The sensation of pruritus, disturbance of sleep due to pruritus, and feeling of sleepiness in the morning were recorded. Mometasone furoate 0.1 per cent cream was applied to all patients once daily. One group received loratadine syrup and another group received placebo syrup. They were followed-up on day 5, 8 and 15. The severity of atopic dermatitis and pruritus significantly decreased after 14 days of treatment in both groups (p < 0.001). There was no difference in therapeutic response between the loratadine and placebo groups (p = 0.99). All signs examined had decreased by the end of the study. The result demonstrated that 0.1 per cent mometasone therapy is very effective for treating childhood atopic dermatitis. Loratadine did not show beneficial effect when combined with good topical corticosteroid but it was safe and had no serious side effect on the children. PMID- 12118497 TI - Placental ratio and fetal growth pattern. AB - INTRODUCTION: Placental hypertrophy and reduced fetal growth have been postulated to be an adaptation to maintain placental function in pregnant women with complications such as malnutrition. If this is true, a pregnancy with impaired fetal growth, resulting in a small for gestational age (SGA) infant, should have an increased placental weight to birthweight ratio (placental ratio) compared to those with appropriate for gestational age (AGA) or large for gestational age (LGA) infants. OBJECTIVES: To determine the relationship between placental ratio and fetal growth pattern. MATERIAL AND METHOD: Labour and delivery data of 1000 deliveries in the Department of Obstetrics & Gynecology, Siriraj Hospital from January 2001 to June 2001 were retrospectively studied to compare the placental ratios among pregnancies with SGA, appropriate for gestational age (AGA) and large for gestational age (LGA) infants. RESULTS: From 96 SGA, 804 AGA and 100 LGA cases, a higher placental ratio was found in the SGA group compared to AGA (0.2074 and 0.1985 respectively, p = 0.013). However, actual placental hypertrophy was not found as demonstrated by a lower placental weight in SGA compared with AGA pregnancies of the same birthweight range. There was no significant difference in placental ratio between the LGA and AGA group, the ratios being 0.2020 and 0.1985 respectively (p = 0.260). Although a positive correlation between placental weight and birthweight was observed in the AGA and LGA groups, it was not demonstrated in the SGA infants. This might influence the placental ratio in the SGA group. CONCLUSION: SGA pregnancies are associated with an increased placental ratio which appears not to be due to placental hypertrophy. As reduced birthweight has been shown to be correlated to diseases in adult life, whether this association between SGA and an increased placental ratio will have an implication in future obstetric care and prediction of diseases in adult life remains to be elucidated. PMID- 12118498 TI - Application of 0.05 per cent legal blood alcohol limits to traffic injury control in Bangkok. AB - A substantial proportion (44%) of traffic injury cases seeking emergency services in public hospitals had a blood alcohol concentration (BAC) of 0.1 per cent or more. To reduce alcohol related traffic injuries and deaths, a law was enacted setting a criminal per se legal blood alcohol limit at 0.05 per cent in 1994. However, not until 1997, was an active public education program undertaken on a national scale to raise awareness against drink driving and to support law enforcement. This includes dissemination of knowledge through multiple channels e.g., roadside posters; stickers on the back of vehicles; sporadic radio and TV programs or spots; public announcements; press reports. In 1999, highly visible sobriety check points were set up as a measure for law enforcement. In order to systematically assess the campaign, multiple methods were used to collect relevant data. This report focused on the outcomes of the campaign based on hospital surveillance data in the emergency rooms of 4 public hospitals from March to November, 2000 on alternate months. It was found that the campaign succeeded in raising public awareness and support for law enforcement against drink driving. However, the proportion of road victims with illegal BAC seeking emergency care did not decline after 17 months of the campaign. Limitations and weaknesses of law enforcement activities were discussed along with recommendation for future action. PMID- 12118499 TI - The prevalence of prostate cancer screening in Thai elderly. AB - PROBLEM: Prostate cancer is the most common cancer in elderly men in Western countries. In the future, it may be an important problem in Thailand. At present, there is no evidence about the prevalence and the outcome of screening in this disease. OBJECTIVES: To determine the prevalence of prostate cancer in elderly Thai men and to identify the most appropriate screening method for detection of prostate cancer in Thailand. MATERIAL AND METHOD: 928 elderly men from communities around Siriraj Hospital were evaluated for prostate cancer by Digital Rectal Examination (DRE) and/or Prostate Specific Antigen (PSA). Transrectal ultrasound guided biopsy (TRUS-Bx) which is the gold standard for definitive diagnosis was performed in cases with an abnormal DRE and/or PSA. If biopsy could not be performed, intermittent follow-up with DRE and/or PSA were recommended. RESULT: The prevalence of prostate cancer in Thai elderly men in the urban community was more than 0.75 per cent and the prevalence of abnormal DRE and PSA was 8.7 and 17.3 per cent respectively. The Positive Predictive Value (PPV) of both tests was 60 per cent and higher than the PPV of an individual test. A screening program for prostate cancer starting with DRE may be more cost effective. CONCLUSION: The prevalence of prostate cancer, abnormal DRE and abnormal PSA in Thai elderly men were more than 0.75, 8.7 and 17.3 per cent respectively which are comparable to the prevalence in Western countries. It is important that we take an interest in this disease. PMID- 12118500 TI - Transient hyperkalemia and hypoaldosteronism in a patient with acute glomerulonephritis. AB - The authors describe a 7-year-old boy with acute glomerulonephritis, who developed acute renal failure in the early course of his disease. While the renal function and other clinical manifestations gradually improved, hyperkalemia occurred unexpectedly, and returned to normal level spontaneously after a short period of symptomatic treatment. With the result of a low transtubular potassium gradient (TTKG) level, it was concluded that hypoaldosteronism was the major cause of hyperkalemia in this patient. PMID- 12118501 TI - Otolaryngological complications of osteopetrosis. AB - Osteopetrosis is a rare inherited bone disease that affects both humans and various mammals. The authors report on two cases of osteopetrosis with otolaryngological complications. One patient had the childhood form and presented with chronic otitis media and brain abscess. The second patient had the adult form and presented with sinusitis from tooth extraction which developed into chronic osteomyelitis of the maxillary bone. PMID- 12118502 TI - Constituents of Clerodendrum bungei. AB - Two new compounds, 5-O-ethylcleroindicin D (1) and bungein A (2), together with 12 known compounds (3-14), were isolated from the aerial parts of the medicinal plant Clerodendrum bungei. The structures of 1 and 2 were elucidated as a perhydrobenzofuran derivative and a peroxide dimer by spectral and chemical evidence. Compounds 3-14 have been obtained from this species for the first time. PMID- 12118504 TI - New thiophenes from Echinops grijisii. AB - A new thiophene-echinoynethiophene A (11), four new natural products, namely 5,5" dichloro-alpha-terthiophene (1), 5-chloro-alpha-terthiophene (2), 5-acetyl alpha terthiophene (3) and 5-carboxyl bithiophene (12), together with seven known thiophenes were isolated and purified from ethanol extract of roots of Echinops grijisii Hance. Their structures were identified on the basis of spectral data. PMID- 12118503 TI - Callus cultures of Annona squamosa for the production of annonaceous acetogenins. AB - Callus cultures of Annona squamosa were induced using different explants including petals, seed contents (megagametophyte and embryo) and fruits (mesocarp). Growth of the calli induced from the explants was found to be influenced by the type, concentration and ratio of auxin vs. cytokinin. The content of squamocin (67.8 microg g(-1) dry weight) in calli cultured on Gamborg B-5 medium containing 5.0 mg l(-1) naphthalene acetic acid and 4.0 mg l(-1) zeatin was nearly seven times higher than that in intact fruits. PMID- 12118505 TI - Cytotoxic macrocyclic trichothecenes from the mycelia of Calcarisporium arbuscula Preuss. AB - A new cytotoxic macrocyclic trichothecene calcarisporin B1 (1), and two known compounds roridin H (2) and roridin J (3) were isolated from the cultured mycelia of Calcarisporium arbuscula Preuss. The structure of 1 was determined to be 8alpha-acetoxy roridin H on the basis of spectral data. The cytotoxic activities of 1-3 were evaluated in vitro. PMID- 12118507 TI - The active constituents from Gualou-xiebai-baijiu-tang part I: active saponins. AB - Two new steroidal saponins and a new triterpenoidal saponin, together with nine known steroidal saponins, were isolated from "Gualou-xiebai-baijiu-tang" consisting of Fructus trichosanthis and Bulbus allii macrostemi. The structures of the three new compounds were determined as 3-O-beta-D-galactopyranosyl hederagenin 28-O-beta-D-xylopyranosyl (1-->6)-beta-D-galactopyranosyl ester (1), spirost 25(27)-ene-2beta,3beta-diol-3-O-beta-D-glucopyranosyl (1-->2)-beta-D galactopyranoside (2) and 26-O-beta-D-glucopyranosyl-22alpha-hydroxy-5beta-furost 25(27)-ene-1beta,3beta,6beta,26-tetraol-3-O-beta-D-galactopyranoside (3), respectively, by means of chemical evidences and spectral analysis. PMID- 12118508 TI - A novel steroid from Tylophora atrofolliculata Metc. AB - A novel steroid, tylophoriside A, was isolated from Tylophora atrofolliculata Metc. The structure was elucidated on the basis of spectroscopic methods and X ray diffraction. PMID- 12118506 TI - A cinnamide derivative from Solanum verbascifolium L. AB - A new cinnamide derivative together with N (p-hydroxyphenylethyl) p-coumaramide and vanillic acid has been isolated from the stems of Solanum verbascifolium L., the structure of the new compound was elucidated as N-2-hydroxy-2 (p hydroxyphenylethyl) p-coumaramide (1) on the basis of physical and chemical evidence and spectral analysis. PMID- 12118509 TI - A new monoterpene from the bark of Eucommia ulmoides. AB - A new monoterpene, eucommidiol (1), was isolated from the bark of Eucommia ulmoides Oliv. (Eucommiaceae), together with a known compound 1,4a,5,7a tetrahydro-7-hydroxymethyl-cyclopenta[c]pyran-4-carboxylic methyl ester. The structure of 1 was characterized as 6,6a-di(hydroxymethyl)-3,3a,4,6a-tetrahydro 2H-cyclopenta[b]furan-2-one on the basis of chemical and spectral evidence including 2DNMR studies. PMID- 12118510 TI - Crassicaulisine, a new sulphonoglycolipid from the red alga Chondria crassicaulis Harv. AB - A new sulphonoglycolipid, crassicaulisine, has been isolated from the red alga Chondria crassicaulis Harv.. Four known compounds were also found from the title plant. The structure of the new compound was elucidated on the basis of chemical reactions and spectroscopic analysis. PMID- 12118511 TI - A new ergostanol saponin from Dioscorea deltoidea Wall var. orbiculata. AB - From the fresh rhizomes of Dioscorea deltoidea Wall var. orbiculata, a novel ergostanol saponin, orbiculatoside A (1), was isolated and identified as 3-O-beta D-glucopyranosyl-ergost-5-ene-3beta, 26-diol-26-O-beta-D-glucopyranosyl(1-->3) [beta-D-glucopyranosyl(1-->2)-beta-D-glucupyranosyl(1-->6)]-beta-D glucopyranoside by various NMR techniques in combination with chemical methods. The new saponin showed strong activity against Pyricularia oryzae, with a MMDC (minimum morphological deformation concentration) value of 28.04 micromol/l and was cytotoxic to cancer cell line K562, HCT-15, A549, HT1080, and A2780a in vitro. PMID- 12118512 TI - A new compound from Geum rivale L. AB - A new compound, 1-O-methyl-6-O-caffeoyl-beta-D-glucopyranose (1), has been isolated from the aerial part of G. rivale, together with five known compounds, cecropiacic acid (2), niga-ichgoside (3), gallic acid (4), 1-o protocatechuoylglucose (5), and sucrose (6). Their structures were elucidated by spectral methods and chemical reactions. PMID- 12118513 TI - Asparosides A and B, two new steroidal saponins from Asparagus meioclados. AB - Asparosides A (1) and B (2), two new saponins, were isolated from the roots of Asparagus meioclados. On the basis of chemical and spectroscopic evidence, their structures were elucidated as 23-O-alpha-arabinopyranosyl-(5beta,25s)-spirostan 3beta,23alpha-diol-3-O-[beta-D-xylopyranosyl(1-->4)]-beta-D-glucopyranoside and 26-O-beta-glucopyranosyl-5beta-furost-20(22)-ene-3beta,26-diol-3-O-[beta-D xylopyranosyl(1-->4)]-beta-D-glucopyranoside, respectively. PMID- 12118514 TI - Three new phlegmariurine B type lycopodium alkaloids from Huperzia serrata. AB - Phlegmariurine B (1), a known alkaloid, along with three new analogous compounds, 2alpha-hydroxyphlegmariurine B (2), 2-oxoyphlegmariurine B (3) and 11 oxophlegmariurine B (4), were isolated from the CHCl3 fraction of total alkaloids of whole plant of the Chinese medicinal herb Huperzia serrata. Their structures were elucidated by spectral analysis. PMID- 12118515 TI - Diversity in antimicrobial resistance and other characteristics among Salmonella typhimurium DT104 isolates. AB - Multiresistant Salmonella enterica subspecies enterica serovar Typhimurium definitive type 104 (S. Typhimurium DT104 or DT104) bacteria are important pathogens in animals and humans. DT104 isolates are often called pentaresistant strains that spread clonally. The purpose of this study was to determine phenotypic, genotypic, and epidemiologic characteristics of 175 S. Typhimurium DT104 strains isolated from food-producing animals in Canada. More than 90% of the isolates were resistant to ampicillin (Amp), chloramphenicol (Chl), florfenicol (Flo), sulfisoxazole (Sul), and tetracycline (Tet), 53% of the isolates were additionally resistant to spectinomycin (Spc) and streptomycin (Str), and 28% to kanamycin (Kan) and neomycin (Neo). Sixty-one percent of the strains harbored a single 60-MDa plasmid, 21% contained 60- and 2.0-MDa plasmids, and 4% had 60, 4.6- and 2.0-MDa plasmids. Resistance to Kan and Neo was encoded by the aminoglycoside aphA-1 gene on 2.0-MDa plasmids, whereas resistance to trimethoprim (Tmp) and Sul was encoded by the dhfrIb gene on 4.6-MDa plasmids. Polymerase chain reactions (PCR) showed the presence of integrons with the ant (3")-Ia aminoglycoside adenyltransferase and the bla(PSE-1) beta-lactamase gene cassettes, and the presence of the flost gene in all but one strain resistant to Spc and Str, Amp, and Chl and Flo, respectively. DT104 isolates from cattle at six feedlots represented a separate clone; they were sensitive to Str and Spc and lacked the ant (3")-Ia gene. Pulsed-field gel electrophoresis (PFGE) using Bln I, Spe I, and Xba I resulted in 15, 12, and 8 PFGE patterns, respectively. In summary, we observed considerable diversity in phenotypic, genotypic, and epidemiological characteristics among the DT104 isolates. PMID- 12118516 TI - Antimicrobial resistance pattern of Escherichia coli causing urinary tract infections, and that of human fecal flora, in the southeast of Iran. AB - Sensitivity of 500 Escherichia coli isolates from urinary tract infections (UTIs, 311 isolate) and fecal samples (189 isolates) was tested against 12 antimicrobial agents using the standard disk diffusion method. Although the rate of resistance to antimicrobial agents was higher in the UTIs, in comparison with the fecal samples, the only significant difference was found in cases of tetracycline (p = 0.008), nalidixic acid (p = 0.038), and trimethoprim-sulfamethoxazole (Sxt,p = 0.05). The pattern of sensitivity to antimicrobial agents with respect to statistically significant difference in the number of sensitive isolates (p < or = 0.01) was: ceftizoxime (99.4%) and ceftriaxone (99.2%) > gentamicin (97.8%), ciprofloxacin (93%) and nitrofurantoin (92%) > cefazoline (85.2%) and nalidixic acid (84.6%) > chloramphenicol (71.6%), cephradine (69.6%) and tetracycline (63.2%) > Sxt (41.6%) > ampicillin (23.2%). Sensitivity of the isolates in respect to sex and age was also determined and compared during this study. Resistance to three or more antimicrobial agents (multidrug resistance, MDR) was found in 209 (41.8%) of the isolates. The high rate of resistance to Sxt and the presence of a high rate of MDR isolates in this area suggest that a reevaluation of the first-line therapeutic may be necessary for the treatment of UTIs in this area. PMID- 12118517 TI - Macrolide-resistance genes in clinical isolates of Streptococcus pyogenes. AB - Macrolide-resistance genes were investigated in 103 macrolide-resistant strains of Streptococcus pyogenes, isolated from children with pharyngotonsillitis. The presence of mef(A), erm(B), and erm(TR) genes was detected by PCR. mef(A) was found in 48 out of 103 (46.6%) strains, whereas erm(B) was detected in 43 isolates (41.7%). All mef(A) strains showed a typical M phenotype (resistance to 14- and 15-membered macrolides, and sensitivity to lincosamides and streptogramin B), whereas erm(B) strains had the MLSB phenotype (resistance to macrolides, lincosamides, and streptogramin B antibiotics). erm(TR) was found in 10 strains, always together with other resistance genes. In seven cases erm(TR) was associated with erm(B), and three cases with mef(A). In two isolates with the M phenotype (1.9%), it was not possible to detect the presence of any of the three macrolide resistance genes tested. Inducible resistance to macrolides was shown for 24 out of the 53 MLSB strains. Analysis of macrorestriction fragment patterns by pulsed-field gel electrophoresis showed that erythromycin-resistant S. pyogenes are polyclonal, however each phenotype, MLSB and M, formed essentially homogeneous groups. PMID- 12118518 TI - Vancomycin-resistant enterococci (VRE) in broiler flocks 5 years after the avoparcin ban. AB - The glycopeptide growth promoter avoparcin was banned from animal production in Denmark in 1995. In this study, we investigated the occurrence of vancomycin resistant enterococci (VRE) in broiler flocks in the absence of the selective pressure exerted by the use of avoparcin. One hundred sixty-two broiler flocks from rearing systems with different histories of avoparcin exposure were investigated for the presence of VRE. Using a direct selective plating procedure, VRE were isolated from 104 of 140 (74.3%) broiler flocks reared in broiler houses previously exposed to avoparcin on conventional and extensive indoor broiler farms. In contrast, only 2 of 22 (9.1%) organic broiler flocks reared on free range farms with no history of previous exposure to avoparcin were VRE-positive. Furthermore, the occurrence of VRE over time in flocks reared in broiler houses previously exposed to avoparcin was investigated. Results obtained by direct selective plating showed no significant decrease in the proportion of VRE positive flocks during the study period (1998-2001). This study demonstrated the extensive occurrence of VRE in broiler flocks more than 5 years after the avoparcin ban in Denmark, and indicates that VRE may persist in the absence of the selective pressure exerted by avoparcin. The results differ markedly from previously published Danish surveillance data on VRE in broilers. This may reflect differences in isolation procedures. PMID- 12118519 TI - Characterization of antimicrobial resistance among Escherichia coli O111 isolates of animal and human origin. AB - Fifty isolates of Escherichia coli serogroup O111 recovered from humans and various animal species over a 24-year period (1976-1999) were examined for typical virulence-associated factors and susceptibilities to antimicrobials of human and veterinary significance. Nine H (flagellar) types were identified including nonmotile (n = 24), 32 (n = 12), negative (n = 5), and 56 (n = 3). Thirty-five (70%) isolates possessed at least one Shiga-toxin-producing E. coli (STEC)-associated virulence determinants (eae, stxl, stx2, hlyA) via PCR analysis. Of these 35 isolates, 20 possessed eae, stxl, and hlyA genes, whereas three isolates possessed eae, stxl, stx2, and hylA genes. Multiple antibiotic resistance was observed in 70% of the 50 E. coli O111 isolates. The majority of isolates displayed resistance to streptomycin, sulfamethoxazole, tetracycline, and kanamycin. Bacterial resistance to ampicillin, gentamicin, chloramphenicol, trimethoprim and apramycin was also observed. Integrons were identified in 23 (46%) of the E. coli isolates assayed, with a 1-kb amplicon being most frequently observed. DNA sequencing of these integrons revealed the presence of the aadA gene, encoding resistance to streptomycin. Two integrons of 1.5 and 2 kb contained the aadA2 and either dfrI or dfrXII genes, encoding resistance to streptomycin and trimethoprim, respectively. Integrons were also identified from isolates dating back to 1982. Isolates were further genetically characterized via ribotyping, which identified 15 distinct ribogroups, with 62% of isolates clustering into four major ribogroups. Certain riboprint patterns from different animal species, including humans, were observed in isolates spanning the 24-year collection period, suggesting the dissemination of specialized pathogenic O111 clones. PMID- 12118521 TI - Evolutionary barriers to quinolone resistance in Streptococcus pneumoniae. AB - It is assumed that bacteria always pay a significant physiological price for the acquisition of resistance to antibiotics. To test whether this was the case for a strain of Streptococcus pneumoniae that develops resistance to fluoroquinolone antibiotics, we selected resistance to these agents in a wild-type strain and measured their fitness in comparative growth experiments. The relative growth rate of a mutant strain selected on ciprofloxacin (parC Serine 79 to Tyrosine) was compared with its susceptible isogenic parent and no significant deficit was found (relative fitness 1.15 95% C.I. +/- 0.2.). A double mutant, however, had a relative fitness of 0.81 (parC Serine 79 to Tyrosine gyrA Serine 81 to Tyrosine). Mutant strains selected on gemifloxacin had only a modest increase in minimum inhibitory concentration; thus, second-round mutants were competed with a first round gyrA Serine 81 to Tyrosine or the susceptible isogenic parent. The growth rate of three double-mutant strains parC Serine 79 to Tyrosine gyrA Serine 81 to Phenylanine, parC Serine 79 to Tyrosine, and Asparagine 83 to Phenylalanine were similar to the isogenic susceptible parent 1.16 (95% C.I. +/- 0.17), 0.99 (95% C.I. +/- 0.05), and 0.95 (95% C.I. +/- 0.05), respectively. These data suggest that mutation in the parC and gyrA genes may, on some occasions, not be associated with a physiological deficit. PMID- 12118520 TI - Investigation of the in vitro activity of streptomycin against Mycobacterium tuberculosis. AB - Streptomycin was the first antibiotic used against Mycobacterium tuberculosis. It was used for years in monotherapy regimens, leading to the emergence of resistance; therefore, its use gradually waned. Given the resistance rates detected with current antituberculosis drugs, the use of streptomycin has gained renewed interest. The mechanism of action of streptomycin is inhibition of protein synthesis of mycobacteria in the ribosome. Resistance emerges when mutations appear in genes encoding 16S rRNA and protein S12. The activity of streptomycin against 1,496 M. tuberculosis strains was investigated; 1,186 and 196 strains corresponded to pulmonary and extrapulmonary specimens, respectively. For 114 strains, the source was not indicated. Initially, the BACTEC 460 TB system was used for antibiotic susceptibility testing. Since 1996, the ESP II system was used. The strains ATCC27294 (sensitive to streptomycin, rifampin, ethambutol, and isoniazid) and ATCC35820 (resistant to streptomycin) were used as controls. An overall resistance rate of 2.2% was obtained. In all cases secondary resistance was observed. Multiresistance was observed in 23 strains. PMID- 12118522 TI - Detection and characterization of class 1 integrons in enterohemorrhagic Escherichia coli. AB - Enterohemorrhagic Escherichia coli (EHEC) strains isolated from humans, cattle, and food and belonging to serogroups O26 (7 strains), O111 (19 strains), and O157 (70 strains) were examined for susceptibility to 11 antimicrobial drugs. Fifty nine strains showing resistance to at least one of the drugs were examined by PCR for the presence of class 1 integrons, which were identified in 17 strains. Integrons were found more frequently in strains belonging to serogroups O111 and O26 than in the O157 isolates. DNA sequence analysis demonstrated that most of the integrons contained the aadA1 gene cassette conferring resistance to streptomycin/ spectinomycin, alone or associated with the drfA1 gene cassette conferring resistance to trimethoprim. One integron, identified in a O157:H7 strain, carried the aadA2 and dfrA12 gene cassettes, conferring resistance to streptomycin/spectinomycin and trimethoprim, and the open reading frame F (OrfF) encoding unknown functions. Most of the integrons were carried by Tn21 derivative transposons and were transferable by conjugation to an E. coli K-12 strain. In conclusion, integrons and antibiotic resistance genes can be frequently found in EHEC strains, particularly E. coli O111 and E. coli O26, and their presence could complicate therapeutic trials. PMID- 12118523 TI - Metallo-beta-lactamase VIM-2 in clinical isolates of Pseudomonas aeruginosa from Portugal. AB - Resistance to carbapenems is emerging, and it is a great problem to therapeutics. Three isolates of Pseudomonas aeruginosa from a Portuguese hospital identified in urine and sputum, in 1995, presented a high-level resistance to imipenem (> 32 mg/L). Afterward, one isolate of P. aeruginosa recovered from urine of an ambulatory patient in 1998 showed high resistance to imipenem and meropenem. The resistance to carbapenems in these strains was associated with the production of a class B beta-lactamase, as was demonstrated by imipenem hydrolysis and inhibition by EDTA. Using primers described for bla(IMP) and bla(VIM), the amplification of the latter was observed in all isolates and a VIM-2 metallo enzyme was identified. The pulsed-field gel electrophoresis (PFGE) patterns of these isolates were indistinguishable, suggesting dissemination to the community of this VIM-2 producer. PMID- 12118524 TI - Variations in the occurrence of the S315T mutation within the katG gene in isoniazid-resistant clinical Mycobacterium tuberculosis isolates from Kuwait. AB - The worldwide threat of drug-resistant tuberculosis (TB) to human health has led to the development of molecular methods for rapidly determining the resistance of clinical Mycobacterium tuberculosis isolates to the two front-line antituberculous drugs, isoniazid and rifampin. The prevalence of the S315T mutation within the katG gene, which confers clinically significant resistance to isoniazid, was determined in isoniazid-resistant clinical M. tuberculosis isolates recovered from TB patients in Kuwait. A total of 67 isoniazid-resistant and 18 susceptible clinical M. tuberculosis isolates were tested. The mutation S315T was found in 46 (69%) of the 67 resistant strains, whereas none of the susceptible strains contained this mutation. The prevalence of this mutation was highest (32 of 40, 80%) in isolates recovered from patients of South Asian origin and lowest in isolates from patients of Middle Eastern origin (8 of 18, 44%). The genotyping performed on isolates carrying the S315T mutation showed that the isolates belong to several different types as they exhibited unique DNA banding patterns. The results point to a varying prevalence of the S315T mutation within the katG gene in clinical M. tuberculosis isolates recovered from patients of different ethnic groupings within the same country. The results also suggest that detection of the S315T mutation in the katG gene may be used as a rapid screening method for identifying isoniazid-resistant clinical M. tuberculosis isolates recovered from majority of patients in some ethnic groupings. PMID- 12118525 TI - Vitamin C therapy ameliorates vascular endothelial dysfunction in treated patients with homocystinuria. AB - OBJECTIVES: We sought to investigate the effects of short- and long-term vitamin C therapy on endothelial dysfunction in patients with homocystinuria. BACKGROUND: Untreated homocystinuria due to cystathionine beta-synthase deficiency is associated with premature atherothrombotic disease; 25% of untreated patients suffer a vascular event by the age of 16 years and 50% by 29 years. Treatment directed at reducing homocysteine accumulation significantly reduces this risk. However, despite 'optimal' treatment and compliance, hyperhomocysteinaemia usually persists and individuals exhibit endothelial dysfunction indicative of an adverse cardiovascular prognosis. Additional intervention is therefore required to further reduce cardiovascular risk. METHODS: We investigated the endothelial effects of acute (2 g single dose) and chronic (1 g/day for 6 months) administration of oral vitamin C in 5 patients with homocystinuria (mean age 26 years, 1 male) and 5 age- and sex-matched controls. Brachial artery endothelium dependent flow-mediated dilatation (FMD) and endothelium-independent responses to nitroglycerin (NTG) were measured using high-resolution ultrasonic vessel wall tracking. RESULTS: Baseline: Plasma total homocysteine was 100.8 +/- 61.6 and 9.2 +/- 1.9 micromol/L in the patient and control groups, respectively (p < 0.001). FMD responses were impaired in the patient group (20 +/- 40 microm) compared with the controls (116 +/- 30 microm) (p < 0.001). Vitamin C administration: FMD responses in the patient group improved both acutely, 160 +/- 65 microm at 4 h (p < 0.001), and chronically, 170 +/- 70 microm at 2 weeks (p < 0.001) and 170 +/- 40 microm at 6 months (p < 0.001). FMD responses in the control group were unaltered (p = 0.526). Within both groups, neither the vascular response to NTG nor plasma homocysteine was altered (p > 0.4). CONCLUSIONS: Vitamin C ameliorates endothelial dysfunction in patients with homocystinuria, independent of changes in homocysteine concentration and should therefore be considered as an additional adjunct to therapy to reduce the potential long-term risk of atherothrombotic disease. PMID- 12118526 TI - Congenital microcephaly and seizures due to 3-phosphoglycerate dehydrogenase deficiency: outcome of treatment with amino acids. AB - Congenital microcephaly, intractable seizures and severe psychomotor retardation characterize 3-phosphoglycerate dehydrogenase (3-PGDH) deficiency, a disorder of L-serine biosynthesis. The enzyme defect results in low concentrations of serine and to a variable degree of glycine in plasma and cerebrospinal fluid. Short-term beneficial effects have been reported of oral treatment with the deficient amino acids. In this paper, we report the first follow-up data of amino acid therapy in five patients treated for 3-7.5 years. Different treatment regimes were used, but a favourable response to amino acids was observed in all patients. A major reduction in seizure frequency occurred in all patients; two patients became free of seizures. Amino acids were well tolerated and no adverse effects were documented. A progress of psychomotor development was only observed in one patient, diagnosed early and treated with a high dosage of L-serine. A favourable outcome of 3-PGDH deficiency depends on early diagnosis and treatment. PMID- 12118527 TI - A new case of CDG-x with stereotyped dystonic hand movements and optic atrophy. AB - We report the clinical findings and the diagnostic work-up of a 17-month-old girl with CDG-x. Predominant clinical signs were, besides psychomotor retardation and truncal hypotonia, stereotyped dystonic hand movements and ophthalmological abnormalities such as optic atrophy, nystagmus and strabismus. Other symptoms that are often found in patients with CDG were not present, such as seizures, microcephaly, cerebellar hypoplasia, dysmorphic features, hepatointestinal disease, coagulopathy or multiorgan involvement. Isoelectric focusing (IEF) of the patient's serum showed a marked elevation of disialotransferrin, thus confirming an IEF type 1 pattern. A generalized glycosylation defect was confirmed also by IEF of a further glycoprotein (alpha1-antitrypsin), an increased carbohydrate deficient transferrin (CDT) serum concentration and an increased CDT/transferrin ratio. All known types of CDG-I, secondary glycosylation abnormalities and variants of amino acid sequence were excluded. PMID- 12118528 TI - Failure of resting echocardiography and cardiac catheterization to identify pulmonary hypertension in two patients with type I Gaucher disease. AB - Pulmonary hypertension (PHT) is a complication of Gaucher disease. Screening with echocardiography is recommended for Gaucher patients. Two patients naive to enzyme replacement therapy are presented in whom resting echocardiography revealed no evidence of PHT. One of the patients also had normal pulmonary artery pressures at cardiac catheterization. The diagnosis of PHT was made with open lung biopsy in one patient and dobutamine echocardiography in the other. In both cases, diagnosis of PHT altered patient management. Resting echocardiographic assessment may fail to identify PHT in patients with Gaucher disease. PMID- 12118529 TI - Successful pregnancy in a woman with mut- methylmalonic acidaemia. AB - We report on a favourable pregnancy in a woman affected by mut- methylmalonic acidaemia. Under vitamin B12 and carnitine therapy she remained symptom-free throughout pregnancy, labour, delivery and the postpartum period and gave birth to a term, healthy female newborn. At follow-up, the child shows normal somatic and neurocognitive development. PMID- 12118530 TI - Treatment of extrapyramidal symptoms in a patient with homozygous homocystinuria. PMID- 12118531 TI - 3-Hydroxyglutarate excretion is increased in ketotic patients: implications for glutaryl-CoA dehydrogenase deficiency testing. AB - Three patients with ketosis had increased excretion of 3-hydroxyglutarate (21.8 37.9 micromol/mmol creatinine; controls 2.3 +/- 1.6), an indicator of glutaryl CoA dehydrogenase deficiency (GDHD), which normalized when the patients were nonketotic. Clinical assessment of all three patients and enzyme studies in one patient were not consistent with GDHD. These findings were compared with those of other ketotic patients, who showed statistically significant increases in 3 hydroxyglutarate excretion (9.4 +/- 5.0 micromol/mmol creatinine; p < 0.01), and with those of a child with confirmed GDHD when she was both ketotic and nonketotic. Secondary increase in 3-hydroxyglutarate excretion during ketosis is a potential confounder in the diagnosis of GDHD. PMID- 12118532 TI - Maple syrup urine disease: mutation analysis in Turkish patients. AB - Maple syrup urine disease (MSUD), the most frequently occurring organic acidaemia in Turkey, is caused by a deficiency of the activity of branched-chain keto acid dehydrogenase enzyme (BCKAD) complex. Mutation analysis of the E1alpha, E1beta, and E2 genes of the BCKAD complex in 12 Turkish MSUD patients yielded three disease-specific mutations and a polymorphism in the E1alpha gene, none in the E1beta gene and one mutation in the E2 gene. Among them, three missense mutations (Q80E, C213Y, T106M) and the F280F polymorphism occurring in the E1alpha gene and the splice site mutation (IVS3 - 1G>A) in the E2 gene were novel. Three of the missense mutations and the splicing mutation occurred homozygously and caused classical MSUD. One patient carried the splicing mutation homozygously and the T106M mutation in the heterozygous state; this patient is the first case having simultaneously two different mutations in two different genes in the BCKAD complex. IVS3 - IG>A splicing mutation detected on the E2 gene causes deletion of the first 14 bp of exon 3 in the mutant mRNA extending between 190 and 204 nt. The deletion spans the cleavage point between mitochondrial targeting and lipoyl bearing site of the E2 protein. PMID- 12118534 TI - Outbreak of multidrug-resistant Salmonella newport--United States, January-April 2002. AB - During January-April 2002, Salmonella serotype Newport was isolated from 47 persons in five states: New York (34 cases), Michigan (five), Pennsylvania (four), Ohio (two), and Connecticut (two). Antimicrobial-susceptibility testing of three isolates by CDC revealed resistance to amoxicillin/clavulanate, ampicillin, cefoxitin, ceftiofur, cephalothin, chloramphenicol, streptomycin, sulfamethoxazole, and tetracycline. In addition, two of three isolates were resistant to kanamycin; two had decreased susceptibility or resistance to ceftriaxone. To determine the cause of the outbreak, the New York State Department of Health (NYSDOH) and CDC conducted a case-control study. This report summarizes the results of this investigation, which implicated exposure to raw or undercooked ground beef as the vehicle of transmission. The findings also highlight the emergence of multidrug-resistant S. Newport in the United States. These strains exhibit decreased susceptibility or resistance to ceftriaxone, thereby complicating empiric therapy for serious Salmonella infections. Clinicians should be informed of the emergence of these S. Newport strains, and persons should refrain from eating undercooked ground beef and wash their hands after handling raw ground beef. PMID- 12118533 TI - Gas chromatographic-mass spectrometric newborn screening for propionic acidaemia by targeting methylcitrate in dried filter-paper urine samples. AB - Propionic acidaemia (PCCD) or deficiency of propionyl-CoA carboxylase (PCC) is one of the most common organic acidaemias. Recent studies have suggested that this disease can cause somatic or cognitive deterioration even in patients without ketosis or metabolic acidosis, or in cases with unusually late onset. This suggests that for this disease a sensitive yet practical screening procedure is required to achieve early treatment. We conducted a pilot study of gas chromatographic-mass spectrometric screening of 12,000 newborns for PCCD using eluates from dried filter-paper urine collected at 4-7 days of age. Methylcitrate (MC) was targeted for PCCD. For bulk screening, 2-hydroxyundecanoate was used as internal standard; for quantification, stable-isotope-labelled MC was used. Urease pretreatment without fractionation allowed satisfactory recovery and reproducibility of the highly polar MC. We detected an asymptomatic male infant with distinctly elevated MC: the creatinine-corrected level relative to 2 hydroxyundecanoate was 4.8 SD above the normal mean. The MC concentration calculated using the stable-isotope-labelled internal standard was 70.6 mmol/mol creatinine 14.7 SD above the normal mean of 3.70. Parallel analysis of the dried blood spot at 4 days of age by tandem MS showed only borderline elevation of propionylcarnitine. The activity of PCC in lymphocytes was 7% of control. Gene analysis revealed that a single missense mutation, TAT to TGT, resulting in Y435C in the beta chain was present in a homozygous form. Dietary treatment including carnitine supplementation decreased this infant's MC level and to date (at 13 months of age), he shows no neurological or somatic abnormalities. PMID- 12118535 TI - Outbreak of Campylobacter jejuni infections associated with drinking unpasteurized milk procured through a cow-leasing program--Wisconsin, 2001. AB - On December 7, 2001, the Sawyer County Department of Health and Human Services in northwestern Wisconsin notified the Wisconsin Division of Public Health about five cases of Campylobacter jejuni enteritis. All of the ill persons drank unpasteurized milk obtained at a local dairy farm. This report summarizes the investigation of these and other cases and of a cow-leasing program used to circumvent regulations prohibiting the sale of unpasteurized milk in Wisconsin. The outbreak highlights the hazards of consuming unpasteurized milk and milk products. PMID- 12118536 TI - Hepatitis B vaccination--United States, 1982-2002. AB - This year marks the 20th anniversary of the implementation in the United States of the world's first vaccine against hepatitis B virus (HBV). In addition to acute disease, persons infected with HBV are at risk for chronic HBV infection and severe morbidity and mortality from cirrhosis and hepatocellular carcinoma. Before 1982, an estimated 200,000-300,000 persons in the United States were infected annually with HBV, including approximately 20,000 children. No practical method of pre-exposure prophylaxis for HBV existed, and the only postexposure prophylaxis available was injection with hepatitis B immune globulin (HBIG). PMID- 12118537 TI - Relationship between intrinsic radiation sensitivity and metastatic potential. AB - PURPOSE: Prior studies emphasized genetic modulation of tumorigenicity, and experimental metastatic potential in cells transfected with oncogenes. Whether the intrinsic radiaton sensitivity of cells might correlate with parallel changes in metastatic potential is unknown. METHODS AND MATERIALS: Rat embryo cells (REC) were transfected with the following oncogenes, and where appropriate, with corresponding selection markers: pCMVneopEJ6.6ras, pEJ6.6ras/v-myc, pEla, and pEJ6.6ras/Ela. Individual transfectant clones and corresponding pooled cellular populations were propagated in selective medium. In vitro cellular radiation sensitivity was determined via clonogenic assays, a minimum of three, by standard techniques and individual SF2 and MID parameters determined. Tumorigenicity was defined as the number of tumors forming following the injection of 1 x 10(5) - 1 x 10(6) cells into the axillary pouch of three different strains of immune deficient mice. Animals were killed once resultant tumors reached a maximum size of 1.5-2.0 cm in maximum diameter. For determination of experimental metastatic potential, between 1 x 10(5) - 1 x 10(6) cells were injected into the tail veins of litter-matched sibling mice in parallel to the tumorigenicity studies. RESULTS: Radiobiologic studies indicate similar levels of radiation sensitivity among REC, mock-transfected REC, Ela, and combined E1a/ras transfectants. pEJ6.6ras, and combined ras/myc transfected pooled cellular populations demonstrated increases in radiation resistance when compared to the pooled radiobiologic data from untransfected and mock-transfected corresponding pooled cellular populations (p <0.05, two-tailed test, SF2, MID). Rat embryo cells, Ela, and mock-transfectants were relatively radiation sensitive and nontumorigenic. pEla/ras was tumorigenic but demonstrated relatively low experimental metastatic potential. Ras, and ras/myc transfectants, demonstrated similar levels of experimental metastatic potential on lung colonization assays. CONCLUSIONS: A good correlation exists between the intrinsic radiation sensitivity and the experimental metastatic potential of transfected REC. The highest levels of radiation resistance in vitro and experimental metastatic potential in vivo were found among REC transfected with ras/myc or activated ras alone. E1a/ ras cotransfected cellular populations, although tumorigenic, were relatively radiation sensitive and nonmetastatic. Further study is needed to formulate a mechanistic explanation for the intriguing correlation between intrinsic radiation sensitivity in vitro and metastatic potential in vivo. PMID- 12118538 TI - Effect of high-dose pentoxifylline on acute radiation-induced lung toxicity in a rat lung perfusion model. AB - PURPOSE: The purpose of this study was to study the effect of high-dose oral pentoxifylline on radiation-induced acute lung injury as assessed with a rat lung perfusion model. METHODS AND MATERIALS: Adult male Sprague-Dawley rats were used throughout this study. A preliminary experiment determined that treatment with 2 g/liter pentoxifylline in drinking water resulted in an average consumption of 1.38 g/m2/day, which is comparable to the maximum tolerated dosage in humans. Seventy-two rats were irradiated to the left hemithorax with single fraction doses ranging from 10 through 18 Gy. Half were treated with 2 g/liter pentoxifylline in drinking water from 1 week before radiation through 8 weeks after radiation. Lung vascular perfusion scanning was performed at 3, 4, 5, 6, and 8 weeks after radiation using 99mTc-macroaggregated albumin. The lung perfusion ratio was defined as the number of counts due to radioactivity within the irradiated left lung region of interest divided by the number of counts within the region of the nonirradiated right lung. This lung perfusion ratio has been shown to decrease with radiation-induced lung injury. RESULTS: Although radiation led to a decreased lung perfusion ratio in all groups, those receiving pentoxifylline maintained higher ratios than irradiated controls from 3-5 weeks, especially for those receiving 15 or 18 Gy. However, from 6 through 8 weeks the irradiated controls exhibited partial recovery of lung perfusion ratio, whereas the pentoxifylline groups did not. By 8 weeks after 15 and 18 Gy, lung perfusion ratios were significantly higher for the irradiated controls than for pentoxifylline-treated rats-a reversal of the pattern observed at 3-5 weeks. CONCLUSIONS: The protection by pentoxifylline against radiation-induced acute lung injury was transient and limited to the first 5 weeks after radiation. Subsequent recovery from lung injury was inhibited by this drug at later times within the acute phase. PMID- 12118539 TI - Commissioning and periodic quality assurance of a clinical electronic portal imaging device. AB - PURPOSE: An electronic portal imaging device (EPID) was recently installed on our dual-energy linear accelerator. Commissioning and quality assurance techniques were developed for the EPID. METHODS AND MATERIALS: A commissioning procedure was developed consisting of five parts: (a) physical operation and safety; (b) image acquisition, resolution, and sensitivity calibration; (c) image storage, analysis, and handling; (d) reference image acquisition; and (e) clinical operations. RESULTS: The physical operation and safety tests relate to the motions of the unit, stability of the unit supports, safety interlocks, and interlock overrides. Imager contrast and spatial resolutions are monitored by imaging a contrast-detail phantom. The imager calibration procedure consists of a no-radiation image to compensate for signal offsets, as well as a "flat-field image." The flat-field image is taken with 5.0 cm of homogeneous phantom material placed at isocenter to provide some photon scatter and to approximate the presence of a patient. Daily quality assurance procedures consists of safety tests and the acquisition and inspection of images of the contrast-detail phantom. After 1 year, the frequency of the daily procedure was reduced to weekly. Quarterly QA procedures are conducted by the physicist and consist of the same procedures conducted in the weekly test. The annual QA procedure consists of a duplication of the commissioning procedure. CONCLUSION: The procedures discussed in this article were applied to an ionization-chamber device. They have been useful in identifying difficulties with the EPID operation, including the need for recalibrating and monitoring the accelerator output stability. PMID- 12118540 TI - Effects of intraoperative irradiation and intraoperative hyperthermia on canine sciatic nerve: neurologic and electrophysiologic study. AB - PURPOSE: Late radiation injury to peripheral nerve may be the limiting factor in the clinical application of intraoperative radiation therapy (IORT). The combination of IORT with intraoperative hyperthermia (IOHT) raises specific concerns regarding the effects on certain normal tissues such as peripheral nerve, which might be included in the treatment field. The objective of this study was to compare the effect of IORT alone to the effect of IORT combined with IOHT on peripheral nerve in normal beagle dogs. METHODS AND MATERIALS: Young adult beagle dogs were randomized into five groups of three to five dogs each to receive IORT doses of 16, 20, 24, 28, or 32 Gy to 5 cm of surgically exposed right sciatic nerve using 6 MeV electrons and six groups of four to five dogs each received IORT doses of 0, 12,16, 20, 24, or 28 Gy simultaneously with 44 degrees C of IOHT for 60 min. IOHT was performed using a water circulating hyperthermia device with a multichannel thermometry system on the surgically exposed sciatic nerve. Neurologic and electrophysiologic examinations were done before and monthly after treatment for 24 months. Electrophysiologic studies included electromyographic (EMG) examinations of motor function, as well as motor nerve conduction velocities studies. RESULTS: Two years after treatment, the effective dose for 50% complication (ED50) for limb paresis in dogs exposed to IORT only was 22 Gy. The ED50 for paresis in dogs exposed to IORT combined with IOHT was 15 Gy. The thermal enhancement ratio (TER) was 1.5. Electrophysiologic studies showed more prominent changes such as EMG abnormalities, decrease in conduction velocity and amplitude of the action potential, and complete conduction block in dogs that received the combination of IORT and IOHT. The latency to development of peripheral neuropathies was shorter for dogs exposed to the combined treatment. CONCLUSION: The probability of developing peripheral neuropathies in a large animal model was higher for IORT combined with IOHT, than for IORT alone. The dose required to produce the same level of late radiation injury to the sciatic nerve was reduced by a factor of 1.5 (TER) if IORT was combined with 44 degrees C of IOHT for 60 min. PMID- 12118541 TI - Treatment planning for carcinoma of the cervix: a patterns of care study report. AB - PURPOSE: The Patterns of Care Study (PCS) of patients treated in 1988-89 included "patterns of treatment planning" for radiotherapy of carcinoma of the uterine cervix. A Consensus Committee of radiation physicists and oncologists established current guidelines and developed questionnaires to assess the treatment planning process (i.e., the general structure, methodology, and tools) of institutions involved in the Patterns of Care Study. This paper reports the findings of the assessment. METHODS AND MATERIALS: The PCS surveyed 73 radiotherapy facilities, of which 21 are academic institutions (AC), 26 hospital-based facilities (HB), and 26 free-standing centers (FS). In total, 242 cases were assessed with 39% from academic centers, 33% from hospital-based centers, and 28% from free standing centers. The survey collected treatment planning information such as the use of computed tomography (CT), simulation procedure, contouring of patient outline, tumor or target delineation, identification of critical structures, method of dose prescription (point or isodose), etc. Data was also obtained concerning implant boosts, e.g., radioisotope used, use of midline block for external beam treatment, availability of remote afterloader, practice of interstitial implants, combination with hyperthermia, etc. RESULTS: There is a high degree of compliance relative to the basic treatment planning standards. For example, 171 cases (out of 173) from AC and HB institutions included simulation and 169 used port film; for cases from FS centers, 61 out of 69 involved simulation and 66 out of 69 included port film. Most institutions used linacs (231 out of 242); in five cases, Co-60 units and in six cases betatron was used. In terms of treatment planning, 53% used skin contours, but only 14% had target volume delineation, with AC and HB being slightly more conscientious in these efforts. Critical organs did not appear to be explicitly considered in external beam treatment planning, with only 3% outlining the bladder, 5% the rectum, and less than 1% the small bowel. Only 11% of the centers used CT in treatment planning, and none reported the use of magnetic resonance imaging (MRI). For patients receiving implants, about 40% had midline blocking during external beam treatment, of which one out of three were shielded by standard blocks and two out of three with customized ones. About 11% of the patients receiving implants were treated with remote afterloading devices, 5% received interstitial implants, and none were treated in combination with hyperthermia. CONCLUSION: The treatment planning aspects of radiotherapy of carcinoma of the cervix have been established by this Patterns of Care Study Survey. There is a high level of uniformity in the approach. Some variations exist among centers in the different strata. PMID- 12118542 TI - Preferential radiosensitization of human prostatic carcinoma cells by mild hyperthermia. AB - PURPOSE: Recent cell culture studies by us and others suggest that some human carcinoma cells are more sensitive to heat than are rodent cells following mild hyperthermia. In studying the cellular mechanism of enhanced thermosensitivity of human tumor cells to hyperthermia, prostatic carcinoma cells of human origin were found to be more sensitive to mild hyperthermia than other human cancer cells. The present study was designed to determine the magnitude of radiosensitization of human prostatic carcinoma cells by mild hyperthermia and to examine whether the thermal radiosensitization is related to the intrinsic thermosensitivity of cancer cells. METHODS AND MATERIALS: Two human prostatic carcinoma cell lines (DU 145 and PC-3) and other carcinoma cells of human origin, in particular, colon (HT 29), breast (MCF-7), lung (A-549), and brain (U-251) were exposed to temperatures of 40-41 degrees C. Single acute dose rate radiation and fractionated radiation were combined with mild hyperthermia to determine thermal radiosensitization. The end point of the study was the colony-forming ability of single-plated cells. RESULTS: DU-145 and PC-3 cells were found to be exceedingly thermosensitive to 41 degrees C for 24 h, relative to other cancer cell lines. Ninety percent of the prostatic cancer cells were killed by a 24 h heat exposure. Prostatic carcinoma cells exposed to a short duration of heating at 41 degrees C for 2 h resulted in a substantial enhancement of radiation-induced cytotoxicity. The thermal enhancement ratios (TERs) of single acute dose radiation following heat treatment 41 C for 2 h were 2.0 in DU-145 cells and 1.4 in PC-3 cells. The TERs of fractionated irradiation combined with continuous heating at 40 degrees C were similarly in the range of 2.1 to 1.4 in prostate carcinoma cells. No significant radiosensitization was observed in MCF-7 and HT-29 cells under the same conditions. CONCLUSION: The present data suggest that a significant radiosensitization of prostatic cancer cells could be obtained by the combined treatment of radiation and mild hyperthermia. Future clinical trials should be aimed at achieving mild heating and fractionated radiation therapy. PMID- 12118543 TI - Chemotherapy, early surgical reassessment, and hyperfractionated abdominal radiotherapy in stage III ovarian cancer: results of a gynecologic oncology group study. AB - PURPOSE: To determine outcomes and treatment toxicities in patients with optimal (< or = 1 cm residual) Stage III ovarian carcinoma treated with three courses of cisplatin-cyclophosphamide, surgical reassessment (SRA), and hyperfractionated whole abdominal irradiation (WAI). METHODS AND MATERIALS: Forty-two eligible patients entered this prospective Phase II study conducted by the Gynecologic Oncology Group (GOG). Disease characteristics were as follows: age range, 32-76 years (median 58); Stage IIIA (n = 1, 2%), IIIB (n = 2, 5%), IIIC (n = 39, 93%); histology-serous papillary (n = 21, 50%); other (n = 21, 50%); Grade 1 (n = 1, 2%); 2 (n = 14, 33%); 3 (n = 27, 54%); residual disease after initial surgery (present: n = 23, 55%; absent: n = 19, 45%). Five patients progressed while on chemotherapy, could not be effectively cytoreduced, and were not eligible for WAI. Of the remaining 37 patients, 35 received WAI. Surgical reassessment was not performed in five patients. RESULTS: Of 37 patients with known SRA status after chemotherapy, 21 (57%) were grossly positive, 4 (11%) were microscopically positive, and 12 (32%) were negative. Based on measurements recorded following initial laparotomy and surgical reassessment, progression during chemotherapy was noted in 40%, stage disease in 37%, and objective response in 23%. Toxicity during hyperfractionated WAI was limited and reversible. No patient beginning WAI failed to complete or required a significant treatment break. Following WAI, six patients underwent laparotomies for abdominal symptoms; five had recurrent disease. Five additional patients were managed conservatively for small bowel obstruction (SBO) or malabsorption, of whom three subsequently developed recurrence. Twenty-two patients having pelvic boosts were significantly more likely to require management for gastrointestinal morbidity (p = 0.0021). Considering all eligible patients, median disease-free and overall survivals were 18.5 and 39 months, respectively. Considering patients completing chemotherapy and WAI, median disease-free and overall survivals were 24 and 46 months, respectively. CONCLUSIONS: (a) Disease progression occurred within three cycles of cisplatin and cyclophosphamide chemotherapy in 40% of patients with optimal (< or = 1 cm residual) Stage III ovarian carcinoma. (b) Following limited chemotherapy, hyper-fractionated WAI was acutely well tolerated. (c) Late radiation-related toxicity was observed in only three patients (8.6%) in the absence of recurrent disease. Late gastrointestinal morbidity was significantly associated with the administration of a pelvic radiotherapy (RT) boost. (d) Short duration chemotherapy followed by SRA and hyperfractionated WAI without a pelvic boost is a promising management option for patients with optimal Stage III ovarian cancer. A Phase III trial will be necessary to determine how this treatment strategy compares with chemotherapy or RT alone in this patient population. PMID- 12118544 TI - Indicators of free radical activity in patients developing radiation pneumonitis. AB - PURPOSE: Radiation pneumonitis is thought to occur as the result of excess free radical generation following radiotherapy. Various in vitro studies have shown that large doses of irradiation can cause membrane lipid peroxidation and the oxidation of protein sulphuryl groups. We, therefore, studied two circulating markers of lipid peroxidation and an indicator of "catalytic iron" (potentially available iron to catalyze the generation of free radicals) in patients undergoing radiotherapy. METHODS AND MATERIALS: The 9,11 diene conjugate of 9,12 linoleic acid, expressed as their molar ratio (percentage molar ratio (MR)) and thiobarbituric acid reactive acid-substances (TBARS), as well as levels of circulating desferrioxamine-chelatable iron assay, were assayed. Serial blood samples were taken over a 3-month period in 25 patients with inoperable nonsmall cell lung cancer. RESULTS: Ten patients developed radiation pneumonitis. The patients who developed pneumonitis showed a tendency for the serum percentage molar ratio to increase after a week. The change in the percentage molar ratio between Time 0 and 1 week of radiotherapy was significantly higher in the group that subsequently developed pneumonitis compared to the group that did not (p = 0.002). The initial serum TBARS levels in patients were not significantly elevated compared to controls and there was no difference in the serum TBARS levels in the pneumonitis and nonpneumonitis groups throughout the study period. After 1 week of radiotherapy the group that subsequently developed pneumonitis had a significantly higher level of desferrioxamine-chelatable iron (DFx-iron) compared with the nonpneumonitis group (p = 0.05). CONCLUSION: These data suggest that both the percentage MR and DFx-iron appear to reflect an increased susceptibility to develop radiation pneumonitis and after 1 week of radiotherapy they indicate patients who are likely to subsequently develop pneumonitis. Hence, these indicators could indicate the group of patients that could benefit from intervention therapies with antioxidants. PMID- 12118545 TI - Pelvic radiation therapy combined with hepatic artery chemotherapy for resected rectal carcinoma with liver metastases. AB - PURPOSE: Patients with hepatic metastases from rectal cancer treated with hepatic artery (HA) chemotherapy have a life expectancy great enough to be at risk for pelvic failure. Therefore, a treatment plan was developed for patients with resected rectal cancer and unresectable hepatic metastases, when the pathologic features of transmural invasion and perirectal lymph node metastases were present. Treatment consisted of concurrent pelvic radiation therapy (RT) and HA 5 fluorouracil (FUra), as systemic levels of FUra are achievable with HA administration, followed by HA fluorodeoxyuridine (FdUrd). METHODS AND MATERIALS: Fifteen patients were offered combined pelvic RT and HA FUra. Radiation was given to an initial dose of 45 Gy to the pelvis, followed by boost treatment for an additional 5.4-10.8 Gy. Concurrent HA chemotherapy was given using FUra or FUra/leucovorin administered in two cycles of 14 days for each cycle. If HA chemotherapy could not be done, then intravenous FUra was given during RT. Following completion of RT and HA FUra, patients were evaluated for treatment with HA FdUrd. RESULTS: Eleven patients received concurrent HA FUra or FUra/leucovorin and pelvic RT. Of these, six continued to receive HA FdUrd after completion of RT, as five patients were found to have progressive hepatic disease. Four patients could not have therapy as outlined, but did receive pelvic RT with concurrent intravenous FUra (two patients), FUra/leucovorin (one patient), or sequential HA FUra (one patient). There were four pelvic recurrences at 1, 4, 14, and 17 months after RT. One was the first site of progression, two occurred simultaneously with other failure, and one occurred after hepatic progression. The liver was the most frequent site of first progression (alone in seven patients; as a component of progression in four patients). Treatment was well tolerated with three Grade > or = 3 toxicities. The median survival was 14 months. CONCLUSIONS: These data support the hypothesis that patients with metastatic rectal cancer are also at risk for pelvic recurrence. The frequency of hepatic progression supports continued aggressive therapy directed to this site. As systemic and regional therapy of metastatic rectal cancer improves, we anticipate that more patients will be at risk for a pelvic recurrence, making it increasingly important to explore the role of pelvic radiation therapy despite the presence of metastatic disease. PMID- 12118546 TI - C-Ki-ras mutations in peripheral blood of pancreatic cancer patients: a marker for early tumor metastasis. AB - PURPOSE: To determine the incidence of circulating minimal malignant clone (CMMC) (harboring c-Ki-ras-2 mutation) in peripheral blood (PB) samples of untreated pancreatic cancer using polymerase chain reaction (PCR) analysis of c-Ki-ras-2 oncogene. METHODS AND MATERIALS: Experiments were carried out in fresh tumor, peritoneal washings (PW), and PB samples of untreated pancreatic adenocarcinoma patients (both resectable and unresectable). Samples were taken from 16 patients diagnosed with pancreatic adenocarcinoma for the PCR analysis of mutated c-Ki-ras oncogene. Five tumor samples, 15 PB samples, and 3 PW samples were analyzed for point mutation of the c-Ki-ras gene at codon 12. RESULTS: Out of five tumor samples analyzed for c-Ki-ras mutation, four were positive at the 12th codon. Out of total 15 PB samples, nine were positive for the c-Ki-ras point mutation at the 12th codon. All the positive PB samples showed a base substitution from GGT to GAT in the second position of the 12th codon. Out of three PW samples, two showed mutation at the second position from GGT to GAT similar to their PB and tumor samples. CONCLUSION: Our study indicated the presence of the tumor cells (CMMC) in PB of pancreatic adenocarcinoma that can be identified by PCR analysis of c-Ki ras oncogene. Patients with presence of CMMC in PB and mutation in PW had aggressive tumors that responded poorly to treatment. PMID- 12118547 TI - Bony landmarks are not an adequate substitute for lymphangiography in defining pelvic lymph node location for the treatment of cervical cancer with radiotherapy. AB - PURPOSE: Curative radiotherapy (RT) for carcinoma of the cervix requires adequate irradiation of regional lymph node groups. The best nonsurgical method of defining lymph node anatomy in the pelvis remains the lymphangiogram (LAG). This study was designed to determine if bony landmarks could accurately substitute for LAG as a means of determining lymph node position for the purpose of pelvic RT treatment planning. METHODS AND MATERIALS: The post-LAG simulation films of 22 patients treated at the Fox Chase Cancer Center for cervical cancer were examined. On anterior/posterior (A/P) simulation films, the distance of lymph nodes was determined from the top, middle, and bottom of the sacroiliac joint, and at the pelvic rim, 1 and 2 cm above the acetabulum. On lateral (LAT) simulation films, lymph node position was measured at points 0, 4, and 8 cm along a line from the bottom of L5 to the anterior aspect of the pubic symphysis. Positive values represent lateral and anterior distances relative to the reference point on A/P and LAT films, respectively. Negative values represent distances in the opposite direction. The adequacy of standard pelvic fields as defined by the Gynecologic Oncology Group (GOG) (A/P: 1.5 cm margin on the pelvic rim; LAT field edge is a vertical line anterior to the pubic symphysis) was also examined. Data are expressed as the mean +/- two standard deviations, (i.e. 95% confidence level). RESULTS: On A/P simulation films, the distance of visualized lymph nodes had mean values of -1.6 +/- 1.7 cm (range -4.1 to -0.4 cm), -1.3 +/- 1.5 cm (range -3.4 to 0.0 cm), and 1.2 +/- 1.8 cm (range -1.0 to 2.6 cm) from the sacro-iliac (SI) joint at the superior, middle, and inferior points, respectively. The mean distance of the nodes from the pelvic rim at points 1 and 2 cm above the acetabulum was 0.3 +/- 1.2 cm (range -0.6 to 1.8 cm) and 0.2 +/- 1.8 cm (range -1.6 to 2.1 cm), respectively. On LAT simulation films, the distance of lymph nodes from points 0, 4, and 8 cm from the previously described reference line had mean values of 2.0 +/- 1.0 cm (range 1.3 to 3.0 cm), 0.9 +/- 3.9 cm (range -1.9 to 5.1 cm), and 1.8 +/- 2.1 cm (range -0.8 to 3.5 cm), respectively. Ten of 22 (45%) patients would have had inadequate nodal irradiation if their fields had been designed according to standard GOG parameters. In all cases, these incompletely irradiated lymph nodes were from the lowest of the lateral external iliac group. CONCLUSION: Great variability in pelvic lymph node location is demonstrated when LAG is used to directly visualize their location. Bony structures are inaccurate landmarks for pelvic lymph node position. The GOG standard pelvic fields are not consistently adequate to cover all lateral external iliac lymph nodes, although the clinical significance of this subgroup of lymph nodes is not known. At this time, LAG remains the ideal radiographic modality to define anatomic location of regional lymph nodes for pelvic RT treatment planning. The clinical importance of the most lateral group of external iliac lymph nodes in various stages of cervical cancer represents a potential area of future research. PMID- 12118549 TI - The field-matching problem as it applies to the peacock three dimensional conformal system for intensity modulation. AB - PURPOSE: Intensity modulated beam systems have been developed as a means of creating a high-dose region that closely conforms to the prescribed target volume while also providing specific sparing of organs at risk within complex treatment geometries. The slice-by-slice treatment paradigm used by one such system for delivering intensity modulated fields introduces regions of dose nonuniformity where each pair of treatment slices abut. A study was designed to evaluate whether or not the magnitude of the nonuniformity that results from this segmental delivery paradigm is significant relative to the overall dose nonuniformity present in the intensity modulation technique itself. An assessment was also made as to the increase in nonuniformity that would result if errors were made in indexing during treatment delivery. METHODS AND MATERIALS: Treatment plans were generated to simulate correctly indexed and incorrectly indexed treatments of 4, 10, and 18 cm diameter targets. Indexing errors of from 0.1 to 2.0 mm were studied. Treatment plans were also generated for targets of the same diameter but of lengths that did not require indexing of the treatment couch. RESULTS: The nonuniformity that results from the intensity modulation delivery paradigm is 11-16% for targets where indexing is not required. Correct indexing of the couch adds an additional 1-2% in nonuniformity. However, a couch indexing error of as little as 1 mm can increase the total nonuniformity to as much as 25%. All increases in nonuniformity from indexing are essentially independent of target diameter. CONCLUSIONS: The dose nonuniformity introduced by the segmental strip delivery paradigm is small relative to the nonuniformity present in the intensity modulation paradigm itself. A positioning accuracy of better than 0.5 mm appears to be required when implementing segmental intensity modulated treatment plans. PMID- 12118548 TI - The use of radiochromic film to measure dose distributions resulting from high dose rate 192Iridium single catheter treatments. AB - PURPOSE: Radiochromic film was used to measure and compare the dose distributions parallel to a high dose rate (HDR) 192Iridium (192Ir) brachytherapy afterloading catheter that resulted from optimized treatment plans using various combinations of prescribed dose magnitude and location as well as source spacing. METHODS AND MATERIALS: Differences exist among clinical investigators for specification of the magnitude and location of prescribed treatment dose for brachytherapy irradiations using HDR 192Ir afterloading. Typical prescriptions for endobronchial irradiation include 5 to 10 Gy at 10 mm or 15 Gy at 6 mm measured from the center of the afterloading catheter. The dose distributions that result from these irradiations are very difficult to quantify by conventional dosimetry methods. This study used radiochromic film to measure the dose distributions resulting from optimized treatment plans for source dwell position separations of 2.5 or 5.0 mm and for a prescribed treatment dose of either 15 Gy at 6 mm or 5 Gy at 10 mm, conditions that have been used at M. D. Anderson Cancer Center for the treatment of endobronchial lesions. An acrylic phantom was designed to allow for measurement of the dose distributions at 0.95 mm (catheter surface), 6 mm, and 10 mm from and parallel to the catheter for sources positioned along either 20 or 80 mm of the catheter. RESULTS: Radiochromic film is shown to be a suitable quality assurance and dosimetry modality for the measurement of the dose distribution along an afterloading catheter resulting from an HDR I92Ir source. Each of the treatment plans was about equally effective in being able to produce a uniform dose distribution at their respective planned target distances. Differences were more apparent when comparing the dose distributions at nontargeted distances. On the catheter surface the dose was very nonuniform and in the case of 2.5 mm source spacing along 20 mm of catheter with target dose planned to 10 mm, the central minimum dose was only 13 to 24% of the dose opposite to the most proximal and distal sources. The absolute doses measured at equivalent distances for the 15 Gy planned to 6 mm treatments are about 1.3 to 1.5 times higher than those measured for the 5 Gy planned to 10 mm treatments. It was also observed that the lateral positioning of the encapsulated source within the afterloading catheter can contribute to dose differences about the catheter that are greatest for measurements made in contact with the catheter surface (24 to 40%) but may also be large at the treatment planning distances of 6 (0 to 15%) and 10 mm (0 to 9%). CONCLUSION: At their respective treatment planning distances of 6 or 10 mm, each of the treatment plans produced dose distributions of comparable uniformity. Against the catheter, relatively more uniform dose distributions with higher minimum doses were obtained for (a) dose prescription at 6 mm, rather than at 10 mm; (b) source separation of 2.5 mm, rather than 5.0 mm (except for a 20 mm active catheter length with dose planned to 10 mm); and (c) longer active length of the catheter of 80 mm, rather than 20 mm. PMID- 12118550 TI - The measurement of linear accelerator isocenter motion using a three-micrometer device and an adjustable pointer. AB - PURPOSE: The small motions of the major axes of a linear accelerator observed during gantry and treatment table rotation were measured to improve beam-target alignment during stereotactic radiosurgery (SRS). METHODS AND MATERIALS: Measurements of gantry isocenter motion and table rotational axis wobble were performed with an adjustable front pointer and a three-micrometer device. Nominal gantry and table isocenters were specified. The gantry motion path and table isocenter coordinates were then applied to offset simulated treatment target coordinates so as to compensate for gantry sag. Target simulation films were examined to document improvement of beam-target alignment. RESULTS: The overall precision of the measurement of gantry and table isocenter coordinates was 0.2 mm. Over gantry rotation of 0 to 360 degrees, the gantry isocenter was found to follow a pinched loop with a maximum point to point distance of 1 mm. Table axis motion was found to be negligible relative to the reproducibility of gantry isocenter motion. Thus, a table isocenter was defined that was invariant to table rotation. CONCLUSION: Results indicate that the three-micrometer device and adjustable front pointer are useful tools for three-dimensional (3D) mapping of gantry, collimator and table isocenters and their motions. It is suggested that such measurements may be useful in the quality assurance of linear accelerators, particularly to improve beam-target alignment during SRS and other high dose external beam therapy. PMID- 12118551 TI - A reticle retrofit and dosimetric consideration for a linear accelerator. AB - PURPOSE: An imperfect reticle system in an accelerator causes uncertainties in source-skin distance (SSD), off-axis distance (OAD), isocenter, and so forth. A reticle was designed and fabricated, and its implications on x-ray and electron beam dosimetry were investigated. METHODS AND MATERIALS: A new reticle frame was dimensioned to fit snugly in the accelerator. The frame was fabricated to carry a pair of adjustable cross wires and to allow the machine operation in the photon and electron modes. The impact of the cross wires on 6 MV photon and 5-10 MeV electron beam parameters such as dose rate (Gy/monitor unit), beam uniformity, surface dose, and so forth, were studied using suitable ion chambers and phantoms. RESULTS: The retrofitted system offered long-term mechanical stability leading to precise SSD, OAD, and isocenter measurements. Changes introduced by the cross wires on the 6 MV photon and 5-10 MeV electron beams are presented. CONCLUSION: Long-term stability of a reticle in an accelerator is important for an accurate patient setup and for making reliable dosimetric measurements. Beam characteristrics have to be studied whenever modifications on a reticle system are made. PMID- 12118552 TI - Volumetric visualization of anatomy for treatment planning. AB - PURPOSE: Delineation of volumes of interest for three-dimensional (3D) treatment planning is usually performed by contouring on two-dimensional sections. We explore the usage of segmentation-free volumetric rendering of the three dimensional image data set for tumor and normal tissue visualization. METHODS AND MATERIALS: Standard treatment planning computed tomography (CT) studies, with typically 5 to 10 mm slice thickness, and spiral CT studies with 3 mm slice thickness were used. The data were visualized using locally developed volume rendering software. Similar to the method of Drebin et al., CT voxels are automatically assigned an opacity and other visual properties (e.g., color) based on a probabilistic classification into tissue types. Using volumetric compositing, a projection into the opacity-weighted volume is produced. Depth cueing, perspective, and gradient-based shading are incorporated to achieve realistic images. Unlike surface-rendered displays, no hand segmentation is required to produce detailed renditions of skin, muscle, or bony anatomy. By suitable manipulation of the opacity map, tissue classes can be made transparent, revealing muscle, vessels, or bone, for example. Manually supervised tissue masking allows irrelevant tissues overlying tumors or other structures of interest to be removed. RESULTS: Very high-quality renditions are produced in from 5 s to 1 min on midrange computer workstations. In the pelvis, an anteroposterior (AP) volume rendered view from a typical planning CT scan clearly shows the skin and bony anatomy. A muscle opacity map permits clear visualization of the superficial thigh muscles, femoral veins, and arteries. Lymph nodes are seen in the femoral triangle. When overlying muscle and bone are cut away, the prostate, seminal vessels, bladder, and rectum are seen in 3D perspective. Similar results are obtained for thorax and for head and neck scans. CONCLUSION: Volumetric visualization of anatomy is useful in treatment planning, because 3D views can be generated without the need for segmentation. When relationships among anatomical structures, rather than geometric models of them, are important, volume rendering presents advantages. The presented algorithm is readily adaptable to distributed parallel implementation on a network of heterogeneous workstations. PMID- 12118553 TI - Intraoperative radiation therapy in recurrent carcinoma of the uterine cervix: report of the French intraoperative group on 70 patients. AB - PURPOSE: To evaluate the feasibility and oncologic results of intraoperative radiation therapy (IORT) for recurrent uterine cervical carcinoma in a cohort of patients treated in seven French institutions. METHODS AND MATERIALS: From 1985 to 1993, 70 patients with pelvic recurrences underwent IORT with/ without external radiation therapy (ERT) and chemotherapy (CT). Treatment modalities for recurrence were IORT alone (40 out of 70), IORT + ERT (30 out of 70), additional chemotherapy (20 out of 70). Gross complete resection (CR) was performed in 30 out of 70 cases, partial resection (PR) in 37 out of 70, and unspecified surgery in 3 out of 70. Sixty-five patients had electron beam IORT and 5, 100 KV photon IORT. Mean IORT cone size, electron beam energy, and dose (calculated at the 90% isodose line) were, respectively, 75 mm (40 to 90), 12 MeV (6 to 20), and 18 Gy (10 to 25) after CR and 80 mm (45 to 100), 15 MeV (7 to 24), and 19 Gy (10 to 30) after PR. RESULTS: Mean follow-up after IORT was 15 months (2 to 69). One, 2- and 3-year overall survival rates were 47, 17, and 8%, respectively; median survival was 11 months and local control, 21%. Median survival and local control rates increased after CR (13 months, 27%) vs. PR (10 months, 17%) and when initial treatment consisted of surgery (S) alone (15 months, 25%) vs. radiation therapy (RT +/- S) (10 months, 16%). However, these differences were not statistically significant. No death-related toxicity was observed. Grade 2 or 3 toxicity was observed in 19 out of 70 patients (27%), including 9 not directly IORT-related complications (13%) (three digestive tract fistulas, one rectal stricture, three urinary fistulas, two infections) and 10 directly IORT-related complications (14%) (five neuropathies, four ureteral obstructions, and one rectal stricture). CONCLUSION: This retrospective study demonstrates the feasibility of IORT. The usefulness of IORT still needs to be evaluated in primary treatment of advanced stages of cervical carcinoma. PMID- 12118554 TI - A comparison of four patient immobilization devices in the treatment of prostate cancer patients with three dimensional conformal radiotherapy. AB - PURPOSE: To determine the variability of patient positioning during three dimensional conformal radiotherapy (3D-CRT) for prostate cancer treated with no immobilization or one of four immunobilization devices, and to determine the effects of patient body habitus and pelvic circumference on patient movement with each individual inmobilization technique. METHODS AND MATERIALS: To see whether our immobilization techniques have improved day-to-day patient movement, a retrospective analysis was carried out. A total of 62 patients treated at one facility on a single machine with 3D-CRT via a four-field box technique (anterior posterior and opposed laterals) in the supine position with either no immobilization or one of four immobilization devices. Five groups of patients were compared: (a) group 1-no immobilization; (b) group 2-alpha cradle from the waist to upper thigh; (c) group 3-alpha cradle from waist to below the knees; (d) group 4-styrofoam leg immobilizer (below knees); and (e) group 5-aquaplast cast encompassing the entire abdomen and pelvis to midthigh with alpha cradle immobilization to their lower legs and feet. Prior to starting radiotherapy, portal films of all four treatment fields were obtained 1 day before treatment. Subsequently, portal films were then obtained at least once a week. Portal films were compared with the simulation films and appropriate changes were made and verified on the next day prior to treatment. A deviation of greater than 0.5 cm or greater was considered to be clincally significant in our analysis. We studied the difference among the types of immobilization and no immobilization by looking at the frequency of movements (overall, and on each of the three axes) that a patient had during the course of his treatment. Using a logistic regression model, the probability of overall and individual directional movement for each group was obtained. In addition, the effects of patient body habitus and pelvic circumference on movement were analyzed. RESULTS: The maximum deviation was 2 cm and the median deviation was 1.2 cm. For each patient, the probability of movement ranged from 0 to 76%, with a mean of 39%. There was no significant difference seen in overall movement with any of the immobilzation devices compared to no immobilization, but there was less vertical (9 vs. 18%; p = 0.03) and AP (6 vs. 15%; p = 0.14) movement with the aquaplast than any other group. However, when examining the lateral direction, the aquaplast had significantly more movement (32 vs. 9%; p < 0.001). When accounting for body habitus and pelvic circumference, no immobilization device was effective in reducing movement in obese patients or in patients with pelvic circumference greater than 105 cm. The aquaplast group had a significantly increased amount of lateral movement with obesity (42 vs. 23%; p < 0.05), and with pelvic circumference >105 cm (33 vs. 29%; p < 0.05). CONCLUSIONS: There was no significant reduction in overall patient movement noted with any of the immobilization devices compared to no immobilization. The aquaplast group had reduced vertical and AP movement of greater than 0.5 cm. There was significantly more lateral movement with aquaplast appreciated in obese patients or patients with pelvic circumferences greater than 105 cm. The aquaplast immobilization appears to be useful in reducing movement in two very clinicaly important dimensions (AP and vertical). Despite our findings, other immobilization may still be useful especially in the treatment of nonobese patients. It is clear that the optimal immobilization technique and patient positioning are yet to be determined. PMID- 12118555 TI - Results of the 1993 association of residents in radiation oncology survey. AB - In 1993, the Association of Residents in Radiation Oncology (ARRO) conducted its tenth annual survey of all residents training in radiation oncology in the United States. The characteristics of current residents are described. Factors influencing the choice of Radiation Oncology as a medical specialty, and posttraining career plans were identified. Residents raised issues on the adequacy of training, problems in work routine, and expressed concerns about board certification and recertification, and about decreased future practice opportunities. PMID- 12118556 TI - Lessons learned from investigations of therapy misadministration events. AB - PURPOSE: Investigation teams composed of Idaho National Engineering Laboratory (INEL), United States Nuclear Regulatory Commission (NRC), and subcontractor personnel performed detailed investigations and analyses of seven misadministration events that were specifically selected on the basis of particular characteristics. These events were analyzed to identify the direct causes, contributing factors, actions to mitigate the event, and the consequences of these events. The INEL also sought to determine the role played by the recent Quality Management Rule. METHODS AND MATERIALS: The investigation teams were multidisciplinary and, depending on the nature of the event, included three or more team members with appropriate expertise in the areas of radiation oncology, medical physics, nuclear medicine technology, risk analysis, and human factors. The investigations focused on the general areas of causes of the event, mitigating actions, and corrective actions. Seven misadministration events were investigated by the teams during 1991 and 1992. RESULTS: Results from the events investigated indicated that (a) the institutional traditions of some licensees contributed to the potential for misadministrations, (b) many misadministrations occurred primarily due to lack of procedures or procedures that were not clearly written, (c) some licensees in this study had not effectively implemented their Quality Management programs, and (d) limited involvement on the part of the Radiation Safety Officer and Authorized Users and changes in routine and unique conditions contribute to the potential for misadministrations. CONCLUSIONS: The project shows that licensees that have experienced misadministration events appear to lack comprehensive safety cultures, where all aspects of daily operations are shaped with patient and staff safety being the primary objective of all activities. PMID- 12118557 TI - A comparison of the structure of radiation oncology in the United States and Japan. AB - PURPOSE: The United States and Japan have very different backgrounds in their medical care systems. In the field of radiation oncology, national surveys on structure have been conducted for both countries and compared to illustrate any similarities and differences present from 1989-1990. METHODS AND MATERIALS: The Patterns of Care Study Facility Survey conducted in 1989 in the United States and the National Survey of Structure in Japan in 1990 were compared to evaluate the equipment pattern, staffing pattern, compliance rate with the "blue book" (3) guideline, and the geographic distribution of institutions. RESULTS: In the United States, a total of 598,184 (49% of the total of newly diagnosed) patients were treated with radiation therapy. In Japan, 62,829 (approximately 15% of the total of newly diagnosed) patients were treated. The numbers of external megavoltage treatment machines were 2,397 in the United States and 494 in Japan. The numbers of full time equivalent (FTE) radiation oncologists were 2,335 in the United States and 366 in Japan. Only 15% of United States facilities and 11% of Japan facilities complied with the narrow blue book guideline for the patients per FTE radiation oncologist (200-250), while the most common ratio was 151-200 patients/FTE in the United States and 51-100 in Japan. In Japan, more than 60% of institutions were staffed by a part-time radiation oncologist (FTE < 1.0). Between geographic regions, there was variation in the percentage of cancer patients treated with radiation therapy for both the United States (42-56%) and Japan (6-25%). CONCLUSION: There is a major difference in the usage of radiation therapy for treating cancer between the United States and Japan with 49% of all new cancer patients treated in the United States and approximately 15% treated in Japan. Equipment structure in the United States is more complete than in Japan with important differences in treatment simulators, treatment planning computers, and support personnel. High dose rate intracavitary radiation is commonly available in Japan and there are geographic differences in radiation oncology utilization in both countries. PMID- 12118558 TI - The effect of delay in treatment on local control by radiotherapy. AB - PURPOSE: The objective of this study was to estimate the effect of delay in initiation of treatment on rates of local control by radiotherapy. METHODS AND MATERIALS: A model of the effects of delay was developed based on the following assumptions: (a) that tumor growth rate is exponential, (b) that a predetermined radiotherapy regimen will kill the same fraction of clonogenic cells in a given tumor whether it is administered early or late, and (c) that the absolute number of cells surviving in a tumor is determined by Poisson statistics. Monte Carlo simulation was used to estimate the expected rate of decrease in local control associated with delay in a population of tumors, which was heterogeneous with respect to doubling time and initial volume. The model was applied to carcinoma of the tonsillar region. RESULTS: It was shown that at some point in the evolution of every case, the probability of local control decreases sharply over a relatively short period of time. The maximum rate of decrease in the probability of local control occurs at the 37% local control level when it reaches 25.5% per tumor doubling time. When heterogeneity with respect to doubling time and stage was taken into account, it was estimated that the local control rate would decrease by approximately 10% per month in a typical series of patients with carcinoma of the tonsillar region. CONCLUSIONS: It was concluded that delay in initiation of radiotherapy may be associated with a clinically important deterioration in local control rates. We recommend that waiting times for radiotherapy should be As Short As Reasonably Achievable (ASARA). PMID- 12118559 TI - A review of human cell radiosensitivity in vitro. AB - The survival curves of 694 human cell lines irradiated in exponentially growing phase in vitro were collected from the literature. Among them, 271 were derived from tumors, 423 were nontransformed fibroblasts and other normal cell strains from healthy people or people with some genetic disorders. Seventy-six different cell types are identified, and a specific radiosensitivity could be associated with each, using D and surviving fraction at 2 Gy. Technical factors such as culture medium, feeder cells, and scoring method were found to affect intrinsic radiosensitivity. In particular, the cell type is not a discriminating factor when cells are studied in agar. Results obtained with cells irradiated in agar must be used cautiously, depending on how the cells were prepared for the experiments. The use of feeder cells narrows the range of radiosensitivity of human cells. For cells irradiated as monolayer, it was possible to build a scale of radiosensitivity according to cell type, ranging, in terms of D from 0.6 Gy for the most sensitive cell lines to more than 4 Gy for the most resistant. Considering that, in most cases, we could estimate the variation of radiosensitivity within each cell type, our classification among cell types can be used by researchers to place their results in the context of the literature. PMID- 12118560 TI - Midwest institute for neutron therapy at Fermilab. PMID- 12118561 TI - Medically inoperable stage I endometrial carcinoma: a few dilemmas in radiotherapeutic management. AB - PURPOSE: The aggressiveness of radiation therapy for patients with medically inoperable endometrial carcinoma is controversial. Patients may die of their underlining medical disease before succumbing to cancer. We try to identify certain subgroup of patients who might benefit most from an aggressive approach and also investigate the impact of residual tumor present in dilatation and curettage (D&C) specimen obtained in second intracavitary implant (ICI). METHODS AND MATERIALS: From 1965 to 1990, 101 patients were treated for clinical clinical Stage I endometrial carcinoma with RT alone due to medical problems. Ages ranged from 39 to 94 years (median 71 years). There were 18 patients with clinical Stage IA and 83 with clinical Stage IB disease. Histology included 44 well differentiated, 37 moderately differentiated, and 20 poorly differentiated tumors. Radiation therapy consisted of external beam only in 3 patients, ICI alone in 26, whole pelvis plus ICI in 10, and whole pelvis plus split field plus ICI in 62. A second D&C was performed on 26 patients at the time of the second ICI. Minimum follow-up was 2 years (median, 6.3 years). RESULTS: The 5-year actuarial disease-free survival (DFS) for the studied cohort is comparable to the expected survival of an age-matched population. Pelvic control was 100% for Stage IA and 88% for Stage IB with 5-year disease-free survivals of 80 and 84%, respectively. We also observed a greater disassociation of DFS and overall survial among patients older than 75 years (84 and 55%, respectively) than in younger patients (84 and 78%, respectively). This is mainly because older patients succumbed to their medical illness. Well-differentiated disease demonstrated the trend toward a better outcome than moderately or poorly differentiated lesions in Stage IB patients (p = 0.05), but not in Stage IA patients. Aggressive radiation therapy approach showed the trend toward a better result in Stage IB patients 75 years of age or younger. There were two failures among 19 patients with no tumor found in the D&C specimen at the time of second implant. In contrast, seven patients with residual tumor seen in the endometrial sample at the time of second implant remain disease free. CONCLUSIONS: Radiation therapy alone is an effective treatment modality for medically inoperable Stage I endometrial carcinoma. Disease-free survival can be translated into longer overall survival in the younger age group, but not in older patients. The latter tend to die of their underlining medical illness. Tumor differentiation influenced the prognosis of Stage IB disease. No tumor seen in the endometrial sampling at the time of second implant did not correlate with a better disease control, and the treatment plan should not be modified on such information. PMID- 12118562 TI - Carcinoma of the uterine cervix. II. Lack of impact of prolongation of overall treatment time on morbidity of radiation therapy. AB - PURPOSE: Several reports document a negative impact of prolongation of overall treatment time in a course of irradiation on tumor control and survival. A correlation has been documented of incidence of significant treatment sequelae with increasing doses of irradiation, volume of the specific organ, and dose per fraction. However, no data were found on the potential correlation of overall irradiation treatment time with significant sequelae. METHODS AND MATERIALS: Records were reviewed of 1,269 patients with carcinoma of the cervix (Stage IB to HI) treated with definitive irradiation (combination of external beam and two intracavitary insertions). Follow-up was obtained in 97% of patients (median, 12 years; minimum, 3 years; maximum, 28 years). The relationships between overall treatment time and time of brachytherapy and incidence of treatment sequelae were analyzed for each stage. RESULTS: Overall incidence of Grades 2 (moderate) sequelae was 7% and of Grade 3 (severe) sequelae, 11%. There was no significant correlation of various incidences of Grade 2 and 3 sequelae with overall treatment times (8% in patients treated in less than 7 weeks, 9% in 7.1 to 9 weeks, and 12% when treatment time was longer than 9 weeks) (p = 0.08). In patients with Stage IB and IIA tumors, incidence of rectal toxicity (mostly proctitis) was comparable in patients treated in less than 7 or 7.1 to 9 weeks (4.1 and 6%, respectively) and slightly higher in those treated in longer periods (11.5%) (p = 0.24). In patients with Stage IIB and III, the incidence of Grade 2 and 3 small bowel morbidity was 2% in those treated in less than 7 weeks, 6% for 7.1 to 9 weeks, and 4.9% for longer times (p < or = 0.01). This increased morbidity was also correlated with total dose of irradiation to the lateral pelvic wall: 5 of 257 (2%) for less than 60 Gy and 21 of 438 (4.8%) for higher doses (p < or = 0.01). There was no significant correlation between the timing of brachytherapy (usually two low dose rate intracavitary insertions performed within 4.5 to 6.5 weeks of initiation of external beam therapy) and significant treatment sequelae. CONCLUSIONS: We observed a varied average incidence of Grade 2 and 3 morbidity in the bladder, rectum, and small intestine with different overall treatment times, without a definite pattern to suggest an impact of prolongation of treatment time on morbidity. Likewise, there was no significant correlation with the timing of intracavitary insertions and morbidity of therapy. Because prolongation of the overall treatment time has a well-documented detrimental effect on pelvic tumor control and survival in carcinoma of the cervix with no significant impact on morbidity, it is imperative to deliver radiation therapy in the shortest possible time and without schedule interruptions. PMID- 12118563 TI - Correlation of pretherapy prostate cancer characteristics with histologic findings from pelvic lymphadenectomy specimens. AB - PURPOSE: The purpose of this study was to identify pretherapy factors associated with pelvic lymph node involvement (LNI) in patients with localized prostatic carcinoma (CaP), and to develop a model that would allow for estimation of this risk at the time of initial diagnosis. METHODS AND MATERIALS: Between January 1988 and December 1992, 2439 patients with clinical Stage T1a-3cN0-XM0 CaP underwent radical retropubic prostatectomy and bilateral pelvic lymph node dissection as sole initial therapy at a single medical institution. Preoperative factors were evaluated for their association with pelvic LNI in univariate and multivariate logistic regression analysis. A model was developed that incorporated independent predictive variables, and probability plots were generated to estimate the likelihood of pelvic LNI in the patient with a new diagnosis of localized CaP. RESULTS: Within clinical tumor stage, three groups (Tla-2a, T2b-c, and T3) were identified in which the observed rate of pelvic LNI was distinctly different. Gleason primary grades were also combined (1-2, 3, and 4-5) because of a similar observation. Univariate analysis identified clinical tumor stage (p < 0.0001), Gleason primary grade (p < 0.0001), and serum prostate specific antigen (p < 0.0001) as factors associated with pelvic LNI. Each of these variables retained independent significance (p < or = 0.0002) in the multivariate model. Patient age (p = 0.12) and history of prior transurethral resection of the prostate (p = 0.36) were not found to correlate with this endpoint. Probability plots provided an estimate of the likelihood for pelvic LNI according to the combination of pretherapy clinical tumor stage, Gleason primary grade, and serum prostate-specific antigen level. CONCLUSION: Clinical tumor stage as determined by digital rectal examination, Gleason primary grade of the diagnostic biopsy specimen, and pretherapy serum prostate-specific antigen value can be combined to estimate the probability of pelvic LNI for the patient with a new diagnosis of localized CaP. This information may be of value in directing the pretherapy diagnostic evaluation, as an aid in radiation therapy treatment planning, and in the conduct of clinical research efforts. PMID- 12118564 TI - Long-term outcome of radical radiation therapy for prostatic carcinoma: 1967 1987. AB - PURPOSE: This study was done to review long-term results of radical radiotherapy for prostate cancer. METHODS AND MATERIALS: The records of 674 patients with Stage T1a, T1b, T2a, T2b, T3, and any T,N1,M0 disease, treated with external beam radiotherapy between January 1, 1967 and December 1987, were reviewed. These patients were treated to an average total dose of 66 Gy, with an average fractional dose of 2.05 Gy, using megavoltage. The duration of follow-up for surviving patients ranged from a minimum of 7 years to more than 20 years. RESULTS: The survival for 151 Stage T1a,T1b patients was 98.5% at 5 years, 93.6% at 10 years, and 75.2% at 15 years. Survival for 346 Stage T2a,b patients was 94.4% at 5 years, 67.9% at 10 years, and 41.5% at 15 years. Survival for 92 Stage T3 patients was 87.3% at 5 years, 54% at 10 years, and 26.6% at 15 years. The survival for 85 any T,N1,M0 patients was 73.9% at 5 years, 34.4% at 10 years, and 8.5% at 15 years. At 15 years, 75.2% of Stage T1a,b patients, 41.5% of Stage T2a,b patients, 21.7% of Stage T3 patients, and 8.5% of Stage T,N1,M0 patients remained free of local recurrence and distant metastases. The elevation of prostatic acid phosphatase prior to radiotherapy was an unfavorable prognostic factor, with impact on both loco-regional recurrences and survival. CONCLUSIONS: The external beam radiotherapy for localized carcinoma of the prostate produced a good loco-regional control, NED, and overall survival. Patients with smaller tumors and low grade fared better than the ones with more aggressive and/or bulky tumors. The weakness of this study is the absence of serial prostate-specific measurements, which were not available during the period under study. The complication rate requiring surgical intervention was low, i.e. 0.4%. PMID- 12118565 TI - High-dose intraoperative radiotherapy for unresectable pancreatic cancer. AB - PURPOSE: The results of high-dose intraoperative radiotherapy (IORT) and/or external beam radiotherapy (EBRT) for unresectable pancreatic cancer were analyzed to evaluate the possible advantages of IORT in combination with EBRT. METHODS AND MATERIALS: Between 1983 and 1993, 115 patients with unresectable adenocarcinoma of the pancreas (53 with non-Stage IV disease and 62 with Stage IV disease) were treated with EBRT + IORT (55 patients), EBRT alone (44 patients), or IORT alone (16 patients). In non-Stage IV patients, the use of EBRT alone was due to the unavailability of IORT and the use of IORT alone was due to refusal of EBRT. The IORT dose was 30-33 Gy and the EBRT dose was 40-60 Gy. A historical control group comprised of 101 patients undergoing palliative surgery alone was also analyzed. RESULTS: Both non-Stage IV and Stage IV patients receiving EBRT with or without IORT had a better prognosis than the nonirradiated historical controls. Among non-Stage IV patients, the median survival of the EBRT + IORT group (8.5 months) and the EBRT group (8 months) was similar, although survival from 12 to 18 months was higher in the former group (38% vs. 10% at 12 months, p = 0.018, and 19% vs. 0% at 18 months, p = 0.023). In Stage IV patients, the prognosis was not influenced by the type of radiotherapy. Multivariate analysis revealed that a pretreatment carbohydrate antigen (CA) 19-9 level < 1000 U/ml was associated with better survival. In non-Stage IV patients with a CA 19-9 level < 1000 U/ ml, EBRT + IORT appeared to produce a better survival than EBRT alone (p = 0.047). This was supported by multivariate analysis. CONCLUSION: High-dose IORT + EBRT may be more effective than EBRT alone in patients with unresectable but localized pancreatic cancer and a low CA 19-9 level. PMID- 12118566 TI - Ten-year results of chemoradiation for anal cancer: focus on late morbidity. AB - PURPOSE: To evaluate (a) long-term survival and (b) the incidence and nature of long-term morbidity/ mortality related to chemoradiation using the anal cancer experience. METHODS AND MATERIALS: From January 1979 to April 1987,34 consecutive patients with Stage I (5 patients), II (15 patients), and II (14 patients) cancers of the anal canal were treated definitively with a chemoradiation regimen combining 41.4 Gy pelvic radiotherapy with two concurrent cycles of 5 fluorouracil and mitomycin C. Cumulative actuarial survival was calculated at 10 years and long-term morbidity was categorized per RTOG/EORTC late toxicity criteria. Specific criteria to grade anal toxicity were devised. RESULTS: Cumulative survival for all 34 patients was 92% at 5 years and 85% at 10 years. The most frequent late toxicity was chronic diarrhea in 17 (50%) patients. Five patients (15%) had Grade 3 or 4 late toxicities. Sexual dysfunction was present in 2 of 26 evaluable patients (7%). CONCLUSIONS: Excellent long-term survival and colostomy-free survival is possible for anal cancer patients treated definitively by chemoradiation. Late effects do not appear to be frequent or intense enough to deter the use of chemoradiation in anal cancer. The biologically expected increase in long-term toxicity when combining radiotherapy and chemotherapy is not substantiated by the results of this study. PMID- 12118567 TI - Interstitial pneumonitis following autologous bone-marrow transplantation conditioned with cyclophosphamide and total-body irradiation. AB - PURPOSE: To assess the influence of different total-body irradiation (TBI) regimens on interstitial pneumonitis (IP), we retrospectively analyzed our clinical data concerning an homogeneous group of patients conditioned with cyclophosphamide (CY) alone and single-dose or fractionated TBI before autologous bone-marrow transplantation (ABMT). METHODS AND MATERIALS: One hundred eighty-six patients with acute nonlymphoblastic leukemia (n = 101), acute lymphoblastic leukemia (n = 62), chronic myeloid leukemia (n = 11), non-Hodgkin's lymphoma (n = 10), and multiple myeloma (n = 2) referred to our department between May 13, 1981 and September 16, 1992, underwent TBI before ABMT. The male-to-female ratio was 123:63 (1.95), and mean and median age was 33 +/- 12 (6-63 years) and 35 years, respectively. Cyclophosphamide alone (60 mg/kg/day on each of 2 successive days) was used as conditioning chemotherapy in all patients. Patients were irradiated according to two techniques: either with single-dose (STBI) (n = 124; 10 Gy administered to the midplane at the level of L4, and 8 Gy to the lungs) or with fractionated (FTBI) (n = 62; 12 Gy in 6 fractions over 3 consecutive days to the midplane at the level of L4, and 9 Gy to the lungs) TBI. The mean instantaneous dose rate was 0.057 +/- 0.0246 Gy/min (0.0264-0.1692 Gy/min). It was < or = 0.048 Gy/min in 48 patients (LOW group), > 0.048 and < or = 0.09 Gy/min in 129 patients (MEDIUM group), and > 0.09 Gy/min in 9 patients (HIGH group). The median follow up period was 5 years (24-120 months). RESULTS: In January 1994, the 5-year overall (including all causes of death) and disease-free survival (DFS) rates were 50 and 48%, respectively. The 5-year DFS was 47.9% in the STBI group, and 47.8% in the FTBI group (p = 0.77). It was 44% in the HIGH group, 53% in the MEDIUM group, and 34% in the LOW group (LOW vs. MEDIUM, p = 0.009). The 5-year IP incidence was 17% in all patients, 16% in the STBI group and 18% in the FTBI group (p = 0.37), but it was significantly higher in patients receiving high instantaneous dose rate TBI (56% in the HIGH, 13% in the MEDIUM, 20% in the LOW groups; HIGH vs. MEDIUM, p = 0.002). However, sex (p = 0.37), age (18% for > 20 vs. 10% for < or = 20 years, p = 0.37), and body weight (> 60 kg vs. < or = 60 kg, p = 0.09) did not influence the IP incidence in univariate analyses. Multivariate analysis (Cox model) revealed that the instantaneous dose rate (p = 0.05), and the age (p = 0.04) were the two independent factors influencing the incidence of IP. CONCLUSION: This retrospective study including only the patients transplanted with ABMT conditioned with CY alone and STBI or FTBI concluded that instantaneous dose rate and age significantly influenced the incidence of IP, whereas sex, body weight, and fractionation did not. PMID- 12118569 TI - Effect of radiation dose rate and cyclophosphamide on pulmonary toxicity after total body irradiation in a mouse model. AB - PURPOSE: Interstitial pneumonitis (IP) is still a major complication after total body irradiation (TBI) and bone marrow transplantation (BMT). It is difficult to determine the exact role of radiation in this multifactorial complication, especially because most of the experimental work on lung damage was done using localized lung irradiation and not TBI. We have thus tested the effect of radiation dose rate and combining cyclophosphamide (CTX) with single fraction TBI on lung damage in a mouse model for BMT. METHODS AND MATERIALS: TBI was given as a single fraction at a high dose rate (HDR, 0.71 Gy/min) or a low dose rate (LDR, 0.08 Gy/min). CTX (250 mg/kg) was given 24 h before TBI. Bone marrow transplantation (BMT) was performed 4-6 h after the last treatment. Lung damage was assessed using ventilation rate (VR) and lethality between 28 and 180 days (LD(50/28-180)). RESULTS: The LD50 for lung damage, +/- standard error (SE), increased from 12.0 (+/- 0.2) Gy using single fraction HDR to 15.8 (+/- 0.6) Gy using LDR. Adding CTX shifted the dose-response curves towards lower doses. The LD50 values for the combined treatment were 53 (+/- 0.2) and 3.5 (+/- 0.2) Gy for HDR and LDR, respectively. This indicates that the combined effect of CTX and LDR was more toxic than that of combined CTX and HDR. Lung damage evaluated by VR demonstrated two waves of VR increase. The first wave of VR increase occurred after 6 weeks using TBI only and after 3 weeks in the combined CTX-TBI treatment, irrespective of total dose or dose rate. The second wave of VR elevation resembled the IP that follows localized thoracic irradiation in its time of occurrence. CONCLUSIONS: Lung damage following TBI could be spared using LDR. However, CTX markedly enhances TBI-induced lung damage. The combination of CTX and LDR is more toxic to the lungs than combining CTX and HDR. PMID- 12118568 TI - Fused pyrazine mono-n-oxides as bioreductive drugs. II Cytotoxicity in human cells and oncogenicity in a rodent transformation assay. AB - PURPOSE: To determine what structural moieties of the fused pyrazine mono-N oxides are determining factors in their in vitro cytotoxicity and oncogenicity. METHODS AND MATERIALS: A new series of experimental bioreductive drugs, fused pyrazine mono-N-oxides, was evaluated in vitro for aerobic and hypoxic cytotoxicity in the HT29 human colon adenocarcinoma cell line by using clonogenic assays. The relative oncogenicities of these compounds were also determined in aerobic cultures of C3H 10T1/2 mouse embryo fibroblasts by using a standard transformation assay. RESULTS: Removal of the 4-methyl piperazine side chain from the parent compound, RB 90740, reduced the potency of the hypoxic cytotoxin. Reduction of the N-oxide function increased the aerobic cytotoxicity and eliminated most of the hypoxic/aerobic cytotoxic differential. The reduced N oxide also had significant oncogenicity, consistent with a mechanism of genotoxicity following bioreduction of RB 90740. CONCLUSION: This new series of bioreductive compounds may be effective in cancer therapy, particularly the lead compound RB 90740. The oncogenic potential of these compounds is similar to that for other cancer therapies. Further studies should include evaluation of these compounds in vivo and the development of analogs with reduced oncogenic potential and retention of the hypoxic/aerobic cytotoxicity differential. PMID- 12118571 TI - Schering-Plough agrees to pay FDA $500 million and bring factories into compliance. PMID- 12118570 TI - Detection of individual hypoxic cells in multicellular spheroids by flow cytometry using the 2-nitroimidazole, EF5, and monoclonal antibodies. AB - PURPOSE: The purpose of this work was to evaluate EF5, a 2-nitroimidazole compound, and anti-EF5 antibodies as a method to quantify radiobiologically hypoxic cells. METHODS AND MATERIALS: Multicellular spheroids of EMT6 mammary sarcoma cells were used as a model to identify hypoxic cells that were resistant to radiation damage. This was accomplished by incubating the spheroids with the 2 nitroimidazole (EF5), which forms hypoxia-dependent adducts with cellular macromolecules that are detected by fluorescent monoclonal antibodies. RESULTS: Cells from spheroids grown for 2 days in sealed flasks had an increased surviving fraction following radiation as compared to fully reoxygenated spheroids, indicating the presence of radiobiological hypoxia. Treatment of the spheroids with EF5 and subsequent immunohistochemical staining of cryosections with an anti EF5 fluorochrome conjugated monoclonal antibody allowed for the identification of EF5-adduct containing cells. Spheroids grown under hypoxic conditions in the presence of EF5 showed limited staining of the peripheral cell layers, intense staining of the interior, and an absence of staining within the necrotic center. In contrast, there was minimal staining in reoxygenated spheroids and no staining in control spheroids incubated in the absence of EF5. Flow cytometric analysis of single cells dissociated from spheroids allowed for the calculation of the percentage of stained cells, as well as the intensity of staining. A comparison of the intensity of staining of EF5 treated hypoxic spheroids with the intensity of staining of single cells incubated with EF5 under controlled oxygen concentrations was used to estimate the oxygen concentration range within spheroids. Selective dissociation of spheroids provided a direct demonstration that the cells containing the highest level of EF5 binding were also the cells with increased radiation resistance. CONCLUSION: This technique provides an excellent means of detecting and quantifying hypoxia, which should be directly applicable in tumors. PMID- 12118572 TI - Mare reproductive loss syndrome returns, but losses are fewer. PMID- 12118573 TI - Promoting animal welfare amidst conflict. PMID- 12118574 TI - Questions reported risks of Salmonella infections. PMID- 12118575 TI - Incorrect dose of detomidine. PMID- 12118576 TI - What is your diagnosis? Aggressive bone lesion involving the left elbow. PMID- 12118577 TI - ECG of the month. Ventricular tachycardia. PMID- 12118578 TI - Is the bitch sexually intact? The ovaries have been removed in this bitch. PMID- 12118579 TI - What is pain? PMID- 12118580 TI - Pain management in laboratory animals--are we meeting the challenge? PMID- 12118581 TI - Pain and distress in agricultural animals. PMID- 12118582 TI - Treatment of pain in dogs and cats. PMID- 12118583 TI - Surgical trauma and chronically painful conditions--within our comfort level but beyond theirs? PMID- 12118584 TI - Alternative methods for the control of pain. PMID- 12118585 TI - Ethical issues regarding pain in animals. PMID- 12118586 TI - Managing pain in human neonates--applications for animals. PMID- 12118587 TI - Evaluation of an in-house enzyme-linked immunosorbent assay for quantitative measurement of serum total thyroxine concentration in dogs and cats. AB - OBJECTIVE: To compare serum total thyroxine (T4) concentrations obtained with an in-house ELISA and a validated radioimmunoassay (RIA). DESIGN: Laboratory trial. SAMPLE POPULATION: 50 canine and 50 feline serum samples submitted for measurement of total T4 concentration with the RIA; samples were selected to represent a wide range of concentrations (< 6 to 167 nmol/L). PROCEDURE: Results of the ELISA and RIA were compared by calculating correlation coefficients, examining linearity, determining bias and precision, and evaluating clinical interpretations. RESULTS: Correlation coefficients for results of the 2 methods were 0.84 for the canine samples and 0.59 for the feline samples. Examination of bias plots revealed large variations in ELISA results, compared with RIA results. For the feline samples, the ELISA consistently overestimated total T4 concentration obtained with the RIA. When results of the 2 methods were categorized (low, borderline low, normal, borderline high, or high), results were discordant for 24 (48%) and 29 (58%) of the canine samples and for 18 (36%) and 28 (56%) of the feline samples (depending on whether borderline high ELISA results were considered normal or high). Reliance on results of the ELISA would have led to inappropriate clinical decisions for 31 (62%) canine samples and 25 (50%) feline samples. The ELISA coefficients of variation for the pooled canine and feline samples were 18 and 28%, respectively. CONCLUSIONS AND CLINICAL RELEVANCE: Substantial discrepancies between ELISA and RIA results for T4 concentrations were detected. Thus, we concluded that the in-house ELISA kit was not accurate for determining serum total T4 concentrations in dogs and cats. PMID- 12118588 TI - Use of enrofloxacin for treatment of large-form Haemobartonella felis in experimentally infected cats. AB - OBJECTIVE: To compare treatment with enrofloxacin and doxycycline with no treatment in cats experimentally infected with Haemobartonella felis. DESIGN: Prospective case-control study. ANIMALS: 16 cats. PROCEDURE: Cats were inoculated with large-form H. felis from a chronically infected donor. Cats were assigned to 1 of 4 treatment groups: doxycycline (5 mg/kg [2.3 mg/lb], p.o., q 12 h), low dose enrofloxacin (5 mg/kg, p.o., q 24 h), high-dose enrofloxacin (10 mg/kg [4.5 mg/lb], p.o., q 24 h), and an untreated control group. Clinical signs, Hct, blood smears, and a polymerase chain reaction (PCR) assay were used to monitor progression of the infection. RESULTS: All cats were confirmed to be infected with H. felis via blood smear evaluations and PCR assay results. Treatment had no effect on Hct during the intratreatment period, but Hct values were significantly greater in the low-dose enrofloxacin group, compared with the control group, during the posttreatment period. During the intratreatment period, H. felis organism counts per 1,000 RBC in the doxycycline treatment and the high-dose enrofloxacin treatment groups decreased at a significantly faster rate than those in the control group. In the posttreatment period, organism counts in the doxycycline treatment group and the low- and high-dose enrofloxacin groups decreased at significantly faster rates than counts in the control group. There was no significant effect of treatment on the number of positive PCR assay results. Two cats treated with enrofloxacin and 1 cat treated with doxycycline completely cleared the H. fe is organism despite presumed immunosuppression caused by glucocorticoids. CONCLUSIONS AND CLINICAL RELEVANCE: Results support the hypothesis that enrofloxacin has anti-H. felis effects. PMID- 12118589 TI - Effect of topical application of fipronil in cats with flea allergic dermatitis. AB - OBJECTIVE: To determine whether topical application of a 10% fipronil solution would control signs of flea allergic dermatitis in cats housed under natural conditions. DESIGN: Multicenter open clinical trial. ANIMALS: 42 client-owned cats with flea allergic dermatitis. PROCEDURES: Study cats along with all other cats and dogs living in the same houses were treated with 10% fipronil solution topically on days 0, 30, and 60. Flea counts and clinical assessments were performed on study cats on days 0, 14, 30, 60, and 90. RESULTS: Percentage reductions in geometric mean flea counts on days 14, 30, 60, and 90, compared with day-0 geometric mean count, were 75, 73, 85, and 94%, respectively. Pruritus score was significantly improved at each examination after day 0, and pruritus was reduced or eliminated in 31 of 40 (78%) cats at the final examination. Similarly, scores for severity of miliary dermatitis and alopecia were significantly improved at each examination, except for alopecia score on day 14. Overall treatment efficacy, assessed on day 90, was excellent for 28 (70%) cats, good for 6 (15%), moderate for 3 (7.5%), and poor for 3 (7.5%). CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that monthly topical application of fipronil is effective for treatment of flea allergic dermatitis in cats housed under natural conditions. PMID- 12118590 TI - Acute necrotizing dermatitis and septicemia after application of a d-limonene based insecticidal shampoo in a cat. AB - A 2-year-old female spayed domestic shorthair cat was examined because of lethargy, inappetance, vocalization, and abnormal aggressive behavior of 1 day's duration. The cat had been groomed the previous day with a d-limonene-based insecticidal shampoo. Skin lesions consisted of coalescing erythematous patches. Despite supportive care, the cat's condition deteriorated. Dermatohistopathologic changes included multifocal areas of acute coagulative epidermal necrosis. The dermis was infiltrated by a dense population of bacilli. d-Limonene toxicosis has been rarely described in dogs and cats. Toxic effects such as hypersalivation, ataxia, shivering, hypothermia, scrotal irritation, hypotension, and erythema multiforme major have been reported. Treatment for septicemia and disseminated intravascular coagulation, along with intensive supportive care, may be necessary. PMID- 12118591 TI - Hypercalcemia and high parathyroid hormone-related protein concentration associated with malignant melanoma in a dog. AB - A 12-year-old Cocker Spaniel with an oral malignant melanoma was evaluated for progressive lethargy and anorexia. No metastases were identified during antemortem evaluation, but severe hypercalcemia was evident. Antemortem diagnostic testing failed to identify a cause for the hypercalcemia. No neoplasms other than the melanoma were identified on postmortem examination. Serum parathyroid hormone-related protein concentration was markedly high, and the melanoma had moderate to marked immunostaining for this protein. Paraneoplastic syndromes are rare in dogs with malignant melanoma. PMID- 12118592 TI - Use of lufenuron as a treatment for fungal endometritis in four mares. AB - Lufenuron, a benzoylphenyl urea derivative, was evaluated as a treatment for endometrial fungal infections in 4 mares. Intrauterine lavage was performed with lufenuron suspended in sterile saline (0.9% NaCl) solution. Cytologic evaluation and fungal culture of the endometrium, as well as subsequent reproductive performance, were used to monitor efficacy of this treatment. Fungal endometritis in mares is associated with infertility. Treatment is often ineffective and costly, and recurrence following treatment is not uncommon. Intrauterine infusions of lufenuron were effective in eliminating fungal endometritis in the 4 mares of this report. PMID- 12118593 TI - Antimicrobial susceptibility patterns of Salmonella isolates from cattle in feedlots. AB - OBJECTIVE: To evaluate the antimicrobial susceptibility patterns of Salmonella isolates from feedlot cattle. DESIGN: Cross-sectional study. SAMPLE POPULATION: 263 Salmonella isolates. PROCEDURES: Fecal samples were collected from the floor of 2 pens in each of 100 feedlots. Two hundred eighty Salmonella isolates were recovered after bacteriologic culture from 38 pens. Of these, 263 isolates were available for antimicrobial susceptibility testing to 16 antimicrobials, using microbroth dilution breakpoint plates. RESULTS: Less than 5% of isolates were resistant to any of the antimicrobials tested, with the exception of sulfamethoxazole (15; 5.7%) and tetracycline (61; 23.2%). Most isolates (197; 74.9%) were susceptible to all antimicrobials tested, whereas 18 (6.8%) were resistant to 2 or more antimicrobials. The percentage of isolates with resistance to any antimicrobial varied by serotype. The percentage of isolates resistant to various antimicrobials was not related to concurrent use of antimicrobials in the feed. CONCLUSIONS AND CLINICAL RELEVANCE: With the exception of tetracycline and sulfamethoxazole, resistance of Salmonella isolates to any of the antimicrobials was uncommon. Most isolates were susceptible to all antimicrobials tested. Antimicrobial resistance was not related to the presence of antimicrobials in the ration being fed at the time of sample collection. PMID- 12118594 TI - Udder cleft dermatitis and sarcoptic mange in a dairy herd. AB - OBJECTIVE: To determine prevalence of udder cleft dermatitis in a dairy herd that was experiencing an outbreak of sarcoptic mange. DESIGN: Clinical survey. ANIMALS: 1,597 Holstein cows and late-gestation heifers. PROCEDURE: Animals were examined for udder cleft dermatitis and for skin lesions consistent with sarcoptic or chorioptic mange. Skin scrapings were collected from 56 cows and examined for ectoparasites. The herd was revisited 1 year later, and prevalences of udder cleft dermatitis and lesions consistent with mange were determined in 506 cows. RESULTS: Of the 1,597 cattle examined, 280 (18%) had udder cleft dermatitis, and 1,397 (87.5%) had lesions consistent with mange. In 43 of 56 (77%) cows, skin scrapings revealed Sarcoptes mites. Udder cleft dermatitis was significantly more common in older than in younger cows. In first-lactation cows, udder cleft dermatitis was less common during the first 4 months of lactation than in the later stages of lactation, but udder cleft dermatitis was identified in cows in all stages of lactation and in cows that were not lactating. The herd was treated with eprinomectin to control mites, and prevalence of lesions consistent with mange 1 year later was only 2.8%. However, prevalence of udder cleft dermatitis was still 12%. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that cows in any stage of lactation and cows that are not lactating can have udder cleft dermatitis but that lesions are more common in older cows. Control of sarcoptic mange was accompanied by a moderate reduction in the prevalence of udder cleft dermatitis but did not eliminate the condition. PMID- 12118595 TI - Congenital erythropoietic protoporphyria in a Limousin calf. AB - Bovine congenital erythropoietic protoporphyria is an uncommon genetic defect in Limousin and Blonde d'Aquitaine cattle that is characterized by severe photosensitization. Clinical signs include intense pruritus and exudative dermatitis involving the face, pinnae, and dorsal aspect of the thorax. Affected cattle have hematologic and serum biochemical values within reference ranges, and their teeth are normochromic. Definitive diagnosis of bovine congenital erythropoietic protoporphyria is accomplished by genetic testing. Affected cattle should be sent to a terminal market. PMID- 12118596 TI - Surgical removal of a seminoma from a black sea bass. AB - An adult black sea bass was examined because of abdominal distention and decreased appetite. A large abdominal swelling was evident and was firm on palpation. Differential diagnoses included neoplasia, abscess or granuloma, hematoma, or swim bladder abnormality. Diagnostic tests included survey radiography, positive-contrast radiography, and computed tomography. The sea bass was anesthetized with tricaine methanesulfonate. A ventral midline abdominal incision was made, and adhesions to the mass were gently dissected. The fish recovered without complications. Radiography was repeated 8 weeks after surgery, and there was no evidence of mass regrowth. To the authors' knowledge, this is the first report of a barium enema being performed in a fish. Although surgical procedures are performed more commonly on fish for research, a few reports of clinical surgical cases have been described. Our experience supports the conclusions of other reports that certain surgical procedures can be performed safely in fish. PMID- 12118597 TI - Monte Carlo simulation of the Leksell Gamma Knife: I. Source modelling and calculations in homogeneous media. AB - The Monte Carlo code PENELOPE has been used to simulate photon flux from the Leksell Gamma Knife, a precision method for treating intracranial lesions. Radiation from a single 6OCo assembly traversing the collimator system was simulated, and phase space distributions at the output surface of the helmet for photons and electrons were calculated. The characteristics describing the emitted final beam were used to build a two-stage Monte Carlo simulation of irradiation of a target. A dose field inside a standard spherical polystyrene phantom, usually used for Gamma Knife dosimetry, has been computed and compared with experimental results, with calculations performed by other authors with the use of the EGS4 Monte Carlo code, and data provided by the treatment planning system Gamma Plan. Good agreement was found between these data and results of simulations in homogeneous media. Owing to this established accuracy, PENELOPE is suitable for simulating problems relevant to stereotactic radiosurgery. PMID- 12118598 TI - Monitor unit calculations for wedged asymmetric photon beams. AB - Algorithms for calculating monitor units (MUs) in wedged asymmetric high-energy photon beams as implemented in treatment planning systems have their limitations. Therefore an independent method for MU calculation is necessary. The aim of this study was to develop an empirical method to determine MUs for points at the centre of wedged fields, asymmetric in two directions. The method is based on the determination of an off-axis factor (OAF) that corrects for the difference in dose between wedged asymmetric and wedged symmetric beams with the same field size. Measurements were performed in a water phantom irradiated with 6 and 18 MV photon beams produced by Elekta accelerators, which are fitted with an internal motorized wedge that has a complex shape. The OAF perpendicular to the wedge direction changed significantly with depth for the 18 MV beam. Dose values measured for a set of 18 test cases were compared with those calculated with our method. The maximum difference found was 6.5% and in 15 cases this figure was smaller than 2.0%. The analytical method of Khan and the empirical method of Georg were also tested and showed errors up to 12.8%. It can be concluded that our simple formalism is able to calculate MUs in wedged asymmetric fields with an acceptable accuracy in most clinical situations. PMID- 12118599 TI - Operator-free, film-based 3D seed reconstruction in brachytherapy. AB - In brachytherapy implants, the accuracy of dose calculation depends on the ability to localize radioactive sources correctly. If performed manually using planar images, this is a time-consuming and often error-prone process-primarily because each seed must be identified on (at least) two films. In principle, three films should allow automatic seed identification and position reconstruction; however, practical implementation of the numerous algorithms proposed so far appears to have only limited reliability. The motivation behind this work is to create a fast and reliable system for real-time implant evaluation using digital planar images obtained from radiotherapy simulators, or mobile x-ray/fluoroscopy systems. We have developed algorithms and code for 3D seed coordinate reconstruction. The input consists of projections of seed positions in each of three isocentric images taken at arbitrary angles. The method proposed here consists of a set of heuristic rules (in a sense, a learning algorithm) that attempts to minimize seed misclassifications. In the clinic, this means that the system must be impervious to errors resulting from patient motion as well as from finite tolerances accepted in equipment settings. The software program was tested with simulated data, a pelvic phantom and patient data. One hundred and twenty permanent prostate implants were examined (105 125I and 15 103Pd) with the number of seeds ranging from 35 to 138 (average 79). The mean distance between actual and reconstructed seed positions is in the range 0.03-0.11 cm. On a Pentium III computer at 600 MHz the reconstruction process takes 10-30 s. The total number of seeds is independently validated. The process is robust and able to account for errors introduced in the clinic. PMID- 12118600 TI - Performance of isotropic light dosimetry probes based on scattering bulbs in turbid media. AB - In a previous paper the calibration of an isotropic light detector in clear media was described and validated. However, in most applications the detector is used to measure light distribution in turbid (scattering) media, that is, in tissues or tissue equivalent optical phantoms. Despite its small diameter (typically 0.8 mm), inserting the detector in a turbid medium may perturb the light distribution and change the fluence rate at the point of measurement. In the present paper we estimate the error in the fluence rate measured by a detector in turbid media after calibration in a clear medium (air), using an optical phantom and detector bulbs of different optical properties. The experimental results are compared with calculations using the diffusion approximation to the transport equation in a spherical geometry. From measurements in optical phantoms and the results of the calculations it appears that introduction of the detector into a water-based turbid medium with refractive index, absorption- and scattering coefficients different from those of the detector bulb may require corrections to the detector response of up to 10-15%, in order to obtain the true fluence rate in that medium. The diffusion model is used to explore the detector response in a number of tissues of interest in photodynamic therapy, using tissue optical properties from the literature. Based on these model calculations it is estimated that in real tissues the fluence rate measured by the detector is up to 3% below the true value. PMID- 12118601 TI - Optical properties of selected native and coagulated human brain tissues in vitro in the visible and near infrared spectral range. AB - Medical laser applications require knowledge about the optical properties of target tissue. In this study, the optical properties of selected native and coagulated human brain structures were determined in vitro in the spectral range between 360 and 1100 nm. The tissues investigated included white brain matter, grey brain matter, cerebellum and brainstem tissues (pons, thalamus). In addition, the optical properties of two human tumours (meningioma, astrocytoma WHO grade II) were determined. Diffuse reflectance, total transmittance and collimated transmittance of the samples were measured using an integrating-sphere technique. From these experimental data, the absorption coefficients, the scattering coefficients and the anisotropy factors of the samples were determined employing an inverse Monte Carlo technique. The tissues investigated differed from each other predominantly in their scattering properties. Thermal coagulation reduced the optical penetration depth substantially. The highest penetration depths for all tissues investigated were found in the wavelength range between 1000 and 1100 nm. A comparison with data from the literature revealed the importance of the employed tissue preparation technique and the impact of the theoretical model used to extract the optical coefficients from the measured quantities. PMID- 12118602 TI - Maps of optical differential pathlength factor of human adult forehead, somatosensory motor and occipital regions at multi-wavelengths in NIR. AB - The optical differential pathlength factor (DPF) is an important parameter for physiological measurement using near infrared spectroscopy, but for the human adult head it has been available only for the forehead. Here we report measured DPF results for the forehead, somatosensory motor and occipital regions from measurements on 11 adult volunteers using a time-resolved optical imaging system. The optode separation was about 30 mm and the wavelengths used were 759 nm, 799 nm and 834 nm. Measured DPFs were 7.25 for the central forehead and 6.25 for the temple region at 799 nm. For the central somatosensory and occipital areas (10 mm above the inion), DPFs at 799 nm are 7.5 and 8.75, respectively. Less than 10% decreases of DPF for all these regions were observed when the wavelength increased from 759 nm to 834 nm. To compare these DPF maps with the anatomical structure of the head, a Monte Carlo simulation was carried out to calculate DPF for these regions by using a two-layered semi-infinite model and assuming the thickness of the upper layer to be the sum of the thicknesses of scalp and skull, which was measured from MRI images of a subject's head. The DPF data will be useful for quantitative monitoring of the haemodynamic changes occurring in adult heads. PMID- 12118604 TI - Application of the photon average trajectories method to real-time reconstruction of tissue inhomogeneities in diffuse optical tomography of strongly scattering media. AB - The possibility of application of the photon average trajectories (PAT) method to real-time reconstruction of tissue inhomogeneities in diffuse optical tomography of strongly scattering media has been substantiated. By this method, the inverse problem is reduced to solution of the integral equation with integration along a conditional PAT. Such an approach allows the standard fast algebraic algorithms commonly used in projection computed tomography to be applied to diffuse optical image reconstruction. To demonstrate the capabilities of the PAT method, a numerical experiment on cross-sectional reconstruction of cylindrical strongly scattering objects with absorbing inhomogeneities has been done. Relative shadows caused by inhomogeneities are simulated via numerical solution of the non stationary diffusion equation. To solve the inverse problem, the QR-factorization least-squares algorithm and the multiplicative algebraic reconstruction technique are used. The results are compared with those obtained by a well-known software package for temporal optical absorption and scattering tomography based on multiple solution of the diffusion equation. It is shown that the PAT method allows reconstruction of the optical structure of objects with comparable accuracy while saving reconstruction time considerably. PMID- 12118603 TI - A new optical method for the non-invasive detection of minimal tissue alterations. AB - Histological analysis, which is used to detect and diagnose most tissue alterations, requires an invasive biopsy procedure and a time-consuming tissue treatment, which limit its efficiency in providing rapid, cost-effective diagnosis and hinder the longitudinal study of tissue alteration. To address these limitations, we have developed a novel procedure, using the features of elastic-scattering spectroscopy, for a real-time, non-invasive analysis of tissues. We have tested whether this approach can detect in vivo changes in mouse skin induced by a single exposure to either complete Freund's adjuvant or 12-O tetradecanoylphorbol-13-acetate, two drugs known to induce discrete alterations of epidermis and dermis, without obvious changes on the skin surface. Here we report that the evaluation of localized absorption and reduced scattering coefficients permitted the detection of changes in skin regions that showed histological alterations, but not in regions which failed to be modified by the drugs. Results show that the optical in vivo analysis of small regions has sufficient specificity and sensitivity to detect minimal alterations of superficial tissues. In view of the prominent involvement of mucosal alterations in most human diseases, including carcinomas, the method provides a useful complement to standard biopsy, notably for the in vivo screening of early in situ epithelial alterations. PMID- 12118605 TI - Image reconstruction from limited angle Compton camera data. AB - The Compton camera is used for imaging the distributions of gamma ray direction in a gamma ray telescope for astrophysics and for imaging radioisotope distributions in nuclear medicine without the need for collimators. The integration of gamma rays on a cone is measured with the camera, so that some sort of inversion method is needed. Parra found an analytical inversion algorithm based on spherical harmonics expansion of projection data. His algorithm is applicable to the full set of projection data. In this paper, six possible reconstruction algorithms that allow image reconstruction from projections with a finite range of scattering angles are investigated. Four algorithms have instability problems and two others are practical. However, the variance of the reconstructed image diverges in these two cases, so that window functions are introduced with which the variance becomes finite at a cost of spatial resolution. These two algorithms are compared in terms of variance. The algorithm based on the inversion of the summed back-projection is superior to the algorithm based on the inversion of the summed projection. PMID- 12118606 TI - Quantitative elasticity imaging: what can and cannot be inferred from strain images. AB - We examine the inverse problem associated with quantitative elastic modulus imaging: given the equilibrium strain field in a 2D incompressible elastic material, determine the elastic stiffness (shear modulus). We show analytically that a direct formulation of the inverse problem has no unique solution unless stiffness information is known a priori on a sufficient portion of the boundary. This implies that relative stiffness images constructed on the assumption of constant boundary stiffness are in error, unless the stiffness is truly constant on the boundary. We show further that using displacement boundary conditions in the forward incompressible elasticity problem leads to a nonunique inverse problem. Indeed, we give examples in which exactly the same strain field results from different elastic modulus distributions under displacement boundary conditions. We also show that knowing the stress on the boundary can, in certain configurations, lead to a well-posed inverse problem for the elastic stiffness. These results indicate what data must be taken if the elastic modulus is to be reconstructed reliably and quantitatively from a strain image. PMID- 12118607 TI - Does amniotic fluid volume affect fetofetal transfusion in monochorionic twin pregnancies? Modelling two possible mechanisms. AB - Clinical evidence suggests that increased amniotic fluid volume due to polyhydramnios increases placental vascular resistance. We have sought to model the possible effects of an increased amniotic fluid volume on the net fetofetal transfusion in monochorionic twin pregnancies. We wanted to compare these effects with the results of previous simulations, which aimed to explain why the twin twin transfusion syndrome (TTTS) placentas more often include bidirectional arteriovenous (AV) rather than AV plus arterioarterial (AA) anastomoses. We extended our mathematical model of TTTS by simulating two different mechanisms that increase the placental vascular resistance as a consequence of polyhydramnios. First, there is an increase in the placental capillary resistance and hence in deep AV and opposite AV (denoted as VA) resistances due to polyhydramnios. Second, there is an increase in the resistance of chorionic veins due to polyhydramnios, assuming that these veins act as Starling resistors. We then simulated the effects of polyhydramnios on different placental anastomotic patterns. The results were as follows. In the first mechanism (polyhydramnios affects AV-VA resistances), an increased amniotic fluid volume hardly affected bidirectional AV, but slightly decreased fetofetal transfusion in AV plus AA anastomoses. However, for these effects to change the natural development of the pregnancy, polyhydramnios needed to persist for approximately 4 weeks, and by comparing the effects of polyhydramnios with the effects of amnioreduction, amnioreduction was more beneficial for normalizing the donor amniotic fluid volume. Therefore, these beneficial effects due to polyhydramnios have no practical clinical significance. In the second mechanism (Starling resistor for chorionic veins), polyhydramnios slightly increased fetofetal transfusion and hence slightly increased TTTS severity in bidirectional AV and AV plus VV, but did not affect AV plus AA anastomoses. In conclusion, we hypothesize that the simulated effects of polyhydramnios are not the primary cause of the fact that TTTS placentas more often include bidirectional AV than AV plus AA anastomoses. Rather, the more likely explanation is the previously identified larger range of AA than VA anastomotic diameters that adequately compensate for the effects of the AV. PMID- 12118608 TI - The theoretical relation of scalp Laplacian and scalp current density of a spherical shell head model. AB - The theoretical relation between the scalp Laplacian (SL) and the scalp current density (SCD) is derived for a spherical shell head model. The result shows that they are related by a function of spatial frequency. For practically available low spatial frequencies, they are approximately linearly related to each other, so the SL estimate may be considered as an approximate SCD estimate in practice. PMID- 12118609 TI - A dosimetric comparison of various multileaf collimators. AB - The dosimetric characteristics of three multileaf collimator (MLC) systems (Elekta, Siemens and Varian) having 10 mm leaf width are compared. A 6 MV photon beam was used from each unit for measurements. Film dosimetry was performed for the measurements and the analysis techniques were exactly duplicated in each system. Two of the collimators have rounded leaf ends (Elekta and Varian) and the third (Siemens) has a flat end that follows beam divergence. A scanning densitometer (Wellhofer with 0.45 mm spot and 0.5 mm step size) was used for film analysis. The dosimetric characteristics studied include: penumbra width (80-20%) as a function of position of the leaf end in the field, inter- and intra-leaf radiation leakage, dose distribution of the tongue and groove, and isodose curves for stepped leaves forming 45 degrees angle beam edge. Results show that MLC designs with divergent and non-divergent leaves produce penumbra (80-20%) widths that are within 2.0 mm of each other. However, the distance of the collimator from the x-ray target plays an important role, and the smallest penumbra width was noted for the Varian MLC despite its rounded leaf-end design. Compared to the other systems, this collimator is positioned about 15 cm closer to the patient which affects the skin dose. The MLC with flat leaf end, although closer to the target, showed slightly poorer penumbra width. Inter-leaf leakage through the leaves is 1.3% for two of the collimators (Elekta and Varian) with the backup jaws and is nearly 1% for the third system (Siemens). The Siemens MLC produces reduced tongue-and-groove effect compared to the other two collimators (Elekta and Varian). The isodose undulation for a stepped edge is found to be significant for the collimator closest to the patient (Varian) and does not depend on the leaf-end shape. There is no perfect MLC system that can be recommended, rather each one has unique advantages and disadvantages that should be weighed with comfort, ease and cost effectiveness for clinical use. PMID- 12118610 TI - Dosimetric characterization of a new miniature multileaf collimator. AB - The dosimetrical characteristics of a new miniature multileaf collimator (ModuLeaf MLC, MRC Systems GmbH, Heidelberg, Germany) attached to the accessory holder of a Siemens accelerator with 6 MV x-rays (PRIMUS, Siemens OCS, Concord, California, USA) have been investigated. In particular, those parameters which are important for the accuracy of the treatment such as output factors, penumbra, field edge precision and transmission/leakage were determined. These data can now be used to implement specific dose calculation procedures for this miniature multileaf collimator in treatment planning systems. PMID- 12118611 TI - Response of the FBX system to a carbon beam: its potential as a dosimeter in heavy particle radiotherapy. AB - The FBX aqueous chemical dosimeter contains 0.2 mol m(-3) ferrous ammonium sulphate, 5.0 mol m(-3) benzoic acid and 0.20 mol m(-3) xylenol orange in 40.0 mol m(-3) sulphuric acid. The dosimeter can measure photon and electron doses in the range 0.1 to 3000 cGy in radiotherapy. The response of this dosimeter was measured for a 53.2 MeV carbon beam in the present work. Our initial result indicates that the sensitivity of the FBX system to the carbon beam as compared to cobalt-60 gamma rays is 25.5%, and thus we believe that the FBX system could be a useful dosimeter for carbon beams and similar heavy ions considered useful in radiotherapy. PMID- 12118612 TI - Sulphate-reducing laboratory-scale high-rate anaerobic reactors for treatment of metal- and sulphate-containing mine wastewater. AB - Upflow anaerobic sludge blanket (UASB) reactors were used in this study to evaluate the feasibility of the sulphate-reducing, anaerobic high-rate process to treat metal- and sulphate-containing mining wastewater (MWW). Four simultaneous reactors, inoculated with different inocula (mesophilic granular sludge from two UASB reactors, one treating sugar refinery wastewater and the other board mill wastewater) and operated with different loadings, were for 95 days fed with synthetic feed consisting of glucose and sulphate. In all reactors, 23-72% of sulphate and 12-93% of COD were removed. Subsequently, two reactors were fed with diluted MWW (zinc as the main metal) for 77 days with hydraulic retention times down to 8 hours. At the onset of the runs (until day 48), over 99.9% of zinc was removed in both reactors, after which removals fell to less than 30-80%. At the end of the runs, the highest zinc content (44 mg g(-1) TS) in the reactor sludges was 21 times higher than that in the inoculum. It cannot be concluded definitively that sulphide precipitation was the only mechanism of metal removal, for biosorption may have had a role to play in the process. PMID- 12118613 TI - Optimization of multistage phenol adsorption by organobentonites: theoretical developments and experimental verification. AB - Material balance equations were developed for representing phenol adsorption by organobentonites in a countercurrent multistage adsorption process. The developed equations were employed to analyze the adsorption performance of the multistage process. Results of the theoretical analyses using empirical Freundlich adsorption isotherm had shown that the multistage process is more efficient than the single-stage process in terms of phenol removal per unit amount of organobentonite. It was also shown that equal division of the total amount of organobentonite among all adsorption stages yields the best overall phenol removal efficiency for the multistage process in comparison with other organobentonite allocations. The Freundlich adsorption isotherm was confirmed by experimental tests to describe well the phenol adsorption by organobentonites. Results of theoretical performance analyses of the multistage adsorption process were experimentally verified using two- and three-stage test examples. PMID- 12118614 TI - Production of S. meliloti using wastewater sludge as a raw material: effect of nutrient addition and pH control. AB - The utilization of wastewater sludge as a raw material for the production of legume inoculant is considered as a new viable alternative for recycling. The effect of addition of nutrient sources (yeast extract and glycerol) and the pH control on S. meliloti was investigated in a shake flask and at controlled pH in a 15 l fermentor. Different concentrations of yeast extract (0.5, 1, 2 and 4 g l( 1)) and glycerol (2.5, 5, 7.5 and 10 g l(-1)) were added to the secondary sludge. The cell yield as well as the generation time were affected by the addition of these nutrient sources. The maximum cell yield (8.85x10(9) cfu ml(-1)) was achieved in 32 hours of incubation with the addition of 4 g l(-1) of yeast extract. This value was 3.2 times higher than from the non-supplemented sludge. Moreover, at this yeast extract concentration, the cell concentration in the stationary phase did not decrease. The addition of glycerol to sludge samples containing 4 g l(-1) of yeast extract further improved the rhizobial growth but not significantly compared with the control. The highest yield (16.5x10(9) cfu ml(-1)) was obtained with 7.5 g l(-1) of glycerol and 4 g l(-1) of yeast extract. In fermentor experiments, pH did not seem to be a limiting factor and the increase of pH up to 8.85 in uncontrolled fermentor seems to have no effect on rhizobial growth and cell yield. PMID- 12118616 TI - Reactor configuration--Part II. Comparative process stability and efficiency of thermophilic anaerobic digestion. AB - Comparative process stability and efficiency of thermophilic anaerobic digestion (55 degrees C) has been evaluated for four different reactor configurations, which are: daily batch-fed single-stage continuously stirred tank reactor (CSIR) (TB), continuously-fed single-stage CSTR (TC), daily batch-fed two-phase CSTR (TTP), and daily batch-fed non-mixed single-stage reactor (TNMR). The results are discussed for periods, 1) start-up until steady state (Days 0-200) and 2) OLR increase from 4% solids at steady state to reactor failure by increasing solids concentration in feed and decreasing the HRT (Days 201-). During the start-up period, the TB showed the worst stability with a pH drop whenever the solids concentration in the feed was increased. Conversely the TNMR reached steady state with 4% feed solids in the shortest time with relatively stable pH and very low VFA. The superior performance of the TNMR confirms the importance of microbial consortia proximity especially for the removal of propionate. The cocktail of inorganic nutrients (Ca, Ni, Fe, and Co) was added daily into all reactors showing high VFA. In the case of the TNMR, complete removal of propionate occurred after supplementation of nutrients. The results indicated that adding nutrients stimulated gas production and facilitated removal of almost all VFA confirming the importance of inorganic nutrients bioavailability. During the long term operation with OLR increases until reactor failure (pH below 5.5), the results show that TB failed at the lowest OLR while the TC and the TNMR reached the highest OLR. Compared with the daily batch fed reactors, the constant pH of the MC seems to be the reason why the MC reached the highest OLR. The superior performance of and the TNMR during both the start-up long-term period confirms the importance of microbial consortia proximity. Two-phase digestion showed little benefit over single stage during the start-up period and no benefit was observed during the long-term period. Additional experiments in which the reactor configuration was changed from CSTR to non-mixed reactor showed significant benefit with respect to gas production and VFA threshold concentration. This once again manifests the importance of microbial consortia proximity. PMID- 12118615 TI - Reactor configuration--Part I. Comparative process stability and efficiency of mesophilic anaerobic digestion. AB - The comparative process stability and efficiency of mesophilic anaerobic digestion (35 degrees C) has been evaluated for four different reactor configurations, which are: daily batch-fed single-stage continuously stirred tank reactor (CSTR) (MB), continuously fed single-stage CSTR (MC), daily batch-fed two phase CSTR (MTP), and daily batch-fed non-mixed single-stage reactor (MNMR). The results are discussed for 2 periods, 1) start-up until steady state (HRT = 20 days, Days 0-200) and 2) OLR increase from 4% solids at steady state to reactor failure by increasing solids concentration in the feed and decreasing the HRT (Days 201-). During the start-up period, the MB showed the lowest stability with a pH drop whenever the solids concentration in the feed was increased. Conversely the MNMR reached steady state with 4% feed solids inthe shortest time with a relatively stable pH and low VFA. The superior performance of the MNMR confirms the importance of microbial consortia proximity especially for the removal of propionate. A cocktail of nutrients (Ca, Ni, Fe, and Co) was added daily into the reactors which showed high VFA except the MNMR which did not exhibit the elevated VFA. The results indicated that adding nutrients stimulated gas production and facilitated removal of almost all VFA except propionate confirming the importance of inorganic nutrients bioavailability. During the long-term operation with OLR increase until reactor failure (pH below 55), the results show that the MC reached the highest OLR while the MNMR failed first. Compared with the daily batch fed reactors, the constant pH of the MC seems to be the reason why the MC reached the highest OLR. Considering the best performance of the MNMR during the start-up period, the failure of the MNMR first is hypothesized to be due to the lack of trace nutrients at high OLR. Overall, two-phase digestion showed no benefit over single stage and all reactors demonstrated comparable VS removal percentage. Additional experiments in which the reactor configuration was changed from CSTR non-mixed reactor showed significant benefit with respect to gas production and lowered propionate threshold concentration. This once again manifests the importance of microbial consortia proximity with respect to the degradation of propionate. PMID- 12118617 TI - Biodegradability enhancement by wet oxidation in alkaline media: delignification as a case study. AB - Nowadays many industries are considering the recycling of process waters as a way of improving environmental safety, preventing pollution, and avoiding the loss of valuable production materials. One industry in the forefront of this trend is the pulp and paper industry. Lignin is a pollutant present in the mill process waters and such macromolecules can cause problems during biological treatment of process waters. Wet oxidation (WO) is a process that can be used as a pre-treatment method for lignin fragmentation and improvement of biodegradability. Wet oxidation (WO) under alkaline conditions permits faster lignin fragmentation than the conventional WO process and, therefore, should favour biodegradability improvement. In this study, the experiments were carried out in a high-pressure batch reactor with an alkali lignin solution at temperatures up to 438 K, an alkali concentration of 1.5-3.5 g l(-1) and an oxygen partial pressure of 0.4 to 1.5 MPa. At an alkali concentration of 3.5 g l(-1)1 and 0.4 MPa of oxygen partial pressure, an increase in BOD/COD ratio was achieved from an initial 11% to 71%. The experiments also showed that the amount of small molecules in the solution measured by Immediately Available BOD (IA BOD) depends on the amount of alkali added and the operating temperature. PMID- 12118618 TI - Detention storage volume for combined sewer overflow into a river. AB - This article discusses the storage volume needed in a combined sewer system tank in order to preserve the water quality. There are a lot of design criteria which do not take into account the conditions of the receiving water, and as a result are inappropriate. A model was used to simulate the performance of a theoretical combined sewer system where a tank was located downstream. Results were obtained from the overflows produced by the rain recorded in Santander (Spain) for 11 years, with several combinations of storage volume and treatment capacity in the wastewater treatment plant. Quality criteria were also proposed for faecal coliforms, BOD, and total nitrogen to evaluate the effects from the overflows in the river water quality. Equations have been obtained which relate the number of overflows, the storage volume and the treatment plant capacity. The bacteriological pollution, quantified by means of faecal coliforms, was the analytical parameter which produced the most adverse effects in the river, so that more storage volume is needed (45 to 180 m3 ha(-1) net) than with other simulated pollutants (5 to 50 m3 ha(-1) net for BOD, and less than 4 m3 ha(-1) net for the total nitrogen). The increase in the treatment plant's capacity, from two to three times the flow in dry weather, reduces the impact on the river water in a more effective way, allowing a reduction of up to 65% in the number of overflows rather than increasing the storage volume. PMID- 12118619 TI - Phosphorus removal and greenhouse gas N2O emission in a lime-induced aerobic sludge granule process. AB - Aerobic sludge granulation was achieved in an activated sludge process continuously fed with lime (Ca2+ 100 mg l(-1) influent) every other day. Eighteen days after lime addition, activated sludge granules with the size of 0.5-2.2 mm were formed, which occupied 10-25% of total sludge volume. Sludge volume index (SVI) was reduced to an average of 50 ml g(-1), which increased average sludge concentration to 3.6-5.0 g VSS l(-1), 1.6-2.1 times of that of control. Greenhouse gas N2O emission was also significantly reduced: N2O concentration from the lime-addition reactor was 5-15 ppmv, 47-61% of that of control, Effluent PO4-P concentration was generally lower than 1 mg l(-1) when average influent PO4 P concentration was 6.07-6.37 mg l(-1). Total phosphorus (TP) and total nitrogen (TN) removal efficiencies were around 89.6% and 14.5-16.1%, over 3.5 and 1 times higher than those of control, respectively. COD removal rate in the lime-addition reactor was 2.05-2.48 kg COD m(-3) d(-1), higher than 1.34-1.61 kg COD m(-3) d( 1) in the control. PMID- 12118620 TI - Treatment of antibiotics wastewater utilizing successive hydrolysis, denitrification and nitrification. AB - A process consisting of anaerobic hydrolysis, denitrification, and oxidation/nitrification was proposed for simultaneous removal of carbon and nitrogen from terramycin crystallizing mother solution (TCMS), and its performance was investigated by treating diluted TCMS in a lab-scale continuous flow column system. Direct denitrification-nitrification of diluted TCMS produced a significant residual of nitrate and nitrite, which disappeared 44 days after startup of anaerobic hydrolysis column. The electron donors available to denitrification were increased by 6 times after diluted TCMS was treated in the hydrolysis column under an HRT of 2.5 h or longer. The reaction rates of organics decomposition, nitrification, and denitrification were also significantly increased due to reduction of terramycin and decomposition of complicated molecules to small molecules during anaerobic hydrolysis. The specific denitrification rate and nitrification rate increased from 0.033 d(-1) and 0.01 d(-1) to 0.045 d(-1) and 0.021 d(-1) respectively after diluted TCMS (dilution ratio: 1:4) was hydrolyzed in anaerobic hydrolysis column at a hydraulic retention time (HRT) of 4 h. PMID- 12118621 TI - Effects of different additives on the performance of spray dryer system during incineration process. AB - The spray dryer system was conventionally employed to remove the SOx, NOx, and HCl in the flue gas. However, the removal efficiency of acid gas in the practical incineration flue gas, which contains dust, heavy metals, and acid gas itself, was seldom mentioned in the literature. The alkaline sorbents possess large specific surface that was a main factor on the adsorption of heavy metals and acid gas. Therefore, the primary objective of this study was focused on the effect of different additives on the removal efficiency of acid gas and heavy metals (Cr, Cd and Pb). The mass and element size distribution of heavy metals in fly ash under different additives were also investigated. The results indicated that the removal efficiency of HCl in the spray dryer system was higher than 97.8%. The effects of additives on the removal efficiency of HCl, however, were undistinguished. In the desulfurization process, the highest removal efficiency was 71.3% when the additive of amorphous SiO2 was added in the spray dryer system. The removal efficiency was 66.0% with the additive of CaCl2 and 63.1% without any additives, respectively. It was also found that the spray dryer system could decrease the concentration of metal in fly ash but increase the amount of fly ash. In addition, amorphous SiO2 in the alkaline sorbent tended to increase the adsorption of heavy metal on reactant, because it could enhance the dispersion of alkaline sorbent. PMID- 12118622 TI - Subsurface irrigation as a microbial delivery tool for bioaugmentation: transport, distribution and survival in large packed soil columns. AB - Traditionally, soil surface inoculation and surface irrigation are used for delivery, transport and distribution of bacteria for agricultural and in-situ environmental decontamination applications. The objective of this study was to test whether subsurface irrigation with a water table management (WTM) system, successfully used previously for nutrient delivery, could also be used to deliver bacteria to soil depths. Twelve stainless steel columns, 1000 mm in length and 200 mm in diameter, were packed with a sandy loam soil. Four experimental treatments, two subsurface irrigation and two surface irrigation, were randomly allocated, in triplicate, to the twelve columns. The transport, implantation and survival of Sinorhizobium (Rhizobium) meliloti, strain A-025, in these columns were monitored at different depths and times after surface and subsurface irrigation. Even though the transport of bacterial cells was slower with the subsurface irrigation regime than with surface irrigation, more bacteria were implanted at different depths with subsurface irrigation. The numbers of bacterial cells distributed with subsurface irrigation were 1.6x10(5), 1.3x10(5), 2.6x10(5) and 2.9x10(5) cfu (colony forming units) x g(-1) of soil at 60, 300, 500 and 700 mm depths, respectively, whereas with surface irrigation the numbers were 2.0x10(4), 3.0x10(4), 1.9x10(5), 9.0x10(2) cfu x g(-1) of soil. These results dearly indicate that subirrigation can be used effectively to bioaugment a sandy loam soil matrix. PMID- 12118623 TI - Drug treatment outcome methodology (1993-1997) strengths, weaknesses, and a comparison to the alcohol field. AB - Although several critiques of the methodology of alcohol treatment outcome studies have been published, similar reviews of the methodology of drug treatment outcome studies are lacking. This paper reviews the methodology of drug treatment outcome studies published from 1993 through 1997 and draws comparisons with the most recent methodological review of alcohol treatment outcome studies. Each drug study was evaluated as to whether the following types of data were reported: (1) demographic, (2) drug use, (3) study characteristics, and (4) outcome and follow up information. Although results for drug studies showed some areas of strength compared to alcohol studies, in general, the weaknesses were similar to or worse than in the alcohol field, including inadequate reporting of demographic and drug use variables. Weaknesses in follow-up procedures were particularly notable. Suggestions for improving the reporting of methodological and outcome variables for drug treatment outcome studies are discussed. PMID- 12118624 TI - Anxiety and explicit alcohol-related memory in problem drinkers. AB - Anxiety is associated with increased craving following in vivo cue exposure in alcoholics. Theoretical accounts [Psychol. Rev. 97 (1990) 147.] have proposed that conscious, deliberate cognitive processes underlie increased craving in drinkers who are trying to abstain. The present study tested the hypothesis that anxiety is associated with biases in explicit (i.e., conscious, deliberate) memory that promote recall of alcohol-related concepts in response to negative affective cues. Fifty-two (seven females) outpatient problem drinkers performed a cued recall task that assessed memory for alcohol-related (ALC), negative affective (NEG), and neutral (NEU) target concepts that had been paired with NEG, ALC, and NEU cues during an incidental study phase. Higher anxiety was associated with increased recall of ALC targets paired with NEG cues. State and trait anxiety were intercorrelated, with higher levels coinciding with a higher frequency of drinking in negative mood states. These findings demonstrate a correspondence between anxiety and alcohol-related memory, and suggest that explicit memory biases may contribute to increased subjective responses (e.g., craving, expectancies) to alcohol stimuli in anxious problem drinkers. PMID- 12118625 TI - Social behaviour and network therapy basic principles and early experiences. AB - The present paper reports on the basic principles of a treatment approach currently being used in a National Multicentre Randomised Controlled Trial of Alcohol Treatments in the United Kingdom (UK Alcohol Treatment Trial). The treatment: Social Behaviour and Network Therapy (SBNT) is novel as a package but has been developed by integrating a number of strategies found to be effective in other treatment approaches. The intervention is based on the notion that to give the best chance of a good outcome people with serious drinking problems need to develop positive social network support for change. A brief review of the evidence supporting social treatments for alcohol problems is followed by an outline of the feasibility work and the basic principles that guided the development of SBNT. Process data from the first 33 trial cases and 2 case vignettes are described and discussed. It is concluded that SBNT is a feasible and coherent treatment approach that can be delivered by a range of therapists in the alcohol field. PMID- 12118626 TI - Motivational interviewing in a group setting with mandated clients: a pilot study. AB - Some clients who are court-ordered to undergo substance abuse treatment are not able or willing to identify treatment-related goals. This situation can cause difficulties in the therapeutic relationship and compromise treatment. To address this problem, we offered a group-based motivational enhancement program prior to standard treatment. Over 2 years, 73 clients attended the motivation group while 94 did not. Compared to those who did not, those who attended the motivation group were significantly less likely to meet criteria for substance abuse dependence, they attended a higher proportion of their treatment sessions, and they were more likely to have completed treatment. After controlling for diagnosis, employment, and age, attending the motivation group was still related to higher rates of attendance and treatment completion. A motivation group may promote better participation in substance abuse treatment programs among persons who do not identify treatment goals. Additional research is needed to draw firm conclusions. PMID- 12118627 TI - College student heavy drinking in social contexts versus alone. AB - Heavy drinking is common among college students and typically occurs in social contexts. Heavy drinking when alone, however, is less common. The present study hypothesized that students who drink heavily when alone (HD-Alone) would differ from college students who only drink heavily in social contexts (Social HD). Forty-nine HD-Alone students (at least one heavy-drinking episode when alone), 213 Social HDs, and 63 non-heavy drinkers (Non-HDs) were compared on alcohol related consequences, drinking milestones, alcohol-outcome expectancies, and symptoms of depression. HD-Alone students reported more negative drinking consequences, earlier onset of regular drinking, more alcohol expectancies, less self-efficacy and motivation to reduce drinking, and higher depression scores than Social HDs and Non-HDs. Findings imply individual differences among heavy drinking college students according to their drinking context. PMID- 12118628 TI - A structural equation model of the effect of poverty and unemployment on alcohol abuse. AB - The short- and long-term effects of poverty and unemployment on alcohol abuse are investigated using structural equation modelling (SEM) to better understand the observed conflicting relationships among them. We studied 795 community residents who provided complete data in both 1989 and 1991 in the Winnipeg Health and Drinking Survey (WHDS), with equal representation of males and females. Results indicate that (a) increased poverty causes increased alcohol use and alcohol problems, and (b) recent unemployment decreases alcohol use while longer unemployment increases it. It is concluded that the effect of unemployment on alcohol abuse changes direction with time and, thus, both cross-sectional and longitudinal data are required to assess any meaningful relationship between them. PMID- 12118629 TI - Depressive symptoms at the age of 12 years and future heavy alcohol use. AB - OBJECTIVE: To study the relationships between depressive and psychosomatic symptoms at the age of 12 years and heavy use of alcohol 3 years later, and to determine whether certain depressive symptoms are more predictive than others in regard to later heavy alcohol use. METHODS: The Children's Depression Inventory (CDI) with added questions concerning psychosomatic symptoms and bullying was used to reveal psychiatric symptoms among children at the age of 12 years. Information concerning alcohol use was obtained from the children at the age of 15 years. RESULTS: Disordered mood at the age of 12 years was not related to heavy alcohol use at the age of 15 years. Perceiving oneself as failing to perform well at school and low self-esteem at the age of 12 years were related to heavy use of alcohol 3 years later among girls, as were interpersonal problems with aggressive tendencies among boys. CONCLUSIONS: Further studies are urgently needed to determine the relationships between the expectations of the school system and deviant behaviors such as substance use among dysphoric children. PMID- 12118630 TI - Abstinence trajectories among treated crack cocaine users. AB - This article reports findings from a study that investigated treatment outcomes among crack/cocaine users over a 18-month period. From a cohort of 229 subjects, three groups emerged: (1) those who had reported ongoing, stable abstinence from crack/cocaine; (2) those who had consistently used during the period; and (3) those who reported cycling between abstinence and use during the follow-up period. Analyses of variance (ANOVA) were conducted to compare the three groups in terms of intake characteristics, including demographic profile, previous treatment, motivational factors, and functioning in seven Addiction Severity Index (ASI) domains. Length of time involved in aftercare and Twelve Step participation after treatment were also contrasted among the three groups. Results showed that subjects who achieved sustained abstinence from crack/cocaine also did better in other domains such as employment, family, legal, and psychiatric than others. Stable abstinence was also significantly associated with a longer period of aftercare and frequent attendance at Twelve Step programs. Logistic regression analyses further estimated the significant impact of the posttreatment factors on the achievement of sustained abstinence. The implications of these findings for treatment services research are discussed. PMID- 12118631 TI - Expectations of the effects of drinking on couple relationship functioning: an assessment of women in distressed relationships who consume alcohol at harmful levels. AB - Based on a cognitive-social learning model of alcohol use, it was hypothesised that women with both alcohol and relationship problems would endorse more positive expectations of the effects of alcohol consumption on their relationship and would report lower relational efficacy than women without relationship or alcohol problems. Measures of relationship-referent alcohol expectancies and relational efficacy were completed by 174 married women with both alcohol and relationship problems (n = 20), alcohol problems alone (n = 26), relationship problems alone (n = 30), or neither problem (n = 98). Women without either alcohol or relationship problems strongly rejected expectations of enhanced relationship functioning (e.g., enhanced intimacy, increased emotional expression) following alcohol consumption, whereas women with both alcohol and relationship problems were ambivalent about these positive expectations. Women with both problems also reported lower relational efficacy than the other groups of women. Negative expectations about the effect of alcohol consumption on relationships in women with low relational efficacy may inhibit harmful drinking. PMID- 12118632 TI - Pathological gambling in treatment-seeking substance abusers. AB - Accumulating evidence suggests that treatment-seeking substance abusers have high rates of gambling problems. However, relatively little is known about the relation between gambling problems and specific psychoactive substances apart from alcohol and methadone-treated opiate addicts. In this study of 580 individuals admitted to a residential addictions program, 10.5% were found to score in the pathological gambling range on the South Oaks Gambling Screen (SOGS) within the past year. The rate of pathological gambling was much higher for cannabis abusers (24%) than for alcohol (4%), cocaine (11.5%), and opiate abusers (4.8%). Men also reported higher rates of pathological gambling (11.9%) than women (7.5%). Individuals with a pathological gambling problem tended to report family histories of gambling problems as well. PMID- 12118633 TI - I. The unethics of 'request' caesarean section. PMID- 12118634 TI - II. The unfacts of 'request' caesarean section. PMID- 12118635 TI - Universal prophylaxis compared with screen-and-treat for Chlamydia trachomatis prior to termination of pregnancy. PMID- 12118636 TI - Adhesion molecules and the normal endometrium. PMID- 12118637 TI - Few women wish to be delivered by caesarean section. AB - OBJECTIVE: To investigate how many women wish to have a caesarean section when asked in early pregnancy, and to identify background variables associated with such a wish. DESIGN: National survey. SETTING: Swedish antenatal clinics. POPULATION: 3,283 Swedish-speaking women booked for antenatal care, at approximately 600 Swedish antenatal clinics, during three weeks spread over one year (1999-2000). METHODS: A questionnaire was mailed shortly after the first antenatal visit. MAIN OUTCOME MEASURES: Women's preferences for mode of delivery. RESULTS: 3,061 women completed the first questionnaire, corresponding to 94% of those who consented to participate after exclusion of reported miscarriages. The background characteristics of the study sample were very similar to a one-year cohort of women giving birth in Sweden during 1999. The result showed that 8.2% of the women would prefer to have a caesarean section. A wish for caesarean section was associated with parity, age, civil status, residential area and obstetric history. Women preferring caesarean section were more depressed and worried, not only about giving birth, but also about other things in life. A multivariate logistic regression model showed three factors being statistically associated with a wish for caesarean section: a previous caesarean section, fear of giving birth and a previous negative birth experience. CONCLUSIONS: Relatively few women wish to have a caesarean section when asked in early pregnancy, and these women seem to be a vulnerable group. PMID- 12118638 TI - What are the implications for the next delivery in primigravidae who have an elective caesarean section for breech presentation? AB - OBJECTIVE: To determine the mode of the next delivery for primigravid women who have an elective caesarean section for breech presentation. DESIGN: Retrospective cohort study. SETTING: University teaching hospital. POPULATION: All primigravid women who had an elective caesarean section for a singleton pregnancy in the years 1992 to 1997 and who delivered their next baby in the hospital before 1999. METHODS: Review of hospital computerised records. MAIN OUTCOME MEASURES: Mode of delivery and fetal presentation at next delivery. RESULTS: Of 194 women who had an elective caesarean section with a breech presentation as a primigravida, 19 (9.8%) had a breech presentation at the time of elective caesarean section for their next baby compared with only two (1.7%) in the 121 women who had an elective caesarean section with a cephalic presentation as a primigravida (RR 5.9 [95% CI 1.4-25.0]). Despite the increased likelihood of another breech presentation, the overall repeat section rate was 43.8% (n = 85) in women with a previous breech presentation (n = 194), compared with 61.2% (n = 74) in women with a previous cephalic presentation (n = 121) (RR 0.72 [95% CI 0.58-0.89]). Of those women allowed to labour after elective caesarean section as a primigravid, the vaginal birth rate was 109/130 (84%) if the presentation previously was breech compared with 47/69 (68%) if the presentation previously was cephalic (RR 1.2 [95% CI 1.03-1.50]). CONCLUSIONS: Women who have an elective caesarean section for a breech presentation in their first pregnancy have about a 1 in 10 chance of having an elective caesarean section for a breech presentation in their second pregnancy. Overall, the incidence of repeat caesarean section for their second baby was 43.8%, and of those allowed to labour, 84% achieved a vaginal delivery. These results compared favourably with women who had an elective caesarean section with a cephalic presentation in their first pregnancy. This information is important in advising primigravid women with a breech presentation about longer term implications of elective caesarean section. It also allows healthcare managers to anticipate the resource implications of any changes in clinical practice for women with a breech presentation in their first pregnancy. PMID- 12118639 TI - High incidence of obstetric interventions after successful external cephalic version. AB - OBJECTIVE: To investigate the delivery outcome after successful external cephalic version (ECV). DESIGN: Case-control study. SETTING: University teaching hospital. POPULATION: The study group consisted of 279 consecutive singleton deliveries at term over a six-year period, all of which had had successful ECV performed. The control group included 28,447 singleton term deliveries during the same six-year period. METHODS: Between group differences were compared with the Mann-Whitney U test or Student's t test where appropriate. Odds ratio and 95% confidence interval (CI) were calculated for categorical variables. Main outcome measures Incidence of and indications for obstetric interventions. RESULTS: The risk of instrumental delivery and emergency caesarean section was higher in the ECV group (14.3% vs 12.8%; OR 1.4; 95% CI 1.0-2.0, and 23.3% vs 9.4%; OR 3.1; 95% CI 2.3 4.1, respectively). The higher caesarean rate was due to an increase in all major indications, namely, suspected fetal distress, failure to progress in labour and failed induction. The higher incidence of instrumental delivery was mainly due to an increase in prolonged second stage. The odds ratio for operative delivery remained significant after controlling for potential confounding variables. There were also significantly greater frequencies of labour induction (24.0% vs 13.4%; OR 2.0; 95% CI 1.5-2.7) and use of epidural analgesia (20.4% vs 12.4%; OR 1.8; 95% CI 1.4-2.4) by women in the ECV group. The higher induction rate is mainly due to induction for post term, abnormal cardiotocography (CTG) and antepartum haemorrhage (APH) of unknown origin. CONCLUSION: The incidence of operative delivery and other obstetric interventions are higher in pregnancies after successful ECV. Women undergoing ECV should be informed about this higher risk of interventions. PMID- 12118640 TI - Does sweeping of membranes beyond 40 weeks reduce the need for formal induction of labour? AB - OBJECTIVE: To assess the efficacy of sweeping of membranes beyond 40 weeks of gestation in reducing the incidence of induction of labour, when induction was planned at 42 weeks. DESIGN: Prospective randomised controlled trial. SETTING: A regional obstetric unit in Hong Kong. POPULATION AND METHODS: A total of 120 women with certain gestational age, determined by early pregnancy ultrasound scan, were recruited from 1st July, 1998 to 31st December, 1999. Sixty women were randomly allocated to sweeping of membranes and the other 60 women acted as control. The satisfaction for women allocated to sweeping of membranes was assessed by a questionnaire after the procedure. The two groups were assessed on intention-to-treat basis. MAIN OUTCOME MEASURES: The incidence of formal induction of labour was compared between the two groups. Possible complications of sweeping of membranes such as rupture of membranes, intrapartum infection, postpartum infection, and neonatal infection were also assessed. Maternal and perinatal outcomes were also assessed. RESULTS: The recruitment to delivery interval was significantly shorter among women who had sweeping of membranes (3.2 versus 4.2 days, P < 0.05). The incidence of induction of labour was comparable (35.5% versus 38%, RR 0.91, 95% CI 0.57 - 1.46). The incidences of caesarean section and assisted vaginal delivery were comparable. The incidences of premature rupture of membranes, intrapartum, and postpartum infection were comparable. The perinatal outcomes were also comparable between the two groups. Up to 70% of women found that this procedure was associated with significant discomfort. One third of these women complained of significant pain. CONCLUSIONS: Sweeping of membranes beyond 40 weeks does not reduce the need for formal induction of labour at 42 weeks. Although it is safe, the majority of women felt uncomfortable during the procedure. PMID- 12118641 TI - Acupuncture treatment during labour--a randomised controlled trial. AB - OBJECTIVE: To investigate acupuncture treatment during labour with regard to pain intensity, degree of relaxation and outcome of the delivery. DESIGN: Randomised controlled trial. SETTING: Delivery ward at a tertiary care centre hospital in Sweden. POPULATION: Ninety parturients who delivered during the period April 12, 1999 and June 4, 2000. METHODS: Forty-six parturients were randomised to receive acupuncture treatment during labour as a compliment, or an alternative, to conventional analgesia. MAIN OUTCOME MEASURES: Assessments of pain intensity and degree of relaxation during labour, together with evaluation of delivery outcome. RESULTS: Acupuncture treatment during labour significantly reduced the need of epidural analgesia (12% vs 22%, relative risk [RR] 0.52, 95% confidence interval [CI] 0.30 to 0.92). Parturients who received acupuncture assessed a significantly better degree of relaxation compared with the control group (mean difference 0.93, 95% CI -1.66 to -0.20). No negative effects of acupuncture given during labour were found in relation to delivery outcome. CONCLUSIONS: The results suggest that acupuncture could be a good alternative or complement to those parturients who seek an alternative to pharmacological analgesia in childbirth. Further trials with a larger number of patients are required to clarify if the main effect of acupuncture during labour is analgesic or relaxing. PMID- 12118642 TI - Sublingual compared with oral misoprostol in term labour induction: a randomised controlled trial. AB - OBJECTIVE: To compare the efficacy and patient acceptability of 50 microg of sublingual misoprostol with 100 microg of oral misoprostol in the induction of labour at term. DESIGN: Non-blinded randomised comparative trial. SETTING: Tertiary level UK Hospital. SAMPLE: Two hundred and fifty women at term with indications for labour induction. METHODS: Fifty micrograms of sublingual misoprostol or 100 microg of oral misoprostol was administered every four hours after random allocation, to a maximum of five doses. MAIN OUTCOME MEASURES: Number of patients delivering vaginally within 24 hours of the induction, mode of delivery, neonatal outcomes and patient acceptability. RESULTS: There was no significant difference in the number of women delivering vaginally within 24 hours of the induction in the sublingual group as compared with the oral group (62.8% vs 59%, RR 1.1, 95% CI 0.6-2.1), or in the mean induction to delivery time (21.8 vs 24.1 h, mean difference 2.3 h 95% CI -2.2 to +6.7). There was no difference in the uterine hyperstimulation rates (1.6% in both groups), operative delivery rates or neonatal outcomes. In the sublingual group, 92.6% found the induction acceptable with 15.8% finding the tablets with an unpleasant taste, while in the oral group it was 96.9% and 4%, respectively. More patients in the oral group thought that they would consider the same method of induction again as compared with the sublingual group (58.6% vs 40%, RR 1.4, 95% CI 1.04-1.9). CONCLUSION: Fifty micrograms of sublingual misoprostol every four hours has the same efficacy and safety profile as compared with 100 microg orally, but the oral route might be preferred by women. PMID- 12118643 TI - The effect of body mass index on three methods of fetal weight estimation. AB - OBJECTIVE: To assess agreement between three methods of estimation of fetal weight and determine the influence of maternal obesity. DESIGN: Prospective observational study. SETTING: A tertiary referral teaching hospital. POPULATION: Unselected women attending for induction of labour. METHOD: Maternal, clinical and ultrasonic estimations of fetal weight were made prior to induction of labour in 96 consenting women. The estimations of fetal weight were performed independently by the three methods. MAIN OUTCOME MEASURE: Bland and Altman plots to show limits of agreement, and intraclass correlation coefficients. RESULTS: Variable levels of accuracy were obtained for maternal, clinical and ultrasound estimates of fetal weight. Ultrasound estimation of fetal weight performed equally best for women of high and low body mass index (BMI). For women of low BMI, the intraclass correlation coefficient (ICC) was 0.90 (95% CI 0.83-0.94) and 0.87 (95% CI 0.77-0.93) for women with high BMI. Despite this, the limits of agreement for ultrasound were in the order of -700 to +500 g. Both maternal and clinical estimation of fetal weight under-estimated true fetal weight in women with low BMI and over-estimated the true fetal weight in women with high BMI. The largest observed mean difference was obtained with clinical palpation in both low and high BMI women. CONCLUSION: The accuracy of ultrasound estimation of fetal weight was better than maternal and clinical estimation of fetal weight and was not influenced significantly by maternal BMI. PMID- 12118644 TI - Relationship between customised birthweight centiles and neonatal anthropometric features of growth restriction. AB - OBJECTIVE: To determine the relationship between customised birthweight centiles (adjusted for maternal and fetal physiological variables) and neonatal anthropometric features of intrauterine growth restriction (IUGR). DESIGN: Observational study. POPULATION: Two-hundred and seventy women with low risk pregnancies participating in a cohort study of serial ultrasound biometry. METHODS: Customised birthweight centiles were calculated following adjustment for maternal weight, height and ethnic origin, gestational age at delivery, birth order, and sex of the infant. Three separate neonatal anthropometric measures were used to define IUGR: subscapular or triceps skinfold thickness <10th centile; ponderal index <25th centile; and mid-arm circumference to occipito frontal circumference ratio (MAC/OFC) <- 1 standard deviation (SD). Relationship of the centiles to these outcomes was evaluated using likelihood ratios (LR) and kappa statistic. These approaches allowed us to examine the strength of the association: an LR of 5-10 would be expected to generate moderate changes in the pre-test probability of IUGR, whereas a kappa value of 0.2-0.4 would reflect fair agreement between customised birthweight centiles and neonatal anthropometric measures. RESULTS: Customised birthweight centile of 10 or less had the following LR values for the various anthropometric criteria for IUGR: 5.1 (95% CI 3-8.5) for low skinfold thickness; 4.3 (95% CI 2.5-7.1) for low ponderal index; and 3.9 (95% CI 2-6.6) for low MAC/OFC ratio. The kappa values were: 0.4 (95% CI 0.26 0.51) for low skinfold thickness; 0.33 (95% CI 0.21-0.46) for low ponderal index; and 0.13 (95% CI 0-0.26) for low MAC/OFC ratio. CONCLUSION: In a low risk population, customised birthweight centiles can only be moderately useful in the identification of neonates with low skinfold thickness and low ponderal index. PMID- 12118645 TI - Maternal morbidity and pregnancy outcome in a cohort of mothers transferred out of perinatal centres during a national census. AB - OBJECTIVE: To record the maternal morbidity and pregnancy outcome in this cohort. DESIGN: Retrospective data collection from a prospectively defined cohort. SETTING: The 37 largest perinatal centres in the UK. POPULATION: 258 in utero transfers recorded during a three-month census (1/4/99-30/6/99). METHODS: A questionnaire regarding the outcome of each mother was sent to the perinatal centre and receiving hospital. RESULTS: Data were returned on 242/258 (94%) mothers. Fifty-eight percent were transferred out of their perinatal centre in preterm labour and 38% had coexisting disease necessitating early delivery. The median gestational age at transfer was 32 weeks (range 23-41). Sixty-one percent delivered at the receiving hospital; 12% were transferred on to a third hospital and 29% ultimately returned to deliver at the original perinatal centre. Fifty two percent of mothers received postnatal care in hospitals other than those defined as a major perinatal centre. One mother delivered during transfer and a further nine within one hour of arrival. One mother received intensive care after delivery and later died, a further 7% required high dependency care postnatally. Data were available on 273/333 (82%) babies. The median gestational age at delivery was 34 weeks (range 24-41). Six infants were stillborn and 187/264 (71%) infants were admitted to a neonatal unit. CONCLUSIONS: This study has documented the maternal morbidity, potential risks and pregnancy outcome of a cohort of mothers transferred out of the largest perinatal centres in the UK because of a shortage of neonatal cots. A national standard for the delivery of high risk perinatal services is needed to uphold good clinical practice guidelines in the care of high risk mothers and their infants. PMID- 12118646 TI - Combined ultrasound and biochemical screening for Down's syndrome in the first trimester: a Scottish multicentre study. AB - OBJECTIVE: To evaluate the use of ultrasound measurements of fetal nuchal translucency (NT) obtained in a routine antenatal clinic setting in combination with appropriate biochemical markers as a first trimester screening test for Down's Syndrome. DESIGN: Multicentre observational study. SETTING: Fifteen Scottish maternity units. POPULATION: Pregnant women (n = 17,229) attending routine antenatal clinics at 10-14 weeks of gestation. METHODS: NT measurements were attempted in all women along with the measurement of maternal serum free beta human chorionic gonadotrophin (F beta hCG) and pregnancy-associated plasma protein-A (PAPP-A). All results were converted to multiples of the appropriate gestational median (MoM) and using a statistical model the risk of an affected pregnancy was derived. No results were given to participating women but all were offered routine second trimester biochemical screening. All cases of Down's Syndrome within the study group were ascertained and the detection rate for each marker was estimated. MAIN OUTCOME MEASURES: Success rate of obtaining NT measurements and overall effectiveness of ultrasound and biochemical markers individually and in combination for the detection of Down's Syndrome pregnancies. RESULTS: NT measurements were obtained in 72.9% of women and blood samples in 98.4%. Forty-five cases of Down's Syndrome were ascertained (2.6/1,000). NT measurements were obtained in 37 cases (median NT 1.65 MoM), blood samples in 42 cases and both NT and blood in 34 cases. In combination with the a priori maternal age risk, observed detection rates at a 5% false positive rate were 20/37 (54%) for NT, 23/42 (55%) for F beta hCG and PAPP-A and 28/34 (82%) for a combination of NT, F beta hCG and PAPP-A using a cutoff risk of 1:250. The effect of failing to obtain NT measurements in all cases reduces the overall detection rate to 62% (i.e. 28/45) if the entire series of affected pregnancies within the study group is considered. CONCLUSIONS: NT in combination with appropriate serum markers has the potential to detect over 80% of Down's Syndrome fetuses in early pregnancy. However, NT measurement is highly operator-dependent. It requires training, external quality control and adequate time to allow accurate measurement, otherwise suboptimal performance will result. PMID- 12118647 TI - Are some perinatal deaths in immigrant groups linked to suboptimal perinatal care services? AB - OBJECTIVE: To test the hypothesis that suboptimal factors in perinatal care services resulting in perinatal deaths were more common among immigrant mothers from the Horn of Africa, when compared with Swedish mothers. DESIGN: A perinatal audit, comparing cases of perinatal deaths among children of African immigrants residing in Sweden, with a stratified sample of cases among native Swedish women. POPULATION AND SETTING: Sixty-three cases of perinatal deaths among immigrant east African women delivered in Swedish hospitals in 1990-1996, and 126 cases of perinatal deaths among native Swedish women. Time of death and type of hospital were stratified. MAIN OUTCOME MEASURES: Suboptimal factors in perinatal care services, categorised as maternal, medical care and communication. RESULTS: The rate of suboptimal factors likely to result in potentially avoidable perinatal death was significantly higher among African immigrants. In the group of antenatal deaths, the odds ratio (OR) was 6.2 (95% CI 1.9-20); the OR for intrapartal deaths was 13 (95% CI 1.1-166); and the OR for neonatal deaths was 18 (95% CI 3.3-100), when compared with Swedish mothers. The most common factors were delay in seeking health care, mothers refusing caesarean sections, insufficient surveillance of intrauterine growth restriction (IUGR), inadequate medication, misinterpretation of cardiotocography (CTG) and interpersonal miscommunication. CONCLUSIONS: Suboptimal factors in perinatal care likely to result in perinatal death were significantly more common among east African than native Swedish mothers, affording insight into socio-cultural differences in pregnancy strategies, but also the suboptimal performance of certain health care routines in the Swedish perinatal care system. PMID- 12118648 TI - Transient hyperthyroidism of hyperemesis gravidarum. AB - OBJECTIVE: To characterise the clinical, biochemical and thyroid antibody profile in women with transient hyperthyroidism of hyperemesis gravidarum. DESIGN: Prospective observational study. SETTING: Hospital inpatient gynaecological ward. POPULATION: Women admitted with hyperemesis gravidarum and found to have hyperthyroidism. METHODS: Fifty-three women were admitted with hyperemesis gravidarum and were found to have hyperthyroidism. Each woman was examined for clinical signs of thyroid disease and underwent investigations including urea, creatinine, electrolytes, liver function test, thyroid antibody profile and serial thyroid function test until normalisation. MAIN OUTCOME MEASURES: Gestation at which thyroid function normalised, clinical and thyroid antibody profile and pregnancy outcome (birthweight, gestation at delivery and Apgar score at 5 minutes). RESULTS: Full data were available for 44 women. Free T4 levels normalised by 15 weeks of gestation in the 39 women with transient hyperthyroidism while TSH remained suppressed until 19 weeks of gestation. None of these women were clinically hyperthyroid. Thyroid antibodies were not found in most of them. Median birthweight in the infants of mothers who experienced weight loss of > 5% of their pre-pregnancy weight was lower compared with those of women who did not (P = 0.093). Five women were diagnosed with Graves' disease based on clinical features and thyroid antibody profile. CONCLUSIONS: In transient hyperthyroidism of hyperemesis gravidarum, thyroid function normalises by the middle of the second trimester without anti-thyroid treatment. Clinically overt hyperthyroidism and thyroid antibodies are usually absent. Apart from a non significant trend towards lower birthweights in the infants of mothers who experienced significant weight loss, pregnancy outcome was generally good. Routine assessment of thyroid function is unnecessary for women with hyperemesis gravidarum in the absence of any clinical features of hyperthyroidism. PMID- 12118649 TI - Effects of vitamin A deficiency during pregnancy on maternal and child health. AB - OBJECTIVE: To examine the association between biochemical vitamin A deficiency in pregnancy and maternal and fetal health. DESIGN: A cross sectional clinical study. SETTING: Antenatal clinic of nutrition unit of Niloufer Hospital catering for a low socio-economic population, and a private nursing home (Swapna nursing home) catering for a high socio-economic population. POPULATION: 736 pregnant women in their third trimester of pregnancy belonging to low (n = 522) and high socio-economic groups (n = 214). METHODS: All the women were subjected to a detailed clinical, anthropometric and obstetric examination. Night blindness was assessed by administering the standard WHO questionnaire. Birthweight and gestational age of the infants, maternal anaemia and development of pregnancy induced hypertension in the mother were recorded. Haemoglobin and serum retinol were estimated at the time of recruitment to the study. MAIN OUTCOME MEASURES: Serum retinol levels, anaemia, pregnancy-induced hypertension, birthweight and gestational age of the infant. RESULTS: Night blindness was observed in 2.9% of the women and subclincal vitamin A deficiency (serum retinol <20 microg/dL with no clinical signs) in 27% of the women. Moderate to severe anaemia was observed in 41.2% of the women, and 15.8% of the women developed pregnancy-induced hypertension. Sixty-one (9.4%) women delivered preterm. Univariate analysis identified a significant association between serum retinol <20 microg/dL and preterm delivery (OR = 1.74, 95% CI 1.03-2.96), maternal anaemia (OR = 1.82, 95% CI 1.28-2.60) and pregnancy-induced hypertension (OR = 1.56, 95% CI 1.02-2.83). After adjusting for the confounding variables (body mass index, parity, age and socio-economic status) in a multivariate analysis, the significant associations between serum retinol <20 microg/dL and preterm delivery (P = 0.02) and anaemia (P = 0.003) persisted, while that for pregnancy-induced hypertension disappeared (P = 0.71). CONCLUSION: The study suggests that subclinical vitamin A deficiency is a problem during the third trimester of pregnancy. Serum concentration of retinol <20 microg/dL appears to indicate a deficient status, and is associated with an increased risk of preterm delivery and maternal anaemia. PMID- 12118650 TI - A randomised controlled trial of the closure or non-closure of peritoneum at caesarean section: effect on post-operative pain. AB - OBJECTIVE: To compare the analgesic requirement in the post-operative period after closure or non-closure of the peritoneum at the caesarean delivery with a standardised anaesthetic and surgical technique. DESIGN: A randomised double blind controlled trial was performed on 100 women who underwent elective caesarean delivery. MAIN OUTCOME MEASURES: Analgesic requirement assessed by morphine usage via patient controlled analgesia pump over the first 24-hour period after surgery, oral analgesia used during the first four days, postoperative pain assessed by a visual analogue scale and a verbal rating scale, and patient satisfaction assessed by verbal rating scale were the main outcome measures. RESULTS: Pain scores at 24 hours were similar in both groups (43.5 in closure and 40.5 in non-closure) but during the first 24 hours the non-closure group had used significantly less morphine than the closure group (0.64 mg/kg of body weight vs 0.82 mg/kg, P = 0.04). The patients in non-closure group had significantly higher satisfaction scores after 24 hours than the closure group. CONCLUSION: Non-closure of both visceral and parietal peritoneum at the caesarean section produces a significant reduction in the post-operative use of patient controlled analgesia pump morphine and significantly higher patient satisfaction at 24 hours post-operatively. PMID- 12118651 TI - Endothelial function is preserved in pregnant women with well-controlled type 1 diabetes. AB - OBJECTIVE: Pregnant women with diabetes mellitus have a higher incidence of adverse pregnancy outcomes. Vascular, and in particular, endothelial function may be significantly modified in diabetes resulting in impaired endothelium-dependent relaxation. This study aims to investigate endothelium-dependent relaxation in pregnant women with pre-existing type I diabetes mellitus. METHODS: Small arteries (mean luminal diameter approximately 295 microm) were isolated from biopsies of subcutaneous fat from pregnant women with pre-existing type I diabetes mellitus, non-diabetic pregnant women, and non-diabetic non-pregnant women. Endothelial and smooth muscle function were determined using wire myography, and the contributions of nitric oxide, vasodilator prostanoid and endothelial hyperpolarisation were studied using specific inhibitors. RESULTS: Arteries obtained from the diabetic pregnant women did not demonstrate any difference in either endothelial or smooth muscle function when compared with non diabetic pregnant women. The contribution of nitric oxide to endothelium dependent relaxation was approximately 20% in the pregnant women regardless of whether they were diabetic, and approximately 11% in the non-pregnant women. Endothelial hyperpolarisation appeared to contribute largely to vasorelaxation in human subcutaneous arteries, and was at least twice that of nitric oxide in pregnant women and fivefold greater in non-pregnant women. CONCLUSIONS: This study provides evidence that pregnant women with well-controlled pre-existing type 1 diabetes mellitus have both normal endothelial and smooth muscle function. Endothelium-dependent hyperpolarisation appears to play a large role in vascular relaxation in human subcutaneous resistance arteries. This study suggests that the problems associated with diabetic pregnancies are unlikely to be due to vascular dysfunction. PMID- 12118652 TI - A longitudinal study of biochemical markers of bone turnover during normal pregnancy and pregnancies complicated by pre-eclampsia. AB - OBJECTIVES: To test the hypothesis that the increased bone turnover observed in established pre-eclampsia is present earlier in pregnancy prior to the diagnosis of pre-eclampsia. DESIGN: A prospective longitudinal study. SETTING: Obstetric Unit at Chelsea and Westminster Hospital, London. POPULATION: Nine women who subsequently developed pre-eclampsia and 17 normal pregnant controls. METHODS: Maternal plasma levels of the cross-linked carboxyl terminal telopeptide of the type I collagen (ICTP), a marker of bone resorption, and of the carboxy-terminal propeptide of type I collagen (PICP), a marker of bone formation, were measured longitudinally in nine women who developed pre-eclampsia and 17 women with normal pregnancy. Serial blood samples were obtained at 16, 20, 24, 28, 32 and 36 weeks of gestation and the levels of ICTP and PICP were measured by radio-immunoassay. RESULTS: ICTP and PICP increased progressively in the normal pregnant and pre eclampsia groups, but the rate of increase was significantly greater in the latter (P = 0.00002 and 0.0008, unpaired t test, for ICTP and PICP summary measures, respectively). In the pre-eclampsia group, positive correlation were observed between ICTP levels at 36 weeks of gestation and plasma uric acid (r = 0.9, P = 0.001) and degree of proteinuria (r = 0.78, P = 0.04). No correlation was observed between the two bone markers and other variables during normal pregnant group. CONCLUSION: These data show that biochemical markers of bone turnover are greater in pregnancies complicated by pre-eclampsia compared with normal pregnancy but only when the disease is clinically evident. PMID- 12118653 TI - Rates of bacterial vaginosis in women undergoing in vitro fertilisation for different types of infertility. AB - OBJECTIVE: To assess whether the rate of bacterial vaginosis (BV) is higher in women with tubal factor infertility compared with those with other causes of infertility. DESIGN: Cross-sectional study. SETTING: Assisted conception unit of a teaching hospital in Leeds. POPULATION: Consecutive women undergoing in vitro fertilisation. METHODS: Women undergoing in vitro fertilisation (IVF) had a vaginal smear taken at the time of their egg collection. The smear was Gram stained and graded as normal, intermediate or BV. MAIN OUTCOME MEASURES: The presence of bacterial vaginosis and the causes of infertility. RESULTS: A total of 749 women were included. The vaginal smears were normal in 63.6%, intermediate in 12.1%, and BV in 24.3%. The rates of BV in women with different types of infertility were 36.4% in tubal factor, 15.6% in male factor, 33.3% in anovulation, 12.5% in endometriosis and 18.9% in unexplained infertility. After controlling for the effects of age and smoking using a multivariate logistic regression model, women with tubal infertility were significantly more likely to have BV than women with endometriosis OR 3.63 (95% CI 1.52-8.67); male factor OR 2.98 (95% CI 1.80-4.90); and unexplained infertility OR 2.20 (95% CI 1.35-3.59). The adjusted figures for the increase of BV in women with anovulation were: endometriosis OR 3.77 (95% CI 1.28-11.08); male factor OR 3.09 (95% CI 1.37 6.96); and unexplained infertility OR 2.29 (95% CI 1.02-5.12). CONCLUSIONS: Women with tubal infertility were three times more likely to have BV than women with endometriosis, male factor or unexplained infertility. These findings support the association between BV, pelvic inflammatory disease (PID) and tubal damage but do not help distinguish between cause and effect. Women with anovulation were also three times more likely to have BV than women with endometriosis or male factor infertility, supporting suggestions of hormonal influence on vaginal flora. PMID- 12118654 TI - Algorithms for combining menstrual and ultrasound estimates of gestational age: consequences for rates of preterm and postterm birth. AB - We compared rates of preterm and postterm birth according to six algorithms for gestational age (GA) estimates based on last menstrual period (LMP) and early ultrasound (EUS): LMP alone, LMP if the discrepancy between the two estimates was within 14 days and otherwise EUS (14-day rule), a 10-day rule, a seven-day rule, a three-day rule and EUS alone. In a sample of 44,623 births in a Canadian tertiary hospital, the choice of algorithms makes a substantial impact on both preterm and postterm birth rates, even when EUS was used for discrepancies over two weeks. PMID- 12118655 TI - Estimating human fetal blood volume on the basis of gestational age and fetal abdominal circumference. PMID- 12118656 TI - Triplet pregnancy in a Jehovah's witness: recombinant human erythropoietin and iron supplementation for minimising the risks of excessive blood loss. PMID- 12118657 TI - Spontaneous expulsion of submucous fibroid after preterm labour. PMID- 12118658 TI - A case of gastrointestinal stromal tumour presenting in pregnancy. PMID- 12118659 TI - Abdominal hysterectomy after insertion of tension-free vaginal tape. PMID- 12118660 TI - Use of the distomedial-proximolateral oblique radiographic view of the elbow joint for examination of the medial coronoid process in dogs. AB - OBJECTIVE: To describe and evaluate a new radiographic view of the elbow joint in dogs that would potentially enhance observation of the medial coronoid process (MCP). SAMPLE POPULATION: 20 cadaver limbs from 10 dogs and clinical examination of 100 elbow joints of 53 dogs. PROCEDURE: Twenty elbow joints from 10 cadavers were imaged by use of mediolateral, flexed mediolateral, craniocaudal, craniolateral-caudomedial oblique (Cr15L-CdMO), and distomedial-proximolateral oblique (Di35M-PrLO) radiographic views before and after placement of 3 lead pellets placed on the cranial, medial, and craniodistal aspect of the MCP. Three examiners independently reviewed these radiographs. One hundred elbow joints of 53 dogs with forelimb lameness and signs of pain elicited on palpation of the elbow joint were examined. These joints were radiographed and treated by use of arthroscopy. Three examiners independently graded the radiographs. RESULTS: The MCP was identified on all Di35M-PrLO views made during the anatomic study. The Di35M-PrLO view had the largest area under the receiving operating characteristic (ROC) curve for detection of abnormalities of the MCP. Fractured and nonfractured MCP could only be significantly differentiated on Di35M-PrLO and mediolateral views. The Di35M-PrLO view had a higher agreement between examiners than other radiographic views for detection of fractures of the MCP. CONCLUSION AND CLINICAL RELEVANCE: The Di35M-PrLO view enhances the identification of anomalies and fragmentation of the MCP in dogs, compared with other radiographic views. The Di35M-PrLO view may be of benefit for early screening of dogs potentially affected with elbow dysplasia. PMID- 12118661 TI - Evaluation of a method to experimentally induce colic in horses and the effects of acupuncture applied at the Guan-yuan-shu (similar to BL-21) acupoint. AB - OBJECTIVE: To evaluate the reliability of a method for inducing colic via small intestinal distention in horses and to examine the analgesic potential of bilateral electroacupuncture (EAP) at the Guan-yuan-shu (similar to BL21) acupoint. ANIMALS: 5 healthy adult horses, each with a gastric cannula. PROCEDURE: A polyester balloon connected to an electronic barostat was introduced into the duodenum via the gastric cannula. At 2 specified intervals (before and after commencement of EAP), the balloon was inflated to a barostat-controlled pressure that induced signs of moderate colic. Each inflation was maintained for 10 minutes. Heart and respiratory rates were continuously recorded. Frequency of various clinical signs of colic was recorded by 2 trained observers during various combinations of balloon inflation and EAP. Each horse received each of 5 treatment protocols (EAP at 20 Hz, sham EAP at 20 Hz, EAP at 80: 120 Hz dense:disperse, sham EAP at 80: 120 Hz dense:disperse, no treatment). Sham EAP was at a point located 2 cm lateral to the Guan-yuan-shu acupoint. RESULTS: Duodenal distention consistently induced a significant increase in frequency of signs of colic. None of the EAP protocols caused a significant reduction in frequency of these clinical signs during distention. CONCLUSIONS AND CLINICAL RELEVANCE: The method described is reproducible and highly controllable method for inducing colic that involved duodenal distention that should be useful in evaluating the efficacy of various analgesic strategies. Bilateral EAP at the Guan-yuan-shu acupoint was ineffective in reducing signs of discomfort induced by this method. PMID- 12118662 TI - Pharmacokinetics after intravenous and oral administration of enrofloxacin in sheep. AB - OBJECTIVE: To compare pharmacokinetics of enrofloxacin administered IV and in various oral preparations to ewes. ANIMALS: 5 mature Katahdin ewes weighing 42 to 50 kg. PROCEDURE: Ewes received 4 single-dose treatments of enrofloxacin in a nonrandomized crossover design followed by a multiple-dose oral regimen. Single dose treatments consisted of an IV bolus of enrofloxacin (5 mg/kg), an oral drench (10 mg/kg) made from crushed enrofloxacin tablets, oral administration in feed (10 mg/kg; mixture of crushed enrofloxacin tablets and grain), and another type of oral administration in feed (10 mg/kg; mixture of enrofloxacin solution and grain). The multiple-dose regimen consisted of feeding a mixture of enrofloxacin solution and grain (10 mg/kg, q 24 h, for 7 days). Plasma concentrations of enrofloxacin and ciprofloxacin were measured by use of high performance liquid chromatography. RESULTS: Harmonic mean half-life for oral administration was 14.80, 10.80, and 13.07 hours, respectively, for the oral drench, crushed tablets in grain, and enrofloxacin solution in grain. Oral bioavailability for the oral drench, crushed tablets in grain, and enrofloxacin in grain was 4789, 98.07, and 94.60%, respectively, and median maximum concentration (Cmax) was 1.61, 2.69, and 2.26 microg/ml, respectively. Median Cmax of the multiple-dose regimen was 2.99 microg/ml. CONCLUSIONS AND CLINICAL RELEVANCE: Enrofloxacin administered orally to sheep has a prolonged half-life and high oral bioavailability. Oral administration at 10 mg/kg, q 24 h, was sufficient to achieve a plasma concentration of 8 to 10 times the minimum inhibitory concentration (MIC) of any microorganism with an MIC < or = 0.29 microg/ml. PMID- 12118663 TI - Effect of withholding feed on concentration and composition of plasma very low density lipoprotein and serum nonesterified fatty acids in horses. AB - OBJECTIVE: To measure and compare the concentration and composition of very low density lipoprotein (VLDL) in plasma and selected lipids in serum of horses fed mixed grass hay ad libitum or denied feed for 36 hours. ANIMALS: 4 healthy adult mares. PROCEDURE: Mares were either fed mixed grass hay ad libitum or denied feed for 36 hours beginning at 8:00 AM. Blood samples were collected every 2 hours during the study period and analyzed for nonesterified fatty acid (NEFA), triglyceride (TG), VLDL, and glucose concentrations and composition of VLDL. RESULTS: Withholding feed significantly increased mean serum concentrations of NEFA. By 36 hours, a 16-fold increase in mean serum NEFA concentration and 2-fold increase in mean plasma VLDL concentration, compared with baseline values, were detected. Mean plasma TG concentrations significantly increased with time in feed deprived horses. Significantly lower overall mean plasma glucose concentrations were detected in feed-deprived horses. Mean percentage of protein in VLDL was significantly lower in feed-deprived horses. Plasma VLDL concentrations varied widely among horses in response to withholding feed. Plasma TG and VLDL concentrations remained unaltered in 2 horses. CONCLUSIONS AND CLINICAL RELEVANCE: Withholding feed significantly increases blood lipid concentrations in horses, but individual horses respond differently. Serum NEFA concentrations were increased in all 4 horses denied feed, indicating mobilization of tissue triglyceride stores. Variation in plasma VLDL concentration in response to withholding feed suggests that its metabolism is strongly influenced by other, as yet undetermined, factors in horses. Differences in the plasma VLDL concentrations among horses in response to withholding feed may be used as an indication of susceptibility to the hyperlipemic syndrome of Equidae. PMID- 12118664 TI - Effects of medetomidine-midazolam, midazolambutorphanol, or acepromazine butorphanol as premedicants for mask induction of anesthesia with sevoflurane in dogs. AB - OBJECTIVE: To characterize the effects of medetomidine-midazolam, midazolam butorphanol, or acepromazine-butorphanol as premedicants for mask induction of anesthesia with sevoflurane in dogs. ANIMALS: 10 healthy Beagles. PROCEDURE: The following premedicants were administered intramuscularly: medetomidine-midazolam (20 microg/kg and 0.3 mg/kg, respectively), midazolam-butorphanol (0.1 and 0.2 mg/kg, respectively), and acepromazine-butorphanol (0.05 and 0.2 mg/kg, respectively). Saline (0.9% NaCI) solution (0.1 ml/kg) was administered intramuscularly as a control. Anesthesia was induced in each dog with sevoflurane in a 100% O2 at a flow rate of 4 L/min developed by a facemask. Vaporizer settings were increased by 0.8% at 15-second intervals until the value corresponding to 4.8% sevoflurane was achieved. Time to onset and cessation of involuntary movements, loss of the palpebral reflex, negative response to tail clamp stimulation, and endotracheal intubation were recorded, and the cardiopulmonary variables were measured. RESULTS: Mask induction with sevoflurane in dogs that received each premedicant resulted in a shorter induction time and milder changes in heart rate, mean arterial blood pressure, cardiac output, and respiratory rate, compared with mask induction without premedicants. Treatment with medetomidine-midazolam resulted in a shorter and smoother induction, compared with acepromazine-butorphanol or midazolam-butorphanol treatment, whereas the cardiovascular changes were greater. Cardiopulmonary variables of dogs during induction following treatment with acepromazine-butorphanol or midazolam-butorphanol were maintained close to the anesthetic maintenance values for sevoflurane, with the exception of mild hypotension that was observed in dogs following acepromazine-butorphanol treatment. CONCLUSION AND CLINICAL RELEVANCE: In dogs use of premedicants provides a smoother and better quality mask induction with sevoflurane. PMID- 12118665 TI - Quantitative genetics of traits associated with hip dysplasia in a canine pedigree constructed by mating dysplastic Labrador Retrievers with unaffected Greyhounds. AB - OBJECTIVE: To determine the genetic influence on expression of traits associated with canine hip dysplasia. ANIMALS: 193 dogs from an experimental canine pedigree. PROCEDURE: An experimental canine pedigree was developed for linkage analysis of hip dysplasia by mating dysplastic Labrador Retrievers with nondysplastic Greyhounds. A statistical model was designed to test the effects of Labrador Retriever and Greyhound alleles on age at detection of femoral capital epiphyseal ossification, 8-month distraction index, and 8-month dorsolateral subluxation score. RESULTS: The additive effect was significant for age at detection of femoral capital epiphyseal ossification. Restricted maximum likelihood estimates (+/-SD) for this trait were 6.4+/-1.95, 10.2+/-2.0, 10.8+/ 3.1, 11.4+/-2.1, and 13.6+/-4.6 days of age for Greyhounds, Greyhound backcross dogs, F1 dogs, Labrador Retriever backcross dogs, and Labrador Retrievers, respectively. The additive effect was also significant for the distraction index. Estimates for this trait were 0.21+/-0.07, 0.29+/-0.15, 0.44+/-0.12, 0.52+/-0.18, and 0.6+/-0.17 for the same groups, respectively. For the dorsolateral subluxation score, additive and dominance effects were significant. Estimates for this trait were 73.5+/-4.1, 71.3+/-6.5, 69.1+/-6.0, 50.6+/-12.9, and 48.4+/-7.7%, respectively, for the same groups. CONCLUSIONS: In this canine pedigree, traits associated with canine hip dysplasia are heritable. Phenotypic differences exist among founder dogs of each breed and their crosses. This pedigree should be useful for identification of quantitative trait loci underlying the dysplastic phenotype. PMID- 12118666 TI - Influence of dietary calcium and phosphorus content in a fixed ratio on growth and development in Great Danes. AB - OBJECTIVE: To study the musculoskeletal development of Great Dane puppies fed various dietary concentrations of calcium (Ca) and phosphorus (P) in fixed ratio by use of dual energy x-ray absorptiometry (DEXA), determination of serum insulin like growth factor 1 and parathyroid hormone concentrations, radiography, and blood chemistry analysis results. ANIMALS: 32 purebred Great Dane puppies from 4 litters. PROCEDURE: At weaning, puppies were assigned randomly to 1 of 3 diets. Blood was collected for biochemical analyses and hormone assays, and radiography and DEXA were performed through 18 months of age. Changes in body weight, bone mineral content, fat tissue weight, lean mass, result of serum biochemical analyses, hormonal concentrations, and radius lengths were analyzed through 18 months of age. RESULTS: Bone mineral content of puppies correlated positively with Ca and P content of the diets fed. Significant differences between groups in bone mineral content, lean mass, and body fat were apparent early. The disparity among groups increased until 6 months of age and then declined until body composition was no longer different at 12 months of age. Accretion rates for skeletal mineral content, fat, and lean tissue differed from each other and by diet group. CONCLUSIONS AND CLINICAL RELEVANCE: Ca and P concentrations in the diet of young Great Dane puppies are rapidly reflected in the bone mineral content of the puppies until 5 to 6 months of age, after which hormonal regulation adjusts absorption and excretion of these minerals. Appropriate Ca and P concentrations in diets are important in young puppies < 6 months of age. PMID- 12118667 TI - Effect of background serum lithium concentrations on the accuracy of lithium dilution cardiac output determination in dogs. AB - OBJECTIVES: To assess the effect of increasing serum lithium concentrations on lithium dilution cardiac output (LiDCO) determination and to determine the ability to predict the serum lithium concentration from the cumulative lithium chloride dosage. ANIMALS: 10 dogs (7 males, 3 females). PROCEDURE: Cardiac output (CO) was determined in anesthetized dogs by measuring LiDCO and thermodilution cardiac output (TDCO). The effect of the serum lithium concentration on LiDCO was assessed by observing the agreement between TDCO and LiDCO at various serum lithium concentrations. Also, cumulative lithium chloride dosage was compared with the corresponding serum lithium concentrations. RESULTS: 44 paired observations were used. The linear regression analysis for the effect of the serum lithium concentration on the agreement between TDCO and LiDCO revealed a slope of -1.530 (95% confidence interval [CI], -2.388 to -0.671) and a y intercept of 0.011 (r2 = 0.235). The linear regression analysis for the effect of the cumulative lithium chloride dosage on the serum lithium concentration revealed a slope of 2.291 (95% CI, 2.153 to 2.429) and a y-intercept of 0.008 (r2 = 0.969). CONCLUSIONS AND CLINICAL RELEVANCE: The LiDCO measurement increased slightly as the serum lithium concentration increased. This error was not clinically relevant and was minimal at a serum lithium concentration of 0.1 mmol/L and modest at a concentration of 0.4 mmol/L. The serum lithium concentration can be reliably predicted from the cumulative lithium dosage if lithium chloride is administered often within a short period. PMID- 12118668 TI - Plasma and urine electrolyte and mineral concentrations in Thoroughbred horses with recurrent exertional rhabdomyolysis after consumption of diets varying in cation-anion balance. AB - OBJECTIVE: To determine whether plasma, urine, and fecal electrolyte and mineral concentrations differ between clinically normal horses and Thoroughbreds with recurrent exertional rhabdomyolysis (RER) after consumption of diets varying in cation-anion balance. ANIMALS: 5 Thoroughbred mares with RER and 6 clinically normal mixed-breed mares. PROCEDURE: Each of 3 isocaloric diets designated as low, medium, and high on the basis of dietary cation-anion balance (DCAB) values of 85, 190, and 380, respectively, were fed to horses for 14 days. During the last 72 hours, 3 horses with RER and 3 control horses had daily urine and fecal samples obtained by total 24-hour collection. Remaining horses had urine samples collected daily by single catheterization. RESULTS: For each diet, no differences existed between horses with RER and control horses in plasma pH, electrolyte concentrations, and creatine kinase activity or in urine pH and renal fractional excretion (FE) values. Plasma pH, strong ion difference, bicarbonate and total carbon dioxide concentrations, and base excess decreased and plasma chloride and ionized calcium concentrations increased with decreasing DCAB. Urine pH decreased with decreasing DCAB. The FE of chloride and phosphorus were greatest for horses fed the low diet. The FE values for all electrolytes exept magnesium did not differ between urine samples obtained by single catheterization and total 24-hour collection. Daily balance of calcium, phosphorus, sodium, chloride, and potassium did not differ significantly among horses fed the various diets. CONCLUSIONS: In clinically normal horses and in horses with RER, the DCAB strongly affects plasma and urine pH and the FE of sodium, potassium, chloride, and phosphorus. PMID- 12118669 TI - Effect of general anesthesia and minor surgical trauma on urine and serum measurements in horses. AB - OBJECTIVE: To characterize the effect of general anesthesia and minor surgery on renal function in horses. ANIMALS: 9 mares with a mean (+/- SE) age and body weight of 9+/-2 years and 492+/-17 kg, respectively. PROCEDURE: The day before anesthesia, urine was collected (catheterization) for 3 hours to quantitate baseline values, and serum biochemical analysis was performed. The following day, xylazine (1.1 mg/kg, IV) was administered, and general anesthesia was induced 5 minutes later with diazepam (0.04 mg/kg, IV) and ketamine (2.2 mg/kg, IV). During 2 hours of anesthesia with isoflurane, Paco2 was maintained between 48 and 52 mm Hg, and mean arterial blood pressure was between 70 and 80 mm Hg. Blood and urine were collected at 30, 60, and 120 minutes during and at 1 hour after anesthesia. RESULTS: Baseline urine flow was 0.92+/-0.17 ml/kg/h and significantly increased at 30 and 60 minutes after xylazine administration (2.14+/-0.59 and 2.86+/-0.97 ml/kg/h respectively) but returned to baseline values by the end of anesthesia. Serum glucose concentration increased from 12+/-4 to 167+/-8 mg/dl at 30 minutes. Glucosuria was not observed. CONCLUSIONS AND CLINICAL RELEVANCE: Transient hyperglycemia and an increase in rine production accompanies a commonly used anesthetic technique for horses. The increase in urine flow is not trivial and should be considered in anesthetic management decisions. With the exception of serum glucose concentration and urine production, the effect of general anesthesia on indices of renal function in clinically normal horses is likely of little consequence in most horses admitted for elective surgical procedures. PMID- 12118670 TI - Pharmacokinetic characteristics and tissue residues for marbofloxacin and its metabolite N-desmethyl-marbofloxacin in broiler chickens. AB - OBJECTIVES: To determine pharmacokinetic characteristics of marbofloxacin after a single IV and oral administration and tissue residues after serial daily oral administration in chickens. ANIMALS: 40 healthy broiler chickens. PROCEDURE: Two groups of chickens (groups A and B; 8 chickens/group) were administered a single IV and oral administration of marbofloxacin (2 mg/kg). Chickens of group C (n = 24) were given serial daily doses of marbofloxacin (2 mg/kg, PO, q 24 h for 3 days). Plasma (groups A and B) and tissue concentrations (group C) of marbofloxacin and its major metabolite N-desmethyl-marbofloxacin were determined by use of high-performance liquid chromatography. Residues of marbofloxacin and N desmethylmarbofloxacin were measured in target tissues. RESULTS: Elimination half life and mean residence time of marbofloxacin in plasma were 5.26 and 4.36 hours after IV administration and 8.69 and 8.55 hours after oral administration, respectively. Maximal plasma concentration was 1.05 microg/ml, and interval from oral administration until maximum concentration was 1.48 hours. Oral bioavailability of marbofloxacin was 56.82%. High concentrations of marbofloxacin and N-desmethyl-marbofloxacin were found in the kidneys, liver, muscles, and skin plus fat 24 hours after the final dose of marbofloxacin; however, marbofloxacin and N-desmethyl-marbofloxacin were detected in only hepatic (27.6 and 98.7 microg/kg, respectively) and renal (39.7 and 69.1 microg/kg, respectively) tissues 72 hours after termination of marbofloxacin treatment. CONCLUSIONS AND CLINICAL RELEVANCE: Analysis of pharmacokinetic data obtained in this study reveals that a minimal therapeutic dose of 2 mg/kg, PO, every 24 hours should be appropriate for control of most infections in chickens. PMID- 12118671 TI - In vitro anion transport alterations and apoptosis induced by phenylbutazone in the right dorsal colon of ponies. AB - OBJECTIVES: To study the functional and structural responses of the right dorsal colon (RDC) of ponies to phenylbutazone (PBZ) in vitro at a concentration that could be achieved in vivo. ANIMALS: 8 adult ponies. PROCEDURE: Short circuit current and conductance were measured in mucosa from the RDC. Tissues incubated with and without HCO3- were exposed to PBZ, bumetanide, or indomethacin. Bidirectional Cl- fluxes were determined. After a baseline flux period, prostaglandin E2 (PGE2) was added to the serosal surfaces and a second flux period followed. Light and transmission electron microscopy were performed. RESULTS: Baseline short circuit current was diminished significantly by PBZ and indomethacin, and increased significantly after addictions of PGE2. After PGE2 was added, Cl- secretion increased significantly in tissues in HCO3- -free solutions and solutions with anti-inflammatory drugs, compared with corresponding baseline measurements and with control tissues exposed to PGE2. Bumetanide did not affect baseline short circuit current and Cl- fluxes. The predominant histologic change was apoptosis of surface epithelial cells treated with PBZ and to a lesser extent in those treated with indomethacin. CONCLUSIONS AND CLINICAL RELEVANCE: Prostaglandin-induced Cl- secretion appeared to involve a transporter that might also secrete HCO3-. Both PBZ and indomethacin altered ion transport in RDC and caused apoptosis; PBZ can damage mucosa through a mechanism that could be important in vivo. The clinically harmful effect of PBZ on equine RDC in vivo could be mediated through its effects on Cl- and HCO3- secretion. PMID- 12118672 TI - Expression of a chemokine by ciliary body epithelium in horses with naturally occurring recurrent uveitis and in cultured ciliary body epithelial cells. AB - OBJECTIVE: To determine whether a chemokine (RANTES)-like protein expressed by ciliary epithelium plays a role in uveitis. SAMPLE POPULATION: 3 clinically normal horses intradermal, 5 eyes from 5 horses with recurrent uveitis, and 10 normal eyes from 5 age- and sex-matched horses. PROCEDURE: Cross-reactivity and sensitivity of recombinant human (rh)-regulated upon activation, normal T-cell expressed and secreted (RANTES) protein were evaluated in horses by use of intradermal hypersensitivity reactions and a chemotaxis assay. Aqueous humor and ciliary body of eyes from clinically normal horses and horses with uveitis were examined for RANTES expression by use of an ELISA and reverse transcription polymerase chain reaction (RT-PCR). Expression of RANTES mRNA and protein content of primary cultures of equine ciliary pigmented epithelial cells (RT-PCR) and culture supernatant (ELISA) were measured 6 or 24 hours, respectively, after cultures were stimulated with interleukin-1beta and tumor necrosis factor-alpha. RESULTS: Strong reactions to intradermal hypersensitivity testing and significant chemotaxis of equine leukocytes to rh-RANTES wereas observed. Aqueous humor of eyes from horses with uveitis contained increased concentrations of rh-RANTES like protein (mean +/- SD, 45.9+/-31.7 pg/ml), compared with aqueous humor from clinically normal horses (0 pg/ml). Ciliary body from horses with uveitis expressed RANTES mRNA, whereas ciliary body from clinically normal horses had low mRNA expression. Stimulated ciliary pigmented epithelial cells expressed increased amounts of rh-RANTES-like protein (506.1+/-298.3 pg/ml) and mRNA, compared with unstimulated samples. CONCLUSIONS AND CLINICAL RELEVANCE: Ciliary epithelium may play a role in recruitment and activation of leukocytes through expression of RANTES. PMID- 12118673 TI - Seasonal interaction of serum vitamin D concentration and bone density in alpacas. AB - OBJECTIVE: To evaluate temporal changes in bone mineral density associated with seasonal variation in serum vitamin D, calcium, and phosphorus concentrations in alpacas. ANIMALS: 5 healthy mature neutered male alpacas. PROCEDURE: Metacarpal bone mineral density was measured at 4 times during a year. Each time alpacas were weighed, blood was collected for determination of serum calcium, phosphorus, and vitamin D concentrations, and samples of feed were analyzed for nutrient content. Vitamin D status was determined by use of an assay that measured serum 25-hydroxycalciferol concentration. Effects of changes in serum vitamin D, calcium, and phosphorus concentration and body weight with season on bone mineral density were determined. RESULTS: Bone mineral density, body weight, and serum vitamin D and phosphorus concentrations varied with season. Bone mineral density, serum vitamin D concentration, and body weight also varied among individual alpacas. Serum vitamin D concentration was lower in January than the previous October and increased from May to the following September. The decrease in bone mineral density lagged behind the decrease in serum vitamin D concentration and was lower in May, compared with the previous October. Body weight was lower in May than the previous October or following September. Solar radiation was highest in July and lowest in December. CONCLUSIONS AND CLINICAL RELEVANCE: Seasonal changes in bone mineral density are associated with changes in serum vitamin D concentrations in alpacas. Changes in bone mineral density associated with a decline in serum vitamin D concentration may predispose some alpacas to developing fractures minimal trauma. PMID- 12118674 TI - Multivariate regression analysis of epidural pressure in cattle. AB - OBJECTIVE: To evaluate the effects of growth, maturity, and pregnancy on epidural pressure in cattle. ANIMALS: 50 healthy Holstein cattle (18 heifers, 23 lactating cows, and 9 pregnant nonlactating cows). PROCEDURE: Each of the cattle was restrained in a standing position. Height of the second lumbar vertebra's transverse process (2LTP) and humeral tuberosity (HT) on the right side as well as abdominal girth (AG) were measured in each animal, and body condition score (BCS) was ascertained. Skin caudal to the first lumbar spinous process was aseptically prepared, and anesthetic was injected. After inserting a 16-gauge 120 mm Tuohy needle in the ligamentum flavum, a calibrated pressure transducer was connected to the needle. Then, the needle was introduced into the epidural space, and epidural pressure was recorded. RESULTS: Mean +/- SD residual epidural pressure of heifers (-9.3+/-3.3 mm Hg) was significantly higher than that of lactating (-174+/-5.5 mm Hg) or nonlactating (-14.5+/-2.4 mm Hg) cows. Stepwise regression of 5 variables revealed that only the difference in height between 2LTP and HT (2LTP - HT) in heifers and only BCS in lactating cows were significantly correlated with residual epidural pressure. For all cattle, the optimal equation (R2 = 0.47) describing the relationship was y = -12.7 + 6.3x, - 0.4x2 - 0.1x3, where y is epidural pressure, x1 is BCS, x2 is 2LTP - HT, and x3 is age. CONCLUSIONS AND CLINICAL RELEVANCE: Negative epidural pressure was detected in standing cattle. Growth, maturity, and pregnancy affect epidural pressure in cattle. PMID- 12118675 TI - Effects on functions of ovine blood mononuclear cells for each of several fatty acids at concentrations found in plasma of healthy and ketotic ewes. AB - OBJECTIVE: To assess effects on functions of peripheral blood mononuclear cells (PBMC) obtained from ewes for each of several fatty acids represented in ovine plasma at concentrations mimicking those of ketotic or healthy ewes. SAMPLE POPULATION: Blood samples obtained from 6 Sardinian ewes. PROCEDURE: The PBMC were cultured in media that contained oleic (OA), palmitic (PA), stearic (SA), linoleic (LA), or palmitoleic (POA) acid at concentrations similar to those of ketotic or healthy ewes. Synthesis of DNA was stimulated by use of concanavalin A or pokeweed mitogen (PWM). Secretion of IgM was stimulated by use of PWM. RESULTS: High concentrations (900, 450, and 225 micromol/L) of OA significantly inhibited DNA synthesis and IgM secretion of PBMC. Conversely, low concentrations (56 or 28 micromol/L) of OA significantly enhanced DNA synthesis of PBMC. High concentrations of PA (600, 300, 150, 75, 375, or 18.7 micromol/L) and SA (300, 150, or 75 micromol/L) significantly inhibited DNA synthesis of PBMC. High concentrations of PA (600, 300, 150, 75, 375, or 18.7 micromol/L) and SA (300, 150, 75, or 38 micromol/L) also significantly inhibited IgM secretion of PBMC. None of the concentrations of LA and POA affected PBMC functions. CONCLUSION AND CLINICAL RELEVANCE: Impaired immunoresponsiveness of ketotic ewes is likely associated with an increase of plasma concentrations of OA, PA, or SA and not with that of LA or POA. At physiologic concentrations, single fatty acids are likely to participate in modulation of immunoresponsiveness by exerting suppressive or stimulatory effects on immune cells. PMID- 12118676 TI - Validation of a cell culture bioassay for detection of petroleum exposure in mink (Mustela vison) as a model for detection in sea otters (Enhydra lutris). AB - OBJECTIVE: To validate a luciferase bioassay, which is based on a recombinant mouse hepatoma cell line, for the detection of exposure to petroleum in mustelid species. ANIMALS: 122 American mink (Mustela vison) and 15 sea otters (Enhydra lutris). PROCEDURES: Mink were exposed to Bunker C fuel oil or Alaska North Slope crude oil externally as a single exposure or internally via low dose concentrations in their ration for 6 months. Serum samples were analyzed for cytochrome P450 1A1 induction by quantification of luciferase activity in the bioassay. Mink liver specimens were also evaluated for cytochrome P450 1A1 induction by quantification of ethoxyresorufin-o-deethylase activity. Serum collected from exposed and unexposed sea otters was also analyzed using the luciferase bioassay. RESULTS: Serum samples from mink externally exposed to petroleum had significantly increased luciferase activities at 1 week after exposure. Serum samples taken at later time points or from mink exposed to either product in the ration did not cause significant luciferase induction. Samples from otters exposed to petroleum had significantly higher luciferase induction as compared with samples from otters not exposed to petroleum at 2 and 8 years after the spill. Cytochrome P450 1A1 activity in liver specimens collected from mink that were internally exposed through diet was significantly increased at the conclusion of our study. CONCLUSION AND CLINICAL RELEVANCE: The luciferase bioassay is a sensitive and specific method for determining recent exposure to petroleum in mink. The lack of luciferase activity in serum samples collected from mink greater than 1 week after experimental exposure was likely attributable to lower overall petroleum exposure in our trial, compared with natural exposures. PMID- 12118677 TI - Evaluation of the perioperative stress response in dogs administered medetomidine or acepromazine as part of the preanesthetic medication. AB - OBJECTIVE: To compare the perioperative stress response in dogs administered medetomidine or acepromazine as part of the preanesthetic medication. ANIMALS: 42 client-owned dogs that underwent elective ovariohysterectomy. PROCEDURE: Each dog was randomly allocated to receive medetomidine and butorphanol tartrate (20 microgram/kg and 0.2 mg/kg, respectively, IM) or acepromazine maleate and butorphanol (0.05 and 0.2 mg/kg, respectively, IM) for preanesthetic medication. Approximately 80 minutes later, anesthesia was induced by administration of propofol and maintained by use of isoflurane in oxygen. Each dog was also given carprofen before surgery and buprenorphine after surgery. Plasma concentrations of epinephrine, norepinephrine, cortisol, and beta-endorphin were measured at various stages during the perioperative period. In addition, cardiovascular and clinical variables were monitored. RESULTS: Concentrations of epinephrine, norepinephrine, and cortisol were significantly lower for dogs administered medetomidine. Concentrations of beta-endorphin did not differ between the 2 groups. Heart rate was significantly lower and mean arterial blood pressure significantly higher in dogs administered medetomidine, compared with values for dogs administered acepromazine. CONCLUSIONS AND CLINICAL RELEVANCE: Results indicate that for preanesthetic medications, medetomidine may offer some advantages over acepromazine with respect to the ability to decrease perioperative concentrations of stress-related hormones. In particular, the ability to provide stable plasma catecholamine concentrations may help to attenuate perioperative activation of the sympathetic nervous system. PMID- 12118678 TI - Bovine adenovirus serotype 7 infections in postweaning calves. AB - OBJECTIVE: To detect bovine adenovirus serotype 7 (BAV-7) infections in calves by use of viral isolation and serologic testing. ANIMALS: 205 postweaning calves. PROCEDURE: 121 calves were assembled by an order buyer through auction markets in eastern Tennessee and transported to New Mexico where they were commingled with 84 healthy ranch-reared calves. Tests included viral isolation in cell culture from peripheral blood leukocytes (PBL) and detection of serum BAV-7 antibodies by use of microtitration viral neutralization. RESULTS: BAV-7 was isolated from PBL of 8 calves and seroconversion to BAV-7 was detected for 38 of 199 (19.1%) calves. Concurrent bovine viral diarrhea virus infections were detected in most calves from which BAV-7 was isolated. CONCLUSIONS AND CLINICAL RELEVANCE: Results of our study indicate that BAV-7 infections can be found in postweaning commingled calves and may develop more commonly in calves with concurrent infections with viruses such as bovine viral diarrhea virus (BVDV). PMID- 12118679 TI - Reliability of goniometry in Labrador Retrievers. AB - OBJECTIVE: To evaluate the reliability of goniometry by comparing goniometric measurements with radiographic measurements and evaluate the effects of sedation on range of joint motion. ANIMALS: 16 healthy adult Labrador Retrievers. PROCEDURE: 3 investigators blindly and independently measured range of motion of the carpus, elbow, shoulder, tarsus, stifle, and hip joints of 16 Labrador Retrievers in triplicate before and after dogs were sedated. Radiographs of all joints in maximal flexion and extension were made during under sedation. Goniometric measurements were compared with radiographic measurements. The influence of sedation and the intra- and intertester variability were evaluated; 95% confidence intervals for all ranges of motion were determined. RESULTS: Results of goniometric and radiographic measurements were not significantly different. Results of measurements made by the 3 investigators were not significantly different. Multiple measurements made by 1 investigator varied from 1 to 6 degrees (median, 3 degrees) depending on the joint. Sedation did not influence the range of motion of the evaluated joints. CONCLUSIONS AND CLINICAL RELEVANCE: Goniometry is a reliable and objective method for determining range of motion of joints in healthy Labrador Retrievers. PMID- 12118681 TI - Linear, logarithmic, and polynomial models of M-mode echocardiographic measurements in dogs. AB - OBJECTIVE: To determine whether logarithmic and polynomial models are superior to simple linear models for predicting reference values for M-mode echocardiographic variables in dogs with a wide range of body weights. ANIMALS: 69 apparently healthy adult male and female dogs of various breeds, ages (range, 1 to 12 years; median, 3.5 years), and body weights (range, 3.9 to 977 kg; median, 25.4 kg). PROCEDURE: Echocardiographic M-mode measurements of the interventricular septum, left ventricular dimension (LVD), left ventricular wall, aorta, and left atrium were obtained. Simple linear, second-order polynomial, third-order polynomial, and logarithmic regression models were determined by use of the least-squares method to describe the relationship between M-mode measurements and body weight. Differences in adjusted R2 values of logarithmic and polynomial models were tested for significance of contribution, compared with the simple linear model. RESULTS: Significant differences in adjusted R2 were found when comparing simple linear with logarithmic or polynomial models for LVD-diastole, LVD-systole, aorta, and left atrium. Differences in adjusted R2 between second-order polynomial, third-order polynomial, and logarithmic models were not significant for any M-mode measurement. CONCLUSIONS AND CLINICAL RELEVANCE: In this study, logarithmic or second-order polynomial models predicted reference values of M mode measurements for size of the cardiac chambers better than simple linear models for dogs with a wide range of body weights. Logarithmic and polynomial models were not superior to simple linear models for M-mode measurements of cardiac wall thickness. PMID- 12118680 TI - Evaluation of the influence of prostaglandin E2 on recombinant equine interleukin 1beta-stimulated matrix metalloproteinases 1, 3, and 13 and tissue inhibitor of matrix metalloproteinase 1 expression in equine chondrocyte cultures. AB - OBJECTIVE: To determine the effects of prostaglandin E2 (PGE2) on recombinant equine interleukin (IL)-1beta-stimulated expression of matrix metalloproteinases (MMP 1, MMP 3, MMP 13) and tissue inhibitor of matrix metalloproteinase 1 (TIMP 1) in vitro. SAMPLE POPULATION: Cultured equine chondrocytes. PROCEDURE: Stationary monolayers of first-passage chondrocytes were exposed to graduated concentrations of PGE2 with or without a subsaturating dose (50 pg/ml) of recombinant equine IL-1beta (reIL-1beta) to induce expression of MMP 1, MMP 3, MMP 13, and TIMP 1, followed by RNA isolation and northern blotting. In subsequent experiments, gene expression was similarly quantified from mRNA isolated from cultures pretreated with phenylbutazone to quench endogenous PGE2 synthesis, followed by exposure to reIL-1beta and exogenous PGE2 (5 mg/ml) with appropriate controls. RESULTS: Exogenous PGE2 (10 mg/ml) significantly reduced reIL-1beta-induced expression of MMP 1, MMP 3, MMP 13, and TIMP 1. Abrogation of cytokine induction with this dose of PGE2 was comparable to that for dexamethasone (10(-5) M) control. Similarly, pretreatment with phenylbutazone, followed by exposure to relL-1beta and PGE2 (5 mg/ml), was associated with a reduced expression of the genes of interest, an effect that was significant for MMP 1, MMP 13, and TIMP 1. CONCLUSIONS AND CLINICAL RELEVANCE: The MMP and TIMP 1 are important mediators in the pathophysiologic events in osteoarthritis. The potential for physiologically relevant regulation of expression of these genes by PGE2 is a consideration in the use of drugs that inhibit prostanoid synthesis in the treatment of equine arthropathies. PMID- 12118682 TI - Understanding compassion fatigue: keys for the caring veterinary healthcare team. PMID- 12118683 TI - Effect of monthly heartworm preventatives on dogs with young heartworm infections. PMID- 12118684 TI - Clinical management of flail chest in dogs and cats: a retrospective study of 24 cases (1989-1999). AB - Cases of flail chest injury for 24 client-owned companion animals following various traumas were evaluated. Concurrently sustained injuries, initial emergency treatments, and definitive treatment and outcome for regimens that utilize stabilization of the flail segment were compared with cases treated with no stabilization. Flail chest was confirmed in 24 animals: 21 dogs and three cats. There was an even division (12 each) of right and left flail segments. The median number of ribs involved was three (range, two to seven). Flail segment stabilization was performed in nine, and 15 were treated with no stabilization. Statistical analysis using multiple data permutations evaluating all combinations failed to reveal a significant difference in outcome between stabilized and unstabilized cases. PMID- 12118685 TI - Cervical injury following a horse kick to the head in two dogs. AB - Two dogs were presented to North Carolina State University Veterinary Teaching Hospital following blunt trauma to the head delivered by a horse kick. On presentation, both dogs had resolving clinical signs directly related to the head trauma, but both also had compromise to their upper airway as a result of indirect injury to the soft tissues of the neck, visible on plain radiographs. One dog made a full recovery following a period of assisted ventilation. The other dog was euthanized at the request of the owner. These injuries illustrate the importance of evaluating the cervical spine and soft tissues of the neck following blunt trauma to the head. PMID- 12118686 TI - Correlation between fine-needle aspiration cytopathology and histopathology of the lung in dogs and cats. AB - Medical records from 28 patients having fine-needle aspiration (FNA) cytopathology and histopathology of pulmonary lesions were reviewed. Clinical signs, thoracic radiographs, cytopathology, histopathology, and complications associated with FNA were evaluated. Correlation between cytopathological and histopathological diagnoses was determined. Cytopathological specimens were classified as neoplastic, inflammatory, or nondiagnostic. Histopathological diagnoses were categorized as neoplastic or inflammatory. No complications were observed following FNA. Diagnoses obtained by FNA cytopathology accurately reflected the diagnosis obtained on histopathological examination in 82% of cases. Fine-needle aspiration cytopathology of the lung is a useful and safe diagnostic tool in dogs and cats with pulmonary parenchymal lesions. PMID- 12118687 TI - Thoracic radiography, bronchoalveolar lavage cytopathology, and pulmonary parenchymal histopathology: a comparison of diagnostic results in 11 cats. AB - The purpose of this study was to compare the diagnostic results and value of thoracic radiography, bronchoalveolar lavage (BAL) fluid cytopathology, and lung histopathology in 11 cats with spontaneous respiratory disease in which radiography and cytopathology were inadequate in establishing a definitive diagnosis. In these cats, radiographic patterns were characterized as bronchial (n=6), interstitial (n=3), and alveolar (n=2); other features included hyperinflation (n=3), bronchiectasis (n=2), pleural fissure lines (n=2), pulmonary nodules (n=2), atelectasis (n=1), and a tracheal mass (n=1). Bronchoalveolar lavage fluid was unremarkable in two cats. Abnormal BAL fluid showed inflammation (n=5), hemorrhage (n=2), epithelial hyperplasia (n=1), or was suspicious for neoplasia (n=1). Histopathological evaluation revealed inflammation (n=8), neoplasia (n=2), and vascular congestion (n=1). The predominant radiographic location of disease correlated with the same histopathological location in seven cats, and the cytopathological class of BAL fluid was consistent with the histopathological class of disease in seven cats. There was poor correlation between the types of cells found in the BAL fluid and the pathologist's prediction of the types of cells likely to be found in the BAL fluid based on the amount and type of airway cellularity seen on histopathological examination. The results of this study suggest that in some cats, BAL fluid cytopathology does not always correlate with the type of pulmonary disease identified on histopathology. In respiratory diseases where radiography and cytopathology fail to provide a definitive diagnosis, histopathological examination of the lung may be necessary. PMID- 12118688 TI - Canine synovial sarcoma: a retrospective assessment of described prognostic criteria in 16 cases (1994-1999). AB - Pertinent patient data and biopsied tissue from 16 cases of canine synovial sarcoma (SS) were reviewed. Histopathological grade, clinical stage, and tissue immunoreactivity to cytokeratin (broad stain, AE1/AE3 and cytokeratin 7) and vimentin were determined and correlated with survival. Effect of treatment on survival was similarly evaluated. Neither clinical stage nor histopathological grade significantly affected survival patterns. Tissues from all cases stained >30% positively with vimentin, whereas no tissue from any case exhibited cytokeratin immunoreactivity. Dogs receiving surgical tumor excision or amputation had a significantly higher survivability than those receiving no treatment (P<0.02). Treatment aggressiveness may be more appropriate than clinical staging or tumor grading in predicting survival. Reliability of diagnosing and prognosticating canine SS with current immunohistochemistry protocols should be questioned. PMID- 12118689 TI - Hematological toxicity and therapeutic efficacy of lomustine in 20 tumor-bearing cats: critical assessment of a practical dosing regimen. AB - Twenty cats with spontaneously arising tumors received oral lomustine at a dose range of 32 to 59 mg/m2 every 21 days. Due to biohazard concerns associated with lomustine capsule reformulation, a standardized 10-mg capsule dosage was used for all cats regardless of body weight. Severe hematological toxicity was infrequent, with the incidence of either grade III or IV neutropenia and thrombocytopenia being 4.1% and 1.0%, respectively. Cats receiving higher cumulative doses of lomustine trended toward a greater likelihood for progressive neutropenia (P=0.07). Two cats with lymphoma, two cats with fibrosarcoma, and one cat with multiple myeloma achieved a measurable partial response to lomustine therapy. Cats treated with higher dosages of lomustine trended toward statistically significant higher response rates (P=0.07). PMID- 12118690 TI - Intertarsal and tarsometatarsal arthrodesis using 2.0/2.7-mm or 2.7/3.5-mm hybrid dynamic compression plates. AB - Proximal intertarsal or tarsometatarsal arthrodesis was performed in four dogs using either 2.0/2.7-mm or 2.7/3.5-mm hybrid dynamic compression plates. Mean radiographic follow-up time was 30.5 weeks (range, 15 to 60 weeks). Mean owner follow-up time was 40.5 weeks (range, 27 to 60 weeks). All cases went on to form stable, healed arthrodeses. Owners reported no noticeable lameness problems long term. Complications occurred in one case and included pressure sores and fracture of the calcaneus. Implant failure did not occur in any case. PMID- 12118691 TI - Severe carpal and tarsal shearing injuries treated with an immediate arthrodesis in seven dogs. AB - The medical records of seven dogs with severe, grade 3, open shearing wounds of the carpus or tarsus that were treated with an immediate arthrodesis were reviewed. Six dogs were managed with a transarticular external skeletal fixator (ESF), and one dog was treated with plate fixation. The soft tissues were managed simultaneously along with the definitive joint stabilization in all cases. Minor complications occurred in four dogs: one dog that required a skin graft, one dog in which a skin graft was recommended but not performed, one dog in which a secondary skin closure was performed, and one dog in which a delayed cancellous bone graft was placed. Major complications occurred in three dogs: two dogs that required restabilization of the arthrodesis and one dog that required implant (i.e., plate) removal due to infection. All dogs healed with acceptable functional and cosmetic results. Further long-term evaluation of five dogs revealed that all but one dog had either excellent or good functional outcomes, and the remaining dog had a fair outcome. Similarly, five dogs had either excellent or good cosmetic outcomes, with the remaining dog having a fair outcome. All owners were very satisfied with the overall results. This clinical study demonstrated that an immediate, definitive management technique, in which bone fusion and wound management are undertaken as simultaneous objectives, is a viable technique of managing severe periarticular shearing injuries. Transarticular stabilization with an ESF is the technique recommended. PMID- 12118692 TI - A urethropexy technique for surgical treatment of urethral prolapse in the male dog. AB - Urethral prolapse is an uncommon condition affecting young male dogs, most commonly English bulldogs. Current described techniques for surgical treatment of urethral prolapse involve manual reduction of prolapsed mucosa and placement of a temporary purse-string suture at the penile tip, or resection of the prolapsed tissue and apposition of urethral and penile mucosa. The incidence of recurrence of urethral prolapse following resection of the prolapse is not known. This report describes a technique for surgical treatment of urethral prolapse in the male dog that minimizes surgical and anesthetic time, is simple to perform, requires minimal equipment, is effective, and is not associated with significant complications or recurrence. Three cases are described. PMID- 12118693 TI - Evaluation of a technique using the carbon dioxide laser for the treatment of aural hematomas. AB - A new technique using the carbon dioxide (CO2) laser for the treatment of aural hematomas is described. The laser is used to make an incision into the hematoma to allow for evacuation of the blood, and then multiple, small incisions are made over the surface of the hematoma to stimulate adhesions between the tissue layers. The CO2 laser was used in this fashion to treat 10 aural hematomas in eight dogs. Follow-up ranged from 1 to 23 months. Owners evaluated the cosmetic results following CO2 laser surgery as excellent in three ears, good in five ears, and fair in two ears. Hematomas were resolved in all 10 cases, although two cases developed serosanguineous fluid accumulation that required percutaneous drainage in one case and a second laser procedure in the other case. PMID- 12118694 TI - Reducing childhood pedestrian injuries. PMID- 12118695 TI - Production of a Pseudomonas lipase in n-alkane substrate and its isolation using an improved ammonium sulfate precipitation technique. AB - Among the various lipidic and non-lipidic substances, normal alkanes within the chain lengths of C-12 to C-20 served as the best carbon substrates for the production of extracellular lipase by Pseudomonas species G6. Maximum lipase production of 25 U/ml of the culture broth was obtained by using n-hexadecane as the sole carbon substrate. The optimum pH of 8 and temperature of 34 + 1 degrees C were demonstrated for the production of lipase in n-hexadecane substrate. The optimum concentration of iron, which played a critical role on the lipase production, was found to be 0.25 mg/l. Lipase production could be enhanced to nearly 2.4-fold by using tributyrin at a concentration of 0.05% (v/v) in the culture medium. High recovery of the lipase protein (83%) from the culture broth was achieved by treating the culture supernatant with Silicone 21 Defoamer followed by ammonium sulfate (60% saturation) fractionation. PMID- 12118696 TI - Effects of anaerobic/aerobic incubation and storage temperature on preservation and deodorization of kitchen garbage. AB - To develop a garbage recycling system for the purpose of the production of lactic acid (LA) to use as raw material for producing biodegradable plastics, the preservation and deodorization of garbage during storage are very important. Anaerobic incubation (i.e., storage) was prove to be more suitable than aerobic incubation during the garbage storage in terms of concentration of LA and soluble sugar, pH value, viable bacteria counts and offensive odour substances. This difference is due to a fact that the growth of putrefactive bacteria such as coliforms and Clostridium spp. appeared to be inhibited by anaerobic fermentation during the storage, because the fermentation caused a drop of garbage pH and generated inhibitory substances, i.e., bacteriocins. Under anaerobic condition, LA concentration in the stored garbage was found to be higher in the order: 37 > 25 > 50 > 5 degrees C, and the concentration of sugar accumulated during the 50 degrees C-storage was the highest. Among the conditions employed, the optimum condition for the storage of kitchen garbage was anaerobic at 5 degrees C. PMID- 12118697 TI - Phytoremediation of subarctic soil contaminated with diesel fuel. AB - The effects of several plant species, native to northern latitudes, and different soil amendments, on diesel fuel removal from soil were studied. Plant treatments included Scots Pine (Pinus sylvestris), Poplar (Populus deltoides x Wettsteinii), a grass mixture (Red fescue, Fesuca rubra; Smooth meadowgrass, Poa pratensis and Perennial ryegrass, Lolium perenne) and a legume mixture (White clover, Trifolium repens and Pea, Pisum sativum). Soil amendments included NPK fertiliser, a compost extract and a microbial enrichment culture. Diesel fuel disappeared more rapidly in the legume treatment than in other plant treatments. The presence of poplar and pine enhanced removal of diesel fuel, but removal under grass was similar to that with no vegetation. Soil amendments did not enhance diesel fuel removal significantly. Grass roots accumulated diesel-range compounds. This study showed that utilisation of selected plants accelerates removal of diesel fuel in soil and may serve as a viable, low-cost remedial technology for diesel contaminated soils in subarctic regions. PMID- 12118698 TI - Pollution attenuation by soils receiving cattle slurry after passage of a slurry like feed solution. Column experiments. AB - Designing soil filtration systems or vegetated filter strips as a means of attenuating water pollution should take into account soil purging capacity. Here we report data on laboratory column trials used to investigate the capacity of a Hortic Anthrosol to attenuate contamination due to downward leaching from cattle slurry applied at the surface. The columns comprised 900 g of soil to a depth of about 20-25 cm, and had been used previously in an experiment involving passage of at least 5 pore volumes of an ion-containing cattle slurry-like feed solution. For the present experiments, the columns were first washed through with distilled water (simulating resting and rain falling after passage of the feed solution), and then received a single slurry dose equivalent to about 300 m3 ha(-1). The columns were then leached with distilled water, with monitoring of chemical oxygen demand (COD) and ion contents in outflow. The results indicated that the pollution-neutralising capacity of the soil was still high but clearly lower than in the earlier experiments with the feed solution. Furthermore, the time-course of COD showed that organic acids were leached through the column even more rapidly than chloride (often viewed as an inert tracer) enhancing the risk of heavy metals leaching and subsequent water pollution. Resting and alternate use of different soil-plant buffer zones would increase the lifespan of purging systems that use soil like the here studied one. PMID- 12118699 TI - Improvement of the composting time for household waste during an initial low pH phase by mesophilic temperature control. AB - Earlier studies indicated that the activity in the initial phase of composting may be reduced when the temperature rises too fast under low pH conditions. A compost reactor experiment on household waste was designed to test whether the degradation time could be reduced by actively preventing the temperature from rising until the pH had reached a certain value. This experiment was performed by monitoring pH in the condensate from the cooled compost gas. The results from 3 + 3 runs with and without temperature control confirmed our hypothesis and a considerable reduction in composting time was achieved. One possible explanation for the results is that the microbes active in the low pH phase are hampered by high temperature. The abrupt rise in pH when the fatty acids are consumed seems to be a good marker of the point when temperature control can be discontinued. PMID- 12118700 TI - Water soluble fractions of rose-scented geranium (Pelargonium species) essential oil. AB - The essential oil of rose-scented geranium (Pelargonium species, family: Geraniaceae) obtained through steam or water plus steam distillation of shoot biomass is extensively used in the fragrance industry and in aromatherapy. During distillation, a part of the essential oil becomes dissolved in the distillation water (hydrosol) and is lost as this hydrosol is discarded. In this investigation, hydrosol was shaken for 30 min with hexane (10:1 proportion) and the hexane was distilled to yield 'secondary' or 'recovered' essential oil. The chemical composition of secondary oil was compared with that of 'primary' oil (obtained directly by distilling shoot biomass of the crop). Primary oil accounted for 93.0% and secondary oil 7.0% of the total oil yield (100.2 ml from 100 kg green shoot biomass). Fifty-two compounds making up 95.0-98.5% of the primary and the secondary oils were characterized through gas chromatography (GC) and gas chromatography-mass spectroscopy (GC--MS). Primary oil was richer in hydrocarbons (8.5-9.4%), citronellyl formate (6.2-7.5%), geranyl formate (4.1 4.7%), citronellyl propionate (1.0-1.2%), alpha-selinene (1.8-2.2%), citronellyl butyrate (1.4-1.7%), 10-epi-gamma-eudesmol (4.9-5.5%) and geranyl tiglate (1.8 2.1%). Recovered oil was richer in organoleptically important oxygenated compounds (88.9-93.9%), commercial rhodinol fraction (74.3-81.2%), sabinene (0.4 6.2%), cis-linool oxide (furanoid) (0.7-1.2%), linalool (14.7-19.6%), alpha terpineol (3.3-4.8%) and geraniol (21.3-38.4%). Blending of recovered oil with primary oil is recommended to enhance the olfactory value of the primary oil of rose-scented geranium. Distillation water stripped of essential oil through hexane extraction can be recycled for distilling the next batch of rose-scented geranium. PMID- 12118701 TI - The utilization of alkali-treated melon husk by broilers. AB - The effects of alkali treatment on chemical constituents of melon husk (MH) and performance characteristics of broilers fed alkali-treated MH (ATMH) diets were investigated. The chemical analysis showed that alkali treatment increased the ash content of MH (from 15.70% to 16.86%) and reduced the crude fibre content (from 29.00% to 14.00%). Result of feed intake was superior on 30% alkali diet with a value of 100.14 g/bird/day. Body weight gain decreased with increase in the level of ATMH in the diet. Highest dressing percentage of 66.33% and best meat/bone ratio of 2.57 were obtained on 10% and 20% alkali diets, respectively. Dietary treatments had significant effect (P < 0.05) on gizzard weight. Up to 20% of maize can be replaced with ATMH in broiler diets to produce good quality poultry carcases and chicken meat with favourable shelf life. PMID- 12118702 TI - Phenolic removal in olive oil mill wastewater by strains of Pleurotus spp. in respect to their phenol oxidase (laccase) activity. AB - The ability of several Pleurotus spp. strains to remove phenolic compounds from an olive oil mill wastewater (OMW) was studied. All strains tested in this work were able to grow in OMW without any addition of nutrients and any pre-treatment, except sterilization. High laccase activity was measured in the growth medium, while 69-76% of the initial phenolic compounds were removed. The black color of OMW became yellow-brown and brighter as the strains grew. The lowest phenolic concentrations were reached after 12/15 days. A decrease of the phytotoxicity, as described by the parameter Germination Index, was noticed in the OMW treated with some Pleurotus spp strains, although this decrease was not proportional to the phenolic removal. A new parameter, namely Phenol-toxicity Index, was considered in the present paper. Using this parameter it was found that the remaining phenolics and/or some of the oxidation products of the laccase reaction in the treated OMW were more toxic than the original phenolic compounds. PMID- 12118703 TI - Production of laccase by a newly isolated strain of Trametes modesta. AB - The effects of the carbon and nitrogen sources, initial pH and incubation temperature on laccase production by Trametes modesta were evaluated using the one-factor-at-a-time method. The final optimisation was done using a central composite design resulting in a four-fold increase of the laccase activity to 178 nkat ml(-1). Response-surface analysis showed that 7.34 g l(-1) wheat bran, 0.87 g l(-1) glucose, 2.9 g l(-1) yeast extract, 0.25 g l(-1) ammonium chloride, an initial pH of 6.95 and an incubation temperature of 30.26 degrees C were the optimal conditions for laccase production. Laccase produced by T. modesta was fully active at pH 4 and at 50 degrees C. The laccase was very stable at pH 4.5 and at 40 degrees C but half-lives decreased to 120 and 125 min at higher temperature (60 degrees C) and lower pH (pH 3). PMID- 12118704 TI - Cadmium removal by living cells of the marine microalga Tetraselmis suecica. AB - Cadmium removal by living cells of the marine microalga Tetraselmis suecica was tested in cultures exposed to different cadmium concentrations (0.6, 3, 6, 15, 30 and 45 mg/l). The EC50 for growth was 7.9 mg Cd/l after six days of exposure. The cadmium removed was proportional to the concentration of this metal in the medium and it was dependent on the time of exposure; cultures with higher cadmium concentration removed a higher amount of this metal. In cultures exposed to 0.6 mg/l, T suecica cells removed 98.1% of added cadmium with 0.392 x 10(-6) microg Cd/cell, whereas in cultures with 45 mg/l only 7.7% was removed with 16.052 x 10( 6) microg Cd/cell. The highest amount of cadmium removed per liter of culture was observed in cultures exposed to 6 mg/l, with 3.577 mg/l of cadmium. After six days of incubation, the higher proportion of cadmium was bioaccumulated intracellularly in all cultures except in 45 mg/l cultures, the percentage of intracellular cadmium being always more than 50%. The highest percentage of bioadsorbed cadmium (60.1%) was found in cells of cultures with the highest cadmium concentration (45 mg/l). Furthermore, a relation between intracellular cadmium and the concentration of sulfhydryl groups was observed. PMID- 12118705 TI - Effect of brewery sludge amendments on some chemical properties of acid soil in pot experiments. AB - A laboratory experiment was performed for an incubation period of 120 days in order to evaluate the changes in chemical properties of an acid soil amended with 0, 15, 30, 60 and 120 t ha-' of brewery sludge (BS). Increasing BS rates and incubation time reduced pH of the soil by 0.3-0.5 unit with respect to the control while soluble salts increased from 0.11 to 0.80 dS m(-1). Organic C, exchangeable cations, soluble cations and anions, NH4+-N and NO3--N contents of the amended soil increased as BS rates increased. In addition, BS application caused a slight increase in cation exchange capacity (CEC) and a slight decrease in exchangeable acidity. PMID- 12118706 TI - The performance and potential of faecal separation and urine diversion to recycle plant nutrients in household wastewater. AB - In household wastewater and biodegradable solid waste, the main proportion of the plant nutrients are found in the toilet water (i.e. in urine and faeces). In order to recover most of these nutrients, with the purpose of decreasing the emission of eutrophicating agents and of increasing their recycling, present waste and wastewater systems have to be changed. If the urine and the faeces are collected, up to 91%, 83%, and 59% for N, P and K, respectively, can be recovered and recycled from the household wastewater. The urine was collected separately in a double flushed urine-diverting toilet and the faeces were separated from the flushwater using two parallel Aquatrons. The Aquatron separates by a combination of a whirlpool effect, gravitation and surface tension. In this study, nitrogen and phosphorus from the faeces were separated to 73% and 58%, respectively, to the dryer fraction. The combination of faecal separation and urine diversion in the house Ekoporten made it possible to collect 60% of N, 46% of P and 43% of K from the wastewater, indicating that this method is an alternative when aiming to recover plant nutrients while still wanting to use water-flushed, urine-diverting toilets, though the potential to collect the nutrients is much higher. PMID- 12118707 TI - Determination of degradation of radioactivity and its kinetics in aerobic composting. AB - In this study, the kinetics of disappearance of radioactivity in aerobic composting was investigated. For this purpose, compost materials were prepared by mixing sugar beet wastes, wine factory wastes (grape wastes), straw and biological treatment sludge in different amounts. While alpha-radioactivity was not initially detected in all composting materials, the composting materials had some beta-radioactivity. In the mixtures of sugar beet wastes--straw-biological treatment sludge (1), sugar beet wastes-wine factory wastes (grape wastes) biological treatment sludge (II) and wine factory wastes (grape wastes) biological treatment sludge (III), the beta-radioactivity reduced by 82%, 58%, 85% respectively of initial values after 52 d. The beta-radioactivity degradation in the composting process could be represented by first-order kinetics and reaction rate constants of mixtures of I, II and III were k = 0.0693 d(-1) (R2 - 0.84), k = 0.0453 d(-1) (R2 = 0.98), k = 0.0234 d(-1) (R2 = 0.97), respectively. PMID- 12118709 TI - Removal of Ni(II) from aqueous solution by adsorption onto hazelnut shell activated carbon: equilibrium studies. AB - Activated carbon prepared from hazelnut shell was used as an adsorbent for the removal of Ni(II) from aqueous solution. Batch mode adsorption studies were carried out by varying initial metal ion concentration, agitation speed, temperature and particle size. A contact time of 180 min was required to reach equilibrium. The equilibrium data were analysed using the Langmuir, Freundlich and Temkin isotherms. The characteristic parameters for each isotherm were determined. The Langmuir isotherm provided the best correlation for Ni(II) onto the activated carbon. Thermodynamical parameters revealed that the adsorption of Ni(II) is exothermic in nature. PMID- 12118708 TI - Flocculation properties of pectin in various suspensions. AB - Pectin had a flocculating activity and its flocculating activities in various suspensions were investigated. Flocculating activity of pectin in a kaolin suspension was markedly stimulated by the addition of Al3+ and Fe3+ to the suspension. Optimum temperature for flocculating activity of pectin in the kaolin suspension was around 30 degrees C and high flocculating activity was obtained when 30 mg/l of pectin and 0.2 mM Fe3+ were added to the suspension. Other inorganic suspensions of activated carbon and acid clay were flocculated by pectin in the presence of Al3+ or Fe3+. Flocculation of organic suspensions such as cellulose and yeast by pectin occurred when 0.1-0.2 mM Fe3+ was present in the suspensions. PMID- 12118710 TI - Effect of distillery effluents on some physiological aspects in maize. AB - A field experiment was conducted for two years to study the effect of application of different distillery effluents: raw spent wash (RSW), biomethanated spent wash (BSW), lagoon sludge (LS), recommended NPK + FYM (farm yard manure) and control (no fertilizer and effluent) on some physiological aspects in maize. The study revealed that the application of distillery effluents resulted in increased leaf area, chlorophyll content, nitrate reductase activity total dry weight and grain yield. Among the effluents, the highest grain yield (36.9 qha(-1)) was obtained in BSW followed by RSW (32.2 qha(-1)) and LS (28.3 qha(-1)). Overall, NPK + FYM treatment recorded the highest grain yield (51.8 qha(-1)). However, to achieve the full manurial potential of the effluents, some amount of fertilizer should be supplemented. PMID- 12118711 TI - Studies on desorption of individual textile dyes and a synthetic dye effluent from dye-adsorbed agricultural residues using solvents. AB - Two solvents, A and B (A: methanol, chloroform, water in the ratio 1:1:1; B: 50% methanol), were used to extract textile dyes adsorbed onto substrates for the purpose of future analyses of the amount of dyes degraded through solid state fermentation (SSF) using white rot fungi. Barley husk, apple pommace and corncob were separately soaked in five different dye solutions and a synthetic textile effluent. A maximum value of 93% desorption of Cibacron Red from corncob was achieved using solvent A. Barley husk was the only substrate from which the synthetic textile effluent could be desorbed, with 82% being recovered using solvent A. PMID- 12118712 TI - Restricted genetic and antigenic diversity of Plasmodium falciparum under mesoendemic transmission in the Venezuelan Amazon. AB - The study of genetic diversity in malaria populations is expected to provide new insights for the deployment of control measures. Plasmodium falciparum diversity in Africa and Asia is thought to reflect endemicity. In comprehensive epidemiological surveys reported here the genetic and antigenic structure of P. falciparum in the Venezuelan Amazon were studied over a 2-year period. DNA polymorphisms in glutamate-rich protein (GLURP), merozoite-surface protein 1 (MSP1) and MSP2 genes, in a multicopy element (PfRRM), all showed low diversity, 1 predominant genotype, and virtually no multi-clonal infections. Moreover, linkage disequilibrium was seen between GLURP, MSP1 and MSP2. Specific antibody responses against MSP1 and MSP2 recombinant antigens reflected the low genetic diversity observed in the parasite population. This is unexpected in a mesoendemic area, and suggests that the low diversity here may not only relate to endemicity but to other influences such as a bottleneck effect. Linkage disequilibrium and a predominant genotype may imply that P. falciparum frequently propagates with an epidemic or clonal population structure in the Venezuelan Amazon. PMID- 12118713 TI - Invasion, and short- and long-term survival of Babesia divergens (Phylum Apicomplexa) cultures in non-bovine sera and erythrocytes. AB - In order to explore the feasibility of producing a Babesia divergens live vaccine free of bovine material contaminants the parasite's ability to grow in human, sheep and horse erythrocytes and serum and serum-free medium was investigated. B. divergens was successfully maintained in bovine erythrocytes overlaid with serum free HL-1 medium. Supplementation of the culture medium with bovine or sheep serum improved parasite growth (monitored by measuring parasitaemia and uptake of tritiated hypoxanthine) whereas horse and human sera reduced parasite growth. As assessed by Giemsa's stained and FITC-labelled blood smears, the parasite invaded all erythrocyte types. Polyparasitism was less common in sheep and horse erythrocytes than in bovine and human erythrocytes. Accole stages were observed in bovine, human and sheep but not in horse erythrocytes. Proliferation following invasion was higher in human but lower in horse and sheep erythrocytes compared with bovine erythrocytes. Long-term cultures of B. divergens reached similar peak parasitaemias in human, sheep and bovine erythrocytes. Attempts to establish long term cultures in horse erythrocytes failed. These results suggest that B. divergens is not host specific at the level of host cell attachment and invasion. Instead, parasite survival appears to be decided once the organism has gained access into the cell. PMID- 12118714 TI - The relationships between intestinal damage and circulating endotoxins in experimental Trypanosoma brucei brucei infections. AB - The involvement of intestinal damage in experimental African trypanosomiasis was investigated in rats infected with Trypanosoma brucei brucei by measuring the urinary excretion of the previously administered non-metabolizable sugar probes, D-mannitol and lactulose, and the flux of FITC-dextran across isolated, everted gut segments. There was increased urinary recovery and flux of the sugar probes across the intestine which were significant (P < 0.05) and maximum at day 21 of the infection, but subsequently reduced, in the terminal stages of infection (day 33 p.i.). In the case of the everted sac studies the reductions were to less than 25% control values (P < 0.001). Levels of circulating endotoxin were increased approximately 3-fold at day 21 p.i., 4-fold at day 33 p.i., compared to controls. At day 21 there was a significant correlation (r = 0.63, P < 0.01) between the log endotoxin levels and the increased sugar excretion expressed as the lactulose/mannitol ratios. Histological studies showed damage to the villi, wall thinning and marked cellular infiltrations, which were very prominent in the proximal jejunum and duodenum. These results demonstrate that during trypanosome infections in rats, increased intestinal leakage and increased circulating endotoxins are significant pathological features. PMID- 12118715 TI - The relationships between endotoxins, nitric oxide and inflammatory cytokines in blood and intestinal tissues in experimental Trypanosoma brucei brucei infections. AB - Increased levels of circulating endotoxins are a feature of both human and experimental African trypanosomiasis. Studies with rats and mice have shown that these may originate from intestinal damage with altered permeability of the gut epithelium. Endotoxins are potent immunomodulatory substances which can initiate the production of a range of cytokines and mediators from different cell types. In rats infected with T.b. brucei we have examined possible associations of the endotoxin increases with increases in levels of TNF-alpha, IL-1beta, IL-6, IFN gamma and nitric oxide (NO). Significant increases in each substance occurred at days 21 and 33 post-infection (p.i.). The increases in cytokines were highly correlated with the endotoxin levels (e.g. at day 21 p.i. the correlation regression values were as follows: TNF-alpha, r = 0.9, P < 0.01; IL-1beta, r = 0.83, P < 0.01; IL-6, r = 0.9, P < 0.01; IFN-gamma, r = 0.7, P < 0.01). There were also strong correlations between the increased levels of several individual cytokines. Biopsies of chopped sections of small intestine tissues of rats showed a parallel production of cytokines, again with significant correlations with the circulating endotoxins. The production of NO and cytokines by the intestine may be associated with the increased transepithelial permeability which occurs during the infection. PMID- 12118716 TI - Phased activity in Heterorhabditis megidis infective juveniles. AB - The infectivity of Heterorhabditis megidis infective juveniles (IJs) increases during storage in water. We investigated whether this change can be related to other features of the IJs' behaviour. IJs were stored in water for 4 weeks at 20 degrees C, and the following parameters were assessed at intervals: infectivity for Galleria mellonella, dispersal in sand, host-finding on agar, and the percentage of IJs active in water. In addition, the behaviour of the IJs in water was described using 7 categories. Immediately after emerging from the host cadaver, IJs were highly active (99% of IJs in water were active and 65% displayed 'waving', the normal method of forward movement). Maximum responsiveness to host volatiles in an agar plate assay was recorded on day 2 (69% of IJs moved from the point of application and 44% of all IJs in the agar arena moved towards a host) and maximum dispersal in sand (5.8 cm) on day 0. These tendencies declined gradually with age, while infectivity underwent a significant increase from 11 nematodes per insect on day 0 to 38 nematodes per insect on day 9. Three phases could be distinguished in the behaviour of H. megidis IJs: an initial dispersal phase, during which infectivity was low; an infective phase, during which dispersal tendency was declining, and a third phase during which all behaviours (dispersal, infectivity and activity) were declining. Over the 4-week storage period, infectivity of H. megidis IJs was correlated (R2 = 0.83) with the percentage time IJs engaged in 'head thrusting' (a behaviour that resembles penetration). There is no evidence that the observed increase in infectivity of H. megidis strain UK211 could be accounted for by a generally greater level of motor activity, nor by an increase in responsiveness to volatile host cues, and it is suggested that it is due to an increased tendency to attempt penetration. PMID- 12118718 TI - Selection pressure towards monoxeny in Camallanus cotti (Nematoda, Camallanidac) facing an intermediate host bottleneck situation. AB - This paper describes the ability of the Asian fish nematode Camallanus cotti to carry out both heteroxeny, i.e. an indirect life-cycle using copepods as intermediate host, and monoxeny, i.e. direct infection and development in the definitive fish host. C. cotti occurs naturally in various freshwater teleosts in Asia. During the past decades it has been disseminated into closed or semi-closed aquaculture systems and aquaria around the world, mainly due to the ornamental fish trade. Under such conditions the species may frequently face a bottleneck situation with regard to the availability of copepods. It is known that C. cotti may reproduce and persist in copepod-free aquaria for several months. In order to investigate whether C. cotti has selected towards monoxeny in water systems lacking copepods, in contrast to the opposite selection pressure when copepods are present, 2 separate infection trials were run. It was shown that the parasite can infect the fish host both indirectly via copepods, and directly. However, C. cotti has significantly higher fitness, expressed as survival to maturity, when transmitted indirectly compared to the direct transmission mode. We suggest that the ability of aquarium populations of C. cotti to carry out a direct life-cycle is favoured by selection in order to avoid extinction whenever copepods are absent. It still remains unknown, however, whether the parasite shows the same characteristics in the wild. PMID- 12118717 TI - SSCP-based identification of members within the Pseudoterranova decipiens complex (Nematoda: Ascaridoidea: Anisakidae) using genetic markers in the internal transcribed spacers of ribosomal DNA. AB - The anisakid nematodes morphologically corresponding with Pseudoterranova decipiens sensu lato (s.l.) (Krabbe, 1878) from different seal or sea lion hosts and geographical origins, previously identified as Pseudoterranova krabbei, P. decipiens (s.s.), P. bulbosa, P. azarasi and P. cattani by multilocus enzyme electrophoresis, were characterized using a DNA approach. Also a population of P. decipiens (s.l.) from Chaenocephalus aceratus, the blackfin icefish, from Antarctica and another from Osmerus eperlanus, the European smelt, from Germany were included in the study. The first (ITS-1) and second (ITS-2) internal transcribed spacers (ITS) of ribosomal DNA (rDNA) were amplified by PCR from individual nematodes and analysed by single-strand conformation polymorphism (SSCP), followed by selective sequencing. While no variation in single-stranded ITS-1 and ITS-2 profiles was detected among samples representing each of the species or populations (with the exception of slight microheterogeneity), SSCP analysis of the ITS-2 amplicons allowed the unequivocal differentiation of all of the 5 sibling species of P. decipiens (s.l.) examined, which was supported by sequence differences in ITS rDNA. Samples representing the P. decipiens (s.l.) population from O. eperlanus had the same SSCP profile as those of P. decipiens (s.s.), which was supported by a lack of nucleotide difference in the ITS between them, suggesting that the former represented P. decipiens (s.s.). Based on SSCP results and ITS sequence data, P. decipiens (s.l.) from C. aceratus was genetically most distinct with respect to all other members of Pseudoterranova examined, which indicated that it may represent P. decipiens E (based on geographical origin) or a distinct species. These findings and the molecular approach taken should have important implications for studying the life-cycles, transmission patterns, epidemiology and population genetics of these anisakid nematodes, and the diagnosis of their infections. PMID- 12118719 TI - Spatial and temporal aspects of urban transmission of Echinococcus multilocularis. AB - High prevalences of Echinococcus multilocularis have been reported from foxes of the city of Zurich, Switzerland. In order to characterize transmission in urban areas, a coproantigen ELISA was evaluated for diagnosing the infection in fox faecal samples collected in the environment. In addition, trapped rodents were investigated for the presence of metacestodes. Faecal samples could reliably be classified as being of fox origin by assessing physical properties as shown by the different parasite spectra of putative fox and dog faecal specimens. From the total of 604 tested putative fox faecal samples 156 (25.8%) were positive in the ELISA with a distinct increase in the proportion of positive samples from the urban to the periurban zone. Furthermore, samples collected in the border zone had significantly more coproantigen-positive results during winter. Prevalence of E. multilocularis in rodents was 9.1% (81/889) for Arvicola terrestris (with 3.5% of the animals harbouring between 14 and 244400 protoscoleces) and 2.4% (2/83) for Clethrionomys glareolus. E. multilocularis-infected A. terrestris were found in 9 of 10 trapping sites in the border zone. The high infection pressure in the periphery of urban areas might pose a risk for infection with E. multilocularis for both domestic carnivores as well as for urban inhabitants. Interventions into the cycle aiming at reducing the infection pressure should therefore focus on these areas. PMID- 12118720 TI - Comparison of the structures of natural and re-established populations of Ascaris in humans in a rural community of Jiangxi, China. AB - To compare the structures of natural and re-established populations of Ascaris in humans, universal (mass) chemotherapy was carried out at the beginning and the end of the study year using pyrantel pamoate. Worms expelled within 48 h of treatment were collected, their sex determined, and measurements made of length, width and weight. Length was used as the criterion for estimating the developmental stage of the worms. In comparison with the natural population, the reestablished population displayed similar sex ratio as well as distribution patterns among individuals and age groups of the host. However, the mean worm burden of the re-established population was significantly decreased, with a reduction of burdens in children aged 5-9 years. Also, the re-established population showed significant changes in population structure and worm measurements in that it comprised more immature and less fertile males, less fertile and more senile females, smaller and lighter males, larger (but not heavier) females than the natural population. The results suggested that the reestablished Ascaris population did not restore to its original status in relation to mean density, composition and fecundity. Therefore, universal treatment once a year should decrease the transmission of Ascaris in humans. Combined with previous results for the same study sites, the present findings also indicated that caution is warranted to avoid misleading conclusions when using prevalence and faecal egg counts as parameters for evaluating the success of control programmes. PMID- 12118721 TI - The time-course of the response to the FMRFamide-related peptide PF4 in Ascaris suum muscle cells indicates direct gating of a chloride ion-channel. AB - We investigated the effects of PF4 on Ascaris suum somatic muscle cells using a 2 electrode current-clamp technique. PF4 is a FaRP (FMRFamide-related peptide), originally isolated from the free-living nematode Panagrellus redivivus. PF4 caused hyperpolarization and an increase in chloride ion conductance when it was applied to the muscle cells of the Ascaris body wall. The delay between the application of the peptide and the appearance of the response was measured and compared with that of gamma-amino butyric acid (GABA), a compound that directly gates ion channels, and with PF1, a FaRP that acts via an intracellular signal transduction mechanism. The PF4 and GABA delay times were not significantly different; they were 1.51+/-0.11 sec and 1.22+/-0.10 sec respectively. The delay following application of PF1, 3.75+/-0.51 sec, was significantly longer. The rapid response to PF4 is consistent with direct gating of a chloride ion channel, which has not been described elsewhere in the literature. PMID- 12118723 TI - Chromosome aberrations in peripheral blood lymphocytes in subjects occupationally exposed to ionizing radiation or chemical clastogens. AB - To get an insight into genotoxic risk in some occupations, in this study the chromosome aberration analysis of peripheral blood lymphocytes was made in 20 physicians and nurses exposed to a low dose of ionizing radiation in a hospital, 12 individuals working with X-rays in a cement factory and 19 technicians working with some chemical toxic agents in the laboratories of a medical school. The control group consisted of 14 sex- and age-matched unexposed persons living in the same district area. The data showed that the total number of chromosome aberrations in 200 scored metaphases in all examined groups were almost the same and inside the low-permitted values. In hospital workers, however, the percentage of acentric and dicentric fragments (1.63 +/- 0.28 vs 0.31 +/- 0.21 and 0.47 +/- 0.18 vs 0.0, respectively) increased predominantly in contrast to cement-factory employees and laboratory workers, where a higher incidence of minutes (0.58 +/- 0.19 vs 0.31 +/- 0.2) or gaps (2.21 +/- 0.37 vs 1.15 +/- 1.15) was noticed. Moreover, in groups exposed to low doses of ionizing radiation (hospital and factory), a positive correlation was found between the total number of chromosome aberrations and the 6-year absorption dose or working period, suggesting an effect of cumulative dosage. The data emphasize that the continuous chromosomal aberration analysis should be obligatory for individuals exposed to various genotoxic substances, even in occupational conditions where according to dosimetric analysis they are exposed to permitted levels of radiation. PMID- 12118722 TI - A phylogenetic hypothesis for the distribution of two genotypes of the pig tapeworm Taenia solium worldwide. AB - Genetic polymorphism was determined among 13 isolates of Taenia solium from various regions using PCR-amplified sequences of 2 mitochondrial genes: cytochrome c oxidase subunit 1 and cytochrome b. The 2 phylogenies obtained were similar to each other regardless of the genes examined. The isolates from Asia (China, Thailand, Irian Jaya and India) formed a single cluster, whereas the isolates from Latin America (Mexico, Peru, Ecuador, Bolivia and Brazil) combined with those from Africa (Tanzania, Mozambique and Cameroon) to form an additional cluster. These results and historical data of swine domestication, distribution of pigs and colonization suggest that T. solium was introduced recently into Latin America and Africa from different regions of Europe during the colonial age, which started 500 years ago, and that the tapeworm of another origin independently spread in Asian countries. PMID- 12118724 TI - Renewal of the culture medium induces a temporary decrease in the gap junctional communication accompanied by degradation and re-establishment of gap junctions in the V79-4 Chinese hamster cell line. AB - To replace the culture medium with a fresh one is a routine action of the mammalian cell culture technique. It is generally assumed that this act per se does not cause any significant physiological response of a cell population that would significantly interfere with the experimental procedures in culture. However, in this series of experiments we demonstrate that the exchange of the culture medium for a fresh one may induce a significant temporary decrease in the GJIC, assessed by the dye coupling method in the V79-4 Chinese hamster cell line. This effect is accompanied by a degradation of gap junctions and their re establishment assessed by the semi-quantitative immunocytochemistry of connexin43. The minimum value of GJIC was reached 45 min after the exchange of the medium. Afterwards, GJIC grew up again, reaching the standard value 3 or 4 h later. This effect does not just result from the exchange of medium as a mechanical action, is not caused by the change of pH and is of quantitative character. The fresh medium loses its capability to reduce GJIC after 3 h of conditioning with the same cells. We found that the value of the early inhibition of GJIC observed during the first 2 h of treatment with the inhibitor of GJIC-EG (applied together with a fresh culture medium)--was indistinguishable from the effect of the exchange of medium itself. Only after that point of time is the EG induced inhibition of GJIC definitely distinguishable. The results demonstrate that a simple exchange of the culture medium, which is generally implemented in various experiments in culture, may cause serious physiological, biochemical and even morphological responses of cells and thus affect the final results of experiments in culture, especially regarding the early effects of drugs. Consequently, to avoid an undesirable response of the cell population reported in this paper we recommend to apply or remove drugs using a medium conditioned with the same cells for at least 3 h. PMID- 12118726 TI - A rat linkage map based on BC x LEW intercross. AB - A genetic linkage map consisting of 258 polymorphic loci has been constructed on the basis of an F2 intercross between the BC/CpbU and LEW/OlaHsd inbred rat strains. When compared to previously published maps a discrepancy was found for rat chromosome 7. The map spans a sex-averaged genetic length of 1790 cM and has an average marker spacing of 7.7 cM. It was estimated that this genetic map is linked to about 90% of the DNA in the rat genome. Because LEW/OlaHsd and BC/CpbU strains differ for dietary cholesterol susceptibility and hepatic copper content, the map is considered to be a valuable tool for studying the genetic background of these complex traits. PMID- 12118725 TI - Tumour-inhibitory effects of dendritic cells administered at the site of HPV 16 induced neoplasms. AB - Experiments were designed to examine whether administration of APC at the site of HPV 16-associated tumours can inhibit tumour growth and whether the efficacy of established dendritic cell lines is comparable to that of fresh BMDC populations. Mice were inoculated s.c. with APC, either bone marrow-derived dendritic cells differentiated in medium supplemented with GM-CSF and IL-4 (BMDC), or with established dendritic cell lines DC2.4 or JAWS II. The pretreated mice, together with untreated controls, were challenged with syngeneic HPV 16-transformed cells MK16 at the site of APC administration. It has been found that both BMDC and dendritic cell lines can inhibit tumour growth and that the efficacy of the established dendritic cell lines DC2.4 and JAWS II was comparable to that of fresh BMDC populations. In vitro induction of proliferative spleen cell responses by co-cultivation with MK16 antigen-pulsed BMDC or MK16 antigen-pulsed dendritic cell lines revealed that both types of APC populations can prime immune reactions directed against syngeneic HPV 16-associated neoplasms. Taken together, the results suggest that local increase in the number of dendritic cells at the site of HPV 16-associated tumours can inhibit progression of the tumours and that the dendritic cell lines which are efficient in this respect can be considered and should be tested in both, preclinical and human systems for delivery of therapeutic vaccines against HPV 16-associated neoplasms. PMID- 12118727 TI - Metabolic characterization of insulin resistance syndrome feature loci in three brown Norway-derived congenic strains. AB - Studies on genetic determination of the insulin resistance syndrome in rat models revealed several susceptibility loci for features of this complex phenotype, i.e. dyslipidemia, insulin resistance and obesity. We analysed the influence of introgression of the RNO4, RNO20 segments of SHR origin and RNO8 segment of PD/Cub origin (all previously shown to be involved in (dys)regulation of carbohydrate and lipid metabolism) onto the genetic background of a common progenitor, the Brown Norway (BN/Cub) rat. The differential segments were genetically characterized in the BN.PD-D8Rat39/D8Rat35 (BN-Lx, RNO8 congenic), BN.SHR-Il6/Cd36 (BN.SHR4, RNO4 congenic) and BN.PD-D8Rat39/D8Rat3, SHR D4Mgh2/Cd36,SHR-D20Wox3/D20Mgh5 (BN-Lx 1K, RNO4, 8, 20 triple congenic) strains and their metabolic profiling was performed. After one week of high-sucrose diet, all congenic strains showed substantially higher levels of serum triglycerides and free fatty acids as well as impaired glucose tolerance in comparison with the BN/Cub progenitor strain. The BN-Lx 1K triple congenic strain displayed the most profound dyslipidemia, glucose intolerance and highest increase of triglyceridemia in response to high-sucrose diet overall, though accompanied with the significantly lowest adiposity index. These results further support the role of genes present within the studied chromosomal regions in observed metabolic disturbances. Furthermore, these findings point to the studied loci within the gene-gene and gene-environment interactions involved in pathogenesis of the insulin resistance syndrome. The set of defined congenic strains provides a possibility of assessing individual features of such a complex phenotype. PMID- 12118728 TI - Modulatory effects of long-chain n-3 fatty acids on cell functions. AB - In recent years, it has been demonstrated that certain fatty acids are involved in the modulation of immune system functions. The mechanisms responsible for these effects are not fully elucidated, but many hypotheses have described numerous changes in the cell functionality as the main factors capable of altering the immune functions. In the present investigation, we have analysed the potential effects of FFA on cell viability, production of superoxide radicals or proteasome activity in assays in vitro. Thus, different FFA, such as OA, EPA or SA have been incorporated to cellular cultures at a concentration of 100 microM. Phospholipase, cyclooxygenase or lipooxygenase inhibitors abolished the loss of thymocyte viability exerted by EPA, the most immunosuppressive fatty acid. Similarly, measurement of the oxidative process by NBT reduction in cells treated with EPA was markedly increased. Nevertheless, the proteasome activity as a mechanism that participates in T-cell activation was not modified by direct action of the different fatty acids on the in vitro cultures. Overall, these results underline the differential role of several fatty acids (particularly long chain n-3 polyunsaturated fatty acids) in order to modulate many functions of the immune system. PMID- 12118729 TI - Characterization of functional domains of human interferon gamma by specific monoclonal antibodies. AB - For the study of functional domains of hIFNgamma, two MABs were characterized. Both MABs, named 1E11 and 5D6, were sequence-specific for hIFNgamma. According to the estimated additivity index, they recognized epitopes located at distinct non overlapping areas of the hIFNgamma molecule. When pre-incubated with hIFNgamma, MAB 1E11 was able to neutralize the antiviral as well as the antiproliferative activity of the cytokine. This indicated that MAB 1E11 was specific either for the domain responsible for binding to the cell receptor or for a domain required for both functions. By contrast, the second MAB 5D6 did not interfere with any of the two hIFNgamma biological activities. The epitope of MAB 5D6 was located between the amino acid residues Leu 135 and Glu 143 by using different forms of C terminally truncated hIFNgamma. These data allow the conclusion that the last nine C-terminal amino acids are not essential for the receptor binding and biological functioning of this cytokine. The possible role of hIFNgamma C terminus in the intracellular cascade of events is discussed. PMID- 12118730 TI - Measuring the outputs of nursing research and development in Australia: the researchers. AB - It is vital for nurses to publish in order to provide evidence of their practice and to increase the knowledge base of their discipline. This paper is one of two that reports on an investigation of the nursing research published by Australian authors from 1995-2000 in 11 nursing journals based in Australia, the UK and the USA. The focus of this article is on the researchers drawn from a total of 509 articles that were content analysed and categorised according to topics of research, paradigm, methods used and funding acknowledgment. The researchers were analysed on the basis of gender, discipline, employment base and location. Publications had from one to 10 authors, averaging two, with 26 authors claiming 23.6% of research articles. The most common discipline area was nursing and universities were the leading area of employment. Authorship was not limited to capital cities reflecting the spread of university campuses in rural areas. Research papers made up 12.5% of possible articles, supporting the notion that few nurses publish research papers in the refereed general nursing journals we focused on. PMID- 12118731 TI - Limit setting: a useful strategy in rehabilitation. AB - Limit setting is a concept familiar to most mental health clinicians, but much less familiar to staff not specifically trained in mental health care. This paper presents guidelines developed for rehabilitation staff on the strategy of limit setting. The aim of these guidelines was to provide a starting point for ongoing education on limit setting and behavioural management for staff working in a non psychiatric rehabilitation environment. Limit setting is presented, not only as a response to challenging behaviour, but also as fundamental to all patient care within the rehabilitation context. The guidelines draw on the concepts of limit setting, acting out, therapeutic relationships and therapeutic milieu as described in the psychiatric literature. A humanistic framework for helping people underpins the guidelines. Principles for selecting and enforcing limits are described. Finally, a list of clarification prompts is provided for clinicians to use when faced with challenging patient behaviour. PMID- 12118732 TI - Home parenteral nutrition: an ethical decision making dilemma. AB - Ethics is a hot topic these days. Home health care providers need not be ethicists, however they do need to be able to identify problems quickly, and know how to address them. This paper explores the ethical issues arising from a narrative analysis involving an advanced cancer patient receiving Total Parenteral Nutrition (TPN) at home. It shows how complicated it is today to make nutrition support decisions that would have been customary less than 30 years ago. For and against arguments of TPN for advanced cancer patients are reviewed. Ethical positions adopted by the medical and nursing professions are explored and contrasted. The importance of patient autonomy, within a holistic notion of care, including decisions incorporating quality of life, are affirmed, providing a challenge to monitoring the status quo in approaches to decision making. PMID- 12118733 TI - Internet-based education for enrolled nurses: could it be e-ffective? AB - There has been an extraordinary growth of technology-mediated learning in the higher education system over the last 10 years (IHEP 2000, p.1), predominantly in the area of distance education. These technological advances provide exciting opportunities for the delivery of education to those who have not previously been able to access on-campus learning, usually because of barriers of distance and cost. While there are a growing number of studies supporting the implementation of distance education programs in nursing education, the use of Internet-based learning as a sole means of education delivery is relatively new. This paper investigates the current literature available regarding the use of Internet-based education delivery for registered and enrolled nurses in undergraduate and postgraduate programs, and reviews contemporary education trends in Victoria, Australia, for enrolled nurses (ENs). The challenge in nursing education currently is to design curricula that will address the health care needs of the future. This is not easy with the rapidly changing environment health care professionals, especially nurses, face in their daily work. To prepare graduates who can function successfully as professional nurses in this new century, nurse educators must examine the dominant trends in health care and education, and analyse whether the processes used to prepare students for practice will result in the desired outcome (Jorgensen et al 1998, p.109). The current growth and impact of web-based and online learning courses has been proposed by some as a major revolution for education, and has been firmly embraced by many tertiary institutions as the way of the future (Sims 1998, p. 21). PMID- 12118734 TI - The participation of volunteers in contemporary palliative care. AB - Historically, in Australia, individuals with widely differing interests, skills and values have engaged collaboratively, in a voluntary capacity, to establish services to assist persons experiencing particular need or hardship. Gradual recognition and acceptance by the State of its social responsibilities to citizens with various needs in areas of health, welfare, education and others, have seen the provision of a range of statutory services available to all Australians. Volunteer participation in the delivery of modern health services, therefore, is not usual; palliative care is an exception rather than a norm. This article explores the relationship between understandings of death and dying in Western culture and the participation of volunteers in contemporary palliative care. The author presents a view that volunteers provide a distinctive contribution to the quality of care delivery and to enrichment of the social environment of the wider community also. The topic is of relevance to all nurses and especially those involved in the care of dying persons and of their families. PMID- 12118735 TI - Exploring nursing leadership. PMID- 12118736 TI - Into the twenty-first century--what future for nursing? PMID- 12118737 TI - The experiences of adults with cerebral palsy during periods of hospitalisation. AB - People with cerebral palsy may have a range of disabilities that can result in daily dependence on others to meet some or all of their basic and more complex care needs. The aim of this New South Wales research was to examine the experiences of adults with cerebral palsy during inpatient admission to a number of public hospitals. A self-selected sample of 31 adults with cerebral palsy completed a questionnaire that collected information related to their disability specific needs and how well these were addressed by nursing staff during admissions to hospitals. Analysis of the data revealed that many respondents felt hospital staff had limited knowledge and skills of caring for people with cerebral palsy, resulting in their basic care needs not being adequately addressed during periods of hospitalisation. Changes in nursing assessment, continuing education and discharge planning are recommended to address these issues. PMID- 12118738 TI - Revascularization during acute myocardial infarction: risks and benefits revisited. AB - BACKGROUND: Indication for immediate revascularization during acute myocardial infarction (MI) is debated. Drug-resistant crescendo angina, as well as hemodynamic compromise, however, often requires acute operation. In this study the differential risks of acute coronary artery bypass grafting with and without MI were stratified. METHODS: Five hundred eighteen patients undergoing isolated coronary artery bypass grafting were investigated. Thirty-nine patients underwent acute revascularization because of enzyme-proven or electrocardiogram-proven MI accompanied by crescendo angina, hemodynamic compromise, or both. They were compared with 33 emergent, 63 urgent, and 383 elective patients without MI. Preoperative risk factors for early mortality and necessity of continuous venovenous hemofiltration were analyzed by means of logistical regression analysis. Perioperative data were compared. RESULTS: Early mortality of the MI cohort was 15.4%, in contrast to 15.2% in emergent, none in urgent, and 2.1% in elective patients. Left internal thoracic artery was used in 87% of MI, 97% of emergent, 94% of urgent, and 97% of elective patients. Intraaortic balloon pump was necessary in 50% of MI patients, 27% of emergent, 6.3% of urgent, and 3.1% of elective cases. Continuous venovenous hemofiltration was performed in 29% of MI patients, 15% of emergent, 4.9% of urgent, and 3.4% of elective patients. Hemodynamic instability significantly increased the odds ratio for early mortality and continuous venovenous hemofiltration. CONCLUSIONS: Patients undergoing acute revascularization carried an elevated risk to die early notwithstanding the presence or absence of acute MI. Liberal use of left internal thoracic artery grafts was not detrimental in acute patients whereas liberal use of intraaortic balloon pump was beneficial. In almost 30% of MI patients, continuous venovenous hemofiltration was not necessary, implying a severely impaired perioperative hemodynamic condition. Immediate revascularization in the presence of acute MI is therefore indicated although it may be addressed as a separate high-risk group. PMID- 12118739 TI - Cerebral oxygen saturation assessed by near-infrared spectroscopy during coronary artery bypass grafting and early postoperative cognitive function. AB - BACKGROUND: Cerebral oxygen saturation (ScO2) can be assessed by near-infrared spectroscopy. We investigated the correlation between early postoperative cognitive performance and intraoperative ScO2 in a prospective observational setting. METHODS: Forty-seven patients undergoing elective coronary artery bypass grafting with cardiopulmonary bypass underwent preoperative and postoperative neuropsychological evaluation. Patients were classified according to the presence or absence of postoperative cognitive dysfunction. Cognitive dysfunction was defined as an individual test score decrease of more than one standard deviation in two or more of the five tests. During operation ScO2 was continuously measured using an INVOS 4100 device. Cerebral oxygen saturation values were analyzed with reference to two cutoff points, which should reflect low cerebral oxygenation: an ScO2 less than 40% and a drop of more than 25% from individual baseline values. The duration and extent of ScO2 values below these two cutoff points was compared between the patients with and without cognitive dysfunction. RESULTS: Sixteen patients (34%) showed postoperative cognitive dysfunction. Cerebral oxygen saturation values less than 40% occurred in 17 patients for a mean (+/- standard error of the mean) of 17.2 +/- 6.5 minutes, whereas a decrease of more than 25% from baseline values occurred in 37 patients for 52.7 +/- 7.8 minutes. The duration and extent below the two cutoff ScO2 values was similar in patients with and without cognitive dysfunction. CONCLUSIONS: Intraoperative regional ScO2 as assessed by near-infrared spectroscopy with the INVOS 4100 device is not predictive for postoperative cognitive performance in patients undergoing coronary artery bypass grafting with cardiopulmonary bypass. PMID- 12118740 TI - Off-pump coronary artery bypass grafting decreases morbidity and mortality in a selected group of high-risk patients. AB - BACKGROUND: The ideal indication for off-pump coronary artery bypass grafting (OPCABG) has yet to be defined. High-risk surgical patients may benefit the most when cardiopulmonary bypass (CPB), aortic cross clamping, and cardioplegic arrest are avoided. The aim of this study was to determine whether off-pump coronary artery bypass grafting might decrease the operative morbidity and mortality in a select group of high-risk patients with multivessel coronary artery disease. METHODS: Utilizing a Parsonnet risk stratification model we analyzed prospectively collected data on a cohort of high-risk coronary artery disease patients, which were operated on with beating-heart technology by the same group of surgeons in a tertiary care university medical center. High-risk patients were defined as those with a Parsonnet score of 15 or greater. RESULTS: Fifty-seven multivessel disease OPCABG patients (over a period of 2 years) had markedly increased Parsonnet scores (24.3 +/- 10.6). The average ejection fraction of the patients was 42% (+/-12.3) and their age ranged from 52 to 85 years (mean 70.6 +/ 10.4, 26% women). Unstable angina was present in 42 patients (74%) and 10 patients underwent OPCABG within 24 hours of the occurrence of acute myocardial infarction. In addition to severe coronary artery disease 32% of the patients presented with congestive heart failure, insulin-dependent diabetes (18%), renal failure (22%), peripheral vascular disease (31%), pulmonary disease (18%), and neurologic disorders (14%). An average of 2.6 +/- 0.9 grafts/patient were performed and the posterior descending artery or marginal branches of the circumflex artery or both were grafted in 90%. The 30-day mortality rate was 3.5% (n = 2). CONCLUSIONS: OPCABG can be performed with a reasonable low morbidity and mortality in this select group of high-risk patients. OPCABG is a reasonable, and might even be preferable, operative strategy in this high-risk group of patients. PMID- 12118741 TI - Severity of coronary artery stenosis at preoperative angiography and midterm mammary graft status. AB - BACKGROUND: The purpose of this study was to evaluate the correlation between the midterm angiographic results of mammary artery grafts and the preoperative stenosis of the target vessel. METHODS: We analyzed preoperative and postoperative angiograms of 93 patients who underwent postoperative midterm (> or = 3 years) angiograms of an internal mammary artery (IMA) to left anterior descending artery graft for clinical or study purposes. Patients were divided into three groups on the basis of the percentage of the coronary artery stenosis at preoperative angiography: < 70%, 70% to 90%, and > 90% stenosis. RESULTS: Preoperative characteristics were similar in the three groups. The overall incidence of IMA occlusion was 19% in the entire population, without significant differences between groups (19% versus 29% versus 14%). The mean mammary artery diameter significantly increased in direct proportion to the severity of the coronary stenosis (2.0 +/- 0.2 mm in the < 70% versus 2.5 +/- 0.3 mm in the 70% to 90% and 2.7 +/- 0.4 mm in the > 90% series; p < 0.05). CONCLUSIONS: Chronic native competitive flow does not significantly affect midterm graft status but does influence mammary graft diameter. PMID- 12118742 TI - Konno aortoventriculoplasty in children and adolescents: from prosthetic valves to the Ross operation. AB - BACKGROUND: In children with complex multilevel left ventricular outflow tract obstruction (LVOTO), the Konno aortoventriculoplasty is used to enlarge the aortic root and increase the size of the aortic valve implanted. We present our experience with the evolution of this surgical approach from prosthetic valves to the Ross operation. METHODS: Between March 1982 and July 2000, 60 patients (36 males and 24 females) had 72 Konno aortoventriculoplasties (prosthetic valve and Konno group [57 patients; redo, 12] and Ross-Konno group [15 patients]). The patients' ages ranged from 5 days to 18 years (mean, 8.2 years). The underlying anatomic diagnoses were congenital aortic stenosis and LVOTO in 51 patients, coarctation of the aorta with bicuspid aortic valve in 13, severe aortic insufficiency associated with a ventricular septal defect in 8, interrupted aortic arch in 6, mitral stenosis in 6, atrioventricular septal defect in 5, and endocarditis in 3. There was no statistical difference in age or diagnosis between the two groups. The prosthetic valve group included 42 mechanical valves, 9 homografts, and 6 xenografts. RESULTS: Follow-up ranged from 10 months to 18.5 years (mean, 9.7 years) in the prosthetic valve and Konno group compared with 5 months to 3.7 years (mean, 2.1 years) in the Ross-Konno group (p < 0.05). There were 10 early deaths and four late deaths in the prosthetic valve and Konno group as compared with one early death and two late deaths in the Ross-Konno group (p = not significant). Reoperations for LVOTO and aortic valve replacement were significantly higher in the prosthetic valve and Konno group as opposed to the Ross-Konno group (16 vs 0, p < 0.05) mainly because of the biological valve and Konno subgroup. There were no statistical differences in reexploration for bleeding, pacemaker insertion, and reoperations for indications other than LVOTO and aortic valve replacement between the two groups. CONCLUSIONS: The Konno aortoventriculoplasty is a good surgical option for complex LVOTO. Patients with a prosthetic valve and Konno carry a high rate of reoperation. Early operative results with the Ross-Konno operation seem promising. PMID- 12118743 TI - Intermediate follow-up of a composite stentless porcine valved conduit of bovine pericardium in the pulmonary circulation. AB - BACKGROUND: In the pediatric population, glutaraldehyde-preserved bovine pericardium conduit containing a stentless porcine valve has been proposed as an alternative to homografts for right ventricular outflow tract reconstruction. METHODS: Between June 1996 and March 2000, a total of 55 patients, 20 with truncus arteriosus, 21 with pulmonary atresia with ventricular septal defect, and 14 with miscellaneous defects, received this conduit. Median age at implantation was 3.4 months (range, 3 days to 19 years), and 27 patients (50%) were less than 3 months old. Clinical outcome, echocardiographic data, and pathologic analysis were recorded. End points for conduit failure were conduit replacement or dilation. A mean follow-up of 27 months (range, 2 to 46 months) was available for 47 survivors. RESULTS: Procedure for conduit obstruction was required in 13 patients. The most common procedure was operation, and all but 3 patients had an unsuccessful balloon angioplasty before reoperation. Actuarial freedom from conduit dilation or reoperation was 93.6% (95% confidence limits, 82% to 99%), 81.9% (95% confidence limits, 64% to 91%), 77.8% (95% confidence limits, 39% to 78%), and 64.3% (95% confidence limits, 26% to 73%) at 1, 2, 3, and 4 postoperative years, respectively. Univariate analysis identified small conduit size as a risk factor for conduit obstruction. CONCLUSIONS: Although this new conduit was not free from progressive obstruction, our clinical results (easy to work and good valvular function) and the availability in small sizes encouraged us to use it as an alternative to small-size homografts when those were not available. PMID- 12118744 TI - Lung and heart-lung transplantation in patients with end-stage cystic fibrosis: the Stanford experience. AB - BACKGROUND: Bilateral lung (BLTx) and heart-lung transplantation have gained wide acceptance as treatment of end-stage lung disease from cystic fibrosis. We reviewed our 13-year experience with thoracic transplantation for cystic fibrosis with an operative approach that favors use of cardiopulmonary bypass for BLTx. METHODS: Sixty-four patients with cystic fibrosis underwent heart-lung transplantation (n = 22, 34.4%) or BLTx (n = 42, 65.6%) between 1988 and 2000. Mean age and weight at transplantation were 29 +/- 8 years and 51 +/- 11 kg, respectively. Mean follow-up for survivors was 4.4 +/- 3.6 years. Immunosuppression regimen included cyclosporine, tapered corticosteroids, azathioprine, and induction therapy with OKT3 (murine monoclonal antibodies) or rabbit antithymocyte globulin. Cardiopulmonary bypass was used in all but 5 patients (7.8%). However, in 8 (19%) of the 42 patients having BLTx, only the grafting of the second lung was performed with cardiopulmonary bypass. RESULTS: The operative mortality rate was 1.6%. The actuarial survival rates at 1 year, 3 years, 5 years and 10 years were 93.2%, 77.7%, 61.8%, and 48.1%, respectively, with no significant difference between BLTx and heart-lung transplantation. The major hospital complications were pneumonia (n = 11, 17.2%) and bleeding (n = 8, 12.5%). Clinically significant reperfusion injury was observed in 6 patients, 3 of whom required reintubation. Freedom from acute lung rejection beyond 1 year was 47.7%. One patient underwent late retransplantation, and 4 required bronchial stenting. Obliterative bronchiolitis accounted for eight (50.0%) of 16 late deaths. CONCLUSIONS: Though postoperative bleeding and pneumonia are still of concern, satisfactory early and intermediate-term results can be expected in patients undergoing BLTx or heart-lung transplantation for cystic fibrosis. Cardiopulmonary bypass can be used for BLTx with no adverse impact on intermediate and long-term outcomes. PMID- 12118745 TI - Total correction of tetralogy of Fallot in the first year of life: late results. AB - BACKGROUND: Correction of tetralogy of Fallot in patients less than 1 year of age offers the advantage of a more normal development; but in the majority of cases exposes the patient to the possibly of a higher mortality with one-stage primary repair, and to the long-term effects of a transannular patch, which is often necessary. METHODS: A retrospective review of total correction of tetralogy of Fallot performed in 63 consecutive patients at less than 1 year of age was made. Risk factors for operative mortality and functional status at follow-up were analyzed. Follow-up was obtained from clinic appointments and telephone questionnaires. RESULTS: The operative mortality was 6%, with three late deaths. Aortic cross-clamp time more than 60 minutes (p = 0.023), cardiopulmonary bypass time more than 90 minutes (p = 0.016), and frequent preoperative respiratory tract infection symptoms (p = 0.008) affected operative survival; whereas age less than 3.0 months or weight less than 6.0 kg did not. Mean follow-up is 11.6 years (+/- 0.6 years, standard error). Actuarial survival is 89% (+/- 4%) and freedom from reoperation is 96% (+/- 4%) at up to 20 years after correction. Eighty-seven percent of patients have normal echocardiographic right ventricular function. Only 4 patients have greater than moderate pulmonary regurgitation by echocardiography. Three of these four patients are asymptomatic. At more than 15 years postoperatively, 88% of patients have good-to-excellent functional status. CONCLUSIONS: Early correction of tetralogy of Fallot at less than 1 year of age can have a low operative mortality and provide excellent asymptomatic long-term survival. PMID- 12118746 TI - Duroflo II heparin bonding does not attenuate cytokine release or improve pulmonary function. AB - BACKGROUND: Comparison of the cytokine generation and leukocyte activation properties of Duroflo II heparin bonded bypass circuit (Baxter Healthcare Corp, Compton, UK) and the conventional cardiopulmonary bypass circuit. Attempt to correlate these to pulmonary dysfunction postoperatively. METHODS: Forty patients undergoing elective, isolated coronary artery bypass grafting were randomly allocated to have either plain extracorporeal circuits (group C) or heparin bonded extracorporeal circuits (group H). Full systemic heparinization was used in all patients. The inflammatory response was assessed by measuring plasma levels of interleukin-6, interleukin-8, interleukin-10, and polymorphonuclear elastase. Gas exchange was assessed by measuring the PaO2/FIO2 ratio. RESULTS: Significant impairment of oxygenation was seen in both groups with the lowest values at the end of the operation before a gradual return to normal during the next 6 hours. There were no differences between the groups in gas exchange or times to extubation. There were significant elevations in all the cytokines, with interleukin-6 levels peaking at 4 hours in group H and 24 hours in group C, before starting to return to normal at 48 hours. The patterns of interleukin-8 and interleukin-10 rise were identical in the two groups. Polymorphonuclear elastase reached a peak at the end of the operation in group H and remained elevated up to 24 hours, whereas levels continued to rise in group C up to 4 hours. There were no significant differences in levels between groups at any time. There were no differences between the groups in blood loss or blood product usage. CONCLUSIONS: Cardiopulmonary bypass induces a systemic inflammatory response with release of cytokines and activation of leukocytes. This correlates with the severe deterioration in pulmonary gas exchange from preoperative levels up to 6 hours postoperatively (p < 0.05). In the presence of systemic heparinization, Duroflo II heparin bondingtf the circuits has minor effects on the pattern of evolution of this inflammatory response. PMID- 12118747 TI - Hypoxic preconditioning in coronary microarteries: role of EDHF and K+ channel openers. AB - BACKGROUND: Hypoxic preconditioning may provide a useful method of myocardial protection in cardiac operations. The present study was designed to investigate the possible mechanisms of preconditioning regarding endothelium-derived hyperpolarizing factor (EDHF) and the effect of a potassium channel opener KRN4884 on the porcine coronary microartery in mimicking hypoxic preconditioning. METHODS: Porcine coronary microartery rings (diameter 200 to 500 microm) studied in a myograph were divided into seven groups: (1) control group; (2) hypoxia reoxygenation group (hypoxia for 60 minutes followed by reoxygenation for 30 minutes); (3) preconditioning group (hypoxia for 5 minutes followed by reoxygenation for 10 minutes before hypoxia reoxygenation); (4) KRN4884 pretreatment group (KRN4884 was added into the myograph chamber 20 minutes before hypoxia reoxygenation); (5) 5-hydroxydecanoate + KRN group (5-hydroxydecanoate was given 20 minutes before KRN4884 pretreatetment); (6) glibenclamide (GBC) + KRN group (GBC was added 20 minutes before KRN4884 pretreatment); and (7) endothelium denuded group (the endothelium was removed). The endothelium-derived hyperpolarizing factor-mediated relaxation to bradykinin was studied in the rings precontracted with U46619 in the presence of N(omega)-nitro-L-arginine and indomethacin. RESULTS: The maximal relaxation induced by bradykinin was reduced in hypoxia reoxygenation (40.7% +/- 2.8% vs 66.9% +/- 2.5% in control, p = 0.000). This reduced relaxation was recovered in either preconditioning (64.6% +/ 4.6%, p = 0.002), or KRN4884 pretreatment (67.1% +/- 3.6%, p = 0.000). The 5 hydroxydecanoate, but not GBC pretreatment abolished the effect of KRN44884 pretreatment (67.1% +/- 3.6% vs 42.9% +/- 3%, p = 0.001). CONCLUSIONS: Hypoxia reoxygenation reduces the relaxation mediated by endothelium-derived hyperpolarizing factor in the coronary microartery. This function can be restored by either hypoxic preconditioning or the K(ATP) channel opener KRN4884, and therefore K(ATP) channel openers may provide similar effect as preconditioning. The mechanism is mainly related to the mitochondrial ATP-sensitive K+ channels. PMID- 12118748 TI - Glove punctures and postoperative skin flora of hands in cardiac surgery. AB - BACKGROUND: Surgical gloves are frequently perforated during operations, including heart operations. This infection risk factor is inadequately studied. METHODS: After preoperative hand disinfection and at the end of 116 heart operations, bacterial samples from hands of surgeons, altogether 800 samples, were taken. Glove punctures were examined with water test. RESULTS: Surgeons changed 70 gloves because of breakage during operations. Additionally, 154 of 400 (39%) gloves had holes in postoperative testing. The breakage rate of gloves increased from 30% in operations shorter than 3 hours to 65% when operations were longer than 5 hours. High bacterial counts of the hands were also more common after prolonged operations. CONCLUSIONS: Glove puncture rates and bacterial counts of hands increase with increasing operation time. We recommend changing of both gloves when a puncture is detected. Before donning new gloves, hands should be disinfected. PMID- 12118749 TI - Subsequent pulmonary resection for bronchogenic carcinoma after pneumonectomy. AB - BACKGROUND: Patients who have undergone a pneumonectomy for bronchogenic carcinoma are at risk of cancer in the contralateral lung. Little information exists regarding the outcome of subsequent lung operation for lung cancer after pneumonectomy. METHODS: The records of all patients who underwent lung resection after pneumonectomy for lung cancer from January 1980 through July 2001 were reviewed. RESULTS: There were 24 patients (18 men and 6 women). Median age was 64 years (range, 43 to 84 years). Median preoperative forced expiratory volume in 1 second was 1.47 L (range, 0.66 to 2.55 L). Subsequent pulmonary resection was performed 2 to 213 months after pneumonectomy (median, 23 months). Wedge excision was performed in 20 patients, segmentectomy in 3, and lobectomy in 1. Diagnosis was a metachronous lung cancer in 14 patients and metastatic lung cancer in 10. Complications occurred in 11 patients (44.0%), and 2 died (operative mortality, 8.3%). Median hospitalization was 7 days (range, 2 to 72 days). Follow-up was complete in all patients and ranged between 6 and 140 months (median, 37 months). Overall 1-, 3-, and 5-year survivals were 87%, 61%, and 40%, respectively. Five year survival of patients undergoing resection for a metachronous lung cancer (50%) was better than the survival of patients who underwent resection for metastatic cancer (14%; p = 0.14). Five-year survival after a solitary wedge excision was 46% compared with 25% after a more extensive resection (p = 0.54). CONCLUSIONS: Limited pulmonary resection of the contralateral lung after pneumonectomy is associated with acceptable morbidity and mortality. Long-term survival is possible, especially in patients with a metachronous cancer. Solitary wedge excision is the treatment of choice. PMID- 12118750 TI - Impact of computed tomography-positron emission tomography fusion in staging patients with thoracic malignancies. AB - BACKGROUND: Positron emission tomography (PET) has been demonstrated to improve staging in patients with thoracic malignancies. This study evaluates the ability of a new imaging technique to improve the spatial resolution and accuracy of PET. METHODS: Patients with known or suspected malignancy (n = 21) who were referred for a dedicated PET scan were also evaluated with a new camera-based PET system, which uniquely allows simultaneous computed tomography (CT) and fusion of the camera-based PET images with the CT images. The dedicated PET scan was obtained 1 hour after intravenous injection of fluorodeoxyglucose. The camera-based PET imaging was fused with the CT images at approximately 2 hours after injection. The camera-based PET and CT-PET fusion images were read independently and blindly by 2 experienced observers and the presence and location of abnormalities was compared with dedicated PET scans. RESULTS: Dedicated PET identified 18 sites in the chest as abnormal. The CT-PET fusion was superior to the camera-based PET alone, concordant with the dedicated PET in 16 of 21 patients compared with 13 of 21 by camera-based PET. The lesions missed by the camera-based PET were less than 1 cm in diameter. Fused CT-PET images provided superior anatomic localization and spatial resolution compared with dedicated PET and camera-based PET. CONCLUSIONS: CT-PET fusion images were more accurate than camera-based PET alone. CT-PET fusion improves the spatial resolution compared with dedicated PET and may improve the availability and efficacy of staging of patients with thoracic malignancies. PMID- 12118751 TI - Surgery as part of combined modality treatment in stage IIIB non-small cell lung cancer. AB - BACKGROUND: The role of surgery after neoadjuvant chemotherapy in patients with stage IIIB non-small cell lung cancer (NSCLC) remains unclear. METHODS: A prospective multicenter trial of neoadjuvant chemotherapy followed by surgery or radiotherapy or both was conducted with 41 patients with stage IIIB NSCLC. End points were toxicity, response, downstaging, complete resectability, and survival. The diagnostic value of repeat mediastinoscopy after neoadjuvant chemotherapy (three courses of gemcitabine/cisplatin) was also studied. RESULTS: Response rate after neoadjuvant chemotherapy was 66% (27 of 41). Fifteen patients underwent repeat mediastinoscopy, which proved to be inadequate in 6 patients. Two repeat mediastinoscopies were false negative. Resection was performed in 18 patients, of which 10 proved to be radical. Hospital mortality was 2.4% (n = 1). Major complications occurred in 6 patients (fistula, empyema, hemorrhage). Histopathologically proven downstaging was seen in 16 patients (39%). Twenty-five patients underwent radiotherapy of whom 14 were diagnosed with stable/progressive disease and 9 with partial/complete response. Median survival for all patients was 15.1 months, for nonresponders 8.4 months and for responders 16.8 months (p = 0.11). Patients with partial/complete response had a mean survival of 21.5 months after resection and 13.0 months after radiotherapy (p = 0.0003). CONCLUSIONS: Radical surgery can be performed in 37% (10 of 27) of the responders resulting in a prolonged survival. Surgery as part of combined modality treatment is feasible in stage IIIB NSCLC. Results of a repeat mediastinoscopy are disappointing and proved to be a not-so-effective restaging tool because of the high number of incomplete procedures and because it yields false negative results. PMID- 12118752 TI - Fluoroscopy-assisted thoracoscopic resection of lung nodules marked with lipiodol. AB - BACKGROUND: To localize small and deeply situated pulmonary nodules under thoracoscopy with roentgenographic fluoroscopy, we developed a marking procedure that uses both lipiodol and colored collagen. METHODS: Pulmonary nodules were marked with 0.4 mL of lipiodol under computed tomography. The visceral pleura near each nodule was marked with 1 mL of colored collagen, ie, a mixture of atelocollagen and methylene blue. Nodules were marked more than 1 day before thoracoscopy. At thoracoscopy, C-arm-shaped roentgenographic fluoroscopy was used to detect the radiopaque nodules. Eighteen nodules in 16 patients were localized by this procedure. The nodules had an average diameter of 7 mm (range: 4 to 10 mm) and were located an average distance of 19 mm (range: 8 to 30 mm) from the pleural surface under computed tomographic measurement. RESULTS: There were no complications from the marking procedure except for pneumothorax in 1 patient who required chest tube drainage for additional marking. All 18 nodules could be easily localized at thoracoscopy. The colored collagen revealed the pleura near the nodules. The lipiodol showed the nodules on the fluoroscopic monitor, which was used to guide the forceps to grasp the nodules. All of the nodules could be resected completely under thoracoscopy without adding minithoracotomy. The pathologic diagnosis was malignant tumor in 9 patients, atypical adenomatous hyperplasia in 3, and benign lesion in 4. CONCLUSIONS: A marking procedure that uses both lipiodol and colored collagen can localize small and deeply situated pulmonary nodules under fluoroscopy and facilitate safe and successful thoracoscopic resection. PMID- 12118753 TI - Serum carcinoembryonic antigen level in surgically resected clinical stage I patients with non-small cell lung cancer. AB - BACKGROUND: There is little general agreement concerning the effectiveness of serum carcinoembryonic antigen (CEA) as a prognostic indicator for non-small cell lung cancer (NSCLC) in clinical stage I patients. We conducted a retrospective study to investigate the relationship between serum CEA level and survival. METHODS: We assessed 297 consecutive patients with clinical stage I NSCLC who underwent surgical resection at Toneyama National Hospital from 1985 to 1998. Serum CEA levels were measured with an enzyme-linked immunosorbent assay kit with the upper limit of normal defined as 7.0 ng/mL based on the 95% specificity level for benign lung disease, in our hospital. RESULTS: There were 56 (19%) patients with serum CEA greater than 7.0 ng/mL. The high CEA group had a median survival time of 50 months and a 5-year survival rate of 49% compared with a 5-year survival rate of 72% (p < 0.0001) for the normal CEA group (n = 241). Patients with postoperatively high CEA levels (n = 15) had the worse prognosis (median survival time 35 months, and 5-year survival 18%) compared with patients whose levels returned to normal (n = 41, median survival time 8.8 months, and 5-year survival 68%; p = 0.01). These differences were also observed in patients with pathologic stage I or II tumors but not in those with pathologic stage III or IV tumors. CONCLUSIONS: Serum CEA level is a useful predictor of survival for patients with clinical stage I NSCLC, and a persistently high CEA level after surgery is an especially strong indicator of a very poor prognosis. PMID- 12118754 TI - Plasma brain natriuretic peptide as a noninvasive marker for efficacy of pulmonary thromboendarterectomy. AB - BACKGROUND: Plasma brain natriuretic peptide (BNP), a cardiac hormone secreted mainly by the cardiac ventricles, has been shown to increase in proportion to the degree of cardiac overload. However, whether plasma BNP may serve as a marker for the efficacy of pulmonary thromboendarterectomy in patients with chronic thromboembolic pulmonary hypertension remains unknown. METHODS: Plasma BNP level was measured in 34 patients with chronic thromboembolic pulmonary hypertension before and 1 month after pulmonary thromboendarterectomy. Right heart catheterization was also performed before and 1 month after the operation. RESULTS: Preoperative plasma BNP level was significantly elevated in patients with chronic thromboembolic pulmonary hypertension compared with control patients (246 +/- 40 vs 13 +/- 2 pg/mL; p < 0.001; n = 34) and was positively correlated with total pulmonary resistance (r = 0.57; p < 0.001). After pulmonary thromboendarterectomy, plasma BNP level in survivors markedly decreased (220 +/- 31 to 54 +/- 9 pg/mL; p < 0.001; n = 32) in association with a reduction of total pulmonary resistance (15.6 +/- 1.0 to 4.5 +/- 0.3 Wood units; p < 0.001). The change in plasma BNP level was closely correlated with that in total pulmonary resistance (r = 0.63; p < 0.001). Importantly, a sustained elevation of plasma BNP (> or = 50 pg/mL) indicated the presence of residual pulmonary hypertension (> or = 5 Wood units) after operation (sensitivity = 73%; specificity = 81%). CONCLUSIONS: Plasma BNP level was strongly associated with the severity of pulmonary hypertension in patients with chronic thromboembolic pulmonary hypertension and thereby may serve as a noninvasive marker for the efficacy of pulmonary thromboendarterectomy. PMID- 12118755 TI - Surgical considerations for pulmonary actinomycosis. AB - BACKGROUND: Diagnosis and treatment of pulmonary actinomycosis is difficult without surgical intervention. METHODS: Thirteen patients (10 men, 3 women; mean age, 62 years) underwent pulmonary resection and were given a pathologic diagnosis of pulmonary actinomycosis at our institution between 1976 and 2001. To clarify when pulmonary actinomycosis should be suspected in patients and the role of surgical intervention, we reviewed preoperative clinical characteristics, computed tomography findings, surgical indication, operative procedure, postoperative clinical course, and outcome. RESULTS: Ten patients (77%) had poor oral hygiene. Twelve patients (92%) were symptomatic, and 10 patients (77%) had hemoptysis. The mean interval between radiographic identification of the abnormality and surgical intervention was 8 months (interquartile range, 3.25 to 8 months). Computed tomography findings in all cases included radiologic opacity with air bronchogram or a low attenuation area. Lung cancer was diagnosed initially because of computed tomography findings of spiculation or pleural indentation, and operation was required in 8 patients (62%). The others were diagnosed with chronic pneumonia, and surgical intervention became necessary because of recurrent hemoptysis or prolonged illness. Six patients underwent lobectomy; the others underwent partial resection or segmentectomy. Neither complication nor recurrence has occurred. CONCLUSIONS: When patients, particularly those with poor oral hygiene, show radiologic opacity with an air bronchogram or low attenuation area on the computed tomography scan, pulmonary actinomycosis should be considered and penicillin should be administered as diagnostic therapy. Surgical intervention may be necessary when frequent hemoptysis has no resolution or lung neoplasm cannot be ruled out. PMID- 12118756 TI - Xenotransplant cardiac chimera: immune tolerance of adult stem cells. AB - BACKGROUND: Bone marrow stromal cells have been shown to engraft into xenogeneic fetal recipients. In view of the potential clinical utility as an alternative source for cellular and gene therapies, we studied the fate of xenogeneic marrow stromal cells after their systemic transplantation into fully immunocompetent adult recipients without immunosuppression. METHODS: Bone marrow stromal cells were isolated from C57B1/6 mice and retrovirally transduced with LacZ reporter gene for cell labeling. We then injected 6 x 10(6) labeled cells into immunocompetent adult Lewis rats. One week later, the recipient animals underwent coronary artery ligation and were sacrificed at various time points ranging from 1 day to 12 weeks after ligation. Hearts, blood, and bone marrow samples were collected for histologic and immunohistochemical studies. RESULTS: Labeled mice cells engrafted into the bone marrow cavities of the recipient rats for at least 13 weeks after transplantation without any immunosuppression. On the other hand, circulating mice cells were positive only for the animals with 1-day-old myocardial infarction. At various time points, numerous mice cells could be found in the infarcted myocardium that were not seen before coronary ligation. Some of these cells subsequently showed positive staining for cardiomyocyte specific proteins, while other labeled cells participated in angiogenesis in the infarcted area. CONCLUSIONS: The marrow stromal cells are adult stem cells with unique immunologic tolerance allowing their engraftment into a xenogeneic environment, while preserving their ability to be recruited to an injured myocardium by way of the bloodstream and to undergo differentiation to form a stable cardiac chimera. PMID- 12118757 TI - Surgical treatment of pulmonary hydatid cysts: is capitonnage necessary? AB - BACKGROUND: Hydatid disease of the lung caused by Echinococcus granulosus is frequently encountered in Mediterranean countries. The ideal surgical method for treating this disease is still unknown. METHODS: Between 1994 and 2001, 71 patients with pulmonary hydatid cysts were treated surgically. There were 41 male and 30 female patients with a mean age of 30.2 years (range, 5 to 70 years). Cystotomy and closure of bronchial openings were performed in all patients. Obliteration of the residual cavity by imbricating sutures from within (capitonnage) was achieved in 39 patients (group 1). There were 34 patients with intact cysts and 37 patients with at least one complicated cyst. The average diameter of the cysts was 6.4 cm, and the mean number of cysts per patient was 1.4. The surgical outcome was assessed in group 1 patients and in patients who had undergone closure of bronchial openings without capitonnage (group 2; n = 32). The groups were comparable in regard to clinical characteristics. RESULTS: There was no mortality. The total hospitalization time (mean +/- standard error of the mean) was 5.0 +/- 5.0 days for group 1 and 5.9 +/- 6.9 days for group 2 (p = 0.91). Stay in the intensive care unit was 1.64 +/- 1.22 days in group 1 and 1.60 +/- 1.52 days in group 2 (p = 0.90). The duration of air leak was 2.56 +/- 4.73 days in group 1 and 2.38 +/- 4.74 days in group 2 (p = 0.87). There was no significant difference between groups in the development of empyema (1 patient in group 2 only) and prolonged air leak (5 patients in group 1 and 4 in group 2). There was also no significant difference in the rate of recurrence (3 patients in group 1 only). CONCLUSIONS: We conclude that capitonnage provides no advantage in operations for pulmonary hydatid cysts. PMID- 12118759 TI - Thymectomy by partial sternotomy for the treatment of myasthenia gravis. AB - BACKGROUND: Myasthenia gravis is an autoimmune disease characterized by weakness and fatigue of voluntary muscles. Surgical treatment of choice for myasthenia gravis has been thymectomy. However, thymectomy indications and surgical approach are still controversial. The purpose of this study is to evaluate the efficacy of partial median sternotomy approach to the thymus. METHODS: From 1973 to 1999, 478 patients with myasthenia gravis underwent thymectomy through a partial median sternotomy. RESULTS: Sixty-one patients (12.7%) had complete remission of symptoms, 299 (62.5%) had a significant improvement, and 83 (17.4%), a mild improvement; whereas 35 patients (7.4%) had no improvement of clinical symptoms. CONCLUSIONS: Partial median sternotomy has shown to be a useful surgical approach to the thymus, as demonstrated by the good functional and aesthetic results, associated with low morbidity and no mortality. PMID- 12118758 TI - Sputum retention after lung operation: prospective, randomized trial shows superiority of prophylactic minitracheostomy in high-risk patients. AB - BACKGROUND: Sputum retention after lung operation is a potentially life threatening condition. The minitracheostomy (Minitrach II, SIMS Portex, Hythe, Kent, UK) is a 4-mm percutaneous cricothyroidotomy device, which allows immediate and repeated aspiration of the tracheobronchial tree by minimally trained staff, and can effectively treat sputum retention. This trial was designed to test the hypothesis that prophylactic minitracheostomy could prevent sputum retention in a high-risk group. METHODS: Between March 1997 and October 1999, 102 patients undergoing lung procedures and considered to be at high risk were prospectively randomized to postoperative, prophylactic minitracheostomy insertion in the recovery room with regular aspiration, or to standard postoperative respiratory therapy. RESULTS: Sputum retention developed in 15 patients (30%) in the standard group (n = 52) compared to 1 patient (2%) in the minitracheostomy group (n = 50) (p < 0.005). There were three deaths related to sputum retention in the standard group compared to none in minitracheostomy group during the perioperative period. CONCLUSIONS: It is possible to identify a group of patients at high risk for sputum retention who will benefit from prophylactic therapy. Minitracheostomy is effective as prophylaxis and treatment. PMID- 12118760 TI - Reconstruction of diaphragm using autologous fascia lata: an experimental study in dogs. AB - BACKGROUND: We investigated whether fascia lata is an appropriate material for reconstruction of the diaphragm. METHODS: A diaphragmatic defect (2 cm by 5 cm) was reconstructed with a patch of autologous fascia lata in the experimental group (n = 12) and with expanded polytetrafluoroethylene in the control group (n = 12). Maximal tensile strength at the sutured region was measured serially. RESULTS: The maximal tensile strength at the sutured region reconstructed with the fascia lata was 1.14 +/- 0.50 kgf 15 days and 2.04 +/- 0.94 kgf 30 days after operation. The values were higher than those of expanded polytetrafluoroethylene (p < 0.0001). These values of fascia lata were close to the original maximal tensile strength of the muscular region of the diaphragm (1.52 to 1.66 kgf). CONCLUSIONS: Reconstruction of diaphragm using autologous fascia lata is safe, easy, and inexpensive, and provides smooth wound healing. The only disadvantage is the necessity of a femoral incision for harvest; nevertheless, it may be worthwhile to use fascia lata in clinical trials to further assess its suitability as a reconstruction material. PMID- 12118761 TI - Early and late mortality after pleurodesis for malignant pleural effusion. AB - BACKGROUND: The purpose of this study is to analyze morbidity and mortality and to determine the relative contribution of each of these potential prognosis variables for predicting morbidity and mortality in patients after pleurodesis by thoracotomy or thoracoscopy. METHODS: Between March 1, 1996, and January 31, 2001, a total of 70 patients underwent pleurodesis for recurrent malignant pleural effusion. Thoracoscopy was performed in 54 patients (77%); pleurodesis was achieved by pleural abrasion (n = 15), pleurectomy (n = 5), and talc insufflation (n = 34). Thoracotomy was performed in 16 patients (23%) who also needed pleurectomy and decortication for a trapped lung. RESULTS: Postoperative complications occurred in 24 patients (34%). Factors adversely affecting morbidity with univariate analysis included: three or four metastatic sites (p = 0.003), and thoracotomy (p = 0.009). Factors adversely affecting morbidity with multivariate analysis included: thoracotomy (p = 0.0005) and number of metastatic sites (p = 0.007). Six patient deaths (8.6%) occurred during hospitalization. Factors adversely affecting in-hospital mortality with univariate analysis included: Eastern Cooperative Oncology Group Performance Status 2 to 3 (p = 0.001), lower preoperative serum hemoglobin (p = 0.001), lower preoperative serum albumin (p = 0.0001), and thoracotomy (p = 0.03). Factors adversely affecting in hospital mortality with multivariate analysis included: preoperative serum albumin less than 60 g/L (p = 0.007) and ECOG Performance Status 2 to 3 (p = 0.008). Twelve patients (17%) died within 90 days after surgery. Factors adversely affecting 3-month mortality with univariate analysis included: ECOG Performance Status 2 to 3 (p = 0.001), lower preoperative serum hemoglobin (p = 0.03), higher preoperative white cells (p = 0.03), lower preoperative serum albumin (p = 0.03), and preoperative thoracentesis more than once per month (p = 0.03). Factors adversely affecting 3-month mortality with multivariate analysis included: ECOG Performance Status 2 to 3 (p = 0.01), preoperative thoracentesis more than once per month (p = 0.03), three or four metastatic sites (p = 0.02), and preoperative white blood cell count > or = 12,000/mm3 (p = 0.03). CONCLUSIONS: Thoracotomy is not indicated in patients with a malignant effusion because of poor survival, a high frequency of complications, and prolonged hospital stay. Pleurodesis thoracoscopy is indicated in patients with good performance status coupled with good nutrition. PMID- 12118762 TI - Long-term follow-up of thoracoscopic pleurodesis for hydrothorax complicating peritoneal dialysis. AB - BACKGROUND: Massive hydrothorax is a significant complication of continuous ambulatory peritoneal dialysis (CAPD) and its ideal management remains undefined. Conservative management in the form of intermittent peritoneal dialysis had limited success. The use of conventional pleurodesis and open thoracotomy were associated with morbidities and limitations. We retrospectively reviewed the long term outcome of 8 patients with massive hydrothorax complicating CAPD, 6 of whom received thoracoscopic pleurodesis. METHODS: Among 397 patients undergoing continuous ambulatory peritoneal dialysis during the period from 1994 to 1998, hydrothorax developed in 8 patients. Four patients were first treated with temporary intermittent peritoneal dialysis using 1-L exchange cycles. Three of them had a recurrence of the hydrothorax whereas only one could resume continuous ambulatory peritoneal dialysis successfully. Two patients then underwent conventional pleurodesis but failed. One of them was switched to hemodialysis. Thoracoscopic pleurodesis was performed for the remaining 2 patients together with 4 other patients with hydrothorax once this complication developed. There were no gross abnormalities including pleuroperitoneal communication sites identified. Talc poudrage was performed in 2 patients and mechanical rub pleurodesis in the other 4 patients. All had uncomplicated procedure and uneventful recovery. RESULTS: One patient after thoracoscopic pleurodesis was soon switched to hemodialysis for an unrelated reason. The other 5 patients resumed continuous ambulatory peritoneal dialysis with no recurrence of hydrothorax for a mean period of 50 months (range 19 to 84). CONCLUSIONS: With thoracoscopic pleurodesis, patients resumed continuous ambulatory peritoneal dialysis without recurrence of hydrothorax on long-term follow-up. PMID- 12118763 TI - Expression of heat shock protein 70 in grossly resected esophageal squamous cell carcinoma. AB - BACKGROUND: The aim of the present study was to use immunohistochemical methods to clarify the clinical implication of heat shock protein (HSP) 70 expression in esophageal squamous cell carcinoma and to investigate the function of HSP70 as a chaperone for p53. METHODS: Seventy-one patients with esophageal squamous cell carcinoma were admitted in the present study. Expression of HSP70 was analyzed by immunohistochemistry and correlated with TNM classification, vessel invasion, p53 expression, and clinical outcome after operation. RESULTS: Overexpression of HSP70 was related to sex (p < 0.05), tumor configuration (p < 0.05), lymph node metastasis (p < 0.01), and lymphatic vessel invasion (p < 0.05). Expression of p53 and HSP70 were not correlated with each other (p = 0.824). Esophageal squamous cell carcinoma with HSP70 expression exhibited a significantly better prognosis compared with HSP70-negative esophageal squamous cell carcinoma in univariate analysis (p < 0.05), but no significance was found in multivariate analysis. CONCLUSIONS: We suggest that HSP70 expression might be of use to assess the progression, lymph node metastasis, and lymphatic vessel invasion of esophageal squamous cell carcinoma. Inasmuch as both lymph node metastasis and HSP70 expression are prognostic variables in esophageal squamous cell carcinoma, examination of HSP70 expression may be of use to assess clinical outcome after operation. PMID- 12118764 TI - Postesophagectomy morbidity, mortality, and length of hospital stay after preoperative chemoradiation therapy. AB - BACKGROUND: Data suggest that preoperative chemoradiation improves survival in patients with stage II and III esophageal tumors. Whether preoperative therapy increases postesophagectomy morbidity and mortality has not been determined. This study evaluates our postoperative results after chemoradiation therapy. METHODS: From 1989 through 1998, 120 consecutive patients underwent chemoradiation therapy followed by esophagectomy at our institution. The medical records for these patients were reviewed to determine patient age, sex, race, cell type, operative technique, complications, deaths, and length of hospital stay (LOS). RESULTS: There were 106 (88%) men and 14 (12%) women with a mean age of 58 (32 to 77) years. White patients predominated (114 of 120, 95%); 98 (82%) had adenocarcinoma and 22 (18%) had squamous cell carcinoma. Operative technique was transhiatal in 91 (76%) patients, three-incision in 23 (19%), Ivor-Lewis in 4 (3%), and thoracoabdominal in 2 (2%). There was 1 death. Complications developed in 44 (37%) patients; 59% (13 of 22) of squamous cell carcinoma patients and 32% (31 of 98) of adenocarcinoma patients developed complications. Respiratory complications occurred in 32% (7 of 22) of squamous cell carcinoma patients and in 3% (3 of 98) of adenocarcinoma patients. Mean length of stay after surgery was 15 days (range 7 to 163). CONCLUSIONS: Postesophagectomy results after chemoradiation therapy are comparable to those reported after esophagectomy alone. Squamous cell carcinoma patients are nearly twice as likely to develop postoperative complications and are more likely to have respiratory complications than adenocarcinoma patients. PMID- 12118765 TI - LVAD support in patients with bioprosthetic valves. AB - The presence of mechanical or bioprosthetic valves has traditionally excluded patients from mechanical circulatory support. However, several centers have now developed algorithms for the surgical management of native or prosthetic valve disease in patients requiring left ventricular assist device insertion. We report adverse events associated with bioprosthetic valves in the mitral and tricuspid positions in 2 patients who received long-term mechanical support. We recommend anticoagulation for all patients with prosthetic valves in the mitral or tricuspid position to avoid thromboembolism, inflow conduit occlusion, or valvular incompetence. PMID- 12118766 TI - Thoratec left ventricular assist device for bridging to recovery in fulminant acute myocarditis. AB - Fulminant acute myocarditis can be the cause of rapid cardiac decompensation that is resistant to maximal medical therapy. Successful weaning from left ventricular mechanical support is very rare in fulminant myocarditis. We report the case of a young patient with viral myocarditis who was successfully weaned from a Thoratec left ventricular assist device with full recovery of myocardial function. PMID- 12118767 TI - Traumatic coronary artery dissection. AB - A 14-year-old boy sustained blunt chest trauma resulting in dissection of the left main coronary artery, postinfarction left ventricular aneurysm, mitral regurgitation, and tricuspid regurgitation. He underwent pericardial patch angioplasty of the left main coronary artery, left ventricular aneurysmectomy, mitral valvuloplasty, and tricuspid annuloplasty. The patient continues to do well 4 years after operation. PMID- 12118768 TI - Combined endovascular and open repair of a penetrating innominate artery and tracheal injury. AB - Endovascular therapy affords the opportunity to decrease surgical morbidity and improve operative planning in complex penetrating injuries of the chest. In this case report we describe a hemodynamically stable patient with a single gunshot wound to the base of the neck (zone I), with combined vascular and tracheal injuries. We present a novel approach to the repair of this type of injury using combined endovascular and open techniques. PMID- 12118769 TI - Pericarditis after trauma resulting in delayed cardiac tamponade. AB - Pericarditis complicating cardiac trauma and resulting in tamponade is uncommon. Possible causes include an autoimmune reaction or an inflammatory response to blood entering the pericardium. We present two patients, one with effusive and one with constrictive pericarditis occurring within 2 weeks of a penetrating trauma close to but not directly involving the heart. These cases illustrate the importance of clinical suspicion and aggressive management in the diagnosis and management of such patients. PMID- 12118770 TI - Pyopneumopericardium caused by mediastinal granuloma. AB - We report the case of a previously healthy 32-year-old man who was seen with flulike symptoms, dyspnea, and chest pain. The diagnosis was pyopneumopericardium, and pericardial tap revealed 1.3 L of purulent material. Computed tomography of the chest demonstrated a calcified mass inferior to the carina. Urgent exploration through a right thoracotomy revealed that the mass was adherent to the esophagus and pericardium. The subcarinal mass was resected. Pathological study demonstrated granulomatous lymph nodes, which were likely due to histoplasmosis. This is among the first reports of granulomatous erosion into the pericardium causing pyopneumopericardium. The patient made a good recovery, and his case demonstrates the importance of early imaging and mediastinal exploration for pyopneumopericardium. PMID- 12118771 TI - Homograft aortic root replacement during pregnancy. AB - Operative cardiac interventions have been performed on pregnant women with varying degrees of success since the late 1950s. Currently, reported maternal mortality for cardiac operations is similar to the mortality rate for nonpregnant female patients. However, fetal mortality remains high, at approximately 20%. Aortic root replacement with an aortic homograft in a 34-year-old pregnant woman with bacterial endocarditis at 18 weeks gestation is presented. Fetal echocardiography during and after bypass was employed. PMID- 12118772 TI - Repair of a complex aortic arch anomaly associated with cutaneous hemangioma. AB - Aortic coarctation and cutaneous hemangioma is a rare association. We describe the case of a neonate with abnormal looping of the aortic arch associated with hemangioma of the head and neck who underwent complex surgical repair without cardiopulmonary bypass. PMID- 12118773 TI - Endovascular treatment of thoracic aortic fistulas. AB - Aortoesophageal and aortobronchial fistulas constitute a problem in therapy because of the high rates of morbidity and mortality associated with operation. From May 1996 to March 2000, we treated by an endovascular procedure one aortoesophageal and three aortobronchial fistulas. There was no postoperative death. We noted one peripheral vascular complication that required a surgical procedure, one postoperative confusion, and one inflammatory syndrome. In one case, because of a persistent leakage after 21 months, we had to implant a second endovascular stent graft. A few weeks later the reopening of this patient's esophageal fistula led to his death by mediastinitis 25 months after the first procedure. The few cases published seem to bear out the interest, observed in our 4 patients, of an endovascular approach to treat complex lesions such as fistulas of the thoracic aorta especially in emergency or palliative cases. PMID- 12118774 TI - Coexistence of sinus rhythm and segmental atrial fibrillation after maze procedure. AB - We present the case of an 80-year-old man with chronic atrial fibrillation associated with mitral regurgitation. The atrial fibrillation was successfully treated with the maze procedure combined with mitral valve replacement. The electrophysiological data are also reported. Recordings of sinus rhythm and intraatrial activity demonstrated the coexistence of sinus rhythm and fibrillation of both atria. This finding indicates that the sinus node was protected from segmental atrial fibrillation by entrance block, and this, in turn, is evidence of the efficacy of the maze procedure. PMID- 12118775 TI - Cardiomyocyte transplantation does not reverse cardiac remodeling in rats with chronic myocardial infarction. AB - BACKGROUND: Several reports have documented the potential benefits of cell transplantation as an alternative to cardiac transplantation. This study was designed to investigate whether cardiomyocyte transplantation is effective in rats with chronic myocardial infarction. METHODS: Syngeneic Lewis rats were used in this study. Chronic myocardial infarction was induced in rats by ligating the left anterior descending artery. Four weeks later, after left ventricular (LV) dysfunction with akinetic regions was confirmed by echocardiography, the rats were randomized into two groups: a group that received fetal cardiomyocyte transplantation (TX group; n = 11); and a group that received an intramyocardial injection of culture medium only (control group; n = 12). RESULTS: Four weeks after treatment, the TX group had smaller end-systolic dimension (LVDs) (7.5 +/- 0.9 vs 8.9 +/- 0.8 mm, p < 0.01) and better fractional shortening (FS) (26.2 +/- 5.9 vs 17.7% +/- 5.1%, p < 0.01) than the control group. However, there were no differences in LV end-diastolic dimension, LVDs, and FS between baseline and post treatment values in the TX group. In addition, plasma levels of atrial natriuretic peptide were not significantly different between the two groups 4 weeks after treatment. In microscopic examination, small amounts of transplanted cardiomyocytes were found only in the periinfarct area, not in the center of scar area, and a thicker ventricular wall in the infarct area was detected in the TX group. CONCLUSIONS: Fetal cardiomyocyte transplantation prevented, but did not reverse, cardiac remodeling that was accompanied with heart failure in myocardial infarction rats. Further investigation is warranted for optimal clinical application to the failing heart. PMID- 12118776 TI - Maze procedure and cor triatriatum repair. AB - A 39-year-old man had cor triatriatum (Lucas-Schmidt type IA) with severe mitral regurgitation and atrial fibrillation. We performed resection of the anomalous septum between the accessory chamber and the left atrium, and conducted mitral valve repair and the maze procedure. The patient regained sinus rhythm and normal pulmonary venous drainage to the left ventricle without mitral regurgitation. Histological examination demonstrated fibrotic myocardial structures in the anomalous septum. The maze procedure and complete excision of the anomalous septum proved to be effective surgical treatment for atrial fibrillation with cor triatriatum. PMID- 12118777 TI - Dilatable banding of a Blalock-Taussig shunt. AB - Dilatable banding has been used in various situations. Sometimes Blalock-Taussig shunt banding is performed to prevent pulmonary overcirculation. Recently several reports have described dilatable pulmonary artery banding. We modified these methods for flow control of a Blalock-Taussig shunt. We report the case of a neonate with truncus arteriosus in which this technique was used. PMID- 12118778 TI - Hemoptysis resulting from unilateral pulmonary artery agenesis. AB - Unilateral pulmonary artery agenesis is a rare congenital anomaly often associated with other cardiovascular abnormalities. It is usually diagnosed and surgically treated in childhood. Subjects without associated cardiac anomalies (isolated unilateral pulmonary artery agenesis) may be asymptomatic or have recurrent respiratory infections. We report a case of left pulmonary artery agenesis in a 35-year-old man complicated by hemoptysis and treated by pneumonectomy. Physicians should be aware of unilateral pulmonary artery agenesis presenting later in life as a source of chronic respiratory symptomatology or hemoptysis. PMID- 12118779 TI - Acute respiratory failure after pleurodesis with doxycycline. AB - Bedside pleurodesis through a tube thoracostomy has been shown to be effective treatment of malignant pleural effusion and pneumothorax with persistent air leak. A variety of agents can be used, and each has been shown to produce rare but potentially serious complications. We report a case of sudden, severe respiratory failure in a 42-year-old man after pleurodesis with 300 mg of doxycycline. His response was consistent with an anaphylactic reaction. After intubation, mechanical ventilation and nebulizer treatments, he rapidly recovered to baseline. On the basis of this report and a review of the literature, we believe that doxycycline may not be an innocuous agent for bedside pleurodesis and that such procedures warrant a monitored setting. PMID- 12118780 TI - Surgical staple metalloptysis after apical bullectomy: a reaction to bovine pericardium? AB - Palliation of symptomatic emphysema may include bullous resection to improve function of the remaining lung. Buttressing staple lines with bovine pericardium partially alleviates postoperative air leak, but can promote inflammation and infection. We report a patient expectorating staples and pericardium 5 years after bilateral apical bullectomy. Previous reporting of this complication in lung volume reduction operation also involved both pericardium and staples, and we propose that an ongoing local inflammatory reaction to these materials may facilitate delayed erosion into airways. PMID- 12118781 TI - Airway obstruction complicating mediastinal tuberculosis: a life-threatening presentation. AB - We discuss the case of a young man with mediastinal tuberculosis who presented with pericarditis, thymic involvement, and respiratory failure because of upper airway obstruction. Although mediastinal tuberculosis is not uncommon, the simultaneous occurrence of these complications is exceedingly rare and in this patient resulted in an acute life-threatening illness. The diagnosis and treatment of this condition can be complex, and these aspects of the patient's care are discussed. PMID- 12118782 TI - Thoracoscopic ultrasonic coagulation of thoracic duct in management of postoperative chylothorax. AB - Chylothorax is a rare but potentially serious complication of cardiac operations. We report here a 72-year-old man who underwent replacement of a descending aneurysm with a synthetic graft for dissecting aneurysm (IIIa). A persistent postoperative chylothorax developed, which necessitated continuous drainage, despite conservative treatment more than 12 days. Thoracoscopic high-frequency ultrasonic coagulation of the thoracic duct without clipping finally stopped chyle production. This method may be useful from the standpoint of minimal access, rapid recovery, less pain, and a shorter operation. PMID- 12118783 TI - Total pericardial defect: risk factor for traumatic aortic type A dissection. PMID- 12118784 TI - Anomalous segmental vein for right upper lobe: an unusual anatomical variation. PMID- 12118785 TI - A simple trick for repairing coronary pseudoaneurysm complicating a Bentall operation. AB - Coronary pseudoaneurysms are a known complication of the Bentall wrap-inclusion method of composite valve grafting. We describe two cases to illustrate a straightforward technique for repair and prevention of coronary pseudoaneurysm formation. PMID- 12118786 TI - Intraoperative confirmation of ulnar collateral blood flow during radial artery harvesting using the "squirt test". AB - Hand ischemia is a major concern after radial artery harvesting for coronary revascularization. Although a number of preoperative tests have been described to assess the adequacy of ulnar collateral blood flow, many of them are subjective and unreliable. In addition, the presence of arterial connections between the radial and ulnar systems in the elbow and forearm and variability in forearm angiology imply that assessment of alternative blood supply to the hand can only be made once collateral branches of the radial artery have been divided. We describe a technique for intraoperative assessment of ulnar collateral blood flow after mobilization and division of collateral branches of the radial artery. PMID- 12118787 TI - Ultrasonic evaluation of graft anastomoses during coronary artery bypass grafting without cardiopulmonary bypass. AB - Performance of the graft-to-coronary anastomosis in coronary artery bypass grafting without cardiopulmonary bypass is more difficult than conventional coronary artery bypass grafting. We report a new method that uses high-frequency epicardial echocardiography to detect technical errors and inadequacies in graft anastomoses. This method improves the operative outcome and enables detection of septal perforator branches and deeply embedded coronary arteries during coronary artery bypass grafting without cardiopulmonary bypass. PMID- 12118788 TI - Salinoma window technique for mediastinal lymph node biopsy. AB - Computed tomographic-guided transthoracic needle biopsy can access virtually all mediastinal lymph node stations, but is limited by the potential for pneumothorax and bleeding. To avoid these possible complications, the extrapleural "salinoma" technique was used for computed tomographic-guided mediastinal biopsies in 15 patients. Sampling methods were coaxial (8), tandem (5), and single pass (2). Diagnostic yield was 93% with no significant bleeding or pneumothorax. The salinoma technique permits biopsy of deep mediastinal lesions to stage pulmonary malignancies, while providing a technique that limits complications. PMID- 12118789 TI - Clinical significance of micrometastasis in lung and esophageal cancer: a new paradigm in thoracic oncology. AB - In the past decade, detection of micrometastatic disease in different clinical samples including pleural lavage, lymph node, bone marrow, and blood has become a rapidly growing area of interest in research of non-small cell lung cancer and esophageal cancer. The results of these studies support the concept that, just as in many other solid malignancies, systemic spread may happen at an early stage in non-small cell lung cancer and esophageal cancer. Such systemic spread is often occult (micrometastases) at the time of primary diagnosis, which may have adverse effects on survival. Improved staging can be expected with information on micrometastases, and a subgroup of patients who will benefit most from adjuvant therapy might be identified. Although reliable and standard methods need to be developed before detection of micrometastasis is incorporated in the routine clinical practice, we suggest that it be considered an important correlate in clinical trials in non-small cell lung cancer and esophageal cancer. PMID- 12118790 TI - Solitary fibrous tumors of the pleura. AB - Solitary fibrous tumor of the pleura is a mesenchymal tumor that has been increasingly recognized over the past few years. The tumor was initially described in the pleura, but it has been reported in many other sites lately. Although the majority of these tumors have a benign course, the malignant form still remains enigmatic. Indeed, the behavior of these tumors is often unpredictable and does not always correlate with histologic findings. In addition, benign tumors may remain unproblematic for several years before changing into a malignant form. In order to define more precisely the clinical behavior of solitary fibrous tumors of the pleura, we reviewed the literature with particular attention to the clinical presentation, histopathologic characteristics, and cytogenetic differentiation of these tumors. A staging system and an algorithm for the management and follow-up of these patients are proposed. PMID- 12118791 TI - Patch-and-glue technique for left ventricular free wall rupture. PMID- 12118792 TI - VHI report card gets a failing grade. Virginia Health Improvement. PMID- 12118793 TI - Is Allen's test not reliable in the selection of patients for radial artery harvest? PMID- 12118794 TI - Safety of low hematocrits during cardiopulmonary bypass. PMID- 12118795 TI - Successful cellular cardiomyoplasty in canine idiopathic dilated cardiomyopathy. PMID- 12118796 TI - Transventricular aortic cannulation. PMID- 12118797 TI - Combined coronary artery bypass grafting and lung cancer operation. PMID- 12118798 TI - Unity and participation: embracing counterintuitive survival skills. PMID- 12118799 TI - Routine enlargement of the small aortic root: a preventive strategy to minimize mismatch. AB - BACKGROUND: We routinely use aortic root enlargement (ARE) as part of one strategy to avoid prosthesis-patient mismatch in patients with relatively small aortic roots who are undergoing aortic valve replacement (AVR). METHODS: We performed a retrospective review of 657 consecutive stented AVR patients at a single institution between 1995 to 2001. Of these, 114 (17%) patients underwent ARE. Root enlargement was selectively performed in patients at risk for prosthesis-patient mismatch, defined as calculated projected indexed effective orifice area (iEOA) less than 0.85 cm2/m2. This involved extension of the aortotomy between the left and noncoronary cusps, valve implantation, and Dacron patch closure of the aorta, thus permitting replacement with a valve size appropriate to body surface area. RESULTS: The mean age of ARE patients was 72.5 +/- 11.0 years, with 32% aged 80 years or more. Of the patients, 61% were female and 27% had undergone previous cardiac operations. Combined procedures included coronary bypass in 57 patients and mitral repair or replacement in 24. The prevalence of mismatch was less than 3%. The ARE required an average of 19 minutes of additional aortic clamp time. The 30-day mortality was 0.9%. Logistic regression showed perfusion time to be the only independent predictor of mortality. CONCLUSIONS: Our results show that ARE can be performed readily and with minimal added risk relative to standard AVR. We also present a preventive strategy to minimize mismatch predicted at time of operation from the reference value of effective orifice area for a given prosthesis and the patient's size. This includes use of ARE to enhance the potential benefit of AVR. PMID- 12118800 TI - Valve replacement in patients on chronic renal dialysis: implications for valve prosthesis selection. AB - BACKGROUND: Reports are sparse describing heart valve replacement in patients with end-stage renal disease. This review assesses a 15-year experience and outcomes after valve replacement in patients on chronic preoperative renal dialysis. METHODS: A computerized database, hospital records, and telephone contact provided outcome data for patients on chronic dialysis undergoing valve replacement between March 22, 1985, and October 13, 2000, in two hospitals. RESULTS: Seventy-two patients underwent 95 valve procedures (74 operations). Ages ranged from 23 years to 84 years (mean, 57 years). Fifty-five aortic, 30 mitral, and 3 tricuspid valve replacements and 7 valvuloplasties were performed. Six of the 74 procedures were reoperative valve replacements. In the 46 patients with reliable long-term (greater than 30 days) follow-up data, significant bleeding or stroke was documented in 17 of 34 patients with a mechanical valve and 1 of 12 patients with a bioprosthetic valve. Overall survival (including two operative deaths) was 72.8% at 3 months, 65.4% at 6 months, 60.5% at 1 year, 39.8% at 2 years, 28.5% at 3 years, and 15.9% at 6 years (Kaplan-Meier). Type of valve implanted did not influence early and late survival. CONCLUSIONS: In this series of patients on chronic dialysis, survival appears to justify valve replacement. However, the sixfold higher incidence of late bleeding or stroke in patients on dialysis with a mechanical valve requiring warfarin suggests that bioprosthetic valves are the valve substitute of choice in patients on chronic dialysis. PMID- 12118801 TI - Minimally invasive tricuspid operation using port access. AB - BACKGROUND: Port-access techniques performed through a right mini-thoracotomy have been extensively described for both the mitral and aortic valves. However, reports of tricuspid valve operations using the port-access approach are rare. A technique for minimally invasive tricuspid valve operation using port access is described. METHODS: Port-access approach was applied to 33 consecutive patients undergoing tricuspid valve repair or replacement. RESULTS: Twelve percent (4 of 33) underwent tricuspid replacement and 88% underwent repair (28 of 33). Perioperative mortality was 6% (2 of 33) and conversion to median sternotomy was 3% (1 of 33). CONCLUSIONS: Port-access tricuspid operations are both feasible and safe with a low conversion rate to conventional median sternotomy. PMID- 12118802 TI - Living, autologous pulmonary artery conduits tissue engineered from human umbilical cord cells. AB - BACKGROUND: Tissue engineering represents a promising approach to in vitro creation of living, autologous replacements with the potential to grow, repair, and remodel. Particularly in a congenital operation, there is a substantial need for such implantation materials. We previously demonstrated fabrication of completely autologous, functional heart valves on the basis of peripheral vascular cells. Presently the feasibility of creating pulmonary artery conduits from human umbilical cord cells was investigated. METHODS: Human umbilical cord cells were harvested and expanded in culture. Pulmonary conduits fabricated from rapidly bioabsorbable polymers were seeded with human umbilical cord cells and grown in vitro in a pulse duplicator bioreactor. Morphologic characterization of the generated neo-tissues included histology, transmission, and scanning electron microscopy. Characterization of extracellular matrix was comprised of immunohistochemistry. Extracellular matrix protein content and cell proliferation were quantified by biochemical assays. Biomechanical testing was performed using stress-strain and burst-stress tests. RESULTS: Histology of the conduits revealed viable, layered tissue and extracellular matrix formation with glycosaminoglycans and collagens I and III. Cells stained positive for vimentin and alpha-smooth muscle actin. Scanning electron microscopy showed confluent, homogenous tissue surfaces. Transmission electron microscopy demonstrated elements typical of viable myofibroblasts, such as collagen, fibrils, and elastin. Extracellular matrix proteins were significantly lower compared with native tissue; the cell content was increased. The mechanical strength of the pulsed constructs was comparable with native tissue; the static controls were significantly weaker. CONCLUSIONS: In vitro fabrication of tissue-engineered human pulmonary conduits was feasible utilizing human umbilical cord cells and a biomimetic culture environment. Morphologic and mechanical features approximated human pulmonary artery. Human umbilical cord cells demonstrated excellent growth properties representing a new, readily available cell source for tissue engineering without necessitating the sacrifice of intact vascular donor structures. PMID- 12118803 TI - Analysis of valve motion after the reimplantation type of valve-sparing procedure (David I) with a new aortic root conduit. AB - BACKGROUND: The reimplantation type of valve-sparing procedure does not allow proper reconstruction of the sinuses of Valsalva. We assessed the valve motion after a reimplantation type (David I) of valve-sparing procedure using a new Dacron conduit that incorporates sinuses of Valsalva. METHODS: Nine consecutive patients undergoing an aortic valve-sparing procedure using the new conduit were studied using two-dimensional transesophageal echocardiography shortly (2 +/- 1 months) after operation to determine root distensibility, expressed as percent change in radius and as pressure strain of the elastic modulus. Next, monodimensional view was used to assess valve motion in its various phases (rapid valve opening velocity, slow closing leaflet displacement, rapid valve closing velocity, maximal leaflet displacement, and leaflet displacement before valve closure). Seven healthy individuals served as control subjects. RESULTS: Root distensibility was reduced at the level of the annulus and sinotubular junction but was similar to control subjects at the level of the sinuses (percent change in radius, 4.1% +/- 0.8% versus 4.5% +/- 1.2%; pressure strain of the elastic modulus, 1,286 +/- 674 g/cm2 versus 1,195 +/- 628 g/cm2). Rapid valve opening (69 +/- 34.4 cm/s versus 51 +/- 11.9 cm/s) and closing (47.6 +/- 16 cm/s versus 36.4 +/- 9 cm/s) velocity as well as slow closing leaflet displacement (24% +/- 4.7% versus 22.1% +/- 7.9%), maximal leaflet displacement (20.1 +/- 4 mm versus 22.7 +/- 1.9 mm), and leaflet displacement before valve closure (15.2 +/- 3 mm versus 17.6 +/- 0.8 mm) were similar to control subjects. CONCLUSIONS: The new aortic root conduit used in a reimplantation type of valve-sparing procedure allows the anatomic reconstruction of the aortic root with leaflet motion similar to that of normal subjects. PMID- 12118804 TI - Morphometric analysis of aortic media in patients with bicuspid and tricuspid aortic valve. AB - BACKGROUND: Patients with bicuspid aortic valves tend to develop dilatation of the ascending aorta. The aim of this study was to analyze whether or not there is any histologic difference in the aortic media of patients with a bicuspid aortic valve or a tricuspid aortic valve. METHODS: A morphometric analysis of the wall of the ascending aorta was performed in 107 patients with bicuspid aortic valves undergoing aortic valve operations. The thickness of the elastic lamellae of the aortic media and the distances between the elastic lamellae were measured with the use of an image analysis system. The histologic specimens of the ascending aorta from 61 surgical patients with tricuspid aortic valve disease served as a control. RESULTS: The patients with bicuspid aortic valves had thinner elastic lamellae of the aortic media (2.71 +/- 0.23 microm) of the ascending aortic wall than the patients with tricuspid aortic valve disease (2.83 +/- 0.23 microm) (p = 0.006). The patients with bicuspid aortic valves also had greater distances between the elastic lamellae (27.21 +/- 8.69 microm) of the ascending aortic wall in comparison with the patients with tricuspid aortic valve disease (24.34 +/- 5.32 microm) (p = 0.033). There was no difference in the total thickness of the aortic media between the groups (p = 0.62). CONCLUSIONS: Patients with a bicuspid aortic valve had thinner elastic lamellae of the aortic media and greater distances between the elastic lamellae than patients with a tricuspid aortic valve. PMID- 12118805 TI - Hydrodynamic function of the second-generation mitroflow pericardial bioprosthesis. AB - BACKGROUND: The hydrodynamic function of the smaller size Mitroflow Synergy stented pericardial bioprostheses has been studied in an in vitro fresh tissue aortic root model and compared with previous studies of free-sewn bioprostheses. METHODS: Three valves of each of the sizes 19, 21, and 23 mm were sutured into fresh tissue aortic roots and tested in a pulsatile flow simulator using two different ventricular input impedance conditions. A high-speed camera was used to study the leaflet opening and closing configurations. Mean pressure difference as a function of root mean square forward flow, effective orifice area, regurgitant volumes, and total energy loss across the valves was measured. RESULTS: Mean pressure difference with respect to root mean square forward flow decreased as the valve size increased. Thus effective orifice area increased as the valve size increased. The open leaflet configuration images showed that all three sizes of Mitroflow valves had a large circular orifice with minimal open leaflet deformation. All valves closed competently with no visible leakage and no closed regurgitant volume. The Mitroflow valves showed better effective orifice areas compared with previously tested frame-mounted porcine bioprostheses but lower effective orifice areas compared with porcine stentless bioprostheses; however, the open leaflet bending deformation was better than for any of the previously tested bioprosthetic valves. CONCLUSIONS: The hydrodynamic function of the Mitroflow Synergy stented pericardial bioprosthesis shows potential for good in vivo hemodynamic performance. The good hemodynamic performance combined with relative ease of implantation technique makes the pericardial valve a good valve in the aortic position, particularly in older patients with small annuli. PMID- 12118806 TI - A comparison of two individual amiodarone regimens to placebo in open heart surgery patients. AB - BACKGROUND: This study compares the ability of two oral amiodarone regimens to reduce the risk of atrial fibrillation (AF) as compared with the placebo among elderly open heart surgery (OHS) patients receiving beta blockade. METHODS: This is a randomized, double-blinded, placebo-controlled trial of 220 patients undergoing OHS. Patients (average age, 73 years) received 7 g of oral amiodarone more than 10 days starting 5 days before OHS (slow load; n = 56), a 6 g oral amiodarone regimen more than 6 days starting 1 day before OHS (fast load; n = 64), or matching placebo in one of the two previously mentioned regimens (n = 100). RESULTS: Patients receiving the slow load amiodarone regimen had a significant reduction in the risk of AF (48.4%; p = 0.013), AF lasting more than 24 hours (76.5%; p = 0.003), symptomatic AF (90.0%; p = 0.002), and recurrent AF (64.5%; p = 0.025) as compared with the placebo. Patients receiving the fast load amiodarone regimen had significant reductions in the risk of AF lasting more than 24 hours (52.6%; p = 0.038) and symptomatic AF (65.0%; p = 0.024), but the incidence of any AF or any recurrence of AF only showed a trend toward significance (34.0% and 45.5%; p = 0.054 and 0.09, respectively). CONCLUSIONS: Oral amiodarone in a slow loading regimen provides significant suppression of all AF factors and can be used when a patient has started it at least 5 days before OHS. If a patient has less than 5 days before OHS, the fast loading regimen is an efficacious alternative as it provides significant benefits in preventing AF from lasting more than 24 hours and for preventing symptomatic AF. Both regimens were well tolerated and safe in elderly patients receiving beta blockade according to the hospital's standard protocol. PMID- 12118807 TI - Surgical outcome of acute type A aortic dissection: analysis of risk factors. AB - BACKGROUND: The aim of this study was to assess the risk factors for the early and late outcome of the surgical treatment of acute type A aortic dissection. METHODS: From 1983 to 2000, a total of 130 patients underwent operation for acute type A aortic dissection. Extent of distal aortic resection included ascending aorta in 19 patients (15%), hemiarch in 29 (22%), and total arch in 82 (63%). In all, 31 preoperative and perioperative variables were analyzed using univariate and multiple logistic regression models for independent predictors of in-hospital mortality and risk of late reoperation. After excluding in-hospital deaths, risk factors for late death were analyzed by Cox proportional hazard analysis. RESULTS: In-hospital mortality was 19.2% (25 of 130 patients). Multivariable analysis indicated that renal/mesenteric ischemia and shock were independent predictors of in-hospital death. At 10 years, the actuarial survival rate including in-hospital mortality was 70.9% +/- 4.7%, and the reoperation event free rate was 73.5% +/- 5.7%. Aortic valve resuspension was an independent predictor of proximal aortic reoperation, whereas nonresection of intimal tear and younger age were independent predictors for distal aortic reoperation. Chronic obstructive pulmonary disease was the only independent predictor for late death. CONCLUSIONS: Patients' preoperative dissection-related complications and comorbidities significantly affect early and late survival rates after surgical treatment of acute type A aortic dissection. PMID- 12118808 TI - Open stent-grafting for aortic arch aneurysm is associated with increased risk of paraplegia. AB - BACKGROUND: Open surgery using the endovascular stent-graft is a novel technique that lessens the invasiveness of surgery for the aortic arch. However, the outcome of this procedure remains uncertain. METHODS: Between November 1996 and July 2000, a total of 19 patients underwent open surgery using an endovascular stent-graft for thoracic aortic aneurysms. There were 15 men (78.9%) and 4 women (21.1%). Patient age ranged from 29 to 82 years (mean 69.3 years, median 74 years). Atherosclerotic thoracic aortic aneurysms were present in 17 patients (89.4%) and aortic dissection in 2 patients (10.5%). RESULTS: Two patients (10.5%) died in the hospital and 4 patients (21.1%) presented with paraplegia postoperatively. Among the 4 patients with postoperative paraplegia, 1 case was complicated with intraoperative aortic dissection. The other 3 patients with paraplegia had spinal cord ischemic time of more than 60 minutes and intraoperative body weight gain of more than 4 kg. Of these 3 patients, hemodynamic instability after cardiopulmonary bypass was observed in 1 patient and cholesterin embolus in the anterior spinal artery was found at autopsy in another. On univariate analysis, age greater than 75 years was the only risk factor associated with paraplegia (p < 0.05). Autopsy findings for the 2 patients showed that the Adamkiewicz arteries were not blocked by the stent-graft in either patient. CONCLUSIONS: Intraoperative aortic dissection, embolization of the intercostal arteries, long ischemic time of the spinal cord, and excessive weight gain during operation may have been associated with the high incidence of paraplegia after open surgery using the endovascular stent-graft. PMID- 12118809 TI - Platelet-derived growth factor-induced expression of c-fos in human vascular smooth muscle cells: implications for long-term graft patency. AB - BACKGROUND: The internal mammary artery (IMA) has been shown to have a significantly superior long-term patency rate when compared with the saphenous vein (SV) graft. Cultured smooth muscle cells (SMCs) from the IMA are more resistant to the mitogenic effects of platelet-derived growth factor (PDGF), when compared with SMCs that are derived from the SV. The radial artery (RA) is currently being used as an alternative to the SV. However, no long-term patency data are available for the RA, and there is no information on the biological behavior of RA-derived SMCs in culture. METHODS: Smooth muscle cell cultures were taken from patients who underwent coronary artery bypass grafting with the IMA, RA, and SV. A quiescent state was induced by serum deprivation for 5 days. Thereafter cells were induced to proliferate by exposure to PDGF-BB. Levels of c fos expression and 3H-thymidine incorporation were used as markers of cell proliferation. RESULTS: We found that even after serum deprivation, c-fos was still detectable; however, basal levels were higher in cells from the SV than cells from either the RA (p = 0.003) or IMA (p = 0.008). After stimulation with PDGF-BB, c-fos expression was greater in SMCs from the SV relative to the RA (p < 0.001) or the IMA (p = 0.02). Finally, relative to the SV, 3H-thymidine in the RA was 0.76 +/- 0.22 (p < 0.05) and 0.39 +/- 0.24 (p < 0.002) in the IMA, respectively. CONCLUSIONS: The data indicate that SMCs from arterial conduits are more resistant to the mitogenic effects of PDGF-BB than those from venous conduits. Our results offer a mechanistic explanation of why arterial conduits might demonstrate patency superior to that of the SV. PMID- 12118810 TI - Sutureless patch technique for postinfarction left ventricular rupture. AB - BACKGROUND: Left ventricular free wall rupture is an uncommon but catastrophic event after myocardial infarction and is associated with a high mortality. After prompt diagnosis some patients may be salvaged with immediate surgical intervention. Surgical techniques used to seal the rupture vary, as few surgeons have experience with this pathologic process. We report our experience using a sutureless patch technique to treat this entity. METHODS: A review of 6 consecutive patients during an 8-year period who were referred to one cardiac unit with postinfarction left ventricular rupture was conducted. RESULTS: There were 3 men and 3 women with an average age of 71.8 years. All were hemodynamically unstable, and 4 were in electromechanical dissociation. Echocardiography confirmed the diagnosis in 5 patients, and cardiac catheterization had been performed in 4 before rupture. All patients were treated promptly with fluid, inotropic agents, and, if needed, cardiopulmonary resuscitation and pericardiocentesis. Resuscitation was continued in the operating room, and the myocardial tear was sealed with a generous patch of unsupported felt secured to the heart with cyanoacrylate glue. Coronary artery bypass grafting was performed in 3 patients if the anatomy was known. All patients survived to the intensive care unit. One death occurred as a result of severe neurologic injury. Five patients were discharged from the hospital, and all were alive 2 months to 7.5 years after operation. CONCLUSIONS: A sutureless patch technique for the treatment of postinfarction rupture is simple, effective, and associated with a favorable outcome. PMID- 12118811 TI - IP telephony--new horizon for telemedicine and e-health. AB - Internet Protocol (IP) based networks are increasingly being used as alternatives to the traditional circuit-switched networks for carrying voice traffic. IP telephony is taking a firm hold in the world telecommunication market for a wide range of applications including telemedicine. The merits of IP telephony for telemedicine lie in cost-savings and new applications and functionalities it may render. In this paper, we try to discuss and analyze the implications of IP telephony on the evolution and deployment of telemedicine and e-health. PMID- 12118812 TI - Using DEA to evaluate efficiency and formulate policy within a Greek national primary health care network. Data Envelopment Analysis. AB - The purpose of this paper is to critically evaluate the relative efficiency of primary health care centers of the principal Greek public insurance provider, the Social Security Institute (IKA). The source of the efficiency data was the Statistical Unit of IKA. Using Data Envelopment Analysis, we analyzed data from 133 centers nationwide. Input variables included the number of personnel, stratified in different categories, and the number of people covered by each health center. The number of pensioners enlisted to each health care facility was used as an index of aging and vulnerability of the covered population. According to the results of the study, centers with the technological infrastructure to perform laboratory and/or radiographic examinations exhibited higher efficiency scores. In addition, centers with eligible covered populations from 10,000 to 50,000 were found as the most efficient. Health sector reforms should be planned on the basis of such analyses. If the model is supplemented with valid demographic, socioeconomic, and epidemiological data, it may become the basis for the creation of a national health care chart, matching available resources to the population and its health care needs. PMID- 12118813 TI - Percutaneous transluminal septal reduction for hypertrophic obstructive cardiomyopathy: report from an international pilot study. AB - Assessing the effectiveness of newer treatments for rare diseases can be challenging because of the small number of patients treated at individual centers. We enrolled patients undergoing percutaneous transluminal septal myocardial ablation (PTSMA) for hypertrophic obstructive cardiomyopathy (HOCM) at five international centers (1 Japan, 2 United Kingdom, and 2 United States). Our study group developed standard data definitions regarding clinical symptom severity, previous HOCM treatment, procedure status, and outcome, and entered patient data directly into a shared, web-based registry system. In the first 10 months of 1998, 51 patients were enrolled in our registry, with 47 ultimately receiving the PTSMA procedure. Although HOCM is consider a single disease, there were significant differences among centers in patient characteristics (age, gender, and family history of HOCM), symptom severity, diagnostic techniques (measurements taken after provocation), and treatment (amount of alcohol used, timing of injection, and number of branches attempted). PMID- 12118814 TI - Negligible electromagnetic interaction between medical electronic equipment and 2.4 GHz band wireless LAN. AB - Wireless LANs using radio waves have recently gained popularity for installation in hospitals. Because electromagnetic waves transmitted from mobile telephones have been shown to cause interference with medical electronic equipment, prudence would seem necessary when introducing radio wave communication devices into hospitals. Therefore, we tested the effect of wireless LAN communication on medical electronic equipment and the effect of electronic equipment on wireless LAN communication. We observed nine pieces of electronic equipment in the operating mode while transmitting radio waves from a wireless LAN. Even when the access point was put very close to the medical electronic equipment surface and data was transmitted, no malfunction of the equipment was observed. The medical electronic equipment caused little change in the effectiveness of the communication device, although radio waves emitted from electric knives and a remote patient monitor reduced the reception rate to about 60%. The communication speed of the wireless LAN was temporarily reduced only when a microwave oven was located close to and facing the access point. Because output in Japan is limited to a maximum of 10 mW wireless LAN following the IEEE802.11b standard should be able to be installed safely in Japanese hospitals. However, wireless LAN access points should not be installed near microwave ovens. PMID- 12118815 TI - Clinical process analysis and activity-based costing at a heart center. AB - Cost studies, productivity, efficiency, and quality of care measures, the links between resources and patient outcomes, are fundamental issues for hospital management today. This paper describes the implementation of a model for process analysis and activity-based costing (ABC)/management at a Heart Center in Sweden as a tool for administrative cost information, strategic decision-making, quality improvement, and cost reduction. A commercial software package (QPR) containing two interrelated parts, "ProcessGuide and CostControl," was used. All processes at the Heart Center were mapped and graphically outlined. Processes and activities such as health care procedures, research, and education were identified together with their causal relationship to costs and products/services. The construction of the ABC model in CostControl was time consuming. However, after the ABC/management system was created, it opened the way for new possibilities including process and activity analysis, simulation, and price calculations. Cost analysis showed large variations in the cost obtained for individual patients undergoing coronary artery bypass grafting (CABG) surgery. We conclude that a process-based costing system is applicable and has the potential to be useful in hospital management. PMID- 12118816 TI - A structural equation modeling approach to examining the predictive power of determinants of individuals' health expenditures. AB - Understanding the determinants of health expenditures is essential for a fair and effective utilization profiling, particularly in the setting of capitation rates in risk-adjustment models. The objective of the study was to examine the relative importance of determinants in predicting future health expenditures, using structural equation modeling. Based on Andersen's behavioral system model, individual determinants along with prior utilization and measures of health status from 1994 are evaluated in a longitudinal design for theirpredictive powerfor health expenditures in 1995. A total of 4,255 policy-holders enrolled in three health plans at Trigon BlueCross/BlueShield of Virginia who responded to a mail survey were included for analysis. Person-level annual charges for health services utilization were used as the dependent variable. Five health scales were excerpted from Health Survey SF-36 to represent an individual's health status. Excluding prior utilization in 1994, health status (gamma = -0.19, p < 0.001) and having diabetes (gamma = 0.08, p < 0.001) are two statistically significant predictors of health expenditures in 1995. Including prior utilization, both health status (gamma = -0.15, p < 0.001) and prior utilization (gamma = 0.15, p < 0.001) are the most important predictors, followed by having diabetes (gamma = 0.08, p < 0.001). Health status is a powerful predictor offuture health expenditures, even when prior utilization is controlled. PMID- 12118817 TI - Categorization and analysis of pain and activity in patients with low back pain using a neural network technique. AB - Low back pain represents a significant medical problem, both in its prevalence and its cost to society. Most episodes of acute low back pain resolve without significant long-term functional impact. However, a minority of patients experience extended chronic pain and disability. In this paper, we have explored new techniques of patient assessment that may prospectively identify this minority ofpatients at risk of developing poor outcomes. We studied 15 patients with acute low back pain and 25 patients with chronic low back pain over 4 month's time. Patients monitored their pain and activity levels continuously over the first 3 weeks. Pain and functional status were assessed at baseline and at 3 weeks following enrollment. Follow-up assessment of functional status and progress were performed at 2 and 4 months. The pain and activity levels were categorized using a self-organizing-map neural network. A back-propagation neural network was trained with the categorization and outcome data. There was a good correlation between the true and predicted values for general health (r = 0.96, p < 0.01) and mental health (r = 0.80, p < 0.01). No significant correlation was found if activity and pain data were not entered into the analysis. Our results show that neural network techniques can be applied effectively to categorizing patients with acute and chronic low back pain. It is our hope that future research will allow these categorizations to be tied to prognostic and therapeutic decisions in patients who present with episodes of back pain. PMID- 12118818 TI - Exploring hospitals' adoption of information technology. AB - This study explores the adoption of information technology (IT) and the association between organizational and market factors, and IT adoption in hospitals. Results suggest that a wide range of amounts and types of IT are adopted. Hospitals with higher overall IT adoption adopt strategic IT most often. Hospitals with lower IT adoption adopt administrative IT most often. Results also show hospital IT adoption to be positively associated with hospital size, location, system membership, ownership, and market competition. PMID- 12118819 TI - Joint magnetic resonance imaging: normal variants and pitfalls related to sports injury. AB - MR imaging abnormalities, such as increased signal within normally hypointense structures, form and attachment abnormalities, fluid collections in joints, tendon sheaths and bursa, or even tumors, such as Morton's neuromas, are common in asymptomatic volunteers. They may be explained by normal physiology, anatomic variability, MR imaging artifacts, or true abnormalities without clinical importance. Although it is not always possible to differentiate such variants or artifacts from clinically relevant findings, it is important to know their potential cause and clinical importance and not to over-report them as abnormality requiring additional imaging or treatment. Thorough knowledge of normal anatomy is crucial in this situation. PMID- 12118821 TI - Straight and rotational instability patterns of the knee: concepts and magnetic resonance imaging. AB - The role of the musculoskeletal radiologist of the twenty-first century is not satisfied by the simple enumeration of findings on imaging studies. In this day of turf battles over the right to interpret images, in the interest of service to referring clinicians, and the optimal care of the patient, it is the responsibility of the radiologist to ascend to a higher level of sophistication in the understanding of the pathology encountered and the implications of our diagnoses. As demonstrated in this overview of the clinical and imaging approach to complex injuries of the knee, it is clear that the physical examination assessment of this patient population can be quite challenging. With a detailed understanding of anatomy, pathology, and what abnormalities change the management of the patient, the radiologist can alert the clinician to potential pitfalls in diagnosis. Neither the clinician nor the radiologist should be satisfied with a single diagnosis, for this introduces the potential of overlooking an associated injury that could preclude the return of a normally functioning articulation. PMID- 12118820 TI - Imaging of sports-related knee injuries. AB - This article reviewed the major sports medicine conditions affecting the knee and their MR imaging appearance. MR imaging of the knee is considered efficacious especially in the setting of indeterminate clinical findings and can stratify patients, guiding further surgical management. MR imaging affects the diagnosis and management of acute knee injury by improving clinician diagnostic certainty, assisting in management decisions, and decreasing the number of arthroscopic procedures. From a societal perspective, knee MR imaging can be considered a cost effective modality when compared with diagnostic and borderline therapeutic arthroscopies (e.g., debridement alone). For these and other reasons, knee MR imaging has shown a substantially increased use for the evaluation of sports medicine conditions. PMID- 12118822 TI - Magnetic resonance imaging of sports-related injuries to the shoulder: impingement and rotator cuff. AB - MR imaging provides clinically useful information in detecting and characterizing sports-related pathology of the rotator cuff and other shoulder disorders in a non-invasive fashion. Complete and partial tears of the rotator cuff, as well as factors contributing to impingement, can be detected and characterized with MR imaging. The size and location of complete tears of the rotator cuff can be accurately determined with MR imaging. PMID- 12118823 TI - Sports-related injuries of the shoulder: instability. AB - With current technology a properly conceived imaging strategy can demonstrate instability lesions in the athlete. Plain radiographs can diagnose acute dislocations and assess successful reductions. In addition, plain radiographs can demonstrate Hill-Sachs and, more importantly for instability, osseous Bankart lesions. In the acute setting, conventional MRI nicely demonstrates labral Bankart, ligamentous. and tendonous injuries that result from dislocations and can lead to instability. In the setting of chronic instability, MR arthrography best evaluates these lesions. In the postoperative shoulder, muitislice CT arthrography may be the modality of choice, but further investigation is needed. If large series validate multislice CT arthrography for the evaluation of postoperative instability lesions, this technique may become widely used in athletes and in other populations where recurrent instability is a problem. Other imaging strategies may also find an increasing central role in evaluating shoulder instability lesions. Indirect MR arthrography, for example, may have a role in assessing these lesions in athletes . Another intriguing technology for this application is the development of high field (0.5 Tesla or greater) open magnets. In such a setting, physiological relationships in the shoulder with motion and stress may be evaluated. Such imaging may farther illuminate our understanding of the stable and unstable shoulder. Unfortunately, with all imaging modalities, whether widely used or experimental, outcomes data is Lacking. How do the various imaging modalities and strategies affect patient outcome? The answer is unknown and needs to be answered before a definitive patient work-up for shoulder instability can be established. PMID- 12118824 TI - Imaging of elbow injuries in the child and adult athlete. AB - Because of the often complex and sometimes poorly remembered history of trauma to the elbow, imaging beyond conventional plain film radiographs is often needed. Usually, this consists of high-resolution MR imaging to evaluate the articular cartilage, supporting ligaments, and tendons about the elbow. Sonography, however, can also be used, especially when there is a targeted clinical question as to the presence of epicondylitis, or to provide guidance for diagnostic or therapeutic injections. PMID- 12118825 TI - Imaging of hip disorders in athletes. AB - Normal hip joint function is fundamental in running-, jumping-, and kicking-based sporting activities. Hip disorders do not account for a large portion of exercise related injuries, but they can pose a clinical dilemma since symptoms tend to be non-specific. Conventional radiographs may demonstrate some causes of hip pain, such as stress fractures and degenerative joint disease. Magnetic resonance (MR) imaging of the hip has proven valuable in the diagnosis of radiographically occult osseous abnormalities and periarticular soft tissue disorders such as stress fractures, avulsion injuries, musculotendinous abnormalities, and bursitis. Conventional MR imaging has been less useful in the evaluation of intra articular lesions including acetabular labral tears, intra-articular loose bodies, and cartilage lesions. The visualization of intra-articular structures and their abnormalities can be improved by the injection of diluted Gadolinium, which distends the capsule and leaks into labral tears. This article will focus on the use of conventional radiography and MR imaging in recreational and professional athletes with painful hip joints, and where possible it will compare MR imaging with other diagnostic modalities such as bone scan and CT. PMID- 12118826 TI - Imaging of athletic injuries to the ankle and foot. AB - Conventional radiographs in conjunction with clinical examination remains the primary method for evaluating the acute athletic injury. In most cases, suspected acute tendon and ligament injuries are initially treated based on physical examination. Magnetic resonance (MR) imaging, with its multiplanar capability and superb soft tissue contrast, is quickly becoming the method of choice for evaluating chronic foot and ankle pain and further defining the extent of tendon and ligament injuries. This article reviews the common acute and chronic (overuse) foot and ankle athletic injuries with an emphasis on imaging characteristics. PMID- 12118827 TI - Imaging of stress fractures in the athlete. AB - Osseous stress fractures and stress reactions represent the effect of abnormal repetitive stress on normal bone. An accurate and thorough clinical history and sequential radiographs often suffice 40 make the diagnosis especially when the fracture occurs in one of the common locations, such as the tibia, metatarsals, or calcaneus. In cases that are atypical in location or clinical presentation the authors rely more on MR imaging, radionuclide bone scanning, and occasionally CT. MR imaging detects early changes of osseous stress injury and allows precise definition of anatomy and extent of injury, and is the preferred modality for evaluating the continuum of osseous manifestations of stress injury. MR imaging is useful in evaluating shin splints, early osseous stress injuries, and overt stress fracture. In the elite athlete prompt diagnosis and early rehabilitation are the goals. PMID- 12118828 TI - Imaging of sports-related muscle injuries. AB - Muscle derangements in athletes have a wide variety of causes, treatments, and prognoses. Given that the cause and severity of sports-related injuries may be difficult to determine clinically in some cases, MR imaging is utilized increasingly to evaluate muscle injuries in athletes. After reviewing useful MR imaging techniques, this article focuses on MR imaging of the most common causes of muscle pain and disability in athletes, including myotendinous strain, delayed onset muscle soreness, muscle contusion, myositis ossificans, muscle laceration, muscle herniation, and compartment syndrome. The differential diagnosis of various signal intensity abnormalities in muscle also is reviewed. PMID- 12118829 TI - Ultrasound in sports medicine. AB - Musculoskeletal sonography has been shown to be effective for many applications related to sports medicine. Some advantages of sonography over MR imaging include portability, accessibility, high resolution, and relative lower cost. More importantly, dynamic imaging under sonography visualization allows diagnoses that cannot be made with routine MR imaging. Additionally, direct imaging correlation with patient symptoms provides important information to the referring clinicians. The disadvantages of sonography include operator dependence and long learning curve. This can be minimized, however, with proper training and standardized technique. Musculoskeletal sonography has proved itself as one of several imaging methods invaluable to the diagnosis of sport medicine-related abnormalities. PMID- 12118830 TI - Photochemotherapy for GvHD. PMID- 12118831 TI - Referral for chronic renal disease: the need for a change in threshold. PMID- 12118832 TI - Creating and maintaining autologous arteriovenous fistulae: the importance of surgical salvage. PMID- 12118833 TI - High molecular weight kininogen adsorption on hemodialysis membranes: influence of pH and relationship with contact phase activation of blood plasma. influence of pre-treatment with poly(ethyleneimine). AB - Protein adsorption is an essential parameter for the evaluation of the hemocompatibility of biomedical materials. In effect, protein adsorption often generates unfavorable, complex, biochemical reactions and precedes cell adhesion on artificial interfaces. It is therefore necessary to modify the surface in contact with blood in order to minimize the onset of undesirable, biochemical, cascade reactions. Adsorption of high molecular weight kininogen (HK), which participates in the contact phase activation of the endogenous blood coagulation cascade, was evaluated at 37 degrees C. This paper also presents the results of contact phase activation tests carried out in vitro with 1:20 diluted human plasma flowing through minidialyzers containing hollow fibers of synthetic hemodialysis membranes. These tests showed that contact phase activation is strongly dependent on pH around the physiological value (ranging from 7.35 to 7.80) for negatively-charged membranes. The same pH effect was observed on the AN69 membrane as regards HK adsorption from binary labeled solutions of 125I-HK and 131I-Fibrinogen at concentrations corresponding to 1% diluted plasma. It is suggested that the influence of pH could be related to imidazole pKa values in histidine residues of kininogen D5H domain. The same study was also conducted with hemodialysis membranes pre-treated with poly(ethyleneimine). Both HK adsorption and contact phase activation proved to be greatly reduced, irrespective of the pH value (between 7.0 and 7.8). Hence, positively-charged poly(ethyleneimine) adsorbed on the membrane through strong ionic interactions with sulfonate groups of the surface probably constitutes a repelling, water swollen layer for the kininogen molecule. In addition, the advantages of the high levels of adsorbance of small molecules on the AN69 membrane leading to blood epuration, due to its high porosity and for some of them to its charge density, should not be lost by such a surface treatment. PMID- 12118834 TI - Lipid-lowering therapy and coagulation/fibrinolysis parameters in patients on peritoneal dialysis. AB - Patients on continuous ambulatory peritoneal dialysis (CAPD) often have abnormalities of lipid metabolism or coagulation and fibrinolysis, these patients may thus be more susceptible to atherosclerosis than those on hemodialysis. It has been reported that hypercoagulability and hyperfibrinolysis are correlated with abnormalities of lipid metabolism. Therefore, we investigated the effect of a decrease in lipids on the coagulation and fibrinolysis system in CAPD patients with hyperlipidemia who received lipid-lowering therapy. The patients included 5 men and 13 women, with a mean age of 52.5 years. Pravastatin sodium (10 mg/day) and ethyl icosapentate (1800 mg/day) were administered concomitantly for 8 weeks. Lipid levels and coagulation/fibrinolysis parameters were measured before and after therapy. The patients were divided into two groups depending on their response to therapy: responders showed a decrease in total cholesterol or triglycerides by at least 20% and non-responders showed less improvement. In the responders, the levels of protein C, tissue plasminogen activator/plasminogen activator inhibitor-I complex, factor XIII, alpha2-plasmin inhibitor, and D-dimer were significantly lower after therapy than before therapy. Protein C, factor XIII, and alpha2-plasmin inhibitor were also significantly decreased after therapy in non-responders, but the extent of the decrease was smaller. The plasminogen level was significantly increased after therapy in non-responders. These findings suggest that a decrease in lipid levels and/or some other action by lipid-lowering agents may correct abnormalities of coagulation and fibrinolysis in CAPD patients. PMID- 12118835 TI - Oxidative stress: the effect of erythropoietin and the dialysis membrane. AB - Dialysis patients run the risk of impaired antioxidative defense and increased free radicals (FR) production. The study was made in order to compare FR-related parameters in ten patients treated with erythropoietin (EPO+) and ten patients not subject to this treatment (EPO-). All patients showed stable hemoglobin levels at > 95 g/L. FR-related parameters were monitored during hemodialysis (HD) using a polysulfon (PS) or a hemophan (H) membrane for 12 of them (6 EPO+ a 6 EPO ). The EPO- group was found to have a higher activity of superoxide dismutase (SOD, 1160 + 218 vs; 882 + 125 IU/gHb, p<0.01) and a higher SOD/glutathione peroxidase (GSHPx) ratio compared with EPO+ (30.5 +/- 7.1 vs; 21.2 + 4.8, p<0.01). A total of 35 healthy volunteers were also examined. When compared with controls EPO- showed higher SOD (p<0.001), lower GSHPx (p<0.05) and a higher SOD/GSHPx ratio (p<0.001). Thiobarbituric acid reacting substances in EPO+ and EPO- were comparable with the levels found in controls. HD using H as well as PS membranes was associated with a decrease in erythrocyte glutathione levels (GSH after 30 minutes; also for H after HD). HD using H and PS membranes resulted in a decrease in the plasma antioxidant capacity (AOC). We can conclude that the intraerythrocyte antioxidant conditions of EPO+ patients are similar to those found in the general population and differ from those in EPO- exhibiting increased SOD and the SOD/GSHPx ratio. HD using the H as well as the PS membrane is accompanied by oxidative stress. PMID- 12118836 TI - Velocities, shear stresses and blood damage potential of the leakage jets of the Medtronic Parallel bileaflet valve. AB - Even nowadays, the essential problem of mechanical heart valve prostheses is the risk of thromboembolic events mainly caused by unnatural hemodynamics, e.g. just a few years ago the Medtronic Parallel (MP) showed unsatisfactory clinical results caused by thrombi. Therefore, in vitro investigations of the whole leakage jets were performed at the MP in mitral position by means of a pulse duplicator using a two channel laser Doppler anemometer. From the measured data, mean velocity profiles and the distribution of Reynolds shear stresses, as a function of the location within the jet, were calculated. From this data the potential of blood damage is evaluated computing a Blood Damage Index (BDI) of hemolysis and platelet damage. Four regurgitant free jets right above the hinges were observed during systole at the inflow side of the MP. The peak velocities at the origin of the jets were in the order of 1.6-2.1 m/s. Two jets experienced maximum turbulent shear stresses around 100 N/m2 within this area. The BDI for platelets of the MP is around ten times higher than the BDI of the St.-Jude Medical. The study shows that besides the flow structure within the hinges of a mechanical heart valve, the whole regurgitant jet has a large blood damage potential. This potential is measurable, respectively calculable and seems to be (on account of it's support of the clinical outcome) one piece of the puzzle that explains the negative trials of the MP. PMID- 12118837 TI - Extracorporeal photochemotherapy after cardiac transplantation: a new therapeutic approach to allograft rejection. AB - Photopheresis (ECP) is a new immunomodulatory therapy in which recipient lymphocytes are treated extracorporeally with 8-methoxypsoralen and ultraviolet light. The treatment seems to induce an inhibition of both humoral and cellular rejection after transplantation. OBJECTIVE: Since recurrent rejection (RR) continues to be a severe complication after heart transplantation (HTx) and the immunosuppressive regimes used for the treatment are often associated with increased morbidity and mortality, we investigated whether ECP could have a beneficial effect on the number and severity of rejection episodes. METHODS: Eleven HTX recipients (5 M and 6 F, mean age 48.5 yrs) with RR were enrolled in the study. ECP was performed at weekly intervals during the 1st month, at 2 week intervals during the 2nd and 3rd month, and then monthly for another 3 months. RESULTS: The fraction of biopsies (EMB) with a grade 0/1A rejection increased during ECP from 46% to 72% while the EMB showing a 3A/3B rejection decreased from 42% to 18%. It is also noteworthy that out of the 78 EMB performed during ECP only one showed a 3B rejection in comparison with 13 out of 110 EMB in the pre ECP period. Six rejection relapses were observed in a total follow-up of 60 months, two of them occurring during the tapering of oral steroid. Four relapses were reversed by ECP, one by i.v. steroids and the last by methotrexate after the failure of both i.v. steroids and ECP. The mean doses of immunosuppressive drugs resulted lower after 6 months of ECP: steroids were reduced from 13 to 8.25 mg/day, cyclosporine from 375 to 285 mg/day, azathioprine from 55 to 35 mg/day. CONCLUSIONS: ECP is a well tolerated treatment. Its administration allows better RR control and significant reduction in immunosuppressive therapy. PMID- 12118838 TI - Photopheresis in cutaneous T-cell lymphoma: five-year experience. AB - BACKGROUND: Cutaneous T-cell lymphoma (CTCL) includes several lymphoproliferative disorders involving mature T-lymphocyte proliferation initially confined to the cutis. These affections, after variable periods, may progress to the blood, limph nodes and visceral organs. Mycosis fungoides (MF) is the most frequent form of CTCL and has an indolent clinical course. The therapy of CTCL depends on the stage of the disease and the patient's general conditions. For advanced cases it includes chemotherapy, retinoids, and interferon-alpha. Since 1987 extracorporeal photochemotherapy (ECP), a novel immunomodulatory approach based on apheresis and photoirradiation of leukocytes, has been successfully introduced for the treatment of advanced CTCL. It can prolong survival of patients with erythrodermic CTCL without significant side effects. OBJECTIVE: To review our five-year experience with ECP in CTCL. METHODS: Since June 1994, 33 CTCL patients have been recruited for ECP, using two different regimens: two procedures on two consecutive days at four-week intervals for six months, or at two-week intervals for three months with progressive tapering in the second three-month period for the more severe forms. Six patients received ECP with IFN-alpha. ECP was done using the photopheresis UVAR system and UVAR XTS (Therakos, West Chester, Pa) and always with 8-MOP liquid formulation injected directly into the buffy coat bag. Lymphocytes in peripheral blood were immunophenotypically characterized for each patient and every ECP session. RESULTS: All patients tolerated ECP well, without significant side effects. Thirty patients are clinically evaluable (at least three ECP cycles). A favourable clinical response was obtained in 80.9% (16/21) of MF patients (complete response 33%, partial response 47.6%) and in 66% (6/9) of patients in the Sezary's syndrome phase (complete response 33.3%, partial response 33.3%). Five of the six patients given IFN-alpha as adjunctive therapy had a PR and one a CR. Four patients are in CR without therapy at follow-ups of 46, 20, 10 and 8 months. There have been no changes in the peripheral lymphocyte immunophenotype during the follow-up. In 19/30 patients the CD95 antigen, correlated with cellular apoptosis, was expressed and was frequently associated with a good clinical response. CONCLUSIONS: In our experience ECP achieved favourable clinical responses in 73% of patients, in monotherapy or in combination with IFN-alpha, without significant side effects. PMID- 12118839 TI - Antiviral therapy for varicella and herpes zoster. AB - Varicella-zoster virus (VZV) causes 2 clinical illnesses, varicella (chickenpox) and herpes zoster (shingles). The purpose of this review is to describe the role of antiviral therapy in the treatment of VZV infections in healthy and immunocompromised children. Acyclovir is the drug of choice for varicella and herpes zoster. The route of administration may be intravenous or oral, depending on the immunocompetence of the host. The clinical impact of acyclovir therapy is related directly to its use early in the clinical course and to the likely susceptibility of the patient to severe or life-threatening VZV infection. Patients who have the most clinical benefit are otherwise healthy adolescents with varicella infection and high-risk populations of immunocompromised children who have varicella or herpes zoster. The morbidity and mortality of VZV infections are reduced substantially by initiating acyclovir treatment early in the course of the disease. PMID- 12118840 TI - Antiviral therapy for cytomegalovirus infections in pediatric patients. AB - Appreciation of the spectrum of illness caused by cytomegalovirus (CMV) infections has increased markedly during the past 2 decades. The number of immunosuppressed patients also has increased during the same time period, reflecting the central tenet that CMV disease is most severe in this patient population. Fortunately, antiviral therapies with activity against CMV also have been identified during this same time course, and they include ganciclovir, foscamet, and cidofovir. Although all 3 of these therapies can have significant toxicities associated with them, nonetheless they are employed with relative frequency to treat potentially life-threatening CMV disease. Ganciclovir is the first-line compound used, followed by foscarnet and cidofovir. This article summarizes those CMV infections that require antiviral therapy and outlines therapeutic options for each. PMID- 12118841 TI - Antiviral therapy for influenza virus infections. AB - Every year, influenza viruses cause global epidemics that result in significant morbidity and mortality. Influenza infections can be serious in children, especially infants and toddlers. Four antiviral agents, amantadine, rimantadine, oseltamivir, and zanamivir, are available for the treatment or prophylaxis of influenza. Experience with the use of these antiviral drugs for influenza in children is limited. Given the small degree of therapeutic gain that is reported from clinical trials, considerations about cost effectiveness are important in deciding whether to use these agents in the treatment of suspected or proven influenza infections in healthy children. PMID- 12118842 TI - Carbapenems. PMID- 12118843 TI - Enterovirus infections: diagnosis and treatment. AB - Enteroviruses cause infections that present in diverse ways and affect people of all ages. Infections peak during summer and fall epidemics and cause 10 to 15 million symptomatic infections annually in the United States. The 70 enteroviral serotypes cause illness that ranges from nonspecific fevers and rashes to life threatening myocarditis or central nervous system disease. These common infections create a significant burden on our society and healthcare system. New developments in rapid diagnosis of enterovirus infections using polymerase chain reaction (PCR) positively affect patient management and have the potential to reduce the healthcare impact of enterovirus infection. The future holds promise for effective antiviral drugs that can treat enterovirus infections and decrease their significant morbidity and mortality. PMID- 12118844 TI - Controversies in isolation and general infection control practices in pediatrics. AB - Current controversies in pediatric isolation and infection control include correct application of standard precautions, importance of providing adequate staffing levels in intensive care units to prevent transmission of infectious agents, use of rapid diagnostic testing and best precautions to prevent transmission of respiratory syncytial virus, best methods to prevent transmission of multidrug-resistant organisms in acute care settings, and preventing transmission of infections to pregnant healthcare workers. Recommendations are evidence-based. PMID- 12118845 TI - Fever of unknown origin in a previously healthy child. PMID- 12118846 TI - Thomas Huckle Weller MD: Nobel Laureate and research pioneer in poliomyelitis, varicella-zoster virus, cytomegalovirus, rubella, and other infectious diseases. AB - In 1954, the Nobel Prize for Medicine was awarded to Drs John Enders, Thomas Weller, and Frederick Robbins for their watershed discovery that growth of poliomyelitis virus occurred in cultures of cells of extraneural origin, first reported in 1949. Their demonstration in 1949 that the Lansing type II strain of poliomyelitis could be grown in cultures of human embryonic tissue set into motion a race to develop a vaccine for the disease that had crippled countless thousands of individuals. The discovery and subsequent recognition were only the beginning of a prolific career for Thomas Huckle Weller, who made numerous contributions to the field of virology, including isolating the varicella-zoster virus (VZV) from cases of chickenpox and zoster, providing suggestive evidence that the same virus is responsible for both diseases; isolating the human cytomegalovirus (CMV) for the first time in tissue culture and suggesting the descriptive name now used for it; establishing Coxsackie viruses as the cause of epidemic pleurodynia: and first isolating rubella virus, the cause of German measles. This article presents a brief biography of Dr Thomas Huckle Weller, one of the field's most important figures, with primary focuses on his work on poliomyelitis, varicella-zoster virus, rubella virus, and cytomegalovirus. PMID- 12118847 TI - Herpes simplex virus infection. AB - Herpes simplex virus (HSV) infections are among the infections most frequently encountered by humans. Two types of HSV infections have been identified-HSV-1, which usually causes orolabial disease, and HSV-2, which is associated more frequently with genital and newborn infections. Usually, HSV causes mild and self limited disease of the mouth and lips or at genital sites. However, on occasion, the disease can be life-threatening. Such is the case with neonatal HSV infection and HSV infections of the central nervous system. Furthermore, in the immunocompromised host, severe infection has been encountered and is a source of morbidity. Even in the immunocompetent host, frequent recurrences, particularly those of the genital tract, can be debilitating. Because HSV does cause genital ulcerative disease, it is associated with an increased risk of acquiring a human immunodeficiency virus infection. During the past 2 decades, selective and specific inhibitors of HSV replication have been developed. These agents, acyclovir, valaciclovir, and famciclovir, all accelerate the events of healing and decrease the probability of excreting the virus when they are taken in a suppressive fashion. The long-term safety of acyclovir has been unequivocally established. Its prodrug, valaciclovir, and the prodrug of penciclovir, famciclovir, have not been used in practice as long and, therefore, less is known about these agents; however, neither is available as a pediatric formulation. PMID- 12118848 TI - Putting one objection to HLA matching on ice. PMID- 12118849 TI - Transplant physicians bear full responsibility for the consequences of kidney donation by a minor. PMID- 12118850 TI - Dendritic cells, tolerance induction and transplant outcome. PMID- 12118851 TI - Hyperhomocysteinemia in renal transplant recipients. AB - Renal transplantation is a commonly performed curative procedure for end-stage renal disease. With the increase in renal allograft half-lives, attention is now being focused on cardiovascular morbidity and death in the renal transplant recipient (RTR) population. Among the more novel cardiovascular disease (CVD) risk factors for which this group is at risk is hyperhomocysteinemia. Hyperhomocysteinemia has been associated with an increased risk of CVD, although prospective randomized trials designed to prove causality are still ongoing. Since plasma total homocysteine levels are inversely related to renal function, RTRs have a greatly increased prevalence of hyperhomocysteinemia. Other determinants of homocysteine include B-vitamins, albumin, age, and genetic polymorphisms. Although RTRs are resistant to the typical B-vitamin doses used to correct hyperhomocysteinemia in the general population, they do respond to supraphysiologic dose therapy. In terms of prevalence, etiology, and treatment of hyperhomocysteinemia, RTRs are very similar to the much larger chronic renal insufficiency population. For this reason, RTRs have been chosen as an ideal study population in investigating the effect of reducing hyperhomocysteinemia on CVD outcomes. PMID- 12118852 TI - Activation of natural killer cells and macrophages by porcine endothelial cells augments specific T-cell xenoresponse. AB - The rejection of xenografts is characterized by infiltration of monocytes and natural killer (NK) cells into the graft, suggesting an important role for the innate immune system in xenorecognition. In this study, purified human NK or T cells were cocultured with porcine endothelial cells, and cytokines were analyzed by ELISA and intracellular FACS. We demonstrated a vigorous human anti-porcine xenoresponse that was associated with a strong T-cell proliferation against porcine endothelial cells. Limiting dilution cloning and T-cell receptor (TCR) Vbeta gene usage revealed a low number of xenoreactive T-cell precursors. We demonstrated that xenogeneic porcine but not allogeneic human endothelial cells induced the early production of interferon (IFN)-gamma by human NK cells but not by CD3+ T cells. Porcine xenoantigen-induced IFN-gamma production was only partially dependent on IL-12. Blocking IL-12 with neutralizing antibodies or by depletion of human macrophages partially decreased IFN-gamma production by CD56+ NK cells. Three-color flow cytometry revealed that IL-12 was produced through a species-specific activation of human macrophages by porcine endothelial cells. Our results indicate that the direct activation of NK cells and macrophages by porcine endothelial cells provides a unique pathway of xenorecognition that augments downstream specific T-cell immunity and represents a powerful effector mechanism in xenograft rejection. PMID- 12118853 TI - The immunobiology of inductive anti-CD40L therapy in transplantation: allograft acceptance is not dependent upon the deletion of graft-reactive T cells. AB - CD40-CD40L costimulatory interactions are crucial for allograft rejection, in that treatment with anti-CD40L mAb markedly prolongs allograft survival in several systems. Recent reports indicate that costimulatory blockade results in deletion of graft-reactive cells, which leads to allograft tolerance. To assess immunologic parameters that were influenced by inductive CD40-CD40L blockade, cardiac allograft recipients were treated with multiple doses of the anti-CD40L mAb MR1, which was remarkably effective at prolonging allograft survival. Acute allograft rejection responses such as IL-2 producing helper cell priming, Th1 priming, and alloantibody production were abrogated by anti-CD40L treatment. Interestingly, the spleens of mice bearing long-term cardiac allografts following inductive anti-CD40L treatment retained precursor donor alloantigen-reactive CTL, IL-2 producing helper cells, and Th1 in numbers comparable to those observed in naive mice. These mice retained the ability to reject donor-strain skin allografts, but were incapable of rejecting the original cardiac allograft, or a second donor-strain cardiac allograft. Further, differentiated effector cells were incapable of mediating rejection following adoptive transfer into mice bearing long-term allografts, suggesting that regulatory cell function, rather than effector cell deletion was responsible for long-term graft acceptance. Collectively, these data demonstrate that inductive CD40-CD40L blockade does not result in the deletion of graft-reactive T cells, but induces the maintenance of these cells in a quiescent precursor state. They further point to a tissue specificity of this hyporesponsiveness, suggesting that not all donor alloantigen reactive cells are subject to this regulation. PMID- 12118854 TI - The use of a minor as a live kidney donor. AB - An analysis of the UNOS database suggests a practice pattern that uses live minor kidney donors in clinical circumstances not endorsed by the recommendations of a recent consensus conference on live organ donation. These data reveal that minor donor kidneys were transplanted more frequently to adult than to pediatric recipients, that only 12% of all recipients were identical twins, and that the use of a minor donor provided no better outcome than that expected from an adult donor. Live organ donation from a minor should only be considered when there is no other living donor available and all other opportunities for transplantation have been exhausted. PMID- 12118855 TI - Successful long-term outcomes using pediatric en bloc kidneys for transplantation. AB - GOAL: The objective of our study was to determine whether acceptable long-term graft survival and function can be achieved using pediatric en bloc renal transplants by employing specific immunologic and selection strategies. MATERIALS AND METHODS: A retrospective analysis of pediatric en bloc kidney transplants at a single institution was performed. A Kaplan-Meier analysis was used to evaluate graft survival. FINDINGS: Fifty-seven adult recipients with at least a 1-year follow-up period were successfully transplanted using pediatric en bloc kidneys between 1993 and 1998. Complete data regarding immunosuppression were available for 53 patients. All patients had a cyclosporine (CsA)- or tacrolimus (TAC)-based regimen with either azathioprine (Aza) or mycophenolate mofetil (MMF) and corticosteroids. All but two received induction with OKT3. One-, 3-, 4-, 5- and 7 year graft survival was calculated to be 88%, 86%, 83%, 68% and 68%, respectively. The mean serum creatinine value at 3 years was 1.0+/-0.4 mg/dL. Thirteen patients (23%) had biopsy-proven rejection. Ten of 19 (53%) patients treated with CsA/Aza had rejection, whereas 2/15 (13%) on CsA/MMF and 1/19 (5%) of patients on TAC/MMF had rejection. Nine patients (16%) had surgical complications. CONCLUSION: Excellent long-term results can be achieved in pediatric en bloc kidney transplantation using OKT3, TAC and MMF in carefully selected adult recipients. PMID- 12118856 TI - Apolipoprotein C-III and E polymorphisms and cardiovascular syndrome, hyperlipidemia, and insulin resistance in renal transplantation. AB - Hyperlipidemia and insulin resistance frequently develop after renal transplantation, contributing to cardiovascular disease. Individual differences in response based upon genetic variations in proteins regulating lipidic and glucose tolerance metabolism could be expected. In the general population, the S2 allelic variant of the apoprotein (apo) C-III gene has been associated with hypertriglyceridemia and an insulin resistant state, whereas the E4 allele of the apo E has been associated with hypercholesterolemia and atherosclerosis. Its influence in renal transplant patients remains to be seen. In order to assess the impact of apo E and C-III major polymorphisms on atherosclerotic vascular disease, lipid profile and impaired glucose tolerance in renal transplant patients, we studied 110 consecutively examined patients undergoing kidney transplantation (age range 24-73 years). Atherosclerotic complications were detected in 25% of patients, with age, male sex and hypercholesterolemia being significant atherosclerotic risk factors. Among the male patients with E4 allele, the odds ratio for coronary disease and global atherosclerosis were 10.2 (95% CI) and 6.4 (95% CI), respectively. There were no significant differences in the frequency of any of the polymorphisms among patients with dyslipidemia and impaired glucose tolerance. As the number of patients in our sample was small, larger studies are needed to verify these issues. While in the studied population C-III polymorphism appears to have little association with the prevalence of atherosclerotic complications, E4 allele should be considered as a genetic marker of coronary artery disease and global atherosclerosis in renal transplant patients. PMID- 12118857 TI - Laparoscopic incisional hernia repair in liver transplant and other immunosuppressed patients. AB - We report the early results of laparoscopic incisional hernia repair in a small group of immunosuppressed patients and compare these results with a cohort of patients with open repair. We describe a modification used to secure the cephalad portion of the Gore-Tex mesh in high epigastric incisional hernias often encountered after liver transplantation. Data were gathered retrospectively for all incisional hernia repairs by our group from March 1996 to January 2001. Twelve of 13 attempted patients had successful completion of their laparoscopic hernia repairs with no reported recurrences to date. Two of these procedures were performed for recurrent hernias. We completed nine of nine attempted laparoscopic hernia repairs in liver transplant patients with epigastric incisional hernias. We repaired two of three attempted lower midline incisional hernias in renal disease patients. One of these patients was soon able to reuse his peritoneal dialysis catheter. A total of 15 patients, 12 with liver transplants, underwent open repair of their incisional hernias. These patients had seven recurrences and/or serious mesh infections with five patients electing repeated operations. In our initial series, laparoscopic mesh repair of incisional hernias is practical and safe in the abdominal organ transplant population with a low incidence of early recurrence and serious infections. PMID- 12118858 TI - The effect of pre-existing ischaemic heart disease on renal dysfunction in cardiac transplant recipients. AB - Renal dysfunction is a recognized complication of cardiac transplantation and can impact on the life expectancy of an already fragile population. A large proportion of these patients require transplantation because of the consequences of ischaemic heart disease (IHD) which, in turn, is often associated with ischaemic nephropathy. We studied the effect of IHD, diagnosed prior to transplantation, on the renal function of recipients who survived more than 6months after surgery. Of the 168 patients transplanted in a single centre over 15 years, 132 were included in the study. Renal dysfunction was defined as a serum creatinine consistently above 200 micromol/L (2.26 mg/dL). Analysis confirmed that IHD was an independent risk factor for developing renal impairment. In transplant recipients with IHD, closer monitoring is warranted to detect and prevent renal dysfunction or to retard its progression. PMID- 12118859 TI - Morbidity from congenital hepatic fibrosis after renal transplantation for autosomal recessive polycystic kidney disease. AB - Presentation of autosomal recessive polycystic kidney disease (ARPKD) ranges from severe renal impairment and a high mortality rate in infancy to older children and adolescents with minimal renal disease and complications of congenital hepatic fibrosis (CHF), cholangitis and portal hypertension. Renal transplantation improves prognosis but it is unclear whether CHF in transplanted children follows the same clinical course as in older children with less severe renal disease. The aim of this study was to evaluate morbidity from CHF in ARPKD post renal transplantation. Data were analyzed for six males and eight females, transplanted for ARPKD (mean age 8.3 years, range 1-22.3 years) at the University of Minnesota between 1972 and 1998. Follow-up was for a mean of 14.5 years (range 3.1-33.6 years). One and 5 years patient survival rates were 93% and 86%, respectively. Overall five patients (36%) died; 4/5 deaths were related to CHF. Causes of death were hepatic failure immediately post transplant (n = 1), septicemia related to bile duct dilatation (n = 3) and multiorgan failure (n = 1). One and 5years graft survival rates were 87% and 70%, respectively. One patient had a combined liver-kidney transplant and two were re-transplanted. Initial signs of CHF were splenomegaly (n = 5), hepatosplenomegaly (n = 4) and gastrointestinal bleed (n = 2). Progression of CHF through childhood included hypersplenism (n = 7), esophageal varices with gastrointestinal bleeding (n = 5) and bile duct dilatation (n = 5). Portal hypertension was treated with portosystemic shunt (n = 3), sclerotherapy (n = 2), banding of varices (n = 1) and transjugular intrahepatic portosystemic shunt (n = 1). Of the nine survivors (mean age 12.8 years) 78% have functioning grafts (one liver-kidney transplant), 63% have portal hypertension and 22% have asymptomatic biliary dilatation. Complications of CHF developed in 79% of children who received a renal transplant for ARPKD. Mortality related to CHF occurred in 29% and accounted for 80% (4/5) of the deaths. PMID- 12118860 TI - A comparison between recipients receiving matched kidney and those receiving mismatched kidney from the same cadaver donor. AB - The optimal allocation of cadaveric kidneys for transplantation with reference to human leukocyte antigen (HLA) match and sharing these organs to a distant center remains controversial. The current analysis was performed using the United Network for Organ Sharing (UNOS) database for cadaveric kidney transplants (Tx) between 1988 and 1997. The graft survivals of zero-mismatch (matched) kidneys with the mate (mismatched) kidneys were compared. There were 2385 donors and 4770 Tx. Significant differences in recipient demographics between matched and mismatched Tx were: fewer African-American race (AA) in the matched group (9.0% vs. 21.9%), higher number of previous Tx (25.5% vs. 14.8%) and elevated mean cold ischemia time (24.0 vs. 22.2 h). Post-Tx dialysis requirements were similar (22.8% vs. 24.1%, p = 0.62) and matched kidneys had to travel more distance (920 vs. 232 miles). Using a Cox model, the matched group had a decreased relative hazard of graft failure of 23.0% (p = 0.0002) or 35% (p < 0.0001) with and without censoring for death. There was significantly better graft survival in the matched recipients in all pairs except AA (matched) and non-AA (mismatched). For older donors (> or = 50 years, n = 1508), the matched grafts survival was marginally significant (p =0.05). Matched kidneys have improved survival compared with the mismatched kidneys despite the longer distance traveled. The benefit of mismatched transplants was predominantly seen in non-AA. PMID- 12118861 TI - Estimated and measured donor creatinine clearance are poor predictors of long term renal graft function and survival. AB - The objective of this study was to evaluate estimated and measured donor renal function in predicting graft function long-term and to identify donor criteria associated with nonacceptable graft prognosis. In 200 consecutive cadaver donors creatinine clearance was measured at explantation and estimated using the Cockcroft formula on admission serum creatinine. Graft function was evaluated in recipients (n = 387) by 24-h creatinine clearance regularly during 3 years after transplantation. Measured creatinine clearance correlated to some extent with long-term graft function, while Cockcroft estimation was slightly superior and similar to using donor age only. Kidneys from donors with intra-operative creatinine clearance < or = 55 mL/min (median 50 mL/min) produced acceptable recipient graft function of 48 mL/min at 3 years and 76% 3-year graft survival. Donor age > or =60 years resulted in clearance at 3 years of 29 mL/min and 78% 3 year graft survival; adding the criteria of admission Cockcroft < or =60 mL/min, graft function at 3 years (28 mL/min) and 3-year graft survival (76%) were similar. In conclusion, creatinine-based estimates of the functional capacity of the donor kidney, calculated or intra-operatively measured, do little to improve the ability of donor age alone to predict long-term allograft function after renal transplantation, and nonacceptable donors are not discriminated. PMID- 12118862 TI - Efficacy and toxicity of a protocol using sirolimus, tacrolimus and daclizumab in a nonhuman primate renal allotransplant model. AB - A regimen combining sirolimus, tacrolimus, and daclizumab has recently been shown to provide adequate immunosuppression for allogeneic islet transplantation in humans, but remains unproven for primarily vascularized allografts. We evaluated this regimen for renal allograft transplantation in mismatched nonhuman primates. Dosages of sirolimus and tacrolimus were adjusted for trough levels of 10-15 ng/mL and 4-6 ng/mL, respectively. Treated monkeys (n = 5) had significantly prolonged allograft survival, with a mean survival of 36 days vs. 7 days in untreated controls (n = 6, p = 0.008). Four of five treated animals, but none of the controls, developed fibrinoid vascular necrosis of the small intestine. A review of gut histology from animals on other immunosuppressive protocols performed by our laboratory suggested that these lesions were a result of sirolimus exposure. In summary, this regimen prolongs the survival of vascularized renal allografts, but is limited by profound GI toxicity in rhesus macaques. PMID- 12118863 TI - Increased beta-myosin heavy chain in acute cellular rejection following human heart transplantation. AB - BACKGROUND: Increased expression of smooth muscle and nonmuscle myosin heavy chains has been previously reported in animal models of cardiac allograft rejection. However, altered expression of beta-myosin heavy chain in human cardiac rejection has not been determined. METHODS: Two-dimensional (2D)-gel electrophoresis of endomyocardial biopsies taken from patients with (Grade 3A, n = 6) and without (Grade 0, n = 6) acute rejection were analyzed. Increased expression of two protein spots (MW approximately 12 kDa) were identified in the presence of acute rejection and were further characterized by mass spectrometry analysis. In patients who had acute rejection, protein expression was subsequently analyzed by immunoblotting on biopsies preceding, during, and following treatment of rejection. RESULTS: Mass spectrometric analysis of the protein spots detected 6 and 22 tryptic peptides, respectively. Protein sequence database search analysis identified the first protein as beta-myosin heavy chain and the second spot consisted of proteins of unidentified nature that may represent novel proteins. Immunoblotting analysis showed 1.4 x fold increase (p < 0.01) of protein expression of beta-myosin heavy chain expression in the presence of acute rejection. CONCLUSIONS: To our knowledge, this is the first 2D-gel study to describe increased expression of beta-myosin heavy chain and other proteins of unidentified nature in association with human cardiac allograft rejection. PMID- 12118864 TI - The perceived urgency of speech warnings: semantics versus acoustics. AB - The relationship between the semantics of words and the acoustics of the way they are spoken is explored. Actors spoke warning signal words in an urgent, nonurgent and monotone style, and participants rated the urgency of the words. Results showed effects for signal word and style of presentation. Acoustic analysis showed that the urgent words were spoken at higher frequency with a broader pitch range and were louder than the nonurgent or monotone words. These acoustic differences were used to synthesize artificial versions of signal words in urgent and nonurgent formats. The urgent words were rated as more urgent than the nonurgent words, a finding attributable to their differing acoustics. Within each speaking style the words were acoustically the same, yet effects for signal word were found, suggesting that semantics is also important in urgency perception. This research has implications for the design and implementation of speech warning systems, particularly those in which urgency mapping is required. PMID- 12118865 TI - Effect of a concurrent auditory task on visual search performance in a driving related image-flicker task. AB - The effect of a concurrent auditory task on visual search was investigated using an image-flicker technique. Participants were undergraduate university students with normal or corrected-to-normal vision who searched for changes in images of driving scenes that involved either driving-related (e.g., traffic light) or driving-unrelated (e.g., mailbox) scene elements. The results indicated that response times were significantly slower if the search was accompanied by a concurrent auditory task. In addition, slower overall responses to scenes involving driving-unrelated changes suggest that the underlying process affected by the concurrent auditory task is strategic in nature. These results were interpreted in terms of their implications for using a cellular telephone while driving. Actual or potential applications of this research include the development of safer in-vehicle communication devices. PMID- 12118866 TI - Contrast sensitivity in a dynamic environment: effects of target conditions and visual impairment. AB - Contrast sensitivity was determined as a function of target velocity (0 degrees - 120 degrees/s) over a variety of viewing conditions. In Experiment 1, measurements of dynamic contrast sensitivity were determined for 24 male and 24 female observers as a function of target velocity for letter stimuli of 2 sizes and 2 durations. Significant main effects were found for target velocity, target size, and target duration, but significant interactions among the variables indicated especially pronounced adverse effects of increasing target velocity for small targets and brief durations. In Experiment 2, the effects of simulated cataracts on dynamic contrast sensitivity were determined for 10 male and 10 female observers. Although the simulated impairment had no effect on traditional acuity scores, dynamic contrast sensitivity was markedly reduced under all conditions but especially with the smaller targets and at higher velocities. Results are discussed in terms of dynamic contrast sensitivity as a useful composite measure of visual functioning that may provide a better overall picture of an individual's visual functioning than does traditional static acuity, dynamic acuity, or contrast sensitivity alone. The measure of dynamic contrast sensitivity may increase understanding of the practical effects of various conditions, such as aging or disease, on the visual system, or it may allow improved prediction of individuals' performance in visually dynamic, situations, such as driving and sports. PMID- 12118867 TI - The far-anchor effect: errors in the perception of motion and implications for aviation safety. AB - The far-anchor effect is responsible for a motion-in-depth illusion that has only recently been recognized. When viewing conditions are limited, motions in depth of a farther target in a two-object display may readily be perceived as opposite motions in depth of the nearer target. The present studies determined whether this error could be avoided through controlled fixation or training with feedback. Under conditions of reduced visibility, participants (college students) viewed 64 two-target presentations varying in the position of the moving target and its direction of motion. Neither fixation instructions nor informational feedback about motion errors affected the occurrence of the basic illusion, nor did a vertical separation of the targets eliminate the main effect, indicating the robustness of the motion illusion under some relatively realistic variations. Such errors in judging motion in depth have significance for both midair collisions between aircraft and ground-incursion accidents under conditions of reduced visibility. Potential applications include the elaboration of examples used in pilot training programs or in training programs for ground personnel. PMID- 12118868 TI - Control performance with three translational degrees of freedom. AB - For multiple degree-of-freedom (DOF) systems, it is important to determine how accurately operators can control each DOF and what influence perceptual, information processing, and psychomotor components have on performance. Sixteen right-handed male students participated in 2 experiments: 1 involving positioning and 1 involving tracking with 3 translational DOFs. To separate perceptual and psychomotor effects, we used 2 control-display mappings that differed in the coupling of vertical and depth dimensions to the up-down and fore-aft control axes. We observed information processing effects in the positioning task: Initial error correction on the vertical dimension lagged in time behind the horizontal dimension. The depth dimension error correction lagged behind both, which was ascribed to the poorer perceptual information. We observed this perceptual effect also in the tracking experiment: Tracking error along the depth dimension was 3.8 times larger than along the other dimensions. Motor effects were also present, with tracking errors along the up-down axis of the hand controller being 1.1 times larger than along the fore-aft axis. These results indicate that all 3 components contribute to control performance. Actual applications of this research include interface design for remote control and virtual reality. PMID- 12118869 TI - Surface textures improve the robustness of stereoscopic depth cues. AB - This research develops design recommendations for surface textures (patterns of color on object surfaces) rendered with stereoscopic displays. In 3 method-of adjustment procedure experiments, 8 participants matched the disparity of a circular probe and a planar stimulus rendered using a single visible edge. The experiments varied stimulus orientation and surface texture. Participants more accurately matched the depth of vertical stimuli than that of horizontal stimuli, consistent with previous studies and existing theory. Participants matched the depth of surfaces with large pixel-to-pixel luminance variations more accurately than they did surfaces with a small pixel-to-pixel luminance variation. Finally, they matched the depth of surfaces with vertical line patterns more accurately than they did surfaces with horizontal-striped texture patterns. These results suggest that designers can enhance depth perception in stereoscopic displays, and also reduce undesirable sensitivity to orientation, by rendering objects with surface textures using large pixel-to-pixel luminance variations. PMID- 12118870 TI - Impact of aviation highway-in-the-sky displays on pilot situation awareness. AB - Thirty-six pilots (31 men, 5 women) were tested in a flight simulator on their ability to intercept a pathway depicted on a highway-in-the-sky (HITS) display. While intercepting and flying the pathway, pilots were required to watch for traffic outside the cockpit. Additionally, pilots were tested on their awareness of speed, altitude, and heading during the flight. Results indicated that the presence of a flight guidance cue significantly improved flight path awareness while intercepting the pathway, but significant practice effects suggest that a guidance cue might be unnecessary if pilots are given proper training. The amount of time spent looking outside the cockpit while using the HITS display was significantly less than when using conventional aircraft instruments. Additionally, awareness of flight information present on the HITS display was poor. Actual or potential applications of this research include guidance for the development of perspective flight display standards and as a basis for flight training requirements. PMID- 12118871 TI - Formal verification of human-automation interaction. AB - This paper discusses a formal and rigorous approach to the analysis of operator interaction with machines. It addresses the acute problem of detecting design errors in human-machine interaction and focuses on verifying the correctness of the interaction in complex and automated control systems. The paper describes a systematic methodology for evaluating whether the interface provides the necessary information about the machine to enable the operator to perform a specified task successfully and unambiguously. It also addresses the adequacy of information provided to the user via training material (e.g., user manual) about the machine's behavior. The essentials of the methodology, which can be automated and applied to the verification of large systems, are illustrated by several examples and through a case study of pilot interaction with an autopilot aboard a modern commercial aircraft. The expected application of this methodology is an augmentation and enhancement, by formal verification, of human-automation interfaces. PMID- 12118872 TI - Agreeing with automated diagnostic aids: a study of users' concurrence strategies. AB - Automated diagnostic aids that are less than perfectly reliable often produce unwarranted levels of disuse by operators. In the present study, users' tendencies to either agree or disagree with automated diagnostic aids were examined under conditions in which (a) the aids were less than perfectly reliable but aided-diagnosis was still more accurate that unaided diagnosis; and (b) the system was completely opaque, affording users no additional information upon which to base a diagnosis. The results revealed that some users adopted a strategy of always agreeing with the aids, thereby maximizing the number of correct diagnoses made over several trials. Other users, however, adopted a probability-matching strategy in which agreement and disagreement rates matched the rate of correct and incorrect diagnoses of the aids. The probability-matching strategy, therefore, resulted in diagnostic accuracy scores that were lower than was maximally possible. Users who adopted the maximization strategy had higher self-ratings of problem-solving and decision-making skills, were more accurate in estimating aid reliabilities, and were more confident in their diagnosis on trials in which they agreed with the aids. The potential applications of these findings include the design of interface and training solutions that facilitate the adoption of the most effective concurrence strategies by users of automated diagnostic aids. PMID- 12118873 TI - The effects of work pace on within-participant and between-participant keying force, electromyography, and fatigue. AB - A laboratory study was conducted to determine the effects of work pace on typing force, electromyographic (EMG) activity, and subjective discomfort. We found that as participants typed faster, their typing force and finger flexor and extensor EMG activity increased linearly. There was also an increase in subjective discomfort, with a sharp threshold between participants' self-selected pace and their maximum typing speed. The results suggest that participants self-select a typing pace that maximizes typing speed and minimizes discomfort. The fastest typists did not produce significantly more finger flexor EMG activity but did produce proportionately less finger extensor EMG activity compared with the slower typists. We hypothesize that fast typists may use different muscle recruitment patterns that allow them to be more efficient than slower typists at striking the keys. In addition, faster typists do not experience more discomfort than slow typists. These findings show that the relative pace of typing is more important than actual typing speed with regard to discomfort and muscle activity. These results suggest that typists may benefit from skill training to increase maximum typing speed. Potential applications of this research includes skill training for typist. PMID- 12118874 TI - Ecological interface design: progress and challenges. AB - Ecological interface design (EID) is a theoretical framework for designing human computer interfaces for complex sociotechnical systems. Its primary aim is to support knowledge workers in adapting to change and novelty. This literature review shows that in situations requiring problem solving, EID improves performance when compared with current design approaches in industry. EID has been applied to industry-scale problems in a broad variety of application domains (e.g., process control, aviation, computer network management, software engineering, medicine, command and control, and information retrieval) and has consistently led to the identification of new information requirements. An experimental evaluation of EID using a full-fidelity simulator with professional workers has yet to be conducted, although some are planned. Several significant challenges remain as obstacles to the confident use of EID in industry. Promising paths for addressing these outstanding issues are identified. Actual or potential applications of this research include improving the safety and productivity of complex sociotechnical systems. PMID- 12118875 TI - The perceived utility of human and automated aids in a visual detection task. AB - Although increases in the use of automation have occurred across society, research has found that human operators often underutilize (disuse) and overly rely on (misuse) automated aids (R. Parasuraman & V. Riley, 1997). Nearly 275 Cameron University students participated in 1 of 3 experiments performed to examine the effects of perceived utility (M. T. Dzindolet, H. P. Beck, L. G. Pierce, & L. A. Dawe, 2001) on automation use in a visual detection task and to compare reliance on automated aids with reliance on humans. Results revealed a bias for human operators to rely on themselves. Although self-report data indicate a bias toward automated aids over human aids, performance data revealed that participants were more likely to disuse automated aids than to disuse human aids. This discrepancy was accounted for by assuming human operators have a "perfect automation" schema. Actual or potential applications of this research include the design of future automateddecision aids and training procedures for operators relying on such aids. PMID- 12118876 TI - Relation between glare and driving performance. AB - The present study investigated the effects of discomfort glare on driving behavior. Participants (old and young; U. S. and Europeans) were exposed to a simulated low-beam light source mounted on the hood of an instrumented vehicle. Participants drove at night in actual traffic along a track consisting of urban, rural, and highway stretches. The results show that the relatively low glare source caused a significant drop in detecting simulated pedestrians along the roadside and made participants drive significantly slower on dark and winding roads. Older participants showed the largest drop in pedestrian detection performance and reduced their driving speed the most. The results indicate that the deBoer rating scale, the most commonly used rating scale for discomfort glare, is practically useless as a predictor of driving performance. Furthermore, the maximum U. S. headlamp intensity (1,380 cd per headlamp) appears to be an acceptable upper limit. PMID- 12118877 TI - Microarray profiling of Erwinia chrysanthemi 3937 genes that are regulated during plant infection. AB - Microarray technology was used to identify genes in Erwinia chrysanthemi 3937 that are specifically up- or down-regulated in a plant host compared with growth in laboratory culture medium. Several genes were plant down-regulated, and almost all of them were homologues of well-known housekeeping genes, such as those encoding metabolic functions, oxidative phosphorylation components, and transcription or translation processes. On the other hand, almost all of the plant up-regulated genes were involved with specialized functions, including already known or new putative virulence factors, anaerobiosis, iron uptake, transporters or permeases, xenobiotic resistance, chemotaxis, and stress responses to reactive oxygen species and heat. A substantial number of the plant up-regulated genes do not appear to be directly involved in damaging the host, but are probably important in adapting the pathogen to the host environment. We constructed insertion mutations in several of the plant up-regulated E. chrysanthemi 3937 genes. Among these, mutations of Bacillus subtilis pps1, Escherichia coli purU, and Pseudomonas aeruginosa pheC homologues reduced virulence on African violet leaves. Thus, new insights were obtained into genes important in bacterial virulence. PMID- 12118878 TI - The soybean GmN6L gene encodes a late nodulin expressed in the infected zone of nitrogen-fixing nodules. AB - Previously, we determined the N-terminal amino acid sequences of a number of putative peribacteroid membrane proteins from soybean. Here, we report the cloning of a gene, GmN6L, that encodes one of these proteins. The protein encoded by GmN6L is similar in sequence to MtN6, an early nodulin expressed in Medicago truncatula roots in response to infection by Sinorhizobium meliloti. The GmN6L gene was strongly expressed in mature nodules but not in other plant organs. GmN6L protein was first detected 2 weeks after inoculation with Bradyrhizobium japonicum and was limited to the infected zone of nodules. GmN6L protein was found in symbiosomes isolated from mature soybean nodules, both as a soluble protein and as a peripheral membrane protein bound to the peribacteroid membrane. These data indicate that GmN6L is a late nodulin, which is not involved in the infection process. Homology between GmN6L and FluG, a protein involved in signaling in Aspergillus nidulans, suggests that GmN6L may play a role in communication between the host and microsymbionts during symbiotic nitrogen fixation. PMID- 12118879 TI - The xanthomonas type III effector protein AvrBs3 modulates plant gene expression and induces cell hypertrophy in the susceptible host. AB - Xanthomonas campestris pv. vesicatoria bacteria expressing the type III effector protein AvrBs3 induce a hypersensitive response in pepper plants carrying the resistance gene Bs3. Here, we report that infection of susceptible pepper and tomato plants leads to an AvrBs3-dependent hypertrophy of the mesophyll tissue. Agrobacterium-mediated transient expression of the avrBs3 gene in tobacco and potato plants resulted in a similar phenotype. Induction of hypertrophy was shown to depend on the repeat region, nuclear localization signals, and acidic transcription activation domain (AAD) of AvrBs3, suggesting that the effector modulates the host's transcriptome. To search for host genes regulated by AvrBs3 in an AAD-dependent manner, we performed a cDNA-amplified fragment length polymorphism analysis of pepper mRNA populations. Thirteen AvrBs3-induced transcripts were identified and confirmed by reverse transcriptase-polymerase chain reaction. Sequence analysis revealed homologies to auxin-induced and expansinlike genes, which play a role in cell enlargement. These results suggest that some of the AvrBs3-induced genes may be involved in hypertrophy development and that xanthomonads possess type III effectors that steer host gene expression. PMID- 12118880 TI - Virulence and differential local and systemic spread of cucumber mosaic virus in tobacco are affected by the CMV 2b protein. AB - A mutant of the Cucumber mosaic virus subgroup IA strain Fny (Fny-CMV) lacking the gene encoding the 2b protein (Fny-CMVdelta2b) induced a symptomless systemic infection in tobacco. Both the accumulation of Fny-CMVdelta2b in inoculated tissue and the systemic movement of the virus appeared to proceed more slowly than for wild-type Fny-CMV. The influence of the 2b protein on virus movement in the inoculated leaf was examined using viral constructs derived from Fny-CMV and Fny-CMVdelta2b expressing the green fluorescent protein. Laser scanning confocal microscopy was used to visualize the movement of these viruses. Whereas the wild type virus spread between the epidermal cells as well as the mesophyll cells, the mutant virus spread less efficiently through the epidermal layer and moved preferentially through the mesophyll. Thus, the 2b protein of Fny-CMV influences the dynamics of movement of the virus both within the inoculated leaf and through the whole plant. We propose that this altered movement profile of Fny-CMVdelta2b results in the absence of disease symptoms in tobacco. PMID- 12118881 TI - Overexpression of Pto induces a salicylate-independent cell death but inhibits necrotic lesions caused by salicylate-deficiency in tomato plants. AB - Tomato plants overexpressing the disease resistance gene Pto (35S::Pto) exhibit spontaneous cell death, accumulation of salicylic acid (SA), elevated expression of pathogenesis-related genes, and enhanced resistance to a broad range of pathogens. Because salicylate plays an important role in the cell death and defense activation in many lesion mimic mutants, we investigated the interaction of SA-mediated processes and the 35S::Pto-mediated defense pathway by introducing the nahG transgene that encodes salicylate hydroxylase. Here, we show that SA is not required for the 35S::Pto-activated microscopic cell death and plays a minor role in defense gene activation and general disease resistance in 35S::Pto plants. In contrast, temperature greatly affects the spontaneous cell death and general resistance in 35S::Pto plants, and high temperature inhibits the cell death. The NahG tomato plants develop spontaneous, unconstrained necrotic lesions on leaves. These lesions also are initiated by the inoculation of a virulent strain of Pseudomonas syringae pv. tomato. However, the NahG-dependent necrotic lesions are inhibited in the NahG/35S::Pto plants. This inhibition is most pronounced under conditions favoring the 35S::Pto-mediated spontaneous cell death development. These results indicate that the signaling pathways activated by Pto overexpression suppress the cellular damage that is caused by SA depletion. We also found that ethylene is dispensable for the 35S::Pto-mediated general defense. PMID- 12118882 TI - Characterization of NADH dehydrogenases of Pseudomonas fluorescens WCS365 and their role in competitive root colonization. AB - The excellent-root-colonizing Pseudomonas fluorescens WCS365 was selected previously as the parental strain for the isolation of mutants impaired in root colonization. Transposon mutagenesis of WCS365 and testing for root colonization resulted in the isolation of mutant strain PCL1201, which is approximately 100 fold impaired in competitive tomato root colonization. In this manuscript, we provide evidence that shows that the lack of NADH dehydrogenase I, an enzyme of the aerobic respiratory chain encoded by the nuo operon, is responsible for the impaired root-colonization ability of PCL1201. The complete sequence of the nuo operon (ranging from nuoA to nuoN) of P. fluorescens WCS365 was identified, including the promoter region and a transcriptional terminator consensus sequence downstream of nuoN. It was shown biochemically that PCL1201 is lacking NADH dehydrogenase I activity. In addition, the presence and activity of a second NADH dehydrogenase, encoded by the ndh gene, was identified to our knowledge for the first time in the genus Pseudomonas. Since it was assumed that low-oxygen conditions were present in the rhizosphere, we analyzed the activity of the nuo and the ndh promoters at different oxygen tensions. The results showed that both promoters are up-regulated by low concentrations of oxygen and that their levels of expression vary during growth. By using lacZ as a marker, it was shown that both the nuo operon and the ndh gene are expressed in the tomato rhizosphere. In contrast to the nuo mutant PCL1201, an ndh mutant of WCS365 appeared not to be impaired in competitive root tip colonization. PMID- 12118883 TI - Analysis of molecular markers genetically linked to the Leptosphaeria maculans avirulence gene AvrLm1 in field populations indicates a highly conserved event leading to virulence on Rlm1 genotypes. AB - Map-based cloning of the avirulence gene AvrLm1 of Leptosphaeria maculans was initiated utilizing a genetic map of the fungus and a BAC library constructed from an AvrLm1 isolate. Seven polymorphic DNA markers closely linked to AvrLm1 were identified. Of these, two were shown to border the locus on its 5' end and were present, with size polymorphism, in both the virulent and the avirulent isolates. In contrast, three markers, J19-1.1, J53-1.3 (in coupling phase with avirulence), and Vir1 (in repulsion phase with avirulence), cosegregated with AvrLm1 in 312 progeny from five in vitro crosses. J19-1.1 and J53-1.3 were never amplified in the virulent parents or progeny, whereas Vir1 was never amplified in the avirulent parents or progeny. J19-1.1 and J53-1.3 were shown to be separated by 40 kb within a 184-kb BAC contig. In addition, the 1.6-cM genetic distance between J53-1.3 and the nearest recombinant marker corresponded to a 121-kb physical distance. When analyzing a European Union-wide collection of 192 isolates, J53-1.3, J19-1.1, and Vir1 were found to be closely associated with the AvrLm1 locus. The results of polymerase chain reaction amplification with primers for the three markers were in accordance with the interaction phenotype for 92.2% (J53-1.3), 90.6% (J19-1.1), and 88.0% (Vir1) of the isolates. In addition, genome organization of the AvrLm1 region was highly conserved in field isolates, because 89.1% of the avirulent isolates and 79.0% of the virulent isolates showed the same association of markers as that of the parents of in vitro crosses. The large scale analysis of field isolates with markers originating from the genetic map therefore confirms (i) the physical proximity between the markers and the target locus and (ii) that AvrLm1 is located in (or close to) a recombination-deficient genome region. As a consequence, map-based markers provided us with high-quality markers for an overview of the occurrence of race "AvrLm1" at the field scale. These data were used to propose hypotheses on evolution towards virulence in field isolates. PMID- 12118884 TI - Mapping of viral genomic regions important in cross-protection between strains of a potyvirus. AB - Cross-protection was tested between potato and tobacco strains of Potato virus A, a member of the genus Potyvirus (PVA), in tobacco plants. Cross-protection was effective only at the initiation of infection. The potato strains provided only weak cross-protection against the tobacco strain, whereas the tobacco strain provided strong cross-protection against potato strains. The tamarillo strain (TamMV) showed cross-protection phenotypes mostly resembling those of the potato strains. Chimera of the PVA strains were utilized to map viral genomic regions important for cross-protection. The coat protein (CP) encoding region and the helper component proteinase (HCpro) affected cross-protection and virus accumulation. An amino acid substitution at the CP N-terminus reduced virus accumulation and the ability to overcome cross-protection, whereas amino acid substitutions introduced to the HCpro increased virus accumulation and the ability to overcome cross-protection. Closer sequence relatedness between the protector and challenger isolate, as determined by the CP-encoding sequence, was correlated with an increased cross-protection ability. Cross-protection was not overcome by inoculation with nonencapsidated viral RNA. Thus, the differences in cross-protection abilities between PVA strains and chimera were not explained with the "re-encapsidation model" described for strains of Tobacco mosaic tobamovirus but may be associated with a virus infection-induced RNA silencing mechanism. PMID- 12118886 TI - Sensitivity of bacterial and fungal plant pathogens to the lytic peptides, MSI 99, magainin II, and cecropin B. AB - In vitro and leaf disk assays of bacterial and fungal plant pathogens were conducted using three cationic lytic peptides, MSI-99, magainin II (MII), and cecropin B (CB). Growth of bacterial organisms was retarded or completely inhibited by low concentrations of these lytic peptides. The peptides also significantly reduced germination of fungal spores and growth of mycelia; however, higher concentrations of peptides were needed to inhibit fungal growth compared with those needed to inhibit bacteria. The relative efficacy of the peptides depended on the microorganism tested, but CB was the most inhibitory to the majority of the bacteria and fungi assayed. MSI-99, a synthetic derivative of MII with increased positive charge, showed equal or two- to fivefold higher antibacterial activity compared to MII in the in vitro assays. MSI-99 was also superior to MII against the oomycete, Phytophthora infestans but was slightly inferior to MII in assays with the true fungi, Penicillium digitatum and Alternaria solani. In the leaf disk assays, pretreating spores of Alternaria solani and Phytophthora infestans with the peptides at concentrations as low as 10 microg per ml led to significant reductions in the size of early blight lesions and prevented development of any late blight lesions on tomato leaf disks. Our results from in vitro and leaf disk assays suggest that MSI-99 can be used as a transgene to generate tomato lines with enhanced resistance to bacterial and fungal diseases of this crop. PMID- 12118885 TI - Induction of trehalase in Arabidopsis plants infected with the trehalose producing pathogen Plasmodiophora brassicae. AB - Various microorganisms produce the disaccharide trehalose during their symbiotic and pathogenic interactions with plants. Trehalose has strong effects on plant metabolism and growth; therefore, we became interested to study its possible role in the interaction of Arabidopsis thaliana with Plasmodiophora brassicae, the causal agent of clubroot disease. We found that trehalose accumulated strongly in the infected organs (i.e., the roots and hypocotyls) and, to a lesser extent, in the leaves and stems of infected plants. This accumulation pattern of trehalose correlated with the expression of a putative trehalose-6-phosphate synthase (EC 2.4.1.15) gene from P. brassicae, PbTPS1. Clubroot formation also resulted in an induction of the Arabidopsis trehalase gene, ATTRE1, and in a concomitant increase in trehalase (EC 3.2.1.28) activity in the roots and hypocotyls, but not in the leaves and stems of infected plants. Thus, induction of ATTRE1 expression was probably responsible for the increased trehalase activity. Trehalase activity increased before trehalose accumulated; therefore, it is unlikely that trehalase was induced by its substrate. The induction of trehalase may be part of the plant's defense response and may prevent excess accumulation of trehalose in the plant cells, where it could interfere with the regulation of carbon metabolism. PMID- 12118887 TI - The presence of diverse IS elements and an avrPphD homologue that acts as a virulence factor on the pathogenicity plasmid of Erwinia herbicola pv. gypsophilae. AB - The pathogenicity of Erwinia herbicola pv. gypsophilae (Ehg) and Erwinia herbicola pv. betae (Ehb) is dependent on a native plasmid (pPATH(Ehg) or pPATH(Ehb)) that harbors the hrp gene cluster, genes encoding type III effectors, phytohormones, biosynthetic genes, and several copies of IS1327. Sequence analysis of the hrp-flanking region in pPATH(Ehg) (cosmid pLA150) revealed a cluster of four additional IS elements designated as ISEhel, ISEhe2, ISEhe3, and ISEhe4. Two copies of another IS element (ISEhe5) were identified on the upstream region of the indole-3-acetic acid operon located on the same cosmid. Based on homology of amino acids and genetic organization, ISEhe1 belongs to the IS630 family, ISEhe2 to the IS5 family, ISEhe3 and ISEhe4 to different groups of the IS3 family, and ISEhe5 to the IS1 family. With the exception of ISEhe4, one to three copies of all the other IS elements were identified only in pathogenic strains of Erwinia herbicola pv. gypsophilae and Erwinia herbicola pv. betae whereas ISEhe4 was present in both pathogenic and nonpathogenic strains. An open reading frame that exhibited high identity (89% in amino acids) to AvrPphD of Pseudomonas syringae pv. phaseolicola was present within the cluster of IS elements. An insertional mutation in the AvrPphDEh, reduced gall size in gypsophila by approximately 85%. In addition, remnants of known genes from four different bacteria were detected on the same cosmid. PMID- 12118888 TI - Potato gene Y-1 is an N gene homolog that confers cell death upon infection with potato virus Y. AB - ADG2 is a DNA sequence mapped to a resistance (R) gene-rich region at the distal end of chromosome XI in potato (Solanum tuberosum subsp. andigena). The gene, in which ADG2 represents the predicted nucleotide-binding domain (NBS), was cloned and characterized. The coding region of the gene (designated as Y-1) is 6,187 bp long and structurally similar to gene N that confers hypersensitive resistance to Tobacco mosaic virus in Nicotiana spp. Both belong to the TIR-NBS-LRR class of genes and show 57% identity at the amino acid sequence level. The introns of Y-1 were spliced as predicted from the sequence. Y-1 cosegregated with Ry(adg), a gene for extreme resistance to Potato virus Y (PVY) on chromosome XI, as tested in a potato-mapping population and with independent potato cultivars. Leaves of the transgenic potato plants expressing Y-1 under the control of Cauliflower mosaic virus 35S promoter developed necrotic lesions upon infection with PVY, but no significant resistance was observed, and plants were systemically infected with PVY. PMID- 12118889 TI - Rhizobium etli mutant modulates carbon and nitrogen metabolism in Phaseolus vulgaris nodules. AB - The aim of this study was to evaluate the biochemical events in root nodules which lead to increased yield when bean is inoculated with a Rhizobium etli mutant (CFN037) having increased respiratory capacity. CFN037-inoculated plants had 22% more nitrogen (N) than did wild-type (CE3)-inoculated plants. Root nodule enzymes involved in nodule carbon and nitrogen assimilation as well as in ureides and amides synthesis were assessed in plants inoculated with CFN037 and the CE3. Our results show that the xylem ureides content was lower while that of amino acids was higher in CFN037- compared with CE3-inoculated plants. Supporting these results, enzymes involved in ureide synthesis were reduced while activity of aspartate aminotransferase, glutamate synthase, sucrose synthase, and glucose-6-P dehydrogenase were increased in CFN037-induced nodules. Glutamate synthase and phosphoenolpyruvate carboxylase transcripts were detected early in the development of nodules induced by CFN037 compared with CE3. However, plants inoculated with strain CE3-vhb, which express the Vitreoscilla sp. hemoglobin and also displays increased respiratory capacity, did not have altered ureide transport in N2-fixing plants. The data suggest that inoculation with special selected mutant strains of R. etli can modulate nodule N assimilation and N transport compounds. PMID- 12118890 TI - Pseudomonas putida strain PCL1444, selected for efficient root colonization and naphthalene degradation, effectively utilizes root exudate components. AB - Previously, we have described the selection of a plant-bacterium pair that is efficient in rhizoremediating naphthalene pollution in microcosm studies. After repeated selection for efficient root tip colonization upon inoculation of seeds of grass cv. Barmultra and for stable and efficient growth on naphthalene, Pseudomonas putida PCL1444 was selected as the most efficient colonizer of Barmultra roots. Here, we report the analysis of Barmultra root exudate composition and our subsequent tests of the growth rate of the bacterium and of the expression of the naphthalene degradation genes on individual exudate components. High performance liquid chromatography analysis of the organic acid and sugar root-exudate components revealed that glucose and fructose are the most abundant sugars, whereas succinic acid and citric acid are the most abundant organic acids. Tn5luxAB mutants of PCL1444 impaired in naphthalene degradation appeared to be impaired in genes homologous to genes of the upper naphthalene degradation pathway present in various Pseudomonas strains and to genes of the lower pathway genes for naphthalene degradation in P. stutzeri. Highest expression for both pathways involved in naphthalene degradation during growth in minimal medium with the carbon source to be tested was observed at the start of the logarithmic phase. Naphthalene did not induce the upper pathway, but a different pattern of expression was observed in the lower pathway reporter, probably due to the conversion of naphthalene to salicylic acid. Salicylic acid, which is described as an intermediate of the naphthalene degradation pathway in many Pseudomonas strains, did induce both pathways, resulting in an up to sixfold higher expression level at the start of the logarithmic phase. When expression levels during growth on the different carbon sources present in root exudate were compared, highest expression was observed on the two major root exudate components, glucose and succinic acid. These results show an excellent correlation between successful naphthalene rhizoremediation by the Barmultra-P. putida PCL1444 pair and both efficient utilization of the major exudate components for growth and high transcription of the naphthalene catabolic genes on the major exudate components. Therefore, we hypothesize that efficient root colonizing and naphthalene degradation is the result of the applied colonization enrichment procedure. PMID- 12118891 TI - Management of hepatitis C infection in renal transplant recipients. PMID- 12118892 TI - Prevention of post-transplant cardiovascular disease--report and recommendations of an ad hoc group. PMID- 12118893 TI - Primed allospecific T cells prevent the effects of costimulatory blockade on prolonged cardiac allograft survival in mice. AB - Costimulatory blockade can induce long-term allograft survival in naive animals, but may not be as effective in animals with previously primed immune repertoires. We attempted to induce long-term graft survival in B10.D2 recipients of B10.A cardiac allografts using donor-specific transfusion (DST) plus anti-CD40 ligand antibody (alphaCD40L). Recipients were either naive mice, or mice previously primed to B10.A or third party alloantigens through engraftment and rejection of skin transplants. Untreated naive mice rejected cardiac transplants by day 15 and contained a high frequency of primed, donor-reactive T cells. Donor-specific transfusion/alphaCD4OL treatment of naive animals induced long-term graft survival associated with low frequencies of donor-reactive T cells. Previous priming of donor-specific T cells through rejection of B10.A, but not third party, skin grafts prevented the effects of DST/alphaCD40L on prolonging survival of B10.A hearts. Moreover, adoptive transfer of CD3+, CD4+ or CD8+ T cells from B10.A skin-graft-primed animals prevented the effects of DST/alphaCD40L. The data demonstrate that animals with immune repertoires containing previously primed, donor-reactive T cells are resistant to the effects of costimulatory blockade. The findings have important implications for ongoing, costimulatory blockade based trials in humans, whose T-cell repertoires are known to contain memory alloreactive T cells. PMID- 12118894 TI - Inability to induce tolerance through direct antigen presentation. AB - Both the direct and indirect antigen presentation pathways are important mechanisms for T cell-mediated allograft rejection. Studies using knockout mice and monoclonal antibodies have demonstrated that CD4+ T cells are both necessary and sufficient for the rejection of allogeneic tissues, including skin, heart, and islet. Furthermore, combined blockade of the CD28/B7 and CD154/CD40 costimulatory pathways induces tolerance in multiple CD4+ T-cell dependent allograft models. In this study, we addressed the T-cell requirement for costimulation in direct antigen presentation. We demonstrated that class II specific alloreactive T-cell receptor transgenic T cells were sufficient to mediate allograft rejection independent of costimulatory blockade. Analysis of the costimulatory capacity of different antigen presenting cell (APC) populations demonstrated that APCs resident within the donor skin, Langerhans cells, are potent stimulators not requiring CD28- or CD154-dependent costimulation for direct major histocompatibility complex (MHC) antigen presentation. These results complement previous work examining the role of costimulation on CD8+ T cells, supporting a model in which the effectiveness of costimulatory blockade in the setting of transplantation may be selective for the indirect pathway of MHC alloantigen presentation. PMID- 12118895 TI - Protective effects of recombinant human antithrombin III in pig-to-primate renal xenotransplantation. AB - Delayed rejection of pig kidney xenografts by primates is associated with vascular injury that may be accompanied by a form of consumptive coagulopathy in recipients. Using a life-supporting pig-to-baboon renal xenotransplantation model, we have tested the hypothesis that treatment with recombinant human antithrombin III would prevent or at least delay the onset of rejection and coagulopathy. Non-immunosuppressed baboons were transplanted with transgenic pig kidneys expressing the human complement regulators CD55 and CD59. Recipients were treated with an intravenous infusion of antithrombin III eight hourly (250 units per kg body weight), with or without low molecular weight heparin. Antithrombin treated recipients had preservation of normal renal function for 4-5 days, which was twice as long as untreated animals, and developed neither thrombocytopenia nor significant coagulopathy during this period. Thus, recombinant antithrombin III may be a useful therapeutic agent to ameliorate both early graft damage and the development of systemic coagulation disorders in pig-to-human xenotransplantation. PMID- 12118897 TI - Treatment of unresectable hepatoblastoma with liver transplantation in the pediatric population. AB - The purpose of our study was to evaluate the outcome of children who underwent liver transplantation as treatment for unresectable hepatoblastoma. We prospectively collected data on 311 consecutive liver transplants performed at Children's Medical Center of Dallas between October 1984 and November 2000. There were nine recipients (five boys, four girls) with a diagnosis of unresectable hepatoblastoma. Postoperative survival of those currently alive ranged from 6 months to 16 years (mean 6.4 years, median 7.7 years). All recipients received preoperative chemotherapy: 67% received postoperative chemotherapy. Mean AFP level prior to transplantation was 1 448000 ng/mL. Mean age at diagnosis was 0.81 years. Mean age at transplantation was 1.87 years. Only two patients experienced acute cellular rejection in the postoperative period. There was a total of three deaths and one recurrence. The only instance in which AFP levels did not decrease to low or undetectable levels post-transplantation was in the patient with recurrent tumor. Liver transplantation has an established role in the treatment of hepatoblastoma. It accounted for 3% of pediatric liver transplants, and provided the only opportunity for survival in otherwise incurable patients. Early diagnosis and treatment were found to be associated with better results. Response to chemotherapy may be an important factor influencing survival. Rising AFP levels after transplantation are associated with recurrence. PMID- 12118896 TI - Acute xenograft rejection mediated by antibodies produced independently of TH1/TH2 cytokine profiles. AB - There is substantial support for the hypothesis that T(H)1 cytokine responses are critical for the normal elaboration of allograft rejection. Recent studies by Wang et al. (1) underscore the importance of T(H)2 responses in xenograft rejection and revealed that T(H)1 cytokines, IL-12 and interferon-gamma (IFN gamma), can negatively regulate the development of humoral responses necessary for xenograft rejection. Their exceptional studies prompted us to test whether the ability of allografts to elicit cellular rejection and xenografts to induce humoral rejection also result from the differential ability to induce T(H)1 and T(H)2 responses. We compared the kinetics of antibody and cytokine (IFN-gamma and IL-4) production in C57BL/6 mice following allograft transplantation with BALB/c hearts and in C57BL/6 and BALB/c mice following transplantation with Lewis rat hearts. We also compared the ability of BALB/c mice, deficient in the ability to produce IL-4 or IFN-gamma, to reject xenografts and produce xenoantibodies. We observed that T(H)1/T(H)2 cytokine production minimally affected the kinetics of graft rejection but regulated the magnitude of IgG subclass production. Anti graft IgM played a critical role in initiating acute antibody-mediated xenograft rejection, and the production antigraft IgM was unaffected by IL-4 or IFN-gamma deficiency. In contrast to the report by Wang et al. (1), we conclude that antibody-mediated xenograft rejection in the concordant Lewis rat heart-to C57BL/6 mouse xenotransplantation model is dependent on anti-IgM production but independent of T(H) cytokine profiles. PMID- 12118898 TI - Recent trends in early outcome of adult patients after heart transplantation: a single-institution review of 251 transplants using standard donor organs. AB - Older age, prior transplantation, pulmonary hypertension, and mechanical support are commonly seen in current potential cardiac transplant recipients. Transplants in 436 consecutive adult patients from 1994 to 1999 were reviewed. There were 251 using standard donors in 243 patients (age range 18-69 years). To emphasize recipient risk, 185 patients who received a nonstandard donor were excluded from analysis. The indications for transplant were ischemic heart disease (n = 123, 47%), dilated cardiomyopathy (n = 82, 32%), and others (n=56, 21%). One hundred and forty-nine (57%) recipients were listed as status I; 5 and 6% were supported with an intra-aortic balloon and an assist device, respectively. The 30-d survival and survival to discharge were 94.7 and 92.7%, respectively; 1-year survival was 89.1%. Causes of early death were graft failure (n = 6), infection (n = 4), stroke (n = 4), multiorgan failure (n = 3) and rejection (n = 2). Predictors were balloon pump use alone (OR= 11.4, p =0.002), pulmonary vascular resistance > 4 Wood units (OR = 5.7, p = 0.007), pretransplant creatinine > 2.0 mg/dL (OR = 6.9, p = 0.004) and female donor (OR = 8.3, p = 0.002). Recipient age and previous surgery did not affect short-term survival. Heart transplantation in the current era consistently offers excellent early and 1-year survival for well selected recipients receiving standard donors. Early mortality tends to reflect graft failure while hospital mortality may be more indicative of recipient selection. PMID- 12118899 TI - Sampling strategy to calculate the cyclosporin-A area under the time concentration curve. AB - The complete area under the time-concentration curve (AUC) is considered to be the gold standard for cyclosporin-A (CyA) monitoring. However, complete AUC is time- and cost-consuming. For this reason, we reviewed 259 4-h AUC (AUC0-4) performed in 74 renal transplanted patients in order to construct an equation to calculate AUC0-4. All samples were drawn from one adult population 13 days following transplantation, in order to allow the cyclosporin metabolism to stabilize. Regression analysis was done either with each or with a combination of two variables. Cyclosporin-A blood concentration at the second hour after the oral dose (C2) was the best predictor of AUC0-4, where AUC0-4 = 451 +(2.73 x C2), R2 = 0.87, p<0.001. The combination of C1 and C2 only, offered a better mathematical improvement to the C2 equation. This equation was further validated in 33 other CyA pharmacokinetic profiles performed in eight patients who had not participated in the equation development. In this new population, the C2 equation excellently predicted the trapezoidal AUC0-4 (R2 = 0.81). Our data shows that C2 can be safely used to estimate AUC0-4. The C2 equation simplifies CyA monitoring because of its high-predictive value and clinical feasibility. PMID- 12118900 TI - Effects of sirolimus on lipids in renal allograft recipients: an analysis using the Framingham risk model. AB - This report describes the effects of sirolimus on plasma lipids, and uses the Framingham risk model to assess the clinical importance of these effects. Lipid data from two large controlled studies of 1295 renal transplant patients were analyzed retrospectively. Sirolimus 2 mg/day and 5 mg/day were compared with placebo or azathioprine, and administered concomitantly with steroids and cyclosporine over 12 months. Hypercholesterolemia and hypertriglyceridemia occurred in all treatment groups and were maximal at 2-3 months. The sirolimus groups evidenced higher lipid levels than the controls, but the elevations diminished over time. At 1 year, the patients given sirolimus 2 mg/day had a mean cholesterol level 17 mg/dL greater and a mean triglyceride level 59 mg/dL greater than the controls. Among the patients given sirolimus 5 mg/day, mean cholesterol was 30 mg/dL greater and mean triglycerides were 103 mg/dL greater than the controls. Treatment with statins and fibrates was effective in reducing cholesterol and triglyceride levels, respectively, in the sirolimus-treated patients. The Framingham risk model predicted that the 17 mg/dL elevation in cholesterol would increase the incidence of coronary heart disease (CHD) by 1.5 new cases per 1000 persons per year and CHD death by 0.7 events per 1000 persons per year. Lipid elevations observed in the sirolimus-treated patients were manageable, improved over time, and responded to lipid-lowering therapy. Based on the Framingham risk model, the CHD risks associated with these cholesterol elevations are small compared with the baseline risks of the transplant population. PMID- 12118901 TI - Ethnicity greatly influences cytokine gene polymorphism distribution. AB - Polymorphisms in the regulatory regions of cytokine genes are associated with high and low cytokine production and may modulate the magnitude of alloimmune responses following transplantation. Ethnicity influences allograft half-life and the incidence of acute and chronic rejection. We have questioned whether ethnic based differences in renal allograft survival could be due in part to inheritance of cytokine polymorphisms. To address that question, we studied the inheritance patterns for polymorphisms in several cytokine genes (IL-2, IL-6, IL-10, TNF alpha, TGF-beta, and IFN-gamma) within an ethnically diverse study population comprised of 216 Whites, 58 Blacks, 25 Hispanics, and 31 Asians. Polymorphisms were determined by allele-specific polymerase chain reaction and restriction fragment length analysis. We found striking differences in the distribution of cytokine polymorphisms among ethnic populations. Specifically, significant differences existed between Blacks and both Whites and Asians in the distribution of the polymorphic alleles for IL-2. Blacks, Hispanics and Asians demonstrated marked differences in the inheritance of IL-6 alleles and IL-10 genotypes that result in high expression when compared with Whites. Those of Asian descent exhibited an increase in IFN-gamma genotypes that result in low expression as compared to Whites. In contrast, we did not find significant ethnic-based differences in the inheritance of polymorphic alleles for TNF-alpha. Our results show that the inheritance of certain cytokine gene polymorphisms is strongly associated with ethnicity. These differences may contribute to the apparent influence of ethnicity on allograft outcome. PMID- 12118902 TI - Limited dose monoclonal IL-2R antibody induction protocol after primary kidney transplantation. AB - This study prospectively compared immunoprophylaxis with a single intraoperative dose (2 mg/kg) of monoclonal interleukin-2 receptor (IL-2R) antibody vs. noninduction in kidney transplant recipients treated with tacrolimus (FK 506), mycophenolate mofetil (MMF) and a prednisone-based immunosuppression regimen. One hundred recipients of first-kidney transplant were enrolled into the study to receive either anti-IL-2R monoclonal antibody, daclizumab (2 mg/kg intraoperatively, limited anti-IL-2R) or no induction (control). Each patient also received oral tacrolimus (dosed to target trough level 10-15 ng/mL), MMF (500 mg bid) and prednisone. The primary efficacy end-point was the incidence of biopsy proven acute rejection during the first 6 months post-transplant. The patients were also followed for 12-month graft function, and graft and patient survival rates. Other than the donor's age being significantly lower in the control group, both groups were comparable with respect to age, weight, gender, race, human leukocyte antigen (HLA)-DR mismatch, panel reactive antibody (%PRA), cold ischemic time, cytomegalovirus (CMV) status, causes of renal failure, and duration and modes of renal replacement therapy (RRT). During the first 6 months, episodes of first biopsy confirmed acute rejection was 3/50 (6%) in the limited anti-IL-2R group and 8/50 (16%) in the controls (p < 0.05). Twelve-month patient 100/98 (%) and graft survival 100/96 (%) were not statistically different. The group receiving limited anti-IL-2R did not have any adverse reactions. Our study demonstrates that a limited (single) 2 mg/kg immunoprophylaxis dose with monoclonal IL-2R antibody (daclizumab) when combined with tacrolimus/MMF/steroid allows significant reduction in early renal allograft rejection to the single digit level. The therapy with anti-IL-2R antibody is simple and is well tolerated. PMID- 12118903 TI - Darusentan: an effective endothelinA receptor antagonist for treatment of hypertension. AB - BACKGROUND: The antihypertensive efficacy and safety of darusentan, a new selective endothelin, antagonist was investigated. METHODS: In a multicenter randomized, double-blind, parallel-group, dose-response study, a 2-week placebo run-in period was followed by a 6-week treatment period and then a 2-week placebo withdrawal period. At baseline before darusentan therapy, the average blood pressure (BP) of the patient population studied was diastolic 103.49 (SD 3.55) and systolic 168.27 (SD 16.63) mm Hg. In total, 392 patients were randomized (darusentan 10 mg: 94 patients, 30 mg: 103 patients, 100 mg: 96 patients, placebo: 99 patients). RESULTS: Darusentan significantly reduced diastolic (mean difference to placebo: 10 mg: -3.7 mm Hg, 95% confidence interval (CI): -6.6, 0.9, P = .009; 30 mg: -4.9 mm Hg, 95% CI: -7.7, -2.2, P = .0005; 100 mg: -8.3 mm Hg, 95% CI: -11.1, -5.5, P = .0001) and systolic BP (mean difference to placebo: 10 mg: -6.0 mm Hg, 95% CI: -11.0, -0.9, P = .02; 30 mg: -7.3 mm Hg, 95% CI: - 12.3, -2.4, P = .004; 100 mg: - 11.3 mm Hg, 95% CI: -16.3, -6.2, P = .0001). Pulse rate remained unchanged in all groups. There was a trend toward more adverse events in the active treatment groups (placebo: 30.3%, 10 mg: 44.7%, 30 mg: 40.8%, 100 mg: 49.0%). Headache was the most commonly reported adverse event, with no relevant difference among treatments. Flushing and peripheral edema were seen in a dose-dependent fashion in the active treatment groups only. CONCLUSION: These data, the first, suggest the therapeutic benefit of selective endothelinA receptor antagonism in human hypertension. PMID- 12118904 TI - Angiotensin converting enzyme inhibition and angiotensin II AT1-receptor blockade reduce the levels of asymmetrical N(G), N(G)-dimethylarginine in human essential hypertension. AB - BACKGROUND: Asymmetrical N(G), N(G)-dimethylarginine (ADMA) is associated with impaired endothelium-dependent vasodilation in humans. METHODS: Twenty young, male, mildly hypertensive subjects were included in a randomized, double-blind, fourfold cross-over study with placebo, enalapril (20 mg/day), eprosartan (600 mg/day), or a combination of both drugs (10 and 300 mg/day, respectively) each over 1 week, followed by a 2-week wash-out phase. After each treatment phase, ADMA concentration was measured. RESULTS: ADMA concentration was 1.69+/-0.59 micromol/L in the placebo phase, and was significantly lower in the enalapril, eprosartan, and combination phases (1.41+/-0.29, 1.42+/-0.43, and 1.38+/-0.30 micromol/L, respectively; all P < 0.05 v placebo). Changes in ADMA levels were independent of the drugs' action on blood pressure (BP). CONCLUSIONS: Levels of ADMA were reduced with enalapril and eprosartan therapy. Our results suggest a specific action of these drugs on ADMA levels that is independent of BP. PMID- 12118905 TI - Ethnicity and unprovoked hypokalemia in the Atherosclerosis Risk in Communities Study. AB - BACKGROUND: Hypertension is more prevalent in the African American population when compared with the European American population in the United States. Unprovoked hypokalemia may lead to hypertension and is associated with several forms of recognized secondary hypertension. METHODS: We investigated the association of ethnicity with unprovoked hypokalemia in the second Atherosclerosis Risk in Communities (ARIC) study examination. Hypokalemia was defined as serum potassium <3.5 mmol/L. RESULTS: A statistically significant association was detected between ethnicity and unprovoked hypokalemia (odds ratio = 5.3; 95% confidence interval = 3.6, 7.7) with unprovoked hypokalemia more prevalent in African Americans both before and after adjustment for important covariates. The unadjusted prevalence for unprovoked hypokalemia was 2.6% for African Americans and 0.5% for European Americans. CONCLUSIONS: We found that the prevalence of unprovoked hypokalemia for African Americans in the ARIC cohort was more than five times that for European Americans. These data suggest that an increased awareness of hypokalemia and its etiology may be indicated for African Americans. PMID- 12118906 TI - Relationships between new risk factors and circadian blood pressure variation in untreated subjects with essential hypertension. AB - Recently a growing amount of interest has been focused on new risk factors for cardiovascular disease, such as insulin, leptin, homocysteine, and urinary albumin excretion (UAE). Furthermore, the absence of a nocturnal blood pressure (BP) decrease is emerging as an index for future target organ damage. In the present study we aimed to determine the relationship between these risk factors and circadian BP variations in essential hypertensive subjects. One hundred six patients, aged 54+/-7 years, with stage I-II untreated hypertension were classified as dippers and nondippers according to the diurnal variation of >10% between mean daytime and nighttime systolic BP (SBP) and diastolic BP (DBP) in 24 h noninvasive ambulatory BP monitoring. Venous blood samples were drawn for determination of insulin, leptin, and homocysteine plasma levels, whereas UAE was evaluated in three consecutive 24-h urine samples. Nondippers compared to dippers had significantly greater hemodynamic load and higher UAE (by 17 mg/24 h, P < .05). The two groups did not differ regarding serum insulin, plasma leptin, and homocysteine levels. In the entire population, leptin was positively correlated with age, body mass index, 24-h DBP, fasting serum insulin, and plasma homocysteine levels, whereas homocysteine levels were significantly related to 24 h SBP and DBP values. Multiple linear regression analyses revealed that only UAE was significantly related with nocturnal SBP and DBP decrease (P < .05 for both). These findings suggest that the increased UAE observed in nondipper hypertensive subjects possibly represents a useful indicator-for future target organ damage. PMID- 12118907 TI - Population-based studies improve outcome in hypertensive patients. AB - In a population-based study, three groups of individuals were examined: 334 hypertensive subjects (group 1) received 7 years of intervention therapy by a Hypertension Team (HyT); 418 subjects (group 2) were simply observed with blood pressure (BP) measurement on demand and given lifestyle advice; and 437 subjects (group 3) had no contact with HyT. Hypertension intervention therapy was then withdrawn, leaving patients to their general practitioners. After phase A, BP was significantly lower than baseline in group 1 (-14.2%, P < .01) and group 2 ( 12.4%, P < .01), whereas it was unchanged in group 3. Cerebrovascular (but not coronary) events were observed less in group 1 (fatal 2.7%, nonfatal 3.7%) than in group 2 (7.2% and 5.2%, respectively, both P < .001 v group 1) or group 3 (8.3% and 6.9%, respectively, both P < .001 v group 1). During a further 7 years of observation (phase B), no between-group differences in mortality were observed. We conclude that simple observation improves BP control, but that intervention is needed to reduce the incidence of stroke. PMID- 12118908 TI - Dopaminergic modulation of aldosterone secretions on changes of sodium intake in aldosterone-producing adenoma. AB - BACKGROUND: Impairment of renal dopamine excretion on high-salt diet (HS) may account for increased blood pressure (BP) in hypertensive subjects. Whether such impairment of dopaminergic activity exists in the adrenal gland is unknown. The purpose of this study was to differentiate degrees of dopaminergic inhibition of aldosterone secretion in patients with aldosterone-producing adenoma (APA). METHODS: A total of 15 patients with unilateral APA were fed a low-salt diet (LS) for 1 week, followed by another week on HS. At the end of each diet period, 24-h ambulatory BP recording, daily urine catecholamine measurement, and a metoclopramide test were performed. RESULTS: A high-salt diet increased both daytime and nighttime BP (P < .001), and urine dopamine excretion (P < .01). Intravenous injection of 10-mg metoclopramide increased plasma aldosterone concentrations (PAC) on both diets. The areas-under-the-curve for PAC between LS and HS were not different, but the area-under-the-curve for PAC increment was greater on HS (P < .05). Six patients with increment areas on HS greater than those on LS by 50% were termed "suppressible," and the remainders as "nonsuppressible." On HS, the so-called suppressible subjects had greater urine dopamine and less urine norepinephrine excretions (P < .05). The nonsuppressible subjects had greater percentage increase of nighttime BP by HS than the suppressible (for systolic BP, 13.1% v 4.5%, P < .01; for mean BP, 12.0% v 5.1%, P < .01, respectively), but no difference in daytime BP. CONCLUSIONS: Two subtypes of APA were defined according to their responses to metoclopramide during salt manipulation. On HS, the nonsuppressible subjects, with less dopaminergic inhibition of aldosterone secretion, had less urinary DA excretion and greater BP elevation. The renal and adrenal dopaminergic activities are regulated in a parallel fashion. PMID- 12118909 TI - Correlation between diastolic impairment and lipid metabolism in mild-to-moderate hypertensive postmenopausal women. AB - BACKGROUND: Many cardiovascular risk factors are found in hypertensive patients. The aim of this study was to evaluate the correlation between cardiac abnormalities (ie, diastolic and left ventricular hypertrophy) with other cardiovascular risk factors in postmenopausal women with hypertension. METHODS: A total of 200 consecutive postmenopausal women (mean age 47.5+/-4 years) with mild to-moderate hypertension that had never been treated were studied. Mean systolic pressure was 163+/-15 mm Hg and mean diastolic pressure 97+/-75 mm Hg. All subjects underwent M-mode two-dimensional echocardiography and cardiac Doppler. The following measurements were made: peak velocity of early left ventricular filling (E); peak velocity of late ventricular filling (A), and the ratio between early and late flow velocity peaks (E/A). The E/A ratio was then normalized for heart rate (E/Ac). Left ventricular mass index normalized for body surface was also measured. In each patient, total cholesterol, high-density lipoprotein (HDL), low-density lipoprotein (LDL), and plasma triglycerides were measured. We evaluated the correlation of E/Ac and left ventricular mass index (LVMI) with the following variables: total cholesterol, HDL, LDL, triglyceridemia, smoking status, systolic and diastolic blood pressure, and body mass index. RESULTS: A significant negative correlation with total cholesterol (r = -0.15, P < .05) and LDL (r = -0.20; P = .005), as well as a significant positive correlation with HDL (r = 0.20, P < .01) were found. No other variable was significantly correlated with E/A. There was no correlation between LVMI and any variable analyzed. CONCLUSION: In postmenopausal women with mild-to-moderate hypertension, high total cholesterol levels and low HDL levels are associated with impaired diastolic function. PMID- 12118910 TI - Linkage of left ventricular early diastolic peak filling velocity to chromosome 5 in hypertensive African Americans: the HyperGEN echocardiography study. AB - BACKGROUND: Altered diastolic filling is an important contributor to several cardiovascular disorders. Multiple lines of evidence suggest a genetic contribution to left ventricular (LV) diastolic filling; however, chromosomal locations harboring genes involved in impaired LV diastolic filling have not been reported. The aim of this study was to identify chromosomal regions contributing to variation of LV transmitral early and late peak filling velocities (E and A velocities), Doppler echocardiographic measures of LV diastolic filling. METHODS: We adjusted E and A velocities for age, age squared, heart rate, body mass index, systolic blood pressure, field center, and antihypertensive medication in sex- and ethnicity-specific linear regression models. Standardized residuals were calculated and used in multipoint variance components linkage analysis (GENEHUNTER). Anonymous markers (Cooperative Human Linkage Center set 8) were available for 167 white hypertensive sibships (397 subjects, mean age 60 years) and 182 African American (393 subjects, mean age 52 years) hypertensive sibships. RESULTS: For E velocity, linkage was detected on chromosome 5 at 133.6 centimorgan (cM) in African Americans (logarithm of the odds [LOD] = 4.13), and suggestive linkage (LOD >1.9) was observed for regions on chromosome 10 (80.8 cM) and chromosome 20 (1.5 cM). For A velocity, suggestive linkage was found for chromosome 12 (GATA85A04) in whites and for chromosome 8 (122.9 cM) in African Americans. Genes contained in and around the linked region are important candidates for diastolic filling (calcium-modulating cyclophilin ligand [5q23], alpha-1B adrenergic receptor [5q23-32]). CONCLUSION: Significant linkage was detected for LV early diastolic peak filling velocity on chromosome 5 in African Americans, indicating that a genomic region may contribute to interindividual variation in LV diastolic filling. PMID- 12118911 TI - T+31C polymorphism of angiotensinogen gene and nocturnal blood pressure decline: the Ohasama study. AB - BACKGROUND: We assessed the association between several polymorphisms of angiotensinogen gene (AGT) and essential hypertension using ambulatory blood pressure (BP). METHODS: We recruited 802 subjects in a rural Japanese community who were aged >40 years and who gave written informed consent for monitoring of their ambulatory BP and genetic analysis (the Ohasama Study). As a polymorphism of AGT, T+31C, which is in complete linkage disequilibrium with M235T, was determined using the TaqMan polymerase chain reaction method. RESULTS: The genotype distribution of AGT/T-+31C in the Ohasama Study was similar to that in another large Japanese population. Although there was no significant difference in 24-h and daytime ambulatory BP values, the nighttime BP was significantly lower in the subjects with TT, resulting in greater decline of nocturnal systolic (P = .090) and diastolic (P = .025) BP in subjects with TT. CONCLUSIONS: AGT/T+31C is associated with the circadian BP variation but not with BP level in the Japanese general population. PMID- 12118912 TI - The apolipoprotein E2 allele modulates activity and maximal velocity of the sodium-lithium countertransporter. AB - BACKGROUND: Alterations in erythrocyte sodium-lithium countertransport (SLC) activity and its maximal velocity (Vmax) are associated with hypertension and hypertriglyceridemia. The presence of apolipoprotein (apo) E variants is associated with hypertriglyceridemia. This study investigated the relationship between apoE phenotype and SLC kinetics. METHODS: Cardiovascular risk factors and SLC kinetics were measured in 171 subjects and 69 controls. Apolipoprotein E phenotypes were determined by Western blotting. RESULTS: Patients were 51% male, aged 56+/-13 years, with a blood pressure (BP) of 134+/-22/81+/-11 mm Hg, total cholesterol of 6.71+/-1.57 (256+/-61 mg/dL); median triglycerides 1.65 mmol/L (146 mg/dL) (range, 0.31 to 9.85 mmol/L; 27 to 872 mg/dL) and high-density lipoprotein (HDL) 1.39+/-0.43 mmol/L (54+/-16.6 mg/dL); fasting glucose 4.91+/ 0.61 mmol/L (88.5+/-11.0 mg/dL); median insulin 11.7 IU/L (range, 3.7 to 39.8 IU/L). Phenotype frequencies were E3/E3 56%, E2/E3 14%, E2/E2 1%, E3/E4 27%, and E4/E4 2%. The SLC activity, Vmax, and sodium affinity (Km) were not significantly different with respect to apoE phenotype in simple analysis by Kruskal Wallis test. However, in multiple regression analysis after exclusion of BP, a strong co correlate of SLC activity, the presence of an apoE2 allele was associated reduced activity (beta = -0.061; P = .01) along with HDL:apoA1 ratio (beta = -0.170; P < .001), whereas for the kinetic parameter Vmax, associations were found with triglyceride (beta = 0.029; P = .04), HDL:apoA1 ratio (beta = -0.186; P = .03) and the presence of an apoE2 allele (beta = -0.089; P = .04). CONCLUSIONS: These findings suggest that the apoE phenotype may modulate SLC activity and that the presence of an apoE2 allele phenotype is associated with lower SLC activity and Vmax. PMID- 12118913 TI - Leptin affects adenylate cyclase activity in H9c2 cardiac cell line: effects of short- and long-term exposure. AB - Leptin has been hypothesized to be a pathophysiologic link between obesity and cardiovascular diseases. Because the adenylate cyclase (AC) system is a main effector of beta-adrenergic receptors and leptin has been shown to modulate AC activity in other cell lines, a leptin impact on cardiac AC activity was hypothesized. Therefore, acute and chronic effects of leptin on a rat cardiac cell line (H9c2) were investigated. Leptin affected both basal (+ 13% at 30 min and -16.4% after 18 h v untreated cells) and catecholamine-stimulated AC activity (isoproterenol + leptin at 30 min or 18 h was +21% v untreated cells; norepinephrine + leptin at 30 min was +38.8% v untreated cells; and norepinephrine + leptin at 18 h was +6% v untreated cells). Thus, long-term leptin treatment was associated with a reduced AC activity and a different responsiveness to catecholamines. The AC activity on leptin treatment was accompanied by changes in levels of proteins structurally or functionally related to AC complex (AC, Gas, Gai, p21-ras). These data indicate that the AC complex is profoundly affected at more than one level by leptin treatment in the H9c2 cardiac cell line. Differences in AC activity after short- and long-term exposure to leptin and the interaction between leptin and catecholamine might provide further insight to the understanding of the development of hypertension and congestive heart failure in obese patients. PMID- 12118914 TI - Early onset of chondroitin sulfate and osteopontin expression in angiotensin II dependent left ventricular hypertrophy. AB - BACKGROUND: Chondroitin sulfate proteoglycan (CSPG) is expressed during embryonic heart development and osteopontin (OPN) is an important mediator of the profibrotic effects of angiotensin II (Ang II). The objective of this study was to analyze extracellular matrix protein (ECMP) expression in Ang II-dependent left ventricular (LV) hypertrophy (LVH), LV dysfunction, and to investigate right ventricular changes. METHODS: We used the hypertensive transgenic rat line TGR(mRen2)27 (Ren2), which provides a well-established model of Ang II-driven cardiac remodeling and progressive LV dysfunction and compared young Ren2 rats at the age of 10 weeks with normotensive Sprague-Dawley (SD) rats (n = 15, each group). RESULTS: Systolic blood pressure and LV weight were elevated in Ren2 compared to SD rats (P < .001). Left ventricular end-diastolic pressure was not altered in Ren2, but +dP/dt(max) and -dP/dt(max) were decreased in Ren2 compared to SD rats (P < .01). Cardiomyocyte widths, interstitial and perivascular fibrosis were increased in left and right ventricles of Ren2 in comparison to SD rats (P < .05). The LV mRNA expression of atrial natriuretic factor, OPN, and collagen I were increased in Ren2 as compared to SD rats (P < .05, respectively). The LV CSPG, collagen I, collagen III, fibronectin, laminin, and OPN contents were elevated in Ren2 compared to SD rats as measured by image analysis and Western blotting (P < .01). CONCLUSIONS: Reactivated expression of CSPG in the adult heart may be an important component of LV ECMP remodeling in LVH. Elevated cardiac OPN expression could mediate the alterations in LV ECMP pattern in Ang II dependent LVH, thus contributing to the development of contractile dysfunction in young Ren2 rats. PMID- 12118915 TI - Insights from Laragh's review course: the role of the renin-angiotensin system in blood pressure regulation. PMID- 12118916 TI - Obesity: its time has come. PMID- 12118917 TI - What can we do about the "epidemic" of obesity. PMID- 12118918 TI - Ethnicity and unprovoked hypokalemia in the ARIC Study. PMID- 12118919 TI - Blunted nighttime blood pressure dipping in postmenopausal women. PMID- 12118920 TI - Laparoscopic surgery for reflux esophagitis and paraesophageal hernia. AB - BACKGROUND: Laparoscopic surgery has become the standard for treatment of several abdominal diseases. We analyzed our data on laparoscopic treatment of reflux esophagitis and paraesophageal hernia. METHODS: Twenty patients (mean age 61 y; 14 men) - 18 with reflux esophagitis and sliding hiatus hernia, and two with paraesophageal and sliding hernia - were operated on using laparoscopy between March 1999 and March 2001. All patients were investigated by upper GI endoscopy, barium study and routine pre-operative work-up. Nineteen patients underwent a modified Nissen fundal wrap along with repair of the diaphragmatic crura; one patient had only crural repair with no fundal wrap. RESULTS: All procedures were completed laparoscopically. The mean operating time was 140 min (range 90 to 240). Eighteen patients were discharged on the third postoperative day and two on the fifth day. One patient had perforation of intrathoracic part of the esophagus during passage of an esophageal bougie; he presented with empyema 10 days after discharge and was treated by intercostal drainage. There were no other complications. All patients have been followed up on an outpatient basis for 3 months to 2 years. All are presently off acid-suppressive therapy. Seventeen patients are free of symptoms; two patients have gas bloat-like symptoms and one has occasional grade I dysphagia. CONCLUSIONS: Laparosopic surgery is a safe and effective method of treating esophagitis and paraesophageal hernia. PMID- 12118921 TI - Magnetic resonance cholangiopancreatography: evaluation in 150 patients. AB - BACKGROUND AND AIMS: Magnetic resonance cholangiopancreatography (MRCP) is a non invasive imaging technique for evaluating the biliary and pancreatic ducts. MRCP has reached a level of resolution and reliability where it may replace diagnostic endoscopic retrograde cholangio-pancreatography (ERCP). We analyzed the results of MRCP in adult patients with biliary or pancreatic disease, and compared the findings with those at surgery or on ERCP. METHODS: Data of 150 patients who underwent MRCP with both single slab and multislice rapid acquisition with relaxation enhancement sequences with half-fourier acquisition single-shot turbo spin echo techniques were analyzed. Patients were divided into four groups according to reason for referral for MRCP: obstructive jaundice (n = 65), chronic/acute pancreatitis (n = 25), screening prior to laparoscopic cholecystectomy (n = 20), and failed ERCP (n = 40). RESULTS: MRCP could accurately identify the level of biliary obstruction in 58 of 61 patients. Characterization of benign or malignant nature of a stricture was possible in 30 of 32 patients when findings of both MRCP and magnetic resonance imaging were analyzed together. MRCP revealed the morphology of the entire pancreatic duct in 13 of 15 patients having ductal changes on endoscopic retrograde pancreatography. CONCLUSION: MRCP has high sensitivity and specificity for detection of biliary dilatation, calculi, strictures and anatomical variants. PMID- 12118922 TI - Comparative analysis of duodenal biopsy tissue orientation supported on vegetable matrix versus filter paper. AB - BACKGROUND: Biopsy tissue may get disoriented if filter paper is used as a supporting medium. Use of vegetable matrix (cucumber) as a supporting medium may obviate the need of lifting the tissue while making blocks and thus avoid disorientation that occurs during this step. OBJECTIVE: To compare the orientation of duodenal biopsy tissue supported on vegetable matrix (cucumber) and on filter paper. METHODS: Over one year, 40 patients (20 with large-volume diarrhea, 20 with dyspepsia) were included in the study. Two pairs of duodenal biopsy tissues were obtained during gastroscopy; one pair was placed on filter paper, the other on vegetable matrix. Tissue and vegetable matrix were embedded together while making blocks, whereas the tissue had to be lifted off in case of filter paper. Sections were stained and assessed for crypt-villous alignment, parallel orientation of crypts and presence of visible muscularis mucosae with the help of a scoring system. RESULTS: Compared to biopsy tissue supported on filter paper, vegetable matrix-supported tissues were better oriented. Scores were rated as bad, good and very good in 8, 11 and 21 vegetable matrix-mounted tissues and in 21, 11 and 8 filter paper-mounted tissues, respectively. CONCLUSION: Duodenal biopsy tissue supported on vegetable matrix (cucumber) is better oriented than that on filter paper. PMID- 12118923 TI - Isolated caudate lobe resection for metastasis from rectal carcinoma. AB - Isolated caudate lobe excision is an uncommon procedure. We report a 41-year-old lady who underwent isolated caudate lobe excision for a solitary metastatic lesion from a previously operated adenocarcinoma of rectum. This is the first such reported case from India. PMID- 12118924 TI - Carbamazepine-induced eosinophilic enteritis. AB - We describe a 15-year-old boy who was on carbamazepine for the past 8 years for seizure disorder, who developed recurrent episodes of small bowel obstruction. Full-thickness small bowel biopsy obtained at laparotomy was consistent with eosinophilic enteritis. He improved clinically after tapering the drug. PMID- 12118926 TI - Rectal schwannoma. AB - Solitary schwannomas constitute only 2%-6% of stromal tumors of the gastrointestinal tract; the stomach is the preferred site of involvement. We report a 35-year-old woman who presented with difficulty in defecation. A mass on the posterior wall of the rectum was excised. Histologically, the lesion was a schwannoma, with S-100 proteins positivity and glial fibrillary acidic protein immunoreactivity. The patient is asymptomatic two years later. PMID- 12118925 TI - Ascites due to pacemaker syndrome. AB - We describe a 65-year-old man who had had a permanent cardiac pacemaker implanted in 1994 and presented now with ascites. He was investigated for 3 months without finding any cause. He was admitted with a colonic bleed due to a polyp; on cardiac evaluation he was found to have pacemaker syndrome. After adjusting the pacemaker settings his ascites and pleural effusion (which had developed later) disappeared. PMID- 12118927 TI - Carcinoma of sigmoid colon presenting as abdominal wall abscess. AB - Abdominal wall abscess is a rare presentation of intra-abdominal disease. We describe a 69-year-old woman with a locally advanced carcinoma of the sigmoid colon presenting as abdominal wall abscess. The diagnosis was suggested by computed tomography of the abdomen. She was treated with resection of the tumor with closure of the rectal stump and proximal end colostomy. No adjuvant therapy was undertaken considering the extent of the disease. She survived for four months after the operation. PMID- 12118928 TI - Association of Dubin-Johnson syndrome and portal vein thrombosis. AB - Dubin-Johnson syndrome is neither complicated by liver cell necrosis nor associated with portal hypertension. We report a 22-year-old man who had recurrent episodes of jaundice (conjugated hyperbilirubinemia) because of Dubin Johnson syndrome and portal hypertension secondary to portal vein thrombosis. The relationship between Dubin-Johnson syndrome and portal vein thrombosis in this case is most likely a chance occurrence. PMID- 12118929 TI - Acute hepatitis E due to travel to endemic area. PMID- 12118930 TI - Is iron deficiency anemia a common presenting manifestation of adult-onset celiac disease? PMID- 12118931 TI - Primary hypertrophic enteropathy. PMID- 12118932 TI - Common carotid artery occlusion causing cerebral infarction in ulcerative colitis. PMID- 12118933 TI - HBeAg-negative chronic hepatitis B. PMID- 12118934 TI - Therapeutic radiological interventional procedures in hepatocellular carcinoma. AB - BACKGROUND: To improve the survival rate of patients with hepatocellular carcinoma (HCC) in whom surgery is not possible, various methods have been developed employing angiographic and percutaneous techniques. We analyzed our experience with various percutaneous therapeutic interventional techniques done for HCC in our center. METHODS: Sixty-one patients with inoperable HCC (mean age 48.9 [SD 13.8] y; 47 men) were treated between January 1997 and December 2000 by transcatheter arterial chemoembolization (TACE) alone (22), TACE with percutaneous alcohol injection (PEI) (20), transcatheter arterial embolization (TAE) with steel coils and gel foam for gastrointestinal bleed (7), percutaneous radiofrequency ablation (1), percutaneous preoperative right portal vein embolization (3) and percutaneous preoperative tumor embolization to reduce blood loss at surgery (8). RESULTS: In 42 patients treated by TACE and PEI and TACE alone, tumor necrosis was scored; over 50% necrosis was seen only after six and nine months in both treatment groups. The survival rates after six and nine months and the median survival were similar in the two groups. Of 7 cases treated with TAE with steel coils and gel foam, the gastrointestinal bleeding stopped in four; in the other three, bleeding did not stop completely although less transfusion was required. In the patient treated by radiofrequency ablation, follow-up contrast-enhanced CT did not show enhancing tumor mass. We noted left lobe enlargement after percutaneous preoperative right portal vein embolization, prior to right hepatectomy. CONCLUSION: In patients with HCC not amenable to surgical intervention, a variety of percutaneous therapeutic interventional techniques may be used. PMID- 12118935 TI - Profile of hepatitis B e antigen-negative chronic hepatitis B. AB - BACKGROUND: Although chronic hepatitis B occurs in hepatitis B e antigen (HBeAg) negative patients, its prevalence and clinical significance are not known. AIM: To determine the prevalence and profile of HBeAg-negative chronic hepatitis B virus (HBV) infection. METHODS: A retrospective analysis of 363 consecutive patients (mean age 36 y; 288 men) with chronic HBV infection was performed. All patients were HBsAg-positive. Tests for liver profile, HBeAg and anti-HBe antibody were performed in all patients. Serum HBV DNA was tested using branched DNA assay in 245 patients. The patients were classified into three groups: no cirrhosis with normal ALT levels, no cirrhosis with elevated ALT levels, and clinical or histological evidence of cirrhosis. RESULTS: Of 363 patients, 141 (39%) were HBeAg-positive and 222 (61%) HBeAg-negative. Of HBeAg-negative patients, 120 (54%) had normal ALT, 45 (20%) had elevated ALT and 57 (26%) had evidence of cirrhosis; corresponding figures in the HBeAg-positive patients were 40 (28%), 66 (47%) and 35 (25%). HBV DNA was positive in 53 of 131 (40%) HBeAg negative patients tested; of these 53 patients, 9 (17%) had normal ALT, 20 (38%) had elevated ALT and 24 (45%) had cirrhosis. Thus, 72% of HBeAg-positive and 46% of HBeAg-negative patients had elevated ALT and/or cirrhosis. Among the latter group, 83% of HBV DNA-positive patients had elevated ALT and/or cirrhosis. Overall, 18% of HBsAg-positive patients had HBeAg-negative, HBV DNA-positive liver disease. CONCLUSION: HBeAg-negative chronic hepatitis B is not an uncommon and benign entity and chronic liver disease develops in a significant proportion of such patients. PMID- 12118936 TI - Microarray, SAGE and their applications to cardiovascular diseases. AB - The wealth of DNA data generated by the human genome project coupling with recently invented high-throughput gene expression profiling techniques has dramatically sped up the process for biomedical researchers on elucidating the role of genes in human diseases. One powerful method to reveal insight into gene functions is the systematic analysis of gene expression. Two popular high throughput gene expression technologies, microarray and Serial Analysis of Gene Expression (SAGE) are capable of producing large amounts of gene expression data with the potential of providing novel insights into fundamental disease processes, especially complex syndromes such as cardiovascular disease, whose etiologies are due to multiple genetic factors and their interplay with the environment. Microarray and SAGE have already been used to examine gene expression patterns of cell-culture, animal and human tissues models of cardiovascular diseases. In this review, we will first give a brief introduction of microarray and SAGE technologies and point out their limitations. We will then discuss the major discoveries and the new biological insights that have emerged from their applications to cardiovascular diseases. Finally we will touch upon potential challenges and future developments in this area. PMID- 12118937 TI - Cytopathological evaluations combined RNA and protein analyses on defined cell regions using single frozen tissue block. AB - The co-existence of multiple cell components in tissue samples is the main obstacle for precise molecular evaluation on defined cell types. Based on morphological examination, we developed an efficient approach for paralleled RNA and protein isolations from an identical histological region in frozen tissue section. The RNA and protein samples prepared were sufficient for RT-PCR and Western blot analyses, and the results obtained were well coincident each other as well as with the corresponding parameters revealed from immunohistochemical examinations. By this way, the sampling problem caused by cell-cross contamination can be largely avoided, committing the experimental data more specific to a defined cell type. These novel methods thus allow us to use single tissue block for a comprehensive study by integration of conventional cytological evaluations with nucleic acid and protein analyses. PMID- 12118938 TI - Overexpression of heme oxygenase-1 protects smooth muscle cells against oxidative injury and inhibits cell proliferation. AB - To investigate whether the expression of exogenous heme oxygenase-1 (HO-1) gene within vascular smooth muscle cells (VSMC) could protect the cells from free radical attack and inhibit cell proliferation, we established an in vitro transfection of human HO-1 gene into rat VSMC mediated by a retroviral vector. The results showed that the profound expression of HO-1 protein as well as HO activity was 1.8- and 2.0-fold increased respectively in the transfected cells compared to the non-transfected ones. The treatment of VSMC with different concentrations of H2O2 led to the remarkable cell damage as indicated by survival rate and LDH leakage. However, the resistance of the HO-1 transfected VSMC against H2O2 was significantly raised. This protective effect was dramatically diminished when the transfected VSMC were pretreated with ZnPP-IX, a specific inhibitor of HO, for 24 h. In addition, we found that the growth potential of the transfected cells was significantly inhibited directly by increased activity of HO-1, and this effect might be related to decreased phosphorylation of MAPK. These results suggest that the overexpression of introduced hHO-1 is potentially able to reduce the risk factors of atherosclerosis, partially due to its cellular protection against oxidative injury and to its inhibitory effect on cellular proliferation. PMID- 12118939 TI - Screen for stage-specific expression genes between tail bud stage and heartbeat beginning stage in embryogenesis of gynogenetic silver crucian carp. AB - A systemic study was initiated to identify stage-specific expression genes in fish embryogenesis by using suppression subtractive hybridization (SSH) technique. In this study, we presented a preliminary result on screen for stage specific expression genes between tail bud stage (TBS) and heartbeat beginning stage (HBS) in gynogenetic silver crucian carp (Carassius auratus gibelio). Two SSH plasmid libraries specific for TBS embryos and HBS embryos were constructed, and stage-specific expression genes were screened between the two stages. 1963 TBS positive clones and 2466 HBS positive clones were sampled to PCR amplification, and 1373 TBS and 1809 HBS PCR positive clones were selected to carry out dot blots. 169 TBS dot blot positive clones and 272 HBS dot blot positive clones were sequenced. Searching GenBank by using these nucleotide sequences indicated that most of the TBS dot blot positive clones could not be found homologous sequences in the database, while known genes were mainly detected from HBS dot blot positive clones. Of the 79 known genes, 20 were enzymes or kinases involved in important metabolism of embryonic development. Moreover, specific expressions of partial genes were further confirmed by virtual northern blots. This study is the first step for making a large attempt to study temporal and spatial control of gene expression in the gynogenetic fish embryogenesis. PMID- 12118940 TI - Recombinant scorpion insectotoxin AaIT kills specifically insect cells but not human cells. AB - The nucleotide sequence deduced from the amino acid sequence of the scorpion insectotoxin AaIT was chemically synthesized and was expressed in Escherichia coli. The authenticity of this in vitro expressed peptide was confirmed by N terminal peptide sequencing. Two groups of bioassays, artificial diet incorporation assay and contact insecticidal effect assay, were carried out separately to verify the toxicity of this recombinant toxin. At the end of a 24 h experimental period, more than 60% of the testing diamondback moth (Plutella xylostella) larvae were killed in both groups with LC50 value of 18.4 microM and 0.70 microM respectively. Cytotoxicity assay using cultured Sf9 insect cells and MCF-7 human cells demonstrated that the toxin AaIT had specific toxicity against insect cells but not human cells. Only 0.13 microM recombinant toxin was needed to kill 50% of cultured insect cells while as much as 1.3 microM toxin had absolutely no effect on human cells. Insect cells produced obvious intrusions from their plasma membrane before broken up. We infer that toxin AaIT bind to a putative sodium channel in these insect cells and open the channel persistently, which would result in Na+ influx and finally cause destruction of insect cells. PMID- 12118941 TI - The intracellular mechanism of alpha-fetoprotein promoting the proliferation of NIH 3T3 cells. AB - AIM: The existence and properties of alpha-fetoprotein (AFP) receptor on the surface of NIH 3T3 cells and the effects of AFP on cellular signal transduction pathway were investigated. METHODS: The effect of AFP on the proliferation of NIH 3T3 cells was measured by incorporation of 3H-TdR. Receptor-binding assay of 125I AFP was performed to detect the properties of AFP receptor in NIH 3T3 cells. The influences of AFP on the [cAMP]i and the activities of protein kinase A (PKA) were determined. Western blot was used to detect the change of K-ras P21 protein expression. RESULTS: The proliferation of NIH 3T3 cells treated with 0-80 mg/L of AFP was significantly enhanced. The Scatchard analysis indicated that there were two classes of binding sites with KD of 2.722 x 10(-9)M (Bmax=12810 sites per cell) and 8.931 x 10(-8)M (Bmax=119700 sites per cell) respectively. In the presence of AFP (20 mg/L), the content of cAMP and activities of PKA were significantly elevated . The level of K-ras P21 protein was upregulated by AFP at the concentration of 20 mg/L. The monoclonal antibody against AFP could reverse the effects of AFP on the cAMP content, PKA activity and the expression of K-ras p21 gene. CONCLUSION: The effect of AFP on the cell proliferation was achieved by binding its receptor to trigger the signal transduction pathway of cAMP-PKA and alter the expression of K- ras p21 gene. PMID- 12118942 TI - Get effective polyclonal antisera in one month. AB - According to the traditional immunization procedure, after the first injection of the sample A (emulsion of aimed antigen and Freund's complete adjuvant) to immunize rabbit, successive injections of the sample B (emulsion of aimed antigen and Freund's incomplete adjuvant) were followed every 2-4 weeks. In general, high titer of the corresponding polyclonal antisera will be observed after 4-5 injections of sample B in 3-4 months. This report presents a simply modified procedure that was able to stimulate the antisera formation in one month and achieve enough avidity to satisfy either Western blot or immunohistochemistry analysis. It just applied an additional injection of the sample A to the rabbit at the 3rd day after the primary immunization injection. You could gain the high titer of the antisera right after the first sample B injection in one month. This method has produced the desired results in three different recombinant antigens with different molecular weight (5.9 KD-55 KD) expressed from prokaryotic or eukaryotic cells. PMID- 12118943 TI - A developmental biological study of aldolase gene expression in Xenopus laevis. AB - We cloned cDNAs for Xenopus aldolases A, B and C. These three aldolase genes are localized on different chromosomes as a single copy gene. In the adult, the aldolase A gene is expressed extensively in muscle tissues, whereas the aldolase B gene is expressed strongly in kidney, liver, stomach and intestine, while the aldolase C gene is expressed in brain, heart and ovary. In oocytes aldolase A and C mRNAs, but not aldolase B mRNA, are extensively transcribed. Thus, aldolase A and C mRNAs, but not B mRNA, occur abundantly in eggs as maternal mRNAs, and strong expression of aldolase B mRNA is seen only after the late neurula stage. We conclude that aldolase A and C mRNAs are major aldolase mRNAs in early stages of Xenopus embryogenesis which proceeds utilizing yolk as the only energy source. aldolase B mRNA, on the other hand, is expressed only later in development in tissues which are required for dietary fructose metabolism. We also isolated the Xenopus aldolase C genomic gene (ca. 12 kb) and found that its promoter (ca. 2 kb) contains regions necessary for tissue-specific expression and also a GC rich region which is essential for basal transcriptional activity. PMID- 12118944 TI - Targeting epidermal Langerhans cells by epidermal powder immunization. AB - Immune reactions to foreign or self-antigens lead to protective immunity and, sometimes, immune disorders such as allergies and autoimmune diseases. Antigen presenting cells (APC) including epidermal Langerhans cells (LCs) play an important role in the course and outcome of the immune reactions. Epidermal powder immunization (EPI) is a technology that offers a tool to manipulate the LCs and the potential to harness the immune reactions towards prevention and treatment of infectious diseases and immune disorders. PMID- 12118945 TI - Iron deficiency, gastric atrophy and Helicobacter pylori. PMID- 12118946 TI - Dyspepsia and dyspepsia subgroups. Not all roads lead to Rome. PMID- 12118947 TI - Surveillance for colitis-associated cancer: we cannot stop now. PMID- 12118948 TI - Epidemiology of duodenal ulcer perforation: a study on hospital admissions in Norfolk, United Kingdom. AB - BACKGROUND: Studies on the incidence of perforated duodenal ulcer are limited and in the United Kingdom, data are largely based on findings observed over two decades ago. To provide updated epidemiological data on duodenal ulcer perforation, the incidence of the disease in Norfolk, United Kingdom was determined. METHODOLOGY: Medical records of patients with duodenal ulcer perforation were reviewed to confirm the diagnosis and obtain information on possible risk factors, namely, Helicobacter pylori infection, smoking and intake of non-steroidal anti-inflammatory drugs. The patients were admitted between 1 January 1996 and 31 December 1998, and were residents of Norfolk, United Kingdom. RESULTS: Sixty-eight cases of duodenal ulcer perforation were identified, 36 (52.9%) were males and 32 (47.1%) were females. The age-standardised incidence rate was 3.77 per 100,000 population per year (95% confidence interval 3.72 3.83). The mean age upon admission for all cases was 72.3 years (standard deviation: 17.8). The mean age for males was 67.7 years (standard deviation: 19.4) and for females 77.6 years (standard deviation: 15.7), which differed significantly (difference in means: 9.9, 95% confidence interval 1.5-18.3). There were 29 deaths (42.7%), of which 19 were females. After adjustment for covariates, the odds ratio of mortality in women was 4.57 (95% confidence interval 1.28-16.29). There were 25 (36.8%) smokers and 22 (32.4%) patients were non-steroidal anti-inflammatory drug users. Helicobacter pylori infection was assessed in only 14 (20.6%) patients; 2 were positive, 3 were negative, and in the rest the results were unrecorded. CONCLUSIONS: The incidence rates were lower compared to previous studies in the United Kingdom conducted in the 1960's and 1980's, which could reflect either improved health care or decreasing exposure to known risk factors. Furthermore, the difference in age distribution of incident cases between males and females may explain the higher mortality in females. PMID- 12118949 TI - Influence of Helicobacter pylori infection on severity of oesophagitis and response to therapy in the elderly. AB - BACKGROUND: The prevalence both of Helicobacter pylori infection and oesophagitis is higher in the elderly, than in adult and young populations. However the relationship between Helicobacter pylori infection and the clinical behaviour of oesophagitis has not yet been clarified. AIM: To evaluate the influence of Helicobacter pylori infection on the severity and clinical outcome after treatment of oesophagitis in elderly patients. METHODS: A total of 271 elderly patients (134 male, 137 female, mean age = 79.2 years, range 65-96) with grade 1 to 3 oesophagitis were studied. At baseline, the patients were divided into 3 groups according to Helicobacter pylori infection: Group 1 = 88 Helicobacter pylori-negative patients; Group 2 = 59 Helicobacter pylori-positive patients and Group 3 = 124 Helicobacter pylori-positive patients who underwent a one-week proton pump inhibitor-based triple therapy for the eradication of Helicobacter pylori infection. All patients were treated with proton pump inhibitors for two months; patients in Group 3 were also treated for one week with proton pump inhibitors plus two antibiotics. After two months, endoscopy and histology were repeated. RESULTS: At baseline, 32.5% of patients were Helicobacter pylori negative and 67.5% were Helicobacter pylori-positive. No baseline differences in severity of oesophagitis were found between Helicobacter pylori negative and positive patients. After proton pump inhibitor therapy, the complete resolution of oesophagitis was observed in 80.7% of Group 1, 76.3% of Group 2 and 75.8% of Group 3 (p=ns). Dividing patients also according to the severity of oesophagitis, no difference in healing rates between the three Groups were observed. CONCLUSIONS: In this elderly population, Helicobacter pylori infection did not influence the severity of oesophagitis at baseline or the response to short-term treatment with proton pump inhibitors. Furthermore, Helicobacter pylori eradication therapy did not influence the healing rate of oesophagitis. PMID- 12118951 TI - Cancer surveillance in ulcerative colitis: critical analysis of long-term prospective programme. AB - BACKGROUND: Patients with longstanding ulcerative colitis are at increased risk of colorectal cancer. In the literature, no agreement has yet been reached regarding prevention strategies. Our report sums up a prospective study started in 1980. METHODS: A total of 65 patients affected by ulcerative colitis for more than seven years were admitted to a regular colonoscopic and biopsy follow-up programme. RESULTS: Some 20 years after the beginning of the study, 23 (35.3%) patients have been operated upon, 2 patients have died but not from cancer 29 (44.66%) patients have abandoned the programme. Only 11 (16.9%) patients have remained under colonoscopic surveillance. CONCLUSION: These results cast some doubts on the significance of such a programme and on its long-term feasibility. PMID- 12118950 TI - Fluctuation in dyspepsia subgroups over time. A three-year follow-up of patients consulting general practice for dyspepsia. AB - BACKGROUND: General practitioners base their treatment and investigation on the symptoms presented by the patients. Subgroups of dyspepsia have been defined in order to guide management. AIM: To study prospectively changes over time in the presentation of dyspepsia according to different subtypes in a general practitioner population. SUBJECTS: Patients consulting the general practitioner because of dyspeptic complaints. METHODS: A random sample of general practitioner patients consulting with different dyspepsia subtypes (ulcer-like, reflux-like, dysmotility-like, uncharacteristic and relapsing dyspepsia) were studied three years after the initial consultation by postal questionnaires to the general practitioners (obtaining information from the patient records) and to the patients (obtaining self-reported symptoms during twelve months). The subtype of dyspepsia at baseline was compared to the subtype in the patient questionnaire. RESULTS: Between 20 and 34% of the patients reported no dyspepsia after three years, with so significant difference between the subtypes. Changes from one subtype to another were common, ulcer-like and reflux-like often changed into dysmotility-like dyspepsia. Dysmotility-like dyspepsia was significantly more stable over time. Patients with more than one episode of dyspepsia changed subtype significantly less. CONCLUSIONS: Most patients who presented with dyspepsia to the general practitioner still reported symptoms three years later. Few patients with dysmotility-like dyspepsia changed subtype over time, whereas changes from one subtype to another were common in other subtypes. This implies that dyspeptic patients could end up having dysmotility-like complaints possibly due to the lack of effective treatment for this condition, compared to the other dyspepsia subtypes. PMID- 12118952 TI - Acute pancreatitis in children. An Italian multicentre study. AB - AIM: To evaluate the clinical, morphological and aetiological aspects of acute pancreatitis in children in Italy. PATIENTS: The hospital records of 50 consecutive patients with acute pancreatitis observed in 5 Italian Pediatric Departments were reviewed. RESULTS: A total of 25 males and 25 females (median age 10.5 years, range 2-17) were studied. Of these patients, 48 (96%) had abdominal pain. The pancreatitis was associated with biliary disease in 10 patients (20%); it was due to viral infection in 6 patients (12%), pancreatic duct abnormalities in 4 (8%, familial chronic pancreatitis in 3 (6%), trauma in 5 (10%) and other causes in 5 (10%); the pancreatitis was of unknown origin in 17 patients (34%). Previous attacks of the disease had occurred in 14 patients. A diagnosis of mild pancreatitis was made in 41 patients (82%) and of severe disease in 9 (18%). One patient with severe pancreatitis died from multiorgan failure. Patients with severe pancreatitis had significantly higher serum concentrations of C-reactive protein than patients with mild pancreatitis. Hospital stay was similar for patients with the mild form and those with the severe form of the disease. CONCLUSIONS: In Italian children, acute pancreatitis is of unknown origin in about one-third of the children and is recurrent in 28% of the cases. The disease is severe in 18% of the cases. PMID- 12118953 TI - Adrenaline plus cyanoacrylate injection for treatment of bleeding peptic ulcers after failure of conventional endoscopic haemostasis. AB - BACKGROUND: Endoscopic therapy is a safe and effective method for treating non variceal upper gastrointestinal bleeding. However failure of therapy, in terms of continuing bleeding or rebleeding, is seen in up to 20%. Cyanoacrylate is a tissue glue used for variceal bleeding that has occasionally been reported as an alternative haemostatic technique in non-variceal haemorrhage. AIM: To retrospectively describe personal experience using cyanoacrylate injection in the management of bleeding ulcers after failure of first-line endoscopic modalities. PATIENTS AND METHODS: Between January 1995 and March 1998, 18 [12 M/6 F, mean age 68.1 years) out of 176 patients, referred to our Unit for non-variceal upper gastrointestinal bleeding, were treated with intralesional injection of adrenaline plus undiluted cyanoacrylate. Persistent bleeding after endoscopic haemostasis or early rebleeding were the indications for cyanoacrylate treatment. RESULTS: Definitive haemostasis was achieved in 17 out of 18 patients treated with cyanoacrylate. One patient needed surgery. No early or late rebleeding occurred during the follow-up. No complications or instrument lesions related to cyanoacrylate were recorded. CONCLUSIONS: In our retrospective series, cyanoacrylate plus adrenaline injection was found to be a potentially safe and effective alternative to endoscopic haemostasis when conventional treatment modalities fail in controlling bleeding from gastroduodenal ulcers. PMID- 12118954 TI - Endoscopic palliation of unresectable malignant oesophageal strictures with self expanding metal stents: comparing Ultraflex and Esophacoil stents. AB - BACKGROUND: Two types of self-expanding metal stents to palliate dysphagia in patients with unresectable malignant oesophageal strictures have been compared. METHODS: From February 1996 to October 2000, 50 metal stents (23 covered Ultraflex and 27 Esophacoil) were placed in 50 patients (40 males, mean age: 67+/ 12 years, range: 33-100, mean dysphagia score: 3.18+/-0.66) with unresectable malignant oesophageal strictures. Patients were followed until death. A retrospective review has been made of a prospectively collected database. RESULTS: The two groups were comparable as far as concerns degree of dysphagia, location and stricture length. Stent placement was successful in all cases. Covered Ultraflex stent was placed in 2 patients with oesophagobronchial fistula. No procedure-related deaths were seen. Early severe complications occurred in 2 patients (perforation in 1 and tumour bleeding in 1, in the Esophacoil group). Nine patients and 1 patient complained of pain following Esophacoil and Ultraflex stent placement, respectively. Late complications were asymptomatic rupture of distal Esophacoil rings in 2 patients, symptomatic Ultraflex stent migration in 2 and tumour overgrowth in 3 (Esophacoil 1, Ultraflex 2). Mean dysphagia score at 4 weeks after stent placement was 1.9+/-0.77. Mean survival was 177+/-109 days (range: 35-603 days). There were no significant differences in technical success, dysphagia palliation, complications (except chest pain) and survival using the two types of stent. CONCLUSIONS: Self-expanding metal stents are safe with high technical success and achieve satisfactory long-term palliation for dysphagia. The covered Ultraflex and Esophacoil stents are equally effective. PMID- 12118955 TI - Epidemiology of inflammatory bowel disease--methodological considerations. AB - The causes and mechanisms of action of inflammatory bowel disease have, so far, eluded discovery. Epidemiological studies have shown that ulcerative colitis tends to level off, whereas Crohn's disease tends to increase. Some of these changes may be due to diagnostic practices and increasing awareness of the disease and Crohn's colitis. The disease varies according to geographical location and a distribution along a north-south axis has been suggested. The differences may be due to study design, or may reflect differences in lifestyle, diet or be due to genetic predisposition triggered by environmental factors. Epidemiological studies designed to investigate such interactions may provide clues to its aetiology. Inflammatory bowel disease could, therefore, serve as a model for the importance of epidemiology when to test or reject the hypothesis of aetiology. PMID- 12118956 TI - Endoscopic retrograde cholangiopancreatography should no longer be used as a diagnostic test: the case in favour. AB - The requirement for diagnostic endoscopic retrograde cholangiopancreatography has decreased considerably in the past 10 years. Alternative imaging techniques are now available for the diagnosis of bile duct stones, pancreatic and biliary tract malignancy and inflammatory diseases such as sclerosing cholangitis and chronic pancreatitis. The imaging techniques include endoscopic ultrasonography, magnetic resonance cholangiopancreatography and helical computed tomography. There is good evidence that these techniques have an accuracy comparable to endoscopic retrograde cholangiopancreatography in the diagnosis of diseases of the bile and pancreatic ducts. All of these methods are less invasive than endoscopic retrograde cholangiopancreatography and have extremely low or negligible complication rates. The choice of technique used depends on local availability and expertise. In future, endoscopic retrograde cholangiopancreatography will be reserved exclusively for therapeutic applications. PMID- 12118957 TI - Endoscopic retrograde cholangiopancreatography should no longer be used as a diagnostic test: the case against. AB - Endoscopic retrograde cholangiopancreatography has been used successfully in diagnosis of pancreatico-biliary diseases. Over the past decade, there have been tremendous developments in radiological technologies which have led to the emergence of new and less invasive modalities like endoscopic ultrasound and spiral computed tomography and magnetic resonance cholangiopancreatography. Understandably, the usefulness of diagnostic endoscopic retrograde cholangiopancreatography is being questioned primarily because of its potential attendant complications. Therefore, this controversial question needs to be debated in the gastroenterology community. In this article, we compare the efficacy of newer diagnostic tools with existing "gold standard" modality- endoscopic retrograde cholangiopancreatography, and put forward our arguments in favour of a continuing role for endoscopic retrograde cholangiopancreatography as a diagnostic tool in certain circumstances. PMID- 12118958 TI - Endoscopic retrograde cholangiopancreatography should no longer be used as a diagnostic test: an independent verdict. PMID- 12118959 TI - Severe infection by varicella virus in an adult with ulcerative colitis: favourable response to acyclovir treatment. PMID- 12118960 TI - Lamivudine-induced muscle mitochondrial toxicity. PMID- 12118961 TI - Prokinetic effect of Alka-Seltzer in healthy subjects. PMID- 12118962 TI - Ciprofloxacin plus piperacillin compared with tobramycin plus piperacillin as empirical therapy in febrile neutropenic patients. A randomized, double-blind trial. AB - BACKGROUND: Therapy with an aminoglycoside and a beta-lactam remains common empirical therapy for febrile neutropenic patients. Concerns of aminoglycoside induced ototoxicity and nephrotoxicity have led to studies of alternate regimens. OBJECTIVE: To determine whether ciprofloxacin-piperacillin is equivalent to tobramycin-piperacillin as empirical therapy for neutropenic fever. DESIGN: Randomized, double-blind multicenter trial. SETTING: Seven U.S. university affiliated hospitals and one private research center. PATIENTS: Febrile (temperature >/= 38 degrees C), neutropenic (neutrophil level < 1 x 10(9) cells/L) hospitalized patients who had leukemia, lymphoma, or solid tumors, or were undergoing bone marrow transplantation. INTERVENTIONS: Patients received piperacillin, 50 mg/kg of body weight intravenously every 4 hours, and ciprofloxacin, 400 mg intravenously every 8 hours, or tobramycin, 2 mg/kg intravenously every 8 hours. MEASUREMENTS: Success was defined as resolution of infection and previously positive cultures without the need to give additional antimicrobial agents. RESULTS: 543 febrile episodes were evaluated, of which 471 were clinically evaluable (234 in the ciprofloxacin-piperacillin group and 237 in the tobramycin-piperacillin group). Success rates in the ciprofloxacin piperacillin group (63 of 234 febrile episodes) and tobramycin-piperacillin group (52 of 237 episodes) were similar (27% vs. 22%, respectively; difference, 5.0 percentage points [95% CI, -2.3 to 12.8 percentage points]), as was survival (96.2% of patients receiving ciprofloxacin-piperacillin versus 94.1% of patients receiving tobramycin-piperacillin; difference, 2.1 percentage points [CI, -2.2 to 6.4 percentage points]). Additions to the initial antimicrobial regimen were the most common reason for treatment failure in both groups (accounting for 67% of failures in the ciprofloxacin-piperacillin group and 72% in the tobramycin piperacillin group; difference, 5.0 percentage points [CI, -13.8 to 3.7 percentage points]). Fevers resolved faster in patients receiving ciprofloxacin piperacillin than in patients receiving tobramycin-piperacillin (mean, 5 vs. 6 days) (P = 0.005). No significant differences in adverse events or toxicity were noted (P = 0.083). CONCLUSION: Ciprofloxacin-piperacillin is as safe and effective as tobramycin-piperacillin for empirical therapy of neutropenic fever. PMID- 12118963 TI - Use of the electrocardiograph-based thrombolytic predictive instrument to assist thrombolytic and reperfusion therapy for acute myocardial infarction. A multicenter, randomized, controlled, clinical effectiveness trial. AB - BACKGROUND: Deciding which patients should receive thrombolytic therapy or percutaneous transluminal coronary angioplasty (PTCA) for acute myocardial infarction (AMI) can be difficult, especially for less-obvious candidates and when consulting physicians are off site. OBJECTIVE: To test whether the electrocardiograph-based Thrombolytic Predictive Instrument (TPI) improves use of thrombolytic and overall reperfusion therapy. DESIGN: 22-month randomized, controlled, clinical effectiveness trial. SETTING: Emergency departments at 28 urban, suburban, and rural hospitals in the United States. PATIENTS: Persons presenting to the emergency department with AMI and ST-segment elevation on an electrocardiogram (ECG). INTERVENTION: TPI predictions automatically printed on ECG text headers. MEASUREMENTS: Percentages of patients receiving thrombolytic therapy, thrombolytic therapy within 1 hour of initial ECG, and overall reperfusion (thrombolytic therapy or PTCA). RESULTS: Of 2875 patients with AMI, 1243 (43.2%) had ST-segment elevation. Of these, 1197 were randomly assigned to study groups; 732 (61.2%) had inferior AMI, and 465 (38.8%) had anterior AMI. A total of 60.5% of controls and 62.1% of TPI patients (P = 0.2) received thrombolytic therapy, 52.5% of controls and 53.3% of TPI patients received thrombolytic therapy within 1 hour (P > 0.2), and 67.6% of controls and 70.3% of TPI patients received overall reperfusion (P = 0.2). Of patients with inferior AMI in the control group versus the TPI group, 61.1% versus 67.6% (P = 0.03) received thrombolytic therapy, 53.2% versus 58.6% (P = 0.08) received thrombolytic therapy within 1 hour, and 67.7% versus 74.7% (P = 0.03) received overall reperfusion. Of patients with anterior AMI in the control group versus the TPI group, 59.5% versus 53.9% (P > 0.2) received thrombolytic therapy, 51.4% versus 45.3% (P > 0.2) received thrombolytic therapy within 1 hour, and 67.6% versus 63.8% (P > 0.2) received overall reperfusion. Among women (n = 398) in the control group versus the TPI group, 48.1% versus 58.2% (P = 0.03) received thrombolytic therapy, 40.5% versus 48.4% (P = 0.10) received thrombolytic therapy within 1 hour, and 55.7% versus 65.7% (P = 0.04) received overall reperfusion. Of patients who required physician consultation by telephone (n = 271) in the control group versus the TPI group, 47.3% versus 63.2% (P = 0.01) received thrombolytic therapy, 41.1% versus 53.6% (P = 0.04) received thrombolytic therapy within 1 hour, and 50.7% versus 66.4% (P = 0.01) received overall reperfusion. CONCLUSIONS: The TPI increased use of thrombolytic therapy, use of thrombolytic therapy within 1 hour, and use of overall coronary reperfusion by 11% to 12% for patients with inferior AMI, 18% to 22% for women, and 30% to 34% for patients with an off-site physician. Although its effect was minimal on patients with high baseline reperfusion rates, the TPI increased use and timeliness of reperfusion in often-missed groups and when involved physicians were off site. PMID- 12118964 TI - Cost-effectiveness analyses of colorectal cancer screening: a systematic review for the U.S. Preventive Services Task Force. AB - PURPOSE: To perform a systematic review of the cost-effectiveness of colorectal cancer screening for the U.S. Preventive Services Task Force. DATA SOURCES: MEDLINE and the British National Health Service Economic Evaluation Database, January 1993 through September 2001. STUDY SELECTION: Original economic evaluations of colorectal cancer screening in average-risk patients were reviewed. The authors sought studies addressing the incremental cost effectiveness of different screening strategies compared with no screening, of different screening strategies compared with one another, and of different ages of screening initiation and cessation. Two investigators independently reviewed each abstract, and potentially eligible articles were retrieved. A four-member working group reached consensus regarding final inclusion or exclusion of articles. DATA EXTRACTION: One reviewer extracted data into evidence tables. The results were checked by other members and discrepancies resolved by consensus. DATA SYNTHESIS: Among 180 potential articles identified, 7 were retained in the final analysis. Compared with no screening, cost-effectiveness ratios for screening with any of the commonly considered methods were generally between 10, 000 dollars and 25, 000 dollars per life-year saved. No one strategy was consistently found to be the most effective or to have the best incremental cost effectiveness ratio. Currently available models provided insufficient evidence to determine optimal starting and stopping ages for screening. CONCLUSIONS: Screening for colorectal cancer appears cost-effective compared with no screening, but a single optimal strategy cannot be determined from the currently available data. Additional data regarding adherence with screening over time, complication rates in real-world settings, and colorectal cancer biology are needed. Additional analyses are necessary to determine optimal ages of initiation and cessation. PMID- 12118966 TI - Unexpected hypoglycemia in a critically ill patient. AB - Administration of the wrong medication is a serious and understudied problem. Because physicians are not directly involved in the drug administration process, they tend to overlook the possibility of adverse drug events and medication errors in their differential diagnoses of patient illnesses or acute deterioration. This article analyzes the case of a patient with iatrogenic hypoglycemia due to administration of the wrong medication: Insulin instead of heparin was used to flush the patient's arterial line. In addition to assessing the results of the institution's "root-cause analysis" of the factors contributing to this particular adverse event and the institution's response, this article reviews the literature on preventing medication errors. Key strategies that might have been helpful in this case include using checklists for common emergency conditions (such as altered level of consciousness) and automated paging for "panic laboratory values," as well as instituting protocols for medication administration. Changing the system of administering medications by bar coding drugs, with checks of the medication, patient, and provider, could have prevented this accident. Finally, organizations need to strive for a "culture of safety" by providing opportunities to discuss errors and adverse events in constructive, supportive environments and by resisting pressure to find a scapegoat. PMID- 12118965 TI - Safety and efficacy of liposomal amphotericin B compared with conventional amphotericin B for induction therapy of histoplasmosis in patients with AIDS. AB - BACKGROUND: In patients with moderate to severe histoplasmosis associated with AIDS, the preferred treatment has been the deoxycholate formulation of amphotericin B. However, serious side effects are associated with use of amphotericin B. OBJECTIVE: To compare amphotericin B with liposomal amphotericin B for induction therapy of moderate to severe disseminated histoplasmosis in patients with AIDS. DESIGN: Randomized, double-blind, multicenter clinical trial. SETTING: 21 sites of the U.S. National Institute of Allergy and Infectious Diseases Mycoses Study Group. PATIENTS: 81 patients with AIDS and moderate to severe disseminated histoplasmosis. MEASUREMENTS: Clinical success, conversion of baseline blood cultures to negative, and acute toxicities that necessitated discontinuation of treatment. RESULTS: Clinical success was achieved in 14 of 22 patients (64%) treated with amphotericin B compared with 45 of 51 patients (88%) receiving liposomal amphotericin B (difference, 24 percentage points [95% CI, 1 to 52 percentage points]). Culture conversion rates were similar. Three patients treated with amphotericin B and one treated with liposomal amphotericin B died during induction (P = 0.04). Infusion-related side effects were greater with amphotericin B (63%) than with liposomal amphotericin B (25%) (P = 0.002). Nephrotoxicity occurred in 37% of patients treated with amphotericin B and 9% of patients treated with liposomal amphotericin B (P = 0.003). CONCLUSION: Liposomal amphotericin B seems to be a less toxic alternative to amphotericin B and is associated with improved survival. PMID- 12118967 TI - Reforming care for persons near the end of life: the promise of quality improvement. AB - Most people in developed countries will live with a serious, eventually fatal, chronic condition for months or years before dying; yet, the delivery of health care services has only just recently begun adapting to this reality. Quality improvement methods have been effective in helping clinical services to make substantial changes quickly. Quality improvement requires stating an aim, measuring success, and testing possible improvements. The testing of changes requires a clinical team to Plan, Do, Study, and Act on new insights (the "PDSA cycle"). Repeated PDSA cycles generate deep understanding of complex systems and make sustainable improvements rapidly. This paper discusses a composite case study in a nursing home setting, which builds on experience with multisite collaborative efforts and introduces quality improvement methods in the context of end-of-life care. PMID- 12118968 TI - Fever, neutropenia, and the second law of thermodynamics. PMID- 12118969 TI - Indwelling urinary catheters: a one-point restraint? PMID- 12118971 TI - Screening for colorectal cancer: recommendation and rationale. AB - This statement summarizes the current U.S. Preventive Services Task Force (USPSTF) recommendation on screening for colorectal cancer and the supporting scientific evidence and updates the 1995 recommendations contained in the Guide to Clinical Preventive Services, 2nd edition. At that time, the USPSTF recommended screening for colorectal cancer with annual fecal occult blood testing, periodic sigmoidoscopy, or the combination of fecal occult blood testing and sigmoidoscopy but concluded that the evidence was insufficient to recommend for or against colonoscopy or barium enema. The complete USPSTF recommendation and rationale statement on this topic, which includes a brief review of the supporting evidence, is available through the USPSTF Web site (http://www.preventiveservices.ahrq.gov), the National Guideline Clearinghouse (http://www.guideline.gov), and in print through the Agency for Healthcare Research and Quality Publications Clearinghouse (telephone, 800-358-9295; e-mail, ahrqpubs@ahrq.gov). The complete information on which this statement is based, including tables and references, is available in the accompanying article in this issue and in the summary of the evidence and systematic evidence review on the Web sites already mentioned. PMID- 12118972 TI - Screening for colorectal cancer in adults at average risk: a summary of the evidence for the U.S. Preventive Services Task Force. AB - PURPOSE: To assess the effectiveness of different colorectal cancer screening tests for adults at average risk. DATA SOURCES: Recent systematic reviews; Guide to Clinical Preventive Services, 2nd edition; and focused searches of MEDLINE from 1966 through September 2001. The authors also conducted hand searches, reviewed bibliographies, and consulted context experts to ensure completeness. STUDY SELECTION: When available, the most recent high-quality systematic review was used to identify relevant articles. This review was then supplemented with a MEDLINE search for more recent articles. DATA EXTRACTION: One reviewer abstracted information from the final set of studies into evidence tables, and a second reviewer checked the tables for accuracy. Discrepancies were resolved by consensus. DATA SYNTHESIS: For average-risk adults older than 50 years of age, evidence from multiple well-conducted randomized trials supported the effectiveness of fecal occult blood testing in reducing colorectal cancer incidence and mortality rates compared with no screening. Data from well conducted case-control studies supported the effectiveness of sigmoidoscopy and possibly colonoscopy in reducing colon cancer incidence and mortality rates. A nonrandomized, controlled trial examining colorectal cancer mortality rates and randomized trials examining diagnostic yield supported the use of fecal occult blood testing plus sigmoidoscopy. The effectiveness of barium enema is unclear. Data are insufficient to support a definitive determination of the most effective screening strategy. CONCLUSIONS: Colorectal cancer screening reduces death from colorectal cancer and can decrease the incidence of disease through removal of adenomatous polyps. Several available screening options seem to be effective, but the single best screening approach cannot be determined because data are insufficient. PMID- 12118973 TI - On being a doctor. Doctor's Luck. PMID- 12118974 TI - Physicians and joint negotiations. PMID- 12118975 TI - Blue light and milk. PMID- 12118976 TI - Fruit and vegetable intake and coronary heart disease. PMID- 12118977 TI - Questioning the treatment of venous thromboembolism. PMID- 12118981 TI - Rapid-onset type 1 diabetes mellitus without pancreatic exocrine dysfunction. PMID- 12118978 TI - Fruit and vegetable intake and coronary heart disease. PMID- 12118982 TI - Hepatotoxicity after prophylaxis with a nevirapine-containing antiretroviral regimen. PMID- 12118984 TI - Summaries for patients. Treatment of fever in hospitalized patients with low white blood cell counts. PMID- 12118985 TI - Summaries for patients. A decision aid for recognizing and treating heart attacks quickly. PMID- 12118986 TI - Summaries for patients. Screening for colorectal cancer: recommendations from the United States Preventive Services Task Force. PMID- 12118987 TI - Summaries for patients. Treatment of histoplasmosis in patients with HIV infection. PMID- 12118988 TI - Poly(dimethylsiloxane) as a material for fabricating microfluidic devices. AB - This Account summarizes techniques for fabrication and applications in biomedicine of microfluidic devices fabricated in poly(dimethylsiloxane) (PDMS). The methods and applications described focus on the exploitation of the physical and chemical properties of PDMS in the fabrication or actuation of the devices. Fabrication of channels in PDMS is simple, and it can be used to incorporate other materials and structures through encapsulation or sealing (both reversible and irreversible). PMID- 12118989 TI - Taming reactive phenol tautomers and o-quinone methides with transition metals: a structure-reactivity relationship. AB - Quinone methides act as important intermediates in organic syntheses, as well as in chemical and biological processes; however, examples of such isolated species are scarce as a result of their high reactivity. Phenol tautomers (keto form of phenol) are also important intermediates in several organic and organometallic reactions; nevertheless, isolated complexes are rare. This Account reviews the recent progress on the synthesis and reactivity of iridium and rhodium o-quinone methide complexes as well as on iridium-mediated ortho functionalization of phenols. This reaction was at the origin of the discovery of a general synthetic procedure to prepare the first metal-stabilized o-quinone methide. PMID- 12118990 TI - Crystal engineering of NLO materials based on metal--organic coordination networks. AB - Crystal engineering, the ability to predict and control the packing of molecular building units in the solid state, has attracted much attention over the past three decades owing to its potential exploitation for the synthesis of technologically important materials. We present here the development of crystal engineering strategies toward the synthesis of noncentrosymmetric infinite coordination networks for use as second-order nonlinear optical (NLO) materials. Work performed mainly in our laboratory has demonstrated that noncentrosymmetric solids based on infinite networks can be rationally synthesized by combining unsymmetrical bridging ligands and metal centers with well-defined coordination geometries. Specifically, coordination networks based on 3D diamondoid and 2D grid structures can be successfully engineered with a high degree of probability and predictability to crystallize in noncentrosymmetric space groups. We have also included noncentrosymmetric solids based on 1D chains and related helical structures for comparison. PMID- 12118991 TI - Proton transfer from alkane radical cations to alkane molecules: selectivity with respect to the site of proton donation and proton acceptance. AB - Information on the donor and acceptor site selectivity in the proton transfer from n-alkane radical cations to n-alkane molecules is gathered from gamma irradiated frozen CCl(3)F/n-alkane solutions (symmetric transfer) and mixed n alkane crystals (asymmetric transfer) by EPR spectroscopy at 77 K and gas chromatographic analysis after melting. The proton-donor site is related very strictly to the structure of the semi-occupied molecular orbital of the parent radical cation, with proton transfer taking place from those C-H bonds that carry appreciable unpaired-electron and positive-charge density. Proton acceptance is restricted to C-H bonds at secondary carbon atoms (no proton transfer to C-C bonds nor to C-H bonds at primary carbon atoms), with a preference for the penultimate position and equal (but considerably lower) transfer to the interior sites. In mixed n-alkane crystals, additional selectivity with respect to the site of proton acceptance results from structural factors in combination with the donor site selectivity (structurally determined acceptor site selectivity). PMID- 12118992 TI - Quasi-nature catalysis: developing C-C bond formations catalyzed by late transition metals in air and water. AB - This Account outlines the recent efforts of developing catalysis under the ambient conditions of air and water for synthetic purposes from the author's laboratory. The discussions are focused on catalytic reactions (mostly C-C bond formations) other than oxidations via late transition metals. It includes the following aspects: (1) copper-catalyzed C-C bond formations; (2) palladium catalyzed C-C bond formations; (3) rhodium-catalyzed C-C bond formations; (4) ruthenium-catalyzed olefin isomerizations and C-H activations. The mechanism, limitations, and synthetic applications of these reactions are also discussed. PMID- 12118993 TI - Polymeric materials containing bile acids. AB - Bile acids are biological compounds in the body that have interesting properties and have been used to make special chemical structures in molecular recognition. Various polymers have been synthesized from bile acids. The materials should preserve some of the properties of bile acids, such as biocompatibility, high stability of the steroid nucleus, reactivity of the side groups, optical activity and self-assembling capacity. The synthesis and applications proposed for such polymers are discussed. PMID- 12118994 TI - Adhesion, friction, and mechanical properties of functionalized alkanethiol self assembled monolayers. AB - We have used interfacial force microscopy to study the adhesion, friction, and mechanical properties of molecular monolayers self-assembled on Au surfaces. This quantitative and stable scanning-probe technique permits detailed studies of these factors. By systematic variation of the chemical nature of the end groups on the monolayers and utilization of standard and intuitive contact-mechanics models, quantitative results are presented of inter- and intrafilm bonding strength as well as the relationship between mechanical behavior and the lateral friction force. PMID- 12118995 TI - Molecular design of transition metal alkynyl complexes as building blocks for luminescent metal-based materials: structural and photophysical aspects. AB - In this Account, the design and successful isolation of a series of soluble di- and polynuclear alkynyl complexes of selected metals with d(8) and d(10) electronic configurations are described. These organometallic complexes are found to exhibit rich photophysical and photochemical properties that are unique to the presence of the alkynyl ligand. By a systematic variation of the nature of the ligands, the metal centers, and certain structural features, the photophysical properties could be readily varied and their spectroscopic origins elucidated. Some of these complexes have been shown to be ideal building blocks for the design of luminescent organometallic oligomers and metal-based functional materials. PMID- 12118996 TI - Hydrogen bridges in crystal engineering: interactions without borders. AB - A hydrogen bond, X-H...A, is an interaction wherein a hydrogen atom is attracted to two atoms, X and A, rather than just one and so acts like a bridge between them. This attraction always increases with increasing electronegativity of X and A, and in the classical view all hydrogen bonds are highly electrostatic and sometimes even partly covalent. Gradually, the concept of a hydrogen bond became more relaxed to include weaker interactions, provided some electrostatic character remains. In the limit, these weak hydrogen bonds have considerable dispersive-repulsive character and merge into van der Waals interactions. A great variety of hydrogen bonds are observed in the solid state and the aim of this article is to highlight some features common to all these bonds and further to suggest that the term hydrogen bridge is perhaps a better descriptor for them. Such a description recognizes an interaction without borders and one that admits of much variation in its relative covalent, electrostatic, and van der Waals content. PMID- 12118997 TI - Tripodal amido complexes: molecular "claws" in main group and transition metal chemistry. AB - The chemistry of transition metal amides has received new impetus in recent years due to the systematic exploitation of the amido function [NR(2)](-) in ligand design. A class of 3-fold symmetrical tripodal amido ligands has proved to be a valuable tool in the stabilization of early transition metal and main group metal complexes and complex fragments. Moreover, promising strategies for their use as chemical reagents and homogeneous catalysts have been developed. An overwiew of the current state of this field is given and the potential for further development will be highlighted. PMID- 12118998 TI - Mapping the reaction of peroxynitrite with CO2: energetics, reactive species, and biological implications. PMID- 12118999 TI - Oxidized phospholipids derived from ozone-treated lung surfactant extract reduce macrophage and epithelial cell viability. AB - Ozone is known to be a highly toxic gas present in the urban air which exerts its effect on pulmonary tissue through its facile chemical reactions with target molecules in the airway. One of the first barriers encountered by ozone is epithelial lining fluid which contains pulmonary surfactant rich in glycerophosphocholine lipids. The reaction of ozone with calf lung surfactant extract was found to result in the production of 1-palmitoyl-2-(9'-oxo-nonanoyl) glycerophosphocholine (16:0a/9-al-GPCho) as an expected product of the ozonolysis of abundant unsaturated phospholipids containing unsaturated fatty acyl groups with a double bond at carbon-9. This oxidized phospholipid was identified as a biologically active product in that it reduced elicited macrophage viability by necrosis with an ED(50) of 6 microM. Further studies of the biological activity of 16:0a/9-al-GPCho revealed that concentrations from 100 to 200 nM initiated apoptosis in pulmonary epithelial-like A549 cells as assessed by TUNEL staining, nuclear size, and caspase-3 activation with loss of viability indicated by reduction of mitochondrial dehydrogenase activity. The release of IL-8, a neutrophil chemokine, from A549 cells was also stimulated by 50-100 nM 16:0a/9-al GPCho. Exposure of calf lung surfactant to low levels of ozone (62.5, 125, and 250 ppb) for various time periods from 2 to 48 h in a feedback-regulated ozone exposure chamber resulted in a dose- and time-dependent increase in the formation of 16:0a/9-al-GPCho as measured by a specific and sensitive LC/MS/MS assay. The quantity of this biologically active chain-shortened glycerophosphocholine lipid generated even at 125 ppb ozone for 2-4 h (50-100 nM) was consistent with this product mediating the toxic effects of ozone on cells in contact with surfactant. PMID- 12119000 TI - Cytochrome P450 3A4-mediated bioactivation of raloxifene: irreversible enzyme inhibition and thiol adduct formation. AB - Raloxifene is a selective estrogen receptor modulator which is effective in the treatment of osteoporosis in postmenopausal women. We report herein that cytochrome P450 (P450)3A4 is inhibited by raloxifene in human liver microsomal incubations. The nature of the inhibition was irreversible and was NADPH- and preincubation time-dependent, with K(I) and k(inact) values estimated at 9.9 microM and 0.16 min(-1), respectively. The observed loss of P450 3A4 activity was attenuated partially by glutathione (GSH), implying the involvement of a reactive metabolite(s) in the inactivation process. Subsequently, GSH adducts of raloxifene were identified in incubations with human liver microsomes; substitution with GSH occurred at the 5- or 7-position of the benzothiophene moiety or at the 3'-position of the phenol ring, with the 7-glutathionyl derivative being most abundant based on LC/MS and NMR analyses. These adducts are postulated to derive from addition of GSH to raloxifene arene oxides followed by dehydration and aromatization. Alternatively, raloxifene may be oxidized to an extended quinone intermediate, which then is trapped by GSH conjugation. The bioactivation of raloxifene most likely is catalyzed by P450 3A4, since the formation of GSH adducts was almost abolished when liver microsomes were pretreated with ketoconazole or with an inhibitory anti-P450 3A4 IgG. The GSH adducts also were detected in incubations of raloxifene with rat or human hepatocytes, while the corresponding N-acetylcysteine adducts were identified in the bile and urine from rats treated orally with the drug at 5 mg/kg. Taken together, these data indicate that P450 3A4-mediated bioactivation of raloxifene in vitro is accompanied by loss of enzyme activity. The significance of these findings with respect to the clinical use of raloxifene remains to be determined. PMID- 12119001 TI - A metabolic activation mechanism of 7H-dibenzo[c,g]carbazole via o-quinone. Part 2: covalent adducts of 7H-dibenzo[c,g]carbazole-3,4-dione with nucleic acid bases and nucleosides. AB - 7H-dibenzo[c,g]carbazole (DBC) is a potent multispecies, multisite carcinogen present in the environment. The metabolic activation pathways of DBC are not completely known. It is hypothesized that DBC may be metabolically activated by oxidation to the reactive Michael acceptor o-quinones, which can form stable and depurinating DNA adducts. The synthesis of DBC-3,4-dione has been previously reported by this research group. In the present article, we describe the synthesis and chemical structural elucidation of nine DBC-nucleic acid adducts produced from reactions of DBC-3,4-dione with Ade, Cyt, 2'-deoxyguanosine (dGuo), 2'-deoxycytidine (dCyd), and Guo. Adducts were isolated from reaction mixtures by HPLC and analyzed using MS including elemental compositions and collision activated dissociation (CAD), (1)H NMR, and two-dimensional chemical shift correlation spectroscopy (COSY) NMR. The adducts, 7-[3,4-dione-DBC-1-yl]-Ade, N4 [3,4-dione-DBC-1-yl]-Cyt, 5-[3,4-dione-DBC-1-yl]-Cyt, two conformational isomers of N2-[3,4-dihydroxy-DBC-1-yl]-dGuo, and two conformational isomers of N2-[3,4 dihydroxy-DBC-1-yl]-Guo, were characterized. Two adducts from reactions of DBC 3,4-dione with dCyd were identified by MS but not fully characterized by NMR due to instability of the adducts. Under similar conditions, the reactions of DBC-3,4 dione with Gua and 2'-deoxyadenosine (dAdo) did not result in an identifiable adduct. Liver DNA adducts from mice treated topically with DBC-3,4-dione (100 microg) in dimethyl sulfoxide/acetone (15/85, 100 microL) were identified with 32P-postlabeling. The major adduct chromatographically matched one of the adducts formed from livers of DBC-treated mouse (adduct 3) using identical conditions. PMID- 12119003 TI - Dinitrobenzene-mediated production of peroxynitrite by neuronal nitric oxide synthase. AB - Neuronal nitric oxide synthase (nNOS) is a modular enzyme that consists of a flavin-containing reductase domain and a heme-containing oxygenase domain, fused by a calmodulin (CaM)-binding sequence. Within the central nervous system, nNOS is localized in the cerebellum. CaM binding to nNOS activates both intradomain as well as interdomain electron transfer, and thus activity. The nNOS reductase shares many characteristics with NADPH-cytochrome P450 reductase (CPR), such as catalyzing the reduction of exogenous electron acceptors such as quinones and nitroarenes. The nitroarene 1,3-dinitrobenzene (1,3-DNB) is a cerebellar neurotoxicant in rats. 1,3-DNB is metabolized by CPR in liver, and it was proposed that metabolism of 1,3-DNB to reactive intermediates is involved in mediating the cerebellar neurotoxicity. We have found that, in a manner similar to CPR, nNOS can interact with 1,3-DNB and generate superoxide anion radical (O2* ). Electron transfer through the nNOS reductase is not limiting for nitric oxide (NO.) and L-citrulline production, even in the presence of certain exogenous electron acceptors such as 1,3-DNB. Therefore, NO., L-citrulline, and O2*- are simultaneously produced by nNOS in the presence of 1,3-DNB and other nitroarenes. The simultaneous production of NO. and O2*- leads to peroxynitrite (ONOO-) formation via the combination of these two radicals at a near-diffusion controlled reaction rate. We present convincing data supporting the hypothesis that in the presence of 1,3-DNB, nNOS is converted from a purely NO. and L citrulline synthase to a ONOO- and L-citrulline synthase, and propose that the resulting nitosative stress plays a role in the cerebellar neurotoxicity of 1,3 DNB. This paper introduces a new and novel enzymatic mechanism with direct toxicological implications whereby nNOS is converted into a ONOO- synthase by certain nitroarenes. PMID- 12119002 TI - Quantification of 2'-fluoro-2'-deoxyuridine and 2'-fluoro-2'-deoxycytidine in DNA and RNA isolated from rats and woodchucks using LC/MS/MS. AB - Apatmers are synthesized using 2'-fluoropyrimdines in place of normal pyrmidines to stabilize them against enzymatic degradation, and thereby improve their therapeutic efficacy. Despite this stabilizing effect, the apatmers can still be degraded by nucleases in the blood. Primer template extension studies have demonstrated that mammalian DNA polymerases can incorporate these 2' fluoropyrimidines into growing strands of DNA. The toxicologic effects of these compounds have been examined in rats and woodchucks, animals known to be susceptible to the toxic effects of other modified pyrimidines. Whether these nucleosides can be incorporated into DNA in vivo has not been established. These studies report the development of methodologies and the results of studies designed to determine if and to what extent 2'-fluoropyrimidines are incorporated into tissue DNA following long-term treatment. Rats were dosed intravenously with either 2'-fluorouridine (2'-FU) or 2'-fluorocytidine (2'-FC) at doses of 5, 50, and 500 mg/kg/day for 90 days. Woodchucks were dosed intravenously with either 2' FU or 2'-FC at doses of 0.75 or 7.5 mg/kg/day for 90 days. The amounts of 2'-FU or 2'-FC in DNA and RNA were quantified using newly developed LC/MS/MS methodologies. Administration of 2'-FU to rats and woodchucks resulted in incorporation of the compound into DNA from liver, spleen, testis, muscle, and kidney. Incorporation also occurred in RNA from rat liver (only tissue examined). Similarly, administration of 2'-FC to rats and woodchucks resulted in incorporation into liver DNA (only tissue examined). These data demonstrate that 2'-fluoropyrimidines are incorporated into DNA and RNA of various tissues of rats and woodchucks following long-term administration. PMID- 12119004 TI - Inhibition of cellular enzymes by equine catechol estrogens in human breast cancer cells: specificity for glutathione S-transferase P1-1. AB - Glutathione S-transferases (GSTs) are a family of detoxification isozymes that protect cells by conjugating GSH to a variety of toxic compounds, and they may also play a role in the regulation of both cellular proliferation and apoptosis. We have previously shown that human GST P1-1, which is the most widely distributed extrahepatic isozyme, could be inactivated by the catechol estrogen metabolite 4-hydroxyequilenin (4-OHEN) in vitro [Chang, M., Shin, Y. G., van Breemen, R. B., Blond, S. Y., and Bolton, J. L. (2001) Biochemistry 40, 4811 4820]. In the present study, we found that 4-OHEN and another catechol estrogen, 4,17beta-hydroxyequilenin (4,17beta-OHEN), significantly decreased GSH levels and the activity of GST within minutes in both estrogen receptor (ER) negative (MDA MB-231) and ER positive (S30) human breast cancer cells. In addition, 4-OHEN caused significant decreases in GST activity in nontransformed human breast epithelial cells (MCF-10A) but not in the human hepatoma HepG2 cells, which lack GST P1-1. We also showed that GSH partially protected the inactivation of GST P1 1 by 4-OHEN in vitro, and depletion of cellular GSH enhanced the 4-OHEN-induced inhibition of GST activity. In addition, 4-OHEN GSH conjugates contributed about 27% of the inactivation of GST P1-1 by 4-OEHN in vitro. Our in vitro kinetic inhibition experiments with 4-OHEN showed that GST P1-1 had a lower K(i) value (20.8 microM) compared to glyceraldehyde-3-phosphate dehydrogenase (GAPDH, 52.4 microM), P450 reductase (PR, 77.4 microM), pyruvate kinase (PK, 159 microM), glutathione reductase (GR, 230 microM), superoxide dismutase (SOD, 448 microM), catalase (562 microM), GST M1-1 (620 microM), thioredoxin reductase (TR, 694 microM), and glutathione peroxidase (GPX, 1410 microM). In contrast to the significant inhibition of total GST activity in these human breast cancer cells, 4-OHEN only slightly inhibited the cellular GAPDH activity, and other cellular enzymes including PR, PK, GR, SOD, catalase, TR, and GPX were resistant to 4-OHEN induced inhibition. These data suggest that GST P1-1 may be a preferred protein target for equine catechol estrogens in vivo. PMID- 12119005 TI - Genotoxicity of trivalent chromium in bacterial cells. Possible effects on DNA topology. AB - Trivalent chromium is a metal required for proper sugar and fat metabolism. However, it has been suggested that it causes DNA damage in in vitro test systems, although in vivo toxicity has not yet been proved. In the present study, the effect of Cr3+ on bacterial cells was tested with the Pro-Tox (C) assay, and its cellular uptake was measured with flame atomic absorption spectroscopy. The potential genotoxicity of Cr3+ was further examined by the study of its influence on a bacterial type II topoisomerase. Cr3+ was shown to cause DNA damage and inhibit topoisomerase DNA relaxation activity, probably by preventing the formation of the covalent link between enzyme and double helix. In addition, Cr3+ decreases the viability and/or proliferation rate of eukaryotic cells such as murine B16 melanoma cells and human MCF-10A neoT ras-transformed human epithelial cells. The possible implication for Cr3+ intake by humans is discussed. PMID- 12119006 TI - MAPKs mediate S phase arrest induced by vanadate through a p53-dependent pathway in mouse epidermal C141 cells. AB - Mitogen-activated protein (MAP) kinases play an important role in mediation of the signal transduction pathway in cellular response to genotoxic stress. Cell growth arrest is considered as an early stage in response to the genotoxic stress. p53 is well-known as a tumor suppression gene involved in both cell growth arrest and apoptosis. The present study investigated the involvement of MAP kinases in vanadate-induced cell growth arrest and the relationship of p53. DNA content analysis showed that vanadate-induced S phase arrest is time- and dose-dependent in p53 wild-type C141 cells but not in p53-deficient C141 cells. Western blotting results indicated that vanadate caused an inactivation of p-cdk2 at Thr160, which is an important kinase for the progression of S phase, and an increase in expression of p21, which is a key for S phase arrest. In p53 deficient cells, vanadate did not induce any observable change in p21 or p-cdk2 level. In addition, vanadate up-regulated phospho-p38 and ERK, two members of MAP kinases. At the same time, vanadate increased the p53 activity as measured by luciferase assay. Addition of PD98059 and SB202190, inhibitors of ERK and p38, respectively, decreased vanadate-induced S phase arrest, reduced p21 levels, restored activation of p-cdk2, and decreased p53 activity. The study demonstrated that vanadate-induced S phase arrest is mediated by both ERK and p38 in a p53 dependent pathway. PMID- 12119007 TI - Kinetics of the biotransformation of maleylacetone and chlorofluoroacetic acid by polymorphic variants of human glutathione transferase zeta (hGSTZ1-1). AB - Glutathione transferase zeta (GSTZ1-1) catalyzes the cis-trans isomerization of maleylacetoacetate and the biotransformation of a range of alpha-haloacids. The objective of this study was to determine the kinetics of the biotransformation of maleylacetone (MA), an analogue of the natural substrate maleylacetoacetate, and chlorofluoroacetic acid (CFA) by polymorphic variants of recombinant hGSTZ1-1. The k(cat) of the four variants of hGSTZ1-1 with MA as the substrate followed the order: 1c-1c > 1b-1b > 1d-1d > 1a-1a whereas the k(cat) for the biotransformation of CFA followed the order: 1a-1a > 1b-1b approximately 1c-1c approximately 1d-1d. The turnover rates of MA were much higher than those of CFA for each variant and ranged from 22-fold (1a-1a) to 980-fold differences (1c-1c). The catalytic efficiencies of hGSTZ1-1 variants with MA as the substrate were much greater than those with CFA as the substrate, but little difference among the polymorphic variants was observed. MA was a mixed inhibitor of all variants with CFA as substrate: the mean competitive inhibition constant (K(ic)(MA)) for all variants was about 100 microM, and the mean uncompetitive inhibition constant (K(iu)(MA)) was about 201 microM. Hence, MA and alpha-haloacids apparently compete for the same active site on the enzyme. DCA-induced inactivation of the four variants showed that the inactivated enzymes show markedly reduced isomerase activities. The residual activities were different for each variant: 1a-1a (12%) > 1b-1b approximately 1c-1c approximately 1d-1d (<5%). This is the first kinetic analysis of polymorphic variants of hGSTZ1-1, and the similarity of the kinetic constants for hGSTZ1-1 variants with either MA or CFA as substrates indicates that few differences in DCA-induced perturbations of tyrosine metabolism would likely be observed in humans. PMID- 12119008 TI - Synthesis, in vitro metabolism, cell transformation, mutagenicity, and DNA adduction of dibenzo[c,mno]chrysene. AB - Polycyclic aromatic hydrocarbons (PAHs) are ubiquitous environmental pollutants. Due to its structural similarity with the potent carcinogen dibenzo[a,l]pyrene (DB[a,l]P) and because of its environmental presence, dibenzo[c,mno]chrysene (naphtho[1,2-a]pyrene, N[1,2-a]P) is of considerable research interest. We therefore developed an efficient synthesis of N[1,2-a]P, and examined its in vitro metabolism by male Sprague Dawley rat liver S9 fraction. Its mutagenic activity in S. typhimurium TA 100 and its morphological cell transforming ability in mouse embryo fibroblasts were evaluated. On the basis of spectral analyses, the in vitro major metabolites were identified as the fjord region dihydrodiol trans-9,10-dihydroxy-9,10-dihydro-N[1,2-a]P (N[1,2-a]P-9,10-dihydrodiol), the K region diols N[1,2-a]P-4,5-dihydrodiol and N[1,2-a]P-7,8-dihydrodiol, and also the 1-, 3-, and 10-hydroxy-N[1,2-a]P; the structure of N[1,2-a]P-9,10-dihydrodiol was also confirmed by independent synthesis. In assays with S. typhimurium TA 100, N[1,2-a]P-9,10-dihydrodiol was half as mutagenic as (+/-)-trans-7,8 dihydroxy-7,8-dihydrobenzo[a]pyrene (B[a]P-7,8-dihydrodiol) at > or =4 nmol/plate. N[1,2-a]P-9,10-dihydrodiol was much more mutagenic than N[1,2-a]P at all dose levels, suggesting that the N[1,2-a]P-9,10-dihydrodiol is the likely proximate mutagen of N[1,2-a]P. Evaluation of morphological cell transformation in C3H10T1/2C18 mouse embryo fibroblasts revealed that N[1,2-a]P was comparable to B[a]P. We further examined the pattern of in vitro adduct formation between calf thymus DNA and (+/-)-anti-9,10-dihydroxy-9,10-dihydro-11,12-epoxy-9,10,11,12 tetrahydro-N[1,2-a]P (N[1,2-a]PDE) and found that dG-adduct formation is 2.9-fold greater than dA-adduct formation. On the basis of our results and those reported in the literature, our working hypothesis is that N[1,2-a]P may be added to the list of potent carcinogens that includes DB[a,l]P. This hypothesis is currently being tested in our laboratory. PMID- 12119009 TI - Comparative tumorigenicity of the environmental pollutant 6-nitrochrysene and its metabolites in the rat mammary gland. AB - Human exposure to the class of nitropolynuclear aromatic hydrocarbons is via inhalation and/or ingestion. Therefore, one of the goals of this study was to determine the propensity of the environmental contaminant 6-nitrochrysene (6-NC) for inducing mammary cancer following its oral administration to female CD rats. 2-Amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), an established mammary carcinogen in the same animal model, was used as a positive control and trioctanoin as a negative control. Thirty-day-old female CD rats were gavaged once weekly for 8 weeks with 6-NC at 50, 25, or 12.5 micromol/rat or PhIP at 50 micromol/rat in 500 microL of trioctanoin. Twenty-three weeks after the last carcinogen administration, rats were decapitated, necropsied, and evaluated histologically. The most common mammary tumors were adenocarcinomas, followed by adenomas and fibroadenomas. The incidence and multiplicity (mean +/- standard deviation) of mammary adenocarcinomas induced by these two carcinogens at the highest dose (6-NC: 90%, 3.73 +/- 2.74; PhIP: 83%, 2.62 +/- 2.58) were significantly higher than those in control rats (10%, 0.10 +/- 0.31). However, there were no statistically significant differences between groups treated with 6 NC and PhIP or among groups receiving various doses of 6-NC. Following its metabolic activation, 6-NC is known to bind covalently to DNA; however, it remains to be determined whether it can also induce DNA base oxidation. Thus, employing the same route of administration, our studies revealed no effect of 6 NC on the basal level of 8-hydroxy-2'-deoxyguanosine (8-OHdG) in the mammary gland in tests at 6, 24, and 48 h after 6-NC treatment and at termination of the carcinogenesis assay in the normal, noninvolved tissue and in mammary tumors. This result suggests that covalent DNA binding of 6-NC metabolites is important in the induction of mammary cancer in rats. Therefore, the other goal of this study was to compare the tumorigenic activities of 6-NC and its metabolites in the rat mammary gland by intramammary administration. This route has also been used in our laboratory to induce mammary cancer in the rat by 6-NC and is employed here to avoid systemic effects and to determine the role of the mammary gland in the metabolic activation of 6-NC and its metabolites. Toward this end, a new method was developed to obtain ample materials of trans-1,2-dihydroxy-1,2 dihydro-6-aminochrysene (1,2-DHD-6-AC); other metabolites were synthesized as reported previously. On the basis of the results, the carcinogenic potency toward the mammary gland is ranked in the following order: 6-NC > 1,2-DHD-6-NC > 6-AC > 6-NCDE > 1,2-DHD-6-AC. Among the metabolites tested, 1,2-DHD-6-NC was the most potent carcinogen. It was significantly more active than its reduced product 1,2 DHD-6-AC. However, the potency of 1,2-DHD-6-NC was not significantly different from 6-AC, a metabolite derived from simple nitroreduction, or from 6-NCDE. Collectively, these results suggest that metabolites derived from both ring oxidation and nitroreduction contribute to the overall carcinogenicity of 6-NC in the rat mammary gland. PMID- 12119010 TI - The detection and identification of 42,43,44,45,46,47,55-heptanor-41 oxoyessotoxin, a new marine toxin from adriatic shellfish, by liquid chromatography-mass spectrometry. AB - The diarrhetic shellfish toxin composition in the digestive glands of mussels collected in June 2001 from the Northern Adriatic sea was investigated by high performance liquid chromatography coupled with electrospray ion trap mass spectrometry. Along with known yessotoxins (1, 3-6), identified by comparison of their retention times and mass spectra with those of appropriate standards, a new marine toxin, 42,43,44,45,46,47,55-heptanor-41-oxoyessotoxin, 7, was detected. MS/MS experiments were used to gain structural information. 7 represents a new addition to the class of yessotoxins. PMID- 12119011 TI - Inhibition of lipid peroxidation in synaptosomes and liposomes by nitrates and nitrites. AB - NO is produced endogenously from arginine by the action of NO synthase, and exogenously by nitrovasodilators, including organic nitrates and nitrites. NO has been proposed as a cytotoxic and cytoprotective agent. There is strong evidence that NO acts as an apparent antioxidant in inhibiting lipid peroxidation, via chain termination, and interestingly lipid nitrates and nitrites have been proposed to be products of this chain termination. Both pro- and antioxidant mechanisms may be drawn for nitrates and nitrites; therefore, their effects on lipid peroxidation were measured in two systems, using tocopherol, thiol, and an NO donor for comparison: (1) rat cerebrocortical synaptosomes with Fe(II)-induced lipid peroxidation measured by thiobarbituric acid reactive substances (TBARS), and (2) phospholipid liposomes with an azo-initiator induction system, quantified by a fluorescent probe of peroxide formation. In contrast to the classical nitrate nitroglycerin, novel nitrates which release NO on reaction with thiols and two novel nitrates which spontaneously generate NO in aqueous solution inhibited lipid peroxidation. i-Amyl nitrite inhibited lipid peroxidation, and its properties were further studied with ESR spectroscopy. The data show that classical nitrites and novel nitrates are not prooxidants, but inhibit lipid peroxidation. PMID- 12119014 TI - Crystal structure of the complex between actin and human vitamin D-binding protein at 2.5 A resolution. AB - A high-affinity complex formed between G-actin and plasma vitamin D-binding protein (DBP) is believed to form part of a scavenging system in the plasma for removing actin released from damaged cells. In the study presented here, we describe the crystal structure of the complex between actin and human vitamin D binding protein at 2.5 A resolution. The complex contains one molecule of each protein bound together by extensive ionic, polar, and hydrophobic interactions. It includes an ATP and a calcium ion bound to actin, but no evidence of vitamin D metabolites bound to the DBP. Both actin and DBP are multidomain molecules, two major domains in actin and three in DBP. All of these domains contribute to the interaction between the molecules. DBP enfolds the end of the actin molecule, principally in actin subdomain 3 but with additional interactions in actin subdomain 1. This orientation is similar to the binding of profilin to actin, as predicted from previous studies. The more extensive interactions of DBP give an affinity for actin some 3 orders of magnitude higher than that for profilin. The larger "footprint" of DBP on actin also leads to an overlap with the actin binding site of gelsolin domain I. PMID- 12119015 TI - A critical residue in the folding pathway of an integral membrane protein. AB - Although a number of common diseases are a direct consequence of membrane protein misfolding, studies of membrane protein folding and misfolding lag well behind those of soluble proteins. Here it is shown that an interfacial residue, Tyr16, of the integral membrane protein diacylglycerol kinase (DAGK) plays a critical role in the folding pathway of this protein. Properly folded Y16C exhibits kinetic parameters and stability similar to wild-type DAGK. However, when unfolded and then allowed to spontaneously fold in the presence of model membranes, Y16C exhibits dramatically lower rates and efficiencies of functional assembly compared to the wild-type protein. The Y16C mutant represents a class of mutations which may be commonly found in disease-related membrane proteins. PMID- 12119013 TI - Hint, Fhit, and GalT: function, structure, evolution, and mechanism of three branches of the histidine triad superfamily of nucleotide hydrolases and transferases. AB - HIT (histidine triad) proteins, named for a motif related to the sequence HphiHphiHphiphi (phi, a hydrophobic amino acid), are a superfamily of nucleotide hydrolases and transferases, which act on the alpha-phosphate of ribonucleotides, and contain a approximately 30 kDa domain that is typically either a homodimer of approximately 15 kDa polypeptides with two active-sites or an internally, imperfectly repeated polypeptide that retains a single HIT active site. On the basis of sequence, substrate specificity, structure, evolution, and mechanism, HIT proteins can be classified into the Hint branch, which consists of adenosine 5'-monophosphoramide hydrolases, the Fhit branch, which consists of diadenosine polyphosphate hydrolases, and the GalT branch, which consists of specific nucleoside monophosphate transferases, including galactose-1-phosphate uridylyltransferase, diadenosine tetraphosphate phosphorylase, and adenylyl sulfate:phosphate adenylytransferase. At least one human representative of each branch is lost in human diseases. Aprataxin, a Hint branch hydrolase, is mutated in ataxia-oculomotor apraxia syndrome. Fhit is lost early in the development of many epithelially derived tumors. GalT is deficient in galactosemia. Additionally, ASW is an avian Hint family member that has evolved to have unusual gene expression properties and the complete loss of its nucleotide binding site. The potential roles of ASW and Hint in avian sexual development are discussed elsewhere. Here we review what is known about biological activities of HIT proteins, the structural and biochemical bases for their functions, and propose a new enzyme mechanism for Hint and Fhit that may account for the differences between HIT hydrolases and transferases. PMID- 12119016 TI - Synthesis and antiviral evaluation of a mutagenic and non-hydrogen bonding ribonucleoside analogue: 1-beta-D-Ribofuranosyl-3-nitropyrrole. AB - Synthetic small molecules that promote viral mutagenesis represent a promising new class of antiviral therapeutics. Ribavirin is a broad-spectrum antiviral nucleoside whose antiviral mechanism against RNA viruses likely reflects the ability of this compound to introduce mutations into the viral genome. The mutagenicity of ribavirin results from the incorporation of ribavirin triphosphate opposite both cytidine and uridine in viral RNA. In an effort to identify compounds with mutagenicity greater than that of ribavirin, we synthesized 1-beta-D-ribofuranosyl-3-nitropyrrole (3-NPN) and the corresponding triphosphate (3-NPNTP). These compounds constitute RNA analogues of the known DNA nucleoside 1-(2'-deoxy-beta-D-ribofuranosyl)-3-nitropyrrole. The 3-nitropyrrole pseudobase has been shown to maintain the integrity of DNA duplexes when placed opposite any of the four nucleobases without requiring hydrogen bonding. X-ray crystallography revealed that 3-NPN is structurally similar to ribavirin, and both compounds are substrates for adenosine kinase, an enzyme critical for conversion to the corresponding triphosphate in cells. Whereas ribavirin exhibits antiviral activity against poliovirus in cell culture, 3-NPN lacks this activity. Evaluation of 3-NPNTP utilization by poliovirus RNA-dependent RNA polymerase (RdRP) revealed that 3-NPNTP was not accepted universally. Rather, incorporation was only observed opposite A and U in the template and at a rate 100-fold slower than the rate of incorporation of ribavirin triphosphate. This diminished rate of incorporation into viral RNA likely precludes 3-NPN from functioning as an antiviral agent. These results indicate that hydrogen bonding substituents are critical for efficient incorporation of ribonucleotides into RNA by viral RdRPs, thus providing important considerations for the design of improved mutagenic antiviral nucleosides. PMID- 12119017 TI - Thermal and conformational stability of seed coat soybean peroxidase. AB - Soybean peroxidase (SBP) obtained from the soybean seed coats belongs to class III of the plant peroxidase superfamily. Detailed circular dichroism and steady state fluorescence studies have been carried out to monitor thermal as well as denaturant-induced unfolding of SBP and apo-SBP. Melting of secondary and tertiary structures of SBP occurs with characteristic transition midpoints, T(m), of 86 and 83.5 degrees C, respectively, at neutral pH. Removal of heme resulted in greatly decreased thermal stability of the protein (T(m) = 38 degrees C). The deltaG degrees (H2O) determined from guanidine hydrochloride-induced denaturation at 25 degrees C and at neutral pH is 43.3 kJ mol(-1) for SBP and 9.0 kJ mol(-1) for apo-SBP. Comparison with the reported unfolding data of the homologous enzyme, horseradish peroxidase (HRP-C), showed that SBP exhibits significantly high thermal and conformational stability. We show that this enhanced structural stability of SBP relative to HRP-C arises due to the unique nature of their heme binding. A stronger heme-apoprotein affinity probably due to the interaction between Met37 and the C8 heme vinyl substituent contributes to the unusually high structural stability of SBP. PMID- 12119018 TI - The structure of PTP-1B in complex with a peptide inhibitor reveals an alternative binding mode for bisphosphonates. AB - Inhibitors of PTP-1B could be therapeutically beneficial in the treatment of type 2 diabetes. Owing to the large number of phosphatases in the cell, inhibitors against PTP-1B must not only be potent but selective as well. N-Benzoyl-L glutamyl-[4-phosphono(difluoromethyl)]-L-phenylalanine-[4-phosphono(difluoro methyl)]-L-phenylalanineamide (BzN-EJJ-amide) is a low nanomolar inhibitor of PTP 1B that shows selectivity over several protein tyrosine phosphatases. To gain an insight into the basis of its potency and selectivity, we evaluated several analogues of the inhibitor and introduced amino acid substitutions into PTP-1B by site-directed mutagenesis. We also determined the crystal structure of PTP-1B in complex with BzN-EJJ-amide at 2.5 A resolution. Our results indicate that the high inhibitory potency is due to interactions of several of its chemical groups with specific protein residues. An interaction between BzN-EJJ-amide and Asp48 is of particular significance, as substitution of Asp48 to alanine resulted in a 100 fold loss in potency. The crystal structure also revealed an unexpected binding orientation for a bisphosphonate inhibitor on PTP-1B, where the second difluorophosphonomethyl phenylalanine (F(2)PMP) moiety is bound close to Arg47 rather than in the previously identified second aryl phosphate site demarked by Arg24 and Arg254. Our results suggest that potent and selective PTP-1B inhibitors may be designed by targeting the region containing Arg47 and Asp48. PMID- 12119019 TI - Cytoplasmic retraction of the amino terminus of human multidrug resistance protein 1. AB - Human multidrug resistance protein 1 (MRP1) is a member of the ATP-binding cassette (ABC) transport superfamily which also includes human multidrug resistance 1 (MDR1) gene product P-glycoprotein (Pgp). Overexpression of MRP1 or Pgp causes multidrug resistance in cancer cells. Different from Pgp, MRP1 contains an extra membrane-spanning domain (MSD1) with a putative extracellular amino terminus in addition to the core structure of two MSDs and two NBDs (nucleotide-binding domains). The structural and functional significance of the additional MSD1 in MRP1 remains elusive. In this study, we generated an IgG1 subclass monoclonal antibody, IU2H10, specific to the amino terminus of human MRP1 and mapped its epitope to 10 amino acids (S8ADGSDPLWD17). It can be used for Western blot, immunoprecipitation, and indirect immunofluorescence studies of human MRP1. However, surprisingly we found that IU2H10 cannot react with MRP1 unless cells are permeabilized. Furthermore, the IU2H10 epitope is exposed extracellularly when the carboxyl-terminal core domain of human MRP1 is deleted. Examination of the amino-terminal sequence of human MRP1 suggests that it consist of mainly coiled structures. These observations provide evidence for a model that is different from the prevailing extracellular location of the amino terminus of human MRP1. It is possible that part of the amino terminus of human MRP1, following exposure to the lumen of the endoplasmic reticulum, is retracted to the cytoplasm. PMID- 12119020 TI - Distinct activation states of alpha5beta1 integrin show differential binding to RGD and synergy domains of fibronectin. AB - alpha5beta1 integrin can occupy several distinct conformational states which support different strengths of binding to fibronectin [Garcia, A. J., et al. (1998) J. Biol. Chem. 273, 34710-34715]. Using a model system in which specific activating monoclonal antibodies were used to achieve uniform activated states, the binding of alpha5beta1 to full-length wild-type fibronectin and mutants of fibronectin in the defined RGD and PHSRN synergy sites was analyzed using a novel method that measures the strength of the coupling between integrin and its ligand. Neither TS2/16- nor AG89-activated alpha5beta1 showed significant mechanical coupling to RGD-deleted fibronectin. However, peptide competition assays demonstrated a 6-fold difference in the binding affinities of these two states for RGD. The mutant synergy site reduced the AG89 (low)-activated state to background levels, but the TS2/16-activated state still retained approximately 30% of the wild-type activity. Thus, these two active binding states of alpha5beta1 interact differently with both the RGD and synergy domains. The failure of the AG89-activated state to show mechanical coupling to either the RGD or synergy domain mutants was unexpected and implies that the RGD domain itself does not contribute significant mechanical strength to the alpha5beta1 fibronectin interaction. The lack of RGD alone to support alpha5beta1 coupling was further confirmed using a synthetic polymer presenting multiple copies of the RGD loop. These results suggest a model in which the RGD domain serves to activate and align the alpha5beta1-fibronectin interface, and the synergy site provides the mechanical strength to the bond. PMID- 12119021 TI - Identification of a peroxide-sensitive redox switch at the CXXC motif in the human mitochondrial branched chain aminotransferase. AB - The human mitochondrial branched chain aminotransferase isoenzyme (hBCATm) must be stored in a reducing environment to remain active. Oxidation or labeling of hBCATm with sulfhydryl reagents results in enzyme inhibition. In this study, we investigated both the structural and biochemical basis for the sensitivity of hBCATm to these reagents. In its native form, hBCATm has two reactive cysteine residues which were identified as Cys315 and Cys318 using iodinated beta-(4 hydroxyphenyl)ethyl maleimide. These are located in the large domain of the homodimer, about 10 A from the active site. The crystal structures show evidence for a thiol-thiolate hydrogen bond between Cys315 and Cys318. Under oxidizing conditions, these cysteine residues can reasonably form a disulfide bond because of the short distance between the sulfur atoms (3.09-3.46 A), requiring only a decrease of 1.1-1.5 A. In addition to Cys315 playing a structural role by anchoring Tyr173, which in the ketimine form increases access to the active site, our evidence indicates that these cysteine residues act as a redox switch in hBCATm. Electrospray ionization mass spectrometry analysis and UV-Vis spectroscopic studies of 5,5'-dithiobis(2-nitrobenzoic acid) labeled hBCATm showed that during labeling, an intrasubunit disulfide bond was formed in a significant portion of the protein. Furthermore, it was established that reaction of hBCATm with H2O2 abolished its activity and resulted in the formation of an intrasubunit disulfide bond between Cys315 and Cys318. Addition of dithiothreitol completely reversed the oxidation and restored activity. Therefore, the results demonstrate that there is redox-linked regulation of hBCATm activity by a peroxide sensitive CXXC center. Future studies will determine if this center has an in vivo role in the regulation of branched chain amino acid metabolism. PMID- 12119022 TI - Structural studies of the L-threonine-O-3-phosphate decarboxylase (CobD) enzyme from Salmonella enterica: the apo, substrate, and product-aldimine complexes. AB - The evolution of biosynthetic pathways is difficult to reconstruct in hindsight; however, the structures of the enzymes that are involved may provide insight into their development. One enzyme in the cobalamin biosynthetic pathway that appears to have evolved from a protein with different function is L-threonine-O-3 phosphate decarboxylase (CobD) from Salmonella enterica, which is structurally similar to histidinol phosphate aminotransferase [Cheong, C. G., Bauer, C. B., Brushaber, K. R., Escalante-Semerena, J. C., and Rayment, I. (2002) Biochemistry 41, 4798-4808]. This enzyme is responsible for synthesizing (R)-1-amino-2 propanol phosphate which is the precursor for the linkage between the nucleotide loop and the corrin ring in cobalamin. To understand the relationship between this decarboxylase and the aspartate aminotransferase family to which it belongs, the structures of CobD in its apo state, the apo state complexed with the substrate, and its product external aldimine complex have been determined at 1.46, 1.8, and 1.8 A resolution, respectively. These structures show that the enzyme steers the breakdown of the external aldimine toward decarboxylation instead of amino transfer by positioning the carboxylate moiety of the substrate out of the plane of the pyridoxal ring and by placing the alpha-hydrogen out of reach of the catalytic base provided by the lysine that forms the internal aldimine. It would appear that CobD evolved from a primordial PLP-dependent aminotransferase, where the selection was based on similarities between the stereochemical properties of the substrates rather than preservation of the fate of the external aldimine. These structures provide a sequence signature for distinguishing between L-threonine-O-3-phosphate decarboxylase and histidinol phosphate aminotransferases, many of which appear to have been misannotated. PMID- 12119023 TI - In vitro selection of hairpin ribozymes activated with short oligonucleotides. AB - We have carried out an in vitro selection to obtain an allosteric hairpin ribozyme, which has cleavage activity in the presence of an exogenous short oligonucleotide as a regulator. Random sequences were inserted in a region corresponding to the hairpin loop of the ribozyme. After 12 rounds of selection, DNA templates were cloned. Of a total of 34 clones, 18 contained the same sequence, and the obtained hairpin ribozymes showed the cleavage activity specifically in the presence of the regulator oligonucleotide. All of the clones contained sequences complementary to the regulator oligonucleotide. The ribozymes with high cleavage activities gained characteristic hairpin loops at the random domain, which were similar to each other. In the absence of the oligonucleotide, the loop domain within the allosteric ribozyme probably forms a slipped hairpin loop, and the complementary sequence, with the regulator oligonucleotide located at the single stranded loop, would allow easy access of the oligonucleotide. The binding of the regulator oligonucleotide triggers a structural change of the hairpin loop to form an active conformation. Furthermore, we constructed an allosteric hammerhead ribozyme by introducing the characteristic hairpin loop. The modified hammerhead ribozyme was also changed to an allosteric ribozyme, which was activated by the addition of the regulator oligonucleotide. The characteristic hairpin loop, which was proved to be regulated by an exogenous oligonucleotide in this report, may be used to control RNA functions in various fields. PMID- 12119026 TI - Vibrational modes of tyrosines in cytochrome c oxidase from Paracoccus denitrificans: FTIR and electrochemical studies on Tyr-D4-labeled and on Tyr280His and Tyr35Phe mutant enzymes. AB - A combined electrochemical and FTIR spectroscopic approach was used to identify the vibrational modes of tyrosines in cytochrome c oxidase from Paracoccus denitrificans which change upon electron transfer and coupled proton transfer. Electrochemically induced FTIR difference spectra of the Tyr-D4-labeled cytochrome c oxidase reveal that only small contributions arise from the tyrosines. Contributions between 1600 and 1560 cm(-1) are attributed to nu8a/8b(CC) ring modes. The nu19(CC) ring mode for the protonated form of tyrosines is proposed to absorb with an uncommonly small signal at 1525-1518 cm( 1) and for the deprotonated form at 1496-1486 cm(-1), accompanied by the increase of the nu19(CC) ring mode of the Tyr-D(4)-labeled oxidase at approximately 1434 cm(-1). A signal at 1270 cm(-1) can be tentatively attributed to the nu7'a(CO) and delta(COH) mode of a protonated tyrosine. Uncommon absorptions, like the mode at 1524 cm(-1), indicate the involvement of Tyr280 in the spectra. Tyr280 is a crucial residue close to the binuclear center and is covalently bonded to His276. The possible changes of the spectral properties are discussed together with the absorbance spectra of tyrosine bound to histidine. The vibrational modes of Tyr280 are further analyzed in combination with the mutation to histidine, which is assumed to abolish the covalent bonding. The electrochemically induced FTIR difference spectra of the Tyr280His mutant point to a change in protonation state in the environment of the binuclear center. Together with an observed decrease of a signal at 1736 cm(-1), previously assigned to Glu278, a possible functional coupling is reflected. In direct comparison to the FTIR difference spectra of the D4-labeled compound and comparing the spectra at pH 7 and 4.8, the protonation state of Tyr280 is discussed. Furthermore, a detailed analysis of the mutant is presented, the FTIR spectra of the CO adduct revealing a partial loss of Cu(B). Electrochemical redox titrations reflect a downshift of the heme a3 midpoint potential by 95 +/- 10 mV. Another tyrosine identified to show redox dependent changes upon electron transfer is Tyr35, a residue in the proposed D-pathway of the cytochrome c oxidase. PMID- 12119025 TI - The microtubule stabilizing agent laulimalide does not bind in the taxoid site, kills cells resistant to paclitaxel and epothilones, and may not require its epoxide moiety for activity. AB - Laulimalide is a cytotoxic natural product that stabilizes microtubules. The compound enhances tubulin assembly, and laulimalide is quantitatively comparable to paclitaxel in its effects on the reaction. Laulimalide is also active in P glycoprotein overexpressing cells, while isolaulimalide, a congener without the drug's epoxide moiety, was reported to have negligible cytotoxic and biochemical activity [Mooberry et al. (1999) Cancer Res. 59, 653-660]. We report here that laulimalide binds at a site on tubulin polymer that is distinct from the taxoid site. We found that laulimalide, while as active as paclitaxel, epothilone A, and eleutherobin in promoting the assembly of cold-stable microtubules, was unable to inhibit the binding of radiolabeled paclitaxel or of 7-O-[N-(2,7-difluoro-4' fluoresceincarbonyl)-L-alanyl]paclitaxel, a fluorescent paclitaxel derivative, to tubulin. Confirming this observation, we demonstrated that microtubules formed in the presence of both laulimalide and paclitaxel contained near-molar quantities, relative to tubulin, of both drugs. Laulimalide was active against cell lines resistant to paclitaxel or epothilones A and B on the basis of mutations in the M40 human beta-tubulin gene. We also report that a laulimalide analogue lacking the epoxide moiety, while less active than laulimalide in biochemical and cellular systems, is probably more active than isolaulimalide. Further exploration of the role of the epoxide in the interaction of laulimalide with tubulin is therefore justified. PMID- 12119024 TI - Preparation and characterization of alpha-D-glucopyranosyl-alpha-acarviosinyl-D glucopyranose, a novel inhibitor specific for maltose-producing amylase. AB - A novel inhibitor against maltose-producing alpha-amylase was prepared via stepwise degradation of a high-molecular-weight acarbose (HMWA) using Thermus maltogenic amylase (ThMA). The structure of the purified inhibitor was determined to be alpha-D-glucopyranosyl-alpha-acarviosinyl-D-glucopyranose (GlcAcvGlc) by (13)C NMR and MALDI-TOF/MS. Progress curves of PNPG2 hydrolysis by various amylolytic enzymes, including MGase, ThMA, and CDase I-5, in the presence of acarbose or GlcAcvGlc indicated a slow-binding mode of inhibition. Analytical ultracentrifugation and X-ray crystallography analyses revealed that the presence of GlcAcvGlc increased the dimerization of ThMA. The formation of dimer complexed with GlcAcvGlc might induce a conformational change in ThMA, leading to a two step inhibition process. The inhibition potency of GlcAcvGlc for MGase, ThMA, and CDase I-5 was 3 orders of magnitude higher than that of acarbose. PMID- 12119027 TI - Identification of N- and C-terminal amino acids of Lhca1 and Lhca4 required for formation of the heterodimeric peripheral photosystem I antenna LHCI-730. AB - Apoproteins of higher plant light-harvesting complexes (LHC) share considerable amino acid sequence identity/similarity. Despite this fact, they occur in different oligomeric states (i.e., monomeric, dimeric, and trimeric). As a step toward understanding the underlying structure requirements for different oligomerization behavior, we analyzed whether amino acids at the N- and C-termini of Lhca1 and Lhca4 are involved in the formation of the heterodimeric LHCI-730. Using altered proteins produced by deletion or site-directed mutagenesis for reconstitution, we were able to identify amino acids required for the assembly of LHCI-730. At the N-terminus of Lhca1, W4 is involved in heterodimerization. This interaction probably depends on aromatic properties because only replacement of W4 by F resulted in dimer formation. Also, at the C-terminus of Lhca1, W seems to play a crucial role for interaction with Lhca4. A detailed analysis by point mutants revealed the importance of an aromatic residue at position 185. One or more other amino acid(s) located downstream of position 188 may exert additional stabilizing effects, presumably in a cooperative way. The scenario for Lhca4 is different. Dimerization broke down only after the deletion of the entire extrinsic N- or C-terminal region, demonstrating that the termini of Lhca4 are not involved in strong interactions with Lhca1 decisive for dimerization. At the N-terminus, dimerization was abolished after the removal of the same number of amino acids at which monomer formation failed. Site-specific mutagenesis of the amino acid decisive for LHC-formation in a deletion study demonstrated that its character is of no importance for dimerization and, therefore, that abolition of dimer formation may be the consequence of a loss in monomer formation. At the C terminus of Lhca4, an even higher number of amino acids than required for monomer formation could be removed without the loss of dimerization. The decisive position is I168, located in the third transmembrane region. Because all point mutants of I168 in the full-length protein yielded dimers, failure of dimerization may be caused by either falling below a critical length of the polypeptide chain, resulting in the loss of too many weak interactions, or by too strong an impairment of Lhca4-folding. Interestingly, N- and C-terminal mutants of Lhca4 not able to form stable monomers formed stable dimers, indicating stabilization of labile monomeric complexes by the Lhca1 subunit in dimerization. Finally, the significance for dimer formation of amino acids in other parts of Lhca1 and Lhca4 which may be involved, besides the amino acids identified here in the specific assembly of the heterodimeric LHCI-730, is discussed. Their identification will result in a better understanding of structure characteristics determining the different oligomerization behavior of LHCs. PMID- 12119028 TI - Effect of binding of Cd2+ on bacterial reaction center mutants: proton-transfer uses interdependent pathways. AB - In bacterial reaction center of Rhodobacter sphaeroides, Cd2+ binds in stoichiometric amount to the protein. In the wild type, this results into a notable decrease of the rates of electron-transfer between the two quinone acceptors after the first (kAB(1)) and second flash (kAB(2)). We have studied these effects in two single mutants, L209PY and L209PF. L209Pro is situated in a protein region rich in hydrogen-bond networks involving water molecules. We show that (1) the combined effects of Cd2+ binding and point mutations have a cumulative consequence in the two mutants, decreasing very substantially the observed rates of electron-transfer. Interestingly, the [Cd2+] titration curves of kAB(2) in the L209PY and L209PF mutants are nearly superimposable to those previously reported for the M17DN and L210DN mutants (Paddock, M. L., Feher, G., and Okamura, M. Y. (2000) Proc. Natl. Acad. Sci U.S.A. 97, 1548-1553). These observations suggest a common effect of all of these mutations (L209, M17, L210) on the protonation state of the histidine cluster to which Cd2+ binds; (2) in the L209PY mutant, the pH titration curves of kAB(1), kAB(2), and k(H)(+), the proton transfer rate at the second flash, are systematically downshifted by 1.5-2 pH units in the presence of 300 microM Cd2+, similarly to the wild type RCs (Gerencser, L., and Maroti, P. (2001) Biochemistry 40, 1850-1860). We propose that Cd2+ binding influences the electrostatics of interdependent ways of proton penetration within the protein, involving at least, directly or indirectly, L209P, L210D, and M17D, probably in conjunction with hydrogen-bonded connected water molecules. PMID- 12119029 TI - Aggregation of LHCII Leads to a Redistribution of the Triplets over the Central Xanthophylls in LHCII. AB - We present laser flash-induced triplet-minus-singlet (TmS(flash)) and absorbance detected-magnetic-resonance (TmS(ADMR)) measurements on the light-harvesting chlorophyll a/b pigment-protein complex (LHCII) from pea. We investigated the influence of LHCII aggregation on xanthophyll triplet formation. The effect of aggregation was previously studied using TmS(ADMR) [van der Vos et al. (1994) Biochim. Biophys. Acta 1208, 243-250] for LHCII from spinach, and it was concluded that aggregation leads to a large increase of the amount of intertrimer triplet transfer. However, a similar study on LHCII from pea with the use of TmS(flash) measurements [Barzda et al. (1998) Biochemistry 37, 546-561] showed much smaller effects. To resolve this apparent discrepancy and to compare the results of TmS(ADMR) and TmS(flash) measurements, we used both techniques to study LHCII from pea, applying an identical aggregation procedure in both cases. It appears that aggregation does not lead to an increase of intertrimer triplet transfer as thought before but to a redistribution of the triplets over the two central xanthophylls (mainly lutein) that are present in each monomeric subunit of LHCII. Moreover, it is argued that the TmS band at 525 nm is due to lutein instead of violaxanthin as was reported in earlier studies. It is concluded that aggregation leads to a change in chlorophyll-xanthophyll interactions, which might explain the large change in excited-state lifetime of chlorophyll a in LHCII upon aggregation. This change in lifetime is possibly related to the phenomenon of nonphotochemical quenching in green plants, which is an important protective regulatory mechanism, that lowers the probability of photoinhibition. PMID- 12119030 TI - Biotin synthase is a pyridoxal phosphate-dependent cysteine desulfurase. AB - Biotin synthase (BioB) is an iron-sulfur dimeric enzyme which catalyzes the last step in biotin synthesis. The reaction consists of the introduction of a sulfur atom into dethiobiotin. It is shown here that BioB displays a significant cysteine desulfurase activity, providing it with the ability to mobilize sulfur from free cysteine. This activity is dependent on pyridoxal 5'-phosphate (PLP) and dithiothreitol and proceeds through a protein-bound persulfide. Like other cysteine desulfurases, BioB binds 1 equiv of PLP. By site-directed mutagenesis, two conserved cysteines, Cys97 and Cys128, are shown to be critical for cysteine desulfuration and are good candidates as the site for a persulfide. Since biotin synthase activity is greatly increased by PLP and cysteine, even though it does not exceed 1 nmol of biotin/nmol of monomer, it is proposed that cysteine desulfuration is intimately linked to biotin synthesis. New scenarios for sulfur insertion into dethiobiotin, in which cysteine persulfides play a key role, are discussed. PMID- 12119031 TI - Spectroscopic and kinetic analyses reveal the pyridoxal 5'-phosphate binding mode and the catalytic features of Treponema denticola cystalysin. AB - To obtain insight into the functional properties of Treponema denticola cystalysin, we have analyzed the pH- and ligand-induced spectral transitions, the pH dependence of the kinetic parameters, and the substrate specificity of the purified enzyme. The absorption spectrum of cystalysin has maxima at 418 and 320 nm. The 320 nm band increases at high pH, while the 418 nm band decreases; the apparent pK(spec) of this spectral transition is about 8.4. Cystalysin emitted fluorescence at 367 and 504 nm upon excitation at 320 and 418 nm, respectively. The pH profile for the 367 nm emission intensity increases above a single pK of approximately 8.4. On this basis, the 418 and 320 nm absorbances have been attributed to the ketoenamine and substituted aldamine, respectively. The pH dependence of both log k(cat) and log k(cat)/K(m) for alpha,beta-elimination reaction indicates that a single ionizing group with a pK value of approximately 6.6 must be unprotonated to achieve maximum velocity. This implies that cystalysin is more catalytically competent in alkaline solution where a remarkable portion of its coenzyme exists as inactive aldamine structure. Binding of substrates or substrate analogues to the enzyme over the pH range 6-9.5 converts both the 418 and 320 nm bands into an absorbing band at 429 nm, assigned to the external aldimine in the ketoenamine form. All these data suggest that the equilibrium from the inactive aldamine form of the coenzyme shifts to the active ketoenamine form on substrate binding. In addition, reinvestigation of the substrate spectrum of alpha,beta-elimination indicates that cystalysin is a cyst(e)ine C-S lyase rather than a cysteine desulfhydrase as claimed previously. PMID- 12119032 TI - Expression, purification, and characterization of two N,N-dimethyltransferases, tylM1 and desVI, involved in the biosynthesis of mycaminose and desosamine. AB - Methylation catalyzed by an S-adenosylmethionine- (AdoMet-) dependent methyltransferase is an effective means to alter the hydrophilicity and/or nucleophilicity of a molecule. While a large number of enzymes capable of catalyzing methylation at carbon, oxygen, sulfur, and nitrogen atoms are known, only a few are able to catalyze N,N-dimethylation. Mycaminose and desosamine are aminohexoses found in several macrolide antibiotics, such as tylosin and methymycin, respectively. Both sugars contain a C-3 N,N-dimethylamino group which has been shown to confer the biological activity of these unusual sugars. Recently, sequence analysis as well as genetic studies has led to the assignment of tylM1 in the tylosin biosynthetic gene cluster and desVI in the methymycin biosynthetic gene cluster as genes encoding the corresponding N,N dimethyltransferases. To verify the proposed roles of the tylM1 and desVI genes, we have overexpressed and purified their encoded products, synthesized the predicted substrates, and characterized the catalytic function of these proteins. Our studies showed that TylM1 and DesVI are homodimeric proteins and have nearly identical biochemical properties. These enzymes do not have strong preference for binding either the unmethylated substrate or the monomethylated intermediate. It is the chemical reactivity of the nitrogen functional group that determines the relative rate of a particular methylation step. Thus, our results not only establish TylM1 and DesVI as new members of a small family of enzymes that are capable of catalyzing N,N-dimethylation of an amino group but also provide evidence indicating that the methylation catalyzed by AdoMet-dependent methyltransferases proceeds in a stepwise manner and is nucleophilic in nature. PMID- 12119033 TI - Timing of epimerization and condensation reactions in nonribosomal peptide assembly lines: kinetic analysis of phenylalanine activating elongation modules of tyrocidine synthetase B. AB - The cyclic decapeptide antibiotic tyrocidine has D-Phe residues at positions 1 and 4, produced during peptide chain growth from L-Phe residues by 50 kDa epimerase (E) domains embedded, respectively, in the initiation module (TycA) and the TycB3 module of the three-subunit (TycABC), 10-module nonribosomal peptide synthetase. While the initiation module clearly epimerizes the aminoacyl thioester Phe1-S-TycA intermediate, the timing of epimerization versus peptide bond condensation at internal E domains has been less well characterized in nonribosomal peptide synthetases. In this study, we use rapid quench techniques to evaluate a three-domain (ATE) and a four-domain version (CATE) of the TycB3 module and a six-domain fragment (ATCATE) of the TycB2(-3) bimodule to measure the ability of the E domain in the TycB3 module to epimerize the aminoacyl thioester Phe-S-TycB3 and the dipeptidyl-S-enzyme (L-Phe-L-Phe-S-TycB3 if L-Phe-D Phe-S-TycB3). The chiralities of the Phe-S-enzyme and Phe-Phe-S-enzyme species over time were determined by hydrolysis and chiral TLC separations, allowing for the clear conclusion that epimerization in the internal TycB3 module occurs preferentially on the peptidyl-S-enzyme rather than the aminoacyl-S-enzyme, by a factor of about 3000/1. In turn, this imposes constraints on the chiral selectivity of the condensation (C) domains immediately upstream and downstream of E domains. The stereoselectivity of the upstream C domain was shown to be L selective at both donor and acceptor sites ((L)C(L)) by site-directed mutagenesis studies of an E domain active site residue and using the small-molecule surrogate D-Phe-Pro-L-Phe-N-acetylcysteamine thioester (D-Phe-Pro-L-Phe-SNAC) and D-Phe-Pro D-Phe-SNAC as donor probes. PMID- 12119034 TI - Effect of variations in the structure of a polyleucine-based alpha-helical transmembrane peptide on its interaction with phosphatidylcholine bilayers. AB - High-sensitivity differential scanning calorimetry (DSC) and Fourier transform infrared (FTIR) spectroscopy were used to study the interaction of an alpha helical transmembrane peptide, acetyl-Lys2-Leu24-Lys2-amide (L24), and odd-chain members of the homologous series of n-saturated diacylphosphatidylcholines. An analogue of L24, in which the lysine residues were all replaced by 2,3 diaminopropionic acid, and another, in which a leucine residue at each end of the polyLeu sequence was replaced by a tryptophan, were also studied. At low peptide concentrations, the DSC thermograms exhibited by these lipid/peptide mixtures are resolvable into two components. One of these components is fairly narrow, highly cooperative, and exhibits properties which are similar to but not identical with those of the pure lipid. In addition, the transition temperature and cooperativity of this component, and its fractional contribution to the total enthalpy change, decrease with an increase in peptide concentration, more or less independently of phospholipid acyl chain length. The other component is very broad and predominates at high peptide concentrations. These two components have been assigned to the chain-melting phase transitions of populations of peptide poor and peptide-enriched lipid domains, respectively. Moreover, when the mean hydrophobic thickness of the PC bilayer is less than the peptide hydrophobic length, the peptide-associated lipid melts at higher temperatures than does the bulk lipid and vice versa. In addition, the chain-melting enthalpy of the broad endotherm does not decrease to zero even at high peptide concentrations, suggesting that these peptides reduce somewhat but do not abolish the cooperative gel/liquid-crystalline phase transition of the lipids with which it is in contact. Our DSC results indicate that the width of the broad phase transition observed at high peptide concentration is inversely but discontinuously related to hydrocarbon chain length. Our FTIR spectroscopic data indicate that these peptides form a very stable alpha-helix under all of our experimental conditions but that small distortions of their alpha-helical conformation are induced in response to mismatch between peptide hydrophobic length and gel-state bilayer hydrophobic thickness. We also present evidence that these distortions are localized to the N- and C-terminal regions of these peptides. Interestingly, replacing the terminal Lys residues of L24 by 2,3-diaminopropionic acid residues actually attenuates the hydrophobic mismatch effects of the peptide on the thermotropic phase behavior of the host PC bilayer, in contrast to the predictions of the snorkel hypothesis. We rationalize this attenuated hydrophobic mismatch effect by postulating that the 2,3-diaminopropionic acid residues are too short to engage in significant electrostatic and hydrogen-bonding interactions with the polar headgroups of the host phospholipid bilayer, even in the absence of any hydrophobic mismatch between incorporated peptide and the bilayer. Similarly, the reduced hydrophobic mismatch effect also observed when the two terminal Leu residues of L24 are replaced by Trp residues is rationalized by considering the lower energetic cost of exposing the Trp as opposed to the Leu residues to the aqueous phase in thin PC bilayers and the higher cost of inserting the Trp as opposed to the Leu residues into the hydrophobic cores of thick PC bilayers. PMID- 12119035 TI - Targeting of nonkaryophilic cell-permeable peptides into the nuclei of intact cells by covalently attached nuclear localization signals. AB - Dermaseptins are a family of antimicrobial peptides that lyse target bacterial cells by destabilization of their membranes. Here we present a novel application of a peptide derived from the dermaseptin S4, S4(13). At nontoxic concentrations, fluorescently labeled S4(13) was able to penetrate intact cultured HeLa cells but essentially failed to enter their nuclei despite its low molecular weight. Covalent attachment of nuclear localization signal (NLS) motifs of the SV40-T antigen and of the HIV-1 Rev protein (ARM) conferred karyophilic properties upon the S4(13). The resulting peptides, which were designated as PV-S4(13) and RR S4(13) penetrated into intact HeLa cells and were able to accumulate within the cells' nuclei. In studies with digitonin-permeabilized cells, nuclear uptake of the PV-S4(13) and the RR-S4(13) peptides showed the same features that characterize active nuclear import. Nuclear import was observed at 37 degrees C, was ATP-dependent, and was inhibited by the free peptides bearing the SV40 NLS and the Rev and Tat ARMs. Microinjected S4(13) remained in the cytoplasm while microinjected RR-S4(13) was translocated into the cells' nuclei. The new type of cell-permeable "karyophilic" peptides described here may be of potential application as a lead compound for therapeutic purposes, as a tool to study nucleocytoplasmic shuttling in intact cells, and for the delivery of peptides to the nucleus. PMID- 12119037 TI - Histone h1(0) and its carboxyl-terminal domain bind in the major groove of DNA. AB - The binding of histone H1(0) to T4 bacteriophage DNA was investigated using thermal denaturation of the DNA titrated with varying concentrations of protein. The H1(0) used was expressed in and purified from a strain of E. coli and is therefore homogeneous with respect to H1 subtype and posttranslational modifications. Two types of T4 DNA were used: wild-type, which contains a modification of the cytosine residues that projects into the major groove: and a mutant type, which lacks the modification of the cytosines. Data were compared to simulated thermal denaturation curves to determine estimates for binding affinity and binding site size in base pairs of the protein. Analysis of the data yielded values of 10(8) M(-1) for K, the binding affinity, and 10 base pairs for n, the number of base pairs covered by one protein, for the mutant T4 DNA. Analysis of the wild-type DNA data suggested that the glucose projecting into the major groove of this DNA decreases the number of sites to which the H1(0) protein can bind, indicating that there are interactions between the protein and the major groove of DNA. The binding site size on this DNA is 10 base pairs, the same as on the unmodified DNA. The affinity for wild-type DNA is slightly higher, 10(9) M( 1). Data were collected and analyzed for binding of two domains of the protein as well, the carboxyl-terminal domain and the central globular domain. Binding of the carboxyl-terminal domain was quantitatively and qualitatively similar to that of the full-length protein. In contrast, binding of the globular domain was quite different: it binds much more weakly, with a K of 6 x 10(4) M(-1), and covers fewer base pairs, with an n of 3. Also, there was no evidence that the globular domain interacts with the major groove of DNA. PMID- 12119036 TI - Role of conserved transmembrane cationic amino acids in the prostaglandin transporter PGT. AB - The prostaglandin transporter "PGT" interacts electrostatically with its anionic substrate, based on inhibition by the disulfonic stilbenes [Chan, B. S. (1998) J. Biol. Chem. 273, 6689-6697], inhibition by the thiol-reactive anion sodium (2 sulfonatoethyl)methanethiosulfonate (MTSES) [Chan, B. S. (1999) J. Biol. Chem. 274, 25564-25570], and the requirement for a negatively charged 1-position carboxyl on the substrate [Itoh, S. (1996) Mol. Pharm. 50, 736-742]. Here we found that modification of positively charged residues on wild-type PGT by arginine- and lysine-specific reagents significantly inhibited transport. We previously found that the binding site of PGT is formed, at least in part, by its membrane-spanning segments [Chan, B. S. (1999) J. Biol. Chem. 274, 25564-25570]. Three charged residues within predicted transmembrane spans (E78, R560, and K613) are conserved in PGT and in related transporters. Substitution of the anionic residue E78 (E78D and E78C) produced an essentially functional transporter, whereas substitution of the cationic residues with neutral residues (R560N and K613Q) resulted in poorly functional transporters. Immunoblotting revealed similar expression levels of wild-type and mutant transporters, and immunostaining indicated correct targeting. Conservative charge substitutions (R560K, K613R, and K613H) resulted in generally functional transporters. In contrast, R560N was nonfunctional, whereas the substrate affinity of K613G decreased greater than 50-fold. Conservative substitutions retaining the charge at position 613 (K613R and K613H) restored the substrate affinity, suggesting a direct role of K613 in substrate binding. Double-neutral mutants E78G/R560C and E78G/K613C were inactive, indicating that these residues are not simply charge paired. Our results suggest that an arginine at position 560 is critical for maximal substrate translocation, and that a positively charged side chain at position 613 contributes to electrostatic binding of the anionic substrate. PMID- 12119038 TI - SH2 domains from suppressor of cytokine signaling-3 and protein tyrosine phosphatase SHP-2 have similar binding specificities. AB - Suppressor of cytokine signaling-3 (SOCS-3) and the protein tyrosine phosphatase SHP-2 both regulate signaling by cytokines of the interleukin-6 family, and this is dependent upon recruitment to tyrosine 757 in the shared cytokine receptor subunit gp130. To better explore the overlap in ligand binding specificities exhibited by these two signaling regulators, we have mapped the phosphopeptide binding preferences of the SH2 domains from SOCS-3 and SHP-2. Degenerate phosphopeptide libraries were screened against recombinantly produced SH2 domains to determine the sequences of optimal phosphopeptide ligands. We found that the consensus ligand binding motif for SOCS-3 was pY-(S/A/V/Y/F)-hydrophobic-(V/I/L) hydrophobic-(H/V/I/Y), while the consensus motif for SHP-2 was pY-(S/T/A/V/I)-X (V/I/L)-X-(W/F). We validated these data through the design of phosphopeptide ligands based on the consensus motifs and found that these bound to SOCS-3 and SHP-2 with high affinity. Finally, we have compared the affinity of SOCS-3 for binding to phosphopeptides representing putative docking sites in the gp130, leptin and erythropoietin receptors. While SOCS-3 binds with much higher affinity to a gp130 phosphopeptide than to phosphopeptides derived from the other receptors, multiple SOCS-3 binding sites are predicted to exist in the leptin and erythropoietin receptors which may compensate for weaker binding to individual sites. PMID- 12119039 TI - New insights into the heme cavity structure of catalase-peroxidase: a spectroscopic approach to the recombinant synechocystis enzyme and selected distal cavity mutants. AB - Catalase-peroxidases (KatGs) are heme peroxidases with homology to yeast cytochrome cperoxidase (CCP) and plant ascorbate peroxidases (APXs). KatGs exhibit a peroxidase activity of broad specificity and a high catalase activity, which strongly depends on the presence of a distal Trp as part of the conserved amino acid triad Arg-Trp-His. By contrast, both CCP and APX do not have a substantial catalase activity despite the presence of the same triad. Thus, to elucidate structure-function relationships of catalase-peroxidases (for which no crystal structure is available at the moment), we performed UV-Vis and resonance Raman studies of recombinant wild-type KatG from the cyanobacterium SynechocystisPCC 6803 and the distal side variants (His123-->Gln, Glu; Arg119- >Ala, Asn; Trp122-->Phe, Ala). The distal cavity of KatG is very similar to that of the other class I peroxidases. A H-bond network involving water molecules and the distal Trp, Arg, and His is present, which connects the distal and proximal sides of the heme pocket. However, distal mutation not only affects the heme Fe coordination state and perturbs the proximal Fe-Im bond, as previously observed for other peroxidases, but also alters the stability of the heme architecture. The charge of the distal residues appears particularly important for maintaining the heme architecture. Moreover, the Trp plays a significant role in the distal H bonding, much more pronounced than in CCP. The relevance of these findings for the catalase activity of KatG is discussed in light of the complete loss of catalase activity in the distal Trp mutants. PMID- 12119040 TI - Substrate specificity and sequence preference of G:T mismatch repair: incision at G:T, O6-methylguanine:T, and G:U mispairs in DNA by human cell extracts. AB - Extracts of two human glioma cell lines (lacking O6-methylguanine DNA methyltransferase) (i.e., A1235 and its alkylation-resistant derivative A1235 MR4) were examined for their ability to execute strand incision at different base mismatches in model (45-bp) DNA. These heteroduplex substrates were of the same sequence except for the presence, at the same site, of one of three mispairs: G:T, O6-methylguanine:T (m6G:T), and G:U. The parental (A1235) extract, when supplemented with ATP and human thymine DNA glycosylase (TDG), acted proficiently on all three substrates, incising immediately 5' to the mismatched thymine or uracil residue. In contrast, the derivative extract, under the same conditions, recognized only the G:U substrate. The activity of the A1235 extract toward the G:T (or m6G:T) substrate was markedly reduced in the absence of ATP, whereas the G:U substrate was incised rapidly by both extracts irrespective of the addition of ATP. These combined data confirm and extend our earlier findings demonstrating that human cells possess two G:T incision activities, one efficient and ATP dependent and the other inefficient and ATP-independent. The derivative extract lacks the former activity but retains the latter activity. In substrate competition assays, the G:U substrate inhibited the ATP-dependent G:T incision activity to a greater extent than did the G:T substrate itself. Given the well known substrate preference of TDG for G:U as compared to G:T, this unexpected result implies that TDG may be an integral component of the ATP-dependent G:T incision machinery in human cells. Finally, the base 5' to the mismatched G in the G:T mispair conferred sequence preference on the A1235 extract in the presence of ATP and TDG, with a pyrimidine (especially cytosine) being much favored over a purine. This latter observation suggests that the ATP-dependent G:T incision activity is designed to repair deaminated 5-methycytosine lesions in CpG islands, the methylation of which is linked to control of gene expression. PMID- 12119041 TI - Conformational stability of legume lectins reflect their different modes of quaternary association: solvent denaturation studies on concanavalin A and winged bean acidic agglutinin. AB - Thermodynamic parameters associated with the unfolding of the legume lectin, WBA II, were determined by isothermal denaturation. The analysis of isothermal denaturation data provided values for conformational stability and heat capacity for WBA II unfolding. To explore the role of intersubunit contact in stability, we carried out similar studies under identical conditions on Concanavalin A, a legume lectin of nearly similar size, buried hydrophobic surface area and tertiary structure to that of WBA II but with a different oligomerization pattern. Both proteins showed a reversible two-state unfolding with guanidine hydrochloride. As expected, the change in heat capacity upon unfolding was similar for both proteins at 3.5 and 3.7 kcal mol(-1) K(-1) for Concanavalin A and WBA II, respectively. Although the deltaG(H20) at the maximum stability of both proteins is around 16 kcal/mol, Concanavalin A exhibits greater stability at higher temperatures. The T(g) obtained for Concanavalin A and WBA II were 21 degrees C apart at 87.2 and 66.6 degrees C, respectively. The higher conformational stability at higher temperatures and the T(g) of Concanavalin A as compared to that of WBA II are largely due to substantial differences in the degree of subunit contact in these dimeric proteins. Ionic interactions and hydrogen bonding between the monomers of the two proteins also seem to play a significant role in the observed stability differences between these two proteins. PMID- 12119042 TI - The membrane-dipped neuronal SNARE complex: a site-directed spin labeling electron paramagnetic resonance study. AB - The formation of the soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) complex is an essential process for membrane fusion and the neurotransmitter release in neurons. As an initial step toward the determination of the membrane topology of the SNARE complex, residues at the membrane-water interface were investigated with site-specific spin labeling electron paramagnetic resonance. EPR analysis revealed that the basic amino acid-rich interfacial region, which is universal for all transmembrane SNARE proteins, inserts into the membrane, eliminating the gap between the core complex and the membrane. The result raises the possibility that core complex formation directly leads to the apposition of two membranes, which could facilitate membrane fusion. PMID- 12119043 TI - Development of high-affinity ligands and photoaffinity labels for the D-fructose transporter GLUT5. AB - The GLUT5 transporter catalyses the specific uptake of D-fructose and can accept this hexose in its furanose and pyranose ring forms. The transporter does not accept fructose epimers and has very limited tolerance of bulky groups substituted at the 2-, 3-, 4- and 5-OH positions [Tatibouet, Yang, Morin and Holman (2000) Bioorg. Med. Chem. 8, 1825-1833]. To further explore whether bulky groups can be tolerated at the primary OH positions, a D-fructose analogue with an allylamine group substitution to replace the 1-OH group was synthesized and was found to be quite well tolerated ( K (i)=27.1 mM). However, this analogue occurs in multiple ring forms. By contrast, 2,5-anhydro-D-mannitol is a symmetrical molecule that occurs only in a furanose ring form in which C-1 and C 6 are equivalent. We have therefore synthesized new 2,5-anhydro-D-mannitol analogues (substituted at the equivalent of the 6-OH of D-fructose) and from studies in Chinese hamster ovary cells expressing GLUT5 cells report that (i) the allylamine derivative of 2,5-anhydro-D-mannitol is well tolerated ( K (i)=2.66 mM); (ii) introduction of a di-nitrophenyl-substituted secondary amine group enhances affinity ( K (i)=0.56 mM); (iii) introduction of amide-linked biotinylated photolabel moieties is possible without loss of affinity relative to 2,5-anhydro-D-mannitol but a small secondary amine spacer between the biotinylated photolabelling moiety and the fructofuranose ring increases affinity (fructose photolabel 2; K (i)=1.16 mM); (iv) introduction of a hydrophilic tartarate spacer between biotin and the diazirine photoreactive groups can be accomplished without reduction in affinity and (v) photoactivation of biotinylated fructose photolabels leads to specific biotin tagging of GLUT5. These data suggest that substitution of a secondary amine group (-NH) to replace the C-6 (or C-1) -OH of 2,5-anhydro-D-mannitol results in compounds of high affinity; the affinity is enhanced over 10-fold compared with D-fructose. PMID- 12119044 TI - Involvement of delta-aminolaevulinate synthase encoded by the parasite gene in de novo haem synthesis by Plasmodium falciparum. AB - The malaria parasite can synthesize haem de novo. In the present study, the expression of the parasite gene for delta-aminolaevulinate synthase (Pf ALAS ) has been studied by reverse transcriptase PCR analysis of the mRNA, protein expression using antibodies to the recombinant protein expressed in Escherichia coli and assay of ALAS enzyme activity in Plasmodium falciparum in culture. The gene is expressed through all stages of intra-erythrocytic parasite growth, with a small increase during the trophozoite stage. Antibodies to the erythrocyte ALAS do not cross-react with the parasite enzyme and vice versa. The recombinant enzyme activity is inhibited by ethanolamine and the latter inhibits haem synthesis in P. falciparum and growth in culture. The parasite ALAS is localized in the mitochondrion and its import into mitochondria in a cell-free import assay has been demonstrated. The import is blocked by haemin. On the basis of these results, the following conclusions are arrived at: PfALAS has distinct immunological identity and inhibitor specificity and is therefore a drug target. The malaria parasite synthesizes haem through the mitochondrion/cytosol partnership, and this assumes significance in light of the presence of apicoplasts in the parasite that may be capable of independent haem synthesis. The Pf ALAS gene is functional and vital for parasite haem synthesis and parasite survival. PMID- 12119045 TI - A non-radioactive method for the assay of many serine/threonine-specific protein kinases. AB - The generation of drugs that modulate the activities of particular protein kinases has become a prime focus of the pharmaceutical and biotechnology industry. Consequently, improved methods for the development of high-throughput screening formats for these enzymes is a high priority. In the present study, we have designed three generic peptide substrates that can be used to assay a diverse range of protein kinases. These peptides share a common seven-residue epitope that includes the site of phosphorylation, and against which we have generated a phospho-specific antibody. Thus a large number of serine/threonine specific protein kinases can be screened using a simple non-radioactive format. PMID- 12119046 TI - Cloning, sequencing and heterologous expression of the gene for lupanine hydroxylase, a quinocytochrome c from a Pseudomonas sp. AB - The gene encoding the enzyme lupanine hydroxylase was isolated by PCR using chromosomal DNA from a lupanine-utilizing Pseudomonas sp. as template and primers based on the sequences of the N- and C-termini of the purified protein. The derived sequence for the mature gene product gave a protein with an M (r) of 72256, in good agreement with the value found by SDS/PAGE of the pure enzyme, and contained the sequences of several peptides obtained after endoproteinase Lys-C digestion of the pure enzyme. The gene, under the transcriptional control of a phoA promotor and with the Escherichia coli alkaline phosphatase signal sequence, was expressed in E. coli containing a plasmid expressing the genes for cytochrome c maturation proteins constitutively. Haem-containing inactive protein in inclusion bodies was renatured and reactivated with pyrroloquinoline quinone (PQQ) and Ca(2+) to give active enzyme. The lupanine hydroxylase (luh) gene coded for a protein with a cleavable 26-residue signal sequence at its N-terminus, required for the transport of the enzyme to its periplasmic location. Analysis of the protein sequence showed that it contains two domains, a large PQQ-binding N terminal domain and a smaller cytochrome c C-terminal domain. Comparison of the derived sequence with those of other proteins showed considerable similarity with other quino(haemo)proteins, including alcohol dehydrogenases from a variety of bacteria. The PQQ-binding domain sequence contains W motifs, characteristic of the eight-bladed "propeller" structure of methanol dehydrogenase, but lacks the unusual disulphide ring structure formed from two adjacent cysteines seen in this enzyme. The C-terminus shares some similarity with bacterial cytochrome c and includes the haem-binding consensus sequence CXXCH. PMID- 12119047 TI - S-S bonds are not required for the sonochemical formation of proteinaceous microspheres: the case of streptavidin. AB - Proteinaceous microspheres can be prepared using the sonochemical method. However, it is known that these proteins should possess at least one cysteine residue in order to obtain stable microspheres using this method. In the present study, we have produced streptavidin microspheres, using the sonochemical method, from streptavidin, which does not have any cysteine residues. PMID- 12119048 TI - Oligolysine-based saccharide clusters: synthesis and specificity. AB - In search of specific and highly selective sugar clusters for cell receptors, such as membrane lectins, various disaccharides were coupled to small peptide cores through an amide bond. In a first step, the reducing disaccharides, i.e. lactose and three different dimannoses, were converted into glycosyl-pyroglutamyl beta-alanine derivatives. The free carboxylic group of these conjugates was then coupled to the alpha and epsilon amino groups of the core peptide (Lys( n )-Ala Cys-NH2) with n =1 to 5, with complete substitution leading to homogeneous glycoclusters. The thiol group of the cysteine residue was used to tag the glycosylated oligolysines upon reaction with fluorescein iodoacetamide. The affinity of these glycoclusters towards two plant lectins was assessed by surface plasmon resonance. The selectivity of their cell uptake was investigated by flow cytometry using two types of cells: a human hepatoma cell line (HepG2 cells) expressing the plasma membrane galactose-specific lectin, and monocyte-derived dendritic cells expressing the plasma membrane mannose-specific lectin. The glycoclusters containing four or five disaccharides were shown to bind plant lectins and cell surface membrane lectins with a narrow selectivity and with a high affinity. PMID- 12119050 TI - Introduction. RSV and RAD: possibilities for prevention? The link between respiratory syncytial virus and reactive airway disease. PMID- 12119049 TI - Abundant expression of Dec1/stra13/sharp2 in colon carcinoma: its antagonizing role in serum deprivation-induced apoptosis and selective inhibition of procaspase activation. AB - The basic helix-loop-helix (bHLH) proteins are intimately associated with developmental events such as cell differentiation and lineage commitment. The HLH domain in the bHLH motif is responsible for dimerization, whereas the basic region mediates DNA binding. Based on sequence alignment and domain analysis, differentially expressed in chondrocytes/stimulated with retinoic acid/split and hairy-related proteins (DEC/STRA/SHARPs) represent a new class of bHLH proteins. The present study describes the functional characterization of DEC1. Subtractive experiments and blotting analyses demonstrated that DEC1 was highly expressed in colon carcinomas, but not in the adjacent normal tissues. Several cell cycle blockers markedly induced DEC1 expression. Stable transfectants with a tetracycline-inducible construct demonstrated that DEC1 caused proliferation inhibition, antagonized serum deprivation-induced apoptosis and selectively inhibited the activation of procaspases. These activities were highly correlated with the abundance of tetracycline-induced DEC1. Stable transfectants expressing a mutant DEC1 (lacking the DNA-binding domain) exhibited neither proliferation inhibition nor apoptotic antagonism, which suggests that DNA binding is required for these actions. Enzymic assays and immunoblotting analyses demonstrated that induction of DEC1 by tetracycline significantly decreased the activation of procaspases 3, 7 and 9 but not procaspase 8. The selective suppression on the activation of procaspases 3, 7 and 9 over procaspase 8 suggests that DEC1 mediated anti-apoptosis is achieved by blocking apoptotic pathways initiated via the mitochondria. The results functionally distinguish DEC1 from other bHLH proteins and directly link this factor to oncogenesis. PMID- 12119051 TI - Asthma: a major pediatric health issue. AB - The incidence, prevalence, and mortality of asthma have increased in children over the past three to four decades, although there has been some decline in the most recent decade. These trends are particularly marked and of greatest concern in preschool children. Internationally, there are huge variations among countries and continents, as demonstrated by the International Study of Asthma and Allergies in Childhood. In general, asthma rates were highest in English-speaking countries (UK, New Zealand, Australia, and North America) and some Latin American countries (Peru and Costa Rica), and lowest in South Korea, Russia, Uzbekistan, Indonesia, and Albania. There is currently no unifying hypothesis to explain these trends or any associated risk factors. Environmental factors that may lead to asthma include air pollution; genetic factors, the hygiene hypothesis, and lifestyle differences also play potentially causative roles. Asthma may develop as a result of persistent activation of the immune system alone or in combination with physiologic airway remodeling in early childhood. Further studies are needed to confirm this hypothesis. PMID- 12119052 TI - Clinical perspectives on the association between respiratory syncytial virus and reactive airway disease. AB - Asthma is a leading cause of morbidity and mortality among children worldwide, as is respiratory syncytial virus (RSV). This report reviews controlled retrospective and prospective studies conducted to investigate whether there is an association between RSV bronchiolitis in infancy and subsequent development of reactive airway disease or allergic sensitization. Findings indicate that such a link to bronchial obstructive symptoms does exist and is strongest for children who experienced severe RSV illness that requires hospitalization. However, it is not yet clear what roles genetic predisposition and environmental or other risk factors may play in the interaction between RSV bronchiolitis and reactive airway disease or allergic sensitization. Randomized, prospective studies utilizing an intervention against RSV, such as a passive immunoprophylactic agent, may determine whether preventing RSV bronchiolitis reduces the incidence of asthma. PMID- 12119053 TI - Potential therapeutic implications of new insights into respiratory syncytial virus disease. AB - Viral bronchiolitis is the most common cause of hospitalization in infants under 6 months of age, and 70% of all cases of bronchiolitis are caused by respiratory syncytial virus (RSV). Early RSV infection is associated with respiratory problems such as asthma and wheezing later in life. RSV infection is usually spread by contaminated secretions and infects the upper then lower respiratory tracts. Infected cells release proinflammatory cytokines and chemokines, including IL-1, tumor necrosis factor-alpha, IL-6, and IL-8. These activate other cells and recruit inflammatory cells, including macrophages, neutrophils, eosinophils, and T lymphocytes, into the airway wall and surrounding tissues. The pattern of cytokine production by T lymphocytes can be biased toward 'T-helper-1' or 'T-helper-2' cytokines, depending on the local immunologic environment, infection history, and host genetics. T-helper-1 responses are generally efficient in antiviral defense, but young infants have an inherent bias toward T helper-2 responses. The ideal intervention for RSV infection would be preventive, but the options are currently limited. Vaccines based on protein subunits, live attenuated strains of RSV, DNA vaccines, and synthetic peptides are being developed; passive antibody therapy is at present impractical in otherwise healthy children. Effective vaccines for use in neonates continue to be elusive but simply delaying infection beyond the first 6 months of life might reduce the delayed morbidity associated with infantile disease. PMID- 12119054 TI - Pathophysiological mechanisms for the respiratory syncytial virus-reactive airway disease link. AB - There is substantial epidemiological evidence supporting the concept that respiratory syncytial virus (RSV) lower respiratory tract infection in infancy may be linked to the development of reactive airway disease (RAD) in childhood. However, much less is known concerning the mechanisms by which this self-limiting infection leads to airway dysfunction that persists long after the virus is cleared from the lungs. A better understanding of the RSV-RAD link may have important clinical implications, particularly because prevention of RSV lower respiratory tract infection may reduce the occurrence of RAD later in life. Among the mechanisms proposed to explain the chronic sequelae of RSV infection is the interaction between the subepithelial neural network of the airway mucosa and the cellular effectors of inflammatory and immune responses to the virus. The body of clinical literature linking RSV and RAD is reviewed herein, as are the cellular and molecular mechanisms of neuroimmune interactions and neural remodeling that may underlie this link, and the possibility that preventing the infection may result in a decreased incidence of its chronic sequelae. PMID- 12119055 TI - Immunoprophylaxis of respiratory syncytial virus: global experience. AB - Respiratory syncytial virus (RSV) infects nearly all children by age 2 years, and it causes considerable illness and death in certain high-risk pediatric populations. Historically, treatment for RSV has been symptomatic, and developing a safe and effective vaccine has been a challenge. Therefore, research efforts have turned to passive immunization as the best option to control RSV. Palivizumab, a genetically engineered humanized monoclonal antibody, has been shown to reduce RSV-related hospitalizations significantly, with few adverse effects. It was approved for use in high-risk children in the USA in 1998 and in Europe in 1999; it is now approved for use in more than 45 countries. The efficacy and safety of palivizumab continue to be supported by both clinical trial and outcomes data. PMID- 12119056 TI - An epidemiological study of respiratory syncytial virus associated hospitalizations in Denmark. AB - Respiratory syncytial virus (RSV) is the most common viral pathogen that causes lower respiratory tract infections in infants. Studies have implicated severe RSV infections early in life as a risk factor for subsequent development of reactive airway disease. We are conducting a study to validate RSV-associated diagnoses in the Danish National Patient Registry, to assess whether the incidence of severe RSV infection is increasing in Denmark, to identify predisposing and protective factors for RSV-associated hospitalization in Denmark, and to examine the association of severe RSV infection with reactive airway disease. The influence of various biological, social and environmental factors on hospitalization for RSV infection will be studied through several population-based registers, including the Danish National Birth Cohort: 'Better health for mothers and children'. The RSV hospitalization cases will be compared with control individuals selected within the same population groups on a case-control or a cohort basis in order to produce estimates of age-adjusted and sex-adjusted relative risks (odds ratio and relative risk) for hospitalization associated with various risk factors. Using register linkage and unique registration of exposures collected through interviews and blood samples from the Danish National Birth Cohort, we will be able to resolve the issues referred to above in a very large sample of Danish children. PMID- 12119057 TI - Prevention and treatment of respiratory syncytial virus bronchiolitis and postbronchiolitic wheezing. AB - Respiratory syncytial virus (RSV) is the primary cause of hospitalization for acute respiratory tract illness in general and specifically for bronchiolitis in young children. The link between RSV bronchiolitis and reactive airway disease is not completely understood, even though RSV bronchiolitis is frequently followed by recurrent episodes of wheezing. Therapy with ribavirin does not appear to significantly reduce long-term respiratory outcome of RSV lower respiratory tract infection, and corticosteroid or bronchodilator therapy may possibly improve outcomes only on a short-term basis. No vaccine against RSV is yet available. It is not known whether prophylaxis with RSV intravenous immune globulin or palivizumab can reduce postbronchiolitic wheezing. PMID- 12119058 TI - Losartan and diabetic nephropathy: commentaries on the RENAAL study. AB - The RENAAL (Reduction of Endpoints in NIDDM with the Angiotensin II Antagonist Losartan) study is a multinational, double-blind, randomized, placebo controlled trial which was recently published. It was aimed to evaluate the effect of the angiotensin receptor blocker losartan in patients with diabetic nephropathy. The primary efficacy measure was the time to the first event of the composite end point of a doubling of serum creatinine, end-stage renal disease, or death. The conclusion was that losartan led to significant improvement in renal outcomes, that was beyond that attributable to blood pressure control in patients with type 2 diabetes and nephropathy. The perusal of the report raises concern, regarding to both the patient population as well as the outcome measures. At randomization, the placebo group included more patients with angina, myocardial infarction and lipid disorders than the losartan group. Information on glucose metabolism was disregarded, and data on antihyperglycemic therapy--which may have undesirable influences on cardiac performance--were not included in a multivariate analysis. In addition, only data on first hospitalization were reported, whilst information on total specific-cause hospitalizations was disregarded, thus potentially masking further unfavorable events. Furthermore, creatinine seems not to be a reliable surrogate end point. Based on its mechanism of action, losartan may possess favorable renoprotective properties. However, due to the methodological flaws and the incomplete data in the RENAAL study, the question of the effectiveness and safety of this drug in diabetic nephropathy remains yet unanswered. PMID- 12119059 TI - How hyperglycemia promotes atherosclerosis: molecular mechanisms. AB - Both type I and type II diabetes are powerful and independent risk factors for coronary artery disease (CAD), stroke, and peripheral arterial disease. Atherosclerosis accounts for virtually 80% of all deaths among diabetic patients. Prolonged exposure to hyperglycemia is now recognized a major factor in the pathogenesis of atherosclerosis in diabetes. Hyperglycemia induces a large number of alterations at the cellular level of vascular tissue that potentially accelerate the atherosclerotic process. Animal and human studies have elucidated three major mechanisms that encompass most of the pathological alterations observed in the diabetic vasculature: 1) Nonenzymatic glycosylation of proteins and lipids which can interfere with their normal function by disrupting molecular conformation, alter enzymatic activity, reduce degradative capacity, and interfere with receptor recognition. In addition, glycosylated proteins interact with a specific receptor present on all cells relevant to the atherosclerotic process, including monocyte-derived macrophages, endothelial cells, and smooth muscle cells. The interaction of glycosylated proteins with their receptor results in the induction of oxidative stress and proinflammatory responses 2) oxidative stress 3) protein kinase C (PKC) activation with subsequent alteration in growth factor expression. Importantly, these mechanisms may be interrelated. For example, hyperglycemia-induced oxidative stress promotes both the formation of advanced glycosylation end products and PKC activation. PMID- 12119060 TI - Are proton pump inhibitors the first choice for acute treatment of gastric ulcers? A meta analysis of randomized clinical trials. AB - BACKGROUND: Gastric ulcers are a frequent problem in the United States. Proton pump inhibitors have been shown to increase healing rates and improve clinical symptoms. The objective of this study is to compare gastric ulcer healing rates for patients treated with a proton pump inhibitor (PPI) (omeprazole, rabeprazole, pantoprazole, or lansoprazole), an histamine 2- receptor antagonist (ranitidine) or placebo. METHODS: A literature search was conducted to identify randomized, controlled clinical trials that included a PPI in at least one treatment arm and assessed the gastric ulcer healing rates endoscopically. The healing rates were estimated for each treatment at specific time points, and Rate Ratios (RR) and 95% confidence intervals (CI) were estimated for each trial. RESULTS: Sixteen trials met the inclusion criteria: four compared a PPI versus placebo, nine compared a PPI versus ranitidine (no trials of rabeprazole versus ranitidine met the inclusion criteria), and three compared a newer PPI (lansoprazole, pantoprazole or rabeprazole) versus omeprazole. In relation to ranitidine, the pooled RR of PPIs (lansoprazole, omeprazole and pantoprazole) was 1.33 (95% CI 1.24 to 1.42) at four weeks. In each trial, greater improvement in the studied clinical symptoms was found with the newer PPIs (rabeprazole, pantoprazole and lansoprazole) when compared to omeprazole. CONCLUSION: In this study treatment with PPIs resulted in higher healing rates than ranitidine or placebo. This evidence suggests that the first choice for gastric ulcer treatment for the greater relief of symptoms is one of the newer PPIs. PMID- 12119063 TI - Perceptions of schizophrenia in multi-cultural Britain. AB - OBJECTIVES: The importance of perceptions of mental health problems from the perspectives of different ethnic populations is especially pertinent within a society such as Britain, which is culturally diverse, but employs a classification of psychopathology which is based upon western models. This study aimed to investigate differences in the lay perceptions of mental health problems, particularly schizophrenia, across different ethnic populations living in Britain. Further it aimed to look at the influence ethnic background had in relation to other variables such as age, gender, religion and contact with people with mental health problems. DESIGN: 190 participants from five broad ethnic groups (Afro-Caribbean, Bangladeshi, Indian, Sub-Saharan African and White British) reported their perceptions of schizophrenia symptoms using the Perceptions of Mental Health Problems Questionnaire. RESULTS: Differences were found across ethnic groups in participants' perceptions of schizophrenia symptoms. Differences were consistently shown across the specific symptoms of hallucinatory behaviour, suspiciousness, unusual thought content and alogia. In particular, in comparison with the white British group, Bangladeshi participants were less likely to view suspiciousness or hallucinatory behaviour as indicative of mental health problems, and the Afro-Caribbean participants were less likely to view unusual thought content as a symptom. As expected, differences in perceptions were also associated with religion, education, gender and contact with people with mental health problems. However, ethnicity was the best predictor of perceptions of schizophrenia symptoms. CONCLUSION: Ethnicity was found to be an important factor in influencing perceptions of schizophrenia. The specific differences found across ethnic groups are useful in beginning to understand more fully public perceptions of mental health problems in Britain today. The findings raise interesting discussion in relation to ethnic and cultural factors in planning services for people from ethnic minorities, and considering in more detail issues concerning diagnosis and engagement. PMID- 12119064 TI - Psychosocial predictors of diet and acculturation in Chinese American and Chinese Canadian women. AB - OBJECTIVE: To examine the influence of diet-related psychosocial constructs on the dietary practices of Chinese populations living in North America. DESIGN: Data are from a cross-sectional survey of 244 women of Chinese ethnicity living in Seattle, WA, USA and Vancouver, BC, Canada. Using an interviewer-administered questionnaire and PRECEDE/PROCEED as our model, we collected information on diet related psychosocial (predisposing, enabling, and reinforcing) factors; consumption of foods reflecting Western and Chinese dietary practices; and past and current consumption of fruits, vegetables and fat. RESULTS: Participants generally believed that there were strong relationships between diet and health, but only about a quarter were aware of nutrition information from the government. Food cost, availability, and convenience did not appear to be major concerns among these participants. Respondents' older relatives and spouses tended to prefer a Chinese diet and also had a strong influence on the household diet. Associations of the psychosocial factors with demographic characteristics, adoption of Western dietary practices, and consumption of fruits and vegetables were informative. For example, older, less educated respondents considered it very important to eat a low fat, high fruit and vegetable diet; while younger, more educated participants who were employed outside the home did not think the Chinese diet is healthier than a typical Western diet (all p < 0.05). Western acculturated respondents were more likely to believe in a relationship between diet and cancer/heart disease and report that preparing Chinese meals is inconvenient (p < 0.05). Respondents with in-family normative pressure to maintain Chinese eating patterns ate more fruits and vegetables (4.4 vs 3.7 servings), while knowledge of nutrition information from the government was associated with increased fruit and vegetable consumption after immigration (all p < 0.05). CONCLUSIONS: Chinese cultural beliefs play an important role in the dietary practices of Chinese living in North America. Therefore, traditional health beliefs, as well as socioeconomic and environmental factors related to diet should be incorporated into the design and implementation of culturally appropriate health promotion programs for Chinese immigrants. PMID- 12119061 TI - A p130Cas tyrosine phosphorylated substrate domain decoy disrupts v-crk signaling. AB - BACKGROUND: The adaptor protein p130Cas (Cas) has been shown to be involved in different cellular processes including cell adhesion, migration and transformation. This protein has a substrate domain with up to 15 tyrosines that are potential kinase substrates, able to serve as docking sites for proteins with SH2 or PTB domains. Cas interacts with focal adhesion plaques and is phosphorylated by the tyrosine kinases FAK and Src. A number of effector molecules have been shown to interact with Cas and play a role in its function, including c-crk and v-crk, two adaptor proteins involved in intracellular signaling. Cas function is dependent on tyrosine phosphorylation of its substrate domain, suggesting that tyrosine phosphorylation of Cas in part regulates its control of adhesion and migration. To determine whether the substrate domain alone when tyrosine phosphorylated could signal, we have constructed a chimeric Cas molecule that is phosphorylated independently of upstream signals. RESULTS: We found that a tyrosine phosphorylated Cas substrate domain acts as a dominant negative mutant by blocking Cas-mediated signaling events, including JNK activation by the oncogene v-crk in transient and stable lines and v-crk transformation. This block was the result of competition for binding partners as the chimera competed for binding to endogenous c-crk and exogenously expressed v crk. CONCLUSION: Our approach suggests a novel method to study adaptor proteins that require phosphorylation, and indicates that mere tyrosine phosphorylation of the substrate domain of Cas is not sufficient for its function. PMID- 12119065 TI - Church rosters: is this a viable mechanism for effectively recruiting African Americans for a community-based survey? AB - OBJECTIVES: The purpose of this report is to describe the process, results, and implications in the phone recruitment of African Americans through church rosters for a survey of diet-and blood pressure-related awareness and hypertension prevalence. DESIGN: The survey was conducted using a non-probability sample of churches and a random selection of participants from church rosters. Recruitment strategies included frequent contact with pastors and church representatives, presentations, standard and tailored recruitment approaches, and bi-annual progress reports. Church representatives provided the rosters and assisted in arranging interviews, which were conducted at church or the participants' homes. RESULTS: Of 742 randomly selected, 315 (42.4%) were ineligible because of an unavailable or unreachable number, a move, discontinued church membership, death, or other reasons. Of the 344 eligible, 45.8% participated, 30.2% refused, 4.4% agreed to participate but did not, and 19.6% were incompletes (called less than three times before recruitment was terminated). Among participants, 70.4% were female, 58.2% had completed college, and the age range was 19-91 years. The survey's sample size goal of 196 was met. CONCLUSIONS: In this study population, over 45% who were eligible participated. Rapport established with church representatives and congregations was critical to the sampling process. Using church rosters can be a low-cost, effective recruitment tool. However, key factors to consider when recruiting African Americans in this manner include: trust, study eligibility criteria, roster accuracy, and time, and generalizability. PMID- 12119066 TI - Geographic variation in cardiovascular disease risk factors among American Indians and comparisons with the corresponding state populations. AB - OBJECTIVES: (1) To compare the prevalence of self-reported CVD, diabetes, hypertension, fair/poor perceived health status, and current tobacco use from three surveys of American Indians - two in the Southeast (Catawba Diabetes and Health Survey [CDHS] and Lumbee Diabetes and Health Survey [LDHS]) and one in the upper Midwest (Inter-Tribal Heart Project [ITHP]). (2) To compare the prevalence estimates from the CDHS, LDHS, ITHP with those for the corresponding state populations (South Carolina, North Carolina, Minnesota and Wisconsin, respectively) derived from the Behavioral Risk Factor Surveillance System (BRFSS). METHODS: Pearson's Chi-square analyses were used to detect statistically significant differences in the age-adjusted prevalence estimates across the study populations. RESULTS: Among these three populations of American Indians, the ITHP participants had the highest prevalence estimates of diabetes (20.1%) and current cigarette smoking (62.8%). The CDHS participants had the highest prevalence estimate of fair/poor perceived health status (32.0%). The LDHS participants had the highest prevalence estimate of chewing tobacco use (14.0%), and the lowest prevalence of CVD. The prevalence estimates of self-reported diabetes were dramatically higher among American Indian participants in the ITHP (20.1%) and CDHS (14.9%) than among participants in the corresponding state BRFSS (5.8% MN and WI and 6.6% SC), as were the estimates for hypertension. CONCLUSION: The substantial variations in prevalence of CVD and its risk factors among Tribal Nations suggests that distinct cultural norms, historic conditions, and important health issues of each American Indian community must be recognized and incorporated into all health promotion programs and policies. PMID- 12119068 TI - Quantitative risk assessment of durable glass fibers. AB - This article presents a quantitative risk assessment for the theoretical lifetime cancer risk from the manufacture and use of relatively durable synthetic glass fibers. More specifically, we estimate levels of exposure to respirable fibers or fiberlike structures of E-glass and C-glass that, assuming a working lifetime exposure, pose a theoretical lifetime cancer risk of not more than 1 per 100,000. For comparability with other risk assessments we define these levels as nonsignificant exposures. Nonsignificant exposure levels are estimated from (a) the Institute of Occupational Medicine (IOM) chronic rat inhalation bioassay of durable E-glass microfibers, and (b) the Research Consulting Company (RCC) chronic inhalation bioassay of durable refractory ceramic fibers (RCF). Best estimates of nonsignificant E-glass exposure exceed 0.05-0.13 fibers (or shards) per cubic centimeter (cm3) when calculated from the multistage nonthreshold model. Best estimates of nonsignificant C-glass exposure exceed 0.27-0.6 fibers/cm3. Estimates of nonsignificant exposure increase markedly for E- and C glass when non-linear models are applied and rapidly exceed 1 fiber/cm3. Controlling durable fiber exposures to an 8-h time-weighted average of 0.05 fibers/cm3 will assure that the additional theoretical lifetime risk from working lifetime exposures to these durable fibers or shards is kept below the 1 per 100,000 level. Measured airborne exposures to respirable, durable glass fibers (or shards) in glass fiber manufacturing and fabrication operations were compared with the nonsignificant exposure estimates described. Sampling results for B sized respirable E-glass fibers at facilities that manufacture or fabricate small diameter continuous-filament products, from those that manufacture respirable E glass shards from PERG (process to efficiently recycle glass), from milled fiber operations, and from respirable C-glass shards from Flakeglass operations indicate very low median exposures of 0, 0.0002, 0.007, 0.008, and 0.0025 fibers (or shards)/cm3, respectively using the NIOSH 7400 Method ("B" rules). Durable glass fiber exposures for various applications must be well characterized to ensure that they are kept below nonsignificant levels (e.g., 0.05 fibers/cm3) as defined in this risk assessment. PMID- 12119069 TI - Benzene metabolism by the isolated perfused lung. AB - Benzene is an occupational hazard and environmental toxicant whose toxic effects are dependent on its metabolism by cytochrome P-450. Most physiologically based pharmacokinetic models assume that benzene is metabolized only in the liver. They may not be completely accurate in predicting metabolism, especially following inhalation exposure, if metabolism by the lung is important. In the current study, the metabolizing capability of the lung was examined in an in vivo simulation using the isolated perfused lung. Lungs from the rabbit, rat, and mouse were used to mimic benzene metabolism following exposure via the pulmonary vasculature. With the isolated perfused mouse lung, three concentrations (55 microM, 120 microM, and 200 microM) were used to evaluate concentration dependence. To evaluate the ability of the lung to metabolize inhaled benzene, the isolated perfused mouse lung was exposed to benzene (approximately 175 ppm) via the trachea. Benzene was metabolized in all species, with phenol being the major metabolite. Phenylsulfate was also detected in perfusate from rabbits and mice but at much lower levels. Benzene metabolism was concentration dependent in mice. The ability of the lung to metabolize benzene during inhalation exposure was demonstrated in the isolated perfused mouse lung. These results demonstrate that the lung can metabolize benzene in an in vivo simulation when exposed via the pulmonary vasculature or via inhalation. PMID- 12119070 TI - Sensory nerve-mediated nasal vasodilatory response to inspired ethyl acrylate. AB - The irritants acrolein, acetaldehyde, and acetic acid induce a rapid sensory nerve-mediated nasal vasodilatory response in the rat. The aim of the current study was to examine acute nasal sensory nerve-mediated acute responses to an irritant ester vapor, ethyl acrylate. For this purpose, the upper respiratory tract of the urethane-anesthetized male F344 rat was isolated by insertion of an endotracheal cannula, and ethyl acrylate-laden air was drawn continuously through that site at a flow rate of 100 ml/min for 50 min. Vascular function was monitored by measuring inert vapor (acetone) uptake throughout the exposure. Nasal flow resistance was also monitored during exposure, and plasma protein extravasation was measured by Evans blue dye leakage. At exposure concentrations of 100 to 400 ppm, ethyl acrylate induced a rapid nasal vasodilatory response, as indicated by increased acetone uptake rates. This response was maintained throughout the exposure. Changes in nasal flow resistance or in Evans blue dye leakage were not observed at these exposure concentrations. The vasodilatory response was diminished in animals pretreated with the sensory nerve toxin capsaicin, providing strong evidence that this response was sensory nerve mediated. Pretreatment with the carboxylesterase inhibitor bis-para-nitro phenolphospahte at a dose sufficient to inhibit nasal carboxylesterase did not alter the response, suggesting that the parent ester, not the carboxylesterase metabolites, is primarily responsible for the sensory-nerve-mediated vasodilatory responses to this ester. PMID- 12119072 TI - Ozone-induced DNA single strand-breaks in guinea pig tracheobronchial epithelial cells in vivo. AB - Two-month-old male guinea pigs (Dunkin-Hartley strain, specific pathogen free), 4 in each group, were exposed to 0.00, 0.45, and 1.00 ppm O3 for 72 h. The trachea with two main bronchi was removed from each animal after O3 exposure. The trachea lavage fluid was used to measure the protein content as an index of altered tracheobronchial epithelial (TE) cell membrane permeability after O3 exposure. The TE cells were isolated and employed for the determination of DNA single strand breaks (SSBs) by fluorometric analysis of DNA unwinding (FADU). The statistical significance level was set at alpha =.05. The results show that neither the yield nor the viability of the TE cell from various O3 treatment groups was different from that of controls. Compared to controls, the protein content was elevated significantly after 0.45 ppm O3 exposure; however, the amount of DNA SSBs was not. The number of DNA SSBs increased significantly in the 1.00 ppm O3 exposure group when compared to controls. Regardless of the alkali incubation time at 15 degrees C, the double-stranded DNA left in the alkali TE cell lysate was a linear function of O3 exposure concentrations. PMID- 12119071 TI - Effects of inhaled ozone on pulmonary immune cells critical to antibacterial responses in situ. AB - The goal of this study was to examine effects from repeated exposure to ozone (O3) on immune cells involved in cell-mediated antibacterial responses in the lungs. Rats exposed to 0.1 or 0.3 ppm O3 for 4 h/day, 5 days/wk, for 1 or 3 wk were analyzed for the ability to clear an intrapulmonary challenge with Listeria monocytogenes or had their lungs processed to obtain pulmonary alveolar macrophages (PAM) and lung-associated lymphocytes for analyses of select cell functions and surface marker expression. The results indicate that repeated inhalation exposure to O3 affected local cell-mediated immunity (CMI) responses as evidenced by effects on clearance of Listeria. However, this modulation was not consistently dependent on exposure concentration or duration. Short-term repeat exposures had more effect on host resistance than did the more prolonged regimen, with rats exposed to 0.1 ppm O3 most adversely impacted. Clearance patterns suggest modifications in innate resistance following 1 wk of exposure to 0.1 ppm O3, but no similar effect following a 3-wk regimen. Exposure to 0.3 ppm O3 appeared to affect both innate and acquired resistance after a 1-wk regimen, but mainly the former after an additional 2 wk of exposure. We conclude that these two mechanisms of resistance are differentially affected by O3 and that distinct time- and O3 concentration-dependent adaptation phenomena evolve for each; that is, in situ adaptation to higher levels of O3 may occur more readily with acquired than with innate/PAM-dependent resistance. A similar pattern of inconsistent effect on PAM and lung-associated lymphocytes was also evident. For example, while 3-wk exposures had a greater effect on PAM reactive oxygen intermediate ROI production, evidence for a significant effect on antibacterial activity was only notable among PAM from rats exposed for 1 wk. Among lung lymphocytes, while 3-wk exposure to 0.1 ppm O3 led to a significant increase in CD25 expression, there was no corresponding increase in responsivity to concanavalin A (ConA); only among cells from 1-wk-exposed rats did lymphoproliferative responses increase. Though investigations of altered immune cell cytokine receptor expression/binding activity are ongoing, results herein provide further evidence to support our longstanding hypothesis that some well documented effects of O3 exposure on human health are quite likely linked to changes in local immune cell (i.e., PAM and lung-associated lymphocytes) functions, with the latter being related to changes in the capacities of these cells to interact with immunoregulatory cytokines. PMID- 12119073 TI - Diesel exhaust affects the abnormal delivery in pregnant mice and the growth of their young. AB - To clarify the toxic effects of diesel exhaust (DE) on delivery in mice and on growth of young, C57Bl-strain females were exposed to 0.3, 1.0, or 3.0 mg diesel exhaust particles (DEP)/m3 or filtered clean air (control) for 4 mo (12 h/day, 7 days/wk). After exposure, some females from each group were examined by necropsy, and the remainders were mated with unexposed males. Estrous females for necropsy who had been exposed to 1.0 mg DEP/m3 had significantly lower uterine weights than the control estrous females. In the mated females, 9.1, 10.0, or 25.0% (0.3, 1.0, or 3.0 mg DEP/m3 of the pregnancies resulted in abnormal deliveries (abortion and unable delivery), but this was not significant. The rate of good nest construction by delivered females exposed to 3.0 mg DEP/m3 was significantly lower. Young were weighed at 11, 14, and 21 days, and weekly from wk 4 to 9 after birth. Body weights of male young of dams exposed to 1.0 or 3.0 mg DEP/m3 were significantly lower at 6 and 8 wk of age. Body weights of female young of dams exposed to 1.0 or 3.0 mg DEP/m3 were also significantly lower at 6, 8, and 9 wk. Vaginal orifices of young female mice whose dams were exposed to 0.3 and 1.0 mg DEP/m3 opened significantly earlier. The young were killed at 30 or 70 days during deep anesthesia, and their body weights, organ weights, and body lengths were measured. Anogenital distance (AGD) of 30-day-old males whose dams were exposed to 0.3 mg DEP/m3 was significantly shorter than that of the controls. Weights of thymus and ovary in 30-day-old females whose dams were exposed to 3.0 mg DEP/m3 were significantly lower. In 70-day-old males of dams exposed to 3.0 mg DEP/m3, body weights were significantly lower and AGD was significantly shorter. Weights of adrenals, testes, and seminal vesicles in 70-day-old males with dams exposed to 1.0 mg DEP/m3 were significantly lower. In 70-day-old females with dams exposed to DE, body weights in the 3.0-mg DEP/m3 group were significantly lower, and weights of adrenals, liver, and thymus in the 1.0-mg DEP/m3 group were significantly lower. Thymus weights in 70-day-old females with dams exposed to 0.3 mg DEP/m3 were significantly lower. Crown-rump length (CR) in 70-day-old females with dams exposed to 1.0 or 3.0 mg DEP/m3 was significantly shorter. These results show that toxic substances in DE might cause abnormal delivery in mice, and that exposed females affected the growth and sexual maturation of their young. PMID- 12119074 TI - Comparison of inflammatory elements in nasal lavage and induced sputum following occupational exposure to moldy-building microbes. AB - Inflammatory processes in the nasal air passages may reflect corresponding processes in the lower airways due to the similarities in histology of nasal mucosa and bronchi. The objective of the current study was to determine whether the levels of inflammatory markers in nasal lavage fluid could be used as predictors of lower respiratory tract inflammation after exposure to microbes in indoor air of moisture-damaged buildings. Differential cell count, immunochemically measured concentrations of proinflammatory cytokines (Interleukins [IL] IL-1, IL-4, IL-6, and tumor necrosis factor alpha [TNFalpha]) and nitric oxide (NO), assessed as nitrite, were analyzed from nasal lavage (NL) and induced sputum (IS) samples of the occupants (n = 60) working in moisture damaged and reference school buildings. The measurements of inflammation markers in NL and IS sample pairs, collected on the same day, were compared. Although the levels of NO (p =.026) and IL-4 (p =.014) in NL predicted their levels in IS in a statistically significant manner, their predictive values (6.9% and 7.8%, respectively) were low. There was no significant correlation between the concentrations of the studied proinflammatory cytokines or differential cell counts in NL and IS samples. Our results indicate that measurement of inflammatory mediators in NL is not per se a reliable method to evaluate the inflammatory status of the lower airways after exposure to indoor air pollutants of moisture damaged building. It is possible that NL is a more sensitive indicator of direct exposure to different irritants in inhaled air than is IS. This may be a reflection of the role of nasal mucosa as the primary physicochemical barrier to inhaled air. PMID- 12119075 TI - The locked-in syndrome: a syndrome looking for a therapy. AB - A review of the scientific literature on locked-in syndrome (LIS) is offered. The clinical features, diagnosis and prognosis of LIS are reviewed, and methods regarding the differential diagnosis of LIS with severe disorders of consciousness are considered. Effective treatment, physiotherapy, and methods of communication are reviewed. Although progress in the field of communication for patients with LIS is promising, it is concluded that there are new possibilities to be pursued and that a more positive outlook in the area of professional care of the patients, as well as more extensive imaginative research will facilitate new and positive strategies for this syndrome. PMID- 12119076 TI - The locked-in syndrome: a syndrome looking for a therapy. AB - The locked-in syndrome (LIS) is a very severe condition caused by a primary vascular or traumatic injury to the brainstem, normally corresponding to a ventral pons lesion due to an obstruction of the basilar artery, and characterized by upper motor neuron quadriplegia, paralysis of lower cranial nerves, bilateral paresis of horizontal gaze and anarthria, and with preserved consciousness. Patients who have suffered this pontine lesion generally have preserved vertical eye movements and movement of the eyelids (blinking), this being their only means of responding to the outside world. A survey was conducted of 44 people diagnosed with LIS, all of them belonging to the Association of Locked-in Syndrome (ALIS) of France. Results of this survey showed that LIS was equally frequent in men and women (51.2% vs. 48.1%) and had occurred at any age between 22-77 years of age (normally between 41-52 years, the mean age being 46.79 years). The average time that transpired post-insult was 71.35 months. The principal cause of LIS was stroke (86.4%), with traumatic brain injury (TBI) being a distant second cause with an incidence of only 13.6%. The diagnosis of LIS was usually made around the middle of the second month after onset (mean of 78.76 days). The principal treatments, when present, were pharmacological and physiotherapy. However, 47.1% of the patients were not receiving treatment of any kind at the time of the survey. Neuropsychologically, 86% had a good attentional level, 97.6% were temporally oriented and 76.7% could read; 18.6% reported memory problems and 24% showed visual deficit (found mainly in patients with LIS originated by TBI); 47.5% reported a good mood state and 12.5% reported feeling depressed; 61.1% reported having sexual desire, but only 30% maintained sexual relations; 78% were capable of emitting sounds and 65.8% could communicate without technical aid; 73.2% enjoyed going out and 81% met with friends at least twice a month. Only 14.3% participated in social activities and 23.8% watched television regularly. Nearly 100% of the patients reported being sensitive to touch to any part of their bodies. This survey suggests diagnostics and rehabilitation procedures. PMID- 12119077 TI - Post-injury substance abuse among persons with brain injury and persons with spinal cord injury. AB - PRIMARY OBJECTIVE: To identify the patient population at greatest risk for post injury adjustment problems, the present study independently examines and compares alcohol and drug use rates in patients with traumatic brain injury (TBI) and patients with spinal cord injury. RESEARCH DESIGN: The two samples were matched with regard to age, gender and mechanism of injury. The study provides a description of post-injury use rates for each population, and describes similarities and differences between the two groups. METHODS AND PROCEDURES: Participants included 30 consecutive Model Systems spinal cord injury (SCI) patients seen for follow-up neuropsychological testing between October 1996-June 1999. An equivalent number of Model Systems TBI patients were matched from a larger sample comprised of 440 consecutive hospital admissions, that returned for a 1-year follow-up neuropsychological evaluation between February 1989-December 1998. All participants were treated in an urban Level I trauma centre and associated inpatient rehabilitation programmes. Information regarding patient demographics, as well as pre- and post-injury psychiatric, employment, academic, criminal, and medical history was obtained via the General Health and History Questionnaire. MAIN OUTCOMES AND RESULTS: With regard to post-injury alcohol use rates, persons with spinal cord injury were more likely to drink on a daily basis. Although not statistically significant, pre-injury drinking rates differed from post-injury rates for both groups. With regard to illicit drug use, persons with TBI differed significantly from persons with SCI. A significant difference was also noted between pre-injury drug use and post-injury drug use for both groups. CONCLUSIONS: Persons who drink post-injury are unlikely to be 'light' or social drinkers. Either people choose to abstain completely or appear to use alcohol frequently. PMID- 12119078 TI - Loewenstein communication scale for the minimally responsive patient. AB - PRIMARY OBJECTIVE: Any sign of communicative ability in patients in vegetative state can provide information about regain of consciousness and conservation of cognitive abilities. The aim of this study was to test the reliability and validity of an instrument designed to measure the degree of communication in minimally responsive patients. MATERIALS: The Loewenstein Communication Scale (LCS) measures five hierarchical functions - mobility, respiration, visual responsiveness, auditory comprehension and linguistic skills (verbal or alternative) - which are divided into five parameters and rated in developmental order on a 5-point scale by level of difficulty. Scores for each function are summed to obtain a quantitative communication profile. METHODS: Forty-two adult patients in vegetative state, as a result of acquired brain injury, were examined with the proposed LCS for the minimally responsive patients by two speech and language clinicians at admission to the Intensive Care Unit (ICU) for brain injured patients and, thereafter, at least once weekly. At the end of the ICU stay, 27 patients who showed signs of recovery and were referred for continued rehabilitation were compared to a group of 15 patients who were not referred for continued rehabilitation, for functional and general LCS scores. The predictive power of the LCS in differentiating between these groups was tested. RESULTS: The LCS was found to have very good reliability with good inter-rater agreement. Patients who eventually continued rehabilitation had significantly higher total scores as well as in the motor, visual and auditory sub-scales. Logistic regression results indicated that these parameters successfully differentiated between the two groups of patients, even after adjusting for age and for scores on the Glasgow Coma Scale. CONCLUSION: The LCS for the minimally responsive patients proved to be reliable and predictive of rehabilitation progress of minimally responsive patients. It may be useful for the interdisciplinary rehabilitation team in planning early individually targeted therapeutic programmes. PMID- 12119079 TI - Functional outcome: a case for mild traumatic brain injury. AB - The purpose of this case study was to demonstrate the successful incorporation of the strengths of process-specific and functional treatment approaches within a therapy programme for a patient with a MTBI. A patient with MTBI and deficits in attention, executive functioning, memory, reasoning and problem solving participated in a 4-month treatment programme. The interactions between the patient's cognitive deficits and problematic activities of daily living were identified. Treatment focused on teaching the patient compensatory strategies to offset the cognitive deficits following a cognitive framework within the context of hierarchically arranged activities of daily living. At the end of this programme, the patient consistently used his compensatory strategies to independently complete activities of daily living that were problematic prior to the receipt of treatment. Implications of this study for the treatment of patients with MTBI suggest that a combined approach is most beneficial in maximizing the recovery of these patients. PMID- 12119080 TI - Scedosporium apiospermum post-traumatic cranial infection. AB - Scedosporium apiospermum is an environmental mould. Human infection caused by this organism is described more and more often. However, only a few case reports demonstrate its role as a telluric contaminant in patients affected by traumatism. This report is the case of a severe post traumatic infection by S. apiospermum in an immunocompetent young man. Surgical drainage associated with systemic therapy was successful. PMID- 12119081 TI - Organic alexithymia: a study of acquired emotional blindness. AB - The phrase 'organic alexithymia' is introduced as a clinically and theoretically useful construct for furthering understanding of alexithymia and the occurrence of alexithymic symptoms in patients with acquired brain injury (ABI). The construct is illustrated by the case study of HR, a 21-year-old man seen at the State Head Injury Unit, Perth, for neuropsychological and clinical assessment 2 years following a motor vehicle accident. HR's case supports the hypotheses that a syndrome substantially similar to alexithymia can occur in patients with acquired brain injury, and that the acquired nature of the organic disorder may cause it to be systematically different from the established syndrome. The case demonstrates the clinical relevance of a construct like organic alexithymia when treating patients with ABI. It also highlights the need to develop instruments capable of identifying the condition and differentiating it from symptoms of depression. PMID- 12119082 TI - Intractable epilepsy and mild brain injury: incidence, pathology and surgical outcome. AB - OBJECTIVE: To study the role of mild brain injury in intractable epilepsy. METHODS: The medical charts of 86 patients who underwent surgery for intractable epilepsy were reviewed in regard to the history of mild brain injury, pathology and surgical outcome. RESULTS: Nine of 86 patients had a previous history of mild brain injury (10.4%) compared to 2.5% of 80 age and sex matched controls. Six of nine patients had non-neoplastic and three had neoplastic lesions. Post-surgical outcome was excellent in eight of nine patients (Engel class IA). CONCLUSIONS: The incidence of mild brain injury was 4-times higher in patients with intractable epilepsy compared to asymptomatic controls. The pathology was variable but in four of nine patients it was compatible with the described pathology in traumatic brain injury. Both groups, with or without brain injury, had good surgical outcome (88% versus 70%). PMID- 12119083 TI - Aggressive behaviour observed within a neurobehavioural rehabilitation service: utility of the OAS-MNR in clinical audit and applied research. AB - The Overt Aggression Scale-Modified for Neurorehabilitation (OAS-MNR) has been proposed as a means of standardizing descriptions of post-acute aggressive behaviour disorders amongst people with acquired brain injury. Single cases in the literature have illustrated its clinical utility. In this paper, its contribution to clinical audit and applied research is explored. The scale was used to record all aggressive behaviour exhibited by 46 patients participating in a neurobehavioural programme during a 14 day period. Data for 3914 acts of aggression were captured. Whilst most was verbal, 443 comprised physical assaults on others. Results suggest the OAS-MNR can be usefully employed to audit practice, and has a role to play in resource allocation: however, efficacy remains best judged at the single case level. Regarding clinical research, it was found that patients with low language function were more likely to physically assault others in the absence of identifiable antecedents. Furthermore, this was more severe and required more intrusive interventions to manage it than aggression shown by the same patients which had identifiable antecedents, or any aggression exhibited by patients whose language was better preserved. It is argued that poor language function creates significant barriers to the treatment of aggression, and, whilst intervention methods with good outcome have previously been described, these are no longer routinely available in the UK. An appeal to reverse this is made. PMID- 12119084 TI - Rehabilitation outcome after primary subarachnoid haemorrhage. AB - Functional outcome of primary subarachnoid haemorrhage survivors was examined following rehabilitation in Singapore. Thirty-nine inpatients admitted over a 4 year period were studied retrospectively. There were 21 (53.8%) males and 18 (46.2%) females, mean age 50.9 (SD 12.7) years, at 37.3 days (SD 16.8) post injury. Seven (17.9%) had dysphagia and 12 (30.8%) had dysphasia; 17 (43.6%) nondysphasics had cognitive impairment; 26 patients (66.7%) had Medical Research Council motor power under grade 4. Modified Barthel Index on Admission (MBIA) and Discharge (MBID) were 45.7 (SD 22.3) and 78.3 (SD 18.9), respectively (p = 0.001). Correlation between MBIA and MBID was statistically significant (r = 0.529, p = 0.001). Mean length of stay in rehabilitation was 42.3 days (SD 27.6). Thirty-four patients (87.2%) were discharged home. Nine out of 24 previously employed patients (37.5%) returned to gainful employment. Despite multiple deficits, the patients made significant functional improvement. PMID- 12119085 TI - Conceptualization and identification of depression in adults with brain damage by clients and rehabilitation clinical staff. AB - PRIMARY OBJECTIVE: To explore reasons used by staff and clients to identify depression and examine how each group differs. RESEARCH DESIGN: A between group comparison on both quantitative and qualitative data. METHODS AND PROCEDURE: Thirty-seven individuals with brain injury and 18 non-psychologist rehabilitation professionals participated in this study. The Inventory for Depressive Symptomatology (IDS), a 28-item instrument of client self-report and clinician ratings was used for comparison. Additionally, qualitative data on presence and reason for identification of depression were collected from both parties. RESULTS: There is substantial disagreement between clinicians and clients with regard to overall identification of depression. However, both groups based diagnoses on mood/cognitive symptoms rather than anxiety or vegetative symptoms. CONCLUSION: The results point to a discrepancy in client and clinician identification and understanding of depression after brain injury, indicating a potential need for education for both groups. PMID- 12119086 TI - Cutoff score on the apathy evaluation scale in subjects with traumatic brain injury. AB - This cross-sectional study was designed to determine a cutoff score on the Apathy Evaluation Scale (AES) that predicts a clinician's designation of a subject with TBI as apathetic or not. Forty-five outpatients with TBI completed the AES-S, and 37 family members, friends, or significant others filled out the AES-I. Three clinicians prospectively gave their impressions of the presence or absence of apathy and retrospectively chose the degree of apathy on a 7-point subjective rating scale. The data was analysed by logistic regression and Receiver Operating Characteristic (ROC) curve. Sensitivity and specificity were calculated. No cutoff score on the AES-S or AES-I was found to have reasonable sensitivity and specificity with respect to the ability to predict the clinician's designation of a subject as apathetic. The AES requires further study if it is to be used to measure apathy following TBI. PMID- 12119087 TI - Quantitive imaging in late traumatic brain injury. Part I: late imaging parameters in closed and penetrating head injuries. AB - OBJECTIVE: To ascertain the value of relatively simple quantitative radiologic measurements after head injury. Despite major advances in neuroradiology, analysis and reporting of imaging studies is based primarily on individual subjective and local experience, rather than on reproducible, standardized parameters; reliance on newer technologies can improve care, but also raises diagnostic costs. DESIGN: Blinded, retrospective, quantitative assessment of computerized tomography studies done some 14 years post-injury. OUTCOME MEASURES: Frontal horn width (FHW); septum-caudate distance (SCD); temporal horn width (THW); interuncal distance (IUD); third ventricle width (3VW); ventricular score (VS); sulcal width (SW); gray-white matter discriminability (GWMD) and subjective assessment of atrophy (SAOA). RESULTS: Diffuse and frontal damage was noted in both closed (CHI) and penetrating (PHI) head injury groups. Enlargement of frontal lobe parameters (septum caudate distance and frontal horn width suggest frontal injury in both closed and penetrating traumatic brain injury (TBI). Temporal horn width and inter-uncal distance were related to VS, 3VW and FHW in closed, but not in penetrating head injury. CONCLUSIONS: Simple linear CSF space measurements are correlated with volumetric and parenchymal measures, and can represent valuable and reliable low-cost quantitative measures of long term brain damage after TBI. PMID- 12119088 TI - Effects of acquired brain injury on adaptive choice and the role of reduced sensitivity to contingencies. AB - The present investigation examined the proposal that brain injury reduces sensitivity to consequences. On a laboratory task, controls and subjects with brain injury made repeated choices between a small amount of money and a larger amount of money followed by a post-reinforcer delay of 3, 12, 18, or 24 seconds. Increasing delays lowered reinforcement densities (i.e. money available per minute) associated with large reinforcers. Consequently, choosing large reinforcers became less adaptive. Results showed subjects with brain injury made less adaptive choices and earned significantly less money than controls, because of a preference for large reinforcers with low reinforcement densities. Maladaptive choice was not attributable to deficits in discriminating reinforcer magnitudes or delays. Results suggest individuals with brain injury may remain sensitive to consequences but fail to discriminate among significant response consequence relations (i.e. contingencies). This view, emphasizing basic learning processes, may account for some problems in skill acquisition and adaptive choice. PMID- 12119089 TI - Diagnostic codes in dentistry--definition, utility and developments to date. AB - Diagnostic codes are computer-readable descriptors of patients' conditions contained in computerized patient records. The codes uniquely identify the diagnoses or conditions identified at initial or follow-up examinations that are otherwise written in English or French on the patient chart. Dental diagnostic codes would allow dentists to access information on the types and range of conditions they encounter in their practices, enhance patient communication, track clinical outcomes and monitor best practices. For the profession, system wide use of the codes could provide information helpful in understanding the oral health of Canadians, demonstrate improvements in oral health, track best practices system-wide, and identify and monitor the progress of high-need groups in Canada. Different systems of diagnostic codes have been implemented by program managers in Germany, the United Kingdom and North America. In Toronto, the former North York Community Dental Services developed and implemented a system that follows the logic used by the Canadian Dental Association for its procedure codes. The American Dental Association is now preparing for the release of SNODENT codes. The addition of diagnostic codes to the service codes already contained in computerized patient records could allow easier analysis of the rich evidence available on the oral health and oral health care of Canadians, thereby enhancing our ability to continuously improve patient care. PMID- 12119090 TI - Investigating the potential for students to provide dental services in community settings. AB - Some dental educational institutions in North America have incorporated community oriented programs into their curriculum. The purpose of this study was to investigate the potential for the clinical placement of Ontario's dental and dental hygiene students in community-based settings. Key informant interviews were used to collect data. The study group consisted of 15 key informants from 9 potential placement sites and 4 educational institutions in Toronto and London, Ontario. The textual data were analyzed qualitatively to identify important issues regarding a clinical placement program. Results showed that there is strong support for the placement of students in community-based clinics; however, the degree to which health centres can accommodate students varies. The majority would not set any limit on the types of dental services that students could provide as long as the services were within the students' competencies. Funding was identified as a barrier to the implementation of such a program, with most of the organizations not able to contribute financially. None would be able to provide sufficient supervision without additional funding. These results indicate that a clinical placement program would be a welcome addition to the training of dental and dental hygiene students, but that external funding for supervision and operational expenses must be available before a program can be instituted. PMID- 12119092 TI - Oral and pharyngeal cancer: knowledge and opinions of dentists in British Columbia and Nova Scotia. AB - Oral and pharyngeal cancers are largely preventable and can be successfully treated when diagnosed at an early stage. Dentists in British Columbia and Nova Scotia were surveyed regarding their knowledge and opinions about oral and pharyngeal cancer. In February 1998 a pretested 41-item survey was mailed to a random sample of dentists in British Columbia and the population of dentists in Nova Scotia. A reminder postcard and one additional mailing were sent to nonrespondents. Of the 670 dentists supplying usable responses (response rate 55.2%) only 56.7% agreed that their knowledge of the subject was current. Most dentists correctly identified tobacco use (99.4%) and alcohol use (90.4%) as risk factors, but fewer correctly identified factors such as the use of spicy foods (57.0%) and poor oral hygiene (46.3%) as not being risk factors. Only 42.5% identified both erythroplakia and leukoplakia, in that order, as the conditions most likely associated with oral cancer. Indices of risk and diagnostic knowledge were constructed by summing the number of correct responses to items in each domain. On 16 risk factors the mean correct score was 9.2, and on 14 diagnostic procedures the mean correct score was 10.0. Only 38.5% of dentists had consistent levels of knowledge on both indices. Differences between the provinces were statistically significant (p < 0.01) for only 2 knowledge items. About three quarters of all dentists (77.0%) were interested in taking continuing education courses. Dentists in British Columbia and Nova Scotia could benefit from undergraduate and continuing education courses to increase their knowledge of risk and diagnostic factors for oral cancer. PMID- 12119091 TI - The effect of dental insurance on the ranking of dental treatment needs in older residents of Durham Region's homes for the aged. AB - The effect of dental insurance on the ranking of dental needs in older adults has not been reported previously. We examined this effect using data obtained from a cross-sectional survey of older adults living in homes for the aged in Durham Region, Ontario. History of dental insurance was obtained during interviews. Dental needs, assessed during clinical examinations, were ranked from no need to urgent need according to the guideline of the American Dental Association. The associations between the rank of dental needs, dental insurance and other factors were analyzed with the Kruskal Wallis test, chi-square test, analysis of variance and multiple logistic regression. Of the 252 participants, 80 (31.7%) had been insured continuously since 1974, 69 (27.4%) had no need for dental treatment and 59 (23.4%) needed urgent dental care. More of the continuously insured than the uninsured residents were dentate (46/80 [57.5%] vs. 75/172 [43.6%], p = 0.04). Ranking of the need for care was not significantly influenced by dental insurance; need of any kind was explained by being dentate (odds ratio 12.3, 95% confidence interval 5.6 27.3). PMID- 12119094 TI - How to pattern an epithelium: lessons from achaete-scute regulation on the notum of Drosophila. AB - The notum of Drosophila is a good model system for the study of two-dimensional pattern formation. Attention has mainly focused on the regulation of the spatial expression of the genes of the achaete-scute complex (AS-C) that results in a stereotyped bristle pattern. Expression of AS-C genes has traditionally been viewed as a consequence of the activity of a group of factors that constitute a prepattern [Stern, 1954. Am. Sci. 42, 213]. The prepattern is thought to be composed of a mosaic of transcription factors that act in combination, through discrete cis-regulatory sequences, to activate expression of genes of the AS-C in small clusters of cells at the sites of each future bristle. Recent results challenge this view and suggest a hierarchy of activity amongst prepattern genes. It is suggested that in the medial notum, the selector-like gene pannier regulates the entire pattern, and is the only factor to directly activate AS-C genes. Other factors may play subsidiary roles. On the lateral notum genes of the iroquois complex appear to regulate the lateral pattern. Regulation of pannier and iroquois depends upon the signalling molecule Decapentaplegic. The majority of genes are expressed in either longitudinal or transverse domains on the notum and we discuss the possibility that pattern formation may rely on these two axial coordinates. We also discuss preliminary results suggesting that prepattern factors also regulate genes required for other, little studied, aspects of notal morphology, such as the muscle attachment sites and pigment distribution. Thus there may be a common prepattern for the entire structure. PMID- 12119093 TI - Oral and pharyngeal cancer: practices and opinions of dentists in British Columbia and Nova Scotia. AB - Oral and pharyngeal cancers are associated with high mortality rates, a situation usually attributed to late-stage diagnosis. Dentists in British Columbia and Nova Scotia were surveyed regarding their practices and opinions related to oral and pharyngeal cancer. In February 1998 a pretested, 41-item survey was mailed to a random sample of dentists in British Columbia (n = 817) and the population of dentists in Nova Scotia (N = 423). A reminder postcard and one additional mailing were sent to nonrespondents. Of the 670 dentists supplying usable responses (response rate 55.2%), only 56.7% agreed that their knowledge of the subject was current. Of 8 health history items, dentists assessed 5 on average, with most (88.0%) asking about the patients' current use of tobacco. A total of 72.7% of the responding dentists performed an oral cancer examination for all edentulous patients at every appointment, but 10.9% never did so. Similarly, 70.7% of the dentists always provided an oral cancer examination at the initial appointment for patients 40 years of age and older, but 9.8% never did so. Undergraduate training related to oral cancer examination was reported as good by only 52.2% of the dentists. About three-quarters of all dentists (77.0%) were interested in taking continuing education courses on this subject. Differences between the 2 provinces were not statistically significant (p > 0.01). Dentists in British Columbia and Nova Scotia could benefit from undergraduate and continuing education courses to increase their knowledge of health history assessment, examination for oral and pharyngeal cancers, and risk reduction strategies, such as counselling about tobacco cessation. PMID- 12119095 TI - Topography of genetic elements of X-chromosome relative to the cell nucleus and to the chromosome X territory determined for human lymphocytes. AB - Topography of three genetic elements--dystrophin (dmd) exons 5-7 (E(1)), 46-47 (E(2)), and centromere of chromosome X (N(X)) were studied relative to cell nuclei and to chromosome X territories of spatially fixed human lymphocytes. Repeated three-dimensional (3D) dual color fluorescence in situ hybridization combined with high-resolution cytometry was used. In addition, the nuclear location of fluorescence weight centers (FWC), spatial volume, and maximal area per one section of chromosome-X territories were investigated. The larger (X(L)) and smaller (X(S)) homologous X-chromosomes were distinguished for each nucleus according to the 3D volume of their territories. The distributions of the [center of nucleus]-to-[genetic element] distances (radial distributions) of dmd exons E(1), E(2), centromere N(X) and FWC were very similar for both homologous X chromosomes of female lymphocytes as well as for the chromosome X of the human male. On the other hand, larger average mutual distances between all pairs of signals (E(1), E(2), N(X), FWC) and larger average maximal area were observed for the larger chromosome (X(L)) in comparison with the smaller one (X(S)). The territory of the larger homologue showed also more irregular surface. The most significant differences between homologous X-chromosomes were found for N(X) E(1), N(X)-E(2) and E(1)-E(2) distances that were in average about twice longer for X(L) as compared with X(S). These parameters correlate to each other and can be used for the reliable determination of more (de)condensed X-chromosome territory. The longer E(1)-E(2) distances for X(L) indicate more open chromatin structure of the dystrophin gene on this chromosome in contrary to closed structure on X(S). Substantially shorter distances of the dystrophin exons from the centromeric heterochromatin in X(S) as compared to X(L) can be explained by silencing effect of centromeres as described in Nature 1 (2000) 137. PMID- 12119096 TI - The human gamma-aminobutyric acid A receptor delta (GABRD) gene: molecular characterisation and tissue-specific expression. AB - Terminal deletions of 1p36 result in a specific and common syndrome characterised by the following: growth delay, distinctive facial anomalies, hearing and visual deficits, heart defects, body asymmetry, moderate to severe psychomotor retardation, epilepsy, and self-abusive behaviour. The human gamma-aminobutyric acid A receptor delta-subunit gene (GABRD) encodes for one of at least 15 ligand gated chloride channels for gamma-aminobutyric acid (GABA), the major inhibitory neurotransmitter in the mammalian brain. Recently we have mapped this gene by radiation hybrid mapping to the critical region of gene loss of the 1p36 deletion syndrome within 1p36.33. The complete complementary DNA (cDNA) sequence of GABRD was generated using assembled sequence of cDNA fragments already available, and 5'-rapid amplification of cDNA ends products. Fine physical mapping of the GABRD gene within this genomic interval was performed by screening bacterial artificial chromosome contigs spanning the critical region of the 1p36 deletion syndrome. The GABRD gene maps immediately proximal to the PRKCZ gene that is located between marker D1S243 and cosmid D1Z2--a region thought to be critical for cognition and speech development. The GABRD gene is expressed most abundantly in brain and has three alternative exons (1A-C) with alternative start codons at the 5'-end. Genomic localisation, function, and expression would suggest that the GABRD gene represents a good candidate for the neurodevelopmental and neuropsychiatric anomalies seen in the 1p36 deletion syndrome. PMID- 12119097 TI - Characterisation of set-1, a conserved PR/SET domain gene in Caenorhabditis elegans. AB - The SET domain is a highly conserved domain shared between proteins of the antagonistic trithorax and Polycomb groups. It has been shown to play an important role in the assembly of either transcriptional activating or repressing protein complexes, and possesses a histone methyl-transferase activity. We report here the characterisation of the Caenorhabditis elegans gene, set-1, encoding a conserved SET-domain protein. We have analysed the developmental expression pattern of set-1 and show that maximal expression is observed early in development when set-1 is ubiquitously expressed. Its expression is more and more restricted as development progress. Gene inactivation by RNA interference shows that set-1 is an essential gene. Functional analysis of set-1 may contribute to the understanding of the molecular role of the SET domain. PMID- 12119098 TI - The odorant-binding proteins of Drosophila melanogaster: annotation and characterization of a divergent gene family. AB - Insect odorant-binding proteins (OBPs) are thought to facilitate the delivery of hydrophobic odorants, such as sex pheromones or food odors, to receptors on sensory neurons. Increasingly, OBP family members are also being found in non sensory tissues where they might carry other types of small hydrophobic molecules. They are identifiable by four or six conserved Cys residues and contain six alpha-helices which enclose a hydrophobic ligand-binding pocket. Through exhaustive BLAST searches we have increased the total number of OBPs identified in Drosophila melanogaster to 38, and have amplified the DNA complementary to RNA corresponding to 21 of these by reverse transcriptase polymerase chain reaction. Isoforms frequently share less than 30% amino acid identity and appear to have radically changed since the separation of the major insect orders. However, their sequences are consistent with known OBP structures. Most are located in clusters of between four and 14 genes and several were unusual in that they contained additions, deletions, or fusions. These hexa helical insect OBPs are structurally unrelated to the functionally analogous lipocalin-like beta-barrel OBPs of vertebrates. As only two lipocalin-like proteins have been found in D. melanogaster, these helical proteins appear to be the dominant carrier of small hydrophobic molecules in insects. PMID- 12119099 TI - Expression of long- and short-type FK506 binding proteins in hyperthermophilic archaea. AB - It has been reported that the hyperthermophilic archaeon, Methanococcus jannaschii, possesses two FKBP (FK506 binding protein) genes in the genome, one being 26 kDa FKBP (long-type FKBP) and the other, 18 kDa FKBP (short-type FKBP). FKBP is a family of peptidyl-prolyl cis-trans isomerases (PPIases). In order to clarify the difference between their roles in archaeal cells, they were expressed in Escherichia coli, and their PPIase and chaperone-like protein-folding activities were investigated. The catalytic efficiency of the PPIase activity of the long-type FKBP was significantly lower than that of short-type FKBP (less than 1/1000) which is comparable to that of human FKBP12. Both FKBPs showed chaperone-like protein-folding activity to enhance the refolding yield of an unfolded protein (Thermoplasma citrate synthase) in vitro. The chaperone-like protein-folding activity of the short type was higher than that of the long type. While the intracellular content of long-type FKBP in M. jannaschii tended to increase, that of short-type FKBP obviously decreased at growth temperatures higher than the optimum of 85 degrees C. In Pyrococcus horikoshii, another hyperthermophilic archaeon, the intracellular content of long-type FKBP did not change with temperature (80-102 degrees C). These results suggest that long-type FKBP functions at any temperature in the cells as a chaperone to maintain the folding states of intracellular proteins. On the other hand, short-type FKBP may be required at lower temperatures. Peptidyl-prolyl cis-trans isomerization is known to be a rate-limiting step in protein-folding and is slower at low temperature. Since the PPIase activity of short-type FKBP was much stronger than that of the long type, it may be required to accelerate the folding of intracellular proteins and for the hyperthermophilic cell to live at low growth temperatures. PMID- 12119100 TI - Molecular characterization of the equine AEG1 locus. AB - Acidic epididymal glycoprotein 1 (AEG1), also called cysteine-rich secretory protein 1 (CRISP1), is a member of the CRISP protein family which is characterized by 16 conserved cysteine residues at the C-terminus. The CRISP proteins are expressed in the male genital tract and are thought to be involved in sperm-egg fusion. Therefore, their genes are of interest as candidate genes for inherited male fertility dysfunctions and as putative quantitative trait loci for male fertility traits. In this report, the cloning and DNA sequence of 90 kb of horse genomic DNA from equine chromosome 20q22 containing the complete equine AEG1 gene are described. The equine AEG1 gene consists of eight exons spanning 31 kb. Analysis of equine AEG1 transcripts did not reveal any evidence for alternative splicing, however three different transcription start sites are used. The first transcription start site is located 20 nt downstream of a TATA box motif. Reverse transcription polymerase chain reaction analysis demonstrated that AEG1 is expressed in different parts of the epididymis, whereas it is hardly detectable in the testis. The naturally occurring diversity of the equine AEG1 gene in different horse breeds was investigated and several polymorphisms are reported, including one that affects the amino acid sequence. Finally, sequence comparisons revealed that the intronless equine PGK2 gene for the testis-specific phosphoglycerate kinase is located approximately 39 kb downstream of AEG1. PMID- 12119101 TI - Genomic analysis of the murine odorant receptor MOR28 cluster: a possible role of gene conversion in maintaining the olfactory map. AB - Genomic analysis was performed for the murine odorant receptor (OR) genes. The MOR28 cluster on chromosome 14 was extensively studied. It contains six OR genes, MOR28, 10, 83, 29A, 29B and 30. The human homolog of this cluster is located on the human chromosome 14, and contains five OR genes, HOR28/10, 83, 29A, 29B and 30. Sequence comparison of these OR gene paralogs and orthologs suggests that the coding homologies are accounted for not only by recent gene duplication, but also by gene conversion among the coding sequences within the cluster. A possible role of gene conversion in the olfactory system is discussed in the context of the olfactory map. PMID- 12119102 TI - Molecular cloning, gene structure, expression profile and functional characterization of the mouse glutamate transporter (EAAT3) interacting protein GTRAP3-18. AB - Glutamate is an important amino acid implicated in energy metabolism, protein biosynthesis and neurotransmission. The Na(+)-dependent high-affinity excitatory amino acid transporter EAAT3 (EAAC1) facilitates glutamate uptake into most cells. Recently, a novel rat EAAT3-interacting protein called GTRAP3-18 has been identified by a yeast two-hybrid screening. GTRAP3-18 functions as a negative modulator of EAAT3-mediated glutamate transport. In order to further understand the function and regulation of GTRAP3-18, we cloned the mouse orthologue to GTRAP3-18 and determined its gene structure and its expression pattern. GTRAP3-18 encodes a 188-residue hydrophobic protein whose sequence is highly conserved amongst vertebrates. Mouse and human GTRAP3-18 genes contain three exons separated by two introns. The GTRAP3-18 gene is found on mouse chromosome 6D3 and on human chromosome 3p14, a susceptibility locus for cancer and epilepsy. GTRAP3 18 protein and RNA were found both in neuronal rich regions of the brain and in non-neuronal tissues such as the kidney, heart and skeletal muscle. Mouse GTRAP3 18 inhibited EAAT3-mediated glutamate transport in a dose-dependent manner. These studies show that GTRAP3-18 is a ubiquitously expressed protein that functions as a negative regulator of EAAT3 function. PMID- 12119103 TI - A computerized database-scan to identify c-MYC targets. AB - The c-MYC oncogene plays a pivotal role in the malignant transformation of various types of human cancer. It is also a key regulator of cellular proliferation, embryonic differentiation and apoptosis. c-MYC encodes a transcription factor that activates target genes in a sequence specific manner through heterodimerization with the ubiquitously expressed factor MAX. Identifying c-MYC target genes is therefore crucial for elucidating the molecular pathways that are downstream of MYC. Most of the c-MYC targets isolated to date as well as targets of other transcription factors have been identified by differential expression or the candidate gene approach. In this paper, we outline a computer-based scan that allows us to create a pool of putative target genes for a transcription factor. The scan is based on a set of criteria including sequence specificity of the c-MYC transcription factor, sequence location and evolutionary conservation of these regulatory elements. Using this procedure, we have identified 12 putative targets for c-MYC. Expression analyses, DNA binding assays and chimeric promoter-reporter experiments suggest that two genes, NM23-H2 and N-RAS, may indeed be direct targets for c-MYC activation. This type of computer-based scan may have a general use to identify targets for other transcription factors. PMID- 12119105 TI - A novel porcine gene, alpha-1-antichymotrypsin 2 (SERPINA3-2): sequence, genomic organization, polymorphism and mapping. AB - A novel porcine gene, alpha-1-antichymotrypsin 2 (SERPINA3-2), a member of the serpin superfamily, was isolated from a porcine genomic library and sequenced. The genomic organization of the approximately 9.0 kb gene was determined on the basis of the porcine liver cDNA of SERPINA3-1 and SERPINA3-2, and comprises five exons and four introns. The coding sequence of SERPINA3-2 shares 86% identity with the paralogue, SERPINA3-1. Porcine SERPINA3-2 was found to be an orthologue of human SERPINA3 (71% identity of the coding sequences) and both genes have a similar genomic organization. Polymorphisms were found in intron 4 of the porcine gene using polymerase chain reaction-restriction fragment length polymorphism. The gene was mapped by linkage analysis and radiation hybrid mapping to the distal end of chromosome 7q, to the gene cluster of the protease inhibitors including PI1 (SERPINA1), PI2, PI3, PI4 (apparently paralogues of SERPINA3), and PO1A and PO1B. SERPINA3-2 is the first porcine serpin gene whose genomic organization has been determined. PMID- 12119104 TI - The mouse Zac1 locus: basis for imprinting and comparison with human ZAC. AB - We identified a maternally methylated CpG island at the mouse Zac1 locus on chromosome (Chr.) 10 in a screen for imprinted genes. The homologous human gene ZAC (also known as LOT1 and PLAGLI) is a candidate gene for transient neonatal diabetes (TNDM), an imprinted disorder associated with paternal duplication for 6q24 and characterized by intrauterine growth retardation and insulin dependence. A mouse model would be indispensable to investigate the basis of the disorder, however, there is apparently no similar phenotype in mice with the corresponding chromosome anomaly. To begin to understand this difference, we have undertaken a comparative analysis of the mouse and human genes. We show that the CpG island is far upstream of the coding body of mouse Zac1, that Zac1 transcripts initiate in a conserved region in the CpG island, and transcripts undergo complex splicing- all properties shared with the human gene. CpG island methylation is present in oocyte DNA and constitutes a germline-specific epigenetic mark. Mice with uniparental disomy (UPD) for Chr. 10 exhibit appropriate parent-of-origin dependent expression of Zac1, indicating that the absence of phenotypes comparable to aspects of human TNDM is not because imprinting of Zac1 is relaxed in these UPD mice. PMID- 12119106 TI - Genes encoding two cystatins in the flesh fly Sarcophaga crassipalpis and their distinct expression patterns in relation to pupal diapause. AB - Two genes encoding cystatins, cysteine proteinase inhibitors, were cloned from the flesh fly Sarcophaga crassipalpis. One, sarcocystatin A (abbreviated Scys-A and SCYS-A for the gene and protein, respectively) was previously cloned from Sarcophaga peregrina, but the other is novel. Here the novel gene is named sarcocystatin B (abbreviated Scys-B and SCYS-B for the gene and protein, respectively). Tissue distribution of the two sarcocystatins differs: the fat body is the major site of Scys-A expression, while Scys-B is present in both the fat body and gut. Both Scys-A and -B are developmentally regulated, but their expression patterns also differ. Scys-A transcripts are present in both diapause- and non-diapause-destined third instar wandering larvae, and are then downregulated throughout pupal diapause. By contrast, Scys-B transcripts are only weakly expressed during the third larval instar but are highly upregulated in early diapause. The upregulation of Scys-B in early diapause suggests a possible role for this proteinase inhibitor in halting development. PMID- 12119107 TI - Human aminopeptidase B (rnpep) on chromosome 1q32.2: complementary DNA, genomic structure and expression. AB - Aminopeptidase B (APB) is a Zn(2+)-metalloexopeptidase, which selectively removes Arg and/or Lys residues from the N-terminus of several peptide substrates. Several data strongly support the hypothesis that this enzyme could participate in the final stages of precursor processing mechanisms and/or in particular inflammatory processes and tumor developments. Therefore, we have cloned the complementary DNA encoding the human APB, a 658-residues protein, containing the canonical "HEXXH(X(18))E", a signature allowing its classification in the M1 family of metallopeptidases. The genomic structure of the human APB gene (rnpep; 1q32.1-q32.2) was also determined. rnpep is bracketed by pre-protein translocase of the inner mitochondrial membrane gene and ETS family transcription factor ELF3 gene. It spans more than 24 kbp and contains 11 exons ranging from 109 to 574 bp. Finally, expression of the human APB messenger RNA (mRNA) was investigated using a pre-made dot-blot. This mRNA seems to be ubiquitous although its expression level varies depending of the cells or tissues considered. PMID- 12119108 TI - A system for deletion and complementation of Candida glabrata genes amenable to high-throughput application. AB - We describe a method for deleting or modifying genes from the pathogenic fungus Candida glabrata as well as a companion vector for complementation or ectopic expression experiments. A linear deletion fragment generated by polymerase chain reaction was used to replace a gene of interest with the C. glabrata gene encoding imidazoleglycerol-phosphate dehydratase (HIS3). As test cases, the chromosomal loci of the C. glabrata genes encoding aminoimidazole ribonucleotide carboxylase (ADE2) and encoding isopropylmalate dehydrogenase (LEU2) were deleted. To facilitate application of the deletion technique to essential genes, we also constructed vectors to allow expression of a complementing copy of the wildtype gene under control of the copper-inducible C. glabrata metallothionein I (MT-1) promoter. One version of the vector carried the Saccharomyces cerevisiae centromere (CEN) and autonomously-replicating sequence (ARS) regions. The C. glabrata ADE2 and LEU2 genes and a transposon-derived neomycin/kanamycin resistance gene were successfully expressed from this vector, with expression of the ADE2 and LEU2 genes complementing the ADE2 and LEU2 deletion mutations, respectively. However, this vector showed regulated expression only for the ADE2 gene. A second version of the vector, which carried an additional C. glabrata CEN and ARS region for stable plasmid maintenance, did show regulated expression for the LEU2 and neomycin/kanamycin resistance genes. This deletion and expression system is potentially applicable to any C. glabrata gene and is amenable to high throughput application. We anticipate that these tools will have broad utility in deletion or modification of specific C. glabrata genes. This approach is also applicable to other yeast fungi. PMID- 12119110 TI - Cloning, characterization and chromosome mapping of the human SMAP1 gene. AB - Stromal membrane associated protein (smap-1) is a new murine cell surface molecule on the stromal cells. The murine smap-1 protein is induced in stromal cells by the contact with erythroid cells, which suggests that this protein may be involved in the haematopoietic progenitor cells to stromal cells interactions. Here we report the structure, map location and expression analysis of the human SMAP1 gene, which cover approximately 100 kb on chromosome 6 between D6S455 and D6S1673 markers. This gene is composed of 11 exons and encodes a 468-amino-acid protein, which shows an 86% of homology with the murine smap-1 protein. The expression of smap-1 in erythropoietic organs as well as the correlation with the erythropoietic activity of the haematopoietic organs suggest that smap-1 is induced in stromal cells by the contact with erythroid cells, defining smap-1 as a key molecule that induced an erythropoietic microenvironment in haematopoietic organs. The high sequence conservation between murine and human SMAP1, as well as its expression in bone marrow, strongly suggest conserved functions of this protein in both organisms. Recently, a constitutional translocation t(6;10)(q13;q22) has been described in a patient with severe aplastic anaemia. SMAP1 gene localizes to 6q13 and is probably implicated in erythropoiesis, therefore it remains as an interesting candidate gene. PMID- 12119109 TI - Physical mapping and characterization of the human Na,K-ATPase isoform, ATP1A4. AB - Four isoforms of the catalytic alpha subunit of the Na,K-ATPase have been previously identified. We characterized and mapped a genomic copy of the human ATP1A4 isoform between D1S2707 and WI-9524, telomeric to a nearby isoform ATP1A2, and within a candidate region at 1q23 for familial hemiplegic migraine (FHM). Human ATP1A4 gene shares 84% identity with the mouse Atp1a4 gene, and both consist of 22 exons and 21 introns. The predicted polypeptide is 1029 amino acids and shares 82 and 79.8% identity, respectively, with human ATP1A2 and ATP1A1. ATP1A4 is larger than other isoforms and most divergent at the N-terminus. ATP1A4 and ATP1A2 are paralogous genes with the same number and organization of putative H-transmembrane domains, conserved exon-intron boundaries, and are found approximately 8.5 kb apart. Expression analysis of the ATP1A4 gene revealed a new major approximately 7.5 kb transcript in human skeletal muscle, with expression also shown in mouse muscle. Predictive analysis of promoter regions identified muscle specific regulatory elements for ATP1A4 and Atp1a4. Mutation analysis among eight affected individuals from a single large, highly penetrant FHM family was negative in ATP1A4 and ATP1A2 although multiple polymorphisms were identified. PMID- 12119111 TI - Histone H1 variants differentially inhibit DNA replication through an affinity for chromatin mediated by their carboxyl-terminal domains. AB - Multiple forms of histone H1 are found in most mammalian tissues, and diversity in their temporal and spatial expression likely corresponds to diversity in function. Here, using Xenopus egg extracts, we show that while the somatic H1s significantly inhibit DNA replication in Xenopus sperm nuclei, little or no inhibition is seen in the case of the testes-specific variant, H1t. We suggest that differences in H1-chromatin interactions might explain some of the diversity in H1 function. To demonstrate this, we show that the somatic H1 variants preferentially assemble into chromatin relative to H1t. Differences in chromatin structure are seen depending on whether chromatin assembly occurs in the presence of somatic H1s or H1t. These data suggest that the mechanistic basis for some of the functional differences of H1 variants lies in their relative affinity for chromatin. Using a series of domain-switch mutants of H1(0) and H1t we identify the H1 carboxyl-terminal domains as the domains responsible for the differential affinity for chromatin and, concurrently, for the differential effects of H1 variants upon DNA replication. PMID- 12119112 TI - Cloning and characterization of feline brain natriuretic peptide. AB - Brain (B-type) natriuretic peptide (BNP) is a cardiac hormone involved in regulation of fluid balance and blood pressure homeostasis of mammalian species. BNP sequence is species-specific and considered to be a significant prognostic and diagnostic marker for cardiac dysfunction. Using conventional polymerase chain reaction and amplification of cDNA 3'- and 5'-ends, a total of 1500 nucleotides encompassing the entire feline BNP gene were characterized. The feline BNP gene is organized in three exons separated by two introns. The complete transcript of 736 nucleotides was characterized, including 396 nucleotides encoding feline preproBNP. The preproBNP consisted of a signal peptide of 26 amino acids and a proBNP of 106 residues. The predicted mature BNP comprised 35 amino acids with likely 26- and 29-aa isomers, including a histidine residue at the C-terminus. Based on the similarity of BNP prepropeptide sequences, a phylogenetic relationship is presented for mammalian species including human, cat, cattle, dog, mouse, rat, sheep and swine. PMID- 12119113 TI - Multiple Rabaptin-5-like transcripts. AB - Rabaptin-5 is an important component of intracellular vesicular transport machinery. In this report, we describe a novel Rabaptin-5gamma transcript which has a 120 nt deletion in coding region of Rabaptin-5 messenger RNA (mRNA). We also cloned the 5'part of Rabaptin-5delta transcript and showed that upstream previously described deletion, Rabaptin-5delta mRNA is identical to Rabaptin-5. Both deletions are in-frame and result in excision of 40 and 43 aa, respectively, from Rabaptin-5 polypeptide chain. We also detected Rabaptin-5delta and Rabaptin 5gamma transcripts in human. The results of Southern blot analysis and human genome sequence analysis strongly suggest that these transcripts along with Rabaptin-5 and Rabaptin-4 are encoded by a single gene, RAB5EP, and differ due to alternative pre-mRNA splicing. Although Rabaptin-5delta and Rabaptin-5gamma transcripts are low-abundant compared to Rabaptin-5 mRNA, their ubiquitous expression patterns might point on significant roles played by encoded by them proteins. PMID- 12119114 TI - U87 RNA, a novel C/D box small nucleolar RNA from mammalian cells. AB - A novel 72 nt small nucleolar RNA (snoRNA) called U87 was found in rat liver cells. This RNA possesses the features of C/D box snoRNA family: boxes C, D', C', D, and 11 nt antisense element complementary to 28S ribosomal RNA (rRNA). The vast majority of C/D box snoRNAs direct site-specific 2'-O-ribose methylation of rRNAs. U87 RNA is suggested to be involved in 2'-O-methylation of a G(3468) residue in 28S rRNA. U87 RNA was detected in different mammalian species with slight length variability. Rat and mouse U87 RNA gene was characterized. Unlike the majority of C/D box snoRNAs U87 RNA lacks the terminal stem required for snoRNA processing. However, U87 gene is flanked by 7 bp inverted repeats potentially able to form a terminal stem in U87 RNA precursor. PMID- 12119115 TI - Identification and characterization of a site-specific tyrosine recombinase within the variable loci of Mycoplasma bovis, Mycoplasma pulmonis and Mycoplasma agalactiae. AB - Three highly mutable loci of the wall-less pathogens Mycoplasma bovis, Mycoplasma pulmonis and Mycoplasma agalactiae undergo high-frequency genomic rearrangements and generate extensive antigenic variation of major surface lipoproteins. Adjacent to each locus, an open reading frame exists as a single chromosomal copy and is predicted to encode a site-specific DNA recombinase exhibiting high homology to the recombinases XerD of Escherichia coli and CodV of Bacillus subtilis. Each of the mycoplasmal proteins are members of the lambda integrase family of tyrosine site-specific recombinases and likely mediates site-specific DNA inversions observed within the adjacent, variable loci. PMID- 12119116 TI - Lipoprotein lipase from rainbow trout differs in several respects from the enzyme in mammals. AB - Previously we found lipase activity with characteristics similar to lipoprotein lipase (LPL) in tissues from rainbow trout [Biochim. Biophys. Acta 1255 (1995) 205], whereas no equivalent to the related hepatic lipase could be found. An equivalent to apolipoprotein CII was also identified and characterized [Gene 254 (2000) 189]. We present here the full nucleotide sequence for LPL from rainbow trout (Oncorhynchus mykiss) and have investigated some properties of the enzyme. In contrast to what has been found in mammals, LPL mRNA was expressed in livers of adult trout. This indicates that trout LPL carries out functions that hepatic lipase has evolved to take over in mammals. Trout LPL was unstable at 37 degrees C compared with bovine and human LPL. Two sequence differences that may relate to the instability are that trout LPL lacks the disulfide bridge in the C-terminal domain and lacks Pro(258). This residue is conserved in LPL from all mammals and has been shown to be critical for enzyme stability at 37 degrees C. On chromatography on heparin-Sepharose trout and chicken LPL eluted at higher salt concentration than bovine (or other mammalian) LPL. The C-terminal end of LPL has been implied in heparin binding and the higher heparin affinity of the trout and chicken enzymes may be because they have 17 and 15 extra amino acid residues at the C-terminal end, of which three residues are positively charged. PMID- 12119117 TI - Nucleotide sequence of phospholipase A(2) gene expressed in snake pancreas reveals the molecular evolution of toxic phospholipase A(2) genes. AB - We have cloned two phospholipase A(2) (PLA(2)) DNA complementary to RNA that contained nucleotide sequences encoding pancreas loop by the reverse transcription-polymerase chain reaction cloning procedure using messenger RNA isolated from Laticauda semifasciata pancreas. Additionally, a gene clone encoding PLA(2) with the pancreatic loop sequence was isolated from a L. semifasciata genomic library. Subsequent sequence analysis revealed that PLA(2) clones encoding group IB" PLA(2). Comparative analysis of group IA and IB" PLA(2) genes revealed that the exon-intron organization is conserved in the genes of both groups. The invaded sequences in the second intron were very similar to those of the L. semifasciata group IA gene. This observation suggested that the integration of the invaded sequences occurred before the divergence of groups IA and IB" during the evolution of PLA(2) gene. The comparative analysis revealed that the arising of group IA PLA(2) occurred by the deletion and substitution of nucleotide sequences in exon III region during the process of accelerated evolution. PMID- 12119118 TI - Petunia actin-depolymerizing factor is mainly accumulated in vascular tissue and its gene expression is enhanced by the first intron. AB - Actin-depolymerizing factor (ADF) is one of the actin cytoskeleton-modulating proteins. We have characterized the accumulation pattern of petunia ADF proteins. PhADF proteins are accumulated in every petunia organ and their accumulation is differentially regulated by developmental signals. Their cellular localization is vascular tissue-preferential in vegetative organs, whereas somewhat different in reproductive organs. In reproductive organs, PhADFs are present in outer integument, endocarp of ovary wall, transmitting tissue of style, and epidermis and endothecium of young anther. From a petunia genomic library, we have isolated a genomic clone encoding PhADF1. Comparison to complementary DNA sequence revealed that the coding region of PhADF1 gene consists of three exons and two introns. Analysis of chimeric gene expression using beta-glucuronidase as a reporter gene in transgenic Arabidopsis revealed that PhADF1 was strongly expressed in every vegetative tissue except petal. In addition, expression of the gene was highly enhanced by its first intron. These results suggest that PhADF1 gene of petunia is mainly expressed in vascular tissues and its expression is regulated by intron-mediated enhancement mechanism. PMID- 12119119 TI - Structure, expression, and functional characterization of the mouse CLP-1 gene. AB - Mouse CLP-1, a potential cardiac transcriptional regulatory factor, is encoded by a single copy gene lacking introns that is expressed into two mRNAs via alternative polyadenylation. Both mRNAs encode the same 41 kDa protein, a novel protein that is 85.3% homologous with a human homologue called HIS1. Mouse CLP-1 is widely expressed in a number of tissues as well as in early development and is localized to the nucleus. The CLP-1 gene promoter is active in different cell types and sequence analysis shows a number of potential binding sites for cardiogenic transcription factors such as Nkx2.5 and GATA-4, indicating a potential role in development. CLP-1 appears to "squelch" the cardiac MLC-2v promoter in a concentration-dependent manner in cardiac but not other cell types, suggesting that CLP-1 may be interacting with a cardiac-specific factor to regulate cardiac MLC-2v expression. The overall expression pattern of CLP-1 is similar to that of LCR-F1 and Oct-1, two widely expressed transcription factors that also play specific roles in the transcription of cell-specific genes. CLP-1 may be a transcriptional mediator capable of interacting with and potentiating cell-specific transcription factors. PMID- 12119121 TI - Predicting three narcosis mechanisms of aquatic toxicity. AB - In this study, the use of solute descriptors (the McGowan's characteristic volume V(X), the excess molar refraction R, the dipolarity/dipolarizability pi(H), the effective or summation hydrogen-bond acidity summation operatoralpha(H) and the effective or summation hydrogen-bond basicity summation operatorbeta(H)) in classifying and predicting the non-polar, polar, and ester narcosis toxicity mechanisms for organic compounds was investigated. Discriminant analysis was performed and the significant discriminating variables were found to be R, pi(H), sigma alpha(H), and (sigma beta (H))(2), the latter of which was created to aid the mechanism classifications. Cross-validation of the non-linear discriminant functions showed a small total error rate of approximately 5% which was reduced to approximately 2% when seven compounds with uncertain a priori mechanism designations were removed. Compared with other methods for toxicity mechanism classification and prediction, the method developed in this study has advantages. It relies on the use of objective numerical values of molecular descriptors that can be calculated and does not require additional experimental work when used for new compounds. The descriptor values can also aid the interpretation of the toxicity mechanism classifications and predictions. Because of the possibility of making incorrect mechanism predictions using a single method, it was recommended that several methods be used together to make the most appropriate mechanism designations and to increase the confidence level associated with the mechanism designations. PMID- 12119122 TI - Metabolism of caffeic acid by isolated rat hepatocytes and subcellular fractions. AB - Caffeic acid (CA) is found in a wide variety of foods such as vegetables, fruits, tea, coffee, and wine. However, enzymes involved in its metabolism have not been identified. In the following, caffeic (CA), chlorogenic (CGA), and dihydrocaffeic (DHCA) acids were incubated with hepatocytes and shown to undergo metabolism by cytochrome P450, catechol-O-methyltransferase (COMT), and beta-oxidation enzymes. Ferulic (FA) or dihydroferulic (DHFA) acids, formed as the result of CA- or DHCA O-methylation by COMT, were also O-demethylated by CYP1A1/2 but not CYP2E1. DHCA or DHFA also underwent side chain dehydrogenation to form CA and FA, respectively, which was prevented by thioglycolic acid, an inhibitor of the beta oxidation enzyme acyl CoA dehydrogenase. The rates of glutathione conjugate formation catalyzed by NADPH/microsomes (CYP2E1) in decreasing order DHCA>CA>CGA trend which was in reverse order to the rates of their O-methylation by COMT. The CA- and DHCA-o-quinones formed by NADPH/P450 likely inhibited COMT but can readily form glutathione conjugates. CA, DHCA and DHFA were inter-metabolized to each other and to FA by isolated rat hepatocytes whereas FA was metabolized only to CA but not to DHCA or DHFA. CA, DHCA, FA, DHFA and CGA showed a dose-dependent hepatocyte toxicity and the LD(50) (2 h), determined were in decreasing order of effectiveness DHCA>CA>DHFA>CGA>FA. In summary, evidence has been provided that O methylation, GSH conjugation, hydrogenation and dehydrogenation are involved in the hepatic metabolism of CA and DHCA. The O-methylation pathway for CA and DHCA is a detoxification route whereas o-quinones formation catalyzed by P450 is the toxification route. PMID- 12119123 TI - Continuous system modeling of equilibrium dialysis for determinations of tissue partitioning of parathion and paraoxon. AB - A tissue/blood partition coefficient, defined as the ratio of tissue chemical concentration to that of the venous outflow of the tissue when at equilibrium, is an important parameter required for physiological based pharmacokinetic models. While many techniques have been developed to quantify tissue/blood partition coefficients for various chemicals, there is no single best approach for their determination. In the current study, equilibrium dialysis of the organophosphorus insecticide parathion and its active metabolite paraoxon was undertaken to assess their partitioning into rat liver. A mass balance analysis of the contents of the dialysis cells suggested that significant levels of parathion and paraoxon were bound to the dialysis membranes. There was no evidence of metabolism of either parathion or paraoxon by the very dilute liver homogenate utilized in the dialysis. In order to investigate the potential impact of binding of a chemical to dialysis membrane during determination of partition coefficients, a computer model of a dialysis system was constructed. The model assumed that all processes occurring within the dialysis cell were first or second order in nature, and that binding to the dialysis membrane occurred symmetrically on both sides of the membrane. Variations in the total number of simulated binding sites on dialysis membrane revealed that increasing the degree of membrane binding resulted in decreased compound on the homogenate and buffer sides of the dialysis cells. However, the final tissue/buffer partition coefficient was unaffected by these alterations in membrane binding, although increased membrane binding prolonged the incubation time required to achieve equilibrium. These simulations suggest that loss of a compound to membrane binding does not preclude the use of equilibrium dialysis for determination of tissue/buffer, and therefore tissue/blood, partition coefficients, provided the dialysis system is allowed to proceed to equilibrium. PMID- 12119124 TI - A biochemical, histochemical, and ultrastructural evaluation of the effect of dimethoate intoxication on rat pancreas. AB - Organophosphorus compounds are widely used in industry, agriculture and for public health purposes. They are among the toxic compounds employed for insect control. The purpose of this work was to study biochemical, histochemical, and histological as well as ultrastructural changes that might occur in the pancreas of adult male Wistar rats as a result of chronic dimethoate intoxication. The treated group received dimethoate orally via gavage (21 mg/kg) daily for 2 months while, the control group was given saline orally (0.1 ml/100 g/day) for the same period. Plasma glucose level was significantly increased while, plasma insulin level was decreased in the intoxicated animals compared with the control group. A patchy reduction of histochemically-detected succinic dehydrogenase enzymatic activity was observed in the pancreas of the intoxicated rats. By contrast, acid phosphatase enzymatic activity was markedly increased in the pancreas of the intoxicated group. No changes were observed in alkaline phosphatase or alpha esterase activities of the intoxicated animals. Light microscopic examination revealed that dimethoate caused patchy degenerative changes of variable severity in many areas of the pancreas affecting both the pancreatic acini and islets of Langerhans. Ultrastructurally, some beta cells revealed dense nuclei with wide perinuclear cisternae. Diminution of the number of beta granules was evident. One month after discontinuation of the dimethoate, all the above mentioned changes induced by dimethoate intoxication persisted. These findings show that chronic exposure to dimethoate insecticide has clear toxic effect on the rat pancreas, which was not reversible within 1 month. Public health education is necessary to raise people awareness about the hazards accompanying the use of such compounds. PMID- 12119126 TI - Normal mammary gland morphology in pubertal female mice following in utero and lactational exposure to genistein at levels comparable to human dietary exposure. AB - The objective of the study was to determine the effect of in utero and lactational exposure to genistein (0, 0.1, 0.5, 2.5 and 10 mg/kg/day) on mammary gland morphology in female B6D2F1 mice at levels comparable to or greater than human exposures. The effect of diethylstilbestrol (DES; 0, 0.1, 1, 10 microg/kg/day) on the mammary gland was also examined as a positive estrogenic control. Pregnant females were treated by daily gavage from gestational day 12 to postnatal day (PND) 20. Female offspring were weaned on PND21 and mammary gland whole mounts were examined for growth (length and area of the epithelial tree), proliferation (number of terminal end buds (TEBs)), and differentiation (density of alveolar buds (ABs)) on PND49. The highest dose of DES induced a significant increase in mammary gland growth (P<0.05) and also decreased the number of TEBs (P<0.06). The density of ABs was not significantly affected by DES. By contrast to DES, genistein had no effect on mammary gland morphology at any dose. These results suggest that in utero and lactational exposure to genistein at levels comparable to or greater than human exposures do not adversely affect mammary gland development in pubertal female B6D2F1 mice. PMID- 12119125 TI - Synergistic cytotoxicity of Delta(9)-tetrahydrocannabinol and butylated hydroxyanisole. AB - We examined the food additive, butylated hydroxyanisole (BHA), for its capacity to modulate the cytotoxic effects of Delta(9)-tetrahydrocannabinol (THC). THC was not cytotoxic when added to cultures of A549 lung tumor cells at concentrations<5 microg/ml, but induced cell necrosis at higher levels with an LC(50)=16-18 microg/ml. BHA alone, at concentrations of 10-200 microM, produced limited cell toxicity but significantly enhanced the necrotic death resulting from concurrent exposure to THC. In the presence of BHA at 200 microM, the LC(50) for THC decreased to 10-12 microg/ml. Similar results were obtained with smoke extracts prepared from marijuana cigarettes, but not with extracts from tobacco or placebo marijuana cigarettes (containing no THC). Two different mechanisms for this synergistic cytotoxicity were investigated. Experiments were repeated in the presence of either diphenyleneiodonium or dicumarol as inhibitors of the redox cycling pathway. Neither of these compounds protected cells from the effects of combined THC and BHA, but rather enhanced necrotic cell death. Measurements of cellular ATP revealed that both THC and BHA reduced ATP levels in A549 cells, consistent with toxic effects on mitochondrial electron transport. The combination was synergistic in this respect, reducing ATP levels to <15% of control. Exposure to marijuana smoke in conjunction with BHA, a common food additive, may promote deleterious health effects in the lung. PMID- 12119127 TI - Ascorbyl radical/ascorbate ratio in plasma from iron overloaded rats as oxidative stress indicator. AB - Oxidative stress has been developed using dietary carbonyl-iron and iron-dextran parenteral administration as models of in vivo iron overload in rats. Carbonyl iron led to a 2-fold increase in plasma iron content, a significant decrease (34%) in ascorbate plasma content and non-significant changes in plasma ascorbyl radical content. Iron-dextran produced a dramatic increase (6.7-fold) in plasma iron content, overwhelming the plasma total iron binding capacity. The ascorbyl radical content increased significantly in iron-dextran treatment (2.6-fold) and plasma ascorbate level was not affected. Ascorbyl radical/ascorbate ratio was significantly higher in both iron treated groups as compared with the control group (4 x 10(-4)+/-1 x 10(-4)). Data reported here indicate that the ascorbyl radical/ascorbate ratio is an appropriate in vivo indicator of oxidative stress under conditions of iron overload. The overall mechanism that describes the ascorbate status in plasma seems to be strongly dependent on the way the excess of iron is stored and thus, to the availability of the catalytically active iron for interacting with the plasma components. On this regard, evaluation of A*/AH( ) ratio did not help to discriminate between the possible involved mechanisms. PMID- 12119128 TI - Oxygenated water does not induce genotoxic effects in the comet assay. AB - Drinking of oxygenated water (i.e. water with increased concentration of physically dissolved oxygen) is said to improve oxygen availability of the body and will do the consumer good. However, increased oxygen concentrations can also lead to an increased production of reactive oxygen species (ROS). If antioxidant defences are not completely efficient, ROS can cause cell injury including DNA damage. We therefore investigated whether drinking of oxygenated water can lead to increased DNA damage in peripheral blood cells of test subjects. We also tested whether direct exposure of V79 Chinese hamster cells to oxygenated medium or oxygenated Hank's solution for various time periods induces DNA damage. Induction of DNA damage was measured with the alkaline comet assay (single cell gel electrophoresis). The comet assay, in particular the modification with FPG post-treatment for the determination of oxidative DNA base damage, has been proven to be extremely sensitive for the detection of oxygen-induced DNA damage. However, both the in vivo and the in vitro studies with the comet assay in the absence and presence of FPG post-treatment did not provide evidence for a genotoxic effect of oxygenated water. PMID- 12119130 TI - Estrogen receptor alpha in mouse splenic lymphocytes: possible involvement in immunity. AB - To elucidate relevance of estrogens to immune responses, we investigated whether estrogen receptor alpha (ERalpha) exists in mouse splenic B cells and T cells and the effect of 17beta-estradiol and endocrine disrupting chemicals (EDCs) on lymphocyte mitogenesis. ERalpha was identified in both male and female mouse splenic cells using RT-PCR. Crude splenic cells were stained with anti-ER antibody, and the distribution of ERalpha in the splenic B cells and part of the splenic T cells was confirmed by flow cytometry. 17beta-Estradiol inhibited B cell mitogenesis at the concentration of 10(-8) M and T cell mitogenesis at the concentration of 10(-6) M. Some EDCs, diethylstilbestrol, bisphenol A, p nonylphenol and di-2-ethylhexylphthalate, suppressed lymphocyte mitogenesis at the concentration of 10(-6)-10(-5) M. We therefore suggest that estrogen may suppress lymphocyte mitogenesis through ERalpha in B and T cells. PMID- 12119129 TI - Cellular activation and neuronal transport of intranasally instilled benzo(a)pyrene in the olfactory system of rats. AB - Nasal tissues can be exposed to benzo(a)pyrene (BaP), e.g. present in diesel exhaust particles and some workplace atmospheres. In this study rats were given 3H-BaP intranasally. Autoradiography and beta-spectrometry were then used to trace cells in the nasal olfactory mucosa having capacity to activate the compound to tissue-bound metabolites. We also examined if deposition of 3H-BaP on the olfactory mucosa results in translocation of labelled material to the brain along olfactory neurons. The results showed that intranasal administration of 3H BaP results in formation of tissue-bound metabolites in sustentacular cells and in the cells of Bowman's glands. Initially the bound material was localised to a higher extent to the sustentacular cells than to the cells of Bowman's glands, whereas at longer survival intervals the uptake in the cells of Bowman's glands dominated. In the latter the covalently bound material was accumulated to a higher extent in the nuclei than in the cytoplasms. We speculate that BaP may interact with the aryl hydrocarbon receptor (AhR) in these cells and that AhR may target activated BaP to the nucleus. Our results further indicated that application of 3H-BaP on the nasal mucosa results in transport of BaP and/or BaP metabolites along the axons of the olfactory neurons to the olfactory bulb. PMID- 12119131 TI - 5-Fluorouracil-induced intestinal toxicity: what determines the severity of damage to murine intestinal crypt epithelia? AB - To elucidate the relationship between 5-fluorouracil (5-FU) distribution and 5-FU induced apoptosis and/or cell cycle arrest, microautoradiography was applied to murine intestinal crypts exposed to [14C] 5-FU by intravenous infusion. The histologic location of apoptotic cells in the crypt did not correlate to that of 5-FU. The temporal profiles of apoptotic and/or mitotic indexes corresponded to those of orally administered 5-FU in a previous study. Two cell cycle-related proteins, p21(WAF/Cip1) and bax, were also investigated in the present study. With time, p21(WAF/Cip1)-positive nuclei apparently migrated up the crypt. Bax positive cytoplasm was observed throughout the crypt epithelial cells, accompanied by the occurrence of apoptosis, and remained until 48 h when the control level recovered. The findings demonstrate that 5-FU mainly exerts an apoptotic effect and/or cell cycle arrest by systemic exposure, and p21 and bax expression determine an individual cell's fate. PMID- 12119132 TI - Expression of hsp 90 in the human kidney and in proximal tubule cells exposed to heat, sodium arsenite and cadmium chloride. AB - The expression of heat shock protein (hsp) 90alpha and beta mRNA and protein were determined in the human kidney and in human proximal tubule (HPT) cells exposed to lethal and sub-lethal concentrations of Cd(+2) under both acute and extended conditions of exposure. Using immunohistochemical analysis, it was demonstrated that hsp 90 was widely distributed in the human adult and fetal kidney. Moderate to strong staining was observed in the straight portions of the distal and proximal tubules, the distal convoluted tubule, the collecting ducts and the parietal epithelium of Bowmans capsule in the glomerulus. Moderate staining was observed in the proximal convoluted tubule of the cortex and the thick loops of Henle within the medulla. In addition, the fetal kidney demonstrated strong staining of the blastema, the 'S-shaped' bodies, and the developing glomeruli. Analysis of hsp 90alpha and beta mRNA expression in total RNA isolated from in situ microdissected proximal tubules or HPT cells demonstrated similar expression levels of both the alpha and beta isoforms in this tubule segment. It was demonstrated that HPT cells exhibited the classic heat shock response when subjected to a physical (heat) or chemical stress (NaAsO(2)). Heat stress, elevated temperature at 42.5 degrees C for 1 h, caused a modest increase in both hsp 90alpha and beta mRNA and protein. Similar results were obtained when the cells were subjected to a classic chemical stress of exposure to 100 microM NaAsO(2) for 4 h. In contrast, acute exposure of HPT cells to 53.4 microM CdCl(2) for 4 h resulted in no consistent increase in hsp 90alpha and beta mRNA or protein. Chronic exposure to Cd(+2) likewise failed to increase either hsp 90 mRNA or protein expression, even at concentrations of Cd(+2) that were lethal to the cells during the time course. This study shows that the HPT has a high basal expression of hsp 90, which is not induced by Cd(+2) exposure. PMID- 12119133 TI - Acute alcohol intoxication decreases cell proliferation and nitric oxide synthase expression in dentate gyrus of rats. AB - To study the dose-dependence of the effect of alcohol on cell proliferation and nitric oxide synthase expression, rats were first divided into the control group, the 0.5 g/kg alcohol-treated group, the 1 g/kg alcohol-treated group, the 2 g/kg alcohol-treated group, and the 4 g/kg alcohol-treated group. To study the duration-dependence of this effect, animals were divided into the control, the 1 day-alcohol-treated group, the 3-days-alcohol-treated group, and the 6-days alcohol-treated group; 2 g/kg of alcohol was applied to rats over the respective number of days. Alcohol injection reduced the number of both 5-bromo-2' deoxyuridine-positive and nicotinamide adenine dinucleotide phosphate-diaphorase positive cells in the dentate gyrus of rats in a dose- and duration-dependent manner. These results suggest that alcohol-induced impairment in memory is due to the inhibition of new cell formation, and it is possible that nitric oxide, its synthesis is affected adversely by alcohol, plays an important role in the regulation of cell proliferation. PMID- 12119134 TI - Porcine reproductive-respiratory syndrome virus infection predisposes pigs for respiratory signs upon exposure to bacterial lipopolysaccharide. AB - This study examined whether an infection with porcine reproductive and respiratory syndrome virus (PRRSV) potentiates respiratory signs upon exposure to bacterial lipopolysaccharides (LPS). Five-week-old conventional pigs were inoculated intratracheally with the Lelystad strain of PRRSV and received 5 days later one or two intratracheal LPS administrations. The necessary controls were included. After LPS administration, pigs were intensively monitored for clinical signs. Additionally, some pigs were euthanatized after a second LPS administration for broncho-alveolar cell analysis and virological examinations of the lungs. Broncho-alveolar lavage (BAL) cells were counted and differentiated. Lung suspensions and BAL fluids were titrated for PRRSV. Exposure of pigs to PRRSV only resulted in a fever for time periods ranging from 1 to 5 days and slight respiratory signs. Exposure of pigs to LPS only resulted in general signs, characterized by fever and depression, but respiratory signs were slight or absent. PRRSV-LPS exposed pigs, on the other hand, developed severe respiratory signs upon LPS exposure, characterized by tachypnoea, abdominal breathing and dyspnoea. Besides respiratory signs, these pigs also showed enhanced general signs, such as fever and depression. Lung neutrophil infiltration was similar in non-infected and PRRSV-infected pigs upon LPS exposure. PRRSV quantities were similar in lungs and BAL fluids of pigs infected with PRRSV only and PRRSV-LPS exposed pigs. These data show a clear synergism between PRRSV and LPS in the induction of respiratory signs in conventional pigs. The synergism was observed in 87% of the pigs. So, it can be considered as reproducible and may be used to test the efficacy of preventive and therapeutic measures. PMID- 12119135 TI - Serological responses of mares and weanlings following vaccination with an inactivated whole virus equine herpesvirus 1 and equine herpesvirus 4 vaccine. AB - Equine herpesvirus 1 (EHV-1) is a major cause of respiratory disease and abortion in horses worldwide. Although some vaccines have been shown experimentally to reduce disease, there are few reports of the responses to vaccination in the field. This study measured antibody responses to vaccination of 159 mares (aged 4 17 years) and 101 foals (aged 3-6 months) on a large stud farm with a killed whole virus EHV-1/4 vaccine used as per the manufacturer's recommendations. Using an EHV glycoprotein D (gD)-specific ELISA and a type-specific glycoprotein G (gG) ELISA, respectively 13.8 and 28.9% of mares, and 42.6 and 46.6% of foals were classed as responding to vaccination. Additionally, 16.4 and 17.6% of mares were classified as persistently seropositive mares. Using both assays, responder mares and foals had lower week 0 mean ELISA absorbances than non-responder mares and foals. Responder mares were ten times more likely to have responder foals, and non-responder mares were six times more likely to have non-responder foals than other mares using the gG ELISA. Mares aged 7 years or less and foals aged 4 months or more were more likely to respond to vaccination than animals in other age groups. There was no association between response of mares and the number of previous vaccinations received and persistently seropositive mares did not respond to vaccination. This study documents the responses of mares and foals to vaccination in a large scale commercial environment in 2000, and suggests that knowledge of antibody status may allow a more selective vaccination strategy, representing considerable savings to industry. PMID- 12119136 TI - Viral infections and bovine mastitis: a review. AB - This review deals with the role of viruses in the aetiology of bovine mastitis. Bovine herpesvirus 1, bovine herpesvirus 4, foot-and-mouth disease virus, and parainfluenza 3 virus have been isolated from milk from cows with clinical mastitis. Intramammary inoculations of bovine herpesvirus 1 or parainfluenza 3 virus-induced clinical mastitis, while an intramammary inoculation of foot-and mouth disease virus resulted in necrosis of the mammary gland. Subclinical mastitis has been induced after a simultaneous intramammary and intranasal inoculation of lactating cows with bovine herpesvirus 4. Bovine leukaemia virus has been detected in mammary tissue of cows with subclinical mastitis, but whether this virus was able to induce bovine mastitis has not been reported. Bovine herpesvirus 2, vaccinia, cowpox, pseudocowpox, vesicular stomatitis, foot and-mouth disease viruses, and bovine papillomaviruses can play an indirect role in the aetiology of bovine mastitis. These viruses can induce teat lesions, for instance in the ductus papillaris, which result in a reduction of the natural defence mechanisms of the udder and indirectly in bovine mastitis due to bacterial pathogens. Bovine herpesvirus 1, bovine viral diarrhoea virus, bovine immunodeficiency virus, and bovine leukaemia virus infections may play an indirect role in bovine mastitis, due to their immunosuppressive properties. But, more research is warranted to underline their indirect role in bovine mastitis. We conclude that viral infections can play a direct or indirect role in the aetiology of bovine mastitis; therefore, their importance in the aetiology of bovine mastitis and their economical impact needs further attention. PMID- 12119137 TI - In vitro development of resistance to enrofloxacin, erythromycin, tylosin, tiamulin and oxytetracycline in Mycoplasma gallisepticum, Mycoplasma iowae and Mycoplasma synoviae. AB - The in vitro emergence of resistance to enrofloxacin, erythromycin, tylosin, tiamulin, and oxytetracycline in three avian Mycoplasma species, Mycoplasma gallisepticum, Mycoplasma synoviae and Mycoplasma iowae was studied. Mutants were selected stepwise and their MICs were determined after 10 passages in subinhibitory concentrations of antibiotic. High-level resistance to erythromycin and tylosin developed within 2-6 passages in the three Mycoplasma species. Resistance to enrofloxacin developed more gradually. No resistance to tiamulin or oxytetracycline could be evidenced in M. gallisepticum or M. synoviae after 10 passages whereas, resistant mutants were obtained with M. iowae. Cross sensitivity tests performed on mutants demonstrated that mycoplasmas made resistant to tylosin were also resistant to erythromycin, whereas mutants made resistant to erythromycin were not always resistant to tylosin. Some M. iowae tiamulin-resistant mutants were also resistant to both macrolide antibiotics. Enrofloxacin and oxytetracycline did not induce any cross-resistance to the other antibiotics tested. These results show that Mycoplasma resistance to macrolides can be quickly selected in vitro, and thus, providing that similar results could be obtained under field conditions, that development of resistance to these antibiotics in vivo might also be a relatively frequent event. PMID- 12119138 TI - Characterization of the in vitro adhesion of Actinobacillus pleuropneumoniae to swine alveolar epithelial cells. AB - Actinobacillus pleuropneumoniae biovar 1 serotypes 2, 5a, 9 and 10 strains were tested for their ability to adhere to alveolar epithelial cells in culture. For the serotypes 5a, 9 and 10 strains, optimal adherence was observed after growth of bacterial cells in a NAD-restricted medium (0.001% NAD). This condition was also associated with the expression of a 55 kDa outer membrane protein (OMP) and of fimbriae. For the serotype 2 strain, adherence and expression of fimbriae and a 55 kDa OMP was less influenced by the growth conditions. The N-terminal amino acid sequence of the 55 kDa OMP had no homology with any known sequence, suggesting that it is an as yet unknown protein. Adherence capabilities were significantly reduced following treatment of the bacteria with proteolytic enzymes or heat. These findings suggest that proteins are involved in adhesion. The hydrophobic bond-breaking agent tetramethylurea was unable to inhibit the adherence of A. pleuropneumoniae to alveolar epithelial cells. Treatment of the bacteria with sodium metaperiodate resulted in lower adhesion scores for the serotypes 2 and 9 strains but the inhibition of adhesion was clearly lower than after treatment with proteolytic enzymes. This indicates that, besides proteins, carbohydrates might also be involved in adhesion of A. pleuropneumoniae to alveolar epithelial cells. The finding that inhibition of adhesion was very high when bacteria were treated with a combination of sodium metaperiodate and pronase also suggests that more than one adhesin is involved. PMID- 12119139 TI - Identification of Corynebacterium pseudotuberculosis isolates from sheep and goats by PCR. AB - The present study was carried out to estimate the prevalence of caseous lymphadenitis (CL) in sheep and goats slaughtered at the local abattoir in Elazig province located in the east of Turkey, between September and December 2000. A total of 2046 sheep and 2262 goat carcasses were examined during the study period and 118 abscessed lymph nodes, 89 from sheep and 29 from goats, were collected. Corynebacterium spp. strains were isolated from 81.4% of the abscesses, giving an overall prevalence of 2.2%. The prevalence was 3.5 and 1.1% in sheep and goats, respectively. PCR on DNA extracted from 96 suspicious isolates, using a pair of Corynebacterium pseudotuberculosis-specific primers, was positive for 93. Although cross-reaction with C. ulcerans, a human/bovine species, was observed, the PCR assay used in this study may successfully be applied for the diagnosis of CL in goats and sheep as an alternative to conventional methods, owing to its advantages of specificity and speed. PMID- 12119140 TI - Comparison between immune responses and resistance induced in BALB/c mice vaccinated with RB51 and Rev. 1 vaccines and challenged with Brucella melitensis bv. 3. AB - BALB/c mice were immunized with live rough Brucella abortus RB51 or smooth Brucella melitensis Rev. 1 vaccines and challenged with a B. melitensis field strain. Protection was assessed by a variety of serological tests and recovery of vaccinal and challenge strains by culture. Mice vaccinated with RB51 gave negative results in the conventional serological tests prior to challenge, namely; standard tube agglutination test (SAT), Rose Bengal plate test (RBPT), buffered acidified plate antigen test (BAPAT), and mercaptoethanol test (MET). Sero-conversion took place to a whole-cell bacterial buffered RB51 antigen after vaccination and persisted for 7 weeks post-vaccination. Mice challenged with B. melitensis were assessed for bacterial load and immune response for 12 weeks after challenge. Protection units were showed that Rev. 1 vaccine was superior to RB51 vaccine in protection of mice against B. melitensis. However, RB51 vaccine has the advantage that it would not elicit antibodies to standard serological tests based on the LPS O antigen. RB51 vaccine could therefore be used for control of B. melitensis infection and avoid confusion in the use of standard sero-diagnostic tests. PMID- 12119141 TI - Application of N-PCR for diagnosis of distemper in dogs and fur animals. AB - The immunofluorescence test, routinely used for laboratory diagnosis of canine distemper virus (CDV) in Poland, is not sufficiently sensitive and specific. Therefore, the application of reverse transcriptase polymerase chain reaction (RT PCR), nested PCR (N-PCR) and Southern blot hybridization for detection of phosphoprotein (P) gene of CDV in peripheral blood mononuclear cells (PBMCs) or internal organs of dogs and fur animals was the aim of these studies. The optimal parameters for two-step PCR were elaborated for reference strains of distemper virus and used for testing biological samples collected from dogs, foxes, ferret and mink with spontaneous distemper. PCR product of 1069bp was obtained in one out of 10 dog blood samples, three out of 14 homogenates of internal organs of dogs and one out of five homogenates of internal organs of fox. Reamplification with the use of CDVa and CDVb primers demonstrated the 429bp fragment in six samples, negative by PCR: two samples collected from dogs, two from foxes, one from mink and one from ferret. The specificity of N-PCR was confirmed by Southern blot hybridization. We conclude that two-step PCR is sensitive and specific method for diagnosis of CDV infection. PMID- 12119142 TI - Cerebral cystine uptake: a tale of two transporters. AB - Transport of cystine across the cell membrane is essential for synthesis of the major cellular antioxidant glutathione. Cystine uptake in the brain occurs by both the Na(+)-independent x(c)(-) cystine-glutamate exchanger and the X(AG)(-) family of high-affinity, Na(+)-dependent glutamate transporters. New evidence concerning the role of cystine transport in the defence against oxidative stress is described. PMID- 12119143 TI - Structure and enzymology of a death-associated protein kinase. AB - The structure of the catalytic domain of death-associated protein kinase (DAPK), a serine/threonine kinase implicated in programmed cell death and tumor suppression, has been determined recently in several crystal forms, including the highest resolution kinase structure available. The structures provide detailed knowledge about ATP-kinase complexes, reveal novel features that might be important in the regulation of enzyme activity, and allow a proteomics-type approach to DAPK enzymology and its role in signal transduction pathways. PMID- 12119147 TI - Apoptosis: coming to terms with biological diversity. PMID- 12119148 TI - Apoptosis: from morphological types of cell death to interacting pathways. PMID- 12119149 TI - Apoptosis: ignoring morphology and focusing on biochemical mechanisms will not eliminate confusion. PMID- 12119151 TI - Asthma -- a need for a rethink? AB - Asthma is a major medical problem but, despite decades of research, the mechanisms that underlie this condition remain elusive. Although the eosinophil has been regarded as a cell that is central to the pathogenesis of asthma, the failure to abrogate asthma symptoms by novel treatments that are designed to suppress the recruitment of eosinophils to the airways challenges this dogma. Our approach to understanding bronchial asthma needs to be broadened to include alterations in the function of afferent nerves that supply airways. Changes in the activity of these nerves offer a possible mechanism by which asthmatic subjects are uniquely responsive to a wide range of physiological and chemical stimuli. Here, we review the current status of asthma research. PMID- 12119152 TI - To make antibodies or not: signaling by the B-cell antigen receptor. AB - Antibodies produced by B cells play an essential role in protecting against disease-causing pathogens. B cells detect the presence of pathogens via B-cell antigen receptors (BCRs), which consist of a transmembrane form of an antibody that is associated with a signaling subunit. Signaling by BCRs not only initiates antibody production but also regulates B-cell development, B-cell survival and the elimination of B cells that recognize components of one's own body. Identifying the intracellular signals generated by BCRs and determining how these signals specify such diverse responses is the key to understanding how the immune system functions normally and how defects in BCR signaling can lead to either immunodeficiency diseases or autoimmune diseases. PMID- 12119153 TI - Novel therapeutic strategies provide the real test for the amyloid hypothesis of Alzheimer's disease. AB - The amyloid and tangle cascade hypothesis is the dominant explanation for the pathogenesis of Alzheimer's disease (AD). A complete knowledge of the metabolic pathways leading to beta-amyloid (A beta) production and clearance in vivo and of the pathological events that lead to fibril formation and deposition into plaques is crucial for the development of an 'anti-amyloid' therapeutic strategy. Important advances in this respect have been achieved recently, revealing new candidate drug targets. Among the most promising potential treatments are beta- and gamma-secretase inhibitors, A beta vaccination, Cu-Zn chelators, cholesterol lowering drugs and non-steroidal anti-inflammatory drugs. Now, the major question is whether these drugs will work in the clinic. PMID- 12119154 TI - Applications of hormesis in toxicology, risk assessment and chemotherapeutics. AB - There is much debate over the fundamental shape of the dose-response curve in the low-dose zone, particularly in the fields of toxicology and risk assessment. The defaults, principally accepted dose-response models in the major texts in these areas and in government regulatory activities, are a threshold model for non carcinogens and a linear model for most carcinogens. We have argued that in properly designed studies the U-shaped hormetic response predominates and is more fundamental. In this article, a broad range of basic issues associated with the acceptance of U-shaped dose responses as central to toxicology, pharmacology and their applications to risk assessment and medicine will be discussed. PMID- 12119155 TI - Are we beta-ARKing up the wrong tree? Casein kinase 1 alpha provides an additional pathway for GPCR phosphorylation. AB - Although originally linked to receptor desensitization, G-protein-coupled receptor (GPCR) phosphorylation has now been implicated in coupling receptors to specific signalling pathways. Generally, this phosphorylation event is thought to be mediated by one of the members of the GPCR kinase (GRK) family. However, recent studies have indicated that protein kinases distinct from the GRK family might also be involved in agonist-mediated GPCR phosphorylation. This review analyses the approaches employed to investigate the nature of GPCR phosphorylation and discusses recent developments implicating other kinases, particularly casein kinase 1 alpha, in the phosphorylation of GPCRs. PMID- 12119158 TI - C(4) photosynthesis in terrestrial plants does not require Kranz anatomy. AB - C(4) photosynthesis in terrestrial plants was thought to require Kranz anatomy because the cell wall between mesophyll and bundle sheath cells restricts leakage of CO(2). Recent work with the central Asian chenopods Borszczowia aralocaspica and Bienertia cycloptera show that C(4) photosynthesis functions efficiently in individual cells containing both the C(4) and C(3) cycles. These discoveries provide new inspiration for efforts to convert C(3) crops into C(4) plants because the anatomical changes required for C(4) photosynthesis might be less stringent than previously thought. PMID- 12119159 TI - New members of the floral organ identity AGAMOUS pathway. AB - The Arabidopsis floral organ identity gene AGAMOUS (AG) specifies stamen and carpel development as well as floral determinacy. Recent reports suggest that the HUA1, HUA2, HEN1 and HEN2 genes function redundantly as components of the AG pathway. The HUA1, HUA2, HEN1 and HEN2 genes encode nuclear proteins that perhaps play a role in RNA metabolism. The HUA and HEN genes function not only on the AG pathway, but also in vegetative development. PMID- 12119165 TI - From elicitins to lipid-transfer proteins: a new insight in cell signalling involved in plant defence mechanisms. AB - Elicitins and lipid-transfer proteins are small cysteine-rich lipid-binding proteins secreted by oomycetes and plant cells, respectively, that share some structural and functional properties. In spite of intensive work on their structure and diversity at the protein and genetic levels, the precise biological roles of lipid-transfer proteins remains unclear, although the most recent data suggest a role in somatic embryogenesis, in the formation of protective surface layers and in defence against pathogens. By contrast, elicitins are known elicitors of plant defence, and recent work demonstrating that elicitins and lipid-transfer proteins share the same biological receptors gives a new perspective to understand the role played by lipid binding proteins, mainly the early recognition of intruders in plants. PMID- 12119166 TI - Pleiotropy, redundancy and the evolution of flowers. AB - Most angiosperm flowers are tightly integrated, functionally bisexual shoots that have carpels with enclosed ovules. Flowering plants evolved from within the gymnosperms, which lack this combination of innovations. Paradoxically, phylogenetic reconstructions suggest that the flowering plant lineage substantially pre-dates the evolution of flowers themselves. We provide a model based on known gene regulatory networks whereby positive selection on a single, partially redundant gene duplicate 'trapped' the ancestors of flower-bearing plants into the condensed, bisexual state approximately 130 million years ago. The LEAFY (LFY) gene of Arabidopsis encodes a master regulator that functions as the main conduit of environmental signals to the reproductive developmental program. We directly link the elimination of one LFY paralog, pleiotropically maintained in gymnosperms, to the sudden appearance of flowers in the fossil record. PMID- 12119167 TI - Mitogen-activated protein kinase cascades in plants: a new nomenclature. AB - Mitogen-activated protein kinase (MAPK) cascades are universal signal transduction modules in eukaryotes, including yeasts, animals and plants. These protein phosphorylation cascades link extracellular stimuli to a wide range of cellular responses. In plants, MAPK cascades are involved in responses to various biotic and abiotic stresses, hormones, cell division and developmental processes. Completion of the Arabidopsis genome-sequencing project has revealed the existence of 20 MAPKs, 10 MAPK kinases and 60 MAPK kinase kinases. Here, we propose a simplified nomenclature for Arabidopsis MAPKs and MAPK kinases that might also serve as a basis for standard annotation of these gene families in all plants. PMID- 12119168 TI - A long way ahead: understanding and engineering plant metal accumulation. AB - Some plants can hyperaccumulate metal ions that are toxic to virtually all other organisms at low dosages. This trait could be used to clean up metal-contaminated soils. Moreover, the accumulation of heavy metals by plants determines both the micronutrient content and the toxic metal content of our food. Complex interactions of transport and chelating activities control the rates of metal uptake and storage. In recent years, several key steps have been identified at the molecular level, enabling us to initiate transgenic approaches to engineer the transition metal content of plants. PMID- 12119170 TI - India joins the GM club. PMID- 12119169 TI - Impact of phyto-oxylipins in plant defense. AB - Phyto-oxylipins are metabolites produced in plants by the oxidative transformation of unsaturated fatty acids via a series of diverging metabolic pathways. Biochemical dissection and genetic approaches have provided compelling evidence that these oxygenated derivatives actively participate in plant defense mechanisms. During the past decade, interest in this field was focused on the biosynthesis of jasmonic acid (one branch of C18 polyunsaturated fatty acid metabolism) and on its relationship to the other plant defense-signaling pathways. However, recently, antisense strategies have revealed that oxylipins other than jasmonates are probably also essential for the resistance of plants to pathogens. PMID- 12119171 TI - Calcium oxalate crystal morphology. PMID- 12119172 TI - Exciting prospects for plants with greater disease resistance. PMID- 12119175 TI - Sodium butyrate stimulates the synthesis of firefly luciferase in transfected CHO cells but levels of BiP chaperone are unaffected. AB - A stably transfected CHO cell line (LUCLEAD) was used where the coding region of native Firefly luciferase was linked to the 3'-UTR of the bovine growth hormone, and the 5'-nucleotides coding for the albumin signal peptide were linked to the N terminal end of the luciferase coding region. Incubation of cells with 1 or 2 mM sodium butyrate (SB) for 72 h had no effect on cell growth since cultures reached confluency at the same time as control cells. Although cell cultures incubated with SB at a concentration of 4 mM were only about 60% confluent the luciferase content was about 5-fold higher than that in control cells. Cells incubated with either 1 or 2 mM SB showed intermediate levels of luciferase content. The amount of the chaperone BiP in the cells was not affected by incubation with SB. The results indicate that SB can be used to effectively promote synthesis of recombinant luciferase. PMID- 12119176 TI - Immunospecific protein of 34.5 kDa from DNA-protein cross-links induced by cis- and trans-diamminedichloroplatinum. AB - Cis-diamminedichloroplatinum(II) (cis-DDP) is one of the most often used anticancer drugs. It is generally accepted that the antitumor activity of the drug results from its interactions with DNA. Trans-diamminedichloroplatinum(II) (trans-DDP) also binds to DNA effectively, but is clinically ineffective. In the present work the lymphocyte nuclear proteins that participate in DNA-protein cross-links induced by cis- and trans-DDP are investigated. In lymphocytes which are incubated without platinum compounds there are DNA-binding proteins in the range of 45-71 kDa. It is shown that additional proteins of 28, 30, 34.5, 45 and 120 kDa are cross-linked with DNA in lymphocytes after 2-h incubation with cis DDP at concentrations of 0.1 and 0.5 mM. Trans-DDP does not bind additional proteins to DNA after the same incubation time. Electrophoretic analysis shows that trans-DDP binds much more of the same nuclear proteins to DNA than cis-DDP after 12-h incubation. In this study a test for the identification of 34.5 kDa protein is also undertaken. This protein appears in the samples obtained after 12 h incubation of lymphocytes with cis- and trans-DDP at 0.5 and 1 mM, especially. The protein of 34.5 kDa from cross-links induced by 1 mM trans-DDP is recognized by antibodies against the protein of the same molecular weight from the nuclear matrix of the lymphocytes. The results obtained here are discussed in relation to the biological activity of diamminedichloroplatinum isomers. PMID- 12119177 TI - Correlation between cell survival, clonogenic activity and micronuclei induction in DMBA-OC-1R cells treated with immunospecific albumin microspheres containing cisplatin and 5-fluorouracil. AB - An ideal chemotherapeutic strategy would be to deliver a high concentration of drug that would be released in sustained small amounts from targeted microspheres to effectively kill only the tumour cells and thus reduce toxicity to normal tissue. Clonogenic and cell survival growth curve assays, as well as the micronucleus assay, were used to determine the feasibility of employing targeted immunomicrospheres in the treatment of cancer. Cells of a rodent ovarian carcinoma cell line, were exposed to cisplatin and 5-fluorouracil, either as free drug or encapsulated in albumin microspheres that were either conjugated to monoclonal antibodies or not. In cell survival growth curve assays, cell survival was reduced to 1.2% of the control when cells were treated with drug-containing immunomicrospheres. 3.2-fold more micronuclei were found in those cells that had been exposed to the drugs in immunomicrospheres than in those subjected to untargeted microspheres. All three assays demonstrated that the targeted immunomicrospheres were more effective in delivering cisplatin and 5-fluorouracil directly to the cells than the unconjugated microspheres, thus suggesting that targeted chemotherapy might be a more effective option in the treatment of cancer. PMID- 12119178 TI - Ex vivo apoptotic potential of peripheral blood mononuclear cells of the elderly human subject. AB - Studies of immunosenescence have led to a detailed knowledge of immune system dysfunctions in the ageing human being. Apoptosis seems to be one of the process regulating an immune response after the antigenic stimulation. We examined whether commonly used methods of assessing apoptosis in the elderly human subject produce comparable results to young subjects. PBMC of young and elderly volunteers were isolated from the venous blood and cultured for 6 or 24 h with antigens of anti-influenza vaccine or PMA. The intensity of apoptosis was measured using the annexinV test, flow cytometric evaluation of DNA content (sub G1 peak in DNA histograms), 'ladder' by DNA gel electrophoresis, and fluorescence microscope. Apoptosis in 6 h-lasting cultures of the elderly was more intense in annexinV test, while it was decreased assessing subG1 peak. Additionally, in the aged group, those changes were associated with cell cycle arrest. Our results suggest that the apoptosis after the stimulation with the vaccine antigens seems to be some kind of activation-induced cell death (AICD). Different patterns of apoptosis after stimulation may be associated with the cell cycle arrest of the PBMC in the elderly. PMID- 12119179 TI - Organization of focal adhesion plaques is disrupted by action of the HIV-1 protease. AB - Focal adhesion plaques were severely affected in human embryonic fibroblasts permeabilized with digitonin and incubated in buffer containing the human immunodeficiency virus type 1 protease (HIV-1 PR). A mutant HIV-1 PR (3271 HIV-1 PR) had no effect on focal adhesion plaques. Similar effects were seen with cells microinjected with either HIV-1 PR or 3271 HIV-1 PR. Immunoblots of the human embryonic fibroblasts demonstrated that a number of focal adhesion plaque proteins were specifically cleaved by HIV-1 PR. These included fimbrin, focal adhesion plaque kinase (FAK), talin, and, to a lesser extent, filamin, spectrin and fibronectin. Proteins detected by antibodies to beta 4 integrin and alpha 3 integrin were also cleaved by the HIV-1 PR. Control experiments demonstrated that the effect and protein cleavages described are due to action of the HIV-1 PR and not to the action of endogenous host cell proteases. PMID- 12119180 TI - Examination of actin polymerization and viscosity induced by cations and ionic strength when cross-linked by alpha-actinin. AB - Increasing potassium chloride concentration from 0 to 100 mM and magnesium chloride from 0 to 2 mM show a parallel rate increase in polymerizing actin, whereas increasing calcium chloride concentration from 0 to 0.2 mM decreases the rate of polymerizing actin. The presence of alpha-actinin has little influence on the polymerization kinetics of actin under these conditions. Viscometric measurements indicate that the presence of various mono- and divalent cations, ionic strength, and alpha-actinin in combination are responsible for changes in the mechanical properties of solutions containing actin. The actin filament dynamic behavior is drastically reduced under these conditions as confirmed by quasi-elastic light scattering. PMID- 12119181 TI - DNA topoisomerase activities in Chinese hamster radiosensitive mutants after X ray treatment. AB - In the last years the attractive hypothesis of a possible involvement of mammalian topoisomerases in DNA repair has been proposed, given their molecular mechanism of action. So far, using asynchronous cultures a lot of controversial results have been reported, without taking into account the frequently dramatic fluctuations of topoisomerase activities depending upon the cell cycle stage and proliferation rate (mainly for topoisomerase II). We have addressed this question making use of G1 synchronous cultures of the Chinese hamster radiosensitive mutants xrs 5 (defective in DNA double strand breaks rejoining) and irs 2 (which shows radioresistant DNA synthesis), as well as their parental lines CHO K1 and V79 respectively, which show a normal radiosensitivity. Cells were irradiated with 5 Gy of X-rays and the activities of topoisomerases I and II in nuclear extracts were studied for comparison with non-irradiated controls in both the mutants and parental cell lines. Our results clearly show a modulation of the topoisomerase activities after irradiation, that varies depending upon the mutation that the different lines bear. While this hypothesis needs further testing, an interesting idea is that DNA topoisomerases might be involved in the cellular response to radiation damage, either through a direct participation in the repair mechanisms or in a preparative step to allow repair to proceed. PMID- 12119182 TI - Insulin-mediated tyrosine phosphorylation of myosin heavy chain and concomitant enhanced association of C-terminal SRC kinase during skeletal muscle differentiation. AB - In this study, we present for the first time: (1) evidence regarding tyrosine phosphorylation of myosin heavy chain, (2) evidence that insulin can phosphorylate myosin, (3) association of myosin with Csk, a signalling molecule, (4) modulation of this association by insulin, and (5) evidence that these interactions are associated with skeletal muscle differentiation. PMID- 12119188 TI - Apolipoprotein A-II, HDL metabolism and atherosclerosis. AB - Apolipoprotein (Apo) A-I and apo A-II are the major apolipoproteins of HDL. It is clearly demonstrated that there are inverse relationships between HDL-cholesterol and apo A-I plasma levels and the risk of coronary heart disease (CHD) in the general population. On the other hand, it is still not clearly demonstrated whether apo A-II plasma levels are associated with CHD risk. A recent prospective epidemiological (PRIME) study suggests that Lp A-I (HDL containing apo A-I but not apo A-II) and Lp A-I:A-II (HDL containing apo A-I and apo A-II) were both reduced in survivors of myocardial infarction, suggesting that both particles are risk markers of CHD. Apo A-II and Lp A-I:A-II plasma levels should be rather related to apo A-II production rate than to apo A-II catabolism. Mice transgenic for both human apo A-I and apo A-II are less protected against atherosclerosis development than mice transgenic for human apo A-I only, but the results of the effects of trangenesis of human apo A-II (in the absence of a co-transgenesis of human apo A-I) are controversial. It is highly suggested that HDL reduce CHD risk by promoting the transfer of peripherical free cholesterol to the liver through the so-called 'reverse cholesterol transfer'. Apo A-II modulates different steps of HDL metabolism and therefore probably alters reverse cholesterol transport. Nevertheless, some effects of apo A-II on intermediate HDL metabolism might improve reverse cholesterol transport and might reduce atherosclerosis development while some other effects might be deleterious. In different in vitro models of cell cultures, Lp A-I:A-II induce either a lower or a similar cellular cholesterol efflux (the first step of reverse cholesterol transport) than Lp A-I. Results depend on numerous factors such as cultured cell types and experimental conditions. Furthermore, the effects of apo A-II on HDL metabolism, beyond cellular cholesterol efflux, are also complex and controversial: apo A-II may inhibit lecithin-cholesterol acyltransferase (LCAT) (potential deleterious effect) and cholesteryl-ester-transfer protein (CETP) (potential beneficial effect) activities, but may increase the hepatic lipase (HL) activity (potential beneficial effect). Apo A-II may also inhibit the hepatic cholesteryl uptake from HDL (potential deleterious effect) probably through the SR-BI depending pathway. Therefore, in terms of atherogenesis, apo A-II alters the intermediate HDL metabolism in opposing ways by increasing (LCAT, SR-BI) or decreasing (HL, CETP) the atherogenicity of lipid metabolism. Effects of apo A-II on atherogenesis are controversial in humans and in transgenic animals and probably depend on the complex effects of apo A-II on these different intermediate metabolic steps which are in weak equilibrium with each other and which can be modified by both endogenous and environmental factors. It can be suggested that apo A-II is not a strong determinant of lipid metabolism, but is rather a modulator of reverse cholesterol transport. PMID- 12119189 TI - Sterol-regulatory element-binding protein (SREBP)-2 contributes to polygenic hypercholesterolaemia. AB - Sterol-regulatory element-binding protein (SREBP)-2 is a key regulator of cholesterol. When cells are deprived of cholesterol, proteolytic cleavage releases the NH(2)-terminal domain of SREBP-2 that binds and activates the promoters of SREBP-2-regulated genes including the genes encoding the low-density lipoprotein (LDL) receptor, 3-hydroxymethyl-3-glutaryl-(HMG-)CoA-synthase, and HMG-CoA-reductase. Thus, SREPB-2 gene activation leads to enhanced cholesterol uptake and biosynthesis. A novel protein polymorphism (SREBP-2-595A/G) discovered in the regulatory domain of human SREBP-2 was investigated regarding its impact on cholesterol homeostasis. In human embryonic kidney (HEK)-293-cells, the cleavage-rate of the SREBP-2-595A-isoform was slightly decreased compared to that of the SREBP-2-595G-isoform. Since cleavage of SREBP-2 activates the LDL receptor mediated uptake of plasma cholesterol, we hypothesized the LDL receptor-mediated uptake to be decreased in homozygous SREBP-2-595A-carriers and thus, plasma total cholesterol (TC) to be higher than in SREBP-2-595G-carriers. Multiple linear regression analysis of population samples from Switzerland (N=1334) and Israel (N=923) demonstrated a significant positive, gene dose-dependent association of the SREBP-2-595A-isoform with higher plasma TC (P=0.001). This cholesterol modulating effect was present in hypercholesterolaemic (DeltaTC=1.05 mmol/l, 14.4%; P=0.002; N=477), but absent in normocholesterolaemic subjects (DeltaTC=0.06 mmol/l, 1.4%; P=0.334; N=1780). In summary, a slightly but constantly decreased cleavage-rate of the SREBP-2-595A-isoform compared to that of the SREBP-2-595G-isoform may lead to a reduced transcriptional activation of the LDL receptor-gene weakening the SREBP-mediated compensation mechanisms, and may, therefore, be a critical factor in the development of polygenic hypercholesterolaemia. PMID- 12119190 TI - pH Heterogeneity of human and rabbit atherosclerotic plaques; a new insight into detection of vulnerable plaque. AB - BACKGROUND: Atherosclerotic plaques are heterogeneous with respect to inflammation, calcification, vascularity, oxygen, and temperature. We hypothesized that they also vary in pH and measured pH in living human carotid endarterectomized atherosclerotic plaques (CEA), Watanabe heritable hyperlipidemic (WHHL) rabbit aortas and human umbilical arteries (HUA). METHODS AND RESULTS: We measured pH of CEA of 48 patients, nine WHHL rabbit aortas and 11 HUA specimens (as controls) using a glass type microelectrode mounted on a micromanipulator in a 37 degrees C incubator. We also used single emission and also dual emission fluorescence ratio imaging microscopy employing pH-sensitive probes to confirm pH heterogeneity. Mean pH measured at 415 points of CEA was 7.55+/-0.32; at 275 points of WHHL rabbit aortas it was 7.40+/-0.43; and in 233 points of HUA it was 7.24+/-0.1. In CEA, pH of yellow (lipid-rich) areas was significantly lower than pH in calcified areas (7.15+/-0.01 vs. 7.73+/-0.01, P<0.0001). The coefficients of variation (heterogeneity) of pH in CEA, WHHL rabbit aortas, and HUA were 0.038+/-0.010, 0.039+/-0.007, and 0.009+/-0.003, respectively (P=0.0001). Fluorescence microscopic imaging confirmed pH heterogeneity in both humans and rabbits but not in HUA. In a variance components analysis 82% of the heterogeneity was due to the within-plaque variation and 2% was attributable to between-plaque variation. CONCLUSIONS: Our findings support the hypothesis of pH heterogeneity in plaques, and suggest a possible role for detecting low pH in the detection of plaque vulnerability. The source of pH heterogeneity particularly acidic pH, its impact on the stability of plaques and its potential clinical utility in locating vulnerable plaques remain to be evaluated. PMID- 12119191 TI - Effect of genetic background and diet on plasma fibrinogen in mice. Possible relation with susceptibility to atherosclerosis. AB - Many epidemiological studies suggest that elevated plasma fibrinogen concentrations form one of the most important independent risk factors in blood for cardiovascular disease and particularly atherosclerosis in humans. To clarify the effect of genetic factors, diets and their interactions on plasma fibrinogen concentrations, we examined plasma fibrinogen levels in four strains of mice, which differ in their susceptibility to cholesterol-induced atherosclerosis. When maintained on basal diet, two strains 129/J and C3H/HeJ exhibited a significantly higher plasma fibrinogen concentration (2.1 and 1.9 mg/ml) than C57BL/6J and BALB/C strains (1.5 and 1.4 mg/ml). The strongest and most rapid (1 week) increase of plasma fibrinogen (by all semi-synthetic diets) is observed in C57BL/6J mice, which are known to be highly susceptible to diet-induced atherosclerosis. After a period of 8 weeks an increase in plasma fibrinogen of approximately 30-50% was observed in all strains on all semi-synthetic diets. Remarkably, no increase was observed in the fibrinogen Aalpha- Bbeta- and gamma chain mRNA levels in the liver on the same diets. These mRNA levels were even decreased by approximately 20-50% in all strains on an extremely atherogenic diet. It was found that: genetic background determines the plasma fibrinogen levels on basal diet; plasma fibrinogen levels are altered by diet; the extent of these changes depends on the genetic background: surprisingly, this increase of fibrinogen in plasma is independent of transcription; the diet-induced increase of fibrinogen was very fast in the very highly atherosclerosis-susceptible strain C57BL/6J having a low basal fibrinogen level, and very slow in the atherosclerosis-resistant strain C3H/HeJ having a high basal fibrinogen level. It might be concluded that it is the kinetics of the response of fibrinogen to diet rather than the actual level, which relates to atherosclerosis susceptibility. PMID- 12119192 TI - Unsaturated fatty acids and their oxidation products stimulate CD36 gene expression in human macrophages. AB - Fatty acids (FA) have been implicated in the control of expression of several atherosclerosis-related genes. Similarly, the CD36 receptor has recently been shown to play an important role in atherosclerosis and other pathologies. The aim of the present study was to evaluate the direct effect of FA and their oxidation products (aldehydes), on the expression of CD36 in both THP-1 macrophages and human monocyte-derived macrophages (HMDM). The FA tested included the saturated FA (SFA) lauric, myristic, palmitic and stearic acid; the monounsaturated FA oleic acid; and the unsaturated FA (UFA) linoleic, arachidonic acid (AA), eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA). Aldehydes used were malondialdehyde (MDA), hexanal, 2,4-decadienal (DDE) and 4-hydroxynonenal (HNE). CD36 expression was measured by RT-PCR, Western blot and immunofluorescence. Incubation of THP-1 macrophages for 24 h with non-cytotoxic concentrations of UFA significantly increased CD36 mRNA expression. By contrast, exposure of THP-1 macrophages to SFA did not affect the levels of CD36 mRNA. Among all UFAs tested, EPA and DHA were the strongest inducers of CD36 mRNA levels, followed by oleic and linoleic acid. Incubation of HMDM with either oleic or linoleic acid significantly increased steady-state CD36 mRNA in a dose-dependent manner. Consistent with the increase of CD36 mRNA expression, incubation of THP-1 macrophages with oleic and linoleic acid for 24 h markedly increased CD36 protein expression. Treatment of THP-1 macrophages with MDA or hexanal for 24 h significantly increased CD36 mRNA expression in a dose dependent manner. In contrast, DDE and HNE significantly decreased this parameter. The data provide evidence for a direct regulatory effect of UFA on CD36 gene expression and support a role for aldehydes in the regulation of CD36 expression by FA. PMID- 12119193 TI - Lovastatin-stimulated superinduction of E-selectin, ICAM-1 and VCAM-1 in TNF alpha activated human vascular endothelial cells. AB - Inhibitors of HMG-CoA reductase (statins) reveal important pharmacological effects in addition to reducing the plasma LDL cholesterol level. In the pathogenesis of arteriosclerosis, transendothelial migration of various leukocytes including monocytes is a crucial step. We, therefore, investigated the expression of E-selectin, intercellular cell adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) in vascular endothelial cells as influenced by lovastatin. Human umbilical vein endothelial cells (HUVECs) express significant amounts of selectins and cell adhesion molecules (CAMs) within a few hours after stimulation with TNF-alpha. This effect is potentiated by 100-200% when the cells are pretreated with 0.1-2.5 microM lovastatin. The lovastatin mediated increase in the cytoplasm and at the cell surface is dose-dependent and significant at lovastatin concentrations comparable to plasma levels in patients under lovastatin treatment. The lovastatin-potentiated increase of E-selectin and CAMs is correlated with a corresponding increase of selectin- and CAM-specific mRNA. We conclude that, in vivo, statin treatment could trigger an enhanced recruitment of macrophages that might support the cholesteryl ester efflux from the arteriosclerotic plaque. PMID- 12119194 TI - Raman spectroscopic investigation of atorvastatin, amlodipine, and both on atherosclerotic plaque development in APOE*3 Leiden transgenic mice. AB - Raman spectroscopy allows quantitative, non-destructive evaluation of entire, intact atherosclerotic plaques. We quantified the anti-atherosclerotic effects of atorvastatin and amlodipine on progression of atherosclerosis using post-mortem Raman spectroscopic plaque imaging in 28 APOE*3 Leiden transgenic mice who were fed a high fat/high cholesterol diet for 28 weeks. Mice were assigned to a control group receiving the diet alone or to groups that received the diet with either 0.01% w/w atorvastatin, 0.002% w/w amlodipine, or the combination. The entire excised aortic arch was scanned with Raman microspectroscopy for quantitation of the distribution of cholesterol and calcification content. When mice had been treated with atorvastatin, cholesterol accumulation and calcification in the aortic arch was reduced by 91 and 98%, respectively, (both P<0.001). Amlodipine did not reduce the cholesterol content but reduced calcification of the aorta by 69% (P<0.05). The combination of amlodipine and atorvastatin was as effective as atorvastatin alone. This study demonstrates the strong atheroprotective potential of atorvastatin. In addition it is demonstrated that amlodipine reduces mineralization of atherosclerotic plaque. No synergistic effect of the combination of amlodipine and atorvastatin on plaque development is demonstrated. This study encourages Raman spectroscopic evaluations of anti atherosclerotic drugs in larger animals and humans in vivo. PMID- 12119195 TI - Reverse cholesterol transport in mice expressing simian cholesteryl ester transfer protein. AB - The role of cholesteryl ester transfer protein (CETP) in atherogenesis remains ambiguous, as both pro and antiatherogenic effects have been described. Expression of CETP increases HDL-cholesteryl ester turnover, but there is no direct evidence whether CETP mobilizes cholesterol in vivo. The rate of cholesterol removal injected into a leg muscle as cationized low density lipoprotein (cat-LDL) was compared in CETP transgenic and control mice. Four days after injection the exogenous cholesterol mass retained in muscle was 65% in CETP transgenic and 70% of injected dose in controls; it decreased to 52-54% by day 8 and negligible amounts remained on day 28. The cat-LDL was labeled with either 3H cholesterol oleate (3H-CE) or 3H-cholesteryl oleoyl ether (3H-COE), a nonhydrolyzable analog of 3H-CE. After injection of 3H-CE cat-LDL, clearance of 3H-cholesterol had a t(1/2) of 4 days between day 4 and 8 but there was little loss of 3H-COE between day 4 and 51. Liver radioactivity on day 4 was 1.7% in controls and 3.4% in CETP transgenics; it was 2.8 and 4.6%, respectively, on day 8. 3H-COE in liver accounted for 60% of label in CETP transgenics. In conclusion, high levels of plasma CETP in mice do not enhance reverse cholesterol transport in vivo but may act on extracellularly located cholesteryl ester. PMID- 12119196 TI - Hepatocyte growth factor is a survival factor for endothelial cells and is expressed in human atherosclerotic plaques. AB - Hepatocyte growth factor (HGF) has multiple effects on target cells upon activation of its receptor, c-Met. In endothelial cells, HGF induces migration, proliferation, and angiogenesis. HGF can also act as an anti-apoptotic factor for several cell types. The signal transduction pathways involved in mediating its anti-apoptotic effects have not been fully clarified. We demonstrated that HGF is anti-apoptotic for human endothelial cells, and identified the signaling pathways by which it mediates its effects. Human umbilical vein endothelial cells (HUVEC) exhibited significant levels of apoptosis after serum deprivation. HGF inhibited apoptosis in a dose dependent manner in serum-deprived cultures. HGF induced the phosphorylation of Akt and Erk1/2, cell survival factors, in a time dependent manner in serum deprived HUVEC. Inhibition of Akt and Erk1/2 activation abolished the anti-apoptotic effects of HGF. The transcription factor, NF-kappaB, can also play a role in promoting cell survival. However, NF-kappaB does not appear to contribute to the anti-apoptotic properties of HGF, as nuclear translocation of NF-kappaB was not detected in HGF-treated cultures. Endothelial cell migration, proliferation, and apoptosis contribute to the pathogenesis of atherosclerosis, and HGF may play a role in the development and progression of vascular lesions. Immunohistochemical analysis of human carotid artery sections demonstrated HGF protein localization within atherosclerotic lesions but not in normal vessels, suggesting that HGF may participate in atherogenesis. PMID- 12119197 TI - Carotid atherosclerosis, intima media thickness and risk factors--an analysis of 1781 asymptomatic subjects in Taiwan. AB - The aim of this study was to investigate the association of intima media thickness (IMT) and plaque with risk factors for atherosclerosis in asymptomatic subjects in Taiwan. Between 1998 and 2001, the study recruited 1781 asymptomatic subjects (1131 men and 650 women [mean age, 49 years; range 18-85 years]). These were examined by B-mode ultrasound to measure the IMT at the far wall of the common carotid artery (CCA) and the extent of plaque formation. A wide range of vascular risk factors including age, gender, smoking, body mass index, blood chemistry, and previous history were surveyed. The mean (+/-S.D.) IMT observed was 0.68 (+/-0.12) and 0.66 (+/-0.11) mm for men and women, respectively, (P=0.0008). The mean (S.D.) IMT of the CCA was 0.66 (+/-0.12) mm on the right side and 0.68 (+/-0.12) mm on the left side (P=0.0004). IMT increased with aging, according to the equation IMT=(0.005xage in years)+0.403 [corrected]. Higher IMT was associated with male gender, and IMT was greater in the left CCA. About 36.9% of subjects had carotid plaques. The percentage of plaque increased with aging. The plaque prevalence was positively associated with IMT. The value of IMT over the cut point of 0.68 mm correlated with obviously increased risk of carotid atherosclerosis. Age, systolic blood pressure and fasting blood sugar were independent risk factors related to both carotid atherosclerosis and thick IMT. PMID- 12119198 TI - Homocysteine levels in men and women of different ethnic and cultural background living in England. AB - This population-based cross-sectional study in South London looks at the total homocysteine (tHcy) levels in groups of different ethnic background and the possible role of environmental factors and the 677C-->T genetic polymorphism of the methylenetetrahydrofolate reductase (MTHFR). Fasting plasma tHcy was measured in 1392 men and women, age 40-59 years; 475 were white, 465 of African origin (of whom 180 were West Africans and 280 Caribbeans) and 452 South Asian (of whom 222 were Hindus and 167 Muslims). The homozygous MTHFR TT variant had observed frequencies of 0.10 in whites, 0.01 in people of African origin and 0.02 in South Asians (P<0.001). tHcy levels were 16% (95% CI 8-26) higher amongst TT than CC. tHcy levels were 25% (21-29) higher in men than women. Levels were significantly higher in South Asians than whites (8% [3-13]). Vegetarians had higher levels than non-vegetarians (25% [18-33]). These differences were present after adjustments for age, sex, smoking, body mass index (BMI), MTHFR 677C-->T polymorphism and socio-economic status. Compared with whites (10.0 [9.7-10.3] micromol/l), and allowing for confounders, Hindus had significantly higher levels of tHcy (12.1 [11.6-12.6] micromol/l). This difference was attenuated by the inclusion of vegetarianism in the model (11.3 [10.8-11.9] micromol/l). In contrast Muslims had similar tHcy levels to whites while both West Africans and Caribbeans had slightly lower levels, though differences were not significant. The reported higher levels of tHcy in South Asians are due to high levels amongst Hindus only. They are in part accounted for by their vegetarianism. These differences in tHcy are large enough to be important contributors to the risk of vascular disease and may be preventable by simple targeted population strategies. PMID- 12119199 TI - A candidate gene study in low HDL-cholesterol families provides evidence for the involvement of the APOA2 gene and the APOA1C3A4 gene cluster. AB - In patients with premature coronary heart disease, the most common lipoprotein abnormality is high-density lipoprotein (HDL) deficiency. To assess the genetic background of the low HDL-cholesterol trait, we performed a candidate gene study in 25 families with low HDL, collected from the genetically isolated population of Finland. We studied 21 genes encoding essential proteins involved in the HDL metabolism by genotyping intragenic and flanking markers for these genes. We found suggestive evidence for linkage in two candidate regions: Marker D1S2844, in the apolipoprotein A-II (APOA2) region, yielded a LOD score of 2.14 and marker D11S939 flanking the apolipoprotein A-I/C-III/A-IV gene cluster (APOA1C3A4) produced a LOD score of 1.69. Interestingly, we identified potential shared haplotypes in these two regions in a subset of low HDL families. These families also contributed to the obtained positive LOD scores, whereas the rest of the families produced negative LOD scores. None of the remaining candidate regions provided any evidence for linkage. Since only a limited number of loci were tested in this candidate gene study, these LOD scores suggest significant involvement of the APOA2 gene and the APOA1C3A4 gene cluster, or loci in their immediate vicinity, in the pathogenesis of low HDL. PMID- 12119200 TI - Impaired carotid and femoral viscoelastic properties and elevated intima-media thickness in peripheral vascular disease. AB - BACKGROUND: We undertook a study to determine whether peripheral vascular disease of the lower extremity (PVD) per se affects the arterial viscoelastic properties and intima-media thickness (IMT) of the carotid and femoral arteries. METHODS: Thirty-five patients with PVD, 35 age- and gender-matched control subjects were examined with ultrasound scan wall tracking system, with the simultaneous measurement of blood pressure for carotid and femoral IMT and viscoelastic properties. RESULTS: Subjects with PVD have significantly impaired carotid elastic properties including compliance (mean (SD): 6.50 (2.39) vs 9.93 (4.07) %mmHg(-1)x10(-2), P<0.001), Petersen's elastic modulus (1.77 (0.69) vs 1.19 (0.63) mmHg x 10(3), P=0.001) and stiffness index (17.92 (7.21) vs 12.10 (6.17), P=0.001) when compared to non-PVD controls. They also have significantly altered femoral elastic properties including Petersen's elastic modulus (5.94 (4.98) vs 3.64 (3.27) mmHg x 10(3), P=0.025) and stiffness index (58.42 (47.76) vs 36.96 (33.43), P=0.033). The carotid (0.85 (0.35) vs 0.59 (0.23) mm, P<0.001) and femoral (1.05 (0.39) vs 0.69 (0.31) mm, P<0.001) IMTs are also significantly elevated in PVD patients. After adjustment for the presumed cardiovascular load assessed on the basis of a cumulative total vascular risk score, as well as age, systolic and diastolic pressure, the carotid viscoelastic indices and the carotid and femoral IMTs remained highly significant. However, the difference in femoral elastic variables was no longer evident. CONCLUSION: PVD per se affects the femoral and carotid wall mechanics and morphology similarly to other cardiovascular risk factors and events. These parameters may provide further information for cardiovascular risk assessment in addition to the classical risk factors and the Framingham equation. Indeed, some guidelines have suggested that additional factors such as the carotid scan may influence the clinician's decision to intervene with therapy. PMID- 12119201 TI - Increased pregnancy loss in young women with aortoiliac disease. AB - BACKGROUND: During clinical evaluation of young women with peripheral arterial occlusive disease, we were surprised by the high prevalence of pregnancy loss in women with segmental stenosis confined to the aortoiliac segment. We wondered if increased occurrence of miscarriage is the result of high expression of vascular and obstetrical risk factors in these patients, or if it is related to localization of disease. In a case-control study designed to investigate risk factors for peripheral arterial occlusive disease in young women, we assessed the risk of miscarriage in these patients according to level of obstruction. METHODS: A total of 202 female patients, aged 18-49 years and 466 healthy control women from a population based case-control study, donated venous blood samples and filled out a structured questionnaire concerning classical cardiovascular risk factors and obstetrical history. In all patients, diagnosis of peripheral arterial occlusive disease was confirmed by intra-arterial angiography. Patients were classified into two groups: those with and those without stenosis of the aortoiliac segment (aortoiliac disease). RESULTS: In 77 of the 202 patients (38%) with peripheral arterial occlusive disease, the obstruction was confined to the aortoiliac segment. The occurrence of miscarriage was high (42%) in young women with aortoiliac disease. Compared to healthy controls, the risk of miscarriage increased 3-fold (OR 3.1; 95% CI 1.8-5.6) in these patients. Adjustment for obstetrical and vascular risk factors did not affect the risk estimate. CONCLUSION: This is the first study that identifies aortoiliac disease as a risk factor for pregnancy loss in young women. The risk of miscarriage is increased 3 fold in women with aortoiliac disease. The presence of vascular and obstetrical risk factors did not affect the strength of the association. Pregnancy loss could be the first sign of insufficient aortic circulation in these patients. PMID- 12119202 TI - Influence of atorvastatin and simvastatin on apolipoprotein B metabolism in moderate combined hyperlipidemic subjects with low VLDL and LDL fractional clearance rates. AB - Subjects with moderate combined hyperlipidemia (n=11) were assessed in an investigation of the effects of atorvastatin and simvastatin (both 40 mg per day) on apolipoprotein B (apoB) metabolism. The objective of the study was to examine the mechanism by which statins lower plasma triglyceride levels. Patients were studied on three occasions, in the basal state, after 8 weeks on atorvastatin or simvastatin and then again on the alternate treatment. Atorvastatin produced significantly greater reductions than simvastatin in low density lipoprotein (LDL) cholesterol (49.7 vs. 44.1% decrease on simvastatin) and plasma triglyceride (46.4 vs. 39.4% decrease on simvastatin). ApoB metabolism was followed using a tracer of deuterated leucine. Both drugs stimulated direct catabolism of large very low density lipoprotein (VLDL(1)) apoB (4.52+/-3.06 pools per day on atorvastatin; 5.48+/-4.76 pools per day on simvastatin versus 2.26+/-1.65 pools per day at baseline (both P<0.05)) and this was the basis of the 50% reduction in plasma VLDL(1) concentration; apoB production in this fraction was not significantly altered. On atorvastatin and simvastatin the fractional transfer rates (FTR) of VLDL(1) to VLDL(2) and of VLDL(2) to intermediate density lipoprotein (IDL) were increased significantly, in the latter instance nearly twofold. IDL apoB direct catabolism rose from 0.54+/-0.30 pools per day at baseline to 1.17+/-0.87 pools per day on atorvastatin and to 0.95+/-0.43 pools per day on simvastatin (both P<0.05). Similarly the fractional transfer rate for IDL to LDL conversion was enhanced 58-84% by statin treatment (P<0.01) LDL apoB fractional catabolic rate (FCR) which was low at baseline in these subjects (0.22+/-0.04 pools per day) increased to 0.44+/-0.11 pools per day on atorvastatin and 0.38+/-0.11 pools per day on simvastatin (both P<0.01). ApoB containing lipoproteins were more triglyceride-rich and contained less free cholesterol and cholesteryl ester on statin therapy. Further, patients on both treatments showed marked decreases in all LDL subfractions. In particular the concentration of small dense LDL (LDL-III) fell 64% on atorvastatin and 45% on simvastatin. We conclude that in patients with moderate combined hyperlipidemia who initially have a low FCR for VLDL and LDL apoB, the principal action of atorvastatin and simvastatin is to stimulate receptor-mediated catabolism across the spectrum of apoB-containing lipoproteins. This leads to a substantial, and approximately equivalent, percentage reduction in plasma triglyceride and LDL cholesterol. PMID- 12119204 TI - Positive Chlamydia pneumoniae serology is associated with elevated levels of tumor necrosis factor alpha in patients with coronary heart disease. AB - Infectious agents are possible stimulators of inflammation in atherogenesis. The aim of this study was to investigate if Chlamydia pneumoniae and Helicobacter pylori were associated with elevated levels of tumor necrosis factor alpha (TNFalpha) and interleukin-6 in coronary heart disease (CHD) patients (n=193) and age- and sex-matched controls (n=193) as markers of increased inflammatory activity. C reactive protein (CRP) and fibrinogen were also included. Serologic status towards the two bacteria was measured and levels of the inflammatory markers were compared between seropositives and seronegatives, each study group being evaluated separately. In CHD patients Chlamydia lipopolysaccharide (LPS) IgA seropositivity predicted elevated TNFalpha levels (P=0.009), still statistically significant after adjustment for traditional cardiovascular risk factors (P=0.005). Chlamydia LPS IgG seropositivity independently predicted fibrinogen levels in CHD patients (P=0.028), while no association between serology and inflammatory markers were observed among controls. H. pylori seropositivity alone was not associated with any increase in the inflammatory markers in any of the two groups. However, in CHD patients seropositivity to both agents predicted higher levels of TNFalpha (P=0.041), CRP (P=0.037) and fibrinogen (P=0.001) compared to double seronegativity. We conclude that C. pneumoniae LPS seropositivity may contribute to increased vascular inflammation in CHD patients, possibly even more pronounced when present in combination with H. pylori seropositivity. PMID- 12119203 TI - Ineffective decrease of serum cholesterol by simvastatin in a subgroup of hypercholesterolemic coronary patients. AB - We measured serum cholesterol concentrations and synthesis markers (e.g. serum lathosterol to cholesterol ratio), and absorption markers (e.g. serum campesterol to cholesterol ratio) of cholesterol in 319 good responders (GR; dose 20 mg up to 1 year) and in 115 poor responders (PR; dose increased at 6 weeks to 40 mg) among Finnish participants in the Scandinavian Simvastatin Survival Study at baseline, 6 weeks and 1 year of the simvastatin treatment. The baseline cholesterol level and the ratios of the absorption markers were higher and those of the synthesis markers lower in PR than GR. The ratios of the precursor sterols were negatively related to the baseline cholesterol in GR only (P=0.003). The cholesterol levels, and the ratios of the precursor sterols were decreased and those of the absorption marker sterols increased less consistently in PR than GR by 20 mg, the group differences being only slightly lessened by the dose addition to 40 mg. One year differences were still frequently significant. The baseline cholesterol concentrations were negatively related to the reduction of the precursor sterol ratios in GR, the change of cholesterol being positively related to those of the synthesis markers and negatively to those of the absorption markers only in PR. Thus, patients needing large statin dose for cholesterol normalization have high absorption and low synthesis of cholesterol, yet baseline synthesis is inversely related to cholesterol level only in GR. The synthesis rate is less markedly reduced by the large than by the small statin dose in the PR, and the reduction is related, in contrast to that in the GR, to lowering of cholesterol. PMID- 12119205 TI - Microalbuminuria and intima-media thickness of the carotid artery in clinically healthy men. AB - Microalbuminuria has been shown to be an independent predictor of cardiovascular disease in different populations. However, the underlying pathophysiological mechanism behind this observation is not known. The purpose of the present cross sectional study was to examine the relation of microalbuminuria to intima-media thickness (IMT) of the common carotid artery in a group of 368 clinically healthy 58-year old men. Urinary albumin excretion (UAE) and IMT of the common carotid artery were measured. Body mass index, WHR, systolic and diastolic blood pressure, heart rate, High-density lipoprotein (HDL) cholesterol and common carotid artery IMT were associated with UAE. A stepwise forward multiple regression showed that systolic blood pressure and WHR could explain 10.4% of the variability in log UAE (systolic blood pressure beta-coefficient 0.0047, SE 0.001, P<0.001; WHR beta-coefficient 0.93, SE 0.30, P=0.002). In conclusion, UAE was significantly associated with IMT of the common carotid artery in clinically healthy men. However, after adjustment for systolic blood pressure and WHR this association was not significant. We suggest that microalbuminuria in healthy subjects is not primarily associated with atherosclerosis but rather to blood pressure and abdominal obesity. PMID- 12119206 TI - Relationship between insulin-resistance and remnant-like particle cholesterol. AB - We investigated the relationship between insulin-resistance (IR) and remnant-like particle cholesterol (RLP-C) using 472 subjects (174 men and 298 women) randomly selected from inhabitants of two rural communities in Japan, Tanno and Sobetsu. The level of fasting immunoreactive insulin (FIRI), fasting blood glucose (FBS), total cholesterol (TC), triglyceride (TG), HDL cholesterol, LDL cholesterol, and RLP-C were measured in each subject. Homeostasis model assessment (HOMA-R) was used as an indicator of IR. The subjects were divided into two groups according to the value of HOMA-R: an IR group of subjects with HOMA-R > approximately equal to 1.73 and a normal (NR) group of subjects with HOMA-R <1.73. There was a significant positive correlation between HOMA-R and RLP-C. The value of RLP-C was higher in the IR group than in the NR group (7.1 vs. 3.9 mg/dl in men and 5.3 vs. 3.6 mg/dl in women). The frequency of hyper RLP cholesterolemia (RLP-C > approximately equal to 7.5 mg/dl) was higher in the IR than in the NR group (23.7 vs. 6.6% in men and 20.3 vs. 6.6% in women). The results of multiple regression analysis showed that HOMA-R was closely related to RLP-C. The results of this study suggest that RLP-C is closely associated with IR syndrome. PMID- 12119207 TI - Genetic variation in aldehyde dehydrogenase 2 and the effect of alcohol consumption on cholesterol levels. AB - Moderate drinkers with a defective alcohol dehydrogenase type 3 (ADH3) genotype have higher high-density lipoprotein (HDL) levels and a decreased risk of coronary artery disease (CAD). We examined the interaction between the aldehyde dehydrogenase type 2 (ALDH2), alcohol intake, and HDL levels in 826 men and 1295 women in a rural town in Japan. The ALDH2 genotype of each subject was determined by polymerase chain reaction (PCR) analysis. HDL was adjusted for the alcohol intake, age, body mass index, smoking status, total cholesterol, triglycerides and HbA1c levels. None of the subjects had a history or ECG suggestive of CAD. The proportions of ALDH2, *1/*1, *1/*2, and *2/*2 (defective homozygote) were 45.8, 46.0, and 8.2%, respectively, for men. Drinking more than two drinks daily was associated with lower HDL levels in men with the defective genotypes compared with men with a normal genotype (55.6+/-0.9 vs. 51.2+/-0.9 mg/dl, mean+/-S.E., P<0.0001). Also, drinking more than 0.5 drinks daily was not associated with beneficial effects on HDL levels in women with defective ALDH2 genotypes. CONCLUSIONS: Alcohol intake did not have beneficial effects on HDL levels in the defective ALDH2 genotype and may not protect against CAD in subjects with defective ALDH2 genotypes. PMID- 12119208 TI - Atorvastatin therapy in hypercholesterolemic patients suppresses cellular uptake of oxidized-LDL by differentiating monocytes. AB - Atherosclerosis is characterized by macrophage foam cells formation, which originate from differentiating blood monocytes that have taken up oxidized LDL (Ox-LDL) at enhanced rate. Statin therapy exhibit pleiotropic effects on many components of atherosclerosis. We have studied the effect of atorvastatin therapy in hypercholesterolemic patients, on the cellular uptake of Ox-LDL by their monocytes during differentiation into macrophages. Eleven hypercholesterolemic men were treated with 20 mg/day of atorvastatin for a period of 1 month. Peripheral blood monocytes harvested from control subjects and from patients before and after atorvastatin therapy were allowed to differentiate in culture for up to 9 days in the presence of 20% autologous serum. In control monocytes/macrophages the cellular uptake of Ox-LDL and the scavenger receptors CD36, SRA-I and SRA-II mRNA expression were upregulated during differentiation, and this upregulation was significantly enhanced in cells from hypercholesterolemic patients. Atorvastatin therapy suppressed the upregulation in Ox-LDL degradation and scavenger receptors expression in differentiating monocytes. These effects could be related at least in part to antioxidant characteristics of atorvastatin. Reduced susceptibility of plasma to free radical induced lipid peroxidation (by 35%), increased plasma total antioxidant status (TAS; by 30%), and increased serum paraoxonase activity (by 53%), were noted following drug therapy. We conclude that atorvastatin therapy in hypercholesterolemic patients reduces the enhanced cellular uptake of Ox-LDL during ex-vivo differentiation of monocytes into macrophages, and decreases cellular scavenger receptors gene expression. These effects may account for the attenuation of atherogenesis in hypercholesterolemic patients following atorvastatin treatment. PMID- 12119210 TI - One-year community-based education program for hypercholesterolemia in middle aged Japanese: a long-term outcome at 8-year follow-up. AB - To examine a long-term effect of community-based education program for hypercholesterolemia and an effect modification by apolipoprotein E polymorphism, we conducted a 1-year randomized clinical trial with 8 year-follow-up. One hundred four persons aged 40-64 years who had serum total cholesterol levels between 6.21 and 7.73 mmol/l (240 and 299 mg/dl) in 1988-89 cardiovascular risk surveys were enrolled in the trial. The intervention group (n=51, 82% for women) attended eight education classes in 1 year, while the control group (n=53, 85% for women) attended only two classes. Both groups were invited to the subsequent annual surveys. The mean serum cholesterol was 0.24-0.26 mmol/l less in the intervention than in the control group at both 6 month and 1 year (P=0.03, each) while the proportion of subjects using hypolipidemic agents was 0 and 6% in both groups, respectively. During 8-year follow-up, the probability of using hypolipidemic agents and/or total cholesterol > or =7.76 mmol/l was 51% in the education group and 69% in the control group; the risk ratio in the intervention vs control groups was 0.62 (95% CI: 0.36-1.06). When stratified by the apolipoprotein E polymorphism examined for 78% of the subjects, the risk ratio was 0.61 (0.31-1.18) among subjects without e4 allele (n=59) and 0.55 (0.14-2.14) among those with e4 allele (n=22). The intervention group had reduced intake of egg, fish egg, butter, mayonnaise and fatty meat compared to the control group at 6-month, 1- and 8-year follow-up. In conclusion, our community-based program was effective in reducing serum total cholesterol levels non-pharmacologically during the first year, and also reduced the likelihood of progressive worsening of hypercholesterolemia during the subsequent 8 years, regardless of the apolipoprotein E polymorphism. PMID- 12119209 TI - Regulation of small dense LDL concentration in Korean and Scottish men and women. AB - Small dense LDL is now emerging as an important risk factor for coronary artery disease. The amount of the LDL III has been reported to differ between ethnic groups. To investigate differences in the distribution of LDL subfractions between Korean and Scottish populations, we measured the plasma concentration and percent distribution of three major LDL subfractions in age-and sex-matched, middle aged, healthy 124 Korean and Scottish subjects (32 Korean men vs. 32 Scottish men; 30 Korean women vs. 30 Scottish women). Body mass index and waist circumference did not differ between the two ethnic groups. Total cholesterol and LDL cholesterol concentrations were higher in Scottish men compared with Korean men (P<0.01), while plasma triglyceride concentration was higher in Korean men and women (P<0.01 in men, P<0.05 in women). HDL cholesterol concentrations in both Korean men and women were lower than that of their Scottish counterparts (P<0.05 in men; P<0.001 in women). Korean men had lower concentrations of total LDL (242+/-65 vs. 325+/-122 mg/dl, P<0.01), LDL I (24+/-18 vs. 60+/-36 mg/dl, P<0.001) and LDL II (110+/-56 vs. 196+/-78 mg/dl, P<0.001). In contrast, LDL III concentration was markedly higher in Korean men (108+/-75 vs. 70+/-65 mg/dl, P<0.05). Likewise, the percent of LDL I (10.0+/-7.3 vs. 19.1+/-10.1%, P<0.001) and LDL II (47.2+/-20.7 vs. 60.1+/-10.9%, P<0.01) were lower in Korean men, while that of LDL III was higher (42.8+/-24.9 vs. 20.8+/-15.0%, P<0.001). In the female population, there were no differences in total LDL and LDL I concentrations between Korean and Scottish. LDL II concentration was lower in Korean women (106+/-53 vs. 151+/-57 mg/dl, P<0.01). Korean women showed a higher percent of LDL III (24.8+/-24.7 vs. 14.2+/-5.9%, P<0.05) and a lower LDL II (47.8+/-19.1 vs. 61.0+/-10.0%, P<0.01). Multiple linear regression revealed that plasma triglyceride concentration was the most important determinant of the LDL III subfraction concentration in Korean men and women and in Scottish men. In Korean men, the LDL III concentration rose linearly through the whole range of plasma triglyceride concentration, whereas in Scottish men, there was a threshold at 108 mg/dl triglyceride above which there was a positive association. Korean women showed the same pattern as Scottish men. We suggest that LDL concentrations and LDL subfraction distributions are regulated differently in these two ethnic groups. The different relationships between triglyceride and LDL III subfraction in Koreans versus Scots suggest that other factors, such as hepatic lipase or cholesteryl ester transfer protein may additionally play a role determining the LDL subfraction profile. PMID- 12119211 TI - Trisomy 4q syndrome: presentation of a new case and review of the literature. AB - We describe the 11th case of a de novo partial trisomy of the long arm of chromosome 4, with the extra segment spanning from 4q27 to 4q35. The aberration resulted from an unbalanced translocation of material from 4q to the short arm of chromosome 7, as evident from fluorescent in situ hybridization. Microsatellite analysis revealed the extra material to originate from the father. The karyotype was interpreted as 46,XX,der(7)t(4;7)(q27;p22). The patient is a 13-year-old girl with severe mental retardation, growth retardation, hearing impairment as well as minor foot, thumb and facial anomalies. Although the extent of the aberration varies between the reported patients, there are nevertheless features in common, suggestive of a trisomy 4q syndrome. The clinical findings most frequently reported are: mental retardation, seizures, microcephaly, hearing impairment and growth retardation, as well as epicanthic folds, high/broad/depressed nasal bridge, malformed ears, tooth and thumb anomalies. Almost the entire long arm of chromosome 4, except band q11, has been involved in trisomies/duplications, but 4q27 and 4q31 seem to be preferentially engaged in the trisomy 4q syndrome. PMID- 12119212 TI - Long-term follow-up of a girl with oro-facio-digital syndrome type I due to a mutation in the OFD 1 gene. AB - In 1954, Papillon-Leage and Psaume described a dominant, X-linked condition which they named oro-facio-digital (OFD). This condition was split into at least nine syndromes, the more common being OFD I. We report a girl with OFD I syndrome followed up for 23 years. Clinical examination showed cleft palate, median cleft lip, multiple oral frenulae, lobulated tongue and brachydactyly. There was no mental retardation. At 19 years of age, renal insufficiency appeared. A renal transplantation was performed. The parents were unaffected. An older brother had hydrocephaly, bilateral optic atrophy and mental retardation. A younger sister is unaffected. A mutation, an insertion of a G leading to a frameshift in the OFD 1 gene, was identified in this patient. PMID- 12119213 TI - True vs. false inv(Y)(p11q11.2): a familial instance concurrent with trisomy 21. AB - A boy with Down syndrome due to a free trisomy 21 also had a metacentric Y chromosome with an arm euchromatic and the other heterochromatic inherited from his phenotypically normal father. This chromosome was mitotically stable and hybridized with the DYZ3 probe precisely at its primary constriction; in addition, a subtelomeric Xp/Yp probe gave the expected signal near the end of the euchromatic arm. So, the proband's karyotype was 47,X,inv(Y)(p11q11.2),+21. Given the high frequency of both chromosome anomalies, we regard its concurrence as a mere coincidence. This observation, along with previous reports, allows us to classify the apparent pericentric inversions of the Y chromosome into two types: "true" inversions characterized by an alphoid single centromere and mitotic stability, and "false" inversions in which a nonalphoid centromere has taken over the usual alphoid centromere; indeed, these chromosomes are dicentric and mitotically unstable. Finally, the inv(Y) polymorphism in man compares with that documented in other mammal species, in which the rearranged Y chromosome neither impairs the fertility nor has other phenotypical consequences. PMID- 12119214 TI - A novel translocation, t(9;21)(q13;q22) rearranging the RUNX1 gene in acute myelomonocytic leukemia. AB - A novel translocation t(9;21)(q13;q22) associated with trisomy 4 has been detected in a patient with acute myelomonocytic leukemia (AML,M4) in relapse. The chromosomal translocation results in rearrangement of the RUNX1 gene at 21q22. The DNA sequence rearranged on chromosome 9 remains unidentified. The diversity of the partners involved in translocations implicating RUNX1 suggests that the functional consequences of the abnormality are more due to the truncation of RUNX1 than to the identity of its partner in the rearrangement. PMID- 12119215 TI - Genetic instability in human mismatch repair deficient cancers. AB - Cancers showing microsatellite instability (MSI-H) are frequent tumors characterized by inactivating alterations of mismatch repair (MMR) genes that lead to an incapacity to recognize and repair errors that occur during DNA replication. These cancers can be inherited as in the human non-polyposis colorectal cancer syndrome, or can occur sporadically in 10-15% of colorectal, gastric and endometrial cancers. MSI-H tumors have different clinicopathological features compared to cancers without this phenotype, termed MSS, and the repertoire of genetic events involved in tumoral progression of both phenotypes is thought to be different. In MSI-H tumors, most of the genetic changes occur at both non-coding and coding microsatellites that are particularly prone to errors during replication due to their repetitive sequence. This mechanism appears to be the main "genetic pathway" by which functional changes with putative oncogenic effects are accumulated in these tumors. PMID- 12119216 TI - French multi-centric study of 2000 amniotic fluid interphase FISH analyses from high-risk pregnancies and review of the literature. AB - This prospective and multi-centric study confirms the accuracy and the limitations of interphase FISH and shows that any cytogenetics laboratory can perform this technique. With regard to the technical approach, we think that slides must be examined by two investigators, because the scoring may be subjective. The main problem with the AneuVysion kit concerns the alpha satellite probes, and especially the chromosome 18 probe, which is sometimes very difficult to interpret because of the high variability of the size of the spots, and this may lead to false negative and uninformative cases. The best solution would be to replace these probes by locus-specific probes. Concerning clinical management, we offer interphase FISH only in very high-risk pregnancies or/and at late gestational age because of the cost of the test. We think that an aberrant FISH result can be used for a clinical decision when it is associated with a corresponding abnormal ultrasound scan. In other cases, most of the time, we prefer to wait for the standard karyotype. PMID- 12119217 TI - The psychopathological phenotype of velo-cardio-facial syndrome. AB - Velo-cardio-facial syndrome (VCFS) is mostly associated with deletions of chromosome 22q11, and is thought to be characterized by an increased frequency of major psychiatric disorders. Sixteen patients adults with VCFS and psychiatric symptoms were evaluated using a semi-structured investigation of history, symptoms, signs and behaviour. All available data were used in consensus meetings to obtain a classifiable diagnostic category. In contrast to other reports, no categorical diagnosis could be established. Instead, a quite specific psychological, behavioural and psychopathological constellation emerged that should most adequately be denominated as a VCFS-psychiatric syndrome. It is concluded that VCFS is associated with a specific psychopathological syndrome. PMID- 12119218 TI - The Peters' plus syndrome: a review. AB - Peters' plus syndrome is an infrequently described entity that combines anomalies in the anterior chamber of the eye with other multiple congenital anomalies, and a developmental delay. Major symptoms are extremely variable anterior chamber anomalies, cupid bow of the upper lip, cleft lip and palate, short stature, broad hands and feet, and variable mental delay. The syndrome follows an autosomal recessive pattern of inheritance. The etiology is unknown, but may involve abnormal neural crest development. A review of the pertinent literature is provided. PMID- 12119219 TI - Granulocyte macrophage-colony stimulating factor augmentation therapy in sepsis: is there a role? PMID- 12119220 TI - Inducible nitric oxide synthase in the tuberculous human lung. PMID- 12119221 TI - The paranasal sinuses and a unique role in airway nitric oxide production? PMID- 12119222 TI - Fluctuations and power laws in pulmonary physiology. PMID- 12119223 TI - A randomized phase II trial of granulocyte-macrophage colony-stimulating factor therapy in severe sepsis with respiratory dysfunction. AB - Granulocyte-macrophage colony-stimulating factor (GM-CSF) stimulates hemopoiesis and effector functions of granulocytes and macrophages and is involved in pulmonary surfactant homeostasis. We investigated whether GM-CSF therapy improved clinically diagnosed severe sepsis and respiratory dysfunction in critically ill patients. This randomized, double-blind, placebo-controlled phase II study added low-dose (3 mcg/kg) intravenous recombinant human GM-CSF daily for 5 days to conventional therapy in 10 patients, with a further eight patients receiving placebo. GM-CSF-treated patients showed improvement in Pa(O(2))/FI(O(2)) over 5 days (p = 0.02) and increased peripheral blood neutrophils (p = 0.08), whereas alveolar neutrophils decreased (p = 0.02). GM-CSF therapy was not associated with decreased 30-day survival or with increased acute respiratory distress syndrome or extrapulmonary organ dysfunction. GM-CSF therapy was associated with increased blood granulocyte superoxide production and restoration or preservation of blood and alveolar leukocyte phagocytic function. We conclude that low-dose GM-CSF was associated with improved gas exchange without pulmonary neutrophil infiltration, despite functional activation of both circulating neutrophils and pulmonary phagocytes. In addition, GM-CSF therapy was not associated with worsened acute respiratory distress syndrome or the multiple organ dysfunction syndrome, suggesting a homeostatic role for GM-CSF in sepsis-related pulmonary dysfunction. PMID- 12119224 TI - Humming greatly increases nasal nitric oxide. AB - The paranasal sinuses are major producers of nitric oxide (NO). We hypothesized that oscillating airflow produced by humming would enhance sinus ventilation and thereby increase nasal NO levels. Ten healthy subjects took part in the study. Nasal NO was measured with a chemiluminescence technique during humming and quiet single-breath exhalations at a fixed flow rate. NO increased 15-fold during humming compared with quiet exhalation. In a two-compartment model of the nose and sinus, oscillating airflow caused a dramatic increase in gas exchange between the cavities. Obstruction of the sinus ostium is a central event in the pathogenesis of sinusitis. Nasal NO measurements during humming may be a useful noninvasive test of sinus NO production and ostial patency. In addition, any therapeutic effects of the improved sinus ventilation caused by humming should be investigated. PMID- 12119225 TI - The effects of intranasal budesonide on allergen-induced production of interleukin-5 and eotaxin, airways, blood, and bone marrow eosinophilia, and eosinophil progenitor expansion in sensitized mice. AB - We have previously demonstrated that allergen inhalation induces expansion of bone marrow eosinophil progenitors in sensitized mice and subjects with asthma and that the inhaled corticosteroid, budesonide, reduced baseline but not allergen-induced increase in bone marrow eosinophil/basophil progenitors (EoB CFU) in subjects with asthma. Here, we evaluated the effects of intranasal budesonide on allergen-induced increases in interleukin (IL)-5 and eotaxin in the airway and peripheral blood, expansion of bone marrow Eo-CFU and eosinophilia in bone marrow, peripheral blood and airway, as well as airway hyperresponsiveness, in ovalbumin (OVA)-sensitized mice. Budesonide treatment attenuated allergen induced eosinophilia in bone marrow, peripheral blood, and airways as well as allergen-induced increases in bone marrow eosinophil progenitors but not allergen induced increases in IL-5 or eotaxin 12 h following the second of two daily exposures to allergen; at later time points treatment was associated with attenuation of IL-5, eosinophilia, Eo-CFU, and airway hyperresponsiveness. These results suggest that a component of the mechanism by which corticosteroid treatment attenuates allergen-induced airway inflammation is through suppression of bone marrow eosinophilopoiesis, and that this is likely not mediated simply through the blocking of IL-5 production at the airway. PMID- 12119226 TI - Surfactant synthesis and kinetics in infants with congenital diaphragmatic hernia. AB - In animal models of congenital diaphragmatic hernia (CDH), surfactant deficiency contributes to the pathophysiology of the disease; however, information on CDH in humans is limited. We compared surfactant disaturated phosphatidylcholine (DSPC) synthesis and metabolism, by stable isotope technology, in newborn infants with CDH and in control subjects. DSPC amount, total proteins, and surfactant protein A (SP-A) from tracheal aspirates were also measured. DSPC and SP-A were significantly lower in 14 infants with CDH than in the eight control subjects. Mean DSPC was 2.3 +/- 1.3 mg/ml of epithelial lining fluid (ELF) in infants with CDH and 4.6 +/- 1.5 mg/ml of ELF in control subjects (p = 0.001). Mean SP-A in infants with CDH and in control subjects was 16.2 +/- 9.3 and 61.2 +/- 30.6 microg/ml of ELF, respectively (p = 0.03). DSPC kinetics was measured in 12 of 14 infants with CDH and in 5 of 8 control subjects. Secretion time was 8.3 +/- 5.5 and 8.5 +/- 2.5 hours and peak time 51.9 +/- 15.2 and 51 +/- 13 hours in infants with CDH and in control subjects, respectively. Fractional synthesis rate was not different for infants with CDH and control subjects (p = 0.4). In conclusion, surfactant DSPC synthesis and kinetics were not significantly deranged in infants with CDH compared with control subjects. Other factors, such as lower surface area or increased DSPC catabolism, may contribute to surfactant pool alteration in CDH. PMID- 12119227 TI - Increased incidence of cardiovascular disease in middle-aged men with obstructive sleep apnea: a 7-year follow-up. AB - The incidence of a cardiovascular disease (CVD) was explored in a consecutive sleep clinic cohort of 182 middle-aged men (mean age, 46.8 +/- 9.3; range, 30-69 years in 1991) with or without obstructive sleep apnea (OSA). All subjects were free of hypertension or other CVD, pulmonary disease, diabetes mellitus, psychiatric disorder, alcohol dependency, as well as malignancy at baseline. Data were collected via the Swedish Hospital Discharge Register covering a 7-year period before December 31, 1998, as well as questionnaires. Effectiveness of OSA treatment initiated during the period as well as age, body mass index (BMI), systolic blood pressure (SBP), diastolic blood pressure (DBP) at baseline, and smoking habits were controlled. The incidence of at least one CVD was observed in 22 of 60 (36.7%) cases with OSA (overnight oxygen desaturations of 30 or more) compared with in 8 of 122 (6.6%) subjects without OSA (p < 0.001). In a multiple logistic regression model, significant predictors of CVD incidence were OSA at baseline (odds ratio [OR] 4.9; 95% confidence interval [CI], 1.8-13.6) and age (OR 23.4; 95% CI, 2.7-197.5) after adjustment for BMI, SBP, and DBP at baseline. In the OSA group, CVD incidence was observed in 21 of 37 (56.8%) incompletely treated cases compared with in 1 of 15 (6.7%) efficiently treated subjects (p < 0.001). In a multiple regression analysis, efficient treatment was associated with a significant risk reduction for CVD incidence (OR 0.1; 95% CI, 0.0-0.7) after adjustment for age and SBP at baseline in the OSA subjects. We conclude that the risk of developing CVD is increased in middle-aged OSA subjects independently of age, BMI, SBP, DBP, and smoking. Furthermore, efficient treatment of OSA reduces the excess CVD risk and may be considered also in relatively mild OSA without regard to daytime sleepiness. PMID- 12119228 TI - Early life factors contribute to the decrease in lung function between ages 18 and 40: the Coronary Artery Risk Development in Young Adults study. AB - Early life factors may influence pulmonary function. We measured forced expiratory volume in 1 second (FEV(1)) in 1985-1986 and 2, 5, and 10 years later in approximately 4,000 black and white men and women initially aged 18-30 years. We estimated the age pattern of FEV(1) according to family smoking status, early diagnosis of asthma, early smoking initiation, adult asthma, and cigarette smoking. FEV(1) followed a quadratic pattern from age of peak through age 40. The pattern varied by race and sex. Early smoking initiation was associated with a faster decrease in FEV(1). Smoking by family members was related to early life asthma and may have contributed to faster FEV(1) decrease by encouraging behaviors such as heavier smoking or earlier smoking initiation. Prevalence of smoking was 28% when no family member smoked, compared with 59% when four or more members smoked. The FEV(1) decline was 8.5% in never-smokers without asthma; 10.1% in nonsmoking individuals diagnosed with asthma; and 11.1% in baseline smokers who smoked 15 or more cigarettes per day. The combination of asthma and heavier smoking was synergistic (17.8% decline). This study delineates an increased rate of decline in those with asthma or in those who smoke cigarettes and implicates early life exposures as contributing to the faster rate of FEV(1) decline. PMID- 12119229 TI - The relationship between individual histologic features and disease progression in idiopathic pulmonary fibrosis. AB - We have retrospectively studied 53 patients with idiopathic pulmonary fibrosis and a histologic diagnosis of usual interstitial pneumonia and evaluated the prognostic significance of four individual histologic features (fibroblastic foci [FF], interstitial mononuclear cell infiltrate, established fibrosis, and intra alveolar macrophages) using a semiquantitative scale of 0-6. An objective count of FF was also undertaken. Using weighted kappa coefficients, interobserver agreement between pathologists was moderate to good (0.56-0.76). Subjective and objective FF scores were strongly associated (R(S) = 0.88, < 0.00005). Mortality was independently linked to a high FF score, p = 0.006, and a low percent predicted carbon monoxide diffusing capacity (DL(CO)), p = 0.01. For pulmonary function, on univariate analysis, the strongest correlations were observed between increasing interstitial mononuclear cell infiltrate or FF scores and greater declines in forced vital capacity (FVC) or DL(CO) at 6 months. Multivariate models revealed that increasing FF scores were independently associated with greater declines in FVC and DL(CO) at both 6 and 12 months. Increasing interstitial mononuclear cell infiltrate scores were also independently linked to functional decline, but only at 6 months. These data suggest a reproducible method on biopsy for predicting rate of disease progression in patients with idiopathic pulmonary fibrosis. PMID- 12119231 TI - Expression of human telomerase reverse transcriptase in lymphangioleiomyomatosis. AB - Telomerase synthesizes nucleotide hexameric repeats (telomeres) at the ends of chromosomes, replacing base sequences that are lost from these sites during each mitotic cycle and protecting these ends against the action of exonucleases and ligases. Therefore, telomerase is essential for maintaining cellular replication. To evaluate the role of telomerase in the proliferation of abnormal smooth muscle cells (lymphangioleiomyomatosis [LAM] cells) in LAM, we performed immunostaining and in situ hybridization studies to identify telomerase protein and messenger RNA (mRNA), respectively, in pulmonary (n = 18) and extrapulmonary (n = 4) lesions from 22 women with LAM (14 untreated and 8 treated with progesterone or tamoxifen). Immunoreactivity and hybridization signals for telomerase were observed in 5 to 20% of LAM cells, mostly of the spindle-shaped type, in 21 of the 22 patients, and were less intense in the treated group. Other types of cells were unreactive in both groups. Telomerase colocalized in the same cells with alpha-smooth muscle actin, but only rarely with HMB-45 antibody (a marker for epithelioid LAM cells); colocalization with proliferating cell nuclear antigen was incomplete. The telomerase-positive LAM cells may constitute the sources of renewal of LAM cells. Modulation of telomerase may be involved in the control of LAM cell proliferation. PMID- 12119230 TI - Analysis of nitric oxide synthase and nitrotyrosine expression in human pulmonary tuberculosis. AB - The role of nitric oxide (NO) in the host-defense against human tuberculosis (TB) is controversial. Although experimental evidence indicates that NO may play an important role in controlling TB, its expression in human tuberculous lungs has not been systematically characterized. We therefore investigated the expression of NO synthases (NOS) and of nitrotyrosine, the latter a marker of NO expression, in surgically resected lungs of eight patients with TB. Immunohistochemical and morphometric analyses revealed that, compared with control subjects, inducible NOS, endothelial NOS, and nitrotyrosine, but not neuronal NOS, were significantly elevated in the inflammatory zone of the tuberculous granulomas, and in the nongranulomatous pneumonitis zone. Tumor necrosis factor-alpha (TNF-alpha) was also significantly increased in tuberculous lungs and was principally localized to the necrotic, and to a lesser extent, the inflammatory and fibrotic areas of the granulomas. The NOS isoforms, nitrotyrosine, and TNF-alpha were expressed by the epithelioid macrophages and giant cells within the granulomas and in alveolar macrophages and epithelial cells in pneumonitis areas. This descriptive study provides evidence that in human TB, NOS isoenzymes and NO are present in specialized areas of the tuberculous granulomas; their precise role in human TB remains to be determined. PMID- 12119232 TI - Protection of human lung cells against hyperoxia using the DNA base excision repair genes hOgg1 and Fpg. AB - Hyperoxia causes pulmonary toxicity in part by injuring alveolar epithelial cells. Previous studies have shown that toxic oxygen-derived species damage DNA and this damage is recognized and repaired by either human enzyme 8-oxoguanine DNA glycosylase (hOgg1) or Escherichia coli enzyme formamidopyrimidine DNA glycosylase (Fpg). To determine whether these DNA repair proteins can reduce O(2) mediated DNA damage in lung cells, A549 lung epithelial cells were transduced with either hOgg1 or Fpg using a retroviral vector containing enhanced green fluorescent protein. Expression of each gene in the transduced cells was confirmed by fluorescent microscopy, Northern blotting, Western blotting, and an enzymatic oligonucleotide cleavage assay. A549 cells expressing either hOgg1 or Fpg were protected from hyperoxia as evidenced by a decrease in DNA damage and a corresponding increase in cell survival. Further, we determined that overexpression of hOgg1 or Fpg partially mitigated the toxic effects of hydrogen peroxide in lung cells. Our data suggest that increased expression of DNA base excision repair genes might represent a new approach for protecting critical lung cells from the toxic effects of hyperoxia. PMID- 12119233 TI - Inflammatory mediator mRNA expression by adenovirus E1A-transfected bronchial epithelial cells. AB - Lung tissue from patients with emphysema and airway obstruction carries excess adenoviral E1A DNA that is expressed as protein in airway surface epithelium and is associated with an increased inflammatory response. To examine mechanisms by which latent adenoviral infection might amplify the inflammatory process, we transfected primary human bronchial epithelial (HBE) cells from three separate patients undergoing lung resection so that they stably expressed adenovirus E1A. Lipopolysaccharide stimulation of the E1A-transfected HBE cells increased intercellular adhesion molecule-1 and interleukin-8 mRNA and protein expression compared with control cells from the same patient. It also induced greater intercellular adhesion molecule-1 promoter activity and greater nuclear factor kappa B binding activity of nuclear extracts in E1A transfectants than controls. E1A-positive transfectants constitutively expressed transforming growth factor beta 1 mRNA and protein, whereas this expression was either very low or not detected in control cells. We conclude that adenoviral E1A transfection transforms primary HBE cells and upregulates their production of mediators that are clinically relevant to the pathogenesis of chronic obstructive pulmonary disease. PMID- 12119234 TI - Thromboxane A(2) receptors mediate pulmonary hypertension in 60% oxygen-exposed newborn rats by a cyclooxygenase-independent mechanism. AB - Endothelin-1 (ET-1) mediates the development of pulmonary hypertension (PHT) in newborn rats exposed to 60% O(2) for 14 days, a model for human chronic neonatal lung injury. ET-1 production by d-14 rat pulmonary artery smooth muscle cells in vitro was markedly increased by thromboxane (TX) A(2) receptor agonists and inhibited by a competitive antagonist. We hypothesized that stimulation of the TX A(2) receptor contributed to O(2)-mediated PHT in vivo. Newborn rat pups received daily intraperitoneal injections of L670596, a competitive TX A(2) receptor antagonist, or 5,5-dimethyl-3-(3-fluorophenyl)4-(4-methylsulfonyl)phenyl-2(5H) furanone (DFU), a cyclooxygenase-2 inhibitor, during 14 days of 60% O(2) or air exposure. L670596, but not DFU, prevented 60% O(2)-mediated right ventricular and small pulmonary vessel smooth muscle hypertrophy. Lung ET-1 content was significantly reduced by L670596 in 60% O(2)-exposed animals. We conclude that TX A(2) receptor activation, though not by TX A(2), caused upregulation of ET-1 and PHT in this model. A likely mediator is the stable lipid peroxidation product, 8 iso-prostane, which acts as an incidental ligand of the TX A(2) receptor and is a potent inducer of ET-1 production by cultured d-14 rat pulmonary artery smooth muscle cells in vitro. PMID- 12119235 TI - Pulmonary alveolar proteinosis: progress in the first 44 years. AB - Pulmonary alveolar proteinosis is a rare clinical syndrome that was first described in 1958. Subsequently, over 240 case reports and small series have described at least 410 cases in the literature. Characterized by the alveolar accumulation of surfactant components with minimal interstitial inflammation or fibrosis, pulmonary alveolar proteinosis has a variable clinical course ranging from spontaneous resolution to death with pneumonia or respiratory failure. The most effective proven treatment--whole lung lavage--was described soon after the first recognition of this disease. In the last 8 years, there has been rapid progress toward elucidation of the molecular mechanisms underlying both the congenital and acquired forms of pulmonary alveolar proteinosis, following serendipitous discoveries in gene-targeted mice lacking granulocyte-macrophage colony-stimulating factor (GM-CSF). Impairment of surfactant clearance by alveolar macrophages as a result of inhibition of the action of GM-CSF by blocking autoantibodies may underlie many acquired cases, whereas congenital disease is most commonly attributable to mutations in surfactant protein genes but may also be caused by GM-CSF receptor defects. Therapy with GM-CSF has shown promise in approximately half of those acquired cases treated, but it is unsuccessful in congenital forms of the disease, consistent with the known differences in disease pathogenesis. PMID- 12119236 TI - Future research directions in idiopathic pulmonary fibrosis: summary of a National Heart, Lung, and Blood Institute working group. AB - Idiopathic pulmonary fibrosis (IPF) is an insidious inflammatory fibroproliferative disease whose cause and course before diagnosis are unknown, and for which existing treatments are of limited benefit. The National Heart, Lung, and Blood Institute convened a working group to develop specific recommendations for future IPF research. Inflammatory and immune processes are involved in IPF pathogenesis, and current therapeutic strategies are aimed at suppressing the inflammation. Recent data suggest that the molecular processes underlying the fibrogenesis may provide new opportunities for therapeutic intervention. Specific areas of future research recommended by the working group include studies to elucidate the etiology of IPF, to develop novel diagnostic techniques and molecular diagnostics, to establish a program for identification of molecular targets for IPF treatment and identification and generation of agonists or antagonists that inhibit fibrogenesis, to foster investigations that couple the use of new technologies (e.g., laser capture microdissection, microarrays, and mass spectroscopic analysis of proteins) with data from the human genome project, to establish a national consortium of Clinical Centers of Excellence to conduct coordinated clinical and laboratory studies of well characterized patients and patient-derived materials, and to stimulate research to develop animal models of persistent and progressive pulmonary fibrosis for evaluation of new intervention approaches. PMID- 12119238 TI - Intramural hematoma in acute aortic syndrome: more than one variant of dissection? PMID- 12119239 TI - Metabolic syndrome: pathophysiology and implications for management of cardiovascular disease. PMID- 12119240 TI - Fish-rich diet, leptin, and body mass. AB - BACKGROUND: Leptin has been implicated in cardiovascular disease. A diet rich in fish has been associated with decreased cardiac and vascular risk. METHODS AND RESULTS: We examined the relationship between diet and leptin in 2 related homogeneous African tribal populations of Tanzania. One tribe consumes freshwater fish as their main diet component (n=279), and the other tribe consumes a primarily vegetarian diet (n=329). In multivariate analysis, plasma leptin levels were associated with type of diet (F=14.3, P<0.001), independent of age, body mass index, body fat, alcohol consumption, or insulin. Both male (2.5+/-2 [fish diet] versus 11.2+/-2.4 [vegetarian diet] ng/mL, P=0.017) and female (5.0+/-1.9 [fish diet] versus 11.8+/-1.4 [vegetarian diet] ng/mL, P=0.007) fish eaters had lower plasma leptin levels than did their vegetable diet counterparts, even though body mass index values were virtually identical. CONCLUSIONS: A diet rich in fish is associated with lower plasma leptin, independent of body fat. These findings may have implications for understanding the reduced cardiovascular risk in subjects on a high-fish diet. PMID- 12119242 TI - Three-dimensional black-blood cardiac magnetic resonance coronary vessel wall imaging detects positive arterial remodeling in patients with nonsignificant coronary artery disease. AB - BACKGROUND: Direct noninvasive visualization of the coronary vessel wall may enhance risk stratification by quantifying subclinical coronary atherosclerotic plaque burden. We sought to evaluate high-resolution black-blood 3D cardiovascular magnetic resonance (CMR) imaging for in vivo visualization of the proximal coronary artery vessel wall. METHODS AND RESULTS: Twelve adult subjects, including 6 clinically healthy subjects and 6 patients with nonsignificant coronary artery disease (10% to 50% x-ray angiographic diameter reduction) were studied with the use of a commercial 1.5 Tesla CMR scanner. Free-breathing 3D coronary vessel wall imaging was performed along the major axis of the right coronary artery with isotropic spatial resolution (1.0x1.0x1.0 mm(3)) with the use of a black-blood spiral image acquisition. The proximal vessel wall thickness and luminal diameter were objectively determined with an automated edge detection tool. The 3D CMR vessel wall scans allowed for visualization of the contiguous proximal right coronary artery in all subjects. Both mean vessel wall thickness (1.7+/-0.3 versus 1.0+/-0.2 mm) and wall area (25.4+/-6.9 versus 11.5+/-5.2 mm(2)) were significantly increased in the patients compared with the healthy subjects (both P<0.01). The lumen diameter (3.6+/-0.7 versus 3.4+/-0.5 mm, P=0.47) and lumen area (8.9+/-3.4 versus 7.9+/-3.5 mm(2), P=0.47) were similar in both groups. CONCLUSIONS: Free-breathing 3D black-blood coronary CMR with isotropic resolution identified an increased coronary vessel wall thickness with preservation of lumen size in patients with nonsignificant coronary artery disease, consistent with a "Glagov-type" outward arterial remodeling. This novel approach has the potential to quantify subclinical disease. PMID- 12119241 TI - Is the pregnancy hormone relaxin also a vasodilator peptide secreted by the heart? AB - BACKGROUND: It has been shown recently that the pregnancy and parturition hormone, relaxin, is secreted by the heart. This study examined the effects of relaxin in small human resistance arteries from the systemic and pulmonary circulations. METHODS AND RESULTS: Arteries were obtained from gluteal biopsies and resected lung tissue and studied with the use of wire myography. Cumulative concentration relaxation curves were constructed in systemic arteries with substance P, epoprostenol, atrial natriuretic peptide, and relaxin (concentration range 10(-13) -10(-7)M). The maximal responses were 88(+/-5)%, 67(+/-10)%, 52(+/ 16)% and 66(+/-16)%, respectively. Endothelium removal virtually abolished the action of relaxin. Relaxin had no vasodilator effect in pulmonary arteries. CONCLUSIONS: Relaxin is a powerful dilator of systemic resistance arteries secreted by the heart that may contribute to cardiovascular regulation. PMID- 12119243 TI - Impact of a specific echocardiographic report comment regarding endocarditis prophylaxis on compliance with American Heart Association recommendations. AB - BACKGROUND: There is substantial underuse and overuse of antibiotic prophylaxis. Current American Heart Association (AHA) recommendations are heavily dependent on echocardiographic data that may not be familiar to referring physicians. We sought to determine the impact of a specific prophylaxis report comment on compliance with AHA recommendations. METHODS AND RESULTS: Using a standardized electronic reporting system, physicians interpreting approximately 50% of echocardiograms included a concluding comment classifying endocarditis risk and stating whether prophylaxis was indicated. The remaining reports were identical in format/content but did not include such comments. Of 1461 eligible outpatients during a 6-month period, 969 (66.3%) responded to a mail survey regarding prophylaxis instructions. Overall, 50% reported taking prophylaxes. Compliance with AHA recommendations was greater among those with a comment (73.2% versus 65.4% for those receiving versus those not receiving comments, respectively; P=0.011), with particular improvement among moderate-risk patients (69.5% versus 59.9%, P=0.024). CONCLUSIONS: An echocardiographic report statement regarding endocarditis risk and need for prophylaxis is a simple low-cost intervention that improves compliance with AHA recommendations. Interpreting physicians should become familiar with AHA recommendations and include a concluding statement addressing prophylaxis. PMID- 12119244 TI - Heritability of coronary artery calcium quantity measured by electron beam computed tomography in asymptomatic adults. AB - BACKGROUND: Electron beam computed tomography is an accurate, noninvasive method to detect and quantify coronary artery calcification (CAC), a marker of subclinical and clinical coronary artery atherosclerosis. CAC quantity predicts future coronary artery disease end points in asymptomatic adults, but measured risk factors explain less than half the variability in CAC quantity. Although several candidate genes for CAC have been identified, the relative importance of genetic influences on CAC quantity has not been assessed in asymptomatic adults in a community. METHODS AND RESULTS: We quantified the relative contributions of measured risk factors and genetic influences on CAC quantity measured by electron beam computed tomography in 698 asymptomatic white adults from 302 families. Before adjusting for any risk factors, 43.5% of the variation in CAC quantity was attributable to genetic factors (P=0.0007). Independent predictors of CAC quantity were identified with stepwise linear regression. After adjusting for these risk factors, including age, sex, fasting glucose level, systolic blood pressure, pack-years of smoking, and LDL cholesterol, 41.8% of the residual variation in CAC quantity was attributable to genetic factors (P=0.0003). CONCLUSIONS: These results demonstrate the importance of genetic factors in subclinical coronary atherosclerosis variation as measured by CAC quantity. The presence of genetic effects suggests that unknown genes that influence CAC quantity are yet to be identified. PMID- 12119245 TI - Sustained ventricular arrhythmias among patients with acute coronary syndromes with no ST-segment elevation: incidence, predictors, and outcomes. AB - BACKGROUND: The prognosis of ventricular arrhythmias among patients with non-ST elevation acute coronary syndromes is unknown. We studied the incidence, predictors, and outcomes of sustained ventricular arrhythmias in 4 large randomized trials of such patients. METHODS AND RESULTS: We pooled the datasets of the Global Use of Streptokinase and tPA for Occluded Arteries (GUSTO)-IIb, Platelet Glycoprotein IIb/IIIa in Unstable Angina: Receptor Suppression Using Integrilin Therapy (PURSUIT), Platelet IIb/IIIa Antagonism for the Reduction of Acute Coronary Syndrome Events in a Global Organization Network (PARAGON)-A, and PARAGON-B trials (n=26 416). We identified independent predictors of ventricular fibrillation (VF) and ventricular tachycardia (VT) and compared the 30-day and 6 month mortality rates of patients who did (n=552) and did not (n=25 864) develop these arrhythmias during the index hospitalization. Independent predictors of in hospital VF included prior hypertension, chronic obstructive pulmonary disease, prior myocardial infarction, and ST-segment changes at presentation. Except for hypertension, these variables also independently predicted in-hospital VT. In Cox proportional-hazards modeling, in-hospital VF and VT were independently associated with 30-day mortality (hazard ratio [HR], 23.2 [95% CI, 18.1 to 29.8] for VF and HR, 7.6 [95% CI, 5.5 to 10.4] for VT) and 6-month mortality (HR, 14.8 [95% CI, 12.1 to 18.3] for VF and HR, 5.0 [95% CI, 3.8 to 6.5] for VT). These differences remained significant after excluding patients with heart failure or cardiogenic shock and those who died <24 hours after enrollment. CONCLUSIONS: Despite the use of effective therapies for non-ST-elevation acute coronary syndromes, ventricular arrhythmias in this setting are associated with increased 30-day and 6-month mortality. More effective therapies are needed to improve the survival of patients with these arrhythmias. PMID- 12119246 TI - Integrated analysis of myocardial blush and ST-segment elevation recovery after successful primary angioplasty: Real-time grading of microvascular reperfusion and prediction of early and late recovery of left ventricular function. AB - BACKGROUND: ST-segment elevation (SigmaSTe) recovery and the angiographic myocardial blush (MB) grade are useful markers of microvascular reperfusion after recanalization of the infarct-related artery. We investigated the ability of a combined analysis of MB grade and SigmaSTe changes to identify different patterns of myocardial reperfusion shortly after primary percutaneous coronary angioplasty (PTCA) and to predict 7-day and 6-month left ventricular (LV) functional recovery. METHODS AND RESULTS: MB grade and SigmaSTe recovery were evaluated shortly after successful primary PTCA (restoration of TIMI grade 3 flow) in 114 consecutive patients with SigmaSTe acute myocardial infarction. LV function was assessed by 2D echocardiograms before PTCA and at 7 days and 6 months thereafter. By combining MB and SigmaSTe changes, 3 main groups of patients were identified. Group 1 patients (n=60) had both significant MB (grade 2 to 3) and SigmaSTe recovery (>50% versus basal SigmaSTe) and a high rate of 7-day (65%) and 6-month (95%) LV functional recovery. In group 2 patients (n=21), who showed MB but persistent SigmaSTe, the prevalence of early LV functional recovery was low (24%) but increased up to 86% in the late phase. Group 3 patients (n=28), who had neither significant MB nor SigmaSTe resolution, had poor early (18%) and late (32%) LV functional recovery. CONCLUSIONS: After successful primary PTCA, integrated analysis of MB and SigmaSTe recovery allows a real-time grading of microvascular reperfusion of the infarct area and predicts the time-course and magnitude of LV functional recovery. PMID- 12119247 TI - Assessment of survival in patients with primary pulmonary hypertension: importance of cardiopulmonary exercise testing. AB - BACKGROUND: Primary pulmonary hypertension (PPH) is a life-threatening disease. Prognostic assessment is an important factor in determining medical treatment and lung transplantation. Whether cardiopulmonary exercise testing data predict survival has not been reported previously. METHODS AND RESULTS: We studied 86 patients with PPH (58 female, age 46+/-2 years, median NYHA class III) between 1996 and 2001 who were followed up in a tertiary referral center. Right heart catheterization was performed and serum uric acid levels were measured in all patients. Seventy patients were able to undergo exercise testing. At the start of the study, the average pulmonary artery pressure was 60+/-2 mm Hg, average pulmonary vascular resistance was 1664+/-81 dyne x s x cm(-5), average serum uric acid level was 7.5+/-0.35 mg/dL, and average peak oxygen uptake during exercise (peak VO(2) was 11.2+/-0.5 mL x kg(-1) x min(-1). During follow-up (mean: 567+/ 48 days), 28 patients died and 16 underwent lung transplantation (1-year cumulative event-free survival: 68%; 95% CI 58 to 78). The strongest predictors of impaired survival were low peak VO(2) (P<0.0001) and low systolic blood pressure at peak exercise (peak SBP; P<0.0001). In a multivariable analysis, serum uric acid levels (all P<0.005) and diastolic blood pressure at peak exercise independently predicted survival (P<0.05). Patients with peak VO(2) < or =10.4 mL x kg(-1) x min(-1) and peak SBP < or =120 mm Hg (ie, 2 risk factors) had poor survival rates at 12 months (23%), whereas patients with 1 or none of these risk factors had better survival rates (79% and 97%, respectively). CONCLUSIONS: Peak VO(2) and peak SBP are independent and strong predictors of survival in PPH patients. Hemodynamic parameters, although also accurate predictors, provide no independent prognostic information. PMID- 12119248 TI - Effects of ventricular premature stimulus coupling interval on blood pressure and heart rate turbulence. AB - BACKGROUND: Heart rate turbulence (HRT) is a promising noninvasive risk stratifier for mortality after myocardial infarction. On the basis of a study of ventricular premature complex coupling interval and sympathetic nerve burst amplitude, we hypothesized that measures of HRT would increase with increased prematurity of the coupling interval. METHODS AND RESULTS: Twenty-eight patients undergoing programmed electrical stimulation were studied (12 with prior myocardial infarction, aged 60+/-18 years). An extrastimulus was delivered from the right ventricular apex after 20 sinus beats with a V-S(2) coupling interval decremented by 20 to 30 ms until refractoriness was reached. Turbulence slope (TS), turbulence timing (TT), and turbulence onset were calculated for each extrastimulus, and the linear regressions of these parameters on coupling interval and compensatory pause were calculated. Arterial blood pressure was measured with arterial catheter or a noninvasive continuous blood pressure transducer (Buffington cuff). TS and turbulence onset were abnormal in 4 and 13 patients, respectively. HRT parameters were significantly correlated with coupling interval or compensatory pause in only 2 or 3 patients for a given regression analysis. This absence of correlation was found likely to be due to lack of correlation between compensatory pause and systolic blood pressure after the compensatory pause. Heart rate and TS were correlated: Patients with high heart rate had low TS and late TT (TS=-2.7+0.01xsinus cycle length, P=0.018; TT=8.8 to 0.005xsinus cycle length, P=0.013). CONCLUSIONS: HRT can be induced by programmed stimulation. In this setting, heart rate affects HRT but not ventricular premature complex prematurity. Induced HRT seems to be a valid method for measuring HRT parameters in patients with few ventricular premature complexes. PMID- 12119249 TI - Use of irbesartan to maintain sinus rhythm in patients with long-lasting persistent atrial fibrillation: a prospective and randomized study. AB - BACKGROUND: Data from studies of angiotensin-converting enzyme inhibitors provide evidence that the renin-angiotensin-aldosterone system plays a role as a mediator of atrial remodeling in atrial fibrillation. The present study has evaluated the effect of treatment with the angiotensin I type 1 receptor blocker irbesartan on maintaining sinus rhythm after conversion from persistent atrial fibrillation. METHODS AND RESULTS: To be included in the present study, patients must have had an episode of persistent atrial fibrillation for >7 days. The patients were then randomized and scheduled for electrical cardioversion. Two groups of patients were compared: Group I was treated with amiodarone, and group II was treated with amiodarone plus irbesartan. The primary end point was the length of time to a first recurrence of atrial fibrillation. From a total of 186 patients assessed in the study, 154 were analyzed with the use of intention-to-treat analysis. Seventy five patients were randomly allocated to group I and 79 to group II. After 2 months of follow-up in the intention-to-treat analysis, the group treated with irbesartan had fewer patients with recurrent atrial fibrillation (Kaplan-Meier analysis, 84.79% versus 63.16%, P=0.008). The Kaplan-Meier analysis of time to first recurrence during the follow-up period (median time, 254 days [range, 60 to 710]) also showed that patients treated with irbesartan had a greater probability of remaining free of atrial fibrillation (79.52% versus 55.91%, P=0.007). CONCLUSIONS: Patients treated with amiodarone plus irbesartan had a lower rate of recurrence of atrial fibrillation than did patients treated with amiodarone alone. PMID- 12119250 TI - Evidence for heritability of abdominal aortic calcific deposits in the Framingham Heart Study. AB - BACKGROUND: Atherosclerosis is a systemic disease that underlies clinical cardiovascular disease. The radiographic finding of abdominal aortic calcific deposits is an indicator of the presence of aortic atherosclerosis and an independent predictor of cardiovascular disease events. Little is known about the heritability of aortic calcification. METHODS AND RESULTS: Original Framingham Heart Study cohort participants (2151) in 1109 extended pedigrees had a lateral lumbar radiograph. The presence and severity of abdominal aortic calcific (AAC) deposits at the levels of the first through fourth lumbar vertebrae was graded by a previously validated rating scale. Correlation coefficients were calculated in pairs of siblings, parent-offspring, and spouses. Age-, sex-, and multivariable adjusted correlation coefficients for AAC were 0.52 for parent-offspring pairs and 0.20 for sibling pairs. In contrast, the multivariable-adjusted correlation for AAC in spouse pairs was -0.02. Using variance component methods implemented in SOLAR, the estimated heritability for age-, sex-, and multivariable-adjusted AAC was 0.49 (P<0.001). Thirty-one percent of the overall variance in AAC deposits was due to measured covariates, and 49% to heritable factors. CONCLUSIONS: In our large, population-based sample, heritable factors play a role in the presence and extent of abdominal aortic calcification. Thus, a substantial proportion of the variation in AAC is due to additive effects of genes, which have yet to be characterized. Measures of aortic atherosclerosis may provide heritable quantitative phenotypes for the genetic dissection of the complex condition of atherosclerosis in human populations. PMID- 12119251 TI - Prognosis of aortic intramural hematoma with and without penetrating atherosclerotic ulcer: a clinical and radiological analysis. AB - BACKGROUND: Advances in imaging techniques have increased the recognition of aortic intramural hematomas (IMHs) and penetrating atherosclerotic ulcers (PAUs); however, distinction between IMH and PAU remains unclear. We intended to clarify differences between IMH coexisting with PAU and IMH not associated with PAU by comparisons of clinical features, imaging findings, and patient outcome to derive the optimal therapeutic approach. METHODS AND RESULTS: We performed a retrospective analysis of 65 symptomatic patients with aortic IMH. There were 34 patients with IMH associated with PAU (group 1) and 31 patients with IMH unaccompanied by PAU (group 2). Involvement of the ascending aorta (type A) was more frequent in group 2 (8 of 31, 26%), whereas most of the patients in group 1 had exclusive involvement of the descending aorta (type B) (31of 34, 91%). Patients were subdivided into 2 categories, those with clinical progression and those with stable disease. Forty-eight percent of patients in group 1 and 8% in group 2 were in the progressive category (P=0.002). Clinical and radiological findings were compared between those group 1 patients who had a progressive disease course (n=12) and those who were stable (n=13). Sustained or recurrent pain (P<0.0001), increasing pleural effusion (P=0.0003), and both the maximum diameter (P=0.004) and maximum depth (P=0.003) of the PAU were reliable predictors of disease progression. CONCLUSIONS: This study suggests a difference in disease behavior that argues for the prognostic importance of making a clear distinction between IMH caused by PAU and IMH not associated with PAU. IMH with PAU was significantly associated with a progressive disease course, whereas IMH without PAU typically had a stable course, especially when limited to the descending thoracic aorta. PMID- 12119252 TI - Beta(2)-adrenergic receptor gene delivery to the endothelium corrects impaired adrenergic vasorelaxation in hypertension. AB - BACKGROUND: Impaired beta-adrenergic receptor (AR)-mediated vasorelaxation in hypertension plays a role in increased peripheral vascular resistance and blood pressure. Because the beta(2)AR is the most abundant vascular AR subtype, we sought to enhance betaAR vasorelaxation by overexpressing beta(2)ARs via adenoviral-mediated gene transfer (ADbeta(2)AR) to the vascular endothelium of the carotid artery. METHODS AND RESULTS: In normotensive Wistar-Kyoto (WKY) and spontaneously hypertensive (SHR) rats, we exposed the right common carotid artery to ADbeta(2)AR in situ for 15 minutes by injection into the lumen while the blood flow was interrupted. Control carotids received an empty vector (ADempty). Three days later, transgene expression and selective endothelial localization were confirmed in infected vessels. Vasoregulation after beta(2)AR overexpression (2 fold) was studied in isolated organ baths. ADbeta(2)AR carotid responses to alpha(1)AR and alpha(2)AR agonists were not affected, whereas responses to epinephrine were altered and betaAR-mediated vasorelaxation was enhanced after beta(2)AR overexpression. As expected, betaAR-mediated vasodilatation in control carotids of SHR rats was significantly less than in similar control WKY carotid arteries. ADbeta(2)AR treatment enhanced betaAR vasorelaxation in SHR to levels similar to those seen in ADbeta(2)AR WKY carotids. CONCLUSIONS: Our results demonstrate a critical role for the endothelium in betaAR-mediated vasorelaxation and suggest that impaired betaAR signaling may account for dysfunctional betaAR vasorelaxation in hypertension rather than impaired endothelium-dependent nitric oxide metabolism. PMID- 12119253 TI - Nifedipine indirectly upregulates superoxide dismutase expression in endothelial cells via vascular smooth muscle cell-dependent pathways. AB - BACKGROUND: Calcium antagonists normalize endothelial dysfunction in many cardiovascular diseases. There is no known receptor, however, for calcium antagonists in endothelial cells (ECs). We hypothesized that vascular smooth muscle cells (VSMCs) are involved in the mechanism underlying the normalization of endothelial dysfunction by calcium antagonists. METHODS AND RESULTS: Coculture studies with ECs and VSMCs were performed to determine whether VSMCs mediate modulation of endothelial superoxide dismutase (SOD) activity and expression induced by the calcium antagonist nifedipine. Nifedipine induced upregulation of SOD activity in rat aortic segments but had no effect on SOD expression or activity in ECs or VSMCs cultured individually. When ECs were cocultured with VSMCs, however, nifedipine upregulated SOD expression and activity in ECs. Nifedipine stimulated vascular endothelial growth factor (VEGF) production from VSMCs, and this stimulation of VEGF production was abolished by HOE-140, an antagonist of the bradykinin B(2) receptor. A neutralizing antibody against VEGF inhibited the upregulation of endothelial SOD by nifedipine. In addition, recombinant VEGF induced an increase in the levels of SOD expression in ECs, and supernatant derived from nifedipine-treated VSMCs enhanced NO production from ECs. This increase in NO production by the supernatant was inhibited by preincubation of ECs with SOD antisense oligodeoxyribonucleotides. CONCLUSIONS: The calcium antagonist nifedipine indirectly upregulates endothelial SOD expression by stimulating VEGF production from adjacent VSMCs. This finding may provide further insight into the mechanism underlying the beneficial effects of calcium antagonists in cardiovascular diseases. PMID- 12119254 TI - Probucol attenuates left ventricular dysfunction and remodeling in tachycardia induced heart failure: roles of oxidative stress and inflammation. AB - BACKGROUND: Oxidative stress and inflammation are potentially involved in the pathogenesis of heart failure (HF). We examined whether antioxidant and antiinflammatory treatment with probucol decreases myocardial oxidative stress and inflammation and attenuates the progression of left ventricular (LV) dysfunction and remodeling (dilatation) in tachycardia-induced HF. METHODS AND RESULTS: We studied 3 groups of dogs: a sham-operated control group and 2 other groups that underwent ventricular pacing at 240 bpm with and without probucol treatment (100 mg/kg IP per week) for 4 weeks. Dogs that underwent ventricular pacing for 4 weeks developed signs of HF, such as a reduction in the LV ejection fraction and increases in the LV end-diastolic dimension and LV end-diastolic pressure. Myocardial oxidative stress, as measured by electron spin resonance spectroscopy with 4-hydroxy-2,2,6,6,-tetramethyl-piperidine-N-oxyl (hydroxy TEMPO), was significantly increased. There was an increase in myocardial monocyte infiltration, monocyte chemoattractant protein-1 expression, and renin angiotensin system and matrix metalloproteinase activity. Probucol treatment prevented increases in oxidative stress, inflammation, and matrix metalloproteinase activity and attenuated LV dysfunction and remodeling. CONCLUSIONS: Probucol attenuated LV dysfunction and remodeling, possibly through its antioxidant and/or antiinflammatory effects in ventricular pacing-induced HF. These data suggest that inflammatory disorders, which cause an abnormal interaction between failing myocardium and activated monocytes, have an important role in the progression of HF. PMID- 12119255 TI - Adverse outcomes of interrupted precordial compression during automated defibrillation. AB - BACKGROUND: Current versions of automated external defibrillators (AEDs) require frequent stopping of chest compression for rhythm analyses and capacity charging. The present study was undertaken to evaluate the effects of these interruptions during the operation of AEDs. METHODS AND RESULTS: Ventricular fibrillation was electrically induced in 20 male domestic swine weighing between 37.5 and 43 kg that were untreated for 7 minutes before CPR was started. Defibrillation was attempted with up to 3 sequential 150-J biphasic shocks, but each was preceded by 3-, 10-, 15-, or 20-second interruptions of chest compression. The interruptions corresponded to those that were mandated by commercially marketed AEDs for rhythm analyses and capacitor charge. The sequence of up to 3 electrical shocks and delays were repeated at 1-minute intervals until the animals were successfully resuscitated or for a total of 15 minutes. Spontaneous circulation was restored in each of 5 animals in which precordial compression was delayed for 3 seconds before the delivery of the first and subsequent shocks but in none of the animals in which the delay was >15 seconds before the delivery of the first and subsequent shocks. Longer intervals of CPR interventions were required, and there was correspondingly greater failure of resuscitation in close relationship to increasing delays. The durations of interruptions were inversely related to the durations of subthreshold levels of coronary perfusion pressure. Postresuscitation arterial pressure and left ventricular ejection fraction were more severely impaired with increasing delays. CONCLUSIONS: Interruptions of precordial compression for rhythm analyses that exceed 15 seconds before each shock compromise the outcome of CPR and increase the severity of postresuscitation myocardial dysfunction. PMID- 12119256 TI - Decompression-triggered positive-pressure ventilation during cardiopulmonary resuscitation improves pulmonary gas exchange and oxygen uptake. AB - BACKGROUND: Intermittent positive-pressure ventilation (IPPV) is the "gold standard" of ventilation during cardiopulmonary resuscitation (CPR), but continuous positive airway pressure (CPAP) is increasingly discussed as an alternative. This study investigated hemodynamics and pulmonary gas exchange applying CPAP enhanced with pressure support ventilation (CPAP(PSV)) during CPR. METHODS AND RESULTS: Twenty-four pigs were subjected to ventricular fibrillation and CPR with CPAP(PSV), CPAP, or IPPV. Measurements were taken before (hemodynamics, blood gases, inert gas measurements) and 10 (hemodynamics, blood gases) and 20 (hemodynamics, blood gases, inert gas measurements) minutes after induction of ventricular fibrillation. Although no significant intergroup differences in hemodynamics were found, arterial partial pressure of oxygen (PaO(2)) was significantly higher during CPAP(PSV) compared with CPAP or IPPV (98+/-10, 61+/-27, and 71+/-30 mm Hg, respectively, P<0.05). CPAP(PSV) resulted in an alveolar-arterial partial pressure of oxygen difference of 56+/-17 mm Hg, whereas during CPAP, 83+/-21 mm Hg was detected, and during IPPV, 98+/-29 mm Hg was detected (P<0.05). Pulmonary blood flow to lung units with a normal VA/Q ratio in percent of cardiac output was 76+/-17% during CPAP(PSV), 61+/-21% during CPAP (P<0.01), and 54+/-13% during IPPV (P<0.01). Oxygen uptake (VO(2)) was significantly higher during CPAP(PSV) than with the other ventilation modes (P<0.05) and comparable to the baseline value in intragroup comparison. Return of spontaneous circulation was recorded in 8 of 8 animals in the CPAP(PSV) group, in 6 of 8 in the CPAP group, and in 3 of 8 in the IPPV group. CONCLUSIONS: CPAP(PSV) provides a straightforward and effective alternative to IPPV or CPAP during CPR that provides significantly higher PaO(2) and VO(2). PMID- 12119257 TI - Platelet glycoprotein IIb/IIIa inhibitors: recognition of a two-edged sword? PMID- 12119258 TI - Images in cardiovascular medicine: Giant right ventricular fibroma in an infant. PMID- 12119259 TI - AHA Guidelines for Primary Prevention of Cardiovascular Disease and Stroke: 2002 Update: Consensus Panel Guide to Comprehensive Risk Reduction for Adult Patients Without Coronary or Other Atherosclerotic Vascular Diseases. American Heart Association Science Advisory and Coordinating Committee. PMID- 12119260 TI - Images in cardiovascular medicine: Magnetic resonance angiography of a congenitally interrupted aortic arch. PMID- 12119263 TI - Protein Kinase A and Human Disease. September 10, 2001. Bethesda, Maryland, USA. Conference proceedings. PMID- 12119264 TI - Mutations of the gene encoding the protein kinase A type I-alpha regulatory subunit (PRKAR1A) in patients with the "complex of spotty skin pigmentation, myxomas, endocrine overactivity, and schwannomas" (Carney complex). AB - Carney complex (CNC) is a familial multiple neoplasia syndrome associated with abnormal skin and mucosal pigmentation. The complex has features overlapping those of McCune-Albright syndrome (MAS) and the other multiple endocrine neoplasias (MENs). CNC is inherited as an autosomal dominant trait, and the responsible genes have been mapped by linkage analysis to loci at 2p16 and 17q22 24. Because of its unusual biochemical features (e.g., paradoxical responses to various endocrine signals) and its clinical similarities to MAS, genes implicated in cyclic nucleotide-dependent signaling, including GNAS1 (which is responsible for MAS), had been considered likely candidates for causing CNC. The gene encoding the protein kinase A (PKA) type I-alpha regulatory subunit (RI alpha), PRKAR1A, had been mapped to 17q22-24; loss-of-heterozygosity (LOH) analysis using polymorphic markers from this region revealed consistent changes in tumors from patients with CNC, including those from one family previously mapped to 17q22-24. Investigation of a polymorphic site within the 5' of the PRKAR1A gene showed segregation with the disease and retention of the allele bearing the disease gene in CNC tumors. Mutations of the PRKAR1A gene were also found to have occurred de novo in sporadic cases of CNC; no mutations were found in kindreds mapping to 2p16. Thus, genetic heterogeneity in CNC was confirmed; in total, 41% of all patients with CNC had mutations in the PRKAR1A gene. All mutations were frameshifts, insertions, and deletions that led to nonsense mRNA and premature termination of the predicted peptide product. Functional studies in CNC tumors suggested that inactivating mutations of the PRKAR1A gene led to nonsense mRNA decay (the mutant peptide product was not present) and were associated with dysregulated PKA activity, increased responsiveness to cAMP, and excess of type II PKA activity. We conclude that the PRKAR1A gene, coding for the RIalpha subunit of PKA, a critical cellular component of a number of cyclic nucleotide dependent signaling pathways, is mutated in a subset of patients with CNC. In their tumors, there is LOH of the normal allele, suggesting that normal RI-alpha may have tumor suppression function in the tissues affected by CNC. An excess of type-II PKA activity was present in affected tissues, which may be responsible for the apparent tumorigenicity of PRKAR1A mutations in endocrine tissues. PMID- 12119265 TI - Dissecting the circuitry of protein kinase A and cAMP signaling in cancer genesis: antisense, microarray, gene overexpression, and transcription factor decoy. AB - Expression of the RI alpha subunit of the cAMP-dependent protein kinase type I (PKA-I) is enhanced in human cancer cell lines, in primary tumors, in transformed cells, and in cells upon stimulation of growth. Signaling via the cAMP pathway may be complex, and the biological effects of the pathway in normal cells may depend upon the physiological state of the cells. However, results of different experimental approaches such as antisense exposure, 8-Cl-cAMP treatment, and gene overexpression have shown that the inhibition of RI alpha/PKA-I exerts antitumor activity in a wide variety of tumor-derived cell lines examined in vitro and in vivo. cDNA microarrays have further shown that in a sequence-specific manner, RI alpha antisense induces alterations in the gene expression profile of cancer cells and tumors. The cluster of genes that define the "proliferation transformation" signature are down-regulated, and those that define the "differentiation-reverse transformation" signature are up-regulated in antisense treated cancer cells and tumors, but not in host livers, exhibiting the molecular portrait of the reverted (flat) phenotype of tumor cells. These results reveal a remarkable cellular regulation, elicited by the antisense RI alpha, superimposed on the regulation arising from the Watson-Crick base-pairing mechanism of action. Importantly, the blockade of both the PKA and PKC signaling pathways achieved with the CRE-transcription factor decoy inhibits tumor cell growth without harming normal cell growth. Thus, a complex circuitry of cAMP signaling comprises cAMP growth regulatory function, and deregulation of the effector molecule by this circuitry may underlie cancer genesis and tumor progression. PMID- 12119266 TI - Regulatory subunits of PKA and breast cancer. AB - Overexpression of the R subunits of PKA (in particular, RI) is associated with high proliferation in normal breast, malignant transformation in the breast, poor prognosis in established breast cancer, and resistance to antiestrogens. These data, together with the observation that successful antiestrogen therapy is associated with reduced expression of RI mRNA, suggest that targeting R subunits is an appropriate therapeutic strategy for breast cancer. Initial experimental results, using antisense RI oligonucleotides, are promising in terms of reducing the growth rate of breast cancer cells and xenografts. While clinical trials designed to target RI subunits have yet to be established (and interventions as preventative measures are even more distant), the concept of these approaches to prevent and treat breast cancer should be developed and exploited. PMID- 12119267 TI - Reinventing the wheel of cyclic AMP: novel mechanisms of cAMP signaling. AB - Mechanisms of cAMP signal transduction have been thoroughly investigated for more than 40 years. From the binding of hormonal ligands to their receptors on the outer surface of the plasma membrane to the cytoplasmic activation of effectors, the ensuing cAMP signaling cascades and the nuclear gene regulatory functions, coupled with the structural elucidation of the cAMP-dependent protein kinase (PKA) and in vivo functional characterizations of each of the components of PKA by homologous recombination gene targeting, our understanding of cAMP-mediated signal transduction has reached its pinnacle. Despite this trove of knowledge, some recent findings have emerged that suggest hitherto novel and alternative mechanisms of cAMP action that could increase the signaling bandwidth of cAMP and PKA in cell growth and transcriptional regulation. This article attempts to review some of these novel and unconventional mechanisms of cAMP and PKA signaling, and to generate further enthusiasm in investigating and validating these new frontiers of the cAMP signal transduction pathway. PMID- 12119268 TI - Role of the PKA-regulated transcription factor CREB in development and tumorigenesis of endocrine tissues. AB - The cAMP pathway plays a major role in the development of endocrine tissues and various molecular defects of key components of this pathway (G protein, receptors, PKA, etc.) have been observed in endocrine tumors. The ubiquitous transcription factor CREB (cAMP-response element binding protein) binds to the cAMP response element (CRE) and stimulates transcription after phosphorylation on Ser(133) by PKA. The CREB family of transcription factors contains three members: CREB, CREM, and ATF-1. Targeted expression of dominant-negative mutants of CREB in transgenic mice leads to somatotrophs or thyroid hypoplasia. GH-secreting adenomas are benign secreting tumors expressing an activated mutant G alpha s protein (Gsp) in about 40% of cases. In GH-secreting adenomas CREB is always expressed and often highly phosphorylated. The CREM isoform ICER is stimulated by cAMP, and its expression is increased in Gsp-harboring tumors. After transfection in pituitary somatotroph cells, activating mutations of Gs protein (Gsp) and overexpression of wild-type G alpha S stimulate transcription of various CRE containing promoters via CREB in a Ser(133)-specific-dependent manner. Activation of the cAMP pathway by ACTH is required for adrenal cortex (AdCx) maintenance and steroidogenesis. CREB is expressed in normal AdCx. Alterations of CRE binding proteins with loss of CREB expression and compensatory overexpression of CREMtau is observed in the human adrenocortical cancer cell line H295R. Similar alterations are found at the protein level in human malignant adrenocortical tumors. In conclusion, the CREB family of transcription factors plays an important role in the development, differentiation, and proliferation of endocrine tissues. Various alterations of the CREB family of transcription factors can be observed in endocrine tumors. PMID- 12119269 TI - The essential role of RI alpha in the maintenance of regulated PKA activity. AB - Cloning of the individual regulatory (R) and catalytic (C) subunits of the cAMP dependent protein kinase (PKA) and expression of these subunits in cell culture have provided mechanistic answers about the rules for PKA holoenzyme assembly. One of the central findings of these studies is the essential role of the RI alpha regulatory subunit in maintaining the catalytic subunit under cAMP control. The role of RI alpha as the key compensatory regulatory subunit in this enzyme family was confirmed by gene knockouts of the three other regulatory subunits in mice. In each case, RI alpha has demonstrated the capacity for significant compensatory regulation of PKA activity in tissues where the other regulatory subunits are expressed, including brain, brown and white adipose tissue, skeletal muscle, and sperm. The essential requirement of the RI alpha regulatory subunit in maintaining cAMP control of PKA activity was further corroborated by the knockout of RI alpha in mice, which results in early embryonic lethality due to failed cardiac morphogenesis. Closer examination of RI alpha knockout embryos at even earlier stages of development revealed profound deficits in the morphogenesis of the mesodermal embryonic germ layer, which gives rise to essential structures including the embryonic heart tube. Failure of the mesodermal germ layer in RI alpha knockout embryos can be rescued by crossing RI alpha knockout mice to C alpha knockout mice, supporting the conclusion that inappropriately regulated PKA catalytic subunit activity is responsible for the phenotype. Isolation of primary embryonic fibroblasts from RI alpha knockout embryos reveals profound alterations in the actin-based cytoskeleton, which may account for the failure in mesoderm morphogenesis at gastrulation. PMID- 12119270 TI - Deficient protein kinase a in systemic lupus erythematosus: a disorder of T lymphocyte signal transduction. AB - Systemic lupus erythematosus (SLE) is an idiopathic autoimmune disease characterized by impaired T lymphocyte immune effector functions. We have identified a disorder of signal transduction in SLE T cells involving the cyclic AMP/protein kinase A (cAMP/PKA) pathway. Cyclic AMP-stimulated PKA-catalyzed protein phosphorylation is markedly diminished owing to profound deficiencies of both type I (PKA-I) and type II (PKA-II) isozyme activities. Deficient PKA-I isozyme is characterized by a significant reduction in the amount of type I regulatory beta subunit (RI beta) steady state mRNA by competitive polymerase chain reaction. This is associated with a 30% decrease in RI alpha protein and a 65% reduction in RI beta protein. Indeed, T cells from approximately 25% of SLE subjects have no detectable RI beta protein. Transient transfection of T cells not expressing RI beta protein with autologous SLE RI beta cDNA bypassed the block in translation, reconstituting PKA activity and augmenting IL-2 production. Of importance was the initial identification of novel RI alpha mRNA mutations characterized by heterogeneous transcript mutations, including deletions, transitions, and transversions. Most mutations are clustered adjacent to GAGAG motifs and CT repeats. By contrast, deficient PKA-II activity is the result of spontaneous dissociation of the cytosolic RII beta(2)C(2) holoenzyme, aberrant RII beta translocation to the nucleus from the cytosol, and retention of RII beta in the nucleus. In conclusion, distinct mechanisms account for deficient PKA-I and PKA-II isozyme activities in SLE T cells. PMID- 12119271 TI - The role of cyclic AMP and its effect on protein kinase A in the mitogenic action of thyrotropin on the thyroid cell. AB - Cyclic AMP has been shown to inhibit cell proliferation in many cell types and to activate it in some. The latter has been recognized only lately, thanks in large part to studies on the regulation of thyroid cell proliferation in dog thyroid cells. The steps that led to this conclusion are outlined. Thyrotropin activates cyclic accumulation in thyroid cells of all the studied species and also phospholipase C in human cells. It activates directly cell proliferation in rat cell lines, dog, and human thyroid cells but not in bovine or pig cells. The action of cyclic AMP is responsible for the proliferative effect of TSH. It accounts for several human diseases: congenital hyperthyroidism, autonomous adenomas, and Graves' disease; and, by default, for hypothyroidism by TSH receptor defect. Cyclic AMP proliferative action requires the activation of protein kinase A, but this effect is not sufficient to explain it. Cyclic AMP action also requires the permissive effect of IGF-1 or insulin through their receptors, mostly as a consequence of PI3 kinase activation. The mechanism of these effects at the level of cyclin and cyclin-dependent protein kinases involves an induction of cyclin D3 by IGF-1 and the cyclic AMP-elicited generation and activation of the cyclin D3-CDK4 complex. PMID- 12119272 TI - Cyclic AMP and the reverse transformation reaction. AB - Traditional methods for cancer treatment have been aimed at killing the cancer cells. Unfortunately this approach all too often is accompanied by harmful killing of normal cells. The present paper describes an experimental program in our laboratory in which cancer cells are treated so as to revert to normal cell behavior. This process, which we have named reverse transformation, appears to offer considerable hope in the treatment of a large number of malignancies. PMID- 12119273 TI - Protein kinase A as target for novel integrated strategies of cancer therapy. AB - We have studied the role of protein kinase A (PKA) in neoplastic transformation, apoptosis, and angiogenesis and its relationship with other signaling molecules, as a basis for developing novel therapeutic strategies. We demonstrated the involvement of PKA type I (PKA-I) in the transduction of mitogenic signals from different sources and demonstrated functional and structural interactions between PKA-I and the activated epidermal growth factor receptor (EGFR). We contributed to the identification and development of several selective inhibitors of PKA-I, such as 8-Cl-cAMP and a hybrid DNA/RNA antisense oligonucleotide of a novel class (AS-PKA-I) and of EGFR, including mAbC225 and ZD1839 (Iressa). All these agents have been investigated in cancer patients. We demonstrated the therapeutic potential of the combined blockade of PKA-I and EGFR, reporting a synergistic antitumor effect when their inhibitors are used in combination. We have also shown that PKA-I and EGFR inhibitors are able to cooperate with selected class of cytotoxic drugs and with ionizing radiation, causing a synergistic inhibition of tumor growth in vitro and in vivo, accompanied by inhibition of expression of growth and angiogenic factors and by suppression of vessel production. Moreover, PKA-I is implicated in a bcl-2-dependent apoptotic pathway, and we have recently reported a cooperative antitumor and proapoptotic effect of AS-PKA-I in combination with an AS-bcl-2. Finally, we have shown that AS-PKA-I also has antitumor and antiangiogenic effects following oral administration and that they can be greatly enhanced in combination with oral ZD1839 and oral taxanes. PMID- 12119274 TI - Protein kinase A and chromosomal stability. AB - All malignant human tumors contain chromosomal rearrangements. Among them, the majority of solid tumors show chromosomal instability, caused by aberrations in chromosomal segregation during cell division. Chromosomal instability, defined as increased probability of formation of novel chromosomal mutations compared to that of normal or control cells, appears to be a feature of tumorigenesis in vivo and in vitro (in cancer cell lines). Several enzymatic kinases are involved in maintaining proper chromosomal segregation and regulating cell cycle progression. One such kinase, cAMP-dependent protein kinase A (PKA), has a functional role in many aspects of cell signaling, metabolism, and proliferation. In this review, we will discuss the potential participation of PKA in chromosomal stability. This role includes the association of PKA with the centrosome, microtubules, and the anaphase-promoting complex/cyclosome (ACP/C), all key aspects of proper chromosomal segregation. PMID- 12119275 TI - Protein kinase A: regulation and receptor-mediated delivery of antisense oligonucleotides and cytotoxic drugs. AB - Protein kinases help regulate eukaryotic cell division. We investigated the regulation of cAMP-dependent protein kinase A (PKA) and casein kinase (CK) type I activity in normal cells and in cancer. To assess this activity in biopsies we suggest a new parameter--the ratio of CK activity and total PKA activity divided by cAMP concentration: CK/PKA/cAMP. In 98 samples of colon mucosa in normal, inflamed, polyp, and adenocarcinoma cells, we found this parameter to be fairly constant in normal conditions and increased 10-fold in colon cancer; the ratio does not depend on the place of biopsy or the patient's age or sex. Experiments with model systems of concanavalin A-stimulated lymphocytes and regenerating rat liver showed that in normal cell proliferation the parameter increases 2-3-fold, as compared to a 30-fold increase in cancer. Unlike normal cells, malignant cells show CK activation and decrease of cAMP; therefore, PKA activity decreases. This suggests a correlation of CK and PKA activity and significant damage to their regulation at pathological changes of tissue proliferation. To further study concerted CK and PKA regulation we used monoclonal antibodies (mAbs) against cAMP dependent protein kinase regulatory subunit RKII beta. We produced 11 antibodies in three groups: inhibiting, which block cAMP binding with RII beta and inhibit holoenzyme formation (RS6); activating, which enhance cAMP binding and do not affect holoenzyme formation (RS28); and neutral (RS17). To investigate mAb influence on protein kinase regulation in live cells we permeabilized pheochromocytoma PC12 by digitonin. When used at 5-microM concentration for 5 min, digitonin allowed us to deliver mAb into PC12 cells at 30-34-nM concentration, leaving 68-75% viable cells. Protein kinase activity was measured within 0.5 and 4 h after incorporation of mAbs into cells. After 30 min incorporation, mAb RS6 blocked PKA activation in PC12 cells under the influence of cAMP; other mAbs showed no effect. mAb RS6 caused a 4-fold increase of free C subunit activity 4 h after incorporation. mAb RS38 decreased R2C2 activity and did not influence C subunit activity. The change of free C subunit activity caused by mAb incorporation was followed by a synchronized, well-balanced change of CK type I activity, which suggests a correlation between the two phosphorylation systems of cell proteins. PMID- 12119276 TI - Gs(alpha) mutations and imprinting defects in human disease. AB - Gs is the ubiquitously expressed heterotrimeric G protein that couples receptors to the effector enzyme adenylyl cyclase and is required for receptor-stimulated intracellular cAMP generation. Activated receptors promote the exchange of GTP for GDP on the Gs alpha-subunit (Gs(alpha)), resulting in Gs activation; an intrinsic GTPase activity of Gs(alpha) deactivates Gs by hydrolyzing bound GTP to GDP. Mutations of Gs(alpha) residues involved in the GTPase reaction that lead to constitutive activation are present in endocrine tumors, fibrous dysplasia of bone, and McCune-Albright syndrome. Heterozygous loss-of-function mutations lead to Albright hereditary osteodystrophy (AHO), a disease characterized by short stature, obesity, and skeletal defects, and are sometimes associated with progressive osseous heteroplasia. Maternal transmission of Gs(alpha) mutations leads to AHO plus resistance to several hormones (e.g., parathyroid hormone) that activate Gs in their target tissues (pseudohypoparathyroidism type IA), while paternal transmission leads only to the AHO phenotype (pseudopseudohypoparathyroidism). Studies in both mice and humans demonstrate that Gs(alpha) is imprinted in a tissue-specific manner, being expressed primarily from the maternal allele in some tissues and biallelically expressed in most other tissues. This likely explains why multihormone resistance occurs only when Gs(alpha) mutations are inherited maternally. The Gs(alpha) gene GNAS1 has at least four alternative promoters and first exons, leading to the production of alternative gene products including Gs(alpha), XL alphas (a novel Gs(alpha) isoform expressed only from the paternal allele), and NESP55 (a chromogranin-like protein expressed only from the maternal allele). The fourth alternative promoter and first exon (exon 1A) located just upstream of the Gs(alpha) promoter is normally methylated on the maternal allele and is transcriptionally active on the paternal allele. In patients with parathyroid hormone resistance but without AHO (pseudohypoparathyroidism type IB), the exon 1A promoter region is unmethylated and transcriptionally active on both alleles. This GNAS1 imprinting defect is predicted to decrease Gs(alpha) expression in tissues where Gs(alpha) is normally imprinted and therefore to lead to renal parathyroid hormone resistance. PMID- 12119277 TI - Regulation of phospholipase D and secretion in mast cells by protein kinase A and other protein kinases. AB - Functions attributed to phospholipase (PL) D include the regulation of intracellular trafficking of Golgi-derived vesicles and secretion of granules from mast cells. We have reported that activation of PLD and secretion in a rat mast cell (RBL-2H3) line is substantially enhanced by cholera toxin, a known activator of protein kinase (PK) A. Here we review the evidence that (1) the synergistic interactions of cholera toxin and other pharmacological agents on mast cell secretion are attributable to the synergistic activation of PLD via PKA, CaM kinase II, and PKC and (2) both PLD1 and PLD2 participate in this process. For example, treatment with cholera toxin, thapsigargin, and phorbol 12 myristate 13-acetate (which activate PKA, CaM kinase II, and PKC, respectively) exhibit synergy in the stimulation of both PLD and secretion. These kinases and PLD are likely confined to membrane components, as similar synergistic interactions could be demonstrated in permeabilized cells. The regulation of PLD and secretion by these kinases is also apparent from studies of inhibitors of PKA and other kinases. Also, by overexpression of either PLD1 or PLD2 it is apparent that both isoforms respond to the same stimuli as endogenous PLD, although PLD1 is largely associated with secretory granules and PLD2 with plasma membrane. The studies reveal interesting differences in the regulation of the translocation of granules (regulated by PKA) and the fusion of these granules with the plasma membrane (regulated by Ca(2+) and PKC). The pathological/physiological implications of the regulation of PLD by PKA require further evaluation in other cell systems. PMID- 12119278 TI - Role of PTEN, a lipid phosphatase upstream effector of protein kinase B, in epithelial thyroid carcinogenesis. AB - Both benign and malignant thyroid disease are well-established components of Cowden syndrome (CS), an autosomal dominant disorder characterized by multiple hamartomas and breast cancer that may be considered a phakomatosis. The susceptibility gene for CS is PTEN, a tumor suppressor gene on 10q23.3 that encodes a lipid phosphatase that lies upstream of protein kinase B (Akt). Interestingly, Carney complex is also a phakomatosis where multiple endocrine neoplasias are prominent and thyroid cancer might be a rare component. One of its susceptibility genes is the regulatory subunit of protein kinase A. Over the course of the last four years, investigators have found the increasing clinical spectrum of syndromes characterized by germline loss-of-function PTEN mutation. In addition to CS, subsets of such disparate syndromes as Bannayan-Riley Ruvalcaba syndrome, Proteus syndrome, and possibly VATER with hydrocephalus and megencephaly with autistic features have been found to have germline PTEN mutations. Paradoxically, somatic intragenic PTEN mutations were rare in uncultured primary epithelial thyroid tumors, although hemizygous deletion occurred in 10-20% of thyroid adenomas and carcinomas. However, with subsequent study, it was discovered that epigenetic silencing of PTEN and perhaps inappropriate subcellular compartmentalization were two novel mechanisms of PTEN inactivation pertinent in thyroid carcinogenesis. Ectopic expression studies in vitro have borne out the importance of PTEN in the pathogenesis of epithelial thyroid neoplasias. PMID- 12119279 TI - Signaling pathways in adrenocortical cancer. AB - Adrenocortical carcinoma is a rare tumor that carries a very poor prognosis. Despite efforts to develop new therapeutic regimens to treat this disease, surgery remains the mainstay of treatment. Laboratory studies of adrenocortical cancers have revealed a wide variety of signaling pathways that can be altered in these neoplasms. Although ACTH signaling through adenylyl cyclase and protein kinase A is important for normal adrenal cellular physiology, there is evidence to suggest that this pathway may inhibit the growth of adrenocortical tumors, and that inactivation of the ACTH receptor may promote tumor formation. Although multiple signal transduction pathways are essential for normal adrenal growth and hormone secretion, efforts to identify events required for neoplastic transformation have met with limited success. Alterations that have frequently been observed in adrenocortical carcinoma include up-regulation of the IGF-II system, as well as mutations in TP53 and RAS. Current studies aim to elucidate the mechanisms of tumor growth by studying proproliferative signaling pathways, such as those involving Akt/PKB and the mitogen-activated protein kinases (MAPKs). Although studies of single pathways have been helpful in guiding investigations, new tools to study the integration and multiplicity of signaling pathways hold the hope of improved understanding of the signaling pathway alterations in adrenocortical cancer. PMID- 12119280 TI - Cyclic AMP-dependent signaling aberrations in macronodular adrenal disease. AB - The adrenal glands are a major source of steroid hormone biosynthesis. In normal physiology, the pituitary hormone corticotropin (ACTH) regulates the secretion of glucocorticoids via its G protein-coupled receptor (ACTHR), the product of the MC2R gene. Aldosterone is another major product of the adrenal gland; its regulation is controlled mainly by the renin-angiotensin system, although ACTH plays a role, too, especially under certain pathological conditions. The adrenal gland also secretes lesser amounts of androgens and intermediate metabolites of all these steroids. Unregulated secretion of any of these hormones can be caused by tumors, adrenocortical adenomas or carcinomas, and/or bilateral (or, rarely, unilateral) hyperplasia. Cortisol-producing hyperplasia of the adrenal glands is caused by two distinct syndromes, both of which have been directly or indirectly associated with protein kinase A signaling: (i) primary pigmented nodular adrenocortical disease (PPNAD) (a micronodular form of bilateral adrenal hyperplasia), either isolated (rarely) or in the context of Carney complex, is caused (in most cases) by mutations of the PRKAR1A gene; and (ii) ACTH independent macronodular adrenal hyperplasia (AIMAH), or massive macronodular adrenal disease (MMAD), has been associated with aberrant (ectopic) expression, and presumably regulation, of various G protein-coupled receptors. AIMAH is a rare, sporadic condition affecting predominantly middle-aged men and women with an almost equal ratio (the latter in contrast to other forms of endogenous Cushing's syndrome). Some familial cases of AIMAH have also been described, and it appears that the pathophysiological phenomena underlying AIMAH may be present in the far more common, sporadic adrenocortical tumors and, perhaps, in the nodular growth detected in the adrenal glands of the elderly in the general population. Thus, the study of ectopic receptor expression and cAMP-dependent PKA activity in AIMAH may have wider implications for adrenal and, indeed, endocrine tumorigenesis. PMID- 12119281 TI - Protein kinase A signaling: "cross-talk" with other pathways in endocrine cells. AB - Protein kinase A (PKA) signaling, in "classic" endocrine cell functioning, is known to mediate cAMP effects, generated through adenylate cyclase as a response to the activation of G protein-coupled receptors (GPCRs). This signaling system is highly versatile; its flexibility is supported by a number of adenylate cyclases, four PKA regulatory and three catalytic subunits, and several phosphodiesterases that close the negative feedback loop of cAMP generation, most molecules that are expressed in a tissue-specific manner. A central question, however, remains: how do the hundreds of GPCRs mediate their specific effects? Tissue specificity of the expression of the various components of the PKA system, albeit necessary, cannot be the only answer. It helps more to view PKA as a central hub that interacts with a variety of other signaling pathways in endocrine cells, not only mediating but also communicating cAMP effects to the mitogen-activated protein kinase (MAPK), protein kinase C and B (PKC and PKB/Akt, respectively). The net result of these complex interactions, evidence for which is reviewed in this chapter, is what we know as "cAMP effects." It is, perhaps, because of this complexity that investigations of PKA signaling in vivo and in vitro often give contradictory results and are difficult to interpret. PMID- 12119282 TI - Jab1 co-activation of c-Jun is abrogated by the serine 10-phosphorylated form of p27Kip1. AB - The cyclin-dependent kinase (cdk) inhibitor p27(Kip1) is a central mediator in the imposition and maintenance of quiescence through the sequestration of G(1) specific cyclin-cdk complexes. Previous studies have implicated the c-Jun co activator protein Jab1 as a regulator of intracellular p27(Kip1) levels. Jab1 has been reported to interact with p27(Kip1) and cause its translocation to the cytoplasm as a prelude to the degradation of the cdk inhibitor. Here we describe experiments that showing phosphorylation of p27(Kip1) at serine 10 leads to the suppression of Jab1 levels with the concomitant inhibition of c-Jun-dependent transcription. This repression is minimized upon quiescence exit through the rapid and preferential loss of the serine 10-phosphorylated form of p27(Kip1) following serum stimulation. Our results, therefore, demonstrate an additional role for p27(Kip1) in the modulation of c-Jun-dependent transcription via Jab1. PMID- 12119283 TI - Inhibitory PAS domain protein (IPAS) is a hypoxia-inducible splicing variant of the hypoxia-inducible factor-3alpha locus. AB - The inhibitory PAS (Per/Arnt/Sim) domain protein, IPAS, functions as a dominant negative regulator of hypoxia-inducible transcription factors (HIFs) by forming complexes with those proteins that fail to bind to hypoxia response elements of target genes. We have previously observed that IPAS is predominantly expressed in mice in Purkinje cells of the cerebellum and in corneal epithelium of the eye where it appears to play a role in negative regulation of angiogenesis and maintenance of an avascular phenotype. Sequencing of the mouse IPAS genomic structure revealed that IPAS is a splicing variant of the HIF-3alpha locus. Thus, in addition to three unique exons (1a, 4a, and 16) IPAS shares three exons (2, 4, and 5) with HIF-3alpha as well as alternatively spliced variants of exons 3 and 6. In experiments using normal mice and mice exposed to hypoxia (6% O(2)) for 6 h we observed alternative splicing of the HIF-3alpha transcript in the heart and lung. The alternatively spliced transcript was only observed under hypoxic conditions, thus defining a novel mechanism of hypoxia-dependent regulation of gene expression. Importantly, this mechanism may establish negative feedback loop regulation of adaptive responses to hypoxia/ischemia in these tissues. PMID- 12119284 TI - Gap junctions between cells expressing connexin 43 or 32 show inverse permselectivity to adenosine and ATP. AB - Gap junctions, composed of proteins from the connexin family, are the only channels that directly connect the cytoplasm of adjacent cells to allow for the intercellular transfer of small hydrophilic molecules. Gap junctional communication is essential for proper development and health in animals and humans. Whereas the study of biological molecules that pass through gap junctions is extremely important, the identification of endogenous transjunctional metabolites is challenging. To help address this problem, we have developed a layered culture system to identify and quantitate the transfer of endogenous molecules that pass between cells through gap junctions. Using these techniques, we have identified several endogenous molecules that showed differential transfer between channels composed of Cx32 versus Cx43. For example, adenosine passed about 12-fold better through channels formed by Cx32. In contrast, AMP and ADP passed about 8-fold better, and ATP greater than 300-fold better, through channels formed by Cx43. Thus, addition of phosphate to adenosine appears to shift its relative permeability from channels formed by Cx32 to channels formed by Cx43. This suggests functional consequence because the energy status of a cell could be controlled via connexin expression and channel formation. PMID- 12119285 TI - Human sterol 27-hydroxylase (CYP27) overexpressor transgenic mouse model. Evidence against 27-hydroxycholesterol as a critical regulator of cholesterol homeostasis. AB - CYP27-overexpressed transgenic mice were generated with the use of a human full length CYP27 coding region cloned into a ubiquitous expression vector. Positive transgenic mice were identified by tail DNA genotyping and high fecal 27 hydroxycholesterol content. The levels of 27-hydroxycholesterol were found to be 3-5 times higher in the circulation and the tissues of the overexpressed mice when compared with littermate controls. There were no gross morphological differences between the overexpressed mice and their controls. Total cholesterol and triglyceride levels were not affected by overexpression of CYP27. Serum lathosterol was also normal, suggesting a normal rate of cholesterol synthesis. Serum levels of 7alpha-hydroxycholesterol were unaffected, suggesting a normal rate of bile acid formation in the pathway involving cholesterol 7alpha hydroxylase. Biliary bile acid composition was slightly affected by CYP27 overexpression in female but not in male mice. Fecal levels of neutral steroids were slightly but significantly increased in overexpressor female mice but not in male mice. Levels of 24-hydroxycholesterol in the circulation were significantly reduced in the overexpressed mice, probably as a consequence of a recently described catabolic pathway involving CYP27. Combined with the results of our previous work on mice with a disruption of the CYP27 gene, the present results suggest that the levels of 27-hydroxycholesterol are not of critical importance for cholesterol homeostasis in mice. PMID- 12119286 TI - Effects of N-ethylmaleimide on conformational equilibria in purified cardiac muscarinic receptors. AB - Muscarinic receptors purified from porcine atria and devoid of G protein underwent a 9-27-fold decrease in their apparent affinity for the antagonists quinuclidinyl benzilate, N-methylscopolamine, and scopolamine when treated with the thiol-selective reagent N-ethylmaleimide. Their apparent affinity for the agonists carbachol and oxotremorine-M was unchanged. Conversely, the rate of alkylation by N-ethylmaleimide, as monitored by the binding of [(3)H]quinuclidinyl benzilate, was decreased by antagonists while agonists were without effect. The receptor also underwent a time-dependent inactivation that was hastened by N-ethylmaleimide but slowed by quinuclidinyl benzilate and N methylscopolamine. The destabilizing effect of N-ethylmaleimide was counteracted fully or nearly so at saturating concentrations of each antagonist and the agonist carbachol. Similar effects occurred with human M(2) receptors differentially tagged with the c-Myc and FLAG epitopes, coexpressed in Sf9 cells, and extracted in digitonin/cholate. The degree of coimmunoprecipitation was unchanged by N-ethylmaleimide, which therefore was without discernible effect on oligomeric size. The data are quantitatively consistent with a model in which the purified receptor from porcine atria interconverts spontaneously between two states (i.e. R R*). Antagonists favor the R state; agonists and N-ethylmaleimide favor the comparatively unstable R* state, which predominates after purification. Occupancy by a ligand stabilizes both states, and antagonists impede alkylation by favoring R over R*. Similarities with constitutively active receptors suggest that R and R* are akin to the inactive and active states, respectively. Purified M(2) receptors therefore appear to exist predominantly in their active state. PMID- 12119287 TI - Specificity of anion exchange mediated by mouse Slc26a6. AB - Recently, CFEX, the mouse orthologue of human SLC26A6, was localized to the brush border membrane of proximal tubule cells and was demonstrated to mediate Cl(-) formate exchange when expressed in Xenopus oocytes. The purpose of the present study was to examine whether mouse Slc26a6 can mediate one or more of the additional anion exchange processes observed to take place across the apical membrane of proximal tubule cells. Influx of [(14)C]formate into Slc26a6 expressing oocytes was inhibited by sulfate, oxalate, and p-aminohippurate (PAH), indicating affinity for these anions. Measurements of uptake of [(14)C]oxalate, [(14)C]PAH, and [(35)S]sulfate indicated that Slc26a6 can mediate transport of oxalate and sulfate but not PAH. Studies of the effect of external anions on [(14)C]oxalate efflux demonstrated Slc26a6-mediated Cl(-)-oxalate, oxalate formate, oxalate-oxalate, and oxalate-sulfate exchange. Two-electrode voltage clamp measurements indicated that Slc26a6-mediated Cl(-)-oxalate exchange is electrogenic. Intracellular pH recordings demonstrated that Slc26a6 can mediate Cl(-)-HCO(3)(-) exchange, but Cl(-)-OH(-) exchange was not detected. The presence of 100 microm oxalate inhibited the rate of Cl(-)-HCO(3)(-) exchange by 60%. We conclude that mouse Slc26a6 has affinity for oxalate, sulfate, and HCO(3)(-) in addition to Cl(-) and formate and can function in multiple exchange modes involving pairs of these anions. In the presence of high oxalate concentrations as found in renal tubular fluid and urine, Slc26a6 may largely function as an electrogenic Cl(-)-oxalate exchanger. PMID- 12119288 TI - Stimulation of enveloped virus infection by beta-amyloid fibrils. AB - Alzheimer's disease is characterized by deposition of beta-amyloid peptide (Abeta) into plaques in the brain, leading to neuronal toxicity and dementia. Human immunodeficiency virus type 1 (HIV-1) infection of the central nervous system can also cause a dementia, and amyloid deposition in the central nervous system is significantly higher in HIV-1-infected individuals compared with uninfected controls. Here we report that Abeta fibrils stimulated, by 5-20-fold, infection of target cells expressing CD4 and an appropriate coreceptor by multiple HIV-1 isolates but did not permit infection of cells lacking these receptors. Abeta enhanced infection at the stage of virus attachment or entry into the cell. Abeta fibrils also stimulated infection by amphotrophic Moloney leukemia virus, herpes simplex virus, and viruses pseudotyped with the envelope glycoprotein of vesicular stomatitis virus. Other synthetic fibril-forming peptides similarly enhanced viral infection and may be useful in gene delivery applications utilizing retroviral vectors. These data suggest that Abeta deposition may increase the vulnerability of the central nervous system to enveloped viral infection and that amyloidogenic peptides could be useful in enhancing gene transfer by enveloped viral vectors. PMID- 12119289 TI - Organization of the receptor-kinase signaling array that regulates Escherichia coli chemotaxis. AB - Motor behavior in prokaryotes is regulated by a phosphorelay network involving a histidine protein kinase, CheA, whose activity is controlled by a family of Type I membrane receptors. In a typical Escherichia coli cell, several thousand receptors are organized together with CheA and an Src homology 3-like protein, CheW, into complexes that tend to be localized at the cell poles. We found that these complexes have at least 6 receptors per CheA. CheW is not required for CheA binding to receptors, but is essential for kinase activation. The kinase activity per mole of bound CheA is proportional to the total bound CheW. Similar results were obtained with the E. coli serine receptor, Tsr, and the Salmonella typhimurium aspartate receptor, Tar. In the case of Tsr, under conditions optimal for kinase activation, the ratio of subunits in complexes is approximately 6 Tsr:4 CheW:1 CheA. Our results indicate that information from numerous receptors is integrated to control the activity of a relatively small number of kinase molecules. PMID- 12119290 TI - Subunit organization in a soluble complex of tar, CheW, and CheA by electron microscopy. AB - The Salmonella and Escherichia coli aspartate receptor, Tar, is representative of a large class of membrane receptors that generate chemotaxis responses by regulating the activity of an associated histidine protein kinase, CheA. Tar is composed of an NH(2)-terminal periplasmic ligand-binding domain linked through a transmembrane sequence to a COOH-terminal coiled-coil signaling domain in the cytoplasm. The isolated cytoplasmic domain of Tar fused to a leucine zipper sequence forms a soluble complex with CheA and the Src homology 3-like kinase activator, CheW. Activity of the CheA kinase in the soluble complex is essentially the same as in fully active complexes with the intact receptor in the membrane. The soluble complex is composed of approximately 28 receptor cytoplasmic domain chains, 6 CheW chains, and 4 CheA chains. It has a molecular weight of 1,400,000 (Liu, I., Levit, M., Lurz, R., Surette, M.G., and Stock, J.B. (1997) EMBO J. 16, 7231-7240). Electron microscopy reveals an elongated barrel like structure with a largely hollow center. Immunoelectron microscopy has provided a general picture of the subunit and domain organization of the complex. CheA and CheW appear to be in the middle of the complex with the leucine zippers of the receptor construct at the ends. These findings show that the receptor signaling complex forms higher ordered structures with defined geometric architectures. Coupled with atomic models of the subunits, our results provide insights into the functional architecture by which the receptor regulates CheA kinase activity during bacterial chemotaxis. PMID- 12119291 TI - Receptor methylation controls the magnitude of stimulus-response coupling in bacterial chemotaxis. AB - Motile prokaryotes employ a chemoreceptor-kinase array to sense changes in the media and properly adjust their swimming behavior. This array is composed of a family of Type I membrane receptors, a histidine protein kinase (CheA), and an Src homology 3-like protein (CheW). Binding of an attractant to the chemoreceptors inhibits CheA, which results in decreased phosphorylation of the chemotaxis response regulator (CheY). Sensitivity of the system to stimuli is modulated by a protein methyltransferase (CheR) and a protein methylesterase (CheB) that catalyze the methylation and demethylation of specific glutamyl residues in the cytoplasmic domain of the receptors. One of the most fundamental unanswered questions concerning the bacterial chemotaxis mechanism is the quantitative relationship between ligand binding to receptors and CheA inhibition. We show that the receptor glutamyl modifications cause adaptation by changing the gain (magnitude amplification) between attractant binding and kinase inhibition without substantially affecting ligand binding affinity. The mechanism adjusts receptor sensitivity to background stimulus intensity over several orders of magnitude of attractant concentrations. The cooperative effects of ligand binding appear to be minimal with Hill coefficients for kinase inhibition less than 2, independent of the state of glutamyl modification. PMID- 12119292 TI - cGMP-dependent protein kinase inhibits serum-response element-dependent transcription by inhibiting rho activation and functions. AB - RhoA, in its active GTP-bound form, stimulates transcription through activation of the serum-response factor (SRF). We found that cGMP inhibited serum-induced Rho.GTP loading and transcriptional activation of SRF-dependent reporter genes in smooth muscle and glial cells in a cGMP-dependent protein kinase (G-kinase) dependent fashion. Serum stimulation of the SRF target gene vinculin was also blocked by cGMP/G-kinase. G-kinase activation inhibited SRF-dependent transcription induced by upstream RhoA activators including Galpha(13) and p115RhoGEF, with Galpha(13)-induced Rho.GTP loading inhibited by G-kinase. G kinase had no effect on the high activation levels of RhoA(63L) or the double mutant RhoA(63L,188A) but inhibited transcriptional activation by these two RhoA mutants to a similar extent, suggesting an effect downstream of RhoA and independent of RhoA Ser(188) phosphorylation. Constitutively active forms of the Rho effectors Rho kinase (ROK), PKN, and PRK-2 induced SRF-dependent transcription in a cell type-specific fashion with ROK being the most efficient; G-kinase inhibited transcription induced by all three effectors without affecting ROK catalytic activity. G-kinase had no effect on RhoA(63L)-induced morphological changes in glial cells, suggesting distinct transcriptional and cytoskeletal effectors of RhoA. We conclude that G-kinase inhibits SRF-dependent transcription by interfering with RhoA signaling; G-kinase acts both upstream of RhoA, inhibiting serum- or Galpha(13)-induced Rho activation, and downstream of RhoA, inhibiting steps distal to the Rho targets ROK, PKN, and PRK-2. PMID- 12119293 TI - CH...O hydrogen bonds at protein-protein interfaces. AB - For the first time, a statistical potential has been developed to quantitatively describe the CH.O hydrogen bonding interaction at the protein-protein interface. The calculated energies of the CH.O pair interaction show a favorable valley at approximately 3.3 A, exhibiting a feature typical of an H-bond and similar to the ab initio quantum calculation result (Scheiner, S., Kar, T., and Gu, Y. (2001) J. Biol. Chem. 276, 9832-9837). The potentials have been applied to a set of 469 protein-protein complexes to calculate the contribution of different types of interactions to each protein complex: the average energy contribution of a conventional H-bond is approximately 30%; that of a CH.O H-bond is 17%; and that of a hydrophobic interaction is 50%. In some protein-protein complexes, the contribution of the CH.O H-bond can reach as high as approximately 40-50%, indicating the importance of the CH.O H-bond at the protein interface. At the interfaces of these complexes, C(alpha)H.O H-bonds frequently occur between adjacent strands in both parallel and antiparallel orientations, having the obvious structural motif of bifurcated H-bonds. Our study suggests that the weak CH.O H-bond makes an important contribution to the association and stability of protein complexes and needs more attention in protein-protein interaction studies. PMID- 12119294 TI - Basal transcription of the mouse sarco(endo)plasmic reticulum Ca2+-ATPase type 3 gene in endothelial cells is controlled by Ets-1 and Sp1. AB - We reported previously that the sarco(endo)plasmic reticulum Ca(2+)-ATPase type 3 (SERCA3) gene is expressed in many tissues and in a subset of cells such as endothelial, epithelial, and lymphoid lineages. Here we analyzed the mechanisms involved in the regulation of transcription of the SERCA3 gene in endothelial cells. The promoter of the murine SERCA3 gene was isolated, and a single transcription initiation site located 301 bp upstream of the translation initiation site was identified. Analysis of the transcriptional activity of fragments of the SERCA3 promoter showed the existence of a minimal promoter region located between bases -97 and +153 that contains one ETS-binding site (EBS) and two Sp1 elements that are essential for basal transcription. Mutation of the EBS or of the Sp1 sites abolished the basal activity of the promoter. We identified Ets-1 and Sp1 among endothelial nuclear factors that recognize the EBS and Sp1 sites on the promoter. Furthermore, transactivation of the -97/+301 promoter fragment by Ets-1 requires the presence of both the EBS and Sp1 sites, suggesting an interaction of the transcription factors on the gene promoter. Finally, overexpression of Ets-1 induced the expression of SERCA3 in endothelial cells and in fibroblasts. PMID- 12119295 TI - The N-terminal domain anchors human topoisomerase I at fibrillar centers of nucleoli and nucleolar organizer regions of mitotic chromosomes. AB - DNA topoisomerase I releases torsion stress created by DNA transcription. In principle, this activity is required in the nucleoplasm for mRNA synthesis and in the nucleoli for rRNA synthesis. Yet, topoisomerase I is mostly a nucleolar protein. Current belief holds that this preference is triggered by the N-terminal domain of the enzyme, which constitutes a nucleolar import signal. Contradicting this view, we show here that nucleolar accumulation of various fragments of topoisomerase I is correlated with their lesser mobility in this compartment and not with the N-terminal domain being intact or present. Therefore, the N-terminal domain is not likely a nucleolar import signal. We show that it rather serves as an adaptor that anchors a subpopulation of topoisomerase I at fibrillar centers of nucleoli and nucleolar organizer regions of mitotic chromosomes. Thus, it provides a steady association of topoisomerase I with the rDNA and with RNA polymerase I, which is maintained in a living cell during the entire cell cycle. PMID- 12119296 TI - Proteasome activity is required for androgen receptor transcriptional activity via regulation of androgen receptor nuclear translocation and interaction with coregulators in prostate cancer cells. AB - Upon binding to androgen, the androgen receptor (AR) can translocate into the nucleus and bind to androgen response element(s) to modulate its target genes. Here we have shown that MG132, a 26 S proteasome inhibitor, suppressed AR transactivation in an androgen-dependent manner in prostate cancer LNCaP and PC-3 cells. In contrast, MG132 showed no suppressive effect on glucocorticoid receptor transactivation. Additionally, transfection of PSMA7, a proteasome subunit, enhanced AR transactivation in a dose-dependent manner. The suppression of AR transactivation by MG132 may then result in the suppression of prostate-specific antigen, a well known marker used to monitor the progress of prostate cancer. Further mechanistic studies indicated that MG132 may suppress AR transactivation via inhibition of AR nuclear translocation and/or inhibition of interactions between AR and its coregulators, such as ARA70 or TIF2. Together, our data suggest that the proteasome system plays important roles in the regulation of AR activity in prostate cancer cells and may provide a unique target site for the development of therapeutic drugs to block androgen/AR-mediated prostate tumor growth. PMID- 12119297 TI - Peptide substrate specificities and protein cleavage sites of human endometase/matrilysin-2/matrix metalloproteinase-26. AB - Human endometase/matrilysin-2/matrix metalloproteinase-26 (MMP-26) is a novel epithelial and cancer-specific metalloproteinase. Peptide libraries were used to profile the substrate specificity of MMP-26 from the P4-P4' sites. The optimal cleavage motifs for MMP-26 were Lys-Pro-Ile/Leu-Ser(P1)-Leu/Met(P1')-Ile/Thr Ser/Ala-Ser. The strongest preference was observed at the P1' and P2 sites where hydrophobic residues were favored. Proline was preferred at P3, and Serine was preferred at P1. The overall specificity was similar to that of other MMPs with the exception that more flexibility was observed at P1, P2', and P3'. Accordingly, synthetic inhibitors of gelatinases and collagenases inhibited MMP 26 with similar efficacy. A pair of stereoisomers had only a 40-fold difference in K(i)(app) values against MMP-26 compared with a 250-fold difference against neutrophil collagenase, indicating that MMP-26 is less stereoselective for its inhibitors. MMP-26 autodigested itself during the folding process. Two of the major autolytic sites were Leu(49)-Thr(50) and Ala(75)-Leu(76), which still left the cysteine switch sequence (PHC(82)GVPD) intact. This suggests that Cys(82) may not play a role in the latency of the zymogen. Interestingly, inhibitor titration studies revealed that only approximately 5% of the total MMP-26 molecules was catalytically active, indicating that the thiol groups of Cys(82) in the active molecules may be dissociated or removed from the active site zinc ions. MMP-26 cleaved Phe(352)-Leu(353) and Pro(357)-Met(358) in the reactive loop of alpha(1) proteinase inhibitor and His(140)-Val(141) in insulin-like growth factor-binding protein-1, probably rendering these substrates inactive. Among the fluorescent peptide substrates analyzed, Mca-Pro-Leu-Ala-Nva-Dpa-Ala-Arg-NH(2) displayed the highest specificity constant (30,000/molar second) with MMP-26. This report proposes a working model for the future studies of pro-MMP-26 activation, the design of inhibitors, and the identification of optimal physiological and pathological substrates of MMP-26 in vivo. PMID- 12119299 TI - Severe abnormalities in the oral mucosa induced by suprabasal expression of epidermal keratin K10 in transgenic mice. AB - Previous studies have demonstrated that keratin K10 plays an important role in mediating cell signaling processes, since the ectopic expression of this keratin induces cell cycle arrest in proliferating cells in vitro and in vivo. However, apart from its well known function of providing epithelial cells with resilience to mechanical trauma, little is known about its possible roles in nondividing cells. To investigate what these might be, transgenic mice were generated in which the expression of K10 was driven by bovine K6beta gene control elements (bK6(beta)hK10). The transgenic mice displayed severe abnormalities in the tongue and palate but not in other K6-expressing cells such as those of the esophagus, nails, and hair follicles. The lesions in the tongue and palate included the cytolysis of epithelial suprabasal cells associated with an acute inflammatory response and lymphocyte infiltration. The alterations in the oral mucosa caused the death of transgenic pups soon after birth, probably because suckling was impaired. These anomalies, together with others found in the teeth, are reminiscent of the lesions observed in some patients with pachyonychia congenita, an inherited epithelial fragility associated with mutations in keratins K6 and K16. Although no epithelial fragility was observed in the bK6(beta)hK10 oral epithelia of the experimental mice, necrotic processes were seen. Collectively, these data show that the carefully regulated tissue- and differentiation-specific patterns displayed by the keratin genes have dramatic consequences on the biological behavior of epithelial cells and that changes in the specific composition of the keratin intermediate filament cytoskeleton can affect their physiology, in particular those of the oral mucosa. PMID- 12119298 TI - Presenilin 1 mutations activate gamma 42-secretase but reciprocally inhibit epsilon-secretase cleavage of amyloid precursor protein (APP) and S3-cleavage of notch. AB - The presenilin 1 (PS1) and presenilin 2 (PS2) proteins are necessary for proteolytic cleavage of the amyloid precursor protein (APP) within its transmembrane domain. One of these cleavage events (termed gamma-secretase) generates the C-terminal end of the Abeta-peptide by proteolysis near residue 710 or 712 of APP(770). Another event (termed gamma-like or epsilon-secretase cleavage) cleaves near residue 721 at approximately 2-5 residues inside the cytoplasmic membrane boundary to generate a series of stable, C-terminal APP fragments. This latter cleavage is analogous to S3-cleavage of Notch. We report here that specific mutations in the N terminus, loop, or C terminus of PS1 all increase the production of Abeta(42) but cause inhibition of both epsilon secretase cleavage of APP and S3-cleavage of Notch. These data support the hypothesis that epsilon-cleavage of APP and S3-cleavage of Notch are similar events. They also argue that, although both the gamma-site and the epsilon-site cleavage of APP are presenilin-dependent, they are likely to be independent catalytic events. PMID- 12119300 TI - Increased affinity and stability of an anti-HIV-1 envelope immunotoxin by structure-based mutagenesis. AB - HIV-infected cells are selectively killed by an immunotoxin in which a truncated form of Pseudomonas exotoxin A is joined to the variable region of a broadly neutralizing antibody (3B3) that recognizes the viral envelope glycoprotein (Env). To improve the efficacy of this molecule, we used three-dimensional structural information and phage selection data to design 23 single and multiple point mutations in the antibody variable region sequences that contact Env. Substituting an aromatic residue for an aspartate in the third complementarity determining region of V(H) increased the potency of the immunotoxin by approximately 10-fold in a cell-killing assay. Detailed analysis of one such mutant, N31H/Q100eY, revealed both a higher affinity for monomeric and cell surface Env and an increased stability against aggregation compared with the starting immunotoxin. Conversion to a disulfide-linked two-chain format further stabilized the protein. N31H/Q100eY retained the ability to bind to Env from multiple viral isolates, to inhibit Env-mediated cell fusion, and to limit spreading viral infection in peripheral blood mononuclear cells. Such site directed mutants may increase the utility of immunotoxins for reducing or eradicating persistent HIV-1 infection in humans. PMID- 12119302 TI - Albumin binding as a general strategy for improving the pharmacokinetics of proteins. AB - Plasma protein binding can be an effective means of improving the pharmacokinetic properties of otherwise short lived molecules. Using peptide phage display, we identified a series of peptides having the core sequence DICLPRWGCLW that specifically bind serum albumin from multiple species with high affinity. These peptides bind to albumin with 1:1 stoichiometry at a site distinct from known small molecule binding sites. Using surface plasmon resonance, the dissociation equilibrium constant of peptide SA21 (Ac-RLIEDICLPRWGCLWEDD-NH(2)) was determined to be 266 +/- 8, 320 +/- 22, and 467 +/- 47 nm for rat, rabbit, and human albumin, respectively. SA21 has an unusually long half-life of 2.3 h when injected by intravenous bolus into rabbits. A related sequence, fused to the anti tissue factor Fab of D3H44 (Presta, L., Sims, P., Meng, Y. G., Moran, P., Bullens, S., Bunting, S., Schoenfeld, J., Lowe, D., Lai, J., Rancatore, P., Iverson, M., Lim, A., Chisholm, V., Kelley, R. F., Riederer, M., and Kirchhofer, D. (2001) Thromb. Haemost. 85, 379-389), enabled the Fab to bind albumin with similar affinity to that of SA21 while retaining the ability of the Fab to bind tissue factor. This interaction with albumin resulted in reduced in vivo clearance of 25- and 58-fold in mice and rabbits, respectively, when compared with the wild-type D3H44 Fab. The half-life was extended 37-fold to 32.4 h in rabbits and 26-fold to 10.4 h in mice, achieving 25-43% of the albumin half-life in these animals. These half-lives exceed those of a Fab'(2) and are comparable with those seen for polyethylene glycol-conjugated Fab molecules, immunoadhesins, and albumin fusions, suggesting a novel and generic method for improving the pharmacokinetic properties of rapidly cleared proteins. PMID- 12119301 TI - Determinants of metabotropic glutamate receptor-5-mediated Ca2+ and inositol 1,4,5-trisphosphate oscillation frequency. Receptor density versus agonist concentration. AB - Diverse patterns of Ca(2+)(i) release differentially regulate Ca(2+)-sensitive enzymes and gene transcription, and generally the extent of agonist activation of phospholipase C-linked G protein-coupled receptors determines the type of Ca(2+) signal. We have studied global Ca(2+) oscillations arising through activation of the metabotropic glutamate receptor mGluR5a expressed in Chinese hamster ovary cells and find that these oscillations are largely insensitive to agonist concentration. Using an inducible receptor expression system and a non competitive antagonist, in conjunction with the translocation of eGFP PH(PLCdelta) to monitor inositol 1,4,5-trisphosphate (InsP(3)) oscillations in single cells, we show that mGluR5a density determines the frequency of these oscillations. The predominant underlying mechanism resulted from a negative feedback loop whereby protein kinase C (PKC) inhibited InsP(3) generation. Down regulation of PKC by prolonged exposure to phorbol ester revealed a second form of Ca(2+)(i) oscillation at low agonist concentrations. These Ca(2+)(i) signals showed features typical of classic repetitive Ca(2+)-induced Ca(2+) release and were sensitive to agonist concentration. Therefore, a single receptor can stimulate two types of InsP(3)-mediated Ca(2+) signal dependent upon feedback inhibition, producing two distinct means of controlling the final pattern of Ca(2+)(i) release. Our results have physiological implications for Ca(2+) signaling in general and emphasize the importance of mGluR5 surface expression for modulating synaptic plasticity. PMID- 12119304 TI - Klotho protein deficiency leads to overactivation of mu-calpain. AB - The klotho mouse is an animal model that prematurely shows phenotypes resembling human aging. Here we report that in homozygotes for the klotho mutation (kl(-/ )), alpha(II)-spectrin is highly cleaved, even before the occurrence of aging symptoms such as calcification and arteriosclerosis. Because alpha(II)-spectrin is susceptible to proteolysis by calpain, we examined the activation of calpain in kl(-/-) mice. m-Calpain was not activated, but mu-calpain was activated at an abnormally high level, and an endogenous inhibitor of calpain, calpastatin, was significantly decreased. Proteolysis of alpha(II)-spectrin increased with decreasing level of Klotho protein. Similar phenomena were observed in normal aged mice. Our results indicate that the abnormal activation of calpain due to the decrease of Klotho protein leads to degradation of cytoskeletal elements such as alpha(II)-spectrin. Such deterioration may trigger renal abnormalities in kl( /-) mice and aged mice, but Klotho protein may suppress these processes. PMID- 12119303 TI - ATPase activity of the MsbA lipid flippase of Escherichia coli. AB - Escherichia coli MsbA, the proposed inner membrane lipid flippase, is an essential ATP-binding cassette transporter protein with homology to mammalian multidrug resistance proteins. Depletion or loss of function of MsbA results in the accumulation of lipopolysaccharide and phospholipids in the inner membrane of E. coli. MsbA modified with an N-terminal hexahistidine tag was overexpressed, solubilized with a nonionic detergent, and purified by nickel affinity chromatography to approximately 95% purity. The ATPase activity of the purified protein was stimulated by phospholipids. When reconstituted into liposomes prepared from E. coli phospholipids, MsbA displayed an apparent K(m) of 878 microm and a V(max) of 37 nmol/min/mg for ATP hydrolysis in the presence of 10 mm Mg(2+). Preincubation of MsbA-containing liposomes with 3-deoxy-d mannooctulosonic acid (Kdo)(2)-lipid A increased the ATPase activity 4-5-fold, with half-maximal stimulation seen at 21 microm Kdo(2)-lipid A. Addition of Kdo(2)-lipid A increased the V(max) to 154 nmol/min/mg and decreased the K(m) to 379 microm. Stimulation was only seen with hexaacylated lipid A species and not with precursors, such as diacylated lipid X or tetraacylated lipid IV(A). MsbA containing the A270T substitution, which renders cells temperature-sensitive for growth and lipid export, displayed ATPase activity similar to that of the wild type protein at 30 degrees C but was significantly reduced at 42 degrees C. These results provide the first in vitro evidence that MsbA is a lipid-activated ATPase and that hexaacylated lipid A is an especially potent activator. PMID- 12119305 TI - A carboxyl-terminal PDZ-interacting domain of scavenger receptor B, type I is essential for cell surface expression in liver. AB - Scavenger receptor B, type I (SR-BI) was recently shown to interact with a PDZ domain-containing protein, PDZK1 (CLAMP/Diphor-1/CAP70/NaPi-Cap1), but the importance of this interaction in vivo in terms of SR-BI function has not been determined. In an effort to elucidate the role of this interaction in vivo, the PDZK1-interacting domain of SR-BI was identified and mutated and expressed liver specifically in mice. The PDZKI-interacting domain on SR-BI was identified as the last three carboxyl-terminal amino acids, Arg-Lys-Leu. A mutant SR-BI (SR BIdel509) that lacked only the leucine in the PDZ-interacting domain failed to interact with PDZK1 in vitro, while showing normal selective uptake function in nonpolarized cells. Transgenic mice with liver overexpression of SR-BIdel509 showed marked accumulation of SR-BI mRNA with only a moderate increase in SR-BI protein in liver, with no reduction in plasma cholesterol levels. Measurement of cell surface SR-BI levels and HDL cholesteryl ester-selective uptake in primary hepatocytes from transgenic mice revealed that SR-BIdel509 was not expressed at the plasma membrane correlating with normal levels of selective uptake compared with hepatocytes from nontransgenic littermates. This study indicates that the PDZK1-interacting domain of SR-BI is essential for cell surface expression of SR BI in liver and suggests that PDZK1 or other PDZ domain proteins may play an important role in regulating SR-BI cell surface expression and hence reverse cholesterol transport. PMID- 12119306 TI - A multidimensional meta-analysis of pharmacotherapy for bulimia nervosa: summarizing the range of outcomes in controlled clinical trials. AB - The empirical literature on pharmacotherapy for bulimia nervosa reveals mixed results. We examined the results of controlled clinical trials of pharmacotherapies for bulimia published from 1980 to 1999. To do this, we employed a multidimensional meta-analysis, a method for aggregating a range of clinically meaningful indicators of outcome (including but not limited to effect size estimates) across studies. We found that pharmacotherapy for bulimia yields a moderate initial effect. However, only a small minority of patients recover, and the average patient continues to meet full DSM-IV criteria for the disorder. Combined pharmacotherapy and short-term psychotherapy appears to produce better results, although most patients continue to show symptoms at termination, and few data are available on sustained recovery over time. In accordance with recent calls in the medical literature for standardization of reporting practices in clinical trials, we suggest that investigators and meta-analysts report a range of indices that bear on efficacy and generalizability to clinical practice. These include exclusion rates and reasons for exclusion, percentage recovered, percentage improved, percentage remaining improved or recovered at follow-up, and percentage seeking additional treatment at follow-up, as well as outcome data for both completer and intent-to-treat samples. PMID- 12119307 TI - Long-term neuropsychiatric consequences of "ecstasy" (MDMA): a review. AB - The recreational drug "ecstasy" (3,4-methylenedioxymethamphetamine, or MDMA) is widely used by young people throughout the world. Experimental studies indicate that MDMA damages serotonergic neurons in animals and possibly in humans. Repeated use may induce long-term neurotoxic effects, with cognitive and behavioral implications. We reviewed both the preclinical and the clinical literature to assess the evidence for persistent neuropsychiatric sequelae in humans. We focused on studies of chronic recreational use and reports of presence or absence of neurological, psychiatric, and psychological problems related to MDMA exposure. These investigations show repeated use of ecstasy to be associated with sleep, mood, and anxiety disturbances, elevated impulsiveness, memory deficits, and attention problems, which may persist for up to 2 years after cessation. In a subset of humans, particularly adolescents, depletion of serotonin by MDMA use may hasten or enhance vulnerability to a wide array of neuropsychiatric problems. Together, the studies reviewed provide substantial evidence that MDMA causes neuronal damage in animals and humans. Additional research is necessary to determine whether the MDMA-induced destruction of serotonergic neurons can have long-term and possibly permanent neuropsychiatric consequences in humans. PMID- 12119308 TI - Complementary and alternative medicine: what is its role? AB - Complementary and alternative medicine (CAM) remedies vary greatly in safety and effectiveness. Then why is their use increasing at a time when evidence for the effectiveness of "orthodox" treatments is greater than ever before? Dazzled by technology, physicians dismiss "nonspecific" treatment effects as mere "placebo" and ignore the effects of caring. Over half of patients with anxiety or depression consult CAM practitioners in any given year. The popularity of CAM attests to its responsiveness to patients' search for more than procedurally oriented care. It reflects biomedicine's failure to give patients the time they need to tell their story and to explain the nature of problems they face, and its failure to provide sufficient information to allow patients their choice among therapeutic alternatives. Effective CAM practices should be incorporated into care, whether treatments are provided by biomedical or CAM practitioners. PMID- 12119309 TI - Processing acute traumatic grief: exacerbation of posttraumatic stress disorder after September 11 in a 9-year-old boy. PMID- 12119310 TI - Psychiatric symptoms in metabolic and other genetic disorders: is our "organic" workup complete? PMID- 12119313 TI - Selective loss and preservation of biographical knowledge: implications for representation. PMID- 12119311 TI - The natural course of depression: Kraepelin and beyond. PMID- 12119314 TI - Selective, non-lateralized impairment of motor imagery following right parietal damage. AB - Using variants of a visually guided pointing task, in which subjects make pointing movements towards targets of varying sizes, we explored motor imagery in a patient with visual neglect. When this patient actually pointed towards targets of different sizes he showed the normal correlation between movement duration (MD) and target size, such that MD increased as target size decreased. In contrast, his imagined movements did not show the same speed-accuracy trade-off observed for actual movements. This was true regardless of the hand used or the initial direction of movement (left versus right). The patient performed normally on several tasks of visual imagery, including size estimation, perceptual discrimination and localization of cities on an imagined map. This patient's performance suggests that the networks in the right parietal lobe play an important role in the generation of internal models of motor movements regardless of the hand used to perform the task. PMID- 12119315 TI - Mixed lateralization of phonological assembly in developmental dyslexia. AB - Developmental phonological dyslexia has been characterized as a deficit in phonological assembly. At a neural level, it is possible that this deficit is represented by weak connectivity between anterior and posterior language systems in the left hemisphere. This study used 3-Tesla functional magnetic resonance imaging to investigate phonological assembly in a developmental phonological dyslexic. The phonological dyslexic showed increased activation in the left hemisphere of the inferior frontal gyrus (BA 44/6) and increased activation in the right hemisphere of the parietal cortex (BA 7), occipital cortex (BA 18), and in the cerebellum, as phonological demands were systematically increased. Converging evidence suggests that the core dysfunction in phonological dyslexia resides in and around the angular gyrus of the left hemisphere. This study supports the compensatory role of posterior regions in the right hemisphere together with the left inferior frontal gyrus. PMID- 12119316 TI - Does the spelling dyslexic read by recognizing orally spelled words? An investigation of a letter-by-letter reader. AB - In this single case report we describe our further observations of a patient (ROC) with a classical spelling dyslexia. A word length effect and a script effect were demonstrated. No evidence of semantic processing was obtained with brief tachistoscopic presentations. Single letter identification was unimpaired. In a series of experiments it was shown that her letter-by-letter reading strategy was mediated by explicit letter naming. In particular, it was observed that with successive presentation of the individual letters there was a stepwise function relating her whole word reading to the single letter presentation times. These durations permitted explicit letter naming. When explicit letter naming was prevented by a simultaneous articulatory suppression task, her letter-by-letter reading was significantly impaired. Her performance was compared with a second patient (MRF) who could read but could not spell. His reading was unaffected by articulatory suppression and significantly impaired by a successive presentation of the individual letters of a word. It is argued that explicit letter naming provides an input to the spelling system and thus can be recognized as orally spelled words. We conclude that in the reading-impaired patient there was a lexical deficit compensated for by the operation of a component of her intact spelling system. PMID- 12119317 TI - Dissociation between attentional set shifting and habit learning: a longitudinal case study. AB - We report a patient (ST) with predominant damage to the right neostriatum, caused by a rare cerebral angiitis. The testing procedure was focused on attentional set shifting (Wisconsin Card Sorting Test; WCST) and habit learning (probabilistic classification learning; PCL). ST showed impairments in the WCST, digit span backward, alphabet span, and PCL procedures, whereas he exhibited spared IQ, short-term verbal memory, object recognition, episodic and semantic memory. After 1 month of steroid therapy, there was a significant improvement in the WCST, digit span backward and alphabet span tests, whereas PCL remained severely impaired. The three control patients with damage to the parietal lobe displayed normal learning rates in PCL. These results suggest that separate frontostriatal mechanisms exist for attentional set shifting and habit learning. PMID- 12119319 TI - Shifting attention and joint attention dissociation in Williams syndrome: implications for the cerebellum and social deficits in autism. AB - An experimental paradigm that assesses one's capacity to perform intermodality attention shifting has proved to be sensitive for persons with cerebellar dysfunction. The basic experiment includes three conditions, auditory focus, visual focus and shift attention. In the auditory focus condition, the participant is instructed to press a joystick button when they hear the target tone and to ignore the other tone and the two visual stimuli. In the visual focus condition, the participant is instructed to press only the button to the target colored square and to ignore the other colored square and the two tones. In the shift attention condition, the participant is instructed to press the button to the first auditory target and then to press to the next visual target. They are instructed to continue to alternate their responses between auditory and visual targets until the trial is complete. Three individuals with Williams Syndrome (WMS), a genetic disorder due to the deletion of the elastin gene, were examined under these experimental conditions. Each participant with WMS had previously completed magnetic resonance imaging, and mid-sagittal area measurements had been made of the vermal lobules I-V and VI-VII. Cases were selected on the basis of cerebellar findings: one case was hypoplastic, one was hyperplastic and one had measurements in a range within one standard deviation of average for normal controls. Each of the WMS participants showed a pattern of being impaired in being able to shift their attention rapidly when cue-to-target intervals were less than 2.5 s. Their performance was very similar to previous reports of persons with cerebellar abnormalities and persons with autism. All three participants improved their target accuracy when given more time to shift their attention. The three participants did not experience performance deficits to either long or short cue-to-target intervals in the auditory focus or visual focus conditions. The results are consistent with the presence of cerebellar dysfunction, and are the first to suggest problems with shifting attention in persons with WMS. However, the three WMS participants demonstrated normal joint attention and had none of the social deficits observed in persons with autism. PMID- 12119320 TI - Is there a schizophasia? A study applying the single case approach to formal thought disorder in schizophrenia. AB - It has been suggested that formal thought disorder, the incoherent speech of schizophrenia, may involve a language disturbance among other abnormalities, or even be a form of dysphasia. Six patients with and seven without formal thought disorder were evaluated on an aphasia test battery. Spontaneous speech was also analysed using Brief Syntactic Analysis. Poor performance on the aphasia test battery was found to be associated with general intellectual impairment but not with formal thought disorder. Naming was preserved in both groups. Patients with formal thought disorder, but not those without, produced semantic errors in their spontaneous speech, and these were unrelated to general intellectual status. The disorder of language in formal thought disorder thus appears to be one of expressive semantic abnormality, which, however, spares naming. Further analysis of two intellectually preserved patients suggested that formal thought disorder may be associated with an additional difficulty in constructing an appropriate model for generating one's own speech. PMID- 12119323 TI - Internal iliac artery embolization with bilateral occlusion before endovascular aortoiliac aneurysm repair-clinical outcome of simultaneous and sequential intervention. AB - PURPOSE: To retrospectively evaluate the clinical outcome of patients after simultaneous or sequential internal iliac artery (IIA) embolization for bilateral IIA occlusion. MATERIALS AND METHODS: Sixteen patients (65-88 y; mean, 75.6 y; two women), 11 with aortobiiliac aneurysms, three with bilateral common iliac artery (CIA)/IIA aneurysms, and two with unilateral CIA/IIA aneurysms, underwent IIA occlusion before endovascular aortoiliac repair. Eight patients underwent simultaneous bilateral IIA embolization before endovascular aortic repair (EVAR). Eight patients had sequential bilateral IIA occlusion. The outcome was assessed by clinical follow-up. RESULTS: There were no severe ischemic complications such as buttock necrosis or acute bowel, bladder, or spinal cord ischemia. Early ischemic complications occurred in 25% (buttock/thigh claudication, n = 3, 18.8%; and sexual dysfunction, n = 1, 6.2%) and had an onset not later than 6 months after intervention: buttock claudication resolved (n = 2) or persisted after aggravation by inferior mesenteric artery embolization for type II endoleak (n = 1). Impotence in a fourth patient persisted. The ischemic complication rate after 6 months was 30% (three of 10) because of a fifth patient who developed ischemic colitis with aggravation of ischemic heart disease after 15 months. The mean follow-up duration was 19.7 months. Patients with simultaneous embolization had a lower complication rate than those with sequential embolization (one of eight [12.5%] vs four of eight [50%], respectively). CONCLUSIONS: IIA embolization for bilateral IIA occlusion can be performed with a complication rate comparable with results of previous studies of unilateral IIA embolization. Chronic buttock claudication may be aggravated by embolization of aortic side branches. Late complications can have an insidious course and be initiated by low-output cardiac failure. Bilateral IIA occlusion is recommended only in patients who are considered unfit for aortic surgery. PMID- 12119321 TI - Empathy deficits in Asperger syndrome: a cognitive profile. AB - Although lack of empathy has been considered a central characteristic of Asperger syndrome, quantitative and qualitative assessments of empathy in this syndrome are lacking. We present two cases of adolescents with Asperger syndrome who show extreme deficits on measures of both cognitive and affective empathy. Analysis of their performance on tasks assessing cognitive and affective processing did not reveal significant impairment in executive functions, nor in their ability to recognize emotions or the ability to create a mental representation of another person's knowledge. However, both patients were unable to integrate the emotional content with mental representations and deduce the other person's emotional state. These results suggest that impaired empathy in individuals with Asperger syndrome may be due to impaired integration of the cognitive and affective facets of the other person's mental state. PMID- 12119324 TI - Angiographic follow-up after suture-mediated femoral artery closure. AB - PURPOSE: Percutaneous closure devices are used in as many as 30% of all endovascular studies. Despite widespread use of these devices, only limited imaging has been performed after percutaneous closure. In this study, arteriograms of patients who had undergone suture-mediated closure with the Perclose device were reviewed. MATERIALS AND METHODS: Between June 1998 and November 2001, 31 patients who had previously undergone closure with use of the Perclose device at our institution returned for additional angiographic procedures. Twenty-one patients underwent closure with use of the Perclose device after embolization, including hepatic artery chemoembolization (n = 18), treatment of hypervascular sacral metastases (n = 2), and bronchial artery embolization (n = 1). Nineteen of these patients had thrombocytopenia. Ten patients underwent closure with use of the Perclose device after diagnosis and treatment of peripheral vascular disease. RESULTS: Of 31 patients, 28 had normal follow-up studies, including one patient who underwent four previous closures. These 28 patients all had normal femoral artery caliber at initial angiography and a platelet count of more than 18,000/mm(3). Two patients with preexisting atherosclerotic change had progression of disease at the puncture site and a third with severe thrombocytopenia developed a small asymptomatic posterolateral pseudoaneurysm. CONCLUSION: In patients with normal femoral arteries, the long term effects of closure with use of the Perclose device, even performed multiple times, appears to be minimal. PMID- 12119325 TI - Stent placement in the treatment of pulmonary artery stenosis secondary to fibrosing mediastinitis. PMID- 12119326 TI - Magnetic resonance imaging outcome after uterine artery embolization for leiomyomata with use of tris-acryl gelatin microspheres. AB - PURPOSE: To determine the imaging outcome after uterine artery embolization (UAE) with use of tris-acryl gelatin microspheres (Embospheres). MATERIALS AND METHODS: A retrospective analysis of magnetic resonance (MR) images was performed comparing studies completed 3-4 months after UAE to those performed before UAE. Twenty-three patients with 61 leiomyomata (as many as three fibroids per patient) were examined. Orthogonal T2, axial T1-weighted fat-saturated, sagittal T2 fast spin-echo, and dynamic T1-weighted sagittal images after Gadolinium injection were analyzed. Two abdominal imaging specialists examined the volume and uterine and leiomyoma perfusion. The Wilcoxon signed-rank test was used for inferences in leiomyoma size difference, infarction, and volume of fibroid tissue perfused. RESULTS: Median volume of all leiomyomata (n = 61) decreased by 52% (P <.001). For dominant fibroids alone (n = 23), a median 52% volume decrease (P <.001) was also noted, whereas the median uterine volume decreased 32%. Median perfused volume of all fibroids decreased from 31 mL to 0 mL, signifying a 100% decrease (P <.001). For dominant fibroids, a 100% median perfused volume decrease from 116 mL to 0 mL was noted (P <.001). Fifty-two of 61 fibroids (85%) and 20 of 23 dominant fibroids (87%) were completely devascularized and two fibroids disappeared. There was no myometrial ischemia identified. CONCLUSIONS: Tris-acryl gelatin microspheres (Embospheres) are an effective embolic agent for UAE, causing infarction and significant decrease in leiomyoma volume. PMID- 12119327 TI - Transcatheter arterial chemoembolization for hepatocellular carcinomas in patients with celiac axis occlusion. AB - PURPOSE: To verify the hypothesis that most instances of celiac axis occlusion in patients with hepatocellular carcinoma (HCC) are caused by diaphragmatic compression and, therefore, transcatheter arterial chemoembolization (TACE) can be performed through the compressed lumen of the celiac axis. MATERIALS AND METHODS: The authors attempted to perform TACE in 36 consecutive patients with HCC and celiac axis occlusion. Spiral computed tomographic (CT) images were available in 26 patients. Initially, catheterization of the hepatic arteries was attempted through the occluded celiac axis. If it failed, catheterization was performed through the pancreaticoduodenal arcades. The causes of celiac axis occlusion were evaluated based on spiral CT and angiographic findings, access routes, technical success rates, and related complications in superselective catheterization of hepatic arteries. RESULTS: Among the 26 patients who underwent spiral CT, diaphragmatic compression of the celiac axis was demonstrated in 23. Selective catheterization of hepatic arteries was possible through the occluded celiac axis in 23 patients (64%). In nine (25%) of the remaining 13 patients, TACE was performed through the dilated pancreaticoduodenal arcades from the superior mesenteric artery. As a procedure-related complication, celiac axis dissection occurred in one patient (3%). CONCLUSION: Most patients with celiac axis occlusion had arcuate ligament compression. In TACE, the celiac artery occlusion could be traversed directly and this should be the initial approach. PMID- 12119328 TI - High-resolution multiphase contrast-enhanced three-dimensional MR angiography compared with two-dimensional time-of-flight MR angiography for the identification of pedal vessels. AB - PURPOSE: The authors prospectively evaluated optimized multiphase high-resolution (HR) Gadolinium (Gd)-enhanced three-dimensional (3D) magnetic resonance (MR) angiography and standard two-dimensional (2D) time-of-flight (TOF) MR angiography for their ability to delineate distal calf and pedal vessels. MATERIALS AND METHODS: Twelve patients (20 limbs) with limb-threatening peripheral arterial occlusive disease underwent HR Gd-enhanced and 2D TOF MR angiography to identify targets for distal bypass. Imaging of the region of the ankle and foot was performed on a 1.5 T system with a head coil. A standard 2D TOF MR angiography sequence was performed first. The HR Gd-enhanced MR angiography sequence was then performed after injection of 0.01-0.2 mmol/kg of gadodiamide, allowing the acquisition of multiple consecutive coronal partitions, each in 18-25 seconds. Two experienced angiographers independently analyzed both studies. Comparison with intraoperative conventional angiography was available in 10 limbs. RESULTS: HR Gd-enhanced MR angiography allowed significantly faster imaging time (P <.0001) and larger coverage area (P <.0001) than 2D TOF MR angiography. All segments seen on 2D TOF MR angiography were visualized on HR Gd MR angiography, and significantly more suitable targets were seen well on HR Gd-enhanced MR angiography than on 2D TOF MR angiography (mean targets per limb: 3.9 +/- 1.9 vs 2.6 +/- 1.5, respectively; P =.02). In addition, HR Gd-enhanced MR angiography allowed better visualization of the arcuate pedal branch than 2D TOF MR angiography (P <.0001). Excellent correlation was demonstrated between HR Gd enhanced MR angiography and intraoperative angiography in 29 segments (binary similarity coefficient, 0.90). A significantly higher percentage of artifacts adversely affected image interpretation with 2D TOF MR angiography than with HR Gd-enhanced MR angiography (14 limbs vs five limbs, P <.001). Artifacts on HR Gd enhanced MR angiography included suboptimal mask in two limbs, venous contamination in one patient (two limbs), and motion artifact in one limb, although the studies remained diagnostic in all cases. CONCLUSION: HR Gd-enhanced MR angiography identified more distal target vessels with greater confidence than 2D TOF MR angiography. Optimized HR Gd-enhanced MR angiography may replace 2D TOF MR angiography as the gold standard examination for evaluation of distal runoff. PMID- 12119329 TI - Inhibition of vascular endothelial growth factor-mediated neointima progression with angiostatin or paclitaxel. AB - PURPOSE: Therapeutic angiogenesis represents a new paradigm for treatment of ischemic vascular syndromes. However, vascular endothelial growth factor (VEGF) enhances the rate and degree of plaque formation. This study evaluates the potential to block these effects nonspecifically with paclitaxel or specifically with angiostatin. MATERIALS AND METHODS: Recombinant human VEGF(165) (rhVEGF) was administrated intramuscularly (2-microg/kg single injection) in combination with adventitial delivery of paclitaxel, angiostatin, or vehicle alone at the site of femoral arterial balloon overdilation injury in New Zealand White rabbits (n = 5 per treatment). Five additional animals with no rhVEGF and no adventitial delivery served as procedural controls. All rabbits were fed according to a 0.25% cholesterol diet beginning 28 days before angioplasty. Treated arteries were harvested after 7 days and evaluated to determine intima-to-media (I/M) ratios, macrophage infiltrate, and endothelial cell density. RESULTS: On histologic analysis, the rhVEGF/gel control group exhibited a mean I/M ratio of 0.337 +/- 0.028, a 77% increase over procedural controls, which exhibited a mean I/M of 0.190 +/- 0.010. rhVEGF/paclitaxel reduced I/M ratios to 0.151 +/- 0.007. In contrast, specific antiangiogenic therapy (rhVEGF/angiostatin) reduced I/M ratios to 0.032 +/- 0.003, a 91% decrease relative to rhVEGF/gel and an 83% decrease relative to procedural controls (P =.001 for each comparison). Local macrophages and endothelial cells also decreased with treatment. CONCLUSIONS: This study shows that paclitaxel and angiostatin each afford local protection against rhVEGF mediated increases in neointima. Angiostatin further prevents progression of underlying neointima. These local therapies may allow broader use of therapeutic angiogenesis while avoiding and treating potentially undesirable effects. PMID- 12119330 TI - In vivo vascular engineering of vein grafts: directed migration of smooth muscle cells by perivascular release of elastase limits neointimal proliferation. AB - PURPOSE: Saphenous vein bypass grafting for coronary revascularization procedures remains limited by accelerated neointima formation. It was hypothesized that creation of a modified chemotactic gradient in vivo could guide migration of smooth muscle cells (SMCs) peripherally instead of in a luminal direction and reduce intimal hyperplasia during vein graft arterialization. MATERIALS AND METHODS: Surgical bypass vein grafting to femoral arteries was performed in adult male New Zealand White rabbits (n = 8 per treatment group; five for 7 d and three for 28 d). Controlled-release microspheres delivering elastase or buffered polymer only were administered perivascularly at the vein graft site. At 7 days, five vein grafts per group were harvested and cross-sections were immunostained with anti-proliferating cell nuclear antigen (PCNA) to determine the number and distribution of proliferating SMCs. At 28 days, three vein grafts per group were harvested and intima-to-media (I/M) ratios were calculated after staining with Verhoeff von Gieson-Masson trichrome stain. RESULTS: Significant early outward directed elastin degradation resulted from elastase treatment. Concurrently, proliferating SMCs migrated peripherally. PCNA(+) cells in the outer half of the wall increased 2.37 fold compared to procedural controls (P <.0001). Directional shifts in SMC migration underlie these results because overall SMC proliferation was not significantly different. At 28 days after vein graft surgery, a 38% reduction (P =.0008) in neointima was observed relative to procedural controls. CONCLUSION: Directional guidance of SMC responses through perivascular elastase release achieves favorable vein graft remodeling characteristics, including limited neointima development. This represents practical evidence that SMC migration can be directionally guided in vivo in a vein graft model and that plaque progression can be prevented by redistributing elastin without decreasing functional vein graft wall stability. PMID- 12119331 TI - Improved coagulation with saline solution pretreatment during radiofrequency tumor ablation in a canine model. AB - PURPOSE: To determine whether pretreatment with local NaCl injection can increase radiofrequency (RF)-induced coagulation in a large animal model. MATERIAL AND METHODS: Multiple canine venereal sarcomas (n = 25) were implanted subcutaneously in eight mildly immunosuppressed dogs (25 mg/kg cyclosporin A twice daily). Tumors were incubated for 8-12 weeks to a diameter of 4.2-6.3 cm (5.1 cm +/- 0.7). Internally cooled RF ablation (1-cm tip; 12 min; pulsed technique; 2,000-mA maximum) was performed. Tumors were pretreated with 6 mL of 18%, 24%, or 36% NaCl injected intratumorally under direct ultrasound guidance after RF electrode insertion, and this treatment was compared to RF treatment without NaCl injection and to 36% NaCl injection without RF ablation. Impedance measurements and remote thermometry were performed. These measurements and resultant coagulation were compared. RESULTS: Significantly greater RF heating (73 degrees C +/- 11 degrees C at 20 mm) was observed when the tumors were treated with 24% or 36% NaCl pretreatment, compared to the 47 degrees C +/- 5 degrees C observed when 18% or no NaCl was injected (P <.02). In the 36% NaCl group, the entire tumor (5.2 cm +/ 0.8 diameter) was completely ablated in every case, with coagulation extending several centimeters into the surrounding tissues. By comparison, control tumors (without NaCl injection) contained coagulation measuring 3.1 cm +/- 0.2, surrounded by viable, well-perfused tumor (P <.01), and 36% NaCl alone produced 2.7 cm +/- 0.6 of patchy necrosis. CONCLUSIONS: Pretreatment with intratumoral injection of small volumes of highly concentrated NaCl markedly increases RF heating and coagulation in a large animal tumor model. The complete destruction of tumors 5 cm in diameter or larger suggests that this substantial increase may be achieved for tumor ablation in clinical practice. PMID- 12119332 TI - Embolization of profunda femoris artery branch pseudoaneurysms with ethylene vinyl alcohol copolymer (onyx). AB - This report describes the use of a new embolic agent, Onyx, which is composed of ethylene vinyl alcohol copolymer dissolved in 8% dimethyl sulfoxide, in the endovascular transcatheter embolization of traumatic profunda femoris artery branch pseudoaneurysms in three cases. Two of the cases involved massive thigh swelling after penetrating injury and the other involved massive hemorrhage at the site of a surgical fixator pin. Diagnostic angiography revealed pseudoaneurysm formation of the profunda femoris artery branches in all cases. Immediate control angiography after the embolization procedures demonstrated complete closure of the pseudoaneurysms. During the follow-up period there was no recurrent bleeding. The advantages of Onyx over conventional liquid embolic agents and coils are discussed. PMID- 12119333 TI - Iliac artery kinking with endovascular therapies: technical considerations. AB - Iliac artery tortuosity should be considered when planning endovascular interventions from a femoral approach. Stiff guide wires across tortuous iliac segments can introduce foreshortening and temporary kinking. Recognition of this phenomenon and its implications is important when making anatomic measurements before endovascular device placement, when assessing iliac runoff, and when considering adjunctive procedures after aortoiliac interventions. Two illustrative cases of external iliac artery kinking are presented, one during an abdominal aortic aneurysm endograft procedure and another encountered during stent placement in an external iliac artery dissection. In both cases, the temporary nature of the deformity was recognized, avoiding unnecessary additional intervention. PMID- 12119334 TI - Detection of room air contamination of angiographic CO2 with use of a gas analyzer. AB - The purpose of this study was to describe a practical method to detect room air contamination in CO2 used for angiography. Samples of CO2 with known room air contamination levels were used in a "bag system" of CO2 delivery and sampled by a gas analyzer commonly used in anesthesia. Nitrogen levels were reliably detected indicating contamination with as little as 2% air. Oxygen levels were reliably detected, indicating contamination with as little as 5% air. Measured CO2 values were unreliable with higher-than-true values at all levels except 100%. All clinically important amounts of N2 and O2 contamination were readily detected by this practical method. PMID- 12119335 TI - Selective spermatic venography and varicocele embolization in men with circumaortic left renal veins. AB - The presence of variant venous anatomy may increase the technical difficulty of percutaneous varicocele embolization. The authors review their experience performing venography of the left spermatic vein and varicocele embolization in 18 men with circumaortic left renal veins. Selective catheterization of the left spermatic vein was achieved in all but one patient, and all patients with positive venographic results underwent successful embolization. The site of the confluence of the left spermatic vein and the renal vein was variable and it was necessary to use jugular and femoral venous approaches to achieve selective left spermatic vein catheterization. Familiarity with anatomic variations associated with circumaortic renal veins and with embolization techniques from jugular and femoral venous routes facilitates percutaneous varicocele embolization in patients with this variant. PMID- 12119336 TI - Transcatheter embolization of an aortobronchial fistula with N-butyl cyanoacrylate. AB - Aortobronchial fistula is a fatal complication after thoracic aortic surgery. So far, treatment options for aortobronchial fistula have included surgical and endovascular stent-graft methods. Herein, a case of an aortobronchial fistula with life-threatening hemoptysis managed with transcatheter embolization of the fistula with N-butyl cyanoacrylate is reported. For the patient with an aortobronchial fistula who cannot be treated by surgical or endovascular stent graft methods, transcatheter embolization of the fistula may be the only available life-saving method. PMID- 12119337 TI - Staged endovascular occlusion of giant idiopathic renal arteriovenous fistula with platinum microcoils and silk suture threads. AB - Large symptomatic aneurysmal renal arteriovenous (AV) fistulas usually present with a flank bruit, uncontrolled hypertension, and high-output cardiac failure. Traditionally, these have been treated surgically because of the risk of inadvertent pulmonary embolism with use of embolization techniques. The authors report a case of a giant renal AV fistula successfully treated with staged embolization with use of metallic coils and silk suture. This resulted in a graded reduction in the extreme flow through the fistula, followed by delayed thrombosis and cure of the patient's symptoms. PMID- 12119338 TI - Common iliac artery occlusion with use of Gianturco coils and ethylene vinyl alcohol liquid embolization agent before aortouniiliac stent-graft deployment. PMID- 12119339 TI - Transjugular intrahepatic portosystemic shunt creation in a patient with situs inversus. PMID- 12119340 TI - Implications of phase variation of a gene (pgtA) encoding a pilin galactosyl transferase in gonococcal pathogenesis. AB - The pilin glycoprotein (PilE) is the main building block of the pilus of Neisseria gonorrhoeae (gonococcus [GC]). GC pilin is known to carry a disaccharide O-glycan, which has an alphaGal attached to the O-linked GlcNAc by a 1-3 glycosidic bond. In this report, we describe the cloning and characterization of the GC gene, pilus glycosyl transferase A (pgtA), which encodes the galactosyl transferase that catalyzes the synthesis of this Gal-GlcNAc bond of pilin glycan. A homopolymeric tract of Gs (poly-G) is present in the pgtA gene of many GC strains, and this pgtA with poly-G can undergo phase variation (Pv). However, in many other GC, pgtA lacks the poly-G and is expressed constitutively without Pv. Furthermore, by screening a large number of clinical isolates, a significant correlation was observed between the presence of poly-G in pgtA and the dissemination of GC infection. Poly-G was found in pgtA in all (24 out of 24) of the isolates from patients with disseminated gonococcal infection (DGI). In contrast, for the vast majority (20 out of 28) of GC isolated from uncomplicated gonorrhea (UG) patients, pgtA lacked the poly-G. These results indicate that Pv of pgtA is likely to be involved in the conversion of UG to DGI. PMID- 12119341 TI - The developmentally regulated expression of Twisted gastrulation reveals a role for bone morphogenetic proteins in the control of T cell development. AB - The evolutionarily conserved, secreted protein Twisted gastrulation (Tsg) modulates morphogenetic effects of decapentaplegic (dpp) and its orthologs, the bone morphogenetic proteins 2 and 4 (BMP2/4), in early Drosophila and vertebrate embryos. We have uncovered a role for Tsg at a much later stage of mammalian development, during T cell differentiation in the thymus. BMP4 is expressed by thymic stroma and inhibits the proliferation of CD4(-)CD8(-) double-negative (DN) thymocytes and their differentiation to the CD4(+)CD8(+) double-positive (DP) stage in vitro. Tsg is expressed by thymocytes and up-regulated after T cell receptor signaling at two developmental checkpoints, the transition from the DN to the DP and from the DP to the CD4(+) or CD8(+) single-positive stage. Tsg can synergize with the BMP inhibitor chordin to block the BMP4-mediated inhibition of thymocyte proliferation and differentiation. These data suggest that the developmentally regulated expression of Tsg may allow thymocytes to temporarily withdraw from inhibitory BMP signals. PMID- 12119342 TI - Identification of the lateral interaction surfaces of human histocompatibility leukocyte antigen (HLA)-DM with HLA-DR1 by formation of tethered complexes that present enhanced HLA-DM catalysis. AB - Human histocompatibility leukocyte antigen (HLA)-DM is a major histocompatibility complex (MHC)-like protein that catalyzes exchange of antigenic peptides from MHC class II molecules. To investigate the molecular details of this catalysis we created four covalent complexes between HLA-DM and the MHC class II allele DR1. We introduced a disulfide bond between the naturally occurring cysteine beta46 on HLA-DM and an engineered cysteine on the end of a linker attached to either the NH(2)- or the COOH terminus of an antigenic peptide that is tightly bound on DR1. We find that when DM is attached to the NH(2) terminus of the peptide, it can, for all linker lengths tested, catalyze exchange of the peptide with a half-life a few minutes (compared with uncatalyzed t(1/2) > 100 h). This rate, which is several orders of magnitude greater than the one we obtain in solution assays using micromolar concentrations of HLA-DM, is dominated by a concentration independent factor, indicating an intramolecular catalytic interaction within the complex. A similar complex formed at the COOH terminus of the peptide shows no sign of DM-specific intramolecular catalysis. Restrictions on the possible interaction sites imposed by the length of the linkers indicate that the face of DR1 that accommodates the NH(2) terminus of the antigenic peptide interacts with the lateral face of HLA-DM that contains cysteine beta46. PMID- 12119343 TI - Two distinct pathways mediated by PA28 and hsp90 in major histocompatibility complex class I antigen processing. AB - Major histocompatibility complex (MHC) class I ligands are mainly produced by the proteasome. Herein, we show that the processing of antigens is regulated by two distinct pathways, one requiring PA28 and the other hsp90. Both hsp90 and PA28 enhanced the antigen processing of ovalbumin (OVA). Geldanamycin, an inhibitor of hsp90, almost completely suppressed OVA antigen presentation in PA28alpha(-/ )/beta(-/-) lipopolysaccharide blasts, but not in wild-type cells, indicating that hsp90 compensates for the loss of PA28 and is essential in the PA28 independent pathway. In contrast, treatment of cells with interferon (IFN)-gamma, which induces PA28 expression, abrogated the requirement of hsp90, suggesting that IFN-gamma enhances the PA28-dependent pathway, whereas it diminishes hsp90 dependent pathway. Importantly, IFN-gamma did not induce MHC class I expressions in PA28-deficient cells, indicating a prominent role for PA28 in IFN-gamma stimulated peptide supply. Thus, these two pathways operate either redundantly or specifically, depending on antigen species and cell type. PMID- 12119344 TI - Delayed cellular maturation and decreased immunoglobulin kappa light chain production in immature B lymphocytes lacking B cell linker protein. AB - B cell linker (BLNK) protein is a component of the B cell receptor (BCR) signaling pathway and BLNK(-/-) mice have a block in B lymphopoiesis at the pro B/pre-B cell stage. To study the effect of BLNK mutation at later stages of B cell development, we introduce an innocuous transgenic BCR into BLNK(-/-) mice and show that two populations of immature B cells distinguishable by their IgM(low (lo)) and IgM(high (hi)) phenotypes are found in the bone marrow of these mice in contrast to a single population of IgM(hi) cells found in control BCR transgenic BLNK(+/+) mice. The mutant IgM(lo) and IgM(hi) cells are at an earlier developmental stage compared with the control IgM(hi) cells as indicated by their differential expression of CD43, B220, and major histocompatibility complex class II antigens and their timing of generation in culture. Thus, in the absence of BLNK the differentiation of immature B cells is delayed. Furthermore, mutant IgM(lo) cells produce equivalent level of immunoglobulin (Ig) mu but less Ig kappa proteins than control and mutant IgM(hi) cells and this defect is attributed to a decrease in the amount of kappa transcripts being generated. Finally, splenic B cells in BCR-transgenic BLNK(-/-) mice are predominantly of the transitional B cell phenotype and are rapidly lost from the peripheral B cell pool. Taken together, the data suggest a role for BLNK and perhaps BCR signaling, in the regulation of kappa light chain expression and continued immature B cell differentiation. PMID- 12119345 TI - Degeneracy of antigen recognition as the molecular basis for the high frequency of naive A2/Melan-a peptide multimer(+) CD8(+) T cells in humans. AB - In contrast with the low frequency of most single epitope reactive T cells in the preimmune repertoire, up to 1 of 1,000 naive CD8(+) T cells from A2(+) individuals specifically bind fluorescent A2/peptide multimers incorporating the A27L analogue of the immunodominant 26-35 peptide from the melanocyte differentiation and melanoma associated antigen Melan-A. This represents the only naive antigen-specific T cell repertoire accessible to direct analysis in humans up to date. To get insight into the molecular basis for the selection and maintenance of such an abundant repertoire, we analyzed the functional diversity of T cells composing this repertoire ex vivo at the clonal level. Surprisingly, we found a significant proportion of multimer(+) clonotypes that failed to recognize both Melan-A analogue and parental peptides in a functional assay but efficiently recognized peptides from proteins of self- or pathogen origin selected for their potential functional cross-reactivity with Melan-A. Consistent with these data, multimers incorporating some of the most frequently recognized peptides specifically stained a proportion of naive CD8(+) T cells similar to that observed with Melan-A multimers. Altogether these results indicate that the high frequency of Melan-A multimer(+) T cells can be explained by the existence of largely cross-reactive subsets of naive CD8(+) T cells displaying multiple specificities. PMID- 12119346 TI - On the role of dendritic cells in peripheral T cell tolerance and modulation of autoimmunity. AB - Recently, it has become clear that dendritic cells (DCs) are essential for the priming of T cell responses. However, their role in the maintenance of peripheral T cell tolerance remains largely undefined. Herein, an antigen-presenting cell (APC) transfer system was devised and applied to experimental allergic encephalomyelitis (EAE), to evaluate the contribution that DCs play in peripheral T cell tolerance. The CD8alpha(-)CD4(+) subset, a minor population among splenic DCs, was found to mediate both tolerance and bystander suppression against diverse T cell specificities. Aggregated (agg) Ig-myelin oligodendrocyte glycoprotein (MOG), an Ig chimera carrying the MOG 35-55 peptide, binds and cross links FcgammaR on APC leading to efficient peptide presentation and interleukin (IL)-10 production. Furthermore, administration of agg Ig-MOG into diseased mice induces relief from clinical EAE involving multiple epitopes. Such recovery could not occur in FcgammaR-deficient mice where both uptake of Ig-MOG and IL-10 production are compromised. However, reconstitution of these mice with DC populations incorporating the CD8alpha(-)CD4(+) subset restored Ig-MOG-mediated reversal of EAE. Transfer of CD8alpha(+) or even CD8alpha(-)CD4(-) DCs had no effect on the disease. These findings strongly implicate DCs in peripheral tolerance and emphasize their functional potency, as a small population of DCs was able to support effective suppression of autoimmunity. PMID- 12119347 TI - Antigen-independent induction of histamine synthesis by immunoglobulin E in mouse bone marrow-derived mast cells. AB - Immunoglobulin (Ig)E-mediated activation of mast cells has long been thought to occur only when Fc(epsilon)RI receptor-bound IgE is cross-linked via multivalent antigens. However, recent studies have raised the possibility that mast cells may be activated by the binding of IgE to the Fc(epsilon)RI receptor in the absence of antigen. Here we demonstrate that IgE binding without antigen induces the expression of histidine decarboxylase (HDC) in mouse interleukin (IL)-3-dependent bone marrow-derived mast cells (BMMCs). The induction of HDC by the binding of IgE was found to require an influx of extracellular calcium ions, which was attenuated by pretreatment with U73122, a phospholipase C inhibitor. Furthermore, the increase in HDC activity upon sensitization with IgE was completely suppressed by pretreatment of BMMCs with protein kinase C inhibitors, such as H7, staurosporine, and Go6976. In addition, immediate activation of the tyrosine kinase Lyn was not detectable upon treatment with IgE. These results suggest that the binding of IgE to its receptor in the absence of antigen results in de novo synthesis of HDC in BMMCs through a signaling pathway distinct to that operating during antigen-stimulated Fc(epsilon)RI activation. PMID- 12119348 TI - CD4(+)CD25(+) regulatory T cells can mediate suppressor function in the absence of transforming growth factor beta1 production and responsiveness. AB - CD4(+)CD25(+) regulatory T cells inhibit organ-specific autoimmune diseases induced by CD4(+)CD25(-) T cells and are potent suppressors of T cell activation in vitro. Their mechanism of suppression remains unknown, but most in vitro studies suggest that it is cell contact-dependent and cytokine independent. The role of TGF-beta1 in CD4(+)CD25(+) suppressor function remains unclear. While most studies have failed to reverse suppression with anti-transforming growth factor (TGF)-beta1 in vitro, one recent study has reported that CD4(+)CD25(+) T cells express cell surface TGF-beta1 and that suppression can be completely abrogated by high concentrations of anti-TGF-beta suggesting that cell-associated TGF-beta1 was the primary effector of CD4(+)CD25(+)-mediated suppression. Here, we have reevaluated the role of TGF-beta1 in CD4(+)CD25(+)-mediated suppression. Neutralization of TGF-beta1 with either monoclonal antibody (mAb) or soluble TGF betaRII-Fc did not reverse in vitro suppression mediated by resting or activated CD4(+)CD25(+) T cells. Responder T cells from Smad3(-/-) or dominant-negative TGF beta type RII transgenic (DNRIITg) mice, that are both unresponsive to TGF-beta1 induced growth arrest, were as susceptible to CD4(+)CD25(+)-mediated suppression as T cells from wild-type mice. Furthermore, CD4(+)CD25(+) T cells from neonatal TGF-beta1(-/-) mice were as suppressive as CD4(+)CD25(+) from TGF-beta1(+/+) mice. Collectively, these results demonstrate that CD4(+)CD25(+) suppressor function can occur independently of TGF-beta1. PMID- 12119349 TI - Human CD4(+)CD25(+) regulatory, contact-dependent T cells induce interleukin 10 producing, contact-independent type 1-like regulatory T cells [corrected]. AB - It has been recently demonstrated that regulatory CD4(+)CD25(+) CD45RO(+) T cells are present in the peripheral blood of healthy adults and exert regulatory function similar to their rodent counterparts. It remains difficult to understand how the small fraction of these T cells that regulate via direct cell-to-cell contact and not via secretion of immunosuppressive cytokines could mediate strong immune suppression. Here we show that human CD4(+)CD25(+) T cells induce long lasting anergy and production of interleukin (IL)-10 in CD4(+)CD25(-) T cells. These anergized CD4(+)CD25(-) T cells then suppress proliferation of syngenic CD4(+) T cells via IL-10 but independent of direct cell contact, similar to the so-called type 1 regulatory T (Tr1) cells. This 'catalytic' function of CD4(+)CD25(+) T cells to induce Tr1-like cells helps to explain their central role for the maintenance of immune homeostasis. PMID- 12119350 TI - Infectious tolerance: human CD25(+) regulatory T cells convey suppressor activity to conventional CD4(+) T helper cells. AB - Regulatory CD4(+)CD25(+) T cells (Treg) are mandatory for maintaining immunologic self-tolerance. We demonstrate that the cell-cell contact-mediated suppression of conventional CD4(+) T cells by human CD25(+) Treg cells is fixation resistant, independent from membrane-bound TGF-beta but requires activation and protein synthesis of CD25(+) Treg cells. Coactivation of CD25(+) Treg cells with Treg cell-depleted CD4(+) T cells results in anergized CD4(+) T cells that in turn inhibit the activation of conventional, freshly isolated CD4(+) T helper (Th) cells. This infectious suppressive activity, transferred from CD25(+) Treg cells via cell contact, is cell contact-independent and partially mediated by soluble transforming growth factor (TGF)-beta. The induction of suppressive properties in conventional CD4(+) Th cells represents a mechanism underlying the phenomenon of infectious tolerance. This explains previously published conflicting data on the role of TGF-beta in CD25(+) Treg cell-induced immunosuppression. PMID- 12119351 TI - The immune regulatory function of lymphoproliferative double negative T cells in vitro and in vivo. AB - Lymphoproliferative (lpr) mice, which lack functional Fas receptor expression and develop autoimmune lymphoproliferative disease, have an accumulation of T cell receptor-alphabeta(+)CD4(-)CD8(-) (double negative T cells [DNTC]) in the periphery. The function of the accumulating DNTC is not clear. In this study we demonstrate that B6/lpr DNTC can dose dependently kill syngeneic CD8(+) and CD4(+) T cells from wild-type B6 mice through Fas/Fas ligand interactions in vitro. We also demonstrate that B6/lpr DNTC that are activated and expand in vivo are able to specifically down-regulate allogeneic immune responses mediated by syngeneic Fas(+)CD4(+) and CD8(+) T cells in vivo. B6/lpr DNTC that have been preactivated in vivo by infusion of either class I- (bm1) or class II- (bm12) mismatched allogeneic lymphocytes are able to specifically enhance the survival of bm1 or bm12, but not third-party skin allografts when adoptively transferred into naive B6(+/+) mice. These findings clearly demonstrate that B6/lpr DNTC have a potent immune regulatory function in vitro and in vivo. They also provide new insights into the mechanisms involved in the development of autoimmune disease in lpr mice. PMID- 12119352 TI - Potential role of phosphatidylinositol 3 kinase, rather than DNA-dependent protein kinase, in CpG DNA-induced immune activation. AB - Unmethylated CpG motifs present in bacterial DNA stimulate a strong innate immune response. There is evidence that DNA-dependent protein kinase (DNA-PK) mediates CpG signaling. Specifically, wortmannin (an inhibitor of phosphatidylinositol 3 kinase [PI3]-kinases including DNA-PK) interferes with CpG-dependent cell activation, and DNA-PK knockout (KO) mice fail to respond to CpG stimulation. Current studies establish that wortmannin actually inhibits the uptake and colocalization of CpG DNA with toll-like receptor (TLR)-9 in endocytic vesicles, thereby preventing CpG-induced activation of the NF-kappaB signaling cascade. We find that DNA-PK is not involved in this process, since three strains of DNA-PK KO mice responded normally to CpG DNA. These results support a model in which CpG signaling is mediated through TLR-9 but not DNA-PK, and suggest that wortmannin sensitive member(s) of the PI3-kinase family play a critical role in shuttling CpG DNA to TLR-9. PMID- 12119353 TI - Molecular adaptation of Borrelia burgdorferi in the murine host. AB - An analysis of expression of 137 lipoprotein genes on the course of murine infection revealed a two-step molecular adaptation by Borrelia burgdorferi, the Lyme disease spirochete. For the first step, regardless whether the initial inocula of B. burgdorferi expressed either all (cultured spirochetes) or less than 40 (host-adapted spirochetes) of the 137 lipoprotein genes, the spirochetes were modulated to transcribe 116 of the genes within 10 d after being introduced to the murine host. This step of adaptation was induced by the microenvironment of the host tissue. During the second step, which was forced by host immune selection pressure and occurred between 17 and 30 d after infection, B. burgdorferi down-regulated most of the lipoprotein genes and expressed less than 40 of the 137 genes. This novel adaptation mechanism could be a critical step for B. burgdorferi to proceed to chronic infection, as the pathogen would be cleared at the early stage of infection if the spirochetes failed to undergo this process. PMID- 12119354 TI - Integrins engage mitochondrial function for signal transduction by a mechanism dependent on Rho GTPases. AB - We show here the transient activation of the small GTPase Rac, followed by a rise in reactive oxygen species (ROS), as necessary early steps in a signal transduction cascade that lead to NFkappaB activation and collagenase-1 (CL 1)/matrix metalloproteinase-1 production after integrin-mediated cell shape changes. We show evidence indicating that this constitutes a new mechanism for ROS production mediated by small GTPases. Activated RhoA also induced ROS production and up-regulated CL-1 expression. A Rac mutant (L37) that prevents reorganization of the actin cytoskeleton prevented integrin-induced CL-1 expression, whereas mutations that abrogate Rac binding to the neutrophil NADPH membrane oxidase in vitro (H26 and N130) did not. Instead, ROS were produced by integrin-induced changes in mitochondrial function, which were inhibited by Bcl-2 and involved transient membrane potential loss. The cells showing this transient decrease in mitochondrial membrane potential were already committed to CL-1 expression. These results unveil a new molecular mechanism of signal transduction triggered by integrin engagement where a global mitochondrial metabolic response leads to gene expression rather than apoptosis. PMID- 12119356 TI - The metalloprotease Kuzbanian (ADAM10) mediates the transactivation of EGF receptor by G protein-coupled receptors. AB - Communication between different signaling pathways enables cells to coordinate the responses to diverse environmental signals. Activation of the transmembrane growth factor precursors plays a critical role in this communication and often involves metalloprotease-mediated proteolysis. Stimulation of G protein-coupled receptors (GPCR) transactivates the EGF receptors (EGFRs), which occurs via a metalloprotease-dependent cleavage of heparin-binding EGF (HB-EGF). However, the metalloprotease mediating the transactivation remains elusive. We show that the integral membrane metalloprotease Kuzbanian (KUZ; ADAM10), which controls Notch signaling in Drosophila, stimulates GPCR transactivation of EGFR. Upon stimulation of the bombesin receptors, KUZ increases the docking and activation of adaptors Src homology 2 domain-containing protein and Gab1 on the EGFR, and activation of Ras and Erk. In contrast, transfection of a protease domain-deleted KUZ, or blocking endogenous KUZ by morpholino antisense oligonucleotides, suppresses the transactivation. The effect of KUZ on shedding of HB-EGF and consequent transactivation of the EGFR depends on its metalloprotease activity. GPCR activation enhances the association of KUZ and its substrate HB-EGF with tetraspanin CD9. Thus, KUZ regulates the relay between the GPCR and EGFR signaling pathways. PMID- 12119355 TI - Localized Ca2+ uncaging reveals polarized distribution of Ca2+-sensitive Ca2+ release sites: mechanism of unidirectional Ca2+ waves. AB - Ca2+-induced Ca2+ release (CICR) plays an important role in the generation of cytosolic Ca2+ signals in many cell types. However, it is inherently difficult to distinguish experimentally between the contributions of messenger-induced Ca2+ release and CICR. We have directly tested the CICR sensitivity of different regions of intact pancreatic acinar cells using local uncaging of caged Ca2+. In the apical region, local uncaging of Ca2+ was able to trigger a CICR wave, which propagated toward the base. CICR could not be triggered in the basal region, despite the known presence of ryanodine receptors. The triggering of CICR from the apical region was inhibited by a pharmacological block of ryanodine or inositol trisphosphate receptors, indicating that global signals require coordinated Ca2+ release. Subthreshold agonist stimulation increased the probability of triggering CICR by apical uncaging, and uncaging-induced CICR could activate long-lasting Ca2+ oscillations. However, with subthreshold stimulation, CICR could still not be initiated in the basal region. CICR is the major process responsible for global Ca2+ transients, and intracellular variations in sensitivity to CICR predetermine the activation pattern of Ca2+ waves. PMID- 12119357 TI - A role for regulated binding of p150(Glued) to microtubule plus ends in organelle transport. AB - A subset of microtubule-associated proteins, including cytoplasmic linker protein (CLIP)-170, dynactin, EB1, adenomatous polyposis coli, cytoplasmic dynein, CLASPs, and LIS-1, has been shown recently to target to the plus ends of microtubules. The mechanisms and functions of this binding specificity are not understood, although a role in encouraging microtubule elongation has been proposed. To extend previous work on the role of dynactin in organelle transport, we analyzed p150(Glued) by live-cell imaging. Time-lapse analysis of p150(Glued) revealed targeting to the plus ends of growing microtubules, requiring the NH2 terminal cytoskeleton-associated protein-glycine rich domain, but not EB1 or CLIP 170. Effectors of protein kinase A modulated microtubule binding and suggested p150(Glued) phosphorylation as a factor in plus-end binding specificity. Using a phosphosensitive monoclonal antibody, we mapped the site of p150(Glued) phosphorylation to Ser-19. In vivo and in vitro analysis of phosphorylation site mutants revealed that p150(Glued) phosphorylation mediates dynamic binding to microtubules. To address the function of dynamic binding, we imaged GFP p150(Glued) during the dynein-dependent transport of Golgi membranes. Live-cell analysis revealed a transient interaction between Golgi membranes and GFP p150(Glued)-labeled microtubules just prior to transport, implicating microtubules and dynactin in a search-capture mechanism for minus-end-directed organelles. PMID- 12119358 TI - Cytoplasmic p21(Cip1/WAF1) regulates neurite remodeling by inhibiting Rho-kinase activity. AB - p21(Cip1/WAF1) has cell cycle inhibitory activity by binding to and inhibiting both cyclin/Cdk kinases and proliferating cell nuclear antigen. Here we show that p21(Cip1/WAF1) is induced in the cytoplasm during the course of differentiation of chick retinal precursor cells and N1E-115 cells. Ectopic expression of p21(Cip1/WAF1) lacking the nuclear localization signal in N1E-115 cells and NIH3T3 cells affects the formation of actin structures, characteristic of inactivation of Rho. p21(Cip1/WAF1) forms a complex with Rho-kinase and inhibits its activity in vitro and in vivo. Neurite outgrowth and branching from the hippocampal neurons are promoted if p21(Cip1/WAF1) is expressed abundantly in the cytoplasm. These results suggest that cytoplasmic p21(Cip1/WAF1) may contribute to the developmental process of the newborn neurons that extend axons and dendrites into target regions. PMID- 12119359 TI - A phosphatidylinositol (4,5)-bisphosphate binding site within mu2-adaptin regulates clathrin-mediated endocytosis. AB - The clathrin adaptor complex AP-2 serves to coordinate clathrin-coated pit assembly with the sorting of transmembrane cargo proteins at the plasmalemma. How precisely AP-2 assembly and cargo protein recognition at sites of endocytosis are regulated has remained unclear, but recent evidence implicates phosphoinositides, in particular phosphatidylinositol (4,5)-bisphosphate (PI[4,5]P2), in these processes. Here we have identified and functionally characterized a conserved binding site for PI(4,5)P2 within mu2-adaptin, the medium chain of the clathrin adaptor complex AP-2. Mutant mu2 lacking a cluster of conserved lysine residues fails to bind PI(4,5)P2 and to compete the recruitment of native clathrin/AP-2 to PI(4,5)P2-containing liposomes or to presynaptic membranes. Moreover, we show that expression of mutant mu2 inhibits receptor-mediated endocytosis in living cells. We suggest that PI(4,5)P2 binding to mu2-adaptin regulates clathrin mediated endocytosis and thereby may contribute to structurally linking cargo recognition to coat formation. PMID- 12119360 TI - Calcium-independent stimulation of membrane fusion and SNAREpin formation by synaptotagmin I. AB - Neurotransmitter release requires the direct coupling of the calcium sensor with the machinery for membrane fusion. SNARE proteins comprise the minimal fusion machinery, and synaptotagmin I, a synaptic vesicle protein, is the primary candidate for the main neuronal calcium sensor. To test the effect of synaptotagmin I on membrane fusion, we incorporated it into a SNARE-mediated liposome fusion assay. Synaptotagmin I dramatically stimulated membrane fusion by facilitating SNAREpin zippering. This stimulatory effect was topologically restricted to v-SNARE vesicles (containing VAMP 2) and only occurred in trans to t-SNARE vesicles (containing syntaxin 1A and SNAP-25). Interestingly, calcium did not affect the overall fusion reaction. These results indicate that synaptotagmin I can directly accelerate SNARE-mediated membrane fusion and raise the possibility that additional components might be required to ensure tight calcium coupling. PMID- 12119361 TI - Autotaxin has lysophospholipase D activity leading to tumor cell growth and motility by lysophosphatidic acid production. AB - Autotaxin (ATX) is a tumor cell motility-stimulating factor, originally isolated from melanoma cell supernatants. ATX had been proposed to mediate its effects through 5'-nucleotide pyrophosphatase and phosphodiesterase activities. However, the ATX substrate mediating the increase in cellular motility remains to be identified. Here, we demonstrated that lysophospholipase D (lysoPLD) purified from fetal bovine serum, which catalyzes the production of the bioactive phospholipid mediator, lysophosphatidic acid (LPA), from lysophosphatidylcholine (LPC), is identical to ATX. The Km value of ATX for LPC was 25-fold lower than that for the synthetic nucleoside substrate, p-nitrophenyl-tri-monophosphate. LPA mediates multiple biological functions including cytoskeletal reorganization, chemotaxis, and cell growth through activation of specific G protein-coupled receptors. Recombinant ATX, particularly in the presence of LPC, dramatically increased chemotaxis and proliferation of multiple different cell lines. Moreover, we demonstrate that several cancer cell lines release significant amounts of LPC, a substrate for ATX, into the culture medium. The demonstration that ATX and lysoPLD are identical suggests that autocrine or paracrine production of LPA contributes to tumor cell motility, survival, and proliferation. It also provides potential novel targets for therapy of pathophysiological states including cancer. PMID- 12119362 TI - Mossy fiber Zn2+ spillover modulates heterosynaptic N-methyl-D-aspartate receptor activity in hippocampal CA3 circuits. AB - Although Zn2+ is contained in large amounts in the synaptic terminals of hippocampal mossy fibers (MFs), its physiological role in synaptic transmission is poorly understood. By using the newly developed high-sensitivity Zn2+ indicator ZnAF-2, the spatiotemporal dynamics of Zn2+ was monitored in rat hippocampal slices. When high-frequency stimulation was delivered to the MFs, the concentration of extracellular Zn2+ was immediately elevated in the stratum lucidum, followed by a mild increase in the stratum radiatum adjacent to the stratum lucidum, but not in the distal area of stratum radiatum. The Zn2+ increase was insensitive to a non-N-methyl-d-aspartate (NMDA) receptor antagonist but was efficiently attenuated by tetrodotoxin or Ca2+-free medium, suggesting that Zn2+ is released by MF synaptic terminals in an activity-dependent manner, and thereafter diffuses extracellularly into the neighboring stratum radiatum. Electrophysiological analyses revealed that NMDA receptor-mediated synaptic responses in CA3 proximal stratum radiatum were inhibited in the immediate aftermath of MF activation and that this inhibition was no longer observed in the presence of a Zn2+-chelating agent. Thus, Zn2+ serves as a spatiotemporal mediator in imprinting the history of MF activity in contiguous hippocampal networks. We predict herein a novel form of metaplasticity, i.e., an experience dependent non-Hebbian modulation of synaptic plasticity. PMID- 12119363 TI - Sequential assistance of molecular chaperones and transient formation of covalent complexes during protein degradation from the ER. AB - BACE457 is a recently identified pancreatic isoform of human beta-secretase. We report that this membrane glycoprotein and its soluble variant are characterized by inefficient folding in the ER, leading to proteasome-mediated ER-associated degradation (ERAD). Dissection of the degradation process revealed that upon release from calnexin, extensively oxidized BACE457 transiently entered in disulfide-bonded complexes associated with the lumenal chaperones BiP and protein disulfide isomerase (PDI) before unfolding and dislocation into the cytosol for degradation. BACE457 and its lumenal variant accumulated in disulfide-bonded complexes, in the ER lumen, also when protein degradation was inhibited. The complexes were disassembled and the misfolded polypeptides were cleared from the ER upon reactivation of the degradation machinery. Our data offer new insights into the mechanism of ERAD by showing a sequential involvement of the calnexin and BiP/PDI chaperone systems. We report the unexpected transient formation of covalent complexes in the ER lumen during the ERAD process, and we show that PDI participates as an oxidoreductase and a redox-driven chaperone in the preparation of proteins for degradation from the mammalian ER. PMID- 12119365 TI - Plant metabolomics: the missing link in functional genomics strategies. PMID- 12119366 TI - Deductions about the number, organization, and evolution of genes in the tomato genome based on analysis of a large expressed sequence tag collection and selective genomic sequencing. AB - Analysis of a collection of 120,892 single-pass ESTs, derived from 26 different tomato cDNA libraries and reduced to a set of 27,274 unique consensus sequences (unigenes), revealed that 70% of the unigenes have identifiable homologs in the Arabidopsis genome. Genes corresponding to metabolism have remained most conserved between these two genomes, whereas genes encoding transcription factors are among the fastest evolving. The majority of the 10 largest conserved multigene families share similar copy numbers in tomato and Arabidopsis, suggesting that the multiplicity of these families may have occurred before the divergence of these two species. An exception to this multigene conservation was observed for the E8-like protein family, which is associated with fruit ripening and has higher copy number in tomato than in Arabidopsis. Finally, six BAC clones from different parts of the tomato genome were isolated, genetically mapped, sequenced, and annotated. The combined analysis of the EST database and these six sequenced BACs leads to the prediction that the tomato genome encodes approximately 35,000 genes, which are sequestered largely in euchromatic regions corresponding to less than one-quarter of the total DNA in the tomato nucleus. PMID- 12119367 TI - Identification, analysis, and utilization of conserved ortholog set markers for comparative genomics in higher plants. AB - We have screened a large tomato EST database against the Arabidopsis genomic sequence and report here the identification of a set of 1025 genes (referred to as a conserved ortholog set, or COS markers) that are single or low copy in both genomes (as determined by computational screens and DNA gel blot hybridization) and that have remained relatively stable in sequence since the early radiation of dicotyledonous plants. These genes were annotated, and a large portion could be assigned to putative functional categories associated with basic metabolic processes, such as energy-generating processes and the biosynthesis and degradation of cellular building blocks. We further demonstrate, through computational screens (e.g., against a Medicago truncatula database) and direct hybridization on genomic DNA of diverse plant species, that these COS markers also are conserved in the genomes of other plant families. Finally, we show that this gene set can be used for comparative mapping studies between highly divergent genomes such as those of tomato and Arabidopsis. This set of COS markers, identified computationally and experimentally, may further studies on comparative genomes and phylogenetics and elucidate the nature of genes conserved throughout plant evolution. PMID- 12119368 TI - Arabidopsis SON1 is an F-box protein that regulates a novel induced defense response independent of both salicylic acid and systemic acquired resistance. AB - One of several induced defense responses in plants is systemic acquired resistance (SAR), which is regulated by salicylic acid and in Arabidopsis by the NIM1/NPR1 protein. To identify additional components of the SAR pathway or other genes that regulate SAR-independent resistance, we performed genetic suppressor screens of mutagenized nim1-1 seedlings, which are highly susceptible to infection by Peronospora parasitica. We isolated the son1 (suppressor of nim1-1) mutant, which shows full restoration of pathogen resistance without the induction of SAR-associated genes and expresses resistance when combined with a salicylate hydroxylase (nahG) transgene. These features indicate that son1-mediated resistance is distinct from SAR. Resistance is effective against both the virulent oomycete Peronospora and the bacterial pathogen Pseudomonas syringae pv tomato strain DC3000. We cloned SON1 and found it to encode a novel protein containing an F-box motif, an element found within the specificity determinant in the E3 ubiquitin-ligase complex. We propose the existence of a novel defense response that is independent of SAR and negatively regulated in Arabidopsis by SON1 through the ubiquitin-proteosome pathway. PMID- 12119369 TI - Role of SCF ubiquitin-ligase and the COP9 signalosome in the N gene-mediated resistance response to Tobacco mosaic virus. AB - The tobacco N gene confers resistance to Tobacco mosaic virus (TMV) and encodes a toll-interleukin-1 receptor/nucleotide binding/Leu-rich repeat class protein. Recent evidence indicates that the Nicotiana benthamiana Rar1 gene (NbRar1), which encodes a protein with a zinc finger motif called CHORD (Cys- and His-rich domain), is required for the function of N. To investigate the role of NbRar1 in plant defense, we identified its interaction partners. We show that the NbRar1 protein interacts with NbSGT1, a highly conserved component of the SCF (Skp1/Cullin/F-box protein)-type E3 ubiquitin ligase complex involved in protein degradation. In addition, we show that NbSGT1 interacts with NbSKP1. Suppression of NbSGT1 and NbSKP1 shows that these genes play an important role in the N mediated resistance response to TMV. Both NbRar1 and NbSGT1 associate with the COP9 signalosome, another multiprotein complex involved in protein degradation via the ubiquitin-proteasome pathway. Silencing of the NbCOP9 signalosome also compromises N-mediated resistance to TMV. Our results reveal new roles for SCF and the COP9 signalosome in plant defense signaling. PMID- 12119370 TI - A surveillance system regulates selective entry of RNA into the shoot apex. AB - Phloem-mobile endogenous RNA is trafficked selectively into the shoot apex. In contrast, most viruses and long-distance post-transcriptional gene silencing (PTGS) signals are excluded from the shoot apex. These observations suggest the operation of an underlying regulatory mechanism. To examine this possibility, a potexvirus movement protein, known to modify cell-to-cell trafficking and PTGS, was expressed ectopically in transgenic plants. These plants were found to be compromised in their capacity to exclude both viral RNA and silencing signals from the shoot apex. The transgenic plants also displayed various degrees of abnormal leaf polarity depending on transgene expression level. Normal patterns of organ development were restored by either virus- or Agrobacterium tumefaciens mediated induction of PTGS. This revealed the presence of an RNA signal surveillance system that acts to allow the selective entry of RNA into the shoot apex. We propose that this surveillance system regulates signaling and protects the shoot apex, in particular the cells that give rise to reproductive structures, from viral invasion. PMID- 12119371 TI - Elicitor-activated phospholipase A(2) generates lysophosphatidylcholines that mobilize the vacuolar H(+) pool for pH signaling via the activation of Na(+) dependent proton fluxes. AB - The elicitation of phytoalexin biosynthesis in cultured cells of California poppy involves a shift of cytoplasmic pH via the transient efflux of vacuolar protons. Intracellular effectors of vacuolar proton transport were identified by a novel in situ approach based on the selective permeabilization of the plasma membrane for molecules of < or = 10 kD. Subsequent fluorescence imaging of the vacuolar pH correctly reported experimental changes of activity of the tonoplast proton transporters. Lysophosphatidylcholine (LPC) caused a transient increase of the vacuolar pH by increasing the Na(+) sensitivity of a Na(+)-dependent proton efflux that was inhibited by amiloride. In intact cells, yeast elicitor activated phospholipase A(2), as demonstrated by the formation of LPC from fluorescent substrate analogs, and caused a transient increase of endogenous LPC, as determined by matrix-assisted laser desorption and ionization time-of-flight mass spectrometry. It is suggested that LPC generated by phospholipase A(2) at the plasma membrane transduces the elicitor-triggered signal into the activation of a tonoplast H(+)/Na(+) antiporter. PMID- 12119372 TI - The subtilisin-like serine protease SDD1 mediates cell-to-cell signaling during Arabidopsis stomatal development. AB - Wild-type stomata are distributed nonrandomly, and their density is controlled by endogenous and exogenous factors. In the Arabidopsis mutant stomatal density and distribution1-1 (sdd1-1), the establishment of the stomatal pattern is disrupted, resulting in stomata clustering and twofold to fourfold increases in stomatal density. The SDD1 gene that encodes a subtilisin-like Ser protease is expressed strongly in stomatal precursor cells (meristemoids and guard mother cells), and the SDD1 promoter is controlled negatively by a feedback mechanism. The encoded protein is exported to the apoplast and probably is associated with the plasma membrane. SDD1 overexpression in the wild type leads to a phenotype opposite to that caused by the sdd1-1 mutation, with a twofold to threefold decrease in stomatal density and the formation of arrested stomata. While SDD1 overexpression was effective in the flp mutant, the tmm mutation acted epistatically. Thus, we propose that SDD1 generates an extracellular signal by meristemoids/guard mother cells and demonstrate that the function of SDD1 is dependent on TMM activity. PMID- 12119373 TI - Nucleocytoplasmic partitioning of the plant photoreceptors phytochrome A, B, C, D, and E is regulated differentially by light and exhibits a diurnal rhythm. AB - The phytochrome family of plant photoreceptors has a central role in the adaptation of plant development to changes in ambient light conditions. The individual phytochrome species regulate different or partly overlapping physiological responses. We generated transgenic Arabidopsis plants expressing phytochrome A to E:green fluorescent protein (GFP) fusion proteins to assess the biological role of intracellular compartmentation of these photoreceptors in light-regulated signaling. We show that all phytochrome:GFP fusion proteins were imported into the nuclei. Translocation of these photoreceptors into the nuclei was regulated differentially by light. Light-induced accumulation of phytochrome species in the nuclei resulted in the formation of speckles. The appearance of these nuclear structures exhibited distinctly different kinetics, wavelengths, and fluence dependence and was regulated by a diurnal rhythm. Furthermore, we demonstrate that the import of mutant phytochrome B:GFP and phytochrome A:GFP fusion proteins, shown to be defective in signaling in vivo, is regulated by light but is not accompanied by the formation of speckles. These results suggest that (1) the differential regulation of the translocation of phytochrome A to E into nuclei plays a role in the specification of functions, and (2) the appearance of speckles is a functional feature of phytochrome-regulated signaling. PMID- 12119374 TI - The Arabidopsis mutant cev1 links cell wall signaling to jasmonate and ethylene responses. AB - Biotic and abiotic stresses stimulate the synthesis of jasmonates and ethylene, which, in turn, induce the expression of genes involved in stress response and enhance defense responses. The cev1 mutant has constitutive expression of stress response genes and has enhanced resistance to fungal pathogens. Here, we show that cev1 plants have increased production of jasmonate and ethylene and that its phenotype is suppressed by mutations that interrupt jasmonate and ethylene signaling. Genetic mapping, complementation analysis, and sequence analysis revealed that CEV1 is the cellulose synthase CeSA3. CEV1 was expressed predominantly in root tissues, and cev1 roots contained less cellulose than wild type roots. Significantly, the cev1 mutant phenotype could be reproduced by treating wild-type plants with cellulose biosynthesis inhibitors, and the cellulose synthase mutant rsw1 also had constitutive expression of VSP. We propose that the cell wall can signal stress responses in plants. PMID- 12119375 TI - Protein-protein interactions between sucrose transporters of different affinities colocalized in the same enucleate sieve element. AB - Suc represents the major transport form for carbohydrates in plants. Suc is loaded actively against a concentration gradient into sieve elements, which constitute the conduit for assimilate export out of leaves. Three members of the Suc transporter family with different properties were identified: SUT1, a high affinity Suc proton cotransporter; SUT4, a low-affinity transporter; and SUT2, which in yeast is only weakly active and shows features similar to those of the yeast sugar sensors RGT2 and SNF3. Immunolocalization demonstrated that all three SUT proteins are localized in the same enucleate sieve element. Thus, the potential of Suc transporters to form homooligomers was tested by the yeast-based split-ubiquitin system. The results show that both SUT1 and SUT2 have the potential to form homooligomers. Moreover, all three Suc transporters have the potential to interact with each other. As controls, a potassium channel and a monosaccharide transporter, expressed in the plasma membrane, did not interact with the SUTs. The in vivo interaction between the functionally different Suc transporters indicates that the membrane proteins are capable of forming oligomeric structures that, like mammalian Glc transporter complexes, might be of functional significance for the regulation of transport. PMID- 12119376 TI - Detection and localization of a chloroplast-encoded HU-like protein that organizes chloroplast nucleoids. AB - Chloroplast DNA (cpDNA) is packed into discrete structures called chloroplast nucleoids (cp-nucleoids). The structure of cpDNA is thought to be important for its maintenance and regulation. In bacteria and mitochondria, histone-like proteins (such as HU and Abf2, respectively) are abundant and play important roles in DNA organization. However, a primary structural protein has yet to be found in cp-nucleoids. Here, we identified an abundant DNA binding protein from isolated cp-nucleoids of the primitive red alga Cyanidioschyzon merolae. The purified protein had sequence homology with the bacterial histone-like protein HU, and it complemented HU-lacking Escherichia coli mutants. The protein, called HC (histone-like protein of chloroplast), was encoded by a single gene (CmhupA) in the C. merolae chloroplast genome. Using immunofluorescence and immunoelectron microscopy, we demonstrated that HC was distributed uniformly throughout the entire cp-nucleoid. The protein was expressed constitutively throughout the cell and the chloroplast division cycle, and it was able to condense DNA. These results indicate that HC, a bacteria-derived histone-like protein, primarily organizes cpDNA into the nucleoid. PMID- 12119377 TI - Missense mutation in the PAS2 domain of phytochrome A impairs subnuclear localization and a subset of responses. AB - Phytochrome A signaling shows two photobiologically discrete outputs: so-called very-low-fluence responses (VLFR) and high-irradiance responses (HIR). By modifying previous screening protocols, we isolated two Arabidopsis mutants retaining VLFR and lacking HIR. Phytochrome A negatively or positively regulates phytochrome B signaling, depending on light conditions. These mutants retained the negative but lacked the positive regulation. Both mutants carry the novel phyA-302 allele, in which Glu-777 (a residue conserved in angiosperm phytochromes) changed to Lys in the PAS2 motif of the C-terminal domain. The phyA 302 mutants showed a 50% reduction in phytochrome A levels in darkness, but this difference was compensated for by greater stability under continuous far-red light. phyA-302:green fluorescent protein fusion proteins showed normal translocation from the cytosol to the nucleus under continuous far-red light but failed to produce nuclear spots, suggesting that nuclear speckles could be involved in HIR signaling and phytochrome A degradation. We propose that the PAS2 domain of phytochrome A is necessary to initiate signaling in HIR but not in VLFR, likely via interaction with a specific partner. PMID- 12119379 TI - Maize HSP101 plays important roles in both induced and basal thermotolerance and primary root growth. AB - HSP101 belongs to the ClpB protein subfamily whose members promote the renaturation of protein aggregates and are essential for the induction of thermotolerance. We found that maize HSP101 accumulated in mature kernels in the absence of heat stress. At optimal temperatures, HSP101 disappeared within the first 3 days after imbibition, although its levels increased in response to heat shock. In embryonic cells, HSP101 concentrated in the nucleus and in some nucleoli. Hsp101 maps near the umc132 and npi280 markers on chromosome 6. Five maize hsp101-m-::Mu1 alleles were isolated. Mutants were null for HSP101 and defective in both induced and basal thermotolerance. Moreover, during the first 3 days after imbibition, primary roots grew faster in the mutants at optimal temperature. Thus, HSP101 is a nucleus-localized protein that, in addition to its role in thermotolerance, negatively influences the growth rate of the primary root. HSP101 is dispensable for proper embryo and whole plant development in the absence of heat stress. PMID- 12119378 TI - Endogenous and silencing-associated small RNAs in plants. AB - A large set of endogenous small RNAs of predominantly 21 to 24 nucleotides was identified in Arabidopsis. These small RNAs resembled micro-RNAs from animals and were similar in size to small interfering RNAs that accumulated during RNA silencing triggered by multiple types of inducers. Among the 125 sequences identified, the vast majority (90%) arose from intergenic regions, although small RNAs corresponding to predicted protein-coding genes, transposon-like sequences, and a structural RNA gene also were identified. Evidence consistent with the derivation of small RNAs of both polarities, and from highly base-paired precursors, was obtained through the identification and analysis of clusters of small RNA loci. The accumulation of specific small RNAs was regulated developmentally. We propose that Arabidopsis small RNAs participate in a wide range of post-transcriptional and epigenetic events. PMID- 12119380 TI - The Arabidopsis SKU5 gene encodes an extracellular glycosyl phosphatidylinositol anchored glycoprotein involved in directional root growth. AB - To investigate how roots respond to directional cues, we characterized a T-DNA tagged Arabidopsis mutant named sku5 in which the roots skewed and looped away from the normal downward direction of growth on inclined agar surfaces. sku5 roots and etiolated hypocotyls were slightly shorter than normal and exhibited a counterclockwise (left-handed) axial rotation bias. The surface-dependent skewing phenotype disappeared when the roots penetrated the agar surface, but the axial rotation defect persisted, revealing that these two directional growth processes are separable. The SKU5 gene belongs to a 19-member gene family designated SKS (SKU5 Similar) that is related structurally to the multiple-copper oxidases ascorbate oxidase and laccase. However, the SKS proteins lack several of the conserved copper binding motifs characteristic of copper oxidases, and no enzymatic function could be assigned to the SKU5 protein. Analysis of plants expressing SKU5 reporter constructs and protein gel blot analysis showed that SKU5 was expressed most strongly in expanding tissues. SKU5 was glycosylated and modified by glycosyl phosphatidylinositol and localized to both the plasma membrane and the cell wall. Our observations suggest that SKU5 affects two directional growth processes, possibly by participating in cell wall expansion. PMID- 12119381 TI - Hypersensitivity of abscisic acid-induced cytosolic calcium increases in the Arabidopsis farnesyltransferase mutant era1-2. AB - Cytosolic calcium increases were analyzed in guard cells of the Arabidopsis farnesyltransferase deletion mutant era1-2 (enhanced response to abscisic acid). At low abscisic acid (ABA) concentrations (0.1 microM), increases of guard cell cytosolic calcium and stomatal closure were activated to a greater extent in the era1-2 mutant compared with the wild type. Patch clamping of era1-2 guard cells showed enhanced ABA sensitivity of plasma membrane calcium channel currents. These data indicate that the ERA1 farnesyltransferase targets a negative regulator of ABA signaling that acts between the points of ABA perception and the activation of plasma membrane calcium influx channels. Experimental increases of cytosolic calcium showed that the activation of S-type anion currents downstream of cytosolic calcium and extracellular calcium-induced stomatal closure were unaffected in era1-2, further supporting the positioning of era1-2 upstream of cytosolic calcium in the guard cell ABA signaling cascade. Moreover, the suppression of ABA-induced calcium increases in guard cells by the dominant protein phosphatase 2C mutant abi2-1 was rescued partially in era1-2 abi2-1 double mutant guard cells, further reinforcing the notion that ERA1 functions upstream of cytosolic calcium and indicating the genetic interaction of these two mutations upstream of ABA-induced calcium increases. PMID- 12119382 TI - Punctuated evolution of mitochondrial gene content: high and variable rates of mitochondrial gene loss and transfer to the nucleus during angiosperm evolution. AB - To study the tempo and pattern of mitochondrial gene loss in plants, DNAs from 280 genera of flowering plants were surveyed for the presence or absence of 40 mitochondrial protein genes by Southern blot hybridization. All 14 ribosomal protein genes and both sdh genes have been lost from the mitochondrial genome many times (6 to 42) during angiosperm evolution, whereas only two losses were detected among the other 24 genes. The gene losses have a very patchy phylogenetic distribution, with periods of stasis followed by bursts of loss in certain lineages. Most of the oldest groups of angiosperms are still mired in a prolonged stasis in mitochondrial gene content, containing nearly the same set of genes as their algal ancestors more than a billion years ago. In sharp contrast, other plants have rapidly lost many or all of their 16 mitochondrial ribosomal protein and sdh genes, thereby converging on a reduced gene content more like that of an animal or fungus than a typical plant. In these and many lineages with more modest numbers of losses, the rate of ribosomal protein and sdh gene loss exceeds, sometimes greatly, the rate of mitochondrial synonymous substitutions. Most of these mitochondrial gene losses are probably the consequence of gene transfer to the nucleus; thus, rates of functional gene transfer also may vary dramatically in angiosperms. PMID- 12119383 TI - Neural correlates of instrumental learning in primary auditory cortex. AB - In instrumental learning, Thorndike's law of effect states that stimulus-response relations are strengthened if they occur prior to positive reinforcement and weakened if they occur prior to negative reinforcement. In this study, we demonstrate that neural correlates of Thorndike's law may be observed in the primary auditory cortex, A1. Adult owl monkeys learned to discriminate tones higher than a standard frequency. Responses recorded from implanted microelectrodes initially exhibited broad spectral selectivity over a four-to five octave range. With training, frequency discrimination thresholds changed from close to one octave to about 1/12 octave. Physiological recordings during the week in which the monkey came under behavioral control signaled by a drop in measured threshold had stronger responses to all frequencies. During the same week, A1 neural responses to target stimuli increased relative to standard and nontarget stimuli. This emergent difference in responsiveness persisted throughout the subsequent weeks of behavioral training. These data suggest that behavioral responses to stimuli modulate responsiveness in primary cortical areas. PMID- 12119384 TI - Cyclic electron transfer in plant leaf. AB - The turnover of linear and cyclic electron flows has been determined in fragments of dark-adapted spinach leaf by measuring the kinetics of fluorescence yield and of the transmembrane electrical potential changes under saturating illumination. When Photosystem (PS) II is inhibited, a cyclic electron flow around PSI operates transiently at a rate close to the maximum turnover of photosynthesis. When PSII is active, the cyclic flow operates with a similar rate during the first seconds of illumination. The high efficiency of the cyclic pathway implies that the cyclic and the linear transfer chains are structurally isolated one from the other. We propose that the cyclic pathway operates within a supercomplex including one PSI, one cytochrome bf complex, one plastocyanin, and one ferredoxin. The cyclic process induces the synthesis of ATP needed for the activation of the Benson-Calvin cycle. A fraction of PSI ( approximately 50%), not included in the supercomplexes, participates in the linear pathway. The illumination would induce a dissociation of the supercomplexes that progressively increases the fraction of PSI involved in the linear pathway. PMID- 12119385 TI - Chaotic mixing deep in the lung. AB - Our current understanding of the transport and deposition of aerosols (viruses, bacteria, air pollutants, aerosolized drugs) deep in the lung has been grounded in dispersive theories based on untested assumptions about the nature of acinar airflow fields. Traditionally, these have been taken to be simple and kinematically reversible. In this article, we apply the recently discovered fluid mechanical phenomenon of irreversible low-Reynolds number flow to the lung. We demonstrate, through flow visualization studies in rhythmically ventilated rat lungs, that such a foundation is false, and that chaotic mixing may be key to aerosol transport. We found substantial alveolar flow irreversibility with stretched and folded fractal patterns, which lead to a sudden increase in mixing. These findings support our theory that chaotic alveolar flow--characterized by stagnation saddle points associated with alveolar vortices--governs gas kinematics in the lung periphery, and hence the transport, mixing, and ultimately the deposition of fine aerosols. This mechanism calls for a rethinking of the relationship of exposure and deposition of fine inhaled particles. PMID- 12119386 TI - Protein-protein interactions among C-4 demethylation enzymes involved in yeast sterol biosynthesis. AB - A Saccharomyces cerevisae microarray expression study indicated that an ORF, YER044C, now designated ERG28, was strongly coregulated with ergosterol biosynthesis. Disruption of the ERG28 gene results in slow growth and accumulation of sterol intermediates similar to those observed in erg26 and erg27 null strains, suggesting that the Erg28p may interact with Erg26p and/or Erg27p. In this study, a peptide from human hemagglutinin protein (HA) epitope tag was added to ERG26 and ERG27 genes, and a Myc tag was added to the ERG28 gene to detect interactions between Erg28p and Erg26p/Erg27p. Differential centrifugation showed that Erg26p, Erg27p, and Erg28p are all membrane-associated proteins. Green fluorescent protein-fusion protein localization studies showed that Erg26p, Erg27p, and Erg28p are all located in the endoplasmic reticulum. Solubilized membrane protein coimmunoprecipitation studies using rabbit anti-Erg25p indicated that Erg25p coimmunoprecipitates with both Erg27p and Erg28p. Erg28p was also shown to reciprocally coimmunoprecipitate with Erg27p. However, no coimmunoprecipitation was observed with Erg26p, most likely because of the poor solubilization of this protein. Sucrose gradient ultracentrifugation studies suggested that Erg25p/Erg26p/Erg27p/Erg28p, along with other proteins in sterol biosynthesis, might form a complex between 66 and 200 kDa. Using an anti-HA column with Erg27p-HA and Erg26p-HA as target proteins, a complex containing Erg25p/Erg26p/Erg27p/Erg28p was identified. Thus, we suggest that Erg28p works as a transmembrane scaffold to tether Erg27p and possibly other C-4 demethylation proteins (Erg25p, Erg26p), forming a demethylation complex in the endoplasmic reticulum. PMID- 12119388 TI - Cooperation of two-domain Ca(2+) channel fragments in triad targeting and restoration of excitation- contraction coupling in skeletal muscle. AB - The specific incorporation of the skeletal muscle voltage-dependent Ca(2+) channel in the triad is a prerequisite of normal excitation-contraction (EC) coupling. Sequences involved in membrane expression and in targeting of Ca(2+) channels into skeletal muscle triads have been described in different regions of the alpha(1S) subunit. Here we studied the targeting properties of two-domain alpha(1S) fragments, green fluorescent protein (GFP)-I x II (1-670) and III x IV (691-1873) expressed alone or in combination in dysgenic (alpha(1S)-null) myotubes. Immunofluorescence analysis showed that GFP-I x II or III x IV expressed separately were not targeted into triads. In contrast, on coexpression the two alpha(1S) fragments were colocalized with one another and with the ryanodine receptor in the triads. Coexpression of GFP-I x II and III x IV also fully restored Ca(2+) currents and depolarization-induced Ca(2+) transients, despite the severed connection between the two channel halves and the absence of amino acids 671-690 from either alpha(1S) fragment. Thus, triad targeting, like the rescue of function, requires the cooperation and coassembly of the two complementary channel fragments. Transferring the C terminus of alpha(1S) to the N-terminal two-domain fragment (GFP-I x II x tail), or transferring the I-II connecting loop containing the beta interaction domain to the C-terminal fragment (III x IV x beta in) did not improve the targeting properties of the individually expressed two-domain channel fragments. Thus, the cooperation of GFP-I.II and III.IV in targeting cannot be explained solely by a sequential action of the beta subunit by means of the I-II loop in releasing the channel from the sarcoplasmic reticulum and of the C terminus in triad targeting. PMID- 12119387 TI - Global analysis of mRNA decay and abundance in Escherichia coli at single-gene resolution using two-color fluorescent DNA microarrays. AB - Much of the information available about factors that affect mRNA decay in Escherichia coli, and by inference in other bacteria, has been gleaned from study of less than 25 of the approximately 4,300 predicted E. coli messages. To investigate these factors more broadly, we examined the half-lives and steady state abundance of known and predicted E. coli mRNAs at single-gene resolution by using two-color fluorescent DNA microarrays. An rRNA-based strategy for normalization of microarray data was developed to permit quantitation of mRNA decay after transcriptional arrest by rifampicin. We found that globally, mRNA half-lives were similar in nutrient-rich media and defined media in which the generation time was approximately tripled. A wide range of stabilities was observed for individual mRNAs of E. coli, although approximately 80% of all mRNAs had half-lives between 3 and 8 min. Genes having biologically related metabolic functions were commonly observed to have similar stabilities. Whereas the half lives of a limited number of mRNAs correlated positively with their abundance, we found that overall, increased mRNA stability is not predictive of increased abundance. Neither the density of putative sites of cleavage by RNase E, which is believed to initiate mRNA decay in E. coli, nor the free energy of folding of 5' or 3' untranslated region sequences was predictive of mRNA half-life. Our results identify previously unsuspected features of mRNA decay at a global level and also indicate that generalizations about decay derived from the study of individual gene transcripts may have limited applicability. PMID- 12119389 TI - The structures of the active center in dark-adapted bacteriorhodopsin by solution state NMR spectroscopy. AB - The two forms of bacteriorhodopsin present in the dark-adapted state, containing either all-trans or 13-cis,15-syn retinal, were examined by using solution state NMR, and their structures were determined. Comparison of the all-trans and the 13 cis,15-syn forms shows a shift in position of about 0.25 A within the pocket of the protein. Comparing this to the 13-cis,15-anti chromophore of the catalytic cycle M-intermediate structure, the 13-cis,15-syn form demonstrates a less pronounced up-tilt of the retinal C12[bond]C14 region, while leaving W182 and T178 essentially unchanged. The N[bond]H dipole of the Schiff base orients toward the extracellular side in both forms, however, it reorients toward the intracellular side in the 13-cis,15-anti configuration to form the catalytic M intermediate. Thus, the change of the N[bond]H dipole is considered primarily responsible for energy storage, conformation changes of the protein, and the deprotonation of the Schiff base. The structural similarity of the all-trans and 13-cis,15-syn forms is taken as strong evidence for the ion dipole dragging model by which proton (hydroxide ion) translocation follows the change of the dipole. PMID- 12119390 TI - Increased cell proliferation, but not reduced cell death, induces lymphocytosis in bovine leukemia virus-infected sheep. AB - Lymphocyte homeostasis is the result of a critical balance between cell proliferation and death. Disruption of this subtle equilibrium can lead to the onset of leukemia, an increase in the number of lymphocytes being potentially due to both of these parameters. The relative importance of cell proliferation vs. apoptosis during pathogenesis induced by the primate T cell lymphotropic viruses and bovine leukemia virus (BLV) has been difficult to assess because of conflicting data from a range of in vitro and ex vivo experimental systems. Here, we aim to resolve this issue by measuring the rates of cell proliferation and death in the BLV-ovine system, an animal model of human T lymphotropic virus (HTLV-1). We use a method based on the i.v. injection of 5-bromodeoxyuridine into BLV-infected sheep. We show that B lymphocytes in BLV(+) asymptomatic sheep proliferate significantly faster than in uninfected controls (average proliferation rate: 0.020 per day vs. 0.011 per day). In contrast, the rates of cell death were not significantly different between aleukemic BLV-infected and control sheep (average death rate 0.089 per day vs. 0.094 per day, respectively). We conclude that the increase in the number of B cells during BLV-induced lymphocytosis results from higher proliferation rates but is not due to a significant decrease in apoptosis, in contrast to data from in vitro (ex vivo) experiments. The imbalance created by the net increase in proliferation in the absence of compensating cell death reveals a complex mechanism of feedback regulation controlling homeostasis in the blood compartment. PMID- 12119391 TI - Ultrasound induced improvement in optical coherence tomography (OCT) resolution. AB - Optical coherence tomography (OCT) is a rapidly emerging technology for high resolution biomedical imaging. The axial resolution of this technology is determined by the bandwidth of the source. Commercial sources generally provide resolutions of 10-20 microm whereas laboratory-based solid state lasers have resolutions of approximately 4 microm. The resolution in tissue depends almost exclusively on detecting single scattered events. However, the phenomenon known as multiple scattering results in a deterioration of resolution as a function of depth. In this study, OCT was combined with ultrasound in an attempt to reduce the effect of multiple scattering. The theory is that, with parallel ultrasound and OCT beams, multiply scattered light with a momentum component significantly perpendicular to the OCT beam will be reduced because the light is Doppler shifted outside the bandpass filter of the OCT detection electronics. A 7.5-MHz ultrasound transducer was used to introduce the photon/phonon interaction. A reflecting metal plate was placed within biological tissue, and the point spread function (PSF) was assessed off the reflector. The PSF was determined in the presence of no ultrasound, pulsed ultrasound, and continuous-wave (CW) ultrasound. CW ultrasound resulted in a 17% improvement (P < 0.001) in resolution and pulsed ultrasound resulted in 8% (P < 0.01). Image noise reduction could also be noted. Combining OCT with a parallel ultrasound beam results in an improvement in resolution through a reduced effect of multiple scattering due to photon/phonon interaction. With higher frequencies, better control of the acoustical beam, and tests in media with higher rates of multiple scattering, improved results are anticipated. PMID- 12119392 TI - Ab initio/GIAO-MP2-calculated structures and (11)B-(13)C NMR chemical shift relationship in hypercoordinate onium-carbonium dications and isoelectronic onium boronium cations. AB - The boronium-carbonium ion continuum was extended to include hypercoordinated onium-carbonium dications and the isoelectronic onium-boronium cation analogs. Structures and (13)C and (11)B NMR chemical shifts of the onium-carbonium dications and the corresponding isoelectronic and isostructural onium-boronium cations were calculated with the ab initio/GIAO-MP2 method. The data show a good linear correlation between (11)B and (13)C NMR chemical shifts, indicating that the same factors that determine the chemical shifts of the boron nuclei also govern the chemical shifts of carbon nuclei of these hypercoordinated onium ions and dications. PMID- 12119393 TI - Self-assembling peptide hydrogel fosters chondrocyte extracellular matrix production and cell division: implications for cartilage tissue repair. AB - Emerging medical technologies for effective and lasting repair of articular cartilage include delivery of cells or cell-seeded scaffolds to a defect site to initiate de novo tissue regeneration. Biocompatible scaffolds assist in providing a template for cell distribution and extracellular matrix (ECM) accumulation in a three-dimensional geometry. A major challenge in choosing an appropriate scaffold for cartilage repair is the identification of a material that can simultaneously stimulate high rates of cell division and high rates of cell synthesis of phenotypically specific ECM macromolecules until repair evolves into steady-state tissue maintenance. We have devised a self-assembling peptide hydrogel scaffold for cartilage repair and developed a method to encapsulate chondrocytes within the peptide hydrogel. During 4 weeks of culture in vitro, chondrocytes seeded within the peptide hydrogel retained their morphology and developed a cartilage like ECM rich in proteoglycans and type II collagen, indicative of a stable chondrocyte phenotype. Time-dependent accumulation of this ECM was paralleled by increases in material stiffness, indicative of deposition of mechanically functional neo-tissue. Taken together, these results demonstrate the potential of a self-assembling peptide hydrogel as a scaffold for the synthesis and accumulation of a true cartilage-like ECM within a three-dimensional cell culture for cartilage tissue repair. PMID- 12119394 TI - Decoupling of unpolluted temperate forests from rock nutrient sources revealed by natural (87)Sr/(86)Sr and (84)Sr tracer addition. AB - An experimental tracer addition of (84)Sr to an unpolluted temperate forest site in southern Chile, as well as the natural variation of (87)Sr/(86)Sr within plants and soils, indicates that mechanisms in shallow soil organic horizons are of key importance for retaining and recycling atmospheric cation inputs at scales of decades or less. The dominant tree species Nothofagus nitida feeds nearly exclusively (>90%) on cations of atmospheric origin, despite strong variations in tree size and location in the forest landscape. Our results illustrate that (i) unpolluted temperate forests can become nutritionally decoupled from deeper weathering processes, virtually functioning as atmospherically fed ecosystems, and (ii) base cation turnover times are considerably more rapid than previously recognized in the plant available pool of soil. These results challenge the prevalent paradigm that plants largely feed on rock-derived cations and have important implications for understanding sensitivity of forests to air pollution. PMID- 12119395 TI - A R2R3-MYB gene, AtMYB30, acts as a positive regulator of the hypersensitive cell death program in plants in response to pathogen attack. AB - Hypersensitive response (HR) is a programmed cell death that is commonly associated with disease resistance in plants. Among the different HR-related early induced genes, the AtMYB30 gene is specifically, rapidly, and transiently expressed during incompatible interactions between Arabidopsis and bacterial pathogens. Its expression was also shown to be deregulated in Arabidopsis mutants affected in the control of cell death initiation. Here, we demonstrate that overexpression in Arabidopsis and tobacco of AtMYB30 (i) accelerates and intensifies the appearance of the HR in response to different avirulent bacterial pathogens, (ii) causes HR-like responses to virulent strains, and (iii) increases resistance against different bacterial pathogens, and a virulent biotrophic fungal pathogen, Cercospora nicotianae. In antisense AtMYB30 Arabidopsis lines, HR cell death is strongly decreased or suppressed in response to avirulent bacterial strains, resistance against different bacterial pathogens decreased, and the expression of HR- and defense-related genes was altered. Taken together, these results strongly suggest that AtMYB30 is a positive regulator of hypersensitive cell death. PMID- 12119396 TI - The molecular basis of the coloration mechanism in lobster shell: beta crustacyanin at 3.2-A resolution. AB - The binding of the carotenoid astaxanthin (AXT) in the protein multimacromolecular complex crustacyanin (CR) is responsible for the blue coloration of lobster shell. The structural basis of the bathochromic shift mechanism has long been elusive. A change in color occurs from the orange red of the unbound dilute AXT (lambda(max) 472 nm in hexane), the well-known color of cooked lobster, to slate blue in the protein-bound live lobster state (lambda(max) 632 nm in CR). Intriguingly, extracted CR becomes red on dehydration and on rehydration goes back to blue. Recently, the innovative use of softer x rays and xenon derivatization yielded the three-dimensional structure of the A(1) apoprotein subunit of CR, confirming it as a member of the lipocalin superfamily. That work provided the molecular replacement search model for a crystal form of the beta-CR holo complex, that is an A(1) with A(3) subunit assembly including two bound AXT molecules. We have thereby determined the structure of the A(3) molecule de novo. Lobster has clearly evolved an intricate structural mechanism for the coloration of its shell using AXT and a bathochromic shift. Blue/purple AXT proteins are ubiquitous among invertebrate marine animals, particularly the Crustacea. The three-dimensional structure of beta-CR has identified the protein contacts and structural alterations needed for the AXT color regulation mechanism. PMID- 12119398 TI - The CcmE protein of the c-type cytochrome biogenesis system: unusual in vitro heme incorporation into apo-CcmE and transfer from holo-CcmE to apocytochrome. AB - Three key steps of cytochrome c biogenesis in many Gram-negative bacteria, the uptake of heme by the heme chaperone CcmE, the covalent attachment of heme to CcmE, and its subsequent release from CcmE to an apocytochrome c, have been achieved in vitro. apo-CcmE from Escherichia coli preferentially bound to ferric, with high affinity (K(d), 200 nM), rather than ferrous heme. The preference for ferric heme was confirmed by competition with 8-anilino-1-naphthalenesulfonate, which bound to a hydrophobic pocket in apo-CcmE. Reduction under certain conditions of the ferric heme-CcmE complex, which has characteristics of a b-type cytochrome, resulted in covalent attachment of heme to the protein. The resulting in vitro-produced holo-CcmE was identical to the in vivo-produced holo-CcmE, proving that unmodified Fe-protoporphyrin IX is incorporated into CcmE. Only noncovalent binding of mesoheme to CcmE was observed, thus implicating at least one vinyl group in covalent binding of heme to CcmE. Heme transferred in vitro from holo-CcmE to apocytochrome c, provided the heme was reduced. The necessity for reduced holo-CcmE might explain the role of the heme chaperone, i.e., prevention of reaction of ferric heme with apocytochrome and thus avoidance of incorrect side products. In addition, an AXXAH mutant of the CXXCH binding motif in the apocytochrome c was unable to accept heme from holo-CcmE. These in vitro results mimic, and thus have implications for, the molecular pathway of heme transfer during c-type cytochrome maturation in many species of bacteria in vivo. PMID- 12119397 TI - R9AP, a membrane anchor for the photoreceptor GTPase accelerating protein, RGS9 1. AB - The regulator of G protein signaling (RGS)-9-1.G(beta 5) complex forms the GTPase accelerating protein for G(alpha t) in vertebrate photoreceptors. Although the complex is soluble when expressed in vitro, extraction of the endogenous protein from membranes requires detergents. The detergent extracts contain a complex of RGS9-1, G(beta 5), G(alpha t), and a 25-kDa phosphoprotein, R9AP (RGS9-1-Anchor Protein). R9AP is encoded by one intronless gene in both human and mouse. Full or partial cDNA or genomic clones were obtained from mice, cattle, human, zebrafish, and Xenopus laevis. R9AP mRNA was detected only in the retina, and the protein only in photoreceptors. R9AP binds to the N-terminal domain of RGS9-1, and anchors it to the disk membrane via a C-terminal transmembrane helix. PMID- 12119399 TI - Correlation of somatic hypermutation specificity and A-T base pair substitution errors by DNA polymerase eta during copying of a mouse immunoglobulin kappa light chain transgene. AB - To test the hypothesis that inaccurate DNA synthesis by mammalian DNA polymerase eta (pol eta) contributes to somatic hypermutation (SHM) of Ig genes, we measured the error specificity of mouse pol eta during synthesis of each strand of a mouse Ig kappa light chain transgene. We then compared the results to the base substitution specificity of SHM of this same gene in the mouse. The in vitro and in vivo base substitution spectra shared a number of common features. A highly significant correlation was observed for overall substitutions at A-T pairs but not for substitutions at G-C pairs. Sixteen mutational hotspots at A-T pairs observed in vivo were also found in spectra generated by mouse pol eta in vitro. The correlation was strongest for errors made by pol eta during synthesis of the non-transcribed strand, but it was also observed for synthesis of the transcribed strand. These facts, and the distribution of substitutions generated in vivo, support the hypothesis that pol eta contributes to SHM of Ig genes at A-T pairs via short patches of low fidelity DNA synthesis of both strands, but with a preference for the non-transcribed strand. PMID- 12119400 TI - Structure of decay-accelerating factor bound to echovirus 7: a virus-receptor complex. AB - Echoviruses are enteroviruses that belong to Picornaviridae. Many echoviruses use decay-accelerating factor (DAF) as their cellular receptor. DAF is a glycosylphosphatidyl inositol-anchored complement regulatory protein found on most cell surfaces. It functions to protect cells from complement attack. The cryo-electron microscopy reconstructions of echovirus 7 complexed with DAF show that the DAF-binding regions are located close to the icosahedral twofold axes, in contrast to other enterovirus complexes where the viral canyon is the receptor binding site. This novel receptor binding position suggests that DAF is important for the attachment of viral particles to host cells, but probably not for initiating viral uncoating, as is the case with canyon-binding receptors. Thus, a different cell entry mechanism must be used for enteroviruses that bind DAF. PMID- 12119401 TI - Glutathionylation of human thioredoxin: a possible crosstalk between the glutathione and thioredoxin systems. AB - To identify proteins undergoing glutathionylation (formation of protein glutathione mixed disulfides) in human T cell blasts, we radiolabeled the glutathione pool with (35)S, exposed cells to the oxidant diamide, and analyzed cellular proteins by two-dimensional electrophoresis. One of the proteins undergoing glutathionylation was identified by molecular weight, isoelectric point, and immunoblotting as thioredoxin (Trx). Incubation of recombinant human Trx with glutathione disulfide or S-nitrosoglutathione led to the formation of glutathionylated Trx, identified by matrix-assisted laser desorption ionization time-of-flight mass spectrometry. The glutathionylation site was identified as Cys-72. Glutathionylation of rhTrx abolished its enzymatic activity as insulin disulfide reductase in the presence of NADPH and Trx reductase. Activity was, however, regained with sigmoidal kinetics, indicating a process of autoactivation due to the ability of Trx to de-glutathionylate itself. These data suggest that the intracellular glutathione/glutathione disulfide ratio, an indicator of the redox state of the cell, can regulate Trx functions reversibly through thiol disulfide exchange reactions. PMID- 12119402 TI - Y586F mutation in murine leukemia virus reverse transcriptase decreases fidelity of DNA synthesis in regions associated with adenine-thymine tracts. AB - Using in vivo fidelity assays in which bacterial beta-galactosidase or green fluorescent protein genes served as reporters of mutations, we have identified a murine leukemia virus (MLV) RNase H mutant (Y586F) that exhibited an increase in the retroviral mutation rate approximately 5-fold in a single replication cycle. DNA-sequencing analysis indicated that the Y586F mutation increased the frequency of substitution mutations 17-fold within 18 nt of adenine-thymine tracts (AAAA, TTTT, or AATT), which are known to induce DNA bending. Sequence alignments indicate that MLV Y586 is equivalent to HIV-1 Y501, a component of the recently described RNase H primer grip domain, which contacts and positions the DNA primer strand near the RNase H active site. The results suggest that wild-type reverse transcriptase (RT) facilitates a specific conformation of the template-primer duplex at the polymerase active site that is important for accuracy of DNA synthesis; when an adenine-thymine tract is within 18 nt of the polymerase active site, the Y586F mutant RT cannot facilitate this specific template-primer conformation, leading to an increase in the frequency of substitution mutations. These findings indicate that the RNase H primer grip can affect the template primer conformation at the polymerase active site and that the MLV Y586 residue and template-primer conformation are important determinants of RT fidelity. PMID- 12119403 TI - Multiple centrosomes arise from tetraploidy checkpoint failure and mitotic centrosome clusters in p53 and RB pocket protein-compromised cells. AB - A high degree of aneuploidy characterizes the majority of human tumors. Aneuploid status can arise through mitotic or cleavage failure coupled with failure of tetraploid G(1) checkpoint control, or through deregulation of centrosome number, thus altering the number of mitotic spindle poles. p53 and the RB pocket proteins are important to the control of G(1) progression, and p53 has previously been suggested as important to the control of centrosome duplication. We demonstrate here that neither suppression of p53 nor of the RB pocket protein family directly generates altered centrosome numbers in any of several mammalian primary cell lines. Instead, amplification of centrosome number occurs in two steps. The first step is failure to arrest at a G(1) tetraploidy checkpoint after failure to segregate the genome in mitosis, and the second step is clustering of centrosomes at a single spindle pole in subsequent tetraploid or aneuploid mitosis. The trigger for these events is mitotic or cleavage failure that is independent of p53 or RB status. Finally, we find that mouse embryo fibroblasts spontaneously enter tetraploid G(1), explaining the previous demonstration of centrosome amplification by p53 abrogation alone in these cells. PMID- 12119404 TI - Designed to penetrate: time-resolved interaction of single antibiotic molecules with bacterial pores. AB - Membrane permeability barriers are among the factors contributing to the intrinsic resistance of bacteria to antibiotics. We have been able to resolve single ampicillin molecules moving through a channel of the general bacterial porin, OmpF (outer membrane protein F), believed to be the principal pathway for the beta-lactam antibiotics. With ion channel reconstitution and high-resolution conductance recording, we find that ampicillin and several other efficient penicillins and cephalosporins strongly interact with the residues of the constriction zone of the OmpF channel. Therefore, we hypothesize that, in analogy to substrate-specific channels that evolved to bind certain metabolite molecules, antibiotics have "evolved" to be channel-specific. Molecular modeling suggests that the charge distribution of the ampicillin molecule complements the charge distribution at the narrowest part of the bacterial porin. Interaction of these charges creates a region of attraction inside the channel that facilitates drug translocation through the constriction zone and results in higher permeability rates. PMID- 12119405 TI - Single residue modification of only one dimer within the hemoglobin tetramer reveals autonomous dimer function. AB - The mechanism of cooperativity in the human hemoglobin tetramer (a dimer of alpha beta dimers) has historically been modeled as a simple two-state system in which a low-affinity structural form (T) switches, on ligation, to a high-affinity form (R), yielding a net loss of hydrogen bonds and salt bridges in the dimer-dimer interface. Modifications that weaken these cross-dimer contacts destabilize the quaternary T tetramer, leading to decreased cooperativity and enhanced ligand affinity, as demonstrated in many studies on symmetric double modifications, i.e., a residue site modified in both alpha- or both beta-subunits. In this work, hybrid tetramers have been prepared with only one modified residue, yielding molecules composed of a wild-type dimer and a modified dimer. It is observed that the cooperative free energy of ligation to the modified dimer is perturbed to the same extent whether in the hybrid tetramer or in the doubly modified tetramer. The cooperative free energy of ligation to the wild-type dimer is unperturbed, even in the hybrid tetramer, and despite the overall destabilization of the T tetramer by the modification. This asymmetric response by the two dimers within the same tetramer shows that loss of dimer-dimer contacts is not communicated across the dimer-dimer interface, but is transmitted through the dimer that bears the modified residue. These observations are interpreted in terms of a previously proposed dimer-based model of cooperativity with an additional quaternary (T/R) component. PMID- 12119406 TI - Energetic communication between mitochondria and nucleus directed by catalyzed phosphotransfer. AB - Exchange of information between the nucleus and cytosol depends on the metabolic state of the cell, yet the energy-supply pathways to the nuclear compartment are unknown. Here, the energetics of nucleocytoplasmic communication was determined by imaging import of a constitutive nuclear protein histone H1. Translocation of H1 through nuclear pores in cardiac cells relied on ATP supplied by mitochondrial oxidative phosphorylation, but not by glycolysis. Although mitochondria clustered around the nucleus, reducing the distance for energy transfer, simple nucleotide diffusion was insufficient to meet the energetic demands of nuclear transport. Rather, the integrated phosphotransfer network was required for delivery of high energy phosphoryls from mitochondria to the nucleus. In neonatal cardiomyocytes with low creatine kinase activity, inhibition of adenylate kinase-catalyzed phosphotransfer abolished nuclear import. With deficient adenylate kinase, nucleoside diphosphate kinase, which secures phosphoryl exchange between ATP and GTP, was unable to sustain nuclear import. Up-regulation of creatine kinase phosphotransfer, to mimic metabolic conditions of adult cardiac cells, rescued H1 import, suggesting a developmental plasticity of the cellular energetic system. Thus, mitochondrial oxidative phosphorylation coupled with phosphotransfer relays provides an efficient energetic unit in support of nuclear transport. PMID- 12119407 TI - The metal reductase activity of some multiheme cytochromes c: NMR structural characterization of the reduction of chromium(VI) to chromium(III) by cytochrome c(7). AB - The redox reaction between CrO(4)(2-) and the fully reduced three-heme cytochrome c(7) from Desulfuromonas acetoxidans to give chromium(III) and the fully oxidized protein has been followed by NMR spectroscopy. The hyperfine coupling between the oxidized protein protons and chromium(III), which remains bound to the protein, gives rise to line-broadening effects on the NMR resonances that can be transformed into proton-metal distance restraints. Structure calculations based on these unconventional constraints allowed us to demonstrate that chromium(III) binds at a unique site and to locate it on the protein surface. The metal ion is located 7.9 +/- 0.4 A from the iron of heme IV, 16.3 +/- 0.7 A from the iron of heme III, and 22.5 +/- 0.5 A from the iron of heme I. Shift changes caused by the presence of unreactive MoO(4)(2-), a CrO(4)(2-) analogue, indicate the involvement of the same protein area in the anion binding. The titration of the oxidation of cytochrome c(7) shows a detailed mechanism of action. The presence of a specific binding site supports the hypothesis of the biological role of this cytochrome as a metal reductase. PMID- 12119408 TI - Transcripts of Vp-1 homeologues are misspliced in modern wheat and ancestral species. AB - The maize (Zea mays) Viviparous 1 (Vp1) transcription factor has been shown previously to be a major regulator of seed development, simultaneously activating embryo maturation and repressing germination. Hexaploid bread wheat (Triticum aestivum) caryopses are characterized by relatively weak embryo dormancy and are susceptible to preharvest sprouting (PHS), a phenomenon that is phenotypically similar to the maize vp1 mutation. Analysis of Vp-1 transcript structure in wheat embryos during grain development showed that each homeologue produces cytoplasmic mRNAs of different sizes. The majority of transcripts are spliced incorrectly, contain insertions of intron sequences or deletions of coding region, and do not have the capacity to encode full-length proteins. Several VP-1-related lower molecular weight protein species were present in wheat embryo nuclei. Embryos of a closely related tetraploid species (Triticum turgidum) and ancestral diploids also contained misspliced Vp-1 transcripts that were structurally similar or identical to those found in modern hexaploid wheat, which suggests that compromised structure and expression of Vp-1 transcripts in modern wheat are inherited from ancestral species. Developing embryos from transgenic wheat grains expressing the Avena fatua Vp1 gene showed enhanced responsiveness to applied abscisic acid compared with the control. In addition, ripening ears of transgenic plants were less susceptible to PHS. Our results suggest that missplicing of wheat Vp-1 genes contributes to susceptibility to PHS in modern hexaploid wheat varieties and identifies a possible route to increase resistance to this environmentally triggered disorder. PMID- 12119410 TI - An anatomy of normal and malignant gene expression. AB - A gene's expression pattern provides clues to its role in normal physiology and disease. To provide quantitative expression levels on a genome-wide scale, the Cancer Genome Anatomy Project (CGAP) uses serial analysis of gene expression (SAGE). Over 5 million transcript tags from more than 100 human cell types have been assembled. To enhance the utility of this data, the CGAP SAGE project created SAGE Genie, a web site for the analysis and presentation of SAGE data (http://cgap.nci.nih.gov/SAGE). SAGE Genie provides an automatic link between gene names and SAGE transcript levels, accounting for alternative transcription and many potential errors. These informatics advances provide a rapid and intuitive view of transcript expression in the human body or brain, displayed on the SAGE Anatomic Viewer. We report here an easily accessible view of nearly any gene's expression in a wide variety of malignant and normal tissues. PMID- 12119409 TI - Haploinsufficiency of Flap endonuclease (Fen1) leads to rapid tumor progression. AB - Flap endonuclease (Fen1) is required for DNA replication and repair, and defects in the gene encoding Fen1 cause increased accumulation of mutations and genome rearrangements. Because mutations in some genes involved in these processes cause cancer predisposition, we investigated the possibility that Fen1 may function in tumorigenesis of the gastrointestinal tract. Using gene knockout approaches, we introduced a null mutation into murine Fen1. Mice homozygous for the Fen1 mutation were not obtained, suggesting absence of Fen1 expression leads to embryonic lethality. Most Fen1 heterozygous animals appear normal. However, when combined with a mutation in the adenomatous polyposis coli (Apc) gene, double heterozygous animals have increased numbers of adenocarcinomas and decreased survival. The tumors from these mice show microsatellite instability. Because one copy of the Fen1 gene remained intact in tumors, Fen1 haploinsufficiency appears to lead to rapid progression of cancer. PMID- 12119411 TI - Molecular portraits of B cell lineage commitment. AB - In an attempt to characterize early B cell development including the commitment of progenitor cells to the B cell lineage, we generated and compared genomewide gene expression profiles of human hematopoietic stem cells (HSCs) and pre-B cells (PBCs) by using serial analysis of gene expression. From more than 100,000 serial analysis of gene expression tags collected from human CD34(+) HSCs and CD10(+) CD19(+) PBCs, 42,399 unique transcripts were identified in HSCs but only 16,786 in PBCs, suggesting that more than 60% of transcripts expressed in HSCs were silenced during or after commitment to the B cell lineage. On the other hand, mRNAs of pre-B cell receptor (pre-BCR)-associated genes are virtually missing in HSCs but account for more than 10% of the transcriptome of PBCs, which also show increased expression of apoptosis-related genes. Both concentration of the transcriptional repertoire on pre-BCR-related genes together with marked up regulation of apoptosis mediators in PBC might reflect selection for the expression of a functional pre-BCR within the bone marrow. Besides known regulator genes of early B cell development such as PAX5, E2A, and EBF, the most abundantly expressed genes in PBCs include ATM, PDGFRA, SIAH1, PIM2, C/EBPB, WNT16, and TCL1, the role of which has not been established yet in early B cell development. PMID- 12119412 TI - Multiple label-free biodetection and quantitative DNA-binding assays on a nanomechanical cantilever array. AB - We report a microarray of cantilevers to detect multiple unlabeled biomolecules simultaneously at nanomolar concentrations within minutes. Ligand-receptor binding interactions such as DNA hybridization or protein recognition occurring on microfabricated silicon cantilevers generate nanomechanical bending, which is detected optically in situ. Differential measurements including reference cantilevers on an array of eight sensors can sequence-specifically detect unlabeled DNA targets in 80-fold excess of nonmatching DNA as a background and discriminate 3' and 5' overhangs. Our experiments suggest that the nanomechanical motion originates from predominantly steric hindrance effects and depends on the concentration of DNA molecules in solution. We show that cantilever arrays can be used to investigate the thermodynamics of biomolecular interactions mechanically, and we have found that the specificity of the reaction on a cantilever is consistent with solution data. Hence cantilever arrays permit multiple binding assays in parallel and can detect femtomoles of DNA on the cantilever at a DNA concentration in solution of 75 nM. PMID- 12119413 TI - Ubiquitin ligase-associated protein SGT1 is required for host and nonhost disease resistance in plants. AB - Homologues of the yeast ubiquitin ligase-associated protein SGT1 are required for disease resistance in plants mediated by nucleotide-binding site/leucine-rich repeat (NBS-LRR) proteins. Here, by silencing SGT1 in Nicotiana benthamiana, we extend these findings and demonstrate that SGT1 has an unexpectedly general role in disease resistance. It is required for resistance responses mediated by NBS LRR and other R proteins in which pathogen-derived elicitors are recognized either inside or outside the host plant cell. A requirement also exists for SGT1 in nonhost resistance in which all known members of a host species are resistant against every characterized isolate of a pathogen. Our findings show that silencing SGT1 affects diverse types of disease resistance in plants and support the idea that R protein-mediated and nonhost resistance may involve similar mechanisms. PMID- 12119415 TI - Molecular recognition of oxygen by protein mimics: dynamics on the femtosecond to microsecond time scale. AB - Molecular recognition by biological macromolecules involves many elementary steps, usually convoluted by diffusion processes. Here we report studies of the dynamics, from the femtosecond to the microsecond time scale, of the different elementary processes involved in the bimolecular recognition of a protein mimic, cobalt picket-fence porphyrin, with varying oxygen concentration at controlled temperatures. Electron transfer, bond breakage, and thermal "on" (recombination) and "off" (dissociation) reactions are the different processes involved. The reaction on-rate is 30 to 60 times smaller than that calculated from standard Smoluchowski theory. Introducing a two-step recognition model, with reversibility being part of both steps, removes the discrepancy and provides consistency for the reported thermodynamics, kinetics, and dynamics. The transient intermediates are configurations defined by the contact between oxygen (diatomic) and the picket-fence porphyrin (macromolecule). This intermediate is critical in the description of the potential energy landscape but, as shown here, both enthalpic and entropic contributions to the free energy are important. In the recognition process, the net entropy decrease is -33 cal mol(-1) K(-1); Delta H is -13.4 kcal mol(-1). PMID- 12119414 TI - Regional analysis of p53 mutations in rheumatoid arthritis synovium. AB - The p53 tumor suppressor protein plays a central role in cell cycle regulation, DNA repair, and apoptosis. Recent studies indicate that DNA damage and somatic mutations in the p53 gene can occur because of genotoxic stress in many tissues, including the skin, colon, and synovium. Although somatic mutations in the p53 gene have been demonstrated in rheumatoid arthritis (RA) synovial tissue and synoviocytes, no information is available on the location or extent of p53 mutations. Using microdissected RA synovial tissue sections, we observed abundant p53 transition mutations, which are characteristic DNA damage caused by oxidative stress. p53 mutations, as well as p53 mRNA expression, were located mainly in the synovial intimal lining rather than the sublining (P < 0.01). Clusters of p53 mutant subclones were observed in some microdissected regions, suggesting oligoclonal expansion. Because IL-6 gene expression is regulated by wild-type p53, IL-6 mRNA expression in microdissected tissues was quantified by using real time PCR. The regions with high rates of p53 mutations contained significantly greater amounts of IL-6 mRNA compared with the low mutation samples (P < 0.02). The microdissection findings suggest that p53 mutations are induced in RA synovial tissues by inflammatory oxidative stress. This process, as in sun exposed skin and inflamed colonic epithelium, provides some of the mutant clones with a selective growth advantage. A relatively low percentage of cells containing p53 mutations can potentially affect neighboring cells and enhance inflammation through the elaboration of proinflammatory cytokines. PMID- 12119416 TI - Crystal structure of monomeric human beta-2-microglobulin reveals clues to its amyloidogenic properties. AB - Dissociation of human beta-2-microglobulin (beta(2)m) from the heavy chain of the class I HLA complex is a critical first step in the formation of amyloid fibrils from this protein. As a consequence of renal failure, the concentration of circulating monomeric beta(2)m increases, ultimately leading to deposition of the protein into amyloid fibrils and development of the disorder, dialysis-related amyloidosis. Here we present the crystal structure of a monomeric form of human beta(2)m determined at 1.8-A resolution that reveals remarkable structural changes relative to the HLA-bound protein. These involve the restructuring of a beta bulge that separates two short beta strands to form a new six-residue beta strand at one edge of this beta sandwich protein. These structural changes remove key features proposed to have evolved to protect beta sheet proteins from aggregation [Richardson, J. & Richardson, D. (2002) Proc. Natl. Acad. Sci. USA 99, 2754-2759] and replaces them with an aggregation-competent surface. In combination with solution studies using (1)H NMR, we show that the crystal structure presented here represents a rare species in solution that could provide important clues about the mechanism of amyloid formation from the normally highly soluble native protein. PMID- 12119418 TI - Association of calnexin with mutant peripheral myelin protein-22 ex vivo: a basis for "gain-of-function" ER diseases. AB - Schwann cell-derived peripheral myelin protein-22 (PMP-22) when mutated or overexpressed causes heritable neuropathies with a previously unexplained "gain of-function" endoplasmic reticulum (ER) retention phenotype. In wild-type sciatic nerves, PMP-22 associates in a specific, transient (t(1/2 ) approximately equal to 11 min), and oligosaccharide processing-dependent manner with the lectin chaperone calnexin (CNX), but not calreticulin nor BiP. In Trembler-J (Tr-J) sciatic nerves, prolonged association of mutant PMP-22 with CNX is found (t(1/2) > 60 min). In 293A cells overexpressing PMP-22(Tr-J), CNX and PMP-22 colocalize in large intracellular structures identified at the electron microscopy level as myelin-like figures with CNX localization in the structures dependent on PMP-22 glucosylation. Similar intracellular myelin-like figures were also present in Schwann cells of sciatic nerves from homozygous Trembler-J mice with no detectable activation of the stress response pathway as deduced from BiP and CHOP expression. Sequestration of CNX in intracellular myelin-like figures may be relevant to the autosomal dominant Charcot-Marie-Tooth-related neuropathies. PMID- 12119417 TI - A crucial role for the putative Arabidopsis topoisomerase VI in plant growth and development. AB - Plant steroid hormones, brassinosteroids (BRs), play important roles throughout plant growth and development. Plants defective in BR biosynthesis or perception display cell elongation defects and severe dwarfism. Two dwarf mutants named bin3 and bin5 with identical phenotypes to each other display some characteristics of BR mutants and are partially insensitive to exogenously applied BRs. In the dark, bin3 or bin5 seedlings are de-etiolated with short hypocotyls and open cotyledons. Light-grown mutant plants are dwarfs with short petioles, epinastic leaves, short inflorescence stems, and reduced apical dominance. We cloned BIN3 and BIN5 and show that BIN5 is one of three putative Arabidopsis SPO11 homologs (AtSPO11-3) that also shares significant homology to archaebacterial topoisomerase VI (TOP6) subunit A, whereas BIN3 represents a putative eukaryotic homolog of TOP6B. The pleiotropic dwarf phenotypes of bin5 establish that, unlike all of the other SPO11 homologs that are involved in meiosis, BIN5/AtSPO11-3 plays a major role during somatic development. Furthermore, microarray analysis of the expression of about 5500 genes in bin3 or bin5 mutants indicates that about 321 genes are down-regulated in both of the mutants, including 18 of 30 BR induced genes. These results suggest that BIN3 and BIN5 may constitute an Arabidopsis topoisomerase VI that modulates expression of many genes, including those regulated by BRs. PMID- 12119419 TI - Self-assembling biomaterials: liquid crystal phases of cholesteryl oligo(L-lactic acid) and their interactions with cells. AB - We report here on the synthesis and characterization of a series of self assembling biomaterials with molecular features designed to interact with cells and scaffolds for tissue regeneration. The molecules of these materials contain cholesteryl moieties, which have universal affinity for cell membranes, and short chains of lactic acid, a common component of biodegradable tissue engineering matrices. The materials were synthesized in good yields with low polydispersities in the range of 1.05-1.15, and their characterization was carried out by small angle x-ray diffraction, transmission electron microscopy, electron diffraction, differential scanning calorimetry, and atomic force microscopy. These molecular materials form layered structures that can be described as smectic phases and can also order into single-crystal stacks with an orthorhombic unit cell. Their layer spacings range from 58 to 99 A, corresponding to bilayers of oligomers with an average of 10 and 37 lactic acid residues, respectively. The self-organized layered structures were found to promote improved fibroblast adhesion and spreading, although the specific mechanism for this observed response remains unknown. The ability of self-assembling materials to present ordered and periodic bulk structures to cells could be a useful strategy in tissue engineering. PMID- 12119420 TI - Role of the E1A Rb-binding domain in repression of the NF-kappa B-dependent defense against tumor necrosis factor-alpha. AB - The adenoviral E1A oncogene sensitizes mammalian cells to tumor necrosis factor alpha (TNF-alpha), in part by repressing the nuclear factor-kappa B (NF-kappa B) dependent defense against this cytokine. Other E1A activities involve binding to either p300/cyclic AMP response element-binding protein (CBP) or retinoblastoma (Rb)-family proteins, but the roles of E1A interactions with these transcriptional regulators in sensitizing cells to TNF-alpha are unclear. E1A expression did not block upstream events in TNF-alpha-induced activation of NF kappa B in NIH 3T3 cells, including degradation of I kappa B-alpha, nuclear translocation of NF-kappa B subunits, and their dimeric binding to kappa B sequences in the nucleus. However, E1A markedly repressed NF-kappa B-dependent transcription and sensitized cells to TNF-alpha induced apoptosis. These E1A effects were selective for kappa B-dependent transcription and for the function of the NF-kappa B p65/RelA subunit. A four amino acid E1A deletion that eliminates binding to Rb-family proteins blocked both repression of TNF-alpha induced transcription and sensitization to apoptosis. In contrast, mutations that eliminate E1A binding to p300/CBP (coactivators of p65/RelA) did not affect either E1A activity. These data suggest that E1A-Rb-binding blocks the NF-kappa B dependent activation response to TNF-alpha by altering the function of p65/RelA at a stage after formation of the transcription factor-enhancer complex. These observations also open questions about the general role of Rb-family proteins in modulation of NF-kappa B-dependent transcription. PMID- 12119421 TI - Characterization of three alternatively spliced isoforms of the Rel/NF-kappa B transcription factor Relish from the mosquito Aedes aegypti. AB - The Rel/NF-kappa B transcription factor Relish performs a central role in the acute-phase response to microbial challenge by activating immune antibacterial peptides. We cloned and molecularly characterized the gene homologous to Drosophila Relish from the mosquito Aedes aegypti. Unlike Drosophila Relish, Aedes Relish has three alternatively spliced transcripts encoding different proteins. First, the predominant Aedes Relish transcript of 3.9 kb contains both the Rel-homology domains and the inhibitor kappa B (I kappa B)-like domain, which is similar to Drosophila Relish and to the mammalian p105 and p100 Rel/NF-kappa B transcription factors. Second, Aedes Relish transcript contains Rel-homology domains identical to those of the major transcript but it completely lacks the I kappa B-like domain-coding region, which has been replaced by a unique 3' untranslated region sequence. In the third transcript, a deletion replaces most of the N-terminal sequence and Rel-homology domains; however, the I kappa B-like domain is intact. All three Aedes Relish transcripts were induced by bacterial injection but not by blood feeding. In vitro-translated protein from the Rel-only construct specifically binds to the kappa B motif from Drosophila cecropin A1 and Aedes defensin genes. PCR and Southern blot hybridization analyses show that these three transcripts originated from the same large inducible mRNA encoded by a single Relish gene. PMID- 12119422 TI - Atm heterozygous mice are more sensitive to radiation-induced cataracts than are their wild-type counterparts. AB - It is important to know whether the human population includes genetically predisposed radiosensitive subsets. In vitro studies have shown that cells from individuals homozygous for ataxia telangiectasia (A-T) are much more radiosensitive than cells from unaffected individuals. Although cells heterozygous for the ATM gene (ATM(+/-)) may be slightly more radiosensitive in vitro, it remained to be determined whether the greater susceptibility of ATM(+/ ) cells translates into an increased sensitivity for late effects in vivo, though there is a suggestion that radiotherapy patients that are heterozygous for the ATM gene may be more at risk of developing late normal tissue damage. We chose cataractogenesis in the lens as a means to assay for the effects of ATM deficiency in a late-responding tissue. One eye of wild-type, Atm heterozygous and homozygous knockout mice was exposed to 0.5-, 1.0-, 2.0-, or 4.0-Gy x rays. The animals were followed weekly for cataract development by conventional slit lamp biomicroscopy. Cataract development in the animals of all three groups was strongly dependent on dose. The lenses of homozygous mice were the first to opacify at any given dose. Most important in the present context is that cataracts appeared earlier in the heterozygous versus wild-type animals. The data suggest that ATM heterozygotes in the human population may also be radiosensitive. This may influence the choice of individuals destined to be exposed to higher than normal doses of radiation, such as astronauts, and may also suggest that radiotherapy patients who are ATM heterozygotes could be predisposed to increased late normal tissue damage. PMID- 12119425 TI - Editorial. PMID- 12119423 TI - Prominent neurodegeneration and increased plaque formation in complement inhibited Alzheimer's mice. AB - Abnormal accumulation of beta-amyloid (Abeta) in Alzheimer's disease (AD) is associated with prominent brain inflammation. Whereas earlier studies concluded that this inflammation is detrimental, more recent animal data suggest that at least some inflammatory processes may be beneficial and promote Abeta clearance. Consistent with these observations, overproduction of transforming growth factor (TGF)-beta1 resulted in a vigorous microglial activation that was accompanied by at least a 50% reduction in Abeta accumulation in human amyloid precursor protein (hAPP) transgenic mice. In a search for inflammatory mediators associated with this reduced pathology, we found that brain levels of C3, the central component of complement and a key inflammatory protein activated in AD, were markedly higher in hAPP/TGF-beta1 mice than in hAPP mice. To assess the importance of complement in the pathogenesis of AD-like disease in mice, we inhibited C3 activation by expressing soluble complement receptor-related protein y (sCrry), a complement inhibitor, in the brains of hAPP mice. Abeta deposition was 2- to 3 fold higher in 1-year-old hAPP/sCrry mice than in age-matched hAPP mice and was accompanied by a prominent accumulation of degenerating neurons. These results indicate that complement activation products can protect against Abeta-induced neurotoxicity and may reduce the accumulation or promote the clearance of amyloid and degenerating neurons. These findings provide evidence for a role of complement and innate immune responses in AD-like disease in mice and support the concept that certain inflammatory defense mechanisms in the brain may be beneficial in neurodegenerative disease. PMID- 12119426 TI - Labor Supply of Poor Residents in Metropolitan Miami, Florida: The Role of Depression and the Co-Morbid Effects of Substance Use. AB - BACKGROUND: Depression represents one of the most common behavioral health problems among the workforce in the United States, with about 1 in every 20 employees experiencing this condition. A recent study estimated that in 1990 the economic costs of depressive disorders in the American workplace amounted to as much as $43 billion, with absenteeism alone accounting for $12 billion. Recently, economists have been focusing attention on the relationship between mental health and labor supply, but a lack of quality data sets containing detailed information on mental health and labor market variables represents a significant barrier to rigorous research. AIMS OF THE STUDY: The primary aims of the present study were to (i) examine the relationship between depression and employment, (ii) conditional on being employed, estimate the effect of depression on annual weeks worked, and (iii) examine the stability of the model estimates to the co-morbid effects of substance use (illicit drugs and alcohol), which has been consistently found to be a correlate of depression. DATA: The study used a unique set of survey data collected between 1996 and 1997 in crime-ridden and low-income neighborhoods of Miami-Dade County, Florida. A targeted sampling strategy was used to recruit chronic drug users (including injection drug users) and non-drug users to examine local health care delivery system characteristics in relation to the population of substance users. The final analysis sample for the present study included 1,274 adults, aged 18 to 65. Depression status was measured from the 20-item Zung Self-Rating Depression Scale (SDS) that classified 384 individuals as depressed and 890 as non-depressed. According to the definition developed by the U.S. Office of National Drug Control Policy for chronic drug use (CDU), about 46 percent of the depressed individuals were found to be CDUs compared to 30 percent of the non-depressed sample. The survey instrument collected information on alcohol use and problem drinking as defined by the 10 item Michigan Alcoholism Screening Test (MAST-10). Based on criteria defined in the MAST-10, 26 percent of the depressed individuals were problematic alcohol users (PAUs) compared to about 16 percent of the non-depressed sample.METHODS: The labor supply measures included employment in the past 30 days and number of weeks worked in the past 12 months. The analysis estimated a univariate probit model of employment as well as a bivariate probit model of depression and employment, which accounted for the possible correlation between the unobserved determinants of depression and employment. The annual weeks worked specification was estimated by a standard Tobit model as well as an instrumental variable (IV) Tobit model, which, in addition to the censoring of the observations, accounted for the possible endogeneity of depression. The stability of the estimated effects of depression to comorbid illicit drug and alcohol use was assessed, by controlling for CDU and PAU in these models. RESULTS: Results from both the univariate probit and the bivariate probit models indicate that depression significantly decreased the probability of being employed. Specifically, depression reduced the probability of employment by an average of 19 percentage points in both models, from a sample average of 43 percent for the non- depressed to 24 percent for the depressed. Estimates from the Tobit models revealed that depression also significantly reduced the number of weeks worked. Conditional on being employed, depressed individuals worked an average of 7 fewer annual weeks than the non-depressed sample in the univariate Tobit model and 8 fewer weeks in the IV Tobit. The findings also showed that the effects of depression on employment and annual weeks worked may be over-estimated if the analysis does not account for the comorbid influence of substance use. IMPLICATIONS FOR HEALTH CARE PROVISION AND USE: The results suggest that prevention and/or treatment of mental health problems such as depression may yield economic benefits by promoting employment and enhancing labor supply. While expansion of public mental health services may not lead to overall increases in employment, it may be justified on social grounds given the high unemployment rate in low-income and crime-ridden neighborhoods. Further insights can be gained by estimating these models with national and international data if one applies appropriate econometric tools to account for complex sample designs. PMID- 12119424 TI - Imaging of metabolites by using a fusion protein between a periplasmic binding protein and GFP derivatives: from a chimera to a view of reality. PMID- 12119427 TI - Drug Treatment as a Crime Fighting Tool. AB - BACKGROUND: The primary approach to reducing crime in the US has been through the criminal justice system. However, drug treatment may be an effective tool in reducing crime. In order to make better use of treatment as an alternative approach, one needs to know if reducing drug use through treatment results in decreased crime. AIMS OF THE STUDY: The objective of this paper is to model and empirically investigate the extent to which a change in drug use that results from treatment reduces crime and whether a change in drug use is causally related to change in crime. We focus on crime-for-profit. METHODS: We use a multi-site dataset of 3,502 inner-city drug users entering treatment. We analyze the change in drug use and crime pre and post treatment. We take first differences to address the omitted variable problem. RESULTS: We find that treatment reduces drug use and that, in turn, reduced drug use has a significant impact on crime. For our study population, reduced drug use seems to be causally related to reduced crime. This finding is robust to specification and subsamples. We estimate that reduced drug use due to treatment is associated with 54% fewer days of crime for profit, ceteris paribus. DISCUSSION: We use a longitudinal data set and a novel approach to analyze the relationship between crime and drugs. We analyze a low-income, inner-city, drug-addicted sample. We use self-reported crime. For our purposes, the use of individual data is an improvement over the use of aggregate level data that has been used in much of the related literature. Limitations of our paper include that we do not have a random sample and that our measure is self-reported in the previous 30 days. IMPLICATIONS FOR HEALTH POLICIES: Our findings suggest that drug treatment may be an effective crime fighting tool. Treatment reduces not only the crime of drug possession, but also crime-for-profit. Current public policy emphasizes use of the criminal justice system, incarceration in particular, as a mechanism to combat crime. Given the huge and growing expense of the criminal justice system, drug treatment might be cost-effective relative to incarceration. California s so called Proposition 36 is based on this yet to be proven premise. Although additional research is required, our findings may help inform the debate on treatment versus criminal justice. We have provided empirically-based findings that reduced drug use due to treatment can result in important reductions in crime. Our findings can serve as a building block for policy development. PMID- 12119428 TI - Profile of Community Mental Health Service Needs in the Moretele District (North West Province) in South Africa. AB - BACKGROUND: The emergence of democracy in South Africa led to a need to transform all public structures, including the health care system. The aim has been to transform these structures in order to bring them in line with the new culture of human rights. Transformation of the whole health care system is motivated by a number of key objectives, which include achieving equity in resource allocation and health service delivery, developing primary health care infrastructure and decentralising services to promote community participation. AIMS OF THE STUDY: In the context of de-institutionalising mental health services in South Africa, this study aimed to investigate community mental health service needs of mental health service users and that of their families in the Moretele district, North-West province, South Africa. METHODS: The study was conducted in three clinics situated in three different communities in the Moretele district. Data collection consisted of : 147 clinical record reviews, 105 interviews with patients followed by a joint interview with a family member, 83 interviews with caregivers and eight interviews with community key informants (traditional healers, a civic leader, a councillor, a retired teacher, and a physician). RESULTS: The majority of service users were males (54%). The mean age was 41 years and 63% had completed primary schooling.Patients were recorded as having only one of two primary diagnoses, namely schizophrenia (57%) or epilepsy (41%). However, a review of prescribed drugs and caregiver interviews showed that there was a presence of mood disorders among service users. The local hospital was service users primary entry point into the mental health care system, followed by traditional healers (30%). Interviews with service users, service providers and caregivers reveal limited knowledge of patient illness. Nevertheless, service users who had epilepsy were more likely to provide details of their illness than those with mental illness. Above half of service users had basic social skills such as bathing, eating, washing and using public transportation independently. Feelings of loneliness and isolation were common among service users in the community; seventy nine percent (79%), for example spent their days entirely in their homes. Only 7% reported contact with friends. Experiences of community discrimination and exploitation of people with mental illness were reported in key informant interviews and by service users themselves. DISCUSSION: The main community mental health service needs identified in this study were: (i) Improved quality of mental health services at clinics. (ii) Better co-ordination of services (clinic, hospital, social work and traditional healers) and removal of barriers to health service utilisation. (iii) Alleviation of the social isolation of mental health service users by building on existing community structures and individuals willing to engage in partnerships with service providers. The authors indentify a need to train primary health care providers in mental health in order to promote the adequate diagnosis and detection of common psychiatric illnesses.They also point out the need for social support interventions to enable people with mental illness to deal with loneliness and isolation and a need for psycho-educational programmes to make patients and caregivers better informed about mental illness. PMID- 12119429 TI - Income and Mental Health: Unraveling Community and Individual Level Relationships. AB - BACKGROUND: The association between individual socioeconomic status (SES) and mental disorder is well-documented, but studies to date have provided limited and sometimes conflicting evidence on the relationship between aspects of socioeconomic environment, including the role of income inequality, and mental disorder. AIMS OF THE STUDY: This paper explores the relationships between mental disorder and individual SES and socioeconomic environment, with particular attention to both the level and dispersion of community income and to their interactions with individual income. METHODS: Cross sectional study using nationally representative, individual level data from the Healthcare for Communities survey merged with supplemental information. Dependent variable is individual mental health status, measured by the 5 item Mental Health Inventory (MHI-5; average 80.6) and an indicator of probable anxiety or mood disorder based on clinical screening instruments (positive for 14.3 percent of respondents in the sample). RESULTS: MHI-5 decreases (indicating worse mental health), and the probability of an anxiety or depressive disorder increases continuously from the highest to the lowest quintiles of family income. Compared to those in the highest income quintile, MHI-5 is more than 10 points lower and the probability of disorder is much greater among individuals in the lowest income quintile. Within-quintile own income level is also strongly associated with mental health among lower income individuals. We find no evidence that higher levels of income inequality are associated with poor mental health outcomes, measured either by the probability of disorder or MHI-5. Regarding income level, MHI-5 is 3.4 to 3.5 points higher among low income individuals in medium or high income states compared to those in low income states. DISCUSSION: The qualitative conclusions are stable across various specifications reported (two different measures of mental health, two geographic levels, and among all individuals and low income individuals alone), and in specifications with alternative parameterizations of the community variables (continuously measured, included as quintiles instead of tertiles, and using other indicators of inequality). Individual income is highly correlated with mental health status; level of state income has some association; community or state income inequality has no detectable relationship with mental health. This analysis provides no support for the hypothesis that income inequality is a stronger determinant of health than individual or family income, a hypothesis that in recent years has received much attention in the popular press and policy debates. Limitations of our analysis include the cross-sectional nature of the analysis, the sample sizes used in derivation of the site variables (though sensitivity analyses showed robust results) and the age of the state data. CONCLUSIONS: The association between individual income and mental health is strong. No support for the income inequality hypothesis is found.IMPLICATIONS FOR HEALTH POLICY FORMULATION: Our findings point to a need for better understanding of the relationship between individual income and mental health outcomes. Our research does not support the notion that policies aimed at diminishing income inequality are an important lever in improving mental health outcomes for individuals. IMPLICATIONS FOR FURTHER RESEARCH: This research does not address whether and how different sources of income-at the individual or community level may affect mental health, and whether the associations observed cross-sectionally also bear out longitudinally. In addition, more research into the relationship between other community characteristics, such as service availability, and mental health outcomes is needed. PMID- 12119430 TI - Cystic dysplasia of the testis: a very rare paediatric tumor of the testis. AB - OBJECTIVES: To describe a case of cystic dysplasia of the testis (CDT), an uncommon cause of scrotal swelling in the pediatric patient. Clinic, therapy, fertility, and radiographic and pathologic findings are discussed and the 30 previously reported cases are reviewed. METHODS: A 9-year-old boy presented with asymptomatic scrotal swelling. A scrotal ultrasound showed a multicystic scrotal mass in the rete testis and an ipsilateral renal agenesis. The growth in size of the mass forced the authors to perform an operative exploration. RESULTS: Intraoperative findings included a multicystic mass in the rete testis of the right testicle. Testicle-sparing total removal of the multicystic mass was performed and the pathologic examination revealed a benign, multilobulated configuration of the cysts in the region of the rete testis. These findings were similar to those found in previously reported cases of CDT. Ipsilateral renal agenesis is the most common associated anomaly. As a pathogenetic factor, mal junction of the Wolffian duct in the 5th week of gestation is most creditable. CONCLUSIONS: CDT is a rare cause of pediatric scrotal mass. When feasible, a testicle-sparing approach should be considered and all patients should undergo evaluation for associated urologic anomalies. PMID- 12119431 TI - Preoperative risk factors for surgery female urethral diverticula. Our experience. AB - INTRODUCTION: This study describes the authors' experience about the most important risk factors in surgical treatment of female urethral diverticula. The attention is focused on the correlation between risk factors and common severe complications after surgical treatment. PATIENTS AND METHODS: Eighteen women underwent transvaginal diverticulectomy from 1990 to December 2000. Voiding cystourethrography and transvaginal ultrasonography were performed preoperatively. As far as location is concerned, 10 women developed a mid urethral diverticula, the remaining had diverticula equally distributed over the distal and proximal urethra. Ten diverticula showed a posterior development, 6 a lateral one and 2 diverticula had horseshoe shape. In 6 cases, diverticulum was larger than 4 cm. RESULTS: Follow-up ranged from 44 to 121 months. We recorded one urethrovaginal fistula, two genuine stress incontinence of new onset, associated in one case with recurrent diverticulum, and one recurrent diverticulum as major complications. CONCLUSIONS: The most important risk factors, evaluated with a statistical analysis, are represented by delayed diagnosis, size over 4 cm, and lateral or horseshoe shape of the diverticulum. PMID- 12119432 TI - Vestibular flap urethroplasty for strictures of the female urethra. Impact on symptoms and flow patterns. AB - INTRODUCTION: To assess the impact on subjective symptoms and flow patterns of a new surgical technique designed to correct strictures of the female distal urethra and urethral meatus. MATERIALS AND METHODS: Seventeen patients (mean age 41.2 years) with symptomatic strictures of either the distal urethra or the urethral meatus entered the study. Patients reporting an AUA score >20, a diagnosis of bladder outlet obstruction according to the Abrams-Griffiths nomogram and the Schaefer linPURR diagram, urethral calibration <20 F and radiologic evidence of the stricture, were considered eligible for surgery. A pedicled flap isolated from the vaginal vestibule was anastomosed with two longitudinal running sutures along the two edges of an opened urethra. RESULTS: In all cases, diffuse fibrosis of the urethral wall was demonstrated at histological examination. Mean (+/- SE) preoperative and 12-month follow-up results were as follows: AUA score 25.2 +/- 2.1 vs. 8.4 +/- 1.2 (p < 0.0001); peak flow rate (ml/s) 13.2 +/- 1.2 vs. 36 +/- 1.5 (p < 0.0001); detrusor pressure at Q(max) (cm H(2)O) 45 +/- 5 vs. 17 +/- 3; residual urine volume (ml) 120 +/- 5 vs. 20 +/- 5 (p < 0.0001). Fifteen patients (88%) showed an unobstructed Abrams Griffiths nomogram and a Schaefer linPURR diagram postoperatively. All but 2 cases (88%) could be calibrated at 28 F postoperatively and showed a normal urethral lumen at voiding cystourethrography. Complications were never noted. CONCLUSIONS: Female patients with symptomatic strictures of the distal urethra or urethral meatus may be treated efficaciously and safely with vestibular flap urethroplasty. Although this technique must be performed under optical magnification it is easy to perform and is not associated with complications. PMID- 12119433 TI - An innovative mobile lithotripter for extracorporeal shock wave lithotripsy and therapy. AB - INTRODUCTION: To assess the performance of a new mobile device for extracorporeal shock wave lithotripsy (ESWL) and therapy (ESWT). MATERIALS AND METHODS: 278 patients underwent 399 treatment for stone disease. 28 patients received 64 treatments for orthopaedic issues such as pseudarthrosis or enthesiopathies resistant to conservative treatment. RESULTS: During ESWL, minor pain symptoms were well resolved with analgesics intravenously. 45% of patients were stone-free and 50% had irrelevant fragments at the time of discharge. 66% of patients underwent a single treatment. Auxiliary measures after ESWL were necessary in 5%. After ESWT, pain symptoms of all patients were reduced on average to 5.2 points on a numerical rating scale from 1 to 10. Patients with bony nonunions produced callus if the bone scan before ESWT showed activity. No complication related to either form of treatment was observed. CONCLUSIONS: This innovative mobile lithotripter fulfils all expectations a user can have in an upt-to-date equipment: good disintegration, low side effects, easy handling, fast installation, dual imaging and suitability for ESWL and ESWT. PMID- 12119434 TI - Evaluation and therapeutic regulation of erectile dysfunction with visual stimulation test. An objective approach by using sildenafil citrate test. AB - AIM: An objective evaluation of the psychogenic cause of erectile dysfunction by performing the visual stimulation tumescence and rigidity (VSTR) test and sildenafil citrate test, together with the effectiveness of sildenafil citrate medication on impotence caused by different etiologies. MATERIAL AND METHODS: Between 1998 and 2000, a total of 36 men (12 patients with diabetic etiology, 5 patients with vasculogenic risk factor) were enrolled in this study. The mean age of patients was 53 (27-67) years. Following standard questionnaires, including a detailed anamnesis from an andrologic viewpoint, VST was performed in an ambulatory setting and beginning with a test dose of 50 mg. At the end of 2 h, the data was evaluated with computer assistance (Rigiscan device) and if a satisfactory erection had not occurred, an additional second dose of sildenafil citrate (50 mg) was given until there was a satisfactory erection. Results obtained from VST: results were classified as group I (fully rigidity, >10 min erection, >70% of rigidity, possible vaginal penetration), group II (unstable erection, 5 min erection, >70% of rigidity, possible vaginal penetration) and group III (tumescence without rigidity, <5 min erection, <70% of rigidity, impossible vaginal penetration). The results obtained during the first 1 h of the VSTR test were regarded as the patient's own erectile condition and later data was accepted as the real effect of sildenafil citrate. The Fisher exact test was used for statistical evaluation including pre- and post-sildenafil effect on erectile rigidity and duration of erection. RESULTS: The erection status of patients was sufficient in 17 (47.2%) in group I, it was insufficient but sufficient enough with an increased dose of sildenafil citrate in 10 (27.7%) in group II, and insufficient without/with full dose of sildenafil citrate in 9 (25%) in group III. Considering rigidity and total erectile period, there was a statistical significant difference between the first two groups with respect to the early and late sildenafil citrate effects on the VSTR test (p < 0.05). Again, 10 patients with known risk factors (diabetes mellitus 5 and vasculogenic 5) in the second group seemed to give a good response to repeated dosage of sildenafil citrate which has been found to be very interesting. However, the rest of the diabetic patients (n = 7) in the third group showed no erection despite the increasing and repeated doses of sildenafil citrate. CONCLUSION: Sildenafil citrate with the VSTR test has effective and reliable results which was regarded as very important to diagnose and determine objectively the amount of therapeutic doses in impotence. In accordance with the literature data, our results also confirm the reliability and the practical nature of the VSTR test, which is less time-consuming and cheaper than the nocturnal penile tumescence and rigidity (NPTR) test. In the VSTR test, necessary doses of medication needed for satisfactory erection were easily regulated in patients with certain kinds of impotence. Additionally, self-criticism advantage of the patients on erection and an unnecessary need for regular sexual partners may make this test preferable in the near future. However, we believe that a large group of patients with other definite parameters are certainly needed in order to obtain more reliable data. PMID- 12119435 TI - Application of the microwave tissue coagulator: is it beneficial to partial nephrectomy? AB - BACKGROUND/AIM: The indications of partial nephrectomy have expanded after the introduction of new techniques for preventing excessive blood loss and avoiding deterioration of the renal function after clamping the renal pedicle. We present our clinical experience of partial nephrectomy for renal tumors using a microwave tissue coagulator. PATIENTS AND METHODS: Between April 1996 and January 2000, 34 patients underwent open partial nephrectomies in the Kobe City General Hospital. The microwave tissue coagulator was used for resection of the renal parenchyma, but in deeper lesions a sharp dissection was performed. Twenty-two patients (groups 1 and 2) underwent partial nephrectomy without vascular control (14 renal pedicles were not disturbed in group 1 patients, and 8 renal pedicles were dissected but not clamped in group 2 patients). Another 12 patients (group 3) underwent vascular control with ligation of the tumor-feeding segmental arteries before parenchymal resection. The patients of group 1 underwent wedge resections, while those of groups 2 and 3 underwent segmental or transverse partial nephrectomies. RESULTS: Complete tumor resection was done in all 34 patients. In group 1, the microwave tissue coagulator was very effective to control the blood loss (mean 330 ml). In larger resections, this method only was inadequate to control the blood loss (mean 489 ml in group 2), so that we needed vascular control. However, despite vascular control, mean blood losses of about 943 ml because of deeper venous bleeding occurred in group 3, and, moreover, postoperative renal infarctions occurred in 2 patients. Other complications were urinary fistula formation in 16 patients (47%) and renal pelvic stenoses in 2 patients (5.8%). All of the urinary fistulas were easily repaired by simple suturing intraoperatively. CONCLUSIONS: Especially in wedge resection, the microwave tissue coagulator achieved safe resection without vascular control which differs from other new techniques. However, in larger resections, a combination with other techniques may be necessary to decrease blood loss and the rate of complications. PMID- 12119436 TI - Brucellar orchitis in Innerwest Anatolia Region of Turkey. A report of 12 cases. AB - Brucellosis, which affects the genitourinary system in rate of 2-20%, is a multiorgan infectious disease. Twelve patients were diagnosed as having brucellar orchitis serologically, clinically and ultrasonographically. In 2 patients, Brucella melitensis was isolated in blood cultures. All the patients were working in cattle dealing. They were treated with 600 mg/day rifampicin plus 200 mg/day doxycycline for 6 weeks and followed up during 1 year. They recovered clinically within 3 weeks. Although they did not have any symptoms or findings, in 4 patients, serological titers did not return to normal after 6 weeks. In 2 of these patients, relapse was seen in the 6th and 8th months, respectively. These 2 patients recovered with 1 g/day ciprofloxacin plus 2 g/day tetracycline for 6 weeks. Relapse did not occur again. Conclusively, brucellosis must be considered as a cause of orchitis in especially endemic regions where cattle dealing is widespread. The patients must be followed for relapse during at least 1 year. PMID- 12119437 TI - Effects of vaginal trauma and oophorectomy on the continence mechanism in rats. AB - PURPOSE: To establish an animal model for studying the effects of vaginal trauma and oophorectomy on the continence mechanism in rats. METHODS: Ninety-six female rats were used in the experiments. The rats were divided into 8 groups that received either no treatment (control), or single vaginal trauma at 0 day and 4 weeks, multiple vaginal traumas, oophorectomy at 4 and 12 weeks, and combined oophorectomy and single vaginal trauma or multiple vaginal traumas at 4 weeks. In vivo experiments were performed to determine abdominal leak point pressure (ALPP) by recording the intravesical pressure obtained during compression of the lower abdomen. In vitro urethral contractility experiments were then performed using isolated urethra and electrical field stimulation, acetylcholine, and norepinephrine. Finally, histological study of the urethral muscles and paraurethral structures was performed. RESULTS: Single or multiple vaginal traumas resulted in a significant reduction of ALPP. The reduced ALPP recovered at 4 weeks after single vaginal trauma. Oophorectomy did not significantly affect ALPP compared to controls; however, when oophorectomy was combined with multiple vaginal traumas, a significant reduction in ALPP occurred. Urethral contractility was reduced after multiple vaginal traumas but was not significantly different from the control after oophorectomy. Histological studies revealed disruption of the ventral part of striated muscles after single or multiple vaginal traumas. Degenerative and hyalinization changes were noted in submucosal and muscle layers after oophorectomy combined with multiple vaginal traumas. CONCLUSIONS: Vaginal trauma can injure the urethral muscles and nerves. Single or multiple vaginal traumas can induce denervation of periurethral muscles and reduce ALPP. With a period of recovery, the urethral resistance increases and ALPP returns. Oophorectomy mainly affected the intrinsic urethral closure mechanism and resulted in a nonsignificantly reduced ALPP; however, a significant reduction of ALPP developed when oophorectomy was combined with multiple vaginal traumas. PMID- 12119438 TI - Smooth muscle within incomplete obliterations of processus vaginalis lacks apoptotic nuclei. AB - INTRODUCTION: Incomplete obliteration of processus vaginalis (PV) has been suggested to result from the persistence of smooth muscle, which should normally disappear after taking part in the descent of testis. Since apoptosis is the mechanism of disappearance, the presence or absence of apoptotic nuclei was evaluated within sacs that result from failed obliteration of PV. MATERIALS AND METHODS: Twenty sacs associated with female inguinal hernia (n = 5), male inguinal hernia (n = 6), hydrocele (n = 5), hydrocele of the cord (n = 2), and undescended testis (n = 2) were evaluated. 10-microm sections were cut from the snap-frozen samples and stained for nuclear DNA fragmentation. RESULTS: Apoptotic nuclei were detected within the vascular structures and mesothelium. However, none of the samples from different diagnostic sources have revealed any apoptotic nucleus within the smooth muscle component. CONCLUSIONS: While the vascular and mesothelial structures within the sacs reveal evidence of apoptosis, the smooth muscle component lacks apoptotic process. The failed apoptosis of smooth muscle may have a role in the persistence of PV. PMID- 12119439 TI - The expression of oncoproteins in transitional cell carcinoma: its correlation with pathological behavior, cell cycle and drug resistance. AB - OBJECTIVE: To investigate the relationship of oncoproteins with histological grade, tumor stage, cell cycle and multidrug resistance (MDR) in bladder cancer. METHODS: The expression of various oncoproteins (p21, EGFR1, erbB-2, c-jun, c myc, bcl-2, Bax) and suppressor gene products (WT-1, RB gene, p53) was studied in normal urothelium, transitional cell carcinoma (TCC) cell lines and their MDR sublines by using specific antibodies and an indirect immunofluorescence flow cytometric method. RESULTS: The total increased rate of measured oncoproteins in TCC cell lines was 62.7%. There was no difference in the expression rate of oncoproteins between low-grade/stage TCC cell lines and high-grade/stage TCC cell lines. All of these TCC cell lines have aneuploidy and increased proliferative activity in the cell cycle, but a correlation with oncoprotein expression was not seen. They had a low expression rate of c-myc, Bax and RB gene when compared to normal urothelium. On the contrary, the expressions of p21, EGFR1, erbB-2, bcl-2, WT-1 and p53 oncoproteins were significantly higher than in normal urothelium (p < 0.05). A long-term cultured MDR subline of TCC8702 (TCC8702/ADR1000) demonstrated a generally decreasing expression of oncoproteins in addition to p53. CONCLUSIONS: The p21, EGFR1, erbB-2, bcl-2, WT-1 and p53 oncoproteins were increased in TCC cells compared to normal urothelium. Oncoprotein expression is related to tumorigenesis of TCC cells, but no correlation was seen between the incidence and tumor differentiation and cell cycle. These oncoproteins decreased gradually in the process of generation of MDR in addition to p53. It indicates that the p53 oncogene is closely related to the acquired MDR of TCC cells. PMID- 12119440 TI - Effect of tumor-infiltrating lymphocyte subsets on prognosis and susceptibility to interferon therapy in patients with renal cell carcinoma. AB - INTRODUCTION: Immunotherapy effectively treats advanced renal cell carcinoma in only a limited number of patients. However, the predicted prognosis for each patient in relation to immune status and response to immunotherapy remains problematic. We analyzed tumor-infiltrating lymphocyte (TIL) subsets to determine whether these correlated with the prognoses for the patients and the response to alpha-interferon therapy. MATERIALS AND METHODS: TIL subsets from resected specimens of 79 patients were analyzed by two-color flow cytometry and then compared with the patients' long-term clinical courses and responses to interferon therapy. RESULTS: In patients with stages III and IV, an increased infiltration of CD4+ cells and decreased CD8+ cells constituted a fair prognostic factor. In 17 patients with metastatic lesions, 8 of 10 patients who had disease progression after interferon therapy showed an increase in CD8+ cells above 25%, whereas 2 responders and 5 patients who had stable disease showed infiltration of CD8+ cells below 25%. CONCLUSIONS: The TIL subset is a prognostic factor for advanced renal cell carcinoma, and its analysis provides a method to predict the susceptibility to interferon therapy. PMID- 12119441 TI - Immunohistochemical study of bcl-2, p53 and Ki-67 expressions in collecting duct carcinoma of the kidney. Five cases and review of the literature on bcl-2, p53 and Ki-67 expressions in renal cell carcinoma and transitional cell carcinoma. AB - OBJECTIVES: This study was designed to examine the expressions of bcl-2, p53 and Ki-67 antigen in collecting duct carcinoma (CDC) of the kidney by an immunohistochemical method. METHODS: The diagnosis of CDC was based on the criteria proposed by Srigley and Eble. The clinical courses of 5 CDC cases examined in this study suggested that 3 cases were low grade and 2 cases high grade. The expressions of bcl-2, p53 and Ki-67 antigen were evaluated in paraffin embedded surgical specimens using anti-bcl-2 and anti-p53 antibodies and Ki-67 antigen, respectively. RESULTS: The expression of bcl-2 was recognized in 3 of 5 cases (60%), p53 expression also in 3 of 5 cases (60%). The Ki-67 labeling index was 7.46 +/- 7.40 (mean +/- SD). CONCLUSIONS: It was suggested that there were two clinical types in CDC; the expression of bcl-2 did not correlate with the CDC patients' clinical courses and the cellular proliferation, and p53 expression was recognized in the CDC patients with highly cellular proliferation. PMID- 12119442 TI - Immunohistochemical studies of caveolin-3 in germ cell tumors of the testis. AB - BACKGROUND/AIM: Caveolin, which is a major constructive component of the caveolar membranes, plays a key role in transcytosis of molecules into cells and regulation of several signal transductions. Caveolin has three isoforms, and recent studies suggest that in some malignant tumors an alteration of caveolin-1 expression correlates with oncogenetic changes. Caveolins have been reported to be negative regulators of inducible nitric oxide synthase (iNOS) which can provoke an antitumor response via infiltrating immune cells. The aim of this study was to examine the expressions of caveolin-1 and caveolin-3 (which is a caveolin-1 homologue localized predominantly in muscle tissue) in testicular cancer. METHODS: We evaluated the expressions of caveolin-1, caveolin-3, and iNOS in 16 seminoma and 10 non-seminoma testicular cancer specimens as well as normal testicular tissue, using a streptavidin-biotin bridge technique on cryostat sections. The expression of caveolin-3 was confirmed by slot-blot analysis. Tumor infiltrating immune cells were also studied immunohistochemically, and the correlation between the number of immune cells and caveolin-3 concentrations was calculated. RESULTS: Immunohistochemistry revealed that caveolin-3, but not caveolin-1, was frequently expressed in seminomas (12 positive out of 16 specimens) without positive staining in their normal counterparts or in nonseminomatous germ cell tumors, except for muscle components in the teratoma. iNOS was not expressed in any tissues examined. Samples with high levels of caveolin-3 tended to have higher degrees of tumor-infiltrating immune cells, although this finding was not statistically significant. CONCLUSION: This is the first report demonstrating the involvement of caveolin-3 in germ cell tumors. PMID- 12119443 TI - A rare retroperitoneal benign tumor. Castleman's disease. AB - A case of retroperitoneal Castleman's disease - a benign lymphoid tumor - is reported. The tumor is excised totally. A 34-month follow-up period is disease free and uneventful. The preoperative workup presented, its clinical behavior and management are discussed. PMID- 12119445 TI - Retroperitoneal schwannoma extending into the intravertebral foramen. AB - A 35-year-old Japanese man first visited the hospital for episodes of numbness in the right medial femoral region and loss of muscular strength in the right thigh. A CT scan of the abdomen showed a 10 x 9-cm well-circumscribed in homogeneous multilayered mass, in contact with the lumbar vertebrae. MRI demonstrated a hypodense and hyperdense mixed mass. Tumor extension into the L(2)/L(3) intravertebral foramen was shown. To avoid spinal cord damage, we chose enucleation of the tumor in the intravertebral foramen. A follow-up CT after 1 year showed no evidence of a recurrent mass. In conclusion, we advocate the nerve sparing operation in cases like that of our patient. PMID- 12119444 TI - Rupture of a renal artery aneurysm in pregnancy. AB - Rupture of a renal artery aneurysm is a well-recognized phenomenon. The rupture usually occurs in late pregnancy. We report a case in whom this occurred in the first trimester of pregnancy. PMID- 12119446 TI - Variants of renal angiomyolipoma closely simulating renal cell carcinoma: difficulties in the histological diagnosis. AB - Renal angiomyolipoma is considered to be a benign renal tumor composed of atypical blood vessels, smooth muscles and fat cells. We report 2 cases of unilateral renal angiomyolipoma. In both cases, our preoperative diagnosis was renal cell carcinoma, because no low density area compatible with fat tissue was noted in the tumors on radiographic evaluation. Through histological examination, both tumors proved to be angiomyolipomas mainly composed of epithelioid cells in 1 case, and spindle-shaped smooth muscle cells mimicking a leiomyoma in the other case. Both patients are well showing no evidence of metastases 16 and 14 months after nephrectomy, respectively. PMID- 12119447 TI - A case of T1C, N3, M1 prostate cancer. PMID- 12119448 TI - Central antinociceptive activity of acetylsalicylic acid is modulated by brain serotonin receptor subtypes. AB - Male Wistar rats were treated with ondansetron (1 and 2 mg/kg s.c.), ketanserin (0.2, 1 and 5 mg/kg s.c.) or NAN-190 (1, 3 and 5 mg/kg i.p.) 15 min before acetylsalicylic acid (ASA, 400 mg/kg i.p.), and 30 min thereafter the pain threshold was evaluated. The antinociceptive activity of ASA in the hot-plate test was variously affected by ondansetron, ketanserin and NAN-190: at the highest dose (2 mg/kg s.c.) ondansetron abolished it while ketanserin (5 mg/kg s.c.) significantly reduced it, and NAN-190 (1-5 mg/kg) did not significantly modify the effect of ASA. Binding experiments indicate that both ondansetron and ketanserin completely prevented the decrease in the maximum number of 5-HT(2) receptors (B(max)) provoked by ASA. These data indicate that the central antinociceptive activity of ASA is modulated in a different manner by serotonin receptor antagonists, and that 5-HT(2) and 5-HT(3) receptors may exert a pivotal role in nociception, alone or in association. PMID- 12119449 TI - Neurotransmitter mechanisms in gabapentin antinociception. AB - The present study was done to investigate the neurotransmitter mechanisms involved in gabapentin antinociception in healthy albino rats. The formalin test was used to asses antinociception. Gabapentin (10-120 mg/kg s.c.) decreased the pain score in the formalin test. In order to study the putative neurotransmitter mechanisms involved in gabapentin action, the effect of gabapentin (30 mg/kg s.c.) alone and in rats pretreated with different receptor blockers, e.g. bicuculine, saclofen, naloxone, mecamylamine, atropine, DL-p-chlorophenylalanine methyl ester hydrochloride, glibenclamide, theophylline, and L-arginine was assessed. Gabapentin decreased the pain score, and the ED(50) of gabapentin was 36.8 +/- 8.2 (30.2-43.1) mg/kg s.c. Pretreatment with different receptor blockers did not modify gabapentin (30 mg/kg s.c.) antinociception except for L-arginine which increased the pain score from 1.68 +/- 0.29 (gabapentin) to 2.29 +/- 0.41. Results suggest the involvement of L-arginine nitric oxide pathways in gabapentin antinociception. PMID- 12119451 TI - LiCl attenuates M(1)AChR-mediated intrathecal pilocarpine-induced reciprocal hindlimb scratching in mice. AB - Pretreatment of mice with a subcutaneous (s.c.) injection of LiCl (1, 3, 10 mmol/kg) 18 h prior to testing produced dose-related attenuation of the reciprocal hindlimb scratching (RHS) response induced by intrathecal (i.t.) administration of pilocarpine. LiCl (10 mmol/kg, 424 mg/kg) pretreatment shifted the pilocarpine (2 microg)-induced RHS dose-response curve about 5-fold to the right compared to vehicle-injected controls (ED(50) = 2.97 and 0.56 microg, respectively). Coadministration of inositol-1,4,5-trisphosphate (IP(3); 5, 10, 20 microg), but not inositol hexaphosphate (0.03-3 microg), myo-inositol (10 microg) or EDTA (1.33 microg), produced dose-related reversal of the LiCl effect. IP(3) administration alone neither produced RHS nor enhanced the pilocarpine-induced RHS in animals not pretreated with LiCl. These findings provide in vivo evidence for a possible link between RHS and a LiCl-sensitive, possibly phosphoinositide related, effector pathway. PMID- 12119450 TI - Inhibition of muscimol on morphine-induced hyperactivity, reverse tolerance and postsynaptic dopamine receptor supersensitivity. AB - This study was performed to investigate the effect of muscimol on morphine induced hyperactivity, reverse tolerance and postsynaptic dopamine receptor supersensitivity in mice. A single administration of morphine induced hyperactivity as measured in mice, and the morphine-induced hyperactivity was inhibited dose-dependently by the administration of the GABA(A) agonist, muscimol (0.3, 0.5 and 1.0 mg kg(-1) i.p.). However, daily repeated administration of morphine caused the development of reverse tolerance against morphine hyperactivity (10 mg kg(-1) s.c.). The administration of muscimol inhibited the development of reverse tolerance against morphine hyperactivity (10 mg kg(-1) s.c.) in mice that had received chronic administration of morphine. Postsynaptic dopamine receptor supersensitivity, as shown by the enhanced ambulatory activity after administration of apomorphine (2 mg kg(-1) s.c.), also developed in reverse tolerant mice. Muscimol also inhibited the development of postsynaptic dopamine receptor supersensitivity induced by the chronic administration of morphine. These results suggest that the hyperactivity, reverse tolerance and postsynaptic dopamine receptor supersensitivity induced by morphine can be inhibited via the activation of GABA(A) receptors. PMID- 12119452 TI - Deoxycholic acid conjugates are muscarinic cholinergic receptor antagonists. AB - In the course of examining the actions of major human bile acids on cholinergic receptors, we discovered that conjugates of lithocholic acid are partial muscarinic agonists. In the present communication, we report that conjugates of deoxycholic acid (DC) act as cholinergic muscarinic receptor antagonists. Chinese hamster ovary (CHO) cells expressing rat M3-muscarinic receptors were used to test bile acids for inhibition of radioligand [N- (3)H-methylscopolamine ((3)H NMS)] binding; alteration of inositol phosphate (IP) formation; mitogen-activated protein (MAP) kinase phosphorylation and cell toxicity. We observed approximately 18.8, 30.3 and 37.1% inhibition of (3)H-NMS binding with DC and its glycine (DCG) and taurine (DCT) conjugates, respectively (all 100 micromol/l, p < 0.01). DCT and DCG inhibited acetylcholine-induced increases in IP formation and MAP kinase phosphorylation (p44 and p42 ERK). DCG and DCT did not alter trypan blue exclusion or lactate dehydrogenase release from CHO-M3 cells. We observed the following rank order of potency (IC(50) micromol/l) for inhibition of (3)H-NMS by muscarinic antagonists and bile acids: NMS (0.0004) > 4-DAMP (0.009) > atropine (0.012) > DCT (170) > DCG (250). None of the bile acids tested were hydrolyzed by recombinant cholinesterase. At concentrations achieved in human bile, DC derivatives are natural muscarinic antagonists. PMID- 12119453 TI - Adjuvant and salvage radiotherapy after radical prostatectomy. AB - BACKGROUND: Following radical prostatectomy, between 15 and 60% of all patients with pT3 prostate cancer experience persistence or increasing levels of prostate specific antigen (PSA) as a sign of tumor persistence or progression within 5 years. Retrospective studies have shown a rate of 35-55% of positive biopsies from the vesicourethral anastomosis in this situation. Best treatment for these disease conditions is under debate, current strategies include adjuvant radiotherapy (RT), 'wait-and-see' and salvage RT or hormone therapy for increasing PSA. RESULTS: A number of retrospective studies have shown an increased rate of local control and 'freedom from treatment failure' following adjuvant RT with doses in the range of 50-60 Gy. However, no survival benefit could be demonstrated by now. Results of three major phase III studies are pending. In case of persisting or increasing PSA levels following radical prostatectomy, 30-70% of these patients will reach an undetectable PSA level after conformal RT with total doses of 60-70 Gy, which will stay undetectable or at least stable within the next 2-5 years in about 50% and therefore offering a chance of cure. When starting RT, PSA should be as low as possible (<2 ng/ml). With higher PSA levels the chance of achieving an undetectable PSA again decreases below 35%. High Gleason scores of 8-10, seminal vesicle involvement and a short PSA doubling time are adverse prognostic factors. Severe late side effects of conformal RT are infrequent (<3%). In contrast, hormonal treatment is of palliative nature in the long run, with a median time to development of metastases of 4-7 years, and can be offered to patients with progressive disease after RT. CONCLUSIONS: Adjuvant RT following radical prostatectomy for pT3 prostate cancer offers higher local control rates and an increase in 'freedom from treatment failure', but no prolongation of survival has yet been shown. In the situation of increasing PSA levels after radical prostatectomy, salvage RT seems to offer a chance of cure in selected patients, although it is difficult to draw firm conclusions because of generally too short follow-up times. PMID- 12119454 TI - Prognostic factors in squamous cell carcinoma of the head and neck. AB - Therapy for squamous cell carcinoma of head and neck relies on surgery, radiotherapy and chemotherapy, mostly a combination thereof. In patients treated with curative intent, the intensity of therapy is adapted to the supposed prognosis and should be defined upon prognostic factors. Besides classical prognostic parameters, T, N and M stage, the presence of extranodal growth (extracapsular spread, ECS), tumor volume, lymph node burden, extent of tumor necrosis, histologic grading, but also type of treatment were determined in consideration of prognosis. The p53 status does not correlate with prognosis in most investigations. The tumor hypoxia seems to be of prognostic value, and strategies to overcome the adverse effect are currently investigated. Not all factors are relevant for all types of treatment. Besides ECS, these new factors so far have rarely been used to stratify prospective combined modality treatment according to the risk of locoregional and distant failure. PMID- 12119455 TI - The value of radiation therapy in the management of aggressive non-Hodgkin's lymphomas. AB - Even though chemotherapy plays a major role in the treatment of aggressive non Hodgkin's lymphomas, the high radiosensitivity of the lymphatic lesions has established radiotherapy as an important component in the management of the disease in localized stages. Nevertheless, the treatment strategies with regard to the stage-adopted indications for radiotherapy, the treatment volume, and the dose remain controversial. This article reviews the available data concerning combined-modality treatment in localized stages and treatment of bulky lesions and residual lymphomas after chemotherapy with emphasis on the role of radiation therapy. PMID- 12119456 TI - New aspects of adjuvant radiotherapy in primarily resectable rectal cancer. AB - Local recurrence rates of approximately 30% have been consistently reported after 'conventional surgery' in stage II + III rectal cancer patients. Postoperative radiotherapy decreases the local recurrence rate only moderately and is not associated with a gain in survival. Preoperative radiotherapy with biologically effective doses > 30 Gy dramatically reduces the relative risk of local recurrences by 57% that translates into significant overall survival benefit. Postoperative 5-fluorouracil-based chemotherapy in combination with radiotherapy has also been shown to increase local tumor control and to result in a significantly better survival. However, an improved surgical technique, the total mesorectal excision (TME), has evolved and is considered standard treatment by many surgeons. With this technique, reduced local recurrence rates of below 10% and improved survival rates have been reported from different institutions. Therefore, a discussion has started, whether or not pre- or postoperative radiotherapy and chemotherapy is beneficial, when TME is used. The preliminary results of a recently published trial on radiotherapy before TME partly resolve the current controversy. An expected local failure rate of approximately 15% and an overall survival rate below 60%, 5 years after TME alone in stage II + III rectal cancer patients suggest the need for additional therapy. Preoperative radiotherapy reduced the risk of local recurrence more than 3 times, indicating that the use of radiotherapy is justified at least in patients with stage II + III disease, even if optimal surgery was used. The unsatisfactory overall survival gives indirect evidence that these patients will also benefit from chemotherapy. However, many questions remain open and an optimal therapeutic schedule has not been established. PMID- 12119457 TI - Intensity-modulated radiotherapy - technology and clinical applications. AB - Intensity-modulated radiotherapy (IMRT) is one of the most important developments in radiooncology of the last years. As an extension of 3D conformal radiotherapy, it provides the possibility of delivering a high radiation dose to the tumor tissue, protecting radiosensible critical organs nearby or even surrounded by the target. This is realized by the overlaying of beams from different directions, which are not homogeneous like in conventional radiotherapy, but inhomogeneous themselves. The sum of the beams from all directions forms finally the desired complex-shaped dose distribution. Because choosing the right intensity modulation of each beam is a nontrivial problem, the best treatment plan could only hardly be found in a trial-and-error process, but has to be computer- optimized. Therefore, IMRT has always to be spoken together with the term of 'inverse planning'. This describes the approach of telling the computerized planning system the desired radiation doses to the target and the allowed maximum doses to organs at risk (OAR), then letting the system compute the optimal modulated beams, called 'fluence profiles', by an iterative algorithm. Because the ideal dose distribution, giving 100% dose to the tumor and 0% dose to the healthy tissue, is never realizable, inverse planning has to be understood as an optimization problem. The present article shows the technical bases of IMRT and inverse planning and then describes important clinical applications. PMID- 12119458 TI - Molecular mechanisms in signal transduction: new targets for the therapy of gynecologic malignancies. AB - Growth factor-mediated signal transduction pathways in gynecologic malignancies are summarized. Targeting critical tyrosine kinase pathways may lead to more specific anticancer regimens in gynecologic oncology. During the past 10 years significant progress in cancer treatment has been made through the discovery of potent specific and well-tolerated inhibitors of signal transduction. Improved understanding of molecules involved in signal transduction pathways will undoubtedly result in an increasing number of compounds under clinical investigation. Some of the described molecular therapeutic approaches are suitable to enrich the conventional chemotherapy. PMID- 12119459 TI - Effective salvage therapy with carboplatin/mitomycin C in metastatic breast cancer. AB - BACKGROUND: Alternative and effective drug regimens in patients with metastatic breast cancer progressing after adriamycin- and taxoid-containing regimens are urgently needed. PATIENTS AND METHODS: In a phase II trial, 43 heavily pretreated patients with metastatic breast cancer were treated with both carboplatin 200 mg/m(2) i.v. and mitomycin C 10 mg/m(2) i.v. on day 1 every 4 weeks. In case of granulocytopenia or thrombocytopenia below grade 3 according to NCI-CTC, the carboplatin dosage was escalated to 300, 400, and 450 mg/m(2) in the next treatment cycle. RESULTS: During the first 3 cycles the dose intensity of carboplatin could be increased from a mean of 50 to 74 mg/m(2)/week. Beyond this value the carboplatin dose intensity decreased because of hematotoxicity. Based on an intention-to-treat analysis, 9 of 43 patients responded to therapy (21%; 95% CI = 10.04-36.04) including 2 complete and 7 partial responses. 15 patients had no change, 13 progressed, and 6 patients were considered nonevaluable. The median time to progression was 3 (range 0-12) months. NCI-CTC grade 3 or 4 granulocytopenia was observed in 14 patients, grade 3 or 4 thrombocytopenia occurred in 32, grade 3 infections in 3, grade 3 hemorrhage in 1, and grade 3 cardiac dysrhythmias in 1 of the patients. CONCLUSIONS: In anthracycline/ taxoid pretreated patients, salvage treatment with a combination of carboplatin and mitomycin C seems to be effective and associated with foreseeable toxicity. Based on our results with an intraindividual dose escalation of carboplatin, a dosage of 300 mg/m(2) in combination with mitomycin C 10 mg/m(2) every 4 weeks seems to represent a recommendable starting dose for future studies. PMID- 12119460 TI - High-dose 5-fluorouracil / folinic acid in combination with three-weekly mitomycin C in the treatment of advanced gastric cancer. A phase II study. AB - BACKGROUND: The 24-hour continuous infusion of 5-fluorouracil (5-FU) and folinic acid (FA) as part of several new multidrug chemotherapy regimens in advanced gastric cancer (AGC) has shown to be effective, with low toxicity. In a previous phase II study with 3-weekly bolus 5-FU, FA and mitomycin C (MMC) we found a low toxicity rate and response rates comparable to those of regimens such as ELF, FAM or FAMTX, and a promising median overall survival. In order to improve this MMC dependent schedule we initiated a phase II study with high-dose 5-FU/FA and 3 weekly bolus MMC. PATIENTS AND METHODS: From February, 1998 to September, 2000 we recruited 33 patients with AGC to receive weekly 24-hour 5-FU 2,600 mg/m(2) preceded by 2-hour FA 500 mg/m(2) for 6 weeks, followed by a 2-week rest period. Bolus MMC 10 mg/m(2) was added in 3-weekly intervals. Treatment given on an outpatient basis, using portable pump systems, was repeated on day 57. Patients' characteristics were: male/female ratio 20/13; median age 57 (27-75) years; median WHO status 1 (0-2). 18 patients had a primary AGC, and 15 showed a relapsed AGC. Median follow-up was 11.8 months (range of those surviving: 2.7 11.8 months). RESULTS: 32 patients were evaluable for response - complete remission 9.1% (n = 3), partial remission 45.5% (n = 15), no change 27.3% (n = 9), progressive disease 15.1% (n = 5). Median overall survival time was 10.2 months [95% confidence interval (CI): 8.7-11.6], and median progression-free survival time was 7.6 months (95% CI: 4.4-10.9). The worst toxicities (%) observed were (CTC-NCI 1/2/3): leukopenia 45.5/18.2/6.1, thrombocytopenia 33.3/9.1/6.1, vomitus 24.2/9.1/0, diarrhea 36.4/6.1/3.0, stomatitis 18.2/9.1/0, hand-foot syndrome 12.1/0/0. Two patients developed hemolytic-uremic syndrome (HUS). CONCLUSIONS: High-dose 5-FU/FA/MMC is an effective and well-tolerated outpatient regimen for AGC (objective response rate 54.6%). It may serve as an alternative to cisplatin-containing regimens; however, it has to be considered that possibly HUS may occur. PMID- 12119461 TI - German national case collection of familial pancreatic cancer - clinical-genetic analysis of the first 21 families. AB - BACKGROUND: The observation of a familial accumulation of ductal pancreatic adenocarcinoma (PC) and the increased risk for PC in certain hereditary tumor syndromes point to a genetic predisposition for PC. In order to evaluate the characteristics of familial PC, a German national case collection for familial pancreas cancer (FaPaCa) was established. PATIENTS AND METHODS: In FaPaCa, families of patients with PC are being collected, who have at least 1 first degree relative with PC or with malignant melanoma. Histopathologic verification of tumor diagnoses, acquisition of clinical data, and full genetic counselling are prerequisites for the enrollment of PC families in FaPaCa. RESULTS: So far, 21 families fulfilled the criteria for partaking in FaPaCa. In 11 families, PC represented the sole tumor entity. Additional tumors included malignant melanoma in 5, breast cancer in 3, and prostatic, colon or lung cancer in 2 families. Compared to the preceding generation, a younger age at diagnosis of PC was observed in the offspring of PC patients (offspring median 53 years vs. parents median 75.5 years). CONCLUSION: The association of PC and breast cancer, and of PC and malignant melanoma suggests predisposing mutations in the BRCA2 or CDKN2A genes in about one third of the FaPaCa families. Mutational analyses in both candidate genes may help to identify individuals who are at an increased risk for developing PC. A shift towards a younger age at diagnosis in our PC families may indicate genetic anticipation and/or changes of patterns of exogenous risk factors. PMID- 12119462 TI - Malignant intestinal non-Hodgkin's lymphoma from the surgical point of view. AB - BACKGROUND: Primary intestinal non-Hodgkin's lymphoma (I-NHL) is much less frequent than gastric lymphoma and has hardly been studied in prospective trails. To the surgeon, patients frequently present with abdominal emergencies. PATIENTS AND METHODS: A consecutive series of patients subjected to surgery because of I NHL between 1998 and 1999 was evaluated retrospectively for characteristic clinical, radiographic and intraoperative findings. Patients with gastric lymphoma were not considered. RESULTS: 10 patients, 8 males and 2 females, with I NHL were subjected to first-line surgery because of painful abdominal tumor, intestinal hemorrhage, obstruction or perforation. I-NHL was located most often in the small bowel (n = 7). It was rare in the colon (n = 2) and the duodenum (n = 1). Median postoperative follow-up was 28 months. Perioperative mortality was 10% (n = 1). Probability of survival 3 years after surgery was 60%. CONCLUSIONS: Patients with I-NHL frequently present with complications of tumor growth, requiring urgent surgical treatment. Irrespective of surgical complications we advocate surgery in cases of resectable disease as first-line treatment. Adjuvant treatment is indicated with respect to resection status and histopathological staging. PMID- 12119463 TI - Primary renal non-Hodgkin's lymphoma - a difficult differential diagnosis. AB - INTRODUCTION: Primary renal lymphoma (PRL) as a clinical entity is not undisputed because the kidneys do not contain lymphatic tissue and the mechanism of development of PRLs is unclear. Most of the few cases reported showed rapid systemic progression and a poor prognosis. Although there are no clearly defined diagnostic criteria for renal lymphomas, abdominal and thoracic computed tomography as well as renal and bone marrow biopsy are recommended. 3 cases of renal lymphoma are reported and their diagnosis and management discussed. CASE REPORTS: Between 1996 and 2001, 3 male patients with renal lymphoma were diagnosed and treated at our institution. In patient No. 1, because of persisting macroscopic hematuria a bilateral PRL was diagnosed by renal biopsy, without any detectable lesions on CT imaging. Patient No. 2 presented with a large renal mass which, on biopsy, was diagnosed as a lymphoma. Patient No. 3 showed lymphoma on renal biopsy and bone marrow involvement. All 3 patients were treated with systemic chemotherapy which resulted in death of disease in 2 patients and a complete remission in 1 patient after adjuvant radiotherapy and nephrectomy. CONCLUSION: PRL represents a rare entity which must nevertheless be considered in cases of unusual renal masses or otherwise unexplained renal symptoms. If diagnosed early, cure is possible, and multimodal treatment should be considered. PMID- 12119464 TI - Is there evidence-based benefit of autologous stem cell transplantation in children with solid tumors? AB - Multidisciplinary care, a better understanding of tumor biology, and advances in chemotherapy have dramatically improved the prognosis of solid tumors in children. The diseasefree survival (DFS) has increased over the last three decades from <20 to 30% in the early 1970s to now 70% after 5-10 years of follow up [1]. This progress has been limited to patients with localized tumors. However, a subset of children with advanced, therapy-resistant or relapsed disease has not derived any benefit from these advances. Despite multimodal conventional therapy, for example in newly diagnosed highrisk neuroblastomas, Ewing family tumors, metastatic rhabdomyosarcomas, or high-grade gliomas, the probability of long-term survival has remained poor, with less than 30%. After tumor progression or recurrence following adequate initial treatment, even less than 10% of patients with solid tumors survived [2]. PMID- 12119465 TI - Cadaveric kidney transplantation for the elderly. PMID- 12119466 TI - Mechanism of chloride deficit in the maintenance of metabolic alkalosis. PMID- 12119467 TI - Rufus of Ephesus and his "Diseases of the Kidneys". AB - The first book on diseases of the kidneys, titled "Diseases of the Kidneys and Bladder", was written by Ruphus of Ephesus at the close of the 1st century AD. Little is known of Rufus, who seems to have attained fame and was considered an equal of Hippocrates and Galen. He was highly respected and extensively quoted by authors of Byzantine, Arabic and Middle Ages medicine. In his description of diseases of the kidneys he makes a concerted effort to correlate structure and function, and to provide a rational explanation of the altered function of the kidneys in disease. The section of his monograph "On Hardening of the Kidneys" is brief, but constitutes the first description of morbid and clinical features of the end-stage kidneys. Like many of his contemporaries, Rufus wrote several monographs on selected topics and organs. That he chose diseases of the kidneys as the subject of one of his monographs makes him especially pertinent to the history of nephrology. PMID- 12119469 TI - Risk factor analysis for long-term tunneled dialysis catheter-related bacteremias. AB - Infection, mainly related to vascular access, is one of the main causes of morbidity and a preventable cause of death in hemodialysis patients. From January 1994 to April 1998 we conducted a prospective study to assess the incidence and risk factors of catheter-related bacteremia. One hundred and twenty-nine tunneled dual-lumen hemodialysis catheters were inserted percutaneously into the internal jugular vein in 89 patients. Bacteremia (n = 56) occurred at least once with 37 (29%) of the catheters (an incidence of 1.1/1,000 catheter-days); local infection (n = 45, 1/1,000 catheter-days) was associated with bacteremia in 18 cases. Death in 1 case was directly related to Staphylococcus aureus (SA) septic shock, and septicemia contributed to deaths in 2 additional cases. Catheters were removed in 48% of the bacteremic episodes. Treatment comprised intravenous double antimicrobial therapy for 15-20 days. Bacteriological data of bacteremia showed 55% involvement of SA. Nasal carriage of SA was observed in 35% of the patients with catheters. Bacteremic catheters were more frequently observed in patients with diabetes mellitus (p = 0.03), peripheral atherosclerosis (p = 0.001), a previous history of bacteremia (p = 0.05), nasal carriage of SA (p = 0.0001), longer catheter survival time (p = 0.001), higher total intravenous iron dose (p = 0.001), more frequent urokinase catheter infusion (p < 0.01), and local infection (p < 0.001) compared with non-bacteremic catheters. Monovariate survival analysis showed that significant initial risk factors for bacteremia were nasal carriage of SA (p = 0.00001), previous bacteremia (p = 0.0001), peripheral atherosclerosis (p = 0.005), and diabetes (p = 0.04). This study confirms the relatively high incidence of bacteremia with tunneled double-lumen silicone catheters and its potential complications. Possible preventive actions are discussed according to the risk factors. PMID- 12119468 TI - Prostacyclin enhances the expression of LPS/INF-gamma-induced nitric oxide synthase in human monocytes. AB - BACKGROUND: Nitric oxide (NO) is an important mediator of inflammatory processes, including macrophage-mediated cellular host defense, and is found to be increased in peritonitis. The ability of human mononuclear cells to contribute to the NO production by expression of active inducible NO synthase (iNOS) is still discussed controversely. AIMS: This study was designed to investigate the influence of prostacyclin receptor (IP receptor) activation on iNOS expression and NO formation in human peripheral blood monocytes. METHOD AND RESULTS: Using reverse transcriptase-polymerase chain reaction, we demonstrated that human monocytes express high levels of IP receptor mRNA. Stimulation of monocytes with the IP receptor selective agonist cicaprost (100 nM) significantly induced cellular cyclic adenosine monophosphate formation, indicating functional coupling of the receptor to G(s). Treatment of cells with lipopolysaccharide (LPS)/interferon gamma (IFN-gamma) further enhanced the IP receptor mRNA expression 2.7 +/- 0.1-fold above basal levels (n = 6). Analysis of iNOS expression revealed barely detectable mRNA levels in unstimulated monocytes which were increased 3.75 +/- 0.3-fold (n = 5) after costimulation with 1 microg/ml LPS and 250 U/ml INF-gamma for 16 h. Further increases of iNOS mRNA expression (9.4 +/- 0.9-fold above basal, n = 5) were obtained, if the monocytes were costimulated with 1 microg/ml LPS, 250 U/ml INF-gamma, and 100 nM cicaprost for 16 h. Measurement of the NO generation correlated with the polymerase chain reaction data: treatment of cells with 1 microg/ml LPS plus 250 U/ml INF-gamma increased the NO(2) production to 2.6 microM, being above the basal level of 2.0 microM, as determined in the cell culture medium. Additional treatment with 100 nM cicaprost further significantly increased the NO(2) production to 3.43 microM. CONCLUSIONS: An IP receptor mediated increase in cyclic adenosine monophosphate formation plays an important role in enhancing LPS/IFN-gamma-induced iNOS expression in human monocytes/macrophages and may, therefore, contribute to the increased production of NO during peritonitis. PMID- 12119470 TI - Inflammatory cytokines and lipopolysaccharide induce Fas-mediated apoptosis in renal tubular cells. AB - BACKGROUND/AIMS: Increased susceptibility of the kidney to acute renal failure (ARF) in the setting of sepsis even in the absence of systemic hypotension is well known. In the hypothesis that the proinflammatory cytokines and lipopolysaccharide (LPS) in gram-negative sepsis can directly cause renal tubular cell apoptosis via Fas- and caspase-mediated pathways, we examined apoptosis and Fas, Fas ligand, FADD expression, as well as PARP cleavage in cultured human proximal tubular cells under the cytokine and LPS-stimulated conditions. METHODS: HK-2 cell, immortalized human proximal tubular cell lines, were treated with 5 and 30 ng/ml of tumor necrosis factor-alpha (TNF-alpha), 5 and 20 ng/ml of interleukin-1beta (IL-1beta) and 30 ng/ml LPS for 24 h. Fas expression was examined by RT-PCR and Fas ligand, Fas-associated protein with death domain (FADD) and poly ADP ribose polymerase (PARP) cleavage were examined by Western blot analysis. Apoptosis was assessed by flow cytometer using Annexin V-FITC and propidium iodide (PI) staining and also by terminal deoxynucleotidyl transferase mediated dUTP nick end labelling (TUNEL) methods. RESULTS: Fas mRNA expression (ratio of Fas/L-19) increased in the TNF-alpha 5, 30 ng/ml and LPS treated group (p < 0.01, p < 0.01, p = 0.02), but there was no difference between the low- and high-dose TNF-alpha groups. Fas ligand protein expression did not increase in the low-dose TNF-alpha treated group, but it increased significantly in the high-dose TNF-alpha treated group (p < 0.01), IL-1beta- and LPS-treated groups (p < 0.01, p = 0.01, p < 0.01, p = 0.02). The intracellular adaptor protein, FADD expression also increased significantly in the high-dose TNF-alpha- and IL-beta-treated groups (p = 0.04, p = 0.04), but in the low-dose TNF-alpha and IL-beta treated group, it did not show statistically significant differences. In the LPS group, FADD expression also showed an increased tendency, but it was not statistically significant (p = 0.09). Western blot for PARP, a DNA repair enzyme mainly cleaved by caspase 3, showed increased 89- and 24-kD PARP cleavage products in TNF-alpha, IL-1beta and LPS treated cells. The degree of apoptosis examined by DNA fragmentation and translocation of membrane phosphatidyl serine significantly increased in cytokines and LPS treated groups. CONCLUSION: These results suggest that Fas- and caspase-mediated apoptosis of tubular cells by inflammatory cytokines and LPS can be one of the possible mechanisms of renal dysfunction in endotoxemia. PMID- 12119471 TI - Dysregulation of podocyte phenotype in idiopathic collapsing glomerulopathy and HIV-associated nephropathy. AB - BACKGROUND: Idiopathic collapsing glomerulopathy (ICG) and HIV-associated nephropathy (HIV-AN) are characterized by severe nephrotic syndrome, collapse and sclerosis of the glomerular tuft with prominent podocyte alterations and extensive tubulointerstitial lesions. We previously showed phenotypic changes in podocytes from patients with diffuse mesangial sclerosis, a severe glomerulopathy sharing several morphological features with collapsing glomerulopathy. The aim of this study was to analyze the podocyte phenotype in ICG and HIV-AN. METHODS: Using immunohistochemical techniques, we studied the podocyte expression of the transcription factor WT1 and its target PAX2, GLEPP1, synaptopodin and vimentin as markers of podocyte maturity and of proliferating cell nuclear antigen (PCNA) as a marker of proliferation. Apoptosis was analyzed by the TUNEL method. Results from renal biopsies of ICG and HIV-AN were compared with those obtained from normal kidney, minimal change nephrotic syndrome (MCNS), focal and segmental glomerulosclerosis (FSGS) and membranous glomerulonephritis (MGN). RESULTS: Abnormal distribution of WT1 and PAX2 and extensive loss of podocyte markers were observed in ICG and HIV-AN; this dysregulation was associated with podocyte proliferation without detectable apoptosis. In contrast, no podocyte changes were detected in MCNS or MGN. In FSGS, phenotypic changes, without proliferation, were restricted to podocytes surrounding focal and segmental glomerular lesions. Increased PCNA expression and apoptosis were observed in ICG and HIV-AN tubular cells. CONCLUSION: Dysregulation of podocyte phenotype and proliferation are present in both ICG and HIV-AN. This suggests that, whatever their etiology, both types of collapsing glomerulopathy share a common pathogenic pathway. Upregulation of cell proliferation and apoptosis observed in tubular epithelial cells is probably involved in the occurrence of severe tubulointerstitial lesions in collapsing glomerulonephritis. PMID- 12119472 TI - Platelet activation markers in patients with nephrotic syndrome. A comparative study of different platelet function tests. AB - BACKGROUND/AIM: Enhanced platelet reactivity may play a significant role in the genesis of the hypercoagulable state of nephrotic syndrome. However, the role of platelet function testing in nephrosis is controversial, partly because the methods used to assess platelet function (platelet aggregation and immunoassays of plasma beta-thromboglobulin and platelet factor 4) have such marked methodological problems. In the present study, we evaluated several tests assessing platelet function in 18 adult patients with idiopathic nephrotic syndrome and normal renal function. METHODS: Platelet function was assessed by measurement of plasma beta-thromboglobulin (enzyme-linked immunosorbent assay, ELISA), plasma P-selectin (ELISA), circulating platelets exposing the activation dependent antigens P-selectin (CD62P) and lysosomal GP53 (CD63) (flow cytometry), and by aggregation response to agonists such as ADP and collagen. Results were compared to those obtained in a group of 16 age- and gender-matched healthy subjects. RESULTS: Levels of plasma beta-thromboglobulin (p = 0.001), plasma P selectin (p < 0.001), and CD62P/CD63-positive platelets (p < 0.001 for both) were increased in nephrotic patients as compared to healthy controls. Platelet hyperaggregability in vitro was found in 13/18 patients. The reproducibility of platelet activation markers, as assessed by blood sample collection a week later from all patients, was found to be higher for plasma P-selectin (Spearman correlation coefficient, R = 0.99) and circulating activated platelets (CD62P: R = 0.97; CD63: R = 0.96) than for plasma beta-thromboglobulin (R = 0.78). CONCLUSIONS: Pronounced platelet activation takes place in nephrotic syndrome and may contribute to the hypercoagulability of nephrosis. Whole blood flow cytometry assay of platelet activation and plasma P-selectin assay may represent useful tests to assess the hypercoagulable state in nephrotic patients. PMID- 12119473 TI - Expression of a functional asialoglycoprotein receptor in human renal proximal tubular epithelial cells. AB - BACKGROUND: The asialoglycoprotein receptor (ASGPR) is a C lectin which binds and endocytoses serum glycoproteins. In humans, the ASGPR is shown mainly to occur in hepatocytes, but does occur extrahepatically in thyroid, in small and large intestines, and in the testis. In the kidney, there has been evidence both for and against its existence in mesangial cells. METHODS: Standard light microscopy examination of renal tissue stained with an antibody against the ASGPR was performed. The mRNA expression for the ASGPR H1 and H2 subunits in primary human renal proximal tubular epithelial cells (RPTEC), in the human proximal tubular epithelial cell line HK2, and in human renal cortex was investigated using reverse-transcribed nested polymerase chain reaction. ASGPR protein expression as well as ligand binding and uptake were also examined using confocal microscopy and flow cytometry (fluorescence-activated cell sorting). RESULTS: Light microscopy of paraffin renal biopsy sections stained with a polyclonal antibody against the ASGPR showed proximal tubular epithelial cell staining of the cytoplasm and particularly in the basolateral region. Renal cortex and RPTEC specifically have mRNA for both H1 and H2 subunits of the ASGPR, but HK2 only expresses mRNA for H1. Using a monoclonal antibody, the presence of the ASGPR in RPTEC was shown by fluorescence-activated cell sorting and immunofluorescent staining. Specific binding and uptake of fluorescein isothiocyanate labelled asialofetuin which is a specific ASGPR ligand was also demonstrated in RPTEC. CONCLUSIONS: Primary renal proximal tubular epithelial cells have a functional ASGPR, consisting of the H1 and H2 subunits, that is capable of specific ligand binding and uptake. PMID- 12119474 TI - Low prevalence of hypercalciuria in Japanese children. AB - BACKGROUND/AIM: There are several factors, such as race, age, sex, and geographical variations, associated with renal stone formation. Although it is known that the prevalence of urolithiasis in Japanese children is low, the reason remains obscure. We hypothesize that the low prevalence of urolithiasis is associated with the urinary calcium excretion. The aim of our study was to investigate the prevalence of hypercalciuria in Japanese children. METHODS: This investigation is a population-based school survey. A group of 529 healthy Japanese children was screened for hypercalciuria by measurement of the urinary Ca/Cr ratio using the morning fasting urine. In addition, the urinary Na/Cr ratio was also calculated for each subject. RESULTS: Hypercalciuria regarded as an urinary Ca/Cr value of more than 0.17 was noted only in 3 out of 529 children (0.6 %), while most cases (494/529, 93.4%) demonstrated hypocalciuria (urinary Ca/Cr <0.05). The mean urinary Ca/Cr value was 0.024 in all subjects combined. Linear regression analysis revealed a positive direct correlation between urinary Ca/Cr and Na/Cr values (rs = 0.14, p < 0.01). The urinary Ca/Cr ratio was not related to age in either sex. CONCLUSIONS: The present study demonstrates that the prevalence of hypercalciuria in Japanese children is low as compared with other countries, even though absorptive hypercalciuria and dietary hypercalciuria might be missed in this setting. This low prevalence of hypercalciuria may be associated with the lower prevalence of urolithiasis in Japanese children. As it is suggested that a low dietary intake of calcium and sodium may play some role in the low urinary calcium excretion, a randomized, controlled study comparing the efficacy of different modes of therapy, such as a low-calcium diet and/or a low-salt diet, might provide valuable information for the prevention of urolithiasis. PMID- 12119476 TI - Ultrapure dialysis fluid lowers the cardiovascular morbidity in patients on maintenance hemodialysis by reducing continuous microinflammation. AB - The aims of our prospective 3-year investigation were (1) to clarify whether high C-reactive protein (CRP) levels are an intermittent or a continuous phenomenon in individual hemodialysis patients and (2) to evaluate a possible relationship between ultrapure dialysis fluid associated CRP levels and an increased prevalence of atherosclerosis in a group of 60 hemodialysis patients treated either with conventional (n = 38) or on-line-produced ultrapure dialysis fluid (n = 22). Primary end points of the study were angiographically confirmed cerebrovascular, cardiovascular, or peripheral vascular events. Measurements of the CRP levels were done every 3 months using a highly sensitive assay. The CRP levels were normal (<0.5 mg/dl) in 45 patients and raised in 15 patients at the time of recruitment. In 87% of the patients with normal CRP levels, ultrapure dialysis fluid was used. The CRP levels measured at recruitment and at various time points thereafter did not differ significantly within patient groups. However, patients with increased CRP concentrations experienced significantly more vascular events as compared with patients with normal CRP levels (11 events vs. 1 event; p < 0.001). The data indicate that continuous induction of acute phase proteins represents a nontraditional vascular risk factor contributing to the development and progression of atherosclerosis in dialysis patients. Ultrapure dialysis fluid lowers cardiovascular morbidity by preventing/reducing chronic microinflammation. PMID- 12119475 TI - Effects of L-arginine and L-NAME on the renal function in hypertensive and normotensive subjects. AB - BACKGROUND/AIM: Renal vasodilation in response to L-arginine has been reported to be diminished in hypertensive (HT) subjects. If this diminished renal vasodilator response indicates disturbance of the renal NO pathway, a diminished renal vasoconstrictor response to NO synthase inhibition may be present in HT subjects as well. The present study was conducted to compare the effects of L-arginine and N(G)-nitro-L-arginine methyl ester (L-NAME) on renal and systemic hemodynamics between HT and normotensive (NT) subjects. METHODS: The responses of renal and systemic vascular resistances (RVR and SVR) and plasma noradrenaline and renin (NOR and PRA) to systemic NO stimulation and inhibition were studied in patients with grade 1 essential HT and age- and sex-matched NT subjects. On separate occasions, after baseline values were obtained, 40-min randomly administered intravenous infusions of L-arginine (12.5 mg.kg(-1).min(-1)) or L-NAME (0.0125 mg.kg(-1).min(-1)) were given. RESULTS: Baseline values of RVR (129 +/- 21 and 162 +/- 10 resistance units) and SVR (15.1 +/- 4.3 and 21.6 +/- 5.1 resistance units) were higher (p < 0.01) in HT than in NT subjects, whereas the baseline values of NOR and PRA were similar. Infusion of L-arginine caused similar decrements in SVR (29 +/- 10 and 31 +/- 11%), but the decrease in RVR was smaller (22 +/- 8 and 35 +/- 12%, respectively; p < 0.05) in HT than in NT subjects. In response to L-NAME, the increments in RVR (66 +/- 10 and 61 +/- 25%) and SVR (36 +/- 21 and 34 +/- 18%) were similar in HT and NT subjects. In both groups, infusion of L-arginine was associated with similar increments, whereas infusion of L-NAME was associated with similar decrements in NOR and PRA. CONCLUSIONS: This study confirms the smaller renal vasodilator response to L-arginine in HT than in NT subjects. Whether this is caused by a disturbance of the renal NO pathway remains doubtful considering the observed similar L-NAME-induced increments in RVR and SVR in the two groups of subjects. PMID- 12119478 TI - Erythrocyte ferritin in patients with chronic renal failure and heterozygous beta thalassemia. AB - AIM: The aim of this research is to study the variance of erythrocyte ferritin (EF) in patients with chronic renal failure (CRF) and heterozygous beta thalassemia (beta-TA), as well as the use of EF as a more reliable index for assessing the body iron status. METHODS: We studied 63 subjects with CRF, 40 subjects with heterozygous beta-TA, 53 subjects with CRF and heterozygous beta-TA and 24 normal subjects. In 11 patients with CRF and heterozygous beta-TA, sternal bone marrow aspiration was performed to evaluate iron stores in the bone marrow. EF was determined in the hemolysate of washed erythrocytes by a radioimmunoassay. RESULTS: EF showed the strongest correlation with bone marrow iron (p < 0.001) in comparison with the remaining hematological parameters that were examined. Patients with CRF without heterozygous beta-TA showed an increase in serum ferritin (SF), even in cases of iron deficiency. In the group of heterozygous beta-TA without renal failure, 22.5% of patients showed an increased EF content up to 150 ag/cell and a tendency for iron overload. Patients with CRF and heterozygous beta-TA showed a high value of EF, up to 200 ag/cell, and iron overload in 22.6%, almost the same proportion as in the previous group. It was also observed that a high value of SF does not indicate iron overload for these patients. In the group of hemodialysis, patients without heterozygous beta-TA who were under erythropoietin (EPO) treatment presented iron deficiency. Many patients with CRF and heterozygous beta-TA who were taking EPO presented iron overload, while very few of them presented iron deficiency. CONCLUSION: These findings suggest that EF is a reliable index for assessing the iron status in patients with CRF and heterozygous beta-TA. PMID- 12119477 TI - Triad of malnutrition, inflammation, and atherosclerosis in hemodialysis patients. AB - BACKGROUND/AIM: As chronic inflammation underlies both atherosclerosis and malnutrition, a possible link between these factors has been suggested in hemodialysis (HD) patients. We designed this study to compare nutritional indices and inflammatory parameters of HD patients with demonstrated atherosclerosis (group I) and HD patients without (group II). METHODS: We included 59 and 57 patients in groups I and II, respectively. The patient groups were matched for the risk factors for atherosclerosis such as age, gender, smoking habits, hypertension, and HD duration. The nutritional status of the patients was evaluated according to laboratory parameters, normalized protein catabolic rate, anthropometric measurements, and subjective global assessment. RESULTS: Laboratory parameters (albumin, prealbumin, total cholesterol, phosphorus, creatinine), normalized protein catabolic rate, and triceps skinfold thickness revealed a significant decline in the nutritional status of the patients with atherosclerosis. We found that the patients with atherosclerosis had significantly higher C-reactive protein, ferritin, and fibrinogen levels when we compared the patient groups for acute-phase reactants. When we assessed malnutrition as being in category B/C (B = mild to moderately malnourished, C = severely malnourished) according to subjective global assessment and inflammation on the basis of a C-reactive protein level > or =10 mg/l, among patients with atherosclerosis, there was a significantly higher proportion of them having malnutrition and inflammation. Additionally, the proportion of patients without any evidence of malnutrition and inflammation was significantly lower in group I than in group II. CONCLUSION: Our study gives evidence for the possible triad of malnutrition, inflammation, and atherosclerosis in HD patients. PMID- 12119479 TI - Changes in bone mineral density, insulin-like growth factor-1 and insulin-like growth factor binding protein-3 in kidney transplant recipients. A longitudinal study. AB - BACKGROUND: Osteopenia is a major complication of renal transplantation (RTx). This cross-sectional and longitudinal study was planned to better define long term bone status and relationship to IGF system components. METHODS: Serial measurements of bone mineral density (BMD) and serum markers were performed in 30 patients prior to RTx and at 6 and 12 months following RTx. Serum concentrations of insulin-like growth factor-1 (IGF-1), insulin-like growth factor binding protein-3 (IGFBP-3), vitamin D, and intact parathyroid hormone (iPTH) were measured. RESULTS: Serum creatinine, phosphate, alkaline phosphatase and osteocalcine levels decreased, serum calcium levels increased and serum iPTH levels did not change significantly after transplantation. The mean BMD of the vertebrae was 0.97 +/- 0.22 g/cm(2) at the time of RTx, 0.87 +/- 0.21 g/cm(2) 6 months post-RTx (p < 0.05), and 0.81 +/- 0.21 g/cm(2) 12 months post-RTx (p < 0.05). Femur BMD also declined from 0.79 +/- 0.16 to 0.72 +/- 14 g/cm(2) at 12 months (p < 0.05). There was a significant increase in the IGF-1 and a significant reduction in the IGFBP-3 concentrations at 6 months post-RTx (p < 0.05). Significant correlations between serum IGF-1 concentrations and vitamin D concentrations were noted only at 6 months. There was no significant correlation between the BMD and serum IGF system. CONCLUSIONS: These results demonstrate a significant loss of BMD after RTx. The circulating levels of IGF-1 and IGFBP-3 stimulated by the reduction in BMD and IGF-1 secretion are increased in order to restore bone formation. PMID- 12119480 TI - Calcium and phosphate plasma levels in dialysis patients after dietary Ca-P overload. Role of gastric acid secretion. AB - In normal subjects, the gastric ionisation of calcium and phosphate seems to be a prerequisite for their intestinal absorption. We investigated the behavior of the plasma calcium and phosphate profile in 30 patients on regular dialysis treatment in the 6 h following a meal containing 1 g of calcium and 2 g of phosphate. Moreover, to assess the role of gastric acidity, the study was repeated after 3 days on omeprazole administration, to nearly abolish gastric acid secretion. Both total plasma calcium and ionized calcium peaked after the meal (at 30 and 120 min, respectively) only in basal study, while no peak was observed after the administration of omeprazole. Surprisingly, both in basal and in the omeprazole study the levels of plasma phosphate did not increase after the test meal. In conclusion, as in normal subjects, the gastric ionization of dietary calcium promotes the intestinal absorption of calcium in uremic patients on dialysis treatment, while the acute gastric acid inhibition by omeprazole reduced the intestinal calcium transport. In contrast, with the "trade off" hypothesis we did not observe any postprandial phosphate peak after the dietary load, and, in contrast with normal subjects, omeprazole administration did not influence the phosphate profile. PMID- 12119481 TI - Effects of oral adsorbent AST-120 (Kremezin) on renal function and glomerular injury in early-stage renal failure of subtotal nephrectomized rats. AB - The aim of the present study was to determine if treatment with an oral adsorbent (AST-120, Kremezin) might decrease the urinary albumin excretion and serum indoxyl sulfate (s-IS), and prevent glomerular sclerosis in early-stage renal failure, i.e. 0.9-1.2 mg/dl of serum creatinine (s-Cr) and 60-95 mg/dl of blood urea nitrogen (BUN), in subtotal (3/4) nephrectomized rats. Levels of s-Cr and s IS in the AST-120-treated rats were significantly lower than those in the untreated control rats. The AST-120-treated rats showed an increase of creatinine clearance. Urinary protein and indoxyl sulfate excretion in the AST-120-treated rats were also significantly lower than those in the untreated control rats. The ratio of glomerular tuft area to the area of Bowman's capsules (GT/BC) in the AST 120-treated rats was significantly lower than that in the untreated control rats. The degree of glomerular sclerosis and tubulointerstitial fibrosis in the AST-120 treated rats was significantly lower than that in the untreated control rats. Furthermore, there was a significant relationship among the degree of GT/BC, glomerular sclerosis, tubulointerstitial fibrosis and the levels of urinary protein excretion. It appears that AST-120 might decrease the accumulation of s Cr and s-IS, and prevent glomerular sclerosis in early stage renal failure in the subtotal nephrectomized rats. PMID- 12119482 TI - Dietary fatty acid supplementation modulates the urinary excretion of calcium and oxalate in the rat. Insight into calcium lithogenesis. AB - BACKGROUND: An anomalous plasma phospholipid polyunsaturated fatty acid composition has been reported in calcium nephrolithiasis, and was proposed to play a crucial role in the pathogenesis of hypercalciuria and hyperoxaluria, well known risk factors for lithogenesis. METHODS: To confirm this hypothesis, we administered rats three different diets rich in coconut, soybean and fish oils, and evaluated their effect on plasma urinary calcium and oxalate excretion, since the quality of fatty acids represents an important factor able to influence the activity of delta-6-desaturase, the rate-limiting enzyme in the biosynthetic pathway of highly unsaturated fatty acids. RESULTS: In comparison with coconut and fish oil, dietary supplementation with soybean oil increased plasma phospholipid arachidonic acid and serum 1,25-vitamin D(3) values, as well as renal tissue calcium content and urinary excretion of sodium, oxalate and calcium. CONCLUSIONS: Our findings demonstrate that the quality of fatty acids may modify the urine excretion of calcium and oxalate, confirming our previous hypothesis of a pathogenetic link between cellular membrane phospholipid polyunsaturated fatty acid composition and calcium nephrolithiasis. In addition, our study provides new insights into the relationship between dietary, environmental factors and renal stone disease. PMID- 12119484 TI - Efficacy and safety of losartan in patients with proteinuria. AB - The renoprotective effect and safety of an angiotensin II receptor blocker, losartan, were studied in 11 diabetic and 14 nondiabetic patients with a daily urinary protein excretion >500 mg. Once-daily losartan was administered to all patients for 16 weeks without diuretics or other antihypertensive agents. A daily dose of 50 mg was given to those patients with a sitting systolic blood pressure of between 130 and 170 mm Hg; 25 mg was given to patients with a systolic blood pressure of between 110 and 130 mm Hg. Sixteen patients (6 diabetic and 10 nondiabetic patients) had a more than 25% decrease of their daily urinary protein excretion (response rate 64%). The mean decrease in these 16 patients was 57 +/- 17% (p < 0.05). Two patients (1 diabetic and 1 nondiabetic) had more than a 25% increase of their urinary protein excretion. The trough sitting systolic blood pressure of all patients (n = 25) decreased from 142 +/- 17 to 125 +/- 13 mm Hg (p < 0.05) and the trough sitting diastolic blood pressure from 87 +/- 11 to 78 +/- 11 mm Hg (p < 0.05). Serum uric acid was measured in 16 patients; a decrease from 7.3 +/- 1.6 to 6.6 +/- 1.4 mg/dl (a 9.6% decrease, p < 0.05) was found after 16 weeks. Our study showed that in both diabetic and nondiabetic proteinuric patients once-daily losartan, given as monotherapy at doses of 25 or 50 mg, was effective in reducing blood pressure, serum uric acid levels, and daily urinary protein excretions. PMID- 12119483 TI - Characterization of TH1/TH2 profile in uremic patients. AB - End-stage renal failure (ESRD) induces a clinical state of immunodeficiency with a higher incidence of infections and a higher mortality due to infectious complications compared with the normal population. Using a newly developed immunofluorescent staining of intracellular cytokines for flow cytometric analysis, we studied Th subsets in 22 healthy control subjects, 28 patients with compensated chronic renal failure (CRF), 25 patients on hemodialysis (HD), and 24 patients on continuous ambulatory peritoneal dialysis (CAPD). Our results demonstrate that the percentage of both interferon-gamma-positive cells and interleukin-4-positive cells increased in compensated CRF patients compared with those in healthy subjects. Moreover, a significantly higher percentage of CD4 positive cells is characterized by a Th1-type cytokine production pattern in HD patients and by a Th2-type cytokine secretion pattern in CAPD patients. These results suggest that the altered Th1/Th2 balance may be associated with the pathogenesis of ESRD. PMID- 12119486 TI - Resolution of nodular glomerular lesions in a patient with light-chain nephropathy. AB - A 37-year-old man developed nephrotic syndrome and renal insufficiency in 1986. He had kappa-light-chain protein both in serum and urine. A renal biopsy showed nodular glomerulosclerosis with deposition of kappa-light-chains in the mesangial area, compatible with light-chain nephropathy. Thereafter, he was treated with steroids and melphalane and the light-chain protein disappeared from both the urine and serum. Although his moderately impaired renal function maintained stable levels for over 10 years, he was diagnosed as having renal cell carcinoma in 1998, and a right nephrectomy was performed. Histopathological examination of a portion of the removed kidney, unaffected by carcinoma, showed mild mesangial proliferation, and both the nodular lesions and light-chain deposits were no longer observed. These observations suggest that an established nodular glomerular lesion may be reversible. PMID- 12119485 TI - ACE gene polymorphism and disease progression of IgA nephropathy in Asians in Singapore. AB - The deletion polymorphism of the angiotensin-converting enzyme (ACE) gene has been considered as a risk factor for IgA nephropathy and for its progression to end-stage renal failure. However, results from various studies are conflicting. We had genotyped the ACE gene in 100 patients with IgA nephropathy, 32 of whom were in end-stage renal failure and in 90 normal adult subjects. All DD cases were subjected to confirmation with a second PCR, performed with the insert specific forward primer. Similar genotype frequencies were obtained for the 90 normal control subjects (II: 47%, ID: 44%, DD: 9%); for the 68 patients not in end-stage renal failure (ESRF) (II: 47%, ID: 46%, DD: 7%) and for the 32 patients with ESRF (II: 53%, ID: 38%, DD: 9%). The genotype frequencies in all 3 series are in Hardy-Weinberg equilibrium. These results suggest that ACE gene polymorphism is not a risk factor for IgA nephropathy and is not a predictor for its progression. Definitive proof of association between ACE gene polymorphism and progression in IgA nephropathy will require a prospective study, controlled for important risk factors, with adequate patient numbers and facility for confirming DD genotypes. PMID- 12119487 TI - Effect of mizoribine on IL-6 release by peripheral blood mononuclear cells. AB - BACKGROUND/AIM: A novel immunosuppressant, mizoribine (MZB), has recently been reported to be effective in the treatment of IgA nephropathy (IgAN), although its mechanism of action remains unknown. This study was conducted to investigate whether the efficacy of MZB on IgAN is exerted by suppression of interleukin-6 (IL-6) release. METHODS: Peripheral blood mononuclear cells (PBMC) were collected from 4 children with IgAN (median age 13.0 years) and 4 control children (median age 5.2 years). PBMC were cultured with medium alone or medium with lipopolysaccharide (LPS), and then incubated with LPS and MZB. Culture supernatants were assayed for lL-6. RESULTS: IL-6 release was increased by LPS in all subjects. Although the median value was higher in IgAN patients (median increase in IL-6 release 1,298.1%) than in controls (median 489.2%), statistical significance was not reached (p > 0.05). 10 mg/ml of MZB suppressed the release of IL-6 in both IgAN patients (median decrease in IL-6 release 39.3%) and controls (median 43.2%), with statistical significance (p < 0.05 and p < 0.01, respectively). CONCLUSION: This study suggests that MZB could suppress IL-6 release in vitro and thus may exert its efficacy on IgAN. PMID- 12119488 TI - Cyclosporin A treatment for membranoproliferative glomerulonephritis type II. AB - A nephrotic patient with membranoproliferative glomerulonephritis type II (MPGN II) was treated with cyclosporin A (CSA) and alternate-day low-dose prednisolone. This patient developed the nephrotic syndrome twice. The second episode of the nephrotic syndrome was steroid resistant, and therefore this patient was treated with a CSA regimen. During treatments with alternate-day low-dose prednisolone and CSA, this patient recovered from the nephrotic syndrome. We conclude that CSA therapy may be effective for patients with the steroid-resistant nephrotic syndrome caused by MPGN II. PMID- 12119489 TI - Cast nephropathy in a case of Waldenstrom's macroglobulinemia. AB - Waldenstrom's macroglobulinemia is a low-grade lymphoplasmacytic lymphoma characterized by a circulating monoclonal IgM. The clinical manifestations are due to deposition of IgM in liver, spleen, and/or lymph nodes. Related symptoms include anemia and complications of the hyperviscosity syndrome. Renal involvement in classical cases of Waldenstrom's macroglobulinemia is rare, and the pathological hallmark finding in the renal biopsy specimen is a thrombotic microangiopathy. We report the case of a 73-year-old female with the diagnosis of pernicious anemia for 2 years before she presented with acute renal failure. A renal biopsy performed suggested the diagnosis of myeloma cast nephropathy. However, bone marrow biopsy specimens and hematological studies did not support this diagnosis. Serum and urinary protein electrophoresis revealed a monoclonal lambda subtype IgM. Ultrasound imaging showed an enlarged spleen. The diagnosis of cast nephropathy in a patient with Waldenstrom's macroglobulinemia was made. She underwent treatment with fludarabine and plasmapheresis/hemodialysis with dramatic improvement of her renal function. PMID- 12119490 TI - A rare cause of pulmonary-renal syndrome. AB - Diseases affecting both the lung and the kidney have grave prognosis and serious diagnostic and therapeutic consequences. Here, 3 cases of pulmonary-renal syndrome caused by antiphospholipid syndrome are reported. The patients presented with dyspnea, renal insufficiency, pulmonary infiltrates on chest X-ray and areas of ground glass attenuation on computed tomography of the lungs. There were no signs of infectious disease, vasculitis or myocardial insufficiency. Clinical findings, antiphospholipid levels and histological findings in transbronchial and/or renal biopsy proved the diagnosis of antiphospholipid syndrome. Antiphospholipid syndrome is a comparatively rare disorder which is relevant in the differential diagnosis of diseases affecting both lung and kidney and requires specific therapeutic measures. PMID- 12119491 TI - Successful treatment of renovascular hypertension due to fibromuscular dysplasia by intravascular ultrasound-guided atherectomy. AB - A 22-year-old man presented with renovascular hypertension, based on a stenosis of the distal portion of the right renal artery with a "string of beads"-like appearance. An intravascular ultrasound image at the renal artery lesion revealed irregularity of the vascular wall. Directional atherectomy was performed and histopathology of atherectomised tissues showed medial fibroplasia, a common type of fibromuscular dysplasia. After atherectomy his hypertension was markedly improved. We report here a case of renovascular hypertension due to fibromuscular dysplasia, successfully diagnosed and treated with IVUS-guided renal atherectomy. PMID- 12119493 TI - Ultrapure dialysis fluid and response to hepatitis B vaccine. PMID- 12119492 TI - Gonadal mosaicism of Frasier syndrome in 3 Chinese siblings with donor splice site mutation of Wilms' tumour gene. AB - Frasier syndrome is a rare human developmental disorder classically affecting 46,XY females and leading to male pseudohermaphroditism and chronic renal failure. We describe a family with both 46,XX and 46,XY females affected by the syndrome due to WT1 splice site mutations. The diagnosis of Frasier syndrome in 1 of the children led to the discovery of the syndrome in 2 other siblings, of whom 1 is asymptomatic. Since the mutation was not found in either parents, gonadal mosaicism was suggested. The implication of family screening for WT1 gene mutation in asymptomatic members is also discussed. PMID- 12119494 TI - Reversible proximal tubular dysfunction in a patient with acute febrile illness, marked hyperbilirubinemia and normal renal function: evidence of leptospirosis. PMID- 12119495 TI - Evaluation of esophageal [correction of osephageal] motor function in chronic renal failure and the role of hemodialysis treatment. PMID- 12119497 TI - Cardiac functions, autonomic nervous system and lipoprotein values in white coat hypertension patients. PMID- 12119496 TI - Low blood pressure, decreased incidence of hypertension, and renal cardiac, and autonomic nervous system functions in patients with sickle cell syndromes. PMID- 12119498 TI - Genetic engineering of allergens: future therapeutic products. AB - Genetic engineering of allergens for specific immunotherapy should aim at the production of modified molecules with reduced IgE-binding epitopes (hypoallergens), while preserving structural motifs necessary for T cell recognition (T cell epitopes) and for induction of IgG antibodies reactive with the natural allergen (blocking antibodies). Common approaches for engineering of hypoallergens usually require knowledge of T and B cell epitopes and involve changing specific base pairs (mutated gene), introduction of a new piece of DNA into the existing DNA molecule (chimeric or hybrid gene), and deletions (truncated gene or fragments). DNA family shuffling has the advantage that it does not require a priori knowledge of structural and functional properties for efficient generation of hypoallergens. The combination of the hypoallergen concept with the Th1-inducing genetic immunization approach might be an attractive alternative for protein-based immunotherapy. PMID- 12119500 TI - Identification and characterization of major allergens in excretory/secretory products of the worm Paragonimus ohirai. AB - Allergens present in the excretory/secretory (ES) products of adult Paragonimus ohirai were biochemically identified. Immunoblot analysis using sera from P. ohirai-infected rats revealed only two allergens to be major proteins in the ES products, with apparent molecular masses (M(r)) of 27 and 29 kD. As the ES products contained a high proportion of acidic and neutral cysteine proteinases, we examined whether or not the allergens and the cysteine proteinases were identical. The acidic and neutral cysteine proteinases were biochemically purified from the ES products and showed M(r) of 27 and 29 kD, respectively. The two cysteine proteinases had almost identical N-terminal amino acid sequences and were reactive with specific IgE in sera from the infected rats. The allergenicity of the cysteine proteinases was confirmed by 48-hour homologous passive cutaneous anaphylaxis. Immunoblot and immunocapture assays using anti-human IgE monoclonal antibody showed that the proteinase allergens were reactive with specific IgE of patients with paragonimiasis westermani. Also, the cysteine proteinases were reactive with specific IgG of both the infected rats and the patients. Therefore the acidic and neutral cysteine proteinases prepared from the ES products of P. ohirai will be useful allergens and antigens for the immunodiagnosis of paragonimiasis. PMID- 12119499 TI - Preparation and characterization of silverfish (Lepisma saccharina) extract and identification of allergenic components. AB - BACKGROUND: Airborne insect antigens represent important aeroallergens which have been widely investigated. Although it has been demonstrated that house dust contains significant silverfish (Lepisma saccharina) levels, none of the extracts obtained so far has been extensively characterized. Thus, we have prepared and characterized a silverfish extract and investigated its IgE-reactive components by testing the reactivity of sera from patients allergic to inhalant insect allergens. METHODS: The extract from silverfish insect bodies was prepared by homogenizing frozen silverfish in Tris-HCl buffer. The soluble material (Sup) was filtered and the insoluble material (Ppt) was resuspended in 100 mM Tris pH 10.6. The two fractions were characterized by biochemical and immunochemical methods. IgE reactivity was investigated on both fractions before and after periodate treatment. RESULTS: Protein content and total carbohydrates was 2 and 3% w/w for Sup and 1 and 0.3% w/w for Ppt. The SDS-PAGE profile of the two fractions showed a different pattern in the MW range of 5-175 kD. Sup and Ppt, probed with allergic sera, showed a complex pattern of IgE reactivity. When periodate-treated fractions were tested, IgE reactivity was either completely abrogated, reduced or not affected, depending on the allergic serum employed. CONCLUSIONS: The results obtained indicate that the classic aqueous-extraction procedures that have been used up to now for other insects might not be completely satisfactory, since several allergenic components are not soluble at the normally used pH. We developed a dedicated extraction procedure allowing the detection of a certain degree of reactivity in sera negative to allergens extracted following classic procedures. PMID- 12119502 TI - Latex sensitization by dermal exposure can lead to airway hyperreactivity. AB - BACKGROUND: Using non-powdered, low-protein natural rubber latex (NRL) gloves has been shown to reduce the elicitation of respiratory symptoms in latex-allergic individuals; however, the role of dermal exposure in the induction of sensitization is not completely understood. OBJECTIVE: These studies were conducted to (1) determine levels of NRL protein in gloves currently in use and (2) evaluate, using a murine model, the potential for dermal exposure to induce NRL sensitization and subsequent airway hyperreactivity upon respiratory challenge. METHODS: Total extractable protein and NRL allergen levels were evaluated from 38 glove samples using the Lowry and CAP inhibition assays, respectively. BALB/c mice were dermally exposed to non-ammoniated latex (NAL, 6.25-25 microg) 5 days/week for 13 weeks and monitored weekly/biweekly for IgE levels. Airway hyperreactivity was determined following respiratory challenge with methacholine (MCH) or NAL proteins on days 60 and 93, respectively. RESULTS: Glove total protein and NRL allergen levels ranged from below the limit of detection to 946 microg/g and from 0.002 to 112 microg/g, respectively. Mice demonstrated dose-dependent increases in total serum IgE levels by day 58 with increased airway hyperreactivity observed upon respiratory challenge with MCH (day 60) or NAL proteins (day 93). CONCLUSIONS: These studies investigated the continued use of gloves with high levels of total extractable protein and NRL allergen. The potential for dermal exposure to induce NRL-specific IgE and airway hyperreactivity upon respiratory challenge suggests there should be continued concern regarding the induction of sensitization in individuals using non powdered latex gloves. PMID- 12119501 TI - Interest of two-dimensional electrophoretic analysis for the characterization of the individual sensitization to latex allergens. AB - BACKGROUND/OBJECTIVE: Latex allergy is a type 1 hypersensitivity reaction that mainly affects high-risk populations such as health care workers, spina bifida affected or multiply-operated children. Ten molecules have so far been identified and registered as latex allergens (Hev b 1 to Hev b 10). The aim of the present investigation was to identify the major latex allergens by an individual analysis of the IgE response of latex-allergic patients to latex proteins separated by two dimensional (2-D) gel electrophoresis. MATERIALS AND METHODS: Latex proteins from a sap or a glove extract were separated by 2-D electrophoresis and transferred to a nitrocellulose membrane. Each membrane was incubated with the serum of one latex-allergic patient. The most frequently recognized latex allergens were characterized in sap and glove extracts using monoclonal antibodies or amino acid microsequencing. RESULTS: The one-dimensional screening of 54 patient sera revealed 4 major bands recognized by IgE. The 2-D analysis of the sensitization to latex allergens allows the identification of allergen isoforms and the characterization of an individual response diversity. Hev b 6.01 was recognized by 88.9% of the patients. Protein spots around 14 kD were recognized by 48.1% of the patients and corresponded to Hev b 6.03 as well as other proteins. A not yet characterized doublet of acidic proteins with molecular masses of 43 and 94 kD was recognized by 20.4% of the sera. Only 5.5% of the sera did not recognize any of these 4 major allergens. Hev b 1 is the main protein from the glove extract but was not constantly found in sap extracts. CONCLUSIONS: One-dimensional electrophoretic analysis of the allergen is usually not sufficient to characterize the individual specificity of the IgE response to latex allergens. Latex-glove proteins which are allergens can be absent from the sap extracts and the sensitization to these allergens could be underestimated. Individual 2-D analysis of the sensitization to latex allergens is useful to define the best allergen mixture required for diagnosis and needed for individual therapy monitoring. PMID- 12119503 TI - The allergenicity of soybean-based products is modified by food technologies. AB - BACKGROUND: Numerous products based on soybean are available and various food technologies are applied for their production. The allergenicity of natural soybean may be modified by these treatments. OBJECTIVES: To compare the allergenicity of native soybean proteins with those of soy milk and texturized protein products. To show additional allergens. METHODS: Three commercial products and two infant formulas were studied: Soybean flour, soy milk, texturized soy proteins, two infant formulas; the first containing total proteins and the second containing a soy protein hydrolysate. Sera from 9 patients allergic to soy protein were tested by immunoblotting (IB). IB inhibition was achieved by incubating sera with protein extract from soybean flour. RESULTS: The SDS-PAGE profile of soybean flour protein and soy milk showed a difference in the proportions of the various protein fractions, with a higher concentration of 37 kD protein in flour and 33-kD protein in milk. Infant formula 1 contained proteins with a molecular weight below 28 kD. The texturized extract contained high proportions of 31- to 34- and 38-kD proteins. Immunoblotting revealed a lack of allergenicity in infant formula. Sera recognizing the 38- and 50-kD proteins in texturized soy protein also recognized the 37- and 49-kD proteins in soybean flour and in soy milk, suggesting a protein glycation by texturization processes. The 30- to 34-kD band in texturized proteins was devoid of any allergenicity. This study seems to indicate that the 30-kD allergen (Gly m Bd 30) disappears during the production of texturized soy protein. CONCLUSION: All technologies applied to soybean-based products induce striking variation in the protein profile and allergenicity. Texturized protein could lack the major allergen Gly m Bd 30. Further studies or texturization might generate modified technologies in order to create hypoallergenic texturized proteins. PMID- 12119504 TI - Murine strain differences in airway inflammation induced by diesel exhaust particles and house dust mite allergen. AB - BACKGROUND: Differences in allergic airway inflammation induced by ovalbumin (OVA) + diesel exhaust particles (DEP) in various murine strains have already been reported. However, there is no report that different murine strains respond differently towards house dust mites or DEP, which are known to aggravate allergic asthma. METHODS: The Dermatophagoides farinae allergens Der f (1 microg) or Der f (1 microg) + DEP (50 microg) were administered intratracheally to two different mouse strains (CBA/JN and C57BL/6N). Histological changes in the lung tissues, asthma-relevant cytokines in the lungs, and allergen-specific immunoglobulins in plasma were investigated. RESULTS: Der f treatment led to the proliferation of goblet cells, production of mucus plugs, and the recruitment of eosinophils and lymphocytes to the airways of the mice. The manifestation of the airway inflammation in the C57BL/6N mouse was much greater than in the CBA/JN mouse. The protein levels of interleukin (IL)-4 and IL-5, regulated on activation, normal T cell expressed, and presumably secreted (RANTES), and eotaxin in the lung tissue of C57BL/6N mice were higher than those in CBA/JN mice by a factor of 1.26 (IL-4), 5.26 (IL-5), 2.07 (RANTES) and 3.27 (eotaxin). DEP aggravated the manifestations of the eosinophilic inflammation in CBA/JN mice through goblet cell proliferation. However, the exact effect of DEP could not be evaluated in C57BL/6N because of its severe enhancement of the inflammation. DEP enhanced the local expression of IL-5, RANTES, and eotaxin in the CBA/JN mouse, and consequently triggered an increased IgG1 production in both strains. Allergen specific IgE antibodies were lower than 1 titer in both mice. CONCLUSION: The murine strain differences in the pathogenesis of allergic airway disease caused by mite allergen might be related to the local expression of the cytokines we screened. The aggravating effect of DEP may be mediated by an increase in the local expression of IL-5, RANTES, eotaxin, and the production of an antigen specific to IgG1. PMID- 12119505 TI - Chymase inhibitor improves dermatitis in NC/Nga mice. AB - BACKGROUND: Mast cell chymase is thought to participate in allergic inflammation, but its precise role remains undetermined. Inbred NC/Nga mice develop skin lesions similar to atopic dermatitis (AD) when they grow up in a conventional environment. To elucidate the possible role of chymase in AD, we examined the effect of a chymase inhibitor on skin lesions of NC/Nga mice. METHODS: NC/Nga mice were given the chymase inhibitor SUN-C8257 daily at 150 mg/kg/day with drinking water, and the severity of the dermatitis was evaluated on day 35 of the experiment. The role of chymase in dermatitis was further investigated in vitro and in vivo using recombinant mouse mast cell protease-4 (mMCP-4). RESULTS: Administration of SUN-C8257 significantly reduced the clinical skin and histological score in NC/Nga mice. SUN-C8257 also inhibited the accumulation of inflammatory cells, such as eosinophils and mast cells, in the affected lesions in this model. mMCP-4 stimulated eosinophil migration in vitro, and intradermal injection of the enzyme resulted in a significant accumulation of inflammatory cells, including eosinophils, at the injection site. Thus amelioration of the skin lesions in NC/Nga mice by SUN-C8257 might be, at least in part, due to the suppression of cell infiltration in the lesions. CONCLUSION: Mast cell chymase may contribute to the pathogenesis of AD, and SUN-C8257 will be beneficial to the treatment of the skin disorder. PMID- 12119506 TI - Chymase inhibitor ameliorates eosinophilia in mice infected with Nippostrongylus brasiliensis. AB - BACKGROUND: Chymase is a chymotrypsin-like serine protease primarily stored in mast cells. Infection with helminth parasites is known to increase the level of mast cell chymase in the jejunum and serum in mice. The aim of the present study is to elucidate the role of chymase in helminth infection. METHODS: Chymase inhibitor SUN-C8257 was administered to mice infected with the nematode Nippostrongylus brasiliensis, and the number of eosinophils in the blood, serum IgE levels and fecal egg counts were determined. RESULTS: Administration of SUN C8257 significantly inhibited blood eosinophilia in BALB/c mice infected with N. brasiliensis. The effect of SUN-C8257 was specific for eosinophils, in that it affected neither the number of total leukocytes nor serum IgE levels. SUN-C8257 did not alter the fecal egg counts in this model, showing that SUN-C8257 has no effect on infectivity and expulsion of the nematode. N. brasiliensis infection induced eosinophilia in mast cell-deficient mice (W/W(v)) as well as their littermates (+/+), and SUN-C8257 inhibited the eosinophilia in +/+ mice but not in W/W(v) mice. These results suggest that the eosinophil number may be regulated by different mechanisms in W/W(v) and +/+ mice, and that the effect of SUN-C8257 on nematode-induced eosinophilia is probably due to chymase inhibition. CONCLUSIONS: Chymase released by activated mast cells may play a role in helminth induced eosinophilia. PMID- 12119507 TI - Hypogammaglobulinemia in steroid-dependent asthmatics correlates with the daily dose of oral prednisolone. AB - BACKGROUND: Steroid-induced adverse effects including suppression of humoral immunity should be considered in steroid-dependent severe asthma. Only a few studies have determined the exact steroid dose that could potentially suppress humoral immunity in asthmatics. METHODS: Randomly selected 100 adult asthmatics treated with inhaled beclomethasone dipropionate (BDP) were classified into three groups based on the dose of steroid to determine the serum IgG, IgA and IgM levels by radioimmunoassay. Relationships between serum immunoglobulin levels and the daily dose and duration of oral prednisolone (PSL) therapy were examined. RESULTS: None of the patients on inhaled corticosteroid alone had hypogammaglobulinemia. Patients on oral PSL at a dose >12.5 mg/day for at least 1 year had low serum IgG. There was no significant correlation between the duration of oral PSL therapy and serum IgG. CONCLUSIONS: Oral PSL can potentially suppress humoral immunity in severe asthma. In asthmatics, hypogammaglobulinemia could develop in those on a daily dose of PSL >12.5 mg, but is independent of the duration of such treatment. No suppression of humoral immunity was noted on inhaled corticosteroid therapy alone, either at low or high dose. PMID- 12119509 TI - [Art therapy: development and evaluation of an observation instrument to systematically analyze patients' paintings from oncology and rheumatology]. AB - INTRODUCTION: Art therapies have been established methods in oncology for many years. To what extent patients' pictures can be systematically described, and what connections to various illnesses are possibly thereby revealed, has been discussed in various specialized medical fields for over a hundred years. The following paper first presents insight into the present stand of research and then presents the results of the study. QUESTION: Are systematic analyses of pictures possible with the use of the instrument we have developed? Are pictorial differences evident within the researched group? METHOD: A previously developed instrument was submitted for expert questioning for the purpose of the study. The instrument that had been modified on the basis of these results could then be used in the study to interpret 162 pictures. Four different raters, three of whom were acquainted with neither the patients nor the pictures, interpreted these pictures independently of one another. RESULTS: The evaluation of the expert interpretations shows that this newly developed instrument is suitable for the differentiated description of patients' pictures. First important indications were drawn in regard to the pictorial differences between the two examined groups of cancer patients and patients with chronic polyarthritis. CONCLUSION: Systematic picture analyses are possible. A further analysis of the expert interpretations is necessary to clarify questions raised by the pictorial differences found in the various diagnostic groups. PMID- 12119508 TI - Murine AIDS induces viremia and functional and phenotypic alterations in blood cells. AB - BACKGROUND: Murine acquired immunodeficiency syndrome (MAIDS) is characterized by generalized lymphoproliferation and progressive immunodeficiency. It is induced by a mixture of two replication-competent murine leukemia viruses (MuLV) and a disease-causing, replication-incompetent defective MuLV. Infection leads to specific phenotypic and functional alterations of lymphocytes in lymphoid organs. METHODS: We analyzed phenotypic, virological and functional parameters in the blood of mice infected with MAIDS virus. RESULTS: Disease progression correlated with increasing viremia, a loss of mitogen responsiveness of T lymphocytes, and the appearance of CD4+ Thy1- T lymphocytes. At >9 weeks after infection, the distribution of leukocyte cell populations became very heterogeneous, and late stage leukemic events were observed in 5 of 23 mice. CONCLUSIONS: Virus titers, mitogen responsiveness and the presence of CD4+ Thy1- T lymphocytes can efficiently be monitored in the blood and serve as diagnostic parameters to monitor disease progression. Acute leukemic events occurring at the terminal stage could be responsible for the death of at least some of the mice with MAIDS. PMID- 12119510 TI - [Cerebral hemodynamics in carbon dioxide applications]. AB - INTRODUCTION: We compare the effect of carbon dioxide (CO(2)) dry and wet applications on cerebral hemodynamics. METHODS: On 22 volunteers measurements were taken during CO(2) application. 10 probands were examined in CO(2) wet application (1,100-1,300 mg/l) and 12 probands in CO(2) dry application (500 g in a 800 l bathtub). The cerebral blood flow velocity (CBFV) in the middle cerebri artery (MCA) was measured as a parameter of cerebral hemodynamics by means of transcranial doppler sonography. Furthermore were recorded CO(2) expiratory concentration (CO(2)et), blood pressure, and sublingual temperature. RESULTS: At CO(2) wet application the CBFV increased during therapy phase by 15% (p = 0.001), parallel to the rise of the CO(2)et by 18% (p = 0.01). During CO(2) dry application CBFV decreased by 11% (p = 0.007), body temperature increased significantly by 0.2 degrees C. CONCLUSION: CO(2) applications have influence on cerebral hemodynamics. Assuming constant diameters of the great brain vessels, CO(2) wet application shows a raising and CO(2) dry application a reducing influence on cerebral blood flow. This influence will attain therapeutic relevance. PMID- 12119511 TI - Antiseptic effect of a topical dermatological formulation that contains Hamamelis distillate and urea. AB - OBJECTIVE: To determine the antimicrobial activity of a distillate of Hamamelis (Aqua Hamamelidis ), United States Pharmacopoeia (USP) 23, and urea formulated as a topical dermatological preparation that contains both active ingredients. METHODS: Using the simple occlusion test and expanded flora test, we conducted in vivo studies in 15 healthy volunteers. We also performed in vitro studies using the agar diffusion test. RESULTS: The occlusion test and expanded flora test demonstrated significant antimicrobial activity for a product containing the Hamamelis distillate (90%) and urea (5%) among other ingredients. The expanded flora test demonstrated significant antimicrobial activity for both Hamamelis distillate and urea. The simple occlusion test showed the same tendency, but results were not significant. The agar diffusion test showed inhibition of Staphylococcus aureus and Candida albicans, among other organisms. Comparison with earlier studies of chlorhexidine digluconate and fuchsine using the same method showed that the antimicrobial activities of Hamamelis distillate and urea were relatively weak. This finding is supported by the weak inhibitory activity observed in the agar diffusion test (using 100% of the finished dosage form). CONCLUSION: Formulations of Hamamelis distillate and urea are mainly used for their antiinflammatory, hydrating, and barrier-stabilizing effects in dermatitis maintenance therapy. As bacterial colonization has a central role in the pathogenesis of atopic dermatitis and intertrigo, the antimicrobial activity of such products is considered a welcome, added benefit. PMID- 12119512 TI - [Blood and tissue eosinophilia, mistletoe lectin antibodies and quality of life in a breast cancer patient undergoing intratumoral and subcutaneous mistletoe therapy]. AB - BACKGROUND: Mistletoe therapy (MT) is a method of complementary medicine whose efficacy is controversially discussed. Until now there is a lack of data of high dose intratumoral application. PATIENT AND METHODS: We are presenting a 3-year follow-up of an 80-year-old woman with metastasized breast cancer (c(2)T3/N1/M1) receiving combined intra- and peritumoral and subcutaneous MT with ABNOBAviscum(r) and concomitant pamitron acid. At time of admission the patient had bone metastases (thoracic vertebra 11 fracture), a lymphangiosis carcinomatosa, bilateral pleural effusions, and a reduced quality of life (QoL). RESULTS: Under MT we induced an eosinophilia and an elevation of eosinophil cationic protein (ECP). Simultaneously, we ascertained a reduction of 50% of Ca 15-3 and a sustained partial tumor remission. After 5 months the mistletoe-lectin 1 antibodies IgG-1 and -3 were maximally increased and we perceived a second Ca 15-3 reduction. After 3 months we observed a benefit in QoL. During the following 5 months the patient gained about 10 kg in weight. In the second year slow tumor progress was observed. After 19 months the patient had pneumonia which caused an MT pause. Subsequently, a combined intratumoral mistletoe and letrozol therapy brought a partial remission. The patient lived without chemo and radiation therapy more than 3 years with good QoL and died after 41 months, after a sepsis and a following stroke without signs of tumor progress. CONCLUSION: In metastasized breast cancer a palliative high-dose local MT can make a contribution to a tumor reduction and a benefit in QoL. PMID- 12119513 TI - Efficacy of distant healing--a proposal for a four-armed randomized study (EUHEALS). AB - BACKGROUND: Distant healing as a treatment modality is frequently used by patients and healers. Some preliminary evidence suggests possible effects. Since patients suffering from multiple chemical sensitivity and chronic fatigue syndrome have only few effective treatment options, distant healing will be offered as a treatment within a formal trial of distant healing. DESIGN AND METHOD: A four-armed randomized trial will include 400 patients with self attributed, environmental problems who fulfil the diagnostic criteria of severe idiopathic chronic fatigue, chronic fatigue syndrome or multiple chemical sensitivity. Patients will be recruited by specialized general practitioners and environmental clinics. They will be treated by healers distributed all over Europe, coming from various healing traditions and nationalities. Each patient will be treated by 3 healers. Healers will have no contact with the patients and will only be provided with the patient's Christian name and a photograph. The patients will be randomized to one of 4 groups in a 2 x 2 factorial design. They will either receive (distant) healing or not, and either know or not know this decision. Thereby the effects of expectation and of time can be disentangled from the specific effects of healing. OUTCOME MEASURE: Primary outcome measure will be the mental health summary scale of the MOS SF-36. The measure will be taken at the beginning and at the end of a 6- month treating or waiting period, respectively. A variety of moderator variables will be considered to evaluate which of these may be predictive of outcome. PMID- 12119516 TI - Bilio-digestive double bypass for nonresectable pancreatic cancer. AB - In spite of extensive preoperative investigation, surgical exploration is often the only way to determine whether a pancreatic cancer is curatively resectable. If curative resection is not possible, palliation of cholestasis and eventual duodenal obstruction is mandatory. This is best achieved by construction of a bilio-digestive double bypass. Many different techniques have been described but considerable rates of delayed gastric emptying have added high morbidity to the procedure. We propose a retrocolic construction technique combining an omega loop with a Roux-en-Y reconstruction which to our knowledge has not been published before. PMID- 12119515 TI - Pyloric drainage (pyloroplasty) or no drainage in gastric reconstruction after esophagectomy: a meta-analysis of randomized controlled trials. AB - BACKGROUND/AIM: A gastric conduit is usually used to reconstruct the foregut after esophagectomy for cancer. The gastric emptying may be impaired after this operation, so some esophageal surgeons routinely add a pyloric drainage procedure (pyloroplasty or pyloromyotomy). We performed a meta-analysis of randomized controlled trials (RCTs) to determine the effect of pyloric drainage on patient outcomes. METHODS: Medline and manual searches were done (completed independently and in duplicate) to identify all published RCTs that addressed the issue of pyloric drainage procedures during gastric conduit reconstruction of the esophagus. The selection process was inclusive; no trials were excluded. Trial validity assessment was done, and a trial quality score was assigned. Early outcomes assessed by meta-analysis included operative mortality, esophagogastric anastomotic leaks, pulmonary morbidity, pyloric drainage complications, fatal pulmonary aspiration, and gastric outlet obstruction. A random-effects model was used, and the relative risk was the principal measure of effect. Systematic semiquantitative review was used for late outcomes such as gastric emptying, bile reflux, nutritional status, and obstructive foregut symptoms. RESULTS: Nine RCTs, that included a total of 553 patients, were selected, with quality scores ranging from 1 to 4 (5-point Jadad scale). Selection and validity agreement was strong. The relative risk (95% CI; p value), expressed as pyloric drainage versus no drainage (treatment vs. control), was 0.92 (0.34, 2.44; p = 0.86) for operative mortality, 0.90 (0.47, 1.76; p = 0.77) for esophagogastric anastomotic leaks, 0.69 (0.42, 1.14; p = 0.15) for pulmonary morbidity, 2.55 (0.34, 18.98; p = 0.36) for pyloric drainage complications, 0.25 (0.04, 1.60; p = 0.14) for fatal pulmonary aspiration, and 0.18 (0.03, 0.97; p = 0.046) for gastric outlet obstruction. Systematic semiquantitative review showed a nonsignificant trend favoring pyloric drainage for the late outcomes of gastric emptying, nutritional status, and obstructive foregut symptoms. For the late outcome of bile reflux, there was a nonsignificant trend favoring the no-drainage group. The scintographic gastric emptying time, expressed as a ratio (pyloric drainage/no drainage), was 0.53. CONCLUSIONS: Data synthesized from existing RCTs show that pyloric drainage procedures reduce the occurrence of early postoperative gastric outlet obstruction after esophagectomy with gastric reconstruction, but they have little effect on other early and late patient outcomes. PMID- 12119517 TI - Aneurysm of the superior mesenteric artery. PMID- 12119518 TI - Laparoscopic wedge resection of gastric submucosal tumors. AB - BACKGROUND/AIMS: The purpose of this study was to evaluate the clinical utility of laparoscopic surgery for gastric submucosal tumor. METHODS: The records of 11 patients who underwent laparoscopic wedge resection (LR group) for gastric submucosal tumors were reviewed and compared with those of 8 patients who underwent open surgery (OS group). RESULTS: Mean operation time was 145 +/- 43 min in the LR group and 127 +/- 33 min in the OS group (p = 0.301). Mean blood loss was 97 +/- 107 and 107 +/- 47 g, respectively (p = 0.387). Patients in the LR group began walking 1.4 +/- 0.7 days after surgery, which was significantly earlier than those in the OS group (2.7 +/- 1.3 days, p = 0.021). The first flatus (1.5 +/- 0.5 vs. 3.1 +/- 0.6 days, respectively, p = 0.0004) and resumption of oral food intake (3.0 +/- 1.7 vs. 4.3 +/- 0.9 days, respectively, p = 0.020) were also earlier in the LR group. White blood cell count on the first postoperative day was lower (7,000 +/- 2,100 vs. 11,900 +/- 3,580/mm(3), respectively, p = 0.004) in the LR group than in the OS group, and the duration of fever (>38.0 degrees C; 0.1 +/- 0.3 vs. 0.9 +/- 0.8 days, respectively, p = 0.014) and the period of postoperative hospitalization (13.2 +/- 3.7 vs. 20.8 +/- 6.1 days, respectively, p = 0.014) were significantly shorter in the LR group than in the OS group. No complications occurred in either group. CONCLUSION: Laparoscopic surgery was superior to open surgery in terms of postoperative recovery time with comparable operation time and blood loss. Laparoscopic wedge resection is a promising surgical alternative for the treatment of gastric submucosal tumors. PMID- 12119520 TI - Morphology, viability and functions of suckling pig hepatocytes cultured in serum free medium at high density. AB - BACKGROUND: In bioartificial liver preparation, serum-contained medium is ordinarily replaced by serum-free medium and hepatocytes are generally cultured at high density. This study was to undertaken to evaluated the dynamic changes in morphology, viability and functions of porcine hepatocytes in serum-free medium at high density. METHODS: Hepatocytes were isolated from suckling pigs by modified two-step in situ collagenase perfusion method and cultured in serum-free medium at high density. Morphology, viability, protein and glucose syntheses, G-6 Pase activity, diazepam transformation of hepatocytes and release of LDH in supernatant during 7 days of culture were evaluated. These measurements were also determined on both groups of hepatocytes cultured at low-density in serum-free medium and serum-contained medium, which served as control groups. RESULTS: Morphology and protein synthesis of hepatocytes cultured in serum-free medium at high density were stable over the course of 7 days. High viability (>90%) was obtained though it declined with time. Diazepam transformation of cells was higher on days 2 and 3. Glucose synthesis of cells declined from day 3 to day 7. G-6-Pase activity of the hepatocytes declined apparently after 1 day of culture and it was maintained at a low level from day 1 to day 7. Release of LDH in supernatant was higher on days 1, 2 and 3. There were no significant differences in viability and functions of hepatocytes except for G-6-Pase activity at low density culture between the serum-free medium group and the serum-contained medium group. The functions of hepatocytes cultured at high density were lower than at low-density culture. CONCLUSIONS: The results showed that the morphology, viability, protein synthesis and diazepam transformation of hepatocytes cultured in serum-free medium at high density were maintained during 7 days of culture. The serum-free medium provided indices of cell viability and functions that were comparable to serum-contained medium. The functions of hepatocyte cultured at high density (1 x 10(7) cells/ml) were lower than at low density (5 x 10(5) cells/ml). PMID- 12119521 TI - Non-compliance with national guidelines in the management of acute pancreatitis in the United kingdom. AB - BACKGROUND: Deficiencies and lack of standardisation of the management of acute pancreatitis in the UK have been reported. National UK guidelines for the management of acute pancreatitis were published in 1998. However, implementation of national guidelines in other areas has been patchy, suggesting that evaluation of the uptake of the pancreatitis guidelines would be appropriate. AIM: Identification of current practice in the management of acute pancreatitis as reported by consultant surgeons, in order to determine how effectively the UK guidelines have been introduced into practice. METHODS: A questionnaire was posted to 1,072 full members of the Association of Surgeons of Great Britain and Ireland. It consisted of 13 questions that aimed to identify the surgeon's practice in the management of patients with acute pancreatitis in relation to key points in the UK guidelines. We compared the practice of hepatobiliary and pancreatic (HBP) vs. non-HBP specialists, and teaching vs. non-teaching hospital surgeons using the chi(2) test. RESULTS: Of 538 responses (50%), 519 were from consultant surgeons. 59 did not look after patients with acute pancreatitis and 89 (17%) had a HBP interest. There were differences between the recommendations in the guidelines and reported practice, particularly in the use of critical care resources and referral to specialist units. Of consultants looking after acute pancreatitis 371 (72%) were non-HBP specialists. There were significant overall differences between the practice of HBP specialists and non-specialists: in severity assessment (Glasgow and C-reactive protein vs. Ranson criteria); indication and timing of requesting computed tomography (routinely at 7-10 days vs. when clinically indicated); nutritional support (enteral feeding vs. no support), and in common bile duct assessment prior to cholecystectomy (intra operative cholangiography vs. endoscopic retrograde cholangiopancreatography). There was no significant difference between practice in teaching and non-teaching hospitals. CONCLUSION: Implementation of national guidelines for the management of acute pancreatitis was greater in the practice of HBP specialists than non specialists. This has implications for the rationale of creating guidelines, and for the strategies associated with their introduction. PMID- 12119519 TI - Manganese deposition in basal ganglia due to perioperative parenteral nutrition following gastrointestinal surgeries. AB - BACKGROUND/AIMS: Serial changes in blood manganese (Mn) levels and brain MRI examinations following perioperative parenteral nutrition (PN) were investigated. METHODS: Six cases undergoing pancreatoduodenectomy (PD), 4 cases undergoing thoracic esophagectomy (TE), 18 cases undergoing total gastrectomy (TG) and 20 cases undergoing colorectal surgeries (CR) with daily administration of 20 micromol of Mn per day were studied. Cases undergoing PD, TE, TG or CR without Mn administration served as controls. RESULTS: Hyperintense lesions in the basal ganglia on T1-weighted MRI and elevated blood Mn levels were recognized after PN in 4 of 6 cases in the PD group, in 3 of 4 cases in the TE group, in 1 of 18 cases in the TG group and in 2 of 20 cases in the CR group. No abnormalities were recognized in the control groups. CONCLUSION: The possible Mn deposition in the basal ganglia caused by perioperative PN should be especially noted in cases undergoing PD or TE. PMID- 12119522 TI - Isolated Roux Loop duct-to-mucosa pancreaticojejunostomy avoids pancreatic leaks in pancreaticoduodenectomy. AB - BACKGROUND: Over the last decade the operative mortality associated with pancreaticoduodenectomy (PD) has decreased. Pancreatic anastomotic leaks resulting in pancreatic bed sepsis and fistulae, however, remain a significant cause of both morbidity and mortality. The optimal method of reconstruction to minimise pancreatic leaks is controversial. AIM: To review the experience of Roux loop duct-to-mucosa pancreaticojejunostomy in a consecutive series of patients undergoing pancreatic head resection. METHODS: Over the 6-year period (1993 1998), 41 patients underwent pancreatic head resections for benign (n = 5) and malignant disease (n = 36). There were 19 males and the median age was 62 years (range 29-83). An isolated Roux loop pancreaticojejunostomy was performed in all cases. RESULTS: Median duration of surgery was 8 h and the median postoperative stay was 16 days. The mean peri-operative blood transfusion was 2.9 units (SD 1.9). The incidence of major complications was 12% and there was 1 death (2.4%). There were no pancreatic leaks or fistulae. CONCLUSIONS: The low complication rate and the absence of pancreatic fistulae in this series would suggest that Roux loop duct-to-mucosa pancreatic reconstruction should be more widely adopted. PMID- 12119523 TI - The implications of subspecialisation on the management and outcome of surgery for rectal carcinoma. AB - BACKGROUND: The management of rectal carcinoma has changed significantly over the last decade. We studied the changing trends in the management of rectal carcinoma over a 7-year period in a district general hospital. METHODS: A retrospective analysis of all patients with histologically proven rectal adenocarcinoma who underwent operative treatment between January 1991 and December 1997 was performed. The type of operative procedure, local recurrence rate and completeness of pathology reporting was documented. RESULTS: There were 200 operative procedures: 102 anterior resections (AR), and 98 abdominoperineal resections (APR). This included 17 palliative resections because of metastatic disease (n = 8) or extensive local invasion (n = 7) or both (n = 2). The APR rate steadily declined from 72% in 1991 to 19% in 1997 (p < 0.005). Subspecialist 'colorectal' surgeons performed only 24% of the operations in 1991 but the figure for 1997 was 85% (p < 0.01). No circumferential resection margin was reported in 1991 but was reported in 85% of the cases in 1997 (p < 0.001). There was a steady increase in stapled anastomoses from 43% in 1991 to 93% in 1997 (p < 0.03). There were 15 local recurrences following 'curative' resection; 7 following APR and 8 following AR (n.s.). CONCLUSION: There was a significant increase in the rate of restorative resection of rectal cancer with a concomitant reduction in permanent stoma formation; this may be attributed to an increase in subspecialisation. Despite this, a low rate of local recurrence was maintained throughout the study period. PMID- 12119524 TI - Systemic interleukin-6 response to colorectal surgery originates from the bowel. AB - BACKGROUND: Surgical trauma evokes a systemic cytokine response which is enhanced in patients with colorectal cancer. The aim of this study was to locate the origin of the systemic cytokine response to colorectal surgery. METHODS: The concentrations of interleukin-6 (IL-6) and tumour necrosis factor-alpha (TNF alpha) were analysed in systemic and mesenteric venous blood in 12 patients operated on with colorectal resections due to cancer or benign lesions. Immunohistochemical staining and analysis of tissue concentrations of IL-6 and TNF-alpha in homogenates from tumours and benign specimens were performed. RESULTS: Mesenteric venous blood contained higher concentrations of IL-6 compared to systemic venous blood after resection, but not before. Tissue concentration of IL-6 was higher in the tumours compared to the benign specimens and immunohistochemistry revealed an abundance of IL-6 and TNF-alpha in malignant epithelium compared to benign mucosa. CONCLUSION: The higher concentration of IL 6 in venous blood from the mesenterium of the resected colonic segment compared to systemic levels, indicates that the bowel is the source of the IL-6 response to surgical trauma in colorectal surgery. PMID- 12119525 TI - The need for interval appendectomy after resolution of an appendiceal mass questioned. AB - BACKGROUND: Our current treatment of an appendiceal mass is initially conservative, followed by an interval appendectomy. The necessity of this routine interval appendectomy is debatable. A study was conducted to evaluate whether surgical factors and pathological features of the excised appendices support interval appendectomy. METHODS: We performed a retrospective study at the University Hospital Groningen and the Deventer Ziekenhuis. All patients diagnosed with an appendiceal mass in the period January 1991 to January 1997 were identified using the hospital database. The medical records of all these patients (n = 233, 108 M, 125 F) were reviewed. The clinical course of the appendiceal mass patients was split up into three distinct episodes: initial diagnosis and treatment of the appendiceal mass, the interval period and the interval appendectomy. Presenting symptoms, findings at clinical examination and additional imaging (ultrasound) were registered, as well as the course of the primary hospitalisation, the interval period, and the interval appendectomy. Results of histological examination of all resected specimens were reviewed. RESULTS: It was found that clinical findings alone were not specific enough to diagnose an appendiceal mass; 47% had a palpable abdominal mass and the median temperature was 38.2 degrees C ranging from 36 to 40.5 degrees C. Ultrasound examination was done in 69% of patients and showed an appendiceal mass in 72%. During the interval period, 4 patients presented with an appendiceal mass needing drainage, and 3 with acute appendicitis requiring emergency appendectomy. At interval appendectomy, histological examination of resection specimen showed a normal appendix without signs of previous inflammation in 30% of cases. In addition, complications due to interval appendectomy were seen in 18% of patients, including sepsis, bowel perforation, small bowel ileus, and various wound abscesses. CONCLUSIONS: We conclude that when causes for the appendiceal mass other than appendicitis are excluded, interval appendectomy seems unnecessary in patients who respond well to initial conservative treatment. PMID- 12119526 TI - Preoperative colonoscopic derotation is beneficial in acute colonic volvulus. AB - AIMS: Analysis of preoperative and operative management of acute colonic volvulus and development of treatment guidelines in a region of low incidence. METHODS: A study of 42 consecutive patients operated for acute colonic volvulus between 1970 and 2000. RESULTS: There were 20 patients with sigmoid volvulus, 21 with cecal volvulus and 1 with volvulus of the transverse colon. All patients presented as emergencies. The correct preoperative diagnosis was possible for sigmoid volvulus in 95% (19/20) of cases and for cecal volvulus in 67% (14/21). Preoperative colonoscopic volvulus derotation was attempted in 19 patients and successfully completed in 9 patients (47%). The success rates for preoperative colonoscopic derotation were 58 (7/12) and 33% (2/6) for sigmoid and cecal volvulus, respectively. Thirty-four patients (81%) underwent colon resections, 26 times as a single-stage procedure, and 8 patients (19%) underwent non-resectional operative techniques. Overall surgical morbidity was 24%, the reoperation rate 9.5% and mortality 12% (5/42). The subgroup of 9 patients with successful non operative volvulus derotation, however, underwent semi-elective single-stage colonic resection without surgical morbidity or mortality. There were no recurrences during a median follow-up period of 9.5 years. CONCLUSION: In the absence of clinical, laboratory or radiological signs of bowel necrosis or perforation, colonoscopic volvulus derotation is recommended in all cases of acute colonic volvulus, followed by semi-elective single-stage colonic resection. PMID- 12119527 TI - Therapeutic fistuloscopy: an alternative approach in the management of postoperative fistulas. AB - BACKGROUND/AIM: Since the treatment of postoperative fistulas remains a difficult problem, we applied endoscopic treatment in such 14 persistent fistulas. METHODS: Fourteen patients presented with postoperative fistulas: 7 patients (low-output group) due to residual cavity after liver hydatid disease surgery and 7 patients (high-output group) after small-bowel resection (n = 3), diverted duodenostomy (n = 1), vertical gastroplasty (n = 1), external pancreatic cyst drainage (n = 1), and transduodenal sphincteroplasty (n = 1). The therapeutic procedures included mechanical removal of silk sutures, necrotic material, and hydatid membranes in the low-output group and fibrin sealing in the high-output group. RESULTS: Fistuloscopy was performed 170-278 days (mean +/- SD 198.7 +/- 36.7 days) and 18 51 days (mean +/- SD 34.0 +/- 11.3 days) postoperatively in low- and high-output fistula patients, respectively, when the average daily output was 20-50 (32.8 +/- 12.5) ml and 200-1,000 (563.1 +/- 319.4) ml, respectively. The low-output group needed only one fistuloscopy session, while the other group required a median number of three sessions plus fibrin sealing, the total amount of fibrin glue used per patient being 2-14 (6.5 +/- 4.4) ml. No procedure-related complication occurred. All fistulas except one healed within 10-33 (21.8 +/- 7.9) days and 2 17 (9.2 +/- 5.1) days in low- and high-output groups, respectively. CONCLUSIONS: We believe fistuloscopy to be a useful tool in the management of gastrointestinal fistulas, but more experience should be gained in using this technique. PMID- 12119528 TI - Gastroduodenal artery stump haemorrhage following pylorus-sparing Whipple procedure: treatment with covered stents. AB - BACKGROUND/AIMS: To report a case of bleeding after pancreatoduodenectomy in a patient with pancreatic leak and portal thrombosis who was successfully treated with an endovascular approach. METHODS: A 58-year-old male, suffering from neoplasm of the distal bile duct, underwent a pylorus-preserving Whipple procedure. On the 18th day, following a sudden drop in pressure and low haematocrit values, the patient underwent surgery. The source of the bleeding was not found. Six days later, following the appearance of bleeding from the abdominal drainage and haematemesis with shock, the patient had an immediate angiography. Bleeding from the gastroduodenal artery stump was evident, the portography showed no portal flow. With respect to the shortness of the stump, safe embolisation with coils, while preserving the common hepatic artery patency, was difficult to obtain. RESULTS: By transcatheter placement of covered stents into the hepatic artery and thereby occluding the origin of the gastroduodenal artery, the bleeding was stopped. After 2 months, CT angiography showed patency of both the common and proper hepatic arteries. Nine months after the procedure the patient is in good health. CONCLUSIONS: Percutaneous placement of covered stents can be the solution in cases where transcatheter embolisation is not recommendable because of portal vein thrombosis. PMID- 12119529 TI - Two unusual cases of adult intussusception. AB - Adult intussusception is very rare. We report 2 unusual cases, a 58-year-old man with a transverse colo-colonic intussusception caused by a malignant sessile polyp that also had an asymptomatic synchronous neoplasm of the kidney, and an 18 year-old female with an ileocecolic intussusception caused by acute appendicitis. This report stresses the point that intussusception in adults may represent an underlying malignancy. The age of the patient and the anatomic location of the intussusception provide significant input as to the etiology and hence the most appropriate surgical procedure. PMID- 12119530 TI - The cerebellum at birth in therian mammals, with special reference to rodents. AB - The stage of cerebellar development at birth was assessed in 23 species of placental mammals. Serial histological sections were examined and five stages in the differentiation of the cerebellar cortical layers were defined. A wide diversity of conditions at birth was found. The available evidence (after parsimony reconstruction) suggests that the last common ancestor of placentals was born with an altricial cerebellum in which the molecular layer was just present between the external granular layer and the prospective Purkinje cell layer. Some placental species have an even more altricial cerebellum at birth (e.g., Muscardinus avellanarius, Sorex araneus), with Mesocricetus auratus as the most altricial species among the taxa studied. In the newborn M. auratus a cerebellar anlage was present with only a loose accumulation of cells located at the dorsal cerebellar anlage above the ventricular neuroepithelial layer. The five species of caviomorph rodents examined here are relatively precocial as far as the cerebellum is concerned. The only other rodent species that has a similarly advanced state was the murid Acomys sp. Most of the life history variables examined were not strongly correlated with the cerebellar stage at birth if at all. However, a significant positive correlation (r(2) = 0.67) was observed between the cerebellar stage at birth and the gestation length and a significant negative correlation (r(2) = 0.31) was observed between cerebellar stage and the average litter size. The weak correlation may be due to sampling among different distantly related clades. The most mature cerebella at birth still had an external granular layer, indicating that the mossy fiber-granule cell connectivity is not yet fully developed and further indicating that this connectivity may depend on external experience to fully mature. All species that have their eyes open at birth also have the most mature cerebelli. The growth of the cortical layers was also studied in a postnatal ontogenetic series of the marsupial Monodelphis domestica. As is the case with placentals, the most advanced stage of cerebellar development coincides with the opening of the eyes. PMID- 12119531 TI - Connectional evidence for dorsal and ventral V3, and other extrastriate areas in the prosimian primate, Galago garnetti. AB - Previously we described patterns of connections that support the concept of V3 in small New World marmoset monkeys, three species of larger New World monkeys, and two species of Old World macaque monkeys. Here we describe a pattern of V1 connections with extrastriate visual cortex in Galago garnetti (also known as Otolemur garnetti) that demonstrates the existence of a V3 in a strepsirhine (prosimian) primate. Injections of fluorochromes or cholera toxin subunit-B (CTB) in V1 labeled cells and terminals in retinotopically matched regions in V2, V3, DL (V4), and MT. Labeled axon terminations were more focused primarily in middle layers of cortex, likely representing 'feedforward' input from V1, whereas labeled cells were more widespread and found in both superficial and deeper cortical layers, indicative of feedback projections. Averaged across injections, V3 had the third largest percentage of labeled cells (11%), following only V2 (47%) and the middle temporal area (MT; 19%). The dorsolateral area (DL, or V4; 9%) also contained a relatively large number of retrogradely labeled cells. These results indicate that V2, V3, DL (V4), and MT are retinotopically connected with V1, and provide major sources of feedback. Other extrastriate areas were less densely connected to V1, and there was no clear indication of labeled terminals. Inferotemporal cortex (IT) provided nearly 7% of feedback connections, whereas the dorsomedial area (DM) contributed about 3%. The remaining areas that have been proposed for galago extrastriate cortex, MTc, MST, FST, LPP and VPP, each accounted for about 1% or less of the total number of labeled cells. Thus, six extrastriate areas, V2, MT, V3, DL (V4), IT, and DM provide over 96% of visual cortex projections to V1. These areas also provide most of the projections to V1 in New and Old World monkeys. PMID- 12119532 TI - Ontogenetic changes in the response properties of the peripheral electrosensory system in the Atlantic stingray (Dasyatis sabina). AB - Adult stingrays use their ampullary electroreceptors to detect prey and locate mates, but the response properties and function of their electrosensory system in the pre-adult stages are unknown. We examined the response properties of Atlantic stingray (Dasyatis sabina) electrosensory primary afferent neurons through ontogeny to determine whether encoding of electrosensory information changes with age, and how it relates to the ontogenetic encoding of biologically relevant electric stimuli. We show that during development electrosensory primary afferents increase resting discharge regularity, show an upward shift in best frequency (BF), an increase in neural sensitivity, and a decrease in bandpass. These ontogenetic changes in the response properties of the stingray electrosense are consistent with sensory adaptations to enhance the avoidance of large predators as young, and increase the location of prey and mates as adults. PMID- 12119533 TI - Tactile hairs on the postcranial body in Florida manatees: a Mammalian lateral line? AB - Previous reports have suggested that the sparsely distributed hairs found on the entire postcranial body of sirenians are all sinus type tactile hairs. This would represent a unique arrangement because no other mammal has been reported to possess tactile hairs except on restricted regions of the body, primarily the face. In order to investigate this issue further, hair counts were made systematically in three Florida manatees (Trichechus manatus latirostris), and hair follicle microanatomy was studied in 110 specimens gathered from 9 animals. We found that the postcranial body possesses approximately 1500 hairs per side, and hair density decreases from dorsal to ventral. External hair length ranged from 2-9 mm, and most hairs were separated from their nearest neighbor by 20-40 mm, resulting in an independent domain of movement for each hair. All hairs exhibited the anatomical characteristics of follicle-sinus complexes typical of tactile hairs, including a dense connective tissue capsule containing an elongated circumferential blood sinus and innervation by 20-50 axons which ascend the mesenchymal sheath. We conclude that this represents a unique distributed underwater tactile system capable of conveying detailed and significant external information concerning approaching animals, water currents and possibly the presence of large stationary features of the environment. Such a system would be analogous to the lateral line in fish, and would be particularly useful in the turbid habitat frequented by Florida manatees. PMID- 12119534 TI - Use of near infrared spectroscopy for estimation of peripheral venous saturation in newborns: comparison with co-oximetry of central venous blood. AB - In the intensive care of sick infants, the global oxygen reserve capacity is estimated by co-oximetry (co-ox) of blood sampled from central venous catheters. Introduction of a noninvasive alternative is desirable. Near infrared spectroscopy (NIRS) offers a technique for noninvasive bedside monitoring of tissue oxygen economy. We studied the relation between peripheral venous oxyhemoglobin saturation (SvO(2)) estimated by venous occlusion and NIRS, and the central SvO(2) measured by co-ox of central venous blood. We report the high reproducibility of NIRS with a test-retest variation of only 2.51 +/- 1.41%. After bias adjusting of NIRS SvO(2) values, a nice correlation (r = 0.96, p > 0.05) between NIRS measurements of peripheral SvO(2) and co-ox of central venous blood was found. The study indicates that NIRS is practical for monitoring relative changes in central venous saturation. This might be useful in the future clinical care of newborns. PMID- 12119535 TI - Vasomotor reactivity in premature neonates at term. AB - In order to assess the specific sympathetic reactivity in premature infants at term, we designed a study to evaluate the peripheral vasomotor response of such infants when exposed to auditory challenges. Testing was performed in 29 premature neonates at term in both quiet and active sleep during a morning session. Two types of noises were used (click and continuous tones) at three frequencies (250, 1,000 and 6,000 Hz) and at three intensities (60, 85 and 110 dBA). Vasomotor response was studied by analyzing with Mathlab software the variability of the plethysmographic wave of the oxymetric pulse. No behavioral awakening was observed in response to any stimulation. When a tachycardia or a bradycardia reaction to the stimuli was observed, all neonates responded with a vasoconstriction. The global mean of the vasoconstrictive response was 18.45%. The overall ANOVA on the vasomotor response revealed significant effects for sleep stages (t: 1.98; p < 0.05), for frequency (t: 3.3; p < 0.001) and for intensity of noise (t: 3.01; p < 0.03) but no significant response with heart rate variability. From these results, we could conclude that the assessment of the vasomotor response is a very sensitive procedure to determine the reactivity of the autonomic nervous system in neonates, and could be used to study such vegetative responses in other stressful situations with good accuracy. PMID- 12119536 TI - Urinary aquaporin-2 excretion in preterm and full-term neonates. AB - The study was undertaken to define the role of aquaporin-2 (AQP2) in renal concentrating performance by measuring urinary AQP2 excretion and urine osmolality in healthy preterm and full-term neonates during early postnatal life. Random urine samples were obtained from 9 full-term newborn infants (mean birth weight 3,218 g, mean gestational age 39.2 weeks) at postnatal ages of 1, 3 and 5 days. Five premature infants with a mean birth weight of 1,570 g and mean gestational age of 30.6 weeks were studied at the end of the 1st week and then weekly up to the 6th week. Urine osmolality (Knauer osmometer), creatinine (modified Jaffe's method) and AQP2 concentrations (radioimmunoassay) were measured. In full-term neonates, urinary AQP2 excretion showed no consistent changes over the age period studied, while urine osmolality decreased significantly with advancing age. In premature infants, urinary AQP2 excretion remained practically unchanged during the first 4 weeks followed by an abrupt increase thereafter. Urine osmolality did not follow the developmental pattern of AQP2 excretion; its mean values varied only from 78 +/- 39 to 174 +/- 146 mosm/l during the experimental period. It is concluded that during the early postnatal period, urinary AQP2 excretion does not serve as a direct marker of the renal action of AVP and the renal capacity to concentrate urine. PMID- 12119537 TI - Induction of human macrophage vascular endothelial growth factor and intercellular adhesion molecule-1 by Ureaplasma urealyticum and downregulation by steroids. AB - Chronic lung disease (CLD) remains a major cause of morbidity for the prematurely born infant. The pathogenesis of CLD is complex and has not been defined entirely. Infection and lung inflammatory events have been thought to play a key role in the development of CLD. However, the contribution of Ureaplasma urealyticum to the development of CLD is debated and steroids produce some improvement in neonates with this disease. The aim of this study was to investigate if U. urealyticum could stimulate macrophages to produce vascular endothelial growth factor (VEGF) and intercellular adhesion molecule-1 (ICAM-1) in vitro, which are potentially associated with both early and later pathological changes in the lung during the development of CLD. In addition, the impact of dexamethasone and budesonide on these processes was examined. We found that U. urealyticum antigen (>/=4 x 10(7) color-changing units/ml) stimulated human macrophages (phorbol 12-myristate 13-acetate-differentiated THP-1 cell line) to produce VEGF and soluble ICAM-1 in a dose-dependent manner (p < 0.05) measured by ELISA. Likewise, cell surface ICAM-1 (CD54) measured by flow cytometry was increased after stimulation with U. urealyticum. This effect was attenuated by budesonide and dexamethasone (p < 0.05). The mRNA expressions of VEGF and ICAM-1 detected by a semi-quantitative reverse transcriptase polymerase chain reaction were also induced in response to U. urealyticum and inhibited by the steroids (p < 0.05). The expression of ICAM-1 was reduced by 85.5% when the TNF-alpha production was neutralized with an anti-TNF-alpha antibody. Our findings imply that U. urealyticum might be involved in the development of CLD of prematurity. PMID- 12119538 TI - Decreased phagocytic capacity of cord blood monocytes in second- and third-born multiplets. AB - OBJECTIVE: To assess the risk to second- and third-borns of developing perinatal infections based on a diminished first-line immune response in comparison to the first-born. STUDY DESIGN: 11 sets of twins and 2 sets of triplets (27-39 weeks of gestation, 740-3,560 g birth weight) admitted consecutively to our neonatal care unit were investigated. All were delivered by cesarean section. Phagocytosis and the expression of TNF-alpha and IL-1 in cord blood were determined immediately and 72 h after birth from peripheral blood monocytes by FACS analysis and ELISA. RESULTS: In all second- and third-borns, phagocytosis of cord blood monocytes was significantly decreased in comparison to the first-borns (p < 0.001, p < 0.015). 72 h after birth phagocytosis determined from peripheral blood showed no significant difference between the first-borns and the second- and third-borns. Cytokine expression revealed no significant differences dependent on birth order. CONCLUSION: We suggest that the observed temporarily diminished first-line immune response of second- and third-born multiplets might be the result of a short-term reduced cellular oxygenation during delivery, but has no influence on further immunological deficiency leading to a higher risk of perinatal infections. PMID- 12119539 TI - Interleukin 8 in the human colostrum. AB - We investigated the postpartum changes in colostral interleukin (IL) 8 concentrations during the first 3 postpartum days and examined the IL-8 production of colostral cells at the levels of both protein and mRNA. Colostrum samples were obtained from healthy mothers after full-term delivery. Colostrum, supernatants, and cell lysates of cultured colostral cells were assayed for IL-8 by enzyme-linked immunosorbent assay. Reverse-transcription polymerase chain reaction was used to test uncultured colostral cells for the production of IL-8 mRNA. The colostral IL-8 concentrations, especially high on day 1 postpartum, declined abruptly during the first 3 postpartum days (mean 19.7, 10.0, and 3.0 ng/ml on days 1, 2 and 3 postpartum, respectively). Colostral cells apparently produced and secreted IL-8 in in vitro culture without stimulant, although in smaller quantities than with lipopolysaccharide stimulation. The majority of uncultured colostral cell samples expressed IL-8 mRNA, suggesting a role of colostral cells, at least in part, as a source of colostral IL-8. PMID- 12119540 TI - Effect of glucose and fatty acid availability on neonatal and adult heart contractility. AB - Changes in contractility of newborn and adult hearts with different substrates were studied under oxygenation and hypoxia. Under oxygenation, insulin (50 mU/l) and low doses of fatty acids (sodium palmitate 0.12 mM) increased the differential ventricular pressure (DVP), while higher doses of fatty acids (from 0.3 to 1.5 mM) decreased it. However, when both low doses of fatty acids and insulin were added simultaneously, tension development decreased. Hypoxia reduced DVP, and low doses of fatty acids restored cardiac force. The contractile response to extracellular glucose concentrations changed during development, and sensitivity to high doses of fatty acids increased with age. In adult cardiomyocytes, glucose uptake was also inhibited by sodium palmitate under oxygenation when cardiac metabolism is fatty acid dependent, but not under hypoxia, when it consumes carbohydrates. Newborn cardiomyocytes consumed more glucose than adult cells, they did not respond to insulin, and palmitate did not completely inhibit glucose uptake neither under oxygenation nor under hypoxia. Substrate availability modified glucose uptake and contractility by independent mechanisms. PMID- 12119541 TI - NMDA receptor modification during graded hypoxia in the cerebral cortex of newborn piglets. AB - Previous studies have shown that the cerebral N-methyl-D-aspartate (NMDA) receptor is altered during hypoxia in newborn piglets. The present study tests whether modification of the glutamate and ion channel sites of the NMDA receptor correlates with the progressive decrease in cerebral energy metabolism induced by hypoxia. Degrees of cerebral hypoxia were attained by exposure of ventilated piglets to decreased oxygen at different concentrations and confirmed by tissue phosphocreatine levels. During graded hypoxia, the number of glutamate sites decreased, the affinity of the ion channel site increased, the inhibition by Zn(2+) increased, the activation by glutamate increased, and the activation by glycine decreased. Therefore, modification of the NMDA receptor correlates with the energy state of the tissue. Alterations in receptor phosphorylation may gradually modify the NMDA receptor and may be initiated by subtle decreases in tissue oxygenation in the newborn brain. PMID- 12119542 TI - Activity of enzymes involved in energy production in the small intestine during suckling-weaning transition of pigs. AB - The effect of weaning on a potential metabolic capacity of key enzymes involved in the energy production by porcine enterocytes was investigated. The activity of citrate synthase, isocitrate dehydrogenase, alpha-ketoglutarate dehydrogenase, glutamate dehydrogenase, alanine aminotransferase and aspartate aminotransferase was determined in the small intestine epithelium of piglets during suckling weaning transition. Investigations were performed on 5-week-old (suckling), 6 week-old (1st week after weaning) and 7-week-old (2nd week after weaning) piglets. The activity of glutamate dehydrogenase decreased (p < 0.05) during the 1st week after weaning, and remained numerically lower during the 2nd week after weaning than in suckling piglets. The activities of isocitrate dehydrogenase and alanine aminotransferase showed the same pattern as the glutamate dehydrogenase activity and decreased numerically during the 1st and 2nd weeks. The activities of citrate synthase and alpha-ketoglutarate dehydrogenase were numerically lower in post-weaned piglets (1st and 2nd weeks) than in suckling piglets. In contrast, the activity of aspartate aminotransferase was high and remained unchanged from week 5 to the 2nd week post-weaning. The activities of alanine and aspartate aminotransferase were positively correlated in suckling piglets (r = 0.98, p < 0.05) and at the 1st week after weaning (r = 0.99, p < 0.01). Also, both aminotransferases were positively correlated to the activity of alpha ketoglutarate dehydrogenase in suckling piglets (r = 0.95, p < 0.05 and r = 0.95, p < 0.05) and to the activity of isocitrate dehydrogenase during the 1st week after weaning (r = 0.99, p < 0.001 and r = 0.99, p < 0.01). The results indicate additional capacity of the tricarboxylic acid (TCA) cycle for transformation of alpha-ketoglutarate from other sources than acetyl-CoA such as glutamine, glutamate and other amino acids. Further, the high activity of aspartate aminotransferase also suggests a high capacity of porcine small intestinal epithelium to provide the TCA cycle with oxaloacetate during the suckling-weaning transition. PMID- 12119544 TI - Partial liquid ventilation with low dose of perflubron and a low stretch ventilation strategy improves oxygenation in a rabbit model of surfactant depletion. AB - To demonstrate the efficacy of partial liquid ventilation (PLV) with a low dose of perflubron (PFB), severe lung injury model followed surfactant-depleted rabbits underwent PLV with either moderate tidal volume (V(T)) and high positive end-expiratory pressure (PEEP) or high V(T) and low PEEP. PLV with low-dose PFB was effective in alleviating hypoxia in a severe lung injury model if adequate PEEP was applied. We concluded that the addition of a low dose of PFB might be beneficial to a low stretch ventilation strategy, which allows a less aggressive approach to achieve adequate oxygenation with a possible reduction in further lung injury. PMID- 12119543 TI - Effect of 7-nitroindazole on bilirubin-induced changes in brain cell membrane function and energy metabolism in newborn piglets. AB - We evaluated the effects of 7-nitroindazole, a selective neuronal nitric oxide synthetase (nNOS) inhibitor, on bilirubin-induced alterations in brain cell membrane function and energy metabolism in the newborn piglets. The decreased cerebral cortical cell membrane Na(+),K(+)-ATPase activity and increased lipid peroxidation products, indicative of bilirubin-induced brain damage, were significantly attenuated by 7-nitroindazole treatment. 7-Nitroindazole also significantly improved the bilirubin-induced reduction in both brain ATP and phosphocreatine levels, decreased blood-to-brain glucose ratio and increased brain lactate level. In summary, 7-nitroindazole significantly attenuated the bilirubin-induced alterations in brain cell membrane function and energy metabolism in the newborn piglet. These findings suggest that nitric oxide produced by nNOS is involved in mediating or facilitating bilirubin-induced cerebral dysfunction. PMID- 12119545 TI - Common perinatal insults diminish cord blood RANTES. AB - We demonstrate that the cord blood RANTES concentrations are reduced in full-term newborns who were born from meconium-stained amniotic fluid as compared with full term newborns born after normal delivery. Since RANTES inhibits immunodeficiency virus (HIV) entry into macrophages, thereby bestowing increased resistance to HIV (including protection in utero), we propose that common perinatal events might precipitate higher perinatal transmission of HIV. PMID- 12119546 TI - Pharmacogenomics. AB - Pharmacogenomics, a revolutionary chapter in the history of pharmacology, has received new impetus from the development and accessibility of molecular biotechnologies, notably DNA chips. The longstanding notion of responders/non responders has given way to a more organic approach, where idiosyncrasy becomes an obsolete concept. This is a major step towards predictive, individualized medicine. In this review, several applications of pharmacogenomics are considered. Genetic polymorphisms of metabolization reactions, mainly with cytochrome P450, explain most of the cases described today. More fundamental and innovative studies have tried to link the structure of receptors or transporters and drug response. A leading topic in neuropsychopharmacology is the relation between the polymorphism of dopaminergic receptors and the efficacy of, or adverse reaction to, neuroleptics. In asthma, the structure of the beta2 adrenergic receptor has been associated with response to treatment. Intrinsic genetic predisposition also plays an important role in cardiovascular diseases, and the role of ion channel mutations will be discussed. Research in oncological molecular epidemiology has explored the connection between the predisposition to certain cancers and specific enzymatic equipment hindering the detoxification of potentially carcinogenic exogenous compounds, or, on the contrary, promoting metabolic activation implicated in the formation of reactive compounds. The search for determinants of addictive behavior is another vast field of pharmacogenomics. Finally, we consider the impact of pharmacogenomics on the methodology of drug development in preclinical and clinical trials. Progress in methods of phenotyping/genotyping should promote diagnosis, guide the choice of drug for an individual (benefit/risk ratio), and determine dosage and regimen. PMID- 12119547 TI - Lactate, glucose and energy metabolism in the ischemic brain (Review). AB - For many decades, lactate was considered to be an end product of anaerobic glycolysis in mammalian tissues with no other function in metabolism. As determination of lactate level became a routine in hospital blood work, fluctuations in its levels were associated with situations other than oxygen lack. It was just a matter of time before elevated lactate blood levels emerged as a 'red flag' for potential malaise and frequently was blamed as the of it. Lactate and its accompanying acidosis are still considered today to be major contributors to selective neuronal damage in cerebral ischemia despite the emergence of alternative, more compelling postulates as to the causes of this damage. The attitude, especially among clinicians, has been difficult to change despite recent findings that strongly indicate lactate to be a possible beneficial intermediate in brain energy metabolism. This review revisits briefly the annals that brought about the lingering negative attitude toward lactate and expands on the more recent findings and debates that have illuminated this monocarboxylate in a different, more positive light. PMID- 12119549 TI - Keratinocyte injury in drug-induced toxic epidermal necrolysis: Simultaneous but distinct topographic expression of CD95R and calprotectin. AB - Keratinocyte injury in drug-induced toxic epidermal necrolysis (TEN) is attributed to a dysregulation in the complex apoptotic machinery. This study was performed to investigate some aspects of the apoptotic pathomechanism involving calprotectin and the CD95 receptor (CD95R, FasR) in TEN. The expression of these two molecules corresponds to calcium-dependent and calcium-independent processes, respectively. Biopsies were collected from blistering skin in 21 TEN patients and from clinically uninvolved skin in 16 of these patients. Immunohistochemistry was performed using specific antibodies directed to CD95R and calprotectin. Half (8/16) of the biopsy specimens taken from clinically uninvolved sites showed the expression of calprotectin throughout the epidermis or restricted to the suprabasal layers. In these samples, a strong CD95R immunoreactivity was often restricted to the basal layer (8/16). Calprotectin was present in all biopsies of bullous skin, especially in suprabasal layers (15/21) or throughout the epidermis (6/21). By contrast, the prominent expression of CD95R was confined almost exclusively to the basal layer (15/21), and more rarely throughout the epidermis (2/21), and it remained sometimes unexpressed (4/21). A clear-cut boundary was often present between the areas labeled by the two antibodies with exceptional overlap between them. The simultaneous expression of calprotectin and CD95R in TEN at distinct levels of the epidermis indicates that the pathomechanism leading to keratinocyte death does not belong to a single process in TEN. Their expression in clinically uninvolved skin suggests that these processes are early and widespread events in TEN. PMID- 12119548 TI - Therapeutic approach to familial hypercholesterolemia by HVJ-liposomes in LDL receptor knockout mouse. AB - Familial hypercholesterolemia (FH) is an inherited disease in humans, which we have used as a model to develop a new strategy of gene therapy. This disease, which is due to mutation in the low density lipoprotein (LDL) receptor gene and results in deficiency of the LDL receptor, is associated with hypercholesterolemia and premature development of coronary heart disease. This disease has been identified as one of the target diseases for gene therapy, because a 50% reduction of cholesterol level would be beneficial in such patients. In this study, we examined the feasibility of gene therapy by the delivery of the human LDL receptor plasmid into the liver via the portal vein. For gene transfer we utilized HVJ-liposome method with which many successful gene transfers have been reported. Administration of the human LDL receptor plasmid by the HVJ-liposome method into the liver resulted in a decrease of total cholesterol level. Moreover, second administration of this gene two weeks after the first administration resulted in sustained lowering of total cholesterol level. Although single administration of plasmid by the HVJ-liposome method induced antibodies against HVJ, this antibody production did not affect gene expression following second administration. These results suggest the possibility of a novel repetitive gene therapy for FH, using human LDL receptor plasmid transfer directly into the liver by the HVJ-liposome method. PMID- 12119550 TI - Human cultured skin fibroblasts express estrogen receptor alpha and beta. AB - Human skin fibroblasts may be the target cells for estrogens. The aim of present study was to confirm the presence of both isoforms of estrogen receptors (ER) in these cells. Experiments were carried out in primary cultures of human skin fibroblasts. ER-alpha and ER-beta mRNAs were measured by quantitative assays based on reverse transcription (RT) of the mRNA and polymerase chain reaction (PCR) amplification of the cDNA. To determine which of the ER isoforms were present and their intracellular locations immunohistochemical staining was performed. MCF-7 culture was a positive control for the immunostaining. The distribution immunostaining of ER-beta protein differed from that of ER-alpha in skin fibroblasts. ER-alpha was detected in both the cytosolic and nuclear compartments of fibroblasts. ER-beta was weakly detectable and was found predominantly in the nuclear compartment. Using the RT-PCR technique mRNA of both ERs was successfully detected in the skin fibroblast cultures with predominantly higher mean level of ER-beta mRNA expression than ER-alpha mRNA. In human culture skin fibroblasts ER-beta co-expresses with ER-alpha. The dominant expression of ER-beta in cultured female skin fibroblasts suggests that ER-beta may play a dominant role in collaboration with ER-alpha in the regulation of estrogen action in skin. PMID- 12119551 TI - Molecular cloning and characterization of human WINS1 and mouse Wins2, homologous to Drosophila segment polarity gene Lines (Lin). AB - WNT signaling molecules play key roles in carcinogenesis and embryogenesis. Drosophila segment polarity gene Lines (Lin) is essential for Wnt/Wingless dependent patterning in dorsal epidermis and also for hindgut development. With Wnt signaling, Lin accumulates in the nucleus to modulate transcription of Wnt target genes through association with beta-catenin/Armadillo and TCF/Pangolin. Here, human WINS1 and mouse Wins2, encoding proteins with Drosophila Lin homologous domain, were isolated using bioinformatics and cDNA-PCR. Human WINS1 encoded 757-amino-acid protein, and mouse Wins2 encoded 498-amino-acid protein. Human WINS1 and mouse Wins2 showed 60.0% total-amino-acid identity. Lin homologous domain of WINS1 and Wins2 showed 29.4% and 27.2% amino-acid identity with that of Drosphila Lin, respectively. In the human chromosome 15q26 region, WINS1 gene was clustered with ASB7 gene encoding ankyrin repeat and SOCS box containing protein 7. Human WINS1 mRNA of 2.8-kb in size was expressed in adult testis, prostate, spleen, thymus, skeletal muscle, fetal kidney and brain. This is the first report on molecular cloning and initial characterization of human WINS1 and mouse Wins2 PMID- 12119552 TI - Expression of minichromosome maintenance-2 in human malignant fibrous histiocytomas: Correlations with Ki-67 and P53 expression, and apoptosis. AB - This study examined the clinicopathological significance of minichromosome maintenance-2 (MCM2) expression in 38 human malignant fibrous histiocytomas (MFHs) and 36 benign fibrohistiocytic tumors (BFHTs) immunohistochemically, and in 9 human sarcoma or carcinoma cell lines, as well as 7 surgical specimens by Western blotting. MCM2 was detected in all the cell lines and surgical specimens as a single band at 120 kDa, while P53 expression was variable. Nuclear expression of MCM2 was noted in tumor but not mitotic cells of all the MFHs and 26 (72.2%) of the BFHTs, the labeling indices (LIs) being 62.0% in the 28 ordinary types, 38.5% in the 10 myxoid types, and 11.2% in the BFHTs with significant difference. Moreover, the LI was significantly higher for MCM2 than that for Ki-67 in the MFHs of both types (p<0.05). No correlation was noted between the MCM2-LI and P53 expression or apoptotic indices, which were significantly higher in the MFHs than BFHTs (p<0.01). These results indicate that MCM2 would correlate with cell proliferation rather than apoptosis in MFHs, and the expression is ubiquitous in proliferating cells, regardless of the expression of P53. Thus, MCM2 might be a reliable marker of proliferating cells in human MFH. PMID- 12119554 TI - Liprin-alpha2 gene, protein tyrosine phosphatase LAR interacting protein related gene, is downregulated by androgens in the human prostate cancer cell line LNCaP. AB - Prostate cancer is one of the most common neoplasms in the USA and Europe. We used differential display PCR (DD-PCR) to identify androgen-regulated genes in prostate cancer. The RNA of LNCaP cells treated with dihydrotestosterone (DHT) was analyzed for differentially expressed genes. Using DD-PCR, we identified a down-regulated cDNA fragment by DHT in LNCaP cells. This fragment was cloned and expressions of this fragment in prostate cancer cell lines were analyzed by RT PCR. Sequence analysis revealed that a cDNA fragment is identical to protein tyrosine phosphatase LAR related gene, liprin-alpha2. liprin-alpha2 was downregulated by dihydrotestosterone (DHT) in LNCaP cells in a time- and androgen concentration-dependent manner. Downregulation by DHT was not inhibited by the protein synthesis inhibitor cycloheximide. This liprin-alpha2 gene was not expressed in androgen independent prostate cancer cell lines PC-3 and DU-145 at the mRNA level. And also, we first revealed here that liprin-alpha2 mRNA is expressed in LNCaP cells as well as human prostate cancer tissues and normal prostate tissues. These data suggest that liprin-alpha2 might play a role in androgen responsive human prostate cancer cell line as well as human prostate cells, and the loss of this gene expression might be associated with the androgen independent characteristics of prostate cancer. PMID- 12119555 TI - Autologous satellite cell seeding improves in vivo biocompatibility of homologous muscle acellular matrix implants. AB - Acellular matrix obtained from homologous muscular tissue has been previously used to repair muscular defects. However, the implants, although not rejected, give rise to an intense inflammatory response and are rapidly replaced by fibrous tissue. In this study we examined the possibility that co-culture with autologous satellite cells can improve the efficiency of homologous acellular matrix as skeletal muscle substitute. Satellite cells, isolated from rat dorsal muscle, were cultured in vitro on homologous acellular matrix obtained by detergent enzymatic treatment of abdominal muscle fragments. Scanning electron microscopy revealed that after 24 h of co-culture satellite cells were attached to the matrix, but still possessed a round shape. After 96 h, seeded cells began to flatten and to differentiate, originating few multinucleated myotubes. Patches of homologous matrix, seeded or not with autologous satellite cells, were implanted in the dorsal muscle of rats. At autopsy, the implants were recovered and processed for light microscopy. Two weeks after surgery, fibrous tissue started to replace the grafts composed only by acellular matrix, which at the 4th week were transformed into a fibrous scar. In contrast, at both times post-surgery the structure of implants containing autologous satellite cells was well preserved. The inflammatory reaction was modest and fibrosis was confined to the periphery of the grafts. It is concluded that the presence of autologous satellite cells is an important factor to preserve the structural integrity and to improve in vivo biocompatibility of homologous muscular acellular matrix implants. PMID- 12119553 TI - Correlation between interleukin 10 and vascular endothelial growth factor expression in human esophageal cancer. AB - Interleukin 10 (IL-10) is an immunosuppressive cytokine produced by T lymphocytes, and is a regulatory molecule for angiogenesis in various cancers. We examined IL-10 and vascular endothelial growth factor (VEGF) gene expression in 45 esophageal cancer patients who underwent surgical resection. Thirty-seven (82.2%) of the 45 esophageal cancers revealed IL-10 gene expression. VEGF121, VEGF165 and VEGF189 isoforms were detected in 93.3% (42/45), 55.6% (25/45) and 26.7% (12/45) of cases, respectively. IL-10 gene expression was significantly correlated with VEGF121 gene expression (P=0.0039, Fisher's test). The results suggested that IL-10 stimulates angiogenic factor gene expression. PMID- 12119556 TI - Effects of galanin on proliferation and apoptosis of immature rat thymocytes. AB - Evidence indicates that some regulatory peptides (endothelins, cholecystokinin and VIP) are involved in the control of thymus growth, and we have investigated whether galanin may be included in this group of peptides. In fact, galanin, a 29 amino acid peptide acting through three subtypes of G protein-coupled receptors (GalR1, GalR2 and GalR3), seems to play a role in the control of the immune system. Reverse transcription (RT)-polymerase chain reaction (PCR) allowed the detection of galanin, GalR1 and GalR3 mRNAs in the thymus cortex of immature (20 day-old) rats, while GalR2 expression was very weak or absent. Immature rats were given three subcutaneous injections (28, 16 and 4 h before sacrifice) of 2 nmol/100 g galanin and or the galanin-receptor antagonist (galanin-A) [D-Thr(6),D Trp(8,9),15-ol]-galanin(1-15), and 0.1 mg/100 g vincristine 3 h before sacrifice. Thymuses were processed for light microscopy and the percentage of metaphase arrested cells (mitotic index) was evaluated. Galanin-A increased the thymus mitotic index, while galanin was ineffective, thereby suggesting that endogenous galanin exerts a maximal tonic inhibitory effect on the proliferative activity of thymocytes in immature rats. Immature rat thymocytes were cultured in vitro for 12 h in the presence of 10(-6) M galanin and/or galanin-A. Hoechst 33342 and propidium iodide were added to the cultures, and the percentage of apoptotic and necrotic cells was determined under fluorescence microscope. Galanin increased apoptotic index, and the effect was prevented by galanin-A. Neither galanin nor galanin-A altered necrotic index. Collectively, our findings indicate that galanin, probably acting through GalR1 and GalR3, exerts antiproliferative and proapoptotic effects on immature rat thymocytes, which makes it likely that this peptide plays a role in the autocrine/paracrine functional regulation of immune system in the rat. PMID- 12119557 TI - Differential chemosensitivity in human intracerebral gliomas measured by flow cytometric DNA analysis. AB - The present study was designed to investigate the chemosensitivity of human intracerebral gliomas using the flow cytometric (FCM) analysis of DNA integrity as an in vitro drug sensitivity testing. We also correlated the in vitro results and the clinical responses to chemotherapy. Forty-one fresh tumor specimens were obtained at surgery, and exposed to 30 currently-available anticancer agents. Drug-induced nuclear damage such as chromatin condensation or DNA degradation were assessed both by morphological observation and FCM analysis of DNA integrity. The FCM assay could be performed in all the cases (100% success rate). The chemosensitivities of anaplastic astrocytoma, oligodendroglial tumors, and medulloblastoma were generally higher than those of glioblastoma multiforme and ependymoma. The in vitro chemosensitivity was markedly heterogeneous among patients with the same histological tumor. Clinical sensitivity was predicted in 86% of all evaluable patients and clinical resistance in 81%; the overall accuracy of the FCM assay was 82%. The FCM analysis of DNA integrity proved to be feasible and sufficiently reliable as a routine clinical examination for the purpose of screening new chemotherapeutic agents and individualizing chemotherapy regimens for patients with intracerebral gliomas. PMID- 12119558 TI - Resveratrol, a chemopreventive agent, disrupts the cell cycle control of human SW480 colorectal tumor cells. AB - Resveratrol is a natural polyphenolic compound produced by a number of plants and found in high amount in peanuts, seeds, grapes or berries as source of human nutrition. Epidemiological studies strongly suggest that resveratrol may act as a cancer chemopreventive compound. The mechanism by which resveratrol inhibits cell proliferation was studied in human colorectal tumor SW480 cell line. The results show that resveratrol strongly inhibits cell proliferation at the micromolar range in a time- and dose-dependent manner. Resveratrol appears to block the cell cycle at the transition --> G2/M since inhibition of [(3)H]-thymidine incorporation is not observed, while there is an increase of the cell number in S phase. During this inhibition process, resveratrol increases the content of cyclins A and B1 as well as cyclin-dependent kinases Cdk1 and Cdk2. Moreover, resveratrol promotes Cdk1 phosphorylation. In conclusion, resveratrol exerts a strong inhibition of SW480 human colorectal tumor cell proliferation at least by modulating cyclin and cyclin-dependent kinase activities. PMID- 12119559 TI - Efficacy of long-term interferon therapy in chronic hepatitis B patients with HBV genotype C. AB - Infection with Hepatitis B virus (HBV) genotype C predominates in Japan. We analyzed the efficacy of interferon (IFN) alpha or beta in the treatment of chronic hepatitis B patients with HBV genotype C and the clinical predictors for therapeutic response. Forty-three genotype C-infected, chronic hepatitis B e antigen (HBeAg)-positive patients (32 men and 11 women with a mean age of 35.6+/ 10.1 years) who had been treated with IFN therapy were retrospectively studied. The patients were classified into two treatment groups. Short-term therapy group was administered a 5-6 MU dose three times weekly for 4 weeks, and the long-term therapy group for 24 weeks. At the end of the follow-up period, 4 (15%) of 27 short-term therapy group patients and 6 (38%) of 16 long-term therapy group patients had normalized serum ALT levels and seroconversion of HBeAg to anti-HBe (p=0.137). Multivariate analysis for parameters most important for the efficacy of IFN therapy was performed using Cox proportional hazard models in order to investigate the association between baseline characteristics of patients and the response to IFN treatment. As a result, the p-values of IFN treatment group and sex were <0.05, and both factors can be recognized as independent significant factors (relative risk, 2.93 and 2.53; p=0.027 and 0.040, respectively). Furthermore, the cumulative rates of seroconversion of HBeAg to anti-HBe analyzed by the Kaplan-Meier method was significantly higher in the female group (p=0.015) and in the long-term IFN therapy group (p=0.0046). In summary, long-term IFN therapy may be more effective than short-term IFN therapy for patients with chronic HBV genotype C infection. PMID- 12119560 TI - Telomerase activity and hTERT mRNA in development and progression of adenoma to colorectal cancer. AB - Telomerase activity and hTERT mRNA expression are upregulated in colorectal cancer. Whether they are inherent in colorectal adenomas, premalignant lesions to cancer, however, remains to be elucidated. We examined telomerase activity by the fluorescence-based telomeric repeat amplification protocol method and analyzed the level of hTERT mRNA by real-time polymerase chain reaction in 74 surgically obtained neoplasms from 29 patients. The specimens were divided into 6 categories according to the criteria of the Vienna Classification. The control comprising 29 non-pathological mucosa were classified into category 1, 6 adenomas indefinite for neoplasia into category 2, 21 non-invasive low grade adenomas into category 3, 23 high grade adenomas or non-invasive carcinomas into category 4, and 15 intramucosal or submucosal carcinomas into category 5. Carcinoma invading beyond the submucosa (9 samples) was referentially subdivided into category 6. Telomerase activity (mean +/- standard error) in 1 categories to 6 were 5.0+/ 1.2, 1.8+/-1.7, 4.3+/-1.6, 20.2+/-2.1, 36.4+/-5.5, and 55.5+/-8.2 units/microg protein, respectively. There were no statistical differences between categories 1 and 2, 1 and 3, and 2 and 3. A significant statistical difference in the other two was observed by the multiple comparison test. The mean levels of hTERT mRNA was 103.1+/-102.4, 103.6+/-103.0, 103.6+/-102.9, 103.7+/-102.9, 104.0+/-103.4, and 104.4+/-104.0 copies/microg total RNA, respectively. There was a significant statistical difference only between category 6 and each of the other categories. These results suggest that telomerase activation occurs during the progression from low-grade to high-grade dysplasia in adenomas and increases steadily with the progression of the degree of dysplasia and invasion during colorectal carcinogenesis, and that hTERT mRNA expression is a feature of the late stage development of colorectal cancer. PMID- 12119561 TI - Expression of survivin is associated with malignant potential in epithelial ovarian carcinoma. AB - Survivin is a new member of the inhibitor of apoptosis family of anti-apoptotic proteins. It has been reported that survivin is expressed during fetal development and in cancer tissues. Because suppression of apoptosis is important for carcinogenesis and tumor growth, we investigated the expression of survivin in human endometrial carcinomas. We analyzed serial frozen sections for survivin protein expression in 26 patients with ovarian epithelial carcinoma and 10 patients with benign cystadenoma of the ovary by fluorescent immunohistochemistry. We analyzed the relationship between the percentages of survivin-stained cells and the characteristics of the patient including histological classification, clinical stage, histological grade, and clinical outcome. Survivin was weakly detected in some benign ovarian cystadenomas (0 12.1%). There was, however, abundant survivin immunoreactivity in the nucleus and/or cytoplasm of the epithelial ovarian carcinoma cells. Scoring on the basis of the percentage of positive cells indicated that survivin expression was significantly associated with PCNA-labeling index, clinical stage, histological grade, clinical outcome, and survival rate (p<0.01, respectively). We conclude that the survivin protein is a defining diagnostic marker for epithelial ovarian carcinomas that may also yield prognostic information. PMID- 12119562 TI - Hepatocyte growth factor ameliorates renal hemodynamic disorder after ischemia/reperfusion. AB - Ischemic injury of the transplanted kidney is one of the causes of reduced graft survival. The purpose of the present experiment was to examine whether hepatocyte growth factor (HGF) would improve acute renal hemodynamic recovery immediately after cold ischemia. Addition of HGF to the preservation solution during 3 h cold ischemia of dog kidney accelerated both recovery of renal blood flow and glomerular filtration rate (GFR). It is suggested that HGF may be useful for preservation of excised kidney for transplantation. As intrarenal arterial infusion of HGF in normal dog kidney had no effects on renal hemodynamics, mechanisms other than direct vasodilator action of HGF appear to be operating in the protection. PMID- 12119563 TI - Molecular cloning and characterization of OSR1 on human chromosome 2p24. AB - During Drosophila hindgut development, bowl, caudal/CDX, brachyenteron/Brachyury/TBX, fork head/FOX, drumstick, lines, and wingless/WNT play important roles. Drosophila bowl gene is homologous to Drosophila odd skipped (odd) gene and odd-skipped related gene (sob). Here, human OSR1, related to Drosophila odd, was isolated using bioinformatics and cDNA-PCR. OSR1 was found to encode 266 amino-acid protein with three C2H2-type zinc fingers, a tyrosine phosphorylation site (Tyr 203), and several putative PXXP SH3 binding motifs. Three zinc fingers and a tyrosine phosphorylation site were conserved among human OSR1, OSR2, Drosophila odd, sob, and bowl. OSR1 showed 63.6% total amino-acid identity with OSR2. OSR1 gene consisting of three exons was located on human chromosome 2p24. OSR1 mRNA of 2.3-kb in size was detected in adult colon, small intestine, prostate, testis, and fetal lung. OSR1 mRNA was significantly up regulated in a pancreatic cancer cell line MIA PaCa-2, and was weakly expressed in gastric cancer cell lines OKAJIMA, MKN45, pancreatic cancer cell lines PANC-1, BxPC-3, AsPC-1, PSN-1, Hs766T, and esophageal cancer cell line TE10. Among 10 cases of primary gastric cancer, OSR1 mRNA was up-regulated in 5 cases, and was down-regulated in 2 cases. This is the first report on molecular cloning and characterization of human OSR1. PMID- 12119564 TI - Molecular cloning, mapping and characterization of a human CK1gamma1 gene. AB - We isolated and sequenced a cDNA clone coding a human protein kinase CK1 (casein kinase 1) by screening a human fetal brain cDNA library. This new cDNA clone of 1756 bp contained an open reading frame, encoding a protein of 438 amino acids with a molecular weight of 50,272 Da and an isoelectric point of 9.37. The entire amino acid sequence of the novel human CK1 was 94% homologous to that of rat CK1gamma1. Northern blot analysis indicated that the human CK1gamma1 was highly expressed in the liver, skeletal muscle, heart and kidney. Furthermore, the human CK1gamma1 gene was mapped to chromosome 15q22 between STS marker D15S159 and D15S125 by polymerase chain reaction analysis of human/rodent hybrid cell panels. PMID- 12119565 TI - Premature ovarian failure and FMR1 premutation co-segregation in a large Brazilian family. AB - Fragile X syndrome is the most common form of inherited mental retardation in men. The molecular mechanism underlying the disease is an amplification of a polymorphic trinucleotide repeat (CGG)n located at 5' end of FMR1 which promotes transcriptional silencing of the gene. Four different classes of alleles could be distinguished in the population based on the size of the repeat, however only large amplifications over 200 CGG are associated with the disease. In the past decade several authors have associated premutated alleles, which harbor expansions from 61 to 200 repeats, with the occurrence of premature ovarian failure (POF). In this work we describe a large Brazilian family in which a POF/premutated woman has transmitted to five out of seven daughters a FMR1 premutated allele. From these five women with premutations, three have experienced premature ovarian failure. Our data clearly indicate a co-segregation pattern of inheritance between POF and fragile X premutation. PMID- 12119566 TI - Effects of distraction on treadmill running time in severely obese children and adolescents. AB - OBJECTIVE: (1) To examine the effects of attentional distraction on running time in an incremental treadmill test in obese youngsters; (2) to investigate whether distraction works at the same extent at the beginning and at the end of residential treatment; and (3) to explore the underlying mechanisms of the possible distraction effects. METHODS: Thirty severely obese youngsters (10 boys, 20 girls, age range 9-17) who were following a 10 month residential treatment, performed a treadmill test until exhaustion in four different sessions using a within subjects design. The two sessions at the beginning of the treatment and the two sessions at the end the treatment were counterbalanced, one with attentional distraction (music) and one without distraction. RESULTS: Obese youngsters ran significantly longer during distraction. This distraction effect seemed to be larger at the beginning compared to at the end of obesity treatment. The absence of differences between the condition with music and the condition without music on perceived bodily symptoms is in line with the idea that it took longer to perceive sufficient bodily sensations to decide to stop the treadmill test in the distraction condition. This interpretation is further corroborated by the physiological data indicating a superior peak performance in the condition with distraction. CONCLUSIONS: Attentional distraction has a positive effect on perseverance in obese youngsters. Further research has to show the usefulness of attentional distraction as a technique to increase exercise adoption and adherence in obesity treatment. PMID- 12119568 TI - Obesity in children and adolescents in Cyprus. Prevalence and predisposing factors. AB - OBJECTIVE: To estimate the prevalence of childhood and adolescent obesity in Cyprus and define possible associated risk factors. STUDY DESIGN: Cross-sectional study of a representative sample of children 6-17 y of age performed during October 1999 to June 2000. METHODS: Anthropometric data were taken using standard methods, from 2467 children. Certain diet and physical activities as well as other socioeconomic family parameters were assessed with the aid of a questionnaire. Obesity and overweight were defined using both the National Health and Nutrition Examination Survey (NHANES) I definition and the newer International Obesity Task Force (IOTF) definition. Logistic regression analyses were performed to estimate the influence of various parameters. RESULTS: The prevalence of obesity in males was 10.3% and in females 9.1% using the NHANES I definition and 6.9 and 5.7%, respectively, using the IOTF definition. The percentages presented a decreasing trend with age. There were an additional 16.9% of males and 13.1% of females defined as overweight with the NHANES I definition and 18.8 and 17.0%, respectively, using the IOTF definition. The most significant associated factor for obesity was parental obesity status. The odds ratio for offspring obesity when both parents were obese ranged from 11.34 (95% CI 1.83 75.50) in females 6 to 11-y-old to 18.09 (95% CI 2.06-158.81) for males 12 to 17 y-old. CONCLUSIONS: The prevalence of childhood and adolescent obesity was estimated for the first time in a representative sample from Cyprus, and this rate is comparable to that observed in North America. These results indicate the need for individual and population measures for the treatment and prevention of pediatric obesity. The rate of obesity differs significantly depending on the method of estimation. PMID- 12119567 TI - Low-density lipoprotein particle size, central obesity, cardiovascular fitness, and insulin resistance syndrome markers in obese youths. AB - OBJECTIVE: (1) To determine the prevalence of small dense low-density lipoprotein (SDLDL) particles in obese youths and (2) to compare youths with SDLDL and large buoyant LDL (LBLDL) subclass phenotypes in total body and abdominal fatness, cardiovascular (CV) fitness, and markers of the insulin resistance syndrome (IRS). DESIGN: For group comparisons, subjects were dichotomized into either SDLDL phenotype group or LBDL phenotype group based on LDL particle size. SUBJECTS: Obese 13 to 16-y-olds (n=80) who had a triceps skinfold greater than the 85th percentile for gender, ethnicity, and age. MEASUREMENTS: LDL particle size, plasma lipids and lipoprotein concentrations, plasma glucose and insulin concentrations, and blood pressures; percentage body fat, visceral adipose tissue (VAT); VO(2) at a heart rate of 170 bpm as an index of CV fitness. RESULTS: The prevalence of the SDLDL phenotype was 54% among the 80 obese youths. Although overall body fatness (ie BMI and percentage body fat) and CV fitness were similar between the two LDL phenotype groups, the SDLDL phenotype group had significantly higher weight, waist circumference and VAT than the LBLDL phenotype group. With respect to the IRS markers, youths with the SDLDL phenotype had significantly higher triacylglycerol (TAG), very low-density lipoprotein cholesterol (VLDLC), apolipoprotein B (apo B), and total cholesterol-to-high-density lipoprotein ratio (TC/HDLC) than youths with the LBLDL phenotype. LDL particle size as a continuous variable was significantly correlated with TAG, VLDLC, apo B, HDLC, and TC/HDLC. Plasma TAG and HDLC concentrations were independent predictors of LDL particle size. CONCLUSION: (1) The SDLDL phenotype was common in obese youths and (2) the relationships of LDL particle size with several of the IRS markers suggested that already in adolescence the expression of the SDLDL phenotype might be an important risk factor for future coronary heart disease mortality and morbidity. PMID- 12119569 TI - Influence of obesity on cardiovascular risk. Twenty-three-year follow-up of 22,025 men from an urban Swedish population. AB - OBJECTIVE: To assess to what extent the incidence of coronary events and death related to smoking, hypertension, hyperlipidemia and diabetes is modified by obesity. DESIGN: Prospective cohort study. SUBJECTS: A total of 22 025 men aged 27 to 61-y-old at entry. MEASUREMENTS: Incidence of coronary events (CE, ie acute myocardial infarctions and deaths due to chronic ischaemic heart disease) and death during 23 y of follow-up was studied in relation to body mass index (BMI), heart rate, blood pressure, blood lipids, glucose and insulin, lifestyle factors, history of angina pectoris, history of cancer, self-reported health and socio economic conditions. RESULTS: At the end of follow-up 20% of the obese men were no longer alive, and 13% had had a coronary event. Incidence of CE was 16% lower (RR (relative risk) 0.84; 95% confidence interval (CI) 0.65-1.10) among underweight (n=1171), 24% higher (RR 1.24; CI 1.12-1.37) among overweight (n=7773), and 76% higher (RR 1.76; 95% CI 1.49-2.08) among obese men (n=1343) than it was among men with normal BMI (n=11 738). The risk associated with overweight and obesity remained statistically significant after adjustment for potential confounders (RR 1.18; CI 1.07-1.31; and 1.39; 1.17-1.65, respectively). The association between BMI and mortality was J-shaped. In all, 1.7% of the obese men were smokers with hypertension, hyperlipidaemia and diabetes, 16.3% were not exposed to any of these risk factors. The cardiovascular risk associated with obesity was small in the absence of other risk factors. Between smoking and obesity there was a statistically significant synergistic effect. CONCLUSIONS: Obesity is associated with an increased incidence of coronary events and death. The risk associated with obesity is substantially increased by exposure to other atherosclerotic risk factors, of which smoking seems to be the most important. The preventive potential of these associations should be evaluated in controlled trials. PMID- 12119570 TI - Adiposity, adipose tissue distribution and mortality rates in the Canada Fitness Survey follow-up study. AB - OBJECTIVE: To compare mortality rates across indicators of adiposity and relative adipose tissue distribution in the Canadian population. SUBJECTS: The sample included 10,323 adult participants 20-69 y of age from the Canada Fitness Survey who were monitored for all-cause mortality over 13 y. METHODS: BMI, waist circumference (WC) and the sum of five skinfolds (SF5) were indicators of adiposity, and the first principal component of skinfold residuals (PC1) represented subcutaneous adipose tissue distribution. Proportional hazards regression was used to estimate relative mortality risk from mortality rates across levels of adiposity and adipose tissue distribution, controlling for the confounding effects of age, smoking status and alcohol consumption. RESULTS: :Significant curvilinear (J-shaped) relationships in men and linear relationships in women were observed between BMI, WC and SF5 and all-cause mortality rates. PC1 was not related to mortality rates in either men or women. In women, the inclusion of the other indicators of adiposity and adipose tissue distribution did not significantly add to the prediction of mortality rates beyond BMI; however, combinations of BMI and both WC and SF5 produced significant models in men. CONCLUSION: The results support the hypothesis that overall level of adiposity is an important predictor of all-cause mortality, more so than the relative distribution of subcutaneous body fat, once overall level of body fatness has been accounted for. PMID- 12119571 TI - Four anthropometric indices and cardiovascular risk factors in Taiwan. AB - OBJECTIVE: To examine the relationships between four anthropometric measurements and cardiovascular risk factors in Taiwan. DESIGN: The data was collected from four nationwide health screen centers in Taiwan from 1998 to 1999. SUBJECTS: A total of 38 556 subjects: 18 280 men and 20 276 women, mean age=37.0+/-11.1 y. None had any known major systemic diseases or were currently on medication. MEASUREMENTS: Individual body weight, height, waist circumference (WC), and cardiovascular risk factors (blood pressure, fasting plasma glucose, triglycerides, total cholesterol level, low-density and high-density-lipoprotein cholesterol level) were assessed and their relationships were examined. RESULTS: In both sexes, with increasing body mass index (BMI), WC, WHpR (waist-to-hip ratio) and WHtR (waist-to-height ratio), there were significantly higher risks of hypertension, impaired fasting glucose, diabetes and dyslipidemia (P<0.001) in almost all age groups. In the age groups older than 65, however, the relationships were statistically inconsistent. CONCLUSIONS: In Taiwan, the four anthropometric indexes (BMI, WC, WHpR, WHtR) are closely related to cardiovascular risk factors. PMID- 12119572 TI - Medical care expenditure associated with body mass index in Japan: the Ohsaki Study. AB - OBJECTIVE: To examine the impact of body mass index (BMI) upon medical care use and its costs in Japan. DESIGN: A population-based prospective cohort study from 1995 to 1998. SUBJECTS: A cohort of 41 967 Japanese adults aged 40-79 y. Subjects who died during the first year of follow-up, or who at baseline reported having had cancer, myocardial infarction, stroke or kidney disease were excluded. MEASUREMENTS: Medical care use and its costs, actual charges, by linkage with the National Health Insurance claim history files after adjustment of smoking, drinking and physical functioning status. RESULTS: There was a U-shaped association between BMI and total medical costs. The nadir of the curve was found at a BMI of 21.0-22.9 kg/m(2). Relative to the nadir, total costs were 9.8% greater among those with BMIs of 25.0-29.9 (rate ratio, 1.10; 95% confidence interval (CI), 1.03-1.17), and 22.3% greater among those with BMIs of 30.0 or higher (rate ratio, 1.22; 95% CI, 1.08-1.37). Estimated excess direct costs attributable to overweight (BMI of 25.0-29.9 kg/m(2)) and obesity (BMI of 30.0 kg/m(2) or higher) represent 3.2% of total health expenditure in the present study, which is within the range reported in Western countries (0.7-6.8%). CONCLUSION: Our prospective data demonstrate that the impact of overweight and obesity upon medical care costs in Japan is as large as in Western countries, despite the much lower mean BMI in Japanese populations. PMID- 12119573 TI - Body weight and obesity in adults and self-reported abuse in childhood. AB - BACKGROUND: Little is known about childhood factors and adult obesity. A previous study found a strong association between childhood neglect and obesity in young adults. OBJECTIVE: To estimate associations between self-reported abuse in childhood (sexual, verbal, fear of physical abuse and physical) adult body weight, and risk of obesity. DESIGN: Retrospective cohort study with surveys during 1995-1997. PATIENTS: A total of 13,177 members of California health maintenance organization aged 19-92 y. MEASUREMENTS: Body weight measured during clinical examination, followed by mailed survey to recall experiences during first 18 y of life. Estimates adjusted for adult demographic factors and health practices, and characteristics of the childhood household. RESULTS: Some 66% of participants reported one or more type of abuse. Physical abuse and verbal abuse were most strongly associated with body weight and obesity. Compared with no physical abuse (55%), being 'often hit and injured' (2.5%) had a 4.0 kg (95% confidence interval: 2.4-5.6 kg) higher weight and a 1.4 (1.2-1.6) relative risk (RR) of body mass index (BMI) > or = 30. Compared with no verbal abuse (53%), being 'often verbally abused' (9.5%) had an RR of 1.9 (1.3-2.7) for BMI > or = 40. The abuse associations were not mutually independent, however, because the abuse types strongly co-occurred. Obesity risk increased with number and severity of each type of abuse. The population attributable fraction for 'any mention' of abuse (67%) was 8% (3.4-12.3%) for BMI > or = 30 and 17.3% (-1.0-32.4%) for BMI > or = 40. CONCLUSIONS: Abuse in childhood is associated with adult obesity. If causal, preventing child abuse may modestly decrease adult obesity. Treatment of obese adults abused as children may benefit from identification of mechanisms that lead to maintenance of adult obesity. PMID- 12119574 TI - Body image and weight change in middle age: a qualitative study. AB - AIMS: To explore men's and women's experiences of weight change in adulthood, body image preferences and beliefs about the health consequences of overweight and to inform the development of a primary care intervention to prevent obesity. SAMPLE: Seventy-two men and women aged 35-55, with a range of BMIs from 22 to 29.9, were identified from two UK general practice registers and invited to participate in an interview about experiences of weight change in adulthood. METHODS: Audio tape recorded, semi-structured interviews were conducted in respondents' homes by trained researchers. Open-ended questions were used to collect experiences of weight change since early adulthood and views about weight change in middle age. Illustrations of a range of men's and women's body shapes were used to prompt discussion of respondents' preferences for male and female body shapes and their perspectives of the health, social and practical problems associated with underweight and overweight. The data were analysed using both quantitative and qualitative methods. RESULTS: Some 87% (33/38) of the women and 59% (20/34) of the men said that they had ever tried to lose weight. At least one instance of successful weight loss was reported by 58% of the women and 47% of the men, although many of these attempts were relatively short-lived and often motivated by specific goals such as a holiday or a wedding. Respondents were sceptical of the possibility of controlling weight without considerable personal sacrifice. Explanations for middle-age weight gain included a sedentary lifestyle, as well as several gender-specific reasons, including hormonal changes and comfort eating for women and beer drinking for men. Nearly all (97%) respondents associated heart disease with overweight, while diabetes was mentioned by only 22% and none mentioned cancer. CONCLUSION: People who have gained weight in middle age may be deterred from trying to prevent further gain by pessimism about the effort required. The efficacy of interventions to encourage relatively small substitutions and changes to diet and physical activity need to be tested. Interventions to help prevent weight gain in middle age could include information about the less widely known health risks such as diabetes and cancer. PMID- 12119575 TI - Effect of oral oleoyl-estrone on adipose tissue composition in male rats. AB - OBJECTIVE: To determine whether the oral administration of oleoyl-estrone has similar mass-decreasing effects on the main different sites of white adipose tissue (WAT). DESIGN: Adult male Zucker lean rats were given a daily oral gavage of oleoyl-estrone (OE, 10 micromol/kg) in 0.2 ml of sunflower oil for 10 days, and were compared with controls receiving only the oil. The mass of the main WAT sites: subcutaneous, epididymal, mesenteric, retroperitoneal, gluteal, perirenal and interscapular, as well as perirenal and interscapular brown adipose tissue (BAT), were dissected and studied. MEASUREMENTS: The tissue weight, DNA, protein, lipid and total cholesterol content, together with the levels of leptin and acyl estrone in the larger WAT and BAT masses, were measured. RESULTS: The weights of WAT depots were correlated with body weight but those of BAT were not. Cell size was maximal for epididymal and mesenteric and minimal for subcutaneous and retroperitoneal WAT and BAT. Differences were detected in DNA, and in protein and lipid content between distinct WAT sites. OE treatment tended to decrease cell number and cell size in WAT; only small differences in composition were found between WAT locations inside the visceral cavity and those outside. Decreases in lipid content were maximal in mesenteric fat. Leptin and acyl-estrone content were fairly uniform at the different WAT sites, except for high concentrations in gluteal WAT. OE induced a greater decrease in leptin and acyl-estrone than in DNA and lipids; changes in these hormones were fairly parallel in all sites. CONCLUSIONS: In general, the differences in composition between visceral and peripheral subcutaneous WAT and their responses to OE were less marked than the individual differences observed between specific sites, regardless of location. WAT sites are fairly diverse in composition, but their response to OE treatment was uniform. OE decreased the weight of WAT through reduction of both cell numbers and size; but did not change the mass or composition of BAT significantly. The effects of OE are more marked in the hormonal signals (leptin and acyl-estrone) from the tissue than in its composition and mass. PMID- 12119576 TI - Estrogen receptor beta is involved in the anorectic action of estrogen. AB - OBJECTIVE: Estrogen has been implicated in feeding behavior and adiposity. This study was undertaken to elucidate the mechanism underlying the anti-obesity and anorectic action of estrogen and the role of estrogen receptor (ER) in the central nervous system. METHODS AND RESULTS: Ovariectomy in 8-week-old female Wistar rats induced hyperphagia along with an increase in body weight and abdominal fat accumulation compared to control sham-operated rats. These changes were fully reversed by subcutaneous replacement of estradiol and were abrogated by pair-feeding. Then, the effects of intracerebroventricular infusion of estradiol, alone or in combination with antisense oligodeoxynucleotides (ODN), for ER in ovariectomized rats were examined. The estradiol group showed 10-20% lower daily food intake, and after the 2-week infusion period a 14% reduction in body weight with a similar reduction in abdominal fat compared to the vehicle group. The inhibitory effect of estradiol on food intake and body weight was blocked by co-administration of ER-beta antisense ODN, whereas ER-alpha antisense ODN did not show any influence. CONCLUSION: These results indicate that ER-beta in the central nervous system is involved in the anorectic action of estrogen. PMID- 12119577 TI - Activities of cytochrome c oxidase and citrate synthase in lymphocytes of obese and normal-weight subjects. AB - BACKGROUND: Obesity represents a heterogeneous group of disorders associated with broad spectrum of metabolic and endocrine abnormalities. The metabolic changes in obesity may also concern the efficacy of mitochondrial system of energy provision. The aim of our study was to analyse activities of mitochondrial enzymes cytochrome c oxidase (COX) and citrate synthase (CS) in isolated lymphocytes of obese and normal-weight subjects. RESULTS: In the group of 304 non obese controls, differences between men and women were found neither in the COX and CS activities nor in the COX/CS ratio in isolated lymphocytes. The activity of COX did not change even with age, whereas the activity of CS decreased significantly resulting in age-dependent increase of the COX/CS ratio (P<0.01). In the group of 60 obese patients aged 17-75 y, the COX activity was 1.2-fold higher (P<0.01) and the CS activity was 1.3-fold lower (P<0.01) compared to 151 non-obese healthy age-matched controls. Consequently, the COX/CS ratio became 1.7 fold higher (P<0.01) in the obese patients compared to the non-obese population, which indicates that both the absolute and relative oxidative capacity are increased. CONCLUSION: Isolated lymphocytes from peripheral blood contribute very little to the overall metabolic turnover, but they may serve as easily available marker cells for studying the changes of mitochondrial energy converting systems in obesity. PMID- 12119578 TI - Prolonged adaptation to fat-rich diet and training; effects on body fat stores and insulin resistance in man. AB - OBJECTIVE: To investigate the effect of prolonged adaptation to training and fat- or carbohydrate-rich diet on body composition and insulin resistance. DESIGN: Longitudinal study. Of three groups two consumed a fat-rich diet, of which one performed regular training (FAT-Train, n=17) and the other maintained normal habitual activity (Fat-Control, n=8). The third group trained and consumed a carbohydrate-rich diet (CHO-Train, n=16). SUBJECTS: Forty-one untrained, healthy male subjects. MEASUREMENTS: Before and after 7 weeks body composition was estimated from skinfold measurements. At rest the respiratory exchange ratio (RER) was determined by the Douglas bag technique. Glycogen was determined in m vastus lateralis and concentrations of insulin and triacylglycerol in serum and glucose, fatty acid and beta-hydroxy-butyrate in plasma was measured. The insulin resistance index was calculated from fasting plasma insulin and glucose values. RESULTS: Across the 7 weeks body weight was reduced (1.3+/-0.3%) in all three groups, however body fat mass was decreased only in the CHO-Train (13%) and maintained in the two FAT-groups. RER at rest was similarly decreased (5%) in the three groups. Plasma insulin tended to decrease (16%) in CHO-Train (P=0.065) and remained unchanged in the two FAT-groups. In contrast plasma glucose (4.6+/-0.1 mmol/l) and plasma FA (453+/-27 micromol/l) remained unchanged across the 7 weeks. The calculated insulin resistance index HOMA-R(mod) was significantly decreased by 19% in CHO-train but remained unchanged in both of the FAT-groups, whereas the calculated insulin secretion index HOMA-beta(mod) was unchanged in all three groups. CONCLUSION: In the present study we demonstrate that despite of a mild energy deficit body fat mass was maintained after prolonged adaptation to fat-rich diet both when normal physical activity was maintained and when training was performed. In contrast a significant decrease in fat mass was observed when carbohydrate-rich diet and training was combined. Furthermore we observed that the insulin resistance index was significantly decreased only when training was combined with a carbohydrate-rich diet. PMID- 12119579 TI - Serum leptin concentration in obese patients with binge eating disorder. AB - BACKGROUND: In steady-state conditions serum leptin concentration is directly related to body fat stores, but is also affected by changes in energy balance. This cross-sectional study investigated the serum leptin concentrations of severely obese patients with binge eating disorder (BED), in whom body fat was greater than normal and, because of eating pattern, rapid and repeated changes in energy balances took place. METHODS: A group of BED obese patients was compared to a group of obese patients with a regular eating pattern with the same body weight, body composition and resting energy expenditure. Serum leptin was measured and the eating attitudes were evaluated by Eating Inventory and Eating Disorder Inventory. RESULTS: In these patients serum leptin concentrations were only weakly correlated to body mass. Furthermore, in BED obese patients serum leptin concentration was higher than in their non binging counterparts. CONCLUSIONS: In obese patients both body fat size and eating behavior influence serum leptin concentration, but BED patients binge eating is not triggered by a low leptin value. PMID- 12119580 TI - Effects of chronic peanut consumption on energy balance and hedonics. AB - OBJECTIVE: To investigate the effects of chronic peanut consumption on energy balance and hedonics. DESIGN: Thirty-week, cross-over, intervention study. Participants were provided 2113+/-494 kJ/day (505+/-118 kcal/day) as peanuts for 8 weeks with no dietary guidance (free feeding-FF), 3 weeks with instructions to add peanuts to their customary diet (addition-ADD) and 8 weeks where peanuts replaced an equal amount of other fats in the diet (substitution-SUB). SUBJECTS: Fifteen, healthy, normal-weight (BMI of 23.3+/-1.8) adults, aged 33+/-9 y. MEASUREMENTS: Dietary intake, appetitive indices, energy expenditure, body weight and hedonics. RESULTS: During FF, peanut consumption elicited a strong compensatory dietary response (ie subjects compensated for 66% of the energy provided by the nuts) and body weight gain (1.0 kg) was significantly lower than predicted (3.6 kg; P<0.01). When customary dietary fat was replaced with the energy from peanuts, energy intake, as well as body weight, were maintained precisely. Participants were unaware that body weight was a research focus. Resting energy expenditure was increased by 11% after regular peanut consumption for 19 weeks (P<0.01). Chronic consumption of peanuts did not lead to a decline in pleasantness or hunger ratings for peanuts nor did it lead to any hedonic shift for selected snack foods with other taste qualities during any of the three treatments. CONCLUSIONS: Despite being energy dense, peanuts have a high satiety value and chronic ingestion evokes strong dietary compensation and little change in energy balance. PMID- 12119582 TI - Weight and dieting: examining levels of weight concern in British adults. AB - OBJECTIVE: Amid rising levels of obesity, not all overweight individuals recognize that their weight is too high. At the same time many of those whose weight is within the normal range are dissatisfied with their body size, providing evidence of inappropriate weight aspirations, especially amongst women. This research examines the nature and level of complacency and over-concern in overweight, underweight and normal-weight individuals. METHODS: Data on weight, perceived overweight and dieting status were collected from a stratified probability sample of 1894 British adults, as part of the Office of National Statistics' Omnibus Survey. RESULTS: Most obese adults correctly perceived themselves as overweight, but many were not trying to lose weight, and only a minority had participated in a programme of weight control. Men's awareness was lower than women's. At the other extreme, few men, but around a quarter of normal weight women felt overweight or were trying to lose weight, but their preferred weight was only slightly below their actual weight. CONCLUSIONS: :These results suggest that weight concern among British women is high, but probably not excessive and there is little evidence for idealization of dangerously low weights. In contrast, many overweight men were unaware of their weight problem. Only around half of those who would benefit from weight reduction were trying to lose weight, and few had received advice from health professionals. In view of the prevalence of obesity, there may be opportunities to provide more guidance on weight control within primary care. PMID- 12119581 TI - Relationships between changes in weight and changes in cardiovascular risk factors in middle-aged French subjects: effect of dieting. AB - OBJECTIVE: To investigate relationships between changes in weight and changes in cardiovascular risk factors in adults, taking intentionality of weight loss into account. DESIGN: Longitudinal study in middle-aged French subjects from the SU.VI.MAX cohort. SUBJECTS: A total of 1773 men (body mass index (BMI) 25.4+/-3.1 kg/m(2), mean+/-s.d) and 2316 women (BMI 23.3+/-3.8 kg/m(2)) aged 45 y and over at baseline. MEASUREMENTS: Weight, height, blood pressure, serum total cholesterol and fasting blood glucose were measured at baseline and after a 2 y follow-up. Self-reported dieting in order to lose weight, smoking status, leisure time physical activity, health events and current treatments were assessed through questionnaires. RESULTS: In multivariate analyses, weight change was positively associated with changes in systolic and diastolic blood pressure (P=0.0002 in women, P=0.0001 in men) and with changes in serum total cholesterol (P=0.008 in women, P=0.02 in men), after adjustment for age, baseline level of each cardiovascular risk factor and current treatments. For example, in men, a weight loss of 5 kg was associated with a decrease of 2.5 mmHg in systolic blood pressure and of 1.5 mmHg in diastolic blood pressure. Taking into account self reported dieting did not modify these associations. CONCLUSIONS: In both genders, systolic and diastolic blood pressure and serum cholesterol increased with weight gain and decreased with weight loss, independently of the intentionality to lose weight. At the population level, modest weight loss may have a substantial impact on cardiovascular risk, and preventing even modest weight gain in adults is an important goal in terms of public health. PMID- 12119584 TI - [Well-informed on health matters--how well? The German 'Clearinghouse for Patient Information'--objective, background and methods]. AB - OBJECTIVE: Health information for lay people in print or electronic form are internationally recognised as useful tools and as necessary in the decision making process of individuals. The effectiveness of offered patient information depends on quality and accessibility. Because much of the available health information is significantly deficient, the Agency for Quality in Medicine developed a programme for assessing the quality of specialised health and medical information for all non-medically trained persons. The German 'Clearinghouse for Patient Information' project is an adjunct to the already established German 'Clearinghouse for Clinical Guidelines'. METHOD: The basis for quality improvement for specialised medical information consists 1. in the determination of demands on quality for patient information published in the German language, 2. development of a standardised instrument for assessing the quality of specialised medical information by non-medically trained persons, 3. establishment of an Internet portal for evaluated and rated material, and 4. the establishment and organisation of relevant partnerships with information providers and users of health and medical information. The principle building block to rate medical information for lay people are the DISCERN criteria for good patient information. The results of the quality rating of specialised medical information are made available to the public with emphasis on promoting transparency of the assessment and rating processes used. RESULTS: The Internet based patient information service of the Agency for Quality in Medicine (www.patienten-information.de 19.06.02) has currently a selection of approximately 800 information items covering fifty disease topics with fully or partially evaluated and rated information. Partnerships to appropriate information providers and users are already established and have been tested in special single projects. CONCLUSION: To advance the process of quality improvement for medical information for non-medical and lay persons, it is essential to develop and to intensify the cooperation with stake holders and partners at different levels. Only through close cooperation with others it is possible to create a basis to continuously develop and improve the information quality assessment instrument further. In addition, effective strategies need to be developed to disseminate these endeavours to patients and consumers of health information in a user-friendly and transparent way. PMID- 12119585 TI - [Conclusions from the Viagra case? Definition of disease in social legislation as exemplified by erectile dysfunction]. AB - The article describes the controversy about the question whether statutory social health insurances are obliged to reimburse the costs for the treatment of erectile dysfunction. Next to the question whether the 'Bundesausschuss der Arzte und Krankenkassen' was entitled to decide it was highly controversial whether erectile dysfunction is a disease according to the laws of social insurance. This enforces more general considerations regarding the possibility to define disease and the relevancy of a concept of disease for the justification and limitation of socially financed services in medicine. PMID- 12119586 TI - [Social inequalities and distribution of the environmental burden among the population (environmental justice)]. AB - The paper deals with the following question: How are the environmental risks distributed across different social groups? Using the term 'environmental justice', it has been discussed in the USA for more than 15 years already that environmental risks are not distributed evenly (i.e., justly). Public attention concentrates on the high environmental burden of the black community, but differences by social status are also addressed. In Germany, there is as yet no comparable discussion. In order to contribute to its development, we focus mainly on the following topics: empirical data from Germany on socio-economic differences in environmental risks concerning the home and the home environment, combination with the discussion on environmental justice, recommendations for research and health policy. The results indicate that also in Germany the lower status groups are exposed to greater environmental risks, and that regional measures of health promotion provide a good chance of reducing this inequality. PMID- 12119587 TI - [Socio-economic differences in the utilisation of screening programmes and health promotion measures in North Rhine-Westphalia, Germany]. AB - This paper examines the effect of socio-economic status on the utilisation of screening programmes and health promotion measures, based on data of a representative health survey in North Rhine-Westphalia (Germany), with a sample of 1,920 respondents. The analysis focuses especially on the utilisation of the cancer screening programme and the 'Health Check-Up' programme, which both have a high rank in German secondary prevention strategies. During the study year and the year before 29.2 % of the relevant male and 56.5 % of the female population took part in cancer screening programmes. At the same time 29.6 % of the males and 30.1 % of the females took part in the 'Health Check-Up' programme. In respect of socio-economic differences, the data confirmed the results of former studies in which non-participation of cancer screening programmes increased with decreasing social class. The social gradient was more pronounced with women than with men. While for the 'Health Check-Up' no socio-economic differences could be found, the participation rates in health promotion measures for women varied significantly with socio-economic status. For both screening programmes generally weaker statistical associations were found if the socio-economic status index was substituted by the educational level of the respondents. The findings suggest that strategies designed to increase participation of such programmes should concentrate more on lower social status persons to prevent the development of inequalities in health. PMID- 12119588 TI - [Expertising according to the federal German social welfare legislation and child and adolescent welfare laws by public health offices--results of an inquiry]. AB - Between December 2000 and February 2001 a survey among public health offices was conducted on the practice of expert reports on recipients of income support. More than 50 % (232) of the German public health offices participated; they carried out 164.000 of these reports in 1999. The analysis of the data shows considerable differences between the states (Bundeslander) concerning reasons and practice of expert reports which cannot be explained by a different social structure of population but indicate that quality of orders from Social Services and the work process in Public Health Offices may be inadequate. The findings result in recommendations for social services and public health offices to develop standards and improve quality, which can help to achieve greater equality. PMID- 12119589 TI - [Health behaviour and health status of nursing staff--a review of the literature]. AB - Presently around 1.1 million nurses are employed in the German health care system. Due to increased workload and staff reduction, studies on the health behaviour and health condition of nursing staff are increasingly important. Hence we reviewed the literature on health behaviour and health condition of nursing staff. Articles available in Medline and Embase from 1978 to 2000 were included, focussing on smoking, alcohol consumption, substance abuse, eating habits, body mass index, dental health status, risk behaviour in traffic, workload, burnout syndrome, sports and recreation, job satisfaction, subjective health status, subjective complaints, vaccinations and quality of life. Health behaviour was divided in unhealthy and healthy behaviour. It became obvious that most of the studies focussed on investigation of unhealthy behaviour, such as smoking, alcohol consumption, substance abuse and poor eating habits. Health promotion was mainly seen as avoiding these unhealthy habits. Only in current studies definitions of health promoting behaviour were developed as a part of a comprehensive life-style. PMID- 12119590 TI - [Execution and quality of attention paid to medical doctoral theses at the faculty of medicine at the University of Wurzburg as seen by the doctoral candidates in their 5th and 6th year of study]. AB - BACKGROUND AND OBJECTIVE: The significance of a doctoral thesis in medicine has been discussed controversially among medical students and faculty members. We examined in a brief but comprehensive study how medical students evaluate their research activities and whether or not alternative thesis models should be implemented. METHODS: A questionnaire comprising 50 questions was evaluated from 160 5th and 6th year medical students at the University of Wuerzburg. RESULTS: Almost two-thirds of our interviewees started working on a thesis project at the beginning of the 4th year of study. 59 % of our participants reported to have neglected the regular study and 56 % said to have prolonged the regular study due to the work on a thesis. Despite considerable variation, the median time spent weekly on a thesis project was 10 hours, independent of where the students worked. The median grade of satisfaction during the whole thesis was rated at 6.5 as assessed on an analogous scale from 1-10 (very satisfied). The research results from 94 students contributed to 164 articles (already published or in press) and the results of 110 students were presented at scientific meetings. Only 50 % of our interviewees were pleased with their published output and the importance of student research activities for the overall German biomedical research was rated 4.4 as assessed on an analogous scale from 1-10 (very important). 70 % of our interviewees preferred defined research periods and 47 % would apply for dual-degree research programmes. The scientific significance of a MD thesis in comparison to a PhD thesis was rated at 2.1 on an analogous scale from 1-5 (5 = equally significant). CONCLUSION: The current German research model might be modified and defined research periods and dual-degree research programmes as a one possible alternative should be considered. PMID- 12119592 TI - Physical and social predictors for pre-term births and low birth weight infants in Taiwan. AB - The purpose of this study was to examine the risk factors associated with pre term labor (PTL) (< 37 gestational weeks) and low birth weight (LBW) (< 2500 gm) infants in a healthy Taiwanese population. From December 1998 through June 1999, a total of 633 healthy pregnant women were recruited at three teaching hospitals in Taipei. Using a prospective study design, the pregnancy outcome information was followed up by telephone or from medical records during the first month postpartum. Data were statistically analyzed by multiple logistic regression. The prevalence of premature births was 5.4%, and the prevalence of LBW infants was 5.1%. Pre-term births were significantly associated with high self-reported fatigue scores (OR = 3.45); extreme maternal age (< 20 and >/= 35 years, OR = 2.38); history of abortion (>/= 2, OR = 3.11); maternal height (.05). Subjects with spouses as their main caregivers had significantly better improvement in QOL after pacemaker implantation. Subjects perceptions of distress from arrhythmia were the most significant determinant in their QOL pre and post pacemaker implantation. The findings suggest that aggressively resolving arrhythmia distress is important for improving QOL. PMID- 12119600 TI - Cost and care quality between licensed nursing homes under different types of ownership. AB - In Taiwan, there is some uncertainty and concern regarding the quality and safety of unlicensed nursing homes, as they are typically crowded and poorly equipped. There are data insufficient regarding the quality of care in licensed nursing homes for the government to reliably assist unlicensed facilities to become licensed. The purpose of this study was to examine the relationship between the different nursing home ownership types and the following dependent variables: (1) operating cost per resident day, (2) RN to resident ratio, (3) facility size, (4) occupancy rate, and (5) quality of care amongst licensed nursing homes nationwide. The descriptive study used a survey design. Data were obtained from 28 licensed nursing homes using self-administered questionnaires, on-site interviews and record reviews. Data were analyzed by Kruskal-Wallis test, Mann Whitney U test and Spearman s correlation. A positive and significant relationship existed between nursing home quality and the RN ratio per resident day. Chain/For-profit and Chain/Non- profit nursing homes tended to have higher operating costs and a better quality of service. Secondary research is still needed to examine the results by detailed cost analysis or by research oriented toward outcomes of residents care. These findings provide basic reference for the government for planning the operation of nursing home facilities and also to assist the many unlicensed nursing homes to ultimately become licensed. The results also present important data for developing reimbursement policies. PMID- 12119601 TI - Genomewide linkage disequilibrium mapping of severe bipolar disorder in a population isolate. AB - Genomewide association studies may offer the best promise for genetic mapping of complex traits. Such studies in outbred populations require very densely spaced single-nucleotide polymorphisms. In recently founded population isolates, however, extensive linkage disequilibrium (LD) may make these studies feasible with currently available sets of short tandem repeat markers, spaced at intervals as large as a few centimorgans. We report the results of a genomewide association study of severe bipolar disorder (BP-I), using patients from the isolated population of the central valley of Costa Rica. We observed LD with BP-I on several chromosomes; the most striking results were in proximal 8p, a region that has previously shown linkage to schizophrenia. This region could be important for severe psychiatric disorders, rather than for a specific phenotype. PMID- 12119602 TI - Major recessive gene(s) with considerable residual polygenic effect regulating adult height: confirmation of genomewide scan results for chromosomes 6, 9, and 12. AB - Segregation and linkage analyses were performed for adult height in a population of 200 Dutch families, each of which was ascertained through a proband with asthma. The best-fit model from the segregation analysis was a major recessive gene with a significant residual polygenic background. Models without a polygenic component were rejected. A genomewide scan was performed, and it confirmed previous linkage results for chromosomes 6q25 (LOD = 3.06, D6S2436), 9p1 (LOD = 2.09, D9S301), and 12q1 (LOD = 1.86, D12S375). Our results provide evidence that a combination of segregation and linkage approaches is valuable in understanding genetic determination of common complex traits. PMID- 12119604 TI - Antiviral market overview. PMID- 12119605 TI - Strategies in the design of antiviral drugs. AB - A decade ago, just five drugs were licensed for the treatment of viral infections. Since then, greater understanding of viral life cycles, prompted in particular by the need to combat human immunodeficiency virus, has resulted in the discovery and validation of several targets for therapeutic intervention. Consequently, the current antiviral repertoire now includes more than 30 drugs. But we still lack effective therapies for several viral infections, and established treatments are not always effective or well tolerated, highlighting the need for further refinement of antiviral drug design and development. Here, I describe the rationale behind current and future drug-based strategies for combating viral infections. PMID- 12119606 TI - Drug discovery by dynamic combinatorial libraries. AB - Dynamic combinatorial chemistry is a recently introduced supramolecular approach that uses self-assembly processes to generate libraries of chemical compounds. In contrast to the stepwise methodology of classical combinatorial techniques, dynamic combinatorial chemistry allows for the generation of libraries based on the continuous interconversion between the library constituents. Spontaneous assembly of the building blocks through reversible chemical reactions virtually encompasses all possible combinations, and allows the establishment of adaptive processes owing to the dynamic interchange of the library constituents. Addition of the target ligand or receptor creates a driving force that favours the formation of the best-binding constituent--a self-screening process that is capable, in principle, of accelerating the identification of lead compounds for drug discovery. PMID- 12119603 TI - A major susceptibility gene for asthma maps to chromosome 14q24. AB - Asthma is a complex genetic disorder with a heterogeneous phenotype, largely attributed to the interactions among many genes and between these genes and the environment. Numerous loci and candidate genes have been reported to show linkage and association to asthma and atopy. Although some studies reporting these observations are compelling, no gene has been mapped that confers a sufficiently high risk of asthma to meet the stringent criteria for genomewide significance. Using 175 extended Icelandic families that included 596 patients with asthma, we performed a genomewide scan with 976 microsatellite markers. The families were identified by cross-matching a list of patients with asthma from the Department of Allergy/Pulmonary Medicine of the National University Hospital of Iceland with a genealogy database of the entire Icelandic nation. We detected linkage of asthma to chromosome 14q24, with an allele-sharing LOD score of 2.66. After we increased the marker density within the locus to an average of one microsatellite every 0.2 cM, the LOD score rose to 4.00. We designate this locus "asthma locus one" (AS1). Taken together, these results provide evidence of a novel susceptibility gene for asthma on chromosome 14q24. PMID- 12119607 TI - Navigating the evolving world of drug discovery. AB - The successful translation of basic scientific discoveries into novel therapeutic strategies for the prevention and treatment of disease depends on integrating all aspects of the drug discovery and development process. PMID- 12119608 TI - The genetic basis of variability in drug responses. AB - It is almost axiomatic that patients vary widely in their beneficial responses to drug therapy, and serious and apparently unpredictable adverse drug reactions continue to be a major public health problem. Here, we discuss the concept that genetic variants might determine much of this variability in drug response, and propose an algorithm to enable further evaluation of the benefits and pitfalls of this enticing possibility. PMID- 12119609 TI - High-throughput crystallography for lead discovery in drug design. AB - Knowledge of the three-dimensional structures of protein targets now emerging from genomic data has the potential to accelerate drug discovery greatly. X-ray crystallography is the most widely used technique for protein structure determination, but technical challenges and time constraints have traditionally limited its use primarily to lead optimization. Here, we describe how significant advances in process automation and informatics have aided the development of high throughput X-ray crystallography, and discuss the use of this technique for structure-based lead discovery. PMID- 12119610 TI - Emerging immune targets for the therapy of allergic asthma. AB - Recent discoveries on the molecular and cellular basis of asthma have markedly altered our understanding of this common respiratory disorder. These insights have come during an unexplained period of rising disease incidence and severity and are now being applied to develop improved therapies. This review explores the latest advances in our understanding of the pathogenesis of allergic asthma, and provides insight into the expanding collaborations between research scientists, clinicians and the pharmaceutical industry in the race to control the asthma epidemic. PMID- 12119612 TI - Moving smaller in drug discovery and delivery. AB - Advances in new micro- and nanotechnologies are accelerating the identification and evaluation of drug candidates, and the development of new delivery technologies that are required to transform biological potential into medical reality. This article will highlight the emerging micro- and nanotechnology tools, techniques and devices that are being applied to advance the fields of drug discovery and drug delivery. Many of the promising applications of micro- and nanotechnology are likely to occur at the interfaces between microtechnology, nanotechnology and biochemistry. PMID- 12119611 TI - Targeting glycosylation as a therapeutic approach. AB - Increased understanding of the role of protein- and lipid-linked carbohydrates in a wide range of biological processes has led to interest in drugs that target the enzymes involved in glycosylation. But given the importance of carbohydrates in fundamental cellular processes such as protein folding, therapeutic strategies that modulate, rather than ablate, the activity of enzymes involved in glycosylation are likely to be a necessity. Two such approaches that use imino sugars to affect glycosylation enzymes now show considerable promise in the treatment of viral infections, such as hepatitis B, and glucosphingolipid storage disorders, such as Gaucher disease. PMID- 12119613 TI - Toxicogenomics and drug discovery: will new technologies help us produce better drugs? AB - Acting on reports in the late 1980s that most drug candidates fail in development, pharmaceutical discovery programmes responded by devising ways to increase the number of chemicals in the pipeline. With discovery now driven primarily by chemistry and high-throughput screening, the biological effects and, in particular, the toxicity of new compounds are largely not appreciated until a compound enters development. Arguably, this paradigm has produced more failures rather than delivering more successes--with more chemicals to examine, much less is known about any single agent before costly development studies are initiated. The emerging field of toxicogenomics is enabling us to ask detailed questions about drug effects very early on, thereby fundamentally changing our approach to drug discovery. PMID- 12119614 TI - Building bridges: affirming culture in health and nursing. PMID- 12119616 TI - Drawing a qualitative distinction of caring practices in a professional context: the case of Chinese nursing. AB - How to draw a qualitative distinction between nursing work and the work of a servant has been a major concern for nurses in China. This article explains the ways in which nurses in China articulate the meaning of caring in practice situations. Seventy nurses in Beijing were invited to share their experience about what caring meant to them as nurses and examples of caring in practice situations. Van Kaam's phenomenologic method of controlled explication was used to analyze the data. The findings reveal that these Chinese nurses are able to articulate the cheng and jing versions of caring practices that emphasize flexible, pluralist, contextualized, individualized, and subjectively informed practices. To further this study, we would suggest nurses be more proactive in starting a dialogue with society so as to seek nursing's legitimate practice that can foster responsive care to patients and society on the one hand and the professional integrity of nursing on the other. PMID- 12119617 TI - Exploring culture in nursing: a theory-driven practice. AB - This article offers useful approaches for understanding culture as an important component for providing holistic care in nursing. Watson's Model of Human Care exemplifies a way to interpret the cultural experience of illness within a holistic framework, helping nurses explain and then detail alternative caring perspectives. A clinical exemplar using this framework will help to explore the interchange between caring and culture. In doing so, Watson's Model of Human Care can be shown to hold great promise for understanding human illness as a cultural event and contribute to holistic care in nursing practice. PMID- 12119618 TI - Describing an explanatory model of HIV illness among aboriginal women. AB - This article describes the explanatory model of human immunodeficiency virus (HIV) illness used by aboriginal women in northern Alberta. Using Kleinman's explanatory model framework, eight women who were HIV positive were interviewed to determine their perspectives on the etiology, pathophysiology, symptomology, course of illness, and methods of treatment for HIV. A comparative analysis was done between the explanatory model of HIV illness as described by participants and the conventional biomedical paradigm of HIV disease. As described by aboriginal women, several aspects of the explanatory model of HIV were congruent with the biomedical paradigm. It was also found that the findings related to etiology and treatment of HIV illness was incongruent with the conventional biomedical paradigm of HIV disease. These findings highlight the relevance of knowing models of illness for health care professionals, particularly nurses who work in communities with a high incidence of HIV/AIDS. These models make care planning of patients with HIV and AIDS more focused and directed. PMID- 12119619 TI - Cultural affirmation and the protection of emotional well-being. AB - This article examines the impact of the Balkan conflict on the culture and emotional health of a community of Serbian Australians. It discusses how an intimate reconnection with their cultural identity, Serbian Australians, without formal mental health service supports, managed the complex and dynamic interplay between homeland events, mainstream media reports, ethnonational bonds, and mental health issues in Australia. Ethnographic techniques revealed that although the Balkan conflict bared a multitude of potent health and emotional concerns for Serbian Australians, their coping was enhanced by an intimate sense of belonging and reassociation with their historical, religious, cultural, and national identities. By engaging in spiritual connections with their culture and ethnicity, the transglobal effects of the Balkan war on Serbians in Australia revealed that mental health and healing could no longer be seen as a localized phenomenon. It must also be seen as something that transcends the nations and communities in which people live. PMID- 12119615 TI - Holism and caring: nursing in the Chinese health care culture. AB - This article examines the concepts of holism and caring within the historical and cultural context of the development of nursing in China. Data were collected in two research projects on the cultural construction of caring in nursing within different Chinese communities. Grounded in the theory of "systematic correspondence" and in Chinese medical tradition, illness is perceived as a state of disharmony between the individual and the natural and social environment. The interplay of caring and curative processes is seen as pertinent to health maintenance. In Chinese culture, it is the moral duty of family members to take care of their sick. Because of this, questions are posed about ways in which the nurse can relate to patients and their family members. Although lay people view nursing as servant's work, nurses cherish "treating the patient as their own family member." The authors contend that these versions of nursing are inappropriate. It is the intention of this article to stimulate discourse on a third version focusing on the patient and the family as the unit of care within their dynamic, natural, and social environments. PMID- 12119620 TI - Effects of psychological distress on blood pressure in adolescents. AB - This cross-sectional survey measured relationships among blood pressure and measures of psychologic distress, family structure, and economic status in a sample of adolescents exposed to Hurricane Hugo. Spielberger's Anger Scale and Derogatis' Brief Symptom Inventory were used. Data analysis revealed 5% of the 1079 adolescents were hypertensive. Multiple regression analyses revealed the following predictors of higher diastolic blood pressure: African-American race, recipient of subsidized lunch, exposure to Hurricane Hugo, and higher anger-in scores in males. The effects of a catastrophic event such as a hurricane on blood pressure and the effects of introjected anger have implications for both health care consumers and providers. PMID- 12119621 TI - Alternative and complementary therapies for labor and birth: an application of Kolcaba's theory of holistic comfort. AB - Although nursing has always used nonpharmacologic interventions for the relief of the discomforts of childbirth, alternative and complementary therapies are becoming more acceptable. Alternative and complementary therapies are based on a balance of body, mind, and spirit. Kolcaba's theory of holistic comfort is proposed as a framework for guiding nurses to use alternative and complementary therapies in the comfort care of laboring women. PMID- 12119622 TI - Acupuncture and oriental body work: traditional and modern biomedical concepts in holistic care--conceptual frameworks and biomedical developments. AB - An introduction to the history and basic concepts underlying acupuncture and Oriental body work was presented in the April 2000 (14:3) issue of Holistic Nursing Practice. The functioning of Yin and Yang organs and meridians are addressed in this article, along with the diagnostic frameworks of Five Elements or Phases and Eight Principle Patterns. Contemporary biomedical developments in the use of acupuncture are discussed, along with research on the endorphin system as a biomediary of acupuncture. Possible applications to nursing are discussed. PMID- 12119623 TI - Therapeutic touch coming of age. AB - Therapeutic Touch, a meditative healing practice created by Dolores Krieger and Dora Kunz, is adapted from the "laying on of hands" for the purpose of helping or healing others. The history of the technique and its influence on the health care system are chronicled in an effort to establish the role Therapeutic Touch has played in transforming people's lives. PMID- 12119624 TI - Use of therapeutic touch in treatment of drug addictions. AB - The purpose of this pilot study is to examine the efficacy of Therapeutic Touch (TT) as a form of treatment intervention with persons who abuse alcohol and/or other drugs. A between-subjects design compared treatment outcomes of three groups of alcohol and other drug abusers over five months. The Addiction Severity Index (ASI), Michigan Alcoholism Screening Test (MAST), Beck Depression Inventory (BI), and Personal Experiences: Therapeutic Touch and Frequency of Alcohol/Drug Use questionnaires were administered to the group receiving TT (Group A) and both control groups (Mimic TT Group B and No Intervention Group C) at various points in the study. Preliminary findings indicate that the use of TT could be effective in prolonging periods of abstinence for alcohol and other drug abusers. PMID- 12119625 TI - The empowering nature of Reiki as a complementary therapy. AB - Reiki is an ancient healing method with roots in both Chinese Medicine and Christian healing. It is a treatment used by individuals as an alternative and complement to Western medical treatment. Reiki has increased in popularity over the past decade, but remains understudied. Methodological and philosophical reasons for why it is difficult to conduct research on the efficacy of Reiki are discussed. The reasons for the increased success of Reiki as an alternative and complementary healing method in the Western world are addressed, as well as the practice of Reiki as a healing method for self and others. PMID- 12119626 TI - Relaxation training as a holistic nursing intervention. AB - For centuries cultures have incorporated strategies that recognize the power of engaging the mind in the process of healing. In this century, relaxation training (RT) is a skill that has been repeatedly validated by nursing, medical and psychology researchers as a complementary intervention that is effective for a wide range of clinical situations. Relaxation can be employed by nurses and patients to offset the negative effects of stress, illness, and surgery while promoting healing and self efficacy. The nurse's knowledge of RT is essential to a holistic nursing approach that supports an innovative model of caring for nursing practice. PMID- 12119627 TI - Prayer: an ancient healing practice becomes new again. AB - Prayer is an ancient healing practice not generally available in our health care system. However, the majority of Americans believe in the healing power of prayer. A recent Time/CNN poll found that 82% of Americans believe that prayer can cure serious illness, 73% believe that praying for others can cure illness, and 64% want their physicians to pray with them. Nurses should know what prayer is, should be familiar with the growing body of literature on the effectiveness of prayer, and understand potential uses of prayer as part of holistic nursing practice. PMID- 12119628 TI - Alternative therapies and nurse practitioners: knowledge, professional experience, and personal use. AB - An anonymous questionnaire was mailed to all members of the Connecticut Nurse Practitioner Group, Inc. to identify their knowledge, and personal and professional experiences with alternative therapies. Of the 202 respondents (a 73% return), 78% described themselves as "slightly" or "quite" knowledgeable about alternative therapies in general and 63% reported personal experience. Approximately half of the nurse practitioners (NP) (49%) indicated they "sometimes" or "usually" ask about their clients' use of such therapies and 65% have referred for or recommended one or more of these modalities. Additionally, some NPs have provided alternative therapies directly (30%). One third (31%) indicated they had received training in one or more therapy(ies) and 65% would be "extremely" or "quite" interested in learning more. PMID- 12119630 TI - Acupuncture and Oriental body work: traditional and biomedical concepts in holistic care: history and basic concepts. AB - Acupuncture and related forms of Oriental body work, such as shiatsu and acupressure, have become increasingly part of the complementary and alternative health care in the last twenty years in the United States. Yet most consumers and professionals, nurses included, know little or nothing about the philosophical and diagnostic underpinnings of these forms of treatment. The frameworks underlying these modalities are intrinsically holistic, and provide an interesting model for nurses and others who are looking for different ways to think about data from patient care. Part I of two articles presents descriptions of practices, and an introduction to the history and basic concepts underlying Chinese acupuncture and related forms of Oriental body work. PMID- 12119629 TI - Therapeutic use of selected herbs. AB - An increasing number of people in the United States are using herbs for health promotion and specific symptom management. Herbs are used to initiate healing through synergistic responses unlike the specific properties of pharmaceuticals. Anecdotal data comprise much of the popular information available about herbs. Scientific studies of the efficacy and safety of herbs, although on the rise, are less available than other drug trials. Clinicians need an appropriate knowledge base for dealing with patients who take herbal preparations as well as the ability to confidently include herbal preparations in their formulary. In this article, five common herbs are reviewed. The effects, clinical studies, side effects, and dosing regimens for aloe vera, arnica, black cohosh, evening primrose oil, and saw palmetto are described. PMID- 12119631 TI - The silent revolution in the use of complementary and alternative therapies continues in the United States. PMID- 12119632 TI - Why write review book chapters? PMID- 12119633 TI - Prevention of obesity--is it possible? AB - Obesity prevention is necessary to address the steady rise in the prevalence of obesity. Although all experts agree that obesity prevention has high priority there is almost no research in this area. There is also no structured framework for obesity prevention. The effectiveness of different intervention strategies is not well documented. Regarding universal prevention little rigorous evaluation has been carried out in larger populations. Obesity prevention has been integrated into community-wide programmes preventing coronary heart disease. Although effective with respect to reduction in cardiovascular risk factors these programmes did not affect mean body mass index (BMI) of the target populations. Selective prevention directed at high risk individuals (e.g. at children with obese parents) exhibited various degrees of effectiveness. However, at present, definitive statements cannot be made because of the limited number of studies as well as limits in study design. Finally, targeted prevention produced promising results in obese children when compared to no treatment. However, there are only very few longterm follow-up data. There is no clear idea about comprehensive interventions studying combinations of different strategies. It is tempting to speculate that predictors of treatment outcome (e.g. psychological and sociodemographic factors) may also serve as barriers to preventive strategies, but this has not yet been investigated. Taken together, obesity prevention should become a high priority research goal. First results of obesity prevention programmes are promising. As well as health promotion and counselling, better school education and social support appear to be promising strategies for future interventions. PMID- 12119634 TI - Clinical aspects of obesity in childhood and adolescence. AB - The level of fatness of a child at which morbidity acutely and/or later in life increases is determined on an acturial basis. Direct measurements of body fat content, e.g. hydrodensitometry, bioimpedance, or DEXA, are useful tools in scientific studies. However, body mass index (BMI) is easy to calculate and is generally accepted now to be used to define obesity in children and adolescents clinically. An increased risk of death from cardiovascular disease in adults has been found in subjects whose BMI had been greater than the 75th percentile as adolescents. Childhood obesity seems to substantially increase the risk of subsequent morbidity whether or not obesity persists into adulthood. The genetic basis of childhood obesity has been elucidated to some extent through the discovery of leptin, the ob gene product, and the increasing knowledge on the role of neuropeptides such as POMC, neuropeptide Y (NPY) and the melanocyte concentrating hormone receptors (for example, MC4R). Environmental/exogenous factors largely contribute to the development of a high degree of body fatness early in life. Twin studies suggest that approximately 50% of the tendency toward obesity is inherited. There are numerous disorders including a number of endocrine disorders (Cushing's syndrome, hypothyroidism, etc.) and genetic syndromes (Prader-Labhard-Willi syndrome, Bardet Biedl syndrome, etc.) that can present with obesity. A simple diagnostic algorithm allows for the differentiation between primary or secondary obesity. Among the most common sequelae of primary childhood obesity are hypertension, dyslipidemia, back pain and psychosocial problems. Therapeutic strategies include psychological and family therapy, lifestyle/behaviour modification and nutrition education. The role of regular exercise and exercise programmes is emphasized. Surgical procedures and drugs used in adult obesity are still not generally recommended in children and adolescents with obesity. As obesity is the most common chronic disorder in industrialized societies, its impact on individual lives as well as on health economics has to be recognized more widely. This review is aimed towards defining the clinical problem of childhood obesity on the basis of current knowledge and towards outlining future research areas in the field of energy homoesostasis and food intake in relation to child health. Finally, one should aim to increase public awareness of the ever increasing health burden and economic dimension of the childhood obesity epidemic that is present around the globe. PMID- 12119635 TI - The peripheral sympathetic nervous system in human obesity. AB - The peripheral sympathetic nervous system is a key factor in the regulation of energy balance in humans. Differences in sympathetic nervous system activity may contribute to variations in 24 h energy expenditure between individuals. beta Adrenoceptors play a more important role than alpha-adrenoceptors in this regulation. The involvement of both beta 1- and beta 2-adrenoceptor subtypes has been demonstrated, the role of the beta 3-adrenoceptor subtype is not yet clear. Normal or increased levels of sympathetic nervous system activity and reduced reactivity appear to be present in established obesity. Furthermore, the sensitivity for beta-adrenoceptor stimulation is impaired in obesity. The blunted reactivity and sensitivity may contribute to the maintenance of the obese state. There are data to suggest that they may also play a role in the aetiology of obesity, because the impairments often remain after weight reduction. Furthermore, a negative correlation between baseline sympathetic nervous system activity and weight gain during follow-up has been found in Pima Indians. Recently, genetic evidence about the involvement of adrenoceptors in obesity has become available. Although the results of association and linkage studies on polymorphisms in the beta 2-, beta 3- and alpha 2-adrenoceptor genes are inconsistent, the functional correlates of some of these polymorphisms (changes in agonist-promoted down-regulation, protein expression levels, lipolytic sensitivity, basal metabolic rate, sympathetic nervous system activity) suggest that they may be important in the aetiology of obesity. PMID- 12119636 TI - The health at any size paradigm for obesity treatment: the scientific evidence. AB - Traditional weight loss (TWL) treatments have been unsuccessful at reducing the prevalence of obesity in the population. Health-care professionals and consumers have criticized TWL treatments as being detrimental to the obese person's health. Consequently, an alternative approach to obesity treatment, the health at any size (H@AS) paradigm, has been proposed. The H@AS paradigm is based on the philosophy that once diet restrictions and barriers to activity have been removed, the individual will develop healthier eating and activity patterns that lead to a naturally healthy body weight. This paper reviews the philosophical foundation and the scientific data that support and oppose the H@AS paradigm and compares it with that of TWL treatments. PMID- 12119637 TI - Insulin resistance and associated metabolic abnormalities in muscle: effects of exercise. AB - Skeletal muscle is a major site of insulin resistance. In addition to glucose transport, oxidative disposal and storage defects, insulin resistant muscle exhibit many other metabolic abnormalities. After a brief review of insulin resistance determinants, we will focus on muscular abnormalities in obesity and type 2 diabetes. Glucose and lipid metabolism defects will be analysed and their interactions discussed. Exercise can improve many of these muscular abnormalities and the mechanisms underlying exercise-induced benefits have been clarified during the past decades. Therefore, exercise training has proved to be useful in the management of insulin resistant states, i.e. mainly obesity, especially in its truncal distribution, and type 2 diabetes. However, exercise prescription remains poorly codified, and results on glycaemic control are sometimes conflicting. In the last part of this review, we will emphasize the pathophysiological basis for an individualized exercise prescription in insulin resistant subjects. PMID- 12119639 TI - Why do we need an obesity review journal year 2000? PMID- 12119638 TI - Very low energy diets in the treatment of obesity. AB - Very low energy diets (VLEDs) are defined as diets which contain energy levels of less than 3.4 MJ (800 kcal) per day and contain daily allowances of all essential nutritional requirements. These diets have been in clinical use for more than 20 years. They are used as the only source of nutrition for 8-16 weeks, which usually achieves a weight loss of 1.5-2.5 kg per week. Before using this type of diet a medical investigation is necessary to evaluate contraindications and to check medication use during the diet. To facilitate maintenance, cognitive behavioural counselling should always be included in a weight reduction programme using a very low energy diet. VLEDs have no serious harmful effects and can safely be used in patients with various chronic diseases. Programmes using VLEDs produce better short-term weight loss than programmes without the diet. However, in randomized controlled trials VLED-based programmes have not achieved significantly better long-term maintenance than conventional programmes. VLEDs are used when rapid weight loss is necessary because of an obesity-related disease. In other patients with obesity it is an alternative to other conservative approaches for treatment of obesity. In type 2 diabetes it may improve long-term glucose metabolism better than conventional weight reducing diets. Some studies suggest that after a VLED-based programme long-term maintenance is better among men than women. This possible gender difference is an important topic for further research. PMID- 12119640 TI - Lessons from obesity management programmes: greater initial weight loss improves long-term maintenance. AB - It is a common belief that weight loss achieved at a slow rate is better preserved than if the weight is lost more rapidly. However, the literature shows that initial weight loss is positively, not negatively, related to long-term weight maintenance. There is evidence from randomised intervention trials to support that a greater initial weight loss induced without changes in lifestyle (e.g. liquid formula diets or anorectic drugs) improves long-term weight maintenance, providing it is followed by a 1-2 years integrated weight maintenance programme consisting of lifestyle interventions involving dietary change, nutritional education, behaviour therapy and increased physical activity. In conclusion, we find evidence to suggest that a greater initial weight loss as the first step of a weight management programme may result in improved sustained weight maintenance. PMID- 12119641 TI - Predictors of weight gain: the biological-behavioural debate. AB - The rapidly increasing prevalence of obesity, in spite of an unchanged gene pool, makes it interesting to search for biological factors which increase the susceptibility at the individual level as well as searching for the responsible environmental factors. Among the identified metabolic factors is a low resting metabolic rate for given body size and composition, a high respiratory quotient (RQ) indicating a low fat oxidation and a low spontaneous physical activity, all factors which are regarded as being under substantial genetic influence. Among the environmental factors, it is low levels of physical activity, increasing inactivity and a high fat diet that are probably the most important ones. In this review we have focused on controversies in this area. Understanding the interaction between the constitutional biological factors and the environmentally determined lifestyle factors it is important to produce better options for both the prevention and treatment of obesity. PMID- 12119642 TI - Obesity: a disease or a biological adaptation? AB - The increase in obesity prevalence is problematic as this condition is associated with health complications such as diabetes and cardiovascular diseases, more particularly when the excess body fat is stored in the deep abdominal region. On the other hand, obesity facilitates the maintenance of body homeostasis probably because of an increased hormonal gradient which favours the regulation of energy balance, to give but one example. The regulation potential of excess body fat is particularly apparent in the reduced-obese state where a reduction of energy expenditure, fat oxidation and some immune system markers, as well as an increase in appetite, stress vulnerability and circulating and adipose tissue organochlorines have been observed. These constitute another category of risk factors which can certainly favour the accumulation of body fat to reestablish body homeostasis on other fronts. Under such conditions, obesity is perceived by the physiologist as a necessary biological adaptation rather than a disease. For health professionals, this emphasizes the importance to seek a reasonable compromise between the favourable reduction of risk to develop metabolic complications by body weight loss and the physiological vulnerability which is also generated by such an intervention. PMID- 12119643 TI - Preventing obesity--in theory and practice? PMID- 12119644 TI - Neuroendocrine mechanisms regulating food intake and body weight. AB - In the field of obesity research, two separate lines of study have emerged which explore the mechanism by which food intake is regulated: short-term control of food intake, and the central regulation of energy balance. The former studies the satiety response during consumption of meals, whereby satiety signalling originating in the gut is transduced into a neural signal that modulates satiety pathways in the brainstem. This review describes a neuroanatomically based model in which leptin and insulin signalling in the hypothalamus governs long-term regulation of energy balance via mechanisms that are integrated with satiety hormone signalling in the brainstem. The functional outcome of this integration is a cumulative meal-to-meal regulation of food intake, that over relatively long intervals serves to maintain stable adipose stores. Our model provides a context within which continued investigation of neuroendocrine mechanisms that control food intake and body weight can be explored, and has potential application to our current understanding of clinical obesity and its treatment. PMID- 12119645 TI - Visceral obesity and metabolic syndrome. AB - There is increasing evidence for the existence of a condition consisting of a cluster of metabolic disorders which include insulin resistance, alterations in glucose and lipid metabolism, increased blood pressure and visceral obesity. The metabolic syndrome is now the favoured definition of the cluster. Each single component of the cluster increases the cardiovascular risk, but the combination of factors is much more important. Insulin resistance is the most frequently associated factor to the singular components of the syndrome: most authors believe that it may be the common aetiological factor. However, visceral obesity seems to be the main driving factor by means of the increased production of free fatty acids whose activity, in turn, might interfere with the action of insulin. Some questions exist about the syndrome because of the frequent lack in the cluster of one of the factors. This does not mean that the missing factor does not belong to the syndrome, but only that it is not yet clinically evident. Weight gain has been shown to be a strong predictor of the metabolic syndrome. This aspect gives strength to treatment and prevention because it means that losing weight or stopping weight increase might reduce the risk of a future appearance of a factor that is still not evident. Interventions to treat visceral obesity by means of losing weight seem to be the most efficacious way to treat the metabolic syndrome thus improving the most widespread cardiovascular risk factor in western countries. PMID- 12119646 TI - The sympathetic nervous system and obesity: role in aetiology and treatment. AB - The sympathetic nervous system (SNS) is an important component of the autonomic nervous system, and thus plays a major role in the maintenance of homeostasis. The SNS is of particular importance in the control of the cardiovascular system and of a number of metabolic processes. Alterations in SNS effects on metabolism have been implicated in the development and maintenance of obesity, and the SNS is a potential therapeutic target in the treatment of obesity. This review provides an overview of the anatomical and physiological aspects of the SNS, before considering the evidence showing a role for the SNS in the development or treatment of obesity. PMID- 12119647 TI - Transgenerational health promotion. AB - The Adult Health and Development Program (AHDP) is an intergenerational, interdisciplinary health promotion and rehabilitation program in which nursing and other college students are paired with older adults to engage in activities to improve their health and well-being. The purpose of this article is to describe resources and strategies used to implement a new program. The University of Delaware's program uses the ACAEM Paradigm developed by the AHDP at the University of Maryland where the program has existed for 27 years. However, the University of Delaware emphasizes the concepts of transition as described by Schumacher and Meleis and adult learning theory. Older adults are often the most open to education and other forms of support when they need help in making a change or transition in their lives. Schumacher and Meleis contend that transitions are a central concept in nursing and propose a model of transition in which education is the primary modality for preparing for transitions. The model also specifies ways to measure desirable transition outcomes. PMID- 12119648 TI - The social environment of nursing homes and the health of older residents. AB - This study investigated the effects of two social environment variables, social support and anomia, on the self-reported health of older nursing home residents. Three specific hypotheses were tested as well as the fit of the data to the proposed theory. A nonrandom, convenience sample of 91 nursing home residents was drawn from four nursing homes. Only whites who could speak and understand English and who were judged to be cognitively intact or only mildly cognitively impaired were included. The data were analyzed using path analysis. Only one hypothesis asserting that anomia will have direct negative effects on self-reported health was fully supported. PMID- 12119649 TI - Care of the older adult following hip fracture. AB - Proximal femoral (hip) fractures are the leading cause of hospitalization for injuries among older persons and constitute a serious health problem. Research shows significant functional losses among older persons following surgery for hip fracture related to a wide range of problems. Because of the physiological, emotional, social, and psychological complications inherent in this particular population, a team approach using a holistic model of care is efficacious when attempting to improve functional outcomes and reduce morbidity and mortality. PMID- 12119650 TI - Abuse of older persons: an overview. AB - As demographics in the United States indicate that by 2050 older persons will constitute one-fourth of the population, statistics indicating abuse of older persons will also increase. As a serious social problem, nurses will need to assess and intervene with families where this type of family violence occurs. The types of abuse of older persons include physical, psychological, sexual, and financial abuse; neglect, self-neglect; and other types such as violation of rights, denial of privacy, and denial of participation in decision making. This article examines types of abuse, who is at risk, identifying characteristics of abusers, and nursing assessment and nursing interventions related to abuse of older persons. PMID- 12119653 TI - Creating clinical experiences in a social work agency on aging. AB - This article describes the growth of a relationship between the fields of social work and nursing that developed into a positive learning experience for senior nursing students. It highlights the growing awareness, complicated by the scarcity of clinical sites, of the importance of including content in curricula related to community-dwelling older adults. The author was able to prepare student assignments from a social work agency on aging. The outcomes of this process included meeting the objectives of community health nursing and a new awareness by nursing students of the collaborative roles of nursing and social work in the community. PMID- 12119652 TI - Computers and caregiving: reaching out and redesigning interventions for homebound older adults and caregivers. AB - This article discusses computer resources for homebound older adults and informal caregivers as an intervention to promote social support and mental health. Published information related to a computer network designed as an intervention for informal caregivers of persons with Alzheimer's disease is included. This information suggests that homebound older adults and informal caregivers can gain valuable information, confidence, and support by using computer resources. A review of the literature supported those findings and suggested that computer technology can facilitate continuing education and the refinement of skills for nurses. Implications for the use of computer resources in nursing education, practice, and research are presented. PMID- 12119655 TI - Weight control, physical activity and cancer--strong links. PMID- 12119654 TI - A review of forgiveness literature with implications for nursing practice. AB - This article describes clinical practice where forgiveness is a central patient/family issue. In this case, the professional nurses learned substantively from the family about the tragic consequences of forgiveness withheld and the transformative nature of forgiveness extended. The concept of forgiveness is defined and forgiveness as a primary, secondary, and tertiary intervention is considered. Implications for professional practice are developed. Practical forgiveness-based assessment questions are included. A review of instrumentation from other disciplines measuring interpersonal dimensions of forgiveness is offered. Significant aspects of the process of forgiveness are elucidated and opportunities for forgiveness-related nursing research are identified. PMID- 12119656 TI - A role for olestra in body weight management. AB - Olestra is a fat substitute made from fatty acids esterified to sucrose and can be used in the preparation of virtually any food made with fat. Foods made with olestra retain the mouthfeel, palatability and satiating effects of their full fat counterparts without providing any digestible energy. Because olestra provides no energy, it has the potential to be a useful tool in weight loss and weight maintenance. Short-term studies of olestra replacement in foods demonstrate that fat replacement leads to a net reduction in fat intake. When excess total energy is available, fat replacement also reduces total energy intake in lean and obese men and women. In longer-term studies in which olestra is incorporated into the daily diet, there is an incomplete compensation for the fat energy replaced by olestra. When overweight men consumed olestra as part of a varied diet over nine months, weight loss continued for the duration of the study, whereas individuals receiving a typical low-fat diet regained most of the initial weight lost. Other studies are underway to examine the usefulness of olestra in long-term weight maintenance following weight loss. Post-marketing surveillance of olestra foods in the United States indicates that substitution of olestra for only 1-2 g of fat d-1 may be sufficient to prevent the average weight gain reported in adults of 0.5-1.0 kg year-1. PMID- 12119651 TI - Wholism for aging families: meeting needs of caregivers. AB - Family caregiving of a frail older person is an increasingly common phenomenon. Caregivers are confronted with new roles and responsibilities that provide both challenges and opportunities. Family issues that require expert nursing attention include role reversal, unresolved conflicts, caregiver immersion, elder mistreatment, and caregiving from a distance. Comprehensive geriatric assessment provides a foundation for intervening with families. Nurses are instrumental in providing a family perspective that meets the needs of frail older persons and their caregivers thus providing wholistic care. PMID- 12119657 TI - Hypothalamic obesity in humans: what do we know and what can be done? AB - Obesity is a common sequel to tumours of the hypothalamic region and their treatment with surgery and radiotherapy. The prevalence of hypothalamic obesity has been underestimated because it may take some years to develop, and the problem has been under-recognized by physicians. Weight gain results from damage to the ventromedial hypothalamus which leads, variously, to hyperphagia, a low metabolic rate, autonomic imbalance, growth hormone (GH) deficiency and various other problems that contribute to weight gain. However, with the exception of GH replacement, few clinical trials have evaluated significant numbers of patients and so the roles of various behavioural, dietary, pharmacological and obesity surgery approaches are controversial. Sufficient knowledge exists to identify those at high risk of hypothalamic obesity so that weight gain prevention approaches can be offered. In those who are already obese, we propose that the principal causal mechanisms in individual patients should be considered as a basis for guiding clinical management. PMID- 12119658 TI - The biomechanics of adiposity--structural and functional limitations of obesity and implications for movement. AB - Obesity is a significant health problem and the incidence of the condition is increasing at an alarming rate worldwide. Despite significant advances in the knowledge and understanding of the multifactorial nature of the condition, many questions regarding the specific consequences of the disease remain unanswered. For example, there is a dearth of information pertaining to the structural and functional limitations imposed by overweight and obesity. A limited number of studies to date have considered plantar pressures under the feet of obese vs. non obese, the influence of foot structure on performance, gait characteristics of obese children and adults, and relationships between obesity and osteoarthritis. A better appreciation of the implications of increased levels of body weight and/or body fat on movement capabilities of the obese would provide an enhanced opportunity to offer more meaningful support in the prevention, treatment and management of the condition. PMID- 12119659 TI - An updated systematic review of interventions to improve health professionals' management of obesity. AB - The objective of this article was twofold (1) to determine the existence and effectiveness of interventions to improve health professionals' management of obesity or the organization of care for overweight and obese people; and (2) to update a previous systematic review on this topic with new or additional studies. The study design was a systematic review of intervention studies, undertaken according to standard methods developed by the Cochrane Effective Practice and Organization of Care (EPOC) Group. Participants were trained health care professionals and overweight and obese patients. The measurements were objective measures of health professionals' practice and behaviours, and patient outcomes including satisfaction, behaviour, psychological factors, disease status, risk factors and measures of body weight, fat, or body mass index (BMI). Twelve studies were included in the original review. A further six were included in this update. Six of the 18 studies were randomized controlled trials of health professional-oriented interventions (such as the use of reminders and training) and one was a controlled before-and-after study to improve collaboration between a hospital clinic and general practitioners (GPs). Ten randomized controlled trials and two controlled clinical trials of interventions comparing either the deliverer of weight-loss interventions or the setting of the delivery of the intervention, were identified. The heterogeneity and generally limited quality of identified studies make it difficult to provide recommendations for improving health professionals' obesity management. To conclude, at present, there are few solid leads about improving obesity management, although reminder systems, brief training interventions, shared care, inpatient care and dietitian-led treatments may all be worth further investigation. Therefore, decisions for the improvement of provision of services must be based on the existing evidence on interventions with patients and good clinical judgement. Further research is needed to identify cost-effective strategies for improving the management of obesity. A full version of this review (including detailed descriptions of the included studies and their methodological quality, and results and excluded studies tables) is available in the Cochrane Library. The Cochrane Library is a database of systematic review and other evidence on the effects of health care, continuously updated as new information emerges. It is available on CD ROM from Update Software. For further information see: http://www.update-software.com/cochrane. PMID- 12119660 TI - Weight control and physical activity in cancer prevention. AB - Overweight and obesity have reached epidemic dimensions worldwide, mainly due to consumption of high energy diets and increased sedentary behaviour. Overweight and insufficient physical activity are clearly associated with cardiovascular diseases and type 2 diabetes. Evidence is also accumulating that they may also increase cancer risk, particularly in the colon, breast and endometrium. This effect seems to be mediated by alterations in the metabolism of endogenous hormones, including sex steroids and insulin, and levels of insulin-like growth factor(IGF)-I and IGF-binding proteins. In light of the beneficial effects of weight control and physical activity for cancer prevention, a healthy lifestyle, keeping a low body weight and exercising most days of the week, is recommended. PMID- 12119661 TI - Gastro-oesophageal reflux disease in obesity: pathophysiological and therapeutic considerations. AB - Gastro-oesophageal reflux disease (GERD) is common in obese patients. Apart from the physical discomfort and the economic burden, GERD may increase morbidity and mortality through its association with oesophageal carcinoma. The pathophysiology of GERD differs between obese and lean subjects. First, obese subjects are more sensitive to the presence of acid in the oesophagus. Second, hiatal hernia, capable of promoting GERD by several mechanisms, is more prevalent among the obese. Third, obese subjects have increased intra-abdominal pressure that displaces the lower oesophageal sphincter and increases the gastro-oesophageal gradient. Finally, vagal abnormalities associated with obesity may cause a higher output of bile and pancreatic enzymes, which makes the refluxate more toxic to the oesophageal mucosa. The altered body composition associated with obesity affects the pharmacokinetics of drugs. There are no data regarding the efficacy of any of the drugs used for GERD treatment. The dosages of cimetidine and ranitidine should be calculated according to the patient's ideal body weight, not their actual weight. Of the operative procedures used for weight loss, Roux-en-Y gastric bypass was found to be most effective for GERD, while gastric banding was associated with a high prevalence of reflux. This review outlines the pathophysiology and the treatment of GERD in obesity with emphasis on the therapeutic considerations in this population of patients. PMID- 12119662 TI - The genetic epidemiology of thinness. AB - Most genetic research in the area of human obesity asks the question 'Why are certain people obese?' Considerably less attention has been paid to the question of why certain people are not obese, particularly given the obesogenic environment that permeates the western culture. We present data from human and animal studies and evolutionary arguments supporting the notion that genetic studies of thinness or obesity resistance may yield important and complementary findings to genetic studies of obesity. We offer strategies for further refining the definition of thinness, weigh the advantages and disadvantages of potential sampling strategies and suggest candidate genes for thinness or obesity resistance. PMID- 12119663 TI - The importance of physical activity in the prevention of overweight and obesity in childhood: a review and an opinion. AB - The prevalence of childhood obesity is increasing and there are a number of theoretical reasons as to why intervention may be more effective in childhood. There are certain risk times for the development of obesity in childhood, which provide a basis for targeted intervention. In addition, tracking data supports the persistence of obesity, at least in later childhood, as well as cardiovascular risk factors. Physical activity is the discretionary component of energy expenditure and there is evidence that falling levels of physical activity are contributing to the obesity epidemic. Physical activity in children is related to developmental stage, is reduced with increasing age and is influenced by parental physical activity. While there is debate about the immediate health benefits of physical activity to children, there are data to support that lower physical activity levels and sedentary behaviours are associated with a higher prevalence of obesity in children. Physical activity is an accepted strategy in the treatment of established obesity (tertiary prevention). The role of physical activity in the prevention of obesity (primary and secondary prevention) is less clear. However a number of recent school-based interventions directed at either increasing physical activity and/or decreasing sedentary behaviours, have shown encouraging results. On balance, increasing physical activity in children is an attractive and non-restrictive approach to obesity prevention. To adopt this approach requires the support and involvement of many community sectors other than health. PMID- 12119664 TI - Obesity and immune function relationships. AB - The immunological processes involved in the collaborative defence of organisms are affected by nutritional status. Thus, a positive chronic imbalance between energy intake and expenditure leads to situations of obesity, which may influence unspecific and specific immune responses mediated by humoral and cell mediated mechanisms. Furthermore, several lines of evidence have supported a link between adipose tissue and immunocompetent cells. This interaction is illustrated in obesity, where excess adiposity and impaired immune function have been described in both humans and genetically obese rodents. However, limited and often controversial information exist comparing immunity in obese and non-obese subjects as well as about the cellular and molecular mechanisms implicated. In general terms, clinical and epidemiological data support the evidence that the incidence and severity of specific types of infectious illnesses are higher in obese persons as compared to lean individuals together with the occurrence of poor antibody responses to antigens in overweight subjects. Leptin might play a key role in linking nutritional status with T-cell function. The complexities and heterogeneity of the host defences concerning the immune response in different nutritional circumstances affecting the energy balance require an integral study of the immunocompetent cells, their subsets and products as well as specific and unspecific inducer/regulator systems. In this context, more research is needed to clarify the clinical implications of the alterations induced by obesity on the immune function. PMID- 12119665 TI - Do stress reactions cause abdominal obesity and comorbidities? AB - 'Stress' embraces the reaction to a multitude of poorly defined factors that disturb homeostasis or allostasis. In this overview, the activation of the hypothalamic-pituitary-adrenal (HPA) axis and the sympathetic nervous system have been utilized as objective measurements of stress reactions. Although long-term activation of the sympathetic nervous system is followed by primary hypertension, consequences of similar activation of the HPA axis have not been clearly defined. The focus of this overview is to examine whether or not repeated activation of these two stress centres may be involved in the pathogenesis of abdominal obesity and its comorbidities. In population studies adrenal hormones show strong statistical associations to centralization of body fat as well as to obesity. There is considerable evidence from clinical to cellular and molecular studies that elevated cortisol, particularly when combined with secondary inhibition of sex steroids and growth hormone secretions, is causing accumulation of fat in visceral adipose tissues as well as metabolic abnormalities (The Metabolic Syndrome). Hypertension is probably due to a parallel activation of the central sympathetic nervous system. Depression and 'the small baby syndrome' as well as stress exposure in men and non-human primates are followed with time by similar central and peripheral abnormalities. Glucocorticoid exposure is also followed by increased food intake and 'leptin resistant' obesity, perhaps disrupting the balance between leptin and neuropeptide Y to the advantage of the latter. The consequence might be 'stress-eating', which, however, is a poorly defined entity. Factors activating the stress centres in humans include psychosocial and socioeconomic handicaps, depressive and anxiety traits, alcohol and smoking, with some differences in profile between personalities and genders. Polymorphisms have been defined in several genes associated with the cascade of events along the stress axes. Based on this evidence it is suggested that environmental, perinatal and genetic factors induce neuroendocrine perturbations followed by abdominal obesity with its associated comorbidities. PMID- 12119666 TI - Self-help in the long-term treatment of obesity. AB - Despite the short-term success of professional behavioural techniques for obesity, weight losses are typically regained following treatment. The long-term maintenance of treatment effects will probably require ongoing, continuing care. Continuing care may be economically feasible when administered through self-help treatment modalities. Self-help confers a number of psychological benefits, such as self-reliance and an increased sense of empowerment. The effectiveness and cost-effectiveness of various modalities of self-help are reviewed, including purely self-prompted help, self-administered manuals, computer-assisted therapy, professionally assisted correspondence courses, and non-profit and commercial self-help groups. Stepped-care models suggest using a combination of these approaches when appropriate. However, logistical difficulties present themselves in stepped-care approaches with obesity, such as the complicating effects of severity and comorbidity on stepped-care status. Self-help groups are a promising venue for the provision of continuing care and as an adjunct to more intensive, specialty therapies. PMID- 12119667 TI - Obesity prevalence and trends in Latin-American countries. AB - The prevalence of obesity in some lower-income and transitional countries is as high as, or even higher than, the prevalence reported in developed nations, and it seems to be increasing rapidly. In most countries, the prevalence of obesity is higher in women than in men, and higher in urban than in rural areas. Preobesity prevalence is very high in most Latin-American countries. Sixty per cent of the population in Venado Tuerto (Argentina) has a body mass index (BMI) of > or = 25 kg m-2, as do 35% of the population in Brazil, 60% in Mexico, 68% in Paraguay and 53% in Peru. Trends are available from Brazil, where marked increases in the prevalence of obesity have occurred, except in women from higher income groups. Women from the higher-income quartiles in urban regions experienced a marked reduction in obesity prevalence from 1989 to 1997 (12.8 to 9.2%). Although data in children is scant, the prevalence of undernutrition is decreasing and the prevalence of obesity is high also in Latin-American children. The prevalence of obesity is high even in minority Indian groups. Rapid changes in dietary structure (in particular associated with urbanization) and major changes in the levels of physical activity, both occupationally and during leisure time, may explain these changes. PMID- 12119668 TI - Recent developments of charge injection and charge transfer in DNA. AB - The question of whether and how electrons migrate through DNA was a matter of controversial discussion over the last ten years. Today, there is no doubt that long distance charge migration through DNA exists and most scientists explain this process by a multistep hopping mechanism. This feature article presents recent developments of our group on the injection of a positive charge into DNA bases and the transfer of the charge between the DNA bases. The influence of the donor, the nature of the bridge, and the distance between the donor and the acceptor are discussed. PMID- 12119669 TI - Combinatorial development of chiral phosphoramidite-ligands for enantioselective conjugate addition reactions. AB - Chiral phosphoramidite ligands embodying bispidine frame work and a binaphthyl phosphoramidite for Cu-catalysed enantioselective conjugate addition reactions were developed employing principles of combinatorial and solid phase chemistry. PMID- 12119670 TI - Electrochemical polymerization of aniline inside ordered macroporous carbon. AB - 3D ordered macroporous multicomponent composite materials have been fabricated by electrochemical deposition of aniline on the inner surface of macroporous carbon; the maximum thickness of polyaniline (PANI) deposited is dependent on the concentration of the aniline as well as the dimension of the windows in the macroporous carbon. PMID- 12119671 TI - Reaction chemistry of metallotexaphyrins: the synthesis and characterization of the first meso-oxotexaphlorin. AB - Ring-oxygenation of metallotexaphyrins, promoted by strong bases, produces oxotexaphlorin, the first example of a meso-oxo functionalized texaphyrin derivative. PMID- 12119672 TI - Conductimetric immunosensor based on poly(3,4-ethylenedioxythiophene). AB - A conductimetric reagentless immunosensor using the biospecific binding pair of goat antirabbit IgG and rabbit IgG has been designed and fabricated using poly (3,4-ethylenedioxythiophene) as the immobilization matrix-cumtransducer. PMID- 12119673 TI - Towards stable analogues of inositol phosphates: stereoselective syntheses of (alpha,alpha-difluoromethyl)phosphonic acid (DFMPA)-containing cyclohexanes. AB - Diels-Alder and conjugate addition reactions were used to prepare precursors to a range of fully functionalized and deoxy inositol phosphate analogues. PMID- 12119674 TI - A unique rhodium-catalyzed rearrangement process: isomerization of an alkyne to a 1,3-diene with concomitant migration of a formyl group. An expeditious route to dienals from readily available 4-alkynals. AB - In CH2Cl2, [Rh(BINAP)]BF4 catalyzes the isomerization of 4-alkynals to dienals with excellent regio- and stereoselectivity; this new process compares favorably with previously reported methods for the synthesis of this class of compounds; a possible pathway for this unusual rearrangement is provided. PMID- 12119675 TI - Synthesis of (+/-)-aculeatins A and B. AB - The first synthesis of two new antiprotozoal and natural products was performed using concomitant deprotecting dithiane-phenolic oxidative reactions to form in one-step the 1,7-dioxadispiro[5.1.5.2]pentadecane core. PMID- 12119676 TI - A new, highly selective synthesis of aromatic aldehydes by aerobic free-radical oxidation of benzylic alcohols, catalysed by n-hydroxyphthalimide under mild conditions. Polar and enthalpic effects. AB - A new selective synthesis of aromatic aldehydes is described, based on catalytic oxidation of benzyl alcohols with molecular oxygen at rt and atmospheric pressure. PMID- 12119677 TI - Creation of a chain-like cationic iron species in montmorillonite as a highly active heterogeneous catalyst for alkane oxygenations using hydrogen peroxide. AB - A chain-like Fe3+ species containing an Fe-O-Fe unit has been prepared within the interlayer space of a montmorillonite, which showed a very high catalytic activity (turnover frequency 386 h-1 and turnover number 23,200) for the oxidation of cyclohexane with H2O2, affording cyclohexyl hydroperoxide as a major product. PMID- 12119678 TI - Continuous green biocatalytic processes using ionic liquids and supercritical carbon dioxide. AB - Soluble Candida antarctica lipase B dissolved in ionic liquids showed good synthetic activity, enantioselectivity and operational stability in supercritical carbon dioxide for both butyl butyrate synthesis and the kinetic resolution of 1 phenylethanol processes by transesterification. PMID- 12119679 TI - A new supramolecular isomer of [Zn(nicotinate)2]n: a novel 4(2).8(4) network that is the result of self-assembly of 4-connected nodes. AB - Self-assembly of Zn(NO3)2 with m-pyridinecarboxylate (nicotinate) under mild conditions affords several products including a novel three-dimensional network [Zn(nicotinate)2]n that has connectivity defined by the circuit symbol 4(2).8(4). PMID- 12119680 TI - Selective oxidation of glycerol to glyceric acid using a gold catalyst in aqueous sodium hydroxide. AB - Glycerol is oxidised to glyceric acid with 100% selectivity using either 1% Au/charcoal or 1% Au/graphite catalyst under mild reaction conditions (60 degrees C, 3 h, water as solvent). PMID- 12119681 TI - Kinetic isotope effects on the dissolution kinetics of solid salicylic acid in aqueous solution: evidence for solubilisation via a proton dissociation recombination mechanism. AB - Quantitative Atomic Force Microscopy measurements made on the dissolving surface of solid salicylic acid in H2O and D2O reveal a kinetic isotope effect (kH/kD = 2.3 +/- 0.6) on the dissolution rate consistent with a transition state in which the proton is dissociated from the dissolving molecule. PMID- 12119682 TI - Second sphere supramolecular chirality: racemic hybrid H-bonded 2-D molecular networks. AB - Neutral hybrid 2-D networks have been generated using a bis-amidinium capable of chelating M(CN)6(3-) anions via hydrogen bonds: the packing of the achiral 2-D networks leads to channels which are occupied by water molecules forming polymeric H-bonded chains; furthermore, owing to the dihapto mode of H-bonding, the presence of supramolecular chirality of the delta' and lambda' types taking place within the second coordination sphere of the metallic centre has been demonstrated. PMID- 12119683 TI - Exfoliation of layered rutile and perovskite tungstates. AB - The layered trirutile phases HMWO6 (M = Nb, Ta) and the layered perovskite H2W2O7 (synthesized by acid leaching of Bi2W2O9) were exfoliated into nanoscale colloids by reaction with quaternary ammonium hydroxides. PMID- 12119684 TI - Monoalkyl, chiral-at-metal 'constrained geometry' complexes as efficient alpha olefin and methyl methacrylate polymerisation catalysts. AB - A new class of monoalkyl or monochloro constrained geometry group 4 complexes has been synthesized; upon activation with aluminium activators they serve as efficient catalysts for olefin polymerisation and for polymerisation of methyl methacrylate. PMID- 12119685 TI - The first fully characterized neutral and cationic rhodium(I)-complexes containing DMSO as the only dative ligand; S-, O- and bridging S,O-bidentate binding modes. AB - Isolation and characterisation of novel neutral and cationic Rh(I) complexes having only DMSO molecules as dative ligands give complexes showing S-, O- and bridging S,O-bidentate binding modes of DMSO. PMID- 12119686 TI - An unexpected Diels-Alder reaction on the fullerene core rather than an expected 1,3-dipolar cycloaddition. AB - Fullerene derivatives resulting from an unexpected Diels-Alder cycloaddition have been obtained by reaction of trans-2-stilbenecarboxaldehyde derivatives with N methylglycine and C60. PMID- 12119687 TI - Rapid, clean, and mild O-acetylation of alcohols and carbohydrates in an ionic liquid. AB - Archetypal O-acetylation reactions of alcohols and carbohydrates proceed rapidly in high yield under mild conditions in a dicyanamide based ionic liquid, that is not only an effective solvent but also an active base catalyst. PMID- 12119689 TI - Synthesis of a new boron carbonitride with a B4C-like structure from the thermolysis of N-alkylated borazines. AB - Thermolysis of N-trialkyl borazines at 500 degrees C produces homogeneous, amorphous boron carbonitride phases, whose compositions are dependent upon the borazine substituent, and whose structures are similar to that of icosahedral boron carbide, B4C. PMID- 12119688 TI - Heterogeneous enantioselective epoxidation of olefins catalysed by unsymmetrical (salen)Mn(III) complexes supported on amorphous or MCM-41 silica through a new triazine-based linker. AB - High enantioselectivity (ee up to 84%) has been achieved in heterogeneous asymmetric epoxidation using a silica-bound unsymmetrical (salen)-manganese(III) complex; amorphous silica can be used in the same way as MCM-41, showing a positive effect in the catalyst recycling. PMID- 12119690 TI - A new catalytic hetero-Heck type reaction. AB - Unsaturated N-chloroamines have been found to cyclise under palladium-catalysis in good yield, the proposed mechanism includes an oxidative addition of the chloroamine to Pd(0), thus opening a new entry to the amides of the late transition metals. PMID- 12119691 TI - Hydroformylation in fluorous solvents. AB - Triaryl-phosphines and -phosphites bearing fluorous ponytails give high rates, good linear selectivity and good retention of catalyst in the fluorous phase during hydroformylation of alkenes in fluorous solvents. PMID- 12119692 TI - Solvent-free mechanochemical synthesis of phosphonium salts. AB - Phosphonium salts have been prepared during high-energy ball-milling of triphenylphosphine with solid organic bromides; the reactions occur at ambient conditions without a solvent; in the case of 2-bromo-2-phenylacetophenone the reaction in a solution usually produces a mixture containing both the C phosphorylated and O-phosphorylated compounds, while the solvent-free mechanically induced transformation results in the thermodynamically favorable C phosphorylated product; the occurrence of the observed transformations during mechanical processing of solid reactants is confirmed by the solid-state 31P NMR spectroscopy and X-ray powder diffraction. PMID- 12119693 TI - Self-assembled calix[6]pyrrole capsules: solid-state encapsulation of different guests in preorganized calix[6]pyrrole capsules. AB - meso-hexamethyl-meso-hexaphenyl calix[6]pyrrole assembles into well-defined dimeric capsules in the crystalline state; the preorganized capsule serves as an efficient host for different organic guests as well as for solvent molecules. PMID- 12119694 TI - The first structurally characterised homoleptic sigma-organotitanium(III) compound. AB - The paramagnetic [EPR, S = 1/2, d1; gav = 1.959(2)] organotitanate(III) anion [TiIII(C6Cl5)4] has been found to be a mononuclear species with nearly tetrahedral geometry (X-ray). PMID- 12119695 TI - The first successful investigation into a cyclodextrin-based enzyme model as an efficient catalyst for luminol chemiluminescent reaction. AB - The chemiluminescence of the luminol-H2O2 system is found for the first time to be remarkably enhanced by the Ce(IV) complexes of cyclodextrin dimers. PMID- 12119696 TI - Computational study of electrophilic addition to acrylate anion:cyclic halonium is the transition structure for degenerate rearrangement of alpha-lactones. AB - The cyclic chloronium or bromonium carboxylate obtained by addition of Cl+ or Br+ to acrylate anion is shown by PCM/B3LYP/6-31+G* calculations to be not an intermediate but a transition structure for interconversion of equivalent halomethyl oxiranones. PMID- 12119697 TI - Electronic state and three-dimensional structure of Mn(III) active sites in manganese-containing aluminophosphate molecular sieve catalysts for the oxyfunctionalisation of alkanes. AB - Combined in situ X-ray absorption spectroscopy measurements and quantum mechanical calculations yield a quantitative three-dimensional structure of the tetrahedrally coordinated Mn(III) active sites in MnAlPO catalysts. PMID- 12119698 TI - Highly facile and stereoselective intramolecular [2 +2]photocycloadditions of bis(alkenoyl)ketenedithioacetals. AB - The conformational change induced by the introduction of a ketenedithioacetal moiety at C-4 of 1,7-substituted-1,6-heptadiene-3,5-diones results in favorable spatial relationships between the alkenoyl groups to effect efficient intramolecular cycloadditions: irradiation of bis(alkenoyl)ketenedithioacetals in solution leads to facile and stereospecific intramolecular [2 + 2] photocycloadditions resulting in the formation of substituted bicyclo[3.2.0]heptane-2,4-diones, the observed conformational rigidity of which is attributed to the push-pull character of the ketenedithioacetal group. PMID- 12119702 TI - Palladium catalysed copolymerisation of ethene with alkylacrylates: polar comonomer built into the linear polymer chain. AB - Copolymerisation of ethene and alkylacrylates is catalysed by palladium modified with di(2-methoxyphenyl)phosphinobenzene-2-sulfonic acid (DOPPBS); a linear polymer is produced in which acrylate units are incorporated into the polyethylene backbone. PMID- 12119700 TI - Multidentate Lewis acids: synthesis, structure and mode of action of a redox based fluoride ion sensor. AB - The mode of action of the bidentate bis(boronate) Lewis acid 2 as a fluoride ion sensor is shown to involve selective anion binding together with an electrochemical response. PMID- 12119701 TI - Total synthesis of fostriecin (CI-920) via a convergent route. AB - Fostriecin, a potent and promising antitumor antibiotic, was stereoselectively synthesized via a convergent route involving a three-segement coupling procedure. PMID- 12119699 TI - Minimalist peptidomimetic cyclophanes as strong organogelators. AB - L-Valine containing cyclophanes have been shown to gelate organic solvents leading to soft materials with a clear expression of their chirality at the supramolecular level. PMID- 12119703 TI - Studies in 3-oxy-assisted 3-aza cope rearrangements. AB - On thermolysis appropriately substituted N-silyloxy-N-allyl enamines undergo smooth 3,3-sigmatropic rearrangments to the corresponding N-silyloxy imino ethers. PMID- 12119704 TI - Phase transformation during cubic mesostructured silica film formation. AB - The structural evolution taking place during CTAB/TEOS based solvent evaporation induced thin film formation has been followed by in-situ time-resolved SAXS; this shows that the final Pm3n cubic structure is formed via the formation of lamellar and hexagonal intermediate structures within the water rich evaporation regime. PMID- 12119705 TI - Syntheses and X-ray crystal structures of dumbbell-shaped bis-fullerene tungsten and molybdenum complexes. AB - In the first neutral bis-fullerene complexes with tungsten(0) and molybdenum(0) [M(eta 2-C60)2(CO)2(dbcbipy)] [M = W and Mo, dbcbipy = 4,4'-di(butyl carboxyl) 2,2'-bipyridine] synthesized in solution, the metal atom coordination is distorted octahedral with two CO groups and the bipy group of dbcbipy in the equatorial plane and the metal atom binding in an eta 2-fashion to C-C bonds of two C60 at axial orientation to give a dumbbell shaped molecule. PMID- 12119706 TI - Biomolecule separation using large pore mesoporous SBA-15 as a substrate in high performance liquid chromatography. AB - Functionalized large-pore mesoporous SBA-15 is utilized for the first time as a good substrate in high performance liquid chromatography (HPLC) to separate biomolecules including peptides and proteins. PMID- 12119707 TI - Remarkable homology in the electronic spectra of the mixed-valence cation and anion radicals of a conjugated bis(porphyrinyl)butadiyne. AB - The pi-radical cation and anion of the dizinc complex of a bis(triarylporphyrinyl)butadiyne, 1+ and 1-, respectively, display remarkably similar near-IR signatures, with intense bands near 1000 and 2500 nm, as predicted by the appropriate frontier-orbital model for inter-porphyrin coupling across the conjugated bridge. PMID- 12119708 TI - Polynitrile-bridged two-dimensional crystal: Eu(III) complex with strong fluorescence emission and NLO property. AB - The first polynitrile coordinated lanthanide complex ([Eu(cda)3(H2O)3].H2O) infinity (cda = carbamyldicyanomethanide anion) has been synthesized and the two dimensional structure coordination polymer exhibits strong fluorescence emission and strong powder second harmonic generation efficiency (16.8 times that of urea). PMID- 12119709 TI - Solution and solid-state studies of 3,4-dichloro-2,5-diamidopyrroles: formation of an unusual anionic narcissistic dimer. AB - 3,4-Dichoro-1H-pyrrole-2,5-dicarboxylic acid bis-phenylamide 3 and 3,4-dichoro-1H pyrrole-2,5-dicarboxylic acid bis-butylamide 4 have been prepared and shown to deprotonate in the presence of basic anions: the X-ray crystal structure of the tetrabutylammonium salt of 3-H+ reveals the formation of a dimer in the solid state. PMID- 12119710 TI - Biphasic dimerisation of methylacrylate--immobilisation and stabilisation of cationic Pd-catalysts in ionic liquids by an ammoniumphosphine ligand. AB - The first continuous, biphasic, Pd-catalysed dimerisation of methylacrylate is described. The biphasic system has been realised by using a tetrafluoroborate ionic liquid as catalyst medium and an ammoniumphosphine ligand to immobilise and stabilise the Pd-catalyst. PMID- 12119711 TI - Synthesis, structure and magnetism of a new manganese carboxylate cluster: [Mn16O16(OMe)6(OAc)16(MeOH)3(H2O)3].6H2O. AB - A new manganese(III/IV) carboxylate cluster [Mn16O16(OMe)6(OAc)16(MeOH)3(H2O)3].6H2O has been structurally characterised and shown to display net antiferromagnetic coupling with preliminary evidence for single-molecule magnetic behaviour. PMID- 12119712 TI - Rational synthesis of alpha-MnO2 single-crystal nanorods. AB - alpha-MnO2 single-crystal nanorods with diameters 20-80 nm and lengths up to 6 microns have been prepared through a low-temperature liquid-phase comproportionation method, which involves no catalysts or templates and may be adjusted to prepare alpha-MnO2 single-crystal nanorods in large scale. PMID- 12119713 TI - Synthesis of a highly enantiomerically enriched silyllithium compound. AB - The highly enantiomerically enriched silyllithium compound lithiomethylphenyl(1 piperidinylmethyl)silane (4) reacts stereospecifically with chlorosilanes, but over a period of several hours slow racemization in solution at room temperature occurs, which can be suppressed by a transmetalation reaction with MgBr2(thf)4. PMID- 12119714 TI - Diastereoselective cascade synthesis of azabicyclo[3.1.0]hexanes from acyclic precursors. AB - The diastereoselective synthesis of azabicyclo[3.1.0]hexanes bearing different substituents on all positions of the cyclopropane ring has been achieved in moderate to good yields. PMID- 12119715 TI - A new fluorescent PET chemosensor for fluoride ions. AB - A new anthracene derivative bearing two phenylurea group at the 1,8-position of anthracene shows a selective fluorescence quenching effect with fluoride ion via a PET mechanism. PMID- 12119716 TI - Hydrolysis of a sulfonamide by a novel elimination mechanism generated by carbanion formation in the leaving group. AB - The alkaline hydrolysis of N-alpha-methoxycarbonyl benzyl-beta-sultam occurs 10(3) times faster than the corresponding carboxylate and with rapid D-exchange at the alpha-carbon: the pH rate profile indicates pre-equilibirum CH ionisation and together with formation of benzoyl formate as a product this suggests a novel mechanism for hydrolysis. PMID- 12119717 TI - Effect of surfactant phase transition on the inclusion behaviour of an amphiphilised porphyrin derivative. AB - The macroscopic morphology of a micelle biomembrane model strongly affects the aggregation state of an included porphyrin derivative. PMID- 12119718 TI - Strong fluorescence enhancement of 2-bromo-3-(1H-indol-3-yl)maleimide upon coordination to a Lewis-acidic metal complex. AB - The emission intensity of an indolyl maleimide derivative increases approximately 80-fold by reversible coordination to (1,4,7,11-tetraazacyclododecane)zinc(II), which makes the system a promising new signalling motif for molecular sensors. PMID- 12119720 TI - Synthesis of a layered zinc phosphate, [NH3(CH2)2NH2(CH2)3NH3][Zn2(PO4)(HPO4)2].H2O, and its transformation to a extra large pore three-dimensional zinc phosphate, [NH3(CH2)2NH2(CH2)3NH3][Zn3(PO4)(HPO4)3]. AB - An unusual two-dimensional zinc phosphate with pendant phosphate groups, projecting into the inter-lamellar space between the layers, has been synthesized and is shown to transform into a three-dimensional structure with 16-membered bifurcated channels, giving evidence for the building up process in the formation of open-framework structures. PMID- 12119719 TI - Base promoted isomerization of aziridinyl ethers: a new access to alpha- and beta amino acids. AB - Aziridinyl ethers are selectively and easily converted to either amino vinyl ethers or alkoxy allylamines by treatment with mixed metal bases (superbases). PMID- 12119722 TI - Amorphous iridium complexes for electrophosphorescent light emitting devices. AB - Iridium complexes with fluorene-modified phenylpyridine ligands are resistant to crystallization and can be used in the fabrication of single layer light emitting diodes. PMID- 12119721 TI - Energies and selectivities for anion binding as a function of host conformational preorganisation. AB - The anion binding of tripodal hosts 2-4 has been studied. Increasing levels of conformational preorganisation of the side arms of the hosts led to increased (Cl ) unaltered (Br-) or decreased (NO3-) binding; it was thus possible to change guest selectivities by about an order of magnitude through conformational preorganisation of the flexible host. PMID- 12119724 TI - Fast and unprecedented chemoselective hydroformylation of acrylates with a fluoropolymer ligand in supercritical CO2. AB - A fluorous polymeric phosphine, when combined with supercritical CO2 (scCO2) and rhodium, effects fast and highly chemoselective hydroformylation of acrylates, one of the least reactive olefins in hydroformylation reactions. PMID- 12119725 TI - Mechanism of the oscillatory decomposition of the dithionite ion in a flow reactor. AB - A simple mechanism consisting of three protonation equilibria and seven redox reactions between sulfur species can describe the large amplitude-sustained temporal pH-oscillations observed during acid-induced decomposition of the dithionite ion in a continuous-flow stirred tank reactor (CSTR) in the temperature range 25-60 degrees C. PMID- 12119723 TI - The fixation of linear versus loop-type peptidic structures by metal coordination: the coordination chemistry of Val-Val- and Val-Val-Val-bridged dicatechol ligands. AB - The Val-Val-bridged dicatechol ligand L1-H4 forms triplybridged dinuclear complexes with titanium(IV) ions, while the more flexible Val-Val-Val derivative L2-H4 leads to mixtures of complexes containing species with a cyclic arrangement of the ligand; with [cis-MoO2]2+ on the other hand, a well-defined macrocycle [(L2)MoO2]2- is formed which possesses a loop-type structure in the peptidic part of the ligand. PMID- 12119726 TI - Preparation of silver nanoparticles in solution from a silver salt by laser irradiation. AB - A new method is proposed for the fabrication of a well-defined size and shape distribution of silver nanoparticles in solution; the method employs direct laser irradiation of an aqueous solution containing a silver salt and a surfactant in the absence of reducing agents. PMID- 12119727 TI - At last. Steamy! PMID- 12119728 TI - Something's burning. PMID- 12119729 TI - Will R&D make Merck hot again? PMID- 12119730 TI - Spared by their low IQ. PMID- 12119731 TI - Eggs on ice. PMID- 12119732 TI - What's in your pipes? PMID- 12119733 TI - Monkeys galore. PMID- 12119734 TI - How to prevent a migraine. PMID- 12119736 TI - The year I chucked conventional medicine. PMID- 12119735 TI - This doctor's peer review suit cost him $240,000. PMID- 12119737 TI - Take-home advice from a stand-up consultant. PMID- 12119738 TI - The Feds ease antitrust rules--cautiously. PMID- 12119739 TI - White coats, drowning horses. PMID- 12119740 TI - Is your practice staffed correctly? PMID- 12119741 TI - Don't "polish" your records. PMID- 12119742 TI - From the analyst's couch. Drugs that target angiogenesis. PMID- 12119743 TI - Angiogenesis modulation in cancer research: novel clinical approaches. AB - Angiogenesis--the formation of new blood vessels--is essential for tumour progression and metastasis. Consequently, the modulation of tumour angiogenesis using novel agents has become a highly active area of investigation in cancer research, from the bench to the clinic. However, the great therapeutic potential of these agents has yet to be realized, which could, in part, be because the traditional strategies that are used in clinical trials for anticancer therapies are not appropriate for assessing the efficacy of agents that modulate angiogenesis. Here, we discuss methods for monitoring the biological activity of angiogenic modulators, and innovative approaches to trial design that might facilitate the integration of these agents into anticancer therapy. PMID- 12119744 TI - Novel animal-health drug targets from ligand-gated chloride channels. AB - The world's three best-selling veterinary antiparasitic drugs ('parasiticides') act on ligand-gated ion channels. The sequencing of the complete genomes of the invertebrate genetic model organisms Caenorhabditis elegans and Drosophila melanogaster has led to the recent cloning of new subunits of 5-hydroxytryptamine gated and histamine-gated chloride channels. Together with L-glutamate-gated chloride channels, which are important targets of known parasiticides, and acetylcholine-gated chloride channels, these new classes of ligand-gated chloride channels, which are known only from invertebrates, add to our understanding of inhibitory neural signalling. They could offer the prospect of being targets for a new generation of selective drugs to control nematode and insect parasites. PMID- 12119745 TI - New treatments for COPD. AB - COPD is one of the most common diseases in the world, and there is a global increase in prevalence, but there are no drugs available at present that halt the relentless progression of this disease. However, a better understanding of the cellular and molecular mechanisms that are involved in the underlying inflammatory and destructive processes has revealed several new targets for which drugs are now in development, and the prospects for finding new treatments are good. PMID- 12119746 TI - Encoding microcarriers: present and future technologies. AB - In answer to the ever-increasing need to carry out many assays simultaneously in drug screening and drug discovery, several microcarrier-based multiplex technologies have arisen in the past few years. The compounds to be screened are attached to the surface of microcarriers, which can be mixed together in a vessel that contains the target analyte. Each microcarrier has to be encoded to know which compound is attached to its surface. In this article, the methods that have been developed for the encoding of microcarriers are reviewed and discussed. PMID- 12119747 TI - Bioequivalence and the immunogenicity of biopharmaceuticals. AB - The expiry of the first patents for recombinant-DNA-derived biopharmaceuticals will open the possibility of marketing generics, if they can be shown to be essentially similar to the innovator product. However, as shown by the problem of immunogenicity, the properties of biopharmaceuticals are dependent on many factors, including downstream processing and formulation. Products from different sources cannot be assumed to be bioequivalent, even if identical genes are expressed in the same host cells and similar production methods are used. Some of the influencing factors are still unknown, which makes it impossible to completely predict biological behaviour, such as immunogenicity, which can sometimes lead to serious side effects. PMID- 12119748 TI - The impact of pharmacogenetics and pharmacogenomics on drug discovery. AB - It is widely perceived at present that pharmacogenetics and pharmacogenomics are about to revolutionize the face of medicine. In a more realistic assessment, the implementation of molecular genetics and biology will provide us with better ways to treat illnesses, and has already begun to do so in an incremental and evolutionary fashion. However, it is unlikely to change fundamentally the direction of medical progress. Advances are most likely to be made in the area of pharmacodynamics, as we learn to differentiate broader conventional clinical diagnoses into separate molecular subtypes. PMID- 12119749 TI - Preclinical safety evaluation of biotechnology-derived pharmaceuticals. PMID- 12119750 TI - Dealing with the data deluge. PMID- 12119751 TI - [A depression should be treated--but how?]. PMID- 12119752 TI - [Reference programs--a tool in every day work]. PMID- 12119753 TI - [The Danish translation of the diagnostic child and adolescent psychiatric interview "Schedule for Affective Disorders and Schizophrenia for School-aged Children, Present and Lifetime Version"]. PMID- 12119754 TI - [The effect of psychotherapy on depression]. AB - Over the last decades, numerous studies of the efficacy of psychotherapy on major depression have been carried out. They provide evidence of the effectiveness of cognitive therapy and interpersonal therapy in the treatment of mild to moderate depression. The effectiveness of psychotherapy in in-patients and patients with severe depression has not been documented. Maintenance interpersonal therapy may contribute to prevent new episodes in recurrently depressive patients, where promising studies also indicate that cognitive therapy has a preventive effect on future depressions. This calls for larger and better controlled studies. PMID- 12119755 TI - [Diagnosis and treatment of depression in general practice. A questionnaire study]. AB - INTRODUCTION: The purpose of this questionnaire study was to assess the knowledge and attitude of general practitioners (GPs) towards the use of diagnostic criteria in clinical practice, and the extent to which diagnostic tools are used as a prerequisite for the treatment of depressive patients in general practice. MATERIAL AND METHODS: A total of 758 out of 1,700 randomly selected GPs responded to the questionnaire. The GPs' demographic data (age, sex, number of years in practice, supplementary education), diagnostic practice (knowledge of depressive core symptoms and diagnostic tools and scales), and clinical practice (assessment of the effectiveness of treatment, duration of treatment, aim of treatment, and use of psychotherapy) were registered. RESULTS: The study showed that GPs, who had taken part in supplementary training in psychiatric issues within the preceding year, to a higher extent than the rest of the GPs used diagnostic tools for all or most patients. This group of GPs also had a significantly greater knowledge of the depressive core symptoms as described in the ICD-10. When compared with the other group of GPs, they also felt themselves more confident of the diagnosis of depression before prescribing antidepressive medication. The vast majority of both groups felt that many depressive patients remained undiagnosed, because they simply do not consult their GPs. They also found that it was often questionable whether the symptoms were caused by life crisis or depression--and whether or not the depressive symptoms of these patients required drug treatment. DISCUSSION: The questionnaire study shows that the GPs' level of knowledge of diagnostics of depression is insufficient. There is an obvious need for supplementary training in general practice, thereby increasing the knowledge and the use of diagnostic criteria in order to make diagnoses of depression more correct and to improve treatment. PMID- 12119756 TI - [A depression rating scale for the ward]. AB - BACKGROUND: The aim of the present study was to develop a quantitative rating scale for depression based on observations in the ward and one that can be administered by the nursing staff. MATERIALS AND METHODS: Based on a literature survey and existing depression rating scales, a new rating scale was developed especially suited for use in inpatient care. The patients were rated simultaneously with the Beck Depression Inventory (BDI) and Clinical Global Impression (CGI). Estimation of construct validity, criterion validity, and item bias was performed with Rasch analysis and analysis of correlation. RESULTS: Statistical analysis revealed that the scale consisted of two sub-scales for mood and behaviour. One item had to be omitted as it was not homogeneous with either of the two sub-scales. DISCUSSION: The present depression rating scales rely on ward observations of patients during admission. This might be the reason why the items concerning behaviour are so strongly represented in this scale. Good correlations were found between the different scales, except for the score on BDI and the sub-scale concerning behaviour. This may be explained by the fact that the BDI is the patient's own judgement of cognitive function and mood. The observations in the ward are to a greater extent built on observation of patient behaviour. It can be used routinely on the psychiatric ward. PMID- 12119757 TI - [Electroconvulsive therapy in Denmark 1999. A nation-wide questionnaire study]. AB - INTRODUCTION: The use of ECT in Denmark has not been systematically evaluated since 1979. Many changes have since taken place in the organisation of psychiatric care and the number of new drugs for the treatment of affective disorders has grown. We wanted to see whether the frequency of treatments and the technical standard of ECT had undergone changes in the same period. MATERIAL AND METHODS: A questionnaire was sent to all psychiatric departments in Denmark concerning the number of ECT sessions and ECT patients in 1999, and technical aspects and organisation of this treatment-modality. RESULTS: The percentage of replies was 100, all departments used ECT, and 86% of the departments adequately reported the number of ECT sessions and ECT-treated patients. The total number of sessions and patients treated was estimated from these answers. We found that the number of ECT sessions had fallen from 19,564 in 1979 to 16,306 in 1999 and the number of ECT patients from 2,332 in 1979 to 1,710 in 1999. ECT is thus given to an average of 5% of all hospitalised psychiatric patients. The number of sessions per patient had increased slightly. The technical quality of treatment with regard to the ECT devices used had improved considerably during the period. DISCUSSION: The use of ECT in Denmark shows a decline since the last evaluation in 1979. However, it still plays an important role in the treatment of hospitalised patients with depression in Denmark. PMID- 12119758 TI - [Cotard's syndrome in depression and maintenance electroconvulsive therapy]. AB - We describe the case of a 65-year-old woman with long-standing, atypical facial pain and fluctuating mental symptoms. A variety of psychiatric diagnoses were made over a period of nine years but no effective treatment was found. Ultimately, the patient was found to suffer from a psychotic depression indicating Cotard's syndrome. Two series of ECT were then given with temporary short remissions. Subsequently, continuation ECT was introduced. To our knowledge, continuation ECT in Cotard's syndrome is rarely, if ever, described in the literature. PMID- 12119759 TI - [Cognitive behavior therapy of a patient with chronic schizophrenia]. AB - A case of successful cognitive behavioural treatment of a 63-year-old woman with chronic schizophrenia is reported. She suffered from medication-resistant auditory hallucinations and was therefore referred for therapy. She continued throughout the therapy on olanzapine 10 mg, the maximum dose tolerated. The therapy went through the stages of engagement, psychological formulation, normalisation, challenging beliefs and providing alternative explanations to her experiences. Socratic questioning was used throughout the therapy as part of the ongoing engagement process. The therapy lasted 21 sessions and she achieved a significant reduction in the PSYRATS rating scale, which was consistent with the markedly improved clinical picture. PMID- 12119761 TI - [Papillary thyroid carcinoma with pulmonary metastases in a child]. AB - Thyroid carcinoma is an uncommon disease in children, thus it is not always suspected early in the evaluation of a child with neck mass. Thyroid carcinoma in children can be treated effectively and has relatively good prognosis if diagnosed early. The lungs are the most common site for distant metastases and in many cases partial remission can be achieved with good quality of life. In this report we describe a child with bilateral neck masses diagnosed to have papillary thyroid carcinoma with neck and pulmonary metastases. This report aims to raise the physicians awareness of the disease in children. We review the diagnostic modalities and treatment of the disease in children. PMID- 12119760 TI - [Fatal attack in patients with bronchial asthma]. AB - What is most notable about asthma deaths is that they continue to occur despite increased understanding of the pathophysiology of asthma, more effective treatment and improvements in asthma education. Studies of fatal and near fatal asthma revealed two distinct groups of patients. The first is a "slow onset-late arrival" group (type I) in whom the final common pathway is a period of poor asthma control, increased use of bronchodilators, late arrival for care of the final attack and underutilization of glucocorticosteroids. This group of patients is characterized by the presence of eosinophilia in the airways. The second group (type II), includes only the minority of death cases. These patients have a sudden, unexpected and rapidly progressive attack. This group of patients is characterized by the presence of neutrophilia in the airways. Two cases of death due to asthma attach, one presenting type I patients and the second type II patients, are described. PMID- 12119762 TI - [Horner's syndrome during epidural analgesia in labor: an alarming sign or a benign phenomenon?]. AB - Horner's syndrome has been reported as a complication of epidural analgesia for labor. In all such cases Bupivacaine 0.5%-0.25% was used. Recently, Bupivacaine of much lower concentration has become a routine anesthetic for labor. We present, for the first time, two parturients who developed Horner's syndrome with diluted Bupivacaine. We suggest that with the absence of other neurological signs, once subdural and subarachnoid injections are excluded, further investigations are not required during the first 24 hours since in most of the cases the Horner's syndrome is due to high rostral spread of the epidural block and disappears within 24 hours. Epidural analgesia can be carefully continued albeit appearance of the syndrome. PMID- 12119763 TI - [Use of health services during the first year after stroke in community dwelling patients]. AB - BACKGROUND: Stroke is the leading cause of disability in the elderly population, and inpatient rehabilitation is the most accepted method of providing rehabilitation to stroke survivors remaining with moderate to severe disability. It is also a major component of the large resources that this patient population demands. Since it is possible to develop alternatives to inpatient rehabilitation, it is important to make a long-term evaluation of its resource allocation, to enable comparison with the other alternatives. OBJECTIVES: To evaluate and classify the health services demanded during the first year after stroke by patients who had received inpatient rehabilitation. METHODS: A prospective follow-up of 75 consecutive stroke survivors who remained with moderate disability and were accepted for inpatient rehabilitation. Utilization of health services was recorded and evaluated considering the patients health status and success in returning to independent daily function. RESULTS: Inpatient rehabilitation lasted about fifty days, started relatively early and enabled participants to regain an acceptable level of basic functional independence. During the year of follow-up, about half of the patients were hospitalized, with stroke related causes contributing significantly. Not all patients were followed up regularly by community health services, and most suffered significant handicap and depression. CONCLUSIONS: This survey justifies the initial investment in inpatient rehabilitation for preparing stroke survivors to continue their lives at home. There is a need to improve continuity of services and to refine the methods of medical and psychosocial follow-up in the community. PMID- 12119764 TI - [The influence of gender on the perception of dyspnea in patients with mild moderate asthma]. AB - BACKGROUND: Men and women respond differently to asthma. Current data suggest that there are more hospital admissions, decreased quality of life (QOL) and increased use of MDIs, in asthmatic females compared to males. PATIENTS AND METHODS: Respiratory muscle strength, 2-agonists consumption, the QOL and the perception of dyspnea (POD) were measured in 20 asthmatic females and 20 males, with mild-persistent to moderate asthma. The results were compared to 40 normal subjects (20 females and 20 males), age matched, who served as a control group. RESULTS: Inspiratory and expiratory muscle strength were significantly lower (p < 0.01), while the POD and the mean daily 2-agonist consumption were significantly higher (p < 0.005 and p < 0.001, respectively), in the females. The females also had significantly lower QOL compared to their male counterparts. CONCLUSIONS: The POD and the mean daily 2-agonist consumption, in asthmatic females, are significantly higher and the QOL significantly lower than that of their male counterparts. The females also had weaker respiratory muscles. Hence, knowing the relation between the POD and the respiratory muscles it can be assumed that this deference in strength may play a role in the gender differences in the response to asthma. PMID- 12119766 TI - [Intercessory prayer for health: a matter of faith, science or both]. PMID- 12119765 TI - [Acute acquired non-traumatic torticollis in hospitalized children]. AB - BACKGROUND: The inflammatory process in torticollis involves the cervical muscles, nerves, and/or the vertebral synovia causing painful and abnormal head position. Most cases of acute acquired non-traumatic torticollis are of benign etiology, but some patients may suffer from a serious disease requiring thorough investigation and hospitalization. OBJECTIVES: To describe the epidemiology, clinical features and the etiology of acute acquired non-traumatic torticollis in our center. METHODS: A retrospective chart review of 45 hospitalized children in the Sapir Medical Center over a 10 year period. RESULTS: We studied 45 children; 23 girls and 22 boys. Their mean age was 6 years (range 6 months to 16 years). All patients reported marked neck pain as their main complaint. Local infection was the most frequent etiology, and 33 (73%) patients were hospitalized during the fall-winter season. Twenty six patients underwent x-ray imaging, and of these, ten (38%) had pathology. The main findings were some degree of atlanto axial subluxation, and widening of the prevertebral space. Treatment included analgesic and antibiotics for suspected bacterial infections. Two patients required surgical drainage. Cervical neck traction was performed on 15 patients. Mean admission time was 4.1 days and was shorter in the pediatric department compared to the ENT and orthopedic departments. CONCLUSIONS: Careful clinical and radiological evaluation for the wide spectrum of clinical entities should be done. Local infections of the respiratory tract are the most common etiology. Conservative treatment usually leads to complete resolution in a short time. PMID- 12119768 TI - [Medicine and intercessory prayer]. PMID- 12119767 TI - [Fatal asthma: who, when, why and how to prevent?]. AB - Despite our understanding of the inflammatory nature of asthma, mortality from the disease has been on the rise. Most deaths follow a protracted course, but some occur abruptly. Many of those who die or who have experienced a near fatal episode share common characteristics (giving hope that many deaths are preventable), such as insufficient steroid therapy, reliance on inhaled bronchodilators, marked peak flow fluctuations, psychiatric disorders and attenuated dyspnea in response to stimuli. No genetic marker has been identified for the at-risk group, but the fall in lung function in response to methacholine dose is high and the ventilatory response to hypoxia is blunted. The last 3 characteristics may be causative. The patient at risk needs to be identified and treated with the goal to normalize lung function (as is usually recommended). However, in this group it may be desirable (though not yet proven) to up-titrate therapy further, with the added goal to minimize peak flow fluctuations and bronchial reactivity. Patients (and families) need to be taught to monitor peak flow and as the disease worsens, by clinical and by written peak flow criteria, to increase inhaled steroids, to use oral steroids, to self inject subcutaneous epinephrine or to sick (urgent) medical attention. This mode of therapy depends on tight patient-physician relation, a pattern of behavior that is not easy to achieve with this group. PMID- 12119769 TI - [Fallopian tube prolapse after hysterectomy]. AB - Fallopian tube prolapse is a rare complication of hysterectomy, characterized by vaginal discharge, abdominal pain, pelvic inflammatory disease and vaginal bleeding. The diagnosis is often delayed, and is usually done after an histopathological examination identifies fallopian tube on biopsy. The advised treatment is surgical resection, which can be done through vaginal incision, abdominally or by laparoscopy. We report a case of fallopian tube prolapse after vaginal hysterectomy in 47-year-old patient in whom the prolapsed-tube was successfully resected vaginally, and review the presentation and surgical methods to correct this rare complication. PMID- 12119770 TI - [West Nile fever: the virus, the mosquito, the disease and our coping]. AB - West Nile fever is a viral disease, transmitted to humans by a mosquito bite. An outbreak of West Nile fever occurred last year in Israel, causing substantial morbidity and mortality. This article reviews the literature regarding this disease, focusing on several recent outbreaks around the world, and with special emphasis on the situation here, in Israel. Recommendations for better coping with future outbreaks to the general public, health authorities and state leaders, are presented at the end of the article. PMID- 12119771 TI - [Euthenasia: attitudes, wishes and behavior of the public, patients and physicians]. AB - The lack of compatibility in conventional medical treatment for patients suffering from terminal diseases or severe disabilities has become one of the more difficult problems of developed countries. Different types of passive and active euthanasia have been adopted as solutions to these new problems. These solutions, however, raise ethical and professional dilemmas, as well as controversies regarding economic and policy-making issues. This situation of social confusion and the publics anxiety regarding end-of-life care are reflected in the literature review, which shows significant discrepancies between physicians and the public in attitudes, preferences and behavior regarding the preferred and existent medical treatment at the end of life. The publics preferences regarding end-of-life care are relatively stable over time. However, the readiness of the public and physicians to accept these solutions increases over time. Considering these controversial solutions and the publics wishes to participate in the decision making process regarding end-of-life care, an increase in open communication and the degree of cooperation among physicians and their patients and family members is suggested. Advancing the treatment methods for severely ill patients, and improving the education of professional caregivers is another essential need. The development of satisfactory treatment alternatives for patients at the end of life may prevent the Israeli society from debating active euthanasia and lead to clear social agreements on passive euthanasia, including the developing of legal options which will enable people, who are interested, to gain control over their end-of-life care. PMID- 12119772 TI - [Aldosterone--classic and non classic effects]. AB - Aldosterone, the major mineralocorticoid and most potent sodium retaining hormone in mammals was discovered in 1952 by Simpson and Tait. Since then numerous studies were carried out to explore the mechanisms underlying the biological actions of this steroid hormone, and the physiological effects of aldosterone remained an attractive area of research. The utilization of sophisticated molecular biology techniques led to the identification of genes encoding major components of the aldosterone-regulated sodium reabsorptive pathway, such as mineralocorticoid receptor. 11b-hydroxysteroid dehydrogenase type 2, epithelia Na+ channel, and sgk and Kras genes. It is clear that aberrations in these genes lead to genetic diseases affecting blood pressure and fluid homeostasis. However, in the last decade several clinical and experimental studies have implicated aldosterone in the pathogenesis of cardiovascular dysfunction, especially heart failure. In this context, aldosterone increases the accumulation of collagen in the fibroblast and myocytes of the cardiac tissue leading to fibrosis and cardiac hypertrophy. Blockade of aldosterone receptors by spironolactone substantially reduced the risk of morbidity and death among patients with severe heart failure. The present review summarizes the recent developments in the molecular mechanisms of aldosterone actions and its involvement in the pathogenesis of heart failure. PMID- 12119773 TI - [Sleeping and eating disorders]. AB - Over the last three decades there has been a dramatic increase in the prevalence of eating disorders (ED) in western society. The main syndromes are anorexia nervosa (AN), bulimia nervosa (BN) and non-specified eating disorders (ED-NOS). These disorders are with high morbidity and life threatening complications. Sleep disturbances are predominant symptom in these disorders. Researches have examined sleep disorders among people suffering from eating disorders, using different methods: sleep polysomnography, actigraph and self report questionnaires. The article presents the diagnostic criteria of eating disorders, sleep structure and review of researches, which examined sleep patterns among people with AN, BN and binge eating disorder (BED). In addition the article reviews the night eating syndrome. This syndrome is considered to be a combination of eating disorder and sleeping disorder. The article describes the characteristic sleep-wake patterns of each syndrome and in comparison to the control group. The discussion suggests some possible explanations for the discrepancies between subjective and objective experience of sleep. PMID- 12119774 TI - [Our experience with two stage ear reconstruction of typical microtia]. AB - One of the most difficult reconstructive procedures is the reconstruction of the auricle, since it has a very difficult form to imitate. During the last four decades there have been numerous reconstructive attempts throughout the world especially for cases of microtia. The principle is to build a framework from cartilage that will be used for the reconstruction. A prominent personality in this field is Dr. Burt Brent, who is identified with this challenge. Dr. Brent has established the method of reconstruction in four stages. In order to limit the operative stages Dr. S. Nagata suggested doing the reconstruction in two stages, using a different cartilage framework and elevating the auricle using temporo-parietal fascial flap and skin graft. His method was improved further by Dr. Francoise Firmin. The first stage is conducted at the age of 9-10, during which the ipsilateral rib cartilage is harvested, carved and put together into a cartilage framework and than inserted into a skin pocket in the area awaiting reconstruction. Half a year later, the auricle is elevated by inserting a wedge cartilage behind it, covering with temporo-parietal fascial flap and skin graft. Our experience using this method in ten microtia cases proves that it is a good choice in total auricle reconstruction for microtia patients. PMID- 12119775 TI - [The effects of Cytomegalic virus (CMV) infection during pregnancy on the developing human fetus]. AB - In most instances Cytomegalic virus causes a sub-clinical infection, rendering the pregnant mother unaware of being infected. However, primary CMV is the most common infection during pregnancy that may have long-term neuro-developmental sequelae in 10%-20% of children born to these mothers. The clinical findings in these children include intrauterine growth restriction (IUGR), microcephaly, hearing and visual impairment, developmental delay, mental retardation and various neurological deficits. Serological diagnosis of primary CMV in the mother is difficult because IgM antibodies persist for long periods of time and both IgG and IgM antibodies may rise during secondary infection that is generally less dangerous to the human embryo in comparison to primary infection. A more reliable diagnostic procedure is to assess fetal infection by polymerase chain reaction (PCR) and CMV culture performed in amniotic fluid at least 6 weeks following presumed maternal infection and past the 21st week of pregnancy. Due to the difficulties in serological diagnosis and the common lack of typical clinical findings, there seems to be no reason to perform routine serological studies for CMV in pregnant mothers or in women prior to planning pregnancy. The best preventive measure is appropriate immunization against CMV, but no effective immunization seems to exist as of today. PMID- 12119776 TI - [Geriatrics 2002]. PMID- 12119777 TI - [Look to the Helse Nord!]. PMID- 12119778 TI - [Travel medicine and advice to travellers]. PMID- 12119779 TI - [20 years without progress?]. PMID- 12119780 TI - [High-resolution computer tomography in cystic fibrosis]. AB - BACKGROUND: Worsening lung disease is the most important factor concerning quality of life and survival rate in the approximately 250 patients with cystic fibrosis in Norway. Pulmonary high-resolution CT has been advocated as a precise diagnostic method, that offers the opportunity to detect slight disease progression. MATERIAL AND METHODS: We examined 21 patients (age 6-34) with high resolution CT. The findings were scored using a modified Bhalla method, where score 0 is normal and score 27 reflects a maximal degree of e.g. bronchiectasis and mucous plugging. RESULTS: Mean high-resolution CT score was 8.0 (range 0-22). Bronchiectasis was found in 17 out of 21 patients, peribronchial wall thickening in 15 patients and mucous plugging in 14 patients. The Bhalla score showed a close correlation with forced expiratory volume in one second (r = -0.844, p < 0.01). INTERPRETATION: High-resolution CT characterises very well the lung pathology in patients with cystic fibrosis. The degree of pathology assessed with a modified Bhalla score correlates well with forced expiratory volume in one second. PMID- 12119781 TI - [Hurler's syndrome--early clinical suspicion]. AB - BACKGROUND: Hurler's syndrome is a rare congenital metabolic disorder and is inevitably lethal when untreated. The presenting symptoms are usually vague, resembling those found in otherwise healthy children. MATERIAL AND METHODS: We present a patient with Hurler's syndrome and discuss the unspecific clinical signs and symptoms seen in these patients based on a review of relevant literature. RESULTS: A two-months-old infant boy was operated for a rightsided inguinal hernia. The anaesthesiologist had difficulty intubating the patient. During the following months the patient had a severe allergic reaction to vaccination, chronic rhinitis with recurrent upper airway infections, and diarrhoea and vomiting. Eventually, failure to gain weight, and psychomotoric delay led to a thorough clinical investigation. Extensive X-ray diagnostics and analysis of urine and serum concluded with Hurler's syndrome. INTERPRETATION: Clinical vigilance is needed in the diagnosis of Hurler's syndrome. Early symptoms, such as rhinitis and hernia, are vague and unspecific. Radiological features, such as broad costae or gibbus, are helpful in obtaining the diagnosis. Decreased level of alpha-L-iduronidase in serum confirms the diagnosis of Hurler's syndrome. Patients tend to be treated symptomatically before the eventual diagnosis of this syndrome. PMID- 12119783 TI - [Postmenopausal hormone replacement therapy and cardiovascular disease]. PMID- 12119782 TI - [Board of appeals for decision on reimbursement of expenses for medical treatment abroad]. AB - In Norway, a National Insurance Board decides whether patients treated abroad should have their expenses reimbursed. Its decisions may be appealed to an independent governmental board of appeals. This article presents the appeals procedure and discusses the experience gained over its first three years. The most important criterion for reimbursement is whether there is a lack of expertise in Norway on the relevant condition, which must be severe. A statement must be sought from a tertiary-level hospital about the proposed intervention. A referral or recommendation is not required. Reimbursement will not be considered for interventions defined as experimental, except in conditions that are so rare that the number of patients in Norway will be so small that one cannot expect large-scale randomised trials organised in this country. During its first three years, the board of appeals considered 184 cases; decisions were revered in 21 cases (11%). The appeals procedure remains well known among the public. Doctors should make themselves familiar with the criteria and the appeals procedure. PMID- 12119784 TI - [Health hazards for immigrants when travelling to their home countries]. AB - Vacations in the home country are important and positive events in the lives of immigrants, events that allow them to maintain contact with their culture, relatives and friends. However, vacations also carry certain health risks, though these risks can to some degree be prevented. Infectious disease is the greatest risk. Some children and adolescents also run the risk of female genital mutilation, forced marriage, and the risk og being left behind in the home country against their will. Among the notifiable diseases registered with the Norwegian Surveillance System for Communicable Diseases (MSIS), five stand out as having a higher incidence in people of foreign background than in people of Norwegian origin: malaria, hepatitis A, shigella infection, typhoid and paratyphoid fever. This higher incidence is partly the result of less use of pre travel vaccines and malaria prophylaxis. Immigrants as a group are exposed to varied risks and should be given high priority in relation to vaccines and malaria prophylaxis for travel abroad. High priority should also be given to preventive health measures designed to reduce the risk of female genital mutilation and other violations against children and young people on visit to their country of origin. PMID- 12119785 TI - [Young children, pregnant women and travelling abroad]. AB - Pregnant women and parents of young children travelling to non-western countries should consider the risks to which they expose themselves and their children. Travelling during these periods of life needs to be particularly well planned. Travel insurance should cover the whole family, and for pregnant women also the risk of premature birth. Travelling long distances during pregnancy involves a certain amount of risk in itself. This risk could be increased if complications were to occur in areas with a lower standard of health service. As a rule, infants and young children easily adapt to new environments but children abroad should be expected to need a doctor at least as often as at home. Some vaccines and antimalarials must not be used for children below a certain age. Only a few vaccines and antimalarials have been systematically studied in pregnant women in order to exclude teratogenicity. We present some aspects of vaccination and malaria prevention, transport, climate and environment, nutrition, food and drinking water hygiene. PMID- 12119787 TI - [Air travel and venous thrombosis]. AB - BACKGROUND AND METHODS: The paper reviews air travel and venous thrombosis. The risk of venous thrombosis increases with distance flown. The mechanisms that cause thrombosis are unknown, but may be related to hypobaric hypoxia, i.e. reduced air pressure inside the aeroplane, stasis of the lower limbs, often referred to as economy class syndrome (ECS), and dehydration. RESULTS: Symptomless deep venous thrombosis may occur in 10% of healthy long-haul airline travellers over 50 years of age, and up to 4.5% of passengers under 50 at high risk of thrombosis. Wearing of elastic compression stockings during the flight is associated with reduced risk of deep venous thrombosis. INTERPRETATION: Passengers should be encouraged to use the muscle-vein pump regularly, ensure adequate hydration, and in some cases wear elastic compression stockings. A majority of passengers will not need any thromboprophylaxis. Aspirin may have a protective role, but its efficacy in preventing venous thrombosis is much smaller than that of low molecular weight heparins. A high prophylactic dose of a low molecular weight heparin should be considered in passengers at high risk of thrombosis. PMID- 12119788 TI - [Physician's responsibility for the nursing home culture]. PMID- 12119786 TI - [Appropriate health insurance for traveller abroad]. AB - BACKGROUND: Increasing numbers of Norwegians are residing abroad temporarily or permanently; one out of three of them will contract illness. There is, however, uncertainty as to what coverage various insurance plans provide. MATERIAL AND METHODS: The insurance terms and conditions referred to in this study have been obtained from the biggest Norwegian insurance companies and from the national insurance scheme in Norway. RESULTS: The national insurance scheme only provide partial coverage of expenses for its members if they fall ill abroad. To be ensured full coverage, travellers must have additional private medical insurance. INTERPRETATION: The terms offered by insurance companies are unclear and should be restated so that individuals travelling abroad know with certainty whether or not they are covered. New types of insurance policies should be offered so that all travellers can be sure that their expenses will be covered in case of illness abroad. PMID- 12119789 TI - [Elderly physicians--and retired physicians]. PMID- 12119790 TI - [Empty guidelines concerning acute low back pain]. PMID- 12119791 TI - [Specialist service for functional diseases]. PMID- 12119792 TI - [Treatment is physician's responsibility!]. PMID- 12119793 TI - [Psychiatric training is no longer compulsory for neurologists]. PMID- 12119794 TI - The global burden of hypertension: good news and bad news. AB - Hypertension is a common disorder and a powerful risk factor for death and disability from heart disease and kidney failure. Worldwide, it affects an estimated 690 million persons and it is the major risk factor for stroke. The good news is that safe and effective treatments are available and in the majority of patients, hypertension can be diagnosed easily and blood pressure can be controlled. Unfortunately, most hypertensive patients have uncontrolled blood pressure. To effectively address this situation, a renewed commitment to refine strategies for controlling blood pressure is necessary. Additional emphasis is also warranted for the prevention of hypertension in the first place. PMID- 12119795 TI - Hypertension detection and control: population and policy implications. AB - The decline in cardiovascular diseases is by far the most remarkable achievement of medicine in the last half of the twentieth century. It can even be said that the treatment of hypertension is the only known medical intervention to have left a clear imprint on mortality trends. Much more remains to be accomplished, however, before even the majority of patients in the United States will be controlled with pharmacologic therapy [27,28]. Significant changes are needed to improve the practice of institutions, the adherence of physicians to appropriate guidelines, and the consistency of pill taking on the part of patients. Global risk evaluation is currently being used more widely in clinical practice to target therapy and improve its overall effectiveness; however, it is perhaps too early to assess its practical value. On theoretic grounds alone, much more needs to be done to refine prediction of cardiovascular risk as a clinical tool. Improvement in information technology, including an electronic medical record and on-line risk equations, will also be required before its full value can be realized. PMID- 12119796 TI - Pathogenesis and clinical physiology of hypertension. AB - Although the definitive cause of primary hypertension remains unknown, its pathogenesis and clinical history are relatively well understood. Genes, environment, and their interaction play crucial roles in the development of hypertension. Elevated blood pressure levels are maintained through alterations in multiple BP regulatory systems including the central and peripheral nervous system, renin-angiotensin-aldosterone system, renal mechanisms, structural and functional changes in the vascular wall and endothelium, and multiple feedback loops involving a wide spectrum of receptors and. PMID- 12119797 TI - Principles and techniques of blood pressure measurement. AB - The gold standard for clinical blood pressure measurement continues to be readings taken by a physician using a mercury sphygmomanometer, but this is changing as mercury is gradually being phased out. The oscillometric technique, which primarily detects mean arterial pressure, is increasingly popular for use in electronic devices. Other methods include ultrasound (used mainly to detect systolic pressure) and the finger cuff method of Penaz, which can record beat-to beat pressure noninvasively from the finger. The preferred location of measurement is the upper arm, but errors may occur because of changes in the position of the arm. Other technical sources of error include inappropriate cuff size and too rapid deflation of the cuff. Clinic readings may be unrepresentative of the patient's true blood pressure because of the white coat effect, which is defined as the difference between the clinic readings and the average daytime blood pressure. Patients with elevated clinic pressure and normal daytime pressure are said to have white coat hypertension. There are three commonly used methods for measuring blood pressure for clinical purposes: clinic readings, self monitoring by the patient at home, and 24-hour ambulatory readings. Self monitoring is growing rapidly in popularity and is generally carried out using electronic devices that work on the oscillometric technique. Although standard validation protocols exist, many devices on the market have not been tested for accuracy. Such devices can record blood pressure from the upper arm, wrist, or finger, but the arm is preferred. Twenty-four-hour ambulatory monitoring has been found to be the best predictor of cardiovascular risk in the individual patient and is the only technique that can describe the diurnal rhythm of blood pressure accurately. Ambulatory monitoring is mainly used for diagnosing hypertension, whereas self-monitoring is used for following the response to treatment. Different techniques of blood pressure measurement may be preferred in certain situations. In infants the ultrasound technique is best, whereas in pregnancy and after exercise the diastolic pressure may be hard to measure using the conventional auscultatory method. In obese subjects it is important to use a cuff of the correct size. PMID- 12119798 TI - The heart, kidney, and brain as target organs in hypertension. AB - The heart, kidney, brain, and arterial blood vessels are prime targets of hypertensive damage. Uncontrolled hypertension accelerates the damage to these organs and results in eventual organ failure and cardiovascular death and disability. Current guidelines for the appropriate treatment and control of hypertension requires an assessment of the presence of target organ damage. When present, evidence of target organ damage determines the urgency and intensity of drug treatment and may also dictate the choice of initial antihypertensive drug class. Thus, evaluation of persons with suspected or established hypertension must include a meticulous search for evidence of target organ damage. Fortunately, treatment with all antihypertensive medications that results in significant BP reductions also reduces fatal and nonfatal hypertensive complications and significantly slows down the progression to organ failure. Because of the important role that adverse activation of the renin-angiotensin aldosterone system plays in target organ damage, drugs that antagonize this system have provided consistent and compelling proof of organ protection in both primary and secondary prevention of adverse outcomes. The challenge now is to use these and all other antihypertensive agents effectively to control BP to target levels in patients with hypertension. Continued emphasis on the adoption of lifestyle changes for prevention of hypertension in the first place or as adjunctive therapy in hypertensive patients is essential. PMID- 12119801 TI - Resistant hypertension, secondary hypertension, and hypertensive crises. AB - Resistant hypertension, secondary hypertension, and hypertensive crises are uncommon but potentially dangerous forms of hypertension that are associated with an increased risk of complications such as myocardial infarction, heart failure, stroke, and renal failure. Appropriate diagnostic screening and selective drug or surgical management can reduce the risk of these complications dramatically. In compliant patients, resistant hypertension occurs most often in obese patients receiving inadequate diuretic therapy. In patients with clinical clues to the diagnosis, the best current screening test for renovascular hypertension is probably the ACE-inhibitor renal scintiscan. Angioplasty is considerably more successful in younger patients with fibrous dysplasia than in older patients with the atherosclerotic variety. Hypertensive crises are divided into BP urgencies and emergencies. In both settings, the reduction in BP should generally be gradual rather than abrupt, with no intent to acutely normalize the BP. PMID- 12119799 TI - Nondrug interventions in hypertension prevention and control. AB - This review was undertaken to address the relation of various factors to HBP and their potential for preventing and controlling this widespread problem. With respect to salt intake and BP, the 1999 Workshop on Sodium and Blood Pressure of the (US) National Heart, Lung, and Blood Institute [5] will serve the reader well as a point of departure. The body of the present review provides more detailed discussion especially of recent epidemiologic research, including the DASH-Sodium trial, published more recently than the proceedings of that workshop. The DASH Sodium trial demonstrates significant increases in SBP and DBP, with sodium intake greater than 65 mmol/d (= 3.7 g NaCl--see equivalencies in Appendix A) and with the usual American diet (versus the DASH diet). These results provide substantial evidence against current dietary practices in many populations where daily intakes of salt are much higher than recommended. We also have addressed alcohol consumption, micronutrients/macronutrients, physical activity and inactivity, obesity, cigarette smoking, and alternative approaches to treatment such as stress reduction/biofeedback, yoga/meditation, and acupuncture. Evidence for the efficacy of certain nonpharmacologic approaches to preventing and controlling HBP is strong. This evidence offers a basis for public health policies and clinical approaches that can greatly affect the incidence and consequences of HBP in the population at large. What is needed now is implementation of the policies and practices addressed here. Unless such action is taken on a large scale, we will have made poor use of the knowledge accrued over decades of research. The clinician is referred to the National Heart, Lung and Blood Institute Web site at www.nhlbi.gov/health/prof/heart/index.htm for resource and guideline information for hypertension. Patients and the general public are referred to the sister web page at www.nhlbi.gov?health?public?heart?index.htm for educational fact sheets and general information on hypertension. PMID- 12119800 TI - Clinical pharmacology of antihypertensive drugs. AB - Systemic hypertension is a major public health problem and is perhaps the most common chronic disorder in most societies. Most patients with vascular disease report hypertension in their medical history. Irrespective of the specialty that one practices, every physician will likely encounter patients with systemic hypertension. Unfortunately, an overwhelming number have so-called "primary" or "essential" hypertension for which a cure has yet to be found. Fortunately, excellent therapy is available to control this modern malady. The field of hypertension continues to evolve rapidly, particularly in the field of therapy. During the past two decades, the treatment of hypertension has moved from a cookbook approach to more scientifically based individualized management. This paradigm shift requires the practitioner to acquire sufficient knowledge about individual drugs and how they work in a given patient. Rapid expansion of available drugs has placed a burden on the clinician to keep up with these advances. We hope that the discussions contained herein will ease that burden somewhat and make the treatment options less cumbersome. This article addresses the practical issues related to selection of antihypertensive drugs and provides an overview of advantages and disadvantages of individual drug classes. The reader should also refer to the JNC VI document [1] to further understand the selection of drug therapy based upon compelling indications. The ultimate aim of hypertension management should always be to achieve target or goal blood pressure levels. PMID- 12119802 TI - Hypertension in orthostatic hypotension and autonomic dysfunction. AB - Disabling orthostatic hypotension dominates the clinical picture of autonomic failure. Nonetheless, severe supine hypertension is observed in about 50% of patients. In patients with multiple system atrophy (Shy-Drager syndrome), supine hypertension is explained by residual sympathetic tone because it can be eliminated with the ganglionic blocker trimethaphan. The cause of hypertension in patients with pure autonomic failure is not known and its understanding may be relevant to essential hypertension. Supine hypertension complicates the treatment of these patients but can be managed by overnight administration of antihypertensive medications. PMID- 12119803 TI - Effective strategies for blood pressure control. AB - Of the estimated 50 million Americans suffering from hypertension, less than a fourth have it under control. One-year costs of failing to control hypertension were recently estimated at $964 million for the total US hypertensive population and $467 million for those being treated. This article will describe effective strategies for blood pressure control at national, state, and community levels. PMID- 12119804 TI - Clinical hypertension. PMID- 12119805 TI - Drugs and children. PMID- 12119806 TI - 3,941,553 births in 1998: including tens of thousands with congenital anomalies and abnormal conditions. AB - Variations are occurring in the birth rates of different population and age groups in this country. In addition to these ongoing developments, there continues to be an increasing number of children born with congenital anomalies and birth injuries. A summary is provided of these developments (based on birth certificate reports) for the general population during the 1990s, and for newborns with congenital anomalies and birth defects during and past twenty-five years. PMID- 12119807 TI - Health of U.S. children in the late 1990s. AB - A summary is presented of the finding from the 1997 National Health Interview Survey on the health of children. Despite major improvements in general health levels, many millions of children continue to have significant unmet general health and oral health needs. PMID- 12119808 TI - Comparison of two tooth-saving preparation techniques for one-surface cavities. AB - The atraumatic restorative treatment technique (ART) is based on removing infected tooth material using only hand instruments and filling the subsequently cleaned cavity with adhesive material such as glass ionomer. As its name suggests, the ART technique should be atraumatic during treatment, as well as for the tooth itself as for the patient. It was primarily developed for treating people living in underserved areas of the world where resources and facilities such as electricity and trained manpower are limited. Many studies have evaluated the ART technique and the results have supported its application. However, a very limited number of studies have compared ART with more conventional techniques. For that reason, a study was conducted in Pakistan, to compare the ART technique with another more conventional treatment technique. The results of this study show that the preparations with hand instruments resulted in smaller sized cavities and therefore may be less traumatic to the tooth. It was also associated with less pain reactions compared to the more conventional technique. Although preparations with hand instruments required more time, this did not seem to affect the survival of restorations. The survival of glass ionomer cement restorations made with hand instruments was comparable with single surface amalgam restorations made with a more conventional technique. Recurrent caries was not associated with any glass ionomer cement restorations made with hand instruments. The retention rate of glass ionomer sealants was low, however one dentist had a sealant retention rate of 81.5 percent that suggests that this procedure can be performed satisfactorily in conjunction with a glass ionomer cement restoration. Operator variances did seem to affect the restorations. Survival of glass ionomer restorations made with both hand and rotary instruments varied for different operators. Similarly, the retention of fissure sealant also varied amongst operators. Operator differences also influenced the extent of tooth substance lost due to cavity preparation. The ART technique is a feasible approach towards the treatment of dental caries especially of one-surface lesions for underserved populations. PMID- 12119809 TI - Cvek pulpotomy: report of a case with five-year follow-up. AB - Partial pulpotomy (Cvek pulpotomy) is the treatment of choice for injured permanent incisor teeth with exposed vital pulp tissue and immature apices. This treatment preserves pulpal function, thus allowing continued root development. The present report describes the case of a permanent incisor with incomplete root end closure that underwent a Cvek pulpotomy, with subsequent apical closure. Five years post pulp therapy, the tooth remained symptom free. PMID- 12119810 TI - Orthodontic treatment in handicapped children: report of four cases. AB - Mentally and physically handicapped children show in the orofacial system motor sensitivity disturbances and malocclusions of varying severity. These dysfunctions affect the breathing and speech ability and inhibit the food intake. Myotherapeutic exercises for strengthening of lip and tongue muscles and orthodontic treatment of the malocclusions help provide esthetic and functional improvements in these patients. The limited compliance necessitates a differentiated procedure during the diagnostic and therapeutic process and demands compromises in some cases. PMID- 12119811 TI - Acellular dermal matrix allograft in the treatment of mucogingival defects in children: illustrative case report. AB - Mucogingival defects can occur in children and are of particular concern when orthodontic treatment is indicated. The rationale for surgical intervention is predicated on the need to repair the mucogingival defect and to establish adequate thickness of attached gingiva. The free gingival graft, usually obtained from the hard palate, is often used to increase the amount of attached gingiva. The prospect of a second surgical site, and its inherent risks and complications, which may include pain, discomfort, and bleeding, is especially undesirable in children. Important to consider is the possibility that a child may not have adequate tissue thickness at the donor site. A case report is presented utilizing the alternative soft tissue graft, Alloderm, to correct a mucogingival defect prior to orthodontic treatment. Adhering to the free gingival autograft technique, an acellular dermal matrix allograft was utilized at the graft site. The patient revealed good post-operative healing, tissue vascularization, and a healthy zone of attached gingiva at the six month follow up visit. Comparable results to the conventional autograft were obtained with less surgical time, surgical sites, and discomfort to the patient. PMID- 12119813 TI - Dens invaginatus in a primary molar: report of case. AB - A case of dens invaginatus in a mandibular second primary molar of an eleven-year old boy is presented. The tooth was extracted and examined by scanning electron microscopy. SEM findings demonstrated the presence of defective enamel and cementum in the pulp chamber. Dentinal tissues were also irregular and had fewer and thinner tubules. This case of dens invaginatus in primary molar is an unusual case of the malformation being the only one in the literature. PMID- 12119812 TI - Comparison of ferric sulfate, formocresol, and a combination of ferric sulfate/formocresol in primary tooth vital pulpotomies: a retrospective radiographic survey. AB - Studies have suggested that formocresol has toxic and carcinogenic potential. A search for an alternative medicament for primary tooth pulpotomies has led to ferric sulfate as a possible alternative. A retrospective study was done in a multipractitioner IHS (Indian Health Service) clinic. Radiographic success or failure was determined for 202 primary tooth pulpotomies performed with either formocresol, ferric sulfate, or a combination procedure of formocresol and ferric sulfate. The post-operative period for the pulpotomies ranged from one month to thirty-six plus months. There was no statistical difference in radiographic failure rates between formocresol, ferric sulfate, or the combination procedure when results were analyzed regardless of post-op period. However, when post-op periods were considered, formocresol performed better at > 36 months and the combination procedure showed significantly more failures at > 36 months. PMID- 12119814 TI - Maxillary canine transpositions in two brothers and one sister: associated dental anomalies and genetic basis. AB - Transposition is an uncommon dental anomaly involving positional interchange of two teeth. The maxillary canine is the tooth more frequently transposed in man. Maxillary canine-first premolar appears to be the most common type of tooth transposition, followed by maxillary canine-lateral incisor transposition. Maxillary canine transpositions are frequently associated with other dental abnormalities such as agenesis and pegshaped incisors. This report describes the presence of transposed canines in one sister and two brothers. The female showed bilateral maxillary canine-first premolar transposition with the left canine fully mesial to its neighboring first premolar, and the right canine blocked-out facially between the first and second premolar. One of the brothers showed full maxillary left canine-lateral incisor transposition. The other brother showed maxillary canine-first premolar transposition and agenesis of maxillary lateral incisors, with the left canine blocked-out facially between the first and second premolar. Findings from this case report and other previously published cases provide strong evidence that maxillary canine transpositions are a disturbance of tooth order and eruptive position resulting from genetic influences within a multifactorial inheritance model. PMID- 12119815 TI - A study of pathology associated with short lingual frenum. AB - Ankyloglossia is a developmental anomaly of the tongue characterized by a short, thick, lingual frenum resulting in limitation of tongue movement. The frenum is attached to the tip of the tongue in varying degrees. This study evaluates short lingual frenum and other associated pathologies (dentofacial anomalies and lingual dysglossia) and investigates whether ankyloglossia is related to speech problems. A thorough intraoral examination was carried out using the WHO regulations for dentofacial anomalies and occlusion problems and a new scale of lingual mobility. This article also discusses the difficulty in diagnosing this condition and finally following to the results of our study, describes the indications for nonsurgical and surgical treatment of this anomaly. PMID- 12119816 TI - Multi-rooted mandibular premolars: report of case. AB - Differential growth of the epithelial diaphragm is needed for the formation of multiple roots in the primary and permanent dentitions. In most instances, the mandibular premolars possess a single root. It is a very unusual event to find a case of bilateral multi-rooted first and second mandibular premolars. This paper presents such a case. PMID- 12119817 TI - Oral self-mutilation in the Lesch-Nyhan syndrome. AB - Lesch-Nyhan Syndrome is a disorder caused by congenital absence of a purine metabolic enzyme, hypoxanthine-guanine phosphoribosyl transferase (HPRT). This syndrome is characterized clinically by mental retardation, chorea, athetosis, hyperuricemia, uricosuria and self-mutilation. This report is of two children, who are cousins, both of whom have Lesch-Nyhan syndrome and presented with severe self-mutilation wounds on their lip(s). Vital pulpotomy and coronal resection was done as a more conservative approach than extracting all offending teeth. By maintaining the root portion of the teeth in the bone, it is expected that preservation of the alveolar bone will be achieved. PMID- 12119818 TI - Menkes disease: report of two cases. AB - Menkes disease is a rare, autosomal recessive disorder characterized by neuronal degeneration, abnormal hair, malformed connective tissue, mental retardation, and a life span of three years. Previously reported dental findings include a high arched palate, delayed eruption of secondary dentition, and open bite. The case of twin seven-year-old males with Menkes disease is presented, along with previously unreported dental findings of spindle-shaped root resorption patterns on the primary maxillary central and lateral incisors. PMID- 12119819 TI - Maxillary molar transposition. AB - To date, five types of maxillary tooth transposition are described in the international literature (canine/first premolar; canine/lateral incisor; lateral incisor/central incisor; canine/central incisor site; canine/first molar site), although the latter two are indeed extreme displacements and not true transpositions. In the present article a sixth type of transposition is recognized and described: the maxillary molar transposition (MxM3M2 and MxM4M3, respectively). Three cases of this anomaly were presented. The occurrence is extremely rare, about 0.04 percent in a sample of 7,000 orthodontic patients. Investigation of other family members (especially parents and siblings) could not substantiate an hypothesis of inheritance of this anomaly. Other possible causes (trauma, caries, tooth agenesis) were also absent in these cases. PMID- 12119820 TI - Parents' self-reported compliance with preventive practices after witnessing their child undergo intravenous sedation for dental treatment. AB - The purpose of this study was to evaluate the self-reported compliance of parents with preventive dental care measures after witnessing their child undergo full mouth rehabilitation under i.v. sedation. Records of 251 pediatric patients who underwent full-mouth rehabilitation under i.v. sedation between 1995 and 1999 at the University Children's Dental Center in Los Angeles, California were evaluated. Demographic data and rates of post sedation follow-up preventive visits were collected. In addition, seventy-five parents participated in a telephone interview and answered five multiple-choice questions that focused on the following areas: nursing habits, diet, hygiene, and follow-up preventive visits pre and post sedation. Parental attitude towards the experience was also assessed. Only 47 percent of the patients returned for at least one post op/recall visit at the Children's Dental Center. Seventy-six percent of parents interviewed stated they made improvements in their child's hygiene post sedation. Only 45 percent of parents stated they made improvements in their child's diet and only 47 percent stated that they increased their child's dental recall visits post sedation. Eighty-eight percent of parents stated that they would be willing to have their children undergo i.v. sedation again if necessary for dental treatment. Parental presence during a child's dental rehabilitation via i.v. sedation does not change the preventive behaviors of these parents post sedation when compared to results of other studies. PMID- 12119821 TI - Effect of xylitol chewing gum on salivary Streptococcus mutans in preschool children. AB - Dental caries remains a significant problem for poor children in the United States. One strategy for treating dental caries is to suppress streptococcus mutans, the chief pathogen responsible for the disease. The purpose of this study was to evaluate the effect of xylitol gum in salivary S. mutans levels in preschool children. Sixty-one children were randomly assigned into the xylitol group and the control group. The xylitol group chewed gum sweetened only with xylitol (XyliFresh100%, Hershey Food Corporation, U.S.A.) three times a day for three weeks. S. mutans counts were tested using the Dentocult-SM Strip Mutans test (Orion Diagnostica, Finland) at baseline and after three weeks. The shift from higher S. mutans scores to lower was greater in the xylitol group than in the control group (p;lt0.05). This study supports the suggestion that chewing xylitol gum may reduce salivary S. mutans levels. Xylitol chewing gum may provide a feasible caries prevention method for preschool children. PMID- 12119822 TI - Factors associated with uncooperative behavior by Brazilian preschool children in the dental office. AB - The purpose of this study was to assess the uncooperative behavior of low income preschool children receiving dental treatment and the factors associated with that behavior. The sample consisted of 177 Brazilian children of both sexes, ages from three to six years. The children were observed during their dental appointments and their behavior was categorized by the Frankl Behavioral Rating Scale. The oral health of the children was evaluated on the basis of a clinical examination and classified using the dmft index. A history taking with the child's parent or guardian provided current medical condition, dental history, and family situation. Of the children, 1 percent presented definitely positive behavior, 59 percent positive behavior, 28 percent negative behavior, and 12 percent definitely negative. A significant association was found between children with negative behavior and the variables of the child's age, parent's or guardian's education, learning or behavioral problems, parent's or guardian's anxiety, child oral health status, general health problems, history of hospitalization, and dental history. PMID- 12119823 TI - Human biting of children and oral manifestations of abuse: a case report and literature review. AB - A ten-month-old female was taken to the Children's Hospital of New York for evaluation of suspected child abuse. The child presented with a severe oral herpetic infection, tongue laceration, and multiple bite marks. Social services confirmed that a parent bit the child's tongue. PMID- 12119824 TI - Rural health issues. AB - The ability to improve one's health and reduce the subsequent risk of disease and disability poses distinct challenges for individuals living in rural areas of the United States. A review of rural health issues provides insight into the dental and general medical realities for 20 percent of the population of this country. PMID- 12119825 TI - The $190 billion conundrum. PMID- 12119826 TI - Broadway: anthrax threat intensifies focus on disaster preparedness. PMID- 12119827 TI - Enforcing prompt-payment regulations: the Texas approach. AB - To ensure that insurance carriers pay providers in a timely manner, Texas has adopted strict payment regulations. Enforcement of the regulations has led to restitution payments for many providers. However, issues such as clean claims, underpayment, discrepancies in payment dates, and self-funded claims continue to present challenges. PMID- 12119828 TI - The hospital cost index: a new way to assess relative cost-efficiency. AB - Hospital senior financial executives routinely review measures of facility costs to assess their organization's cost-efficiency. The measures they typically use for this activity are cost per adjusted discharge and cost per adjusted patient day. These measures have flaws that limit their usefulness for comparing cost efficiency of different hospitals. A more effective measure is the hospital cost index (HCI), which adjusts for case-mix complexity in both inpatient and outpatient operations and can be computed from publicly available databases. The highly reliable HCI enables senior financial executives to make comparison among specific hospitals, including direct competitors. PMID- 12119829 TI - Quantifying value for physician order-entry systems: a balance of cost and quality. AB - Healthcare CFOs commonly demand hard data to prove that an investment in computerized physician order entry (CPOE) will be worthwhile. However, a balanced analysis of cost and quality of the CPOE system has advantages over traditional return-on-investment appraisals. Montefiore Medical Center (MMC), Bronx, New York, assessed the value of its CPOE system by quantifying cost and quality measures rather than relying solely on dollar returns. MMC collected process times for medication ordering before and after the CPOE system was introduced and demonstrated that the use of CPOE increased medication ordering efficiency by 92 percent. MMC calculated additional process times for ward clerks, nurses, and pharmacists before and after CPOE implementation and determined the time saved per employee. From that number, the dollars potentially saved per employee and total potential dollar value of time savings per year were calculated. PMID- 12119830 TI - Labor analytics software can help control labor costs. AB - One of the largest costs in healthcare organizations is labor. As volume of activity fluctuates, the labor supply needs to be adjusted appropriately. Having an appropriate quantity and skill mix of full- and part-time employees affects labor costs per hour and labor efficiency, two labor-cost control points that department managers need to monitor to make effective hiring decisions. Managers can use software applications for operating budgets, productivity monitoring, position control, and staffing/scheduling to manage labor expenses. Knowing the type of information each of these cost-control tools provides is critical to making effective decisions related to labor expenses. Because of their unique expertise, healthcare financial managers can serve as a resource to help the organization's department managers manage labor costs. PMID- 12119831 TI - CMS issues new forms for Medicare provider enrollment. AB - In its continual efforts to manage Medicare provider enrollment, CMS has revised its provider application forms. Along with the revised forms is a new requirement for a three-year cycle revalidation process that could present a considerable challenge to large providers. Also of note are longer processing periods for initial enrollment, suspension of payments if current address information is not maintained, and deactivation of a provider's number if 12 months pass without claim activity. Improvements that providers may appreciate include electronic access to forms and a decrease in the amount of information required to enroll. PMID- 12119832 TI - Managing organizational improvement in a resource-challenged environment. PMID- 12119833 TI - Providers may incur payer penalties under HIPAA. PMID- 12119834 TI - Aim for success with a career action plan. PMID- 12119835 TI - Sowing the seeds of long-term care insurance growth. PMID- 12119836 TI - Be proactive and avoid costly lawsuits. PMID- 12119837 TI - Breaking free of restraints. PMID- 12119838 TI - Three myths about HIPAA compliance. PMID- 12119839 TI - Consumerism: automating patient and provider interactions. AB - Healthcare is essentially an information industry that is seeking to automate its processes and use Internet technology to support its consumers. The healthcare industry understands that, without computerization, it would be nearly impossible to reduce medical errors, analyze the delivery of care, and provide real-time clinical decision support to healthcare providers to support patient care via the Internet. Without progressive steps, use of Internet technology to care for patients will be limited. The only way to meet the information challenges of tomorrow is to begin capitalizing on today's technology and implement the necessary tools now. Most important, healthcare organizations need to adopt a full array of technology to support their consumers in the electronic world of today and tomorrow. PMID- 12119840 TI - Nurturing leadership for telemedicine: clinician champions. PMID- 12119841 TI - An interview with John Mack, president of the Internet Healthcare Coalition. Interview by Richard D Lang. PMID- 12119842 TI - Experiences with knowledge application in medical care. AB - IT interventions, such as provider order entry and knowledge repositories, are improving the application of medical knowledge. One health system's experiences are reviewed, including challenges faced and organizational factors that supported the process. PMID- 12119843 TI - The consult is real, the visit is virtual. AB - A web-based service was developed through which patients can obtain second opinions from physicians. Factors addressed included incorporating e-appointments into a medical practice, consolidating different approaches to second opinions, complying with disparate state licensing regulations, and communicating the program's features to providers and patients. PMID- 12119844 TI - Meeting the needs of customers with health CRM. AB - Customer relationship management (CRM) is a business strategy, supported by applications and technologies, that can fundamentally transform how healthcare delivery organizations manage patient and physician interactions, reduce cost, improve customer-facing processes, drive market and revenue growth, and manage regulatory compliance processes. PMID- 12119845 TI - Enterprise identity management strategies. AB - The need for an enterprise-wide person-centric approach to patient identification is presented, along with the challenges to achieving that objective and guidelines to the actions and technology that will support the enterprise view. PMID- 12119846 TI - HIPAA strengthens business case for electronic report distribution systems. AB - HIPAA may finally force healthcare organizations to make long-postponed decisions to increase the use of automation and technology for report distribution. For most, the direct benefits will far outweigh the costs. PMID- 12119847 TI - Training T.I.P.S. PMID- 12119848 TI - Integrated disease management pilot for diabetes. AB - A New Zealand diabetes program integrates management systems for hospitals and physician practices through a shared integrated care server (ICS), which supports collaborative patient management, virtual consults, and clinical feedback. PMID- 12119851 TI - Health Web site accreditation: opportunities and challenges. PMID- 12119850 TI - The transformation of an IS department. AB - The story of one IS department's journey from chaos to efficiency all the way to innovative and award-winning is told, including details of significant changes in approach, technology, processes, and quality measurement. PMID- 12119849 TI - Telemedicine in Kansas 1994-2001. AB - A longitudinal study of organizational issues affecting telemedicine adoption and diffusion in Kansas is presented. Key strategies highlighted include a centralized scheduling and coordinating process and leadership to oversee state activities. PMID- 12119852 TI - Management and conduct of randomized controlled trials. AB - In preparing to undertake a clinical trial, it may be helpful to keep in mind Fredrickson's description of clinical trials (31): "Field trials are indispensable. They will continue to be an ordeal. They lack glamour, they strain our resources and patience, and they protract the moment of truth to excruciating limits. Still, they are among the most challenging tests of our skills. I have no doubt that when the problem is well chosen, the study is appropriately designed, and that when all the populations concerned are made aware of the route and the goal, the reward can be commensurate with the effort. If, in major medical dilemmas, the alternative is to pay the costs of perpetual uncertainty, have we really any choice?" PMID- 12119854 TI - Sample size calculations for randomized controlled trials. PMID- 12119855 TI - Design of randomized trials. PMID- 12119853 TI - Analysis of randomized controlled trials. AB - Although the sophistication and flexibility of the statistical technology available to the data analyst have increased, some durable, simple principles remain valid. Hypothesis-driven analyses, which were anticipated and specified in the protocol, must still be kept separate and privileged relative to the important, but risky data mining made possible by modern computers. Analyses that have a firm basis in the randomization are interpreted more easily than those that rely heavily on statistical models. Outcomes--such as quality of life, symptoms, and behaviors--that require the cooperation of subjects to be measured will come to be more and more important as trials move away from mortality as the main outcome. Inevitably, such trials will have to deal with more missing data, especially because of dropout and noncompliance. There are fundamental limits on the ability of statistical methods to compensate for such problems, so they must be considered when studies are designed. Finally, it must be emphasized that the availability of software is not a substitute for experience and statistical expertise. PMID- 12119856 TI - Ethics in the design and conduct of clinical trials. PMID- 12119857 TI - Regulatory issues for clinical trials in humans. PMID- 12119858 TI - Special issues related to randomized trials of primary prevention. PMID- 12119859 TI - Community-based health intervention trials: an overview of methodological issues. PMID- 12119860 TI - Economic endpoints in clinical trials. AB - Economic endpoints are increasingly common in randomized clinical trials. As a relatively new addition to the field, methods of measuring and analyzing cost data are still evolving. These developments will be stimulated by the demands of the public for efficient and effective medical care, which will be based on the findings of clinical trials. PMID- 12119861 TI - Beyond intention to treat. PMID- 12119862 TI - Expectations of pediatric sport participation among pediatricians, patients, and parents. AB - As the number of pediatric and adolescent athletes involved in competitive sports continues to grow, and as the competitors in youth sports trend toward a "win at all costs" mentality, pediatricians are increasingly being asked to provide sports medicine treatment and counseling for athletic children. This article outlines the demographic changes in the pediatric and adolescent athlete population in the United States and explains how the pediatrician can become a more effective caregiver to the athletic patient. PMID- 12119863 TI - Pediatric neurodevelopment and sports participation. When are children ready to play sports? AB - A fundamental knowledge of normal child and adolescent development is essential to providing a developmentally appropriate sports experience for the child, and to providing guidance to parents regarding their child's sport participation. This article reviews neurodevelopment, normal child and adolescent development relevant to sport participation, and developmental readiness to participate in sports. Neurodevelopmental maturation is a complex, continuous process. The sense of social comparison is not achieved until after 6 years of age, and the ability to understand the competitive nature of sports is generally not achieved until 9 years of age. By about 12 years of age, most children are mature enough to comprehend the complex tasks of sports and are physically and cognitively ready to participate in competitive sports with appropriate supervision. PMID- 12119864 TI - Special issues and concerns for the high school- and college-aged athletes. AB - Increasing numbers of high school- and college-aged students are participating in sports. As sport participation and intensity increases, the frequency of associated heat related illnesses and acute and chronic overuse injuries will continue to become more prevalent. The sports medicine physician plays an essential role not only in the practice and treatment of injuries but also in educating athletes about safe and healthy exercise habits that will provide life long benefits. PMID- 12119865 TI - The female athlete. Before and beyond puberty. AB - As the twentieth century progressed, the female athlete became an accepted participant of sports at all levels. This article reviews various aspects of female sports participation. After an historical perspective, selected comments are provided on psychologic and physiologic aspects. Concepts of adolescent gynecology are reviewed, including breast and menstrual problems and pregnancy. Other areas reviewed include iron deficiency anemia, stress urinary incontinence, and sports injuries in female athletes. PMID- 12119867 TI - Upper extremity injuries in youth sports. AB - This article provides an overview of common upper extremity injuries in youth sports, including injuries to the shoulder, elbow, and wrist. Pain in the shoulder and elbow is common among youngsters who participate in throwing sports, raquet sports, and swimming, while wrist pain is common among young gymnasts. Acute trauma to the shoulder and elbow can occur in almost any sporting activity. This article provides descriptions of common injuries and guidelines for treatment. PMID- 12119866 TI - Catastrophic pediatric sports injuries. AB - The high school sports of wrestling, gymnastics, ice hockey, baseball, track, and cheerleading should receive closer attention to prevent injury. Safer equipment and sport-specific conditioning should be provided and injuries strictly monitored. Greater attention must also be paid to swimming and diving techniques, and continued observation is needed for heat stroke and heat intolerance in sports such as football, wrestling, basketball, track and field, and cross country. An increased awareness of commotio cordis in sports other than baseball should include ice hockey, football, track field events, and lacrosse. American football because of the sheer numbers and associated catastrophic injury potential must continue to be monitored at the highest medical levels! PMID- 12119868 TI - Lower extremity injuries in youth sports. AB - The lower extremity is the most commonly injured anatomic area in sports. An appropriate history and physical examination is sufficient to diagnose most injuries. By knowing the position where a joint has the greatest and least bony stability, one can predict the likelihood of bony and ligament injury. Such information is helpful in determining which structures are most susceptible to injury as well as identifying when radiographs or other diagnostic studies are indicated. An accurate diagnosis is crucial for planning effective therapy and in determining the need for surgical referral. An understanding of common injury patterns is also helpful in differentiating sources of musculoskeletal pain that are not injury related. PMID- 12119869 TI - Low back pain in the adolescent athlete. AB - There are several other entities such as infection, tumors, and fractures that I have not covered in this article. These entities are not common in adolescent athletes but must always be considered when athletes do no respond to typical treatment protocols for the problems I have discussed. The most important theme to take from this article is that low back pain in adolescent athletes is a problem that should not be ignored but instead fully evaluated because structural problems are quite common in this patient population. PMID- 12119870 TI - Chronic musculoskeletal pain in young athletes. AB - Chronic musculoskeletal pain is becoming increasingly common in young athletes. When these athletes do not respond well to standard treatments, for example physical theraphy and anti-inflammatories, other diagnoses must be considered, such as reflex sympathetic dystrophy, fibromyalgia, and/or overtraining syndrome. PMID- 12119871 TI - The case for large-size mutations. PMID- 12119873 TI - Use of "protective devices" for cellular telephones--technical information statement. PMID- 12119872 TI - The first accurate measurement of systolic and diastolic blood pressure. PMID- 12119874 TI - Golden accomplishments in biomedical engineering. PMID- 12119875 TI - Voices of experience. Interview by Frederik Nebeker and Michael Geselowitz. PMID- 12119876 TI - From Einthoven's galvanometer to single-channel recording with electronics, neuroscientists get inside the membrane. PMID- 12119877 TI - The truth shall set you free. Development of the polygraph. PMID- 12119878 TI - [Therapy refractory intervertebral disk complaints. When is minimally invasive treatment possible?]. PMID- 12119880 TI - [Is it Alzheimer disease? Recognizing the first signs of dementia syndrome]. AB - Today, early diagnosis of different forms of dementia is possible with a relatively easy-to-apply set of diagnostic tools. The quality of the medical history, including the information supplied by relatives and friends, together with the psychological and physical findings, determine the diagnostic hit rate. Laboratory investigations and imaging techniques are indispensable for detecting the underlying cause of existing dementia. Standardized rapid screening procedures for the identification of mild cognitive deficits have speeded up the reliable selection of suspicious cases needing referral to a specialist. Identification of individual day-to-day problems and non-cognitive disorders round off the diagnosis of dementia, and establish the conditions for planning a suitable treatment strategy. PMID- 12119879 TI - [Still many uncertainties in stroke management. New guidelines should change this]. PMID- 12119881 TI - [Starting Alzheimer therapy in early stages whenever possible. Activities of daily living remain intact longer]. AB - In recent years, the efficacy of symptomatic antidementive drugs in the treatment of Alzheimer's disease (AD) has been well documented. A prerequisite for maximally effect antidementive treatment is early diagnosis and a subsequent specific diagnostic clarification. A further essential is early initiation of treatment to delay progression of the disease and thus early loss of daily skills and independence ending in the need for intensive nursing care. Currently, cholinesterase inhibitors, the efficacy of which has been confirmed in placebo controlled multicenter studies, are recommended for the treatment of mild to moderate AD. Further substances with proven efficacy are memantine, ginkgo biloba extract EGb761 and certain classical nootropics. To treat behavioral and other psychological disturbances, symptom-related substances such as selective serotonin reuptake inhibitors and atypical neuroleptics should be employed. In addition to their positive effect on cognitive disturbances, cholinesterase inhibitors also have an appreciable impact on concomitant psychopathological symptoms. PMID- 12119882 TI - [Family physician or gastroenterologist?]. PMID- 12119883 TI - [New approaches in therapy of dementia. Can Alzheimer disease be prevented soon?]. PMID- 12119884 TI - [Aortic stent versus open prosthesis implantation. How can the aneurysm be excluded?]. AB - The diagnosis of an infrarenal aortic aneurysm mandates not only regular ultrasonographic monitoring, but also careful instruction of the patient about an emergency, that is, symptoms associated with rupture, and how to react. If ultrasound reveals a clear increase in the size of the aneurysm, or if a diameter of 5 cm is reached, the indication for surgery is established. Two options are then available: implantation of an aortic stent, which has the advantage of being a minimally invasive procedure, or open prosthesis implantation. However, the benefits and risks of both options must be carefully weighed up, since the patients are often elderly and have cardiopulmonary problems. Postoperative surveillance of the patient comprises three- to six-monthly follow-up with ultrasound or CT scan. PMID- 12119885 TI - ["Diabetic autonomic neuropathy" series. Epidemiology, pathogenesis, basic therapy and prognosis]. PMID- 12119886 TI - [Medicine behind bars. Illness is usual not grounds for modifying imprisonment]. AB - The article describes the possibilities of medical care in prison as exemplified by the Kassel 1 prison with its attached central hospital facility. The main areas of medical care covered there are the treatment of wounds, conservative treatment of fractures, ophthalmology, ENT, urology, dentistry and, via external consultant physicians, also internal medicine, pneumology, dermatology and gynecology. Such infections as hepatitis B and C, syphilis, and tuberculosis have a greater prevalence among prisoners--in contrast to other infections afflicting people housed under similar living conditions, such as in communal living facilities, which show no such increased prevalence. Furthermore, there is a relatively high percentage of injuries, including those that are self-inflicted. The problem of certifying a prisoner medically unfit to tolerate imprisonment is discussed. PMID- 12119887 TI - [Better prognosis in hypertension and left ventricular hypertrophy. LIFE Study sets new standard]. PMID- 12119888 TI - [Coxib spares the stomach. This is true also for patients with preventive aspirin administration]. PMID- 12119889 TI - [Corticosteroid-free therapy for the neurodermatitis patient. Rapid cessation for tormenting urticaria]. PMID- 12119890 TI - [Heart failure. What helps in dyspnea?]. PMID- 12119891 TI - [Preventing chronification. So pain isn't branded into memory]. PMID- 12119892 TI - [New US guidelines. Myocardial infarct is now treated more aggressively]. PMID- 12119893 TI - [TNF-alpha antibody in rheumatoid arthritis. Efficacy-risk relationship is positively evaluated]. PMID- 12119894 TI - [Is dyspepsia an infection? H. pylori eradication is only effective in chronic symptoms]. PMID- 12119895 TI - [There are now evidence-based guidelines for breast carcinoma. There are evidence based guidelines for breast carcinoma. Individual therapy out? (interview by Dr. Judith Neumaier)]. PMID- 12119896 TI - [Hypertensive patient who smokes suffers from dyspnea. Is it the lung, the heart or both?]. PMID- 12119897 TI - [Online risk for psychiatric patients. "Internet suicide" as cause of death]. PMID- 12119898 TI - [Acute abdomen--even if not much therapy is in your hands: your family physician know-how is sought]. PMID- 12119899 TI - [Respiratory manifestations of reflux disease. Gastric acidity--poison for larynx, teeth and respiratory tract]. AB - Gastroesophageal reflux is now a generally accepted risk factor for the development of adenocarcinoma of the esophagus. Less well known is the relationship of reflux disease (GERD) and respiratory disorders. Among the extra esophageal manifestations of reflux disease is reflux laryngitis, which affects up to 78 patients with chronic hoarseness, Reinke's edema, laryngeal stricture, postnasal drip, asthma and non-cardiac chest pain. Despite popular opinion, changes in lifestyle (for example, cessation of smoking and drinking, avoidance of fatty foods) do not result in an improvement in symptoms. The treatment of choice for GERD is the use of proton pump inhibitors (PPI) in the form of stepdown therapy; in individual cases as symptom-orientated on-demand therapy. PMID- 12119900 TI - [Initial measures in reflux manifestations. Endoscopy or therapy attempt with proton pump inhibitors]. PMID- 12119901 TI - [Proton pump inhibitors according to need. Always for heartburn]. AB - Gastroesophageal reflux disease (GERD) is one of the most common medical problems. The large majority of patients so affected have no endoscopically evident disease, or only mild erosive reflux esophagitis. Since both forms of GERD are probably non-progressive, on-demand therapy aimed at adequately controlling symptoms is safe treatment. Proton pump inhibitors (PPI) are the drugs of choice for acute and long-term treatment. In 1999, the first study reporting on on-demand therapy with omeprazole was published. A clear superiority over placebo was found, as was also dependence of the effect on the dose. In the recent past, publications have reported on on-demand therapy with the s-isomer of omeprazole--esomeprazole. With success rates of around 90%, on-demand therapy has proven to be highly effective. On average, the patients take one 20 mg tablet of esomeprazole on every third to fourth day. PMID- 12119902 TI - [Emergencies in general practice. Drowning accidents and "near-drowning"]. PMID- 12119903 TI - [Hypertension in general practice. Not harmless nervousness in the presence of the physician!]. PMID- 12119905 TI - [Dermatologic manifestations of internal diseases. l2: Gout, paraneoplasias and striae distensae]. PMID- 12119904 TI - [Market hypertension, but normotensive in general practice. Much more frequent than assumed!]. PMID- 12119906 TI - [Long-term study with kava special extract. Anxiety symptoms decrease over time]. PMID- 12119908 TI - [General practice without computer communications. Will the money faucet be turned off?]. PMID- 12119907 TI - [Incidental finding before cataract operation. Thoracic space-occupying lesion of uncertain origin]. PMID- 12119909 TI - [When statins alone are not enough. Putting a vice grip on cholesterol]. PMID- 12119910 TI - [Lipid control in the double pack. Combined drugs to achieve the goal]. PMID- 12119911 TI - [Therapy of hypertension. Why the systolic value is decisive]. PMID- 12119912 TI - [Wrinkles, lack of vitality, but also fibromyalgia and arteriosclerosis. A hormone is supposed to eliminate these manifestations]. PMID- 12119913 TI - [Fear of long-term injections. Insulin therapy--soon treatment also without injection]. PMID- 12119914 TI - Nursing practice and the preoccupation with power over death. PMID- 12119915 TI - The spiritual aspects of dying at home. AB - Paterson and Zderad's humanistic nursing theory can be used to meet the spiritual needs of terminally ill persons in the home setting. The spiritual needs, as identified by Highfield and Cason, are applied to the hospice patient. The comforts of the home environment and humanistic nursing practice are integrated in the "meetings" between dying persons, their families, and hospice nurses. These meetings contribute to fulfilling the spiritual needs of terminally ill persons. Hospice nurses practicing holistic nursing and using caring behaviors help dying persons develop a "more-being" in themselves as the triad of person, family, and nurse share the lived experiences. PMID- 12119916 TI - Guilt, shame, and religious and spiritual pain. AB - Hospice care is dedicated to alleviating the pain of dying people. In addition to physical, social, and psychological pain, religious or spiritual pain can add to the struggles of many patients. Religious pain is rooted in guilt leading toward punishment and experienced as fear. It is resourced through the positive teachings of the patient's religious legacy. Spiritual pain is rooted in shame leading a patient to abandon hope in God's love. It is resourced through bringing unconditional love to the patient's sense of self-hatred and inner criticism. PMID- 12119917 TI - Easing elders' pain. AB - Pain is an undertreated, understudied problem in the growing elder population. Clinicians need to consider the compelling evidence that a significant majority of this population experience pain that interferes with quality of life and normal functioning. The barriers to adequate pain management must be addressed and misconceptions corrected. It is critical to identify both acute and chronic pain. Pain assessment and reassessment need to be performed regularly and used in treatment selection. Consideration of physiologic changes that occur in the aged will guide choices of pharmacologic and non-pharmacologic therapies. Pain assessment and treatment must be recognized as fundamental care issues. PMID- 12119918 TI - The importance of culture throughout all of life and beyond. AB - Culture is a critical component of what shapes human experience throughout life. Unfortunately, at the end of life cultural considerations often take a back seat to considerations related to disease processes and functional ability. This article describes why culture is so vital at all stages of life. It also attempts to provide insight into how death, dying, grief, and loss are viewed from a cross cultural perspective with a focus on African Americans. It presents practice implications for health care professionals. PMID- 12119919 TI - A relaxation training program to increase self-efficacy for anxiety control in Alzheimer family caregivers. AB - This study investigated whether relaxation training would increase caregivers' self-efficacy for controlling anxiety associated with difficult behaviors by care recipients. Thirty-six individuals caring for relatives with Alzheimer's disease were taught four types of relaxation techniques. Caregivers' self-efficacy increased following the intervention. Length of time spent in the caregiving role and duration of time since care recipients' diagnosis were significantly related to caregiver self-efficacy before the intervention, but became non-significant following the intervention. An insignificant decrease in incidence of reported behavioral problems occurred following the intervention. PMID- 12119920 TI - Therapeutic music for patients with psychiatric disorders. AB - Many patients with psychiatric disorders struggle with poor skills in coping, communication, socialization, and self-expression that may result in dysfunctional behavioral, cognitive, and emotional responses. Therapeutic music offers a noninvasive approach to strengthen these skills and effect behavior change. At a regional inpatient psychiatric hospital in Colorado, a program of therapeutic music was developed using Rogers' theory of unitary human beings as the theoretical framework. This article describes the approach used to strengthen coping skills in communication, socialization, and self-expression. Suggestions are made for developing a program of music with similar patient populations. PMID- 12119921 TI - Healing practices of the people of Belize. AB - Few countries in the world offer health care enthusiasts the chance to explore holistic models more than the tiny country of Belize in Central America. Situated in the lush rain forests of the tropics, Belize provides the context for practitioners who are trying to blend treatment modalities as old as the Mayan Empire with the unrelenting influx of intensive care technologies from the West. Herein are the stories of a cadre of healers in Belize who model an integration of the traditional and the professional and how they have accomplished it. PMID- 12119922 TI - Blood transfusion: merits of component therapy. I. The clinical use of red cells, platelets, and granulocytes. PMID- 12119923 TI - Is the nipple the preferred landmark for selecting chest electrode positions in infants? Preliminary report. AB - The relation on roentgenograms between the intercostal spaces and routinely selected chest electrode Position 4 on the one hand and the mammary nipple on the other was determined in 15 infants. The nipple was more frequently located over the fifth intercostal space than the electrode position selected by palpation. This difference is ascribed to poor delineation of the angle of Louis in this age group. PMID- 12119924 TI - Bell's palsy in infants associated with varicella-zoster virus infection. PMID- 12119925 TI - Nodular toxic goiter (Plummer's disease) in a child. PMID- 12119926 TI - Breech presentation among infants with familial dysautonomia. PMID- 12119927 TI - Hepatoma associated with androgen therapy for aplastic anemia. PMID- 12119928 TI - Variations in salicylamide glucuronide formation in normal and in G-6-PD deficient children. PMID- 12119929 TI - Fetal blood lead values in a rural area. AB - Lead levels were determined from neonatal samples of blood collected at a general care hospital. The cord blood lead values of babies from urban maternal residences were significantly higher than those from rural environments. The over all cord blood lead value was (mean +/- S.D.) 9.4 +/- 3.7 micrograms of lead per 100 ml. of whole blood. This level represented a considerably lower value than others previously reported and suggests a lower "normal" value for our population. PMID- 12119930 TI - Ophthalmoplegia in maple syrup urine disease. PMID- 12119931 TI - A family with Mobius syndrome. AB - A classic Mobius syndrome, including bilateral abducens-facial paralysis, was found in a newborn boy. At least 15 maternal relatives showed partial features of the syndrome. As potential features of Mobius syndrome also were found in the father's side of the family, a recessive mode of inheritance is suggested. PMID- 12119932 TI - Use of the telephone by pediatric house staff: a technique for pediatric care not taught. AB - Pediatric house staff had difficulty taking an adequate telephone history when given a complaint of cough, vomiting, diarrhea, or rash. Increased time in training did not seem to improve history taking. Those responsible for pediatric training programs need to consider instructing house staff in the use of the telephone. PMID- 12119933 TI - The syndrome of rudimentary testes: occurrence in five siblings. PMID- 12119934 TI - Cyclophosphamide and the prepubertal gonad: a negative report. AB - Serum determinations of follicle-stimulating hormone and luteinizing hormone have been carried out in 13 prepubertal and adult patients who had been treated with courses of either oral or intravenous cyclophosphamide. All results were within the normal range for the patients' ages and sexual development. Although these results establish that gonadal endocrine function and pituitary-gonadal feedback relations may not be destroyed by cyclophosphamide, the possibility remains that prolonged cyclophosphamide therapy in young patients will result in some impairment of future fertility. PMID- 12119935 TI - Incidence of kanamycin resistance among Escherichia coli isolates from neonates. AB - The antimicrobial susceptibilities of 368 E. coli strains isolated from well neonates, well pediatric outpatients, and ill neonates were determined, and the incidence of antibiotic-resistant strains was correlated with epidemiologic data. Ninety-five per cent of these isolates were susceptible to kanamycin and 98 per cent to gentamicin. Most of the difference in susceptibility of these E. coli strains to kanamycin and gentamicin appeared to be on the basis of weight for weight activity. Kanamycin-susceptible strains were isolated from 96 per cent of well neonates and well outpatients, and 94 per cent of ill neonates, age three days or less. An increase in the incidence of resistance to kanamycin, streptomycin, and ampicillin among strains isolated from neonates was related to prolonged hospitalization and/or prior antibiotic therapy. This study demonstrates the need for analyzing antimicrobial susceptibility data from newborn nurseries in an epidemiologic context. PMID- 12119936 TI - Preliminary report of the committee on phototherapy in the newborn infant. PMID- 12119937 TI - Comment: hazard of ultraviolet radiation from fluorescent lamps to infant during phototherapy. PMID- 12119938 TI - Comment: another view of phototherapy. PMID- 12119939 TI - Maternal expectations and attitudes toward toilet training: a comparison between clinic mothers and private practice mothers. AB - Mothers at inner city clinics differ from middle-class suburban mothers with private pediatricians in their attitudes and expectations regarding toilet training, such as the ideal age to initiate bladder and bowel training, ideal age for completion of toilet training, response to the child who soils after a program of toilet training has been initiated, and source of information to guide in toilet training. Possible explanations to account for these differences are explored; the implications for training health personnel are discussed. PMID- 12119940 TI - Pediatric cardiac resuscitation team: a 6 year study. AB - A cardiopulmonary resuscitation team based in the pediatric intensive care unit responded to 239 cardiac arrests over a 6 year period. Patients with respiratory problems and infectious diseases requiring isolation composed the largest groups in whom unexpected arrests occurred. There was a 78 per cent rate of initial response to therapy and a 47 per cent survival rate. This is almost twice the survival rate of the best previously reported study and five times the average. PMID- 12119942 TI - Congenital renal anomalies associated with congenital adrenal hyperplasia. PMID- 12119941 TI - Craniofacial and limb defects secondary to aberrant tissue bands. PMID- 12119943 TI - Birth weight in relation to blood group. PMID- 12119944 TI - "Collagen-like" syndrome with antithyroid therapy. PMID- 12119945 TI - Fatal neonatal postexchange transfusion hepatitis. PMID- 12119946 TI - Arthritis associated with inflammatory bowel disease in children. AB - Arthritis occurred in 23 of 136 (17 per cent) children and teenagers with inflammatory bowel disease, in 18 of 86 (21 per cent) patients with ulcerative colitis, and 5 of 50 (10 per cent) with granulomatous bowel disease. Eighteen children had peripheral arthritis which characteristically affected only a few large joints and was of brief duration and benign outcome. Five boys had spondylitis which was progressive and inseparable clinically from ankylosing spondylitis. Occurrence of joint manifestations was not associated with severity of bowel disease. Anemia and growth retardation occurred frequently. Mucocutaneous lesions were associated with peripheral arthritis but not with spondylitis. No patient had iridocyclitis. The possibility of bowel disease should be considered in children presenting with arthritis, particularly if gastrointestinal complaints, mucocutaneous lesions, anemia, or growth retardation are associated with pauciarticular arthritis. Peripheral arthritis is benign and regresses with improvement of underlying bowel disease but spondylitis is progressive and requires recognition and management for prevention of deformity. PMID- 12119947 TI - Infection and chronic granulomatous disease. PMID- 12119948 TI - Growth standards for American negro (black) boys and girls. PMID- 12119949 TI - Insulin and sodium bicarbonate treatment of diabetic acidosis. PMID- 12119950 TI - Insulin and sodium bicarbonate treatment of diabetic acidosis. PMID- 12119951 TI - Lead in red blood cells and plasma. PMID- 12119952 TI - Comparison of the antibody response to streptococcal cellular and extracellular antigens in acute pharyngitis. AB - The antibody response to the group A carbohydrate moiety of the streptococcal cell wall is of special interest because of its postulated role in the pathogenesis of rheumatic valvulitis. The immune response to this somatic antigen was measured in 159 children with culture-proved group A streptococcal pharyngitis and was compared with that to two extracellular antigens of the Group A streptococcus: streptolysin O and streptococcal DNase B. The data suggest that the maximum anti-A-carbohydrate rise occurs soon after the onset of streptococcal pharyngitis in a fashion similar to the response to some streptococcal extracellular antigens. However, the anti-A-carbohydrate antibody response appeared to be a less sensitive indicator of streptococcal upper respiratory tract infection. PMID- 12119953 TI - Hypocomplementemic and normocomplementemic acute nephritis in children: a comparison with respect to etiology, clinical manifestations, and glomerular morphology. AB - Of 182 patients with acute glomerulonephritis, 20 had normal C3 levels at onset. Normocomplementemic and hypocomplementemic patients were similar with respect to incidence and site of preceding streptococcal infection, elevation of ASO titer, distribution by age, sex, race, season, and year,?and glomerular morphology by light and electron microscopy. They differed in that the normocomplementemic patients tended to have normal serum C5 levels and, for reasons not clear, reduced serum albumin and elevated cholesterol levels. The consistent absence by immunofluorescence of IgG in the glomeruli of five hypocomplementemic patients and its presence in five normocomplementemic patients was considered a chance observation. The data suggest that in each group the nephritis was poststreptococcal and that the mechanism producing poststreptococcal glomerulonephritis is capable of acting independently of that activating circulating C3. PMID- 12119954 TI - Hypertension and narrowing of the renal arteries in infancy. AB - The clinical histories of two infants who had severe hypertension and stenoses of the renal arteries are reported. In both children blood pressure fell significantly after surgical correction of renal ischemia. However, both infants had persistent mild hypertension for several months after surgical treatment. Similar results were found in other infants with renovascular hypertension, reported previously (11 cases). The mortality rate in this total group of 13 infants reviewed was high if renal ischemia was not corrected surgically (4/4). The reason why some infants have persistent hypertension after apparently adequate surgical relief of renal ischemia is not known. PMID- 12119955 TI - Plasma lead levels in normal and lead-intoxicated children. AB - Lead reaches tissues through the plasma, where the concentration of this ion is characteristically low. Little information exists on the mechanisms which determine the dynamics of lead in plasma. Measurement of plasma lead levels have been determined in normal and lead-intoxicated children, newborn infants, and children with sickle-cell disease. The results in all groups were remarkably similar and constant over a wide range of blood lead concentration and regardless of hematocrit. These results lend further support to the postulate that the red cell represents a repository for lead, maintaining plasma lead concentration within closely defined limits, and that methods other than an isolated measurement of plasma lead will be necessary to uncover a presumably dynamic transport system between red cell and plasma. PMID- 12119956 TI - Febrile illness in early infancy associated with ECHO virus infection. AB - Twenty-three infants less than six weeks of age with fever of 100.4 degrees F, or higher and no evidence of bacterial infection were seen at the Cincinnati General Hospital from July to December, 1971. Seventeen of these were admitted to the hospital; 15 were treated with penicillin and kanamycin for possible sepsis. Viral isolation was attempted on 21 of these infants, and ECHO viruses were isolated from 14 (66.7 per cent), compared to three (14.3 per cent) of 21 controls. Eleven of the 14 ECHO viruses isolated were type 4, and the other 3 were types 6, 11, and 25 respectively. Acute and convalescent serum was obtained from 11 of the 21 infants; seven had a fourfold rise in antibody to ECHO virus type 4. Most of the children had fever with irritability, six of the 23 had a fine maculopapular rash, and three had aseptic meningitis. This study suggests that ECHO viruses may be a significant cause of febrile illness in young infants during the summer and fall and may account for illnesses which lead to hospitalization as possible sepsis. PMID- 12119957 TI - Neutrophil granulocyte function in patients with pulmonary infection. AB - Neutrophil granulocyte function was evaluated in patients with pulmonary disease to determine if a change in function occurred during active pulmonary infections. Twenty-five patients with cystic fibrosis of the pancreas were studied. Ten patients were asymptomatic and 15 patients had symptoms and signs of active pulmonary infection. Leukotaxis, random migration, and nitroblue tetrazolium dye reduction by neutrophils were compared in the asymptomatic and symptomatic patients. The leukotactic activity of circulating neutrophils was markedly increased in all of the patients with active pulmonary infection and spontaneous nitroblue tetrazolium dye reduction by neutrophils was increased in 13 of 15 patients. Bacterial infection confined to the respiratory tract is capable of stimulating circulating neutrophils making them more responsive to chemotactic factors as well as increasing nitroblue tetrazolium reduction. PMID- 12119958 TI - Hyperreactivity to cow's milk in an infant with LE and tart cell phenomenon. AB - LE and tart cells were demonstrated in a black male infant whose serum contained milk-percipitating antibodies and who had pulmonary infiltrates. Immunoblasts, plasmocytoid lymphocytes, and an LE cell were found in a milk-stimulated skin window. The presence of LE cells corresponded to the presence of ENA antibody. Tart cells varied with oral milk challenge. A large Arthus type of skin reaction to injected milk was demonstrated. An oral feeding of milk resulted in a decrease in plasma C3. Lymphocyte transformation resulted from in vitro milk stimulation. ENA (extractable nuclear antigen) antibody and resulting LE cell formation possibly represented the combination of nuclear protein with milk antigen. The pulmonary infiltrates may represent a hypersensitivity pneumonitis characterized by both Arthus and cell-mediated reaction to milk. PMID- 12119959 TI - Anorexia nervosa: clinical observations in a successful treatment plan. AB - This poorly understood disease has been studied in 27 girls and two boys. Metabolic aberrations included electrocardiographic abnormalities, hypovitaminosis A, hypercarotenemia, abnormal oral glucose tolerance curves, decreased xylose excretion, decreased excretion of luteinizing (LH) and follicle stimulating hormones (FSH), decreased serum thyroxine levels, absence of diurnal variation of plasma concentrations of glucocorticoids, elevation of blood urea nitrogen, bone marrow hypoplasia, and leukopenia. With restoration of proper nutrition, most of these abnormalities (except FSH and LH) were corrected. An effective method for inpatient treatment is described. There were no deaths during this period of study. PMID- 12119960 TI - Familial congenital absence of adrenal glands; evaluation of glucocorticoid, mineralocorticoid, and estrogen metabolism in the perinatal period. AB - The patient was the fourth of affected male siblings. Cortisol (1.3 micrograms per cent), cortisone (9.6), and corticosterone sulfate (0.1) concentrations were low in cord blood. The larger amount of cortisone may have originated from maternal cortisol. Aldosterone was undetectable in cord blood, indicating lack of fetal secretion or maternofetal transfer. Unexpectedly normal concentrations of 11-deoxycorticosterone (DOC) sulfate in cord serum could represent maternal transfer of DOC, with subsequent fetal sulfurylation. Low estrone and estradiol concentrations in maternal and cord serum were consistent with absence of the fetal adrenals. Despite the low levels of the steroids, the propositus had a normal lecithin-sphingomyelin ratio at 38 weeks' gestation. Circulatory insufficiency developed within half an hour after birth and responded to gluco- and mineralocorticoid therapy. The three untreated siblings died between 14 and 67 hours of age. It is evident that early recognition of this condition may be lifesaving. PMID- 12119961 TI - A familial syndrome of isolated "aplasia" of the anterior pituitary. Diagnostic studies and treatment in the neonatal period. AB - A male newborn infant developed hypoglycemia, collapsed, and convulsed at eight hours of age. The diagnosis of pituitary "aplasia" was suspected, because of a previously affected female sibling, and treatment with glucocorticoids was instituted. Diagnostic studies revealed a deficiency of thyrotropin, growth hormone, and prolactin. He is now six months of age and is thriving on replacement therapy. Analysis of previous reports of this entity indicates that isolated "aplasia" of the anterior pituitary is a genetic syndrome with an autosomal recessive mode of transmission. The course in this patient suggests that this disorder, if diagnosed, is amenable to therapy. PMID- 12119962 TI - Argininosuccinic aciduria: investigation of an affected family. AB - Two siblings with argininosuccinic aciduria were studied by an interdisciplinary team. Considerable variability in the clinical expression of this disorder was observed in comparison with previously reported cases. Detection of the heterozygous state in the parents and in a fetal sibling was demonstrated by tissue assay for argininosuccinase activity. Mental retardation and neurologic deficits in the affected children were irreversible with dietotherapy but growth was progressive and the general course was relatively benign. PMID- 12119963 TI - A pattern of craniofacial and limb defects secondary to aberrant tissue bands. AB - Seven infants have been studied who have a similar pattern of severe craniofacial and limb anomalies associated with aberrant tissue bands. The craniofacial anomalies consist of unusual encephaloceles, facial clefts, and cranial plus midfacial distortion; the limb anomalies consist of constrictions, amputations, and pseudosyndactyly. These defects are interpreted as being secondary to the early distorting and disruptive effects of aberrant tissue bands, the etiology for which is undetermined. Survival beyond the neonatal period has occurred in four of the five liveborn patients, indicating that early death should not necessarily be anticipated. Recurrence risk is apparently negligible. PMID- 12119964 TI - Lymphoepithelioma in childhood. AB - Nine black children with lymphoepithelioma, a rare malignancy of childhood, are the subject of this report. Unique clinical features included tender cervical lymphadenopathy with torticollis, trismus, epistaxis, and change in voice quality. A nasopharyngeal mass was demonstrable in seven children on careful examination, but none was resectable. Treatment with radiation alone or radiation plus cyclophosphamide resulted in complete tumor regression in eight of the nine children. Local recurrence or distant metastases occurred in four within 10 months, two of whom responded to additional radiation plus cyclophosphamide or adriamycin. At present, four children are alive without disease for periods of seven to 78 months, two are alive for seven to 53 months and are in remission from recurrent disease, and three have died with distant metastases. Freedom from disease for one year was associated with a favorable prognosis. Adjuvant chemotherapy appears warranted in view of the high incidence of local recurrence and distant metastases. PMID- 12119965 TI - Re-look at service as an innovation for nursing. AB - Service as a form of scholarship of application is examined from the perspectives of academic and practice experiences of professional nurses. Emphasis is placed on the evolution of services. Data and case studies are provided to illustrate varied ways in which nurses are expertly providing service to the profession and community. The need to explore further the innovative contributions nurses make through scholarly service is stressed. PMID- 12119966 TI - The beat of the drum. AB - This article describes the development of a nurse-managed clinic in an urban environment. The clinic provides primary care services to persons who do not have health insurance. One primary population served is urban American Indians. Perceived barriers to care are discussed. Cultural sensitivity is addressed and patient situations are presented. PMID- 12119967 TI - Care of homeless men in the community. AB - The author describes health care provided to homeless men by community health nursing students who conduct a service learning project at an inner city mission. Service learning is an educational method that combines the academic learning objectives of community nursing students and the health care educational and service needs of a vulnerable population of homeless men. Holistic and interdisciplinary nursing skills are practiced. Clients learn healthier life style choices and to determine more appropriately the need and source for professional health care when problems are identified. PMID- 12119968 TI - Faculty practice at a homeless shelter for women and children. AB - Homelessness in America has significantly increased in recent years. Exact numbers of homeless persons in the United States are difficult to assess, though estimates of homeless persons range from 250,000 to 3 million. The homeless population has shifted to include women and children, including two parent families. Providing health care for the homeless is one of the most important and challenging health issues today. There are many barriers to providing adequate health care. The purpose of this article is to describe the complexity of the role and the experiences of a pediatric nurse practitioner at a clinic in a homeless shelter that houses approximately 30 women and children. PMID- 12119969 TI - Faculty practice and health promotion in a community correctional facility. AB - Faculty practice and health promotion are topics of interest to nursing faculty to prepare students and themselves for the future. Health promotion with vulnerable populations is a focus for the 21st century health care delivery system. Faculty practice emphasizing health promotion with a vulnerable population meets the needs of faculty as they strive to meet personal and professional goals. The development of a faculty practice serving a vulnerable population represented by clients in a community corrections facility will be described. Scope of practice, the development of a research focus, teaching approaches, and future initiatives are presented. PMID- 12119970 TI - Massage therapy as a nursing practice. AB - Nursing is a profession that can be practiced in many unique ways. Nursing care can be provided through conventional means in traditional settings or practiced in domains labeled as alternative or complementary. This article describes massotherapy as an alternative therapy of a holistic nursing practice. The elements of a business plan, including planning and marketing are addressed. PMID- 12119971 TI - Moving towards health oriented patient education (HOPE). AB - The economics of prevention supports reimbursement of nurse practitioners for patient education. The role has undergone historical change, shifting from imparting disease-oriented health education (DOPE) toward empowering patients to use their own resources to the fullest to attain health. Nurse practitioners are well suited to provide care that facilitates behavior change and health-oriented patient education (HOPE). Essentials for effective patient education include use of an open communication style, written instructions, and the address of barriers. Adult literacy and reader-friendliness must be considered when assembling written materials. PMID- 12119972 TI - Toward holistic theory-based intervention for dementia behavior. AB - Four million Americans suffer from some form of dementia. Over 50% of these individuals exhibit behaviors that are perceived as "disturbing" by family and caregivers. The Need-driven Dementia-compromised Behavior Model was developed by a group of nurse researchers to study and understand these behaviors. The model changes the view of dementia behaviors as "disturbing" to that of behavior as signifying potentially understandable needs. This article reviews the model and demonstrates how interventions derived from the model can be used to respond to dementia behaviors in a holistic fashion. PMID- 12119973 TI - Social support and strain of family caregivers of older adults. AB - To assess the direct effects and interactive models of social support, caregivers to functionally impaired older adults were identified by hospital personnel. Within a week of referral, family caregivers were interviewed in the home about strain, depressive symptomatology, caregiving appraisal, informal social support, and coping. Caregiving appraisal significantly explained strain and depressive symptomatology. The interaction of social support with strain did not moderate or lessen depressive symptomatology. These findings suggest that nursing continue to examine the effect of home health care on strain and depressive symptomatology of caregivers of older adults. PMID- 12119975 TI - Businesspeak is the new language of healthcare. PMID- 12119974 TI - Using cognitive strategies to enhance bladder control and comfort. AB - Compromised urinary bladder syndrome (CUBS), a combination of frequency and incontinence, causes multiple discomforts for community-dwelling adults. A holistic intervention--audiotaped cognitive strategies--was designed to augment the effects of an educational program designed to treat CUBS. CUBS was operationalized with a voiding diary, and comfort related to bladder health was operationalized in a questionnaire. In this quasi-experimental design the outcomes were measured at four time points. Repeated measures multivariate analyses of variance and nonparametric analyses were conducted to assess differences between the two groups. Results indicated that the treatment group had more comfort and improved CUBS compared with the control group. PMID- 12119976 TI - Creating violence-free healthy cities for our youth. AB - Communities are seeking ways to improve health by reducing violence. The Social Ecology Model of Adolescent Interpersonal Violence Prevention is presented as a tool for communities to use in identifying and addressing the multiple factors impacting engagement in or avoidance of violent behaviors. The CityNet Healthy Cities model is presented as a process for communities to use in developing broad based actions groups capable of having an impact on violence. Together these models can be used by health professionals as they work with communities. Suggestions for community assessment, evaluation of available programs, committee development, and implementation are provided using the two models. PMID- 12119977 TI - Using peer volunteers to promote mammography education in senior citizens' housing facilities. AB - A peer volunteer intervention to encourage attendance at mammography education programs was developed and evaluated as part of a multi-intervention study to improve mammography by women 65 years and older. Approximately five to seven women from each of 20 racially and socioeconomically diverse senior citizens' housing facilities were recruited. After completing a training session, they distributed invitations and spoke personally with fellow residents about attending the upcoming program. Attendance rates for facilities with the peer volunteer program were compared with those in facilities without the volunteers. Results of a logistic regression analysis found the volunteer program, in addition to facility size, to have had a statistically significant effect on attendance at the mammography education presentations. PMID- 12119978 TI - The grief of parents of murdered children: a suggested model for intervention. AB - The grief of parents who have lost children to murder is extreme, prolonged, and unique. The authors, themselves bereaved by homicide, suggest a model to facilitate understanding such grief and to guide intervention. The model postulates that homicide survivor grief occurs in three arenas: the private, personal world; the public world of the media; and the world of the criminal justice system. Each arena contains features unique to homicide survivor bereavement. These features are described for the reader. PMID- 12119979 TI - Asthma in the inner city: a growing public health problem. AB - Asthma in the inner city impacts people of all ages and is most pronounced in African Americans and other minorities. During the past decade, the prevalence of asthma has increased by 42%, a rate consistently higher in African Americans. Along with the increase in asthma prevalence, the costs associated with this disease have also risen dramatically. In addition, asthma is the leading cause of school absenteeism and also contributes to lost productivity. This article focuses on the epidemiology of asthma in urban areas and identifies various risk factors that are important in achieving control of this disease. Suggestions for future interventions are discussed. PMID- 12119981 TI - Parent to parent: preventing adolescent exposure to HIV. AB - A 2-year pilot program implemented by public health nurses (PHNs) in partnership with the leaders of parent associations of 10 urban high schools encouraged communication about health-promoting sexual behavior between adults and adolescents. The purpose was to assess the feasibility of involving parents in health promotion efforts in schools. During the first year, PHNs prepared a cohort of 17 parent trainers to make presentations to parent groups about the reality of teens' risk for human immunodeficiency virus (HIV) exposure, facts to counter myths about outcomes of sex education, and the skills teens need to implement healthy choices, including abstinence, correct condom use, and talking about sex with a potential sexual partner. Each parent trainer was then scheduled to make two presentations at parent association meetings. Criteria for assessing the impact of extending the program, including broader diffusion in the community, are proposed. This approach may be effective for community-based education about other health issues. PMID- 12119982 TI - Learning in the city: what is the value added? AB - As health care services shift to the ambulatory and community setting, health professions educators must consider revamping educational programs to reflect the new venues of care. This transformation extends far beyond the location of services. The dynamic between the health care provider and the patient shifts to provider and customer. Services should be geared to the concerns of the individual and tailored to meet the realities of the person's life. This is true in all settings; and particularly true in the city. This article discusses the unique challenges of the urban environment and the benefits students gain by having clinical experiences based the city and working with members of these communities. PMID- 12119983 TI - Routine encounters of nurses practicing in the urban trenches of health care. PMID- 12119980 TI - A community-developed, community-based lead poisoning prevention program: Lead Awareness North Philly Style. AB - This work describes a demonstration/research project to implement and evaluate community-developed, community-based strategies that address childhood lead poisoning. The project will increase knowledge of childhood lead poisoning as an environmental health risk, as well as hazard, exposure, and outcome surveillance for lead as an environmental agent. At the end of the second year of this study the data indicate increased knowledge about lead poisoning and that more children are being tested for lead poisoning in the experimental census tracts (CTs), when compared with the control CTs, with lower lead levels. PMID- 12119984 TI - Long-term efficacy of dietary treatment of obesity: a systematic review of studies published between 1931 and 1999. AB - METHODS: MEDLINE surveys were carried out and reference lists were cross-checked to identify publications on long-term outcome for dietary treatment of obesity. 898 papers were identified, 17 fulfilled our planned criteria for inclusion (dietary treatment; adults; follow-up period > or = 3 years; follow-up rate > or = 50% of original study group; information on one of the success criteria: maintenance of all weight initially lost (or further weight reduction) or maintenance of at least nine to 11 kg of initial weight loss; obesity complications of the patient group not over-represented; English, German or Scandinavian languages). RESULTS: The 17 included publications (here of three publications on randomized clinical trials with control group relevant for this review) reported on 21 study groups, comprising 3030 patients. Of these 2131 (70%) were followed-up for 3-14 years (median 5 years). Mean initial weight loss ranged from four to 28 kg (median 11 kg). Overall, 15% (median, range 0-49%) of followed-up patients fulfilled one of the criteria for success. Overall, success rates seemed stable for up to 14 years of observation. Diet combined with group therapy lead to better long-term success rates (median 27%) than did diet alone (median 15%) or diet combined with behaviour modification (median 14%). Active follow-up was generally associated with better success rates than was passive follow-up (19% vs. 10%). Conventional diet seemed to be most efficacious in addition with group therapy, whereas VLCD apparently was most efficacious if combined with behaviour modification and active follow-up. CONCLUSION: The literature on long-term follow-up of dietary treatment of obesity, although limited and inhomogeneous, points to an overall median success rate of 15% and a possible adjuvant effect of group therapy, behaviour modification and active follow-up. PMID- 12119985 TI - Orlistat for the treatment of obesity: rapid review and cost-effectiveness model. AB - The aim of this study is to clarify the potential benefits, disbenefits and costs of Orlistat for the treatment of obesity. The method was a search for relevant systematic reviews and randomized controlled trials, in Medline, Pre-Medline, Embase and the Cochrane Library, using Orlistat and its synonyms. Identified trials were appraised using a standard appraisal checklist and trial data were extracted for use in cost-effectiveness modelling. Three large multicentre, randomized placebo controlled trials were included in the rapid review. On average, Orlistat results in obese people losing an additional 3-4% of their initial body weight over diet alone during a 2 year period. There was no strong evidence that this short-term weight loss would have a longer-term impact on morbidity and mortality. The cost utility of Orlistat treatment was estimated at around 46,000 Pounds per Quality Adjusted Life Year gained (extreme values sensitivity analysis 14,000 Pounds to 132,000 Pounds). This rapid review raises some important questions about the potential value of Orlistat in the treatment of obesity. Further research is needed, not only to clarify the longer-term impact of Orlistat treatment, but also to uncover the longer-term impact on mortality and morbidity from short-term weight loss. PMID- 12119986 TI - An assessment of the safety and efficacy of sibutramine, an anti-obesity drug with a novel mechanism of action. AB - Sibutramine is a combined serotonin(5-HT) and noradrenaline (NA)re-uptake inhibitor. Sibutramine works predominantly through its two pharmacologically active metabolites (i.e. primary and secondary amines) which induce marked weight loss by affecting both food intake and energy expenditure. It is able to enhance the physiological process of satiety, and to stimulate thermogenesis, increasing the efferent sympathetic activity to thermogenically active brown fat. There is a dose-related reduction in body weight in clinical trials with sibutramine, with weight loss up to 11% below baseline, which can last up to 18 months with continued treatment. When weight loss is induced with a very low calorie diet (VLCDL), patients randomized to the sibutramine treatment continued to lose weight over a 1 year period, reaching 15% below baseline, whereas the placebo treated patients regained some weight. Sibutramine improves metabolic fitness, by decreasing the biochemical risk factors associated with obesity, such as plasma triglycerides, total cholesterol and low density lipoprotein (LDL) cholesterol, glucose and insulin, and increasing HDL-cholesterol. In controlled studies, 84% of sibutramine-treated patients reported side effects, most commonly including dry mouth, constipation and insomnia, compared with 71% of patients receiving placebo. A small increase in heart rate and blood pressure also occurs and persists for as long as treatment is continued, which, therefore, requires monitoring. Nevertheless, successful treatment of moderately hypertensive obese patients with sibutramine has been demonstrated without undue blood pressure problems and even a mean lowering of blood pressure associated with weight loss. Finally, sibutramine does not have the potential for abuse that is characteristic of amphetamine and it is indistinguishable from placebo in abuse potential studies. PMID- 12119987 TI - Redefining type 2 diabetes: 'diabesity' or 'obesity dependent diabetes mellitus'? AB - Type 2 diabetes is considered by diabetes physicians as a complex and heterogeneous disease with a poorly understood aetiology, apart from the fact that there is a strong genetic propensity that becomes overt when exposed to a typical Western lifestyle. Our clinical targets are now moving from controlling the disease to preventing it. Do we need to await more research on the aetiology and pathophysiology before establishing a preventive strategy? No, the pathophysiology may be poorly understood, but there is now solid evidence that type 2 diabetes is a disease of fatness. New, controlled, clinical trials show that as little as 5% weight loss is sufficient to prevent most obese subjects with impaired glucose tolerance developing type 2 diabetes. Since type 2 diabetes is obesity dependent, and obesity is the main aetiogical cause of type 2 diabetes, we propose the term 'diabesity' should be adopted. PMID- 12119988 TI - The human uncoupling protein-1 gene (UCP1): present status and perspectives in obesity research. AB - Energy expenditure through brown adipose tissue thermogenesis contributes either to maintenance of body temperature in a cold environment or to wasted food energy, i.e. cold-induced or diet-induced thermogenesis. Both mechanisms are due to a specific and unique protein: the uncoupling protein-1. Uncoupling protein-1 is exclusively expressed in mitochondria of brown adipocytes where it uncouples respiration from ATP synthesis, dissipating the proton gradient as heat. In humans, although uncoupling protein-1 can be detected, the inability to quantify brown adipose tissue makes it difficult to argue for a role for uncoupling protein-1 in thermogenesis and energy expenditure. This review summarizes data supporting the existence of brown adipocytes and the role of UCP1 in energy dissipation in adult humans. Understanding the mechanisms which regulate transcription and expression of the human UCP1 gene will facilitate the identification of molecules able to increase the levels of this protein in order to modulate energy expenditure in adult humans. PMID- 12119989 TI - The mechanism of effect of growth hormone on preadipocyte and adipocyte function. AB - Growth hormone (GH) is not only the major regulator of postnatal somatic growth but also exerts profound effects on body composition through a combination of anabolic, lipolytic and antinatriuretic actions. GH enhancement of the lipolytic activity of adipose tissue in combination with a reduction of triglyceride accumulation via inhibition of lipoprotein lipase activity appears to be the major mechanism by which GH results in a reduction of the total fat mass. Recently, much progress has been made in understanding the molecular mechanism by which GH affects cellular function. This review provides a brief discourse and summary of the mechanism of effects of GH on preadipocyte/adipocyte function. It is intended to provide a functional understanding of the mechanism of action of GH as it relates to adipogenesis and adipocyte function. PMID- 12119990 TI - Biological determinants of spontaneous physical activity. AB - A decline in daily physical activity levels is clearly a major factor contributing to the current obesity epidemic affecting both developed and developing countries in the world. This escalating problem is associated with increased morbidity and mortality and reduced psychosocial health. Thus, increasing physical activity has become the strategy of choice in public health strategies to prevent obesity. Efforts to improve levels of physical activity in the population rely upon an accurate understanding of the determinants of physical activity. Most research has focused on environmental and social influences, while the potential for physical activity to be controlled by intrinsic biological processes has been largely overlooked. This review presents some of the compelling and diverse evidence that has emerged recently showing that physical activity energy expenditure is a critical factor in both the successful regulation of energy balance in normal individuals, as well as the abnormal regulation of energy balance that characterizes obesity. Although the metabolic and genetic factors involved in these regulatory processes remain mostly unidentified, some novel discoveries have been made in this area recently and these are described within this review. PMID- 12119991 TI - Does physical activity prevent weight gain--a systematic review. AB - This paper is a systematic review of research data on associations between physical activity and weight gain, with or without prior weight reduction. The selected studies were restricted to Caucasian (white) adults. Most studies with data on physical activity collected at follow-up, found an inverse association between physical activity and long-term weight gain. This finding was present in studies both with and without prior weight reduction. Prospective studies with physical activity measured at baseline, and randomized weight reduction interventions, gave inconsistent results regarding the effects of increased physical activity on weight change. The weighted mean weight regain in randomized studies with or without exercise training was 0.28 and 0.33 kg/month, respectively. Based on observational studies, it seemed that an actual increase in energy expenditure of physical activity of approximately 6300-8400 kJ/week (1500-2000 kcal/week) is associated with improved weight maintenance. This is more than was prescribed in most randomized trials, and certainly more than the participants actually achieved. Adherence to a prescribed exercise programme remains a big challenge. Before new methods to improve exercise adherence are found, the role of prescribed physical activity in prevention of weight gain remains modest. PMID- 12119992 TI - Taste, food intake and obesity. AB - Research in human eating behaviour prior to 1990 has shown that taste impacts the palatability and selection of food for intake; sensory-specific satiety; satiation; and thermic effect of food. Research in the last decade has added information to these areas; expanded the field to comparisons of the impact of 'wanting' vs. 'liking' food on intake, and provided insight into the relationship of food intake and brain functioning through new imaging techniques. This article will review literature from the last decade on research in the area of taste and its impact on food intake. Emphasis will be placed on differences seen between lean and obese humans and how these may contribute to the development of human obesity. Suggestions for future research directions will also be discussed. PMID- 12119993 TI - Quality of life and obesity. AB - Interest in the quality of life of patients with different diseases continues to grow. Recent years have witnessed a dramatic rise in the prevalence of obesity worldwide, stimulating interest in the health and quality of life consequences of this phenomenon. The body of research on the quality of life of obese individuals has grown to a point that a review of this literature is warranted. Numerous studies have demonstrated that obese persons experience significant impairments in quality of life as a result of their obesity, with greater impairments associated with greater degrees of obesity. Weight loss has been shown to improve quality of life in obese persons undergoing a variety of treatments. Further research is needed to clarify whether quality of life differs among subsets of obese persons. Until recently, there has been little standardization of quality of life measures in obesity. The SF-36 has been used in a number of studies of obese persons. Several obesity-specific instruments have also been developed and have shown great promise. The quality of life of obese individuals is an important issue that should be included in weight management treatment and research. PMID- 12119994 TI - The influence of obesity on hyperandrogenism and infertility in the female. AB - Although a critical mass of adipose tissue is essential for the normal development of female reproductive function, obesity has been shown to produce menstrual disturbances and subfertility. The severity of obesity and the distribution of fat tissue are important factors that influence the female reproductive system. The pathogenetic mechanistic links between them aren't clearly elucidated. Obese women, especially those with upper body obesity, have insulin resistance and hyperinsulinaemia, hyperandrogenaemia, increased peripheral aromatization of androgens to oestrogens, altered gonadotrophin secretion, decreased sex hormone binding globulin, decreased growth hormone (GH) and insulin like growth factor binding proteins (IGFBPs), increased leptin levels and altered neuroregulation of the hypothalamic-pituitary-gonadal axis. These have been considered as some of the links in the sequence of events of the disrupted ovulatory process. The mechanisms of these actions and their influence on female reproductive function are discussed below. PMID- 12119996 TI - Mitochondrial uncoupling as a target for drug development for the treatment of obesity. AB - Mitochondrial proton cycling is responsible for a significant proportion of basal or standard metabolic rate, so further uncoupling of mitochondria may be a good way to increase energy expenditure and represents a good pharmacological target for the treatment of obesity. Uncoupling by 2,4-dinitrophenol has been used in this way in the past with notable success, and some of the effects of thyroid hormone treatment to induce weight loss may also be due to uncoupling. Diet can alter the pattern of phospholipid fatty acyl groups in the mitochondrial membrane, and this may be a route to uncoupling in vivo. Energy expenditure can be increased by stimulating the activity of uncoupling protein 1 (UCP1) in brown adipocytes either directly or through beta 3-adrenoceptor agonists. UCP2 in a number of tissues, UCP3 in skeletal muscle and the adenine nucleotide translocase have also been proposed as possible drug targets. Specific uncoupling of muscle or brown adipocyte mitochondria remains an attractive target for the development of antiobesity drugs. PMID- 12119995 TI - The biology of white adipocyte proliferation. AB - Expanded adipose tissue mass increases the risk for many clinical conditions including diabetes, hypertension, coronary atherosclerotic heart disease, and some forms of cancer. Therefore, it is imperative that we understand the mechanisms by which fat pads expand. The enlargement of fat cells during the development of obesity has been previously hypothesized to be a triggering factor for the proliferation of new fat cells. There is now a preponderance of evidence that adipose tissue is a source of growth factors such as IGF-I, IGF binding proteins, TNF alpha, angiotensin II, and MCSF that are capable of stimulating proliferation. The relative importance of these autocrine/paracrine factors in the normal control of preadipocyte proliferation is unknown. In addition, the proliferative response of preadipocytes to the paracrine milieu is undoubtedly modulated by neural inputs to fat tissue and/or serum factors. Together, these multiple regulatory controls orchestrate overall and region-specific adipose tissue cellularity responses associated with the development of hyperplastic obesity. Both in vivo and in vitro studies are needed to understand the complex, interacting physiological mechanisms by which growth of this important organ is regulated. PMID- 12119997 TI - Sleep disordered breathing--a new component of syndrome x? AB - Sleep disordered breathing (SDB) is a complication of obesity estimated to occur in about 4-6% of overweight individuals. These respiratory disturbances during sleep incorporate a number of conditions including snoring, upper airway resistance syndrome and obstructive sleep apnoea syndrome (OSAS). It is thought that as well as having deleterious effects on sleep quality these conditions may also promote cardiovascular and hormonal changes leading to an elevated blood pressure and an increased incidence of cardiovascular morbidity. Evidence reviewed here points to an alteration in sympathovagal balance, baroreceptor sensitivity, insulin resistance and leptin, growth hormone and lipid levels. Whether these changes are a consequence of the associated obesity or the SDB itself remains to be proven. PMID- 12119998 TI - Use of beta-blockers in obesity hypertension: potential role of weight gain. AB - Beta-blockers are the most frequently used drugs for the treatment of hypertension. Apart from concerns regarding potential adverse metabolic effects on lipids or insulin sensitivity, beta-blockers can also cause weight gain in some patients. This fact appears little known to clinical practitioners and trialists. Thus, only a minority of clinical trials with beta-blockers report weight changes during treatment. In trials that do report weight changes, beta blockers are associated with a weight gain of 1.2 (range -0.4-3.5) kg. This may be attributable to the fact that beta blockade can decrease metabolic rate by 10%. Beta-blockers may also have other negative effects on energy metabolism. Obesity management in overweight hypertensive patients may therefore be more difficult in the presence of beta-blocker treatment. We therefore question the use of beta-blockers as first-line therapy for overweight or obese patients with uncomplicated hypertension. PMID- 12119999 TI - Digestive system involvement in cystic fibrosis. AB - Cystic fibrosis is a hereditary disease well known to paediatricians. Over recent years, its prevalence among the adult population has dramatically increased; thus becoming a disease increasingly seen in adult practice. Cystic fibrosis is a multi-organ disease, with a wide spectrum of clinical manifestations involving many organs. The aim of this article is to review the digestive system manifestations of this disease: the involvement of the gastrointestinal tract, liver, biliary system and pancreas, with a special emphasis on the adult population. PMID- 12120000 TI - Frequent deletions of tumor suppressor genes in pure pancreatic juice from patients with tumoral or nontumoral pancreatic diseases. AB - BACKGROUND/AIMS: K-ras codon 12 mutation is the most frequent genetic alteration in pancreatic cancer. Sensitivity and specificity of K-ras are not high enough to detect all pancreatic cancers, especially at early stage. This study investigated whether detection of p16 and/or DPC4 deletions along with K-ras mutation in DNA samples could improve the definition of patients at risk of pancreatic cancer. METHODS: K-ras mutations were investigated by sequencing. p16 and DPC4 homozygous deletions were studied using comparative multiplex polymerase chain reaction of DNA in pancreatic juice sampled during endoscopic retrograde pancreatography in 57 patients with either pancreatic cancer (group I, 18 patients), chronic pancreatitis (group II, 20 patients), or nontumoral pancreatobiliary disease (group III, 19 patients). RESULTS: The frequencies of Ki-ras mutations were 61% in group I, 10% in group II, and 10.5% in group III. The frequencies of p16 exon 2 and DPC4 deletions were, respectively, 28 and 36% in group I, 50 and 58% in group II, and 24 and 36% in group III. CONCLUSIONS: The combination of p16 and DPC4 deletions with K-ras mutation does not improve the diagnosis of pancreatic cancer based on K-ras mutation alone. These data suggest that tumor suppressor gene inactivation can occur with a high frequency during nonmalignant pancreatic diseases. PMID- 12120001 TI - Juicy genes. Molecular analysis of pancreatic secretions. PMID- 12120002 TI - Endoscopic ultrasound-guided gallbladder bile aspiration in idiopathic pancreatitis carries a significant risk of bile peritonitis. AB - BACKGROUND: Direct microscopic examination of bile for the presence of microlithiasis is often performed during the evaluation of patients with idiopathic pancreatitis. Bile sampled from the duodenum and/or the common bile duct may not represent gallbladder bile, and thus may be inadequate for the diagnosis of microlithiasis. AIM: We sought to determine the safety and efficacy of endoscopic ultrasound (EUS)-guided fine-needle aspiration (FNA) of gallbladder bile in patients with idiopathic pancreatitis. METHODS: Patients with idiopathic pancreatitis underwent EUS with a linear echoendoscope. After excluding potential causes of pancreatitis such as common bile duct stones or pancreatic lesions, the gallbladder was identified. The gallbladder lumen was entered using a 22-gauge FNA needle via the duodenal wall. Bile was aspirated and analyzed for the presence of cholesterol monohydrate crystal, calcium bilirubinate granules, calcium carbonate microspheroliths and mucin gel strands. RESULTS: Three patients underwent EUS-guided FNA of gallbladder bile. Two of these patients developed bile peritonitis within one hour of the procedure prompting us to discontinue the study. One patient's gallbladder bile contained microlithiasis. This patient had bile aspirated from the common bile duct via ERCP 3 weeks prior to EUS. However, analysis of that bile sample failed to show microlithiasis. CONCLUSION: Unfortunately, transduodenal EUS-guided FNA of gallbladder bile using a 22-gauge needle carries a significant risk of bile peritonitis. PMID- 12120003 TI - Chronic pancreatitis and diabetes mellitus. A retrospective analysis of 156 ERCP investigations in patients with insulin-dependent and non-insulin-dependent diabetes mellitus. AB - BACKGROUND: Pancreatic exocrine dysfunction has been described frequently in IDDM and NIDDM patients. Most authors tried to explain this finding as a diabetic complication. On the other hand, diabetes secondary to chronic pancreatitis (CP) might be more common than believed so far. AIM OF THE STUDY: In this study we evaluated pancreatograms of patients with known diabetes mellitus in order to detect ductal morphology changes characteristic for CP. METHODS: Consecutive diabetic patients admitted for ERCP for different reasons were evaluated retrospectively concerning ERCP findings, especially pancreatic duct changes (Cambridge classification), diabetes type, duration and therapy. RESULTS: 156 patients (76 male, 80 female; mean age 60 years (19-93)) were studied (38 IDDM; 118 NIDDM). Pancreatic ducts were classified as normal in 23.3%, CP degree I in 22.7%, CP degree II in 32.7% and CP degree III in 21.3%. The duct changes did not correlate with diabetes type (p = 0.19), diabetes duration (p = 0.38), diabetes therapy (p = 0.5) or age (p = 0.48). CONCLUSION: Since CP should be defined by morphological and functional changes, it must be concluded that a substantial number of patients with a primary diagnosis of diabetes mellitus may have CP as a concomitant disease or, more likely, as a cause for their diabetic state. PMID- 12120004 TI - Effect of chronic hypoxia on glutathione status and membrane integrity in the pancreas. AB - BACKGROUND: Our recent study has shown that chronic hypoxia could upregulate significantly a local renin-angiotensin system in the pancreas. The activation of such a local renin-angiotensin system may provide an alternate mechanism that leads to the generation of reactive radical species in the pancreas during chronically hypoxic exposure. The present study aims at elucidating the antioxidant status in the pancreas during varying degrees of chronic hypoxia. METHODS: Sprague-Dawley rats were exposed to an isobaric hypoxic (10% oxygen) chamber for a period up to 28 days. The glutathione status and membrane integrity of the pancreas were studied with a time course of chronic hypoxia (3, 7, 14, 21 and 28 days). The effect of chronic hypoxia on changes of oxidative states in the pancreas was assessed based on the measurements of glutathione, malondialdehyde, alpha-amylase and DNA fragmentation using biochemical assays. RESULTS: Pancreatic glutathione was decreased drastically after 3-day hypoxia and its level was almost completely recovered after 7-day hypoxia. Malondialdehyde was not affected while DNA fragmentation was increased significantly in a time-dependent manner during the course of chronic hypoxia. Membrane integrity of the pancreatic cells was improved, as evidenced by the decrease of plasma alpha-amylase during the time-course study of chronic hypoxia. CONCLUSION: Pancreatic glutathione was depleted only in the early period of chronic hypoxia followed by a rapid recovery, suggesting that adaptive response of the pancreas may occur during chronic hypoxia. The enhancement of glutathione-dependent antioxidant capacity during chronic hypoxia prevented oxidative damage to the membrane of the pancreatic cells. PMID- 12120005 TI - Wirsung and Santorini: the men behind the ducts. AB - During the 16th and 17th centuries, several important discoveries were accomplished by anatomists whose contribution has enlightened the most important anatomic structures of the pancreas. Following the earliest discoveries, researchers of several medical specialities further investigated the ductal pancreatic system. The accessory pancreatic duct with its minor papilla, the main pancreatic duct and the papilla major along with the confluence of the main pancreatic duct with the bile duct and pancreas divisum, have been the objects of interest of several personalities of the medical history. Eponyms in pancreatic anatomy were given to remember some of them, although anatomical misattributions are frequent and controversial. The aim of the authors was to dedicate a small tribute to the researchers who have written, during the last 500 years, important chapters of the medical history and who dedicated their lives to study the pancreatic ducts and their duodenal endings. Furthermore, a brief outlook was dedicated to the impact of anatomic variations and of embryologic anomalies of the pancreatic ducts in our clinical practice and in our actual understanding of duct-related diseases. The authors are confident that the genial curiosity of few extraordinary personalities of the past and the opportunities provided by modern technology continue to play a major role that may finally add wisdom to decision making in dealing with duct-related biliopancreatic diseases and safety to diagnostic and therapeutic procedures employed. PMID- 12120006 TI - Treatment with 5-fluorouracil enhances radiosensitivity of the human pancreatic cancer cell line MiaPaCa-2. AB - BACKGROUND AND PURPOSE: Several clinical studies have suggested that the combination of radiation therapy and 5-fluorouracil (5-FU) may improve outcome of patients with pancreatic cancer. However, there are few experimental studies supporting this treatment. AIM OF THE STUDY: To examine the radiosensitivity of human pancreatic cancer cells and its modulation by 5-FU. MATERIAL AND METHODS: MiaPaCa-2, PANC-1 and NP-18 cells growing as monolayer culture were treated with radiation and 5-FU. In addition, 5-FU was studied administered either pre- or postradiation, both as pulse or continuous exposure. Cell survival was determined by the in vitro clonogenic assay. RESULTS: In MiaPaCa-2 cell line, both radiation and 5-FU alone reduced cell survival. The addition of 5-FU to radiation caused a significant net decrease of cell survival. Pulse exposure of 5-FU decreased survival after 2 Gy and mean inactivation dose by 1.64; continuous exposure decreased survival after 2 Gy and mean inactivation dose by about 2.4. Timing of 5-FU exposure did not modify survival. However, when adjusting for 5-FU killing effect and cell multiplicity, only continuous exposure significantly enhanced radiation cell killing. CONCLUSION: Both pulse and continuous exposure increase radiation cell killing, but only continuous exposure may radiosensitize MiaPaCa-2 cells. PMID- 12120007 TI - Differential activation of p42ERK2 and p125FAK by cholecystokinin and bombesin in the secretion and proliferation of the pancreatic amphicrine cell line AR42J. AB - BACKGROUND: AR42J rat pancreatic acinar carcinoma cells have retained the potential to secrete digestive enzymes in addition to their ability to proliferate upon stimulation with regulatory peptides. We investigated the involvement of p42ERK2 and p125FAK (extracellular signal-regulated protein kinase and focal adhesion protein kinase, respectively) by cholecystokinin and bombesin stimulation with regard to secretion and mitogenesis. METHODS: The p42ERK2 activity was measured by kinase assay and the activation of p125FAK by antiphosphotyrosine Western blot analysis of p125FAK immunoprecipitates. The expression of both kinases was determined by Western blot analysis, the amylase secretion by colorimetry, and the DNA synthesis by [3H]thymidine incorporation. RESULTS: p42ERK2 and p125FAK were activated by cholecystokinin and bombesin with maximum stimulation at concentrations above 10 nM. Bombesin was a weaker activator of p42ERK2 and p125FAK, causing only half of the kinase activity induced by stimulation with cholecystokinin. PD98059 was shown to inhibit p42ERK2, while tyrphostin 25 blocked p125FAK tyrosine phosphorylation. Preincubation of AR42J cells with PD98059 or tyrphostin 25 was without influence on cholecystokinin- or bombesin-stimulated secretion in normal or 72-hour dexamethasone-pretreated cells. In contrast, inhibition of both protein kinases leads to reduced cholecystokinin-stimulated [3H]thymidine incorporation rates. CONCLUSIONS: Cholecystokinin induced proliferation of AR42J cells by strong activation of p42ERK2 and p125FAK. Bombesin failed to stimulate DNA synthesis, probably due to its reduced potency to stimulate these kinases. Both protein kinases are not implicated in the process of enzyme secretion. PMID- 12120008 TI - Effects of Celebrex and Zyflo on BOP-induced pancreatic cancer in Syrian hamsters. AB - BACKGROUND/AIMS: Selective inhibition of eicosanoid synthesis decreases inflammation, however, it is still unknown whether oxidative stress and carcinogenesis might be influenced in ductal pancreatic ductal cancer as well. METHODS: 120 male hamsters were randomized into 8 groups (n = 15). While control group 1-4 received 0.5 ml normal saline s.c. weekly for 16 weeks, groups 5-8 were injected 10 mg BOP/kg body weight to induce pancreatic cancer. After establishment of pancreatic cancer, groups 1 and 5 received no therapy, groups 2 and 6 were fed 7 mg Celebrex daily, groups 3 and 7 were given 28 mg Zyflo and groups 4 and 8 received Celebrex and Zyflo orally daily in weeks 17-32. In week 33, all animals were sacrificed, macroscopic size of pancreatic carcinomas was measured, incidence of pancreatic cancer was analyzed histopathologically and activities of antioxidative enzymes and concentration of products of lipid peroxidation in tumor-free and pancreatic intratumoral tissue were determined. RESULTS: Incidence and size of macroscopic pancreatic carcinomas were decreased by single therapy with Zyflo as well as combined therapy (Zyflo + Celebrex). Activities of antioxidative enzymes were increased and the concentration of products of lipid peroxidation was decreased in tumor-free pancreas. On the other hand, lipid peroxidation was increased in pancreatic tumors. CONCLUSION: Zyflo alone or in combination with Celebrex reduce tumor growth in pancreatic cancer and thus might be a new therapeutic option in advanced pancreatic cancer. PMID- 12120009 TI - Power Doppler ultrasonography for the assessment of vascular invasion by pancreatic cancer. AB - AIM: To investigate the diagnostic accuracy of power Doppler ultrasonography (US) in assessing the vascular invasion by pancreatic cancer. METHODS: A prospective study of 40 consecutive patients with pancreatic cancer (head 35, body 5) was performed. All patients underwent surgery. The relationships between tumor and each vessel were classified into four types according to the closest circumferential contact of the tumor with the vessel. A type 0 indicated no contact; type 1 indicated less than one third contact; type 2 indicated one third to 99% contact, and type 3 indicated encasement. Vascular invasion was diagnosed in types 2 and 3. The diagnostic accuracy was evaluated in the portal vein and in the splanchnic arteries (celiac artery, common hepatic artery, and superior mesenteric artery). The power Doppler US findings were confirmed by the operative findings. The results of power Doppler US were compared with those of CT scan and angiography. RESULTS: Portal vein invasion was confirmed in resected specimens in 23 cases and by operative findings in 5 cases. For the diagnosis of portal vein invasion, sensitivity, specificity, and overall accuracy of power Doppler US were, respectively, 79.3, 90.9, and 82.5%. The respective values were 79.3, 100, and 85% for CT and 72.4, 81.8, and 75% for angiography. For the diagnosis of arterial invasion, sensitivity, specificity, and overall accuracy of power Doppler US were 80, 92, and 90%, respectively. The corresponding values were 47, 88, and 73% for CT and 47, 100, and 80% for angiography. CONCLUSION: Power Doppler US proved to be useful for the diagnosis of vascular invasion by pancreatic cancer. PMID- 12120010 TI - Two cases of pancreatic tuberculosis in nonimmunocompromised patients. A diagnostic challenge and a rare cause of portal hypertension. AB - Pancreatic tuberculosis is very rare, especially in immunocompetent patients, and represents a diagnostic challenge. We describe 2 cases of pancreatic tuberculosis mimicking carcinoma on CT scan. In the first case, explorative laparotomy revealed granulomatous inflammation suggestive of tuberculosis. Cultured smears from the pancreatic tail tested positive for Mycobacterium tuberculosis, and the patient responded well to antituberculous medication. In the second case, fine needle aspirate revealed tuberculosis. This case is unique with regard to development of portal hypertension in pancreatic tuberculosis. Antituberculous medication achieved little improvement, then the patient was lost to follow-up. In suspicion of carcinoma the patient underwent laparotomy in another hospital. Malignancy was excluded, and a purulent necrotic pancreas was resected. The patient finally improved without any antituberculous medication and remains well. Both patients were tested HIV-negative. We summarize the etiology, clinical presentation, diagnosis and treatment of a diagnostic dilemma, which should be considered in clinical practice. PMID- 12120011 TI - Benign inflammatory pancreatic mucinous cystadenomas mimicking locally advanced cystadenocarcinomas. Presentation of 3 cases. AB - We report 3 cases of benign mucinous cystadenoma of the pancreas mimicking, both clinically and on imaging findings, locally advanced cystadenocarcinoma spreading to neighbouring organs (stomach, splenic and mesenteric vessels, and diaphragm). Surgical resection was performed in light of the suspicion of invasive carcinoma in all 3 cases. Histological examination of the resected specimens showed entirely benign mucinous cystadenomas associated with marked peri-tumoural inflammation that accounted for the pre-operative misdiagnoses. All 3 patients are alive 40, 47 and 54 months after surgery without evidence of tumour relapse. These cases indicate that surgery must be considered in patients with cystic tumour of the pancreas suggesting locally advanced cystadenocarcinoma, even when pre-operative imaging findings suggest tumour extension into neighbouring organs. PMID- 12120012 TI - Who owns grief. Interview by James S. Russell. PMID- 12120013 TI - Children's Medical Center. Augusta, Georgia. PMID- 12120014 TI - Sekii Ladies Clinic. Furukawa, Miyagi, Japan. PMID- 12120015 TI - Dental clinic. Orlandia, Brazil. PMID- 12120016 TI - Mott Children's Center. Puyallup, Washington. PMID- 12120017 TI - The house of the future has arrived. Researchers at MIT are revolutionizing house design and construction so that aging baby boomers can grow old at home. PMID- 12120018 TI - Digital tools for age-smart housing. PMID- 12120019 TI - How quality managers can stop information overload. PMID- 12120020 TI - Revised disclosure standard presents new compliance challenges. PMID- 12120021 TI - Sentinel event leads to safety checklist. PMID- 12120022 TI - DP, CM skills may stem bed-capacity problems. PMID- 12120023 TI - Health care seeks causes for discharge delays. PMID- 12120024 TI - Paperless records will improve quality. PMID- 12120025 TI - In praise of our unsung nursing leaders. PMID- 12120026 TI - Managing febrile children: when and how to treat. PMID- 12120027 TI - Nursing leadership--looking beyond the lamplight. PMID- 12120028 TI - Questioning the commissioner's impartiality. PMID- 12120029 TI - Regulating health professionals. PMID- 12120030 TI - Government must assist MECA progress. PMID- 12120031 TI - Examining with humility our role as nurses. PMID- 12120032 TI - Learning from student bullying experiences. PMID- 12120033 TI - Changing the horizontal violence culture. PMID- 12120034 TI - NZNO accepts midwifery as a separate profession. PMID- 12120035 TI - A study on a possibility of predicting early relapse in leprosy using a ND-O-BSA based ELISA. AB - Serological methods have been used for detecting infection with Mycobacterium leprae. We have applied a serological test to explore the possibility it could detect a bacterial relapse among patients who have been cured with chemotherapy. More specifically we used an indirect enzyme-linked immunosorbant assay (ELISA) using the natural disaccharide (ND) of the phenolic glycolipid antigen of M. leprae linked to bovine serum albumin as antigen. Antibody levels were measured in sera from normal controls, active leprosy cases, cured leprosy patients, and relapsing leprosy patients. We correlated antibody levels with the type of leprosy, the bacterial index, and with relapse among cured leprosy patients. In our hands, the ND-ELISA, when applied to screening for infection with M. leprae, had excellent sensitivity, specificity, positive and negative predictive values, and both a low false positive rate and a low false negative rate. Antibody levels gradually increased among active patients from the tuberculoid to the lepromatous end of the leprosy spectrum. There was a year-by-year fall in antibody levels in patients responding to chemotherapy. Antibody levels and the bacterial index were correlated using the Spearman's rank correlation method. Serial antibody levels were measured in 666 leprosy patients after being cured with dapsone monotherapy. Over a three year follow up, 95 multibacillary patients became antibody positive and 12 of them had bacterial relapses of their disease. In contrast, among 335 cases that remained antibody negative, only one relapse was seen. Among 44 paucibacillary cured patients who became antibody positive, there was one relapse. There were 192 such patients who remained antibody negative and one relapsed. The risk of relapse is 6.7 times higher among cured multibacillary patients compared to cured paucibacillary patients. Overall, the cumulative relapse rate among antibody positive cases was 13.7%, compared to 0.4% among antibody negative patients. We conclude that the ND-ELISA is a useful tool both for screening for early infection with M. leprae and for predicting a relapse in cured patients, particularly in cured multibacillary patients. PMID- 12120036 TI - The paper grip test for screening on intrinsic muscle paralysis in the foot of leprosy patients. AB - Plantar intrinsic foot muscles provide structure to the foot during walking and thus regulate mechanical foot sole stresses. When paralyzed, for instance in leprosy patients with neuropathy of the distal part of the tibial nerve, there is a high prevalence of plantar ulceration and deformities, especially when muscle weakness goes together with loss of foot sole sensibility. These patients should get immediate care involving education, special footwear and reconstructive surgery before further foot impairment and deformity becomes manifest. Thus far, in leprosy patients little attention is paid to screening of plantar intrinsic muscles activity. This can be done with a new simple and non-invasive method, the Paper Grip Test (PGT). There are two variants for detecting intrinsic muscle weakness of the foot, PGT1 for the great toe and PGT2 for the combined lesser toes. In this study, 517 leprosy patients and 170 healthy volunteers were investigated with the PGT. Sensibility of the foot sole was tested by means of a 10 gram monofilament. Specificity to the PGT1 is found to be about 95.3% which is considered good for physical diagnostic tests. PGT2 is less specific than PGT1. Individual muscle power and understanding of the patient seems to influence the outcome of the test to a certain extent. Sensitivity can only be calculated when the diagnosis is confirmed by electromyography. Especially patients with anesthetic feet, females, older patients and patients with PN-, BB- or LL-types of leprosy appeared to have a higher prevalence of intrinsic foot muscle weakness. All results were analyzed by means of the bivariate Pearson correlation analysis and proved to be statistically significant (p = < 0.05). It is concluded that the PGT1, more than the PGT2, is a useful screening test on the function of plantar intrinsic foot muscles in leprosy patients in hospitals and during fieldwork in developing countries. PMID- 12120037 TI - Light and ultrastructural study of sciatic nerve lesions induced using intraneural injection of viable Mycobacterium leprae in normal and immunosuppressed Swiss white mice. AB - Freshly harvested M. leprae were microinjected into the sciatic nerves of nonimmunosuppressed (non-TR) and immunosuppressed (TR) mice using the technique described by Wisniewski and Bloom. The lesions thus induced, on bypassing the blood-nerve barrier, were biopsied at regular intervals beginning 24 hr and followed up to one year. The fate of M. leprae and the ensuing inflammation and nerve damage were studied using light and electron microscopy. The lesions in both non-TR and TR mice at 24 hr showed an influx of polymorphonuclear leukocytes and an increase in mast cells. The influx and peaking of lymphocytes were delayed by two weeks and 6 weeks, respectively, in TR mice, but the density of lymphocytes at the peak intervals was comparable in both. The plasma cells denoting the humoral response were seen in both, but there was a delay of 3 weeks in non-TR mice. The lesions in non-TR mice showed differentiation of macrophages into epithelioid cells and the formation of giant cells depicting borderline tuberculoid leprosy (BT), Whereas in TR mice, the macrophages showed foamy cytoplasmic changes depicting borderline lepromatous leprosy (BL). Other significant observations common to both non-TR and TR mice were: a) The lesions remained highly localized and showed signs of regression at the 6th and the 12th month intervals. b) The characteristic segmental demyelination and some attempt at remyelination were seen at the site. c) The influx of lymphocytes concorded well with demyelination. d) Bacteria were only seen in the macrophages and never in the Schwann cells or endothelial cells. e) Bacteria persisted in the macrophages, but appeared progressively degenerate at the 6th and 12th post inoculation months, suggesting loss of viability. The study shows that there was a very effective containment of the infection and that the Schwann cells were resistant to M. leprae infection in the neural milieu. Nerve damage and Schwann cell bacillation do not go hand-in-hand. PMID- 12120038 TI - Detection of Mycobacterium leprae DNA by PCR in blood sample from nine-banded armadillo: preliminary results. PMID- 12120039 TI - "A natural sweat test" in leprosy. PMID- 12120040 TI - Single-dose ROM treatment for multilesion paucibacillary leprosy--further observations. PMID- 12120041 TI - A phase 2 open trial of pentoxifylline for the treatment of leprosy reactions. PMID- 12120042 TI - Successful treatment of the paralyzed lower eyelid due to Hansen's disease by implanting auricular cartilage. PMID- 12120043 TI - Intraocular lens implantation in leprosy. AB - The preoperative, operative and postoperative ocular complications in 48 eyes of 39 leprosy patients who underwent standard extracapsular cataract extraction and posterior chamber intraocular lens implantation, by the same surgeon, were studied retrospectively. Seventeen were male and 22 were female. Thirteen (33%) were paucibacillary (PB) while 26 (67%) were multibacillary (MB) patients. Three patients were smear-positive at the time of surgery. Grade 2 deformity that included claw hands, absorbed fingers, saddle noses and foot drop were present in 64% of the patients. None of the patients had any previous intraocular inflammation although one patient had previously had a Type 1 reaction and 5 patients had previously had Type 2 reactions. Preoperative complications like corneal opacities (3 eyes) and lagophthalmos (5 eyes) were not associated with lower vision postoperatively. No significant operative complications like vitreous loss, endothelial damage or iris tear were encountered, except in one eye where there was a posterior capsular tear. Seventeen eyes (35%) developed uveitis of 3+ or more in the immediate postoperative period, but abated with routine topical steroid eye drops. Six months after surgery 7 out of 47 eyes (15%) had developed posterior capsular opacities. There were no significant differences (p = > 0.05) in the visual acuity outcomes or in ocular complications when MB patients were compared with PB patients. Smear-positive patients were not significantly different from smear-negative patients when postoperative complications were compared. Visual outcomes in the 23 eyes followed up at two years after surgery were 6/18 or higher, except in one eye which had sustained a severe injury one year after surgery. IOLs were found to be safe and beneficial in this series of patients, but a much larger prospective study with matched normal controls is needed to prove the safety and efficacy of IOLs in leprosy patients. PMID- 12120044 TI - Squalene-hopene cyclase: catalytic mechanism and substrate recognition. AB - Rapid progress on the catalytic mechanism and substrate recognition by squalene hopene cyclase, which has occurred only in the last several years, is reported. A series of site-directed mutation experiments and some squalene analogues have provided deep insight into the polycyclization mechanism and catalytic sites in conjunction with the information from X-ray crystal data. PMID- 12120045 TI - Three-dimensional imaging of YB56 by high-resolution electron microscopy. AB - The three-dimensional potential map of YB56 was obtained by inverse Fourier transformation of three-dimensional phases and amplitudes in three high resolution images taken along the [100], [110] and [111] directions of YB56 crystals; the size of the imaging region was 14 nm x 14 nm x approximately 4 nm, and the image directly showed the three-dimensional potential map of the crystal, a useful method for three-dimensional structure analysis in nanoscale regions. PMID- 12120046 TI - Susceptibility of a heterogeneous catalyst, Rh/gamma-alumina, to rapid structural change by exposure to NO. AB - Metal particles in a Rh/gamma-Al2O3 catalyst of differing particle size are oxidised by NO/He within 5 seconds at 313 K; rapid, highly exothermic dissociative chemisorption of NO is the initial step. PMID- 12120047 TI - Helix versus zig-zag: control of supramolecular topology via carboxylic acid conformations in ortho-substituted phenyl amines. AB - ortho-Substituted phenyl amines form supramolecular helices or zig-zag structures, depending on the conformation of the carboxylic acid substituents- which can be controlled by an intramolecular hydrogen-bonding interaction. PMID- 12120048 TI - Synthesis and reactivity of a ferrocene-derived PCP-pincer ligand. AB - The 1,3-bis(diphosphinomethyl)ferrocene 3 readily reacts with [(C2H4)2RhCl]2 to form an equilibrating pair of diastereomers 8a and 8b by C-H insertion into the ferrocene. PMID- 12120050 TI - An organic template approach for the synthesis of selectively functionalised tetraazacycloalkanes. AB - Selectively functionalised tetraazacycloalkanes are obtained from the open-chain tetraamine by using a bisaminal moiety acting both as a template agent and as a N protecting group. PMID- 12120049 TI - Efficient kinetic resolution in hydroboration of 1,2-dihydronaphthalenes. AB - 1-Substituted 1,2-dihydronaphthalenes undergo kinetic resolution during asymmetric hydroboration with Rh-QUINAP complexes. PMID- 12120051 TI - Unusual cyclo-tetra and hexa peptidation of bis-boc-cystine with cystine-di-OMe: one step preparation of the novel 32- and 48-membered cyclotetracystine and cyclohexacystine. AB - The unprecedented formation of 32- and 48-membered macrocycles that inscribe 4 and 6 cystine units, in the peptidation of bis-Boc-cystine with cystine di-OMe is reported. PMID- 12120052 TI - The first samarium(II)-mediated stereoselective spirocyclization onto an aromatic ring. AB - The first samarium(II)-mediated spirocyclisation onto an aromatic ring was achieved by the reaction of methyl 4-(4-oxoalkyl)benzoates with SmI2 in the presence of i-PrOH and HMPA, yielding methyl 1-alkyl-1-hydroxyspiro-[4.5]dec-6 ene-8-carboxylates in moderate to high yields. PMID- 12120053 TI - Redox control of the P450cam catalytic cycle: effects of Y96F active site mutation and binding of a non-natural substrate. AB - Spectroelectrochemistry measurements are used to demonstrate that active site mutation and binding of an non-natural substrate to P450cam (CYP101) reduces the shift in the redox potential caused by substrate-binding, and thereby results in slower catalytic turnover rate relative to wild-type enzyme with the natural camphor substrate. PMID- 12120055 TI - Hexa(2-pyridyl)[3]radialene: self-assembly of a hexanuclear silver array. AB - Reaction of the new ligand hexa(2-pyridyl)[3]radialene with silver tetrafluoroborate results in the formation of a M6L2 cage with an encapsulated mu 3-fluorido anion. PMID- 12120054 TI - Mixed nitrosyl/phosphinidene and nitrene/phosphinidene clusters of ruthenium. AB - Reaction of the aminophosphinidene complex [Ru5(CO)15(mu 4-PNPri2)] 1 with [PPN][NO2] (PPN = Ph3P=N=PPh3) led to the mixed nitrosyl/phosphinidene cluster complex [PPN][Ru5(CO)13(mu-NO)(mu 4-PNPri2)] 2 which is transformed into the novel nitrene/phosphinidene cluster [Ru5(CO)10(mu-CO)2(mu 3-CO)(mu 4-NH)(mu 3 PNPri2)] 3 via treatment with triflic acid. PMID- 12120056 TI - Hexakis(2-pyridyl)- and hexakis(3-pyridyl)[3]radialene: novel, water-soluble [3]radialenes with potential utility for supramolecular chemistry. AB - The titled [3]radialenes are the first, water-soluble hexaaryl[3]radialenes with considerably high electron affinity, their alkali metal reduction giving rise to anion-radicals and dianions stepwise as fairly stable species in degassed tetrahydrofuran. PMID- 12120057 TI - Para-acyl calix[4]arenes: amphiphilic self-assembly from the molecular to the mesoscopic level. AB - Studies of the properties of a series of amphiphilic calixarenes show that they can form stable monolayers, Langmuir-Blodgett layers and solid lipid nanoparticles, the mesostructures were investigated by atomic force microscopy and the crystal structure of one compound shows a partially interdigitated tilted bilayer structure. PMID- 12120058 TI - [30]heptaphyrin(1.1.1.1.1.0.0): an aromatic expanded porphyrin with a 'figure eight' like structure. AB - The synthesis, characterization, and X-ray structure of a heptapyrrolic macrocycle, [30]heptaphyrin(1.1.1.1.1.0.0) is reported; despite showing aromatic features, it exhibits a 'figure eight'-like structure in the solid state. PMID- 12120061 TI - Evidence for through-framework electron transfer in intrazeolite photochemistry. Case of Ru(bpy)3(2+) and methylviologen in novel delaminated ITQ-2 zeolite. AB - A sample of novel delaminated zeolite ITQ-2 containing Ru(bpy)3(2+) on the external cups and MV2+ included in the independent and not connected channels has been prepared; emission and time-resolved laser flash photolysis has shown unambigously that photoinduced electron transfer from Ru(bpy)3(2+) to MV2+ occurs through the zeolite framework. PMID- 12120059 TI - Silica-based powders and monoliths with bimodal pore systems. AB - Porous pure and doped silicas with pore sizes at two length scales (meso/macroporous) have been prepared and shaped both as powders and monoliths through a one-pot surfactant assisted procedure by using a simple template agent and starting from atrane complexes as inorganic precursors. PMID- 12120060 TI - Synthesis of dicobalt hexacarbonyl 5-p-tolylethynyl-2'-deoxyuridine. AB - Reactions of 5-p-tolylethynyl-2'-deoxyuridine and 3',5'-di-O-acetyl-5-p tolylethynyl-2'-deoxyuridine with Co2(CO)8 in THF gave 5-p-tolC2[CO2(CO)6]-2' deoxyuridine and 3',5'-di-O-acetyl-5-p-tolC2[CO2(CO)6]-2'-deoxyuridine (92 and 66%). PMID- 12120062 TI - Possible high-pressure structures of sulfur trioxide. AB - Calculations with the linearized augmented plane wave method indicate that several high-density forms of sulfur trioxide should be accessible at pressures above 29 GPa, with densities up to 1.7 times larger than the presently known forms of solid SO3. PMID- 12120063 TI - Calixarenes as ligands for transition-metal catalysts: a bis(calix[4]arene-11,23 dicarboxylato) dirhodium complex. AB - A novel dirhodium tetracarboxylate complex is described in which two calix[4]arene macrocycles, bridged at the upper rim by a Rh-Rh unit, serve as ligands and whose solid-state structure shows an unusual coordination of a toluene molecule in the axial position at each rhodium atom. PMID- 12120064 TI - Compartmental Schiff-base ligands as selective double-loaded extractants for copper(II). AB - The Robson compartmental macrocyclic ligand derived from the condensation of derivatives of 2,6-diformylphenol and diamines has been prepared for the first time in its free ligand form; competitive three-phase transport and two-phase extraction studies confirm high double-loaded selectivity for the binding and delivery of Cu(II). PMID- 12120065 TI - Microwave-assisted preparation of dialkylimidazolium tetrachloroaluminates and their use as catalysts in the solvent-free tetrahydropyranylation of alcohols and phenols. AB - Microwave-assisted preparation of dialkylimidazolium tetrachloroaluminates, [CnMIM][AlCl4] and their application as recyclable catalysts for the efficient and eco-friendly protection of alcohols as tetrahydropyranyl (THP) ethers are described. The same catalyst can also be utilized for the deprotection of THP ethers. PMID- 12120066 TI - Unusually long cooperative chain of seven hydrogen bonds. An alternative packing type for symmetrical phenols. AB - Conformational flexibility in a symmetrical tris-phenol leads to close packed structures that are also characterised by an extended though finite cooperative chain of hydrogen bonds. PMID- 12120068 TI - Tetraalkylammonium pentaorganosilicates: the first highly stable silicates with five hydrocarbon ligands. AB - Several tetraalkylammonium pentaorganosilicates, derived from 9,9-spirobi(9H,9 silafluorene) were prepared from the corresponding lithium silicates and isolated and characterized as storable high melting solids. PMID- 12120067 TI - A ring expansion reaction of 1,3-oxathiolanes. AB - 1,3-Oxathiolanes are efficiently converted, via sulfur ylide intermediates, to 1,4-oxathianes by ring expansion with a silylated diazoacetate in the presence of a copper catalyst. PMID- 12120069 TI - Fabrication of compact silver nanoshells on polystyrene spheres through electrostatic attraction. AB - Nanoshells composed of close-packed silver nanocrystals have been fabricated on polystyrene spheres via direct electrostatic attraction at appropriate pH; the thickness and roughness of the shell can be readily controlled through a layer-by layer technique. PMID- 12120070 TI - Problems and solutions for alkene polymerisation catalysts incorporating Schiff bases; migratory insertion and radical mechanisms of catalyst deactivation. AB - Steric blocking of an intramolecular 1,2-migratory insertion reaction of a zirconium salicylaldiminato complex leads to a long-lived catalyst for ethene polymerisation, but promotes a new radical catalyst decomposition mechanism in certain instances; kinetic and thermodynamic parameters for both pathways have been established. PMID- 12120071 TI - Crosslinking a palladium(II) polymer gives a laminated sheet structure. AB - Chains of a zigzag coordination polymer containing bis(pyridyl) bridging groups between palladium(II) centres can be arranged to give a laminated sheet structure by a biomimetic approach in which hydrogen bonding involving amide groups is the key feature. PMID- 12120072 TI - Evidence that gold(III) porphyrins are not electrochemically inert: facile generation of gold(II) 5,10,15,20-tetrakis(3,5-di-tert-butylphenyl)porphyrin. AB - Gold(III) porphyrin 1 is shown to undergo reduction at the central metal ion to give the first known gold(II) porphyrin overturning the long held assumption that reduction of such complexes only occurs at the macrocycle. PMID- 12120073 TI - Anion sensing 'venus flytrap' hosts: a modular approach. AB - A series of podands based on three hydrogen bonding 'arms' have been prepared and their affinities for simple inorganic anions measured. PMID- 12120074 TI - Ligand movement modulates the rate of proton transfer in reactions of [Fe4S4Cl4]2 . AB - Substitution of the first chloro-ligand in [Fe4S4Cl4]2- by 4-RC6H4S- (R = CF3, Cl, H, Me or MeO), in the presence of [H2N(CH2)3CH2]+, involves initial binding of thiolate, followed by protonation and finally chloride dissociation; the rate of protonation is facilitated by electron-withdrawing R-substituents indicating that Fe-thiolate bond length changes modulate proton transfer. PMID- 12120075 TI - A stable metal coordination polymer gel based on a calix[4]arene and its 'uptake' of non-ionic organic molecules from the aqueous phase. AB - Here we report a coordination polymer based, stable organogel (termed 'metallogel' to emphasize the essential role of metal-ligand bonding during the gelation), which 'uptakes' neutral organic molecules from the aqueous phase. PMID- 12120076 TI - Synthesis and characterization of [Cu(Me2oxpn)Ni(NO2)(tmen)](ClO4): a single ferrimagnetic dinuclear CuII-NiII complex acting as weak molecule-based magnet. AB - The new heterodinuclear complex [Cu(Me2oxpn)Ni(NO2)(tmen)](ClO4), that exhibits strong antiferromagnetic intramolecular coupling between CuII and NiII ions (ferrimagnetic behavior), shows ferromagnetic ordering at low temperature, due likely to a small canting phenomenon; it is one of the very few compounds made from isolated molecules that lead to cooperative magnetic behavior. PMID- 12120077 TI - Directly observed covalent coupling of quantum dots to single-wall carbon nanotubes. AB - Carboxylate chemistry is used to covalently couple metal nanoparticles to defect sites in controllably oxidized single-walled carbon nanotube termini and side walls, and this process monitored by atomic force microscopy. PMID- 12120078 TI - Toward constructing nanoscale hydroxo-lanthanide clusters: syntheses and characterizations of novel tetradecanuclear hydroxo-lanthanide clusters. AB - Two novel tetradecanuclear hydroxo-lanthanide complexes, formulated as Ln14(mu 4 OH)2(mu 3-OH)16(mu-eta 2-acac)8(eta 2-acac)16, Ln = Tb, 1; Eu, 2; acac = acetylacetonato, were synthesized and characterized by single-crystal X-ray diffraction, elemental analysis and phosphorescence spectroscopy. PMID- 12120079 TI - Zipping up 'the crushed fullerene' C60H30:C60 by fifteen-fold, consecutive intramolecular H2 losses. AB - MALDI TOF-MS of tribenzo[l:1':1"]benzo[1,2-e:3,4-e':5,6-e"]triacephenanthrylene (1a, C60H30) gives C60.+ by multiple intramolecular cyclodehydrogenation reactions. PMID- 12120080 TI - Synthesis of the RuIV amido complex [TpRu(CO)(PPh3)(NHPh)][OTf]2 (Tp = hydridotris(pyrazolyl)borate; OTf = trifluoromethanesulfonate) and deprotonation to form an octahedral and d4 imido complex: computational study of RuIV-imido bonding. AB - Deprotonation of [TpRu(CO)(PPh3)(NHPh)][OTf]2 yields the thermally unstable d4 imido complex [TpRu-(CO)(PPh3)(NPh)][OTf]; a computational study of the bonding of the imido complex provides a foundation for discussion of its instability in terms of pi-conflict. PMID- 12120081 TI - Quasi-solid-state dye-sensitized solar cells using room temperature molten salts and a low molecular weight gelator. AB - A dye-sensitized solar cell fabricated using the room temperature molten salt, 1 hexyl-3-methylimidazolium iodide, iodine and a low molecular weight gelator as a quasisolid-state electrolyte showed a 5.0% light-to-electricity conversion efficiency under AM 1.5 irradiation, and high-temperature stability. PMID- 12120082 TI - Hydride encapsulation in s-block metal inverse crown chemistry. AB - A new category of 'inverse crown' complex has been established through the synthesis of the disodium-dimagnesium diisopropylamide [Na2Mg2(N(Pri)2)4(mu H)2.(toluene)2], the first such complex to exhibit hydride encapsulation, a non planar octagonal 'host' ring or solvent stabilisation. PMID- 12120083 TI - Size quantized formation and self-assembly of gold encased silver nanoparticles. AB - Mixing aqueous dispersions of thiocyanate ion coated small (< 3.5 nm diameter) gold nanoparticles and EDTA covered larger (> 22 nm diameter) silver nanoparticles, results in the formation of robust gold encased silver nanoparticles; in contrast to using larger (> 11 nm diameter) gold nanoparticles which forms chained structures. PMID- 12120084 TI - Nitrones are suitable ligands for heme models: X-ray crystal structure of the first metalloporphyrin nitrone complex. AB - The metalloporphyrin nitrone complexes [M(oep)-(CO)(DMPO)] (M = Ru, Os; oep = octaethylporphyrinato dianion; DMPO = 5,5-dimethyl-1-pyrroline N-oxide) have been prepared: the crystal structure of the Ru complex reveals a sole eta 1-O binding mode of the nitrone ligand to the metal center. PMID- 12120085 TI - Picosecond time-resolved infrared spectroscopic investigation of excited state dynamics in a PtII diimine chromophore. AB - This is the first report describing the use of picosecond time-resolved infrared (TRIR) spectroscopy to probe a d8 metal chromophore, Pt(4,4'-(CO2Et)(2)-2,2' bpy)Cl2: monitoring changes in the v(CO) vibrations allows for an assignment of the lowest excited state to an MLCT with an 8.7 ps lifetime. PMID- 12120086 TI - Molybdenum alkynyls as alkynyl transfer reagents. AB - Molybdenum alkynyl complexes [Mo(C identical to CR)(eta 3-allyl)(CO)2(phen)] feature long Mo-Calkynyl bond distances and propensity to undergo the cleavage of these bonds, a property that allowed their use as acetylide transfer reagents. PMID- 12120087 TI - Investigation of the factors controlling the regioselectivity of the hydroboration of fluoroolefins. AB - Either Markovnikov or anti-Markovnikov regioselectivity can be achieved at will during the hydroboration-oxidation of perfluoroalkyl(aryl)ethylenes by varying the hydroborating agent. PMID- 12120088 TI - Aqueous enantioselective hydrogenation of methyl 2-acetamidoacrylate with Rh MeDuPHOS occluded in PDMS. AB - A new chiral heterogeneous catalytic system obtained by occlusion of the Rh MeDuPHOS complex in a polydimethylsiloxane film was tested in the asymmetric hydrogenation of methyl 2-acetamidoacrylate in aqueous medium. PMID- 12120089 TI - Rhodium-catalysed addition of arylboronic acids to oxabenzonorbornadienes. AB - The facile addition reaction of boronic acids to oxabenzonorbornadienes was achieved using a catalytic amount of a rhodium(I) complex having P(OEt)3 ligands, affording cis-2-aryl-1,2-dihydro-1-naphthol stereoselectively, and in good yield without concomitant deboronation of the boronic acid. PMID- 12120090 TI - Apoptosis-based anticancer drugs. PMID- 12120091 TI - Efficacy at G-protein-coupled receptors. AB - At present, the drug-discovery process centres on ligands that either block or produce physiological responses. However, there are therapeutic uses for ligands that do neither of these things, but which still affect receptors in other ways. This review discusses the intimate relationship between the affinity of a ligand for its receptor, and the probability that the binding of the ligand will produce some change in the receptor, resulting in efficacy. This, in turn, argues that ligands that have affinity should be tested more broadly, for a wider range of efficacies, to detect hidden therapeutic activities. PMID- 12120092 TI - Apoptosis-based therapies. AB - Many of today's medical illnesses can be attributed directly or indirectly to problems with apoptosis--a programmed cell-suicide mechanism. Disorders in which defective regulation of apoptosis contributes to disease pathogenesis or progression can involve either cell accumulation, in which cell eradication or cell turnover is impaired, or cell loss, in which the cell-suicide programme is inappropriately triggered. Identification of the genes and gene products that are responsible for apoptosis, together with emerging information about the mechanisms of action and structures of apoptotic regulatory and effector proteins, has laid a foundation for the discovery of drugs, some of which are now undergoing evaluation in human clinical trials. PMID- 12120093 TI - The potential for novel anti-inflammatory therapies for coronary artery disease. AB - Although drugs that lead to cholesterol and lipid lowering have proved to have significant effects in lowering cardiovascular morbidity and mortality, coronary artery disease remains a principal cause of death worldwide. There is a clear need to discover further therapeutic approaches to control this disease adequately. This review focuses on the mechanisms that have been implicated in the recruitment, activation and differentiation of inflammatory monocytes/macrophages in nascent vascular lesions into lipid-laden foam cells. These mechanisms might provide attractive targets for novel therapies for coronary artery disease. PMID- 12120094 TI - Drug and gene targeting to the brain with molecular Trojan horses. AB - Getting drugs and genes into the brain is a tall order. This is because the presence of the blood-brain barrier prevents many molecules from crossing into the brain. Overcoming this problem will have a profound effect on the treatment of many neurological disorders, allowing larger water-soluble molecules to pass into the brain. Transport vectors, such as endogenous peptides, modified proteins or peptidomimetic monoclonal antibodies, are one way of tricking the brain into allowing these molecules to pass. This article will review such molecular Trojan Horses, and the progress that has been made in the delivery of drugs and genes to the brain. PMID- 12120095 TI - Novel drug development opportunities for heparin. AB - The glycosaminoglycan heparin has been used in the clinic as an anticoagulant for more than 50 years. A fully characterized sequence in native heparin is known to be responsible for this activity. However, heparin is a complex polysaccharide, which has an array of properties that are unrelated to its anticoagulant activity. Recent research has provided us with an increased understanding of the specific structural requirements for the various actions of heparin, indicating that it might be possible to create 'tailor-made' sequences based on the heparin template to isolate specific therapeutic activities. This research should provide the basis for novel drug treatments for a range of diseases, including cancer and various inflammatory diseases. PMID- 12120096 TI - Animal experimentation: the continuing debate. AB - The use of animals in research and development has remained a subject of public debate for over a century. Although there is good evidence from opinion surveys that the public accepts the use of animals in research, they are poorly informed about the way in which it is regulated, and are increasingly concerned about laboratory-animal welfare. This article will review how public concerns about animal experimentation developed, the recent activities of animal-rights groups, and the opportunities and challenges facing the scientific community. PMID- 12120098 TI - New money for old rope? AB - Randomized, placebo-controlled, clinical trials tell us whether a drug works, but they normally fail to reveal whether a drug works better than existing therapies. The increasing consideration now being given to the cost-effectiveness of available treatment options should spur a move towards more comparative trials. PMID- 12120097 TI - Metabonomics: a platform for studying drug toxicity and gene function. AB - The later that a molecule or molecular class is lost from the drug development pipeline, the higher the financial cost. Minimizing attrition is therefore one of the most important aims of a pharmaceutical discovery programme. Novel technologies that increase the probability of making the right choice early save resources, and promote safety, efficacy and profitability. Metabonomics is a systems approach for studying in vivo metabolic profiles, which promises to provide information on drug toxicity, disease processes and gene function at several stages in the discovery-and-development process. PMID- 12120099 TI - Beyond body mass index. AB - Body mass index (BMI) is the cornerstone of the current classification system for obesity and its advantages are widely exploited across disciplines ranging from international surveillance to individual patient assessment. However, like all anthropometric measurements, it is only a surrogate measure of body fatness. Obesity is defined as an excess accumulation of body fat, and it is the amount of this excess fat that correlates with ill-health. We propose therefore that much greater attention should be paid to the development of databases and standards based on the direct measurement of body fat in populations, rather than on surrogate measures. In support of this argument we illustrate a wide range of conditions in which surrogate anthropometric measures (especially BMI) provide misleading information about body fat content. These include: infancy and childhood; ageing; racial differences; athletes; military and civil forces personnel; weight loss with and without exercise; physical training; and special clinical circumstances. We argue that BMI continues to serve well for many purposes, but that the time is now right to initiate a gradual evolution beyond BMI towards standards based on actual measurements of body fat mass. PMID- 12120100 TI - Interventions for preventing obesity in childhood. A systematic review. AB - BACKGROUND: The prevalence of obesity and overweight is increasing worldwide. Obesity in children impacts on their health in both short- and long-term. Obesity prevention strategies are poorly understood. OBJECTIVE: To assess the effectiveness of interventions designed to prevent obesity in childhood. SEARCH STRATEGY: Electronic databases were searched from January 1985 to October 1999. SELECTION CRITERIA: Data from randomized control trials and non-randomized trials with concurrent control group were included. A priori, studies with follow up of 1 year minimum were selected however, this was subsequently amended to include studies with a minimum follow up of three months. DATA COLLECTION & ANALYSIS: Two reviewers independently extracted data and assessed study quality. MAIN RESULTS: Seven studies were included, three long-term (> 1 years) and four short-term (> 3 months and < 1 years). The studies included were diverse in terms of study design and quality, target population, theoretical underpinning of intervention approach, and outcome measures. As such, it was not appropriate to combine study findings using statistical methods. CONCLUSIONS: Two of the long-term studies (one focused on dietary education and physical activity vs. control, and the other only on dietary education vs. control), resulted in a reduction in the prevalence on obesity, but the third, which focused on dietary education and physical activity, found no effect. Of the four short-term studies, three focused simply on physical activity/reduction of sedentary behavious vs. control. Two of these studies resulted in a reduction in the prevalence of obesity in intervention groups compared with control groups, and another study found a non significant reduction. The fourth study focused on dietary education and physical activity, and did not find an effect on obesity, but did report a reduction in fat intake. Overall, the findings of the review suggest that currently there is limited quality data on the effectiveness of obesity prevention programmes and as such no generalizable conclusions can be drawn. The need for well-designed studies that examine a range of interventions remains a priority. PMID- 12120102 TI - Obesity and health-related quality of life. AB - Although it is well documented that obesity is strongly associated with morbidity and mortality, less is known about the impact of obesity on functional status and health-related quality of life (HRQL). However, in recent years research has been conducted to estimate the impact of obesity on HRQL, and to determine the effects of weight reduction on HRQL. The majority of published studies indicate that obesity impairs HRQL, and that higher degrees of obesity are associated with greater impairment. Obesity-associated decrements on HRQL tend to be most pronounced on physical domains of functioning. Studies of the effect of obesity surgery among morbidly obese patients indicate that this procedure produces significant and sustained improvements in the majority of HRQL indices; among mild-to-moderately obese persons, modest weight reduction derived from lifestyle modification also appears to improve HRQL, at least in the short term. Additional research is needed to (1) further characterize the effect that obesity has on HRQL; (2) estimate the short- and long-term effects of various methods of weight reduction (e.g. surgery, lifestyle modification) on HRQL; (3) improve both the conceptualization and measurement of HRQL to incorporate the personal preferences and values of the patient; and (4) develop ways to enhance and sustain positive changes in HRQL, even if weight maintenance is elusive. PMID- 12120103 TI - Obesity in the elderly--a future matter of concern? AB - In most studies mean body weight increases with age up to about age 60 and then levels off, but information about the association between body weight and mortality at higher ages is sparse, since most studies published are cross sectional, thus introducing a bias in selectivity. Some studies actually suggest a protective effect of overweight in the oldest age groups. Indices of visceral obesity may be better indicators of risk than body mass index (BMI) in these age groups. Not only actual weight, but also weight development over the last decades may predict outcome. Most clinical trials exclude older patients and little is known about benefits of diets or drugs inducing weight loss in these age groups. More information is available suggesting multiple benefits of physical activity. Mechanical complications of obesity, such as osteoarthritis and static respiratory complications seem to improve with weight loss even at higher ages. For health economic reasons it will become important to address treatment strategies in the elderly in the future, since they will constitute a large segment of the population. Recent studies suggest that bariatric surgery, previously considered contraindicated in obese patients above age 60 can be safely performed even in patients above age 70 and with the same benefits as for younger subjects. PMID- 12120104 TI - The medical-care cost burden of obesity. AB - Recent years have seen a dramatic rise in the prevalence of obesity in many countries, stimulating interest in the health and economic consequences of this phenomenon. In this article, we provide a systematic review of the literature on the medical-care cost burden of obesity. Relevant studies were identified using a computerized search of the medical literature for English-language articles published between 1990 and 2001. The 18 studies that met all criteria for inclusion in the review can be classified as modelling or database studies and further distinguished as cross-sectional or longitudinal in nature. The majority of studies that have been conducted are cross-sectional modelling studies, including 10 studies reporting the burden of obesity to national health systems. These suggest that obesity accounts for 5.5-7.0% of national health expenditures in the United States and 2.0-3.5% in other countries for which estimates have been reported. Other studies highlight the burden of obesity from other perspectives, including employers and health plans, as well as the impact of obesity on future disease risks and associated medical-care costs. Despite various methodological limitations, discussed herein, this body of research leads to the inescapable conclusion that obesity exacts an immense economic toll in various countries throughout the world. PMID- 12120101 TI - Childhood overweight: a contextual model and recommendations for future research. AB - The prevalence of overweight among children has doubled within the past two decades. Increases in the rate of childhood overweight are of particular concern due to the negative health and psychological effects noted among overweight children. As shown by previous research, the development of childhood overweight involves a complex set of factors from multiple contexts that interact with each other to place a child at risk of overweight. This multifaceted system can be conceptualized using Ecological Systems Theory (EST). EST highlights the importance of considering the context(s), or ecological niche, in which a person is located in order to understand the emergence of a particular characteristic. In the case of a child, the ecological niche includes the family and the school, which are in turn embedded in larger social contexts including the community and society at large. In this review, EST is used as a framework with which to summarize research assessing predictors of childhood overweight. Specifically, child characteristics that place children at risk of the development of overweight (including dietary intake, physical activity, and sedentary behaviour) will be reviewed while taking into consideration the influence of the familial environment, the school environment, and the community and larger social environments. It is concluded that future research needs to adopt a broader contextual approach in order to understand and intervene against the processes leading to the development of overweight among children and that the use of theories or paradigms such as EST will facilitate developing and testing models of causal processes. PMID- 12120105 TI - The safety and efficacy of pharmaceutical and herbal caffeine and ephedrine use as a weight loss agent. AB - Since passage of the Dietary Supplement Health and Education Act of 1994, the sale of herbal dietary supplements containing caffeine and ephedrine for weight loss has become widespread in the United States. Reports of adverse events associated with the use of these non-prescription supplements have raised concerns in the United States regulatory community. Restricting the use of these products is now being considered. Such restriction should be based upon controlled clinical trials. This review of the literature in Medline relative to the use of caffeine and ephedrine in the treatment of obesity concludes that caffeine and ephedrine are effective in causing weight loss. Caffeine and ephedrine give equivalent weight loss to Diethylpropion and superior weight loss compared to dexfenfluramine. Caffeine and ephedrine have a long history of safe, non-prescription use. The adverse events accompanying acute dosing are mild and transient. Adverse events with caffeine and ephedrine reach and remain at placebo levels after 4-12 weeks of continuous treatment, but data from randomized trials up to 6 months only are available. Obesity is chronic, requires chronic treatment, its incidence is increasing and it has few effective treatments. The benefits of caffeine and ephedrine in treating obesity appear to outweigh the small associated risks. Restriction of dietary herbal supplements containing caffeine and ephedrine, often with other ingredients, should be based on controlled clinical trials of these products. PMID- 12120106 TI - The dilemma of decision-making: processing thinking critical to nursing. PMID- 12120107 TI - Assessing critical thinking in baccalaureate nursing students: a longitudinal study. AB - The purpose of this study was to evaluate the attainment of critical thinking skills of students before and after curriculum revision of a baccalaureate nursing program. The California Critical Thinking Skills Test (CCTST) was used to measure the critical thinking ability of the students at program entry, midpoint, and at exist. The sample consisted of three cohorts of students: cohort 1 (n = 55) was the baseline class before curriculum revision, whereas cohorts 2 (n = 55) and 3 (n = 73) were the first two classes to experience the revised curriculum. The results revealed that cohort 2 achieved significantly higher critical thinking scores than the baseline cohort. Cohort 2 also improved dramatically on all subscales from test 1 to test 3. However, cohort 3 failed to demonstrate improved critical thinking scores over time. Findings have implications for measuring critical thinking. PMID- 12120108 TI - Evaluation of critical thinking outcomes of a BSN program. AB - Following a curriculum revision, which emphasized critical thinking, a school of nursing selected the California Critical Thinking Skills Test (CCTST) as a standardized outcomes measure for its bachelor's of science in nursing (BSN) program. Students in three tracks of the program were administered the CCTST on entry into the nursing curriculum and again on exit. Paired t tests for dependent samples were used to analyze pretest-posttest differences of all students (N = 136) in the program. Results for students in each of the three tracks demonstrated significantly improved (P < or = .05) CCTST scores on all subscales and total scores, with one exception. RN to BSN students' scores on the Analysis subscale approached but did not reach significance (P = .058). Implications for evaluation are discussed. PMID- 12120109 TI - Acupuncture as complementary therapy for back pain. AB - Back pain affects more than 9 million people in the United States and accounts for 25% of disability in all work-related injuries. It has a lifetime prevalence of 60%-90% and it costs more than $50 billion annually to manage back pain. The incidence of back pain in nurses is over 80% and accounts for more than 150 million working days per year being lost. Western treatment of back pain is controversial and often ineffective. Acupuncture as a therapeutic intervention is practiced widely in the United States. Research has demonstrated that acupuncture may benefit those who suffer from back pain when they have failed to respond to previous treatment by drugs, bed rest, epidural injection, physiotherapy, osteopathy, chiropractics, and surgery. Acupuncture is a powerful and complementary therapy for back pain. PMID- 12120110 TI - The influence of a baccalaureate program on traditional, RN-BSN, and accelerated students' critical thinking abilities. AB - Although there are multiple ways of achieving baccalaureate-level nursing education, all graduates must demonstrate critical thinking abilities to practice competently. The purpose of this study was to measure the changes in critical thinking abilities of students pursuing various pathways in the same baccalaureate nursing curriculum. Traditional, registered nurse-bachelor of science (RN-BSN), and accelerated students completed the Watson-Glaser Critical Thinking Appraisal (WGCTA) at the beginning and end of their nursing course sequence. Findings revealed a significant difference between the pre- and post WGCTA scores of traditional students (t = -2.84, P = .007) and RN-BSN students (t = -2.28, P = .029), but not of accelerated students. Similarities between the curricular pathways that could account for the results were analyzed. Further research is needed to determine the best combination of liberal arts and nursing courses for the development of critical thinking abilities. PMID- 12120111 TI - Surrounded by ocean, a world apart ... the experience of elder women living alone. AB - The purpose of this phenomenological study was to discover, describe, and understand the lived experience of some of Maine's most isolated elder women, those living alone on islands off the coast. Nine women, age 80 and older, were interviewed. Three dominant themes essential to this rural island lifeway emerged: no one is an island, no one lives alone; securely anchored in safe harbor; and weathering the storms. Results were that these women remain actively engaged in life, feeling valued and connected, safe and secure; they are resourceful, resilient, and determined. PMID- 12120112 TI - The integration of mind-body-soul and the practice of humanistic nursing. AB - Nursing has shifted the focus of its praxis toward a commitment to holistic care. This holistic perspective suggests each nurse must bring an authentic self as the essential element of therapeutic participation with another human being. The art of the therapeutic use of self in relationships involves an aesthetic process of ongoing reflected action, whereby an individual strives toward a sense of harmony and balance within oneself and with the world. The artful use of self provides an opportunity for expansion and personal growth and actualizes the potential to expand the good of all. PMID- 12120113 TI - Spirituality in nursing from a Japanese perspective. AB - The purpose of this study was to explore the concept of spirituality and its expression among persons in a Japanese farming community. It was found that spirituality was described as "living in harmony with nature and surrounding people." Common expressions of this spirituality were through faith and ways of worship, prayer, search for inner peace, hope and thanksgiving, including spiritual well-being. The latter was defined as showing thanks to nature, ancestors, and people; caring for surrounding people; and peacefully embracing the concept of death. To provide congruent spiritual care, it was suggested that nurses need to expand their own knowledge and understanding of spirituality; integrate spirituality in their nursing care; and improve communication with their clients and their families. PMID- 12120114 TI - Trance and suggestion: timeless interventions and implication for nurses in the new millennium. AB - Trance is a normal psychophysiological phenomenon. Suggestion is the purposeful use of that phenomenon and the heart of hypnosis. Suggestion deepens and enriches trance and promotes beneficial changes, or healing, within the individual. This article addresses the traditional historical and cultural roots of the hypnotic experience and use of "trance" and "suggestion." How nurses often unknowingly use these phenomena and can intentionally use trance and suggestion in their practices is explored. PMID- 12120115 TI - Using student writing assignments to assess critical thinking skills: a holistic approach. AB - This work offers an example of one school's holistic approach to the evaluation of critical thinking by using student writing assignments. Faculty developed tools to assess achievement of critical thinking competencies, such as analysis, synthesis, insight, reflection, open mindedness, and depth, breadth, and appropriateness of clinical interventions. Faculty created a model for the development of program-specific critical thinking competencies, selected appropriate writing assignments that demonstrate critical thinking, and implemented a holistic assessment plan for data collection and analysis. Holistic assessment involves the identification of shared values and practices, and the use of concepts and language important to nursing. PMID- 12120116 TI - Colorectal cancer screening and surveillance: the present and the future. AB - Colorectal cancer screening is now proven to be effective and is widely endorsed by guidelines committees. Several forms of screening have merit, but the screening tests used in individual patients are often dictated by insurers. The use of screening colonoscopy is expected to increase. Gastroenterologists will play the central role in ensuring the safe and cost-effective application of colonoscopy to screening and surveillance. PMID- 12120117 TI - Dyspepsia, irritable bowel syndrome, and constipation: review and what's new. AB - Functional gastrointestinal disorders are collections of symptoms attributable to the gastrointestinal tract in the absence of mucosal, structural, or biochemical disease. The two most common disorders, irritable bowel syndrome and functional dyspepsia, have common etiopathogenetic features, notably psychosocial disturbances, dysmotility, heightened sensitivity, and, possibly, an association with a postinfective state. The third condition is constipation, in which transit disorders and abnormal evacuation represent disturbances of function that are amenable to therapy. This review is an update of the control mechanisms, pathophysiology, investigation, and potential pharmacotherapies of these disorders. Serotonergic and adrenergic agents and neurotrophic factors are among the novel approaches that may have a significant impact on these disorders. PMID- 12120118 TI - Esomeprazole magnesium (Nexium). AB - Esomeprazole, the S-isomer of omeprazole, is a new proton pump inhibitor. Esomeprazole provides better control of intragastric pH than omeprazole. It is more effective in treating erosive esophagitis in patients with GERD than omeprazole. Esomeprazole can maintain the healing of erosive esophagitis when used daily or on demand. It is also effective for the eradication of Helicobacter pylori infections. The incidence and type of adverse events associated with esomeprazole therapy are infrequent and likely to be similar to omeprazole. PMID- 12120119 TI - Advances in liver disease. Highlights from the 51st Annual Meeting of the American Association for the Study of Liver Diseases, October 27-31, 2000, Dallas, TX. PMID- 12120120 TI - The versatile chemistry of the [B20H18]2- ions: novel reactions and structural motifs. AB - Among the polyhedral [closo-BnHn]2- ion series (n = 5-12 inclusive) the aromatic [closo-B10H10]2- ion is both readily available and quite reactive. Among its many reactions which retain its cage structure one finds the oxidative dimerization reaction in which two [closo-B10H12]2- ions each formally lose a hydride ion and undergo dimerization of the resulting [closo-B10H9]- ions to produce the [trans B20H18]2- ion. The two-component [closo-B10H9]- ions of the latter are linked together by a pair of unique B-B-B bonds which provide unprecedented reactivity to the structure. Among these reactions are the two-electron reduction to a set of three interconvertible [B20H18]4- ions having intercage B-B bonds and the related reductive substitution reaction in which [trans-B20H18]2- undergoes attack by nucleophile, L, to produce [B20H18L]2-. The latter species is formally a substituted [B20H19]3- (L = H) ion formed by B-B bond protonation of one of the isomeric [B20H18]4- ions. These and a variety of novel reactions are described here along with interrelated reaction mechanisms considered for the first time. PMID- 12120121 TI - Electrospray ionisation mass spectrometric detection of weak non-covalent interactions in nogalamycin-DNA complexes. AB - We demonstrate the use of electrospray ionisation mass spectrometry (ESI-MS) in high salt solutions for the analysis of weak non-covalent complexes of the anthracycline antibiotic nogalamycin with novel DNA hairpin structures; high signal-to-noise ratios for the complexes in the absence of bound Na+ ions permits relative binding affinities to be estimated. PMID- 12120122 TI - Evidence of an equilibrium between selenides and osmium(VIII) reagents and selenoxides and osmium(VI) reagents. AB - Driving the equilibrium between selenides and osmium(VIII) reagents with selenoxides and osmium(VI) by a subsequent reaction (rearrangement of allyl selenoxides to allyl alcohols or addition of osmium(VIII) species on C=C double bonds) to one side, allows the transformation of methyl geranyl selenides to linalool and of methyl citronellyl selenoxide to 6,7-dihydroxy citronellyl selenide. PMID- 12120123 TI - First characterisation of rotational conformers in a chiral nitroxide by EPR spectroscopy. AB - The detection and characterisation in liquid solution by EPR spectroscopy of the rotational conformers of a nitroxide radical containing a chiral centre is reported for the first time. PMID- 12120124 TI - A new method for fluoride determination by using fluorophores and dyes anchored onto MCM-41. AB - A new colourimetric and fluorimetric method for fluoride determination in aqueous samples based on the specific reaction between fluoride and silica has been developed and applied on real samples. PMID- 12120125 TI - A single-source route to CdS nanorods. AB - We report the preparation of CdS nanorods using a thiosemicarbazide complex of cadmium [Cd(NH2CSNHNH2)2Cl2]. The precursor was decomposed in tri-n octylphosphine oxide (TOPO) at 280 degrees C to give TOPO capped CdS nanoparticles; nano-dimensional rods of the material are clearly visible in transmission electron microscopy (TEM); the particles have been further characterised by X-ray diffraction (XRD) and selected area electron diffraction (SAED) and optical measurements. PMID- 12120126 TI - Intermolecular recognition and crystal packing in molybdenum and tungsten coordination polymers as deduced from powder X-ray diffraction data. AB - The molecular conformations and packing of [(tBuCO2)3M2(mu-X)M2(O2CtBu)3], where M = Mo and W, and X = oxalate and perfluoroterephthalate, determined in the solid state from powder X-ray diffraction analysis, reveal one-dimensional coordination polymers involving pivalate-oxygen to metal interactions (X = perfluoroterephthalate), and oxalate--as well as pivalate-oxygen to metal bonds (X = oxalate), and allows explanation of the unusual state-dependent chromic properties of these compounds. PMID- 12120127 TI - Reaction of singlet oxygen with Ir(I) and Rh(I) thiolato complexes: oxidative addition vs. S-oxidation. AB - Singlet oxygen reacts with Ir(I) and Rh(I) thiolato complexes to form the corresponding Ir(III) and Rh(III) peroxo thiolato complexes which do not undergo intramolecular oxidation of the thiolate moiety. PMID- 12120128 TI - The first genuine observation of fluorescent mononuclear phthalocyanine aggregates. AB - An initial photophysical study of a tetra-solketal-substituted zinc phthalocyanine is reported; at low temperature this compound exhibits very strong aggregation, and a new red shifted emission peak is observed, lambda max approximately 750 nm, attributed to a fluorescent phthalocyanine dimer. PMID- 12120129 TI - Self-immobilized catalysts for ethylene polymerization: neutral, single-component salicylaldiminato phenyl nickel(II) complexes bearing allyl substituents. AB - A new family of self-immobilized ethylene polymerization catalysts, derived from neutral, single-component salicylaldiminato phenyl nickel complexes, is described. PMID- 12120130 TI - Methylidynetricobalt nonacarbonyl catalyzed cyclotrimerization of alkynes. AB - A cobalt carbonyl cluster, methylidynetricobalt nonacarbonyl, catalyzed inter- and intramolecular cyclotrimerization of alkynes producing substituted benzene derivatives in good to excellent yields. PMID- 12120131 TI - Hydroxy-substituted oligopyridine dicarboxamide helical foldamers. AB - As shown by X-ray diffraction and NMR studies, short oligopyridine dicarboxamides bearing benzyloxy, hydroxy and hydroxylate moieties adopt very robust single helical conformations, even in water. PMID- 12120132 TI - Selective functionalisation of TNT for sensitive detection by SERRS. AB - Selective chemical functionalisation of 2,4,6-trinitrotoluene to a surface enhanced resonance Raman active species for sensitive detection. PMID- 12120133 TI - Folding of aromatic oligoimides of trans-1,2-diaminocyclohexane. AB - Chiral oligomeric diimides prepared from pyromellitic dianhydride, (R,R)-1,2 diaminocyclohexane and phthalic anhydride fold into M or P helical conformers; trimer 1 folds into the P conformer in the crystal but the M conformer dominates in solution; longer chain oligomers 2 and 3 form preferentially P conformers in solution, as a result of intermolecular interactions. PMID- 12120134 TI - Reduction of alpha-aryloxy carbonyl compounds with samarium(II) iodide. A new traceless linker for the solid phase synthesis of carbonyl compounds. AB - A new linker for the solid phase synthesis of functionalised carbonyl compounds which is cleaved under mild, neutral conditions using samarium(II) iodide has been developed; the manipulation of an immobilised gamma-butyrolactone has been carried out to illustrate the utility of the linker. PMID- 12120135 TI - A new bifunctional catalyst for tandem Heck-asymmetric dihydroxylation of olefins. AB - A new bifunctional catalyst consisting of active palladium and osmium species anchored on silica gel through a mercaptopropyl spacer and a cinchona alkaloid respectively has been prepared for the first time and used in the heterogeneous tandem Heck-asymmetric dihydroxylation of olefins to afford diols with excellent yields and enantiomeric excesses (ee's) in presence of N-methylmorpholine N-oxide or K3Fe(CN)6 as cooxidants. PMID- 12120136 TI - A C-terminal domain of the membrane copper pump Ctr1 exchanges copper(I) with the copper chaperone Atx1. AB - A cloned C-terminal domain of the yeast high-affinity copper uptake pump Ctr1 exchanges Cu(I) rapidly with the yeast copper chaperone Atx1: 10(-2) < Kex < 10(+2). PMID- 12120137 TI - Construction of molecular wires based on a gold(I) bis-sigma-acetylide building block incorporated into ruthenium(II) polypyridyl complexes. AB - An Ru(II)-Au(I)-Ru(II) triad has been synthesized from [Ru(bpy)2(3 ethynylphenanthroline)]2+ with Au(tht)Cl and characterized by spectroscopic means such as NMR and ESI-MS; the Ru(II)-Au(I)-Ru(II) triad shows an intense emission at 620 nm upon excitation at 360 nm, which suggests an efficient energy transfer from the Au site to Ru sites via extended pi-conjugation through the ethynyl units. PMID- 12120138 TI - Aurophilicity as a cofactor in crystal engineering. Dicyanoaurate(I) anion as a building block in a novel Co(II)-Au(I) bimetallic assembly. AB - A 2D grid-shaped cyanide-bridged Co(II)-Au(I) bimetallic coordination polymer, [Co(DMF)2(Au(CN)2)2], has been prepared from the [Au(CN)2]- building block; sheets associate pair-wise by aurophilic interactions and the compound exhibits zeolite-like properties. PMID- 12120139 TI - Olefin self-metathesis as a new entry into xenotransplantation antagonists bearing the Galili antigen. AB - A hexameric disaccharide cluster bearing the terminal Gal alpha related xenotransplantation antigen was constructed using a sequence of ruthenium carbenoid catalyzed olefin self-metathesis of monoallylated tribenzyl pentaerythritol followed, after interconversion of benzyl ethers into para iodobenzyl ethers, by a single step Sonogashira cross-coupling of six prop-2-ynyl glycosides onto a hexameric aryl iodide scaffold. PMID- 12120140 TI - Zeolite-coated quartz fibers as media for photochemical and photophysical studies. AB - Zeolite-coated optical fibers are useful as media to carry out asymmetric photochemical reactions and for sensing polyaromatic compounds. PMID- 12120141 TI - Aurophilic complexes as gold atom sources in organic media. AB - The decomposition, either thermal or under H2, of [O(AuIPR3)3](CF3SO3) (R = Ph 1, R = Me 2) in organic solvents has been studied by 31P(1H) NMR, UV-vis spectroscopy and TEM; during the reaction, the phosphine acts as an efficient oxygen trap and gold nanoparticles are produced which may be stabilized by PVP in acetonitrile (mean diameter 4.5 nm) or oleylamine in toluene (mean diameter 9 nm). PMID- 12120142 TI - Synthesis and crystal structure of a novel decanuclear silver cluster complex [Ag(SC6H2Pri(3)-2,4,6)](10).2CHCI3.C2H5OH. AB - The cluster [Ag(SC6H2Pri(3)-2,4,6)]10 (1) contains a 20-membered ring of alternating silver and sulfur atoms, which is compactly folded; the sulfur atoms are doubly bridging and each silver atom exhibits linear two-coordination in the primary Ag-S interactions. PMID- 12120143 TI - Towards ruthenium(II) polypyridine complexes with prolonged and predetermined excited state lifetimes. AB - The excited state lifetime of a Ru(bpy)3-motif is linearly related to the number of appended pyrenyl chromophores, but independent of connectivity; values for nine complexes range from 0.8 to 18.1 microseconds. PMID- 12120145 TI - Nanotubes in Si-doped titanium dioxide. AB - Silicon-doped metal oxide nanotubes are formed in Si-doped titanium dioxide prepared by a sol-gel technique. PMID- 12120144 TI - Spectroscopic detection of short-lived anthracene derivative cation radicals using an electron transfer stopped-flow method with the tris(2,4 dibromophenyl)amine cation radical. AB - Dynamic transformation profiles of short-lived cation radicals of anthracene derivatives, including 1-methyl, 2-methyl and unsubstituted anthracenes, could be observed using an electron transfer stopped-flow method by adopting the tris(2,4 dibromophenyl)amine cation radical as a reaction initiator. PMID- 12120146 TI - A highly efficient titanium-based olefin polymerisation catalyst with a monoanionic iminoimidazolidide pi-donor ancillary ligand. AB - The titanium complex Cp[1,3-(2',6'-Me2C6H3)2(CH2N)2C=N]Ti(CH2Ph)2, with a monoanionic eta 1-iminoimidazolidide ancillary ligand, is shown to be a highly efficient catalyst for olefin polymerisation when activated with the Lewis acid B(C6F5)3. PMID- 12120147 TI - Oxidative coupling of platinum arylamides: temperature dependent C-H or C-F cleavage. AB - The oxidation of an arylamido platinum complex leads to C-C coupling with selective C-H versus C-F bond cleavage depending on the reaction temperature. PMID- 12120148 TI - Au2trien: a dinuclear gold(III) complex with unprecedented structura features. AB - The X-ray structure of a dinuclear gold(III) complex, Au2trien, shows the presence of two square planar gold(III) centers bridged by a nitrogen donor, in a very unusual fashion. PMID- 12120149 TI - An expedient synthesis of tetrakis(cyclopropylmethyl)methane. AB - Synthesis of tetrakis(cyclopropylmethyl)methane, a new symmetric product has been described using the radical mediated gem-diallylation of cyclopropylmethyl xanthate as a key step and its single crystal X-ray analysis established its C2 symmetry. PMID- 12120150 TI - Ultrasound promoted Suzuki cross-coupling reactions in ionic liquid at ambient conditions. AB - Palladium catalyzed Suzuki cross-coupling reactions of halobenzenes including chlorobenzenes with phenylboronic acid have been achieved at ambient temperature (30 degrees C) in the absence of a phosphine ligand using the ionic liquid 1,3-di n-butylimidazolium tetrafluoroborate [bbim][BF4] with methanol as co-solvent under ultrasonic irradiation. PMID- 12120151 TI - Engineered biosynthesis of novel spinosyns bearing altered deoxyhexose substituents. AB - Novel spinosyns have been prepared by biotransformation, using a genetically engineered strain of Saccharopolyspora erythraea, in which the beta-D-forosamine moiety in glycosidic linkage to the hydroxy group at C17 is replaced by alpha-L mycarose. PMID- 12120152 TI - Direct catalytic asymmetric aldol reactions of aldehydes. AB - The development of a direct catalytic enantioselective aldol reaction of aldehydes with activated carbonyl compounds catalyzed by chiral amines is presented and the potential demonstrated by the synthesis of optically active beta-hydroxycarboxylic acid derivatives. PMID- 12120153 TI - Remarkable co-catalysis by copper(I) oxide in the palladium catalyzed cross coupling of arylboronic acids with ethyl bromoacetate. AB - Copper(I) oxide can effectively co-catalyze the Suzuki type cross-coupling reactions of arylboronic acids with ethyl bromoacetate. As an alternative protocol for introducing the methylenecarboxy group into functionalized molecules, this reaction occurs in the absence of highly toxic thallium compounds or special ligands and should be convenient and practical. PMID- 12120155 TI - One-step heterogeneously catalytic oxidation of o-cresol by oxygen to salicylaldehyde. AB - Salicylaldehyde (selectivity = 57.3% at a conversion = 73.8%) was prepared for the first time by the oxidation of o-cresol in a single step using impregnated CuCo/C catalysts. PMID- 12120154 TI - A new strategy towards the total synthesis of phenanthridone alkaloids: synthesis of (+)-2,7-dideoxypancratistatin as a model study. AB - A new strategy towards the synthesis of phenanthridone alkaloids has been reported through the synthesis of (+)-2,7-dideoxypancratistatin from D-(-)-quinic acid employing PET initiated carbocyclization of an electron rich aromatics by silylenol ether as a key step. PMID- 12120156 TI - Formation and crystal structure of an unexpected inclusion complex of a metal free phthalocyanine and oxalic acid. AB - Treatment of 3-(2,4-dimethyl-3-pentyloxy)phthalonitrile (2) with CeCl3 in the presence of 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU) in n-pentanol gives the corresponding metal-free phthalocyanine 3, which unexpectedly traps the oxalic acid in the crystal lattice forming a 1:1 inclusion complex. PMID- 12120157 TI - The catalytic activity of alumina supported Ru nanoparticles for NO/CH4 reaction. AB - Alumina supported colloidal Ru nanoparticles, with an initial average size of 4.8 nm, show high activity for NO conversion for T > or = 450 degrees C and remarkably high selectivity (approximately 80%) to syngas at 600 degrees C. PMID- 12120159 TI - Formal syntheses of heliannuols A and D, allelochemicals from Helianthus annus. AB - A synthesis of heliannuol A 1 is described involving hydrogenolysis of the cyclopropane fused benzoxepane compound 23 to generate the benzoxocane ring system of 1 and a fragmentation of methyl ether 25 furnished 4-methoxycurcuphenol 29, an advanced intermediate to heliannuol D 4. PMID- 12120158 TI - Water-soluble aminoxyls (nitroxides): 2-methyl-2-[(N-(4-tert butylphenyl)oxyl]propanesulfonate and (1-oxyl-2,5,5-trimethylpyrrolidin-2 yl)methanesulfonate. AB - Two charged aminoxyls, ammonium (1-oxyl-2,5,5-trimethylpyrrolidin-2 yl)methanesulfonate and ammonium 2-methyl-2-[(N-(4-tert butylphenyl)oxyl]propanesulfonate, obtained by methods easily adaptable to the preparation of various other aminoxyls are totally soluble in water but are partially associated in other solvents. PMID- 12120160 TI - 1:1 complexes of 5-(4-[N-tert-butyl-N-aminoxyl]phenyl)pyrimidine with manganese(II) and copper(II) hexafluoroacetonylacetonate. AB - Mn(hfac)2 and Cu(hfac)2 form 1:1 complexes with 5-(4-[N-tert-butyl-N aminoxyl]phenyl)pyrimidine that exhibit strong metal-nitroxide exchange; spin polarization models do not explain the antiferromagnetic exchange behavior between spin sites in these complexes. PMID- 12120161 TI - Expression of the prohelicity of bis-cyclomanganated 2,3-diphenylquinoxaline through reactions with diaryldiazomethanes. AB - The reactions of bis-cyclomanganated 2,3-diphenylquinoxaline with diazodiphenylmethane and 9-diazofluorene allowed the formation of a new oligomeric and dinuclear manganospiralene and the ready preparation of a pentacyclic helix comprising two (eta 5-fluorenyl)Mn(CO)3 fragments, whose helicity can be locked upon one-dimensional linear coordination to silver cation. PMID- 12120162 TI - Palladium-mediated cross-coupling reactions with supported reagents in supercritical carbon dioxide. AB - Commercially available polystyrene supported amine and phosphine resins facilitate palladium-mediated Heck and Suzuki reactions in supercritical carbon dioxide (scCO2). PMID- 12120163 TI - The aromaticity and Mobius characteristics of carbeno[8]heteroannulenes and triplet state annulenes. AB - Two types of annulene which may show significant Mobius aromatic character and bond and twist delocalisation are proposed; triplet states with 4n + 2 occupancy of the p pi array of atomic orbitals and a novel 8-pi carbeno[8]heteroannulene ring system 1 where the Huckel highly antiaromatic nature as a planar system can be attenuated or even reversed by the C2 symmetric Mobius distortion. PMID- 12120164 TI - Transmetallation of beta-allenyl silanes: efficient synthesis of dienyl chlorostannanes and chlorostibines. AB - The reaction of 1-trimethylsilylbuta-2,3-diene with tin tetrachloride, antimony trichloride or antimony pentachloride gave the corresponding buta-1,3-dien-2-yl halostannane or stibine derivatives; this ligand exchange was extended to other beta-allenylsilanes. PMID- 12120165 TI - Ni-complex-catalysed addition polymerisation of 2-phenyl-1-methylenecylopropane to afford a polymer with cyclopropylidene groups. AB - pi-Allyl-nickel complexes initiated addition polymerisation of 2-phenyl-1 methylenecyclopropane to give a polymer with three-membered rings; the formed polymer showed a high Tg and negligible thermal decomposition up to 300 degrees C. PMID- 12120166 TI - Pyrolysis and UV photoelectron spectroscopy of bicyclo[3.2.0]hept-6-en-2-one; preparation and detection of cyclohepta-2(Z),4(E)-dien-1-one. AB - Flash vacuum pyrolysis of bicyclo[3.2.0]hept-6-en-2-one (1) in the source chamber of a UV photoelectron (PE) spectrometer using a CW CO2 laser as a directed heat source facilitated an electrocyclic ring expansion to yield the transient species cyclohepta-2(Z),4(E)-dien-1-one (2), the PE spectrum of which was compared to that of an authentic sample of cyclohepta-2(Z),4(Z)-dien-1-one (4) and confirmed a conrotatory ring opening of 1 that obeys the Woodward-Hoffmann rules. PMID- 12120167 TI - Palladium-catalyzed cyclization of alkenyl beta-keto esters in the presence of chlorotrimethylsilane. AB - PdCl2(CH3CN)2 catalyzed the cyclization of alkenyl beta-keto esters in the presence of a stoichiometric amount of SiMe3Cl to form 2 carboalkoxycyclohexanones in good yield with excellent regioselectivity. PMID- 12120168 TI - Synthesis and structural characterization of imido-lanthanide complexes with a metal-nitrogen multiple bond. AB - Treatment of an amido-ytterbium complex with n-BuLi leads to the isolation and structural characterization of a mixed amido-imido-ytterbium or imido-ytterbium complex, respectively, depending upon the molar ratios of the reactants; the Yb-N distance and the linearity of the imido N atom strongly suggest the presence of a formal Yb=N multiple bond in these novel complexes. PMID- 12120169 TI - Unique crown thioether complexes of f elements: the crystal structure of U(III) and La(III) complexes of 1,4,7-trithiacyclononane. AB - The crystal structures of the first U(III) complex of a crown thioether and of its La(III) analog have been determined; a stronger M-S interaction is observed for U(III) with respect to La(III) and Ce(III) in solution and in the solid state. PMID- 12120170 TI - Liquid-crystalline fullerene-oligophenylenevinylene conjugates. AB - Functionalization of C60-oligophenylenevinylene derivatives with a cyanobiphenyl terminated dendromesogen leads to new donor-acceptor systems with liquid crystalline properties. PMID- 12120171 TI - Reductive complexation of cycloheptatrienes by iron pentacarbonyl and catalytic sodium borohydride. AB - Fe(CO)5 and a catalytic amount of sodium borohydride react with cycloheptatrienes in protic solvents to yield the corresponding tricarbonyl(eta 4-1,3-diene)iron complexes in a one-pot procedure, which has been found to be particularly efficient for the synthesis of the useful tricarbonyl(cyclo-heptadiene)iron complex. PMID- 12120172 TI - Synthesis, structure, and reactions of a nitroxyl complex of iridium(III), cis,trans-IrHCl2(NH=O)(PPh3)2. AB - Reaction of Ir(NO)(PPh3)3 with anhydrous HCl results in addition of 2 equivalents of HCl with formal protonation of the nitrosyl ligand, affording the unusual six co-ordinate nitroxyl complex cis,trans-IrHCl2(NH=O)(PPh3)2. PMID- 12120173 TI - Chemical studies of the radical scavenging mechanism of bisorbicillinol using the 1,1-diphenyl-2-picrylhydrazyl radical. AB - A potent antioxidant, bisorbicillinol, which is a member of the bisorbicillinoid family isolated from the culture broth of Trichoderma sp. USF-2690, produces a stable radical-terminated symmetric dimer by donating two hydrogen atoms to the 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical. PMID- 12120174 TI - High-selectivity, high-flexibility glass hollow-fiber membrane for gas separation. AB - A high gas selectivity, high flexibility glass hollow-fiber membrane based on spinodal phase separation has been prepared by direct winding from glass melt, followed by acid leaching processing. PMID- 12120175 TI - Best of DDW 2001. Highlights from the 2001 Digestive Disease Week. May 20-23, 2001, Atlanta, GA. PMID- 12120176 TI - Current treatment for chronic hepatitis C. AB - Treatment of chronic hepatitis C has seen phenomenal progress over the last 10 years. The short courses of interferon monotherapy that were used in the early 1990s led to sustained improvement in liver disease and durable loss of detectable virus in fewer than 10% of patients. Longer courses improved the durability of those responses slightly. The major recent advance in treatment is the addition of the nucleoside analogue ribavirin to the interferon regimen. Combination of these two drugs for 6 to 12 months results in a sustained virologic response in 30% to 40% of previously untreated patients. Long-acting pegylated interferon has just been approved by the FDA and promises to improve the ease of administration. Furthermore, when used in combination with ribavirin, pegylated interferon increases the sustained virologic response rate to more than 50%. PMID- 12120177 TI - Training and competence in gastrointestinal endoscopy. AB - Patients, physicians, and health care providers want assurances that individuals performing gastrointestinal endoscopic procedures are competent and adequately credentialed. Definition of competence, however, has been an elusive goal. Most organizations, including professional societies and hospital privileging committees, have relied on estimated numbers of procedures performed or subjective assessment by a proctor as a surrogate marker of competence. Increasingly, objective assessment of performance is recognized as important in determining competence. Recent data have shown that learning curves for trainees are substantially more gradual than generally thought, and that the number of procedures required to achieve basic technical proficiency is much higher. Emerging data demonstrate that there is substantial variation in outcomes of endoscopy in clinical practice, related in part to the prior training, subspecialty background, ongoing case volume, and the individual endoscopist. Outcome variations correlate with both technical success and complications. Strategies for assessing competence in trainees and those in practice include numbers of procedures performed, subjective or objective assessment by a proctor, and self-assessment by the trainee. In the future, it is hoped that computers will be increasingly used to document outcomes of endoscopy in training and clinical practice as a part of routine report generation. PMID- 12120178 TI - Pegylated interferons. AB - Interferon therapy for chronic hepatitis C is not a cure, but it is able to decrease the viral load and may decrease the risk of complications (e.g., cirrhosis, liver failure, liver cancer). Pegylation of the interferon increases the amount of time the interferon remains in the body by increasing the size of the interferon molecule. Increasing molecule size slows the absorption, prolongs the half-life, and decreases the rate of interferon clearance. Thus the duration of biological activity is increased with pegylated interferon over nonpegylated interferon. The peginterferon alfa products offer an advantage over nonpegylated interferon alfa products because of less frequent administration. Tolerability of the pegylated interferons is comparable to the nonpegylated formulations. Monotherapy with these agents produces a better response in some patients than monotherapy with the nonpegylated formulation. Combination therapy with ribavirin is more effective than monotherapy. Studies comparing peginterferon alfa-2b and peginterferon alfa-2a in the treatment of chronic hepatitis C have not been performed. PMID- 12120179 TI - One hundred years of NSAID gastropathy: are coxibs the answer? AB - One hundred years after the introduction of aspirin, greater understanding of the mechanism of action of NSAIDs has led to the development of selective COX-2 inhibitors. These have been shown to reduce pain and inflammation with reduced risk of GI complications. However, questions remain regarding such issues as restriction of their use to patients at high risk for complications, cost effectiveness, effectiveness compared with prostaglandin replacement or acid reduction therapy, and safety in patients also taking aspirin for platelet inhibition. PMID- 12120180 TI - Gastroesophageal reflux disease--state of the art. AB - Gastroesophageal reflux disease (GERD) is defined as chronic symptoms or mucosal damage produced by the abnormal reflux of gastric contents into the esophagus; heartburn, its most common manifestation, occurs in 7% to 10% of the U.S. population on a daily basis. In addition, many so-called extraesophageal or atypical symptoms, including chronic cough, laryngitis and other otolaryngologic conditions, asthma, and unexplained chest pain, can be associated with GERD, but these patients appear to have a decreased frequency of heartburn, making the diagnosis of GERD difficult. All patients can be successfully managed with appropriate, titrated use of pharmacologic therapy. Antireflux surgery should thus be considered as an option only for patients who cannot afford or choose not to continue long-term medical therapy and for the rare patient with side effects or resistance to proton pump inhibitors. Endoscopic therapy for reflex should be considered as an experimental technology needing continuing evaluation. PMID- 12120181 TI - Dyspepsia, non-ulcer dyspepsia, and Helicobacter pylori. AB - Since the discovery of Helicobacter pylori and its role in peptic ulcer disease, two strategies for low-cost treatment of uninvestigated dyspepsia have emerged: "test and treat" and "test and scope." The efficacy of these strategies is examined, with a review of several recent studies. Controversy exists as to the usefulness of eradication of H. pylori in the treatment of non-ulcer dyspepsia. Results of four large trials are presented as an illustration of this controversy. Guidelines for the management of dyspepsia and non-ulcer dyspepsia are reviewed. PMID- 12120182 TI - Budesonide modified-release capsules. AB - Budesonide modified-release capsule is an effective form of therapy for the treatment of Crohn's disease located in the distal ileum, ileocecal region, and ascending colon. Because some of the benefit of budesonide therapy results from local effects, this agent will not be very effective in the treatment of patients with extensive colitis or left-side colitis. Budesonide is equal to less effective than prednisolone or prednisone therapy in the treatment of active Crohn's disease, but is associated with fewer glucocorticoids adverse reactions. PMID- 12120183 TI - Update on medical management of inflammatory bowel disease: ulcerative colitis. AB - Significant advances have been made regarding our understanding of the etiopathogenesis of inflammatory bowel disease. This review focuses on the most recent applications of medical therapy for ulcerative colitis. Therapeutic approaches continue to evolve regarding inductive and maintenance strategies with oral and topical aminosalicylates, systemic and non-systemic corticosteroids, and cyclosporine and alternative immunomodulators. As further investigations continue to discern microbiological and immunoinflammatory targets, future therapies may include both probiotics and novel biological agents. PMID- 12120184 TI - Emerging foodborne pathogens: keeping your patients and your families safe. AB - Changes in food production and societal pressures have led to a continuing increase in the incidence of foodborne illness. Many pathogens are associated with specific foods, e.g., E. coli O157:H7 with hamburgers or Salmonella with eggs. The U.S. Food and Drug Administration has recently approved irradiation for sterilization of meat, but public acceptance of irradiated food is low. Because contaminated foods are seldom detected before they reach store shelves, care in food preparation by professional and home cooks is crucial. PMID- 12120186 TI - Report from the ACG. Highlights from the American College of Gastroenterology. 66th Annual Scientific Meeting. October 19-23, 2001, Las Vegas, NV. PMID- 12120185 TI - Tegaserod for the treatment of constipation-predominant irritable bowel syndrome. AB - Tegaserod, a potent, partial serotonin 4 receptor (5-HT4) agonist, is an effective agent for the treatment of females with constipation-predominant irritable bowel syndrome. Tegaserod enhances gastric motility, stimulates peristaltic reflux and intestinal secretion, inhibits visceral sensitivity, and/or shortens colonic transit time. This agent may help women who have failed to respond to diet and exercise, laxatives, and other forms of therapy. The optimal dose of tegaserod is 6 mg twice daily and results in decreased number of days per month with pain, bloating, and days without bowel movements. Tegaserod is less effective in males than females in the treatment of constipation predominant irritable bowel syndrome. Tegaserod is well tolerated. Diarrhea is the most frequent adverse effect. The diarrhea tends to occur most frequently during the first few months of therapy and decreases with continued administration. PMID- 12120187 TI - Effect of herbal medicine keishi-to (TJ-45) and its components on rat pancreatic acinar cell injuries in vivo and in vitro. AB - BACKGROUND: In an attempt to clarify the mechanism of the effect of a herbal medicine, Saiko-keishi-to (TJ-10), which is a combination of Keishi-to (TJ-45) and Sho-saiko-to (TJ-9), we investigated the effects of these two herbal medicines and their components on pancreatic acinar cell injury models in vivo and in vitro. METHODS: Four-week-old male WBN/Kob rats were fed an MB-3 pellet diet containing herbal medicine (TJ-9, TJ-10 and TJ-45). Expressions of pancreatitis-associated protein (PAP) and manganese superoxide dismutase (Mn-SOD) were analyzed with a reverse transcription-polymerase chain reaction (RT-PCR) and immunohistochemistry. The herbal medicines and two of their components, Keihi (Cinnamomi cortex) and Shakuyaku (Paeoniae radix alba), were tested in vitro using an arginine-treated rat pancreatic acinar AR4-2J cell injury model. The inducible nitric oxide synthase (iNOS) was assayed in in vitro experiments. RESULTS: TJ-45-treated WBN/Kob rats showed no evidence of pancreatitis whereas there were pathological changes of chronic pancreatitis in TJ-9-treated WBN/Kob rats. PAP was not expressed and Mn-SOD expression was increased in the TJ-10-, and TJ-45-treated rats. The herbal medicines and two components suppressed PAP mRNA expression and enhanced Mn-SOD and iNOS mRNA expression in arginine-treated AR4-2J cells. CONCLUSION: These results suggest that the herbal medicine TJ-45 is effective for chronic pancreatitis caused by pancreatic ischemia. PMID- 12120189 TI - Co-existent chronic pancreatitis and pancreatic neuroendocrine tumor. Case report and review of the literature. AB - BACKGROUND: Few reports exist in the literature regarding neuroendocrine tumors either presenting as, or associated with, chronic pancreatitis. We report a case of chronic pancreatitis with a coexisting neuroendocrine tumor (gastrinoma) of the body of the pancreas. The available literature is reviewed. METHODS: Patient data including history, surgical procedure, histology and radiology investigations were collected and summarized. A Medline search using the key words 'pancreatitis' and 'neuroendocrine tumors' was performed for the years 1966 1999. Cited references in the relevant papers not listed in Medline databases were also evaluated. RESULTS: A 64-year-old female patient was operated on for unclear cystic lesions in the head and tail of the pancreas. Intraoperatively, a gastrinoma was incidentally discovered in the body of the pancreas. It did not appear to be obstructing the main pancreatic duct. The patient underwent a distal pancreatectomy with pancreatico-jejunostomy. Four months postoperatively, she is doing well with no signs of tumor recurrence. The Medline search revealed 125 publications, of which only 17 dealt with either acute or chronic pancreatitis associated with neuroendocrine tumors. When all available data were included, there were 26 cases of neuroendocrine tumors associated with acute pancreatitis. Additionally, 11 cases were associated with chronic pancreatitis, of which only 3 appear to be merely coexistent with chronic pancreatitis without an apparent cause-and-effect relationship between these two entities. CONCLUSION: We report the rare co-existence of chronic pancreatitis and a neuroendocrine tumor (gastrinoma) of the pancreas. The cause-and-effect relationship between neuroendocrine tumors of the pancreas and chronic pancreatitis continues to be uncertain. However, when the etiology of chronic pancreatitis is unclear, rare neuroendocrine tumors of the pancreas might be considered. Questions remain with regard to the potential role of chronic pancreatitis in the pathogenesis of pancreatic neuroendocrine tumors. PMID- 12120188 TI - Abnormal differentiation of islet cells in pancreatic cancer. AB - Pancreatic cancer in many patients is associated with altered glucose metabolism and abnormalities in pancreatic islet hormones at serum and tissue levels. Our previous studies have indicated a tendency of islet cells to differentiate toward ductal cell lineage, but the specificity of these findings for pancreatic cancer was not investigated. In the present study, we examined the immunoreactivity of pancreatic islets to antibodies against tumor-associated antigens DU-PAN-2, TAG 72 and CA19-9 in tissues from the normal pancreas, chronic pancreatitis and pancreatic cancer. Although no immunoreactive islet cells were found in the 12 normal pancreases and 20 chronic pancreatitis patients, 25 of 37 pancreatic cancer tissues showed the expression of these antigens, primarily CA19-9 and TAG 72, where the number of immunoreactive cells varied considerably from case to case. In 4 cases over 50% and in 2 of them more than 75% of the islets showed positive staining of 60-70% of islet cells within each islet. The presence of intrainsular ductular structures expressing the same antigen as the surrounding islet cells suggested transformation of antigen expressing islet cells to ductal cells. All but four islets were within or around the cancer favoring the notion that factors produced by cancer cells are responsible for the altered islet cell differentiation. PMID- 12120190 TI - Solid pseudopapillary neoplasm of the pancreas: report of a case after a 10-year follow-up and review of the literature. AB - A solid pseudopapillary neoplasm (SPN) is an extremely rare tumour of the pancreas that frequently occurs in young females and is mostly benign. SPN is a low-grade malignant tumour that may evolve years before symptoms start. However, the pathogenesis of this tumour remains unclear and there are no adequate reports of long-term results to evaluate the management and the long-term surgical control. We describe a new case of SPN with a 10-year follow-up, and review the world literature that accounts for approximately 322 cases. Moreover, a review of the current management and surgical tendencies in the treatment of SPN is considered. An SPN pancreatic tumour occurred in a 24-year-old female who complained of episodic mild abdominal pain sustained by a palpable epigastric mass. The tumour mass was detected by ultrasound and computer tomography and was localised at the tail of the pancreas adherent to the spleen. The preoperative diagnosis was uncertain and en-block distal pancreatectomy and splenectomy were performed. The size of the mass which weighed 300 g was 11 x 12 x 8 cm, and the tumour was strictly adherent and invaded the splenic hilum. Histologic examination confirmed a complete resection of the primary SPN that locally invaded spleen. The postoperative period was uneventful and after a 10-year follow-up the patient is free of symptoms. SPN should be considered in the differential diagnosis of large pancreatic masses, especially in young females. Radical resection, where technically feasible, should be considered the therapy of choice as it is a safe and effective control of the disease. PMID- 12120191 TI - Bacterial infection of pancreatic necrosis: role of bacterial translocation, impact of antibiotic treatment. PMID- 12120192 TI - Body composition, muscle function and psychological changes in patients undergoing operation for hepatic or pancreatic disease. AB - BACKGROUND: There is currently a dearth of data with respect to changes in body composition, physiological function and pSychological state in patients undergoing operative treatment for pancreatic or hepatic disease although marked changes in these variables have been reported in colorectal surgical patients. METHODS: In 36 patients (37 operations) we have studied the effect of a pancreatic or hepatic operation (with and without nutritional support) on body fat and body protein (assessed by Dual energy X-ray absorptiometry (DEXA) and anthropometry), respiratory function (measured by spirometry and vitalography), voluntary muscle function (measured by hand dynamometry) and psychological state (measured by use of the hospital anxiety and depression score and visual analogue scale for fatigue) over a 1-week period postoperatively. RESULTS: On the 3rd postoperative day there were significant changes in: grip strength 307 (135-499) to 249 (85-461) N; FEV1 2.28 (0.48-3.98) to 1.02 (0-2.42) litres/min; FVC 2.90 (0.75-5.02) to 1.28 (0.22-3.31) litres; anxiety score 7 (0-17) to 6 (1-20); depression score 3 (0-10) to 5 (0-20), and fatigue 3.9 (0.4-10) to 6.8 (1.0-9.7). These persisted on day 7 by which time mid-arm circumference and total body fat (by DEXA) had fallen from 30.1 (21.1-45.0) to 29.5 (20.2-43.2) cm, and 20.7 (5.8 53.7) to 20.4 (6.6-53.5) kg, respectively. CONCLUSION: We conclude that operative treatment for pancreatic or hepatic disease has an adverse effect on body composition, physiological function and psychological state. PMID- 12120193 TI - Differential roles of endogenous nitric oxide on neural regulation of basal exocrine pancreatic secretion in intact and denervated pancreas. AB - BACKGROUND: Autonomic nerves and humoral factors regulate pancreatic secretion. Nerves containing nitric oxide (NO) synthase (NOS) are in close proximity and located within cholinergic, adrenergic and sensory nerve bundles. Yet, the interactive mechanisms between various nerve populations remain elusive. AIMS: To evaluate the role of endogenous NO in basal exocrine pancreatic secretion in the extrinsically denervated rat pancreas. METHODS: Male Sprague-Dawley rats were assigned to 2 groups of 11 animals. The first group of sham-operated animals served as controls. In the second group extrinsic pancreatic innervation was surgically interrupted. One week later, after selective catheterization of the celiac axis and the bile-pancreatic duct, the animals received intra-arterial infusions of NG-nitro-L-arginine (L-NNA; 0.48 mg/kg b.w./h) followed by intra arterial infusions of L-arginine (110 mg/kg b.w./h). Total protein and amylase were measured in bile-pancreatic secretions collected at 15-min intervals. RESULTS: In controls, total protein and amylase output showed a biphasic secretion pattern with an increase during L-NNA infusion followed by a decrease when the infusion ceased and further augmentation 1 h later. In denervated animals, L-NNA caused a sustained decrease in pancreatic secretion followed by an increase 1 h later. Infusion of L-arginine at the time of maximum decrease slowed the second phase of protein and amylase output in sham-operated rats, but accentuated the onset of secretion in denervated animals. CONCLUSION: Inhibition of endogenous NO release was shown to increase baseline secretion in the intact pancreas. Superposition of extrinsic denervation on neural NOS-blockade decreased basal exocrine secretion, indicating that intra-pancreatic NO release is regulated by extra-pancreatic nerves. PMID- 12120194 TI - Staging acute pancreatitis: where are we now? AB - Acute pancreatitis is a potentially lethal disease in about 20% of all the cases. Early identification of those patients with the severe type of the disease is of a great importance as intensive care treatment and other therapeutic manipulations can apply to alter the clinical course, and finally the outcome. Therefore, there is a need for precise criteria for severity prediction--that can be easily applied with high accuracy and sensitivity--very early after the onset of acute pancreatitis. Although no 'ideal' predictor exists so far, APACHE II score, C-reactive protein and the trypsinogen activation peptide, could be used in clinical practice. Other prognostic markers such as interleukin-6 and interleukin-8 could be useful in the near future, as soon as proper assays will be available. Furthermore, the development of logistic models, based on different parameters concerning the severity of the individual patient, is another option for the future. PMID- 12120195 TI - Pathogenesis of infection in pancreatic inflammatory disease. PMID- 12120196 TI - Infections in pancreatic inflammatory disease: clinical trials for antibiotic prophylaxis. AB - The clinical significance and incidence of the specific pancreatic infections during severe pancreatitis, such as infected pancreatic necrosis and pancreatic abscesses, is very well known in the literature. The up-to-date knowledge in both the microbiological and pharmacological field related to pancreatitis suggests a series of antibiotics which are potentially useful in the prophylaxis of pancreatic infections. Here we review the most important controlled clinical experiences reported up to now. PMID- 12120197 TI - Acute pancreatitis: threat of fungal infection. PMID- 12120198 TI - The enteral factor in pancreatic infection. PMID- 12120199 TI - Has blood glucose level measured on admission to hospital in a patient with acute pancreatitis any prognostic value? AB - BACKGROUND: Early detection of pancreatic necrosis allows better management of the disease. Contrast-enhanced computed tomography (CT) as the gold standard for detecting pancreatic necrosis is expensive. AIM OF THE STUDY: This study was to evaluate for the first time whether blood glucose estimation on hospital admission--a simple, cheap, readily available laboratory parameter--may detect pancreatic necrosis and have prognostic value in acute pancreatitis. METHODS: Single blood glucose estimation upon hospital admission was evaluated prospectively for detecting pancreatic necrosis and as a prognostic indicator. The study included 241 nondiabetic patients with a first attack of acute pancreatitis. All underwent CT within 72 h of admission. RESULTS: High blood glucose (> 125 mg/dl) correlated significantly with complex high clinical and biochemical prognostic scores (Ranson, Imrie), a high Balthazar score, pancreatic pseudocysts, and a long hospital stay, but not with organ failure, indication for artificial ventilation, dialysis, surgery, length of intensive care, and mortality. Pancreatic necrosis detection sensitivity of high blood glucose was 83%, specificity 49%, positive predictive value 28%, and negative predictive value 92%. CONCLUSION: A patient with normal blood glucose on admission is unlikely to have pancreatic necrosis. Contrast-enhanced CT would not be needed unless the patient fails to improve. PMID- 12120200 TI - Serum free fatty acid concentration in patients with acute pancreatitis. AB - BACKGROUND: The involvement of lipolytic enzymes and liberated fatty acids in ethiology of acute pancreatitis (AP) has been implicated. AIM: To analyze the level of FFA in the patients with AP in relation to severity of the disease. MATERIAL AND METHODS: The study included 36 patients with acute edematous AP (group I), 29 patients with necrotizing AP: 16 without (group IIA) and 13 with complications (group IIB), and 12 control subjects. Serum levels of total FFA (by enzymatic method) and the individual fatty acids of the FFA pool (by gas-liquid chromatography) were measured during the first 4 days after admission. RESULTS: A significant increase in the mean total serum FFA was noted for all the groups with the highest values on admission (p < 0.02-0.01). The per cent contribution was significantly higher as compared to control group for oleic acid (group I, p < 0.02, group IIA, p < 0.05, group IIB, p < 0.005), linoleic acid (group IIB, p < 0.02) and arachidonic acid (group IIA, p < 0.05, group IIB, p < 0.02). Significantly lower percentage was noted for stearic acid (all three groups: p < 0.01, p < 0.005, p < 0.01, respectively) and for palmitic acid (only group IIB: p < 0.005). The ratio of saturated to polyunsaturated fatty acids was significantly lower than in control group on each day of study for group IIB only (p < 0.005-p < 0.001). CONCLUSION: Polyunsaturated fatty acids, mainly linoleic and arachidonic, may be involved in the development of complications in acute pancreatitis. PMID- 12120201 TI - Fatal outcome in acute pancreatitis: its occurrence and early prediction. AB - BACKGROUND/AIMS: This study aims to determine predictability of death in acute pancreatitis at a secondary-care hospital in Germany. METHODS: This study is part of an ongoing study on the epidemiology of acute pancreatitis and covers 368 patients with a first attack of acute pancreatitis in Luneburg county from 1988 to 1999. Early and late mortality were defined as < or = 1 weeks and > 1 week after admission. The following parameters were used to establish on admission likelihood of death: admission within 24 h or later with an acute attack, abdominal tenderness, signs of peritonitis, amylase and lipase in serum, leukocytes, hematocrit, potassium, sodium, calcium, creatinine after rehydration, blood glucose, bilirubin, serum glutamate-oxalacetate transaminase (SGOT), serum lactate dehydrogenase (SLDH), arterial pO2, APACHE II score, Ranson and Imrie scores. RESULTS: Of the 368 patients 17 (5%) died, 7 early because of multiple organ failure and 10 late because of septic complications. Mortality rates in interstitial and necrotising pancreatitis were 3 and 17%, respectively. Only an elevated serum creatinine (> 2.0 mg/dl) and a blood glucose > 250 mg significantly correlated with mortality. Ranson and Imrie scores were also significantly correlated with mortality; however, they were not obtained on admission, but only after 48 h. In univariate analysis, APACHE II score > or = 6 on admission and lipase > 1,000 U/l on admission provided a high sensitivity and negative predictive value for early and late mortality patients. CONCLUSION: Approximately half of the deaths in acute pancreatitis occur because of multiple organ failure or septic complications. New approaches have to be found to counteract these severe complications. A fatal outcome may be predicted by simple laboratory parameters such as a high serum creatinine and blood glucose. An APACHE II score > or = 6 and a lipase level on admission > or = 1,000 U/l indicate severe pancreatitis. PMID- 12120202 TI - SPINK1/PSTI mutations are associated with tropical pancreatitis in Bangladesh. A preliminary report. AB - BACKGROUND/AIMS: Tropical pancreatitis (TP) refers to a severe type of idiopathic chronic pancreatitis that develops in children in tropical regions of Africa and southern Asia. Phenotypically TP is subdivided into fibrocalculous pancreatic diabetes (FCPD) and tropical calcific pancreatitis without diabetes mellitus (TCP). Recently an association was identified between idiopathic pancreatitis in the USA and Europe and mutations in the serine protease inhibitor, Kazal type 1 (SPINK1) gene (previously termed pancreatic secretory trypsin inhibitor, PSTI). Our aim was to determine if either form of TP has a genetic basis. METHODS: We studied 8 well-characterized patients from Bangladesh with FCPD, 4 with TCP and 4 controls without pancreatic disease. The entire SPINK1 gene was sequenced in these patients. RESULTS: We detected two disease-associated SPINK1 mutations (N34S/IVS1 - 37T > C and IVS3 + 2T > C) in 6 of 8 patients from Bangladesh with FCPD but not in 4 patients with TCP (p < 0.03) or 4 controls (p < 0.03). CONCLUSIONS: We conclude that SPINK1 mutations are associated with FCPD in Bangladesh. Since SPINK1 mutations in Europeans and North Americans are associated with idiopathic chronic pancreatitis that is phenotypically different from FCPD, we further conclude that mutated SPINK1 markedly increases the risk of developing a variety of pancreatic diseases possibly through a chronic elevation of active trypsin within the pancreas. PMID- 12120203 TI - Autoimmune pancreatitis: possibilities of CT characterization. AB - BACKGROUND: Pancreatitis is defined as an inflammatory disease of the pancreas, leading to morphological and pathological changes. Recently, an autoimmune pathogenesis of this disease has been proposed. This type of pancreatitis should be differentiated from other pancreatic diseases, since appropriate therapy is effective and morphological changes and pancreatic function can recover to normal levels. AIM OF THE STUDY: To assess the possibility of distinguishing autoimmune pancreatitis from other pancreatic diseases with an analogous clinical presentation on the basis of CT findings alone. METHODS: The CT images of 7 patients with proven autoimmune pancreatitis, along with those of 20 patients with other pancreatic diseases, but with an analogous clinical presentation, were retrospectively evaluated in a blinded fashion by 2 radiologists. In particular, the radiologists had to search for the typical signs of autoimmune pancreatitis. Discordant cases were further analyzed in the presence of a 3rd radiologist. The final diagnosis was acquired by means of a majority or overall consensus. The sensitivity, specificity, positive and negative predictive values of CT were calculated against each of the diseases (autoimmune pancreatitis; other pancreatic diseases), in order to evaluate the diagnostic value of the scan. RESULTS: After the consensus evaluation, the correct diagnosis was reached in 25/27 (92.5%) cases, with only 2/27 wrong diagnoses (autoimmune pancreatitis diagnosed as another pancreatic disease and vice versa). The sensitivity and specificity of CT against autoimmune pancreatitis were 86 and 95%, respectively. The positive and negative predictive values were 89 and 93%, respectively. CONCLUSION: Patients with autoimmune pancreatitis demonstrate imaging findings that enable the correct diagnosis by dynamic CT, even in the presence of nonspecific clinical symptoms. The precise classification of the disease is extremely important, since simple steroid therapy then represents the correct treatment, and leads to complete recovery. PMID- 12120204 TI - Detailed tissue expression of bcl-2, bax, bak and bcl-x in the normal human pancreas and in chronic pancreatitis, ampullary and pancreatic ductal adenocarcinomas. AB - BACKGROUND: The aim of this study was to evaluate expression of the bcl-2 family of apoptosis regulating proteins in normal and diseased human pancreatic tissues. METHOD: Expression of bcl-2, bax, bcl-x, bak and p53 was determined in formalin fixed paraffin wax-embedded archival specimens of normal pancreatic tissue (n = 7), chronic pancreatitis (n = 7), pancreatic ductal adenocarcinoma (n = 23) and ampullary cancer (n = 7) by immunohistochemistry using specific antibodies. RESULTS: In normal pancreas and chronic pancreatitis tissues, bcl-2, bax and bcl x were predominantly expressed in ductal epithelial cells while p53 was not detected. In pancreatic ductal adenocarcinoma and ampullary cancer, bcl-2 was not detected compared with expression seen in normal acini (p < 0.01), minor (p < 0.001) and major ducts (p < 0.01), bax expression was reduced with respect to minor ducts (p < 0.01) but no different from normal acini or major ducts. bak and bcl-x were more strongly expressed in malignant epithelia compared with acini and major ducts but reduced when compared with minor ducts (p < 0.01). Overexpression of p53 was identified in 11 (48%) of 23 pancreatic adenocarcinomas and 4 (57%) of 7 ampullary cancers. Differential survival of individual patients was predicted by the relative level of bcl-x expression but not bax or bak, such that strong expression of bcl-x was associated with a median postoperative survival of 171 days when compared with 912 days for diminished expression (p < 0.001) of bcl-x. CONCLUSION: Pancreatic and ampullary cancer are associated with absent bcl-2 expression. bax, bak and bcl-x expression was reduced compared with normal minor ducts whilst bak and bcl-x expression was increased when compared with major ducts. bcl-x expression correlates with survival following resection and may represent a potential prognosis marker. PMID- 12120205 TI - Wegener's granulomatosis with onset of acute pancreatitis and rapid progress. A case report. AB - Although Wegener's granulomatosis is a rare disorder, the clinical and histological characteristics are well known. However, Wegener's granulomatosis with the onset of acute pancreatitis has rarely been reported. We discuss the case of Wegener's granulomatosis in a 65-year-old man, presenting with acute pancreatitis and whose disease progressed rapidly. PMID- 12120206 TI - On the role of islet amyloid polypeptide in glucose intolerance and anorexia of pancreatic cancer. PMID- 12120207 TI - Cyclooxygenases and lipoxygenases as potential targets for treatment of pancreatic cancer. AB - Pancreatic adenocarcinoma is characterized by poor prognosis, late diagnosis and lack of response to conventional therapies. The incidence of this disease shows no sign of declining in the Western world. Thus, new targets need to be identified for pancreatic cancer treatment. In particular, new chemotherapeutic agents would be extremely beneficial for control of unresectable cancer and metastatic lesions as well as for prevention of this deadly disease. Mounting evidence suggests that both lipoxygenases (LOXs) and cyclooxygenases (COXs), the key enzymes for arachidonic acid metabolism, have a profound influence on the development and progression of several human cancers. Recent evidence suggests that both COX and LOX pathways are important in pancreatic cancer. Results from immunocytochemical, RT-PCR, and Western blotting studies have shown that COX, specifically COX-2, is upregulated in human pancreatic cancer cell lines as well as human pancreatic cancer tissues compared with normal ductal cells and normal pancreas specimens. Agents that block COX enzymes significantly inhibit pancreatic cancer growth both in vitro and in vivo, in parallel with induction of apoptosis. Expression of both 5-LOX and 12-LOX is also seen in pancreatic cancer, although compared to the expression of COX this has not been extensively investigated. Chemical inhibitors or antisense oligonucleotides that block either 5-LOX or 12-LOX cause marked inhibition of pancreatic cancer cell proliferation. On the other hand, LOX metabolites stimulate growth of the tumor cells and reverse LOX-inhibitor-induced growth inhibition, suggesting the specific role of LOX in regulating pancreatic cancer cell proliferation. Although questions still need to be answered, such as the underlying mechanisms for COX and LOX-induced growth inhibition, both COX and LOX pathways are potential targets for pancreatic cancer treatment and chemoprevention. COX and LOX enzyme inhibitors are available and have been shown to be relatively safe in the treatment of other diseases. PMID- 12120208 TI - Role of endogenous insulin in pancreatic secretion in rats. AB - BACKGROUND: Pancreatic exocrine tissue is influenced by islet hormones through the islet-acinar portal system. We investigated the role of endogenous insulin in pancreatic secretion in vivo in rats. METHODS: Anesthetized rats were prepared by cannulation of the pancreatic duct, and pure pancreatic juice was collected. Pancreatic secretion was stimulated by intravenous infusion of cholecystokinin (CCK) (0.06 microgram/kg/h) with or without intravenous infusion of glucose (0.25 and 0.5 g/kg/h). The effect of intravenous infusion of glucose (0.5 g/kg/h) on pancreatic secretion stimulated by intraduodenal infusion of casein (400 mg/h) was also studied. RESULTS: Intravenous infusion of glucose (0.5 g/kg/h) produced a significant elevation of plasma insulin and glucose levels. The basal pancreatic secretion was not influenced by glucose. Pancreatic secretion of juice volume, amylase, and trypsin in response to stimulation by CCK was significantly augmented by glucose (0.5 g/kg/h). Intravenous infusion of glucose also resulted in significant increases in casein-stimulated pancreatic juice volume, amylase and trypsin outputs. CONCLUSION: Endogenously released insulin plays an important role in potentiating pancreatic secretion stimulated by exogenous CCK and intraduodenal infusion of casein. PMID- 12120209 TI - Hemorrhagic complications of pancreatitis: radiologic evaluation with emphasis on CT imaging. AB - OBJECTIVE: To analyze and describe the incidence, pathophysiology, radiographic diagnosis and the initial management of hemorrhagic complications associated with pancreatitis. MATERIAL AND METHODS: Among 1,910 patients diagnosed of having pancreatitis in the last 10 years, 26 developed hemorrhagic complications (1.3%). These complications were detected from 2 months to 8 years after one or several episodes of pancreatitis with a mean of 2.3 years. Radiographic studies were reviewed and clinical management and outcome were recorded. RESULTS: Ten patients had CT evidence of pancreatic necrosis, 12 patients chronic pancreatitis, and 17 patients pancreatic pseudocysts. The cause of hemorrhage was bleeding pseudoaneurysm in 16 patients (61%), diffuse bleeding with pancreatic necrosis in 5 patients (19.5%) and hemorrhagic pseudocysts in 5 patients (19.5%). Intra abdominal hemorrhage developed in 21 patients and gastro-intestinal bleeding in 5 patients. Arterial embolization was attempted in 12 patients and was successful in 9 patients (75%). Surgery was used in 16 patients and the overall mortality rate was 11%. CONCLUSIONS: Hemorrhagic complications are rarely seen and are usually late sequelae of pancreatitis. They develop because of leaking or ruptured pseudoaneurysms, diffuse bleeding in pancreatic necrosis, and hemorrhagic pseudocysts. Early detection followed by angiography, embolization and/or surgery has decreased mortality rates. PMID- 12120210 TI - Antisense oligonucleotides specific to mutated K-ras genes inhibit invasiveness of human pancreatic cancer cell lines. AB - BACKGROUND/AIMS: Point mutations of the K-ras gene are detected in > 90% of human pancreatic cancers and may play an important role in tumorigenesis. However, correlations between mutant K-ras and the invasive activity of the tumor have remained unclarified. METHODS: 17-merphosphorothioate antisense oligonucleotides targeting K-ras point mutations were transfected into three kinds of human pancreatic cancer cell lines (MIAPaCa-2, PANC-1, and BxPC-3), and the invasive activity was investigated using an in vitro chemoinvasion assay. RESULTS: Antisense oligonucleotides strongly inhibited the invasive activity of the cell lines with mutant K-ras genes (MIAPaCa-2, PANC-1), but not in that with a wild type K-ras (BxPC-3). CONCLUSION: Antisense oligonucleotides specific to mutated K ras genes inhibited the invasiveness of human pancreatic cancer cell lines. Specific antisense therapy to the point mutation of K-ras might be a new anticancer strategy for pancreatic cancer. PMID- 12120211 TI - Neural hormonal regulation of exocrine pancreatic secretion. AB - Exocrine pancreatic secretion is regulated by hormone-hormonal and neural hormonal interactions involving several regulatory peptides and neurotransmitter from the gut, the pancreas and the vagus nerve. The roles of the gastrointestinal peptides including secretin, CCK, neurotensin, motilin, PYY and pancreatic islet hormones including insulin, pancreatic polypeptide and somatostatin have been established. Interactions among secretin, CCK and neurotensin produce synergistic stimulatory effect. Motilin modulates the cyclic pattern of pancreatic secretion while local insulin provides a permissive role for the action of secretin and CCK at physiological concentration. Somatostatin, PYY and pancreatic polypeptide are inhibitory regulators, acting either on the release of secretin and CCK or on the action of the two stimulatory hormones. The vagal afferent-efferent pathway mediates the actions of many of these regulatory peptides, particularly of secretin and CCK. Acetylcholine and nitric oxide are the neurotransmitters known to mediate the actions of secretin and CCK. Serotonin (5-HT) released from enterochromaffin cells in the intestinal mucosa and nerve terminals of the enteric nervous system and intrapancreatic nerves may be involved in both stimulatory and inhibitory mechanism through its various receptor subtypes. 5-HT also mediates the action of secretin and CCK. The regulatory roles of neuropeptides, PACP and GRP, are now established, whereas those of others are being uncovered. Pancreatic juice provides both positive and negative feedback regulation of pancreatic secretion through mediation of both secretin- and CCK releasing peptides. Three CCK-releasing peptides have been purified: monitor peptide from pancreatic juice, diazepam-binding inhibitor from porcine intestine, and luminal CCK-releasing factor from rat intestinal secretion. All have been shown to stimulate CCK release and pancreatic enzyme secretion. Pancreatic phospholipase A2 from pancreatic juice and intestinal secretion appears to function as a secretin-releasing peptide. However, the detailed map of neurohormonal regulatory pathways of exocrine pancreatic secretion is yet to be constructed. PMID- 12120212 TI - Insights into the molecular basis of ligand binding by the cholecystokinin receptor. AB - The receptor for the peptide hormone, cholecystokinin, is a G-protein-coupled receptor in the rhodopsin/beta-adrenergic receptor family. A number of methodological approaches have been utilized to gain insights into the molecular basis for natural peptide ligand binding and activation of this physiologically important receptor. Insights into this have come from sequence analysis, ligand and receptor structure-activity data, receptor mutagenesis, conformational analysis of ligand and receptor fragments, and photoaffinity labeling. In this work, we review the contributions of each of these complementary approaches and provide a current integrated view of the active complex of cholecystokinin bound to its receptor. PMID- 12120213 TI - Growth factors in pancreatic health and disease. AB - In this article the role of different growth factors and their receptors in the pathogenesis of chronic pancreatitis and pancreatic cancer is discussed. The expression of members of the epidermal growth factor family, the fibroblast growth factor family, the transforming growth factor-beta family, the platelet derived growth factor family, the nerve growth factor family, the insulin-like growth factor family and their signaling receptors is presented, and a correlation of the molecular data with clinical and pathological changes is performed. A number of these growth factors and their receptors are markedly overexpressed in chronic pancreatitis and pancreatic cancer. In chronic pancreatitis, overexpression of growth factors and their receptors contributes to tissue remodeling and fibrogenesis. In contrast to chronic pancreatitis, pancreatic cancer is associated with a variety of genetic alterations, including mutations in tumor suppressor genes and cell cycle regulators. In the presence of these genetic disturbances, enhanced expression of growth factors and their receptors contributes to cell proliferation and enhances the aggressiveness of pancreatic cancer cells. In summary, growth factors and their receptors are often altered in chronic pancreatitis and pancreatic cancer and contribute to various pathogenetic aspects in these disorders. PMID- 12120214 TI - From acinar cell damage to systemic inflammatory response: current concepts in pancreatitis. AB - Acute pancreatitis represents a local inflammatory disorder with severe systemic consequences. Significant progress in understanding the pathophysiology of acute pancreatitis has been achieved in recent years. However, there is no clear concept about initialization and propagation of the disease both in experimental models and in humans. Furthermore, reliable strategies to evaluate prognosis and perform therapy are still missing. The review focuses on mechanisms originating from acinar cells leading to a systemic inflammatory response in experimental pancreatitis. PMID- 12120215 TI - TGF alpha transgenic mice. A model of pancreatic cancer development. AB - Pancreatic cancer is a devastating disease with a fatal prognosis due to late diagnosis and resistance to radiation and chemotherapy. The average survival after diagnosis is still 3 to 8 months. In the last few years genetic alterations in cancer-causing genes have been identified in tumors and putative premalignant lesions using microdissection techniques. However, the functional consequence of these genetic alterations for pancreatic growth and differentiation is unknown. TGF alpha overexpressed in the pancreas causes the development of tubular structures and fibrosis. Mice older than one year develop ductal pancreatic cancer. Crossbreeding these mice with p53 knockout mice dramatically accelerated tumor development. Moreover, tumors developing in these mice show frequently biallelic deletion of the Ink4a locus or LOH of SMAD4. These mice represent the first model of pancreatic adenocarcinomas with genetic alterations as well as growth characteristics similar to the human disease. PMID- 12120216 TI - Pancreaticopleural fistula imaged with magnetic resonance pancreatography. PMID- 12120217 TI - Genetic testing for hereditary pancreatitis: guidelines for indications, counselling, consent and privacy issues. PMID- 12120218 TI - Pancreatic cancer in hereditary pancreatitis: consensus guidelines for prevention, screening and treatment. PMID- 12120219 TI - Third International Symposium on Inherited Diseases of the Pancreas. PMID- 12120220 TI - Gene mutations in children with chronic pancreatitis. AB - In the last few years, several genes have been identified as being associated with hereditary and idiopathic chronic pancreatitis (CP), i.e. PRSS1, CFTR and SPINK1. In this study, we investigated 164 unrelated children and adolescents with CP for mutations in disease-associated genes by direct DNA sequencing, SSCP, RFLP and melting curve analysis. In 15 patients, we detected a PRSS1 mutation (8 with A16V, 5 with R122H, 2 with N29I), and in 34 patients, a SPINK1 mutation (30 with N34S, 4 with others). SPINK1 mutations were predominantly found in patients without a family history (29/121). Ten patients were homozygous for N34S, SPINK1 mutations were most common in 'idiopathic' CP, whereas patients with 'hereditary' CP predominantly showed a PRSS1 mutation (R122H, N29I). In patients without a family history, the most common PRSS1 mutation was A16V (7/121). In conclusion, our data suggest that CP may be inherited in a dominant, recessive or multigenetic manner as a result of mutations in the above-mentioned or as yet unidentified genes. This challenges the concept of idiopathic CP as a nongenetic disorder and the differentiation between hereditary and idiopathic CP. Therefore, we propose to classify CP as either 'primary CP' (with or without a family history) or 'secondary CP' caused by toxic, metabolic or other factors. PMID- 12120222 TI - Hereditary pancreatitis in Japan: a review of pancreatitis-associated gene mutations. AB - Pancreatitis-associated gene mutations have been reported in patients with hereditary pancreatitis and idiopathic pancreatitis in the Caucasian population and involve the cationic trypsinogen gene, the pancreatic secretory trypsin inhibitor gene and the cystic fibrosis transmembrane conductance regulator gene. In the Japanese population, mutational screening analyses of these genes have shown several mutations. The present study reviews previous reports from Japan in order to evaluate the racial specificity of pancreatitis-associated gene mutations. PMID- 12120221 TI - Hereditary pancreatitis in North America: the Pittsburgh-Midwest Multi-Center Pancreatic Study Group Study. AB - BACKGROUND: Hereditary pancreatitis (HP) was defined on a clinical basis alone until the first cationic trypsinogen gene (PRSS1) mutation was discovered through the initial phase of the current Pittsburgh Midwest Multi-Center Pancreatic Study Group (MMPSG) HP study in 1996, making genetic testing available. AIM: To evaluate the regional distribution of HP in the United States, and to compare the study's gene mutation database with the pedigree databases to determine whether family history alone predicts the likelihood of detecting mutations in the cationic trypsinogen gene. METHODS: Probands of families with HP, familial pancreatitis and idiopathic chronic pancreatitis were recruited through referrals from MMPSG collaborating centers, other physicians and self-referral of patients who had learned of the study through the World Wide Web (www.pancreas.org). Pedigrees were constructed, detailed questionnaires were completed and a blood sample was drawn for each proband and participating family members. The birthplace and current location of each patient was recorded, DNA was analyzed for known mutations and the pattern of phenotype inheritance was determined from analysis of each pedigree. RESULTS: A total of 717 individuals were ascertained; 368 (51%) had clinical pancreatitis confirmed and the rest were primarily unaffected family members used for linkage studies. Forty-six clinically unaffected individuals were silent mutation carriers (11% of mutation-positive individuals). HP was most common in Minnesota, New York and the central mid Atlantic states plus Kentucky and Ohio. One hundred and fifteen of 150 kindreds fulfilled the strict definition of an HP family, and 60 (52%) had PRSS1 mutations. Of the families with a detected mutation, 11% did not fulfill the clinical definition of an HP kindred. CONCLUSIONS: The distribution of HP within the United States shows major regional differences. The etiology of HP can be identified in a small majority of HP families through genetic testing. However, family history alone is not a good predictor of finding a mutation in the cationic trypsinogen (PRSS1) gene. PMID- 12120223 TI - Acute and chronic pancreatitis in patients with inborn errors of metabolism. AB - Acute and chronic recurrent pancreatitis have been reported in patients with a variety of inborn errors of metabolism. Among these are hyperlipidaemias, various disorders of branched-chain amino acid degradation, homocystinuria, haemolytic disorders, acute intermittent porphyria and several amino acid transporter defects. Some of these disease entities are exceedingly rare. In most of these disorders, pancreatitis is not very common and, with the exception of lipoprotein lipase and apolipoprotein C-II deficiency, is neither the leading nor the clinically most distressing manifestation of the underlying metabolic defect. The majority of these syndromes are, however, inherited, and often entire kindreds are carriers of well-defined germline mutations that can, to varying degrees, be associated with pancreatitis. We have reviewed the clinical, biochemical and genetic characteristics of those inborn errors of metabolism because interesting information can be gained from the in regard to the pathophysiology of pancreatitis and because they need to be distinguished from other hereditary causes of the disease. PMID- 12120224 TI - SPINK1 mutations are associated with multiple phenotypes. AB - Mutations in the gene encoding for the pancreatic secretory trypsin inhibitor or serine protease inhibitor, Kazal type I (SPINK1) have been associated with different entities of chronic pancreatitis. While there is no doubt about the involvement of SPINK1 mutations in pancreatic inflammatory disease, much controversy has arisen regarding which alterations are associated with disease and what type of disease model should be applied when the SPINK1 gene is examined. This article presents the existing data on SPINK1 mutations in idiopathic chronic pancreatitis, familial pancreatitis, hereditary pancreatitis and tropical pancreatitis. The possible role of SPINK1 mutations and polymorphisms in pancreatic disease is discussed. PMID- 12120225 TI - The pathobiochemistry of hereditary pancreatitis: studies on recombinant human cationic trypsinogen. AB - BACKGROUND/AIMS: This study attempts to identify the biochemical alterations in human cationic trypsinogen and trypsin caused by the hereditary pancreatitis associated mutations Arg117-->His and Asn21-->Ile. METHODS: Recombinant wild-type and mutant human cationic trypsinogens were expressed in Escherichia coli and purified to homogeneity, and trypsin autolysis and trypsinogen autoactivation were characterized. RESULTS: Both mutations significantly enhanced the autoactivation of human cationic trypsinogen. In addition, the Arg117-->His mutation inhibited autocatalytic inactivation of trypsin, while the Asn21-->Ile mutation had no such effect. CONCLUSIONS: The findings support the notion that enhanced trypsinogen activation in the pancreas is the common initiating step in hereditary pancreatitis, whereas trypsin stabilization plays a role in cases associated with the Arg117-->His mutation. PMID- 12120226 TI - Hereditary pancreatic cancer. AB - Hereditary pancreatic cancer (PC) appears to be exceedingly heterogeneous, as evidenced by its association with a variety of integrally associated diverse cancers and/or differing mendelian inherited cancer syndromes, which include the Lynch syndrome II variant of hereditary nonpolyposis colorectal cancer, hereditary breast-ovarian cancer syndrome in families with the BRCA2 mutation, hereditary pancreatitis, Peutz-Jeghers polyposis and the familial atypical multiple-mole melanoma syndrome in families with the CDKN2A (p16) germline mutation. Because of this heterogeneity, we provide a conservative estimate that about 5% (1,460) of PC cases in the US annually are hereditary. Although this number is relatively small, members of hereditary PC families serve as excellent models for studying the etiology, natural history, biomarkers, pathogenesis, potential carcinogenic exposures and their perturbation of underlying genetic events, and treatment of PC. These individuals would benefit greatly from method(s) capable of detecting cancer at an early stage, and such knowledge would also be useful for improving the diagnosis of the much more common 'sporadic' form of PC. PMID- 12120227 TI - Gene-environment interactions in pancreatic cancer. AB - Exocrine pancreatic cancer remains a major cause of cancer death in Western populations. Despite many efforts, little is known about its etiology. Tobacco is the only established cause, although the proportion of cases of exocrine pancreatic cancer attributed to it is only 30%. A family history of pancreatic cancer accounts for 10% of the cases of this disease. A large proportion of cases are due to yet unrecognized factors. The combined contribution of genetic susceptibility and environmental factors has rarely been considered. A higher risk of exocrine pancreatic cancer has been observed for patients with hereditary pancreatitis who smoked. It has also been suggested that CFTR mutations and alcohol could interact in the development of exocrine pancreatic cancer. Common variants in a large number of genes could act as low-penetrance alleles. Little is known about their role as susceptibility markers for exocrine pancreatic cancer, except for metabolic enzymes. A few studies have assessed the association between polymorphisms in these genes and exocrine pancreatic cancer. Surprisingly, none of them observed an interaction with tobacco consumption. The reality may be more complex; exocrine pancreatic cancer is genetically heterogeneous and it could involve many somatic and heritable mutations. Gene gene interactions and endogenous factors, among others, can contribute to tumor development. Future epidemiological studies should consider all of these aspects together. PMID- 12120228 TI - Inherited pancreatic cancer: surveillance and treatment strategies for affected families. AB - BACKGROUND: Nearly 10% of pancreatic cancers are hereditary in origin, and in some individuals, the risk of pancreatic cancer approaches 50%. A number of defined syndromes can predispose families to pancreatic cancer, although many of the mechanisms that result in familial pancreatic cancers are unknown. This article reviews current knowledge regarding familial pancreatic cancers and highlights the rationale for screening and surveillance. Methods for screening and surveillance of these high-risk individuals are described that allow the detection of pancreatic dysplasia, the precursor to pancreatic cancer. We also describe a single-center experience with the management and surveillance of familial pancreatic cancer kindreds. METHODS: Thirty-five patients from 13 familial pancreatic cancer kindreds underwent screening and/or surveillance. Endoscopic ultrasound (EUS) is the initial test of choice. Endoscopic retrograde cholangiopancreatography (ERCP) is reserved for symptomatic individuals or to investigate abnormal findings on EUS. In the proper clinical setting, patients with abnormal findings on both EUS and ERCP are candidates for total pancreatectomy. RESULTS: Twelve of 35 patients were noted to have abnormal findings on EUS and ERCP. All of these individuals underwent pancreatectomy, 10 total and 2 partial. The patients who underwent partial pancreatectomy are currently awaiting resection of the pancreatic remnant. Histopathologic examination of all 12 specimens demonstrated pancreatic dysplasia (the precursor lesion to pancreatic cancer). These specimens had no evidence of pancreatic cancer; nor were any of the resected pancreata normal. Follow-up of the 35 high risk patients at present varies from 1 to 48 months, and none of the patients under surveillance have developed pancreatic cancer. CONCLUSION: The screening and surveillance of high-risk members of familial pancreatic cancer kindreds using EUS and ERCP is an effective method for identifying individuals with pancreatic dysplasia prior to the onset of invasive pancreatic cancer. The surveillance needs to be performed by a team of specialists who have experience in dealing with pancreatic cancer and its precursors. PMID- 12120229 TI - Molecular diagnosis of early pancreatic ductal adenocarcinoma in high-risk patients. AB - The prevalence of pancreatic cancer in the general population is too low--even in high-prevalence areas such as Northern Europe and North America (8-12 per 10(5) population)--relative to the diagnostic accuracy of present detection methods to permit primary screening in the asymptomatic adult population. The recognition that the lifetime risk of developing pancreatic cancer for patients with hereditary pancreatitis (HP) is extremely high (20% by the age of 60 years and 40% by the age of 70 years) poses considerable challenges and opportunities for secondary screening in those patients without any clinical features of pancreatic cancer. Even for secondary screening, the detection of cancer at a biological stage that would be amenable to cure by surgery (total pancreatectomy) still requires diagnostic modalities with a very high sensitivity and specificity. Conventional radiological imaging methods such as endoluminal ultrasound and endoscopic retrograde pancreatography, which have proved to be valuable in the early detection of early neoplastic lesions in patients with familial pancreatic cancer, may well be applicable to patients with HP but only in those without gross morphological features of chronic pancreatitis (other than parenchymal atrophy). Unfortunately, most cases of HP also have associated gross features of chronic pancreatitis that are likely to seriously undermine the diagnostic value of these conventional imaging modalities. Pre-malignant molecular changes can be detected in the pancreatic juice of patients. Thus, the application of molecular screening in patients with HP is potentially the most powerful method of detection of early pancreatic cancer. Although mutant (mt) K-ras can be detected in the pancreatic juice of most patients with pancreatic cancer, it is also present in patients with non-inherited chronic pancreatitis who do not progress to pancreatic cancer (at least in the short to medium term), as well as increasingly in the older population without pancreatic disease. Nevertheless, the presence of mt-K-ras may identify a genuinely higher-risk group, enabling additional diagnostic imaging and molecular resources to be focussed on such a group. What is clear is that prospective multi-centre studies, such as that being pursued by the European Registry of Hereditary Pancreatitis and Familial Pancreatic Cancer (EUROPAC), are essential for the development of an effective secondary screening programme for these patients. PMID- 12120231 TI - Pancreatic cancer: factors regulating tumor development, maintenance and metastasis. AB - Pancreatic cancer has one of the poorest prognoses of all gastrointestinal malignancies. Today, it is the fourth or fifth leading cause of cancer-related deaths in Western industrialized countries, and the incidence has been increasing throughout the past decades. Insensitivity to growth-inhibitory and apoptotic signals as well as self-sufficiency of growth-promoting factors are hallmarks of the pathogenesis of this malignancy. In pancreatic cancer, a variety of growth factors and their receptors are expressed at increased levels. For example, the concomitant presence of the epidermal growth factor (EGF) receptor and its ligand EGF is associated with enhanced tumor aggressiveness and shorter survival following tumor resection. Furthermore, a number of other growth factors and their receptors, such as nerve growth factor and its receptor, are overexpressed in pancreatic cancer and contribute to its malignant phenotype. Besides factors which directly promote cell proliferation, a variety of other factors such as galectins are upregulated, which influences the tumor environment and the invasiveness of pancreatic cancer cells. In addition, tumor suppressor genes such as KAI1 are expressed at reduced levels, thereby enhancing the ability of pancreatic cells to form metastases. A complex disturbance of factors is present in pancreatic cancer, resulting in a distinct growth advantage which clinically results in rapid tumor progression and poor patient survival. PMID- 12120230 TI - Update of familial pancreatic cancer in Germany. AB - BACKGROUND/AIMS: The prevalence of familial pancreatic cancer (FPC) and the characteristics of FPC have not yet been well investigated in the German population. Therefore, a German case collection for FPC was established in July 1999 to collect and evaluate data on FPC families. METHODS: The prevalence of pancreatic cancer (PC) as well as other tumours and diseases was studied in families with at least 2 first-degree relatives with histologically confirmed PC, and in families of patients with PC and a first-degree relative with malignant melanoma. All participating family members were genetically counselled and evaluated by a standardised questionnaire. RESULTS: In an 18-month period, 73 independent kindreds with potential FPC contacted the national case collection. So far, 20 kindreds have fulfilled the criteria for FPC and have undergone complete workups. Most families revealed an autosomal dominant pattern of inheritance. Twelve families revealed an isolated accumulation of PC. Importantly, in 8 of 20 (35%) families, additional tumour types such as melanoma, breast and prostate cancer occurred. CONCLUSION: The observed phenotypic heterogeneity indicates an association with predisposing tumour suppressor genes p16 and BRCA2 in up to 30% of FPC families. Mutation analysis of these candidate genes might lead to the identification of the predisposing gene defect in a proportion of FPC families. PMID- 12120232 TI - Timing and extent of surgical intervention in patients from hereditary pancreatic cancer kindreds. AB - Our knowledge of the molecular and genetic etiology of hereditary pancreatic cancer has expanded considerably and is steadily increasing. However, there are only a few hard data available regarding the clinical and surgical management of these patients. Surgery is currently performed when we detect dysplastic changes in the pancreas or when cancer is suspected. Of the available diagnostic modalities, endoscopic ultrasonography has proven so far to be the most useful for detecting dysplastic changes in the pancreases of patients from hereditary pancreatic cancer kindreds. It seems reasonable, once dysplasia has been diagnosed in a high-risk patient, to proceed to total pancreatectomy. The multifocal nature of dysplastic lesions precludes any type of operation that would leave behind pancreatic tissue. Currently, prophylactic whole-organ resection in the absence of premalignant lesions cannot be recommended since we do not know the exact risk for the development of cancer. PMID- 12120233 TI - Neonatal screening for cystic fibrosis: long-term clinical balance. AB - BACKGROUND/AIMS: Very few studies have been performed on the long-term clinical advantages of neonatal screening programs for cystic fibrosis (CF) and these have been inconclusive. This is a preliminary report of two observational cohort studies on this subject. METHODS: In the first study, CF patients born between 1973 and 1981 in northeastern Italy were split into 4 groups according to the modality of diagnosis: screening by meconium test (58 patients); meconium ileus (45 patients); symptoms and pancreatic insufficiency (PI; 75 patients), or symptoms and pancreatic sufficiency (PS; 19 patients). The patients were followed for up to 26 years by three CF centers sharing common treatment protocols. In the second study, two cohorts of CF patients born between 1983 and 1992 were compared. Patients from one cohort (126 patients) were born in the Veneto region, where a neonatal screening program had been established based on immunoreactive trypsinogen. Patients from the other cohort (152 patients) were born in Sicily, where an intensive program of early diagnosis by symptoms was implemented. The cohorts were comparable for CF incidence, CFTR genotypes, gender proportion and common treatment protocols. Statistical analyses were performed by Kaplan-Meier survival curves, a Cox proportional hazard model for survival and cross-sectional comparisons by 2-year periods for weight z score, height z score and body mass index. RESULTS: In the first study, the patients detected by newborn screening (PI) showed better survival and nutritional status compared to patients diagnosed through meconium ileus or symptom presentation with PI. PS patients diagnosed by symptoms showed the best outcome, but most of them had a mild genotype. In the second study, the Veneto cohort showed better outcome with regard to survival and nutritional status over 16 years of follow-up. CONCLUSIONS: Observational cohort studies cannot give definitive evidence of the clinical benefit of neonatal CF screening; however, data have been accumulated which strongly suggest a better clinical outcome for CF patients born in an area where a screening program is performed. PMID- 12120234 TI - CFTR and cationic trypsinogen mutations in idiopathic pancreatitis and neonatal hypertrypsinemia. AB - BACKGROUND/AIMS: The CFTR gene has been shown to be involved in sporadic idiopathic pancreatitis (IP) and neonatal hypertrypsinemia with normal sweat chloride test (NHNST). The cationic trypsinogen gene (Try4) is responsible for hereditary pancreatitis. The aim of the present study was to find a correlation between mutations in the two genes and the two phenotypes. METHODS: Analysis of some known gene mutations and complete gene screening by denaturing gradient gel electrophoresis and DNA sequencing were undertaken. Thirty-two sporadic IP patients were investigated for the CFTR study, while 13 sporadic IP patients plus 4 hereditary pancreatitis families (24 tested individuals) were examined for the Try4 study. Fifty neonates with NHNST were investigated for the study of both genes. RESULTS: CFTR mutations were more frequently observed in sporadic IP cases with a common cystic fibrosis mutation or borderline sweat chloride than in cases with a negative sweat test. Try4 mutations were found in 1 out of the 13 sporadic IP cases tested. CONCLUSIONS: The CFTR gene may be involved in IP and NHNST, while the Try4 gene may be involved in IP, but not in NHNST, in this limited series of observations. PMID- 12120235 TI - Shwachman-Diamond syndrome: clinical phenotypes. AB - The clinical phenotype of Shwachman-Diamond syndrome (SDS) is extremely heterogeneous, showing a wide range of abnormalities and symptoms. The main characteristics of the syndrome are exocrine pancreatic dysfunction, haematologic abnormality and growth retardation. At diagnosis, especially when made in infancy, symptoms of pancreatic insufficiency are always present. This condition could be considered as a transient pancreatic insufficiency. In fact, several studies have shown that, with advancing age, about 40-60% of patients become pancreatic sufficient. Observations on the evolution of pancreatic activity lead us to believe that the diagnosis of SDS must be considered even in the absence of signs and symptoms of pancreatic insufficiency. Intermittent neutropenia is the most common haematological finding in SDS, but more of the bone marrow cellular elements can be involved. In recent years, recombinant human granulocyte colony stimulating factor has been used in some SDS subjects with severe neutropenia and frequent infection. The major haematological problem in the disease is the appearance of acute myeloid leukaemia; however, its prevalence is difficult to establish. Growth retardation is a typical manifestation. Weight and length are deficient at birth and remain below normal over time. Some studies show that SDS patients present short stature rather than malnutrition and this would suggest an inherent growth problem. A broad spectrum of skeletal abnormalities has been found to be associated with this syndrome. Short ribs with broadened anterior ends and metaphyseal dyschondroplasia of the long bone are the most common findings. Elevated liver enzymes and hepatomegaly are present in the first years of life with subsequent improvement without complications. Developmental delay, learning disorders and attention deficit disorders are also reported. PMID- 12120236 TI - The genes in pancreatic carcinoma. PMID- 12120237 TI - Hereditary pancreatitis: a model for understanding the genetic basis of acute and chronic pancreatitis. AB - Progress in understanding pancreatic diseases has been limited by a number of factors. Primary problems include the absence of good animal models, and difficulty in understanding the origin of pancreatic disease since the disease is usually manifest by the progressive destruction of the gland itself. Beginning in 1995, our laboratory, with the support of the Midwest Multicenter Pancreatic Study Group, began investigating the genetic basis of hereditary pancreatitis. Utilization of information becoming available through the human genome project allowed us to map and identify the hereditary pancreatitis gene as cationic trypsinogen (PRSS1). Molecular modeling, and subsequent experimental evidence, has solved key elements of the mysteries surrounding the origin of acute pancreatitis and the progression of acute pancreatitis to chronic pancreatitis. The availability of new genetic information and genomic tools should produce a revolution in our understanding of pancreatic diseases. PMID- 12120238 TI - Pancreatic cancer genetics. AB - Cancer is a multi-stage process resulting from accumulation of genetic changes in the somatic DNA of normal cells. Although in the majority of cases the changes occur only in the cancer cells there is a small proportion of cancers where a germline mutation confers an increased risk for cancer. Cancer susceptibility genes have effects that range from high to low penetrance with a corresponding high to lower likelihood for cancer in the carriers. Pancreatic cancer-prone families have been identified and some of the germline mutations responsible elucidated. Germline mutations in the BRCA2, CDKN2A/p16, hMSH2, hMLH1, hPMS1, hPMS2, LKB1/STK1, and PRSS1 genes have been associated with increased risk for pancreatic cancer. The concept of screening high-risk groups for pancreatic cancer is emerging, preferably in specialised centres with a multidisciplinary team approach. PMID- 12120239 TI - Ethical and policy issues in genetic testing. PMID- 12120240 TI - Use of DNA arrays/microarrays in pancreatic research. AB - In recent years, enormous technical advances in experimental protocols as well as robotic and bioinformatic techniques have allowed DNA array/microarray technology to emerge as the leading technology in the field of functional, disease-related genome analysis. Multiple applications exist for DNA arrays/microarrays including comparative genomic analysis to identify chromosomal imbalances (Matrix-CGH), the study of mutations and genetic polymorphisms, and the study of gene expression (expression profiling). Expression profiling is the most widely used application of DNA array/microarray technology and allows to measure gene expression of thousands of genes simultaneously. The present review describes the basic principles of expression profiling analyses and outlines some applications in pancreatic cancer research. PMID- 12120241 TI - Lineage commitment and cellular differentiation in exocrine pancreas. AB - Exocrine pancreatic cell types comprise greater than 90% of parenchymal cell mass in the adult pancreas. However, the factors regulating differentiation of acinar and ductal epithelial cells remain incompletely characterized. Like pancreatic islet cells, acinar and ductal cells arise from pluripotent precursors within embryonic pancreatic epithelium. Recent studies have suggested that a common pool of pluripotent stem cells is responsible for generating both endocrine and exocrine cell types, and that specific signaling pathways regulate a critical balance between endocrine and exocrine lineage commitment. PMID- 12120242 TI - Early pancreatic cancer. AB - The results of surgical treatment for ordinary carcinoma of the pancreas, even now considered the only means for cure, have been dismal. In order to define early pancreatic cancer, aiming amelioration of surgical results, early pancreatic cancer has been seeked. It may be readily conceivable that the smaller the tumor size, the earlier the lesion. The relationship between tumor size and surgical results was reviewed from the literature, some of which included articles written in Japanese. Tumor size < or = 2 cm in diameter is not always an early cancer. Tumor < 1 cm could be an early cancer but does not definitely reveal long-term survival. An increase of pancreatic cancer in Japan may be strongly related with the increased elderly population. Small cystic lesions which develop in elderly persons seem to indicate carcinogenesis of ordinary ductal cancer of the pancreas. Carcinoma in situ may be an early pancreatic cancer. Early pancreatic cancer is defined as an intraductal adenocarcinoma without any invasion or with minimal invasion to the stroma, regardless of size or extent of the lesion. PMID- 12120243 TI - Pancreatic carcinoma: what is the best imaging test? PMID- 12120244 TI - Imaging pancreatic cancer: the role of multidetector CT with three-dimensional CT angiography. AB - Over the past two decades, there have been significant technical advancements in computed tomography (CT). This has allowed CT to remain the gold standard for the evaluation of pancreatic pathology despite the advent of other imaging modalities, including MRI, PET and endoscopic ultrasound. Initially, CT scanners could only obtain 10-mm-thick slices at a rate of 4 slices per minute. Today, the current state of the art is multidetector CT (MDCT) technology, which allows the entire pancreas to be imaged by 1-mm slices in under 20 s. In addition, these new scanners allow true volume acquisition. The resultant data sets can be displayed not only as axial slices but also as a three-dimensional (3D) volume. The detail of these reconstructions when performed with volume rendering and maximum intensity projection techniques allows a detailed vascular mapping with accuracy that may exceed classic angiography. The use of thin collimation and dual-phase acquisition also improves the detection of hepatic metastasis as well as other sites of extrapancreatic disease. This article reviews the current state of the art of pancreatic imaging with specific emphasis on the use of MDCT, volume acquisitions and 3D arterial- and venous-phase vascular mapping. The advantages of these techniques and their impact on diagnosis and patient management are also addressed. PMID- 12120245 TI - Accuracy of endoscopic ultrasound in diagnosing and staging pancreatic carcinoma. AB - BACKGROUND: The role of endosonography in diagnosing and staging pancreatic adenocarcinoma is evolving. The aim of this review is to present recently published material comparing the performance of endosonography relative to other imaging modalities when evaluating a patient with a suspected or known carcinoma of the pancreas. METHODS: Medline was searched using the terms 'endosonography' and 'pancreas neoplasms'. References from retrieved papers were reviewed to identify other reports. Emphasis was placed on peer-reviewed material published within the past 3 years that included comparison with other imaging modalities. RESULTS: Despite advances in cross-sectional imaging modalities, endosonography remains the most sensitive and specific method to identify pancreatic mass lesions. Resectability determination of pancreatic carcinoma is best done with dual-phase helical CT, although endosonography may have slightly improved accuracy for lymph node assessment. Endosonography-guided fine-needle aspiration biopsy has high sensitivity (93%) and specificity (100%) when employed in patients with masses in whom pancreatic cancer is suspected but prior biopsies are negative. CONCLUSIONS: Endosonography can aid in diagnosing patients with pancreatic neoplasms through definitive inclusion or exclusion of a mass lesion as well as biopsy confirmation of malignancy. The role of endosonography in determination of resectability has been eclipsed by dual-phase helical CT. PMID- 12120246 TI - Radiologic imagings of cystic neoplasms of the pancreas. AB - An increasing number of cystic neoplasms of the pancreas have been detected with the progress of radiologic modalities. These cystic tumors include solid pseudopapillary tumor, serous cystadenoma, mucinous cystic tumor, and intraductal papillary mucinous tumor. The latter two lesions have been reported under a clinical nick-name of 'mucin-hypersecreting' cystic tumor. In the diagnosis of cystic neoplasms of the pancreas, clinical findings such as age, sex and location of lesion were informative. US and CT were useful to check up the presence of cystic lesion of the pancreas. ERCP was useful to examine the cystic lesion but it has some limitations. ERCP did not visualize the cystic lesion when the lesion was filled with excessive mucin. MRI (T2-weighted image) and magnetic resonance cholangiopancreatography were powerful to display the entire cystic lesion. In the assessment of grade of malignancy, EUS was valuable to demonstrate the precise internal structures such as mural nodule. In this communication, radiologic diagnosis of cystic tumor of the pancreas was briefly reviewed focussing on 'mucin-hypersecreting' tumor of the pancreas. PMID- 12120247 TI - Role of endoscopic ultrasound in the diagnosis of cystic lesions of the pancreas. AB - Endoscopic ultrasound (EUS) is an ideal imaging technique for pancreatic cystic lesions. Ultrasound is exquisitely sensitive for detecting and characterizing cysts arising in solid organs, and when the transducer is placed on an endoscope, high resolution imaging of the pancreas is achieved. Linear EUS can also guide needle aspiration of pancreatic cystic lesions and through the use of aspiration cytology, cystic tumors of the pancreas can be diagnosed. Since cytology is a relatively insensitive test, cyst fluid tumor markers such as CEA have been employed to improve the sensitivity for the detection of malignancy. Cyst fluid CEA values are uniformly low in serous cystadenomas, higher in mucinous lesions, and markedly elevated in mucinous cystadenocarcinomas. Through the use of these techniques, the ability to detect and diagnose early malignancies of the pancreas will be greatly enhanced. PMID- 12120248 TI - Cystic neoplasms of the pancreas. AB - Cystic tumors of the pancreas have become increasingly prevalent. In large series, more than 90% of pancreatic cystic neoplasms are accounted for by mucinous cystadenomas and cystadenocarcinomas, serous cystadenomas, and intraductal papillary mucinous tumors. Differentiating cystic neoplasms from pseudocysts can almost always be accomplished by clinical and radiological means, but in doubtful cases, when observation is contemplated, or when it is important to determine preoperatively the type of cystic neoplasm, cyst fluid analysis is useful. This can be readily obtained by endoscopic ultrasound, and analysis of enzymes, viscosity, cytology and a variety of tumor markers allows for a better differential diagnosis. PMID- 12120249 TI - Cystic lesions and neoplasms of the pancreas. The features are becoming clearer. AB - This review focuses on the most common cystic pancreatic lesions and neoplasms (i.e. pseudocysts, intraductal papillary-mucinous neoplasms, mucinous cystic neoplasms, serous cystic neoplasms, solid pseudopapillary neoplasms and ductal adenocarcinomas with cystic features) and discusses their clinicopathological features. The progress made in recent years has made it possible to clearly distinguish between the various entities and to unravel some of the problems involved. PMID- 12120250 TI - Site-specific therapeutic effects of protease inhibitors: effect of route of administration in experimental pancreatitis. AB - Inappropriate local and systemic activation of trypsin arising from trypsinogen mediates key steps in the pathogenesis of acute pancreatitis. Trypsin presumably causes direct injury to cells, but also activates other proteases and causes secondary effects such as inducing the expression of adhesion molecules on endothelium and leukocytes, and stimulating leukocytes to secrete cytokines, tissue-damaging enzymes, oxygen radicals and matrix metalloproteinases. Protease inhibition interferes with this cascade of events. Here we review experimental studies on protease inhibition and their potential application and report our findings of site-specific therapeutic effects of the novel protease inhibitor nafamostat (FUT-175) in experimental acute pancreatitis. PMID- 12120251 TI - Continuous regional application of protease inhibitor in the treatment of acute pancreatitis. An experimental study using closed duodenal obstruction model in dogs. AB - In clinical settings, the effectiveness of protease inhibitors in the treatment of acute pancreatitis has been still controversial. With the concept that sufficient tissue concentration of protease inhibitor in the pancreas has to be included to achieve its potent inhibitory effect, we applied a continuous regional intra-arterial (CRI) application of low-molecular-weight protease inhibitor, nafamostat mesilate (FUT-175), for closed duodenal loop obstruction model in mongrel dogs. The use of CRI application led to a higher concentration of FUT-175 in the pancreatic tissue (4,453 +/- 758 ng/g) when compared with that applied intravenously (905 +/- 48 ng/g). Consequently, pancreatic parenchyma in CRI application animals was remarkably preserved, as assessed by the lower extent of pancreatic necrosis (12.4 +/- 2.6% in CRI vs. 25.6 +/- 1.9% in intravenous). Additionally, the elevation of trypsin-like activity in the pancreas was significantly inhibited in CRI animals. Based on these findings, the dose as well as the route of protease inhibitors should be carefully considered to achieve its beneficial effect. PMID- 12120253 TI - A return to patent good sense. PMID- 12120252 TI - Benefit of continuous regional arterial infusion of protease inhibitor and antibiotic in the management of acute necrotizing pancreatitis. AB - Although we have reported the beneficial effect of continuous regional arterial infusion (CRAI) of protease inhibitor and antibiotic on acute necrotizing pancreatitis (ANP), the optimal timing of the initiation of CRAI therapy has not been clarified. The present study was conducted to evaluate whether the difference of the timing of CRAI therapy may affect the clinical course and outcome in ANP. 73 patients with ANP were stratified into three groups according to the interval between the onset and initiation of CRAI therapy as follows: group I (32 patients in whom CRAI therapy was initiated within 48 h after the onset); group II (22 patients in whom CRAI therapy was initiated between 48 and 72 h after the onset), and group III (19 patients in whom CRAI was initiated more than 72 h after the onset). The mortality rate was 3.2% in group I, 9.1% in group II, and 26.3% in group III. The mortality rate was significantly low in group I compared with that in group III. The frequency of respiratory failure in group I was also significantly low compared with that in group III. CRP and APACHE II score were reduced rapidly in both groups I and II after the initiation of CRAI therapy. These results suggested that the optimal timing of CRAI therapy in ANP should be considered to be within 72 h after the onset. PMID- 12120254 TI - Supreme Court rules on Festo. PMID- 12120255 TI - Selective anticancer drugs. PMID- 12120256 TI - Glivec (STI571, imatinib), a rationally developed, targeted anticancer drug. AB - In the early 1980s, it became apparent that the work of pioneers such as Robert Weinberg, Mariano Barbacid and many others in identifying cancer-causing genes in humans was opening the door to a new era in anticancer research. Motivated by this, and by dissatisfaction with the limited efficacy and tolerability of available anticancer modalities, a drug discovery programme was initiated with the aim of rationally developing targeted anticancer therapies. Here, we describe how this programme led to the discovery and continuing development of Glivec (Gleevec in the United States), the first selective tyrosine-kinase inhibitor to be approved for the treatment of a cancer. PMID- 12120257 TI - Nucleic-acid therapeutics: basic principles and recent applications. AB - The sequencing of the human genome and the elucidation of many molecular pathways that are important in disease have provided unprecedented opportunities for the development of new therapeutics. The types of molecule in development are increasingly varied, and include antisense oligonucleotides and ribozymes. Antisense technology and catalytic nucleic-acid enzymes are important tools for blocking the expression of abnormal genes. One FDA-approved antisense drug is already in the clinic for the treatment of cytomegalovirus retinitis, and other nucleic-acid therapies are undergoing clinical trials. This article reviews different strategies for modulating gene expression, and discusses the successes and problems that are associated with this type of therapy. PMID- 12120258 TI - Optical biosensors in drug discovery. AB - Optical biosensors that exploit surface plasmon resonance, waveguides and resonant mirrors have been used widely over the past decade to analyse biomolecular interactions. These sensors allow the determination of the affinity and kinetics of a wide variety of molecular interactions in real time, without the need for a molecular tag or label. Advances in instrumentation and experimental design have led to the increasing application of optical biosensors in many areas of drug discovery, including target identification, ligand fishing, assay development, lead selection, early ADME and manufacturing quality control. This article reviews important advances in optical-biosensor instrumentation and applications, and also highlights some exciting developments, such as highly multiplexed optical-biosensor arrays. PMID- 12120259 TI - New horizons--alternative routes for insulin therapy. AB - Since the introduction of insulin therapy 80 years ago, the lives of millions of patients with diabetes have been saved, prolonged and immeasurably improved. However, restoring normal glucose levels in diabetic patients through administering insulin by subcutaneous injection has proved virtually impossible. The consequences for patients are serious complications, including diabetic retinopathy and nephropathy, which tend to result from persistent hyperglycaemia. Maximizing glucose control in diabetic patients requires several daily injections. In an effort to reduce this burden, alternative and less-intrusive routes for the administration of insulin are being explored. PMID- 12120260 TI - Genome-based pharmacogenetics and the pharmaceutical industry. AB - Pharmacogenetic capabilities have changed markedly since The SNP Consortium made a dense single-nucleotide polymorphism (SNP) map freely available in 2001. For more than 40 years, pharmacokinetics and pharmacodynamics of drug-metabolizing molecules were the focus of practical applications. Today, it is possible to use SNP-mapping technologies to create a genetic profile of each individual that can be used to identify patterns of susceptibility genes for common diseases as well as genetic risk/efficacy factors that are related to the effects of drugs. PMID- 12120261 TI - Virtual screening using grid computing: the screensaver project. PMID- 12120262 TI - Growth factors in development and diseases of the exocrine pancreas. PMID- 12120264 TI - Natural course of chronic pancreatitis. AB - Although the three leading symptoms of chronic pancreatitis, pain, exocrine and endocrine pancreatic insufficiency, are well known, only a few long-term studies have correlated these symptoms with the natural course of the disease. Besides these symptoms, numerous pancreatic complications and/or pancreatitis-associated diseases may affect the course and determine the prognosis of chronic pancreatitis. Their influence, however, has not been studied in detail. This review has two major aims: The first one is to give an up-to-date survey of present knowledge on the natural course of the disease with a view to the leading symptoms under special consideration of how the duration of the disease, continual alcohol abuse as well as endoscopic procedures and surgical treatment affect pain. Included is also what is known about chronic pancreatitis as a precondition of pancreatic and extrapancreatic carcinoma. The effect of chronic pancreatitis on the socio-economic status of patients is discussed and the mortality rate of the disease. The second aim of this review is to stimulate pancreatologists from different centers to consolidate resources in order to perform larger controlled studies than any one single center can undertake and work out common criteria for the diagnosis of chronic pancreatitis and follow-up of its course. PMID- 12120263 TI - Estradiol has a direct impact on the exocrine pancreas as demonstrated by enzyme and vigilin expression. AB - BACKGROUND: Estrogen receptors have been found in the exocrine pancreas; however, the exact role of estrogen in pancreatic enzyme synthesis and secretion remains to be elucidated. Vigilin, a multi-KH domain protein, is part of a tRNA containing ribonucleoprotein complex and may be a suitable marker for stimulation of the translational machinery. In the present study, we investigated the influence of estradiol and compared it to CCK on the expression of vigilin, trypsin and amylase in rat pancreatic acini. METHODS: Acini were isolated and incubated with CCK or estradiol. The change in amylase and trypsin levels in the medium and in cell extracts were determined using a photometric method. The change in vigilin mRNA and protein expression were determined by RT-PCR and Western blotting, respectively. RESULTS: Treatment of isolated exocrine pancreatic cells with estradiol caused stimulation of amylase and trypsin production and inhibition of secretion, while treatment with CCK showed only a minor effect on enzyme production and resulted mainly in a stimulation of secretion. Further we found an increase in vigilin mRNA and protein expression in acini stimulated with both CCK-8 and estradiol. CONCLUSION: Our data suggest that estradiol may play a role in inducing exocrine enzyme production but not secretion, and that vigilin, as a marker for translational activity, is stimulated in parallel to the pancreatic enzymes: amylase and trypsin. PMID- 12120265 TI - In vivo octreotide administration acutely reduces exocrine granule size in the human pancreas. AB - BACKGROUND: Octreotide has been found to be effective in the prevention of postoperative complications of pancreatic surgery, but the benefit of preoperative octreotide administration has not been assessed. AIMS: To evaluate the ability of octreotide in reducing the amount of digestive enzymes in the pancreas before surgery, a morphometric ultrastructural study of the gland was undertaken in patients undergoing demolitive pancreatic surgery. METHODS: Twenty three inpatients received saline (n = 8) or octreotide (100 micrograms s.c.) before surgery either once (n = 5), or three (n = 5) or six (n = 5) times at 8 hour intervals. At surgery, biopsies of the pancreas were taken and processed for electron microscopy. Several parameters were assessed in exocrine cells by means of ultrastructural morphometry. RESULTS: A single administration of octreotide significantly reduced the exocrine granule number and the mean and total granular surface sectional area, and the ratio between granule area and cytoplasmic area. Repeated octreotide administrations were associated with partial (3 administrations) and complete (6 administrations) recovery of all parameters to control values. CONCLUSION: Preoperative administration of octreotide, the synthetic analogue of somatostatin, acutely reduces exocrine granule number and size in the pancreatic cell. This finding can partially explain the prophylactic effect of the drug on early complications of pancreatic surgery. Such an effect is not maintained over multiple administrations of the somatostatin analogue. Possible explanations for this latter finding are discussed. PMID- 12120266 TI - Site-specific localization of protein kinase C isoforms in rat pancreas. AB - BACKGROUND: Protein kinase C (PKC), a major signal-transducing enzyme, is recognized to play an important role in the regulation of pancreatic exocrine and endocrine secretion, and yet the distribution of PKC isoforms in rat pancreas has remained unclarified. AIM OF THE STUDY: We examined the precise localization of PKC isoforms to elucidate the role of PKC in the normal rat pancreas. METHODS: Male Sprague-Dawley rats were used throughout the experiment. For Western blot analysis, the islet of Langerhans and the acinar tissue were separated by the collagenase digestion method. Also, the whole pancreas was taken out and immunohistochemistry performed. RESULTS: According to Western blot analysis, PKC alpha, -gamma, -delta, -epsilon, -zeta, and -lambda were detected in both acinar and islet cells while PKC-beta II were observed exclusively in the islet. PKC beta I was not observed. On immunohistochemistry, the immunoreactivities of PKC isoforms were observed as follows: PKC-alpha, weakly in some endocrine cells and ductal epithelium; PKC-beta II, mainly in the islet center; PKC-gamma, in the islet, intrapancreatic ganglia and ductal epithelium; PKC-delta, in the islet periphery, weakly in some acinar cells and ductal epithelium; PKC-epsilon, strongly in the islet, acinar cell and ductal epithelium; PKC-zeta, in the islet, acinar cell and ductal epithelium; PKC-lambda in some endocrine cells and ductal epithelium. CONCLUSION: These results suggest that the intrapancreatic site specific existence of PKC isoforms may regulate pancreatic exocrine and endocrine functions via a PKC-mediated signal transduction. PMID- 12120268 TI - Evaluation of morphology and microcirculation of the pancreas by ex vivo and in vivo reflectance confocal microscopy. AB - BACKGROUND: Near-infrared reflectance confocal microscopy (CM) provides non invasive real-time images of thin virtual horizontal tissue sections with high resolution and contrast. AIM: Aim of the study was to characterize morphology, microcirculation and leukocyte-endothelial interaction (LEI) in normal pancreas by in vivo and ex vivo CM. METHODS: For CM we used water immersion objective lenses of high numerical aperture and near-infrared wavelengths. Experimentally measured lateral optical resolution is 0.5-1 micron and the axial resolution is 3 5 microns. The maximum depth of resolution was 300-400 microns. For ex vivo imaging, freshly excised pancreatic tissue from rats was studied by reflectance CM and conventional histopathology. For in vivo CM, the pancreatic head was exteriorized on a specially constructed stage for imaging the microcirculation and LEI. Images were obtained in real time at rates of 30 frames/s and later analyzed off-line to evaluate LEI and functional capillary density (FCD). RESULTS: CM allowed high resolution visualization of normal pancreas acinar cells, ducts, islets, capillaries and LEI in postcapillary venules. Histological images and optical sections from ex vivo CM can be correlated. Cellular morphology is better analyzed by conventional histology, but angioarchitecture and connective tissue structure are better evaluated by CM. FCD (265.7 +/- 16.6 cm-1) and LEI (rolling leukocytes 5.3 +/- 1.6/100 microns/sticking leukocytes 1.5 +/- 0.9/100 microns) were evaluated by in vivo CM in the normal pancreas. CONCLUSIONS: CM findings in tissues ex vivo correlate with those of classical histology but add informative details of connective tissue structure and angioarchitecture. Potential future applications for in vivo CM include real-time analysis of microcirculation, leukocyte-endothelial interaction and angiostructure in inflammatory and malignant pancreatic diseases. PMID- 12120267 TI - Pancreatitis-associated protein levels in pancreatic juice from patients with pancreatic diseases. AB - BACKGROUND: Pancreatitis-associated protein (PAP), the acute-phase protein of the pancreas, is overexpressed in acute pancreatitis. Serum PAP levels were reported to be useful as an indicator of the severity, prognosis and healing of acute pancreatitis. Although PAP was originally identified in pancreatic juice, there has been no clinical report on PAP levels in pancreatic juice. This study was conducted to determine levels of PAP in pancreatic juice (PJ-PAP) in various human pancreatic diseases. METHODS: PAP levels in endoscopically aspirated PJ were measured by enzyme-linked immunosorbent assay in 86 patients with pancreatic diseases. RESULTS: 55% of 22 patients with pancreatic cancer (PC) and 25% of 49 patients with chronic pancreatitis (CP) were positive (> 350 ng/ml) for PJ-PAP. PJ-PAP levels were significantly higher in PC than in CP, in which PJ-PAP was also significantly higher than in 15 control subjects. There was no significant correlation between PJ-PAP and serum PAP, and combination assay of serum PAP and/or PJ-PAP detected 80% of PC cases and 44% of CP cases. CONCLUSIONS: We have demonstrated that human PAP could be detected in pancreatic juice from patients with pancreatic diseases. Determination of PAP in pancreatic juice might be helpful for early detection of pancreatic injury. PMID- 12120269 TI - Body composition, physiological function and psychological changes in patients with predicted severe acute pancreatitis. AB - BACKGROUND: Serious sepsis and major blunt trauma have adverse effects on body composition, physiological function and psychological state. The effect of severe acute pancreatitis on these groups of variables has not yet been reported. METHODS: We have studied the effect of predicted severe acute pancreatitis (admission APACHE II score > or = 6) on body fat and mid arm muscle circumference (assessed by dual-energy X-ray absorptiometry and anthropometry), respiratory function (measured by spirometry and vitalography), voluntary muscle function (measured by hand dynamometry) and psychological state (measured by use of the hospital anxiety and depression score and visual analogue scale for fatigue) on admission, and three and seven days thereafter. RESULTS: The median APACHE II score of the 15 patients (7 men) in this study was 10 (6-13). The patients showed significant improvement in respiratory function (FEV1 1.3, 1.6, 2.3 litres/s, p < 0.01; FVC 1.5, 2.0, 2.9 litres, p < 0.01; PEFR 304, 372, 409 litres/min, p = 0.01 but no change in voluntary muscle function (210, 205, 213 N, p = 0.41) over the 7 day study period. Psychological state improved in terms of fatigue (2.4, 4.2, 7.0, p < 0.01) and depression (6, 9, 4, p = 0.12) but not in anxiety (8, 6, 7, p = 0.07). Body fat measured by DEXA (17.3 kg on admission, 16.7 kg on day 7, p = 0.13) and the mid upper arm muscle circumference (262, 248, 251 mm, p = 0.10) did not change implying that energy and nitrogen balance over the 7-day study period was achieved. CONCLUSION: Predicted severe acute pancreatitis has an adverse effect on respiratory function and psychological state similar to that observed in serious sepsis. Improvement in these variables is apparent over 7 days of effective treatment. Further research is indicated to examine the effect of nutritional support on these variables. PMID- 12120270 TI - Management of patients with extended pancreatic necrosis. AB - BACKGROUND: Extended pancreatic necrosis pose a considerable therapeutic problem in patients with necrotizing pancreatitis. AIM: Factors that limit conservative treatment in patients with extended pancreatic necrosis were analyzed. METHODS: The clinical course of 61 patients with an extent of necrosis of more than 50% of the gland (according to contrast-enhanced CT scan) were analysed with special regard to systemic complications. Indications for surgical treatment were either persistent organ failure or pancreatic infection. RESULTS: 10 patients were managed by conservative treatment, 51 (84%) patients underwent operation. Indications for surgery were sepsis with or without organ failure in 17 patients, persistent organ failure in another 17 patients, persistent SIRS in 13 patients and local complications in 4 patients. Pancreatic infection was present in 25 patients. The incidence of systemic complications did not differ between infected and sterile necrosis, but they occurred earlier in sterile necrosis. CONCLUSIONS: Persistent organ failure is limiting conservative treatment during the early course in patients with sterile necrosis. The latter course is characterized by a high incidence of pancreatic infection and septic organ failure. PMID- 12120271 TI - Successful extracorporeal shock wave lithotripsy for sibling pancreatic duct stones. AB - We present a case of 2 brothers with idiopathic chronic pancreatitis associated with pancreatic duct stones which could be successfully disintegrated by extracorporeal shock wave lithotripsy (ESWL). An obvious etiology for the pancreatolithiasis, like alcohol or biliary disease, was lacking and point mutations of the cationic trypsinogen gene exons 2 and 3 were not detected in the long arm of the 7th chromosome. However, a hereditary etiology could not be precluded since pancreatolithiasis occurred in the siblings. There has been no recurrence of pancreatic stones during 42 months of follow-up periods, for both. ESWL, the least invasive therapy, appeared applicable and effective for pancreatolithiasis in the present cases. PMID- 12120272 TI - Acute pancreatitis-induced hypomagnesemia. AB - We describe the case of an alcoholic patient admitted to our hospital with alcohol-induced acute pancreatitis who developed severe hypomagnesemia (serum magnesium 0.36 mmol/l) during hospitalization. The underlying pathogenetic mechanisms of hypomagnesemia are discussed. PMID- 12120273 TI - The death of an all-star. PMID- 12120274 TI - Breeding grounds. PMID- 12120275 TI - Mysteries of the beach. PMID- 12120276 TI - Obesity market overview. PMID- 12120277 TI - PXR, CAR and drug metabolism. AB - Mechanisms that protect the body from a diverse array of harmful chemicals are also involved in drug metabolism, and can cause adverse drug-drug interactions. Two closely related orphan nuclear hormone receptors--the pregnane X receptor (PXR) and the constitutive androstane receptor (CAR)--have recently emerged as transcriptional regulators of cytochrome P450 expression that couple xenobiotic exposure to oxidative metabolism. In this review, we provide an examination of the roles of PXR and CAR as xenobiotic sensors, and discuss the application of this knowledge to toxicological screening in drug discovery. PMID- 12120278 TI - Therapeutic strategies for human amyloid diseases. AB - Amyloid diseases are a large group of a much larger family of misfolding diseases. This group includes pathologies as diverse as Alzheimer's disease, immunoglobulin-light-chain disease, reactive amyloid disease and the familial amyloid polyneuropathies. These diseases are generally incurable at present, although some drugs are known to transiently slow the progression of Alzheimer's disease. As we increase our understanding of the causative mechanisms of these disorders, the likelihood of success for a given therapeutic strategy will become clearer. This review will look at small-molecule and macromolecular approaches for intervention in amyloid diseases other than Alzheimer's disease, although select examples from Alzheimer's disease will be discussed. PMID- 12120279 TI - Obesity therapy: altering the energy intake-and-expenditure balance sheet. AB - Obesity is associated with numerous health complications, which range from non fatal debilitating conditions such as osteoarthritis, to life-threatening chronic diseases such as coronary heart disease, diabetes and certain cancers. The psychological consequences of obesity can range from lowered self-esteem to clinical depression. Despite the high prevalence of obesity and the many advances in our understanding of how it develops, current therapies have persistently failed to achieve long-term success. This review focuses on how fat mass can be reduced by altering the balance between energy intake and expenditure. PMID- 12120280 TI - Histone-deacetylase inhibitors: novel drugs for the treatment of cancer. AB - The opposing actions of histone acetyltransferases (HATs) and histone deacetylases (HDACs) allow gene expression to be exquisitely regulated through chromatin remodelling. Aberrant transcription due to altered expression or mutation of genes that encode HATs, HDACs or their binding partners, is a key event in the onset and progression of cancer. HDAC inhibitors can reactivate gene expression and inhibit the growth and survival of tumour cells. The remarkable tumour specificity of these compounds, and their potency in vitro and in vivo, underscore the potential of HDAC inhibitors as exciting new agents for the treatment of cancer. PMID- 12120281 TI - Ethical perspectives on pharmacogenomic profiling in the drug development process. AB - Pharmacogenomics, which is a field that encompasses the study of genetic polymorphisms that underlie individual differences in drug response, is rapidly advancing. The potential for the widespread use of pharmacogenomics in the drug development process merits an examination of its fundamental impact on clinical trial design and practice. This article provides a critical analysis of some of the issues that pertain to pharmacogenomics in the drug development process. In particular, four areas will be discussed: clinical-trial design; subject stratification; some new social risks; and economic concerns. Recommendations are offered for addressing the issues that are discussed and anticipating the regulatory needs for pharmacogenomics-based trials. PMID- 12120282 TI - Protein kinases--the major drug targets of the twenty-first century? AB - Protein phosphorylation regulates most aspects of cell life, whereas abnormal phosphorylation is a cause or consequence of disease. A growing interest in developing orally active protein-kinase inhibitors has recently culminated in the approval of the first of these drugs for clinical use. Protein kinases have now become the second most important group of drug targets, after G-protein-coupled receptors. Here, I give a personal view of some of the most important advances that have shaped this field. PMID- 12120283 TI - A postmodern moral tale: the ethics of research relationships. AB - Once upon a time, universities were ivory towers--or, at least, most people saw them as such. Academics prided themselves on not being influenced by, or answerable to, anyone except the academy, and perceived themselves as free from conflicts of interest arising from relationships with people or institutions outside the walls of the university. None of this holds true today and, as the case of Dr Nancy Olivieri illustrates, universities are facing some difficult times in adjusting to these changes. PMID- 12120284 TI - Pharmaceutical consultancy: a view to a skill. PMID- 12120285 TI - Expanding the genetic code. AB - The ability to incorporate unnatural amino acids into proteins directly in living cells will provide new tools to study protein and cellular function, and may generate proteins or even organisms with enhanced properties. Due to the limited promiscuity of some synthetases, natural amino acids can be substituted with close analogs at multiple sites using auxotrophic strains. Alternatively, this can be achieved by deactivating the editing function of some synthetases. The addition of new amino acids to the genetic code, however, requires additional components of the protein biosynthetic machinery including a novel tRNA-codon pair, an aminoacyl-tRNA synthetase, and an amino acid. This new set of components functions orthogonally to the counterparts of the common 20 amino acids, i.e., the orthogonal synthetase (and only this synthetase) aminoacylates the orthogonal tRNA (and only this tRNA) with the unnatural amino acid only, and the resulting acylated tRNA inserts the unnatural amino acid only in response to the unique codon. Using this strategy, the genetic code of Escherichia coli has been expanded to incorporate unnatural amino acids with a fidelity rivaling that of natural amino acids. This methodology is being applied to other cell types and unnatural analogs with a variety of functionalities. PMID- 12120286 TI - A novel ribbon-candy-like supramolecular architecture of cadmium(II) terephthalate polymer with giant rhombic channels: twofold interpenetration of the 3D 8(2)10-a net. AB - The hydrothermal reaction of H2tp (tp = terephthalate), [Ph3PCH2Ph]Cl and water with Cd(O2CCH3)(2).2H2O gives rise to a novel ribbon-candy-like supramolecular architecture with twofold interpenetration of an unprecedented 3D 8(2)10-a net formed by polymer ([Ph3PCH2Ph][Cd(tp).Cl].2H2O]n containing giant rhombic channels, which displays strong fluorescent emission in the solid state. PMID- 12120287 TI - Hydrogen-bonded extended structure of the 1:3 adduct of a C3 symmetric cobalt(III) complex with a tripod-ligand involving three imidazolate groups and hydroquinone or resorcinol. AB - The assembly reaction arising from hydrogen bonding between a chiral C3 symmetric cobalt(III) complex and a tripod-ligand involving three imidazolate groups [tris[2-(((2-methylimidazolato-4-yl)methylidene)amino)ethyl]amine]cobalt(III) and either hydroquinone or resorcinol gave the 1:3 adducts, with 3D extended structures showing the template effect of the complex. PMID- 12120288 TI - Ni(II) N4-macrocycle grafted crown ether: caesium cobalt(III) bis(dicarbollide) coordination polymer. AB - A mixture of a Ni(TMTAA) grafted crown ether and Cs[Co(C2B9H11)2] in toluene CH2Cl2 affords a 1:1 complex, comprised of layers of infinite two-dimensional polymeric arrays separated by layers of the cobalticarborane anion which themselves form two B-H...Cs+ hydrogen interactions. PMID- 12120289 TI - Super acid catalysis in supercritical fluid reaction media for the formation of linear alkyl benzenes. AB - High catalytic activity is demonstrated for the formation of linear alkylbenzenes using a perfluorosulfonic acid catalyst in supercritical fluid reaction media: enhanced alkylation activity is observed in fluoroform (CHF3) compared to carbon dioxide. PMID- 12120290 TI - Dimethylargenate is a stable species in the gas phase. AB - The formation of dimethylargenate from silver diacetate has been explored using multistage mass spectrometry experiments in a quadrupole ion trap and DFT theory. PMID- 12120291 TI - Formation of oligotriazoles catalysed by cucurbituril. AB - The catalytic activity of cucurbituril in 1,3-dipolar cycloadditions has been applied to the synthesis of oligotriazoles. PMID- 12120292 TI - Dissipative water intrusion in hydrophobic MCM-41 type materials. AB - Texture-related features of water intrusion in hydrophobised MCM-41 silicas render these materials especially suitable for energy dissipation in mechanical dampers. PMID- 12120293 TI - Planar and central chiral [2.2]paracyclophane-based N,O-ligands as highly active catalysts in the diethylzinc addition to aldehydes. AB - The application of planar and central chiral [2.2]paracyclophane-based N,O ligands in asymmetric diethylzinc additions to various aldehydes is described revealing an unusual substrate spectrum for the catalysts and their remarkably high activity. PMID- 12120294 TI - First examples of the catalytic asymmetric ring-opening of meso 1,2-dioxines utilising cobalt(II) complexes with optically active tetradentate Schiff base ligands: formation of enantio-enriched cyclopropanes. AB - The combination of chiral cobalt beta-ketoiminato or cobalt salen complexes and meso 1,2-dioxines leads to catalytic asymmetric ring-opening affording enantio enriched cis gamma-hydroxy enones; subsequent capture by an ylide affords enantio enriched cyclopropanes. PMID- 12120295 TI - Enhancing the reactivity of uranium(VI) organoimido complexes with diazoalkanes. AB - Diphenyldiazomethane effects a two-electron oxidation of the uranium(IV) monoimido complex (C5Me5)2U(=N-2,4,6-t-Bu3C6H2) to give the uranium(VI) mixed bis(imido) complex, (C5Me5)2U(=N-2,4,6-t-Bu3C6H2)(=N-N=CPh2), which undergoes a rare cyclometallation reaction upon mild thermolysis to afford a uranium(IV) bis(amide) complex that results from net addition of a C-H bond of an ortho tert butyl group across the N=U=N core. PMID- 12120296 TI - Chelating bis-carbene rhodium(III) complexes in transfer hydrogenation of ketones and imines. AB - Chelating rhodium(III) carbene complexes are accessible via a simple synthesis and are catalytically active for hydrogen transfer from alcohols to ketones and imines. PMID- 12120297 TI - Nanotube composites: novel SiO2 coated carbon nanotubes. AB - A novel route to nanocomposites consisting of multi-walled carbon nanotubes (MWNTs) embedded in amorphous SiOx is reported; the material has been characterised by high resolution transmission electron microscopy (HRTEM) and high resolution electron energy loss spectroscopy (HREELS); for the first time, and based on our observations, we propose theoretical models accounting for stable SiOx/tube interfaces using density functional based tight binding (DFTB). PMID- 12120298 TI - A novel multidecker sandwich complex from the reaction of ferrocene with GaCl3. AB - A redox reaction takes place between GaCl3 and an excess of ferrocene, leading to the multidecker sandwich complex 1, [-Ga(C5H5)Fe(C5H5)Ga(C5H5)Fe(C5H5) ]n[GaCl4]2n, which features an array of alternating Ga(I) and Fe(II) ions bridged by cyclopentadienyl moieties. PMID- 12120299 TI - Asymmetric deprotonation and dearomatising cyclisation of N-benzyl benzamides using chiral lithium amides: formal synthesis of (-)-kainic acid. AB - Chiral lithium amides deprotonate N-benzyl benzamides enantioselectively, initiating an asymmetric dearomatising cyclisation to enantiomerically enriched isoindolinones. PMID- 12120300 TI - Novel reactions of phosphorus(III) azides and isocyanates: unusual modes of cycloaddition with dipolarophiles and an unexpected case of ring expansion. AB - New modes of 1,3-dipolar cycloaddition are uncovered by the isolation of [CH2(6-t Bu-4-Me-C6H2O)2]P(C(CO2Me)C(CO2Me)N[NP(N3)(OC6H2-6-t-Bu-4-Me)2CH2]N) (3) and [CH2(6-t-Bu-4-Me-C6H2O)2]P(C(CO2Me)C(CO2Me)C(O)N) (4) on treating [CH2(6-t-Bu-4 Me-C6H2O)2]P-X [X = N3 (1) and NCO (2)] with the dipolarophile MeO2CC identical to CCO2Me; compound 4 undergoes an unprecedented ring expansion upon addition of 2-(methylamino)ethanol to afford the spirocycle [CH2(6-t-Bu-4-Me C6H2O)2]P(OCH2CH2N(Me)CH(CO2Me)CH(CO2Me)C(O)N) (5). PMID- 12120301 TI - Total synthesis and determination of the stereochemistry of 2-amino-3 cyclopropylbutanoic acid, a novel plant growth regulator isolated from the mushroom Amanita castanopsidis Hongo. AB - The unknown stereostructure of 2-amino-3-cyclopropylbutanoic acid 1, a novel plant growth regulator isolated from the mushroom Amanita castanopsidis Hongo, was determined to be (2S,3S)-2 through its racemic and enantioselective syntheses employing the chelate-enolate Claisen rearrangement as a key step. PMID- 12120302 TI - 1D silver(I) complex of nitronyl nitroxide with strong spin-spin interaction through silver(I) ion. AB - A 1D silver(I) complex of nitronyl nitroxide was prepared and its structure was determined by X-ray diffraction analysis; magnetic studies indicate that the spin spin interaction of nitronyl nitroxides through silver(I) ions along the chain are fairly strong (J/kb = -84 K). PMID- 12120303 TI - Functionalisation of carbon dioxide by an iron(II) complex. AB - Addition of carbon dioxide to trans-Fe(dmpe)2(SCHNEt)H 2 affords the iminium carboxylate trans-Fe(dmpe)2(SCHN+(Et)CO2-)H 4, which rearranges to the ferracyle cis-Fe(dmpe)2(SCH2N(Et)C(O)O-kappa S,O) 5. PMID- 12120304 TI - Cryptophanols, new versatile compounds for the synthesis of functionalized cryptophanes and polycryptophanes. AB - After deprotection with a palladium catalyst, mono-allylated cryptophane-A (1, 2) and cryptophane-E (3) gave the new cryptophanols 4, 5 and 6, respectively, which are important key compounds for the preparation of monofunctionalized cryptophanes as well as for the design of large supramolecular hosts such as the bis-cryptophanes 7 and 8. PMID- 12120305 TI - Palladium-catalysed asymmetric arylation of tert-cyclobutanols via enantioselective C-C bond cleavage. AB - Palladium-catalysed arylation of tert-cyclobutanols with aryl bromide involving enantioselective C-C bond cleavage affords chiral ketones with moderate to good enantioselectivity. PMID- 12120306 TI - Novel catalysis of dendrimer-bound Pd(0) complexes: sterically steered allylic amination and the first application for a thermomorphic system. AB - Phosphinated dendrimer-bound Pd(0) complex catalysts show high stereoselectivity for allylic amination due to the surface congestion of dendrimers and can be easily recycled without loss of activity under thermomorphic conditions. PMID- 12120307 TI - 1,2-diphenylethylenediamine linked chiral Ti(IV) complex--a new entry to the highly enantioselective silylcyanation of aliphatic and aromatic aldehydes. AB - Highly enantioselective silylcyanation of aliphatic and aromatic aldehydes was achieved by using a 1,2-diphenylethylenediamine linked chiral Ti(IV) complex as the catalyst. PMID- 12120308 TI - Catalytic one-pot synthesis of [5.5.5.6]fenestrane systems via a dicobalt octacarbonyl-catalyzed tandem cycloaddition of dienediynes. AB - Catalytic one-pot synthesis of fenestrane derivatives from dienediynes was developed: fenestranes were synthesized in high yields by a dicobalt octacarbonyl catalyzed tandem cycloaddition of dienediynes. PMID- 12120309 TI - In situ STM imaging of surface dissolution and rearrangement of a Pt-Fe alloy electrocatalyst in electrolyte solution. AB - The formation of a Pt skin layer with predominantly (111)-oriented facets induced by dissolution of Fe atoms in a Pt-Fe alloy for fuel cell applications is investigated for the first time by using in situ electrochemical STM in 0.1 M HClO4 solution. PMID- 12120310 TI - Palladium nanoparticles stabilised by polyfluorinated chains. AB - Palladium nanoparticles can be prepared by reduction of palladium(II) chloride in the presence of different compounds featuring long perfluorinated carbon chains. PMID- 12120311 TI - Novel water clusters in the crystalline state: structures of a symmetrical, cyclic hexamer and an 'opened-cube' octamer. AB - Six- and eight-membered hydrogen-bonded water clusters of novel structure types have been found in crystalline hydrates. PMID- 12120312 TI - [M(N3)2(L)]n: building 3-D MII-azido networks with new topologies. AB - Novel 3D topologies combining diazine and azido bridges between MnII magnetic centres have been obtained and characterised by low-temperature magnetic measurements. PMID- 12120313 TI - Deuterium NMR spectroscopy is a versatile and economical tool for monitoring reaction kinetics in ionic liquids. AB - Time-resolved 2H NMR spectroscopy is used to monitor the progress of and gain kinetic information for a variety of reactions in different ionic media. PMID- 12120314 TI - A non-aqueous organometallic route to highly monodispersed copper nanoparticles using [Cu(OCH(Me)CH2NMe2)2]. AB - Good quality, highly monodispersed capped copper metal nanoparticles have been synthesised in a non-hydrolytic approach using thermal decomposition of the Cu(II) precursor [Cu(OCH(Me)CH2NMe2)2] in a hot coordinating solvent without further reducing agents; the copper nanoparticles have been characterised by optical spectroscopy (UV/VIS), electron microscopy (TEM), electron diffraction (SAED), and dynamic light scattering (DLS). PMID- 12120315 TI - Pyrene-neomycin conjugate: dual recognition of a DNA triple helix. AB - We report the synthesis of pyrene-neomycin conjugate and its ability to stabilize DNA/RNA triple helices. PMID- 12120316 TI - Fabrication of a nano-scale gap by selective chemical deposition. AB - An electrode nanogap of 45 nm has been prepared by a new method in which the initial gap of 1-2 microns obtained by conventional lithography was shortened by selective chemical deposition of copper onto the electrodes. PMID- 12120317 TI - [Cu(pyrazine-2-carboxylate)2]2Cd4I8: unprecedented 1-D serpentine inorganic chains and regular 2-D metal-organic square grids in a 3-D framework. AB - A novel three-dimensional coordination polymer containing an unprecedented one dimensional serpentine motif conjoined with a two-dimensional square grid motif is presented. PMID- 12120318 TI - Single phase preparation of monodispersed silver nanoclusters using a unique electron transfer and cluster stabilising agent, triethylamine. AB - A simple and reproducible single phase preparation of 2.5 nm silver nanoclusters is described using silver benzoate along with triethylamine (TEA) and dodecanethiol (DDT); these spontaneously self-assemble to a two-dimensional array whereas in the absence of thiol only polydispersed nanoclusters are obtained. PMID- 12120319 TI - Labile coodination dendrimers. AB - Addition of anionic benzylsulfate dendrons to dynamic mixtures of Ag+ and triphosphine ligands results in the assembly of loosely-bonded cage-core dendrimers. PMID- 12120320 TI - Novel low temperature solution deposition of perpendicularly orientated rods of ZnO: substrate effects and evidence of the importance of counter-ions in the control of crystallite growth. AB - Perpendicularly orientated ZnO rods have been grown on thin ZnO templates, from aqueous solutions of zinc acetate and hexamethylenetetraamine (HMT); growth along the c-axis of the ZnO crystallites is promoted by the presence of acetate in the bath. PMID- 12120321 TI - Atomic force microscopy studies of the KF (100) surface: formation of water-rich surface phases in moist N,N-dimethylformamide. AB - A water rich surface phase is observed in the system KF-DMF-H2O by atomic force microscopy; the effects on surface morphology and likely implications for halogen exchange reactions using KF are discussed. PMID- 12120322 TI - An isolated cyclo-tetraarsendiide: low temperature synthesis and crystal structure of bis-pentaamminesodium tetraarsendiide-ammonia (1/3) [Na(NH3)5]2As(4).3NH3. AB - Reduction of an excess of arsenic with a solution of sodium in liquid ammonia yields the thermally unstable title compound [Na(NH3)5]2As(4).3NH3, which contains isolated square-planar As4(2-) ions with a mean As-As bond length of 2,345 A. PMID- 12120323 TI - Synthesis, crystal and molecular structure of [Sn(eta 4-P2C2But2)]: the first non transition metal 1,3-diphosphacyclobutadienyl compound. AB - Treatment of [Zr(eta 5-C5H5)2(PCBut)2] with SnCl2 led to the novel monomeric (1,3 diphosphacyclobutadienyl)tin(II) half sandwich complex [Sn(eta 4-P2C2But2)] which has been characterised by multinuclear NMR spectroscopy and in the solid state by a single crystal X-ray diffraction study. PMID- 12120325 TI - Preliminary evidence for sinapyl acetate as a lignin monomer in kenaf. AB - 9-Acetylated syringyl 8-8-linked dehydrodimers are degradation products released from kenaf lignins, implicating sinapyl acetate as a lignin precursor. PMID- 12120324 TI - In vitro selection and evaluation of RNA aptamers that recognize arginine-rich motif model peptide on a quartz-crystal microbalance. AB - To study RNA-peptide interactions, we performed an in vitro selection of RNA on a simple alpha-helical peptide-immobilized quartz-crystal microbalance (QCM) and evaluated the association constants (10(7) M-1) of the selected RNA to the model peptide on the same QCM plate. PMID- 12120326 TI - First example of a covalently bound dimeric inverted porphyrin. AB - Reaction of 5,10,15,20-tetraphenyl-2-aza-21-carbaporphyrinatonickel(II) with dihalomethanes in the presence of a proton scavenger gives a 2,21'-CH2-linked dimer of Ni(II) inverted porphyrins with the yield reaching 90% and its 2,2' linked isomer as a minor product. PMID- 12120327 TI - Direct thioesterification from carboxylic acids and thiols catalyzed by a Bronsted acid. AB - In the presence of a catalytic amount of trifluoromethanesulfonic acid, free carboxylic acids reacted with free thiols directly to afford the corresponding thioesters in high yields. PMID- 12120328 TI - Synthesis of a novel mesoporous tin phosphate, SnPO4. AB - Novel mesoporus tin phosphates have been synthesized using alkyltrimethylammonium bromide (alkyl = C8-C18) as surfactant; the structure of the materials is stable at 500 degrees C. UV-VIS spectra show tetrahedral coordination of tin. PMID- 12120329 TI - Facile synthesis of ketones from 1,1-disilylethenes via oxidation of gem disilylalkanes. AB - The oxidation of gem-disilylalkanes, which can be derived from 1,1-disilylethene, alkyllithiums and alkyl halides, affords the corresponding ketones. PMID- 12120330 TI - Novel preparation of beta,beta'-connected porphyrin dimers. AB - Porphyrin dimers were prepared from beta,beta'-dipyrrole derivatives via the double pyrrolylmethylation followed by double [2 + 2] MacDonald porphyrin synthesis. PMID- 12120331 TI - Efficient soluble polymer-supported sharpless alkene epoxidation catalysts. AB - High chemical yields and good enantiomeric excesses are obtained by using soluble polymer-supported tartrate ester in the epoxidation of trans-hex-2-en-1-ol using Ti(OPri)4/tert-butyl hydroperoxide. PMID- 12120332 TI - A novel type of formation of zwitterionic compounds, containing two phosphorus atoms of opposite charge and different coordination number. AB - The oxidation of bis[bis(dialkylamino)phosphinyl]methane 1 with 5,5,5-trifluoro-4 (trifluoromethyl)penta-3-en-2-one 2 unexpectedly gave zwitterionic compound 7 which according to X-ray analysis contains two phosphorus atoms of opposite charge and different coordination number (lambda 4P(+); lambda 6P(-)) with a direct P-H bond at the hexacordinated phosphorus. PMID- 12120333 TI - Synthesis and application of chiral bisphosphines through lithiation-conjugate addition tandem cyclization of chiral alpha,beta,psi,omega-unsaturated bisphosphine oxide. AB - Upon treatment with lithium diisopropylamide achiral and chiral alpha,beta,psi,omega-unsaturated bisphosphine oxides underwent lithiation conjugate addition tandem cyclization to afford the corresponding endo-alpha,beta unsaturated cyclic bisphosphine oxides; sequential stereoselective reduction of the cyclized bisphosphine oxide gave the corresponding trans- and cis bisphosphines that were successfully applicable in a catalytic asymmetric hydrogenation as chiral bisphosphine ligands. PMID- 12120334 TI - Amidation of silyl enol ethers and cholesteryl acetates with chiral ruthenium(II) schiff-base catalysts: catalytic and enantioselective studies. AB - Chiral ruthenium(II)-salen complexes [RuII(salen)(PPh3)2] catalyse asymmetric aziridination of alkenes with up to 83% ees, asymmetric amidation of silyl enol ethers with up to 97% ees, and allylic amidation of cholesteryl acetates with good regioselectivity. PMID- 12120335 TI - Transcyclometalation, a versatile methodology for multiple metal-carbon bond formation with multisite ligands. AB - Transcyclometalation has been successfully applied for the multi-platination and ruthenation of 'chartwheel'-type ligand systems containing six potential metal binding sites, thus providing a method for multiple metal-carbon bond formation via C-H bond activation which is superior to established cyclometalation protocols. PMID- 12120336 TI - One-step synthesis of alkenyl ketone complexes from Cp*RhCl2(PPh3), alkyne and H2O in the presence of KPF6. AB - The reaction of Cp*RhCl2(PPh3) 1 with 1-alkyne and H2O in the presence of KPF6 afforded the alkenyl ketone complex [Cp*Rh(PPh3)(CPh=CHCOCH2R)](PF6) [R = p-tolyl (3a), R = Ph (3b)], whereas Cp*IrCl2(PPh3) 2 or [(eta 6-C6Me6)RuCl2(PPh3) gave the corresponding [Cp*IrCl(CO)(PPh3)](PF6) 5a and [(eta 6 C6Me6)RuCl(CO)(PPh3)](PF6). PMID- 12120337 TI - The d2/d3 alkyne redox pair [WF2(PhC identical to CPh)Tp']z (z = +1 or 0): missing links in a 'redox family tree'. AB - The d2/d3 redox pair [WF2(PhC identical to CPh)Tp']z [z = +1 or 0, Tp' = hydrotris(3,5-dimethylpyrazolyl)borate] is the missing link in a 'redox family tree' relating the d6 tricarbonyls [M(CO)3L]- to the d2 trihalides [MX3L] (M = Mo or W, L = Cp or Tp') by a series of stepwise reactions involving sequential one electron oxidation followed by ligand substitution. PMID- 12120339 TI - Aza-pinacol rearrangement: acid-catalyzed rearrangement of aziridines to imines. AB - A series of di-, tri-, and tetra-substituted N-tosylaziridines [N-(toluene-p sulfonyl)aziridines] 1, prepared by aziridination of the corresponding alkenes with N-[(tolyl-p-sulfonyl)imino]phenyliodinane (TsN = IPh), was found to undergo a BF3-catalyzed rearrangement (aza-pinacol rearrangement) under mild conditions to give the corresponding N-tosylimines 2 generally in satisfactory yields. PMID- 12120338 TI - A novel molecular assembly mode of ferulic acid derivatives. AB - Ferulic acid derivative assembles with three kinds of non-covalent interactions, i.e., metal coordination, hydrogen bonding and CH-pi interaction: X-ray crystallographic study illustrated the molecular assembly mode. PMID- 12120340 TI - Excited state intramolecular redox reaction of 2-(hydroxymethyl)anthraquinone in aqueous solution. AB - The title compound undergoes a novel excited state intramolecular redox reaction in which the 'distal' side chain benzylic alcohol is oxidized to the aldehyde and the carbonyl moieties of anthraquinone reduced, with evidence suggesting that the primary photochemical process is a deprotonation of the benzylic C-H proton (by water) mediated by the solvent. PMID- 12120341 TI - Novel synthesis of organic nanowires and their optical properties. AB - Aligned nanowires of organic luminescent material were prepared by introducing the organic luminants into nanochannels of variable size in an anodic aluminum oxide (AAO) membrane, and the emission spectra from these nanowire arrays exhibited novel size-dependent luminescent properties. PMID- 12120342 TI - Supported diazonium salts--convenient reagents for the combinatorial synthesis of azo dye. AB - Resin supported diazonium salts were synthesised. These were observed to be stable to storage and to provide a convenient means of compound handling and were employed in the solution synthesis of a 6 x 6 azo dye library. PMID- 12120343 TI - One-pot synthetic route to homoallylketones via selective combination of alkyne, ethylene and aldehyde mediated by AlCl3 and zirconocene. AB - Zirconacyclopentenes generated in situ from alkynes and ethylene undergo selective reaction with aldehydes to afford homoallylketones in yields exceeding 70% in the presence of AlCl3. PMID- 12120344 TI - Seed-mediated growth of large, monodisperse core-shell gold-silver nanoparticles with Ag-like optical properties. AB - Large, monodisperse core-shell Au-Ag nanoparticles with Ag-like optical properties have been prepared by the seeding growth method in micellar media. PMID- 12120345 TI - Alkene metathesis catalysis in ionic liquids with ruthenium allenylidene salts. AB - Ring closing metathesis of dienes in 1-butyl-3-methylimidazolium salts in the presence of ruthenium allenylidene salts as catalyst is described. PMID- 12120347 TI - Divergent approach to imino sugar C-glycosides using imino glycals: application to the stereocontrolled synthesis of (+)-deoxoprosophylline. AB - Tri-O-acetyl imino glucal 2 is readily made and shown to undergo a variety of Lewis acid mediated carbon-carbon bond forming reactions at C-1 of the piperidine nucleus. In all the reactions studied, the beta-anomer is predominant. PMID- 12120346 TI - Spontaneous disproportionation of rhodium(I) bisoxazolinates to rhodium(II). AB - [RhI(t-Bu2-boxate)(C2H4)2] spontaneously disproportionates to the mononuclear [RhII(t-Bu2-boxate)2], whereas [RhI(Ph2-boxate)(C2H4)2] is stable against disproportionation. PMID- 12120348 TI - Lipophilic derivatives of cyclam as new inhibitors of tumor cell growth. AB - Two new lipophilic tetraazamacrocycles were prepared and, in contrast to non lipophilic analogs, found to be potent inhibitors of tumor cell growth in vitro with IC50 values below 10 micromolar. PMID- 12120349 TI - A polyoxometallate-templated coordination polymer: synthesis and crystal structure of [Cu3(4,4'-bipy)5(MeCN)2]PW12O(40).2C6H5CN. AB - The spherical phosphotungstate ion, PW12O40(3-), has been used as a non coordinating anionic template for the construction of a novel, three-dimensional Cu(I) coordination polymer. PMID- 12120351 TI - Access to 2,5-disubstituted tetrahydrofurans from Grignard reagents and hemiacetal derivatives. AB - The first diastereoselective addition of Grignard reagents onto cyclic oxocarbenium ions, obtained from glycosyl acetates, to afford 2,5-disubstituted tetrahydrofurans, is reported. PMID- 12120350 TI - Magneto-switchable electrogenerated biochemiluminescence. AB - Magnetic-field-stimulated 'ON' and 'OFF' biochemiluminescence is accomplished by electrocatalyzed reduction of naphthoquinone-functionalized magnetic particles in the presence of a biocatalytic peroxidase/luminol system. PMID- 12120352 TI - Construction of positively-charged layered assemblies assisted by cyclodextrin complexation. AB - A beta-cyclodextrin dimer is found to be effective in preparing a layer-by-layer architecture of positively charged ferrocene-appended poly(allylamine) presumably on the basis of strong beta-cyclodextrin-ferrocene host-guest interaction. PMID- 12120353 TI - Synthesis of high silicon content SAPO(4)-5 using anionic surfactants in a hexanol/aqueous two phase media. AB - High silicon content SAPO(4)-5 (up to 0.511 atoms per unit cell) has been synthesised, using sodium 3-bromopropanesulfonate, sodium 1-butanesulfonate, sodium naphthalene-1-sulfonate or sodium n-decyl sulfate as surfactants; the SiO2 in the reaction gels ranged up to 3.0 (molar ratio), silicon incorporation was confirmed by XRD, XRF, TG-DTA, FT-IR and SEM techniques. PMID- 12120354 TI - Efficient synthesis of biaryl lactones by domino retro-Michael-aldol lactonization reactions. AB - Biaryl lactones were prepared by novel domino retro-Michael-aldol-lactonization reactions of 2,3-dihydropyrans. PMID- 12120355 TI - Synthesis and structure of a new macrotricyclic ligand that encapsulates lithium and transition metal ions. AB - A new macrotricyclic ligand, L3 with an N4S2 donor set that encapsulates lithium and transition metal ions has been synthesized from the tetraazamacrocycle cyclam. PMID- 12120356 TI - Titanium-catalyzed enantioselective alkynylation of aldehydes. AB - A simple and practical method to make chiral propargylic alcohols has been developed: in the presence of a titanium alkoxide catalyst prepared in situ from titanium tetraisopropoxide and (R)-H8-binaphthol, a variety of aromatic aldehydes were converted to the corresponding chiral propargylic alcohols with very good enantioselectivities (up to 96.2% e.e.) and yields. PMID- 12120357 TI - Colour indicator for enantiomeric excess and assignment of the configuration of the major enantiomer of an amino acid ester. AB - A colour indicator for the full range of enantiomeric excess (-100%-->100% ee) is presented which is based on visual colour inspection of a liquid crystal doped with the analyte, i.e. the methyl ester of amino acid phenylglycine, providing the enantiomeric excess and allowing the assignment of the major enantiomer. PMID- 12120358 TI - Lyotropic mesomorphism in some thermotropic, polycatenar complexes of silver(I). AB - A wide range of mesophases are induced on treating polycatenar complexes of silver(I) with a wide range of organic solvents. PMID- 12120359 TI - Controlling allostery using redox chemistry. AB - The binding of a hydrogen-bonding receptor to its substrate is reversibly regulated by varying the oxidation state of a copper allosteric cofactor. PMID- 12120360 TI - Remarkable effect of nitrogen dioxide for N-hydroxyphthalimide-catalyzed aerobic oxidation of methylquinolines. AB - Aerobic oxidation of methylquinolines was successfully achieved by the use of N hydroxyphthalimide/Co(OAc)2/Mn(OAc)2 as catalyst in the presence of a small amount of nitrogen dioxide as an initiator. PMID- 12120361 TI - Snapshots of a working catalyst: possibilities and limitations of in situ spectroscopy in the field of heterogeneous catalysis. AB - Probing events taking place in a heterogeneous catalyst under reaction conditions has attracted a lot of attention in recent years. It requires the development of characterization techniques and the construction of in situ spectroscopic reaction cells, which allow the identification of reaction intermediates and active sites in a working catalyst. The ultimate goals are the development of quantitative structure/composition-activity/selectivity relationships and the gathering of fundamental insight in short- and/or long-term deactivation mechanisms of heterogeneous catalysts. The use of in situ infrared, Raman, UV VIS, electron paramagnetic resonance, nuclear magnetic resonance, X-ray absorption and Mossbauer spectroscopy; the construction of valuable spectroscopic reaction cells and the possibilities and limitations to monitor real time catalytic events are reviewed. Several case studies describing the use of in situ spectroscopy in catalysis research are discussed. PMID- 12120362 TI - What can atomic force microscopy tell us about protein folding? AB - Force spectroscopy has emerged as a new tool to study protein folding, in which force replaces the chemical denaturant used in traditional folding experiments. This new technique complements older methods and allows a range of new questions to be investigated. What sort of protein is able to resist mechanical unfolding, and to what extent is mechanical stability dictated by fold or function? What is the effect of force on the unfolding energy surface? Do proteins unfold by the same pathway in mechanical and chemical denaturation experiments? Answers to these are starting to emerge based on a combination of experimental and computational approaches. We present some of the forced unfolding experiments to date and simple methods for characterizing the unfolding potential from the results. Several studies have also begun a more fine-grained description of mechanical unfolding, for example by invoking intermediates to explain features seen in unfolding traces and by using mutagenesis to try to localize the origin of mechanical stability. We propose further experimental approaches to this goal using the protein engineering method to characterize transition states, similar to those used in conventional folding experiments. However, it is likely that a high-resolution picture of mechanical unfolding will only emerge through a combined interpretation of careful experimental work and computer simulation. PMID- 12120363 TI - Solid-phase template-directed synthesis of a [2]rotaxane using a solid-phase stopper. AB - The first synthesis of a rotaxane by solid phase chemistry has been achieved, using the resin bead as a 'Mega' stopper during the synthesis. One of the advantages of this methodology over traditional solution routes include the ability to use mass action to drive the chemistry, without complicating the purification process. PMID- 12120365 TI - Silica-polyoxazoline hybrid with nanosized hollow enclosing porphyrin in hybrid walls. AB - A silica-polyoxazoline hybrid with nanosized hollows and porphyrin moieties was fabricated via an emulsion templated sol-gel route. PMID- 12120364 TI - In vivo cleavage of a target RNA by copper kanamycin A. Direct observation by a fluorescence assay. AB - A novel fluorescence assay to monitor in vivo cleavage chemistry of RNA target sequences has been established and used to demonstrate the activity of copper aminoglycoside mediated degradation of RNA in bacterial cells. PMID- 12120366 TI - Synthesis and properties of ionic liquids derived from the 'chiral pool'. AB - New chiral ionic liquids have been synthesised which are directly derived from the 'chiral pool' and therefore readily available in kg scale; NMR-measurements indicate that these liquids may be interesting solvents for enantioselective reactions and useful in chiral separation techniques. PMID- 12120367 TI - Highly efficient use of NaOCl in the Ru-catalysed oxidation of aliphatic ethers to esters. AB - The selectivity of alpha-oxidation of ethers to esters via Ru-NaOCl can be dramatically improved by pH control, at high substrate to catalyst ratios using a stoichiometric amount of hypochlorite in biphasic media at room temperature. PMID- 12120368 TI - The biosynthesis of bisvertinolone: evidence for oxosorbicillinol as a direct precursor. AB - Biosynthetic incorporation of labeled sodium acetate into oxosorbicillinol in Trichoderma sp. USF-2690 suggests that oxosorbicillinol is derived from six acetate units, and subsequent bioconversion of the labeled oxosorbicillinol to bisvertinolone in the fermentation of the strain suggests that bisvertinolone is biosynthesized from oxosorbicillinol and sorbicillinol in a Michael-type reaction. PMID- 12120369 TI - Unprecedented zwitterion in quinonoid chemistry. AB - The first 12 pi-electron zwitterionic structure in quinonoid chemistry is described with the N,N,O,O-molecule 6 in which the positive charge is pi delocalized between the nitrogen atoms and the negative charge between the oxygen atoms; depending on the crystallization solvent, a 1D-tape-like H-bonded network can be generated in the solid-state. PMID- 12120370 TI - Immobilized alpha-diazophosphonoacetate as a versatile key precursor for palladium catalyzed indole synthesis on a polymer support. AB - Rh(II)-catalyzed N-H insertion reaction of immobilized alpha diazophosphonoacetate with 2-haloanilines followed by Horner-Emmons reaction gave immobilized enaminoesters, which were efficiently cyclized to indoles via intramolecular palladium catalyzed reaction on a polymer support. PMID- 12120371 TI - Molecular design and evaluation of quinoxaline-carbohydrate hybrids as novel and efficient photo-induced GG-selective DNA cleaving agents. AB - Quinoxaline, found in antitumor quinoxaline antibiotics, was found to cleave double stranded DNA at the 5' side guanine of 5'-GG-3' site on irradiation with long wavelength UV light without any additive; furthermore, a bis(quinoxaline carbohydrate) hybrid system was very effective for DNA cleavage. PMID- 12120372 TI - Discriminating between lanthanide ions: self-assembly of heterodimetallic triple stranded helicates. AB - A bis-terdentate segmental ligand has been designed which self-assembles with lanthanide ions of different size to yield heterodimetallic triple-stranded helicates. PMID- 12120373 TI - Tandem radical and non-radical reactions mediated with thiols--a new method of cleavage of allylic amines. AB - Thiyl radical promotes the isomerisation of allylic amines into enamines via two consecutive hydrogen atom abstraction steps, and the subsequent polar addition of the corresponding thiol to the enamine results in the cleavage of the C-N bond via a thioaminal intermediate: this reaction provides a mild, metal-free methodology for the deprotection of allylated primary and secondary amines. PMID- 12120374 TI - Design of 3-D coordination networks: topology and metrics. AB - Using 4,4',4"-tricyanotriphenylmethanol 1 as a heterotetradentate tecton with C3v symmetry bearing three CN and one OH group, under self-assembly conditions a 3-D coordination network was obtained in the presence of Ag+ cations acting as a tetrahedral metallic tecton; due to the metrics of 1 (three long and one short distance between the central C atom and N and O coordination sites, respectively), the 3-D network is of pseudo-diamondoid type with different cavity sizes; although a two-fold homo-interpenetration is observed for the 3-D networks, the remaining space is occupied by CHCl3, MeOH solvent molecules and SbF6- anions. PMID- 12120375 TI - A novel C-C bond formation reaction with 1-methoxymethylindolylborate. AB - The reaction of 1-methoxymethylindolylborates 2 with electrophiles in the presence of benzaldehyde enabled the novel construction of tri-substituted indoles in a 'one-pot' procedure. PMID- 12120376 TI - Conjugate addition of radicals generated from diacyloxyiodobenzenes to dehydroamino acid derivatives; a synthesis of diaminopimelic acid analogues. AB - Radical decomposition of bis((2S)-N-benzyloxycarbonyl-2-aminopentan-5-carboxy-1 methyl ester)iodobenzene followed by decarboxylation and subsequent conjugate addition with a series of selectively protected dehydroamino acids leads to new analogues of diaminopimelic acid. PMID- 12120377 TI - Polyclonal antibody-catalysed hydrolysis of a beta-lactam. AB - We report the first example of antibody-catalysed hydrolysis of a beta-lactam where the antibodies were generated by a simple transition-state analogue; in this example the antibodies are polyclonal. PMID- 12120378 TI - Rapid assembly of highly-functionalised difluorinated cyclooctenones via ring closing metathesis. AB - Building block methodology from trifluoroethanol and ring-closing metathesis using a Furstner modification of Grubbs' conditions allows the rapid synthesis of novel difluorinated cyclooctenones. PMID- 12120379 TI - Direct observation of eta 2-imine formation through beta-H abstraction between amide ligands. Neutron and X-ray diffraction structure of a dihydride imine ditantalum complex. AB - Reactions of Ta(NMe2)5 with D2SiR'Ph (R' = Me, Ph) were found to give a dideuteride eta 2-imine complex (Me2N)3Ta(mu-D)2(mu-N-eta 2-N,C CH2NMe)Ta(NMe2)3(1-d2) through C-H activation of an amide ligand via beta-H abstraction, and the structure of 1 was confirmed by single crystal neutron and X ray diffraction. PMID- 12120380 TI - Direct conversion of macrocyclic furans into macrocyclic isothiazoles. AB - The first calixhetarenes with more than one heteroatom in the constituent rings are prepared in one step by treatment of calix[4]furan 1a and calix[6]furan 1b with ethyl carbamate, thionyl chloride and pyridine to give 2, 3, 4 and 5, 6, 7 respectively; these products have been characterised by X-ray crystallography which reveals that in 2 all eight heteroatoms lie on one face of the macrocyle. PMID- 12120381 TI - Synthesis of (chlorovinyl)cobaloxime complexes, model complexes of proposed intermediates in the B12-catalyzed dehalogenation of chlorinated ethylenes. AB - (Chlorovinyl)cobaloxime complexes, (dmgH)2(py)CoR (R = cis-dichlorovinyl, trans-2 chlorovinyl or 1-chlorovinyl, dmgH = dimethylglyoximate) were synthesized from the reaction of chlorinated ethylenes with in situ generated (dmgH)2Co(I) in the presence of pyridine. PMID- 12120382 TI - Ion-neutral complexes formation and 1,3-proton transfer in the chemical ionization of alkylcyclohexyl benzoates. AB - CI, CID, labelling experiments and DFT calculations are used for the elucidation of the mechanism for the decomposition of cyclohexyl benzoates, which proceeds through 1,3-H shift and two equilibrating ion-neutral complexes. PMID- 12120383 TI - Dendritic chiral auxiliaries on silica: a new heterogeneous catalyst for enantioselective addition of diethylzinc to benzaldehyde. AB - This article reports the first work on the use of silica supported dendritic chiral auxiliaries for the enantioselective addition of diethylzinc to benzaldehyde: the control of dendrimer propagation on the silica surface is of prime importance to obtain enhanced conversion, selectivity, and enantioselectivity. PMID- 12120384 TI - Electrochemical synthesis of fully sulfonated n-dopable polyaniline: poly (metanillic acid). AB - Electrochemical homopolymerization of metanillic acid has been achieved for the first time using a 4:1 acetonitrile-water mixture to get 100% sulfonated polyaniline which is soluble in both organic and aqueous solvents, electrically conducting, and is n-dopable. PMID- 12120385 TI - pH-regulated formation of amyloid-like beta-sheet assemblies from polyglutamate grafted polyallylamine. AB - A novel artificial protein with simple primary structure, poly(gamma-methyl-L glutamate)-grafted polyallylamine, has been prepared and the resultant peptide has shown a unique property of pH-regulated conformation and morphology. PMID- 12120386 TI - Catalytic asymmetric synthesis of O-acetyl cyanohydrins from KCN, Ac2O and aldehydes. AB - A (salen)titanium catalyst has been found to induce the asymmetric addition of potassium cyanide and acetic anhydride to aldehydes, giving enantiomerically enriched cyanohydrin esters with up to 92% enantiomeric excess using just 1 mol% of the catalyst. This is the first report of the asymmetric synthesis of cyanohydrin derivatives using a cyanide source which is non-volatile and inexpensive. PMID- 12120387 TI - Deracemisation and stereoinversion of alpha-amino acids using D-amino acid oxidase and hydride reducing agents. AB - The deracemisation and stereoinversion of both cyclic and acyclic DL-alpha-amino acids, using porcine kidney D-amino acid oxidase (DAAO) and a hydride reducing agent (NaCNBH3-NaBH4), has been investigated. PMID- 12120388 TI - Diels-Alder chemistry of 2-diethoxyphosphinylcyclohex-2-enones. A new approach to complex phosphonates and synthetic applications of the beta-keto phosphonate system. AB - Enone phosphonates 1 and 2 were found to be excellent dienophiles for the Diels Alder reaction, giving phosphonate-containing polycycles, and the phosphonate group of the resulting adducts facilitated both the installation of an angular alkyl group via a reductive alkylation process and the regioselective generation of a ring junction double bond via an intramolecular Wadsworth-Horner-Emmons reaction. PMID- 12120389 TI - A new and short method for the synthesis of 2,4-methanoproline. AB - 2,4-Methanoproline, a supposed non-proteinogenic anti-feedant, was synthesised in 5 steps starting from allyl benzyl ether 3 in 10% overall yield with an intramolecular nucleophilic substitution as the key step for the formation of the bicyclic skeleton. PMID- 12120390 TI - Structure and exchange in silicon-linked tetraradicals. AB - The EPR spectroscopy, crystallography, and magnetic susceptibility of tetrakis(N oxyl-2,2,6,6-tetramethylpiperidin-4-yl)silane and tetrakis(4-N-tert-butyl-N aminoxylphenyl)silane show that silicon acts as a weak intramolecular exchange linker for polynitroxides, although both tetraradicals show onset of inter-spin exchange at reduced temperatures. PMID- 12120391 TI - A novel route for synthesis of gamma-butyrolactone through the coupling of hydrogenation and dehydrogenation. AB - A coupling process of the hydrogenation of maleic anhydride and the dehydrogenation of 1,4-butanediol has been invented for the synthesis of gamma butyrolactone over a Cu-Zn catalyst, realizing optimal hydrogen utilization and better energy efficiency. PMID- 12120392 TI - New layered manganese oxide halides. AB - The first layered manganese(III) oxide chlorides, Sr2MnO3Cl and Sr4Mn3O8-yCl2, have been synthesised; Sr2MnO3Cl adopts a K2NiF4 type structure with sheets of MnO5 square based pyramids linked through oxygen and separated by SrCl layers; it is the end member of a new family of Ruddlesden-Popper type manganese oxide halides which includes the three-layer member Sr4Mn3O8-yCl2 also reported herein. PMID- 12120393 TI - 1-Allyl-3-propylthiourea modified mesoporous silica for mercury removal. AB - 1-Allyl-3-propylthiourea modified mesoporous silica has high adsorption capacity for mercury ions and its regeneration can be accomplished by washing with 10% thiourea in aqueous 0.05 M HCl. PMID- 12120395 TI - A simple route towards tubular ZnO. AB - Thermal treatment of Zn(NH3)(4)2+ precursor in ethanol solvent led to the formation of the tubular ZnO which exhibited strong ultraviolet photoluminescence around 385 nm at room temperature; TEM images showed the hollow tubules with approximately 450 nm in diameter and approximately 4 microns in length were built up by ZnO polycrystals. PMID- 12120394 TI - Selective phosphatidylethanolamine translocation across vesicle membranes using synthetic translocases. AB - Two sulfonamide derivatives of tris(aminoethyl)amine selectively facilitate the translocation of a fluorescent phospholipid probe containing the phosphoethanolamine head-group across vesicle membranes. PMID- 12120396 TI - The acylation of an acyl complex resulting in a labile OCO tridentate ligand. AB - The reaction of propionic anhydride with [fac-Ru(C(O)Et)-(CO)2(H2O)3][CF3SO3] produces a new propylidin dipropionato group, which behaves as a tridentate ligand giving the neutral complex Ru(CEt(OC(O)Et)2)(CO)2(CF3SO3). PMID- 12120397 TI - A modular approach to porphyrin oligomers using metal ions as connectors. AB - Porphyrins bearing two external coordination sites allowed the stepwise preparation of polymetallic oligomers connected by metal centers. PMID- 12120398 TI - Highly efficient Grignard-type imine additions via C-H activation in water and under solvent-free conditions. AB - A highly effective Cu-Ru catalyzed addition of terminal alkynes to imines via C-H activation has been achieved in water or under solvent-free conditions. PMID- 12120399 TI - Novel photoreaction of N-alkyl(p-methoxyphenyl)arylamines assisted by protic acids. AB - A novel photochemical transformation from N-alkyl(p-methoxyphenyl)arylamines (1a 1f) to 1,2,4-trihydro(4aH)-carbazol-3-ones (2a-2f) is reported with the assistance of protonation at the dihydrocarbazole intermediate followed by sequential formal [1,5]hydrogen, [1,3]hydrogen shifts and proton assisted hydrolysis. PMID- 12120400 TI - Site-selective mono-titanation of conjugated diynes with a Ti(II) alkoxide reagent. Concise preparation of stereo-defined enynes and dienynes. AB - Conjugated diynes underwent selective mono-titanation with a Ti(II) reagent to give 1:1 diyne-titanium alkoxide complexes, which reacted with proton, aldehyde, and another acetylene to give stereo-defined enynes, enynols, and dienynes. PMID- 12120402 TI - Bimetallic nanoparticles aligned at the tips of carbon nanotubes. AB - Cluster-derived bimetallic nanoparticles have been deposited onto multi-wall carbon nanotubes and shown to be generally homogeneously dispersed, of uniform small sizes, of the same composition as the starting mixed-metal clusters, and to have a tendency to align at the tips of the tubules. PMID- 12120401 TI - Electrochemical activation of carbon dioxide in ionic liquid: synthesis of cyclic carbonates at mild reaction conditions. AB - Electrocatalytic cycloaddition of carbon dioxide to epoxides in room temperature ionic liquids as reaction media without any additional supporting electrolyte and catalyst could be conducted with high to excellent performances under mild conditions. PMID- 12120403 TI - Synthesis of bent [4]phenylene (cyclobuta[1,2-a:3,4-b']bisbiphenylene) and structure of a bis(trimethylsilyl) derivative: the last [4]phenylene isomer. AB - The syntheses of the title compounds were accomplished by cobalt-catalyzed alkyne cyclotrimerizations using two strategies; the properties of the bent phenylene frame reflect the combined effects of benzocyclobutadienofusion of the component [3]phenylene substructures. PMID- 12120404 TI - Stereoselective intramolecular hydrogen abstraction by a chiral benzophenone derivative. AB - An unprecedented stereoselective photoreduction of a chiral BZP is observed in steady state as well as in time-resolved studies. PMID- 12120405 TI - The effect of delta lambda chirality on molecular organization in two-dimensional films of a Ru(II) complex with a mesogenic ligand. AB - A novel amphiphilic Ru(II) complex, [Ru(acac)2L] (acac = acetylacetonato, L = 5,5'-bis(4-octylphenyloxycarbonyl)-2,2'-bipyridyl), in which L undergoes SmC, SmA and N liquid crystal phases, exhibits a remarkable chirality effect on its monolayer state: that is, a racemic mixture gives a monolayer consisting of spike like aggregates of 1.2 nm (in height) x 50 nm (in diameter), whereas the delta enantiomer gives a uniform monolayer. PMID- 12120406 TI - Selective anion effects in chiral complexes of iridium via diffusion and HOESY data: relevance to catalysis. AB - 1H and 19F Pulsed Gradient Spin Echo (PGSE) diffusion data, together with 1H, 19F HOESY results are shown to distinguish between different types of anion/cation interactions in chiral dihydrido-P,N-complexes of Ir(III); in CD2Cl2 the diffusion coefficients, D, for the BArF and the Ir-cation suggest ion-pairing whereas the D-values for PF6-reveal independent motion; the PF6- approaches the cation via a specific pathway; the combined PGSE/HOESY approach offers a unique opportunity for exploring anion effects in organometallic/catalytic chemistry. PMID- 12120407 TI - A novel catalyst for hydrazine decomposition: molybdenum carbide support on gamma Al2O3. AB - An alumina-supported Mo2C catalyst is found to be as active as a conventionally used Ir/gamma-Al2O3 catalyst for catalytic decomposition of hydrazine tested in a monopropellant thruster. PMID- 12120408 TI - From the analyst's couch. Trends in development cycles. PMID- 12120409 TI - Combinatorial informatics in the post-genomics ERA. AB - The multitude of potential drug targets emerging from genome sequencing demands new approaches to drug discovery. A chemogenomics strategy, which involves the generation of small-molecule compounds that can be used both as tools to probe biological mechanisms and as leads for drug-property optimization, provides a highly parallel, industrialized solution. Key to the success of this strategy is an integrated suite of chemi-informatics applications that can allow the rapid and directed optimization of chemical compounds with drug-like properties using 'just-in-time' combinatorial chemical synthesis. An effective embodiment of this process requires new computational and data-mining tools that cover all aspects of library generation, compound selection and experimental design, and work effectively on a massive scale. PMID- 12120411 TI - Cytochrome P450 enzymes in the generation of commercial products. AB - Cytochrome P450 enzymes are remarkably diverse oxygenation catalysts that are found throughout nature. Although most of the interest in the pharmaceutical industry has focused on the role of cytochrome P450s in drug development, these enzymes also offer potential in the discovery not only of drugs, but also of other useful chemicals. Potential applications range from the use of cytochrome P450s as drug targets, to the use of randomly generated mutants of cytochrome P450s to produce libraries of new chemicals and drugs. PMID- 12120410 TI - New therapeutics that modulate chemokine networks. AB - Chemokines are small cytokines that control a wide variety of biological and pathological processes, from immunosurveillance to inflammation, and from viral infection to cancer. The numerous known chemokine receptors have given hope that selective receptor antagonism might be possible, which could allow us to control which cells are recruited and activated at any time and in any place. As chemokine receptors are G-protein-coupled receptors, which are classical targets for the pharmaceutical industry, it is hoped that chemokines could be the first cytokines for which small-molecule receptor antagonists could be developed. Recently, reports of chemokine-receptor antagonists, both in vitro and in animal models of disease, have been published. It is anticipated that this field could produce clinically useful therapies in the next few years. PMID- 12120412 TI - SOD mimetics are coming of age. AB - The list of pathophysiological conditions that are associated with the overproduction of superoxide anions expands every day. The most exciting realization is that there seems to be a similarity between the tissue injury that is observed in various disease states, as superoxide anions produce tissue injury and associated inflammation in all tissues in similar ways. Tissue injury and inflammation form the basis of many disease pathologies, including ischaemia and reperfusion injuries, radiation injury, hyperoxic lung damage and atherosclerosis. This commonality provides a unique opportunity to manipulate numerous disease states with an agent that removes superoxide anions. PMID- 12120413 TI - Potential cardioprotective actions of no-releasing aspirin. AB - The use of low doses of aspirin on a daily basis has increased greatly in the past 20 years, based on observations that it can significantly reduce the risk of heart attacks and strokes. However, aspirin can also cause severe damage to the stomach. A modified version of aspirin that releases nitric oxide has been developed that seems to offer important advantages over its 103-year-old parent- namely, improved protection for the heart without the unwanted effects on the stomach. PMID- 12120414 TI - Telomere maintenance as a target for anticancer drug discovery. AB - Maintenance of telomeres--specialized complexes that protect the ends of chromosomes--is undertaken by the enzyme complex telomerase, which is a key factor that is activated in more than 80% of cancer cells that have been examined so far, but is absent in most normal cells. So, targeting telomere-maintenance mechanisms could potentially half tumour growth across a broad spectrum of tumour types, with little cytotoxic effect outside tumours. Here, we describe the current understanding of telomere biology, and the application of this knowledge to the development of anticancer drugs. PMID- 12120415 TI - New safe medicines faster: a proposition for a pan-European research effort. AB - Providing support for downstream drug development has not traditionally been a primary focus of public research funding programmes. However, the European Commission has decided to include the drug development process in the European Union's Sixth Framework Programme for Research and Technological Development (2002-2006). If the present proposal is adopted, research consortia and networks must be ready to exploit the proposed funds. 'Integrated Projects' and 'Networks of Excellence' will allow complex topics to be tackled simultaneously by many research groups and associated stakeholders, but these will be very demanding to implement. PMID- 12120416 TI - A bridge between academia and the drug discovery pipeline. PMID- 12120417 TI - Effects of free fatty acids on insulin secretion in obesity. AB - The prevalence of obesity in Western society has reached epidemic proportions and its aetiological role in the development of type 2 diabetes has made finding an effective treatment for the condition of crucial importance. Of the many consequences of obesity, derangements in glucose metabolism present one of the greatest problems to health. While the role of obesity in causing insulin resistance has received much attention, the effect of obesity on beta-cell failure and the consequent development of type 2 diabetes requires re-emphasis. In this review, the current understanding of the effects of elevated free-fatty acids on beta-cell function will be examined, including a discussion of potential mechanisms. In particular, dysregulation of biochemical pathways and alterations in key enzymes, proteins and hormones will be considered as grounds for the progression to a diabetic phenotype. PMID- 12120418 TI - Metabolically active components of fat-free mass and resting energy expenditure in humans: recent lessons from imaging technologies. AB - Imaging technologies, i.e. magnetic resonance imaging (MRI), computer tomography (CT) and dual-energy X-ray absorptiometry (DEXA), are precise and accurate techniques used to study lean body mass and adipose tissue distribution. CT and MRI can also be used to assess metabolically active components of fat-free mass (FFM). (Throughout this article, metabolic activity is defined with respect to oxidative metabolism.) To date a total of 116 in vivo measurements of organ masses (OM), in combination with the measurement of resting energy expenditure (REE), have been reported. These data suggest that MRI- or CT-derived OM explains part (approximately 5-10%) of the interindividual variance in REE. The data also suggest that REE can be reconstructed from detailed body composition analysis. Calculating REE from the sum of individual OM multiplied by a constant organ tissue-respiration rate showed a high correlation between calculated and measured REE, with only small and non-significant differences of 83-96 kJ d-1. In addition to CT- and MRI-derived OM, data are available of 244 obese and non-obese subjects regarding the association between regional components of lean body mass (LBM, assessed by DEXA) and REE. These results suggest that measurement of LBM distribution also provides the opportunity to adjust for the non-linearity of REE on body mass. Assessment of metabolically active components of FFM or LBM may also add to our understanding of malnutrition-, obesity- and disease states related variance in REE. There is need for (1) standardization of imaging technology in body composition research; (2) reference data on detailed body composition, also including more recent autopsy data; (3) reducing the number of assumptions in model-based predictions; and (4) a combination of imaging technologies with in vivo measurements of individual OM respiration. PMID- 12120419 TI - A review of the literature of Bardet-Biedl disease and report of three cases associated with metabolic syndrome and diagnosed after the age of fifty. AB - Bardet-Biedl syndrome (BBS) is a genetic autosomal-recessive disease (formerly grouped with Laurence-Moon-Biedl syndrome but considered today as a separate entity) characterized by abdominal obesity, mental retardation, dysphormic extremities (syndactyly, brachydactyly or polydactyly), retinal dystrophy or pigmentary retinopathy, hypogonadism or hypogenitalism (limited to male patients) and kidney structural abnormalities or functional impairment. The expression and severity of the various clinical BBS features show inter- and intrafamilial variability. This study focuses on three cases of familial BBS--two sisters and one brother (66, 64 and 51 years of age, respectively)--with the main cardinal findings of the disease plus a classic 'metabolic syndrome' (characterized by abdominal obesity, atherogenic dyslipidaemia, raised blood pressure, insulin resistance with or without glucose intolerance, and prothrombotic risk and proinflammatory states). One female patient (not affected by reproductive dysfunction) had three healthy offspring, while the other two patients were unmarried. Another severely affected brother died at 70 years of age; two other brothers are lean but affected by nephropathy, retinopathy, slight mental retardation, polydactyly, hypertension and thrombotic diseases, and had healthy offspring. BBS is a rather rare but severe syndrome that is often mis- or undiagnosed. Ophthalmologists, endocrinologists and nephrologists should be aware of BBS because of its adverse prognosis--early onset of blindness, associated findings of metabolic syndrome and increased vascular risk, and severe renal impairment (the most frequent cause of reduced survival and death early in life). PMID- 12120420 TI - Dietary fat is a major player in obesity--but not the only one. PMID- 12120421 TI - Dietary fat plays a major role in obesity: no. AB - The percentage of dietary energy from fat has been suggested to be an important determinant of body fat, and this presumed effect has been invoked to justify the general promotion of low-fat diets. Dietary fat and the prevalence of obesity are lower in poor countries than in affluent countries. However, these contrasts are seriously confounded by differences in physical activity and food availability; within areas of similar economic development, per capita intake of fat and the prevalence of obesity have not been positively correlated. Randomized trials are the preferable method for evaluating the effect of dietary fat on adiposity because they avoid problems of confounding that are difficult to control in other studies. In short-term trials, a small reduction in body weight is typically seen in individuals randomized to diets with a lower percentage of calories from fat. In a meta-analysis of these trials, it was estimated that a decrease in 10% of energy from fat would reduce weight by 16 g d-1, which would correspond to a 9-kg weight loss by 18 months. However, compensatory mechanisms appear to operate because in trials lasting one year or longer, fat consumption within the range of 18-40% of energy has consistently had little, if any, effect on body fatness. Moreover, within the United States (US), a substantial decline in the percentage of energy from fat during the last two decades has corresponded with a massive increase in obesity, and similar trends are occurring in other affluent countries. Diets high in fat do not account for the high prevalence of excess body fat in Western countries; reductions in the percentage of energy from fat will have no important benefits and could further exacerbate this problem. The emphasis on total fat reduction has been a serious distraction in efforts to control obesity and improve health in general. PMID- 12120422 TI - From instinct to intellect: the challenge of maintaining healthy weight in the modern world. AB - The global obesity epidemic is being driven in large part by a mismatch between our environment and our metabolism. Human physiology developed to function within an environment where high levels of physical activity were needed in daily life and food was inconsistently available. For most of mankind's history, physical activity has 'pulled' appetite so that the primary challenge to the physiological system for body weight control was to obtain sufficient energy intake to prevent negative energy balance and body energy loss. The current environment is characterized by a situation whereby minimal physical activity is required for daily life and food is abundant, inexpensive, high in energy density and widely available. Within this environment, food intake 'pushes' the system, and the challenge to the control system becomes to increase physical activity sufficiently to prevent positive energy balance. There does not appear to be a strong drive to increase physical activity in response to excess energy intake and there appears to be only a weak adaptive increase in resting energy expenditure in response to excess energy intake. In the modern world, the prevailing environment constitutes a constant background pressure that promotes weight gain. We propose that the modern environment has taken body weight control from an instinctual (unconscious) process to one that requires substantial cognitive effort. In the current environment, people who are not devoting substantial conscious effort to managing body weight are probably gaining weight. It is unlikely that we would be able to build the political will to undo our modern lifestyle, to change the environment back to one in which body weight control again becomes instinctual. In order to combat the growing epidemic we should focus our efforts on providing the knowledge, cognitive skills and incentives for controlling body weight and at the same time begin creating a supportive environment to allow better management of body weight. PMID- 12120423 TI - Long-term weight development after pregnancy. AB - For some women pregnancy is a trigger for developing overweight and obesity. Seventy-three per cent of 128 female patients at our Obesity Unit indicated that they had retained more than 10 kg after each of their pregnancies, and for this subgroup weight development after pregnancy was of crucial importance for their future health. Although mean weight increases after pregnancy generally are modest, there are wide individual variations. In studies at the Obesity Unit, weight retention ranging from up to 26.5 kg one year after pregnancy to a loss of 12.3 kg was reported, although the mean weight gained was only 0.5 kg. Numerous studies have analysed factors explaining weight development after pregnancy and delivery, with a range of subjects from several hundred thousand women to fewer than one-hundred, but overall it has been surprisingly difficult to identify strong predictors of weight development. Numerous confounders have been identified; in a review up to 31 such confounders were reported. Methodological problems include weight development over time also in non-pregnant women and problems of identifying the optimal time-point when the overall impact of the pregnancy on weight development should be evaluated. Lactation has consistently been found to play a small role in explaining weight retention up to one year after delivery. Few studies have examined the role of physical activity during pregnancy and after delivery to explain weight development. Our own ongoing follow-up of women who gave birth during 1984-85, the so-called SPAWN (Stockholm Pregnancy and Women's Nutrition) study, illustrates that 15 years after delivery, a significant proportion of the 1423 women initially studied were available for re-examination. Drop-out analyses indicate that for most variables under study, the remaining women were representative for the initial sample. Pregnancy and weight development are intertwined in a complex pattern, which includes a change in lifestyle factors, such as eating behaviour, physical activity, smoking cessation and degree of lactation, but which are still not fully understood. PMID- 12120424 TI - Obesity, haemostasis and the fibrinolytic system. AB - Obese patients are at risk for the development of cardiovascular diseases, which can in part be explained by disturbances in the haemostatic and fibrinolytic systems. Indeed, obese subjects tend to have higher values of fibrinogen, factor VII, factor VIII, von Willebrand factor and plasminogen activator inhibitor compared to non-obese subjects. Abdominal obesity, in particular, has been shown to be associated with disturbances in fibrinogen, factor VIII and von Willebrand factor, while less consistent results have been found for factor VII. Recently it has been demonstrated that the adipocyte itself is able to produce plasminogen activator inhibitor-1, possibly explaining the high levels found in obesity. Different studies have investigated the association between haemostatic and fibrinolytic parameters and the insulin resistance syndrome, often present in obese subjects. Fibrinogen has been found to be related to insulin, but it has been suggested that this relationship is not independent of the accompanying inflammatory reaction. Results from studies on the relationship between insulin resistance and factor VII, factor VIII and von Willebrand factor levels are inconsistent. In contrast, plasminogen activator inhibitor-1 has been found to correlate with all components of the insulin resistance syndrome, and can be considered as a true component of this metabolic syndrome. Weight loss seems to have a beneficial effect on factor VII--probably mediated through a reduction in triglycerides. Data on factor VIII and von Willebrand factor are scarce but weight loss does not seem to have an effect. Fibrinogen does not seem to be reduced by modest weight loss and a more substantial weight loss seems necessary to lower fibrinogen levels. In contrast, both modest and substantial weight loss have been found to significantly reduce plasminogen activator inhibitor-1 levels. In conclusion, the increased cardiovascular risk observed in obesity could in part be explained by the association between insulin resistance and components of the fibrinolytic and haemostatic systems. Whether this relationship is truly causal or indirect needs to be elucidated further. PMID- 12120425 TI - Medicare pay raise likely. But drug benefit probably won't pass this year. PMID- 12120426 TI - Playing it cool. Tenet keeps calm, will pay $56 million to settle billing fraud allegations at hospitals. PMID- 12120428 TI - Surmounting the cost barrier. To help the uninsured, government must cut mandates, subsidize private plans. PMID- 12120427 TI - Drugmakers quiet in GPO drama. But devicemakers push senate to enact changes. PMID- 12120429 TI - The Boston cleanup. Paul Levy earned a reputation by helping to restore the city's polluted harbor. Now his project is Beth Israel Deaconess. PMID- 12120430 TI - Stocking up on savings. Through supply-side automation and improvements in the purchasing process, a New York healthcare system has slashed millions in costs. Even more is on the way. PMID- 12120431 TI - Straight talk: new approaches in health care. HIPAA: deadlines are looming. Are providers prepared? AB - This is the fourth installment in a series of group discussions by top executives on key issues in healthcare today. Modern Healthcare and PricewaterhouseCoopers present Straight Talk. This session tackles the Health Insurance Portability and Accountability Act of 1996, or HIPAA, and where providers are today in the compliance process and where they need to go. The discussion was held on June 4, 2002 at Modern Healthcare's Chicago headquarters. The moderator was Jeffrey P. Fusile, Healthcare Consulting Partner with PricewaterhouseCoopers, Atlanta. The act protects consumers' health-insurance coverage after job changes. It also mandates significant modifications in the way providers handle the submission of claims and other related transactions and provides protection for the privacy and security of patients' health information. The law requires providers to comply with regulations governing electronic transactions and code sets by October 2003- assuming they file for an extension by October 2002--and privacy regulations by April 2003. The security compliance date has not yet been determined, but it is widely agreed that much of the security rules' requirements will be necessary to honor an organization's privacy commitments in April 2003. PMID- 12120433 TI - AMA approves plans to become umbrella group. Now here's the hard part: AMA must get support of specialty societies to influence Washington. PMID- 12120432 TI - A matter of financial faith. Roman Catholic megasystem Ascension inks merger deal with smaller not-for-profit seeking to preserve its charitable mission. AB - Roman Catholic giant Ascension Health is showing that not only investor-owned chains can offer a financial sanctuary to a troubled hospital. Last week, the nation's largest not-for-profit system agreed to absorb Carondelet Health System and its eight remaining hospitals. PMID- 12120434 TI - The cutaneous manifestations of HIV infection. AB - Dermatologic disease is extremely common and varied in HIV-infected patients. While some cutaneous findings are nearly exclusive to HIV-seropositive individuals, many are found in the general population. However, HIV-infected individuals often have an increased prevalence or severity, atypical presentations, or difficulty with treatment of the disease. Immune reconstitution with HAART significantly reduces the prevalence of many dermatologic diseases, but also has associated cutaneous side effects. Correct and early diagnosis of skin disease in HIV-infected individuals allows for early management and improved quality of life. Because dermatologic manifestations may be the first clue of HIV infection, offering HIV testing to affected individuals can lead to early diagnosis and treatment of HIV infection and, ideally, a decrease in disease progression and transmission. PMID- 12120435 TI - A review of measles virus. AB - Measles virus is an epidemic disease with a worldwide distribution. Since the development of the live attenuated vaccine, the incidence of reported measles cases has declined by greater than 99% in the United States. Measles causes a systemic illness manifested by a characteristic prodrome and pathognomonic rash. Although usually a self-limited disease, measles can cause severe complications, especially in adults and the immunocompromised. We discuss a vaccination schedule for preschool and school-aged children. Recent research has demonstrated clinical benefit in patients with severe measles virus infections that are treated with ribavirin and vitamin A supplementation. PMID- 12120436 TI - Enterovirus infections: a review of clinical presentation, diagnosis, and treatment. AB - The enteroviruses, RNA viruses of the Picornaviridae family, are ubiquitous pathogens which include more than 70 different serotypes that infect people of all ages and tend to occur seasonally in the summer and fall. Clinical manifestations may vary diversely with one serotype, while multiple serotypes can present with identical symptoms and may mimic bacterial infections. Most enterovirus infections cause benign, self-limiting disease; however, they can also produce severe and sometimes fatal illnesses such as meningitis, encephalitis, myocarditis, neonatal sepsis, and polio. Severe enterovirus infections are being diagnosed and treated earlier with better prognostic outcomes due to the advances of polymerase chain reaction technology in accurately detecting virus in patient fluids as well as the recent development of new antiviral therapies. PMID- 12120437 TI - Rubella. AB - Rubella is still a public health concern in this country. With the great numbers of immigrants welcomed in the United States also comes the risk of diseases that are little known or thought to be of little concern. Obviously, there is a need for continued vaccination of citizens, not only at approximately 1 year of age and school age but also in terms of continued screening for and immunization of susceptible women of childbearing age. Policies need to be developed to address the issue of rubella infection and susceptibility in areas where there is a high population density of people from countries that may not have such strict immunization practices. Vaccination at workplaces that employ high numbers of foreign-born workers or that involve a close working environment, education of workers and health care professionals in the recognition of rubella and its potentially devastating effects, and continued surveillance for and reporting of cases in high-risk areas may be of benefit. PMID- 12120438 TI - The cutaneous manifestations of viral hepatitis. AB - Viral hepatitis is a common disease with significant morbidity and mortality that is found throughout the world. It is a known cause of acute and chronic liver disease. Because the disease can be asymptomatic in its acute stages, the diagnosis is often missed. This article reviews the extrahepatic manifestations of the disease, in particular the visible cutaneous lesions, which offer clues to the diagnosis of viral hepatitis. Familiarity with the most common skin lesions associated with each type of hepatitis virus will be useful in identifying previously undiagnosed individuals. This article serves as a review of the most widely published associations between skin diseases and specific subtypes of viral hepatitis. PMID- 12120439 TI - Herpes simplex viruses 1 and 2. AB - An increased understanding of the pathogenesis and transmission of HSV infections and the development of sensitive type-specific diagnostic tests have helped develop effective prophylactic and therapeutic antiviral drug regimens. Effective medications have been available for quite some time, but the most optimal regimens are still under investigation. Advances in the treatment of atypical presentations of HSV infection (such as the use of cidofovir gel for the treatment of acyclovir-resistant HSV) are promising. Newer treatments, such as resiquimod, actually may alter the course of HSV infection, reducing the severity and frequency of recurrences. Vaccines are being explored as preventive and therapeutic measures against HSV. PMID- 12120440 TI - Clinical manifestations of varicella-zoster virus infection. AB - Infections by VZV, the virus that causes chickenpox and herpes zoster, usually are diagnosed by the classic clinical presentations. In immunocompromised patients, however, the atypical presentation can make the diagnosis more challenging. Although varicella typically follows an uncomplicated course in children, adults and immunocompromised patients can develop complications involving several organs; some complications may be fatal. Prevention of disease with the vaccine is ideal. When varicella or zoster infection does occur, proper treatment should be initiated, depending on the age and immune status of the patient. PMID- 12120441 TI - Cutaneous manifestations of Epstein-Barr virus infection. AB - The most common cutaneous manifestations of EBV include IM, OHL, and cutaneous lymphoproliferative disorders. Infectious mononucleosis is a self-limited manifestation of acute EBV infection. The transient rash that occurs quite commonly in patients with IM who have received antibiotic therapy is an erythematous, maculopapular eruption, usually located on the trunk and upper extremities. Oral hairy leukoplakia occurs in immunosuppressed HIV-positive and HIV-negative individuals. In HIV-positive individuals, it serves as an indicator of disease severity and rapid progression to AIDS. The presence of OHL in an individual should prompt the clinician to perform a through history-taking and investigation of immune status. Cutaneous lymphoproliferative disorders associated with EBV occur in individuals with congenital or acquired immunodeficiency syndromes. PMID- 12120442 TI - Cytomegalovirus infections. AB - Over the past two decades, there has been an escalation in the number of patients undergoing immunosuppressive therapy following solid organ or bone marrow transplantation, as well as a dramatic increase in the incidence of AIDS. As a result, human cytomegalovirus (HCMV)--once considered a neonatal disease--has captured great interest and importance as a major pathogen in both immunocompromised and immunocompetent patients. Like other members of herpesviridae family, HCMV establishes latency in myeloid lineage cells with potential for reactivation. The natural history of HCMV infection can be divided into primary infection, latency, and reinfection. This review article briefly discusses the molecular pathogenesis of HCMV, then focuses on the clinical picture of this disease, with emphasis on the skin pathology. Diagnostic methods and treatments are also discussed. PMID- 12120443 TI - Human herpesviruses 6 and 7. AB - Human herpesviruses 6 and 7 are newly discovered viruses that belong to the genus Roseolavirus within the subfamily Betaherpesvirinae. These ubiquitous viruses may cause primary or chronic persistent infection or remain in a state of latency for many years, until a decrease in the immunologic state of the host leads to reactivation of infection. Several diseases have been linked with HHV-6 and HHV 7. In the dermatologic arena, a definite association has been proven only for HHV 6 and exanthema subitum (roseola infantum), whereas the role of HHV-7 in the pathogenesis of pityriasis rosea remains a matter of debate. PMID- 12120444 TI - Human herpesvirus 8. AB - Human herpesvirus 8 (HHV 8), also known as Kaposi's sarcoma-associated herpesvirus (KSHV) is a g2 herpesvirus and the most recently identified human tumor virus. HHV 8 has been consistently implicated in the pathogenesis of all clinical variants of Kaposi's sarcoma, as well as in the plasma cell variant of multicentric Castleman's disease and primary effusion lymphomas. Pathogenicity of the virus is increased in the host who is immunosuppressed, either iatrogenically or through H1V-1 infection. The HHV 8 genome contains several homologues of cellular genes that regulate cell growth and differentiation, and the exact mechanisms of the virus' oncogenicity using molecular piracy are still being investigated and elucidated. In this article, the authors review the epidemiology, transmission, clinical manifestations, and molecular genetics of HHV 8 infection and provide a summary of the current treatment modalities available to the clinician. PMID- 12120445 TI - Human papillomavirus: a review. AB - Human papillomavirus infection remains a great source of morbidity and mortality. Progress in understanding the structure of HPV and its pathogenesis has led to a wide variety of possible new treatment modalities to combat HPV-related disease. Most HPV infections (whether high risk or low risk) resolve without any medical intervention. Persistent or progressive disease, however, remains difficult to treat. Although currently available therapies have proved efficacious and tolerable in the treatment of nongenital and genital warts, no single therapy is uniformly effective in eradicating persistent HPV infection. Cytodestructive methods, such as cryotherapy, remain the primary treatment modality for nongenital warts. Immune response modifiers, such as imiquimod, currently show the greatest promise in treating HPV-induced anogenital lesions, both with respect to complete response and in preventing recurrence. Human papillomavirus infection is one of the most common sexually transmitted diseases in the world, and cervical cancer still causes significant morbidity and mortality. Pap smear tests have greatly reduced the incidence and mortality of cervical cancer in developed countries. Additional research will focus on primary and secondary prevention strategies. Vaccines against high-risk HPV types are promising modalities currently under investigation to prevent HPV infections and possibly to treat them. PMID- 12120446 TI - Parvovirus B19: a review. AB - Infection with parvovirus B19 may result in a wide range of dermatologic manifestations. The specific skin findings include erythema infectiosum and papular purpuric "gloves-and-socks" syndrome. The nonspecific findings include reticular erythema, maculopapular eruptions, and petechiae and purpura, as well as other less frequently described findings. Associations with other dermatologic diseases, such as erythema multiforme and erythema nodosum, also have been described. A role in the pathogenesis of various collagen vascular disorders has been suggested and is under investigation. The diagnosis of infection rests on the typical clinical findings. Whenever parvovirus B19 infection is diagnosed, the physician must ensure that neither the patient nor his or her contacts is a member of certain vulnerable populations. In these populations, infection with parvovirus B19 may result in devastating complications. The vulnerable populations include those with hematologic disease, immunosuppressed patients, and pregnant women. Treatment of infection in the healthy immunocompetent individual is asymptomatic, and the acute infections typically resolve without complications. PMID- 12120447 TI - A review of the dermatologic manifestations of poxvirus infections. AB - Physicians need to be familiar with the presentation of poxvirus infections. The poxvirus infections that are common, such as MCV, are rarely serious; however, physicians need to understand them because they can be bothersome to patients and require reassurance and, if available, treatment. The more rare poxvirus infections, such as variola, need to be recognized because they are generally serious. It is important to consider that these infections can suggest underlying systemic disease, as in the case of severe, recalcitrant MCV infection, which may be indicative of HIV infection or another immunocompromised state. PMID- 12120448 TI - Management of pyoderma gangrenosum. AB - The management of pyoderma gangrenosum (PG) requires a structured approach to establishing diagnosis of the disease and assessment of the patient. Clinical management of active PG lesions should be carried out in coordination with other specialists (such as nurses and pain managers) and often necessitates a flexible, innovate attitude to therapy, because the needs of individual patients may vary widely. Although there is no single successful treatment for this disease, certain types of PG lesions are recognized to respond more readily to accepted therapies than others. We outline guidelines to the management of the patient with PG and discuss alternative therapies. PMID- 12120449 TI - New therapeutic options for chronic wounds. AB - Increased research and understanding of acute and chronic wounds has led to the development of new therapies to stimulate and improve healing of difficult wounds. These include various growth factors, animal-derived wound coverings, and bioengineered human skin tissue. PMID- 12120451 TI - Holding on to hope. Police grill a drifter for clues in a vexing kidnap case. PMID- 12120450 TI - Code blue in Jerusalem. PMID- 12120452 TI - Staring across a great divide. PMID- 12120453 TI - Diabolical, haunting terror--here and now. PMID- 12120454 TI - And the doctor shall heal: the Israel-Palestinian conflict. PMID- 12120455 TI - Smallpox: a possible comeback. PMID- 12120456 TI - Medical management of terrorist attacks. PMID- 12120457 TI - Hospital management of a bioterror event. PMID- 12120458 TI - Preparedness of the Israeli health system for a biologic warfare event. AB - The threat of a disease outbreak resulting from biologic warfare has been of concern for the Israeli health system for many years. In order to be prepared for such an event the health system has formulated doctrines for various biologic agents and defined the logistic elements for the procurement of drugs. During the last 4 years, and especially after the West Nile fever epidemic in 2000, efforts to prepare the healthcare system and the relevant organizations were accelerated. The Director-General of the Ministry of Health nominated a Supreme Steering Committee to fill in the gaps and upgrade the preparedness of the health system for an unusual disease outbreak. This committee and its seven subcommittees established appropriate guidelines, communication routes among different organizations, and training programs for medical personnel. The anthrax outbreak in the United States found the healthcare system in the hub of the preparation process, and all modes of action were intensified. Further work by hospitals, primary care clinics and all other institutes should be increased to maintain a state of proper preparedness. PMID- 12120459 TI - The epidemiologic pyramid of bioterrorism. AB - Recent events have drawn world attention to "mythological diseases"--such as anthrax, plague and smallpox--which have been out of the spotlight for some decades. Much of our current knowledge of epidemic intervention and disease prevention was acquired over history through our experience with these diseases, such that the sudden panic over the reemergence of these historically well-known entities is perplexing. Over time, changes in the balance of the epidemiologic triangle have driven each of these disease systems towards a new equilibrium with which we are not familiar. While the pathogens may be similar, these are not the diseases of the past. These new disease systems are insufficiently described by the classic epidemiologic triangle, which lacks a dimension necessary for providing a valid model of the real-world effects of bioterror-related disease. Interactions within the classic epidemiologic triangle are now refracted through the prism of the global environment, where they are mediated, altered, and often amplified. Bioterror-associated diseases must be analyzed through the epidemiologic pyramid. The added dimension represents the global environment, which plays an integral part in the effects of the overall disease system. The classic triangle still exists, and continues to function at the base of the new model to describe actual agent transmission, but the overall disease picture should be viewed from the height of the fourth apex of the pyramid. The epidemiologic pyramid also serves as a practical model for guiding effective interventional measures. PMID- 12120460 TI - Surveillance for early detection and monitoring of infectious disease outbreaks associated with bioterrorism. AB - The appearance of "new" infectious diseases, the reemergence of "old" infectious diseases, and the deliberate introduction of infectious diseases through bioterrorism has highlighted the need for improved and innovative infectious disease surveillance systems. Traditional current surveillance systems are generally based on the recognition of a clear increase in diagnosed cases before an outbreak can be identified. For early detection of bioterrorist-initiated outbreaks, the sensitivity and timeliness of the systems need to be improved. Systems based on syndromic surveillance are being developed using technologies such as electronic reporting and the internet. The reporting sources include community physicians, public health laboratories, emergency departments, intensive care units, district health offices, and hospital admission and discharge systems. The acid test of any system will be the ability to provide analyses and interpretations of the data that will serve the goals of the system. Such analytical methods are still in the early stages of development. PMID- 12120461 TI - Preparation for an outbreak of smallpox in Israel. AB - Because of its high case-fatality rate, its very high transmission potential, and the worldwide shortage of effective vaccine, smallpox tops international lists of over a dozen possible bioterror and biologic warfare agents. In a scenario involving aerosol variola virus release, tens to hundreds of first-generation cases would ensue, as would hundreds to thousands of subsequent cases resulting from person-to-person transmission. A smallpox outbreak in Israel must not be regarded as a doomsday event: the methods of smallpox outbreak control are known and will be implemented. The rapidity with which organized outbreak control measures are competently executed will determine how many generations of cases occur before the outbreak is brought under control. Planning, vaccine stockpiling, laboratory expansion, professional training and public education, all carried out well in advance of an epidemic, will minimize the number of casualties. The reinstitution of routine smallpox vaccination in Israel, as in other countries, must be given serious consideration, since it has the potential for eliminating the threat of smallpox as a bioterror agent. PMID- 12120462 TI - Hemorrhagic fevers and bioterror. PMID- 12120463 TI - Defense against biologic warfare with superantigen toxins. AB - BACKGROUND: Superantigens produced by Staphylococcus aureus and Streptococcus pyogenes are among the most lethal of toxins. Toxins in this family trigger an excessive cellular immune response leading to toxic shock. OBJECTIVES: To design an antagonist that is effective in vivo against a broad spectrum of superantigen toxins. METHODS: Short peptide antagonists were selected for their ability to inhibit superantigen-induced expression of human genes for cytokines that mediate shock. The ability of these peptides to protect mice against lethal toxin challenge was examined. RESULTS: Antagonist peptide protected mice against lethal challenge with staphylococcal enterotoxin B and toxic shock syndrome toxin-1, superantigens that share only 6% overall amino acid homology. Moreover, it rescued mice undergoing toxic shock. Antagonist peptides show homology to a beta strand/hinge/alpha-helix domain that is structurally conserved among superantigens, yet remote from known binding sites for the major histocompatibility class II molecule and T cell receptor that function in toxic T cell hyperactivation. CONCLUSIONS: The lethal effect of superantigens can be blocked with a peptide antagonist that inhibits their action at the top of the toxicity cascade before activation of T cells occurs. Superantigenic toxin antagonists may serve not only as countermeasures to biologic warfare but may be useful in the treatment of staphylococcal and streptococcal toxic shock, as well as in some cases of septic shock. PMID- 12120464 TI - Mental health consequences of bioterrorism. PMID- 12120465 TI - A novel mode of infection with hepatitis B: penetrating bone fragments due to the explosion of a suicide bomber. PMID- 12120466 TI - Radiation terrorism--the medical challenge. AB - Ionizing radiation can cause acute as well as chronic and late illnesses, and is a well-known health hazard. Its use by terrorists and nations in the form of a non-conventional weapon is no longer impossible. The release of radioactive materials with the accompanying contamination and radiation has the potential of causing serious medical problems. In analyzing the different radiologic terrorism scenarios, a scheme is proposed for the triage and evacuation of injured, contaminated and non-contaminated casualties from the scene itself as well as from the periphery. Knowledge, plans and drills will lessen the impact of those potential attacks and prepare us to respond to such events. PMID- 12120467 TI - Adverse reaction to atropine and the treatment of organophosphate intoxication. AB - Atropine is the drug of choice for treatment of organophosphate nerve agent and insecticide intoxication and has been used for this indication for several decades. Adverse reactions to atropine may occur, and are of two types: toxic and allergic. Toxic reaction, the most common form, results from the anti-muscarinic effects of the drug. Since it is most probably related to interpersonal variation in sensitivity to atropine, toxic effects may appear at the usual therapeutic doses. The second type, allergic reaction, includes local manifestations, usually after the administration of eyedrops, and systemic reaction in the form of anaphylaxis. Since most patients manifest only a mild reaction, allergy testing is not performed and the prevalence of allergy to atropine is therefore not known. Severe allergic reaction to atropine is rare, as evidenced by the small number of case reports in the literature despite the drug's extensive use. Alternative anti-muscarinic drugs recommended for OP poisoning include glycopyrrolate and scopolamine. Glycopyrrolate is a peripheral anti-muscarinic drug that has been studied in comparison to atropine for many clinical indications, while scopolamine is an anti-muscarinic drug with both peripheral and central effects. An acceptable alternative regimen for patients with proven allergy to atropine is a combination of glycopyrrolate with centrally active drugs such as benzodiazepines or scopolamine. PMID- 12120468 TI - Simulation-based training of medical teams to manage chemical warfare casualties. AB - With chemical warfare becoming an imminent threat, medical systems need to be prepared to treat the resultant mass casualties. Medical preparedness should not be limited to the triage and logistics of mass casualties and first-line treatment, but should include knowledge and training covering the whole medical spectrum. In view of the unique characteristics of chemical warfare casualties the use of simulation-assisted medical training is highly appropriate. Our objective was to explore the potential of simulator-based teaching to train medical teams in the treatment of chemical warfare casualties. The training concept integrates several types of skill-training simulators, including high tech and low tech simulators as well as standardized simulated patients in a specialized simulated setting. The combined use of multistimulation modalities makes this maverick program an excellent solution for the challenge of multidisciplinary training in the face of the looming chemical warfare threat. PMID- 12120469 TI - Training Israeli medical personnel to treat casualties of nuclear, biologic and chemical warfare. AB - Recent events have significantly increased concern about the use of biologic and chemical weapons by terrorists and other countries. Since weapons of mass destruction could result in a huge number of casualties, optimizing our diagnostic and therapeutic skills may help to minimize the morbidity and mortality. The national demands for training in medical aspects of nuclear, biologic and chemical warfare have increased dramatically. While Israeli medical preparedness for non-conventional warfare has improved substantially in recent years especially due to extensive training programs, a standardized course and course materials were not available until recently. We have developed a core curriculum and teaching materials for a 1 or 2 day modular course, including printed materials. PMID- 12120471 TI - The experience of one institution dealing with terror: the El Aqsa Intifada riots. AB - BACKGROUND: During a period of 13 months--1 October 2000 to 31 October 2001--586 terror assault casualties were treated in the trauma unit and emergency department of Hadassah University Hospital (Ein Kerem campus); 27% (n = 158) were hospitalized and the rest were discharged within 24 hours. OBJECTIVES: To analyze the special requirements of a large number of victims who received treatment during a short period. METHODS: Data were attained from the main admitting office and the trauma registry records. Factors analyzed included age, gender, mechanism of injury, anatomic site of injury, Injury Severity Score, and length of stay. RESULTS: Males comprised 81% of the hospitalized patients. The majority of the injuries (70%) were due to gunshot wounds and 31% of the hospitalized patients were severely injured (ISS > or = 16). Twelve patients died, yielding a mortality rate of 7.5%. CONCLUSION: The nature of the injuries was more complex and severe than trauma of other etiologies, as noted by the mean length of stay (10.2 vs. 7.2 days), mean intensive care unit stay (2.8 vs. 0.9 days), and mean operations per patient (0.7 vs. 0.5). The mean insurance cost for each hospitalized terror casualty was also higher than for other trauma etiologies (US$ 3,200 vs. 2,500). PMID- 12120470 TI - The medico-legal investigation of the El Aqsah Intifada. AB - BACKGROUND: The majority (n = 445) of the Israeli and Palestinian fatal victims of the El Aqsah Intifada was examined at the National Center of Forensic Medicine in Tel Aviv. Analysis of the trauma sustained and the anthropologic profile of both the victims and the perpetrators elucidates the trends and contrasts them with the phenomenon in the past. OBJECTIVES: The purpose of the forensic investigation of mass casualty incidents is manifold: establishing the minimal number of individuals involved, identifying the victims and perpetrators, collecting material evidence, and determining the modus operandi. METHODS: The postmortem examination includes external description of the bodies and their injuries, photo-documentation, and sampling of tissues. Radiography, dental examination, and a ten-print card of each cadaver are also recorded. RESULTS: The modus operandi of the current Intifada is somewhat different from that of the previous wave of terrorism and includes more road shootings and vehicular terrorism. In addition, three suicide bombers who detonated explosive devices within crowded areas were young women, and the age of the perpetrators has increased from up to 35 years to individuals as old as 47, thus greatly enlarging the potential number of suicide terrorists. Virologic and biologic tests have been introduced to examine the tissues of the suicide bombers since they are possible sources of contagion to the wounded victims. CONCLUSION: The results of the medico-legal investigation of victims and perpetrators of terrorism enable us to establish the modus operandi and the profile of potential perpetrators, which can help in the prevention of similar attacks. Documentation of the different types of injuries in fatal victims of explosion and shooting contributes to improving the awareness of the medical staff treating the wounded of similar attacks. Further investigation into the reliability of virologic and biologic tests conducted on postmortem tissue is recommended. PMID- 12120472 TI - Rectal penetrating injuries from blast trauma. PMID- 12120473 TI - Blast injury of the ear in a confined space explosion: auditory and vestibular evaluation. AB - BACKGROUND: The ear is the most frequent organ affected during an explosion. Recognition of possible damage to its auditory and vestibular components, and particularly the recovery time of the incurred damage, may help in planning the optimal treatment strategies for the otologic manifestations of blast injury and preventing deleterious consequences. OBJECTIVE: To report the results of the oto vestibular initial evaluation and follow-up of 17 survivors of a suicide terrorist attack on a municipal bus. METHODS: These 17 patients underwent periodic ear inspections and pure tone audiometry for 6 months. Balance studies, consisting of electronystagmography and computerized dynamic posturography were performed at the first time possible. RESULTS: Complaints of earache, aural fullness and tinnitus resolved, whereas dizziness persisted in most of the patients. By the end of the follow-up, 15 (55.6%) of the eardrum perforations had healed spontaneously. Hearing impairment was detected in 33 of the 34 tested ears. Recovery of hearing was complete in 6 ears and partial in another 11. ENG and CDP were performed in 13 patients: 5 had abnormal results on CDP while the ENG was normal in all the patients. Of the seven patients who complained of vertigo, only one improved and was free of symptoms 1 month after the explosion. CONCLUSION: Exposure to a high powered explosion in a confined space may result in severe auditory and vestibular damage. Awareness of these possible ear injuries may prevent many of the deleterious consequences of such injuries. PMID- 12120474 TI - Terror and rehabilitation of two family members with spinal cord injury. PMID- 12120475 TI - The role of radiology in terror injuries. AB - Although one might think that nothing could be further apart than "terror" and "medicine," in reality, medicine is intimately involved in the rescuing of those who are injured in terrorist attacks and in identifying and determining the cause of death in those who do not survive. Radiology has an important role in the workup of trauma patients in general, and in patients injured during the course of a terrorist attack in particular. Radiologic examinations determine the location and severity of injuries and are used to follow injured patients, particularly when complications occur. Conventional X-rays and CT scans are useful to detect the presence of foreign bodies, such as bullets, shrapnel and nails, which are often combined with the explosive charge in suicide bombings. Both can also be used for postmortem examinations. Although biologic, chemical and radiologic warfare constitute a real threat for the future, it is essential that we be familiar with the more "conventional" forms of terror that we face today. PMID- 12120476 TI - Primary care clinic attenders under war stress. AB - BACKGROUND: The threat to the individual's physical integrity and well-being as well as to those of significant others, the disruption of normal patterns of life, and property losses make wartime a highly stressful condition. OBJECTIVES: To assess the level of psychological distress in primary care attenders in a district of Jerusalem (Gilo) that experienced long-term exposure to gunfire. METHODS: A self-administered questionnaire exploring emotional distress (anxiety and depression symptoms), gunfire exposure, patterns of help-seeking behavior, and prescription of sedative or hypnotic drugs was administered to a sample of 125 consecutive attenders to a general practitioner during a 10 week period in the autumn of 2001. Eighty-four attenders residing in Gilo were compared with 41 attenders residing in neighborhoods that had not been under fire. T-tests and Mann-Whitney two-sample tests were used to determine statistical significance of differences. RESULTS: The mean distress score was significantly higher among the Gilo residents than among their counterparts in other neighborhoods (1.1 +/- 0.8 vs. 0.8 +/- 0.5, t = 1.73, P < 0.01); 15.5% of the former reported probable clinically significant distress. Emotional distress was associated with periods of intensive gunfire exposure, psychological care-seeking behavior, and the prescription of sedative or hypnotic drugs. No significant differences in distress levels were found between those living in zones of Gilo that were at differential gunfire risk, nor between those whose houses and cars were or were not damaged. CONCLUSIONS: War-related life events would seem to be associated with elevated emotional distress. A motivated primary care physician could easily and reliably ascertain the attenders' psychological status and identify those requiring psychological support. These identification and intervention stages are facilitated if the specialized services are community-based. PMID- 12120477 TI - Organophosphate poisoning: a multihospital survey. AB - BACKGROUND: Organophosphates are frequently used as insecticides in the household and in agricultural areas, thus posing a risk for accidental exposure. OBJECTIVES: To describe the characteristics, clinical course and outcome of 97 patients admitted to emergency rooms with a diagnosis of acute OP poisoning. METHODS: The clinical details of 97 patients were collected from 6 different hospitals in Israel. Diagnosis of intoxication was based on clinical findings, butyrylcholinesterase levels and, in several cases, the material brought to the hospital. Demographic, intoxication and clinical data were analyzed. RESULTS: The study group comprised 64 men and 33 women whose age range was 1-70 years (mean 19.8 +/- 17.1); more than one-third of the patients were less than 10 years old. Accidental exposure was the cause of intoxication in 51.5% of the patients, and suicide in 20.6% of exposures. Intoxication occurred at home in most patients (67%), and the route of intoxication was oral in 65% of them. The patients arrived at the hospital 20 minutes to 72 hours after intoxication. Nine patients were asymptomatic; 53 presented with mild intoxication, 22 with moderate, and 13 had severe intoxication, 5 of whom died. There was a direct correlation between the degree of inhibition of butyrylcholinesterase levels and the severity of intoxication. Treatment included decontamination and antidotal medication. Duration of hospitalization ranged between 1 and 14 days (average 2.9 days). CONCLUSIONS: Organophosphates may cause severe morbidity and mortality. Medical staff should therefore be aware of the clinical manifestations and the antidotal treatment for this poisoning. PMID- 12120478 TI - Ocular injuries following sulfur mustard exposure: clinical characteristics and treatment. AB - The chemical warfare agent sulfur mustard affects primarily the eyes, skin and respiratory tract. Of these, ocular injury is the most immediate and distressing. Learning to recognize ocular injury enables the treating physician to provide early and suitable treatment, which will reduce complications and allow the victim a rapid recovery. PMID- 12120479 TI - The history and threat of biological warfare and terrorism. AB - The inevitable conclusion is that the availability of biological warfare agents and supporting technologic infrastructure, coupled with the fact that there are many people motivated to do harm to the United States, means that America must be prepared to defend her homeland against biological agents. Some have argued to the contrary, that the threat and risks from a biological weapon attack are not to be considered serious, because [39]: They've not been used yet on a large scale so they probably won't be in the near future. Their use is so morally repugnant that they probably won't be used at all. The technologic hurdles associated with isolating, growing, purifying, weaponizing, and disseminating adequate quantities of pathologic agents are so high that only the most advanced laboratories could attempt the process. Similar to a 'nuclear winter,' the aftermath of a biological attack is so unthinkable that none would attempt it. Unfortunately, the trends associated with biotechnology globalization, terrorist group dynamics, and global/regional politics render these beliefs untenable and inappropriate, as recent events have underscored. To that end, the United States has accelerated its program of defense against biological weapons, as it must. Biological weapons are such dreadful weapons of uniqueness and complexity that a specific defense strategy is paramount. Elements of this program include pharmaceutical stockpiles, heightened surveillance systems, energized vaccine development programs, and comprehensive training initiatives. Although the depth and breadth of these efforts are unprecedented, above all these efforts is the absolute necessity for medical and public health care professionals to be educated and actively involved. These professionals are the sine qua non of future defensive readiness. This is just the start; unfortunately, there is no end yet in sight. PMID- 12120480 TI - Threats in bioterrorism. I: CDC category A agents. AB - Although once considered unlikely, bioterrorism is now a reality in the United States since the anthrax cases began appearing in the fall of 2001. Intelligence sources indicate there are many countries and terrorist organizations that either possess biological weapons or are attempting to procure them. In the future it is likely that we will experience additional acts of bioterrorism. The CDC category A agents represent our greatest challenge because they have the potential to cause grave harm to the medical and public health systems of a given population. Thus, it is imperative that plans be developed now to deal with the consequences of an intentional release of any one or more of these pathogens. PMID- 12120481 TI - Threats in bioterrorism. II: CDC category B and C agents. AB - A variety of agents have potential for use as weapons of biological terrorism. Knowledge of the likely organisms may be useful in preparations to mitigate the effects of a BT event. Recognition of the clinical presentation of these organisms could help physicians identify them quickly, allowing more appropriate management and possible prophylaxis of others who may have been exposed. Although many of these agents do not have specific treatments, it is important to recognize those that do. It is also important to know which infections require isolation because of potential for person-to-person spread. Table 3 summarizes important features of the CDC category B and C agents. The list of agents discussed in this article is by no means exhaustive. It is always possible that some "mad scientist" could modify an existing organism or engineer some new agent for use in biological terrorism. The possibilities are limited only by the ingenuity and depravity of those individuals who would take part in such an attack. PMID- 12120482 TI - Diagnostic analyses of biological agent-caused syndromes: laboratory and technical assistance. AB - The impact of a bioterrorism attack can be greatly reduced by collaboration among primary healthcare providers, laboratories, the veterinary community, public health officials, and emergency response personnel. Improved communication and coordination are essential to make this happen. As a first-line provider, the emergency physician must keep in mind the possibility of bioterrorism and alert the laboratory so that samples can be processed in the correct fashion. New and exciting developments in laboratory organization, communication, and diagnostic capabilities will ensure that all patients receive the best possible care. PMID- 12120483 TI - Medical management of the suspected victim of bioterrorism: an algorithmic approach to the undifferentiated patient. AB - We have purposely expanded on the well-known ATLS paradigm to aid EHCPs in their approach to a potential bioterrorism event. By building on a process that is already familiar, we hope this will aid the EHCP to remember a systematic approach to such an incident. By following this ten-step process, we believe that all EHCPs, and especially those practicing at the first echelons of care in urgent care clinics and EDs, can approach the daunting problem of biological defense with a good deal more confidence. This same model advocated for bioterrorism also may apply to natural infectious disease epidemics, particularly of emerging or re-emerging diseases, that might not be optimally managed by reliance on the conventional public health strategy that requires physician dependent definitive diagnosis and active reporting mechanisms. The authors hope the acquired knowledge and skills one might gain will rarely be needed, but if the events surrounding the dispersal of anthrax-contaminated mail in the fall of 2001 are any indication of the future, such competencies will be invaluable. PMID- 12120484 TI - Medical management of vulnerable populations and co-morbid conditions of victims of bioterrorism. AB - Planning for the medical response to bioterrorism has primarily focused around the needs of the population as a whole. There has been little discussion pertaining to certain vulnerable groups such as children, pregnant women, or immunocompromised patients, yet they will likely comprise a significant subset of the exposed population. In addition, they will be at increased risk for morbidity and mortality following an attack. The emergency response to bioterrorism will be more complex as it relates to these vulnerable populations. Careful consideration of their special needs, some of which are presented in this article, may refine our efforts. PMID- 12120485 TI - Emergency mental health management in bioterrorism events. AB - The United States has not suffered significant psychosocial or medical consequences from the use of biological weapons within its territories. This has contributed to a "natural" state of denial at the community level. This denial could amplify the sense of crisis, anxiety, fear, chaos, and disorder that would accompany such a bioterrorist event. A key part of primary prevention involves counteracting this possibility before an incident occurs. Doing so will require realistic information regarding the bioterrorism threat followed by the development of a planned response and regular practice of that response. Unlike in natural disasters or other situations resulting in mass casualties, emergency department physicians or nurses and primary care physicians (working in concert with epidemiologic agencies), rather than police, firemen, or ambulance personnel, will be most likely to first identify the unfolding disaster associated with a biological attack. Like community leaders, this group of medical responders must be aware of its own susceptibility to mental health sequelae and performance decrement as the increasing demands of disaster response outpace the availability of necessary resources. A bioterrorist attack will necessitate treatment of casualties who experience neuropsychiatric symptoms and syndromes. Although symptoms may result from exposure to infection with specific biological agents, similar symptoms may result from the mere perception of exposure or arousal precipitated by fear of infection, disease, suffering, and death. Conservative use of psychotropic medications may reduce symptoms in exposed and uninfected individuals, as may cognitive-behavioral interventions. Clear, consistent, accessible, reliable, and redundant information (received from trusted sources) will diminish public uncertainty about the cause of symptoms that might otherwise prompt persons to seek unnecessary treatment. Training and preparation for contingencies experienced in an attack have the potential to enhance delivery of care. Initiating supportive social, psychotherapeutic, and psychopharmacologic treatments judiciously for symptoms and syndromes known to accompany the traumatic stress response can aid the efficient treatment of some patients and reduce long-term morbidity in affected individuals. Preventive strategies and planning must take into account the idea that specific groups within the population are at higher risk for psychiatric morbidity. First responders comprise one group at psychologic risk in this situation, and healthcare providers comprise another. These and other high-risk groups will benefit from the same supportive interventions developed for the community as a whole. PMID- 12120486 TI - Bioterrorism preparedness. I: The emergency department and hospital. AB - Fundamental precepts in hospital-based planning for bioterrorist events include having a comprehensive well-rehearsed disaster plan that is based on a threat and vulnerability analysis. JCAHO Environment of Care Standards and an "all-hazards" approach to disaster planning and management form the basis for a solid bioterrorism response plan. During preparation, education and training are imperative. Clinicians must maintain a high index of suspicion for use of bioterrorism agents, be able to make a rapid diagnosis, and promptly initiate empiric treatment. Other personnel from administration, security, public relations, laboratory, pharmacy, and facilities management should be familiar with the plan, know when and how to activate it, and understand their roles in the response. A recognized incident command system should be used. Hospital leadership must be aware of the facility's capabilities and capacities, and should have plans for expansion of services to meet the surge in demand. The command center should coordinate emergency personnel teams, decontamination, security, acquisition of supplies, and notification of public health and other authorities and the media. If the plan is ever implemented, stress management with psychologic support will play an important role in recovery. PMID- 12120487 TI - Bioterrorism preparedness. II: The community and emergency medical services systems. AB - Disaster planning is an arduous task. Perhaps no form of disaster is more difficult to prepare for than one resulting from the intentional, covert release of a biological pathogen or toxin. The complexities of response operations and the perils of inadequate preparation cannot be overemphasized. Even with detailed planning, deviations from anticipated emergency operations plans are likely to occur. Several federal programs have been initiated to assist communities in enhancing their preparedness for events involving biological and other agents of mass destruction. Many of these, such as the Metropolitan Medical Response Systems (MMRS) Program [37,38], will be discussed elsewhere. Community preparedness will be enhanced by: 1. Implementing a real-time public health disease surveillance program linking local healthcare, emergency care, EMS, the CDC, local law enforcement, and the FBI 2. Improved real-time regional patient and healthcare capacity status management 3. Development of affordable, accurate biological agent detection systems 4. Incorporation of standardized education and training curricula (appropriate for audience) on terrorism and biological agents into healthcare training programs 5. Expansion of federal and state programs to assist communities in system development 6. Increased public awareness and education programs. PMID- 12120488 TI - Bioterrorism preparedness. III: State and federal programs and response. AB - Management of a bioterrorism event will begin with early detection and intervention at the local level. Any large-scale event will require rapid state and federal assistance. Federal initiatives targeting bioterrorism have increasingly become a complex web of executive and legislative actions, frequently initiated in reaction to specific events, and often unrelated to this threat. Multiple executive and legislative branch actions have resulted in a proliferation of federal programs, and coordination of these efforts remains a significant challenge. Still, great strides have been taken to improve our defensive posture against this emerging threat, and, at all levels, governmental authorities and agencies are much better prepared to respond to such events than they were a decade ago. The events of September 11, 2001 and subsequent events are clear indicators that the timeline for preparedness has been significantly compressed. Federal emergency operations, historically designed more for recovery than response, seemed up to the task in the wake of the World Trade Center and Pentagon attacks, although there was criticism of federal responsiveness to the subsequent anthrax incidents [71,72], and the timeliness of federal resources in the event of a large-scale outbreak resulting from a bioterrorism attack has yet to be truly tested. The recent establishment of the Office of Homeland Security and the Homeland Security Council holds promise that some of these inefficiencies may be rectified and overall coordination of programs will improve. Continued improvements in the effectiveness of the federal government in meeting the challenges of this and other emerging threats to homeland security will require: Establishment of consensus standards, metrics, and measures of effectiveness for all aspects of disaster, epidemic, and terrorism management at the local, regional, state, and federal levels Delineation of expected, quantifiable state and local capabilities to mitigate, prepare, respond, and recover from all disasters, including those caused by terrorist actions Development of predefined or clear and rapidly discernible criteria for deployment of state and federal emergency resources Full accountability of program costs and expenditures Continued consolidation or coordination of the many overlapping and at times redundant federal programs. PMID- 12120489 TI - Future challenges in preparing for and responding to bioterrorism events. AB - The future success of our preparations for bioterrorism depends on many issues as presented in this article. If these issues are properly addressed, the resulting improvements in bioterrorism preparations will allow us to better deter and mitigate a bioterrorism incident and will also provide us with the added benefit of improvements in early detection, diagnosis, and treatment of natural disease outbreaks. Emergency physicians must take an active leading role in working with the various disciplines to produce a better-prepared community. PMID- 12120490 TI - Culture-centrism and holistic care in nursing practice. PMID- 12120491 TI - Current concepts in the management of hypertensive crisis: emergencies and urgencies. AB - Hypertensive emergencies and hypertensive urgencies represent a large percentage of major medical emergencies and have the potential of producing serious organ damage or death if not treated promptly and selectively. Several classifications of antihypertensive agents are discussed, with emphasis on selecting agents appropriate for patients' hypertension manifestations and comorbid situations. Epidemiology and evaluation of hypertension, as well as common pharmacokinetics of several common and new oral and parenteral antihypertensive agents, are described. Special nursing considerations of medication administration and gerontology concepts are included. PMID- 12120492 TI - Measuring quality of care for essential hypertension. AB - Published guidelines set the standard of care for essential hypertension. The authors offer an audit tool based on national standards to evaluate how actual treatment measures up to the published standard of care. The measure of quality of care is based on three assumptions: Recording essential observations is an integral part of optimal health care What is recorded took place and what is not recorded did not take place If documentation of essential events agrees with audit criteria, the outcome of care will most likely be optimal PMID- 12120494 TI - Selected complementary methods and nursing care of the hypertensive client. AB - The National Institutes of Health and the World Health Organization have targeted the treatment of mild or borderline hypertension as a critical health care issue. Conventional practitioners' focus on more intensive treatment for blood pressure elevations in the lower ranges is accompanying the consumer-driven movement toward the use of complementary methods. Over 60 million Americans used herbal therapies in the past year, and visits to complementary care practitioners are expected to increase beyond the 425 million now made annually. The purpose of this article is to identify four herbs that consumers commonly select for the treatment of hypertension and identify nursing care considerations for their use. Additionally, the article reviews research on the effectiveness of acupuncture for hypertension, along with nursing care implications for patients. PMID- 12120493 TI - The health risk of hypertension in south Texas: a demographic profile. AB - This article provides a demographic profile of the hypertensive adult in South Texas. Hypertension, a silent disease because of its lack of overt signs and symptoms, poses a major public health problem. Few prevalence studies have been completed reflecting Mexican-American populations. South Texas hypertension trends are comparable to national trends. However, the awareness of hypertension in South Texas is lower than that of the national population. The density of risk factors may be responsible for the patterns of South Texas hypertension. PMID- 12120495 TI - Nurturing hope and spirituality in the nursing home. AB - Nurturing body, mind, and spirit is part of holistic care, and yet often the primary focus of care in nursing homes is physical needs. As part of a larger study examining factors related to hope among institutionalized elders, spirituality emerged as the only significant predictor of hope. Findings supported the active presence of hope despite age and functional limitations. The significant contribution of spirituality to hope calls for attention to the provision of opportunities to support and enhance spiritual practices in the nursing home setting. Nurses in nursing homes have the opportunity to establish close relationships with residents over time, often substituting for family and friends no longer available. Because length of stay is long, more time is available to enter into meaningful spirit-sharing relationships with residents. Suggestions for interventions that nurture hope and spirituality within a holistic and caring framework are presented. PMID- 12120496 TI - The development of critical thinking among students in baccalaureate nursing education. AB - Students entering nursing education present with a range of critical-thinking skills and dispositions that are affected by genetic influences, modeling by others, and formal learning experiences. The experience of higher education further contributes, positively or negatively, to the development of those skills and dispositions over time. A conceptual framework is presented as a model to explain the relationship between variables contributing to critical thinking as it is manifested in clinical judgment. A developmental model is presented to align the development of cognitive capabilities and the application of those capabilities in the subprocesses of the nursing process. Implications for nurse educators related to the understanding of critical-thinking development of students are discussed. PMID- 12120498 TI - Hypertension and holistic care. PMID- 12120497 TI - Validity evidence for using a general critical thinking test to measure nursing students' critical thinking. AB - This study examined validity evidence for using a general test of critical thinking skills and dispositions to measure nursing students' critical-thinking abilities. Content evidence indicated strong support for the theoretical framework underlying the test but less support for the way in which the critical thinking constructs were specifically measured. Scores related to critical thinking skills demonstrated significant but low correlations with grade point averages, were moderately correlated with SAT scores, and were uncorrelated with scores related to critical-thinking dispositions. The evidence suggests that nursing programs may need to reconsider how critical thinking should be measured and evaluated. PMID- 12120499 TI - Issues and trends in care of the hypertensive client. AB - This article discusses issues and trends in care of the hypertensive client that pertain to practice, education, research, and health policy. Clinical practice issues center around obtaining accurate blood pressure measurements and educating patients about health promotion interventions and use of medications. These topics are important in the education of the beginning nurse student as well. Researchers have studied risk factors, incidence, and prevalence in global populations; use of pharmacologic agents; and investigative methods employing technology. Researchers are beginning to explore more holistic approaches to controlling risk factors before hypertension occurs. Funding for public education regarding prevention and control of hypertension is a number one health policy issue. PMID- 12120500 TI - Fuelling the pipeline. AB - Drug pipelines need to be fed, not just with promising compounds, but also with promising scientists. Despite the high-tech nature of the enterprise, the skills most in demand when companies are recruiting can often be the most traditional. PMID- 12120501 TI - Structural proteomics. PMID- 12120502 TI - Building drug delivery into tissue engineering. AB - The creation of efficient methods for manufacturing biotechnology drugs--many of which influence fundamental but complex cell behaviours, such as proliferation, migration and differentiation--is creating new opportunities for tissue repair. Many agents are potent and multifunctional; that is, they produce different effects within different tissues. Therefore, control of tissue concentration and spatial localization of delivery is essential for safety and effectiveness. Synthetic systems that can control agent delivery are particularly promising as materials for enhancing tissue regeneration. This review discusses the state of the art in controlled-release and microfluidic drug delivery technologies, and outlines their potential applications for tissue engineering. PMID- 12120503 TI - Regulators of G-protein signalling as new central nervous system drug targets. AB - G-protein-coupled receptors (GPCRs) are major targets for drug discovery. The regulator of G-protein signalling (RGS)-protein family has important roles in GPCR signal transduction. RGS proteins contain a conserved RGS-box, which is often accompanied by other signalling regulatory elements. RGS proteins accelerate the deactivation of G proteins to reduce GPCR signalling; however, some also have an effector function and transmit signals. Combining GPCR agonists with RGS inhibitors should potentiate responses, and could markedly increase the agonist's regional specificity. The diversity of RGS proteins with highly localized and dynamically regulated distributions in brain makes them attractive targets for pharmacotherapy of central nervous system disorders. PMID- 12120504 TI - Allosteric binding sites on cell-surface receptors: novel targets for drug discovery. AB - Cell-surface receptors are the targets for more than 60% of current drugs. Traditionally, optimizing the interaction of lead molecules with the binding site for the endogenous agonist (orthosteric site) has been viewed as the best means of achieving selectivity of action. However, recent developments have highlighted the fact that drugs can interact with binding sites on the receptor molecule that are distinct from the orthosteric site, known as allosteric sites. Allosteric modulators could offer several advantages over orthosteric ligands, including greater selectivity and saturability of their effect. PMID- 12120505 TI - NMR in drug discovery. AB - NMR spectroscopy has evolved into an important technique in support of structure based drug design. Here, we survey the principles that enable NMR to provide information on the nature of molecular interactions and, on this basis, we discuss current NMR-based strategies that can identify weak-binding compounds and aid their development into potent, drug-like inhibitors for use as lead compounds in drug discovery. PMID- 12120506 TI - Chemical database techniques in drug discovery. AB - Chemical databases are becoming a powerful tool in drug discovery. Database searches based on possible requirements for biological activity can identify compounds that might be suitable for further analysis or indicate novel ways to achieve the desired activity. What considerations are involved in the construction and searching of chemical databases? PMID- 12120507 TI - Development of therapeutics: opportunities within complementary and alternative medicine. AB - Whereas other components of the National Institutes of Health support the discovery and subsequent development of novel chemical entities into drugs, the National Center for Complementary and Alternative Medicine (NCCAM) studies complex natural products that are marketed as dietary supplements. This article contrasts the regulatory framework for dietary supplements and drugs, outlines the challenges of evaluating dietary supplements for safety and clinical effectiveness, and describes the evolving drug model for botanicals. PMID- 12120508 TI - The role of pharmacology in drug discovery. AB - The International Union of Pharmacology (IUPHAR) enthusiastically welcomes the decision by the Nature Publishing Group to launch its new journal, Nature Reviews Drug Discovery. The title of the new journal poses interesting questions for pharmacologists. Why 'Drug Discovery'? Would we have preferred 'Pharmacology'? And do these distinctions even matter, as aren't all pharmacologists involved somehow in drug discovery? PMID- 12120510 TI - Can Master Craftsmen be mass-produced? AB - Pharmaceutical companies have not realized the promised productivity gains from novel research technologies. Discovery 'industrialization' also demands profound changes to both people skills and organization for the benefits to be delivered. PMID- 12120509 TI - Indirect monitoring of lung inflammation. AB - The assessment of airway inflammation by non-invasive methods could provide a signal to start anti-inflammatory treatment before the onset of symptoms and the impairment of lung function. It could also be useful in the follow-up of patients with lung disease, and for guiding drug treatment. Measuring inflammatory markers in exhaled breath condensate is potentially the easiest way to quantify lung inflammation. The clinical applicability of this method could facilitate the practice of respiratory medicine. PMID- 12120511 TI - The scanning probe microscopy of metalloproteins and metalloenzymes. AB - In recent years, the concept of microscopy and the ability to study processes at a truly molecular level have been revolutionised by the development of a family of instruments based on acquiring data through the scanning of a proximal probe across a surface. Scanning Probe Microscopes (SPMs) enable surface-confined structures to be resolved at angstrom-resolution, in real time, and under a variety of controllable conditions. Despite initial difficulties, much progress has been in the application of this technology to the high-resolution analysis of biological systems; these have varied from complex cellular systems to molecular biopolymers. Studying the interactions of protein with surfaces has been intrinsic to the development of our understanding of blood coagulation, fibrinolysis, thrombus formation and the synthesis of biocompatible materials. The specific interactions of metalloproteins and enzymes with electrode surfaces remains central to the understanding of the bioelectrochemical processes and to the development of biosensing devices. Though ellipsometry, Raman, microcalorimetry, surface plasmon resonance, and other spectroscopic methods, can provide much information on these interfaces, the acquired data are averaged over a large number of molecular species with a low spatial resolution. Proximal probe methods have much to offer in this regard and have revolutionized our ability to monitor such interactions. PMID- 12120512 TI - Metal-induced regulation of fullerene complexation with double-calix[5]arene. AB - The metal-induced regulation of fullerene complexation with double-calix[5]arene is described. The receptor shows strong binding to C70 only in the presence of Cu+. PMID- 12120513 TI - The role of template in the synthesis of meso-hexamethyl-meso-hexaphenyl calix[6]pyrrole: trihalogenated compounds as templates for the assembly of a host with a trigonal cavity. AB - Trihalogenated compounds act as effective and selective templates in the template assisted synthesis of meso-hexaphenyl-calix[6]pyrrole. PMID- 12120514 TI - Mesoporous aluminophosphates from a single-source precursor. AB - Mesoporous aluminophosphates with a strict ratio of A1:P = 1:1 have been synthesized from a single-source molecular precursor. PMID- 12120515 TI - Formation of a molecular spin ladder induced by a supramolecular cation structure. AB - A novel molecular spin ladder structure of a nickel dithiolate complex has been constructed using a supramolecular cation composed of anilinium and 18-crown-6. PMID- 12120516 TI - Selective oxidation of cyclooctane to cyclootanone with molecular oxygen in the presence of compressed carbon dioxide. AB - The oxidation of cyclooctane (1) to cyclooctanone (3) with molecular oxygen and acetaldehyde (2) as a co-reductant occurs efficiently in the presence of compressed CO2. Up to 20% yields of 3 are obtained under optimised multiphase conditions. PMID- 12120518 TI - Alkylation of heme by the antimalarial drug artemisinin. AB - The peroxide function of artemisinin has been activated by iron(II)-heme generated in situ from iron(III)-protoporphyrin-IX and glutathione, a biologically relevant reductant. In mild conditions, this reaction produced a high yield (85%) of heme derivatives alkylated at alpha-, beta-, and delta-meso positions by a C4-centered radical derived from artemisinin. PMID- 12120517 TI - Oxygenative cleavage of catechols including protocatechuic acid with molecular oxygen in water catalysed by water-soluble non-heme iron(III) complexes in relevance to catechol dioxygenases. AB - Catechol dioxygenase model oxygenations have been performed for the first time in water by using water-soluble nonheme iron(III) complexes, enabling the oxygenation of protocatechuic acid and other catechols. PMID- 12120519 TI - A model recognition switch. Electrochemical control and transduction of imidazole binding by electrode-immobilized microperoxidase-11. AB - Electrode-immobilized microperoxidase-11 exhibited a titratable potentiometric response to imidazole, demonstrating both molecular recognition and the capability for "switchable" changes in the affinity of an immobilized redox receptor for a target ligand. PMID- 12120520 TI - A tetranuclear nickel(II) complex assembled from an asymmetric compartmental ligand and bearing an intramolecular [H3O2]-bridge. AB - The asymmetric di-aminic compartmental ligand HL5 forms a tetranuclear nickel(II) complex in which the core is assembled from two confacial bioctahedra [Ni...Ni, approximately 2.90 A]; the open faces of the bioctahedra are joined at the O atoms of two mu-cresolato bridges [Ni...Ni, 3.72 A], and the shared faces of the bioctahedra are linked by a tetradentate (mu 4, eta 2)-perchlorate anion and by an unusual tetradentate (mu 4, eta 2)-[H3O2]-bridge. PMID- 12120521 TI - Self-assembly of double stranded dinuclear titanium(IV)-Schiff base complexes and formation of intramolecular mu-oxo bridges. AB - The reaction of titanium isopropoxide with a Schiff base ligand containing an isobutenyl linker leads to double stranded dinuclear titanium(IV)-Schiff base complexes through self-assembly with concomitant formation of intramolecular mu oxo bridges upon hydrolysis. PMID- 12120522 TI - Selective hydrogenation of maleic anhydride to gamma-butyrolactone over Pd/Al2O3 catalyst using supercritical CO2 as solvent. AB - A selective hydrogenation of maleic anhydride to either gamma-butyrolactone or succinic anhydride over simple Pd/Al2O3 catalyst under supercritical CO2 medium is described for the first time which has considerable promise for both lab-scale as well as industrial selective hydrogenations of low vapor pressure compounds without employing environmentally harmful organic solvents. PMID- 12120523 TI - A novel scoop-shaped conformation of C-methylcalix[4]resorcinarene in a bilayer structure. AB - A new scoop-shaped conformation of C-methylcalix[4]resorcinarene has been identified; it is a hybrid of the previously observed crown- and flattened cone conformations. PMID- 12120525 TI - The positive effect on hole transport behaviour in anisotropic gels consisting of discotic liquid crystals and hydrogen-bonded fibres. AB - The hole mobility of a discotic liquid-crystalline triphenylene compound is enhanced by the gelation with hydrogen-bonded fibrous aggregates. PMID- 12120524 TI - Synthesis of D-erythro-sphingosine and D-erythro-ceramide. AB - A 1,2-metallate rearrangment of a higher order cuprate derived from an alpha lithiated xylal derivative and tridecyllithium is the key step in a synthesis of D-erythro-sphingosine and ceramide. A convenient method for preparing alpha lithiated glycals from alpha-phenylsulfinyl glycals is also described. PMID- 12120526 TI - Erythrocyte-like liposomes prepared by means of amphiphilic cyclodextrin sulfates. AB - A novel class of sulfated glycolipids with excellent self-assembling capacity to form stable monolayers at an air-water interface and specific erythrocyte-like liposomes was synthesised from alpha, beta, and gamma-cyclodextrins as starting materials. PMID- 12120528 TI - Metal catalysed Michael additions in ionic liquids. AB - The ionic liquid [bmim][BF4] (1-n-butyl-3-methylimidazolium tetrafluoroborate) was used as innovative solvent for a recyclable catalytic system active in metal promoted Michael additions employing Ni(acac)2 as catalyst. PMID- 12120527 TI - A new supramolecular assembly obtained from the combination of silver(I) cations with a thiophosphorylated cavitand. AB - The new tetra-thiophosphonatocavitand 1 in its iiii configuration extracts quantitatively Ag+ ions from aqueous solutions; the tetranuclear complex [1(2).Ag4Pic4] was selectively formed and characterized in the solid state by X ray diffraction which revealed the formation of a new dimeric assembly through Ag+ coordination. PMID- 12120529 TI - Mixed-anion complexes with a bipyrazolyl ligand. A new entry to a realm of three dimensional five-connected coordination topologies. AB - Cross-linking of corrugated square grid coordination layers by anionic bridging groups generates five-connected coordination networks; the 3D topologies of mixed anion cobalt(II) and nickel(II) complexes with a tetramethyl-substituted 4,4' bipyrazolyl ligand are supported by mu-SO4(2-) functions and exist as neutral or cationic five-connected arrays involving additional terminal (NCS-) or non coordinated (NO3-, ClO4-) groups. PMID- 12120530 TI - Intramolecular Diels-Alder reactions of N-substituted oxazolones. AB - The first intramolecular Diels-Alder reactions of simple trienes featuring an N substituted oxazolone as the dienophilic component have been investigated and are reported herein. PMID- 12120531 TI - Soccer-playing metal oxide giant spheres: a first step towards patterning structurally well defined nano-object collectives. AB - In context with the challenge to assemble giant molecules into patterns with limited size, molybdenum oxide giant spheres (with a molecular mass of about 16 kDa) could be 'kicked out' like soccer balls into the gas phase using matrix assisted laser desorption and ionization (MALDI) and detected by TOF mass spectrometry while cluster collectives ranging from dimers to pentamers were observed. PMID- 12120532 TI - Perfluorocarbon-stabilized silver nanoparticles manufactured from layered silver carboxylates. AB - Perfluorocarboxylate-stabilized silver nanoparticles have been prepared uniformly via the thermal decomposition of layered silver perfluorocarboxylates (AgCO2(CF2)nCF3, n = 10, 12, 14 and 16). PMID- 12120533 TI - Three component coupling reactions of N-acetyl-2-azetine- rapid stereoselective entry to 2,3,4-trisubstituted tetrahydroquinolines. AB - N-Acetyl-2-azetine undergoes Lewis acid catalysed [4 + 2]-cycloaddition with imines derived from aromatic amines and gave a 1:1 mixture of exo-endo diastereoisomeric azetidine cycloadducts which reacted further with aromatic amine, to give 2,3,4-trisubsitituted tetrahydroquinolines in good to excellent yield, predominantly as one diastereoisomer. PMID- 12120534 TI - Photophysics, aggregation and amplified quenching of a water-soluble poly(phenylene ethynylene). AB - The fluorescence, absorption and fluorescence quenching properties of an anionic poly(phenylene ethynylene) are investigated in H2O and MeOH solutions. PMID- 12120535 TI - Carbon ceramic electrode modified with redox liquid. AB - A carbon ceramic electrode modified with a redox liquid, butylferrocene, exhibiting in aqueous salt solution electrochemical behaviour resulting from the redox process of the modifier and ion transfer across the liquid-liquid interface has been prepared. PMID- 12120536 TI - A novel and efficient oxidative biaryl coupling reaction of phenol ether derivatives using a combination of hypervalent iodine(III) reagent and heteropoly acid. AB - A novel and efficient oxidative biaryl coupling reaction of phenol ether derivatives using a combination of hypervalent iodine(III) reagent, phenyliodine(III) bis(trifluoroacetate) (PIFA), and heteropoly acid has been developed. PMID- 12120538 TI - Synthesis of the first 1,2,3,4-azatriphospholene complex. AB - Synthesis of the first 1,2,3,4-azatriphospholene complex was achieved by heating a solution of a P-phenyl-substituted 7-phosphanorbornadiene tungsten complex and triphenylphosphonio cyanomethylide, whereby CH-insertion products were formed in a competing reaction; these results also provide first evidence for the ability of electrophilic terminal phosphanediyl complexes to react at the ylide carbon atom and at the carbonitrile nitrogen atom of Wittig-ylides having a nitrile functional group; the structures of both complexes were established through X-ray single-crystal diffraction studies. PMID- 12120537 TI - Low temperature selective catalytic reduction (SCR) of NO with NH3 over Fe-Mn based catalysts. AB - Fe-Mn based transition metal oxides (Fe-Mn, Fe-Mn-Zr and Fe-Mn-Ti) show nearly 100% NO conversion at 100-180 degrees C for selective catalytic reduction of NO with NH3 under the applied conditions with a space velocity of 15,000 h-1. PMID- 12120539 TI - Sodium and potassium 3-sila-beta-diketiminates show new coordination modes. AB - The reaction between the lithium 3-sila-beta-diketiminate (1) and the appropriate MOBut yielded the crystalline sodium (2) or potassium (3) 3-sila-beta diketiminate in high yield; X-ray crystal data and NMR spectra show new coordination modes for the [N(R)C(Ar)Si(R)C(Ar)N(R)]- ligand (R = SiMe3, Ar = C6H3Me(2)-2,6). PMID- 12120540 TI - Reactive species formed from N-benzyloxycarbonyl alpha-aminophosphonochloridates and triethylamine: probable identity and implications for synthesis. AB - The sterically congested phosphonochloridate BnOCONHCMe2P(O)(OMe)Cl reacts rapidly with Et3N to give what is thought to be an oxazaphospholine oxide 7 (and Et3NHCl); unhindered BnOCONHCH2P(O)(OMe)Cl seems to react in the same way, in which case the product is not a phosphonylammonium salt as has been suggested. PMID- 12120541 TI - Mimicking dye-based functions of natural blue-light photoreceptors by studying photoinduced energy and electron transfer in a pyrene-isoalloxazine(flavin) phenothiazine triad. AB - An artificial system containing phenothiazine as electron donor, isoalloxazine as flavinoid redox-mediator and pyrene as antenna has been built up in order to model photoinduced energy and electron transfer processes of natural blue-light photoreceptors. PMID- 12120543 TI - The first supraicosahedral p-block metallacarboranes. AB - Reaction of Na2[7,9-C2B10H12] or its C-dimethyl analogue with SnCl2 in THF affords 4,1,6-closo-SnC2B10H12 or 1,6-Me2-4,1,6-closo-SnC2B10H10 respectively, the first 13-vertex metallacarboranes involving p-block metals; spectroscopic, structural and theoretical studies, including an analysis of fluctionality in solution, are reported. PMID- 12120542 TI - Core modified oxybenziporphyrins: new aromatic ligands for metal-carbon bond activation. AB - Successful syntheses of two new aromatic core modified oxybenziporphyrins by a simple '3 + 1' methodology and the first aromatic core modified oxybenziporphyrin palladium complex are reported. PMID- 12120544 TI - Photoinduced alkyl group exchange of ethylzinc alkoxides: X-ray crystal structure of an iodomethylzinc methoxide. AB - Irradiation of a solution of ethyl zinc alkoxides and CH2I2 leads to clean formation of iodomethylzinc alkoxides; these intermediates are important species generated in stereoselective cyclopropanation processes; no alkyl group exchange is observed in the absence of irradiation; the solid-state structure of (MeO)8Zn7(CH2I)6 is also reported. PMID- 12120545 TI - A new class of macrocyclic complexes formed via nickel-promoted macrocyclisation of dioxime with dinitrile. AB - o-Phthalonitrile couples with chelating dioxime on nickel(II), with formation of a dinuclear nickel(II) macrocyclic complex--the first representative of a new class of imine-appended macrocycles. PMID- 12120546 TI - Direct monitoring of DNA cleavages catalyzed by an ATP-dependent deoxyribonuclease on a 27 MHz quartz-crystal microbalance. AB - Kinetic studies of enzymatic DNA cleavage reactions (the enzyme binding, hydrolysis along DNA strands, and then release of the enzyme from the completely hydrolyzed ssDNA) were carried out on a 27 MHz quartz-crystal microbalance. PMID- 12120547 TI - [Zn8(SiO4)(C8H4O4)6]n: the firstborn of a metallosilicate-organic hybrid material family (C8H4O4 = isophthalate). AB - [Zn8(SiO4)(C8H4O4)6]n (C8H4O4 = isophthalate), synthesized by hydrothermal reaction, possesses a diamondoid framework structure constructed from hexahedron like Zn8(SiO4) cores and C8H4O4 linkers and remains stable up to 500 degrees C in air, representing the first member of a new class of metallosilicate-organic hybrid materials. PMID- 12120548 TI - The first gas chromatographic resolution of carnitine enantiomers. AB - The enantiomers of carnitine are converted on-line in the injection port of a gas chromatograph into beta-hydroxy-gamma-butyrolactones and are separated on a derivatized beta-cyclodextrin chiral stationary phase. PMID- 12120549 TI - Light-assisted chemical deposition of highly (0001) oriented zinc oxide film. AB - Highly (0001) oriented zinc oxide (ZnO) films of smooth layer type and hexagonal columns have been prepared on quartz glass substrates at temperature as low as 323 K by UV light assisted chemical deposition from an aqueous solution containing hydrated zinc nitrate and dimethylamine-borane (DMAB). PMID- 12120550 TI - A chromophore-labeled poly(N-propargylamide): a new strategy for a stimuli responsive conjugated polymer. AB - A new stimuli-responsive conjugated polymer was synthesized, where, following the change of secondary conformation from helical to disordered state, the fluorescence property of the side chain chromophore changes upon sensing external stimuli. PMID- 12120551 TI - Novel monodentate spiro phosphorus ligands for rhodium-catalyzed hydrogenation reactions. AB - Novel monodentate phosphorus ligands containing the 1,1'-spirobiindane backbone have been synthesized and applied in the asymmetric rhodium-catalyzed hydrogenation of functionalized olefins, providing excellent enantioselectivities (up to 99.3% ee). PMID- 12120552 TI - A novel and direct synthesis of indoles via catalytic reductive annulation of nitroaromatics with alkynes. AB - Indoles are produced regioselectively and in moderate yields from the reactions of nitroaromatics with alkynes catalyzed by [CpM(CO)2]2 (1; [(eta 5 C5H5)Fe(CO)2]2; [(eta 5-C5Me5)-Fe(CO)2]2; [(eta 5-C5Me5)Ru(CO)2]2) under carbon monoxide. PMID- 12120553 TI - Electrochemical generation of ferrate in acidic media at boron-doped diamond electrodes. AB - An extremely strong oxidant, ferrate (Fe(VI) or FeO4(2-), has been produced electrochemically in an acidic aqueous medium for the first time. PMID- 12120554 TI - Selectfluor F-TEDA-BF4 mediated and solvent directed iodination of aryl alkyl ketones using elemental iodine. AB - Reactions of aryl alkyl ketones with methanol solution of elemental iodine and 1 fluoro-4-chloromethyl-1,4-diazoniabicyclo[2.2.2]octane bis(tetrafluoroborate) (Selectfluor F-TEDA-BF4) result in the formation of corresponding alpha-iodo ketones, while switch over of the regioselectivity can be directed by using acetonitrile as the solvent and selective iodination of the aromatic site of target molecules is thus achieved. PMID- 12120555 TI - Butane and propane oxidation by engineered cytochrome P450cam. AB - The haem monooxygenase cytochrome P450cam has been engineered to oxidise the gaseous alkanes butane and propane to butan-2-ol and propan-2-ol, respectively, by the use of bulky amino acid substitutions to reduce the volume of the substrate pocket and thus improve the enzyme-substrate fit: the F87W/Y96F/T101L/V247L mutant oxidizes butane with a turnover rate of 750 min-1 and 95% yield based on NADH consumed while the wild-type enzyme has an activity of 0.4 min-1 with 4% yield. PMID- 12120556 TI - Vesicle formation induced by metal ions from micelle-forming sodium hexadecylimino diacetate in dilute aqueous solution. AB - In dilute aqueous solution, micelle-forming sodium hexadecylimino diacetate assembles into vesicles induced by Cu(II), Co(II) and Ni(II) ions. PMID- 12120558 TI - The synthesis of [2], [3] and [4]rotaxanes and semirotaxanes. AB - The cucurbituril-catalysed synthesis of [2], [3] and [4]semirotaxanes allows access to regioselectively pure, 1,3-disubstituted mono-, bis- and tris-triazoles in high yield after dethreading. PMID- 12120559 TI - Asymmetric Michael addition of formaldehyde N,N-dialkylhydrazones to alkylidene malonates. AB - Enantiopure formaldehyde N,N-dialkylhydrazones 1 smoothly react with prochiral alkylidene malonates 2 in the presence of MgI2 to afford the corresponding Michael adducts 3 in excellent yields and good diastereoselectivities; direct racemization-free BF3.OEt2-catalyzed thiolysis of the hydrazone C=N bond affords the corresponding dithioketals 7 in optically pure or enantiomerically enriched form. PMID- 12120557 TI - Cyclohexadiene-trans-diols as versatile starting material in natural product synthesis: short and efficient synthesis of iso-crotepoxide and ent-senepoxide. AB - A new synthesis of ent-senepoxide and iso-crotepoxide starting from microbially produced(+)-trans-2,3-dihydroxy-2,3-dihydrobenzoic acid via regio- and stereoselective epoxidation is described. PMID- 12120561 TI - Synthesis and molecular structure of titanium complexes containing a reduced TEMPO radical. AB - Two titanium compounds containing monoanionic ligands derived from TEMPO were synthesized and structurally characterized, demonstrating the flexibility of the coordination modes adopted by the ligand. PMID- 12120560 TI - Local disruption of DNA-base stacking by bulky base surrogates. AB - A novel biphenyl base surrogate disrupts 2-aminopurine base stacking while maintaining duplex integrity. PMID- 12120562 TI - Design of bimodal mesoporous silicas with interconnected pore systems by ammonia post-hydrothermal treatment in the mild-temperature range. AB - Bimodal (4 and 8 nm) mesoporous silicas with interconnected three-dimensional structure were synthesized by mild-temperature post-synthesis hydrothermal treatment of MCM-41 mesoporous materials in ammonia solution. PMID- 12120563 TI - Synthesis and structure of a heptanuclear nickel(II) complex uniquely exhibiting four distinct binding modes, two of which are novel, for a hydroxamate ligand. AB - The reaction of 2-(dimethylamino)phenylhydroxamic acid (2-dmAphaH) with NiSO(4).6H2O gives the complex [Ni7(2-dmAphaH-1)2(2-dmApha)8(H2O)2]SO(4).15H2O uniquely exhibiting four distinct hydroxamate binding modes, two of which are novel, and showing both antiferromagnetic and ferromagnetic interactions in contrast to [Cu5(2-dmAphaH-1)4(HSO4)2(MeOH)2].2MeOH, a strongly antiferromagnetic metallacrown formed with CuSO(4).5H2O. PMID- 12120564 TI - Solid-phase synthesis of apicidin A and a cyclic tetrapeptoid analogue. AB - The solid-phase synthesis of the antiprotozoal cyclic tetrapeptide apicidin A is reported and its synthetic accessibility is contrasted with that of a structurally similar reduced cyclic tetrapeptoid analogue. PMID- 12120565 TI - Ruthenium bis(bipyridine) sulfoxide complexes: new catalysts for alkene epoxidation. AB - The ruthenium bis(bipyridine) sulfoxide complexes Ru-1 and Ru-2 exhibit high catalytic activity for epoxidation of unfunctionalized olefins in the presence of [bis(acetoxy)iodo]benzene; with the chiral catalyst, Ru-2, asymmetric induction up to 94% was observed for beta-methylstyrene. PMID- 12120566 TI - Ion pair recognition by Zn-porphyrin/crown ether conjugates: visible sensing of sodium cyanide. AB - Synthesis and complexation behavior of ditopic neutral receptors composed of both a Lewis-acidic binding site (zinc porphyrin moiety) and a Lewis-basic binding site (crown ether moiety) are reported; the receptors bound only NaCN in a ditopic fashion with a color change, and in contrast other sodium salts bound to the receptors in a monotopic fashion without a color change. PMID- 12120567 TI - Synthesis of gamma-methylene oxacycles and alpha- and beta-alkylidene lactones via silicon-assisted ring opening of cyclopropyl carbinols. AB - Cyclopropyl carbinols bearing a (tert-butyldiphenylsilyl)methyl substituent undergo silicon-assisted regioselective ring cleavage and the resulting beta silyl carbocation is intramolecularly trapped with hydroxy and ester functions to generate gamma-methylene oxacycles and alpha- and beta-alkylidene lactones without the cleavage of the silicon function. PMID- 12120568 TI - First Ritter-type reaction of alkylbenzenes using N-hydroxyphthalimide as a key catalyst. AB - The first Ritter-type reaction of alkylbenzenes with nitriles has been successfully achieved by the use of N-hydroxyphthalimide (NHPI) as a key catalyst. Thus, treatment of ethylbenzene with ammonium hexanitratocerate(IV) (CAN) in the presence of a catalytic amount of NHPI in EtCN under argon produced the corresponding amide in good selectivity. PMID- 12120569 TI - New examples of mixed seleno-sulfides; reactions with triphenylphosphine. AB - The formation of mixed seleno-sulfides by means of activation of methanephosphonoseleno(thio)ic acids with arylsulfenyl chloride is rationalized on the basis of NMR and identification of the products of their reactions with triphenylphosphine. PMID- 12120570 TI - The first perfluoroacetylacetonate metal complexes: as unexpectedly robust as tricky to make. AB - First metal complexes containing the perfluoroacetylacetonato ligand have been prepared by the reaction of anhydrous Ln(OAc)3 (Ln = Eu, Tb, Tm) with heptafluoroacetylacetone, optionally in the presence of other ligands Ph3PO, bpy, and PyO. PMID- 12120571 TI - Practical tethering of vitamin B1 on a silica surface via its phosphate group and evaluation of its activity. AB - A convenient immobilization of thiamine pyrophosphate molecules on a silica surface through the phosphate group is developed, leading to a very active heterogenized biocatalyst for pyruvate decarboxylation. PMID- 12120572 TI - Amplification of a cyclic mixed-metalloporphyrin tetramer from a dynamic combinatorial library through orthogonal metal coordination. AB - A cyclic porphyrin tetramer, consisting of two bis-phosphine substituted zinc(II) porphyrin units and two Rh(III)TPP units, is selected and amplified virtually quantitatively from a dynamic combinatorial library using 4,4'-bipy as a scaffold and using orthogonal binding modes. PMID- 12120573 TI - Enantioselective synthesis of 2-arylpiperidines from chiral lactams. A concise synthesis of (-)-anabasine. AB - Cyclodehydration of achiral or racemic aryl-delta-oxoacids with (R) phenylglycinol stereoselectively affords chiral non-racemic bicyclic lactams, from which the enantiodivergent synthesis of (R)- and (S)-2-phenylpiperidine, the diastereodivergent synthesis of cis- and trans-3-ethyl-2-phenylpiperidine, and the enantioselective synthesis of the piperidine alkaloid (-)-anabasine is reported. PMID- 12120574 TI - Discovery of a novel synthetic phosphatase from a bead-bound combinatorial library. AB - Using split/pool encoded synthesis and a colorimetric catalysis assay, a number of synthetic phosphatase catalysts were developed. PMID- 12120576 TI - Promotion of cyclization of linear pentapeptides and heptapeptide by different univalent metal ions. AB - Univalent metal ions such as Na+, K+ and Cs+ can enhance not only the cyclization yields of some linear pentapeptides and heptapeptide but also their cyclization rates while some bivalent and trivalent metal ions such as Mg2+, Ca2+, Zn2+, Fe2+, Ni2+ and Cr3+ elevate neither the cyclization yields nor the cyclization rates and some of them prevent the cyclization. PMID- 12120575 TI - Oligodeoxynucleotides containing amide-linked LNA-type dinucleotides: synthesis and high-affinity nucleic acid hybridization. AB - Synthesis of three different amide-linked LNA-type dinucleotides and their incorporation into 9-mer and 17-mer oligodeoxynucleotides is described; compared to the reference DNA-RNA duplex, incorporation of one of the three dimers (5' DNA*LNA dimer) induced significantly increased duplex thermostabilities. PMID- 12120577 TI - Hollow nanoparticles via stepwise complexation and selective decomplexation of poly(ethylene imine). AB - The preparation of hollow nanoparticles with amino groups on the inner side via the stepwise complexation and selective decomplexation of poly(ethylene imine) is presented. PMID- 12120578 TI - Copper(I) catalysed cyclisation of unsaturated N-benzoyloxyamines: an aminohydroxylation via radicals. AB - A new catalytic aminohydroxylation via radicals has been developed leading to cyclic aminoalcohols. PMID- 12120579 TI - Novel Cp2LnX-mediated coupling-cyclization of propargyl bromide: a new construction of the benzene ring skeleton. AB - In the presence of Cp2LnX-HgCl2, the treatment of RC identical to CCH2Br with Mg leads to the formation of benzene derivatives C6H4R2-1,2 (R = H, Ph) in moderate yield, which provides a new method for the construction of the benzene ring skeleton. PMID- 12120580 TI - Mechanistic switch leading to highly efficient chirality transfer in Pd(0) catalyzed coupling-cyclization of aryl iodides with 1:1 acid-base salts of 2,3 allenoic acids and L-(-)-cinchonidine or D-(+)-/L-(-)-alpha-methylbenzylamine. Enantioselective synthesis of highly optically active 3-aryl polysubstituted butenolides. AB - An efficient methodology provides an easy access to highly optically active polysubstituted butenolides starting from aryl halides and 1:1:salts of optically active 2,3-allenoic acid-base via an oxidative addition-coordinative cyclization reductive elimination mechanism, which led to the high efficiency of this chirality isomerization reaction. PMID- 12120581 TI - Reduction of chromium in ethylene polymerisation using bis(imido)chromium(VI) catalyst precursors. AB - Hybrid density functional calculations on [Cr(NR)2C3H7(C2H4)]+ (R = H, tBu) have revealed a facile reductive elimination reaction involving beta-hydrogen transfer from the alkyl chain, suggesting that the active species in ethylene polymerisation with bis(imido)chromium(VI) precursors contains a reduced chromium atom. PMID- 12120584 TI - Proceedings of the International Symposium on Biological Reactive Intermediates VI. Paris, France, 2000. PMID- 12120582 TI - Helical structure of heterochiral RNA dimers: helical sense of ApA is determined by chirality of 3'-end residue. AB - Heterochiral ApAs (ALpAL and ALpAL) have been synthesized and investigation of their helical structures by means of spectroscopic techniques indicates that the chirality of the 3'-end residue is the primary factor for determining the helical sense of ApA. PMID- 12120586 TI - Hematology 2001. American Society of Hematology Education Program Book. Orlando, Florida, USA. December 7-11, 2001. Proceedings. PMID- 12120585 TI - Proceedings of the VIII International Symposium on Nidoviruses (Coronaviruses and Arteriviruses). May 20-25, 2000. Lake Harmony, Pennsylvania, USA. PMID- 12120587 TI - From measurement to clinical outcome. Festschrift in honour of Per Bech on the occasion of his 60th birthday. PMID- 12120588 TI - Nitric Oxide: From Molecular Level to Clinical Application. September 17-20, 2001. Abstracts. PMID- 12120589 TI - The chemistry of the cultural heritage environment, diagnostics and conservation. Proceedings of the conference organized by the Division of Environmental Chemistry of the Italian Chemical Society and the Free University of Urbino. Urbino, Italy. October 4-5, 2000. PMID- 12120590 TI - Barriers to incident reporting. PMID- 12120591 TI - [Galactosemia]. PMID- 12120592 TI - Proceedings of the 2nd Israeli-Polish Neurological Conference. Tel Aviv, Israel. 23 June 2000. PMID- 12120593 TI - Variability among Cryptosporidium parvum genotype 1 and 2 immunodominant surface glycoproteins. AB - Published genomic differences between Cryptosporidium parvum genotype 1 (human derived) and genotype 2 (animal and human-derived) isolates suggest that these may belong to two distinct species. This is of significant interest since genotype 1 isolates are associated with sporadic cases of human cryptosporidiosis in 30-40 % of cases in contrast to 60-70 % of cases caused by genotype 2. The lower genetic sequence similarity between genotype 1 and 2 surface glycoproteins (gp40/15) suggests that antigenic differences should also occur, a feature that was investigated in this study. Using immune and convalescent serum samples from gnotobiotic piglets previously inoculated with genotype 1 and 2 isolates, we demonstrated that C. parvum gp15 was immunodominant for both genotype 1 and 2 isolates. Lower genetic sequence similarity between genotype 1 and 2 Cpgp40/15 did correspond to gp15 protein differences as detected by Western blot. Moreover, we confirmed that gp15 contains epitopes that are also immunodominant. Deglycosylation of C. parvum proteins resulted in decreased ability of gp15, gp23 and gp900 to react with homologous polyclonal antibodies, suggesting that these proteins also express carbohydrate epitopes. Taken together, our data suggest that there is a high phenotypic variability between C. parvum genotype 1 and 2 isolates at the level of gp15. We contemplate that gp15 surface glycoprotein plays an important role in the biology of C. parvum as a potent inducer of immune response and a possible virulence factor. PMID- 12120594 TI - Abstracts of the International Workshop on Nijmegen Breakage Syndrome. Prague, Czech Republic, 25-28 April 2002. PMID- 12120595 TI - Proceedings of a festschrift honouring the scientific career of George H. Collier. 18 October 2001, New York, New York, USA. PMID- 12120596 TI - [VIII Symposium on health hazards in the workplace. May 30-June 1, 2001, Lodz]. PMID- 12120597 TI - Abstracts of the 1st Congress of Gastroenterology and Hepatology of Bosnia and Herzegovina. Sarajevo, May 29-31, 2002. PMID- 12120598 TI - Shadows. PMID- 12120599 TI - Keeping kids safe. Injury prevention programs in EMS. PMID- 12120600 TI - Upbeat generation. BLS journey in ozone park. PMID- 12120601 TI - Poisoning experts: who do you call? PMID- 12120602 TI - No room in the womb. PMID- 12120603 TI - Interest of a new salicylic acid derivative in the prevention of acne relapses. PMID- 12120604 TI - Efficacy and skin tolerability of Saliker in the treatment of dandruff. PMID- 12120605 TI - A test battery to insure the safety of topical products especially designed for sensitive skin: application for the development of a daily cream. PMID- 12120606 TI - In vitro effect of a spa water on the migratory and stimulatory capacities of human Langerhans calls. PMID- 12120607 TI - Modulatory effects of selenium and strontium salts on keratinocyte-derived inflammatory cytokines. PMID- 12120608 TI - Ocular tolerance and efficiency of two solutions applied to non-infectious blepharitis. PMID- 12120609 TI - In vivo determination of the skin protection capacity of a day-cream by means of ICL-S. PMID- 12120611 TI - Efficacy and safety of a new salicylic acid derivative as a complement of vitamin A acid in acne treatment. PMID- 12120612 TI - Nudging acne by topical beta-lipohydroxy acid (LHA), a new comedolytic agent. PMID- 12120610 TI - Dermo-epidermal stimulation elicited by a salicylic acid derivative: a comparison with salicylic acid and all trans-retinoic acid. PMID- 12120614 TI - Chronic UVA exposure: protective effect on skin induced pigmentation by a daily use of a day care cream containing broad band sunscreen. PMID- 12120613 TI - UV-induced production of immunosuppressive mediators in human skin: prevention by a broadspectrum sunscreen. PMID- 12120615 TI - Decrease in skin ascorbic acid concentration with age. PMID- 12120616 TI - Interest of a 5% vitamin C w/o emulsion in the treatment of skin aging: effects on skin relief. PMID- 12120617 TI - Topically applied ascorbic acid helps to restructure chronically photodamaged human skin. PMID- 12120618 TI - Effect of a water/oil emulsion containing ascorbic acid on collagen neosynthesis in human full thickness skin discs in culture. PMID- 12120620 TI - A comparative ultrastructural study of hydroxyacids induced desquamation. PMID- 12120619 TI - Stimulation of collagen biosynthesis by topically applied vitamin C. PMID- 12120621 TI - Mechanism of action of a lipophilic salicylic acid derivative on normal skin. PMID- 12120623 TI - The high frequency of coeliac disease and other autoimmune diseases in subjects affected by Type I (insulin-dependent) diabetes mellitus and in their first degree relatives. PMID- 12120622 TI - Why is the ADA classification of diabetes mellitus not always applied? PMID- 12120624 TI - Complex carbohydrates--structure and function with respect to the glycoconjugate composition of the cupula of the semicircular canals. AB - The review briefly describes the structure and function of complex carbohydrates of glycoproteins and proteoglycans both in general and with particular respect to the potential roles sugar chains may play in the cupula, i.e. the molecular organization of these constituents, their biophysical properties, and their biological functions. PMID- 12120626 TI - [Extra pounds despite sports and diets. Genetics can also be involved]. PMID- 12120625 TI - [Guidelines for cardiovascular prevention in diabetic patients]. PMID- 12120627 TI - [Simple corrections in therapy of type 2 diabetes. Lowering HbA1c below 7%]. PMID- 12120628 TI - The time it takes to detect changes in speed and direction of visual motion. AB - We studied the ability of human observers to detect abrupt changes in velocity of motion of a random dot pattern. The pattern moved horizontally for 0.9 s at velocity V0, then changed to V1 either in speed, or in direction for a time T and returned to the initial motion. The threshold duration for detection of the change was measured for initial speeds of 2, 4, 8 and 16 deg/s. The time to detect a velocity reversal was equal to that for detection of an increase in speed by a factor of three. The time to detect an abrupt cessation of motion was equal to the time for detection of an increase in speed by a factor of two. The time to detect a direction change, the speed being constant, decreased gradually with increasing angle between V0 and V1 from 12 to 180 degrees and with increasing V0; the detection time was a function of (V1-V0) almost independent of the value of V0. This finding supports the hypothesis of Dzhafarov et al. (Percept Psychophys 1993;54:373-750), that the visual system effectively reduces the detection of velocity changes (from V0 to V1) to the presumably more simple detection of a motion onset, from 0 to (V1-V0). The characteristics of the detection process in the cases of uni- and two-dimensional velocity changes are discussed. PMID- 12120629 TI - Atraumatic bloodless removal of intramedullary hemangioblastomas of the spinal cord. AB - OBJECT: The goal of this study was to summarize the author's personal experience in the surgical treatment of 19 patients with intramedullary spinal cord hemangioblastomas. METHODS: All cases were from the author's private practice and were treated between 1967 and 1990. In all cases the intramedullary hemangioblastomas were totally removed by using a unique microsurgical technique of bipolar coagulation, which is fully described in this paper. A bipolar forceps was used to shrink each tumor and detach it from its feeding and draining vessels. Tumor resection was successfully accomplished in all patients. Blood loss was minimal, averaging less than 100 ml, and what little bleeding occurred did so during laminotomy or laminectomy. No bleeding occurred during tumor removal, and no transfusions were given. All patients were symptomatic preoperatively, and all recovered or improved following surgery. CONCLUSIONS: The technique described in this paper makes tumor removal safe, effective, and relatively easy. PMID- 12120630 TI - Unusual case of extradural choroid plexus papilloma of the sacral canal. Case report. AB - An unusual case of a sacral, extradural choroid plexus papilloma involving the S1 3 level is described. This 50-year-old woman presented with a 4-month history of pain involving her right buttock, perineum, and leg. Contrast-enhanced magnetic resonance (MR) imaging of the spine revealed a well-defined, mildly enhancing sacral canal mass at the S1-3 level; its appearance was consistent with that of a benign tumor. Intraoperatively, the lesion was found to be extradural in location and was entwined among nerve roots in the sacral canal. Microscopic examination of the gross totally resected tumor revealed typical features of a choroid plexus papilloma. Despite performing a thorough neuroimaging workup (craniospinal contrast-enhanced MR imaging) for an intracranial or spinal primary mass, none was found. The choroid plexus appeared entirely normal; however, both a cavum septum pellucidum and a cavum vergae were noted. Extraneural choroid plexus papilloma, specifically intrasacral, extradural choroid plexus papilloma has not been previously reported. The present example is thought to have arisen either from ectopic choroid plexus tissue or perhaps by metaplasia from ependymal rests. PMID- 12120631 TI - Vertebral metastatic chemodectoma: imaging and therapeutic octreotide. Case report. AB - The authors report on the use of external-beam radiotherapy and octreotide in a 32-year-old woman who presented with spinal cord compression secondary to metastatic chemodectoma. Scintigraphy studies were used to confirm the presence of somatostatin receptors. Magnetic resonance imaging, and in particular spinal angiography, were performed to define the extent of spinal metastatic disease. The literature on current investigation and management of vertebral metastatic chemodectoma is reviewed. PMID- 12120632 TI - Pilocytic astrocytoma of a spinal nerve root. Case report. AB - A case of pilocytic astrocytoma involving a spinal nerve root is reported. A 39 year-old woman presented with a 1-year history of progressive pain and numbness, predominantly in the S-1 dermatome. Magnetic resonance (MR) imaging revealed an intradural lesion at the tip of the conus medullaris. The intradural tumor was excised as was the sacrificed nerve root. Histological examination showed a pilocytic astrocytoma in which there were unusual features of calcification and ossification. At 3-year follow-up review MR imaging demonstrated no residual tumor. To the best of the authors' knowledge, this is the first case of a primary pilocytic astrocytoma, a tumor typically of central nervous system origin, arising from a spinal nerve root. PMID- 12120633 TI - Delayed intracranial migration of cervical sublaminar and interspinous wires and subsequent cerebellar abscess. Case report. AB - Delayed complications associated with sublaminar and interspinous wiring in the pediatric cervical spine are rare. The authors present a case of delayed complication in which a cervical fusion wire migrated into the cerebellum, causing subsequent cerebellar abscess 2 years after posterior cervical arthrodesis. A craniotomy was required to remove the wire and drain the abscess. Despite their history of safety and successful fusion, procedures involving sublaminar and interspinous wiring carry a risk of neurological injury secondary to wire migration. A thorough neuroimaging evaluation is required in patients who have undergone fusion and who have neurological complaints to detect late instrumentation-related sequelae. PMID- 12120634 TI - Circumferential fracture of the skull base causing craniocervical dislocation. Case report. AB - Fractures of the craniocervical junction are common in victims of high-speed motor vehicle accidents; indeed, injury to this area is often fatal. The authors present the unusual case of a young woman who sustained a circumferential fracture of the craniocervical junction. Despite significant trauma to this area, she suffered remarkably minor neurological impairment and made an excellent recovery. Her injuries, treatment, and outcome, as well as a review of the literature with regard to injuries at the craniocervical junction, are discussed. PMID- 12120635 TI - Use of an aiming device in posterior atlantoaxial transarticular screw fixation. Technical note. AB - Posterior atlantoaxial transarticular screw fixation is an excellent procedure associated with high fusion rates. There is, however, the potential risk of vertebral artery (VA) injury. The authors designed a special aiming device that allows a cannulated screw to be inserted accurately in the most posterior part of the C1-2 joint via the most posterior and medial part of the isthmus of C-2; this screw pathway most safely avoids VA injury. The instruments include an aiming device and a flexible screw-inserting system. The tip of the aiming device is placed on the ridge of the C-2 isthmus just posterior to the atlantoaxial joint. The guide wire should then pass 1 mm below the device tip. The system consists of flexible guide wires, a drill, a tap, and a screwdriver, and the screw is inserted easily via a posterior approach in which the patient's back is not obstructive. Ten patients with atlantoaxial subluxation or osteoarthritis underwent surgery in which the device was used. In all cases, the screws were inserted safely without causing VA injury, although preoperative computerized tomography (CT) reconstructions revealed a high-risk high-riding unilateral VA in three patients. Postoperative CT reconstructions demonstrated that all screws but one were inserted as planned, and successfully cleared the vertebral groove. In conclusion, this newly designed device is practical and useful for the accurate insertion of screws, thus avoiding VA injury during atlantoaxial transarticular screw fixation. PMID- 12120636 TI - Initial stability of cervical spine fixation: predictive value of a finite element model. Technical note. AB - The purpose of this study was to generate a validated finite element (FE) model of the human cervical spine to be used to analyze new implants. Digitized data obtained from computerized tomography scanning of a human cervical spine were used to generate a three-dimensional, anisotropic, linear C5-6 FE model by using a software package (ANSYS 5.4). Based on the intact model (FE/Intact), a second was generated by simulating an anterior cervical fusion and plate (ACFP) C5-6 model in which monocortical screws (FE/ACFP) were used. Loading of each FE model was simulated using pure moments of +/- 2.5 Nm in flexion/extension, axial left/right rotation, and left/right lateral bending. For validation of the models, their predicted C5-6 range of motion (ROM) was compared with the results of an earlier, corresponding in vitro study of six human spines, which were tested in the intact state and surgically altered at C5-6 with the same implants. The validated model was used to analyze the stabilizing effect of a new disc spacer, Cenius (Aesculap AG, Tuttlingen, Germany), as a stand-alone implant (FE/Cenius) and in combination with an anterior plate (FE/Cenius+ACFP). In addition, compression loads at the upper surface of the spacer were investigated using both models. As calculated by FE/Intact and FE/ACFP models, the ROM was within 1 standard deviation of the mean value of the corresponding in vitro measurements for each loading case. The FE/Cenius model predicted C5-6 ROM values of 5.5 degrees in flexion/extension, 3.1 degrees in axial rotation (left and right), and 2.9 degrees in lateral bending (left and right). Addition of an anterior plate resulted in a further decrease of ROM in each loading case. The FE/Cenius model predicted an increase of compression load in flexion and a decrease in extension, whereas in the FE/Cenius+ACFP model an increase of graft compression in extension and unloading of the graft in flexion were predicted. The current FE model predicted ROM values comparable with those obtained in vitro in the intact state as well as after simulation of an ACFP model. It predicted a stabilizing potential for a new cage, alone and in combination with an anterior plate system, and predicted the influence of both loading modality and additional instrumentation on the behavior of the interbody graft. PMID- 12120637 TI - Combined anterior-posterior fusion for cervical spondylotic myelopathy in patients with athetoid cerebral palsy. AB - OBJECT: Cervical spondylotic myelopathy (CSM) or myeloradiculopathy, frequent in adults with athetoid cerebral palsy, is a serious secondary disability in patients with an existing congenital handicap. Although several surgical procedures have been described for CSM in adults with athetoid cerebral palsy, none has had satisfying long-term results. The object of this study was to evaluate the effectiveness and safety of combined anterior-posterior fusion with wave-shaped rods and its influence on the stability of other spinal segments. METHODS: Twenty-three patients with CSM and athetoid cerebral palsy underwent posterior fusion with wave-shaped rods combined with anterior interbody fusion with internal fixation; 20 patients, 17 men and three women, were followed for more than 5 years. This procedure yielded good results. The mean follow-up period was 8.7 years (range 5-17 years). At 1-year follow-up examination, ambulation had improved in 12 patients. Upper-extremity pain, deltoid muscle weakness, and ability to self-feed improved in almost all patients. Myelopathy recurred in one patient 8.5 years after surgery. The mean motion angle at the adjacent level to the fixed segment did not change postoperatively, but the mean motion between C-1 and C-2 increased and slight atlantoaxial subluxation occurred postoperatively in five patients. CONCLUSIONS: Combined anterior-posterior fusion can effectively improve neurological function in patients with CSM and athetoid cerebral palsy, even in those with severe involuntary movements. Postoperative rigid external fixation is not required. PMID- 12120638 TI - Inverted-hook occipital clamp system in occipitocervical fixation. Technical note. AB - The authors describe an occipitocervical fixation procedure in which they use inverted occipital hooks inserted through a burr hole drilled in the squamous part of the occipital bone. Fifteen patients with unstable lesions of the occipitocervical junction underwent occipitocervical internal fixation. The mean follow-up period was 21 months (range 2-63 months). No implant failed, and postoperative immobilization was not required. The placement of a posterior occipitocervical graft (for which fusion is uncertain) can be avoided in certain conditions. PMID- 12120639 TI - A simple and reliable behavioral analysis of locomotor function after spinal cord injury in mice. Technical note. AB - To establish a simple and reliable method to assess the behavioral function after spinal cord injury (SCI) in mice, the authors used an automated animal movement analysis system, SCANET. Two different SCI lesions were created in adult female BALB/c and C57BL/6 mice by transecting both the posterior columns and the left lateral and anterior funiculi or only the left lateral and anterior funiculi at T 8. Control mice underwent laminectomy only. The SCANET system consists of a cage equipped with two crossing sensor frames arranged at different heights, by which small (M1) and large (M2) horizontal movements and the vertical movement involved in rearing (RG) can be monitored. The authors assessed locomotor function by determining the M1, M2, and RG scores; to this end, they used the SCANET system and a previously established behavior test, the 21-point open-field Basso-Beattie Bresnahan (BBB) Locomotor Rating Scale. The results indicated that the RG scores were significantly and consistently different between the spinal cord-injured and control mice, irrespective of the mouse strain or injury model, but that M1 and M2 scores were not. Moreover, there was a statistically positive correlation between the RG score and the BBB Scale score. For the assessment of locomotor function after SCI, use of the SCANET sytem in behavioral analysis is simple and the method is highly reproducible. The analysis of vertical movement is useful for assessing the recovery of limb function in mice following thoracic hemisection. PMID- 12120640 TI - Cervicothoracic fusion performed using dual-diameter rods and polyaxial lateral mass screws. Case illustration. PMID- 12120641 TI - Spondyloarthropathy-induced atlantoaxial collapse. Case illustration. PMID- 12120642 TI - Synovial chondromatosis of the cervical spine. Case illustration. PMID- 12120643 TI - Solitary bone cyst in L-2. Case illustration. PMID- 12120644 TI - First appearance and sense of the term "spinal column" in ancient Egypt. Historical vignette. AB - In contemporary anatomical nomenclature terms have often been derived from an ancient perspective of the human body. The term "spinal column" was used in ancient Egypt, symbolized by the "djet column." The authors discuss the first appearance of the term "spinal column," taking the ancient Egyptian interaction between religion and daily life into consideration, and they describe the different segments of the spinal column known to the ancient Egyptians. Inspection of medical papyri provides insight into the knowledge held by ancient Egyptian physicians and embalmers. It is assumed that hieroglyphs were used to depict the different vertebral regions of the spinal column (cervical, thoracic, and lumbar). The knowledge was gleaned, in fact, in pursuit of religious goals. The djet might be an example of how anatomical knowledge can improve understanding of a symbol that was previously thought to have a primarily religious meaning. The authors maintain that modern medical knowledge is useful for making a more precise and anatomically correct interpretation of the presumed sense intended by the ancient Egyptians. PMID- 12120645 TI - Central cord syndrome. PMID- 12120646 TI - Transoral resection. PMID- 12120647 TI - Spine surgery in morbidly obese patients. AB - OBJECT: The diagnosis, treatment, and postoperative care of morbidly obese patients undergoing spinal surgery require modifications for body habitus. With a growing percentage of the United States population becoming morbidly obese, the surgeon may need elective or emergency treatment plans that address the special needs of these patients. The authors retrospectively reviewed the diagnosis, treatment, and postoperative care of the severely obese patient undergoing spinal surgery. METHODS: To assess the associated results and complications of management that required modification for body habitus, 12 patients were included in the study (nine females); the mean age was 50 years and mean weight was 320 lb. Cases of cervical (two cases), thoracic (four cases), and lumbar surgeries (six cases) were included. The follow-up period ranged from 6 months to 2 years. Patients presented with myelopathy (five cases), radicular pain and weakness (four cases), radiculopathy (two cases), and cauda equina syndrome (one patient). Chronic progressive neurological deterioration secondary to spinal cord compression was demonstrated in nine patients and acute pain and/or weakness secondary to nerve root compression was observed in three patients. CONCLUSIONS: The authors found that although morbidly obese patients may present late in the course of their symptoms and require modifications in the use of standard neuroimaging, operative facilities, and treatment plans, open mindedness and persistence can yield satisfactory results in most cases. PMID- 12120648 TI - Traumatic central cord syndrome: results of surgical management. AB - OBJECT: The authors compare clinical outcomes demonstrated in patients with traumatic central cord syndrome (CCS) who underwent early (< or = 24 hours after injury) or late (> 24 hours after injury) surgery. METHODS: The clinical characteristics, radiographic findings, surgery-related results, length of hospital stay (LOS), and clinical outcomes obtained in 50 patients with surgically treated traumatic CCS were reviewed retrospectively. Shorter intensive care unit (ICU) stay and LOS were observed in all patients who underwent early surgery compared with those who underwent late surgery. In patients with CCS secondary to acute disc herniation or fracture/dislocation who underwent early surgery significantly greater overall motor improvement was observed than in those who underwent late surgery (p = 0.04). Overall motor outcome in patients with CCS secondary to spinal stenosis or spondylosis who underwent early surgery was not significantly different from that in those who underwent late surgery (p = 0.51). Worse motor outcomes were found in patients who were older than 60 years of age and in whom initial bladder dysfunction was present (p = 0.03 and 0.02, respectively) compared with younger patients without bladder dysfunction. CONCLUSIONS: Early surgery is safe and more cost effective than late surgery for the treatment of traumatic CCS, based on ICU stay and LOS and improved overall motor recovery, in patients whose CCS was related to acute disc herniation or fracture. In the setting of spinal stenosis or spondylosis, early surgery was safe but did not improve motor outcome compared with late surgery. PMID- 12120649 TI - Magnetic resonance imaging correlation in pediatric spinal cord injury without radiographic abnormality. AB - OBJECT: The authors conducted a study to determine correlations between clinical syndromes and early magnetic resonance (MR) imaging-documented findings in children with spinal cord injury without radiographic abnormality (SCIWORA). METHODS: The authors retrospectively reviewed the records obtained in 20 patients who presented with SCIWORA to the Children's Hospital of Buffalo between 1992 and 1999. Initial neurological syndromes, subsequent hospital course and outcome, and early MR imaging findings obtained using conventional sequences on a 1.5-tesla unit were recorded. Neurological syndromes on presentation were complete (Frankel Grade A) in two patients (10%), severe partial (Frankel Grade C) in one patient (5%), and mild partial (Frankel Grade D) in 17 patients (85%). Partial neurological deficits resolved in 14 (78%) of 18 patients within 72 hours and lasted more than 72 hours in four patients (22%). Magnetic resonance imaging was performed in both patients presenting with complete injuries and in 17 of 18 patients presenting with partial neurological deficits. The studies were obtained within 24 hours in 17 patients (85%). Neuroimaging revealed spinal cord swelling at the cervical level in one of the children with complete injury and cord edema with associated hemorrhage at cervical and thoracic levels in the other. Neural and extraneural elements were shown to be normal in all 17 patients with partial injuries who underwent MR imaging, including in the four patients with partial motor deficits lasting more than 72 hours. CONCLUSIONS: In this series, the predominant neurological presentation of SCIWORA was a mild, partial syndrome that resolved within 72 hours. Magnetic resonance imaging revealed abnormal features only in those patients with complete neurological deficits. These findings suggest that in the acute setting conventional MR imaging sequences may lack the sensitivity to demonstrate neural and extraneural abnormalities associated with partial or temporary neurological deficits of SCIWORA, even when those deficits persist beyond 72 hours. PMID- 12120650 TI - Bone morphogenetic protein-2 application by a poly(D,L-lactide)-coated interbody cage: in vivo results of a new carrier for growth factors. AB - OBJECT: Growth factors such as bone morphogenetic protein-2 (BMP-2) have been proven to promote spine fusion and to overcome the disadvantages of an autologous bone graft. The optimum method to deliver such growth factors remains a matter of discussion. The purpose of this study was to determine the safety and efficacy of a new poly(D,L-lactide) (PDLLA) carrier system for BMP-2 and to compare this carrier system with a collagen sponge carrier in a sheep cervical spine interbody fusion model. METHODS: Thirty-two sheep underwent C3-4 discectomy and fusion: Group 1, titanium cage (eight animals); Group 2, titanium cage coated with a PDLLA carrier (eight animals); Group 3, titanium cage coated with a PDLLA carrier including BMP-2 (150 microg) (eight animals); and Group 4, titanium cage combined with a collagen sponge carrier including BMP-2 (150 microg) (eight animals). Blood samples, body weight, and temperature were assessed. Radiographs were obtained pre- and postoperatively and after 1, 2, 4, 8, and 12 weeks. At the same time points, disc space height, intervertebral angle, and lordosis angle were measured. After the sheep were killed 12 weeks postoperatively, flexion-extension radiography was performed to evaluate fusion sites. Quantitative computerized tomography scans were obtained to assess bone mineral density (BMD), bone mineral content (BMC), and bone callus volume (BCV). Biomechanical testing was performed in flexion, extension, axial rotation, and lateral bending. Stiffness, range of motion, neutral, and elastic zone were determined. Histomorphological and morphometrical analyses were performed, and polychrome sequential labeling was used to determine the timeframe of new bone formation. There were no differences among the groups concerning blood counts, body weight, and temperature. Compared with the noncoated cages, all PDLLA-coated cages showed significantly higher values for BMD of the callus, as well as slightly higher values for BMC, BCV, and the bone volume/total volume ratio. In comparison with the cage-alone group, the BMP-2 groups showed significantly higher values for BMD and biomechanical stiffness. Histomorphological, -morphometrical, and polychrome sequential labeling analyses demonstrated greater progression of callus formation in the BMP 2 groups than in any other group. Compared with BMP-2 delivered using a collagen sponge carrier, BMP-2 application with a PDLLA carrier resulted in a higher BCV and a greater progression of interbody callus formation in the histomorphometrical analysis. CONCLUSIONS: The use of cervical spine interbody fusion cages coated with PDLLA as a delivery system for growth factors was effective. In this 12-week follow-up study, the PDLLA coating showed no adverse effects. The slight but not significant positive effect of the PDLLA carrier on interbody fusion might be a result of the degradation process of the biodegradable carrier. Compared with collagen sponge delivery of BMP-2, the PDLLA coated interbody cages significantly increased the results of interbody bone matrix formation. In this new combination (implant + PDLLA + growth factor) the cage represents a "real fusion" cage, because it not only serves as a mechanical device for spinal fixation but also as a local drug delivery system. PMID- 12120651 TI - Ultrastructural scoring of graded acute spinal cord injury in the rat. AB - OBJECT: There is a need for an accurate quantitative histological technique that also provides information on neurons, axons, vascular endothelium, and subcellular organelles after spinal cord injury (SCI). In this paper the authors describe an objective, quantifiable technique for determining the severity of SCI. The usefulness of ultrastructural scoring of acute SCI was assessed in a rat model of contusion injury. METHODS: Spinal cords underwent acute contusion injury by using varying weights to produce graded SCI. Adult Wistar rats were divided into five groups. In the first group control animals underwent laminectomy only, after which nontraumatized spinal cord samples were obtained 8 hours postsurgery. The weight-drop technique was used to produce 10-, 25-, 50-, and 100-g/cm injuries. Spinal cord samples were also obtained in the different trauma groups 8 hours after injury. Behavioral assessment and ultrastructural evaluation were performed in all groups. When the intensity of the traumatic injury was increased, behavioral responses showed a decreasing trend. A similar significant negative correlation was observed between trauma-related intensity and ultrastructural scores. CONCLUSIONS: In the present study the authors characterize quantitative ultrastructural scoring of SCI in the acute, early postinjury period. Analysis of these results suggests that this method is useful in evaluating the degree of trauma and the effectiveness of pharmacotherapy in neuroprotection studies. PMID- 12120652 TI - Preinjury cervical alignment affecting spinal trauma. AB - OBJECT: The authors tested the hypothesis that initial alignment of the head-neck complex affects cervical spine injury mechanism, trauma rating, injury classification based on stability, and fracture pattern. METHODS: Thirty intact human cadaveric head-neck complexes were prepared by fixing the thoracic end in polymethylmethacrylate. The cranium was unconstrained. The initial spinal alignment was described in terms of eccentricity, defined as the anteroposterior position of the occipital condyles with respect to the T-1 vertebral body. The specimens were subjected to impact loading delivered using an electrohydraulic testing device. Outcomes after injury were identified using radiography and computerized tomography. The mechanisms of injury were classified according to fracture pattern into compression-extension, compression-flexion, hyperflexion, and vertical compression. Trauma was graded according to the Abbreviated Injury Scale rating system. Based on clinical assessment, injuries were classified as stable or unstable. Injuries were also classified into bone fracture or nonfracture groups. Analysis of variance tests were used to determine the influence of eccentricity on spinal injury outcomes. Eccentricity significantly influenced the mechanism of injury (p < 0.0001), trauma rating (p < 0.005), and fracture (p < 0.0001) classification. Statistically significant differences, however, were not apparent when the classification of injury was based on stability considerations. CONCLUSIONS: Spinal alignment is a strong determinant of the biomechanics of impact-induced cervical spine injury. PMID- 12120653 TI - Functional recovery after complete contusion injury to the spinal cord and transplantation of human neuroteratocarcinoma neurons in rats. AB - OBJECT: Human neuroteratocarcinoma (hNT)-derived neurons are differentiated postmitotic neurons derived from a human teratocarcinoma cell line following treatment with retinoic acid. In preclinical transplantation studies investigators have demonstrated both their safety as a source of neurons for transplantation and efficacy in treating stroke-related behavioral deficits. The objective of this study was to examine whether hNT neurons transplanted in an area of complete spinal cord contusion would improve electrophysiological measures of spinal cord function. METHODS: Complete spinal cord contusion injury, defined as the complete loss of motor evoked potentials (MEPs), was produced in 30 rats at T-8. Ten rats with contused spinal cords underwent transplantation with hNT neurons within the site of contusion immediately after injury (immediate transplant group). Ten rats underwent hNT neuron transplantation following a 2 week evaluation for loss of MEPs (delayed transplant group). Ten other rats with contusion injury served as a spinal cord injury control group, and 10 rats underwent only a T-8 laminectomy and served as non-injured controls. All rats survived 8 weeks after transplantation. In the delayed transplant group significant functional recovery was observed, as demonstrated by return of MEPs and a modest improvement of motor function. Immunohistochemical analysis showed the survival, integration, and long fiber outgrowth of the grafted hNT neurons. CONCLUSIONS: These findings suggest that the transplantation of the hNT neurons may be an effective means of reestablishing electrical connectivity of the injured spinal cord. PMID- 12120654 TI - A collagen-based sealant to prevent in vivo reformation of epidural scar adhesions in an adult rat laminectomy model. AB - OBJECT: The authors investigated the effect of a collagen-based sealant, Gel Amidon Oxyde (GAO), in preventing the reformation of epidural scar adhesions in an adult rat model of laminectomy. METHODS: Thirty-two adult Sprague-Dawley rats underwent a complete L5-6 laminectomy, after which the dura mater was exposed and the left adjacent L-4 and L-5 nerve roots were exposed. The surgical wound was then closed; 1 month later it was reopened. The epidural scar adhesions that developed were observed and carefully removed, leaving clean dura and nerve roots reexposed. In 16 experimental rats, GAO was placed onto the reexposed dura and around the nerve roots before it polymerized. No treatment was performed in 16 control rats. Postoperatively, all rats were healthy and without neurological deficit. The incisions healed within 1 week regardless of the treatment with the GAO. Three months after reoperation, magnetic resonance imaging revealed that important epidural adhesions were present in the control rats but not in the experimental rats. These findings were then confirmed by gross anatomical examination in which a white tissue layer was found over the dura without adhesions in the experimental animals, whereas significant epidural scar adhesions were demonstrated in the controls. Histological evaluation of the laminectomy site also showed that the peridural space in the experimental rats was larger than that in the controls. CONCLUSIONS: The authors found that GAO may be a safe and effective antiscarring adhesion biomaterial in vivo. When placed into the laminectomy site, GAO may prove beneficial in preventing the formation and reformation of epidural scar adhesions in humans. PMID- 12120655 TI - Percutaneous pedicle screw fixation of the lumbar spine: preliminary clinical results. AB - OBJECT: Standard techniques for pedicle screw fixation of the lumbar spine involve open exposures and extensive muscle dissection. The purpose of this study was to report the initial clinical experience with a novel device for percutaneous posterior fixation of the lumbar spine. METHODS: An existing multiaxial lumbar pedicle screw system was modified to allow screws to be placed percutaneously by using an extension sleeve that permits remote manipulation of the polyaxial screw heads and remote engagement of the screw-locking mechanism. A unique rod-insertion device was developed that linked to the screw extension sleeves, allowing for a precut and -contoured rod to be placed through a small stab wound. Because the insertion device relies on the geometrical constraint of the rod pathway through the screw heads, minimal manipulation is required to place the rods in a standard submuscular position, there is essentially no muscle dissection, and the need for direct visual feedback is avoided. Twelve patients (six men and six women) who ranged in age from 23 to 68 years underwent pedicle screw fixation in which the rod-insertion device was used. Spondylolisthesis was present in 10 patients and osseous nonunion of a prior interbody fusion was present in two. All patients underwent successful percutaneous fixation. Ten patients underwent single-level fusions (six at L5-S1, three at L4-5, and one at L2-3), and two underwent two-level fusions (one from L3-5 and the other from L4 S1). The follow-up period ranged from 10 to 19 months (mean 13.8 months). CONCLUSIONS: Although percutaneous lumbar pedicle screw placement has been described previously, longitudinal connector (rod or plate) insertion has been more problematic. The device used in this study allows for straightforward placement of lumbar pedicle screws and rods through percutaneous stab wounds. Paraspinous tissue trauma is minimized without compromising the quality of spinal fixation. Preliminary experience involving the use of this device has been promising. PMID- 12120656 TI - Oxidative stress and heat shock protein response in human paraspinal muscles during retraction. AB - OBJECT: The need for wide dissection and forceful retraction of paraspinal muscles often required for posterolateral lumbar fusion and fixation may severely jeopardize the muscles, structurally and functionally. The underlying pathophysiology of muscle damage may involve both mechanical and ischemic mechanisms. On the other hand, the surgery-related stress may trigger certain protective responses within the insulted paraspinal muscles. This study was conducted to assess the relationship between the oxidative stress and the stress response mediated by heat shock protein 70 (HSP70) induction within paraspinal muscles being retracted. METHODS: Multifidus muscle specimens were surgically obtained before, during, and after retraction in patients with lumbar spondylolisthesis undergoing posterolateral lumbar fusion, pedicle fixation, and laminectomy. Muscle samples were analyzed to determine HSP70 and malondialdehyde (MDA) levels. Both HSP70 expression and MDA production within multifidus muscle cells were increased significantly by retraction. Expression of HSP70 then decreased after a peak at 1.5 hours of retraction, whereas MDA levels remained elevated even after release of retractors for reperfusion of the muscles. Analysis of histopathological and immunohistochemical evidence indicated that the decline of HSP70 synthesis within muscle cells after prolonged retraction was the result of severe muscle damage. CONCLUSIONS: Results of this study highlight the deleterious effect of intraoperative retraction on human paraspinal muscles at the cellular and molecular levels. The authors also found that intraoperative maneuvers aimed at reducing the oxidative stress within the paraspinal muscles may help to attenuate surgery-related paraspinal muscle damage. PMID- 12120657 TI - Distribution of genes for parathyroid hormone (PTH)-related peptide, Indian hedgehog, PTH receptor and patched in the process of experimental spondylosis in mice. AB - OBJECT: Little is known about the molecular mechanisms underlying the process of spondylosis. The authors determined the extent of genetic localization of major regulators of chondrogenesis such as Indian hedgehog (Ihh) and parathyroid hormone (PTH)-related peptide (PTHrP) and their receptors during the development of spondylosis in their previously established experimental mouse model. METHODS: Experimental spondylosis was induced in 5-week-old ICR mice. The cervical spines were chronologically harvested, and histological sections were prepared. Messenger (m) RNA for PTHrP, Ihh, PTH receptor (PTHR; a receptor for PTHrP), patched (Ptc; a receptor for Ihh), bone morphogenetic protein (BMP)-6, and collagen type X (COL10; a marker for mature chondrocyte) was localized in the tissue sections by performing in situ hybridization. In the early stage, mRNA for COL10, Ihh, and BMP-6 was absent; however, mRNA for PTHrP, PTHR, and Ptc was detected in the anterior margin of the cervical discs. In the late stage, evidence of COL10 mRNA began to be detected, and transcripts for Ihh, PTHrP, and BMP-6 were localized in hypertrophic chondrocytes adjacent to the bone-forming area in osteophyte. Messenger RNA for Ptc and PTHR continued to localize at this stage. In control mice, expression of these genes was absent. CONCLUSIONS: The localization of PTHrP, Ihh, BMP-6, and the receptors PTHR and Ptc demonstrated in the present experimental model indicates the possible involvement of molecular signaling by PTHrP (through the PTHR), Ihh (through the Ptc), and BMP-6 in the regulation of chondrocyte maturation leading to endochondral ossification in spondylosis. PMID- 12120658 TI - Neural function preservation and early mobilization after resection of metastatic sacral tumors and lumbosacropelvic junction reconstruction. Report of three cases. AB - The management of tumors that metastasize to the sacrum remains controversial. Typically, resection of such tumors and reconstruction of the lumbopelvic junction requires sacrifice of neural elements resulting in neurological dysfunction and prolonged periods of bed rest. This severely affects the quality of life in patients in whom there is frequently a limited life expectancy. The authors describe three patients who underwent subtotal resection of metastatic sacral tumors. Postoperatively, good outcome was demonstrated in all patients. The authors present a technique for debulking and reconstruction that provides immediate spinopelvic junction stability and allows for early mobilization. Quality of life is significantly improved compared with that resulting from either medical treatment or traditional surgery. PMID- 12120659 TI - Intradural chordoma without bone involvement. Case report and review of the literature. AB - Chordomas are most commonly of extradural origin and associated with bone destruction. The authors describe a rare case of an intradural chordoma. To the best of their knowledge, this is the first case in which multiple lesions were found intradurally without bone involvement; the lesions were disseminated through the thoracolumbosacral spinal cord and extended into the paraspinal muscles, and metastatic lesions in the cervical cord were also present. PMID- 12120661 TI - Digoxin products for oral use; revocation of conditions for marketing. Final rule. AB - The Food and Drug Administration (FDA) is revoking the regulation establishing conditions for marketing digoxin products for oral use. This regulation is no longer necessary because the products, which are new drugs, can be regulated under the approval process for new drug applications (NDAs) and abbreviated new drug applications (ANDAs) as set forth in the Federal Food, Drug, and Cosmetic Act (the act). PMID- 12120660 TI - Local recurrence after S2-3 sacrectomy in sacral chordoma. Report of four cases. AB - Excision is the treatment of choice in cases of sacral chordoma. Local recurrences, however, have often been observed even after total en bloc resection. The authors assessed outcomes in four cases of tumor recurrence in patients who underwent total en bloc S2-3 resection for sacral chordomas that were located below S-3. The primary recurrences were located at either side of the lateral portion of the remaining sacrum in all patients. In two patients in whom preoperative magnetic resonance imaging indicated no invasion of the tumor into surrounding soft tissues, recurrence in the resected end of the gluteus maximus or piriformis muscle was also observed. The authors therefore recommend that the S2-3 sacrectomy should be performed over an adequate margin, including a part of sacroiliac joints at the bilateral portions of the sacrum and soft tissues such as the gluteus maximus or piriformis muscle. PMID- 12120662 TI - Medicare program; criteria for submitting supplemental practice expense survey data under the physician fee schedule. Interim final rule with comment period. AB - This interim final rule revises criteria that we apply to supplemental survey information supplied by physician, non-physician, and supplier groups for use in determining practice expense relative value units under the physician fee schedule. This interim final rule solicits public comments on the revised criteria for supplemental surveys. PMID- 12120663 TI - Occupational injury and illness recording and reporting requirements. Final rule. AB - The Occupational Safety and Health Administration (OSHA) is revising the hearing loss recording provisions of the Occupational Injury and Illness Recording and Reporting Requirements rule published January 19, 2001 (66 FR 5916-6135), scheduled to take effect on January 1, 2003 (66 FR 52031-52034). This final rule revises the criteria for recording hearing loss cases in several ways, including requiring the recording of Standard Threshold Shifts (10 dB shifts in hearing acuity) that have resulted in a total 25 dB level of hearing above audiometric zero, averaged over the frequencies at 2000, 3000, and 4000 Hz, beginning in year 2003. PMID- 12120664 TI - Dosimetry, dosinference and doswaggery. PMID- 12120665 TI - Monte Carlo calculation of the radiation field at aircraft altitudes. AB - Energy spectra of secondary cosmic rays are calculated for aircraft altitudes and a discrete set of solar modulation parameters and rigidity cut-off values covering all possible conditions. The calculations are based on the Monte Carlo code FLUKA and on the most recent information on the interstellar cosmic ray flux including a detailed model of solar modulation. Results are compared to a large variety of experimental data obtained on the ground and aboard aircraft and balloons, such as neutron, proton, and muon spectra and yields of charged particles. Furthermore, particle fluence is converted into ambient dose equivalent and effective dose and the dependence of these quantities on height above sea level, solar modulation, and geographical location is studied. Finally, calculated dose equivalent is compared to results of comprehensive measurements performed aboard aircraft. PMID- 12120666 TI - Spectral emissions and dosimetry of metal tritide particulates. AB - Inference of intakes and doses from inhalation of metal tritide particles has come under scrutiny because of decommissioning and decontamination of US Department of Energy facilities. Since self-absorption of radiation is very significant for larger particles, interpretation of counting results of metal tritide particles by liquid scintillation requires information about emission spectra. Similarly, inference of dose requires knowledge of charged particle and photon spectra. The PENELOPE Monte Carlo radiation transport computer code was used to compute spectral emissions and other dosimetric quantities for tritide particulates of Sc, Ti, Zr, Er, and Hf. Emission fractions, radial absorbed dose distributions, specific energy distributions and related frequency-mean specific energies and lineal energies, and the emitted spectra of electrons and bremsstrahlung photons are presented for selected particulates with diameters ranging from about 0.01 microm to 25 microm. Results characterising the effects of uncertainties associated with the composition and density of the tritides are also presented. Emission spectra are used to illustrate trends in the relationship between apparent and observed activity as a function of particle type and size. Emissions from metal tritide particles are weakly penetrating, and electron emission spectra tend to 'harden' as particle size increases. Microdosimetric considerations suggest that the radiation emitted by metal tritides can be classified as a low linear energy transfer radiation source. For cells less than about 7 microm away from the surface of a metal tritide, the primary dose component is due to electrons. However, bremsstrahlung radiation may deposit some energy tens, hundreds or even thousands of micrometres away from the surface of a tritide particle. The data and analyses presented in this report will help improve the accuracy of dose determinations for particulates of five metal tritides. Future work on the spectral emissions and dosimetry of metal tritide particulates needs to consider the contributions of so-called internal bremsstrahlung, an additional form of bremsstrahlung radiation emitted during beta decay. PMID- 12120667 TI - Monitoring bremsstrahlung from 4 MeV electrons by nuclear excitation and Al2O3 thermoluminescence dosemeters. AB - Isomer excitation by gamma,gamma' reactions and aluminium oxide thermoluminescence dosemeters (TLDs) have been used to monitor bremsstrahlung from the 4 MeV electron beam of a linear accelerator type LPR4 produced on a 0.9 mm Pt converter foil. Natural indium and osmium as well as TLDs have been irradiated at different distances (2-11 cm) and angles (0 degrees-90 degrees). Dose rates measured by TLDs were 5-110 kGy x h(-1). Isomer excitation of 115In (half-life 4.5 h) was used for monitoring bremsstrahlung of energies above 1 MeV, while that of 189Os (half-life 5.8 h) extended the available range down to 200 keV. Isomer production yields measured by gamma spectrometry and found to be about 10(-19)-10(-18) Bq per nucleus were calibrated against dose rate. A graphical method based on a semiempirical formula was used to evaluate the bremsstrahlung flux as well as the dose rate from the activity of isomeric monitors with uncertainties below 20%. The method is simple, of linear response in a large scale, independent of temperature, and able to monitor extremely high gamma intensities. PMID- 12120668 TI - Application of the alkaline comet assay in biodosimetry: assessment of in vivo DNA damage in human peripheral leukocytes after a gamma radiation incident. AB - The alkaline comet assay was employed in the assessment of DNA damage in leukocytes of a worker incidently exposed to gamma radiation (221 mSv, 60Co source). The comet tail lengths and tail moments were studied. By using the alkaline comet assay immediately after accidental exposure a high level of DNA damage was recorded. The highest levels of DNA damage were recorded one day and one week after the radiation incident. Later on, a decrease in both comet parameters was observed. Although the level of DNA damage was diminished during a one year period, it was still elevated compared to normal values recorded in leukocytes of a healthy, unexposed person. The results obtained indicate that the alkaline comet assay is a rapid and sensitive microdosimetric technique and is suitable for in vivo human biomonitoring, especially in cases of incidental exposure to ionising radiation. PMID- 12120669 TI - Breast dose variability in a bi-racial population undergoing screening mammography. AB - This study evaluated individual and population dose variability during screening mammography, among 570 white and black women in South Carolina, USA. Aspects of dosimetry that were considered include compressed breast thickness (CBT), number of films per screening session, and dose in previous or subsequent sessions. Breast dose was log-normally distributed in the population, with a geometric mean of 6.6 mGy per session. Doses were significantly higher for black women, for women with high CBT or who receive more than two views per breast, and for the mediolateral oblique, compared to the craniocaudal, view. No relationship was observed between age and dose. Total dose per breast varied by a factor of 20 across the study population, but the individual's dose varied little among repeat screening sessions, especially after adjusting for the number of films received per session. These results may inform assessments of the projected risks of inducing breast cancer from screening mammography. PMID- 12120670 TI - Mammography dose in relation to body mass index, race, and menopausal status. AB - Mammography dose increases with compressed breast thickness (CBT), but few studies have examined other correlates of dose. The purpose of this study was to evaluate the relation between factors such as race, age, body mass index (BMI), CBT, and menopausal status and mammography screening dose, measured for 509 women in a US population. A multiple linear regression model was developed for dose, based on consideration of these factors as well as examination characteristics. BMI and number of films during examination were positively related to dose. After adjusting for these factors, high CBT also leads to higher dose. Whites receive lower doses than black women but differences are slight after controlling for the effects of CBT and BMI, which were significantly higher among black women. Pre menopausal women receive higher doses, after adjusting for all other factors, than post-menopausal women. Jointly, these factors account for approximately 75% to 80% of the variability in dose among this study population. Because rates of overweight are increasing in the US, average doses from mammography may be increasing as well. PMID- 12120671 TI - Measurements performed in Goiania after a new intervention action in 2001. AB - Brazil's worst radiological accident took place in 1987, in the city of Goiania. In 1999 and 2000, detailed measurements of 137Cs contamination were performed in junkyard II, one of the places involved in the accident. High values of 137Cs activity per unit mass were found in soil layers at depths between 10 and 40 cm from the surface, reaching values as high as 175 kBq x kg(-1). High values of 137Cs concentration in fruits and plants were also observed. Moreover, values of ambient dose equivalent rate at 1 m above the ground were found to be higher than the limit of 1.0 microSv x h(-1) set by the Brazilian National Nuclear Energy Commission (CNEN) in 1987. In February 2000, the CNEN was informed about the results of our measurements. As consequence, in August 2001, the CNEN performed a new intervention action in the area, covering all its extension with a concrete layer and removing some plants and trees. The new remedial action reduced the dose rate to approximately 13% of the value prior to covering the site in concrete, reaching values below the CNEN limit, as demonstrated by the measurements presented here. PMID- 12120672 TI - Evaluation of patient exposure in computerised tomogram in Poland. AB - The increasing number of computerised tomography (CT) procedures performed in Poland in recent years has resulted in a growing contribution of these examinations to the whole exposure of the population to ionising radiation from medical sources. (The number of CT examinations in Poland was 170,000 in 1995 and 460,000 in 1999.) An evaluation is presented of doses to patients in CT examinations performed with different types of CT unit. To evaluate the exposure to patients dose linear product (DLP) was measured using a NOMEX dosemeter with a pencil chamber (PTW, Frieburg) and the cylindrical PMMA phantoms 'head' and 'body'. CTDI values were evaluated according to current methodology as described in European Guidelines (EUR 16262). The measurements were performed for seven types of CT unit made by different companies. The CTDI values were also compared to reference levels recommended by IAEA. In conclusion it was found that the value of collective effective dose (2200 man.Sv), has increased in Poland nearly 4 times in comparison to 1995, whereas the number of CT examinations increased nearly 3 times in this period. For most of the 'controlled' CT scanners the values of CTDI in head procedures are near to or higher than the IAEA Reference Level (50 mGy); this can result from the protocols, which are chosen without a dose analysis. PMID- 12120674 TI - Anatomy and embryology of the external ear and their clinical correlation. AB - The vascular anatomy for design and execution of various flaps of the auricular region is outlined with emphasis on clinical correlation. A new classification of various flap compositions from the postauricular region based on histologic and anatomic observations is proposed, together with a corresponding clinical example used in different clinical situations. Sound knowledge of the vascular pattern surrounding the auricle provides immense versatility in performing flap operations in this region. A summary of the controversy and updates on auricular embryology is provided in relation to various congenital malformations. PMID- 12120673 TI - Evaluation of absorbed dose rate and annual effective dose equivalent due to terrestrial gamma radiation in rocks in a part of Southwestern Nigeria. AB - The average outdoor absorbed dose rate in air and the average annual effective dose equivalent due to terrestrial gamma radiation from 40K, 238U and 232Th in rocks in Ondo and Ekiti States, Southwestern Nigeria have been evaluated from measurements of the concentrations of these radionuclides in this environmental material. The concentration measurements were obtained using a very sensitive gamma spectroscopic system consisting of a 7.6 cm x 7.6 cm NaI(Tl) scintillation detector coupled to a computerised ACCUSPEC installation. The average absorbed dose rate and average annual effective dose equivalent was found to be 8.33 +/- 2.76 nGy x h(-1) and 8.7 +/- 2.9 microSv x y(-1) respectively. PMID- 12120675 TI - Acquired ear defects. AB - Trauma and tumor are the causes of acquired ear defects that surgeons are frequently called on to treat. Trauma may result in hematoma or laceration of the ear. In addition, both trauma and tumor excision may result in skin or cartilage loss. Prevention of recurrence depends on complete excision of the tumor m both lateral margins and depth. Tumors of the ear (as well as the nose) are often incompletely excised because of the difficulty in reconstruction. The absence of subcutaneous tissue in the ear allows fixation and perichondrial involvement to occur early. The tumor affinity for perichondrium usually prohibits penetration into the cartilage itself. PMID- 12120676 TI - Reconstruction of the ear after skin and perichondrium loss. AB - Because no two auricular defects are exactly the same, the choice of a suitable method for ear reconstruction is essential. Location and size of the defect influence the choice of technique needed for reconstruction. The method of reconstruction varies if there is skin loss, skin and perichondrium loss, or full thickness loss. The skin surrounding the defect should be examined to determined if it is lacerated, burned, or scarred to decide whether or not it can be used in reconstruction. A plan of treatment should be decided and explained fully to the patient. A small area of skin loss can be closed by undermining of the edges and direct closure. If this cannot be performed because the defect is too large, the perichondrium is then examined to decide whether or not it is intact. PMID- 12120677 TI - Reconstruction of the ear after skin and cartilage loss. AB - Small defects (less than 1.5 cm) of the helix or antihelix of the middle third of the ear can be converted to a wedge-shaped excision and primary repair performed. In some cases, small Burrow's triangles on either side of the wedge must be exised from the scapha or antihelix to allow for closure without distortion or cupping. In addition, the resection may go across the conchal rim and include the bowl to allow for rotation without deformity. PMID- 12120678 TI - Total ear reconstruction in postburn deformity. AB - The external ear enjoys a special place in society all over the world. It is meant to be flaunted and adorned. Ear piercing is routine, with a range of jewelry pieces concentrating in enhancing its natural beauty. Persons with deformed ears have to limit their range of hair styles. There is a definite need to reconstruct the deformed ear of both sexes. To achieve desirable results in ear reconstruction is a difficult task. Although cartilage fabrication is an important step in ear reconstruction in postburn deformity of the ear, the final outcome is mainly decided by the quality and quantity of skin available in the auricular region for draping of framework. PMID- 12120679 TI - Ear replantation. AB - Most trauma to the ear is minimal and can heal with excellent results if meticulous repair is performed immediately after the accident. Amputation of the ear, on the other hand, is a very serious problem and can lead to severe deformity. Many reattachment and reconstruction techniques are described; however, reattachment of the severed ear continues to be a major challenge that can lead to severe deformity. To date, only a few successful cases of microvascular ear replantation have been reported. Bone-anchored prosthesis or plastic reconstruction may be recommended if the patient refuses surgery, in patients with multiple health problems, or in the case of a reconstructive attempt that has failed and the patient prefers to wear a prosthesis for the rest of his or her life. PMID- 12120680 TI - Microvascular ear replantation. AB - Microvascular ear replantation is a rare event, having been reported only 25 times since the first case in 1980. It requires a lengthy operative time and hospital stay, results in multiple blood transfusions, and has a significant failure rate. Nevertheless, a successful ear replantation is a dramatic demonstration of the power of microsurgery to restore a lost part. When successful, it obviates the need for other complex reconstructive efforts and provides an unsurpassed aesthetic result. This article reviews the history of microsurgical and nonmicrosurgical ear replantation and presents recommendations for treatment. PMID- 12120681 TI - Reconstruction of congenital and acquired earlobe deformity. AB - The appearance and symmetry of the auricle is crucial for the maintenance of facial cosmetic harmony. The earlobe is considered to be an important attribute of beauty in most cultures, and earlobe decoration with color or earring is a common practice in many societies. A reconstructive technique for congenital or acquired deformity of the earlobe is described. In addition, the postauricular chondrocutaneous flap is very convenient for reconstruction of various auricular parts. PMID- 12120682 TI - Microtia repair with rib cartilage grafts: a review of personal experience with 1000 cases. AB - Surgical construction of the auricle with autogenous tissues is a unique marrying of science and art. Although the surgeon's facility with both sculpture and design is imperative, the surgical result is equally influenced by adherence to sound principles of plastic surgery and tissue transfer. The material reviewed in this article is derived from clinical experience with congenital microtia: 1094 completed ears in 1000 patients (94 cases were bilateral). This article focuses on total repair of major congenital ear defects, but includes relevant supplementary input from experience gained by managing more than 125 traumatic auricular deformities. PMID- 12120683 TI - Otoplasty for prominent ears. AB - Protruding ears may be a source of psychological distress in either sex and at any age. A truly gratifying psychological response to a well performed otoplasty is the rule. If the neonate with protruding ears, mildly constricted ears, Stahl's ear, or cryptotia is seen by a plastic surgeon during the baby's first days of life, the timing is auspicious. If the process of shaping the ear with molding splints and Steri-Strips is promptly initiated, correction of the problem without surgery is a realistic expectation. The urgency and the effectiveness of early nonsurgical treatment of such ears is not yet widely appreciated by those responsible for primary medical care of neonates. The plastic surgeon is in a position to increase awareness of the availability and effectiveness of this technique. PMID- 12120684 TI - The constricted ear. AB - The constricted ear may be described best as a pursestring closure of the ear. The deformity may include lidding of the upper pole with downward folding, protrusion of the concha, decreased vertical height, and low ear position relative to the face. The goals of surgical correction should include obtaining symmetry and correcting the intra-auricular anatomy. The degree of intervention is based on the severity of the deformity and may range from simple repositioning, soft tissue rearrangement, or manipulation of the cartilage. Multiple surgical techniques are described. PMID- 12120685 TI - Alternative surgical methods of treatment for the constricted ear. AB - There have been numerous articles published on surgical correction of the constricted ear where the auricular defect was described in detail or classified in addition to their suggested surgical treatment methods or techniques. As for the surgical method introduced by Stephenson and modified by Musgrave it is worthy to note that the technique for the expansion and reinforcement of the auricular cartilage is useful, especially for corrective surgery for cryptotia cases. All articles concerned with surgery involving with the auricle are of importance to young surgeons who seek to pursue a clinical practice involving corrective and reconstructive surgery of the auricle. One must consider not only the selection of the surgical method or technique but also the importance in planning the surgery to correct the defect and in the selection of the material required to correct the defect or to reconstruct the auricle. The ultimate goal is to attain consistent, satisfactory, and favorable results, the appearance of a normal auricle. PMID- 12120686 TI - Cryptotia: principles and management. AB - Adequate treatment of the cryptotic ear must address both recreation of the upper pole and correction of the vertical height. The degree of soft tissue manipulation ia dictated by the severity of the deformity. Often this must be combined with repositioning or reconstruction of the cartilaginous skeleton. PMID- 12120687 TI - Nonsurgical treatment of various auricular deformities. AB - Nonsurgical treatment does not always correct all auricular deformities. However, we believe that all types of deformities can be treated if the gradual and continuous correction is made. Therefore, it is recommended that nonsurgical treatment should be tried first, even in older children. Even if the correction with the splint is not satisfactory, the improved form with the splint will make it easier to obtain a good, delicate form by surgery at a later stage. PMID- 12120688 TI - Multiple sclerosis in its European matrix. PMID- 12120689 TI - Cerebral plasticity in multiple sclerosis: insights from fMRI. AB - Functional magnetic resonance imaging (fMRI) allows noninvasive localization of cerebral activation with relatively high spatial and temporal resolution. The considerable potential for the elucidation of the mechanisms of brain function has made it a useful tool to investigate the neural substrate of motor, sensory and cognitive functions. Understanding derived from these basic cognitive neuroscience investigations is beginning to be applied to clinically relevant problems. In this article, applications to multiple sclerosis (MS) are reviewed, which address the challenging notion that adaptive cerebral plasticity may have an important influence on the relationship between MS pathology and its clinical expression. PMID- 12120690 TI - Metabolic differences between multiple sclerosis subtypes measured by quantitative MR spectroscopy. AB - We used quantitative magnetic resonance (MR) spectroscopic imaging with T1-based image segmentation to evaluate the subtypes of multiple sclerosis (MS) (eight patients each group of relapsing-remitting [RR], secondary progressive [SP] and primary progressive [PP]). There was no significant difference in age between the PP group with the RP, SP or control group. We found that the metabolite ratio of choline/NA from the periventricular white matter region was not significantly different between the RR and SP groups. Using an ANOVA, the ratios of periventricular choline/NA or creatine/NA of these combined groups were significantly higher than the PP and control groups. Quantification of these data suggest that the major cause of the elevation of these parameters is due to an increase in choline and creatine in the RR group while NA is decreased in the SP group. Thus, early PP disease appears to be relatively intact with respect to neuronal loss. PMID- 12120691 TI - Quantitative 1H MRS imaging 14 years after presenting with a clinically isolated syndrome suggestive of multiple sclerosis. AB - clinically isolated syndromes (CIS) are events suggestive for emerging multiple sclerosis (MS). A majority of patients develop MS within months or years whilst others remain clinically isolated. The goal of this study was to investigate whether biochemical metabolites detectable by 'H magnetic resonance spectroscopy (MRS) may serve to distinguish between these two groups. We investigated 41 patients 14 years after presentation with a CIS and 21 controls with combined quantitative short echo 'H MRS and magnetic resonance imaging (MRI) and assessed disability according to the Expanded Disability Status Scale (EDSS). At follow up, 32 had developed MS, and 9 still had CIS. Compared with controls, MS patients demonstrated significantly higher concentrations of myo-inositol (Ins) in normal appearing white matter (NAWM) and lesions. Lesions also demonstrated a reduced N acetyl-aspartate (NAA) level and an increase in choline-containing compounds (Cho). The NAWM Ins concentration was correlated with EDSS (r = 0.48, p = 0.005). MS normal appearing cortical grey matter (CGM) exhibited a decreased NAA. Patients who remained CIS did not differ significantly from controls in any MRS measure. Metabolite changes in normal appearing white and grey matter in MS indicate diffuse involvement of the entire MS brain, which was not seen in the persisting CIS patients. Elevated Ins in MS NAWM appeared functionally relevant It may indicate glial cell proliferation or gliosis. PMID- 12120692 TI - T1 histograms of normal-appearing brain tissue are abnormal in early relapsing remitting multiple sclerosis. AB - OBJECTIVE: To use both whole-brain and normal-appearing brain tissue (NABT) T1 relaxation time histograms to investigate abnormalities in early relapsing remitting (RR) multiple sclerosis (MS). BACKGROUND: In patients with established MS, both lesions and NABT exhibit an increase in T1 relaxation time. By using T1 histogram analysis, it is hoped that such changes in early disease can be detected. METHOD: Twenty-seven patients and 14 age- and sex-matched controls underwent magnetic resonance imaging (MRI) of the brain, which included the following sequences: 1) proton density (PD)- and T2-weighted fast spin echo (FSE) to measure T2 lesion load, 2) PD- and T1-weighted gradient echos from which T1 relaxation was calculated, and 3) T1-weighted SE imaging pre- and post-triple dose (0.3 mmol/kg) gadolinium (Gd-DTPA) to measure T1 hypointense and gadolinium enhancing lesion loads, respectively. All patients had RR MS with disease duration <3 years (median 1.7 years). Statistical parametric mapping (SPM) 99 was used to segment brain from cerebrospinal fluid (CSF), and lesions were segmented using a local thresholding technique. RESULTS: Both whole-brain and NABT histograms were abnormal for all six T1 histogram parameters that were measured. For NABT, the mean T1 was 1,027 (+/- 74) ms in patients and 969 (+/- 41) ms in controls (p=0.003). There was little difference between the global and NABT histograms, which indicates that most of the whole-brain histogram abnormality derives from normal-appearing tissues. There was a correlation between the Nine Hole Peg Test and NABT T1 measures. CONCLUSION: There are widespread abnormalities of NABT in early RR MS, which were sensitively detected by T1 relaxation time histogram analysis. As such, T1 histogram analysis appears promising for studying the natural history of early RR MS, and in the monitoring of response to treatment PMID- 12120693 TI - Cerebral blood flow velocity changes to visual stimuli in patients with multiple sclerosis. AB - We assessed the blood flow velocity (BFv) changes to visual stimuli using transcranial Doppler (TCD) in patients with multiple sclerosis (MS) during an exacerbation period by means of vasoneuronal coupling. Eighty-four patients (19 men, 75 women) and 45 healthy subjects (14 men, 31 women) were studied. Both posterior cerebral arteries (PCAs) were simultaneously monitored by TCD sonography during 10 cycles of 20 s eyes open observing complex moving visual images, and 20 s eyes closed at the end of every cycle. TCD sonography was performed at least at the first 2 days of exacerbation. Mean cerebral BFv throughout the procedure (p=0.003, p=0.001; right and left sides, respectively), velocity at rest (p=0.001, p<0.001), and velocity at stimulation (p=0.021, p=0.01) on both PCAs were significantly lower in patients than controls. However, BFv changes to visual stimulation on both sides were significantly higher in patients (p=0.01, p=0.031) compared to controls. There were negative correlations between P100 latencies and relative blood flow changes on both sides, but it was not significant on the left side. These results may suggest that patients with MS during exacerbation have more reactive vessels in the posterior circulation and! or more reactive neurons in the occipital cortex. PMID- 12120694 TI - Intrathecal synthesis of matrix metalloproteinase-9 in patients with multiple sclerosis: implication for pathogenesis. AB - Matrix metalloproteinase-9 (MMP-9) was detected by zymography and enzyme-linked immunosorbent assay (ELISA) in matched serum and cerebrospinal fluid (CSF) samples from patients with neurological diseases. Patients with relapsing remitting multiple sclerosis (RR-MS) had serum and CSF MMP-9 levels comparable to those from patients with inflammatory neurological diseases (INDs), but higher than patients with non-inflammatory neurological diseases (NINDs) and healthy donors (HDs). MMP-9 increased in active RR-MS in comparison with inactive RR-MS implying that MMP-9 in MS is related with clinical disease activity. A correlation between the CSF/serum albumin (Q(AIb)) and CSF/serum MMP-9 (Q(MMP-9)) was observed in IND and NIND but not in RR-MS patients, indicating that CSF MMP-9 levels in NIND and IND patents could be influenced by serum MMP-9 and blood-brain barrier (BBB) permeability properties. MS patients had higher values of Q(MMP 9):Q(Alb)(MMP-9 index) than IND and NIND patients suggesting that in MS the increase in CSF MMP-9 could be due to intrathecal synthesis of MMP-9. A significant inverse correlation was found between MMP-9 and its endogenous inhibitor TIMP-1 in RR-MS indicating that in MS patients both the increase in MMP 9 and the decrease in TIMP-1 serum levels could contribute to BBB disruption and T-lymphocyte entry into the CNS. PMID- 12120695 TI - Cortisol is increased in postmortem cerebrospinal fluid of multiple sclerosis patients: relationship with cytokines and sepsis. AB - Hypothalmo-pituitary-adrenal (HPA) axis activity is altered in patients with multiple sclerosis (MS), resulting in elevated basal levels and enhanced response of cortisol in stimulation tests. HPA axis hyperactivation in MS is thought to be the result of complex interactions of genetic, immunologic, and neuroendocrinological mechanisms. In order to investigate whether cytokine levels in the central nervous system are associated with the activation of the HPA axis in MS, we measured cortisol, interleukin (IL)-6, IL-10 and TNF-alpha levels in postmortem cerebrospinal fluid (CSF) of 18 patients with severe MS and 50 controls. We also investigated the cortisol and cytokine levels in the CSF of a group of MS patients and controls who died with sepsis, in order to see whether acute infectious situations affect the association between cortisol and cytokines. The cortisol levels in MS patients were increased by 80% in comparison to controls (p=0.008). There was no difference in IL-6 levels between the groups, while IL-10 and TNF-alpha levels of the majority of subjects were below detection limits. There was a positive correlation between cortisol and IL-6 only in control patients with sepsis (r=0.89, p=0.019), but not within the MS patents with sepsis or MS and control groups without sepsis. Cortisol levels in postmortem serum and CSF were highly correlated (r>0.78, p<0.001). We concluded that the basal level of cortisol is significantly increased in the CSF of MS patients and that IL-6 is not responsible for this rise. The relationship between cortisol and IL-6 in sepsis is discussed. PMID- 12120696 TI - Anti-viral properties of interferon beta treatment in patients with multiple sclerosis. AB - Viral infections are potentially associated with the etiology and pathogenesis of multiple sclerosis (MS). It has been speculated that the treatment efficacy of interferon beta (IFN beta) in MS may relate to its anti-viral properties. The study was undertaken to evaluate the in vivo anti-viral effects of IFN beta-1a in patients with MS. Human herpesvirus-6 (HHV-6) was studied as an example for being a latent neurotropic virus. IFN beta used at concentrations of approximately 0.5 microg/ml was shown to significantly reduce in vitro HHV-6 replication in a susceptible T-cell line. Sera derived from 23 MS patients treated with IFN beta 1a were examined for serum cell-free DNA of HHV-6 as an indicator for viral replication and the reactivity of IgM antibodies to a recombinant HHV-6 virion protein containing a known immunoreactive region. The results were compared with those of control sera obtained from untreated MS (n=29) and healthy individuals (n=21). The findings indicated that IFN beta treatment significantly reduced HHV 6 replication as evident by decreased cell-free DNA in treated MS specimens. The results correlated with decreased IgM reactivity to the HHV-6 antigen in treated MS patients compared to untreated controls, suggesting reduced exposure to HHV-6. The findings were confirmed in paired sera obtained from seven MS patients before and after the treatment The study provides new evidence indicating that IFN beta has potent in vivo anti-viral effects that may contribute to the treatment efficacy in MS. PMID- 12120698 TI - Multiple sclerosis in the tropics: genetic association to STR's loci spanning the HLA and TNF. AB - Clear evidence has been presented correlating gene polymorphisms at 6p21.3-21.4 (containing HLA and TNF) and the predisposition to acquire multiple sclerosis (MS). In a previous study, we found that polymorphisms at HLA DQAI were associated with being or not being predisposed to MS in individuals inhabiting the tropics, where the prevalence of MS is significantly lower than in subtropical areas. Here, we tested the hypothesis that polymorphisms at D6S276, D6S265, D6S273 and D6S291 microsatellite loci are in strong linkage disequilibrium with a major genetic factor predisposing to MS. These microsatellites span the 6p21.3 region with intervals of 5 cM establishing particular landmarks for the HLA and TNF loci. Thirty-five MS patients and 35 controls, age, sex, social, ethnically and geographically matched healthy individuals, were studied. After testing the fit of gene frequencies to the normal distribution and performing the correlation for multiple comparisons, we found significant differences among the case and the control frequencies for the allele 202 belonging to the marker D6S276 (Pc=0.00455) and for the allele 114 belonging to the marker D6S265 (Pc=0.0084). For these two alleles at different loci, we found higher frequencies in the cases than in the controls. A nonsignificant p value was found in testing the existence of linkage disequilibrium among the studied loci in the cases and in the controls. In conclusion, the current study adds evidence to the established association among polymorphisms of genes located at 6p21.3-21.4 and MS. Furthermore, because of the distribution of the tested microsatellite loci, the more probable critical region could be correlated with the TNF neighborhood. PMID- 12120697 TI - Thiopurine methyltransferase activity in a Spanish population sample: decrease of enzymatic activity in multiple sclerosis patients. AB - The present study was performed in order to obtain the thiopurine methyltransferase (TPMT) activity frequency distribution histogram in a Spanish population. A total of 3640 Spanish clinical laboratory samples were evaluated, which included 1249 patients with Crohn's disease, 589 with ulcerative colitis, 348 with multiple sclerosis (MS), 487 with several autoimmune diseases different from the above-mentioned diseases and 967 a donor group. We have measured the TPMT activity in red blood cells (RBCs) by a radiochemical method, using S adenosyl-L-[methyl-3H]methionine as methyl donor. The different groups present in their entirety a normal distribution histogram and a wide range of TPMT activity from 0 to 41 U/ml RBCs. The differences found between the Spanish population TPMT activity frequency distribution histogram and the pattern previously described in a North American population were not due to azathioprine treatment or gender. The effect of autoimmune diseases on TPMT activity was evaluated: the enzymatic activity was similar in the donor group (19.9 +/- 6.3 U/ml RBCs) and in the patients with Crohn's disease (20.0 +/- 5.8 U/ml RBCs) and ulcerative colitis (19.7 +/- 6.1 U/ml RBCs); however, it decreased significantly (p<0.0001) in MS patients (17.1 +/- 6.1 U/ml RBCs) with respect to the donor group. In conclusion, our results show that the Spanish population TPMT distribution is closer to that of the Jewish population of Israel than to North American populations, and that in MS the enzymatic activity of TPMT decreases significantly. This observation may take into account the usage of azathioprine as therapeutic agent in Spanish MS patients. PMID- 12120699 TI - Autonomic instability, as measured by pupillary unrest, is not associated with multiple sclerosis fatigue severity. AB - Multiple sclerosis (MS) fatigue is one of the most common symptoms in MS, but its pathophysiology is still not understood Sympathovagal imbalance was suggested as a reason for fatigue in chronic fatigue syndrome. We examined the role of an imbalance in the central autonomic nervous system (ANS) as a cause of MS fatigue in 51 MS patients and a control group of 22 healthy volunteers. Fatigue was assessed with the revised MS Fatigue Severity Scale (FSS) and the Modified Fatigue Impact Scale (MFIS). Depression was evaluated with the Beck Depression Inventory (BDI). Disintegration of the central ANS expressed by pupillary fatigue waves was measured with pupillography and documented in the pupillary unrest index (PUI). All subjects had less than five points on the seven-point Stanford Sleepiness Scale and were therefore not sleepy. MS patients had significant higher mean FSS scores (p=0.001) and mean MFIS scores (p=0.003) than our control group. Mean BDI scores were significant higher (p=0.001) in the MS group, but were in the lowest score range (0-10 points) in both groups. Surprisingly, we found a statistically significant inverse correlation between PUI values and either FSS scores (p=0.001; r=-0.521) or MFIS scores (p=0.002; r=-0.423) in the MS group, but not in healthy participants. We therefore conclude that autonomic instability, as measured by pupillary unrest is not associated with MS fatigue severity. PMID- 12120700 TI - Fluency in multiple sclerosis: which measure is best? AB - Tests of verbal fluency provide brief and sensitive measures of the deficits in rapidly retrieving overlearned information common in multiple sclerosis (MS). Production of words that begin with the letters F, A, and S is the verbal fluency measure most often used with patients who are fluent in English. However, because of frequency of words beginning with certain letters varies from one language to another, it is unlikely that any fixed set of letters will be appropriate for multicenter trials that involve patients who are fluent in different languages. A possible alternative involves using semantic fluency categories that contain such a large number of exemplars that no fluent speaker of any language could exhaust the category in the allotted response time. To examine the potential usefulness of semantic fluency measures, 203 MS patients and 87 healthy controls generated words that begin with F, A, or S or were exemplars of the categories animals and parts of the body. Receiver operating characteristic (ROC) curve analyses indicated that sensitivities and specificities for the three fluency measures in discriminating patients from controls were quite similar, especially if patients with global cognitive impairment were excluded. PMID- 12120701 TI - The use of multiple sclerosis databases at neurological university hospitals in Germany. AB - The development of easy-to-use, clinically oriented multiple sclerosis (MS) database programs has been started, thus paving the way for MS centers to computerize their patient records and to improve quality management To evaluate the prevalence of such programs at German neurological hospitals, a questionnaire was designed and sent to all clinic directors. With a return of more than 92%, it became evident that MS databases are still being used only by a minority of 22% on a regular basis. We did not recognize the predominance of a single program. A new MS database system that is being presently implemented in Germany is described. PMID- 12120702 TI - Quantitative analysis of substance P, neurokinin A and calcitonin gene-related peptide in gingival crevicular fluid associated with painful human teeth. AB - The aim of this study was to investigate the presence of substance P (SP), neurokinin A (NKA) and calcitonin gene-related peptide (CGRP) in the gingival crevicular fluid of teeth diagnosed with pain of pulpal origin compared with clinically healthy teeth, and to detect any changes in the levels of these neuropeptides in gingival crevicular fluid after removal of the pulp from the painful teeth. Gingival crevicular fluid was collected at baseline from one interproximal site at a painful and a non-painful contralateral tooth from 54 adult patients. Sampling was repeated after 1 wk in a subset of 21 subjects. Samples were analysed for SP, NKA, and CGRP using radioimmunoassay. The mean levels of SP and NKA were significantly higher in gingival crevicular fluid from painful teeth compared with non-painful teeth. The level of SP in the GCF of painful teeth fell significantly 1 wk after pulpectomy. In contralateral teeth, there were no significant differences in the levels of SP and NKA after 1 wk. It is concluded that SP and NKA are present in significantly greater amounts in the GCF of painful teeth compared with healthy teeth. PMID- 12120703 TI - Oral lichen planus has a high rate of TP53 mutations. A study of oral mucosa in icelanD. AB - Oral squamous cell carcinoma (OSCC) is a world-wide health problem. In addition to external exposure (smoking and alcohol), certain oral lesions may increase the risk of oral cancer (e.g. leukoplakia, erythroplakia, and oral lichen planus). TP53 has been implicated in OSCC, but there are limited studies of mutations in premalignant oral lesions. In this study, 55 samples from OSCC, 47 from hyperkeratotic (HK) oral mucosa, clinically diagnosed as white patches, 48 samples from oral lichen planus (OLP), and 12 biopsies from normal oral mucosa were studied immunohistochemically for expression of TP53 protein. From all the carcinoma samples and selected non-malignant samples showing moderate or strong TP53 protein expression, malignant cells or TP53-positive nuclei were microdissected and screened for mutations in exons 5-8 by constant denaturation gel electrophoresis. Moderate to strong TP53 protein staining was seen in 56% of OSCC, 32% of OLP but only in 13% of HK. All OLP samples showed a characteristic pattern of positive nuclei confined to the basal layer, whereas TP53 staining was seen in suprabasal nuclei in HK. Mutation rate was 11 out of 52 for OSCC, three out of 20 tested for HK and, remarkably, nine out 27 tested for OLP. There was no correlation between TP53 protein staining and TP53 mutations. No associations were found with anatomical sites or disease progression. The unexpectedly high mutation rate of OLP might explain the premalignant potential of this lesion. PMID- 12120704 TI - Accuracy of computer-automated caries detection in digital radiographs compared with human observers. AB - The aim of this study was to compare diagnostic accuracy of a caries detection program with that of human observers. A total of 190 extracted teeth were radiographed with two Trophy RVG (RadioVisioGraphy) digital sensor systems. Four observers scored the approximal surfaces in all images on a disease severity scale. Each observer thereafter used the Logicon Caries Detector (LCD) program to analyse the surfaces in the digital images and recorded their outcome. To determine the true absence or presence of caries, histological validation was used. Sensitivities, specificities, positive and negative predictive values were calculated and differences between the diagnostic methods tested. Specificities for the outcome with the LCD were significantly lower for three observers than when they themselves assessed the RVG images and, correspondingly, the positive predictive values were lower for the LCD outcome for three of the observers. Sensitivity was also lower for two observers on the diagnostic threshold caries in dentine. It was concluded that the automated caries detection program is less accurate than human observers in detecting approximal caries lesions. PMID- 12120705 TI - Comparison of factors potentially related to the occurrence of dental erosion in high- and low-erosion groups. AB - Soft drink intake, method of drinking, pH variations, plaque topography, and various salivary, microbial and clinical factors were compared in Saudi men with high (n = 10, mean = 20.5 yr) and low (n = 9, mean = 20.3 yr) dental erosion. pH measurements were carried out with a microtouch electrode at six different intraoral locations after the subjects had consumed 330 ml of regular cola-type drink in their customary manner. The results showed that higher intake of cola type drinks was more common in the high- (253 l yr(-1)) than in the low-erosion group (140 l yr(-1)). High erosion was associated with a method of drinking whereby the drink was kept in the mouth for a longer period (71 s vs. 40 s). pH after drinking did not differ between the groups for any of the six measuring sites. Plaque accumulation on the palatal surfaces of maxillary anterior teeth and urea concentration in unstimulated saliva were lower in high-erosion subjects. Aside from these, there were no differences in salivary and microbial factors between the groups. First molar cuppings, buccal cervical defects, and mouth breathing were more common in the high- than in the low-erosion group. In summary, consumption of cola-type drink, method of drinking, amount of palatal plaque on anterior teeth, and salivary urea concentration are factors associated with dental erosion. PMID- 12120706 TI - Actinobacillus actinomycetemcomitans proportion of subgingival bacterial flora in relation to its clonal type. AB - We investigated whether certain Actinobacillus actinomycetemcomitans clones occur in elevated proportions in subgingival flora, and if the proportions relate to other bacteria in the samples. A total of 121 A. actinomycetemcomitans strains from 121 patients with periodontitis were serotyped and 60 strains were also genotyped. The 121 strains were divided into three groups and the 60 strains into two groups according proportion of A. actinomycetemcomitans. The samples from the 60 patients with genotyped strains were cultured for five other species. Among the 121 strains, serotype b occurred significantly more frequently in the high- (n = 14, proportions > 5%, mean = 18.09, SD = 20.07%) than low- (n = 49, proportions < or = 0.1%), mean = 0.04, SD = 0.03%) or intermediate-proportion groups (n = 58, proportions > 0.5%, mean = 1.31, SD = 1.24%). Genotype 3 occurred significantly more frequently in samples with low A. actinomycetemcomitans proportions (n = 28, < or = 0.1%, mean = 0.04, SD = 0.03%) than in those with high proportions (n = 32, > 0.1%, mean = 5.70, SD = 14.60%). No differences were seen in the detection frequencies or proportions of the five bacterial species between the samples with low or high A. actinomycetemcomitans proportions. The results indicate that certain clonotypes of A. actinomycetemcomitans may preferentially occur as low proportions, suggesting their controlled growth. Conversely, some serotype b clones may have a competitive advantage in subgingival flora. PMID- 12120707 TI - Selection of dairy bacterial strains as probiotics for oral health. AB - The aim of the present study was to select bacterial strains with potential properties as oral probiotics, namely for the prevention of dental caries. We examined 23 dairy microorganisms, out of which we identified two Streptococcus thermophilus and two Lactcoccus lactis strains that were able to adhere to saliva coated hydroxyapatite beads to the same extent as Streptococcus sobrinus OMZ176. Two of them, Strep. thermophilus NCC1561 and Lactoc. lactis ssp. lactis NCC2211, were further successfully incorporated into a biofilm mimicking the dental plaque. Furthermore, they could grow in such a biofilm together with five strains of oral bacterial species, representative of supragingival plaque. In this system, Lactoc. lactis NCC2211 was able to modulate the growth of the oral bacteria, and in particular to diminish the colonization of Streptococcus oralis OMZ607, Veillonella dispar OMZ493, Actinomyces naeslundii OMZ745 and of the cariogenic Strep. sobrinus OMZ176. These findings encourage further research with selected non-pathogenic dairy bacterial strains with the aim to decrease the cariogenic potential of dental plaque. PMID- 12120708 TI - Influence of secondary colonizers and human plasma on the adherence of Porphyromonas gingivalis in vitro. AB - The influence of secondary colonizers (Fusobacterium nucleatum and Actinomyces naeslundii) and the effect of human plasma on the adherence of Porphyromonas gingivalis were investigated. Hydroxyapatite (HAP) discs coated with Streptococcus sanguis were immersed in a 3H-labeled bacterial cell suspension of F. nucleatum or A. naeslundii and then in a 14C-labeled P. gingivalis cell suspension. Bacterial cells on the discs were pyrolysed to quantify the radioisotopes released. The cell numbers of secondary colonizers on the discs increased with immersion time and this, in turn, resulted in significantly elevated adherence of P. gingivalis. These two secondary colonizers had very similar positive effects on the adherence of P. gingivalis. Human plasma significantly inhibited the adherence of P. gingivalis and secondary colonizers to S. sanguis-coated HAP discs. Adherence of P. gingivalis and A. naeslundii was strongly inhibited by plasma, while that of F. nucleatum was affected the least. Treatment with plasma, after immersion of streptococcal-coated discs in individual cell suspension of secondary colonizers, also reduced subsequent adherence of P. gingivalis. The rate of decrease was much smaller in F. nucleatum. These results indicate that both F. nucleatum and A. naeslundii enhance the adherence of P. gingivalis, and that the former may play a more important role in the establishment of P. gingivalis in dental plaque where plasma-derived components are present. PMID- 12120709 TI - Secretion from submucosal salivary glands of the ferret in response to a cholinesterase inhibitor applied onto the oral mucosa. AB - Parasympathomimetics or cholinesterase inhibitors taken orally may relieve dry mouth symptoms, but this route of administration is often associated with adverse systemic reactions. In the present study, an animal model was worked out aimed at stimulating the submucosal glands and avoiding systemic effects. In the anesthetized ferret, saliva from the parotid, sublingual and submandibular glands was prevented from reaching the mouth. Vehicle or physostigmine was applied topically for 10 min on the buccal and labial mucosa on one side, while the other side served as a control. Fluid from each side was collected every 5 min Mean basal secretion was 0.17 mg 5 min(-1). The response to physostigmine 0.1% did not exceed that of the vehicle. At higher concentrations the responses lasted 80-120 min. Peak secretion was nine (0.25% physostigmine), 13 (0.5% physostigmine) and 40 (1% physostigmine) times higher than baseline. At 1% physostigmine, the secretion from the control side was elevated, and a small flow from the duct cannulated parotid gland occurred, indicating systemic effects. Arterial blood pressure was well maintained. Additional observations on the duct-cannulated zygomatic gland showed that secretion from this gland increased already within the 10-min period of application of physostigmine to the overlying mucosa. The distance between the mucosa and the zygomatic gland was only about 1 mm. Physostigmine-induced secretion was abolished by atropine. Local gland stimulation may be an attractive alternative for the treatment of dry mouth. PMID- 12120710 TI - Immunohistochemical study on pulpal response in rat molars after cavity preparation by Er:YAG laser. AB - While Er:YAG laser systems are in extensive use for caries removal and cavity preparation, the effects of such treatment on pulp tissue remain unclear. This study evaluates these systems using immunohistochemical methods and compares the results with information gained from treatment using conventional burs. Cervical cavities were prepared in the upper first molars of rats, using either an Er:YAG laser or a conventional tungsten-carbide bur. At intervals of 5 min, 6 h, 12 h, 1 d, 3 d and 7 d after cavity preparation, the teeth were processed for immunohistochemical analyses of tissue non-specific alkaline phosphatase, OX6 positive major histocompatibility complex class II antigen-expressing cells and PGP 9.5-immunoreactive nerve fibers. DNA fragmentation was detected by the terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) method. Tissue non-specific alkaline phosphatase was observed mainly in the subodontoblastic layer under the cavity lesion, from 5 min, in both groups. The immunoreactivity was more pronounced in the laser group, but by 7 d no significant differences were recognizable. At 12 h, TUNEL-positive cells were detected around the odontoblastic layer in both groups. From 3 d to 7 d, a limited number of positive cells were still visible in the group that underwent standard treatment. Clear similarities in the distribution patterns of OX6 immunopositive cells and PGP 9.5-immunoreactive nerve fibers were also noted. From 12 h to 1 d, OX6-positive cells accumulated along the pulp-dentin border, extending their processes into the dentinal tubules. Numerous bead-like PGP 9.5 immunoreactive nerve fibers were observed under the odontoblastic layer at 7 d. These results demonstrated that there was no appreciable difference in the manner in which pulp tissue responded to treatment with either Er:YAG laser or a conventional drill. This would seem to indicate the usefulness of the Er:YAG laser system in the removal of caries and cavity preparation. PMID- 12120711 TI - Sequential expressions of MMP-1, TIMP-1, IL-6, and COX-2 genes in induced periapical lesions in rats. AB - To elucidate the pathogenesis of periapical lesion-associated bone resorption, a disease model of Wistar rat molar was employed. After lesion induction, the mRNAs encoding for matrix metalloproteinase-1 (MMP-1), tissue inhibitor of metalloproteinase-1 (TIMP-1), interleukin-6 (IL-6), and cyclooxygenase-2 (COX-2) in the developing lesions were detected by in situ hybridization at day 5, 10, 15 and 20, respectively. At day 5, MMP-1, IL-6 and COX-2 mRNAs appeared predominantly in macrophages. During day 15 to day 20, increased expressions of these mediators were also found in osteoblasts but to a lesser extent compared with those in macrophages. MMP-1 mRNA was also detected in osteoclasts. In contrast, expression of the TIMP-1 gene was noted primarily in osteoblasts and was less pronounced compared with that of MMP-1. The mediator-expressing cells aggregated in the vicinity of bone resorption areas and their numbers increased with time. These data suggest that macrophages and osteoblasts are involved in the development of periapical lesions, and that they promote bone resorption by producing MMP-1, IL-6 and COX-2. In addition, administration of a specific COX-2 inhibitor, meloxicam, reduced the extent of periapical bone resorption by 43% and simultaneously diminished the numbers of cells synthesizing MMP-1 and IL-6 mRNAs. These results further elucidate the significance of COX-2 in disease progression of periapical lesions as it modulates indirectly the production of MMP-1 and IL 6. PMID- 12120712 TI - Gene transfer mediated by different viral vectors following direct cannulation of mouse submandibular salivary glands. AB - The salivary gland has been suggested as an accessible organ for gene transfer to express recombinant proteins locally in the saliva, as well as for secretion to the blood circulation. The aim of this study was to evaluate the efficiency of gene transfer to salivary glands using different viral vectors: adenovirus, vaccinia, herpes simplex type 1 (HSV), and two retroviral vectors (murine leukemia virus (MuLV) and lentivirus). We show, by in situ staining and beta galactosidase reporter activity assay, that the adenoviral and vaccinia vectors were able to deliver the reporter gene efficiently to acinar and duct cells. The HSV vector was less efficient and infected only the acinar cells. The lentiviral vector infected acinar and duct cells, but at a relatively low efficiency. The MuLV vector did not infect the salivary gland unless cell proliferation was induced. Host immune responses to viral infection, inflammation, apoptosis and lymphocyte infiltration, in the transduced glands, were assessed. The DNA viral vectors induced local lymphocyte infiltration and apoptosis. In contrast, the retroviral vectors did not induce an immune response. Our results describe the outcome of salivary gland infection with each of the five different viral vectors and indicate their advantages and limitations for transferring genes to the salivary glands. PMID- 12120713 TI - On the origin of intrinsic matrix of acellular extrinsic fiber cementum: studies on growing cementum pearls of normal and bisphosphonate-affected guinea pig molars. AB - Cementum pearls (CPs) belong to a type of acellular extrinsic fiber cementum (AEFC) that form on the maturing enamel of guinea pig molars. This study aimed to elucidate the forming process of intrinsic matrix of AEFC using the CPs of normal and bisphosphonate-affected guinea pig molars as experimental models. A group of guinea pigs were subjected to continuous administration of 1-hydroxyethylidene 1,1-bisphosphonate (HEBP) for 2 wk to inhibit mineralization of growing CPs. Fenestration of the enamel organ and migration of periodontal cells on to the exposed surface of maturing enamel appeared to be unaffected by HEBP, whereas de novo formation as well as growth of pre-existing CPs did not proceed under the same conditions. Immunoreactions for osteopontin were located exclusively on the mineralized matrix of preformed CPs, implying the absence of additional deposition or accumulation of putative intrinsic cementum matrix on the affected CPs, where the propagation of mineral phase had been arrested. In both normal and HEBP-treated groups, distinct enzymatic reactions for alkaline phosphatase appeared on the cells of the periodontal ligament associated closely with the sites of CP formation, and along the mineralization front of CPs. These observations suggest that the mineralization process per se plays a central role in the deposition of AEFC matrix and that alkaline phosphatase of periodontal cells penetrating through the enamel organ to the maturing enamel surface plays a key role in the mineralization process of CPs. PMID- 12120714 TI - Modified Class II open sandwich restorations: evaluation of interfacial adaptation and influence of different restorative techniques. AB - The sandwich technique with resin-modified glass ionomer cement (RMGIC) has been proposed to relieve the contraction stresses of direct resin composite (RC) restorations. The aim of this study was to evaluate the interfacial adaptation to enamel and dentin of modified Class II open RMGIC/RC sandwich restorations and the influence of different light curing techniques and matrix bands. Forty box shaped Class II fillings were placed in vivo in premolars scheduled for extraction after one month. In groups I and II, a metal matrix was used; RC was inserted with horizontal (group I) and diagonal (group II) increments and cured with indirect/direct light. Group III was performed as group II, but a transparent matrix was used. Group IV was as group II, but with a separating liner between RMGIC and RC. Group V was a closed sandwich restoration. Interfacial quality was studied using SEM replica technique. Gap-free interfacial adaptation to enamel was observed for RMGIC in 70%, for RC in 70% and to dentin for RMGIC in 81%, for RC in 56%. No significant differences were seen between the experimental groups. At the cervical margins, RMGIC showed significantly better adaptation to enamel than RC, 74% and 42%, respectively. In conclusion, the investigated restorations showed a high percentage of gap-free interfacial adaptation in vivo. Interfacial adaptation to dentin and to cervical enamel was significantly better for RMGIC than for RC. PMID- 12120715 TI - Comparison of auditory steady-state response and auditory brainstem response thresholds in children. AB - Recently, auditory steady-state responses (ASSRs) have been proposed as an alternative to the auditory brainstem response (ABR) for threshold estimation. The goal of this study was to investigate the degree to which ASSR thresholds correlate with ABR thresholds for a group of sedated children with a range of hearing losses. Thirty-two children from the University of Iowa Hospitals and Clinics ranging in age from 2 months to 3 years and presenting with a range of ABR thresholds participated. Strong correlations were found between the 2000-Hz ASSR thresholds and click ABR thresholds (r = .96), the average of the 2000- and 4000-Hz ASSR thresholds and click ABR thresholds (r = .97), and the 500-Hz ASSR and 500-Hz toneburst ABR thresholds (r = .86). Additionally, it was possible to measure ASSR thresholds for several children with hearing loss that was great enough to result in no ABR at the limits of the equipment. The results of this study indicate that the ASSR may provide a reasonable alternative to the ABR for estimating audiometric thresholds in very young children. PMID- 12120716 TI - Prediction of hearing threshold in infants using auditory steady-state evoked potentials. AB - This retrospective study examines the relationship between auditory steady-state evoked potential (ASSEP) thresholds determined in infancy and subsequently obtained behavioral hearing levels in children with normal hearing or varying degrees of sensorineural hearing loss. Overall, the results from 211 subjects showed that the two test techniques were highly correlated, with Pearson r values exceeding .95 at each of the audiometric test frequencies between 500 and 4000 Hz. Analysis of the findings for babies with significant hearing loss (moderate to profound levels) showed similar threshold relationships to those obtained in previous studies involving adults and older children. The results for infants with normal or near-normal hearing did, however, differ from those reported for older subjects, with behavioral thresholds typically 10 to 15 dB better than would have been predicted from their ASSEP levels. PMID- 12120717 TI - Advantages and caveats when recording steady-state responses to multiple simultaneous stimuli. AB - This article considers the efficiency of evoked potential audiometry using steady state responses evoked by multiple simultaneous stimuli with carrier frequencies at 500, 1000, 2000, and 4000 Hz. The general principles of signal-to-noise enhancement through averaging provide a basis for determining the time required to estimate thresholds. The advantage of the multiple-stimulus technique over a single-stimulus approach is less than the ratio of the number of stimuli presented. When testing two ears simultaneously, the advantage is typically that the multiple-stimulus technique is two to three times faster. One factor that increases the time of the multiple-response recording is the relatively small size of responses at 500 and 4000 Hz. Increasing the intensities of the 500- and 4000-Hz stimuli by 10 or 20 dB can enhance their responses without significantly changing the other responses. Using multiple simultaneous stimuli causes small changes in the responses compared with when the responses are evoked by single stimuli. The clearest of these interactions is the attenuation of the responses to low-frequency stimuli in the presence of higher-frequency stimuli. Although these interactions are interesting physiologically, their small size means that they do not lessen the advantages of the multiple-stimulus approach. PMID- 12120718 TI - The auditory steady-state response: full-term and premature neonates. AB - Two studies were aimed at developing the auditory steady-state response (ASSR) for universal newborn hearing screening. First, neonates who had passed auditory brainstem response, transient evoked otoacoustic emission, and distortion-product otoacoustic emission tests were also tested with ASSRs using modulated tones that varied in frequency and level. Pass rates were highest (> 90%) for amplitude modulated tones presented at levels > or = 69 dB SPL. The effect of modulation frequency on ASSR for 500- and 2000-Hz tones was evaluated in full-term and premature infants in the second study. Full-term infants had higher pass rates for 2000-Hz tones amplitude modulated at 74 to 106 Hz compared with pass rates for a 500-Hz tone modulated at 58 to 90 Hz. Premature infants had lower pass rates than full-term infants for both carrier frequencies. Systematic investigation of ASSR threshold and the effect of modulation frequency in neonates is needed to adapt the technique for screening. PMID- 12120719 TI - The auditory steady-state response: clinical observations and applications in infants and children. AB - Two studies illustrate the use of the auditory steady-state response (ASSR) in the pediatric clinical audiology setting. A protocol for estimating bone conduction thresholds from ASSR was developed. Bone-conducted narrow-band noise was used to mask the ASSR for a 1.0-kHz modulated tone. The amount of bone conducted noise needed to mask the ASSR may distinguish between infants and children with conductive hearing losses and those with sensory losses. The amount of bone-conducted noise may also be used to estimate bone-conduction thresholds; however, the accuracy of this technique needs verification with behavioral methods to determine thresholds for bone-conducted pure tones in infants. When ASSR tests are used as part of the diagnostic evaluation for infants and children at risk for hearing loss, the results yield information about the audiometric contour and residual hearing, which aid in treatment and habilitation decisions. PMID- 12120720 TI - The new contract: renaissance or requiem for general practice? PMID- 12120721 TI - Chronic obstructive pulmonary disease and primary care. PMID- 12120722 TI - X-rays for back pain? PMID- 12120723 TI - Endoscopy in primary care--a survey of current practice. AB - BACKGROUND: Long waiting lists in district general hospitals and savings from fundholding led to the setting up of a number of endoscopy units in primary care. Concerns have been expressed over safety, supervision and cost effectiveness. Increasingly, general practitioners (GPs) are being encouraged to become specialists and offer intermediate care. Endoscopy is frequently cited as an example of intermediate care that could be offered by primary care specialists. This is the first survey of such a service. AIM: To examine whether endoscopy in primary care can be considered to be a safe procedure. DESIGN OF STUDY: A questionnaire-based survey. SETTING: Twenty-eight general practice units performing endoscopy in primary care. METHOD: Units performing endoscopy in primary care were identified using the Primary Care Society of Gastroenterology (PCSG) database and following an appeal in the GP press. A postal questionnaire was sent to each unit covering its history, throughput, and case-mix, experience of endoscopists, supervision, audit and CME, equipment, waiting times and complication rates. RESULTS: Of the 28 units identified, 27 (96%) replied to the questionnaire, 13 units provided both upper and lower bowel examination, six oesophago-gastro-duodenoscopy (OGD) only, and eight lower bowel only. Units had been openfor an average of five years (range = 2 to 18 years), and 41 doctors and 68 nurse assistants provided the service. The average experience of endoscopists was 16 years (range = 6 to 25 years), and 36,455 procedures had been performed by the time of the survey (24,195 OGD and 12,260 lower bowel examinations). Ninety six per cent of the units undertook audit. Urgent waiting times were 1.2 weeks and routine 3.4 weeks (range = 1.0 to 6.0). The annual throughput of 22 units in the past year was 8,478 procedures (4506 OGD, 3,972 lower bowel examinations). Out of 24,195 OGDs there were three reported complications (one perforation of pharyngeal pouch, treated conservatively, one chest pain after over-insufflation, and one slow recovery after intravenous sedation); there was no mortality. Out of 12,260 lower bowel procedures there was one perforated caecal carcinoma after flexible sigmoidoscopy (died), three perforations at colonoscopy and seven other minor complications. CONCLUSIONS: Endoscopy in primary care appears to be a safe procedure. This good safety record is probably attributable to careful case selection and minimal use of intravenous sedation. PMID- 12120724 TI - Feasibility and effectiveness of a pulmonary rehabilitation programme in a community hospital setting. AB - BACKGROUND: Pulmonary rehabilitation programmes run in secondary care have proved to be one of the most effective interventions for patients with chronic obstructive pulmonary disease (COPD). AIM: To assess whether a pulmonary rehabilitation programme, similar to that run in secondary care, could be established in a primary care-run community hospital and whether it could achieve similar benefits in patents with moderately severe COPD. DESIGN OF STUDY: Uncontrolled prospective intervention study SETTING: A primary care-run community hospital. METHOD: Thirty-four patients with COPD aged between 5 and 80 years of age (mean = 70years) with a forced expiratory volume (FEV1) of 30 to 50% (mean = 40%) predicted were enrolled in a programme established in the activities room at Honiton Community Hospital. Patients were assessed at the start, on completion of the programme, and six months after completion, using spirometry, shuttle-walking distance, and short form-36 (SF-36) and chronic respiratory questionnaire (CRQ) scores. RESULTS: All but one patient completed the programme. There were significant improvements in the walking distance (by a mean of 100 m), in the SF 36, and in all domains of the CRQ. There was no significant change in the FEV1 or forced vital capacity. CONCLUSION: Pulmonary rehabilitation programmes can be run in community hospitals. They appear to be as effective as those run in secondary care and patients may find them easier to access. PMID- 12120725 TI - A survey of access to medical services in nursing and residential homes in England. AB - BACKGROUND: Residential and nursing homes make major demands on NHS services. AIM: To investigate patterns of access to medical services for residents in homes for older people. DESIGN OF STUDY: Telephone survey. SETTING: All nursing and dual registered homes and one in four residential homes located in a stratified random sample of 72 English primary care group/trust (PCG/T) areas. METHOD: A structured questionnaire investigating home characteristics, numbers of general practitioners (GPs) or practices per home, homes' policies for registering new residents with GPs, existence of payments to GPs, GP services provided to homes, and access to specialist medical care. RESULTS: There were wide variations in the numbers of GPs providing services to individual homes; this was not entirely dependent on home size. Eight percent of homes paid local GPs for their services to residents; these were more likely to be nursing homes (33%) than residential homes (odds ratio [OR] = 10.82, [95% CI = 4.48 to 26.13], P<0.001) and larger homes (OR for a ten-bed increase = 1.51 [95% CI = 1.28 to 1.79], P<0.001). Larger homes were more likely to encourage residents to register with a 'home' GP (OR for a ten-bed increase = 1.16 [95% CI = 1.04 to 1.31], P = 0.009). Homes paying local GPs were more likely to receive one or more additional services, over and above GPs' core contractual obligations. Few homes had direct access to specialist clinicians. CONCLUSION: Extensive variations in homes' policies and local GP services raise serious questions about patient choice, levels of GP services and, above all, about equity between residents within homes, between homes and between those in homes and in the community. PMID- 12120726 TI - Selecting persistent glue ear for referral in general practice: a risk factor approach. AB - BACKGROUND: Glue ear (otitis media with effusion) is the most common reason for surgical intervention in children. AIM: To determine the yield and predictive value of a set of risk factors that predict persistence of glue ear over the interval from general practice referral to ear, nose and throat (ENT) consultation to ensure the appropriateness of referrals. DESIGN OF STUDY: Nested case control study SETTING: Sixteen ENT departments in the UK. METHOD: With the aid of audiometry and tympanometry, diagnostic information was collected on 548 children from 16 ENT departments after referral by their general practitioner (GP), as a lead-in to a clinical trial, the Trial of Alternative Regimens in Glue Ear Treatment (TARGET). Using cases and controls, children were classified as either having or not having persistent glue ear. Parental reports on an extensive list of risk factors were also collected. RESULTS: After adjustment for time waiting to be seen from GP referral and age at referral, four main significant factors emerged for persistence of glue ear. These were: referral between July and December (OR [odds ratio] = 1. 73, 95% CI = 1.15 to 2.6); having a mother who smokes ten or more cigarettes per day (OR = 1.7, 95% CI = 1.1 to 2.8); multiple upper airway symptoms (OR = 2.2, 95% CI = 1.5 to 3.2; and siblings with a history of glue ear (OR = 1.6 for one sibling versus none). CONCLUSION: For a child who is referred between July and December; who has two or more upper airway symptoms, who has a sibling who has had glue ear, and who has a mother who smokes ten or more cigarettes per day, the odds of having persistent glue ear are over ten times that of a child without adverse values on these factors. PMID- 12120727 TI - Questionnaire survey of users of NHS walk-in centres: observational study. AB - BACKGROUND: NHS walk-in centres have recently been established throughout England to improve access to primary health care. AIM: To determine the characteristics and experiences of people consulting NHS walk-in centres compared with general practice. DESIGN OF STUDY: Observational study using questionnaires. SETTING: Thirty-eight walk-in centres and 34 neighbouring general practices. METHOD: People attending randomly selected survey sessions at walk-in centres or neighbouring general practices on a 'same-day' basis were given a self administered questionnaire. This collected data about socio-demographic characteristics, reasons for consulting, attitudes to continuity, satisfaction, enablement, referrals, and intentions. RESULTS: Walk-in centre visitors were more likely to be owner-occupiers (55% versus 49%; P<0.001), to have further education (25% versus 19%; P = 0.006), and to be white (88% versus 84%; P< 0.001) than general practice visitors. Main reasons for attending a walk-in centre were speed of access and convenience. Walk-in centre visitors were more likely to attend on the first day of illness (28% versus 10%; P<0.001), less likely to expect a prescription (38% versus 70%, P<0.001), and placed less importance on continuity of care (adjusted odds ratio = 0.58; 95% CI = 0.50 to 0.68) than general practice visitors. People were more satisfied with walk-in centres (adjusted mean difference = 6.6%; 95% CI = 5.0% to 8.2%). Enablement scores were slightly higher in general practice (adjusted mean difference = 0.40; 95 % CI = 0.11 to 0. 6). Following the consultation 13% of walk-in centre visitors were referred to general practice, but 32% intended to make an appointment. CONCLUSION: NHS walk in centres improve access to care, but not necessarily for those people with greatest health needs. People predominantly attend with problems of recent onset as an alternative to existing health providers, and are very satisfied with the care received. These benefits need to be considered in relation to the cost, and in comparison with other ways of improving access to health care. PMID- 12120728 TI - Defining frequent attendance: evidence for routine age and sex correction in studies from primary care settings. AB - Primary care patients who attend significantly more frequently than the norm present a clinical challenge and implications for resource usage. However, studies are methodologically diverse in their definitions of frequent attendance, and do not always standardise for age and sex. This study shows that studies that do not correct for age and sex variations in consultation will miss a significant proportion of patients whose behaviour differs widely from their peers. It suggests a simple correction that could be utilised in primary care studies without requiring sophisticated statistical analyses. PMID- 12120729 TI - Membership of the Royal College of General Practitioners and recognition of depression in primary care. AB - To determine the effect of membership of the Royal College of General Practitioners (RCGP) on recognition of depression, odds ratios, Lowess-smoothed plots, and regression models were produced for recognition rates in a representative sample of 20,818 patient contacts for members and non-members of the RCGP Membership of the RCGP did not appear to convey greater ability to discriminate between depressed and non-depressed patients, but members were more likely to recognise depression in both depressed and non-depressed individuals. PMID- 12120730 TI - A pilot study of pulmonary rehabilitation in primary care. AB - Pulmonary rehabilitation is an effective intervention for patients with chronic obstructive pulmonary disease (COPD). It is usually available only through selected hospitals. A pilot study was undertaken to see if pulmonary rehabilitation performed by the primary health care team in one practice was feasible. Fourteen patients were recruited; 13 completed the programme and one year of follow-up. The programme was well received by patients and staff. There were not enough suitable patients among a practice list of 10,500 to justify the running of this programme for a single practice; one primary care group would suffice PMID- 12120731 TI - Headache in primary care: how important is diagnosis to management? AB - Headache is a common presentation in primary care. The classification of headache was overhauled by the International Headache Society (IHS) in 1988, and the past decade has seen rapid growth in the understanding of headache disorders. The IHS places particular importance on precise headache diagnosis. This paper discusses the relevance of such an approach to primary care. A review of the literature revealed a dearth of evidence regarding headache management in primary care settings. The evidence from other settings is considered and gaps in the literature highlighted. PMID- 12120732 TI - Effects of physical activity in mild to moderate COPD: a systematic review. AB - Pulmonary rehabilitation has become an evidence-based treatment for patients with severe chronic obstructive pulmonary disease (COPD). However, large numbers of patients who suffer from mild to moderate COPD receive treatment from their general practitioners (GPs). To encourage compliance, advice given to patients in general practice should be clear, practical, and acceptable. This is particularly true of the advice that is given by GPs to improve their patients' physical condition by walking, cycling or swimming, as recommended by the Dutch College of General Practitioners in their guideline for the treatment of COPD. We performed a literature search on the effects of physical activity in patients with mild to moderate COPD on exercise tolerance, dyspnoea and quality of life (QOL). We also looked at the numbers of hospitalisation days and exacerbations, expressed as oral prednisolone courses. The literature search included Medline (1983 to 1999), EMBASE (1984 to 2000), and the Cochrane Library (2000). All hits were screened for subject and language and abstracts were selected on the basis of a protocol that included disease severity, hypothesis, outcome parameters, and control group. Review articles on physical exercise and COPD were examined and reference lists of selected articles were screened for relevant studies The broad literature search generated 4,968 articles and, after exclusion according to title and abstract, 35 original studies and 27 review articles were analysed. Of these, five original studies fitted the criteria and none of the review articles was selected. A positive influence of physical activity on exercise tolerance in mild to moderate COPD was reported in four out of five studies. There was no clear effect on dyspnoea or QOL, probably because of the low numbers of subjects. No studies that addressed the number of hospitalisation days or prednisolone courses as outcomes were included. Physical exercise training (usually as part of a package of rehabilitation) can improve the fitness of patients with mild or moderate COPD, but it has not been shown to benefit QOL or dyspnoea significantly, or indeed long-term disease progression. PMID- 12120733 TI - A case of mural dyslexia. PMID- 12120734 TI - Influenza ear? PMID- 12120735 TI - Cataract: a pre-referral protocol. PMID- 12120737 TI - 'Homelessness and health' conference. PMID- 12120736 TI - Abdominal aortic aneurysm. PMID- 12120738 TI - Response shift, responsiveness or recall bias? PMID- 12120739 TI - GP telephone consultations. PMID- 12120740 TI - Chronic fatigue syndrome. PMID- 12120741 TI - Prescribing costs and patterns: authors' response. PMID- 12120742 TI - Commentary on the EBOR trial report. PMID- 12120743 TI - Screening for atrial fibrillation. PMID- 12120744 TI - Intuition, creativity, dialogue, tacit knowledge and ... evidence? PMID- 12120751 TI - Pharmacological aspects of erectile dysfunction. AB - Erectile dysfunction (ED) is a common problem with a prevalence of approximately 50% in men aged 40 to 70. There are several etiologies for ED including vasculogenic, neurogenic, hormonal and/or psychogenic factors; one-fourth of ED cases can be drug-related. Penile erection involves a complex interaction between the CNS and local factors. It is a neurovascular event modulated by psychological and hormonal factors. Pharmacologically, neural modulation and endocrine status are very important to attaining penile erection. There have been several significant advances for the pharmacologic treatment of ED. Treatments include agents that are not only orally effective, but possess either local or central acting mechanisms of action. Apomorphine, a centrally-acting agent, is effective in the treatment of ED. Sildenafil, another orally effective agent, acts by inhibiting cyclic GMP-specific phosphodiesterase Type V. Testosterone can be effective transdermally. Non-orally active agents include alprostadil and papaverine. Phentolamine and trazodone are effective in selected cases. Some agents can interact with other medications. Several pharmacological agents, some with central-acting mechanisms and some with Iocally-acting vascular effects, are therapeutically useful in the treatment of ED. PMID- 12120752 TI - Studies for the emetic mechanisms of ipecac syrup (TJN-119) and its active components in ferrets: involvement of 5-hydroxytryptamine receptors. AB - Ipecac syrup, prepared from a galentical ipecac, contains the nauseant alkaloids cephaeline and emetine. The involvement of receptors and serotonin- and dopamine metabolizing enzymes in the emesis induced by ipecac syrup and these components was investigated. 1) In ferrets, the selective 5-HT3-receptor antagonist ondansetron (0.5 mg/kg, p.o.) prevented each emesis induced by TJN-119 (0.5 mL/kg, p.o.), cephaeline (0.5 mg/kg, p.o.) and emetine (5.0 mg/kg, p.o.), but the intraperitoneal administration of the selective dopamine D2-receptor antagonist sulpiride failed to significantly suppress the TJN-119, cephaeline and emetine induced emesis at a dose of 0.1 mg/kg that blocked apomorphine-induced emesis. 2) In the receptor binding assays, cephaeline and emetine had a distinct affinity to 5-HT4 receptor, but no or weak affinity to 5-HT1A, 5-HT3, nicotine, M3, beta1, NK1, and D2 receptors. 3) Cephaeline and emetine did not affect activities of metabolic enzymes of 5-HT and dopamine (MAO-A, MAO-B, tryptophan 5-hydroxylase and tyrosine hydroxylase) in vitro. These results suggest that 5-HT3 receptor plays an important role in the emetic action of TJN-119, cephaeline and emetine, and the 5-HT4 receptor may be involved in their mechanisms. PMID- 12120753 TI - Influence of angiotensin II type 1-receptor antagonist CV11974 on infarct size and adjacent regional function after ischemia-reperfusion in dogs. AB - The presence of nonischemic regional dysfunction at the adjacent region of the ischemic myocardium was demonstrated in clinical studies. Recent studies demonstrated an angiotensin II type 1 (AT1)-receptor antagonist reduced myocardial ischemia-reperfusion injury. We investigated the role of the adjacent region after reperfusion by studying the effects of AT1-receptor antagonist on myocardial function and infarct size. We investigated 12 open-chest anesthetized dogs undergoing 90 min of left anterior descending coronary artery occlusion followed by 4 h of reperfusion. Six dogs injected with an AT1-receptor antagonist (CV11974) immediately after reperfusion were compared with 6 control dogs. Percent systolic shortening (%SS) was measured by two sets of the pair sonomicrometer crystals implanted to adjacent and remote nonischemic myocardium. After 4 h of reperfusion, infarct size was measured. There were no significant differences of the %SS at baseline between two regions. In both groups, %SS at adjacent region after reperfusion was significantly decreased as compared with remote region. There were no significant differences between the two groups. Infarct size, as a percentage of the area at risk, was smaller in the AT, group than in control group (25.49+/-7.53% vs 68.58+/-26.88% P<0.01). AT1-receptor antagonist reduces infarct size. This effect is not related to the change of regional myocardial function at adjacent region after reperfusion. PMID- 12120754 TI - Stimulatory effects of centrally injected nitric oxide donors on gastric acid secretion in anesthetized rats. AB - The effects of centrally injected nitric oxide (NO) donors on gastric acid secretion were investigated in continuously perfused stomach of anesthetized rats. The lateral cerebroventricular (LV) injection of NOC5 (30 - 100 microg) and NOC12 (10 - 100 microg) dose-dependently stimulated gastric acid secretion. The LV injection of NOC18 (30 microg) also stimulated gastric acid secretion. The other type of NO donor, sodium nitroprusside (3 - 30 microg, LV), also dose dependently stimulated gastric acid secretion. The effect of NOC5 at 100 microg was blocked by carboxy-PTIO, an NO scavenger, and by cervical vagotomy. Furthermore, NOC12 (30, 100 microg) dose-dependently stimulated gastric acid secretion in pylorus-ligated conscious rats. These results suggest that centrally injected NO donors stimulate gastric acid secretion in both conscious and anesthetized rats through vagus activation. PMID- 12120755 TI - Laxative and anti-diarrheal activity of polycarbophil in mice and rats. AB - We investigated the laxative and anti-diarrheal activity of polycarbophil, an insoluble hydrophilic polymer, in comparison with other agents used for treating functional bowel disorder (FBD). In naive rats, polycarbophil (500 mg/kg) increased fecal weight and water contents without producing diarrhea. Carboxymethylcellulose (CMC) did not produce evident changes in bowel movement. Picosulfate markedly produced diarrhea. Loperamide, trimebutine and granisetron decreased stool output dose-dependently. Constipation, indicated by decrease in fecal weight, was produced by loperamide and clonidine in rats. Polycarbophil (500 mg/kg) and CMC increased fecal weight without diarrhea. Conversely trimebutine further decreased fecal weight in constipated rats. Polycarbophil (500 mg/kg) suppressed diarrhea induced by castor oil, and at 250-500 mg/kg, it produced shaped stools in animals with stools loosened by prostaglandin E2, serotonin or carbachol in mice. Polycarbophil (500 mg/kg) also reduced stools in rats with stool output increased by wrap restraint stress (WRS). CMC had no effect in the diarrhea models, except for carbachol-induced diarrhea, and WRS induced evacuation. Loperamide, trimebutine and granisetron inhibited diarrhea production and WRS-induced evacuation, except for carbachol-induced diarrhea. The results show that polycarbophil prevents constipation and diarrhea without inducing diarrhea or constipation, which is different from the other agents. Hydrophilic polymers such as polycarbophil will be promising agents for the treatment of FBD. PMID- 12120756 TI - Antioxidative and hypolipidemic effects of barley leaf essence in a rabbit model of atherosclerosis. AB - The antioxidative and hypolipidemic effects of barley leaf essence (BL) were investigated in a rabbit model of atherosclerosis. Twenty-four New Zealand White male rabbits were assigned randomly into four dietary groups. The normal group was fed regular rabbit chow and the control group was fed a chow containing 0.5% cholesterol and 10% corn oil. The BL group and the probucol group were fed the same diet as the control group plus 1% (w/w) BL or 1% (w/w) probucol, respectively. The plasma levels of total cholesterol, triacylglycerol, lucigenin chemiluminescence (CL) and luminol-CL were increased in the control group compared to the normal group; and they were decreased in the BL group and the probucol group compared to the control group. The value of T50 of red blood cell hemolysis and the lag phase of low-density lipoprotein oxidation increased in the BL group and in the probucol group compared to the controls. Ninety percent of the intimal surface of the thoracic aorta was covered with atherosclerotic lesions in the control group, but only 60% of the surface was covered in the BL group. This 30% inhibition of hyperlipidemic atherosclerosis by BL was associated with a decrease in plasma lipids and an increase in antioxidative abilities (as measured by T50, lag phase and CL). These results suggest that the antioxidant and hypolipidemic effects of BL could be useful in the prevention of cardiovascular disease in which atherosclerosis is important. PMID- 12120757 TI - Production of nitric oxide, but not prostacyclin, is reduced in klotho mice. AB - A novel murine model of aging (kl/kl mice) has been developed by in vivo mutagenesis. We analyzed endothelial function in this strain. Ring preparations of the thoracic aorta were obtained from 6- to 9-week old wild-type (+/+) and heterozygous (kl/+) klotho mice. The aortas of kl/+ mice showed an exaggerated contractile response to norepinephrine and attenuated vasodilator responses to acetylcholine and lecithinized superoxide dismutase (SOD) compared to +/+ mice. The response to sodium nitroprusside was unaltered in kl/+ mice. The contraction in response to norepinephrine was augmented by treatment with N(G)-nitro-L arginine methyl ester (L-NAME, 10(-5) M) to a greater extent in +/+ mice than in kl/+ mice. Treatment with L-NAME abolished the vasodilator responses to both acetylcholine and lecithinized SOD. NO metabolites (NO2- and NO3-) and cGMP concentrations in the urine were significantly reduced in kl/+ mice compared to +/+ mice. However, the urinary excretion of 6-keto-prostaglandin F1alpha was unaltered. There was little immunostaining for NO synthase and vascular endothelial growth factor (VEGF) in the aorta of kl/+ mice. No immunostaining for NO synthase was noted in the aorta of kl/kl mice. The expression of the klotho gene product may have a role in the regulation of VEGF expression and is tightly linked to endothelial release of NO. PMID- 12120758 TI - Amlodipine inhibits pro-inflammatory cytokines and free radical production and inducible nitric oxide synthase expression in lipopolysaccharide/interferon-gamma stimulated cultured vascular smooth muscle cells. AB - Overproduction of nitric oxide (NO) from inducible nitric oxide synthase (iNOS) is importantly involved in the pathogenesis of endotoxemia and atherosclerosis. Calcium antagonists are commonly used as cardiovascular drugs and have a beneficial effect on prolonging survival in various models of endotoxin shock. The present study was to investigate the effect of a calcium antagonist amlodipine on nitrite, tumor necrosis factor-alpha (TNF-alpha) and interleukin 1beta (IL-1beta) formation and iNOS induction both in lipopolysaccharide (LPS) and interferon-gamma (IFN-gamma)-treated rat aortic smooth muscle cells (RASMC) and in a rat model of endotoxemia. Incubation with amlodipine (0.1 - 10 microM) for 24 h resulted in a significant and dose-dependent attenuation in medium nitrite, TNF-alpha and IL-1beta formation as well as iNOS protein expression in LPS/IFN-gamma-treated RASMC. In addition, amlodipine inhibited leucigenin-induced superoxide formation in RASMC. In the rat endotoxic model, the serum nitrite/nitrate, TNF-alpha and IL-1beta levels as well as iNOS protein expression of lungs were also suppressed by administration of amlodipine (50 microg/kg, i.v.). These results suggest that amlodipine may exert vascular beneficial effects by suppressing pro-inflammatory cytokines and free radical generation as well as iNOS induction in smooth muscle cells during activation of inflammatory mechanism. PMID- 12120759 TI - Effects of Kampo extracts on drug metabolism in rat liver microsomes: Rhei Rhizoma extract and Glycyrrhizae Radix extract inhibit drug oxidation. AB - In the present study, the effects on drug oxidations in rat liver microsomes in vitro using 126 Kampo extracts were investigated. Although the effects of inhibition on drug oxidations were dependent on the Kampo extracts and probe reactions studied, most of the Kampo extracts showed inhibitory effects on both N demethylations of aminopyrine and erythromycin in rat liver microsomes. Among the Kampo extracts studied herein, Daio-kanzo-to exhibited the most remarkable inhibitory effect on both reactions. The Rhei Rhizoma extract inhibited not only aminopyrine and erythromycin N-demethylations, but also phenacetin O deethylation, 7-ethoxycoumarin O-deethylation, ethanol oxidation and tolbutamide 4-hydroxylation in rat liver microsomes. The Glycyrrhizae Radix extract also showed a remarkable inhibitory effect on phenacetin O-deethylation as well as aminopyrine and erythromycin N-demethylations. In contrast, the Glycyrrhizae Radix extract virtually showed no effect on ethanol oxidation. PMID- 12120760 TI - Inhaled pinacidil, an ATP-sensitive K+ channel opener, and moguisteine have potent antitussive effects in guinea pigs. AB - We investigated whether inhaled pinacidil and moguisteine inhibit capsaicin induced coughs in guinea pigs. Inhaled pinacidil (15 - 60 microg/ml), an ATP sensitive K+ channel opener, and moguisteine (15 - 60 microg/ml) each dose dependently inhibited the number of capsaicin-induced coughs. The antitussive effects of pinacidil and moguisteine were significantly antagonized by pretreatment with glibenclamide (10 mg/kg, i.p.), an ATP-sensitive K+ channel blocker. However, pretreatment with naloxone methiodide (10 mg/kg, s.c.) had no significant effect on the antitussive effects of either pinacidil or moguisteine. On the other hand, inhaled dihydrocodeine (15 - 60 microg/ml) also dose dependently suppressed the number of capsaicin-induced coughs. The antitussive effect of inhaled dihydrocodeine was significantly antagonized by pretreatment with naloxone methiodide (10 mg/kg, s.c.), but not by glibenclamide (10 mg/kg, i.p.). These results indicate that inhaled pinacidil and moguisteine both attenuate capsaicin-induced coughs. Pinacidil and moguisteine may exert their antitussive effects through the activation of ATP-sensitive K+ channels in the tracheobronchial tract. Furthermore, it is possible that ATP-sensitive K+ channels may be involved in the antitussive effects of peripherally acting non narcotic antitussive drugs. PMID- 12120761 TI - Effect of anti-CD14 antibody on experimental periodontitis induced by Porphyromonas gingivalis lipopolysaccharide. AB - The lipopolysaccharide (LPS) released by Porphyromonas gingivalis, a Gram negative bacterium found in the periodontal pockets of patients with periodontitis, induces bone resorbing activity in vivo. We previously showed that a receptor for LPS on human gingival fibroblasts and gingival epithelial cells is CD14. In this study, we established a mouse model of experimental periodontitis by applying a P. gingivalis LPS solution to the buccal region of mice. P. gingivalis LPS-induced bone resorption and interleukin-6 production in the gingival tissues were significantly inhibited by pretreatment with anti-CD14 antibody for 5 weeks prior to LPS treatment. This result suggests that anti-CD14 antibody may be usable as a prototype for the development of drugs for the treatment of periodontal disease. PMID- 12120762 TI - Capsazepine inhibits thermal hyperalgesia but not nociception triggered by protease-activated receptor-2 in rats. AB - Protease-activated receptor-2 (PAR-2), expressed in sensory neurons, triggers thermal hyperalgesia, nociceptive behavior and spinal Fos expression in rats. In the present study, we examined if the nociceptive processing by PAR-2 is mediated by trans-activation of capsaicin receptors. The thermal hyperalgesia following an intraplantar (i.pl.) administration of the PAR-2-activating peptide SLIGRL-NH2 was completely abolished by the capsaicin receptor antagonist capsazepine. In contrast, neither the nociceptive behavior nor spinal Fos expression in response to i.pl. SLIGRL-NH2 were attenuated by capsazepine. Our data imply that trans activation of capsaicin receptors by PAR-2 might be involved in the PAR-2 triggered thermal hyperalgesia, but not nociception. PMID- 12120763 TI - Antagonistic interaction between BIIE 0246, a neuropeptide Y Y2-receptor antagonist, and omega-conotoxin GVIA, a Ca2+ channel antagonist, in presynaptic transmitter releases in dog splenic arteries. AB - Isolated dog splenic arteries were perfused with Krebs-Henseleit solution at 37 degrees C, using the cannula inserting method. Periarterial nerve electrical stimulation (10-V amplitude; 1-ms duration; 30-s trains of pulses; 1, 4 and 10 Hz) readily caused double peaked vasoconstrictions, i.e., 1st peaked response was mostly inhibited by alpha,beta-methylene ATP and the 2nd one, by prazosin. These responses were consistently inhibited by omega-conotoxin GVIA (omega-CTX), whereas they were facilitated by BIIE 0246, a neuropeptide Y (NPY) Y2-receptor antagonist. The omega-CTX-induced blocking effects of transmitter release were significantly antagonized by BIIE 0246. It is possible that the NPY Y2 receptor activity may partially be linked to presynaptic Ca2+ channels. PMID- 12120765 TI - Functional study on nitroxidergic nerve in isolated dog pulmonary arteries and veins. AB - In dog pulmonary arterial and venous strips without endothelium under treatment with prazosin, nicotine induced relaxation that was abolished by N(G)-nitro-L arginine, hexamethonium and methylene blue. L-Arginine antagonized the N(G)-nitro L-arginine action. Neurogenic relaxations tended to be more evident in the vein. Nitric oxide (NO)-induced relaxations were greater in the veins than in the arteries. Concentrations of NO to induce the same magnitude of relaxation as that to nicotine were higher in the arteries. In conclusion, dog pulmonary arteries and veins are innervated by nitroxidergic (nitrergic) nerves, and NO is released by nerve stimulation with nicotine in a larger amount in the artery than the vein. PMID- 12120764 TI - Cardiovascular effects of L-158,809, a new angiotensin type 1 receptor antagonist, assessed using the halothane-anesthetized in vivo canine model. AB - L-158,809 is a new angiotensin II type 1 receptor antagonist. We simultaneously assessed its antagonistic potency and cardiovascular effects with the halothane anesthetized in vivo canine model (n = 5). L-158,809 was intravenously infused over 10 min at escalating doses of 0.03, 0.3 and 3 mg/kg. Angiotensin II (0.1 microg/kg, i.v.)-induced vasopressor and negative inotropic responses were significantly suppressed from the low dose L-158,809. Meanwhile, L-158,809 did not affect any of the cardiovascular parameters except that QTc was slightly shortened after the high dose administration. These results support the previous in vitro knowledge that L-158,809 is a highly selective angiotensin II receptor antagonist. PMID- 12120766 TI - Organ culture as a useful method for studying the biology of blood vessels and other smooth muscle tissues. AB - The benefit of organ culture is to retain the original structural relationship between various cell species and their interactions and enable us to study the long-term effects of exogenous stimuli. Organ culture methods have been used especially in the studies of the proliferative vascular diseases, such as atherosclerosis and restenosis. We describe here that organ culture is a useful in vitro method to study the biology of vascular and other smooth muscle organs. PMID- 12120767 TI - Effect of pulp protection technique on the clinical performance of amalgam restorations: three-year results. AB - This study evaluated the influence of the pulp protection technique on clinical performance of amalgam restorations after three years, with particular reference to post-operative sensitivity and secondary caries. One hundred and twenty (120) Class II amalgam restorations (68 premolars, 52 molars; 78 MOD, 42 OD/MO) were placed in 30 participants (four restorations per participant). The restorations were divided into four groups according to the pulp protection technique used: copal varnish; 2% neutral sodium fluoride; adhesive resin and no pulp protection. The parameters evaluated were post-operative sensitivity, staining of the dental structure, tooth vitality, partial or total loss of the restoration and secondary caries. One hundred and eight (108) restorations were available for evaluation after three years. No partial or total loss of restorations had occurred; all teeth were vital, no tooth structure staining or secondary caries was detected in any of the restored teeth. Post-operative sensitivity was observed only in two restorations at baseline and at seven-days. The three-year clinical performance of teeth restored with a high copper dispersed phase amalgam was not affected by the choice of pulp protection technique. PMID- 12120768 TI - Clinical evaluation of a medium-filled flowable restorative material as a pit and fissure sealant. AB - This clinical study evaluated the retention rate and caries protection of a medium-filled (46% volume) flowable restorative material (CuRay-Match, OMNII Oral Pharmaceuticals, West Palm Beach, FL 33409, USA) compared to an unfilled sealant (Delton, Dentsply Caulk, Milford, DE 19963, USA). Using a half-mouth design, sealants were applied on randomly assigned caries-free first and/or second permanent molars of 32 children ranging in age from 6-11 years. A total of 118 teeth were etched, dried and sealed. Teeth were evaluated at one, six and 18 month intervals. After one month, 52 teeth sealed with unfilled sealant were intact compared with 46 sealed with a medium-filled resin, and after six months, 36 teeth sealed with an unfilled sealant were intact compared with 27 that were sealed with a medium-filled resin. After 18 months, 29 teeth were still fully sealed with an unfilled sealant, whereas 18 were sealed with a medium-filled resin. The difference between the two groups was not statistically significant. Regarding caries development, four teeth sealed with a medium-filled material and five teeth sealed with an unfilled sealant were decayed after 18 months. These results indicate that a medium-filled flowable restorative material did not perform better in retention rate and caries increment when compared to an unfilled conventional sealant. However, the effect of the additional techniques, such as the use of bonding agent and fissurotomy on retention rates should be evaluated in further studies. PMID- 12120770 TI - Effects of pre-soaked retraction cords on the microcirculation of the human gingival margin. AB - In the direct treatment of cervical lesions and to improve the impression-making procedure prior to fabricating indirect restorations, exposure of the gingival sulcus and control of hemorrhage or gingival fluid seepage are a prerequisite. For gingival displacement, cords impregnated with medicaments are widely used. In this investigation, the authors first studied in vitro the time course of fluid absorption by retraction cords immersed in test solutions. Thereafter, in a clinical trial, they examined the microcirculatory responses of the gingival margin after subgingival insertion and removal of retraction cords pre-soaked in solutions containing saline, AlCl3, Fe2(SO4)3 or epinephrine. Blood flow was recorded using laser Doppler technique. Blood perfusion fell markedly upon inserting the retraction cord, and this response was invariably present with all the compounds tested. After five-minutes, the decrease became less apparent with cords that were impregnated with physiological saline, AlCl3 or Fe2(SO4)3. Removing the cord elicited a prompt, marked and sustained increase in gingival microcirculation. However, removal of the cord impregnated with 0.1% epinephrine failed to reverse the decreased perfusion, and blood flow to the gingival margin remained low over an additional 20-minute observation. The results of this study indicate that with the exception of epinephrine, all retraction substances tested produced gingival hyperemia for operative procedures. Only epinephrine exhibited sustained vasoconstrictor response. It is anticipated that using laser Doppler flowmetry may be a suitable technique to evaluate the appropriate concentration of epinephrine that would elicit topical vasoconstriction. PMID- 12120769 TI - Histopathologic study on pulp response to single-bottle and self-etching adhesive systems. AB - This study compared the pulp response to seven adhesive resins (three single bottle and four self-etching primers) and their companion resin composite systems with a commercial calcium hydroxide material when applied to exposed monkey pulps. The control group was capped with Dycal (DY), while the experimental groups were capped with one of the following adhesive resin systems: AQ Bond (AQ), Single Bond (SB), Imperva Fluorobond (IF), One Step (OS), Prime&Bond NT (PBNT), Perme Bond F (PBF) and One-up Bond F (OBF). Histopathologic evaluation of pulp tissue disorganization, inflammatory cell infiltration, reparative dentin formation and bacterial penetration at the 3rd, 30th and 90th post-operative days was done using light microscopy. Data were analyzed using the Kruskal-Wallis test followed by the Least Significant Difference Test to determine differences between the control group and the experimental groups at each observation period. The correlation of inflammatory cell infiltration and bacterial presence was investigated by the Kendall correlation analysis. All tests were performed at a 95% level of confidence. The pulpal responses of groups DY, SB, OS, PBF and OBF were generally characterized by none-to-mild pulp tissue disorganization and inflammatory cell infiltration. Also, initiation of reparative dentin formation was found earlier in Group DY, resulting in more complete dentin bridges at the 30- and 90-day observation periods. Groups AQ, IF and PBNT had significantly more inflammatory cell infiltration and a lower incidence of reparative dentin formation than Group DY. A significant correlation was detected between inflammatory cell infiltration and the presence of bacteria. It is concluded that the pulp response to SB, OS, PBF and OBF is not significantly different from the calcium hydroxide preparation. However, calcium hydroxide capping resulted in a higher incidence and faster rate of reparative dentin formation. PMID- 12120771 TI - Fracture resistance of premolars with bonded class II amalgams. AB - This study evaluated the fracture resistance of maxillary premolars with MOD cavity preparation and simulated periodontal ligament. The teeth were restored with silver amalgam (G1), Scotchbond Multi-Purpose Plus and silver amalgam (G2) and Panavia F and silver amalgam (G3). After restorations were made, the specimens were stored at 37 degrees C for 24 hours at 100% humidity and submitted to the compression test in the Universal Testing Machine (Instron). The statistical analysis of the results (ANOVA and Tukey Test) revealed that the fracture resistance of group 2 (G2=105.720 kgF) was superior to those of groups 1 (G1=72.433 kgF) and 3 (G3=80.505 kgF) that did not differ between them. PMID- 12120772 TI - In vitro surface analysis of active and arrested dentinal caries using a pH imaging microscope. AB - This in vitro study evaluated the pH value of active and arrested caries lesions in deep dentinal caries using a pH-imaging microscope (SCHEM-100, HORIBA Ltd, Kyoto, Japan). Buccal-lingual cut sections of extracted human teeth that had either active or arrested dentinal caries lesions were placed on the pH-imaging sensor of the microscope. The pH values were compared statistically by one-way ANOVA and Fisher's PLSD test (p<0.05). In addition, both types of caries lesions were compared with a caries detector solution for this pH-imaging characterization. For both active and arrested lesions in dentin, the lowest pH values in the caries lesions (range from 5.3 to 6.6) were lower than that of intact dentin (range from 6.8 to 7.4). There were statistical differences between the lowest pH value within the active lesion (range from 5.3 to 5.8) and the arrested lesion (range from 6.3 to 6.6) (p<0.05). Although the arrested lesion was unstainable and impermeable to the dye, there was a close relationship between dye staining and pH-imaging characterization within the active lesion from visual inspection. PMID- 12120773 TI - A scanning electron microscopic study of different caries removal techniques on human dentin. AB - Scanning electron microscopy (SEM) evaluated the effect of different caries removal techniques on human dentin topography. Thirty-six extracted human carious mandibular molars were randomly assigned to six groups according to caries removal technique. Carious tissue was removed by hand excavation, bur excavation, air-abrasion, laser ablation, chemomechanical methods and sono-abrasion. The remaining dentin surfaces were replicated and gold-coated. The surfaces were examined using SEM and distinct differences in appearance were observed among specimens treated with different caries removal techniques. While hand-excavated, bur-excavated and air-abraded carious dentin surfaces were covered with a residual smear layer, sono-abrasion with patent dentinal tubules completely removed the smear layer. A few patent orifices of dentinal tubules were observed in dentin subjected to laser ablation and chemo-mechanical caries removal. PMID- 12120774 TI - Comparative analysis of pulpal circulatory reaction to an acetone-containing and an acetone-free bonding agent as measured by vitalmicroscopy. AB - Despite great progress in the production of new dental polymers, application of these products is still controversial. The unlined utilization of cytotoxic adhesive materials on pulpal dentin can adversely influence the pulp, leading to alterations in local microcirculation that can be an early sign of pathological changes. In a previous study by the authors, the effect of an acetone-free bondmaterial was examined on the vascular diameter of pulpal vessels by means of vitalmicroscopy. In this study, experiments comparing experimental data provided by an acetone-containing bondmaterial to these earlier findings with acetone-free ones have been performed. Thirty male Sprague-Dawley rats (weighing 333+/-9 g) were used for this investigation. The first lower incisor was prepared for vitalmicroscopy. Changes in vessel diameter were recorded prior to and 5, 15, 30 and 60 minutes after the investigated materials (Scotchbond Multi-Purpose Dental Adhesive System or Prime & Bond 2.1) were administered on dentin as recommended by the manufacturer. In control rats (saline administration), the vessel diameter was stable during the experiment. In the presence of acetone-free bondmaterial (Scotchbond), the vessel diameter was increased during the experimental period in relation to the baseline (12.15+/-2.85%; 16.36+/-2.39%; 14.16+/-3.48%; 12.12+/ 3.72%). In the presence of acetone-containing bondmaterial (Prime & Bond 2.1), a similar result was observed (10.56+/-2.27%; 16.13+/-2.94%; 17.88+/-2.54%; 14.54+/ 3.16%). The differences between the control values and those registered with test groups were significant (p<0.05; ANOVA). There was no significant difference among the test groups. The results of this study suggest that dental bond materials applied on a very thin layer of dentin may affect the blood supply to the dental pulp. However, no stasis or prestasis has been detected, indicating a possible reversible effect. The authors could not show any statistical difference between the vasodilatation caused by the acetone-containing and the acetone-free bond material. PMID- 12120775 TI - Effect of thickness of flowable resins on marginal leakage in class II composite restorations. AB - Despite limited scientific evaluation, there is an increased use of low elastic modulus flowable resin composite (FRC) as a stress-relieving gingival increment in Class II restorations. This study compared marginal leakage in preparations with gingival margins in enamel or dentin/cementum (sub-CEJ and supra-CEJ) after FRC was used as a gingival increment to hybrid resin composite used alone. In addition, the extent of leakage around restorations with or without the use of FRC gingival increments when light curing the resin composites from occlusal direction only or buccal, lingual and occlusal directions was compared. Sixty extracted human molars were prepared with two identical Class II (MO and OD) preparations (30 were 1 mm sub-CEJ and 30 were 1 mm supra-CEJ) and randomly assigned to six groups. After etching, dentin-bonding agent was applied to all prepared tooth surfaces according to the manufacturer's specifications. One of three different thicknesses of FRC (0.5 mm, 1 mm or 2 mm) was placed on the gingival floor, cured and a hybrid resin composite was placed occlusally to complete the restoration. The control preparation on each tooth was restored in the same manner, except that a hybrid resin composite was used for both the gingival and occlusal increments. The restored teeth were thermocycled (300 cycles), then immersed in 50% silver nitrate prior to the hemi-section and measured for leakage under a light microscope. The data were evaluated using paired measures analysis of variance (ANOVA). Most of the occlusal margins showed no leakage, while almost every gingival margin demonstrated some silver nitrate penetration regardless of whether it was located sub or supra-CEJ, although significantly less leakage was found in restorations with supra-CEJ margins (p=0.0001). Among supra-CEJ restorations, there was a pronounced reduction in leakage as FRC thickness increased (p=0.0005). In the teeth restored with the gingival-margin located supra-CEJ, the 2 mm thickness FRC gingival increment showed significantly less leakage (p<0.01) compared with the 0.5 mm thickness of FRC gingival increment. The direction of the curing light did not affect the extent of leakage (p>0.05). The use of FRC material as a gingival increment sub CEJ in posterior hybrid resin restorations produced no significant difference in leakage (p>0.05). The results of this study indicated that restorations located supra-CEJ (with gingival margins in enamel) with 2 mm thick FRC gingival increments demonstrated significantly less leakage than did those with 0.5 mm FRC. When the margin of the restoration was located sub-CEJ (in dentin/cementum), neither the thickness nor the presence of FRC as a gingival increment significantly influenced the marginal leakage. PMID- 12120776 TI - Surface texture of resin-modified glass ionomer cements: effects of finishing/polishing systems. AB - This study investigated the surface texture of two resin-modified glass ionomer cements (RMGICs) in the vertical and horizontal axis after treatment with different finishing/polishing systems. Class V preparations were made on the buccal and lingual/palatal surfaces of freshly extracted teeth. The cavities on each tooth were restored with Fuji II LC (GC) and Photac-Fil Quick (ESPE) according to manufacturers' instructions. Immediately after light-polymerization, gross finishing was done with 8-flute tungsten carbide burs. The teeth were then randomly divided into four groups and finished/polished with (a) Robot Carbides (RC); (b) Super-Snap system (SS); (c) OneGloss (OG) and (d) CompoSite Points (CS). The sample size for each material-finishing/polishing system combination was eight. The mean surface roughness (microm) in vertical (RaV) and horizontal (RaH) axis was measured using a profilometer. Data was subjected to ANOVA/Scheffe's tests and Independent Samples t-test at significance level 0.05. Mean RaV ranged from 0.59-1.31 and 0.83-1.52, while mean RaH ranged from 0.80 1.43 and 0.85-1.58 for Fuji II LC and Photac-Fil, respectively. Results of statistical analysis were as follows: Fuji II LC: RaV-RC, SS0.01). The SEM observations showed that both adhesive systems etched the primary enamel deeper than the permanent enamel, suggesting that the action of acid etch seemed to be more intense on primary enamel than on permanent enamel. Bonding of the adhesive systems to primary enamel was almost identical to permanent enamel. PMID- 12120778 TI - The effect of rebonding on microleakage of class V aesthetic restorations. AB - This in vitro study evaluated the effect of rebonding on microleakage of a resin composite, a condensable resin and two polyacid-modified resin composite restorations. Standardized cylindrical Class V dentin cavities were prepared on the buccal root surfaces of 240 extracted bovine incisive teeth. The prepared teeth were randomly assigned to four groups of 60 teeth and restored with the following restorative systems: I--(ZS) Z100/Scotchbond Multi-Purpose Plus; II- (SS) Solitaire/Solid Bond; III--(FS) Freedom/Stae; IV--(FSB) F200/Single Bond. Thirty teeth of each group were rebonded with a low-viscosity resin (Fortify/BISCO), according to the manufacturer's instructions. The remaining teeth received no treatment. All teeth were thermocycled for 5,000 cycles and brushed by hand three times a day for 10 days using a toothbrush and a slurry of dentifrice and water. Specimens were stained in a 2% methylene blue solution and longitudinally sectioned with diamond disks. Microleakage was scored on a scale of 0 to 3. The Kruskal-Wallis test showed statistically significant differences among the groups (h=156.54; alpha<0.05). Pairwise comparison by means of the least significant difference showed that (SS) and (FS) with or without rebonding were not statistically different from each other. These groups showed the highest microleakage differences from (ZS) and (FSB) with or without rebonding. (ZS) with rebonding showed the lowest microleakage that was not statistically different from (ZS) without rebonding and (FSB) with rebonding. PMID- 12120780 TI - Curing light intensity effects on wear resistance of two resin composites. AB - This in vitro study evaluated the wear resistance of resin composite polymerized using four different light-curing systems. For this, a well-defined cylindrical cavity preparation (4.0 mm in diameter x 3.0 mm in depth) was made in a ceramic block (n=4 per material/light condition). Uncured material, either a universal hybrid composite (Herculite XRV) or a flowable hybrid composite (Revolution Formula 2), was packed and light-cured from the top surface only with one of the four light-curing units: 1) a conventional quartz-tungsten-halogen light, 2) a soft-start light, 3) an argon-ion laser or 4) a plasma-arc curing light. After storing the specimens in deionized water at 37 degrees C for 24 hours, the excess cured material was ground through successive grits up to a final 1200-grit SiC abrasive. The specimens were placed in deionized water at 37 degrees C for an additional 24 hours. Wear simulation was performed using a four-station Leinfelder-type three-body wear device. A slurry of water and unplasticized polymethylmethacrylate beads, simulating an artificial food bolus, was placed on the surface of each resin-composite-restored ceramic block. The entire cycling procedure was carried out 400,000 times. Impressions of each resin composite surface were taken with polyvinylsiloxane and epoxy replicas were made. Wear analyses were conducted by generating tracings across the worn surface of epoxy replicas using profilometer scans. For the universal hybrid composite and the flowable hybrid composite, the lowest wear occurred in specimens that were cured using the conventional quartz-tungsten-halogen light, and the highest wear was detected on those specimens made using the argon-ion laser. For both resin composites, the mean wear for specimens cured using the argon-ion laser was significantly higher than that of the specimens cured with the three other lights, which were statistically similar. PMID- 12120781 TI - Polyadenylate polymerase (PAP) and 3' end pre-mRNA processing: function, assays, and association with disease. AB - Polyadenylate polymerase (PAP) is one of the enzymes involved in the formation of the polyadenylate tail of the 3' end of mRNA. Poly (A) tail formation is a significant component of 3' processing, a link in the chain of events, including transcription, splicing, and cleavage/polyadenylation of pre-mRNA. Transcription, capping, splicing, polyadenylation, and transport take place as coupled processes that can regulate one another. The poly(A) tail is found in almost all eukaryotic mRNA and is important in enhancing translation initiation and determining mRNA stability. Control of poly(A) tail synthesis could possibly be a key regulatory step in gene expression. PAP-specific activity values are measured by a highly sensitive assays and immunocytochemical methods. High levels of PAP activity are associated with rapidly proliferating cells, it also prevents apoptosis. Changes of PAP activity may cause a decrease in the rate of polyadenylation in the brain during epileptic seizures. Testis-specific PAP may play an important role in spermiogenesis. PAP was found to be an unfavorable prognostic factor in leukemia and breast cancer. Furthermore, measurements of PAP activity may contribute to the definition of the biological profile of tumor cells. It is crucial to know the specific target causing the elevation of serum PAP, for it to be used as a marker for disease. This review summarizes the recently accumulated knowledge on PAP including its function, assays, and association with various human diseases, and proposes future avenues for research. PMID- 12120782 TI - Effect of alcohol on lipids and lipoproteins in relation to atherosclerosis. AB - Several studies indicate that light-to-moderate alcohol consumption is associated with a low prevalence of coronary heart disease. An increase in high-density lipoprotein (HDL) cholesterol is associated with alcohol intake and appears to account for approximately half of alcohol's cardioprotective effect. In addition to changes in the concentration and composition of lipoproteins, alcohol consumption may alter the activities of plasma proteins and enzymes involved in lipoprotein metabolism: cholesteryl ester transfer protein, phospholipid transfer protein, lecithin:cholesterol acyltransferase, lipoprotein lipase, hepatic lipase, paraoxonase-1 and phospholipases. Alcohol intake also results in modifications of lipoprotein particles: low sialic acid content in apolipoprotein components of lipoprotein particles (e.g., HDL apo E and apo J) and acetaldehyde modification of apolipoproteins. In addition, "abnormal" lipids, phosphatidylethanol, and fatty acid ethyl esters formed in the presence of ethanol are associated with lipoproteins in plasma. The effects of lipoproteins on the vascular wall cells (endothelial cells, smooth muscle cells, and monocyte/macrophages) may be modulated by ethanol and the alterations further enhanced by modified lipids. The present review discusses the effects of alcohol on lipoproteins in cholesterol transport, as well as the novel effects of lipoproteins on vascular wall cells. PMID- 12120783 TI - Principles derived from the study of simple skills do not generalize to complex skill learning. AB - We review research related to the learning of complex motor skills with respect to principles developed on the basis of simple skill learning. Although some factors seem to have opposite effects on the learning of simple and of complex skills, other factors appear to be relevant mainly for the learning of more complex skills. We interpret these apparently contradictory findings as suggesting that situations with low processing demands benefit from practice conditions that increase the load and challenge the performer, whereas practice conditions that result in extremely high load should benefit from conditions that reduce the load to more manageable levels. The findings reviewed here call into question the generalizability of results from studies using simple laboratory tasks to the learning of complex motor skills. They also demonstrate the need to use more complex skills in motor-learning research in order to gain further insights into the learning process. PMID- 12120784 TI - Stimulus-response compatibility and psychological refractory period effects: implications for response selection. AB - The purpose of this paper was to provide insight into the nature of response selection by reviewing the literature on stimulus-response compatibility (SRC) effects and the psychological refractory period (PRP) effect individually and jointly. The empirical findings and theoretical explanations of SRC effects that have been studied within a single-task context suggest that there are two response-selection routes-automatic activation and intentional translation. In contrast, all major PRP models reviewed in this paper have treated response selection as a single processing stage. In particular, the response-selection bottleneck (RSB) model assumes that the processing of Task 1 and Task 2 comprises two separate streams and that the PRP effect is due to a bottleneck located at response selection. Yet, considerable evidence from studies of SRC in the PRP paradigm shows that the processing of the two tasks is more interactive than is suggested by the RSB model and by most other models of the PRP effect. The major implication drawn from the studies of SRC effects in the PRP context is that response activation is a distinct process from final response selection. Response activation is based on both long-term and short-term task-defined S-R associations and occurs automatically and in parallel for the two tasks. The final response selection is an intentional act required even for highly compatible and practiced tasks and is restricted to processing one task at a time. Investigations of SRC effects and response-selection variables in dual-task contexts should be conducted more systematically because they provide significant insight into the nature of response-selection mechanisms. PMID- 12120786 TI - Learning to vary and varying to learn. AB - We compared two sources of behavior variability: decreased levels of reinforcement and reinforcement contingent on variability itself. In Experiment 1, four groups of rats were reinforced for different levels of response-sequence variability: one group was reinforced for low variability, two groups were reinforced for intermediate levels, and one group was reinforced for very high variability. All of the groups experienced three different reinforcement frequencies for meeting their respective variability contingencies. Results showed that reinforcement contingencies controlled response variability more than did reinforcement frequencies. Experiment 2 showed that only those animals concurrently reinforced for high variability acquired a difficult-to-learn sequence; animals reinforced for low variability learned little or not at all. Variability was therefore controlled mainly by reinforcement contingencies, and learning increased as a function of levels of baseline variability. Knowledge of these relationships may be helpful to those who attempt to condition operant responses. PMID- 12120785 TI - Activity-based anorexia: ambient temperature has been a neglected factor. AB - Activity-based anorexia refers to the self-starvation of rats exposed to experimental conditions that combine restricted access to food with access to an activity wheel. This paper compares previous studies of this phenomenon in relation to the ambient temperatures (AT) that were employed. On this basis, and from some more direct evidence, we argue that AT is an important, but neglected, factor in activity-based anorexia research. More attention to AT is needed in future research, since its neglect threatens the validity of conclusions drawn from those studies. Furthermore, direct examination of the effect of AT on activity-based anorexia will allow a better understanding of the mechanisms underlying this phenomenon and the possible clinical implications for the treatment of human anorexia nervosa. PMID- 12120787 TI - Self-control in honeybees. AB - Self-control means choosing a large delayed reward over a small immediate reward; impulsiveness is its opposite. The metabolic hypothesis states that the amount of self-control across species correlates negatively with metabolic rate (Tobin & Logue, 1994). Foraging honeybees have high metabolic rates; the metabolic hypothesis would predict little self-control in bees. But foraging bees work for the long-term good of their hive, conditions that seem to require self-control. In three experiments, we gave bees the choice between (1) a sweeter delayed reward and a less sweet immediate reward and (2) a large delayed reward and a small immediate reward. Bees showed much self-control, inconsistent with the metabolic hypothesis. PMID- 12120788 TI - A new effect of luminance gradient on achromatic simultaneous contrast. AB - A new effect in the domain of achromatic simultaneous contrast has been observed. A middle gray region placed at the center of an area filled by a linear achromatic gradient from black (outer part) to white (inner part) is perceived as being much darker than an identical middle gray region surrounded by a reversed gradient. By using a matching task in two experiments, it has been shown that this phenomenon is much stronger than the classical achromatic simultaneous contrast effect. The new effect is interpreted in terms of the albedo hypothesis. PMID- 12120790 TI - A diffusion model account of response time and accuracy in a brightness discrimination task: fitting real data and failing to fit fake but plausible data. AB - A brightness discrimination experiment was performed to examine how subjects decide whether a patch of pixels is "bright" or "dark," and stimulus duration, brightness, and speed versus accuracy instructions were manipulated. The diffusion model (Ratcliff, 1978) was fit to the data, and it accounted for all the dependent variables: mean correct and error response times, the shapes of response time distributions for correct and error responses, and accuracy values. Speed-accuracy manipulations affected only boundary separation (response criteria settings) in the model. Drift rate (the rate of accumulation of evidence) in the diffusion model, which represents stimulus quality, increased as a function of stimulus duration and stimulus brightness but asymptoted as stimulus duration increased from 100 to 150 msec. To address the argument that the diffusion model can fit any pattern of data, simulated patterns of plausible data are presented that the model cannot fit. PMID- 12120789 TI - Spatial-frequency uncertainty and cuing effects on psychometric functions for contrast detection. AB - In the present study, the effects of spatial-frequency uncertainty and cuing on psychometric functions for contrast detection of sinusoidal gratings are examined. For this purpose, psychometric functions were collected from 4 subjects under fixed-frequency, randomized-frequency, and cued-frequency conditions. The experiment was conducted with a temporal two-alternative forced-choice task, and five spatial frequencies in the range of 0.5 and 8.0 c/deg and seven contrast levels for each frequency were used. The results showed that the psychometric functions for the randomized-frequency condition were shallower than those for the fixed-frequency condition, supporting the single-band model for the uncertainty effects (Hubner, 1993a, 1993b). For the cued-frequency condition, the slopes of the functions were not clearly different from those for the randomized condition. These results clearly differ from those of Hubner (1996b), which showed, in the spatial two-alternative forced-choice task, steeper psychometric functions for the randomized-frequency condition than those for the fixed- and cued-frequency conditions, supporting the multiple-band model (Hubner, 1993a, 1993b). The difference suggests that the single-band model applies to the uncertainty effects in the temporal forced-choice task, whereas the multiple-band model does so in the spatial forced-choice task. PMID- 12120791 TI - Successor states in a four-state ambiguous figure. AB - The satiation theory of ambiguous figures holds that interpretation shifts are caused by fatigue of neural arrangements responsible for the prevailing interpretation. A four-state multistable figure is introduced, in which two depicted cubes can be seen as connected or unconnected and as facing up or facing down. Observers viewed the figure for 4 min. When descriptive labels were used for the interpretations, shifts to interpretations that shared neither dimension were significantly more frequent than shifts that conserved orientation or connection/disconnection. However, all types of transitions were equally likely when arbitrary letter codes were used, implying that the putatively fatigued assemblies can be dedicated to whole figures or to their characteristics, depending on observer expectations. PMID- 12120792 TI - Hemispheric asymmetries in categorical judgments of direction versus coordinate judgments of velocity of motion. AB - Three experiments on visual field differences in motion perception are reported. Experiment 1 employed circular stimuli that grew or shrank either quickly or slowly. Experiments 2 and 3 employed circles that moved upward or downward either quickly or slowly. Judgments based on categorical equivalence classes (i.e., grow/shrink, upward/downward) generally yielded small and nonsignificant right visual field advantages. Judgments based on the precise coordinates of motion (i.e., quickly/slowly) yielded significant left visual field advantages across all three experiments. Results are interpreted in light of Kosslyn's (1987) model of hemispheric differences in the processing of categorical versus coordinate spatial relations. PMID- 12120793 TI - The Doppler effect is not what you think it is: dramatic pitch change due to dynamic intensity change. AB - Historically, auditory pitch has been considered to be a function of acoustic frequency, with only a small effect being due to absolute intensity. Yet we found that when tones are Doppler shifted so that frequency drops, the pitch dramatically rises and falls, closely following the pattern of dynamic intensity change. We show that continuous intensity change can produce pitch variation comparable to a frequency change approaching an octave. This effect opposes and is an order of magnitude larger than the well-known effect of discrete intensity change in the frequency range employed. We propose that the perceptual interaction of continuous changes in pitch and loudness reflects a natural correlation between changes in frequency and intensity that is neurally encoded to facilitate the parsing and processing of meaningful acoustic patterns. PMID- 12120794 TI - Eye gaze is not unique: automatic orienting in response to uninformative arrows. AB - Recent studies (Driver et al., 1999; Friesen & Kingstone, 1998; Langton & Bruce, 1999) have argued that the perception of eye gaze may be unique, as compared with other symbolic cues (e.g., arrows), in being able to automatically trigger attentional orienting. In Experiment 1, 17 participants took part in a visuospatial orienting task to investigate whether arrow cues might also trigger automatic orienting. Two arrow cues were presented for 75 msec to the left and right of a fixation asterisk. After an interval of either 25 or 225 msec, the letter O or X appeared. After both time intervals, mean response times were reliably faster when the arrows pointed toward, rather than away from, the location of the target letter. This occurred despite the fact that the participants were informed that the arrows did not predict where the target would appear. In Experiment 2, the same pattern of data was recorded when several adjustments had been made in an attempt to rule out alternative explanations for the cuing effects. Overall, the findings suggest that the eye gaze is not unique in automatically triggering orienting. PMID- 12120795 TI - Attentional coding of categorical relations in scene perception: evidence from the flicker paradigm. AB - The purpose of the present investigation was to determine whether the positions of objects in a scene are coded relative to one another categorically (i.e., above, below, or side of; Experiment 1) and to determine whether spatial position in scene perception is coded preattentively or only under focused attention (Experiment 2). In Experiment 1, participants viewed alternating versions of a scene in which one of the objects in the scene changed its categorical relationship to the closest object in the scene, changed only its metric relationship to the closest object in a scene, or appeared and disappeared. Participants were faster at detecting changes that disrupted categorical relations than at detecting changes that disrupted only metric relations. In Experiment 2, this categorical advantage still occurred even when participants were cued to the location of the change. These results suggest that categorical spatial relations are being coded in scene perception and that attention is required in order to encode spatial relations. PMID- 12120796 TI - Recognizing the un-real McCoy: priming and the modularity of face recognition. AB - Fodor (1983) has proposed that face perception is carried out by an informationally encapsulated module, whose operation is unaffected by context or expectancies. We tested the modularity hypothesis by examining whether discriminations between normal and distorted versions of famous faces can be primed, either by the name of an associated person (semantic context) or by a valid cue as to the identity of the target face (expectancy). A preliminary experiment showed that, in the absence of priming, discriminations between normal and distorted versions of a face were unaffected by whether the target faces were familiar or not, confirming that these judgments tap perceptual, not postperceptual (semantic), coding processes. In Experiment 1, accuracy was significantly higher when target face pairs were preceded by related name primes, as compared with unrelated ones. In Experiment 2, reaction times were significantly faster for targets preceded by a valid identity cue than for targets preceded by an invalid one. Neither effect could be explained as a speed accuracy tradeoff. These results fail to support Fodor's conjecture that face processing is encapsulated. PMID- 12120797 TI - Pitch characteristics of infant-directed speech affect infants' ability to discriminate vowels. AB - "Baby talk" or speech directed to prelinguistic infants is high in pitch and has exaggerated pitch contours (up/down patterns of pitch change) across languages and cultures. Using an acoustic model, we predicted that the large pitch contours of infant-directed speech should improve infants' ability to discriminate vowels. On the other hand, the same model predicted that high pitch would not benefit, and might actually impair, infants' ability to discriminate vowels. We then confirmed these predictions experimentally. We conclude that the exaggerated pitch contours of infant-directed speech aid infants' acquisition of vowel categories but that the high pitch of infant-directed speech must serve another function, such as attracting infants' attention or aiding emotional communication. PMID- 12120798 TI - The influence of the lexicon on speech read word recognition: contrasting segmental and lexical distinctiveness. AB - The neighborhood activation model (NAM; P. A. Luce & Pisoni, 1998) of spoken word recognition was applied to the problem of predicting accuracy of visual spoken word identification. One hundred fifty-three spoken consonant-vowel-consonant words were identified by a group of 12 college-educated adults with normal hearing and a group of 12 college-educated deaf adults. In both groups, item identification accuracy was correlated with the computed NAM output values. Analysis of subsets of the stimulus set demonstrated that when stimulus intelligibility was controlled, words with fewer neighbors were easier to identify than words with many neighbors. However, when neighborhood density was controlled, variation in segmental intelligibility was minimally related to identification accuracy. The present study provides evidence of a common spoken word recognition system for both auditory and visual speech that retains sensitivity to the phonetic properties of the input. PMID- 12120799 TI - Privileged access to action for objects relative to words. AB - We compared action (pour or twist?) and contextual/semantic (found in kitchen?) decisions made to pictures of objects, nonobjects, and words. Although there was no advantage for objects over words in contextual/semantic decisions, there was an advantage for objects over words and nonobjects in action decisions. For objects, both action and contextual/semantic decisions were faster than naming; for words, the opposite occurred. These results extend the early results of Potter and Faulconer (1975) that there is privileged access to semantic memory for objects relative to that for words and privileged access to phonology for words. Our data suggest that, for objects, there is privileged access to action knowledge rather than to all forms of semantic knowledge and that this is contingent on learned associations between objects and actions. PMID- 12120800 TI - The accessibility of characters in single sentences: proper names, common nouns, and first mention. AB - Accessibility of characters in two-character sentences (e.g., The butler helped Calvin at the wedding reception) was investigated with a probe recognition task. Probes were either the first character (e.g., butler) or the second character (e.g., Calvin) in a sentence and were designated by proper names or common nouns crossed with name or noun nonprobes. Results show that (1) probes in first position are more accessible than those in second position, but not when noun probes are paired with name nonprobes, (2) characters designated by names are generally more accessible than those designated by nouns, and (3) the first name in a sentence is more available than other characters, regardless of position. Thus, accessibility of characters in a sentence seems dependent on discourse function, with named characters seen as main characters, rather than on nondiscourse-related factors, such as temporal distinctiveness. PMID- 12120801 TI - Dissociation between priming and recognition in the expression of sequential knowledge. AB - Exposure to a repeating sequence of target stimuli in a speeded localization task can support both priming of sequence-consistent responses and recognition of sequence components. Here, a task is introduced in which measures of priming and recognition are obtained concurrently, and it is demonstrated that priming of sequence-consistent responses occurs even when test stimuli are not recognized. The results show that sequence knowledge can be expressed in the absence of conscious recognition. However, we also show that this result is consistent with a simple model in which priming and recognition depend on exactly the same underlying memory strength variable. PMID- 12120802 TI - Context matching and judgments of recency. AB - An experiment was done to test a context-matching explanation of memory for recency under steady-state conditions. Subjects went through a list of 550 names, in which individual names were repeated at lags of 5-30 other items. The names were shown in two different styles or contexts. An old versus new recognition decision was made on each name, and each old response was followed by a numerical judgment of recency (JOR). When first- and second-presentation contexts were the same, recognition hit rates were higher, and mean JORs were shorter (more recent), than when the two contexts were different. The JOR result is as predicted by the context-matching hypothesis. PMID- 12120803 TI - The emergence of item-specific encoding effects in between-subjects designs: perceptual interference and multiple recall tests. AB - The perceptual-interference effect occurs when interference with word perception (by backward masking) enhances later memory for the word. In terms of the item specific-relational framework (Hunt & McDaniel, 1993), this effect is similar to other manipulations that enhance item-specific encoding (such as the generation effect). One similarity is that item-specific effects typically do not arise in between-subjects designs. However, the present experiment demonstrates that a between-subjects perceptual-interference effect emerges over multiple recall tests. Furthermore, perceptual interference produces both more intertest gains (indexing enhanced item-specific processing) and more intertest losses (indexing disrupted relational encoding) compared with the intact (control) condition. Finally, delaying the mask to a point at which it no longer interferes with perception (266 msec) eliminates both the perceptual-interference recall advantage and the increase in intertest gains. This condition still produces more intertest losses, however. Together, these results imply that a delayed mask disrupts relational encoding but produces no item-specific enhancement, dissociating the two effects of the perceptual-interference manipulation. PMID- 12120804 TI - Directed forgetting of actions by younger and older adults. AB - Memory for actions that are performed is substantially better than memory for descriptions of actions (e.g., Earles, 1996). In fact, people may form memories for actions even if they do not intend to or want to remember them. The directed forgetting paradigm was used to test the ability of younger and older adults to intentionally forget simple actions (also known as subject-performed tasks, or SPTs). Participants were asked to perform the action described by a verb-noun pair (e.g., break toothpick) or to read the pair, but not to perform the action. Following each pair, the participants were told either to remember or to forget the pair. Younger adults intentionally forgot verbally encoded pairs significantly better than did older adults. Actions that were performed, however, were difficult for both younger and older adults to intentionally forget. The performance of an action thus seems to result in strong item-specific processing that makes the action difficult to intentionally forget even for younger adults who can successfully intentionally forget verbally encoded items. PMID- 12120805 TI - Part-set cuing of false memories. AB - Part-set cuing inhibition describes the common finding that re-presenting items from a word list can reduce subjects' overall recall performance for studied items. Do part-set cuing effects occur for false memories as well? In the present experiments, subjects studied lists of words drawn from Roediger and McDermott (1995). After studying each list, subjects completed math problems and then recalled the list items either with or without accompanying list cues. In Experiment 1, the recall cues consisted of items drawn randomly from the original list. In Experiment 2, an additional type of cued recall task was added in which the even numbered list items were used as cues. Taken together, these experiments demonstrate robust part-set cuing effects for critical nonpresented items. In addition, they show that whereas recall of critical words is reduced by the presence of cues at test, retrieval cues do not affect critical words and studied words in exactly the same manner. PMID- 12120806 TI - Quantile maximum likelihood estimation of response time distributions. AB - We introduce and evaluate via a Monte Carlo study a robust new estimation technique that fits distribution functions to grouped response time (RT) data, where the grouping is determined by sample quantiles. The new estimator, quantile maximum likelihood (QML), is more efficient and less biased than the best alternative estimation technique when fitting the commonly used ex-Gaussian distribution. Limitations of the Monte Carlo results are discussed and guidance provided for the practical application of the new technique. Because QML estimation can be computationally costly, we make fast open source code for fitting available that can be easily modified to use QML in the estimation of any distribution function. PMID- 12120807 TI - Effect of valerian, Valeriana edulis, on sleep difficulties in children with intellectual deficits: randomised trial. AB - Serious sleep problems are common in children with an intellectual deficit (ID), and are often the source of much distress for both the child and caregivers. As yet, no satisfactory long-term treatment exists for intransigent sleep difficulties in children with an ID. Valerian, Valeriana spp., has been used for thousands of years to induce relaxation and sleep. Scientific investigation of valerian's sleep promoting ability in humans, whilst limited, has yielded promising findings. This initial study aimed to explore valerian's potential for assisting in the treatment of sleep problems in children with an ID. Five children with varying intellectual deficits and different primary sleep problems underwent eight continuous weeks of monitoring via sleep diaries, adhering to a double blind, placebo controlled and randomised design. Compared to baseline and placebo, valerian treatment led to significant reductions in sleep latencies and nocturnal time awake, lengthened total sleep time and improved sleep quality. The treatment was apparently most effective in children with deficits that involved hyperactivity. Although the findings are preliminary and in need of replication, there is evidence to suggest that valerian may be useful in the safe and effective long-term treatment of intransigent sleep difficulties in children with ID's, and therefore warrants further investigation. PMID- 12120808 TI - Incidence and clinical features of liver injury related to Kampo (Japanese herbal) medicine in 2,496 cases between 1979 and 1999: problems of the lymphocyte transformation test as a diagnostic method. AB - We retrospectively examined the summaries of all admission records of patients from 1979 to 1999 in our department, and selected for further study all liver injuries suspected of being related to Kampo medicines. Among 2,496 summaries, 30 summaries described liver disorders suspected of being related to Kampo medicines. Whether there was a causal relationship between the use of Kampo medication and the occurrence of liver injury was assessed according to the criteria described by Haller and Benowitz (2000), independently of the results of the lymphocyte transformation test (LTT). Among 30 events, we concluded that 9 were definitely unrelated, and 6 were probably unrelated to the use of Kampo medicines. Nine events (0.36% of 2,496 patient admissions and 0.06% of 14,616 outpatients) were considered possibly related, and only 6 events (0.24% of 2,496 patient admissions and 0.04% of 14,616 outpatients) were judged to be definitely or probably related to Kampo medicines. Low-grade eosinophilia was observed in a few patients of these "related" groups, and no fever or rash was observed in these "related" groups. Other clinical features, including type of liver injury, duration of Kampo medicine-use, recovery period and laboratory data, were not different from liver injuries associated with western drugs. Most patients in the definitely "unrelated" group were positive in the LTT for the suspect Kampo medicine, suggesting that the LTT may be unreliable for the diagnosis of Kampo medicine-induced liver injury. From 1979 to 1999, our use of Kampo medicines to treat patients resulted in a low rate of liver injury and no fatalities. PMID- 12120809 TI - Induction of apoptosis in a human erythroleukemic cell line K562 by tylophora alkaloids involves release of cytochrome c and activation of caspase 3. AB - Tylophora alkaloids are plant products known for their antiasthamatic and antiproliferative activities. The underlying cellular changes resulting from inhibition of proliferation were investigated. Tylophora alkaloids induced apoptosis in K562 cells with characteristic apoptotic features like nuclear condensation, apoptotic body formation, flipping of membrane phosphatidylserine, activation of caspase 3 and release of mitochondrial cytochrome c. These studies suggest that the Tylophora alkaloids, in addition to their antiproliferative effects also induce apoptosis in erythroleukemic cells. These observations imply that Tylophora alkaloids could be useful molecules for their antiproliferative activity and for induction of apoptosis in tumor cells. PMID- 12120810 TI - Effects of veratrine and paeoniflorin on isolated mouse vas deferens. AB - In this study, we attempted to identify the interactions and mechanisms between veratrine and paeoniflorin on isolated mouse vas deferens. Paeoniflorin had no effect on isolated mouse vas deferens. Veratrine (1 x 10(-5) approximately 1 x 10(-3) g/ml) could directly induce contraction of isolated rat and mouse vas deferens. The concentration induced by veratrine (1 x 10(-5) g/ml) was completely inhibited by Ca2+-free solution and verapamil (1 x 10(-5) M), in both the epididymal and the prostatic portions of isolated mouse vas deferens. Naloxone (1 x 10(-5) M) did not alter the contraction induced by veratrine (1 x 10(-5) g/ml) in either the epididymal or the prostatic portions of isolated mouse vas deferens. Paeoniflorin (4.8 x 10(-5) g/ml) inhibited the contraction induced by veratrine (1 x 10(-5) g/ml) in both the epididymal and the prostatic portions of isolated mouse vas deferens. Paeoniflorin (4.8 x 10(-5) g/ml) potentiated norepinephrine (1 x 10(-5) M)-induced phasic contraction in the epididymal portion, but decreased contractions in the prostatic portion. Paeoniflorin (4.8 x 10(-5) g/ml) increased KCI (56 mM)-induced phasic contraction in the epididymal portion, but decreased the tonic contraction in either the epididymal or the prostatic portion. Veratrine (1 x 10(-5) g/ml)-induced contractions could be decreased by pretreatment with ryanodine (1 x 10(-5) M) in both the epididymal and the prostatic portions. Pretreatment with the combination of paeoniflorin (4.8 x 10(-5) g/ml) and ryanodine (1 x 10(-5) M) did not potentiate the inhibition of paeoniflorin in the veratrine-induced contraction in both the epididymal and the prostatic portions of isolated mouse vas deferens. PMID- 12120811 TI - Antioxidant property of Celastrus paniculatus willd.: a possible mechanism in enhancing cognition. AB - In the present study aqueous, methanolic, chloroform and petroleum ether extracts of seeds of Celastrus paniculatus were investigated for their effect on cognitive functions in rats. Male Wistar rats weighing 200-250 g each were used to study effect on learning and memory through use of the shuttle-box, step-through, step down and elevated plus maze paradigms. Only the aqueous seed extract (200 mg/kg body wt. for 14 days) showed an improvement in learning and memory in both the shuttle-box and step-through paradigms. Therefore, further experiments were conducted using the aqueous extract at 100, 200 and 300 mg/kg body wt. doses in different paradigms of cognition. All three doses of the aqueous extract increased the number of avoidances in the shuttle-box and step-through latency the in step-through apparatus, but no significant difference was observed between the doses tested. In the step-down apparatus, the 200- and 300-mg/kg body wt. doses of aqueous extract showed a significant increase in step-down latency, whereas no significant difference was observed in the elevated-plus-maze paradigm between drug-treated and vehicle-treated groups. Since the behavioral impairments are associated with oxidative stress, we investigated the effect of the aqueous extract on oxidative stress parameters. Among the three doses tested, only 200 and 300 mg/kg body wt. stimulated a significant decrease in the brain levels of malondialdehyde, with simultaneous significant increases in levels of glutathione and catalase. The present findings indicate that the aqueous extract of Celastrus paniculatus seed has cognitive-enhancing properties and an antioxidant effect might be involved. PMID- 12120812 TI - Antioxidant, cyclooxygenase and topoisomerase inhibitory compounds from Apium graveolens Linn. seeds. AB - Cyclooxygenase inhibitory and antioxidant bioassay-directed extraction and purification of celery seeds yielded sedanolide (1), senkyunolide-N (2), senkyunolide-J (3), 3-hydroxymethyl-6-methoxy-2,3-dihydro-1H-indol-2-ol (4), L tryptophan (6), and 7-[3-(3,4-dihydroxy-4-hydroxymethyl-tetrahydro-furan-2-yloxy) 4,5-dihydroxy-6-hydroxymethyl-tetrahydro-pyran-2-yloxy]-5-hydroxy-2-(4-hydroxy-3 methoxy-phenyl)-chromen-4-one (7). The structures of compounds 1-7 were determined using spectroscopic methods. Compound 4 is reported here for the first time. At 250 pg ml(-1), compounds 1-4, 6 and 7 displayed prostaglandin H endoperoxide synthase-I (COX-I) and prostaglandin H endoperoxide synthase-II (COX II) inhibitory activities at pH 7. The acetylated product (5) of compound 4 also inhibited COX-I and COX-II enzymes when tested at 250 microg ml(-1). Compounds 6 and 7 exhibited good antioxidant activity at concentrations of 125 and 250 microg ml(-1). Only compounds 1-3 exhibited topoisomerase-I and -II enzyme inhibitory activity at concentrations of 100, 200 and 200 microg ml(-1), respectively. PMID- 12120813 TI - The fruiting body and its caterpillar host of Cordyceps sinensis show close resemblance in main constituents and anti-oxidation activity. AB - Cordyceps (summer-grass, winter-worm), one of the most valued traditional Chinese medicines, is used commonly for the replenishment of body health. It consists of the dried fungus Cordyceps sinensis growing on caterpillar larvae. For medication, the fruiting body (fungus) and the worm (caterpillar) are used together. However, the pharmacological efficiency and the main constituents of the individual parts have not been determined. In the present study the water extracts from the fruiting body and worm of natural Cordyceps were analyzed for their content of nucleosides and polysaccharides; the results showed that the worm had chemical composition similar to the fruiting body. In addition, both the fruiting body and worm of Cordyceps showed similar potency in their anti oxidation activities in the xanthine oxidase assay, the induction of hemolysis assay and the lipid-peroxidation assay. These results suggest that the function of the worm in Cordyceps is to provide a growth medium for the fruiting body, and that eventually, the worm is totally invaded by C. sinensis mycelia. PMID- 12120814 TI - Anti-inflammatory and antioxidant activities of cat's claw (Uncaria tomentosa and Uncaria guianensis) are independent of their alkaloid content. AB - Cat's claw is an herbal medicine from the Amazon that is used widely to treat inflammatory disorders. The purpose of this study was to characterize the antioxidative and antiinflammatory properties of cat's claw, Uncaria tomentosa (UT) and Uncaria guianensis (UG). Alkaloids and flavanols were determined using reversed-phase HPLC; scavenging of 1,1-diphenyl-2-picrilhydrazyl (DPPH), hydroxyl radicals, and lipid peroxidation by spectrophotometry; and TNFalpha production by ELISA. Anti-inflammatory activity was assessed in vitro by inhibition of TNFalpha and nitrite production from RAW 264.7 cells exposed to LPS (50 ng/ml) and in vivo using the indomethacin-induced gastritis model. Apoptosis was assessed using the TUNEL technique and TNFalpha mRNA by in situ RT-PCR. In each of the antioxidant assays tested, UG was more potent than UT (P < 0.01). The total oxindole and pentacyclic alkaloid content of UT was 35-fold > UG. The IC50 value for inhibition of TNFalpha production was significantly (P < 0.01) higher for UT (14.1 ng/ml) vs UG (9.5 ng/ml), yet at concentrations that were considerable lower than that required for antioxidant activity. Non-alkaloid HPLC fractions from UT decreased LPS-induced TNFalpha and nitrite production in RAW 264.7 cells (P < 0.01) at a concentration range comparable to the parent botanical. Oral pretreatment for 3 d with UT protected against indomethacin-induced gastritis, and prevented TNFalpha mRNA expression and apoptosis. These results indicate that while both species of cat's claw provide effective antioxidant and anti inflammatory activities, U. guianensis is more potent. In conclusion, the presence of oxindole or pentacyclic alkaloids did not influence the antioxidant and anti-inflammatory properties of cat's claw. PMID- 12120815 TI - Daidzein, coumestrol and zearalenone affect lipogenesis and lipolysis in rat adipocytes. AB - Daidzein, coumestrol and zearalenone - compounds called phytoestrogens, considered as active biological factors affecting many important physiological and biochemical processes appeared to be also significant regulators of adipocyte metabolism. In our experiments the influence of daidzein (0.01, 0.1 and 1 mM), coumestrol (0.001, 0.01 and 0.1 mM), zearalenone (0.01, 0.1 and 1 mM) and estradiol (0.01, 0.1 and 1 mM) on basal and insulin-stimulated (1 nM) lipogenesis from glucose and acetate was tested in adipocytes isolated from growing (160 +/- 5 g b.w) male Wistar rats. All tested compounds significantly attenuated glucose conversion to lipids. In the case of daidzein and coumestrol, this effect was probably due to inhibition of glycolysis. Daidzein (0.01, 0.1 and 1 mM), coumestrol (0.01 and 0.1 mM) and zearalenone (0.01, 0.1 and 1 mM) affected also basal and epinephrine-stimulated (1 microM) lipolysis. Daidzein (0.01 and 1 mM) augmented basal glycerides breakdown in adipocytes. The epinephrine-induced lipolysis was dependent on daidzein concentration and its stimulatory (0.1 mM) or inhibitory (1 mM) influence was observed. Zearalenone changed lipolysis only at the concentration of 1 mM and its effect was contradictory in the absence or presence of epinephrine (the stimulatory or inhibitory effect, respectively). Results obtained in experiments with inhibitors (insulin, 1 nM and H-89, 50 microM) and activators (dibutyryl-cAMP, 1 mM and forskolin, 1 microM) of lipolysis allowed us to assume that daidzein augmented basal lipolysis acting on PKA activity. The inhibitory effect of daidzein and zearalenone on epinephrine induced lipolysis is probably due to restriction of HSL action. The influence of coumestrol on glycerides breakdown was less marked. Estradiol augmented only epinephrine-stimulated lipolysis. PMID- 12120816 TI - Extract of Ocimum canum lowers blood glucose and facilitates insulin release by isolated pancreatic beta-islet cells. AB - Aqueous extract of Ocimum canum Sim, (Lamiaceae) is used by some Ghanaians to manage diabetes mellitus. In vivo modulation of levels of fasting blood glucose by 0. canum extract was evaluated in type-II diabetes mellitus using the C57BL/KsJ db/db genetically diabetic animal model, and its effects on glucose stimulated insulin release in vitro were monitored using isolated rat pancreatic beta-islet cells. The results showed that fasting blood glucose levels and body weight decreased significantly (p < 0.05) in diabetic and non-diabetic C57BL/KsJ mice, which were administered aqueous extract of 0. canum. In vitro, the 0. canum extract significantly enhanced insulin release from isolated rat pancreatic beta islet cells. Insulin release was found to be dependent on glucose concentration and increased with increasing O. canum concentration in the incubation medium up to an optimum extract concentration of 0.03 mg/ml. Release of the hormone decreased beyond this concentration of extract in the medium. Addition to the medium of Desmodium adscendens, a plant preparation used to manage inflammatory disorders, did not increase but rather inhibited insulin secretion by the pancreatic beta-islet cells. These results could explain the use of 0. canum in Ghanaian folk medicine to manage diabetes mellitus. PMID- 12120817 TI - Herbal medicinal products during pregnancy? PMID- 12120818 TI - Anti-inflammatory and antimicrobial activities of triterpenoids from Strobilanthes callosus nees. AB - The anti-inflammatory and antimicrobial activities of the 95% ethanol extract, benzene fraction and isolated triterpenoids of Strobilanthes callosus were investigated. In the carrageenan-induced paw edema inflammation model, the taraxerol showed a high reduction of edema, but the antimicrobial effect observed was lower at the two doses employed. These results confirm the use of this plant in folk medicine as an anti-inflammatory and antimicrobial herbal drug. PMID- 12120819 TI - Extrahepatic conditions and hepatic encephalopathy in elderly patients. AB - PURPOSE: Extrahepatic conditions can cause, exacerbate, or mimic hepatic encephalopathy in any patient with advanced liver disease, particularly in older persons. The aim of this study was to characterize the clinical features and frequency of extrahepatic conditions and the effect of therapeutic interventions upon the encephalopathy. DESIGN: Survey. SETTING: Inner city community hospital. METHODS: Retrospective chart review of 294 elderly patients (age 65-97) with liver disease and suspected hepatic encephalopathy, during a 15-year period, that included 188 men and 106 women. RESULTS: Extrahepatic conditions were found in 64 patients (22%); 29 (10%) patients had > 1 extrahepatic condition. Category and frequency of the extrahepatic conditions found in these 64 patients were as follows: urinary tract infection, 21 (33%); cellulitis/infected pressure ulcers, 16 (25%); pneumonia, 16 (25%); septicemia (with positive blood cultures), 10 (16%); silent myocardial infarction, 10 (16%); drug toxicity (nonsteroidal anti inflammatory drugs, sedatives, hypnotics, antidiabetics), 6 (9%); meningitis, 6 (9%); head injury, 5 (8%); stroke, 5 (8%); and subdural hematoma, 5 (8%). CONCLUSION: A significant proportion of elderly patients with liver disease and presumptive diagnosis of hepatic encephalopathy may have extrahepatic condition(s), and the treatment of the latter may improve clinical outcome of such patients. A high index of suspicion, low threshold of diagnostic measures, and prompt treatment of any associated extrahepatic condition are essential to prevent significant morbidity and mortality of these patients. PMID- 12120820 TI - Lower serum digoxin concentrations in heart failure and reassessment of laboratory report forms. AB - Serum digoxin concentrations (SDC) have been used clinically since the early 1970s. Whereas the therapeutic range for SDC is frequently cited as either 0.8 to 2.0 ng/mL or 0.5 to 2.0 ng/mL, studies over the past decade suggest an upper limit of 1.0 ng/mL for treating heart failure. The same upper limit for SDC is suggested for patients with heart failure and atrial fibrillation with rapid ventricular response. Reducing the upper limit of the therapeutic range to 1.0 ng/mL on computerized and paper laboratory report forms may guide clinicians to avoid unnecessarily high SDC, thus minimizing risk of digoxin toxicity without sacrificing therapeutic benefit for heart failure. PMID- 12120821 TI - Therapeutic applications of monoclonal antibodies. AB - Researchers have sought therapeutic applications for monoclonal antibodies since their development in 1975. However, murine-derived monoclonal antibodies may cause an immunogenic response in human patients, reducing their therapeutic efficacy. Chimeric and humanized antibodies have been developed that are less likely to provoke an immune reaction in human patients than are murine-derived antibodies. Antibody fragments, bispecific antibodies, and antibodies produced through the use of phage display systems and genetically modified plants and animals may aid researchers in developing new uses for monoclonal antibodies in the treatment of disease. Monoclonal antibodies may have a number of promising potential therapeutic applications in the treatment of asthma, autoimmune diseases, cancer, poisoning, septicemia, substance abuse, viral infections, and other diseases. PMID- 12120823 TI - Cocaine-associated chest pain. PMID- 12120822 TI - A 41-year-old man with fatigue, weight loss, hypercalcemia, and hepatosplenomegaly. PMID- 12120824 TI - Idiopathic intracranial hypertension with primary aldosteronism: report of 2 cases. AB - Although unconfirmed, the syndrome idiopathic intracranial hypertension (IIH), commonly seen in overweight 20- to 50-year-old women, has been proposed to have its origins in an endocrine-based disturbance of electrolytes. Herein we report on 2 women with IIH and primary aldosteronism (PAL). Aged 57 and 55 (patients 1 and 2), each had a longstanding history of mild-to-moderate arterial hypertension, recurrent hypokalemia, and headaches. They were found to have IIH at ages 51 and 45. PAL was diagnosed at ages 57 and 35, respectively, due to proven left adrenal adenoma in patient 1; and presumptive adrenal nodular hyperplasia in patient 2. This is the first report to appear in the English medical literature that describes an association between IIH and PAL. It raises the prospect that in some cases of IIH associated with arterial hypertension, an autonomous production of aldosterone should be considered. PMID- 12120825 TI - Changes in blood macrophage colony-stimulating factor levels after cesarean section in normotensive pregnancy and preeclampsia. AB - BACKGROUND: Macrophage colony-stimulating factor (M-CSF) stimulates the proliferation and differentiation of placental trophoblasts and may regulate trophoblast invasion into the placental bed. M-CSF levels in peripheral blood show a significant increase in preeclampsia. Thus, the present study examined changes in blood levels of M-CSF before and after cesarean section and compared them between normotensive and preeclamptic pregnant women. METHODS: Peripheral blood was collected before, 1 day after, and 10 days after cesarean section from 27 women, 12 of whom were preeclamptic pregnant patients with a mean blood pressure of 162/98 mm Hg and 15 were age- and gestational age-matched normotensive pregnant women (normotensive control subjects). Peripheral blood was also collected once from 15 age-matched healthy, normal cycling women (nonpregnant control subjects). M-CSF level was determined by the sandwich enzyme linked immunosorbent assay (ELISA) method using 3 antibodies. RESULTS: In normotensive and preeclamptic pregnancies, the M-CSF levels increased significantly (P < 0.01) 1 day after surgery but then decreased significantly (P < 0.01) at 10 days after surgery. Before and 1 day after surgery, the M-CSF levels were significantly higher (P < 0.01) in preeclamptic patients than in normotensive control subjects, but not at 10 days after surgery. CONCLUSIONS: The blood M-CSF levels were significantly higher in preeclampsia than in normotensive pregnancies, before cesarean section. The M-CSF levels in the circulation at 1 day after surgery increased significantly. The increase was about 270 U/mL net and at similar levels in 2 groups. Thus, increases in M-CSF levels after cesarean section may occur via similar mechanisms in normotensive and preeclamptic pregnancies. The M-CSF level in normotensive pregnancies and preeclampsia decreased and returned to the normal level at 10 days after cesarean section. PMID- 12120826 TI - Apparent failure of endocarditis prophylaxis caused by penicillin-resistant Streptococcus mitis. AB - Antibiotic resistance among viridans streptococci has increased with Streptococcus mitis being more resistant than other viridans species. In a case presented in this report, it is possible that antibiotic resistance contributed to an apparent failure of endocarditis prophylaxis. The patient had undergone periodontal surgery on 2 separate occasions and in both instances was administered 2 g of amoxicillin orally 1 hour before each procedure. He subsequently developed a subacute illness and had multiple blood cultures drawn that grew S. mitis with a minimum inhibitory concentration of 1.0 microg/mL for penicillin. Transesophageal echocardiogram provided further evidence of infective endocarditis with vegetations seen on the anterior leaflet of the mitral valve. Combination therapy with high-dose intravenous aqueous crystalline penicillin G and gentamicin sulfate for 4 weeks was curative. Clindamycin, rather than amoxicillin, has since been used as dental prophylaxis for subsequent procedures. PMID- 12120827 TI - Association between state level drinking and driving countermeasures and self reported alcohol impaired driving. AB - OBJECTIVES: In 1999, alcohol related motor vehicle crashes in the United States claimed 15786 lives and injured more than 300000 persons. Drinking and driving behavior is shaped by individual and environmental level influences. In this study, the association between each state's driving under the influence of alcohol (DUI) countermeasures and self reported alcohol impaired driving was explored. METHODS: Mothers Against Drunk Driving's (MADD's) Rating the States 2000 survey, which graded states on their DUI countermeasures from 1996-99, was used as an index of each state's comprehensive DUI prevention activities. Information on alcohol impaired driving from residents of each state was obtained from the 1997 Behavioral Risk Factor Surveillance System (BRFSS) survey. The association between the MADD state grades and alcohol impaired driving was assessed using multiple logistic regression. RESULTS: Of the 64162 BRFSS respondents who reported drinking any alcohol during the past month, 2.1% of women and 5.8% of men reported at least one episode of alcohol impaired driving in the past month. Those living in states with a MADD grade of "D" were 60% more likely to report alcohol impaired driving than those from states with a MADD grade of "A" (odds ratio 1.6, 95% confidence interval 1.3 to 2.1). The association existed for men and women. CONCLUSION: These findings suggest that stronger state level DUI countermeasures are associated with lower rates of self reported alcohol impaired driving. PMID- 12120828 TI - Personal and situational influences on drink driving and sober driving among a cohort of young adults. AB - OBJECTIVES: To compare personal and situational influences on incidents involving drink driving with those involving sober driving. METHODS: Information on a range of road safety practices was sought in face to face interviews conducted with 969 members of the Dunedin Multidisciplinary Health and Development Study cohort at age 26 years. A total of 750 study members reported an incident that involved the opportunity to consume alcohol and also travel by motor vehicle. Of these, 87 were classified as "drink drive incidents" and 663 as "sober drive incidents". RESULTS: Study members who were male, of lower socioeconomic status, had no school qualifications, or were dependent on alcohol or marijuana at age 21 were significantly more likely to report a drink drive incident at age 26. Compared with the sober drive incidents, the drink drive incidents were more commonly associated with driving alone, drinking at bars, and no advanced planning. For drink drive incidents the amount of alcohol consumed was influenced by the conviviality of the occasion, whereas for sober drive incidents it was the need to drive. One quarter of those reporting drink drive incidents stated they had used marijuana and/or LSD at the event at which they drank. CONCLUSIONS: Drink drive and sober drive incidents differed, particularly with regard to decisions made before the event. Prevention efforts could usefully be targeted toward these decisions. PMID- 12120829 TI - Older driver involvements in police reported crashes and fatal crashes: trends and projections. AB - OBJECTIVES: Older drivers have become a larger part of the driving population and will continue to do so as the baby boomers reach retirement age. The purpose of this study was to identify the potential effects of this population increase on highway safety. METHODS: Driver involvement rates for all police reported crashes were calculated per capita, per licensed driver, and per vehicle-mile of travel for 1990 and 1995. Also, driver involvement rates for fatal crashes were calculated for 1983, 1990, and 1995. Based on current crash rates per licensed driver and estimates of the future number of licensed drivers, projections of crashes involving drivers aged 65 and older were made for years 2010, 2020, and 2030. RESULTS: Driver crash involvement rates per capita decreased with age, but fatal involvement rates per capita increased starting at age 70. The same pattern existed for involvement rates per licensed driver. For both all crashes and fatal crashes, involvement rates per mile driven increased appreciably at age 70. Using projections of population growth, it was estimated that for all ages there will be a 34% increase in the number of drivers involved in police reported crashes and a 39% increase in the number involved in fatal crashes between 1999 and 2030. In contrast, among older drivers, police reported crash involvements are expected to increase by 178% and fatal involvements may increase by 155% by 2030. Drivers aged 65 and older will account for more than half of the total increase in fatal crashes and about 40% of the expected increase in all crash involvements; they are expected to account for as much as 25% of total driver fatalities in 2030, compared with 14% presently. CONCLUSIONS: By most measures, older drivers are at less risk of being involved in police reported crashes but at higher risk of being in fatal crashes. Although any projections of future crash counts have inherent uncertainty, there is strong evidence that older drivers will make up a substantially larger proportion of drivers involved in fatal crashes by 2030 because of future increases in the proportion of the population aged 65 and older, and trends toward increased licensure rates and higher annual mileage among older persons. Countermeasures to reduce the anticipated death toll among older drivers should address the increased susceptibility to injury of older vehicle occupants in crashes. PMID- 12120830 TI - The nature of newspaper coverage of homicide. AB - OBJECTIVES: Previous research has shown that some homicides are more likely than others to receive newspaper coverage (for example, homicides by strangers). The present investigation examined whether, once the decision has been made to report on a homicide, the nature of the coverage (that is, how much visibility is given to a story, what information is included, and how a story is written) differs according to two key variables, victim ethnicity, and victim-suspect relationship. SETTING: Los Angeles, California (USA). METHODS: Homicide articles from the 1990-94 issues of the Los Angeles Times were stratified according to the predictors of interest (victim ethnicity and victim-suspect relationship) and a sample was drawn. Data that characterized two primary aspects of newspaper coverage, prominence and story framing (including background information, story focus, use of opinions, story tone, and "hook" or leading introductory lines) were abstracted from the articles. Descriptive statistics and cross tabulations were generated. Multivariate analyses were conducted to examine the predictive value of victim ethnicity and victim-suspect relationship on the nature of the newspaper coverage. RESULTS: Newspaper coverage of homicide was generally factual, episodic, and unemotional in tone. Victim-suspect relationship, but not victim ethnicity, was related to how a story was covered, particularly the story frame. Homicides by intimates were covered consistently differently from other types of homicides; these stories were less likely to be opinion dominated, be emotional, and begin with a "hook". CONCLUSION: Victim-suspect relationship was related to the nature of coverage of homicides in a large, metropolitan newspaper. Given the agenda setting and issue framing functions of the news media, these findings have implications for the manner in which the public and policy makers perceive homicides and, consequently, for the support afforded to various types of solutions for addressing and preventing violence. PMID- 12120831 TI - Residential fire related deaths and injuries among children: fireplay, smoke alarms, and prevention. AB - BACKGROUND: The aim of the study was to describe the epidemiology of residential fire related deaths and injuries among children, and identify risk factors for these injuries through a linked dataset for the city of Dallas, Texas. METHODS: Data for all residential fires were linked with fire related injury data, using fire department records, ambulance transports, hospital admissions, and medical examiner records, for children 0-19 years of age. Causes of fires, including fireplay (children playing with fire or combustibles), arson and other causes, were determined by fire department investigation. RESULTS: From 1991-98, 76 children were injured in residential fires (39 deaths, 37 non-fatal). The highest rates occurred in the youngest children (<5 years) and in census tracts with lowest income. Fireplay accounted for 42% (32/76) of all injuries, 62% (15/24) of deaths in children 0-4 years, and 94% (13/14) of deaths from apartment and mobile home fires. Most of the fireplay related injuries (27/32, 84%) were from children playing with matches or lighters. Most started in a bedroom. Smoke alarms showed no protective efficacy in preventing deaths or injuries in fires started by fireplay or arson, but there was significant protective efficacy for a functional smoke alarm in fires started from all other causes (p<0.01). CONCLUSIONS: Residential fire related injuries among children in Dallas occurred predominantly in the youngest ages (<5 years) and in poor neighborhoods. Most of the deaths, especially those in apartments and mobile homes, resulted from fireplay. Smoke alarms appeared to offer no protection against death or injury in fireplay associated fires, possibly from the nature of the child's behavior in these fires, or from the placement of the smoke alarm. Prevention of childhood residential fire related deaths may require interventions to prevent fireplay in order to be successful. PMID- 12120832 TI - Citywide trauma experience in Kampala, Uganda: a call for intervention. AB - OBJECTIVES: To describe injuries and their emergency care at five city hospitals. SETTING: Data were collected between January and December 1998 from casualty departments of the five largest hospitals of Kampala city, Uganda, with bed capacity ranging from 60 to 1200. METHODS: Registry forms were completed on trauma patients. All patients with injuries were eligible. Outcome at two weeks was determined for admitted patients. RESULTS: Of the 4359 injury patients, 73% were males. Their mean age was 24.2 years, range 0.1-89, and a 5-95 centile of 5 50 years. Patients with injuries were 7% of all patients seen. Traffic crashes caused 50% of injuries, and were the leading cause for patients > or = 10 years. Fifty eight per cent of injuries occurred on the road, 29% at home, and 4% in a public building. Falls, assaults, and burns were the main causes in homes. Fourteen per cent of injuries were intentional. Injuries were severe in 24% as determined with the Kampala trauma score. One third of patients were admitted; two thirds arrived at the hospital within 30 minutes of injury, and 92% were attended within 20 minutes of arrival. CONCLUSIONS: Injuries in Kampala are an important public health problem, predominantly in young adult males, mostly due to traffic. The majority of injuries are unintentional. Hospital response is rapid, but the majority of injuries are minor. Without pre-hospital care, it is likely that patients with serious injuries die before they access care. Preventive measures and a pre-hospital emergency service are urgently needed. PMID- 12120833 TI - Socioeconomic differences in injury risks in childhood and adolescence: a nation wide study of intentional and unintentional injuries in Sweden. AB - STUDY OBJECTIVE: To measure socioeconomic differences in injuries among different age groups of children and adolescents. SUBJECTS: Children under 20 living in Sweden between 1990 and 1994 (about 2.6 million). METHOD: A cross sectional study based on record linkage between 15 Swedish national registers. Children were divided into four age groups and allocated to four household socioeconomic status groups. Absolute and relative risks were compiled using children of high/intermediate level salaried employees as the comparison group. Four diagnostic groups were considered: fall, traffic, interpersonal violence, and self inflicted injuries. RESULTS: Injury incidences were relatively low and socioeconomic differences negligible in the 0-4 year olds. Thereafter, significant socioeconomic differences were observed in all diagnostic groups except falls. The highest absolute differences were in traffic injuries, especially among 15-19 year olds, and in self inflicted injuries among 15-19 year old girls. Relative differences were highest in both categories of intentional injuries for the age group 10-14. Social circumstances in the household other than family socioeconomic status affected the social pattern of intentional but not that of unintentional injuries. CONCLUSIONS: Socioeconomic differences in injury risks are not necessarily constant over age. Inequalities are particularly high in absolute terms among adolescents 15-19 years old for traffic injuries and in relative terms among 10-14 year olds for intentional injuries. PMID- 12120834 TI - Missing the target: a comparison of buyback and fatality related guns. AB - OBJECTIVES: To determine whether the firearms recovered in buyback programs in a large urban community are the types most closely associated with firearm fatalities in the same geographic area. METHODS: The type, caliber, and manufacturer of 941 handguns recovered in Milwaukee County 1994-96 buyback programs were compared with 369 homicide related and 125 suicide related handguns used in Milwaukee during 1994-97. RESULTS: Buyback handguns differed substantially from those used in homicide and suicide. One third of buyback handguns were semiautomatic pistols versus two thirds of homicide related handguns (p<0.001) and 40% of suicide related handguns (p=NS). Over 75% of buyback handguns were small caliber compared with 24% of homicide and 32% of suicide handguns (p<0.001). The top two manufacturers of buyback handguns represented 30% of these guns but only 5% of fatality related handguns (p<0.001). Companies currently out of business manufactured 15% of buyback handguns versus 7% of fatality related handguns (p<0.001). CONCLUSIONS: Handguns recovered in buyback programs are not the types most commonly linked to firearm homicides and suicides. Although buyback programs may increase awareness of firearm violence, limited resources for firearm injury prevention may be better spent in other ways. PMID- 12120835 TI - Risk factors associated with non-fatal adolescent firearm injuries. AB - STUDY OBJECTIVES: To identify behavioral, environmental, and sociodemographic risk factors associated with non-fatal firearm injuries among inner city adolescents in the United States. DESIGN: A case-control study in which patients with firearm injury serve as cases and those with medical conditions serve as controls. SETTING: A level I trauma center in a metropolitan area serving a predominately lower socioeconomic status population. PARTICIPANTS: Cases were 45 consecutive patients 11-18 years presenting to the emergency department with non fatal firearm injury; controls were 50 age and gender matched patients presenting with acute medical problems. OUTCOME MEASURE: Odds ratios (OR) and associated 95% confidence interval (CI) as estimates of the magnitude of association between risk factors and non-fatal firearm injury. RESULTS: After adjusting for age, gender and socioeconomic status, multivariate analysis identified four risk factors independently associated with firearm injury: living with less than two parents (OR 3.8, 95% CI 1.2 to 12.2), skipping class (OR 7.1, 95% CI 1.7 to 28.9), previous arrest (OR 6.2, 95% CI 1.9 to 20.7), and being African-American (OR 4.2; 95%CI 1.4 to 14.9). CONCLUSION: Risk factors for adolescents sustaining a non-fatal firearm injury are sociodemographic and environmental, not just behavioral. Thus interventions that foster protective and supportive environments may help prevent firearm injuries. PMID- 12120836 TI - Safety attitudes and beliefs of junior Australian football players. AB - OBJECTIVES: To describe the safety attitudes and beliefs of junior (aged 16-18 years) Australian football players. SETTING: Six Victorian Football League Under 18 (VFL U18) clubs in Victoria, Australia. METHODS: Cross sectional survey. Altogether 103 players completed a self report questionnaire about their safety beliefs and perceptions of support when injured, across three contexts in which they played: VFL U18 club, local club, and school. RESULTS: Although only 6% believed it was safe to play with injuries, 58% were willing to risk doing so. This increased to almost 80% when players perceived that their chances of being selected to play for a senior elite team would be adversely affected if they did not play. There were significant differences in the perceived level of support for injured players and in the ranking of safety as a high priority across the three settings. In general, the VFL U18 clubs were perceived as providing good support for injured players and giving a high priority to safety issues, but local clubs and particularly schools were perceived to address these issues less well. CONCLUSIONS: Junior Australian football players have certain beliefs and perceptions in relation to injury risk that have the potential to increase injuries. These negative beliefs need to be addressed in any comprehensive injury prevention strategy aimed at these players. PMID- 12120838 TI - Research on injury prevention: topics for systematic review. AB - BACKGROUND: Duplication should be avoided in research and only effective intervention programs should be implemented. OBJECTIVE: To arrive at a consensus among injury control investigators and practitioners on the most important research questions for systematic review in the area of injury prevention. DESIGN: Delphi survey. METHODS: A total of 34 injury prevention experts were asked to submit questions for systematic review. These were then collated; experts then ranked these on importance and availability of research. RESULTS: Twenty one experts generated 79 questions. The prevention areas with the most number of questions generated were fires and burns, motor vehicle, and violence (other than intimate partner), and the least were other interventions (which included Safe Communities), and risk compensation. These were ranked by mean score. There was good agreement between the mean score and the proportion of experts rating questions as important or very important. Nine of the top 24 questions were rated as having some to a substantial amount of research available, and 15 as having little research available. CONCLUSIONS: The Delphi technique provided a useful means to develop consensus on injury prevention research needs and questions for systematic review. PMID- 12120837 TI - Are we blind to injuries in the visually impaired? A review of the literature. AB - OBJECTIVES: To review the literature on the risks and types of injuries associated with visual impairment, and to identify pertinent areas for future research. METHODS: A search of bibliographic databases was conducted in April 2000 for studies published since 1980 and selected studies that met two or more of the following criteria: formal ophthalmic assessment was used; adjustment for confounding variables; large sample size including numbers of visually impaired; and clear definitions and outcomes. RESULTS: Thirty one studies were selected. The majority of these studies (20) assessed falls (including eight on hip fracture and four on multiple falls), eight studies reported traffic related injuries, and three studies assessed occupational injury. The evidence on falls, which relate predominantly to older people, suggests that those with reduced visual acuity are 1.7 times more likely to have a fall and 1.9 times more likely to have multiple falls compared with fully sighted populations. The odds of a hip fracture are between 1.3 and 1.9 times greater for those with reduced visual acuity. Studies of less severe injuries and other causes of injury were either poorly designed, underpowered, or did not exist. CONCLUSIONS: There are substantial gaps in research on both injuries to which people with visual impairment are especially susceptible and in evaluating interventions to reduce these injuries. It is recommended that in future studies the minimum data captured includes: formal ophthalmic assessment of visual fields and visual acuity, outcome measurement, control for confounders, and the costs of health care resource use and any interventions. PMID- 12120839 TI - Child bicyclist injuries: are we obtaining enough information in the emergency department chart? AB - OBJECTIVE: The purpose of this study was to assess the range of information relevant to bicyclist injury research that is available on routinely completed emergency department medical records. METHODS: A retrospective chart review of emergency department medical records was conducted on children who were injured as bicyclists and treated at an urban level I pediatric trauma center. A range of variables relevant to bicyclist injury research and prevention was developed and organized according to the Haddon matrix. Routinely completed free text emergency department medical records were assessed for the presence of each of the targeted elements. In addition, medical records of seriously injured patients (for whom a more structured medical record is routinely used) were compared to free form records of less seriously injured patients to identify differences in documentation that may be related to the structure of the medical record. RESULTS: Information related to previous medical history (96% of records), diagnosis (89%), documentation of pre-hospital care (82%), and child traumatic contact points (81%) were documented in the majority of medical records. Information relevant to prevention efforts was less commonly documented: identification of motor vehicle/object involved in crash (58%), the precipitating event (24%), the location of the crash (23%), and documentation of helmet use (23%). Records of seriously injured patients demonstrated significantly higher documentation rates for pre-hospital care and child traumatic contact points, and significantly lower documentation rates for previous medical history, child kinematics, main body parts impacted, and location of injury event. CONCLUSIONS: Routinely completed free text emergency department medical records contain limited information that could be used by injury researchers in effective surveillance. In particular information relating to the circumstances of the crash event that might be used to design or target prevention efforts is typically lacking. Routine use of more structured medical records has the potential to improve documentation of key information. PMID- 12120840 TI - Safe hot tap water and the risk of scalds and legionella infection. PMID- 12120841 TI - Regulations, legislation, and classification. PMID- 12120842 TI - An introduction to the Barell body region by nature of injury diagnosis matrix. AB - INTRODUCTION: The Barell body region by nature of injury diagnosis matrix standardizes data selection and reports, using a two dimensional array (matrix) that includes all International Classification of Diseases (ICD)-9-CM codes describing trauma. AIM: To provide a standard format for reports from trauma registries, hospital discharge data systems, emergency department data systems, or other sources of non-fatal injury data. This tool could also be used to characterize the patterns of injury using a manageable number of clinically meaningful diagnostic categories and to serve as a standard for casemix comparison across time and place. CONCEPT: The matrix displays 12 nature of injury columns and 36 body region rows placing each ICD-9-CM code in the range from 800 to 995 in a unique cell location in the matrix. Each cell includes the codes associated with a given injury. The matrix rows and columns can easily be collapsed to get broader groupings or expanded if more specific sites are required. The current matrix offers three standard levels of detail through predefined collapsing of body regions from 36 rows to nine rows to five rows. MATRIX DEVELOPMENT: This paper presents stages in the development and the major concepts and properties of the matrix, using data from the Israeli national trauma registry, and from the US National Hospital Discharge Survey. The matrix introduces new ideas such as the separation of traumatic brain injury (TBI), into three types. Injuries to the eye have been separated from other facial injuries. Other head injuries such as open wounds and burns were categorized separately. Injuries to the spinal cord and spinal column were also separated as are the abdomen and pelvis. Extremities have been divided into upper and lower with a further subdivision into more specific regions. Hip fractures were separated from other lower extremity fractures. FORTHCOMING DEVELOPMENTS: The matrix will be used for the development of standard methods for the analysis of multiple injuries and the creation of patient injury profiles. To meet the growing use of ICD-10 and to be applicable to a wider range of countries, the matrix will be translated to ICD-10 and eventually to ICD-10-CM. CONCLUSION: The Barell injury diagnosis matrix has the potential to serve as a basic tool in epidemiological and clinical analyses of injury data. PMID- 12120844 TI - Health literacy: new wine in old bottles? PMID- 12120843 TI - UNICEF's child injury league table. An analysis of legislation: more mixed messages. AB - This paper presents a summary table and discussion of legislation related to child injury prevention in member countries of the Organisation for Economic Cooperation and Development. The table is an expanded version of the one which appeared in the UNICEF Report Card "Child Deaths by Injury in Rich Countries" (2001). A commentary is provided on the variations in legislation between countries in terms of range and form of measures and an estimate of degree of enforcement. As legislation is generally considered a powerful tool in injury prevention, the paper examines whether those countries with the widest range of legislation and the strongest enforcement have made the most progress in reducing child injury deaths since the 1970s. It also considers whether a commitment to extensive legislation is reflected in a country's position in the UNICEF league table of injury death. The initial conclusion to these two basic issues is that no clear picture can be seen and we thus need to know far more about the relationship between legislation and societies and cultures as they vary from place to place. This paper hopes to stimulate more widespread debate about the role of legislation in different countries. PMID- 12120845 TI - Patient information on phantom limb pain: a focus group study of patient experiences, perceptions and opinions. AB - Educating patients about their condition is regarded as a fundamental step in pain management. This study used focus groups with patients to explore their experiences and perceptions of the information on phantom pain that they received before and after amputation, and their views on improving this information. Thirty-one patients with a lower limb amputation attended one of seven focus groups. The majority reported phantom pain although there were individual variations in character, severity and persistence. There were wide variations in what people were told from occasional reports of good information to instances of people reporting little or no information from professionals. There were strong feelings that information should be given before or soon after amputation with a preference for verbal one-to-one explanations. Professionals, particularly nurses and surgeons, were regarded as the best source of information, although peer support was seen to be important. These findings indicate that people require timely up-to-date information on phantom pain which sensitively addresses the variability of the experience and provides the foundation for ongoing pain management. We propose that the information process could be improved by ensuring that professionals use standard information for patients derived from purposefully written sections in national guidelines. PMID- 12120846 TI - Psychosocial correlates of sun-protective practices of preschool staff toward their students. AB - The skin cancer rate in the US has been increasing faster than that for other cancers. Most skin cancers are related to sun exposure and the majority of exposure occurs before adulthood. Thus, children are an important target group to study and preschools can be useful avenues for delivering sun-protection messages. The current study examines the behaviors of preschool staff in protecting students from sun exposure and investigates factors related to sun protective practice. Preschool staff (n = 245) were surveyed about their sun protective practices toward students as the cross-sectional baseline measurement for a larger project. The primary aim of this study was to investigate correlates of staff's sun-protective behavior toward students. A theoretical model of psychosocial constructs that combined components of the Theory of Planned Behavior and Social Cognitive Theory was evaluated using structural equation modeling. Self-efficacy and perceived norms were the strongest correlates of behavior. A hypothesized link between expectancy and behavior was not supported. The roles of self-efficacy and perceived norms in the preschool context are discussed as they relate to staffs behavior. PMID- 12120847 TI - Measuring perceptions of innovation adoption: the diffusion of a federal drug prevention policy. AB - The purpose of this paper is to describe the testing of a new scale to assess the perceived attributes of a federal drug prevention policy. The 17-item scale was administered to 107 Safe and Drug Free Schools (SDFS) coordinators in 12 states as a part of a larger investigation examining the diffusion of a federal drug prevention policy. In developing this scale, the authors drew from theory, previously validated measures, expert review and pre-testing with SDFS coordinators. Factor analysis revealed three underlying constructs representing relative advantage/compatibility, complexity and observability. The constructs found were internally consistent with a Cronbach's alpha ranging from a high of 0.89 for relative advantage/compatibility to a low of 0.71 for observability. Each of these constructs was correlated with a district's adoption of the policy in predictable ways. The construct of relative advantage/compatibility appears to be especially useful in assessing policy adoption. This scale was developed to assess a specific innovation; however, we believe that it can be easily adapted to understand the adoption of other health education interventions. PMID- 12120848 TI - Peer-led sex education--characteristics of peer educators and their perceptions of the impact on them of participation in a peer education programme. AB - The RIPPLE study is a randomized controlled trial of peer-led sex education in English secondary schools. In 1997, 27 schools were recruited and randomly allocated to a programme of peer-led sex education or to act as control schools. In experimental schools peer educators in Year 12 (aged 16/17 years) were recruited in two successive cohorts and, having received a standardized training programme, delivered classroom-based sex education sessions to Year 9 students (aged 13/14 years). This paper is the first of two focusing on data gathered from these peer educators. Through analysis of pre-(n = 505) and post- (n = 331) programme questionnaire data, the paper describes the profile of peer educators and examines the impact on them of their involvement. Compared to the students receiving the peer-led sex education, more peer educators were female, white, high academic achievers and less socially disadvantaged. Peer educators reported positive changes in sexual knowledge and changes towards more liberal attitudes, and believed the programme would have a positive impact on their confidence in relationships and on their sexual behaviour. There was an increase in confidence about communication and interaction in groups. The paper discusses the methodological difficulties of assessing how involvement in such a programme impacts on peer educators. PMID- 12120849 TI - What influences peer-led sex education in the classroom? A view from the peer educators. AB - This paper is the second of two presenting data gathered from peer educators in the RIPPLE study-a randomized controlled trial of peer-led sex education in English secondary schools. Peer educators were recruited from Year 12 students (aged 16/17 years) in 13 schools in two successive cohorts in 1997 and 1998. Following a standardized training programme they delivered sex education sessions to Year 9 students (aged 13/14 years). Through analysis of 18 focus group discussions and of post-programme questionnaire data (n = 301), this paper aims to identify the issues and processes considered by peer educators to be important in implementing a peer education programme, and to examine peer educators' views on the relationship between themselves and the Year 9 students. Methodological issues arising when collecting, analysing and presenting such data are discussed, and some recommendations are outlined for carrying out school-based peer education. PMID- 12120850 TI - The challenge of evaluating complex interventions: a framework for evaluating media advocacy. AB - New health promotion and public health approaches such as media advocacy pose particular evaluation challenges. Evaluation is important to provide feedback to media advocacy practitioners on how to enhance their efforts, and to funders and researchers seeking to assess media advocacy's effectiveness as a health promotion strategy. The media advocacy evaluation literature contains some examples of promising evaluation approaches but is still evolving. A comprehensive framework for the evaluation of media advocacy is presented. Building on existing approaches to evaluation in media advocacy and on current thinking regarding evaluation in health promotion, it proposes a series of indicators and research methods for evaluating media advocacy at the levels of formative, process and outcome evaluation. The framework can be used to encourage strategic reflection on the media advocacy process, to guide evaluation of specific interventions, and to demonstrate to funders the importance and complexity of evaluation in this promising field. PMID- 12120852 TI - Access to information and the pursuit of equity. PMID- 12120851 TI - A health intervention programme for children with albinism at a special school in South Africa. AB - The genetic condition albinism has a high frequency among the Sotho people of northern South Africa. Affected children have pale hair, eyes and skin-a dramatic contrast to the normal dark pigmentation. Their visual performance is poor and many attend special schools for the visually impaired. Children with albinism experience problems that are, on the one hand, physiological, and, on the other, social-psychological and educational in nature. In this self-report study 38 children at a rural special school described their eye and skin problems, a direct result of their lack of pigmentation, as well as strategies they adopted to manage their condition. A further section of the study deals with the social adaptation difficulties experienced by these children. The questionnaire tested for local belief systems about albinism and how these impact on the socialization of children with albinism. The intervention strategy proposed in this study is based on the assumption that any attempt to address both the health and social problems should be of a holistic, interactionist nature, and be based on the values and belief systems of the local community. In addressing the physical problems, the proposed intervention programme focuses on sensible sun protection habits from a young age and the active participation of the children. To alleviate the social problems a team (interactionist) approach including children, teachers, parents, health officials and the wider community is recommended. PMID- 12120853 TI - Annotation: The 'effects' of parenting reconsidered: findings, challenges, and applications. AB - BACKGROUND: Questions remain concerning the 'effects' of parenting on behavioural/emotional problems in children. This annotation discusses recent findings concerning the parenting 'effects' literature and identifies areas in need of further research. METHOD: The review begins by examining theories and definitions of parenting, and then considers research findings on the predictors of parent-child relationships and their effects on behavioural/emotional adjustment in children. Evidence for causal processes are then examined in light of findings emphasizing the need to consider the impact of larger systems on child's well-being, bi-directional processes in parent-child interactions, and alternative hypotheses suggested by behavioural genetics. RESULTS: Different kinds of evidence suggest strong links between parent-child relationship quality and children's well-being, but difficulties remain for drawing causal connections. The need for greater integration among research traditions and the need for theory development are highlighted. In addition, although a substantial and robust research base exists on parent-child relationships, the applicability of these findings to clinical settings is uncertain. CONCLUSIONS: Substantial progress has been made in our understanding of the nature of parent-child relationships and their developmental effects, but a number of basic conceptual and methodological and clinical questions continue to need rigorous study. PMID- 12120854 TI - Practitioner review: The effectiveness of systemic family therapy for children and adolescents. AB - BACKGROUND: Systemic family therapy has become a widely used intervention in child and adolescent mental health services over the last twenty years. METHODS: This paper reviews the development of systemic family therapy, briefly describes the theory and techniques associated with the most prominent contemporary strands of systemic practice, and examines the empirical justification for using systemic family therapies with children and adolescents. RESULTS: There is a paucity of well-designed randomised controlled trials of systemic therapies with children and adolescents and those trials that do exist evaluate older structural and strategic therapies. Methodological limitations of existing research include the use of unrepresentative participants, small sample sizes and wide age ranges. There is a lack of credible no-treatment or alternative treatment controls, tests of clinical as opposed to statistical significance, and conceptually relevant outcome measures that examine underlying interactional mechanisms. The term 'family therapy' encompasses a wide range of interventions and it is not always clear what treatment intervention has been delivered. Nevertheless, there is good evidence for the effectiveness of systemic family therapies in the treatment of conduct disorders, substance misuse and eating disorders, and some support for their use as second-line treatments in depression and chronic illness. CONCLUSIONS: Systemic family therapy is an effective intervention for children and adolescents but further well-designed outcome studies are needed using clearly specified, manualised forms of treatment and conceptually relevant outcome measures. PMID- 12120855 TI - Parental expressed emotion in depressed adolescents: prediction of clinical course and relationship to comorbid disorders and social functioning. AB - BACKGROUND: High expressed emotion (EE) predicts worse clinical course for a number of disorders. High EE is more frequent in parents of disordered children than normal controls. It is uncertain whether EE and its components are disorder specific, whether EE is more closely related to parent characteristics or child characteristics, and whether EE predicts clinical course independently of clinical variables that reflect severity of disorder. EE has not been investigated in adolescent depression. METHOD: The 57 participants in this study were a sub-sample of a longitudinal study of the clinical course of depression. Adolescents and parents were recruited from consecutive referrals to all psychiatric outpatient clinics and inpatient units in a geographic catchment area. The association between EE and one-year clinical outcome of major depressive disorder was tested and associations between EE and characteristics of the adolescent, the parent, and the family were examined. RESULTS: EE was independent of socio-demographic characteristics, comorbid diagnoses, and parental depression. High EE was associated with worse adolescent social functioning according to either adolescent or parent report. High EE was associated with the presence of more depression symptoms. Low EE predicted major depression remission in participants without comorbid attention deficit/hyperactivity disorder (ADHD), but this association was not independent of the association between social functioning and depression remission. CONCLUSIONS: The findings indicate a need to examine possible protective effects of low EE. Relationships between EE, social functioning, and depression persistence and remission require further examination. PMID- 12120856 TI - Do parent and child behaviours differentiate families whose children have obsessive-compulsive disorder from other clinic and non-clinic families? AB - BACKGROUND: Limited research has been conducted investigating parent and child behaviour during family interactions in families who have a child with obsessive compulsive disorder (OCD). While a number of authors in the field of childhood OCD have suggested possible parent and child behaviours that are characteristic of these families, few studies have attempted to explore these in a methodologically sound approach. METHOD: This study compared the observed behaviours of parents and children in families whose children were diagnosed with OCD, to families whose children were diagnosed with other anxiety disorders, externalising disorders and no clinical problems. During family discussions, parent and child behaviours and affect were coded using a Likert-scale system. Family members were rated on behavioural dimensions of control, warmth, doubt, avoidance, confidence, positive problem solving, and rewarding independence. RESULTS: Results indicated that parents and children in the OCD group could be clearly differentiated from families in the other groups based on parent and child behaviour. Mothers and fathers of OCD children were less confident in their child's ability, less rewarding of independence, and less likely to use positive problem solving. Children in the OCD group showed less positive problem solving, less confidence in their ability to solve the problem, and they displayed less warmth during their interactions with their parents. CONCLUSIONS: Parents and children in families where there is a child with OCD behave in a different manner during family interactions to other families. These findings offer interesting and important exploratory information relating to observed parent and child behaviour across different clinical and non-clinical groups. Limitations of this study are addressed and directions for further research are discussed. PMID- 12120857 TI - The development of impulsivity, fearfulness, and helpfulness during childhood: patterns of consistency and change in the trajectories of boys and girls. AB - BACKGROUND: The objective of the present study was to describe the development of boys and girls during the elementary-school years on three dimensions that conceptually and empirically represent risk for maladjustment. METHOD: Every year between kindergarten and grade six, teachers rated the impulsivity, fearfulness, and helpfulness dimensions among a sample of 1,865 children representative of kindergarten boys and girls in the province of Quebec (Canada) in 1986-87. A group-based trajectory method was used to 1) identify groups of boys and girls following distinct-level trajectories of behaviours (on each dimension) during the elementary-school years; 2) estimate the proportion of children in each of the identified trajectory groups; and 3) estimate the patterns of consistency and variations in trajectories. RESULTS: The results indicated that the best models comprised three distinct-level trajectory groups on fearfulness and helpfulness (a low, moderate, and high group) and four distinct-level trajectory groups on impulsivity. The helpfulness and fearfulness trajectory groups were generally more stable than the impulsivity groups. The broad patterns of development were similar across sexes. However, there were more boys on the higher impulsivity trajectories and low helpfulness trajectory, while there were more girls on the high fearfulness trajectory. CONCLUSION: We found that behavioural consistency over middle childhood varied across trajectory groups and across dimensions, and we identified sex differences in the distribution of children in the different trajectory groups that may reflect gender-specific risks for psychopathology. PMID- 12120858 TI - Associations between behaviour problems and verbal and nonverbal cognitive abilities and disabilities in early childhood. AB - BACKGROUND: We investigated associations between behaviour problems and verbal and nonverbal cognitive abilities at 2, 3 and 4 years of age both for the entire distribution and for the lowest 5% and 10% of the verbal and nonverbal cognitive disabilities. METHODS: A community sample of 4,000 pairs of twins born in England and Wales in 1994 and 1995 was assessed by their parents at 2, 3 and 4 years using the Revised Rutter Parent Scale for Preschool Children (RRPSPC, behaviour problems), the MacArthur Communicative Development Inventory (MCDI, verbal development), and the Parent Report of Children's Abilities (PARCA, nonverbal cognitive development). RESULTS: For the entire sample, behaviour problem scores were modestly associated with lower MCDI and PARCA scores - correlations were less than .30. Similarly modest effect sizes were found for relationships between behaviour problem scores and the lowest 5% and 10% of the MCDI and of the PARCA distributions. Associations were stronger for nonverbal than for verbal development, increased from 2 to 3 to 4 years, and, at the extremes of the distributions, were stronger for boys than for girls. Multivariate genetic analyses indicated that both genetic and shared environmental factors mediate the links between behaviour problems and cognitive development both for the total distribution and for the extremes. Genetic links may be stronger for the extremes than for the total sample. CONCLUSIONS: We conclude that, in this community sample of young children, associations between behaviour problems and verbal and nonverbal cognitive development are generally modest for the entire distribution and are no greater at the extremes than expected on the basis of the associations for the entire distribution. PMID- 12120859 TI - Young adult academic outcomes in a longitudinal sample of early identified language impaired and control children. AB - BACKGROUND: The long-term academic consequences of childhood language impairment are both theoretically and clinically important. An unbiased appraisal of these outcomes, however, requires carefully designed, longitudinal research. METHOD: A group of children first identified as having speech and/or language impairment in a community-based, longitudinal study at 5 years of age and matched controls were re-examined during young adulthood (age 19). A comprehensive battery of speech and language, cognitive and achievement tests, psychiatric interviews, and questionnaires were completed by subjects, their parents and teachers. RESULTS: While children with early speech problems showed only a few academic differences from controls in young adulthood, early language impaired (LI) young adults lagged significantly behind controls in all areas of academic achievement, even after controlling for intelligence. Further, rates of learning disabilities (LD) were significantly higher in the LI group than both the controls and community base rates. Concurrent individual difference variables, including phonological awareness, naming speed for digits, non-verbal IQ, verbal working memory, and executive function, all contributed unique variance to achievement in specific areas. CONCLUSION: Early LI rather than speech impairment is clearly associated with continued academic difficulties into adulthood. These results speak to the need for intensive, early intervention for LI youngsters. PMID- 12120860 TI - Shape constancy in autism: the role of prior knowledge and perspective cues. AB - BACKGROUND: Evidence suggests that individuals with autism may not attend to contextual information (conceptual or perceptual) when processing stimuli (Frith 1989; Shah & Frith, 1983). METHOD: We investigated the role of prior knowledge and perspective cues when judging the shape of a slanted circle in individuals with and without autism. Individuals adjusted a shape on a computer screen to appear the same as a slanted circle. RESULTS: Participants in all groups (autistic, moderate learning difficulties, children aged 9 years and adults) exaggerated circularity. Strikingly, however, individuals with autism were unique in exaggerating circularity significantly far less when perspective cues surrounding the slanted circle were eliminated. Prior knowledge that the shape was a slanted circle provoked a strong exaggeration effect in participants without autism, but not in those with autism. CONCLUSIONS: Perhaps classifying the stimulus as a 'circle' was sufficient to provoke a strong exaggeration effect in those without (but not with) autism. In this domain, we show that perception in autism may be less influenced by prior knowledge, and therefore less 'top down'. PMID- 12120861 TI - The severity and nature of motor impairment in Asperger's syndrome: a comparison with specific developmental disorder of motor function. AB - BACKGROUND: The aims of this study were to measure objectively the extent and severity of motor impairment in children with Asperger's syndrome and to determine whether the motor difficulties experienced by such children differed in any way from those classified as having a Specific Developmental Disorder of Motor Function (SDD-MF). Criteria derived from ICD 10-R were used to identify 11 children with Asperger's syndrome and a matched group of 9 children with a Specific Developmental Disorder of Motor Function. Children in both groups were required to have a verbal IQ of 80 or greater on the WISC IIIR. METHOD: The Autism Diagnostic Interview (Revised; Lord, Rutter, & LeCouteur, 1994) was used to identify features of AS in the first group and to exclude them in the latter. The Movement Assessment Battery for Children (Henderson & Sugden, 1992) provided a standardised test of motor impairment. A Gesture Test based on that by Cermak, Coster, and Drake (1980) was used to assess the child's ability to mime the use of familiar tools and to imitate meaningless sequences of movements. RESULTS: All the children with Asperger's syndrome turned out to meet our criterion for a diagnosis of motor impairment, five of the six most severely motor impaired children in the whole study being from this group. Performance of the Asperger group was also slightly poorer on the Gesture Test. The profile of performance on each test was examined in detail but no evidence of group differences in the pattern of impairment was found. CONCLUSIONS: This study is consistent with others suggesting a high prevalence of clumsiness in Asperger's syndrome. Our findings also attest to the widespread prevalence of motor impairment in developmental disorders and the problems such co-morbidity poses for attempts to posit discrete and functionally coherent impairments underlying distinct syndromes. PMID- 12120862 TI - Gaze behavior of children with pervasive developmental disorder toward human faces: a fixation time study. AB - BACKGROUND: The abnormal gaze behavior of autistic children toward human faces, as observed in daily-life situations, are investigated in two fixation time studies. It has been argued that faces are a special kind of stimuli for normal individuals and that this might not be the case for autistic children. METHODS: A group of high-functioning autistic children (including a group of sub-threshold PDD-NOS children) was compared with a group of normal children, with respect to their fixation behavior for photographs of human faces. Using an infrared eye tracking device, fixation times for the whole face and for the facial elements of faces were compared between the two groups. The first study dealt with faces having an emotional expression. The second study dealt with neutral faces presented either upright or upside-down. RESULTS: Results of the two studies showed that autistic children have the same fixation behavior as normal children for upright faces, with or without an emotional expression. Furthermore, results of the second study showed that normal children spent less time looking at upside down faces, but that the fixation times of autistic children were not influenced by the orientation of the faces. CONCLUSIONS: These results plead against the notion that the abnormal gaze behavior in everyday life is due to the presence of facial stimuli per se. Furthermore, the absence of a face orientation effect in autistic children might be a reflection of a lack of holistic processing of human faces in autism. PMID- 12120863 TI - Conversational behaviour of children with Asperger syndrome and conduct disorder. AB - BACKGROUND: Social communication problems in individuals who have Asperger syndrome constitute one of the most significant problems in the syndrome. This study makes a systematic analysis of the difficulties demonstrated with the use of language (pragmatics) in adolescents who have Asperger syndrome. METHOD: Recent advances in discourse analysis were applied to conversational samples from a group of children with Asperger syndrome and a matched control group of children with severe conduct disorder. Two types of conversation were sampled from each group, differing in emotional content. RESULTS: The results showed that in these contexts children with Asperger syndrome were no more verbose as a group than controls, though they showed a tendency to talk more in more emotion-based conversations. Children with Asperger syndrome, as a group, performed similarly to control subjects in ability to respond to questions and comments. However, they were more likely to show responses which were problematic in both types of conversation. In addition, individuals with Asperger syndrome showed more problems in general conversation than during more emotionally and socially loaded topics. The group with Asperger syndrome was found to contain a small number of individuals with extreme verbosity but this was not a reliable characteristic of the group as a whole. PMID- 12120864 TI - Equilibrium structure and stability in a frequency-dependent, two-population diploid model. AB - We investigate the equilibrium structure for an evolutionary genetic model in discrete time involving two monoecious populations subject to intraspecific and interspecific random pairwise interactions. A characterization for local stability of an equilibrium is found, related to the proximity of this equilibrium with evolutionarily stable strategies (ESS). This extends to a multi population framework a principle initially proposed for single populations, which states that the mean population strategy at a locally stable equilibrium is as close as possible to an ESS. PMID- 12120865 TI - An alternative stochastic model of generation of oligodendrocytes in cell culture. AB - According to our previous model, oligodendrocyte--type 2 (O-2A) astrocyte progenitor cells become competent for differentiation in vitro after they complete a certain number of critical mitotic cycles. After attaining the competency to differentiate, progenitor cells divide with fixed probability p in subsequent cycles. The number of critical cycles is random; analysis of data suggests that it varies from zero to two. The present paper presents an alternative model in which there are no critical cycles, and the probability that a progenitor cell will divide again decreases gradually to a plateau value as the number of completed mitotic cycles increases. In particular all progenitor cells have the ability to differentiate from the time of plating. The Kiefer-Wolfowitz procedure is used to fit the new model to experimental data on the clonal growth of purified O-2A progenitor cells obtained from the optic nerves of 7 day old rats. The new model is shown to fit the experimental data well, indicating that it is not possible to determine whether critical cycles exist on the basis of these experimental data. In contrast to the fit of the previous model, which suggested that the addition of thyroid hormone increased the limiting probability of differentiation as the number of mitotic cycles increases, the fit of the new model suggests that the addition of thyroid hormone has almost no effect on the limiting probability of differentiation. PMID- 12120866 TI - Structured population on two patches: modeling dispersal and delay. AB - We derive from the age-structured model a system of delay differential equations to describe the interaction of spatial dispersal (over two patches) and time delay (arising from the maturation period). Our model analysis shows that varying the immature death rate can alter the behavior of the homogeneous equilibria, leading to transient oscillations around an intermediate equilibrium and complicated dynamics (in the form of the coexistence of possibly stable synchronized periodic oscillations and unstable phase-locked oscillations) near the largest equilibrium. PMID- 12120867 TI - The influence of drug treatment on the maintenance of schistosome genetic diversity. AB - Drug treatment of patients with schistosomiasis may select for drug-resistant parasites. In this article, we formulate a deterministic model with multiple strains of schistosomes (helminth parasites with a two-host life cycles) in order to explore the role of drug treatment in the maintenance of a polymorphism of parasite strains that differ in their resistance levels. The basic reproductive numbers for all strains are computed, and are shown to determine the stabilities of equilibria of the model and consequently the distribution of parasite phenotypes with different levels of drug tolerance. Analysis of our model shows that the likelihood that resistant strains will increase in frequency depends on the interplay between their relative fitness, the cost of resistance, and the degree of selection pressure exerted by the drug treatments. PMID- 12120868 TI - The effect of density-dependent treatment and behavior change on the dynamics of HIV transmission. AB - In this work, we propose a model for heterosexual transmission of HIV/AIDS in a population of varying size with an intervention program in which treatment and/or behavior change of the infecteds occur as an increasing function of the density of the infected class in the population. This assumption has socio-economic implications which is important for public health considerations since density dependent treatment/behavior change may be more cost-saving than a program where treatment/behavior change occurs linearly with respect to the number of infecteds. We will make use of the conservation law of total sexual contacts which enables us to reduce the two-sex model to a simpler one-sex formulation. Analytical results will be given. Unlike a similar model with linear treatment/behavior change in Hsieh (1996) where conditions were obtained for the eradication of disease, we will show that density-dependent treatment/behavior change cannot eradicate the disease if the disease is able to persist without any treatment/behavior change. This work demonstrates the need to further understand how treatment/behavior change occurs in a society with varying population. PMID- 12120869 TI - Oscillations in a refractory neural net. AB - A functional differential equation that arises from the classic theory of neural networks is considered. As the length of the absolute refractory period is varied, there is, as shown here, a super-critical Hopf bifurcation. As the ratio of the refractory period to the time constant of the network increases, a novel relaxation oscillation occurs. Some approximations are made and the period of this oscillation is computed. PMID- 12120870 TI - A new mathematical model for avascular tumour growth. AB - The early development of solid tumours has been extensively studied, both experimentally via the multicellular spheroid assay, and theoretically using mathematical modelling. The vast majority of previous models apply specifically to multicell spheroids, which have a characteristic structure of a proliferating rim and a necrotic core, separated by a band of quiescent cells. Many previous models represent these as discrete layers, separated by moving boundaries. Here, the authors develop a new model, formulated in terms of continuum densities of proliferating, quiescent and necrotic cells, together with a generic nutrient/growth factor. The model is oriented towards an in vivo rather than in vitro setting, and crucially allows for nutrient supply from underlying tissue, which will arise in the two-dimensional setting of a tumour growing within an epithelium. In addition, the model involves a new representation of cell movement, which reflects contact inhibition of migration. Model solutions are able to reproduce the classic three layer structure familiar from multicellular spheroids, but also show that new behaviour can occur as a result of the nutrient supply from underlying tissue. The authors analyse these different solution types by approximate solution of the travelling wave equations, enabling a detailed classification of wave front solutions. PMID- 12120871 TI - Complex ligand-protein systems: a globally convergent iterative method for the n x m case. AB - When n types of univalent ligands are competing for the binding to m types of protein sites, the determination of the system composition at equilibrium reduces to the solving of a non-linear system of n equations in C = [0; 1](n). We present an iterative method to solve such a system. We show that the sequence presented here is always convergent, regardless of the initial value in C. We also prove that the limit of this sequence is the unique solution in C of the non-linear system of equations. PMID- 12120872 TI - A model for actin-filament length distribution in a lamellipod. AB - A mathematical model is derived to describe the distributions of lengths of cytoskeletal actin filaments, along a 1 D transect of the lamellipod (or along the axis of a filopod) in an animal cell. We use the facts that actin filament barbed ends are aligned towards the cell membrane and that these ends grow rapidly in the presence of actin monomer as long as they are uncapped. Once a barbed end is capped, its filament tends to be degraded by fragmentation or depolymerization. Both the growth (by polymerization) and the fragmentation by actin-cutting agents are depicted in the model, which takes into account the dependence of cutting probability on the position along a filament. It is assumed that barbed ends are capped rapidly away from the cell membrane. The model consists of a system of discrete-integro-PDE's that describe the densities of barbed filament ends as a function of spatial position and length of their actin filament "tails". The population of capped barbed ends and their trailing filaments is similarly represented. This formulation allows us to investigate hypotheses about the fragmentation and polymerization of filaments in a caricature of the lamellipod and compare theoretical and observed actin density profiles. PMID- 12120873 TI - Radiation breakage of DNA: a model based on random-walk chromatin structure. AB - Monte Carlo computer software, called DNAbreak, has recently been developed to analyze observed non-random clustering of DNA double strand breaks in chromatin after exposure to densely ionizing radiation. The software models coarse-grained configurations of chromatin and radiation tracks, small-scale details being suppressed in order to obtain statistical results for larger scales, up to the size of a whole chromosome. We here give an analytic counterpart of the numerical model, useful for benchmarks, for elucidating the numerical results, for analyzing the assumptions of a more general but less mechanistic "randomly located-clusters" formalism, and, potentially, for speeding up the calculations. The equations characterize multi-track DNA fragment-size distributions in terms of one-track action; an important step in extrapolating high-dose laboratory results to the much lower doses of main interest in environmental or occupational risk estimation. The approach can utilize the experimental information on DNA fragment-size distributions to draw inferences about large-scale chromatin geometry during cell-cycle interphase. PMID- 12120874 TI - Comparative in-vitro activities of ertapenem against aerobic bacterial pathogens isolated from patients with complicated intra-abdominal infections. AB - The in vitro activities of ertapenem, ceftriaxone, amoxicillin-clavulanate, ampicillin-sulbactam, and piperacillin-tazobactam were compared against 1018 aerobic bacterial pathogens isolated from 531 patients with complicated intra abdominal infection. Enterobacteriaceae accounted for 66.3% of the aerobic bacteria; Escherichia coli was the most common isolate. The ertapenem minimal inhibitory concentration was < or = 2 microg/mL for 74.6% of isolates and > or = 8 microg/mL for 21.9% (including isolates of enterococci, methicillin-resistant Staphylococcus aureus, Acinetobacter baumannii, and Pseudomonas aeruginosa). Against Enterobacteriaceae, ertapenem was the most potent and the most active drug evaluated (100% susceptible), followed by ceftriaxone (98% susceptible), piperacillin-tazobactam (96% susceptible), amoxicillin-clavulanate (80% susceptible), and ampicillin-sulbactam (64% susceptible). Piperacillin-tazobactam was the only drug evaluated with clinically useful activity against P. aeruginosa. In summary, ertapenem was highly active in vitro against many clinically important aerobic intra-abdominal bacterial pathogens, especially Enterobacteriaceae. PMID- 12120875 TI - In-vitro activity of clinafloxacin compared to ciprofloxacin against Acinetobacter baumannii strains isolated from intensive care unit patients. AB - The activity of clinafloxacin was compared to that of ciprofloxacin against 154 Acinetobacter baumannii strains isolated from patients treated in Intensive Care Units. Minimum inhibitory concentrations (MICs) were determined by the Epsilometer test method. The majority (87.6%) of the A. baumannii strains tested were resistant to ciprofloxacin (MIC range 0.125->32, MIC50 = >32, MIC90 = >32). On the contrary, only 9.7% of the strains tested were resistant to clinafloxacin (MIC range 0.023->4, MIC50 = 0.75, MIC90 = 2). Due to its superior activity shown against A. baumannii strains, compared to ciprofloxacin, clinafloxacin may be added to the therapeutic armamentarium for hospital-acquired infections caused by A. baumannii in Intensive Care Units. PMID- 12120876 TI - In-vitro susceptibility of quinolone-resistant clinical isolates of Escherichia coli to fosfomycin trometamol. AB - Escherichia coli (E. coli) is the most commonly isolated microorganism in uncomplicated lower urinary tract infections (UTI). Due to the increased isolation of E. coli strains resistant to quinolones, it is important to have available alternative drugs to this class of antibiotics as therapy for UTIs caused by this pathogen. Among the large number of currently available antimicrobial agents, fosfomycin trometamol is a useful alternative due to its peculiar microbiological and pharmacokinetic properties. Therefore, we tested the in vitro susceptibility of 79 quinolone-resistant clinical urinary isolates of E. coli to fosfomycin trometamol in comparison with amoxicillin, chloramphenicol, cotrimoxazole, netilmicin, nitrofurantoin and tetracycline. Fosfomycin trometamol showed high activity with a MIC90 of 4 mg/l. While no strains were resistant to fosfomycin trometamol, 83.5%, 63.3%, 58.2%, and 48.1% of the isolates were resistant to tetracycline, amoxicillin, chloramphenicol and cotrimoxazole, respectively. Nitrofurantoin and netilmicin resistance was present only in 12.7% and 6.3% of the strains, respectively. In conclusion, fosfomycin trometamol has retained its activity against quinolone-resistant strains of E. coli and cross resistance with other classes of antimicrobial agents is not presently a problem. The strains tested did present high levels of resistance to other classes of antibiotics. PMID- 12120877 TI - Remarkable substituent effects on antimicrobial activities of 1,3 diorganylimidazolidinium salts. AB - In vitro antibacterial and antifungal activities of a series of 25 diazolidinium salts, 1,3-diorganylimidazolidinium (1), together with 1,3-dialkylpyrimidinium (2), were evaluated against standard strains: Escherichia coli (ATCC 25922), Staphylococcus epidermidis (ATCC 12228), Staphylococcus aureus (ATCC 29213), Enterococcus faecalis (ATCC 29212), Enterobacter cloacae (ATCC 13047), Pseudomonas aeruginosa (ATCC 27853) and Candida albicans (ATCC 10239). Selective and effective antibacterial activity against one gram-negative (P. aeruginosa) and two gram-positive (E. faecalis and S. aureus) bacteria were found in salts 1a, 1i and 1j, in contrast to modest to poor activity observed in the rest of the salts. The enhanced antibacterial activity is clearly linked to the introduction of two bulky mesityl or mesitylbenzyl substituents on the nitrogen atoms of the imidazoline skeleton, while the side chain of the backbone of the molecule has no influence. PMID- 12120878 TI - Effect of medium composition on static and cidal activity of amphotericin B, itraconazole, voriconazole, posaconazole and terbinafine against Aspergillus fumigatus: a multicenter study. AB - The effect of the medium composition on the fungistatic (MIC) and fungicidal (MLC) activity of amphotericin B, itraconazole, voriconazole, posaconazole and terbinafine against four Aspergillus fumigatus strains has been investigated by four European laboratories. MICs were determined by broth microdilution, using RPMI 1640 and Antibiotic Medium 3 (AM3), three times in three independent determinations by the four laboratories. MLCs were determined for the three independent determinations by the four laboratories, subculturing 100 microl from each well showing no visible growth after 48 hours. Except for a 2-dilution difference observed in three cases, no differences were observed between MICs determined on the two media. In contrast, a 3- to 6-dilution discrepancy between the MLCs was observed for the azoles. Endpoints on RPMI were higher than those on AM3. A 1-2 dilution difference was noted between both the endpoints of amphotericin B and of terbinafine. The highest inter- and intra-laboratory agreements were reached on AM3. The azoles showed a medium-dependent fungicidal activity. PMID- 12120879 TI - Emerging antimicrobial resistances among Proteus mirabilis in Europe: report from the MYSTIC Program (1997-2001). Meropenem Yearly Susceptibility Test Information Collection. AB - Resistance patterns that are currently problematic in Europe can vary greatly within the same species over time, among various patient populations and among geographic regions on the same continent. The results from the Meropenem Yearly Susceptibility Test Information Collection (MYSTIC) Program, which monitors carbapenem resistance rates in institutions using meropenem, were used to determine resistance differences among Proteus mirabilis. MIC results from 688 P. mirabilis strains were classified into 4 patient care groups: ICU (n=426), neutropenia patients (NP; n=145), general wards (n=97) and cystic fibrosis patients (CF; n=20). A total of 40 centers from 12 European countries have participated since 1997, divided into 3 geographic regions (East, North, South). All testing was performed by NCCLS reference methods and interpretive criteria, including screening of extended-spectrum beta-lactamase (ESBL) phenotypes. Over the monitored interval the resistance rates varied for each agent without a clear trend toward a greater rate. Rank order of susceptibility was: meropenem (99%) > piperacillin/tazobactam (TAZ; 96%) > cefepime (95%) > ceftazidime (CAZ; 94%) > imipenem (IPM; 92%). Ciprofloxacin (CIP) was the least active agent tested (MIC90 4 microg/ml; 86% susceptible). Unexpectedly, 3.6% of P. mirabilis were imipenem resistant (MIC, > or = 16 microg/ml). Greater rates of resistance were found for strains from NP and CF patients, and from eastern or southern European sites, usually associated with epidemic clusters. Generally susceptible species such as P. mirabilis have recently emerged as therapeutic problems in European medical centers following mutations that compromise CIP, CAZ and aminoglycoside use. Imipenem also showed decreased susceptibility of greater than 7% compared to less than 1% for meropenem. Continued surveillance by the MYSTIC Program appears to be a prudent practice to focus effective empiric treatment regimens. PMID- 12120880 TI - Antibiotic resistance of bacteria associated with community-acquired urinary tract infections in the southern area of the Gaza Strip. AB - The aim of this study was to identify the microorganisms that cause "community acquired" urinary tract infections among adults and to investigate their resistance to fourteen selected antimicrobial agents. The uropathogens identified in 121 positive midstream urine cultures from the 270 subjects included in this study were Escherichia coli (57.9%), Proteus species (9.9%), Enterobacter species (7.4%), Klebsiella species (6.6%), Pseudomonas species (5.8%), Staphylococcus saprophyticus (5.0%), Enterococcus species (3.3%), Acinetobacter species (2.5%), Citrobacter species (0.8%) and Staphylococcus aureus (0.8%). Isolates were subjected to antimicrobial susceptibility testing and a high proportion of the isolates was found to be resistant to amoxycillin (73.6%), doxycycline (68.6%) and trimethoprim-sulfamethoxazole (66.1%). The most effective drugs against all the isolates were ciprofloxacin (95.9%), amikacin (95.0%) and ceftazidime (94.2%). A high percentage of multiple-drug resistance was also observed for the majority of the isolates. PMID- 12120881 TI - An open, comparative pilot study of thiamphenicol glycinate hydrochloride vs clarithromycin in the treatment of acute lower respiratory tract infections due to Chlamydia pneumoniae. AB - The aim of this study was to evaluate the efficacy and tolerability of thiamphenicol glycinate hydrochloride (TGH) i.m. versus clarithromycin in acute lower respiratory infections due to Chlamydia pneumonia. 113 patients with suspected pneumonia were screened. 40 patients with IgM and/or IgA titers > or = 1:16 and/or IgG titers > or = 1:512 were assigned to 10 days of treatment with TGH 1500 mg daily or clarithromycin 1000 mg daily. 34 patients were considered a clinical success. 33 patients were a radiological success. 22 patients showed a decrease in IgG values. 3 patients had an increase in IgG values. Blood/urine values presented no clinically significant variations. Clinical efficacy was similar in both treatment groups. These are the first results confirming in vivo the recent in vitro evidence that TGH is effective against acute lower respiratory tract infections due to C. pneumoniae, thus representing an alternative therapy to clarithromycin. PMID- 12120882 TI - Repeated point prevalence survey of nosocomial infections in a Greek university hospital. AB - Two point prevalence surveys of nosocomial infections (NIs) were carried out in a Greek University hospital on an annual basis in 1998 and 1999. The overall prevalence of NIs was 9.5% and 9.1% in the first and second study, respectively. The average length of stay of patients in the hospital (ALOS) was 7.7 and 9.6 days in these two studies, respectively. Of the 97 NIs detected, the most frequent were lower respiratory tract infections (36%). Urinary tract infections, bloodstream infections, surgical site infections, and gastrointestinal infections were found in 25.8%, 19.6%, 7.2% and 4.1% of patients, respectively. The prevalence of antibiotic usage was 55.6% in 1998 and 54.1% in 1999. Empiric antibiotic therapy prevailed over prophylactic and rational therapies. These percentages are higher than those reported from other countries, emphasizing the need for rational antibiotic usage to decrease pharmacy expenses and discourage the development of resistant microorganisms. A nationwide network of surveillance of NIs in Greece is now being developed using these experiences. PMID- 12120883 TI - Recent clinical evidence of the efficacy and safety of thiamphenicol glycinate acetylcysteinate and thiamphenicol glycinate. AB - Thiamphenicol is a broad-spectrum antimicrobial agent active against penicillin resistant Streptococcus pneumoniae, Staphylococcus aureus VISA strains, most methicillin-resistant isolates and atypical pathogens such as Mycoplasma pneumoniae and Chlamydia pneumoniae). Thiamphenicol is present as glycinate hydrochloride (TG) and glycinate acetylcysteinate (TGA) esters in the parenteral and aerosol dosage form. This multicenter, double-blind, randomized clinical trial aimed to evaluate the efficacy and tolerability of aerosol administration of TGA, compared to TG, in the treatment of acute and/or exacerbated infections of the respiratory tract. Results showed that both treatments ameliorated the symptoms (frequency and severity of cough, difficulty in expectoration) associated with the evaluated pathologies, i.e. tracheobronchitis, acute and exacerbated chronic bronchitis. The investigators rated both treatments Good or Very Good in 90% of patients at the end of treatment, with "Very Good" for patients treated with TGA (37%) compared to 28% of patients treated with TG. Both treatments were well tolerated with fewer than 5% of patients experiencing an adverse event. PMID- 12120884 TI - Comparison of different treatment combinations for chronic hepatitis B infection. AB - Chronic hepatitis B virus (HBV) infection is a leading cause of cirrhosis and hepatocellular carcinoma worldwide. Its prevalence approaches 10% in hyper endemic areas. The aim of treating chronic HBV infection is to halt progression of liver injury by suppressing viral replication or eliminating infection. This study was planned to evaluate the advantages of combination therapy with interferon-alpha plus second-generation nucleoside analogues (lamivudine or famciclovir), or vaccination with a pre-S2 and S proteins containing vaccine in chronic HBV infection. 29 patients were divided into three groups and were treated with the following combinations: (1) IFN-alpha2a 9 million units 3x week for 6 months with HBV vaccine 20 microg given on 0, 1 and 2 months; (2) IFN alpha2a 6 million units 3x week plus famciclovir 250 mg 3x day for 6 months; (3) IFN-alpha2a 6 million units 3x week plus lamivudine 100 mg/day for 6 months. Complete response was suspected in 3 patients in group 1, in 4 patients in group 2, and in 7 patients in group 3. Partial response was suspected in 4, 1 and 2 patients in groups 1, 2 and 3, respectively. The results of the present study suggest that the combination of IFN-alpha with lamivudine is more effective than the combination of IFN-alpha with HBV vaccination or famciclovir. PMID- 12120885 TI - Effects of rifaximin administration on the intestinal microbiota in patients with ulcerative colitis. AB - The effect of rifaximin on the intestinal bacterial population was studied in a clinical trial. Twelve patients with ulcerative colitis were administered rifaximin 1800 mg/day in 3 treatment periods of 10 days, each followed by 25 days of wash-out. Fecal samples were collected at the beginning and at the end of each treatment period to perform microbiological examinations. Titer variations of enterococci, coliforms, lactobacilli, bifidobacteria, Bacteroides spp., and Clostridium perfringens as well as their susceptibility to rifaximin during the different phases of the study were evaluated. The presence of Candida spp. was also monitored. After each wash-out period, concentrations of the intestinal microbial groups tested returned to initial values, showing that the administration of high doses of rifaximin does not significantly modify the colonic microbiota. Rifaximin-resistant isolates were also found, particularly in bacteria belonging to Bifidobacterium genus, included as probiotics in several fermented foods and in pharmaceutical preparations. PMID- 12120886 TI - A dose finding study of carboplatin and gemcitabine in advanced non-small cell lung cancer. AB - The excellent activity of the cisplatin-gemcitabine combination and favorable toxicological profile of carboplatin are the basis of carboplatin-gemcitabine combination therapy for non-small cell lung cancer. We carried out a dose-finding study with the aim of establishing the maximum tolerated dose (MTD) of carboplatin on day 1 in combination with gemcitabine at the dose of 1000 mg/m2 on days 1 and 8 in a 21-day cycle. The starting dose level for carboplatin was the area under the concentration time curve (AUC) 4 mg/ml/min. 18 patients were treated and a dose limiting toxicity was observed in 2 cases at the level of AUC 6 mg/ml/min. AUC 5 mg/ml/min was considered as the MTD for carboplatin in our regimen. Notably, 7 objective responses were observed. PMID- 12120888 TI - Asymptomatic acute pancreatitis due to tamoxifen-induced severe hypertriglyceridemia in a patient with diabetes mellitus and breast cancer. AB - We report tamoxifen-induced hypertriglyceridemia and asymptomatic acute pancreatitis in a 51 year-old women with type 2 diabetes mellitus and stage III-b infiltrative ductal carcinoma, admitted to the hospital with weakness, oliguria and glucose dysregulation. On admission, there was no fever, abdominal or back pain, rebound tenderness, nausea, or vomiting. Following 1 year of tamoxifen treatment, triglycerides increased from 400 to 1344 mg/dl (blood urea nitrogen 52 mg/dl, creatinine 2.0 mg/dl, glucose 341 mg/dl). Hypertriglyceridemia was considered to be due to either diabetic dyslipidemia and/or tamoxifen. On computerized tomography, pancreatic enlargement, heterogenity, hypodensity and a pancreatic pseudocyst (5 x 7.5 cm diameter) were found. Acute pancreatitis was suspected, and serum amylase level was found to be increased (273 IU/L). Tamoxifen was discontinued and gemfibrozil was started. Triglycerides decreased to 301 mg/dl and amylase decreased to 66 IU/L a week later and remained normal thereafter. This case indicates that tamoxifen-induced hypertriglyceridemia may cause acute pancreatitis without classical symptoms which might be due to autonomic neuropathy in diabetic patients. Effects on lipid metabolism should be considered and triglycerides should be closely followed in patients on tamoxifen. PMID- 12120887 TI - Ralitrexed (Tomudex) or Nordic-FLv regimen in metastatic colorectal cancer: a randomized phase II study focusing on quality of life, patients' preferences and health economics. AB - Raltitrexed (Tomudex) is proven effective in metastatic colorectal cancer. Between 1998-2000, 25 patients were included in a randomized phase II study comparing raltitrexed (13 patients) and the Nordic FLv regimen (12 patients). 23 patients were evaluable for response. The overall response rate was 2/12 (1 CR, 1 PR) in the raltitrexed arm and 1/11 (1 CR) in the Nordic FLv arm, respectively. There was no difference in overall survival (raltitrexed--14.7 months, Nordic FLv -15.4 months). 23 patients were evaluable for Quality of Life (QoL) analysis. 23/25 and 17/21 questionnaires (EORTC QLQ C-30) were returned at baseline and first evaluation. Raltitrexed tended to be the most toxic regimen, when looking at nausea and vomiting, appetite loss, diarrhea and global QoL. However, most patients (65%) recommended the raltitrexed treatment schedule. The total treatment cost was equal in both arms (about 6,800 EURO/patient) and the hospital/hospital hotel stay costs accounted for more than half of it. PMID- 12120889 TI - Culturing Chlamydophila pneumoniae. PMID- 12120890 TI - Effects of subinhibitory concentrations of ibuprofen isobuthanolammonium on virulence factors of uropathogenic Escherichia coli. PMID- 12120891 TI - Pathophysiology of ageing. AB - Ageing is characterized by a gradual decline in organ functional reserves which reduces the ability to maintain homeostasis under conditions of stress. Introduction of cell culture and molecular biology techniques has provided new experimental tools for the analysis of ageing at the molecular level. During ageing progressive degeneration of cells and loss of regenerative capacity are enhanced and with time the alterations caused by them ultimately lead to death. In this paper the current knowledge of the mechanisms of ageing is summarized. PMID- 12120892 TI - Serum response factor: discovery, biochemistry, biological roles and implications for tissue injury healing. AB - Serum response factor (SRF) is a transcription factor, which binds to a serum response element (SRE) associated with a variety of genes including immediate early genes such as c-fos, fosB, junB, egr-1 and -2, neuronal genes such as nurr1 and nur77 and muscle genes such as actins and myosins. By regulating expression of these genes, SRF controls cell growth and differentiation, neuronal transmission as well as muscle development and function. SRF can be activated by a variety of agents, including serum, lysophosphatidic acid (LPA), lipopolysaccharide (LPS), 12-O-tetradecanoylphorbol-13-acetate (TPA), cytokines, tumor necrosis factor-alpha (TNFalpha), agents that increase intracellular Ca2+, T-cell virus1 activator protein, hepatitis B virus activator proteins pX, activated oncogenes and protooncogenes as well as extracellular stimuli such as antioxidant and UV light. SRF itself is regulated by both cellular signal transduction pathways and interaction with other transcription factors e.g. Sp1, ATF6 and myogenic regulatory factors. Its biological function is best elucidated for myocardium. Specific cardiac SRF transgenesis demonstrated that overexpression of SRF caused hypertrophic cardiomyopathy in mouse and the mouse died of heart failure within 6 months after birth. Other transgenic data suggested that sufficient SRF was needed for embryogenesis and early development. Since SRF is important regulator of numerous genes involved in cell growth and differentiation, including muscle and neural components, SRF may also play a crucial role in tissue injury and ulcer healing, e.g. healing of gastrointestinal ulcers. PMID- 12120893 TI - Activation of peroxisome proliferator-activated receptor gamma suppresses inducible cyclooxygenase and nitric oxide synthase during oral mucosal ulcer healing. AB - BACKGROUND: Peroxisome proliferator-activated receptor-gamma (PPARgamma) is a ligand-dependent transcription factor, belonging to the steroid hormone receptor family, known to play a pivotal role in the resolution of inflammation. In this study, we investigated the effect of a specific PPARgamma ligand, ciglitazone, on the course of buccal mucosal ulcer healing by analyzing mucosal activity of inducible nitric oxide synthase (NOS-2) and the expression cyclooxygenases (COX-1 and COX-2) responsible for prostaglandin (PG) generation. METHODS: Groups of rats with experimentally induced buccal mucosal ulcers were administered twice daily for up to 10 days with ciglitazone at 5, 10, and 15 mg/kg or the vehicle, and their mucosal tissue subjected to assessment of ulcer healing rate and biochemical measurements. RESULTS: The ulcer onset, characterized by up regulation of NOS-2 and COX-2 protein expression, was reflected in a marked increase in the mucosal PGE2 generation and NOS-2 activity, whereas healing was accompanied by a drop in PGE2 and NOS-2 activity, and a decrease in COX-2 and NOS 2 protein expression. The mucosal expression of COX-1 protein, however, remained unchanged. Administration of ciglitazone led to a significant dose-dependent acceleration in the mucosal reduction of PGE2 generation and NOS-2 activity, and produced a marked decline in COX-2 and NOS-2 protein expression, but the rate of ulcer healing and the expression of COX-1 protein remained unaffected. CONCLUSIONS: Our findings thus suggest that the products of induced NOS-2 and COX 2 enzymes, associated with mucosal inflammatory responses to injury, do not play a significant role in oral mucosal ulcer healing. PMID- 12120894 TI - The influence of cholecystokinin on gastric myoelectrical activity in duodenal ulcer following Helicobacter pylori eradication--an electrogastrographic study. AB - Cholecystokinin (CCK) plays an important role in the regulation of postprandial gastric motor activity which was found to be abnormal in duodenal ulcer patients. This study was designed to compare the influence of CCK on gastric myoelectrical function in duodenal ulcer patients and healthy controls. Fifteen patients with active duodenal ulcer and Helicobacterpylori (H. pylori) infection and 15 healthy controls were included into this study. Electrogastrography (EGG) was performed before and 4 weeks after the eradication of H. pylori in ulcer patients and in healthy controls. We compared EGG parameters in the fasting and postprandial period and during intravenous infusion of caerulein, an analog of CCK with or without addition of loxiglumide, a specific CCK-1 receptor antagonist. The amplitude of fasting EGG in duodenal ulcer patients was similar to that in control subjects and was not affected by H. pylori eradication. In contrast, the amplitude of postprandial EGG was markedly increased in duodenal ulcer patients when compared to that in healthy controls and it was significantly reduced following the eradication of H. pylori. The blockade of CCK-1 receptors with loxiglumide in healthy controls or H. pylori eradicated ulcer patients significantly enhanced postprandial EGG amplitude almost to the level observed in the infected duodenal ulcer patients, but failed to affect this amplitude in ulcer patients. Exogenous caerulein, an analog of CCK, failed to affect EGG amplitude in duodenal ulcer patients with H. pylori infection, but it reduced significantly EGG amplitude in these patients after H. pylori eradication and in control subjects. This inhibitory effect of caerulein in H. pylori negative ulcer patients and healthy controls was abolished by the addition of loxiglumide. Ulcer patients showed significant dysrhythmia with tachygastria up to 20% of the recording time both under basal conditions and postprandially and H. pylori eradication was followed by a significant decrease in tachygastria to about 5%, the value being similar to that in healthy controls. We conclude that the amplitude and frequency of gastric myoelectrical activity are enhanced in duodenal ulcer patients and impaired in response to CCK but these changes can be normalized by successful H. pylori eradication. PMID- 12120895 TI - The influence of epidermal growth factor on the course of ischemia-reperfusion induced pancreatitis in rats. AB - Acute pancreatitis is accompanied by the enhanced expression of EGF in the pancreas and the administration of EGF was found to exhibit the beneficial effect on edematous cerulein-induced pancreatitis. Therefore, we decided to determine the influence of EGF on necro-hemorrhagic pancreatitis induced by ischemia and reperfusion (I/R). Acute pancreatitis was induced in rats by restricting the pancreatic blood flow (PBF) in the inferior splenic artery for 30 min using microvascular clips. EGF was administered three times daily (10 microg/kg per dose s.c.) starting immediately after the clips removal. Rats were sacrificed on day 1, 3, 5, 10 and 21 following ischemia. PBF was measured using a laser Doppler flowmeter. Morphological signs of pancreatitis, as well as the levels of plasma amylase, lipase, interleukin-1beta and interleukin-10 concentration and pancreatic cell proliferation were examined. RESULTS: Ischemia with reperfusion caused acute necro-hemorrhagic pancreatitis with a histological and biochemical manifestation of pancreatic damage, followed by a spontaneous regeneration. The administration of EGF caused the reduction in the histological signs of pancreatic damage, such as necrosis, edema and leukocyte infiltration, and accelerated the pancreatic repair. Also, EGF treatment significantly attenuated the reduction in pancreatic blood flow and DNA synthesis. The activity of plasma amylase and lipase, as well as plasma interleukin-1beta and interleukin-10 concentrations were decreased in EGF treated animals. CONCLUSIONS: EGF exerts beneficial influence on the course of I/R induced pancreatitis and this effect seems to be related to the reduction in the activation of pro-inflammatory interleukin cascade, the improvement of PBF, and the increase in pancreatic cell growth. PMID- 12120896 TI - Effect of static handgrip on plasma adrenomedullin concentration in patients with heart failure and in healthy subjects. AB - Adrenomedullin (ADM) release is enhanced in pheochromocytoma, chronic heart failure (HF), hypertension and renal diseases. This study was designed to test the hypothesis that ADM secretion increases also in response to acute stimuli, such as static effort and to compare plasma ADM response to this stimulus in patients with chronic HF and healthy persons. Eight male HF patients (II/III class NYHA) and eight healthy subjects (C) performed two 3-min bouts of static handgrip at 30% of maximal voluntary contraction, alternately with each hand without any break between the bouts. At the end of both exercise bouts and in 5 min of the recovery period, plasma ADM and catecholamines were determined. In addition, heart rate, blood pressure, and stroke volume (SV) were measured. The baseline plasma ADM and noradrenaline levels were higher, whilst plasma adrenaline and SV were lower in HF patients than in C group. The 1st exercise bout caused an increase in plasma ADM from 3.32 +/- 0.57 to 4.98 +/- 0.59 pmol l( 1) (p<0.01) in C and from 6.88 +/- 0.58 to 7.80 +/- 0.43 pmol x l(-1) (p<0.02) in HF patients. The 2nd exercise bout did not produce further elevation in plasma ADM and during recovery the hormone concentration declined to pre-exercise or lower values. There were no differences between groups in exercise-induced increases in plasma ADM. Plasma ADM correlated with SV (r = -0.419) and with noradrenaline concentrations (r = 0.427). It is concluded that static exercise causes the short-lasting increase in plasma ADM concentration which is similar in healthy subjects and in patients with mild heart failure. PMID- 12120897 TI - The opposite effects of cyclic AMP-protein kinase a signal transduction pathway on renal cortical and medullary Na+,K+-ATPase activity. AB - Cyclic AMP-protein kinase A (PKA) pathway plays an important role in signal transduction in renal tubular cells, however, its role in transport regulation is not completely established. The aim of this study was to investigate in vivo the effect of PKA on renal Na, K-ATPase activity. The study was performed in male Wistar rats. The animals were anaesthetized with pentobarbital and investigated drugs were infused through the catheter inserted into the abdominal aorta. Na+,K+ ATPase activity was assayed in an isolated microsomal fraction of the renal cortex and medulla. Cell-permeable cAMP analogue, dibutyryl-cAMP (db-cAMP), dose dependently stimulated Na+,K+-ATPase in the renal cortex and inhibited in the renal medulla. Maximal stimulation (+38.5%) and inhibition (-46.8%) were observed at a dose of 10(-6) mol/kg/min. Measurement of Na+,K+-ATPase activity at different Na' concentrations revealed that in the renal cortex db-cAMP increased Vmax of the enzyme without any effect on sodium affinity, whereas in the renal medulla decrease in Vmax was accompanied by decreased sodium affinity, evidenced by elevated K(0.5) for sodium. The effect of db-cAMP was mimicked by the infusion of either adenylate cyclase activator, forskolin, or inhibitor of phosphodiesterase, IBMX. Both stimulatory and inhibitory effects of db-cAMP were prevented by pretreatment with protein kinase A inhibitor, KT 5720 (10(-8) mol/kg/min) but not by inhibitor of protein kinase G, KT 5823. The inhibitory effect in the renal medulla was partially blocked by pretreatment with either ethoxyresorufin or 17-ODYA - two nonspecific inhibitors of cytochrome P450 dependent arachidonate metabolism, whereas an inhibitor of epoxygenase, miconazole, was not effective. Infusion of 20-hydroxyeicosatetraenoic acid (20 HETE) at a dose of 10(-10) mol/kg/min decreased medullary Na+,K+-ATPase activity by 24.2%. Exogenous protein phosphatases inhibitor, okadaic acid (OA, 10(-8) - 10(-7) mol/kg/min) caused dose-dependent decrease in renal medullary Na+,K+ ATPase activity, maximally by 31.9%, but had no effect in the renal cortex. The effects of OA and db-cAMP in the renal medulla were not additive. When OA administration (10(-7) mol/kg/min) was followed by 20-HETE (10(-10) mol/kg/min), medullary Na+,K-ATPase activity decreased by 48.6% and was similar as after db cAMP. We conclude, that cAMP-PKA pathway activates Na+,K+-ATPase in the renal cortex and inhibits in the renal medulla. The inhibitory effect is partially mediated by cytochrome P450-dependent arachidonate metabolites and possibly also by PKA-dependent inhibition of protein phosphatases. PMID- 12120898 TI - The effect of amphetamine sensitization on mouse immunoreactivity. AB - Recent studies indicate a role of the immune system in the behavioral effects of amphetamine in rodents. In the present study we attempted to find a connection between the behavioral changes induced by repeated, intermittent administration of amphetamine and some immunological consequences of sensitization to amphetamine in mice. Male Albino Swiss mice were treated repeatedly (for 5 days) with amphetamine (1 mg/kg, i.p.). On day 9, they received a challenge dose of amphetamine (1 mg/kg). Acute administration of amphetamine increased their locomotor activity by ca. 40%. In animals treated repeatedly with amphetamine, the challenge dose of the psychostimulant induced behavioral sensitization, i.e. the higher locomotor activation as compared with that after its first administration to mice. Immune functions were evaluated by the ability of splenocytes to proliferate and to produce cytokines such as interferon gamma (IFN gamma), interleukin (IL)-4 and IL-10. Acute amphetamine administration significantly decreased, by ca. 30% and 25%, the proliferation of splenocytes in response to an optimal and a suboptimal dose of concanavalin A (Con A), respectively, and increased their ability to produce IL-4. Chronic intermittent treatment with amphetamine significantly decreased, by ca. 65% and 50%, the proliferative response of T cells to an optimal and a suboptimal dose of Con A, respectively, and diminished by 20% the metabolic activity of splenocytes. The above data showed that both acute and chronic amphetamine administration diminished some aspects of the cell-mediated immunity; nevertheless, immunosuppression was particularly evident in amphetamine-sensitized mice. Our findings seem to indicate possible importance of monitoring and correcting immune changes in the therapy of amphetamine addiction. PMID- 12120899 TI - Effect of allopregnanolone on d-[3H]-aspartate release and [3H]-glutamate uptake in the hippocampus of kainate-treated mice. AB - In order to determine whether the status epilepticus leads to alterations in the neurosteroid effect on excitatory amino acid transmission, we studied the influence of allopregnanolone on aspartate release and glutamate uptake in mouse hippocampus at various times after kainate administration. No significant differences in the K+-stimulated D-[3H]-aspartate release from the hippocampi of saline- and kainate-treated mice were observed; however, that parameter tended to fall in tissues collected I h after kainate administration. Allopregnanolone significantly attenuated the K+-stimulated D-[3H]-aspartate release from the hippocampi of control animals, as well at 24 h and 7 days after kainate injection; in contrast it did not affect amino acid release from the hippocampi collected 1 h after kainate administration. Kainate administration had no effect on [3H]-glutamate uptake after 1 and 24 h, but elevated that parameter on day 7. Allopregnanolone (10 and 100 microM) did not affect [3H]-glutamate uptake in control and kainate-treated mice. In conclusion, the present study indicates a loss of the inhibitory effect of allopregnanolone on the potasium-stimulated D [3H]-aspartate release from mouse hippocampus during the kainate-induced status epilepticus; moreover, it excludes involvement of this neurosteroid in the regulation of hippocampal [3H]-glutamate uptake in both control and kainate treated mice. PMID- 12120900 TI - 7-Nitroindazole enhances amphetamine-evoked dopamine release in rat striatum. an in vivo microdialysis and voltammetric study. AB - The intracellular second messenger nitric oxide (NO) is implicated in a variety of physiological functions, including release and uptake of dopamine (DA). In the described study, in vivo microdialysis and differential pulse voltammetric techniques were used to determine the involvement of NO in release of DA and its metabolites (dihydroxyphenylalanine, DOPAC; homovanillic acid, HVA) in neostriatum of freely moving rats. While the NO donor molsidomine (30.0 mg/kg; MOLS) and neuronal NO synthase- (nNOS-) inhbitor 7-nitroindazole (10.0 mg/kg; 7 NI) had no effect on the basal in vivo microdialysate level of DA, 7-NI specifically enhanced D,L-amphetamine-(1.0 mg/kg i.p.; AMPH) evoked release of DA. Basal or AMPH effects on DOPAC and HVA levels were not influenced by MOLS or 7-NI. Findings indicate that nitrergic systems have an important role in mediating effects of AMPH on dopaminergic systems. PMID- 12120901 TI - Propranolol modifies platelet serotonergic mechanisms in rats. AB - Though the mechanisms for the vascular actions of vasodilatory beta-blockers are mostly determined, some of their interactions with monoaminergic systems are not elucidated. Because there are evidences supporting a possible involvement of serotonin (5-HT) in the actions of beta-blockers, we studied the effect of propranolol on peripheral serotonergic mechanisms in normotensive and Goldblatt two-kidney - one clip (2K1C) hypertensive rats. In both groups of animals propranolol decreased systolic blood pressure, significantly increased whole blood serotonin concentration and at the same time it decreased platelet serotonin level. The uptake of the amine by platelets from hypertensive animals was lower than that of normotensive animals and it was decreased by propranolol only in the latter. In both groups propranolol inhibited potentiation of ADP induced platelet aggregation by serotonin. In conclusion, this study provides evidence that propranolol modifies platelet serotonergic mechanisms in normotensive and renal hypertensive rats. PMID- 12120903 TI - Expression of serum response factor in normal rat gastric mucosa. AB - Serum response factor (SRF) is a transcription factor that is involved in cell proliferation, muscle and neuron development and maintenance. Its expression in gastric mucosa remains unknown. In this study we demonstrated that SRF is expressed in normal rat gastric tissue as two isoforms and localized mainly to smooth muscle cells of muscularis mucosae and its extensions into the lamina propria, to muscularis propria, and musculature of the vascular system. To a lesser extent, SRF is also expressed in the gastric epithelium of the mucosal neck area (proliferative zone) and in the endothelial cells of microvessels. These data suggest that the main role of SRF in normal gastric tissue is to maintain muscular support and contraction and possibly epithelial regeneration. PMID- 12120902 TI - Effect of adrenergic antagonists and cyclooxygenase inhibitors on the nicotine induced hypothalamic-pituitary-adrenocortical activity. AB - Nicotine is a potent stimulus for the hypothalamic-pituitary-adrenal (HPA) axis. Systemic nicotine acts via central mechanisms to stimulate by multiple pathways the release of ACTH from the anterior pituitary corticotrops and corticosterone from the adrenal cortex. Nicotine may stimulate indirectly the hypothalamic paraventricular nucleus, the site of the corticotropin-releasing hormone (CRH) neurons which activates ACTH release. In the present studies an involvement of adrenergic system and prostaglandins synthesized by constitutive cyclooxygenase (COX-1) and inducible cyclooxygenase (COX-2) in the nicotine-induced HPA response in rats was investigated. Nicotine (2.5-5 mg/kg i.p.) significantly increased plasma ACTH and corticosterone levels measured 1 hr after administration. Adrenergic receptor antagonists or COX inhibitors were injected i.p. 15 min prior to nicotine and the rats were decapitated 1 hr after the last injection. Prazosin (0.01-0.1 mg/kg), an alpha1-adrenergic antagonist, significantly decreased the nicotine-evoked ACTH and corticosterone secretion. Yohimbine (0.1-1.0 mg/kg), an alpha2-adrenergic antagonist, moderately diminished ACTH response, and propranolol (0.1-10 mg/kg), a beta-adrenergic antagonist, did not significantly alter the nicotine-induced hormones secretion. Pretreatment with piroxicam (0.2 2.0 mg/kg), a COX-1 inhibitor, considerably impaired the nicotine-induced ACTH and corticosterone secretion. Compound NS-398 (0.2-5.0 mg/kg), a selective COX-2 blocker did not markedly alter these hormones secretion, and indomethacin (2 mg/kg), a non-selective COX inhibitor significantly diminished ACTH response. These results indicate that systemic nicotine stimulates the HPA axis indirectly, and both adrenergic system and prostaglandins are significantly involved in this stimulation. Noradrenaline, stimulating postsynaptic alpha1-adrenergic receptors, and prostaglandins, synthesized by COX-1 isoenzyme, are of crucial significance in the nicotine-induced ACTH and corticosterone secretion. PMID- 12120904 TI - Centrally applied vasopressin intensifies hypotension and bradycardia after hemorrhage in shr rats. AB - Spontaneuosly hypertensive rats (SHR) have been shown to exhibit several alterations in function of the intrabrain vasopressinergic system. The present study was designed to find out whether centrally administered vasopressin (AVP) may influence the cardiovascular adaptation to hypotensive hypovolemia in SHR rats. Two series of experiments were performed on conscious 17 SHR rats chronically implanted with lateral cerebral ventricle (LCV) cannulas and with femoral artery catheters. Mean arterial pressure (MAP) and heart rate (HR) were monitored before and after arterial bleeding (1,3% body weight) performed during LCV infusion of 1) artificial cerebrospinal fluid 5 microl/hour (aCSF); and 2) arginine vasopressin, 100 ng/hour/5 microl of aCSF (AVP). Central administration of aCSF and AVP had no effect on MAP and HR under resting conditions. Hemorrhage evoked significant hypotension (p<0.001) and bradycardia (p<0.001). During central infusion of AVP hemorrhage resulted in significantly greater hypotension than during central infusion of aCSF alone (p<0,05). The results provide evidence that centrally applied vasopressin significantly modulates cardivascular adjustments to hypotensive hemorrhage in SHR. PMID- 12120905 TI - Methotrexate twice weekly vs once weekly in rheumatoid arthritis: a pilot double blind, controlled study. AB - Methotrexate (MTX) is the most commonly used disease-modifying antirheumatoid drug used in patients with rheumatoid arthritis. It is usually given on a weekly schedule, but as the half-life of its active compound, the polyglutamate MTX, is 3 days, we did a pilot study to see if MTX twice weekly is superior to MTX once weekly. Eighty patients with rheumatoid arthritis (RA) fulfilling the American College of Rheumatology (ACR) criteria were enrolled into a double-blind, controlled trial of 16-week duration. Patients achieving ACR scores of 20 and 50 were determined at 8 and 16 weeks. In addition, the incidence of hepatotoxicity was also studied. Of the 80 patients, 66 completed 8 weeks of study, whereas 53 completed 16 weeks of study. Most withdrawals were because of inefficacy. At 8 weeks, ACR 20 response (24/34 in once weekly and 22/32 in twice weekly) and ACR 50 response (16/34 in once weekly and 16/32 in twice weekly) in both groups were similar. Even at 16 weeks, there was no significant difference in ACR 20 and ACR 50 responses. Intent-to-treat analysis also yielded similar results. Five patients in each group had a mild reversible rise in transaminases. Our study suggests that MTX twice weekly has no advantage over once weekly regarding efficacy, and thus the currently used once weekly regimen is good enough till alternate dosing schedules are evaluated in a large multicentric trial. PMID- 12120906 TI - Hearing loss and middle ear involvement in rheumatoid arthritis. AB - In this controlled study, hearing and middle ear functions were investigated in 37 patients with rheumatoid arthritis (RA) and 35 controls in order to study the prevalence and the nature of hearing loss in RA. The prevalence of the hearing impairment was significantly higher in the RA group, and the majority was bilateral (P<0.001). Of the patients, 35.1% had sensorineural (SN), 24.3% had conductive loss, and 10.8% had a mixed type of hearing loss. The hearing loss was positively correlated to the Steinbrocker functional index. The prevalence of abnormal tympanograms was 37.8%, while it was 17.1% in the control group. The probable site of involvement responsible for the SN loss was the cochlea, and discontinuity of the ossicles, rather than stiffness, was responsible for the conductive hearing loss. The presence of a mixed type of hearing loss suggested a multifocal involvement of the audiologic system in RA. PMID- 12120907 TI - A multicenter, case control study of risk factors for low tibial speed of sound among residents of urban areas in Turkey. AB - Risk factors which have been associated with low bone mass are multifactorial and represent regional differences between and within countries. The purpose of this study was to evaluate the possible risk factors of low tibial speed of sound (tSOS), which determines cortical bone status among residents of urban regions in Ankara, Izmir, and Istanbul, in Turkey, and also to compare groups of different socioeconomic status (SES). A total of 1,026 subjects (63% women and 53% of low socioeconomic status) 40-70 years old were included in the study. Risk factors of osteoporosis were determined using the European Vertebral Osteoporosis Study (EVOS) questionnaire, and the bone status was screened by tSOS. Socioeconomic status was found to be among the major risk factors of low tSOS in our population (odds ratio 0.39, 95% confidence interval 0.26-0.58), besides the well-known risk factors such as age and gender. Therefore, we suggest that SES is an important determinant of cortical bone status. Additionally, our results confirmed the correlation between tSOS and the clinical determinants of bone mass. PMID- 12120908 TI - Monofluorophosphate combined with hormone replacement therapy in postmenopausal osteoporosis. An open-label pilot efficacy and safety study. AB - A 3-year, open-label, monocenter study was performed on 60 patients with postmenopausal established osteoporosis treated with monofluorophosphate and calcium supplement (MFP/Ca) combined with hormone replacement therapy (HRT). Bone mineral density (BMD) after 3 years increased by 15.5% in the lumbar spine (L2 L4) and by 2.3% in the femur neck. During the 3 years, a total of six new vertebral fractures (NewVF) occurred in five patients (8.3% incidence) and a total of six nonvertebral fractures (Non-VF) occurred in six patients (10% incidence). Back pain score already decreased significantly after 6 months of treatment and, at the end of the 3rd treatment year, the pain score had decreased by 84%. The treatment was well tolerated, with only few mild or moderate adverse events. The results were compared with those of a previous study conducted in the same center with a similar protocol with a calcium supplement on patients suffering from postmenopausal established osteoporosis and treated with MFP/Ca but without HRT. It could be inferred that with the 3-year treatment, the combination of MFP/Ca with HRT protects from NewVF in one patient of every two treated. The comparison suggests that the MFP/Ca with HRT combination could be more effective than MFP/Ca alone in protecting from NewVF and from Non-VF, justifying further double blind, prospective randomized studies pursuing this investigation. PMID- 12120909 TI - Comparison of open carpal tunnel release and local steroid treatment outcomes in idiopathic carpal tunnel syndrome. AB - To compare the efficacy of local steroid injection and open carpal tunnel release, a symptom and functional status questionnaire (Boston Questionnaire) and sensory and motor nerve conduction studies were performed in 90 patients with electrophysiologically proven idiopathic carpal tunnel syndrome, of whom 44 were treated surgically and 46 by two-dose steroid injection. Electrophysiologic studies and the Boston Questionnaire were applied before and at the 3rd and 6th months after treatment. Both groups showed significant improvement at first follow-up. The surgically treated group showed a significant and further improvement of symptoms and conduction values between the 3rd- and 6th-month evaluations, whereas no significant change was observed in the patient group treated by steroid injection. By the end of follow-up, 5% of the hands in the open carpal tunnel release (OCTR) group and 13% of the hands in the local steroid injection (LSIG) group showed electrophysiological worsening, and 5% of the hands in the OCTR group and 22% of the hands in the LSIG group showed symptomatic worsening. Our results show that steroid injection provides an improvement comparable with that from surgical release of the median nerve at a 3-month interval. However, this improvement is not long-lasting. PMID- 12120910 TI - Posterior tibial tendon dysfunction and MR imaging in rheumatoid arthritis. AB - We present the case of a patient with long-standing rheumatoid arthritis and an acute onset of total dysfunction of the posterior tibial tendon. On MRI, a rupture of the tendon was apparent. Intraoperatively, however, massive tenosynovitis with stricture of the tendon was identified as the cause of posterior tibial tendon dysfunction. This case illustrates a pitfall in MRI imaging with potential diagnostic and therapeutic consequences. PMID- 12120912 TI - Serum beta 2-microglobulin reflects disease activity in Behcet's disease. AB - Behcet's disease (BD) is a systemic remitting vasculitis characterized by orogenital ulceration and uveitis. The disease is not associated with specific laboratory abnormalities. Hence, the activity of BD is generally assessed by clinical findings and--to some extent--by nonspecific markers of inflammation. This study was performed to investigate the relative efficiency of erythrocyte sedimentation rate (ESR), C-reactive protein (CRP), serumamyloid A protein (SAA), and beta 2-microglobulin (beta 2-m) levels as markers of disease activity in patients with BD. The study population consisted of 20 patients with active BD, 23 patients with inactive BD, and 27 healthy adults serving as the control group. Serum beta 2-m, SAA, ESR, and CRP levels of patients with BD were found to be significantly higher than those in the healthy control group. They were also higher in patients with active disease than in those in remission and controls. The levels of SAA. ESR, and CRP in inactive patients were also significantly higher than the controls. No statistically significant difference was noted between beta 2-m levels of patients with inactive BD and healthy controls. Serum beta 2-m levels can beregarded as a more discriminative marker of activation in BD. The high diagnostic value of SAA levels indicate that it can also be accepted as a marker of disease activity in BD. PMID- 12120911 TI - Localization of extrapulmonary tuberculosis in the synovial membrane, skin, and meninges in a patient with systemic lupus erythematosus and IgG deficiency. AB - We report on a 31-year-old female patient with systemic lupus erythematosus (SLE) for 24 years who had a past history of skin tuberculosis (lupus vulgaris), long term corticosteroid therapy, and IgG deficiency. She presented with monoarthritis and concomitant meningitis from skin tuberculosis after 5 years. The diagnosis of joint and meningeal tuberculosis was defined with clinical symptoms--signs and typical histopathological findings of involved synovium. Clinical improvement was achieved with antituberculous therapy. Cutaneous, articular, and cerebral manifestations of tuberculosis might have been confused with some of the lupus manifestations or lupus activation. It should be kept in mind that tuberculosis may be encountered in SLE due to the nature of the underlying disease and/or its therapy. It is also worth mentioning that, in this patient, tissues involved with extrapulmonary tuberculosis were the primary areas of involvement with SLE. PMID- 12120913 TI - Hemostatic parameters in Behcet's disease: a reappraisal. AB - Behcet's disease (BD) is a relapsing vasculitis in which orogenital ulceration is a prominent feature. The disease presents a tendency to thrombosis. The prevalence of venous or arterial thrombosis in BD reaches 40% in some series. Molecular markers of imbalance have been extensively studied in BD. The aim of this paper is to review current data on essential hemostatic parameters. The precise pathogenetic mechanism(s) underlying the prothrombotic state of BD is unknown. Thrombophilic factors could contribute to thrombosis in BD. Vasculitic endothelial injury may trigger or enhance the pathological hemostatic process. PMID- 12120914 TI - Acrochordon and impaired carbohydrate metabolism. AB - Acrochordons were reported to have a probable association with diabetes mellitus but detailed data about this relation has not been introduced yet. We evaluated 120 patients with acrochordon for the presence of impaired carbohydrate metabolism. Overt diabetes mellitus (DM) was found in 88 patients and glucose intolerance was detected in 6 patients; 4 patients had reactive hypoglycemia. We concluded that acrochordons may be skin markers of underlying impaired carbohydrate metabolism and the patients with acrochordon should be evaluated for the presence of diabetes mellitus. PMID- 12120915 TI - Involvement of the skeletal GH-IGF system in an experimental model of diabetes induced growth retardation. AB - Uncontrolled diabetes is associated with growth retardation. We investigated the effect of insulin-dependent diabetes on animal growth and IGF-I gene expression in the epiphyseal growth plate region of the long bones. We also studied the effect of GH administration on somatic growth in the diabetic state. Streptozotocin (STZ)-injected diabetic rats had a decreased somatic growth rate in comparison to controls. GH administration (2.5 U/kg day) in the diabetic animals (DGH group) prevented this decrease. Serum IGF-I levels were decreased in both diabetic and DGH animals. Within 72 h from diabetes onset, IGF-I mRNA levels in epiphyseal growth plate homogenates decreased whereas IGF-I receptor mRNA levels increased in diabetic animals. The decrease in IGF-I mRNA transcript levels was localized to the metaphyseal region by in situ hybridization. We conclude that in the STZ-induced diabetic state, the reduction in linear growth is associated with a parallel decrease in IGF-I gene expression at the epiphyseal growth plate area. Diabetic growth retardation can be reversed with GH administration, which does not reconstitute serum IGF-I levels. Therefore, we speculate that GH in this model may act locally through the skeletal GH-IGF-I system. PMID- 12120916 TI - Universal screening and intensive metabolic management of gestational diabetes: cost-effectiveness in Italy. AB - This study retrospectively evaluated two groups of pregnant women. Group A women (n=1,338) were universally screened for gestational diabetes mellitus (GDM) and GDM patients were intensively treated. In Group B (n=4,035), screening was performed only in women at high risk for GDM and treatment was conventional. This study confirms the validity of a cost-effective screening program for the diagnosis of GDM and that selective screening may be an option only in a situation where healthcare resources are very scarce and/or universal screening of any kind is not feasible. Once the diagnosis of GDM has been made, metabolic management with an intensive approach is important to reduce maternal and fetal morbidity. Diagnosis of GDM and intensive treatment represent a cost for the public health system, but permit a significant monetary savings in terms of costs linked to maternal and neonatal morbidity. PMID- 12120917 TI - Effect of increased glucose load on maternal-fetal transport of alpha aminoisobutyric acid in the perfused human placenta: in vitro study. AB - The role of hyperglycemia on modulation of maternal-fetal transport of amino acids in humans is little understood. Hence, we have explored the effect of increased glucose load on transport kinetics of a model non-metabolizable amino acid, alpha-aminoisobutyric acid (AIB), in the human placenta in vitro. Transport kinetics of AIB in maternal-fetal direction was studied using perfusion of isolated human placental lobules. NCTC (National Culture and Tissue Collection) 135 medium, diluted with Earle's buffered salt solution was used as the perfusate and tritiated water was used as the reference marker. Effect of increased glucose load on transport kinetics of study and reference substances was studied in normal term placentae (n=5; gestational age, 38.5 +/- 0.5 weeks) in succeeding experimental phases, after a control perfusion phase with physiological glucose concentration. AIB transport fraction (TF), relative to tritiated water TF, averaged 54.8% in control euglycemic phase while in hyperglycemic concentration phases of 27.8 and 55.6 mM, the AIB TF index averaged 42.4% and 38.2%, respectively. Analysis of variance revealed that the difference was statistically significant. Similarly, absorption rate index of the amino acid was also significantly lower in the hyperglycemic perfusion phases compared to control euglycemic phase. We conclude that hyperglycemia may play a deleterious role in limiting maternal-fetal transport of A-type amino acids in the in vivo state. PMID- 12120918 TI - Additive effect of overweight and type 2 diabetes in the appearance of coronary heart disease but not of stroke: a cross-sectional study. AB - We evaluated the possible additive effect of overweight and diabetes in the occurrence of coronary heart disease (CHD) and stroke, and their interaction with other established risk factors. In a cross-sectional study, we evaluated the frequency of CHD and stroke in four groups of subjects: (1) lean non-diabetic subjects (n=250); (2) lean diabetic subjects (n=269); (3) overweight non-diabetic subjects (n=203); and (4) overweight diabetic subjects (n=446). CHD was more frequent among diabetic subjects, and even more among overweight diabetic subjects; stroke was more frequent among diabetic subjects, but equally frequent in overweight and in lean diabetic subjects. At multiple logistic regression analysis, age, arterial hypertension, diabetes were independent risk factors for CHD and for stroke; BMI and hyperlipidemia were risk factors only for CHD. CHD was an independent risk factor for stroke, and stroke was a risk factor for CHD. We conclude that obesity and diabetes are additional risk factors for CHD but not for stroke. The value of established risk factors such as arterial hypertension and hyperlipidemia in determining the appearance of CHD and stroke is maintained in the presence overweight and diabetes. Finally, CHD is frequently associated with stroke, suggesting a common process of atherosclerosis underlying both diseases. PMID- 12120919 TI - Early effects of diabetes on inducible nitric oxide synthase in the kidney. AB - NO may be responsible for the glomerular hyperfiltration observed in diabetic kidney by inducing vasodilation of the afferent arteriole. The aim of this study was to evaluate which isoform of nitric oxide synthase (NOS) is responsible for increased renal production of NO in diabetic kidney. Thirty male WKY rats were divided into 6 groups. Five rats were sacrificed immediately, five after 20 days. In the other rats, diabetes was induced by streptozotocin. The four diabetic groups were sacrificed respectively after 5, 10, 15 and 20 days. Urine excretion of NO metabolites was assayed; immunochemistry showed the presence of inducible (iNOS) and endothelial constitutive (ecNOS) synthases in the kidney. Urinary excretion of NO metabolites increased significantly in diabetic rats five days after the induction of diabetes and at the end of the study whereas it was unchanged in the control group. Renal ecNOS remained unchanged throughout the study in all rats whereas iNOS increased significantly in diabetic rats from the fifth day until the end of the study. The results demonstrate that iNOS is activated in the kidney of rats, soon after the induction of diabetes, thus suggesting its involvement in the increased production of NO observed immediately after the onset of diabetes. PMID- 12120920 TI - Blood glucose self-monitoring from abdominal skin: a precise and virtually pain free method. AB - For many diabetic patients, years of blood glucose self-monitoring (SM) with readings taken several times daily is an inevitable aspect of insulin therapy. We investigated whether SM from abdominal skin might be an alternative to the established fingertip method. A total of 63 diabetic patients and 16 nondiabetic volunteers determined their blood glucose in parallel in capillary blood from the tip of the finger and from abdominal skin 5 times daily on 5 successive days. The blood samples were collected from the two test regions using lancing devices, and the SM determinations were all done with a meter. Consecutive specific enzymatic glucose determinations in blood from the fingertip served as the reference method. The results of the SM from abdominal skin, a method perceived as virtually painless, were in close correlation with the control laboratory determinations and with SM from the finger (Pearson's r, 0.94 and 0.95). The comparison of SM method for abdomen vs. finger laboratory control gave a linear regression equation of y=8.35+0.94x (r=0.94). Error grid analysis revealed: range A, 93.6%; range B, 5.4%; range C, 0.05%; range D, 1.0%; and range E, 0%. Bland and Altman analysis yielded the mean of the differences, 0.2 mg/dl; 2 SD, 32 mg/dl; minimum, -162 mg/dl; maximum, 148 mg/dl. Laboratory glucose determinations in capillary blood from the fingertip and from abdominal skin led in 99.7% of the cases to concordant therapeutic decisions in the diabetics; the sample material was therefore equivalent. The practical aspects (afterbleeding, number of punctures, test strip consumption) of SM from the two regions showed no essential differences. However, only 22% of the diabetic patients investigated continued to perform SM from abdominal skin on a longer basis. In a further 5 adipose diabetic patients (BMI, 32 kg/M2), SM from abdominal skin was not practicable, as there was insufficient blood to collect. SM from abdomal skin is a simple, virtually pain-free and precise method. It provides certain diabetic patients with an alternative to the established method of SM from the fingertip. PMID- 12120921 TI - NAO reports on successes and shortcomings in controlling FMD. National Audit Office. PMID- 12120922 TI - Trilostane treatment of 78 dogs with pituitary-dependent hyperadrenocorticism. AB - The efficacy of trilostane in the treatment of canine pituitary-dependent hyperadrenocorticism (PDH) was evaluated in 78 dogs with the condition which were treated for up to three years. The drug appeared to be well tolerated by almost all the dogs, and only two developed clinical signs and biochemical evidence of hypoadrenocorticism. Polyuria and polydipsia completely resolved in 70 per cent of the dogs that had these problems, and skin changes resolved in 62 per cent of the dogs that had skin abnormalities. There was a significant reduction (P<0.001 in each case) in both the mean basal and post-adrenocorticotrophic hormone (ACTH) cortisol concentrations after a mean of 12.3 days of treatment. The post-ACTH cortisol concentration decreased to less than 250 nmol/litre in 81 per cent of the dogs within one month of the start of treatment and in another 15 per cent at some later time. The median survival time of the 26 dogs which died was 549 days, and 51 of the dogs were alive at the completion of the study. One was lost to follow up after 241 days treatment. PMID- 12120923 TI - Evaluation of aerosol transmission of porcine reproductive and respiratory syndrome virus under controlled field conditions. AB - The aim of this study was to determine whether porcine reproductive and respiratory syndrome virus (PRRSV) could be transmitted by aerosol under field conditions. A total of 210 five-month-old PRRSV-negative pigs were housed in a mechanically ventilated finishing facility containing 11 pens. Pen 1 contained 10 pigs (indirect contact controls) and pen 2 remained empty, providing a barrier of 2.5 m from the remaining pigs in pens 3 to 11. Fifteen or 16 of the pigs in each of pens 3 to 11 were infected experimentally with a field isolate of PRRSV and the other six or seven pigs served as direct contact controls. Five days after the pigs were infected, two trailers containing 10 five-week-old PRRSV-naive sentinel pigs were placed along each side of the building; one was placed 1 m from the exhaust fans on one side of the building, and the other was placed 30 m from the fans on the other side, and the sentinel pigs remained in the trailers for 72 hours. They were then moved to separate buildings on the same site, 30 and 80 m, respectively, from the infected barn, and their PRRSV status was monitored for 21 days. The direct and indirect contact control pigs became infected with PRRSV but the sentinel pigs did not. PMID- 12120924 TI - Degrees of aversion shown by rats and mice to different concentrations of inhalational anaesthetics. AB - The distress associated with the induction of anaesthesia with halothane, isoflurane, enflurane and carbon dioxide was investigated in rats and mice by measuring the level of aversion they displayed on exposure to low, medium and high concentrations of these agents. The animals were exposed to each agent in a test chamber containing air or gas mixtures, which they were able to enter and leave at will, and the level of aversion was assessed in terms of the initial withdrawal and total dwelling times in the chamber. Comparisons between the anaesthetic and air-control treatments indicated that concentrations of the agents recommended for the rapid and efficient induction of anaesthesia were associated with some degree of aversion. Carbon dioxide was by far the most aversive gas for both rats and mice, with the least aversive being halothane for rats, and halothane and enflurane for mice. With all the anaesthetics, the level of aversion increased as the concentration increased. PMID- 12120925 TI - Reporting of sheep lameness conditions to veterinarians in the Scottish borders. PMID- 12120926 TI - Thromboembolic meningoencephalitis due to Haemophilus somnus in feedlot cattle in Argentina. PMID- 12120927 TI - Modelling and FMD. PMID- 12120928 TI - Staphylococcus aureus septicaemia in a killer whale. PMID- 12120929 TI - Equine viral arteritis in the UK. PMID- 12120930 TI - RCVS fees for 2003. PMID- 12120931 TI - RCVS fees for 2003. PMID- 12120932 TI - Tail docking of dogs. PMID- 12120933 TI - Perioperative antibiotic prophylaxis--still room for improvement. PMID- 12120934 TI - Vancomycin-resistant Staphylococcus aureus: a real and present danger? AB - The glycopeptide antibiotics, vancomycin and teicoplanin, are the mainstay of therapy for infections involving strains of Staphylococcus aureus that are resistant to methicillin and gentamicin. Durng the last 5 years, clinical isolates of S. aureus showing reduced susceptibility to glycopeptides have been reported from many countries around the world, often associated with prolonged glycopeptide therapy. Detection and monitoring of such strains has been hindered by the fact that vancomycin (or glycopeptide)-intermediate S. aureus (VISA) isolates may be missed on conventional disk sensitivity tests. Effective control measures are required to prevent the increasing occurrence and spread of such strains in both the hospital and community settings. An important aspect of control is promoting the judicious use of glycopeptides. The recent introduction of the alternative antibiotics quinupristin/dalfopristin and linezolid, which are active against S. aureus strains resistant to many other classes of agent, should facilitate this process. PMID- 12120935 TI - Klebsiella bacteremia in children in southern Israel (1988-1997). AB - BACKGROUND: Klebsiella spp. have emerged in recent years as a major cause of gram negative bacteremia in infants and children. We therefore aimed to document the epidemiology, antibiotic susceptibility pattern and outcome of both community acquired and nosocomial Klebsiella spp. bacteremias in children. PATIENTS AND METHODS: From 1988-1997, 177 episodes of Klebsiella bacteremia, representing 15% of all gram-negative bacteremias, occurred at the Soroka Medical Center in 166 children aged 0-14 years. RESULTS: The overall incidence of Klebsiella bacteremia in southern Israel during the study period was 0.13/1,000, with an increase from 0.1 to 0.2/1,000 children from 1988-1992 to 1993-1997 (p = 0.02). 113 and 64 episodes were recorded in Bedouin Arabs and Jewish children, respectively. The incidence of Klebsiella bacteremia was significantly higher in Bedouins compared to Jewish children (p < 0.001). The incidence of Klebsiella bacteremia increased significantly among Jewish children from 1993-1997 compared to 1988-1992. The incidence of Klebsiella bacteremia was 2/1,000 admissions, with an increase from 1.8 to 2.2/1,000 from 1993-1997 compared to 1988-1992. The incidence of Klebsiella bacteremia was significantly higher among hospitalized Bedouin children compared to Jewish children (3.1 vs 1.4/1,000 admissions, p < 0.001). There were 48 (27%), 24 (14%) and 98 (55%) Klebsiella bacteremia episodes at the pediatric departments, pediatric intensive care unit (PICU) and neonatal intensive care unit (NICU), respectively. 76% of Klebsiella bacteremia episodes were nosocomial; 66% occurred at NICU. 71% and 90% of Klebsiella bacteremia episodes occurring at NICU and PICU, respectively, were nosocomial. The overall incidence of nosocomial infections was 1.5/1,000 admissions, with an increase from 1.2 to 1.8/1,000 from 1993-1997 compared to 1988-1992 (p = 0.03). The resistance rates of Klebsiella spp. to piperacillin, ceftriaxone, ceftazidime and gentamicin were 34%, 17%, 17% and 14%, respectively. A significant increase in the resistance rates to ceftriaxone and ceftazidime was observed from 1993-1997 compared to 1988-92 (21.9% vs 7.8%, p = 0.05 and 21.9% vs 5%, p = 0.03). A significant increase in resistance to ceftriaxone was recorded at PICU and NICU (from 12% and 0%, respectively, from 1988-1992, to 61% and 16%, respectively, from 1993-1997, p = 0.02). Overall mortality rate of Klebsiella bacteremia was 13% (21/167 cases, 12 and eight at PICU and NICU, respectively). CONCLUSION: An increase in Klebsiella bacteremia was recorded in southern Israel during the 10 years of the study. A marked increase in the rate of nosocomial Klebsiella bacteremia occurred at all departments. Resistance to third-generation cephalosporins emerged frequently at PICU and NICU during the last period of the survey. PMID- 12120936 TI - Clinical dysentery in hospitalized children. AB - BACKGROUND: Clinical dysentery is a severe presentation of an enteric infection. The aim of the study was to evaluate the impact of a serious bacterial etiology in clinical dysentery in hospitalized children and determine if children at high risk can be identified on the basis of clinical or laboratory parameters. PATIENTS AND METHODS: A prospective study design was used. The study population included 60 children admitted to our department with clinical dysentery over a 16 month period. Fresh stool specimens were collected on days 1, 2 and 3. The clinical and laboratory data of the children were analyzed. RESULTS: Clinical dysentery accounted for 1.7% of all pediatric hospitalizations during this period. Stool cultures were positive for Shigella spp. in 18 children (30%), and Salmonella spp. in 15 children (25%), Campylobacter jejuni was identified in one patient (2%). There were no significant differences in clinical characteristics or laboratory parameters between children with positive and negative stool cultures. CONCLUSION: 40% of the children hospitalized for clinical dysentery were eligible for antibiotic treatment. Early administration of empiric antibiotic treatment is justified in children hospitalized for clinical dysentery in Israel. Clinical or laboratory parameters were unable to differentiate those with clinical dysentery at risk of serous bacterial pathogens in stool. PMID- 12120937 TI - Hepatitis B vaccination: long-term follow-up of the immune response of preterm infants and comparison of two vaccination protocols. AB - BACKGROUND: We conducted a 3-year follow-up study of long-term antibody persistence following vaccination of low-risk preterm infants with recombinant hepatitis B vaccine (HBV). Two three-dose protocols were compared: vaccination beginning within 24 h of birth to initial vaccination delayed until a weight of 2,000 g was reached. SUBJECTS AND METHODS: The study population included 136 children, divided into three groups: children born prematurely (< or = 35 weeks, n = 57), children born at term (> or = 37 weeks, n = 39), both groups receiving the first dose of HBV within 24 h of birth, and children born prematurely (< or = 35 weeks, n = 40), who received the first dose of HBV when a weight of 2,000 g was reached. All infants received the second hepatitis vaccination 1 month after the first, and the third dose 6 months after the first. Hepatitis B surface antibody (AntiHBs) was measured at an age of 3-3.5 years (at least 2.5 years after completion of the three-dose HBV series). An AntiHBs level of > or = 10 IU/l was considered positive. RESULTS: At 3-3.5 years of age, a higher percentage of the premature-delayed vaccination group had a positive AntiHBs level (92.5%) compared to both the premature (54.4%, p < 0.001) and full-term groups (71.8%, p < 0.05) vaccinated soon after birth. The premature-delayed vaccination group also had a significantly higher geometric mean concentration (GMC) (119 vs 14.2 IU/l, p < 0.001 and 119 vs 32.7 IU/l, p < 0.005, respectively). CONCLUSION: Delaying vaccination of premature infants against hepatitis B until a weight of 2,000 g was reached resulted in both a significantly higher percentage of children with positive antibody levels and a significantly higher GMC at 3-3.5 years of age as compared to early-vaccinated preterm and full-term infants. The known short-term advantage of delayed vaccination of preterm infants was shown to persist for at least the first 3 years of life. PMID- 12120938 TI - Hepatitis D virus infection in Thailand: HDV genotyping by RT-PCR, RFLP and direct sequencing. AB - BACKGROUND: Hepatitis D virus (HDV) is a degenerate RNA virus or virusoid that requires the surface coat of hepatitis B virus (HBV), i.e. hepatitis B surface antigen (HBsAg), in order to become infectious. Three distinct genotypes of the virus have been classified. In this study, HDV genotypes were determined by restriction fragment length polymorphism (RFLP) and direct sequencing. In Thailand, simultaneous HDV/HBV infections are particularly prevalent among intravenous drug users (IVDU). PATIENTS AND METHODS: A total of 743 IVDU sera were screened for HBV infection. HBsAg-positive samples were subjected to serological analysis for anti-HDV. RFLP analysis using the endonucleases Xho I and Sma I was performed on the PCR amplified HDV genome to establish the prevailing HDV genotypes. RESULTS: 55 sera (7%) had detectable HBsAg; all 55 were subsequently subjected to serological analysis for anti-HDV, 12 (21.8%) of which were positive. Eight (66%) specimens had detectable HDV-RNA by RT-PCR. All polymorphisms were shown to be genotype I, a finding confirmed by direct sequencing. 36 HBsAg-positive sera obtained from the blood bank to serve as controls were negative for anti-HDV. CONCLUSION: Our data show that HDV infection is still limited among IVDU and that the pattern of polymorphism closely resembles that of the western HDV genotype I. PMID- 12120939 TI - Multiple sclerosis and human herpesvirus 6. AB - BACKGROUND: A possible but as yet unproven relationship has been proposed between the onset or persistence of multiple sclerosis (MS) symptoms and herpesviruses, including, most recently, human herpesvirus 6 (HHV-6). A study was conducted to investigate the presence of HHV-6 DNA and the synthesis of antibodies against HHV 6, cytomegalovirus (CMV) and Epstein-Barr virus (EBV) in serum and cerebrospinal fluid (CSF) of patients with MS. MATERIALS AND METHODS: PCR and ELISA were used to detect HHV-6 DNA and specific antibodies against HHV-6, CMV and EBV in 211 samples (139 sera and 72 CSF). There were three groups of samples: group I, paired samples of serum and CSF from 41 MS patients; group II, paired samples of serum and CSF from 31 patients with neurological diseases other than MS (OND); group III, 67 serum samples from 27 different MS patients undergoing serologic follow-up. RESULTS: No HHV-6 DNA was found in any sample. Group I sera showed elevated anti-HHV-6 IgG and IgA levels. In group II, anti-CMV IgG was detected in one CSF sample and anti-HHV-6 IgM in one serum sample. Group III sera showed high concentrations of anti-HHV-6 IgG, IgA and IgM. CONCLUSION: Given the clinical implications of the presence of antibodies against HHV-6 in MS patients, a viral reactivation cannot be excluded as an environmental factor. PMID- 12120940 TI - Effect of dexamethasone and pentoxifylline in combination with amoxicillin in the treatment of penicillin-insensitive pneumococcal pneumonia in guinea pigs. AB - A fatal guinea pig model of pneumococcal pneumonia was developed in order to evaluate the efficacy of amoxicillin combined with either pentoxifylline or dexamethasone. Parameters assessed were survival time and lung changes (alterations, bacterial colony counts, inducible nitric oxide synthase [iNOS] and cyclooxygenase-2 [COX-2] protein expression). Animals receiving pentoxifylline (50 mg/kg) showed higher survival rates than controls (p < 0.05). Animals which received amoxicillin (50 mg/kg), alone or combined, showed significantly higher survival rates than controls (p < 0.05). Animals dying in spite of receiving amoxicillin alone or combined had lung colony counts significantly lower than those that did not receive the antibiotic (p < 0.001), but their lungs showed identical changes. The correlation between COX-2 protein expression and mortality was rather high (r = 0.75). The addition of either dexamethasone or pentoxifylline to amoxicillin improved neither survival rates nor lung pathology when compared with the antibiotic alone. PMID- 12120941 TI - Infection of the finger in a general pediatrician. PMID- 12120942 TI - The fecal microflora of 1-3-month-old infants during treatment with eight oral antibiotics. AB - We examined the fecal microflora of 1-3-month-old infants during treatment with phenoxymethylpenicillin, amoxycillin, pivampicillin, cefaclor, cefadroxil, loracarbef, erythromycin or cotrimoxazole. Escherichia coli increased during treatment with penicillins or cephalosporins, but was not affected by erythromycin or cotrimoxazole. Other enterobacteria were acquired or increased during treatment with all agents except cotrimoxazole. Enterococci persisted or increased during phenoxymethylpenicillin, cephalosporin or cotrimoxazole treatment, whereas erythromycin and the other penicillins suppressed them. Bacteroides, bifidobacteria and lactobacilli were suppressed to undetectable levels in most infants during treatment with all agents, except phenoxymethylpenicillin and loracarbef. PMID- 12120943 TI - Combination of quinupristin/dalfopristin and glycopeptide in severe methicillin resistant staphylococcal infections failing previous glycopeptide regimens. AB - BACKGROUND: We report our experience with quinupristin/ dalfopristin in combination with a glycopeptide in the treatment of severe staphylococcal infections failing previous glycopeptide regimens. PATIENTS AND METHODS: Five patients, affected by persistent bacteremia (n = 2), post-cardiothoracic surgery infection (n = 2) and post-traumatic bone infection (n = 1) due to methicillin resistant Staphylococcus aureus (MRSA, n = 4) methicillin-resistant coagulase negative Staphylococcus (MRCNS, n = 1) and unsuccessfully treated with antibiotics including a glycopeptide, were treated with a quinupristin/ dalfopristin and glycopeptide combination. RESULTS: Three patients were clinically cured; one patient with MRSA thoracic aorta prosthetic infection relapsed after 3 months; one patient was lost to follow-up. CONCLUSION: Quinupristin/dalfopristin, in combination with a glycopeptide, is an effective treatment option for severe methicillin-resistant staphylococcal infections failing previous glycopeptide regimens. PMID- 12120944 TI - The practice of perioperative antibiotic prophylaxis in eight German hospitals. AB - BACKGROUND: Although there is consensus in the international literature on the benefits of using perioperative antibiotic prophylaxis (PAP), there is still considerable scope for improving its use in many hospitals. MATERIALS AND METHODS: In this study, data on the practice of PAP were recorded in eight German hospitals within the framework of a prospective controlled interventional study for the surveillance and prevention of nosocomial infections. RESULTS: A total of 627 surgical procedures (appendectomies, other colorectal procedures, total prosthetic hip replacement) were assessed; 397 with PAP and 224 without PAP; six procedures could not be evaluated. Of the 397 PAP recorded, only 180 (45.3%) were performed correctly in accordance with international standards as a preoperative single dose (19/59 PAP in appendectomies, 72/188 PAP in other colorectal procedures, 89/150 PAP in total prosthetic hip replacement). CONCLUSION: There is still great uncertainty regarding the point in time at which PAP should be administered and its duration. Additional efforts are necessary to improve PAP in accordance with published evidence-based guidelines. PMID- 12120945 TI - Laryngopharyngitis by Corynebacterium ulcerans. AB - A 71-year-old female patient was hospitalized with membranous laryngopharyngitis typical of classical diphtheria. A toxigenic strain of Corynebacterium ulcerans was isolated from the throat. The patient was treated for 6 days with amoxicillin clavulanic acid and recovered without complications. This second reported case of diphtheric laryngopharyngitis caused by C. ulcerans in Switzerland is a reminder that C. ulcerans should be included as a possible agent in patients with classical diphtheria symptoms. PMID- 12120946 TI - Recurrent septicemia due to Campylobacter fetus and Campylobacter lari in an immunocompetent patient. AB - We describe a severe and recurrent septicemia due to Campylobacter in a 75-year old immunocompetent patient. Two Campylobacter strains were detected in several blood cultures. Campylobacter fetus and Campylobacter lari were identified with PCR tests based on species-specific nucleotide sequences for the 16S rRNA gene. PMID- 12120947 TI - Brachyspira (Serpulina) pilosicoli spirochetemia in an immunocompromised patient. AB - The case of an elderly immunocompromised man with non-Hodgkin's lymphoma who presented with fever, abdominal pain and bloody diarrhea is described. Brachyspira pilosicoli was isolated from culture. The patient was treated with penicillin G i.v. and became afebrile. B. pilosicoli is a recently recognized enteric pathogen of humans and animals. Intestinal spirochetosis should be included in the differential diagnosis of any immunocompromised or critically ill patient with dysentery. PMID- 12120948 TI - Actinomyces neuii and neonatal sepsis. AB - Actinomyces neuii belongs to the coryneform bacteria. In the case presented here this gram-positive rod had to be considered the pathogen causing not only the chorioam nionitis but also the neonatal sepsis. Conventional therapeutic regimes are effective due to the high sensitivity of A. neuii to beta-lactam antibiotics. PMID- 12120949 TI - Methicillin-resistant Staphylococcus aureus meningitis: has the time come for an alternative to vancomycin? PMID- 12120950 TI - Stereochemistry of the hydroperoxides formed during autoxidation of CLA methyl ester in the presence of alpha-tocopherol. AB - The initial steps in the autoxidation of CLA methyl ester are poorly understood. The aim of this study was to determine the stereochemistry of the hydroperoxides formed during autoxidation of CLA methyl ester in the presence of a good hydrogen atom donor. For this purpose, 9-cis,11-trans CLA methyl ester was autoxidized in the presence of alpha-tocopherol under atmospheric oxygen at 40 degrees C in the dark. The CLA methyl ester hydroperoxides were isolated, reduced to the corresponding hydroxy derivatives, and separated by HPLC. The stereochemistry of seven hydroxy-CLA methyl esters was investigated. The position of the hydroxy group was determined by GC-MS. The geometry as well as the position of the double bonds in the alkyl chain was determined by NMR. In addition, the 13C NMR spectra of six hydroxy-CLA methyl esters were assigned using COSY, gradient heteronuclear multiple bond correlation, gradient heteronuclear single quantum correlation, and total correlation spectroscopy experiments. The autoxidation of 9-cis,11-trans CLA methyl ester in the presence of a good hydrogen atom donor is stereoselective in favor of one geometric isomer, namely the 13-(R,S)-hydroperoxy-9-cis,11-trans octadecadienoic acid methyl ester. Three types of conjugated diene hydroperoxides are formed as primary hydroperoxides: trans,trans hydroperoxides (12-OOH-8t,10t and 9-OOH-10t,12t), a cis,trans hydroperoxide with the trans double bond adjacent to the hydroperoxide-bearing carbon atom (13-OOH-9c,11t), and a new type of cis,trans lipid hydroperoxide with the cis double bond adjacent to the hydroperoxide-bearing carbon atom (8-OOH-9c,11t). In addition, three nonkinetic hydroperoxides (13-OOH-9t,11t, 8-OOH-9t,11t, and 9-OOH-10t,12c) are formed. This study supports the theory that CLA methyl ester autoxidizes at least partly through an autocatalytic free radical reaction. The complexity of the hydroperoxide mixture is due to formation of two different pentadienyl radicals. Moreover, the stereoselectivity in favor of one geometric isomer can be explained by the selectivity of the two previous steps: the preferential formation of a W conformer of the pentadienyl radical over the Z-conformer, and regioselectivity of the oxygen addition to the pentadienyl radical. PMID- 12120951 TI - Regiospecific enzymatic oxygenation of cis-vaccenic acid during aerobic senescence of the halophilic purple sulfur bacterium Thiohalocapsa halophila. AB - A regiospecific oxygenation of the allylic carbon 10 of cis-vaccenic acid has been observed in senescent cells of the halophilic purple sulfur bacterium Thiohalocapsa halophila incubated under aerobic conditions in darkness. The results obtained strongly suggest that these enzymatic processes involve the initial dioxygenase-mediated formation of 1 O-hydroperoxyoctadec-cis-11-enoic acid, which is not accumulated in the cells of T. halophila owing to its high cytotoxic properties. Deuterium labeling and GC-MS analyses enabled us to demonstrate that subsequent enzymatic conversions of this allylic hydroperoxide involved reduction, cleavage, isomerization, and saturation reactions. Some of the specific oxidation products thus formed could constitute potential T. halophila biomarkers. PMID- 12120952 TI - Dietary fish oil and vitamin E enhance doxorubicin effects in P388 tumor-bearing mice. AB - In this study, four kinds of rodent diets, CO, FO, CVe, and FVe, were used by addition of canola oil, oil mixture (fish oil + canola oil), canola oil plus vitamin E, and oil mixture plus vitamin E, respectively, to a basic diet, AIN 93G, to investigate the influence of dietary fish oil and vitamin E on doxorubicin (DOX) treatment in P388 ascitic mice. Animal life span (LS) and heart damage were recorded in mice fed the four different diets and treated with different doses of DOX. The optimal doses of DOX for antitumor effect as manifested by increased LS were 6.0 and 9.0 mg/kg. Both fish oil and vitamin E significantly enhanced this effect. On the other hand, DOX at 12.0 mg/kg induced severe heart damage, which was also significantly aggravated by both fish oil and vitamin E, as shown by both decreased LS and increased serum creatine phosphokinase activity. Fish oil and vitamin E appeared to enhance the antitumor effect of optimal doses of DOX but to aggravate cardiotoxicity owing to DOX overdose. PMID- 12120953 TI - Effect of palm oil carotene on breast cancer tumorigenicity in nude mice. AB - Biological therapies are new additions to breast cancer treatment. Among biological compounds, beta-carotene has been reported to have immune modulatory effects, in particular, enhancement of natural killer cell activity and tumor necrosis factor-alpha production by macrophages. The objective of this study was to investigate the effect of palm carotene supplementation on the tumorigenicity of MCF-7 human breast cancer cells injected into athymic nude mice and to explore the mechanism by which palm carotenes suppress tumorigenesis. Forty-eight 4-wk old mice were injected with 1 x 10(6) MCF-7 cells into their mammary fat pad. The experimental group was supplemented with palm carotene whereas the control group was not. Significant differences were observed in tumor incidence (P< 0.001) and tumor surface area and metastasis to lung (P< 0.005) between the two groups. Natural killer (NK) cells and B-lymphocytes in the peripheral blood of carotene supplemented mice were significantly increased (P < 0.05 and P < 0.001, respectively) compared with controls. These results suggest that palm oil carotene is able to modulate the immune system by increasing peripheral blood NK cells and B-lymphocytes and suppress the growth of MCF-7 human breast cancer cells. PMID- 12120955 TI - Lauric acid is desaturated to 12:1n-3 by hepatocytes and rat liver homogenates. AB - Lauric acid desaturation was investigated and described in liver homogenates and in cultured rat hepatocytes. The identification of the desaturated product of lauric acid has been performed using the oxidative cleavage method, and we showed that the obtained monoene was mainly 12:1n-3. This result suggests that lauric acid desaturation could be the first step in the biosynthesis of alpha-linolenic acid in animal cells. PMID- 12120954 TI - Antihypertensive effects of a dietary unsaturated FA mixture in spontaneously hypertensive rats. AB - The aim of the present study was to investigate whether a mixture of dietary n-6 and n-3 PUFA could lower blood pressure in spontaneously hypertensive rats (SHR) of different ages. In addition, we studied how such a treatment could normalize the FA composition of plasma TAG and cholesterol esters (CE), and of red blood cell (RBC) total lipids. SHR (ages 4, 19, and 50 wk) were fed a normal diet (control groups) or a semisynthetic diet containing a mixture of gamma-linolenic acid (GLA), EPA, and DHA (experimental groups). Systolic blood pressure was measured at regular intervals. After 11 wk of consuming this diet, plasma TAG and CE were separated by TLC and analyzed for their FA composition. Total FA composition of RBC was also determined. The degree to which blood pressure was elevated was reduced in SHR after 11 wk of diet. The largest decrease was obtained with the oldest animals. In RBC, EPA and DHA contents increased. In plasma TAG and CE, EPA, DHA, and GLA increased whereas arachidonic acid decreased. The n-6 and n-3 unsaturated FA mix slowed the development of hypertension in young SHR and decreased blood pressure in adult and aged SHR. In addition, the present treatment altered the n-3 and n-6 PUFA content of SHR lipids to that seen in normotensive rats. PMID- 12120956 TI - LDL binding to lipid emulsion particles: effects of incubation duration, temperature, and addition of plasma subfractions. AB - Lipid emulsions used in parenteral nutrition interact with lipoproteins leading to exchanges of lipids and acquisition of several apolipoproteins (apo). It has been previously observed that, during in vitro incubation of emulsions with purified LDL, a variable fraction of LDL binds to TG-rich emulsion particles. The purpose of this study was to better characterize such an interaction. Two emulsions containing 20% soybean oil (Endolipid, B. Braun AG, Melsungen, Germany) or fish oil were incubated with LDL, either alone or in the presence of various plasma subfractions, for different durations and at different temperatures. The fraction named M-LE (containing TG-rich particles modified after incubation) was separated by ultracentrifugation or gel filtration chromatography, and the apoB content was measured as an index of LDL binding to TG-rich emulsion particles. The formation of such complexes was visualized by freeze-fracture electron microscopy. LDL binding was not influenced by the method used for M-LE isolation. Binding occurred quickly, did not increase with prolonged incubation, was inversely related to increasing incubation or ultracentrifugation temperature, and withstood 40 h of ultracentrifugation at 163,000 x g. The presence of glycerol or excess phospholipids in the emulsion did not markedly affect the formation of the complexes. In contrast, adding very small amounts of lipoprotein poor plasma (d > 1.210 g/mL) or HDL markedly reduced the process, and albumin had no effect. The TG composition of the emulsion influenced the binding of LDL to TG rich particles, since more apoB was found in M-LE from fish oil than from soybean oil emulsion. PMID- 12120957 TI - Transfer of lipids between hemolymph and hepatopancreas in the shrimp Macrobrachium borellii. AB - Crustacean lipids are transported in the hemolymph by an HDL. The hepatopancreas is the most important and active organ regarding lipid metabolism, so we studied the interchange of FA and acylglycerols between both components of the hepatopancreas-hemolymph system in the decapod crustacean Macrobrachium borellii. The hepatopancreas and a sole plasma lipoprotein were labeled by in vivo incubations with 14C palmitic acid injected into the hemolymph. Then they were incubated in vitro with unlabeled hepatopancreas and hemolymph, and the transfer of lipids between them was measured by radiochromatographic techniques. It was determined in vivo that more than 80% of the circulating palmitic acid was taken up by the hepatopancreas and incorporated into PC and TAG. Both classes of lipids, but mainly PC, were transferred back from tissues to the hemolymph. Lipid transfer was also demonstrated in vitro. The transfer of PC (30% of labeling) as well as that of FFA (48% of labeling) from hemolymph to hepatopancreas was determined. On the other hand, FFA were released more efficiently than the acylglycerols from intact hepatopancreas to hemolymph, and they were the only lipid transferred when the hepatopancreas had been previously washed. PMID- 12120958 TI - Lipid, FA, and sterol composition of New Zealand green lipped mussel (Perna canaliculus) and Tasmanian blue mussel (Mytilus edulis). AB - The lipid, FA, and sterol composition of the New Zealand green lipped mussel (NZGLM, Perna canaliculus) and of the Tasmanian blue mussel (TBM, Mytilus edulis) were compared using TLC-FID and GC-MS. The respective mussel species were obtained from three different sites in both New Zealand (NZ) and Tasmania. Lipid class distribution of both mussel species was characterized by a high proportion of phospholipid (PL, 57-79%) and TG (10-25%), FFA (7-12%), and sterols (ST, 12 18%). The NZGLM had higher proportions of TG, FFA, and ST (P < 0.01), whereas the TBM had a higher proportion of PL (P < 0.01). There were higher proportions of total PUFA, saturated FA, n-3 FA, and hydroxy and nonmethylene-interrupted FA (P < 0.05) in the TBM compared with the NZGLM. The major FA in the NZGLM were 16:0 (15-17%), 20:5n-3 (14-20%), and 22:6n-3 (11-17%). The same FA dominated lipids in the TBM, although there were significantly higher proportions of 16:0 (P = 0.000) and 22:6 n-3 (P = 0.003) and lower proportions of 20:5n-3 (P = 0.0072) in the TBM. A novel PUFA, 28:8n-3, was detected in both mussels with higher amounts in the TBM, which probably reflects a greater dietary contribution of dinoflagellates for this species. Cholesterol was the dominant sterol in both mussels. Other major sterols included brassicasterol, 22-methylcholesterol, trans 22-dehydrocholesterol, and desmosterol. There were significant differences (P < 0.05) between the NZGLM and TBM for 12 of the 20 sterols measured. Six sterols showed significant site differences for the NZGLM, and 10 for the TBM. The differences in the FA and sterol composition between the two species may be due to the diet of the NZGLM being more diatom-derived and the diet of the TBM having a greater dinoflagellate component. PMID- 12120959 TI - Two novel glucosylceramides from gonads and body walls of the Patagonian starfish Allostichaster inaequalis. AB - From the water-insoluble lipid fraction of the chloroform/methanol/water extract of the gonads and body walls of the Patagonian starfish Allostichaster inaequalis, two new glucosylceramides (4 and 7) were isolated together with the known phalluside-1 (1) and two glucosylceramides (2 and 3) previously isolated from the starfish Cosmasterias lurida. The new compounds were characterized as (2S,3R,4E,8E,10E)-1-(beta-D-glucopyranosyloxy)-3-hydroxy-2-[(R)-2-hydroxy-15 tetracosenoyl] amino-4,8,10-octadecatriene (4) and (2S,3R,4E,15Z)-1-(beta-D glucopyranosyloxy)-3-hydroxy-2-[(R)-2-hydroxyhexadecanoyl] amino-4,15-docosadiene (7) by means of spectroscopic and chemical methods. PMID- 12120960 TI - Positional distribution of CLA in TAG of lamb tissues. AB - The content and positional distribution of CLA in TAG fractions of lamb tissues was examined with either preformed CLA or the linoleic acid precursor of CLA in the diet as experimental treatments. The CLA content of phospholipid (PL) from these tissues was also examined. Thirteen lambs were randomized to the following dietary treatments: (i) control diet (no supplement); (ii) CLA supplementation (0.33 g d(-1) for 21 d prior to weaning) to milk-replacer of pre-ruminating lambs, or (iii) feeding linoleic acid-rich oil (6% safflower oil on a dry matter basis) to weaned ruminating lambs. At slaughter, tissue samples were procured from diaphragm, rib muscle, and subcutaneous (SC) adipose tissue. Safflower oil supplementation in the diet resulted in an increase in CLA content of the TAG from diaphragm, rib muscle, and SC adipose tissue by about threefold (P< 0.05) on a mol% basis. CLA was localized to the sn-1/3 positions of TAG. Animals that received pre-formed CLA, however, had increased proportions of CLA at the sn-2 position of TAG from SC adipose tissue, suggesting that there were tissue specific dietary effects and possible age-related effects on the mode of FA incorporation into TAG. Safflower oil supplementation in the diet had no effect on the CLA content of PL from diaphragm, rib muscle, and SC adipose tissue, suggesting that CLA was preferentially incorporated into the TAG of these tissues. PMID- 12120961 TI - Analysis of diastereomeric DAG naphthylethylurethanes by normal-phase HPLC with on-line electrospray MS. AB - Normal-phase HPLC resolution of sn-1,2(2,3)- and x-1,3-DAG generated by partial Grignard degradation from natural TAG was carried out with both (R)-(-) and (S) (+)-1-(1-naphthyl)ethylurethane derivatives. The diastereomeric sn-1,2- and sn 2,3-DAG derivatives were resolved using two Supelcosil LC-Si (5 microm, 25 cm x 4.6 mm i.d.) columns in series and an isocratic elution with 0.37% isopropanol in hexane at a flow rate of 0.7 mL/min. The DAG were detected by UV absorption at 280 nm and were identified by electrospray ionization MS in the positive ion mode following postcolumn addition of chloroform/methanol/30% ammonium hydroxide (75:24.5:0.5, by vol) at 0.6 mL/min. Application of the method to a stereospecific analysis of the molecular species of TAG of rat VLDL showed that the TAG composition of VLDL circulating under basal conditions differs markedly from that of VLDL secreted by the liver during inhibition of serum lipases. The inhibition of serum lipases resulted in a significant proportional decrease in 16:0 and PUFA and an increase in 18:0 and oligoenoic FA in the sn-1-position, whereas the FA compositions in the sn-2- and sn-3-positions were much less affected. PMID- 12120962 TI - Method for analysis of 4-hydroxy-2-(E)-nonenal with solid-phase microextraction. AB - A simple analytical method for 4-hydroxy-2-(E)-nonenal (HNE) using solid-phase microextraction (SPME) fiber was developed. HNE or the derivative of HNE formed by reaction with 2,4-dinitrophenylhydrazine (DNPH) was extracted from the sample solution by immersing the SPME fiber into the solution, and the amount of HNE was quantified by HPLC. The extraction conditions of HNE and HNE-DNPH were examined, using standard solutions, with respect to fiber coating, NaCl concentration, rate of stirring, adsorption temperature, and adsorption time. The recovery of HNE reached 80%, and the quantification limits of HNE and HNE-DNPH using standard compounds were 14.1 pmol/10 mL and 486.5 fmol/10 mL, respectively. This method can be applied to the detection of HNE in oxidized oil or samples of porcine liver. PMID- 12120963 TI - A critique of 50-m CP-Sil 88 capillary columns used alone to assess trans unsaturated FA in foods: the case of the TRANSFAIR study. PMID- 12120964 TI - Transsexualism/gender identity disorder. PMID- 12120965 TI - Cardiac rehabilitation service provision in Ireland: the Irish Association of Cardiac Rehabilitation survey. AB - BACKGROUND: The first national survey of cardiac rehabilitation services was conducted. AIMS: To establish levels of service provision, service formats, and geographic distribution of cardiac rehabilitation services in 1998. METHODS: Public hospitals in the Republic (n=41) and Northern Ireland (n=12) which provide services to cardiac patients were surveyed. RESULTSl Response rate was 81%. Twelve centres (29%) in the Republic and nine (75%) in Northern Ireland provided cardiac rehabilitation. There was wide geographic variability in service provision. Most centres were unable to identify the proportion of eligible patient participation. Most programmes were established for less than five years. All programmes had multidisciplinary teams, multicomponent courses and co ordinators. Additionally, 44% of hospitals without programmes had plans regarding programme establishment. CONCLUSIONS: The findings highlight the underdeveloped but rapidly expanding nature of cardiac rehabilitation services in Ireland. They provide a baseline from which to address rehabilitation needs and to judge the success of the National Cardiovascular Health Strategy in addressing these needs. PMID- 12120966 TI - Evaluation of stroke management in an Irish university teaching hospital: the Royal College of Physicians stroke audit package. AB - BACKGROUND: There are few data regarding the standard of stroke care in Ireland. AIM: To investigate the level of documentation for 13 key areas of stroke management. METHODS: Using a validated stroke audit package, this study reviewed the medical records of 100 consecutive patients hospitalised with acute stroke. RESULTS: Documentation of stroke symptoms, risk factors, general examination and investigations (cranial computer tomography [CT] and carotid Dopplers) were satisfactory. Neurological documentation was variable, with power (87%), sensation (70%) and eye movements (63%) being the most frequently recorded features, while cognition (3%), visual fields (13%), gait (7%), incontinence (1%) and swallowing (0%) were infrequently recorded. Diagnostic formulation and an acute management plan were documented in less than half of patients, whereas cranial CT (93%) and carotid Dopplers (93%) were well documented. Secondary preventive measures were documented in two-thirds of patients at follow-up. CONCLUSIONS: These results serve as a baseline from which to initiate and monitor improvements in the service at our hospital, including the involvement of neurologists in stroke care, and will also allow assessment of the impact of such changes. PMID- 12120967 TI - A prospective comparison of pedal ergometry with conventional treadmill testing in the investigation of lower extremity pain. AB - BACKGROUND: Investigation of lower extremity pain is compromised by comorbid disorders that may interfere with conventional testing. AIMS: To compare pedal ergometry with conventional treadmill testing. METHODS: A prospective study was performed where patients presenting with a diagnosis of intermittent claudication were assessed by both methods of testing. RESULTS: Of 78 patients studied with both tests, no exercise-induced ankle pressure changes occurred in 26, two were unable to complete either test despite normal pressure measurements, while 24 had exercise-induced pressure drop detected by both tests. Of patients who completed pedal ergometry, 21 were unable to complete the treadmill test, 14 of whom had negative ergometry, while seven had a pressure drop detected by pedal ergometry. Three had pressure changes with pedal ergometry, but not with treadmill testing and two had pressure changes on the treadmill not reproduced by pedal ergometry. CONCLUSIONS: Pedal ergometer is more sensitive than treadmill testing in detecting arterial insufficiency, as indicated by a 20% or greater fall in ankle pressure, and more suitable in a subgroup of patients unable to tolerate conventional treadmill testing. PMID- 12120968 TI - Inhaled nitric oxide in combination with volume resuscitation refines a porcine model of endotoxic shock. AB - BACKGROUND: Existing porcine models of endotoxic shock poorly represent the human situation. AIMS: To assess whether the cardiovascular profile of a porcine model could be improved by refining the protocol. METHODS: In 30 pigs, right and left heart pressures and cardiac output were measured. Lipopolysaccharide (LPS) was administered as a bolus (n=12), as a 30 minute infusion (n=6) or as a 30 minute infusion along with inhaled NO and volume resuscitation (n=6) and six sham treated pigs received normal saline. Haemodynamic values were measured over three hours. RESULTS: LPS increased pulmonary vascular resistance (PVR) (13.3 +/- 1.4 to 37.0 +/- 3.9kPa/l per sec, p<0.05) and reduced cardiac output (6.0 +/- 0.6 to 4.8 +/- 0.41/min). Mortality was 50% within 30 minutes. Inhaled NO and volume resuscitation controlled pulmonary vascular resistance (PVR) and preserved CO. Systemic vascular resistance (SVR) declined in the first hour (118.4 +/- 11.8 to 65.8 +/- 8.2kPa/l per sec, p<0.05) and remained low. CONCLUSIONS: Porcine models of endotoxaemia based on LPS administration are a poor model of human septic shock, but can be improved by regulating PVR and supporting CO which may contribute to future studies of septic shock PMID- 12120970 TI - Exploration of the acute scrotum: a retrospective analysis of 100 consecutive cases. AB - BACKGROUND: Acute scrotal pain is a common urological emergency. Urgent exploration is the standard means of management, as no investigation can confidently exclude torsion of testis from the differential diagnoses. In our department, all patients up to the age of 40 years, presenting with acute scrotal pain undergo emergency scrotal exploration. AIM: This study assesses the outcome of such a policy in one unit. METHODS: The notes of 100 consecutive males who underwent exploration of scrotum have been reviewed. RESULTS: Testicular torsion was diagnosed in 33% of patients, with torsion of the appendix testis being the next most common entity. In the group with testicular torsion 12% required orchidectomy, and all had presented at least 24 hours after the onset of pain. The perioperative morbidity of scrotal exploration was very low. CONCLUSION: These data justify the policy of emergency exploration for all cases of acute scrotal pain up to the age of 40 years. PMID- 12120969 TI - Diagnostic and therapeutic ERCP: a large single centre's experience. AB - BACKGROUND: Most large published series on endoscopic retrograde cholangiopancreatography (ERCP) are multicentre-based and consequently reflect varying experience. AIMS: To assess morbidity and mortality rates of ERCP in a single tertiary referral centre. METHODS: A series of 1,758 consecutive ERCPs performed in 1,148 patients between 1991 and 1994 were reviewed to evaluate indications, findings, procedures, success, complication and mortality rates. RESULTS: There were 1,108 (63%) successful initial ERCPs, 11% failed cannulation attempts and 26% follow-up ERCPs. The desired duct was successfully cannulated in 96.5% of cases. Initial cannulation failure rate was 8.8%. Twenty-seven per cent had normal ERCPs, 30% had choledocholithiasis and 22% had strictures. Fifty-five per cent had therapeutic ERCPs. Major complications occurred in 3.5% with four ERCP-related deaths (0.35%). Therapeutic ERCP had a higher incidence of major complications compared to diagnostic ERCP: 4.6% vs 2.1%, (p=0.02); and mortality rate was 0.5% vs 0.2%, (p=0.4). Significant haemorrhage secondary to biliary sphincterotomy, pre-cut papillotomy and snare papillectomy accounted for most of the difference (1.6%). CONCLUSIONS: The majority of ERCPs were performed in elderly patients, over half of whom required therapeutic ERCP. Therapeutic ERCP carried significantly higher complication rate compared with diagnostic ERCP. Unsuccessful cannulation and follow-up ERCP accounted for 11% and 26% of ERCP workload, respectively. PMID- 12120971 TI - The increasing prevalence of childhood sickle-cell disease in Ireland. AB - BACKGROUND: Ireland has been relatively free of sickle-cell disease (SCD) and a care policy for the disease has not been established. AIM: To determine the prevalence of childhood SCD in Ireland and to predict requirements for a comprehensive care and assessment programme. METHODS: We retrospectively analysed the data of children with SCD presenting with sickle-cell crisis to one institution from January 1999 to April 2001. We also determined the nature and severity of the presenting haemoglobinopathy phenotype. RESULTS: Ninety-two patients with haemoglobinopathy have been registered with the Paediatric Haematology Service. The majority are from Nigeria with a smaller number from Angola and the Congo. Sixty have sickle-cell trait, 23 SCD, four haemoglobin SC disease and two haemoglobin E (HbE). There have been 32 sickle-cell crises. The majority were haemolytic or splenic sequestration events with a smaller number of aplastic and vaso-occlusive events and one osteomyelitis. CONCLUSION: The increasing number of children presenting with SCD as a result of the increasing refugee numbers requires a comprehensive care approach similar to that required for paediatric haemophilia to ensure optimum care. PMID- 12120972 TI - Views of Irish general practitioners on screening for cervical cancer. AB - BACKGROUND: A national cervical screening programme is being established in Ireland and there is little information on the level of resources required. AIMS: To obtain information on attitudes of Irish general practitioners (GPs) and on the resources needed by them in relation to participation in the programme. METHODS: An anonymous postal questionnaire was sent to a random sample of 600 GPs, approximately 25% of the total population of GPs in Ireland. RESULTS: A response rate of 87.5% was obtained. The majority (88%) would participate in a national programme. Those who would not were more likely to be in single-handed practice, aged over 44 years, have no ancillary staff, no computer and be in rural practice. GPs were in favour of a special fee for smear taking, a training programme and management guidelines on the test report. They did not want bonus payments for achieving targets or the report to be sent to the client as well as the doctor. CONCLUSION: GPs will support a national cervical screening programme but a number of organisational issues must be discussed with them to ensure a successful programme. PMID- 12120973 TI - Deaths in general practice: an Irish national profile. AB - BACKGROUND: There is little information on general practitioner (GP) involvement in terminal care. Aim This study explores general practice experience of the care of dying patients. METHODS: One hundred and forty-two GPs offered to participate in a study of consecutive deaths during three months, to a maximum of five cases per practice. Data were collected on patient characteristics, cause and place of death, terminal care and GP notification of deaths. RESULTS: One hundred and three GPs (73%) completed data collection. Participating GPs were younger and more likely to be in group practice. There were 297 deaths reported: 34% of practices had five deaths or more but 20% had no death. Seventy-five per cent of patients had one GP consultation in the final three months, 60% had at least one hospital admission and 38.8% of deaths occurred at home. Mean home visit, surgery consultation and phone consultation rates were 5.4, 1.8 and 3.6 respectively. In 88% of cases, the GP was informed of the death within one week. CONCLUSIONS: GPs are notified rapidly of deaths in all groups and causes. In the majority, the GP has had recent clinical contact and has often been heavily involved in care. Most deaths and care occur outside the cancer-related sphere. PMID- 12120975 TI - Self-castration. AB - BACKGROUND: Deliberate genital self-mutilation is rare. Successful self castration has been reported in a small number of individuals. METHOD: This study reports one such case of self-castration in a transsexual who was dissatisfied with waiting times for sex reassignment surgery (SRS). RESULT AND CONCLUSION: The apparent triggering factor in this case appears to be depression related to the lengthy waiting times for SRS. As neither psychotherapy nor hormonal manipulation is successful, consideration should be given to improving the resources for patients with genuine gender dysmorphism. PMID- 12120974 TI - Patients' views on out-of-hours care in general practice in Dublin. AB - BACKGROUND: Little is known regarding patients' views and levels of satisfaction with out-of-hours care in Irish general practice despite significant recent changes in service delivery. AIMS: This study aimed to record patients' experience of out-of-hours care on a specific occasion and elicit their satisfaction with out-of-hours care in general. METHODS: Patients requesting out of-hours care in three south inner city Dublin practices in June and July 2000 were identified and sent an anonymous postal questionnaire. RESULTS: Two hundred and forty patients were identified and 58% responded to the questionnaire. The approximate call rate was 195 calls per 1,000 patients per year. Sixty-one per cent of patients used the co-operative service, 28% received a house call and 3% received telephone advice only; 86% are currently satisfied with out-of-hours care. CONCLUSIONS: The majority of patients are satisfied with the current out-of hours service. Telephone consultation rates are significantly lower than other countries. These findings need to be considered before the widespread introduction of systems involving increased telephone consultations. PMID- 12120976 TI - Primary lymphoma of the bladder: a report of three cases. AB - BACKGROUND: Primary lymphoma of the bladder is rare and its management is an evolving field. AIMS: To highlight primary lymphoma of the bladder as a possible diagnosis in cases of bladder neoplasm and to illustrate the currently favoured management options. METHODS: Three cases of primary bladder lymphoma are reported and management is reviewed. RESULTS: Each of the three cases was managed differently with each management approach yielding a favourable outcome. CONCLUSION: Chemotherapy combined, if necessary, with surgery or radiation therapy, should be the standard of care, depending on the full histological diagnosis. PMID- 12120977 TI - Quinolone-associated tendonitis: a potential problem in COPD? AB - BACKGROUND: Quinolones have traditionally had limited application in the area of community-acquired respiratory tract infections due to poor cover against Streptococcus pneumoniae. This trend is changing with the broader spectrum of newer fluoroquinolones. A rare serious side effect of fluoroquinolones is tendinopathy. AIMS: This study describes two cases of levofloxacin-associated tendinopathy in patients with severe chronic obstructive pulmonary disease (COPD) and the implications and mechanisms involved are discussed. CONCLUSIONS: The finding of two cases of levofloxacin-induced tendinopathy in our patients suggests that the problem may be more frequent than previously considered. Patients with COPD treated with fluoroquinolones may have other risk factors for tendinopathy such as advanced age, corticosteroid use and renal impairment and merit vigilance for signs of tendonitis. PMID- 12120978 TI - Greek medicine from Asclepius to Hippocrates. PMID- 12120979 TI - The seventeenth century. PMID- 12120980 TI - Tuberculosis complicated by ARDS. PMID- 12120981 TI - Inferior dislocation of the patella: an unusual cause of a locked knee. PMID- 12120982 TI - Biliary sepsis in a patient on anti-TNFalpha therapy. PMID- 12120983 TI - The vacuolar-atpase of Paramecium multimicronucleatum: gene structure of the B subunit and the dynamics of the V-ATPase-rich osmoregulatory membranes. AB - Previous studies have shown that the vacuolar-ATPase (V-ATPase) of the contractile vacuole complexes (CVCs) in Paramecium multimicronucleatum is necessary for fluid segregation and osmoregulation. In the current study, immunofluorescence showed that the development of a new CVC begins with the formation of a new pore around which the collecting canals form. The decorated membranes are then deposited around the newly formed collecting canals. Quick freeze deep-etch techniques reveal that six 10-nm-wide V-ATPase V, sectors, tightly packed into a 20 x 30-nm rectangle, form two rows of these compacted sectors that helically wrap around the cytosolic side of decorated membrane tubules. During new CVC formation, packing of decorated tubules around mature CVCs was temporarily disrupted so that some of these decorated tubules became transformed into decorated vesicles. Freeze-fracturing of these decorated vesicles revealed a highly pitted E-face and a particulate P-face. The V-ATPase was purified for the first time in any ciliated protozoan and shown to contain, as in other cells, the V1 subunits A to E, and four 14-20 kDa polypeptides. The B subunit was cloned and found to be encoded by one gene containing four short introns. This subunit has 510 amino acid residues with a predicted molecular weight of 56.8 kDa, a value similar to B subunits of other organisms. Except for the N- and C-termini, it has a 75% sequence identity with other B subunits, suggesting that the B subunits in Paramecium, like other species, have been conserved and that the entire surface of this subunit may be important in interacting with other subunits. PMID- 12120984 TI - Fine structure of the myxosporean, Henneguya curimata n. sp., parasite of the Amazonian fish, Curimata inormata (Teleostei, Curimatidae). AB - Henneguya curimata n. sp. (Myxozoa, Myxobolidae) is described from the kidney of the teleost Curimata inormata collected in an estuarine region of the Amazon River, near Belem. Brazil. This myxosporean produces large cysts (0.6-1.2 mm in diam.) that represent plasmodia containing all life cycle stages, including spores. The spore body is ellipsoidal (approximately 16.6 microm in length and approximately 6.2 microm in width), and each valve presents a tapering tail (approximately 19.1 microm in length). These valves surround the binucleate sporoplasm cell and two ellipsoidal polar capsules located side-by-side at the same level, measuring 6.5 x 1.2 microm each and containing 10-11 coils of the polar filament. On the basis of its host specificity and on data collected by light and electron microscopy, the organism, H. curimata n. sp. is distinguished as a new species. The taxonomic affinities and morphological comparisons with other similar species of the same genus are discussed. PMID- 12120986 TI - Entamoeba histolytica Schaudinn, 1903 and Entamoeba dispar Brumpt, 1925: differences in their cell surfaces and in the bacteria-containing vacuoles. AB - Entamoeba histolytica Schaudinn, 1903 and Entamoeba dispar Brumpt. 1925 are two of eight species of Entamoeba that sometimes inhabit the human colon. The former is an invasive organism capable of causing life-threatening intestinal and extra intestinal disease: the latter appears not to be invasive. Because the two species, when viewed by light microscopy appear morphologically similar, they were long regarded as a single species. However, recent biochemical. immunological, and genetic studies provided convincing evidence that they belong to separate species. Our ultrastructural studies revealed distinct differences in at least two features of the trophozoites. 1) The cell surfaces of the trophozoites of each species differ with regard to structures exposed on the surface, and the distribution and arrangement of intra-membranous proteins. 2) The phagocytosis of bacteria differs in respect to the formation of the phagocytic vacuoles. Loose vacuoles containing several bacteria were seen in E. histolytica whereas tight vacuoles containing a single bacterium were observed in E. dispar. Furthermore, bacteria were found only within vacuoles in E. histolytica; in E. dispar, bacteria were found within vacuoles and some were found free in the cytoplasm. PMID- 12120985 TI - In vitro effect of nitazoxanide against Entamoeba histolytica, Giardia intestinalis and Trichomonas vaginalis trophozoites. AB - Nitazoxanide, a 5-nitrothiazolyl derivative, is effective in the treatment of a broad range of parasitic infections. In vitro, it is active against several protozoa, including Cryptosporidium parvum, Blastocystis hominis, and Giardia intestinalis. The objective of this study was to determine the in vitro effect of nitazoxanide on the growth and morphology of three anaerobic protozoa (Entamoeba histolytica, Giardia intestinalis, and Trichomonas vaginalis) and to compare these effects with those of metronidazole and albendazole. A subculture method was used to determine the concentrations required to inhibit growth by 50% or 90% (IC50 and IC90,). Nitazoxanide exhibited IC50, and IC90 values of 0.017 and 0.776 microg/ml respectively, against E. histolytica, 0.004 and 0.067 microg/ml against G. intestinalis, and 0.034 and 2.04 6 microg/ml against T. vaginalis. Based on the IC90 values, nitazoxanide was more toxic than metronidazole and albendazole against E. histolytica; albendazole and nitazoxanide were more toxic than metronidazole against G. intestinalis; and metronidazole was the most toxic drug against T. vaginalis. The effects of nitazoxanide on trophozoite ultrastructure of all three parasites included cell swelling and distorted cell shape, a redistribution of vacuoles, plasma membrane damage, and the formation of extensive empty areas in the cytoplasm of the protozoa. PMID- 12120987 TI - The fine structure of the gamont of Pterospora floridiensis (Apicomplexa: Eugregarinida). AB - Transmission electron microscopy of the gamont stage of Pterospora floridiensis has revealed a number of features. The gamont's surface varies from smooth to crenulate, with numerous pockets and folds. The pellicle is composed of an outer membrane, a middle lucent region, and an inner dense layer comprised of two tightly appressed membranes. Short ridges on the pellicle are 200-300+ nm long, 75-100 nm wide, and have a height of approximately 50 nm. The thickness of the pellicle is 100 nm when measured from the inner membrane to the top of a ridge. The ridges are formed by the plasma membrane and an underlying structure that is circular in cross-section. The surface folds and the pellicular ridges are distributed over the soma and the cell's unusual branching arms, though both are reduced near the junction between two gamonts in syzygy, and are absent at the central area of the junctional site. The cell has numerous active Golgi complexes associated with vesicles, as well as scattered dense mitochondria, lipid droplets, and paraglycogen granules. The nucleus has a large (13 microm) endosome, eccentrically located, and peripheral chromatin along the inner nuclear membrane. PMID- 12120988 TI - Morphological, small subunit rRNA, and physiological characterization of Trimyema minutum (Kahl, 1931), an anaerobic ciliate from submarine hydrothermal vents growing from 28 degrees C to 52 degrees C. AB - A thermophilic strain of Trimyema minutum was isolated from the hydrothermally heated sea floor at Vulcano Island (Italy) and cultivated monoxenically on Marinobacter sp. and Methanococcus thermolithotrophicus. It can be propagated strictly anaerobically and is sensitive to oxygen: if exposed to air at 48 degrees C all cells die within 60 min. It grows from 0.45-7.2% (w/v) salt and at pH 6.0-8.0. The isolate is the most extreme thermophilic ciliate which ever has been cultivated, exhibiting an optimal growth temperature of 48 degrees C (doubling time 6 h). Growth occurs between 28 degrees C and 52 degrees C. Trimyema minutum is redescribed using live observation and silver impregnation. Its morphology and the small subunit ribosomal RNA sequence is distinctly different from that of T. compressum, but morphology is highly similar to that of T. shoalsia Nerad et al. 1995, which is thus probably a junior synonym of T. minutum. To stabilize the bewildering species taxonomy in Trimyema, we suggest to recognize our population as a neotype of T. minutum. PMID- 12120989 TI - How oxymonads lost their groove: an ultrastructural comparison of Monocercomonoides and excavate taxa. AB - Despite being amongst the more familiar groups of heterotrophic flagellates, the evolutionary affinities of oxymonads remain poorly understood. A re interpretation of the cytoskeleton of the oxymonad Monocercomonoides hausmanni suggests that this organism has a similar ultrastructural organisation to members of the informal assemblage 'excavate taxa'. The preaxostyle, 'R1' root, and 'R2' root of M. hausmanni are proposed to be homologous to the right, left, and anterior roots respectively of excavate taxa. The 'paracrystalline' portion of the preaxostyle, previously treated as unique to oxymonads, is proposed to be homologous to the I fibre of excavate taxa. Other non-microtubular fibres are identified that have both positional and substructural similarity to the distinctive B and C fibres of excavate taxa. A homologue to the 'singlet root', otherwise distinctive for excavate taxa, is also proposed. The preaxostyle and C fibre homologue in Monocercomonoides are most similar to the homologous structures in Trimastix. suggesting a particularly close relationship. This supports and extends recent molecular phylogenetic findings that Trimastix and oxymonads form a clade. We conclude that oxymonads have an excavate ancestry, and that the 'excavate taxa' sensu stricto form a paraphyletic assemblage. PMID- 12120990 TI - Antigenic diversity of Encephalitozoon hellem demonstrated by subspecies-specific monoclonal antibodies. AB - Encephalitozoon hellem is a unicellular, obligate intracellular microsporidian species detected and isolated in HIV-infected patients presenting with keratoconjunctivitis, sinusitis, tracheobronchitis, nephritis, cystitis, and disseminated infection. A total of 24 monoclonal antibodies were produced against E. hellem and characterized. The monoclonal antibodies were of the immunoglobulin (Ig) G and Ig M subclasses, and, when incorporated into indirect immunofluorescence and immunoblotting assays, reacted against 13 isolates of E. hellem originating from three geographic regions. These monoclonal antibodies did not react with one strain each of either Encephalitozoon intestinalis or Encephalitozoon cuniculi, demonstrating their specificity. Two monoclonal antibodies reacted with all karyotype B-E. hellem isolates but did not react with karyotype A-isolates from North America and the Netherlands, thus demonstrating antigenic diversity among E. hellem isolates. These results add to the increasing evidence for diversity among E. hellem, which therefore may be reclassified into subspecies. PMID- 12120992 TI - Out of Africa. PMID- 12120991 TI - Alpha-proteobacterial relationship of apicomplexan lactate and malate dehydrogenases. AB - We have cloned and sequenced a lactate dehydrogenase (LDH) gene from Cryptosporidium parvum (CpLDH1). With this addition, and that of four recently deposited alpha-proteobacterial malate dehydrogenase (MDH) genes, the phylogenetic relationships among apicomplexan LDH and bacterial MDH were re examined. Consistent with previous studies, our maximum likelihood (ML) analysis using the quartet-puzzling method divided 105 LDH/MDH enzymes into five clades, and confirmed that mitochondrial MDH is a sister clade to those of y proteobacteria, rather than to alpha-proteobacteria. In addition, a Cryptosporidium parvum MDH (CpMDH1) was identified from the ongoing Cryptosporidium genome project that appears to belong to a distinct clade (III) comprised of 22 sequences from one archaebacterium, numerous eubacteria, and several apicomplexans. Using the ML puzzling test and bootstrapping analysis with protein distance and parsimony methods, the resulting trees not only robustly confirmed the alpha-proteobacterial relationship of apicomplexan LDH/MDH, but also supported a monophyletic relationship of CpLDH1 with CpMDHI. These data suggest that, unlike most other eukaryotes, the Apicomplexa may be one of the few lineages retaining an alpha-proteobacterial-type MDH that could have been acquired from an ancestral alpha-proteobacterium through primary endosymbiosis giving rise to the mitochondria, or through an unknown lateral gene transfer (LGT) event. PMID- 12120993 TI - HIV epidemics in Africa: what explains the variations in HIV prevalence? AB - There are large differences in the prevalence of HIV infection between different regions in sub-Saharan Africa, ranging from less than 10% in pregnant women in most of West Africa, to over 25% in pregnant women in large cities in Eastern and Southern Africa. These differences in HIV prevalence are in many instances due to differences in rate of spread of the virus. The multicenter study on factors determining the differential spread of HIV in four African cities tried to identify factors that could explain differences in spread of HIV between different regions in sub-Saharan Africa. The study was conducted in four cities, including two cities with a relatively low HIV prevalence (Cotonou in Benin and Yaounde in Cameroon) and two cities with a high HIV prevalence (Kisumu in Kenya and Ndola in Zambia). The difference in HIV prevalence between the four cities could not be explained by differences in sexual behavior. Any differences in sexual behavior were outweighed by differences in factors that influence HIV transmission, i.e. male circumcision and HSV-2 infection. These findings have important implications for the design of interventions. PMID- 12120994 TI - Neutralizing antibody responses to HIV-1 infection. AB - Neutralizing antibodies represent an important component of immune control in many viral infections. In HIV-1 infection, almost all individuals develop antibodies capable of neutralizing autologous viruses in vitro; however, the role of these antibodies in vivo still remains unclear. Their absence during the acute phase of infection, when the viral levels are brought under control, suggests they play a minor role in immune control and that cellular immune responses are more critical during this time. However, during chronic infection these antibodies may be important in preventing cell-to-cell spread and they still represent our best hope of providing sterilizing immunity (i.e., prevention of infection) by vaccination. Significant advances over the last few years in understanding the structure of the envelope glycoproteins have renewed interest in the role of neutralizing antibodies and the possibility that immunogens capable of stimulating a neutralizing antibody response can be developed. PMID- 12120995 TI - HIV receptors and cellular tropism. AB - Viruses use specific cell surface receptors to bind to and subsequently gain entry into their host cells. Some retroviruses such as HIV-1 and HIV-2 utilize one receptor for high-affinity binding (CD4), and a separate coreceptor to mediate fusion of the viral envelope with the cell membrane (CCR5 or CXCR4). The identification of these receptors explains the cellular tropism of HIV, and hence its pathogenesis leading to immune deficiency (T-helper cell depletion), the wasting syndrome (macrophage infection), and dementia (microglia infection). HIV can infect cells by membrane fusion at the cell surface and by receptor-mediated endocytosis. Knowledge of the HIV receptors has led to practical developments such as inhibitory drugs, reasons for genetic resistance to infection, and should inform the judicious choice of candidate vaccines. PMID- 12120996 TI - The development of HIV-1 subtype C vaccines for Southern Africa. AB - One in nine people in South Africa are estimated to be HIV-1 infected, with the majority of these infections being due to HIV-1 subtype C. Until recently, most HIV-1 candidate vaccines were not based on subtype C genes. In response to this epidemic, therefore, the South African AIDS Vaccine Initiative (SAAVI) was established to facilitate the development and testing of candidate HIV-1 subtype C vaccines. The first HIV-1 subtype C candidate vaccine is due to be, tested at the end of 2002, and is based on Venezuelan encephalitis virus replicons expressing Gag protein. The next candidate vaccines to be tested will be DNA and modified vaccinia Ankara vaccines expressing subtype C genes. PMID- 12120997 TI - Alphavirus replicon particles as candidate HIV vaccines. AB - Replicon particles based on Venezuelan equine encephalitis virus (VEE) contain a self-replicating RNA encoding the VEE replicase proteins and expressing a gene of interest in place of the viral structural protein genes. Structural proteins for packaging of replicon RNA into VEE replicon particles (VRPs) are expressed from separate helper RNAs. Aspects of the biology of VEE that are exploited in VRP vaccines include 1) expression of very high levels of immunogen, 2) expression of immunizing proteins in cells in the draining lymph node, and 3) the ability to induce mucosal immunity from a parental inoculation. Results of experiments with VRPs expressing green fluorescent protein or influenza virus hemagglutinin (HA) demonstrated that specific mutations in the VRP envelope glycoproteins affect both targeting in the draining lymph node and efficiency of the immune response in mice. VRPs expressing either the matrix-capsid portion of Gag, the full-length envelope gp160, or the secreted gp140 of cloned SIVsm H-4i were mixed in a cocktail and used to immunize macaques at 0, 1, and 4 months. Neutralizing antibodies against SIVsm H-4 were induced in 6 of 6 vaccinates and CTL in 4 of 6. An intrarectal challenge with the highly pathogenic SIVsm E660 was given at 5 months. A vaccine effect was seen in reduced peak virus loads, reduced virus loads both at set point and at 41 weeks postchallenge, and preserved or increased CD4 counts compared to controls. A candidate VRP HIV vaccine expressing Clade C Gag contains a sequence that is very close to the South African Clade C consensus and was selected from a recent seroconverter in the Durban cohort to represent currently circulating genotypes in South Africa. A GMP lot of this vaccine has been manufactured and tested for a phase I trial in the first months of 2002. PMID- 12120998 TI - Bacterial multidrug resistance mediated by a homologue of the human multidrug transporter P-glycoprotein. AB - Most ATP-binding cassette (ABC) multidrug transporters known to date are of eukaryotic origin, such as the P-glycoproteins (Pgps) and multidrug resistance associated proteins (MRPs). Only one well-characterized ABC multidrug transporter, LmrA, is of bacterial origin. On the basis of its structural and functional characteristics, this bacterial protein is classified as a member of the P-glycoprotein cluster of the ABC transporter superfamily. LmrA can even substitute for P-glycoprotein in human lung fibroblast cells, suggesting that this type of transporter is conserved from bacteria to man. The functional similarity between bacterial LmrA and human P-glycoprotein is further exemplified by their currently known spectrum of substrates, consisting mainly of hydrophobic cationic compounds. In addition, LmrA was found to confer resistance to eight classes of broad-spectrum antibiotics, and homologs of LmrA have been found in pathogenic bacteria, supporting the clinical and academic value of studying this bacterial protein. Current studies are focused on unraveling the mechanism by which ABC multidrug transporters, such as LmrA, couple the hydrolysis of ATP to the translocation of drugs across the membrane. Recent evidence indicates that LmrA mediates drug transport by an alternating two-site transport mechanism. PMID- 12120999 TI - Molecular epidemiology of TB: challenging dogmas and asking new questions. AB - Traditional epidemiological methods provide insight into the dynamics of diseases such as tuberculosis. These traditional techniques have limitations and rely on a number of assumptions. The application of molecular techniques to the study of epidemiology has allowed us to gain new insights into the biology of the organism Mycobacterium tuberculosis and the dynamics of the disease. We have been enabled to push the limits of understanding of the epidemiology of this disease, allowing us to challenge the old clinical dogmas, ask new questions, design new strategies, and measure the efficacy of such new interventions to combat this age old scourge. Among the dogmas challenged are that infection outside the home is commonplace, so-called relapse cases may in fact be largely reinfection, and active transmission may be more common than previously thought and reactivation disease relatively uncommon. These findings alone demand urgent attention and the design of optimal intervention strategies to reduce the burden of disease. PMID- 12121000 TI - Human genetic susceptibility to tuberculosis and other mycobacterial diseases. AB - The existence of a genetic component in mycobacterial disease susceptibility is no longer in doubt and the investigations now being conducted aim to determine which genes are involved, to what extent, and in which disease phenotype they are relevant. In certain rare instances of susceptibility to poorly pathogenic mycobacteria, the genetic component is clear. The approaches employed to elucidate common disease susceptibility include linkage studies, particularly genome-wide linkage analysis of both tuberculosis and leprosy, and association studies. A number of candidate genes have shown association with tuberculosis, and in many cases, on replication of the study, association has been confirmed in a disparate population, indicating the wider importance of the gene in the disease process. In other instances, associations appear to be particular to a population or a subtype of disease. PMID- 12121001 TI - Prediction of drug resistance in M. tuberculosis: molecular mechanisms, tools, and applications. AB - Management of Tuberculosis is complicated by the emergence of drug resistant strains of Mycobacterium tuberculosis and this poses a threat to the success of Tuberculosis control programmes. Drug susceptibility testing by culture is time consuming and technically difficult. It is known that resistance to drugs is due to a number of genomic mutations in specific genes of M. tuberculosis. These mutations in combination with molecular techniques can be used as markers for drug resistance, since drug susceptible isolates lack the corresponding gene mutations. This review focuses on molecular mechanisms, methods and applications as a possible new diagnostic tool for the early molecular detection of drug resistance in M. tuberculosis. PMID- 12121002 TI - Transcriptomics and proteomics: tools for the identification of novel drug targets and vaccine candidates for tuberculosis. AB - The availability of the complete genome sequence of Mycobacterium tuberculosis has revolutionised many areas of tuberculosis research and facilitated functional genomics studies. Most notably, transcriptomics and proteomics have become important tools in gaining increased understanding of the biology of M. tuberculosis and offer the promise of being able to deliver novel drug targets and vaccine candidates. PMID- 12121003 TI - Thiol metabolism of the trypanosomatids as potential drug targets. AB - Trypanosomatids produce significant amounts of four major low molecular mass thiols, trypanothione, glutathionylspermidine, glutathione, and ovothiol A. Of these, only glutathione is present in cells of the host. All four low molecular mass thiols are directly or indirectly maintained in a reduced state by trypanothione reductase. Available evidence, from gene disruption studies, indicate that this is an essential enzyme. Attempts to exploit trypanothione reductase as a chemotherapeutic target lead to the design of competitive and irreversible inhibitors of the enzyme. A promising route involves the design of redox cyclers interacting specifically with trypanothione reductase as subversive substrates. Progress in studies on the biosynthesis of ovothiol A is summarized. PMID- 12121004 TI - A chemical approach towards understanding the mechanism and reversal of drug resistance in Plasmodium falciparum: is it viable? AB - Genetic and biochemical approaches to studies of drug resistance mechanisms in Plasmodium falciparum have raised controversies and contradictions over the past several years. A different and novel chemical approach to this important problem is desirable at this point in time. Recently, the molecular basis of drug resistance in P. falciparum has been associated with mutations in the resistance genes, Chloroquine Resistance Transporter (PfCRT) and the P-glycoprotein homologue (Pgh1). Although not the determinant of chloroquine resistance in P. falciparum, mutations in Pgh1 have important implications for resistance to other antimalarial drugs. Because it is mutations in the aforementioned resistance genes rather than overexpression that has been associated with drug resistance in malaria, studies on mechanisms of drug resistance and its reversal by chemosensitisers should benefit from a chemical approach. Target-oriented organic synthesis of chemosensitisers against proteins implicated in drug resistance in malaria should shed light on mechanism of drug resistance and its reversal in this area. The effect of structurally diverse chemosensitisers should be examined on several putative resistance genes in P. falciparum to deal with antimalarial drug resistance in the broadest sense. Therefore, generating random mutations of these resistance proteins and subsequent screening in search of a specific phenotype followed by a search for mutations and/or chemosensitisers that affect a specific drug resistance pathway might be a viable strategy. This diversity oriented organic synthesis approach should offer the means to simultaneously identify resistance proteins that can serve as targets for therapeutic intervention (therapeutic target validation) and chemosensitisers that modulate the functions of these proteins (chemical target validation). PMID- 12121005 TI - Human papillomavirus (HPV) infection in Southern Africa: prevalence, immunity, and vaccine prospects. AB - Human papillomavirus (HPV) associated cancers are more prevalent in developing countries compared to developed countries. The major cancer caused by HPV is cervical cancer. The humoral immune response to HPV can be a marker of past infection but may also reflect persistent infection and cervical disease. IgA antibodies to HPV in oral fluid were also found to be markers of cervical disease. Cell mediated immunity is important in clearing HPV infection and for regression of the associated lesions: this means that women infected with HIV have a high prevalence of co-infection with HPV. Good cervical screening programmes can control HPV associated cervical neoplasia. However, in countries such as South Africa, where these programmes are inadequate, there is a need for an HPV vaccine. The development of HPV vaccines is reviewed. There is a call for an inexpensive vaccine that will be accessible to the women that do not have access to adequate screening programmes and are therefore at the greatest risk of cervical cancer. PMID- 12121006 TI - Kaposi's sarcoma biology. AB - Cancer remains a major burden for HIV-infected individuals. The majority of cancers associated with HIV infection are driven by oncogenic viruses like Epstein-Barr virus, Kaposi's sarcoma-associated herpesvirus and human papillomavirus. Kaposi's sarcoma is a tumour of endothelium and is the most common malignancy in HIV infected individuals. AIDS-related cancers are providing critical insight into cancer immunity and viral oncogenesis. PMID- 12121007 TI - Oesophageal cancer in Africa. AB - Oesophageal cancer, the eighth most frequent cancer in the world occurs as two main subtypes, squamous cell carcinoma (more prevalent in developing countries) and adenocarcinoma (more common in developed countries). Certain populations of central, eastern, and southern Africa display very high frequencies of oesophageal squamous cell carcinoma, presenting a serious health burden to the continent. Most patients are diagnosed at a late stage because of the asymptomatic development of the disease, with associated poor prognosis. A better understanding of the aetiological agents and molecular mechanisms involved in the development of oesophageal squamous cell carcinoma may offer opportunities to reduce exposure to environmental risk factors and also allow early diagnosis or predict response to therapy. Epidemiologic studies have identified smoking, alcohol consumption, diets poor in fresh fruit and vegetables, consumption of foods contaminated with Fusarium verticillioides, and HPV infection as risk factors associated with the development of this disease in Africa. Although we have an incomplete understanding of the molecular events involved in the development of oesophageal squamous cell carcinoma, advances have been made that suggest lines of future exploration. South African patients with oesophageal squamous cell carcinoma display a lower incidence of point mutations in the p53 gene than described elsewhere, suggesting that the profile of aetiological agents may be different than described for other high-risk areas for oesophageal cancer. Recent studies suggest that RARbeta and COX II is frequently downregulated and upregulated, respectively, in oesophageal squamous cell carcinomas. These results suggest potential therapeutic opportunities that can be exploited to combat the high incidence of this disease in Africa. PMID- 12121008 TI - Medicinal chemistry and chemical biology of new generation taxane antitumor agents. AB - P-glycoprotein (P-GP)-based multidrug resistance (MDR) and undesirable side effects are significant drawbacks to the clinical use of paclitaxel and docetaxel. Extensive SAR studies of taxanes in these laboratories led to the discovery of new generation taxanes that are highly active against not only drug sensitive but also drug-resistant human cancer cell lines as well as tumor xenografts in mice. One of these second generation taxanes, SB-T-110131 (IDN5109), exhibited excellent pharmacological profile in the preclinical studies and has been selected for clinical development (recoded as Bay 59-8862), which is currently in the phase II clinical trials. Bay 59-8862 is orally active with high bioavailability, showing excellent activity against a variety of drug-resistant tumors. "Advanced second generation taxanes" show essentially no difference in cytotoxicity against drug-resistant and drug-sensitive cell lines, virtually overcoming MDR. Photoaffinity labeling of P-GP using photoreactive radiolabeled paclitaxel analogs has disclosed the paclitaxel-binding domain of P-GP. Highly efficient taxane-based MDR reversal agents (TRAs) have also been developed, which can recover the cytotoxicity of paclitaxel to practically the original level against paclitaxel-resistant MDR expressing cancer cells. Highly promising results have emerged from the study of taxane-monoclonal antibody (MAb) immunoconjugates, which have been proved to specifically deliver extremely cytotoxic agents to tumor in an animal model. PMID- 12121009 TI - Risk stratification of older patients. AB - Cardiovascular disease is a growing global health threat due in part to demographic shifts, in particular the growing elderly population in which cardiovascular disease is highly likely to develop. Older individuals have numerous coexisting conditions that contribute to increased risk for morbidity and mortality. In the past, physicians primarily used diastolic blood pressure (BP) as the indicator for measuring relative risk. However, since the early 1970s observational studies have found that systolic BP rather than diastolic BP is a better predictor of cardiovascular events, particularly in the older population. The Sixth Report of the Joint National Committee on Prevention, Detection, Evaluation, and Treatment of High Blood Pressure (JNC-VI) recommended specific goals for the treatment of hypertensive patients. Specifically, JNC-VI reinforced the notion of more aggressive BP goals for patients at the greatest risk for an event. The approach to patients who are not at goal is different from earlier guidelines. Therapy is now aimed at building therapeutic regimens. Currently, there are five classes of agents that have been shown to reduce the morbidity and mortality associated with cardiovascular and renal disease: diuretics, beta blockers, angiotensin converting enzyme inhibitors, calcium antagonists, and angiotensin receptor blockers. If a patient is not at goal and there is no response to an agent from one of these classes or troublesome side effects develop, then and only then should that drug be stopped and a different one used. Otherwise, a multidrug regimen is constructed and additional agents added in logical fashion. A majority of patients with higher risk can attain control at the specified BP goal safely and effectively by using a combination of agents. PMID- 12121010 TI - Recent landmark clinical trials: how do they modify the therapeutic paradigm? AB - Despite intense investigation and clinical attention, many challenges remain in the management of the hypertensive patient. It is clear that hypertension remains inadequately controlled worldwide, with the control rate in the United States approximating 27%. Furthermore, several recent studies have underscored that it is frequently difficult to attain control at goal blood pressure (BP) with monotherapy and that adequate control of hypertension based on the newer more intensified BP goals necessitates multiple drug therapy. Indeed, in the recently published landmark trials of angiotensin I receptor antagonists, including the Irbesartan Diabetic Nephropathy Trial (IDNT) and Reduction of Endpoints in Non insulin-dependent Diabetes Mellitus with the Angiotensin II Antagonist Losartan (RENAAL), multiple antihypertensive drugs were required to attain goal. A pivotal class of drug required to comprise this regimen is the calcium antagonists. For example, in RENAAL, 78% of patients randomized to losartan required add-on therapy with a calcium antagonist. Calcium antagonists are an important and often necessary component of this multiple drug regimen. PMID- 12121011 TI - Calcium-channel blockade and cardiovascular prognosis: recent evidence from clinical outcome trials. AB - This article has three purposes: 1) to summarize recent findings of the Syst-Eur Trial; 2) to provide a short overview of the large trials in hypertension that have compared older with newer drug classes; and 3) to update the results of a meta-regression analysis that addressed the question of: to what extent blood pressure (BP) lowering can explain the findings of recent outcome trials in hypertensive patients or high-risk patients with normotension or hypertension. The Syst-Eur trial showed that in older patients with isolated systolic hypertension, drug treatment starting with a dihydropyridine calcium channel blocker reduced the risk of stroke and of all cardiovascular complications. Furthermore, this treatment regimen improved the prognosis of diabetic patients; reduced the incidence of proteinuria; and prevented dementia, in particular Alzheimer's disease. The pooled evidence from nine recently published actively controlled outcome trials involving 62,605 hypertensive patients proved that calcium channel blockers have the same long-term efficacy and safety as the older drug classes. Compared with diuretics and beta-blockers, calcium channel blockers may offer greater protection against stroke and less protection against myocardial infarction, resulting in similar overall cardiovascular benefit. A meta-regression analysis including 30 trials and 149,407 hypertensive or high risk patients showed that BP gradients largely accounted for most-if not all-of the differences in outcome. These findings emphasize the desirability of tight BP control. The hypothesis that angiotensin converting enzyme inhibitors or alpha blockers might influence outcome beyond their BP lowering-effects was not confirmed. PMID- 12121012 TI - Calcium antagonists in hypertension: from hemodynamics to outcomes. AB - Hypertension, by definition, is a hemodynamic disorder. A high cardiac output and a normal systemic vascular resistance characterize the young hypertensive patient. As hypertension progresses, resistance becomes progressively elevated and cardiac output returns to normal. The elderly patient with hypertension has very high systemic vascular resistance and low cardiac output. Antihypertensive drugs should not only lower arterial pressure but also bring other hemodynamic parameters as well as functional and structural changes of the cardiovascular system back to normal. With the notable exception of the classic beta-blockers, all antihypertensive drug classes, including the vasodilating beta-blockers, increase or maintain cardiac output and lower systemic vascular resistance. Calcium antagonists, although a very heterogeneous group, have been shown to have a similar effect on systemic hemodynamics. Initially, the short-acting agents (even verapamil) produce a reflex increase in heart rate and cardiac output with a decrease in systemic vascular resistance. This reflexive cardiac acceleration is not seen with the extended-release or longer-acting formulations, which usually maintain cardiac output and decrease systemic resistance. Lercanidipine is a novel calcium antagonist that has been shown to differ from other dihydropyridines in that the incidence of vasodilatory edema for any given decrease in blood pressure is less pronounced. Whereas all dihydropyridine calcium antagonists dilate the afferent arteriole in the kidney, preclinical studies have shown that lercanidipine also produces dilation of the efferent vessel. Similar balanced pre- and postcapillary vasodilation may be an explanation for the lower incidence of vasodilatory edema seen clinically with lercanidipine. These micro- and macrovascular features make lercanidipine an attractive new member in the arsenal of the powerful dihydropyridine calcium antagonists. PMID- 12121013 TI - Orbital complications of sinusitis in children. AB - BACKGROUND: Orbital complications of sinusitis are uncommon but can result in significant morbidity if not appropriately managed. OBJECTIVE: This study was conducted to evaluate the clinical presentation, diagnosis, management, and outcome of orbital complications of sinusitis in children treated at our institution over a 10-year period. METHODS: The study retrospectively reviewed cases of 139 children with evidence of orbital complications of sinusitis admitted to the Montreal Children's Hospital between January 1990 and March 2000. Factors assessed included the clinical presentation, radiologic findings, management, and outcome (length of admission, complications). Complications were classified as preseptal if they did not penetrate the periorbita. Postseptal complications were defined as those penetrating the periorbita and were further subdivided into cellulitis and abscess categories. RESULTS: Seventy-two percent of patients presented with preseptal cellulitis, 19% with orbital cellulitis, and 9% with subperiosteal abscess. Ophthalmoplegia and proptosis at presentation were found to be predictors of postseptal disease, although computed tomography (CT) was necessary to differentiate between cellulitis and abscess. Preseptal disease resolved with antibiotics in all cases. Postseptal disease was treated medically and in some cases surgically, although surgery did not affect outcome. CONCLUSION: Preseptal complications of sinusitis can be diagnosed clinically without a CT scan and should be treated with an appropriate course of intravenous antibiotics. Postseptal complications of sinusitis can be diagnosed by the presence of ophthalmoplegia or proptosis and mandate a CT scan to differentiate abscess from orbital cellulitis. Management of these patients should include intravenous antibiotics, reserving surgery for selected cases. PMID- 12121014 TI - Use of skin staplers in head and neck surgery: prospective clinical study. AB - OBJECTIVE: To compare the advantages and disadvantages of skin staplers versus conventional sutures in head and neck surgery. SUBJECTS: Fifty consecutive patients who underwent extensive surgery in the head and neck area were included in this prospective trial. METHODS: Patients were randomized into two equal groups: one group had their wounds closed with staplers and the other with monofilament sutures. The main outcome measures were speed of suturing, wound healing and cosmetic result, complications, and cost of either method. RESULTS: Cosmetic results were good in both groups. Neither group had any complications. Wound closure speed was 18.9 +/- 1.4 cm/min when using skin staplers and 1.9 +/- 0.4 cm/min for conventional suturing (p < .001). CONCLUSION: Skin staplers significantly reduce wound closure time and yield similar cosmetic results with no complications and with only a slightly higher cost of suturing material. PMID- 12121016 TI - Effects of a noncompressive hematoma on free flap viability. AB - Microvascular free tissue transfer has become an integral component of head and neck reconstructive surgery and historically has a greater than 95% success rate. Hematomas have long been known to cause skin flap necrosis by a variety of mechanisms including direct pressure necrosis, cellular damage secondary to the effects of free radicals, and inflammation. Compression from a hematoma is known to cause free flap compromise. To our knowledge, however, there have been no studies documenting the effect of a noncompressive hematoma surrounding the vascular pedicle on free flap viabiliry. This animal study was designed to assess the effects of a noncompressive hematoma on free flap survival. A right groin cutaneous free flap was created in 32 male Spraque Dawley rats. The rats were then divided into three groups: (1) 10 rats had no fluid added to the anastomotic compartment; (2) 11 rats had 0.5 cc of blood, taken from the femoral artery at the time it was cut, instilled in the anastomotic compartment via the 18-gauge angiocath; and (3) 11 rats had 0.5 cc of pentastarch instilled in the anastomotic compartment via the 18-gauge angiocath. There was 100% free flap survival in all three groups. All vessels examined at the time of sacrifice revealed intact anastomoses and no evidence of luminal clots. This study strongly suggests that in the cutaneous groin free flap model, a perianastomotic noncompressive hematoma has no effect on free flap survival. PMID- 12121015 TI - Is hyperventilation-induced nystagmus more common in retrocochlear vestibular disease than in end-organ vestibular disease? AB - Hyperventilation-induced nystagmus (HVIN) has previously been shown by the senior author to be common in patients with both acoustic neuromas and following resection. The recurrent study's aim was to examine if HVIN was specific for retrocochlear pathology. To test this, the incidence of HVIN in 24 patients with confirmed acoustic neuroma was compared with its incidence in 38 patients with end-organ vestibular disease (defined as a greater than 25% reduction in caloric testing). Hyperventilation was carried out for 90 seconds. The results showed that 58% of the acoustic neuroma group were positive for HVIN versus 18% of the end-organ group. This difference was very significant on chi-square testing (p < .002). Hyperventilation-induced nystagmus appears to be much more prevalent in retrocochlear pathology than in end-organ pathology. PMID- 12121017 TI - Outcome of neck dissection for node-positive melanoma. AB - OBJECTIVE: To determine the rates of regional recurrence for node-positive melanoma after neck dissection alone. DESIGN: Retrospective review from a single tertiary care institution. METHODS: Data were obtained for all patients receiving neck dissection from 1990 to 1997 at Sunnybrook and Women's Health Sciences Center. Analysis was performed using the Kaplan-Meier method. RESULTS: Thirty-one patients underwent neck dissection for node-positive melanoma in the study period. The rate of regional recurrence was 31% at 5 years. The mean time to recurrence was 78 months. CONCLUSION: Neck dissection alone may be appropriate for some patients, but those with more advanced neck disease are likely to benefit from adjuvant radiotherapy. PMID- 12121018 TI - Retrospective study of the progression of oral premalignant lesions to squamous cell carcinoma: a South Wales experience. AB - The premalignant potential of leukoplakia is controversial. This study is intended to determine whether chronic oral "white lesions" demonstrated premalignant potential by investigating the population presented to the University Hospital of Wales in Cardiff between January 1996 and June 2000. Patient demographics are presented. During this period, 510 biopsies were performed on suspicious white lesions and 311 cases of oral cancer were diagnosed. During the same period, 11 of the 311 patients developed malignancy despite initial benign diagnosis. Thirty-four of the 510 biopsies were initially diagnosed as oral lichen planus (OLP), whereas 3 of the previously mentioned 11 were diagnosed as OLP and later developed squamous cell carcinoma. This represents an incidence of detected premalignancy in 3.5% of oral cancer cases. On re-examination of the initial biopsy specimens, these 3 cases met criteria for OLP in some areas, whereas the others appeared hyperkeratotic. These specimens may represent a subtype of premalignant chronic white lesions rather than exemplify the premalignant potential of OLP. Further direction of research will be discussed. PMID- 12121019 TI - The sheep as an experimental animal for tracheal surgery. AB - Resection and end-to-end anastomosis of the trachea are the preferred treatments for various benign and malignant diseases involving the trachea. Various suture techniques and suture materials have been reported; however, some basic questions are still unsolved. Most basic work has been done in dogs, but they are expensive to acquire and keep. Therefore, we evaluated the sheep as an experimental animal for tracheal surgery. We report on our results with sheep regarding anaesthesia, surgical procedure, and tracheal cross-sectional area. Sheep proved to be excellent experimental animals for surgical research regarding tracheal resections and end-to-end anastomosis. PMID- 12121020 TI - Medialization of the vocal cord with fat injection. AB - A simple way of obtaining a smooth paste of fat for injection for medialization of the vocal cord has been described. PMID- 12121021 TI - Cavernous sinus thrombosis secondary to sinusitis. PMID- 12121022 TI - Vascular tumours of the nasal septum. PMID- 12121023 TI - Paranasal sinus teratocarcinosarcoma with intradural extension. AB - Teratocarcinosarcoma, although a rare neoplastic entity, should be considered as a differential diagnosis in any middle-aged adult presenting with a history of intermittent unilateral epistaxis and nasal obstruction. Tissue biopsy may fail to reveal a full spectrum of histologic heterogeneity in these tumours, and definitive diagnosis is usually made with tumour resection. Aggressive treatment including surgery followed by adjuvant radiation therapy is advocated and confers a better rate of survival than radiotherapy alone. Our current report is unique in two respects. First, disease recurrence is usually manifested very early on, leading some authors to conclude that a neoplastic-free interval of 3 years or longer probably indicates a good chance of being cured. Our patient, in contrast, experienced a disease-free interval of 4 years before evidence of recurrence emerged. Second, intracranial extension with brain parenchymal involvement has not been previously reported despite the tumour's proximity to the anterior cranial fossa and its locally aggressive behaviour with frequent bony invasion. Despite intracranial invasion, our patient experienced a long disease-free interval. As such, even advanced disease should be treated aggressively. PMID- 12121024 TI - Metastatic malignant melanoma of the pyriform sinus. PMID- 12121025 TI - Dermoid cyst of the tongue: magnetic resonance imaging. PMID- 12121026 TI - Hearing loss caused by lightning strike: case report and review of the literature. PMID- 12121027 TI - Metastatic bone marrow involvement by squamous cell carcinoma of the tonsil. PMID- 12121028 TI - Nasopharyngeal tuberculosis with massive cervical lymphadenopathy. PMID- 12121031 TI - An unusual case of dysphagia: ectopic salivary gland pleomorphic adenoma of the parapharyngeal space. PMID- 12121029 TI - Osteogenic sarcoma metastasizing to the larynx. PMID- 12121030 TI - Post-traumatic bilateral facial nerve palsy. PMID- 12121032 TI - Poliovirus-induced hearing loss. PMID- 12121033 TI - Microtubule associated protein (tau) gene variability in patients with frontotemporal dementia. AB - Frontotemporal dementia represents up to 10% of all dementias and is, next to Alzheimer's disease and Lewy body disease, the third most common cause of degenerative dementia. The term "frontotemporal dementia" covers a range of conditions, including Pick's disease, frontal lobe degeneration and dementia associated with motor neurone disease. Neuropathologically FTD is characterised by atrophy of the frontal and temporal lobes of the cerebral cortex, often with additional subcortical changes. Both familial and more frequently sporadic forms of FTD can be recognised. Recently, mutations in the microtubule-associated protein (tau) gene have been found in families with frontotemporal dementia and parkinsonism linked to chromosome 17 (FTDP-17). The identification of mutations in the tau gene indicates that the protein plays a central role in the process of neurodegeneration. Epidemiology of frontotemporal dementias in Poland remains still unknown. A prevalence of tau mutations among Polish patients has not been established yet. Here, we report results of a mutational analysis of the tau gene among Polish FTD patients. No pathogenic mutation was found in the analysed sample. The study confirmed that the frequency of tau mutations is very low and depends strongly on the clinical criteria used to select patients. Mutations in the tau gene account only for a small number of FTD cases with a clear autosomal dominant pattern of disease inheritance. Therefore there should exist additionalgenetic and non-genetic factors contributing to the pathogenesis of both familial (linked and non-linked to chromosome 17) and sporadic forms of FTD. PMID- 12121035 TI - Early infantile form of spinal muscular atrophy (Werdnig-Hoffmann disease) with prolonged survival. AB - According to several previously used classification systems, spinal muscular atrophy (SMA) type I has been characterised by an early onset (< 6 months), severe course (patients are never able to sit without support) and fatal prognosis (death before 2-4 years). We report 36 out of 349 SMA type I patients (10%) who had an onset before 6 months and never learnt to sit but survived at least beyond their fifth birthday, in spite of total immobility and frequent respiratory infections. These data support a classification avoiding age at death as a criterion. In the subgroup of type I with prolonged survival, there was no difference in life span between those individuals who had an onset of disease at birth or before and those with symptoms starting after 3 months. Life span may be related to the ability to withstand repeated respiratory insult, as several of the sibships with children affected by similar neuromuscular compromise showed discordant ages at death. "Chronic" SMA I and SMA II may represent overlapping phenotypes as our index patients with SMA I had a sib with SMA. PMID- 12121034 TI - N-methyl-d-aspartate receptor-mediated processing of beta-amyloid precursor protein in rat hippocampal slices: in vitro--superfusion study. AB - Abnormal proteolytic degradation of the beta amyloid precursor protein (beta-APP) may result in accumulation of potentially neurotoxic beta amyloid (betaA). The role of various receptors in the regulation of beta-APP processing has been suggested. This study aimed to determine how NMDA receptors and Ca2+ ions regulate proteolysis of beta-APP in rat hippocampus in vitro. Adult rat hippocampal slices were superfused with NMDA-containing media, and immunoreactivity of soluble beta-APP derivatives was detected in dialysates. Application of 100 microM and 250 microM NMDA for 20 min in Ca2+-containing medium induced dose-dependent release of aminoterminal beta-APP derivatives, and a fragment of Abeta sequence, whereas carboxy-terminal fragments of beta-APP were only slightly detected. This indicates activation of beta-APP processing, and release of its soluble cleavage products. This effect was inhibited by NMDA receptor antagonist 1 microM MK-801 and 100 microM CPP in Ca2+-free medium, thus indicating that NMDA receptors and calcium ions mediate proteolytic non amyloidogenic degradation of the beta-APP. PMID- 12121036 TI - Suicide muscle cell programme-apoptosis. Ultrastructural study. AB - To establish the ultrastructural criteria of suicide muscle cell programme apoptosis we investigated the muscle biopsies of acute fatal spinal muscular atrophy (SMA) infants with gene survival motor neurone (SMN) exon 7 deletion. The muscle cell apoptosis was characterised by precisely sequential morphology of both cell nucleus and sarcoplasm. The most characteristic feature was the aggregation of nuclear chromatin initially into peripherally located ring and later into uniformly dense, dark masses. Accompanying features included sarcoplasmic condensation cell shrinking and fragmentation into multiple bodies. The coexistence of muscle cells at different stages of apoptosis in the same tissue section was the most prominent and specific feature of suicide muscle cell programme. The appearance of an identical sequence of ultrastructural changes in muscle as observed in other tissues suggests that a unique mechanism is involved in genetically programmed cell death. PMID- 12121038 TI - Early occurring neuronal migration and maturation disorders. AB - This case report presents severe malformations of the central nervous system as a result of a pathological process occurring very early during development. Abnormal maturation and migration of neuroblasts were seen in all cerebral structures. Such changes are most often genetically determined, but in our case the analysis of their aetiology was not available. PMID- 12121037 TI - CT angiography in the diagnosis of spontaneous extracerebral vertebral artery dissection. AB - Spontaneous vertebral artery dissection is an underrecognised cause of cerebral ischaemia, most often seen in young adult patients. For this diagnosis cerebral angiography is the most sensitive and specific method. We describe a 30-year-old, male, with cerebellar stroke, in whom spontaneous vertebral artery dissection was diagnosed. Our diagnosis was confirmed by cerebral angiography and CT angiography. In our case anticoagulation (in acute phase) was introduced subcutaneously and later continued orally. The outcome was very good with complete remission. CT angiography seems to be a promising non-invasive method for the diagnosis of vertebral artery dissection. The prognosis in the patient with this diagnosis is relatively good (if managed properly), but the disease must always be treated as serious and life-threatening. PMID- 12121039 TI - The effect of thymus ubiquitin complex on neuropathological changes within the brain of pregnant rabbits with experimental model of antiphospholipid syndrome. AB - The present study was designed to assess the impact of thymus ubiquitin complex (TUC) on morphological changes within the central nervous system (CNS) in pregnant rabbits with experimental antiphospholipid syndrome (APS). We also compared the intensity of neuropathological appearance and serological markers of the APS treated with the TUC. Post-mortem neuropathological investigations were done in 47 female New Zealand rabbits. The material was divided into 4 groups: Group I-23 pregnant animals with APS; Group II-7 pregnant rabbits with APS, treated with TUC; Group III-7 pregnant rabbits without APS, treated with TUC; Group IV (the control one)-110 pregnant animals without APS, untreated with TUC. The APS was induced by subcutaneous injections of cardiolipin. There were two main abnormalities within the CNS of pregnant rabbits, which could be a result of the APS: the thrombo-necrotic foci of nervous tissue, and perivascular or meningeal inflammatory infiltrates. It turned out that TUC application reduced the morphological abnormalities, but it did not eliminate them entirely. The number of cases with thrombo-necrotic changes was reduced by approximately 18%, while perivascular infiltrates by 36% respectively. The TUC application did not influence the meningeal infiltration. The platelet count significantly increased (p < 0.001), the activated partial thromboplastin time shortened (p < 0.001), and the number of immunised animals, demonstrating positive immunofluorescence test, significantly decreased after the TUC administration. The inflammatory changes within the CNS are the integral component of the experimental model of APS in pregnant rabbits. The TUC may decrease the inflammatory component of the APS in the CNS. The inflammatory infiltrates seem to be more responsive to TUC application and parallel to APS serological improvement after treatment than necrotic changes. PMID- 12121040 TI - Bacterial colonization and invasion in pigs experimentally exposed to Streptococcus suis serotype 2 in aerosol. AB - A recently developed porcine model for aerogenous infection with Streptococcus suis serotype 2 was applied in a study of the phases of bacterial colonization and initial invasion. Eighteen pigs were exposed to aerosolized S. suis serotype 2 after pre-exposure to mild acetic acid in aerosol. The animals were killed consecutively within the first six days after challenge. After death, all animals were necropsied and examined by bacteriology, histopathology, and immunohistochemistry. Systemic infection was established in four out of 18 animals exposed to S. suis serotype 2. All systemically infected animals developed clinical signs and lesions typical of the infection. In four additional animals, subclinical infection was demonstrated by re-isolation of S. suis from the palatine tonsil. However, in all 18 challenged animals, immunohistochemistry demonstrated S. suis serotype 2 antigen in the palatine and/or nasopharyngeal tonsils. In all four systemically infected animals, S. suis serotype 2 antigen was also found in the mandibular lymph node. These observations point towards the tonsils as possible portals of entry for S. suis serotype 2 with subsequent lymphogenous spread. Thus, the present findings parallel the proposed pathogenesis for S. suis serotype 1 infection in pigs. PMID- 12121041 TI - Prevalence of and resistance to anti-microbial drugs in selected microbial species isolated from bulk milk samples. AB - The prevalence of strains of Staphylococcus aureus, coagulase-negative (CN) staphylococci, Listeria monocytogenes, Escherichia coli, Enterococcus faecalis, E. faecium and Bacillus cereus, was investigated in 111 bulk milk samples. Staphylococcus aureus was isolated from 38 samples, CN staphylococci from 63 samples, E. coli from 49 samples, E. faecalis or E. faecium from 107 samples, and L. monocytogenes from two samples. Bacillus cereus was not found in any of the samples and three samples were free of any of the selected species. Sensitivity to the anti-microbial drugs amikacin, ampicillin, ampicillin + sulbactam, cephalothin (CLT), cephotaxime, clindamycin, chloramphenicol (CMP), co trimoxazole, erythromycin (ERY), gentamicin, neomycin, norfloxacin, oxacillin, penicillin, streptomycin (STR), tetracycline (TTC) and vancomycin was tested using the standard dilution technique. Minimum inhibitory concentration (MIC) characteristics (MIC50, MIC90, MIC range) were determined for each microbial species. Resistance against one or more anti-microbial drugs was found in 93% of S. aureus, 40% of CN staphylococci, 73% of E. coli, 88% of E.faecalis, 55% of E.faecium, and one L. monocytogenes strain. Most of the strains, particularly enterococci, were resistant to STR, TTC, and ERY (MIC50 4 microg/ml). A high percentage of staphylococci were resistant to beta-lactam antibiotics. High resistance to CLT was found in 11 strains of E. coli (MIC 256 microg/ml) and strains resistant to CMP (MIC90 16 microg/ml) were detected. The highest numbers of resistance phenotypes were found in E. coil (16) and CN staphylococci (12). Eighteen identical resistance phenotypes were demonstrated in indicator bacteria (E. coli, E. faecalis, E. faecium) and pathogens (S. aureus, CN staphylococci) isolated from the same bulk milk sample. The obtained resistance data were matched against the herd owners' information on therapeutic use of the drugs. This confrontation could not explain the findings of strains resistant to ERY or CMP. Our findings are evidence of selection of resistant strains among not only pathogenic agents, but also among indicator bacteria which can become significant carriers of transmissible resistance genes. PMID- 12121042 TI - Immunohistochemical detection of Mycoplasma agalactiae in formalin-fixed, paraffin-embedded tissues from naturally and experimentally infected goats. AB - Samples from the mammary tissue of 14 lactating goats (12 naturally infected and two experimentally infected) were examined for the presence of Mycoplasma agalactiae. A monoclonal antibody (5G12) was applied to formalin-fixed, paraffin wax-embedded sections and labelled by the avidin-biotin peroxidase complex (ABC) method. Histological examination of tissue sections revealed strong immunoreactivity in all animals included in the study. Mycoplasma agalactiae antigen was mainly detected in the cellular debris at the periphery of purulent exudates present within lactiferous sinuses, and lactiferous and interlobular ducts. In addition, M. agalactiae organisms appeared in the cytoplasm of the epithelium of ducts, and in infiltrating macrophages and neutrophils within the ducts, alveoli, interstitial tissue and regional lymph node sinuses. It is concluded that this monoclonal antibody-based immunohistochemical technique is an efficient and specific method for the post-mortem detection of M. agalactiae in cases of clinical mastitis as well as being a useful tool for the study of the route of infection and cellular types involved during mastitis caused by this organism. PMID- 12121043 TI - Effect of infectious status and parity on somatic cell count and California mastitis test in pampinta dairy ewes. AB - The relationship between somatic cell counts (SCC) and California mastitis test (CMT) results according to the infectious status of mammary halves and parity of Pampinta dairy ewes was evaluated. Tests were associated to bacteriological analysis and classified into three groups: uninfected (negative culture), infected by minor pathogens and infected by major pathogens. Coagulase-negative Staphylococcus (32.4%), Micrococcus spp. (32.4%), Corynebacterium spp. (5.4%), and Bacillus spp. (1.4%) were the minor pathogens isolated, while Staphylococcus aureus (27%) and Escherichia coli (1.4%) were the major pathogens isolated. A good correlation was found between the CMT and SCC, which included inflammatory and epithelial cells (r = 0.64; P < 0.0001). SCC averages for the CMT scores shown in parentheses were 223 576 (0); 245,248 (1); 397,778 (2); 1,159,109 (3) and 2,460,833 (4) cells/ml. The correlation between SCC and the infectious status of udder halves was 0.58 (P < 0.0001). The relationship between SCC and CMT profiles and infectious status studied by a discriminant analysis showed, with an accuracy of 65%, three infectious status groups. SCC arithmetic means were 244,470 cells/ml for negative culture, 1,044,100 cells/ml for minor pathogens and 2,045,652 cells/ ml for major pathogens. With the exception of 1-year-old ewes, no significant differences were observed in SCC as affected by age or parity. PMID- 12121045 TI - Plasma haptoglobin concentration in swine after challenge with Streptococcus suis. AB - Eight 15-week-old pigs, reared under specific pathogen-free conditions, were inoculated with Streptococcus suis serotype 2. The animals were monitored before and after challenge by measuring rectal temperature, recording specific clinical symptoms and collecting blood samples for haptoglobin determination. Twenty-four hours after infection, the average haptoglobin plasma concentration of the animal group increased significantly and reached a maximum 4 days post-inoculation, followed by a constant mean level until the end of the trial on day 10. In spite of individual differences between the animals, an increase in haptoglobin concentration of at least 2.5 times above normal was observed in all infected pigs 1 day after challenge. Twenty-four hours after challenge, lameness was observed in five animals and an elevated body temperature was observed in seven of the eight experimental infected animals. These are the classical clinical symptoms of streptococcal infection. Haptoglobin was shown to increase in acute S. suis infection in pigs. PMID- 12121044 TI - Glomerulonephritis associated with simultaneous canine adenovirus-1 and Dirofilaria immitis infection in a dog. AB - This article describes a case of glomerulonephritis and immunocomplex (IgM, IgG and C3c) deposition in the mesangium and basement membranes of a 2-year-old dog with canine viral hepatitis and dirofilariasis. The deposits observed in the mesangium were in the vicinity of cells with viral replication. However, no clear relationship was found between viral replication and the deposition of immunocomplexes in the glomerular capillary basement membranes, which may be the reason why these deposits have only been tentatively related to the concomitant infestation by Dirofilaria immitis. PMID- 12121046 TI - Characterization of Mycoplasma hyosynoviae strains by amplified fragment length polymorphism analysis, pulsed-field gel electrophoresis and 16S ribosomal DNA sequencing. AB - Mycoplasma hyospnoviae strains from Denmark, Germany, Japan, Sweden, the Netherlands and the UK were examined for variations in the genomic DNA and within the 16S ribosomal RNA (rRNA) gene. Variations in the chromosomal DNA among 57 isolates recovered from the respiratory tract and joints of pigs, were investigated by analysis of amplified fragment length polymorphisms of the Bg/II and MfeI restriction sites and by pulsed-field gel electrophoresis of a BssHII digest of chromosomal DNA. Both methods allowed unambiguous differentiation of the analysed strains and showed similar discriminatory potential for the differentiation of M. hyosynoviae isolates. Concordant results obtained with the two whole-genome fingerprinting techniques evidence the considerable intraspecies genetic heterogeneity of M. hyosynoviae. Sixteen field strains of M. hyosynoviae and the type strain S16(T) were further examined for variation within the 16S rRNA gene. Ten field strains possessed the 16S rDNA sequences identical to the type strain, while the remaining six strains had sequences that differed by one to two nucleotides from that obtained from the type strain. PMID- 12121047 TI - Prevalence of antibodies to classical swine fever, Aujeszky's disease, porcine reproductive and respiratory syndrome, and bovine viral diarrhoea viruses in wild boars in Croatia. AB - During the hunting season in February 1999, a total of 44 blood samples were collected from wild boars shot in the area of Moslavacka gora. These blood samples were examined by enzyme immunoassay for the presence of antibodies to classical swine fever (CSFV), Aujeszky's disease (ADV), bovine viral diarrhoea (BVDV), and porcine reproductive and respiratory syndrome (PRRSV) viruses. Out of 44 serum samples examined, 17 (38.63%) were positive for CSFV, 24 (54.54%) were positive for ADV and two (4.54%) were positive for BVDV. All sera were negative for PRRSV. The results, recorded for the first time in Croatia, supported the hypothesis that wild boar act as a potential reservoir of CSFV, ADV and BVDV, and thus have a role in the epidemiology of these diseases. PMID- 12121048 TI - Molecular characterization of phenotypically CAMP-negative Streptococcus agalactiae isolated from bovine mastitis. AB - In the present study three phenotypically CAMP-negative Streptococcus agalactiae, isolated from three cows with mastitis, were characterized by molecular analysis. An identification of the S. agalactiae was performed by conventional methods and by PCR amplification of species specific parts of the 16S rRNA gene and the 16S 23S rDNA intergenic spacer region. In addition all three phenotypically CAMP negative isolates harboured a normal sized CAMP-factor encoding cfb gene indicating a reduced expression of CAMP-factor or a gene defect elsewhere along the pathway of expression. The clonal identity of the three isolates could be demonstrated by macrorestriction analysis of their chromosomal DNA. PMID- 12121049 TI - Identification of immunodominant antigens of adult Haemonchus contortus (Nematoda: Trichostrongylidae). PMID- 12121050 TI - Are antiphospholipid antibodies an independent risk factor for atherosclerosis? AB - The purpose of this study was to check whether antiphospholipid antibodies (aPL) could be an independent risk factor for atherosclerosis. Eighty-five subjects were studied: 45 with primitive antiphospholipid antibody syndrome and 40 controls affected by deep vein thrombosis secondary to known causes. The two groups were homogeneous for age, sex, and risk factors for atherosclerosis. All the subjects submitted to echo-color doppler of the carotid arteries, femoral arteries, and abdominal aorta. The cases were then subdivided into three subgroups on the basis of the positivity to the three subpopulations of aPL. Results demonstrate that there is no correlation between aPL and atherosclerosis. The different positivity to aPL does not modify this conclusion. PMID- 12121051 TI - Reference intervals of the dilute tissue thromboplastin inhibition and dilute Russell's viper venom tests revisited. AB - Reference intervals for two well-recognized tests for the lupus anticoagulant were determined using 98 healthy subjects. The purpose of the study was to compare the reference intervals for the dilute tissue thromboplastin inhibition test on this group of healthy subjects calculated by parametric and nonparametric statistical methods, and to compare these results with results obtained on subsets of 20 and 40 randomly selected individuals from the group of 98. The same procedures were followed for the dilute Russell's viper venom test. Results were recorded in seconds of clotting times and in ratios (subject/mean of that set or subset). Statistical analysis revealed Gaussian distribution of the results in the large group as well as in each subset for both tests. The results showed more variation between sets of the dilute tissue thromboplastin inhibition test than of the dilute Russell's viper venom test. Nonparametrically calculated reference intervals were wider than those determined by the parametric method, especially if confidence intervals are provided for both reference limits in a group of 94 controls or in a subset of 40 subjects. The nonparametric technique uses all data for the calculation of reference interval of such sample sizes no matter how widely spread the results are. There was no significant difference between the reference intervals of subsets and the whole group (p > 0.05) calculated by both statistical techniques. Very few outliers were observed among these subjects in both tests. PMID- 12121052 TI - Hemostatic abnormalities following bone marrow transplantation. AB - Hemostatic abnormalities in 26 patients following bone marrow transplantation (BMT) were examined. In the event-free survival group, the plasma levels of antithrombin (AT) and protein C (PC) were significantly decreased 1 and 2 weeks after BMT, and the plasma levels of thrombomodulin (TM) and tissue plasminogen activator-plasminogen activator inhibitor-1 complex (tPA-PAI-I complex) were significantly increased from 4 weeks to 13 weeks after BMT. Excepting AT, there was no significant difference in hemostatic parameters before BMT among the event free survival, 6-month survival, and death within 6 months groups. On day 0 following BMT, only plasma AT levels were significantly lower in the 6-month survival group than in the death within 6 months group. From 1 to 3 weeks after BMT, plasma levels of AT or PC were significantly lower in the death within 6 months group than in the 6-month survival group. From 1 to 5 weeks after BMT, the plasma levels of TM and tissue type plasminogen activator-plasminogen activator inhibitor-I complex (tPA-PAI-I complex) were significantly higher in the 6-month survival group than in the death within 6 months group. From 1 to 13 weeks after BMT, the plasma levels of D-dimer or soluble fibrin monomer (SFM) were significantly higher in the death within 6 months group than in the 6-month survival group. There was no remarkable difference in plasma levels of thrombin antithrombin comlex or plasmin-plasmin inhibitor complex following BMT between these groups of patients. These findings suggest that the decrease in the plasma AT or PC level reflects early occurrence of complications of prognostic significance and that the increase in vascular endothelial cell markers such as plasma levels of TM or tPA-PAI-I complex reflects occurrence of complications during the middle course of BMT. Plasma levels of D-dimer and SFM may be useful markers for predicting complications associated with poor prognosis after BMT. PMID- 12121053 TI - Facilitated thrombolysis: dethrombosis? AB - Intracoronary thrombus formation results from the combined effects of platelet aggregation and fibrin formation. Fibrinogen plays a significant role in each of these components. Dethrombosis is a therapeutic approach that allows dissolution of a recent thrombus while avoiding the potent fibrinolytic therapies. The less aggressive strategies suggested to achieve dethrombosis are antifibrin, antiplatelet, and defibrinogenation. Glycoprotein receptor antagonists have been beneficial in patients undergoing percutaneous transluminal coronary angioplasty; however, little is known about their potency of inducing reperfusion in acute myocardial infarction. Early use of the strategies suggested to induce dethrombosis (antifibrin, antiplatelet, and defibrinogenation) may facilitate fibrinolysis and primary angioplasty and further induce reperfusion. Nevertheless, the theoretical arguments of facilitated thrombolysis (dethrombosis) have not yet been fruitful as a major clinical benefit except in patients undergoing primary percutaneous coronary intervention. PMID- 12121054 TI - Is family history sufficient to identify women with risk of venous thromboembolism before commencing the contraceptive pill? AB - Oral contraceptives (OGs) increase risk of venous thromboembolic disease (VTE). The incidence of thomboembolic disease in healthy young women who are not taking OCs is 0.4-0.8/10,000, and in healthy young women using OCs, it is 3-4/10,000. To assess whether a family history of thromboembolism is a suitable tool to identify women who should not be given OCs, 50 women who suffered a VTE while taking OCs were studied. Only 16% of these women had family history which is why in our opinion, it is not a sufficient safeguard to recommending the use of OCs. PMID- 12121055 TI - Difference in mortality after hip fracture is associated with postdischarge prescription of antithrombotic prophylaxis: a case-control study. AB - This study was undertaken to determine whether a prolongation of pharmaceutical antithrombotic prophylaxis beyond hospitalization for hip fracture is associated with a reduced mortality rate. One hundred seventy-nine cases with hip fracture (patients older than 50 years of age) admitted to local general hospitals in 1999 who received postdischarge prescription of any antithrombotic agent (heparin, oral anticoagulants, antiplatelet drugs) and 179 age- and sex-matched patients with hip fracture who did not were included. Postdischarge mortality was assessed at 90 days. Compared with patients who did not receive postdischarge prescription of antithrombotic agents, those who did had an odds ratio of 0.22 (95% confidence interval 0.08-0.59) for all causes of mortality. This result did not change after excluding nonvascular mortality (odds ratio, 0.17; confidence interval, 0.03 0.73; p=0.011). Patients admitted to the hospital for hip fracture are at high risk of death after discharge if they are not given antithrombotic treatment. To substantiate these data, ad hoc prospective randomized trials are needed. PMID- 12121056 TI - Postprandial lipemia is associated with platelet and monocyte activation and increased monocyte cytokine expression in normolipemic men. AB - The activation of platelets and monocytes has been implicated in the development of cardiovascular diseases. We asked the question if postprandial lipemia following a fat- containing meal is associated with platelet and monocyte activation and increased platelet-monocyte interaction. Thirteen healthy, normal weight, normolipemic males, 20 to 49 years, consumed a 40% fat meal of whole foods. Blood samples were obtained at fasting and 3 1/2 and 6 hours after ingestion. Triglyceride levels increased to 48% over baseline at 3 1/2 hours postconsumption and returned to fasting levels by 6 hours. Multiparameter flow cytometry using monoclonal antibodies showed that the percentage of platelets expressing surface P-selectin and the activated conformation the GPIIb-IIa receptor was significantly higher at 3 1/2 hours compared to fasting. The percentage of platelet-monocyte aggregates increased by 36% at 3 1/2 hours and 43% at 6 hours postconsumption. The percentage of monocytes expressing intracellular tumor necrosis factor-alpha (TNF-alpha) increased seven and eightfold at 3 1/2 and 6 hours, respectively. The expression of interleukin-1beta (IL-1beta increased in a similar manner. These data suggest activation of platelets and monocytes after a moderate fat meal. Repetitive activation of platelets and monocytes could be an early event in the initiation and development of atherosclerosis. PMID- 12121057 TI - Oral contraceptives and venous thromboembolism: which are the safest preparations available? AB - Oral contraceptive therapy is associated with a fourfold increased risk of venous thromboembolism as compared with age-matched non-users. The composition of oral contraceptives has varied considerably during the past two to three decades. The estrogen content (ethinylestradiol) has decreased and is now less than 0.03 mg/pill. This was done on the assumption that estrogen was the main culprit for thrombotic complications. Subsequently it was found that the progestins contained in the pill could also play a thrombogenic role. This was particularly maintained to be so for the third-generations progestins, namely gestodene or desogestrel. These gonane progestins have been widely used since the early 1990s, because they appeared to have a lesser androgenic effect. A careful and impartial evaluation of the literature seems to indicate that third-generation progestins are associated with a slight increase in thrombotic risk. However, the significance of this difference remains to be proven. In fact, a relative risk of only two in retrospective studies may have limited effect and disappear in prospectives studies. The role of associated risk factors, both congenital and acquired, has been often overlooked in most of the papers dealing with the subject. This may be important. Preparations containing third-generation progestins are probably associated with a slight increase in thrombosis risk. It is the responsibility of the physician to select the preparation most suited for a given patient. As a general rule it may be safe to start with a preparation containing second generation progestins. However there is no need for "a pill scare" and it does not seem justified to have women already taking pills containing third-generation progestins to switch to other preparations. If a woman taking preparations containing third-generation progestins experience symptoms, it is probably safe to advise that patient not to take any oral contraceptive pill in the future, regardless of the type. The same is true for women who experience symptoms while taking second-generation progestins preparations. PMID- 12121058 TI - MIXCON-LA: a precise, sensitive and specific aPTT-based assay for detection of lupus anticoagulant. AB - Lupus anticoagulants (LA) are associated with an increased risk of thrombosis and laboratory detection is of major importance. Multiple tests are available for screening and confirmation, but they differ in sensitivity and specificity, frequently lacking the ability to discriminate between the presence of LA, heparin, and oral anticoagulants. Based on the test-principle of the Lupus Ratio test, an automated, sensitive APTT-based assay, using mixtures of a lupussensitive and a lupusinsensitive APTT-reagent with normal plasma for detection of lupus anticoagulants was developed. Ninety-nine healthy volunteers, ten patients treated with unfractionated heparin intravenously, 19 patients taking stable oral anticoagulation, five patients with hemophilia A, and 15 patients with antiphospholipid-antibody-syndrome (APS) were investigated. In all patients, two APTTs were performed, one with each reagent, on 1:1 mixtures of test plasma and normal plasma (MIXCON-LA assay). The ratio between the two clotting times was divided by the corresponding ratio for the normal plasma. This final lupus ratio (LR) was used for evaluation. The within-series imprecision and the between-series imprecision were excellent with coefficients of variation between 1.5% and 1.9%. The mean +/- 2 SD of the LR of the 99 healthy volunteers was used as reference range (LR: 0.95-1.07). All patients treated either with heparin or oral anticoagulants remained negative in the MLXCON-LA assay (specificity, 100%), while one of five patients with hemophilia A, in whom a factor VIII-inhibitor developed, showed a false-positive result. In 13 of 15 patients with APS, an increased ratio was observed (sensitivity, 87%). This assay system allows precise, specific, and sensitive detection of lupus anticoagulants. PMID- 12121059 TI - Effect of clopidogrel on platelet aggregation and plasma concentration of fibrinogen in subjects with cerebral or coronary atherosclerotic disease. AB - Acetylsalicylic acid inhibits thromboxane A2 production and reduces the risk of vascular occlusive events by 20% to 25%. Ticlopidine inhibits ADP-dependent platelet aggregation and reduces the same risk by 30% to 35%, but produces some adverse effects. Clopidogrel is a ticlopidin-related antiplatelet drug, with the same mechanism of action; it reduces the expression of the glycoprotein IIb/IIIa, the fibrinogen receptor on the platelet surface. Clopidogrel has the same clinical efficacy of ticlopidin and has a decreased incidence of adverse effects. The effect of one daily dose of 75 mg of clopidogrel on platelet function in 90 subjects was evaluated; 41 with coronary artery disease and 49 with cerebral vascular disease. Before treatment and after 6 and 12 weeks, bleeding time and fibrinogen plasma concentration were also evaluated. There was a reduction in 5 microM ADP-induced platelet aggregation of 38%+/-27% at 6 weeks and 44%+/-29% at 12 weeks in patients with coronary artery disease; 35%+/-41%, 29%+/-59% in the cerebral vascular disease group; and 36%+/-36% and 35%+/-49% in the total group. Reduction of 20 microg/mL collagen-induced platelet aggregation was not significant in any group. Plasma fibrinogen levels did not vary during treatment. Bleeding time was significantly prolonged in all studied groups. There were no hemorrhagic complications; only digestive discomfort in less than 3% of patients. Clopidogrel efficiently reduces ADP-induced platelet aggregation and prolongs bleeding time and is a safe and efficacious antiplatelet drug. PMID- 12121060 TI - Low-molecular-weight heparin for vertebral artery dissection. AB - A case of vertebral artery dissection and consequent basilar artery thrombosis in a 6-year-old patient is reported. The patient was treated with subcutaneous low molecular-weight heparin at therapeutic doses (enoxaparin, 100 IU/kg twice daily) for 3 weeks, then reduced to 2,000 IU/day for 3 more weeks. After this time a magnetic resonance angiogram was obtained that showed complete recanalization of the left basilar artery. Enoxaparin proved to be a safe and useful therapy for thrombosis accompanying vertebral artery dissection. PMID- 12121061 TI - Ticlopidine-induced marrow aplasia treated with cyclosporine. AB - A 50-year-old diabetic and hypertensive male patient is reported who had ticlopidine-induced marrow aplasia partially responsive to colony-stimulating factors and corticosteroids, but experienced complete recovery with cyclosporine. There is no consensus on the treatment of ticlopidine-induced marrow aplasia. Although many cases are reported to recover with colony-stimulating factors and corticosteroids, others are unresponsive or partially responsive. Our patient also did not completely respond to these medications, but was successfully treated with cyclosporine alone. Alone or in combination with corticosteroids, cyclosporine is an effective drug of choice for the resistant patients. PMID- 12121062 TI - Equivalence of capillary versus venous INR results and patient- versus professional-determined INR values using the CoaguChek S system. PMID- 12121063 TI - Tentative guidelines and practical suggestions to avoid venous thromboembolism during oral contraceptive therapy. AB - Oral contraceptive therapy (OCT) is widely used in the world. It is usually safe and effective but side effects are occasionally seen. Venous thromboembolism is one of the most feared side effects. To avoid this complication adequate guidelines are needed. These have to take into account family history, personal history, and suitable laboratory investigations. The presence of an idiopathic venous thrombosis in the family or in the personal history is of paramount importance. However it is often difficult to ascertain whether a venous thrombosis is idiopathic or not. Even when there is doubt, a coagulation study should be carried out. An adequate coagulation study in this case should include at least an evaluation of antithrombin, protein C, and protein S. A search for homozygosity of factor V Leiden appears advisable. These defects represent absolute contraindications to the use of OCT. Relative contraindications may be represented by other minor coagulation disorders such as heterozygous factor V Leiden, fibrinolysis defects, and a G-to-A 20210 prothrombin abnormality. Other noncoagulation-related conditions such as hypertension, liver damage, and obesity may represent absolute or relative contraindications to the use of OCT. PMID- 12121065 TI - Hemapheresis listening post. PMID- 12121064 TI - Positive selection of CD34+ cells by immunoadsorption: factors affecting the final yield and hematopoietic recovery in patients with hematological malignancies and solid tumors. AB - There is a progressive increase in the use of selected hematopoietic progenitor cells after myeloablative therapy in patients affected by malignancies. Our goal was to determine which blood parameters, in the starting cell population, influence the concentration of CD34+ progenitors and the removal of unwanted cells in the final product. Also, we evaluated the hematopoietic recovery and toxicity associated with peripheral blood stem cell infusion. We retrospectively reviewed 53 procedures of positive selection of CD34+ cells, performed with the Ceprate SC immunoadsorption system, in 47 paticnts affected by various hematologic malignancies and solid tumors. An increased percentage of CD34+ cells in the starting fraction was associated both with the final purity and enrichment of CD34+ cells and with a decreased percentage of CD3+ and CD19+ cells in the final product. A low platelet count before selection had a borderlinc influence on the recovery of CD34+ cells. Forty patients received a median of 5 x 10(6) CD34+ cells per kg; the absolute neutrophil count (ANC) reached 0.5 x 10(9)/l in a median of 10 days whereas a PLT count above 20 x 10(9)/l was observed in 14 days. The reinfusion of selected CD34+ cells, containing a very low amount of dymethylsulfoxide. was well tolerated and no adverse reactions were observed. Autologous transplantation with selected CD34+ cells is a safe and well-tolerated procedure in patients affected by hematologic malignancies and solid tumors. Positive selection of CD34+ cells seems to be related to the quality of the apheresis products, particularly to the initial CD34+ cell and PLT content. PMID- 12121066 TI - Transfusion in Japan. PMID- 12121067 TI - Board certification of medical doctors and medical technologists in transfusion medicine in Japan. AB - The Japan Society of Blood Transfusion (JSBT) organized a Board for certification of medical doctors in Transfusion Medicine (Board certified medical doctors by the JSBT) and a Board for certification of medical technologists for transfusion medicine (qualified medical technologists in Transfusion Medicine). For certified medical doctors the JSBT co-ordinated with the Japanese Association of Medical Science and with the Japan Medical Association, and for qualified medical technologists the JSBT co-organized with the Japanese Society of Laboratory Medicine, the Japanese Association of Medical Technologist and the College of Clinical Pathology of Japan. By May 2001, 259 of the certified medical doctors and 875 of the qualified medical technologists have been certified and are participating actively to increase the level of transfusion practice at individual facilities. PMID- 12121068 TI - Application of peripheral blood stem cells (PBSC) mobilized by recombinant human granulocyte colony stimulating factor for allogeneic PBSC transplantation and the comparison of allogeneic PBSC transplantation and bone marrow transplantation. AB - Recombinant human granulocyte colony stimulating factor (rhG-CSF)-mobilized peripheral blood stem cells (PBSC) are now widely used for allogeneic PBSC transplantation (alloPBSCT). Large numbers of hematopoietic progenitor cells mobilized by rhG-CSF would be considered equivalent or better than bone marrow (BM) cells and would be used as an alternative to BM for allogeneic hematopoietic stem cell transplantation. The complications associated with the administration of rhG-CSF and apheresis in PBSC collection in formal donors are well tolerated and usually acceptable in the short term but some hazardous adverse events such as splenic rupture and cardiac arrest are reported although the incidence is very low. Protective means and stopping rules for safe donation in the collection of PBSC are established. The characteristics of PBSC were clarified; the expression of some adhesion molecules such as CD49d on CD34 positive cells of PBSC have been shown to be low compared to BM stem cells. In alloPBSCT compared with allogeneic BM transplantation (alloBMT), the incidence and frequency of graft versus host disease (GVHD) is of concern because high number of T lymphocytes are infused in alloPBSCT. The incidence and severity of acute GVHD are not increased but chronic GVHD is higher in alloPBSCT compared with alloBMT. The outcome of alloPBSCT and BMT are almost equivalent and conclusive results regarding survival are not yet available. PMID- 12121069 TI - Blood program in Japan: current status. PMID- 12121070 TI - Changes in haematological parameters following captive bolt stunning in relation to the increased level of Syntaxin 1-B, a CNS marker in blood. PMID- 12121071 TI - Red cell storage lesion assessed by the levels of potassium, haemoglobin and Annexin V in supernatants. AB - The conventional and a new marker of global cellular lesion (Annexin V) are used to assess the processing/storage-induced changes in four types of RBC and filtered blood at 4 degrees and 22 degrees C, stored for a period of 35 days, in multi-satellite packs. It appears that mechanical trauma and presence of leucocytes and residual platelets have potential to increase levels of all markers of storage lesion, but to a variable extend. We have also provided new evidence that multi-satellite packs can be safely used for up to 35 days for small volume transfusion to sick premature infants, in a well-managed system by administering several transfusions from the same donation, hence reducing donor exposure. PMID- 12121072 TI - Changing perspectives of apheresis in India in the twentieth century. AB - Globally, the last century (20th) has seen many changes in terms of amazing advancements and applications of modern science, medicine, biotechnology and computers. One of the dramatic outcomes of such great inventions and discoveries was that the medical profession got the wonderful tool of immunomodulation and haemapheresis which has changed the outlook of the entire spectrum of immune complex disorders both in terms of the great success in treatment and the rehabilitation from so-called incurable diseases. Additionally this has resulted in a better quality of life and lower morbidity and mortality. Organ transplants have become a reality and immunomodulation plays a pivotal role in their success. India also has tasted this sweet recipe of modern medicine and has imbibed it in its medical culture. India is not only famous for the Taj Mahal and Hawa Mahal (Agra and Jaipur) but also for its brain (of brain drain/otherwise) and betz cells which have brought glories and laurels in the field of medicine, mathematics, engineering, literature, physics and computers, etc. to the native countries and India equally. Apheresis has now invaded the Indian scenario also and there is a growing interest, demand and application for it in the clinical field. But, being a unique country, the problems encountered are also unique. This paper deals with an overview of the changing perspectives of apheresis in India in the 20th century. PMID- 12121073 TI - Asymmetric sulfoxidation and amine binding by H64D/V68A and H64D/V68S Mb: mechanistic insight into the chiral discrimination step. AB - The H64D/V68A and H64D/V68S mutants of Myoglobin are found to oxidize thioanisole with high enantioselectivity and reactivity. These mutants are also capable of enantioselective binding of alpha-methylbenzylamine, which mimics an expected sulfoxidation intermediate. The kinetic study of the amine binding shows that the Fe-O bond cleavage in the intermediate may be the chiral discrimination step of the sulfoxidation. PMID- 12121074 TI - Controlled assembly of mesoscale structures using DNA as molecular bridges. AB - Mesoscale polyhedral structures from binary mixtures of microspheres of specific size ratios were prepared by using DNA as a molecular bridge. Carboxy-modified polystyrene beads were decorated with fluorescently labeled single-stranded DNA via carboxydiimide chemistry. Fluorescent resonance electron transfer in a confocal microscopy setting was utilized to corroborate DNA hybridization. Tetrahedrons were made by combining DNA-containing 0.818 and 0.211 mum beads, while octahedrons were obtained by bridging 0.818 and 0.364 mum beads. Confocal data in the reflection mode and SEM provide evidence for the formation of mesoscale building blocks. PMID- 12121075 TI - Point of attachment and sequence of immobilized peptide-acridine conjugates control affinity for nucleic acids. AB - Screening of combinatorial libraries by spatial arraying strategies requires library members to be solid-phase immobilized. However, for nucleic acid ligands that bind via intercalation, immobilization may inhibit binding if the tethering functionality is present at the edge of the heterocyle that approaches the duplex during the binding reaction. We report here a method for immobilizing peptide acridine conjugates (PACs) via either their C- or their N-terminus, corresponding to functionalization at either the 4- or the 9-position of acridine, respectively, and for assaying the nucleic acid binding properties of the resulting resins. We find that both the amino acid sequence of the PAC as well as its point of attachment to the solid support are important in determining affinity for duplex nucleic acids. These results have implications for the design of future on-bead and microarray-based selections and in understanding the nucleic acid binding of functionalized intercalators. PMID- 12121076 TI - Nonstatistical dynamics in deep potential wells: a quasiclassical trajectory study of methyl loss from the acetone radical cation. AB - The loss of methyl radical from the acetone radical cation, following its formation from the enol isomer, has been found to be nonergodic. The origins of this behavior are probed computationally by a quasiclassical trajectory simulation on an AM1-SRP potential energy surface. Experimental reaction outcomes are qualitatively reproduced, and evidence is presented that coherent excitation of the CCO angle bending during isomerization is a significant factor in the nonstatistical behavior. PMID- 12121078 TI - Synthesis and characterization of mesoporous indium tin oxide possessing an electronically conductive framework. AB - The new mesoporous transparent conducting oxide based on indium-tin-oxide, meso ITO, has been synthesized by a modified sol-gel method, using CTAB as the surfactant. Critical was the employment of triethanolamine to control the rate of hydrolysis and inhibit deposition of the bulk oxides. Removal of the surfactant by calcination yielded a relatively well-ordered worm-hole motif arrangement of pores visible in the TEM and stable to 400 degrees C. BET measurements revealed no hysteresis in the absorption-desorption isotherm, consistent with a narrow pore-size distribution (between 20 and 40 A depending on the In:Sn ratio); surface areas ranged between 270 and 310 m2/g. This colorless material is the first mesoporous oxide exhibiting substantial framework conductivity, with a conductivity at 25 degrees C of 1.2 x 10-3 S/cm. This distinguishes it from mesoporous mixed-valence transition-metal oxides that exhibit weak hopping semiconductor behavior and much lower conductivity. PMID- 12121077 TI - Generation of functionalized asymmetric benzynes with TMP-zincates. Effects of ligands on selectivity and reactivity of zincates. AB - We have developed new methods for preparing functionalized benzynes through deprotonative zincation as a key reaction using R2Zn(TMP)Li, and we also describes dramatic ligand effects on the benzyne formation. Deprotonative zincation of various meta-substituted bromobenzenes with Me2Zn(TMP)Li proved effective for the one-pot generation of various 3-functionalized benzynes, particularly those electrophilic substituents such as ester, amide, and cyano. On the other hand, zincation with tBu2Zn(TMP)Li, followed by electrophilic trapping (with I2) proved a powerful tool for the preparation of 1,2,3-trisubstituted aromatic compounds.8 The resultant 1,2,3-trisubstituted benzenes are available as precursors for generation of 3-substituted benzynes by halogen-zinc exchange reactions with Me3ZnLi. These methods offer far greater generality than previous methods for the synthesis of functionalized asymmetric benzynes, and should be of value in new syntheses of various natural products and functional materials. In addition, these results underline the utility of spectator ligands on the central metal of ate complexes as a tunable functionality in the development of new ate complex-promoted reactions. PMID- 12121079 TI - Gas-phase ozonolysis of alkenes: formation of OH from anti carbonyl oxides. AB - Gas-phase ozone-alkene reactions are known to produce the hydroxyl radical (OH) in high yields. Most mechanistic studies to date have focused on the role of syn carbonyl oxides; however, OH production from ethene ozonolysis indicates a second, poorly understood OH-forming channel, which may contribute to OH production in the ozonolysis of substituted alkenes as well. Using laser-induced fluorescence, we have measured OH and OD yields from the ozonolysis of two partially deuterated alkenes, cis- and trans-3-hexene-3,4-d2. OD is formed from both alkenes, indicating a pathway of hydroxyl-radical formation involving vinylic hydrogens, accounting for one-third of total OH formation from cis-3 hexene. The lack of a significant kinetic isotope effect suggests this pathway is the "hot acid" channel, arising from rearrangement of anti carbonyl oxides. Measured yields also allow for the estimation of syn:anti carbonyl oxide ratios, approximately 50:50 for trans-3-hexene and approximately 20:80 for cis-3-hexene, qualitatively consistent with our understanding of ozonide decomposition pathways. PMID- 12121080 TI - The reversible formation of a single-bonded (C(60)(-))(2) dimer in ionic charge transfer complex: Cp*(2)Cr.C(60)(C(6)H(4)Cl(2))(2). The molecular structure of (C(60)(-))(2). AB - A new ionic complex of C60 with decamethylchromocene, Cp*2Cr.C60(C6H4Cl2)2 (1), has been obtained. The fullerides are monomeric in 1 at room temperature, whereas they form a single-bonded (C60-)2 dimer at low temperatures, the structure of which has been studied by the X-ray diffraction on a single crystal at 100 K. The length of the intercage C-C bond is 1.597(7) A and the interfullerene distance is equal to 9.28 A. A phase transition attributed to the reversible C60*- dimerization is observed in the 220-200 K range. The transition is accompanied by changes in the unit cell parameters, the decrease of the magnetic moment from 4.20 muB (S = 3/2, 1/2) to 3.88 muB (S = 3/2) and the appearance of EPR signal from Cp*2Cr+, simultaneously. PMID- 12121081 TI - Protein prosthesis: a semisynthetic enzyme with a beta-peptide reverse turn. AB - beta-Amino acids are incorporated into an enzyme by using the method of expressed protein ligation. In the resulting semisynthetic enzyme, an R-nipecotic acid-S nipecotic acid module replaces Asn113 and Pro114 of ribonuclease A. The semisynthetic enzyme not only retains full catalytic activity but also gains conformational stability. Thus, structural elements can be replaced with foldameric equivalents to endow proteins with more desirable properties. PMID- 12121082 TI - Isocyanate formation in the catalytic reaction of CO + NO on Pd(111): an in situ infrared spectroscopic study at elevated pressures. AB - The catalytic CO + NO reaction to form CO2, N2, and N2O has been studied on a Pd(111) surface at pressures up to 240 mbar using in situ polarization modulation infrared reflection absorption spectroscopy (PM-IRAS). At 240 mbar, for a pressure ratio of PCO:PNO = 3:2 and under reaction conditions, besides adsorbed CO, the formation of isocyanate (-NCO) was observed. Once produced at 500-625 K, the isocyanate species was stable within the entire temperature range studied (300-625 K). On the other hand, its formation required a total CO + NO pressure of at least 0.6 mbar, illustrating the importance of in situ infrared experiments under high-pressure conditions. The significance of the isocyanate formation for the CO + NO reaction on Pd(111) is discussed. PMID- 12121084 TI - GaCl(3)-catalyzed ortho-ethynylation of phenols. AB - Phenols are ethynylated at the ortho position with silylated chloroethyne in the presence of a catalytic amount of GaCl3 and lithium phenoxide. The lithium salt is essential for the catalysis, and addition of 2,6-di(tert-butyl)-4 methylpyridine inhibits desilylation and hydration of the products. The reaction can be applied to various substituted phenols giving the ortho-ethynylated products in high yields, and the turnover numbers based on GaCl3 are between 8 and 10. The reaction mechanism involves addition of in situ formed phenoxygallium to the haloethyne followed by the elimination of GaCl3. PMID- 12121083 TI - Site-isolated luminescent europium complexes with polyester macroligands: metal centered heteroarm stars and nanoscale assemblies with labile block junctions. AB - The synthesis of a series of polymeric Eu(III) complexes with polyester ligands, along with supporting emission spectra, luminescence lifetimes, and, for a Eu block copolymer film, atomic force microscopy (AFM) data, is presented. Dibenzoylmethane was derivatized with a hydroxyl initiator site (dbmOH, 1) for tin octoate catalyzed ring opening polymerization of dl-lactide. The resulting poly(lactic acid) macroligand, dbmPLA (2), was combined with EuCl3 to generate Eu(dbmPLA)3 (3). Chelation of both dbmPLA and a polycaprolactone-functionalized bipyridine ligand (bpyPCL2) led to the Eu(III)-centered heteroarm star Eu(dbmPLA)3(bpyPCL2) (4). Unpolarized emission spectra and luminescence lifetimes were recorded for the Eu polymers in CH2Cl2 and for Eu(dbmPLA)3, as a film. Solution data for Eu(dbm)3 and Eu(dbm)3(bpy) were collected for comparison. For Eu tris(dbm) complexes, data were fit to a double exponential decay, indicating the presence of multiple species. Relative amounts of the longer lifetime component increase in the series Eu(dbm)3 solutions to Eu(dbmPLA)3 solutions to Eu(dbmPLA)3 films, perhaps suggesting benefits of the "polymer shell effect" and the diminishment of aquo adducts known to shorten lifetimes. As with the nonpolymeric analogue, data for Eu(dbmPLA)3(bpyPCL2) fit to a single-exponential decay. The sharpness of the feature at 579.7 nm, attributable to the 5D0 --> 7F0 transition in the emission spectrum of 4, lends further support for a homogeneous sample. AFM studies of "as cast" thin films of 4 reveal a lamellar structure with a 17.5 nm repeat. These microstructures, inferred to contain Eu luminophores at the glassy PLA-crystalline PCL domain interfaces, are modified by thermal treatment. PMID- 12121085 TI - Excited-state symmetry breaking in cofacial and linear dimers of a green perylenediimide chlorophyll analogue leading to ultrafast charge separation. AB - Photoexcitation of chromophoric dimers constrained to a symmetric pi-stacked geometry by their molecular structure usually produces excimers independent of solvent polarity, while dimers with edge-to-edge perpendicular pi systems undergo excited-state symmetry breaking in highly polar solvents leading to intradimer charge separation. We present direct evidence for symmetry breaking in the lowest excited singlet state of a symmetric cofacial dimer of 1,7-bis(pyrrolidin-1'-yl) perylene-3,4:9,10-bis(dicarboximide) (5PDI) in the low polarity solvent toluene to produce a radical ion pair quantitatively. This dimer, cof-5PDI2, was synthesized by attaching two 5PDI chromophores via imide groups to a xanthene spacer. For comparison, a linear symmetric dimer, lin-5PDI2, was prepared in which the 5PDI chromophores are linked end-to-end via a N-N single bond between their imides. The edge-to-edge pi systems of the 5PDI chromophores within lin 5PDI2 are perpendicular to one another. Ground-state absorption spectra of both 5PDI dimers show exciton coupling, which is consistent with the orientation of the 5PDI chromophores relative to one another. Ultrafast transient absorption spectroscopy following excitation of the dimers with 700 nm, 100 fs laser pulses shows that quantitative intradimer electron transfer occurs in cof-5PDI2 in toluene with tau = 0.17 ps followed by charge recombination to the ground state with tau = 222 ps. Similar measurements on lin-5PDI2 reveal that photoinduced electron transfer does not occur in toluene, but occurs in more polar solvents such as 2-methyltetrahydrofuran, wherein tau = 55 ps for charge separation and tau = 99 ps for charge recombination. Excited-state symmetry breaking in 5PDI dimers provides new routes to biomimetic charge separation and storage assemblies that can be more easily prepared and modified than those based on multiple tetrapyrrole macrocycles. PMID- 12121086 TI - Quantitative rate constants for radical reactions in the nanopores of cotton. AB - The understanding of radical reactions in nanostructured materials is important for developing new synthetic procedures and controlling degradation reactions. To develop this area, an easy method for measuring quantitative rate constants of some radical reactions in nanostructures is required. A simple method for measuring the rate constant of dye bleaching, kdye, by organic radicals in such materials is introduced, involving the measurement of microsecond bleaching kinetics by diffuse reflectance spectroscopy, following laser flash creation of the radicals. Using wet and dry cotton as model substrates, we obtained kdye of 2 hydroxy-2-propyl and 1-hydroxy-1-cyclohexyl radicals with reactive red 3 and reactive orange 4 and compared them to solution-phase values. Surprisingly, the reactions in cotton follow simple liquid-phase kinetics and are diffusion controlled. A cage effect in cotton is also found. PMID- 12121087 TI - First-principles study of aqueous hydroxide solutions. AB - Car-Parrinello molecular dynamics simulations have been carried out for aqueous NaOH and KOH solutions under ambient conditions over a wide range of concentrations. From these simulations, we have observed a continuous change of the water structure with added hydroxide, characterized by a significant shift of the second peak of the OO radial distribution functions to shorter distances. At the highest concentration investigated, the normal tetrahedral coordination of pure water is completely missing, a result that is consistent with a recent neutron diffraction experiment. The added hydroxide also gives rise to some unique spectroscopic features, including a "free" O-H stretch, a broadening of the normal water OH stretching band, and a large blue shift of both the librational band and the low-frequency translation. These results are in good agreement with the experimental data. Finally, it was demonstrated that the structural and dynamical behavior is inextricably linked to the formation of compact hydroxide-water complexes. PMID- 12121088 TI - Reversible intramolecular hydride transfer between tantalum and an aromatic ring: an eta(5)-cyclohexadienyl complex as a masked hydride. AB - The methyl triflate complex (2,6-Mes2C6H3N=)(2,6-Mes2C6H3NH)TaMe(OSO2CF3) reacts cleanly with H2 to give the eta5-cyclohexadienyl complex 3. Complex 3 therefore results from the insertion of an arene ring into an M-H bond, which has been proposed as the first step in catalytic arene hydrogenations. This insertion is reversible, such that 3 behaves as a "masked hydride" in its reaction chemistry. PMID- 12121089 TI - The "diindene" ArInInAr (Ar = C(6)H(3)-2,6-Dipp(2), Dipp = C(6)H(3)-2,6 Pr(i)(2)). Dimeric versus monomeric in(I) aryls: para-substituent effects in terphenyl ligands. AB - The synthesis and structure of a "diindene" with significant metal-metal bonding are described. It has an In-In distance of 2.9786(5) A, an In-In-C angle of 121.23(6) degrees , and an In-In bond order that is probably less than unity. PMID- 12121090 TI - Structure of the photocolored 2-(2',4'-dinitrobenzyl)pyridine crystal: two-photon induced solid-state proton transfer with minor structural perturbation. AB - The X-ray structure of the blue phototautomer of 2-(2',4'-dinitrobenzyl)pyridine (DNBP) produced by two-photon excitation in a single crystal is the first direct evidence of a product in the DNBP photochromic family. A nitro-assisted proton transfer mechanism is attributed to the photocoloration of solid DNBP. PMID- 12121091 TI - Synthesis and structures of [[HC(CMeNAr)(2)]Ge(S)X] (Ar = 2,6-iPr(2)C(6)H(3), X = F, Cl, Me): structurally characterized examples with a formal double bond between group 14 and 16 elements bearing a halide. AB - Treatment of [{HC(CMeNAr)2}GeX] (Ar = 2,6-iPr2C6H3, X = Cl (1), F (2)), with elemental sulfur at room temperature smoothly afforded the [{HC(CMeNAr)2}Ge(S)X] (X = Cl (3), F (4)). Compound 4 can also be obtained from 3 with the fluorination reagent Me3SnF. Reaction of 3 with MeLi led to the formation of [{HC(CMeNAr)2}Ge(S)Me] (5). Single-crystal X-ray structural analyses indicate compounds 3-5 are monomeric. The germanium centers adopt four coordinated sites and reside in distorted tetrahedral environment. Compounds 3 and 4 are structurally characterized examples with a formal double bond between group 14 and 16 elements bearing a halide. PMID- 12121092 TI - Novel histidine-heme covalent linkage in a hemoglobin. AB - When treated with dithionite at neutral pH, the recombinant hemoglobin from Synechocystis sp. PCC 6803 reconstituted with ferric heme undergoes a rapid chemical reaction resulting in the attachment of the heme group to the polypeptide chain. The nature of the cross-linked species was studied by NMR and mass spectral methods. 1H NMR data indicated that the 2-vinyl group was the reacting moiety of the heme. Mass spectrometry of pepsin digests located the site of attachment within a 12-mer at the C-terminal end of the protein. Homonuclear and 1H-15N NMR data identified the modified residue as His117, which underwent addition to the vinyl Calpha through the imidazole Nepsilon. Dithionite treatment of the globin reconstituted with Zn protoporphyrin IX sample did not lead to 2 vinyl group modification, suggesting that the chemical reduction of the heme iron facilitated the attachment. PMID- 12121093 TI - Syn stereochemistry of cyclic ether formation in 1,8-cineole biosynthesis catalyzed by recombinant synthase from Salvia officinalis. AB - Regiospecifically labeled geranyl diphosphates ((2E,6E)-[1,1,8,8,8-(2)H(5)]- and (2E,6Z)-[1,1,9,9,9-(2)H(5)]-GDP) and D(2)O incorporation, in concert with NMR spectrometry, were employed to demonstrate a unique intramolecular syn-facial protonation-cyclization mechanism of action of 1,8-cineole synthase. PMID- 12121094 TI - An oligomeric ser-pro dipeptide mimetic assuming the polyproline II helix conformation. AB - A hexapeptide assembled from dipeptide building blocks assumes the secondary structure of the polyproline II (PPII) helix. Side chain-to-backbone hydrogen bonds stabilize peptide torsions next to the fused ring systems. The solution structure of the polyhydroxylated peptide was studied by spectroscopic methods. PMID- 12121095 TI - Decoupling optical and potentiometric band gaps in pi-conjugated materials. AB - Syntheses, optical spectroscopy, potentiometric studies, and electronic structural calculations are reported for two classes of conjugated (porphinato)metal oligomers that feature a meso-to-meso ethyne-bridged linkage topology. One set of these systems, bis[(5,5'-10,20-bis[3,5-bis(3,3-dimethyl-1 butyloxy)phenyl]porphinato)zinc(II)]ethyne (DD), 5,15-bis[[5'-10',20'-bis[3,5 di(3,3-dimethyl-1-butyloxy)phenyl]porphinato)zinc(II)]ethynyl]-10,20-bis[3,5-di(9 methoxy-1,4,7-trioxanonyl)phenyl]porphinato)zinc(II) (DDD), and 5,15-bis[[15' ' (5'-10',20'-bis[3,5-bis(3,3-dimethyl-1-butyloxy)phenyl]porphinato)zinc(II)]-[(5' '-10' ',20' '-bis[3,5-di(9-methoxy-1,4,7 trioxanonyl)phenyl]porphinato)zinc(II)]ethyne]ethynyl]-10,20-bis[3,5-di(9-methoxy 1,4,7-trioxanonyl)phenyl]porphinato)zinc(II) (DDDDD), constitute highly soluble analogues of previously studied examples of this structural motif having simple 10,20-diaryl substituents, while a corresponding set of conjugated oligomers, [(5 10,20-bis[3,5-bis(3,3-dimethyl-1-butyloxy)phenyl]porphinato)zinc(II)]-[(5'-15' ethynyl-10',20'-bis[10,20-bis(heptafluoropropyl)porphinato)zinc(II)]ethyne (DA), 5,15-bis[[5'-10',20'-bis[3,5-di(3,3-dimethyl-1 butyloxy)phenyl]porphinato)zinc(II)]ethynyl]-10,20 bis(heptafluoropropyl)porphinato]zinc(II) (DAD), and 5,15-bis[[15' '-(5'-10',20' bis[3,5-bis(3,3-dimethyl-1-butyloxy)phenyl]porphinato)zinc(II)]-[(5' '-(10' ',20' '-bis(heptafluoropropyl)porphinato)zinc(II)]ethyne]ethynyl]-10,20-bis[3,5-di(9 methoxy-1,4,7-trioxanonyl)phenyl]porphinato)zinc(II) (DADAD), features alternating electron-rich and electron-poor (porphinato)zinc(II) units. Electrooptic and computational data for these species demonstrate that it is possible to engineer conjugated oligomeric structures that possess highly delocalized singlet (S1) excited states yet manifest apparent one-electron oxidation and reduction potentials (E1/20/+ and E1/2-/0 values) that are essentially invariant with respect to those elucidated for their constituent monomeric precursors. PMID- 12121096 TI - Structure of a tethered cationic 3-aminopropyl chain incorporated into an oligodeoxynucleotide: evidence for 3'-orientation in the major groove accompanied by DNA bending. AB - The structure of the dodecamer d(CGCGAATXCGCG)(2), in which X = Z3dU, 5-(3 aminopropyl)-2'-deoxyuridine, was determined. At neutral pH, Z3dU introduced a positive charge into the major groove. NMR spectroscopy revealed that the Z3dU omega-aminopropyl moiety oriented in the 3'-direction from the site of modification. Watson-Crick base pairing remained intact throughout the dodecamer. The presence of the charged amino group in the major groove resulted in a 0.24 ppm upfield shift of one (31)P NMR resonance in the 3'-direction at the phosphodiester linkage between nucleotides C(9) and G(10). Molecular dynamics calculations restrained by distances obtained from (1)H NOE data and torsion angles obtained from (1)H NMR (3)J coupling data, and in which the omega-amino group was constrained to be proximate to G(10)O(6), predicted from the (31)P NMR data and molecular modeling (Dande, P.; Liang, G.; Chen, F.-X.; Roberts, C.; Nelson, M. G.; Hashimoto, H.; Switzer, C.; Gold, B. Biochemistry 1997, 36, 6024 6032), were consistent with experimental NOEs. These refined structures exhibited bending. The distance from the amino group to the 5'-phosphate oxygen of Z3dU was >5 A, which indicated that in this dodecamer the Z3dU amino group did not participate in a salt bridge to its 5'-phosphate. PMID- 12121097 TI - Equilibrium constants for dehydration of water adducts of aromatic carbon-carbon double bonds. AB - Equilibrium constants (K(de)) are reported for the dehydration of hydrates of benzene, naphthalene, phenanthrene, and anthracene. Free energies of formation of the hydrates (DeltaG(o) (f)(aq)) are derived by combining free energies of formation of the parent (dihydroaromatic) hydrocarbon with estimates of the increment in free energy (DeltaG(OH)) accompanying replacement of a hydrogen atom of the hydrocarbon by a hydroxyl group. Combining these in turn with free energies of formation of H(2)O and of the aromatic hydrocarbon products furnishes the desired equilibrium constants. The method depends on the availability of thermodynamic data (i) for the hydrocarbons from which the hydrates are derived by hydroxyl substitution and (ii) for a sufficient range of alcohols to assess the structural dependence of DeltaG(OH). The data comprise chiefly heats of formation and standard entropies in the gas phase and free energies of transfer from the gas phase to aqueous solution (the latter being derived from vapor pressures and solubilities). They also include experimental measurements of equilibrium constants for dehydration of alcohols, especially cyclic, allylic, and benzylic alcohols. In general DeltaG(OH) depends on whether the alcohol is (a) primary, secondary, or tertiary; (b) allylic or benzylic; and (c) open chain or cyclic. Differences in geminal interactions of the hydroxyl group of the alcohol with alpha-alkyl and vinyl or phenyl groups account for variations in DeltaG(OH) of 5 kcal mol(-1). Weaker variations which arise from beta-vinyl/OH or beta-phenyl/OH interactions present in the aromatic hydrates but not in experimentally studied analogues are estimated as 1.0 kcal mol(-1). Equilibrium constants for dehydration may be expressed as their negative logs (pK(de)). Reactions yielding the following aliphatic, aromatic, and antiaromatic unsaturated products then have pK(de) values: +4.8, ethene; +15.0, ethyne; +22.1, cyclopropene; +28.4 cyclobutadiene; -22.2, benzene; -14.6, naphthalene; -9.2, phenanthrene; -7.4, anthracene. Large positive values are associated with formation of strained or antiaromatic double bonds and large negative values with aromatic double bonds. Trends in pK(de) parallel those of heats of hydrogenation. The results illustrate the usefulness of a substituent treatment for extending the range of currently available free energies of formation. In addition to hydroxyl substituent effects, DeltaG(OH), values of DeltaG(pi) for substitution of a pi-bond in a hydrocarbon are reported. PMID- 12121098 TI - Protonated benzofuran, anthracene, naphthalene, benzene, ethene, and ethyne: measurements and estimates of pK(a) and pK(R). AB - Aqueous solvolyses of acyl derivatives of hydrates (water adducts) of anthracene and benzofuran yield carbocations which undergo competitive deprotonation to form the aromatic molecules and nucleophilic reaction with water to give the aromatic hydrates. Trapping experiments with azide ions yield rate constants k(p) for the deprotonation and k(H2O) for the nucleophilic reaction based on the "azide clock". Combining these with rate constants for (a) the H(+)-catalyzed reaction of the hydrate to form the carbocation and (b) hydrogen isotope exchange of the aromatic molecule (from the literature) yields pK(R) = -6.0 and -9.4 and pK(a) = 13.5 and -16.3 for the protonated anthracene and protonated benzofuran, respectively. These pK values may be compared with pK(R) = -6.7 for naphthalene hydrate (1-hydroxy-1,2-dihydronaphthalene), extrapolated to water from measurements by Pirinccioglu and Thibblin for acetonitrile-water mixtures, and pK(a) = -20.4 for the 2-protonated naphthalene from combining k(p) with an exchange rate constant. The differences between pK(R) and pK(a) correspond to pK(H2O), the equilibrium constant for hydration of the aromatic molecule (pK(H2O) = pK(R) - pK(a)). For naphthalene and anthracene values of pK(H2O) = +13.7 and +7.5 compare with independent estimates of +14.2 and +7.4. For benzene, pK(a) = 24.3 is derived from an exchange rate constant and an assigned value for the reverse rate constant close to the limit for solvent relaxation. Combining this pK(a) with calculated values of pK(H2O) gives pK(R) = -2.4 and -2.1 for protonated benzenes forming 1,2- and 1,4-hydrates, respectively. Coincidentally, the rate constant for protonation of benzene is similar to those for protonation of ethylene and acetylene (Lucchini, V.; Modena, G. J. Am. Chem Soc. 1990, 112, 6291). Values of pK(a) for the ethyl and vinyl cations (-24.8) may thus be derived in the same way as that for the benzenonium ion. Combining these with appropriate values of pK(H2O) then yields pK(R) = -39.8 and -29.6 for the vinyl and ethyl cations, respectively. PMID- 12121100 TI - Interaction of anions with perfluoro aromatic compounds. AB - The complexes formed by a variety of anions with perfluoro derivatives of benzene, naphthalene, pyridine, thiophene, and furan have been calculated using DFT (B3LYP/6-31++G**) and MP2 (MP2/6-31++G** and MP2/6-311++G**) ab initio methods. The minimum structures show the anion interacting with the pi-cloud of the aromatic compounds. The interaction energies obtained range between -8 and 19 kcal mol(-1). The results obtained at the MP2/6-31++G** and MP2/6-311++G** levels are similar. However, the B3LYP/6-31++G** results provide longer interaction distances and smaller interaction energies than do the MP2 results. The interaction energies have been partitioned using an electrostatic, polarization, and van der Waals scheme. The AIM analysis of the electron density shows a variety of topologies depending on the aromatic system considered. PMID- 12121099 TI - Enantioselective total syntheses of manzamine a and related alkaloids. AB - As a prelude to undertaking the total syntheses of the complex manzamine alkaloids, a series of model studies were conducted to establish the scope and limitations of intramolecular [4 + 2] cycloadditions of N-acylated vinylogous ureas with the trienic substrates 17a,b, 28a,b, and 34. These experiments clearly demonstrated that the geometry of the internal double bond and the presence of an electron-withdrawing group on the diene moiety were essential for the facile and stereoselective formation of the desired cycloadducts. The enantioselective syntheses of the manzamine alkaloids ircinol A (75), ircinal A (5), and manzamine A (1) were then completed by employing a convergent strategy that featured a novel domino Stille/Diels-Alder reaction to construct the tricyclic ABC ring core embodied in these alkaloids. Thus, the readily accessible chiral dihydropyrrole 58 was first converted in a single chemical operation into the key tricyclic intermediate 60. Two ring-closing metathesis reactions were then used to form the 13- and 8-membered rings leading to Z-72 and 74, the latter of which was quickly elaborated into ircinal A (5) via ircinol A (75). The synthetic 5 thus obtained was converted into manzamine A (1) following literature precedent. This concise synthesis of ircinal A required a total of 24 operations from commercially available starting materials with the longest linear sequence being 21 steps. PMID- 12121101 TI - Combinations of microphase separation and terminal multiple hydrogen bonding in novel macromolecules. AB - The synthesis and characterization of terminal multiple hydrogen-bonded (MHB) polymers, such as poly(styrene) (PS), poly(isoprene) (PI), and microphase separated PS-b-PI block copolymers, possessing controlled molecular weights and narrow molecular distributions are described. Hydroxyl-terminated polymeric precursors were prepared using living anionic polymerization and subsequent quantitative termination with ethylene oxide. MHB polymers were synthesized in a controlled fashion via end-group modification of these well-defined macromolecular alcohols with excess isophorone diisocyanate and subsequent derivatization of the isocyanate-terminated polymeric intermediate with methyl isocytosine. The glass transition temperatures of the terminal MHB polymers were reproducibly higher than both nonfunctionalized and hydroxyl-terminated polymers at nearly equivalent number average molecular weights. Thin-layer chromatography analysis indicated that the interaction of terminal MHB polymers with silica was stronger as compared to both nonfunctionalized and hydroxyl-terminated polymers. Rheological characterization indicated that the melt viscosity at constant shear rate for various MHB polymers was more than 100 times higher than those for nonfunctionalized and hydroxyl-terminated polymers. Interestingly, the melt viscosity of MHB polymers was higher than those of nonfunctionalized polymers with twice the number average molecular weight. In addition, DSC and rheological characterization also suggested that terminal MHB polymers formed aggregates and not simple dimers in the melt state, and the aggregates were observed to completely dissociate at 80 degrees C. PMID- 12121102 TI - Molecular packing and morphology of oligo(m-phenylene ethynylene) foldamers. AB - Understanding and controlling solid-state morphologies and molecular conformations is the key to optimizing the properties of materials. As an example for the influence of small chemical changes on solid-state structures, we studied oligo(m-phenylene ethynylene) foldamers, where the introduction of an endo-methyl group induces a transition from an extended all-transoid to a helical all-cisoid conformation. The resulting structural changes were analyzed by X-ray diffraction (XRD), polarized optical microscopy (POM), and low-dose high-resolution electron microscopy (LD-HREM) over several length scales from the molecular to the mesoscopic level. The strong tendency of the endo-methyl oligomer 1 to form stable compact helices in solution resulted in round droplets with an ordered hexagonal columnar (Col(ho)) liquid crystalline structure, where shrinkage during the crystallization resulted in the formation of a banded texture. On the other hand, the endo-hydrogen oligomer 2 exhibited a very different morphology; its extended linear shape was maintained during crystallization and resulted in an extended lamellar structure, which was determined by a compromise between crystalline packing and minimization of the surface area. Another pronounced difference between both molecular structures was the ability of the extended lamellar "crystals" to bend, whereas the helices form either straight or disordered domains. In addition, both materials exhibit strong surface effects, which extend considerably inside the droplet and induce uniform bending of the supramolecular structures. PMID- 12121103 TI - Elucidation of the chiral recognition mechanism of cinchona alkaloid carbamate type receptors for 3,5-dinitrobenzoyl amino acids. AB - A cinchona alkaloid having extraordinary chiral discriminatory powers (alpha = 32.6 for dinitrobenzoyl leucine) is developed as a chiral stationary phase (CSP) for chromatography. An explanation of how chiral discrimination takes place is presented. Using a soluble analogue of the CSP, we found that NMR spectrometry indicates that 1:1 complexes exist for both optical isomers interacting with the CSP, that the free base form of the CSP exists in an open/closed ratio of 35/65 but that the protonated, bound-state form is exclusively in the anti-open conformation, and that significant intermolecular NOEs exist for the more stable diastereomeric complex but not for the less stable complex. Stochastic molecular dynamics simulations were carried out in solvents of low and high dielectric. The chromatographic retention orders and free energy differences of analyte binding to CSP were reproduced computationally as were the observed intra- and intermolecular NOEs. Data from the simulation were used to evaluate the intermolecular forces responsible for analyte binding as well as to discern fragments of the CSP doing most of the work of holding the complexes together. The enantiodifferentiating forces and the parts of the CSP most responsible for chiral discrimination are described. Moments of distributions of key dihedral angles and distances between centroids were used to assess the relative rigidity of the competing diastereomeric complexes. Simultaneous multiple-contact ion pairing, hydrogen bonding, and pi-stacking are possible for the longer retained enantiomer only. An X-ray crystallographic study of the more stable complex confirms the conclusions derived from chromatography, NMR spectroscopy, and molecular modeling. PMID- 12121104 TI - Dyeing uric acid crystals with methylene blue. AB - The growth of anhydrous uric acid (UA) and uric acid dihydrate (UAD) crystals from supersaturated aqueous solutions containing methylene blue, a cationic organic dye, has been investigated. Low concentrations of dye molecules were found to be included in both types of crystal matrixes during the growth process. Incorporation of dye into UA crystals occurs with high specificity, affecting primarily [001] and [201] growth sectors, while UAD crystals grown from solutions of similar dye concentration show inclusion but little specificity. The orientation of the UA-trapped species was determined from polarization data obtained from visible light microspectrometry. To achieve charge neutrality, a second anionic species must also be included with the methylene blue into UA and UAD crystal matrices. Under high pH conditions, crystallization of 1:1 stoichiometric mixtures of methylene blue and urate yields methylene blue hexahydrate (MBU.6(H2O). The crystal structure of MBU.6(H2O) reveals continuous pi-pi stacks of planes of dye cations and urate anions mediated by water molecules. This structure provides an optimal geometry for methylene blue-urate pairs and additional support for the incorporation of these dimers in uric acid single-crystal matrices. The strikingly different inclusion patterns in UA and UAD demonstrate that subtle changes in the crystal surfaces and/or growth dynamics can greatly affect recognition events. PMID- 12121105 TI - Design and synthesis of a novel class of sugar-peptide hybrids: C-linked glyco beta-amino acids through a stereoselective "acetate" Mannich reaction as the key strategic element. AB - A new type of sugar-amino acid hybrid, which is comprised of a sugar unit (gluco , galacto-, or mannopyranose) linked through a C-glycosidic linkage to the beta position of an alpha-unsubstituted beta-amino acid unit, is presented. It is hypothesized that these new compounds, or the oligomeric peptides derived therefrom, might possess the structural features of beta-amino acid oligomers and the chemical and enzymatic resistance of C-glycosides to hydrolysis. The synthetic strategy is based on a new Mannich-type reaction between a chiral acetate enolate equivalent and alpha-amido sulfones derived from the corresponding sugar-C-glycoside aldehydes. While the sugar-C-glycoside aldehyde partner is prepared from well-established transformations on known sugar precursors, the lithium enolate derived from (1R)-endo-2-acetylisoborneol 3 is employed as the key element. This Mannich approach proceeds with essentially perfect diasteromeric control leading to the new beta-amino carbonyl adducts in good yields. Further, cleavage of the camphor auxiliary is smoothly performed by oxidative treatment with ammonium cerium nitrate (CAN). Complementarily, direct peptide-type coupling of the beta-amino carbonyl Mannich adducts with an alpha- or beta-amino acid residue and subsequent CAN-promoted detachment of the auxiliary yields dipeptide fragments bearing a sugar-containing aliphatic side chain and is a process that can be iterated. A preliminary conformational study based on the combination of experimental NMR data and molecular mechanics and molecular dynamics (MD) of one particular adduct is also provided. PMID- 12121106 TI - Fluoride-selective binding in a new deep cavity calix[4]pyrrole: experiment and theory. AB - A new "super-extended cavity" tetraacetylcalix[4]pyrrole derivative was synthesized and characterized, and X-ray crystal structures of complexes bound to fluoride and acetonitrile were obtained. The binding behavior of this receptor was investigated by NMR titration, and the complex was found to exclusively bind fluoride ions in DMSO-d(6). This unusual binding behavior was investigated by Monte Carlo free energy perturbation simulations and Poisson calculations, and the ion specificity was seen to result from the favorable electrostatic interactions that the fluoride gains by sitting lower in the phenolic cavity of the receptor. The effect of water present in the DMSO on the calculated free energies of binding was also investigated. Owing to the use of a saturated ion solution, the effect of contaminating water is small in this case; however, it has the potential to be very significant at lower ion concentrations. Finally, the adaptive umbrella WHAM protocol was investigated and optimized for use in binding free energy calculations, and its efficiency was compared to that of the free energy perturbation calculations; adaptive umbrella WHAM was found to be approximately two times more efficient. In addition, structural evidence demonstrates that the protocol explores a wider conformational range than free energy perturbation and should therefore be the method of choice. This paper represents the first complete application of this methodology to "alchemical" changes. PMID- 12121107 TI - A facile approach to architecturally defined nanoparticles via intramolecular chain collapse. AB - A novel approach is presented for the controlled intramolecular collapse of linear polymer chains to give well-defined single-molecule nanoparticles whose structure is directly related to the original linear polymer. By employing a combination of living free radical polymerization and benzocyclobutene (BCB) chemistry, nanoparticles can be routinely prepared in multigram quantities with the size being accurately controlled by either the initial degree of polymerization of the linear chain or the level of incorporation of the BCB coupling groups. The latter also allows the cross-link density of the final nanoparticles to be manipulated. In analogy with dendritic macromolecules, a significant reduction of up to 75% in the hydrodynamic volume is observed on going from the starting random coil linear chains to the corresponding nanoparticles. The facile nature of the living free radical process also permits wide variation in monomer selection and functional group incorporation and allows novel macromolecular architectures to be prepared. Furthermore, the use of block copolymers functionalized with benzocyclobutene groups in only one of the blocks gives, after intramolecular collapse, a hybrid architecture in which a single linear polymer chain is attached to the globular nanoparticle. PMID- 12121108 TI - Organometallic dendrimers based on (tetraphenylcyclobutadiene)cyclopentadienylcobalt modules. AB - The synthesis and characterization of novel organometallic polyphenylene dendrimers containing 24 or 44 phenyl rings and one cyclobutadiene(cyclopentadienyl)cobalt unit is reported. The dendrimers are made by the convergent CpCo(CO)(2)-mediated dimerization of di- or tetraethynyltolanes followed by a divergent core extension utilizing tetraphenylcyclopentadienone. The obtained dendrimers are air and water stable, soluble materials that show interesting differences in their hydrodynamic properties as evidenced by gel permeation chromatography. Scanning pulse voltammetry in solution shows that the dendrimers are oxidized at potentials ranging from 0.8 to 0.83 V. The more sterically encumbered the dendrimer, the higher its oxidation potential, that is, the more difficult oxidation is. PMID- 12121109 TI - Genetic construction of truncated and chimeric metalloproteins derived from the alpha subunit of acetyl-CoA synthase from Clostridium thermoaceticum. AB - In this study, a genetics-based method is used to truncate acetyl-coenzyme A synthase from Clostridium thermoaceticum (ACS), an alpha(2)beta(2) tetrameric 310 kDa bifunctional enzyme. ACS catalyzes the reversible reduction of CO(2) to CO and the synthesis of acetyl-CoA from CO (or CO(2) in the presence of low potential reductants), CoA, and a methyl group bound to a corrinoid-iron sulfur protein (CoFeSP). ACS contains seven metal-sulfur clusters of four different types called A, B, C, and D. The B, C, and D clusters are located in the 72 kDa beta subunit, while the A-cluster, a Ni-X-Fe(4)S(4) cluster that serves as the active site for acetyl-CoA synthase activity, is located in the 82 kDa alpha subunit. The extent to which the essential properties of the cluster, including catalytic, redox, spectroscopic, and substrate-binding properties, were retained as ACS was progressively truncated was determined. Acetyl-CoA synthase catalytic activity remained when the entire beta subunit was removed, as long as CO, rather than CO(2) and a low-potential reductant, was used as a substrate. Truncating an approximately 30 kDa region from the N-terminus of the alpha subunit yielded a 49 kDa protein that lacked catalytic activity but exhibited A-cluster-like spectroscopic, redox, and CO-binding properties. Further truncation afforded a 23 kDa protein that lacked recognizable A-cluster properties except for UV-vis spectra typical of [Fe(4)S(4)](2+) clusters. Two chimeric proteins were constructed by fusing the gene encoding a ferredoxin from Chromatium vinosum to genes encoding the 49 and 82 kDa fragments of the alpha subunit. The chimeric proteins exhibited EPR signals that were not the simple sum of the signals from the separate proteins, suggesting magnetic interactions between clusters. This study highlights the potential for using genetics to simplify the study of complex multicentered metalloenzymes and to generate new complex metalloenzymes with interesting properties. PMID- 12121110 TI - Growth and structure evolution of novel tin oxide diskettes. AB - The novel SnO diskettes have been synthesized by evaporating either SnO or SnO(2) powders at elevated temperature. Disregard the source material being SnO or SnO(2), the SnO diskettes are formed at a low-temperature region of 200-400 degrees C. Two types of diskette shapes have been identified: the solid-wheel shape with a drop center rim (type I) and the diskette with cone peak(s) and spiral steps (type II). The diskettes are determined to be tetragonal SnO structure (P4/nmm), with their flat surfaces being (001). The formation of the SnO diskettes is suggested to result from a solidification process. The structural evolution from SnO diskettes to SnO(2) diskettes has been investigated by oxidizing at different temperatures. The result shows that the phase transformation from SnO to SnO(2) occurs in two processes of decomposition and oxidization, and the decomposition process consists of two steps: first from SnO to Sn(3)O(4) and then from Sn(3)O(4) to SnO(2). PMID- 12121111 TI - Mechanism of vinylic and allylic carbon-fluorine bond activation of non perfluorinated olefins using Cp*(2)ZrH(2). AB - Cp(2)ZrH(2) (1) (Cp = pentamethylcyclopentadienyl) reacts with vinylic carbon fluorine bonds of CF(2)=CH(2) and 1,1-difluoromethylenecyclohexane (CF(2)=C(6)H(10)) to afford Cp(2)ZrHF (2) and hydrodefluorinated products. Experimental evidence suggests that an insertion/beta-fluoride elimination mechanism is occurring. Complex 1 reacts with allylic C-F bonds of the olefins, CH(2)=CHCF(3), CH(2)=CHCF(2)CF(2)CF(2)CF(3), and CH(2)=C(CF(3))(2) to give preferentially 2 and CH(3)-CH=CF(2), CH(3)-CH=CF-CF(2)CF(2)CF(3), and CF(2)=C(CF(3))(CH(3)), respectively, by insertion/beta-fluoride elimination. In the reactions of 1 with CH(2)=CHCF(3) and CH(2)=CHCF(2)CF(2)CF(2)CF(3), both primary and secondary alkylzirconium olefin insertion intermediates were observed in the (1)H and (19)F NMR spectra at low temperature. A deuterium labeling study revealed that more than one olefin-dihydride complex is likely to exist prior to olefin insertion. In the presence of excess 1 and H(2), CH(2)=CHCF(3) and CH(2)=CHCF(2)CF(2)CF(2)CF(3) are reduced to propane and (E) CH(3)CH(2)CF=CFCF(2)CF(3), respectively. PMID- 12121112 TI - Regular intergrowth in the AFI-type crystals: influence on the intracrystalline adsorbate distribution as observed by interference and FTIR-microscopy. AB - Interference microscopy and FTIR microscopy are applied to study intracrystalline concentration profiles of methanol in CrAPO-5 zeolite crystals. By using both techniques, the high spatial resolution of interference microscopy is complemented by the ability of FTIR spectroscopy to pinpoint adsorbates by their characteristic IR bands. For the first time two-dimensional concentration profiles of an unprecedented quality are reported which show a nonhomogeneous distribution of adsorbate in zeolite crystal under equilibrium with the adsorbate vapor. These nonhomogeneous profiles are attributed to regular intergrowth effects in CrAPO-5. A possible internal structure of CrAPO-5 crystals is suggested. PMID- 12121113 TI - Hydrogenation without a transition-metal catalyst: on the mechanism of the base catalyzed hydrogenation of ketones. AB - The hydrogenation of unsaturated organic substrates such as olefins and ketones is usually effected by homogeneous or heterogeneous transition-metal catalysts. On the other hand, a single case of a transition-metal-free and purely base catalyzed hydrogenation of ketones was reported by Walling and Bollyky some 40 years ago. Unfortunately, the harsh reaction conditions (ca. 200 degrees C, >100 bar H(2), potassium tert-butoxide as base) limit the substrate spectrum of this reaction to robust, nonenolizable ketones such as benzophenone. We herein present a mechanistic study of this process as a basis for future rational improvement. The base-catalyzed hydrogenation of ketones was found to be irreversible, and it shows first-order kinetics with respect to the substrate ketone, hydrogen, and catalytic base. The rate of the reaction depends on the type of alkali ion present (Cs > Rb - K >> Na >> Li). Using D(2) instead of H(2) revealed a rapid base-catalyzed isotope exchange/equilibration between the gas phase and the solvent as a concomitant reaction. The degree of deuteration of the product alcohols did not indicate a significant kinetic isotope effect. It is proposed that both ketone reduction and isotope exchange proceed via similar six-membered cyclic transition states involving the H(2)(D(2))-molecule, the alkoxide base, and the ketone (solvent alcohol in the case of isotope exchange). Mechanistic analogies are pointed out which apparently exist between the base-catalyzed hydrogenation of ketones studied here and the Ru-catalyzed asymmetric ketone hydrogenation developed by Noyori. In both cases, heterolysis of the hydrogen molecule appears to be assisted by a Bronsted-base (i.e., alkoxide), the latter being bound to the substrate ketone or the catalyst ligand, respectively, by a bridging Lewis-acidic alkali ion. PMID- 12121114 TI - NMR and calculational studies on the regioselective lithiation of 1 methoxynaphthalene. AB - 1-Methoxynaphthalene (1) undergoes regioselective lithiation in position 2 (n BuLi/TMEDA) or in position 8 (t-BuLi), respectively. The detected formation of a n-BuLi/1 complex (1:1 n-BuLi/1 mixture) appears to have only minor influence on the regioselectivity (both products are obtained). The exchange of hydrogen atom H2 by deuterium results in a remarkably reduced reaction rate for the lithiation with n-BuLi in THF-d(8). This isotope effect and the formation of the thermodynamically less favorable 2-lithio compound suggest a kinetically controlled mechanism. The lack of an isotope effect for the reaction of 8 deuterio-1-methoxynaphthalene with t-BuLi and the formation of the thermodynamically preferred 8-lithiated product indicate a thermodynamically controlled mechanism. Slow conversion of the 2- into the 8-lithiated species (at higher temperatures) gives further evidence that n-BuLi and t-BuLi afford the kinetically and thermodynamically preferred products, respectively. PMID- 12121115 TI - Analysis of heparan sulfate oligosaccharides with ion pair-reverse phase capillary high performance liquid chromatography-microelectrospray ionization time-of-flight mass spectrometry. AB - Heparan sulfate, a cell surface bound glycosaminoglycan polysaccharide, has been implicated in numerous biological functions. Heparan sulfate molecules are highly complex and diverse, yet deceivingly look simple and similar, rendering structure -function correlation tedious. Current chromatographic and mass spectrometric techniques have limitations for analyzing glycosaminoglycan samples that are in low abundance and that are large in size, due to their highly acidic nature arising from a large number of sulfate and of carboxylate groups. A new methodology was developed using capillary ion-paired reverse-phase C18 HPLC directly coupled to ESI-TOF-MS to address the above issues. On the basis of HS disaccharide analysis, dibutylamine was found to be the best suited for HS analysis among many ion-pairing agents investigated. Next, analysis of oligosaccharides derived from heparosan, the precursor for heparan sulfate, was undertaken to demonstrate its greater applicability in a more complex structural analysis. The established chromatographic conditions enabled the characterization of heparosan oligosaccharides of sizes up to tetracontasaccharide with high resolution in a single run and were amenable to negative ion electrospray MS in which sodium adduction and fragmentation were avoided. To date, these are the largest nonsulfated HS precursor oligosaccharides to be characterized by LC/MS. Finally, the current methodology was applied to the characterization of the biologically important ATIII binding pentasaccharide and its precursors, which differ from each other by sulfation pattern and/or degree of sulfation. All of these pentasaccharides were well-resolved and characterized by the LC/MS system with (34)SO(4) as a mass spectral probe. This newly developed methodology facilitates the purification and rapid characterization of biologically significant HS oligosaccharides, and will thus expedite their synthesis. These findings should undoubtedly pave the way in deciphering multiple functional arrangements, ascribed to many biological activities, which are predictably embedded in a single large chaotic, yet well-organized HS polysaccharide chain. Development of newer techniques for HS oligosaccharide analysis is greatly needed in the postgenome era as attention shifts to the functional implications of proteins and carbohydrates in general and HS in particular. PMID- 12121116 TI - Kinetics and mechanism of hydroxyl radical and OH-adduct radical reactions with nitroxides and with their hydroxylamines. AB - Stable nitroxide radicals are potent antioxidants and are among the most effective non-thiol radioprotectants, although they react with hydroxyl radicals more slowly than typical phenolic antioxidants or thiols. Surprisingly, the reduced forms of cyclic nitroxides, cyclic hydroxylamines, are better reductants yet have no radioprotective activity. To clarify the reason for this difference, we studied the kinetics and mechanisms of the reactions of nitroxides and their hydroxylamines with (*)OH radicals and with OH-adducts by using pulse radiolysis, fluorimetric determination of phenolic radiation products, and electron paramagnetic resonance spectrometric determination of nitroxide concentrations following radiolysis. Competition kinetics with phenylalanine as a reference compound in pulse radiolysis experiments yielded rate constants of (4.5 +/- 0.4) x 10(9) M(-1) s(-1) for the reaction of (*)OH radical with 2,2,6,6 tetramethylpiperidine-N-oxyl (TPO), 4-hydroxy-TPO (4-OH-TPO), and 4-oxo-TPO (4-O TPO), (3.0 +/- 0.3) x 10(9) M(-1) s(-1) for deuterated 4-O-TPO, and (1.0 +/- 0.1) x 10(9) M(-1) s(-1) for the hydroxylamine 4-OH-TPO-H. The kinetic isotope effect suggests the occurrence of both (*)OH addition to the aminoxyl moiety of 4-O-TPO and H-atom abstraction from the 2- or 6-methyl groups or from the 3- and 5 methylene positions. This conclusion was further supported by final product analysis, which demonstrated that (*)OH partially oxidizes 4-O-TPO to the corresponding oxoammonium cation. The rate constants for the reactions of the nitroxides with the OH-adducts of phenylalanine and terephthalate have been determined to be near 4 x 10(6) M(-1) s(-1), whereas the hydroxylamine reacted at least 50 times slower, if at all. These findings indicate that the reactivity toward (*)OH does not explain the differences between the radioprotective activities of nitroxides and hydroxylamines. Instead, the radioprotective activity of nitroxides, but not of hydroxylamines, can be partially attributed to their ability to detoxify OH-derived secondary radicals. PMID- 12121117 TI - The elusive atomic rationale for DNA base pair stability. AB - A systematic analysis of the electrostatic interaction between 27 natural DNA base pairs was carried out, based on ab initio correlated wave functions and the topology of the electron density. Using high rank multipole moments we show that the atomic partitioning of the interaction energy contains many substantial contributions between distant atoms. Profiles of cumulative energy versus internuclear distance show large fluctuations and provide an electrostatic fingerprint of the partitioning of interaction energy in a complex. A quantified comparison between each pair of energy profiles, one for each base pair, makes clear that there is no correlation between the total base pair interaction energy and the shape of the profile. In other words, base pairs with similar interaction energy are not stable for the same reasons in terms of atomic partitioning. In summary, simple rules to rationalize the pattern of energetic stability of naturally occurring base pairs in terms of subsets of atoms are elusive. Our work cautions against inappropriate use of Jorgensen's secondary interaction hypothesis. PMID- 12121118 TI - Passive and active oxidation of Si(100) by atomic oxygen: a theoretical study of possible reaction mechanisms. AB - Reaction mechanisms for oxidation of the Si(100) surface by atomic oxygen were studied with high-level quantum mechanical methods in combination with a hybrid QM/MM (Quantum mechanics/Molecular Mechanics) method. Consistent with previous experimental and theoretical results, three structures, "back-bond", "on-dimer", and "dimer-bridge", are found to be the most stable initial surface products for O adsorption (and in the formation of SiO(2) films, i.e., passive oxidation). All of these structures have significant diradical character. In particular, the "dimer-bridge" is a singlet diradical. Although the ground state of the separated reactants, O+Si(100), is a triplet, once the O atom makes a chemical bond with the surface, the singlet potential energy surface is the ground state. With mild activation energy, these three surface products can be interconverted, illustrating the possibility of the thermal redistribution among the initial surface products. Two channels for SiO desorption (leading to etching, i.e., active oxidation) have been found, both of which start from the back-bond structure. These are referred to as the silicon-first (SF) and oxygen-first (OF) mechanisms. Both mechanisms require an 89.8 kcal/mol desorption barrier, in good agreement with the experimental estimates of 80-90 kcal/mol. "Secondary etching" channels occurring after initial etching may account for other lower experimental desorption barriers. The calculated 52.2 kcal/mol desorption barrier for one such secondary etching channel suggests that the great variation in reported experimental barriers for active oxidation may be due to these different active oxidation channels. PMID- 12121119 TI - Chemical bonding in biguanidinium dinitramide and biguanidinium bis-dinitramide from experimental X-ray diffraction data. AB - The electron density and related properties of biguanidinium dinitramide (BIGH)(DN) and biguanidinium bis-dinitramide (BIGH(2))(DN)(2) crystals (space groups P1 and C2/c) have been determined from low-temperature (90(1) K) X-ray diffraction experiments. The Hansen-Coppens multipole model as implemented in the XD program gave R = 0.0247 and 0.0201 (all reflections) which allowed the calculation of the electron density and Laplacian distributions. The bonding (3, 1) critical points were also found. The analysis of the results shows that Bader's topological theory provides a more useful description of the chemical bonding in the studied crystals as compared to the classical analysis of deformation densities. The hydrogen bonding in the crystals was analyzed. The atomic charges were integrated over the atomic basins. PMID- 12121120 TI - Investigations of polyelectrolyte adsorption at the solid/liquid interface by sum frequency spectroscopy: evidence for long-range macromolecular alignment at highly charged quartz/water interfaces. AB - IR-visible sum frequency spectroscopy (SFS) was employed to investigate the molecular level details of the adsorption of the positively charged polyelectrolyte, polydiallyldimethylammonium chloride (PDDA), at the quartz/water interface. Below pH 9.0, signal from the interfacial water structure was visible, but none from the adsorbed polymer could be detected. This indicated that the PDDA was not well enough aligned at the interface under these conditions to elicit a sum frequency response. At more basic pH values (>or=9.6), however, adsorbed PDDA molecules became well-ordered as indicated by the presence of CH stretch peaks from methylene and methyl groups. The intensities of the CH stretch modes were independent of the adsorbed amount of PDDA at pH 12.3 but decreased as the pH of the bulk solution was lowered. The conditions for polymer alignment fell outside the parameters where layer-by-layer growth of oppositely charged polyelectrolytes was possible because the net charge on the surface under high pH conditions remained negative. PMID- 12121121 TI - New group 2 chemistry: a multiple barium-nitrogen bond in the CsNBa molecule. AB - The existence of a series of triatomic molecules with the general formula MNM', where M is an alkaline metal (K, Rb, Cs), and M' is an alkaline earth metal (Ca, Sr, Ba), has been predicted by quantum chemical methods. Among these, the CsNBa molecule shows a feature not found before, the presence of a multiple bond between barium and nitrogen. As a consequence of this novel bonding situation, the molecule is linear. The same holds for all Ba triatomics, MNBa, independent of the nature of the alkali M atom, and for all Sr compounds, MNSr. The presence of a multiple bond makes CsNBa, and other related Ba and Sr molecules, particularly stable and appealing experimentally. The systems with the alkaline earth metal M' = Ca, on the other hand, turned out to be bent. Calculations have also been performed on the negative ions BaN(-) and CaN(-), which form a well defined entity in the MNM' systems (M' = Ba, Ca). The results show that the two ions have a different electronic structure in the ground state, which is one reason for the different properties of the MNM' systems and explains why the molecules containing the BaN(-) moiety are linear, while those containing CaN(-) are bent. PMID- 12121123 TI - Novel cyanine dye-labeled dideoxynucleoside triphosphates for DNA sequencing. AB - Single color cyanine dye-labeled (Cy 5.0 and Cy 5.5) dideoxynucleoside-5' triphosphates, or 'terminators', containing different spacer lengths were synthesized and evaluated for efficacy in DNA sequencing methods using a modified thermally stable DNA polymerase. The single color cyanine dye terminators were formulated into two separate sets of sequencing mixes, one for Cy 5.0 and the other for Cy 5.5, and evaluated on different automated sequencing platforms. Each set of mixes included two pyrimidine terminators with 17-atom linkers and two purine terminators with 10-atom linkers between the dye and the nucleotide. The two sets of cyanine dye-labeled terminators chosen for this cycle sequencing study produced improved band patterns with band uniformity similar to that obtained with dye-primer sequencing methods. PMID- 12121124 TI - Establishment of intein-mediated protein ligation under denaturing conditions: C terminal labeling of a single-chain antibody for biochip screening. AB - Intein-mediated protein ligation is a recently developed method that enables the C-terminal labeling of proteins. This technique requires a correctly folded intein mutant that is fused to the C-terminus of a target protein to create a thioester, which allows the ligation of a peptide with an N-terminal cysteine (1, 2). Here we describe the establishment of this method for the labeling, under denaturing conditions, of target proteins that are expressed insolubly as intein fusion proteins. A GFPuv fusion protein with the Mycobacterium xenopi gyrA intein was expressed in inclusion bodies in Escherichia coli and initially used as a model protein to verify intein cleavage activity under different refolding conditions. The intein showed activity after refolding in nondenaturing and slightly denaturing conditions. A construct of the same intein with an anti neutravidin single-chain antibody was also expressed in an insoluble form. The intein-mediated ligation was established for this single chain antibody-intein fusion protein under denaturing conditions in 4 M urea to prevent significant precipitation of the fusion protein during the first refolding step. Under optimized conditions, the single-chain antibody was labeled with a fluorescent peptide and used for antigen screening on a biochip after final refolding. This screening procedure allowed the determination of binding characteristics of the scFv for avidin proteins in a miniaturized format. PMID- 12121125 TI - Peptide arrays for highly sensitive and specific antibody-binding fluorescence assays. AB - We report a novel generation of peptide arrays fabricated by site-specific ligation of glyoxylyl peptides onto glass slides covered by a semicarbazide sol gel layer. These arrays allowed the highly sensitive and specific detection of antibodies in very small blood samples from infected individuals using three model peptidic epitopes (HCV Core and NS4, EBV Capsid) in an immunofluorescence assay. Comparison with standard enzyme-linked immunosorbent assays (ELISAs) demonstrated a large gain in sensitivity and specificity. These unique properties, combined with the possibility to immobilize glycoproteins such as antibodies, offer the possibility to perform sandwich immunofluorescent assays in a highly parallel format. PMID- 12121127 TI - Quantitation of the tumor-targeting properties of antibody fragments conjugated to cell-permeating HIV-1 TAT peptides. AB - Human monoclonal antibodies are promising agents for the development of more selective anticancer therapeutics. However, the tumor-targeting efficiency of most anticancer antibodies is severely limited by their poor penetration into the tumor mass. Recent studies have shown that a peptide derived from the HIV TAT protein could improve the distribution of cytoplasmic reporter proteins when administered systemically as fusion proteins or cross-linked chimeras. In this article, we tested by quantitative biodistribtution analysis whether conjugation to TAT peptides could improve the tumor targeting properties of scFv(L19)-Cys: an engineered human antibody fragment specific for the ED-B domain of fibronectin, a marker located in the modified extracellular matrix surrounding tumor neovasculature. Our results show that TAT peptides, consisting either of L-amino acids or D-amino acids, can efficiently transduce target cells when conjugated to fluorophores and/or antibody fragments, suggesting a receptor-independent cell entry mechanism. However, conjugation of scFv(L19)-Cys to TAT peptides resulted in a severely reduced tumor targeting performance compared to the unconjugated antibody, as measured in murine F9 teratocarcinoma-bearing mice, after intravenous injection of the radiolabeled antibody preparations. Our results outline the usefulness of TAT peptides for the efficient in vitro transduction of cells with globular proteins. In particular, the use of TAT peptides composed of D-amino acids may significantly reduce proteolytic degradation. At the same time, the poor biodistribution properties of antibody-TAT conjugates cast doubts over the applicability of this methodology for the delivery of biopharmaceuticals in vivo. PMID- 12121126 TI - DOTA-D-Tyr(1)-octreotate: a somatostatin analogue for labeling with metal and halogen radionuclides for cancer imaging and therapy. AB - The goal of this study was to evaluate a somatostatin receptor ligand, DOTA-D Tyr(1)-octreotate (DOTA-DY1-TATE), that has the chelator 1,4,7,10 tetraazacyclotetradecane-N,N',N'',N'"-tetraacetic acid (DOTA) attached to the D Tyr(1) residue, allowing radiolabeling with both radiohalogens and radiometals. A potential advantage of having a chelator attached to the Tyr(1) residue is that halogen radiolabels may residualize or remain trapped in tumor cells rather than clear from the tumor. DOTA-DY1-TATE was synthesized by solid-phase methods and radiolabeled with (61)Cu, (64)Cu, and (125)I in high radiochemical purity and specific activity. A competitive binding assay demonstrated that (nat)Cu-DOTA-DY1 TATE and DOTA-(nat)I-DY1-TATE had comparable affinity to (nat)In-DTPA-OC in AR42J rat pancreatic tumor cells membranes. (61)Cu-DOTA-DY1-TATE had a dissociation constant (K(d)) of 176.4 pM and a receptor concentration (B(max)) of 244.4 fmol/mg. A tumor uptake of 1.515 %ID/g was determined for (64)Cu-DOTA-DY1-TATE and 0.814 %ID/g for DOTA-(125)I-DY1-TATE in AR42J tumor bearing Lewis rats at 1 h postinjection. DOTA-(125)I-DY1-TATE remained in the tumor at a higher concentration out to 4 h postinjection, suggesting that the iodine may have residualized in the tumor cells. MicroPET imaging of (64)Cu-DOTA-DY1-TATE in AR42J tumor bearing rats and SCID mice at 2 h postinjection showed significant uptake and good contrast in the thigh tumors in the rat model and in the neck and thigh tumors of the mouse. This study demonstrates that DOTA-DY1-TATE is a somatostatin analogue that can be labeled with both metal and halogen radionuclides, and its (64)Cu- and (125)I-radiolabeled compounds showed somatostatin receptor-mediated uptake in normal and tumor tissues. PMID- 12121128 TI - Development of EGF-conjugated liposomes for targeted delivery of boronated DNA binding agents. AB - Liposomes are of interest as drug delivery tools for therapy of cancer and infectious diseases. We investigated conjugation of epidermal growth factor, EGF, to liposomes using the micelle-transfer method. EGF was conjugated to the distal end of PEG-DSPE lipid molecules in a micellar solution and the EGF-PEG-DSPE lipids were then transferred to preformed liposomes, either empty or containing the DNA-binding compound, water soluble acridine, WSA. We found that the optimal transfer conditions were a 1-h incubation at 60 degrees C. The final conjugate, (125)I-EGF-liposome-WSA, contained approximately 5 mol % PEG, 10-15 EGF molecules at the liposome surface, and 10(4) to 10(5) encapsulated WSA molecules could be loaded. The conjugate was shown to have EGF-receptor-specific cellular binding in cultured human glioma cells. PMID- 12121129 TI - Cholesterol inclusion in liposomes affects induction of antigen-specific IgG and IgE antibody production in mice by a surface-linked liposomal antigen. AB - In the previous study, we investigated the induction of ovalbumin (OVA)-specific antibody production in mice by OVA-liposome conjugates made using four different lipid components, including unsaturated carrier lipid and three different saturated carrier lipids. All of the OVA-liposome conjugates tested induced IgE selective unresponsiveness. The highest titer of anti-OVA IgG was observed in mice immunized with OVA-liposomes made using liposomes with the highest membrane fluidity, suggesting that the membrane fluidity of liposomes affects their adjuvant effect. In this study, liposomes with five different cholesterol inclusions, ranging from 0% to 43% of the total lipid, were made, and the induction of OVA-specific antibody production by OVA-liposome conjugates was compared among these liposome preparations. In contrast to the results in the previous study, liposomes that contained no cholesterol and possessed the lowest membrane fluidity demonstrated the highest adjuvant effect for the induction of IgG antibody production. In addition, when the liposomes with four different lipid compositions were used, OVA-liposome conjugates made using liposomes that did not contain cholesterol induced significantly higher levels of anti-OVA IgG antibody production than did those made using liposomes that contained cholesterol and, further, induced significant production of anti-OVA IgE. These results suggest that cholesterol inclusion in liposomes affects both adjuvanticity for IgG production and regulatory effects on IgE synthesis by the surface-coupled antigen of liposomes. PMID- 12121130 TI - Steps toward high specific activity labeling of biomolecules for therapeutic application: preparation of precursor [(188)Re(H(2)O)(3)(CO)(3)](+) and synthesis of tailor-made bifunctional ligand systems. AB - Two kit preparations of the organometallic precursor [(188)Re(H(2)O)(3)(CO)(3)](+) in aqueous media are presented. Method A uses gaseous carbon monoxide and amine borane (BH(3).NH(3)) as the reducing agent. In method B CO(g) is replaced by K(2)[H(3)BCO(2)] that releases carbon monoxide during hydrolysis. Both procedures afford the desired precursor in yields >85% after 10 min at 60 degrees C. HPLC and TLC analyses revealed 7 +/- 3% of unreacted (188)ReO(4)(-) and <5% of colloidal (188)ReO(2). Solutions of up to 14 GBq/mL Re-188 have been successfully carbonylated with these two methods. The syntheses of two tailor-made bifunctional ligand systems for the precursor [(188)Re(H(2)O)(3)(CO)(3)](+) are presented. The tridentate chelates consist of a bis[imidazol-2-yl]methylamine or an iminodiacetic acid moiety, respectively. Both types of ligand systems have been prepared with alkyl spacers of different length and a pendent primary amino or carboxylic acid functionality, enabling the amidic linkage to biomolecules. The tridentate coordination of the ligands to the rhenium-tricarbonyl core could be elucidated on the macroscopic level by X-ray structure analyses and 1D and 2D NMR experiments of two representative model complexes. On the nca level, the ligands allow labeling yields >95% with [(188)Re(H(2)O)(3)(CO)(3)](+) under mild reaction conditions (PBS buffer, 60 degrees C, 60 min) at ligand concentrations between 5 x 10(-4) M and 5 x 10(-5) M. Thus, specific activities of 22-220 GBq pe micromol of ligand could be achieved. Incubation of the corresponding Re-188 complexes in human serum at 37 degrees C revealed stabilities between 80 +/- 4% and 45 +/- 10% at 24 h, respectively, and 63 +/- 3% and 34 +/- 3% at 48 h postincubation in human serum depending on the chelating system. Decomposition product was mainly (188)ReO(4)( ). The routine kit-preparation of the precursor [(188)Re(H(2)O)(3)(CO)(3)](+) in combination with tailor-made ligand systems enables the organometallic labeling of biomolecules with unprecedented high specific activities. PMID- 12121131 TI - Novel poly(ethylene glycol) derivatives for preparation of ribosome-inactivating protein conjugates. AB - This study describes the synthesis, characterization, and reactivity of new methoxypoly(ethylene glycol) (mPEG) derivatives containing a thioimidoester reactive group. These activated polymers are able to react with the lysyl epsilon amino groups of suitable proteins, generating an amidinated linkage and thereby preserving the protein's positive charge. mPEG derivatives of molecular weight 2000 and 5000 Da were used, and two spacer arms were prepared, introducing chains of different lengths between the hydroxyl group of the polymer and the thioimidate group. These mPEG derivatives were used to modify gelonin, a cytotoxic single-chain glycoprotein widely used in preparation of antitumoral conjugates, whose biological activity is strongly influenced by charge modification. The reactivity of mPEG thioimidates toward lysil epsilon-amino groups of gelonin was evaluated, and the results showed an increased degree of derivatization in proportion to the molar excesses of the polymer used and to the length of the alkyl spacer. Further studies showed that the thioimidate reactive is able to maintain gelonin's significant biological activity and immunogenicity. On the contrary, modification of the protein with N-hydroxysuccinimide derivative of mPEG strongly reduces the protein's cytotoxic activity. Evaluation of the pharmacokinetic behavior of native and PEG-grafted gelonin showed a marked increase in plasma half-life after protein PEGylation; in particular, the circulating life of the conjugates increased with increased molecular weight of the polymer used. The biodistribution test showed lower organ uptake after PEGylation, in particular by the liver and spleen. PMID- 12121132 TI - Identification of two tamoxifen target proteins by photolabeling with 4-(2 morpholinoethoxy)benzophenone. AB - Our quest to identify target proteins involved in the activity of tamoxifen led to the design of photoaffinity ligand analogues of tamoxifen able to cross-link such proteins. A new tritiated photoprobe, 4-(2-morpholinoethoxy)benzophenone (MBoPE), was synthesized and used to identify proteins involved in tamoxifen binding in rat liver. MBoPE, which has structural features in common with the potential antagonist of the intracellular histamine receptor (N,N-diethyl-2-[(4 phenylmethyl)phenoxy]ethanamine HCl: DPPE) is unable to bind the estrogen receptor although it does compete with tamoxifen for an antiestrogen binding site (AEBS). This tritiated benzophenone derivative was obtained by metal-catalyzed halogen-tritium replacement reaction. Because of its high specific activity, four target proteins could be photolabeled, three of which were identified with M(r) of 60,000, 49,500, and 14,000, while the fourth at 27,500 was in too low an amount and could not be sequenced. The 49.5 kDa protein corresponded by mass spectrometry to the microsomal epoxide hydrolase already identified with an aryl azide photoprobe [Mesange, F., et al. (1998) Biochem. J. 334, 107-112]. The 60 and 14 kDa proteins were identified as the carboxylesterase (ES10) and the liver fatty acid binding protein (L-FABP), respectively. The inhibitory effect of tamoxifen on carboxylesterase activity and the competitive efficacy of oleic acid on [(3)H]tamoxifen binding suggest that both proteins are AEBS subunits. Moreover, treatment of hepatocytes with antisense mRNA directed against ES10 or L FABP abolished both tamoxifen and MBoPE binding. On the basis of previous pharmacological arguments, the 27.5 kDa protein might correspond to the sigma I receptor. Altogether, these results confirm that the microsomal epoxide hydrolase is a target for tamoxifen and provide evidence of two new target proteins implicated in cell lipid metabolism. PMID- 12121133 TI - Polyethylene glycol conjugates of methotrexate varying in their molecular weight from MW 750 to MW 40000: synthesis, characterization, and structure-activity relationships in vitro and in vivo. AB - Poly(ethylene glycol)s (PEGs) are potential drug carriers for improving the therapeutic index of anticancer agents. In this work, the anticancer drug methotrexate (MTX) was activated with N,N'-dicyclohexylcarbodiimide (DCC) and coupled to amino group bearing PEGs of MW 750, 2000, 5000, 10 000, 20,000, and 40,000. First, the activation process of MTX with DCC in the presence and absence of N-hydroxysuccinimide was analyzed through HPLC. Preincubation of methotrexate with DCC alone at 0 degrees C proved to be favorable with respect to the amount of activated species and the formation of byproducts. MTX-PEG conjugates were synthesized according to this procedure, isolated through size-exclusion chromatography, and characterized through analytical HPLC, MALDI-TOF spectrometry, and gel permeation chromatography. In a cell-free assay, all of the drug polymer conjugates inhibited the target enzyme of MTX, dihydrofolate reductase (DHFR), to a similar extent, but were not as active as free MTX. Additionally, incubation of the MTX-PEG40000 conjugate for 6 days at 37 degrees C in phosphate buffered saline (pH 7.4), in cell-conditioned medium, or in human serum revealed no significant release of methotrexate. These results, taken together, indicate that release of MTX from polymer conjugates is not necessary for an effective interaction with the active site of dihydrofolate reductase. Evaluation of the in vitro cytotoxicity of the MTX-PEG conjugates in two adherent and three suspension human tumor cell lines revealed that the IC(50) values of the tested compounds increased with the size of the drug-polymer conjugates. The most effective compound tested in these assays was the free drug MTX itself (IC(50) value ranging from approximately 0.01 to 0.05 microM), while the IC(50) values of the polymer conjugates were higher (IC(50) value for MTX-PEG750, 2000 and 5000: approximately 0.6-3 microM; for MTX-PEG10000 and 20000: approximately 2 7 microM; and for MTX-PEG40000: > 6 microM). Subsequently, MTX-PEG5000, MTX PEG20000, and MTX-PEG40000 were evaluated in a human mesothelioma MSTO-211H xenograft model, and their antitumor effects were compared with free methotrexate and the albumin conjugate MTX-HSA, a conjugate that is currently in phase II clinical trials. In contrast to the in vitro results, the high molecular weight MTX-PEG conjugates exhibited the highest in vivo antitumor activity: At a dose of 40 and 80 mg/kg MTX-PEG5000 was less active than MTX at its optimal dose of 100 mg/kg; MTX-PEG20000 at a dose of 40 mg/kg showed antitumor efficacy comparable to MTX, but MTX-PEG40000 at a dose of 20 mg/kg was superior to MTX and demonstrated antitumor activity of the same order as MTX-HSA (20 mg/kg). PMID- 12121134 TI - Modifications in synthesis strategy improve the yield and efficacy of geldanamycin-herceptin immunoconjugates. AB - Geldanamycin (GA) was modified with N-tert-butyloxycarbonyl-1,3-diaminopropane to introduce a latent primary amine. After deprotection, this primary amine provided a site for introduction of a maleimide group that enabled linkage to proteins. This maleimido derivative of geldanamycin (GMB-APA-GA) was linked to the monoclonal antibody Herceptin after the antibody had been modified with Traut's reagent to introduce thiol groups. By this sequence, a new immunoconjugate (H:APA GA) was generated that showed greater antiproliferative activity than the previously reported analogous immunoconjugate created with a 1,4-diaminobutane spacer derivative of geldanamycin to form an immunoconjugate, H:ABA-GA. Both immunoconjugates inhibited in vitro the growth of MDA-361/DYT2 cells, a cell line overexpressing the HER2 antigen, while Herceptin alone was ineffective. However, H:APA-GA showed better efficacy than H:ABA-GA (IC(50) = 0.2 vs 0.58 mg/mL and cell doubling time >12 vs 6 days, respectively). Results of the in vivo therapy experiments in a xenograft model were consistent with the in vitro findings. Treatment with Herceptin prolonged the survival of the tumor-bearing mice when compared with the control group, but H:ABA-GA and H:APA-GA were each more efficacious than unmodified Herceptin. However, unlike H:ABA-GA, the immunoconjugate H:APA-GA caused stable tumor regression (in 25% of the recipients), showing a qualitative improvement with potential clinical relevance. PMID- 12121135 TI - Selective adsorption of serum albumin on biomedical polyurethanes modified by a poly(ethylene oxide) coupling-polymer with cibacron blue (F3G-A) endgroups. AB - A tri-block-coupling polymer, "PEO-MDI-PEO" ["poly(ethylene oxide)-4,4'-methylene diphenyl diisocyanate-poly(ethylene oxide)", abbreviated "MPEO"], was used to react with a triazine dye, Cibacron Blue F3G-A (ciba), in an alkaline environment. The product of this nucleophilic reaction was a penta-block-coupling polymer, "ciba-PEO-MDI-PEO-ciba" (abbreviated "cibaMPEO"). The cibaMPEO-modified poly(ether urethane) (PEU) surfaces were prepared by dip-coating and detected by XPS. The surface enrichment of both ciba endgroups and poly(ethylene oxide) spacer-arms was revealed. On the modified surfaces, bovine serum albumin (BSA) adsorbing experiments were carried out, respectively, in the low and high BSA bulk-concentration solutions, and accordingly, the methods of radioactive (125)I probe and ATR-FTIR were, respectively, employed for the characterization. The competitive adsorption of BSA and bovine serum fibrinogen (Fg) in the BSA-Fg binary solutions was also studied using a (125)I-probe, and through which the reversibly BSA-selective adsorption on cibaMPEO-modified PEU surfaces was confirmed. Finally, the improvement of blood-compatibility on the modified surfaces was verified by the plasma recalcification time (PRT) test. PMID- 12121136 TI - Chemical coupling of a monoclonal antisurfactant protein-B antibody to human urokinase for targeting surfactant-incorporating alveolar fibrin. AB - Intraalveolar fibrin formation is a common histopathological finding in acute inflammatory and chronic interstitial lung diseases. Incorporation of hydrophobic surfactant components into polymerizing fibrin results in a severe loss of surface activity, altered mechanical and structural clot properties, and a reduced susceptibility toward fibrinolytic degradation. Such events have been implicated in atelectasis formation, impairment of gas exchange, and provocation of fibroproliferative changes. In an effort to address the unique features of alveolar fibrin, we designed a hybrid molecule consisting of a monoclonal antibody against surfactant protein SP-B (8B5E) and the catalytic domain of urokinase (B-chain), which was termed MABUC. The urokinase B-chain was prepared by limited reduction of human two-chain-urokinase and subsequent affinity purification and coupled to the antibody using a heterobifunctional cross-linker. Purification of the chimeric protein included gel filtration chromatography and affinity chromatography. An ELISA-like microtiter plate assay, based on the immunological detection of the SP-B moiety and the fibrinolytic activity of the u PA domain, was developed for the detection of the hybrid molecule. Chromogenic substrate assays, (125)I-based fibrin plate assays, and active site titration were performed to analyze the specific fibrinolytic activity of the conjugate. MABUC was found to fully retain the ability of SP-B binding and the fibrinolytic activity of u-PA. In addition, MABUC was noted to be 1.5-2-fold more effective in the dissolution of surfactant embedding clots and to be approximately 3-fold more resistant against PAI-1, the predominant fibrinolysis inhibitor in the alveolar compartment, as compared to the native u-PA. The superiority of MABUC was particularly prominent (>5-fold efficacy) when investigating clot material incorporating both PAI-1 and surfactant, as a mimicry of alveolar fibrin. We conclude that urokinase and 8B5E can be cross-linked chemically, thus yielding a fibrinolytic enzyme with enhanced substrate specifity for surfactant-containing clots and higher PAI-1 resistance as compared to native u-PA. PMID- 12121137 TI - Poly(ethylenimine-co-L-lactamide-co-succinamide): a biodegradable polyethylenimine derivative with an advantageous pH-dependent hydrolytic degradation for gene delivery. AB - A biodegradable gene transfer vector has been synthesized by linking several low molecular weight (MW) polyethylenimine (PEI, 1200 Da) blocks using an oligo(L lactic acid-co-succinic acid) (OLSA, 1000 Da). The resulting copolymer P(EI-co LSA) (8 kDa) is soluble in water and degrades via base-catalyzed hydrolytic cleavage of amide bonds. With regard to its application as a gene transfer agent, the polymer showed an interesting pH dependency of degradation. At pH 5, when DNases are highly active, the degradation proceeds at a slower rate than at a physiological pH of 7.4. PEI and P(EI-co-LSA) spontaneously formed complexes with plasmid DNA. Whereas the complexes formed with PEI were not stable and aggregated, forming particles of up to 1 microm hydrodynamic diameter, P(EI-co LSA) formed complexes, which were about 150 nm in size and of narrow size distribution. The latter complexes were stable, due to their high surface charge (zeta-potential + 18 mV). Similar to low MW PEI, the copolymer exhibited a low toxicity profile. At the same time, the copolymer showed a significant enhancement of transfection activity in comparison to the low MW PEI. This makes P(EI-co-LSA) a promising candidate for long-term gene therapy where biocompatibility and biodegradability become increasingly important. PMID- 12121138 TI - Synthesis of peptide-oligonucleotide conjugates with single and multiple peptides attached to 2'-aldehydes through thiazolidine, oxime, and hydrazine linkages. AB - 2'-Deoxyoligonucleotides and 2'-O-methyloligoribonucleotides carrying one or more 2'-aldehyde groups were synthesized and coupled to peptides containing an N terminal cysteine, aminooxy, or hydrazide group to give peptide-oligonucleotide conjugates incorporating single or multiple peptides in good yield. The facile conjugation method allows specific coupling in aqueous solution of unprotected oligonucleotides containing aldehyde groups to unprotected N-terminally modified peptides and other small molecules. A 12-mer 2'-O-methyloligoribonucleotide complementary to the HIV-1 TAR RNA stem-loop and containing two conjugated copies of an 8-mer model laminin peptide was hardly affected in TAR RNA binding and showed a similar level of inhibition of HIV-1 Tat-dependent in vitro transcription compared to the unconjugated 2'-O-methyloligoribonucleotide. Advantages of this conjugation method include (1) the ability to attach more than one peptide or other small molecule to oligonucleotide at defined nucleoside residue locations; (2) a conjugation route that does not affect significantly oligonucleotide binding to RNA structures; and (3) three alternative, facile, and mild conjugation reaction types that do not require use of a large excess of peptide reagent. PMID- 12121139 TI - Intracellular delivery of nanometric DNA particles via the folate receptor. AB - The size of condensed DNA particles is a key determinant for both diffusion to target cells in vivo and intracellular trafficking. The smallest complexes are obtained when each DNA molecule collapses individually. This was achieved using a designed cationic thiol-detergent, tetradecyl-cysteinyl-ornithine (C(14)COrn). The resulting particles were subsequently stabilized by air-induced dimerization of the detergent into a disulfide lipid on the DNA template. Particles are anionic (zeta potential = -45 mV), and their size (30 nm) corresponds to the volume of a single plasmid DNA molecule. The electrophoretic mobility of the condensed DNA, though quasi-neutralized, was found higher than that of the extended DNA. Moreover, the dimerized (C(14)COrn)(2) lipid was found to be an efficient transfection reagent for various cell lines. In an attempt to achieve extended circulation times and to target tumors by systemic delivery, we have coated the particles with PEG-folate residues. Plasmid DNA was condensed into monomolecular particles as described above and coated by simple mixing with DPPE PEG-folate. Physicochemical measurements showed particles coated with 2% of DPPE PEG(3400)-folate remain monomolecular and are stable in the cell-culture medium. Caveolae-mediated cell entry was demonstrated by ligand-dependence, by competition with excess folic acid as well as by confocal microscopy. PMID- 12121140 TI - Differential conjugation of tat peptide to superparamagnetic nanoparticles and its effect on cellular uptake. AB - Surface modification of superparamagnetic contrast agents with HIV-1 tat peptide has emerged as a promising means for intracellular magnetic labeling and noninvasive tracking of a large number of cell types with MRI. To achieve efficient intracellular delivery of the nanoparticles, we investigated the effect on cellular uptake of superparamagnetic iron oxide particles by varying the number of attached tat peptides. First, we report here a modified P2T method in measuring the numbers of surface attachments per particle through disulfide linkage. The method was shown to have desirable simplicity and reproducibility. With the P2T method as a tool, conjugates with progressively higher ratios of peptide-to-particle were synthesized. We were able to demonstrate that higher numbers of tat peptide facilitate the cellular uptake of iron oxide nanoparticles in a nonlinear fashion. Cells labeled with these optimized preparations were readily detectable by MR imaging. The increase in sensitivity could allow in vivo tracking of 100-fold lower cell concentration than currently described. PMID- 12121141 TI - Polyethylenimine-graft-poly(ethylene glycol) copolymers: influence of copolymer block structure on DNA complexation and biological activities as gene delivery system. AB - For two series of polyethylenimine-graft-poly(ethylene glycol) (PEI-g-PEG) block copolymers, the influence of copolymer structure on DNA complexation was investigated and physicochemical properties of these complexes were compared with the results of blood compatibility, cytotoxicity, and transfection activity assays. In the first series, PEI (25 kDa) was grafted to different degrees of substitution with PEG (5 kDa) and in the second series the molecular weight (MW) of PEG was varied (550 Da to 20 kDa). Using atomic force microscopy, we found that the copolymer block structure strongly influenced the DNA complex size and morphology: PEG 5 kDa significantly reduced the diameter of the spherical complexes from 142 +/- 59 to 61 +/- 28 nm. With increasing degree of PEG grafting, complexation of DNA was impeded and complexes lost their spherical shape. Copolymers with PEG 20 kDa yielded small, compact complexes with DNA (51 +/- 23 nm) whereas copolymers with PEG 550 Da resulted in large and diffuse structures (130 +/- 60 nm). The zeta-potential of complexes was reduced with increasing degree of PEG grafting if MW >or= 5 kDa. PEG 550 Da did not shield positive charges of PEI sufficiently leading to hemolysis and erythrocyte aggregation. Cytotoxicity (lactate dehydrogenase assay) was independent of MW of PEG but affected by the degree of PEG substitution: all copolymers with more than six PEG blocks formed DNA complexes of low toxicity. Finally, transfection efficiency of the complexes was studied. The combination of large particles, low toxicity, and high positive surface charge as in the case of copolymers with many PEG 550 Da blocks proved to be most efficient for in vitro gene transfer. To conclude, the degree of PEGylation and the MW of PEG were found to strongly influence DNA condensation of PEI and therefore also affect the biological activity of the PEI-g-PEG/DNA complexes. These results provide a basis for the rational design of block copolymer gene delivery systems. PMID- 12121142 TI - Cathepsin B-labile dipeptide linkers for lysosomal release of doxorubicin from internalizing immunoconjugates: model studies of enzymatic drug release and antigen-specific in vitro anticancer activity. AB - The anticancer drug doxorubicin (DOX) has been linked to chimeric BR96, an internalizing monoclonal antibody that binds to a Lewis(y)-related, tumor associated antigen, through two lysosomally cleavable dipeptides, Phe-Lys and Val Cit, giving immunoconjugates 72 and 73. A self-immolative p aminobenzyloxycarbonyl (PABC) spacer between the dipeptides and the DOX was required for rapid and quantitative generation of free drug. DOX release from model substrate Z-Phe-Lys-PABC-DOX 49 was 30-fold faster than from Z-Val-Cit-PABC DOX 42 with the cysteine protease cathepsin B alone, but rates were identical in a rat liver lysosomal preparation suggesting the participation of more than one enzyme. Conjugates 72 and 73 showed rapid and near quantitative drug release with cathepsin B and in a lysosomal preparation, while demonstrating excellent stability in human plasma. Against tumor cell lines with varying levels of BR96 expression, both conjugates showed potent, antigen-specific cytotoxic activity, suggesting that they will be effective in delivering DOX selectively to antigen expressing carcinomas. PMID- 12121143 TI - Introduction of lanthanide(III) chelates to oligopeptides on solid phase. AB - The synthesis of oligopeptide building blocks for the introduction of nonluminescent and luminescent lanthanide(III) chelates to the oligopeptide structure on the solid phase is described. The oligopeptide conjugates synthesized were used in DELFIA-based receptor binding assay (motilin) as well as in LANCE time-resolved fluorescence quenching assay (caspase-3). PMID- 12121144 TI - Synthesis of nonluminescent lanthanide(III) chelates tethered to an aminooxy group and their applicability to biomolecule derivatization. AB - Synthesis of nonluminescent lanthanide(III) chelates tethered to an aminooxy group (i.e., 1-[4-(6-aminooxyhexamido)benzyl]diethylenetriaminetetraacetic acid lanthanides(III), 6a-d, where Ln(3+) is Eu, Dy, Sm, and Tb) is described. Their applicability to biomolecule derivatization is demonstrated by allowing them to react with a synthetic oligopeptide, a protein, two synthetic drugs, and a steroid. The oligopeptide and protein were linked to 6 after preoxidation of their N-terminal serine residues, while the drugs and the steroid reacted via their ketone functionality. Also some application data is included. PMID- 12121145 TI - 99mTc-Labeling of a hydrazinonicotinamide-conjugated LTB(4) receptor antagonist useful for imaging infection and inflammation. AB - This report describes the (99m)Tc labeling of a hydrazinonicotinamide (HYNIC) conjugated LTB(4) receptor antagonist (SG380). The ternary ligand technetium complex [(99m)Tc(SG38)(tricine)(TPPTS)] (RP517) was prepared using a non-SnCl(2) containing formulation ((2001) J. Pharm. Sci. 90, 114-123). Unlike other HYNIC conjugated small biomolecules, SG380 is lipophilic and has low solubility in the kit matrix. Using a combination of a solubilizing agent (Lysolecithin) and a cosolvent (ethanol), we have developed a new formulation for routine preparation of RP517. Using this formulation, RP517 can be prepared in high radiochemical purity (RCP > 90%) and remains stable in the kit matrix for at least 6 h. We also prepared the corresponding (99)Tc analogue, [(99)Tc]RP517. An HPLC concordance experiment using RP517 and [(99)Tc]RP517 showed that the same technetium complex was prepared at both the tracer and macroscopic levels. The LC-MS data are completely consistent with the 1:1:1:1 composition for Tc:SG380:tricine:TPPTS. PMID- 12121146 TI - Combined thioether/hydrazone chemoselective ligation reactions for the synthesis of glycocluster-antigen peptide conjugates. AB - Hydrazone/thioether ligation reactions show promise for the synthesis of clustered glycosides-antigen conjugates. Due to its propensity to aggregate, tetanus toxoid-derived epitopic peptide TT(830-846) was elected to further evaluate this three-component ligation process. This difficult sequence was supplemented by a hydrazine or a glyoxylyl group either at its C- or N-terminus. The peptide-hydrazines or peptide-aldehydes thus obtained were coupled with glyoxylyl- (or hydrazino-) N-chloroacetylated-L-lysinyl trees and 2-thioethyl alpha-D-mannopyranoside. As anticipated the ligations were controlled by the nature of the peptide and proved difficult for the C-terminal aldehyde derivative. However, when the process was performed in absence of buffer and using mannitol as a dispersing agent, all combinations finally led to the expected glycoconjugates in 40-60% purified yields. PMID- 12121147 TI - Improving the labeling of S-acetyl NHS-MAG(3)-conjugated morpholino oligomers. AB - S-Acetyl MAG(3) (S-acetylmercaptoacetyltriglycine) has been used as a chelator for the (99m)Tc labeling of a variety of biomolecules. The objective of this study was to improve upon the labeling of morpholino (MORF), a DNA analogue, as a model biomolecule. A 15mer MORF with a primary amine was conjugated with NHS MAG(3) in the usual manner, and the MORF-MAG(3) was purified over a P4 column as before. The conjugate was radiolabeled using stannous ion as usual, and the impurities were identified using size exclusion high-performance liquid chromatography (SE HPLC). Various methods were then investigated to remove the impurities. With tartrate as the transchelator, two impurities were identified as labeled MAG(3) and labeled tartrate. The labeled MAG(3) could not be removed by simply repurifying the conjugate using the usual pH 5.2 NH(4)OAc buffer before labeling. However, this impurity could be completely removed if the conjugate was adjusted to pH 7.6 and heated before repurification. The labeled tartrate impurity was removed by heating during labeling. On the basis of these observations, the following procedure for purification of the conjugation mixture and subsequent labeling was adopted. After MORF was conjugated with NHS-MAG(3) and purified over P4 with pH 5.2 NH(4)OAc eluant, the oligomer fractions were combined, adjusted to pH 7.6, and heated in a boiling water bath for 20 min. The conjugated oligomer was then repurified over P4 for storage at refrigerator temperatures. Labeling is achieved simply by adding fresh stannous ion to a solution of the MORF-MAG(3) in pH 7.6 containing tartrate followed by (99m)Tc pertechnetate. After the mixture is heated for 20 min in boiling water, the labeling efficiency is always over 90% as determined by size exclusion HPLC and paper chromatography and the specific activities can exceed 7 mCi/microg. By making several relatively simple changes to the routine procedure used to conjugate and radiolabel biomolecules with (99m)Tc via MAG(3), a modified procedure was developed that results in labeling efficiency high enough to avoid postlabeling purification. PMID- 12121148 TI - Visible light induced biohydrogen production from sucrose using the photosensitization of Mg chlorophyll-a. AB - A photoinduced hydrogen production system, coupling sucrose degradation with invertase and glucose dehydrogenase (GDH) and hydrogen production with colloidal platinum as a catalyst using the visible light-induced photosensitization of Mg chlorophyll-a (Mg Chl-a), has been developed. Continuous hydrogen gas production was observed when the reaction mixture containing sucrose, invertase, GDH, nicotinamide adenine dinucleotide (NAD(+)), Mg Chl-a, methyl viologen (MV(2+), an electron relay reagent), and colloidal platinum was irradiated by visible light. PMID- 12121149 TI - Comparison of yttrium and indium complexes of DOTA-BA and DOTA-MBA: models for (90)Y- and (111)In-labeled DOTA-biomolecule conjugates. AB - Yttrium and indium complexes of 1,4,7,10-tetraaza-4,7,10-tris(carboxymethyl)-1 cyclododecylacetylbenzylamine (DOTA-BA) and 1,4,7,10-tetraaza-4,7,10 tris(carboxymethyl)-1-cyclododecylacetyl-R-(+)-alpha-methylbenzylamine (DOTA-MBA) were prepared in order to study solution structures of (90)Y- and (111)In-labeled DOTA-biomolecule conjugates. (90)Y and (111)In complexes M(L) (M = (90)Y and (111)In; L = DOTA-BA and DOTA-MBA) were prepared from the reaction of MCl(3) with DOTA-BA and DOTA-MBA, respectively, in ammonium acetate buffer. A reverse phase HPLC method revealed that both (90)Y and (111)In complexes show only one radiometric peak in their radio-HPLC chromatograms. It was also found that (111)In(DOTA-BA) and (111)In(DOTA-MBA) are more hydrophilic than their corresponding (90)Y analogues, suggesting different coordination spheres in (111)In and (90)Y complexes of the same DOTA conjugate. Complexes M(L) (M = Y and In; L = DOTA-BA and DOTA-MBA) were prepared and characterized by HPLC, LC-MS, and NMR ((1)H and (13)C) methods. The HPLC concordance experiments for (90)Y(DOTA MBA)/Y(DOTA-MBA) and (111)In(DOTA-MBA)/In(DOTA-MBA) show that the same complex is prepared at both tracer and macroscopic levels. The NMR data ((1)H and (13)C) clearly demonstrates that Y(DOTA-BA) and Y(DOTA-MBA) exist in solution as one predominant isomer. VT NMR data ((1)H and (13)C) show that In(DOTA-BA) and In(DOTA-MBA) are fluxional at room temperature while Y(DOTA-BA) and Y(DOTA-MBA) become fluxional only at elevated temperatures. The fluxionality of these complexes is due to rapid rotation of acetate/acetamide chelating arms and inversion of ethylenic groups of the macrocyclic ring. PMID- 12121151 TI - Not another 'NOF'! PMID- 12121152 TI - Abdominal aortic aneurysms: which surgeon and what procedure? PMID- 12121150 TI - Inhibition of telomerase by linear-chain fatty acids: a structural analysis. AB - In the present study, we have found that mono-unsaturated linear-chain fatty acids in the cis configuration with C(18) hydrocarbon chains (i.e. oleic acid) strongly inhibited the activity of human telomerase in a cell-free enzymic assay, with an IC(50) value of 8.6 microM. Interestingly, fatty acids with hydrocarbon chain lengths below 16 or above 20 carbons substantially decreased the potency of inhibition of telomerase. Moreover, the cis-mono-unsaturated C(18) linear-chain fatty acid oleic acid was the strongest inhibitor of all the fatty acids tested. A kinetic study revealed that oleic acid competitively inhibited the activity of telomerase ( K (i)=3.06 microM) with respect to the telomerase substrate primer. The energy-minimized three-dimensional structure of the linear-chain fatty acid was calculated and modelled. A molecule width of 11.53-14.26 A (where 1 A=0.1 nm) in the C(16) to C(20) fatty acid structure was suggested to be important for telomerase inhibition. The three-dimensional structure of the telomerase active site (i.e. the substrate primer-binding site) appears to have a pocket that could bind oleic acid, with the pocket being 8.50 A long and 12.80 A wide. PMID- 12121153 TI - Treatment of acute malignant colorectal obstruction with self-expandable metallic stents. AB - BACKGROUND: Malignant left-sided large bowel obstruction from intraluminal and extrinsic causes constitutes a surgical emergency. When conservative measures fail, emergent surgery is usually required, which carries increased morbidity and mortality compared with elective resections. In many situations, a stoma is created and further surgery may be required later to re-establish bowel continuity. We present an initial series of patients in whom self-expandable metallic stents (SEMS) were deployed to allow bowel decompression in place of emergency surgery for acute left-sided colorectal obstruction. METHODS: From April 1999 to January 2001, patients who were admitted to the colorectal unit with clinical and radiological features of intestinal obstruction were considered for endolumenal stenting. Stenting was performed under radiological guidance. RESULTS: Sixteen patients underwent endolumenal stenting (age range: 23-90 years, eight men and eight women). There were two technical failures, two delayed perforations and one sealed perforation related to the stent. Three patients underwent elective resection and anastomosis after successful bowel decompression and mechanical bowel preparation. Eight patients with advanced malignancy or multiple medical disease had good bowel decompression after stent deployment and did not undergo any surgery. CONCLUSION: SEMS is a useful alternative in the management of acute left-sided colorectal obstruction for a select group of patients. An algorithm for management of acute left-sided large bowel obstruction incorporating the use of SEMS is proposed. PMID- 12121154 TI - Effect of cervical hard collar on intracranial pressure after head injury. AB - BACKGROUND: Patients suffering head trauma are at high risk of having a concomitant cervical spine injury. A rigid cervical collar is usually applied to each patient until spinal stability is confirmed. Hard collars potentially cause venous outflow obstruction and are a nociceptive stimulus, which might elevate intracranial pressure (ICP). This study tested the hypothesis that application of a hard collar is associated with an increase in ICP. METHODS: A prospective series of 10 head-injured patients with a postresuscitation Glasgow coma scale score of nine or less had ICP measurements before and after cervical hard collar application. RESULTS: Nine out of 10 patients had a rise in ICP following application of the collar. The difference in pre- and postapplication ICP was statistically significant (P < 0.05). CONCLUSIONS: Early assessment of the cervical spine in head-injured patients is recommended to minimize the risk of intracranial hypertension related to prolonged cervical spine immobilization with a hard collar. PMID- 12121155 TI - Stenting for obstructing colorectal malignancy: an interim or definitive procedure. AB - BACKGROUND: The purpose of this paper is to review and report our experience with colorectal stenting in the management of malignant large bowel obstruction. METHODS: Twelve consecutive patients with malignant left-sided large bowel obstruction between June 1998 and January 2001 underwent insertion of self expanding metallic stents. One patient required two stents. Eight stents were inserted under fluoroscopic guidance, and five were inserted with combined fluoroscopic and endoscopic guidance. Patients were followed up until death, stent removal or the time of publication. RESULTS: Thirteen stents were inserted. Eleven patients with acute large bowel obstruction had relief of obstruction with stenting, and one of these patients required a second stent because relief had been incomplete. One patient was stented in order to subsequently close a problematic stoma. Technical success was 92.9% and clinical success was 100%. Three patients proceeded to surgical resection. In nine patients, the stent was left as the definitive procedure. Of these, six patients have died within 4 months. Complications included one case of migration, one case of reobstruction, one intestinal haematoma and one case of cheesewiring. CONCLUSIONS: Colorectal stenting is an important treatment modality for malignant large bowel obstruction. It may be definitive treatment alone, or a bridge to elective surgical resection. PMID- 12121156 TI - Tube sigmoidostomy: a modification of the antegrade colonic evacuation. AB - BACKGROUND: The Malone appendicostomy enema has established its place in the treatment of paediatric faecal incontinence. More recently, the laparoscopic appendicostomy enema has gained favour. In the situation where the appendix has already been removed, caecal stomal tubes can be constructed to facilitate antegrade colonic wash-outs. These techniques may not provide an accept-able result if there is a significant time delay from the introduction of the enema to the result. Wash-outs through the more distal colon -- either by retrograde techniques with a rectal tube or, more recently, through a plastic stomal device inserted percutaneously into the sigmoid colon (the Chait tube) -- can result in rapid evacuation. Rectal wash-outs may not provide an adequate lavage and can be technically difficult for some children to perform, while many other children find external devices cosmetically unacceptable. We present a modification of this distal colonic technique that can allow greater control, can be used in the absence of an appendix and avoids the need for an external device. METHODS: Two paediatric patients with spina bifida and faecal incontinence underwent tube sigmoidostomy formation. In both patients, the appendix was not available to be used for colonic wash-outs. RESULTS: Both patients reported excellent success with this procedure. They are continent, able to cannulate the stoma and irrigate independently. CONCLUSIONS: The technique can provide excellent continence control in patients who are not successful with a right-sided Malone appendicostomy. It also overcomes the need for a catheterizable appendix, and an external device. PMID- 12121157 TI - Management of liver trauma with implications for the rural surgeon. AB - BACKGROUND: The majority of patients with liver trauma can be managed conservatively. However, the unstable patient requires emergency laparotomy to control bleeding. Controversy exists regarding the primary surgical management of these injuries. This is of particular relevance for the isolated rural general surgeon. METHODS: The literature was reviewed by searching MEDLINE databases from 1966 to the present time. The majority of the evidence presented is level 3, with interpretations and recommendations based on the experience of the senior authors. RESULTS: In the majority of patients, conservative management remains the mainstay of treatment. However, haemodynamic -instability requires urgent laparotomy. Perihepatic packing should be used to arrest bleeding. Primary anatomical resection is rarely indicated, especially in non-specialist centres. CONCLUSION: In the remote rural setting, severe liver trauma remains a daunting condition for the general surgeon to manage. Primary surgical treatment should be perihepatic packing, stabilization and urgent transfer; there is no place for primary anatomical resection outside specialist units. PMID- 12121158 TI - Influence of preoperative medical status and delay to surgery on death following a hip fracture. AB - BACKGROUND: A retrospective study of 138 patients with fractured hips was undertaken to determine if a delay to surgery beyond 24 h influenced 1-year mortality. In particular, the results of the subgroup of patients who were in the American Society of Anesthesiologists (ASA) Class 3 or 4 were considered. METHODS: Patients were found using ICD-9 database information. One-year mortality data was collected from Births, Deaths and Marriages -- the New Zealand government agency that collects and stores statistics on these events. RESULTS: The 1-year mortality rate was 17.4%. Age, operation type and time to surgery did not significantly affect 1-year mortality. American Society of Anesthesiologists' Class 1 and 2 patients had a significantly lower 1-year mortality (5.3%) than ASA Class 3 and 4 patients (22.4%) (P = 0.02). CONCLUSION: Time to surgery did not significantly affect 1-year mortality within each ASA Class. PMID- 12121159 TI - Primary melioidotic prostatic abscess: presentation, diagnosis and management. AB - INTRODUCTION: In South-East Asia and Northern Australia, melioidosis (infection with Burkholderia pseudomallei) is a known cause of severe community-acquired sepsis. However, melioidosis presenting primarily as prostatic abscesses is very rare. METHODS: The presenting features, investigations and management outcome of five patients who developed melioidotic prostatic abscesses from 1997 to 2000 were reviewed in the present study. RESULTS: The mean age at presentation was 53 years (range: 29-69). Old age and diabetes mellitus were predisposing factors. All patients had a fever of at least 38.5 degrees C and presented with obstructive urinary symptoms culminating in urinary retention. Presence of prostatic abscess was demonstrated by transrectal ultrasound in all cases. The abscesses were drained with transurethral resection of the prostate. One patient required re-resection while another patient developed severe septic shock requiring intensive care and -inotropic support. There was no mortality in our series. CONCLUSIONS: Elderly diabetic men presenting with fever and urinary tract obstruction in endemic areas may harbour an unusual but potentially life threatening melioidotic prostatic abscess. Transrectal ultrasound and bacteriological confirmation are mandatory. Prompt surgical drainage coupled with appropriate antibiotics are keys to a favourable outcome. PMID- 12121160 TI - Impact of splenectomy on circulating T-lymphocyte subsets in stage III gastric cancer. AB - BACKGROUND: The role of splenectomy remains unclear in patients with gastric cancer who undergo total gastrectomy. The aim of this study was to prospectively evaluate the impact of splenectomy on circulating T-lymphocyte subsets and survival in advanced gastric cancer. METHODS: Analysis of lymphocyte subsets was performed in 40 patients with American Joint Committee on Cancer (AJCC) stage III gastric adenocarcinoma located on the upper one-third of the stomach, who underwent a curative total gastrectomy with or without splenectomy. Circulating T lymphocyte subsets were measured on venous blood by using flow cytometry and monoclonal antibodies at preoperative day 1, and postoperative months 1, 3, 6, 12 and 18. RESULTS: The proportion of lymphocytes and the values of CD3, CD8, CD16 and CD25 subsets were higher in the splenectomy group of patients at postoperative month 3. In the spleen preservation group at the same point of treatment, the proportion of granulocytes and the values of CD4 and CD4 : CD8 ratio were higher. Except for CD16 levels, all T-lymphocyte subsets showed no significant difference between splenectomy and spleen preservation groups after postoperative month 3. Increased CD16 levels in the splenectomy group were not associated with improvement in patients' 5-year survival rates. CONCLUSION: These results suggest that the long-term impact of splenectomy does not play an important role in postoperative quantitative changes of circulating T-lymphocyte subsets of patients with stage III gastric cancer who have undergone total gastrectomy. Furthermore, splenectomy does not give a prognostic benefit, based on tumour recurrence and survival of patients with proximal one-third gastric cancer who undergo total gastrectomy. PMID- 12121161 TI - Retropharyngeal abscess in children. AB - BACKGROUND: Retropharyngeal abscess (RPA) is an uncommon, potentially fatal condition found more frequently in children than adults. Prompt diagnosis and surgical management of this condition is imperative to prevent complications including airway obstruction and mediastinitis. Few studies have been dedicated to paediatric retropharyngeal abscess. METHODS: A retrospective analysis of 21 cases of retropharyngeal abscess at the Sydney Children's Hospital over a 12-year period was performed. RESULTS: There were 12 boys and nine girls involved in the analysis. Their ages ranged from 3 months to 12 years. Common -presenting symptoms and signs included fever, dysphagia, neck swelling and torticollis. Respiratory compromise was present in 29% of the children at presentation. Foreign body ingestion accounted for 10% of cases. Seventeen cases were managed with surgical drainage. Surgical approaches adopted included transoral (70%), external cervical approach (20%) and a combined approach in 10%. There was no mortality. Mediastinitis occurred in two patients, one of whom also had recurrent laryngeal nerve palsy. No other serious complications occurred. CONCLUSION: Retropharyngeal abscess should be considered in all children presenting with neck pain and dysphagia. Prompt diagnosis and institution of appropriate medical and surgical therapy is imperative to prevent complications such as airway obstruction. The management of this condition should occur in a paediatric institution with appropriate medical, surgical and intensive care -facilities. PMID- 12121162 TI - POSSUM: a re-evaluation in patients undergoing surgery for rectal cancer. The Physiological and Operative Severity Score for Enumeration of Mortality and Morbidity. AB - BACKGROUND: The problem of directly comparing morbidity and mortality rates between institutions without some sort of adjustment for case mix is well documented. Scoring systems have been developed to allow comparisons to be made. The Physiological and Operative Severity Score for the enUmeration of Mortality and morbidity (POSSUM) is one such system. It was designed to predict operative mortality and morbidity in differing settings and to be independent of case mix. The present study examines the use of POSSUM in colorectal practice in Saudi Arabia. METHODS: Patients referred to King Faisal Specialist Hospital between 1990 and 1998 for primary management of an histologically proven rectal cancer were identified. POSSUM mortality and morbidity scores and Portsmouth Physiological and Operative Severity Score (P-POSSUM) mortality scores were calculated separately for each patient, and predicted rates were compared with observed rates in the patients studied. RESULTS: There were 70 men (mean age: 55.6 years; range: 25-87) and 75 women (mean age: 52.8 years; range: 26-84). One hundred and six patients underwent 'curative' surgery. Abdominoperineal resection was the most frequently performed procedure. Major anastomotic leakage following anterior resection occurred in two of fifty patients. One patient developed a pulmonary embolism but no patient developed postoperative myocardial infarction. Two patients died. The median and mean physiological and operative severity scores were 13 (range: 12-37) and 17 (range: 8-37) and 14.68 and 18.36, respectively. The overall POSSUM-predicted (using median scores) morbidity and mortality rates were 35.4% and 6.7%. The P-POSSUM-predicted (using mean scores) mortality rate was 3.5%. Observed morbidity and mortality rates were 54.5% and 1.4%. CONCLUSION: POSSUM failed to predict outcomes accurately in patients undergoing surgery for rectal cancer in Saudi Arabia. P-POSSUM also overpredicted mortality but to a lesser extent. Patient's 'wellness' and the previously identified inability of POSSUM to accurately predict death in low-risk populations may explain these findings. Care must be exercised in using the POSSUM formulae for risk adjustment in different settings. PMID- 12121163 TI - Intraoperative magnetic resonance: the future of surgery. AB - Intraoperative magnetic resonance imaging (iMRI) is a new development in medicine that bridges the specialties of surgery and radiology. Deficiencies in the visualization of anatomical architecture and the perception of tumour boundaries in conventional open surgery have led to the integration of imaging within surgery. The superior soft tissue and multiplanar imaging features of magnetic resonance (MR) make this imaging modality superior to that of alternatives. The unique properties of MR to detect heat change and perfusion, and diffusion characteristics of tissue enhance the usefulness of this medium. Concurrent developments in computer aided image guidance and thermoablative technology, herald the era of minimally invasive tumour ablation. Applications have been developed for areas such as neurosurgery, general surgery, gynaecology and urology. PMID- 12121164 TI - Survey of current cementing techniques in total knee replacement. AB - BACKGROUND: Current techniques in total knee replacement (TKR) vary greatly. Cementing techniques in TKR are no exception. Identification of current trends would aid ongoing research into future developments. METHODS: A questionnaire was sent to all orthopaedic surgeons in Queensland, Australia. A range of factors influencing cementing techniques for the tibial plateau were investigated. RESULTS: Eighty-five percent of questionnaires were returned. Ninety-one percent of surgeons regularly cement the tibial plateau, 90% use some form of lavage prior to cement application, 95% of surgeons apply cement by hand only. Techniques aimed at increasing cement penetration (cement gun, intraosseous suction) are used infrequently. CONCLUSIONS: In contrast to total hip replacement, measures are not often taken to ensure that there is penetration of cement or a complete cement mantle into the tibial plateau in TKR. This may be because of a perceived lack of evidence in modern literature. This study also illustrates the wide variation in technique in a relatively common orthopaedic operation. PMID- 12121165 TI - The Mann-Williamson procedure: a historical contribution. PMID- 12121166 TI - Primary hydatid cysts of psoas muscle. AB - BACKGROUND: Hydatid cysts may occur in any area of the body, but they usually localize to the liver and the lungs. Primary localization in muscle is not common, accounting for 2-3% of all sites; even rarer is the development of multiple cysts. METHODS: The patient presented with a painless abdominal mass which gradually increased in size to a diameter of approximately 16 cm. Organ imaging scan revealed multiple hydatid cysts within the right psoas muscle. Because of the proximity of the lesions to the iliac vessels, ureter and nerves to the lower limb, percutaneous drainage and alcoholization under local anaesthesia were -performed with the aim of reducing the size of the cysts and sterilizing them prior to definitive surgery. This procedure was not effective. Two weeks after percutaneous treatment the patient underwent surgery. RESULTS: At operation the cysts were localized and successfully removed under ultrasound guidance. Postoperative stay was -uneventful. Two years after surgery the patient has no evidence of recurrent hydatid disease. CONCLUSIONS: Ultrasonography is the preferred method for detecting muscular hydatid cyst and for guiding the surgeon during resection. PMID- 12121167 TI - Isolated dissection of a non-aneurysmal popliteal artery. AB - Two cases of isolated dissection of a non-aneurysmal, non-atherosclerotic popliteal artery are reported. Isolated dissection of the popliteal artery is a distinct entity that may have far reaching effects on patient's mobility if not adequately treated. This lesion can be seen in non-aneurysmal, non atherosclerotic vessels and in patients with no predisposing medical causes. It requires a high degree of suspicion for diagnosis because of its rarity. Surgical treatment is the best option and results in a good outcome. PMID- 12121169 TI - Autoimmune pancreatitis. AB - The recommended treatment for a focal mass in the head of the pancreas is pancreaticoduodenectomy. Preoperative biopsy is not advised in patients who are candidates for resection because of the documented risk of tumour dissemination along the needle tract and significant false negative results.1 Autoimmune pancreatitis is a relatively uncommon condition that can present as a pancreatic mass and mimic malignancy. It may respond to glucocorticoid therapy, and further assessment of such treatment is indicated.2 Such experience will only accumulate if wider knowledge of this condition leads to clinical suspicion. PMID- 12121168 TI - Chicken bone perforation of a sigmoid diverticulum. PMID- 12121170 TI - Initial experience of vocal cord evaluation using grey-scale, real-time, B-mode ultrasound. PMID- 12121180 TI - 'Complementary and alternative medicine' population based studies: a growing focus on allergy and asthma. PMID- 12121181 TI - Was the lung as target organ in food allergy underestimated? PMID- 12121182 TI - Allergy as a global problem: 'think globally, act globally'. PMID- 12121183 TI - Asthma and allergy: a worldwide problem of meanings and management? PMID- 12121184 TI - Intrinsic properties of allergens and environmental exposure as determinants of allergenicity. PMID- 12121185 TI - Asthma and atopy - a total genome scan for susceptibility genes. AB - BACKGROUND: Allergic asthma is an increasingly common disease of complex inheritance. Several studies have suggested candidate regions, but genetic heterogeneity, ethnic differences and varying study designs may in part explain the lack of identified and confirmed susceptibility genes. Investigation of different populations will further clarify the topic. We therefore evaluated allergic asthma and increased total and specific IgE in 39, 45 and 57 sib-pairs from 100 Danish allergy families. METHODS: Affected sib-pairs meeting a narrow phenotype definition were selected for the three phenotypes atopy, allergic asthma and increased total IgE. We performed a total genome scan using 446 microsatellite markers and obtained nonparametric linkage results from the MAPMAKER/SIBS computer program. RESULTS: Our study revealed four candidate regions (MLS > 2) on chromosome 1p36, 3q21-q22, 5q31 and 6p24-p22, and 15 candidate regions (1 < MLS < 2) that may contain susceptibility genes for asthma and atopy. We did not find linkage to the candidate genes TNF-beta, FcER1beta and Il4R-alpha, except for weak support for linkage of the asthma phenotype to TNF beta (MLS = 1.18). CONCLUSIONS: We found evidence for two new asthma and atopy loci, 1p36 and 3q21-q22, and supported linkage in the Danish population to seven previously reported candidate regions. PMID- 12121186 TI - Determinants of undiagnosed asthma. AB - BACKGROUND: The conformity between asthma diagnosed in a clinical setting and asthma from an epidemiological point of view has not been much studied. METHODS: History, symptoms, and clinical findings in persons with and without a previous physician diagnosed asthma were studied among persons classified as having asthma according to an epidemiological definition. RESULTS: Smoking was more prevalent in persons not diagnosed as having asthma prior to the study (n = 96), than in those with a previous asthma diagnosis (n = 119), (60% vs. 41%, P = 0.006). Positive skin prick test (SPT) was less common (39.6% vs. 58.0%, P = 0.009), lung function expressed as FEV1.0 in percentage of predicted was better (92.1 vs. 85.4, P = 0.018), reversibility to a bronchodilator was smaller (mean reversibility 4.0% vs. 6.2% of predicted, P = 0.003) and symptoms were less frequently reported (P < 0.001) by those who had not a physician diagnosed asthma prior to the study. CONCLUSION: The asthma diagnosis is often overlooked by the medical profession in patients with mild disease. Furthermore, smoking seems to worsen asthma exacerbations in patients with mild disease. PMID- 12121187 TI - Alternative medicine in allergies - prevalence, patterns of use, and costs. AB - BACKGROUND: There is evidence that the use of alternative medicine (AM) for allergies has increased. However, little is known from population-based studies about what determines its use. The objective of this study was to evaluate the patterns of use of AM for allergies. METHODS: A population-based nested case control study was conducted in 2000-01 using computer-assisted telephone interviews. Three hundred and fifty-one adults participated (median age 46 years) with allergies including hay fever, asthma, atopic eczema, and food hypersensitivity. Information was obtained on demographics, prevalence, motivation, information, type of AM, provider, costs, willingness to pay, and subjective assessment of AM. RESULTS: 26.5% of participants used AM because of their allergies. Compared to nonusers, this group of users was significantly younger (median age 43 vs 47; p=0.004) and better educated (school education > 8 year vs75% and >95% of benign glands (verified on H&E). Cases that showed no staining were repeated to ensure no false negatives. Both observers agreed with respect to percentage of staining in 96% of the cases. Twenty-nine of 30 cases (97%) showed staining in >95% of benign glands with CK5/6. In contrast, K903 staining was seen in <50% of benign glands in five of 30 (17%), 50-75% in nine of 30 (30%), and >75% in 10 of 30 (33%), with only two cases (7%) showing >95% staining for K903. In four cases (13%) the K903 failed to stain any tissue even after repeat staining. K903 was conspicuously negative in atrophic glands in three of 30 cases (10%). Neither K903 nor CK5/6 stained malignant glands. Using a cut-off of >75% staining in benign glands the sensitivity of CK5/6 and K903 was 97% and 40%, respectively. CONCLUSIONS: CK5/6 has superior sensitivity and reliability compared with that of K903 when evaluating routine prostate needle biopsies, including improved staining of atrophic prostatic glands. While K903 is traditionally used to differentiate benign glands from malignant glands, these results support the use of CK5/6 as an effective and reliable substitute for K903 in routine clinical practice. PMID- 12121236 TI - Primary thymic epithelial tumours of the pleura mimicking malignant mesothelioma. AB - AIMS: To illustrate the macroscopic, light microscopic and immunophenotypic similarities that exist between primary pleural thymic epithelial tumours and diffuse malignant mesothelioma. To investigate the expression of the mesothelial markers, cytokeratin (CK) 5/6, calretinin and thrombomodulin in a series of mediastinal thymic epithelial tumours. METHODS AND RESULTS: Over a 10-year period, 64 diffuse pleural tumours of non-mesothelial histogenesis were identified in the files of referrals to the South Wales regional thoracic centre (Llandough Hospital, Cardiff). Of these, five pleural tumours were diagnosed as primary pleural thymic epithelial neoplasms. From the files of the Mesopath group, Caen, three additional cases of thymic epithelial tumours with pleural involvement were identified. The study group comprised eight cases (four males, four females) with median age at presentation of 56 years (range 19-75 years). In one case there was a history of asbestos exposure. Macroscopically, seven tumours formed diffuse pleural masses. No mediastinal abnormality or intraparenchymal lesions were seen in five cases. By light microscopy, seven thymic epithelial neoplasms showed a lobulated architecture, one appeared extensively cystic. The tumours were of varied morphological subtypes: one medullary (WHO Type A), two mixed (WHO Type AB), three predominantly cortical (WHO Type B1) and two cortical (WHO Type B1). The subtypes morphologically mimicked sarcomatoid, biphasic, lymphohistiocytoid variant and epithelioid mesothelioma. The pleural thymic epithelial tumours showed immunoreactivity with broad spectrum cytokeratin AE1/AE3 (8/8; 100%), CK5/6 (8/8; 100%), and 1/8 (13%) expressed thrombomodulin. Calretinin showed variable nuclear and cytoplasmic expression in all cases, but equivocally in the thymic epithelial cell component. In 7/8 (88%) the thymic epithelial cells exhibited focal aberrant expression of CD20. Epithelial membrane antigen (EMA) showed focal expression in the perivascular and organoid areas in 6/8 (75%) cases. Carcinoembryonic antigen (CEA) and CD34 were uniformly negative. In 4/8 (50%) cases the lymphoid cell component was of immature phenotype expressing CD99, terminal deoxynucleotidyl transferase (TdT) and lymphoid precursors had a high proliferation fraction with Ki67. In the series of 20 primary mediastinal thymic epithelial tumours tested, mesothelial marker expression revealed CK5/6 (20/20), thrombomodulin (3/20; 15%) and calretinin (0/20; 0%). Varying amounts of calretinin-positive stromal cells were present. CONCLUSION: Primary pleural thymic epithelial tumours are rare but may mimic malignant mesothelioma by forming diffuse serosal-based masses. In addition, both tumours may show morphological diversity (with epithelial, spindled and mixed components present). An awareness that thymic epithelial tumours may variably express the mesothelial markers CK5/6, calretinin and thrombomodulin prevents misdiagnosis. In the distinction from malignant mesothelioma a lobulated architecture and organoid features favour a thymic epithelial neoplasm. The presence of aberrant CD20 expression in a cytokeratin-positive epithelial neoplasm and/or the presence of an immature lymphoid population (by demonstration of CD1a, CD2, CD99 and TdT) indicates a thymic epithelial neoplasm. In contrast, nuclear calretinin expression favours malignant mesothelioma. PMID- 12121237 TI - Granulomatous ulcerative colitis: a re-appraisal of the mucosal granuloma in the distinction of Crohn's disease from ulcerative colitis. AB - AIMS: To determine whether the presence and location of giant cells or granulomas in relation to crypts distinguishes between ulcerative colitis and Crohn's disease. METHODS AND RESULTS: Twenty-nine large bowel mucosal biopsy specimens showing giant cells and/or granulomas in a background more typical of ulcerative colitis than Crohn's disease were collected between 1986 and 1996. Each was subject to detailed independent analysis by three histopathologists. Follow-up of the cases was by examination of all previous and subsequent gastrointestinal surgical or biopsy material and by scrutiny of the clinical notes by a gastroenterologist. On the basis of the accumulated histological data 10 of these 29 cases were accorded the diagnosis of ulcerative colitis. In nine of these 10 cases the clinical diagnosis, where known, was in keeping with this and all nine contained only crypt-associated giant cells and/or granulomas. The tenth case contained a solitary free-standing granuloma and clinically the patient had perianal disease, suggesting that the true diagnosis was Crohn's disease. CONCLUSIONS: Isolated giant cells and well-defined epithelioid granulomas distant from crypts do not, as a rule, occur in ulcerative colitis, and hence their presence in a colonoscopic biopsy showing features of chronic inflammatory bowel disease is a strong pointer towards the diagnosis of Crohn's disease. Crypt associated giant cells and granulomas can occur in ulcerative colitis and in themselves are unreliable features for the discrimination between Crohn's disease and ulcerative colitis. PMID- 12121238 TI - Differences in p53 and cadherin-catenin complex expression between histological subtypes in diffusely infiltrating gastric carcinoma. AB - AIMS: The aim of this study was to elucidate possible clinicopathological differences between diffusely infiltrating gastric carcinoma of 'pure type' (poorly differentiated carcinoma without any glandular component) and 'mixed type' (coexistence of poorly differentiated carcinoma and intramucosal glandular component). METHODS AND RESULTS: The clinicopathological features and immunohistochemical expression of p53 and intercellular adhesion molecules (E cadherin and alpha-, beta- and gamma-catenins) were compared between the patients with pure (n=59) and mixed (n=56) types of diffusely infiltrating gastric carcinoma. Intestinal metaplasia (P < 0.01), prominent lymphatic permeation (P < 0.001) and lymph node metastasis (P < 0.05) were more frequently observed in mixed type than in pure type, while survival probability did not differ between the two groups. The frequency of p53 over-expression was higher in mixed type (56%) than in pure-type (19%) (P < 0.001). In mixed type, p53 expression was not different between glandular and poorly differentiated components. By contrast, the expression of adhesion molecules was more frequently preserved in glandular components than in poorly differentiated components. CONCLUSIONS: These two subtypes seem to be different in nature and biological behaviour. The preservation of adhesion molecules in mixed type may be associated with higher incidence of lymphatic permeation and lymph node metastasis. PMID- 12121239 TI - Expression of multidrug resistance-associated protein 2 (MRP2) in normal human tissues and carcinomas using tissue microarrays. AB - AIMS: Using tissue microarrays, this study analysed the expression of the multidrug resistance protein, MRP2, by immunohistochemistry with two different MRP2 antibodies. This is the first study to address the expression of MRP2 in various common human neoplasms. METHODS AND RESULTS: Immunohistochemistry was performed on zinc formalin-fixed tissue to evaluate normal tissues and carcinomas using two antibodies against MRP2 (EAG5, a polyclonal antibody, and M2-lll-6, a monoclonal antibody). Immunostaining was localized in neoplastic cells mainly on the cell membrane with M2-lll-6 and cell membrane and cytoplasm with EAG5. In normal tissues MRP2 was expressed in liver, gastrointestinal tract, and kidney tubular epithelial cells with both antibodies. MRP2 was seen in nine of 22 renal cell carcinomas, eight of 13 gastric carcinomas, 25 of 49 breast carcinomas, 14 of 32 lung carcinomas, 39 of 50 colon carcinomas, and 16 of 17 ovarian carcinomas. There was < 10% variability between the two antibodies. MRP2 expression was highest in moderate to poorly differentiated tumours from colon, lung, gastric, and ovarian carcinomas and in grade 2 and 3 breast and renal carcinomas. CONCLUSION: The expression of MRP2 in many solid human tumours indicates that inherent drug resistance may play an important role as a biomarker for predictive chemotherapy treatment. PMID- 12121240 TI - A method to fix lipids for staining fat embolism in paraffin sections. AB - AIMS: To develop a method to preserve lipids in formalin-fixed tissues for staining in paraffin sections, and to illustrate its use in lung and brain of a fat embolism case, and in examples of fatty liver and atheroma. METHODS AND RESULTS: A saturated solution of linoleic acid in 70% ethylene glycol was prepared and tissues were exposed to this for 3 days at 56 degrees C. These tissues were treated with 2% chromic acid at 4 degrees C for 24 h followed by 24 h in 5% sodium bicarbonate, with appropriate rinsing between solutions. Paraffin sections of these tissues were stained with a lipid-soluble dye such as Oil Red O. Examples of fat embolism, fatty liver, and atheroma were shown photographically as illustrations of expected results. CONCLUSIONS: The demonstration of fat embolism with good quality tissue detail is made practical by the method, which is convenient and inexpensive. The method appears to be generally applicable to tissue lipids of various sorts, as exemplified by adipose tissue, fatty liver, and atheroma. PMID- 12121242 TI - Localized pleural metastatic adenosarcoma of the uterine cervix mimicking a malignant solitary fibrous tumour: CD10 has no value in differential diagnosis. PMID- 12121243 TI - Intraductal papilloma of the parotid gland in a child. PMID- 12121244 TI - Cutaneous cellular fibrous histiocytoma metastasizing to the lungs. PMID- 12121245 TI - Cytological atypia in endometrial polyps and immunostaining for p16, p53 and Ki67. PMID- 12121247 TI - Guest editorial: mental health then and now: the past and future contribution of Health and Social Care in the Community to policy and practice. PMID- 12121248 TI - Street-level bureaucrats? heart disease, health economics and policy in a primary care group. PMID- 12121249 TI - Experience and meaning of user involvement: some explorations from a community mental health project. AB - With an increased interest in and policy commitment to involving service users in the planning and delivery of health service provision, there is a clear need to explore both the rhetoric and realities of what user involvement entails. In the present paper, by drawing upon an evaluation of a community-based exercise facility for people with mental health problems, the authors explore ways in which the reality of user involvement is subject to a range of configurations within health services. The paper describes a piece of qualitative research that was undertaken within a participatory framework to explore the nature of user involvement within the facility. The data have been analysed using a grounded theory approach to provide insights into: the organisational context in which user involvement takes place; factors which encourage meaningful participation on the part of service users; perceived barriers to user involvement; and issues of sustainability and continuity. This research approach has enabled the authors to explore the views and experiences of users, service providers and referral agencies in relation to the nature and potential for user involvement. The findings illustrate ways in which user involvement may take place under both flexible and formal arrangements across a variety of activities. The present paper provides an account of some of the meanings and experiences of what 'successful' user participation may involve and the conditions which underpin 'success'. The authors conclude that successful and meaningful user involvement should enable and support users to recognise their existing skills, and to develop new ones, at a pace that suits their particular circumstances and personal resources. This process may require adaptation not only by organisations, but also by service providers and non-involved users. PMID- 12121250 TI - Comparing and contrasting the role of family carers and nurses in the domestic health care of frail older people. AB - Care in the community has been constructed on the basis of professional support for carers who, as a result of community care policy that has released highly dependant people from residential care and long-stay wards, are carrying out a wide range of tasks, including complex health care activities. The present paper examines the health care activities currently undertaken by family carers and the way in which they work with, and are supported by, professional nurses in the home. It compares and contrasts the approaches of both groups to care-giving for this client group. The authors conclude by making some suggestions for improving the way in which family carers and nurses work together in the home. PMID- 12121251 TI - Combining informal care and work: supporting carers in the workplace. AB - The UK Government is concerned that women and men who care for disabled or sick relatives, or elderly people, and who also wish to take part in paid work should have increased opportunities to do so. However, many informal carers find combining work and care difficult; some may 'choose' to give up paid employment completely. The present paper draws on the findings from two projects to explore the extent to which the needs of employees with caring responsibilities are supported in the workplace. The two projects examined evidence from a study of informal carers assessed under the 1995 Carers Act, identified the difficulties which they face in their workplace and observed the strategies which they developed to help sustain the two roles. From this, a model of support for working carers was developed which includes leave policies, carer-friendly working arrangements, access to a (private) telephone, and supportive line managers and co-workers. This support model was tested on the employment policies of 13 employers to see how 'carer-friendly' they were. Most of the organisations studied were able to provide appropriate support for carers identified in the model. Questions were then raised about different aspects of carer-friendly working arrangements, including whether carers should receive any special treatment that is not available to their colleagues, the role of line managers, and the relationship between seniority and opportunities to combine work and care. PMID- 12121252 TI - Solving inequities in provider distribution: loan repayment. AB - The distribution of primary health care professionals in England and Wales is inequitable, with relatively lower concentrations of professionals in deprived areas. The objective of the present study was to determine whether graduate health professionals would be willing to work in under-served areas in return for educational loan repayment. The study group consisted of a convenience sample of 50 newly qualified and trainee general practitioners, and 50 newly qualified community nurses and health visitors in mid- and west Wales. At interview, the subjects were presented with descriptions of general practices and asked to indicate their preferred practice. Practice descriptions varied systematically in terms of location (i.e. urban, suburban and rural), population deprivation (i.e. deprived or mixed affluent/deprived) and availability of loan repayment (i.e. none or loans paid off over a period of between one and 4 years). The main outcome was the probability that a practice with loan repayment was chosen. Compared with a suburban practice, a one-year loan repayment option made the rural and urban deprived practices 1.6 times and 1.2 times more likely to be chosen, respectively. Nurses were generally more willing than doctors to work in a deprived area in return for loan repayment. The findings suggest that loan repayment may offset health professionals' aversion to working in deprived areas. Such a scheme needs to be piloted to see whether it does offer value for money in recruiting health professionals to under-served areas. PMID- 12121253 TI - Social construction of the managerialism of needs assessment by health and social care professionals. AB - Managerialism in community care has not only radically changed organisational structures delivering care, but the assessment of health and social care needs, the justifications for the assessments, and the experience of those who require publicly funded services. The present paper describes the social construction of the managerialism of needs assessment by health and social care professionals, and illustrates this through the identification of older people as a particular kind of client. The argument draws on 'third way', modernity and postmodernity thinking to show needs assessment as a socially constructed area of welfare. The empirical work in this study is based on the views of 38 health and social care professionals obtained by semi-structured in-depth interviews and a postal questionnaire. The views of these professionals show that the social construction of needs assessment takes place in managing the matching of eligibility criteria against types of services. The key to this process is the application of the concept of management that places health and social care professionals in roles where they are acting for state, voluntary or private agencies, and not in all contexts working together with older people. The study shows that professionals identify older people into two groups or 'classes', i.e. those having health needs as distinct from those with social care. The techniques used amount to an exercise of power by professionals over older people. Change is necessary to break down the dominance by professionals in the needs assessment process. A broader concept of the 'third way' vision by Giddens (1998) is also required to achieve greater relevance to how health and social care is organised, and how relations between professionals and older people are integrated into the idea and practice of participatory care. Therefore, the emancipatory side of modernity remains a largely unfinished project. PMID- 12121254 TI - Prioritising home care needs: research with older people from three ethnic minority community groups. AB - This paper draws on research with three minority ethnic community groups in Manchester. The aim of the study was to examine and prioritise social care needs. Focus groups were chosen as a way of beginning discussion about these issues. The findings suggest that some quality issues are relevant to all communities of older people. However, some were specific to the community groups. This paper describes some of the lessons learned about running focus groups with people whose first language is not English and suggests some considerations for future research. PMID- 12121255 TI - Access to medicines: cost as an influence on the views and behaviour of patients. AB - The present paper explores how charges for medicines incurred by patients influence their decisions for managing acute or chronic conditions, and whether prescription cost and affordability issues are discussed in the general practitioner (GP)-patient encounter. People suffering from dyspepsia, hay fever or hypertension, or those taking hormone replacement therapy, were recruited through three community pharmacies in the North-west of England. Six focus groups were conducted with a total of 31 participants, the majority of whom were non exempt from prescription charges. The management behaviour of those participants who had to pay for their prescriptions, particularly those from less-affluent or deprived backgrounds, was influenced by cost. However, cost was not the overriding influence, with other factors, such as symptom or disease severity, effectiveness, or necessity of treatment, playing a more important part in participants' management decisions. Cost as an issue was reflected in the various strategies used by participants to reduce medication cost, such as not having some prescribed items dispensed, taking a smaller dose or buying a cheaper over the-counter product. Despite the use of numerous strategies, participants did not talk to their GPs about issues of cost and affordability. Participants felt that paying for prescriptions was their problem. There was a belief that discussing cost issues could jeopardise the doctor-patient relationship. Although not the dominant factor, medication cost nevertheless influenced participants when deciding how to manage their condition. Awareness of the existence of prepayment certificates, which can be bought by patients who require regular medication, was low, and this should be addressed through improved information/dissemination. Despite the high level of prescription items exempt, the current level of the prescription charge is still a barrier to obtaining prescription medicines under the National Health Service to those on lower incomes. PMID- 12121256 TI - Good practice towards homeless drug users: research evidence from Scotland. AB - Evidence of large numbers of people who are both homeless and drug dependent, the complexity of their needs, and the many difficulties which they can encounter when trying to access assistance highlight the importance of basic standards of good practice in working with homeless drug users. This is particularly relevant given the growth of new managerialism and the expansion of social care markets occurring within the UK public sector since the 1980s. Within this context, the aim of the present paper is to further understanding of how best to provide support to homeless drug users by examining good practice from the perspectives of both service providers and service users. Data were collected from 48 semi structured interviews (12 with staff and 36 with clients) conducted in six case study agencies (three homelessness agencies and three drug agencies). Interviews were audio-recorded and the data were analysed using Framework. Findings from the study revealed that good practice related to five broad areas. These were: (1) staffing; (2) agency environment; (3) support provided; (4) service delivery; and (5) agency aims and objectives. Similarities between the views of service providers and users were evident. However, differences of opinion suggested that the best definitions of good practice are achieved by consultation with a range of stakeholders (including staff and clients). Data also showed that good practice is fundamentally related to the qualitative and intangible aspects of service provision, and not just to more easily quantifiable inputs, processes, outputs and outcomes. The paper concludes by arguing that the challenge for new managerialism is to build evaluation frameworks which can accommodate this complexity, and thus, begin to portray good practice in a more accurate and meaningful light. PMID- 12121263 TI - Guest editorial: public and user 'involvement' in the UK National Health Service. PMID- 12121264 TI - The experiences of non-medical health professionals undertaking community-based health assessments for people aged 75 years and over. AB - Our objective was to explore the perceptions of allied health professionals who conducted over 3000 home-based health assessments within the general-practice dominated Australian primary healthcare system. A series of semistructured qualitative interviews were carried out within the Department of Veterans' Affairs 'Preventive Care Trial', where health assessments are undertaken by health professionals in the homes of participating veterans and war widows. Health professionals were employed within the Preventive Care Trial to conduct assessments in 10 areas of New South Wales and Queensland. Subjects were mainly registered nurses, but also included a social worker, an occupational therapist, a physiotherapist and a psychologist. The health professionals described positive attitudes towards the health assessments, and showed that they have the broad range of necessary personal and professional skills to undertake them. Home visits were seen as an essential component and the most useful aspects included direct observation of home safety and medications. This study demonstrates that health assessments for older people can be acceptably and competently undertaken by suitably qualified allied health professionals, and that an effective collaborative approach to patient care can be achieved through such a system. PMID- 12121265 TI - Information that informs rather than alienates families with disabled children: developing a model of good practice. AB - The importance to families with disabled children of relevant and accessible information about services has been illustrated in numerous studies and was re emphasised by the Department of Health's 'quality protects' initiative. Indeed, the provision of information and the importance of keeping families informed is frequently viewed as a significant factor within both the concept of empowerment and the facilitation of enabling and participatory processes for service users and their families. However, although there has been considerable research highlighting parents' information needs, there has been significantly less exploration of how parents would actually like to receive this information. This paper seeks to bridge this knowledge gap and also discusses the empowering potential of user-friendly information. Drawing upon data collected from focus group discussions with parents caring for children with a range of disabilities or chronic illnesses, this paper explores how the families of service users would like to receive information. In particular, it examines the criteria by which parents judge the quality of information and their ideas as to what constitutes good practice, especially in terms of how information is presented, its content and the way it is delivered. Using these ideas and criteria, the paper begins to develop a model of good information practice that is both three-dimensional and personally interactive. Indeed, parents' desire for a combination of personal guidance and good-quality information, whether in the form of in-depth booklets or shorter directories, is viewed as being of paramount importance and, furthermore, as having an important empowering potential. PMID- 12121266 TI - Health and social factors for health visitor caseload weighting: reliability, accuracy and current and potential use. AB - The reliability and validity of data collection and recording for a health visitor caseload weighting system operating in a large combined acute and community trust in Bristol was examined. Client families were recruited from all participating (service) health visitors and were interviewed by a research health visitor attached to the project. The presence or absence of 28 health needs factors, selected for their known impact on health in the longer term, was subsequently compared with the service health visitor's own caseload records and with the computer record derived from them. Substantial differences were determined in the records obtained between the service and research health visitors and between the written and computerised records. Whilst the mean total score recorded for each family by the service health visitor was 2.0, the average of the differences in the total number of health factors recorded by the service and research health visitors was 1.9. Discrepancies were mostly associated with differences in interpretation of definitions, knowledge of recent events, changing circumstances and issues of confidentiality. Health factors at particular risk of being misinterpreted and those associated with other health factors were identified in order to propose a reduced factor set with greater inherent reliability and validity. At the level of the ward, the caseload weighting score, as currently defined, is highly correlated with standard deprivation indices in widespread common use. The results of this study indicate the need for users of the caseload weighting data to decide on the primary function of this data set - area-based community profiling or identification of at-risk client families in the community. This decision will inform further efforts to identify the most useful factors, tighten definitions, streamline data collection and train health visitors in their use. Interpretation of data will be facilitated by a scientifically developed scoring system. This work will assist trusts, both locally and nationally, in rationalising their allocation of health visiting activities to areas of greatest need. PMID- 12121267 TI - Variant Creutzfeldt-Jakob disease: costs borne by families. AB - The objectives of this study were: (1) to estimate the costs borne by families caring for patients with variant Creutzfeldt-Jakob disease (vCJD); (2) to contextualise results to recent policy initiatives, and (3) to consider the methodological problems of estimating costs of care. Semi-structured interviews and a follow-up postal questionnaire, eliciting costs to families both before and after the patient's death, were carried out. Participants included 19 families of patients with vCJD. Cost profiles were constructed, detailing key time and financial costs associated with their relative's illness and death accruing to families. Main outcome measures included total, median and ranges of relevant cost elements. Sensitivity analyses, comparing high and low cost estimates, were undertaken. The total time cost to families before patient's death ranged between 605 and 9230 hours (median 2006 hours). Applying low cost estimates, families incurred between pound 2616 and pound 39 588 (median pound 14 481) in forgone earnings and between pound 2699 and pound 18 558 (median pound 8049) in marginal sundry costs before the patient's death. The value of care provided by families ranged between pound 0 and pound 87 303 (median pound 9652) at low cost estimates. Many families continued to incur costs after the patient's death, with low cost estimates per week ranging between pound 0 and pound 176 (median pound 29). Costs to families associated with vCJD were substantial and greatly exceeded benefit entitlements. These costs were high even if patients received care in hospital, varied as the illness progressed and continued after patients' deaths. The National Carers Strategy does not consider fully the needs of some groups of carers or the full range or magnitude of potential costs to families associated with caring. PMID- 12121268 TI - Community-backed drug initiatives in the UK: a review and commentary on evaluations. AB - This article presents a review of evaluations of community-backed initiatives for drug users in the UK. Community-backed responses have taken a number of approaches. Despite accessing over 250 publications relating to initiatives in the course of this study, a paucity of published evaluative evidence on 'what works?' and 'why does it work?' was apparent. The readily accessible literature is examined under five broad types of initiative: self-help groups, parents' groups, residents' groups, community development groups and diversionary activity groups. In a further commentary on evaluation issues for such drug initiatives, it is argued that they are under-evaluated, partly because they are on the margins of drug services and partly because of a lacuna of support in evaluation skills. PMID- 12121269 TI - Parenting issues that may be addressed through a confidential helpline. AB - This study identifies what parents might expect from a confidential helpline and highlights areas of parental concern in the task of child-rearing. A community sample of 424 families from four schools was collected. Although many issues raised concerned parents, those particularly cited were behaviour management, school bullying and drug and alcohol problems. It is clear that there is a perceived need by parents for input into the parenting process; parents were seeking information and advice rather than support. The question of which aspects of parenting can be developed, either through a helpline or other services, is discussed. PMID- 12121270 TI - Working with families in Tower Hamlets: an evaluation of the Family Welfare Association's Family Support Services. AB - This paper describes an evaluation carried out by South Bank University of the work of the Family Welfare Association's (FWA's) Family Support Services (FSSs) in Tower Hamlets, London. Tower Hamlets is a multi-racial area in east London that, according to the 1991 census, has high levels of poverty, overcrowding and unemployment. Increasing poverty and social exclusion, which further entrench inequalities in health, are reported by sources such as government, health and social services and research as requiring innovative local responses to meet pressing welfare needs. The evaluation reported here examined three projects: Family Support, Building Bridges and Quality Protects - these are referred to collectively as FSSs. The evaluation shows that FSSs are innovative services that demonstrate effective ways of working with vulnerable families affected by experiences of racism, bullying, mental health difficulties, domestic violence or child abuse. In common with other successful initiatives in the UK and abroad, FSSs are aimed to be non-stigmatising, non-intrusive and responsive to the ethnicity, views and specific needs of families. This paper focuses on the participatory work of FSSs with families to illustrate effective methods of quality support, detail outcomes, and draw lessons for policy and practice. PMID- 12121271 TI - Two bi-allelic single nucleotide polymorphisms within the promoter region of the horse tumour necrosis factor alpha gene. AB - Primers based on GenBank sequences within the 5' untranslated region (UTR) of the human and horse tumour necrosis factor alpha (TNF-alpha) genes were designed and used to amplify a 522-bp product. Sequencing of five clones derived from five independent PCRs obtained from three different animals of three different breeds (Old Kladruber, Akhal-Teke and Shetland Pony) revealed a high level of sequence identity to the TNF-alpha promoter regions of other species. The existing GenBank horse sequences were confirmed and extended upstream by 230 nucleotides. Based on the sequence obtained, a new horse-specific forward primer was designed to amplify a 213-bp PCR product, which was screened for polymorphism using single strand conformation polymorphism (SSCP). Three allelic variants of the horse TNF alpha gene were identified and sequenced (GenBank accession numbers ADF 349558 60). Two single nucleotide polymorphisms explained the existence of the three SSCP alleles detected: C/T and T/C single base pair substitutions at positions 137 and 147, respectively. Differences in allelic frequencies between Old Kladruber and Akhal-Teke breeds were observed. PMID- 12121272 TI - Analysis of candidate genes on chromosome 19 in coeliac disease: an association study of the KIR and LILR gene clusters. AB - Coeliac disease is strongly heritable, with more than half of the genetic susceptibility estimated to come from genes outside the HLA region. Several candidate regions have been suggested from genome-wide linkage studies including chromosome 19q13.4 where linkage has been replicated between populations. The natural killer (NK) cell immunoglobulin-like receptors (KIRs) and leukocyte immunoglobulin-like receptor (LILR, also known as ILT and LIR) gene clusters lie within this region in the leukocyte receptor cluster (LRC). KIR molecules are involved in cytotoxic lymphocyte function and expressed by intraepithelial T and NK cells in the duodenum. We studied 132 unrelated UK Caucasian coeliac patients and their parents together with a control group of 171 UK Caucasians. PCR-SSP for KIR2DL1, KIR2DL2, KIR2DL3, KIR2DL5, LILRA3 (ILT6), LILRA3 deletion and an LILRA3 exon 3 single nucleotide polymorphism (SNP) allowed classification of KIR genotypes into five categories and determination of homozygosity or heterozygosity for the common A and B type KIR haplotypes (as defined in the text) and for the LILRA3 deletion. Case-control analysis found no association of the five KIR genotype categories, the A or B KIR haplotypes, the LILRA3 gene deletion or the LILRA3 exon 3 SNP with coeliac disease. A transmission disequilibrium test also found no association of the A and B KIR haplotypes or the LILRA3 gene deletion with coeliac disease. PMID- 12121273 TI - Frequencies of the -330 (T --> G) IL-2 and -590 (T --> C) IL-4 gene polymorphisms in a population from south-eastern Brazil. AB - Polymorphisms in the promoter regions of cytokine genes may affect their transcription. A T/G substitution at position -330 of the interleukin-2 (IL-2) gene and a T/C substitution at position -590 of the interleukin-4 (IL-4) gene have been described previously. The -590 (T --> C) IL-4 gene polymorphism was associated with asthma and atopy in US and Japanese populations. Population genetics is a useful tool for determination of the biological significance of genetic polymorphisms. The aim of this study was to investigate the frequencies of polymorphisms in the promoter regions of the IL-2 and IL-4 genes in a population from south-eastern Brazil and to compare them with those published for other populations. Allele frequencies were estimated in 114 unrelated individuals from Sao Paulo State. These subjects had an average age of 41.2 years (+/- 12.4 years) and the ethnic composition of the sample was: 78.07% Caucasian, 11.4% Black and 10.53% Mulatto. DNA from subjects was extracted from epithelial buccal cells, and the PCR-RFLP technique was employed to investigate the -330 (T --> G) IL-2 and -590 (T --> C) IL-4 gene polymorphisms. The allele frequency of the IL-2 gene polymorphism obtained in our study was similar to that found in UK Caucasoid groups. The T allele frequency of the IL-4 gene polymorphism observed in the Caucasian Brazilian group was similar to that found in UK and Australian populations, while the frequency observed for the Black Brazilian group was similar to that found in Japanese and Kuwaiti Arab populations. The results for the -330 (T --> G) IL-2 and -590 (T --> C) IL-4 polymorphisms are consistent with the high contribution of European lineages to the population in south-eastern Brazil. PMID- 12121274 TI - HLA-DRB1*12 is associated with protection against complicated typhoid fever, independent of tumour necrosis factor alpha. AB - We investigated whether HLA-DR2 or -DR12 alleles in 63 Javanese patients with complicated or non-complicated typhoid fever were associated with severity of disease. No association was observed between HLA type and susceptibility to disease. However, in patients we did find a negative association of DR12 (DRB1*12021) with complicated typhoid fever (P = 0.05; OR = 0.3; 95% CI: 0.1 1.0). No effect of DR2 (DRB1*1502) on outcome (P = 0.46; OR = 1.5; 95% CI: 0.5 4.5) was demonstrated. The odds ratio for DR12 remained unchanged after adjusting for DR2. Tumour necrosis factor alpha (TNF-alpha) production capacity in lipopolysaccharide (LPS)-stimulated whole blood culture, as measured by non equilibrium radioimmunoassay, was significantly lower in complicated than in non complicated cases (P = 0.02), confirming previous data. No significant correlation of either DR12 (P = 0.47) or DR2 (P = 0.89) was found with TNF-alpha production capacity. Apparently, protection against complications by DR12 is attributable to other mechanisms. PMID- 12121275 TI - FcgammaRIIA, FcgammaRIIIA and FcgammaRIIIB polymorphisms in Spanish patients with systemic lupus erythematosus. AB - Linkage studies on human families suggest that receptors for the Fc fragments of immunoglobulin G (IgG) (FcgammaRs) could be implicated in the susceptibility to, or the progression of, some autoimmune diseases. In this work we analyse the possible role of polymorphic variants of FcgammaRIIA, FcgammaRIIIA and FcgammaRIIIB genes in the susceptibility to systemic lupus erythematosus, the prototype systemic autoimmune disease. A total of 276 Spanish patients (34 male and 242 female) with systemic lupus erythematosus were included in this cross sectional study. The FcgammaRIIA-131, FcgammaRIIIA-176 and FcgammaRIIIB-NA1/NA2 polymorphisms were investigated in the patient group as well as in 194 ethnically matched controls using polymerase chain reaction-amplification refractory mutation system (PCR-ARMS). Statistical comparisons of genotype frequencies were performed using the chi2 test. In the case of the FcgammaRIIIB-NA1/NA2 polymorphism, an increase in the frequency of homozygous NA2/NA2 in patients was found (61.2 vs. 51.0%; P = 0.03; OR = 1.5; 95% CI = 1.03-2.24), as well as a decrease in the frequency of the NA1/NA2 genotype (28 vs. 38.7%; P = 0.02; OR = 0.6; 95% CI = 0.41-0.92). These associations were independent of patient gender and HLA-DRB1 specificities. With respect to the FcgammaRIIA-131 and FcgammaRIIIA 176 polymorphisms, no differences were found between patients and controls. Patient stratification according to their lupus-related nephritis status gave similar genotypic distribution patterns in both disease categories in all the cases. PMID- 12121276 TI - Structural basis of allotypes of ecto-nucleotide pyrophosphatase/phosphodiesterase (plasma cell membrane glycoprotein PC-1) in the mouse and rat, and analysis of allele-specific xenogeneic antibodies. AB - Ecto-nucleotide pyrophosphatase/phosphodiesterases (E-NPPs) have been implicated in bone calcification, type II diabetes, control of purinergic signalling and tumour invasion. The gene for the plasma cell membrane glycoprotein PC-1 in the mouse (Enpp1) has been known since 1970 to exist in two allelic forms, but their structural basis was heretofore unknown. We show that the Enpp1a and Enpp1b alleles differ by only two amino acids, at positions 650 and 679 in the C terminal nuclease-like domain. Histidine 650 but not arginine 679 forms an essential part of the Enpp1a epitope recognized by monoclonal antibody IR-518. Sequences of LEW and LOU rats and the rat glioma cell line C6 differ from that of the mouse by about 60 amino acids. The LOU and C6 cell line sequences differ by only three amino acids, but differ from the LEW sequence by 10 amino acids. All three rat strains possess the mouse Enpp1b allele at positions 650 and 679. Despite numerous other differences from the mouse, rats immunized with Enpp1a mouse cells have generated monoclonal antibodies specific for the Enpp1a allele, suggesting that amino acids 650 and 679 may be particularly immunogenic. The cytoplasmic tails of the mouse and rat are highly conserved, but are significantly different from human cytoplasmic tails. PMID- 12121277 TI - Codominant expression of the polymorphic MICA alloantigens encoded by genes in the HLA region. AB - The HLA-related, polymorphic MHC class I-related chain A (MICA) gene encodes a 383-amino acid polypeptide, with three extracellular domains (alpha1, alpha2 and alpha3), a transmembrane region and a cytoplasmic tail. We have previously shown that freshly isolated endothelial cells, fibroblasts, keratinocytes and monocytes express MICA, while peripheral blood CD4+, CD8+ or CD19+ lymphocytes do not. This polymorphic MICA molecule is a target for specific alloantibodies in sera from kidney, heart and lung transplant recipients, although its possible role during graft rejection remains to be demonstrated. In this study we investigated whether there is codominance in the expression of MICA. We isolated RNA from a heterozygous cell line (HCT116), previously shown by sequencing-based typing to be MICA*001/MICA*00902, as well as 12 clones derived from it. Thereafter, we retrotranscribed the RNA into cDNA, and performed a molecular typing using MICA sequence specific oligonucleotides (SSOP). Using this approach, we detected the RNA encoding MICA*001 and MICA*00902 in all the clones and in the parental cell line, indicating that MICA is codominantly expressed. This codominant expression was further confirmed by cloning and sequencing plasmids encoding these two alleles produced from the same HCT116 RNA preparation. We also produced the two recombinant MICA proteins (MICA*001 and MICA*00902). They reacted with rabbit anti-MICA polyclonal antibodies by ELISA and Western blot, indicating that the plasmids carrying the cDNA inserts probably encode functional MICA proteins. This strongly suggests that, like the HLA class I and class II proteins, MICA is codominantly expressed. The codominant expression of the polymorphic, HLA-like MICA alloantigens may have implications for the immune response elicited by the allograft in organ transplantation. PMID- 12121278 TI - Unbalanced amounts of HLA-DQA1 allele mRNA: DQA1*03 shows high and DQA1*0501 low amounts of mRNA in heterozygous individuals. AB - Genes of the HLA-DR, DQ region confer strong susceptibility to type 1 diabetes mellitus (IDDM). A possible mechanism of susceptibility is a difference in the amounts of transcripts of predisposing and neutral or protective haplotypes. In this study we developed an assay to compare the amounts of mRNA of two distinct HLA-DQA1 alleles in peripheral blood lymphocytes (PBLs) of heterozygous individuals, using a quantitative RT-PCR with an internal standard covering all HLA-DQA1 specifities. We also developed an algorithm to calculate the amounts of mRNA for two distinct alleles in heterozygous individuals based on the comparison to the same internal standard. In total, 37 HLA-DQA1 heterozygous individuals were analysed, including patients with IDDM (n = 14) and healthy controls (n = 23). Intra-individually, we observed different amounts of mRNA for different HLA DQA1 alleles in the order: HLA-DQA1*03 > *01 > *0201 > *05. This order was observed in all individuals. We also observed a variation in the ratio of these unbalanced amounts of mRNA in individuals with the same HLA-DQA1 allele combinations. In all allele combinations the average ratio was increased in patients with IDDM compared to the control samples. HLA-DQA1*03 positive and DQA1*03, *05 heterozygous patients had the highest average ratios. Nevertheless, based on limited sample numbers, these differences did not reach significance. We therefore conclude that variations between HLA-DQA1 alleles are not limited to the nucleotide sequence but are also found at the level of amounts of mRNA. PMID- 12121279 TI - CD80 (B7-1) and CD86 (B7-2) genes and genetic susceptibility to coeliac disease. AB - The role of the costimulatory molecules B7-1 (CD80) and B7-2 (CD86) in T-cell activation makes them good candidates for coeliac disease susceptibility genes. We conducted a genetic linkage study of the CD80/86 gene region in the general Finnish population and in a local subisolate. No linkage was found in either population. PMID- 12121280 TI - A novel DRB1*09 allelic sequence in the Jing ethnic minority of China. AB - A new DRB1 allele, DRB1*0902, has been identified in an individual of the Jing ethnic minority. Its sequence was confirmed by sequencing of PCR products and clones. This allele differed by three nucleotides from DRB1*09012 at positions 157, 161 and 166, and resulted in amino acid motif substitution from VAES to DAEY. PMID- 12121281 TI - The interleukin-1 receptor antagonist gene: a single-copy variant of the intron 2 variable number tandem repeat (VNTR) polymorphism. AB - A variable number tandem repeat (VNTR) polymorphism exists within intron 2 of the human interleukin-1 receptor antagonist gene, consisting of perfect repeats of an 86-bp sequence. Five allelic variants have been identified wherein the number of repeats varies from two to six. This is the first report of a rare, single-copy allele designated IL1RN*0. PMID- 12121282 TI - Buffalo (Bubalus bubalis) interleukin-2: sequence analysis reveals high nucleotide and amino acid identity with interleukin-2 of cattle and other ruminants. AB - A 4400-bp genomic sequence and a 332-bp truncated cDNA sequence of the interleukin-2 (IL-2) gene of Indian water buffalo (Bubalus bubalis) were amplified by polymerase chain reaction and cloned. The coding sequence of the buffalo IL-2 gene was assembled from the 5' end of the genomic clone and the truncated cDNA clone. This sequence had 98.5% nucleotide identity and 98% amino acid identity with cattle IL-2. Three amino acid substitutions were observed at positions 63, 124 and 135. Comparison of the predicted protein structure of buffalo IL-2 with that of human and cattle IL-2 did not reveal significant differences. The putative amino acids responsible for IL-2 receptor binding were conserved in buffalo, cattle and human IL-2. The amino acid sequence of buffalo IL-2 also showed very high identity with that of other ruminants, indicating functional cross-reactivity. PMID- 12121283 TI - A recently described polymorphism in the CD28 gene on chromosome 2q33 is not associated with susceptibility to type 1 diabetes. AB - Type 1 diabetes mellitus is an autoimmune disease with a strong genetic background. The CTLA4 gene region (IDDM12) has been implicated in genetic susceptibility to type 1 diabetes by genome scanning and both family- and population-based analyses. As the genes encoding the costimulatory molecules CTLA4 and CD28, which compete for the receptor B7, reside close together on chromosome 2q33 and have high sequence homology, we investigated a recently described polymorphism in intron 3 of the CD28 gene and the CLTA4 codon 17 polymorphism in 176 patients with type 1 diabetes and 220 healthy controls. Whereas CTLA4 was found to be associated with type 1 diabetes, the frequency of the CD28 polymorphism did not differ between patients and controls, either in the entire sample or after stratification for CTLA4 genotype. Thus, the CD28 intron 3 polymorphism does not appear to be associated with susceptibility to type 1 diabetes. PMID- 12121286 TI - Behavior and physiology of mice lacking the GABAA-receptor delta subunit. AB - PURPOSE: Mice with a targeted disruption of the gamma-aminobutyric acid (A) receptor (GABA(A)R) delta subunit exhibited spontaneous seizures and a strikingly selective attenuation of responses to neuroactive steroids, but not to other neuromodulators. This study further characterized the behavior and physiology of the delta mutants. METHODS: Seizure susceptibility to pentylenetetrazol (PTZ) injection, intracellular recordings, and whole-cell patch recordings in dentate granule neurons was determined. RESULTS: Susceptibility to PTZ-induced convulsive seizures was significantly higher in -/- versus +/+ mice. In intracellular recordings, the evoked inhibitory postsynaptic potentials (IPSPs) had much faster decay tau in -/- mutants (25 +/- 3.5 ms) compared with +/+ controls (51 +/- 5.2 ms). In whole-cell patch recordings, the decay tau of pharmacologically isolated miniature spontaneous GABA(A)R-mediated synaptic currents (mIPSCs) from -/- mice was only slightly, but significantly faster (5.7 +/- 0.13 ms; n = 18) than that of +/+ controls (6.8 +/- 0.32 ms; n = 20). Furthermore, mIPSC prolongation by bath application of alphaxalone (10 microM), was much smaller in -/- mice (52 +/- 12%; n = 3) compared with +/+ littermates (192 +/- 14%; n = 3). CONCLUSIONS: Faster decay of evoked IPSPs and mIPSCs is consistent with altered GABA(A)R subunit composition in the delta mutants, but suggests predominant extrasynaptic delta subunit location in the dentate gyrus of normal mice. Faster-decaying IPSPs likely contribute to the proepileptic phenotype of the delta mutants. Reduced neurosteroid sensitivity might also contribute to seizure susceptibility. It remains to be determined whether delta subunit-containing GABA(A)Rs represent the actual target of neurosteroids or whether the behavioral and physiologic sensitivity to neurosteroids is indirectly altered in the delta-/- mice. PMID- 12121287 TI - Antiepileptic actions of neuropeptide Y in the mouse hippocampus require Y5 receptors. AB - PURPOSE: Recent evidence suggests an antiepileptic role for neuropeptide Y (NPY) in the central nervous system. The precise receptor subtypes mediating the inhibitory actions of NPY in the hippocampal formation, however, remain unclear. In vitro studies suggest a role for Y2 receptors in modulating excitatory hippocampal synaptic transmission and epileptiform discharge. In vivo studies implicate Y5 receptors. Here we used pharmacologic tools and Y5-receptor knockout mice to examine the role of Y5 receptors in mediating the antiexcitatory and antiepileptic actions of NPY in the hippocampal formation. METHODS: Hippocampal slices were obtained from age-matched wild-type (WT; 129 s3/svimj) and Y5 receptor knockout (Y5R KO) mice generated on the same background strain. Extracellular or whole-cell voltage-clamp recordings were obtained in area CA3 pyramidale. Evoked population spikes or excitatory postsynaptic currents were monitored during bath application of NPY, NPY13-36, or human pancreatic polypeptide (hPP). In some slices, zero-magnesium cerebrospinal fluid (CSF) was used to evoke spontaneous epileptiform discharges. RESULTS: NPY and NPY agonists with preference for either Y2 (NPY13-36) or Y5 (hPP) receptor subtypes caused a significant reduction in population spike and excitatory postsynaptic current (EPSC) amplitudes in slices from WT mice. NPY (and NPY agonists) also suppressed zero-magnesium epileptiform burst discharge in slices from WT mice. In contrast, bath application of NPY, NPY13-36, or hPP had no effect in slices from Y5R KO mice. CONCLUSIONS: NPY modulates excitatory synaptic transmission and inhibits limbic seizure activity in the mouse hippocampus. The antiepileptic actions of NPY, in the mouse, appear to require activation of hippocampal Y5 receptors. PMID- 12121288 TI - Progesterone reduces pentylenetetrazol-induced ictal activity of wild-type mice but not those deficient in type I 5alpha-reductase. AB - PURPOSE: To investigate the importance of progesterone (P4) metabolism by the 5alpha-reductase type I enzyme in mitigating P4 antiseizure effects. METHODS: Ovariectomized, female homozygous and heterozygous 5alpha-reductase type I knockout mice (n = 23) and their wild-type siblings (n = 31) were administered P4 (1.0 mg), and their pentylenetetrazol (PTZ)-induced ictal behaviors were compared with those of vehicle-administered mice (n = 49). RESULTS: Mice deficient in the 5alpha-reductase type I enzyme administered P4, or vehicle-administered control mice, had significantly shorter latencies and increased incidence of PTZ-induced hindlimb extension and death than did wild-type mice administered P4. CONCLUSIONS: These data suggest that P4's metabolism by the 5alpha-reductase type I enzyme may mitigate some of P4's antiseizure effects in the PTZ-induced seizure model. PMID- 12121289 TI - Involvement of the neuropeptide orphanin FQ/nociceptin in kainate and kindling seizures and epileptogenesis. AB - PURPOSE: To investigate the role of orphanin FQ/nociceptin (OFQ/N) in epilepsy, we analyzed (a) proOFQ/N (the OFQ/N precursor) and ORL-1 (the OFQ/N receptor) messenger RNA (mRNA) levels in the kainate and in the kindling models of epilepsy in the rat; and (b) seizure expression in proOFQ/N knockout mice. METHODS: Epilepsy models: kainate and kindling. Northern blot analysis, radioactive in situ hybridization. RESULTS: Increased proOFQ/N mRNA levels were found in the thalamus (reticular nucleus) after kainate administration. In contrast, ORL-1 gene expression decreased dramatically in the amygdala, hippocampus, thalamus, and cortex after kainate administration. OFQ/N knockout mice displayed reduced susceptibility to kainate-induced seizures, in that (a) lethality was reduced, (b) latency to generalized seizure onset was significantly prolonged, and (c) behavioral seizure scores were significantly reduced. Furthermore, kindling progression was delayed in OFQ/N-/- mice. CONCLUSIONS: These data indicate that limbic seizures are associated with increased OFQ/N release in multiple brain areas, causing downregulation of ORL-1 receptors and activation of OFQ/N biosynthesis in selected areas, and support the notion that the OFQ/N-ORL-1 system may play a facilitatory role in ictogenesis and in epileptogenesis. PMID- 12121290 TI - Short-term plasticity of hippocampal neuropeptides and neuronal circuitry in experimental status epilepticus. AB - PURPOSE: We used a model of self-staining status epilepticus (SSSE), induced by brief intermittent stimulation of the perforant path in unanesthetized rats, to study the mechanism of initiation and of maintenance of SSSE and the role of neuropeptides in those processes. METHODS: The perforant path was stimulated intermittently for 7 min (ineffective stimulation) or 30 min (generating SSSE). Peptides and their agonists and antagonists were delivered either intraperitoneally, or directly into the hippocampus through a implanted cannula. Behavior and electroencephalogram (EEG) were recorded through a videotape telemetry system with automatic spike and seizures detection programs, which were supplemented by manual review of the records to confirm the diagnosis. Immunocytochemistry and enzyme-linked immunosorbent assay followed published methods. RESULTS: Initiation of SSSE was blocked by many agonists of inhibitory neurotransmitters or neuromodulators, and by many antagonists of excitatory synapses, and was facilitated by agents with the opposite action, suggesting the activation of a complex circuit with multiple potential entry points. Once SSSE was established, however, only N-methyl-d-aspartate (NMDA)-receptor ligands and a few neuropeptides had major effects on its maintenance. Galanin and dynorphin had powerful anticonvulsant roles in the maintenance phase of SSSE, whereas somatostatin and neuropeptide Y suppressed seizures only transiently. SSSE seemed to induce maladaptive changes in neuropeptides: it depleted the hippocampus of the galanin- and dynorphin-immunoreactive (IR) fibers, which normally function as endogenous anticonvulsants; whereas it induced overexpression of the proconvulsant neuropeptides substance P and neurokinin B; however, late in the course of SSSE, galanin-IR interneurons appeared in the dentate hilus. CONCLUSIONS: Initiation of SSSE seems to involve a circuit with many points of entry, and blockage of any point along this circuit inhibits the development of SSSE. Far fewer agents alter the maintenance phase of SSSE. Galanin, dynorphin, somatostatin, and neuropeptide Y have anticonvulsant roles, matching the previous described convulsant role of substance P and neurokinin B. Galanin and dynorphin seem to undergo maladaptive changes, which appear to play an important role of the maintenance phase of SSSE. Later, the de novo expression of inhibitory neuropeptides in novel cells in hippocampus coincides with the waning of seizures and may play a role in their termination. PMID- 12121291 TI - Functional role of inflammatory cytokines and antiinflammatory molecules in seizures and epileptogenesis. AB - PURPOSE: We investigated the changes in the expression of proinflammatory cytokines and related molecules in the rodent hippocampus after the induction of limbic seizures. We then studied the effects of pharmacologic intervention on the interleukin (IL)-1 system on limbic seizures and the susceptibility to seizures of transgenic mice overexpressing the naturally occurring antagonist of IL-1 (IL 1Ra) in astrocytes. METHODS: Limbic seizures were induced in rodents by intrahippocampal injection of kainic acid or bicuculline methiodide or by electrical stimulation of the hippocampus causing status epilepticus (SE). Seizure activity was recorded by EEG analysis and behavioral observation according to Racine's scale. Cytokine expression in the hippocampus was studied by reverse transcriptase-polymerase chain reaction (RT-PCR) followed by Southern blot quantification of the various messenger RNAs (mRNAs) and by immunocytochemistry. RESULTS: We found that limbic seizures rapidly and transiently enhanced IL-1beta, IL-6, and tumor necrosis factor (TNF)-alpha mRNA in the hippocampus with a peak effect at 6 h after SE. Immunoreactivity of the various cytokines was increased in glia. The increase of IL-1Ra was delayed because the peak effect was observed at 24 h after SE. Moreover, IL-1Ra was not produced in large excess, as during peripheral inflammation but in a molar ratio to IL-1beta of 1:1. Intrahippocampal injection of IL-1beta worsened seizure activity, whereas IL-1Ra was a powerful anticonvulsant in various models of limbic seizures. Transgenic mice overexpressing IL-1Ra in astrocytes were less sensitive to bicuculline-induced seizures. CONCLUSIONS: This study shows that limbic seizures in rodents rapidly and reversibly induce proinflammatory cytokines in glia and suggests that changes in the IL-1Ra/IL-1beta ratio in brain may represent an effective physiopathologic mechanism to control seizures. PMID- 12121292 TI - TsTx toxin isolated from Tityus serrulatus scorpion venom-induced spontaneous recurrent seizures and mossy fiber sprouting. AB - PURPOSE: TsTx is a scorpion alpha-type toxin that binds to site 3 of the Na+ channels in a voltage-dependent mode, slowing or blocking the inactivation mechanism of these channels (Possani et al., Eur J Biochem 1999). This binding increases depolarization time of the channel and consequently induces excessive neurotransmitter release. Previously we reported that hippocampal injection of TsTx induces clonic convulsions, electrographic seizures, and hippocampal damage. This investigation was designed to characterize the long-term behavioral, electroencephalographic (EEG), and histopathologic features after a single TsTx injection into the rat hippocampus. METHODS: Cannulas and electrodes were stereotaxically implanted in the CA1 dorsal hippocampus of rats. Three days after surgery, the animals were injected into the hippocampus with 1 microl of TsTx, 2 microg (n = 9) or 0.1 M phosphate buffer (n = 5). After injection, EEG records and behavioral observations were made during 10 h. For a period of 4 months, the animals were observed through direct visual observation for 8 h/day, 5 days/week for occurrence of convulsive seizures. At the end of the experiment, the animals were processed for histologic analyses. Sections 40- and 20-microm thick were stained according to neo-Timm or Nissl methods, respectively. Cell counts in the cresyl violet-stained sections were performed within the hippocampal pyramidal cell layers (CA1, CA3, and CA4) and granule cell of the dentate gyrus. RESULTS: Fifteen minutes after TsTx injection, facial automatisms, rearing, masticatory jaw movements, sniffing, and wet-dog shakes were observed. In approximately 1 h, limbic convulsions characterized by forelimb clonus, rearing, and falling after generalized clonic convulsion with jumping, wild running, and falling were observed. EEG showed isolated spikes and clusters of spikes that started in the hippocampus and evolved to the cortex, isolated seizures, and epileptic discharges delayed 1-2 min. These recurred repeatedly characterizing the status epilepticus (SE). SE was characterized in 77.5% of animals. Seizures were no longer observed 24 h after the injection. Spontaneous recurrent seizures (SRSs) occurred 31-49 days after TsTx injection. All animals that showed SE in the acute period developed SRSs. Facial myoclonus, generalized clonus, forelimb clonus, rearing, and falling characterized the seizures. The seizure frequency was 1-2 per animal per week. All rats injected with TsTx had significantly fewer cells in CA1, CA3, and CA4 subfield of the hippocampal formation compared with the animals of the control group (p < 0.05, analysis of variance, followed by Tukey test). Neo-Timm-positive granules, normally absent in the supragranular layer, were present in dentate gyrus of rats with TsTx-induced SRSs. CONCLUSIONS: Our results suggest that SRSs observed in this study may be a consequence of the TsTx-induced SE. All animals injected with the toxin showed massive neuronal loss in the hippocampal subfields CA1, CA3, and CA4, but only those that had SRSs showed mossy fiber sprouting in the supragranular layer of the dentate gyrus. This shows synaptic reorganization that also is observed in human epileptogenic tissue. These results indicate that TsTx toxin may be a useful tool for studies on neuronal lesions and/or experimental epilepsy. Studies on the mechanisms involved are under investigation. PMID- 12121293 TI - Behavioral, morphologic, and electroencephalographic evaluation of seizures induced by intrahippocampal microinjection of pilocarpine. AB - PURPOSE: We studied, by means of video-EEG and neo-Timm histochemistry, the behavioral, electrophysiologic, and structural characteristics of seizures induced by intrahippocampal microinjection of pilocarpine (HIP-PILO), a selective model of temporal lobe epilepsy (TLE). METHODS: We investigated the behavioral and electrophysiologic (hippocampus and amygdala EEG) evaluation of status epilepticus (SE) induced by HIP-PILO and the consequent spontaneous recurrent seizures (SRSs). We evaluated hippocampal structural rearrangements after SE by means of neo-Timm staining. RESULTS: HIP-PILO induced SE in 17 (71%) of 24 animals. Although three animals displayed spontaneous remission of SE (not used in analysis) before the established SE duration (90 min), none of those undergoing SE died. Of SE animals, 10 (71%) of 14 had SRSs. All animals with SE had clear-cut mossy fiber sprouting (MFS) in the inner molecular layer of the dentate gyrus and epileptiform activity in hippocampus and amygdala. CONCLUSIONS: HIP-PILO rats displayed SE, SRS, MFS, and limbic epileptiform activity, without animal loss after SE. Thus, our data support this as a selective and efficient model of TLE. PMID- 12121294 TI - Short-term effects of pilocarpine on rat hippocampal neurons in culture. AB - PURPOSE: Status epilepticus (SE) has been considered an epileptogenic factor in humans. In the pilocarpine (PILO) model, after a brief period marked by SE, the rats exhibit recurrent spontaneous seizures, mimicking the clinical features of temporal lobe epilepsy. The aim of our study was to identify the molecular actions of PILO that could account for its ability to induce SE. METHODS: Whole cell mode of the patch-clamp technique was applied to cultured hippocampal neurons (2-3 weeks old) in the absence and in the presence of PILO (1-10 microM), to study the spontaneous activity, the evoked, and the miniature postsynaptic currents. The postsynaptic currents were isolated pharmacologically. RESULTS: PILO (1 and 10 microM) caused an initial increase followed by a decrease in the frequency of spontaneous activity. The increase in the frequency of excitatory postsynaptic currents (EPSCs) and inhibitory PSCs (IPSCs) was blocked by atropine (1 microM), indicating that this effect is mediated through muscarinic receptors. PILO also promoted a brief increase of the amplitude of IPSCs indirectly evoked by stimulation of a neuron synaptically connected to the neuron under study. Conversely, PILO promoted a sustained increase on the amplitude of electrically evoked EPSCs. In presence of tetrodotoxin (TTX; 300 nM), PILO (1 microM) increased the frequency of miniature EPSCs and IPSCs without changing their amplitude during the first 3 min of application. CONCLUSIONS: These results indicate that PILO acting through muscarinic receptor causes an imbalance between excitatory and inhibitory transmission that can result in the generation of SE observed in animals acutely treated with PILO. PMID- 12121295 TI - Epileptogenesis during development: injury, circuit recruitment, and plasticity. AB - PURPOSE: To use animal models of variable seizure induction in rats at different developmental stages to determine contributing factors for spontaneous seizures resulting from status epilepticus (SE) early in life. METHODS: Two models of SE with distinct modes of seizure induction, lithium-pilocarpine (LiPC) and perforant path stimulation (PPS), were used at different ages. Multiple methods of determining neurodegeneration during an acute period and plastic changes in those monitored during the chronic phase were used. RESULTS: Different modes of seizure induction lead to varying types and extents of damage, dependent on the age of the animals at the time of insult. LiPC resulted in injury to animals as young as 2 weeks and became widespread in animals 3 weeks old, whereas widespread damage after PPS was not seen until P35. Rats at an age with widespread damage in response to seizures also showed extensive immediate-early gene activation and often developed spontaneous seizures and features of hippocampal plasticity seen in the epileptic brain. CONCLUSIONS: SE early in life results in multiple consequences to the developing brain. These changes, coexisting in the nonepileptic brain, can overlap in a maladaptive combination to result in the diseased state of epilepsy. The consequence of early seizures in immature animals is a function of both the developmental stage and the method of seizure induction. PMID- 12121296 TI - Dynamic changes of status epilepticus-induced neuronal degeneration in the mediodorsal nucleus of the thalamus during postnatal development of the rat. AB - PURPOSE: Status epilepticus (SE) was previously found to induce damage in the mediodorsal nucleus of the thalamus (MD) in both adult and immature rats. This study was designed to describe age-related changes of SE-induced neuronal degeneration in this part of the brain. METHODS: SE was induced by LiCl/pilocarpine in five age groups of rats (P12-P25). Distribution of degenerating neurons was studied at various time intervals from 4 h up to 1 week using Fluoro Jade B (FJB) staining. For P12 and P25 rats, an interval of 3 months was added. RESULTS: Damaged neurons were found in all age groups during a 1-week period after SE. Patterns of neuronal degeneration, however, changed in an age related manner. In animals at P12 and P15, FJB-labeled neurons were located in the central and lateral segment of the MD. In the P18 group, degenerating neurons occurred in all three segments of the MD, with a prevalence in central and lateral subdivisions. In contrast, in P21 and P25 rats, FJB-labeled neurons were predominantly located in the central and medial segments. Degenerating neurons were still present 3 months after SE in the medial segment in P25 animals, whereas no labeled neurons were detected in the P12 group at this time. CONCLUSIONS: Our data demonstrate that the pattern of neuronal degeneration in MD is mainly related to age at SE onset. In addition to damage occurring during the acute phase of SE, a population of degenerating neurons was detected in P25 animals during the chronic period 3 months after SE. PMID- 12121297 TI - Interaction of excitatory amino acid agonists with cortical afterdischarges in developing rats. AB - PURPOSE: To determine the role of excitatory amino acids (EAAs) in genesis of two types of epileptic afterdischarges. METHODS: Cortical stimulation and recording electrodes were implanted in 12-, 18-, and 25-day-old rats. Epileptic afterdischarges were induced by rhythmic stimulation of sensorimotor cortex. The stimulation was repeated 6 times with 20-min intervals. Ten minutes after the first afterdischarge, N-methyl-d-aspartate, homocysteine, or kainic acid was injected. The doses were chosen individually for different age groups to be subconvulsive. Type and duration of afterdischarges as well as type and severity of motor correlates were evaluated. RESULTS: N-methyl-d-aspartate prolonged afterdischarges only in 12-day-old rats, whereas two other drugs did it in all age groups. Motor correlates of afterdischarges were changed to flexion seizures in 12-day-old rats after N-methyl-d-aspartate and homocysteine; in 25-day-old rats homocysteine led to generalized tonic-clonic seizures (i.e., both patterns seen after substantially higher doses of these drugs in nonstimulated rats). Seizures lasted tens of minutes. Kainic acid did not change the motor pattern in any age group, but nonconvulsive EEG seizures were recorded in the interstimulation periods mainly in 18- and 25-day-old rats. Increased transition into the limbic type of afterdischarges appeared only after homocysteine in 18- and 25-day-old rats. CONCLUSIONS: A mutual potentiation of epileptic phenomena was induced by two agents. The actions of N-methyl-d-aspartate and kainic acid differ in all age groups; the effects of homocysteine were identical with those of N-methyl-d-aspartate in 12-day-old rats but not later. Only homocysteine augmented transition into the limbic type of afterdischarges. PMID- 12121298 TI - Seizures decrease postnatal neurogenesis and granule cell development in the human fascia dentata. AB - PURPOSE: There is considerable controversy whether childhood seizures damage existing neurons and/or adversely affect neurogenesis and synaptogenesis. This study addressed this question by examining fascia dentata neurogenesis, cell death, and aberrant axon connections in hippocampi from children with extratemporal seizure foci. METHODS: Surgically resected (n = 53) and age comparable autopsy (n = 22) hippocampi were studied for neuronal densities, polysialic acid (PSA) neural cell adhesion molecule (NCAM) immunoreactivity (IR), TUNEL, and neo-Timm's histochemistry. RESULTS: Compared with autopsy cases, hippocampi from children with frequent seizures showed (a) decreased fascia dentata granule cell densities; (b) decreased PSA NCAM IR cell densities in the stratum granulosum, infragranular, and hilar regions; (c) no positive TUNEL stained cells; and (d) aberrant supragranular mossy fiber axon connections. CONCLUSIONS: These results indicate that severe seizures during early childhood are associated with anatomic signs of decreased postnatal granule cell neurogenesis (PSA NCAM IR) and aberrant mossy fiber axon connections (neo-Timm's) without evidence of seizure-induced cell death (TUNEL). In humans, these results support the concept that seizures do not damage existing neurons, but adversely affect processes involved with normal postnatal neuronal development such as neurogenesis and axon formation. Such alterations probably negatively affect normal brain development, and/or promote epileptogenesis. PMID- 12121299 TI - Epileptogenesis after self-sustaining status epilepticus. AB - PURPOSE: To describe the natural history of chronic epilepsy after experimental self-sustaining status epilepticus (SSSE) and to correlate patterns of SSSE with ictal, interictal, and plastic changes that characterize chronic epilepsy. METHODS: SSSE was induced in adult Wistar rats by 30-min intermittent electrical stimulation of the perforant path. In some animals, SSSE was treated by short term administration of antiepileptic drugs (AEDs). After SSSE, EEG and animal behavior were monitored for or =1 year in the rat. The silence is only behavioral, because EEG paroxysmal activity is seen in every animal. In this model of SSSE, the timing of treatment is a major determinant of outcome. Early treatment reduces the incidence of chronic epilepsy, whereas late treatment only reduces its severity. The possibility that this reduction of the severity of epilepsy may led to spontaneous remissions merits further study. PMID- 12121300 TI - Rate of interictal events and spontaneous seizures in epileptic rats after electrical stimulation of hippocampus and its afferents. AB - PURPOSE: Deep brain stimulation has been used by several investigators to prevent the occurrence of spontaneous seizures in patients with intractable epilepsy. With the kainic acid rat model of spontaneous recurrent seizures, we examined the consequences of subthreshold electrical stimulation of commissural pathways and perforant path on the synaptic plasticity, rate of interictal epileptiform events (IIEs), and spontaneous seizures in kainic acid (KA)-treated rats epileptic rats. METHODS: Recording microelectrodes were implanted bilaterally in the dentate gyrus and entorhinal cortex. Stimulating electrodes were implanted into the rostral part of the right hippocampus and right perforant path. Stimulation was performed daily with 200 Hz/0.5-s duration trains, 10-min duration 1-Hz train, or 2-h, 1-Hz train and 2-h duration, 50-Hz trains. Integrated amplitude of evoked potentials, rate of IIE, and spontaneous seizures was analyzed before and after stimulation. RESULTS: High-frequency tetani evoked long-term potentiation in 50% of epileptic rats compared with 100% of control rats. No long-term depression was observed after 1-Hz train. Decrease rate of IIE was found during 1-Hz and 50-Hz stimulation and returned to the basal level within 30-60 min. No significant change of the spontaneous seizure rate was found. CONCLUSIONS: Epileptic brain is less prone to plastic changes compared with the normal brain. Daily 2-h electrical stimulation with either low or high frequency does not have a long term effect on the rate of interictal events and spontaneous seizures. PMID- 12121301 TI - Delayed sclerosis, neuroprotection, and limbic epileptogenesis after status epilepticus in the rat. AB - PURPOSE: Hippocampal sclerosis and massive neurodegeneration in other parts of the limbic system are considered hallmarks of temporal lobe epilepsy. Using the rat model of kainate-induced status epilepticus, we sought to determine if limbic sclerosis after an excitotoxic insult follows a delayed type of neurodegeneration and is thus accessible to neuroprotective intervention after the insult. Effective pharmacologic neuroprotection after status epilepticus also addresses the old question of whether degenerative morphologic changes after an epilepsy inducing event like status epilepticus are the primary cause of epileptogenesis (i.e., the development of recurrent spontaneous seizures) during the following weeks. METHODS: Female Wistar rats after 90 min of generalized status epilepticus were used. Molecular biologic and histologic techniques were used to demonstrate markers of delayed cell death (apoptosis) 48 h after the status. The neuroprotective effects of i.c.v. injections of caspase inhibitors and systemic injections of the anticonvulsant drugs (AEDs) dizocilpine and retigabine after the status epilepticus were studied. The effect of neuroprotective intervention on the development of recurrent spontaneous seizures was investigated by behavioral observation of the rats. RESULTS: After generalized status epilepticus in Wistar rats, massive sclerosis of the hippocampus and the piriform cortex occurred. TUNEL labeling and electron microscopy revealed that apoptosis is involved in the degenerative processes. Immunohistochemical analysis of the time course of the expression of the proapoptotic protein Bax suggested a maximal induction of apoptosis 24-48 h after the status. Application of caspase inhibitors before or after the status did not reduce lesions, although Bax labeling was reduced. Injection of dizocilpine and to a lower extent also of retigabine after the status prevented limbic neurodegeneration and expression of markers of apoptosis. However, the neuroprotection by dizocilpine did not prevent the development of recurrent spontaneous seizures. CONCLUSIONS: Prolonged seizure activity can induce delayed sclerosis in the hippocampus and other parts of the limbic system. This delayed cell loss can be prevented by neuroprotective drugs after a status epilepticus. However, the damage in limbic brain regions is not the main reason for limbic epileptogenesis and the occurrence of recurrent spontaneous seizures. PMID- 12121302 TI - Damage, reorganization, and abnormal neocortical hyperexcitability in the pilocarpine model of temporal lobe epilepsy. AB - PURPOSE: Clinical, neuropathological, and electrophysiological data have shown that limbic structures are involved in the pathogenesis of temporal lobe epilepsy (TLE). In most cases, limbic-originated seizures frequently spread to extrahippocampal areas. It is unclear whether such distant circuitries, especially the neocortex, exhibit abnormal electrophysiology as consequences of a chronic epileptogenic process. The present research studied neuropathological abnormalities and in vitro electrophysiological properties of sensorimotor neocortex in pilocarpine-treated epileptic rats. METHODS: Adult epileptic animals showing six to seven seizures/week and saline-injected rats were selected for neurohistology. Coronal sections were sampled throughout the anteroposterior extent of the diencephalon and stained with cresyl violet (Nissl). Immunocytochemistry (ICC) was performed using anti-neurofilament (SMI-311) antibody. Extracellular (layer II/III) and intracellular (layer V) recordings were performed in coronal sensorimotor neocortical slices. Several electrophysiological aspects were examined such as evoked responses, intrinsic properties, and firing patterns of layer V pyramidal cells. RESULTS: Nissl staining showed a significant decrease of cortical thickness in epileptic rats when compared with controls, particularly in superficial layers (II-IV). Such abnormalities were also revealed by SMI-311 staining. SMI-311-labeled dendrite arborizations were more complex in layers I-II of epileptic rats. Epileptic rats manifested several abnormalities in extracellular field responses including hyperresponsiveness and presence of alpha-amino-3-hydroxy-5-methyl-4 isoxazolepropionate (AMPA)-mediated polysynaptic activity. Although no significant changes were observed concerning passive intrinsic properties, it was possible to detect a higher proportion of bursting neurons distributed in layer V (60%) of epileptic rats compared with 22% in control slices. CONCLUSIONS: Taken together, our findings indicate damage, reorganization, and chronic hyperexcitability of sensorimotor neocortex in experimental TLE. PMID- 12121303 TI - Characterization of reactive astrocytes in the chronic phase of the pilocarpine model of epilepsy. AB - PURPOSE: Astrogliosis is a prominent finding in human temporal lobe epilepsy. Work in animal models of temporal lobe epilepsy, however, have mostly concentrated on the acute phases of the studied models or have failed to demonstrate reactive gliosis during the chronic phases of such models. METHODS: Here we used the pilocarpine model of chronic seizures and Cajal's gold sublimate technique for the staining reactive astrocytes to study this issue. RESULTS: For half of the animals (nine of 17) subject to pilocarpine-induced status epilepticus (SE), when assessed 60 days later, variable levels of reactive astrocytes were seen in many thalamic, hippocampal, amygdalar, and neocortical areas. For the remaining half of the animals, however (eight of 17), despite a similar SE induction, as well as for controls, we could not detect stained reactive astrocytes. CONCLUSIONS: We hypothesize that these results might underlie possible differences in the frequency of recurrent spontaneous seizures. PMID- 12121304 TI - Effect on epileptogenesis of carbamazepine treatment during the silent period of the pilocarpine model of epilepsy. AB - PURPOSE: This study addresses the question of epileptogenesis by investigating the effects of carbamazepine (CBZ) on the silent period of the pilocarpine model of epilepsy. METHODS: Adult Wistar rats were subjected to status epilepticus (SE) induced by pilocarpine and treated with CBZ or saline, i.p, during 56 days. Latency for the first spontaneous seizure, incidence, frequency, and duration of seizures were monitored for < or =56 days after treatment. Hippocampal histologic analysis was performed. RESULTS: CBZ reduced the frequency and duration of the seizures and the hippocampal damage. CONCLUSIONS: CBZ did not abort the epileptogenesis but minimized the expression of seizures and hippocampal damage. PMID- 12121305 TI - How mutations in the nAChRs can cause ADNFLE epilepsy. AB - PURPOSE: The linkage between autosomal dominant nocturnal frontal lobe epilepsy (ADNFLE) and neuronal nicotinic acetylcholine receptor has been strongly reinforced by the report of five distinct mutations in the two genes coding for the major brain alpha4beta2 nicotinic acetylcholine (ACh) receptors. As a first step toward understanding the basic mechanisms underlying this genetically transmissible neurologic disorder, we examined the similarities and differences of the functional properties displayed by naturally occurring mutant forms of this ligand-gated channel. METHODS: Functional studies of neuronal nicotinic ACh receptors reconstituted in Xenopus oocytes were designed to analyze the common traits displayed by the different mutations associated with ADNFLE. RESULTS: Coexpression of the control and mutated alleles harboring the alpha4S248F mutation obtained from patient DNAs yielded ACh-evoked currents of amplitude comparable to the control responses but with a higher sensitivity and desensitization to the natural agonist. Alternatively, the other mutants (alpha4L776ins3, alpha4S252L, and beta2V287M) displayed an increased ACh sensitivity without pronounced desensitization. In addition, whereas a reduction of calcium permeability was observed for the mutants (alpha4S248F and alpha4L776ins3), no significant modification of ionic selectivity could be detected in the alpha4S252L mutation. Hence increase in ACh sensitivity is the only common characteristic so far observed between the four naturally occurring mutant receptors investigated. CONCLUSIONS: Analyses of functional properties of four nAChR mutants associated with ADNFLE indicate that a gain of function of these mutant receptors may be at the origin of the neuronal network dysfunction that causes the epileptic seizures. These data are discussed in the context of our latest knowledge of the pyramidal cell function. PMID- 12121306 TI - The genetic absence epilepsy rat from Strasbourg (GAERS), a rat model of absence epilepsy: computer modeling and differential gene expression. AB - PURPOSE: We present results obtained by computer modeling of the thalamic network and differential gene expression analysis in a rat strain with absence epilepsy, the genetic absence epilepsy rat from Strasbourg (GAERS). METHODS: (a) Computer modeling used equations from the Hodgking-Huxley model with a circuit of 13 reticular thalamic (nRt) and 39 thalamocortical (TC) neurons; (b) gene-expression analysis using differential mRNA display (DD), in situ hybridization, Northern blotting, and competitive reverse transcriptase-polymerase chain reaction (RT PCR). RESULTS: (a) Computer modeling showed an increased network synchrony in the thalamic circuit as the value of conductance of low-voltage activated calcium channel (LVACC) is increased. (b) Using differential mRNA display, a 40% upregulation of the H-ferritin mRNA in the hippocampus was demonstrated. Looking for some candidate genes of the VACC family, no difference was found in the alpha1G mRNA expression between GAERS and control animals, whereas a decreased expression of the alpha1E subunit was observed in the cerebellum and the brainstem of the GAERS. This phenomenon was not observed in young animals when the epileptic phenotype is not expressed. CONCLUSIONS: The use of computer modeling appeared to be an efficient way to evaluate the impacts of electrophysiologic findings in vivo from single cells on an entire circuit. No clear single gene defect was revealed so far in GAERS. More information could arise from linkage analysis. However, some brain structures like hippocampus or cerebellum classically not known to be involved in the production of absence spike-and-wave discharges could in fact participate in the development of this phenotype. PMID- 12121307 TI - Ictogenesis and epileptogenesis in EL mice. AB - PURPOSE: In EL mice, ictogenesis is established at age approximately 10 weeks, whereas epileptogenesis is induced through an experience of repetitive seizures during development. An "abnormal neural plasticity" has been suggested to be involved in these pathologic processes. It also is known that two isoforms of nitric oxide (NO) synthetase (nNOS and eNOS) are essential for the long-term potentiation (LTP), a plastic response of neurons. It appears, therefore, that these NO synthetases might play a major role in the establishment of abnormal neural plasticity. The purpose of the present study was to investigate ictogenesis and epileptogenesis by observing alterations of NO synthetases as well as immediate early gene (IEG) expressions and gamma-aminobutyric acid (GABA) distributions in the brain during development and with respect to seizure history. METHODS: IEG (c-fos and zif) expression in EL mice were analyzed by in situ hybridization with 35S. Distribution of GABA concentrations and glutamic acid decarboxylase (GAD) activities in the parietal cortex of EL mice was quantitatively determined using ultramicroenzymatic chemistry (Lowry, 1978). Three isoforms of NOS were assayed by immunoblotting analysis for hippocampal tissues of EL mice and their control animals, DDY mice. DNA fragmentation was detected with the TUNEL method. RESULTS: In EL mouse brains, IEG expression was related to the seizure history, seizure threshold, and age. Even in the interictal period, the animals expressed IEG continuously when their seizure thresholds were very low. Among various IEG expression sites in the brain, hippocampal CA1 was the most remarkable. These IEG expression sites were almost identical to the brain regions of EL mice where GABA concentrations and GAD activities were altered. Unexpectedly, the eNOS content of EL was very small, although eNOS appears to be responsible for NO that mediates an increase in local cerebral blood flow during focal seizures. nNOS, iNOS, and to a lesser extent, eNOS were essential to establish both ictogenicity and epileptogenicity. DNA fragmentation was observed in the hippocampus of EL mice in the interictal period. CONCLUSIONS: Continuous IEG expression and abnormal GABAergic function are involved in the epileptogenesis of EL mice. Transiently expressed IEG, on the other hand, is associated with the ictogenesis. It is conceivable that an excess amount of iNOS (and subsequent increase in harmful antimicrobial NO) and a lesser amount of eNOS (and subsequent decrease in NO or endothelium-derived relaxing factor, EDRF) may work together to contribute to a focus complex and ictogenesis. Drugs that suppress iNOS and/or potentiate eNOS may be promising candidates for a new type of antiepileptic agent. PMID- 12121308 TI - Clinical and genetic heterogeneity in familial temporal lobe epilepsy. AB - PURPOSE: Familial forms of temporal lobe epilepsy have been described recently. A locus on ch 10q has been linked to partial epilepsy with auditory symptoms. We investigated the proportion of families segregating temporal lobe epilepsy (TLE) linked to ch 10q and sought to establish genotype-phenotype correlations. METHODS: We studied 15 unrelated families segregating TLE. A total of 153 individuals, including 79 patients, were analyzed in this study. Family members were genotyped for four polymorphic dinucleotide repeat markers: D10S185, D10S574, D10S577, and D10S192, which flank the 15-cM candidate interval on ch 10q. Two-point lod scores were calculated for each family separately. RESULTS: Fourteen of our families had ictal semiology of mesial temporal onset of seizures and magnetic resonance imaging (MRI) abnormalities in the mesial structures; only one family, with seven affected individuals, reported auditory symptoms and had normal MRIs. Pedigree analysis showed an autosomal dominant transmission with 0.75 penetrance. Only two families had informative lod scores. A large family, with 22 affected individuals segregating mesial TLE, had negative lod scores for all four markers genotyped. The lod scores were significantly negative (less than -2.00) up to 0.05 for D10S185, 0.10 for D10S574, 0.25 for D10S577, and 0.15 for D10S192. The single family with auditory symptoms had positive lod scores for all markers genotyped, with a Z max of 1.52 at 0.0 for D10S574. CONCLUSIONS: We identified two different clinical groups of families segregating TLE. Most families identified in this study had mesial TLE. Only one single family segregating lateral TLE was found. We significantly excluded linkage between familial mesial TLE and the locus on ch 10q. In addition, we showed evidence for linkage between one family with lateral TLE and markers on ch 10q. This is strong evidence for clinical and genetic heterogeneity among familial forms of TLE. PMID- 12121309 TI - Upregulation of cystatin C expression in the rat hippocampus during epileptogenesis in the amygdala stimulation model of temporal lobe epilepsy. AB - PURPOSE: Cystatin C is a cysteine proteinase inhibitor with widespread distribution in body fluids and tissues, abundant in the cerebrospinal fluid and in brain tissue. There is an implied role for cystatin C in several neurologic disorders, but the actual function of cystatin C in the brain remains unknown. Moreover, the reports on the distribution of cystatin C in the brain are controversial. We present the data on the distribution of cystatin C in normal brain tissue and during epileptogenesis. METHODS: Epileptogenesis was triggered by inducing self-sustained status epilepticus (SSSE) with a 20- to 30-min electrical stimulation of the amygdala in rats. Animals were monitored continuously for 2 weeks with video-EEG to ascertain that they were in an epileptogenic phase. RESULTS: Analysis of double-stained immunopreparations indicated that in normal brain, cystatin C is expressed mainly in microglia. In epileptogenic animals, immunostaining was increased in the microglia as well as in the neuropil at 4 days, 1 week, and 2 weeks after SSSE. Moreover, the density of cystatin C-positive microglia was associated with the severity of neuronal damage in the CA1 subfield of the hippocampus. CONCLUSIONS: This is the first report linking cystatin C with epileptogenesis and epilepsy. Further studies will explore the potential neuroprotective functions of this protein during epileptogenesis and whether the manipulation of its expression or function will have therapeutic implications. PMID- 12121310 TI - Estrogen treatment protects GABA(B) inhibition in the dentate gyrus of female rats after kainic acid-induced status epilepticus. AB - PURPOSE: We used the paired-pulse inhibition paradigm to determine whether the cell loss in the hilus of the dentate gyrus of female rats after kainic acid (KA) induced status epilepticus (SE) is associated with functional changes in the dentate gyrus. Additionally, we determined whether the lost function could be preserved by using estrogen neuroprotection. METHODS: Female rats were ovariectomized and treated either with estrogen replacement (four doses of 2 microg of estradiol every 24 h: two doses before SE, two doses after) or oil. SE was induced by I.P. administration of KA (16 mg/kg) and terminated after 5 h with pentobarbital. After 2 days, hippocampal/dentate gyrus slices were prepared. Population spikes were recorded in the granule cell layer as a response to mixed perforant-path stimulation (10- to 1,000-ms interstimulus intervals). Ratios of the test response to conditioning response were evaluated. Gamma-aminobutyric acid type B (GABAB) receptors were blocked with 400 microM CGP 35348. RESULTS: In slices from oil-treated rats, SE induced a loss of paired-pulse inhibition in the dentate gyrus at the interstimulus intervals marking intermediate facilitation and late depression. There was no such loss of paired-pulse inhibition in estrogen-treated rats. CGP 35348 was unable to alter paired-pulse inhibition in slices form oil-treated rats. In slices from estrogen-treated rats, CGP decreased paired-pulse inhibition at 50-150-ms interstimulus intervals. Comparison of paired-pulse inhibition in slices from oil-treated rats with slices from estrogen treated rats with CGP 35348 revealed a GABAB-independent difference at interstimulus intervals >300 ms. CONCLUSIONS: Our study demonstrated that there is a complete loss of GABAB receptor-mediated inhibition after KA-induced SE in the dentate gyrus. Pretreatment with estrogen can save GABAB-receptor function, probably by neuroprotection of neurons containing the postsynaptic GABAB receptors. PMID- 12121311 TI - Altered expression of neuropeptide Y, -Y(1), and -Y(2) receptors in the hippocampus of patients with mesial temporal lobe epilepsy. PMID- 12121312 TI - Dendritic targeting of mRNAs for plasticity genes in experimental models of temporal lobe epilepsy. AB - PURPOSE: To analyze whether the subcellular localization of the messenger RNAs (mRNAs) coding for the neurotrophin brain-derived neurotrophic factor (BDNF), its receptor TrkB, and the alpha and beta subunits of calcium-calmodulin-dependent kinase II (CaMKII) are modified after pilocarpine and kindled seizures. METHODS: Epilepsy models: pilocarpine and kindling. Analysis of mRNA levels in the dendrites: high-resolution, nonradioactive in situ hybridization. RESULTS: Nonstimulated rats: BDNF, TrkB, and CaMKII-beta mRNAs localized in the soma and in the proximal dendrites of hippocampal pyramidal cells, and in the soma only of dentate gyrus (DG) granule cells; CaMKII-alpha mRNA localized throughout the dendritic length in neurons of all hippocampal subfields. Pilocarpine seizures: increased staining levels of CaMKII-alpha mRNA throughout the whole dendritic length in all hippocampal subfields; induction of CaMKII-beta, BDNF, and TrkB mRNAs dendritic targeting in CA1, CA3, and DG neurons. Class 2 kindled seizures: increase in dendritic staining intensity for CaMKII-alpha in CA1, CA3, and DG neurons; induction of dendritic localization of CaMKII-beta, BDNF, and TrkB mRNAs in CA3 neurons. Fully kindled seizures: no change in the subcellular distribution of BDNF, TrkB and CaMKII-beta mRNAs; reduction of CaMKII-alpha mRNA dendritic staining, as compared with unstimulated kindled animals. CONCLUSIONS: Data provide evidence that BDNF, TrkB, and CaMKII-alpha and -beta mRNAs are accumulated in the dendrites of specific hippocampal neurons during pilocarpine seizures and kindling development. The dendritic targeting of these genes may be causally involved in epileptogenesis and thus may represent a new therapeutic target for some forms of partial epilepsy. PMID- 12121313 TI - Extracellular matrix components are altered in the hippocampus, cortex, and cerebrospinal fluid of patients with mesial temporal lobe epilepsy. AB - PURPOSE: This work studied the profile of glycosaminoglycans (GAGs) in the hippocampus, cortex, and cerebrospinal fluid of patients with temporal lobe epilepsy (TLE). METHODS: The GAGs were analyzed by agarose gel electrophoresis, enzymatic degradation, and enzyme-linked immunosorbent assay (ELISA). RESULTS: The hippocampus of TLE patients showed increased levels of chondroitin sulfate and hyaluronic acid against normal levels of these GAGs in the neocortex and cerebrospinal fluid (CSF). CONCLUSIONS: These results suggest that these matrix components could be involved in the pathophysiology of TLE. PMID- 12121314 TI - Changes in flip/flop splicing of astroglial AMPA receptors in human temporal lobe epilepsy. AB - PURPOSE: Recent data suggested a role for glial cells in epilepsy. This study sought to identify and functionally characterize AMPA receptors expressed by astrocytes in human hippocampal tissue resected from patients with intractable temporal lobe epilepsy. METHODS: Patch-clamp and fast application methods were combined to investigate astrocytes in situ and after fresh isolation from the stratum radiatum of the hippocampal CA1 subfield. Relying on presurgical and histopathologic analysis, we divided human specimens into two groups, Ammon's horn sclerosis (AHS) and lesion-associated epilepsy. RESULTS: Fast application of glutamate and kainate evoked receptor currents in all cells studied. Reversal potential analysis revealed an intermediate Ca2+ permeability of the receptor channels that did not vary between the two groups of patients. However, preapplication of the AMPA receptor-specific modulator, cyclothiazide, disclosed differences in flip-flop splicing. This treatment considerably enhanced the receptor conductance, with potentiation being significantly stronger in cells from AHS specimens compared with lesion-associated cells, suggesting upregulation of AMPA receptor flip splice variants in astrocytes of the sclerotic tissue. CONCLUSIONS: Compelling evidence has been accumulated showing direct and rapid signaling between neurons and glial cells. Our data suggest that in AHS patients, neuronally released glutamate will lead to an enhanced and prolonged depolarization of astrocytes, which might be involved in seizure generation and spread in this particular condition of human temporal lobe epilepsy. PMID- 12121315 TI - Coupling of electrical and metabolic activity during epileptiform discharges. AB - Changes in electrical activity, ionic microenvironments, and intracellular Ca concentration were measured during recurrent seizures induced by low Mg in slices and slice cultures. In both preparations, initial seizure-like events (SLEs) changed after some time into drug-refractory late recurrent discharges. In slice cultures, there was considerable cell loss in all hippocampal areas after 2 h of status epilepticus. During recurrent SLEs, the NAD(P)H autofluorescence declined, as did intramitochondrial calcium signals, indicating mitochondrial damage. At the same time, ethidium signals indicated increased radical oxygen species production. These alterations could be reduced by alpha-tocopherol, which also protected slice cultures against status epilepticus-induced cell death. PMID- 12121316 TI - Glutamatergic modulation of GABAergic signaling among hippocampal interneurons: novel mechanisms regulating hippocampal excitability. AB - PURPOSE: Because interneurons play a central role in regulating the excitability of the hippocampal formation, it is important to understand the mechanisms that modulate gamma-aminobutyric acid (GABA)ergic signaling among them. This study addresses the modulation of GABA release from interneuron terminals by presynaptic glutamate receptors. METHODS: Whole-cell recordings were obtained from CA1 stratum radiatum interneurons in guinea pig hippocampal slices. Selective agonists and blockers of glutamate receptors were used to study modulation of GABAergic transmission by group III metabotropic receptors or kainate receptors. Antidromic action-potential initiation also was analyzed by stimulating the axons of interneurons. RESULTS: Agonists of group III metabotropic glutamate receptors attenuated monosynaptic GABAergic signals in interneurons, but not in pyramidal neurons, in agreement with anatomic evidence on the distribution of these receptors. Submicromolar kainate enhanced the frequency and amplitude of spontaneous GABAergic signals in interneurons. Kainate also depolarized the axons of hippocampal interneurons, and triggered spontaneous ectopic action potentials in axons. Synaptically released glutamate reproduced many of the effects of both agonists, implying that these receptors can sense the ambient glutamate concentration, and therefore indirectly respond to the excitatory traffic in the hippocampus. When the two classes of receptors were stimulated simultaneously, complex interactions were obtained. CONCLUSIONS: Group III metabotropic receptors and kainate receptors profoundly affect GABAergic signaling among interneurons of the hippocampus. Glutamatergic modulation of GABAergic signaling among interneurons represents a novel class of mechanisms that potentially plays a major role in determining the initiation, propagation, and termination of seizures. PMID- 12121317 TI - Determinants of ictal epileptiform patterns in the hippocampal slice. AB - PURPOSE: The transition from an interictal to an ictal pattern of epileptiform activity is a strategic target for antiepileptic drug (AED) action. Both the muscarinic agonist pilocarpine and the selective group I metabotropic glutamate receptor (mGluR) agonist (RS)-3,5-dihydroxyphenylglycine (DHPG) produce prolonged synchronous activity in the hippocampal slice that resembles ictal discharges. We evaluated the role of synaptic mechanisms and release of calcium from intracellular stores in the generation of prolonged ictal oscillations. METHODS: Pilocarpine (10 microM) in 7.5 mM[K+]o or DHPG (100 microM) in 5 mM[K+]o artificial cerebrospinal fluid (ACSF) were bath applied to hippocampal slices, and extracellular recordings were made from the CA3 region. The pattern of activity was characterized as ictal if prolonged oscillations of discharges occurred at >2 Hz lasting for >3 s. The pattern of epileptiform activity was characterized and compared with the pattern observed after bath application of pharmacologic agents. RESULTS: The AMPA/kainic acid (KA) glutamate receptor blocker DNQX (20 microM) dampened and stopped ictal oscillations; however, antagonism of N-methyl-d-aspartate (NMDA) or gamma-aminobutyric acid (GABAA) receptors had minimal effects on ictal patterns. Ictal discharges were suppressed by dantrolene (30-100 microM), which blocks release of calcium from intracellular stores, or thapsigargin (1-5 microM), which inhibits the adenosine triphosphatase (ATPase) that maintains intracellular calcium stores. The L-type calcium channel antagonist nifedipine (1 microM) blocked ictal activity produced by pilocarpine or DHPG. CONCLUSIONS: Ictal discharges produced by pilocarpine or DHPG depended on intact synaptic transmission mediated by AMPA/KA receptors, release of calcium from intracellular stores, and L-type calcium channel activation. The results suggest that muscarinic and group I mGluRs activate a positive-feedback system that creates calcium oscillations and prolonged neuronal synchronization mediated by recurrent excitatory synaptic connections in the CA3 region of the hippocampus. PMID- 12121318 TI - Veratridine, but not elevated K+, inhibits excitatory amino acid transporter activity in rat hippocampal slices. AB - PURPOSE: Excitatory amino acid transporter (EAAT) activity prevents Glu from reaching toxic levels, but their contribution to epileptogenesis remains controversial. We examined how the convulsant veratridine causes inhibition of EAAT activity and how it differs from the effects of another convulsant, high (50 mM) K+, that also increases Na+ conductance. METHODS: Transverse rat hippocampal slices were incubated for 1 h with 100 microM veratridine in oxygenated artificial cerebrospinal fluid (aCSF) with or without extracellular Ca2+. The medium was replaced by 50 microM[(3)H]glutamate in aCSF, and the slices incubated for 10 min at 37 degrees C. The slices were washed 3 times with cold aCSF after removal of the extracellular medium, and the radioactivity was quantified after solubilization of the slices. RESULTS: Veratridine caused a time- and dose dependent decrease, whereas high K+ had no effect on EAAT activity. The effects of veratridine on EAAT activity were not prevented by tetrodotoxin (TTX; 10 microM). Coincubation of ouabain with veratridine resulted in further decreases of EAAT activity. Removal of extracellular Ca2+ potentiated the inhibitory effects of veratridine (and other convulsants) on EAAT activity. Chelation of intracellular Ca2+ with BAPTA also increased the inhibitory effects of veratridine on EAAT activity. CONCLUSIONS: Veratridine caused changes Ca2+ dynamics that led to inhibition of EAAT activity. Such changes in EAAT activity can contribute to the sustained epileptiform activity caused by veratridine. PMID- 12121319 TI - Changes in nonlinear signal processing in rat hippocampus associated with loss of paired-pulse inhibition or epileptogenesis. AB - PURPOSE: To study acute and chronic physiological effects of perforant path stimulation using paired-pulse and nonlinear signal analysis techniques (Wiener kernel analysis). METHODS: Two to 3-month-old Wistar rats were implanted with stimulating electrodes in the perforant path and recording electrodes in the granule cell layer. Loss of paired-pulse inhibition was produced with 2 Hz continuous and 20 Hz (10 s/min) intermittent stimulation for periods of 1-15 min (0.1 ms, 20 v pulses). Some animals received 30-60 min of stimulation, a model for status epilepticus/epileptogenesis. Responses to paired-pulse or white noise inputs were recorded sequentially. RESULTS: Loss of inhibition with brief 1-3 min of stimulation, measured by increase paired-pulse ratio (P2/P1 ISI 40 ms) from 0.25 (+/-0.27) pre- to 1.02 (+/-0.18) post-stimulation (p < 0.001), lasted 43 (+/ 15) min. For 30-60 min of stimulation, the paired-pulse ratios were 0.088 (+/ 0.11), 1.59 (+/-0.036), 0.06 (+/-0.11), 0.82 (+/-0.22) for pre-, immediate post-, 1 week post-, and 1 month poststimulation, respectively (p < 0.025). Compared to prestimulation values, Wiener kernel amplitudes for immediate, 1 week, and 1 month poststimulation were 24% (+/-13%), 72% (+/-17%), and 31% (+/-21%), respectively (p < 0.05). Wiener kernels 1 month poststimulation showed response prolongation with increased opportunity for excitatory interactions of inputs (particularly those separated by 4 ms). CONCLUSIONS: Brief perforant path stimulation causes sustained loss of inhibition in the dentate, possibly an early event in the transition to status epilepticus. Stimulation for 30-60 min causes chronic changes in paired-pulse and white noise (Wiener kernel) responses. Transient recovery occurs by 1 week, but later new features appear (including delayed/late inhibition and potential excitatory cross-talk) that might favor epileptic seizures. PMID- 12121320 TI - Detecting epileptic spikes. AB - We present a technique for automatic detection of epileptic spikes in electroencephalogram (EEG) recordings. We used a nonlinear modeling method that enables us to detect rapidly and accurately epileptic behavior in the EEG signal. Our approach is illustrated by an application to interictal activity from a patient with focal epilepsy. PMID- 12121321 TI - Do mossy fibers release GABA? AB - PURPOSE: Mossy fibers are the sole excitatory projection from dentate gyrus granule cells to the hippocampus, forming part of the trisynaptic hippocampal circuit. They undergo significant plasticity during epileptogenesis and have been implicated in seizure generation. Mossy fibers are a highly unusual projection in the mammalian brain; in addition to glutamate, they release adenosine, dynorphin, zinc, and possibly other peptides. Mossy fiber terminals also show intense immunoreactivity for the inhibitory neurotransmitter gamma-aminobutyric acid (GABA), and immunoreactivity for GAD67. The purpose of this review is to present physiologic evidence of GABA release by mossy fibers and its modulation by epileptic activity. METHODS: We used hippocampal slices from 3- to 5-week-old guinea pigs and made whole-cell voltage clamp recordings from CA3 pyramidal cells. We placed stimulating electrodes in stratum granulosum and adjusted their position in order to recruit mossy fiber to CA3 projections. RESULTS: We have shown that electrical stimuli that recruit dentate granule cells elicit monosynaptic GABAA receptor-mediated synaptic signals in CA3 pyramidal neurons. These inhibitory signals satisfy the criteria that distinguish mossy fiber-CA3 synapses: high sensitivity to metabotropic glutamate-receptor agonists, facilitation during repetitive stimulation, and N-methyl-D-aspartate (NMDA) receptor-independent long-term potentiation. CONCLUSIONS: We have thus provided compelling evidence that there is a mossy fiber GABAergic signal. The physiologic role of this mossy fiber GABAergic signal is uncertain, but may be of developmental importance. Other evidence suggests that this GABAergic signal is transiently upregulated after seizures. This could have an inhibitory or disinhibitory effect, and further work is needed to elucidate its actual role. PMID- 12121322 TI - Electrophysiologic abnormalities of the hippocampus in the pilocarpine/cycloheximide model of chronic spontaneous seizures. AB - PURPOSE: Mossy fiber sprouting (MFS) and synaptic reorganization in the dentate gyrus (DG) is considered one of the physiopathologic mechanisms in temporal lobe epilepsy. Supragranular MFS can be blocked by cycloheximide (CHX) without interfering with the genesis of spontaneous recurrent seizures. The aim of this study was to investigate electrophysiologic properties of the hippocampus in the CHX/pilocarpine (CHX/PILO) model as compared with the conventional PILO model. METHODS: In vitro electrophysiology was performed 2 months after status epilepticus (SE) induction using extracellular recordings in hippocampal slices from PILO (n = 8) and CHX/PILO animals (n = 10). Field potential responses were evoked in the CA1 and DG regions during perfusion with normal artificial cerebrospinal fluid (aCSF) and aCSF containing 3.5, 5, or 8 mM K+ without or with bicuculline added. Neo-Timm staining was used for the assessment of supragranular MFS. RESULTS: Evoked potentials in PILO- and CHX/PILO-treated rats displayed small-amplitude polyspiking activity (epileptiform responses) in CA1 and an apparently normal isolated population spike in DG. More important, PILO and CHX/PILO animals did not differ regarding electrophysiologic abnormalities, even under high K+ or high K+/bicuculline. Analysis of the neo-Timm staining revealed strong supragranular MFS in PILO-injected rats and significantly less staining in CHX/PILO rats. Thus, occurrence of abnormal stimulus responses and high K+- or high K+/bicuculline-induced epileptiform activities did not depend on the degree of MFS. CONCLUSIONS: We therefore suggest that other mechanisms such as anomalous intrinsic bursting and disinhibition rather than MFS might account for the increased hippocampal hyperexcitability in this model. PMID- 12121323 TI - AlphaCaMKII and NMDA-receptor subunit expression in epileptogenic cortex from human periventricular nodular heterotopia. AB - PURPOSE: Periventricular nodular heterotopia (PNH) is the most common human brain dysgenesis, very frequently characterized by focal drug-resistant epilepsy. To understand the cellular mechanisms underlying its intrinsic hyperexcitability, we investigated the expression of glutamate-receptor subunits and related proteins in four human patients affected by PNH. METHODS: PNH was diagnosed by means of magnetic resonance imaging. The epileptogenic area was revealed by depth electrode recordings and removed during epilepsy surgery. Sections from the removed cerebral tissue were analyzed by means of immunocytochemistry (ICC), with antibodies directed against N-methyl-d-aspartate (NMDA)-receptor subunits, the alpha subunit of the Ca2+/calmodulin-dependent kinase II (alphaCaMKII), and its active phosphorylated form. RESULTS: The ICC data demonstrated that the subcortical heterotopic nodules were consistently characterized by lower expression of alphaCaMKII and its activated form. In more pronounced cases (i.e., when the extension of the nodules to the neocortex determined clear layering abnormalities), the heterotopic tissue also was characterized by a decreased expression of NMDA-receptor subunits, which was particularly evident in the dendritic compartment. CONCLUSIONS: These data suggest the existence of an alteration of alphaCaMKII and the NMDA-receptor complex in the epileptogenic brain tissue of human PNH, which may play a role in the basic mechanisms of hyperexcitability associated with this brain dysgenesis. PMID- 12121324 TI - Immunohistochemical study of six cases of Taylor's type focal cortical dysplasia: correlation with electroclinical data. AB - PURPOSE: Cortical specimens from six patients operated on for drug-resistant epilepsy diagnosed as Taylor's type focal cortical dysplasia were submitted to neuropathological and immunohistochemical studies. METHODS: All patients were submitted to presurgical investigations including clinical and neuropsychological evaluations, EEG/video telemetry of ictal and interictal events, magnetic resonance imaging, and ictal and interictal single-photon emission computed tomography (SPECT). Recordings from electrocorticography (ECoG) were obtained in four cases and from subdural electrode implantation in two. Postsurgical follow up was assessed according to Engel's score. Immunohistochemistry (IHC) was processed for parvalbumin (PV), calbindin D28-K (CB), nonphosphorylated neurofilaments (SMI-311), glial fibrillary acidic protein (GFAP) in all cases. RESULTS: We found continuous/quasi-continuous spikes and sharp-wave patterns in three cases and frequent repetitive bursting of polyspikes and ECoG seizures in two cases. Every patient showed cortical dyslamination, abnormal and giant neurons, and balloon cells. GFAP immunoreactivity was found in astrocytes and some balloon cells that were less intensely stained. Nonphosphorylated neurofilaments SMI-311 immunoreactivity was found in normal and giant neurons and in some balloon cells, making visible thin neuropils. PV immunoreactivity was present in normal interneurons and in fibers in layers IV-V. PV-negative balloon cells were surrounded by abundant PV-positive fibers. CB immunoreactivity was found mostly in interneurons in layers II-III. CONCLUSIONS: Our research is inconclusive. More cases should be investigated, and we must draw more accurate anatomic correlations between the ECoG recordings and surgical specimens studied with IHC. PMID- 12121325 TI - Neurotrophin receptor immunoreactivity in severe cerebral cortical dysplasia. AB - PURPOSE: Cerebral cortical dysplasia (CD) is one of the important causes of intractable epilepsies and characterized histologically by disorganized cortical lamination and cytomegalic dysplastic neurons. Although it has been suggested that neurotrophins play an important role in differentiation, growth, and survival of developmental neurons, their pathogenetic role in CD has rarely been investigated. METHODS: To know the pathogenetic role of various neurotrophins on dysplastic neurons, immunohistochemical staining was performed using antibodies against NGFRp75, trkA, trkB, and trkC in surgical specimens of 20 patients with CD. RESULTS: TrkB and trkC were strongly expressed in dysplastic neurons of severe CD, and NGFRp75 was also expressed in some dysplastic neurons. CONCLUSIONS: It is known that brain-derived neurotrophic factor (BDNF) and neurotrophin-3 (NT-3) contribute to the differentiation of neuronal precursor cells, dendritic and axonal arborization, synaptic plasticity, and cellular hyperexcitability, so increased expression of trkB and trkC may have a critical pathogenetic role in cytoskeletal abnormalities and epileptogenicity in dysplastic neurons of CD. PMID- 12121326 TI - Evidence that ATP participates in the pathophysiology of pilocarpine-induced temporal lobe epilepsy: fluorimetric, immunohistochemical, and Western blot studies. AB - PURPOSE: This study was performed to study the role of adenosine triphosphate (ATP) in the brain of pilocarpine-induced chronic epileptic rats. METHODS: ATP mediated changes in intracellular calcium were studied by the fura-2 method. Immunohistochemistry and Western blotting methods were used to localize and quantify P2X7 receptors in these animals. RESULTS: The fluorimetric study in chronic rats revealed a biphasic response indicating the presence of P2X7 receptors. The Western blotting study showed an increase of 80% of P2X7 expression in chronic rats compared with the control group. P2X7 immunoreactivity resembled mossy fiber sprouting at the dentate gyrus of epileptic animals. CONCLUSIONS: These results suggest that purinergic receptors may participate in the pathophysiology of temporal lobe epilepsy. PMID- 12121327 TI - Evaluation of opioid peptide and muscarinic receptors in human epileptogenic neocortex: an autoradiography study. AB - PURPOSE: The main goal of the present study was to evaluate possible alterations in opioid peptide and muscarinic receptors in human neocortical epileptic focus and the surrounding area removed from patients with pharmacologically resistant epilepsy and epilepsy secondary to cerebral lesion by tumor or other causes. METHODS: In vitro quantitative autoradiography experiments were carried out to label mu, delta, and muscarinic receptors of neocortical epileptic focus and surrounding area obtained from patients with pharmacologically resistant primary epilepsy and epilepsy caused by tumors and angioma cavernosa, and compared with neocortex obtained from patients with dementia and tumors without epilepsy. RESULTS: The mu receptor levels were lower in surrounding areas (-46%). The delta receptor binding was reduced in epileptic focus obtained from patients with epilepsy secondary to cerebral lesion (-25%) and surrounding areas (-31%). In contrast, muscarinic receptor levels were higher in the focus from patients with primary epilepsy (layers I-II, 52%; layers III-IV, 44%; layers V-VI, 36%). CONCLUSIONS: It is suggested that the increased muscarinic receptors in the epileptic focus and the decreased mu and delta receptors in the surrounding area are associated with the initiation and propagation of seizure activity in human epileptogenic neocortex. PMID- 12121328 TI - Loss and sprouting of nitric oxide synthase neurons in the human epileptic hippocampus. AB - PURPOSE: Nitric oxide (NO) has been implicated in a variety of functions, including the control of synaptic plasticity and sensory signaling. Current evidence suggests that this unconventional neurotransmitter mediates N-methyl-d aspartate (NMDA) receptor-linked excitotoxicity. This study describes the expression of neuronal NO synthase (nNOS) immunoreactivity (IR) in hippocampi from patients with temporal lobe epilepsy (TLE). METHODS: Hippocampi from patients with clinical symptoms, neuroimaging, and EEG typical of hippocampal sclerosis (HS; n = 22) were compared with those from patients with neocortical temporal lesions (NONHS; n = 4) and autopsy (AUT; n = 18) patients for total cells, and nNOS-IR neuron and puncta densities. RESULTS: Compared with AUT, HS hippocampi had significantly less nNOS-IR neuron densities in the fascia dentata (FD); hilus, and CA4, CA3, CA2, and CA1 subfields. HS hippocampi had significantly greater nNOS-IR puncta densities in the FD, as compared with AUT and NONHS. CONCLUSIONS: Our results show that hippocampi from TLE patients exhibit a loss of nNOS-IR neurons and an abnormal FD innervation. The release of NO can influence the dynamics of ionic channels and neurotransmitter release, thus affecting neuronal membrane potential. Because the NOergic transmission does not obey the topographic constraints imposed on conventional transmitters, target cells can be stimulated even in regions with severe deafferentation. The plastic changes described here may contribute to abnormal hippocampal excitability. PMID- 12121330 TI - The projected health care burden of Type 2 diabetes in the UK from 2000 to 2060. AB - AIMS/HYPOTHESIS: To predict the incidence and prevalence of Type 2 diabetes in the UK, the trends in the levels of diabetes-related complications, and the associated health care costs for the period 2000-60. METHODS: An established epidemiological and economic model of the long-term complications and health care costs of Type 2 diabetes was applied to UK population projections from 2000 to 2060. The model was used to calculate the incidence and prevalence of Type 2 diabetes, the caseloads and population burden for diabetes-related complications, and annual NHS health care costs for Type 2 diabetes over this time period. RESULTS: The total UK population will not increase by more than 3% at any time in the next 60 years. However, the population over 30 will increase by a maximum of 11% by 2030. Due to population ageing, in 2036 there will be approximately 20% more cases of Type 2 diabetes than in 2000. Cases of diabetes-related complications will increase rapidly to peak 20-30% above present levels between 2035 and 2045, before showing a modest decline. The cost of health care for patients with Type 2 diabetes rises by up to 25% during this period, but because of reductions in the economically active age groups, the relative economic burden of the disease can be expected to increase by 40-50%. CONCLUSION/INTERPRETATION: In the next 30 years Type 2 diabetes will present a serious clinical and financial challenge to the UK NHS. PMID- 12121331 TI - The evolution of diabetes information systems. AB - Diabetes information systems have already evolved rapidly in recent years along a developmental pathway initiated by the St Vincent Declaration, fuelled by the rapid pace of IT development in the 1990s and now endorsed by the emerging NHS information strategy. They will be central to the delivery of 'patient-centred' care and essential to supporting and monitoring the diabetes national service framework implementation. Widespread experience has identified three key principles. Firstly the need for a core data set that supports both service delivery and quality development. Secondly, because of the multiprofessional, multisector nature of diabetes care, there is a need to reconcile information from many diverse sources into unitary diabetes care records. Thirdly the crucial importance of making data collection a by-product of every day care delivery (i.e. no duplicate data entry). The work of many local innovators, allied to the increasing experience of the Diabetes UK sponsored UKDIABS project has generated substantial expertise. With the aid of new extraction/analysis tools such as QUIDS and a consistent approach to assessment, this work has hopefully laid secure foundations for monitoring the implementation of the national service framework. Furthermore, parallel developments under the aegis of the National electronic Library for Health (NeLH) should enable those involved with diabetes care to access relevant knowledge and information with ease. Increasingly user friendly ways by which patients can interact with their electronic records and linked knowledge sources will create many new opportunities. Diabetes information systems are likely to be at the forefront of diabetes care delivery in the future, providing patients and professionals with timely and accurate data for the organization and delivery of care. PMID- 12121332 TI - The GP perspective: problems experienced in providing diabetes care in UK general practice. AB - AIMS: To describe the problems and barriers perceived by general practitioners (GPs) whilst providing diabetes care in primary care in England and Wales and to identify those health authorities (HAs) in which primary care reported the most and least difficulty. DESIGN: Descriptive postal survey using a self-administered questionnaire. SUBJECTS: One thousand eight hundred and seventy-three randomly sampled GP practices (one in five practices). RESULTS: One thousand three hundred and twenty (70%) responded. Getting patients to alter their lifestyles was perceived as causing the most difficulty in managing individual patients, followed by lack of time, patients' nonattendance, noncompliance with medical regimens and poor communication with secondary care. The greatest barriers to practices providing desirable care were lack of time/under-funding and keeping up to date in the area of diabetes, followed by lack of space, inadequate chiropody, dietetics, ophthalmology and access to secondary care. There are important differences between HAs in the difficulties experienced by primary care teams and we have ranked the HAs accordingly. CONCLUSION: The study has identified what problems need tackling in order to assist primary care to deliver good quality diabetes care, and has highlighted HAs where primary care needs further help, and HAs where examples of good practice may be found and useful lessons learnt. This should form the basis of a needs-based research and development programme for diabetes in primary care. PMID- 12121333 TI - Diabetes UK funded surveys of the structural provision of primary care diabetes services in the UK. AB - AIM: To describe diabetes service provision in primary care in the UK. METHODS: Postal questionnaires were sent to all UK primary care organizations (PCOs), and to a sample of general practices in England and all practices in Wales and Scotland. The data collection period ended on 30 April 2001. RESULTS: Seventy nine per cent of the PCOs and 40% of the practices provided usable information. There is evidence that respondents were not significantly biased in relation to their interest in diabetes care. Diabetes was included as a Health Improvement Programme (or equivalent) priority by 62% of PCOs and had been identified as a clinical governance priority by 27%. Sixty-five per cent had information about the ethnic composition of their general population, 57% had an estimate of the number of people with diabetes. Sixty-nine per cent had a local diabetes register but this was said to cover the entire local population in only 64% of these. At least one audit of diabetes care had been carried out (in the previous 5 years) in 75% and, in 76%, clinical guidelines on diabetes care were made available to practices. In the practices, 80% had a designated lead person for diabetes. Seventy-three per cent had at least one general practitioner with a special interest and 87% at least one nurse. Seventy-two per cent of practices ran specific diabetes clinics and 51% had a screening policy. Eighty-six per cent considered that they had adequate systems in place for the delivery of diabetes care. However, only 6% were able to offer a dedicated diabetes telephone help or advice line and only 9% an evening out of hours clinic. Regular practice meetings were held to discuss diabetes in 35%, whereas 39% had a formal shared care protocol. Fourteen per cent held regular joint meetings with the hospital-based team and in 38% there was membership of Diabetes UK for at least one partner or the practice itself. A third (34%) of responding practices were unsure whether a Local Diabetes Services Advisory Group or equivalent existed in their area. Geographical differences in service provision were identified with, for example, practices in London having fewer components in place that were specifically related to the provision of diabetes care. Single-handed practices, wherever they were situated, had in place fewer staff and facilities specifically for diabetes care. PMID- 12121334 TI - Association of British Clinical Diabetologists (ABCD): survey of specialist diabetes care services in the UK, 2000. 2. Workforce issues, roles and responsibilities of diabetes specialist nurses. AB - AIM: To examine the provision and role of diabetes specialist nurses (DSNs), and the content of patient education programmes in the UK. METHOD: A postal survey of secondary care providers of diabetes services in the UK in 2000. RESULTS: Following two reminders, a 77% response rate demonstrated 2.5 (median) whole time equivalent DSNs per 250 000 population, with only 13% of centres meeting the recommended staffing level of four per 250 000 population. The vast majority carried out work both in hospital and in the community, the proportion a reflection of who employed and managed staff. There was a wide variation in the qualifications required and the nursing gradings of DSNs, and regional variation in the number of grade I nurses, with the greatest proportion based in the South east of England. The vast majority (96%) provided patient education, and where it existed (in 60% of responses), were the major providers of a patient help line (90%). Although key providers of patient education, there had been no specific education for this task in over 20% of responses. There was broad consistency in the topics covered at educational sessions, although advice on footwear (76%) and home urine glucose monitoring (73%) were least frequently documented. The issuing of literature and cards for patient use was also very variable. Over 25% of bids for diabetes service improvement were for additional DSNs, but only 48% of these were successful. CONCLUSIONS: There has been an improvement in staffing levels of DSNs over the last 10 years but the numbers are many fewer than recommended in national strategy documents, with evidence that despite expansion being given a high profile, such efforts are often unsuccessful. There was also evidence of considerable variation in the qualifications and gradings of DSNs throughout the UK and indeed in their day-to-day roles, and the content of patient education programmes. This suggests the need for a nationally co-ordinated approach to training and recruitment. PMID- 12121335 TI - Association of British Clinical Diabetologists (ABCD): survey of specialist diabetes care services in the UK, 2000. 3. Podiatry services and related foot care issues. AB - AIM: To examine the provision of, and variations in, podiatry and other services for diabetic foot care in the UK. METHOD: A postal survey of secondary care providers of diabetes services in the UK in 2000. RESULTS: Following two reminders a 77% response rate was achieved. The responses indicated that 97% had a state-registered podiatrist attached to the service, providing three (median) sessions each week for diabetes care, although only 44% had availability at all diabetic clinics, and only 3% had availability at paediatric diabetic services. Podiatry access at all diabetic clinics increased the likelihood of associated preventative as opposed to reactive ('trouble shooting') care (P < 0.05). All individuals with feet at 'high risk' of ulceration had access to 'at least 2 monthly review' in 15% of trusts, and with active foot ulceration at least weekly in 43%. Over 70% used at least one form of equipment to assess peripheral neuropathy, but peripheral blood flow was only formally measured in 13%. Although podiatry input to patient education was common (84%), only 6% had received formal training in education. Guidelines and strategies for management of active foot problems were available in 50-74% of cases. Orthotic input was highly variable, and absent in 15% of responses. Podiatrist fitting and application of foot protective apparatus was only recorded in 22-61% of responses. Access to isotopic and/or MR foot imaging and peripheral angiography and angioplasty was recorded in 75-83% of responses. Separate specialist foot clinics were available in 49%, and where this was the case the use of newer foot ulcer healing applications was higher (P < 0.01). Clear regional differences were apparent in the nature of the service, the use of newer treatments, and in access to an orthotist, a local 'dedicated' foot surgeon or a separate diabetic foot clinic. Of 245 documented bids for service improvements, only 19 related to foot care and only 21% of bids were successful. CONCLUSIONS: Despite an increase in podiatry support to diabetes care over the last 10 years, the level of access and the nature of the services provided is much less than recommended in many advisory documents. The strategy of a co-ordinated 'team' approach to foot care still takes place in less than 50% of centres. There are clear regional differences in diabetes foot care services. Both providers and purchasers of diabetes services may not have given sufficient attention to this area, given the relatively small number of documented bids for service improvements in this area, and the very low success rate of such bids. PMID- 12121336 TI - Association of British Clinical Diabetologists (ABCD): survey of specialist diabetes care services in the UK, 2000. 4. Dietetic services and nutritional issues. AB - AIMS: To examine the provision of, and variations in, dietetic services for diabetes in secondary care in the UK. METHOD: A postal survey of all secondary care providers of diabetes services. RESULTS: There was a 77% response rate. A dedicated dietician supported diabetes services in 73% of responses, but only 45% were able to see newly diagnosed patients within 1 month. Only 3% of responses documented that dietetic services provided the recommended minimum 22 h weekly input to diabetes care, and an annual dietetic review was said to be available in 15%. An opportunity for more frequent visits was most likely if there was poor glycaemic control (78% of responses), particularly when services were provided by a dedicated diabetes dietician. Although dieticians frequently provided input to patient education (88%), specific training for this purpose and provision for continuing education of these individuals was less common (14% and 63%, respectively). Nutritional guidelines were available in 74%, but only 31% of responses documented current guidelines on obesity management. Of bids for additional dietetic resources, only 21% had been successful. There was evidence of regional variation in service provision, and no greater provision of dietetic services in areas with a large South Asian population and an expected high prevalence of diabetes. In broad terms, dietetic services for diabetes care had not altered in comparison with a similar survey in 1997. CONCLUSIONS: The level of dietetic support of secondary care diabetes services remains dramatically lower than recommended in advisory documents, and appears to have changed little over the last 3 years. This is compounded by marked regional differences, and was no better in areas with a higher than average prevalence of diabetes. The survey also highlights the need for more co-ordinated and structured education and training of dieticians as well as more consistency in nutritional guidelines. PMID- 12121337 TI - Current status of screening for diabetic retinopathy in the UK. AB - This article reviews the current status of retinopathy screening schemes in the UK. There is evidence that high-quality diabetic retinopathy screening schemes are in existence but provision is patchy. Many health authorities have ad hoc screening programmes reaching only about 60% of patients, with unacceptable or undocumented efficacy and minimal quality control. Several models of screening are currently in use with the current preferred option being camera-based screening. Digital imaging systems offer the best prospects for image acquisition, although at present evidence of adequate effectiveness only exists for 35 mm film-based systems. The final report of the National Diabetic Retinopathy Screening Programme commissioned by the UK National Screening Committee for inclusion into the national service framework for diabetes, is thus eagerly awaited and should set standards for screening programmes, in order to improve the care of all those with diabetes. Quality assurance will be the main driver in the immediate future of improvements in screening programmes. Research data will provide the evidence to refine techniques and set targets in the longer term, with the emphasis on cost-effectiveness and quality of life. PMID- 12121338 TI - Maternity services for women with diabetes in the UK. AB - Diabetic pregnancy is high risk in those regions within the UK that have reported on such an outcome. There is a paucity of information about diabetic pregnancy outcome in the rest of the UK but it seems appropriate to assume that poor outcome is widespread. Previously reports have highlighted problems with the delivery of preconception care and the lack of agreement about the screening and diagnosis of gestational diabetes. The provision of maternity services for women with diabetes in the UK is reviewed. Service development is traced from the 1980s to more recent surveys from Scotland and the former Northern region of England. Service provision has improved over the years in those regions that have agreed and audited standards for provision of care. There is a lack of information about current services within the UK but this will be partly addressed by the forthcoming survey of organization of services co-ordinated by the Confidential Enquiry into Stillbirths and deaths in Infancy (CESDI), which will survey England, Wales and Northern Ireland. In Scotland the Scottish Intercollegiate Guidelines 'Management of diabetes in pregnancy' has been implemented and audited. A similar guideline of agreed standards of care is needed for the rest of the UK. PMID- 12121339 TI - Renal services for people with diabetes in the UK. AB - Diabetic nephropathy is a serious complication of diabetes that can lead to endstage renal failure (ESRF). It is now the most common cause of ESRF in patients accepted onto renal replacement therapy (RRT) programmes in the UK. Rates of diabetic ESRF are more common in ethnic minority populations. The risk of developing diabetic ESRF is higher in Type 1 diabetes but in absolute terms more patients with Type 2 diabetes develop ESRF and are treated. There is still unmet need for RRT amongst patients with diabetes who develop ESRF. The shortage of organ donors, especially amongst ethnic minorities, means that dialysis is the mainstay of treatment in patients with diabetes and ESRF. This is now largely hospital haemodialysis with an increasing proportion being delivered in satellite units. Demand for RRT from patients with diabetes will increase due to demographic change and the increasing prevalence of diabetes, particularly Type 2, in the population. To meet this challenge closer liaison between those primarily caring for patients with diabetes (primary care physicians and diabetologists) and nephrologists is required to ensure effective surveillance of renal function, to increase early referral and to agree protocols of subsequent care. Continued expansion of high-quality RRT is needed that ensures equity of access with particular targeting in areas with large ethnic minority populations. A national priority must be an increase in the kidney transplant rate. PMID- 12121340 TI - Diabetes care in childhood and adolescence. AB - The presentation of diabetes in young people has changed significantly over recent years. Not only has there been a rising incidence of Type 1 diabetes, especially in young children, but also there is an increasing recognition of Type 2 diabetes. Young people are also increasingly being diagnosed with genetic defects of B-cell function and with diabetes in association with cystic fibrosis and other chronic diseases. There have also been significant changes in the pattern of paediatric diabetes care. This is increasingly being provided by a specialized paediatric multidisciplinary team in each health district working to agreed national standards. Despite improvements, diabetes control is still suboptimal with a high incidence of complications being reported in young adults. The challenge over the next few years is the provision of a uniform, equitable and first class paediatric service throughout the UK together with the introduction of new approaches to care, aiming to improve individual diabetic control and reduce long-term complications. Increased collaboration with adult colleagues is needed to enable the transition of care in adolescence to a service that young adults perceive to meet their needs, encourage their attendance and improve their diabetes control and quality of life. A national paediatric diabetes register together with regular audit will encourage these objectives. PMID- 12121341 TI - Diabetes in the elderly: problems of care and service provision. AB - Diabetes is common in the elderly and old UK citizens, affecting between 10% and 25%. There is considerable associated morbidity and mortality, with dementia being a common problem. The diabetic elder is also at risk of drug side-effects. Most of the evidence base for treatment is based on trials performed in younger diabetic subjects or older nondiabetic subjects; however, we can practice evidence-biased medicine whilst awaiting the results of ongoing trials. The older persons national service framework (NSF) may share some similarities with the diabetes NSF; it was 1 year late, and had no clear funding, amongst several other worries. Residential care, which is more likely to be required by diabetic elders, is also under-funded with major concerns about the quality of care for the diabetic resident. The little evidence that we have regarding care of the older diabetic person also suggests inadequacies. Given the likelihood that we will have to manage with present resources, managed clinical networks may be one way to cope. PMID- 12121342 TI - The Audit Commission review of diabetes services in England and Wales, 1998-2001. AB - AIMS OF THE AUDIT COMMISSION: The Audit Commission has a statutory duty to promote the best use of public money. It does this through value for money studies, such as that reported in Testing Times[1]. This work has been followed with a review of innovative practice in commissioning. These initiatives aim to support the implementation of the diabetes national service framework. The Audit Commission also appoints external auditors to NHS organizations who assess probity and value for money in the NHS; the latter by applying national studies locally and by carrying out local studies. METHODS: Research for Testing Times consisted of structured visits to nine acute trusts, a telephone survey of 26 health authorities and a postal survey of 1400 people with diabetes and 250 general practitioners. Local audits used a subset of the original research tools. Case studies were identified through a cascade approach to contacts established during Testing Times and through self-nomination. RESULTS: Rising numbers of people with diabetes are placing increasing pressure on hospital services. Some health authorities and primary care organizations have reviewed patterns of service provision in the light of the increasing demands. These reviews show wide variations in patterns of routine care. In addition, there is a widespread lack of data on the delivery of structured care to people with diabetes. People with diabetes report delays in gaining access to services, and insufficient time with staff. There are insufficient arrangements in place for providing information and learning opportunities to support self-management. CONCLUSION: As the number of people with diabetes continues to rise, the potential for providing more care in a primary care setting needs to be explored. This will enable specialist services to focus more effectively on those with the most complex needs. PMID- 12121343 TI - Improvements in the provision of hospital diabetes services in an English region 1988-98. AB - AIMS: To investigate the changes in provision of hospital-based services for patients with diabetes in an English region over a 10-year period. METHODS: Questionnaires were completed by lead clinicians in hospitals in the Northern region of England in 1988 and repeated in 1998. Information was sought on diabetes service provision including the staff and their working practices. Data are presented to demonstrate changes during the 10 years. RESULTS: During a 10 year period the number of consultants providing specialized diabetes services increased from 16 to 25 (to become one per 126 240 population). Their outpatient sessions changed from 34 to 55.5 per week, with a decrease in nonspecialists providing diabetes services. Reductions occurred in registrar numbers providing sessions from 23 to 15 and senior house officers from 16 to 14. Increases occurred in other health care professionals: diabetes specialist nurses from 19 to 30.3 whole time equivalents (WTEs); dieticians from 16 to 32.3 WTEs and chiropodists from 8 to 23 WTEs. The numbers of specialized clinics and units providing services from diabetes care centres increased. Improved facilities in clinics and access to laboratory tests were available to all units. Diabetes registers came into use in 12 of 16 units, but there have been difficulties in providing funding. 'Out-of-hours' advice has moved towards advising their patients to see their general practitioners or the accident and emergency department of the hospitals. CONCLUSIONS: The number of diabetes professional staff and the provision of specialized diabetes services have increased during a 10-year period in the Northern region of England. However, they still fall far short of recommended staffing levels and services are far from comprehensive in most districts. PMID- 12121345 TI - Inherited disorders of cholesterol biosynthesis. AB - For many decades, cholesterol has been considered an important structural component of cellular membranes and myelin, and a precursor of steroid hormones and bile acids. Moreover, the recognition that high cholesterol levels (hypercholesterolemia) are a major risk factor for the development of heart disease and atherosclerosis has gained enormous attention not only in medicine, medical and pharmacological research, but also from the general public. The discovery of a crucial role of cholesterol in human embryogenesis and the recent identification of a number of inherited disorders of cholesterol biosynthesis also show that low cholesterol levels (hypocholesterolemia) may have severe consequences for human health and development. In the past few years, seven distinct inherited disorders have been linked to different enzyme defects in the cholesterol biosynthetic pathway by the finding of abnormally increased levels of intermediate metabolites in patients followed by the demonstration of disease causing mutations in genes encoding the implicated enzymes. Patients afflicted with these disorders are characterized by multiple morphogenic and congenital anomalies including internal organ, skeletal and/or skin abnormalities. PMID- 12121346 TI - History of genetics through philately: Sir William Osler (1849-1919) and the Osler-Weber-Rendu syndrome. PMID- 12121347 TI - LDL receptor mutation genotype and vascular disease phenotype in heterozygous familial hypercholesterolaemia. AB - Patients with homozygous familial hypercholesterolaemia (FH) caused by receptor negative, low-density lipoprotein (LDL) receptor gene mutations have higher concentrations of LDL-cholesterol in plasma and earlier onset of cardiovascular disease (CVD) than patients homozygous for receptor-defective, LDL receptor mutations. In contrast, it is uncertain whether the severity of atherosclerotic disease differs in heterozygous FH caused by receptor-negative and receptor defective mutations. The present authors investigated the influence of LDL receptor mutation type on the clinical phenotype in 31 patients with heterozygous FH caused by the receptor-negative, Trp23-stop mutation and in 31 patients heterozygous for the receptor defective Trp66-Gly mutation. Untreated levels of plasma LDL-cholesterol and calculated cholesterol-years score did not differ significantly between the two groups of patients. Detection of vascular disease was based on two approaches: (1) measurement of coronary calcification by spiral computed tomography (CT) scanning; and (2) ultrasonic measurement of carotid intima-media thickness (IMT). Age was significantly correlated to the presence of coronary calcification, but controlling for relevant cofactors, there was no evidence that the receptor-negative mutation caused more calcification than the receptor-defective mutation. Furthermore, carotid IMT was significantly influenced by plasma concentrations of Lp(a) and triglycerides, as well as by age, sex and smoking status, but again, there was no statistically significant effect of LDL receptor gene mutational type. The similarity in vascular phenotypes was probably caused by a similar life-long burden of LDL-cholesterol in the two groups of patients. PMID- 12121348 TI - Type III hyperlipoproteinema with apolipoprotein E2/2 genotype in Japan. AB - Limited information is available concerning type III hyperlipoproteinemia (HLP) in the Asian population. Therefore, clinical and biochemical characteristics of type III HLP were examined in 16 Japanese patients. Mean plasma triglyceride (TG) and total cholesterol (chol) levels were 381 mg/dl and 253 mg/dl, respectively, and the mean very low density lipoprotein (VLDL)-chol/plasma TG ratio was 0.27, which were lower than those reported in Western countries. Eighty percent of the patients had high plasma remnant-like particles (RLP)-chol levels above 50 mg/dl and a high RLP-chol/plasma TG ratio above 0.1. Twelve patients (75.0%) were obese. Seven patients (43.8%) had type 2 diabetes mellitus and four patients (25.0%) had impaired glucose tolerance. Six patients (37.5%) had coronary heart disease (CHD), but none had peripheral vascular disease or xanthomas. TG-rich lipoproteins from type III HLP patients with diabetes mellitus stimulated cholesteryl ester synthesis by human macrophages significantly (p < 0.001) more than those from type III HLP patients without diabetes mellitus. In conclusion, the Japanese type III HLP patients had lower plasma TG and total chol levels and a lower VLDL-chol/plasma TG ratio, but CHD was more common. The patients were characterized by a high frequency of obesity and/or glucose intolerance. The TG rich lipoproteins from type III HLP patients with diabetes mellitus were more atherogenic. PMID- 12121349 TI - Endothelial nitric oxide synthase gene polymorphisms in Fabry's disease. AB - The gene encoding endothelial nitric oxide synthase (eNOS) is involved in abnormalities in nitric oxide (NO) synthesis that mediates functional damage of vascular cells, especially of endothelial cells (ECs), a common characteristic in cardiovascular diseases. In Fabry's disease, the characteristic mutation in the alpha-galactosidase A (alpha-gal A) gene induces large deposits of glycosphingolipids, particularly concentrated in ECs, a process associated with endothelial dysfunction. To determine whether in addition to alpha-gal A gene mutations, eNOS genetic variations are implicated in this process, we examined the genotypes of the missense Glu298Asp (G894T) variant in exon 7 and 27-bp tandem repeats in intron 4 (4b/a) in 19 patients with Fabry's disease, and 39 normal volunteers. The results showed that both varials have a significant association with Fabry's disease. The frequencies of mutant Glu/Asp + Asp/Asp genotypes and Asp allele are significantly higher in Fabry's disease (68.4%, p = 0.044, and 47.4%, p = 0.022, respectively) than in controls (46.7% and 25%, respectively). The frequencies of eNOS 4b/a polymorphisms are also significantly different in Fabry's disease when compared to controls. The mutant 4b/a + 4a/a genotype frequencies are 55.5% (p = 0.032) and 4a allele 27.8% (p = 0.05) compared with controls (23.1% and 12.8%, respectively). These results indicate that more than half of the patients with Fabry's disease carry the Glu298Asp variant ( approximately 68%) and/or the 4b/a polymorphism ( approximately 55%). To the best of our knowledge, this is the first report showing an influence of eNOS gene polymorphisms in patients with Fabry's disease. PMID- 12121350 TI - Effects of apolipoprotein A-IV genotype on glucose and plasma lipoprotein levels. AB - The effects of apolipoprotein (apo) A-IV genotype on serum glucose, total cholesterol, low density lipoprotein (LDL) cholesterol, high density lipoprotein (HDL) cholesterol, triglyceride and glucose concentrations were ascertained in a population of 373 men and 361 women with a mean age of about 57 years. Subjects were evaluated at entry into a lifestyle intervention program. Apolipoprotein A IV genotype variations at residues 347 and 360 were examined, as these mutations affect the sequence of apo A-IV, a major protein constituent of intestinal triglyceride-rich lipoprotein and HDL. With regard to the apo A-IV 360 mutation, 16.4% of the females and 13.4% of the males carried the apo A-IV 2-allele, almost entirely in the heterozygous state. No effect of the apo A-IV 1/2 genotype was observed in either men or women on total cholesterol, LDL cholesterol, HDL cholesterol, triglyceride, the total cholesterol (TC)/HDL ratio, or on A-I, A-IV and apo B levels. This was also the case for the apo A-IV 347 mutation. However, women with the apo A-IV 360 1/2 genotype had significantly (p < 0.005) higher glucose levels (105.5 mg/dl) compared with the 1/1 wild-type (94.0 mg/dl). All analyses were also adjusted for age, body mass index, medications, alcohol use and cigarette smoking. The prevalence of the 347 mutation was somewhat higher than the 360 mutation, with 29% of the females and 32.0% of the males being heterozygous for this mutation, and 3.9% of the females and 5.4% of the males being homozygous for this mutation. These data are consistent with the concept that the apo A-IV 360 and 347 genotypes have no significant effect on apo A-IV levels and other lipid parameters in either gender. However, apo A-IV 360 1/2 genotype did have a significant effect on serum glucose levels in women. PMID- 12121351 TI - Homozygosity in Huntington's disease: new ethical dilemma caused by molecular diagnosis. AB - Huntington's disease (HD) is a degenerative disorder of the central nervous system with autosomal dominant inheritance. Genetic counseling has always been difficult in this disorder with anguish, depression and denial being very common in both the patient and family members. The discovery of the causal gene has led to precise diagnostic procedures allowing homozygotes for the disease to be identified. Contrary to what occurs in some other autosomal dominant diseases, the course of the disease is not more severe in the homozygote than in the heterozygote. The present authors describe a family comparing two affected siblings: one is heterozygotic and the other homozygous for the HD mutation. They confirm that the age and symptoms of onset did not differ significantly between the subjects; however, the disease seemed to have a more severe progression in the heterozygote than in the homozygote. The authors discuss the ethical dilemma derived from the genetic counseling of a homozygotic patient, given the fact that all his offspring will be affected. Letting the offspring know about their 100% probability of inheriting the disorder is equivalent to delivering a non requested predictive test, while not informing them constitutes withholding crucial information from the individual. PMID- 12121352 TI - An Italian severe Salla disease variant associated with a SLC17A5 mutation earlier described in infantile sialic acid storage disease. AB - The present study reports two Italian brothers affected by severe Salla disease (sialic acid storage disease), a slowly progressive autosomal recessive neurodegenerative disorder prevalent in the Finnish population. Mutations of the SLC17A5 gene, which encodes a protein called sialin, are the primary cause of both Salla disease and infantile sialic acid storage disease (ISSD), a clinically distinct severe disorder. All Finnish patients with Salla disease show a R39C mutation. Both patients showed moderate intellectual disability, spastic ataxic syndrome, hypomyelination and cerebellar atrophy on magnetic resonance imaging (MRI), and lysosomal storage, all typical of Salla disease. Mutation analysis of the SLC17A5 gene in the younger brother revealed no R39C mutation, but a 15-bp deletion in exon 6 on one of the alleles. This mutation is the same described in French-Canadian patients with ISSD. Salla disease must be suspected in patients with unexplained psychomotor retardation associated with ataxia and/or pyramidal symptoms, and MRI findings consistent with cerebral hypomyelination, irrespective of the patient's ethnic origin. A mutation screening based on R39C change does not exclude Salla disease outside Finland. Conversely, mutations found in ISSD can be expected, even in patients showing the Salla phenotype (e.g. symptoms at the milder end of the spectrum). PMID- 12121353 TI - Facial clefting in an Arab town in Israel. AB - To study the prevalence of cleft palate and cleft lip with or without cleft palate in an Israeli Arab town, questionnaires were sent to the parents of 1375 pupils in grades 1 and 2 in all seven primary schools in the town of Taibe, and 1281 responded. The information requested included data about siblings and members of the parental generation to give a total of 16 174, and the presence of consanguinity and history of exposure to medication, radiation, smoking or alcohol during pregnancy. There were four affected individuals among the index cases, of whom two had cleft palate only and two cleft lip with cleft palate, giving prevalence rates for each of these of 1.56/1000. Adding to these the number of affected siblings gave a total of 10 affected individuals; two with cleft palate only (0.39/1000) and eight with cleft lip with or without cleft palate (1.56/1000). Among the parental generation, of 16 reported affected individuals, two had cleft palate only (0.18/1000) and 14 cleft lip with or without cleft palate (1.26/1000). The overall prevalence rate for all 26 affected individuals was 1.6/1000; four of these had cleft palate only (0.24/1000) and 22 had cleft lip with or without cleft palate (1.36/1000). There were no cases whose mothers had been exposed to medication, radiation, smoking or alcohol during pregnancy. The effect of consanguinity was not significant (P < 0.92). This study shows that the prevalence of facial clefting in an Israeli Arab community is consistent with that in the general population worldwide. PMID- 12121354 TI - Mutation in the cartilage-derived morphogenetic protein-1 (CDMP1) gene in a kindred affected with fibular hypoplasia and complex brachydactyly (DuPan syndrome). AB - The present authors have previously described a consanguineous Pakistani family with fibular hypoplasia and complex brachydactyly (DuPan syndrome) inherited as an autosomal recessive trait. All affected individuals showed either reductions or absence of bones in the limbs, and appendicular bone dysmorphogenesis with unaffected axial bones. Obligate heterozygote parents were phenotypically normal. Mutations in the cartilage-derived morphogenetic protein 1 (CDMP1) gene have been reported in two acromesomelic chondrodysplasias (i.e. Hunter-Thompson type and Grebe type) which are phenotypically related to DuPan syndrome. CDMP1, a member of the transforming growth factor beta super-family of secreted signalling molecules, has been reported to regulate limb patterning and distal bone growth. Therefore, the present authors examined genomic DNA from the family with DuPan syndrome for mutations in the CDMP1 gene. Affected individuals were homozygous for a missense mutation, T1322C, in the coding region of the CDMP1 gene. This mutation was not found in 44 control subjects of Pakistani origin. The T1322C change predicts a leu441pro substitution in the mature domain of the CDMP1 protein. This is likely to cause a conformational change in the CDMP1 protein that influences the expression of genes which are required for normal bone development. This finding extends the spectrum of phenotypes produced by defects in the CDMP1 gene. PMID- 12121355 TI - Homozygosity for the V37I Connexin 26 mutation in three unrelated children with sensorineural hearing loss. AB - Mutations in the Connexin 26 (Cx26) gene have been found to account for approximately 20% of all childhood deafness. This number approaches 50% in documented recessive cases of hearing loss. Two mutations, 35delG and 167delT, account for the majority of reported mutations in this gene, but to date, more than 60 mutations have been described. No other single gene has yet been identified that contributes this significantly to the aetiology of hearing loss. Several mutations in this gene have been found to predominate in specific ethnic populations (167delT in Ashkenazi Jews and 235delC in Japanese individuals). While the majority of mutations found in Cx26 result in frame shifts and premature terminations, a number of missense mutations have also been identified. The V37I missense mutation has been reported as both a polymorphism and as a potentially disease-causing missense mutation. The present authors have identified three unrelated individuals with sensorineural hearing loss who are homozygous for this mutation. One individual is of Philippine ancestry, another is from a Chinese and Cambodian background, while the third is of Chinese ancestry, raising the possibility that this mutation may be more frequent among populations in eastern Asia. PMID- 12121356 TI - Tubulopathy, endocrinopathies and encephalomyopathy in a child with a novel large scale mitochondrial DNA deletion. PMID- 12121357 TI - Esophageal and duodenal atresia in a girl with a 12q24.3-qter deletion. PMID- 12121359 TI - The trouble with tonometry. PMID- 12121360 TI - Relationship between intraocular pressure and systemic health parameters in a Korean population. AB - PURPOSE: This study aimed to evaluate the relationship between intraocular pressure (IOP) and age and obesity, adjusted for systemic health parameters such as sex and mean blood pressure, in a Korean population. METHODS: A total of 13 212 healthy participants underwent automated multiphasic tests, including tonometry, automated perimetry, fundus photography, blood pressure and body mass index (BMI). Six age groups were used, divided by decades ranging from 20-29 years to 70+ years. The association between IOP and systemic health para-meters was examined using cross-sectional analysis. RESULTS: The median age of participants was 47.6 years (range 20-84 years), and 6684 (50.6%) of participants were men. The mean IOP of participants was 15.5 mmHg. The mean IOP, blood pressure and BMI values were significantly higher in men than in women (P < 0.05). The overall prevalence of ocular hypertension, defined as IOP >21 mmHg without signs of glaucomatous visual field loss or optic disc damage, was 6.1% in men and 2.5% in women. Intraocular pressure was associated with mean blood pressure, sex, age and BMI by multiple regression analysis (P < 0.05). The relationship between IOP and age adjusted for sex, mean blood pressure and BMI had a significantly negative tendency for both sexes (P < 0.05). Body mass index had a significantly positive relation with IOP after controlling for age, sex and mean blood pressure in men (P < 0.05), but not in women. CONCLUSIONS: In this Korean population, after multiple adjustment, IOP was found to decrease with age and to increase with BMI in men. PMID- 12121361 TI - Analysis of risk factors that may be associated with progression from ocular hypertension to primary open angle glaucoma. AB - BACKGROUND: As a multifactorial disease, glaucoma may be associated with pressure dependent and pressure-independent factors. Ocular hypertension (OHT) may develop into primary open angle glaucoma (POAG) for many patients. Groups with OHT and POAG were compared for pressure-dependent and independent risk factors. A high prevalence of any factor(s) could indicate a contribution to progression from OHT to POAG. METHODS: A sample of patients with POAG (n = 438) and with OHT (n = 301) were selected from those attending a tertiary referral private glaucoma practice, and data were collected regarding age and intraocular pressure at the time of diagnosis, sex, family history of glaucoma, systemic hypertension, diabetes, Raynaud's phenomenon, migraine and myopia. RESULTS: After multivariate analysis, older age at time of diagnosis (chi(2)(5) = 73.89, P < 0.001), myopia (odds ratio [OR] = 1.5, 95% confidence interval [CI] 1.0-2.2; P < 0.05), a family history of glaucoma (OR = 1.6, 95% CI 1.1-2.3; P < 0.01) and a high intraocular pressure (chi(2)(4) = 16.96; P = 0.002) were found to be more prevalent among those with POAG. No other significant differences could be found between the two groups. CONCLUSION: Patients who have OHT may be at higher risk of developing POAG if they also have myopia, a family history of glaucoma or are of older age. PMID- 12121362 TI - Posterior capsule opacification after cataract surgery in remote Australian Aboriginal patients. AB - PURPOSE: The aim of this retrospective study was to determine the rate of visually significant posterior capsular opacification formation after cataract surgery for Australian Aborigines living in rural or remote areas in the 'Top End' of the Northern Territory, Australia, and then to assess these patients' outcomes after capsulotomy. METHODS: Aboriginal patients living in remote areas of the Top End of the Northern Territory who underwent cataract surgery between 1994 and 1999 were identified from records at the three major hospitals in the region. The presence of posterior capsular opacification (PCO) was determined by clinical examination. The primary endpoint for this study was the presence of axial opacification of the posterior capsule and the need for subsequent Nd:YAG posterior capsulotomy to improve sight. Linear regression analysis of the time from surgery to follow up and the number of eyes requiring Nd:YAG capsulotomy was performed. Operated eyes were grouped according to the interval between surgery and follow up (Group 1: follow up within 1 year of surgery, n= 25; Group 2: follow up 1-3 years after surgery, n= 42; Group 3: follow up 3-5 years after surgery, n= 51). RESULTS: One hundred and eighteen operated eyes were examined. Eyes in Group 3 were found to have the highest incidence of visually significant PCO (27.5%). There were more eyes requiring capsulotomy after 3 years than after 1 year following surgery. Linear regression analysis revealed an odds ratio of 1.4 (P = 0.07). All nine eyes in the 1-3 year group that had developed visually significant PCO had undergone extra-capsular cataract extraction. CONCLUSIONS: For the remote Aboriginal patient who has undergone cataract surgery, there is a relatively minor chance of developing PCO within the first postoperative year regardless of the type of surgery undertaken. This study illustrates that the longer the time after surgery the greater the chance of developing visually significant PCO. For the remote Aboriginal patient there is a high chance (approximately 28%) of developing visually significant PCO within 5 years after cataract surgery. These figures are lower than those reported from other parts of Australia. PMID- 12121363 TI - Use of contact lenses to correct aphakia in children. AB - BACKGROUND: Contact lenses play an important role in the management of childhood aphakia. However, the use and care of the lenses can be challenging. The aim of this study was to determine how parents coped and what problems they encountered. METHODS: Thirty-one families of aphakic children who were fitted with contact lenses between 1990 and 1997 were contacted and invited to participate in a telephone survey. RESULTS: Thirty families agreed to participate. Eighteen families (60%) were still using contact lenses successfully at the time of the study. Four families (13.3%) stopped using contact lenses within a year because of overwhelming difficulties handling the lenses. There was an initial learn-ing process that was often stressful and frustrating. The learning curve lasted from 1 to 24 months (first quartile 2.0 months; median 3.3 months; third quartile 15.0 months). The personality of the parent, temperament of the child and the frequency at which problems occurred may all have contributed to how each family coped with the lenses. Common initial problems included inserting the lenses into the eyes of an uncooperative child and the loss of lenses. Other problems included irritation associated with dryness, and the financial cost of maintaining the lenses. CONCLUSION: Although most parents initially found contact lenses daunting, 70% of families surveyed eventually learned to cope well. Parents felt that proper education of both parents and children on why the lenses were required, a good appreciation of problems that may occur, a strong social support network and adequate financial aid were important factors in learning to cope. PMID- 12121364 TI - Phacoemulsification in filtered chronic angle closure glaucoma eyes. AB - PURPOSE: To report the results of phacoemulsification in eyes with chronic angle closure glaucoma having a functional filtering bleb. METHODS: A prospective and non-comparative study was undertaken in 60 consecutive eyes of 44 patients having a functional filtering bleb for chronic angle closure glaucoma and undergoing cataract surgery. Only those cases who had intraocular pressure 0.05 using paired t-test). The central anterior chamber depth was statistically significant (P = 0.002) when preoperative (1.90 +/- 1.27 mm) was compared to postoperative at 6 months (2.11 +/- 1.27 mm). CONCLUSION: Phacoemulsification in eyes with a functional filtering bleb for chronic angle closure glaucoma is challenging. However, with slight modifications in technique, it can be accomplished without compromising the functioning of the bleb. PMID- 12121365 TI - Vitreous surgery in highly myopic retinal detachment resulting from a macular hole. AB - PURPOSE: This study aimed to assess the therapeutic effect of vitreous surgery in conjunction with photocoagulation for highly myopic retinal detachment resulting from a macular hole. METHODS: Sixty-two consecutive highly myopic patients (65 eyes) with retinal detachment from macular holes underwent vitreous surgery. Gas tamponade in conjunction with laser photocoagulation was performed in 46 eyes (44 cases, group 1); gas tamponade only was performed in 12 eyes (11 cases, group 2); and silicone oil tamponade only was performed in seven eyes (seven cases, group 3). Additional laser photocoagulation was given in group 1 if necessary. The anatomical and functional success rates were compared between the groups. RESULTS: Primary retinal reattachment was achieved in 43 eyes (93.5%) in group 1, seven eyes (58.3%) in group 2 and four eyes (57.1%) in group 3. Final visual acuity was 6/60 or more in 24 eyes (52.2%) in group 1, in six eyes (50.0%) in group 2 and in three eyes (42.9%) in group 3. The initial retinal reattachment rate was significantly higher in group 1 than in group 2 (P = 0.0075) and group 3 (P = 0.0248). The macular hole was completely closed in 18 eyes in group 1 and one eye in group 2 after 2 months or longer. A thin fibrous membrane and scar could be easily noticed beneath the macula in 15 eyes. CONCLUSION: Vitreous surgery in conjunction with laser photo-coagulation can improve the surgical success rate for highly myopic retinal detachment resulting from a macular hole. PMID- 12121366 TI - Ocular contusion caused by elastic cords: a retrospective analysis using the Erlangen Ocular Contusion Registry. AB - PURPOSE: Elastic cords are common tools to secure luggage to the roof of motor vehicles or the carrier of bicycles. In comparison with other variants of ocular contusion, patients with elastic cord injuries appear to suffer more severe ocular damage. METHODS: The charts of 398 consecutive patients of the Erlangen Ocular Contusion Registry who had been hospitalized for ocular contusion over a 10-year period were retrospectively reviewed. The acute clinical findings of 23 patients with elastic cord injury were compared with 375 patients with other contusions. Open globe injuries were excluded. RESULTS: A total of 5.8% of the patients were injured by elastic cords; their mean age was 36.6 +/- 17.0 years (range 8-66; median 34 years). The following morphological changes were documented in elastic cord injuries versus other ocular contusions: hyphema 100% versus 75%; angle recession 92% versus 72%; iridodialysis 17% versus 9%; traumatic cataract 14% versus 10%; lens dislocation 38% versus 13%; choroidal rupture 19% versus 6%; peripheral Berlin's oedema 57% versus 35%; and central Berlin's oedema 29% versus 12%. The difference of the incidence was significant for lens dislocation (P < 0.01), choroidal rupture (P = 0.024), and central (P = 0.029) and peripheral Berlin's oedema (P = 0.043). The final visual outcome was lower in patients with elastic cords injuries (P = 0.008). CONCLUSION: Elastic cord injuries induce more severe ocular damage than most other contusions. The elastic cords producing industry is called on to develop safer luggage fixation systems and warn against the potential harmful consequences of injury to the eye. PMID- 12121367 TI - Effectiveness of local anaesthesia for external dacryocystorhinostomy. AB - PURPOSE: To establish the effectiveness, complications and patient acceptance of local anaesthesia (LA) with intra-venous sedation, for external dacryocystorhinostomy (DCR). METHODS: Data were prospectively collected over a 4 year period (1997-2000) on all patients undergoing external DCR under LA with intravenous sedation by one surgeon and one anaesthetist, using a standardized technique. RESULTS: Of 183 DCR procedures, 145 (76.5%) were performed using LA in 123 patients aged 24-84 years (median 64 years). Eleven simultaneous bilateral DCR procedures were performed under LA. Another 11 patients had a contralateral DCR at a later date. In only three DCR (2.1%) was supplementation of LA required during the procedure because of pain. The only complication of the technique was one retrobulbar haemorrhage from the medial peribulbar injection. In this patient, the procedure was completed satisfactorily without further sequelae. All patients found the technique acceptable and all 11 patients who returned for surgery on the opposite side at a later date elected to have surgery under LA again. CONCLUSIONS: External DCR can be satisfactorily and safely performed under LA with a high level of patient acceptance. PMID- 12121368 TI - Traumatic chiasmal syndrome: a series of 19 patients. AB - PURPOSE: To present a clinical series of 19 patients with traumatic chiasmal syndrome. METHODS: A retrospective study was performed. This included all patients with traumatic chiasmal syndrome seen in the neuro-ophthalmology clinic at the Royal Adelaide Hospital between January 1970 and January 2000. RESULTS: Of the 19 study patients, most were young males involved in motor accidents. Two thirds had skull fractures. Three-quarters of patients had a final visual acuity of 6/12 or better in at least one eye. Ten patients had a complete optic nerve palsy. The incidence of diabetes insipidus in this study was 37%. The incidence of cranial nerve lesions, hypopituitarism, carotid cavernous fistula, and other deficits were documented. Magnetic resonance imaging and surgical findings were consistent with known mechanisms of chiasmal injury. CONCLUSIONS: Trauma is a rare cause of chiasmal syndrome. Patients with bitemporal field defects should be questioned about prior head injury. In the acute setting, magnetic resonance imaging is the most useful investigation. The treating practitioner should anticipate and treat associated endocrine, ocular motility, and other disorders. Mechanisms of damage to the optic chiasm after trauma include direct tearing, contusion haemorrhage and contusion necrosis. These mechanisms should not be considered mutually exclusive. Unilateral temporal hemianopia with a fellow blind eye is not necessarily the result of chiasmal disruption. PMID- 12121370 TI - Retinal manifestations of ophthalmic artery hypoperfusion. AB - Ophthalmic artery hypoperfusion is a relatively uncommon clinical entity. This study illustrates the posterior segment findings of ophthalmic artery hypoperfusion in a series of nine patients. Colour photographs and relevant fluorescein angiograms highlighting the findings are shown. The retinal manifestations of ophthalmic artery hypoperfusion in this series of patients include midperipheral haemorrhages, dilated retinal veins, optic disk collaterals, optic disk neo-vascularization, cotton wool spots, grey intraretinal lesions, fundus pallor, optic disk swelling and choroidal infarcts. Recognition of the ophthalmic changes in this condition may lead to detection of carotid artery disease, the surgical and medical treatment of which has important bearing on patient management. PMID- 12121369 TI - Orbital lipomatous haemangiopericytoma: an unusual variant. AB - Haemangiopericytoma (HPC) of the orbit is a rare tumour occurring in all age groups and presenting with slowly progressive proptosis, ocular motility impairment and visual loss. Although most are benign tumours that are resectable at diagnosis, one-third of these tumours demonstrate malignant features and recur locally or metastasize, usually with a fatal outcome. In 1995, a morphologically unique variant, termed lipomatous haemangiopericytoma (L-HPC), was described. Only one case has been reported previously arising in the orbit. In this paper, a case is reported of L-HPC occurring in the orbit and consider the prognostic implications of this HPC variant. PMID- 12121371 TI - Acquired hyperopia with choroidal folds. AB - Acquired hyperopia with choroidal folds is an uncommon but well recognized condition that is often benign and classically shows flattening of the posterior pole of the affected eye, variable enlargement of the optic nerve complex, and a space between the optic nerve and its sheath. A case is presented that shows the perineural space is best seen on magnetic resonance imaging. Lumbar puncture to exclude increased intracranial pressure should be performed even in the absence of papilloedema. PMID- 12121372 TI - Orbital foreign body misdiagnosed as superior orbital rim fracture. AB - Orbital foreign bodies may be difficult to diagnose clinically and radiologically. In cases where a foreign body is suspected, both the mechanism of injury and the composition of the offending material need to be taken into account. A case is described of an orbital foreign body misdiagnosed as a superior orbital rim fracture, resulting in persistent ptosis and diplopia, and leading to delayed recovery for the patient, a commercial airline pilot. PMID- 12121373 TI - Rapid seroconversion to Treponema pallidum and HIV positivity in a patient with retinal vasculitis. AB - A patient with idiopathic retinal vasculitis was found to have rapid conversion of serological tests for Treponema pallidum and HIV. The diagnosis was confirmed by Venereal Disease Research Laboratory (VDRL) testing and dark ground illumination for T. pallidum, and enzyme-linked immunosorbent assay (ELISA) and Western blot assay tests for HIV. Following treatment with intravenous penicillin, the fundus lesions resolved and visual acuity recovered from inaccurate light projection to 6/24 in the right eye and from counting fingers close to face to 2/60 in the left eye. Serological tests for T. pallidum in patients with concurrent HIV infection may be unpredictable. Hence, it is important to repeat these tests even in the early treatment phase of patients with retinal vasculitis who have shown initial seronegativity. This enables earlier diagnosis and initiation of specific treatment. PMID- 12121374 TI - Familial amyloidotic polyneuropathy presenting with rubeotic glaucoma. AB - A 78-year-old man with familial amyloidotic polyneuropathy type I (Met30), presented with rubeotic glaucoma 9 months following an uncomplicated vitrectomy for vitreous amyloidosis. There was retinal neovascularization and extensive retinal vascular closure. In the preceding 9 months, episodes of 'uveitis' and high intraocular pressure are thought to be due to amyloid protein released into the aqueous leading to trabecular meshwork obstruction and high intraocular pressures, thus compounding the ocular ischaemia created by amyloid vascular closure. The patient underwent pan-retinal photocoagulation and Molteno implant surgery. The rubeosis regressed and pressure control was gained but sight was lost. PMID- 12121375 TI - Garlic and the strabismus surgeon. AB - Potential side-effects caused by complementary medicines are often not anticipated. A case is reported of bilateral retro-bulbar haemorrhages with elevated intraocular pressure during strabismus surgery that occurred as a result of odourless garlic tablet ingestion prescribed by a naturopath. A near catastrophic event was averted with rapid recognition and medical treatment. This is believed to be the first such report of haemorrhaging in ophthalmic surgery induced by garlic. It is concluded that unregulated naturopathic prescribing is potentially dangerous; doctors need to ask specifically about naturopathic potions. PMID- 12121376 TI - Pituitary apoplexy presenting with an orbital bruit. AB - Pituitary apoplexy is a sight-threatening syndrome in which a pituitary adenoma undergoes sudden enlargement as a result of haemorrhage, infarction or both. Classic features of the syndrome include sudden severe headaches, reduced consciousness, visual impairment, ophthalmoplegia and/or endocrinological disturbance. Pituitary apoplexy has been reported following cardiac bypass surgery. The case is reported of a 68-year-old man who presented with left external and internal ophthalmoplegia, complete ptosis, mild chemosis, reduced vision, and an orbital bruit following coronary artery bypass grafting. Carotid angiography showed the left internal carotid artery to be bowed anteriorly and narrowed. Magnetic resonance imaging demonstrated features consistent with pituitary apoplexy. It is believed that an orbital bruit has not previously been reported in pituitary apoplexy. PMID- 12121377 TI - Analysis of the United Kingdom solar eclipse public health campaign 1999. PMID- 12121378 TI - Holistic impressions of an ophthalmologist having cataract surgery himself. PMID- 12121379 TI - Cost of pterygium. PMID- 12121391 TI - Diseases of the generative nail apparatus. Part II: nail bed. AB - Nail bed insults may affect the resultant nail product in a number of ways. If focal in nature, the severity and duration of the insult will determine the appearance of the nail product. Widespread insults may alter the size, shape or colour of the nail plate. Nail bed insults tend to present with more immediacy than nail matrical insults, as there is no need to wait until the nail grows out until the sign is obvious. We are less interested in the history of the event, than we are with the clinical sign being presented to us. Should treatment be required for focal nail bed insults, treatment is likely to be local and curative. Widespread nail bed insults may be associated with systemic diseases and if a result of local problems these may be difficult to treat. PMID- 12121392 TI - Teledermatology: influence of zoning and education on a clinician's ability to observe peripheral lesions. AB - Teledermatology can benefit rural and remote communities, where specialist dermatological services may not be readily available. Regarding store-and- forward teledermatology, we hypothesized that the site of a lesion in an image (zoning) may influence a clinician's ability to observe target lesions, and that education on image viewing may improve use of this technology. We examined this by conducting both pre- and post-education studies. The education on image viewing consisted of a presentation on the outcome of the first study-survey on image viewing. The first study demonstrated that zoning influences a clinician's visual attention and that significant, concurrent lesions in the periphery may be missed. The second study demonstrated that brief education could produce a measurable change in observing peripheral lesions. These findings have medico legal implications and suggest that further education in the use of such technology is necessary in order to optimize patient care and prevent potential errors. PMID- 12121393 TI - Treatment of interdigital tinea pedis with 25% and 50% tea tree oil solution: a randomized, placebo-controlled, blinded study. AB - Tea tree oil has been shown to have activity against dermatophytes in vitro. We have conducted a randomized, controlled, double-blinded study to determine the efficacy and safety of 25% and 50% tea tree oil in the treatment of interdigital tinea pedis. One hundred and fifty-eight patients with tinea pedis clinically and microscopy suggestive of a dermatophyte infection were randomized to receive either placebo, 25% or 50% tea tree oil solution. Patients applied the solution twice daily to affected areas for 4 weeks and were reviewed after 2 and 4 weeks of treatment. There was a marked clinical response seen in 68% of the 50% tea tree oil group and 72% of the 25% tea tree oil group, compared to 39% in the placebo group. Mycological cure was assessed by culture of skin scrapings taken at baseline and after 4 weeks of treatment. The mycological cure rate was 64% in the 50% tea tree oil group, compared to 31% in the placebo group. Four (3.8%) patients applying tea tree oil developed moderate to severe dermatitis that improved quickly on stopping the study medication. PMID- 12121394 TI - Survey of phototherapy practice by dermatologists in Australia. AB - A postal survey was sent to all dermatologists in Australia to determine current phototherapy practices. Questionnaires were returned by 158 (57%) of 277 dermatologists, of whom 112 (71%) provided phototherapy. Large variations existed in attitudes and practice, including indications, contraindications, dosage schedules, equipment maintenance, response to adverse events, and follow-up arrangements. Cumulative ultraviolet (UV) doses for psoralen and UVA (PUVA) were not calculated by 21%, while 30% did not calculate cumulative doses for UVB. Written informed consent was not obtained by 32%. Phototherapist dermatologists reported 25 patients developing melanoma following PUVA. Only 30% of Australian dermatologists organize regular follow up of patients after phototherapy. Australians have the highest rates of melanoma and non-melanoma skin cancers in the world, because of their ancestry and high solar exposure. This makes it inappropriate for Australian dermatologists to rely entirely on foreign safety data when assessing the risks and benefits of phototherapy in Australian patients. There is a need for standardized Australian guidelines that can be prospectively assessed to ensure phototherapy is used to maximize efficacy and minimize risks in Australian patients, given their unique ancestral mix and outdoor lifestyle. PMID- 12121395 TI - Efficacy, cutaneous tolerance and cosmetic acceptability of desonide 0.05% lotion (Desowen) versus vehicle in the short-term treatment of facial atopic or seborrhoeic dermatitis. AB - The differences between topical corticosteroids are based mainly on their potency, safety and patient acceptability. The aim of this study was to evaluate a mild- to mid-potent topical corticosteroid, desonide 0.05%, on these three parameters in an Australian cohort of patients with facial seborrhoeic or atopic dermatitis. Eighty-one adult patients were randomized to receive desonide 0.05% lotion or its vehicle, applied twice daily for 3 weeks under double-blind conditions. In the active treatment group, 88% of patients had their skin condition cleared or almost cleared and only two patients experienced cutaneous adverse events (rash and pruritus). The acceptability of the lotion was high; 95% of patients stated they would use this topical corticosteroid again. These data support the short-term use of desonide 0.05% lotion as a suitable agent for the short-term treatment of facial dermatitis. PMID- 12121396 TI - Idiopathic calciphylaxis. AB - A 68-year-old woman presented with a painful necrotic ulcer on her right calf and necrotic breakdown of a left below-knee amputation stump as a result of calciphylaxis. No cause could be identified and corrected. Treatment comprised wound care, substituting low molecular weight heparin for warfarin, hyperbaric oxygenation and etidronate disodium, none of which were helpful and major limb amputations were considered the best option for this patient. PMID- 12121397 TI - Fatal group A streptococcal necrotizing fasciitis and toxic shock syndrome in a patient with psoriasis and chronic renal impairment. AB - A 78-year-old woman presented with rapid onset of skin pain which evolved into oedema, discoloration and infarction. She was diagnosed with group A beta haemolytic streptococcus (Streptococcus pyogenes) necrotizing fasciitis and streptococcal toxic shock syndrome. The patient had a past history of psoriasis and end-stage renal impairment. Despite treatment with multiple antibiotics in an intensive care unit, the skin infarction involving the upper trunk continued to expand and the patient died within 24 hours of hospital admission. Group A streptococcus and Staphylococcus aureus were cultured from a tissue biopsy. Renal failure and compromised skin barrier function are known to predispose to invasive streptococcal infections, but necrotizing fasciitis has only rarely been reported in association with psoriasis. This case illustrates the fulminant nature of the infection. PMID- 12121398 TI - Staphylococcal scalded skin syndrome complicating acute generalized pustular psoriasis. AB - A 60-year-old woman with psoriasis vulgaris treated with oral cyclosporin and acitretin developed an acute generalized pustular eruption with erythema and associated fever consistent with acute generalized pustular psoriasis. She was admitted to hospital and, despite intravenous fluid replacement, developed acute renal failure. In addition, she developed staphylococcal septicaemia. After transfer to the intensive care unit because of deteriorating renal function, a sudden onset of widespread flaccid blistering (Nikolsky sign positive) and superficial erosions was noted. Histology of a biopsied blister revealed subcorneal splitting of the epidermis consistent with staphylococcal scalded skin syndrome. The patient was treated with intravenous dicloxacillin and the blistering gradually improved over 10 days. PMID- 12121399 TI - Late-onset warfarin necrosis. AB - A 43-year-old woman developed tenderness and induration of her thighs and lower abdomen, 56 days after commencing warfarin for aortic and mitral valve replacements. Investigations showed elevated inflammatory markers, mild renal impairment, normal echocardiogram and low protein C and S levels consistent with warfarin therapy. Three weeks later, purpuric areas evolved into large tender haemoserous bullae, which broke down to form ulcers. Histology confirmed the clinical impression of warfarin-induced skin necrosis with dermal and subcutaneous venular thrombi. Despite cessation of warfarin and commencement of heparin, the lesions progressed. When the patient became febrile, blood cultures grew Pseudomonas aeruginosa, which was treated with intravenous imipenem and vancomycin. Wound swabs grew methycillin-resistant Staphylococcus aureus and the antibiotics were changed. The patient developed septic shock and, despite intensive care management, her condition deteriorated and she died 9 weeks after the onset of the skin symptoms. PMID- 12121400 TI - Stage IV CD30+ anaplastic large cell lymphoma: response to acitretin and interferon-alpha. AB - Retinoids and interferon (IFN)-alpha induce differentiation, affect cell proliferation and alter various immune parameters. In combination, their effects may be additive or even synergistic in the treatment of malignancy. We present a 53-year-old woman with stage IV CD30+ anaplastic large cell lymphoma with brain, lung and skin involvement. The patient had been on methotrexate for rheumatoid arthritis. After a combination of oral acitretin 50 mg daily and IFN-alpha 3 million units subcutaneously 3 times per week, the skin lesions cleared within 2 months, lung lesions by 5 months and brain lesions by 7 months. Although we cannot exclude that methotrexate played a role in the development of this lymphoma and that its withdrawal contributed to the clearance of lesions, we propose that the patient's disease responded to the combination of acitretin and IFN-alpha. PMID- 12121401 TI - Allergic contact dermatitis following exposure to essential oils. AB - Allergic contact dermatitis from the topical use of essential oils is not widely recognized as an occupational hazard. Four cases of allergic contact dermatitis to essential oils occurring in three aromatherapists and one chemist with a particular interest in aromatherapy are described. All presented with predominantly hand dermatitis and demonstrated sensitization to multiple essential oils. One patient developed a recurrence of cutaneous symptoms following ingestion of lemongrass tea. Workers within this industry should be aware of the sensitization potential of these products and the risk of limiting their ability to continue employment. PMID- 12121402 TI - Short-term resolution of psoriasis after total thyroidectomy for euthyroid multinodular goitre. AB - A 36-year-old Chinese female with an 8-year history of chronic, generalized plaque psoriasis demonstrated a marked improvement of the disease after removal of an intercurrent euthyroid multinodular goitre. Thyroxine was commenced immediately postoperatively. No thyroid antibodies were detected and thyroid function and calcium levels remained within normal limits both pre- and postoperatively. Four weeks following surgery, narrow-band ultraviolet B (nbUVB) therapy was recommenced for recurrent psoriasis. The manifestations of psoriasis at this stage were less severe than before thyroidectomy and responded well to treatment, whereas before surgery the response to therapy had been poor. One year following total thyroidectomy, the patient received very effective psoriasis control with nbUVB therapy. The possible role of surgery and thyroid hormones in altering the pathogenesis of psoriasis in the acute setting is clearly of interest and warrants further research consideration. PMID- 12121403 TI - Localized cutaneous necrosis associated with the antiphospholipid syndrome. AB - A 34-year-old woman with systemic lupus erythematosus and high titres of antiphospholipid antibodies was admitted to hospital suffering a viral illness but developed haemorrhagic and necrotic areas on the neck and anterior chest 7 days following cessation of warfarin. Anticoagulation had been initiated following a retinal vein thrombosis, but was ceased on day 4 of admission when she was found to be excessively anticoagulated (international normalized ratio (INR) > 10). However, at the time of developing the cutaneous lesions, the INR was sub-therapeutic. Histology of a skin biopsy from the neck revealed thrombosis of upper dermal blood vessels without vasculitis, consistent with antiphospholipid antibody-related skin necrosis. This case illustrates one of the cutaneous features that can occur in patients with elevated titres of antiphospholipid antibodies and the importance of closely monitoring anticoagulation in such patients. PMID- 12121404 TI - Lupus panniculitis clinically simulating alopecia areata. AB - A 27-year-old woman with a known history of lupus erythematosus presented with two circumscribed patches of non-scarring alopecia closely resembling alopecia areata. Scalp biopsy showed a predominantly subcutaneous and deep dermal lymphocytic infiltrate that surrounded the deep follicular segments and hair bulbs, as well as the eccrine glands. There was associated hyaline fat sclerosis. The epidermis, infundibular and isthmus segments of follicles were relatively spared and lacked the lichenoid inflammation and fibrosis seen with lupus erythematosus. The biopsy findings illustrate that the deep variant of lupus panniculitis may be concentrated around the hair bulbs and deep temporary segments of hair follicles and spare the permanent stem cell-rich follicular segments. This pattern is capable of producing a temporary hair-loss, clinically simulating alopecia areata. The clinical history, presence of subtle erythema and scalp tenderness on physical examination, as well as the biopsy findings, were important clues in distinguishing our case from a true combination of alopecia areata and lupus erythematosus. PMID- 12121405 TI - Contact dermatitis to phenol-formaldehyde resin in two plywood factory workers. AB - Two cases of occupational allergic contact dermatitis in plywood mill workers are presented. Although both patients thought they were allergic to sawdust, they were in fact allergic to a phenol-formaldehyde resin used to bond the plywood sheets together. Both patients only developed their dermatitis when they came into contact with uncured glue. PMID- 12121406 TI - Peptides in the treatment of inflammatory skin disease. PMID- 12121408 TI - The needs of cancer patients and their families from rural and remote areas of Queensland. AB - This study examines the impact of travelling for treatment on cancer patients and their families. Twenty-eight consecutive cancer patients, who were receiving radiation therapy treatment and 19 family carers, completed a structured needs assessment questionnaire and an in-depth interview. Both patients and carers reported moderate to high levels of unmet psychological need. Carers were found to have higher levels of anxiety than patients, although both groups had higher anxiety levels than the general population. Taking more responsibility for household tasks and organising new living arrangements for the family were the most frequently identified demands of a dual burden of caring. Nearly 40% of carers reported some disruption to their schedule and half reported experiencing financial difficulties. The qualitative interviews highlight the disruption that parents and children experience under the present system, particularly in relation to the demands of family life and the need to maintain some level of continuity and security for children. PMID- 12121409 TI - Identifying and overcoming the barriers to Aboriginal access to general practitioner services in Rural New South Wales. AB - The Wiradjuri General Practitioners and Aboriginal Health Workers Project aimed to help improve Aboriginal health in central western New South Wales (NSW) by identifying and overcoming the barriers to the Aboriginal population's access to general practitioner services. The central strategy of the project was to convene three rounds of consultative meetings in five towns: Bathurst, Orange, Cowra, Forbes and Condobolin. These meetings brought together Aboriginal community members, general practitioners and Aboriginal health workers to express and define local issues and problems, and to propose solutions. The solutions included general practitioner outreach clinics, a focus on prewinter immunisation, bulk-billing of Aboriginal patients, Aboriginal cultural awareness training for all general practice personnel, employing Aboriginal staff in general practice, and closer professional interaction between general practitioners and Aboriginal health workers. Most participants evaluated the meetings as positive and constructive. The project substantiates the perceived need for alternative models for the funding and delivery of general practitioner services to Aboriginal communities. PMID- 12121410 TI - Comparison of roles and professional development needs of nurse executives working in metropolitan, provincial, rural or remote settings in Queensland. AB - This research provides a profile of nurse executives' roles, career opportunities and professional development needs across metropolitan, provincial, rural and remote settings in Queensland. A cross-sectional survey was posted to all Directors/Assistant Directors of Nursing in the Queensland Public Health Sector (n = 281), with a response rate of 52.3% (n = 147). Findings indicated that the role of nursing executives has expanded and diversified, with multiple role responsibilities increasing with distance from the metropolitan area. Nurse executives in remote areas were less satisfied with the quality of supervision and mentorship they received, and least likely to participate in career enhancing activities. Metropolitan nursing executives utilised more of the career enhancing opportunities provided by the Queensland Public Health Sector than did provincial, rural or remote nursing executives. Provincial nurse executives were the group most likely to network, attend conferences and participate in informal educational activities. Professional development needs, although generally common to all groups, were more practice specific for those in rural and remote areas. Many nurse executives begin their careers in rural or remote areas where limited opportunities for career development may be detrimental to their future development. PMID- 12121411 TI - Farm-related fatal injury of young and older adults in Australia, 1989-1992. AB - This paper describes the types of, and circumstances surrounding, unintentional farm-related fatal injuries involving young and older adults in Australia. Information was obtained from an inspection of coronial files for the period 1989 1992. Around 14% of all farm-related fatalities in Australia during 1989-1992 were of young adults aged 15-24 years and approximately one-quarter were of older adults aged > or = 55 years. Young adults were commonly fatally injured in motor vehicle incidents and in incidents involving firearms. Tractors were the most common agent involved in fatal incidents involving older adults. Intervention measures to prevent fatalities of older adults in agriculture should focus on the safe use of tractors, while for young adults it appears prevention efforts should centre around safe use of firearms and operation of motor vehicles on the farm. Ways to overcome barriers to the use of injury prevention measures in rural Australia should be further explored. PMID- 12121412 TI - Increasing rural activity and curriculum content in the Adelaide University Medical School. AB - This study aimed to document the level of rural activity and curriculum content in the Adelaide University Medical School. A questionnaire was distributed to all heads of departments within the Medical School and additional information was obtained from reports and discussions with key personnel. There has been an increase in the proportion of students with a rural background enrolled from 9% in 1994 to 22% in 2000. There has also been an increase in the number of weeks available for rural placements from 12 (mostly optional) to 29 weeks (some mandatory), and an increase in the number of departments offering rural placements. There has also been improved academic support to rural practitioners and many departments directly provide services in rural communities. A new combined University Department of Rural Health (UDRH)/Rural Clinical School associated with the Adelaide University Medical School aims to provide at least half of all clinical training to 25% of all medical students of Australian origin. PMID- 12121413 TI - Long-term employment outcomes in a rural area following traumatic brain injury. AB - One of the major sequelae following traumatic brain injury is a change in employment status. This poses significant challenges for rehabilitation services. Several studies have investigated the longer-term outcome after traumatic brain injury. Longer-term outcome in an economically disadvantaged rural environment with limited rehabilitation services has not been studied extensively. A group of individuals (n = 65) who sustained a traumatic brain injury were compared regarding pre- and post-employment status. The moderate and severely injured sub groups showed a significant change in employment status. Ideally rehabilitation programs should target re-employment as an outcome. Limited employment opportunities in rural areas may make this more difficult to achieve. PMID- 12121414 TI - Clathrin-protein interactions. AB - There is a complex network of protein-protein and protein-lipid interactions that underlie clathrin-mediated vesicular traffic in all compartmentalized cells from yeast to man. Major progress has been made in the determination of the three dimensional structures of many of the components. Recently, there has been an explosion in the identification and characterization of clathrin binding partners. This review integrates the structural and biochemical information that is currently available to present a unified view of how many clathrin binding partners interact with clathrin. PMID- 12121415 TI - The Golgi apparatus: balancing new with old. AB - Most models put forward to explain cellular processes do not stand the test of time. The 'lucky' few that are able to survive extensive experimental tests and peer critique may eventually become dogmas or paradigms. When this happens, the amount of experimental data required to overturn the paradigm is extensive. To some, such inertia may seem prohibitive to scientific progress but rather, in our opinion, this helps to maintain a degree of coherence. It is needed so that experiments and interpretations may be conducted within relatively safe boundaries. In the field of protein transport in the secretory pathway, we have enjoyed a relatively stable and productive period for quite some time (more than 30 years!). It is only very recently that the field has entered into a phase where all bets seem to be off. As in any paradigm shift, the accumulation of experimental observations inconsistent with the old dogma eventually reached a critical point. As we 'reluctantly' dispense with the long-standing paradigm of forward vesicular transport, we face a time that is bound to be trying as well as exciting. PMID- 12121417 TI - Cargo selection in vesicular transport: the making and breaking of a coat. AB - Intracellular traffic is mediated by vesicular/tubular carriers. The carriers are formed by the activity of cytosolic coat proteins that are recruited to their target membranes and deform these membranes into buds and vesicles. Specific interactions between recruited coat subunits and short peptide sequences (transport motifs) on cargo proteins direct the incorporation of cargo into budded vesicles. Here, we focus on cargo selection reactions mediated by COPII and AP-2/clathrin vesicle coat complexes to explore common mechanisms by which coat assembly support localized and selective cargo sorting. Recent findings suggest that multiple, low-affinity interactions are employed in a cooperative manner to support coat assembly and enable cargo recognition. Thus low-binding affinities between coat subunits and transport motifs are transiently transformed into high-avidity, multivalent and selective interactions at vesicle bud sites. The temporal and regulated nature of the interactions provide the key to cargo selection. PMID- 12121416 TI - Protein dislocation from the endoplasmic reticulum--pulling out the suspect. AB - Proteins that fail to fold properly as well as constitutive or regulated short lived proteins of the endoplasmic reticulum are subjected to proteolysis by cytosolic 26S proteasomes. This process is known as endoplasmic reticulum associated protein degradation. In order to become accessible to the proteasome of this system substrates must first be retrogradely transported from the endoplasmic reticulum into the cytosol, in a process termed dislocation. This export step seems to be accompanied by polyubiquitination of such molecules. Surprisingly, protein dislocation from the endoplasmic reticulum seems to require at least some components that mediate import into this compartment. However, protein import and export display differences in the mechanism that provides the driving force and ensures directionality. Of special interest is the cytoplasmic Cdc48p/Npl4p/Ufd1p complex, which is required for the degradation of various endoplasmic reticulum-associated protein degradation substrates and seems to function in a step after polyubiquitination but before proteasomal digestion. In this review, we will summarize our knowledge on protein export during endoplasmic reticulum-associated protein degradation and discuss the possible function of certain components involved in this process. PMID- 12121418 TI - Accumulation of a GPI-anchored protein at the cell surface requires sorting at multiple intracellular levels. AB - Proteins modified by glycosylphosphatidylinositol membrane anchors have become popular for investigating the role of membrane lipid microdomains in cellular sorting processes. To this end, trypanosomatids offer the advantage that they express these molecules in high abundance. The parasitic protozoan Trypanosoma brucei is covered by a dense and nearly homogeneous coat composed of a glycosylphosphatidylinositol-anchored protein, the variant surface glycoprotein, which is essential for survival of the parasite in the mammalian blood. Therefore, T. brucei must possess mechanisms to selectively and efficiently deliver variant surface glycoprotein to the cell surface. In this study, we have quantified the steady-state distribution of variant surface glycoprotein by differential biotinylation, by fluorescence microscopy and by immunoelectron microscopy on high-pressure frozen and freeze-substituted samples. These three techniques provide very similar estimates of the fraction of variant surface glycoprotein located on the cell surface, on average 89.4%. The intracellular variant surface glycoprotein (10.6%) is predominantly located in the endosomal compartment (75%), while 25% are associated with the endoplasmic reticulum, Golgi apparatus and lysosomes. The density of variant surface glycoprotein in the plasma membrane including the membrane of the flagellar pocket, the only site for endo- and exocytosis in this organism, is 48-52 times higher than the density in endoplasmic reticulum membranes. The relative densities of the Golgi complex and of the endosomes are 2.7 and 10.8, respectively, compared to the endoplasmic reticulum. This data set provides the basis for an analysis of the dynamics of sorting. Depending on the intracellular itinerary of newly formed variant surface glycoprotein, the high surface density is achieved in two (endoplasmic reticulum -> Golgi complex --> cell surface) or three enrichment steps (endoplasmic reticulum --> Golgi complex --> endosomes --> cell surface), suggesting sorting between several membrane compartments. PMID- 12121419 TI - Peroxisome remnants in pex3delta cells and the requirement of Pex3p for interactions between the peroxisomal docking and translocation subcomplexes. AB - During peroxisomal matrix protein import, the peroxisomal targeting signal receptors recognize cargo in the cytosol and interact with docking and translocation subcomplexes on the peroxisomal membrane. Using immunoprecipitations of multiple protein components, we show that in Pichia pastoris the docking subcomplex consists of the unique peroxins Pex13p, Pex14p and Pex17p, whereas the putative translocation subcomplex has all three RING finger peroxins, Pex2p, Pex10p and Pex12p, as unique constituents. We identify Pex3p as a shared component of both subcomplexes. In pex3delta cells, the unique constituents of the docking subcomplex interact as they do in wild-type cells, but the assembly of the translocation subcomplex is impaired and its components are present at reduced levels. Furthermore, several interactions detected in wild type cells between translocation and docking subcomplex components are undetectable in pex3delta cells. Contrary to previous reports, pex3delta cells have peroxisome remnants that pellet during high-speed centrifugation, associate with membranes on floatation gradients and can be visualized by deconvolution microscopy using antibodies to several peroxins which were not available earlier. We discuss roles for Pex3p in the assembly of specific peroxisomal membrane protein subcomplexes whose formation is necessary for matrix protein import. PMID- 12121420 TI - EHD3: a protein that resides in recycling tubular and vesicular membrane structures and interacts with EHD1. AB - Here we report the characterization of an eps15 homology (EH) domain containing protein designated EHD3. EHD3 was mapped to human chromosome 2p22-23, while the murine Ehd3 homolog was mapped to chromosome 17p21. Both the human and the mouse genes contain a polymorphic (CA) repeat in their 3'UTR. One 3.6-kb Ehd3 transcript was mainly detected in adult mouse brain and kidney and at day 7 of mouse development. On the other hand, human tissues exhibited two, 4.2- and 3.6 kb, EHD3 RNA species. They were predominantly expressed in heart, brain, placenta, liver, kidney and ovary. EHD3, expressed as a green fluorescent fusion protein was localized to endocytic vesicles and to microtubule-dependent, membrane tubules. There was a clear colocalization of EHD3-positive structures and transferrin-containing recycling vesicles, implying that EHD3 resides within the endocytic recycling compartment. Shuffling the N-terminal domain of EHD1 (previously shown to reside in the transferrin-containing, endocytic recycling compartment) with that of EHD3 resulted in a chimeric EHD protein that was localized mainly to tubules instead of the endocytic vesicles, implicating the N terminal domain as responsible for the tubular localization of EHD3. Mutant EHD3 forms, missing the N-terminal or the C-terminal domains, lost their tubular localization. Results of two-hybrid analyses indicated that EHD1 and EHD3 interact with each other. In addition, EHD1 and EHD3 could be coimmunoprecipitated from cellular extracts, confirming the interaction implied by two-hybrid analysis. Moreover, coexpression of EHD1 and EHD3 resulted in their colocalization in microtubule-dependent tubules as well as in punctate forms. Based on its specific intracellular localization and its interaction with EHD1, we postulate that EHD3 localizes on endocytic tubular and vesicular structures and regulates their microtubule-dependent movement. PMID- 12121421 TI - The mu2 subunit of the clathrin adaptor AP-2 binds to FDNPVY and YppO sorting signals at distinct sites. AB - The endocytic sorting signal on the low-density lipoprotein receptor for clathrin mediated internalization is the sequence FDNPVY in the receptor's cytosolic tail. We have used a combination of surface plasmon resonance and crosslinking with a photoactivated peptide probe to demonstrate the interaction between FDNPVY containing peptides and the mu2 chain of purified AP-2 clathrin adaptors (the complexes responsible for plasma membrane sorting). We show that recognition of the FDNPVY signal is mediated by a binding site in the mu2-subunit that is distinct from the site for the more general YppO sorting signal, another tyrosine based sequence also recognized by mu2-adaptin. These results suggest the possibility that low-density lipoprotein receptor uptake may be modulated specifically and independently of other proteins in the clathrin pathway. PMID- 12121422 TI - Anterior cruciate ligament injuries in young females playing soccer at senior levels. AB - The aim of this investigation was to study the consequences of anterior cruciate ligament injuries in female soccer players. Special interest was focused on young female soccer players (< 16 years) sustaining anterior cruciate ligament injuries when playing at a senior level, which means playing together with players 19 years or older. In Sweden, all players belonging to an organized soccer club are insured by the same insurance company, the Folksam Insurance Company. Data of all soccer-related knee injuries in females reported to the Folksam Insurance Company between 1994 and 1998 were collected. A questionnaire was sent to 978 females who were registered to have sustained a knee injury before the age of 20 years. The response rate was 79%. Three hundred and ninety-eight female soccer players who had sustained an anterior cruciate ligament injury before the age of 19 years were analysed. Most of their anterior cruciate ligament injuries had been diagnosed using arthroscopy or magnetic resonance imaging (84%). Thirty-eight percent of the players had been injured before the age of 16 years. Of these, 39% were injured when playing in senior teams. When playing in senior teams 59% of the players below the age of 16 years and 44% of the players 16 years or older sustained their ACL injuries during contact situations. At the time of this investigation (2-7 years after the anterior cruciate ligament injury), altogether 78% (n = 311) reported that they had stopped playing soccer. The most common reason (80%) was symptoms from their anterior cruciate ligament-injured knee. It appears that many young female soccer players injure their anterior cruciate ligament when playing at a senior level. Therefore, we suggest that female soccer players under the age of 16 years should be allowed to participate only in practice sessions but not games at a senior level. PMID- 12121423 TI - Arthroscopic treatment of impingement of the ankle reduces pain and enhances function. AB - A consecutive series of 105 patients with a median age of 35 (16-62) years who were operated on with arthroscopic resection for impingement of the ankle using standardized technique without distraction is presented. All patients complained of painful dorsiflexion and had failed to respond to conservative treatment. A total of 177 diagnoses were found, soft tissue impingement or synovitis in 89, anterior bony impingement in 44, chondral lesion in 20, loose bodies in 16 and osteoarthritis in eight. At follow-up after 2 years, 65 patients were pain free while 28 patients had experienced reduction of pain. Gait was improved in 30/41 patients and 22 resumed sporting activities. The results were graded excellent in 67, good in 25, fair in six and poor in seven patients. There were four deep infections and one synovial fistula in this series. The deep infections all responded well to arthroscopic synovectomy and intravenous antibiotics. In one patient persistent symptoms were recorded. Ankle arthroscopy yielded good results in the treatment of anterior impingement of the ankle as it effectively reduced pain and enhanced function. PMID- 12121424 TI - Range of motion training in brace vs. plaster immobilization after anterior cruciate ligament reconstruction: a prospective randomized comparison with a 2 year follow-up. AB - The purpose of this prospective and randomized study was to compare rehabilitation with early range of motion (ROM) training vs immobilization following anterior cruciate ligament (ACL) reconstruction. Fifty patients, undergoing an ACL reconstruction with a bone-patellar tendon-bone graft, were postoperatively allocated randomly to either a plaster cast or a brace for 5 weeks. The brace group had ROM exercises from postoperative day 7. The commencement of ROM exercises was postponed 4 weeks for the plaster group compared to the brace group, but progressed subsequently with equal speed. There was no difference between the groups in the ROM of flexion or extension 20 weeks after the ACL reconstruction and later. Twenty-four months after surgery, the muscle strength deficit in the hamstring muscles (isokinetic measurements; percent difference, injured vs uninjured) was significantly larger in the brace group (mean +/- SD: 5.9 +/- 7.8%, P < 0.01) than in the plaster group (- 0.9 +/- 11.8%, NS) (brace vs plaster group, P < 0.05). Furthermore, there was also a tendency in the brace group to a larger strength deficit in the quadriceps muscle (brace: 11.1 +/- 13.2%, P < 0.001; plaster: 3.8 +/- 12.9%, NS) (brace vs plaster group, P= 0.07). There was no difference between the groups in the total sagittal knee laxity, as measured with an arthrometer, or in the subjective knee function or activity level (Lysholm score together with the Tegner activity level) between the groups. It is concluded that the postoperative treatment with early range of motion training after ACL reconstruction gave as good ROM, knee stability, subjective knee function and activity level as the treatment with immobilization. It is hypothesized that the larger strength deficit observed after rehabilitation with early range of motion training is secondary to the more intensive training and physical therapist involvement that was demanded in order to achieve full ROM following immobilization. PMID- 12121425 TI - Estimated moments at L5/S1 level and muscular activation of back extensors for six prone back extension exercises in healthy individuals. AB - The purpose of this study was to identify the prone back extension (PBE) exercises that offer the most resistance to the back extensor muscles. Twenty males with no previous history of low back injury performed two repetitions of eight exercises. These consisted of two maximal isometric voluntary activations (MVA) and six different PBE exercises. The participants, while lying prone on a bench, were asked to raise either their trunk or legs to the horizontal position and hold for one second (static phase) before returning to the original position. The exercises differed according to the particular segments that were raised into extension. To assess the PBE exercises, a biomechanical model was used to estimate the level of load resistance (%MVA) relative to the maximum voluntary activation strength (MVA). In addition, electromyography (EMG) was included to measure the level of muscle activity (%MVE) relative to the maximum voluntary activation EMG (MVE) value reached during the MVA. A significant relationship of 0.86 was found between the mass of the subject and the peak reaction moment during the MVA. Peak level of load resistance averaged around 50% MVA with the exception of exercise no. 5 where the subject had to raise in extension his trunk, legs and arms (65% MVA). The erector spinae (ES) activity level reached a peak value of 61% MVE for exercise no. 5. In the exercises where only the trunk was lifted, the peak level of ES was below 47% MVE and the average activity during the static phase reached approximately 26% MVE. For healthy individuals PBE exercises can be considered low intensity exercises (< 50% MVA; except exercise no. 5) and can be used to improve the endurance of the back muscles. PMID- 12121426 TI - Load-displacement properties of the human triceps surae aponeurosis and tendon in runners and non-runners. AB - The load-displacement and stress-strain characteristics of the human triceps surae tendon and aponeurosis, in vivo, was examined during graded maximal voluntary plantarflexion efforts in runners who trained 80 km/ week or more and age-matched non-runners. Synchronous real-time ultrasonography of triceps surae tendon and aponeurosis displacement, electromyography of the gastrocnemius, soleus and dorsiflexor muscles, and joint angular rotation were obtained. Tendon cross-sectional area and ankle joint moment arm were obtained from magnetic resonance imaging. Tensile tendon force was calculated from the joint moments and tendon moment arm and stress was obtained by dividing force by cross-sectional area. Strain was obtained from the displacements normalized to tendon length. Antagonist coactivation and small amounts of ankle joint rotation significantly affected tensile tendon force and aponeurosis and tendon displacement, respectively (P < 0.01). Plantarflexion moment was similar in runners (138 +/- 27 Nm, mean +/- SEM) and non-runners (142 +/- 17 Nm). Tendon moment arm was alike in non-runner (58.3 +/- 0.2 mm) and runners (55.1 +/- 0.1 mm). Similarly, there was no difference in tendon tensile force between runners (2633 +/- 465 N) and non runners (2556 +/- 401 N). The cross-sectional area of the Achilles tendon was larger in runners (95 +/- 3 mm2) than non-runners (73 +/- 3 mm(2)) (P < 0.01). The load-deformation data yielded similar stiffness (runners 306 +/- 61 N/mm, non runners 319 +/- 42 N/mm). The maximal strain and stress was 4.9 +/- 0.8% and 38.2 +/- 9.8 MPa in non-runners and 4.1 +/- 0.8% and 26.3 +/- 5.1 MPa in runners. The larger tendon cross-sectional area in trained runners suggests that chronic exposure to repetitive loading has resulted in a tissue adaptation. PMID- 12121427 TI - Walking trials in postmenopausal women: effect of one vs two daily bouts on aerobic fitness. AB - We compared the effects of one vs two daily bouts of walking on aerobic fitness and body composition in postmenopausal women. One hundred and thirty-four subjects were randomized into exercise groups or a control group and 130 completed the study. The subjects walked 5 d/week for 15 weeks at 65% of their maximal aerobic power expending 300 kcal (1255 kJ) in exercise in one (Group S1) or two daily sessions (Group S2). VO(2max) was measured in a direct maximal treadmill test. Body mass index (BMI) was calculated and the percentage of body fat (fat%) estimated using skinfold measurements. The net change in the VO(2max) was 2.5 mL min/kg (95% CI 1.5, 3.5) (8.7%) in Group S1 and 2.5 mL min/kg (95% CI 1.5, 3.5) (8.8%) in Group S2. The net change in body mass was -1.2 kg (95% CI 1.9, -0.5) in Group S1 and -1.1 kg (95% CI -1.8, -0.4) in Group S2. The net fat% change was -2.1% (95% CI-2.7, -1.4) in Group S1 and -1.7% (95% CI-2.3, -1.0) in Group S2. Exercise improved the maximal aerobic power and body composition equally when walking was performed in one or two daily bouts. PMID- 12121428 TI - Prevalence of stress incontinence in nulliparous elite trampolinists. AB - During trampoline jumping the pelvic floor is exposed to high forces. There has been a general belief that physically fit women have a strong pelvic floor as a result of their regular training, thus preventing urinary incontinence. The aim of this study was to survey the prevalence of stress urinary incontinence in female elite trampolinists. The prevalence of urinary incontinence was assessed by a questionnaire, sent to all 35 elite trampolinists (mean age 15, range 12-22 years) in Sweden. Eighty percent of the trampolinists reported involuntary urinary leakage, but only during trampoline training. The leakage started after 2.5 (range 1-4) years of training. Age (P < 0.001), duration of training (P = 0.04), and training frequency (P = 0.01) were significantly associated with leakage. All women above 15 years of age (n = 23) reported urinary leakage (P < 0.001). Eighteen incontinent women continued the study and their leakage was verified by a pad test. The leakage averaged 28 g during a jump session. The muscle strength was measured with perineometry in 10 women and showed good strength in the pelvic floor muscles. PMID- 12121429 TI - Dynamic balance in former elite male athletes and in community control subjects. AB - Dynamic balance was studied in 105 former elite athletes (aged 45-68) and in 966 community control subjects (aged 27-67). Subjects walked barefoot along a 5-m long test track, stepping only on two pads which they alternately put down in front of them and picked up from behind while standing on one foot. A better test result (short completion time) was associated with lower body mass index (in both groups), with jumping height and physical activity during the previous year (in former athletes), and with younger age, better general health, and better perceived physical fitness (in control subjects). Test results in former athletes were, on average, comparable to those of 24-30 years younger community control subjects. PMID- 12121430 TI - Tug of war: introduction to the sport and an epidemiological injury study among elite pullers. AB - The objective of this study was to describe injury patterns among elite tug of war athletes, using a survey method. The setting was the World Tug of War (TOW) Championships in 1998. All 990 male and female athletes asked to participate. Demographic data, participation history, and injury history during TOW, including injury number and type, were collected. A total of 252 (187 males and 65 females) athletes completed the survey (25% response rate). Although males had been competing longer than females (9.3 +/- 6.9 years vs. 6.5 +/- 5.0 years, t-test, P < 0.005), the percentage of males (64/187 = 32%) and females (24/65 = 37%) females reporting TOW injuries was similar. Strains and sprains comprised over half of all injuries, and the back (42%), shoulder-upper limb (23%), and knee (17%) were most commonly injured. Injury patterns were similar among males and females. Elite TOW athletes represent a diverse age range (15-55 years) of males and females who have competed for 5-10 years. About one-third will have suffered past TOW-related injuries, typically involving sprains or strains of the low back, shoulder girdle-upper extremity, and knee. The possibility of an increased injury rate among female TOW athletes warrants further study. Considerations for future research are discussed. PMID- 12121432 TI - Current state of pneumococcal vaccines. AB - Streptococcus pneumoniae is a leading cause of bacterial pneumonia, meningitis, and acute otitis media in children and adults worldwide. According to World Health Organization estimates, at least 1 million children under 5 years of age die each year from pneumococcal pneumonia. The emergence of resistant strains necessitates the development of an effective vaccine with a large serotype coverage. The 11 most common serotypes cause 72-83% of all serious pneumococcal diseases worldwide. Currently marketed 23-valent pneumococcal polysaccharide vaccine provides large serotype coverage and offers a less expensive option. However, it is efficacious only in adults but not in infants. Conjugate vaccines offer a solution by generating immunological memory already at early age. A recently licensed 7-valent conjugate vaccine is immunogenic and efficacious in infants. Its serotype coverage might be sufficient in Europe and North America, but not in Africa, Asia and Oceania. A need exists to develop pneumococcal vaccines with lower cost and larger serotype coverage. Several 11-valent pneumococcal conjugate vaccines are being evaluated in phase I-III trials. This study reviews the current state of pneumococcal problem and pneumococcal vaccines in clinical use. PMID- 12121433 TI - The development of primary and secondary lymphoid tissues in the nurse shark Ginglymostoma cirratum: B-cell zones precede dendritic cell immigration and T cell zone formation during ontogeny of the spleen. AB - Secondary lymphoid tissue and immunoglobulin (Ig) production in mammals is not fully developed at birth, requiring time postnatally to attain all features required for adaptive immune responses. The immune system of newborn sharks - the oldest vertebrate group having adaptive immunity - also displays immature characteristics such as low serum IgM concentration and high levels of IgM1gj, an innate-like Ig. Primary and secondary lymphoid tissues in sharks and other cartilaginous fish were identified previously, but their cellular organization was not examined in detail. In this study of nurse shark lymphoid tissue, we demonstrate that the adult spleen contains well-defined, highly vascularized white pulp (WP) areas, composed of a central T-cell zone containing a major histocompatibility complex (MHC) class II+ dendritic cell (DC) network and a small number of Ig+ secretory cells, surrounded by smaller zones of surface Ig+ (sIg+) B cells. In neonates, splenic WPs are exclusively B-cell zones containing sIgM+-MHC class IIlow B cells; thus compartmentalized areas with T cells and DCs, as well as surface Ig novel antigen receptor (sIgNAR)-expressing B cells are absent at birth. Not until the pups are 5 months old do these WP areas become adult-like; concomitantly, sIgNAR+ B cells are readily detectable, indicating that this Ig class requires a 'mature immune-responsive environment'. The epigonal organ is the major site of neonatal B lymphopoiesis, based on the presence of developing B cells and recombination-activating gene 1 (RAG1)/terminal deoxynucleotidyl transferase (TdT) expression, indicative of antigen receptor rearrangement; such expression persists into adult life, whereas the spleen has negligible lymphopoietic activity. In adults but not neonates, many secretory B cells reside in the epigonal organ, suggesting, like in mammals, that B cells home to this primary lymphoid tissue after activation in other areas of the body. PMID- 12121434 TI - Rat complement factor H: molecular cloning, sequencing and quantification with a newly established ELISA. AB - Factor H (FH) is the predominant soluble regulatory protein of the complement system. With a concentration of 300-600 microg/ml in human plasma it acts as a cofactor for the FI-mediated cleavage of the component C3b to iC3b. Furthermore, it competes with factor B for binding to C3b and C3(H2O) and promotes the dissociation of the C3bBb complex (i.e. it has decay accelerating activity). FH is a monomer of about 155 kDa which comprises 20 short consensus repeats (SCR), each of which is composed of nearly 60 amino acid residues. For the screening of a rat liver cDNA library, we used two hybridization probes which had been produced by polymerase chain reaction (PCR). The probes were generated using degenerated primers which corresponded to conserved parts of the human and the murine factor H nucleotide sequences. The entire rat sequence spanned 4240 nucleotides with an open reading frame of 3708 nucleotides. These were preceded by 23 nucleotides of the 5' untranslated region, followed by a stop codon and a 3' untranslated region of 478 nucleotides including the polyadenylation-signal up to the beginning of the poly A tail. Comparison of the rat cDNA-derived coding sequence revealed identities of 74% to the human and 87% to the mouse FH nucleotide sequence. The translation product of rat FH mRNA was 1236 aa in length (leader sequence included) with an identity of 63% to the human and 81.5% to the murine protein. The degree of glycosylation of rat FH-Mr is about 9.5%. To quantitate FH in rat serum and supernatants of primary cultures of rat hepatocytes (HC), a reliable and sensitive sandwich-enzyme-linked immunosorbent assay (ELISA) was established. The concentration of FH in rat serum was calculated to be 238 microg +/- 21 microg/ml (mean +/- SD). Its concentration in the culture supernatants of HC was upregulated about three-fold by interferon (IFN)-gamma (100 U/ml). PMID- 12121435 TI - Distinct phenotypic adhesion molecule expression on human cord blood progenitors during early Eosinophilic commitment: upregulation of beta(7) integrins. AB - Increasing levels of proinflammatory cells, including eosinophils and basophils, are seen at the site of allergen challenge in allergic disease of the airways. Mechanisms for the recruitment of these cell types could involve either specific upregulation of adhesion molecule and chemoattraction, or the initiation of proliferation and differentiation of inflammatory cell progenitors derived from the bone marrow. In this study, we demonstrate, in two systems of eosinophilic basophilic lineage-committed granulocytes of relative immaturity, that eosinophilic differentiation in vivo implies the induction of a distinct adhesion phenotype, characterized by the upregulation of beta(7) integrin and downregulation of beta(1) and alpha(5) integrins. Moreover, the eosinophilic differentiation induced an upregulation of complement receptor type 1 and type 3, and the expression was further enhanced upon a short-course in vitro activation with ionomycin. These data indicate a sequential alteration of disparate members of the integrin family during eosinophilic-basophilic differentiation, which may attribute to specific adhesion requirements at distinct stages of cell maturation. PMID- 12121436 TI - Restricted specificity of intermolecular spreading to endogenous La (SS-B) and 60 kDa Ro (SS-A) in experimental autoimmunity. AB - Intermolecular spreading of humoral autoimmunity to different components of the Ro (SS-A) and La (SS-B) ribonucleoprotein (RNP) complex has been reported following immunization with a single component of the complex. Although the immune response to the immunizing antigen is polyclonal and diversified, little is known about the specificity of the recruited autoimmune responses to the endogenous Ro and La antigens which drive B-cell spreading. To determine the specificity of intermolecular spreading to La, we examined sera from 52 kDa Ro (Ro52)- and 60 kDa Ro (Ro60)-immunized C3H/HeJ mice for reactivity with recombinant fragments spanning endogenous mouse (m)La by enzyme-linked immunosorbent assay (ELISA) and immunoblotting. Sera from mice primed and boosted with recombinant Ro52 and Ro60 showed reactivity restricted to the COOH-terminal fragment of mLa (aa361-415). The recruited anti-La response was species-specific, cross-reacting weakly with the corresponding region on the human La molecule, and was abrogated by the preabsorption of the Ro-immune sera with mLa 361-415. Analogous experiments using recombinant mRo60 fragments spanning the mRo60 molecule revealed a similar pattern of oligoclonality in the specificity of anti Ro60 autoimmunity following active immunization with La and Ro52. These results suggest that intermolecular-intrastructural T-B help is limiting in this model, and reveal unsuspected immunodominance of selected Ro-La epitopes in the spreading of the autoantibody response to these structures. The focusing of the recruited autoantibody response to these COOH-terminal regions of the Ro and La polypeptides may also reflect the surface accessibility of these regions in La-Ro RNP. PMID- 12121437 TI - Different role of Apaf-1 in positive selection, negative selection and death by neglect in foetal thymic organ culture. AB - Apoptotic protease-activating factor 1 (Apaf-1) is a component of the apoptosome which is required for the activation of procaspase-9. As Apaf-1 knockout (KO) (Apaf-1-/-) mice die before birth, the role of Apaf-1 during thymic selection was investigated using 5 day foetal thymic organ culture (FTOC) of thymi obtained at gestational day 15. There was a lower ratio of CD4 single-positive (SP) to CD8 SP cells and decreased apoptosis of CD4+CD8+ (DP) thymocytes from Apaf-1-/- mice compared with wild-type. To determine if these defects resulted in increased production of neglected thymocytes, the Apaf-1-/- mice were crossed with the T cell receptor (TCR)-alpha-chain KO mice. There was no difference in thymocyte development in the thymi of TCR-alpha-/-Apaf-1-/- and TCR-alpha-/-Apaf-1+/+ mice 5 days after FTOC. To determine if Apaf-1 is involved in apoptosis during death by negative or positive selection, FTOC of the thymus of Apaf-1-/- Db/HY TCR alphabeta transgenic (Tg) mice was carried out. There was decreased apoptosis of the HY clonal-specific M33+ thymocytes and an increased percentage of the autoreactive CD8+M33+ thymocytes in male, but not female Apaf-1-/- Db/HY TCR Tg mice. Our data suggest that Apaf-1 is not involved in positive selection or death by neglect, but may have a partial role in negative selection during early thymic T-cell development. PMID- 12121438 TI - Genetic aspects and microenvironment affect expression of CD18 and VLA-4 in experimental tuberculosis. AB - Control of infection by Mycobacterium tuberculosis is dependent on macrophage activation and efficient migration of effector T-cell populations. Lymphocyte differentiation is associated with changes in cell surface phenotype and alterations in the migratory pattern of these cells. In this study, we investigated the expression of adhesion receptors involved in activation and migration process in experimental tuberculosis. We observed that susceptible BALB/c mice infected with virulent M. tuberculosis by intraperitoneal route presented downmodulation of very late antigen 4 (VLA-4) and unchanged levels of CD18 and CD44hi on peritoneal lymphocytes. On the other hand, lymphocytes from resistant C57BL/6 mice infected by the same route showed unchanged levels of VLA 4 and upregulation of CD18 and CD44hi. However, when BALB/c mice were infected by intratracheal route, lung lymphocytes presented a different pattern of CD18, CD44hi and VLA-4 expression from that observed on peritoneal cells, characterized by unchanged levels of VLA-4 and upregulation of CD18 and CD44hi- coincidentally the same phenotype found on peritoneal cells from C57BL/6. These results suggest that susceptibility and resistance to M. tuberculosis infection, depending on the experimental model, are related to the expression of CD18, CD44hi and VLA-4. Moreover, the microenvironment at the site of infection seems to differentially regulate the expression of these receptors. Thus, the up- or downmodulation of these adhesion receptors is probably associated with differential recruitment of T cells at the site of infection, which may or may not mediate protection in experimental tuberculosis. PMID- 12121439 TI - Expression of human protein tyrosine phosphatase epsilon in leucocytes: a potential ERK pathway-regulating phosphatase. AB - The expression of protein tyrosine phosphatase epsilon (PTPepsilon) was studied in human tissues and blood cells. High mRNA expression was observed in peripheral blood leucocytes, particularly in monocytes and granulocytes which revealed at least four distinct transcripts. In lymphocytes, PTPepsilon expression was induced after 12-O-tetradecanoylphorbol-13-acetate (TPA) or antigen-receptor stimulation, indicating that PTPepsilon plays a role in the events taking place after antigen engagement. Previously, PTPepsilon has been shown to be involved in regulating voltage-gated potassium channel activity, insulin-receptor signalling and Janus kinase-signal transducers and activators of transcription (STAT) signalling. Transfection of cells with different PTPepsilon constructs and activator protein-1 reporter gene indicates that the catalytic activity of PTPepsilon is involved in the regulation of the mitogen-activated protein kinase cascade. In particular, the extracellular signal-regulated kinases (ERK1/2) were shown to be inhibited in both phosphorylation status and enzymatic activity after overexpression of PTPepsilon. Thus, PTPepsilon emerges as a phosphatase with a potential to regulate the ERK1/2 pathway either directly or indirectly through its catalytic activity. PMID- 12121440 TI - The cellular response associated with cervical intraepithelial neoplasia in HIV+ and HIV- subjects. AB - This study investigates local alterations in T-cell and macrophage subsets that occur in cervical epithelial neoplasia (CIN), in the presence and absence of human immunodeficiency virus (HIV) infection. Ectocervical biopsies from 10 women with CIN who were infected with HIV, and 10 women with CIN but no HIV infection were studied by immunocytochemistry. Significantly increased proportions of activated CD8+ T cells were seen in all CIN biopsies, and these proportions were further increased in the presence of HIV infection. Levels of CD8+TIA-1+ cells were particularly increased in the CIN+HIV+ group. There was a lack of expression of CD28 on the CD8+ cells of the epithelium of CIN+HIV+ samples. A significant reduction in the proportion of epithelial inductive D1+ macrophages and an increase in D1+D7+-suppressive cells were observed in the CIN+HIV+ cohort. The lack of expression of CD28 on the CD8+ cells of the epithelium of CIN+HIV+ samples in combination with the reduced CD4+ T-cell numbers seen in the presence of HIV infection may contribute to the development of higher grade CIN in this susceptible group. This may be aggravated by the reduction in the D1+ epithelial inductive macrophages, which might reflect recruitment of more suppressive D1+D7+ cells. This would further compromise the ability of the local T-cell system to respond to antigens and thus contribute to the development of neoplasia at this site. These results suggest that the increase in activated CD8+ T cells is a consequence rather than a cause of CIN. PMID- 12121441 TI - Restriction in T-cell receptor repertoire in a patient affected by trichothiodystrophy and CD4+ lymphopenia. AB - Molecular analysis of T-cell receptor (TCR) repertoire, by measuring the CDR3 heterogeneity length of beta-variable regions (spectratyping), is useful for acquiring novel information on the status of immune system in primary immunodeficiency. Here, we evaluate TCR repertoire in a child with trichothiodystrophy (TTD) and combined immunodeficiency (CID). Spectratyping revealed marked alterations of TCR repertoire distribution: 21 and 10 out of 27 TCR Vbeta (TCRBV) families and subfamilies were skewed in CD8+ and CD4+ subsets, respectively. These findings revealed, for the first time in a TTD patient with CID, a marked reduction in the TCR repertoire complexity, which may reflect alterations in the mechanisms regulating the generation and homeostasis of T cells. PMID- 12121443 TI - The broad-spectrum potato cyst nematode resistance gene (Hero) from tomato is the only member of a large gene family of NBS-LRR genes with an unusual amino acid repeat in the LRR region. AB - The Hero gene of tomato is a broad spectrum resistance gene that confers a high level of resistance to all pathotypes of the potato cyst nematodes Globodera rostochiensis and partial resistance to G. pallida. The gene was identified by map-based cloning, sequencing and complementation analysis of two susceptible tomato lines with an array of 13 overlapping cosmids spanning a total distance of 135 kb. Hero encodes a protein with a nucleotide-binding site (NBS) and a leucine rich-repeat (LRR) domain and is a member of a gene family of 14 highly homologous genes, which are clustered within a continuous 118-kb region. The isolated Hero gene displayed resistance to various G. rostochiensis pathotypes and partial resistance to G. pallida pathotype Pa2/3 in transgenic tomato lines. None of the Hero homologues conferred resistance to G. rostochiensis pathotypes. Hero can be distinguished from its homologues by the length of a compound hexanucleotide microsatellite, which codes for a charged and repetitive amino acid domain within the LRR. We propose that the expansion of this microsatellite may be involved in the evolution of the Hero resistance gene. PMID- 12121444 TI - The plasma membrane oxidase NtrbohD is responsible for AOS production in elicited tobacco cells. AB - A cDNA encoding a protein, NtrbohD, located on the plasma membrane and homologue to the flavocytochrome of the neutrophil NADPH oxidase, was cloned in tobacco. The corresponding mRNA was accumulated when tobacco leaves and cells were treated with the fungal elicitor cryptogein. After elicitation with cryptogein, tobacco cells transformed with antisense constructs of NtrbohD showed the same extracellular alkalinization as the control, but no longer produced active oxygen species (AOS). This work represents the first demonstration of the function of a homologue of gp91-phox in AOS production in elicited tobacco cells. PMID- 12121445 TI - Searching limiting steps in the expression of chloroplast-encoded proteins: relations between gene copy number, transcription, transcript abundance and translation rate in the chloroplast of Chlamydomonas reinhardtii. AB - We performed a systematic investigation of the quantitative relationship between genome copy number, transcription, transcript abundance and synthesis of photosynthetic proteins in the chloroplast of the green algae Chlamydomonas reinhardtii grown either in mixotrophic or phototrophic conditions. The chloroplast gene copy number is lower in the latter condition and the half-life and accumulation levels of most chloroplast transcripts are significantly reduced, although the relative rates of protein synthesis remain similar. Our study shows that, in most instances, chloroplast protein synthesis is poorly sensitive to changes in gene copy number or transcript abundance in the chloroplast. Treatment with 5-fluoro-2'-deoxyuridine, that inhibits chloroplast DNA replication and decreases extensively the number of copies of the chloroplast genome, had limited effects on the abundance of most chloroplast transcripts and little if any effect on the rates of protein synthesis. When using rifampicin, that selectively inhibits chloroplast transcription, we found no direct correlation between the level of transcripts remaining in the chloroplast and the rates of chloroplast protein synthesis. For two chloroplast genes, a 90% decrease in the amount of transcript did not cause a drop in the rate of synthesis of the corresponding protein product. Overall, our results demonstrate that there is no gene dosage effect in the chloroplast and that transcript abundance is not limiting in the expression of chloroplast-encoded protein. PMID- 12121446 TI - Conifer reproductive development involves B-type MADS-box genes with distinct and different activities in male organ primordia. AB - The Norway spruce MADS-box genes DAL11, DAL12 and DAL13 are phylogenetically related to the angiosperm B-function MADS-box genes: genes that act together with A-function genes in specifying petal identity and with C-function genes in specifying stamen identity to floral organs. In this report we present evidence to suggest that the B-gene function in the specification of identity of the pollen-bearing organs has been conserved between conifers and angiosperms. Expression of DAL11 or DAL12 in transgenic Arabidopsis causes phenotypic changes which partly resemble those caused by ectopic expression of the endogenous B genes. In similar experiments, flowers of Arabidopsis plants expressing DAL13 showed a different homeotic change in that they formed ectopic anthers in whorls one, two or four. We also demonstrate the capacity of the spruce gene products to form homodimers, and that DAL11 and DAL13 may form heterodimers with each other and with the Arabidopsis B-protein AP3, but not with PI, the second B-gene product in Arabidopsis. In situ hybridization experiments show that the conifer B like genes are expressed specifically in developing pollen cones, but differ in both temporal and spatial distribution patterns. These results suggest that the B function in conifers is dual and is separated into a meristem identity and an organ identity function, the latter function possibly being independent of an interaction with the C-function. Thus, even though an ancestral B-function may have acted in combination with C to specify micro- and megasporangia, the B function has evolved differently in conifers and angiosperms. PMID- 12121447 TI - Comparative analysis of plastid transcription profiles of entire plastid chromosomes from tobacco attributed to wild-type and PEP-deficient transcription machineries. AB - Transcription of plastid chromosomes in vascular plants is accomplished by at least two RNA polymerases of different phylogenetic origin: the ancestral (endosymbiotic) cyanobacterial-type RNA polymerase (PEP), of which the core is encoded in the organelle chromosome, and an additional phage-type RNA polymerase (NEP) of nuclear origin. Disruption of PEP genes in tobacco leads to off-white phenotypes. A macroarray-based approach of transcription rates and of transcript patterns of the entire plastid chromosome from leaves of wild-type as well as from transplastomic tobacco lacking PEP shows that the plastid chromosome is completely transcribed in both wild-type and PEP-deficient plastids, though into polymerase-specific profiles. Different probe types, run-on transcripts, 5' or 3' labelled RNAs, as well as cDNAs, have been used to evaluate the array approach. The findings combined with Northern and Western analyses of a selected number of loci demonstrate further that frequently no correlation exists between transcription rates, transcript levels, transcript patterns, and amounts of corresponding polypeptides. Run-on transcription as well as stationary RNA concentrations may increase, decrease or remain similar between the two experimental materials, independent of the nature of the encoded gene product or of the multisubunit assembly (thylakoid membrane or ribosome). Our findings show (i) that the absence of photosynthesis-related, plastome-encoded polypeptides in PEP-deficient plants is not directly caused by a lack of transcription by PEP, and demonstrate (ii) that the functional integration of PEP and NEP into the genetic system of the plant cell during evolution is substantially more complex than presently supposed. PMID- 12121448 TI - Evidence for the presence and activity of a complete antioxidant defence system in mature sieve tubes. AB - The phloem is the major route for the transport of solutes and nutrients from source to sink organs in plants. The functional transport phloem consists of parenchymal tissue, enucleate sieve elements, and the intimately connected companion cells. The general absence of a nucleus and functional ribosomes in sieve tubes poses problems especially for damage avoidance and repair of sieve element components. To examine how sieve tubes can remain functional during oxidative stress, we analysed phloem sap of cucumber and pumpkin plants with respect to the presence of antioxidant defence enzymes, their enzymatic activity, and activity changes after exposure to drought stress. Using 1D SDS-PAGE and nano ESI MS/MS, the presence of proteins such as cytosolic Cu/Zn superoxide dismutase, monodehydroascorbate reductase, and peroxidase could be shown. Moreover, activities for several antioxidant enzymes (superoxide dismutase, dehydroascorbate reductase, peroxidase) in phloem exudate could be demonstrated. The activity of these enzymes in phloem sap from cucumber and pumpkin plants increased in response to drought stress. The presented results together with earlier findings provide evidence supporting the presence of a complete machinery of antioxidant defence enzymes and detoxifying metabolites important for avoiding damage to essential components of the sieve elements due to oxidative stress. PMID- 12121450 TI - A novel method for the construction of genome wide transcriptome maps. AB - Expression profiling by cDNA-AFLP is commonly used to display the transcriptome of a specific tissue, treatment or developmental stage. In this paper, cDNA-AFLP has been used to study transcripts expressed in segregating populations from Arabidopsis thaliana and potato (Solanum tuberosum). The genetic differences between the offspring genotypes are thus visualized as polymorphisms in the transcriptome. We show that polymorphic transcripts can be used as genetic markers and allow the construction of a linkage map. The resulting map shows that, in contrast to genomic markers, the transcriptome-derived markers did not cluster in particular areas of the chromosome, and that cDNA-AFLP markers are targeted specifically to transcriptionally active regions. The cDNA-AFLP markers used in mapping are derived from DNA polymorphisms in transcripts, rather than differences in expression regulation. The high potential of transcriptome markers as opposed to (anonymous) genomic markers for applications in genetic analyses, marker-assisted breeding and map-based cloning is discussed. PMID- 12121449 TI - A nucleus-encoded maize protein with sigma factor activity accumulates in mitochondria and chloroplasts. AB - Plants contain nuclear gene families that encode proteins related to the principal sigma factors of eubacteria. As sigma factors function in transcription, the plant proteins have been presumed or demonstrated to associate with the eubacteria-like RNA polymerase of chloroplasts. In maize, five sig cDNA sequences have been reported, and four of the products are present in plastids as predicted. However, in vitro chloroplast import assays and computer algorithms gave ambiguous results with the fifth protein, ZmSig2B. Unlike the other maize sigma factors, ZmSig2B is expressed throughout developing seedling leaves, as well as in roots and etiolated tissues. To determine the subcellular location of ZmSig2B, we have now used immunoblot assays to show that it co-purifies with both mitochondria and plastids. Its NH2-terminal 153 amino acids, translationally fused to green fluorescent protein (GFP), targeted GFP to chloroplasts and mitochondria in bombarded maize leaves. A putative role for ZmSig2B in mitochondrial transcription is supported by its presence in a maize mitochondrial transcription extract. ZmSig2B also exhibits the expected properties of a chloroplast sigma factor: recombinant ZmSig2B binds to a chloroplast promoter and initiates transcription in vitro when combined with Escherichia coli core RNA polymerase. Therefore ZmSig2B is an unusual nucleus-encoded sigma factor that appears to function in both chloroplasts and mitochondria. PMID- 12121451 TI - Constructing plant radiation hybrid panels. AB - Radiation hybrid (RH) mapping, a somatic cell genetic technique, has been developed in animal systems as a general approach for the construction of long range physical maps of chromosomes. This statistical method relies on X-ray induced breakage of chromosomes to determine the physical distance between markers, as well as their order on the chromosome. The method can be applied to single chromosomes or across the whole genome. The generation of plant (barley) radiation hybrids and their culture in vitro is described here. PCR-based marker systems are used to verify hybrid status and to demonstrate genome coverage. RH panels of the type generated can be used for physical mapping, map-based cloning, or sequence contig assembly. RH resources will greatly aid the physical characterisation of crop plants with large genomes. PMID- 12121452 TI - Analysis of surface growth in shoot apices. AB - A salient feature of shoot meristem growth is the maintenance of distinct anatomical and morphological features despite a continuous flux of cells. To investigate how meristem organization is self-perpetuated, we developed a protocol for the analysis of meristem growth in 3-D. Our protocol uses a non destructive replica method to follow the pattern of cell expansion and cell divisions on the meristem surface over several days. Algorithms to reconstruct the meristem surface and compute its curvature and rate of extension were implemented. We applied this approach to the shoot apical meristem of Anagallis arvensis and showed that a subcellular resolution of extension rates can be achieved. This is the first detailed quantitative analysis of meristem geometry and surface expansion in 3-D. This new approach will be useful to connect cellular activities such as cell expansion, cell division, and differential gene expression with overall meristem morphogenesis. PMID- 12121453 TI - Ecophysiology of Antarctic vascular plants. AB - Most of the ice and snow-free land in the Antarctic summer is found along the Antarctic Peninsula and adjacent islands and coastal areas of the continent. This is the area where most of the Antarctic vegetation is found. Mean air temperature tends to be above zero during the summer in parts of the Maritime Antarctic. The most commonly found photosynthetic organisms in the Maritime Antarctic and continental edge are lichens (around 380 species) and bryophytes (130 species). Only two vascular plants, Deschampsia antarctica Desv. and Colobanthus quitensis (Kunth) Bartl., have been able to colonize some of the coastal areas. This low species diversity, compared with the Arctic, may be due to permanent low temperature and isolation from continental sources of propagules. The existence of these plants in such a permanent harsh environment makes them of particular interest for the study of adaptations to cold environments and mechanisms of cold resistance in plants. Among these adaptations are high freezing resistance, high resistance to light stress and high photosynthetic capacity at low temperature. In this paper, the ecophysiology of the two vascular plants is reviewed, including habitat characteristics, photosynthetic properties, cold resistance, and biochemical adaptations to cold. PMID- 12121454 TI - CO2-concentrating mechanisms in Egeria densa, a submersed aquatic plant. AB - Egeria densa is an aquatic higher plant which has developed different mechanisms to deal with photosynthesis under conditions of low CO2 availability. On the one hand it shows leaf pH-polarity, which has been proposed to be used for bicarbonate utilization. In this way, at high light intensities and low dissolved carbon concentration, this species generates a low pH at the adaxial leaf surface. This acidification shifts the equilibrium HCO3-/CO2 towards CO2, which enters the cell by passive diffusion. By this means, E. densa increases the concentration of CO2 available for photosynthesis inside the cells, when this gas is limiting. On the other hand, under stress conditions resulting from high temperature and high light intensities, it shows a biochemical adaptation with the induction of a C4-like mechanism but without Kranz anatomy. Transfer from low to high temperature and light conditions induces increased levels of phosphoenolpyruvate carboxylase (PEPC, EC 4.1.1.31) and NADP-malic enzyme (NADP ME, EC 1.1.1.40), both key enzymes participating in the Hatch-Slack cycle in plants with C4 metabolism. Moreover, one PEPC isoform, whose synthesis is induced by high temperature and light, is phosphorylated in the light, and changes in kinetic and regulatory properties are correlated with changes in the phosphorylation state of this enzyme. In the present review, we describe these two processes in this submersed angiosperm that appear to help it perform photosynthesis under conditions of extreme temperatures and high light intensities. PMID- 12121455 TI - A protein kinase activated by darkness phosphorylates nitrate reductase in Komatsuna (Brassica campestris) leaves. AB - Although it has been shown that leaf nitrate reductase (NR: EC 1.6.6.1) is phosphorylated by subjecting plants to darkness, there is no evidence for the existence of dark-activated or dark-induced NR kinase. This study was undertaken to investigate the occurrence of a protein kinase phosphorylating NR in response to dark treatments. Immediately after transferring Komatsuna (Brassica campestris L.) plants to darkness, we observed rapid increases in the phosphorylating activity of the synthetic peptide, which is designed for the amino acid sequence surrounding the regulatory serine residue of the hinge 1 region of Komatsuna NR, in crude extracts from leaves. The activity reached a maximum after 10 min of darkness. Inactivation states of NR estimated from relative activities with or without Mg2+ were correlated to activities of the putative dark-activated protein kinase. Using the synthetic peptide as a substrate, we purified a protein kinase from dark-treated leaves by means of successive chromatographies on Q-Sepharose, Blue Sepharose, FPLC Q-Sepharose, and ATP-gamma-Sepharose columns. The purified kinase had an apparent molecular mass of 150 kDa with a catalytic subunit of 55 kDa, and it was Ca2+-independent. The purified kinase phosphorylated a recombinant cytochrome c reductase protein, a partial protein of NR, and holo NR, and inactivated NR in the presence of both 14-3-3 protein and Mg2+. The kinase also phosphorylated synthetic peptide substrates designed for sucrose phosphate synthase and 3-hydroxy-3-methylglutaryl-Coenzyme A reductase. Among inhibitors tested, only K252a, a potent and specific serine/threonine kinase inhibitor, completely inhibited the activity of the dark-activated kinase. The activity of the purified kinase was also specifically inhibited by K252a. Taken together with these findings, results obtained suggest that the putative dark-activated protein kinase may be the purified kinase itself, and may be responsible for in vivo phosphorylation of NR and its inactivation during darkness. PMID- 12121456 TI - Cloning of a vacuolar invertase from Belgian endive leaves (Cichorium intybus). AB - Although a lot of vacuolar invertase (EC 3.2.1.26) cDNAs are available from a diversity of plant species, up to now no sequence information is available on invertases from any dicot fructan-containing species. Therefore, we describe the cloning of vacuolar acid invertase cDNA from etiolated Belgian endive leaves (Cichorium intybus L. var. foliosum cv. Flash), formed throughout the forcing process of the witloof chicory roots. Full-length cDNA was obtained by a combination of RT-PCR, PCR and 5'- and 3' RACE RT-PCR, starting with primers based on conserved amino acid sequences. The cloned chicory acid invertase groups together with vacuolar type invertases and fructan biosynthetic enzymes. A putative role for vacuolar type invertases in fructan synthesizing plants is discussed. PMID- 12121457 TI - Overexpression of a bacterial indole-3-acetyl-l-aspartic acid hydrolase in Arabidopsis thaliana. AB - Transgenic Arabidopsis lines (ecotype Col-0) carrying the Enterobacter agglomerans IaaspH gene under CaMV 35S promoter control were more sensitive to exogenous indole-3-acetyl aspartic acid (IAA-Asp) and metabolized [2'-14C]IAA-Asp more rapidly than control lines. Free IAA, total IAA and IAN levels in independent transgenic lines that accumulated IaaspH mRNA varied insignificantly from control levels, yet IAA-Asp levels were significantly reduced. The transgenic lines were grown in a variety of conditions and subjected to morphometric analysis. All three lines showed statistically significant differences in rosette diameter (in soil), root and hypocotyl length (on agar). These effects were transient in some cases and did not manifest themselves under all growth conditions tried. The two independent lines with single T-DNA insertions had lower seed set compared to control lines. PMID- 12121458 TI - Induction of tolerance to fast desiccation in black spruce (Picea mariana) somatic embryos: relationship between partial water loss, sugars, and dehydrins. AB - Events associated with the induction of tolerance to fast desiccation in black spruce (Picea mariana) somatic embryos were investigated. An experimental approach using an initial period of partial water loss was developed to induce either no, partial, or complete tolerance to fast desiccation. Tolerance to subsequent fast desiccation was not promoted by decreasing embryo water content from 1.5 to 1.1 g H2O g-1 DW (g g-1) throughout the first 24 h of slow desiccation. However, tolerance increased from 10 to 95% germination during the second 24-h period of slow desiccation after partial water loss from 1 to 0.55 g g-1. Emphasis was also placed on the relationship between observed tolerance, and sugar and dehydrin contents. Compared to controls, sucrose content in embryos doubled after 24 h of slow desiccation and more than tripled after 48 h. Conversely, starch content was decreased by one half after 24 h and by three quarters after 48 h. Sucrose abundance and raffinose occurrence after 48 h of slow desiccation were congruent with complete tolerance to fast desiccation. The period of slow desiccation between 24 and 48 h also increased the content of a 24 kDa dehydrin and the appearance of a 42-kDa dehydrin. The relationship between partial water loss, sugars and dehydrins is discussed with respect to tolerance to fast desiccation in black spruce somatic embryos. PMID- 12121459 TI - Effects of water stress on antioxidant enzymes of leaves and nodules of transgenic alfalfa overexpressing superoxide dismutases. AB - The antioxidant composition and relative water stress tolerance of nodulated alfalfa plants (Medicago sativa L. x Sinorhizobium meliloti 102F78) of the elite genotype N4 and three derived transgenic lines have been studied in detail. These transgenic lines overproduced, respectively, Mn-containing superoxide dismutase (SOD) in the mitochondria of leaves and nodules, MnSOD in the chloroplasts, and FeSOD in the chloroplasts. In general for all lines, water stress caused moderate decreases in MnSOD and FeSOD activities in both leaves and nodules, but had distinct tissue-dependent effects on the activities of the peroxide-scavenging enzymes. During water stress, with a few exceptions, ascorbate peroxidase and catalase activities increased moderately in leaves but decreased in nodules. At mild water stress, transgenic lines showed, on average, 20% higher photosynthetic activity than the parental line, which suggests a superior tolerance of transgenic plants under these conditions. However, the untransformed and the transgenic plants performed similarly during moderate and severe water stress and recovery with respect to important markers of metabolic activity and of oxidative stress in leaves and nodules. We conclude that the base genotype used for transformation and the background SOD isozymic composition may have a profound effect on the relative tolerance of the transgenic lines to abiotic stress. PMID- 12121460 TI - Oxygen-mediated cold-acclimation in cucumber (Cucumis sativus) seedlings. AB - Cold acclimation of etiolated cucumber seedlings, consisting of cooling at 12 degrees C for 48 h followed by a warming period at 25 degrees C, led to tolerance to subsequent chilling at 2 degrees C. Tolerance, as evidenced by freedom from chilling injury and continued growth, developed during the warming period in a time-course manner for 12 h but decreased with prolonged warming. A similar increase and subsequent decrease was also observed in the content of palmitic, linoleic and linolenic acids in total lipid fraction from cucumber hypocotyl tissue. During the warming period supra-ambient oxygen stimulated, whereas subambient oxygen inhibited, the increase in fatty acid content as well as development of chilling tolerance. A strong correlation between oxygen-mediated changes in fatty acid content and associated development of cold tolerance suggests that both these processes are interrelated. Cold acclimation, but not cold stress, led to an increase followed by a decrease in CO2 evolution suggesting that a respiratory upsurge is yet another feature of cold acclimation in cucumbers. PMID- 12121461 TI - A gene encoding stellacyanin is induced in Capsicum annuum by pathogens, methyl jasmonate, abscisic acid, wounding, drought and salt stress. AB - A stellacyanin cDNA clone (CASLP1) was isolated from a pepper cDNA library from hypersensitive response (HR) lesions of leaves infected with an avirulent strain of Xanthomonas campestris pv. vesicatoria. The deduced amino acid sequences of CASLP1 are homologous to those of stellacyanins from cucumber, maize, pea and Arabidopsis. The CASLP1 mRNA was not constitutively expressed in all organs of pepper, but strongly induced and accumulated in pepper tissues infected with X. campestris pv. vesicatoria, Colletotrichum coccodes, Phytophthora capsici or C. gloeosporioides. In situ hybridization results revealed that CASLP1 transcripts were strongly localized in the phloem areas of vascular bundles in infected tissues of pepper stems and fruits. CASLP1 mRNA accumulation was found in lower pepper leaves infected by either virulent or avirulent strains of X. campestris pv. vesicatoria and non-pathogenic Pseudomonas fluorescens, but not in uninoculated upper leaves. Induction of this CASLP1 gene occurred in pepper leaves applied with methyl jasmonate (MeJA), but not with ethylene, salicylic acid, dl-beta-amino-n-butyric acid and benzothiadiazole. Accumulation of CASLP1 transcripts was locally or systemically induced in pepper leaves upon mechanical wounding and was activated in a MeJA-dependent manner. The CASLP1 transcript was also strongly induced in leaf and stem tissues after exposure of pepper plants to abscisic acid, salt and drought. PMID- 12121462 TI - Boron requirement in the Discaria trinervis (Rhamnaceae) and Frankia symbiotic relationship. Its essentiality for Frankia BCU110501 growth and nitrogen fixation. AB - The essentiality of boron (B) for nitrogen fixation in heterocystous cyanobacteria and rhizobial symbioses has been widely established. However, nothing is known about the possible involvement of the micronutrient in actinorhizal symbioses. Therefore, the effect of boron (B) deficiency on the establishment of the Discaria trinervis-Frankia BCU110501 symbiosis was investigated. Nodulation was diminished in B-deficient D. trinervis or in plants inoculated with Frankia grown in the absence of B. These poorly nodulated plants showed a reduction of shoot and root weight and small size. Because depletion of the micronutrient during growth of the actinomycete altered the infection capacity of Frankia, we also studied growth, structure and nitrogen fixation of free-living Frankia BCU110501. Growth was delayed in B-deficient BAP media (+N cultures), and completely inhibited in B-deprived N-free BAP media (-N cultures), suggesting that B is required to enhance growth of Frankia and essential for the development of nitrogen fixing activity. Ultrastructural study of B-deficient cells showed an alteration of filament walls both in +N and especially in -N cultures, indicating a possible role of the microelement in the maintenance of these structures. Moreover, the stability of vesicle envelopes was impaired in the absence of B and, hence, nitrogenase occurrence and nitrogen fixation were totally absent. The results show that B is required for both partners to establish an effective symbiosis. PMID- 12121463 TI - Chilling tolerance of maize, cucumber and rice seedling leaves and roots are differentially affected by salicylic acid. AB - Salicylic acid (SA) is one component of a complex signalling pathway that is induced by a number of biotic and abiotic stresses. Exposing seedling radicles to aqueous solutions of 0.5 mM salicylic acid for 24 h before chilling at 2.5 degrees C for 1-4 days reduced the chilling-induced increase in electrolyte leakage from maize and rice leaves, and cucumber hypocotyls, but not from their radicles. The SA treatments that induced chilling tolerance in the aerial portion of the seedlings did not induce chilling tolerance in the radicles, even though the SA treatments were applied to the radicles. A comparison of activity among five antioxidant enzymes showed that SA did not alter enzyme activities in the radicles, but that chilling tolerance induced by SA in the aerial portions of maize and cucumber plants was associated with an increase in the activity of glutathione reductase and guaiacol peroxidase. PMID- 12121464 TI - Effect of regurgitant from Leptinotarsa decemlineata on wound responses in Solanum tuberosum and Phaseolus vulgaris. AB - The effect of regurgitant from Leptinotarsa decemlineata Say larvae on wound induced responses was studied using two plant species, Solanum tuberosum L. and Phaseolus vulgaris L. Wounding of one leaf of intact S. tuberosum plants differentially affected ethylene production and activities of peroxidase and polyphenol oxidase. Only polyphenol oxidase activity was stimulated by wounding in both wounded and systemic leaves. Peroxidase activity was not affected by wounding. Wounding caused only a transient increase of ethylene production from wounded leaves. The application of regurgitant to wound surfaces stimulated ethylene production as well as activities of peroxidase and polyphenol oxidase in both wounded and systemic leaves. Wounding significantly enhanced ethylene production and polyphenol oxidase activity in wounded and systemic leaves of P. vulgaris. The application of regurgitant caused an amplification of ethylene production, peroxidase activity, and polyphenol oxidase activity, in both wounded and systemic leaves of bean plants. Several substances were tested for their role as possible endogenous signals in P. vulgaris. Hydrogen peroxide and methyl jasmonate appeared as potential local and systemic signals of ethylene formation in wounded bean plants. Local ethylene production in leaf discs was differentially affected by the regurgitant application in potato versus bean plants. While all tested concentrations of regurgitant caused stimulation of ethylene formation from potato leaf discs, ethylene production was completely inhibited by increasing concentrations of the regurgitant in bean leaf discs. Our data present evidence that ethylene may play an important role in the interaction between plants and herbivores at the level of recognition of a particular herbivore leading to specific induction of signalling cascades. PMID- 12121465 TI - Two subtilisin-like proteases from soybean. AB - Two subtilisin-like proteases (SLP) were identified in soybean (Glycine max [L.] Merr.). The first, SLP-1, was localized in seed coats early in seed development, but became undetectable with anti-SLP-1 antibodies as seed fill progressed. A partial purification of SLP-1 was achieved using a two step chromatographic procedure. NH2-terminal sequence analysis of the partially purified enzyme permitted primers to be designed that were used to amplify cDNA encoding SLP-1. A genomic clone encoding SLP-1 was also obtained. Characterization of the cDNA and partially purified SLP-1 revealed the initial translation product was an 82 694 MW precursor. After removal of a signal peptide, the mature protein was formed by removal of an NH2-terminal propeptide. A COOH-terminal peptide also appeared to be removed from some of the protease molecules. DNA blot analysis suggested that at least one additional SLP gene was present in soybean. The second gene, SLP-2, was subsequently cloned and characterized. Although the coding regions for SLP-1 and SLP-2 were homologous, their promoters were quite divergent. RT-PCR revealed that SLP-2 message was found in the mature plant and in cotyledons of germinating seeds. Although SLP-2 mRNA could be identified in developing seeds, the message was at least an order of magnitude less abundant than that for SLP-1, and it was mis-spliced such that a chain termination event would preclude obtaining a product. As with SLPs from other organisms, the functions of the soybean proteases are unknown. However, SLP-1 is one of only a few proteins from soybean seed coats that have been described. PMID- 12121466 TI - Identification of a grapefruit cDNA belonging to a unique class of citrus dehydrins and characterization of its expression patterns under temperature stress conditions. AB - Citrus fruits are sensitive to low temperatures and this often results in the development of chilling injuries during postharvest storage. In order to gain more insight into the molecular mechanisms involved in the acquisition of fruit chilling tolerance, we initiated a grapefruit (Citrus paradisi, cv. Marsh Seedless) flavedo cDNA sequencing project and used it to identify a cDNA similar to other Poncirus trifoliata and Citrus unshiu dehydrin genes reported to be responsive to low temperatures. The grapefruit dehydrin cDNA, designated cor15, encodes a predicted polypeptide of 15.1 kDa, that is almost completely identical with other reported citrus dehydrin proteins, except that it contains two large amino acid repeats, whereas P. trifoliata COR11 has only one such repeat and P. trifoliata COR19 and C. unshiu COR19 have three repeats. Together, the various grapefruit, P. trifoliata and C. unshiu dehydrins form a closely related and unique dehydrin gene family that differs from most other plant dehydrins in having an unusual K-segment similar to that of gymnosperms and in having a serine cluster (S-segment) at an unusual position at the carboxy-terminus. The grapefruit cor15 gene is consistently expressed in the fruit peel tissue at harvest, but its message levels dramatically decrease during storage at 2 degrees C. However, a pre-storage hot water treatment, which enhances fruit chilling tolerance, elicited retention of the constant level of cor15 gene expression during cold storage and eliminated its decline. The hot water treatment had no inductive effect on cor15 gene expression when the fruit were held at non chilling temperatures. The effects of other stresses, such as exposure to ethylene, UV irradiation and wounding, on cor15 gene expression, were temporary and persisted for 1-2 days after the treatments. PMID- 12121467 TI - Characterization of mutants with reduced seed dormancy at two novel rdo loci and a further characterization of rdo1 and rdo2 in Arabidopsis. AB - Seed dormancy and germination are complex traits that are controlled by many genes. Four mutants in Arabidopsis thaliana exhibiting a reduced dormancy phenotype, designated rdo1, rdo2, rdo3, and rdo4, have been characterized, both genetically and physiologically. Two of these mutants, rdo1 and rdo2, have been described before, the other two represent novel loci. The mutants mapped on chromosome 1 (rdo3), chromosome 2 (rdo2 and rdo4), and chromosome 3 (rdo1). None of these loci has been related to dormancy before. All four mutants show pleiotropic effects in the adult plant stage, which are different for each mutant. None of the mutants is deficient in ABA. Compared to Ler (wild-type), ABA sensitivity is not altered either, thereby excluding the possibility that ABA is involved in causing the reduced dormancy phenotype. The GA requirement was studied by using the GA biosynthesis inhibitor paclobutrazol, and genetically by generating double mutants with the GA-deficient mutant ga1-3. The results obtained by these two methods were comparable for all but one mutant: rdo1. In a GA-deficient background, rdo1, rdo2 and rdo3, all show sensitivity to GA between that of ga1-3 and ga1-3 aba1. However, when using paclobutrazol rdo1 exhibited the same sensitivity as rdo4 and wild-type. Analysis of double mutants among the rdo mutants revealed a very complex and inconsistent pattern. PMID- 12121468 TI - Isolation and characterization of a Chlamydomonas gene that encodes a putative blue-light photoreceptor of the phototropin family. AB - In the search for a Chlamydomonas reinhardtii photoreceptor that may mediate blue light-induced responses we identified a gene that encodes a protein with a structure typical for that of members of the phototropin family, i.e. two LOV domains that may function in flavin mononucleotide binding and a ser/thr kinase domain. The amino acid sequences of these domains are closely related to those of higher plant phototropins. This single-copy gene (Phot) encodes a protein with a calculated molecular mass of 81.4 kDa which is distinctly smaller than the homologous proteins of higher plants that exhibit molecular masses around 120 kDa. Expression analyses revealed rather constant levels of Phot mRNA and Phot protein in vegetative cells incubated in the dark and in cells undergoing gametogenesis. Only vegetative cells in the light showed a reduced expression of the Phot gene. Cell fractionation studies revealed that the protein is membrane associated. In higher plants, phototropins were shown to be bound to the plasma membrane. However, the expression of a Phot-GFP gene fusion in tobacco protoplasts revealed an association of the fusion protein with the endogenous membrane network of the cell. PMID- 12121469 TI - Interaction of Porphyromonas gingivalis with KB cells: comparison of different clinical isolates. AB - The ability of different Porphyromonas gingivalis strains (15 clinical isolates and ATCC 33277) to attach to and invade KB cells, in relation to other properties such as release of interleukin (IL)-6 and IL-8, cytotoxicity, proteolytic activity and types of fimbriae genes present, was examined. A hierarchical cluster analysis based on adherence and internalization resulted in four groups. Eight of the 15 clinical isolates belonged to a cluster group whose adherence and internalization were about 10% those of the ATCC strain. A negative correlation between lysine-specific protease activity and adherence was found. In all cases the released concentrations of IL-6 and IL-8 were very low. Only one strain was found to be cytotoxic to KB cells. Principal components analysis demonstrated correlations between adherence, internalization and autoaggregation. Most strains had fimA type I and II, type I being associated with elastase-like activity. The ability of P. gingivalis to invade epithelial cells may be a key factor for maintaining periodontal disease. PMID- 12121470 TI - Identification of the peptide motifs that interact with HLA-DR8 (DRB1*0802) in Streptococcus mutans proteins. AB - A glucosyltransferase (GTF) and a surface protein antigen (PAc) of Streptococcus mutans have been suggested as possible components of an effective dental caries vaccine. To identify antigenic peptides in GTF and PAc that bind to MHC class II (HLA-DR8, DRB1*0802) molecules, we investigated binding activities to DR8 molecules of overlapping synthetic peptides at several sites in GTF and in the alanine-rich repeating region of PAc using an ELISA-inhibition competitive binding assay for the interaction between the HLA-DR molecule and the PAc (316 334) peptide. Six GTF peptides and 10 PAc peptides strongly bound to the HLA-DR8 molecule. In a homology analysis of the amino acid sequences of the six GTF peptides, two binding motifs were found in L/Y--Y/L-A/N and Y/L--N/G/E--Y-V/L/P. Moreover, a new binding motif in PAc was found in L--Y-A. It is suggested that these binding motifs could be useful in designing a dental caries vaccine in humans. PMID- 12121471 TI - Expression and roles of herpesvirus entry mediators A and C in cells of oral origin. AB - The roles of viral glycoprotein D (gD) and cellular herpesvirus entry mediators A (HveA) and C (HveC) in herpes simplex virus entry into oral cells were determined. Studies with purified truncated forms of gD-1, HveA and HveC indicated that these molecules may be involved in herpes simplex virus entry into oral cells. Moreover, HveA was expressed similarly in primary cultures of gingival keratinocytes and fibroblasts, whereas HveC was expressed at higher levels in gingival keratinocytes, as determined by RT-PCR and immunocytochemical staining. Further analysis using immunohistochemistry demonstrated that both HveA and HveC were expressed in epithelial cells, fibroblasts and vascular endothelial cells in gingival tissues. However, only HveC was detected in nerve fibers. Also, HveA was detected throughout the epidermis, whereas HveC was pronounced in the strata basale and spinosum. In conclusion, this study characterized HveA and HveC, molecules that may participate in entry of herpes simplex virus into oral cells. PMID- 12121472 TI - Molecular mediators of Porphyromonas gingivalis-induced T-cell apoptosis. AB - Porphyromonas gingivalis produces virulence factors which can modify the molecular and cellular components of the host immune response. In the present work we investigated the role of specific virulence factors from P. gingivalis in the induction of apoptosis in Jurkat T cells. P. gingivalis culture supernatants mimicked the effect of butyric acid on T-cell apoptosis and this effect was associated with an increase in histone H4 acetylation. A role for proteases was excluded in experiments which demonstrated that neither protease inhibitors nor use of P. gingivalis mutants defective in protease synthesis had any effect on the stimulation of T-cell apoptosis in this system. PMID- 12121474 TI - Intracellular and extracellular pHs of Streptococcus mutans after addition of acids: loading and efflux of a fluorescent pH indicator in streptococcal cells. AB - A pH-sensitive fluorescent dye, 2', 7'-bis-(2-carboxyethyl)-5 and 6 carboxyfluorescein (BCECF), was used to determine intracellular pH (pH(in)). The efflux of BCECF loaded into oral streptococcal cells was determined after incubation of the cells at 35 degrees C for 20 min in the presence and absence of glucose. In the absence of glucose, the fluorescence of intracellular BCECF in Streptococcus mutans, Streptococcus sanguis, Streptococcus salivarius and Streptococcus sobrinus decreased only very slightly, indicating that the dye could be useful for pH(in) determination. In the presence of glucose, however, the fluorescence decreased by 57%. Thus, the pH(in) of S. mutans cells was measured by the BCECF method in the absence of glucose at various acidic pH levels by adding lactic, acetic and hydrochloric acids to the cell suspensions. The pH(in) was almost equal to the extracellular pH (pH(out)) for pH(out) values of between 8 and 5, indicating that protons permeated easily across the S. mutans cell membrane. For pH(out) between 5 and 4, pH(in) was constant at around 5, suggesting that the cell membrane was impermeable to protons, or that a cytoplasmic buffering system functioned. pH(in) decreased at pH(out) values of < 4. The constant pH(in) at acidic pH(out) levels could protect intracellular components, such as proteins, against acidification by sugar fermentation. PMID- 12121473 TI - Detection of cytolethal distending toxin activity and cdt genes in Actinobacillus actinomycetemcomitans isolates from geographically diverse populations. AB - A cytolethal distending toxin (CDT) found in Actinobacillus actinomycetemcomitans inhibits the eukaryotic cell cycle, which may contribute to the pathogenic potential of the bacterium. The presence of the cdtABC genes and CDT activity were examined in 40 clinical isolates of A. actinomycetemcomitans from Brazil, Kenya, Japan and Sweden. Thirty-nine of 40 cell lysates caused distension of Chinese hamster ovary cells. At least one of the cdt genes was detected in all strains examined. The three cdt genes were detected, by PCR, in 34 DNA samples. DNA from one strain from Kenya did not yield amplicons of the cdtA and cdtB genes and did not express toxic activity. Restriction analysis was performed on every amplicon obtained. PCR-RFLP patterns revealed that the three cdt genes were conserved. These data provided evidence that the cdt genes are found and expressed in the majority of the A. actinomycetemcomitans isolates. Although a quantitative difference in cytotoxicity was observed, indicating variation in expression of CDT among strains, no clear relationship between CDT activity and periodontal status was found. PMID- 12121475 TI - Kinetics of KB and HEp-2 cell responses to an invasive, cytolethal distending toxin-producing strain of Actinobacillus actinomycetemcomitans. AB - The periodontal pathogen Actinobacillus actinomycetemcomitans produces cytolethal distending toxin (CDT), a complex multicomponent toxin that arrests the growth of many types of eukaryotic cell. The kinetics of the effects of CDT-containing extracts, from an invasive strain of this bacterium, were examined on epithelial like cells routinely used in invasion studies. Both KB and HEp-2 cells were exquisitely sensitive to the effects of the CDT with TD50 of 30 and 300 pg of total bacterial protein, respectively. Initial cell morphology changes were relatively rapid, occurring within the first 13 h of exposure. CDT-treated KB cells increased in size to 4-5 times the size of untreated controls. Cytotoxicity was irreversible when attached cells were incubated, for a minimum of 120 min, with nanogram quantities of CDT-containing extract. As cultures aged, the cells became more resistant to the effects of the CDT-containing extracts. These findings have important implications for understanding the ability of A. actinomycetemcomitans to invade and multiply in epithelial cells. PMID- 12121476 TI - Streptococcus mutans binding to solid phase dextran mediated by the glucan binding protein C. AB - Streptococcus mutans GbpC is a wall-anchored surface protein which is involved in dextran-dependent aggregation. The GbpC phenotype is observed only in cells grown under stress conditions. In order to detect the GbpC protein of S. mutans, we isolated the wall fraction following digestion of the cell wall of this organism by N-acetylmuramidase, and detected the GbpC protein from S. mutans cells by western analysis with anti-GbpC serum. Interestingly, S. mutans cells exhibiting the negative dextran(alpha-1,6 glucan)-dependent aggregation (ddag) phenotype expressed the protein and could bind to immobilized dextran. PMID- 12121477 TI - Adhesion of viridans group streptococci to sialic acid-, galactose- and N acetylgalactosamine-containing receptors. AB - The binding of 10 viridans group streptococci to sialic acid-, galactose (Gal)- and N-acetylgalactosamine (GalNAc)-containing receptors was defined by analysis of the interactions between these bacteria and structurally defined glycoconjugates, host cells and other streptococci. All interactions with sialic acid-containing receptors were Ca(2+)-independent as they were not affected by ethyleneglycoltetraacetic acid (EGTA), whereas all interactions with Gal- and GalNAc-containing receptors were Ca(2+)-dependent. Recognition of sialic acid-, Gal- and GalNAc-containing receptors varied widely among the strains examined, in a manner consistent with the association of each of the three lectin-like activities with a different bacterial cell surface component. PMID- 12121478 TI - gbpC and pac gene mutations detected in Streptococcus mutans strain GS-5. AB - The Streptococcus mutans gbpC gene encoding cell wall-anchoring glucan-binding protein C is involved in the dextran(alpha-1,6 glucan)-dependent aggregation (ddag) of this organism. Unlike cells of other strains of S. mutans, strain GS-5 cells did not exhibit dextran(alpha-1,6 glucan)-dependent aggregation under any conditions. We therefore hypothesized that the gbpC gene may be mutated in strain GS-5. Sequencing analysis of the 1752-nucleotide GS-5 gbpC gene revealed a point mutation that switched codon 65 to a TAA termination codon. Strain GS-5 was previously reported also to have a mutation in the pac gene encoding the cell wall-anchored major protein antigen. The laboratory-maintained strain GS-5 is regarded as having lower cariogenicity than the original isolate. The decreased cariogenicity developed during the laboratory culture of strain GS-5 may have been caused by mutations in an environment lacking appropriate selective pressures. PMID- 12121479 TI - Melatonin protects against gastric ulceration and increases the efficacy of ranitidine and omeprazole in reducing gastric damage. AB - The antiulcer effect of melatonin on gastric lesions caused by restraint-cold stress was studied with the intent of determining the mechanism of action of this agent. Melatonin dose-dependently prevented restraint-cold stress-induced gastric damage with around 90% inhibition at a dose of 60 mg/kg BW. When compared with already marketed antiulcer drugs such as ranitidine and omeprazole, melatonin was found to be more effective than ranitidine but less effective than omeprazole in preventing stress ulcer. As stress-induced gastric lesions are mainly caused by oxidative damage because of hydroxyl radicals (*OH), the effect of melatonin in scavenging the.OH generated during stress conditions in vivo as well as in an in vitro model system were studied. The results indicate that melatonin caused an 88% reduction of endogenous *OH during stress in vivo, an observation confirmed in an established in vitro system. Furthermore, a decrease in the activity of gastric peroxidase (GPO) and an increase in the gastric mitochondrial superoxide dismutase (Mn-SOD) activity because of restraint-cold stress was attenuated by melatonin pretreatment indicating that the indole possibly exerts its gastroprotective effects through its direct as well as indirect antioxidant activities. Moreover, in separate experiments, cotreatment of rats with melatonin and ranitidine or omeprazole was found to protect against stress ulceration in doses at which either of these alone could not protect the stomach. The findings raise the possibility of melatonin being considered as an effective gastroprotective agent individually or as a cotreatment with either ranitidine and omeprazole. PMID- 12121480 TI - Potentiation of antiproliferative effects of tamoxifen and ethanol on mouse hepatoma cells by melatonin: possible involvement of mitogen-activated protein kinase and induction of apoptosis. AB - Melatonin, the major secretory product of the pineal gland, is in focus of many research areas because of its ability to scavenge free oxygen radicals and thereby protect cells and tissues from radical damage. Some studies suggest melatonin may be a possible therapeutic agent with potential clinical applications against pathological states due to reactive oxygen species. Here, we investigated the effects of melatonin on the mouse hepatoma cell line HEPA 1-6, coincubated with ethanol, and tamoxifen, respectively. Cell proliferation rates were detected by the 3-[4,5 dimethylthiazol-2-y1]-2,5-diphenyltetrazolium bromide (MTT) proliferation assay. A dose-dependent inhibition of the proliferative activity by melatonin was observed from 640 microM to 3 mM, which was significantly higher (P < 0.01) than with the solvent (ethanol) alone. Concentrations of 320 microM and less had no effect on cell proliferation. This antiproliferative effect might be because of the prolonged activation of mitogen activated protein kinase which was activated by phosphorylation 15 min after the induction with melatonin. Furthermore, apoptosis was found to be enhanced by melatonin (75% more than with the solvent alone, P < 0.001). Finally, we show that the inhibitory effect of tamoxifen (25 microM) is markedly enhanced by the coincubation with melatonin (1.3 mM) up to 75% (P < 0.001). These data show that the antiproliferative effects of tamoxifen and ethanol, respectively, on mouse hepatoma cell line HEPA 1-6 are enhanced by melatonin. Although at the conditions described here the antiproliferative effects of melatonin occur at supraphysiological concentrations, these data may help to support clinical studies where melatonin is given simultaneously with tamoxifen or other standard chemotherapeutica. PMID- 12121481 TI - Melatonin potentiates 5-HT(1A) receptor activation in rat hypothalamus and results in hypothermia. AB - Effects of melatonin on both thermoregulatory responses and hypothalamic serotonin release were assessed in unanesthetized rats at three different ambient temperatures (Ta). Systemic administration of melatonin (30-120 mg/kg, i.p) caused a decrease in both colonic temperature and hypothalamic serotonin (5-HT) release in rats at both Ta 8 and 22 degrees C. The hypothermia was brought about by a decrease in metabolic rate at Ta 8 degrees C, whereas at Ta 22 degrees C the hypothermia was produced by both a decrease in metabolic rate and an increase in cutaneous temperature. However, in the heat (Ta 31 degrees C), neither thermoregulatory responses nor hypothalamic 5-HT release was affected by the same amount of administered melatonin. The melatonin-induced hypothermia and decreased 5-HT release in the hypothalamus were attenuated by selective depletion of brain 5-HT produced by intracerebroventricular injection of 5,7-dihydroxytryptamine. Furthermore, the melatonin-induced hypothermia was almost completely abolished by treatment with a 5-HT2A receptor agonist (DOI) or a 5-HT1A receptor antagonist [( )-pindolol]. The data indicate that melatonin potentiates the 5-HT1A receptor activation in the hypothalamus and results in hypothermic effects which can be antagonized by the expected hyperthermic effect of DOI. PMID- 12121482 TI - Melatonin's unique radical scavenging properties - roles of its functional substituents as revealed by a comparison with its structural analogs. AB - Melatonin's O-methyl and N-acetyl residues are not only the basis of its amphilicity enabling the molecule to enter all organs and all subcellular compartments, but are also decisive for its antioxidant properties. We have compared melatonin's redox chemistry with that of several structural analogs: tryptamine, N-acetyltryptamine, serotonin, N-acetylserotonin, 5 methoxytryptamine, 6-chloromelatonin and 2-iodomelatonin. Scavenging of hydroxyl radicals (*OH) was measured in a scavenger competition assay based on ABTS cation radical (ABTS(*)+) formation. The capability of undergoing single-electron transfer reactions was studied using an ABTS(*)+ reduction assay, reflecting the more general property of scavenging organic cation radicals. Direct scavenging of superoxide anions (O2(*)-), under non-catalyzed conditions, was investigated in a hematoxylin autoxidation assay. Measurements of chemiluminescence were used for studying scavenging of O2(*)- under catalyzed conditions, either by hemin mediated interaction or by combination with the respective indolyl cation radicals. Light emission was determined in the absence or presence of the *OH scavenger dimethylsulfoxide and the O2(*)- scavenger Tiron. Products formed by oxidation of the respective indoles in a moderately alkaline, hemin-catalyzed H2O2 system were analyzed by thin-layer chromatography and fluorometry. Absence of either the O-methyl or the N-acetyl residue causes marked diminutions in the capacities of scavenging *OH and ABTS(*)+ as well as in chemiluminescence emitted during oxidation. The importance of the N-acetyl group is insofar remarkable as it seems, at first glance, to be isolated from the indolic moiety; interactions between side chain and indolic moiety are therefore decisive for melatonin's redox properties. The 5-hydroxylated compounds are not generally more efficient scavengers, but particularly better reducers of ABTS(*)+; in the alkaline H2O2 system generating *OH and O2(*)-, melatonin was much more rapidly oxidized than the 5-hydroxylated and non-substituted analogs. Oxidative products formed from any of the compounds studied contained much less of substituted kynuramines as in the case of melatonin, indicating that radical chain termination by O2(*)- is considerably more efficient with melatonin. These findings are supported by measurements of chemiluminescence, which largely reflects pyrrole ring cleavage as a result of combination with superoxide anions. In this regard, only 6 chloromelatonin equalled melatonin, whereas the efficiency of 2-iodomelatonin was much lower, another indication for the importance of 2,3-dioxygenation. PMID- 12121483 TI - Physiological concentrations of melatonin inhibit the nitridergic pathway in the Syrian hamster retina. AB - In the present work, the effect of melatonin on the hamster retinal nitridergic pathway was examined. When the retinas were incubated in the presence of low concentrations (1 pM-10 nM) of melatonin for 15 min, a significant decrease of nitric oxide synthase (NOS) activity was observed. However, when crude retinal homogenates were preincubated with melatonin for 15 min, no changes in NOS activity were detected, despite the fact that under the same conditions trifluoperazine, a calmodulin inhibitor, significantly decreased enzymatic activity. Kinetic analysis showed that melatonin decreased the V(max) of retinal NOS without changes in the K(m). On the other hand, low concentrations (100 pM) of melatonin significantly reduced retinal L-arginine influx. A decrease in the V(max) of L-arginine uptake was observed in the presence of melatonin, whereas the K(m) remained unchanged. Melatonin significantly inhibited the accumulation of cyclic guanosine monophosphate (cGMP) levels induced by both L-arginine and sodium nitroprusside (SNP). In summary, the present results indicate that melatonin could be a potent inhibitor of the retinal nitridergic pathway. PMID- 12121484 TI - The effects of melatonin on Ca(2+) homeostasis in endothelial cells. AB - The effect of melatonin on the Ca(2+) signaling process in bovine aortic endothelial cells (BAE) and in primary cultured vascular endothelial cells from normotensive Sprague Dawley (SDR) and genetically hypertensive (SHR) rats was investigated using the Ca(2+) indicator Fura-2. Acute applications of melatonin failed to initiate a Ca(2+) response in the three cell types considered. However, preincubating SHR aortic endothelial cells with exposure to melatonin increased the internal Ca(2+) release triggered by bradykinin (BK) and ATP while stimulating the related agonist-evoked Ca(2+) entry. This effect appeared specific for SHR cells, as a similar incubation period failed to alter the Ca(2+) responses in BAE and SDR cells. Because of the known overproduction of free radicals in SHR cells, the effect of melatonin on Ca(2+) signaling was also tested in SDR and BAE cells exposed to the superoxide anion radical. Melatonin reversed the deleterious action of free radicals on Ca(2+) signaling in both cases, suggesting that its stimulatory effect in SHR was linked to its antioxidative properties. Finally, experiments where melatonin was applied between successive BK stimulation periods showed an enhancement of the agonist evoked Ca(2+) entry in BAE and SDR cells. This effect appeared to be independent of the production of second messengers as no specific binding sites for melatonin, including MT1, MT2 and MT3 receptors, could be detected in BAE cells. We conclude that melatonin improves Ca(2+) signaling in dysfunctional endothelial cells characterized by an overproduction of free radicals while stimulating the agonist-evoked Ca(2+) entry in normal endothelial cells through a mechanism not related to its antioxidative properties. PMID- 12121485 TI - Neuroprotection by melatonin against ischemic neuronal injury associated with modulation of DNA damage and repair in the rat following a transient cerebral ischemia. AB - In the present study, double fluorescence staining combined with confocal laser scanning microscopy analysis were used to examine the effects of melatonin on ischemia-induced neuronal DNA strand breaks and its possible mechanisms in a transient middle cerebral artery (MCA) occlusion model. Results showed that melatonin dose-dependently reduced infarct areas and decreased both DNA double and single strand breaks (DSB and SSB) and enhanced cell viability in the peri ischemic brain regions. Furthermore, Bcl-2 induction in the ischemic brain was further enhanced by melatonin treatment. Double staining analysis indicated that the cells costained for Bcl-2 and TdT-mediated-deoxyuridine triphosphate (dUTP) nick-end labeling (TUNEL), a DSB marker, displayed a relative regular morphology compared with the cells only stained with TUNEL. Transient ischemia induced an expression of excision repair cross-complementing factor 6 (ERCC6) mRNA, a gene essential for the preferential repair of nuclear excision repair, in the injured neurons. Double labeling showed that ERCC6 only co-localized with proliferating cell nuclear antigen (PCNA), a member of the nuclear excision repair complex, but not with TUNEL. Melatonin further and statistical significantly up-regulated ERCC6 mRNA expression in the peri-ischemic region of rat brains. The results suggest that neuroprotection by melatonin against ischemic injury may be related to modulation of apoptosis and DNA repair capacity. PMID- 12121486 TI - Acute modifications in the levels of daytime melatonin do not influence leptin in postmenopausal women. AB - Melatonin shows a clear circadian rhythm with peak values at night, and may act directly with fat cells. Leptin, the anorexic hormone synthesized mainly by adipocytes, is produced in a circadian fashion, similar to that of melatonin. Accordingly, in the present study, we investigated whether melatonin may contribute to the rise in circulating leptin. The study was performed in postmenopausal women with 2 months of treatment with placebo or estradiol (50 microg/day). Melatonin was administered in doses of 1 mg by mouth versus placebo. In experiment 1, melatonin was administered at 08:30 hr. In experiment 2, at 08:30 hr and 10:30 hr, and in experiment 3 at 15:30 hr. Three blood samples, one every 15 min, were collected prior to the administration of melatonin and 2 hr after the administration of the single melatonin dose or the second melatonin administration (experiment 2). Following its administration, circulating melatonin reached pharmacological levels. In the three experiments, levels of leptin were not modified by the daytime administration of melatonin. These data indicate that, at least in daytime hours, acute modifications in daytime melatonin levels do not influence levels of leptin of postmenopausal women either without or with estradiol replacement. Accordingly, the metabolic, endocrine, reproductive and biological modifications induced by acute daytime melatonin in women do not seem to be mediated by modifications in circulating leptin. PMID- 12121487 TI - Role of CSF in the transport of melatonin. PMID- 12121488 TI - Variation in reproductive behaviour within a sex: neural systems and endocrine activation. AB - Intrasexual variation in reproductive behaviour, morphology and physiology is taxonomically widespread in vertebrates, and is as biologically and ecologically significant as the differences between the sexes. In this review, we examine the diverse patterns of intrasexual variation in reproductive behaviours within vertebrates. By illustrating the genetic, cellular, hormonal and/or neural mechanisms underlying behavioural variation in a number of species, another level of complexity is added to studies of brain organization and function. Such information increases our understanding of the unique and conserved mechanisms underlying sex and individual differences in behaviour in vertebrates as a whole. Here, we show that intrasexual variation in behaviour may be discrete or continuous in nature. Moreover, this variation may be due to polymorphism at a single genetic locus or many loci, or may even be the result of phenotypic plasticity. Phenotypic plasticity simply refers to cases where a single genotype (or individual) can produce (or display) different phenotypes. Defined in this way, plasticity subsumes many different types of behavioural variation. For example, some behavioural phenotypes are established by environmental factors during early ontogeny, others are the result of developmental transitions from one phenotype early in life to another later in life, and still other strategies are facultative with different behaviours displayed in different social contexts. PMID- 12121489 TI - Changes in basal hypothalamic chicken gonadotropin-releasing hormone-I and vasoactive intestinal polypeptide associated with a photo-induced cycle in gonadal maturation and prolactin secretion in intact and thyroidectomized starlings (Sturnus vulgaris). AB - Chicken gonadotropin-releasing hormone-I (GnRH-I) and the avian prolactin releasing hormone, vasoactive intestinal polypeptide (VIP), were measured in the basal hypothalamus in male starlings during photo-induced gonadal growth and the subsequent development and maintenance of reproductive photorefractoriness. Comparisons were made with thyroidectomized birds, which maintain breeding condition irrespective of changes in photoperiod. In intact birds, basal hypothalamic GnRH-I increased four-fold after photostimulation and then decreased 115-fold over 12 weeks to values characteristic of long-term photorefractoriness. Pituitary and plasma prolactin increased after photostimulation, reaching peak values when the testes were regressing, and returned to low values in long-term photorefractory birds. Basal hypothalamic VIP did not change after photostimulation in intact birds. In photostimulated thyroidectomized birds, values for basal hypothalamic GnRH-I and VIP, and for pituitary and plasma prolactin, remained no different to those of nonphotostimulated intact birds. These observations confirm that reproductive photorefractoriness is related to a decrease in hypothalamic GnRH-I. However, photorefractoriness in terms of prolactin secretion is not similarly related to a decrease in basal hypothalamic VIP. The mechanisms responsible for the decrease in prolactin in long-term photorefractory birds and for the total lack of photoperiodic responses in thyroidectomized birds remain unresolved. PMID- 12121490 TI - The effect of endotoxin administration on the secretory dynamics of oxytocin in follicular phase mares: relationship to stress axis hormones. AB - The primary aim of this study was to define the secretory dynamics of oxytocin and vasopressin in pituitary venous effluent from ambulatory horses during acute endotoxaemia, a stimulus that may release both hormones. Our secondary aim was to investigate the role of oxytocin in regulating adrenocorticotropic hormone (ACTH) secretion by comparing oxytocin, vasopressin, corticotropin-releasing hormone (CRH) and ACTH secretory profiles during endotoxaemia and by monitoring the ACTH response to oxytocin administration. Pituitary venous blood was collected nonsurgically continuously and divided into 1-min segments from eight follicular phase mares. Four mares were sampled for 30 min before and 3.5 h after receiving an i.v. infusion of bacterial endotoxin (TOX). Four control mares were sampled for 2.5 h without infusion of TOX. Another three follicular phase mares were given 5 U of oxytocin to replicate the peak response to TOX and pituitary blood collected every 1 min for 10 min before and 15 min after injection. Endotoxin raised the secretion rates of all hormones measured. All hormones were released episodically throughout the experiment, with TOX increasing the amplitude of peaks in each hormone. Peaks in oxytocin and vasopressin were coincident in each treated mare. Similarly, ACTH peaks were coincident with peaks of oxytocin and vasopressin in each treated mare, and with peaks of CRH in three mares. However, oxytocin administration did not affect ACTH secretion. We conclude that during endotoxaemia in horses: (i) oxytocin and vasopressin are secreted synchronously; (ii) oxytocin is unlikely to be acting as an ACTH secretagogue since inducing peak oxytocin concentrations observed during TOX does not raise ACTH; and therefore (iii) the close relationship between oxytocin and ACTH secretion is circumstantial and due to the fact that oxytocin secretion is concurrent with that of vasopressin, a proven ACTH secretagogue in horses. PMID- 12121491 TI - Hormonal and behavioural responses of paradoxical sleep-deprived rats to the elevated plus maze. AB - Activation of the hypothalamic-pituitary-adrenal (HPA) axis is observed immediately after 96 h of paradoxical sleep (PS) deprivation. However, when individually or group PS-deprived rats are challenged with a mild stressor, they exhibit a facilitation of the corticosterone response, and a faster return to basal levels than control rats. Because the housing condition influences coping behaviour, we tested whether the type of PS deprivation (individually or in group) influenced anxiety-like behaviour in the elevated plus-maze and the accompanying adrenocorticotropin (ACTH) and corticosterone responses. Individually (I-DEP) or group deprived (G-DEP) rats and their appropriate control groups were either killed immediately after 96 h of sleep deprivation (time-point 0 or 'basal') or exposed to a 5-min test on the elevated plus maze and sampled 5, 20 or 60 min after test onset. Control of I-DEP rats showed reduced locomotor activity and augmented anxiety-like behaviour, replicating the effects of social isolation. Although I-DEP rats exhibited higher motor activity than cage control rats, these groups did not differ in regard to the percentage of entry and time spent in the open arms. G-DEP rats, in turn, ambulated more, entered and remained longer in the open arms, exhibiting less anxiety-like behaviour. PS-deprived rats exhibited higher ACTH and corticosterone 'basal' secretion than control rats. For all groups, peak ACTH secretion was reached at the 5-min time-point, returning to unstressed basal levels 60 min after the test, except for G-DEP rats, which showed a return at 20 min. Peak levels of corticosterone occurred at 5 min for PS deprived groups and at 20 min for control groups. G-DEP rats showed a return to 'basal' unstressed levels at 20 min, whereas the I-DEP and control groups did so at 60 min. A negative correlation between exploration in the open arms and hormone concentrations was observed. These data indicate that housing condition influences the subsequent behaviour of PS-deprived rats in the EPM which, in turn, seems to determine the secretion profile of ACTH and corticosterone in response to the test. PMID- 12121492 TI - Ghrelin stimulates insulin secretion from the pancreas of normal and diabetic rats. AB - Ghrelin is a novel 28-amino acid gut-brain peptide, which was first isolated in the rat stomach. This study examined the effect of ghrelin on insulin secretion from the isolated pancreas of normal and diabetic rats. Diabetes was induced by a single dose of streptozotocin. Four weeks after the induction of diabetes, pancreatic tissue fragments of normal and diabetic rats were treated with different concentrations (10(-12), 10(-9) and 10(-6) M) of ghrelin. Ghrelin evoked large and significant increases in insulin secretion from the pancreas of both normal and diabetic rats. In the pancreas of normal rats, diltiazem (calcium channel antagonist) or a combination of atropine (muscarinic cholinergic receptor antagonist), propranolol (beta-adrenergic receptor antagonist) and yohimbine (alpha2-adrenergic receptor antagonist) significantly reduced the stimulatory effect of ghrelin on insulin secretion. Diltiazem and yohimbine failed to inhibit ghrelin-evoked insulin release in diabetic rat pancreas. Ghrelin-immunoreactivity cells was observed in 2.6% and 3.8% of the total cell population in the islet of Langerhans of normal and diabetic rats, respectively. PMID- 12121493 TI - Progestin receptor immunoreactivity within steroid-responsive vasopressin immunoreactive cells in the male and female rat brain. AB - Progestin receptor immunoreactivity is found in the same regions of the bed nucleus of stria terminalis (BST) and centromedial amygdala (CMA) as steroid responsive vasopressin immunoreactive (AVP-ir) cells. To test whether AVP-ir cells express progestin receptors, brains of male rats were stained immunocytochemically for arginine vasopressin as well as progestin receptors. In BST and CMA, over 95% of AVP-ir cells contained progestin receptor immunoreactivity. In contrast, none of the AVP-ir cells in the suprachiasmatic, supraoptic and paraventricular nuclei expressed progestin receptor immunoreactivity. To study whether progestin receptor expression in AVP-ir cells in the BST and CMA is responsive to gonadal steroids, male and female rats were castrated and implanted with either empty capsules or capsules filled with testosterone or oestradiol, respectively. Ten days later, brains were processed for AVP and progestin receptor immunoreactivity. Although there was no effect of hormonal status on the percentage of colocalized cells, the level of progestin receptor immunoreactivity was higher in rats that received gonadal steroids than those that did not. The presence of progestin receptor immunoreactivity in steroid responsive AVP-ir cells, and the responsiveness of this expression to gonadal hormones, is consistent with the possibility that the effects of gonadal steroids on AVP-ir expression in the BST and CMA may be mediated at least in part by progestin receptors. PMID- 12121494 TI - Comparison between the influence of the intravenous and intracerebroventricular injection of a nitric oxide donor on adrenocorticotropic hormone release and hypothalamic neuronal activity. AB - We investigated the ability of the nitric oxide (NO) donor 3-morpholino sydnonimine (SIN-1) to release adrenocorticotropic hormone (ACTH) and up-regulate hypothalamic neurones following its intravenous (i.v.) injection. i.v. SIN-1 (0.2 1.8 mg/kg) produced dose-related increases in plasma ACTH levels which were blocked by prior neutralization of endogenous corticotropin-releasing factor (CRF) but not by vasopressin antibodies. In contrast, the intracerebroventricular (i.c.v.) injection of 50-microg SIN-1 released significantly larger amounts of ACTH, a response blunted by either CRF or vasopressin antibodies. While i.c.v. SIN-1 markedly up-regulated transcripts of the immediate early gene NGFI-B in the paraventricular nucleus (PVN) of the hypothalamus, no such response was observed following the i.v. injection of up to 2.0 mg/kg SIN-1. Finally, we found no evidence that the influence of the peripheral administration of SIN-1 on ACTH secretion is mediated by altered pituitary responsiveness to CRF or vasopressin. The fact that NO has a profound hypotensive influence in the periphery suggests that it may have released ACTH through this mechanism, although the absence of PVN neuronal response in regions that are activated by decreased blood pressure casts some doubt on this hypothesis. As the systemic injection of arginine derivatives that block NOS activity potently augment the ACTH response to circulating pro-inflammatory cytokines or vasopressin, the present findings indicate that the mechanisms responsible for this phenomenon are distinct from those responsible for ACTH released by i.v. SIN-1. PMID- 12121495 TI - Effects of adrenalectomy and of mineralocorticoid receptor/glucocorticoid receptor ligands in female Brown Norway and Fischer 344 rats and f1 hybrids. AB - Our previous studies suggested that the mineralocorticoid receptor (MR) of Brown Norway (BN) male rats is active independently of the presence of its ligands (i.e. constitutively active), and that glucocorticoid receptor (GR)-mediated mechanisms are more efficient in BN than in Fischer 344 (F344) male rats. Such functional differences in corticosteroid receptors led us to compare the effect of adrenalectomy (ADX) and MR/GR-mediated actions (treatments with deoxycorticosterone, DOC and RU 28362, respectively) on female rats from both strains, and, within the framework of a genetic study, to investigate how these differences were inherited in rats of the first generation (F1) born from the crossbreeding between BN and F344 inbred rats. This study extends our previous hypotheses of a constitutive activation of MR and of a greater efficiency of GR in males to females of the BN strain. In both strains, female rats were less sensitive to ADX and to treatments with DOC or RU 28362 than males. Globally, F1 hybrid BNxF344 rats inherited the functional characteristics of MR and GR of BN rats. PMID- 12121496 TI - Ghrelin acts on leptin-responsive neurones in the rat arcuate nucleus. AB - Leptin decreases food intake and increases energy expenditure in rodents by inhibiting neurones in the hypothalamic arcuate nucleus. The growth hormone secretagogue (GHS) ghrelin is known to stimulate food intake and to be the endogenous ligand for the GHS-receptor, which is strongly expressed in the arcuate nucleus, like the leptin receptor (Ob-R). In this study, we analysed the effect of systemic ghrelin administration on Fos expression in the arcuate nucleus on neurones expressing Ob-R. Injection of ghrelin (0.2 mg/kg, i.p) significantly increased the number of neurones expressing Fos protein in the ventromedial arcuate nucleus. Fifty-seven percent of all Fos-positive cells in the ventromedial arcuate nucleus were also positive for Ob-R staining. Furthermore, we investigated electrophysiologically the effect of ghrelin and leptin on the activity of arcuate neurones in an in-vitro slice preparation. Ghrelin stimulated the electrical activity dose-dependently in 80% of all cells tested (n=49) with a threshold concentration of 10(-11) M; only 8% were inhibited and 12% did not respond. The effect of ghrelin (10(-7) M) was weakly antagonized by the peptidic GHS-receptor antagonist (D-Lys3)-GHRP-6 (10(-4) M), which also showed a much weaker affinity (IC(50), 0.9 x 10(-6) M) to the GHS-receptor than ghrelin (IC(50), 0.3 x 10(-9) M). Ghrelin increased the electrical activity in 76% of all cells which were inhibited by leptin (n=17). These data show that ghrelin interacts with the leptin hypothalamic network in the arcuate nucleus. The opposite effect of leptin and ghrelin on neurones in the arcuate nucleus may serve as a neurophysiological correlate of the orexigenic and anorectic effects of ghrelin and leptin. PMID- 12121497 TI - Involvement of cAMP-response element binding protein in corticotropin-releasing factor (CRF)-induced down-regulation of CRF receptor 1 gene expression in rat anterior pituitary cells. AB - Corticotropin-releasing factor (CRF) is a major secretagogue of adrenocorticotopic hormone from the anterior pituitary and a key activator of the hypothalamic-pituitary-adrenal axis. We previously reported that CRF down regulates expression of the CRF type-1 receptor (CRF-R1) mRNA in cultured rat anterior pituitary cells. The present study was conducted to clarify the signal transduction systems involved in CRF-induced down-regulation of CRF-R1 gene expression in the anterior pituitary. Northern blot analysis revealed that, under serum-free conditions, 10 nM CRF decreased CRF-R1 mRNA levels in cultured rat anterior pituitary cells as we reported previously. Treatment with 5 mM 8-Br-cAMP reduced CRF-R1 mRNA levels within 2 h. The mRNA level fell to 37+/-3% of the basal level at 2 h and remained low for 16 h after treatment. This CRF-induced reduction of CRF-R1 mRNA expression was inhibited completely by pretreatment with protein kinase A (PKA) inhibitor (1 microM H-89). Further examination revealed that after pretreatment with 10 microM of antisense oligodeoxynucleotide for cyclic AMP-response element binding protein (CREB), the CRF-induced inhibition of CRF-R1 mRNA was partially decreased to 79+/-4% of the control level 2 h after administration of CRF. These findings indicate that CRF may down-regulate CRF-R1 mRNA expression via a cAMP-PKA-mediated mechanism in rat anterior pituitary cells, and that CREB may mediate at least a portion of this inhibitory effect. PMID- 12121498 TI - Habituation and cross-sensitization of stress-induced hypothalamic-pituitary adrenal activity: effect of lesions in the paraventricular nucleus of the thalamus or bed nuclei of the stria terminalis. AB - Habituation of the hypothalamic-pituitary-adrenal (HPA) response to chronic intermittent restraint stress (30 min/day for 15 days) and the cross sensitization to a heterotypic stress [i.p. lipopolysaccharide (LPS)] were investigated in intact male Sprague Dawley rats, and in rats bearing quinolinic acid lesions to the medial anterior bed nuclei of the stria terminalis (BST) or anterior region of the paraventricular nucleus of the thalamus (PVT). In intact animals, a single period of restraint increased plasma corticosterone levels at 30 min and led to an increase in corticotropin-releasing hormone (CRH) mRNA levels in the PVN at 3 h. LPS had a smaller effect on corticosterone and more variable effect on CRH mRNA. Chronic intermittent restraint stress caused a decrease in body weight and increase in adrenal weights, with concomitant increase in basal corticosterone levels. These animals also displayed marked habituation of the corticosterone and CRH mRNA responses to the homotypic stress of restraint, but no loss of the corticosterone response to the heterotypic stress of LPS and a cross-sensitization of the CRH mRNA response. This pattern of stress responses in control and chronically stressed animals was not significantly affected by lesions to the PVT or BST, two areas which have been implicated in the coping response to stress. Thus, these data provide evidence for independent adaptive mechanisms regulating HPA responses to psychological and immune stressors, but suggest that neither the medial anterior BST nor the anterior PVT participate in the mechanisms of habituation or cross-sensitization. PMID- 12121500 TI - Disease recurrence after liver transplantation: need we worry? PMID- 12121501 TI - Hepatitis B: weighing the costs of treatment. PMID- 12121502 TI - Hepatorenal syndrome in patients with cirrhosis. AB - Type 1 hepatorenal syndrome (HRS) is a severe complication of end-stage cirrhosis. Type 1 HRS is an acute functional renal failure (i.e. glomerular hypofiltration) with no other explanation than the presence of the circulatory and neurohumoral alterations associated with severe chronic liver disease. Plasma volume expansion does not improve renal function. In contrast, administration of the vasopressin analog terlipressin, a splanchnic and systemic vasoconstrictor, may improve renal function and be used while awaiting liver transplantation. PMID- 12121503 TI - Randomized open trial for comparison of proton pump inhibitors in triple therapy for Helicobacter pylori infection in relation to CYP2C19 genotype. AB - BACKGROUND AND AIMS: Genetic polymorphism of cytochrome P450 (CYP) 2C19 influences the efficacy of Helicobacter pylori eradication therapy with a proton pump inhibitor (PPI) and amoxicillin. However, in triple therapy (PPI plus amoxicillin and clarithromycin), little is known about the impact of CYP2C19 polymorphism, or the use of rabeprazole, which is not well metabolized by CYP2C19. The efficacy of three PPI (omeprazole, lansoprazole, and rabeprazole) in a 1-week triple regimen were compared in relation to CYP2C19 polymorphism. METHOD: One hundred and eighty-three patients were randomized to receive one of the following regimens: amoxicillin 500 mg t.i.d., clarithromycin 200 mg t.i.d., and PPI (omeprazole 20 mg, lansoprazole 30 mg, or rabeprazole 10 mg) b.i.d. CYP2C19 polymorphism was analyzed by PCR restriction fragment length polymorphism. RESULTS: Intention-to-treat-based overall cure rates for omeprazole, lansoprazole or rabeprazole regimens were 83.1% (95% confidence interval (CI): 69-89%), 86.7% (CI: 75-93%), and 76.6% (CI: 64-85%), respectively, without significant difference. The cure rate of the rabeprazole regimen (but not the lansoprazole or omeprazole regimens) tended to be correlated with CYP2C19 genotypes (P = 0.076). In patients with a homozygous extensive metabolizer genotype, the per protocol-based cure rate with rabeprazole (62.5%) was significantly lower than that with lansoprazole (90.0%; P = 0.038). CONCLUSION: The overall cure rate of 1-week triple therapy for H. pylori eradication was not significantly different between regimens with omeprazole, lansoprazole or rabeprazole, but the impact of CYP2C19 genetic polymorphism on the cure rate appeared to differ between these PPI. PMID- 12121504 TI - Serum ferritin and Helicobacter pylori infection in children: a sero epidemiologic study in Korea. AB - BACKGROUND: Helicobacter pylori infection is known to affect iron metabolism and serum ferritin levels, which are reduced in adults with H. pylori infection. The aim of the present study was to investigate the association between H. pylori infection and iron status in healthy Korean children. METHODS: The H. pylori seropositivity in 753 schoolchildren aged 6-12 years was screened for using an ELISA and confirmed by western blot analyses. Serum ferritin levels were measured using an immunoradiometric assay in 36 H. pylori-seropositive children and in 72 age- and gender-matched seronegative controls. RESULTS: The median serum ferritin levels were significantly lower in H. pylori-seropositive children than in seronegative controls (24 vs 39 ng/mL; P < 0.001). The prevalence of iron deficiency (ferritin < 15 ng/mL) in H. pylori-seropositive children was significantly higher (13.9%) than in seronegative children (2.8%). This association persisted after adjusting for age and their socioeconomic status (odds ratio, 5.6; 95% confidence interval, 1.0-30.6). CONCLUSION: Serum ferritin levels are reduced in children with H. pylori infection. The H. pylori infection may lead to iron deficiency in children. PMID- 12121505 TI - Increase in colorectal epithelial apoptotic cells in patients with ulcerative colitis ultimately requiring surgery. AB - BACKGROUND AND AIMS: Up to one-third of patients with ulcerative colitis (UC) need to undergo surgery, but the factors that exacerbate inflammation remain unclear. The authors hypothesize that excessive apoptosis reported in active UC may disrupt epithelial defenses and exacerbate the disease. The aim of the present study was to clarify whether apoptotic epithelial cells and histiocytes engulfing them increased in patients with active UC who ultimately require surgery (UC-S) rather than those receiving medication alone (UC-M). METHODS: The study included 29 patients with UC-S, 35 with UC-M, 18 with infectious colitis, and 16 healthy controls. Apoptotic cells were detected by terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL). Using biopsy specimens taken from the most severely inflamed rectosigmoid mucosa as determined endoscopically, the apoptotic index (apoptotic cells/epithelial cells,%) and density (per mm2) of lamina propria histiocytes positive for CD68 were then evaluated. Statistical differences were tested with the Mann-Whitney U test. RESULTS: The apoptotic indices in UC-M patients were significantly higher than those in controls (P < 0.05) but almost equal to those in infectious colitis patients. In the upper and lower halves of the mucosa, both apoptotic indices and histiocyte densities were significantly higher for UC-S than in UC-M (P < 0.01). Ratios of the mean apoptotic index for UC-S to that for UC-M exceeded 3.4, while ratios of the mean histiocyte density were limited to approximately 1.6. CONCLUSIONS: The results suggest that epithelial apoptosis is a non-specific phenomenon and that an increased number of apoptotic cells exceeding histiocyte phagocytic capacity may play a part in the disruption of epithelial defenses and further accelerate mucosal inflammation. PMID- 12121506 TI - Efficacy of granulocyte-macrophage colony-stimulating factor or lamivudine combination with recombinant interferon in non-responders to interferon in hepatitis B virus-related chronic liver disease patients. AB - BACKGROUND AND AIMS: Non-response to interferon (IFN) monotherapy is a major therapeutic problem in the management of chronic hepatitis B infection. The efficacy of combination therapy to enhance the immunomodulatory effect of IFN by combining granulocyte-macrophage colony-stimulating factor (GMCSF) or decreasing viral load by adding an antiviral agent such as lamivudine was evaluated prospectively. METHODS: Twenty-four patients with chronic hepatitis B who were non-responders to previous IFN therapy were randomized to receive an IFN and GMCSF (group A, n = 10) or IFN and lamivudine (group B, n = 14) combination for 6 months. The end-of-treatment response was assessed by hepatitis B virus (HBV)-DNA and hepatitis B e antigen (HBeAg) determination. RESULTS: All patients successfully completed both the treatment schedules. The mean age, alanine aminotransferase (ALT) levels, liver histology, HBV-DNA levels and distribution of HBV genotypes were comparable between the two groups. At the end of treatment there was a significant decrease in mean ALT levels. The HBV-DNA and HBeAg loss was seen in six of 10 (60%) patients in group A and in seven of 14 (50%) patients in group B. During a mean follow-up of 15 +/- 3 months, two of six (33%) patients in group A and three of seven (43%) patients in group B relapsed with HBV-DNA and HBeAg positivity, which meant an overall sustained response of 40% and 28%, respectively. None of the factors such as HBV viral load, ALT levels or liver histology could predict the non-response to combination therapy or occurrence of relapse. There was a trend in patients with genotype A compared with genotype D towards non-response to therapy, although the difference was not significant. CONCLUSIONS: The combination of IFN plus GMCSF or lamivudine was effective in non responders to IFN monotherapy. Larger studies using such combination therapies would be helpful in improving treatment strategies for chronic hepatitis B. PMID- 12121507 TI - Clinicopathologic features of patients with hepatocellular carcinoma seropositive for alpha-fetoprotein-L3 and seronegative for des-gamma-carboxy prothrombin in comparison with those seropositive for des-gamma-carboxy prothrombin alone. AB - BACKGROUND: There has been no study of the clinicopathologic features of patients with hepatocellular carcinoma (HCC) who are seropositive for lectin-reactive alpha-fetoprotein (AFP-L3) alone, or seropositive for AFP-L3 and seronegative for des-gamma-carboxy prothrombin (DCP) in comparison with those who are seropositive for DCP alone. Thus, the present comparative study was performed. METHODS: The clinicopathologic features of HCC patients with either one or two tumors who underwent a hepatectomy (n = 88) were compared among the following five groups according to the seropositivity of AFP, AFP-L3 and DCP: (i) group A, seropositive for AFP above 100 ng/mL, AFP-L3 above 15% and DCP above 100 mAU/mL; (ii) group B, seropositive for AFP-L3 and seronegative for DCP below 40 mAU/mL; (iii) group C, seronegative for AFP below 20 ng/mL, AFP-L3 below 15% and seropositive for DCP; (iv) group D, seropositive for AFP and seronegative for AFP-L3 and DCP; and (v) group E, seronegative for AFP, AFP-L3 and DCP. RESULTS: Group B patients showed a higher incidence of infiltrative-type HCC with an irregular margin (P < 0.05) and a higher frequency of poorly differentiated HCC (P < 0.01) compared with group C patients. Group A patients had larger tumors and more massive-type tumors than group B patients. Our HCC cases showed that advanced clinicopathologic features were demonstrated in the order of group B, group C and group D. Group A and B patients and group D and E patients showed similar characteristics. CONCLUSIONS: Hepatocellular carcinoma patients who were seropositive for AFP-L3 and seronegative for DCP demonstrated clinicopathologic features of more advanced HCC compared with those who were seropositive for DCP alone. PMID- 12121508 TI - Hepatic nodules with early enhancement during computed tomography portography: report of six cases. AB - AIM: To study the clinicopathologic characteristics of hepatic nodular lesions with high attenuation (increased portal blood flow) compared with surrounding hepatic parenchyma on computed tomography (CT) during arterial portography (CTAP). METHODS: For six lesions found in six patients demonstrated as a high attenuated mass by CTAP, CT during hepatic arteriography (CTHA; n = 3 patients), digital subtraction hepatic arteriography (n = 6) and conventional helical CT (n = 6) were evaluated retrospectively and compared with histopathologic findings (n = 4). Pathologic diagnosis was atypical adenomatous hyperplasia, nodule-in-nodule hepatocellular carcinoma (HCC) in one resected lesion each and overt HCC in two biopsied lesions. Two patients did not undergo any therapy and were followed up. RESULTS: The average size of lesions was 2.2 cm (range 1.2-3.5 cm). The CTAP revealed high attenuation in all six lesions; entirely within the lesion (n = 4 lesions) or peripherally with a central low attenuation (n = 2). In contrast, CTHA showed low attenuated lesions; entirely within the mass (n = 2) or peripherally with a central high-attenuated spot (n = 1). Hepatic arteriogram revealed only two hypervascular lesions; entirely and partially in one each. In the arterial phase of helical CT, all but one lesion were iso- or hypo attenuated. In two patients who were followed up to 39 and 55 months without therapy, neither tumor growth nor hemodynamic change of the lesion was recognized on CT. CONCLUSIONS: Even though the incidence of hepatic nodular lesions demonstrated as high attenuating on CTAP is low, all but one lesion in the current series showed iso- or hypo-attenuation on CTHA and/or helical CT, suggesting the hemodynamics are reciprocal between CTAP and CTHA. One exceptional lesion that showed high attenuation on both CTAP and conventional CT was pathologically advanced HCC. Based on the follow-up study of two untreated patients, this kind of lesion with high attenuation on CTAP seems to grow slowly. PMID- 12121509 TI - Aldehydic products of lipid peroxidation do not directly activate rat hepatic stellate cells. AB - BACKGROUND AND AIM: Activation of hepatic stellate cells (HSC) results in the transdifferentiation of the resting (quiescent) phenotype to one characterized by loss of vitamin A droplets, increased alpha-smooth muscle actin (SMA) expression and increased collagen production. Aldehydic products of lipid peroxidation have been shown to increase collagen production by cultured fibroblasts and by passaged HSC, but it is unclear whether these products of lipid peroxidation can initiate the activation of HSC. In the present study the effects were examined of two aldehydic products of lipid peroxidation, malondialdehyde (MDA) and 4 hydroxynonenal (HNE), on activation of rat HSC in early culture as measured by SMA and desmin expression, and collagen production. METHODS: The HSC from normal rat liver were plated in plastic wells and exposed to either MDA (5-200 micromol/L), HNE (0.1-20 micromol/L) or vehicle for either 3 or 7 days. The cells were then harvested; SMA and desmin levels were measured by western blotting. Collagen production was measured by radiolabeled proline incorporation after 6 h of aldehyde exposure. RESULTS: Malondialdehyde (100 and 200 micromol/L) decreased SMA expression during the 3-day and 7-day exposures compared with controls. 4 Hydroxynonenal (20 micromol/L) decreased SMA expression significantly while no effects were observed with lower concentrations compared with controls during the 3-day exposure. Seven-day exposure to HNE (0.1-20 micromol/L) failed to alter SMA expression compared with controls. Exposure to MDA or HNE did not influence desmin expression or collagen production. CONCLUSIONS: Aldehydic products of lipid peroxidation do not directly activate HSC in early culture and alternative pathways may be responsible for HSC activation during oxidative stress. PMID- 12121510 TI - High, but not low, dietary retinoids aggravate manifestation of rat liver fibrosis. AB - BACKGROUND: Although there is strong implication that retinoids regulate Ito cell proliferation and collagen synthesis, results from in vivo studies on the relationship between vitamin A and liver fibrosis are conflicting. The present study focuses on the role of vitamin A in carbon tetrachloride (CCl4)-induced fibrosis by chronic feeding of rats with either a vitamin A-supplemented or depleted diet. METHODS AND RESULTS: In animals with high dietary hepatic retinoid levels, liver fibrosis was more pronounced and was associated with an increased CCl4-toxicity resulting in high mortality (73%). Enhanced liver fibrosis was confirmed by in vivo fluorescence microscopic determination of both collagen deposits (7.4 +/- 1.1 vs 3.9 +/- 0.3% in high vitamin A diet-fed and standard diet-fed fibrotic animals, respectively; P < 0.05) and rarefication of sinusoids (1.5 +/- 0.2 vs 2.4 +/- 0.2 sinusoids/200 microm in high vitamin A diet-fed and standard diet-fed fibrotic animals, respectively; P < 0.05). It was further associated with decreased bile flow and increased parenchymal cell damage. CCl4 reduced hepatic retinoid levels in high vitamin A diet-fed animals, but restored hepatic retinoid levels in animals fed with a vitamin A-deficient diet, implying major interference of vitamin A metabolism with hepatotoxic agents such as CCl4. Low vitamin A feeding did not modulate liver fibrogenesis and caused no mortality. CONCLUSIONS: These results show that the vitamin A status of the liver plays an important role in liver fibrogenesis. While dietary vitamin A shortage does not promote liver fibrogenesis, high levels of vitamin A have the potential to increase systemic and hepatic toxicity of CCl4. Thus, the narrow therapeutic window for nutritional vitamin A substitution must take into account that liver fibrotic patients may display enhanced susceptibility to the adverse effects of vitamin A. PMID- 12121511 TI - Genetic testing for HFE hemochromatosis in Australia: the value of testing relatives of simple heterozygotes. AB - BACKGROUND: It is unclear whether screening of relatives of C282Y and H63D heterozygotes (other than compound heterozygotes) for hemochromatosis will detect sufficient numbers of cases to justify introduction of this screening strategy. METHODS: Conditional probabilities were determined using published Australian allele frequencies and penetrance data to determine the detection rate of hemochromatosis by testing the siblings and offspring of heterozygotes (subjects with only one HFE mutation). RESULTS: The number of individuals who are at risk of developing increased body iron stores because of HFE mutations is substantially higher (1 in 80) than previously estimated. In addition, 33% of the Australian population are heterozygous for either C282Y or H63D. Based on population estimates, the relative risk to the offspring of C282Y and H63D heterozygotes of developing increased iron stores is 4.1 and 1.5, respectively, while the relative risk to each sibling is 2.3 and 1, respectively. The risk of developing clinical features of hemochromatosis or hepatic fibrosis is likely to be substantially lower. CONCLUSIONS: Although the detection rate from testing the families of unaffected heterozygotes is low, this can be justified as a clinically useful screening strategy. At the present time this strategy should be restricted to first-degree relatives of heterozygotes. Further studies are recommended to determine if cascade genetic screening is a cost-effective alternative to general population screening. PMID- 12121513 TI - Hepatobiliary and pancreatic: aspartate aminotransferase elevation in a bed ridden old man. PMID- 12121512 TI - Gastrointestinal: giant inflammatory colonic diverticulum. PMID- 12121514 TI - Ethanol injection therapy of an isolated bile duct associated with a biliary cutaneous fistula. AB - Bile leakage after hepatic resection often results in the formation of a biliary cutaneous fistula. Such a fistula, when caused by an isolated bile duct in the remnant liver, can be intractable. We report a successful case of ethanol injection therapy of an isolated bile duct. A 73-year-old man underwent right hepatic resection for hepatocellular carcinoma. Bile leakage occurred after surgery, and the patient developed a biliary-cutaneous fistula. Fistulography revealed an isolated bile duct in the remnant portion of the caudate lobe without communication to the main biliary system. As conservative management with simple drainage was ineffective, injection therapy with ethanol was performed with a balloon occlusion catheter. After 11 therapy sessions, the bile duct was eradicated, and the biliary- cutaneous fistula was completely healed. The post treatment course was uneventful. Ethanol injection therapy can be a choice for management of patients with a biliary fistula caused by an isolated bile duct. PMID- 12121516 TI - Resistant pathogens: considerations in geriatrics and infectious disease. PMID- 12121517 TI - Antimicrobial resistance and aging: beginning of the end of the antibiotic era? AB - Throughout the history of mankind, infectious diseases have remained a major cause of death and disability. Although industrialized nations, such as the United States, have experienced significant reductions in infection-related mortality and morbidity since the beginning of the "antibiotic era," death and complications from infectious diseases remain a serious problem for older persons. Pneumonia is the major infection-related cause of death in older persons, and urinary tract infection is the most common bacterial infection seen in geriatric patients. Other serious and common infections in older people include intra-abdominal sepsis, bacterial meningitis, infective endocarditis, infected pressure ulcers, septic arthritis, tuberculosis, and herpes zoster. As a consequence, frequent prescribing of antibiotics for older patients is common practice. The large volume of antibiotics prescribed has contributed to the emergence of highly resistant pathogens among geriatric patients, including methicillin-resistant Staphylococcus aureus, penicillin-resistant Streptococcus pneumoniae, vancomycin-resistant enterococci, and multiple-drug-resistant gram negative bacilli. Unless preventive strategies coupled with newer drug development are established soon, eventually clinicians will be encountering infections caused by highly resistant pathogens for which no effective antibiotics will be available. Clinicians could then be experiencing the same frustrations of not being able to treat infections effectively as were seen in the "pre-antibiotic era." PMID- 12121518 TI - Resistant pathogens in urinary tract infections. AB - Antimicrobial susceptibility of bacteria causing urinary tract infection (UTI) has evolved over several decades as antimicrobial exposure has repeatedly been followed by emergence of resistance. Older populations in the community, long term care facilities, or acute care facilities have an increased prevalence of resistant bacteria isolated from UTI. Resistant isolates are more frequent in long-term care populations than the community. Resistant isolates include common uropathogens, such as Escherichia coli or Proteus mirabilis, and organisms with higher levels of intrinsic resistance, such as Pseudomonas aeruginosa or Providencia stuartii. Isolation of resistant organisms is consistently associated with prior antimicrobial exposure and higher functional impairment. The increased likelihood of resistant bacteria makes it essential that a urine specimen for culture and susceptibility testing be obtained before instituting antimicrobial therapy. Therapy for the individual patient must be balanced with the possibility that antimicrobial use will promote further resistance. Antimicrobial therapy should be avoided unless there is a clear clinical indication. In particular, asymptomatic bacteriuria should not be treated with antimicrobials. Where symptoms are mild or equivocal, urine culture results should be obtained before initiating therapy. This permits selection of specific therapy for the infecting organism and avoids empiric, usually broad-spectrum, therapy. Where empirical therapy is necessary, prior infecting organisms should be isolated, and recent antimicrobial therapy, as well as regional or facility susceptibility patterns, should be considered in antimicrobial choice. Where empirical therapy is used, it should be reassessed 48 to 72 hours after initiation, once pretherapy cultures are available. PMID- 12121519 TI - Resistant pathogens in respiratory tract infections in older people. AB - Bronchitis, bronchiectasis, and pneumonia are the most common respiratory tract infections observed in older people and are the leading causes of morbidity and mortality associated with infection. Accurate diagnosis of respiratory tract infections in older people is problematic because of the lack of clear symptoms and signs that are usually seen in younger patients. In addition, the increasing prevalence of bacterial resistance to antibiotic therapy highlights the importance of appropriate therapy. The following review examines the issues associated with the accurate diagnosis of respiratory tract infections, optimal therapy for older patients, and the mechanisms of emerging bacterial resistance to antibiotic therapy. PMID- 12121520 TI - Management of antibiotic-resistant bacteria. AB - Optimizing the management of antibiotic resistance is an important strategy in improving outcomes for infectious diseases in older persons. Strategies that manage antibiotic resistance must take into account all clinical settings, because resistant pathogens previously seen only in acute care facilities are becoming increasingly common in long-term care facilities. Recently, modest improvement in therapeutic options for the treatment of infections due to resistant pathogens has become available because of the development of newer antibiotics. Some of these drugs are briefly discussed in this review, but the best strategy is to limit the potential for the development of resistance and transmission of these pathogens. This can best be accomplished by minimizing misuse of antibiotics and maximizing adherence to basic hygiene standards. PMID- 12121522 TI - Uncovering the secret: giving voice to the experiences of nurses who misuse substances. AB - AIM OF THE STUDY: The aim of this study was to gain insight into the experience of being a nurse with a substance misuse problem. RATIONALE: Several members of the research team work in a withdrawal programme specifically designed for nurses by nurses. Gaining insight into nurses' experiences will contribute to the development of appropriate guidelines to assist with prevention, identification and intervention strategies. BACKGROUND: Published research is almost exclusively from North America. Much of it represents work completed in the 1980s, with very few research-based accounts from the perspective of nurses, highlighting that this is a poorly researched and understood problem in Australia. RESEARCH METHODS: This phenomenological study used in-depth, unstructured interviews either face to face or via the telephone with a purposive sample of 12 nurses who had experienced problem substance use. FINDINGS: Five major themes were identified: nurses' justification for using substances, the fear surrounding being 'discovered', the personal meaning for nurses, the professional impact and the turning point in their road to recovery. DISCUSSION: The five themes derived from the data are inextricably linked to the nature of nursing and of the nursing workplace. Information about potential risk factors and the experiences of nurses with substance misuse problems is critical to the development of prevention and treatment strategies. CONCLUSIONS: Issues of occupational health and safety are raised due to the increasingly demanding and often traumatic nursing work environment. Fears about stigma and loss of their nursing identity highlight nurses' perceptions that treatment programmes are often punitive in nature. Guidelines for the workplace addressing issues such as confidentiality, stress reduction and strategies for handling situations where a colleague is suspected of having a problem are beginning steps that will help address the problem of substance misuse in the nursing profession. PMID- 12121523 TI - How prevalent is violence towards nurses working in general hospitals in the UK? AB - BACKGROUND: Violence in nursing is not a new phenomenon but in recent years much greater emphasis has been placed on the problem in the United Kingdom (UK). A number of official reports, media stories and national initiatives have focused attention on the problem in this country. However, it is not clear whether violence and abuse have in fact become more prevalent. At present little is known about the scale of the problem for general nurses working in general hospitals in the UK. AIM OF THE STUDY: A realistic assessment of the scale of this problem should facilitate a meaningful debate about the interventions needed to counter it and support the requests for funding that will be required. This study aimed to establish the utility of existing research findings, to include relevant but previously unused sources and to synthesize the results. METHOD: A systematic search of the literature pertinent to the aim of the study was followed by a critical review. The focus was on research originating in the UK, including some general research on occupational violence which included data on nurses. FINDINGS: Overall, the research findings are limited. The best available evidence suggests nurses as a whole do face a high level of risk compared with all workers and this excess risk holds for general nurses. The data support a figure of more than 9.5% of general nurses working in general hospitals assaulted (with or without injury) in any 1 year. Trends over time are impossible to identify at present. CONCLUSIONS: Efforts to combat the problem should include greater emphasis on the problem outside accident and emergency departments, prioritizing preregistration training in the management of aggression, and further research. Better reporting should also be a priority. PMID- 12121524 TI - Developing and supporting extended nursing roles: the challenges of NHS walk-in centres. AB - BACKGROUND: National Health Service (NHS) walk-in centres offer immediate access to nurse-led primary care. Forty 'pilot' centres are now open, of which nine are in London. In providing clinical assessment, health information and a limited range of treatments to users, they exemplify new nursing roles that are emerging in primary care. AIM: To describe the emerging roles of walk-in centre nurses, and explore the causes of role stress and review arrangements for training, development and support. METHODS; Semi-structured interviews with 29 managers and nurses from the nine London walk-in centres and with 10 stakeholders providing other primary care services near to three of the walk-in centres. Findings. Walk in centre nurses are drawn from a wide range of clinical backgrounds. Emerging roles include diagnosing, developing clinical management plans, prescribing and discharging patients. Most find the roles challenging, but at times also stressful. There is no consensus on the most appropriate nursing skill mix for a walk-in centre nor on the core competencies required by the nurses working there. As a result there is no standardization of induction, training or support for walk-in centre nurses - particularly on the balance between taught and apprenticeship-type training. CONCLUSIONS: Agreement is required on the competencies required for walk-in centre nursing and on appropriate ways to develop and support nurses in their practice. Lessons could be learned from general practitioner training. PMID- 12121525 TI - A framework for the role of Registered Nurses in the specialty practice of rehabilitation nursing in Australia. AB - AIM OF THE STUDY: This study sought to explore systematically the role of Registered Nurses working in rehabilitation in Australia. BACKGROUND: Rehabilitation has been identified as an important aspect of health care. However, evidence of a comprehensive investigation of the nurses' role in rehabilitation cannot be found. From Australia, in particular, no research has been published in this area. METHODS: This study used a qualitative approach by engaging 13 nurses in one-to-one interviews and a further 21 in focus group discussions. Thematic analysis was conducted on the interview and focus group data. FINDINGS: Seven domains of practice were identified and are suggested as a framework for the specialty practice of rehabilitation nursing. They capture the 'how' and 'what' of rehabilitation nursing practice. Central to this practice is a rehabilitative approach to patient care, teaching and coaching, and continual assessment. The nurses explained in detail how these aspects of rehabilitation nursing differentiate their practice from that of their acute care colleagues. CONCLUSION: The rehabilitative approach is one of a variety of approaches to nursing care, but should not be seen as the exclusive domain of rehabilitation nurses. Rehabilitation belongs in every nurse's toolkit. PMID- 12121526 TI - Applying a phenomenological method of analysis derived from Giorgi to a psychiatric nursing study. AB - BACKGROUND: The experience of mental ill health is fundamentally disempowering. The processes of psychiatric hospital care and treatment may also add to the personal feeling of disempowerment. This disempowerment is partly due to the failure of others to afford a proper hearing to the person's story of his/her experiences and problems in life. Hence, there is a need to investigate patients' experiences of being mentally ill with psychosis and being helped in a psychiatric hospital. AIM: This paper describes the application of a phenomenological method of analysis derived from Amadeo Giorgi to an investigation of psychiatric patients' experiences about being mentally ill with psychosis and being helped in a psychiatric hospital ward in Northern Finland. METHOD: This phenomenological study was conducted with nine voluntary adult patients recovering from psychosis. In 1998, patients were interviewed regarding their experiences of psychosis and being helped. The verbatim transcripts of these interviews were analysed using Giorgi's phenomenological method. Giorgi's method of analysis aims to uncover the meaning of a phenomenon as experienced by a human through the identification of essential themes. Patients' experiences of psychosis and being helped were clustered into a specific description of situated structure and a general description of situated structure. FINDINGS: The Giorgian method of phenomenological analysis was a clear-cut process, which gave a structure to the analyses and justified the decisions made while analysing the data. A phenomenological study of this kind encourages psychiatric nurses to focus on patients' experiences. CONCLUSION: Phenomenological study and Giorgi's method of analysis are applicable while investigating psychiatric patients' experiences and give new knowledge of the experiences of patients and new views of how to meet patients' needs. PMID- 12121527 TI - Unstructured interviews: challenges when participants have a major depressive illness. AB - BACKGROUND: There is debate about undertaking sensitive research with vulnerable populations. Primarily, the literature has focused on informed consent, confidentiality and the principle of beneficence, with little discussion about data collection methods. AIM: This paper discusses the challenges of conducting unstructured interviews when participants have a major depressive illness. Issues arose during a phenomenological study that explored the meaning of being nurtured with seven people who were hospitalized for depression. FINDINGS: The depressive illness and treatment were found to impact on participants' articulation and recalling of their experience, and raised ethical concerns about their informed consent. Personal engagement with participants raised the ethical issue of research vs. therapy. Furthermore, participants being in a hospital complicated the necessity for privacy. The methodological issue of bracketing of assumptions was deemed to be important to 'see' the phenomenological relevance of patients' experiences. CONCLUSION: Knowledge and experience are required when conducting unstructured interviews. Debates about the challenges of unstructured interviewing needs to be highlighted in research texts to assist novice researchers. Support from an experienced research mentor would assist novice interviewers through the interview process and provide post-interview debriefing. PMID- 12121528 TI - Psychometric evaluation of the Demands of Immigration Scale with Taiwanese Chinese immigrants: a pilot study. AB - BACKGROUND: Each year thousands of people move across national borders and become immigrants of another country. They face several demands and sources of distress during resettlement. The Demands of Immigration (DI) Scale developed by Aroian et al. (1998), is the only instrument available to nurses (and other clinicians and researchers) to measure immigration-specific distress. This scale, however, is written in English and has only been tested with former Soviet immigrants in the Boston area of United States of America (USA) for psychometrics. AIM: This instrumentation pilot study is designed to evaluate the readability and psychometric properties of the Chinese version of the DI Scale. INSTRUMENT: This selected scale is a 23-item, 6-point Liket type scale. Six dimensions are included: loss, novelty, occupation adjustment, language accommodation, discriminations, and not feeling at home. High scores indicate high levels of distress related to the demands of immigration. METHODS: The study uses a descriptive, cross-sectional design with a multimethod approach. Seventy-three Taiwanese-Chinese immigrants (> or = 18 years) in the USA responded to the scale and a demographic questionnaire. Eighteen of them contributed to the interview data that were collected for assessing content validity of the scale. Observations during the interviews and participants' questions were also documented for the evaluation. Scale format, wording of items, distribution of responses, internal consistency, test-retest reliability, content validity, and construct validity are examined. FINDINGS AND CONCLUSION: Analyses suggest that the Chinese version is easy to read and understand. The internal consistency and test-retest reliability are satisfactory. This scale could be used with Taiwanese Chinese immigrants as a generic measure of immigration-related distress. Nonetheless, three main problems with its use with Taiwanese-Chinese immigrants and Chinese immigrants at large are revealed in the study. Each problem is discussed. Suggestions for further development of the Chinese DI Scale are addressed. PMID- 12121529 TI - From practice to theory: tightening the link via three fieldwork strategies. AB - BACKGROUND: Theory that is directly linked with clinical experience has far greater relevance for nursing practice; however, problems continue to persist with the establishment of that linkage. The use of fieldwork as a strategy for developing and linking theory has enormous potential for enhancing the number of theories that truly fit the clinical practice realities of nursing. Traditionally, fieldwork has been conducted in contexts that are unfamiliar to the researcher. However, nurses have a wealth of clinical understanding in their areas of practice. Conduct of inquiry in a familiar practice area may facilitate identification of significant, clinically relevant concepts and expedite theory development that is closely linked to nursing practice. AIM OF THE PAPER: wo-step fieldwork approach and three specific strategies are presented that have been found useful for developing theory that is tightly aligned with nursing practice. The strategies of theoretical selectivity, theoretical integration, and theory creation are described and an example of each presented. The strategies are an extension of the Hybrid Model of Concept Development. These strategies identify alternative paths for concept clarification and theoretical congruence through fieldwork in clinical settings. The focus of theoretical selectivity is elaboration of concept definition with selection among theories. In contrast, the focus of theoretical integration is the linking of a central concept with an existing theory or perspective in the prefieldwork phase and then using the full fieldwork to evaluate its usefulness. Lastly, the theory creation strategy emphasizes describing a phenomenon of interest, identifying influencing factors, and beginning to create relational statements. It is closest to standard inductive theory development. PMID- 12121530 TI - Issues in educating health professionals to meet the diverse needs of patients and other service users from ethnic minority groups. AB - AIM: The main aim of the study was to undertake training needs analysis among a multi-professional group for the purpose of improving care for ethnic minority patients and other service users. BACKGROUND: Evidence from the literature identifies that some of the explanations advanced for the failure of health professionals to meet the needs of ethnic minorities include lack of understanding of cultural diversities, racism, racial stereotyping, lack of knowledge, exclusivity, and ethnocentrism. While these issues have been addressed in different countries, little work has been carried out to examine these from the perspective of health professionals caring for ethnic minorities. This study is therefore an attempt to find out what health professionals know about caring for patients and other service users from minority ethnic groups and their perception of training needs in this area of work. METHODS: A pre- and post training design phase structured the qualitative approach. A purposive sample of individuals working across five health service organizations located in a multi racial city yielded a multi-professional group of participants. Views of 22 participants were obtained by semi-structured interviews at a pretraining phase. Training needs of health professionals drew on Walklin's (1992) six stages used to structure data collection, data analysis and delivery of training. The post training phase used questionnaires to evaluate immediate learning that based on a 4-week period of reflection and applied to practice. The questionnaires were complemented by a facilitator-lead focus group. RESULTS: The majority of the participants confirmed that no attention was given in their initial education to the health care needs of minority ethnic groups. Instead, participants engaged in self-initiated learning to improve their knowledge and understanding. The issue of communication was viewed with dissatisfaction and seen as affecting the sufficiency of caring for these patients. All participants rated meeting the needs of ethnic minorities as very important and believed that they had gained a better understanding of the concepts of ethnicity and race and resources available in local communities as a result of the training. They also reported changes in thinking about ethnic minorities and had started to acquire greater confidence to engage with colleagues about different cultural values and practices and the implications of these for caring. While a quarter of the participants had transferred some learning to practice, the majority were not able to bring about any change. This majority response challenged the sustainability of learning about ethnic minorities when training takes place away from the context in which professionals practise. CONCLUSION: Training embedded in clinical and nonclinical environment where patients and other service users and professionals interact is offered as a major finding. PMID- 12121531 TI - Practice nurses and clinical guidelines in a changing primary care context: an empirical study. AB - BACKGROUND: Practice Nurses form an increasingly large proportion of the English National Health Service primary care workforce and the delegation to them of clinical work from General Practitioners has attracted some academic attention. Central to this process are clinical guidelines, which provide the interface between the movement towards 'evidence-based practice' and a range of government driven policy developments in primary care. AIMS: To identify the attitudes of practice nurses to clinical guidelines; to investigate the impact of guidelines on nurse/physician relationships; and to describe the impact of the changing primary care context on nurses. METHODS: We interviewed a sample of 29 Practice Nurses three times during a 16-month period to clarify their attitudes towards guidelines, their use of guidelines in practice and their assessment of guidelines' importance. We gathered further data on organizational culture and perceptions of national reforms of primary care structures. RESULTS: We found that practice nurses are generally supportive of clinical guidelines. Moreover, nurses' role and influence within primary care is in a process of transition to one in which they may undertake responsibility for influencing General Practitioners' clinical behaviour so as to adhere to guidelines. Practice nurses themselves recognize and welcome this, though with some reservations. CONCLUSIONS: Our findings support the proposal that explicit codification of the scientific basis of the work of lower paid groups may enhance their relative professional status. PMID- 12121533 TI - Cytomegalovirus and cyclosporin-induced gingival overgrowth in children with liver grafts. AB - OBJECTIVE: To determine whether cytomegalovirus (CMV) is associated with gingival overgrowth in paediatric liver graft recipients treated with cyclosporin. STUDY DESIGN: Thirty-four children, 25 of whom were under 5 years of age, who had undergone liver transplantation, were examined. An Index of Severity of Gingival Overgrowth was used to measure the prevalence and severity of the gingival overgrowth. The trough cyclosporin level was recorded and the CMV status of the patient matched to the dental findings. The association between the severity of gingival overgrowth and CMV infection was examined using the contingency coefficient. An anova was used to assess the association between the circulating trough cyclosporin concentration and the severity of gingival overgrowth. Pearson's Product Moment Correlation Coefficient was used to examine the association between the duration of exposure to cyclosporin and the severity of gingival overgrowth. RESULTS: There was a significant inverse association between the duration of exposure to cyclosporin and the severity of gingival overgrowth. There was no relationship between the trough cyclosporin concentration and the severity of gingival overgrowth. There was no association between CMV and gingival overgrowth. CONCLUSION: Gingival overgrowth was related to the duration of cyclosporin therapy but was neither more prevalent nor more severe in subjects who were CMV seropositive. PMID- 12121535 TI - Parental anxiety as a possible predisposing factor to child dental anxiety in patients seen in a suburban dental hospital in Nigeria. AB - INTRODUCTION: The aim of this study was to determine the relationship between parents' anxiety level and that of the child patient. METHODS: The Short Form of the Dental Anxiety Survey Schedule was administered to 81 children who were attending the dental clinic for the first time. The Dental Anxiety Scale was also used to collect relevant information from the parents. RESULTS: There was no statistically significant correlation between the anxiety level of the mother (r = -0.02, P = 0.82) or the father (r = -0.59, P = 0.62) and that of their child. However, bivariate analysis showed a closer association between the anxiety levels of the mother and the child (P = 0.055) compared to that between the father and the child (P = 0.475) although this was again found not to be statistically significant. CONCLUSION: Assessment and management of the anxiety level of the mother may be needed in some cases, both to manage the child effectively and to break the cycle of dental care anxiety in families. PMID- 12121534 TI - Supernumerary teeth: review of the literature and a survey of 152 cases. AB - INTRODUCTION: A review of the literature relating to supernumerary teeth is presented along with a survey of 152 cases. METHODS: The study population consisted of 152 children who visited the department of Paediatric Dentistry at the Jordan University Hospital. Patients ranged in age from 5 to 15 years. Supernumeraries were detected by clinical examination and radiographs. RESULTS: Males were affected more than females with a sex ratio of 2.2 : 1. Seventy-seven percent of the patients had one supernumerary tooth, 18.4% had double teeth, and 4.6% had three or more supernumeraries. Ninety percent of the supernumerary teeth occurred in the premaxilla, of which 92.8% were in the central incisor region and of these latter 25% were located in the midline. The other 10.4% of the supernumeraries were located in the premolar, canine, molar, and lower central incisor regions. Two cases were of non-syndrome supernumerary teeth. Seventy-five percent of the supernumeraries were conical, 83.1% were in the normal vertical position and 26.5% were erupted. Conical-shaped supernumerary teeth had a significantly higher rate of eruption compared to the tuberculate type. PMID- 12121536 TI - General anaesthesia for exodontia in children: experience of a dental teaching hospital in relation to changes in national guidance. AB - OBJECTIVES: To examine, in relation to changes in national guidance, trends in the provision of general anaesthesia for exodontia in children. DESIGN: Retrospective analysis of hospital records. SETTING: University Dental Hospital, Cardiff, UK. METHODS: Data were drawn from records of services provided in each of four index months during 1989, 1991, 1997 and 1999 (the years immediately preceding/following the Poswillo Report of 1990 and the General Dental Council's revised guidance of 1998). RESULTS: In the first year after publication of the Poswillo report, there was a 19.6% increase in use of 'chair case' general anaesthesia over the four index months. Subsequently, upon the emergence of additional providers, patient flow in 1997 had fallen to 65.8% of that seen at baseline. Following the General Dental Council's revised guidance of 1998, there was a further reduction in the provision of 'chair case' general anaesthesia (to 40.8% of that seen at baseline). CONCLUSIONS: In this unit, throughput of children receiving general anaesthesia for exodontia has fluctuated in periods marked by changing national guidance. However, it is not possible to attribute such fluctuations in patient flow solely to this factor. PMID- 12121537 TI - Sealing ability of a single bond adhesive in primary teeth. An in vivo study. AB - AIM: To compare the ability of a single bond adhesive to its preceding multiple step dentin bonding agent in reducing microleakage around class V composite resin in primary teeth. MATERIAL AND METHODS: Twelve children, between 9 and 11 years old, with non-carious primary canines indicated for extraction for orthodontic purposes, were selected for this study. Each child had facial surfaces of his or her upper and/or lower canines prepared for class V cavities. One canine was randomly selected for restoration using the Scotchbond Multi Purpose Plus System (group I) and its antimere for the Single Bond adhesive (group II). Thirty cavities were restored by Z-100 composite. Materials were handled according to the instructions of the manufacturer. All teeth were extracted one month later, immersed in 2% basic fuchsin then sectioned to evaluate dye penetration. RESULTS: No statistically significant difference was found in the degree of microleakage between the two materials. CONCLUSIONS: Neither of the two adhesive systems was able to completely prevent leakage of class V restorations. Additional preventive measures should be implemented whenever using composite resin restorations in children. PMID- 12121539 TI - Teeth grinding, tongue and lip biting in a 24-month-old boy with meningococcal septicaemia. Report of a case. AB - This paper describes the management of a 24-month-old boy who presented with self inflicted trauma to his lower lip and tongue, and teeth grinding, 21 days after developing meningococcal septicaemia. A decision to observe and prescribe palliative therapy was made. Extraction of the lower right deciduous canine, which had become non-vital, possibly due to bruxism, was carried out. PMID- 12121538 TI - Oral submucous fibrosis in a 12-year-old Bangladeshi boy: a case report and review of literature. AB - Oral submucous fibrosis is commonly seen in the adult population of the ethnic minorities in the UK, although its presentation in a child is rare. Whilst the condition is considered multifactorial and irreversible, we present a case of oral submucous fibrosis in a 12-year-old Bangladeshi boy whose cessation of habitual betel nut chewing and forcible mouth chewing exercises led to an improvement in his mouth opening, although his ability to protrude his tongue remained unaltered. The clinical features, pathogenesis and management of submucous fibrosis are described. The widespread use of betel quid among Asians in the UK is summarized and the importance of its recognition as a precancerous condition is emphasized. PMID- 12121540 TI - Impacted supernumerary tooth developed under palatal polyp. AB - We present a rare case of an infant's palatal polyp associated with an impacted supernumerary tooth. We have previously reported three cases of palatal polyps in infants [1]. In one case, after surgical removal of the palatal polyp at the age of 1 year and 8 months, the lesion subsequently began swelling. A periapical radiograph at the age of 2 years and 9 months revealed a small calcified mass in the maxillary left incisor region. The swelling was kept under observation, the calcified mass developed gradually and was removed surgically at the age of 5 years and 5 months. The removed calcified mass was clinically diagnosed as an impacted supernumerary tooth and this was confirmed histologically. Histological findings did not indicate any relationship between the palatal polyp and the impacted supernumerary tooth. PMID- 12121545 TI - Polymorphous light eruption. PMID- 12121541 TI - Space-regaining treatment for a submerged primary molar: a case report. AB - The aetiology of submerged primary molars is not known and optimal treatment has still not been established. If submerged primary teeth are left untreated, the occlusal consequences are space-loss due to tipping of adjacent teeth and/or overeruption of the opposing teeth. Normally, treatment consists of extraction of the deciduous teeth or observation for normal exfoliation. Here we report a case where the submerged primary molar was extracted surgically after space-loss had been regained. PMID- 12121546 TI - Becker's melanosis associated with fibrous dysplasia. PMID- 12121547 TI - Infraorbital eyelid edema as the presenting sign of bronchogenic carcinoma. PMID- 12121548 TI - Purpura annularis telangiectoides with vasculitic ulcers treated with colchicine. PMID- 12121549 TI - Overlap collagen vascular disease as a marker for development of primary biliary cirrhosis. PMID- 12121550 TI - Exercise-induced anaphylaxis in a marathon runner. PMID- 12121551 TI - Anonychia congenita totalis: a case report and review of the literature. PMID- 12121552 TI - Mees' lines in a patient following acute arsenic intoxication. PMID- 12121553 TI - Development of localized myxedema in a skin graft. PMID- 12121554 TI - Cutaneous Rosai-Dorfman disease preceding inguinal lymphadenopathy. PMID- 12121555 TI - Rejuvenating facial massage--a bane or boon? AB - BACKGROUND: Facial massage is an extremely popular form of beauty treatment and is thought to rejuvenate the skin. We decided to study the benefits and untoward effects of this form of facial beauty treatment. METHODS: One hundred and forty two women (aged 17-63 years), who had received facial beauty treatment in three well-established beauty parlours in New Delhi, were entered into the study and observed for a period of 12 weeks after the facial beauty treatment. Twenty-seven of the subjects had a repeat facial beauty treatment 4-6 weeks after entry into the study, giving a total of 169 massage episodes observed. Immediate and delayed effects of the beauty treatment were examined. RESULTS: Facial beauty treatment generally consists of three steps: vigorous massaging of the face with creams, steaming (using a hot towel or a steaming gadget), and application of a face mask containing adsorbents and astringents. In our study, the creams used for massage included "off the shelf" creams manufactured by standard cosmetic companies in 95 (56.3%) subjects, herbal creams in 61 (36.1%), and creams containing exotic ingredients, such as gold salts, in 13 (7.7%). Sixty-one (36.1%) patients developed erythema and puffiness within 15 min to 2 h after the beauty treatment. This lasted for 2-6 h. Forty-one (24.3%) women underwent the procedure of comedone extraction after steaming. In 12 (7.1%) of these women, persistent erythema was noticed at the site of comedone extraction. Eight (4.7%) women developed mild dermatitis on the face, 2-7 days after the facial beauty treatment. Patch testing with constituents used in the facial beauty treatment was positive in four patients (herbal cream, 1; witch hazel, 1; orange face pack, 1; and gold cream, 1). In 47 (33.1%) subjects, an acneiform eruption was observed 3-10 weeks after the facial beauty treatment (mean, 6.1 +/- 3 weeks). Thirteen (27.7%) of these subjects had taken the facial beauty treatment for the first time, whereas 34 (72.3%) developed an acneiform eruption after every facial massage. The predominant lesions were deep-seated nodules, although a few comedones, especially closed ones, were present in some patients. Lesions were always present on the cheeks, an area of focus during the facial massage, and healed with hyperpigmentation. The benefits of facial beauty treatment, as mentioned by the subjects, included a feeling of freshness and rejuvenation in 84 (59.1%), keeping the skin supple in 76 (53.5%), feeling of warmth and tightening of the skin in 71 (50%), and delaying the onset of wrinkles in 21 (14.8%). CONCLUSIONS: Although there are several subjective benefits with facial beauty treatment, there may be immediate side-effects, such as erythema and edema, as well as delayed problems, such as dermatitis and acneiform eruption, in about one third of patients. PMID- 12121557 TI - Squamous cell carcinoma in a patient with Netherton's syndrome. AB - A 29-year-old white woman with a history of Netherton's syndrome presented with two squamous cell carcinomas on the right dorsal hand and the left upper arm. She reported a 2-year history of these lesions, which were originally treated as warts. She denied excessive sun exposure, immunosuppressive therapy, or a previous history of skin cancer. Her past medical history included acute renal failure, multiple urinary tract infections, meningitis, and recurrent otitis media as a child. In addition, she had an ovarian abscess at 4 years of age with resulting salpingo-oophorectomy. She also reported a history of severe myopia, glaucoma, and multiple ocular infections with a resulting corneal scar. In addition to atopic dermatitis, she had a 10-year history of psoriasis. Her medications included topical steroids and emollients for atopic dermatitis and psoriasis, in addition to Timolol ophthalmic drops for glaucoma. Her family history was significant for a 22-year-old sister with Netherton's syndrome (Fig. 1). She denied any history of skin cancer in her sister or other members of her family. On physical examination, she had an exfoliative erythroderma, madarosis, and diffuse patchy alopecia. In the bilateral axilla, she had well-defined pink scaly plaques which were confirmed as psoriasis by biopsy. On the right dorsal hand, she had a 1.5 x 1.0 cm pink verrucous plaque (Fig. 2). On the left upper arm, she had a 1.5 x 0.8 cm pink scaly plaque. Biopsies of both sites confirmed squamous cell carcinomas. Both lesions were completely excised with 4 mm margins. PMID- 12121556 TI - Prayer marks. AB - BACKGROUND: Prayer marks (PMs) are asymptomatic, chronic skin changes that consist mainly of thickening, lichenification, and hyperpigmentation, and develop over a long period of time as a consequence of repeated, extended pressure on bony prominences during prayer. METHODS: Three hundred and forty-nine Muslims and 24 non-Muslims were examined for the appearance of PMs at different body sites. RESULTS: The prospective study of 349 Muslims (both males and females) with regular praying habits showed the occurrence of PMs on specific locations, such as the forehead, knees, ankles, and dorsa of the feet, leading to dermatologic changes consisting of lichenification and hyperpigmentation. The incidence of PMs was significantly higher in males than in females. Older subjects (over 50 years of age) demonstrated a significantly higher frequency of lichenification and hyperpigmentation, suggesting that repeated pressure and friction for prolonged periods are the causative factors for the development of PMs. Histologic examination of skin biopsies from the affected sites showed compact orthokeratosis, hypergranulosis, dermal papillary fibrosis, and dermal vascularization. PMs were not associated with any risk of secondary complications, such as erythema, bullous formation, and infections. CONCLUSIONS: PMs are commonly occurring dermatologic changes in Muslims who pray for prolonged periods. PMID- 12121558 TI - Autoimmune and rheumatic manifestations and antinuclear antibody study in HIV infected Thai patients. AB - BACKGROUND: There have been reports concerning an association between human immunodeficiency virus (HIV) infection and autoimmune and rheumatic diseases. OBJECTIVE: The purpose of this study was to examine autoimmune and rheumatic manifestations in HIV-infected patients and their correlation with antinuclear antibody (ANA) tests. METHODS: The clinical and laboratory results of HIV infected patients attending the Department of Dermatology, Siriraj Hospital, Bangkok, Thailand, from February 1999 to January 2000, were analyzed. Laboratory studies included serum CD4 lymphocyte count and ANA tests. RESULTS: Sixty-two patients were enrolled prospectively in the study. Myalgia was the most common clinical presentation (50%). Others included photosensitivity (on history) (39%), arthralgia (26%), vasculitis (18%), sicca complex (10%), arthritis (7%), and Reiter's syndrome (2%). A history of hair loss was given by 23% of patients. A positive ANA test was detected in 3%. No cases of systemic lupus erythematosus, scleroderma, or dermatomyositis were seen. CONCLUSIONS: Autoimmune and rheumatic manifestations were not uncommonly detected in patients with HIV infection. HIV infection may sometimes mimic systemic lupus erythematosus clinically. PMID- 12121560 TI - Treatment of axillary hyperhidrosis with botulinum-A toxin. AB - BACKGROUND: Severe axillary hyperhidrosis is a source of great embarrassment and considerable emotional stress to individuals afflicted with this condition. Existing topical and surgical therapies are either ineffective or associated with unacceptable morbidity. We attempt to determine the effect of botulinum-A toxin (Dysport) in the treatment of axillary hyperhidrosis. PATIENTS AND METHODS: After visualization of hyperhidrosis using the iodine-starch test, 10 patients with axillary hyperhidrosis underwent intradermal injection with 125 units of Dysport on each axilla. Patients were observed for 7 months after treatment. RESULTS: The treatment was well tolerated without side-effects. All patients experienced relatively complete anhidrosis of the axillary skin after about 1 week for periods ranging from 4 to 7 months. CONCLUSIONS: Botulinum-A toxin may offer a fast, safe, and highly effective therapeutic option for severe hyperhidrosis. PMID- 12121559 TI - Childhood lichen planus: a study of 87 cases. AB - AIMS: We undertook this study to analyze the clinical profile of childhood lichen planus (LP) prevailing in north India, and to highlight differences from and similarities with adult LP. METHODS: Clinical records of children with LP, who attended the Pediatric Dermatology Clinic, Nehru Hospital, Postgraduate Institute of Medical Education and Research, Chandigarh, India, from July 1988 to December 2000, were analyzed. RESULTS: Eighty-seven patients with LP were examined during a 12.5-year period of observation. These patients formed 2.5% of the total number of pediatric dermatology patients and 0.6% of the total number of new dermatology outpatients. The male : female ratio was 1.1 : 1. The age at onset was between 8 months and 12 years (mean, 7.1 years). Classical LP was observed in 53 (60.9%) patients, followed by actinic LP in 10 (11.5%), and lichen planus hypertrophicus (LPH) and linear LP in eight (9.2%) each. Other forms seen were eruptive, follicular, lichen planopilaris, atrophic, and bullous LP. The involvement of skin alone was observed in 75 (86.2%) children and mucosa alone only in one (1.1%) child. Concomitant skin and mucosal involvement was seen at the time of presentation in four patients, while mucosal involvement occurred later during the course of the disease in seven patients. Lesions appeared earlier in boys than in girls. CONCLUSIONS: The natural history of LP in children was essentially similar to that in adults. Unusual features, such as involvement of the palms and soles and upper eyelids, were observed. Actinic LP, mimicking melasma, as reported in adult women, also seems to occur in children. PMID- 12121561 TI - Evaluation of inflammatory parameters in physical urticarias and effects of an anti-inflammatory/antiallergic treatment. AB - BACKGROUND: Physical urticaria (PU) includes a heterogeneous group of urticarias whose etiopathogenic aspects are still obscure and whose therapeutical management is often difficult. We have previously demonstrated the efficacy of a sequential treatment with nimesulide, a unique nonsteroidal anti-inflammatory drug, and ketotifen in various forms of PU. METHODS: The expression of some inflammatory parameters was evaluated in 10 patients affected with some forms of PU. In particular, serum levels of interleukin (IL)-4, IL-1beta, tumor necrosis factor (TNF)-alpha, adhesion molecules (sELAM, sICAM-1 and sVCAM), soluble receptors (sIL-2R, sCD30, sCD23) and IgE were assessed. Moreover, the cutaneous expression of IL-1beta, TNF-alpha and ICAM-1 was studied on biopsy specimens taken from nonlesional skin of patients. The same parameters were further evaluated in both skin and serum following treatment with nimesulide and ketotifen, in order to better understand the possible effects of these agents on the inflammatory network of PU. RESULTS: Before therapy we could detect significantly higher serum levels of IL-1beta and TNF-alpha (P < 0.001) and of circulating adhesion molecules in comparison with controls (sELAM, sICAM: P < 0.001; sVCAM: P < 0.02); after treatment, a significant reduction of each was observed (P < 0.05). Simultaneously, a high expression of IL-1beta, TNF-alpha and ICAM-1 was detected in all skin specimens at the baseline, with a relevant decrease following therapy. CONCLUSIONS: These results confirm the therapeutical value of treatment with nimesulide and ketotifen in PU and suggest that these agents are able to reduce the release and the expression of some inflammatory molecules that are up regulated in PU. PMID- 12121562 TI - Unusual location of cutaneous leishmaniasis on the hallux in a Brazilian patient. AB - A 70-year-old white Brazilian woman from a rural area had a 2-year history of a painful lesion on her left toe. The lesion increased progressively in size followed by toenail destruction. She was treated with systemic antibiotics for secondary bacterial infection several times without any clinical response. Physical examination showed an erythematous swelling on the first toe with an irregular 2.5 cm ulcer with a raised edge that was infiltrating and destroying the toenail. The bottom of the ulcer was granular and partially covered with a crust (Fig. 1). Laboratory studies showed a strong positive Montenegro intradermal reaction (2.5 mm). Other intradermal reactions were also performed, such as purified protein derivative (PPD) and sporotrichin, which were negative. On X-ray examination of the left foot, bone destruction of the distal phalanx of the first toe and a soft tissue swelling were observed (Fig. 2). A biopsy was taken and the histologic picture showed a chronic inflammatory change with tuberculoid-type granuloma and necrosis suggesting leishmaniasis, although parasites were not observed. Based on the clinical, histologic, and immunologic aspects, we concluded that this was a case of leishmaniasis. Methylglucamine (Glucantime) was introduced at a total dose of 17 g of the salt (10 mg/kg daily for 40 days). Immediately after the start of treatment, the lesion began to improve, and 4 months later the lesion had healed completely and the dystrophic nail had started to grow (Fig. 3). PMID- 12121563 TI - Treatment of cutaneous leishmaniasis with a combination of allopurinol and low dose meglumine antimoniate. AB - The objective of this study was to compare the efficacy of a combination of allopurinol (AL) and low-dose meglumine antimoniate (MA) with standard-dose MA in cutaneous leishmaniasis caused by Leishmania major. An open, controlled study was performed. Seventy-two patients were randomly selected from volunteers with cutaneous leishmaniasis living in a hyperendemic area. Exclusion criteria included pregnancy, nursing vs. gestation, age less than 5 years, and duration of disease of more than 4 months. Each patient received MA (60 mg/kg/day) or AL (20 mg/kg/day) plus low-dose MA (30 mg/kg/day) for 20 days, and was followed up for 30 days after cessation of treatment. The study was completed as planned in 66 patients. Complete healing occurred in 74.2% of patients in the MA group and in 80.6% of patients in the MA + AL group. No difference was found between the two groups with respect to side-effects. The combination of AL and MA increases the anti-leishmanial effects of antimoniate. In this study, it was confirmed that low dose MA plus AL is as effective as high-dose MA in the treatment of cutaneous leishmaniasis caused by L. major. PMID- 12121564 TI - Short-term methotrexate therapy in psoriasis: a study of 197 patients. AB - Methotrexate (MTX) is one of the most effective antipsoriatic drugs available. Although it is undeniably hepatotoxic, it can be used safely in most patients with severe psoriasis if established guidelines are followed. Current opinion on the monitoring of hepatic damage is divided, however, and the need for repeated liver biopsies during MTX therapy is being re-examined. We have used MTX in a short-term protocol in our patients with psoriasis for the past 20 years, and have recently attempted to minimize or eliminate the need for liver biopsies using this regimen. Data on 244 psoriatics who were given MTX from 1981 to 2000 have been reviewed. Our protocol entailed the use of weekly oral MTX at the full therapeutic dose during episodes of peak disease activity, with tapering off of MTX when the disease subsided in response to treatment combined with natural/seasonal remission. Intensive topical and heliotherapy were encouraged throughout to facilitate the earliest possible drug withdrawal and the longest possible drug-free interval before the next relapse. Strict inclusion criteria were applied before starting MTX. A total of 243 cycles of MTX therapy have been given to 197 evaluable patients. More than 75% improvement occurred in 88% of patients in 8.5 +/- 5.1 weeks. The mean cumulative dose was 709.3 +/- 369.2 mg and the mean duration of follow-up was 16.5 +/- 9.1 months. Fifteen (6.1%) patients had serious adverse effects requiring the cessation of therapy. Only three patients had deranged liver function tests. Thirty-four pre-MTX and 13 post MTX liver biopsies were taken, which revealed grade I or II changes that were nonprogressive. Our experience with short-term MTX therapy has enabled us to safely administer MTX to our patients with minimal recourse to liver biopsy. In developing countries, where advanced noninvasive methods for the assessment of liver damage are unaffordable or unavailable, this interrupted, short-term regimen may present an acceptable and safe method of using MTX in carefully selected patients with severe psoriasis. PMID- 12121565 TI - Rapid response to infliximab in severe pustular psoriasis, von Zumbusch type. AB - Tumor necrosis factor-alpha (TNF-alpha) is a chemokine secreted by T cells which is thought to play a critical role in the pathophysiology of psoriasis. The monoclonal antibody, infliximab, complexes with TNF-alpha, rendering it inactive. A recent clinical trial has reported the clinical benefit and safety of infliximab in moderate to severe plaque psoriasis. We report a case of rapid response and clinical benefit using infliximab in severe pustular psoriasis of von Zumbusch. PMID- 12121566 TI - A novel sustained-release matrix based on biodegradable poly(ester amide)s and impregnated with bacteriophages and an antibiotic shows promise in management of infected venous stasis ulcers and other poorly healing wounds. AB - Healing of poorly vascularized and venous stasis ulcers is often refractory to therapy, particularly when they are infected. Systemic antibiotic therapy may be of little benefit in this setting because of poor penetration of the antibiotic into the wound and the frequent associated emergence of bacterial strains resistant to common antimicrobial agents. Given the clinical significance of these problems, there is a need to explore alternative management approaches for these difficult-to-treat wounds. PhagoBioDerm is a novel wound-healing preparation consisting of a biodegradable polymer impregnated with an antibiotic and lytic bacteriophages, which was recently licensed for sale in the Republic of Georgia (one of the former Soviet Union republics). In 1999-2000, in Tbilisi, Georgia, 107 patients who had ulcers that had failed to respond to conventional therapy were treated with PhagoBioDerm alone or in combination with other interventions. The wounds/ulcers healed completely in 67 (70%) of 96 patients for whom follow-up data were available. In 22 cases in which microbiologic data were available, healing was associated with the concomitant elimination of, or a reduction in, specific pathogenic bacteria in the ulcers. Our findings suggest that this slow-release biopolymer is safe and of possible benefit in the management of refractory wounds, and they support the apparent utility of bacteriophages in this setting. Further studies, including carefully designed clinical trials, will be required to rigorously evaluate the efficacy of this novel wound dressing preparation. PMID- 12121567 TI - Reactive oxygen species generation by human spermatozoa: a continuing enigma. PMID- 12121568 TI - Impact of obesity on hypogonadism in the andropause. AB - Obesity is an issue that is increasingly affecting ageing men. With ageing, there is a decline in androgens as well. There are implications for the health of ageing men as a result of hypogonadism. Overall, there seems to be an inverse relationship between body mass index and testosterone levels, as is also demonstrated in our cross-sectional study. Obesity seems to depress the production of testosterone. It has been hypothesized that there is increased aromatization of testosterone to oestradiol and alteration of the hypothalamic pituitary-adrenal axis in obese older men. Some hormones can affect obesity in ageing men including leptin, insulin, dehydroepiandrostenedione and growth hormone. The relationship of obesity to these hormones in ageing men will be reviewed. Testosterone replacement in ageing men can alter body composition whereby fat is exchanged for muscle. These studies will also be reviewed. Further studies in this field are recommended to evaluate long-term benefits and risks. PMID- 12121569 TI - Quantification of cyclin A1 and glyceraldehyde-3-phosphate dehydrogenase expression in testicular biopsies of infertile patients by fluorescence real-time RT-PCR. AB - The objective of this study was to establish a classification of meiotic disorders by detection of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and cyclin A1 mRNA in patients presenting with non-obstructive azoospermia. GAPDH and cyclin A1 expression levels were detected in 37 histologically classified testicular tissue specimens by one-line RT-PCR. Levels of cyclin A1 expression (NCyclinA1) were high in tissue specimens with full spermatogenesis [3.519 +/- 3.141 (mean +/- SD)] but only minimal in those without haploid germ cells. A lack of cyclin A1 expression was seen in most sertoli-cell-only syndrome (SCOS) specimens. The criteria for an ideal internal standard were only partially met by GAPDH, as we detected decreased expression in tissue samples with maturation arrest [(268.2 +/- 106.2 relative gene expression (mean +/- SD)] and SCOS (201.7 +/- 95.2) compared to those with normal histological findings (325.1 +/- 129.3). It was concluded that the level of cyclin A1 mRNA expression predicts meiotic disorders during spermatogenesis. GAPDH is not an ideal internal standard for specific gene expression in testicular tissue specimens. PMID- 12121570 TI - Effects of dihydrotestosterone on cardiac inward rectifier K(+) current. AB - There are well-described sexually dimorphic differences both in the electrocardiogram and in the propensity to develop drug-induced arrhythmias. The QT interval and the risk of ventricular proarrhythmia are reduced in males compared with females. Inward rectifier potassium current (IK(1)) is a primary determinant of the ventricular resting membrane potential, and an important contributor to myocardial excitability. METHODS AND RESULTS: Using the whole-cell patch-clamp technique, we evaluated the effects of dihydrotestosterone (DHT) on IK(1) in ventricular myocytes from castrated rabbits that were treated with either replacement DHT or vehicle-control for 3 weeks. Compared with the DHT treated group, myocytes from the control animals had a significant reduction in inward IK(1) conductance (p < 0.005) and rectification ratio (RR) (p < 0.04) with no significant change in peak outward current. Acute DHT superfusion of the myocytes increased inward IK(1) conductance from baseline (p < 0.05) and increased the RR (p < 0.05). Testosterone has been reported to increase intracellular ornithine decarboxylase activity in ventricular tissue, which would increase intracellular polyamines, known modulators of IK(1) rectification. We found that inclusion of the intracellular polyamines spermidine and putrescine in the pipette solution caused a decrease in inward IK(1), accompanied by an increase in peak outward current and a reduction in the RR. CONCLUSION: In summary, DHT modulates IK(1) in a chronic, as well as, an acute fashion. These effects are not because of altered intracellular polyamines. DHT may modulate myocardial excitability through effects on IK(1). PMID- 12121571 TI - Evaluation of the acrosome reaction and viability in buffalo spermatozoa using two staining methods: the effects of heparin and calcium ionophore A23187. AB - In this study, we investigated the effect of heparin and calcium ionophore A23187 on in vitro induction of buffalo sperm acrosome reaction (AR). Two methods for detection of the AR and viability were employed. Fluorescein isothiocyanate conjugated Arachis hypogea agglutinin (FITC-PNA) was used as a vital stain in combination with ethidium homodimer-1 (EthD-1) to determine the acrosome status of viable spermatozoa. In another experiment, trypan blue replaced EthD-1 to differentiate live and dead spermatozoa having undergone AR. The results from the two methods were significantly correlated (r > 0.9). Four different staining patterns were found in both methods. The FITC-PNA intensely labels the acrosome region of acrosome-intact spermatozoa. EthD-1 and trypan blue stained red and blue at the post-acrosomal region of dead spermatozoa, respectively. Spermatozoa incubated with heparin showed a significant increase ( p < 0.05) in the percentage of live acrosome-reacted sperm after 30 min incubation period. This trend continued and was significantly different over the entire incubation period when compared with the control group at the same interval. In the ionophore treated group, the proportion of changes in live acrosome-intact and live acrosome-reacted spermatozoa was statistically significantly different ( p < 0.001) when compared with those treated with heparin at the same interval. The AR occurred sooner and to a greater extent when incubated with the ionophore but at 5 h of incubation the percentage of false acrosomal reaction was significantly higher than those in the control and heparin-treated groups. The results in this study indicated that the in vitro induction of AR by heparin and calcium ionophore evaluated by both methods could be used to assess sperm fertilizing capacity for in vitro fertilization of this species. PMID- 12121572 TI - A comparison of the NADPH oxidase in human sperm and white blood cells. AB - The mechanism of reactive oxygen species (ROS) generation in human sperm has recently been found to depend upon a novel NADPH-oxidase (NOX5) resembling the multicomponent NADPH-oxidase of white blood cells (WBCs). The purpose of our study was to compare the ROS producing activity of NOX5 in sperm and NADPH oxidase of WBCs, and to investigate the role of protein kinase C (PKC) in NOX5 activation. A combination of electron paramagnetic resonance (EPR), chemiluminescence (CL), and nitroblue tetrazolium (NBT) dye reduction were used to monitor ROS production by sperm. The involvement of PKC in NOX5 activation was investigated using myristate acetate (PMA), and the PKC inhibitor GF-109203X. The presence of b cytochrome in NOX5 was investigated by spectrophotometry. PMA stimulated WBCs produced superoxide dismutase -- inhibitable EPR signals for both superoxide and hydroxyl radicals. Sperm did not produce these spectra with or without PMA stimulation. WBCs generated significantly increased levels of CL and reduced NBT after PMA stimulation; whereas sperm did not increase the CL response or reduce NBT. Adenosine triphosphate (ATP) levels in WBCs were significantly reduced after PMA stimulation, whereas sperm ATP levels did not change. The characteristic spectra of b cytochrome observed after dithionite reduction of WBCs was not observed with sperm under similar conditions. These results indicate that the ROS producing activity of NOX5 is significantly lower than the WBC NADPH oxidase, and suggest that the activation mechanism of NOX5 in sperm is independent of PKC. PMID- 12121573 TI - Interactive effect of semen and cervical mucus quality on postcoital test outcome: analysis from an andrological point of view. AB - This study analysed the relationship between semen quality and the postcoital test (PCT) outcome in 616 couples, especially focusing on the interactive effect of semen and cervical mucus quality. When PCTs performed in the presence of unfavourable mucus were excluded, a significant correlation was found between semen parameters and PCT outcome. In oligo/asthenozoospermia, 46.7% of PCT outcomes were negative, while the remaining were positive. Notably, in the presence of an optimal mucus score, 39% of PCT outcomes were good (> or = 7 forward motile spermatozoa/high power field). In normozoospermia, 16% of PCT were negative. A suboptimal cervical mucus quality significantly affected the PCT outcome in the presence of oligo/asthenozoospermia, but not in normozoospermia. In couples with repeated PCT, a better mucus score was associated with a significant improvement of the PCT outcome. When the outcome of two PCTs performed in the same couples with an unmodified mucus score was compared, a good consistency of the results was observed. In conclusion, the PCT can provide information in additional to that obtained from conventional semen analysis, as the interactive effect of semen/cervical mucus cannot be accurately inferred from the separate evaluation of the two members of a couple. PMID- 12121574 TI - Regional differences in semen qualities in the Baltic region. AB - Recent prospective studies of male reproductive health have shown differences between several European countries. Our objective was to evaluate the current situation in the two Baltic States Estonia and Lithuania. In 1997-99 we investigated semen parameters, levels of reproductive hormones and general health factors of 196 men from the general population in Lithuania; from Estonia, 79 men from the general population and 118 soldiers were investigated. Adjusted for interlaboratory differences and abstinence period, sperm concentration of Lithuanian and Estonian men from the general populations were shown to be 55 and 67 million/mL, respectively. The Estonian soldiers had the highest sperm concentration, 82 million/mL. The frequencies of morphologically normal spermatozoa were 6.2, 7.7 and 9.6%, respectively. In contrast to the semen qualities, highest Inhibin B levels were detected in the Lithuanians (233 pg/mL) followed by Estonian men from the general population (220 pg/mL) and Estonian soldiers (185 pg/mL). The soldiers had also the lowest level of testosterone and oestradiol. The sperm counts of the Estonian and Lithuanian men investigated here are higher than recently shown for Norwegian, Danish, Estonian and Finnish men. Comparisons should be cautiously drawn as the groups are not completely comparable. Still, even within the Baltic region, geographically close and sharing common recent social history, differences in semen quality and levels of reproductive hormones are apparent. PMID- 12121575 TI - Re: Gonzales, G. F. & Villena, A. (2001) True corrected seminal fructose level: a marker of the function of seminal vesicles in infertile men. International Journal of Andrology 24, 255-60. PMID- 12121577 TI - Cloning and characterisation of hAps1 and hAps2, human diadenosine polyphosphate metabolising Nudix hydrolases. AB - BACKGROUND: The human genome contains at least 18 genes for Nudix hydrolase enzymes. Many have similar functions to one another. In order to understand their roles in cell physiology, these proteins must be characterised. RESULTS: We have characterised two novel human gene products, hAps1, encoded by the NUDT11 gene, and hAps2, encoded by the NUDT10 gene. These cytoplasmic proteins are members of the DIPP subfamily of Nudix hydrolases, and differ from each other by a single amino acid. Both metabolise diadenosine-polyphosphates and, weakly, diphosphoinositol polyphosphates. An apparent polymorphism of hAps1 has also been identified, which leads to the point mutation S39N. This has also been characterised. The favoured nucleotides were diadenosine 5',5"'-pentaphosphate (kcat/Km = 11, 8 and 16 x 10(3) M(-1) x s(-1) respectively for hAps1, hAps1-39N and hAps2) and diadenosine 5',5"'-hexaphosphate (kcat/Km = 13, 14 and 11 x 10(3) M(-1) x s(-1) respectively for hAps1, hAps1-39N and hAps2). Both hAps1 and hAps2 had pH optima of 8.5 and an absolute requirement for divalent cations, with manganese (II) being favoured. Magnesium was not able to activate the enzymes. Therefore, these enzymes could be acutely regulated by manganese fluxes within the cell. CONCLUSIONS: Recent gene duplication has generated the two Nudix genes, NUDT11 and NUDT10. We have characterised their gene products as the closely related Nudix hydrolases, hAps1 and hAps2. These two gene products complement the activity of previously described members of the DIPP family, and reinforce the concept that Ap5A and Ap6A act as intracellular messengers. PMID- 12121578 TI - "Sheepish about being Bullish". PMID- 12121579 TI - Recent advances in the management of superficial bladder tumors. AB - Superficial bladder tumors are by far the most common form of bladder cancer managed by practicing urologists. Indeed up to 75% of initial tumors fall in this category and because of the high recurrence rate, superficial tumors represent over 90% of tumor events being treated. New diagnostic methods have been developed to improve the sensitivity of tumor detection of both cystoscopy and urinary cytology. Risk stratification of patients based on simple clinical parameters provides new opportunities for adapting monitoring strategies as well as providing a rationale for the use of intravesical chemotherapy and immunotherapy. Finally, recent pathological substratification of stage T1 grade 3 may allow a stepwise approach to the management of this high risk tumor. Collectively, all these advances bring us a stepforward in tailoring the management of superficial bladder tumor patients by minimizing the burden of unnecessary investigations and treatments while safely identifying those patients with high risk disease that deserve more aggressive treatment. PMID- 12121580 TI - The changing face of prostate cancer in British Columbia 1988-2000. AB - OBJECTIVE: To evaluate changes of prostate cancer incidence, referrals, stage, treatment and outcomes delivered in British Columbia since the 1980's. MATERIALS AND METHODS: Examination of the BC Provincial Tumour Registry, BC Cancer Agency (BCCA) and BC Medical Services Plan databases. RESULTS: The number of incident cases increased linearly from 1980 through 1990. Between 1991 and 1995 a harvesting effect was seen due to unofficial PSA screening, balanced by a post harvest effect between 1995 and 1998. Since 1999 the incidence has resumed the linear trend extrapolated from the 1980's. The age-standardised incidence rate has recently risen in younger (<65yrs) men. The incidence of metastatic cancer has dropped from 14% of cases referred to the BCCA in 1988 to 3.5% in 2000. A steady proportionate increase in T1 and T2 referrals has occurred since 1988. PSA levels at referral are lower (mean PSA 10 nmol/L in 2000 versus 15 nmol/L in 1990. Gleason scores are higher, likely reflecting changes of interpretation of pathological grade. The number of men receiving any curative therapy has increased from 43% in 1990 to 53% by 1999, and the proportion treated with surgery has increased from 30% in 1990 to 50% by 2000. Mortality rates have been falling since 1991, and BC has the lowest mortality rate in Canada. CONCLUSIONS: Predictions of incidence have been beset by unanticipated external factors, and have underestimated actual incidence. Stage migration towards better prognosis tumours occurring in younger men has led to the increased use of surgery and brachytherapy and decreased use of external radiation. PMID- 12121581 TI - A randomized trial to evaluate effectiveness and cost effectiveness of naturopathic cranberry products as prophylaxis against urinary tract infection in women. AB - PURPOSE: To determine, from a societal perspective, the effectiveness and cost effectiveness of concentrated cranberry tablets, versus cranberry juice, versus placebo used as prophylaxis against lower urinary tract infection (UTI) in adult women. MATERIALS AND METHODS: One hundred fifty sexually active women aged 21 through 72 years were randomized for one year to one of three groups of prophylaxis: placebo juice + placebo tablets versus placebo juice + cranberry tablets, versus cranberry juice + placebo tablets. Tablets were taken twice daily, juice 250 ml three times daily. Outcome measures were: (1) a >50% decrease in symptomatic UTI's per year (symptoms + >or= 100 000 single organisms/ml) and (2) a >50% decrease in annual antibiotic consumption. Cost effectiveness was calculated as dollar cost per urinary tract infection prevented. Stochastic tree decision analytic modeling was used to identify specific clinical scenarios for cost savings. RESULTS: Both cranberry juice and cranberry tablets statistically significantly decreased the number of patients experiencing at least 1 symptomatic UTI/year (to 20% and 18% respectively) compared with placebo (to 32%) (p<0.05). The mean annual cost of prophylaxis was $624 and $1400 for cranberry tablets and juice respectively. Cost savings were greatest when patients experienced >2 symptomatic UTI's per year (assuming 3 days antibiotic coverage) and had >2 days of missed work or required protective undergarments for urgency incontinence. Total antibiotic consumption was less annually in both treatment groups compared with placebo. Cost effectiveness ratios demonstrated cranberry tablets were twice as cost effective as organic juice for prevention. CONCLUSIONS: Cranberry tablets provided the most cost-effective prevention for UTI. PMID- 12121582 TI - Adrenal tumors associated with inadequately treated congenital adrenal hyperplasia. AB - We describe a case of salt-losing congenital adrenal hyperplasia due to 21 hydroxylase deficiency complicated by a right adrenal adenoma. The development of adrenal adenoma or carcinoma in-patients with congenital adrenal hyperplasia (CAH) is rare; the etiology is not clear but is thought to be related to inadequate glucocorticoid therapy. Tumor formation is postulated to be a consequence of ACTH hypersecretion, which results from the lack of glucocorticoid synthesis. Our patient underwent clitorectomy and multiple constructive procedures as a newborn baby; she was managed with hormone replacement for many years. However while she took adequate mineralocortocoid dosage, she chronically tended to take inadequate doses of glucocorticoid seeking to increase her muscle ability. She developed a 6.5 cm adrenal tumor. She was managed by a hand-assisted laparoscopic radical adrenalectomy. The tumor was histologically consistent with adrenal adenoma. The importance of compliance with her medications was emphasized. PMID- 12121583 TI - Simultaneous renal cell carcinoma of the kidney and transitional cell carcinoma of the bladder. AB - Herein we report a rare combination of two urological tumors simultaneously: renal cell carcinoma of the kidney and transitional cell carcinoma of the bladder. A 67 year old male presented with hematuria. He was diagnosed with left renal cell carcinoma and transitional cell carcinoma of the bladder. Subsequently this patient underwent left nephrectomy (palliative) to control the pain and transurethral resection of bladder tumor. The clinical picture, radiographic and histological finding are presented, as well as the treatment offered. PMID- 12121584 TI - Testicular block using intra-testicular lidocaine: a new anaesthetic technique for percutaneous testis biopsy. AB - PURPOSE: We describe a simple technique to deliver local anaesthetic for percutaneous testis biopsies. MATERIALS AND METHODS: With the testis held firmly, a 25 gage needle is used to inject lidocaine, without epinephrine, into the skin and dartos superficial to the testis, then the needle is advanced through the tunica albuginea and 0.5 mL to 1.0 mL of lidocaine is injected directly into the testis. The testis becomes slightly more turgid with the injection. A percutaneous biopsy is then immediately performed. RESULTS: Intra-testicular lidocaine, (without need of a cord block or any sedation) was used on a total of 45 consecutive patients having percutaneous testicular biopsies. Procedure time was short (averages less than 5 minutes) and anaesthesia was profound. There was no change in the number of seminiferous tubules for evaluation compared to biopsies on men using a cord block. Only 1/45 men had a post-procedure testicular hematoma (this resolved in 4 weeks). CONCLUSIONS: Intra-testicular lidocaine appears to be a simple, rapid and safe method to provide anaesthesia for a percutaneous testis biopsy. PMID- 12121587 TI - Expectant management with selective delayed intervention for favorable-risk prostate cancer. AB - The optimal management of clinically localized prostate cancer remains unresolved. Management options range from a conservative approach to definitive treatment. While evidence suggests that expectant management yields similar 10 year survival rates and quality-adjusted life years compared to definitive treatment, this approach alone will deprive some patients with potentially curable disease of the opportunity for curative therapy. Effective management of localized prostate cancer requires differentiation between patients with biologically aggressive disease, in whom curative therapy is strongly warranted, and those with indolent malignancy, in whom conservative management would be equally efficacious. A comparison of surveillance studies in the literature yields a striking similarity: every series contains a substantial subset of long term survivors, particularly in patients with favorable clinical parameters. We describe a prospective clinical study to evaluate a novel approach in which the decision between definitive therapy and conservative management is determined by the rate of prostate-specific antigen (PSA) increase or the development of early, rapid clinical and/or histologic progression. We enrolled 250 patients followed with active surveillance with selective delayed intervention. Patients were followed with active surveillance until they met criteria defining significant PSA, clinical, or histologic progression. At a median follow-up of 42 months, 60 patients came off observation, while 140 have remained on observation. We conclude that an approach of active surveillance with selective intervention for patients with rapid biochemical or clinical progression is feasible, and that PSA doubling time appears useful in guiding intervention in patients managed initially with expectant management. A policy of close monitoring with selective intervention for the 15% who progress rapidly is appealing, and is currently being investigated in several clinical trials. PMID- 12121588 TI - Role of radical prostatectomy in high-risk prostate cancer. AB - Many methods exist to define high-risk prostate cancer. These include clinical stage, serum PSA, and pathological features such as Gleason score and the number of positive biopsies. Partin tables are widely used to stratify patients according to risk of adverse pathological features at surgery, and to identify those more likely to remain free of recurrent disease following surgery. The priority in most patients with localized prostate cancer remains the selection of a treatment that will provide them with the best chance for cure. While treatment related morbidity is an important issue, we believe that side effects of surgery or radiation therapy are not increased in patients with high-risk cancer. Results from a small number of population studies indicate a highly significant improvement in disease-specific survival for radical prostatectomy compared to radiotherapy, and it appears that this difference may become more pronounced as the grade of the cancer increases. While acknowledging the need for adjuvant radiotherapy and/or hormonal therapy, we suggest that radical prostatectomy may offer a better primary treatment option for patients with high-grade cancer. However, urologists must be prepared for higher failure rates when performing this surgery in patients with high-risk disease compared to those with low-risk disease. PMID- 12121589 TI - Laparoscopic prostatectomy: here to stay. AB - In 2002, open retropubic radical prostatectomy remains the standard of care for localized carcinoma of the prostate. However, the laparoscopic approach offers several appealing advantages and is being practiced more widely. The more commonly performed technique is the transperitoneal "Montsouris" technique with defined steps which are described. There are a number of pointers that are learned with experience. The most remarkable aspect of laparoscopic prostatectomy is the relatively pain-free and trouble-free postoperative course. Patients can be discharged within 2 days, have very little analgesic requirement, and feel well faster. The catheter can be removed in 4 to 7 days. Cancer control appears comparable to that of open surgery, although results are relatively preliminary and are stated in terms of biochemical freedom from disease. The functional results continue to be reported, but in an experienced surgeon's hands, continence appears to recover more quickly and appears to be similar to that achieved after open surgery. Potency can be preserved with nerve sparing. Laparoscopic radical prostatectomy is a procedure that will be more widely practiced in North America as several of the technological limitations are overcome and as surgeons become more experienced with laparoscopic surgery. PMID- 12121590 TI - Risk stratification in clinically localized prostate cancer. AB - Clinical outcomes in patients with localized prostate cancers are heterogeneous. In recent years, analyses of large datasets from multiple centres have yielded a better understanding of how to measure risk in localized prostate cancer. Regardless of whether patients are treated with prostatectomy, radiotherapy, brachytherapy, or expectant management, three factors appear correlated with clinical outcome: biopsy Gleason score, clinical T stage, and serum prostate specific antigen (PSA). Partin Tables, derived from these parameters and recently updated and refined, may be used to estimate the risk of metastasis and to assess certain aspects of surgical management in clinically localized disease. Partin tables, however, are limited by the fact that pathologic stage does not always predict clinical outcome. Nomograms that employ serum PSA, biopsy Gleason score, and clinical T-stage have been developed with the aim of predicting clinical recurrence after radical prostatectomy or radiation therapy. Three risk categories for clinically localized prostate cancer have recently been developed by the Canadian Genito-Urinary Radiation Oncologists Consensus Conference, which group cases according to serum PSA, T-stage, and biopsy Gleason score. Additional factors have been assessed in the hopes of improving the prediction of outcome in clinically localized disease, but none of these has consistently been demonstrated to add independent value to the principal parameters of serum PSA, T stage, and Gleason score. Virtually all predictive nomograms, algorithms, and tables incorporate a combination of these three parameters. While these tools may be useful in prognosticating an individual case, several limitations preclude their widespread use. The greatest benefit to date of risk stratification is its use in comparing outcomes of series of patients treated with various modalities, and in clinical trial design and analysis. PMID- 12121591 TI - The role of chemotherapy in advanced prostate cancer. AB - The development of hormone resistance is an unfortunate final common pathway in most patients with advanced prostate cancer, resulting in a narrowing of therapeutic options for the clinician, and limited median survival of 10-12 months for the patient. While cytotoxic chemotherapy has been utilized for many years, its efficacy has been disappointing. Quality of life assessments are increasingly used in assessing response in hormone-resistant prostate cancer (HRPC), and PSA has emerged as an important surrogate marker of response in both local and advanced disease. Estramustine and the taxanes have been investigated, as monotherapy and in combination, in the treatment of HRPC in phase 2 and 3 clinical trials, a number of which are ongoing. Substantial advances in the management of HRPC over the past decade have led to renewed optimism that improvement in survival can be achieved, and support the belief that chemotherapy plays a role in this pursuit. In tandem with the development of new agents, refined means of assessing response have been developed, and represent a key component of new research strategies in HRPC. PMID- 12121592 TI - Prostate cancer: risk categories and role of hormones and radiotherapy. AB - The Genito-Urinary Radiation Oncologists of Canada (GUROC) have produced a consensus statement on radiotherapy in prostate cancer. This paper summarizes the consensus statement with regard to risk grouping and the role of hormones and radiotherapy. Survival is the most important outcome in the assessment of patients treated with radiotherapy. Other outcomes of interest include disease free survival, metastatic-free survival, local control, biochemical measures, toxicity, efficacy, and quality of life. Risk groupings based on prognostic data are increasingly used in the management of prostate cancer. These groupings have been correlated to prognosis in several studies, and are helpful in identifying optimum treatments, and as a research tool to evaluate new treatments and modalities. Adjuvant hormone treatment with radiotherapy has been demonstrated in two studies (Bolla and RTOG 85-31) to be beneficial in patients with locally advanced prostate cancer. Neoadjuvant hormone treatment in patients with low- and intermediate-risk disease is being evaluated in a RTOG study and its utility in these patients will be clarified when the study results are available. The GUROC consensus statement recommends that patients with high-risk non-metastatic prostate cancer be treated with adjuvant hormone therapy for 2-3 years. Part of this hormone treatment may be administered in a neoadjuvant fashion. Adjuvant hormone treatment should not be routinely used in low- and intermediate-risk prostate cancer. Neoadjuvant hormone treatment is recommended prior to radiotherapy in patients with bulky tumors. The results of ongoing research will further clarify the use of hormone treatment with radiotherapy. PMID- 12121593 TI - Controversies in the management of localized prostate cancer: consensus development by Canadian urologists. AB - This consensus statement emerged from the 2nd Canadian Uro-Oncology Congress, held January 16-20, 2002. The attendees at this meeting comprised approximately 125 urologists from across Canada, representing both community and academic perspectives. The group heard presentations by experts in the field addressing the spectrum of controversies in prostate cancer. After each session, the meeting broke into workshop sessions where attendees discussed the controversies raised by the speakers. Present in each workshop was a reporter who summarized the consensus of the group. These summaries were collected and integrated into a set of questions. At the end of the congress, the attendees voted on the questions and controversies identified through the workshop process. PMID- 12121594 TI - Increasing incidence of nontyphi Salmonella bacteremia among children living in southern Israel. AB - OBJECTIVES: To determine if the epidemiology of Salmonella gastroenteritis and childhood bacteremia among the two ethnic populations (Jews and Bedouins) living in southern Israel has changed in recent years. METHODS: Retrospective review of laboratory records and medical charts of patients from whom non-typhi salmonellae were isolated from stool and blood cultures in the 1990-1995 period. RESULTS: The overall incidence of enteric Salmonella infections was 123.5 per 100 000 inhabitants and remained stable during the study period. The incidence of bacteremia among children younger than 4 years increased from 9.3 per 100 000 in the 1990-1992 period to 26.8 per 100 000 in the 1993-1995 period (P 0.01). This increment was especially caused by Salmonella virchow and S. enteritidis, which were also isolated with increasing frequency from stool cultures. The Bedouin population was underrepresented among stool isolates, whereas its representation among blood isolates closely resembled the fraction of this ethnic group in the overall population of the area. CONCLUSIONS: The incidence of enteric salmonellosis has remained stable at a high endemic level in recent years in southern Israel. The incidence of children's bacteremia has experienced a significant increase, associated with S. virchow and S. enteritidis. Differences in the utilization of medical services may explain differences in the epidemiology of Salmonella infections found in the two resident ethnic groups. PMID- 12121595 TI - Foodborne disease in our global village: a multinational investigation of an outbreak of Salmonella serotype Enteritidis phage type 4 infection in Puerto Vallarta, Mexico. AB - OBJECTIVES: In late 1996, a multinational investigation was launched following an outbreak of diarrheal illness that caused the disruption of an international scientific conference at a first-class hotel in Puerto Vallarta, Mexico. METHODS: A questionnaire was mailed to all American and to selected international attendees. Additional copies of the questionnaire were provided for any family members who may have attended the conference. A case was defined as an illness with three or more loose stools during a 24-h period in a conference attendee or accompanying family member, with illness lasting 2 or more days and onset between 6 and 9 November 1996. RESULTS: Questionnaires were returned by 81% (232/288) of American attendees, 47% (18/38) of selected international attendees, and 25 family members; 30% (83/275) of respondents met the case definition. Ill persons resided in at least seven countries. Salmonella serotype Enteritidis phage type 4 was isolated from stool specimens from patients residing in Canada, the UK, and the USA. Attending a hotel banquet on 6 November was associated with illness; 42% (82/194) of banquet attendees became ill versus 3% (1/37) of non-attendees (relative risk (RR)515.6, 95% confidence interval (CI)52.3-108.9). The only banquet food item associated with illness was chili rellenos; 53% (58/109) of persons who ate chili rellenos were ill versus 22% (12/55) of those who did not (RR52.4, 95% CI51.4-4.1). Chili rellenos ingredients included shelled eggs and cheese; Salmonella was isolated from the leftover cheese but the isolate was not serotyped. CONCLUSIONS: Salmonella may be a cause of traveler's diarrhea and can result in outbreaks even among travelers who follow routine precautions (i.e. staying in a first-class hotel and eating hot foods). International collaboration in investigating similar outbreaks, including sharing subtyping results, will be necessary for long-term prevention. Global Salm-Surv, an international network of Salmonella reference laboratories coordinated by the World Health Organization, may facilitate such collaboration. PMID- 12121596 TI - The decision-making process in antibacterial treatment of pediatric upper respiratory infections: a national prospective office-based observational study. AB - BACKGROUND: The identification of patient management practices and the sources of medical information is crucial for rationalizing the treatment of respiratory tract infections, whose high incidence, especially in children, makes them one of the maior areas of unnecessary health expenditure. MATERIALS AND METHODS: This national prospective study was designed to investigate the diagnostic and prescribing habits of 100 office-based pediatricians managing upper respiratory tract infections in 1111 pediatric patients (604 males, mean age 6.7962.77 years; 507 females, mean age 6.7362.8 years) sequentially enrolled when an antibiotic treatment was deemed necessary. RESULTS: The most frequently diagnosed diseases were acute tonsillopharyngitis (56.2%) and acute otitis media (18.1%). Penicillins were prescribed in 34.3% of the cases, cephalosporins in 38.1%, and macrolides in 26.1%: oral drugs accounted for 92.2% of the prescriptions. The treatments were administered once or twice daily in 75.8% of the patients, and prescribed for 8 days in more than 80%; 76.7% also received supportive or symptomatic treatment (antipyretics, corticosteroids, cough suppressants and non steroidal anti-inflammatory drugs). Laboratory or radiologic investigations were rarely requested. The main sources of medical information indicated by the participating pediatricians were pharmaceutical companies (35.6%) and meeting or congress reports (27.3%). CONCLUSIONS: The results indicate that more active education is still needed to improve the decision-making processes of office based pediatricians. PMID- 12121597 TI - Drugs in the parallel market for the treatment of urethral discharge in Dakar: epidemiologic investigation and physicochemical tests. AB - OBJECTIVE: Sexually transmitted diseases (STDs) constitute a major public health concern in developing countries. Their interest lies mainly in their diagnosis and their early treatment. Owing to lack of health education and poor living conditions inherent in underdevelopment, self-medication is common practice in these Third World countries. Therefore, the illegal sale of drugs is an important phenomenon in Africa and Asia. METHODS: An investigation, with a view to evaluating the importance of drug sales in the parallel market for the treatment of urethral discharge in Dakar, was carried out in 50 different locations in the working-class districts of the capital from 13 February to 6 March 1997. These drugs, obtained from vendors in the illegal market, were tested and analyzed using the standard physicochemical methods. RESULTS: The most frequently proposed drugs to treat male urethritis are: ampicillin 250-mg capsules (44%); oxytetracyline 250-mg capsules (24%); and cotrimoxazole 450-mg tablets (12%). In most cases (88%), these drugs were sold unpackaged; 12% were sold in blisters. Furthermore, in 90% of cases, the expiry date was not indicated. The dosage and duration of treatment were correct in only 6% of cases. The physicochemical analysis was based on the external appearance, the identification, and the dosage of the active principle. Of these drugs offered by street vendors, 53.1% had an unusual appearance; they were mainly ampicillin 250 mg (21 of 22 samples) and oxytetracycline 250 mg (6 of 12 samples). Furthermore, all active principles were identified as positive, with the exception of ampicillin 250 mg, for which only one sample of the 22 was positive; the others appeared to be flour, with no trace of the active principle. CONCLUSION: Given the extent of these illegal sales of drug and their harmful consequences for the health of the population, adequate measures should be taken to eradicate this blight. For this mission to succeed, public authorities, health professionals and populations should combine their efforts. PMID- 12121598 TI - Antibody response to Haemophilus influenzae type b tetanus conjugate vaccine with two doses given at 3 and 5 months of age. AB - BACKGROUND: In developed countries, the use of Hib conjugate vaccines has led to the near disappearance of invasive Hib disease, but costs have limited its use in developing countries. In order to identify more economical vaccination schedules, we carried out a trial to evaluate the immunogenicity of an alternative two-dose PRP-T regimen, based on a previous report in which carrier priming could be obtained with prior diphtheria-tetanus-pertussis (DTP) vaccination. METHODS: Healthy infants were enrolled to receive the PRP-T given at 3 and 5 months of age, with DTP vaccination given at 2, 4 and 6 months of age. Serum specimens were obtained at 3, 6 and 15 months of age. IgG anti-Hib titer determination was performed using enzyme-linked immunosorbent assay to evaluate serologic response and its duration. RESULTS: One-hundred and seventeen infants were enrolled. The geometric mean titer (GMT) of antibody to PRP was low in the pre-immunization samples (0.13 mg/mL), achieving high values after two doses of PRP-T (27.42 mg/mL), with all titers over 1 mg/mL; the GMT at 15 months was 5.45 mg/mL; 94.6% of infants had serologic responses after the two doses of vaccination, with average intervals of 27 and 22 days between DTP and PRP-T first-to-first and second-to-second administrations, respectively. However, these intervals were 11 and 3 days for infants who did not have serologic responses (P50.0013 and 0.0030, respectively). CONCLUSIONS: These results indicate that two doses of PRP-T can induce high antibody titers using the proposed schedule; moreover, the GMT assessed at 15 months of age was also protective. The enhanced immune response observed in the study could be explained by the previous administration of the DTP vaccine, since the longer the interval between DTP and PRP-T, the better the response to Hib vaccine. The PRP-T vaccine given at 3 and 5 months of age may be an economical alternative to the current proposed schedule, which could make the introduction of Hib vaccination in developing countries more feasible, considering the relatively high cost of this vaccine. PMID- 12121599 TI - Effect of age on outcome of secondary dengue 2 infections. AB - OBJECTIVE: Dengue hemorrhagic fever/dengue shock syndrome (DHF/DSS) is a growing global health problem. It is not known how age affects the outcome of secondary dengue infections. In an island setting, a large DHF/DSS outbreak in Cuba occurred in 1981. Involved were individuals, 3-40 year old, whose only lifetime dengue exposure was to DEN-1 in 1977 and DEN-2 in 1981. In this report we calculate age-specific DHF/DSS hospitalization and death rates based on secondary DEN 2 infections. METHODS: Published and unpublished hospital and seroepidemiologic data from the 1981 DHF/DSS outbreak were used for the analysis. RESULTS: Children, aged 3 and 4 years, with secondary DEN-2 infections were found to have a high death rate (25.4/10 000 secondary DEN-2 infections). The death rate fell with increasing age, being 15.9-fold lower in the 10-14-year age group. The death rate for children aged 3-14 years was 14.5-fold higher than in young adults aged 15-39 years. The death rate rose somewhat in adults aged 50 years and older. DHF/DSS hospitalization rates showed the same trend as death rates. CONCLUSIONS: Age is an important variable in the outcome of secondary DEN-2 infections. DHF/DSS case fatality and hospitalization rates are highest in young infants and the elderly. The risk that a child will die during a secondary DEN-2 infection is nearly 15-fold higher than the risk in adults. PMID- 12121600 TI - Long-term memory cellular immune response to dengue virus after a natural primary infection. AB - OBJECTIVES: This study was conducted to examine the memory T-cell response to dengue virus 20 years after a primary infection. We took advantage of the exceptional epidemiologic situation in Cuba, where the population initially suffered two large successive epidemics due to dengue virus 1 and 2 respectively over a 4-year period. Thereafter, no dengue virus circulation was subsequently observed, except for the Santiago de Cuba municipality. DESIGN: T-cell response was evaluated in peripheral blood mononuclear cells (PBMCs) from 20 individuals with history of a primary infection by dengue virus 1 or 2. Methods previously shown to induce lymphoproliferation of CD4+ memory T-cell subpopulations were used. We evaluated the proliferative responses generated in those PBMCs after stimulation with dengue virus 1, 2, 3 and 4 antigens in a serotype-specific and serotype-crossreactive way. RESULTS: Serotype-specific and serotype-crossreactive lymphoproliferative responses in all PBMCs donated by dengue immune donors were observed. The serotype-crossreactive response for dengue 2 was stronger than for the rest of the serotypes. CONCLUSIONS: This is the first report of cellular memory lymphocyte response specific for dengue virus detected 20 years after a primary infection by dengue. PMID- 12121601 TI - Immunogenicity and reactogenicity of a combined hepatitis A and B vaccine in healthy Chilean subjects. AB - OBJECTIVES: A combination vaccine against hepatitis A and B provides the opportunity for simultaneous protection against both diseases with a single vaccine. This clinical study investigated the reactogenicity and immunogenicity of a combined hepatitis A and B vaccine (Twinrix, GlaxoSmithKline Biologicals, Rixensart, Belgium) in healthy Chilean adults between 18 and 40 years of age. METHODS: In total, 345 healthy, seronegative health care workers were enrolled and randomized to three groups who received one of three lots of Twinrix on a 0-, 1- and 6-month schedules. Blood samples were screened 1 month after each dose for anti-HAV and anti-HBs antibodies. Reactogenicity after each dose was assessed using diary cards. RESULTS: The nature and incidence of symptoms were similar to those reported for other Twinrix studies. Very few symptoms were scored as severe. Upon completion of the vaccination, all subjects had anti-HAV antibodies with titers $6000 mIU/mL, and all but one were protected against hepatitis B, with titers $4000 mIU/mL. CONCLUSIONS: We have demonstrated the high immunogenicity and tolerance of the combined hepatitis A and B vaccine. Combined vaccination has the advantage of offering dual protection with a reduction in the number of injections needed, lower associated costs, and a positive impact on compliance. PMID- 12121603 TI - Non-smoker with pulmonary nodules. PMID- 12121602 TI - Clinical and epidemiologic trends in HIV/AIDS patients in a hospital setting of Yaounde, Cameroon: a 6-year perspective. AB - OBJECTIVES: In order to appreciate the impact of the HIV/AID pandemic in Yaound , Cameroon, an evaluation of the clinical and epidemiologic trends in HIV/AIDS patients was undertaken in a hospital setting. METHODS: A rapid assessment method was used to collect data. Patient record examination, interviews and direct observation were employed. RESULTS: Of 875 cases studied in the hospital during a 6-year period, 43.7% were males and 56.3% females. A total of 5.4% of all the cases were seen in 1993 compared to 30.5% in 1998. The number of admissions per patient ranged from 0 to 4, with a median duration of admission of 14 days (range 0-343 days). The 25-44-year age group was mostly affected (63.4% cases) and 10.1% were in the 0-14-year age group. About 27% of cases died in hospital, mainly between 1996 and 1997. The predominant clinical manifestations included persistent fever and diarrhea, excessive weight loss, chronic cough and profound asthenia. Opportunistic infections and cancers also formed part of the picture. CONCLUSIONS: The increasing clinical and epidemiologic trends of the HIV/AIDS pandemic within the hospital show the devastation and socio-economic impact, especially on the Cameroonian youth and women. Intense public health measures must be put in place to educate and cater for the vulnerable groups in society. PMID- 12121604 TI - The promise of osseointegration: two decades later. PMID- 12121605 TI - Effects of a mechanical barrier on the integration of cortical onlay bone grafts placed simultaneously with endosseous implant. AB - BACKGROUND: Previous experimental studies on onlay bone graft integration have shown either advantages or disadvantages to the use of mechanical barriers. This indicates that the role played by the biologic properties of transplanted bone and membrane in graft revascularization and bone remodeling has not yet been established. The outcomes regarding osseointegration of titanium dental implants applied in such a condition are still contradictory. PURPOSE: The rabbit's radius model that is grafted onto the mandibular lower border and covered by membrane can reproduce a challenging experimental situation to preliminarily study the factors involved in osseointegration under deprived blood vessels source. MATERIALS AND METHODS: Fourteen New Zealand White rabbits had a 2.5-cm segment of the right radius osteoectomized and fixed onto the right mandibular lower border using titanium screws. Two screw-shaped titanium implants (2.5 mm wide yen 2.5 mm long) were installed 7 mm apart in the mid length of the grafted bone. In experimental sites, the graft with the implants and graft-host bone junction were covered by expanded polytetrafluoroethylene (e-PTFE) membrane; control sites were left uncovered. Eight animals from the experimental group and six animals from the control group were sacrificed at 6 and 24 weeks after surgery. Ground sections obtained from en bloc tissues containing graft, implants, and recipient bone were subjected to histologic evaluation and histomorphometric analysis (area occupied by the graft and bone-to-implant contact). RESULTS: The graft showed significantly more resorption after 24 weeks than at 6 weeks (p < or =.05) irrespective of the treatment (with or without membrane), although the amount of new bone was greater at 24 weeks in sites where a membrane was covering the graft. Compared with 6 weeks postoperatively, the bone-to-implant contact was considerably improved at 24 weeks (p < or =.05), and the membrane seemed beneficial for implant osseointegration when compared with unprotected sites (p .05). As a result of graft resorption, the amount of soft tissue was considerably expanded in sites beneath membrane, accompanied by a sustained process of trabecular bone deposition close to the barrier. CONCLUSIONS: Cortical onlay grafts covered by membrane demonstrated delayed remodeling, probably as a consequence of a hindered process of graft revascularization. Grafts covered by membrane might rely on previous host bone resorption both to become revascularized and to remodel. The findings that the membrane-protected grafts were most resorbed at 24 weeks might be attributable to better implant osseointegration, because the fixtures were exposed to greater mechanical stimulation in these sites. key words: bone regeneration, implant osseointegration, onlay bone-graft PMID- 12121606 TI - The Marius implant bridge: surgical and prosthetic rehabilitation for the completely edentulous upper jaw with moderate to severe resorption: a 5-year retrospective clinical study. AB - BACKGROUND: Patients seeking replacement of their upper denture with an implant supported restoration are most interested in a fixed restoration. Accompanying the loss of supporting alveolar structure due to resorption is the necessity for lip support, often provided by a denture flange. Attempts to provide a fixed restoration can result in compromises to oral hygiene based on designs with ridge laps. An alternative has been an overdenture prosthesis, which provides lip support but has extensions on to the palate and considerations of patient acceptance. The Marius bridge was developed as a fixed bridge alternative offering lip support that is removable by the patient for hygiene purposes, with no palatal extension beyond normal crown-alveolar contours. PURPOSE: Implant supported restorative treatment of completely edentulous upper jaws, as an alternative to a complete denture, is frequently an elective preference, and it requires significant patient acceptance beyond the functional improvement of chewing. Patients with moderate to severe bone resorption and thin ridges present additional challenges for adequate bone volume and soft-tissue contours. The purpose of this investigation was to develop a surgical and prosthetic implant treatment protocol for completely edentulous maxillae in which optimal lip support and phonetics is achieved in combination with substantial implant anchorage without bone grafting. MATERIALS AND METHODS: The Marius bridge is a complete-arch, double-structure prosthesis for maxillae that is removable by the patient for oral hygiene. The first 45 consecutive patients treated by one person (YF) in one center with this concept are reported, with 245 implants followed for up to 5 years after prostheses connection. RESULTS: The cumulative fixture survival rate for this 5-year retrospective clinical study was 97%. Five fixtures failed before loading, in five different patients, and two fixtures in the same patient failed at the 3-year follow-up visit. None of the bridges failed, giving a prostheses survival rate of 100%. The complications were few and mainly prosthetic: nine incidences of attachment component complications, one mesobar fracture, and three reports of gingivitis. All complications were solved or repaired immediately, with minimal or no interruption of prostheses use. CONCLUSIONS: Satisfactory medium-term results of survival and patient satisfaction show that the Marius bridge can be recommended for implant dentistry. The technique may reduce the need for grafting, because it allows for longer implants to be placed with improved bone anchorage and prostheses support. key words: clinical follow-up, complete arch, double structure, esthetics, maxilla, phonetics, tilted implants PMID- 12121607 TI - Bone reactions to oxidized titanium implants with electrochemical anion sulphuric acid and phosphoric acid incorporation. AB - BACKGROUND: The importance of the surface properties of implants for a successful osseointegration has been emphasized. It is generally known that bone response to implant surfaces is considerably related to the various surface properties. PURPOSE: The purpose of this study was to investigate bone tissue reactions to multifactorial biocompatibility of the surface oxide of electrochemically oxidized titanium implants. The ultimate objective was to improve surface quality, resulting in enhancement of clinical outcomes of osseointegrated implants. MATERIALS AND METHODS: Three different surface types of commercially pure titanium (c.p. Ti) implants were prepared. Turned implants were used for controls and test implants were prepared by the micro arc oxidation (MAO) method, either in sulphuric acid (S implants) or in phosphoric acid (P implants). Implants were inserted in the femur and tibia of 10 mature New Zealand White rabbits. The bone response was evaluated by biomechanical tests, histology, and histomorphometry. The follow-up time was 6 weeks. RESULTS: The mean peak values of the removal torque showed significant differences between control and test S implants (p =.022) but showed no significant differences between control and test P implants (p =.195) or between test S and test P implants (p =.457). In addition, the histomorphometric comparisons of the bone-to-metal contact around entire implants demonstrated 186% increase in S implants (p =.028) and 232% increase in P implants (p =.028) compared with the paired control groups. Quantification of the bone area in the threads did not show any significant differences. CONCLUSIONS: The present results suggest that the primary mode of action in strong bone response to S implants is mechanical interlocking, and to P implants, it is biochemical interaction. It is possible that the phosphate groups in the titanium oxide of P implants provide potential chemical bonding sites for calcium ions and hydroxyapatite of the bone matrix during biologic mineralization. key words: bone responses, histomorphometry, oxidized implants, removal torque test, surface oxide properties PMID- 12121608 TI - Precision surgical template for implant placement: a new systematic approach. AB - The importance of a precise surgical template for implant placement cannot be overstated. The radiographic template carries both clinical and radiographic information for the planning of fixture angulation and location. This article describes a systematic approach to the fabrication of a dual-purpose radiographic surgical template. The simple steps result in the accurate transfer of radiographic information to the surgical template with no need for complex equipment or maneuvers. key words: dental implants, implant placement, radiographic template, surgical template PMID- 12121609 TI - Simplified methods of implant treatment in the edentulous lower jaw. A controlled prospective study. Part I: one-stage versus two-stage surgery. AB - BACKGROUND: The original protocol for Branemark System implants in the mandible was a two-stage procedure with 3 months healing time. With five or six implants and a cast framework of gold, the treatment is rather expensive, and simplified methods would be desirable. PURPOSE: The goal of this controlled serial study was to investigate the outcome of a simplified procedure with one-stage surgery, four Branemark implants, shortened healing time, and a new titanium-acrylic fixed full prosthesis. MATERIALS AND METHODS: Eighty-two patients were treated in three different groups at two specialist centers. All patients were provided with four implants, loaded with a Procera All-in-One bridge (Nobel Biocare, Gothenburg, Sweden) after 12 weeks. In group A (n = 30), one-stage surgery was combined with two-piece implants. In group B (n = 30), the control group, two-stage surgery and two-piece implants were used. In group C (n = 22), one-stage surgery was combined with one-piece implants. Marginal bone level was rated from radiographs at implant insertion, at baseline, and after 1 year. RESULTS: The survival rate after 1 year for group A was 93.3%; group B, 97.5%; and group C, 93.2%. The differences were not statistically significant. Between fixture insertion and baseline, the average bone loss for group A was 1.2 mm; group B, 1.3 mm; and group C, 1.3 mm. No complications in the form of bridge loosening or acrylic fractures were recorded during the first year. CONCLUSIONS: The survival rates and the marginal bone changes did not differ significantly between the one-stage groups and the control group. The survival rate and the marginal bone changes were similar for one-piece and two-piece implants. Four implants were sufficient to support full fixed prostheses in the mandibles. The Procera All-in-One bridges proved to be of high quality, and no complications were experienced. key words: endosseous implants, nonsubmerged implants, one-piece implants, prospective clinical study, submerged implants PMID- 12121610 TI - Clinical experience of CNC-milled titanium frameworks supported by implants in the edentulous jaw: a 3-year interim report. AB - BACKGROUND: The use of computer numeric controlled (CNC)-milled titanium frameworks is a new technique for framework fabrication, and few clinical reports have been made on this treatment modality. PURPOSE: The goal of this study was to report the clinical performance of implant-supported prostheses with CNC-milled titanium frameworks in the edentulous jaw and to compare the results with prostheses provided with conventional cast frameworks during the first 3 years of function. MATERIALS AND METHODS: A consecutive group of 126 edentulous patients were provided by random distribution with 67 prostheses with CNC-milled titanium frameworks in 23 upper and 44 lower jaws and 62 conventional prostheses with gold alloy castings in 31 upper and 31 lower jaws. Radiographic 1-year data and clinical 3-year data were collected for both the titanium and control group. RESULTS: One prosthesis was lost in each group owing to loss of implants, and the overall 3-year prosthesis cumulative survival rate was 98.2% for both groups. Patients with smoking habits experienced significantly more implant failures than nonsmokers (p =.006). Few problems were observed. No metal fractures were seen in the test group, whereas two frameworks and one abutment screw fractured in the control group. Resin veneer fractures were the most common complication, with a slightly higher incidence observed in the control group. CONCLUSIONS: Computer numeric controlled-milled titanium frameworks can be used as an alternative to conventional castings in the edentulous jaw, presenting clinical performance similar to that of conventional cast frameworks during the first 3 years of function. key words: computer numeric controlled, implant supported, prostheses, titanium PMID- 12121611 TI - Histologic evaluation of a human immediately loaded titanium implant with a porous anodized surface. AB - BACKGROUND: Several researchers have demonstrated, in the past decade, in clinical and histologic studies, that immediate loading can be successfully used in implant dentistry. Many factors are thought to be of importance in obtaining mineralized tissues at the interface. One of these factors is the implant surface characteristics. Recently, an implant with a new porous anodized surface has been introduced in the market. PURPOSE: The goal of this study was to histologically evaluate a clinically retrieved immediately loaded implant with a porous anodized surface. MATERIALS AND METHODS: After a 6-month loading period, an immediately loaded implant with a porous anodized surface that had been inserted in the posterior maxilla was retrieved. RESULTS: Histology showed that mineralized tissue was present at the interface and the bone-implant contact percentage was about 60%. No gaps or fibrous tissue or inflammatory infiltrate were present at the interface. CONCLUSIONS: Results show that immediate loading of dental implants has no untoward effects on the formation of mineralized tissues at the interface. Immediate loading is a possible alternative in implant dentistry. key words: dental implants, histology, immediate loading, oxide coating, porous surface PMID- 12121612 TI - Translation is required to remove Y14 from mRNAs in the cytoplasm. AB - BACKGROUND: Y14 is an RNA binding protein which is part of a multiprotein complex, the exon-exon junction complex (EJC), that assembles on the exon-exon junctions of mRNAs produced by splicing. The position-specific binding of Y14 persists on mRNAs after their export to the cytoplasm. Thus, Y14, together with its interacting proteins, has the capacity to communicate to the cytoplasm the processing history of the mRNA, including the position of the removed introns, information that is likely to be important for defining premature termination codons. How Y14 and other components of the EJC are removed from mRNAs into the cytoplasm has not been determined. RESULTS: We show that Y14 but not another EJC component, Aly/REF, is present in polysome profile fractions containing one ribosome per mRNA. Using reporter constructs in an in vitro splicing/translation coupled system, we show that Y14 remains associated with untranslated mRNAs but is removed from translationally active mRNAs. Importantly, mRNAs whose translation in vivo is prevented by the presence of strong secondary 5' UTR structure retain Y14 in the cytoplasm. CONCLUSIONS: These findings indicate that Y14 remains associated with mRNAs in the cytoplasm until they are translated, and translation is required to remove Y14 from mRNAs. Thus, the process of translation removes the splicing-dependent EJC protein imprints, which most likely function in the surveillance of mRNAs to define premature termination codons and possibly also in modulating the translation activity of cytoplasmic mRNAs. PMID- 12121613 TI - Activation of phosphoinositide 3-kinase gamma by Ras. AB - BACKGROUND: Type I phosphoinositide 3-kinases are responsible for the hormone sensitive synthesis of the lipid messenger phosphatidylinositol(3,4,5) trisphosphate. Type IA and IB subfamily members contain a Ras binding domain and are stimulated by activated Ras proteins both in vivo and in vitro. The mechanism of Ras activation of type I PI3Ks is unknown, in part because no robust in vitro assay of this event has been established and characterized. Other Ras effectors, such as Raf and phosphoinositide-phospholipase Cepsilon, have been shown to be translocated into the plasma membrane, leading to their activation. RESULTS: We show that posttranslationally lipid-modified, activated N-, H-, K-, and R-Ras proteins can potently and substantially activate PI3Kgamma when using a stripped neutrophil membrane fraction as a source of phospholipid substrate. We have found GTPgammaS-loaded Ras can significantly (6- to 8-fold) activate PI3Kgamma when using artificial phospholipid vesicles containing their substrate, and this effect is a result of both a decrease in apparent Km for phosphatidylinositol(4,5)-bisphosphate and an increase in the apparent Vmax. However, neither in vivo nor in the two in vitro assays of Ras activation of PI3Kgamma could we detect any evidence of a Ras-dependent translocation of PI3Kgamma to its source of phospholipid substrate. CONCLUSIONS: Our data suggest that Ras activate PI3Kgamma at the level of the membrane, by allosteric modulation and/or reorientation of the PI3Kgamma, implying that Ras can activate PI3Kgamma without its membrane translocation. This view is supported by structural work that has suggested binding of Ras to PI3Kgamma results in a change in the structure of the catalytic pocket. PMID- 12121614 TI - Apoptosis protection by the Epo target Bcl-X(L) allows factor-independent differentiation of primary erythroblasts. AB - BACKGROUND: Erythropoietin (Epo) is required for correct execution of the erythroid differentiation program. Erythropoiesis requires Bcl-X(L), a major late target of Epo-receptor signaling. Mice lacking Bcl-X(L) die around embryonic age E12.5, forming normal erythroid progenitors but lacking functional red cells. Recently, serum-free culture conditions for expansion of murine red cell progenitors were developed, yielding cells capable of in vivo-like terminal differentiation into enucleated erythrocytes, in response to Epo/insulin. Here we address whether Epo function during terminal maturation involves a cytokine independent "default program," requiring only apoptosis inhibition through Epo dependent upregulation of Bcl-X(L). RESULTS: Exogenous expression of Bcl-X(L) or Bcl-2 in primary murine erythroblasts or clonal erythroblast lines derived from p53(-/-) mice allowed these cells to undergo terminal erythroid maturation, in the complete absence of cytokines. A potential autocrine Epo loop was ruled out by respective neutralizing antibodies. Importantly, sustained proliferation of Bcl-X(L)-expressing immature erythroblasts still required respective factors (Epo, stem cell factor [SCF], and the glucocorticoid receptor ligand dexamethasone [Dex]). Epo-independent differentiation in these Bcl-X(L)- or Bcl-2 expressing, primary erythroblasts was thus triggered by removal of the renewal factors SCF and Dex. This initiated the maturation-specific expression cascade of erythroid transcription factors, followed by differentiation divisions (characterized by a short G1 phase and decrease in cell size), hemoglobin accumulation, and enucleation. CONCLUSIONS: During erythroid maturation, Epo regulates red cell numbers via apoptosis inhibition, caused by Epo-dependent upregulation of the antiapoptotic protein Bcl-X(L). This allows "default" terminal differentiation of apoptosis-protected, committed erythroblasts, independent of any exogenous signals. PMID- 12121615 TI - Purification and functional characterization of SET8, a nucleosomal histone H4 lysine 20-specific methyltransferase. AB - BACKGROUND: Covalent modifications of histone N-terminal tails play fundamental roles in regulating chromatin structure and function. Extensive studies have established that acetylation of specific lysine residues in the histone tails plays an important role in transcriptional regulation. Besides acetylation, recent studies have revealed that histone methylation also has significant effects on heterochromatin formation and transcriptional regulation. Histone methylation occurs on specific arginine and lysine residues of histones H3 and H4. Thus far, only 2 residues on histone H4 are known to be methylated. While H4 arginine 3 (H4-R3) methylation is mediated by PRMT1, the enzyme(s) responsible for H4-lysine 20 (H4-K20) methylation is not known. RESULTS: To gain insight into the function of H4-K20 methylation, we set out to identify the enzyme responsible for this modification. We purified and cloned a novel human SET domain-containing protein, named SET8, which specifically methylates H4 at K20. SET8 is a single subunit enzyme and prefers nucleosomal substrates. We find that H4-K20 methylation occurs in a wide range of higher eukaryotic organisms and that SET8 homologs exist in C. elegans and Drosophila. We demonstrate that the Drosophila SET8 homolog has the same substrate specificity as its human counterpart. Importantly, disruption of SET8 in Drosophila reduces levels of H4-K20 methylation in vivo and results in lethality. Although H4-K20 methylation does not correlate with gene activity, it appears to be regulated during the cell cycle. CONCLUSIONS: We identified and characterized an evolutionarily conserved nucleosomal H4-K20-specific methyltransferase and demonstrated its essential role in Drosophila development. PMID- 12121616 TI - A CDK-activating kinase network is required in cell cycle control and transcription in fission yeast. AB - Cyclin-dependent kinases (CDKs) involved in cell cycle control require activation by phosphorylation, but CDK-activating kinase (CAK) has diverged between metazoans and budding yeast. Fission yeast has two CAKs: the essential Mcs6 complex, homologous to the metazoan CDK7 complex implicated in cell cycle control and transcription; and Csk1, a nonessential ortholog of budding yeast Cak1. Both can activate the major CDK, Cdc2, but Csk1 can also activate Mcs6, so it was unclear whether the pathway is a linear cascade or a network. Here, we show that a mutation, mcs6-13, which selectively abrogates CDK activation, blocks both G1/S and G2/M transitions, but only when csk1(+) is absent. In contrast, gradual depletion or rapid inactivation of Mcs6 in csk1(+) cells causes cell separation defects or growth arrest, respectively, accompanied by decreased phosphorylation of RNA polymerase II (RNAP II), but not of Cdc2. Finally, neither cell cycle arrest nor CAK failure is recapitulated by a second mutation in mcs6-13 that prevents Mcs6 activation by Csk1, indicating that Csk1 activates Cdc2 directly in vivo. Thus, Mcs6 acts in concert with Csk1 to activate Cdc2 and independently to support transcription and facilitate cell separation. Csk1 likewise has multiple physiologic targets, including Mcs6 and Cdc2. PMID- 12121618 TI - The ubiquitin-interacting motifs target the endocytic adaptor protein epsin for ubiquitination. AB - The covalent attachment of ubiquitin to proteins is an evolutionarily conserved signal for rapid protein degradation. However, additional cellular functions for ubiquitination are now emerging, including regulation of protein trafficking and endocytosis. For example, recent genetic studies suggested a role for ubiquitination in regulating epsin, a modular endocytic adaptor protein that functions in the assembly of clathrin-coated vesicles; however, biochemical evidence for this notion has been lacking. Epsin consists of an epsin NH(2) terminal homology (ENTH) domain that promotes the interaction with phospholipids, several AP2 binding sites, two clathrin binding sequences, and several Eps15 homology (EH) domain binding motifs. Interestingly, epsin also possesses several recently described ubiquitin-interacting motifs (UIMs) that have been postulated to bind ubiquitin. Here, we demonstrate that epsin is predominantly monoubiquitinated and resistant to proteasomal degradation. The UIMs are necessary for epsin ubiquitination but are not the site of ubiquitination. Finally, we demonstrate that the isolated UIMs from both epsin and an unrelated monoubiquitinated protein, Eps15, are sufficient to promote ubiquitination of a chimeric glutathione-S-transferase (GST)-UIM fusion protein. Thus, our data suggest that UIMs may serve as a general signal for ubiquitination. PMID- 12121617 TI - Targeted ablation of connexin26 in the inner ear epithelial gap junction network causes hearing impairment and cell death. AB - Mutations in the gene encoding the gap junction protein connexin26 (Cx26) are responsible for the autosomal recessive isolated deafness, DFNB1, which accounts for half of the cases of prelingual profound hereditary deafness in Caucasian populations. To date, in vivo approaches to decipher the role of Cx26 in the inner ear have been hampered by the embryonic lethality of the Cx26 knockout mice. To overcome this difficulty, we performed targeted ablation of Cx26 specifically in one of the two cellular networks that it underlies in the inner ear, namely, the epithelial network. We show that homozygous mutant mice, Cx26(OtogCre), have hearing impairment, but no vestibular dysfunction. The inner ear developed normally. However, on postnatal day 14 (P14), i.e., soon after the onset of hearing, cell death appeared and eventually extended to the cochlear epithelial network and sensory hair cells. Cell death initially affected only the supporting cells of the genuine sensory cell (inner hair cell, IHC), thus suggesting that it could be triggered by the IHC response to sound stimulation. Altogether, our results demonstrate that the Cx26-containing epithelial gap junction network is essential for cochlear function and cell survival. We conclude that prevention of cell death in the sensory epithelium is essential for any attempt to restore the auditory function in DFNB1 patients. PMID- 12121619 TI - Establishment of hindbrain segmental identity requires signaling by FGF3 and FGF8. AB - The hindbrain (brainstem) of all vertebrates follows a segmental developmental strategy and has been the focus of intense study not only for its intrinsic interest but also as a model for how more complex regions of the brain are patterned. Segmentation ultimately serves to organize the development of neuronal populations and their projections, and regional diversity is achieved through each segment having its own identity. The latter being established through differential expression of a hierarchy of transcription factors, including Hox genes, Krox20, and Kreisler/Valentino. Here we identify a novel signaling center in the zebrafish embryo that arises prior to establishment of segmental patterning and which is located centrally within the hindbrain territory in a region that corresponds to the presumptive rhombomere 4. We show that signaling from this region by two members of the FGF family of secreted proteins, FGF3 and FGF8, is required to establish correct segmental identity throughout the hindbrain and for subsequent neuronal development. Spatiotemporal studies of Fgf expression suggest that this patterning mechanism is conserved during hindbrain development in other vertebrate classes. PMID- 12121620 TI - Ran localizes around the microtubule spindle in vivo during mitosis in Drosophila embryos. AB - The GTPase Ran regulates multiple cellular functions throughout the cell cycle, including nucleocytoplasmic transport, nuclear membrane assembly, and spindle assembly. Ran mediates spindle assembly by affecting multiple spindle assembly pathways: microtubule dynamics, microtubule motor activity, and spindle pole assembly. Ran is predicted to facilitate spindle assembly by remaining in the GTP bound state around the chromatin in mitosis. Here, we directly test the central tenet of this hypothesis in vivo by determining the cellular localization of Ran pathway components in Drosophila embryos. We find that, during mitosis, RCC1, the nucleotide exchange factor for Ran, is associated with chromatin, while Ran and RanL43E, an allele locked in the GTP-bound state, localize around the spindle. In contrast, nuclear proteins redistribute throughout the embryo upon nuclear envelope breakdown (NEB). Thus, in vivo RanGTP has the correct spatial localization within the cell to modulate spindle assembly. PMID- 12121621 TI - Cryptochrome-deficient mice lack circadian electrical activity in the suprachiasmatic nuclei. AB - The mammalian master clock driving circadian rhythmicity in physiology and behavior resides within the suprachiasmatic nuclei (SCN) of the anterior hypothalamus. Circadian rhythms are generated by a set of clock genes via intertwined negative and positive autoregulatory transcription-translation feedback loops. The Cryptochrome 1 and 2 genes are indispensable for molecular core oscillator function, as evident from the arrhythmic wheel-running behavior and lack of rhythmic clock gene expression in mCry1/mCry2 double-mutant mice in constant darkness. In the present study, using real-time multiunit electrode activity recordings in hypothalamic slices, we show that SCN neurons from mCry deficient mice kept in constant darkness lack circadian oscillations in firing patterns. This proves that cryptochromes, and thus an intact circadian clockwork, are prerequisites for circadian electrical activity in SCN neurons. Interestingly, when mCry-deficient mice were kept in normal light-dark conditions and SCN slices were prepared 2 hr after the beginning of the day, a single noncircadian peak in neuronal activity was detected. This light-induced rise in electrical activity of the SCN may explain why mCry-deficient mice lack the arrhythmic short bouts of wheel-running activity and instead show apparently normal behavior in normal day-night cycles. PMID- 12121622 TI - Color and luminance contrasts attract independent attention. AB - Paying attention can improve vision in many ways, including some very basic functions such as contrast discrimination, a task that probably reflects very early levels of visual processing. Electrophysiological, psychophysical, and imaging studies on humans as well as single recordings in monkey show that attention can modulate the neuronal response at an early stage of visual processing, probably by acting on the response gain. Here, we measure incremental contrast thresholds for luminance and color stimuli to derive the contrast response of early neural mechanisms and their modulation by attention. We show that, for both cases, attention improves contrast discrimination, probably by multiplicatively increasing the gain of the neuronal response to contrast. However, the effects of attention are highly specific to the visual modality: concurrent attention to a competing luminance, but not chromatic pattern, greatly impedes luminance contrast discrimination; and attending to a competing chromatic, but not luminance, task impedes color contrast discrimination. Thus, the effects of attention are highly modality specific, implying separate attentional resources for different fundamental visual attributes at early stages of visual processing. PMID- 12121623 TI - Role of the arabidopsis DRM methyltransferases in de novo DNA methylation and gene silencing. AB - Proper DNA methylation patterning requires the complementary processes of de novo methylation (the initial methylation of unmethylated DNA sequences) and maintenance methylation (the faithful replication of preexisting methylation). Arabidopsis has two types of methyltransferases with demonstrated maintenance activity: MET1, which maintains CpG methylation and is homologous to mammalian DNMT1, and CHROMOMETHYLASE 3 (CMT3), which maintains CpNpG (N = A, T, C, or G) methylation and is unique to the plant kingdom. Here we describe loss-of-function mutations in the Arabidopsis DOMAINS REARRANGED METHYLASE (DRM) genes and provide evidence that they encode de novo methyltransferases. drm1 drm2 double mutants retained preexisting CpG methylation at the endogenous FWA locus but blocked de novo CpG methylation that is normally associated with FWA transgene silencing. Furthermore, drm1 drm2 double mutants blocked de novo CpNpG and asymmetric methylation and gene silencing of the endogenous SUPERMAN (SUP) gene, which is normally triggered by an inverted SUP repeat. However, drm1 drm2 double mutants did not show reactivation of previously established SUPERMAN epigenetic silenced alleles. Thus, drm mutants prevent the establishment but not the maintenance of gene silencing at FWA and SUP, suggesting that the DRMs encode the major de novo methylation enzymes affecting these genes. PMID- 12121624 TI - Nectin couples cell-cell adhesion and the actin scaffold at heterotypic testicular junctions. AB - Actin-based cell-cell adherens junctions (AJs) are crucial not only for mechanical adhesion but also for cell morphogenesis and differentiation. While organization of homotypic AJs is attributed mostly to classic cadherins, the adhesive mechanism of heterotypic AJs in more complex tissues remains to be clarified. Nectin, a member of a family of immunoglobulin-like adhesion molecules at various AJs, is a possible organizer of heterotypic AJs because of its unique heterophilic trans-interaction property. Recently, nectin-2 (-/-) mice have been shown to exhibit the defective sperm morphogenesis and the male-specific infertility, but the role of nectin in testicular AJs has not been investigated. We show here the heterotypic trans-interaction between nectin-2 in Sertoli cells and nectin-3 in spermatids at Sertoli-spermatid junctions (SspJs), heterotypic AJs in testes. Moreover, each nectin-based adhesive membrane domain exhibits one to-one colocalization with each actin bundle underlying SspJs. Inactivation of the mouse nectin-2 gene causes not only impaired adhesion but also loss of the junctional actin scaffold at SspJs, resulting in aberrant morphogenesis and positioning of spermatids. Localization of afadin, an adaptor protein of nectin with the actin cytoskeleton, is also nectin-2 dependent at SspJs. These results indicate that the nectin-afadin system plays essential roles in coupling cell cell adhesion and the cortical actin scaffold at SspJs and in subsequent sperm morphogenesis. PMID- 12121625 TI - Ran binds to chromatin by two distinct mechanisms. AB - Ran GTPase plays important roles in nucleocytoplasmic transport in interphase and in both spindle formation and nuclear envelope (NE) assembly during mitosis. The latter functions rely on the presence of high local concentrations of GTP-bound Ran near mitotic chromatin. RanGTP localization has been proposed to result from the association of Ran's GDP/GTP exchange factor, RCC1, with chromatin, but Ran is shown here to bind directly to chromatin in two modes, either dependent or independent of RCC1, and, where bound, to increase the affinity of chromatin for NE membranes. We propose that the Ran binding capacity of chromatin contributes to localized spindle and NE assembly. PMID- 12121626 TI - The basic helix-loop-helix factor olig2 is essential for the development of motoneuron and oligodendrocyte lineages. AB - Sonic hedgehog (Shh), an organizing signal from ventral midline structures, is essential for the induction and maintenance of many ventral cell types in the embryonic neural tube. Olig1 and Olig2 are related basic helix-loop-helix factors induced by Shh in the ventral neural tube. Although expression analyses and gain of-function experiments suggested that these factors were involved in motoneuron and oligodendrocyte development, they do not clearly define the functional differences between Olig1 and Olig2. We generated mice with a homozygous inactivation of Olig2. These mice did not feed and died on the day of birth. In the spinal cord of the mutant mice, motoneurons are largely eliminated and oligodendrocytes are not produced. Olig2(-/-) neuroepithelial cells in the ventral spinal cord failed to differentiate into motoneurons or oligodendrocytes and expressed an astrocyte marker, S100beta, at the time of oligodendrogenesis. Olig1 or Olig3, other family members, were expressed in the descendent cells that should have expressed Olig2. We concluded that Olig2 is an essential transcriptional regulator in motoneuron and oligodendrocyte development. Our data provide the first evidence that a single gene mutation leads to the loss of two cell types, motoneuron and oligodendrocyte. PMID- 12121627 TI - Drosophila Bruce can potently suppress Rpr- and Grim-dependent but not Hid dependent cell death. AB - Bruce is a large protein (530 kDa) that contains an N-terminal baculovirus IAP repeat (BIR) and a C-terminal ubiquitin conjugation domain (E2). BRUCE upregulation occurs in some cancers and contributes to the resistance of these cells to DNA-damaging chemotherapeutic drugs. However, it is still unknown whether Bruce inhibits apoptosis directly or instead plays some other more indirect role in mediating chemoresistance, perhaps by promoting drug export, decreasing the efficacy of DNA damage-dependent cell death signaling, or by promoting DNA repair. Here, we demonstrate, using gain-of-function and deletion alleles, that Drosophila Bruce (dBruce) can potently inhibit cell death induced by the essential Drosophila cell death activators Reaper (Rpr) and Grim but not Head involution defective (Hid). The dBruce BIR domain is not sufficient for this activity, and the E2 domain is likely required. dBruce does not promote Rpr or Grim degradation directly, but its antiapoptotic actions do require that their N termini, required for interaction with DIAP1 BIR2, be intact. dBruce does not block the activity of the apical cell death caspase Dronc or the proapoptotic Bcl 2 family member Debcl/Drob-1/dBorg-1/Dbok. Together, these results argue that dBruce can regulate cell death at a novel point. PMID- 12121628 TI - Flaws bug television gene drama. AB - MEDIAWATCH: Rejection of scientific advice by the writers of a BBC thriller based on the consequences of genetic modification of wheat with an antibiotic resistance gene renders the plot absurd, writes Bernard Dixon. PMID- 12121629 TI - Eternal vigilance on an Amazon floodplain. AB - Deep and puzzling evolutionary questions are still being raised by the unfashionable work of systematic biologists. James S. Albert describes some such work and argues why we need much more of it in the face of unprecedented loss of species through human activity. PMID- 12121631 TI - Caenorhabditis elegans par genes. PMID- 12121632 TI - Comment on Severin, F.F., and Hyman, A.A. (2002). Pheromone induces programmed cell death in S. cerevisiae. Curr. Biol. 12, R233-R235. PMID- 12121633 TI - Chromosome Organization: Reaching out to Embrace New Models. AB - New information on the architecture of protein complexes involved in the dynamic organization of chromosomes challenges existing models of their structures and provides fresh insight into their mechanisms of action. PMID- 12121634 TI - Planar polarity: location, location, location. AB - Recent studies have shown that many proteins that regulate planar polarity in the fly eye are organized into discrete membrane sites which may be crucial for coordinating groups of cells. PMID- 12121635 TI - Gravitropism: lateral thinking in auxin transport. AB - Plant stems and roots orient themselves with respect to directional illumination and gravity by differential growth on either side of the organ. A model formulated in the 1930s proposed that tropic growth curvature arose from growth hormone redistribution. Studies with Arabidopsis are beginning to reveal the mechanism. PMID- 12121636 TI - Neural development: axon regeneration derailed by dendrites. AB - Maturing neurons gradually lose the ability to regenerate axons. In the retina, signals from neighboring cells have been found to induce a perinatal switch from extension of axons to extension of dendrites, a change that may contribute to regeneration failure. PMID- 12121637 TI - Chromosome dynamics: new light on Aurora B kinase function. AB - Aurora B family kinases play an essential role in chromosome segregation and cytokinesis. Recent work suggests that the kinase activity is required for bipolar chromosome orientation, kinetochore assembly, spindle checkpoint and microtubule dynamics. Aurora B also has additional functions in chromosome condensation and cohesion. PMID- 12121638 TI - RNA export: searching for mRNA identity. AB - Efficient eukaryotic gene expression hinges on the ability of mRNA to travel from the nucleus to its cytoplasmic destination. Recent work lends insight into features that allow diverse mRNAs to be recognized by shared export machinery. PMID- 12121639 TI - Genetic code: introducing pyrrolysine. AB - Monomethylamine methyltransferase of the archaebacterium Methanosarcina barkeri contains a novel amino acid, pyrrolysine, encoded by the termination codon UAG. Initial studies suggest that pyrrolysine may be co-translationally inserted during protein synthesis, probably by a mechanism analogous to that operating during selenocysteine incorporation. PMID- 12121640 TI - Cell fusion: an EFFicient sculptor. AB - Mutations in the eff-1 gene of Caenorhabditis elegans, which prevent all cell cell fusions in the nematode's epidermis, have revealed developmental roles for cell fusion. An extracellular fusogen-like domain in EFF-1 suggests it might direct the fusion of lipid bilayers. PMID- 12121641 TI - Rice genomes: a grainy view of future evolutionary research. AB - The draft genome sequences from two subspecies of rice are powerful new tools for gene discovery in the grasses. Genome-wide comparisons of gene content and order will also shed new light on evolutionary processes. PMID- 12121642 TI - FHA: a signal transduction domain with diverse specificity and function. AB - The structure of the FHA domain of the Chfr mitotic checkpoint protein described in this issue of Structure represents one of only a few known structures of this newly discovered phosphoprotein binding domain with diverse function and specificity. PMID- 12121643 TI - Another twist in helix C and a missing pocket. AB - PKB/Akt reveals a major role for helix C in the regulation of activity in the first structure of an AGC family protein kinase in its low-activity form. PMID- 12121644 TI - Crystal structure of the FHA domain of the Chfr mitotic checkpoint protein and its complex with tungstate. AB - The Chfr mitotic checkpoint protein is frequently inactivated in human cancer. We determined the three-dimensional structure of its FHA domain in its native form and in complex with tungstate, an analog of phosphate. The structures revealed a beta sandwich fold similar to the previously determined folds of the Rad53 N- and C-terminal FHA domains, except that the Rad53 domains were monomeric, whereas the Chfr FHA domain crystallized as a segment-swapped dimer. The ability of the Chfr FHA domain to recognize tungstate suggests that it shares the ability with other FHA domains to bind phosphoproteins. Nevertheless, differences in the sequence and structure of the Chfr and Rad53 FHA domains suggest that FHA domains can be divided into families with distinct binding properties. PMID- 12121645 TI - SH3-SH2 domain orientation in Src kinases: NMR studies of Fyn. AB - The regulatory domains of Src family kinases SH3 and SH2 suppress Src activity when bound to the catalytic domain. Here, the isolated SH3-SH2 fragment from the Src family member Fyn (FynSH32) is studied by NMR. The properties of this fragment are expected to be similar to the domains in the active state, where they are dissociated from the catalytic domain. Crosscommunication between SH3 and SH2 of FynSH32, measured by chemical shift perturbation, was found to be small. Diffusion and alignment anisotropy measurements showed that SH3 and SH2 of peptide-bound FynSH32 are significantly coupled but still exhibit some interdomain flexibility. The observed average domain orientation indicates that a large SH3-SH2 domain closure is required to reach the inactive state. The implications of these results for Src regulation are discussed. PMID- 12121646 TI - The 1.1 A crystal structure of human TGF-beta type II receptor ligand binding domain. AB - Transforming growth factor beta (TGF-beta) is involved in a wide range of biological functions including development, carcinogenesis, and immune regulation. Here we report the 1.1 A resolution crystal structure of human TGF beta type II receptor ectodomain (TBRII). The overall structure of TBRII is similar to that of activin type II receptor ectodomain (ActRII) and bone morphogenic protein receptor type IA (BRIA). It displays a three-finger toxin fold with fingers formed by the beta strand pairs beta1-beta2, beta3-beta4, and beta5-beta6. The first finger in the TBRII is significantly longer than in ActRII and BRIA and folds tightly between the second finger and the C terminus. Surface charge distributions and hydrophobic patches predict potential TBRII binding sites. PMID- 12121647 TI - A dynamic analysis of the rotation mechanism for conformational change in F(1) ATPase. AB - Molecular dynamics trajectories for the bovine mitochondrial F(1)-ATPase are used to demonstrate the motions and interactions that take place during the elementary (120 degrees rotation) step of the gamma subunit. The results show how rotation of the gamma subunit induces the observed structural changes in the catalytic beta subunits. Both steric and electrostatic interactions contribute. An "ionic track" of Arg and Lys residues on the protruding portion of the gamma subunit plays a role in guiding the motions of the beta subunits. Experimental data for mutants of the DELSEED motif and the hinge region are interpreted on the basis of the molecular dynamics results. The trajectory provides a unified dynamic description of the coupled subunit motions involved in the 120 degrees rotation cycles of F(1)-ATPase. PMID- 12121648 TI - Molecular basis of mitomycin C resistance in streptomyces: structure and function of the MRD protein. AB - Mitomycin C (MC) is a potent anticancer agent. Streptomyces lavendulae, which produces MC, protects itself from the lethal effects of the drug by expressing several resistance proteins. One of them (MRD) binds MC and functions as a drug exporter. We report the crystal structure of MRD and its complex with an MC metabolite, 1,2-cis-1-hydroxy-2,7-diaminomitosene, at 1.5 A resolution. The drug is sandwiched by pi-stacking interactions of His-38 and Trp-108. MRD is a dimer. The betaalphabetabetabeta fold of the MRD molecule is reminiscent of methylmalonyl-CoA epimerase, bleomycin resistance proteins, glyoxalase I, and extradiol dioxygenases. The location of the binding site is identical to the ones in evolutionarily related enzymes, suggesting that the protein may have been recruited from a different metabolic pathway. PMID- 12121649 TI - Structural basis of von Willebrand factor activation by the snake toxin botrocetin. AB - The A1 domain of von Willebrand factor (vWF) mediates platelet adhesion to sites of vascular injury by binding to the platelet receptor glycoprotein Ib (GpIb), an interaction that is regulated by hydrodynamic shear forces. The GpIb binding surface of A1 is distinct from a regulatory region, suggesting that ligand binding is controlled allosterically. Here we report the crystal structures of the "gain-of-function" mutant A1 domain (I546V) and its complex with the exogenous activator botrocetin. We show that botrocetin switches the mutant A1 back toward the wild-type conformation, suggesting that affinity is enhanced by augmenting the GpIb binding surface rather than through allosteric control. Functional studies of platelet adhesion under flow further suggest that the activation mechanism is distinct from that of the gain-of-function mutation. PMID- 12121650 TI - Molecular mechanism for the regulation of human mitochondrial NAD(P)+-dependent malic enzyme by ATP and fumarate. AB - The regulation of human mitochondrial NAD(P)+-dependent malic enzyme (m-NAD-ME) by ATP and fumarate may be crucial for the metabolism of glutamine for energy production in rapidly proliferating tissues and tumors. Here we report the crystal structure at 2.2 A resolution of m-NAD-ME in complex with ATP, Mn2+, tartronate, and fumarate. Our structural, kinetic, and mutagenesis studies reveal unexpectedly that ATP is an active-site inhibitor of the enzyme, despite the presence of an exo binding site. The structure also reveals the allosteric binding site for fumarate in the dimer interface. Mutations in this binding site abolished the activating effects of fumarate. Comparison to the structure in the absence of fumarate indicates a possible molecular mechanism for the allosteric function of this compound. PMID- 12121651 TI - Structural and thermodynamic characterization of the DNA binding properties of a triple alanine mutant of MATalpha2. AB - Triply mutated MATalpha2 protein, alpha2-3A, in which all three major groove contacting residues are mutated to alanine, is defective in binding DNA alone or in complex with Mcm1 yet binds with MATa1 with near wild-type affinity and specificity. To gain insight into this unexpected behavior, we determined the crystal structure of the a1/alpha2-3A/DNA complex. The structure shows that the triple mutation causes a collapse of the alpha2-3A/DNA interface that results in a reorganized set of alpha2-3A/DNA contacts, thereby enabling the mutant protein to recognize the wild-type DNA sequence. Isothermal titration calorimetry measurements reveal that a much more favorable entropic component stabilizes the a1/alpha2-3A/DNA complex than the alpha2-3A/DNA complex. The combined structural and thermodynamic studies provide an explanation of how partner proteins influence the sequence specificity of a DNA binding protein. PMID- 12121652 TI - Structure of CcmG/DsbE at 1.14 A resolution: high-fidelity reducing activity in an indiscriminately oxidizing environment. AB - CcmG is unlike other periplasmic thioredoxin (TRX)-like proteins in that it has a specific reducing activity in an oxidizing environment and a high fidelity of interaction. These two unusual properties are required for its role in c-type cytochrome maturation. The crystal structure of CcmG reveals a modified TRX fold with an unusually acidic active site and a groove formed from two inserts in the fold. Deletion of one of the groove-forming inserts disrupts c-type cytochrome formation. Two unique structural features of CcmG-an acidic active site and an adjacent groove-appear to be necessary to convert an indiscriminately binding scaffold, the TRX fold, into a highly specific redox protein. PMID- 12121653 TI - Noncompetitive antibody neutralization of IL-10 revealed by protein engineering and x-ray crystallography. AB - IL-10 is a dimeric cytokine that must engage its high-affinity cell surface receptor, IL-10R1, to induce multiple cellular activities. Here we report the 1.9 A crystal structure of an engineered IL-10 monomer (IL-10M1) in complex with a neutralizing Fab fragment (9D7Fab). 9D7Fab and IL-10R1 bind distinct nonoverlapping surfaces on IL-10M1. Antagonism of the IL-10M1/IL-10R1 interaction is the result of 9D7Fab-induced conformational changes in the CD loop of IL-10M1 that indirectly alter the structure of the IL-10R1 binding site. A single mutation (Ile87Ala) in the same CD loop region of the Epstein-Barr virus IL-10 (ebvIL-10) also reduces IL-10R1 binding affinity, suggesting that ebvIL-10 and 9D7Fab use similar allosteric mechanisms to modulate IL-10R1 affinity and biological activity. PMID- 12121654 TI - Stabilization of globular proteins via introduction of temperature-activated elastin-based switches. AB - To investigate whether swapping native turns of a globular protein with an elastin-based turn sequence (VPGVG) can increase its thermostability, we have performed molecular dynamics simulations of wild-type chymotrypsin inhibitor 2 (CI2) and variants containing elastin-based turns at 10 degrees C and 40 degrees C. Wild-type CI2 is more stable at 10 degrees C, while both of the variant forms are more stable at 40 degrees C. Detailed analyses indicate that the elastin based turns do indeed contribute to the inverse temperature behavior of the modified proteins. Therefore, swapping a wild-type turn sequence with an elastin based turn provides a novel way to both improve stability of target proteins at body temperature and to possibly introduce a temperature-sensitive switch. PMID- 12121655 TI - Crystal structure of a human rhinovirus that displays part of the HIV-1 V3 loop and induces neutralizing antibodies against HIV-1. AB - We report the 2.7 A resolution structure of a chimeric rhinovirus, MN-III-2, that displays part of the HIV-1 gp120 V3 loop and elicits HIV-neutralizing antibodies. The V3 loop insert is dominated by two type I beta turns. The structures of two adjacent tripeptides resemble those of analogous segments in three Fab/V3 loop peptide complexes. Although two of the three corresponding antibodies bind and neutralize MN-III-2 well, only one of the three can bind without significant rearrangement. These results suggest that the V3 loop insert: (1) can share some local conformational similarity to V3 loop sequences presented on different structural frameworks; (2) must be able to adopt multiple conformations, even in a relatively constrained environment; and (3) may mimic the conformational variability of the epitope on HIV-1, increasing the likelihood of eliciting appropriate neutralizing immune responses. PMID- 12121656 TI - Structure-based discovery of a novel, noncovalent inhibitor of AmpC beta lactamase. AB - beta-lactamases are the most widespread resistance mechanisms to beta-lactam antibiotics, and there is a pressing need for novel, non-beta-lactam drugs. A database of over 200,000 compounds was docked to the active site of AmpC beta lactamase to identify potential inhibitors. Fifty-six compounds were tested, and three had K(i) values of 650 microM or better. The best of these, 3-[(4 chloroanilino)sulfonyl]thiophene-2-carboxylic acid, was a competitive noncovalent inhibitor (K(i) = 26 microM), which also reversed resistance to beta-lactams in bacteria expressing AmpC. The structure of AmpC in complex with this compound was determined by X-ray crystallography to 1.94 A and reveals that the inhibitor interacts with key active-site residues in sites targeted in the docking calculation. Indeed, the experimentally determined conformation of the inhibitor closely resembles the prediction. The structure of the enzyme-inhibitor complex presents an opportunity to improve binding affinity in a novel series of inhibitors discovered by structure-based methods. PMID- 12121657 TI - Further checkpoints in Th1 development. AB - Tight control of Th1 immunity is essential to prevent immunopathology. Central to control of the IFN-gamma gene is the transcription factor T-bet, whose induction is Stat-1 dependent. IL-12 is dominant in directing Th1 development, while synergizing with IL-18 for IFNgamma production from differentiated Th1 cells. In this issue of Immunity, IL-27 is described, which acts in synergy with IL-12 early in Th1 development from naive T cells via the receptor TCCR/WSX-1. We review the coordination of these checkpoints in Th1 development and function. PMID- 12121658 TI - Activated T cell death in vivo mediated by proapoptotic bcl-2 family member bim. AB - At the end of the T cell response, the majority of the activated T cells die. We activated Vbeta8(+) T cells with staphylococcal enterotoxin B (SEB) in vivo and monitored the expansion and deletion of Vbeta8(+) T cells. We found that, in response to SEB, activated T cells died in vivo in the absence of Fas or TNF-R signaling but not when they overexpressed human Bcl-2. We also found that Vbeta8(+) T cells from Bim-deficient mice are resistant to SEB-induced deletion. While Bim levels did not change, endogenous Bcl-2 levels within Vbeta8(+) T cells decrease following SEB injection. Thus, the death of superantigen-stimulated T cells in vivo is mediated by Bim and may be modulated by a decrease in Bcl-2. PMID- 12121659 TI - The CD28 signaling pathway regulates glucose metabolism. AB - Lymphocyte activation initiates a program of cell growth, proliferation, and differentiation that increases metabolic demand. Although T cells increase glucose uptake and glycolysis during an immune response, the signaling pathways that regulate these increases remain largely unknown. Here we show that CD28 costimulation, acting through phosphatidylinositol 3'-kinase (PI3K) and Akt, is required for T cells to increase their glycolytic rate in response to activation. Furthermore, CD28 controls a primary response pathway, inducing a level of glucose uptake and glycolysis in excess of that needed to maintain cellular ATP/ADP levels or macromolecular synthesis. These data suggest that CD28 costimulation functions to increase glycolytic flux, allowing T cells to anticipate energetic and biosynthetic needs associated with a sustained response. PMID- 12121660 TI - IL-27, a heterodimeric cytokine composed of EBI3 and p28 protein, induces proliferation of naive CD4+ T cells. AB - An efficient Th1-driven adaptive immune response requires activation of the T cell receptor and secretion of the T cell stimulatory cytokine IL-12 by activated antigen-presenting cells. IL-12 triggers Th1 polarization of naive CD4(+) T cells and secretion of IFN-gamma. We describe a new heterodimeric cytokine termed IL-27 that consists of EBI3, an IL-12p40-related protein, and p28, a newly discovered IL-12p35-related polypeptide. IL-27 is an early product of activated antigen presenting cells and drives rapid clonal expansion of naive but not memory CD4(+) T cells. It also strongly synergizes with IL-12 to trigger IFN-gamma production of naive CD4(+) T cells. IL-27 mediates its biologic effects through the orphan cytokine receptor WSX-1/TCCR. PMID- 12121661 TI - CR3 (CD11b/CD18) and CR4 (CD11c/CD18) are involved in complement-independent antibody-mediated phagocytosis of Cryptococcus neoformans. AB - IgM and IgA to the Cryptococcus neoformans capsular glucuronoxylomannan (GXM) promote complement-independent phagocytosis by macrophages with efficiency comparable to that of IgG1. IgM- and IgA-mediated phagocytosis of C. neoformans was proportional to CR3 expression, inhibited by Abs to CR3 (CD11b/CD18) and CR4 (CD11c/CD18), and dramatically reduced with macrophages of CD18-deficient mice. IgM and IgA promoted ingestion of yeast cells by CHO cells expressing CR3 and CR4. In contrast, IgG1-mediated phagocytosis was only partially inhibited by Abs to CR3 and CR4. Phagocytosis by IgM and IgA but not IgG1 was inhibited by soluble GXM, which binds CD18. Involvement of CR in antibody-mediated complement independent phagocytosis indicates a new link between innate and adaptive immune systems. PMID- 12121662 TI - Identification and characterization of thymic epithelial progenitor cells. AB - T cell differentiation and repertoire selection depend critically on several distinct thymic epithelial cell types, whose lineage relationships are unclear. We have investigated these relationships via functional analysis of the epithelial populations within the thymic primordium. Here, we show that mAbs MTS20 and MTS24 identify a population of cells that, when purified and grafted ectopically, can differentiate into all known thymic epithelial cell types, attract lymphoid progenitors, and support CD4(+) and CD8(+) T cell development in nude mice. In contrast, other epithelial populations in the thymic primordium can fulfill none of these functions. These data establish that the MTS20(+)24(+) population is sufficient to generate a functional thymus in vivo and thus argue strongly that all thymic epithelial cell types derive from a common progenitor cell. PMID- 12121663 TI - Th1 cells regulate hematopoietic progenitor cell homeostasis by production of oncostatin M. AB - Regulation of hematopoietic progenitor cell homeostasis is crucial for maintenance of innate immunity and the ability of the body to respond to injury and infection. In this report, we demonstrate that progenitor cell numbers and cycling status in vivo are dramatically increased in mice deficient in Stat6 and decreased in mice deficient in Stat4, targeted mutations which also alter T helper cell polarization. Experiments using mice that have T cell restricted transgenic expression of Stat4 or Stat6 or have been in vivo depleted of T cell subsets demonstrate that CD4(+) T cells regulate progenitor cell activity. Injection of the Th1 cytokine Oncostatin M but not other cytokines into Stat4 deficient mice recovers progenitor cell activity to wild-type levels. Thus, T helper cells actively regulate hematopoietic progenitor cell homeostasis. PMID- 12121664 TI - Distinct structure and signaling potential of the gamma delta TCR complex. AB - Alpha beta and gamma delta T cells are distinguished by the clonotypic subunits contained within their TCRs. Although the alpha beta TCR has been well characterized, much less is known about the gamma delta TCR. Here, we report that, unlike alpha beta T CRs, most gamma delta TCRs expressed on ex vivo gamma delta T cells lack CD3 delta. Despite this structural difference, signal transduction by the gamma delta TCR is superior to that of the alpha beta TCR, as measured by its ability to induce calcium mobilization, ERK activation, and cellular proliferation. Additionally, the TCR complexes expressed on primary gamma delta T cells contain only zeta zeta homodimers; however, following activation and expansion, Fc epsilon R1 gamma is expressed and is included in the gamma delta TCR complex. These results reveal fundamental differences in the primary structure and signaling potential of the alpha beta- and gamma delta TCR complexes. PMID- 12121665 TI - Immature CD4(+)CD8(+) thymocytes form a multifocal immunological synapse with sustained tyrosine phosphorylation. AB - The immunological synapse formed during mature T cell activation consists of a central cluster of TCR and MHC molecules surrounded by a ring of LFA-1 and ICAM 1. We examined synapse formation in thymocytes undergoing activation in a lipid bilayer system by following the movement of fluorescent MHC and ICAM-1 molecules. Immature CD4(+)CD8(+) thymocytes formed a decentralized synapse with multiple foci of MHC accumulation corresponding to areas of exclusion of ICAM-1. The MHC clusters and ICAM-1 holes were mobile and transient and correlated with active and sustained signaling, as shown by staining with antibodies against phosphotyrosine and activated Lck. Our findings show that signaling in immature thymocytes can result from a novel, multifocal pattern of receptor accumulation. PMID- 12121666 TI - Salp15, an ixodes scapularis salivary protein, inhibits CD4(+) T cell activation. AB - Tick saliva has pleiotropic properties that facilitate persistence of the arthropod upon the host. We now describe a feeding-inducible protein in Ixodes scapularis saliva, Salp15, that inhibits CD4(+) T cell activation. The mechanism involves the repression of calcium fluxes triggered by TCR ligation and results in lower production of interleukin-2. Salp15 also inhibits the development of CD4(+) T cell-mediated immune responses in vivo, demonstrating the functional importance of this protein. Salp15 provides a molecular basis for understanding the immunosuppressive activity of I. scapularis saliva and vector-host interactions. PMID- 12121667 TI - Redundant and alternative roles for activating Fc receptors and complement in an antibody-dependent model of autoimmune vitiligo. AB - Complement and Fc receptor (FcR)-positive cells mediate effector functions of antibodies. Antibody-dependent immunity against the melanosome membrane glycoprotein gp75/tyrosinase-related protein-1 (TYRP-1) of melanocytes leads to autoimmune hypopigmentation (vitiligo) in mice. Hypopigmentation occurred in mice deficient in activating FcR containing the common gamma subunit (Fc gamma R gamma(-/-)) and in mice deficient in the C3 complement component. Mice doubly deficient in both Fc gamma R gamma and C3 did not develop hypopigmentation, suggesting that complement and Fc gamma R formed redundant mechanisms. Following passive immunization with antibody, no further adaptive immune responses were required. Chimeric Fc gamma R gamma(-/-),C3(-/-) mice reconstituted with bone marrow from either Fc gamma R gamma(-/-) or C3(-/-) mice or adoptively transferred with Fc gamma R gamma(+/-) macrophages did develop antibody-mediated hypopigmentation. Thus, either complement or macrophages expressing activating Fc gamma R can independently and alternatively mediate disease in a model of autoimmune vitiligo. PMID- 12121668 TI - Inactivation of Notch1 impairs VDJbeta rearrangement and allows pre-TCR independent survival of early alpha beta Lineage Thymocytes. AB - Notch proteins influence cell fate decisions in many developmental systems. During lymphoid development, Notch1 signaling is essential to direct a bipotent T/B precursor toward the T cell fate, but the role of Notch1 at later stages of T cell development remains controversial. We have recently reported that tissue specific inactivation of Notch1 in immature (CD44(-) CD25(+)) thymocytes does not affect subsequent T cell development. Here, we demonstrate that loss of Notch1 signaling at an earlier (CD44(+)CD25(+)) developmental stage results in severe perturbation of alpha beta but not gamma delta lineage development. Immature Notch1(-/-) thymocytes show impaired VDJ beta rearrangement and aberrant pre-TCR independent survival. Collectively, our data demonstrate that Notch1 controls several nonredundant functions necessary for alpha beta lineage development. PMID- 12121670 TI - High dose chemotherapy and autologous hematopoietic stem cell transplantation for rheumatoid arthritis: a review. AB - A new treatment approach, involving intense immunosuppression and autologous hematopoietic stem cell transplantation (SCT), has emerged in recent years for the treatment of severe, refractory rheumatic autoimmune diseases including rheumatoid arthritis (RA). The rationale of this strategy is based on the concept of immunoablation by intense immunosuppression with subsequent regeneration of naive T lymphocytes derived from reinfused hematopoietic progenitor cells. Patients with a therapy-refractory, progressively erosive disease who are at risk of functional disability and early mortality are considered eligible for treatment with autologous SCT. The goal is long-term improvement of disease activity and quality of life. However, when offering SCT to RA patients these benefits should be balanced against toxicities and treatment-related mortality. In several patients with intractable RA, long-term remissions were observed with this strategy, but failures have been reported as well. Only small numbers of RA patients have been treated thus far. Although different treatment protocols have been used, high dose chemotherapy as a means to achieve immunoablation has been invariably used in all studies. In this review we discuss background, clinical results, protocols, and future prospects of high dose chemotherapy and autologous SCT for RA. PMID- 12121669 TI - Autoregulation of NFATc1/A expression facilitates effector T cells to escape from rapid apoptosis. AB - Threshold levels of individual NFAT factors appear to be critical for apoptosis induction in effector T cells. In these cells, the short isoform A of NFATc1 is induced to high levels due to the autoregulation of the NFATc1 promoter P1 by NFATs. P1 is located within a CpG island in front of exon 1, represents a DNase I hypersensitive chromatin site, and harbors several sites for binding of inducible transcription factors, including a tandemly arranged NFAT site. A second promoter, P2, before exon 2, is not controlled by NFATs and directs synthesis of the longer NFATc1/B+C isoforms. Contrary to other NFATs, NFATc1/A is unable to promote apoptosis, suggesting that NFATc1/A enhances effector functions without promoting apoptosis of effector T cells. PMID- 12121671 TI - NK cells become Ki-67+ in MLC and expand depending on the lack of ligand for KIR on stimulator cells in IL-2 supplemented MLC. AB - Mixed lymphocyte culture (MLC) response was measured with the use of Ki-67 monoclonal antibody and responding cells were verified by CD3 and CD56 surface markers staining. Stimulator cells were discriminated from responder cells on the basis of forward and side scatter. Allogeneic, but not autologous MLC had Ki-67+ responder cells in lymphocyte gate at the end of the culture. In allogeneic MLC T cells and natural killer (NK) cells were in a similar proportion Ki-67+ (mean +/- SD: 59.25% +/- 9.72% versus 61.75% +/- 13.2%). Ki-67+ NK cells had higher CD56 mean fluorescence intensity than those lacking Ki-67 (745+/-357 versus 196+/-56 p < 0.0001). NK cells contribution to responding lymphocytes was positively correlated with the percentage of Ki67+ cells in NK population by the end of the culture (r = 0.74, p = 0.002). NK cells response in MLC increased upon supplementation of the culture medium with human recombintant interleukin-2 (IL 2). Responder cells from single individual were tested with 8 Bw4+ and 8 Bw4- as well as with 9 CNK1+ and 9 CNK1- stimulators. In IL-2 supplemented MLC killer inhibitory receptor expressing cells expanded when ligands for this receptor were absent in stimulating population. Consequently, stimulator cells lacking Bw4 promoted NKB1+ cells expansion (7.2% +/- 3% versus 3.6% +/- 1%, p = 0.0031), whereas HLA-C NK1 negative stimulators promoted CD158a+ cells expansion (9.6% +/- 4.8% versus 6% +/- 2.6%, p = 0.0385). PMID- 12121672 TI - Human immune response in schistosomiasis: the role of P24 in the modulation of cellular reactivity to Schistosoma mansoni antigens. AB - Schistosome antigenic components are being tested as vaccine candidates with various degrees of success, but there are only few reports using multivalent antigens to stimulate an appropriate immune response that leads to resistance or granuloma modulation. We investigated the in vitro response of peripheral blood mononuclear cells (PBMC) from chronic intestinal schistosomiasis individuals to PIII, a multivalent antigen prepared from Schistosoma mansoni adult worm antigen, and response to P24, a single antigen obtained from PIII. Treatment of PBMC with either PIII or P24 caused significant decrease in cellular proliferation and granuloma formation induced by S. mansoni antigens, and a significant elevation in IL-10 and TNF-alpha but not in IFN-gamma production. Moreover, P24 promoted an elevation in TNF-alpha level on the in vitro granuloma reaction, when cocultured with polyacrylamide beads (PB) coupled to S. mansoni antigens. These findings suggest that, besides inducing protective immunity, PIII and P24 antigens seem to be important in the regulation of in vitro granuloma formation through stimulation of IL-10 and TNF-alpha production in human schistosomiasis. The more pronounced effect of P24 on reducing the in vitro granulomatous reaction could be associated with a balance between IL-10 and TNF-alpha production. PMID- 12121673 TI - The HLA class I A locus affects susceptibility to type 1 diabetes. AB - Human leukocyte antigen A (HLA-A) genotypes were determined for samples from 283 multiplex, Caucasian, type 1 diabetes families from the Human Biological Data Interchange (HBDI) using an immobilized probe assay. Distribution of HLA-A alleles transmitted to patients was significantly different from that in affected family-based controls (AFBAC) (p = 0.004). Transmission disequilibrium test (TDT) analysis revealed differential transmission of several HLA-A alleles from parents to affected offspring. HLA class II DRB1 and DQB1 loci were also typed, allowing assignment of HLA-A alleles to haplotypes and calculation of linkage disequilibrium values. Some of the apparent effects of HLA-A alleles on type 1 diabetes susceptibility were attributable to linkage disequilibrium with DR and DQ alleles, although others were not. The differences in frequencies between patients and controls of alleles A*0101, A*2402, and A*3002 could not be explained by linkage disequilibrium alone. Our results suggest an important role for class I antigens in modulating susceptibility to type 1 diabetes. PMID- 12121674 TI - Relative HLA-DRB1*04 allele frequencies in five United States populations found in a hematopoietic stem cell volunteer donor registry and seven new DRB1*04 alleles. AB - The frequencies of 29 HLA-DRB1*04 alleles were determined for five major U.S. populations found within a hematopoietic stem cell volunteer donor registry. One hundred sixty-one DRB1*04 positive individuals from each of the self-described groups, Caucasians, African-Americans, Asian/Pacific Islanders, Hispanics, and Native Americans, were randomly chosen from a database of 82,979 unrelated persons. Subjected to polymerase chain reaction-sequence-specific oligonucleotide probe (PCR-SSOP) typing, these 805 individuals carried a total of ten different DRB1*04 alleles, ranging from DRB1*0401 to DRB1*0411 with DRB1*0409 conspicuously absent from all five groups. The distribution of DRB1*04 alleles varied among the groups, with DRB1*0401 being predominant in Caucasians, African-Americans, and Native Americans. DRB1*0404 and DRB1*0407 were the two most commonly observed alleles in Hispanics, whereas DRB1*0405 and DRB1*04031 were most common in Asian/Pacific Islanders. The remaining 18 DRB1*04 alleles known at the time of the study were not observed. Although not observed in the frequency study, seven previously unreported DRB1*04 alleles are also described. PMID- 12121675 TI - High variability of HLA-B27 alleles in ankylosing spondylitis and related spondyloarthropathies in the population of northern Spain. AB - The distribution of B27 alleles (B*2701-23) was characterized by PCR-SSP in ankylosing spondylitis and related spondyloarthropathies (SpA) in a sample of B27 positive patients from northern Spain. Six B27 alleles were identified: B*2705,02,03,07,08 and B*2713. B*2705 and 02 were the most common alleles in the SpA studied: ankylosing spondylitis (AS) (n = 89), reactive arthritis (ReA) (n = 11), psoriatic arthritis (PsA) (n = 29), and inflammatory bowel disease (IBD) (n = 21). B*2707 and B*2708 were found in PsA patients and B*2703 in one patient with IBD. B*2713 was identified in a healthy control family. B*2713 has not been reported to be represented in either ethnic group. Thus, this population shows higher levels of B27 diversity than other Caucasian groups. PMID- 12121676 TI - Pediatric celiac disease in India is associated with multiple DR3-DQ2 haplotypes. AB - The role of human leukocyte antigen (HLA) DQ2 heterodimer (DQA1*0501-DQB1*0201) in presenting gluten peptides to effector T cells in celiac disease (CD) has been well documented. Because HLA-DQ2 is carried on DR3 haplotypes due to linkage disequilibrium, such haplotypes are encountered more frequently in patients with autoimmune disease. This study analyzed 35 North Indian children below 15 years of age and diagnosed to have CD as per the ESPGAN criteria, which included histopathologic alterations in duodenal biopsies, clinical response to gluten withdrawal, and presence of antiendomysial antibodies. The HLA class I and class II alleles were determined by polymerase chain reaction-sequence-specific primers, sequence-specific oligonucleotide probe, and reverse line strip molecular techniques. A statistically significant positive association of the disease with HLA-DRB1*03 (94.2% versus 22.1% in controls, chi(2) = 73.4, p = 7.54E-11), and a negative association with DRB1*15 (chi(2) = 7.4, p = 6.5E-03) and DRB1*13 alleles was observed. The HLA-DQB1*0201 was observed in all the 35 patients (100%), whereas the DQ2 heterodimer alpha(0)beta(0) occurred in 97.1% of CD patients (31.4% in double dose, 65.7% in single dose) and revealed significant deviation from healthy controls (chi(2) = 102.08, p = 7.56E-11). Further analysis revealed involvement of multiple DR3+ve haplotypes with CD in Indians, of which A26-B8-DR3 was the most common DR3 haplotype among patients (34.28%, chi(2) = 40.57, p = 2.65E-10) followed by Ax-B21-DR3 (11.4%) (chi(2) = 13.8, p = 2E-04) and the classical Caucasian haplotype A1-B8-DR3 (5.7%). The former two haplotypes are characteristic of Asian Indians and are involved in the development of CD. We conclude that the high risk DR3 haplotypes that play a crucial role in the development of CD are unique in Asian Indians. Detailed analysis of these haplotypes in Indian patients with autoimmune diseases may help understand the influence of other intervening genes within the major histocompatibility complex. PMID- 12121677 TI - An unequal crossover event in RCCX modules of the human MHC resulting in the formation of a TNXB/TNXA hybrid and deletion of the CYP21A. AB - The central region of the human major histocompatibility complex contains tandemly arranged genes of RP, C4, CYP21, and TNX. The C4 gene region is prone to rearrangements that generates duplications, conversions, and deletions. Diversity in gene number and size causes reorganization and may lead to genetic disorders. The RP, C4, CYP21, and TNX genes form a genetic unit called RCCX. We describe molecular studies on RCCX haplotypes revealing a unique recombination giving rise to a TNXB/TNXA hybrid gene, CYP21A deletion and CYP21B duplication on one chromosome of the propositus. His other chromosome carries a deletion of CYP21A TNXA-RP2-C4B genes, resulting in the total absence of CYP21A genes and the presence of three CYP21B genes in the genome. PMID- 12121678 TI - Genetic association between interleukin-10 gene promoter region polymorphisms and type 1 diabetes age-at-onset. AB - This study investigated whether interleukin-10 (IL-10) gene promoter region polymorphisms are associated with susceptibility to or clinical presentation of type 1 diabetes. The frequency of -1082G/A, -819C/T, and -592C/A polymorphisms was analyzed in 128 Japanese patients with type 1 diabetes and in 107 healthy control subjects in a case-controlled study. The allelic and haplotypic frequencies of the IL-10 gene promoter region polymorphisms were similar in patients with type 1 diabetes and in control subjects. However, the -819T and 592A allele were associated with adult-onset (>18 years) of the disease (p = 0.037). Furthermore, the frequency of ATA haplotype was increased in adult-onset patients than that in early-onset patients (< or =18 years; p = 0.037). Among the genotypes comprising ATA haplotype, the frequency of ATA/ATA was significantly higher in adult-onset patients than in early-onset patients (p = 0.004). These results suggest that the IL-10 gene promoter polymorphisms are associated with the age-at-onset in Japanese patients with type 1 diabetes. PMID- 12121679 TI - Association between interleukin-6 gene polymorphism and human T-cell leukemia virus type I associated myelopathy. AB - We studied cytokine gene polymorphisms in the promoter region, including interleukin (IL)-6, IL-1beta, and IL-10, in Japanese patients with human T-cell leukemia virus type I (HTLV-I) associated myelopathy (HAM) (n = 65), asymptomatic HTLV-I carriers (n = 143), and HTLV-I seronegative, normal controls (n = 160). There was a significant difference between HAM patients and HTLV-I carriers in the distribution of IL-6 promoter polymorphism at position -634 (chi(2) = 9.90, p = 0.0071). The IL-6 genotype was also significantly different between HAM patients and normal controls (chi(2) = 11.53, p = 0.0033), while a similar distribution was observed in IL-1beta and IL-10 polymorphisms among HAM patients, carriers, and normal controls. The results suggest that IL-6 gene region may contribute to susceptibility to HAM, and that aberrant cytokine productions could be involved in the development of HAM. PMID- 12121680 TI - Neither high-pass filtering nor mathematical differentiation of the EMG signals can considerably reduce cross-talk. AB - Using mathematical simulation of motor unit potentials (MUPs), detected by a point and rectangular plate electrode, we have shown that the muscle tissue does not act like a low-pass frequency filter on MUPs. Depending on the electrode type and its longitudinal position, the relative weight of the terminal phases (reflecting the excitation extinction) in MUPs and thus of high frequencies in the MUP power spectrum, increase with the MU depth. Therefore, high-pass filtering or differentiating signals detected neither monopolarly nor bipolarly could eliminate the cross-talk produced by high frequency components of MUPs from deep MUs. Such methods could be effective against the main components but not against the MUP leading edge and terminal phases. To reduce the cross-talk, position of the detecting electrodes should correspond to anatomy of muscles producing the cross-talk. Monopolar electrode should be located above the ends of the muscles. Cross-talk of the muscles located beyond the muscle of interest could be higher than that produced above the end-plate of deep muscles. On the contrary, under detection by a longitudinal bipolar electrode, the cross-talk is much smaller above the end-plate region or beyond deep muscles. The cross-talk is the greatest above the ends of the deep muscles. PMID- 12121681 TI - Reduction of the muscle activity associated with self-imposed electrical stimulation of mixed nerves supplying lower limb muscles in man. AB - Similarly to brief hammer taps self-imposed on the limb segments, self-triggered electrical stimulation delivered to the tibial, femoral or peroneal nerves are associated with anticipatory reduction in the muscle activity (RMA) of the target leg muscles. The anticipatory RMA starts shortly before the expected perturbation and lasts until the onset of the impact. The purpose of the present work is to study to what extent the selective activation of the different homonymous and heteronymous afferents could modify the central programs and the associated time and space distribution of the anticipatory RMA pattern in the target leg muscles. Our results show that the anticipatory RMA pattern is a result mainly of a pre programmed non-specific central command, rather than consequence of the specific composition of the afferent volleys originating from different homonymous and heteronymous nerves. The commands for the voluntary movement triggering the stimulation and the associated anticipatory RMA are closely linked but independently controlled. By their synchronisation and co-ordination the central nervous system accounts the differences in length of the particular motor pathways. It is suggested, that in contrast to the classic anticipatory postural adjustments, the anticipatory RMA is presumably an alternative mode of feed forward control diminishing the undesired effects of the afferent inputs. PMID- 12121682 TI - A motion of forearm supination with maintenance of elbow flexion produced by electrical stimulation to two elbow flexors in humans. AB - Motions of the forearm induced by electrical stimulation to two elbow flexors (brachioradialis: BR, biceps brachii: BB) were examined in five healthy human subjects. Stainless steel wire electrodes were implanted percutaneously into each motor point of the muscles. The muscles were stimulated separately with a computer-controlled multi-channel stimulator. The motions were taken with a digital video system. Angular changes of the motions in elbow flexion/extension and forearm pronation/supination were measured. Electromyograms (EMG) of BR, BB, and the triceps brachii (TB) were recorded. Electrical stimulation to BR induced a motion of flexion and that to BB motions of flexion and supination. The stimulation to BR with an adequate intensity provided holding of flexion with the prone forearm in all the subjects. In this situation, additional stimulation to BB resulted in motions of flexion and supination. However, the additional stimulation accompanied with a decrease of the stimulation intensity for BR provided a motion of supination with maintenance of the flexion in all the subjects. Since during the stimulation BR, BB, and TB showed no voluntary contraction in EMG, it is suggested that modulation of contraction between BR and BB by the stimulation can produce force in supination with keeping constant force in flexion to support the weight below the elbow. PMID- 12121683 TI - Development of dynamic knee stability after acute ACL injury. AB - Recently, a training program that includes perturbation of support surfaces has been shown to allow most active individuals with ACL injury who pass a screening examination to successfully return to high level activities. The purpose of this study was to identify the effect of this rehabilitation program on involved side muscle activation during walking in subjects with acute ACL rupture and to determine if the activation changes were coincident with improved function. Nine subjects with an acute, unilateral ACL injury or rupture of an ACL graft, who met the screening examination criteria, received ten sessions of rehabilitation that included perturbation training. Motion analysis of five self-paced walking trials were performed before and after training. Electromyographic (EMG) data were collected during stance. After training during walking, the vastus lateralis (VL) integral of activity increased, and relationships between muscles were significantly altered. During walking, VL activation variables were dependent on lateral hamstrings (LH) and/or the soleus (SOL) activation, while no relationships were found before training. Function improved after training, and all subjects returned to their pre-injury activities without experiencing instability. The relationships formed between muscles post-training suggests that perturbation training enhances dynamic knee stability by inducing a well coordinated strategy among muscles that affect tibial translation. PMID- 12121684 TI - Upper-limb surface electro-myography at maximum supination and pronation torques: the effect of elbow and forearm angle. AB - Forearm pronation and supination, and increased muscular activity in the wrist extensors have been both linked separately to work-related injuries of the upper limb, especially humeral epicondylitis. However, there is a lack of information on forearm torque strength at ranges of elbow and forearm angles typical of industrial tasks. There is a need for strength data on forearm torques at different upper limb angles to be investigated. Such a study should also include the measurement of muscular activity for the prime torque muscles and also other muscles at possible risk of injury due to high exertion levels during tasks requiring forearm torques.Twenty-four male subjects participated in the study that involved maximum forearm torque exertions for the right arm, in the pronation and supination directions, and at four elbow and three forearm rotation angles. Surface EMG (SEMG) was used to evaluate the muscular activity of the pronator teres (PT), pronator quadratus (PQ), biceps brachi (BB), brachioradialis (BR), mid deltoid (DT) and the extensor carpi radialis brevis (ECRB) during maximum torque exertions. Repeated measures ANOVA indicated that both direction and forearm angle had a significant effect on the maximum torques (p<0.05) while elbow angle and the interactions were highly significant (p<0.001). The results revealed that supination torques were stronger overall with a mean maximum value of 16.2 Nm recorded for the forearm 75% prone. Mean maximum pronation torque was recorded as 13.1 Nm for a neutral forearm with the elbow flexed at 45 degrees. The data also indicated that forearm angle had a greater effect on supination torque than pronation torque. Supination torques were stronger for the mid-range of elbow flexion, but pronation torques increased with increasing elbow extension. The strength profiles for the maximum torque exertions were reflected in the EMG changes in the prime supinators and pronators. In addition, the EMG data expressed as the percentage of Maximum Voluntary Electrical activity (MVE), revealed high muscular activity in the ECRB for both supination (26-43% MVE) and pronation torques (17-55% MVE). The results suggest that the ECRB acts as a stabiliser to the forearm flexors for gripping during pronation torques depending on forearm angle, but acts as a prime mover in wrist extension for supination torques with little effect of elbow and forearm angle. This indicates a direct link between forearm rotations against resistance and high muscular activity in the wrist extensors, thereby increasing stress on the forearm musculo-skeletal system, especially the lateral epicondyle. PMID- 12121685 TI - Neuromuscular fatigue of elbow flexor muscles of dominant and non-dominant arms in healthy humans. AB - The purpose of this study was to assess differences in fatigue-related changes in variables related to structures within the neuromuscular system, between the dominant and non-dominant elbow flexor muscles of right-handed individuals. Two experimental sessions were performed on the right arm and one on the left arm. For each session, maximum voluntary torque, level of voluntary activation, M-wave amplitude, twitch/train or twitch/doublet torque ratio and EMG median frequency were obtained before and up to 20 min after a sustained maximum isometric fatigue task. Our main results were: 1) reproducible fatigue-induced changes in all variables of interest between the two sessions performed with the right arm, 2) significantly greater failure in voluntary activation and neuromuscular propagation with sustained activity for the non-dominant compared with dominant side, and 3) no effect of dominance on MVC torque, endurance time, and fatigue induced changes in EMG median frequency and elicited torques. These results suggest that the preferential use of elbow flexor muscles with the dominant arm leads to more fatigue resistance in certain structures/mechanisms of the neuromuscular system, but not in others. PMID- 12121686 TI - Influence of interelectrode distance and force level on the spectral parameters of surface electromyographic recordings from the lumbar muscles. AB - In order to study the influence of interelectrode distance and force level on the electromyographic (EMG) spectral parameters and on their reliability, bipolar surface EMG measurements were performed on the lumbar muscles of 15 subjects. Two test contractions (45 s) at 40% of maximal voluntary contraction (MVC) were performed, one with 2 cm interelectrode distance and the other with 4 cm, followed by two contractions at 80% MVC with the same change in interelectrode distance. Increasing the interelectrode distance from 2 to 4 cm caused a significant mean decrease (about 8%) in the initial median frequency. It is shown that this shift is of an order of magnitude that may be expected from the bipolar electrode filter factor, and we further conclude that the observed individual variations in the shift are likely to be connected to fluctuations in the shape of the power spectrum and to variations in conduction velocity. No significant change was found for the median frequency slope when changing the interelectrode distance. Increasing the force (from 40 to 80% MVC) also caused a significant mean decrease (about 10%) in the initial median frequency. The median frequency slope became significantly more negative by more than 200%. We conclude, however, that torque fluctuations during the fatigue contractions should have had only minor influence on the standard error of measurement of the initial median frequency and of the median frequency slope. PMID- 12121687 TI - Electromyographic analysis of trapezius and sternocleidomastoideus muscles during nasal and oral inspiration in nasal- and mouth-breathing children. AB - The purpose of this study was to evaluate sternocleidomastoideus (SCM) and trapezius (superior fibers) muscle activity patterns in mouth-breathing children, and to compare them with nasal-breathing children. Forty-six children, of both sexes, ranging from 8 to 12 years old, were evaluated through electromyography. The selected children were divided into two groups; Group I, was made up of 26 mouth-breathing children and Group II of 20 nasal-breathing children. Electromyographic recordings were obtained through surface electrodes in the SCM and trapezius muscles, bilaterally, during oral and nasal inspiration. Root-mean square (RMS) data expressed in microvolts (microV), were analyzed using the Kruskall-Wallis statistical test. From the results obtained, we concluded that there was a significant difference in the muscle activity between the groups, with higher activity during nasal inspiration in the mouth-breathing group. During oral inspiration, there was no significant difference between groups. Within the groups, only the mouth-breathing group showed higher activity during nasal inspiration. PMID- 12121688 TI - Electromyography of trunk muscles in isometric graded axial rotation. AB - This study was conducted to determine the pattern, magnitude, and phasic inter relationship of the trunk muscles in maximal isometric and graded isometric axial rotational contractions and compare them with those previously observed from the same subjects in the same experimental session in dynamic conditions. In 50 normal young healthy subjects (27 male and 23 female), after a suitable skin preparation, bipolar silver-silver chloride recessed pregelled surface electrodes were placed on external oblique, internal oblique, rectus abdominis, pectoralis major, latissimus dorsi, erector spinae at T(10) and L(3) levels bilaterally with 2 cm interelectrode distance. EMG signals from grounded subjects were suitably preamplified and amplified by a fully isolated system. These subjects were stabilized in an upright-seated posture in the Axial Rotation Tester (AROT), which was placed in isometric mode for force and rotation output from the AROT. The 14 channels of EMG, the force and the rotation were sampled at 1 kHz. The subjects initially registered their isometric maximal voluntary contraction (MVC) on both sides which was used for reference and then performed their 25%, 50% and 75% of MVC bilaterally in an isometric mode in a random order. The EMG magnitude, the slope of the rise of the EMG, and the phasic interrelationship of muscles were analyzed. The results showed that female sample generated only 65% of torque of their male counterparts. There were no significant differences between the male and the female samples in the EMG variables. Exertions to the left and to the right were not significantly different from each other for the measured variables. However, the magnitude contribution of the muscles and the slope of rise of EMG were significantly different in two directions (p<0.001). The phasic interrelationship of the external obliques, the latissimus dorsi and the erector spinae were different from other muscles (p<0.01). With the increasing grades of contraction the latissimus dorsi and the external obliques increased their magnitude significantly whereas that of the erectores spinae underwent a decrease in proportionate terms (but not in absolute magnitude) suggesting their role as stabilizers but not as rotators. PMID- 12121689 TI - Influence of disposable, concentric needle electrodes on muscle enzyme and lactate serum levels. AB - Several studies addressed the question whether needle-EMG causes elevation of muscle enzymes [aspartate-aminotransferase, alanine-aminotransferase, lactate dehydrogenase, creatine-phosphokinase (CPK), isoenzyme-MB, aldolase] and lactate with conflicting results. However, these studies used sterilizable needle electrodes and different protocols and methods to record EMGs and determine muscle enzymes. This study examined if muscle enzymes are elevated immediately after or 24 h following EMGs with disposable, concentric needle-electrodes, and if they are dependent on age, sex, muscle, number of investigated sites and previous CPK-elevation. In 53 subjects, 24 woman, 29 men, aged 17-88 years, muscle enzymes were determined before, immediately after and 24 h following EMG with disposable, concentric needle-electrodes. Muscle enzymes were not different before, immediately after and 24 h following the EMG. Muscle enzymes were not different between patients 60 years of age. Apart from higher CPK in men than women, muscle enzymes were not different between the genders. Apart from CPK, muscle enzymes were not different between the brachial biceps and anterior tibial muscle. Muscle enzymes were not different if 20 sites were investigated and were independent on pre-EMG CPK-levels. In conclusion this study shows that muscle enzymes do not increase immediately or 24 h following EMG with disposable, concentric needle-electrodes, irrespective of age, gender, muscle, number of investigated sites and pre-EMG CK-levels. PMID- 12121690 TI - Factors influencing outcome of the American Board of Surgery Certifying examination: an observational study. AB - BACKGROUND: Because of the content of the American Board of Surgery (ABS) certifying (oral) examination, there is a perception that those in some subspecialty surgical training programs at the time of the examination may have a lower pass rate. In addition, the format of the oral examination has prompted the use of specialized preparation such as "mock orals" and commercial courses. The purpose of this study was to correlate the pass rates on the ABS certifying examination with the practice status and methods of specialized preparation. MATERIALS AND METHODS: A survey covering demographic information, type of surgical practice at the time of the examination, methods of preparation, and results of the examination was distributed to 1997 and 1998 graduates via a request to surgical residency program directors. RESULTS: One hundred one of 268 program directors supplied 717 names. There were 465 responses. Surveys distributed by the other 167 program directors resulted in an additional 81 responses. Four hundred ten (75%) of the respondents had taken the certifying examination. The total pass rate was 91%. There were no significant differences in the pass rate between those in private practice general surgery; those in academic general surgery; Thoracic, Vascular, or Plastic Surgery Fellows; those in other surgical fellowships; and those in the military or research. No significant differences in the pass rates were noted between those who prepared with formal mock orals, with informal mock orals, with a commercial course, with combinations of the three, and with no specialized preparation. CONCLUSION: Performance on the ABS certifying examination was not influenced by the candidate's practice status at the time of the examination. A substantial percentage of examinees either are exposed to or perceive the need to pursue specialized preparation for the examination, a behavior that in general produces good results. PMID- 12121691 TI - Uptake of indocyanine green by hepatocytes under inflow occlusion of the liver. AB - BACKGROUND: It is not clear that hepatic venous backflow actually contributes to hepatic tissue oxygenation under inflow occlusion of the liver. In order to prove that substances delivered via the hepatic vein can be utilized and/or metabolized in hepatocytes during inflow occlusion, hepatic uptake in bile and excretion of indocyanine green (ICG) were investigated in pigs. MATERIALS AND METHODS: Animals were divided into two groups: an inflow occlusion (IO) group (N = 6) and a total hepatic vascular exclusion (THVE) group (N = 3) using a bypass. One milligram of ICG per kilogram body weight was administered at the beginning of blood flow occlusion, the retention rate in the blood (ICG R) measured, and the ICG in the hepatic tissue measured by near-infrared (NIR) spectroscopy. Furthermore, the ICG concentration was measured in bile excreted by intermittent perfusion of the liver. RESULTS: ICG R declined with time in both groups; however, ICG R in the IO group decreased much faster than in the THVE group. There were significant differences between the two groups after 30 min of occlusion (P < 0.05). ICG in the hepatic tissue could be detected as a peak at 805 nm 10 min after ICG injection, and the peak became steeper with time. On the other hand, ICG was not detected at all in the hepatic tissue after 180 min in the THVE group. ICG was excreted in the bile after 60 min under IO and increased with time. On the contrary, ICG was not excreted in the bile at all under THVE. There were significant differences between the two groups after 90 min (P < 0.05). CONCLUSION: These results indicate that ICG can be extracted in hepatocytes and excreted in bile under IO of the liver. Consequently, substances such as oxygen and drugs, which are delivered via the hepatic vein, can be utilized and/or metabolized in hepatocytes under IO. PMID- 12121693 TI - Diabetic hyperglycemia: a facilitating factor in systemic capillary leak. AB - BACKGROUND: Diabetes is a known risk factor for increased morbidity and mortality following trauma and serious injuries. The current studies were designed to investigate the effects of diabetes on both local and systemic injury following mesenteric ischemia-reperfusion in rats. MATERIALS AND METHODS: Rats received streptozotocin (65 mg/kg ip) to induce diabetes or vehicle and were studied 8 weeks later. In another group of normal rats, hyperglycemia (blood sugar >200 mg/dl) was induced by intravenous infusion of a glucose solution. Rats were subjected to 10 or 30 min of superior mesenteric artery occlusion followed by 1 or 4 h of reperfusion or a sham operation. Intestinal mucosal injury, neutrophil infiltration, and changes in capillary leak were determined. Flow cytometry was used to assess neutrophil "priming state." RESULTS: Euglycemic and acutely hyperglycemic rats exhibited no mucosal injury after 10 min of ischemia, but did show significant damage after 30 min of ischemia followed by 1 h of reperfusion. Euglycemic rats had significant capillary leak following 30 min of ischemia and 4 h of reperfusion that was associated with an increase in the neutrophil priming state. In acute hyperglycemia, leak occurred after 30 min of ischemia and only 1 h of reperfusion. Diabetic rats exhibited significant mucosal injury after 10 min of ischemia and 1 h of reperfusion that was associated with significant capillary leak and increased neutrophil priming state. CONCLUSION: Altered neutrophil activity contributes to the increased susceptibility to local intestinal and systemic injury in diabetes. PMID- 12121692 TI - Hepatic vascular response to elevated intraperitoneal pressure in the rat. AB - The rat is increasingly being used to study the physiological response to elevated intra-abdominal pressure (IAP) during laparoscopic surgery. Although decreased portal venous flow associated with the elevated IAP has been reported in large animals, little information is available in rats. Furthermore, the relative blood flow changes in the hepatic artery and portal vein have not been reported. Therefore, this study was performed to elucidate the change in systemic and splanchnic circulation, including hepatic arterial and portal venular flow, during pneumoperitoneum in rats. Sprague-Dawley rats were assigned into either a ventilated or nonventilated group and then subjected to various levels of IAP (0, 5, 10, and 20 mm Hg) using carbon dioxide gas. At each pressure, both cardiac output and splanchnic organ flow were determined using fluorescent microspheres. There was no obvious hemodynamic difference between the ventilated and nonventilated groups. Mean arterial pressure and cardiac index were significantly lower with 20 mm Hg of IAP compared to 0 mm Hg in both groups. Flow to the spleen, stomach, duodenum, total intestine, and portal vein was all decreased by increasing IAP (P < 0.05 at 20 mm Hg compared to 0 mm Hg) and was significantly correlated to the decrease in cardiac index. However, the hepatic arterial flow was relatively preserved throughout all levels of IAP, suggesting activation of the hepatic arterial buffer response. We conclude that the decreased splanchnic flow during pneumoperitoneum largely depends on the decreased cardiac index. Hepatic artery flow, however, is selectively preserved and may provide protection for liver function during sustained elevations in IAP. PMID- 12121694 TI - The effect of phosphodiesterase inhibition on gallbladder motility in vitro. AB - BACKGROUND: Coffee consumption decreases the risk of gallstone disease. The proposed motility effects of caffeine, a phosphodiesterase (PDE) inhibitor, are unknown. The aim of our study was to determine the effect of caffeine and specific PDE inhibitors on gallbladder motility in vitro. METHODS: Gallbladder muscle strips from opossums were attached to force transducers. Baseline tone, electrical field stimulation (EFS)-induced nitric oxide-mediated off responses, and changes in the cholecystokinin (CCK)-8 dose-response relationship were measured. Caffeine; vinpocetine (VIN), type I PDE inhibitor; erythro-9-[2-hydroxy 3-nonyl]adenine HCl (EHNA), type II PDE inhibitor; zarderverine (ZARD), type III/IV PDE inhibitor; Ro 20-1724, type IV PDE inhibitor; and zaprinast (ZAP), type V PDE inhibitor were added to the tissue baths. RESULTS: Caffeine and all specific PDE inhibitors decreased baseline tone. Caffeine, VIN, EHNA, ZARD, Ro 20 1724, and ZAP decreased the EFS-induced off response. Caffeine (10(-5) M) and EHNA increased the CCK-8 dose-response contractions. This effect was not inhibited by atropine. Caffeine increased the tissue levels of cyclic 3',5' guanosine monophosphate but not cyclic 3',5'-adenosine monophosphate. Caffeine decreases baseline tone and the off response via type I-V PDE pathways. Caffeine also increases CCK-induced gallbladder contractions via type II PDE pathways. CONCLUSION: These effects may be a mechanism that contributes to the decreased gallstone formation with caffeine consumption. PMID- 12121695 TI - Very late survival after vascular surgery. AB - PURPOSE: The aim of this study was to define very late survival in veterans who routinely underwent preoperative assessment of left ventricular function using radionuclide ventriculography (RNVG) before elective major vascular surgery from 7/84 to 7/88 at one Veterans Affairs Medical Center. METHODS: RNVG defined left ventricular ejection fraction (EF) and determined the presence of ventricular wall motion abnormalities. Patients undergoing elective vascular surgery (n = 310) who had preoperative RNVG were then followed over the years using direct contact, VA administrative databases, and, most recently, the Social Security Death Index. RESULTS: Follow-up was 6.64 +/- 4.62 years (range 0 to 16.2 years). Current survival is 10% (11/107) after carotid surgery, 12% (10/82) after aortic aneurysm repair, 15% (17/111) after extremity reconstruction, and 0% (0/10) after visceral artery reconstruction (ns). There was no statistically significant difference in mortality between the different types of vascular surgery at 30 days or at 1, 5, and 10 years after surgery (ns). Actual survival rates at 5 years after carotid surgery, aneurysm repair, extremity reconstruction, and visceral reconstruction were 55, 61, 59, and 50%, respectively. Stepwise logistic regression analysis was performed which included preoperatively defined cardiovascular risk factors, type of surgery, and results of RNVG. The final regression model indicated that age, diabetes, smoking at the time of surgery, and low EF were independently associated with overall mortality while angina, prior myocardial infarction (MI), and type of operation were not. Mean survival duration with normal EF (>50%) was 7.99 years versus 4.78 years with low EF (P < 0.001). No patient with severe left ventricular dysfunction (EF < or = 35%; n = 39) or who had postoperative cardiac complications (MI, CHF, ventricular arrhythmia; n = 38) survived to the present. CONCLUSIONS: Very late survival after major vascular surgery was related to the presence of diabetes, active smoking at the time of surgery, left ventricular function, and postoperative cardiac complications. Since there was no association of overall mortality with angina or prior MI, an aggressive approach to coronary evaluation in such patients might not alter very late survival. PMID- 12121696 TI - Laparoscopic ventral hernia repair: an initial institutional experience. AB - BACKGROUND: Ventral and incisional hernias remain a problem for surgeons with reported recurrence rates of 25-50% for open repairs. Laparoscopic approaches offer several theoretical advantages over open repairs. MATERIALS AND METHODS: All patients undergoing a laparoscopic ventral hernia repair from April to December 2000 were prospectively entered in a database. Patients underwent repair with expanded polytetrafluoroethylene dual mesh. Full-thickness abdominal wall nonabsorbable sutures and 5-mm tacks were placed circumferentially. RESULTS: Of 32 patients, 15 underwent incisional repair, 13 had repair of a recurrent incisional hernia, and 4 had repair of a primary abdominal wall defect. Two procedures [2/32; 6.3%] were converted to open, one for loss of abdominal domain and one for neovascularization due to cirrhosis. There were two early recurrences [2/30; 6.7%]. Both of these failures occurred in patients with hernia defects extending to the inguinal ligament, preventing placement of full-thickness abdominal wall sutures inferiorly. Average operating time was 128 +/- 42 min (range 37-225 min). Average length of stay was 1.8 days [range 0-7 days]. There were no transfusion requirements or wound infections. One patient underwent a small bowel resection after completion of repair. One patient required drainage of a seroma 4 weeks after the procedure. CONCLUSIONS: Laparoscopic ventral hernia repair can be safely performed with an acceptable early recurrence rate, operative time, length of stay, and morbidity. Securing the mesh with full thickness abdominal wall sutures in at least four quadrants remains a key factor in preventing early recurrence. PMID- 12121697 TI - CT scan in the management of acute appendicitis. AB - BACKGROUND: Recent studies demonstrate a 98% accuracy of a CT scan in the diagnosis of acute appendicitis. We aimed to determine the accuracy and clinical value of CT scans in patients suspected of having acute appendicitis. PATIENTS AND MATERIALS: We reviewed outcomes of 125 patients over a 5-month period who had CT scans for the initial diagnosis of acute appendicitis. CT scan interpretations were correlated with surgical and pathologic findings. Follow-up was attempted in all patients who did not undergo appendectomy. RESULTS: CT scans and clinical courses were complete in 110 patients (88%); 14 patients were lost to follow-up and 1 was excluded. One patient had two CT scans. Thus, there were 111 CT scans available for review. Radiologic interpretation of these CT scans yielded 36 positive (33%), 67 negative (60%), and 8 indeterminate (7%), resulting in a sensitivity of 90%, a specificity of 89%, a PPV of 78%, and a NPV of 96%. CONCLUSIONS: CT scan may be useful in the diagnosis of acute appendicitis, but the reported high accuracy rate was not reproduced at our institution. CT scan was not clinically useful in 21% of patients. We conclude that a CT scan may be beneficial in the diagnosis of appendicitis with selected patients who have equivocal findings. Thus, at our institution, the accuracy of a CT scan does not justify its routine use in patients with clinical findings of appendicitis. PMID- 12121698 TI - Role of L-selectin in the development of ventilator-associated pneumonia in patients after major surgery. AB - BACKGROUND: The circulating level of soluble L-selectin (sL-selectin) has been reported to be low in adult respiratory distress syndrome and acute lung injury. This study explores the role of L-selectin in the development of ventilator associated pneumonia (VAP) in patients undergoing major surgery. PATIENTS AND METHODS: Thirty-four patients who underwent esophagectomy were maintained by mechanical ventilation in a surgical intensive care unit. Fourteen patients developed VAP by postoperative day (POD) 7, while 20 patients did not. The plasma levels of soluble adhesion molecules and elastase were measured serially by ELISA or EIA. The expression of L-selectin on polymorphonuclear neutrophils (PMNs) was analyzed by flow cytometry. RESULTS: In multiple logistic regression analysis, only the preoperative plasma level of sL-selectin was significantly associated with VAP. The plasma sL-selectin level before surgery was significantly lower in the patients who developed VAP compared with the patients who did not develop VAP. After surgery, the level of sL-selectin did not change. The plasma level of soluble intercellular adhesion molecule-1 increased in the patients with and without VAP. The plasma level of soluble vascular cell adhesion molecule-1 was significantly higher in the patients with VAP. L-selectin expression on PMNs showed a peak on POD 2 in the patients without VAP, whereas it was impaired in the patients with VAP. CONCLUSIONS: Determination of the preoperative plasma level of sL-selectin may help to identify patients at high risk for VAP after esophagectomy. PMID- 12121699 TI - Experimental monitoring of hepatic glucose, lactate, and glutamate metabolism by microdialysis during surgical preparation of the liver hilus. AB - Mechanical liver manipulation can lead to hepatic microcirculation (MC) impairment. The pathobiochemical relevance of this phenomenon is not fully understood. Microdialysis (MD) allows a quantification of metabolic products in interstitial fluid, thus enabling analysis of the hepatic metabolic state during changes of liver perfusion. The aim of the study was to quantify the functional effects of standardized surgical liver preparation both on liver metabolism and microperfusion. Two groups of animals (pigs, n = 25) were formed: In the trial group (TG; n = 13) the liver was mobilized, followed by hilar preparation. In the control group (CG; n = 12) mobilization of the liver without hilar dissection was performed. Surgical manipulation was followed by an observation in both groups. Hepatic interstitial glucose, lactate, and glutamate concentrations were detected by MD and liver MC by thermodiffusion. During liver mobilization MC decreased significantly in both groups (TG; 86.7 +/- 2.0 to 73.4 +/- 2.3 ml/100 g min; and CG; 88.3 +/- 3.1 to 71.9 +/- 2.2 ml/100 g/min). In the trial group levels decreased further during hilar preparation reaching minimal values of 65.6 +/- 2.8. After preparation MC recovered to baseline. Glucose, lactate, and glutamate concentrations increased significantly during liver mobilization in the trial (glucose; 0.52 +/- 0.13 to 0.88 +/- 0.19 mmol/L; lactate; 0.34 +/- 0.07 to 0.54 +/- 0.07 mmol/L; glutamate; 34.5 +/- 3.6 to 52.6 +/- 8.0 micromol/L) and control group (glucose; 0.58 +/- 0.06 to 0.95 +/- 0.13 mmol/L; lactate; 0.30 +/- 0.06 to 0.49 +/- 0.07 mmol/L; glutamate; 32.9 +/- 2.36 to 56.1 +/- 5.12 micromol/L). Throughout hilus preparation maximum values could be measured in TG (glucose; 1.69 +/- 0.34; lactate; 0.90 +/- 0.18; glutamate; 63.5 +/- 7.2). After termination of mobilization or preparation baseline concentrations were reached again. MD allows monitoring of metabolic changes in hepatic parenchyma. Surgical liver preparation leads to changes of intrahepatic glucose, lactate, and glutamate levels (without alterations of parameters in systemic plasma) along with hepatic MC impairment. Reconstitution of hepatic MC was accompanied by rapid normalization of metabolic parameters. By measuring specific parameters, MD could prove to be of use for functional assessment of metabolic effects due to MC disturbances. PMID- 12121700 TI - Coordinate regulation of secretory stress proteins (PSP/reg, PAP I, PAP II, and PAP III) in the rat exocrine pancreas during experimental acute pancreatitis. AB - BACKGROUND: Pancreatic stone protein (PSP/reg) is a constitutively secreted protein in pancreatic juice. Pancreatitis-associated protein (PAP) belongs to the same family of proteins. PAP is highly increased during acute pancreatitis, while no exact data exist regarding PSP/reg protein synthesis and secretion. Recently, an attempt to determine PSP/reg and PAP levels in sera of rats with acute pancreatitis showed a significant increase in PAP but failed to demonstrate changes in PSP/reg. Others reported that surgical manipulation of the pancreas, including sham controls, affected mRNA levels of PSP/reg. Neither report determined protein levels of PSP/reg. METHODS: Rats were treated intraperitoneally with a supramaximal dose of caerulein to induce pancreatitis, a physiological dose of caerulein, or a saline injection. Pancreata were analyzed for PAP and PSP/reg using ELISAs. RNA was extracted for Northern blot analysis of PAP I, II, and III and PSP/reg mRNA. RESULTS: Experimental induction of acute pancreatitis caused a coordinate increase in both PSP/reg and PAP. PAP showed an acute response and returned to low levels within 48 h while PSP/reg exhibited a more sustained response. Intraperitoneal application of a physiological dose of caerulein and even a saline injection caused an increase in PSP/reg. CONCLUSION: PSP/reg and PAP levels are increased through similar mechanisms by physiological and supramaximal doses of caerulein. However, PSP/reg regulation appears to sustain high levels while PAP levels are more transient. Since the regulation of this protein family is affected even under mild stress, we define them as secretory stress proteins. PMID- 12121701 TI - IL-10 is not protective in intestinal ischemia reperfusion injury. AB - BACKGROUND: Ischemia/reperfusion of the small intestine disrupts gut barrier function, increases bacterial translocation, and activates systemic pro inflammatory responses. Pharmacological treatment with the anti-inflammatory cytokine interleukin-10 (IL-10) following ischemia to muscle reduces the severity of local and systemic inflammation. While endogenous IL-10 is protective in murine models of acute endotoxemia, its physiological role during direct gut injury is unknown. PATIENTS AND MATERIALS: Mice genetically deficient in IL-10 (IL-10(-/-)) and their normal littermates (IL-10(+/+)) underwent 20 to 50 min of gut ischemia by occlusion of the superior mesenteric artery. RESULTS: Both short- and long-term (>16 h) survival after reperfusion of IL-10(-/-) mice was identical to that of the wild-type littermates, with 50% mortality observed at 35 min of occlusion. The small bowel demonstrated discrete gross areas of hemorrhage and ischemia localized to the jejunum. No significant difference in the extent or time for occurrence of macroscopic or microscopic intestinal damage to the small bowel was observed in IL-10(-/-) or IL-10(+/+) mice, despite the marked elevation in serum IL-6. CONCLUSIONS: The absolute serum concentration of IL-6 in the presence or the absence of IL-10 does not affect local or systemic response to ischemic intestinal injury. These results also demonstrate that the anti inflammatory cytokine IL-10 does not play a significant local or systemic protective role in this model of ischemia/reperfusion. PMID- 12121702 TI - Ischemic preconditioning prior to aortic cross-clamping protects high-energy phosphate levels, glucose uptake, and myocyte contractility. AB - BACKGROUND: We hypothesized that indices of myocyte contractility and metabolism could be preserved with ischemic preconditioning in a model of reversible ischemia similar to that occurring during routine cardiac surgery. Regional measures of metabolism and function have not been studied in conjunction with individual myocyte function during postischemic recovery of preconditioned myocardium. PATIENTS AND METHODS: In 16 dogs, myocardium supplied by the left anterior descending artery (LAD) was preconditioned with intermittent LAD ischemia and reperfusion. Following preconditioning, the heart was made globally ischemic for 20 min at normothermia by aortic cross-clamping while on cardiopulmonary bypass. In 10 animals, serial measurements of LAD and remote region adenosine triphosphate (ATP) levels, glucose uptake, and wall thickening were obtained with full-thickness drill biopsies, positron emission tomography (PET), and 2-D echocardiography, respectively. In the remaining 6 animals, cardiac myocytes were isolated after 1 h of reperfusion for measurement of myocyte contractility and intracellular calcium transients. RESULTS: ATP levels were higher in the preconditioned LAD region than in the remote region at end of ischemia (3.17 +/- 0.33 nmol/mg vs 2.59 +/- 0.30 nmol/mg, P = 0.006). Similarly, preconditioned region glucose uptake was 40% higher than remote region glucose uptake at 2 days postischemia (0.35 +/- 0.06 micromol/min/g vs 0.25 +/- 0.05 micromol/min/g, P = 0.019). There were no differences in regional wall thickening as measured by 2-D echo either immediately following ischemia or at 2 days. Individual myocyte contractile response to increasing concentrations of extracellular calcium was preserved in cells from preconditioned myocardium, but it was severely depressed in remote region myocytes. CONCLUSIONS: We conclude that regional ischemic preconditioning prior to prolonged ischemia protects myocardial glucose uptake and myocyte contractile function. The beneficial effects on glucose metabolism suggest that preconditioning may have sustained protective effects on cell metabolism. PMID- 12121703 TI - A novel implantable collagen gel assay for fibroblast traction and proliferation during wound healing. AB - BACKGROUND: A novel implantable assay for studying cellular behavior in the wound environment was developed. The assay is unique in that it combines the more quantitative nature of in vitro assays with the greater physiological relevance of in vivo wound healing models. MATERIALS AND METHODS: Cells were seeded in a physiologically relevant biological matrix, a collagen gel, contained within a semipermeable tube, and then exposed to soluble factors of the wound environment at different stages of the wound healing response. Gels were harvested at prescribed time points, and cell proliferation rates and gel compaction were measured. These data were combined with our theory for cell-matrix mechanical interactions to estimate the cell traction exerted by the cells leading to gel compaction. Cell morphology and alpha-smooth muscle actin expression were also characterized. RESULTS: The proliferation of and traction exerted by fibroblasts exposed to the soluble wound environment were different from those in similar collagen gels maintained in culture in complete medium. Proliferation and traction also varied over the course of the wound healing response. Traction was higher and proliferation lower in day 1-5 wounds compared to day 7-11 wounds. Recovered cells no longer stained for alpha-smooth muscle actin, in contrast to cells maintained in culture. CONCLUSIONS: Changes in the soluble wound environment that occur as the wound healing response proceeds alter fibroblast traction and migration. We have developed a new assay that employs a physiologically relevant biological matrix and allows the effects of the dynamic soluble wound environment on cellular traction, proliferation, and other phenomena such as protein expression to be quantified. PMID- 12121704 TI - Comparison of Celsior and UW solution in experimental pancreas preservation. AB - BACKGROUND: The University of Wisconsin solution (UW) is the gold standard for pancreas preservation. Celsior (CEL) was formulated specifically for heart preservation. Recently, experimental and clinical experience has been reported on the application of CEL to abdominal organs. In this animal study, pancreas preservation with CEL was compared with that in UW solution. PATIENTS AND MATERIALS: Heterotopic, allogeneic pancreaticoduodenal transplantation was performed in female Gottingen Minipigs (n = 12 donors, n = 12 recipients). The grafts were flushed and stored for 6 h at 4 degrees C in UW or CEL. The recipients were randomized into two groups receiving either UW (n = 6)- or CEL (n = 6)-preserved grafts with a follow-up of 5 days. Blood flow (laser Doppler), partial oxygen tension, histological changes, endothelin-1 (plasma, immunohistochemistry), lipase, amylase, trypsinogen activation peptide, and C reactive protein (CRP) were measured. RESULTS: Partial oxygen tension was lower in the CEL group (P < 0.05). However, blood flow did not differ between UW- and CEL-preserved organs. The histomorphologic analysis of the pancreatic grafts revealed significantly less edema in the UW-preserved organs. Serum levels of amylase, lipase, CRP, and TAP taken from the central venous blood were comparable in the two groups, except for higher amylase values 36 h after reperfusion in the CEL group compared to the UW group (P < 0.05). Likewise, TAP taken from the portal venous effluent of the graft was found to be higher in the CEL group than in UW (P < 0.05). Endothelin-1 serum levels rose significantly during reperfusion without differences between the two groups. ET-1 immunohistochemistry revealed increased local ET-1 during reperfusion in all grafts. However, the ET-1 immunostaining in the CEL group was more pronounced than that in the UW group (P < 0.05). CONCLUSIONS: Our results suggest that CEL solution is not as effective in preventing pancreatic ischemia/reperfusion damage as the standard UW solution in experimental pancreas transplantation. Increased ET-1 immunostaining and reduced p(ti)O(2) in the CEL group indicate increased microcirculatory damage in the CEL group. PMID- 12121705 TI - Oral hypoglycemic sulfonylurea glimepiride preserves the myoprotective effects of ischemic preconditioning. AB - BACKGROUND: To investigate whether the sulfonylurea glimepiride affects the myoprotective effects of ischemic preconditioning (IPC), isolated rabbit hearts were perfused with Krebs-Henseleit solution. METHODS: Eight hearts underwent IPC consisting of two cycles of 5 min global ischemia and reperfusion. Six hearts received a 5-min infusion of 10 microM glimepiride, six hearts received a 5-min infusion of 50 microM glimepiride, and seven hearts received a 5-min infusion of 10 microM glibenclamide before IPC. Seven hearts received a 5-min infusion of the selective mitochondrial K(ATP) channel opener diazoxide (50 microM). Other hearts received a 5-min infusion of 10 microM glimepiride (n = 6), 50 microM glimepiride (n = 6), or 10 microM glibenclamide (n = 7) before diazoxide. Seven hearts served as a control. All groups then were subjected to 1 h of regional ischemia, followed by 1 h of reperfusion. LV pressures, monophasic action potential duration (APD(50)), and infarct size were measured. RESULTS: Both IPC and diazoxide significantly prolonged APD(50) and preserved diastolic function at 60 min of reperfusion compared to control. In addition, both groups reduced infarct size compared to control. Glibenclamide, but not glimepiride reversed these effects. CONCLUSION: Glimepiride offers less cardiovascular effects than glibenclamide, possibly due to its lower affinity for the mitochondrial K(ATP) channels. PMID- 12121706 TI - Cold storage sensitizes rat femoral artery to an endotoxin-induced decrease in endothelium-dependent relaxation. AB - Cold-stored arteries, tissues or organs are transferred in vascular, reconstructive and transplantation surgery. The function of transferred vessels and tissues diminishes when infection complicates transplantation, thereby contributing to morbidity. To evaluate the mechanisms involved, the effects of cold storage on basal vascular reactivity and the sensitivity to the vascular effects of endotoxin were tested in isolated rat femoral artery segments. A crossover design was followed, so that prior to cold storage 4 vessels were incubated for 2 h at 37 degrees C with endotoxin (Escherichia coli 0127:B8, 50 microg mL(-1)) in Krebs solution and 4 with Krebs solution only, while, after cold storage, segments from the former vessels were incubated with Krebs solution only and segments from the latter with endotoxin in Krebs solution. Vascular reactivity was tested in a wire myograph by the addition of depolarizing 125 mM KCl or norepinephrine (NE) as well as the endothelium-dependent vasodilator acetylcholine (ACh) and endothelium-independent vasodilator sodium nitroprusside (SNP). Cold storage did not affect vascular reactivity in the absence of endotoxin. Endotoxin decreased maximum response to NE prior to storage and sensitivity to SNP prior to and after cold storage. After cold storage, endotoxin decreased relaxation to ACh and increased vasoconstriction in response to KCl and NE (P < 0.05). We conclude that cold storage does not alter endothelial and smooth muscle function but sensitizes rat femoral artery to an endotoxin-induced decrease in endothelium-dependent relaxation and thereby to an increase in vasoconstrictor responses, whereas endotoxin alone only decreases receptor dependent vasoconstrictor responses and sensitivity to NO donors. This may explain in part the detrimental effect of infection on function of cold-stored arterial grafts and tissue/organ transfers. PMID- 12121707 TI - Growth hormone increases circulating neutrophil activation and provokes lung microvascular injury in septic peritonitis rats. AB - BACKGROUND: Growth hormone (GH) has been shown to increase mortality in critical illness, illustrating the need for better understanding of GH treatment. The neutrophil is a key mediator in producing organ injury following shock and trauma and is regulated by GH. Therefore, the purpose of the present study was to examine the effects of GH on circulating neutrophil activation and subsequent lung injury induced by sepsis in rats. MATERIALS AND METHODS: Sepsis was induced in male Wistar rats via cecal ligation and puncture (CLP). Recombinant human growth hormone (1 IU/kg) was given subcutaneously right after CLP with an additional injection at 12 h after CLP. CD11b expression and an oxidative burst of neutrophils were detected using flow cytometric analysis. Lung myeloperoxidase activity was determined as an index of tissue neutrophil accumulation. Lung microvascular injury was assessed by quantitating extravasation of Evan's blue dye into lung parenchyma. RESULTS: Growth hormone significantly increased sepsis induced expression of CD11b and sepsis-induced circulating neutrophil activation. Also growth hormone increased neutrophil accumulation in lungs induced by sepsis. Lung microvascular injury was aggravated by growth hormone treatment in septic rats. CONCLUSIONS: It is worthwhile to rethink GH administration in critical illness and further studies are required to determine the safety and clinical benefits of GH administration in critical illness. PMID- 12121708 TI - Immobilization of human thrombomodulin to expanded polytetrafluoroethylene. AB - BACKGROUND: The success of synthetic grafts for vascular reconstruction remains limited by thrombosis and intimal hyperplasia. In addition to the well-described antithrombotic effects of thrombomodulin, we have demonstrated that recombinant human thrombomodulin (rTM) inhibits arterial smooth muscle cell proliferation induced by thrombin. This study investigated the binding of functional rTM to expanded polytetrafluoroethylene (ePTFE). METHODS: Immobilization of rTM was achieved by either (1) a direct coating or (2) a two-step binding process using a water-soluble condensing cross-reaction agent EDAC to modify the ePTFE surface followed by binding of rTM. The samples were then subjected to a tangential shaken wash. The evidence of bound rTM was evaluated by both morphologic and functional studies. RESULTS: SEM, BSI, and X-ray microanalysis identified that the two-step binding method resulted in significantly greater binding of rTM molecules to ePTFE pre- and post a 7-h wash than the direct coating method. With the two-step binding method rTM ranging from 0.25 to 12.5 microg immobilized to ePTFE-activated protein C (APC) in a concentration-dependent manner by more than 6000-fold compared to the buffer control (P < 0.04) and 50-85% more than direct coating (P < 0.004). With direct coating, the level of APC dropped significantly to near 40% of the preshaken level at 2 h and diminished to 26% at 7 h. Whereas, the level of APC with the two-step binding stabilized at 51 and 49% after being shaken 2 and 7 h, respectively. CONCLUSION: Functional rTM binding to ePTFE was significantly improved with a new two-step binding method. PMID- 12121709 TI - Arterial embolization hyperthermia in porcine renal tissue. AB - BACKGROUND: Arterial embolization hyperthermia (AEH) consists of arterially embolizing tumors with ferromagnetic particles that generate hysteretic heating on exposure to an alternating magnetic field. It was the objective of this study to evaluate AEH using the kidney of a large animal as a tumor model. METHODS: Between 50 and 400 mg of ferromagnetic microspheres (32 microm in diameter) was arterially infused into the kidneys of three pigs. Temperature probes were inserted into renal tissue, skin, and subcutaneous fat. Each subject was then exposed to an alternating magnetic field for 5 min, while under a general anesthetic. A femoral artery catheter was used to monitor the cardiac pulse. Three days after treatment the renal tissue was chemically analyzed for iron content, which was then correlated with tissue heating rates. RESULTS: There was a linear relationship between heating rate and iron concentration (N = 18, correlation = 0.72, P < 0.001) that suggested tissue iron concentrations in the range of 1.55 to 4.05 mg/g would yield tissue heating rates of 0.5 to 1.0 degrees C/min. No temperature increases were detected in control renal tissue (N = 6). The median increase in skin temperature after 5 min of heating was 0.8 degrees C (N = 6, min = 0.7 degrees C, max = 1.3 degrees C), and that in subcutaneous fat was 1.1 degrees C (N = 6, min = 0.8 degrees C, max = 1.2 degrees C). There was no detectable stimulation of cardiac or skeletal muscle or peripheral nerves during treatment. All subjects had uneventful 3-day posttreatment survivals. CONCLUSION: This study has shown that AEH can target deep-seated, vascularized tissue in a large animal with therapeutic temperatures (> 42 degrees C), and that the treatment is safe and well tolerated. Further assessment of treatment schedules should allow for a human trial in the near future. PMID- 12121710 TI - Effects of estrogen, progesterone, and combination exposure on interleukin-1 beta induced expression of VCAM-1, ICAM-1, PECAM, and E-selectin by human female iliac artery endothelial cells. AB - The cardioprotective effect of hormone replacement therapy (HRT) in healthy, postmenopausal women is well documented. Little work has been performed on the effect of HRT in peripheral arteries. Recent work suggests that HRT may adversely affect the patency of peripheral grafts. This study investigates the in vitro effects of estrogen and/or progesterone exposure on adhesion molecule expression by normal human female iliac artery endothelial cells (HIAEC). Control and interleukin-1 beta (IL-1 beta)-stimulated HIAEC monolayers were labeled with fluorescent-tagged antibodies against the adhesion molecules VCAM-1, ICAM-1, PECAM, and E-selectin. FACS analysis was used to measure antibody-labeled adhesion molecule expression. ICAM-1 and PECAM were found to be constitutively expressed. VCAM-1 and ICAM-1 expression were significantly upregulated by IL-1 beta, while E-selectin was neither constitutively expressed nor upregulated by IL 1 beta. Pretreatment with estrogen or progesterone alone decreased IL-1 beta stimulated VCAM-1 and ICAM-1 expression, but not to statistically significant levels. Combined hormone exposure significantly decreased, in an additive fashion, the expression of VCAM-1 and ICAM-1 in stimulated cells. This study supports extension of the beneficial effects ascribed to HRT to include the peripheral arterial endothelium of healthy women. PMID- 12121711 TI - Effective treatment of gut barrier dysfunction using an antioxidant, a PAF inhibitor, and monoclonal antibodies against the adhesion molecule PECAM-1. AB - BACKGROUND: Oxygen free radicals (OFRs), platelet activating factor (PAF), cell adhesion molecules, and transmigration of polymorphonuclear leukocytes through the gut barrier are probably all essential in the development of gut barrier dysfunction following intestinal ischemia and reperfusion (I/R). Pretreatment and early treatment of I/R with the OFRs-scavenger (NAC), the PAF inhibitor lexipafant, and monoclonal antibodies against the adhesion molecule PECAM-1 (anti PECAM-1-Mab) have been reported to be effective in the prevention or recovery of gut barrier dysfunction and result in a decrease in cytokine levels. Less is known about the effect of treatment inserted during the late stage of I/R. The objective of this study was to evaluate the potential therapeutic value of single or combination therapy with NAC, lexipafant, and anti-PECAM-1-MAb administered late during intestinal I/R in the rat. METHODS: NAC, lexipafant, and anti-PECAM-1 MAb were administrated, alone or in combination, after 3 h of reperfusion following 40 min of superior mesenteric arterial ischemia in the rat. Intestinal endothelial and epithelial barrier permeability, myeloperoxidase (MPO) activity, interleukin-1 beta (IL-1 beta), and protease inhibitor levels were evaluated after 12 h of reperfusion. RESULTS: Intestinal endothelial and epithelial permeability significantly increased in rats with I/R and saline treatment. Proteolytic activity in plasma was indicated by low levels of the three measured plasma protease inhibitors. Intestinal mucosal MPO content increased significantly. These changes were, to different degrees, reduced by late inserted treatment with NAC, lexipafant, or anti-PECAM-1-MAb. Alterations in systemic levels of IL-1 beta paralleled the changes found in gut barrier permeability and leukocyte trapping. Systemic antithrombin III levels and increased barrier permeability in remote organs were partly restored, especially by multimodal therapy. CONCLUSION: Treatment with NAC, lexipafant, and/or monoclonal antibodies against PECAM-1, inserted at a later stage of I/R, reduced the severity of I/R associated intestinal dysfunction and decreased the systemic concentrations of IL 1 beta, local leukocyte recruitment (MPO), and partly restored plasma protease inhibitor levels. PMID- 12121712 TI - Antiproteolytic action of insulin in burn-injured rats. AB - BACKGROUND: Negative nitrogen balance is a typical metabolic response to burn injury resulting in decreased muscle mass and activity. Since insulin is an anabolic hormone, using insulin as a prophylactic agent in burned patients has received some attention. The present study was carried out to investigate the systemic effect of insulin on burn injury-induced muscle wasting. PATIENTS AND METHODS: A 15-20% total body surface area (TBSA) scald burn injury was inflicted on the shaved dorsum of rats. Rats were treated with a daily subcutaneous insulin injection for 3 days (0.25-1.0 U/day). After the treatment, a variety of insulin dependent physiological parameters were monitored. Overall body protein degradation rates were determined by measuring the urinary tyrosine. Also, protein degradations were measured in diaphragm muscles, splenocytes, and peripheral blood mononuclear cells to directly confirm the antiproteolytic activity of insulin. RESULTS: Administration of insulin to burn-injured rats restored body weight primarily by reducing accelerated protein degradation and regaining the intracellular protein content in individual skeletal muscle. The measured physiological parameters showed no possible side effects. Protein degradation in immune cells was also suppressed after the therapy. CONCLUSION: Results indicate that lower dose insulin particularly suppresses protein degradation without causing secondary effects. It may be a useful approach to preventing burn injury-induced muscle wasting and also has a potential to improve immune response. PMID- 12121713 TI - Hepatic ischemia-reperfusion promotes liver metastasis of colon cancer. AB - BACKGROUND: The effects of hepatic ischemia/reperfusion (I/R) on liver metastasis have not been fully examined. We examined hepatic I/R and liver metastasis of colorectal cancer in a rat model; we also quantitated expression of E-selectin (ELAM-1) mRNA after I/R. MATERIALS AND METHODS: Rats underwent 30 or 60 min of 70% partial hepatic ischemia. After 60 min of reperfusion, rat colon adenocarcinoma cells (RCN-H4) were inoculated intrasplenically. The number of tumor nodules on the liver surface was determined 3 weeks later. Expression of E selectin mRNA was determined at 1, 3, and 6 h after ischemia by quantitative RT PCR. RESULTS: Hepatic I/R promoted liver metastasis of RCN-H4 and induced the expression of E-selectin mRNA in both clamped and unclamped liver lobes. The number of tumor nodules and the expression of E-selectin mRNA after 60 min of ischemia was greater than that after 30 min. CONCLUSIONS: Hepatic I/R, especially with a long duration of ischemia, induces expression of E-selectin and promotes liver metastasis of colon cancer in rats. PMID- 12121718 TI - Central role of peroxisomes in isoprenoid biosynthesis. AB - Peroxisomes contain enzymes catalyzing a number of indispensable metabolic functions mainly related to lipid metabolism. The importance of peroxisomes in man is stressed by the existence of genetic disorders in which the biogenesis of the organelle is defective, leading to complex developmental and metabolic phenotypes. The purpose of this review is to emphasize some of the recent findings related to the localization of cholesterol biosynthetic enzymes in peroxisomes and to discuss the impairment of cholesterol biosynthesis in peroxisomal deficiency diseases. PMID- 12121719 TI - A lipid based depot (DepoFoam technology) for sustained release drug delivery. AB - Encapsulation of drugs into multivesicular liposomes (DepoFoam) offers a novel approach to sustained-release drug delivery. While encapsulation of drugs into unilamellar and multilamellar liposomes, and complexation of drugs with lipids, resulted in products with better performance over a period lasting several hours to a few days after intravascular administration, DepoFoam-encapsulation has been shown to result in sustained-release lasting over several days to weeks after non vascular administration. The routes of administration most viable for delivery of drugs via DepoFoam formulations include intrathecal, epidural, subcutaneous, intramuscular, intra-atricular, and intraocular. DepoFoam particles are distinguished structurally from unilamellar vesicles, multilamellar vesicles, and neosomes in that each particle comprises a set of closely packed non-concentric vesicles. The particles are tens of microns in diameter and have large trapped volume, thereby affording delivery of large quantities of drugs in the encapsulated form in a small volume of injection. A number of methods based on a manipulation of the lipid and aqueous composition can be used to control the rate of sustained-release from a few days to several weeks. PMID- 12121720 TI - Multi-subunit acetyl-CoA carboxylases. AB - Acetyl-CoA carboxylase (ACC) catalyses the first committed step of fatty acid synthesis, the carboxylation of acetyl-CoA to malonyl-CoA. Two physically distinct types of enzymes are found in nature. Bacterial and most plant chloroplasts contain a multi-subunit ACC (MS-ACC) enzyme that is readily dissociated into its component proteins. Mammals, fungi, and plant cytosols contain the second type of ACC, a single large multifunctional polypeptide. This review will focus on the structures, regulation, and enzymatic mechanisms of the bacterial and plant MS-ACCs. PMID- 12121721 TI - Storage (irritative) and voiding (obstructive) symptoms as predictors of benign prostatic hyperplasia progression and related outcomes. AB - OBJECTIVES: To assess the utility of voiding and filling symptom subscores in predicting features of benign prostatic hyperplasia (BPH) progression, including acute urinary retention (AUR) and prostate surgery. METHODS: The Proscar Long term Efficacy and Safety Study (PLESS) was a 4-year study designed to evaluate the effects of finasteride versus placebo in men with lower urinary tract symptoms (LUTS), clinical evidence of BPH, and no evidence of prostate cancer. A self-administered questionnaire was employed to quantify LUTS at baseline. Receiver operating characteristics (ROC) curves were used to assess baseline characteristics from patients treated with placebo as predictors of outcomes. The characteristics assessed included the overall symptom score (Quasi-AUA SI), separate voiding and filling subscores, prostate volume (PV) and serum prostate specific antigen (PSA) levels. RESULTS: PV and PSA were superior to the symptom scores at predicting episodes of spontaneous AUR and all types of AUR. The Quasi AUA SI and the filling and voiding subscores were effective at predicting progression to surgery; however, PSA was more effective at predicting this outcome. To better evaluate symptoms as predictors of surgery, patients who experienced a preceding episode of AUR were excluded from the surgery analysis. In the absence of preceding AUR, the best predictors of future surgery were the Quasi-AUA SI and the filling subscore. CONCLUSIONS: Among men with LUTS, clinical BPH and no history of AUR, the overall symptom score and storage subscore are useful parameters to aid clinicians in identifying patients at risk for future prostate surgery. PV and PSA were the best predictors of AUR, while PSA was the best predictor of prostate surgery (for all indications). PMID- 12121722 TI - Is the higher prevalence of benign prostatic hyperplasia related to lower urinary tract symptoms in Korean men due to a high transition zone index? AB - OBJECTIVE: Asian men generally have smaller prostate gland sizes than their Western counterparts. Nonetheless, the prevalence of lower urinary tract symptoms (LUTS) is similar between native Asian men and men in the Western hemisphere. The purpose of this study was to determine if the enlargement of the transition zone volume (TZV) relative to the overall prostate volume (PV) might account for the prevalence of LUTS among Korean men despite having "smaller" prostates. METHODS: Three hundred and seventy consecutive age-matched men (94 Caucasian, 94 Hispanic, 93 African-American, and 89 Korean) with LUTS were evaluated utilizing the International Prostate Symptom Score (IPSS), peak flow rate (Q(max)), serum PSA and transrectal ultrasound (TRUS). The ratio of TZV to total PV was used to determine the transition zone index (TZI). RESULTS: Mean baseline IPSS and Q(max) were significantly different (p<0.001 and p<0.03) for Korean men (19.9+/-7.6 and 11.3+/-4.2) in comparison to African-American (14.6+/-3.7, 12.6+/-4.1), Caucasian (13.4+/-4.3, 12.5+/-3.8), and Hispanic (13.9+/-3.1, 11.9+/-4.5) men. Regardless of race, TZI correlated with IPSS (r=0.31, p<0.01) and Q(max) (r=0.26, p<0.04). Mean TZI was significantly (p<0.001) higher in Korean and African-American men (0.45+/-0.08 and 0.44+/-0.05, respectively) than Caucasian and Hispanic men (0.39+/-0.03 and 0.38+/-0.02, respectively). CONCLUSIONS: Among age-matched, ethnically diverse men with moderate to severe LUTS, Korean men demonstrated more clinical symptoms and a higher ratio of transition zone enlargement relative to total prostate in comparison to Caucasian, Hispanic, and African-American men. The clinical significance of these findings remains to be determined. PMID- 12121723 TI - Gleason score predicts androgen independent progression after androgen deprivation therapy. AB - OBJECTIVE: The Gleason system is the most widely utilized histologic grading system for prostate cancer and a powerful predictor of cancer behavior. In this study, we evaluated the prognostic value of the Gleason grading system in predicting progression to androgen independent prostate cancer (AIPC). METHODS: Records from 150 patients with advanced or metastatic prostate cancer treated with androgen deprivation therapy (ADT) were retrospectively reviewed. Androgen independent progression was defined as two consecutive elevations of serum prostate specific antigen (PSA) above the nadir value. Kaplan-Meier and the Cox proportional hazards methods were used to assess potential predictors of progression to AIPC. RESULTS: Patients with low and moderate Gleason scores experienced significantly longer remissions compared to those with Gleason score of 8-10 (p=0.0006, Log-Rank test). The cumulative hazard of progressing to AIPC increased by almost 70% for each unit increase in total Gleason score. CONCLUSION: In this patient cohort the Gleason score was the only independent predictor of progression to AIPC. PMID- 12121724 TI - Visualization of prostate cancer with 11C-choline positron emission tomography. AB - BACKGROUND AND OBJECTIVE: Visualization of prostate cancer with positron emission tomography (PET) using 2-[18F]-2-deoxy-D-glucose (FDG) as radiopharmaceutical is limited by the low uptake of FDG in the tumor and by radioactivity excreted into the bladder. More specific PET radiopharmaceuticals would be welcome. Carbon-11 labeled choline (CHOL) is a new radiopharmaceutical potentially useful for tumor imaging as it is incorporated in the cell membranes as phosphatidylcholine. We prospectively studied the visualization of prostate cancer using CHOL PET. METHODS: A total of 25 consecutive patients with histologically proven prostate cancer and five patients with a benign prostate were included. PET images were performed with an ECAT HR(+) using 400MBq CHOL. Data acquisition was started at 5 minutes post-injection. Attenuation-corrected images were evaluated visually. Standardized uptake values (SUV) were calculated of the normal prostate gland and of the prostate tumor tissue. RESULTS: The normal prostate was visualized with a mean SUV of 2.3 (range 1.3-3.2). The primary tumor could be visualized with a mean SUV of 5.0 (range 2.4-9.5). Lymph node metastases >5mm could be identified. Non-specific uptake of CHOL was noticed in the intestines. Little to no radioactivity in the bladder was observed. CONCLUSION: Carbon-11-choline is avidly taken up in prostate cancer, both primary tumor and lymph node metastases, in the virtual absence of urinary radioactivity. These results confirm the early results obtained by others and permit further clinical research on the value of CHOL PET as a metabolic imaging technique in areas where conventional imaging have a limited sensitivity. PMID- 12121725 TI - Seminal secretory capacity of the male accessory sex glands in chronic pelvic pain syndrome (CPPS)/chronic prostatitis with special focus on the new prostatitis classification. AB - OBJECTIVE: The aim of the study was to evaluate the secretory dysfunction of the male accessory glands in men with inflammatory versus non-inflammatory chronic pelvic pain syndrome (CPPS). METHODS: One hundred and twelve consecutive patients symptomatic for chronic pelvic pain were included into the study. All underwent a combined granulocyte analysis in expressed prostatic secretions (EPS) and a four glass-test followed by ejaculate analysis. Patients were subgrouped according to elevated granulocyte counts in prostatic secretions, leukocytes in semen, or any of both. The content/total enzyme activity of the secretory seminal plasma parameters gamma-glutamyl-transferase (gamma-GT), fructose, and alpha-glucosidase representing the secretory capacity of the prostate gland, the seminal vesicles, and the epididymes, respectively, were investigated. RESULTS: The only significant findings were a reduced total enzyme activity of gamma-GT in men stratified according to elevated granulocyte counts in prostatic secretions (p=0.022; cutpoint 9.85U per ejaculate; sensitivity 61.1%, specificity 58.8%, AUC 0.6347) and in men with any inflammatory sign (p=0.033; cutpoint 9.9U per ejaculate, sensitivity 63%, specificity 58.33%, AUC 0.6404). CONCLUSIONS: Secretory damage of the prostate gland in men with inflammatory CPPS is demonstrable provided that increased granulocytes in prostatic secretions are part of the diagnostic criteria. However, because of the low sensitivity and specificity of gamma-GT it cannot be recommended as diagnostic tool to detect inflammatory disease on the basis of reduced secretory capacity. PMID- 12121726 TI - The efficacy and safety of the tension-free vaginal tape procedure do not depend on the method of analgesia. AB - OBJECTIVE: The original tension-free vaginal tape (TVT) method, described by Ulmsten et al., routinely uses local anaesthesia during the procedure. Since the anaesthetic effect after local application of lidocaine hydrochloride was not always satisfactory we decided to introduce the spinal anaesthesia during this operation. The aim of the present study was to compare local and spinal anaesthesia with respect to their efficacy and safety in the TVT procedure. METHODS: 103 women, with objectively confirmed stress urinary incontinence, were randomised into the study. Sixty-seven women were anaesthetised locally and 36 patients spinally. All TVT procedures were performed as originally described. Objective assessment of the influence of anaesthesia on intra-abdominal pressure at rest and during the cough test was done using a rectal catheter and a central venous pressure manometer. The efficacy of the TVT procedure was based on a gynaecological examination with a cough test and a three-degree subjective scale: complete cure, improvement or failure. RESULTS: The success of the TVT procedure performed under local anaesthesia is comparable with that achieved under spinal analgesia (p=0.42). The number of complications that occurred in the two groups does not differ significantly (p=0.57). CONCLUSIONS: Spinal anaesthesia impairs the ability to cough effectively during the TVT procedure. However, the efficacy and safety of the operations performed under this type of anaesthesia are comparable with the efficacy and safety of operations done under local anaesthesia. PMID- 12121727 TI - Cystometrical sensory data from a normal population: comparison of two groups of young healthy volunteers examined with 5 years interval. AB - OBJECTIVE: To describe the pattern of sensations reported during standardized cystometry in a group of healthy young volunteers and compare them with a group examined 5 years before. METHODS: A group of 50 young healthy volunteers without any symptoms or history reported the sensations they felt during cystometry. These results were compared with those of another group of 38 young healthy volunteers examined in the same lab in 1995 by another investigator. RESULTS: All participants perceived a first sensation of bladder filling, first desire to void and strong desire to void. Each sensation was easily distinguishable from the others. The volumes at which these sensations came up varied widely. The ratio between volumes at consecutive sensations and at full bladder was fairly constant. All but two parameters were not significantly different from those found in 1995. CONCLUSIONS: Our data give additional weight to previous findings that there exists a normal pattern of sensations reported during cystometric bladder filling. This sensory pattern probably corresponds with specific physiological mechanisms as suggested before. Deviations from this pattern indicate or illustrate pathology. PMID- 12121728 TI - Intravesical meglumine gamma-linolenic acid in superficial bladder cancer: an efficacy study. AB - OBJECTIVES: Gamma-linolenic acid (GLA) is known to be cytotoxic to malignant cells. We assessed the efficacy of the novel intravesical formulation, meglumine gamma-linolenic acid (MeGLA), in a phase II trial, in patients with recurrent, superficial bladder cancer. PATIENTS AND METHODS: Thirty patients with recurrent, superficial transitional cell carcinoma (TCC) were recruited. The tumour pattern was recorded at flexible cystoscopy. Patients received a single intravesical instillation of 50ml of either 50mg (1mg/ml) (15 patients), or 125mg (2.5mg/ml) (15 patients) of MeGLA in water, retained for one hour. At subsequent cystoscopy, the tumour patterns were recorded, prior to undertaking routine cystodiathermy. Biopsies were obtained for histological assessment. Responses were divided into complete, partial or none. RESULTS: All 30 patients retained the drug for 1 hour without significant local or systemic side effects. There were 4 (13%) complete responses, 9 (30%) partial responses, and 17 (57%) non-responders. Histology showed no evidence of damage to surrounding urothelium. CONCLUSIONS: Our data confirms the safety and tolerability of MeGLA, which is consistent with findings from a previous phase I trial. A response rate of 43% also indicates that MeGLA has a significant cytotoxic effect against TCC and the results are similar to those obtained using standard, single-dose, intravesical regimens. PMID- 12121729 TI - Conservative management of upper urinary tract tumors. AB - PURPOSE: We determined the immediate and long-term results of endoscopic management of upper tract transitional cell in regard to rates of tumor recurrence and preservation of renal function. MATERIALS AND METHODS: From January 1990 to July 1999, 61 patients (mean age 66.2 years) underwent endoscopic management of upper tract cell carcinoma. Of the patients 20 (32%) had a solitary kidney. Tumors were resected in a one time procedure by ureteroscopy only in 31.5%, by percutaneous nephroscopy in 29% or both in 8%; multiple treatment was necessary in 31.5% of cases using percutaneous nephroscopy only. RESULTS: Immediate nephrectomy was done in six cases for high grade (three patients), insufficient local control (two cases) or patient's choices (one case). There were six cases of benign tumors excluded from survival Kaplan Meier analysis. With a mean follow-up of 39.9 months, the rate of kidney preservation, recurrence free rate, global survival and specific survival rates were, respectively, 81%, 68%, 77%, and 84%. CONCLUSIONS: Nephron sparing percutaneous management of upper tract cell carcinoma is applicable in a significant number of patients with a filling defect of upper urinary tract TCC. In carefully selected patients the results are at least comparable to other forms of management of tumor control and preservation of renal function. PMID- 12121730 TI - Treatment of uretero-intestinal and ureterovesical strictures by Acucise balloon catheter. AB - OBJECTIVE: Acucise balloon catheter has been proposed as an alternative to open surgery for the treatment of strictures of the ureteropelvic junction because of its low morbidity and the short hospital stay following the endoscopic procedure. The objective of this study was to evaluate the results of this technique applied to patients developing strictures after surgical reimplantation of the ureterovesical (UV) or uretero-intestinal (UI) junction. MATERIAL AND METHODS: Between March 1997 and January 2000, 12 strictures (11 patients) were treated by Acucise balloon catheter via an antegrade and/or retrograde approach with double J stenting for an average of 6 weeks (range: 4-12 weeks): six uretero-ileal strictures (three Bricker, one uretero-ileoplasty, one enterocystoplasty and one Kock pouch) and six ureterovesical strictures (Lich-Gregoir or Paquin UV reimplantations after gynaecological, vascular or endoscopic surgery). The median postoperative follow-up was 16 months (range: 10-36 months). A good result was defined by the absence of recurrence of the stricture evaluated both clinically and radiologically (regression of stasis measured by IVU and/or ultrasonography). RESULTS: The mean operating time was 70 minutes and the mean hospital stay was 4.8 days (range: 3-14 days). Only one intraoperative complication was observed (migration of the double J stent to the kidney). The operation was successful in eight patients (75%). The success rate was 83% for ureterovesical strictures and 50% for uretero-ileal strictures. A history of previous irradiation appeared to be a factor of failure. CONCLUSION: The Acucise procedure is a minimally invasive and effective (75% success rate) treatment option for the treatment of postoperative stricture after ureteric reimplantations. In our department, this option is considered to be first-line treatment, as surgical reimplantation is reserved for failures of the endoscopic technique. PMID- 12121731 TI - Botulinum a toxin and detrusor sphincter dyssynergia: a double-blind lidocaine controlled study in 13 patients with spinal cord disease. AB - OBJECTIVE: To compare the efficacy and tolerance of botulinum A toxin (BTx) versus lidocaine (L), applied in the external urethral sphincter with a single transperineal injection in order to treat detrusor sphincter dyssynergia (DSD) in spinal cord injured patients. METHODS: Thirteen patients (1F, 12 M) suffering from chronic urinary retention due to DSD were randomised to receive one transperineal injection of 100 IU BTx Botox degrees in 4 ml of 9% saline (botulinum group, (BG)) or 4 ml of 0.5% L (lidocaine group, (LG)). The main criteria of efficacy was post-voiding residual urine volume (PRUV), assessed three times daily on day one (D1), D7 and D30 after each injection. Other criteria were micturition diary, satisfaction score (SS), maximal urethral pressure (MUP), maximum detrusor pressure (DP) and type of DSD, recorded on D0 and D30. RESULTS: In the BG, there was a significant decrease in PRUV (D7: -141.4 ml (p<0.03); D30: -159.4 ml (p<0.01)), in MUP (D30: -32 cm H(2)O, p<0.04) whereas no significant improvement was shown in the LG. SS was higher in BG than LG (p<0.02). DSD improved in BG whereas it remained unchanged in LG. All LG patients also received one injection of BTx on D30. They still presented improvement in PRUV and MUP 1 month later (D30'). Tolerance appeared satisfactory in both groups. CONCLUSIONS: The preliminary results of this initial randomised double blind study clearly demonstrated the superiority of BTx compared to L in improving clinical symptoms and urethral hypertonia associated with DSD in spinal cord injured patients. PMID- 12121732 TI - Long-term outcome of the retained ureteral stump after lower pole heminephrectomy in duplex kidneys. AB - OBJECTIVES: Duplication of the ureter and renal pelvis is the most common upper urinary tract anomaly in childhood. The anatomical and functional divisions between upper and lower moieties of duplex kidney are extremely variable. The underlying pathological condition associated with a lower moiety is usually massive vesicoureteral reflux (VUR) to the lower collecting system and only rare obstruction. The non-functioning upper moiety is usually associated with obstructive ectopic ureter (with or without ureterocele). Most lower pole heminephrectomies are carried out for non-functioning lower moieties. In most cases, the lower defunctionalised segment of the ureter is left in situ. Complete ureterectomy is usually performed if presence of VUR into the lower end of the corresponding ureter is shown. There is little information on the long-term outcome of residual ureteral 'stumps'. The purpose of our study was to review the long-term outcome of retained ureteral stumps in children undergoing heminephrectomy for non-functioning lower pole moieties in duplex kidneys. MATERIALS AND METHODS: The medical records of 19 patients who underwent 20 lower pole heminephrectomies for a non-functioning lower pole moiety of a duplex kidney between January 1990 and December 2000 were reviewed retrospectively. Median age at heminephrectomy was 4.5 years (range: 1 month to 12 years). Indications for heminephrectomy in the 20 renal units was reflux nephropathy in 16 (80%) and obstructive nephropathy in 4 (20%). All corresponding ureters were taken down as low as possible and transfixed through the heminephrectomy incision. Median follow-up was 8.5 years (range: 1-11 years). RESULTS: Eight (40%) showed VUR into the stump after lower pole heminephrectomy. Two of these underwent subureteral endoscopic correction of VUR with polytetrafluoroethylene paste and resection of the stump was carried out in remaining two patients for recurrent urinary tract infections (UTI). Remaining four of the eight patients demonstrated spontaneous resolution of VUR during follow-up. CONCLUSIONS: Our data suggest that the vast majority of patients with residual ureteral stumps after lower pole heminephrectomy do not require stump resection at long-term follow-up. PMID- 12121733 TI - Induction of antibodies against prostate-specific membrane antigen (PSMA) by vaccination with a PSMA DNA vector. AB - INTRODUCTION AND OBJECTIVES: Prostate-specific membrane antigen (PSMA) is a 750 amino acid surface protein expressed primarily in prostate epithelium, and is upregulated 10-fold in prostate cancer. It is therefore an attractive target for immunotherapy. However, most reported antibodies to PSMA apparently recognize epitopes in the residue 43-570 region of the extracellular domain, and upon binding are rapidly removed from the cell surface by internalization. This would potentially limit their ability to mediate Fc-dependent cytoxicity. In this study, we constructed a DNA expression vector, pV/TM-PSMc, in which this region was deleted from full-length PSMA cDNA. Mice were vaccinated with pV/TM-PSMc DNA to determine whether humoral responses directed against PSMA-positive human prostate cancer cells could be induced by this C-terminal region. METHODS: Polymerase chain reaction (PCR)-based techniques were used to delete codons 50 570 from the coding region of human PSMA cDNA, thereby joining the C-terminal end (PSMc) to the N-terminal cytoplasmic/transmembrane domain (TM). This truncated product, TM-PSMc, was cloned into the vector pNGVL3 (pV). The resulting vector, pV/TM-PSMc, was confirmed by DNA sequencing, and by expression studies using reverse transcriptase (RT)-PCR for transcripts and immunohistochemical (IHC) staining with the PSMA monoclonal antibody (mAb) 7E11.C5. BALB/c mice were injected in the tibialis anterior muscle four times, at biweekly intervals, with 100 microg vector DNA per injection. One week after the last injection, blood was drawn for serum preparation. The serum was assayed for antibodies against PSMA by IHC staining of LNCaP, a PSMA-positive human prostate cancer line. Expression in vaccinated muscle cells was determined by RT-PCR assay for TM-PSMc transcripts. RESULTS: NIH3T3 cells transfected with pV/TM-PSMc stained positively by IHC reaction with mAb 7E11.C5. 48h after one intramuscular (i.m.) injection of mice with 100 microg pV/TM-PSMc vector DNA, TM-PSMc transcripts were detectable in muscle RNA by RT-PCR analysis. Anti-serum from pV/TM-PSMc-DNA vaccinated mice, at a dilution of 1:20, intensely IHC-stained both live and fixed LNCaP cells. CONCLUSIONS: These results demonstrate that anti-PSMA humoral responses were induced by i.m. injection of mice with pV/TM-PSMc DNA. Antibodies in the anti serum were directed against extracellular epitopes of native PSMA expressed by human prostate cancer cells. Vaccination with DNA expression vectors such as pV/TM-PSMc may provide an immunotherapeutic approach for the treatment of prostate cancer. PMID- 12121735 TI - Fibroblast growth factors (FGFs) in the cochlear nucleus of the adult mouse following acoustic overstimulation. AB - To see if fibroblast growth factors (FGFs) might function in the central changes following auditory overstimulation we tracked immunostaining in the cochlear nucleus of adult mice with monoclonal antibodies to FGFs (FGF-1, FGF-2) and FGF receptor. After exposure nearly all outer hair cells died, while inner hair cell and fiber loss were restricted to a region midway along the cochlear spiral. FGFs staining in the cochlear nucleus appeared in hypertrophied astrocytes in the regions of nerve fiber degeneration only. For normal-sized astrocytes there was an increase in the number stained and the intensity of staining across all frequency domains, but not in neurons. The increases were modest at 3-7 days, pronounced at 14 days, modest again by 30 days, and back to control levels by 60 days. FGF receptor staining of neurons occurred equally in all mice, exposed or not. The findings suggest that astrocytes play a role in the central responses to acoustic overstimulation and cochlear damage, involving FGFs, possibly regulating the activity of intrinsic neurons or signaling axonal growth. Not limited to regions of cochlear nerve fiber and inner hair cell loss, the changes in FGFs may represent a reaction to outer hair cell damage which spreads broadly across the central pathways. PMID- 12121736 TI - Stereocilia defects in waltzer (Cdh23), shaker1 (Myo7a) and double waltzer/shaker1 mutant mice. AB - Mutations in myosin VIIa (Myo7a) and cadherin 23 (Cdh23) cause deafness in shaker1 (sh1) and waltzer (v) mouse mutants respectively. In humans, mutations in these genes cause Usher's syndrome type 1B and D respectively, as well as certain forms of non-syndromic deafness. Examination of the organ of Corti from shaker1 and waltzer mice has shown that these genes are required for the proper organisation of hair cell stereocilia. Here we show that at embryonic day 18.5, the outer hair cells of Cdh23(v) homozygote mutant mice appear immature, projecting fewer recognisable stereocilia than heterozygote controls, and by post natal day (P) 4 their stereocilia are arranged in a disorganised pattern rather than in the regular 'V'-shape seen in heterozygotes. Inner hair cell stereocilia are also disorganised in Cdh23(v) mutant homozygotes. Myo7a was expressed normally in the hair cells of P0 Cdh23(v2J) mutants demonstrating that cadherin 23 is not required for Myo7a expression at this stage. No stereocilia defects were observed in P4 Cdh23(v)/Myo7a(4626SB) double heterozygotes (+/Cdh23(v) +/Myo7a(4626SB)) and neither the Cdh23(v) nor Myo7a(4626SB) homozygote phenotypes were affected by the presence of one mutant copy of Myo7a or Cdh23 respectively. The hair cell phenotype of double homozygote mutant mice did not differ from single Myo7a(4626SB) homozygote mutants. Finally, we found no significant correlation between loss of hearing and double heterozygosity for mutations in Cdh23 and Myo7a in mice aged between 7.5 and 10 months. These findings suggest that Cdh23 and Myo7a are both required for establishing and/or maintaining the proper organisation of the stereocilia bundle and that they do not genetically interact to affect this process nor to cause age-related hearing loss. PMID- 12121738 TI - Can fishes resolve temporal characteristics of sounds? New insights using auditory brainstem responses. AB - Numerous fish species produce broad-band pulsed sounds with a distinct temporal patterning which is thought to be important during intraspecific communication. In order to determine whether fishes are able to utilize temporal characteristics of acoustic signals, time resolution was determined in four species of otophysines and anabantoids by analyzing auditory brainstem responses (ABRs) to double-click stimuli with varying click periods. At click periods of 3.5 ms, two distinct ABRs were clearly detectable in all species. The minimum pulse period resolvable by the auditory system was below 1.5 ms in each species and slightly intensity-dependent. No differences were found between vocal and non-vocal species within each taxon. Comparisons of the time resolution data to the pulse periods of intraspecific sounds in the vocal species showed that the otophysine Platydoras costatus and the anabantoid Trichopsis vittata are likely to process each pulse within a series of intraspecific sounds. However, as non-vocal and vocal species have a similar minimum resolvable click period, the high temporal resolution capacities of the auditory system of fish might not represent special adaptations for intraspecific acoustic communication. Nonetheless, we suggest that temporal characteristics of naturally occurring conspecific and heterospecific sounds provide reliable information for acoustic communication. PMID- 12121737 TI - Effects of caloric restriction and aging on the auditory function of rhesus monkeys (Macaca mulatta): The University of Wisconsin Study. AB - The present study is part of a larger project that investigates the effect of caloric restriction on longevity in the rhesus monkey. The purpose of the present study was to document presbycusis and the effect of caloric restriction on presbycusis in monkeys. The control group had 35 monkeys allowed to eat freely and the caloric-restricted group (CR) had 33 monkeys with a 30% reduction in caloric intake. Monaural and binaural auditory brainstem response (ABR) and middle latency response (MLR) were obtained from 27 female and 41 male monkeys that were 11-23 years of age and had been in the study for 102, 42, or 36 months when tested. Significant findings were the following: (1) wave I amplitudes were larger for females and for younger monkeys, and amplitudes decreased in aging males but not in aging females; (2) wave IV amplitudes were larger for females than males, and amplitudes for CR females were larger than for female controls, whereas the amplitudes from control and CR males were not different; (3) wave Pa latencies were shorter for females, and shorter latencies were maintained for aging females but not for aging males; (4) interwave interval IV-Pa was shorter for females, and intervals lengthened for aging males but not aging females; (5) binaural wave IV amplitude decreased faster with age for control monkeys than for CR monkeys, and the L+R Pa amplitude decreased with age. Additional trends were identified for longitudinal monitoring as monkeys enter old age. PMID- 12121739 TI - Involvement of the mitochondrial permeability transition in gentamicin ototoxicity. AB - Aminoglycosides may induce irreversible hearing loss in both animals and humans. In order to study the nature and mechanisms underlying gentamicin-induced cell death in the inner ear, the cochlear neurosensory epithelia were dissected from guinea pigs and incubated with 0.5-10 mM gentamicin. Concentration-dependent loss of cell viability was detected by the inability of damaged cells to exclude propidium iodide. Outer hair cells were most sensitive towards gentamicin toxicity, followed by inner hair cells whereas Deiters and Hensen cells were not affected by the gentamicin concentrations used. The iron chelators 2,2'-dipyridyl and deferoxamine provided partial protection against gentamicin-induced hair cell death while the calcium chelator Quin-2 AM had no effect. Gentamicin (0.5-1 mM) induced condensation of chromatin typical for apoptosis. Using the fluorescent dye tetramethyl-rhodamine methyl ester and laser scanning microscopy we could visualize a loss of the mitochondrial membrane potential in damaged outer hair cells about 1 h before cell death occurred. Cyclosporin A, an inhibitor of the mitochondrial permeability pore, provided partial protection against gentamicin toxicity. This strongly suggests an involvement of the mitochondrial permeability transition in gentamicin-induced apoptosis. PMID- 12121740 TI - Expression of heregulin and ErbB/Her receptors in adult chinchilla cochlear and vestibular sensory epithelium. AB - Immunolabeling of heregulin, a growth factor that enhances cell proliferation in damaged utricles, and one of its binding receptors, ErbB-2, has been briefly described in the P3 rat cochlea and utricle [Zheng et al. (1999) J. Neurocytol. 28, 901-912]. However, little is known about the distribution of heregulin and its three binding receptors in adult animals. Here we describe the immunolabeling patterns for heregulin, ErbB-2, ErbB-3 and ErB-4 in the cochlea, spiral ganglion, utricle and saccule of the adult chinchilla using confocal microscopy. Heregulin immunolabeling was intense along the apical pole of Deiters cells and Hensen cells and along the membrane of supporting cells of the utricle and saccule; light immunolabeling was present in the outer layer of the spiral prominence and cytoplasm of spiral ganglion neurons. In the cochlea, intense to moderate ErbB-2 immunolabeling was evident in the cytoplasm of pillar cells, outer hair cells (OHCs), border cells, stria vascularis and spiral ligament; moderate ErbB-2 immunolabeling was present in the cytoplasm of the hair cell and supporting cell layers of the utricle and saccule. In the cochlea, light ErbB-3 immunolabeling was present in the inner hair cells, OHCs, marginal and intermediate cell layers of the stria vascularis and spiral ganglion neurons; moderate ErbB-3 immunolabeling was present in the cytoplasm of hair cells and supporting cells of the utricle and saccule. In the cochlea, utricle and saccule, ErbB-4 immunolabeling was intense in the nuclei and light to moderate in the cytoplasm and membrane of sensory cells and supporting cells. These results suggest that heregulin acting through ErbB receptors and various receptor complexes may play an important role in cell proliferation and survival in the cochlea and vestibular system. PMID- 12121741 TI - Inner ear pathology in the mucopolysaccharidosis VII mouse. AB - Mucopolysaccharidosis type VII (MPS VII, Sly syndrome) is caused by dysfunction of the acid hydrolase beta-D-glucuronidase. The defect results in the accumulation of incompletely degraded glycosaminoglycans within lysosomes of a wide array of cell types. MPS VII is associated with mixed (conductive and sensorineural) hearing loss, vision defects, shortened stature, mental retardation and decreased lifespan. Whether the sensorineural component of hearing loss in MPS VII involves degeneration of cochlear sensory cells is not yet clear. The MPS VII mouse resembles its human counterpart in all major aspects, and has been the focus of extensive research seeking to correct MPS VII and other lysosomal storage diseases. The value of potential treatments for this hearing loss can be determined only if cochlear pathology in this model is well characterized. We examined threshold sensitivity, frequency tuning, hair cell density and the appearance of the cochlea and vestibular organs in MPS VII mice ranging from 1.0 to 7.5 months of age. At all ages, lysosomal storage is pronounced within cells of spiral limbus, spiral prominence, spiral ligament and glial cells, but not within organ of Corti, stria vascularis, or neurons. Within the vestibular maculae and cristae, both hair cells and supporting cells also show lysosomal storage. Although hearing thresholds are never normal, reduction in the sharpness of frequency tuning is not apparent until 2.5 months of age, suggesting that the sensorineural component of hearing loss begins in adulthood. No evidence was found for cell loss within the organ of Corti, or any other structure, however. Our results suggest that sensorineural hearing loss in the MPS VII mouse is not caused by degeneration, but may arise from alterations in mass and stiffness of cochlear structures or impaired sensory cell function. They also indicate a possible vestibular component in MPS VII. PMID- 12121742 TI - Comparison of binaural auditory brainstem responses and the binaural difference potential evoked by chirps and clicks. AB - Rising chirps that compensate for the dispersion of the travelling wave on the basilar membrane evoke larger monaural brainstem responses than clicks. In order to test if a similar effect applies for the early processing stages of binaural information, monaurally and binaurally evoked auditory brainstem responses were recorded for clicks and chirps for levels from 10 to 60 dB nHL in steps of 10 dB. Ten thousand sweeps were collected for every stimulus condition from 10 normal hearing subjects. Wave V amplitudes are significantly larger for chirps than for clicks for all conditions. The amplitude of the binaural difference potential, DP1-DN1, is significantly larger for chirps at the levels 30 and 40 dB nHL. Both the binaurally evoked potential and the binaural difference potential exhibit steeper growth functions for chirps than for clicks for levels up to 40 dB nHL. For higher stimulation levels the chirp responses saturate approaching the click evoked amplitude. For both stimuli the latency of DP1 is shorter than the latency of the binaural wave V, which in turn is shorter than the latency of DN1. The amplitude ratio of the binaural difference potential to the binaural response is independent of stimulus level for clicks and chirps. A possible interpretation is that with click stimulation predominantly binaural interaction from high frequency regions is seen which is compatible with a processing by contralateral inhibitory and ipsilateral excitatory (IE) cells. Contributions from low frequencies are negligible since the responses from low frequencies are not synchronized for clicks. The improved synchronization at lower frequencies using chirp stimuli yields contributions from both low and high frequency neurons enlarging the amplitudes of the binaural responses as well as the binaural difference potential. Since the constant amplitude ratio of the binaural difference potential to the binaural response makes contralateral and ipsilateral excitatory interaction improbable, binaural interaction at low frequencies is presumably also of the IE type. Another conclusion of this study is that the chirp stimuli employed here are better suited for auditory brainstem responses and binaural difference potentials than click stimuli since they exhibit higher amplitudes and a better signal-to-noise ratio. PMID- 12121743 TI - Effects of noise and cue enhancement on neural responses to speech in auditory midbrain, thalamus and cortex. AB - Speech perception depends on the auditory system's ability to extract relevant acoustic features from competing background noise. Despite widespread acknowledgement that noise exacerbates this process, little is known about the neurophysiologic mechanisms underlying the encoding of speech in noise. Moreover, the relative contribution of different brain nuclei to these processes has not been fully established. To address these issues, aggregate neural responses were recorded from within the inferior colliculus, medial geniculate body and over primary auditory cortex of anesthetized guinea pigs to a synthetic vowel consonant-vowel syllable /ada/ in quiet and in noise. In noise the onset response to the stop consonant /d/ was reduced or eliminated at each level, to the greatest degree in primary auditory cortex. Acoustic cue enhancements characteristic of 'clear' speech (lengthening the stop gap duration and increasing the intensity of the release burst) improved the neurophysiologic representation of the consonant at each level, especially at the cortex. Finally, the neural encoding of the vowel segment was evident at subcortical levels only, and was more resistant to noise than encoding of the dynamic portion of the consonant (release burst and formant transition). This experiment sheds light on which speech-sound elements are poorly represented in noise and demonstrates how acoustic modifications to the speech signal can improve neural responses in a normal auditory system. Implications for understanding neurophysiologic auditory signal processing in children with perceptual impairments and the design of efficient perceptual training strategies are also discussed. PMID- 12121744 TI - Transgene expression in neonatal mouse inner ear explants mediated by first and advanced generation adenovirus vectors. AB - The mouse serves as a valuable model for treatment leading to the prevention and therapy of inner ear disease. Transgenic correction of genetic inner ear disease in mice may help develop treatment for human genetic inner ear disease. In mutations involving hair cells (HCs) or supporting cells (SCs), it is necessary to insert the wild-type transgenes directly into these cells. We used inner ear explants to characterize the transgenic expression using adenovirus-mediated reporter genes (bacterial lacZ). The variable parameters were the age of the explants (P1-P5), the type of vector (first and advanced generation adenovirus) and the genotype of the mouse (wild-type versus shaker-2 mutant). Transduction of cochlear HCs was detected at P1 and in some of the P3 cochleae. Low efficiency transduction of SCs was observed in P1 explants, but the efficiency increased with age and reached high levels at P5. The pattern of transduction was similar regardless of the genotype and the type of vector used. The data demonstrate that differentiating HCs and SCs in mouse explants can be transduced by adenovirus vectors, suggesting that cultures of mouse ears are a valuable model for developing inner ear gene therapy protocols. PMID- 12121745 TI - Amikacin ototoxicity enhanced by Ginkgo biloba extract (EGb 761). AB - An animal study was realized to investigate the possible beneficial effect of EGb 761 as an antioxidant agent on amikacin ototoxicity by measuring distortion product otoacoustic emissions (DPOAEs). Twenty-eight adult rats were grouped equally as follows. GROUP AMIKACIN: rats received amikacin 600 mg/kg/day intramuscularly between postnatal days (PND) 30 and PND44. Group amikacin/EGb 761: rats received amikacin 600 mg/kg/day intramuscularly between PND30 and PND44 and EGb 761 100 mg/kg/day orally between PND30 and PND50. Group EGb 761: rats received equivolume saline intramuscularly between PND30 and PND44 and EGb 761 100 mg/kg/day orally between PND30 and PND50. NO TREATMENT GROUP: rats received nothing. Group amikacin was found to be affected only on the last measurement day of study (PND57). The frequencies greater than 2002 Hz were significantly reduced compared with the amplitudes of PND30 (P<0.05). Group amikacin/EGb 761 was most and earliest affected by amikacin-induced ototoxicity. DPOAE amplitudes were found in this group to be decreased at 2-6 kHz starting on PND50. The results of Group EGb 761 and No treatment group were not significantly changed. For the DPOAE input/output amplitude thresholds, Group amikacin (P<0.05) and Group amikacin/EGb 761 (P<0.01) had significantly elevated thresholds on PND57, except at 5 kHz for Group amikacin (P=0,06). According to the results of the study, EGb 761 may be regarded as a facilitating drug for the development of amikacin ototoxicity. The results of the present study may warn against concomitant use of aminoglycosides, specifically amikacin, with EGb 761. PMID- 12121746 TI - The effect of eye position and background noise on vertical sound localization. AB - The influence of the eye position in combination with a simultaneously presented background (frame) sound on the auditory localization of a single (target) sound source was investigated in the median vertical plane (MVP). Five loudspeakers ranged from -20 degrees to +20 degrees with the center speaker at 0 degrees elevation. Listeners verbally estimated the position of a 500-ms noise stimulus, which was presented while they were fixating visual targets in various elevations. The perceived sound elevation appeared to be shifted 8.6 degrees on average towards the direction of eccentric gaze. When the target was temporarily embedded in a 2-s frame sound (emitted by one of the two outer loudspeakers) and the gaze was fixated at a straight-ahead position, the listeners consistently shifted the apparent target positions about 4 degrees away from the frame locations. This auditory contrast effect, which is consistent with earlier findings, remained even if the gaze direction deviated from straight ahead. It is concluded that the auditory contrast effect exists independently from the eye position effect and that the two effects act separately on auditory sound localization. PMID- 12121747 TI - Interaction of excitation and inhibition in inferior collicular neurons of the big brown bat, Eptesicus fuscus. AB - Neurons in the inferior colliculus (IC) of the midbrain receive convergent excitatory and inhibitory inputs from both lower and higher auditory nuclei. Interaction of these two opposing inputs shapes different response properties of IC neurons. In this study, we examined this interaction of excitation and inhibition in IC neurons using a probe (excitatory pulse) and a masker (inhibitory pulse) under different stimulation conditions. Inhibition of probe elicited responses by a masker, i.e. masking, occurred when the masker was presented at certain inter-pulse intervals (the temporal window) in relation to the probe. At the best inter-pulse interval, masking was maximal such that a neuron had the minimal number of impulses, the longest response latency, and the smallest excitatory frequency tuning curve. The temporal window for masking expanded with increasing masker duration. The inhibition decreased with increasing probe intensity but increased with increasing masker intensity. Increasing masker intensity also produced progressive shrinkage in excitatory frequency tuning curves. Similarly, increasing probe intensity produced progressive shrinkage of inhibitory frequency tuning curves. Possible mechanisms underlying the time and intensity dependence of inhibition are discussed. PMID- 12121748 TI - Evidence for interactions across frequency channels in the inferior colliculus of awake chinchilla. AB - As a result of cochlear processing, information about acoustic broadband signals is distributed across many parallel frequency channels. Periodic modulations of signal envelopes - conspicuous in particular in harmonic signals - may extend across a wide frequency range and give rise to temporal response patterns in the auditory nerve, particularly useful for recombination of constituents and the separation of the signals from background noise. Herein we report evidence that across frequency processing as necessary for binding of related signal components occurs already in the auditory midbrain of mammals. Extracellular recordings were made from 231 multi and single units in the inferior colliculus of awake chinchillas. Loud pure tones evoked onset type excitation (26%) and suppression of spontaneous rate (60%) not only in the range of the units' characteristic frequency (CF), but also in a frequency range far above CF. About 80% of all units tuned to CFs below 3 kHz gave sustained responses to low level stimuli of high frequencies (>2CF) provided the tones were sinusoidally amplitude modulated (SAM) with a unit specific modulation frequency although none of the spectral components of the amplitude modulation alone was sufficient to evoke such a response, even at high intensities. Low level high carrier SAM responses and wide band onset responses as well as inhibition must have their origin in a non-linear across frequency channel interaction of neuronal information. Many aspects of these responses cannot be explained by peripheral distortion in the cochlea. We therefore propose a mechanism of integration across frequency channels that may originate within the inferior colliculus and/or the nuclei of the lateral lemniscus. This process may lead to the binding of information that shares a common periodicity and may thereby help to distinguish different acoustic objects. PMID- 12121749 TI - Protection and treatment of sensorineural hearing disorders caused by exogenous factors: experimental findings and potential clinical application. AB - During the last decade, there have been numerous interesting findings regarding the roles of neurotrophins, nitric oxide, reactive oxygen species, glutamate receptors, and shock protein in the auditory system. These findings have provided a scientific basis for the development of techniques to protect the auditory system against trauma as well as for the treatment of peripheral hearing disorders. This review focuses on recent advances in experimental prevention and treatment of hearing impairment which are expected to be of clinical value in the near future. Viral vector and non-viral vector gene therapy and transplantation of stem cells are discussed as potential treatments of irreversible sensorineural inner ear damage. PMID- 12121750 TI - Affinity analysis of lectin interaction with immobilized C- and O- gylcosides studied by surface plasmon resonance assay. AB - A biosensor based on the surface plasmon resonance (SPR) principle was used for kinetic analysis of lectin interactions with different immobilized saccharide structures. A novel affinity ligands beta-D-glycopyranosylmethylamines derived from common D-aldohexoses linked to the carboxymethyl dextran layer of the SPR sensor surface served for interactions with a wide range of lectins. The method of preparation and use of the beta-D-mannopyranosyl glycosylated sensor surface was described. The results of affinity analysis of lectin-ligand interactions were evaluated and compared with data obtained from measurements using commercially available p-aminophenyl alpha-D-glycopyranosides. Possible applications and advantages of C- and O-glycosylated SPR biosensors are discussed. PMID- 12121751 TI - Interaction process between ionic surfactant and protein probed by series piezoelectric quartz crystal technique. AB - A method for probing the interaction process between ionic surfactant and protein was developed with series piezoelectric quartz crystal (SPQC) sensing technique. It was based on the sensitive response of the SPQC sensor to the change in solution conductivity. A new relationship between the sensor response and the properties of ionic species in solution was derived. The method was used to examine the interaction process of two surfactants, cetyltrimethylammonium bromide (CTAB) and sodium dodecyl sulfate (SDS), with lysozyme in aqueous solution. The obtained experimental results were in agreement with those of other methods from references. These results had been discussed. It was shown that the new method developed here was a useful and promising tool for probing the ionic surfactant-protein interaction process and might find more applications in similar studies. PMID- 12121752 TI - An approach to increased polyplex gene delivery by peptides selected from a phage display library. AB - Phage display libraries were screened for peptides to be incorporated in nonviral gene delivery vehicles. Cells in culture were incubated with heptamer random peptide libraries displayed on M13 bacteriophages in three to five copies per phage. Surface-adherent phages were removed or inactivated and the cells were fractionated in a nuclear pellet and supernatant. Bacteriophages from each of the two fractions were amplified and reincubated with the cells. Three successive rounds of selection were performed. Eighteen sequenced clones revealed 14 different sequences. Two sequences were homologous to segments of the HIV gp120 protein. For three sequences, the corresponding synthetic peptides were generated and attached via avidin-biotin to polylysine-condensed plasmid DNA containing a reporter gene. The addition of the peptides led to 8-14 times increase in the expression of the reporter. PMID- 12121753 TI - Energy transfer method in membrane studies: some theoretical and practical aspects. AB - Some applications of resonance energy transfer (RET) method to distance estimation in membrane systems are considered. The model of energy transfer between donors and acceptors randomly distributed over parallel planes localized at the outer and inner membrane leaflets is presented. It is demonstrated that RET method can provide evidence for specific orientation of the fluorophore relative to the lipid-water interface. An approach to estimating the depth of the protein penetration in lipid bilayer is suggested. PMID- 12121754 TI - Immobilization of invertase on rice husk using polyethylenimine. AB - Washed and dried rice husk was coated with 2% polyethylenimine (PEI). Invertase was immobilized on this support through adsorption followed by cross-linking with 2% glutaraldehyde. Immobilized enzyme was reused for the hydrolysis of sucrose without loss in activity. This approach may serve as a simple technique in the future for the covalent immobilization of enzymes on lignocellulosic supports. PMID- 12121755 TI - Selective extraction of lecithin:cholesterol acyltransferase (EC 2.3.1.43) from human plasma. AB - A method for the rapid extraction of lecithin:cholesterol acyltransferase (LCAT) from human plasma or serum has been developed. The method is based on direct treatment of acidified plasma of fully conserved enzyme activity, with the strong ion exchanger Q-Sepharose, which under the experimental conditions bound all LCAT but only about 10% of the total protein content of the plasma, no albumin and essentially no lipoproteins. This corresponds to a 10-fold purification. Only traces of apolipoprotein A-I remained in the quantitatively desorbed LCAT preparation which, however, contained a residual fraction of apolipoprotein D and acidic plasma proteins. The present one-step procedure for extraction of LCAT in high yields from human plasma represents a simple and efficient alternative to the first step in previously described methods for preparation of the enzyme to homogeneity. PMID- 12121756 TI - Fe-only hydrogenases: structure, function and evolution. AB - Hydrogenases are enzymes capable of catalyzing the oxidation of molecular hydrogen or its production from protons and electrons according to the reversible reaction: H(2)<==>2H(+)+2e(-). Most of these enzymes fall into to major classes: NiFe and Fe-only hydrogenases. Extensive spectroscopic, electrochemical and structural studies have shed appreciable light on the catalytic mechanism of hydrogenases. Although evolutionarily unrelated, NiFe and Fe-hydrogenases share a common, unusual feature: an active site low-spin Fe center with CO and CN coordination. We have recently focused our attention on Fe-hydrogenases because from structural studies by us and others, it appears to be a simpler system than the NiFe counterpart. Thus the primary hydrogen binding site has been identified and plausible, electron, proton and hydrogen pathways from and to the buried active site may be proposed from the structural data. The extensive genome sequencing effort currently under way has shown that eukaryotic organisms contain putatively gene coding sequences that display significant homology to Fe hydrogenases. Here, we summarize the available evidence concerning the mechanism of these enzymes and carry out a structural comparison between Fe-hydrogenases and related proteins of unknown metal content from yeast, plant, worm, insect and mammals. PMID- 12121757 TI - Molecular and cellular mechanisms of iron homeostasis and toxicity in mammalian cells. AB - Iron is an essential metal for almost all living organisms due to its involvement in a large number of iron-containing enzymes and proteins, yet it is also toxic. The mechanisms involved in iron absorption across the intestinal tract, its transport in serum and delivery to cells and iron storage within cells is briefly reviewed. Current views on cellular iron homeostasis involving the iron regulatory proteins IRP1 and IRP2 and their interactions with the iron regulatory elements, affecting either mRNA translation (ferritin and erythroid cell delta aminolaevulinate synthase) or mRNA stability (transferrin receptor) are discussed. The potential of Fe(II) to catalyse hydroxyl radical formation via the Fenton reaction means that iron is potentially toxic. The toxicity of iron in specific tissues and cell types (liver, macrophages and brain) is illustrated by studies with appropriate cellular and animal models. In liver, the high levels of cyoprotective enzymes and antioxidants, means that to observe toxic effects substantial levels of iron loading are required. In reticuloendothelial cells, such as macrophages, relatively small increases in cellular iron (2-3-fold) can affect cellular signalling, as measured by NO production and activation of the nuclear transcription factor NF kappa B, as well as cellular function, as measured by the capacity of the cells to produce reactive oxygen species when stimulated. The situation in brain, where anti-oxidative defences are relatively low, is highly regionally specific, where iron accumulation in specific brain regions is associated with a number of neurodegenerative diseases. In the brains of animals treated with either trimethylhexanoylferrocene or aluminium gluconate, iron and aluminium accumulate, respectively. With the latter compound, iron also increases, which may reflect an effect of aluminium on the IRP2 protein. Chelation therapy can reduce brain aluminium levels significantly, while iron can also be removed, but with greater difficulty. The prospects for chelation therapy in the treatment and possible prevention of neurodegenerative diseases is reviewed. PMID- 12121758 TI - Physico-chemical aspects of hemozoin (malaria pigment) structure and formation. AB - Recent literature dealing with the chemical composition, structure and mechanism of formation of hemozoin and its synthetic counterpart, beta-hematin, is reviewed. PMID- 12121759 TI - Mechanisms of compound I formation in heme peroxidases. AB - The formation of compound I is the first step in the reaction mechanism of plant heme peroxidases. This intermediate stores two oxidizing equivalents from hydrogen peroxide as an oxyferryl iron center and a radical, either on the porphyrin ring or on a tryptophan residue. Site-directed mutagenesis has proved to be a most useful tool for the identification of the intermediates involved and the resulting nature of the compound I formed. Although there is no doubt that an acid-base mechanism operates in heme peroxidase during the formation of compound I, the roles of several distal pocket residues are currently the subject of intensive research. It is now generally accepted that the conserved distal histidine in the active site of heme peroxidases is the acid-base catalyst that promotes the heterolytic cleavage of hydrogen peroxide. Other residues, such as the distal arginine and asparagine, participate in a range of roles assisting catalysis by the distal histidine. Recent advances in the elucidation of the mechanism at the molecular level are discussed. Another aspect related to the nature of compound I is the location of the radical center. Novel radical species have been detected in the reactions of ascorbate peroxidase, lignin peroxidase and several mutants of horseradish peroxidase. Detailed kinetic and spectroscopic studies of these radical species have provided important insights about the factors that control porphyrin-protein radical exchange. The wide range of data being obtained on compound I will lead to an understanding of its vital function in peroxidase catalysis and the physiological roles played by these enzymes. PMID- 12121760 TI - Does P450-type catalysis proceed through a peroxo-iron intermediate? A review of studies with microperoxidase. AB - Recent stopped-flow kinetics demonstrated the existence of an intermediate before the occurrence of the final product of the reaction of both iron-containing microperoxidase-8 (Fe(III)MP-8) and manganese-containing microperoxidase-8 (Mn(III)MP-8) with H(2)O(2). The intermediate was assigned to be (hydro)peroxo iron. With both mini-catalysts the final state obtained after 30-40 ms showed a resemblance to PorM(IV)MP-8[double bond]O(R(+)*); (R(+)*) is a radical located at the peptide. Quantum mechanical calculations indicate that hydroperoxo-iron is inactive as a catalytic intermediate in cytochrome P450 (P450)-type catalysis. Instead, the calculations suggest that peroxo-iron acts as the catalytic intermediate in P450-type catalysis. In addition, the calculations demonstrate that, although less likely, the possibility that oxenoid-iron acts as a catalytic intermediate in P450 catalysis cannot be fully excluded. An interesting aspect of the reactions catalysed by MP-8 is the possibility that, in view of the reversibility of the reactions between (hydro)peroxo-iron and oxenoid-iron, H(2)O plays a decisive role, at least in some cytochromes P450, in the removal of halogens, avoiding the production of compounds hazardous to the organism. PMID- 12121761 TI - Metal binding and structure-activity relationship of the metalloantibiotic peptide bacitracin. AB - Bacitracin is a widely used metallopeptide antibiotic produced by Bacillus subtilis and Bacillus licheniformis with a potent bactericidal activity directed primarily against Gram-positive organisms. This antibiotic requires a divalent metal ion such as Zn(2+) for its biological activity, and has been reported to bind several other transition metal ions, including Mn(2+), Co(2+), Ni(2+), and Cu(2+). Despite the widespread use of bacitracin since its discovery in the early 1940s, the structure-activity relationship of this drug has not been established and the coordination chemistry of its metal complexes was not fully determined until recently. This antibiotic has been suggested to influence cell functioning through more than one route. Since bacterial resistance against bacitracin is still rare despite several decades of widespread use, this antibiotic can serve as an ideal lead for the design of potent peptidyl antibiotics lacking bacterial resistance. In this review, the results of physical (including NMR, EPR, and EXAFS) and molecular biological studies regarding the synthesis and structure of bacitracin, the coordination chemistry of its metal derivatives, the mechanism of its antibiotic actions, its influence on membrane function, and its structure and function relationship are discussed. PMID- 12121762 TI - Reactivity of recombinant and mutant vanadium bromoperoxidase from the red alga Corallina officinalis. AB - Vanadium bromoperoxidase (VBPO) from the marine red alga Corallina officinalis has been cloned and heterologously expressed in Esherichia coli. The sequence for the full-length cDNA of VBPO from C. officinalis is reported. Steady state kinetic analyses of monochlorodimedone bromination reveals the recombinant enzyme behaves similarly to native VBPO from the alga. The kinetic parameters (K(m)(Br )=1.2 mM, K(m)(H(2)O(2))=17.0 microM) at the optimal pH 6.5 for recombinant VBPO are similar to reported values for enzyme purified from the alga. The first site directed mutagenesis experiment on VBPO is reported. Mutation of a conserved active site histidine residue to alanine (H480A) results in the loss of the ability to efficiently oxidize bromide, but retains the ability to oxidize iodide. Kinetic parameters (K(m)(I-)=33 mM, K(m)(H(2)O(2))=200 microM) for iodoperoxidase activity were determined for mutant H480A. The presence of conserved consensus sequences for the active sites of VBPO from marine sources shows its usefulness in obtaining recombinant forms of VBPO. Furthermore, mutagenesis of the conserved extra-histidine residue shows the importance of this residue in the oxidation of halides by hydrogen peroxide. PMID- 12121763 TI - Site-directed mutagenesis for cysteine residues of cobalt-containing nitrile hydratase. AB - Three cysteine residues, which are completely conserved among alpha-subunits in all nitrile hydratases, are thought to be the ligands of a metal ion in the catalytic center of this enzyme. These cysteine residues (i.e. alpha C102, alpha C105 and alpha C107) in the high-molecular-mass nitrile hydratase (H-NHase) of Rhodococcus rhodochrous J1 were replaced with alanine by site-directed mutagenesis using the R. rhodochrous ATCC12674 host-vector system, and the resultant transformants were investigated. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) for the cell-free extracts of each mutant transformant revealed that four mutant transformants (i.e. alpha C105A, alpha C107A, alpha C102A/C105A and alpha C105A/C107A) showed predominant alpha- and beta-subunit protein bands with a mobility identical to those of the native H NHase, while three mutant transformants (i.e. alpha C102A, alpha C102A/C107A and alpha C102A/C105A/C107A) did not produce the corresponding proteins. The purified former four mutant enzymes showed neither enzymatic activity nor the maximum absorption at 410 nm which was detected in the wild type H-NHase. They also did not contain cobalt ions. Based upon these findings, these three cysteine residues were found to be essential for the active expression of H-NHase. PMID- 12121764 TI - Engineering the proximal heme cavity of catalase-peroxidase. AB - Catalase-peroxidases (KatGs) are prokaryotic heme peroxidases with homology to yeast cytochrome c peroxidase (CCP) and plant ascorbate peroxidases (APXs). KatGs, CCP and APXs contain identical amino acid triads in the heme pocket (distal Arg/Trp/His and proximal His/Trp/Asp), but differ dramatically in their reactivities towards hydrogen peroxide and various one-electron donors. Only KatGs have high catalase activity in addition to a peroxidase activity of broad specificity. Here, we investigated the effect of mutating the conserved proximal triad on KatG catalysis. With the exception of W341F, all variants (H290Q, W341A, D402N, D402E) exhibited a catalase activity <1% of wild-type KatG and spectral properties indicating alterations in heme coordination and spin states. Generally, the peroxidase activity was much less effected by these mutations. Compared with wild-type KatG the W341F variant had a catalase and halogenation activity of about 40% and an even increased overall peroxidase activity. This variant, for the first time, allowed to monitor the hydrogen peroxide mediated transitions of ferric KatG to compound I and back to the resting enzyme. Compound I reduction by aromatic one-electron donors (o-dianisidine, pyrogallol, aniline) was not influenced by exchanging Trp by Phe. The findings are discussed in comparison with the data known from CCP and APX and a reaction mechanism for the multifunctional activity of the W341F variant is suggested. PMID- 12121765 TI - P(M) and P(R) forms of cytochrome c oxidase have different spectral properties. AB - The reaction between bovine heart cytochrome c oxidase and dioxygen was monitored at room temperature in the visible and Soret regions following photolysis of the mixed-valence CO-bound enzyme. Time-resolved optical absorption difference spectra were collected between 50 ns and 1.7 ms by a gated multichannel analyzer. Singular value decomposition and global exponential fitting resolved three processes with apparent lifetimes of 2.2+/-0.5, 17+/-4 and 160+/-30 micros. The spectra of the intermediates were extracted based on a sequential kinetic mechanism and compared to the corresponding intermediate spectra observed during the reaction of the fully reduced enzyme with dioxygen. The first process is associated with a conformational change at heme a(3) upon dissociation of CO from Cu(B)(+) and concomitant back-electron transfer from heme a(3) to heme a. This is followed by O(2) binding to heme a(3) forming compound A (A(M)), with a spectrum identical to that observed upon O(2) binding to heme a(3) in the fully reduced enzyme (A(R)). Intermediate A(M) decays into P(M), the spectrum of which is equivalent to that of the 607 nm form, generated upon addition of H(2)O(2) to the oxidized enzyme at alkaline pH values (P(H)). However, the spectrum of P(M) is significantly different from the corresponding intermediate observed upon the reaction of dioxygen with the fully reduced enzyme (P(R)). The spectral differences between P(M) and P(R) may arise from the different number of redox equivalents at the binuclear site, with a tyrosine radical in the P(M) state, and tyrosine or tyrosinate in P(R), or may be the consequence of a more complex reaction mechanism in the case of the fully reduced enzyme. PMID- 12121766 TI - Contribution of heme-propionate side chains to structure and function of myoglobin: chemical approach by artificially created prosthetic groups. AB - Horse heart myoglobin was reconstituted with mesohemin derivatives methylated at the 6- or 7-position to evaluate the role of the heme-6-propionate or heme-7 propionate side chain in the protein. The association and dissociation of the O(2) binding for the deoxymyoglobin with 6-methyl-7-propionate mesoheme are clearly accelerated. Furthermore, the myoglobin with 6-methyl-7-propionate mesoheme shows fast autoxidation from oxymyoglobin to metmyoglobin compared to the myoglobin with 6-propionate-7-methyl heme and the reference protein. These results indicate the 6-propionate plays an important physiological role in the stabilization of oxymyoglobin because of the formation of a salt-bridge with the Lys45. The acceleration of CO binding rate is observed for the myoglobin with 6 propionate-7-methyl mesoheme, suggesting that the replacement of the 7-propionate with a methyl group has an influence on the His93-heme iron coordination. The structural perturbation of His93 imidazole was also supported by 1H NMR spectra of cyanide and deoxy forms of the myoglobin with 6-propionate-7-methyl mesoheme. Thus, it is found that the 7-propionate regulates the hydrogen-bonding network and His93-heme iron coordination in the proximal site. PMID- 12121767 TI - Theoretical study of the discrimination between O(2) and CO by myoglobin. AB - Combined quantum chemical and molecular mechanics geometry optimisations have been performed on myoglobin without or with O(2) or CO bound to the haem group. The results show that the distal histidine residue is protonated on the N(epsilon 2) atom and forms a hydrogen bond to the haem ligand both in the O(2) and the CO complexes. We have also re-refined the crystal structure of CO[bond]myoglobin by a combined quantum chemical and crystallographic refinement. Thereby, we probably obtain the most accurate available structure of the active site of this complex, showing a Fe[bond]C[bond]O angle of 171 degrees, and Fe[bond]C and C[bond]O bond lengths of 170-171 and 116-117 pm. The resulting structures have been used to calculate the strength of the hydrogen bond between the distal histidine residue and O(2) or CO in the protein. This amounts to 31-33 kJ/mol for O(2) and 2-3 kJ/mol for CO. The difference in hydrogen-bond strength is 21-22 kJ/mol when corrected for entropy effects. This is slightly larger than the observed discrimination between O(2) or CO by myoglobin, 17 kJ/mol. We have also estimated the strain of the active site inside the protein. It is 2-4 kJ/mol larger for the O(2) complex than for the CO complex, independent of which crystal structure the calculations are based on. Together, these results clearly show that myoglobin discriminates between O(2) and CO mainly by electrostatic interactions, rather than by steric strain. PMID- 12121768 TI - NMR studies on interaction of lauryl maltoside with cytochrome c oxidase: a model for surfactant interaction with the membrane protein. AB - Interaction of lauryl maltoside (LM) surfactant with bovine heart cytochrome c oxidase (CcO) has been studied by NMR techniques. Detailed 2-D (1)H and (13)C NMR techniques were used to assign the NMR signals of the surfactant nuclei. Paramagnetic dipolar shift of the surfactant (13)C NMR signals were used to identify the atoms close to the enzyme. The diamagnetic carbon monoxide complex of CcO did not cause any shift in the surfactant NMR spectra suggesting that the paramagnetic centres of the native CcO cause the shifts by dipolar interactions. The results showed that the polar head groups of the surfactant comprised of two maltoside rings are more affected, while the hydrophobic tail groups did not show any significant change on binding of the surfactant to the enzyme. This indicated that surfactant head groups possibly bind to the enzyme surface and the hydrophobic tail of the surfactant forms micelles and remains away from the enzyme. Based on the results, we propose that the membrane bound enzyme is possibly stabilised in aqueous solution by association with the micelles of the neutral surfactant so that the polar heads of the micelles bind to the polar surface of the enzyme. These micelles might form a 'belt like' structure around the enzyme helping it to remain monodispersed in the active form. PMID- 12121769 TI - Primary structure of a Japanese lacquer tree laccase as a prototype enzyme of multicopper oxidases. AB - The cDNA library of the Japanese lacquer tree (Rhus vernicifera) was constructed by the reverse transcription of mRNA. A cDNA encoding laccase was amplified by PCR using primers based on the N-terminal amino acid sequences of the purified laccase and its peptide fragments formed by digestions with chymotrypsin and trypsin, and subcloned. The laccase cDNA clone contained a single, large open reading frame of 1599 nucleotides, encoding a protein of 533 amino acids with a calculated molecular mass of 58981 Da. The lacquer laccase was found to have 42 to 62% identity with other plant laccases and 20 to 24% identity with microorganism laccases at the deduced amino acid level. Differing from microorganism laccases the lacquer laccase utilizes a Met residue in addition to one Cys and two His residues to construct the type 1 Cu site. The secondary structure of the lacquer laccase was predicted to mainly consist of the beta structure (28.7%) and loop and random structures (67.0%). The alpha-helix content was predicted to be only 4.3%. The location of these secondary structures was assumed to be very similar to those of ascorbate oxidase and fungal laccase, the crystal structures of which have been determined. PMID- 12121770 TI - Spectroscopic and functional characterization of Cu-containing nitrite reductase from Hyphomicrobium denitrificans A3151. AB - The Cu-containing nitrite reductase from Hyphomicrobium denitrificans (HydNIR) has been spectroscopically and functionally characterized. The visible absorption spectrum implies that the enzyme has two type 1 Cu ions in one subunit (ca. 50 kDa). The electron paramagnetic resonance (EPR) spectrum of HydNIR is simulated assuming the sum of three distinct S = 1/2 systems: two type 1 Cu signals (axial and rhombic symmetries) and one type 2 Cu signal. The intramolecular electron transfer reaction from the type 1 Cu to the type 2 Cu at pH 6.0 does not occur in the absence of nitrite, but a very slow electron transfer reaction is observed in the presence of nitrite. The apparent first-order rate constants for the intramolecular electron transfer reactions (k(ET(intra))) in the presence of nitrite and also the apparent catalytic rate constants (k(cat)) of HydNIR decrease gradually with increasing pH in the range of pH 4.5-7.5. These pH profiles are substantially similar to each other, suggesting that the intramolecular electron transfer process is linked to the subsequent nitrite reduction process. PMID- 12121771 TI - Spectroscopic and structural characterization of copper(II) and palladium(II) complexes of a lichen substance usnic acid and its derivatives. Possible forms of environmental metals retained in lichens. AB - The metal binding properties of a phenolic lichen substance usnic acid (UA) and its acetyl and enamine derivatives 9-O-acetylusnic acid (MAUA), 7,9-di-O acetylusnic acid (DAUA), Delta(2,11)-enaminousnic acid (EUA), and N-substituted Delta(2,11)-enaminousnic acids have been studied by synthetic and spectroscopic methods, and the structures of copper(II) and palladium(II) complexes have been established by the X-ray diffraction method. Cu(II) reacted with UA and DAUA to give the binary complexes Cu(UA)(2) x H(2)O and Cu(DAUA)(2), respectively, and Cu(bpy) (bpy=2,2'-bipyridine) formed ternary complexes with UA and DAUA. Pd(II) also reacted with UA, DAUA, EUA, and N-substituted Delta(2,11)-enaminousnic acids to give the corresponding binary complexes. All the isolated complexes are insoluble in water and soluble in most organic solvents. They exhibited very strong absorption and circular dichroism spectral peaks in the UV region. The (1)H-NMR spectrum in CDCl(3) of the Pd(II) complex of N-phenyl-Delta(2,11) enaminousnic acid (PEUA), Pd(PEUA)(2) x C(6)H(6), showed that the C(4)-proton signal suffered a large upfield shift (0.86 ppm) due to the ring current effect of the N-phenyl moiety. X-Ray crystal structure analysis has been performed for Cu(bpy)(UA)(ClO(4)) x CH(3)OH, Pd(MEUA)(2) x C(6)H(6), and Pd(PEUA)(2) x C(6)H(6). Cu(bpy)(UA)(ClO(4)) x CH(3)OH has a square-pyramidal structure with the two nitrogen atoms of bpy and the two oxygen atoms of the mono-deprotonated B ring of UA in the equatorial positions, while Pd(II) binds with two molecules of MEUA or PEUA in the trans configuration through the nitrogen and oxygen atoms with deprotonation. The N-phenyl ring of PEUA in Pd(PEUA)(2).C(6)H(6) was revealed to be located close to the C(4) proton as indicated by (1)H-NMR. Isolation of Cu(2)(bpy)(2)(UA)(NO(3))(2) x 2H(2)O suggests that UA has two metal binding sites that can form polymeric complexes. The present results substantiate the metal binding ability and the structures of the complexes of usnic acid and other substances from lichens as biomonitors of environmental metal ions. PMID- 12121772 TI - Structures and electronic properties of the catecholatoiron complexes in relation to catechol dioxygenases: chlorocatecholatoiron complexes are compared to the 3,5 di-tert-butylcatecholatoiron complex in the solid state and in solution. AB - Chlorocatecholatoiron complexes, [Fe(TPA)(4Cl[bond]Cat)]BPh(4) and [Fe(TPA)(3Cl[bond]Cat)]BPh(4), (4Cl[bond]Cat and 3Cl[bond]Cat: 4- and 3 chlorocatecholates, respectively; TPA: tris(2-pyridylmethyl)amine) were isolated as intermediates for the oxygenative cleavage of chlorocatechols by nonheme iron complexes. Geometric structures of these complexes together with [Fe(TPA)(DTBC)]BPh(4) (DTBC: 3,5-di-tert-butylcatecholate) as reference were analyzed by X-ray absorption spectroscopy (EXAFS) in the solid state and in solution. Structure of the DTBC complex in the solid state was shown to be noticeably different from the other complexes as seen in the magnetic susceptibility and spectroscopic data. Electronic and magnetic properties of these complexes were studied by X-ray absorption (XANES), electronic (VIS) and ESR spectroscopies, and magnetic susceptibility. Electron transfer from the catecholate ligand to the Fe(III) center was indicated by the Fe[bond]K edge values in XANES spectra and by the LMCT bands in electronic spectra. Magnetic susceptibility and ESR data indicated that at low temperatures the complexes are in equilibrium between the low (S=1/2) and high-spin (S=5/2) ferric states with the latter component increasing with temperature. Remarkable differences between the spin states in solid and in solution were observed with the DTBC complex. PMID- 12121774 TI - Biomimetic hydrolytic activation by Fe(III) aggregates: structures, reactivity and properties of novel oxo-bridged iron complexes. AB - The tetranuclear aggregate (enH(2))[Fe(4)(mu(3)-O)(heidi)(4)(mu-O,O' O(2)CNHC(2)H(4)NH(3))] x 4H(2)O contains a novel bidentate zwitterionic carbamic acid ligand. Magnetic studies indicate that the unsymmetrical Fe(4) core is ferrimagnetic with an S=4 ground state. Similar ligands have been obtained on rectangular tetranuclear aggregates [M(4)(mu-O)(mu-OH)(hpdta)(2)(mu-X)(2)](n-) (M[double bond]Fe, Al, Ga). The carbamic acid ligands are considered to result from the hydrolytic activation (fixation) of atmospheric CO(2) by the aggregate precursor to give a carbonato intermediate, which then reacts with the organic diamine used as base in the synthesis. Similar aggregates with acetate ligands result from hydrolytic activation of the DMA used as cosolvent. Closely related mechanisms for these two activation processes are proposed, which are also related to the accepted mechanisms for carbonic anhydrase and urease. PMID- 12121773 TI - Photo-induced oxidation of a dinuclear Mn(2)(II,II) complex to the Mn(2)(III,IV) state by inter- and intramolecular electron transfer to Ru(III)tris-bipyridine. AB - To model the structural and functional parts of the water oxidizing complex in Photosystem II, a dimeric manganese(II,II) complex (1) was linked to a ruthenium(II)tris-bipyridine (Ru(II)(bpy)(3)) complex via a substituted L tyrosine, to form the trinuclear complex 2 [J. Inorg. Biochem. 78 (2000) 15]. Flash photolysis of 1 and Ru(II)(bpy)(3) in aqueous solution, in the presence of an electron acceptor, resulted in the stepwise extraction of three electrons by Ru(III)(bpy)(3) from the Mn(2)(II,II) dimer, which then attained the Mn(2)(III,IV) oxidation state. In a similar experiment with compound 2, the dinuclear Mn complex reduced the photo-oxidized Ru moiety via intramolecular electron transfer on each photochemical event. From EPR it was seen that 2 also reached the Mn(2)(III,IV) state. Our data indicate that oxidation from the Mn(2)(II,II) state proceeds stepwise via intermediate formation of Mn(2)(II,III) and Mn(2)(III,III). In the presence of water, cyclic voltammetry showed an additional anodic peak beyond Mn(2)(II,III/III,III) oxidation which was significantly lower than in neat acetonitrile. Assuming that this peak is due to oxidation to Mn(2)(III,IV), this suggests that water is essential for the formation of the Mn(2)(III,IV) oxidation state. Compound 2 is a structural mimic of the water oxidizing complex, in that it links a Mn complex via a tyrosine to a highly oxidizing photosensitizer. Complex 2 also mimics mechanistic aspects of Photosystem II, in that the electron transfer to the photosensitizer is fast and results in several electron extractions from the Mn moiety. PMID- 12121776 TI - Reactions of peroxyl radicals with Fe(H(2)O)(6)(2+). AB - The reactions of RO(2)* radicals with Fe(H(2)O)(6)(2+) were studied, R[double bond]H; CH(3); CH(2)COOH; CH(2)CN; CH(2)C(CH(3))(2)OH; CH(2)OH; CHCl(2)/CCl(3). All these processes involve the following reactions: Fe(H(2)O)(6)(2+)+RO(2)*<==>(H(2)O)(5)Fe(III)[bond]OOR(2+) K(1) approximately 250 M(-1); (H(2)O)(5)Fe(III)[bond]OOR(2+)+H(3)O(+)/H(2)O- >Fe(H(2)O)(6)(3+)+ROOH+H(2)O/OH(-); (H(2)O)(5)Fe(III)[bond]OOR(2+)+2Fe(H(2)O)(6)(2+)-->3Fe(H(2)O)(6)(3+)+ROH; 2 RO(2)*-->Products; RO(2)*+(H(2)O)(5)Fe(III)[bond]OOR(2+)- >Fe(H(2)O)(6)(2+)+products. The values of k(1) and k(3) [reaction is clearly not an elementary reaction] approach the ligand exchange rate of Fe(H(2)O)(6)(2+), i.e. these reactions follow an inner sphere mechanism and the rate determining step is the ligand exchange step. The rate of reaction is several orders of magnitude faster than that of the Fenton reaction. Surprisingly enough the K(1) values are nearly independent of the redox potential of the radical and are considerably higher than calculated from the relevant redox potentials. These results indicate that the ROO(-) ligands considerably stabilise the Fe(III) complex, this stabilisation is smaller for radicals with electron withdrawing groups which raise the redox potential of the radical but decrease the basicity of the ROO(-) ligands, two effects which seem to nearly cancel each other. Finally, the results clearly indicate that reaction (5) is relatively fast and affects the nature of the final products. The contribution of these reactions to oxidation processes involving 'Fenton-like' processes is discussed. PMID- 12121775 TI - Synthesis, structure and catecholase activity of dinuclear copper and zinc complexes with an N(3)-ligand. AB - The preparation and characterization of dinuclear [M(II)(dbcat)(idpa)](2) (M[double bond]Zn (1), Cu (3); dbcat[double bond]3,5-di-tert-butylcatecholate; idpa[double bond]3,3'-iminobis(N,N-dimethylpropylamine)) complexes are described. Crystallographic characterization of the complex [Cu(II)(dbcat)(idpa)](2) has shown that the co-ordination geometry around copper(II) ions is distorted square pyramidal (triclinic, P-1, a=10.576(1) A, b=11.927(1) A, c=12.621(1) A, alpha=77.89(1) degrees, beta=88.65(1) degrees, gamma=70.21(1) degrees, V=1462.7(2) A(3), Z=2, R=0.0387). Both 1 and 3 were suitable catalysts for the catalytic oxidation of dbcatH(2) to dtbq (dtbq=3,5-di-tert-butyl-1,2 benzoquinone) with dioxygen at ambient conditions in good yields. However, on the basis of kinetic studies the copper- and zinc-catalyzed reactions showed different mechanisms. In the first case valence tautomerism [Cu(II)(dbcat)(idpa)]<==>[Cu(I)(dbsq)(idpa)] precedes the reaction with O(2), while with the zinc complex metal-bound catecholate reacts directly with O(2) with the formation of free superoxide anion. PMID- 12121777 TI - Examination of the effects of oxidation and ring closure on the cytotoxicities of the platinum complexes of N-(2-hydroxyethyl)ethane-1,2-diamine and ethane-1,2 diamine-N,N'-diacetic acid. AB - The crystal structures, electrochemical properties and cytotoxicities of platinum(II) and platinum(IV) complexes of the multidentate ligands N-(2 hydroxyethyl)ethane-1,2-diamine (NNOH) and ethane-1,2-diamine-N,N'-diacetic acid (H(2)enda) are reported. In the platinum(II) state the NNOH and H(2)enda ligands act as bidentate ligands, coordinating through the two amine groups with the hydroxyethyl and carboxylate groups remaining uncoordinated. Oxidation with hydrogen peroxide followed by refluxing yields the ring closed Pt(IV) complexes in which the NNOH and H(2)enda ligands are deprotonated and coordinate via the two amine groups and either the deprotonated alcohol group in the case of NNO or both carboxylato groups in the case of enda. The platinum(IV) complex of NNO is 2 to 5-fold more active against a panel of cisplatin sensitive and resistant human tumour cell lines than is the platinum(II) complex, whereas in the case of enda, the reverse is true. Ring closure to occupy both axial sites apparently leads to deactivation of platinum(IV) complexes, but a single closure does not necessarily do so. PMID- 12121778 TI - Circular dichroism study of the irreversibility of conformational changes induced by polyamine-linked dinuclear platinum compounds. AB - In this work, the reversibility of both the B-->Z and B-->A conformational change in polymer DNA induced by polynuclear platinum compounds was studied. The compounds examined were: [[trans-PtCl(NH(3))(2)](2)[NH(2) (CH(2))(6)NH(2)]](2+) (BBR3005); [[trans-PtCl(NH(3))(2)](2)[mu-spermine-N1,N12]](4+) (BBR3535); [[trans PtCl(NH(3))(2)](2)[mu-spermidine-N1,N8]](3+) (BBR3571); [[trans PtCl(NH(3))(2)](2)[mu-BOC-spermidine]](2+) (BBR3537); and [[trans PtCl(NH(3))(2)](2)[mu-trans-Pt(NH(3))(2)(H(2)N(CH(2))(6)NH(2))(2)]](4+) (BBR3464). The conformational changes were assessed by circular dichroism and the reversibility of the transitions was tested by subsequent titration with the DNA intercalator ethidium bromide (EtBr). Fluorescent quenching was also used to assess the ability of ethidium bromide to intercalate into A and/or Z-DNA induced by the compounds. The results were compared with those produced by the simple hexamminecobalt cation [Co(NH(3))(6)](3+). The data suggest that while conformational changes induced by electrostatic interactions are confirmed to be reversible, covalent binding induces irreversible changes in both the A and Z conformation. The relevance of these changes to the novel biological action of polynuclear platinum compounds is discussed. PMID- 12121779 TI - Aryl substituted ruthenium bis-terpyridine complexes: intercalation and groove binding with DNA. AB - The non-covalent interaction of five novel ruthenium(II) bis-terpyridine complexes with calf thymus DNA and, where appropriate, with poly[d(G-C)](2) and poly[d(A-T)](2) is described. Each complex is functionalised with aryl tail groups in the 4' position of the terpyridine ligands ((i) 9-anthracenyl, (ii) 4,4'-biphenyl, (iii) beta-naphthyl, (iv) 9-phenanthrenyl, and (v) 1-pyrenyl). Circular dichroism and linear dichroism show that the binding of three of the complexes (phenanthrenyl, anthracenyl and pyrenyl) at low metal complex concentration is dominated by intercalation of the aryl tail groups between the DNA bases. The complex with the biphenyl tail predominantly exhibits groove binding with no significant tail intercalation. The naphthyl derivative binds both by intercalation and a non-intercalative mode even at low metal complex concentrations. At high metal complex concentrations, aggregation of the complexes on the DNA is observed. Resonance light scattering indicates that the aggregates are of low nuclearity along the groove. PMID- 12121780 TI - Synthesis and characterization of water-insoluble and water-soluble dibutyltin(IV) porphinate complexes based on the tris(pyridinyl)porphyrin moiety, their anti-tumor activity in vitro and interaction with DNA. AB - The water-insoluble and water-soluble organotin(IV)porphinate complexes based on the tris-(4-pyridinyl)porphyrin and tris(N-methyl-4-pyridiniumyl)porphyrin moieties were synthesized and characterized by elemental analysis, (1)H NMR, IR and electrospray ionization mass spectra. The in vitro activity of the compounds against P388 leukemia and A-549 was determined. The results show that the anti tumor activities of organotin(IV)porphinate is related to the water solubility of the compounds and the central ion in the porphyrin ring. The interaction between the water-soluble dibutyltin(IV) porphinate (7 and 10) complexes and DNA has been investigated. The result shows that compounds 7 and 10 cause DNA hypochromism measured by A(260), a slight increase in the viscosity of the DNA, and an increase in the melting point of DNA by 2.9 and 1.6 degrees C, respectively at DNA(base)/Drug(Por) ratios of 60. The binding constants to DNA were 1.35+/-0.16 x 10(7) M(-1) (7) and 1.45+/-0.12 x 10(6) M(-1) (10) determined using EB competition method based on the porphyrin concentration, which is 20 and five times greater than that of precursor porphyrins [5-p,o-(carboxy)methoxyphenyl 10,15,20-tris(N-methyl-4-pyridiniumyl)] porphyrin (p,o-tMPyPac) to DNA. Electrophoresis test shows that the compounds cannot cleave the DNA. According to the electrophoresis test result and all the above results, the cytotoxic activity against P388 and A-549 tumor cells appears not to come from the cleavage of DNA caused by the compounds but from the high affinity of compounds to DNA. PMID- 12121781 TI - Synthetic procedures to monomethyl-platinum(II) complexes containing nitrogen ligands of biological relevance. AB - The complex trans-bis(dimethylsulfoxide)chloromethylplatinum(II) (1) is fairly soluble in water, where it undergoes multiple equilibria involving the formation of geometrically distinct [Pt(H(2)O)(DMSO)Cl(CH(3))] aqua-species. On reacting an aqueous solution of 1 with monodentate nitrogen donor ligands L, such as pyridines or amines, two well distinct patterns of behavior can be recognized: (i) a single stage fast substitution of one DMSO by the entering ligand, yielding a complex of the type trans(C,N)-[Pt(DMSO)(L)Cl(CH(3))] which contains four different groups coordinated to the metal and which undergoes a slow conversion into its cis-isomer, (ii) a double substitution affording cationic complex ions of the type cis-[Pt(L)(2)(DMSO)(CH(3))](+). When this latter reaction is carried out using sterically hindered ligands, slow rotation of the bulk ligand around the Pt[bond]N bond allows for the identification of head-to-head and head-to-tail rotamers in solution, through (1)H NMR spectrometry. The addition of chloride anion to 1 leads to the anionic species cis-[Pt(DMSO)Cl(2)(CH(3))](-), where a molecule of DMSO still remains coordinated to the metal center, despite its quite fast rate of ligand exchange (k(exch) with free DMSO=12+/-1 s(-1)). The reaction of complex 1 with bidentate ligands, such as ethylenediamine (en) or simple amino acids, leads to the cationic species [Pt(en)(DMSO)(CH(3))](+) or to the neutral [Pt(DMSO)(N[bond]O)(CH(3))], (where N[bond]-O[double bond]GlyO(-), AlaO(-)). PMID- 12121783 TI - Hydrolytically active Eu(III) and Ce(IV) EF-hand peptides. AB - A chimeric peptide (P4) has been designed to incorporate an EF-hand metal-binding loop into the context of the helix-turn-helix DNA binding motif of the engrailed homeodomain. This construct binds lanthanides, and in the presence of these metals, promotes the cleavage of supercoiled DNA and model phosphate esters (bisnitrophenyl phosphate). P4 binds lanthanides with moderate affinities (Eu(III), log K(a)=4.85; and Ce(IV), log K(a)=5.23). The structure of P4 is enhanced by metal binding, but the increase in secondary structure observed by CD is small, and suggests the metallopeptide is also quite flexible. Despite this flexibility, the efficient cleavage of DNA at low concentrations is dependent on the metallopeptide, and not on peptide or metal alone. This enhanced reactivity suggests the designed DNA-binding EF-hand peptides deliver the metal to the DNA for catalysis, even without rigid secondary structure. PMID- 12121782 TI - Depleted uranium-catalyzed oxidative DNA damage: absence of significant alpha particle decay. AB - Depleted uranium (DU) is a dense heavy metal used primarily in military applications. Published data from our laboratory have demonstrated that DU exposure in vitro to immortalized human osteoblast cells (HOS) is both neoplastically transforming and genotoxic. DU possesses both a radiological (alpha particle) and a chemical (metal) component. Since DU has a low-specific activity in comparison to natural uranium, it is not considered to be a significant radiological hazard. In the current study we demonstrate that DU can generate oxidative DNA damage and can also catalyze reactions that induce hydroxyl radicals in the absence of significant alpha particle decay. Experiments were conducted under conditions in which chemical generation of hydroxyl radicals was calculated to exceed the radiolytic generation by one million-fold. The data showed that markers of oxidative DNA base damage, thymine glycol and 8 deoxyguanosine could be induced from DU-catalyzed reactions of hydrogen peroxide and ascorbate similarly to those occurring in the presence of iron catalysts. DU was 6-fold more efficient than iron at catalyzing the oxidation of ascorbate at pH 7. These data not only demonstrate that DU at pH 7 can induced oxidative DNA damage in the absence of significant alpha particle decay, but also suggest that DU can induce carcinogenic lesions, e.g. oxidative DNA lesions, through interaction with a cellular oxygen species. PMID- 12121784 TI - Structures of HO(2)-Co(III)bleomycin A(2) bound to d(GAGCTC)(2) and d(GGAAGCTTCC)(2): structure-reactivity relationships of Co and Fe bleomycins. AB - HO(2)-Co(III)bleomycin is a model for HO(2)-Fe(III)bleomycin, which initiates single and double strand cleavage of DNA. In order to enlarge the understanding of its structure and reactivity, three-dimensional structures of HO(2) Co(III)bleomycin bound to two DNA oligomers, d(GAGCTC)(2) (I) and d(GGAAGCTTCC)(2) (II), that have 5'-GC-3' binding sites, have been determined by nuclear magnetic resonance (NMR) methods. Besides previously recognized determinants of binding selectivity, a probable hydrogen bond was detected between the pyrimidinyl acetamido NH(2) and the carbonyl of cytosine base paired to G at the recognition site. Another hydrogen bond between the NH of the dimethylsulfonium R group and N7 of guanine opposite cytosine at the GC site may contribute to specification of the pyrimidine. Substitution of G with inosine shifted HO(2)-Co(III)Blm A(2)[bond]I and Fe(III)Blm[bond]I into fast exchange on the NMR time scale, supporting the role of the 2-amino group in site specification for each molecule. The conformationally stable metal-domain linker established a close-packed adduct with the minor groove in which the hydroperoxide ligand occupies a sterically constrained pocket that is isolated from the solvent. The hydroperoxide group is directed toward one of the two cytosine H4' hydrogens but is sterically blocked from access to the other by the drug. These findings enlarge the structural understanding of selective binding of Co(III)/Fe(III)Blm species at G-pyrimidine sites. They also rationalize the instability of a number of ligands bound to Co(III)/Fe(III)Blm at specific binding sequences and the relative unreactivity of Fe(III)Blm[bond]I with ascorbate as well as its lack of interaction with spin labels. PMID- 12121785 TI - DNA-binding and cleavage studies of macrocyclic copper(II) complexes. AB - Three hexaaza macrocyclic copper (II) complexes with different functional groups have been synthesized and characterized by elemental analysis and infrared spectra. Absorption and fluorescence spectral, cyclic voltammetric and viscometric studies have been carried out on the interaction of [CuL(1)]Cl(2) (L(1)[double bond]3,10-bis(2-methylpyridine)-1,3,5,8,10,12 hexaazacyclotetradecane), [CuL(2)]Cl(2) (L(2)[double bond]3,10-bis(2 propionitrile)-1,3,5,8,10,12-hexaazacyclotetradecane) and [CuL(3)]Cl(2) (L(3)=3,10-bis(2-hydroxyethyl)-1,3,5,8,10,12-hexaazacyclotetradecane) with calf thymus DNA. The results suggest that three complexes can bind to DNA by different binding modes. The spectroscopic studies together with viscosity experiments and cyclic voltammetry suggest that [CuL(1)](2+) could bind to DNA by partial intercalation via pyridine ring into the base pairs of DNA. [CuL(2)](2+) may bind to DNA by hydrogen bonding and hydrophobic interaction while [CuL(3)](2+) may be by weaker hydrogen bonding. The functional groups on the side chain of macrocycle play a key role in deciding the mode and extent of binding of complexes to DNA. Noticeably, the three complexes have been found to cleave double-strand pUC18 DNA in the presence of 2-mercaptoethanol and H(2)O(2). PMID- 12121786 TI - Effect of divalent cations and cytosine protonation on thermodynamic properties of intermolecular DNA double and triple helices. AB - The contribution of divalent cations and cytosine protonation to conformation and stability of duplex and triplex formation were intensively investigated and characterized by ultraviolet (UV), circular dichroism (CD), differential scanning calorimetry (DSC), and electrophoresis mobility shift assay (EMSA). CD spectra showed that the divalent cations investigated would not significantly distort nucleotide geometry, while UV and DSC melting experiments revealed that the cation binding abilities to duplexes and triplexes were clearly dependent on the types of cations under near physiological conditions. The calorimetric enthalpies were generally underestimated relative to the corresponding van't Hoff enthalpies for Hoogsteen and Watson-Crick transitions, but free energy changes derived from the DSC measurements were in good agreement with those derived from the UV measurements. The adjacent placing of the C(+) x G.C triplets in triplexes lowered the stabilities of not only Hoogsteen base-pairing but also Watson-Crick base-pairing. The protonation contribution of the given cytosine residues might depend on the local and global structure of the protonated cytosine complex. A rigid structural targeted-strand would favor the protonation of cytosine residues. The apparent pK(a) values for parallel duplex and triplex investigated were determined to be 6.4 and 7.6, respectively, which are considerably heightened by 2.1 and 3.3 pH unit as compared to the intrinsic pK(a) value of the free cytosine residues. PMID- 12121787 TI - Picosecond Kerr-gated time-resolved resonance Raman spectroscopy of the [Ru(phen)(2)dppz](2+) interaction with DNA. AB - To investigate the basis of the 'light-switch' effect, the solvent dependence of the Kerr-gated picosecond-time resolved resonance Raman (TR(3)) spectra of [Ru(bpy)(2)dppz](2+), [Ru(phen)(2)dppz](2+), and the modified complex [Ru(phen)(2)cpdppzOMe](2+) and a dimer [mu-C4(cpdppz)(2)-(phen)(4)Ru(2)](4+) were studied. The investigation focussed on comparing the behaviour of [Ru(phen)(2)dppz](2+) in acetonitrile, ethanol, H(2)O, D(2)O, and DNA. The data are consistent with a model wherein excitation induces metal-to-ligand charge transfer (MLCT) to any of the ligands (termed the 'precursor' state) which, by interligand electron transfer (ILET), produces an excited state localised on the dppz ligand, MLCT(1). In water this state relaxes with a characteristic time of approximately 6 ps to a non-emissive state (MLCT(2)). The TR(3) spectra in water, acetonitrile and DNA are all distinctly different. However, the early (4 ps) water spectrum resembles the spectrum in DNA. This interesting observation suggests that the DNA-bound excited state of the complex can be thought of as a model for the initial, poorly solvated state in water. PMID- 12121788 TI - Synthesis, structure, spectroscopic and in vitro antitumour studies of a novel gallium(III) complex with 2-acetylpyridine (4)N-dimethylthiosemicarbazone. AB - The reaction of 2-acetylpyridine 4N-dimethylthiosemicarbazone (HL) with GaCl(3) in absolute ethanol in 1:1 molar ratio yielded the complex [GaL(2)][GaCl(4)]. The crystal structure of the gallium(III) complex has been determined by X-ray diffraction methods. Infrared, electronic, ESI mass and (1)H, (13)C, (15)N and (71)Ga NMR spectra, as well as the thermal behaviour are reported. The cytotoxicity assay in several human cancer cell lines (SW480, SK-BR-3 and 41M) suggests that the gallium(III) complex might be endowed with promising antitumour properties. In vitro cytotoxic activity exceeds that of all other tested gallium(III) complexes and is slightly higher than that of HL. PMID- 12121789 TI - Interactions of cisplatin and transplatin with proteins. Comparison of binding kinetics, binding sites and reactivity of the Pt-protein adducts of cisplatin and transplatin towards biological nucleophiles. AB - In this manuscript we report on the interactions of cis-DDP (cisplatin, cis diamminedichloroplatinum(II)) and trans-DDP (transplatin, trans diamminedichloroplatinum(II)) with two model proteins, ubiquitin (Ub) and horse heart myoglobin (Mb), and attempt to answer the question whether proteins that have methionine-Pt adducts can transfer the platinum to biological nucleophiles and particularly to DNA. Our study shows that cisplatin and transplatin form different adducts with ubiquitin: transplatin forms one major adduct, trans [Pt(Ub)(NH(3))(2)Cl], while cisplatin forms four distinct adducts, [Pt(Ub)(NH(3))(2)Cl], [Pt(Ub)(NH(3))(2)(H(2)O)], [Pt(Ub)(NH(3))(2)], and [Pt(Ub)(NH(3))]. When binding ubiquitin, Met1 is the preferred binding site of cisplatin, but not of transplatin. Cisplatin binds faster than transplatin to both ubiquitin and horse heart myoglobin. Both cisplatin and transplatin adducts form stable ternary adducts when reacted with 5'-guanosine monophosphate (5'-GMP) or a tetranucleotide. No transfer of the Pt moiety from the proteins to the nucleotides was observed. Glutathione efficiently removes the platinum from preformed adducts of both cisplatin and transplatin with ubiquitin. PMID- 12121790 TI - (153)Sm(3+) and (111)In(3+) DTPA derivatives with high hepatic specificity: in vivo and in vitro studies. AB - Two DTPA derivatives, a mono-amide derivative containing an iodinated synthon, DTPA-IOPsp (L(1)) and the ligand DTPA(BOM)(3) (BOM=benzyloxymethyl) (L(2)), radiolabelled with (153)Sm(3+) and (111)In(3+), were studied as potential hepatospecific gamma scintigraphic agents. In vivo studies with Wistar rats show that the main excretory pathway for all the chelates studied is the hepatobiliary system. The complexes of L(2) show even greater hepatobiliary specificity than L(1), perhaps as a consequence of longer blood circulation times due to their strong affinity towards HSA. The (153)Sm(3+) chelates are also more hepatospecific than the corresponding (111)In(3+) chelates. The La(3+) and In(3+) chelates of L(1) and L(2) show some structural and dynamic differences in aqueous solution, as studied by (1)H NMR spectroscopy. While only two nona-coordinated isomers were observed for the La(3+) complexes with both ligands, its number is much larger in the In(3+) complexes, with both octa- and hepta-coordinated species (with unbound side arms), as well as structural isomers for each coordination number. PMID- 12121791 TI - Studies on the kinetic stabilities of the Gd(3+) complexes formed with the N mono(methylamide), N'-mono(methylamide) and N,N"-bis(methylamide) derivatives of diethylenetriamine-N,N,N',N",N"-pentaacetic acid. AB - High kinetic stability is an important requirement for the Gd(3+) complexes used as contrast enhancement agents in magnetic resonance imaging. The kinetic stabilities of the Gd(3+) complexes formed with DTPA-N-mono(methylamide) (L(3)), DTPA-N'-mono(methylamide) (L(2)) and DTPA-bis(methylamide) (L(1)) are characterized by the rates of the exchange reactions with Eu(3+) and the endogenous Cu(2+) and Zn(2+). The exchange reactions occur via the proton assisted dissociation of the complexes and direct attack of the exchanging metal ions on the complex. On the basis of the line-shape analysis of the 1H NMR spectra of the LaL(2), obtained in the pH range 2.5-3.5, we assume that for the proton-assisted dissociation of the complexes the formation of an intermediate containing a free iminodiacetate group must be followed with the rupture of the metal-central nitrogen bond. At about pH > or = 5, the reactions between GdL(2) or GdL(3) and Cu(2+) or Zn(2+) proceed predominantly by direct reaction of the reactants, through the formation of dinuclear intermediates. The contribution of the proton-assisted dissociation is highly important for GdL(1), but its reaction with Zn(2+) is significantly slower than the reactions of GdL(2) and GdL(3). The overall rates of dissociation of GdL(1), GdL(2), GdL(3) and Gd(DTPA)(2-) through H(+) (pH 7.4), Cu(2+) (1 x 10(-6) M) and Zn(2+) (1 x 10(-5) M)-assisted reactions are surprisingly very similar. Replacement of one or two carboxylates with amide groups results in significantly decreased stability constants, but has practically no effect on the kinetic stability of the Gd(3+) complexes, indicating the lower reactivity of the amide groups with Cu(2+) and Zn(2+). PMID- 12121792 TI - Structure-dependent metallokinetics of antidiabetic vanadyl-picolinate complexes in rats: studies on solution structure, insulinomimetic activity, and metallokinetics. AB - The insulinomimetic effect of vanadium is the most remarkable and important among its several biological actions. Vanadyl ion (+4 oxidation state of vanadium) and its complexes have been found to normalize the blood glucose levels of both type 1 and 2 diabetic animals. We have developed insulinomimetic vanadyl complexes having different coordination modes, emphasizing the possible usefulness of vanadyl-picolinate [VO(pa)(2)] and its related complexes with the VO(N(2)O(2)) coordination mode. In order to apply these complexes clinically in the future, the relationship between the chemical structure, insulinomimetic action, organ distribution of vanadium, and blood disposition of vanadyl species must be closely investigated. In the present investigation, we studied the blood disposition of the vanadyl-picolinate complexes in healthy rats, and tried to understand comprehensively the relationship between the structures, insulinomimetic activity, and metallokinetic parameters of the complexes, which had been recently prepared and specifically synthesized for the present study, by using an in vivo blood circulation monitoring -- electron spin resonance (BCM ESR) method for analyzing ESR signals due to paramagnetic metal ions and complexes in the blood in real time. Metallokinetic parameters were estimated based on the blood clearance curves in terms of a two-compartment pharmacokinetic model, and vanadyl species were indicated to be distributed in peripheral tissues and gradually eliminated from the circulating blood, depending on their chemical structures. Vanadyl concentrations in the blood of rats given bis(5 iodopicolinato)oxovanadium(IV) [VO(5ipa)(2)] and bis(3 methylpicolinato)oxovanadium(IV) [VO(3mpa)(2)] with electron-withdrawing and donating groups, respectively, remained significantly higher and longer, due to their slower clearance rates from the blood, than in rats given other complexes, suggesting that the high exposure and long residence of vanadyl species bring about the high normoglyceric effect in diabetic animals. We then examined the relationship between insulinomimetic activity and metallokinetic parameters in the family of VO(pa)(2) for further development of insulinomimetic vanadyl complexes. IC(50), the 50% inhibitory concentration of the complexes on the free fatty acid release from isolated rat adipocytes treated with epinephrine, was found to be sufficiently correlated with metallokinetic parameters such as area under the concentration curve, mean residence time, total clearance, and distribution volume at steady-state. Furthermore, the in vivo antidiabetic activity of the complexes was enhanced with increasing exposure and residence of vanadyl species in the blood of animals. On the basis of these results, we concluded that in vitro insulinomimetic activity, metallokinetic character, and in vivo antidiabetic action of vanadyl-picolinate complexes are closely related to their chemical structures. PMID- 12121793 TI - Supramolecular structures of metronidazole and its copper(II), cobalt(II) and zinc(II) coordination compounds. AB - In this work we present the synthesis and structural and spectroscopic characterization of Cu(II), Co(II) and Zn(II) coordination compounds with the antibiotic metronidazole ([double bond]emni). Coordination to metal ions is through its imidazolic nitrogen, while the hydroxyethyl and nitro groups act as supramolecular synthons. [Co(emni)(2)Br(2)], and [Zn(emni)(2)X(2)] (X(-)=Cl, Br) stabilize zig-zag chains, and a 2D supramolecular structure is formed by inter chain contacts through inter-molecular hydrogen-bonding. Pleated sheet or layers are formed by [Co(emni)(2)Cl(2)] and [Cu(emni)(2)Cl(H(2)O)](2)Cl(2), respectively. The dinuclear Cu(II) compound [Cu(emni)mu(O(2)CMe)(2)](2) gives a one-dimensional zig-zag arrangement. The contribution of metal ions in metronidazole coordination compounds is shown in the stabilization of the different aggregate structures. PMID- 12121794 TI - Does tobacco marketing undermine the influence of recommended parenting in discouraging adolescents from smoking? AB - OBJECTIVE: The tobacco industry contends that parenting practices, not marketing practices, are critical to youth smoking. Our objective was to examine whether tobacco-industry marketing practices undermine the protective effect of recommended authoritative parenting against adolescent smoking. DESIGN AND SETTING: Receptivity to tobacco advertising and promotions was assessed in 1996 from a representative sample of California adolescent never-smokers aged 12 to 14 years. A follow-up survey of 1641 of these adolescents was conducted in 1999 that included measures of the key components of authoritative parenting: parental responsiveness, monitoring, and limit setting. MAIN OUTCOME MEASURE: Smoking initiation in adolescents. RESULTS: Adolescents in families with more authoritative parents were half as likely to smoke by follow-up as adolescents in families with less-authoritative parents (20% vs 41%, p <0.0001). In families with more-authoritative parents, adolescents who were highly receptive to tobacco industry advertising and promotions were significantly more likely to smoke (odds ratio=3.52, 95% confidence interval =1.10-11.23), compared to those who were minimally receptive. This effect was not significant in adolescents in families with less-authoritative parents. The overall attributable risk (adjusted for exposure to peer smokers) of smoking from tobacco-industry advertising and promotions was 25%. However, an estimated 40% of adolescent smoking in families with more-authoritative parents was attributable to tobacco-industry advertising and promotions; this was five times the attributable risk seen in families with less-authoritative parents (8%). CONCLUSION: The promotion of smoking by the tobacco industry appears to undermine the capability of authoritative parenting to prevent adolescents from starting to smoke. PMID- 12121795 TI - Domestic violence compared to other health risks: a survey of physicians' beliefs and behaviors. AB - BACKGROUND: Physicians routinely confront patient risk behaviors once considered private, including tobacco use, alcohol abuse, and HIV/STD-risk behavior. We compared physicians' behaviors and beliefs on screening and intervention for domestic violence with each other risk. METHODS: Survey of nationwide, random sample of 610 primary care physicians from the American Medical Association Physician Masterfile. RESULTS: Fewer primary care physicians screened for domestic violence than for other risks (p <0.001); once domestic violence was identified, however, physicians intervened with equal or greater frequency than for other risks. Fewer believed that they knew how to screen or intervene for domestic violence compared with other risks, and significantly fewer believed that domestic violence interventions were successful compared with interventions for tobacco and HIV/STD risks (Bonferroni adjusted p<0.001). CONCLUSIONS: Lower domestic violence screening rates may reflect physicians' beliefs that they do not know how to screen or intervene, and that interventions are less successful for domestic violence than for other risks. We may improve screening rates by educating physicians that a simplified role, as for other risks, can be effective for domestic violence. PMID- 12121796 TI - Sunburn prevalence among adults in the United States, 1999. AB - BACKGROUND: Exposure to high levels of sunlight, such as a sunburn, is a strong determinant of melanoma risk. METHODS: To describe statewide and U.S. estimates of sunburn prevalence in the United States and determine demographic and behavioral predictors of sunburn, we analyzed data from the 1999 Behavioral Risk Factor Surveillance System, a population-based telephone survey conducted in all 50 states, the District of Columbia, and Puerto Rico. RESULTS: Of 156,354 adults aged > or =18 years, 31.7% (95% confidence interval, 31.3%-32.1%) reported a sunburn in the past year; of adults aged 18 to 29 years, 57.5% reported such a sunburn. Reporting was highest among white, non-Hispanic males (44.1%), followed by white non-Hispanic females (35.3%), and lowest among black non-Hispanic males and females (5.1% and 5.3%, respectively). Statewide period prevalence of sunburn among whites was highest (>45%) in Wisconsin, Utah, Wyoming, Washington, DC, and Indiana, and lowest (<30%) in Puerto Rico, Arizona, Tennessee, Oklahoma, and New York. CONCLUSIONS: Nationwide and statewide skin cancer prevention efforts should target young adults. Periodic monitoring of sunburn is important in evaluating the effectiveness of those efforts. PMID- 12121797 TI - Vaccinating first-year college students living in dormitories for Meningococcal disease: an economic analysis. AB - BACKGROUND: Surveillance of meningococcal disease among U.S. college students found an elevated rate of this disease among first-year students living in dormitories. OBJECTIVE: This study examines the economics of routinely vaccinating a cohort of 591,587 incoming first-year students who will live in dormitories for > or =1 years. METHODS: A cost-benefit model (societal perspective) was constructed to measure the net present value (NPV) of various vaccination scenarios, as well as the cost/case and cost/death averted. Input values included hospitalization costs from $10,924 to $24,030 per hospitalization; immunization costs (vaccine plus administration costs) from $54 to $88 per vaccine; 30 nonfatal, vaccine-preventable cases over a 4-year period (includes 3 with sequelae); 3 premature deaths; value of human life from $1.2 million to $4.8 million; and long-run sequelae costs from $1298 to $14,600. Sensitivity analyses were also conducted on vaccine efficacy (80% to 90%); discount rate (0% to 5%); and coverage (60% to 100%). RESULTS: The costs of vaccination outweighed the benefits gained with NPVs ranging from -$11 million to -$49 million. The net cost per case averted ranged from $0.6 million to $1.9 million. The net cost per death averted ranged from $7 million to $20 million. The break-even costs of vaccination (when NPV=$0) at 60% coverage ranged from $23 (90% vaccine efficacy) to $5 (80% efficacy). CONCLUSIONS: The model showed that the vaccination program is not cost-saving. Key variables influencing the results were the low number of vaccine-preventable cases and the high cost of vaccination. However, from the perspective of students and parents, the cost of vaccination might be worth the real or perceived benefit of reducing the risk to an individual student of developing meningococcal disease. PMID- 12121798 TI - Fragmentation of immunization history among providers and parents of children in selected underserved areas. AB - OBJECTIVE: We assessed fragmentation of children's immunization history among providers and parents of children aged 12 to 35 months in four selected underserved areas. STUDY DESIGN: Area probability cluster sample surveys were conducted in 1997-1998 in northern Manhattan, San Diego, Detroit, and rural Colorado. Surveys consisted of face-to-face interviews with parents followed by record checks with all named immunization providers. We used Advisory Committee on Immunization Practices recommendations to determine up-to-date (UTD) status with vaccinations. The UTD status for each child was determined in four ways: (1) according to the parent-held immunization records, (2) according to the records of the child's most recent provider, (3) according to the records of the child's second most recent provider, and (4) according to provider and parent-reconciled information. RESULTS: In all four areas, the majority of records of the most recent provider agreed with the reconciled information. However, in all areas, the percentage of children UTD according to provider- and parent-reconciled information was higher than the percentage of children UTD according to information from only the child's most recent provider or from only parent-held immunization records. Across all sites, the percentage of children UTD with the DTP/DTaP vaccine was 2% to 9% lower, according to the most recent provider's information than according to reconciled information. Similar results were seen for other vaccines. The most recent provider not having complete immunization history was significantly associated with not being UTD in New York and having received unnecessary immunizations in San Diego and Detroit. CONCLUSION: For most children, although the records of the most recent provider give accurate data for clinical decision making, the immunization histories of some children in these underserved areas are fragmented between providers and parents. This can limit the provider's ability to vaccinate children appropriately. PMID- 12121799 TI - Effects of a computer-based, telephone-counseling system on physical activity. AB - BACKGROUND: There is increasing interest in developing interventions to promote physical activity (PA) that do not involve face-to-face contact with health professionals. We developed a fully automated PA counseling system (telephone linked communication, TLC-PA) that was delivered via telephone. DESIGN: A randomized, controlled trial with 298 adult, sedentary members (mean age, 45.9 years; 72% women; 45% white; and 45% African American) of a multi-site medical practice. The comparison group (TLC-Eat) received an automated intervention promoting healthy eating, which was also delivered via telephone. INTERVENTION: The TLC-PA promoted moderate-intensity PA (MI-PA) based on the transtheoretical model of behavior change and social cognitive theory. The system was available to participants for 6 months. MAIN OUTCOMES: Energy expenditure in MI-PA, proportion of participants who met recommendations for MI-PA, and motivational readiness for PA. MEASURES: Self-reports of PA behavior and motivational readiness at baseline, 3 months, and 6 months. RESULTS: At 3 months, intention-to-treat analyses showed that the TLC-PA group was more likely to meet recommendations for MI- or vigorous intensity PA (VI-PA) compared to the TLC-Eat group (TLC-PA=26% vs TLC-Eat=19.6%, p=0.04). Among study completers, TLC-PA subjects reported significantly higher daily kilocalorie energy expenditure in MI-PA (2.3 kcal/kg/d vs 2.0 kcal/kg/d, p=0.02); a larger proportion met recommendations for MI- or VI-PA (31.2% vs 21.3%, p=0.02) and were in more advanced stages of motivational readiness than TLC-Eat subjects (TLC-PA=52.5% vs TLC-Eat=42.2%, p=0.04). Results were not maintained at 6 months. The proportion of TLC-PA users decreased significantly over the intervention period. CONCLUSIONS: A fully automated counseling system had positive short-term effects on PA among sedentary adults. Lack of maintenance of effects may be due to a decrease in the number of participants who continued to use the system. PMID- 12121800 TI - One- and two-year predictors of decline in physical activity among inner-city schoolchildren. AB - BACKGROUND: Alarming secular declines in physical activity (PA) have been observed among youth over the last decade. A better understanding of the predictors of these declines is crucial to identifying those children most at risk and to developing interventions that target youth before the onset of decline. This report identifies 1- and 2-year predictors of decline in PA among fourth- and fifth-grade students from inner-city neighborhoods in Montreal, Canada. METHODS: Data for this study were collected in classroom questionnaires each May/June from 1993 to 1997. Analyses for this paper were completed in 2001. The cohort included active (at least one PA per day) children with baseline and 1 year (n =1873) or 2-year (n =509) follow-up data. RESULTS: In boys, 1-year predictors of decline to an inactive status identified in generalized estimating equations analysis included moderate (vs high) baseline PA (odds ratio [OR]=1.66, 95% confidence interval [CI]=0.91-3.05); low PA self-efficacy (OR=1.67, 95% CI=1.03-2.71); born outside Canada (OR=2.13; 95% CI=1.31-3.46); Asian origin (OR=1.81; 95% CI=1.03-3.16) and no participation in school teams (OR=1.81, 95% CI=0.93-3.55). In girls, these 1-year predictors included moderate PA (OR=1.91, 95% CI=1.10-3.32); low PA self-efficacy (OR=1.70, 95% CI=1.15-2.49); watching four or more TV programs per day (OR=1.40, 95% CI=0.97-2.02); mother unemployed (OR=1.54, 95% CI=1.07-2.23); and grade five (vs grade four) (OR=1.35, 95% CI=0.94 1.93). Two-year predictors in boys included moderate baseline PA (OR=2.52, 95% CI=0.84-7.50), and born outside Canada (OR=1.96, 95% CI=0.91-4.20). In girls, these 2-year predictors included moderate baseline PA (OR=2.75, 95% CI=1.01 7.49); no participation in school teams (OR=2.14, 95% CI=0.92-5.00); watching four or more TV programs per day (OR=1.93, 95% CI=0.99-3.74); and born outside Canada (OR=1.85, 95% CI=0.96-3.55). CONCLUSIONS: Reduced TV viewing among girls and increased participation in school sports teams in boys and girls may prevent declines in PA among pre-adolescents from inner-city neighborhoods. PMID- 12121801 TI - Pedestrian fatalities by race/ethnicity in Arizona, 1990-1996. AB - PURPOSE: To explore rates of pedestrian fatalities in Arizona, and how rates and circumstances of pedestrian deaths differ by race/ethnicity, urban or rural residence, age, and gender. METHODS: Using the Fatality Analysis Reporting System and the National Center for Health Statistics' Multiple Cause of Death file, pedestrian fatalities in Arizona from 1990 through 1996 were classified by gender, race/ethnicity, and urban or rural residence. Age-adjusted rates were calculated and adjusted for the proportion of rural residence. Age analyses compared pedestrian fatality rates in 10-year age groups by race/ethnicity. Conditions associated with pedestrian deaths were examined, including the time and day of occurrence, alcohol involvement, and degree of pedestrian contribution to the crash. RESULTS: American Indians had rates of pedestrian deaths 6 to 13 times those of non-Hispanic whites. Elevated rates for American Indians were found in urban and rural areas, in both genders, in all age groups in men, and in five of nine age groups in women. American-Indian pedestrian death rates and relative risks (RRs) were higher in rural areas than in urban areas. Compared to non-Hispanic whites, urban Hispanic males had an elevated RR of 1.56, rural Hispanic females had an RR of 2.45, and urban African-American (AA) females had an RR of 2.33. However, significantly elevated rates, compared to non-Hispanic whites, were limited to Hispanic males aged <5 years and African-American females aged 65 to 74 years. In all race/ethnic groups, except rural Hispanics, men had higher rates than women, although American-Indian women had higher rates than non Hispanic whites, African Americans, and Hispanic men. Rural residence accounted for 27% of the excess American-Indian pedestrian mortality. Sixty-one percent of urban, American-Indian pedestrian deaths occurred on weekends, compared to 29% among non-Hispanic whites and 46% among Hispanics. American Indians had six times the rate of alcohol-related pedestrian deaths as non-Hispanic whites in urban areas and 16 times that respective rate in rural areas. Hispanics had an alcohol- involvement RR of 1.82 in urban areas, but the RR was not elevated in rural areas. When blood alcohol was measured, the blood alcohol concentration was >0.20 g/dL in 64.4% of American Indians, 35% of Hispanics, and 29% of non-Hispanic whites. CONCLUSION: A major disparity in pedestrian fatalities exists for both American-Indian men and women in urban and rural areas. Other racial/ethnic groups have elevated pedestrian fatality rates that are gender and residence specific, and are limited to specific age groups. Much of the American-Indian excess mortality is alcohol related and associated with residence in rural areas. PMID- 12121802 TI - Obstetrical textbooks: recommendations about drinking during pregnancy. AB - BACKGROUND: For the past 2 decades, public health authorities have recommended that pregnant women abstain from alcohol. We reviewed obstetrical textbooks published over the last 4 decades to identify trends in recommendations for drinking during pregnancy. The study was begun in 2000 and completed in 2001. DESIGN: Eighty-one texts were identified from a national listing service (n =51) and local library shelves (n =30). RESULTS: Only 14 (17%) of the texts contained a consistent recommendation that pregnant women should not drink alcohol. Although there was a slight upward trend toward recommendations for abstinence in more recent texts, only 24% of the 29 texts published after 1990 were in this category. Fifty-three percent of all texts and 52% of texts published after 1990 contained a sentence condoning drinking at some level. The remaining texts (30%) contained no recommendations. CONCLUSIONS: Many texts, even those published recently, have not embraced public health recommendations and, in some instances, contradict them. PMID- 12121806 TI - Sensitization does not develop to cocaine-induced potentiation of the antinociceptive effect of morphine. AB - Repeated intermittent cocaine administration produces a progressive increase (sensitization) in the motor stimulatory action of cocaine. Previous studies have shown that cocaine produces antinociception and also enhances the antinociceptive effect of opioid analgesics. The present study was designed to investigate if sensitization to these effects of cocaine develops. In the first part of the study, we determined if acute cocaine administration (3, 10, 30 mg/kg, intraperitoneal [i.p.]) increases the antinociceptive effect of morphine (5 mg/kg, subcutaneous [s.c.]) in rats using the hot plate test. Cocaine (30 mg/kg, i.p.), alone, produced a small but significant antinociceptive effect at 15 min after drug administration. When administered 15 min prior to morphine, cocaine dose-dependently enhanced the effect of morphine (5 mg/kg, s.c.) at the time (45 min post-cocaine) when cocaine by itself did not significantly change the hot plate latency. In the second part of the study, we examined if sensitization develops to cocaine-induced antinociception and its ability to increase the antinociceptive effect of morphine. Naive rats were injected with either saline or cocaine (30 mg/kg) once daily for 3 days and tested on the hot plate apparatus either 24 h or 1 wk after the last cocaine injection. Some of the rats from each group were also tested for motor stimulation induced by cocaine (5 mg/kg, i.p.) 24 h after the hot plate test to confirm that sensitization had occurred to the motor stimulatory action of the drug. Additional rats were treated with saline or cocaine for 3 days, but neither treated with morphine nor tested on the hot plate apparatus, and tested for behavioral sensitization to the motor stimulatory action of cocaine (5 mg/kg, i.p.) 24 h or 1 wk later. Sensitization developed to the motor stimulatory effect of cocaine in both groups, regardless of morphine treatment on the prior day. Sensitization also developed to the antinociceptive effect of cocaine 24 h but not 1 wk after the last cocaine injection. No sensitization was observed in the ability of cocaine to enhance the antinociceptive effect of morphine. Overall, our data suggest that while cocaine enhanced the antinociceptive effect of morphine, sensitization did not develop to this action of cocaine. PMID- 12121805 TI - Maternal dietary docosahexanoic acid content affects the rat pup auditory system. AB - Previous studies of the effects of dietary docosahexanoic acid (DHA), 22:6n3, on neurodevelopment have focused mainly on visual-evoked potentials and indices of visual activity, measures that may be confounded by effects on the retina rather than on neural pathways. We investigated the effect of pre- and postnatal maternal dietary DHA content on auditory brainstem conduction times (ABCTs), the appearance of the auditory startle reflex (ASR), and 2',3'-cyclic nucleotide 3' phosphodiesterase (CNPase) activity in brainstem homogenates. Timed pregnant dams were fed, beginning on day 2 of gestation and throughout lactation, a purified diet containing one of three levels of DHA (0, 1, or 3% of total fatty acids, or 0, 0.4 or 1.2% of total energy). On postnatal day (PND) 3, pups were randomly crossfostered within diet groups to minimize litter effects and culled to 10 per litter. Cerebrums and milk from culled pups stomachs were collected for lipid analysis. The timing of appearance of the ASR was determined between PND 10 through 14 and ABCTs were measured in pups on PND 24 and 31. Pups were sacrificed on PND 31 and cerebrums were removed. In each of two replicated studies, pups in the 1% DHA group weighed significantly less on PND 3 and they gained significantly less weight from PND 3 to 31 compared with pups in the 0 or 3% groups (p<0.01). The auditory studies were not conducted on the 1% DHA group since measures of auditory function are in part a function of somatic growth. The tissue fatty acid data for the 1% DHA group did not show unexpected findings. Higher dietary DHA was reflected in milk and pup cerebrums, and levels of arachidonic acid were inversely related to levels of DHA. In the pups of dams fed diets containing 3% versus 0% DHA, the ASR appeared significantly later (p<0.001) and the ABCTs were longer (p<0.05) on PND 31. CNPase activity levels were not different between the 0 and 3% DHA groups. This study demonstrated that the auditory brainstem response is sensitive for identifying effects of diet on neurodevelopment, and that diets supplemented with high levels of DHA may exert a negative influence on central nervous system development, potentially through effects on myelin. This study suggests the need for further studies of pre- and postnatal long chain polyunsaturated fatty acid dietary supplementation. PMID- 12121808 TI - Cutaneous vascular responses evoked by noxious stimulation in rats with the spinal nerve ligation-induced model of neuropathy. AB - Antidromic activation of nociceptive nerve fibres innervating the skin produces an axon reflex that involves extravasation and vasodilation of cutaneous blood vessels. We determined whether the axon reflex of the hindlimb skin is influenced by an experimental model of neuropathy induced by unilateral ligation of spinal nerves L(5) and L(6) in the rat. Ligation of spinal nerves induced symptoms mimicking tactile allodynia, as indicated by a marked decrease of the hindlimb withdrawal threshold to mechanical stimulation. The axon reflex induced by antidromic electrical stimulation of nociceptive fibres innervating the plantar skin ipsilateral to the ligation was attenuated according to determination of extravasation response and blood flow response. Lidocaine block or transection of the sciatic nerve of the neuropathic limb did not induce any change in basal blood flow of the plantar skin. The results indicate that ligation of spinal nerves induces an attenuation of the axon reflex. This attenuation reflects a decrease in the efferent function of primary afferent nociceptors innervating the hypersensitive skin of the hindpaw. The attenuation of antidromically-induced vascular responses was not caused by overriding sympathetic activity, as indicated by lack of blood flow effects by lidocaine blocks or a transection of the sciatic nerve. PMID- 12121807 TI - Central nucleus of the amygdala and the control of tonic immobility in guinea pigs. AB - Tonic immobility (TI) is a temporary state of profound motor inhibition induced by situations that supposedly generate intense fear, with the objective to protect the animal from attacks by predators. A previous study by our group demonstrated that cholinergic stimulation of the central, basolateral, and lateral posterior nuclei of the amygdala decreases the duration of TI in guinea pigs. In the current study, we attempted to investigate the effects of cholinergic, opioidergic, and gamma-aminobutyric acid (GABA)ergic stimulation of the central amygdala (CEA) on TI modulation. We observed that both cholinergic (carbachol, 2.7 nmol/0.2 microl) and opioidergic (morphine, 2.2 and 4.4 nmol/0.2 microl) stimulation of the CEA decreased the duration of TI and that these effects could be reversed by pretreatment with naloxone (1.3 and 2.7 nmol/0.2 microl). Our results also showed that microinjection of the GABAergic agonist muscimol (0.26 nmol/0.2 microl) reduced the duration of TI episodes, whereas microinjection of the GABAergic antagonist bicuculline (BIC, 1 nmol/0.2 microl) increased it. Thus, the present experiments demonstrated that cholinergic, opioidergic, and GABAergic systems of the CEA have an inhibitory action on the duration of TI in guinea pigs. Furthermore, the current study suggests an interaction of cholinergic and opioidergic mechanisms. In addition, the GABAergic circuit of the CEA has a tonic inhibitory influence on the duration of TI and is mediated by GABA(A) receptors. PMID- 12121809 TI - Dynamic determination and possible mechanism of amino acid transmitter release from rat spinal dorsal horn induced by the venom and a neurotoxin (BmK I) of scorpion Buthus martensi Karsch. AB - In the present communication, we determined the dynamic release of amino acid transmitters from spinal dorsal horn induced by scorpion Buthus martensi Karsch (BmK) venom and a neurotoxin (BmK I). The results found that glutamate and aspartate release could be evoked significantly within the initial 30 min with the applied doses of either 0.05 and 0.01 mg BmK venom or 0.01 and 0.002 mg BmK I. However, gamma-aminobutyric acid (GABA) release could be largely evoked during the second 30 min by the venom, but not by BmK I. The result suggested that nociceptive afferent fibers could be activated to induce excitatory amino acid release from spinal dorsal horn by nociceptive factors such as BmK I, but the delayed release of GABA might be attributed to the modulating role of some antinociceptive components in the venom. PMID- 12121811 TI - Impaired learning in a spatial working memory version and in a cued version of the water maze in rats with MPTP-induced mesencephalic dopaminergic lesions. AB - A lesion in the substantia nigra pars compacta (SNc) of rats induced by intra nigral administration of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) caused specific loss of dopamine and its nonconjugated metabolites in the dorsal striatum and in the prefrontal cortex (PFC), but not in the hippocampus or the ventral striatum (nucleus accumbens). This lesion did not alter the motor performance of the rats or learning of a spatial reference memory task in the water maze but impaired learning of a spatial working memory task and also of a cued version of the water maze. The results are discussed by relating the selective memory deficits observed in these water maze tasks to the PFC, dorsal striatum, and hippocampus. Some parallels between the memory deficits in these SNc-lesioned rats and Parkinson's disease patients are also discussed. PMID- 12121810 TI - Chemical preconditioning with 3-nitropropionic acid: lack of induction of neuronal tolerance in gerbil hippocampus subjected to transient forebrain ischemia. AB - Chemical preconditioning using the mitochondrial toxin, 3-nitropropionic acid (3 NP) has been reported to induce neuroprotection against subsequent global ischemia. To investigate the underlying mechanisms, Mongolian gerbils were pretreated with either vehicle or 3-NP at the dose of 3 or 10 mg/kg, intraperitoneal, 3 days prior to a 5-min bilateral carotid artery occlusion followed by either 48 h or 7 days of blood recirculation. Neuronal damage was assessed by a cresyl violet/fuchsin acid staining. Induction of heat shock protein 72 (HSP72) and manganese superoxide dismutase (MnSOD) expression was evaluated by Western blotting. Astroglial and microglial activation was detected by immunohistochemistry (glial fibrillary acid protein) and by histochemistry (isolectin B4 staining), respectively. Present data show that the hippocampal neuronal damage induced by ischemia were of similar extent between the vehicle- and 3-NP-treated gerbils, whatever the dose tested, indicating that 3-NP did not induce tolerance to transient forebrain ischemia under our experimental conditions. The lack of difference in the post-ischemic level of HSP72 and MnSOD protein expression and in the intensity of astroglial and microglial activation represents further indirect indications of the absence of 3-NP preconditioning effect. In conclusion, although chemical preconditioning with 3-NP is a well established phenomenon at least in vitro and in models of focal ischemia, the relevance of 3-NP as a preconditioning molecule towards global brain ischemia remains an open question. PMID- 12121812 TI - Changes of cortical epileptic afterdischarges under the influence of convulsant drugs. AB - Convulsant drugs picrotoxin (0.5 and/or 1 mg/kg, intraperitoneal (i.p.)) and pentylenetetrazol (10 and/or 20 mg/kg, i.p.) were used to compromise GABAergic inhibition, caffeine (75 and/or 150 mg/kg, i.p.) to antagonize adenosinergic system to study the role of inhibition in cortical epileptic afterdischarges. Rats with implanted cortical stimulation and registration electrodes were stimulated four times at 10-min intervals, drugs were injected between the first and second stimulation. Four different phenomena were evaluated: movements directly bound to stimulation were intensified by all three drugs, i.e., excitability of the cerebral cortex was increased. Incidence of two types of afterdischarges (spike-and-wave rhythm and "limbic" type) was not changed by any drug, i.e., the transition of epileptic activity into limbic structures was not increased. Afterdischarges were most efficiently prolonged by caffeine, i.e., caffeine probably interferes with mechanism(s) arresting cortical afterdischarges. The intensity of clonic seizures accompanying spike-and-wave afterdischarges, i.e., spread of epileptic activity into the motor system was only transiently increased by picrotoxin, the effects of caffeine did not reach the level of statistical significance. Our results indicate various mechanisms and diverse role of the two inhibitory systems in generation of evaluated phenomena. PMID- 12121813 TI - Translocation of protein kinase C by halothane in cholinergic cells. AB - Protein kinase C (PKC) is a signal transducing enzyme that is an important regulator of multiple physiologic processes and a potential molecular target for volatile anaesthetic actions. However, the effects of these agents on PKC activity are not yet fully understood. Volatile anaesthetics increase intracellular calcium concentration ([Ca(2+)](i)) in a variety of cells, thus their effects on PKC activity may be indirect due to [Ca(2+)](i) increase. Alternatively, the anaesthetics could directly stimulate PKC activity. In order to distinguish these two possibilities in intact cells, we used a fully functional green fluorescent protein conjugated PKCbetaII (GFP-PKCbetaII) and confocal microscopy to evaluate the dynamic redistribution of PKC in living SN56 cells, a cholinergic cell line, in response to halothane. Halothane induced PKC translocation in SN56 cells transfected with GFP-PKCbetaII. This effect was not suppressed by dantrolene, a drug that blocks halothane-induced Ca(2+) release from intracellular stores in these cells. These findings indicate that halothane induces PKC translocation in SN56 cells independently of its ability to release calcium from internal stores. PMID- 12121814 TI - Systemic angiotensin II alters intrinsic heart rate through central mechanisms. AB - Angiotensin II (ang II)-induced increases in intrinsic heart rate (IHR), and the resulting tachycardia, may contribute to development of renal hypertension. Whether circulating ang II affects the cardiac pacemaker through peripheral mechanisms or through actions in the central nervous system (CNS) has not been directly tested. These studies determined the role of a central site of ang II action, the tissue surrounding the anteroventral third ventricle (AV3V), in increased IHR induced by systemic ang II. Blood pressure and heart rate were measured in male rats with lesions of the AV3V region and in control-operated animals during i.v. infusion (3 h) of ang II, norepinephrine, or vehicle. IHR was evaluated at the end of the infusion period. Systemic ang II increased blood pressure equally in both experimental groups. However, heart rate was reduced only in animals with AV3V lesions. Furthermore, ang II increased IHR only in control-operated rats. Changes in blood pressure, heart rate, and IHR in response to norepinephrine infusion were similar between animals with AV3V lesions and control-operated rats. These data demonstrate that systemic ang II mediates IHR through actions in the CNS, specifically the AV3V region. PMID- 12121815 TI - Glial fibrillary acidic protein immunodetection and immunoreactivity in the anterior and posterior medial amygdala of male and female rats. AB - The medial amygdala (MeA) has receptors for gonadal hormones and modulates reproductive behaviors in rats. Adult male and female rats were used for the immunodetection, a less accurate technique, and the immunohistochemistry for the astrocytic marker glial fibrillary acidic protein (GFAP) in the anterior and posterior MeA. Both procedures were done using polyclonal anti-GFAP and were quantified by densitometry. The first technique provided no evidence for a difference between sexes in the immunocontent of GFAP in any region of the MeA (p > 0.1). Nevertheless, the measure of the intensity of GFAP immunoreactivity (GFAP IR) showed that females had a higher GFAP-IR in the posterodorsal (p < 0.01) and in the posteroventral subregions of the MeA (p < 0.01) than males. No sex difference was found in its anterodorsal part (p > 0.1). The present results point out the differences between these two above-mentioned techniques but add a new finding to the previously described sexual dimorphism in the MeA, i.e., the GFAP-IR. Data also suggest that probably astrocytes can be affected by sex steroids in this brain area. It is likely that this regionally specific difference in the GFAP-IR may contribute to the distinct functional roles that the MeA subregions have in male and female rats. PMID- 12121816 TI - Monoamine metabolism and sympathetic nervous activation following subarachnoid haemorrhage: influence of gender and hydrocephalus. AB - Subarachnoid haemorrhage is a serious condition, often accompanied by cerebral vasospasm and hydrocephalus, which may result in delayed cerebral ischaemia and neurological deterioration. While the mechanisms responsible remain unknown, activation of the sympathetic nervous system, leading to elevated levels of circulating catecholamines is, at least in part, implicated. In this study, we sought to examine the importance of sympathetic nervous activation and its relation to brain monoaminergic neurotransmission in 25 patients following subarachnoid haemorrhage by examining plasma and cerebrospinal fluid levels of the catecholamines noradrenaline, adrenaline and dopamine, and their metabolites. Total body sympathetic activity was concurrently assessed using isotope dilution methodology. In the early phase following subarachnoid haemorrhage patients exhibited markedly elevated rates of spillover of noradrenaline to plasma (9.11 +/- 1.12 vs. 3.39 +/- 0.26 nmol/min, p < 0.01), with rates being higher in those patients in whom hydrocephalus developed (11.15 +/- 1.40 vs. 7.90 +/- 1.41 nmol/min, p = 0.05). The degree of sympathetic nervous activation tended to be higher in females compared with males. Lower cerebral perfusion pressures were observed in those patients in whom cerebrospinal fluid concentrations of noradrenaline and dopamine metabolites were high. A marked sympathetic nervous activation, more pronounced in women and in those with hydrocephalus, occurs following subarachnoid haemorrhage. The diminished cerebral perfusion seen following subarachnoid bleeding may occur as a result of activation of central catecholaminergic neurones. PMID- 12121817 TI - Volume expansion suppresses the tooth-pulp evoked jaw-opening reflex related activity of trigeminal neurons in rats. AB - The aim of the present study is to clarify whether physiological stimulation of vagal afferents modulates the activity of the trigeminal spinal nucleus oralis (TSNO) neurons related to the tooth-pulp (TP)-evoked jaw-opening reflex (JOR) in pentobarbital-anesthetized rats. The activity of TSNO neurons and the amplitude of digastric electromyogram (dEMG) increased proportionally during 1.0-3.5 times the threshold for JOR. The amplitude of the dEMG of 14 out of 17 rats was suppressed by physiological stimulation of vagal afferents after intravenous infusion of Ficoll. Out of 23, 18 TSNO unit activities in 14 rats were also suppressed by Ficoll infusion. This suppressive effect of unit and dEMG activities returned to the control level within 25 min. After administration of naloxone (0.5 and 1.0 mg/kg, i.v.) the suppressive effect of Ficoll infusion on the activity of TSNO neurons (5/7) was significantly attenuated compared to the control (p < 0.01). The inhibition TSNO neuronal and dEMG activities by Ficoll infusion was volume-dependent in a range of 5-10% of total blood volume. Furthermore, right vagus nerve ligation greatly inhibited the suppressive effect of Ficoll-induced TSNO activity. These results therefore suggest that low pressure cardiopulmonary baroreceptors whose afferents travel in the vagus nerve inhibit the pulpal nociceptive transmission. PMID- 12121819 TI - Pregnenolone sulfate, dehydroepiandrosterone sulfate and allotetrahydrodeoxycorticosterone affect rapid tolerance to the hypothermic effect of ethanol. AB - Our previous study showed that the neurosteroids pregnenolone sulfate (PS) and epipregnanolone stimulated and blocked, respectively, the demonstration of chronic tolerance to the incoordinating effect of ethanol. The aim of the present study was to investigate the influence of three neurosteroids on the demonstration of tolerance to ethanol-induced hypothermia in mice using the rapid tolerance paradigm. The first experiment defined the doses of ethanol that did or did not induce rapid tolerance to ethanol-induced hypothermia. In the second, the influence of pretreatment of mice with PS (0.08 or 0.15 mg/kg, i.p.) or dehydroepiandrosterone sulfate (DHEAS; 0.15 or 0.20 mg/kg, i.p.) before ethanol (4.0 g/kg, i.p.) on rapid tolerance was studied. The third experiment examined the effect of allotetrahydrodeoxicorticosterone (ALLOT; 0.10 or 0.20 mg/kg, i.p.) before ethanol (4.0 g/kg, i.p.) on rapid tolerance. Results showed that pretreatment with PS or with DHEAS significantly facilitated the demonstration of rapid tolerance, whereas pretreatment with ALLOT interfered with the demonstration of tolerance to the hypothermic effect. PMID- 12121818 TI - Evidence that toxicity of lipopolysaccharide upon cholinergic basal forebrain neurons requires the presence of glial cells in vitro. AB - The cholinergic system of the basal forebrain is affected in brains of dementia patients and during neuroinflammation. The aim of this study was to establish a method to cultivate basal forebrain cholinergic neurons in dissociated, pure neuronal cultures and to apply this method to study the effect of acute and chronic experimentally-induced inflammation using lipopolysaccharide. Purity of the cultures, degrees of neuronal dissociation, connectivity and neuronal survival were investigated by immunocytochemistry for microtubule-associated protein-2 (neurons), glial fibrillary acidic protein (astroglia), complement receptor 3 (microglia), choline acetyltransferase and the neurotrophin receptor p75 (cholinergic neurons). Neuronal cultures only contained <7% astrocytes and <1% microglia when using a "sandwich-technique". Acute (1, 10 microg/ml) as well as chronic (0.1, 1 microg/ml) treatment with lipopolysaccharide did neither affect total number of neurons, nor number of p75-positive neurons or enhance expression of major histocompatibility complex I or II. Our results suggest that lipopolysaccharide-induced degeneration of both microtubule-associated protein-2 like immunoreactive as well as specific killing of cholinergic forebrain neurons in vitro are mediated by glial cells. PMID- 12121820 TI - Medial versus lateral parabrachial nucleus lesions in the rat: effects on mercaptoacetate-induced feeding and conditioned taste aversion. AB - The two experiments of the present study examined the influence of bilateral electrophysiologically-guided ibotenic acid lesions of the medial (gustatory) and lateral (viscerosensory) subdivisions of the parabrachial nucleus (PBN) on lipoprivic feeding and on the acquisition of a conditioned taste aversion. In Experiment 1, mercaptoacetate (0, 400, 600, or 800 micromol/kg) failed to enhance food intake in normal rats maintained and tested on standard laboratory chow. In the same procedure, rats with lesions of the medial or lateral PBN consumed less food during baseline but nonetheless were sensitive to the orexigenic action of mercaptoacetate. In Experiment 2, both types of PBN lesions prevented acquisition of a conditioned taste aversion induced by the oral self administration of lithium chloride. The results suggest that PBN neurons essential for conditioned taste aversion are not involved in the mercaptoacetate-induced feeding of rats maintained and tested on standard laboratory chow. PMID- 12121821 TI - Epileptiform spikes desynchronize and diminish fast (gamma) activity of the brain. An "anti-binding" mechanism? AB - Fast (20-100 Hz) rhythms of electrical activity of the brain have been suggested to be important for perception and cognition providing a mechanism for temporal binding of neural activities underlying mental representations. Also, fast rhythms often precede epileptiform discharges in patients and some experimental models. Generalized slow (2-3 Hz) spike activity after systemic kainic acid (KA) in the rat has been shown to be preceded by intense gamma activity. A relationship between the intensified gamma rhythms and the subsequent spike activity was studied during kainate-induced acute epileptogenesis. Power, multiple coherence and phase were analyzed at frequencies 1-100 Hz in the EEG recorded from the hippocampal-neocortical structures of the rat. Gamma rhythms, extremely intense and highly coherent at the onset of discharges, were followed by a slow rhythm of epileptiform spikes/sharp waves. During this spike activity and immediately afterwards, the gamma power and coherence were significantly decreased. These data show an antagonism between gamma rhythms and spike activity and ability of the latter to desynchronize and suppress the former. They are supportive to the hypothesis that epileptiform spike activity may result from the extreme activation of the "anti-binding" mechanism controlling temporal binding at high frequencies. It is suggested that when fast activity is abnormally intensified, "over-binding" with global synchrony of gamma rhythms can occur in the neural networks. It may lead to inadequate synaptic modifications. To prevent this process, epileptiform discharge develops as a protective mechanism suppressing fast activity. This proposal has implications for our understanding of temporal binding in the brain and how its excessive activation may precipitate the development of pathological states. PMID- 12121822 TI - Differential discharge patterns of rhythmical activity in trigeminal motoneurons during fictive mastication and respiration in vitro. AB - Rhythmical activity in trigeminal motoneurons (TMNs) was studied in an in vitro neonatal rat brainstem preparation that retains functionally active circuits for oral-motor behaviors. Whole-cell current-clamp recording from TMNs demonstrated rhythmical activities during both spontaneously generated respiratory activity and neurochemically induced rhythmical oral-motor activity. TMNs showed spontaneous rhythmical (0.08 +/- 0.04 Hz) activities of burst-firing pattern during inspiration synchronized with inspiratory activities recorded in hypoglossal nerves. During rhythmical oral-motor activity induced by bath application of N-methyl-d,l-aspartic acid and the GABA(A) receptor antagonist bicuculline methiodide, TMNs showed only a rhythmical (5.6 +/- 0.8 Hz) pattern of single-spike discharge. TMNs never showed a burst-firing pattern during rhythmical oral-motor activity even when membrane potentials were shifted either to depolarized or hyperpolarized levels. Rhythmical activity in TMNs exhibited different discharge patterns between rhythmical oral-motor activity and respiratory activity generated in vitro. PMID- 12121824 TI - Antiangiogenic mechanisms of diet-derived polyphenols. AB - Accumulating evidence demonstrates that polyphenols in natural products are beneficial against human lethal diseases such as cancer and metastasis. The underlying mechanisms of anti-cancer effects are complex. Recent studies show that several polyphenols, including epigallocatechin-3-gallate (EGCG) in green tea and resveratrol in red wine, inhibit angiogenesis when administrated orally. These polyphenols have direct effects on suppression of angiogenesis in several standard animal angiogenesis models. Because angiogenesis is involved in many diseases such as cancer, diabetic retinopathy and chronic inflammations, the discovery of these polyphenols as angiogenesis inhibitors has shed light on the health beneficial mechanisms of natural products, which are rich in these molecules. At the molecular level, recent studies have provided important information on how these molecules inhibit endothelial cell growth. Perhaps the greatest therapeutic advantage of these small natural molecules over large protein compounds is that they can be administrated orally without causing severe side effects. It is anticipated that more polyphenols in natural products will be discovered as angiogenesis inhibitors and that these natural polyphenols could serve as leading structures in the discovery of more potent, synthetic angiogenesis inhibitors. PMID- 12121823 TI - Effect of cognitive load on postural control. AB - The present study reports findings from two experiments on the relation between a mental task (silent backward counting) and posture. The first experiment included 30 normal subjects and the second experiment 20 normal subjects. In Experiment 1 postural sway and performance of the mental task were measured in a 2 x 2 dual task design (with or without mental task and calf stimulation). In Experiment 2 a similar design was used, the only difference being that during trials without the mental task, subjects were instructed to focus on their balance and provide a rating of body sway. Results showed that balance perturbation led to decreased performance on the cognitive task in Experiment 1, but not in Experiment 2. The mental task led to less body sway, while focused attention attenuated the effect. In conclusion, control of body sway and cognitive functioning are to some extent related. PMID- 12121825 TI - Dietary fat and atherosclerosis. PMID- 12121826 TI - Decreased aortic early atherosclerosis in hypercholesterolemic hamsters fed oleic acid-rich TriSun oil compared to linoleic acid-rich sunflower oil. AB - Previous studies have demonstrated that low density lipoprotein (LDL) enriched in polyunsaturated fatty acids (PUFA) are more susceptible to oxidation (ex vivo) than those containing monounsaturated fatty acids (MUFA). To test whether this observation was associated with various parameters considered to be related with the development of early aortic atherosclerosis, hamsters were fed commercial hypercholesterolemic diets (HCD) containing either the PUFA, sunflower oil (SF) or the MUFA, TriSun oil (TS) at 10% with 0.4% cholesterol (wt/wt). LDL isolated from hamsters fed TS had significantly longer lag phase (30%, P < 0.05), a decreased propagation phase (-62%, P < 0.005), and fewer conjugated dienes formed (-37%, P < 0.007) compared to hamsters fed SF. Aortic vasomotor function, measured as degree of aortic relaxation, was significantly greater in the TS vs SF-fed hamsters whether acetylcholine or the calcium ionophore A23187 was used as the endothelium-dependent agonist. As a group, the SF-fed hamsters had significantly more early atherosclerosis than hamsters fed TS (46%, P < 0.006). When animals across the two diets were pair-matched by plasma LDL-C levels, there was an 82% greater mean difference (P < 0.002) in early atherosclerosis in the SF versus the TS-fed hamsters. While there were no significant associations with plasma lipids and lipoprotein cholesterol, early atherosclerosis was significantly correlated with lag phase (r = -0.67, p < 0.02), rate of LDL conjugated diene formation (r = 0.74, p < 0.006) and maximum dienes formed (r = 0.67, p < 0.02). Compared to TS-fed animals, aortic sections from hamsters fed the SF-containing diet revealed that the cytoplasm of numerous foam cells in the subendothelial space reacted positively with the monoclonal anti-bodies MDA-2 and NA59 antibody, epitopes found on oxidized forms of LDL. The present study suggests that compared to TS, hamsters fed the SF-diet demonstrated enhanced LDL oxidative susceptibility, reduced aortic relaxation, greater early aortic atherosclerosis and accumulation of epitopes found on oxidized forms of LDL. PMID- 12121827 TI - Dietary orotic acid affects antioxidant enzyme mRNA levels and oxidative damage to lipids and proteins in rat liver. AB - We investigated the effects of the dietary addition of orotic acid on liver antioxidant enzymes, mRNA levels of these enzymes, and peroxidative products by comparing casein with soy protein as the source of dietary protein. Rats fed the casein diet accumulated more liver lipids than those fed the soy protein diet when orotic acid was added. The addition of orotic acid lowered both the activity of liver Cu, Zn-superoxide dismutase and the level of Cu, Zn-superoxide dismutase mRNA. The addition of orotic acid led to a significant increase in the contents of conjugated dienes and protein carbonyls in the liver. In addition, dietary soy protein protected the increase in the levels of lipids and proteins peroxide induced by orotic acid. The addition of orotic acid to the casein diet increased the activities of both serum ornithine carbamoyltransferase and alanine aminotransferase. Thus, liver damage might result from the increased superoxide anion due to the decrease in the activity of hepatic superoxide dismutase, as well as increase in the production of hepatic peroxidative products in rats fed the casein diet with orotic acid. PMID- 12121828 TI - Norbixin ingestion did not induce any detectable DNA breakage in liver and kidney but caused a considerable impairment in plasma glucose levels of rats and mice. AB - From the seeds of Bixa orellana are extracted the carotenoids bixin and norbixin that have been widely used for coloring food. In this study, the toxicity of norbixin, purified or not (annatto extract containing 50% norbixin), was investigated in mice and rats after 21 days of ingestion through drinking water. Mice were exposed to doses of 56 and 351 mg/kg (annatto extract) and 0.8, 7.6, 66 and 274 mg/kg (norbixin). Rats were exposed to doses of 0.8, 7.5 and 68 mg/kg (annatto extract) and 0.8, 8.5 and 74 mg/kg (norbixin). In rats, no toxicity was detected by plasma chemistry. In mice, norbixin induced an increase in plasma alanine aminotransferase activity (ALT) while both norbixin and annatto extract induced a decrease in plasma total protein and globulins (P < 0.05). However, no signs of toxicity were detected in liver by histopathological analysis. No enhancement in DNA breakage was detected in liver or kidney from mice treated with annatto pigments, as evaluated by the comet assay. Nevertheless, there was a remarkable effect of norbixin on the glycemia of both rodent species. In rats, norbixin induced hyperglycemia that ranged from 26.9% (8.5 mg/kg norbixin, to 52.6% (74 mg/kg norbixin, P < 0.01) above control levels. In mice, norbixin induced hypoglycemia that ranged from 14.4% (0.8 mg/kg norbixin, P < 0.05) to 21.5% (66 mg/kg norbixin, P < 0.001) below control levels. Rats and mice treated with annatto pigments showed hyperinsulinemia and hypoinsulinemia, respectively indicating that pancreatic beta-cells were functional. More studies should be performed to fully understand of how species-related differences influences the biological fate of norbixin. PMID- 12121829 TI - Effects of genistein and daidzein on the cell growth, cell cycle, and differentiation of human and murine melanoma cells(1). AB - Genistein and daidzein are two major isoflavonoids in dietary soybean that have inhibition effect on the cell growth of different tumor cell lines. We previously reported the anti-tumor activities of genistein and daidzein in human co1on tumor (HCT) cells and their different ability to enhance the activation of murine lymphocytes. In the present study, the effect of genistein and daidzein on the cell growth, cell cycle progression, and differentiation of murine K1735M2 and human WM451 cel1s was investigated. It was found that genistein could inhibit the cell growth of two metastatic melanoma cell lines, murine Kl735M2 and human WM45l in a dose-dependent manner. Flow cytometry showed that genistein could cause arrest of both Kl735M2 and WM45l at G(2)/M phase, while daidzein increased the cell numbers at S phase, decreased the cell numbers at G(1) phase. Detection of melanin and morphological observation showed that genistein can induce Kl735M2 and WM45l to produce dendrite-like structure and produce more melanin by 80%. In contrast, daidzein only retarded the growth of K1735M2 and did not induce differentiation in either K1735M2 or WM451. These results suggest that genistein and daidzein in soybean can inhibit certain malignant phenotype of melanoma via different mechanisms and be potential medical candidates for melanoma cancer therapy. PMID- 12121830 TI - Effects of beta-carotene, vitamin C and E on antioxidant status in hyperlipidemic smokers. AB - Smoking can accelerate the consumption of the stored antioxidant vitamins and increase the oxidative stress in the hyperlipidemic patients. The study investigated the effects of combined beta-carotene, vitamin C, and vitamin E on plasma antioxidant levels, erythrocyte antioxidative enzyme activities, and LDL lipid peroxides. Male hyperlipidemic smokers (35-78 years old) were randomly divided into two antioxidant supplemented groups: intervention 1 (I1, n = 22) (15 mg beta-carotene/day, 500 mg vitamin C/day, and 400 mg alpha-tocopherol equivalent/day) and intervention 2 (I2, n = 20) (30 mg beta-carotene/day, 1000 mg vitamin C/day, and 800 mg alpha-tocopherol equivalent/day). After 6-week supplementation, plasma beta-carotene, vitamin C, vitamin E, and erythrocyte glutathione levels increased significantly by 200%, 98%, 129%, and 39%, respectively, in the I1 group, and by 209%, 216%, 197%, and 32%, respectively, in the I2 group. Plasma Fe(+2) concentrations and Fe(+2)/Fe(+3) decreased significantly in both groups. Except erythrocyte glutathione peroxidase activity in the I1 group, erythrocyte catalase, glutathione peroxidase, and superoxide dismutase activities increased significantly in both groups. Lipid peroxides in LDL decreased significantly by 56% and 72% in the I1 and I2 groups, respectively. However, the levels of plasma iron, erythrocyte glutathione, and LDL lipid peroxides, and the activities of erythrocyte antioxidative enzymes did not differ between two groups. In conclusion, combined antioxidant supplements increased plasma antioxidant levels and antioxidative enzyme activities, and lowered LDL lipid peroxides in male hyperlipidemic smokers. Higher dosage of the supplements did not have an additive effect. PMID- 12121831 TI - The effect of dietary menhaden, olive, and coconut oil fed with three levels of vitamin E on plasma and liver lipids and plasma fatty acid composition in rats. AB - The effect of dietary fats with varying degrees of unsaturation in the presence of different concentrations of vitamin E on tissue lipid levels was studied in rats. Rats were fed either menhaden oil, olive oil or coconut oil at 15% levels with either 0.1, 0.3 or 0.6 mg/g of vitamin E as alpha-tocopherol for four weeks. Rat serum and liver were analyzed for total cholesterol, HDL-cholesterol, triacylglycerol and phospholipids. In addition, fatty acid composition of serum lipids was also analyzed. Serum total cholesterol and triacylglycerol were significantly lower in rats fed menhaden oil than in those fed olive or coconut oil, while the HDL-cholesterol was significantly higher in serum of rats fed menhaden and olive oil than in those fed coconut oil. Levels of vitamin E in the diet had only a significant effect on serum cholesterol and liver phospholipids. The Pearson correlation coefficient showed a significant positive relationship between serum triacylglycerol and total cholesterol, and a negative correlation between triacylglycerol and HDL-cholesterol, and between total and HDL cholesterol.In the liver, total cholesterol was significantly higher in rats fed coconut oil than in rats fed menhaden oil. Total liver phospholipids were lower in rats fed either coconut oil or olive oil compared to those fed menhaden oil, especially with higher levels of vitamin E intake. Higher levels of vitamin E in the diet appear to increase triacylglycerol and phospholipids in livers of rats fed menhaden oil. In the liver a significant negative correlation was observed between phospholipids and cholesterol. The type and degree of unsaturation (polyunsaturated fatty acids in menhaden oil, monounsaturated fatty acids in olive oil and saturated fatty acids in coconut oil) significantly affected plasma and tissue lipids. PMID- 12121832 TI - The development and clinical use of trastuzumab (Herceptin). AB - HER-2 is a member of the c-erbB family of receptor tyrosine kinases and is overexpressed by 20-30% of human breast cancers. HER-2 overexpression is an independent adverse prognostic factor and may also predict for response to both chemotherapy and endocrine agents. Trastuzumab is a humanised monoclonal antibody that binds with high affinity to the extracellular domain of HER-2. In HER-2 overexpressing preclinical models trastuzumab has been shown to have a marked antiproliferative effect and demonstrates synergy with a number of cytotoxic drugs. Several phase II and phase III clinical trials have now been performed in patients with advanced breast cancer that overexpress HER-2. Trastuzumab was initially shown to be active and well tolerated as a single agent in heavily pretreated women. Subsequently, studies of first-line treatment for metastatic breast cancer have demonstrated an improvement in survival for trastuzumab when used in combination with either paclitaxel or an anthracycline-cyclophosphamide regimen compared with chemotherapy alone. Unexpectedly, the combination of trastuzumab and the anthracycline-containing regimen was associated with a significant incidence of cardiac dysfunction. The benefit of trastuzumab is generally confined to patients whose tumours have gene amplification as detected by fluorescence in situ hybridisation (FISH) and this is tightly associated with immunohistochemical (IHC) staining at the highest (3+) level. A small number of patients have IHC 2+ tumours together with FISH evidence of gene amplification and may also derive benefit from treatment. Trastuzumab has also been shown to be effective when used as first-line monotherapy for advanced breast cancer. Trials to date have employed trastuzumab in a weekly schedule, but there is emerging evidence that a three-weekly regimen may be as effective. Trastuzumab has shown encouraging activity when used with other agents including docetaxel and vinorelbine. The combination of trastuzumab, docetaxel, and platinum salts also appears to be very active. The role of trastuzumab as adjuvant therapy for early breast cancer is being tested in a number of large randomised trials. PMID- 12121833 TI - Vitamin A and apoptosis in prostate cancer. AB - Apoptosis represents an effective way to eliminate cancer cells. Unfortunately, advanced prostate tumors eventually progress to androgen-independent tumors, which are resistant to current therapeutic approaches that act by triggering apoptosis. Vitamin A and its natural and synthetic analogs (retinoids) induce apoptosis in prostate cancer cells in vitro and in animal models, mainly through induction of retinoic acid receptor-beta (RARbeta). Expression levels of RARbeta, however, are significantly reduced in hormone-independent prostate cancer cells. Recently, a new class of synthetic retinoids related to 6-[3-(1-adamantyl)-4 hydroxyphenyl]-2-naphthalene carboxylic acid (AHPN) (also called CD437) that effectively induces apoptosis of both hormone-dependent and -independent prostate cancer cells in a retinoid receptor-independent manner was identified and has drawn a lot of attention in the field. The apoptotic effect of AHPN requires expression of orphan receptor TR3 (also called nur77 or NGFI-B). Paradoxically, TR3 expression is also induced by androgen and other mitogenic agents in prostate cancer cells to confer their proliferation. The recent finding that TR3 migrates from the nucleus to mitochondria to trigger apoptosis in response to AHPN suggests that the opposing biological activities of TR3 are regulated by its subcellular localization. Thus, agents that induce translocalization of TR3 from the nucleus to mitochondria will have improved efficacy against prostate cancer. TR3, therefore, represents an unexplored molecule that may be an ideal target for developing new agents for prostate cancer therapy. PMID- 12121834 TI - A critical reappraisal of MIB-1 labelling index significance in a large series of pituitary tumours: secreting versus non-secreting adenomas. AB - Pituitary tumours are usually benign neoplasia, but may have a locally aggressive or malignant evolution. This study aimed to identify factors which mostly influence their proliferative activity, in order to clarify its value for clinical and research purposes. The proliferative index was determined in a prospective series of 132 pituitary tumours as the percentage of monoclonal antibody MIB-1-immunopositive cells and referred to as the MIB-1 labelling index (LI). Its distribution was analysed according to both univariate and multivariate models. A life-threatening pituitary tumour is presented separately. The mean LI was 1.24+/-1.59%, with significant differences between clinically secreting (CS) and clinically non-secreting (CNS) adenomas. In CS adenomas (n=65), LI was highly variable and markedly influenced by pre-operative pharmacological treatment (0.80+/-1.03 vs 2.06+/-2.39% in treated vs untreated cases, P=0.009); it decreased with patient's age (P=0.025, r=0.28) and increased with tumour volume and invasiveness. The influence of pre-operative treatment and macroscopic features on LI in this group was confirmed by multivariate analysis. In CNS adenomas (n=67), LI distribution was less variable than in CS adenomas (P<0.0001), it was age-independent and correlations with tumour volume, invasiveness or recurrence did not reach significance. In a rapidly growing parasellar tumour, the mean LI was 24% at first surgery and exceeded 50% at second surgery performed 4 months later. LI should be interpreted according to hormone secretion and pre-operative treatment. Unusually high LI values deserve particular attention. PMID- 12121835 TI - Gene therapy of prostate cancer: current and future directions. AB - Prostate cancer (PCA) is the second most common cause of death from malignancy in American men. Developing new approaches for gene therapy for PCA is critical as there is no effective treatment for patients in the advanced stages of this disease. Current PCA gene therapy research strategies include cytoreductive approaches (immunotherapy and cytolytic/pro-apoptotic) and corrective approaches (replacing deleted or mutated genes). The prostate is ideal for gene therapy. It is an accessory organ, offers unique antigens (prostate-specific antigen, prostate-specific membrane antigen, human glandular kallikrein 2 etc.) and is stereotactically accessible for in situ treatments. Viral and non-viral means are being used to transfer the genetic material into tumor cells. The number of clinical trials utilizing gene therapy methods for PCA is increasing. We review the multiple issues involved in developing effective gene therapy strategies for human PCA and early clinical results. PMID- 12121836 TI - Magnetic resonance imaging for primary breast cancer management: current role and new applications. AB - Techniques for magnetic resonance (MR) imaging of the breast have been evolving over the past decade. The opportunities for integration of MR imaging into clinical breast cancer management and clinical research are increasing. In this paper, we will review the principles behind the creation of standard and MR images and use this as a platform to evaluate clinical studies and indications for the use and study of MR. In particular, we will focus on those areas where MR has the capability of changing care and/or improving our understanding of the biology of breast cancer. In addition, we will address areas where MR is not yet capable of adding value or where MR may lead to unnecessary procedures. PMID- 12121837 TI - Guiding principles for research involving animals and human beings. PMID- 12121838 TI - Baroreceptor reflex function. PMID- 12121839 TI - Physiological significance of alpha(2)-adrenergic receptor subtype diversity: one receptor is not enough. AB - Alpha(2)-adrenergic receptors mediate part of the diverse biological effects of the endogenous catecholamines epinephrine and norepinephrine. Three distinct subtypes of alpha(2)-adrenergic receptors, alpha(2A), alpha(2B), alpha(2C), have been identified from multiple species. Because of the lack of sufficiently subtype-selective ligands, the specific biological functions of these receptor subtypes were largely unknown until recently. Gene-targeted mice carrying deletions in the genes encoding for individual alpha(2)-receptor subtypes have added important new insight into the physiological significance of adrenergic receptor diversity. Two different strategies have emerged to regulate adrenergic signal transduction. Some biological functions are controlled by two counteracting alpha(2)-receptor subtypes, e.g., alpha(2A)-receptors decrease sympathetic outflow and blood pressure, whereas the alpha(2B)-subtype increases blood pressure. Other biological functions are regulated by synergistic alpha(2) receptor subtypes. The inhibitory presynaptic feedback loop that tightly regulates neurotransmitter release from adrenergic nerves also requires two receptor subtypes, alpha(2A) and alpha(2C). Similarly, nociception is controlled at several levels by one of the three alpha(2)-receptor subtypes. Further investigation of the specific function of alpha(2)-subtypes will greatly enhance our understanding of the relevance of closely related receptor proteins and point out novel therapeutic strategies for subtype-selective drug development. PMID- 12121840 TI - Phosphate transport in the duodenum and jejunum of goats and its adaptation by dietary phosphate and calcium. AB - Endogenous P(i) recycling is a characteristic feature of the P homeostasis in ruminants. A pronounced salivary P(i) secretion into the rumen is balanced by a high intestinal P(i) absorption and an almost complete renal P(i) reabsorption. In monogastric animals, the major P(i) transport mechanism across the apical membrane of the enterocyte is an Na(+)-dependent transport mediated by NaPi cotransporter type IIb. In ruminants, an Na(+)-, as well as an H(+)-dependent, P(i) transport system seems to exist in the small intestines. Therefore, morphological localization, type of ionic dependence, and ability to adapt to dietary P or Ca restriction of duodenal and jejunal P(i) transport were characterized in goats. In the duodenum, there was an H(+)-dependent, Na(+) sensitive P(i) transport system that did not belong to the NaPi type II family and was not influenced by dietary P or Ca restriction. In contrast, in the jejunum, there was an Na(+)-dependent, H(+)-sensitive P(i) transport mainly mediated by NaPi IIb. P restriction stimulated the NaPi IIb protein expression, resulting in higher P(i) transport capacity. PMID- 12121841 TI - Vascular responses in vivo to 8-epi PGF(2alpha) in normal and hypercholesterolemic pigs. AB - Hypercholesterolemia (HC) is characterized by increased circulating 8-epi prostaglandin-F(2alpha) (isoprostane), a vasoconstrictor, marker, and mediator of increased oxidative stress, whose vascular effects might be augmented in HC. Anesthetized pigs were studied in vivo with electron beam computed tomography after a 12-wk normal (n = 8) or HC (n = 8) diet. Mean arterial pressure (MAP), single-kidney perfusion, and glomerular filtration rate (GFR) were quantified before and during unilateral intrarenal infusions of U46619 (10 ng x kg(-1) x min(-1)) or isoprostane (1 microg x kg(-1) x min(-1)). Basal renal perfusion and function were similar, and isoprostane infusion elevated its systemic levels similarly in normal and HC (333 +/- 89 vs. 366 +/- 48 pg/ml, respectively, P < 0.01 vs. baseline). Both drugs markedly and comparably decreased cortical perfusion and GFR in both groups, whereas medullary perfusion decreased significantly only in HC. Moreover, MAP increased significantly only in HC (+9 +/ 3 and +11 +/- 3 mmHg, respectively, P130 diverse skeletal muscles were analyzed to detect common elements that account for variability in shortening velocity and force production. Body mass was found to be a significant predictor of shortening velocity in terrestrial and flying animals, with smaller animals possessing faster muscles. Although previous studies of terrestrial mammals revealed similar trends, the current study indicates that this pattern is more universal than previously appreciated. In contrast, shortening velocity in muscles used for swimming and nonlocomotory functions is not significantly affected by body size. Although force production is more uniform than shortening velocity, a significant correlation with shortening velocity was detected in muscles used for locomotion, with faster muscles tending to produce more force. Overall, the contractile properties of skeletal muscles are conserved among phylogenic groups, but have been significantly influenced by other factors such as body size and mode of locomotion. PMID- 12121851 TI - Developmental changes in cardiac recovery from anoxia-reoxygenation. AB - The developing cardiovascular system is known to operate normally in a hypoxic environment. However, the functional and ultrastructural recovery of embryonic/fetal hearts subjected to anoxia lasting as long as hypoxia/ischemia performed in adult animal models remains to be investigated. Isolated spontaneously beating hearts from Hamburger-Hamilton developmental stages 14 (14HH), 20HH, 24HH, and 27HH chick embryos were subjected in vitro to 30 or 60 min of anoxia followed by 60 min of reoxygenation. Morphological alterations and apoptosis were assessed histologically and by transmission electron microscopy. Anoxia provoked an initial tachycardia followed by bradycardia leading to complete cardiac arrest, except for in the youngest heart, which kept beating. Complete atrioventricular block appeared after 9.4 +/- 1.1, 1.7 +/- 0.2, and 1.6 +/- 0.3 min at stages 20HH, 24HH, and 27HH, respectively. At reoxygenation, sinoatrial activity resumed first in the form of irregular bursts, and one-to-one atrioventricular conduction resumed after 8, 17, and 35 min at stages 20HH, 24HH, and 27HH, respectively. Ventricular shortening recovered within 30 min except at stage 27HH. After 60 min of anoxia, stage 27HH hearts did not retrieve their baseline activity. Whatever the stage and anoxia duration, nuclear and mitochondrial swelling observed at the end of anoxia were reversible with no apoptosis. Thus the embryonic heart is able to fully recover from anoxia/reoxygenation although its anoxic tolerance declines with age. Changes in cellular homeostatic mechanisms rather than in energy metabolism may account for these developmental variations. PMID- 12121852 TI - Heat acclimation in rats: modulation via lipid polyunsaturation. AB - Heat acclimation of rats has been shown to enhance endurance of rat hearts to ischemic insult and acute heat stress. Common protective features have been shown to be operative during both these stress-inducing conditions. To explore the role of membrane lipid composition in the adaptive response, we analyzed two major parameters that impact membrane dynamics and order, the nonesterified cholesterol levels and the acyl chain composition of phospholipids, in rat heart and salivary glands, both major thermoregulatory organs, in short- and long-term heat acclimated rats. Before exposure to heat, control salivary gland tissue has a higher cholesterol-to-phospholipid mole ratio (0.32 +/- 0.02) than heart (0.14 +/ 0.01), and the acyl chains of its phospholipids are 50% more saturated. The remodeling strategies of the tissues after exposure to heat differed. Heart cholesterol levels increased after short-term heat acclimation (approximately 50%), whereas salivary gland cholesterol levels decreased in acute heat stress and long-term heat acclimation (approximately 32%). Remodeling of phospholipid acyl chains, particularly an increase in docosahexaenoic acid, was a protective strategy in both tissues (57% in heart and >100% in salivary glands). Modifying membrane lipid composition by treating rats with liposomes composed of egg phosphatidylcholine (PC) before exposure to heat resulted in a 38% increase in endurance to thermal stress. The density and affinity of muscarinic receptors of submaxillary salivary glands, involved in the acclimation response, were measured in control and PC liposome-treated rats, and then both groups were subjected to short-term heat acclimation. After PC treatment the well-established compensatory upregulation of the muscarinic receptors and concomitant decrease in their affinity was blunted. The substantial increase in the thermal endurance of heat challenged intact rats after treatment with PC liposomes (600 vs. 200 min) suggests that membrane lipid composition plays a role in the ability of these tissues to respond to heat stress. PMID- 12121853 TI - Role of corticotropin-releasing hormone in stressor-induced alterations of sleep in rat. AB - Corticotropin-releasing hormone (CRH) mediates responses to a variety of stressors. We subjected rats to a 1-h period of an acute stressor, physical restraint, and determined the impact on subsequent sleep-wake behavior. Restraint at the beginning of the light period, but not the dark period, increased waking and reduced rapid eye movement sleep without dramatically altering slow-wave sleep (SWS). Electroencephalogram (EEG) slow-wave activity during SWS and brain temperature were increased by this manipulation. Central administration of the CRH receptor antagonist astressin blocked the increase in waking after physical restraint, but not during the period of restraint itself. Blockade of CRH receptors with astressin attenuated the restraint-induced elevation of brain temperature, but not the increase of EEG slow-wave activity during subsequent SWS. Although corticosterone increased after restraint in naive animals, it was not altered by this manipulation in rats well habituated to handling and injection procedures. These results suggest that under these conditions central CRH, but not the hypothalamic-pituitary-adrenal axis, is involved in the alterations in sleep-wake behavior and the modulation of brain temperature of rats exposed to physical restraint. PMID- 12121854 TI - Functional properties of human muscle fibers after short-term resistance exercise training. AB - The aim of this study was to assess the relationships between human muscle fiber hypertrophy, protein isoform content, and maximal Ca(2+)-activated contractile function following a short-term period of resistance exercise training. Six male subjects (age 27 +/- 2 yr) participated in a 12-wk progressive resistance exercise training program that increased voluntary lower limb extension strength by >60%. Single chemically skinned fibers were prepared from pre- and posttraining vastus lateralis muscle biopsies. Training increased the cross sectional area (CSA) and peak Ca(2+)-activated force (P(o)) of fibers containing type I, IIa, or IIa/IIx myosin heavy chain by 30-40% without affecting fiber specific force (P(o)/CSA) or unloaded shortening velocity (V(o)). Absolute fiber peak power rose as a result of the increase in P(o), whereas power normalized to fiber volume was unchanged. At the level of the cross bridge, the effects of short-term resistance training were quantitative (fiber hypertrophy and proportional increases in fiber P(o) and absolute power) rather than qualitative (no change in P(o)/CSA, V(o), or power/fiber volume). PMID- 12121855 TI - Central respiratory activity of the tadpole in vitro brain stem is modulated diversely by nitric oxide. AB - Nitric oxide (NO) is a potent central neuromodulator of respiration, yet its scope and site of action are unclear. We used 7-nitroindazole (7-NI), a selective inhibitor of endogenous neuronal NO synthesis, to investigate the neurogenesis of respiration in larval bullfrog (Rana catesbeiana) isolated brain stems. 7-NI treatment (0.0625-0.75 mM) increased the specific frequency of buccal ventilation (BV) events, indicating influence on BV central rhythm generators (CRGs). The drug reduced occurrence, altered burst shape, and disrupted clustering of lung ventilation (LV) events, without altering their specific frequency. LV burst occurrence and clustering also differed between pH conditions. We conclude that NO has diverse effects on respiratory rhythmogenesis, being necessary for the expression of respiratory rhythms, inhibiting the frequency of BV CRG, and affecting both shape and clustering of LV bursts through conditional modulation of LV CRG. We confirm central chemosensitivity in these preparations and demonstrate chemomodulation of LV burst clustering and occurrence but not specific frequency. Results support distinct oscillators underlying LV and BV CRGs. PMID- 12121856 TI - Pregnancy and acute baroreflex resetting in conscious rabbits. AB - To test the hypothesis that acute resetting of baroreflex control of heart rate (HR) is enhanced during pregnancy, we determined whether the rightward shift in the baroreflex relationship between arterial pressure and HR after arterial pressure is raised [~25 mmHg for 30 min, due to infusion of phenylephrine (PE) or methoxamine (Meth)] is greater in late pregnant compared with nonpregnant conscious rabbits. Baroreflex function was assessed by monitoring HR responses to both stepwise steady-state changes (n = 14) and rapid ramp changes (n = 10) in arterial pressure. Pregnancy decreased reflex gain, increased reflex minimum HR, and shifted the curves to a lower pressure level, when either the steady-state or ramp method was used (all changes, P < 0.05). When PE was used to increase pressure, resetting of steady-state curves was observed both before and during pregnancy, but the magnitude of the resetting was less in the pregnant rabbits. Further inspection of the data revealed that the size of the shift in pregnant rabbits was inversely related to the dose of PE. Because the pressure rise was the same in all experiments, PE appears to nonspecifically counteract acute resetting. When Meth was used instead to increase pressure, resetting of steady state curves was similar in pregnant and nonpregnant rabbits and was unrelated to dose. Similarly, when reflex curves were generated using the ramp method, and either Meth or low doses of PE were used to increase pressure, no differences in the degree of resetting were observed between pregnant and nonpregnant rabbits. In summary, high doses of PE counteract acute resetting of baroreflex control of HR. More importantly, while baroreflex function is depressed, the ability of the baroreflex to reset appears to be preserved during pregnancy. PMID- 12121857 TI - Sympathetic-renal interaction in chronic arterial pressure control. AB - The effects of neonatal sympathectomy of donors or recipients on posttransplantation arterial pressure were investigated in spontaneously hypertensive rats (SHR) by renal transplantation experiments. Conscious mean arterial pressure (MAP) and renal vascular resistance were 136 +/- 1 mmHg and 15.5 +/- 1.2 mmHg x ml(-1) x min x g in sympathectomized SHR (n = 8) vs. 158 +/- 4 mmHg (P < 0.001) and 20.8 +/- 1.1 mmHg x ml(-1) x min x g (P < 0.05) in controls (n = 10). Seven weeks after transplantation of a kidney from neonatally sympathectomized SHR donors, MAP in SHR recipients (n = 10) was 20 mmHg lower than in controls transplanted with a kidney from hydralazine-treated SHR (n = 10) (P < 0.05) associated with reduced sodium sensitivity of MAP. Neonatal sympathectomy also lowered MAP in F1-hybrids (F1H; SHR x Wistar-Kyoto rats). Within 6 wk after transplantation, renal grafts from untreated SHR increased MAP by 20 mmHg in sympathectomized F1H (n = 10) and by 35 mmHg in sham-treated F1H (n = 8) (P < 0.05). Neonatal sympathectomy induces chronic changes in SHR kidney function leading to a MAP reduction even when extrarenal sympathetic tone is restored. Generalized reduction in sympathetic tone resets the kidney-fluid system to reduced MAP and blunts the extent of arterial pressure rise induced by an SHR kidney graft. PMID- 12121858 TI - Acute sympathoexcitatory action of angiotensin II in conscious baroreceptor denervated rats. AB - Angiotensin II (ANG II) has complex actions on the cardiovascular system. ANG II may act to increase sympathetic vasomotor outflow, but acutely the sympathoexcitatory actions of exogenous ANG II may be opposed by ANG II-induced increases in arterial pressure (AP), evoking baroreceptor-mediated decreases in sympathetic nerve activity (SNA). To examine this hypothesis, the effect of ANG II infusion on lumbar SNA was measured in unanesthetized chronic sinoaortic denervated rats. Chronic sinoaortic-denervated rats had no reflex heart rate (HR) responses to pharmacologically evoked increases or decreases in AP. Similarly, in these denervated rats, nitroprusside-induced hypotension had no effect on lumbar SNA; however, phenylephrine-induced increases in AP were still associated with transient decreases in SNA. In control rats, infusion of ANG II (100 ng x kg(-1) x min(-1) iv) increased AP and decreased HR and SNA. In contrast, ANG II infusion increased lumbar SNA and HR in sinoaortic-denervated rats. In rats that underwent sinoaortic denervation surgery but still had residual baroreceptor reflex-evoked changes in HR, the effect of ANG II on HR and SNA was variable and correlated to the extent of baroreceptor reflex impairment. The present data suggest that pressor concentrations of ANG II in rats act rapidly to increase lumbar SNA and HR, although baroreceptor reflexes normally mask these effects of ANG II. Furthermore, these studies highlight the importance of fully characterizing sinoaortic-denervated rats used in experiments examining the role of baroreceptor reflexes. PMID- 12121859 TI - Effects of fetal ovine adrenalectomy on sympathetic and baroreflex responses at birth. AB - Studies were performed to test the hypothesis that the absence of adrenal glucocorticoids late in gestation alters sympathetic and baroreflex responses before and immediately after birth. Fetal sheep at 130-131 days gestation (term 145 days) were subjected to bilateral adrenalectomy before the normal prepartum increase in plasma cortisol levels. One group of fetuses (n = 5) received physiological cortisol replacement with a continuous infusion of hydrocortisone (2 mg x day(-1) x kg(-1) for 10 days), whereas the other group received 0.9% NaCl vehicle (n = 5). All animals underwent a second surgery 48 h before the study for placement of a renal nerve recording electrode. Heart rate (HR), mean arterial blood pressure (MABP), renal sympathetic nerve activity (RSNA), and baroreflex control of HR and RSNA were studied before and after cesarean section delivery. At the time of study (140-141 days gestation), fetal plasma cortisol concentration was undetectable in adrenalectomized (ADX) fetuses and 58 +/- 9 ng/ml in animals receiving cortisol replacement (ADX + F). Fetal and newborn MABP was significantly greater in ADX + F relative to ADX animals. One hour after delivery, MABP increased 13 +/- 3 mmHg and RSNA increased 91 +/- 12% above fetal values in ADX + F (both P < 0.05) but remained unchanged in ADX lambs. The midpoint pressures of the fetal HR and RSNA baroreflex function curves were significantly greater in ADX + F (54 +/- 3 and 56 +/- 3 mmHg for HR and RSNA curves, respectively) than ADX fetuses (45 +/- 2 and 46 +/- 3 mmHg). After delivery, the baroreflex curves reset toward higher pressure in ADX + F but not ADX lambs. These results suggest that adrenal glucocorticoids contribute to cardiovascular regulation in the late-gestation fetus and newborn by modulating arterial baroreflex function and sympathetic activity. PMID- 12121860 TI - Characterization of pituitary IGF-I receptors: modulation of prolactin and growth hormone. AB - There have been no studies in any vertebrate that have localized insulin-like growth factor (IGF)-I receptors in prolactin (PRL) cells or that have correlated pituitary binding to the potency of IGF-I in regulating both PRL and growth hormone (GH) secretion. We show that IGF-I binds with high affinity and specificity to the pituitary gland of hybrid striped bass (Morone saxatilis x M. chrysops). IGF-I and IGF-II were equipotent in inhibiting saturable (125)I-IGF-I binding, whereas insulin was ineffective. IGF-I binds with similar affinity to the rostral pars distalis (>95% PRL cells) as the whole pituitary gland and immunohistochemistry colocalizes IGF-I receptors and PRL in this same region. Des(1-3)IGF-I, a truncated analog of IGF-I that binds with high affinity to IGF-I receptors but weakly to IGF-I binding proteins (IGFBPs), showed a similar inhibition of saturable (125)I-IGF-I binding, but it was more potent than IGF-I in stimulating PRL and inhibiting GH release. These results are the first to localize IGF-I receptors to PRL cells, correlate IGF-I binding to its efficacy in regulating GH and PRL secretion, as well as demonstrate that IGFBPs may play a significant role in modulating the disparate actions of IGF-I on PRL and GH secretion. PMID- 12121861 TI - ICAM-1 and VCAM-1 mediate endotoxemic myocardial dysfunction independent of neutrophil accumulation. AB - Both intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) have been implicated in neutrophil-mediated lung and liver injury during sepsis. However, the role of these adhesion molecules as well as the contribution of neutrophils in myocardial dysfunction during sepsis remains to be determined. The purpose of this study was to examine the role of ICAM-1, VCAM-1, and neutrophils in lipopolysaccharide (LPS)-induced myocardial dysfunction. Mice were subjected to LPS (0.5 mg/kg ip) or vehicle (normal saline), and left ventricular developed pressure (LVDP) was determined by the Langendorff technique. LVDP was depressed by nearly 40% at 6 h after LPS. Immunofluorescent staining revealed a temporal increase in myocardial ICAM-1/VCAM 1 expression and neutrophils after LPS. Antibody blockade of VCAM-1 reduced myocardial neutrophil accumulation and abrogated LPS-induced cardiac dysfunction. Antibody blockade or absence of ICAM-1 (gene knockout) also abrogated LPS-induced cardiac dysfunction but did not reduce neutrophil accumulation. Neutrophil depletion (vinblastine or antibody) did not protect from LPS-induced myocardial dysfunction. Our results suggest that although endotoxemic myocardial dysfunction requires both ICAM-1 and VCAM-1, it occurs independent of neutrophil accumulation. PMID- 12121862 TI - Microvessel formation from mouse embryonic aortic explants is oxygen and VEGF dependent. AB - To delineate the roles of O(2) and vascular endothelial growth factor (VEGF) in the process of angiogenesis from the embryonic aorta, we cultured mouse embryonic aorta explants (thoracic level to lateral vessels supplying the mesonephros and metanephros) in a three-dimensional type I collagen gel matrix. During 8 days of culture under 5% O(2), but not room air, the addition of VEGF to explants stimulated the formation of CD31-positive, Flk-1-positive, Gs-IB(4)-positive structures in a concentration-dependent manner. Electron microscopy showed the structures to be capillary-like. VEGF-induced capillary-like structure formation was inhibited by sequestration of VEGF via addition of soluble Flt-1 fusion protein or anti-VEGF antibodies. Expression of Flk-1, but not Flt-1, was increased in embryonic aorta cultured under 5% O(2) relative to room air. Our data suggest that low O(2) upregulates Flk-1 expression in embryonic aorta in vitro and renders it more responsive to VEGF. PMID- 12121863 TI - 17Beta-estradiol decreases hypoxic induction of erythropoietin gene expression. AB - Exposure to chronic hypoxia induces erythropoietin (EPO) production to facilitate oxygen delivery to hypoxic tissues. Previous studies from our laboratory found that ovariectomy (OVX) exacerbates the polycythemic response to hypoxia and treatment with 17beta-estradiol (E2-beta) inhibits this effect. We hypothesized that E2-beta decreases EPO gene expression during hypoxia. Because E2-beta can induce nitric oxide (NO) production and NO can attenuate EPO synthesis, we further hypothesized that E2-beta inhibition of EPO gene expression is mediated by NO. These hypotheses were tested in OVX catheterized rats treated with E2-beta (20 microg/day) or vehicle for 14 days and exposed to 8 or 12 h of hypoxia (12% O(2)) or normoxia. We found that E2-beta treatment significantly decreased EPO synthesis and gene expression during hypoxia. E2-beta treatment did not induce endothelial NO synthase (eNOS) expression in the kidney but potentiated hypoxia induced increases in plasma nitrates. We conclude that E2-beta decreases hypoxic induction of EPO. However, this effect does not appear to be related to changes in renal eNOS expression. PMID- 12121864 TI - Lack of TNF-alpha attenuates intimal hyperplasia after mouse carotid artery injury. AB - This study sought to determine the influence of tumor necrosis factor-alpha (TNF alpha) on intimal hyperplasia (IH) and characterize the mechanisms of transcriptional regulation after vascular injury. A murine model of wire carotid artery injury was employed to induce IH in wild-type (WT) and TNF-alpha-deficient [TNF(-/-)] animals. Three days after injury, TNF-alpha and nuclear factor-kappaB (NF-kappaB) protein expression was markedly increased in the injured WT carotid artery compared to control. Injury increased TNF-alpha and NF-kappaB mRNA expression 100- and 7.5-fold, respectively. Compared with WT specimens, injury in TNF(-/-) animals decreased both NF-kappaB mRNA and protein nearly 7.5- and 4 fold, respectively. Expression of the NF-kappaB-dependent cytokine monocyte chemotactic protein 1 was markedly diminished in injured TNF(-/-) animals. Finally, TNF(-/-) animals demonstrated a sevenfold reduction in IH compared with WT animals. Cumulatively, these data mechanistically link TNF-alpha and NF-kappaB in vivo and suggest an important influence of TNF-alpha on postinjury IH. PMID- 12121865 TI - Sympathetic nerve regulation to heating is altered in senescent rats. AB - Renal and splanchnic sympathetic nerve discharge (SND) responses to increased (38 41 degrees C) internal temperature were determined in anesthetized young (3-6 mo old), mature (12 mo old), and senescent (24 mo old) Fischer 344 (F344) rats. We hypothesized that SND responses would be altered in senescent and mature rats as demonstrated by attenuated sympathoexcitatory responses to heating and by the absence of hyperthermia-induced SND pattern changes. The following observations were made. 1) Renal and splanchnic SND responses were significantly increased during heating in young and mature but not in senescent rats. 2) At 41 degrees C, renal and splanchnic SND responses were higher in young compared with senescent rats, and renal SND was higher in mature than in senescent rats. 3) Heating changed the SND bursting pattern in young, but not in mature or senescent, rats. 4) SND responses to heating did not differ between baroreceptor-innervated (BRI) and sinoaortic-denervated (SAD) senescent rats but were higher in SAD compared with BRI young rats. These results demonstrate an attenuated responsiveness of sympathetic neural circuits to heating in senescent F344 rats. PMID- 12121866 TI - Selective REM sleep deprivation during daytime I. Time course of interventions and recovery sleep. AB - Although repeated selective rapid eye movement (REM) sleep deprivation by awakenings during nighttime has shown that the number of sleep interruptions required to prevent REM sleep increases within and across consecutive nights, the underlying regulatory processes remained unspecified. To assess the role of circadian and homeostatic factors in REM sleep regulation, REM sleep was selectively deprived in healthy young adult males during a daytime sleep episode (7-15 h) after a night without sleep. Circadian REM sleep propensity is known to be high in the early morning. The number of interventions required to prevent REM sleep increased from the first to the third 2-h interval by a factor of two and then leveled off. Only a minor REM sleep rebound (11.6%) occurred in the following undisturbed recovery night. It is concluded that the limited rise of interventions during selective daytime REM sleep deprivation may be due to the declining circadian REM sleep propensity, which may partly offset the homeostatic drive and the sleep-dependent disinhibition of REM sleep. PMID- 12121867 TI - Selective REM sleep deprivation during daytime. II. Muscle atonia in non-REM sleep. AB - One of the hallmarks of rapid eye movement (REM) sleep is muscle atonia. Here we report extended epochs of muscle atonia in non-REM sleep (MAN). Their extent and time course was studied in a protocol that included a baseline night, a daytime sleep episode with or without selective REM sleep deprivation, and a recovery night. The distribution of the latency to the first occurrence of MAN was bimodal with a first mode shortly after sleep onset and a second mode 40 min later. Within a non-REM sleep episode, MAN showed a U-shaped distribution with the highest values before and after REM sleep. Whereas MAN was at a constant level over consecutive 2-h intervals of nighttime sleep, MAN showed high initial values when sleep began in the morning. Selective daytime REM sleep deprivation caused an initial enhancement of MAN during recovery sleep. It is concluded that episodes of MAN may represent an REM sleep equivalent and that it may be a marker of homeostatic and circadian REM sleep regulating processes. MAN episodes may contribute to the compensation of an REM sleep deficit. PMID- 12121868 TI - Dynamic baroreflex control of blood pressure: influence of the heart vs. peripheral resistance. AB - The aim in the present experiments was to assess the dynamic baroreflex control of blood pressure, to develop an accurate mathematical model that represented this relationship, and to assess the role of dynamic changes in heart rate and stroke volume in giving rise to components of this response. Patterned electrical stimulation [pseudo-random binary sequence (PRBS)] was applied to the aortic depressor nerve (ADN) to produce changes in blood pressure under open-loop conditions in anesthetized rabbits. The stimulus provided constant power over the frequency range 0-0.5 Hz and revealed that the composite systems represented by the central nervous system, sympathetic activity, and vascular resistance responded as a second-order low-pass filter (corner frequency approximately 0.047 Hz) with a time delay (1.01 s). The gain between ADN and mean arterial pressure was reasonably constant before the corner frequency and then decreased with increasing frequency of stimulus. Although the heart rate was altered in response to the PRBS stimuli, we found that removal of the heart's ability to contribute to blood pressure variability by vagotomy and beta(1)-receptor blockade did not significantly alter the frequency response. We conclude that the contribution of the heart to the dynamic regulation of blood pressure is negligible in the rabbit. The consequences of this finding are examined with respect to low frequency oscillations in blood pressure. PMID- 12121869 TI - Opposite effects of iv amiodarone on cardiovascular vagal and sympathetic efferent activities in rats. AB - It is unknown whether amiodarone exerts a direct central action on the cardiovascular autonomic nervous system. This study was designed to evaluate the effects of acute amiodarone administration on vagal and sympathetic efferent nerve discharges. Experiments were carried out in 25 decerebrate unanesthetized rats. In one group, vagal activity was recorded from preganglionic fibers isolated from the cervical vagus nerve. In another group, sympathetic recordings were obtained from fibers isolated from the cervical sympathetic trunk in intact conditions or after barodenervation. Recordings were performed before and for 60 min after amiodarone (50 mg/kg iv) administration. In all groups, amiodarone induced bradycardia and hypotension. Vagal activity increased immediately, reaching a significant difference after 20 min (260 +/- 131% from 16.4 +/- 3.3 spikes/s) and was unmodified by the barodenervation. At difference, sympathetic activity after an initial and short-lasting increase (150 +/- 83% from 24.8 +/- 5.7 spikes/s) began to decrease significantly after 20 min (36 +/- 17%) throughout the experiment. The initial increase in sympathetic activity was not observed in barodenervated animals. These changes in vagal and sympathetic activity could play an important role in contributing to the antiarrhythmic action of amiodarone. PMID- 12121870 TI - The epithelial cell cytoskeleton and intracellular trafficking. II. Intestinal epithelial cell exosomes: perspectives on their structure and function. AB - Intestinal epithelial cells (IEC) are located at the strategic interface between the external environment and the most extensive lymphoid compartment in the body. Besides their central role in the absorption of nutrients, they also provide sample information to the immune system on soluble or particulate antigens present in the intestinal lumen. Like professional antigen-presenting cells, IEC have recently been shown to secrete 30- to 90-nm diameter vesicles named exosomes from their apical and basolateral surfaces. These vesicles carry molecules that are implicated in adhesion and antigen presentation, such as major histocompatibility complex (MHC) class I molecules, MHC class II molecules, CD63, CD26/dipeptidyl-peptidase IV, tetraspan proteins, and A33 antigen. IEC exosomes therefore, constitute a link by which IEC may influence antigen presentation in the mucosal or systemic immune system independent of direct cellular contact with effector cells. PMID- 12121871 TI - Lipopolysaccharides in liver injury: molecular mechanisms of Kupffer cell activation. AB - Endogenous gut-derived bacterial lipopolysaccharides have been implicated as important cofactors in the pathogenesis of liver injury. However, the molecular mechanisms by which lipopolysaccharides exert their effect are not entirely clear. Recent studies have pointed to proinflammatory cytokines such as tumor necrosis factor-alpha as mediators of hepatocyte injury. Within the liver, Kupffer cells are major sources of proinflammatory cytokines that are produced in response to lipopolysaccharides. This review will focus on three important molecular components of the pathway by which lipopolysaccharides activate Kupffer cells: CD14, Toll-like receptor 4, and lipopolysaccharide binding protein. Within the liver, lipopolysaccharides bind to lipopolysaccharide binding protein, which then facilitates its transfer to membrane CD14 on the surface of Kupffer cells. Signaling of lipopolysaccharide through CD14 is mediated by the downstream receptor Toll-like receptor 4 and results in activation of Kupffer cells. The role played by these molecules in liver injury will be examined. PMID- 12121872 TI - Controversy: PPARgamma as a target for treatment of colorectal cancer. AB - Colorectal cancer (CRC) represents a significant cause of morbidity and mortality worldwide. Recently, ligands for the nuclear hormone receptor peroxisome proliferator-activated receptor gamma (PPARgamma) have exhibited promise in the treatment of CRC. For example, activation of PPARgamma reduces the proliferation of cultured CRC cells grown in vitro or in vivo using the nude mouse xenograft model of tumor growth. Furthermore, agonists of the receptor also reduce the development of preneoplastic lesions in a model of carcinogen-induced CRC in rats. However, ligands for the receptor paradoxically enhance intestinal adenoma formation in another murine model of intestinal polyposis, the APC(Min) mice. These disparate results may be due to the inherent limitations of the APC(Min) mouse as a model for humans with CRC. Finally, genetic studies identifying loss of function mutations of PPARgamma in human CRC specimens strongly suggest a tumor suppressive role for the receptor during the development of CRC. PMID- 12121873 TI - Different modes of NF-kappaB/Rel activation in pancreatic lobules. AB - The eukaryotic transcription factor nuclear factor-kappaB (NF-kappaB)/Rel is activated by a large variety of stimuli. It has been demonstrated that NF kappaB/Rel is induced during the course of cerulein pancreatitis. Here, we show that NF-kappaB/Rel is differentially activated in pancreatic lobules. Cerulein induces NF-kappaB/Rel via activation of IkappaB kinase (IKK), which causes degradation of IkappaBalpha but not IkappaBbeta. Tumor necrosis factor-alpha mediated IKK activation leads to IkappaBalpha and IkappaBbeta degradation. In contrast, oxidative stress induced by H(2)O(2) activates NF-kappaB/Rel independent of IKK activation and IkappaBalpha degradation; instead IkappaBalpha is phosphorylated on tyrosine. H(2)O(2) but not cerulein-mediated NF-kappaB/Rel activation can be blocked by stabilizing microtubules with Taxol. Inhibition of tubulin polymerization with nocodazole causes NF-kappaB/Rel activation in pancreatic lobules. These results propose three different pathways of NF kappaB/Rel activation in pancreatic acinar cells. Furthermore, these data demonstrate that microtubules play a key role in IKK-independent NF-kappaB/Rel activation following oxidative stress. PMID- 12121874 TI - PGF(2alpha)-induced contraction of cat esophageal and lower esophageal sphincter circular smooth muscle. AB - Lower esophageal sphincter (LES) tone depends on PGF(2alpha) and thromboxane A(2) acting on receptors linked to G(i3) and G(q) to activate phospholipases and produce second messengers resulting in muscle contraction. We therefore examined PGF(2alpha) signal transduction in circular smooth muscle cells isolated by enzymatic digestion from cat esophagus (Eso) and LES. In Eso, PGF(2alpha)-induced contraction was inhibited by antibodies against the alpha-subunit of G(13) and the monomeric G proteins RhoA and ADP-ribosylation factor (ARF)1 and by the C3 exoenzyme of Clostridium botulinum. A [(35)S]GTPgammaS-binding assay confirmed that G(13), RhoA, and ARF1 were activated by PGF(2alpha). Contraction of Eso was reduced by propranolol, a phospholipase D (PLD) pathway inhibitor and by chelerythrine, a PKC inhibitor. In LES, PGF(2alpha)-induced contraction was inhibited by antibodies against the alpha-subunit of G(q) and G(i3), and a [(35)S]GTPgammaS-binding assay confirmed that G(q) and G(i3) were activated by PGF(2alpha). PGF(2alpha)-induced contraction of LES was reduced by U-73122 and D609 and unaffected by propranolol. At low PGF(2alpha) concentration, contraction was blocked by chelerythrine, whereas at high concentration, contraction was blocked by chelerythrine and CGS9343B. Thus, in Eso, PGF(2alpha) activates a PLD- and protein kinase C (PKC)-dependent pathway through G(13), RhoA, and ARF1. In LES, PGF(2alpha) receptors are coupled to G(q) and G(i3), activating phosphatidylinositol- and phosphatidylcholine-specific phospholipase C. At low concentrations, PGF(2alpha) activates PKC. At high concentration, it activates both a PKC- and a calmodulin-dependent pathway. PMID- 12121875 TI - Stimulation of the gastrin-cholecystokinin(B) receptor promotes branching morphogenesis in gastric AGS cells. AB - Epithelial organization is maintained by cell proliferation, migration, and differentiation. In the case of the gastric epithelium, at least some of these events are regulated by the hormone gastrin. In addition, gastric epithelial cells are organized into characteristic tubular structures (the gastric glands), but the cellular mechanisms regulating the organization of tubular structures (sometimes called branching morphogenesis) are uncertain. In the present study, we examined the role of the gastrin-cholecystokinin(B) receptor in promoting branching morphogenesis of gastric epithelial cells. When gastric cancer AGS-G(R) cells were cultured on plastic, gastrin and PMA stimulated cell adhesion, formation of lamellipodia, and extension of long processes in part by activation of protein kinase C (PKC) and phosphatidylinositol (PI)-3 kinase. Branching morphogenesis was not observed in these circumstances. However, when cells were cultured on artificial basement membrane, the same stimuli increased the formation of organized multicellular arrays, exhibiting branching morphogenesis. These effects were reversed by inhibitors of PKC but not of PI-3 kinase. We conclude that, in the presence of basement membrane, activation of PKC by gastrin stimulates branching morphogenesis. PMID- 12121876 TI - Role of luminal ATP in regulating electrogenic Na(+) absorption in guinea pig distal colon. AB - Extracellular ATP regulates a variety of functions in epithelial tissues by activating the membrane P2-receptor. The purpose of this study was to investigate the autocrine/paracrine regulation by luminal ATP of electrogenic amiloride sensitive Na(+) absorption in the distal colon from guinea pigs treated with aldosterone by measuring the amiloride-sensitive short-circuit current (I(sc)) and (22)Na(+) flux in vitro with the Ussing chamber technique. ATP added to the luminal side inhibited the amiloride-sensitive I(sc) and (22)Na(+) absorption to a similar degree. The concentration dependence of the inhibitory effect of ATP on amiloride-sensitive I(sc) had an IC(50) value of 20-30 microM, with the maximum inhibition being approximately 50%. The effects of different nucleotides and of a nucleoside were also studied, the order of potency being ATP = UTP > ADP > adenosine. The effects of ATP were slightly, but significantly, reduced in the presence of suramin in the luminal solution. The inhibitory effect of luminal ATP was more potent in the absence of both Mg2+ and Ca2+ from the luminal solution. Pretreatment of the tissue with ionomycin or thapsigargin in the absence of serosal Ca2+ did not affect the percent inhibition of amiloride-sensitive I(sc) induced by ATP. Mechanical perturbation with a hypotonic luminal solution caused a reduction in amiloride-sensitive I(sc), this effect being prevented by the presence of hexokinase, an ATP-scavenging enzyme. These results suggest that ATP released into the luminal side by hypotonic stimulation could exert an inhibitory effect on the electrogenic Na(+) absorption. This effect was probably mediated by a P2Y(2) receptor on the apical membrane of colonic epithelial cells, and a change in the intracellular Ca2+ concentration may not be necessary for this process. PMID- 12121878 TI - Increased activity and expression of iNOS in human duodenal enterocytes from patients with celiac disease. AB - The activity of nitric oxide synthase (NOS) was assayed in enterocytes isolated from human duodenal biopsies to determine its role in celiac disease. Patients were categorized into groups with irritable bowel syndrome, iron-deficiency anemia, B(12)/folate deficiency, and treated and untreated celiac disease. Enterocytes isolated from all groups showed 1400W-inhibitable Ca2+-independent NOS activity with a pH level and temperature optimum of 9.4 and 37 degrees C, respectively. Western blotting showed that enterocytes expressed the inducible NOS protein and proteins with nitrated tyrosine residues, the latter being indicative of nitric oxide-driven peroxynitrite and/or free-radical damage. Endothelial NOS was seen only in the lamina propria. Patients with celiac disease had higher NOS activity than other patient groups. Treatment of the condition led to a fall in activity. Enzyme-linked immunosorbent assay demonstrated cGMP production by the enterocyte fraction, but cGMP levels did not correlate with NOS activity. These results suggest that inducible NOS is constitutively expressed in human duodenal enterocytes, is increased in patients with untreated celiac disease, and is partially corrected when such patients are treated. We found no evidence to support a role for nitric oxide in the formation of cGMP within the small intestine. Furthermore, we were unable to demonstrate a role for peroxynitrite/free radical damage in the pathophysiology of celiac disease. PMID- 12121877 TI - Helicobacter pylori induces apoptosis of rat gastric parietal cells. AB - Gastric Helicobacter pylori infection may lead to multifocal atrophic corpus gastritis associated with loss of epithelial cells as well as glandular structures. The current work investigated H. pylori effects on cell death of isolated, nontransformed rat parietal cells (PC). Highly enriched rat PC (>97%) were isolated from gastric mucosa and cultured in serum-free medium over 24 h. The cells were cocultured over 8 h with cytotoxin-associated immunodominant protein (cagA)(+)/vacuolating toxin (vacA)(+) or with cagA(-)/vacA(-) H. pylori laboratory strains and also with H. pylori mutants deleted in several genes of the cag pathogenicity island. Staphylococcus aureus or Campylobacter jejuni were used as controls. Apoptosis was determined by terminal deoxynucleotidyl transferase dUTP nick-end labeling staining and electron microscopy. Interleukin (IL)-8 and cytokine-induced neutrophil chemoattractant (CINC)-1 secretion was measured by ELISA. Activation of nuclear factor-kappaB (NF-kappaB) was studied in nuclear extracts of PC by electrophoretic mobility shift assay. Apoptosis of PC was induced in a concentration- and time-dependent manner by cagA(+)/vacA(+) H. pylori strains but not by cagA(-)/vacA(-) negative strains or by the cagE knockout mutant. S. aureus and C. jejuni had no effect. PC showed no IL-8 or CINC 1 secretion on exposure to cagA(+)/vacA(+) H. pylori. cagA(+)/vacA(+) strains induced activation of NF-kappaB complexes in nuclear extracts of PC, which were composed of p65 and p50 subunits. No significant stimulation of NF-kappaB activation was detected by incubation of PC with the cagE knockout mutant. Preincubation of PC with antisense but not missense oligodeoxynucleotides against the p65 subunit significantly reduced DNA binding to the kappaB recognition sequence. The p65 oligonucleotides as well as the proteasome inhibitor N-CBZ isoleucin-glutamin-(o-t-butyl-)-alanin-leucin and the nitric oxide synthase inhibitor N(G)-monomethyl-L-arginine completely prevented PC apoptosis induced by cagA(+)/vacA(+) strains. In summary, cagE presence appears to be essential for H. pylori-induced apoptosis of gastric parietal cells, and this effect is dependent on the activation of NF-kappaB and production of nitric oxide. PMID- 12121879 TI - Acid- and bile-induced PGE(2) release and hyperproliferation in Barrett's esophagus are COX-2 and PKC-epsilon dependent. AB - Barrett's esophagus (BE) results from acid and bile reflux and predisposes to cancer. To further understand the mechanisms of acid- and bile-induced hyperproliferation in BE, we investigated the release of PGE(2) in response to acid or bile salt exposure. Biopsies of esophagus, BE, and duodenum were exposed to a bile salt mixture as a 1-h pulse and compared with exposure to pH 7.4 for up to 24 h, and PGE(2) release, cyclooxygenase-2 (COX-2), and protein kinase C (PKC) expression were compared. Similar experiments were also performed with acidified media (pH 3.5) alone, in the presence or absence of bisindolylmaleimide (BIM), a selective PKC inhibitor, and NS-398, a COX-2 inhibitor. One-hour pulses of bile salts or acid significantly enhanced proliferation, COX-2 expression, and PGE(2) release in BE. In contrast, the combination pulse of acid and bile salts had no such effect. Treatment with either BIM or NS-398 led to a dramatic decrease in PGE(2) release in BE explants and a suppression of proliferation. The acid- or bile salt-mediated hyperproliferation is related to PGE(2) release. Acid- and bile salt-induced induction of COX-2 and PKC may explain, at least in part, the tumor-promoting effects of acid and bile in BE. PMID- 12121880 TI - Cortical processing of human gut sensation: an evoked potential study. AB - The rectum has a unique physiological role as a sensory organ and differs in its afferent innervation from other gut organs that do not normally mediate conscious sensation. We compared the central processing of human esophageal, duodenal, and rectal sensation using cortical evoked potentials (CEP) in 10 healthy volunteers (age range 21-34 yr). Esophageal and duodenal CEP had similar morphology in all subjects, whereas rectal CEP had two different but reproducible morphologies. The rectal CEP latency to the first component P1 (69 ms) was shorter than both duodenal (123 ms; P = 0.008) and esophageal CEP latencies (106 ms; P = 0.004). The duodenal CEP amplitude of the P1-N1 component (5.0 microV) was smaller than that of the corresponding esophageal component (5.7 microV; P = 0.04) but similar to that of the corresponding rectal component (6.5 microV; P = 0.25). This suggests that rectal sensation is either mediated by faster-conducting afferent pathways or that there is a difference in the orientation or volume of cortical neurons representing the different gut organs. In conclusion, the physiological and anatomic differences between gut organs are reflected in differences in the characteristics of their afferent pathways and cortical processing. PMID- 12121881 TI - Modulation of lipid synthesis, apolipoprotein biogenesis, and lipoprotein assembly by butyrate. AB - Short-chain fatty acids (SCFAs) are potent modulators of the growth, function, and differentiation of intestinal epithelia. In addition, high-fiber diets may protect against the development of atherosclerosis because of their cholesterol lowering effects due, in large part, to SCFA production, liver sterol metabolism, and bile acid excretion. Although the small gut plays a major role in dietary fat transport and contributes substantially to plasma cholesterol and lipoprotein homeostasis, the impact of SCFAs on intestinal lipid handling remains unknown. In the present study, the modulation of lipid synthesis, apolipoprotein biogenesis, and lipoprotein secretion by butyrate was investigated in Caco-2 cells plated on permeable polycarbonate filters, which permit separate access to the upper and lower compartments of the monolayers. Highly differentiated and polarized cells (20 days of culture) were incubated for 20 h with 20 mM butyrate in the apical medium. In the presence of [14C]oleic acid, butyrate led to a significant reduction of secreted, labeled triglycerides (27%; P < 0.01) and phospholipids (25%; P < 0.05). Similarly, butyrate significantly decreased the incorporation of [14C]acetate into exported cholesteryl ester (49%; P < 0.005). As expected from these results, with [14C]oleic acid as a precursor, butyrate significantly (P < 0.05) diminished the delivery of radiolabeled chylomicrons and very low-density lipoproteins. In parallel, [35S]methionine pulse labeling of Caco-2 cells revealed the concomitant inhibitory effect of butyrate on the synthesis of apolipoproteins B-48 (28%; P < 0.05) and A-I (32%; P < 0.01). Collectively, our data indicate that butyrate may influence lipid metabolism in Caco-2 cells, thus suggesting a potential regulation of intestinal fat absorption and circulating lipoprotein concentrations. PMID- 12121882 TI - Substance P-evoked Cl(-) secretion in guinea pig distal colonic epithelia: interaction with PGE(2). AB - Interaction between substance P (SP) and PGE(2) on Cl(-) secretion in the guinea pig distal colonic epithelia was investigated. A short-circuit current (I(sc)) was measured as an index of ion transport. Mucosa preparations deprived of muscle and submucosa of distal colon were mounted in the Ussing flux chamber and treated with TTX and piroxicam to remove the influences of neuronal activity and endogenous PG synthesis, respectively. Although SP (10(-7) M) itself evoked little increase in I(sc), exogenous PGE(2) concentration dependently enhanced the response of SP. The effect of PGE(2) on the SP-evoked response was mimicked by forskolin and 8-bromoadenosine cAMP. Depletion of Ca2+ from the bathing solution reduced the PGE(2)-dependent response of SP. Effects of PGE(2), SP, and SP in the presence of PGE(2) on intracellular Ca2+ concentration ([Ca2+](i)) in isolated crypt cells were measured by the confocal microscope fluorescence imaging system. SP, but not PGE(2), temporally evoked an increase in [Ca2+](i) but declined to the baseline within 3 min. A return of the SP-evoked increase in [Ca2+](i) was slower in the presence of PGE(2) than SP alone. These results suggest that PGE(2) synergistically enhances SP-evoked Cl(-) secretion via an interaction between the intracellular cAMP and [Ca2+](i) in the epithelial cells. In conclusion, SP and PGE(2) could cooperatively induce massive Cl(-) secretion in guinea pig distal colon at epithelial levels. PMID- 12121883 TI - Trans-10,cis-12-conjugated linoleic acid inhibits Caco-2 colon cancer cell growth. AB - A commercially available mixture of conjugated linoleic acid (CLA) isomers decreases colon cancer cell growth. We compared the individual potencies of the two main isomers in this mixture [cis-9,trans-11 (c9t11) and trans-10,cis-12 (t10c12)] and assessed whether decreased cell growth is related to changes in secretion of insulin-like growth factor II (IGF-II) and/or IGF-binding proteins (IGFBPs), which regulate Caco-2 cell proliferation. Cells were incubated in serum free medium with different concentrations of the individual CLA isomers. t10c12 CLA dose dependently decreased viable cell number (55 +/- 3% reduction 96 h after adding 5 microM t10c12 CLA). t10c12 CLA induced apoptosis and decreased DNA synthesis, whereas c9t11 CLA had no effect. Immunoblot analysis of 24-h serum free conditioned medium using a monoclonal anti-IGF-II antibody revealed that Caco-2 cells secreted both a mature 7,500 molecular weight (M(r)) IGF-II and higher M(r) forms of IGF-II. The levels of the higher M(r) and the mature form of IGF-II were decreased 50 +/- 3% and 22 +/- 2%, respectively, by 5 microM t10c12 CLA. c9t11 CLA had no effect. Ligand blot analysis of conditioned medium using 125I-labeled IGF-II revealed that t10c12 CLA slightly decreased IGFBP-2 production; c9t11 CLA had no effect. Exogenous IGF-II reversed t10c12 CLA-induced growth inhibition and apoptosis. These results indicate that CLA-inhibited Caco-2 cell growth is caused by t10c12 CLA and may be mediated by decreasing IGF-II secretion in Caco-2 cells. PMID- 12121885 TI - Multichannel intraluminal impedance accurately detects fasting, recumbent reflux events and their clearing. AB - Multichannel intraluminal impedance (MII) is a new diagnostic test for gastroesophageal reflux disease (GERD). The objective of this report is to determine the accuracy of MII in detecting individual reflux events (REs) identified by pH probe and manometry, as well as their clearing in patients with severe GERD compared with normal volunteers. Ten severe GERD patients and 10 normal volunteers underwent simultaneous manometry [7 sites: gastric, lower esophageal sphincter, esophagus (4), pharynx], pH, and MII (6 sites in esophagus) for 15 min in the left and right recumbent posture while fasting. We found that patients had 30-fold more REs than normal volunteers (41 +/- 11 vs. 1.3 +/- 0.4), and 95% of all REs were detected by MII. An average 15-fold fall in impedance with liquid and fivefold rise with gas made REs and their composition easy to detect with MII. In the right recumbent posture, nearly all REs detected by MII were liquid (98%, 98/100). In contrast, all 283 REs detected by MII in the left recumbent posture were gas. Nearly all REs detected by MII were cleared (98%, 368/374). Mean acid clearing time was threefold longer (47 s) than clearing time by either manometry (15 s) or MII (13 s), primarily due to acid rereflux, i.e., additional acid REs during acid clearing. We conclude that MII is accurate in detecting REs identified by manometry and/or pH probe, their composition, and their clearing. PMID- 12121884 TI - Mechanical properties in the human gastric antrum using B-mode ultrasonography and antral distension. AB - The aims of this study were to investigate gastric antral geometry and stress strain properties by using transabdominal ultrasound scanning during volume controlled distensions in the human gastric antrum. Seven healthy volunteers underwent stepwise inflation of a bag located in the antrum with volumes up to 60 ml. The stretch ratio and Cauchy stress and strain were calculated from measurements of pressure, diameter, and wall thickness. A second distension series was conducted in three volunteers during administration of the anticholinergic drug butylscopolamine. Analysis of stretch ratios demonstrated positive strain in the circumferential direction, negative strain in the radial direction, and no strain in the longitudinal direction. The stress-strain relation was exponential and did not differ without or with the administration of butylscopolamine. The wall stress was decomposed into its active and passive components. The well-known length-tension diagram from in vitro studies of smooth muscle strips was reproduced. The maximum active tension appeared at a volume of 50 ml, corresponding to a stretch ratio of 1.5. We conclude that the method provides measures of antral biomechanical wall properties and can be used to reproduce the muscle length-tension diagram in humans. PMID- 12121886 TI - Rectal and colonic distension elicit viscerovisceral reflexes in humans. AB - Colonic transit is slowed in patients with disordered rectal evacuation, but the mechanism of this phenomenon is unclear. Our objective was to investigate rectocolonic inhibitory reflexes in humans to provide potential insight into patients with obstructed defecation. In 30 healthy subjects, a barostat manometric assembly recorded colonic tone and phasic activity in the descending colon during rectal distension and recorded rectal tone during colonic distension. Phasic distensions were 8, 16, and 32 mmHg above balloon operating pressure, and staircase inflations were comprised of balloon inflation then deflation in 2-mmHg increments at 30-s intervals from 0 to 36 mmHg. Colonic balloon volumes increased to a similar extent during phasic rectal distensions 8, 16, and 32 mmHg above operating pressure, reflecting reduced colonic tone; balloon volumes also increased and phasic pressure activity decreased during staircase rectal distensions. In contrast, rectal balloon volume declined, reflecting increased tone during phasic and staircase colonic distensions. Thus rectal distension inhibited colonic motor activity, indicative of a viscerovisceral inhibitory reflex. PMID- 12121887 TI - Role of pp60(c-src) and p(44/42) MAPK in ANG II-induced contraction of rat tonic gastrointestinal smooth muscles. AB - We examined the role of mitogen-activated protein kinase (p(44/42) MAPK) in ANG II-induced contraction of lower esophageal sphincter (LES) and internal anal sphincter (IAS) smooth muscles. Studies were performed in the isolated smooth muscles and cells (SMC). ANG II-induced changes in the levels of phosphorylation of different signal transduction and effector proteins were determined before and after selective inhibitors. ANG II-induced contraction of the rat LES and IAS SMC was inhibited by genistein, PD-98059 [a specific inhibitor of MAPK kinases (MEK 1/2)], herbimycin A (a pp60(c-src) inhibitor), and antibodies to pp60(c-src) and p(120) ras GTPase-activating protein (p(120) rasGAP). ANG II-induced contraction of the tonic smooth muscles was accompanied by an increase in tyrosine phosphorylation of p(120) rasGAP. These were attenuated by genistein but not by PD-98059. ANG II-induced increase in phosphorylations of p(44/42) MAPKs and caldesmon was attenuated by both genistein and PD-98059. We conclude that pp60(c src) and p(44/42) MAPKs play an important role in ANG II-induced contraction of LES and IAS smooth muscles. PMID- 12121888 TI - Impaired gastrocolonic response and peristaltic reflex in slow-transit constipation: role of 5-HT(3) pathways. AB - Colonic motility is modulated by the 5-hydroxytryptamine (5-HT)(3)-dependent gastrocolonic response and 5-HT(3)-independent peristaltic reflex. We compared descending colon tone responses to antral distension, duodenal lipid perfusion, and colonic distension after double-blind placebo or granisetron in 13 healthy volunteers and nine slow-transit constipated patients. Antral distension (100-300 ml) and duodenal lipids (3 kcal/min) evoked increases in colon tone in volunteers, which were blunted in constipated patients (P < 0.05). Granisetron (10 microg/kg) reduced responses to antral distension and lipids in volunteers and to lipids in constipated patients (P < 0.05). The ascending contraction of the peristaltic reflex was blunted in constipated patients (P < 0.05), whereas descending responses were similar. Granisetron did not modify the peristaltic reflex. Colonic responses to bethanechol were similar in patients and volunteers. In conclusion, antral distension- and duodenal lipid-activated gastrocolonic responses and ascending contractions of the peristaltic reflex are impaired with slow-transit constipation with loss of both 5-HT(3)-dependent and -independent function. Thus abnormalities of neural reflex modulation of colonic motor function may play pathophysiological roles in slow-transit constipation. PMID- 12121889 TI - Intestinal preconditioning prevents systemic inflammatory response in hemorrhagic shock. Role of HO-1. AB - Intestinal ischemia-reperfusion has been implicated in the systemic inflammatory response and organ injury in hemorrhagic shock, but the exact role of the intestine has never been directly demonstrated. Preconditioning (PC) with brief periods of intermittent ischemia is a known potent anti-ischemic intervention and thus can be used as a tool to assess the role of local intestinal ischemia reperfusion injury in systemic inflammatory response. Thus rats were first subjected to sham surgery or intestinal preconditioning with four cycles of 1-min ischemia and 10 min of reperfusion 24 h before hemorrhagic shock followed by resuscitation. PC reduced fluid requirements, lung edema, and lactate and tumor necrosis factor-alpha production. These effects were abolished by the heme oxygenase-1 (HO-1) inhibitor tin protoporphyrin (Sn-PP). PC induced more than fivefold in intestinal HO-1 expression. These results suggest that intestinal ischemia-reperfusion is a major trigger for inflammatory response and organ injury in nonseptic shock. HO-1 appears to play an important role in the protective effect of intestinal preconditioning. PMID- 12121890 TI - Multiple transcription factors in 5'-flanking region of human polymeric Ig receptor control its basal expression. AB - The polymeric Ig receptor (pIgR) is a critical component of the mucosal immune system and is expressed in largest amounts in the small intestine. In this study, we describe the initial characterization of the core promoter region of this gene. Expression of chimeric promoter-reporter constructs was supported in Caco-2 and HT-29 cells, and DNase I footprint analysis revealed a large protein complex within the core promoter region. Site-directed mutagenesis experiments determined that elements within this region serve to either augment or repress basal activity of the human pIgR promoter. Band shift assays of overlapping oligonucleotides within the core promoter identified eight distinct complexes; the abundance of most complexes was enhanced in post-confluent cells. In summary, we report the characterization of the human pIgR promoter and the essential role that eight different nuclear complexes have in controlling basal expression of this gene in Caco-2 cells. PMID- 12121891 TI - Glucocorticoid regulation and glycosylation of mouse intestinal type IIb Na-P(i) cotransporter during ontogeny. AB - We sought to characterize expression of an apically expressed intestinal Na-P(i) cotransporter (Na-P(i)-IIb) during mouse ontogeny and to assess the effects of methylprednisolone (MP) treatment. In control mice, Na-P(i) uptake by intestinal brush-border membrane vesicles was highest at 14 days of age, lower at 21 days, and further reduced at 8 wk and 8-9 mo of age. Na-P(i)-IIb mRNA and immunoreactive protein levels in 14-day-old animals were markedly higher than in older groups. MP treatment significantly decreased Na-P(i) uptake and Na-P(i)-IIb mRNA and protein expression in 14-day-old mice. Additionally, the size of the protein was smaller in 14-day-old mice. Deglycosylation of protein from 14-day old and 8-wk-old animals with peptide N-glycosidase reduced the molecular weight to the predicted size. We conclude that intestinal Na-P(i) uptake and Na-P(i)-IIb expression are highest at 14 days and decrease with age. Furthermore, MP treatment reduced intestinal Na-P(i) uptake approximately threefold in 14-day-old mice and this reduction correlates with reduced Na-P(i)-IIb mRNA and protein expression. We also demonstrate that Na-P(i)-IIb is an N-linked glycoprotein and that glycosylation is age dependent. PMID- 12121893 TI - Interstitial cells of Cajal and electrical activity in ganglionic and aganglionic colons of mice. AB - An antibody directed against Kit protein was used to investigate the distribution of interstitial cells of Cajal (ICC) within the murine colon. The ICC density was greatest in the proximal colon and decreased along its length. The distribution of the different classes of ICC in the aganglionic colons of lethal spotted (ls/ls) mice was found to be similar in age-matched wild-type controls. There were marked differences in the electrical activities of the colons from ls/ls mutants compared with wild-type controls. In ls/ls aganglionic colons, the circular muscle was electrically quiescent compared with the spontaneous spiking electrical activity of wild-type tissues. In ls/ls aganglionic colons, postjunctional neural responses were greatly affected. Inhibitory junction potentials were absent or excitatory junction potentials inhibited by atropine were observed. In conclusion, the distribution of ICC in the ganglionic and aganglionic regions of the colons from ls/ls mutants appeared similar to that of wild-type controls. The electrical activity and neural responses of the circular layer are significantly different in aganglionic segments of ls/ls mutants. PMID- 12121892 TI - Rat liver myofibroblasts and hepatic stellate cells differ in CD95-mediated apoptosis and response to TNF-alpha. AB - Hepatic stellate cells (HSC), particularly activated HSC, are thought to be the principle matrix-producing cell of the diseased liver. However, other cell types of the fibroblast lineage, especially the rat liver myofibroblasts (rMF), also have fibrogenic potential. A major difference between the two cell types is the different life span under culture conditions. Although nearly no spontaneous apoptosis could be shown in rMF cultures, 18 +/- 2% of the activated HSC (day 7) were apoptotic. Compared with activated HSC, CD95R was expressed in 70% higher amounts in rMF. CD95L could only be detected in activated HSC. Stimulation of the CD95 system by agonistic antibodies (1 ng/ml) led to apoptosis of all rMF within 2 h, whereas activated HSC were more resistant (5.3 h/ 40% of total cells). Although transforming growth factor-beta downregulated apoptosis in both activated HSC and rMF, tumor necrosis factor-alpha (TNF-alpha) upregulated apoptosis in rMF. Lack of spontaneous apoptosis and CD95L expression in rMF and the different reaction on TNF-alpha stimulation reveal that activated HSC and rMF belong to different cell populations. PMID- 12121894 TI - Reduction of spontaneous and irradiation-induced apoptosis in small intestine of IGF-I transgenic mice. AB - Insulin-like growth factor I (IGF-I) may promote survival of putative stem cells in the small intestinal epithelium. Mitosis and apoptosis were quantified in crypts of nonirradiated and irradiated IGF-I transgenic (TG) and wild-type (WT) littermates. The mean apoptotic index was significantly greater in WT vs. TG littermates. After irradiation, apoptotic indexes increased, and WT mice showed a more dramatic increase in apoptosis than TG mice at the location of putative stem cells. After irradiation, no mitotic figures were observed in WT crypts, whereas mitosis was maintained within the jejunal epithelium of TG mice. The abundance and localization of Bax mRNA did not differ between nonirradiated littermates. However, there was more Bax mRNA in TG vs. WT mice after irradiation. Bax mRNA was located along the entire length of the irradiated crypt epithelium, but there was less Bax protein observed in the bottom third of TG mouse crypts compared with WT littermates. IGF-I regulates cell number by stimulating crypt cell proliferation and decreasing apoptosis preferentially within the stem cell compartment. PMID- 12121895 TI - Orexins in rat dorsal motor nucleus of the vagus potently stimulate gastric motor function. AB - Orexins regulate food intake, arousal, and the sleep-wake cycle. They are synthesized by neurons in the lateral hypothalamus and project to autonomic areas in the hindbrain. Orexin A applied to the dorsal surface of the medulla stimulates gastric acid secretion via a vagally mediated pathway. We tested the hypothesis that orexins in the dorsal motor nucleus (DMN) of the vagus regulate gastric motor function. Multibarelled micropipette assemblies were used to administer vehicle, L-glutamate, orexins A (1 and 10 pmol) and B (10 pmol), and a dye marker into this site in anesthetized rats. When the pipette was positioned in the DMN rostral to the obex (where excitation of neurons by L-glutamate evoked an increase in contractility), orexins A and B increased intragastric pressure and antral motility. In contrast, 10 pmol orexin A microinjected into the DMN caudal to the obex (where L-glutamate evokes gastric relaxation through a vagal inhibitory pathway) did not significantly alter gastric motor function. In separate immunocytochemical studies, orexin receptor 1 was highly expressed in neurons in the DMN. Specifically, it was present in retrogradely labeled preganglionic neurons in the DMN that innervate the stomach. These data are consistent with the idea that orexin A stimulates vagal excitatory motor neurons. These are the first data to suggest that orexins in the DMN have potent and long lasting effects to increase gastric contractility. PMID- 12121896 TI - Immunolocalization of electroneutral Na(+)-HCO cotransporters in human and rat salivary glands. AB - Patterns of salivary HCO secretion vary widely among species and among individual glands. In particular, virtually nothing is known about the molecular identity of the HCO transporters involved in human salivary secretion. We have therefore examined the distribution of several known members of the Na(+)-HCO cotransporter (NBC) family in the parotid and submandibular glands. By use of a combination of RT-PCR and immunoblotting analyses, the electroneutral cotransporters NBC3 and NBCn1 mRNA and protein expression were detected in both human and rat tissues. Immunohistochemistry demonstrated that NBC3 was present at the apical membranes of acinar and duct cells in both human and rat parotid and submandibular glands. NBCn1 was strongly expressed at the basolateral membrane of striated duct cells but not in the acinar cells in the human salivary glands, whereas little or no NBCn1 labeling was observed in the rat salivary glands. The presence of NBCn1 at the basolateral membrane of human striated duct cells suggests that it may contribute to ductal HCO secretion. In contrast, the expression of NBC3 at the apical membranes of acinar and duct cells in both human and rat salivary glands indicates a possible role of this isoform in HCO salvage under resting conditions. PMID- 12121897 TI - In situ correlation of cytokine secretion and apoptosis in Helicobacter pylori associated gastritis. AB - Tumor necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma) are important for the pathogenesis of Helicobacter pylori-associated gastritis and peptic ulcer disease. Gastric biopsies from H. pylori-positive and -negative patients were used to examine the in situ correlation of TNF-alpha and IFN-gamma with epithelial cell apoptosis, bacterial load, and histological parameters of gastritis. From the same patients, we isolated H. pylori-specific T cell lines and clones and examined their ex vivo release of proinflammatory cytokines. We found a highly significant correlation of TNF-alpha and IFN-gamma production with activity and grade of gastritis (P < 0.01), H. pylori density (P = 0.01), epithelial cell apoptosis (P < 0.001), and Fas/Fas-ligand expression (P < 0.001). T cell lines and clones were all TCR-alphabeta(+) and showed T helper 1 functional phenotype. With the use of serial histological sections, this study showed for the first time the in situ correlation of TNF-alpha and IFN-gamma with epithelial cell apoptosis, bacterial load, and histological severity of disease and emphasizes the role of these cytokines in the pathophysiology of H. pylori associated disease. PMID- 12121898 TI - Clindamycin as an antimalarial drug: review of clinical trials. PMID- 12121899 TI - Influence of clarithromycin on early atherosclerotic lesions after Chlamydia pneumoniae infection in a rabbit model. AB - Chlamydia pneumoniae may play a role in atherogenesis and vascular diseases, and antibiotics may prove useful in these conditions. Three groups of New Zealand White rabbits (24 per group) were infected via the nasopharynx with C. pneumoniae on three separate occasions (2 weeks apart). Group I was untreated and sacrificed at 12 weeks; group II received clarithromycin at 20 mg/kg/day for 8 days, beginning 5 days after each inoculation (early treatment); and group III received a similar dose of clarithromycin starting 2 weeks after the third inoculation and continued for 6 weeks thereafter (delayed treatment). To test for a possible anti inflammatory effect of clarithromycin, two other groups of uninfected rabbits (12 animals in each) were fed 0.5% cholesterol-enriched chow, and one of these groups was treated with clarithromycin at 30 mg/kg/day for 6 weeks. Of 23 untreated infected rabbits, 8 developed early lesions of atherosclerosis, whereas 2 of the 24 early-treated group II had similar changes (P = 0.036 [75% efficacy]). However, in the delayed-treatment group, group III, 3 of 24 rabbits developed early lesions of atherosclerosis, thus demonstrating 62.5% reduction compared to the untreated controls (P = 0.07 [trend to statistical significance]). C. pneumoniae antigen was detected in 8 of 23 group I (untreated) rabbits versus 1 of 24 of the early-treated (group II) rabbits and 4 of 24 animals in the delayed group III (P = 0.009 and 0.138, respectively). All of the untreated, cholesterol fed rabbits had moderate to advanced atherosclerosis (grade III or IV); clarithromycin had no effect on reducing the prevalence of but did reduce the extent of atherosclerosis in the cholesterol-fed rabbits by 17% compared to untreated controls. Thus, clarithromycin administration modified C. pneumoniae induced atherosclerotic lesions and reduced the ability to detect organism in tissue. Early treatment was more effective than delayed treatment. PMID- 12121900 TI - Comparative stability studies of antipseudomonal beta-lactams for potential administration through portable elastomeric pumps (home therapy for cystic fibrosis patients) and motor-operated syringes (intensive care units). AB - The stability of antipseudomonal beta-lactams in concentrated solutions was examined in view of their potential administration by continuous infusion with external pumps (for intensive care patients) or with portable pumps carried under clothing (for cystic fibrosis patients). Aztreonam (100 g/liter), piperacillin (128 g/liter, with tazobactam), and azlocillin (128 g/liter) remained 90% stable for up to more than 24 h at 37 degrees C (mezlocillin [128 g/liter] was stable at 25 degrees C but not at 37 degrees C). Ceftazidime (120 g/liter), cefpirome (32 g/liter), and cefepime (50 g/liter) remained 90% stable for up to 24, 23.7, and 20.5 h at 25 degrees C but only for 8, 7.25, and 13 h at 37 degrees C, respectively. The control of temperature therefore appears to be critical for all three cephalosporins that cannot be recommended for use in portable pumps carried under clothes for prolonged periods for reasons of stability. Cefpirome and cefepime solutions developed an important color change (from light yellow to dark red) upon exposure when stored at 30 degrees C or higher. Degradation of ceftazidime was accompanied by the liberation of pyridine which, at 37 degrees C, was in excess of what is allowed by the U.S. Pharmacopeia, i.e., 1.1 mg/liter, after 8 and 12 h for drug concentrations of 12 and 8.3%, respectively. Imipenem and meropenem are too unstable (10% degradation at 25 degrees C after 3.5 and 5.15 h, respectively) to be recommended for use by continuous infusion. Faropenem, examined in comparison with imipenem and meropenem, proved as stable as aztreonam or piperacillin. PMID- 12121901 TI - Two diarylurea electron transport inhibitors reduce Staphylococcus aureus hemolytic activity and protect cultured endothelial cells from lysis. AB - Reduction in electron transport is associated with decreased production in alpha toxin despite the fact that Staphylococcus aureus is able to grow from 1 CFU to >10(7) CFU. Similarly, under anaerobic conditions, S. aureus does not produce alpha-toxin. Although the pathways that connect oxidative metabolism and toxin production are unknown, agents are available that exhibit greater inhibition of plant versus mammalian electron transport. Herbicides block electron transport in plants by inhibiting the formation of phosphoquinol (QH(2)) in plants. Commercial use in farming is possible because these compounds are much less active against the quinones found mammalian mitochondria. Because bacterial electron transport systems are closer to plant than mammalian systems, we hypothesized that inhibitors of respiration might be able to reduce S. aureus electron transport and block the production of alpha-toxin. We studied two compounds and found that the effective dose for the inhibition of bacterial respiration was 50 to >3,500 times lower than the concentration required to cause similar inhibition of rat mitochondrial respiration. Compounds I and II also reduced toxin production in S. aureus without causing overt toxicity to cultured endothelial cells. Finally, the compounds reduced the damage caused by S. aureus when cocultured with the endothelial cells. This raises the possibility that compounds that inhibit bacterial respiration might be prove valuable for the prevention of toxin production in S. aureus. PMID- 12121902 TI - Tn5406, a new staphylococcal transposon conferring resistance to streptogramin a and related compounds including dalfopristin. AB - We characterized a new transposon, Tn5406 (5,467 bp), in a clinical isolate of Staphylococcus aureus (BM3327). It carries a variant of vgaA, which encodes a putative ABC protein conferring resistance to streptogramin A but not to mixtures of streptogramins A and B. It also carries three putative genes, the products of which exhibit significant similarities (61 to 73% amino acid identity) to the three transposases of the staphylococcal transposon Tn554. Like Tn554, Tn5406 failed to generate target repeats. In BM3327, the single copy of Tn5406 was inserted into the chromosomal att554 site, which is the preferential insertion site of Tn554. In three other independent S. aureus clinical isolates, Tn5406 was either present as a single plasmid copy (BM3318), as two chromosomal copies (BM3252), or both in the chromosome and on a plasmid (BM3385). The Tn5406 carrying plasmids also contain two other genes, vgaB and vatB. The insertion sites of Tn5406 in BM3252 were studied: one copy was in att554, and one copy was in the additional SCCmec element. Amplification experiments revealed circular forms of Tn5406, indicating that this transposon might be active. To our knowledge, a transposon conferring resistance to streptogramin A and related compounds has not been previously described. PMID- 12121903 TI - Hexapeptide derivatives of glycopeptide antibiotics: tools for mechanism of action studies. AB - Hexapeptide (des-N-methylleucyl) derivatives of LY264826 were prepared in order to examine further the role of N-substituted hydrophobic side chains in defining the mechanisms of action of semisynthetic glycopeptide antibiotics. The hexapeptide of LY264826 binds to the cell wall intermediate analog L-Lys-D-Ala-D Ala with a 100-fold lower affinity than LY264826 and inhibits Micrococcus luteus almost 200-fold more poorly than LY264826 does. Alkylation of the 4-epi vancosamine moiety of the disaccharide significantly enhanced the antibacterial activity of the hexapeptide. Alkylation did not affect the binding affinity for D alanyl-D-alanine residues; however, it did enhance dimerization 7,000-fold and enhanced binding to bacterial membrane vesicles noticeably compared with the levels of dimerization and binding for the unsubstituted hexapeptide. The findings from this study complement those presented in an earlier report (N. E. Allen, D. L. LeTourneau, and J. N. Hobbs, Jr., J. Antibiot. 50:677-684, 1997) and are consistent with the conclusion that the enhanced antibacterial activities of semisynthetic glycopeptide antibiotics derive from the ability of the hydrophobic side chain to markedly affect both dimerization and binding to bacterial membranes. PMID- 12121904 TI - Analysis of penicillin-binding protein genes of clinical isolates of Streptococcus pneumoniae with reduced susceptibility to amoxicillin. AB - The recent emergence of pneumococcal isolates exhibiting an unusual resistance phenotype of higher amoxicillin MICs in relation to the penicillin MICs prompted an analysis of the pbp genes from three such strains isolated in France. For comparison, three amoxicillin-susceptible strains were included in the study. DNA sequence analysis of the pbp2x, pbp2b, and pbp1a genes revealed extensive sequence divergence in all six isolates compared to the sequences of the genes of penicillin-susceptible strain R6. With the exception of pbp2b, no amino acid mutations were unique to the resistant isolates. Transformation experiments with cloned pbp genes isolated from one of the resistant isolates demonstrated a stepwise development of amoxicillin resistance involving penicillin-binding proteins (PBPs) 2X, 2B, and 1A. Full resistance, equivalent to that of the donor strain, was achieved only when genomic DNA was transformed into R6(2x/2b/1a) mutants, suggesting that full resistance development in this isolate is mediated by a non-PBP determinant. Moreover, the recently identified murMN resistance determinant does not appear to have any impact on resistance in this isolate. This determinant (from the French isolate) was, however, able to transform an R6 mutant harboring pbp2x, pbp2b, and pbp1a genes from a Hungarian clone with an extremely high level of penicillin resistance so that it had increased levels of penicillin resistance. These results indicate that the development of high-level beta-lactam resistance is a complex process and that the involvement of MurMN in penicillin resistance appears to be dependent on specific mutations in PBPs 2X, 2B, and/or 1A. Furthermore, an additional (as yet unidentified) non-PBP-mediated resistance determinant is required for full resistance development in some pneumococci. PMID- 12121905 TI - Effects of gender, AIDS, and acetylator status on intrapulmonary concentrations of isoniazid. AB - The objective of the present study was to evaluate the effects of gender, AIDS, and acetylator status on the steady-state concentrations of orally administered isoniazid in plasma and lungs. Isoniazid was administered at 300 mg once daily for 5 days to 80 adult volunteers. Subjects were assigned to eight blocks according to gender, presence or absence of AIDS, and acetylator status. Blood was obtained prior to administration of the first dose, 1 h after administration of the last dose, and at the completion of bronchoscopy and bronchoalveolar lavage (BAL), which was performed 4 h after administration of the last dose. The metabolism of caffeine was used to determine acetylator status. Standardized bronchoscopy was performed without systemic sedation. The volume of epithelial lining fluid (ELF) recovered was calculated by the urea dilution method. Isoniazid concentrations in plasma, BAL fluid, and alveolar cells (ACs) were measured by high-performance liquid chromatography. AIDS status or gender had no significant effect on the concentrations of isoniazid in plasma at 1 or 4 h. Concentrations in plasma at 4 h and concentrations in ELF were greater in slow acetylators than fast acetylators. The concentration in plasma (1.85 +/- 1.60 microg/ml [mean +/- standard deviation; n = 80]) at 1 h following administration of the last dose was not significantly different from that in ELF (2.25 +/- 3.50 microg/ml) or ACs (2.61 +/- 5.01 microg/ml). For the entire study group, concentrations in plasma at 1 h were less than 1.0, 2.0, and 3.0 microg/ml for 34.7, 60, and 82.7% of the subjects, respectively; concentrations in ELF were less than 1.0, 2.0, and 3.0 microg/ml in 30 (37.5%), 53 (66.0%), and 58 (72.5%) of the subjects, respectively; and concentrations in ACs were less than 1.0, 2.0, and 3.0 microg/ml in 43 (53.8%), 59 (73.8%), and 65 (81.3%) of the subjects, respectively. The concentrations of orally administered isoniazid in plasma were not affected by gender or the presence of AIDS. The concentrations in plasma at 4 h and the concentrations in ELF, but not the concentrations in ACs, were significantly greater in slow acetylators than fast acetylators. Concentrations in plasma and lungs were low compared to recommended therapeutic concentrations in plasma and published MICs of isoniazid for Mycobacterium tuberculosis. The optimal concentrations of isoniazid in ACs and ELF are unknown, but these data suggest that the drug enters these compartments by passive diffusion and achieves concentrations similar to those found in plasma. PMID- 12121906 TI - Potent and selective inhibition of human cytomegalovirus replication by 1263W94, a benzimidazole L-riboside with a unique mode of action. AB - Benzimidazole nucleosides have been shown to be potent inhibitors of human cytomegalovirus (HCMV) replication in vitro. As part of the exploration of structure-activity relationships within this series, we synthesized the 2 isopropylamino derivative (3322W93) of 1H-beta-D-ribofuranoside-2-bromo-5,6 dichlorobenzimidazole (BDCRB) and the biologically unnatural L-sugars corresponding to both compounds. One of the L derivatives, 1H-beta-L ribofuranoside-2-isopropylamino-5,6-dichlorobenzimidazole (1263W94), showed significant antiviral potency in vitro against both laboratory HCMV strains and clinical HCMV isolates, including those resistant to ganciclovir (GCV), foscarnet, and BDCRB. 1263W94 inhibited viral replication in a dose-dependent manner, with a mean 50% inhibitory concentration (IC(50)) of 0.12 +/- 0.01 microM compared to a mean IC(50) for GCV of 0.53 +/- 0.04 microM, as measured by a multicycle DNA hybridization assay. In a single replication cycle, 1263W94 treatment reduced viral DNA synthesis, as well as overall virus yield. HCMV mutants resistant to 1263W94 were isolated, establishing that the target of 1263W94 was a viral gene product. The resistance mutation was mapped to the UL97 open reading frame. The pUL97 protein kinase was strongly inhibited by 1263W94, with 50% inhibition occurring at 3 nM. Although HCMV DNA synthesis was inhibited by 1263W94, the inhibition was not mediated by the inhibition of viral DNA polymerase. The parent benzimidazole D-riboside BDCRB inhibits viral DNA maturation and processing, whereas 1263W94 does not. The mechanism of the antiviral effect of L-riboside 1263W94 is thus distinct from those of GCV and of BDCRB. In summary, 1263W94 inhibits viral replication by a novel mechanism that is not yet completely understood. PMID- 12121907 TI - Preclinical and toxicology studies of 1263W94, a potent and selective inhibitor of human cytomegalovirus replication. AB - 1263W94 is a novel benzimidazole compound being developed for treatment of human cytomegalovirus infection. No adverse pharmacological effects were demonstrated in safety pharmacology studies with 1263W94. The minimal-effect dose in a 1-month rat study was 100 mg/kg/day, and the no-effect dose in a 1-month monkey study was 180 mg/kg/day. Toxic effects were limited to increases in liver weights, neutrophils, and monocytes at higher doses in female rats. 1263W94 was not genotoxic in the Ames or micronucleus assays. In the mouse lymphoma assay, 1263W94 was mutagenic in the absence of the rat liver S-9 metabolic activation system, with equivocal results in the presence of the S-9 mix. Mean oral bioavailability of 1263W94 was >90% in rats and approximately 50% in monkeys. Clearance in rats and monkeys was primarily by biliary secretion, with evidence of enterohepatic recirculation. In 1-month studies in rats and monkeys, mean peak concentrations and exposures to 1263W94 increased in near proportion to dose. Metabolism of 1263W94 to its primary metabolite, an N-dealkylated analog, appeared to be mediated via the isozyme CYP3A4 in humans. 1263W94 was primarily distributed in the gastrointestinal tract of rats but did not cross the blood brain barrier. In monkeys, 1263W94 levels in the brain, cerebrospinal fluid, and vitreous humor ranged from 4 to 20%, 1 to 2%, and <1%, of corresponding concentrations in plasma, respectively. The high level of binding by 1263W94 to human plasma proteins (primarily albumin) was readily reversible, with less protein binding seen in the monkey, rat, and mouse. Results of these studies demonstrate a favorable safety profile for 1263W94. PMID- 12121908 TI - Alkoxyalkyl esters of cidofovir and cyclic cidofovir exhibit multiple-log enhancement of antiviral activity against cytomegalovirus and herpesvirus replication in vitro. AB - The incidence of cytomegalovirus (CMV) retinitis is declining in AIDS patients but remains a significant clinical problem in patients with organ transplants and bone marrow transplants. Prophylaxis with ganciclovir (GCV) or valganciclovir reduces the incidence of CMV disease but may lead to the emergence of drug resistant virus with mutations in the UL97 or UL54 gene. It would be useful to have other types of oral therapy for CMV disease. We synthesized hexadecyloxypropyl and octadecyloxyethyl derivatives of cyclic cidofovir (cCDV) and cidofovir (CDV) and found that these novel analogs had 2.5- to 4-log increases in antiviral activity against CMV compared to the activities of unmodified CDV and cCDV. Multiple-log increases in activity were noted against laboratory CMV strains and various CMV clinical isolates including GCV-resistant strains with mutations in the UL97 and UL54 genes. Preliminary cell studies suggest that the increase in antiviral activity may be partially explained by a much greater cell penetration of the novel analogs. 1-O-Hexadecyloxypropyl-CDV, 1 O-octadecyloxyethyl-CDV, and their corresponding cCDV analogs are worthy of further preclinical evaluation for treatment and prevention of CMV and herpes simplex virus infections in humans. PMID- 12121909 TI - Steady-state pharmacokinetics of lamivudine in human immunodeficiency virus infected patients with end-stage renal disease receiving chronic dialysis. AB - The steady-state pharmacokinetics of lamivudine were evaluated in 11 subjects with human immunodeficiency virus infection and end-stage renal disease, 9 of whom were receiving hemodialysis and 2 of whom were receiving chronic ambulatory peritoneal dialysis (CAPD). All subjects received 150 mg of lamivudine daily for at least 2 weeks prior to sampling for determination of the pharmacokinetics of lamivudine over a 24-h period on 2 consecutive days. On the first day, subjects received 150 mg of oral lamivudine and underwent dialysis (hemodialysis or CAPD). On the second day, subjects received another 150 mg of oral lamivudine but dialysis was not performed. For the subjects undergoing hemodialysis, the geometric mean predose serum lamivudine concentration was 1.14 microg/ml (95% confidence interval [CI], 0.83 to 1.58 microg/ml), the geometric mean maximum concentration in serum (C(max)) was 3.77 microg/ml (95% CI, 3.01 to 4.71 microg/ml), and the geometric mean area under the serum concentration-time curve from time zero to 24 h (AUC(0-24)) was 49.8 microg. h/ml (95% CI 39.1 to 63.6 microg. h/ml). Hemodialysis removed approximately 28 mg of lamivudine but had no significant effect on C(max) or AUC(0-24). In the absence of hemodialysis, the geometric mean lamivudine terminal elimination half-life was 17.2 h (95% CI, 10.5 to 28.1 h), whereas the geometric mean intradialysis half-life of lamivudine was 5.3 h (95% CI, 3.4 to 8.2 h). The pharmacokinetics of lamivudine in subjects undergoing CAPD were similar to those in subjects undergoing hemodialysis. CAPD removed 24 mg of lamivudine over a 24-h period but had no effect on C(max) or AUC(0-24). Pharmacokinetic modeling suggests that a lamivudine dose of 25 mg daily in hemodialysis subjects would provide serum exposure similar to that provided by a dose of 150 mg twice daily in patients with normal renal function. PMID- 12121910 TI - Identification of small-molecule inhibitors of nucleoside triphosphate hydrolase in Toxoplasma gondii. AB - Approximately 150,000 small-molecule compounds were tested by a robotic screening assay for their ability to inhibit nucleoside triphosphate hydrolase (NTPase), a novel enzyme of the tachyzoite form of Toxoplasma gondii. Five unrelated species of compounds were found to inhibit the activities of both NTPase isoforms (NTPase isoform I [NTPase-I] and NTPase-II). The 50% inhibitory concentrations (IC(50)s) ranged from 0.1 to 20 microM, and in general, the IC(50)s were similar for both NTPase isoforms. However, the activity of NTPase-I was 20 times more sensitive than the activity of NTPase-II to one of the inhibitors: 9-hydroxy-10 (pentachlorophenoxy)stearic acid. The five compounds identified also prevented tachyzoite replication in vitro, with IC(50)s ranging from approximately 7 to > or =50 microM. The most effective of these initial compounds, 2-phenylthio indole, was used to identify six additional, structurally related compounds, which were tested for their inhibitory effects on enzyme activities and tachyzoite replication. Surprisingly, these compounds were competitive inhibitors of NTPase-I but noncompetitive inhibitors of NTPase-II. Modifications to the indole and phenol rings resulted in alterations of activity, thus providing insight into the structural features that are important for inhibition of T. gondii NTPases. PMID- 12121911 TI - Class 1 integron containing a new gene cassette, aadA10, associated with Tn1404 from R151. AB - The carbenicillin, gentamicin, kanamycin, streptomycin, spectinomycin, sulfonamide, and tobramycin resistance determinants found on Pseudomonas aeruginosa plasmid R151 have previously been shown to translocate to another plasmid, R388, and it was inferred that a transposon, Tn1404, carried the resistance determinants. Sequencing of the cassette array from the plasmid known as R388::Tn1404 revealed two known gene cassettes, oxa10 and aadB, and a previously unidentified cassette determining resistance to streptomycin and spectinomycin, here designated aadA10, in the order oxa10-aadB-aadA10. These cassettes replaced the dfrB2-orfA cassette array of R388, indicating that movement of the resistance determinants from R151 to R388 resulted from recombinational exchange between two class 1 integrons rather than transposition. The AadA10 protein is most closely related to AadA6 (85% identical) and AadA7 (80% identical). The aadA10 cassette found here has only a simple site containing a 7-bp spacer derived from attI1 in place of a 59-base element and is likely to represent a derivative of the complete cassette. IntI1-mediated deletion of the aadA10 cassette was not detected, indicating that this single simple site is either inactive or only weakly active. PMID- 12121912 TI - Methods for data mining from large multinational surveillance studies. AB - Traditionally, large surveillance studies have been analyzed by the use of the MICs at which 90% of isolates tested are inhibited (MIC(90)s), MIC(50)s, frequency distributions, and percent susceptibility. In the past, these approaches have proved satisfactory for the monitoring of resistance. From these traditional uses, one can readily detect an increase in MICs for organism and drug combinations. Now that large surveillance studies have been conducted for a number of years and databases have grown to include a large number of datum points, new approaches to the extraction of useful information from these studies are needed. The present study proposes approaches, including the use of antibiotypes, principal components analysis, phylogenetics, and population genetic analysis, to the evaluation of data from large multinational surveillance studies. Application of these types of analyses can be used to describe genetic diversity, analyze changes in susceptibility patterns over time, and possibly, shed light on the origins and evolution of antimicrobial resistance. As global surveillance studies become more common and new questions concerning the evolution of resistance are raised, innovative approaches to analysis of the data will increase in importance. PMID- 12121913 TI - Efficacy of intravenous liposomal amphotericin B (AmBisome) against coccidioidal meningitis in rabbits. AB - The efficacy of intravenously administered liposomal amphotericin B (AmBisome [AmBi]) for the treatment of experimental coccidioidal meningitis was compared with those of oral fluconazole (FLC) and intravenously administered conventional amphotericin B (AMB). Male New Zealand White rabbits were infected by intracisternal inoculation of arthroconidia of Coccidioides immitis. Starting 5 days postinfection, animals received one of the following: 5% dextrose water diluent; AMB given at 1 mg/kg of body weight; AmBi given at 7.5, 15, or 22.5 mg/kg intravenously three times per week for 3 weeks; or oral FLC given at 80 mg/kg for 19 days. One week after the cessation of therapy, all survivors were euthanatized, the numbers of CFU remaining in the spinal cord and brain were determined, and histological analyses were performed. All AmBi-, FLC-, or AMB treated animals survived and had prolonged lengths of survival compared with those for the controls (P < 0.0001). Treated groups had significantly lower numbers of white blood cells and significantly lower protein concentrations in the cerebrospinal fluid compared with those for the controls (P < 0.01 to 0.0005) and had fewer clinical signs of infection (e.g., weight loss, elevated temperature, and neurological abnormalities including motor abnormalities). The mean histological scores for AmBi-treated rabbits were lower than those for FLC treated and control rabbits (P < 0.016 and 0.0005, respectively); the scores for AMB-treated animals were lower than those for the controls (P < 0.0005) but were similar to those for FLC-treated rabbits. All regimens reduced the numbers of CFU in the brain and spinal cord compared with those for the controls (P < or =0.0005). AmBi-treated animals had 3- to 11-fold lower numbers of CFU than FLC treated rabbits and 6- to 35-fold lower numbers of CFU than AmB-treated rabbits. Three of eight animals given 15 mg of AmBi per kg had no detectable infection in either tissue, whereas other doses of AmBi or FLC cleared either the brain or the spinal cord of infection in fewer rabbits. In addition, clearance of the infection from both tissues was achieved in none of the rabbits, and neither tissue was cleared of infection in AMB-treated animals. Overall, these data indicate that intravenously administered AmBi is superior to oral FLC or intravenous AMB and that FLC is better than AMB against experimental coccidioidal meningitis. These data indicate that AmBi may offer an improvement in the treatment of coccidioidal meningitis. Additional studies are warranted. PMID- 12121914 TI - Characterization of a novel plasmid-mediated cephalosporinase (CMY-9) and its genetic environment in an Escherichia coli clinical isolate. AB - An Escherichia coli strain, HKYM68, which showed resistance to broad-spectrum cephalosporins was isolated from a sputum specimen in Japan. The high-level resistance of the strain to ceftazidime, cefpirome, and moxalactam was carried by a self-transferable plasmid. The beta-lactamase gene responsible for the resistance was cloned and sequenced. The deduced amino acid sequence of this gene product, CMY-9, had a single amino acid substitution (E85D), the residue reported to be part of the recognition site for the R1 side chain of beta-lactams, compared with the amino acid sequence of CMY-8 and also had 78% identity with the amino acid sequence of CepH, a chromosomal cephalosporinase of Aeromonas hydrophila. A sul1-type class 1 integron containing an aacA1-orfG gene cassette was identified upstream of bla(CMY-9) and ended with a truncated 3' conserved segment. The following 2.1 kb was almost identical to the common region of integrons In6 and In7 and the integron of pSAL-1, except that orf513 encoding a putative transposase was identified instead of orf341 due to addition of a single nucleotide. bla(CMY-9) was closely located downstream of the end of the common region. These observations are indicative of the exogenous derivation of bla(CMY 9) from some environmental microorganisms such as aeromonads. PMID- 12121915 TI - Mutations in cytochrome b resulting in atovaquone resistance are associated with loss of fitness in Plasmodium falciparum. AB - Drug resistance in malarial parasites has become a major obstacle in the control of the disease. Strategies are urgently needed to control the development of resistance and to possibly reverse existing resistance. One key element required to reverse malaria drug resistance is for the parasites to "pay" a biological "cost" or suffer a loss of fitness when acquiring resistance to antimalarial drugs. Such a situation would be a disadvantage to the resistant parasites in the absence of drug pressure. We compared here the relative fitness of atovaquone resistant Plasmodium falciparum K1 clones with single and double base mutations in their cytochrome b genes to their parent clones during erythrocytic stages in the absence of drug pressure. We found that the double amino acid mutation (M133I and G280D) is associated with a 5 to 9% loss of fitness and that the single amino acid change of M133I did not result in any detectable loss of fitness. Molecular modeling of the interaction of P. falciparum cytochrome b with ubiquinone led to the prediction that a loss of fitness of the malaria parasites would result from the G280D mutation due to its close proximity to the putative ubiquinone-binding site. This appears to have resulted in a weakening of the cytochrome b-ubiquinone complex, thereby causing the electron transport chain to become less efficient. Our results suggest that the prevalence of resistant parasites may decrease after the drug usage is discontinued. PMID- 12121916 TI - Immunomodulatory and protective effects of moxifloxacin against Candida albicans induced bronchopneumonia in mice injected with cyclophosphamide. AB - In a previous study, moxifloxacin was shown to ameliorate immunosuppression and enhance cytokine production in several tissues, including the lungs of cyclophosphamide-injected mice. We examined here the effects of moxifloxacin on Candida albicans lung infection in cyclophosphamide-injected mice. Mice were injected on day 0 with 250 mg of cyclophosphamide/kg, and on days 1 to 4 they were given moxifloxacin at 22.5 mg/kg/day compared to controls given ceftazidime at 75 mg/kg/day or saline. On day 6, C. albicans (10 7 CFU/mouse) was inoculated intratracheally, and animals were observed for the development of bronchopneumonia, weight loss, mortality, the presence of C. albicans, and lung cytokine production. Histopathology on day 10 postinoculation revealed bronchopneumonia in 50, 67, and 0% of saline-, ceftazidime-, and moxifloxacin treated mice, respectively (P < 0.05). The mortality rates were 28, 17, and 5%, respectively (P < 0.05), and weight loss occurred at 20, 32, and 0%, respectively (P < 0.05). By day 15, C. albicans was eliminated from all moxifloxacin-treated mice but was still isolated from lung homogenates of 50 to 60% of the saline- and ceftazidime-treated groups. Among the cytokines tested on days 0 to 15, we found an increased production of tumor necrosis factor alpha, KC (functional interleukin-8), and gamma interferon in the lungs of ceftazidime- and saline treated controls compared to the moxifloxacin pretreatment that abolished their secretion. In conclusion, moxifloxacin protected cyclophosphamide-injected mice from C. albicans-induced lung infection and significantly reduced pneumonia, weight loss, and mortality despite the lack of direct antifungal activity. This is most likely due to an immunomodulating activity conferred by moxifloxacin, as shown in this model and in our previous studies. Its potential protective role should be studied in patients undergoing chemotherapy and immune suppression. PMID- 12121917 TI - N-arylsulfonyl hydrazones as inhibitors of IMP-1 metallo-beta-lactamase. AB - Members of a family of N-arylsulfonyl hydrazones have been identified as novel inhibitors of IMP-1, a metallo-beta-lactamase of increasing prevalence. Structure activity relationship studies have indicated a requirement for bulky aromatic substituents on each side of the sulfonyl hydrazone backbone for these compounds to serve as efficient inhibitors of IMP-1. Molecular modeling has provided insight into the structural basis for the anti-metallo-beta-lactamase activity exhibited by this class of compounds. PMID- 12121918 TI - Tetracycline rapidly reaches all the constituent cells of uropathogenic Escherichia coli biofilms. AB - We have developed a method for visualizing Escherichia coli cells that are exposed to tetracycline in a biofilm, based on a previous report that liposomes containing the E. coli TetR(B) protein fluoresce when exposed to this antibiotic. By our method, cells devoid of TetR(B) also exhibited tetracycline-dependent fluorescence. At 50 microg of tetracycline ml(-1), planktonic cells of a uropathogenic E. coli (UPEC) strain developed maximal fluorescence after 7.5 to 10 min of exposure. A similar behavior was exhibited by cells in a 24- or 48-h UPEC biofilm, as examined by confocal laser microscopy, regardless of whether they lined empty spaces or occupied densely packed regions. Further, a comparison of phase-contrast and fluorescent images of corresponding biofilm zones showed that all the cells fluoresced. Thus, all the biofilm cells were exposed to tetracycline and there were no pockets within the biofilm where the antibiotic failed to reach. It also appeared unlikely that niches of reduced exposure to the antibiotic existed within the biofilms. PMID- 12121919 TI - Overexpression of Sbe2p, a Golgi protein, results in resistance to caspofungin in Saccharomyces cerevisiae. AB - Caspofungin inhibits the synthesis of 1, 3-beta-D-glucan, an essential cell wall target in fungi. Genetic studies in the model yeast Saccharomyces cerevisiae have shown that mutations in FKS1 and FKS2 genes result in caspofungin resistance. However, direct demonstration of the role of gene overexpression in caspofungin resistance has been lacking. We transformed wild-type S. cerevisiae with an S. cerevisiae URA3-based GAL1 cDNA library and selected transformants in glucose synthetic complete plates lacking uracil (glucose SC minus uracil plates). We then moved the transformants to galactose SC minus uracil plates containing caspofungin (1 microg/ml) and looked for caspofungin-resistant colonies. We retested the candidates (true positives were sensitive on glucose caspofungin and resistant on galactose caspofungin media, respectively). We identified 16 caspofungin-resistant candidates. Restriction analysis and hybridization confirmed that 15 of the 16 clones were identical. We sequenced one of the cDNA clones and found that it contained the cDNA for SBE2. SBE2 has been described in S. cerevisiae to encode a Golgi protein involved in the transport of cell wall components (B. Santos and M. Snyder, Mol. Biol. Cell, 11:435-452, 2000). The SBE2 cDNA plasmid conferred again galactose-dependent caspofungin resistance when transformed back into the wild-type S. cerevisiae. Finally, the SBE2 deletion mutant was hypersensitive to caspofungin. In conclusion, overexpression of Sbe2p under the regulated control of the GAL1 promoter results in caspofungin resistance in S. cerevisiae. This transport pathway may provide insight into the tolerance or lack of sensitivity to caspofungin of some pathogenic fungi. PMID- 12121921 TI - Comparison of visual and spectrophotometric methods of broth microdilution MIC end point determination and evaluation of a sterol quantitation method for in vitro susceptibility testing of fluconazole and itraconazole against trailing and nontrailing Candida isolates. AB - Visual determination of MIC end points for azole antifungal agents can be complicated by the trailing growth phenomenon. To determine the incidence of trailing growth, we performed testing of in vitro susceptibility to fluconazole and itraconazole using the National Committee for Clinical Laboratory Standards broth microdilution M27-A reference procedure and 944 bloodstream isolates of seven Candida spp., obtained through active population-based surveillance between 1998 and 2000. Of 429 C. albicans isolates, 78 (18.2%) showed trailing growth at 48 h in tests with fluconazole, and 70 (16.3%) showed trailing in tests with itraconazole. Of 118 C. tropicalis isolates, 70 (59.3%) showed trailing growth in tests with fluconazole, and 35 (29.7%) showed trailing in tests with itraconazole. Trailing growth was not observed with any of the other five Candida spp. tested (C. dubliniensis, C. glabrata, C. krusei, C. lusitaniae, and C. parapsilosis). To confirm whether or not isolates that showed trailing growth in fluconazole and/or itraconazole were resistant in vitro to these agents, all isolates that showed trailing growth were retested by the sterol quantitation method, which measures cellular ergosterol content rather than growth inhibition after exposure to azoles. By this method, none of the trailing isolates was resistant in vitro to fluconazole or itraconazole. For both agents, a 24-h visual end point or a spectrophotometric end point of 50% reduction in growth relative to the growth control after 24 or 48 h of incubation correlated most closely with the result of sterol quantitation. Our results indicate that MIC results determined by either of these end point rules may be more predictive of in vivo outcome for isolates that give unclear visual end points at 48 h due to trailing growth. PMID- 12121920 TI - Inhibition of cyclin-dependent kinase 1 by purines and pyrrolo[2,3-d]pyrimidines does not correlate with antiviral activity. AB - We have previously shown that a series of nonnucleoside pyrrolo[2,3-d]pyrimidines selectively inhibit the replication of herpes simplex virus type 1 (HSV-1) and human cytomegalovirus (HCMV). These compounds act at the immediate-early or early stage of HCMV replication and have antiviral properties somewhat similar to those of roscovitine and olomoucine, specific inhibitors of cyclin-dependent kinases (cdks). In the present study we examine the hypothesis that pyrrolo[2,3 d]pyrimidines exert their antiviral effects by inhibition of cellular cdks. Much higher concentrations of a panel of pyrrolo[2,3-d]pyrimidine nucleoside analogs with antiviral activity were required to inhibit recombinant cdk1/cyclin B compared to the submicromolar concentrations required to inhibit HCMV and HSV-1 replication. 4,6-Diamino-5-cyano-7-(2-phenylethyl)pyrrolo[2,3-d]pyrimidine (compound 1369) was the best inhibitor of cdk1 and cyclin B, with a 50% inhibitory concentration (IC(50); 14 microM) similar to that of roscovitine; it was competitive with respect to ATP (K(i) = 14 microM). The potency of compound 1369 against cdk1 and cyclin B was similar to its cytotoxicity (IC(50)s, 32 to 100 microM) but not its antiviral efficacy (IC(50)s, 0.02 to 0.3 microM). Thus, our results indicated the null hypothesis. In contrast, roscovitine was only weakly active against HSV-1 (IC(50), 38 microM) and HCMV (IC(50), 40 microM). These values were similar to those derived by cytotoxicity and cell growth inhibition assays, thereby suggesting that roscovitine is not a selective antiviral. Therefore, we propose that inhibition of cdk1 and cyclin B is not responsible for selective antiviral activity and that pyrrolo[2,3-d]pyrimidines constitute novel pharmacophores which compete with ATP to inhibit cdk1 and cyclin B. PMID- 12121923 TI - Stress-based identification and classification of antibacterial agents: second generation Escherichia coli reporter strains and optimization of detection. AB - Escherichia coli strains bearing single-copy fusions between the lacZ reporter gene and the cspA, ibp, or P3rpoH stress promoters offer a simple means to detect sublethal concentrations of antibacterial agents interfering with prokaryotic translation or cell envelope integrity while simultaneously providing information on the mechanism of action of the test compound (A. A. Bianchi and F. Baneyx, Appl. Environ. Microbiol. 65:5023-5027, 1999). Here, we expand the usefulness of this system by (i) demonstrating that a fusion between the SOS-inducible sulA promoter and lacZ is a highly specific probe for the detection of antimicrobial agents that ultimately interfere with DNA replication, (ii) showing that inactivation of the tolC gene allows efficient detection of very low concentrations of model antibiotics (including aminoglycosides) whereas polymyxin B-mediated outer membrane permeabilization facilitates the identification of intermediate concentrations of hydrophobic compounds, and (iii) validating the potential of detector strains and sensitization strategies for high-throughput screening using a reproducible and internally consistent 96-well microplate assay. PMID- 12121922 TI - Sulfadoxine-pyrimethamine resistance in the rodent malaria parasite Plasmodium chabaudi. AB - We have studied resistance to sulfadoxine-pyrimethamine (S/P) in the rodent malaria parasite Plasmodium chabaudi. A stable S/P-resistant mutant, AS(50S/P), was selected by drug treatment of a clone, AS(PYR), already resistant to pyrimethamine. The sequences of the P. chabaudi dhfr and dhps genes were obtained and found to be identical in AS(50S/P) and AS(PYR), showing that resistance to S/P in AS(50S/P) was not due to additional mutations in either gene. AS(50S/P) was crossed with a drug-sensitive clone, AJ, and 16 independent recombinant progeny were obtained. These clones were phenotyped for their susceptibility to S/P and to sulfadoxine and pyrimethamine separately. Pyrimethamine resistance was invariably associated with S/P resistance, but no correlation was found between resistance to S/P and resistance to sulfadoxine. Quantitative trait locus analysis of the progeny with 31 chromosome-specific markers showed that mutant P. chabaudi dhfr, or one or more genes closely linked to it, was a major determinant of S/P resistance. In addition, the inheritance of genes on chromosomes 5 and 13 from the sensitive parent appeared to contribute to the level of resistance observed. These results demonstrate that the S/P resistance of the AS(50S/P) mutant of P. chabaudi does not involve mutation in dhps and is not due simply to a combination of two genes determining resistance to pyrimethamine and sulfadoxine separately. PMID- 12121925 TI - PCR-based ordered genomic libraries: a new approach to drug target identification for Streptococcus pneumoniae. AB - Described here are the development and validation of a novel approach to identify genes encoding drug targets in Streptococcus pneumoniae. The method relies on the use of an ordered genomic library composed of PCR amplicons that were generated under error-prone conditions so as to introduce random mutations into the DNA. Since some of the mutations occur in drug target-encoding genes and subsequently affect the binding of the drug to its respective cellular target, amplicons containing drug targets can be identified as those producing drug-resistant colonies when transformed into S. pneumoniae. Examination of the genetic content of the amplicon giving resistance coupled with bioinformatics and additional genetic approaches could be used to rapidly identify candidate drug target genes. The utility of this approach was verified by using a number of known antibiotics. For drugs with single protein targets, amplicons were identified that rendered S. pneumoniae drug resistant. Assessment of amplicon composition revealed that each of the relevant amplicons contained the gene encoding the known target for the particular drug tested. Fusidic acid-resistant mutants that resulted from the transformation of S. pneumoniae with amplicons containing fusA were further characterized by sequence analysis. A single mutation was found to occur in a region of the S. pneumoniae elongation factor G protein that is analogous to that already implicated in other bacteria as being associated with fusidic acid resistance. Thus, in addition to facilitating the identification of genes encoding drug targets, this method could provide strains that aid future mechanistic studies. PMID- 12121924 TI - Small-colony mutants of Staphylococcus aureus allow selection of gyrase-mediated resistance to dual-target fluoroquinolones. AB - Fluoroquinolones acting equally through DNA gyrase and topoisomerase IV in vivo are considered desirable in requiring two target mutations for emergence of resistant bacteria. To investigate this idea, we have studied the response of Staphylococcus aureus RN4220 to stepwise challenge with sparfloxacin, a known dual-target agent, and with NSFQ-105, a more potent sulfanilyl fluoroquinolone that behaves similarly. First-step mutants were obtained with both drugs but only at the MIC. These mutants exhibited distinctive small-colony phenotypes and two- to fourfold increases in MICs of NSFQ-105, sparfloxacin, and ciprofloxacin. No changes were detected in the quinolone resistance-determining regions of the gyrA, gyrB, grlA, or grlB gene. Quinolone-induced small-colony mutants shared the delayed coagulase response but not the requirement for menadione, hemin, or thymidine characteristic of small-colony variants, a subpopulation of S. aureus that is often defective in electron transport. Second-step mutants selected with NSFQ-105 had gyrA(S84L) alterations; those obtained with sparfloxacin carried a gyrA(D83A) mutation or a novel gyrB deletion (DeltaRKSAL, residues 405 to 409) affecting a trypsin-sensitive region linking functional domains of S. aureus GyrB. Each mutation was associated with four- to eightfold increases in MICs of NSFQ-105 and sparfloxacin, but not of ciprofloxacin, which we confirm targets topoisomerase IV. The presence of wild-type grlB-grlA gene sequences in second step mutants excluded involvement of topoisomerase IV in the small-colony phenotype. Growth revertants retaining mutant gyrA or gyrB alleles were quinolone susceptible, indicating that resistance to NSFQ-105 and sparfloxacin was contingent on the small-colony mutation. We propose that small-colony mutations unbalance target sensitivities, perhaps through altered ATP or topoisomerase levels, such that gyrase becomes the primary drug target. Breaking of target parity by genetic or physiological means eliminates the need for two target mutations and provides a novel mechanism for stepwise selection of quinolone resistance. PMID- 12121927 TI - Assessment of azithromycin in combination with other antimalarial drugs against Plasmodium falciparum in vitro. AB - Initial field malaria prophylaxis trials with azithromycin revealed insufficient efficacy against falciparum malaria to develop azithromycin as a single agent. The objective of this in vitro study was to determine the best drug combination(s) to evaluate for future malaria treatment and prophylaxis field trials. In vitro, azithromycin was tested in combination with chloroquine against 10 representative Plasmodium falciparum isolates. Azithromycin was also assessed in combination with eight additional antimalarial agents against two or three multidrug-resistant P. falciparum isolates. Parasite susceptibility testing was carried out with a modification of the semiautomated microdilution technique. The incubation period was extended from the usual 48 h to 68 h. Fifty percent inhibitory concentrations (IC(50)s) were calculated for each drug alone and for drugs in fixed combinations of their respective IC(50)s (1:1, 3:1, 1:3, 4:1, 1:4, and 5:1). These data were used to calculate fractional inhibitory concentrations and isobolograms. Chloroquine-azithromycin studies revealed a range of activity from additive to synergistic interactions for the eight chloroquine-resistant isolates tested, while an additive response was seen for the two chloroquine sensitive isolates. Quinine, tafenoquine, and primaquine were additive to synergistic with azithromycin, while dihydroartemisinin was additive with a trend toward antagonism. The remaining interactions appeared to be additive. These results suggest that a chloroquine-azithromycin combination should be evaluated for malaria prophylaxis and that a quinine-azithromycin combination should be evaluated for malaria treatment in areas of drug resistance. PMID- 12121926 TI - Distribution of mef(A) in gram-positive bacteria from healthy Portuguese children. AB - We screened 615 gram-positive isolates from 150 healthy children for the presence of the erm(A), erm(B), erm(C), erm(F), and mef(A) genes. The mef(A) genes were found in 20 (9%) of the macrolide-resistant isolates, including Enterococcus spp., Staphylococcus spp., and Streptococcus spp. Sixteen of the 19 gram-positive isolates tested carried the other seven open reading frames (ORFs) described in Tn1207.1, a genetic element carrying mef(A) recently described in Streptococcus pneumoniae. The three Staphylococcus spp. did not carry orf1 to orf3. A gram negative Acinetobacter junii isolate also carried the other seven ORFs described in Tn1207.1. A Staphylococcus aureus isolate, a Streptococcus intermedius isolate, a Streptococcus sp. isolate, and an Enterococcus sp. isolate had their mef(A) genes completely sequenced and showed 100% identity at the DNA and amino acid levels with the mef(A) gene from S. pneumoniae. PMID- 12121928 TI - Efficacies of entecavir against lamivudine-resistant hepatitis B virus replication and recombinant polymerases in vitro. AB - Entecavir (ETV) is a potent and selective inhibitor of hepatitis B virus (HBV) replication in vitro and in vivo that is currently in clinical trials for the treatment of chronic HBV infections. A major limitation of the current HBV antiviral therapy, lamivudine (3TC), is the emergence of drug-resistant HBV in a majority of treated patients due to specific mutations in the nucleotide binding site of HBV DNA polymerase (HBV Pol). To determine the effects of 3TC resistance mutations on inhibition by ETV triphosphate (ETV-TP), a series of in vitro studies were performed. The inhibition of wild-type and 3TC-resistant HBV Pol by ETV-TP was measured using recombinant HBV nucleocapsids, and compared to that of 3TC-TP. These enzyme inhibition studies demonstrated that ETV-TP is a highly potent inhibitor of wild-type HBV Pol and is 100- to 300-fold more potent than 3TC-TP against 3TC-resistant HBV Pol. Cell culture assays were used to gauge the potential for antiviral cross-resistance of 3TC-resistant mutants to ETV. Results demonstrated that ETV inhibited the replication of 3TC-resistant HBV, but 20- to 30-fold higher concentrations were required. To gain further perspective regarding the potential therapeutic use of ETV, its phosphorylation was examined in hepatoma cells treated with extracellular concentrations representative of drug levels in plasma in ETV-treated patients. At these concentrations, intracellular ETV-TP accumulated to levels expected to inhibit the enzyme activity of both wild-type and 3TC-resistant HBV Pol. These findings are predictive of potent antiviral activity of ETV against both wild-type and 3TC resistant HBV. PMID- 12121929 TI - Simple fibroblast-based assay for screening of new antimicrobial drugs against Mycobacterium tuberculosis. AB - In this study, we propose a simple and reproducible host-cell-based assay for the screening of antimycobacterial drugs that is suitable for drug discovery. The method evaluates both antimycobacterial activity of the drugs and their cytotoxicity to host cells. The basis of this simple fibroblast-based assay (SFA) is that cells of human lung fibroblast cell line MRC-5, which are highly sensitive to mycobacterial cytotoxicity, are killed by virulent Mycobacterium tuberculosis strain H(37)Rv bacilli in response to the viability of bacilli. Clinically used antimycobacterial drugs inhibited the mycobacterial cytotoxicity to MRC-5 cells in a dose-dependent manner. MICs of isoniazid, streptomycin, rifampin, and ethambutol determined by this SFA (0.428, 1.816, 0.013, and 3.465 microg/ml, respectively) were within 1 log of MICs determined by the broth dilution test (BDT) using Middlebrook 7H9 medium. The MIC of pyrazinamide, which exhibits bactericidal activity only at a high dose by BDT (1,231 microg/ml at pH 6.6 and 492 microg/ml at pH 5.8), was 3.847 microg/ml in the modified method of SFA. On the other hand, sodium azide, a toxic agent for both mammalian cells and bacteria, exhibited cytotoxicity to fibroblasts at a dose lower than that required to inhibit mycobacterial growth. Thus, this fibroblast-based method enabled us to evaluate both antibacterial activity of drugs and their cytotoxicity to human cells within a short period of time. PMID- 12121930 TI - Trends in antimicrobial resistance among urinary tract infection isolates of Escherichia coli from female outpatients in the United States. AB - The Infectious Diseases Society of America advocates trimethoprim sulfamethoxazole (SXT) as initial therapy for females with acute uncomplicated bacterial cystitis in settings where the prevalence of SXT resistance does not exceed 10 to 20%. To determine trends in the activities of SXT, ampicillin, ciprofloxacin, and nitrofurantoin among urine isolates of Escherichia coli from female outpatients, susceptibility testing data from The Surveillance Network (TSN) Database-USA (n = 286,187) from 1995 to 2001 were analyzed. Resistance rates among E. coli isolates to ampicillin (range, 36.0 to 37.4% per year), SXT (range, 14.8 to 17.0%), ciprofloxacin (range, 0.7 to 2.5%), and nitrofurantoin (range, 0.4 to 0.8%) varied only slightly over this 7-year period. Ciprofloxacin was the only agent studied that demonstrated a consistent stepwise increase in resistance from 1995 (0.7%) to 2001 (2.5%). In 2001, SXT resistance among E. coli isolates was >10% in all nine U.S. Bureau of the Census regions. At institutions testing > or =100 urinary isolates of E. coli (n = 126) in 2001, ampicillin (range, 27.3 to 98.8%) and SXT (range, 7.5 to 47.1%) resistance rates varied widely while ciprofloxacin (range, 0 to 12.9%) and nitrofurantoin (range, 0 to 2.8%) resistance rates were more consistent. In 2001, the most frequent coresistant phenotypes were resistance to ampicillin and SXT (12.0% of all isolates; 82.3% of coresistant isolates) and resistance to ampicillin, ciprofloxacin, and SXT (1.4% of all isolates; 9.9% of coresistant isolates). Coresistance less frequently included resistance to nitrofurantoin (3.5% of coresistant isolates) than resistance to ciprofloxacin (15.8%), SXT (95.7%), and ampicillin (98.1%). In conclusion, among urinary isolates of E. coli from female outpatients in the United States, national resistance rates to SXT were relatively consistent (14.8 to 17.0%) from 1995 to 2001 but demonstrated considerable regional and institutional variation in 2001. Therapies other than SXT may need to be considered in some locations. PMID- 12121931 TI - Pharmacokinetics and safety of voriconazole following intravenous- to oral-dose escalation regimens. AB - In this study, the safety, tolerability, and pharmacokinetics of intravenous (i.v.)- to oral-dose regimens of voriconazole were evaluated with a group of 42 healthy men, 41 of whom completed the study. Two cohorts of subjects participated in the study. Cohort 1 (n = 28) took part in two study periods, each consisting of 14 days separated by a minimum 7-day washout. In one of the periods, 14 subjects received 6 mg/kg i.v. twice a day (b.i.d.) on day 1 followed by 3 mg/kg i.v. b.i.d. on days 2 to 7 and were then switched to 200 mg orally b.i.d. for days 8 to 14. In the other period, subjects received 6 mg/kg i.v. b.i.d. on day 1 followed by 5 mg/kg i.v. b.i.d. on days 2 to 7 and were then switched to 400 mg orally b.i.d. for days 8 to 14. The remaining 14 subjects in cohort 1 received a matching placebo throughout the study. In cohort 2 (n = 14), 7 subjects received 6 mg/kg i.v. b.i.d. on day 1 followed by 4 mg/kg i.v. b.i.d. on days 2 to 7 and were then switched to 300 mg orally b.i.d. for days 8 to 14. The remaining seven subjects in cohort 2 received a matching placebo. Blood samples were taken prior to dosing on days 1 to 6 and on days 8 to 13. Blood samples were drawn prior to dosing and at frequent intervals up to 12 h following the morning dose on days 7 and 14 of each study period. The samples were assayed for voriconazole by a high performance liquid chromatography method. The maximum concentration in plasma (C(max)) occurred at the end of the 1-h i.v. infusion and between 1.4 and 1.8 h after oral administration. Voriconazole exhibited nonlinear pharmacokinetics, possibly due to saturable metabolism. For cohort 1, both C(max) and the area under the concentration-time curve within a dosage interval (AUC(tau)) increased disproportionately with dose for both i.v. and oral dosing. For i.v. dosing, a 1.7-fold increase in dose resulted in 2.4- and 3.1-fold increases in C(max) and AUC(tau), respectively. Similarly, a 2-fold increase in oral dosing resulted in 2.8- and 3.9-fold increases in C(max) and AUC(tau), respectively. The mean values for C(max) observed following oral dosing were lower than those obtained after i.v. administration, ranging from 62.7 to 89.6% of the i.v. value. After the switch from i.v. to oral dosing, most subjects achieved steady state by day 4, and mean minimum concentrations in plasma remained above clinically important MICs. The pharmacokinetic profiles for saliva followed a pattern similar to those observed for plasma; there was a highly significant correlation between plasma and saliva voriconazole concentrations (P < 0.0001). Voriconazole was well tolerated; the most commonly reported adverse events in voriconazole-treated subjects were mild to moderate headache, rash, and abnormal vision. Visual function tests detected no further abnormalities during voriconazole treatment. PMID- 12121932 TI - Safety, pharmacokinetics, and pharmacodynamics of cyclodextrin itraconazole in pediatric patients with oropharyngeal candidiasis. AB - The safety, pharmacokinetics, and pharmacodynamics of cyclodextrin itraconazole (CD-ITRA) oral suspension were investigated in an open sequential dose escalation study with 26 human immunodeficiency virus (HIV)-infected children and adolescents (5 to 18 years old; mean CD4(+)-cell count, 128/microl) with oropharyngeal candidiasis (OPC). Patients received CD-ITRA at either 2.5 mg/kg of body weight once a day (QD) or 2.5 mg/kg twice a day (BID) for a total of 15 days. Pharmacokinetic sampling was performed after the first dose and for up to 120 h after the last dose, and antifungal efficacy was evaluated by standardized scoring of the oropharynx. Apart from mild to moderate gastrointestinal disturbances in three patients (11.5%), CD-ITRA was well tolerated. Two patients (7.6%) discontinued treatment prematurely due to study drug-related adverse events. After 15 days of treatment, the peak concentration of drug in plasma (C(max)), the area under the plasma concentration-time curve (AUC) from 0 to 24 h (AUC(0-24)), the concentration in plasma at the end of the dosing interval (predose) (C(min)), and the terminal half-life of itraconazole (ITRA) were (means and standard deviations) 0.604 +/- 0.53 microg/ml, 6.80 +/- 7.4 microg. h/ml, 0.192 +/- 0.06 microg/ml, and 56.48 +/- 44 h, respectively, for the QD regimen and 1.340 +/- 0.75 microg/ml, 23.04 +/- 14.5 microg. h/ml, 0.782 +/- 0.19 microg/ml, and 104.22 +/- 94 h, respectively, for the BID regimen. The mean AUC based accumulation factors for ITRA on day 15 were 4.14 +/- 0.9 and 3.53 +/- 0.6, respectively. A comparison of the dose-normalized median AUC of the two dosage regimens revealed a trend toward nonlinear drug disposition (P = 0.05). The mean metabolic ratios (AUC of hydroxyitraconazole/AUC of ITRA) at day 15 were 1.96 +/- 0.1 for the QD regimen and 1.29 +/- 0.2 for the BID regimen, respectively (P < 0.05). The OPC score (range, 0 to 13) for all 26 patients decreased from a mean of 7.46 +/- 0.8 at baseline to 2.8 +/- 0.7 at the end of therapy (P < 0.001), demonstrating antifungal efficacy in this setting. The relationships among C(max), C(min), AUC(0-12), C(max)/MIC, C(min)/MIC, AUC(0-12)/MIC, time during the dosing interval when the plasma drug concentrations were above the MIC for the infecting isolate, and the residual OPC score at day 15 for the entire study population fit inhibitory effect pharmacodynamic models (r, 0.595 to 0.421; P, <0.01 to <0.05). All patients with fluconazole-resistant isolates responded to treatment with CD-ITRA; however, there was no clear correlation between the MIC of ITRA and response to therapy. In conclusion, CD-ITRA was well tolerated and efficacious for the treatment of OPC in HIV-infected pediatric patients. Pharmacodynamic modeling revealed significant correlations between plasma drug concentrations and antifungal efficacy. Based on this documented safety and efficacy, a dosage of 2.5 mg/kg BID can be recommended for the treatment of OPC in pediatric patients > or =5 years old. PMID- 12121933 TI - Efficacy of caspofungin alone and in combination with voriconazole in a Guinea pig model of invasive aspergillosis. AB - The antifungal activity of caspofungin acetate (CAS) alone and in combination with voriconazole (VRC) was evaluated in an immunosuppressed transiently neutropenic guinea pig model of invasive aspergillosis. Guinea pigs were immunosuppressed with triamcinolone at 20 mg/kg of body weight/day subcutaneously beginning 4 days prior to lethal intravenous challenge with Aspergillus fumigatus and were made temporarily neutropenic with cyclophosphamide administered at 150 mg/kg intraperitoneally (i.p.) 1 day prior to challenge. Therapy with i.p. CAS at 1 and 2.5 mg/kg/day (with and without oral VRC at 5 mg/kg/day), oral VRC at 5 mg/kg/day, or i.p. amphotericin B (AMB) at 1.25 mg/kg/day was begun 24 h after challenge and was continued for 5 days. Mortality occurred in 12 of 12 untreated controls, whereas mortality occurred in 4 of 12 and 6 of 12 guinea pigs treated with CAS at 1 and 2.5 mg/kg/day, respectively, and in 3 of 12 guinea pigs treated with AMB. No mortality occurred among animals treated with CAS at 1 mg/kg/day plus VRC at 5 mg/kg/day, CAS at 2.5 mg/kg/day plus VRC at 5 mg/kg/day, or VRC at 5 mg/kg/day alone. Both CAS regimens increased the survival times and reduced the colony counts in tissue compared with those for the controls. Treatment with VRC and AMB significantly reduced the colony counts in the tissues of selected animals compared with those in the tissues of the controls. Treatment with VRC and AMB also resulted in reductions in colony counts in tissues compared with those in the tissues of animals treated with CAS (the difference was not statistically significant) and improved the survival times but did not sterilize tissues. Combination therapies with CAS plus VRC at either dose reduced colony counts in tissues 1,000-fold over those for the controls and were the only regimens that significantly reduced the numbers of positive cultures. The combinations of CAS plus VRC were highly effective in this model and should be further evaluated for use against invasive aspergillosis. PMID- 12121934 TI - Antileishmanial activity of the antiulcer agent omeprazole. AB - The benzimidazole compound omeprazole, used widely for the treatment of peptic ulcer disease, inhibits the growth of Leishmania donovani, the causative agent of visceral leishmaniasis. Promastigotes cultured at acidic pH and amastigotes within infected macrophages are reduced 90% or more with 150 microM omeprazole. Antiparasitic action of the drug is due to its inhibition of the P-type K(+),H(+) ATPase on the surface membrane. This enzyme is important for pH homeostasis and the maintenance of proton motive force across the membrane in Leishmania. The drug is effective only at acidic pH, a condition that mimics the in vivo environment within the phagolysosomal vesicles where the amastigote form of the parasite resides. Omeprazole deserves consideration as an alternative to currently available chemotherapeutics, which have severe toxic side effects. PMID- 12121935 TI - Ciprofloxacin in polyethylene glycol-coated liposomes: efficacy in rat models of acute or chronic Pseudomonas aeruginosa infection. AB - In a previous study in experimental Klebsiella pneumoniae pneumonia, the therapeutic potential of ciprofloxacin was significantly improved by encapsulation in polyethylene glycol-coated ("pegylated") long-circulating (STEALTH) liposomes. Pegylated liposomal ciprofloxacin in high doses was nontoxic and resulted in relatively high and sustained ciprofloxacin concentrations in blood and tissues, and hence an increase in the area under the plasma concentration-time curve (AUC). These data correspond to data from animal and clinical studies showing that for fluoroquinolones the AUC/MIC ratio is associated with favorable outcome in serious infections. Clinical failures and the development of resistance are observed for marginally susceptible organisms like Pseudomonas aeruginosa and for which sufficient AUC/MIC ratios cannot be achieved. In the present study the therapeutic efficacy of pegylated liposomal ciprofloxacin was investigated in two rat models of Pseudomonas aeruginosa pneumonia. In the acute model pneumonia developed progressively, resulting in a rapid onset of septicemia and a high mortality rate. Ciprofloxacin twice daily for 7 days was not effective at doses at or below the maximum tolerated dose (MTD). However, pegylated liposomal ciprofloxacin either at high dosage or given at low dosage in combination with free ciprofloxacin on the first day of treatment was fully effective (100% survival). Obviously, prolonged concentrations of ciprofloxacin in blood prevented death of the animals due to early-stage septicemia in this acute infection. However, bacterial eradication from the left lung was not effected. In the chronic model, pneumonia was characterized by bacterial persistence in the lung without bacteremia, and no signs of morbidity or mortality were observed. Ciprofloxacin administered for 7 days at the MTD twice daily resulted in killing of more than 99% of bacteria in the lung; this result can also be achieved with pegylated liposomal ciprofloxacin given once daily. Complete bacterial eradication is never observed. PMID- 12121936 TI - DNA gyrase and topoisomerase IV mutations associated with fluoroquinolone resistance in Proteus mirabilis. AB - Mutations associated with fluoroquinolone resistance in clinical isolates of Proteus mirabilis were determined by genetic analysis of the quinolone resistance determining region (QRDR) of gyrA, gyrB, parC, and parE. This study included the P. mirabilis type strain ATCC 29906 and 29 clinical isolates with reduced susceptibility (MIC, 0.5 to 2 microg/ml) or resistance (MIC, > or =4 microg/ml) to ciprofloxacin. Susceptibility profiles for ciprofloxacin, clinafloxacin, gatifloxacin, gemifloxacin, levofloxacin, moxifloxacin, and trovafloxacin were correlated with amino acid changes in the QRDRs. Decreased susceptibility and resistance were associated with double mutations involving both gyrA (S83R or -I) and parC (S80R or -I). Among these double mutants, MICs of ciprofloxacin varied from 1 to 16 microg/ml, indicating that additional factors, such as drug efflux or porin changes, also contribute to the level of resistance. For ParE, a single conservative change of V364I was detected in seven strains. An unexpected result was the association of gyrB mutations with high-level resistance to fluoroquinolones in 12 of 20 ciprofloxacin-resistant isolates. Changes in GyrB included S464Y (six isolates), S464F (three isolates), and E466D (two isolates). A three-nucleotide insertion, resulting in an additional lysine residue between K455 and A456, was detected in gyrB of one strain. Unlike any other bacterial species analyzed to date, mutation of gyrB appears to be a frequent event in the acquisition of fluoroquinolone resistance among clinical isolates of P. mirabilis. PMID- 12121937 TI - In vitro activities of position 2 substitution-bearing 6-nitro- and 6-amino benzothiazoles and their corresponding anthranilic acid derivatives against Leishmania infantum and Trichomonas vaginalis. AB - 6-Nitro- and 6-amino-benzothiazoles bearing different chains in position 2 and their corresponding anthranilic acid derivatives were investigated for their in vitro antiparasitic properties against parasites of the species Leishmania infantum and Trichomonas vaginalis compared to their toxicity towards human monocytes. Biological investigations established that the antiprotozoal properties depended greatly on the chemical structure of the position 2 substitution-bearing group. Compound C1, 2-[(2-chloro-benzothiazol-6-yl) amino] benzoic acid, demonstrated an interesting antiproliferative activity towards parasites of the species T. vaginalis, while compound C11, 2-([2-[(2 hydroxyethyl) amino]-benzothiazol-6-yl] amino) benzoic acid, exhibited a promising activity against parasites of the species L. infantum in their intracellular amastigote form. Additional experiments established that compound C11, which was poorly toxic against the promastigote and the extracellular amastigote forms of the parasite, could improve host-protective mechanisms against Leishmania by preventing parasite internalization by macrophages and stimulating NO production, by means of a mechanism synergistically enhanced by the presence of gamma interferon. PMID- 12121938 TI - In vitro and in vivo activities of tigecycline (GAR-936), daptomycin, and comparative antimicrobial agents against glycopeptide-intermediate Staphylococcus aureus and other resistant gram-positive pathogens. AB - Tigecycline (GAR-936) and daptomycin are potent antibacterial compounds in advanced stages of clinical trials. These novel agents target multiply resistant pathogenic bacteria. Daptomycin is principally active against gram-positive bacteria, while tigecycline has broad-spectrum activity. When tested by the standard protocols of the National Committee for Clinical Laboratory Standards in Mueller-Hinton broth II, tigecycline was more active than daptomycin (MICs at which 90% of isolates tested are inhibited, 0.12 to 1 and 0.5 to 16 microg/ml, respectively) against staphylococcal, enterococcal, and streptococcal pathogens. Daptomycin demonstrated a stepwise increase in activity corresponding to an increase in the supplemental concentration of calcium. When tested in base Mueller-Hinton broth supplemented with 50 mg of calcium per liter, daptomycin demonstrated improved activity (MIC(90)s, 0.015 to 4 microg/ml). The activity of daptomycin, however, equaled that of tigecycline against the glycopeptide intermediate Staphylococcus aureus (GISA) strains only when the test medium was supplemented with excess calcium (75 mg/liter). Tigecycline and daptomycin demonstrated in vivo efficacies against GISA, methicillin-resistant S. aureus, and methicillin-susceptible S. aureus strains in an intraperitoneal systemic murine infection model. These data suggest that tigecycline and daptomycin may offer therapeutic options against clinically relevant resistant pathogens for which current alternatives for treatment are limited. PMID- 12121939 TI - Novel nucleoside analogue MCC-478 (LY582563) is effective against wild-type or lamivudine-resistant hepatitis B virus. AB - The emergence of resistant hepatitis B virus (HBV) with the L528M mutation and/or the M552V and M552I mutations in the polymerase gene following long-term lamivudine treatment is becoming an important clinical problem. The aim of this study was to investigate the susceptibility of wild-type and lamivudine-resistant HBV to MCC-478 (LY582563), a novel nucleoside analogue derivative of phosphonomethoxyethyl purine. The susceptibility of wild-type HBV and lamivudine resistant mutants (M552I, M552V, and L528M/M552V) to MCC-478 was examined by transient transfection of full-length HBV DNA into human hepatoma cells. HBV DNA replication was monitored by Southern blot hybridization, and the effective concentration required to reduce replication by 50% (EC(50)) was determined. The replicative intermediates of wild-type and lamivudine-resistant mutants were progressively diminished by treatment with increasing doses of MCC-478. The MCC 478 EC(50)s were 0.027 microM for wild-type HBV (about 20 times more efficient than lamivudine), 2.6 microM for M552I, 3.3 microM for M552V, and 2.0 microM for L528M/M552V. Wild-type HBV and lamivudine-resistant mutants are susceptible to MCC-478. MCC-478 appears to be a candidate for the treatment of HBV infection and exhibits potent activity against lamivudine-resistant HBV. PMID- 12121940 TI - In vitro activities of quinupristin-dalfopristin and cefepime, alone and in combination with various antimicrobials, against multidrug-resistant staphylococci and enterococci in an in vitro pharmacodynamic model. AB - Use of combinations of antimicrobials that together achieve synergistic activities against targeted microorganisms is one potential strategy for overcoming bacterial resistance. As the incidence of infections caused by multidrug-resistant staphylococci and enterococci increases, the importance of devising additional synergistic drug combinations for these bacteria is magnified. We evaluated a number of antimicrobial combinations, with a focus on quinupristin-dalfopristin (Q-D), cefepime, and linezolid, using a previously described in vitro pharmacodynamic model. The combination of Q-D with either linezolid or vancomycin, as well as the combination of cefepime-vancomycin, resulted in enhanced killing (> or =2-log(10) increase in killing versus the most active single agent) against methicillin-resistant Staphylococcus aureus (MRSA) 494. An improved effect (<2 log(10) kill increase in kill) against MRSA 494 was noted for cefepime plus either Q-D or linezolid, as well as linezolid-vancomycin. Similar relationships were observed for a methicillin-susceptible S. aureus isolate (isolate 1199). Against methicillin-resistant S. epidermidis R444, enhanced killing was achieved with the combination of cefepime-linezolid, while improvement was noted for vancomycin with either cefepime or linezolid. The combination of cefepime and vancomycin also achieved enhanced killing against a glycopeptide-intermediate-susceptible S. aureus isolate (isolate 992). The combination of linezolid and doxycycline achieved an enhanced effect against vancomycin-resistant Enterococcus faecalis (VREFc) and E. faecium. Q-D plus ampicillin or linezolid resulted in similar enhancement of activity against the VREFc isolate. The results of this study suggest a number of novel antimicrobial combinations that may be useful against staphylococci and enterococci. Combination regimens including cefepime, Q-D, and/or linezolid warrant further investigation for the treatment of refractive infections due to multidrug resistant gram-positive pathogens. PMID- 12121941 TI - Three-dimensional quantitative structure-activity relationship and comparative molecular field analysis of dipeptide hydroxamic acid Helicobacter pylori urease inhibitors. AB - A homology model of Helicobacter pylori urease was developed by using the crystal structure of urease from Klebsiella aerogenes (EC 3.5.1.5) as a template. The acetohydroxamic acid moiety was docked into the active pocket of the enzyme model, followed by relaxation of the complex by use of molecular dynamics. The resulting conformation was used as a template to construct 24 potential dipeptide hydroxamic acid inhibitors with which comparative molecular field analysis (CoMFA) was performed. The resulting model provided a cross-validation correlation coefficient (q(2)(L00)) of 0.610, a conventional r(2) value of 0.988, and an F (Fisher indication of statistical significance) value of 294.88. We were able to validate the CoMFA model by using the 50% inhibitory concentrations of six compounds that were not included in the construction of the model. A very good structural correlation was observed between the amino acids in the model urease's active pocket and the contour maps derived from the CoMFA model. This correlation, accompanied by the validation supplied by use of the CoMFA data, illustrates that the model can aid in the prediction and design of novel H. pylori urease inhibitors. PMID- 12121943 TI - Efficacy of proton pump inhibitor drugs against Plasmodium falciparum in vitro and their probable pharmacophores. AB - The substituted benzimidazoles omeprazole, lansoprazole, rabeprazole, and pantoprazole were found to have in vitro activity against three different isolates of Plasmodium falciparum: D6 (which is chloroquine and pyrimethamine sensitive), W2 (chloroquine and pyrimethamine resistant), and TM91C235 (multidrug resistant). Lansoprazole and rabeprazole were the most effective against all three isolates, with a 50% inhibitory concentration (IC(50)) range of 7 to 11 microM. Omeprazole showed intermediate activity against D6 and W2 isolates, with IC(50)s of 27 to 28 microM, but had poor activity against TM91C235, with an IC(50) of 76 microM. Pantoprazole was the least effective, with IC(50)s of 73 microM against D6, 53 microM against W2, and 39 microM against TM91C235. A pharmacophore model describing the important features responsible for potent activity of the drugs was developed using computational techniques of semiempirical quantum chemical methods and the three-dimensional QSAR procedure of the CATALYST software. The important features of the pharmacophore, according to the findings based on the CATALYST procedures, are the hydrogen bond acceptor and donor sites at the benzimidine nitrogen atoms and the two aromatic hydrophobic sites in the molecules. AM1 quantum chemical calculations identified the electrostatic potential surface surrounding the sulfoxide atom as crucial for potent activity. PMID- 12121942 TI - Short-chain aliphatic polysulfonates inhibit the entry of Plasmodium into red blood cells. AB - Several steps in the pathogenesis of a Plasmodium falciparum infection depend on interactions of parasite surface proteins with negatively charged sugars on the surface of host cells such as sialate residues or glycosaminoglycans. For these reasons, our previous studies examining agents that interfere with heparan sulfate-protein binding during amyloidogenesis suggested that short-chain aliphatic polysulfonates may prove useful as antimalarial agents. A series of related polysulfonates were synthesized and assessed both in tissue culture with the asexual stages of P. falciparum in human red blood cells and in vivo by use of Plasmodium berghei infections in mice. Poly(vinylsulfonate sodium salt) (molecular weight range, 1,500 to 3,000) proved effective in interfering with P. falciparum merozoite entry into human red blood cells and significantly delaying the increase in the level of P. berghei parasitemia in mice. The concept that anionic molecules that mimic large polysaccharide structures may have antimalarial properties has been suggested and examined previously. Our results suggest that related anionic agents [poly(vinylsulfonate sodium salt)-like molecules] orders of magnitude smaller than those previously considered may prove useful in abrogating merozoite entry into erythrocytes and may potentially block sporozoite entry into liver cells. Structure-activity studies conducted to enhance these properties may provide compounds with scope for significant further analysis and development. PMID- 12121944 TI - Transport of an antifungal trypsin inhibitor isolated from corn across the blood brain barrier. AB - We determined whether an antifungal 14-kDa protein trypsin inhibitor isolated from corn is able to cross the blood-brain barrier. We found that it completely crossed the blood-brain barrier by means of a saturable mechanism at a rate of 0.153 microl/g. min, with about 0.082% of the intravenously injected dose being taken up per gram of brain. PMID- 12121945 TI - Salicylate reduces susceptibility of Mycobacterium tuberculosis to multiple antituberculosis drugs. AB - Salicylate induces multiple antibiotic resistance in various bacterial species. Here we investigated the effect of salicylate on the susceptibility of Mycobacterium tuberculosis to a range of antituberculosis (anti-TB) drugs. In the presence of salicylate, the killing effects of isoniazid (INH), rifampin (RMP), ethambutol (EMB), streptomycin (STR), and p-aminosalicylate (PAS) were reduced, as shown with a tetrazolium redox dye viability assay and a bacterial survival assay. Salicylate-induced resistance was more pronounced for PAS, STR, and EMB but was not apparent for INH and RMP when salicylate and the anti-TB agents were incorporated into 7H11 plates. The significance of these findings for TB treatment needs to be further evaluated in vivo. PMID- 12121946 TI - The AcrAB-TolC efflux pump contributes to multidrug resistance in the nosocomial pathogen Enterobacter aerogenes. AB - We identified the genes encoding the AcrA-AcrB-TolC efflux pump in Enterobacter aerogenes and constructed acrAB and tolC mutants from a multidrug-resistant isolate. Both derivatives were more susceptible to antibiotics than the parental strain. Sequence analysis and complementation experiments revealed that the multidrug-resistant isolate is an acrR mutant. PMID- 12121947 TI - gyrA polymorphism in Campylobacter jejuni: detection of gyrA mutations in 162 C. jejuni isolates by single-strand conformation polymorphism and DNA sequencing. AB - Mutations in the quinolone resistance-determining region of the gyrA gene from 138 ciprofloxacin-resistant (MIC, > or =4 microg/ml) and 24 ciprofloxacin susceptible (MIC, < or =1 microg/ml) clinical Campylobacter jejuni isolates were subjected to single-strand conformation polymorphism analysis and sequencing. All of the isolates could be assigned to three genotypic clusters based on silent mutations. All resistant isolates had a point mutation at codon 86. PMID- 12121948 TI - Therapeutic efficacies of GW471552 and GW471558, two new azasordarin derivatives, against pneumocystosis in two immunosuppressed-rat models. AB - Two new azasordarins, GW471552 and GW471558, were studied in vivo for treatment of Pneumocystis carinii pneumonia. In the Wistar rat spontaneous pneumonia model, both azasordarins significantly reduced the number of P. carinii cysts per gram of lung homogenate when administered at 1 mg/kg of body weight twice a day for 10 days. In a nude rat inoculation model, both compounds showed therapeutic efficacy at 0.25 mg/kg twice a day for 10 days. PMID- 12121949 TI - Longitudinal assessment of antipneumococcal susceptibility in the United States. AB - The prevalence of antimicrobial resistance among 4,940 U.S. pneumococcal isolates collected during 1999 was as follows: penicillin, 16.2%; amoxicillin-clavulanate, 12.2%; cefuroxime, 28.1%; ceftriaxone, 3.6%; trimethoprim-sulfamethoxazole, 30.3%; azithromycin, 21.4%; levofloxacin, 0.6%; and moxifloxacin, 0.1%. Compared to the previous 1997-1998 study (Jones et al., Antimicrob. Agents Chemother. 44:2645-2652, 2000), increases were noted for resistance to penicillin (3.7%; P < 0.001), amoxicillin-clavulanate (3.9%; P < 0.001), cefuroxime (5.7%; P < 0.001), azithromycin (2.4%; P = 0.014), trimethoprim-sulfamethoxazole (15.4%; P < 0.001), and levofloxacin (0.3%; P = 0.017). Resistance to ceftriaxone (0.1%; P = 0.809) and moxifloxacin (0.03%; P = 0.570) decreased. Concurrently, multidrug resistance increased (P < 0.001) from 6.3% to 11.3%. PMID- 12121950 TI - Novel complex sul1-type integron in Escherichia coli carrying bla(CTX-M-9). AB - For the present report, a novel complex class 1 integron, In60, was characterized. Part of this integron includes the bla(CTX-M-9) gene and its downstream nucleotide sequence, which shares 81% and 78% nucleotide identity with those of kluA-1 beta-lactamase and orf3 of K. ascorbata, respectively. Furthermore, a new insertion sequence, IS3000, has been found in In60. PCR analysis indicates that integron In60 is present in 33 of 34 nonclonal enterobacterial isolates carrying the putative beta-lactamase CTX-M-9. PMID- 12121952 TI - Activities of six different quinolones against clinical respiratory isolates of Streptococcus pneumoniae with reduced susceptibility to ciprofloxacin in Spain. AB - Six quinolones were tested on 817 consecutive pneumococcal isolates for which ciprofloxacin MICs were high (> or =2 microg/ml); the isolates had been collected during two recent Spanish surveillance studies. For strains for which the ciprofloxacin MIC was >or =4 microg/ml, the MICs at which 90% of the isolates tested against gemifloxacin, moxifloxacin, gatifloxacin, sparfloxacin, levofloxacin, and ofloxacin were inhibited were 0.25, 1, 1, 1, 4 and 16 microg/ml, respectively, and the corresponding prevalences of resistance were 0, 1, 4.5, 9.5, 8.4 and 23%. The proportion of isolates for which the ciprofloxacin MIC is high has increased over time. PMID- 12121951 TI - In vitro evaluation of AZD2563, a novel oxazolidinone, against 603 recent staphylococcal isolates. AB - AZD2563, a novel oxazolidinone, and a selection of comparator drugs that included linezolid, erythromycin, clindamycin, quinolones, and gentamicin were tested against 384 Staphylococcus aureus (176 oxacillin-resistant S. aureus [ORSA]) and 219 coagulase-negative staphylococci (CoNS; 162 oxacillin resistant) by reference microdilution (all strains) and agar dilution (30 strains) methods. The following results were noted for AZD2563. (Note that, for comparison only, a breakpoint of < or =4 microg/ml [the breakpoint of linezolid] was used for this study, although a susceptibility breakpoint for AZD2563 has not been determined.) For S. aureus, the MIC at which 50% of the isolates tested are inhibited (MIC(50)) was 1 microg/ml, the MIC(90) was 2 microg/ml, and the percent susceptibility was 100%. For CoNS, the MIC(50) was 0.5 microg/ml, the MIC(90) was 1 microg/ml, and the percent susceptibility was 100%. ORSA and OR-CoNS strains were equally inhibited by AZD2563 and linezolid. AZD2563 demonstrated antistaphylococcal activity comparable to that of linezolid. PMID- 12121953 TI - Chemiosmotic mechanism of antimicrobial activity of Ag(+) in Vibrio cholerae. AB - Although the antimicrobial effects of silver salts were noticed long ago, the molecular mechanism of the bactericidal action of Ag(+) in low concentrations has not been elucidated. Here, we show that low concentrations of Ag(+) induce a massive proton leakage through the Vibrio cholerae membrane, which results in complete deenergization and, with a high degree of probability, cell death. PMID- 12121954 TI - Incidence, epidemiology, and characteristics of quinolone-nonsusceptible Streptococcus pneumoniae in Croatia. AB - Among 585 Streptococcus pneumoniae strains isolated in 22 Croatian hospitals 21 strains (3.6%) were quinolone nonsusceptible. MICs of all quinolones were high for seven strains tested with the same serotype (23F) and mutations in gyrA, parC, and parE. The remaining 14 strains were more heterogeneous and had mutations only in parC and/or parE, and the MICs of quinolones were lower for these strains. PMID- 12121956 TI - Transmission electron microscopic study of antibiotic action on Klebsiella pneumoniae biofilm. AB - The penetration of ampicillin and ciprofloxacin through biofilms formed by Klebsiella pneumoniae was confirmed by transmission electron microscopic observation of antibiotic-affected cells at the distal edge of the biofilm. Because the bacteria nevertheless survived antibiotic treatment, some protective mechanism other than inadequate penetration must have been at work in the biofilm. PMID- 12121955 TI - Role of active efflux in association with target gene mutations in fluoroquinolone resistance in clinical isolates of Vibrio cholerae. AB - Quinolones are among the drugs of choice in the management of cholera caused by Vibrio cholerae. In this study, we demonstrate that, in addition to mutations detected in the target genes gyrA and parC, proton motive force-dependent efflux is involved in quinolone resistance in clinical isolates of V. cholerae. PMID- 12121957 TI - Rapid detection of a molecular marker for chloroquine-resistant falciparum malaria. AB - A PCR-based technique using molecular beacons was developed to detect the chloroquine resistance-associated pfcrt K76T point mutation in Plasmodium falciparum. One hundred thirty African clinical isolates were tested by the new method in comparison with the PCR-restriction fragment length polymorphism method. This rapid and inexpensive genomic assay could expand the possibilities for monitoring chloroquine resistance. PMID- 12121958 TI - Induction of apoptosis by a nonnucleoside human immunodeficiency virus type 1 reverse transcriptase inhibitor. AB - Inhibition of human immunodeficiency virus type 1 reverse transcriptase (RT) by both nucleoside and nonnucleoside RT inhibitors profoundly inhibits virus replication. Nucleoside RT inhibitors are known to be toxic, but there is little information regarding the toxicities of nonnucleoside RT inhibitors (NNRTI). We demonstrate that efavirenz (an NNRTI) induces caspase- and mitochondrion dependent apoptosis of Jurkat T cells and human peripheral blood mononuclear cells. The clinical relevance of these observations is not yet clear. PMID- 12121959 TI - Two novel vaginal microbicides (polystyrene sulfonate and cellulose sulfate) inhibit Gardnerella vaginalis and anaerobes commonly associated with bacterial vaginosis. AB - This is the first report demonstrating the in vitro inhibitory activity of two novel microbicides (cellulose sulfate and polystyrene sulfonate) against bacterial vaginosis (BV)-associated bacteria. Vaginal application of these microbicides not only may reduce the risk of acquisition of human immunodeficiency virus and other sexually transmitted infection-causing organisms but may also decrease the incidence of BV. PMID- 12121960 TI - Extrusion of penem antibiotics by multicomponent efflux systems MexAB-OprM, MexCD OprJ, and MexXY-OprM of Pseudomonas aeruginosa. AB - The high intrinsic penem resistance of Pseudomonas aeruginosa is due to the interplay among the outer membrane barrier, the active efflux system MexAB-OprM, and AmpC beta-lactamase. We studied the roles of two other efflux systems, MexCD OprJ and MexXY-OprM, in penem resistance by overexpressing each system in an AmpC and MexAB-OprM-deficient background and found that MexAB-OprM is the most important among the three efflux systems for extrusion of penems from the cell interior. PMID- 12121961 TI - Biguanide-atovaquone synergy against Plasmodium falciparum in vitro. AB - The synergistic potential of a range of biguanides, their triazine metabolites, tetracyclines, and pyrimethamine in combination with atovaquone has been assessed. All five biguanides tested interacted synergistically with atovaquone against Plasmodium falciparum in vitro. All of the other compounds tested were either additive or antagonistic. PMID- 12121962 TI - Efficacy of orally delivered cochleates containing amphotericin B in a murine model of aspergillosis. AB - Cochleates containing amphotericin B (CAMB) were administered orally at doses ranging from 0 to 40 mg/kg of body weight/day for 14 days in a murine model of systemic aspergillosis. The administration of oral doses of CAMB (20 and 40 mg/kg/day) resulted in a survival rate of 70% and a reduction in colony counts of more than 2 logs in lungs, livers, and kidneys. Orally administered CAMB shows promise for the treatment of aspergillosis. PMID- 12121963 TI - In vitro susceptibilities of zygomycetes to combinations of antimicrobial agents. AB - Combinations of antimicrobial agents were tested against 35 strains of zygomycetes. The interaction between amphotericin B and rifampin was synergistic or additive. Flucytosine alone was inactive and, upon combination with amphotericin B, synergy was not achieved. The combination of amphotericin B with terbinafine was synergistic for 20% of strains, and the interaction between terbinafine and voriconazole was synergistic for 44% of strains. Antagonism was not observed. PMID- 12121964 TI - Topoisomerase mutations associated with in vitro selection of resistance to moxifloxacin in Streptococcus pneumoniae. AB - We analyzed the frequencies of selection, the order of acquisition, and the mutations selected on moxifloxacin in two wild-type pneumococcal strains, R6 and 5714. The first selection step showed either a single GyrA mutation or no mutation in any of the quinolone resistance-determining regions. Second-step mutants selected had either a second mutation in ParC or in ParE. Moxifloxacin could belong to these fluoroquinolones, which preferentially target GyrA though probably acting equally through both gyrase and topoisomerase IV. PMID- 12121965 TI - Activities of ABT-773 against microaerophilic and fastidious organisms. AB - ABT-773 was tested against 317 fastidious isolates; it inhibited 99% of organisms at a concentration of 4.0 microg/ml. With ampicillin-sulbactam and levofloxacin, only 2 and 6% of these strains, respectively, were resistant at the breakpoint. With clindamycin, penicillin G, and metronidazole, 22, 26, and 58% of the strains, respectively, were resistant. PMID- 12121966 TI - Resazurin microtiter assay plate: simple and inexpensive method for detection of drug resistance in Mycobacterium tuberculosis. AB - A method for detecting multidrug-resistant Mycobacterium tuberculosis by using a reduction of resazurin is described. Eighty clinical isolates were evaluated against isoniazid and rifampin; results at 7 days were compared with those of the proportion method. Specificity and sensitivity were excellent. The method is simple, inexpensive, and rapid and might be used with other antituberculosis drugs. PMID- 12121967 TI - Hematologic effects of linezolid: summary of clinical experience. AB - Linezolid has been associated with reversible myelosuppression. Clinical trial data were evaluated for anemia, thrombocytopenia, and neutropenia. Thrombocytopenia and a slight increased risk for anemia were evident at > or =2 weeks of linezolid treatment. Hematologic abnormalities were consistent with mild, reversible, duration-dependent myelosuppression. Appropriate monitoring is warranted with linezolid use. PMID- 12121968 TI - The presenilin 1 deltaE9 mutation gives enhanced basal phospholipase C activity and a resultant increase in intracellular calcium concentrations. AB - We studied effects of the familial Alzheimer's disease presenilin 1 (PS1) exon 9 deletion (PS1-DeltaE9) mutation on basal and carbachol-stimulated phosphoinositide (PI) hydrolysis and intracellular Ca(2+) concentrations ([Ca(2+)](i)) in human SH-SY5Y neuroblastoma cells. We demonstrate that PS1 DeltaE9 cells have an enhanced basal PI hydrolysis and [Ca(2+)](i) as compared with both wild type PS1 (PS1-WT) and nontransfected (NT) cells. Both were reversed by the phospholipase C (PLC) inhibitor neomycin. The PS1-DeltaE9-related high basal [Ca(2+)](i) was also reversed by xestospongin C confirming that this effect was inositol trisphosphate receptor-mediated. Carbachol gave a greater stimulation of [Ca(2+)](i) in PS1-DeltaE9 cells that took longer to return to basal as compared with responses seen in NT and PS1-WT cells. This long tail-off effect seen in PS1-DeltaE9 cells after carbachol stimulation was reversed by xestospongin C and dantrolene, suggesting that it was mediated by inositol trisphosphate receptor and ryanodine receptor amplification of Ca(2+). Ruthenium red only reduced carbachol peak elevations of [Ca(2+)](i) in NT and PS1-WT cells and not in PS1-DeltaE9 cells. No significant between cell type differences were seen for basal and carbachol-stimulated [Ca(2+)](i) with either ryanodine or the endoplasmic reticulum Ca(2+) ATPase inhibitor cyclopiazonic acid. Immunostaining experiments revealed that for all the cell types PS1 is present at the plasma membrane and co-localizes with N-cadherin, a component of the cell-cell adhesion complex. Immunoblotting of cell extracts for PLC-beta1 showed that, compared with NT and PS1-WT cells, the PS1-DeltaE9 transfectants gave a relative increase in levels of the calpain generated N-terminal fragment (100 kDa) over full-length (150 kDa) PLC-beta1. Our results suggest that the PS1-DeltaE9 mutation causes upstream changes in PI signaling with enhanced basal PLC activity as a primary effect that leads to a higher [Ca(2+)](i). This may provide a novel mechanism by which the PS1-DeltaE9 mutation sensitizes cells to apoptotic stimuli and enhanced amyloid beta generation. PMID- 12121969 TI - Proteasome-mediated degradation of Smac during apoptosis: XIAP promotes Smac ubiquitination in vitro. AB - During apoptosis, Smac (second mitochondria-derived activator of caspases)/DIABLO, an IAP (inhibitor of apoptosis protein)-binding protein, is released from mitochondria and potentiates apoptosis by relieving IAP inhibition of caspases. We demonstrate that exposure of MCF-7 cells to the death-inducing ligand, TRAIL (tumor necrosis factor-related apoptosis-inducing ligand), results in rapid Smac release from mitochondria, which occurs before or in parallel with loss of cytochrome c. Smac release is inhibited by Bcl-2/Bcl-xL or by a pan caspase inhibitor demonstrating that this event is caspase-dependent and modulated by Bcl-2 family members. Following release, Smac is rapidly degraded by the proteasome, an effect suppressed by co-treatment with a proteasome inhibitor. As the RING finger domain of XIAP possesses ubiquitin-protein ligase activity and XIAP binds tightly to mature Smac, an in vitro ubiquitination assay was performed which revealed that XIAP functions as a ubiquitin-protein ligase (E3) in the ubiquitination of Smac. Both the association of XIAP with Smac and the RING finger domain of XIAP are essential for ubiquitination, suggesting that the ubiquitin-protein ligase activity of XIAP may promote the rapid degradation of mitochondrial-released Smac. Thus, in addition to its well characterized role in inhibiting caspase activity, XIAP may also protect cells from inadvertent mitochondrial damage by targeting pro-apoptotic molecules for proteasomal degradation. PMID- 12121970 TI - Spontaneous calcium oscillations control c-fos transcription via the serum response element in neuroendocrine cells. AB - In excitable cells the localization of Ca2+ signals plays a central role in the cellular response, especially in the control of gene transcription. To study the effect of localized Ca2+ signals on the transcriptional activation of the c-fos oncogene, we stably expressed various c-fos beta-lactamase reporter constructs in pituitary AtT20 cells. A significant, but heterogenous expression of c-fos beta lactamase was observed in unstimulated cells, and a further increase was observed using KCl depolarization, epidermal growth factor (EGF), pituitary adenylate cyclase-activating polypeptide (PACAP), and serum. The KCl response was almost abolished by a nuclear Ca2+ clamp, indicating that a rise in nuclear Ca2+ is required. In contrast, the basal expression was not affected by the nuclear Ca2+ clamp, but it was strongly reduced by nifedipine, a specific antagonist of l-type Ca2+ channels. Spontaneous Ca2+ oscillations, blocked by nifedipine, were observed in the cytosol but did not propagate to the nucleus, suggesting that a rise in cytosolic Ca2+ is sufficient for basal c-fos expression. Inactivation of the c-fos promoter cAMP/Ca2+ response element (CRE) had no effect on basal or stimulated expression, whereas inactivation of the serum response element (SRE) had the same marked inhibitory effect as nifedipine. These experiments suggest that in AtT20 cells spontaneous Ca2+ oscillations maintain a basal c-fos transcription through the serum response element. Further induction of c-fos expression by depolarization requires a nuclear Ca2+ increase. PMID- 12121971 TI - COM crystals activate the p38 mitogen-activated protein kinase signal transduction pathway in renal epithelial cells. AB - Interaction of calcium oxalate monohydrate (COM) crystals with renal cells has been shown to result in altered gene expression, DNA synthesis, and cell death. In the current study the role of a stress-specific p38 MAP kinase-signaling pathway in mediating these effects of COM crystals was investigated. Exposure of cells to COM crystals (20 microg/cm(2)) rapidly stimulated strong phosphorylation and activation of p38 mitogen-activated protein kinase (p38 MAP kinase) and re initiation of DNA synthesis. Inhibition of COM crystal binding to the cells by heparin blocked the effects of COM crystals on p38 MAPK activation. We also show that specific inhibition of p38 MAPK by 4-(4-fluorophenyl)-2-(4 methylsulfonylphenyl)-5-(4-pyridyl) imidazole (SB203580) or by overexpression of a dominant negative mutant of p38 MAP kinase abolishes COM crystal-induced re initiation of DNA synthesis. The inhibition is dose-dependent and correlates with in situ activity of native p38 MAP kinase, determined as mitogen-activated protein kinase-activated protein kinase-2 (MAPKAP kinase-2) activity in cell extracts. In summary, inhibiting activation of p38 MAPK pathway abrogated the DNA synthesis in response to COM crystals. These data are the first demonstrations of activation of the p38 MAPK signaling pathway by COM crystals and suggest that, in response to COM crystals, this pathway transduces critical signals governing the re-initiation of DNA synthesis in renal epithelial cells. PMID- 12121972 TI - Analysis of the life cycle of stat6. Continuous cycling of STAT6 is required for IL-4 signaling. AB - Signal transducer and activator of transcription (Stat)6 is a transcription factor important for the development of Th2 cells and regulation of gene expression by IL-4 and IL-13. It is known that Stat6 is rapidly activated in response to IL-4; however, the fate of activated Stat6 is less clear. We examined the fate of activated Stat6 and found that during continuous exposure to IL-4, Stat6 activity was sustained for 72 h and that the maintenance of a constant level of activated Stat6 did not require new protein synthesis. In contrast, when cells were pulsed with IL-4 and then incubated in the absence of IL-4, the half life of Stat6 phosphorylation and DNA binding activity was less than 1 h. Stat6 did not accumulate in the nucleus, and protein degradation did not play a major role in the disappearance of activated Stat6. Inhibition of kinase activity by staurosporine or the JAK inhibitor, AG490, revealed that maintenance of Stat6 activation in the continuous presence of IL-4 required ongoing phosphorylation of latent cytoplasmic Stat6 molecules. Cells treated with an inhibitor of nuclear export, leptomycin B, were unable to maintain Stat6 activation. Thus, the maintenance of Stat6 activation requires a constant cycle of activation, deactivation, nuclear export, and reactivation. PMID- 12121973 TI - Protein kinase C-epsilon promotes survival of lung cancer cells by suppressing apoptosis through dysregulation of the mitochondrial caspase pathway. AB - The serine/threonine protein kinase C (PKC) has been implicated in the regulation of drug resistance and cell survival in many types of cancer cells. However, the one or more precise mechanisms remain elusive. In this study, we have identified and determined the mechanism by which PKC-epsilon, a novel PKC isoform, modulates drug resistance in lung cancer cells. Western blot analysis demonstrates that expression of PKC-epsilon, but not other PKC isoforms, is associated with the chemo-resistant phenotype of non-small cell lung cancer (NSCLC) cell lines. Northern blotting and nuclear run-on transcription analysis further reveals that the failure of expression of PKC-epsilon in the chemo-sensitive phenotype of small cell lung cancer (SCLC) cells results from transcriptional inactivation of the gene. Importantly, forced expression of PKC-epsilon in NCI-H82 human SCLC cells confers a significant resistance to the chemotherapeutic drugs, etoposide and doxorubicin. Resistance is characterized by a significant reduction in apoptosis in PKC-epsilon-expressing cells. Treatment of NCI-H82 cells with etoposide induces a series of time-dependent events, including the release of cytochrome c from the mitochondria to the cytosol, activation of caspase-9 and caspase-3, and cleavage of poly(ADP-ribose) polymerase (PARP). All of these events are blocked by PKC-epsilon expression. Furthermore, caspase-specific inhibitors, z-VAD-fmk and z-DEVD-fmk, significantly attenuate the accumulation of sub-G(1) population and block the PARP cleavage in response to etoposide. These results suggest that PKC-epsilon prevents cells from undergoing apoptosis through inhibition of the mitochondrial-dependent caspase activation, thereby leading to cell survival. Finally, down-regulation of PKC-epsilon expression by the antisense cDNA in NSCLC cells results in increased sensitivity to etoposide. Taken together, our findings suggest an important role for PKC-epsilon in regulating survival of lung cancer cells. PMID- 12121974 TI - Gamma-synuclein promotes cancer cell survival and inhibits stress- and chemotherapy drug-induced apoptosis by modulating MAPK pathways. AB - Synucleins are a family of highly conserved small proteins predominantly expressed in neurons. Recently we and others have found that gamma-synuclein is dramatically up-regulated in the vast majority of late-stage breast and ovarian cancers and that gamma-synuclein over-expression can enhance tumorigenicity. In the current study, we have found that gamma-synuclein is associated with two major mitogen-activated kinases (MAPKs), i.e. extracellular signal-regulated protein kinases (ERK1/2) and c-Jun N-terminal kinase 1 (JNK1), and have shown that over-expression of gamma-synuclein leads to constitutive activation of ERK1/2 and down-regulation of JNK1 in response to a host of environmental stress signals, including UV, arsenate, and heat shock. We also tested the effects of gamma-synuclein on apoptosis and activation of JNK and ERK in response to several chemotherapy drugs. We have found that gamma-synuclein-expressing cells are significantly more resistant to the chemotherapeutic drugs paclitaxel and vinblastine as compared with the parental cells. The resistance to paclitaxel can be partially obliterated when ERK activity is inhibited using a MEK1/2 inhibitor. Activation of JNK and its downstream caspase-3 by paclitaxel or vinblastine is significantly down-regulated in gamma-synuclein-expressing cells, indicating that the paclitaxel- or vinblastine-activated apoptosis pathway is blocked by gamma synuclein. In contrast to paclitaxel and vinblastine, etoposide does not activate JNK, and gamma-synuclein over-expression has no apparent effect on this drug induced apoptosis. Taken together, our data indicate that oncogenic activation of gamma-synuclein contributes to the development of breast and ovarian cancer by promoting tumor cell survival under adverse conditions and by providing resistance to certain chemotherapeutic drugs. PMID- 12121975 TI - On the role of exon and intron sequences in trans-splicing utilization and cap 4 modification of the trypanosomatid Leptomonas collosoma SL RNA. AB - In trypanosomatid protozoa the biogenesis of mature mRNA involves addition of the spliced leader (SL) sequence from the SL RNA to polycistronic pre-mRNA via trans splicing. Here we present a mutational analysis of the trypanosomatid Leptomonas collosoma SL RNA to further our understanding of its functional domains important for trans-splicing utilization. Mutant SL RNAs were analyzed for defects in modification of the hypermethylated cap structure (cap 4) characteristic of trypanosomatid SL RNAs, for defects in the first step of the reaction and overall utilization in trans-splicing. Single substitution of the cap 4 nucleotides led to undermethylation of the cap 4 structure, and these mutants were all impaired in their utilization in trans-splicing. Abrogation of the sequence of the Sm-like site and sequences downstream to it also showed cap modification and trans splicing defects, thus providing further support for a functional linkage between cap modifications and trans-splicing. Further, we report that in L. collosoma both the exon and intron of the SL RNA contribute information for efficient function of the SL RNA in trans-splicing. This study, however, did not provide support for the putative SL RNA-U6 small nuclear RNA (snRNA) interaction at the Sm site like in the nematodes, suggesting differences in the bridging role of U6 in the two trans-splicing systems. PMID- 12121976 TI - Molecular characterization of the human La protein.hepatitis B virus RNA.B interaction in vitro. AB - The La protein was recently identified as a host factor potentially involved in the cytokine-induced post-transcriptional down-regulation of hepatitis B virus (HBV) RNA. The La binding site was mapped to a predicted stem-loop structure within a region shared by all HBV RNAs, and it was concluded that the La protein might be an HBV RNA-stabilizing factor. To characterize the RNA binding mediated by the different RNA recognition motifs (RRMs) of the human La protein, several La deletion mutants were produced and analyzed for HBV RNA binding ability. The data demonstrate that the first RRM is not required for binding, whereas the RNP 1 and RNP-2 consensus sequences of the RRM-2 and RRM-3 are separately required for binding, indicating a cooperative function of these two RRMs. Furthermore, the results suggest that multimeric La disassembles into monomeric La upon binding of HBV RNA.B. By gel retardation assay the affinity of the wild type human La.HBV RNA.B interaction was determined in the nanomolar range, comparable to the affinity determined for the mouse La.HBV RNA.B interaction. This study identified small regions within the human La protein mediating the binding of HBV RNA. Hence, these binding sites might represent targets for novel antiviral strategies based on the disruption of the human La.HBV RNA interaction, thereby leading to HBV RNA degradation. PMID- 12121977 TI - Cell cycle promoting activity of JunB through cyclin A activation. AB - JunB, a major component of the AP-1 transcription factor, is known to act antagonistically to c-Jun in transcriptional regulation and is proposed to be a negative regulator of cell proliferation. Employing fibroblasts derived from E9.5 junB(-/-) mouse embryos we provide evidence for a novel cell cycle promoting role of JunB. Despite a normal proliferation rate, primary and immortalized junB(-/-) fibroblasts exhibited an altered cell cycle profile, which was characterized by an increase in the population of S-phase cells, while that of cells in G(2)/M phase was diminished. This delay in G(2)/M-transition is caused by impaired cyclin A-CDK2 and cyclin B-CDC2 kinase activities and counteracts the accelerated S-phase entry. Cells lacking JunB show severely delayed kinetics of cyclin A mRNA expression due to the loss of proper transcriptional activation mediated via binding of JunB to the CRE element in the cyclin A promoter. Upon reintroduction of an inducible JunB-ER(TM) expression vector the cell cycle distribution and the cell cycle-associated cyclin A-CDK2 kinase activity could be restored. Thus, cyclin A is a direct transcriptional target of JunB driving cell proliferation. PMID- 12121979 TI - Activation of Go-coupled dopamine D2 receptors inhibits ERK1/ERK2 in pituitary cells. A key step in the transcriptional suppression of the prolactin gene. AB - In pituitary lactotrophs the prolactin gene is stimulated by neuropeptides and estrogen and is suppressed by dopamine via D2-type receptors. Stimulatory signals converge on activation of the mitogen-activated protein kinases ERK1/2, but dopamine regulation of this pathway is not well defined. Paradoxically, D2 agonists activate ERK1/2 in many cell types. Here we show that in prolactin secreting GH4ZR7 cells and primary pituitary cells, dopamine treatment leads to a rapid, pronounced, and specific decrease in activated ERK1/2. The response is blocked by D2-specific antagonists and pertussis toxin. Interestingly, in stable lines expressing specific pertussis toxin-resistant Galpha subunits, toxin treatment blocks dopamine suppression of MAPK in Galpha(i2)- but not Galphao expressing cells, demonstrating that G(o)-dependent pathways can effect the inhibitory MAPK response. At the nuclear level, the MEK1 inhibitor U0126 mimics the D2-agonist bromocryptine in suppressing levels of endogenous prolactin transcripts. Moreover, a good correlation is seen between the IC(50) values for inhibition of MEK1 and suppression of prolactin promoter function (PD184352 > U0126 > U0125). Both dopamine and U0126 enhance the nuclear localization of ERF, a MAPK-sensitive ETS repressor that inhibits prolactin promoter activity. In addition, U0126 suppression is transferred by tandem copies of the Pit-1-binding site, consistent with mapping experiments for dopamine responsiveness. Our data suggest that ERK1/2 suppression is an obligatory step in the dopaminergic control of prolactin gene transcription and that bidirectional control of ERK1/2 function in the pituitary may provide a key mechanism for endocrine gene control. PMID- 12121978 TI - A 29-kDa protein associated with p67phox expresses both peroxiredoxin and phospholipase A2 activity and enhances superoxide anion production by a cell-free system of NADPH oxidase activity. AB - Production of toxic oxygen metabolites provides a mechanism for microbicidal activity of the neutrophil. The NADPH oxidase enzyme system initiates the production of oxygen metabolites by reducing oxygen to form superoxide anion (O(2)()). With stimulation of the respiratory burst, cytosolic oxidase components, p47(phox), p67(phox), and Rac, translocate to the phagolysomal and plasma membranes where they form a complex with cytochrome b(558) and express enzyme activity. A 29-kDa neutrophil protein (p29) was identified by co immunoprecipitation with p67(phox). N-terminal sequence analysis of p29 revealed homology to an open reading frame gene described in a myeloid leukemia cell line. A cDNA for p29 identical to the open reading frame protein was amplified from RNA of neutrophils. Significant interaction between p29 and p67(phox) was demonstrated using a yeast two-hybrid system. A recombinant (rh) p29 was expressed in Sf9 cells resulting in a protein with an apparent molecular weight of 34,000. The rh-p29 showed immunoreactivity with the original rabbit antiserum that detected p47(phox) and p67(phox). In addition, rh-p29 exhibited PLA(2) activity, which was Ca(2+) independent, optimal at low pH, and preferential for phosphatidylcholine substrates. The recombinant protein protected glutathione synthetase and directly inactivated H(2)O(2). By activity and sequence homology, rh-p29 can be classified as a peroxiredoxin. Finally, O(2)() production by plasma membrane and recombinant cytosolic oxidase components in the SDS-activated, cell free NADPH oxidase system were enhanced by rh-p29. This effect was not inhibited by PLA(2) inhibitors. Thus, p29 is a novel protein that associates with p67 and has peroxiredoxin activity. This protein has a potential role in protecting the NADPH oxidase by inactivating H(2)O(2) or altering signaling pathways affected by H(2)O(2). PMID- 12121980 TI - Inhibition of the inositol trisphosphate receptor of mouse eggs and A7r5 cells by KN-93 via a mechanism unrelated to Ca2+/calmodulin-dependent protein kinase II antagonism. AB - KN-93, a Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) inhibitor, concentration-dependently and reversibly inhibited inositol 1,4,5-trisphosphate receptor (IP(3)R)-mediated [Ca(2+)](i) signaling in mouse eggs and permeabilized A7r5 smooth muscle cells, two cell types predominantly expressing type-1 IP(3)R (IP(3)R-1). KN-92, an inactive analog, was ineffective. The inhibitory action of KN-93 on Ca(2+) signaling depended neither on effects on IP(3) metabolism nor on the filling grade of Ca(2+) stores, suggesting a direct action on the IP(3)R. Inhibition was independent of CaMKII, since in identical conditions other CaMKII inhibitors (KN-62, peptide 281-309, and autocamtide-related inhibitory peptide) were ineffective and since CaMKII activation was precluded in permeabilized cells. Moreover, KN-93 was most effective in the absence of Ca(2+). Analysis of Ca(2+) release in A7r5 cells at varying [IP(3)], of IP(3)R-1 degradation in eggs, and of [(3)H]IP(3) binding in Sf9 microsomes all indicated that KN-93 did not affect IP(3) binding. Comparison of the inhibition of Ca(2+) release and of [(3)H]IP(3) binding by KN-93 and calmodulin (CaM), either separately or combined, was compatible with a specific interaction of KN-93 with a CaM-binding site on IP(3)R-1. This was also consistent with the much smaller effect of KN-93 in permeabilized 16HBE14o(-) cells that predominantly express type 3 IP(3)R, which lacks the high affinity CaM-binding site. These findings indicate that KN-93 inhibits IP(3)R-1 directly and may therefore be a useful tool in the study of IP(3)R functional regulation. PMID- 12121981 TI - Interaction between the N-terminal and middle regions is essential for the in vivo function of HSP90 molecular chaperone. AB - At the primary structure level, the 90-kDa heat shock protein (HSP90) is composed of three regions: the N-terminal (Met(1)-Arg(400)), middle (Glu(401)-Lys(615)), and C-terminal (Asp(621)-Asp(732)) regions. In the present study, we investigated potential subregion structures of these three regions and their roles. Limited proteolysis revealed that the N-terminal region could be split into two fragments carrying residues Met(1) to Lys(281) (or Lys(283)) and Glu(282) (or Tyr(284)) to Arg(400). The former is known to carry the ATP-binding domain. The fragments carrying the N-terminal two-thirds (Glu(401)-Lys(546)) and C-terminal one-third of the middle region were sufficient for the interactions with the N- and C terminal regions, respectively. Yeast HSC82 that carried point mutations in the middle region causing deficient binding to the N-terminal region could not support the growth of HSP82-depleted cells at an elevated temperature. Taken together, our data show that the N-terminal and middle regions of the HSP90 family protein are structurally divided into two respective subregions. Moreover, the interaction between the N-terminal and middle regions is essential for the in vivo function of HSP90 in yeast. PMID- 12121982 TI - Staring, a novel E3 ubiquitin-protein ligase that targets syntaxin 1 for degradation. AB - Syntaxin 1 is an essential component of the neurotransmitter release machinery, and regulation of syntaxin 1 expression levels is thought to contribute to the mechanism underlying learning and memory. However, the molecular events that control the degradation of syntaxin 1 remain undefined. Here we report the identification and characterization of a novel RING finger protein, Staring, that interacts with syntaxin 1. Staring is expressed throughout the brain, where it exists in both cytosolic and membrane-associated pools. Staring binds and recruits the brain-enriched E2 ubiquitin-conjugating enzyme UbcH8 to syntaxin 1 and facilitates the ubiquitination and proteasome-dependent degradation of syntaxin 1. These findings suggest that Staring is a novel E3 ubiquitin-protein ligase that targets syntaxin 1 for degradation by the ubiquitin-proteasome pathway. PMID- 12121983 TI - Calpain-mediated X-linked inhibitor of apoptosis degradation in neutrophil apoptosis and its impairment in chronic neutrophilic leukemia. AB - The number of neutrophils in the blood and tissues is controlled by constitutive apoptotic programmed cell death and clearance by phagocytes such as macrophages. Here, we found that calpains cleave the X-linked inhibitor of apoptosis (XIAP) in vitro, producing fragments that are unable to inhibit caspase-3. These fragments were detected in normal neutrophils but were unstable and rapidly degraded. Calpain inhibition delayed tumor necrosis factor-alpha-induced apoptosis of normal neutrophils, consistent with a role for calpains in regulating the onset of apoptosis. Interestingly, neutrophils from three patients with chronic neutrophilic leukemia, a rare syndrome characterized by accumulation of mature neutrophils, exhibited decreased mu-calpain expression, diminished calpain activity, and impaired XIAP degradation. Neutrophils from these patients displayed a delay in spontaneous, Fas-stimulated, and tumor necrosis factor-alpha induced apoptosis. These observations suggest that calpain-mediated XIAP degradation contributes to initiation of apoptosis in normal neutrophils and dysfunction of this regulatory pathway can lead to pathological neutrophil accumulation. PMID- 12121984 TI - Association of Helicobacter pylori vacuolating toxin (VacA) with lipid rafts. AB - A variety of extracellular ligands and pathogens interact with raft domains in the plasma membrane of eukaryotic cells. In this study, we examined the role of lipid rafts and raft-associated glycosylphosphatidylinositol (GPI)-anchored proteins in the process by which Helicobacter pylori vacuolating toxin (VacA) intoxicates cells. We first investigated whether GPI-anchored proteins are required for VacA toxicity by analyzing wild-type Chinese hamster ovary (CHO) cells and CHO-LA1 mutant cells that are defective in production of GPI-anchored proteins. Whereas wild-type and mutant cells differed markedly in susceptibility to aerolysin (a bacterial toxin that binds to GPI-anchored proteins), they were equally susceptible to VacA. We next determined whether VacA physically associates with lipid rafts. CHO or HeLa cells were incubated with VacA, and Triton-insoluble membranes then were separated by sucrose density gradient centrifugation. Immunoblot analysis revealed that a substantial proportion of cell-associated toxin was associated with detergent-resistant membranes (DRMs). DRM association required acid activation of the purified toxin prior to contact with cells, and acid activation also was required for VacA cytotoxicity. Treatment of cells with methyl-beta-cyclodextrin (a cholesterol-depleting agent) did not inhibit VacA-induced depolarization of the plasma membrane, but interfered with the internalization or intracellular localization of VacA and inhibited the capacity of the toxin to induce cell vacuolation. Treatment of cells with nystatin also inhibited VacA-induced cell vacuolation. These data indicate that VacA associates with lipid raft microdomains in the absence of GPI anchored proteins and suggest that association of the toxin with lipid rafts is important for VacA cytotoxicity. PMID- 12121985 TI - Separable gating mechanisms in a Mammalian pacemaker channel. AB - Despite permeability to both K(+) and Na(+), hyperpolarization-activated cyclic nucleotide-gated (HCN) pacemaker channels contain the K(+) channel signature sequence, GYG, within the selectivity filter of the pore. Here, we show that this region is involved in regulating gating in a mouse isoform of the pacemaker channel (mHCN2). A mutation in the GYG sequence of the selectivity filter (G404S) had different effects on the two components of the wild-type current; it eliminated the slowly activating current (I(f)) but, surprisingly, did not affect the instantaneous current (I(inst)). Confocal imaging and immunocytochemistry showed G404S protein on the periphery of the cells, consistent with the presence of channels on the plasma membrane. Experiments with the wild-type channel showed that the rate of I(f) deactivation and I(f) amplitude had a parallel dependence on the ratio of K(+)/Na(+) driving forces. In addition, the amplitude of fully activated I(f), unlike I(inst), was not well predicted by equal and independent flow of K(+) and Na(+). The data are consistent with two separable gating mechanisms associated with pacemaker channels: one (I(f)) that is sensitive to voltage, to a mutation in the selectivity filter, and to driving forces for permeating cations and another (I(inst)) that is insensitive to these influences. PMID- 12121986 TI - Analysis of the L116K variant of CooA, the heme-containing CO sensor, suggests the presence of an unusual heme ligand resulting in novel activity. AB - CooA is the CO-sensing transcriptional activator from Rhodospirillum rubrum, in which CO binding to its heme prosthetic group triggers a conformational change of CooA that allows the protein to bind its cognate target DNA sequence. By a powerful in vivo screening method following the simultaneous randomization of the codons for two C-helix residues, 113 and 116, near the distal heme pocket of CooA, we have isolated a series of novel CooA variants. In vivo, these show very high CO-independent activities (comparable with that of wild-type CooA in the presence of CO) and diminished CO-dependent activities. Sequence analysis showed that this group of variants commonly contains lysine at position 116 with a variety of residues at position 113. DNA-binding analysis of a representative purified variant, L116K CooA, revealed that this protein is competent to bind target DNA with K(d) values of 56 nm for Fe(III), 36 nm for Fe(II), and 121 nm for Fe(II)-CO CooA forms. Electron paramagnetic resonance and electronic absorption spectroscopies, combined with additional mutagenic studies, showed that L116K CooA has a new ligand replacing Pro(2) in both Fe(III) and Fe(II) states. The most plausible replacement ligand is the substituted lysine at position 116, so that the ligands of Fe(III) L116K CooA are Cys(75) and Lys(116) and those in the Fe(II) form are His(77) and Lys(116). A possible explanation for CO-independent activity in L116K CooA is that ligation of Lys(116) results in a repositioning of the C-helices at the CooA dimer interface. This result is consistent with that repositioning being an important aspect of the activation of wild-type CooA by CO. PMID- 12121987 TI - Proteomic analysis reveals alterations in the renal kallikrein pathway during hypoxia-induced hypertension. AB - Obstructive sleep apnea syndrome (OSAS), a disorder characterized by episodic hypoxia (EH) during sleep, is associated with systemic hypertension. We used proteomic analysis to examine differences in rat kidney protein expression during EH, and their potential relationship to EH-induced hypertension. Young male Sprague-Dawley rats were exposed to either EH or sustained hypoxia (SH) for 14 (EH14/SH14) and 30 (EH30/SH30) days. Mean arterial blood pressure was significantly increased only in EH30 (p < 0.0002). Kidney proteins were resolved by two-dimensional-PAGE and were identified by MALDI-MS. Renal expression of kallistatin, a potent vasodilator, was down-regulated in all animals. Expression of alpha-1-antitrypsin, an inhibitor of kallikrein activation, was up-regulated in EH but down-regulated in SH. Western blotting showed significant elevation of B(2)-bradykinin receptor expression in all normotensive animals but remained unchanged in hypertensive animals. Proteins relevant to vascular hypertrophy, such as smooth muscle myosin and protein-disulfide isomerase were up-regulated in EH30 but were down-regulated in SH30. These data indicate that EH induces changes in renal protein expression consistent with impairment of vasodilation mediated by the kallikrein-kallistatin pathway and vascular hypertrophy. In contrast, SH induced changes suggest the kallikrein- and bradykinin-mediated compensatory mechanisms for prevention of hypertension and vascular remodeling. To test the hypothesis suggested by the proteomic data, we measured the effect of EH on blood pressure in transgenic hKLK1 rats that overexpress human kallikrein. Transgenic hKLK1 animals were protected from EH-induced hypertension. We conclude that EH induced hypertension may result, at least in part, from altered regulation of the renal kallikrein system. PMID- 12121988 TI - Regulation of the nonreceptor tyrosine kinase Brk by autophosphorylation and by autoinhibition. AB - Brk (breast tumor kinase) is a nonreceptor tyrosine kinase that is most closely related to the Frk family of kinases, and more distantly to Src family kinases. Brk was originally identified in a screen for tyrosine kinases that are overexpressed in human metastatic breast tumors. To shed light on the activity and regulation of Brk and related tyrosine kinases, we expressed and purified Brk using the Sf9/baculovirus system. We characterized the substrate specificity of Brk using synthetic peptides, and we show that the kinetic parameters K(m) and k(cat) both play a role in specificity. We carried out mass spectrometry experiments to show that Brk autophosphorylates within the predicted kinase activation loop and at additional sites in the N terminus. Autophosphorylation increases enzyme activity of wild-type Brk but not of a Y342A mutant form of Brk. We also carried out experiments to address the possible involvement of the Src homology (SH) 2 and SH3 domains of Brk in enzyme regulation. Mutation of a C terminal tyrosine (Tyr-447) increases enzyme activity and SH2 domain accessibility, consistent with a role for this residue in autoinhibition. A proline-rich peptide activates Brk, suggesting that the SH3 domain is also involved in maintaining an inactive form of Brk. These biochemical results for Brk may aid in the understanding of other tyrosine kinases in the Frk family. PMID- 12121989 TI - Tyrphostins protect neuronal cells from oxidative stress. AB - Tyrphostins are a family of tyrosine kinase inhibitors originally synthesized as potential anticarcinogenic compounds. Because tyrphostins have chemical structures similar to those of the phenolic antioxidants, we decided to test the protective efficacy of tyrphostins against oxidative stress-induced nerve cell death (oxytosis). Many commercially available tyrphostins, at concentrations ranging from 0.5 to 200 microm, protect both HT-22 hippocampal cells and rat primary neurons from oxytosis brought about by treatment with glutamate, as well as by treatment with homocysteic acid and buthionine sulfoximine. The tyrphostins protect nerve cells by three distinct mechanisms. Some tyrphostins, such as A25, act as antioxidants and eliminate the reactive oxygen species that accumulate as a result of glutamate treatment. These tyrphostins also protect cells from hydrogen peroxide and act as antioxidants in an in vitro assay. In contrast, tyrphostins A9 and AG126 act as mitochondrial uncouplers, collapsing the mitochondrial membrane potential and thereby reducing the generation of reactive oxygen species from mitochondria during glutamate toxicity. Finally, the third group of tyrphostins does not appear to be effective as antioxidants but rather protects cells by increasing the basal level of cellular glutathione. Therefore, the effects of tyrphostins on cells are not limited to their ability to inhibit tyrosine kinases. PMID- 12121991 TI - Reproducibility of oligonucleotide microarray transcriptome analyses. An interlaboratory comparison using chemostat cultures of Saccharomyces cerevisiae. AB - Assessment of reproducibility of DNA-microarray analysis from published data sets is complicated by the use of different microbial strains, cultivation techniques, and analytical procedures. Because intra- and interlaboratory reproducibility is highly relevant for application of DNA-microarray analysis in functional genomics and metabolic engineering, we designed a set of experiments to specifically address this issue. Saccharomyces cerevisiae CEN.PK113-7D was grown under defined conditions in glucose-limited chemostats, followed by transcriptome analysis with Affymetrix GeneChip arrays. In each of the laboratories, three independent replicate cultures were grown aerobically as well as anaerobically. Although variations introduced by in vitro handling steps were small and unbiased, greater variation from replicate cultures underscored that, to obtain reliable information, experimental replication is essential. Under aerobic conditions, 86% of the most highly expressed yeast genes showed an average intralaboratory coefficient of variation of 0.23. This is significantly lower than previously reported for shake-flask-culture transcriptome analyses and probably reflects the strict control of growth conditions in chemostats. Using the triplicate data sets and appropriate statistical analysis, the change calls from anaerobic versus aerobic comparisons yielded an over 95% agreement between the laboratories for transcripts that changed by over 2-fold, leaving only a small fraction of genes that exhibited laboratory bias. PMID- 12121990 TI - A single enzyme catalyses formation of Trypanothione from glutathione and spermidine in Trypanosoma cruzi. AB - Protozoa of the order Kinetoplastida differ from other organisms in their ability to conjugate glutathione (l-gamma-glutamyl-cysteinyl-glycine) and spermidine to form trypanothione [N(1),N(8)-bis(glutathionyl)spermidine], a metabolite involved in defense against chemical and oxidant stress and other biosynthetic functions. In Crithidia fasciculata, trypanothione is synthesized from GSH and spermidine via the intermediate glutathionylspermidine in two distinct ATP-dependent reactions catalyzed by glutathionylspermidine synthetase (GspS; EC ) and trypanothione synthetase (TryS; EC ), respectively. Here we have cloned a single copy gene (TcTryS) from Trypanosoma cruzi encoding a protein with 61% sequence identity with CfTryS but only 31% with CfGspS. Saccharomyces cerevisiae transformed with TcTryS were able to synthesize glutathionylspermidine and trypanothione, suggesting that this enzyme is able to catalyze both biosynthetic steps, unlike CfTryS. When cultures were supplemented with aminopropylcadaverine, yeast transformants contained glutathionylaminopropylcadaverine and homotrypanothione [N(1),N(9)-bis(glutathionyl)aminopropylcadaverine], metabolites that have been previously identified in T. cruzi, but not in C. fasciculata. Kinetic studies on recombinant TcTryS purified from Escherichia coli revealed that the enzyme displays high-substrate inhibition with glutathione (K(m) and K(i) of 0.57 and 1.2 mm, respectively, and k(cat) of 3.4 s(-1)), but obeys Michaelis-Menten kinetics with spermidine, aminopropylcadaverine, glutathionylspermidine, and MgATP as variable substrate. The recombinant enzyme possesses weak amidase activity and can hydrolyze trypanothione, homotrypanothione, or glutathionylspermidine to glutathione and the corresponding polyamine. PMID- 12121992 TI - SNAP-23 is a target for calpain cleavage in activated platelets. AB - The role of calpain in platelet function is generally associated with aggregation and clot retraction. In this report, data are presented to show that one component of the platelet secretory machinery, SNAP-23, is specifically cleaved by calpain in activated cells. Other proteins of the membrane fusion machinery, e.g. syntaxins 2 and 4 and alpha-SNAP, are not affected. In vitro studies, using permeabilized platelets, demonstrate that cleavage is time- and calcium dependent. Analysis of SNAP-23 cleavage products suggests that the calpain cleavage site(s) is in the C-terminal third of the molecule potentially between the cysteine-rich acyl attachment sites and the C-terminal coiled-coil domain. The time course of cleavage is most consistent with late calpain-mediated events such as pp60(c-src) cleavage, but not early events such as protein-tyrosine phosphatase-1B activation. SNAP-23 cleavage is inhibited by calpeptin, calpastatin, calpain inhibitor IV, and E-64d, but not by caspase 3 inhibitor III or cathepsin inhibitor I. When tested for their effect on secretion, none of the calpain-specific inhibitors significantly affected release of soluble components from any of the three platelet granule storage pools. These results indicate that SNAP-23 cleavage occurs after granule release and therefore may play a role in affecting granule membrane exteriorization. This is consistent with the ultrastructural morphology of calpeptin-treated platelets after activation. PMID- 12121993 TI - Identification of a short highly conserved amino acid sequence as the functional region required for posttranscriptional autoregulation of the cystathionine gamma synthase gene in Arabidopsis. AB - Cystathionine gamma-synthase (CGS) catalyzes the first committed step of Met biosynthesis in plants. We have previously shown that expression of the gene for CGS is feedback-regulated at the level of mRNA stability, and that the amino acid sequence encoded by the first exon of the CGS gene itself is responsible for the regulation (Chiba, Y., Ishikawa, M., Kijima, F., Tyson, R. H., Kim, J., Yamamoto, A., Nambara, E., Leustek, T., Wallsgrove, R. M., and Naito, S. (1999) Science 286, 1371-1374). To identify the functional region within CGS exon 1, deletion analysis was performed. The results showed that the 41-amino acid region of exon 1 highly conserved among plants is necessary and sufficient for the regulation. Analyses of in vivo and in vitro generated mutations that abolish the regulation identified the functionally important amino acids as 11-13 residues within this conserved region. The importance of these residues was confirmed by deletion analysis within the conserved region. These studies identified the functional region of CGS exon 1 required for the posttranscriptional autoregulation of the CGS gene as (A)RRNCSNIGVAQ(I), with uncertainty of the first and last residues. This sequence is almost perfectly conserved among CGS sequences of higher plants but cannot be found elsewhere in the public databases. PMID- 12121994 TI - A novel calcium-stimulated adenylyl cyclase from Trypanosoma cruzi, which interacts with the structural flagellar protein paraflagellar rod. AB - Trypanosoma cruzi adenylyl cyclases are encoded by a large polymorphic gene family. Although several genes have been identified in this parasite, little is known about the properties and regulation of these enzymes. Here we report the cloning and characterization of TczAC, a novel member of T. cruzi adenylyl cyclase family. The TczAC gene is expressed in all of the parasite life forms and encodes a 1,313-amino acid protein that can complement a Saccharomyces cerevisiae mutant deficient in adenylyl cyclase activity. The recombinant enzyme expressed in yeasts is constitutively active, has a low affinity for ATP (K(m) = 406 microm), and requires a divalent cation for catalysis. TczAC is inhibited by Zn(2+) and the P-site inhibitor 2'-deoxyadenosine 3'-monophosphate, suggesting some level of conservation in the catalytic mechanism with mammalian adenylyl cyclases. It shows a dose-dependent stimulation by Ca(2+) which can be reversed by high concentrations of phenothiazinic calmodulin inhibitors. However, bovine calmodulin fails to stimulate the enzyme. Using a yeast two-hybrid screen it was found that TczAC interacts through its catalytic domain with the paraflagellar rod protein, a component of the flagellar structure. Furthermore, we demonstrate that TczAC can dimerize through the same domain. These results provide novel evidence of the possible localization and regulation of this protein. PMID- 12121995 TI - Drugs mediate the transcriptional activation of the 5-aminolevulinic acid synthase (ALAS1) gene via the chicken xenobiotic-sensing nuclear receptor (CXR). AB - Heme is an essential component in oxygen transport and metabolism in living systems. In non-erythropoietic cells, 5-aminolevulinate synthase (ALAS1) is the first and rate-limiting enzyme in the heme biosynthesis pathway. ALAS1 expression and heme levels are increased in vivo by drugs and other chemical inducers of cytochrome P450 hemoproteins through mechanisms that are poorly understood. In the present studies, a chicken genomic cosmid library was employed to isolate a major portion of the ALAS1 gene. Two drug-responsive enhancer sequences, 176 and 167 base pairs in length, were identified in the 5'-flanking region of the gene in reporter gene assays in the hepatoma cell line LMH. The relative potency of inducers to activate these enhancers corresponds to induction of ALAS1 mRNA levels in LMH cells. Analysis of putative transcription factor binding sites within the enhancers revealed DR5 and DR4 type recognition sequences for nuclear receptors. Drug activation of the enhancer elements was reduced at least 60% after mutagenesis of individual nuclear receptor binding sites and was virtually eliminated following alteration of both recognition sites within the respective elements. Electrophoretic mobility shift assays and transactivation studies demonstrate direct interactions between the nuclear receptor binding sites and the recently described chicken xenobiotic-sensing receptor, (CXR) implicating drug activation mechanisms for ALAS1 similar to those found in inducible cytochrome(s) P450. This is the first report describing direct transcriptional activation of ALAS1 by drugs via drug-responsive enhancer sequences. PMID- 12121996 TI - A vacuolar-type H+-pyrophosphatase governs maintenance of functional acidocalcisomes and growth of the insect and mammalian forms of Trypanosoma brucei. AB - Vacuolar proton pyrophosphatases (V-H(+)-PPases) are electrogenic proton pumps found in many organisms of considerable industrial, environmental, and clinical importance. V-H(+)-PPases of several parasites were shown to be associated with acidic vacuoles named acidocalcisomes, which contain polyphosphate and calcium. In this work we functionally characterized a Trypanosoma brucei V-H(+)-PPase gene by using double-stranded RNA interference methodology to produce inducible V-H(+) PPase-deficient strains of procyclic and bloodstream forms (PFiVP1 and BFiVP1). Acidocalcisomes of these mutated parasites lost acidity and contained 90% less polyphosphate. PFiVP1 did not release calcium after the addition of nigericin, and its total acidity was reduced by 70%. This mutant also failed to stabilize its intracellular pH on exposure to external basic pH >7.4 and recovered from intracellular acidification at a slower rate and to a more acidic final intracellular pH. In the absence of T. brucei V-H(+)-PPase expression, PFiVP1 and BFiVP1 grew at a slower rate with doubling times of 27 h instead of 15 h, and 10 h instead of 7.5 h, respectively. Moreover, BFiVP1 could not grow over 5 x 10(5) cells/ml corresponding to a cell density reduction of five times for bloodstream form stationary phase growth. PMID- 12121997 TI - Cyclic AMP-dependent transcriptional up-regulation of phosphodiesterase 4D5 in human airway smooth muscle cells. Identification and characterization of a novel PDE4D5 promoter. AB - Phosphodiesterase 4D (PDE4D), part of the complex cAMP-specific PDE4 family, plays a pivotal role in the regulation of airway smooth muscle relaxation by catalyzing the hydolysis of cAMP. Its gene on chromosome 5q12 encodes 5 splice variants, which show tissue-dependent expression and regulation. The genomic arrangement of PDE4D was determined using in silico methods, and a putative promoter of one of the protein kinase A-activated, long isoforms, PDE4D5 was identified. Promoter-luciferase constructs, transiently transfected into a beta(2) adrenoreceptor-expressing CHO-K1 cell line, were used to demonstrate that the PDE4D5 promoter up-regulated reporter gene expression in response to increased cell cAMP. Site-directed mutagenesis of the cAMP-response element (CRE) at position -201 identified this as the principal component of the mechanism underlying this cAMP responsiveness. In the second part of this study, cAMP dependent induction of PDE4D5 transcript in primary cultured human airway smooth muscle cells (hASMs) was demonstrated using both qualitative reverse transcriptase PCR and quantitative real-time PCR. Isolated PDE4D5 isoenzyme activity, measured after selective immunoprecipitation from hASMs, confirmed that this increase in expression led to an up-regulation of functional activity. We present evidence for cAMP-driven PDE4D5 up-regulation in hASMs and suggest a CRE containing, isoform-specific promoter as the primary mechanism. PMID- 12121998 TI - Threonine 79 is a hinge residue that governs the fidelity of DNA polymerase beta by helping to position the DNA within the active site. AB - DNA polymerase beta (pol beta) is an ideal system for studying the role of its different amino acid residues in the fidelity of DNA synthesis. In this study, the T79S variant of pol beta was identified using an in vivo genetic screen. T79S is located in the N-terminal 8-kDa domain of pol beta and has no contact with either the DNA template or the incoming dNTP substrate. The T79S protein produced 8-fold more multiple mutations in the herpes simplex virus type 1-thymidine kinase assay than wild-type pol beta. Surprisingly, T79S is a misincorporation mutator only when using a 3'-recessed primer-template. In the presence of a single nucleotide-gapped DNA substrate, T79S displays an antimutator phenotype when catalyzing DNA synthesis opposite template C and has similar fidelity as wild type opposite templates A, G, or T. Threonine 79 is located directly between two helix-hairpin-helix motifs located within the 8-kDa and thumb domains of pol beta. As the pol beta enzyme closes into its active form, the helix-hairpin-helix motifs appear to assist in the production and stabilization of a 90 degrees bend of the DNA. The function of the bent DNA is to present the templating base to the incoming nucleotide substrate. We propose that Thr-79 is part of a hydrogen bonding network within the helix-hairpin-helix motifs that is important for positioning the DNA within the active site. We suggest that alteration of Thr-79 to Ser disrupts this hydrogen bonding network and results in an enzyme that is unable to bend the DNA into the proper geometry for accurate DNA synthesis. PMID- 12121999 TI - A novel set of Wnt-Frizzled fusion proteins identifies receptor components that activate beta -catenin-dependent signaling. AB - Wnt proteins initiate the canonical (beta-catenin-regulated) signaling cascade by binding to seven-transmembrane spanning receptors of the Frizzled (Fz) family together with the coreceptors LRP5 and -6, members of the low density lipoprotein receptor-related protein family (LRP). Several reports have shown physical and functional associations between various Wnt, LRP, and Frizzled molecules; however, the underlying mechanisms for selectivity remain poorly understood. We present data on a novel set of Wnt-Fz fusion constructs that are useful for elucidating mechanisms of Wnt signal transduction specificity in both Xenopus embryos and 293T cells. In 293T cells, coexpression of several Wnt-Fz fusion proteins with LRP6, but not LRP5, significantly activated a Wnt-responsive promoter, Optimized TOPFlash. Interestingly, Wnt proteins from both the Wnt1 and Wnt5A classes, when fused to the same Frizzled, can synergize with LRP6 to activate signaling and induce secondary axes in Xenopus embryos. However, when several Wnt-Fz constructs containing different Frizzled molecules were tested, it was found that all Frizzled molecules are not equivalent in their ability to activate the canonical Wnt pathway in this context. The data suggest that the distinction between the two Wnt classes lies not in intrinsic differences in the molecules but via the Frizzled molecules with which they interact. PMID- 12122000 TI - Measuring kinesin's first step. AB - A variety of models have recently emerged to explain how the molecular motor kinesin is able to maintain processive movement for over 100 steps. Although these models differ in significant features, they all predict that kinesin's catalytic domains intermittently separate from each other as the motor takes 8-nm steps along the microtubule. Furthermore, at some point in this process, one molecule of ATP is hydrolyzed per step. However, exactly when hydrolysis and product release occur in relation to this forward step have not been established. Furthermore, the rate at which this separation occurs as well as the speed of motor stepping onto and release from the microtubule have not been measured. In the absence of this information, it is difficult to critically evaluate competing models of kinesin function. We have addressed this issue by developing spectroscopic probes whose fluorescence is sensitive to motor-motor separation or microtubule binding. The kinetics of these fluorescence changes allow us to directly measure how fast kinesin steps onto and releases from the microtubule and provide insight into how processive movement is maintained by this motor. PMID- 12122001 TI - Dendritic cell (DC)-specific intercellular adhesion molecule 3 (ICAM-3)-grabbing nonintegrin (DC-SIGN, CD209), a C-type surface lectin in human DCs, is a receptor for Leishmania amastigotes. AB - Dendritic cells (DCs) play a critical role in the initiation of the immunological response against Leishmania parasites. However, the receptors involved in amastigote-dendritic cell interaction are unknown, especially in absence of opsonizing antibodies. We have studied the interaction of Leishmania pifanoi axenic amastigotes with the C-type lectin DC-specific intercellular adhesion molecule (ICAM)-3-grabbing nonintegrin (DC-SIGN, CD209), a receptor for ICAM-2, ICAM-3, human immunodeficiency virus gp120, and Ebola virus. L. pifanoi amastigotes interact with immature human dendritic cells and CD209-transfected K562 cells in a time- and dose-dependent manner. Leishmania amastigote binding to human dendritic cells and DC-SIGN-transfected cells is inhibited by a function blocking DC-SIGN-specific monoclonal antibody. More importantly, this monoclonal antibody dramatically reduces internalization of Leishmania amastigotes by immature human DCs. These results constitute the first description of a nonviral pathogen ligand for DC-SIGN and provide evidence for a relevant role of DC-SIGN in Leishmania amastigote uptake by dendritic cells. Our finding has important implications for Leishmania host-cell interaction and the immunoregulation of cutaneous leishmaniasis. PMID- 12122002 TI - Inhibition of prostate tumor angiogenesis by the tumor suppressor CEACAM1. AB - We have previously shown that CEACAM1, a cell-adhesion molecule, acts as a tumor suppressor in prostate carcinoma. Expression of CEACAM1 in prostate cancer cells suppresses their growth in vivo. However, CEACAM1 has no effect on the growth of prostate cancer cells in vitro. This difference suggests that the antitumor effect of CEACAM1 may be due to inhibition of tumor angiogenesis, perhaps by increased secretion of antiangiogenic molecules from the cells. In this study, we have demonstrated that expression of CEACAM1 in DU145 prostate cancer cells induced the production of a factor or factors that specifically blocked the growth of endothelial but not epithelial cells. Conditioned medium from the CEACAM1-expressing cells but not control luciferase-expressing cells inhibited endothelial cell migration up a gradient of stimulatory vascular endothelial growth factor in vitro and inhibited corneal neovascularization induced by basic fibroblast growth factor in vivo. Moreover, conditioned medium from CEACAM1 expressing cells induced endothelial cell apoptosis in vitro. Only medium conditioned by CEACAM1 mutants that were able to suppress tumor growth in vivo could cause endothelial cell apoptosis. These observations suggest that CEACAM1 mediated tumor suppression in vivo is, at least in part, due to the ability of CEACAM1 to inhibit tumor angiogenesis. PMID- 12122003 TI - Transmembrane topology of AgrB, the protein involved in the post-translational modification of AgrD in Staphylococcus aureus. AB - The accessory gene regulator (agr) of Staphylococcus aureus is the central regulatory system that controls the gene expression for a large set of virulence factors. This global regulatory locus consists of two transcripts: RNAII and RNAIII. RNAII encodes four genes (agrA, B, C, and D) whose gene products assemble a quorum sensing system. RNAIII is the effector of the Agr response. Both the agrB and agrD genes are essential for the production of the autoinducing peptide, which functions as a signal for the quorum sensing system. In this study, we demonstrated the transmembrane nature of AgrB protein in S. aureus. A transmembrane topology model of AgrB was proposed based on AgrB-PhoA fusion analyses in Escherichia coli. Two hydrophilic regions with several highly conserved positively charged amino acid residues among various AgrBs were found to be located in the cytoplasmic membrane as suggested by PhoA-AgrB fusion studies. However, this finding is inconsistent with the putative transmembrane profile of AgrB by computer analysis. Furthermore, we detected an intermediate peptide of processed AgrD from S. aureus cells expressing AgrB and a 6 histidine tagged AgrD. These results provide direct evidence that AgrB is involved in the proteolytic processing of AgrD. We speculate that AgrB is a novel protein with proteolytic enzyme activity and a transporter facilitating the export of the processed AgrD peptide. PMID- 12122004 TI - Crystal structure of horse carbonmonoxyhemoglobin-bezafibrate complex at 1.55-A resolution. A novel allosteric binding site in R-state hemoglobin. AB - Bezafibrate, an antilipidemic drug, is known as a potent allosteric effector of hemoglobin. The previously proposed mechanism for the allosteric potency of this drug was that it stabilizes and constrains the T-state of hemoglobin by specifically binding to the large central cavity of the T-state. Here we report a new allosteric binding site of fully liganded R-state hemoglobin for this drug. The high resolution crystal structure of horse carbonmonoxyhemoglobin in complex with bezafibrate reveals that the bezafibrate molecule lies near the surface of the E-helix of each alpha subunit and the complex maintains the quaternary structure of the R-state. Binding is caused by the close fit of bezafibrate into the binding pocket, which is composed of some hydrophobic residues and the heme edge, suggesting the importance of hydrophobic interactions. Upon binding of bezafibrate, the distance between Fe and the N epsilon(2) of distal His E7(alpha 58) is shortened by 0.22 A in the alpha subunit, whereas no significant structural changes are transmitted to the beta subunit. Oxygen equilibrium studies of R-state-locked hemoglobin with bezafibrate in a wet porous sol-gel indicate that bezafibrate selectively lowers the oxygen affinity of one type of subunit within the R-state, consistent with the structural data. These results disclose a new allosteric mechanism of bezafibrate and offer the first demonstration of how the allosteric effector interacts with R-state hemoglobin. PMID- 12122005 TI - Molecular mimicry of an HLA-B27-derived ligand of arthritis-linked subtypes with chlamydial proteins. AB - HLA-B27 is strongly associated with spondyloarthropathies, including ankylosing spondylitis and reactive arthritis. The latter disease is triggered by various Gram-negative bacteria. A dodecamer derived from the intracytoplasmic tail of HLA B27 was a natural ligand of three disease-associated subtypes (B*2702, B*2704, and B*2705) but not of two (B*2706 and B*2709), weakly or not associated to spondyloarthropathy. This peptide was strikingly homologous to protein sequences from arthritogenic bacteria, particularly to a region of the DNA primase from Chlamydia trachomatis. A synthetic peptide with this bacterial sequence bound in vitro disease-associated subtypes equally as the natural B27-derived ligand. The chlamydial peptide was generated by the 20 S proteasome from a synthetic 28-mer with the sequence of the corresponding region of the bacterial DNA primase. Molecular modeling suggested that the B27-derived and chlamydial peptides adopt very similar conformations in complex with B*2705. The results demonstrate that an HLA-B27-derived peptide mimicking arthritogenic bacterial sequences is a natural ligand of disease-associated HLA-B27 subtypes and suggest that the homologous chlamydial peptide might be presented by HLA-B27 on Chlamydia-infected cells. PMID- 12122006 TI - Cyclin F is degraded during G2-M by mechanisms fundamentally different from other cyclins. AB - Cyclin F, a cyclin that can form SCF complexes and bind to cyclin B, oscillates in the cell cycle with a pattern similar to cyclin A and cyclin B. Ectopic expression of cyclin F arrests the cell cycle in G(2)/M. How the level of cyclin F is regulated during the cell cycle is completely obscure. Here we show that, similar to cyclin A, cyclin F is degraded when the spindle assembly checkpoint is activated and accumulates when the DNA damage checkpoint is activated. Cyclin F is a very unstable protein throughout much of the cell cycle. Unlike other cyclins, degradation of cyclin F is independent of ubiquitination and proteasome mediated pathways. Interestingly, proteolysis of cyclin F is likely to involve metalloproteases. Rapid destruction of cyclin F does not require the N-terminal F box motif but requires the COOH-terminal PEST sequences. The PEST region alone is sufficient to interfere with the degradation of cyclin F and confer instability when fused to cyclin A. These data show that although cyclin F is degraded at similar time as the mitotic cyclins, the underlying mechanisms are entirely distinct. PMID- 12122007 TI - Impaired renal NaCl absorption in mice lacking the ROMK potassium channel, a model for type II Bartter's syndrome. AB - ROMK is an apical K(+) channel expressed in the thick ascending limb of Henle (TALH) and throughout the distal nephron of the kidney. Null mutations in the ROMK gene cause type II Bartter's syndrome, in which abnormalities of electrolyte, acid-base, and fluid-volume homeostasis occur because of defective NaCl reabsorption in the TALH. To understand better the pathogenesis of type II Bartter's syndrome, we developed a mouse lacking ROMK and examined its phenotype. Young null mutants had hydronephrosis, were severely dehydrated, and approximately 95% died before 3 weeks of age. ROMK-deficient mice that survived beyond weaning grew to adulthood; however, they had metabolic acidosis, elevated blood concentrations of Na(+) and Cl(-), reduced blood pressure, polydipsia, polyuria, and poor urinary concentrating ability. Whole kidney glomerular filtration rate was sharply reduced, apparently as a result of hydronephrosis, and fractional excretion of electrolytes was elevated. Micropuncture analysis revealed that the single nephron glomerular filtration rate was relatively normal, absorption of NaCl in the TALH was reduced but not eliminated, and tubuloglomerular feedback was severely impaired. These data show that the loss of ROMK in the mouse causes perturbations of electrolyte, acid-base, and fluid volume homeostasis, reduced absorption of NaCl in the TALH, and impaired tubuloglomerular feedback. PMID- 12122008 TI - Selective interleukin-12 synthesis defect in 12/15-lipoxygenase-deficient macrophages associated with reduced atherosclerosis in a mouse model of familial hypercholesterolemia. AB - Targeted gene disruption or overexpression of 12/15-lipoxygenase in mice on the genetic background of apolipoprotein E or low density lipoprotein-receptor (LDL R) deficiency has implicated 12/15-lipoxygenase in atherogenesis. The data support indirectly a role for 12/15-lipoxygenase in the oxidative modification of low density lipoprotein. In this study we set out to explore other potential mechanisms for 12/15-lipoxygenase in atherosclerosis using apolipoprotein B mRNA editing catalytic polypeptide-1/LDL-R double-deficient mice, a model highly related to the human condition of familial hypercholesterolemia. 12/15 Lipoxygenase deficiency in this strain led to approximately 50% decrease in aortic lesions in male and female mice at 8 months on a chow diet in the absence of cholesterol differences. While studying 12/15-lipoxygenase-deficient macrophages in culture, we discovered a remarkable selective defect (75-90% decrease) in interleukin-12 production but not in tumor necrosis factor-alpha or nitric oxide release, in response to lipopolysaccharide in the presence or absence of interferon-gamma priming. The lipopolysaccharide/interferon-gamma response was associated with a 33-50% decrease in nuclear interferon consensus sequence-binding protein, which is consistent with interferon consensus sequence binding protein containing protein complex-dependent regulation of the interleukin-12 p40 gene. The decrease in interleukin-12 production was recapitulated in vivo in mouse aortas of the triple knockout group and was reflected in a marked decrease in interferon-gamma expression. The data provide support for a novel mechanism linking the 12/15-lipoxygenase pathway to a known immunomodulatory Th1 cytokine in atherogenesis. PMID- 12122009 TI - Midkine binds to anaplastic lymphoma kinase (ALK) and acts as a growth factor for different cell types. AB - Midkine (MK) is a developmentally regulated, secreted growth factor homologous to pleiotrophin (PTN). To investigate the potential role of MK in tumor growth, we expressed MK in human SW-13 cells and studied receptor binding, signal transduction, and activity of MK. The MK protein stimulates soft agar colony formation in vitro and tumor growth of SW-13 cells in athymic nude mice, as well as proliferation of human endothelial cells from brain microvasculature and umbilical vein (HUVEC) in the low ng/ml range. MK binds to anaplastic lymphoma kinase (ALK), the receptor for PTN, with an apparent K(d) of 170 pm in intact cells, and this receptor binding of MK is competed by PTN with an apparent K(d) of approximately 20 pm. Monoclonal antibodies raised against the extracellular ligand-binding domain of ALK inhibit ALK receptor binding of MK as well as MK stimulated colony formation of SW-13 cells. Furthermore, MK stimulates ALK phosphorylation in WI-38 human fibroblasts and activates PI3-kinase and MAP kinase signal transduction in WI-38, HUVEC, neuroblastoma (SH SY-5Y) and glioblastoma (U87MG) cells that express the ALK protein. We conclude that MK can act as a growth, survival, and angiogenic factor during tumorigenesis and signals through the ALK receptor. PMID- 12122010 TI - A cold-regulated nucleic acid-binding protein of winter wheat shares a domain with bacterial cold shock proteins. AB - The molecular mechanisms of cold acclimation are still largely unknown; however, it has been established that overwintering plants such as winter wheat increases freeze tolerance during cold treatments. In prokaryotes, cold shock proteins are induced by temperature downshifts and have been proposed to function as RNA chaperones. A wheat cDNA encoding a putative nucleic acid-binding protein, WCSP1, was isolated and found to be homologous to the predominant CspA of Escherichia coli. The putative WCSP1 protein contains a three-domain structure consisting of an N-terminal cold shock domain with two internal conserved consensus RNA binding domains and an internal glycine-rich region, which is interspersed with three C terminal CX(2)CX(4)HX(4)C (CCHC) zinc fingers. Each domain has been described independently within several nucleotide-binding proteins. Northern and Western blot analyses showed that WCSP1 mRNA and protein levels steadily increased during cold acclimation, respectively. WCSP1 induction was cold-specific because neither abscisic acid treatment, drought, salinity, nor heat stress induced WCSP1 expression. Nucleotide binding assays determined that WCSP1 binds ssDNA, dsDNA, and RNA homopolymers. The capacity to bind dsDNA was nearly eliminated in a mutant protein lacking C-terminal zinc fingers. Structural and expression similarities to E. coli CspA suggest that WCSP1 may be involved in gene regulation during cold acclimation. PMID- 12122011 TI - CD40-mediated activation of NF-kappa B in airway epithelial cells. AB - We have reported previously that airway epithelial cells (AEC) express CD40 and that activation of this molecule stimulates the expression of inflammatory mediators, including the chemokine RANTES (regulated on activation normal T cell expressed and secreted). Because NF-kappaB regulates the expression of many inflammatory mediators, such as RANTES, we utilized CD40-mediated induction of RANTES expression to investigate the mechanisms that underlie CD40-mediated activation of NF-kappaB in AEC. Results demonstrate that, in AEC, intact NF kappaB sites were required for CD40-mediated activation of the RANTES promoter. To examine activation of NF-kappaB binding directly, electrophoretic mobility shift analyses were performed. These analyses revealed that CD40 ligation stimulated NF-kappaB binding and that the activated NF-kappaB complexes were composed of p65 subunits. Additional studies focused on the CD40-triggered signaling pathways that facilitate NF-kappaB activation. Findings show that CD40 engagement activated the IkappaB kinases IKK-alpha and IKK-beta and stimulated IkappaBalpha phosphorylation. Analyses also examined the role of tumor necrosis factor-associated factor (TRAF) molecules in CD40-mediated NF-kappaB activation within AEC. Stable transfectants expressing wild-type or mutant forms of the cytoplasmic domain of CD40 suggested that TRAF3, but not TRAF2, binding was essential for CD40-mediated RANTES expression. Further studies indicated that exogenous expression of wild-type TRAF3 enhanced activation of the RANTES promoter, whereas exogenous expression of wild-type TRAF2 inhibited this activation; TRAF3-mediated enhancement was dependent upon NF-kappaB. Together, these findings suggest that, in AEC, ligation of CD40 regulates the expression of inflammatory mediators, such as RANTES, via activation of NF-kappaB. Moreover, these results suggest that CD40-mediated signaling in AEC differs with previously reported findings observed in other cell models, such as B lymphocytes. PMID- 12122012 TI - Requirements for heterodimerization between the orphan nuclear receptor Nurr1 and retinoid X receptors. AB - The nuclear receptor nurr1 is a transcription factor involved in the development and maintenance of neurons synthesizing the neurotransmitter dopamine. Although the lack of nurr1 expression has dramatic consequences for these cells either in terms of differentiation or survival, the mechanisms by which nurr1 controls gene transcription still remain unclear. In the intent to understand better the modalities of action of this nuclear receptor, we have undertaken a systematic analysis of the transcriptional effects and DNA binding properties of nurr1 as a monomer or when forming dimers with the different isotypes of the retinoic X receptor (RXR). Here, we show that nurr1 acts as a gene activator independently of RXR and through an AF2-independent mechanism. In addition, heterodimerization with RXR is isotype-specific, involves multiple domains in the C-terminal region of nurr1, and requires RXR binding to DNA. RXR(alpha)-nurr1 and RXRgamma-nurr1 heterodimers bind direct repeat response elements and display no specific requirements with respect to half-site spacing. However, the retinoid responsiveness of DNA-bound heterodimers requires the reiteration of at least three nurr1 binding sites, thereby limiting retinoid-induced nurr1 transcriptional activity to specific direct response elements. PMID- 12122013 TI - Stabilization of telomere length and karyotypic stability are directly correlated with the level of hTERT gene expression in primary fibroblasts. AB - Telomere shortening and lack of telomerase activity have been implicated in cellular senescence in human fibroblasts. Expression of the human telomerase (hTERT) gene in sheep fibroblasts reconstitutes telomerase activity and extends their lifespan. However, telomere length is not maintained in all cell lines, even though in vitro telomerase activity is restored in all of them. Cell lines expressing higher levels of hTERT mRNA do not exhibit telomere erosion or genomic instability. By contrast, fibroblasts expressing lower levels of hTERT do exhibit telomere shortening, although the telomeres eventually stabilize at a shorter length. The shorter telomere lengths and the extent of karyotypic abnormalities are both functions of hTERT expression level. We conclude that telomerase activity is required to bypass senescence but is not sufficient to prevent telomere erosion and genomic instability at lower levels of expression. PMID- 12122014 TI - Csk homologous kinase (CHK) and ErbB-2 interactions are directly coupled with CHK negative growth regulatory function in breast cancer. AB - Our previous studies demonstrated that Csk homologous kinase (CHK) acts as a negative growth regulator of human breast cancer through inhibition of ErbB-2/neu mediated Src family kinase activity (Bougeret, C., Jiang, S., Keydar, I., and Avraham, H. (2001) J. Biol. Chem. 276, 33711-33720. The interaction between the CHK SH2 domain and Tyr(P)(1248) of the ErbB-2 receptor has been shown to be specific and critical for CHK function. In this report, we investigated whether the interaction of the CHK SH2 domain and ErbB-2 is directly related to the inhibition of heregulin-stimulated Src kinase activity. We constructed three CHK SH2 domain binding mutants: G129R (enhanced binding), R147K (inhibited binding), and R147A (disrupted binding). NMR spectra for the domains of each construct were used to evaluate their interaction with a Tyr(P)(1248)-containing ErbB-2 peptide. G129R showed enhanced binding to ErbB-2, whereas binding was completely disrupted by R147A. The enhanced binding mutant showed chemical shift changes at the same residues as wild-type CHK, indicating that this mutant has the same binding characteristics as the wild-type protein. Furthermore, inhibition of heregulin stimulated Src kinase activity was markedly diminished by R147A, whereas G129R mediated inhibition was stronger as compared with wild-type CHK. These results indicate that the specific interaction of CHK and ErbB-2 via the SH2 domain of CHK is directly related to the growth inhibitory effects of CHK. These new CHK high affinity binding constructs may serve as good candidates for inhibition of the ErbB-2/Src transduction pathway in gene therapy studies in breast cancer. PMID- 12122015 TI - Microfibril-associated glycoprotein-2 interacts with fibrillin-1 and fibrillin-2 suggesting a role for MAGP-2 in elastic fiber assembly. AB - Elastic fibers are composed of the protein elastin and a network of 10-12 nm microfibrils. The microfibrillar proteins include, among others, the fibrillins and microfibril-associated glycoproteins-1 and -2 (MAGP-1 and MAGP-2). Little is known about how microfibrillar proteins interact to support fiber assembly. We used the C-terminal half of MAGP-2 in a yeast two-hybrid library screen to identify relevant ligands. Six of 13 positive clones encoded known microfibrillar proteins, including fibrillin-1 and -2. Deletion analysis of partial fibrillin-1 and -2 clones revealed a calcium-binding epidermal growth factor repeat containing region near the C terminus responsible for binding. This region is distinct from the region of fibrillin-1 reported by others to bind MAGP-1. The MAGP-2 bait was unable to interact productively with other epidermal growth factor repeats in fibrillin-1, demonstrating specificity of the interaction. Deletion analysis of the MAGP-2 bait demonstrated that binding occurred in a core region containing 48% identity and 7 conserved cysteine residues with MAGP-1. Immunoprecipitation of MAGP-2 from transfected COS-7 cells resulted in the coprecipitation of fibrillin. These results demonstrate that MAGP-2 specifically interacts with fibrillin-1 and -2 and suggest that MAGP-2 may help regulate microfibrillar assembly. The results also demonstrate the utility of the yeast two-hybrid system to study protein-protein interactions of the extracellular matrix. PMID- 12122016 TI - Regulation of pituitary adenylate cyclase-activating polypeptide gene transcription by TTF-1, a homeodomain-containing transcription factor. AB - Pituitary adenylate cyclase-activating polypeptide (PACAP) is an important hypophysiotrophic factor as well as a regulator for immune, reproductive, and neural tissues. We recently found that TTF-1, a homeodomain-containing transcription factor essential for the development of the fetal diencephalon, is postnatally expressed in the hypothalamic area and plays a transcription regulatory role for certain neurohormones. Based on the similarity of synthesis sites between PACAP and TTF-1 and, moreover, on the presence of conserved core TTF-1 binding motifs in the 5'-flanking region of the PACAP gene, we sought to uncover a regulatory role of TTF-1 in PACAP gene transcription. The TTF-1 homeodomain binds to six of the seven putative binding domains observed in the 5' flanking region of the PACAP gene. In the C6 glioma cell-line, TTF-1 activates the PACAP promoter in a dose-dependent manner. This transactivation of PACAP by TTF-1 was totally removed when the core TTF-1 binding motif at -369 was deleted. RNase protection assays showed that TTF-1 and PACAP mRNAs have daily fluctuations in the rat hypothalamus. They both were at low levels during the day and high levels during the night. Intracerebroventricular administration of an antisense TTF-1 oligodeoxynucleotide significantly decreased the PACAP mRNA level as well as TTF-1 protein content in the rat hypothalamus, suggesting that TTF-1 also regulates PACAP transcription in vivo. Moreover, the TTF-1 promoter was inhibited by molecular oscillators of CLOCK and BMAL-1. Taken together, these data suggest that TTF-1 plays an important regulatory role in the gene transcription for PACAP, which may be important for the generation of a daily rhythm of hypothalamic PACAP gene expression. PMID- 12122018 TI - A nucleophile activation dyad in ribonucleases. A combined X-ray crystallographic/ab initio quantum chemical study. AB - Ribonucleases (RNases) catalyze the cleavage of the phosphodiester bond in RNA up to 10(15)-fold, as compared with the uncatalyzed reaction. High resolution crystal structures of these enzymes in complex with 3'-mononucleotide substrates demonstrate the accommodation of the nucleophilic 2'-OH group in a binding pocket comprising the catalytic base (glutamate or histidine) and a charged hydrogen bond donor (lysine or histidine). Ab initio quantum chemical calculations performed on such Michaelis complexes of the mammalian RNase A (EC ) and the microbial RNase T(1) (EC ) show negative charge build up on the 2'-oxygen upon substrate binding. The increased nucleophilicity results from stronger hydrogen bonding to the catalytic base, which is mediated by a hydrogen bond from the charged donor. This hitherto unrecognized catalytic dyad in ribonucleases constitutes a general mechanism for nucleophile activation in both enzymic and RNA-catalyzed phosphoryl transfer reactions. PMID- 12122017 TI - Head involution defective (Hid)-triggered apoptosis requires caspase-8 but not FADD (Fas-associated death domain) and is regulated by Erk in mammalian cells. AB - The molecular machinery of apoptosis is evolutionarily conserved with some exceptions. One such example is the Drosophila proapoptotic gene Head involution defective (Hid), whose mammalian homologue is not known. Hid is apoptotic to mammalian cells, and we have examined the mechanism by which Hid induces death. We demonstrate for the first time a role for the extracellular signal-related kinase-1/2 (Erk-1/2) in the regulation of Hid function in mammalian cells. Bcl-2 and an inhibitor of caspase-9 blocked apoptosis, indicative of a role for the mitochondrion in this pathway, and we provide evidence for a role for caspase-8 in Hid-induced apoptosis. Thus, apoptosis was blocked by an inhibitor of caspase 8, deletion of caspase-8 rendered cells resistant to Hid-induced apoptosis, and Hid associated with caspase-8 in cell lysates. The Fas-associated death domain (FADD) was dispensable for the apoptotic function of Hid, indicating that Hid does not require extracellular death receptor signaling for the activation of caspase-8. In activated T cells, the cytokine interleukin-2 blocked caspase-8 processing and apoptosis, suggesting that survival cues from trophic factors may target a Hid-like intermediate present in mammalian cells. Thus, this study shows that Hid engages with conserved components of cellular death machinery and suggests that apoptotic paradigms characterized by FADD-independent activation of caspase-8 may involve a Hid-like molecule in mammalian cells. PMID- 12122020 TI - Sequences of the mouse N-acetylglucosaminyltransferase V (Mgat5) mRNA and an mRNA expressed by an Mgat5-deficient cell line. AB - N-acetylglucosaminyltransferase V, encoded by the Mgat5 gene, plays an important regulatory role in the synthesis of complex N-glycans, including the Lewis antigens. The elevated expression of this enzyme is regulated during development and is highly associated with cellular transformation and malignancy. In addition, mammary tumors produced in Mgat5 gene knockout mice show markedly reduced metastases, indicating that it plays an important role in this process. The expression of this transferase is important for normal T cell proliferation and function. The amino acid and mRNA sequences for this protein are very highly conserved among the reported species. We report here the sequence of the mouse Mgat5 mRNA. This sequence is very similar to the other mammalian sequences with 95%, 91%, and 87.8% overall sequence identity to rat, hamster, and human mRNAs, respectively. In addition, we have identified a 63-bp insertion mutation, introducing a premature stop codon in the coding region of the Mgat5 mRNA expressed in a N-acetylglucosaminyltransferase V-deficient cell line, PhaR(2.1). PMID- 12122019 TI - Real-time observation of coiled-coil domains and subunit assembly in intermediate filaments. AB - We have utilized electron paramagnetic resonance spectroscopy to study secondary structure, subunit interaction, and molecular orientation of vimentin molecules within intact intermediate filaments and assembly intermediates. Spectroscopy data prove alpha-helical coiled-coil structures at individual amino acids 316-336 located in rod 2B. Analysis of positions 305, 309, and 312 identify this region as conforming to the helical pattern identified within 316-336 and thus demonstrates that, contrary to some previous predictions, this region is in an alpha-helical conformation. We show that by varying the position of the spin label, we can identify both intra- and inter-dimer interactions. With a label attached to the outside of the alpha-helix, we have been able to measure interactions between positions 348 of separate dimers as they align together in intact filaments, identifying the exact point of overlap. By mixing different spin-labeled proteins, we demonstrate that the interaction at position 348 is the result of an anti-parallel arrangement of dimers. This approach provides high resolution structural information (<2 nm resolution), can be used to identify molecular arrangements between subunits in an intact intermediate filament, and should be applicable to other noncrystallizable filamentous systems as well as to the study of protein fibrils. PMID- 12122021 TI - Studies on the metal binding sites in the catalytic domain of beta1,4 galactosyltransferase. AB - The catalytic domain of bovine beta1,4-galactosyltransferase (beta4Gal-T1) has been shown to have two metal binding sites, each with a distinct binding affinity. Site I binds Mn(2+) with high affinity and does not bind Ca(2+), whereas site II binds a variety of metal ions, including Ca(2+). The catalytic region of beta4Gal-T1 has DXD motifs, associated with metal binding in glycosyltransferases, in two separate sequences: D(242)YDYNCFVFSDVD(254) (region I) and W(312)GWGGEDDD(320) (region II). Recently, the crystal structure of beta4Gal-T1 bound with UDP, Mn(2+), and alpha-lactalbumin was determined in our laboratory. It shows that in the primary metal binding site of beta4Gal-T1, the Mn(2+) ion, is coordinated to five ligands, two supplied by the phosphates of the sugar nucleotide and the other three by Asp254, His347, and Met344. The residue Asp254 in the D(252)VD(254) sequence in region I is the only residue that is coordinated to the Mn(2+) ion. Region II forms a loop structure and contains the E(317)DDD(320) sequence in which residues Asp318 and Asp319 are directly involved in GlcNAc binding. This study, using site-directed mutagenesis, kinetic, and binding affinity analysis, shows that Asp254 and His347 are strong metal ligands, whereas Met344, which coordinates less strongly, can be substituted by alanine or glutamine. Specifically, substitution of Met344 to Gln has a less severe effect on the catalysis driven by Co(2+). Glu317 and Asp320 mutants, when partially activated by Mn(2+) binding to the primary site, can be further activated by Co(2+) or inhibited by Ca(2+), an effect that is the opposite of what is observed with the wild-type enzyme. PMID- 12122022 TI - Molecular analysis of a novel family of complex glycoinositolphosphoryl ceramides from Cryptococcus neoformans: structural differences between encapsulated and acapsular yeast forms. AB - Complex glycoinositolphosphoryl ceramides (GIPCs) have been purified from a pathogenic encapsulated wild-type (WT) strain of Cryptococcus neoformans var. neoformans and from an acapsular mutant (Cap67). The structures of the GIPCs were determined by a combination of tandem mass spectrometry, nuclear magnetic resonance spectroscopy, methylation analysis, gas chromatography-mass spectrometry, and chemical degradation. The main GIPC from the WT strain had the structure Manp(alpha1-3)[Xylp(beta1-2)] Manp(alpha1-4)Galp(beta1-6)Manp(alpha1 2)Ins-1-phosphoryl ceramide (GIPC A), whereas the compounds from the acapsular mutant were more heterogeneous in their glycan chains, and variants with Manp(alpha1-6) (GIPC B), Manp(alpha1-6) Manp(alpha1-6) (GIPC C), and Manp(alpha1 2)Manp(alpha1-6)Manp(alpha1-6) (GIPC D) substituents linked to the nonreducing terminal mannose residue found in the WT GIPC A were abundant. The ceramide moieties of C. neoformans GIPCs were composed of a C(18) phytosphingosine long chain base mainly N-acylated with 2-hydroxy-tetracosanoic acid in the WT GIPC while in the acapsular Cap67 mutant GIPCs, as well as 2-hydroxy-tetracosanoic acid, the unusual 2,3-dihydroxy-tetracosanoic acid was characterized. In addition, structural analysis revealed that the amount of GIPC in the WT cells was fourfold less of that in the acapsular mutant. PMID- 12122023 TI - Large-scale preparation, purification, and characterization of hyaluronan oligosaccharides from 4-mers to 52-mers. AB - Hyaluronan (HA) was depolymerized by partial digestion with testicular hyaluronidase and separated into size-uniform HA oligosaccharides from 4-mers to 52-mers by anion exchange chromatography after removal of the hyaluronidase. The purity and size of each HA oligosaccharide was confirmed by using HPLC analyses, FACE, and ESI-MS. (1)H and (13)C NMR assignments and elemental analyses were obtained for each HA oligosaccharide. Endotoxins, proteins, and DNA were absent or in trace amounts in these HA oligosaccharides. Gram/mg-scale hyaluronan oligosaccharides were obtained from 200 g of HA starting material. These pure, size-uniform, and large range of HA oligosaccharides will be available for investigating important biological functions of HA, such as for the determination of the size(s) of HA oligosaccharides that induce angiogenesis or mediate inflammatory responses, and to interact with HA-binding proteins and receptors both in in vitro and in vivo studies. PMID- 12122024 TI - Characterization of mycobacterial protein glycosyltransferase activity using synthetic peptide acceptors in a cell-free assay. AB - Synthetic peptides derived from a 45-kDa glycoprotein antigen of Mycobacterium tuberculosis were shown to function as glycosyltransferase acceptors for mannose residues in a mannosyltransferase cell-free assay. The mannosyltransferase activity was localized within both isolated membranes and a P60 cell wall fraction prepared from the rapidly growing mycobacterial strain, Mycobacterium smegmatis. Incorporation of radiolabel from GDP-[(14)C]mannose was inhibited by the addition of amphomycin, indicating that the glycosyl donor for the peptide acceptors was a member of the mycobacterial polyprenol-P-mannose (PPM) family of activated glycosyl donors. Furthermore, a direct demonstration of transfer from the in situ generated PP[(14)C]Ms was also demonstrated. It was also found that the enzyme activity was sensitive to changes in overall peptide length and amino acid composition. Because glycoproteins are present on the mycobacterial cell surface and are available for interaction with host cells during infection, protein glycosyltransferases may provide novel drug targets. The development of a cell-free mannosyltransferase assay will now facilitate the cloning and biochemical characterisation of the relevant enzymes from M. tuberculosis. PMID- 12122025 TI - Molecular cloning, gene organization, and expression of mouse Mpi encoding phosphomannose isomerase. AB - Phosphomannose isomerase (PMI) interconverts fructose-6-P (Fru-6-P) and mannose-6 P (Man-6-P), linking energy metabolism to protein glycosylation. We have cloned the mouse Mpi cDNA, analyzed its genomic organization, and studied the expression in different tissues. The Mpi gene has eight exons covering 7.2 kb. The structure and intron-exon boundaries are essentially the same as its human ortholog with 85% amino acid identity. Mpi is alternatively spliced at the 3' end, resulting in three messages with different 3'-untranslated regions. Mpi expression is regulated at both the transcription and translation levels, with the highest expression level in testis. Rabbit antibodies prepared against mouse PMI expressed in E. coli recognize a single 47-kDa band. Immunohistochemistry of mouse tissues shows general cytosolic staining in all cells. In testis, staining is intense in round spermatids and residual bodies, moderate in pachytene spermatocytes, and weak in spermatogonia and spermatozoa. In contrast, northern blot analysis shows comparable transcripts of 1.8 and 1.6 kb in pachytene spermatocytes and round spermatids, suggesting delayed translation of PMI. The stage-specific expression of PMI in testis may be important for KDN synthesis, which requires Man-6-P, or it may be needed to ensure sufficient glycosylation precursors in cells that do not utilize glucose and instead rely on lactate and pyruvate. PMID- 12122026 TI - Macrophage C-type lectin on bone marrow-derived immature dendritic cells is involved in the internalization of glycosylated antigens. AB - Bone marrow-derived dendritic cells (DCs) were examined for the expression of the murine macrophage C-type lectin specific for galactose and N-acetylgalactosamine (mMGL). Flow cytometric analysis after double staining for MHC class II and mMGL with specific monoclonal antibodies indicated that mMGL was expressed on immature DCs with low to moderate levels of MHC class II and down-regulated during maturation. Immature DCs bound and internalized alpha-N-acetylgalactosaminides conjugated to soluble polyacrylamide (alpha-GalNAc polymers), whereas mature DCs and bone marrow cells did not. The two-color flow cytometric profiles indicated that the degree of alpha-GalNAc polymer bindings exactly coincided with the intensity of the binding of a mMGL-specific monoclonal antibody LOM-14. The internalized alpha-GalNAc polymers seemed to be transported to MHC class II compartments. Thus, mMGL is transiently expressed on bone marrow-derived DCs during their development and maturation and suggested to be involved in the uptake of glycosylated antigens for presentation. PMID- 12122027 TI - Spatial analysis of ocular dominance patterns in monocularly deprived cats. AB - Monocular deprivation during a critical period in postnatal development leads to a shift in functional and anatomical ocular dominance at the expense of the deprived eye. We analyzed the complete two-dimensional pattern of [(3)H]proline labeled afferents in primary visual cortex (area 17) of cats monocularly deprived of vision at eye opening. Substantial shrinkage of deprived eye territory in favor of the normal eye extended into optic disc and monocular segment representations. However, small domains of deprived eye afferents were distributed evenly over the entire visual field representation. Interestingly, normal and deprived eye afferents overlapped extensively in the ipsilateral and in the peripheral contralateral visual field representation of the deprived eye, so that ipsi- and contralateral ocular dominance patterns are not at all complementary. We suggest that this could be the result of both an earlier maturation of crossed versus uncrossed visual pathways and of a maturational gradient within area 17 leading to a lower vulnerability of the central visual field representation to monocular deprivation. Quantitative analysis, using a triangulation algorithm which confirmed previously described larger spacing of adjacent ocular dominance columns in strabismic cats, revealed no difference in spacing of ocular dominance domains in area 17 between monocularly deprived and normals cats. In addition, column spacing was very similar in the same animal and in littermates, indicating that the genetic influence on columnar layout is stronger than previously assumed. PMID- 12122028 TI - Hearing after congenital deafness: central auditory plasticity and sensory deprivation. AB - The congenitally deaf cat suffers from a degeneration of the inner ear. The organ of Corti bears no hair cells, yet the auditory afferents are preserved. Since these animals have no auditory experience, they were used as a model for congenital deafness. Kittens were equipped with a cochlear implant at different ages and electro-stimulated over a period of 2.0-5.5 months using a monopolar single-channel compressed analogue stimulation strategy (VIENNA-type signal processor). Following a period of auditory experience, we investigated cortical field potentials in response to electrical biphasic pulses applied by means of the cochlear implant. In comparison to naive unstimulated deaf cats and normal hearing cats, the chronically stimulated animals showed larger cortical regions producing middle-latency responses at or above 300 microV amplitude at the contralateral as well as the ipsilateral auditory cortex. The cortex ipsilateral to the chronically stimulated ear did not show any signs of reduced responsiveness when stimulating the 'untrained' ear through a second cochlear implant inserted in the final experiment. With comparable duration of auditory training, the activated cortical area was substantially smaller if implantation had been performed at an older age of 5-6 months. The data emphasize that young sensory systems in cats have a higher capacity for plasticity than older ones and that there is a sensitive period for the cat's auditory system. PMID- 12122029 TI - Cognitive and neural mechanisms of decision biases in recognition memory. AB - In recognition memory tasks, stimuli can be classified as "old" either on the basis of accurate memory or a bias to respond "old", yet bias has received little attention in the cognitive neuroscience literature. Here we examined the pattern and timing of bias-related effects in event-related brain potentials (ERPs) to determine whether the bias is linked more to memory retrieval or to response verification processes. Participants were divided into a High Bias and a Low Bias group according to their bias to respond "old". These groups did not differ in recognition accuracy or in the ERP pattern to items that actually were old versus new (Objective Old/New Effect). However, when the old/new distinction was based on each subject's perspective, i.e. when items judged "old" were compared with those judged "new" (Subjective Old/New Effect), significant group differences were observed over prefrontal sites with a timing (300-500 ms poststimulus) more consistent with bias acting early on memory retrieval processes than on post retrieval response verification processes. In the standard old/new effect (Hits vs Correct Rejections), these group differences were intermediate to those for the Objective and the Subjective comparisons, indicating that such comparisons are confounded by response bias. We propose that these biases are top-down controlled processes mediated by prefrontal cortex areas. PMID- 12122030 TI - Machine psychology: autonomous behavior, perceptual categorization and conditioning in a brain-based device. AB - In studying brain activity during the behavior of living animals, it is not possible simultaneously to analyze all levels of control from molecular events to motor responses. To provide insights into how levels of control interact, we have carried out synthetic neural modeling using a brain-based real-world device. We describe here the design and performance of such a device, designated Darwin VII, which is guided by computer-simulated analogues of cortical and subcortical structures. All levels of Darwin VII's neural architecture can be examined simultaneously as the device behaves in a real environment. Analysis of its neural activity during perceptual categorization and conditioned behavior suggests neural mechanisms for invariant object recognition, experience-dependent perceptual categorization, first-order and second-order conditioning, and the effects of different learning rates on responses to appetitive and aversive events. While individual Darwin VII exemplars developed similar categorical responses that depended on exploration of the environment and sensorimotor adaptation, each showed highly individual patterns of changes in synaptic strengths. By allowing exhaustive analysis and manipulation of neuroanatomy and large-scale neural dynamics, such brain-based devices provide valuable heuristics for understanding cortical interactions. These devices also provide the groundwork for the development of intelligent machines that follow neurobiological rather than computational principles in their construction. PMID- 12122031 TI - Receptor-activated calcium signals in tangentially migrating cortical cells. AB - Recent studies have shown that the two main types of cortical neurons, pyramidal and nonpyramidal, have different origins and use different migratory routes- radial and tangential respectively. The role of neurotransmitters in radial migration is well known; however, there are no data about their effect on intracellular calcium--[Ca(2+)](i)--in tangentially migrating cells. We have performed ratiometric and confocal calcium imaging of 1,1'-dioctodecyl-3,3,3',3' tetramethylindocarbocyanine labelled tangentially migrating neurons in the intermediate zone cells of fetal rat coronal slices. Superfusion with N-methyl-D aspartic acid (NMDA) leads to an increase in [Ca(2+)](i), which is blocked by the antagonist APV or the presence of Mg(2+) in the medium. Kainate produced an increase in [Ca(2+)](i) that could be blocked by the non-NMDA antagonist CNQX. Muscimol, an agonist of GABAa-receptors, produced a transitory increase in [Ca(2+)](i) that was blocked by the specific antagonist bicuculline or the presence of tetrodotoxin in the medium. We conclude that tangentially migrating cells display consistent [Ca(2+)](i) changes in response to agonists of NMDA, non NMDA and GABAa receptors, suggesting that these cells are quite mature and homogeneous. The endogenous activation of these receptors may have either a direct effect on tangential migration or modulate the response of migrating cells to external cues. PMID- 12122032 TI - Neurophysiological evidence of error-monitoring deficits in patients with schizophrenia. AB - The present study was designed to investigate the time-course of neural activity underlying the disruption of response monitoring in patients with schizophrenia. Event-related brain potentials were recorded from 12 patients with schizophrenia and from 12 age-matched controls while they performed a computerized version of the Stroop color-naming task. In control participants, but not in patients with schizophrenia, intrusion errors elicited an error-related negativity (ERN) that peaked at approximately 40 ms after the response and was maximum over the central region of the scalp. Brain electrical source analysis revealed an anterior cingulate generator for the ERN. Patients also showed reduced error-related slowing of response time following intrusion errors. These findings provide neuro physiological evidence indicating that deficits in error monitoring in schizophrenia arise from a disruption of error-detection processes, possibly attributable to anterior cingulate dysfunction. PMID- 12122033 TI - Modulation of motor and premotor activity during imitation of target-directed actions. AB - Behavioral studies reveal that imitation performance and the motor system are strongly influenced by the goal of the action to be performed. We used functional magnetic resonance imaging (fMRI) to assess the effect of explicit action goals on neural activity during imitation. Subjects imitated index finger movements in the absence and presence of visible goals (red dots that were reached for by the finger movement). Finger movements were either ipsilateral or contralateral. The pars opercularis of the inferior frontal gyrus showed increased blood oxygen level-dependent fMRI signal bilaterally for imitation of goal-oriented actions, compared with imitation of actions with no explicit goal. In addition, bilateral dorsal premotor areas demonstrated greater activity for goal-oriented actions, for contralateral movements and an interaction effect such that goal-oriented contralateral movements yielded the greatest activity. These results support the hypothesis that areas relevant to motor preparation and motor execution are tuned to coding goal-oriented actions and are in keeping with single-cell recordings revealing that neurons in area F5 of the monkey brain represent goal-directed aspects of actions. PMID- 12122034 TI - Regional brain shape abnormalities persist into adolescence after heavy prenatal alcohol exposure. AB - We assessed regional brain shape abnormalities and spatial relationships between brain shape and abnormalities observed in the underlying tissue in children and adolescents prenatally exposed to large quantities of alcohol. We used high resolution, 3-D, structural magnetic resonance imaging data and novel, whole brain, surface-based image analysis procedures to study 21 subjects with heavy prenatal alcohol exposure (8-22 years, mean age 12.6 years) and 21 normally developing control subjects (8-25 years, mean age 13.5 years). Significant brain size and shape abnormalities were observed in the alcohol-exposed subjects in inferior parietal/ perisylvian regions bilaterally, where their brains appeared to be narrower than those of the controls in the same general location where they also had increased gray matter density. Highly significant decreased brain surface extent or reduced brain growth was also observed in the ventral aspects of the frontal lobes most prominent in the left hemisphere. For the first time in this report we have mapped brain morphologic abnormalities to the cortical surface in subjects with prenatal alcohol exposure and have shown that the size and shape of the brain is altered in these individuals. The results imply that brain growth continues to be adversely affected long after the prenatal insult of alcohol exposure to the developing brain and the brain regions most implicated, frontal and inferior parietal/ perisylvian, may be consistent with behavioral deficits characteristic of individuals prenatally exposed to alcohol. PMID- 12122035 TI - Distributed cortical systems in visual short-term memory revealed by event related functional magnetic resonance imaging. AB - The spatio-temporal distribution of brain activity as revealed by non-invasive functional imaging helps to elucidate the neuronal encoding and processing strategies required by complex cognitive tasks. We investigated visual short-term memory for objects, places and conjunctions in humans using event-related time resolved functional magnetic resonance imaging that permitted segregation of encoding, retention and retrieval phases. All conditions were accompanied by the activation of a widespread network of parietal and prefrontal areas during the retention phase, but this retention-related activity showed additional modulations depending on task instructions. These modulations confirmed a posterior-anterior and right-left dissociation for spatial versus non-spatial memory and revealed that conjunction memory does not rely on a linear addition of the component processes. PMID- 12122036 TI - Oscillations in the alpha band (9-12 Hz) increase with memory load during retention in a short-term memory task. AB - To study the role of brain oscillations in working memory, we recorded the scalp electroencephalogram (EEG) during the retention interval of a modified Sternberg task. A power spectral analysis of the EEG during the retention interval revealed a clear peak at 9-12 Hz, a frequency in the alpha band (8-13 Hz). In apparent conflict with previous ideas according to which alpha band oscillations represent brain "idling", we found that the alpha peak systematically increased with the number of items held in working memory. The enhancement was prominent over the posterior and bilateral central regions. The enhancement over posterior regions is most likely explained by the well known alpha rhythm produced close to the parietal-occipital fissure, whereas the lateral enhancement could be explained by sources in somato-motor cortex. A time-frequency analysis revealed that the enhancement was present throughout the last 2.5 s of the 2.8 s retention interval and that alpha power rapidly diminished following the probe. The load dependence and the tight temporal regulation of alpha provide strong evidence that the alpha generating system is directly or indirectly linked to the circuits responsible for working memory. Although a clear peak in the theta band (5-8 Hz) was only detectable in one subject, other lines of evidence indicate that theta occurs and also has a role in working memory. Hypotheses concerning the role of alpha band activity in working memory are discussed. PMID- 12122037 TI - Spaceflight induces changes in the synaptic circuitry of the postnatal developing neocortex. AB - The establishment of the adult pattern of neocortical circuitry depends on various intrinsic and extrinsic factors, whose modification during development can lead to alterations in cortical organization and function. We report the effect of 16 days of spaceflight [Neurolab mission; from postnatal day 14 (P14) to P30] on the neocortical representation of the hindlimb synaptic circuitry in rats. As a result, we show, for the first time, that development in microgravity leads to changes in the number and morphology of cortical synapses in a laminar specific manner. In the layers II/III and Va, the synaptic cross-sectional lengths were significantly larger in flight animals than in ground control animals. Flight animals also showed significantly lower synaptic densities in layers II/III, IV and Va. The greatest difference was found in layer II/III, where there was a difference of 344 million synapses per mm(3) (15.6% decrease). Furthermore, after a 4 month period of re-adaptation to terrestrial gravity, some changes disappeared (i.e. the alterations were transient), while conversely, some new differences also appeared. For example, significant differences in synaptic density in layers II/III and Va after re-adaptation were no longer observed, whereas in layer IV the density of synapses increased notably in flight animals (a difference of 185 million synapses per mm(3) or 13.4%). In addition, all the changes observed only affected asymmetrical synapses, which are known to be excitatory. These results indicates that terrestrial gravity is a necessary environmental parameter for normal cortical synaptogenesis. These findings are fundamental in planning future long-term spaceflights. PMID- 12122038 TI - Phosphorylation of the postsynaptic density-95 (PSD-95)/discs large/zona occludens-1 binding site of stargazin regulates binding to PSD-95 and synaptic targeting of AMPA receptors. AB - Dynamic regulation of AMPA-type receptors at the synapse is proposed to play a critical role in alterations of the synaptic strength seen in cellular models of learning and memory such as long-term potentiation in the hippocampus. Stargazin, previously identified as an AMPA receptor (AMPAR)-interacting protein, is critical for surface expression and synaptic targeting of AMPARs. Stargazin interacts with postsynaptic density-95/discs large/zona occludens-1 (PDZ) proteins via a C-terminal PDZ binding motif. Interestingly, the C terminal of stargazin also predicts phosphorylation at a threonine residue critical for PDZ protein binding. Because protein phosphorylation regulates synaptic plasticity, we characterized this site and the effects of stargazin phosphorylation on AMPAR function. In vitro peptide phosphorylation assays and Western blot analysis with phospho-stargazin-specific antibodies indicate that the critical threonine within the stargazin PDZ binding site is phosphorylated by protein kinase A. This phosphorylation disrupts stargazin interaction and clustering with postsynaptic density-95 (PSD-95) in transfected COS-7 cells. Furthermore, a stargazin construct with a Thr-to-Glu mutation that mimics phosphorylation fails to cluster at synaptic spines and downregulates synaptic AMPAR function in cultured hippocampal neurons. These data suggest that phosphorylation of the stargazin PDZ ligand can disrupt stargazin interaction with PSD-95 and thereby regulate synaptic AMPAR function. PMID- 12122040 TI - Automatic postural responses are delayed by pyridoxine-induced somatosensory loss. AB - Pyridoxine given in large doses is thought to destroy selectively the large diameter peripheral sensory nerve fibers, leaving motor fibers intact. This study examined the effects of pyridoxine-induced somatosensory loss on automatic postural responses to sudden displacements of the support surface in the standing cat. Two cats were trained to stand on four force plates mounted on a movable platform. They were given pyridoxine (350 mg/kg, i.p.) on 2 successive days (0 and 1). Electromyographic (EMG) activity was recorded from selected hindlimb muscles during linear ramp-and-hold platform displacements in each of 12 directions at 15 cm/sec. In control trials onset latencies of evoked activity in hindlimb flexor and extensor muscles ranged from 40 to 65 msec after the onset of platform acceleration. After injection the EMG latencies increased over days, becoming two to three times longer than controls by day 7. Excursions of the body center of mass (CoM) in the direction opposite to that of platform translation were significantly greater at day 7 compared with controls, and the time at which the CoM subsequently reversed direction was delayed. Both animals were ataxic from day 2 onward. Histological analysis of cutaneous and muscle nerves in the hindlimb revealed a significant loss of fibers in the group I range. Our results suggest that large afferent fibers are critical for the timing of automatic postural responses to ensure coordinated control of the body CoM and balance after unexpected disturbances of the support surface. PMID- 12122039 TI - Role for reelin in the development of granule cell dispersion in temporal lobe epilepsy. AB - The reelin signaling pathway plays a crucial role during the development of laminated structures in the mammalian brain. Reelin, which is synthesized and secreted by Cajal-Retzius cells in the marginal zone of the neocortex and hippocampus, is proposed to act as a stop signal for migrating neurons. Here we show that a decreased expression of reelin mRNA by hippocampal Cajal-Retzius cells correlates with the extent of migration defects in the dentate gyrus of patients with temporal lobe epilepsy. These results suggest that reelin is required for normal neuronal lamination in humans, and that deficient reelin expression may be involved in migration defects associated with temporal lobe epilepsy. PMID- 12122041 TI - Altered electromyographic activity pattern of rat soleus muscle transposed into the bed of antagonist muscle. AB - Human patients suffering from motor paralysis of the leg can learn, to some extent, how to use the transposed antagonistic muscle in place of the damaged or ineffective muscle. Experiments on animals showed opposite results, although in a few experiments the functional reorganization of the activity of the transposed muscle was not excluded. In our experiments, we performed transposition of the soleus (Sol) with a preserved innervation into the bed of the removed extensor digitorum longus (EDL) in 6-d-old pups and 3-month-old rats. The locomotor and reflex EMG activity of the transposed Sol (trSol) was recorded in chronic experiments 3-4 months later. Our results showed that the EMG activity of the Sol might be modified when the muscle is transposed into the bed of the antagonistic muscle EDL. The modification consisted of an additional burst of activity during the swing phase and of reflex response of the muscle to the plantar flexion. This modification was present in all animals operated on at 6 d of age and in two of six adults. After chronic experiments, we excluded the possibility that the flexor-like activity was induced by additional innervation of the trSol by the cut end of the EDL nerve. We suggest that the observed modifications of activity of trSol might be caused by readjustment of the spinal network organization to the new functional demands. Two factors might be responsible for this readjustment: the immaturity of the nervous system at the moment of surgery and preserved afferent innervation of the transposed muscle. PMID- 12122042 TI - Eighteen-month-old Fischer 344 rats fed a spinach-enriched diet show improved delay classical eyeblink conditioning and reduced expression of tumor necrosis factor alpha (TNFalpha ) and TNFbeta in the cerebellum. AB - Diets high in antioxidant properties are known to reverse some deficits in neuronal and cognitive function that occur in aging animals. Antioxidants are also known to reduce levels of proinflammatory factors in the CNS. We report here that 6 weeks of a spinach-enriched diet ameliorates deficits in cerebellar dependent delay classical eyeblink learning and reduces the proinflammatory cytokines tumor necrosis factor alpha (TNFalpha) and TNFbeta in the cerebelli of eyeblink-trained animals. Eighteen-month-old Fischer 344 rats were given spinach enriched lab chow or regular lab chow for 6 weeks. The rats were then given 6 d of 30 trials per day training using a 3 kHz tone conditioned stimulus and airpuff unconditioned stimulus. Rats were killed 3 weeks after eyeblink training. Cytokine expression was measured using RNase protection assay analysis in the eyeblink-trained animals and in a group of young control animals given regular lab chow diet. Old animals on the spinach-enriched lab chow diet learned delay eyeblink conditioning significantly faster than old animals on the regular diet. Cerebelli from older animals on the spinach-enriched diet had significantly less TNFalpha and TNFbeta than cerebelli from older animals on the control diet. PMID- 12122043 TI - Enhanced inhibition of synaptic transmission by dopamine in the nucleus accumbens during behavioral sensitization to cocaine. AB - Neural adaptations in the nucleus accumbens (NAc), a key component of the mesolimbic dopamine (DA) system, are thought to mediate several of the long-term behavioral sequelas of chronic in vivo exposure to drugs of abuse. Here, we examine whether the modulation of excitatory synaptic transmission by DA in the NAc shell is modified after chronic cocaine exposure that induced behavioral sensitization. The DA-induced inhibition of AMPA receptor-mediated synaptic responses was enhanced in cocaine-treated mice, an effect that was caused by activation of D1-like receptors. DA did not enhance NMDA receptor-mediated synaptic responses in saline- and cocaine-treated mice or in the dorsal striatum of control mice. We hypothesize that the enhanced inhibitory effects of DA on synaptic transmission in the NAc are one of a number of adaptations that contribute to a decrease in excitatory drive to NAc after exposure to drugs of abuse. PMID- 12122044 TI - Sympathoexcitation by bradykinin involves Ca2+-independent protein kinase C. AB - Bradykinin has long been known to excite sympathetic neurons via B(2) receptors, and this action is believed to be mediated by an inhibition of M-currents via phospholipase C and inositol trisphosphate-dependent increases in intracellular Ca(2+). In primary cultures of rat superior cervical ganglion neurons, bradykinin caused an accumulation of inositol trisphosphate, an inhibition of M-currents, and a stimulation of action potential-mediated transmitter release. Blockade of inositol trisphosphate-dependent signaling cascades failed to affect the bradykinin-induced release of noradrenaline, but prevented the peptide-induced inhibition of M-currents. In contrast, inhibition or downregulation of protein kinase C reduced the stimulation of transmitter release, but not the inhibition of M-currents, by bradykinin. In cultures of superior cervical ganglia, classical (alpha, betaI, betaII), novel (delta, epsilon), and atypical (zeta) protein kinase C isozymes were detected by immunoblotting. Bradykinin induced a translocation of Ca(2+)-independent protein kinase C isoforms (delta and epsilon) from the cytosol to the membrane of the neurons, but left the cellular distribution of other isoforms unchanged. This activation of Ca(2+)-independent protein kinase C enzymes was prevented by a phospholipase C inhibitor. The bradykinin-dependent stimulation of noradrenaline release was reduced by inhibitors of classical and novel protein kinase C isozymes, but not by an inhibitor selective for Ca(2+)-dependent isoforms. These results demonstrate that bradykinin B(2) receptors are linked to phospholipase C to simultaneously activate two signaling pathways: one mediates an inositol trisphosphate- and Ca(2+)-dependent inhibition of M-currents, the other one leads to an excitation of sympathetic neurons independently of changes in M-currents through an activation of Ca(2+)-insensitive protein kinase C. PMID- 12122045 TI - Limbic seizures induce P-glycoprotein in rodent brain: functional implications for pharmacoresistance. AB - The causes and mechanisms underlying multidrug resistance (MDR) in epilepsy are still elusive and may depend on inadequate drug concentration in crucial brain areas. We studied whether limbic seizures or anticonvulsant drug treatments in rodents enhance the brain expression of the MDR gene (mdr) encoding a permeability glycoprotein (P-gp) involved in MDR to various cancer chemotherapeutic agents. We also investigated whether changes in P-gp levels affect anticonvulsant drug concentrations in the brain. Mdr mRNA measured by RT PCR increased by 85% on average in the mouse hippocampus 3-24 hr after kainic acid-induced limbic seizures, returning to control levels by 72 hr. Treatment with therapeutic doses of phenytoin or carbamazepine for 7 d did not change mdr mRNA expression in the mouse hippocampus 1-72 hr after the last drug administration. Six hours after seizures, the brain/plasma ratio of phenytoin was reduced by 30% and its extracellular concentration estimated by microdialysis was increased by twofold compared with control mice. Knock-out mice (mdr1a/b -/-) lacking P-gp protein showed a 46% increase in phenytoin concentrations in the hippocampus 1 and 4 hr after injection compared with wild-type mice. A significant 23% increase was found in the cerebellum at 1 hr and in the cortex at 4 hr. Carbamazepine concentrations were measurable in the hippocampus at 3 hr in mdr1a/b -/- mice, whereas they were undetectable at the same time interval in wild-type mice. In rats having spontaneous seizures 3 months after electrically induced status epilepticus, mdr1 mRNA levels were enhanced by 1.8-fold and fivefold on average in the hippocampus and entorhinal cortex, respectively. Thus, changes in P-gp mRNA levels occur in limbic areas after both acute and chronic epileptic activity. P-gp alterations significantly affect antiepileptic drugs concentrations in the brain, suggesting that seizure-induced mdr mRNA expression contributes to MDR in epilepsy. PMID- 12122046 TI - Presynaptic mitochondrial calcium sequestration influences transmission at mammalian central synapses. AB - Beyond their role in generating ATP, mitochondria have a high capacity to sequester calcium. The interdependence of these functions and limited access to presynaptic compartments makes it difficult to assess the role of sequestration in synaptic transmission. We addressed this important question using the calyx of Held as a model glutamatergic synapse by combining patch-clamp with a novel mitochondrial imaging method. Presynaptic calcium current, mitochondrial calcium concentration ([Ca(2+)](mito), measured using rhod-2 or rhod-FF), cytoplasmic calcium concentration ([Ca(2+)](cyto), measured using fura-FF), and the postsynaptic current were monitored during synaptic transmission. Presynaptic [Ca(2+)](cyto) rose to 8.5 +/- 1.1 microM and decayed rapidly with a time constant of 45 +/- 3 msec; presynaptic [Ca(2+)](mito) also rose rapidly to >5 microM but decayed slowly with a half-time of 1.5 +/- 0.4 sec. Mitochondrial depolarization with rotenone and carbonyl cyanide p trifluoromethoxyphenylhydrazone abolished mitochondrial calcium rises and slowed the removal of [Ca(2+)](cyto) by 239 +/- 22%. Using simultaneous presynaptic and postsynaptic patch clamp, combined with presynaptic mitochondrial and cytoplasmic imaging, we investigated the influence of mitochondrial calcium sequestration on transmitter release. Depletion of ATP to maintain mitochondrial membrane potential was blocked with oligomycin, and ATP was provided in the patch pipette. Mitochondrial depolarization raised [Ca(2+)](cyto) and reduced transmitter release after short EPSC trains (100 msec, 200 Hz); this effect was reversed by raising mobile calcium buffering with EGTA. Our results suggest a new role for presynaptic mitochondria in maintaining transmission by accelerating recovery from synaptic depression after periods of moderate activity. Without detectable thapsigargin-sensitive presynaptic calcium stores, we conclude that mitochondria are the major organelle regulating presynaptic calcium at central glutamatergic terminals. PMID- 12122047 TI - The relationship between intracellular free iron and cell injury in cultured neurons, astrocytes, and oligodendrocytes. AB - Iron is an essential element for cells but may also be an important cytotoxin. However, very little is known about iron transport, redox status, or toxicity specifically inside cells. In this study, we exploited the sensitivity of fura-2 to quenching by ferrous iron (Fe(2+)) to detect intracellular free iron ([Fe(2+)](i)) in neurons, astrocytes, and oligodendrocytes in primary culture. All cell types exposed to Fe(2+) in the presence of the ionophore pyrithione rapidly accumulated Fe(2+) to a similar extent. The heavy-metal chelators bipyridyl and N,N,N',N'-tetrakis(2-pyridalmethyl)ethyl-enediamine rapidly reversed the increase in [Fe(2+)](i), whereas desferrioxamine had little effect. Interestingly, the Fe(2+)-mediated quenching of fura-2 fluorescence was reversed in a concentration-dependent manner by hydrogen peroxide. This was likely caused by the oxidation of Fe(2+) to Fe(3+) inside the cell. Acute exposure of cells to Fe(2+) was only toxic when the metal was applied together with pyrithione, showing that Fe(2+) is only toxic when elevated inside cells. Interestingly, only neurons and oligodendrocytes were injured by this elevation in [Fe(2+)](i), whereas astrocytes were unaffected, although [Fe(2+)](i) was elevated to the same degree in each cell type. These studies provide a novel approach for detecting [Fe(2+)](i) in a manner sensitive to the redox state of the metal. These studies also provide a model system for the study of the toxic consequences of elevated [Fe(2+)](i) in neural cells. PMID- 12122048 TI - Synaptic calcium-channel function in Drosophila: analysis and transformation rescue of temperature-sensitive paralytic and lethal mutations of cacophony. AB - Voltage-gated calcium channels play a key role in chemical synaptic transmission by providing the calcium trigger for regulated neurotransmitter release. Genes encoding the primary structural subunit, alpha1, as well as accessory subunits of presynaptic calcium channels have now been identified in a variety of organisms. The cacophony (cac) gene in Drosophila, also known as nightblind A, encodes a voltage-gated calcium-channel alpha1 subunit homologous to vertebrate alpha1 subunits implicated in neurotransmitter release. A recent genetic screen in our laboratory isolated cac(TS2), a conditional cac mutant exhibiting rapid paralysis at elevated temperatures. This mutant has allowed synaptic electrophysiology after acute perturbation of a specific calcium-channel gene product, demonstrating that cac encodes a primary calcium channel functioning in neurotransmitter release. Here we report the molecular lesion in cac(TS2), a missense mutation within a calcium-dependent regulatory domain of the alpha1 subunit, as well as phenotypic rescue of temperature-sensitive and lethal cac mutations by transgenic expression of a wild-type cac cDNA. Notably, rescue of rapid, calcium-triggered neurotransmitter release was achieved by neural expression of a single cDNA containing a subset of alternative exons and lacking any conserved synaptic-protein interaction sequence. Possible implications of these findings are discussed in the context of structure-function studies of synaptic calcium channels, as well as alternative splicing and mRNA editing of the cac transcript. PMID- 12122049 TI - A dual role for the SDF-1/CXCR4 chemokine receptor system in adult brain: isoform selective regulation of SDF-1 expression modulates CXCR4-dependent neuronal plasticity and cerebral leukocyte recruitment after focal ischemia. AB - The chemoattractant stromal cell-derived factor-1 (SDF-1) and its receptor CXC chemokine receptor 4 (CXCR4) are key modulators of immune function. In the developing brain, SDF-1 is crucial for neuronal guidance; however, cerebral functions of SDF-1/CXCR4 in adulthood are unclear. Here, we examine the cellular expression of SDF-1 isoforms and CXCR4 in the brain of mice receiving systemic lipopolysaccharide (LPS) or permanent focal cerebral ischemia. CXCR4 mRNA was constitutively expressed in cortical and hippocampal neurons and ependymal cells. Hippocampal neurons targeted the CXCR4 receptor to their somatodendritic and axonal compartments. In cortex and hippocampus, CXCR4-expressing neurons exhibited an overlapping distribution with neurons expressing SDF-1 transcripts. Although neurons synthesized SDF-1alpha mRNA, the SDF-1beta isoform was selectively expressed by endothelial cells of cerebral microvessels. LPS stimulation dramatically decreased endothelial SDF-1beta mRNA expression throughout the forebrain but did not affect neuronal SDF-1alpha. After focal cerebral ischemia, SDF-1beta expression was selectively increased in endothelial cells of penumbral blood vessels and decreased in endothelial cells of nonlesioned brain areas. In the penumbra, SDF-1beta upregulation was associated with a concomitant infiltration of CXCR4-expressing peripheral blood cells, including macrophages. Neuronal SDF-1alpha was transiently downregulated and neuronal CXCR4 was transiently upregulated in the nonlesioned cerebral cortex in response to ischemia. Although endothelial SDF-1beta may control cerebral infiltration of CXCR4-carrying leukocytes during cerebral ischemia, the neuronal SDF-1alpha/CXCR4 system may contribute to ischemia-induced neuronal plasticity. Thus, the isoform-specific regulation of SDF-1 expression modulates neurotransmission and cerebral infiltration via distinct CXCR4-dependent pathways. PMID- 12122050 TI - Serial analysis of gene expression identifies metallothionein-II as major neuroprotective gene in mouse focal cerebral ischemia. AB - We applied serial analysis of gene expression (SAGE) to study differentially expressed genes in mouse brain 14 hr after the induction of focal cerebral ischemia. Analysis of >60,000 transcripts revealed 83 upregulated and 94 downregulated transcripts (more than or equal to eightfold). Reproducibility was demonstrated by performing SAGE in duplicate on the same starting material. Metallothionein-II (MT-II) was the most significantly upregulated transcript in the ischemic hemisphere. MT-I and MT-II are assumed to be induced by metals, glucocorticoids, and inflammatory signals in a coordinated manner, yet their function remains elusive. Upregulation of both MT-I and MT-II was confirmed by Northern blotting. MT-I and MT-II mRNA expression increased as early as 2 hr after 2 hr of transient ischemia, with a maximum after 16 hr. Western blotting and immunohistochemistry revealed MT-I/-II upregulation in the ischemic hemisphere, whereas double labeling demonstrated the colocalization of MT with markers for astrocytes as well as for monocytes/macrophages. MT-I- and MT-II deficient mice developed approximately threefold larger infarcts than wild-type mice and a significantly worse neurological outcome. For the first time we make available a comprehensive data set on brain ischemic gene expression and underscore the important protective role of metallothioneins in ischemic damage of the brain. Our results demonstrate the usefulness of SAGE to screen functionally relevant genes and the power of knock-out models in linking function to expression data generated by high throughput techniques. PMID- 12122051 TI - Ischemia induces a translocation of the splicing factor tra2-beta 1 and changes alternative splicing patterns in the brain. AB - Alternative splice-site selection is regulated by the relative concentration of individual members of the serine-arginine family of proteins and heterogeneous nuclear ribonucleoproteins. Most of these proteins accumulate predominantly in the nucleus, and a subset of them shuttles continuously between nucleus and cytosol. We demonstrate that in primary neuronal cultures, a rise in intracellular calcium concentration induced by thapsigargin leads to a translocation of the splicing regulatory protein tra2-beta1 and a consequent change in splice-site selection. To investigate this phenomenon under physiological conditions, we used an ischemia model. Ischemia induced in the brain causes a cytoplasmic accumulation and hyperphosphorylation of tra2-beta1. In addition, several of the proteins binding to tra2-beta1, such as src associated in mitosis 68 and serine/arginine-rich proteins, accumulate in the cytosol. Concomitant with this subcellular relocalization, we observed a change in alternative splice-site usage of the ICH-1 gene. The increased usage of its alternative exons is in agreement with previous studies demonstrating its repression by a high concentration of proteins with serine/arginine-rich domains. Our findings suggest that a change in the calcium concentration associated with ischemia is part of a signaling event, which changes pre-mRNA splicing pathways by causing relocalization of proteins that regulate splice-site selection. PMID- 12122052 TI - Inflammation-dependent cerebral deposition of serum amyloid a protein in a mouse model of amyloidosis. AB - The major pathological hallmark of amyloid diseases is the presence of extracellular amyloid deposits. Serum amyloid A (SAA) is an apolipoprotein primarily produced in the liver. Serum protein levels can increase one thousandfold after inflammation. SAA is the precursor to the amyloid A protein found in deposits of systemic amyloid A amyloid (AA or reactive amyloid) in both mouse and human. To study the factors necessary for cerebral amyloid formation, we have created a transgenic mouse that expresses the amyloidogenic mouse Saa1 protein in the brain. Using the synapsin promoter to drive expression of the Saa1 gene, the brains of transgenic mice expressed both RNA and protein. Under noninflammatory conditions, transgenic mice do not develop AA amyloid deposits in the brain; however, induction of a systemic acute-phase response in transgenic mice enhanced amyloid deposition. This deposition was preceded by an increase in cytokine levels in the brain, suggesting that systemic inflammation may be a contributing factor to the development of cerebral amyloid. The nonsteroidal anti inflammatory agent indomethacin reduced inflammation and protected against the deposition of AA amyloid in the brain. These studies indicate that inflammation plays an important role in the process of amyloid deposition, and inhibition of inflammatory cascades may attenuate amyloidogenic processes, such as Alzheimer's disease. PMID- 12122053 TI - Interleukin-18 involvement in hypoxic-ischemic brain injury. AB - Inflammation is a critical factor for development of hypoxic-ischemic (HI) brain injury. Interleukin-18 (IL-18) is a proinflammatory cytokine expressed in microglia and processed by caspase-1. Our aim was to characterize the expression of IL-18 and its receptor in relation to caspase-1 and IL-1beta after HI and to evaluate to what extent IL-18 contributes to HI brain injury. Seven-day-old rats were subjected to HI, and brain tissue was sampled at different time points (3 hr to 14 d) after insult. The mRNA for IL-18 and caspase-1 were analyzed with reverse transcriptase PCR, protein was analyzed by Western blot (IL-18, caspase 1) or ELISA (IL-1beta), and the regional distribution was assessed by immunohistochemistry. HI was also induced in C57BL/6 mice, and brain injury in IL 18-deficient animals was compared with that in wild-type animals. The expression of mRNA/protein for caspase-1 and IL-18 in brain homogenates increased progressively at 12 hr to 14 d after HI, whereas IL-1beta peaked at 8 hr. A widespread expression of caspase-1 and IL-18 protein in microglia was found in the HI hemisphere. The IL-18 receptor was expressed on neurons of the cerebral cortex and thalamus. IL-1beta was primarily found in microglia in the habenular nucleus of the thalamus. The infarct volume was reduced by 21% (p = 0.01), and the neuropathology score was significantly decreased in the cerebral cortex ( 35%), hippocampus (-22%), striatum (-18%), and thalamus (-17%) in mice with IL-18 deficiency compared with wild-type mice. In conclusion, we found that IL-18 expression in microglia was markedly increased after HI and that IL-18 appears to be important for the development of HI brain injury. PMID- 12122054 TI - Neurofilament-M interacts with the D1 dopamine receptor to regulate cell surface expression and desensitization. AB - We used the yeast two-hybrid assay to identify novel proteins that interact with the D(1) dopamine receptor. The third cytoplasmic loop (residues 217-273) of the rat D(1) receptor was used as bait to identify clones encoding interacting proteins from a rat brain cDNA library. This identified two clones encoding the C terminus of rat neurofilament-M (NF-M) (residues 782-846). The NF-M clone did not interact with the third cytoplasmic loops of the rat D(2), D(3), or D(4) receptors, but showed weak interaction with that of the D(5) receptor. Coexpression of full-length NF-M with the D(1) receptor in HEK-293 cells resulted in >50% reduction of receptor binding accompanied by a reduction in D(1) receptor mediated cAMP accumulation. NF-M had no effect on the expression of other dopamine receptor subtypes. Using a D(1) receptor-green fluorescent protein chimera and confocal fluorescence microscopy, we found that NF-M reduced D(1) receptor expression at the cell surface and promoted accumulation of the receptor in the cytosol. Interestingly, the D(1) receptors that were expressed at the cell surface in the presence of NF-M were resistant to agonist-induced desensitization. Cellular colocalization of NF-M and the D(1) receptor in the rat brain was examined by epifluorescence microscopy. These experiments showed that approximately 50% of medium-sized striatal neurons expressed both proteins. Colocalization was also observed in pyramidal cells and interneurons within the frontal cortex. Similar immunohistochemical analyses using NF-M-deficient mice showed decrements in D(1) receptor expression compared with control mice. These results suggest that NF-M interacts with the D(1) receptor in vivo and may modify its expression and regulation. PMID- 12122055 TI - Stomatin-related olfactory protein, SRO, specifically expressed in the murine olfactory sensory neurons. AB - We identified a stomatin-related olfactory protein (SRO) that is specifically expressed in olfactory sensory neurons (OSNs). The mouse sro gene encodes a polypeptide of 287 amino acids with a calculated molecular weight of 32 kDa. SRO shares 82% sequence similarity with the murine stomatin, 78% with Caenorhabditis elegans MEC-2, and 77% with C. elegans UNC-1. Unlike other stomatin-family genes, the sro transcript was present only in OSNs of the main olfactory epithelium. No sro expression was seen in vomeronasal neurons. SRO was abundant in most apical dendrites of OSNs, including olfactory cilia. Immunoprecipitation revealed that SRO associates with adenylyl cyclase type III and caveolin-1 in the low-density membrane fraction of olfactory cilia. Furthermore, anti-SRO antibodies stimulated cAMP production in fractionated cilia membrane. SRO may play a crucial role in modulating odorant signals in the lipid rafts of olfactory cilia. PMID- 12122056 TI - Activation of presynaptic P2X7-like receptors depresses mossy fiber-CA3 synaptic transmission through p38 mitogen-activated protein kinase. AB - P2X(7) receptor subunits form homomeric ATP-gated, calcium-permeable cation channels. In this study, we used Western blots and immunocytochemistry to demonstrate that P2X(7) receptors are abundant on presynaptic terminals of mossy fiber synapses in the rat hippocampus. P2X(7)-immunoreactive protein was detected using a specific P2X(7) antibody in Western blots of protein isolated from whole hippocampus and from a subcellular fraction containing mossy fiber synaptosomes. P2X(7) immunoreactivity was colocalized with syntaxin 1A/B-immunoreactivity in mossy fiber terminals in the dentate hilus and stratum lucidum of CA3. Extracellular and whole-cell voltage-clamp recordings in CA3 revealed that bath application of the potent P2X(7) agonist 2',3'-O-(4-benzoylbenzoyl)-ATP (Bz-ATP) caused a long-lasting inhibition of neurotransmission at mossy fiber-CA3 synapses. Consistent with a presynaptic action at mossy fiber synapses, Bz-ATP had no significant effect on neurotransmission at associational-commissural synapses in CA3 but increased paired-pulse facilitation during depression of mossy fiber evoked currents. In addition, Bz-ATP had no postsynaptic effect on holding current or conductance of CA3 neurons. Bz-ATP-induced mossy fiber synaptic depression was blocked by the P2X(7) antagonist oxidized ATP but not by the P2X(1-3,5,6) antagonist pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid or the P2Y antagonist reactive blue 2. Finally, an antagonist of p38 MAP kinase activation [4-(4-fluorophenyl)2-(4-methylsulfinylphenyl)5-(4-pyridyl)imidazole] but not extracellular signal-regulated kinase 1/2 MAP kinase (2'-amino-3' methoxyflavone) blocked the synaptic depression mediated by Bz-ATP, suggesting that this presynaptic inhibition was mediated by activation of p38 MAP kinase. The results of the present study demonstrate that activation of presynaptic P2X(7) receptors depresses mossy fiber-CA3 synaptic transmission through activation of p38 MAP kinase. PMID- 12122057 TI - Sequential nuclear accumulation of the clock proteins period and timeless in the pacemaker neurons of Drosophila melanogaster. AB - Antisera against the circadian clock proteins Period (PER) and Timeless (TIM) were used to construct a detailed time course of PER and TIM expression and subcellular localization in a subset of the ventrolateral neurons (vLNs) in the Drosophila accessory medulla (AMe). These neurons, which express pigment dispersing factor, play a central role in the control of behavioral rhythms. The data revealed several unexpected features of the circadian clock in Drosophila. First, TIM but not PER was restricted to the cytoplasm of vLNs throughout most of the early night. Second, the timing of TIM and PER nuclear accumulation was substantially different. Third, the two subsets of vLNs, the large and small vLNs, had a similar timing of PER nuclear accumulation but differed by 3-4 hr in the phase of TIM nuclear accumulation. These aspects of PER and TIM expression were not predicted by the current mechanistic model of the circadian clock in Drosophila and are inconsistent with the hypothesis that PER and TIM function as obligate heterodimers. The differing profiles of TIM and PER nuclear accumulation suggest that PER and TIM have distinct functions in the nuclei of vLNs. PMID- 12122058 TI - Mapping the binding site of the neuroprotectant ifenprodil on NMDA receptors. AB - Ifenprodil is a noncompetitive antagonist of NMDA receptors highly selective for the NMDA receptor 2B (NR2B) subunit. It is widely used as a pharmacological tool to discriminate subpopulations of NMDA receptors, and derivatives are currently being developed as candidate neuroprotectants. Despite numerous studies on the mechanism of action of ifenprodil on NMDA receptors, the structural determinants responsible for the subunit selectivity have not been identified. By combining functional studies on recombinant NMDA receptors and biochemical studies on isolated domains, we now show that ifenprodil binds to the N-terminal leucine/isoleucine/valine-binding protein (LIVBP)-like domain of NR2B. In this domain, several residues, both hydrophilic and hydrophobic, were found to control ifenprodil inhibition. Their location in a modeled three-dimensional structure suggests that ifenprodil binds in the cleft of the LIVBP-like domain of NR2B by a mechanism (Venus-flytrap) resembling that of the binding of Zn on the LIVBP-like domain of NR2A. These results reinforce the proposal that the LIVBP-like domains of NMDA receptors, and possibly of other ionotropic glutamate receptors, bind modulatory ligands. Moreover, they identify the LIVBP-like domain of the NR2B subunit as a promising therapeutic target and provide a framework for designing structurally novel NR2B-selective antagonists. PMID- 12122059 TI - Intrinsic program for migration of cerebellar granule cells in vitro. AB - Cerebellar granule cells exhibit distinct modes of migration in different cortical layers. The role of external cues in controlling these alterations has been suggested, but the significance of internal programs is not well understood. In the present study, we examined autonomous changes of migratory behavior of isolated granule cells in microexplant cultures of the postnatal mouse cerebellum. We found that isolated granule cells sequentially go through three characteristic phases of migration without cell-cell contact. In the first phase (0-20 hr in vitro) granule cells exhibit the highest rate of turning behavior and have multiple short processes. The length of the movement cycle is shortest. In the second phase (20-40 hr in vitro), granule cells extend a long and thick process and exhibit an elongated cycle of movement. Their speed is fastest, whereas the rate of turning is lowest. In the third phase (40-60 hr in vitro), granule cells slow down their movement and slightly increase their turnings. The length of the movement cycle further increases. At the end, the cells become permanently stationary, extend a lamellipodium around the soma, and emit several thin processes. Interestingly, granule cells sequentially develop four different modes of turning. These results indicate that internal (intrinsic) programs control alterations of granule cell behavior in a stage-dependent manner, suggesting that such programs independent of local cell-cell contacts may be essential for granule cell translocation in the developing cerebellum. PMID- 12122060 TI - Microtubule reconfiguration during axonal retraction induced by nitric oxide. AB - Axonal retraction is induced by different types of physiological cues and is responsible for the elimination of mistargeted axons. There is broad agreement that alterations in the cytoskeleton underlie axonal retraction. The prevailing view is that axonal retraction involves a wholesale depolymerization of microtubules and microfilaments. However, axons retracting physiologically display a very different morphology than axons induced to retract by experimental depolymerization of microtubules. Experimental depolymerization of microfilaments actually prevents retraction rather than causing it. We have proposed an alternative hypothesis, namely that axonal retraction involves a backward retreat of cytoskeletal elements rather than their wholesale depolymerization. In the present study, we sought to test this hypothesis with regard to microtubules. When a donor of nitric oxide was applied to cultured chick sensory neurons, the majority of axons retracted dramatically within 30-60 min. Retracting axons were characterized by an enlarged distal region, a thin trailing remnant, and sinusoidal bends along the shaft. Quantitative immunofluorescence analyses showed no detectable loss of microtubule mass during retraction, even with regard to the most labile microtubules. Instead, microtubules were reconfigured into coiling and sinusoidal bundles to accommodate the shortening of the axon. Stabilization of microtubules by taxol did not prevent the retraction, even at concentrations of the drug that actually caused microtubule levels to increase. The retractions induced by nitric oxide were remarkably similar to those observed when motor proteins are manipulated, suggesting that these retractions may result from alterations in the activities of the motors that configure microtubules. PMID- 12122062 TI - The projections of early enteric neurons are influenced by the direction of neural crest cell migration. AB - The enteric nervous system arises from the neural crest. In embryonic mice, vagal neural crest cells enter the developing foregut at approximately embryonic day 9.5 (E9.5) and then migrate rostrocaudally to colonize the entire gastrointestinal tract by E14.5. This study showed that a subpopulation of vagal crest-derived cells, very close to the migratory wavefront, starts to differentiate into neurons early, as shown by the expression of neuron-specific proteins and the absence of Sox10. Many of the early differentiating neurons transiently exhibited tyrosine hydroxylase (TH) immunoreactivity. The TH cells were demonstrated to be the progenitors of nitric oxide synthase (NOS) neurons. Immunohistochemistry, lesions, and DiI tracing were used to examine the projections of developing enteric neurons. The axons of first neurons in the gut (the TH-NOS neurons) projected in the same direction (caudally), and traversed the same pathways through the mesenchyme, as the migrating, undifferentiated, vagal crest-derived cells. To examine if the direction of migration and direction of axon projection are linked, coculture experiments were set up in which vagal crest-derived cells migrated either rostrocaudally (as they do in vivo), or caudorostrally (which they do not normally do), to colonize explants of embryonic aneural hindgut. The direction in which neurons projected was correlated with the direction of cell migration, but migration direction appears to be not the only mechanism influencing axon projection. Peristaltic reflexes involve both orally (rostrally) projecting neurons and anally (caudally) projecting neurons. Because few rostrally projecting neurons could be detected before birth, the full circuitry for peristaltic reflexes appears to develop after birth. PMID- 12122061 TI - Directional guidance of oligodendroglial migration by class 3 semaphorins and netrin-1. AB - Oligodendrocytes, the myelin-forming cells of the CNS, are generated from multiple foci distributed along the developing neural tube. Little is known about the endogenous guidance cues controlling the migration of oligodendrocyte precursor cells (OPCs) from their site of emergence toward their final destination, mainly the future white matter tracts. During embryonic development, the optic nerve is populated by OPCs originating in the diencephalon that migrate from the chiasm toward the retina. Here we show that OPCs migrating into the embryonic optic nerve express the semaphorin receptors neuropilin-1 and -2, as well as deleted in colorectal cancer (DCC) and, to a lesser extend unc5H1, two of the netrin-1 receptors. Using a functional migration assay, we provide evidence that Sema 3A and netrin-1 exert opposite chemotactic effects, repulsive or attractive, respectively, on embryonic OPCs. In addition, we show that Sema 3F has a dual effect, chemoattractive and mitogenic on embryonic OPCs. The localization of cells expressing Sema 3A, Sema 3F, and netrin-1 is consistent with a role for these ligands in the migration of OPCs in the embryonic optic nerve. Altogether, our results suggest that the migration of OPCs in the embryonic optic nerve is modulated by a balance of effects mediated by members of the semaphorin and netrin families. PMID- 12122063 TI - Rac1-mediated endocytosis during ephrin-A2- and semaphorin 3A-induced growth cone collapse. AB - Negative guidance molecules are important for guiding the growth of axons and ultimately for determining the wiring pattern in the developing nervous system. In tissue culture, growth cones at the tips of growing axons collapse in response to negative guidance molecules, such as ephrin-A2 and semaphorin 3A. The small GTPase Rac1 is involved in growth cone collapse, but the nature of its role is not clear. Rac1 activity assays showed that Rac1 is transiently inactivated after treatment with ephrin-A2. Ephrin-induced growth cone collapse, however, correlated with resumption of Rac1 activity. We demonstrate that Rac1 is required for endocytosis of the growth cone plasma membrane and reorganization of F-actin but not for the depolymerization of F-actin during growth cone collapse in response to ephrin-A2 and semaphorin 3A. Rac1, however, does not regulate constitutive endocytosis in growth cones. These findings show that in response to negative guidance molecules, the function of Rac1 changes from promoting actin polymerization associated with axon growth to driving endocytosis of the plasma membrane, resulting in growth cone collapse. Furthermore, Rac1 antisense injected into the embryonic chick eye in vivo caused the retinotectal projection to develop without normal topography in a manner consistent with Rac1 having an obligatory role in mediating ephrin signaling. PMID- 12122064 TI - A critical function of the pial basement membrane in cortical histogenesis. AB - Mice with a targeted deletion of the nidogen-binding site of laminin gamma1 were used to study the function of the pial basement membrane in cortical histogenesis. The pial basement membrane in the mutant embryos assembled but was unstable and disintegrated at random segments. In segments with a disrupted basement membrane, radial glia cells were retracted from the pial surface, and radially migrating neurons, including Cajal-Retzius cells and cortical plate neurons, passed the meninges or terminated their migration prematurely. By correlating the disruptions in the pial basal lamina with changes in the morphology of radial glia cells, the aberrant migration of Cajal-Retzius cells, and subsequent dysplasia of cortical plate neurons, the present data establish a causal relationship of proper cortical histogenesis with the presence of an intact pial basement membrane. PMID- 12122065 TI - Myelination is altered in insulin-like growth factor-I null mutant mice. AB - Increasing evidence indicates that insulin-like growth factor-I (IGF-I) has an important role in oligodendrocyte development. In this study, we examined myelination during postnatal development in IGF-I knock-out (KO) mice by assessing myelin staining, the expression of myelin basic protein (MBP) and proteolipid protein (PLP), two major myelin-specific proteins, and the number of oligodendrocytes and their precursors. For comparison, we also measured the expression of median subunit of the neuron-specific intermediate filament, M neurofilament (M-NF), to obtain an index of the effects of IGF-I deficiency on neurons. We found that myelin staining, MBP and PLP expression, and the percentage of oligodendrocytes and their precursors are significantly reduced in all brain regions of developing IGF-I KO mice but are similar to controls in adult IGF-I KO mice. In contrast, the abundance of M-NF was decreased in both the developing and adult brain of IGF-I KO mice. We also found that IGF-II protein abundance is increased in the brains of IGF-I KO mice. Our data indicate, therefore, that myelination during early development is altered in the absence of IGF-I by mechanisms that involve a reduction in oligodendrocyte proliferation and development. Although neuronal actions cannot be excluded in the myelin normalization, the reduced axonal growth suggested by the reduced M-NF expression makes a role for neuronal factors less compelling. These data suggest that IGF-I plays a significant role in myelination during normal early development and that IGF-II can compensate in part for IGF-I actions on myelination. PMID- 12122066 TI - Relations between brain pathology and temporal lobe epilepsy. AB - Temporal lobe epilepsy, the most common type of epilepsy in adult humans, is characterized clinically by the progressive development of spontaneous recurrent seizures of temporal lobe origin and pathologically by hippocampal neuronal loss and mossy fiber sprouting. In this study, we sought to test the prominent hypothesis that neuronal loss and mossy fiber sprouting play a critical role in the genesis and progression of temporal lobe epilepsy. Rats receiving a single kainic acid injection experienced a single sustained episode of epileptic status with massive neuronal loss and mossy fiber sprouting, whereas rats receiving triple kainic acid injections experienced two priming episodes and one sustained episode of epileptic status with no detectable neuronal loss and mossy fiber sprouting. Early in the process of chronic seizure development, primed rats that failed to show detectable neuronal loss and mossy fiber sprouting exhibited a starting date and a frequency of spontaneous recurrent seizures similar to those of nonprimed rats that showed massive neuronal loss and mossy fiber sprouting. However, nonprimed rats displayed significantly prolonged episodes of spontaneous recurrent seizures over the whole process of chronic seizure development and more frequent severe seizures later in the process. Similar results were observed in both Fischer-344 and Wistar rats as well as in the rat pilocarpine preparation of temporal lobe epilepsy. These results fail to reveal a relation between neuronal loss-mossy fiber sprouting and the genesis of temporal lobe epilepsy but suggest that neuronal loss, mossy fiber sprouting, or both contribute to the intensification of chronic seizures. PMID- 12122067 TI - Synchronous neuronal activity is a signal for axonal sprouting after cortical lesions in the adult. AB - The ability of the adult brain to form new connections in areas denervated by a lesion (axonal sprouting) is more widespread than previously thought, but mechanisms remain unknown. We have previously demonstrated an unexpected, robust axonal sprouting of contralateral corticostriatal neurons into the denervated striatum after ischemic cortical lesions. We now take advantage of marked differences in the degree of axonal sprouting from contralateral homotypic cortex after two types of cortical lesions to define the role of neuronal activity in this response. Thermal-ischemic lesions (TCL) of sensorimotor cortex, which induce axonal sprouting, produced two sequential patterns of low-frequency, synchronized neuronal activity that are not seen after similarly sized aspiration lesions, which do not induce axonal sprouting. An early rhythm of synchronous neuronal activity occurred in perilesion cortex on day 1 after lesion, with a frequency range of 0.2-2 Hz. A later pattern of activity occurred on days 2 and 3 after lesion, with a frequency range of 0.1-0.4 Hz. This second rhythm synchronized neuronal activity across widespread areas, including the cortical areas that contain the cell bodies of the sprouting axons. TTX was used to block this patterned neuronal activity and determine whether axonal sprouting was prevented. Chronic TTX infusion into the lesion site blocked the synchronous neuronal activity after TCL as well as axonal sprouting. Thus, both after different types of lesions and in the blockade experiments axonal sprouting was strongly correlated with synchronous neuronal activity, suggesting a role for this activity in anatomical reorganization after brain lesion in the adult. PMID- 12122068 TI - The type 1 interleukin-1 receptor is essential for the efficient activation of microglia and the induction of multiple proinflammatory mediators in response to brain injury. AB - Interleukin-1 (IL-1) is induced immediately after insults to the brain, and elevated levels of IL-1 have been strongly implicated in the neurodegeneration that accompanies stroke, Alzheimer's disease, and multiple sclerosis. In animal models, antagonizing IL-1 has been shown to reduce cell death; however, the basis for this protection has not been elucidated. Here we analyzed the response to penetrating brain injury in mice lacking the type 1 IL-1 receptor (IL-1R1) to determine which cellular and molecular mediators of tissue damage require IL-1 signaling. At the cellular level, fewer amoeboid microglia/macrophages appeared adjacent to the injured brain tissue in IL-1R1 null mice, and those microglia present at early postinjury intervals retained their resting morphology. Astrogliosis also was mildly abrogated. At the molecular level, cyclooxygenase-2 (Cox-2) and IL-6 expression were depressed and delayed. Interestingly, basal levels of Cox-2, IL-1, and IL-6 were significantly lower in the IL-1R1 null mice. In addition, stimulation of vascular cell adhesion molecule-1 mRNA was depressed in the IL-1R1 null mice, and correspondingly, there was reduced diapedesis of peripheral macrophages in the IL-1R1 null brain after injury. This observation correlated with a reduced number of Cox-2+ amoeboid phagocytes adjacent to the injury. In contrast, several molecular aspects of the injury response were normal, including expression of tumor necrosis factor-alpha and the production of nerve growth factor. Because antagonizing IL-1 protects neural cells in experimental models of stroke and multiple sclerosis, our data suggest that cell preservation is achieved by abrogating microglial/macrophage activation and the subsequent self-propagating cycle of inflammation. PMID- 12122069 TI - Purified adult ensheathing glia fail to myelinate axons under culture conditions that enable Schwann cells to form myelin. AB - Several studies have suggested that olfactory ensheathing glia (EG) can form Schwann cell (SC)-like myelin. Because of possible misinterpretation attributable to contaminating SCs, the capacity of EG to produce myelin needs to be explored further. Therefore, we compared the abilities of adult EG, purified by immunopanning with p75 antibody, and adult SCs to produce myelin when cocultured with purified dorsal root ganglion neurons (DRGNs) in serum-free and serum containing media. In both media formulations, the number of myelin sheaths in SC/DRGN cultures was far higher than in EG/DRGN cultures; the number of sheaths in EG/DRGN cultures was equal to that in purified DRGN cultures without added cells. The latter result demonstrates that myelination by a few SCs remaining in purified DRGN cultures may occur, suggesting that myelin in EG/DRGN cultures could be SC myelin. Striking differences in the relationship of EG and SC processes to axons were observed. Whereas SCs displayed relatively short, thick processes that engulfed axons in small bundles or in individual cytoplasmic furrows and segregated larger axons into one-to-one relationships, EG extended flattened sheets that partitioned or only partially encircled fascicles of axons, sometimes spanning the entire culture. SCs exhibited behavior typical of SCs in peripheral nerves, whereas EG exhibited characteristics resembling those of EG in olfactory nerves. In sum, p75-selected EG from adult animals did not exhibit an SC-like relationship to axons and did not form myelin. PMID- 12122070 TI - D1 dopamine receptor regulation of microtubule-associated protein-2 phosphorylation in developing cerebral cortical neurons. AB - This study addresses the hypothesis that the previously described capacity of D1 dopamine receptors (D1Rs) to regulate dendritic growth in developing cortical neurons may involve alterations in the phosphorylation state of microtubule associated protein-2 (MAP2). The changes in phosphorylation of this protein are known to affect its ability to stabilize the dendritic cytoskeleton. The study involved two systems: primary cultures of mouse cortical neurons grown in the presence of the D1R agonists, SKF82958 or A77636, and the cortex of neonatal transgenic mice overexpressing the D1A subtype of D1R. In both models, a decrease in dendritic extension corresponded with an elevation in MAP2 phosphorylation. This phosphorylation occurred on all three amino acid residues examined in this study: serine, threonine, and tyrosine. In cultured cortical neurons, D1R stimulation-induced increase in MAP2 phosphorylation was blocked by the protein kinase A (PKA) inhibitor, H-89, and mimicked by the PKA activator, S(p)-cAMPS. This indicates that D1Rs modulate MAP2 phosphorylation through PKA-associated intracellular signaling pathways. We also observed that the elevations in MAP2 phosphorylation in neuronal cultures in the presence of D1R agonists (or S(p) cAMPS) were maintained for a prolonged time (up to at least 96 hr). Moreover, MAP2 phosphorylation underwent a substantial increase between 24 and 72 hr of exposure to these drugs. Our findings are consistent with the idea that D1Rs can modulate growth and maintenance of dendrites in developing cortical cells by regulating the phosphorylation of MAP2. In addition, our observations suggest that MAP2 phosphorylation by long-term activation of D1Rs (and PKA) can be divided into two phases: the initial approximately 24-hr-long phase of a relatively weak elevation in phosphorylation and the delayed phase of a much more robust phosphorylation increase taking place during the next approximately 48 hr. PMID- 12122071 TI - Maturation and death of adult-born olfactory bulb granule neurons: role of olfaction. AB - Young neurons born in the subventricular zone (SVZ) of adult mice migrate to the olfactory bulb (OB) where they differentiate into granule cells (GCs) and periglomerular interneurons. Using retroviral labeling of precursors in the SVZ, we describe five stages and the timing for the maturation of newly formed GCs: (1) tangentially migrating neuroblasts (days 2-7); (2) radially migrating young neurons (days 5-7); (3) GCs with a simple unbranched dendrite that does not extend beyond the mitral cell layer (days 9-13); (4) GCs with a nonspiny branched dendrite in the external plexiform layer (days 11-22); and (5) mature GCs (days 15-30). Using [3H]thymidine, we show that the maximum number of labeled GCs is observed around day 15 after injection. Interestingly, between days 15 and 45 after birth, soon after the cells developed spines, the number of [3H]thymidine labeled GCs declined by 50%. Using anosmic mice, we found that sensory input was critical for the survival of GCs from day 15 to 45 after labeling. However, the number and morphology of 15-d-old cells in the granule cell layer was similar in anosmic and wild-type mice. We infer that the lack of activity did not have an effect on the generation, migration, and early differentiation of granule cells. Soon after young GCs matured, and presumably became synaptically connected, their survival depended on the level of activity that they received. This selection mechanism might allow the construction of specific OB circuits based on olfactory experience and suggests possible functions of OB cell replacement. PMID- 12122072 TI - Diets enriched in foods with high antioxidant activity reverse age-induced decreases in cerebellar beta-adrenergic function and increases in proinflammatory cytokines. AB - Antioxidants and diets supplemented with foods high in oxygen radical absorbance capacity (ORAC) reverse age-related decreases in cerebellar beta-adrenergic receptor function. We examined whether this effect was related to the antioxidant capacity of the food supplement and whether an antioxidant-rich diet reduced the levels of proinflammatory cytokines in the cerebellum. Aged male Fischer 344 rats were given apple (5 mg dry weight), spirulina (5 mg), or cucumber (5 mg) either in 0.5 ml water by oral gavage or supplied in the rat chow daily for 14 d. Electrophysiologic techniques revealed a significant decrease in beta-adrenergic receptor function in aged control rats. Spirulina reversed this effect. Apple (a food with intermediate ORAC) had an intermediate effect on cerebellar beta adrenergic receptor physiology, and cucumber (low ORAC) had no effect, indicating that the reversal of beta-adrenergic receptor function decreases might be related to the ORAC dose. The mRNA of the proinflammatory cytokines tumor necrosis factor alpha (TNFalpha) and TNFbeta was also examined. RNase protection assays revealed increased levels of these cytokines in the aged cerebellum. Spirulina and apple significantly downregulated this age-related increase in proinflammatory cytokines, whereas cucumber had no effect, suggesting that one mechanism by which these diets work is by modulation of an age-related increase in inflammatory responses. Malondialdehyde (MDA) was measured as a marker of oxidative damage. Apple and spirulina but not cucumber decreased MDA levels in the aged rats. In summary, the improved beta-adrenergic receptor function in aged rats induced by diets rich in antioxidants is related to the ORAC dose, and these diets reduce proinflammatory cytokine levels. PMID- 12122074 TI - Detection and discrimination of relative spatial phase by V1 neurons. AB - Edge-like and line-like features result from spatial phase congruence, the local phase agreement between harmonic components of a spatial waveform. Psychophysical observations and models of early visual processing suggest that human visual feature detectors are specialized for edge-like and line-like phase congruence. To test whether primary visual cortex (V1) neurons account for such specificity, we made tetrode recordings in anesthetized macaque monkeys. Stimuli were drifting equal-energy compound gratings composed of four sinusoidal components. Eight congruence phases (one-dimensional features) were tested, including line-like and edge-like waveforms. Many of the 137 single V1 neurons (recorded at 45 sites) could reliably signal phase congruence by any of several response measures. Across neurons, the preferred spatial feature had only a modest bias for line like waveforms. Information-theoretic analysis showed that congruence phase was temporally encoded in the frequency band present in the stimuli. The most sensitive neurons had feature discrimination thresholds that approached psychophysical levels, but typical neurons were substantially less sensitive. In single V1 neurons, feature discrimination exhibited various dependences on the congruence phase of the reference waveform. Simple cells were over-represented among the most sensitive neurons and on average carried twice as much feature information as complex cells. However, the distribution of the indices of optimal tuning and discrimination of relative phase was indistinguishable in simple and complex cells. Our results suggest that phase-sensitive pooling of responses is required to account for human psychophysical performance, although variation in feature selectivity among nearby neurons is considerable. PMID- 12122073 TI - Group I metabotropic glutamate receptor (mGluR)-dependent long-term depression mediated via p38 mitogen-activated protein kinase is inhibited by previous high frequency stimulation and activation of mGluRs and protein kinase C in the rat dentate gyrus in vitro. AB - The induction of synaptic plasticity is known to be influenced by the previous history of the synapse, a process termed metaplasticity. Here we demonstrate a novel metaplasticity in which group I metabotropic glutamate receptor (mGluR) dependent long-term depression (LTD) of synaptic transmission is regulated by previous mGluR activation. In these studies, the group I mGluR-dependent LTD induced by the selective agonist (RS)-3,5-dihydroxyphenylglycine (DHPG-LTD) was inhibited by previous preconditioning brief high-frequency stimulation (HFS), regardless of whether the preconditioning HFS induced long-term potentiation. Blockade of NMDA receptors during the preconditioning HFS did not alter the inhibition of DHPG-LTD by the HFS. However, antagonism of mGluRs during the preconditioning HFS did prevent the inhibition of DHPG-LTD by the HFS. In addition, blocking PKC stimulation during the preconditioning HFS also prevented the inhibitory effect of HFS on DHPG-LTD. The DHPG-LTD itself was not inhibited by blocking PKC stimulation but was inhibited by blocking the p38 mitogen activated protein kinase (MAPK) pathway. Thus, whereas the DHPG-LTD is mediated via activation of the p38 MAPK pathway, the inhibitory effects of preconditioning HFS on DHPG-LTD are mediated via stimulation of group I/II mGluRs, activation of PKC, and subsequent blocking of the functioning of group I mGluR. PMID- 12122075 TI - Space and time maps of cone photoreceptor signals in macaque lateral geniculate nucleus. AB - We studied neurons in the central visual field representation of the lateral geniculate nucleus (LGN) in macaque monkeys by mapping their receptive fields in space and time. The mapping was performed by reverse correlation of a spike train of a neuron with pseudorandom, binary level stimuli (m-sequence grids). Black and white m-sequence grids were used to map the receptive field for luminance. The locations of receptive field center and surround were determined from this luminance map. To map the contribution of each cone class to the receptive field, we designed red-green or blue-yellow m-sequence grids to isolate the influence of that cone (long, middle, or short wavelength-sensitive: L, M, or S). Magnocellular neurons generally received synergistic input from L and M cones in both the center and the surround. A minority had cone-antagonistic (M-L) input to the surround. Red-green opponent parvocellular neurons received opponent cone input (L+M- or M+L-) that overlapped in space, as sampled by our stimulus grid, but that had somewhat different extents. For example, an L+ center parvocellular neuron would be L+/M- in both center and surround, but the L+ signal would be stronger in the center and the M- signal stronger in the surround. Accordingly, the luminance receptive field would be spatially antagonistic: on-center/off surround. The space-time maps also characterized LGN dynamics. For example, magnocellular responses were transient, red-green parvocellular responses were more sustained, and blue-on responses were the most sustained for both luminance and cone-isolating stimuli. For all cell types the surround response peaked 8-10 msec later than the center response. PMID- 12122077 TI - Dendroarchitecture of relay cells in thalamic barreloids: a substrate for cross whisker modulation. AB - A double-labeling protocol was used to determine how the dendroarchitecture of relay cells relates to the three-dimensional structure of barreloids in the ventral posterior medial nucleus of the rat thalamus. Single barreloids were retrogradely labeled by injecting Fluoro-Gold in identified barrel columns, and single relay cells activated by the same whisker, or by an adjacent whisker located on the same arc, were juxtacellularly labeled with biotinylated dextran. Results show that the dendritic field of relay cells is asymmetric, variously oriented with respect to the geometry of the barreloids, and that all cells extend dendrites in surrounding barreloids. Extrabarreloid dendrites are of small size (<1.5 microm) and represent up to 54% (range, 11-54%) of the total dendritic length. In contrast, the thick proximal dendrites remain confined to the home barreloid of the cell, being directed toward its center or along its margin. There is a trend for cells located dorsally in barreloids to form more elaborate trees with a larger proportion of extrabarreloid dendrites. Electron microscopic examination of labeled cells shows that extrabarreloid dendrites are exclusively contacted by synaptic terminals of cortical and reticular thalamic origin, whereas intrabarreloid dendrites also receive contacts from lemniscal terminals. Because corticothalamic and reticular thalamic cells establish point-to-point connections with homotopic barreloids, it is proposed that the spatial arrangement of dendrites determines the combination of whisker deflection that best modulates cell firing. Because relay cell responses are direction sensitive, maximal modulation would occur if dendritic field orientation relates to the direction selectivity of responses. PMID- 12122076 TI - Metabotropic glutamate 2 receptors modulate synaptic inputs and calcium signals in striatal cholinergic interneurons. AB - Striatal cholinergic interneurons were recorded from a rat slice preparation. Synaptic potentials evoked by intrastriatal stimulation revealed three distinct components: a glutamatergic EPSP, a GABA(A)-mediated depolarizing potential, and an acetylcholine (ACh)-mediated IPSP. The responses to group II metabotropic glutamate (mGlu) receptor activation were investigated on the isolated components of the synaptic potentials. Each pharmacologically isolated component was reversibly reduced by bath-applied LY379268 and ((2S,1'R,2'R,3'R)-2-(2,3 dicarboxylcyclopropyl)-glycine, group II agonists. In an attempt to define the relevance of group II mGlu receptor activation on cholinergic transmission, we focused on the inhibitory effect on the IPSP, which was mimicked and occluded by omega-agatoxin IVA (omega-Aga-IVA), suggesting a modulation on P-type high voltage-activated calcium channels. Spontaneous calcium-dependent plateau potentials (PPs) were recorded with cesium-filled electrodes plus tetraethylammonium and TTX in the perfusing solution, and measurements of intracellular calcium [Ca2+]i changes were obtained simultaneously. PPs and the concomitant [Ca2+]i elevations were significantly reduced in amplitude and duration by LY379268. The mGlu-mediated inhibitory effect on PPs was mimicked by omega-Aga-IVA, suggesting an involvement of P-type channels. Moreover, electrically induced ACh release from striatal slices was reduced by mGlu2 receptor agonists and occluded by omega-Aga-IVA in a dose-dependent manner. Finally, double-labeling experiments combining mGlu2 receptor in situ hybridization and choline acetyltransferase immunocytochemistry revealed a strong mGlu2 receptor labeling on cholinergic interneurons, whereas single-label isotopic in situ hybridization for mGlu3 receptors did not show any labeling in these large striatal interneurons. These results suggest that the mGlu2 receptor mediated modulatory action on cell excitability would tune striatal ACh release, representing an interesting target for pharmacological intervention in basal ganglia disorders. PMID- 12122078 TI - Neural correlates of structure-from-motion perception in macaque V1 and MT. AB - Structure-from-motion (SFM) is the perception of three-dimensional shape from motion cues. We used a bistable SFM stimulus, which can be perceived in one of two different ways, to study how neural activity in cortical areas V1 and MT is related to SFM perception. Monkeys performed a depth-order task, where they indicated in which direction the front surface of a rotating SFM cylinder display was moving. To prevent contamination of the neural data because of eye position effects, all experiments with significant effects of radius, vergence, and velocity were excluded. As expected, the activity of approximately 50% of neurons in V1 and approximately 80% of neurons in MT is affected by the stimulus. Furthermore, the activity of 20% of neurons in area V1 is modulated with the percept. This proportion is higher in MT, where the activity of >60% of neurons is modulated with the percept. In both areas, this perceptual modulation occurs only in neurons with activity that is also affected by the stimulus. The perceptual modulation is not correlated with neural tuning properties in area V1, but it is in area MT. Together, these results suggest that V1 is not directly involved in the generation of the SFM percept, whereas MT is. The perceptual modulation in V1 may be attributable to top-down feedback from MT. PMID- 12122079 TI - Tyrosine phosphorylation of the NR2B subunit of the NMDA receptor in the spinal cord during the development and maintenance of inflammatory hyperalgesia. AB - The present study examined the levels of NMDA receptor NR2 subunit tyrosine phosphorylation in a rat model of inflammation and correlated it with the development of inflammation and hyperalgesia. Hindpaw inflammation and hyperalgesia were induced by intraplantar injection of complete Freund's adjuvant. Proteins from the spinal cord (L4-L5) were immunoprecipitated with anti NR2A or anti-NR2B antibodies and used for subsequent analysis using 4G-10, a specific anti-phosphotyrosine antibody. Compared with naive rats, there was a rapid and prolonged increase in tyrosine phosphorylation of the NR2B, but not NR2A, subunit after inflammation. The increase in NR2B tyrosine phosphorylation was dependent on primary afferent drive because (1) the phosphorylation correlated with the temporal profile of inflammation and hyperalgesia, (2) shorter-duration noxious stimulation produced a rapid and shorter-lasting increase in phosphorylation, and (3) local anesthetic block of the injected paw reversibly blocked inflammation-induced NR2B tyrosine phosphorylation and delayed hyperalgesia. The increase in NR2B tyrosine phosphorylation was abolished by intrathecal pretreatment with genistein, a tyrosine kinase inhibitor; PP2, an Src family tyrosine kinase inhibitor; AIDA, a group I metabotropic glutamate receptor antagonist; L733,060, an NK1 tachykinin receptor antagonist, and chelerythrine, a protein kinase C inhibitor. In addition, intrathecal PP2 delayed the onset of mechanical hyperalgesia and allodynia. These findings correlate in vivo NMDA receptor tyrosine phosphorylation with the development and maintenance of inflammatory hyperalgesia and suggest that signal transduction upstream to NR2B tyrosine phosphorylation involves G-protein-coupled receptors and PKC and Src family protein tyrosine kinases. PMID- 12122080 TI - Concurrent activation of dopamine D1 and D2 receptors is required to evoke neural and behavioral phenotypes of cocaine sensitization. AB - Repeated exposure to psychomotor stimulants produces a striking behavioral syndrome involving repetitive, stereotypic behaviors that occur if an additional exposure to the stimulant is experienced. The same stimulant exposure produces specific alterations in gene expression patterns in the striatum. To identify the dopamine receptor subtypes required for the parallel expression of these acquired neural and behavioral responses, we treated rats with different D1-class and D2 class dopamine receptor agonists and compared the responses of drug-naive rats with those of rats given previous intermittent treatment with cocaine. In rats exposed to repeated cocaine treatment, the effects of a subsequent challenge treatment with either a D1-class agonist (SKF 81297) or a D2-class agonist (quinpirole) were not significantly different from those observed in drug-naive animals: the drugs administered singly did not induce robust stereotyped motor behaviors nor produce significantly striosome-predominant expression of early genes in the striatum. In contrast, challenge treatment with the D1-class and D2 class agonists in combination led to marked and correlated increases in stereotypy and striosome-predominant gene expression in the striatum. Thus, immediately after repeated psychomotor stimulant exposure, only the concurrent activation of D1 and D2 receptor subclasses evoked expression of the neural and behavioral phenotypes acquired through repeated cocaine exposure. These findings suggest that D1-D2 dopamine receptor synergisms underlie the coordinate expression of both network-level changes in basal ganglia activation patterns and the repetitive and stereotypic motor response patterns characteristic of psychomotor stimulant sensitization. PMID- 12122081 TI - Interneuronal basis of the generation of related but distinct motor programs in Aplysia: implications for current neuronal models of vertebrate intralimb coordination. AB - Coordination of two sets of movements, protraction-retraction versus opening closing, of the feeding apparatus (the radula) in ingestive and egestive motor programs of Aplysia resembles vertebrate intralimb coordination in that the relative timing of the two sets of movements differs in the two motor programs. In both ingestion and egestion, radula protraction and retraction alternate, whereas radula closure shifts its phase relative to protraction-retraction. In egestion, the radula closes in protraction; in ingestion, the radula closes in retraction. In both ingestive and egestive motor programs elicited by the command like neuron, cerebral-buccal interneuron-2 (CBI-2), the protraction and retraction movements are mediated by the same sets of controller interneurons. In contrast, radula closure is mediated by two controller interneurons, B20 and B40, that are preferentially active in egestion and ingestion, respectively. In egestion, B20, active in protraction, drives closure motorneuron B8 in protraction, whereas in ingestion, B40, also active in protraction, uses a functionally novel mechanism, fast inhibition and slow excitation, to drive B8 in retraction. Our findings are summarized in a neural model that permits a conceptual comparison of our model with two previous hypothetical models of intralimb coordination in spinal circuits that were proposed by Grillner (1981, 1985) and Berkowitz and Stein (1994). Although our model supports the existence of separate controllers for different movements as in the Grillner (1981, 1985) model; in terms of basic mechanisms, our model is similar to the Berkowitz and Stein (1994) model because the closure movement is mediated by separate controllers in different programs, and thus both models can be classified as recruitment models. PMID- 12122082 TI - Phrenic long-term facilitation requires spinal serotonin receptor activation and protein synthesis. AB - Respiratory long-term facilitation (LTF) is a form of serotonin-dependent plasticity induced by intermittent hypoxia. LTF is manifested as a long-lasting increase in respiratory amplitude (and frequency) after the hypoxic episodes have ended. We tested the hypotheses that LTF of phrenic amplitude requires spinal serotonin receptor activation and spinal protein synthesis. A broad-spectrum serotonin receptor antagonist (methysergide) or protein synthesis inhibitors (emetine or cycloheximide) were injected intrathecally in the cervical spinal cord of anesthetized rats. Control rats, injected with vehicle (artificial CSF), exhibited an augmented phrenic burst amplitude after three 5 min episodes of hypoxia (78 +/- 15% above baseline, 60 min after hypoxia; p < 0.05), indicating LTF. Pretreatment with methysergide, emetine, or cycloheximide attenuated or abolished phrenic LTF (20 +/- 4, 0.2 +/- 11, and 20 +/- 2%, respectively; all p > 0.05). With protein synthesis inhibitors, phrenic LTF differed from control by 15 min after intermittent hypoxia. As an internal control against unintended drug distribution, we measured respiratory LTF in hypoglossal (XII) motor output. At 60 min after intermittent hypoxia, all treatment groups exhibited similar XII LTF (artificial CSF, 44 +/- 10%; methysergide, 40 +/- 5%; emetine, 35 +/- 9%; and cycloheximide, 57 +/- 29%; all p < 0.05), suggesting that drugs were restricted at effective doses to the spinal cord. We conclude that phrenic LTF requires spinal serotonin receptor activation and protein synthesis. Serotonin receptors on phrenic motoneuron dendrites may induce new protein synthesis, thereby giving rise to phrenic LTF. PMID- 12122083 TI - Dopamine release in the dorsal striatum during cocaine-seeking behavior under the control of a drug-associated cue. AB - Compulsive drug use is characterized by a pattern of drug seeking and consumption that becomes progressively habitual and less and less modifiable by external and internal factors. Although traditional views would posit that nigrostriatal dopamine (DA) neurons originating in the substantia nigra and innervating the dorsal striatum are primarily concerned with motor functions, recent studies have implicated the dorsal striatum in mediating stimulus-response (habit) learning. In this study, in vivo microdialysis in combination with a second-order schedule of cocaine reinforcement was used to investigate the role of the dorsal striatal dopamine innervation in well established drug-seeking behavior under the control of a drug-associated cue [light conditioned stimulus (CS+)]. Rats were initially trained to self-administer cocaine under a continuous reinforcement schedule where a response on one of two identical levers led to a 20 sec presentation of a light CS+ and an intravenous cocaine infusion (0.75 mg/kg). The response requirement for the CS+ and cocaine was then progressively increased until stable responding was established under a second-order schedule of reinforcement. During microdialysis, rats were presented with the cocaine-associated CS+ either noncontingently or contingent on responding during a session of cocaine-seeking behavior. The results showed a marked increase in DA release in the dorsal striatum during drug-seeking, when cocaine cues were presented contingently, but not when the same cue was presented noncontingently. These data indicate a possible involvement of the dopaminergic innervation of the dorsal striatum in well established, or habitual, cocaine-seeking behavior. PMID- 12122084 TI - Dynamic interactions between local surface cues, distal landmarks, and intrinsic circuitry in hippocampal place cells. AB - A number of computational models of hippocampal place cells incorporate attractor neural network architecture to simulate key findings in the place cell literature, including the properties of pattern completion, firing in the absence of visual input, and nonlinear responses to environmental manipulations. To test for evidence of attractor dynamics, ensembles of place cells were recorded using multiple-tetrode techniques. After many days of experience in an environment with salient local surface cues on a circular track and salient distal landmarks on the wall, the local surface cues were rotated as a set in opposition to the distal landmarks. The amount of mismatch between the local and distal sets of cues varied from 45 to 180 degrees. If place cells were parts of strong attractors, then their place fields should follow either the local cues or the distal cues as an integrated ensemble. Instead, in single recording sessions, some place cells were controlled by the distal landmarks, other cells were controlled by the local cues, and other cells became silent or gained new fields. In some cases, individual place fields split in half, following both the local and distal cues. If place cells are indeed parts of attractor networks in the hippocampus, then the attractors may be weak relative to the inputs from external sources, such as representations of the sensory environment and representations of heading direction, in a familiar, well explored environment. PMID- 12122085 TI - Leptin regulates growth hormone-releasing factor, somatostatin, and alpha melanocyte-stimulating hormone but not neuropeptide Y release in rat hypothalamus in vivo: relation with growth hormone secretion. AB - It is known that leptin, an adipocyte-derived hormone, exerts a stimulatory effect on growth hormone (GH) secretion in various animal species. However, no previous study examined in vivo whether leptin affects the secretion of GH releasing factor (GRF), somatostatin (SRIH), and some other closely relevant neurohormones in the hypothalamus. Therefore, in this study we investigated the effects of direct leptin infusion into the hypothalamus on the in vivo release of GRF, SRIH, alpha-melanocyte-stimulating hormone (alpha-MSH), and neuropeptide Y (NPY) in freely moving adult male rats using the push-pull perfusion. Leptin was infused into the median eminence-arcuate nucleus complex at three different concentrations, i.e., 1.0 (normal feeding level), 3.0, and 10 ng/ml (mild obesity level). In normally fed rats, only 10 ng/ml leptin was able to stimulate GH secretion, whereas in 3 d fasted rats, GH release was dose-dependently stimulated by 1.0 and 3.0 ng/ml leptin, although its 10 ng/ml dose did not produce additional effects. The facilitation of GH secretion occurred as increased pulse amplitudes without significant changes in the pulse frequency. During the leptin infusion, the hypothalamic GRF increased and SRIH decreased in magnitudes that approximately paralleled those of GH changes. Leptin stimulated the release of alpha-MSH in the fasted but not fed rats. It is likely that the fasting-induced increase in the hypothalamic alpha-MSH sensitivity to leptin is relevant to ingestive behavior involving leptin. Leptin was without effect on NPY release in either the fed or fasted group. Although it is certain that NPY mediates at least part of the metabolic actions of leptin, NPY is unlikely to be involved in the acute effects of leptin on GH, GRF, and SRIH secretion. These results demonstrate for the first time that leptin can alter the in vivo release of both GRF and SRIH in rat hypothalamus concurrently with the stimulation of GH secretion. PMID- 12122086 TI - Concurrent autoreceptor-mediated control of dopamine release and uptake during neurotransmission: an in vivo voltammetric study. AB - Receptor-mediated feedback control plays an important role in dopamine (DA) neurotransmission. Recent evidence suggests that release and uptake, key mechanisms determining brain extracellular levels of the neurotransmitter, are governed by presynaptic autoreceptors. The goal of this study was to investigate whether autoreceptors regulate both mechanisms concurrently. Extracellular DA in the caudate-putamen and nucleus accumbens, evoked by electrical stimulation of the medial forebrain bundle, was monitored in the anesthetized rat by real-time voltammetry. Effects of the D2 antagonist haloperidol (0.5 mg/kg, i.p.) on evoked DA levels were measured to evaluate autoreceptor control mechanisms. Two strategies were used to resolve individual contributions of release and uptake to the robust increases in DA signals observed after acute haloperidol challenge in naive animals: pretreatment with 3beta-(p-chlorophenyl)tropan-2beta-carboxylic acid p-isothiocyanatophenylmethyl ester hydrochloride (RTI-76; 100 nmol, i.c.v.), an irreversible inhibitor of the DA transporter, and kinetic analysis of extracellular DA dynamics. RTI-76 effectively removed the uptake component from recorded signals. In RTI-76-pretreated rats, haloperidol induced only modest increases in DA elicited by low frequencies and had little or no effect at high frequencies. These results suggest that D2 antagonism alters uptake at all frequencies but only release at low frequencies. Kinetic analysis similarly demonstrated that haloperidol decreased V(max) for DA uptake and increased DA release at low (10-30 Hz) but not high (40-60 Hz) stimulus frequencies. We conclude that presynaptic DA autoreceptors concurrently downregulate release and upregulate uptake, and that the mechanisms are also independently controlled during neurotransmission. PMID- 12122087 TI - Interactions between heterotypic stressors and corticosterone reveal integrative mechanisms for controlling corticotropin-releasing hormone gene expression in the rat paraventricular nucleus. AB - Although the convergence of neural and humoral afferent information onto paraventricular neuroendocrine corticotropin-releasing hormone (CRH) neurons is a major determinant for adaptive stress responses, the underlying integrative mechanisms are poorly understood. To dissect the relative contributions made by neural afferents and corticosterone to these processes, we determined how the concurrent application of two heterotypic physiological stressors, chronic dehydration (produced by drinking hypertonic saline) and sustained hypovolemia (produced by subcutaneous injections of polyethylene glycol), is interpreted by the synthetic and secretory activity of CRH neurons using in situ hybridization and plasma ACTH measurements. These two stressors are encoded by relatively simple, distinct, and well defined sets of neural afferents to CRH neurons. Both increase plasma corticosterone, but they have opposing actions on CRH gene expression when applied separately. In the first experiment, we showed that chronic dehydration suppresses CRH gene transcription after hypovolemia, but not the preproenkephalin and c-fos mRNA responses or ACTH secretion. In the second, we showed that negative feedback actions of corticosterone do not suppress CRH gene activation after hypovolemia, but instead determine the prestress lower limit of a range within which the CRH gene then responds. Collectively, these data show that at least two processes are integrated to control how the CRH gene responds to multiple stimuli. First, the presence of corticosterone, which although permissive for appropriately activating the CRH gene during hypovolemia, does not mediate the suppressed gene response. Second, neural afferent-driven processes that encode dehydration play a central role in suppressing CRH activation. PMID- 12122088 TI - Temporal encoding for auditory computation: physiology of primary afferent neurons in sound-producing fish. AB - Many fish rely on sounds for communication, yet the peripheral structures containing the hair cells are simple, without the morphological specializations that facilitate frequency analysis in the mammalian cochlea. Despite this, neurons in the midbrain of sound-producing fish (Pollimyrus) have complex receptive fields, extracting features from courtship sounds. Here we present an analysis of the initial encoding of sounds by the primary afferents and demonstrate that the representation of sound undergoes a substantial transformation as it ascends to the midbrain. Afferents were isolated as they coursed from the sacculus through the medulla. Tones (100 Hz-1.2 kHz) elicited synchronized spikes [vector strength (VS) >0.9] on each stimulus cycle [coefficient of variation (CV) <1.1], with little spike rate adaptation. Most afferents (67%) were spontaneously active and began synchronizing 10 dB below rate threshold. Rate thresholds for the most sensitive afferents (65 dB) were close to behavioral thresholds. The distribution of characteristic frequencies and best sensitivities was matched to the spectrum of sounds of this species and to its audiogram. Three clusters of afferents were identified, one including afferents that generated spike bursts and had v-shaped response areas (bursters), and two others that included entrained afferents with broad response areas (entrained types I and II). All afferents encoded the timing of clicks within click trains with time-locked spikes, and none showed selectivity for interclick intervals. Understanding the computations that yield complex receptive fields is an essential goal for auditory neuroscience, and these data on primary encoding advance this goal, allowing a comparison of inputs with feature-extracting midbrain neurons. PMID- 12122089 TI - Adjuvant chemotherapy for postmenopausal lymph node-negative breast cancer: it ain't necessarily so. PMID- 12122090 TI - Attribution of deaths following cancer treatment. PMID- 12122091 TI - Final report of Hanford Thyroid Disease Study released. PMID- 12122093 TI - Researchers push for sharing of trial results with participants. PMID- 12122094 TI - Can increased treatment of hepatitis C stem the tide of liver cancer? PMID- 12122096 TI - Endocrine responsiveness and tailoring adjuvant therapy for postmenopausal lymph node-negative breast cancer: a randomized trial. AB - BACKGROUND: The role of adjuvant chemotherapy in postmenopausal patients with lymph node-negative breast cancer is controversial. After demonstrating the efficacy of chemotherapy combined with tamoxifen for postmenopausal patients with lymph node-positive disease, the International Breast Cancer Study Group launched a randomized trial (Trial IX) to evaluate the role of adjuvant chemotherapy preceding treatment with tamoxifen for patients with lymph node-negative disease. METHODS: After stratification by estrogen receptor (ER) status, patients were randomly assigned to receive three 28-day courses of "classical" adjuvant CMF chemotherapy (cyclophosphamide at 100 mg/m(2) on days 1-14, orally; methotrexate at 40 mg/m(2) on days 1 and 8, intravenously; and 5-fluorouracil at 600 mg/m(2) on days 1 and 8, intravenously) followed by tamoxifen (20 mg/day, orally for 57 months) (CMF-->tamoxifen) or to receive tamoxifen alone (20 mg/day, orally for 60 months). We enrolled 1669 eligible patients, 382 (23%) with ER-negative tumors, 1217 (73%) with ER-positive tumors, and 70 (4%) with unknown ER status. The median follow-up was 71 months. All statistical tests were two-sided. RESULTS: The added benefit of CMF followed by tamoxifen over tamoxifen alone was statistically significantly dependent on ER status (tests for interaction: P =.01 for disease-free survival [DFS] and P =.07 for overall survival [OS]). For patients with ER-negative tumors, the addition of CMF statistically significantly improved DFS (5-year DFS = 84% for CMF-->tamoxifen versus 69% for tamoxifen alone; difference = 15%; 95% confidence interval [CI] = 6% to 24%; risk ratio [RR] = 0.52; 95% CI = 0.34 to 0.79; P =.003) and OS (5-year OS = 89% for CMF- >tamoxifen versus 81% for tamoxifen alone; difference = 8%; 95% CI = 0% to 16%; RR = 0.51; 95% CI = 0.30 to 0.87; P =.01). By contrast, for patients with ER positive tumors, addition of CMF provided no benefit in terms of DFS (5-year DFS = 84% for CMF-->tamoxifen versus 85% for tamoxifen alone; difference = -1; 95% CI = -6% to 4%; RR = 0.99; 95% CI = 0.75 to 1.30; P =.92) or OS (5-year OS = 95% for CMF-->tamoxifen versus 93% for tamoxifen alone; difference = 2%; 95% CI = -1% to 5%; RR = 0.95; 95% CI = 0.64 to 1.40; P =.80). CONCLUSIONS: Postmenopausal patients with lymph node-negative breast cancer benefited substantially from adjuvant chemotherapy if their cancer was ER-negative (i.e., endocrine nonresponsive). In contrast, if their cancer was ER-positive (i.e., endocrine responsive), they obtained no benefit from the combination treatment compared with tamoxifen alone. PMID- 12122097 TI - Are deaths within 1 month of cancer-directed surgery attributed to cancer? AB - BACKGROUND: Cancer mortality should include not only deaths from cancer but also deaths from cancer treatment. By convention, deaths within 30 days of a surgical procedure are considered treatment-related deaths in the calculation of operative mortality-that is, the chance of dying from surgery. How cause of death is attributed in patients who die within 1 month of cancer-directed surgery is unknown. METHODS: The National Cancer Institute's Surveillance, Epidemiology, and End Results (SEER) program data from 1994 through 1998 were used to examine the cause of death in patients diagnosed with one of 19 common solid tumors who had died within 1 month of diagnosis and had also received cancer-directed surgery. We determined the proportion of deaths not attributed to the cancer and the magnitude of the undercount in cancer-specific mortality. RESULTS: Among 4135 patients with only one cancer who died within 1 month of diagnosis and cancer directed surgery, the proportion of deaths not attributed to the coded cancer was 41% (1714/4135), ranging from 13% (1/8) for cervical cancer to 81% (13/16) for laryngeal cancer. Selected intermediate values include 25% (14/56) for esophageal cancer, 34% (177/525) for lung cancer, 42% (719/1695) for colorectal cancer, 59% (110/186) for breast cancer, and 75% (80/106) for prostate cancer. Restricting the analysis to deaths following specific major procedures (e.g., esophagectomy, pneumonectomy, colectomy) had little effect on the findings. If all deaths within 1 month of cancer-directed surgery were attributed to cancer, cancer mortality would rise about 1%. CONCLUSION: Some deaths that are conventionally attributed to surgery are not being attributed to the cancer for which the surgery was performed. Although the estimated effect of this misclassification on overall cancer mortality is modest, it may be indicative of more widespread confusion about how to code treatment-related deaths of patients with cancer. PMID- 12122098 TI - Diagnosis of genito-urinary tract cancer by detection of minichromosome maintenance 5 protein in urine sediments. AB - BACKGROUND: Because cystoscopy is invasive and expensive and urine cytology has low sensitivity, alternative methods for detecting bladder cancer are sought. Minichromosome maintenance (Mcm) proteins have been used as diagnostic markers for cervical cancer. We investigated whether one Mcm protein, Mcm5, can be used to detect urothelial cancer cells in urine sediments. METHODS: We used two monoclonal antibodies against His-tagged human Mcm5 (amino acids 367-582) in an immunofluorometric assay to measure Mcm5 levels in cells in the urine of 353 patients who presented with hematuria or lower urinary tract symptoms or who were undergoing follow-up cystoscopy for urothelial neoplasia. Urine samples were also subjected to routine cytologic analysis. Patients underwent upper urinary tract imaging and cystoscopy within 12 hours of producing the urine sample. Data were analyzed by comparing areas under a nonparametric receiver operating characteristics (ROC) curve and by McNemar's test and Fisher's exact test. All statistical tests were two-sided. RESULTS: At the assay cut point where the false negative and false-positive rates were the same, the Mcm5 test detected primary and recurrent bladder cancers with 87% (95% confidence interval [CI] = 77% to 94%) sensitivity and 87% (95% CI = 83% to 91%) specificity. At the cut point where the specificities of urine cytology and the Mcm5 test were equal (97%, 95% CI = 95% to 99%), the Mcm5 test was statistically significantly (P<.001) more sensitive than urine cytology, 73% (95% CI = 61% to 83%) versus 48% (95% CI = 35% to 60%). At the lower detection limit of the Mcm5 test, sensitivity was highest, 92% (95% CI = 83% to 97%) and specificity was 78% (95% CI = 72% to 83%). Patients with prostate cancer had higher levels of Mcm5 in their urine sediments than did men without malignancy (P<.001). CONCLUSIONS: Elevated levels of Mcm5 in urine sediments are highly predictive of bladder cancer. PMID- 12122099 TI - Coexpression of glucose transporter 1 and matrix metalloproteinase-2 in human cancers. AB - BACKGROUND: Cancer cells express higher levels of glucose transporter proteins (Gluts) than do normal cells. Glut-1 overexpression is associated with invasiveness. Because matrix metalloproteinase-2 (MMP-2) is also overexpressed in cancer cells and is associated with invasiveness, we tested the hypothesis that Glut-1 may regulate MMP-2 expression. METHODS: We transiently transfected Glut-1 complementary DNA (cDNA) or Glut-1 antisense oligonucleotides in the human rhabdomyosarcoma cell line RD and analyzed MMP-2 mRNA expression and cell invasiveness. Empty vector and sense oligonucleotides were used for controls. We analyzed MMP-2 promoter activity in transfectants with a luciferase reporter assay and with p53 and Ets-1 gel mobility shift assays. Eight human cancer cell lines and 80 human cancer specimens were analyzed for coexpression of Glut-1 and MMP-2 by western blot and immunohistochemical analyses, respectively. RESULTS: Overexpression of Glut-1 in RD cells increased MMP-2 expression 4.3-fold (95% confidence interval [CI] = 3.7-fold to 4.9-fold) and invasiveness 3.2-fold (95% CI = 2.6-fold to 3.8-fold) relative to control transfected cells. Conversely, suppression of Glut-1 expression by antisense oligonucleotides decreased MMP-2 expression by 71.5% (95% CI = 71.1% to 71.9%) and invasiveness by 53.0% (95% CI = 47.5% to 58.5%). Glut-1-mediated MMP-2 expression involved the binding of the transcription factor p53 but not Ets-1. All eight human cancer cell lines coexpressed Glut-1 and MMP-2 by western blotting, and 45 of 80 human tumor tissues coexpressed Glut-1 and MMP-2 by immunohistochemistry. CONCLUSIONS: MMP-2 expression and cell invasiveness are tightly associated with Glut-1 expression in human cancer cell lines. Because suppression of Glut-1 decreased MMP-2 expression and cancer cell invasion, Glut-1 could be a target for therapy of various cancers that overexpress Glut-1. PMID- 12122100 TI - An analysis of DNA repair as a determinant of survival in patients with non-small cell lung cancer. AB - BACKGROUND: Non-small-cell lung cancer (NSCLC) is frequently resistant to chemotherapy, and this resistance has been associated with elevated nucleotide excision repair (NER) in tumor tissue. We hypothesized that patients with NSCLC who had effective systemic (host) NER would have poorer survival than patients with suboptimal NER and that the association between NER effectiveness and survival would be most marked in patients receiving chemotherapy. METHODS: 375 patients with newly diagnosed NSCLC were accrued for a case-control study between July 1995 and December 1999. NER activity was estimated as the DNA repair capacity (DRC) measured in the patient's peripheral lymphocytes by the host cell reactivation assay. Cox proportional hazards models were used to assess the association between DRC and survival. All statistical tests were two-sided. RESULTS: For every unit (percentage) increase in DRC, the relative risk (RR) of death was 1.05 (95% confidence interval [CI] = 1.00 to 1.10; P =.05) for the 345 patients for whom weight loss information was available and 1.06 (95% CI = 1.00 to 1.12; P =.03) for the 275 patients with complete follow-up information. In 86 patients treated with chemotherapy only, the RR of death increased to 1.11 (95% CI = 1.02 to 1.21; P =.01) for every unit (percentage) increase in DRC. Of those 86 patients, patients in the top quartile of the DRC distribution were at twice the RR of death as those in the lowest quartile (RR = 2.72; 95% CI = 1.24 to 5.95; P =.01). Effective DRC was not a risk factor for death in patients who were not treated with chemotherapy. CONCLUSIONS: Our data suggest that effective host DRC may be associated with poorer survival in patients with NSCLC who are treated with chemotherapy. PMID- 12122101 TI - Insulin-like growth factor-I (IGF-I) and IGF binding protein-3 as predictors of advanced-stage prostate cancer. AB - BACKGROUND: Plasma levels of insulin-like growth factor-I (IGF-I) have been associated with the risk of prostate cancer. However, the association of IGF-I with specific tumor stage and grade at diagnosis, which correlate with risk of recurrence and mortality, has not been studied rigorously. To determine whether plasma levels of IGF-I and its main circulating binding protein, IGF binding protein-3 (IGFBP-3), predict more aggressive forms of prostate cancer, we investigated the association between plasma levels of each and specific stages and grades of prostate cancer. METHODS: We examined 530 case patients and 534 control subjects in a nested case-control study in the prospective Physicians' Health Study. Patients with prostate cancer diagnosed from 1982 through 1995 were matched to control subjects by age and smoking status. IGF-I and IGFBP-3 were measured in prospectively collected plasma samples. Conditional logistic regression models were used to estimate the relative risks (RRs) for prostate cancer associated with IGF-I and IGFBP-3, stratified on grade (Gleason score > or = 7 versus <7) and stage (early = stage A or B prostate cancer versus advanced = stage C or D prostate cancer). All statistical tests were two-sided. RESULTS: Plasma levels of IGF-I and IGFBP-3 were predictors of advanced-stage prostate cancer (RR = 5.1, 95% confidence interval [CI] = 2.0 to 13.2 for highest versus lowest quartiles of IGF-I; RR = 0.2, 95% CI = 0.1 to 0.6 for highest versus lowest quartiles of IGFBP-3) but not of early-stage prostate cancer. Neither was differentially associated with Gleason score. Men with high IGF-I levels and low IGFBP-3 levels had an RR for advanced-stage prostate cancer of 9.5 (95% CI = 1.9 to 48.4) compared with men with low levels of both. Combining IGF-I and IGFPB-3 measurements with a standard prostate-specific antigen (PSA) measurement for prostate cancer screening increased the specificity (from 91% to 93%) but decreased sensitivity (from 40% to 36%) compared with measurement of PSA alone. CONCLUSIONS: Circulating levels of IGF-I and IGFBP-3 may predict the risk of developing advanced-stage prostate cancer, but their utility for screening patients with incident prostate cancer may be limited. PMID- 12122102 TI - Development of an improved vaccine for anthrax. PMID- 12122103 TI - Herpes simplex viruses: is a vaccine tenable? PMID- 12122104 TI - The surprising role of vascular K(ATP) channels in vasospastic angina. PMID- 12122105 TI - Regulation of renin expression and blood pressure by vitamin D(3). PMID- 12122106 TI - Making room for T cells. PMID- 12122107 TI - The TSH receptor reveals itself. PMID- 12122108 TI - Viral induction of a chronic asthma phenotype and genetic segregation from the acute response. AB - Paramyxoviral infections cause most of the acute lower respiratory tract illness in infants and young children and predispose to the development of chronic wheezing, but the relationship between these short- and long-term viral effects are uncertain. Here we show that a single paramyxoviral infection of mice (C57BL6/J strain) not only produces acute bronchiolitis, but also triggers a chronic response with airway hyperreactivity and goblet cell hyperplasia lasting at least a year after complete viral clearance. During the acute response to virus, same-strain ICAM-1-null mice are protected from airway inflammation and hyperreactivity despite similar viral infection rates, but the chronic response proceeds despite ICAM-1 deficiency. Neither response is influenced by IFN-gamma deficiency, but the chronic response is at least partially prevented by glucocorticoid treatment. In contrast to viral infection, allergen challenge caused only short-term expression of asthma phenotypes. Thus, paramyxoviruses cause both acute airway inflammation/hyperreactivity and chronic airway remodeling/hyperreactivity phenotypes (the latter by a hit-and-run strategy, since viral effects persist after clearance). These two phenotypes can be segregated by their dependence on the ICAM-1 gene and so depend on distinct controls that appear critical for the development of lifelong airway diseases such as asthma. PMID- 12122109 TI - The Polycomb-group gene Rae28 sustains Nkx2.5/Csx expression and is essential for cardiac morphogenesis. AB - The Polycomb-group (PcG) gene Rae28 is a mammalian homologue of the Drosophila gene polyhomeotic. PcG genes are known to maintain transcription states, once initiated, probably by regulating chromatin structure. Since homozygous Rae28 deficient (Rae28(-/-)) mice displayed cardiac anomalies similar to congenital heart diseases in humans, we examined the role of Rae28 in cardiac morphogenesis at the molecular level. In Rae28(-/-) embryos, expression of the cardiac selector gene Nkx2.5/Csx (Nkx2.5) was initiated properly but was not sufficiently sustained later in development. This impaired expression of Nkx2.5 in the maintenance phase proved to have a crucial effect on cardiac morphogenesis, as demonstrated by the results of a genetic complementation experiment in which the cardiac anomalies were suppressed by overexpression of human NKX2.5/CSX1 in Rae28(-/-) embryos. Ubiquitous expression of exogenous Rae28 likewise restored the impaired Nkx2.5 expression in Rae28(-/-) embryos, further supporting the notion that Rae28 sustains Nkx2.5 expression in cardiomyocytes. Thus, our data show that a mammalian PcG gene can play a key role in organogenesis by helping to maintain the expression of a selector gene. PMID- 12122110 TI - T cell homeostatic proliferation elicits effective antitumor autoimmunity. AB - Development of tumor immunotherapies focuses on inducing autoimmune responses against tumor-associated self-antigens primarily encoded by normal, unmutated genes. We hypothesized that such responses could be elicited by T cell homeostatic proliferation in the periphery, involving expansion of T cells recognizing self-MHC/peptide ligands. Herein, we demonstrate that sublethally irradiated lymphopenic mice transfused with autologous or syngeneic T cells showed tumor growth inhibition when challenged with melanoma or colon carcinoma cells. Importantly, the antitumor response depended on homeostatic expansion of a polyclonal T cell population within lymph nodes. This response was effective even for established tumors, was characterized by CD8(+) T cell-mediated tumor specific cytotoxicity and IFN-gamma production, and was associated with long-term memory. The results indicate that concomitant induction of the physiologic processes of homeostatic T cell proliferation and tumor antigen presentation in lymph nodes triggers a beneficial antitumor autoimmune response. PMID- 12122111 TI - Hepatocyte-specific inhibition of NF-kappaB leads to apoptosis after TNF treatment, but not after partial hepatectomy. AB - One of the earliest TNF-dependent events to occur during liver regeneration is the activation of the transcription factor NF-kappaB through TNF receptor type 1. NF-kappaB activation in the liver can have both antiapoptotic and proliferative effects, but it is unclear which liver cell types, hepatocytes or nonparenchymal cells (NPCs), contribute to these effects. To specifically evaluate the role of hepatocyte NF-kappaB, we created GLVP/DeltaN-IkappaB(alpha) transgenic mice, in which expression of a deletion mutant of IkappaB(alpha) (DeltaN-IkappaB(alpha)) was induced in hepatocytes after injection of mifepristone. In control mice, injection of 25 microg/kg TNF caused NF-kappaB nuclear translocation in virtually all hepatocytes by 30 minutes and no detectable apoptosis, while in mice expressing DeltaN-IkappaB(alpha), NF-kappaB nuclear translocation was blocked in 45% of hepatocytes, leading to apoptosis 4 hours after TNF injection. In contrast, expression of DeltaN-IkappaBalpha in hepatocytes during the first several hours after partial hepatectomy did not lead to apoptosis or decreased proliferation. As NF-kappaB activation was not inhibited in liver NPCs, it is likely that these cells are responsible for mediating the proliferative and antiapoptotic effects of NF-kappaB during liver regeneration. PMID- 12122112 TI - Episodic coronary artery vasospasm and hypertension develop in the absence of Sur2 K(ATP) channels. AB - K(ATP) channels couple the intracellular energy state to membrane excitability and regulate a wide array of biologic activities. K(ATP) channels contain a pore forming inwardly rectifying potassium channel and a sulfonylurea receptor regulatory subunit (SUR1 or SUR2). To clarify the role of K(ATP) channels in vascular smooth muscle, we studied Sur2 gene-targeted mice (Sur2(-/-)) and found significantly elevated resting blood pressures and sudden death. Using in vivo monitoring, we detected transient, repeated episodes of coronary artery vasospasm in Sur2(-/-) mice. Focal narrowings in the coronary arteries were present in Sur2(-/-) mice consistent with vascular spasm. We treated Sur2(-/-) mice with a calcium channel antagonist and successfully reduced vasospastic episodes. The intermittent coronary artery vasospasm seen in Sur2(-/-) mice provides a model for the human disorder Prinzmetal variant angina and demonstrates that the SUR2 K(ATP) channel is a critical regulator of episodic vasomotor activity. PMID- 12122113 TI - Thyroid-stimulating autoantibodies in Graves disease preferentially recognize the free A subunit, not the thyrotropin holoreceptor. AB - Graves disease is directly caused by thyroid-stimulating autoantibodies (TSAb's) that activate the thyrotropin receptor (TSHR). We observed upon flow cytometry using intact cells that a mouse mAb (3BD10) recognized the TSHR ectodomain with a glycosidylphosphatidylinositol (ECD-GPI) anchor approximately tenfold better than the same ectodomain on the wild-type TSHR, despite the far higher level of expression of the latter. The 3BD10 epitope contains the N-terminal cysteine cluster critical for TSAb action. Consequently, we hypothesized and confirmed that TSAb (but not thyrotropin-blocking autoantibodies [TBAb's]) also poorly recognize the wild-type TSHR relative to the ECD-GPI. Despite poor recognition by TSAb of the holoreceptor, soluble TSHR A subunits (known to be shed from surface TSHR) fully neutralized autoantibody-binding activity. These data indicate that the epitope(s) for TSAb's, but not for TBAb's, are partially sterically hindered on the holoreceptor by the plasma membrane, the serpentine region of the TSHR, or by TSHR dimerization. However, the TSAb epitope on the soluble A subunit is freely accessible. This observation, as well as other evidence, supports the concept that A subunit shedding either initiates or amplifies the autoimmune response to the TSHR, thereby causing Graves disease in genetically susceptible individuals. PMID- 12122114 TI - Signaling through estrogen receptors modulates telomerase activity in human prostate cancer. AB - Sex steroid hormone receptors play a central role in all stages of prostate cancer. Here, we tested whether estrogen receptor (ER) signaling contributes to telomerase activation, an early event in prostate tumorigenesis. Following 17beta estradiol (E(2)) treatment, both mRNA encoding the catalytic subunit of human telomerase (hTERT) and telomerase activity were promptly induced in human prostate normal epithelial cells, fresh explants from benign prostate hyperplasia, and prostate cancer explants and cell lines. Reporter expression studies and in vivo chromatin immunoprecipitation assays revealed E(2)-dependent hTERT promoter induction and showed that both ERalpha and ERbeta bound this sequence. Crucially, addition of the anti-estrogen 4-hydroxytamoxifen caused a differential recruitment in vivo of ERalpha and ERbeta onto the hTERT promoter and inhibited telomerase activity. Treatment with the aromatase inhibitor letrozole, which prevented testosterone-mediated interaction between ER and the hTERT estrogen response element, resulted in a negative regulation of telomerase activity. Thus, intracellular conversion of androgens to estrogens may contribute to the etiopathogenesis of prostate cancer. Given the present evidence for direct control of hTERT gene expression and telomerase activity in the prostate by the ER, we suggest that this transcriptional regulator represents a possible therapeutic target in prostate cancer. PMID- 12122115 TI - 1,25-Dihydroxyvitamin D(3) is a negative endocrine regulator of the renin angiotensin system. AB - Inappropriate activation of the renin-angiotensin system, which plays a central role in the regulation of blood pressure, electrolyte, and volume homeostasis, may represent a major risk factor for hypertension, heart attack, and stroke. Mounting evidence from clinical studies has demonstrated an inverse relationship between circulating vitamin D levels and the blood pressure and/or plasma renin activity, but the mechanism is not understood. We show here that renin expression and plasma angiotensin II production were increased severalfold in vitamin D receptor-null (VDR-null) mice, leading to hypertension, cardiac hypertrophy, and increased water intake. However, the salt- and volume-sensing mechanisms that control renin synthesis are still intact in the mutant mice. In wild-type mice, inhibition of 1,25-dihydroxyvitamin D(3) [1,25(OH)(2)D(3)] synthesis also led to an increase in renin expression, whereas 1,25(OH)(2)D(3) injection led to renin suppression. We found that vitamin D regulation of renin expression was independent of calcium metabolism and that 1,25(OH)(2)D(3) markedly suppressed renin transcription by a VDR-mediated mechanism in cell cultures. Hence, 1,25(OH)(2)D(3) is a novel negative endocrine regulator of the renin-angiotensin system. Its apparent critical role in electrolytes, volume, and blood pressure homeostasis suggests that vitamin D analogues could help prevent or ameliorate hypertension. PMID- 12122116 TI - Essential role of Gas6 for glomerular injury in nephrotoxic nephritis. AB - Growth-arrest specific gene 6 (Gas6) is a vitamin K-dependent growth factor for mesangial and epithelial cells. To investigate whether Gas6 is essential for progressive glomerular injury, we constructed Gas6(-/-) mice and examined the role of Gas6 in accelerated nephrotoxic nephritis (NTN), a model of progressive glomerulonephritis. We found less mortality and proteinuria in Gas6(-/-) mice than in wild-type mice following injection of nephrotoxic serum. Glomerular cell proliferation, glomerular sclerosis, crescent formation, and deposition of fibrin/fibrinogen in glomeruli were also reduced in Gas6(-/-) mice. Furthermore, administering Gas6(-/-) mice recombinant wild-type Gas6, but not Gas6 lacking a previously characterized N-terminal gamma-carboxyl group, induced massive proteinuria, glomerular cell proliferation, and glomerulosclerosis, comparable to responses seen in wild-type mice. These data indicate that Gas6 induces glomerular cell proliferation in NTN and suggest that this factor contributes to glomerular injury and the progression of chronic nephritis. PMID- 12122117 TI - Functional redundancy of Rab27 proteins and the pathogenesis of Griscelli syndrome. AB - Griscelli syndrome (GS) patients and the corresponding mouse model ashen exhibit defects mainly in two types of lysosome-related organelles, melanosomes in melanocytes and lytic granules in CTLs. This disease is caused by loss-of function mutations in RAB27A, which encodes 1 of the 60 known Rab GTPases, critical regulators of vesicular transport. Here we present evidence that Rab27a function can be compensated by a closely related protein, Rab27b. Rab27b is expressed in platelets and other tissues but not in melanocytes or CTLs. Morphological and functional tests in platelets derived from ashen mice are all within normal limits. Both Rab27a and Rab27b are found associated with the limiting membrane of platelet-dense granules and to a lesser degree with alpha granules. Ubiquitous transgenic expression of Rab27a or Rab27b rescues ashen coat color, and melanocytes derived from transgenic mice exhibit widespread peripheral distribution of melanosomes instead of the perinuclear clumping observed in ashen melanocytes. Finally, transient expression in ashen melanocytes of Rab27a or Rab27b, but not other Rab's, restores peripheral distribution of melanosomes. Our data suggest that Rab27b is functionally redundant with Rab27a and that the pathogenesis of GS is determined by the relative expression of Rab27a and Rab27b in specialized cell types. PMID- 12122122 TI - Jean-Martin Charcot (1825-1893). The man behind the joint disease. AB - Jean-Martin Charcot was one of the most celebrated French physicians of the 19th century. A masterful teacher and a captivating lecturer, Charcot created the foundations of neurology as an independent discipline, and transformed the Salpetriere hospital, in Paris, into one of the world's greatest teaching centers for clinical neurologic research. His name is attached to the distinct pathologic entity, Charcot's joint disease, that he so meticulously described. This article reviews the highlights of Charcot's career and his clinicoanatomic studies of patients with tabetic arthropathies. PMID- 12122118 TI - Treatment of cardiomyopathy and rhabdomyolysis in long-chain fat oxidation disorders using an anaplerotic odd-chain triglyceride. AB - The current dietary treatment of long-chain fatty acid oxidation defects (high carbohydrate with medium-even-chain triglycerides and reduced amounts of long chain fats) fails, in many cases, to prevent cardiomyopathy, rhabdomyolysis, and muscle weakness. We hypothesized that the apparent defect in energy production results from a depletion of the catalytic intermediates of the citric acid cycle via leakage through cell membranes (cataplerosis). We further hypothesized that replacing dietary medium-even-chain fatty acids (precursors of acetyl-CoA) by medium-odd-chain fatty acids (precursors of acetyl-CoA and anaplerotic propionyl CoA) would restore energy production and improve cardiac and skeletal muscle function. We fed subjects with long-chain defects a controlled diet in which the fat component was switched from medium-even-chain triglycerides to triheptanoin. In three patients with very-long-chain acyl-CoA dehydrogenase deficiency, this treatment led rapidly to clinical improvement that included the permanent disappearance of chronic cardiomyopathy, rhabdomyolysis, and muscle weakness (for more than 2 years in one child), and of rhabdomyolysis and weakness in the others. There was no evidence of propionyl overload in these patients. The treatment has been well tolerated for up to 26 months and opens new avenues for the management of patients with mitochondrial fat oxidation disorders. PMID- 12122123 TI - Medical treatment of Charcot's arthropathy. AB - Charcot's arthropathy is a devastating condition affecting diabetic patients with peripheral neuropathy, resulting in a foot at risk for ulceration and amputation. Early diagnosis is important for the institution of appropriate treatment, which may help prevent disease progression and foot deformity. This article discusses the pathogenesis and treatment options available for the disorder. PMID- 12122119 TI - The MEKK1-JNK pathway plays a protective role in pressure overload but does not mediate cardiac hypertrophy. AB - Mitogen-activated protein kinase kinase kinase (MEKK1) mediates activation of c Jun NH(2)-terminal kinase (JNK). Although previous studies using cultured cardiac myocytes have suggested that the MEKK1-JNK pathway plays a key role in hypertrophy and apoptosis, its effects in cardiac hypertrophy and apoptosis are not fully understood in adult animals in vivo. We examined the role of the MEKK1 JNK pathway in pressure-overloaded hearts by using mice deficient in MEKK1. We found that transverse aortic banding significantly increased JNK activity in Mekk1(+/+) but not Mekk1(-/-) mice, indicating that MEKK1 mediates JNK activation by pressure overload. Nevertheless, pressure overload caused significant levels of cardiac hypertrophy and expression of atrial natriuretic factor in Mekk1(-/-) animals, which showed higher mortality and lung/body weight ratio than were seen in controls. Fourteen days after banding, Mekk1(-/-) hearts were dilated, and their left ventricular ejection fraction was low. Pressure overload caused elevated levels of apoptosis and inflammatory lesions in these mice and produced a smaller increase in TGF-beta and TNF-alpha expression than occurred in wild type controls. Thus, MEKK1 appears to be required for pressure overload-induced JNK activation and cytokine upregulation but to be dispensable for pressure overload-induced cardiac hypertrophy. MEKK1 also prevents apoptosis and inflammation, thereby protecting against heart failure and sudden death following cardiac pressure overload. PMID- 12122124 TI - From acute to chronic: monitoring the progress of Charcot's arthropathy. AB - The monitoring of Charcot's arthropathy in patients with diabetes mellitus is twofold: 1) assessment of disease activity as the condition progresses from the acute to the chronic phase, and 2) identification of structural abnormalities and complications that may arise as a result of the disease. The former guides the clinician as to the duration of primary treatment, and the latter provides important information regarding the long-term prognosis and facilitates clinical decision making regarding other treatments including surgery, footwear, and orthoses. The mainstay of assessing disease activity remains thorough and regular assessment of swelling, temperature differences, and bony abnormalities. Radiographic assessment performed at baseline and periodically throughout the course of the disease will show stages of early fracture and fragmentation followed by eventual trabecular bridging, ankylosis of the affected joints, and sclerosis, heralding the chronic phase of the disease. Radiographic assessment also provides visualization of bony deformities and prominences. In addition to these assessments, changes may be further quantified by the use of infrared dermal thermography and quantitative bone scanning techniques. Careful clinical monitoring of patients is essential to optimize treatment for acute Charcot's arthropathy and improve the long-term outcome for patients presenting with this condition. PMID- 12122125 TI - Charcot's arthropathy of the foot. PMID- 12122126 TI - Use of subatmospheric (VAC) therapy to improve bioengineered tissue grafting in diabetic foot wounds. AB - The use of bioengineered tissue and topical subatmospheric pressure therapy have both been widely accepted as adjunctive therapies for the treatment of noninfected, nonischemic diabetic foot wounds. This article describes a temporally overlapping method of care that includes a period of simultaneous application of bioengineered tissue (Apligraf, Novartis Pharmaceuticals Corp, East Hanover, New Jersey) and subatmospheric pressure therapy delivered through the VAC (Vacuum Assisted Closure) system (KCI, Inc, San Antonio, Texas). Future descriptive and analytic works may test the hypothesis that combined therapies used at different and often overlapping periods during the wound-healing cycle may be more effective than a single modality. PMID- 12122127 TI - Maggot debridement therapy: a primer. AB - Treatment of chronic wounds of the lower extremity requires a systematic, multidisciplinary approach as well as flexibility in order to achieve acceptable, consistent short-term and long-term results. Maggots, once considered an obsolete therapeutic modality, can be a useful addition to the armamentarium of the foot and ankle specialist. This article describes the use of maggot debridement therapy for intractable wounds of the lower extremity. PMID- 12122128 TI - How and why to surgically debride neuropathic diabetic foot wounds. AB - Wound debridement, when systematically performed, may be as important as off loading in reducing the prevalence of chronic inflammatory by-products in a wound and thus in converting a chronic wound into an acute one. Although it has been suggested that aggressive surgical debridement of wounds may be beneficial, there have been few, if any, technical descriptions of this aspect of therapy. It is therefore the purpose of this article to describe the general principles, process, and technique of outpatient surgical debridement of noninfected, nonischemic neuropathic diabetic foot wounds performed at the authors' institutions. The authors hope to foster further discussion leading to improvement in the process and the prevalence of such debridement. PMID- 12122129 TI - Technique for fabrication of an "instant total-contact cast" for treatment of neuropathic diabetic foot ulcers. AB - Addressing pressure reduction in the treatment of diabetic foot wounds is a critical component of therapy. The total-contact cast has proven to be the gold standard of treatment because of its ability to reduce pressure and facilitate patient adherence to the off-loading regimen. Removable cast walkers have proven to be as effective as total-contact casts in pressure reduction, but this has not translated into equivalent time to healing. A simple technique to convert the removable cast walker into a device that is not as easily detached from the lower extremity, thereby encouraging the use of this device over a 24-hour period, is presented in this article. The procedure involves wrapping the cast walker with cohesive bandage or plaster of Paris. In the authors' opinion, this technique addresses many of the disadvantages of the total-contact cast, resulting in an adequate compromise in this aspect of care. PMID- 12122132 TI - Lactate -- the forgotten fuel! PMID- 12122133 TI - Background potassium channels move into focus. PMID- 12122134 TI - Do seminiferous tubules secrete a fluid rich in KHCO3? PMID- 12122135 TI - Quantitative evolutionary design. AB - The field of quantitative evolutionary design uses evolutionary reasoning (in terms of natural selection and ultimate causation) to understand the magnitudes of biological reserve capacities, i.e. excesses of capacities over natural loads. Ratios of capacities to loads, defined as safety factors, fall in the range 1.2 10 for most engineered and biological components, even though engineered safety factors are specified intentionally by humans while biological safety factors arise through natural selection. Familiar examples of engineered safety factors include those of buildings, bridges and elevators (lifts), while biological examples include factors of bones and other structural elements, of enzymes and transporters, and of organ metabolic performances. Safety factors serve to minimize the overlap zone (resulting in performance failure) between the low tail of capacity distributions and the high tail of load distributions. Safety factors increase with coefficients of variation of load and capacity, with capacity deterioration with time, and with cost of failure, and decrease with costs of initial construction, maintenance, operation, and opportunity. Adaptive regulation of many biological systems involves capacity increases with increasing load; several quantitative examples suggest sublinear increases, such that safety factors decrease towards 1.0. Unsolved questions include safety factors of series systems, parallel or branched pathways, elements with multiple functions, enzyme reaction chains, and equilibrium enzymes. The modest sizes of safety factors imply the existence of costs that penalize excess capacities. Those costs are likely to involve wasted energy or space for large or expensive components, but opportunity costs of wasted space at the molecular level for minor components. PMID- 12122136 TI - Autonomic function in mice lacking alpha5 neuronal nicotinic acetylcholine receptor subunit. AB - Neuronal acetylcholine nicotinic receptors (nAChR) are composed of 12 subunits (alpha2-10, beta2-4), of which alpha3, alpha5, alpha7, beta2 and beta4 subunits are known to exist in the autonomic nervous system (ANS). alpha5 subunits possess unique biophysical and pharmacological properties. The present study was undertaken to examine the functional role and pharmacological properties of the nAChR alpha5 subunits in the ANS using mice lacking alpha5 nAChR subunits (alpha5 /-). These mice grew to normal size showing no obvious physical or neurological deficit. They also showed normality in thermoregulation, pupil size and resting heart rate under physiological conditions. The heart rate and rectal temperature did not differ between alpha5-/- and wild-type mice during exposure to cold stress. An impairment of cardiac parasympathetic ganglionic transmission was observed during high frequency vagal stimulation, which caused cardiac arrest in all wild-type animals while alpha5-/- mice were more resistant. Deficiency of alpha5 subunits strikingly increased the sensitivity to a low concentration of hexamethonium, leading to a nearly complete blockade of bradycardia in response to vagal stimulation. Such a concentration of hexamethonium only slightly depressed the effects of vagal stimulation in control mice. Deficiency of alpha5 subunits significantly increased ileal contractile responses to cytisine and epibatidine. These results suggest that alpha5 subunits may affect the affinity and sensitivity of agonists and antagonists in the native receptors. Previous studies revealed that alpha5 subunits form functional receptors only in combination with other alpha and beta subunits. Thus, the data presented here imply that alpha5 subunits modulate the activity of nAChR in autonomic ganglia in vivo. PMID- 12122137 TI - Reciprocal intraglomerular excitation and intra- and interglomerular lateral inhibition between mouse olfactory bulb mitral cells. AB - How patterns of odour-evoked glomerular activity are transformed into patterns of mitral cell action potentials (APs) in the olfactory bulb is determined by the functional connectivity of the cell populations in the bulb. We have used paired whole-cell voltage recordings from olfactory bulb slices to compare the functional connectivity of mitral cells to the known anatomy of the mitral cell network. Both inhibitory and excitatory coupling were observed between pairs of mitral cells. Inhibitory coupling was seen as an increased frequency of small, asynchronous GABAergic IPSPs following APs in the presynaptic cell. Excitatory coupling was short in latency, beginning about 1.3 ms after the presynaptic AP and was mediated by both NMDA and AMPA receptors. Mitral cell pairs were coupled by excitation if and only if their apical dendrites terminated in the same glomerulus. The excitatory coupling between mitral cells resembles conventional fast synaptic transmission in its time course, amplitude and latency, despite the absence of evidence for anatomically defined synapses between mitral cells. PMID- 12122138 TI - Molecular identification of Kvalpha subunits that contribute to the oxygen sensitive K+ current of chemoreceptor cells of the rabbit carotid body. AB - Rabbit carotid body (CB) chemoreceptor cells possess a fast-inactivating K+ current that is specifically inhibited by hypoxia. We have studied the expression of Kvalpha subunits, which might be responsible for this current. RT-PCR experiments identified the expression of Kv1.4, Kv3.4, Kv4.1 and Kv4.3 mRNAs in the rabbit CB. There was no expression of Kv3.3 or Kv4.2 transcripts. Immunocytochemistry with antibodies to tyrosine hydroxylase (anti-TH) and to specific Kv subunits revealed the expression of Kv3.4 and Kv4.3 in chemoreceptor cells, while Kv1.4 was only found in nerve fibres. Kv4.1 mRNA was also found in chemoreceptor cells following in situ hybridization combined with anti-TH antibody labelling. Kv4.1 and Kv4.3 appeared to be present in all chemoreceptor cells, but Kv3.4 was only expressed in a population of them. Electrophysiological experiments applying specific toxins or antibodies demonstrated that both Kv3.4 and Kv4.3 participate in the oxygen-sensitive K+ current of chemoreceptor cells. However, toxin application experiments confirmed a larger contribution of members of the Kv4 subfamily. [Ca2+]i measurements under hypoxic conditions and immunocytochemistry experiments in dispersed CB cells demonstrated the expression of Kv3.4 and Kv4.3 in oxygen-sensitive cells; the presence of Kv3.4 in the chemoreceptor cell membrane was not required for the response to low PO2. In summary, three Kv subunits (Kv3.4, Kv4.1 and Kv4.3) may be involved in the fast inactivating outward K+ current of rabbit CB chemoreceptor cells. The homogeneous distribution of the Kv4 subunits in chemoreceptor cells, along with their electrophysiological properties, suggest that Kv4.1, Kv4.3, or their heteromultimers, are the molecular correlate of the oxygen-sensitive K+ channel. PMID- 12122139 TI - Spatial characteristics of sarcoplasmic reticulum Ca2+ release events triggered by L-type Ca2+ current and Na+ current in guinea-pig cardiac myocytes. AB - Ca2+ signals in cardiac muscle cells are composed of spatially limited elementary events termed Ca2+ sparks. Several studies have also indicated that Ca2+ signals smaller than Ca2+ sparks can be elicited. These signals have been termed Ca2+ quarks and were proposed to result from the opening of a single Ca2+ release channel of the sarcoplasmic reticulum. We used laser-scanning confocal microscopy to examine the subcellular properties of Na+ current (I(Na))- and L-type Ca2+ current (I(Ca,L))-induced Ca2+ transients in voltage-clamped ventricular myocytes isolated from guinea-pigs. Both currents, I(Na) and I(Ca,L), evoked substantial, global Ca2+ transients. To examine the spatiotemporal properties of such Ca2+ signals, we performed power spectral analysis of these Ca2+ transients and found that both lacked spatial frequency components characteristic for Ca2+ sparks. The application of 10 microM verapamil to partially block L-type Ca2+ current reduced the corresponding Ca2+ transients down to individual Ca2+ sparks. In contrast, I(Na)-induced Ca2+ responses were still spatially homogeneous and lacked Ca2+ sparks even for small current amplitudes. By using high resistance patch pipettes (> 4 MOmega) to exaggerate the loss of voltage control during I(Na), Ca2+ sparks appeared superimposed on a homogeneous Ca2+ release component and were exclusively triggered during the flow of I(Na). In the presence of 10 microM ryanodine both I(Ca,L) and I(Na) elicited small, residual Ca2+ transients that were spatially homogeneous but displayed distinctively different temporal profiles. We conclude that I(Na) is indeed able to cause Ca2+ release in guinea pig ventricular myocytes. In contrast to I(Ca,L)-induced Ca2+ transients, which are built up from the recruitment of individual Ca2+ sparks, the I(Na)-evoked cellular responses were always homogeneous, indicating that their underlying elementary Ca2+ release event is distinct from the Ca2+ spark. Thus, I(Na) induced Ca2+ transients are composed of smaller Ca2+ signals, most likely Ca2+ quarks. PMID- 12122140 TI - Physical mobilization of secretory vesicles facilitates neuropeptide release by nerve growth factor-differentiated PC12 cells. AB - It has been speculated that neurosecretion can be enhanced by increasing the motion, and hence, the availability of cytoplasmic secretory vesicles. However, facilitator-induced physical mobilization of secretory vesicles has not been observed directly in living cells, and recent experimental results call this hypothesis into question. Here, high resolution green fluorescent protein (GFP) based measurements in nerve growth factor-differentiated PC12 cells are used to test whether altering dense core vesicle (DCV) motion affects neuropeptide release. Experiments with mycalolide B and jasplakinolide demonstrate that neuropeptidergic DCV motion at the ends of processes is proportional to F-actin. Furthermore, Ba2+ increases DCV mobility without detectably modifying F-actin. Finally, we show that altering DCV motion by changing F-actin or stimulating with Ba2+ proportionally changes sustained neuropeptide release. Therefore, increasing DCV mobility facilitates prolonged neuropeptide release. PMID- 12122141 TI - Myocardial and skeletal muscle glucose uptake during exercise in humans. AB - The purpose of this study was to investigate the effects of exercise on myocardial glucose uptake and whether the pattern of glucose uptake is the same as in skeletal muscle. Glucose uptake was measured using positron emission tomography (PET) and 2-[18F]fluoro-2-deoxy-D-glucose ([18F]FDG). Twelve healthy men were studied during rest, while 14 subjects were studied after 35 min of bicycle exercise corresponding to 30, 55 and 75 % of maximal oxygen consumption (*VO2,max)) on three separate days. [(18)F]FDG was injected 10 min after the start of exercise and exercise continued for a further 25 min. Myocardial and skeletal muscle PET scanning was commenced directly after the completion of the exercise bout. As compared to the resting state, exercise doubled myocardial glucose uptake at the 30 % (P = 0.056) and 55 % intensity levels (P < 0.05), while at the 75 % intensity level glucose uptake was reduced significantly compared to the lower exercise intensities. There was no significant difference between the highest intensity level and the resting state (P = 0.18). At rest and during low-intensity exercise, myocardial glucose uptake was inversely associated with circulating levels of free fatty acids. However, during higher exercise intensities when plasma lactate concentrations increased significantly, this association disappeared. In contrast to myocardial responses, skeletal muscle glucose uptake rose in parallel with exercise intensity from rest to 30 % and then 55 % *VO2,max) (P < 0.001) and tended to increase further at the intensity of 75 % *VO2,max) (P = 0.065). In conclusion, these results demonstrate that myocardial glucose uptake is increased during mild- and moderate-intensity exercise, but is decreased during high-intensity exercise. This finding suggests that the increased myocardial energy that is needed during high-intensity exercise is supplied by substrates other than glucose. PMID- 12122144 TI - New light shed on fluid formation in the seminiferous tubules of the rat. AB - In this study the effects of perfusing isolated seminiferous tubules of the testes are reported for the first time. Initial perfusion studies (fast rate perfusion) resulted in gross morphological damage to the seminiferous tubules. The recorded transepithelial potential (V(t)) was close to 0 mV. Slow perfusion rates eliminated morphological damage to the perfused tubules. These tubules exhibited a V(t) of -5.4 +/- 1.8 mV which was significantly different (P < 0.0001) from tubules that were perfused at a fast rate. Additional non-perfusion electrophysiological experiments (oil-gap and agar probe techniques) provided the confirmation that tubules not morphologically compromised produced a higher V(t) which was not statistically different (P < 0.0001) from slowly perfused tubules. A revised hypothesis on fluid secretion is postulated. In brief, that the seminiferous tubule is solely responsible for the production of its luminal fluid. This hypothesis is contrary to the long standing 'Tuck' hypothesis which suggested that the source of luminal fluid in the seminiferous tubule originated from secretions of Sertoli cells as well as from distal testicular structures, e.g. the rete testis. PMID- 12122142 TI - Neurotrophin modulation of voltage-gated potassium channels in rat through TrkB receptors is time and sensory experience dependent. AB - The whole-cell configuration of the patch-clamp technique, immunoprecipitation experiments and unilateral naris occlusions were used to investigate whether the voltage-gated potassium channel Kv1.3 was a substrate for neurotrophin-induced tyrosine phosphorylation and subsequent functional modulation of current properties in cultured rat olfactory bulb (OB) neurons. Membrane proteins of the OB included all three Trk receptor kinases, but the truncated form of the receptor, lacking an intact kinase domain, was the predominant form of the protein for TrkA and TrkC, while TrkB was predominantly found as the full-length receptor. Acute (15 min) stimulation of OB neurons with bath application of 50 ng ml(-1) brain-derived neurotrophic factor (BDNF), which is a selective ligand for TrkB, caused suppression of the whole-cell outward current and no changes in the kinetics of inactivation or deactivation. Acute stimulation with either nerve growth factor or neurotrophin-3 failed to evoke any changes in Kv1.3 function in the OB neurons. Chronic exposure to BDNF (days) caused an increase in the magnitude of Kv1.3 current and speeding of the inactivation and deactivation of the channel. Acute BDNF-induced activation of TrkB receptors significantly increased tyrosine phosphorylation of Kv1.3 in the OB, as shown using a combined immunoprecipitation and Western blot analysis. With unilateral naris occlusion, the acute BDNF-induced tyrosine phosphorylation of Kv1.3 was increased in neurons lacking odour sensory experience. In summary, the duration of neurotrophin exposure and the sensory-dependent state of a neuron can influence the degree of phosphorylation of a voltage-gated ion channel and its concomitant functional modulation by neurotrophins. PMID- 12122143 TI - Characterization of four types of background potassium channels in rat cerebellar granule neurons. AB - Cerebellar granule neurons express a standing outward (background) K+ current (I(K,SO)) that regulates the resting membrane potential and cell excitability. As several tandem-pore (2P) K+ channel mRNAs are highly expressed in cerebellar granule cells, we studied whether, and which, 2P K+ channels contribute to I(K,SO). I(K,SO) was highly sensitive to changes in extracellular pH and was partially inhibited by acetylcholine, as reported previously. In cell-attached patches from cultured cerebellar granule neurons, four types of K+ channels were found to be active when membrane potential was held at -50 mV or +50 mV in symmetrical 140 mM KCl. Based on single-channel conductances, gating kinetics and modulation by pharmacological agents and pH, three K+ channels could be considered as functional correlates of TASK-1, TASK-3 and TREK-2, which are members of the 2P K+ channel family. The fourth K+ channel (Type 4) has not been described previously and its molecular correlate is not yet known. Based on the measurement of channel current densities, the Type 2 (TASK-3) and the Type 4 K+ channels were determined to be the major sources of I(K,SO) in cultured cerebellar granule neurons. The Type 1 (TASK-1) and Type 3 (TREK-2) activities were relatively low throughout cell growth in culture (1-10 days). Similar to TASK-1 and TASK-3, the Type 4 K+ channel was highly sensitive to changes in extracellular pH, showing a 78 % inhibition by changing the extracellular pH from 7.3 to 6.3. The results of this study show that three 2P K+ channels and an additional pH-sensing K+ channel (Type 4) comprise the I(K,SO) in cultured cerebellar granule neurons. Our results also show that the high sensitivity of I(K,SO) to extracellular pH comes from the high sensitivity of Type 2 (TASK-3) and Type 4 K+ channels. PMID- 12122145 TI - Stimulation-dependent regulation of the pH, volume and quantal size of bovine and rodent secretory vesicles. AB - Trapping of weak bases was utilized to evaluate stimulus-induced changes in the internal pH of the secretory vesicles of chromaffin cells and enteric neurons. The internal acidity of chromaffin vesicles was increased by the nicotinic agonist 1,1-dimethyl-4-phenyl-piperazinium iodide (DMPP; in vivo and in vitro) and by high K+ (in vitro); and in enteric nerve terminals by exposure to veratridine or a plasmalemmal [Ca2+]o receptor agonist (Gd3+). Stimulation induced acidification of chromaffin vesicles was [Ca2+]o-dependent and blocked by agents that inhibit the vacuolar proton pump (vH+-ATPase) or flux through Cl- channels. Stimulation also increased the average volume of chromaffin vesicles and the proportion that displayed a clear halo around their dense cores (called active vesicles). Stimulation-induced increases in internal acidity and size were greatest in active vesicles. Stimulation of chromaffin cells in the presence of a plasma membrane marker revealed that membrane was internalized in endosomes but not in chromaffin vesicles. The stable expression of botulinum toxin E to prevent exocytosis did not affect the stimulation-induced acidification of the secretory vesicles of mouse neuroblastoma Neuro2A cells. Stimulation-induced acidification thus occurs independently of exocytosis. The quantal size of secreted catecholamines, measured by amperometry in cultured chromaffin cells, was found to be increased either by prior exposure to L-DOPA or stimulation by high K+, and decreased by inhibition of vH+-ATPase or flux through Cl- channels. These observations are consistent with the hypothesis that the content of releasable small molecules in secretory vesicles is increased when the driving force for their uptake is enhanced, either by increasing the transmembrane concentration or pH gradients. PMID- 12122146 TI - Spatial heterogeneity of transmembrane potential responses of single guinea-pig cardiac cells during electric field stimulation. AB - Changes in transmembrane voltage (V(m)) of cardiac cells during electric field stimulation have a complex spatial- and time-dependent behaviour that differs significantly from electrical stimulation of space-clamped membranes by current pulses. A multisite optical mapping system was used to obtain 17 or 25 microm resolution maps of V(m) along the long axis of guinea-pig ventricular cells (n = 57) stained with voltage-sensitive dye (di-8-ANEPPS) and stimulated longitudinally with uniform electric field (2, 5 or 10 ms, 3-62 V cm(-1)) pulses (n = 201). The initial polarizations of V(m) responses (V(mr)) varied linearly along the cell length and reversed symmetrically upon field reversal. The remainder of the V(m) responses had parallel time courses among the recording sites, revealing a common time-varying signal component (V(ms)). V(ms) was depolarizing for pulses during rest and hyperpolarizing for pulses during the early plateau phase. V(ms) varied in amplitude and time course with increasing pulse amplitude. Four types of plateau response were observed, with transition points between the different responses occurring when the maximum polarization at the ends of the cell reached values estimated as 60, 110 and 220 mV. Among the cells that had a polarization change of > 200 mV at their ends (for fields > 45 V cm(-1)), some (n = 17/25) had non-parallel time courses among V(m) recordings of the various sites. This implied development of an intracellular field (E(i)) that was found to increase exponentially with time (tau = 7.2 +/- 3.2 ms). Theoretical considerations suggest that V(ms) represents the intracellular potential (phi(i)) as well as the average polarization of the cell, and that V(mr) is the manifestation of the extracellular potential gradient resulting from the field stimulus. For cells undergoing field stimulation, phi(i) acts as the cellular physiological state variable and substitutes for V(m), which is the customary variable for space-clamped membranes. PMID- 12122147 TI - Loads, capacities and safety factors of maltase and the glucose transporter SGLT1 in mouse intestinal brush border. AB - Safety factors are defined as ratios of biological capacities to prevailing natural loads. We measured the safety factor of the mouse intestinal brush-border hydrolase maltase in series with the glucose transporter SGLT1, for comparison with previous studies of sucrase and lactase. Dietary maltose loads increased 4 fold from virgin to lactating mice. As in previous studies of intestinal adaptive regulation, that increase in load without change in dietary composition resulted in an increase in maltase and SGLT1 capacities mediated non-specifically by an increase in intestinal mass, without change in maltase or SGLT1 activities per milligram of tissue. Maltase and SGLT1 capacities increased only sublinearly with load during lactation, such that safety factors decreased with load: from 6.5 to 2.4 for maltase, and from 1.1 to 0.5 for SGLT1. The apparently high safety factor for maltase may be related to the multiple natural substrates hydrolysed by the multiple sites of maltase activity. The apparently low safety factor for SGLT1 is made possible by the contribution of hindgut fermentation to carbohydrate digestion. SGLT1 activity is paradoxically higher for mice consuming sucrose than for mice consuming maltose, despite maltose hydrolysis yielding double the glucose load yielded by sucrose hydrolysis, and despite glucose constituting the load upon SGLT1. PMID- 12122148 TI - The focal adhesion protein paxillin regulates contraction in canine tracheal smooth muscle. AB - The adapter protein paxillin localizes to the focal adhesions of adherent cells and has been implicated in the regulation of cytoskeletal organization and cell motility. Paxillin undergoes tyrosine phosphorylation in response to the contractile stimulation of tracheal smooth muscle. We therefore hypothesized that paxillin may be involved in regulating smooth muscle contraction. Tracheal smooth muscle strips were treated with paxillin antisense oligonucleotides to inhibit the expression of paxillin protein selectively. Paxillin antisense or sense was introduced into muscle strips by reversible permeabilization and strips were incubated with antisense or sense for 3 days. Paxillin antisense selectively depressed paxillin expression, but it did not affect the expression of vinculin, focal adhesion kinase, myosin light chain kinase, myosin heavy chain or myosin light chain. Tension development in response to stimulation with ACh or KCl was markedly depressed in paxillin-depleted muscle strips. Active force and paxillin protein expression were restored by incubation of antisense-treated strips in the absence of oligonucleotides. The depletion of paxillin did not inhibit the increase in intracellular free Ca2+, myosin light chain phosphorylation or myosin ATPase activity in response to contractile stimulation. The concentration of G actin was significantly lower in unstimulated paxillin-depleted smooth muscle tissues than in normal tissues. While stimulation with acetylcholine caused a decrease in G-actin in normal muscle strips, it caused little change in the G actin concentration in paxillin-depleted muscle strips, suggesting that paxillin is necessary for normal actin dynamics in smooth muscle. We conclude that paxillin is required for active tension development in smooth muscle, but that it does not regulate increases in intracellular Ca2+, myosin light chain phosphorylation or myosin ATPase activity during contractile stimulation. Paxillin may be important in regulating actin filament dynamics and organization during smooth muscle contraction. PMID- 12122149 TI - Independence of extracellular tortuosity and volume fraction during osmotic challenge in rat neocortex. AB - The structural properties of brain extracellular space (ECS) are summarised by the tortuosity (lambda) and the volume fraction (alpha). To determine if these two parameters were independent, we varied the size of the ECS by changing the NaCl content to alter osmolality of bathing media for rat cortical slices. Values of lambda and alpha were extracted from diffusion measurements using the real time ionophoretic method with tetramethylammonium (TMA+). In normal medium (305 mosmol kg(-1)), the average value of lambda was 1.69 and of alpha was 0.24. Reducing osmolality to 150 mosmol kg(-1), increased lambda to 1.86 and decreased alpha to 0.12. Increasing osmolality to 350 mosmol kg(-1), reduced lambda to about 1.67 where it remained unchanged even when osmolality increased further to 500 mosmol kg(-1). In contrast, alpha increased steadily to 0.42 as osmolality increased. Comparison with previously published experiments employing 3000 M(r) dextran to measure lambda, showed the same behaviour as for TMA+, including the same constant lambda in hypertonic media but with a steeper slope in the hypotonic solutions. These data show that lambda and alpha behave differently as the ECS geometry varies. When alpha decreases, lambda increases but when alpha increases, lambda rapidly attains a constant value. A previous model allowing cellular shape to alter during osmotic challenge can account qualitatively for the plateau behaviour of lambda. PMID- 12122150 TI - Ionotropic P2X purinoreceptors mediate synaptic transmission in rat pyramidal neurones of layer II/III of somato-sensory cortex. AB - Fast P2X receptor-mediated excitatory postsynaptic current (EPSC) was identified in pyramidal neurones of layer II/III of somato-sensory cortex in acutely isolated slices obtained from the brain of 17- to 22-day-old rats. The EPSCs were elicited by electrical stimulation of vertical axons originating from layer IV-VI neurones at 0.1 Hz in the presence of bicuculline. When the glutamatergic EPSC was blocked by saturating concentrations of glutamate receptor inhibitors 2,3 dioxo-6-nitro-1,2,3,4-tetrahydrobenzo-[f]-quinoxaline-7-sulphonamide (NBQX) and D (-)-2-amino-5-phosphonopentanoic acid (D-AP5), a small EPSC component was recorded from 90 % of neurones tested. This residual EPSC was not affected by selective blockers of nicotinic (hexamethonium) or serotonin (N-(1-azabicyclo [2.2.2]oct-3-yl)-6-chloro-4-methyl-3-oxo-3,4-dihydro-2H-1,4-benzoxazine-8 carboxamide hydrochloride, Y-25130) receptors, but it was reversibly inhibited by the antagonists of P2X receptors NF023 (8,8'-[carbonylbis(imino-3,1 phenylenecarbonylimino)]bis-1,3,5-naphthalene-trisulphonic acid), NF279 (8,8' [carbonylbis (imino-4,1-phenylenecarbonylimino-4,1-phenylenecarbonylimino)]bis 1,3,5-naphthalene-trisulphonic acid) and PPADS (pyridoxal phosphate-6-azophenyl 2',4'-disulphonic acid). Application of ATP (10 microM) or alpha,beta-methylene ATP (10 microM) to pyramidal neurones, acutely isolated from cortical slices, evoked inward currents (30 to 200 pA) in 65 % of cells tested. The relative calcium/caesium permeability (P(Ca)/P(Cs)) of P2X receptors was 12.3 as estimated from the reversal potential of ATP-induced current measured at different extracellular calcium concentrations. We concluded that P2X purinoreceptors are activated during synaptic transmission in neocortex. PMID- 12122151 TI - Modulation of volume-sensitive chloride current by noradrenaline in rabbit portal vein myocytes. AB - The effect of noradrenaline on the volume-sensitive chloride current (I(Cl(swell))) was studied with conventional whole-cell recording techniques in freshly dispersed isolated smooth muscle cells of the rabbit portal vein. In the absence of receptor antagonists, noradrenaline produced an increase in the amplitude of I(Cl(swell)) in some cells and a decrease in others. In the presence of the beta-adrenoceptor antagonist propranolol, noradrenaline increased I(Cl(swell)) and in the presence of the alpha(1)-adrenoceptor antagonist prazosin, noradrenaline reduced I(Cl(swell).) The phospholipase C (PLC) inhibitor U73122 reduced the amplitude of I(Cl(swell)) whereas the inactive analogue U73343 had no effect. The phorbol esters phorbol-12-myristate-13-acetate (PMA) and phorbol-12,13-dibutyrate (PDBu) increased the amplitude of I(Cl(swell)) by approximately 60 and 100 %, respectively, in a voltage-independent fashion. Inhibitors of protein kinase C (PKC) chelerythrine and calphostin-C decreased the amplitude of I(Cl(swell)) in a concentration-dependent but voltage-independent manner. Bath application of 8-Br-cAMP decreased I(Cl(swell)) by about 60 % whereas the inhibitor of protein kinase A (PKA) KT5720 increased the amplitude of I(Cl(swell)) by approximately 80-90 %. In the presence of propranolol, chelerythrine prevented the increase of I(Cl(swell)) by noradrenaline; in the presence of prazosin, KT5720 blocked the inhibitory action of noradrenaline. The results show that in rabbit portal vein myocytes noradrenaline enhances I(Cl(swell)) by acting on alpha(1)-adrenoceptors and reduces I(Cl(swell)) by stimulating beta-adrenoceptors. The data suggest that the potentiating and inhibitory effects of noradrenaline are mediated, respectively, by PKC and PKA. PMID- 12122152 TI - Break excitation alone does not explain the delay and amplitude of anodal current induced vasodilatation in human skin. AB - In iontophoresis experiments, a 'non-specific' current-induced vasodilatation interferes with the effects of the diffused drugs. This current-induced vasodilatation is assumed to rely on an axon reflex due to excitation of cutaneous nociceptors and is weaker and delayed at the anode as compared to the cathode. We analysed whether these anodal specificities could result from a break excitation of nociceptors. Break excitation is the generation of action potentials at the end of a square anodal DC current application, which are generally weaker than those observed at the onset of a same application at the cathode. In eight healthy volunteers, we studied forearm cutaneous laser Doppler flow (LDF) responses to: (1) anodal and cathodal 100 microA current applications of 1, 2, 3, 4 or 5 min; (2) 100 microA anodal applications of 3 min with a progressive ending over 100 s (total charge 23 mC); these were compared to square ended 100 microA anodal applications of the same total charge (23 mC) or duration (3 min); (3) a 4 min 100 microA anodal application with a 333 msec break at half time. Results (mean +/- S.D.) are expressed as percentage of heat-induced maximal vasodilatation (%MVD). Onset (T(vd)) and amplitude (LDF(peak)) of vasodilatation were determined. We observed that: T(vd) was linearly related to the duration of current application at the anode (slope = 1.01, r(2) = 0.99, P < 0.0001) but not at the cathode (slope = 0.03, r(2) = 0.02, n.s.). Progressive ending of anodal current did not decrease LDF(peak) (63.3 +/- 24.6 %MVD) as compared to square ending of current application of the same duration (36.9 +/- 22.2 %MVD) or the same total charge (57.1 +/- 23.5 %MVD). A transient break of anodal current did not allow for the vasodilatation to develop until current was permanently stopped. We conclude that, during iontophoresis, anodal break excitation alone cannot account for the delay and amplitude of the vascular response. PMID- 12122153 TI - Finger movement is associated with attenuated cutaneous reflexes recorded from human first dorsal interosseous muscle. AB - Cutaneomuscular reflexes (CMR) have been recorded from the first dorsal interosseous muscle (1DI) of the preferred hand, somatosensory evoked potentials (SEP) were recorded from the contralateral sensory cortex and the sensory nerve action potential (SNAP) was recorded from the median nerve of 15 adult subjects whilst electrically stimulating the digital nerves of the index finger. Subjects performed the following tasks (a) a sustained abduction of the index finger against resistance at 10-20 % maximum voluntary contraction (MVC), and (b) abduction of the index finger as in (a) whilst performing self paced low amplitude tapping of the (i) index finger, (ii) thumb, (iii) middle finger, (iv) little finger and (v) ipsilateral foot. The E2 component of the CMR and the N20/P25 components of the SEP were significantly reduced during finger tapping (P < 0.05). This reduction was independent of which finger was tapping (P > 0.05). There was a significant (qualitative) relationship between the decrease in the size of the E2 component of the CMR and the N20/P25 components of the SEP ((2) test; P < 0.05). There were no significant changes in E1 and I1 (P > 0.05). The size of the SNAP was independent of task (P > 0.05). The size of the E1, I1, E2 components of the CMR, and the N20/P25 components of the SEP were unaltered during foot tapping (P > 0.05, n = 4). We conclude that the decrease in size of the E2 component associated with finger tapping results from gating of the digital nerve input. PMID- 12122154 TI - Synchronized oscillations caused by disinhibition in rodent neocortex are generated by recurrent synaptic activity mediated by AMPA receptors. AB - During disinhibition the neocortex generates synchronous activities. In neocortical slices application of GABA(A) and GABA(B) receptor antagonists transformed slow oscillations into large amplitude spike-wave discharges that contained a rhythmic ~10 Hz neocortical oscillation. The 10 Hz oscillations caused by disinhibition were highly region specific and were generated only in frontal agranular regions of neocortex, such as the primary motor cortex, but not in granular neocortex. Pharmacological manipulations showed that the 10 Hz oscillations were critically dependent on alpha-amino-3-hydroxy-5-methylisoxazole 4-propionate (AMPA) receptors. Current source density (CSD) analyses in slices using 16-site silicon probes revealed that the 10 Hz oscillations were expressed with large current sinks in the upper layers and smaller current sinks in the lower layers that precede them. The results indicate that blocking GABA(B) receptors in the agranular neocortex unmasks recurrent synaptic activity mediated by AMPA receptors that results in the generation of these oscillations. PMID- 12122155 TI - Short- and medium-term plasticity associated with augmenting responses in cortical slabs and spindles in intact cortex of cats in vivo. AB - Plastic changes in the synaptic responsiveness of neocortical neurones, which occur after rhythmic stimuli within the frequency range of sleep spindles (10 Hz), were investigated in isolated neocortical slabs and intact cortex of anaesthetized cats by means of single, dual and triple simultaneous intracellular recordings in conjunction with recordings of local field potential responses. In isolated cortical slabs (10 mm long, 6 mm wide and 4-5 mm deep), augmenting responses to pulse-trains at 10 Hz (responses with growing amplitudes from the second stimulus in a train) were elicited only by relatively high-intensity stimuli. At low intensities, responses were decremental. The largest augmenting responses were evoked in neurones located close to the stimulation site. Quantitative analyses of the number of action potentials and the amplitude and area of depolarization during augmenting responses in a population of neurones recorded from slabs showed that the most dramatic increases in the number of spikes with successive stimuli, and the greatest increase in depolarization amplitude, were found in conventional fast-spiking (FS) neurones. The largest increase in the area of depolarization was found in regular-spiking (RS) neurones. Dual intracellular recordings from a pair of FS and RS neurones in the slab revealed more action potentials in the FS neurone during augmenting responses and a significant increase in the depolarization area of the RS neurone that was dependent on the firing of the FS neurone. Self-sustained seizures could occur in the slab after rhythmic stimuli at 10 Hz. In the intact cortex, repeated sequences of stimuli generating augmenting responses or spontaneous spindles could induce an increased synaptic responsiveness to single stimuli, which lasted for several minutes. A similar time course of increased responsiveness was obtained with induction of cellular plasticity. These data suggest that augmenting responses elicited by stimulation, as well as spontaneously occurring spindles, may induce short- and medium-term plasticity of neuronal responses. PMID- 12122156 TI - Frequency-selective augmenting responses by short-term synaptic depression in cat neocortex. AB - Thalamic stimulation at frequencies between 5 and 15 Hz elicits incremental or 'augmenting' cortical responses. Augmenting responses can also be evoked in cortical slices and isolated cortical slabs in vivo. Here we show that a realistic network model of cortical pyramidal cells and interneurones including short-term plasticity of inhibitory and excitatory synapses replicates the main features of augmenting responses as obtained in isolated slabs in vivo. Repetitive stimulation of synaptic inputs at frequencies around 10 Hz produced postsynaptic potentials that grew in size and carried an increasing number of action potentials resulting from the depression of inhibitory synaptic currents. Frequency selectivity was obtained through the relatively weak depression of inhibitory synapses at low frequencies, and strong depression of excitatory synapses together with activation of a calcium-activated potassium current at high frequencies. This network resonance is a consequence of short-term synaptic plasticity in a network of neurones without intrinsic resonances. These results suggest that short-term plasticity of cortical synapses could shape the dynamics of synchronized oscillations in the brain. PMID- 12122157 TI - Physiological basis of fractal complexity properties of heart rate variability in man. AB - The diagnostic and prognostic power of the fractal complexity measure 'alpha' of detrended fluctuation analysis (DFA) has remained mysterious because there has been no explanation of its meaning, particularly in relation to spectral analysis. First, we present a mathematical analysis of the meaning of alpha, in weighted power-spectral terms. Second, we test this hypothesis and observe correlations between DFA-based and weighted spectral methods of 0.97 (P < 0.0001) for alpha1 and 0.98 (P < 0.0001) for alpha2. Third, we predict mathematically that even in conventional (unweighted) spectral analysis there should be approximate counterparts to DFA, namely that alpha1 and alpha2 behave broadly in proportion to the conventional (unweighted) ratios LF/(HF + LF) and VLF/(LF + VLF), respectively, where HF is high frequency, LF is low frequency and VLF is very low frequency. Fourth, we test this hypothesis by physiologically manipulating spectral ratios in healthy volunteers in two ways. The effect of 0.1 Hz controlled breathing on LF/(HF + LF) correlates markedly with the effect on alpha1 (r = 0.73, P = 0.01); the effect on VLF/(LF + VLF) correlates markedly with that on alpha2 (r = 0.76, P < 0.01). Likewise, with voluntary periodic breathing the reduction in alpha2 correlates strongly with that in VLF/(LF + VLF) (r = 0.88, P < 0.001); effects on alpha1 and LF/(HF + LF) again clearly correlate (r = 0.73, P = 0.01). Finally, we examine published literature to identify previously undiscussed evidence of the relationship between alpha1 and LF/(HF + LF). We conclude that the alpha1 and alpha2 indices are simply frequency-weighted versions of the spectral ratios LF/(HF + LF) and VLF/(LF + VLF), respectively, multiplied by two (giving a range of 0-2). We can now understand fractal manifestations of physiological abnormalities: depressed baroreflex sensitivity low LF/HF low LF/(HF + LF) low alpha1, while periodic breathing high VLF/LF high VLF/(LF + VLF) high alpha2. Prognostic associations of alpha are no longer mysterious. PMID- 12122158 TI - Dietary protein adequacy and lower body versus whole body resistive training in older humans. AB - This study assessed the effects of long-term consumption of the United States Recommended Dietary Allowance (RDA) for protein by older people who were sedentary or performed resistive training (RT) on body composition, skeletal muscle size and protein metabolism, and if the number of muscle groups trained influenced the muscle hypertrophy response to RT. Twelve men and 17 women (age range 54-78 years) completed this 14 week controlled diet and exercise study. Throughout the study, each subject completely consumed daily euenergetic menus that provided the RDA of 0.8 g protein kg(-1) day(-1). From study weeks 3-14 (weeks RT1-RT12), 10 subjects (four men, six women) performed whole body RT (WBRT), nine subjects (four men, five women) performed lower body RT (LBRT) and 10 subjects (four men, six women) remained sedentary (SED). Both the LBRT and WBRT groups performed knee extension and flexion exercises, and the WBRT group also performed chest press and arm pull exercises (three sets per exercise at 80 % of one repetition maximum, 3 days per week for 12 weeks). From week 2 (baseline) to week RT12, muscle strength increased in muscle groups trained in the LBRT and WBRT groups, and was not changed in the SED group. From baseline to week RT12, whole body muscle mass and protein-mineral mass were not changed, fat free mass (P = 0.004) and total body water (P = 0.013) were decreased, and percentage body fat was increased (P = 0.011) in these weight-stable older people, independent of group assignment. The RT-induced increases in mid-thigh muscle area (from computed tomography scans) were comparable in the LBRT and WBRT groups (2.13 +/- 1.26 cm(2) and 2.17 +/- 1.24 cm(2), respectively), and were different from those in the SED group, which lost muscle area (-1.74 +/- 0.57 cm(2); group-by-time P < 0.05). From baseline to week RT12, 24 h urinary total nitrogen excretion decreased (P < 0.001), nitrogen balance shifted from near equilibrium to positive, whole body leucine oxidation (from the infusion of L [(13)C]leucine) decreased (P < 0.05) and net (postabsorptive vs. postprandial) leucine balance (P < 0.05) increased from near equilibrium to positive, with no differences in responses over time among the three groups. In conclusion, the number of muscle groups trained did not influence whole body protein metabolism or RT-induced muscle hypertrophy in older people. Most of these data are consistent with a successful adaptation to the RDA for protein. However, research should continue to question whether the decreases in fat-free mass and total body water observed in all subjects, and the decrease in mid-thigh muscle area in the SED group, are physiological accommodations, and whether the RDA for protein might be marginally inadequate for older people to maintain skeletal muscle. PMID- 12122160 TI - Envenomation and consumption of poisonous seafood. PMID- 12122161 TI - Neutralising antibodies to the beta interferons. PMID- 12122163 TI - Functional reorganisation of memory following traumatic brain injury: a study with H(2)(15)O PET. PMID- 12122159 TI - Age-dependent variations in the directional sensitivity of balance corrections and compensatory arm movements in man. AB - We investigated the effects of ageing on balance corrections induced by sudden stance perturbations in different directions. Effects were examined in biomechanical and electromyographic (EMG) recordings from a total of 36 healthy subjects divided equally into three age groups (20-34, 35-55 and 60-75 years old). Perturbations consisted of six combinations of support-surface roll (laterally) and pitch (forward-backward) each with 7.5 deg amplitude (2 pure pitch, and 4 roll and pitch) delivered randomly. To reduce stimulus predictability further and to investigate scaling effects, perturbations were at either 30 or 60 deg s(-1). In the legs, trunk and arms we observed age-related changes in balance corrections. The changes that appeared in the lower leg responses included smaller stretch reflexes in soleus and larger reflexes in tibialis anterior of the elderly compared with the young. For all perturbation directions, onsets of balance correcting responses in these ankle muscles were delayed by 20-30 ms and initially had smaller amplitudes (between 120-220 ms) in the elderly. This reduced early activity was compensated by increased lower leg activity after 240 ms. These EMG changes were paralleled by comparable differences in ankle torque responses, which were initially (after 160 ms) smaller in the elderly, but subsequently greater (after 280 ms). Findings in the middle-aged group were generally intermediate between the young and the elderly groups. Comparable results were obtained for the two different stimulus velocities. Stimulus-induced trunk roll, but not trunk pitch, changed dramatically with increasing age. Young subjects responded with early large roll movements of the trunk in the opposite direction to platform roll. A similarly directed but reduced amplitude of trunk roll was observed in the middle-aged. The elderly had very little initial roll modulation and also had smaller stretch reflexes in paraspinals. Balance-correcting responses (over 120-220 ms) in gluteus medius and paraspinals were equally well tuned to roll in the elderly, as in the young, but were reduced in amplitude. Onset latencies were delayed with age in gluteus medius muscles. Following the onset of trunk and hip balance corrections, trunk roll was in the same direction as support-surface motion for all age groups and resulted in overall trunk roll towards the fall side in the elderly, but not in the young. Protective arm movements also changed with age. Initial arm roll movements were largest in the young, smaller in the middle aged, and smallest in the elderly. Initial arm roll movements were in the same direction as initial trunk motion in the young and middle aged. Thus initial roll arm movements in the elderly were directed oppositely to those in the young. Initial pitch motion of the arms was similar across age groups. Subsequent arm movements were related to the amplitude of deltoid muscle responses which commenced at 100 ms in the young and 20-30 ms later in the elderly. These deltoid muscle responses preceded additional arm roll motion which left the arms directed 'downhill' (in the direction of the fall) in the elderly, but 'uphill' (to counterbalance motion of the pelvis) in the young. We conclude that increased trunk roll stiffness is a key biomechanical change with age. This interferes with early compensatory trunk movements and leads to trunk displacements in the direction of the impending fall. The reversal of protective arm movements in the elderly may reflect an adaptive strategy to cushion the fall. The uniform delay and amplitude reduction of balance-correcting responses across many segments (legs, hips and arms) suggests a neurally based alteration in processing times and response modulation with age. Interestingly, the elderly compensated for these 'early abnormalities' with enlarged later responses in the legs, but no similar adaptation was noted in the arms and trunk. These changes with age provide an insight into possible mechanisms underlying falls in the elderly. PMID- 12122164 TI - Coffee and subarachnoid haemorrhage. PMID- 12122165 TI - The palmomental reflex: a useful clinical sign? AB - The palmomental reflex, an involuntary contraction of the mentalis muscle of the chin caused by stimulation of the thenar eminence, can be tested easily and rapidly. Its presence may alert the clinician to the possibility of cerebral pathology. However, the reflex is often present in normal people and may be absent in disease states. Testing merely for the presence or absence of the reflex therefore lacks both specificity and sensitivity. A strong, sustained, and easily repeatable contraction of the mentalis muscle, which can be elicited by stimulation of areas other than the palm, is more likely to indicate cerebral damage. PMID- 12122166 TI - Clinically important change in quality of life in epilepsy. AB - BACKGROUND: Health related quality of life (HRQOL) is increasingly recognised as an important outcome in epilepsy. However, interpretation of HRQOL data is difficult because there is no agreement on what constitutes a clinically important change in the scores of the various instruments. OBJECTIVES: To determine the minimum clinically important change, and small, medium, and large changes, in broadly used epilepsy specific and generic HRQOL instruments. METHODS: Patients with difficult to control focal epilepsy (n = 136) completed the QOLIE-89, QOLIE-31, SF-36, and HUI-III questionnaires twice, six months apart. Patient centred estimates of minimum important change, and of small, medium, and large change, were assessed on self administered 15 point global rating scales. Using regression analysis, the change in each HRQOL instrument that corresponded to the various categories of change determined by patients was obtained. The results were validated in a subgroup of patients tested at baseline and at nine months. RESULTS: The minimum important change was 10.1 for QOLIE-89, 11.8 for QOLIE-31, 4.6 for SF-36 MCS, 3.0 for SF-36 physical composite score, and 0.15 for HUI-III. All instruments differentiated between no change and minimum important change with precision, and QOLIE-89 and QOLIE-31 also distinguished accurately between minimum important change and medium or large change. Baseline HRQOL scores and the type of treatment (surgical or medical) had no impact on any of the estimates, and the results were replicated in the validation sample. CONCLUSIONS: These estimates of minimum important change, and small, medium, and large changes, in four HRQOL instruments in patients with epilepsy are robust and can distinguish accurately among different levels of change. The estimates allow for categorisation of patients into various levels of change in HRQOL, and will be of use in assessing the effect of interventions in individual patients. PMID- 12122167 TI - Reproductive dysfunction in women with epilepsy: recommendations for evaluation and management. AB - BACKGROUND: Epilepsy is commonly associated with reproductive endocrine disorders. These include polycystic ovary syndrome (PCOS), isolated components of this syndrome such as polycystic ovaries, hyperandrogenaemia, hypothalamic amenorrhoea, and functional hyperprolactinaemia. OBJECTIVE: To summarise the currently known relations between epilepsy and reproductive endocrine disorders. METHODS: A review of clinical experience and published reports. RESULTS: The most likely explanations for endocrine disorders related to epilepsy or antiepileptic drugs are: (1) a direct influence of the epileptogenic lesion, epilepsy, or antiepileptic drugs on the endocrine control centres in the brain; (2) the effects of antiepileptic drugs on peripheral endocrine glands; (3) the effects of antiepileptic drugs on the metabolism of hormones and binding proteins; and (4) secondary endocrine complications of antiepileptic drug related weight changes or changes of insulin sensitivity. Regular monitoring of reproductive function at visits is recommended, including questioning about menstrual disorders, fertility, weight, hirsutism, and galactorrhoea. Particular attention should be paid to patients on valproate and obese patients or those experiencing significant weight gain. Single abnormal laboratory or imaging findings without symptoms may not constitute a clinically relevant endocrine disorder. However, patients with these kinds of abnormalities should be monitored to detect the possible development of a symptomatic disorder associated with, for example, menstrual disorders or fertility problems. CONCLUSIONS: If a reproductive endocrine disorder is found, antiepileptic drug treatment should be reviewed to ensure that it is correct for the particular seizure type and that it is not contributing to the endocrine problem. The possible benefits of a change in treatment must be balanced against seizure control and the cumulative side effect of alternative agents. PMID- 12122168 TI - Visual association test to detect early dementia of the Alzheimer type. AB - BACKGROUND: The visual association test (VAT) is a brief learning task based on imagery mnemonics. The test materials consist of six line drawings of pairs of interacting objects or animals-for example, an ape holding an umbrella. The person is asked to name each object and, later, is presented with one object from the pair and asked to name the other. OBJECTIVE: To verify that the task induces robust incidental or effortless learning (study 1), and to study the efficiency of the test as a discriminator between early dementia of the Alzheimer type (DAT) and non-demented people (study 2) and non-DAT types of dementia (study 3). METHODS: Study 1: two groups of elderly volunteers were administered the VAT. The stimuli were presented in the interactive fashion to group A-for example, a monkey carrying an umbrella (n=83)-and side by side to group B-for example, separate pictures of a monkey alone and an umbrella alone (n=79). Group B received learning instructions, but group A did not. Study 2: three groups of subjects were selected from a population based follow up study: incident DAT cases (n=24), cognitively declining subjects not diagnosed with dementia (n=21), and stable non-demented subjects (n=204). Test performance of the non-demented group at baseline was compared with that of patients with DAT at the time of their diagnosis, of patients with DAT a year before their diagnosis, and of non demented declining subjects at baseline. Study 3: subjects were patients referred for neuropsychological assessment because of suspected dementia. They were diagnosed by consensus criteria of various dementia syndromes. RESULTS: Study 1: recall was more than twice as high in group A as in group B. Thus interactive presentation, even in the absence of learning instructions, enhances learning. Study 2: at a level of 97.5% specificity, the VAT had a sensitivity of 87.5% for DAT cases at the time of diagnosis and 66.7% one year before diagnosis. The cognitively declining group scored significantly lower on the VAT at baseline than the non-demented group. The VAT discriminated more effectively than both the MMSE and the six item picture learning task from the CAMCOG. Study 3: VAT scores were significantly lower in patients with DAT (n=48) than in patients with vascular dementia (n=37), frontotemporal dementia (n=9), or subcortical dementia (n=15), but not lower than in patients with Lewy body dementia (n=7). Mean mini mental state examination scores of these groups were not significantly different. The VAT discriminated patients with DAT from patients with other types of dementia more effectively than a prose recall test. Sensitivity was 79% and specificity 69%. CONCLUSIONS: The VAT detects with high specificity a sizeable proportion of patients with DAT a year before the diagnosis, and a low VAT score is relatively uncommon in patients with non-DAT dementia. PMID- 12122169 TI - Differentiation of dementia with Lewy bodies from Alzheimer's disease using a dopaminergic presynaptic ligand. AB - BACKGROUND: Dementia with Lewy bodies (DLB) is one of the main differential diagnoses of Alzheimer's disease (AD). Key pathological features of patients with DLB are not only the presence of cerebral cortical neuronal loss, with Lewy bodies in surviving neurones, but also loss of nigrostriatal dopaminergic neurones, similar to that of Parkinson's disease (PD). In DLB there is 40-70% loss of striatal dopamine. OBJECTIVE: To determine if detection of this dopaminergic degeneration can help to distinguish DLB from AD during life. METHODS: The integrity of the nigrostriatal metabolism in 27 patients with DLB, 17 with AD, 19 drug naive patients with PD, and 16 controls was assessed using a dopaminergic presynaptic ligand, (123)I-labelled 2beta-carbomethoxy-3beta-(4 iodophenyl)-N-(3-fluoropropyl)nortropane (FP-CIT), and single photon emission tomography (SPET). A SPET scan was carried out with a single slice, brain dedicated tomograph (SME 810) 3.5 hours after intravenous injection of 185 MBq FP CIT. With occipital cortex used as a radioactivity uptake reference, ratios for the caudate nucleus and the anterior and posterior putamen of both hemispheres were calculated. All scans were also rated by a simple visual method. RESULTS: Both DLB and PD patients had significantly lower uptake of radioactivity than patients with AD (p<0.001) and controls (p<0.001) in the caudate nucleus and the anterior and posterior putamen. CONCLUSION: FP-CIT SPET provides a means of distinguishing DLB from AD during life. PMID- 12122170 TI - Progressive ventricular enlargement in patients with clinically isolated syndromes is associated with the early development of multiple sclerosis. AB - BACKGROUND: In patients with clinically isolated syndromes (CIS) suggestive of multiple sclerosis (MS), the extent of brain magnetic resonance imaging (MRI) lesion load influences the probability and time to development of clinically definite MS. Cerebral atrophy is recognised in established MS, but its time of onset and whether, in early disease, it is related to MRI lesion load or clinical outcome is less certain. OBJECTIVES: This study investigated ventricular enlargement over one year in CIS patients and explored its relation with lesion load and clinical outcome. METHODS: A semi-automated thresholding technique for measuring ventricular volume (MIDAS) was applied to MRI scans in a cohort of 55 patients with CIS, recruited consecutively and imaged within three months of the onset of symptoms and again after one year. RESULTS: Clinical MS had developed after one year in 16 of 40 patients with an abnormal baseline T2 scan and 2 of 15 with a normal scan. Significant ventricular enlargement was seen in 27 of 55 patients who fulfilled the new McDonald MRI criteria for MS using all available MRI at clinical follow up (median increase 0.3 cm(3), p=0.005) Significant increase in ventricular volume was also seen in the 18 of 55 patients who developed clinical MS over the follow up period (median increase 0.5 cm(3), p=0.006). There were significant but modest correlations between baseline lesion measures and subsequent ventricular enlargement. CONCLUSIONS: (1) Lesions and atrophy are both associated with early relapse leading to a diagnosis of clinical MS; (2) while lesions contribute to the development of atrophy, atrophy may also develop by other mechanisms. This suggests that MR measures have a complementary role in monitoring the course of MS, even from the earliest clinical stage. PMID- 12122172 TI - Differential effects of three interferon betas on neutralising antibodies in patients with multiple sclerosis: a follow up study in an independent laboratory. AB - OBJECTIVE: To evaluate the incidence and the prevalence of neutralising antibodies (NABs) to three interferon beta (IFNbeta) products in patients with multiple sclerosis (MS). METHODS: Sera were tested from 125 patients with relapsing-remitting MS. Patients were treated with IFNbeta-1b (Betaferon, n = 29) 8 MIU subcutaneously every other day, IFNbeta-1a (Avonex, n = 44) 30 microg intramuscularly once weekly, or IFNbeta-1a (Rebif, n = 36) 22 microg subcutaneously three times weekly for 6 to 18 months. An additional 16 patients were treated with Rebif 22 microg intramuscularly once or twice weekly. NABs were assessed using the cytopathic effect assay before treatment and every three months during treatment. Patients with two or more consecutive positive samples were considered to be persistent NAB positive (NAB+). RESULTS: At baseline, no patients were NAB+. NABs developed during the first three months of treatment and continued to develop until month 18. Over 18 months of treatment, the risk of being persistent NAB+ was 31% for Betaferon, 15% for Rebif, and 2% for Avonex (Betaferon versus Avonex, p = 0.001; Betaferon versus Rebif, p = 0.19; Rebif versus Avonex, p = 0.04). In all patients with one or more NAB+ samples, the risk of becoming NAB+ was 38% for Betaferon, 18% for Rebif, and 7% for Avonex (Betaferon versus Avonex, p = 0.0007; Betaferon versus Rebif, p = 0.10; Rebif versus Avonex, p = 0.07). At month 18, the prevalence of persistent NAB+ patients was 31.6% for Betaferon, 18.7% for Rebif, and 4% for Avonex. Numbers of NAB+ patients observed were similar with intramuscular Rebif and with subcutaneous Rebif. CONCLUSION: The three IFNbeta preparations have different degrees of immunogenicity, with Betaferon producing the highest incidence of NABs and Avonex the lowest. These differences should be considered by neurologists when selecting treatment for their patients with MS because NABs can reduce both bioavailability and clinical efficacy of IFNbeta. PMID- 12122175 TI - Sensitivity of clinical and behavioural tests of spatial neglect after right hemisphere stroke. AB - OBJECTIVES: The lack of agreement regarding assessment methods is responsible for the variability in the reported rate of occurrence of spatial neglect after stroke. The aim of this study was to assess the sensitivity of different tests of neglect after right hemisphere stroke. METHODS: Two hundred and six subacute right hemisphere stroke patients were given a test battery including a preliminary assessment of anosognosia and of visual extinction, a clinical assessment of gaze orientation and of personal neglect, and paper and pencil tests of spatial neglect in the peripersonal space. Patients were compared with a previously reported control group. A subgroup of patients (n=69) received a behavioural assessment of neglect in daily life situations. RESULTS: The most sensitive paper and pencil measure was the starting point in the cancellation task. The whole battery was more sensitive than any single test alone. About 85% of patients presented some degree of neglect on at least one measure. An important finding was that behavioural assessment of neglect in daily life was more sensitive than any other single measure of neglect. Behavioural neglect was considered as moderate to severe in 36% of cases. A factorial analysis revealed that paper and pencil tests were related to two underlying factors. Dissociations were found between extrapersonal neglect, personal neglect, anosognosia, and extinction. Anatomical analyses showed that neglect was more common and severe when the posterior association cortex was damaged. CONCLUSIONS: The automatic rightward orientation bias is the most sensitive clinical measure of neglect. Behavioural assessment is more sensitive than any single paper and pencil test. The results also support the assumption that neglect is a heterogeneous disorder. PMID- 12122174 TI - Bladder dysfunction in acute transverse myelitis: magnetic resonance imaging and neurophysiological and urodynamic correlations. AB - AIMS: To evaluate micturition abnormalities in acute transverse myelitis and correlate these with evoked potentials, magnetic resonance imaging (MRI), and urodynamic findings. SETTING: Tertiary care teaching hospital. PATIENTS: 18 patients with acute transverse myelitis, aged 4-50 years; 15 had paraparesis and three quadriparesis. METHODS: Patients with acute transverse myelitis had a neurological evaluation and tibial somatosensory and motor evoked potential studies in the lower limbs. Spinal MRI was carried out using a 1.5 T scanner. Urodynamic studies were done using Dantec UD 5500 equipment. Neurological outcome was determined on the basis of Barthel index score at six months as poor, partial, or complete. In some patients, urodynamic studies were repeated at six and 12 months. RESULTS: Spinal MRI in 14 of the 18 patients revealed T2 hyperintense signal changes extending for at least three spinal segments in 13; one patient had normal MRI. In the acute stage, 17 patients had a history of urinary retention and one had urge incontinence. On follow up at six months two patients regained normal voiding, retention persisted in six, and storage symptoms developed in 10, of whom five also had emptying difficulties. Urodynamic studies showed an areflexic or hypocontractile bladder in 10, detrusor hyperreflexia with poor compliance in two, and detrusor sphincter dyssynergia in three. Early abnormal urodynamic findings commonly persisted at the six and 12 months examinations. Persistent abnormalities included detrusor hyperreflexia, dyssynergia, and areflexic bladder. The urodynamic abnormalities correlated with muscle tone and reflex changes but not with sensory or motor evoked potentials, muscle power, MRI signal changes, sensory level, or six months outcome. CONCLUSIONS: Bladder dysfunction is common in acute transverse myelitis and may be the only sequel. Urodynamic study is helpful in evaluating the bladder dysfunction and also in its management. PMID- 12122178 TI - Diffusion weighted magnetic resonance imaging in a case of acute Wernicke's encephalopathy. PMID- 12122176 TI - Effectiveness of bed rest after mild traumatic brain injury: a randomised trial of no versus six days of bed rest. AB - BACKGROUND: Outcome after mild traumatic brain injury (MTBI) is determined largely by the appearance of post-traumatic complaints (PTC). The prevalence of PTC after six months is estimated to be between 20 and 80%. Bed rest has been advocated to prevent PTC but its effectiveness has never been established. OBJECTIVE: To evaluate the effect of bed rest on the severity of PTC after MTBI. METHODS: Patients presenting with MTBI to the emergency room were randomly assigned to two intervention strategies. One group was advised not to take bed rest (NO) and the other to take full bed rest (FULL) for six days after the trauma. The primary outcome measures were severity of PTC on a visual analogue scale and physical and mental health on the medical outcomes study 36 item short form health survey (SF-36) at two weeks and three and six months after the trauma. RESULTS: Between October 1996 and July 1999, 107 (54 NO, 53 FULL) patients were enrolled. Outcome variables in both groups clearly improved between two weeks and six months. After adjustment for differences in baseline variables, most PTC tended to be somewhat more severe in the FULL group six months after the trauma, but no significant differences were found. Neither were there any significant differences in the outcome parameters between the two groups after three months. Two weeks after the trauma, most PTC in the FULL group were slightly less severe than those in the NO group, and physical subscores of the SF 36 in the FULL group were slightly better. These differences were not significant. Patients in the FULL group reported significantly less dizziness during the intervention period. CONCLUSIONS: As a means of speeding up recovery of patients with PTC after MTBI, bed rest is no more effective than no bed rest at all. Bed rest probably has some palliative effect within the first two weeks after the trauma. PMID- 12122177 TI - Functional reorganisation of memory after traumatic brain injury: a study with H(2)(15)0 positron emission tomography. AB - OBJECTIVE: To study the effects of moderate to severe traumatic brain injury (TBI) on the functional neuroanatomy supporting memory retrieval. METHODS: Subjects were six patients who had sustained a moderate to severe TBI about four years before scanning and had since made a good recovery. Eleven healthy young adults matched to the patients for age and education served as controls. An established H(2)(15)0 positron emission tomography paradigm was used to elicit brain activations in response to memory retrieval. TBI patients' patterns of brain activation were compared statistically with those of control subjects. Both group and individual case data were analysed. RESULTS: Both TBI patients and controls engaged frontal, temporal, and parietal regions known to be involved in memory retrieval, yet the TBI patients showed relative increases in frontal, anterior cingulate, and occipital activity. The hemispheric asymmetry characteristic of controls was attenuated in patients with TBI. Reduced activation was noted in the right dorsomedial thalamus. Although local aspects of this pattern were affected by the presence of focal lesions and performance differences, the overall pattern was reliable across patients and comparable to functional neuroimaging results reported for normal aging, Alzheimer's disease, and other patients with TBI. CONCLUSIONS: The TBI patients performed memory tasks using altered functional neuroanatomical networks. These changes are probably the result of diffuse axonal injury and may reflect either cortical disinhibition attributable to disconnection or compensation for inefficient mnemonic processes. PMID- 12122179 TI - Increase in focal concentration of deoxyhaemoglobin during neuronal activity in cerebral ischaemic patients. AB - BACKGROUND AND PURPOSE: Blood oxygenation level dependent contrast functional magnetic resonance imaging (BOLD-fMRI) has been applied to functional mapping in brain disorders, based on the assumption that normal adults and patients with brain disorders exhibit similar evoked cerebral blood oxygenation (CBO) changes. This study compared evoked CBO changes measured by near infrared spectroscopy (NIRS) with the activation mapping obtained by BOLD-fMRI in patients with cerebral ischaemia. METHODS: The study involved six normal adults and six patients with cerebral ischaemia. Hand grasping was performed as a motor task. All patients could perform the task similarly to the controls at the time of examination, but single photon emission computed tomography demonstrated low baseline cerebral blood flow and a decreased haemodynamic reserve in the primary sensorimotor cortex on the lesion side. Using NIRS, concentration changes of deoxyhaemoglobin (Deoxy-Hb), oxyhaemoglobin (Oxy-Hb), and total haemoglobin (Total-Hb) were measured in the primary sensorimotor cortex contralateral to the task. BOLD-fMRI signals were measured by 1.5 T magnetic resonance imaging using an echo-planar technique. Activation maps were calculated by statistical parametric mapping. RESULTS: In the controls, Deoxy-Hb decreased in association with increases of Oxy-Hb and Total-Hb in the primary sensorimotor cortex during the task. However, in the patients, Deoxy-Hb increased significantly from baseline, while Oxy-Hb and Total-Hb also increased, indicating the presence of rCBF increases in response to neuronal activation. BOLD-fMRI demonstrated only limited activation areas in the primary sensorimotor cortex on the lesion side. CONCLUSION: The CBO changes in patients with cerebral ischaemia differed from those of normal adults; Deoxy-Hb was increased in activation areas of the patients. This implies that BOLD-fMRI may overlook activation areas in the patients unless both increases and decreases of signal are taken into consideration. PMID- 12122180 TI - Risk factors for aneurysmal subarachnoid haemorrhage: the Tromso study. AB - OBJECTIVES: To conduct a population based case-control study with premorbid registration of potential risk factors to address the difficulty in identifying risk factors for aneurysmal subarachnoid haemorrhage (SAH). SAH is rare in prospective studies, and retrospective studies may have a selection bias. METHODS: The Tromso health study is a population based survey of risk factors for cardiovascular disease in 27 161 subjects. 26 cases of aneurysmal SAH were identified in which risk factors were registered before the bleeding. Four age and sex matched controls were selected for each case. A backward logistic regression analysis was conducted and odds ratios (ORs) for significant risk factors were calculated. Systolic and diastolic blood pressure, cigarette smoking habits, serum concentrations of lipoproteins, body mass index, and coffee consumption were analysed. RESULTS: The crude annual incidence rate of aneurysmal SAH was 8.84/100 000 population. The proportion of current smokers was significantly (p = 0.003) higher in patients with SAH (73.1%) than in controls (41.3%). Drinking more than five cups of coffee per day was more common among patients (85%) than controls (59%) (p = 0.004). Mean (SD) systolic blood pressure was higher (p = 0.017) in patients (154.0 (32.5)) than in controls (136.3 (23.3)). Regression analysis showed that cigarette smoking (p = 0.04), systolic blood pressure (p < 0.0001), and coffee consumption (p = 0.004) were independent risk factors for SAH. The OR of current smokers versus never smokers was 4.55 (95% confidence interval (CI) 1.08 to 19.30) and the OR of drinking more than five cups of coffee a day was 3.86 (95% CI 1.01 to 14.73). The OR of an increase in systolic blood pressure of 20 mm Hg was 2.46 (95% CI 1.52 to 3.97). CONCLUSIONS: Cigarette smoking and hypertension are significant independent risk factors for aneurysmal SAH. A high coffee consumption may also predispose patients to aneurysmal SAH. PMID- 12122181 TI - Comparison of the psychometric characteristics of the functional independence measure, 5 item Barthel index, and 10 item Barthel index in patients with stroke. AB - OBJECTIVES: To compare the reliability, validity, and responsiveness of the motor subscale of the functional independence measure (FIM), the original 10 item Barthel index (BI), and the 5 item short form BI (BI-5) in inpatients with stroke receiving rehabilitation. METHODS: 118 inpatients with stroke at a rehabilitation unit participated in the study. The patients were tested with the FIM motor subscale and original BI at admission to the rehabilitation ward and before discharge from the hospital. The distribution, internal consistency, concurrent validity, and responsiveness of each measure were examined. RESULTS: The BI and FIM motor subscale showed acceptable distribution, high internal consistency (alpha coefficient > or = 0.84), high concurrent validity (Spearman's correlation coefficient, r(s) > or = 0.92, intraclass correlation coefficient (ICC) > or = 0.83), and high responsiveness (standardised response mean > or = 1.2, p < 0.001). The BI-5 exhibited a notable floor effect at admission but this was not found at discharge. The BI-5 showed acceptable internal consistency at admission and discharge (alpha coefficient > or = 0.71). The concurrent validity of the BI 5 was poor to fair at admission (r(s) = 0.74, ICC < or = 0.55) but was good at discharge (r(s) > or = 0.92, ICC > or = 0.74). It is noted that the responsiveness of the BI-5 was as high as that of the BI and the FIM motor subscale. CONCLUSIONS: The results showed that the BI and FIM motor subscale had very acceptable and similar psychometric characteristics. The BI-5 appeared to have limited discriminative ability at admission, particularly for patients with severe disability; otherwise the BI-5 had very adequate psychometric properties. These results may provide information useful in the selection of activities of daily living measures for both clinicians and researchers. PMID- 12122182 TI - Cognitive dysfunction after isolated brain stem insult. An underdiagnosed cause of long term morbidity. AB - Cognitive dysfunction adversely influences long term outcome after cerebral insult, but the potential for brain stem lesions to produce cognitive as well as physical impairments is not widely recognised. This report describes a series of seven consecutive patients referred to a neurological rehabilitation unit with lesions limited to brain stem structures, all of whom were shown to exhibit deficits in at least one domain of cognition. The practical importance of recognising cognitive dysfunction in this group of patients, and the theoretical significance of the disruption of specific cognitive domains by lesions to distributed neural circuits, are discussed. PMID- 12122183 TI - Viewing less to see better. AB - OBJECTIVE: To assess the efficacy, as well as the long term duration, of a new procedure for the rehabilitation of visuospatial neglect in patients with right hemisphere stroke. METHODS: Patients with right unilateral hemispheric damage identified with neglect were assigned to a treatment (T+) or a control (T-) group. The treatment consisted in abolishing all visual inputs from the right hemispace for one week by means of specially devised hemiblinding goggles. Patients' visuospatial abilities were tested and compared between groups immediately after the week of treatment. Both groups were further assessed one week after treatment suspension for evaluation of long term beneficial effects. RESULTS: Following the treatment, a substantial amelioration of visuospatial neglect symptoms was selectively observed in the T+ group. In contrast, untreated patients showed only weak signs of recovery. Most important, the amelioration obtained in the T+ group of patients was not ephemeral, being significantly maintained after a further period of one week, even after suspension of the treatment. CONCLUSION: The protracted efficacy of the proposed "hemiblinding technique" may have important implications for the recovery of visuospatial neglect and may be a very promising tool for investigating both the cognitive and the neural basis of neglect rehabilitation. PMID- 12122184 TI - Detection of preclinical motor neurone loss in SOD1 mutation carriers using motor unit number estimation. AB - OBJECTIVE: To determine the pattern of motor neurone loss in amyotrophic lateral sclerosis (ALS). In particular, to determine whether there is a gradual life long presymptomatic motor neurone loss or, alternatively, a sudden catastrophic loss just before the onset of symptoms. METHOD: The statistical motor unit number estimation (MUNE) technique was used in a longitudinal study of 19 asymptomatic carriers of the Cu, Zn superoxide dismutase 1 (SOD1) gene. MUNE results were compared with those of 34 age and sex matched SOD1 negative family controls and 23 population controls. Motor neurone loss was also estimated in 12 patients with sporadic ALS. 84 subjects (43 male and 41 female patients) with an age range from 16-73 years were followed up over three years, both clinically and by MUNE, every six months. RESULTS: In 2 of the 19 mutation carriers, there was a sudden reduction in MUNE several months before the onset of weakness. The patients with symptomatic sporadic ALS also had a reduced MUNE, but there was no detectable loss of motor neurones in the remainder of the subjects. CONCLUSION: MUNE can be used to detect preclinical loss of motor units in familial ALS. Normal numbers of motor neurones were maintained in 17 SOD1 mutation carriers over the three year period. There was an abrupt loss of motor neurones just before the onset of symptomatic weakness in two SOD1 mutation carriers. These results suggest that some form of trigger may initiate rapid cell loss and death of motor neurones just before the onset of symptoms. PMID- 12122185 TI - Prolonged hemiplegic migraine associated with unilateral hyperperfusion on perfusion weighted magnetic resonance imaging. PMID- 12122186 TI - Slowly progressive Foix-Chavany-Marie syndrome associated with chronic herpes simplex encephalitis. PMID- 12122187 TI - Lesion responsible for WEMINO syndrome confirmed by magnetic resonance imaging. PMID- 12122188 TI - A relapsing-remitting type of ocular myasthenia gravis without typical muscle fatiguability. PMID- 12122189 TI - Acute dopaminergic challenge tests to assess postural/kinetic tremor of different origin: a case report. PMID- 12122190 TI - Distal myopathy with tubular aggregates: a new phenotype associated with multiple deletions in mitochondrial DNA? PMID- 12122191 TI - Topiramate induced manic episode. PMID- 12122192 TI - Headache. PMID- 12122195 TI - Surfing for headache. PMID- 12122193 TI - Chronic daily headache. PMID- 12122194 TI - New daily persistent headache. PMID- 12122196 TI - Migraine: diagnosis and management. PMID- 12122197 TI - Colloid cyst of the IIIrd ventricle. PMID- 12122198 TI - US guidelines on neuroimaging in patients with non-acute headache: a commentary. PMID- 12122199 TI - Trigeminal autonomic cephalgias. PMID- 12122200 TI - Facial pain: neurological and non-neurological. PMID- 12122201 TI - Acute headache in the emergency department. PMID- 12122202 TI - The Chiari malformations. PMID- 12122203 TI - Multivesicular bodies and multivesicular endosomes: the "ins and outs" of endosomal traffic. AB - Multivesicular endosomes (MVEs) are complex intracellular organelles that function in endocytosis. A major function of the endocytic pathway is to sort internalized macromolecules and membrane proteins. Appropriately sorted proteins, such as epidermal growth factor (EGF) receptor (EGFR), are incorporated into MVEs before transport to the lysosomal compartment, where degradation occurs. Thus, MVEs operate in the endosome-to-lysosome portion of the pathway. In yeast cells, where MVE formation has been extensively studied, the pathway terminates in the yeast vacuole, which is equivalent to the vertebrate lysosome. MVEs arise by invagination of the limiting membrane of an endosomal vesicle such that many small internal vesicles are formed, hence the term "multivesicular endosome." In part, the internalization and targeting of membrane proteins to the MVE involves ubiquitin, a small protein associated with protein degradation. In reticulocytes and certain antigen-presenting cells, MVEs are routed to the plasma membrane rather than the lysosome, releasing small vesicles called "exosomes" back into the extracellular space. PMID- 12122204 TI - Folate deficiency disturbs hepatic methionine metabolism and promotes liver injury in the ethanol-fed micropig. AB - Alcoholic liver disease is associated with abnormal hepatic methionine metabolism and folate deficiency. Because folate is integral to the methionine cycle, its deficiency could promote alcoholic liver disease by enhancing ethanol-induced perturbations of hepatic methionine metabolism and DNA damage. We grouped 24 juvenile micropigs to receive folate-sufficient (FS) or folate-depleted (FD) diets or the same diets containing 40% of energy as ethanol (FSE and FDE) for 14 wk, and the significance of differences among the groups was determined by ANOVA. Plasma homocysteine levels were increased in all experimental groups from 6 wk onward and were greatest in FDE. Ethanol feeding reduced liver methionine synthase activity, S-adenosylmethionine (SAM), and glutathione, and elevated plasma malondialdehyde (MDA) and alanine transaminase. Folate deficiency decreased liver folate levels and increased global DNA hypomethylation. Ethanol feeding and folate deficiency acted together to decrease the liver SAM/S adenosylhomocysteine (SAH) ratio and to increase liver SAH, DNA strand breaks, urinary 8-oxo-2'-deoxyguanosine [oxo(8)dG]/mg of creatinine, plasma homocysteine, and aspartate transaminase by more than 8-fold. Liver SAM correlated positively with glutathione, which correlated negatively with plasma MDA and urinary oxo(8)dG. Liver SAM/SAH correlated negatively with DNA strand breaks, which correlated with urinary oxo(8)dG. Livers from ethanol-fed animals showed increased centrilobular CYP2E1 and protein adducts with acetaldehyde and MDA. Steatohepatitis occurred in five of six pigs in FDE but not in the other groups. In summary, folate deficiency enhances perturbations in hepatic methionine metabolism and DNA damage while promoting alcoholic liver injury. PMID- 12122205 TI - The threshold for polyglutamine-expansion protein aggregation and cellular toxicity is dynamic and influenced by aging in Caenorhabditis elegans. AB - Studies of the mutant gene in Huntington's disease, and for eight related neurodegenerative disorders, have identified polyglutamine (polyQ) expansions as a basis for cellular toxicity. This finding has led to a disease hypothesis that protein aggregation and cellular dysfunction can occur at a threshold of approximately 40 glutamine residues. Here, we test this hypothesis by expression of fluorescently tagged polyQ proteins (Q29, Q33, Q35, Q40, and Q44) in the body wall muscle cells of Caenorhabditis elegans and show that young adults exhibit a sharp boundary at 35-40 glutamines associated with the appearance of protein aggregates and loss of motility. Surprisingly, genetically identical animals expressing near-threshold polyQ repeats exhibited a high degree of variation in the appearance of protein aggregates and cellular toxicity that was dependent on repeat length and exacerbated during aging. The role of genetically determined aging pathways in the progression of age-dependent polyQ-mediated aggregation and cellular toxicity was tested by expressing Q82 in the background of age-1 mutant animals that exhibit an extended lifespan. We observed a dramatic delay of polyQ toxicity and appearance of protein aggregates. These data provide experimental support for the threshold hypothesis of polyQ-mediated toxicity in an experimental organism and emphasize the importance of the threshold as a point at which genetic modifiers and aging influence biochemical environment and protein homeostasis in the cell. PMID- 12122206 TI - Genome sequence of a serotype M3 strain of group A Streptococcus: phage-encoded toxins, the high-virulence phenotype, and clone emergence. AB - Genome sequences are available for many bacterial strains, but there has been little progress in using these data to understand the molecular basis of pathogen emergence and differences in strain virulence. Serotype M3 strains of group A Streptococcus (GAS) are a common cause of severe invasive infections with unusually high rates of morbidity and mortality. To gain insight into the molecular basis of this high-virulence phenotype, we sequenced the genome of strain MGAS315, an organism isolated from a patient with streptococcal toxic shock syndrome. The genome is composed of 1,900,521 bp, and it shares approximately 1.7 Mb of related genetic material with genomes of serotype M1 and M18 strains. Phage-like elements account for the great majority of variation in gene content relative to the sequenced M1 and M18 strains. Recombination produces chimeric phages and strains with previously uncharacterized arrays of virulence factor genes. Strain MGAS315 has phage genes that encode proteins likely to contribute to pathogenesis, such as streptococcal pyrogenic exotoxin A (SpeA) and SpeK, streptococcal superantigen (SSA), and a previously uncharacterized phospholipase A(2) (designated Sla). Infected humans had anti-SpeK, -SSA, and Sla antibodies, indicating that these GAS proteins are made in vivo. SpeK and SSA were pyrogenic and toxic for rabbits. Serotype M3 strains with the phage-encoded speK and sla genes increased dramatically in frequency late in the 20th century, commensurate with the rise in invasive disease caused by M3 organisms. Taken together, the results show that phage-mediated recombination has played a critical role in the emergence of a new, unusually virulent clone of serotype M3 GAS. PMID- 12122207 TI - Comprehensive analysis of gene expression in Nicotiana tabacum leaves acclimated to oxidative stress. AB - The molecular mechanisms by which plants acclimate to oxidative stress are poorly understood. To identify the processes involved in acclimation, we performed a comprehensive analysis of gene expression in Nicotiana tabacum leaves acclimated to oxidative stress. Combining mRNA differential display and cDNA array analysis, we estimated that at least 95 genes alter their expression in tobacco leaves acclimated to oxidative stress, of which 83% are induced and 17% repressed. Sequence analysis of 53 sequence tags revealed that, in addition to antioxidant genes, genes implicated in abiotic and biotic stress defenses, cellular protection and detoxification, energy and carbohydrate metabolism, de novo protein synthesis, and signal transduction showed altered expression. Expression of most of the genes was enhanced, except for genes associated with photosynthesis and light-regulated processes that were repressed. During acclimation, two distinct groups of coregulated genes ("early-" and "late response" gene regulons) were observed, indicating the presence of at least two different gene induction pathways. These two gene regulons also showed differential expression patterns on an oxidative stress challenge. Expression of "late-response" genes was augmented in the acclimated leaf tissues, whereas expression of "early-response" genes was not. Together, our data suggest that acclimation to oxidative stress is a highly complex process associated with broad gene expression adjustments. Moreover, our data indicate that in addition to defense gene induction, sensitization of plants for potentiated gene expression might be an important factor in oxidative stress acclimation. PMID- 12122208 TI - alpha -Synucleinopathy and selective dopaminergic neuron loss in a rat lentiviral based model of Parkinson's disease. AB - Parkinson's disease (PD) is characterized by the progressive loss of substantia nigra dopaminergic neurons and the presence of cytoplasmic inclusions named Lewy bodies. Two missense mutations of the alpha-synuclein (alpha-syn; A30P and A53T) have been described in several families with an autosomal dominant form of PD. alpha-Syn also constitutes one of the main components of Lewy bodies in sporadic cases of PD. To develop an animal model of PD, lentiviral vectors expressing different human or rat forms of alpha-syn were injected into the substantia nigra of rats. In contrast to transgenic mice models, a selective loss of nigral dopaminergic neurons associated with a dopaminergic denervation of the striatum was observed in animals expressing either wild-type or mutant forms of human alpha-syn. This neuronal degeneration correlates with the appearance of abundant alpha-syn-positive inclusions and extensive neuritic pathology detected with both alpha-syn and silver staining. Lentiviral-mediated expression of wild-type or mutated forms of human alpha-syn recapitulates the essential neuropathological features of PD. Rat alpha-syn similarly leads to protein aggregation but without cell loss, suggesting that inclusions are not the primary cause of cell degeneration in PD. Viral-mediated genetic models may contribute to elucidate the mechanism of alpha-syn-induced cell death and allow the screening of candidate therapeutic molecules. PMID- 12122209 TI - Processing of glycans on glycoprotein and glycopeptide antigens in antigen presenting cells. PMID- 12122210 TI - The emergence of humans: the coevolution of intelligence and longevity with intergenerational transfers. AB - Two striking differences between humans and our closest living relatives, chimpanzees and gorillas, are the size of our brains (larger by a factor of three or four) and our life span (longer by a factor of about two). Our thesis is that these two distinctive features of humans are products of coevolutionary selection. The large human brain is an investment with initial costs and later rewards, which coevolved with increased energy allocations to survival. Not only does this theory help explain life history variation among primates and its extreme evolution in humans; it also provides new insight into the evolution of longevity in other biological systems. We introduce and apply a general formal demographic model for constrained growth and evolutionary tradeoffs in the presence of life-cycle transfers between age groups in a population. PMID- 12122211 TI - Biomaterial adherent macrophage apoptosis is increased by hydrophilic and anionic substrates in vivo. AB - An in vivo rat cage implant system was used to identify potential surface chemistries that prevent failure of implanted biomedical devices and prostheses by limiting monocyte adhesion and macrophage fusion into foreign-body giant cells while inducing adherent-macrophage apoptosis. Hydrophobic, hydrophilic, anionic, and cationic surfaces were used for implantation. Analysis of the exudate surrounding the materials revealed no differences between surfaces in the types or levels of cells present. Conversely, the proportion of adherent cells undergoing apoptosis was increased significantly on anionic and hydrophilic surfaces (46 +/- 3.7 and 57 +/- 5.0%, respectively) when compared with the polyethylene terephthalate base surface. Additionally, hydrophilic and anionic substrates provided decreased rates of monocyte/macrophage adhesion and fusion. These studies demonstrate that biomaterial-adherent cells undergo material dependent apoptosis in vivo, rendering potentially harmful macrophages nonfunctional while the surrounding environment of the implant remains unaffected. PMID- 12122212 TI - The crystal structure of human CD21: Implications for Epstein-Barr virus and C3d binding. AB - Human complement receptor type 2 (CD21) is the cellular receptor for Epstein-Barr virus (EBV), a human tumor virus. The N-terminal two short consensus repeats (SCR1-SCR2) of the receptor interact with the EBV glycoprotein gp350/220 and also with the natural CD21 ligand C3d. Here we present the crystal structure of the CD21 SCR1-SCR2 fragment in the absence of ligand and demonstrate that it is able to bind EBV. Based on a functional analysis of wild-type and mutant CD21 and molecular modeling, we identify a likely region for EBV attachment and demonstrate that this region is not involved in the interaction with C3d. A comparison with the previously determined structure of CD21 SCR1-SCR2 in complex with C3d shows that, in both cases, CD21 assumes compact V-shaped conformations. However, our analysis reveals a surprising degree of flexibility at the SCR1-SCR2 interface, suggesting interactions between the two domains are not specific. We present evidence that the V-shaped conformation is induced by deglycosylation of the protein, and that physiologic glycosylation of CD21 would result in a more extended conformation, perhaps with additional epitopes for C3d binding. PMID- 12122213 TI - Riboflavin is a component of the Na+-pumping NADH-quinone oxidoreductase from Vibrio cholerae. AB - Flavins are cofactors in many electron-transfer enzymes. Typically, two types of flavins perform this role: 5'-phosphoriboflavin (FMN) and flavin-adenine dinucleotide (FAD). Both of these are riboflavin derivatives, but riboflavin itself has never been reported to be an enzyme-bound component. We now report that tightly bound riboflavin is a component of the NADH-driven sodium pump from Vibrio cholerae. PMID- 12122214 TI - Redox systems of the cell: possible links and implications. PMID- 12122216 TI - Another arrow in the Drosophila quiver. PMID- 12122215 TI - How are the ABC transporters energized? PMID- 12122217 TI - Initial arm muscle activation in a planar ballistic arm movement with varying external force directions: a simulation study. AB - It has been shown in previous research that the initial phase of EMG for a punching movement remained almost unchanged regardless of whether an external force was applied to the arm. The purpose of the present study was to explain this finding with the help of simulations. A two-dimensional model of the arm actuated by 6 Hill-type muscles was used to simulate a punching movement in the horizontal plane from a prescribed starting position with 90 elbow flexion. Input to the model was the stimulation of the muscles, and output were, among others, muscle forces and segmental accelerations. A genetic algorithm was used to determine the muscle onset times that minimized movement duration and targeting error. In a subsequent forward simulation, the optimized muscle onset times for an unloaded punching movement were superimposed on the isometric stimulation necessary to hold the arm in the starting position while an external force was applied to the arm. The resulting movement was only slightly different from the unloaded movement. It appeared that because of the low level of isometric muscle force prior to the movement, and the high level of stimulation during the movement, muscle force was increased at a rate that was almost independent of the prior force level. These results confirmed the suggestion that the initial phase of EMG in ballistic movements is more related to the rate of change of force than to the absolute force level. It is hypothesized that this may simplify the task of the nervous system in the choice of initial muscle activity in ballistic arm movements because no adjustments to varying external forces are required PMID- 12122218 TI - The one-target advantage: advanced preparation or online processing? AB - The one-target advantage refers to a shorter movement time for one- target aiming movements, in comparison to aiming attempts followed by a second movement. Theoretical explanations of the one-target advantage vary in the extent to which they attribute this phenomenon to prior planning or to online control mechanisms. In this research, we attempted to gain insight into the control of sequential aiming movements by manipulating the availability of online feedback during this first or second movement component. When the participants vision was occluded during the first movement (Experiment 1) or during the second movement (Experiment 2), their performance was affected, showing that vision was important for online control of the movement sequence. A one-target advantage was found when the second movement was in the same direction as the first, but not when it was reversed with respect to the home button. Both prior planning and online control processes contribute to the one-target advantage. The degree to which these processes are important for limb control depends on the specific task demands PMID- 12122219 TI - A new look at posturographic analysis in the clinical context: sway-density versus other parameterization techniques. AB - In order to identify useful guidelines for the clinical practitioner as regards the use of static posturographic analysis, we collected a set of posturograms from 3 groups of participants (normal participants, Parkinsonian patients, and osteo-porotic patients), according to the Romberg test. From each posturogram, we extracted global parameters (in the time domain and frequency domain) and structural parameters (based on diffusion plots and sway-density plots), with a total of 38 parameters. The discriminative power of each parameter was evaluated by means of statistical analysis in relation to the condition effect (open vs. closed eyes) and the pathology effect (normal participants vs. patients). The initial set of 38 parameters was reduced to 24 by identifying clear redundancies, and then to 18 by eliminating the parameters that did not pass the condition effect with normal participants. These parameters were analyzed for reliability and discriminative power in the general framework of a biomechanic model of postural stabilization. At the end of this analysis, we suggested that a set of 4 parameters is particularly valuable in the clinical practice: 2 global parameters (sway-path and frequency band of the posturogram) and 2 structural parameters (mean value of peaks and mean inter-peak distance in the sway-density plots). PMID- 12122220 TI - Symmetry of discrete and oscillatory elbow movements: does it depend on torque that the agonist and antagonist muscle can exert? AB - The hypothesis that strength of active muscles affects the symmetry of the velocity profiles of voluntary movements was tested. In particular, it was assumed that the duration of acceleration and deceleration phases reflects the ability of the antagonistic muscles to exert torque in such a way that stronger muscle requires less time for action. Twelve subjects performed consecutive 50 flexions and extensions in blocks of either discrete or oscillatory movements. They were tested under high and moderate speed conditions, as well as within different ranges of elbow joint angles. The symmetry ratio (SR; acceleration time divided by deceleration time) was calculated in order to assess movement symmetry. The results demonstrated SR > 1 under most of the discrete and, particularly, oscillatory movement conditions. A velocity-associated increase in SR was recorded, while different ranges of elbow movements, assumed to provide different torques of the agonist and antagonist muscles, also provided different SR. The findings were generally in line with the predicted effects of movement conditions on muscle strength, particularly those related to elbow angle and elbow angular velocity. Deviations from the ideal movement symmetricity have usually been interpreted as either weakness of various motor control models and hypotheses, or as a sub-optimal control of movements in certain subject populations; the present study suggests an alternative interpretation based upon the ability of active muscles to exert torque PMID- 12122221 TI - Grip and load force coordination during a manual transport movement: findings in healthy participants. AB - During transport of an object using the precision grip with thumb and index finger, a modulation of the grip force is needed in response to the forces evoked by the movement. We measured the grip force (GF) and the load force (LF) in 10 healthy participants moving a 640-g object forward and upward. The task was repeated with various speeds. There were considerable changes with speed of the LF trajectory but not of the GF trajectory. A loss of synergy between GF and LF appeared in fast lifts. This is in contrast to the close coupling between load force and grip force repeatedly demonstrated during simple lifts. We suggest that (a) speed should be considered as an input parameter for movement planning, and (b) regulation of GF and of LF are independent under certain conditions. We discuss whether the grip-load force synergy should be considered a special case rather than a more general principle PMID- 12122222 TI - Inter-digit individuation and force variability in the precision grip of young, elderly, and Parkinson's disease participants. AB - We examine the force fluctuations in the control of grip force to determine if force variability increases or decreases in relation to the degree of inter-digit individuation. This relation was examined in young (n = 7) and elderly (n = 7) participants, and in participants diagnosed with Parkinson's disease (n = 7). Force was produced under different force levels (5%, 25%, 50% MVC) with and without visual feedback. Force variability was assessed using the standard deviation and root mean square error, and inter-digit individuation was examined using cross-approximate entropy. Force variability increased with the force level, the removal of visual feedback, and also in the Parkinson's disease compared to the young and elderly matched control participants. There was a reduction in the degree of inter-digit individuation, with increases in force level, the removal of visual feedback, and in Parkinson's disease participants compared to the matched controls. Overall, there was a negative correlation between the degree of inter-digit individuation and force variability. The force fluctuations in precision grip revealed a continuum for the degree of inter-digit individuation in which task constraints, aging, and Parkinson's disease alter the coupling between the digits in controlling grip force. PMID- 12122223 TI - The relative roles of feedforward and feedback in the control of rhythmic movements. AB - A simple pendulum model is used to study how feedforward and feedback can be combined to control rhythmic limb movements. I show that a purely feedforward central pattern generator (CPG) is highly sensitive to unexpected disturbances. Pure feedback control analogous to reflex pathways can compensate for disturbances but is sensitive to imperfect sensors. I demonstrate that for systems subject to both unexpected disturbances and sensor noise, a combination of feedforward and feedback can improve performance. This combination is achieved by using a state estimation interpretation, in which a neural oscillator acts as an internal model of limb motion that predicts the state of the limb, and by using alpha-gamma coactivation or its equivalent to generate a sensory error signal that is fed back to entrain the neural oscillator. Such a hybrid feedforward/feedback system can optimally compensate for both disturbances and sensor noise, yet it can also produce fictive locomotion when sensory output is removed, as is observed biologically. CPG behavior arises due to the interaction of the internal model and a feedback control that uses the predicted state. I propose an interpretation of the neural oscillator as a filter for processing sensory information rather than as a generator of commands. PMID- 12122224 TI - Visual perception of movement kinematics and the acquisition of "action prototypes". AB - Recognizing a class of movements as belonging to a "nominal" action category, such as walking, running, or throwing, is a fundamental human ability. Three experiments were undertaken to test the hypothesis that common ("prototypical") features of moving displays could be learned by observation. Participants viewed moving stick-figure displays resembling forearm flexion movements in the sagittal plane. Four displays (presentation displays) were first presented in which one or more movement dimensions were combined with 2 respective cues: direction (up, down), speed (fast, slow), and extent (long, short). Eight test displays were then shown, and the observer indicated whether each test display was like or unlike those previously seen. The results showed that without corrective feedback, a single cue (e.g., up or down) could be correctly recognized, on average, with the proportion correct between .66 and .87. When two cues were manipulated (e.g., up and slow), recognition accuracy remained high, ranging between .72 and .89. Three-cue displays were also easily identified. These results provide the first empirical demonstration of action-prototype learning for categories of human action and show how apparently complex kinematic patterns can be categorized in terms of common features or cues. It was also shown that probability of correct recognition of kinematic properties was reduced when the set of 4 presentation displays were more variable with respect to their shared kinematic property, such as speed or amplitude. Finally, while not conclusive, the results (from 2 of the 3 experiments) did suggest that similarity (or "likeness") with respect to a common kinematic property (or properties) is more easily recognized than dissimilarity. PMID- 12122225 TI - Learning a new bimanual coordination pattern is influenced by existing attractors. AB - The present study investigates whether the acquisition of a rhythmical bimanual coordination pattern is influenced by existing intrinsic coordination tendencies. Participants were required to learn 1 of 5 new coordination patterns, whose relative phase phi was either 36, 60, or 90 degrees away from the 0 degree and 180 degree attractors, respectively. They performed 35 trials, each consisting of 2 conditions: In the augmented feedback condition, continuous visual guidance was provided, while in the normal feedback condition participants were required to rely on normal vision of their arms. We found that all to-be-learned patterns were performed with higher accuracy in the visually guided condition, whereas interference with pre-existing coordination tendencies was more pronounced in the normal vision condition. Comparing the learning progress of the 5 groups, we found for patterns close to anti-phase, a smaller improvement and significantly larger phase errors than for patterns close to in-phase. This indicates that the acquisition of a new phase relationship is influenced by existing attractors and that the 180 degree attractor interfered more strongly with the to-be-learned pattern than the 0 degree attractor. PMID- 12122226 TI - Goal-equivalent joint coordination in pointing: affect of vision and arm dominance. AB - This study used the uncontrolled manifold (UCM) approach to study joint coordination underlying the control of task-related variables important for success at reaching and pointing to targets. More combinations of joint motions are available to the control system to achieve task success than are strictly necessary. How this abundance of motor solutions is managed by the nervous system and whether and how the availability of visual information affects the solution to joint coordination was investigated in this study. The variability of joint angle combinations was partitioned into 2 components with respect to control of either the hand's path or the path of the arm's center of mass (CM). The goal equivalent variability (GEV) component represents trial-to-trial fluctuations of the joint configuration consistent with a stable value of the hand or CM path. The other component, non-goal-equivalent variability (NGEV), led to deviations away from the desired hand or CM path. We hypothesized a style of control in which the NGEV component is selectively restricted while allowing a range of goal equivalent joint combinations to be used to achieve stability of the hand and CM paths. Twelve healthy right-handed subjects reached across their body to the center of a circular target with both the right and left arms and with their eyes open or closed on different trials. When repeating the task with the same arm under identical task conditions, subjects used a range of goal-equivalent joint configurations to control the entire trajectory of both the hand's and the arm's CM motion, as well as the terminal position of the pointer-tip. Overall joint configuration variability was consistently larger in the middle of the movement, near the time of peak velocity. The style of joint coordination was qualitatively similar regardless of the arm used to point or the visual condition. Quantitative differences in the structure of joint coordination were present for the non dominant arm, however, when pointing in the absence of vision of the hand and target. The results of this study suggest that the nervous system uses a control strategy that provides for a range of goal-equivalent, rather than unique, joint combinations to stabilize the values of important task-related variables, while selectively restricting joint configurations that change these values. The possible advantage of this style of control is discussed. Absence of vision during reaching affected joint coordination only quantitatively and only for the less skilled left arm, suggesting that the role of visual information may be greater when organizing the motor components of this arm. PMID- 12122227 TI - Regular exercise (3x45 min/wk) decreases plasma viscosity in sedentary obese, insulin resistant patients parallel to an improvement in fitness and a shift in substrate oxidation balance. AB - Exercise training decreases blood viscosity in athletes parallel with metabolic improvements mostly characterized by an increase in insulin sensitivity. Patients with low insulin sensitivity exhibit a host of metabolic disorders that may also benefit from regular training. However, the hemorheologic aspects of training in such subjects are not known and we aimed at characterizing them. SUBJECTS: Thirty two obese insulin resistant subjects were tested before and after 2 months. Twenty-one of them were trained (3x45 min/wk) at a level defined by exercise calorimetry and corresponding to the power at which lipid oxidation reaches a maximum (LIPOX(max )) and eleven served as controls. The two groups were matched for age and body mass index. There was no weight change in controls while the 2 months training period decreased weight by 2.5 kg (p<0.02). This change was totally explained by a loss in fat mass (-2.7 kg, p<0.02) while fat free mass remained unchanged. Blood rheology was unchanged in the control group while training improved plasma viscosity eta(pl) (before: 1.43+/-0.03 mPa.s; after: 1.35+/-0.03 mPa.s, p<0.02). There was no change in either hematocrit, red cell rigidity or red cell aggregation. The balance of substrates oxidation shifted towards a higher use of lipids (point of crossover where subjects oxidize 70% carbohydrates 30% lipids: before 39.3+/-6.9 watts; after 70.8+/-6 watts, p<0.001; point where lipid oxidation is maximal (LIPOX(max )) before: 16.5+/-1.4 watts; after: 21.4+/-1.3 watts, p<0.001) and V(O(2max )) increased by 74% (p<0.01). Consistent with observations in athletes, the metabolic and ergometric improvements induced by training reduces eta(pl) in sedentary, insulin resistant patients, but at those low levels training does not appear to induce "autohemodilution" (as reflected by hematocrit) neither it improves red cell deformability or aggregation. The reliability of eta(pl) as simple and unexpensive marker of efficiency of training in insulin resistant patients should be further evaluated. PMID- 12122228 TI - Postexercise red cell aggregation is negatively correlated with blood lactate rate of disappearance. AB - In three separate studies, we have observed that the rise in blood lactate during exercise is correlated to blood viscosity and red cell aggregation. Whether these results were related to an effect of blood rheology on lactate production by muscles or on lactate disappearance remains unknown. The modelling of postexercise lactate kinetics allows a fair evaluation of lactate production by muscles (gamma1) and lactate disappearance (gamma2), the latter being easily measurable with simplified protocols. We thus investigated the relationships between pre- and postexercise blood rheology and gamma2. Ten subjects (2 female and 8 males; age 16-45 yr, weight 62-106.5 kg) exhibiting a wide range of gamma2 (from 2 to 7.7x10(-2) min(-1)) underwent a maximal exercise-test with postexercise calculation of gamma2 with the simplified formula gamma2=0.0724+0.755(Lac8-Lac20)/(Lac8.Deltat)-0.00684Lac20 where Lac8 and Lac20 are lactate concentrations 8 and 10 min after exercise stop at the level of VO2max, as previously reported. During exercise whole blood viscosity eta(b) increased (+15%, p<0.01) due to a rise in hematocrit (p<0.05) and plasma viscosity (+0.08+/-0.03 mPa.s, p<0.05), while red cell rigidity was unchanged. Red cell aggregation (Myrenne M1) increased by 11% (p<0.05). Postexercise M1 (measured at VO2max) was the only hemorheologic parameter correlated to gamma2 (r=-0.697, p=0.037). We find once again a statistical relationship between lactate at exercise and red cell aggregation. Microcirculatory adaptations influenced by red cell aggregation may influence lactate disposal (as reflected by gamma2), adding its effect to that of the balance between carbohydrates and fat oxidation which is the major determinant of blood lactate concentrations at exercise in physiological conditions. PMID- 12122229 TI - Haemorheological profile in metabolic Syndrome. AB - Metabolic Syndrome (MetS) has been defined as a clinical condition including impaired glucose tolerance or diabetes mellitus and/or insulin-resistance, associated with two or more of the following components: arterial hypertension, central obesity, dyslipidaemia, microalbuminuria. In a group of subjects with MetS we examined the macrohaemorheological profile, demonstrating a significant increase of blood, plasma and serum viscosity and a decrease of whole blood filterability. The results show that in these subjects a secondary hyperviscosity condition is present, but also that several significant correlations are present between the haemorheological variables and some aspects of MetS, especially those reflecting central obesity (waist to hip ratio) and insulin-resistance. The altered haemorheological profile likely contributes to explain the high cardiovascular risk present in MetS, but it may also participate, through its influence on haemodynamic pattern, in the pathogenesis of insulin-resistance. PMID- 12122230 TI - The hemorheological effects of raloxifene in postmenopausal women with osteoporosis. Results of a 3-year placebo-controlled clinical trial. AB - Raloxifene, the prototype of the selective estrogen receptor modulators, has been associated with an increased risk of venous thromboembolism. As hemorheological factors may be involved in thrombus formation this placebo-controlled study investigated whether raloxifene was associated with changes in determinants of blood viscosity. Fifty-seven post-menopausal women were randomly assigned to receive placebo, raloxifene 60 mg/day, or raloxifene 120 mg/day for 36 months. Venous blood samples were collected at baseline and at 12-monthly intervals and used to measure hematocrit, whole blood and plasma viscosity and plasma fibrinogen concentration. Time- and treatment-related changes in the grouped and pooled data was analysed using ANOVA with repeated measures and correlation matrices. The mean values of all the hemorheological indices showed small inconsistent changes within the normal reference range over the 36-month period of the study. There was a small but significant decrease over time in high shear rate blood viscosity and plasma viscosity in raloxifene-treated subjects compared to those receiving placebo (p<0.05). Correlation analyses showed the anticipated relationships between blood viscosity and hematocrit and plasma viscosity levels and also between plasma viscosity and plasma fibrinogen concentration. No subject developed a thromboembolic vascular event during the study. These results show that compared with placebo treated-subjects, long-term raloxifene treatment in post-menopausal women, at a dose of either 60 or 120 mg daily, was not associated with adverse changes in hemorheological factors that may contribute to venous thromboembolism. PMID- 12122231 TI - The effects of cholesterol levels on hemorheological parameters in diabetic patients. AB - Red blood cell (RBC) deformability is an important hemorheological parameter to determine the passage of RBC through narrow capillaries and the reduction of blood viscosity under high shear rates. Although it has been substantial evidence that diabetes mellitus (DM) and hypercholesterolemia increase the risk of coronary heart disease, the mechanism is unclear. In this study the relationship between hemorheological parameters and plasma cholesterol in type 2 diabetic patients (n=55, mean age 43.4+/-9.2 years) was examined. Type 2 diabetic patients were classified as normocholesterolemic (n=25; cholesterol < or = 200 mg/dl) and hypercholesteroloemic (n=30; cholesterol > 200 mg/dl) subgroups. Hypercholesterolemic type 2 diabetic patients had the highest blood and plasma viscosity and the lowest RBC deformability. The results were significantly different from normocholesterolemic type 2 diabetic patients (p<0.001). Our data suggest that elevated plasma cholesterol may impair RBC deformability and increase in blood and plasma viscosity by an additional effect to hyperglycemia in type 2 diabetic patients. PMID- 12122232 TI - The effect of atorvastatin on hemorheological parameters in rabbits fed on a normal diet. AB - The effects of statins have been investigated mostly in hyperlipidemic states so far. We analysed blood cholesterol, triglyceride, albumin, fibrinogen and gammaglobulin levels, haematocrite, hemoglobin, erythrocyte, leukocyte and platelet counts, blood and plasma viscosity and erythrocyte rigidity in 12 rabbits fed on a normal diet (chow) which were given 1 mg/kg/day atorvastatin for 4 weeks. Compared to the baseline levels, erythrocyte rigidity (k=0.12+/-0.05 vs. k=0.7+/-0.02) and gammaglobulin levels (1.03+/-0.23 g/dl vs. 0.78+/-0.27 g/dl) decreased significantly (p=0.008 and p=0.025, respectively). Blood lipids, hematological variables, blood and plasma viscosity did not change statistically. Our findings imply that in a normolipemic state, statins given in low doses may improve erythrocyte rigidity without altering blood lipids in short term. Decreased plasma gammaglobulin levels may be reflecting their immunomodulatory effects. PMID- 12122233 TI - The role of erythrocytapheresis in secondary erythrocytosis therapy. PMID- 12122234 TI - Complexity analysis of placental blood flow in normal and high-risk pregnancies. AB - Various strategems of complexity analysis of microvascular blood flow were carried out in several fields of medicine in the past, as such as angiology, ophthalmology and neurology. The introduction of colour-angio-mode, a special form of colour coded Doppler sonography, now makes possible to perform complexity analysis of the placental blood displacement even in the absence of information about hydrodynamic details such as directionality, velocity profile and number of displaced blood cells. Algorithms were developed which allows to extract information concerning the time averaged power of phonon-erythrocytes collision events (from the square of the frequencies of back scattered ultrasound recorded during 166 ms) in 20,000 to 40,000 regions of interest. The obtained values are being displayed as false coloured pixels on a video-screen, we succeeded to obtain quantitative data about displacement rates.In cross-sectional and longitudinal studies we generated typical diagrams displaying the "occurrence rate" of various powers of displacement over time. By this mode of display contour plots can be generated, showing a large amount of low intensity pixels and a small amount of high intensity pixels representing the parenchymatous blood flow inside the placenta. As was to be expected, interdependencies between the placental blood flow and the maternal and fetal heart rates as well as the maternal breathing can be found. While there was only limited influence of maternal and fetal heart rate on the placental blood flow, maternal breathing showed striking influence. Surprisingly, during expiration the power of placental blood movement was decreased, and there was a marked increase during inspiration. In cases of severe intrauterine growth retardation, colour pixel intensities were seen to transiently vanish during end-expiration. The power of placental blood displacement was marked increased subsequent to reducing maternal hematocrit during hemodilution therapy by infusion of artificial colloids. These interdependencies could be confirmed by ex vivo examinations perfusing and percolating the placenta after birth in a hemodynamic model. Additionally, we found interdependencies between fetal and maternal blood displacement inside the placenta. By modelling the decrease of fetal inotropic power in the ex vivo examinations, increase in the power of maternal blood displacement in the intervillous space. The two types of placental blood flow are known to be determined by many factors. While it is currently impossible to measure all these parameters determining an parenchymatous blood flow, it is possible to obtain useful informations about the physiologic and pathophysiologic changes of placental blood flow using colour-angio-mode as a tool of complexity analysis based on the distribution of local blood displacement. This new knowledge can help to understand clinically relevant changes in the individual patient as well their underlying causes. PMID- 12122235 TI - Stress and strain distribution behavior in the bone due to the effect of cancellous bone, dental implant material and the bone height. AB - The stress and strain distribution in the bone surrounding a dental implant have been analyzed using the finite element and optimization techniques. The effect of removing cancellous bone completely or not was investigated. Two models were used, the first model without cancellous bone and the second with it. The elastic modulus of the implant material and the length of the implant neck or the height of bone surrounding the implant were used as design variables in the two models. In the first model a higher level of stress in the cortical bone surrounding the neck of the implant was found. While in the second model, it was found surrounding the tip of the implant. The result indicates that the stress concentration factor in the bone of the first model is reduced by 30% compared to the initial design. However, when the implant was surrounded by sleeve of cancellous bone (second model) the stress concentration is reduced by 16% for cortical bone and 15% for cancellous bone. This reduction help to reduce fatigue failure and bone resorption. PMID- 12122236 TI - Age related constitutive laws and stress distribution in human main coronary arteries with reference to residual strain. AB - An analytical model has been used to simulate the effects of tissue aging on residual strain, constitutive relations and stiffness parameter in the main right and left (ramus circumflexus) human coronary arteries, based on experimental data. The experimental opening angle theta scatters considerably with age. The optimum angle theta(op) approximately equals 70 degrees, which makes the circumferential stress uniform in the arterial wall at a normal blood pressure, is approximately constant throughout aging. Above age of the 15 years the estimated and experimental values of theta are greater than theta(op) and therefore the mechanical load of the inner layers of the media and the intima decreases and the adventitia is overloaded. On the basis of nonlinear regression analysis, age-related constitutive laws of arterial wall circumferential stiffness have been determined. Above the age of 30, arterial wall hardening increases rapidly. The left coronary artery is stiffer than the right artery for groups from 35 to 45 years of age. Hyperelasticity theory has been used to identify age-related multiaxial stress through wall thickness. A theoretical model based on the reduced Green strain provides a very good representation of the coronary artery circumferential mechanical response and predicts its nearly isotropic behavior. Bio-composite material forms non-homogeneous stresses and, in the course of aging, it increases the adventitia loading. In groups aged from 10 to 15 years, whose coronary artery residual strains are low, the circumferential stress distribution has a classic form. Stiffness parameter beta gradually increases with age and this increase is significant above the age of 60. Parameter beta tends to decrease when the opening angle theta increases. PMID- 12122237 TI - Stabilization of nontoxic PVC formulation for gamma irradiation sterilization, I. Effect of additives. AB - In order to investigate the fundamental data for the resistance of gamma radiation sterilization of polyvinylchloride (PVC), the formulations of the additives such as plasticizers of dioctylphthalate, trioctyltrimellitate and polyester, second plasticizers, Ca/Zn nontoxic metallic stabilizers with powder, paste and liquid state, and phosphite stabilizers have been carried out. The control and irradiated PVC samples with 1.5, 2.5 and 4.0 Mrads were characterized by mechanical tester, colorimetry, and extractant in water. The effect of plasticizers observed in the order of dioctylphthalate approximately equal trioctyltrimellitate > polymeric plasticizer. It was observed in the order of Ca/Zn metallic stabilizers of paste > liquid approximately equal powder state for the color change and liquid > paste > powder for the extractant. The mechanism of the discoloration of PVC in our experiment was predominant the formation of polyene by the dehydrochlorination rather than the formation of keton and aldehyde by the oxidation and chain dissociation by the measurement of gel permeation chromatography and mechanical property. The proposed mechanisms of stabilization and discolorization with various additives are also discussed. PMID- 12122238 TI - Effect of loading rate on the apparent fracture toughness of acrylic bone cement. AB - In this study, a determination is made of the effect of loading rate, v (0.1 mm min(-1) versus 1.0 mm min(-1) versus 10 mm min(-1)) on the value of the plane strain fracture toughness, K(Ic), of three commercial formulations of acrylic bone cement (Osteopal), CMW3, and Copal), that are characterized as "low-", "medium-", and "high-" viscosity brands, respectively). For all formulations, K(Ic) increases with increase in v. However, while this trend is statistically significant for CMW3 and Copal, this is not so for Osteopal. The CMW3 and Copal results are explained in terms of changes of the molecular relaxation transitions in the cement and the thermal state at the crack tip of the test specimen. Two implications of the findings are discussed. In the case of Osteopal, a recommendation for further study is made. PMID- 12122240 TI - Automatic quantitative analysis of bipolysaccharide liposome by image processing method. AB - This paper established the extracting method and algorithm for liposome images on the basis of analyzing the form feature of liposome, and its corresponding automatic analysis software. Some algorithms, such as the logical algorithms of mathematical morphology, the grayscale-gradient co-occurrence matrix-based method and the image enhancement method of grayscale image, were used to restrain the noise effectively and increase the definition of image. The result on bipolysaccharide liposome indicates that the method increases the measuring speed and precision. PMID- 12122239 TI - Therapeutic effect of in vivo sustained estradiol release from poly (lactide-co glycolide) microspheres on bone mineral density of osteoporosis rats. AB - Poly (lactide-co-glycolide or PLGA) microspheres containing 0.3% (w/w) of estradiol were prepared by a solvent evaporation method. These PLGA microspheres had a wide particle distribution between 0.5 and more than 100 microm. The average size was 76 microm. Physicochemical properties of the microspheres were characterized by X-ray diffraction patterns, FT-IR spectra and DSC. In vitro estradiol release was maintained at a constant rate from these PLGA microspheres for 1 month. The loaded drug was totally recovered in the collection buffer within this time period. In vivo experiments were performed on Wistar rats that had received ovariectomy. These rats were fed with a vitamin D-deficient and Ca deficient diet. The combination of ovariectomy and diet induced osteoporosis. PLGA microspheres containing either 50, 100, or 200 microg estradiol were injected into these rats. The plasma estradiol in each rat was monitored for 50 days. These in vivo drug release patterns were found to be different from the one obtained from in vitro release. The Ca-AUC was not significant different among various dosages administered. However, bone mineral density for rats after the injection of estradiol loaded microspheres was higher than that obtained for the control. This suggested that all estradiol microspheres administration induced bone generation in osteoporosis rats. PMID- 12122241 TI - Comparison of the biological activity of grade GUR 1120 and GUR 415HP UHMWPE wear debris. AB - This study evaluated the in vitro response of murine macrophages to clinically relevant polyethylene particles from two grades of UHMWPE at varying volume doses. Clinically relevant UHMWPE wear debris was generated in vitro using a tri pin-on-disc tribometer. The debris was observed using a scanning electron microscope and analysed by image analysis. There was no significant difference in the wear rates of the two grades of UHMWPE. Analysis of the wear debris showed that GUR 415HP produced a higher percentage of mass of debris in the submeter size range compared to GUR 1120. The wear debris was co-cultured with C3H murine peritoneal macrophages at particle volume (microm(3)): cell number ratios of 100 :1, 50 :1, 10 :1 and 1 :1 for both grades of UHMWPE and additionally at 0.5 :1 and 0.1 :1 for grade GUR 415HP. The secretion of TNF-alpha was determined by ELISA. Significantly elevated levels of TNF-alpha were secreted at 100 :1 ratio when macrophages were challenged with wear debris from GUR 1120 and at 10 :1 and 1 :1 for debris from GUR 415HP. The results suggested that the greater percentage mass of debris in the submicrometer size range from GUR 415HP lead to a substantial increase in biological activity for this grade of UHMWPE on a volume for volume basis when compared with GUR 1120. PMID- 12122242 TI - Quantitative analysis of UHMWPE wear debris isolated from the periprosthetic femoral tissues from a series of Charnley total hip arthroplasties. AB - Submicrometer and micrometer-sized UHMWPE wear particles have been associated with osteolysis and failure of total hip replacements. A previous study by Tipper et al. examined the wear debris isolated from the acetabular periprosthetic tissues from 18 Charnley total hip replacements, and compared this data to the tribological variables of the prostheses. The present study aimed to isolate the UHMWPE wear debris from the femoral periprosthetic tissues from 10 of the same cohort of patients, and compare it with the debris isolated from the corresponding acetabular tissues. A variety of particle morphologies were observed, discrete submicrometer particles, along with flakes and fibrils. The particle size distributions ranged from 0.1 to >250 microm, however, the largest particles were only found in samples when the femoral head damage was characterised as low (R(pm) < 0.2 microm). The mode of the frequency distribution of particles was in the range of 0.1-0.5 microm for all the femoral tissues. Considerable variations were found in the mass distributions of the wear particles as a function of size for different patients. The net mass of debris isolated from the femoral tissues was significantly lower (p < 0.05, Student's t test) than from the corresponding acetabular tissues. This along with considerable spatial variation in the net mass of debris isolated from the different regions of the same sample of acetabular tissue, indicates that the transportation of the debris has a marked effect on the net mass of debris accumulated in different tissues. PMID- 12122243 TI - Long-term durability of porous hydroxyapatite with low-pressure system to support osteogenesis of mesenchymal stem cells. AB - In this study, we hypothesize that loading more marrow-derived mesenchymal stem cells (MSCs) into porous material by using a low-pressure system during subculture, creating a composite which combines MSCs and a novel mechanical reinforced porous hydroxyapatite, can result in more bone tissue formation in vivo. Within 26 weeks postimplantation, we examined in vivo bone formation of the experimental group with 100 mmHg pressure applied to porous HA blocks loaded with MSCs. For in vivo testing, the 2-week subcultured HA/MSC composites were implanted into subcutaneous sites of syngeneic rats. These implants were harvested at 13 and 26 weeks after implantation. SEM showed that the pore surface is covered by osteoblasts as well as collagenous extracellular matrix at 13 weeks. Light microscopy revealed the quantity of bone at 26 weeks was greater than at 13 weeks. These results showed that the novel mechanical reinforced porous HA combined with MSC has more potential for bone formation at 100 mmHg, making this method very efficient for bone reconstruction. PMID- 12122244 TI - Activation and extinction models for platelet adhesion. AB - Adherent platelets are an important part of both thrombus formation and in certain stages of atherogenesis. Platelets can be activated by potent chemicals released from adherent platelets and adhere far more readily than unactivated ones. An analytical and numerical model is presented utilising high Peclet number for the activation and adhesion of platelets in shear flows. The model uses a similarity transformation, which characterises the relationship between convective, diffusive transport and the bulk platelet activating reaction mechanism. A first order surface reaction mechanism is used to model platelet adhesion at the wall (cell) surface. The reduced Damkohler number, M, characterises the importance of the bulk reaction and includes both convective and diffusive terms. For a high rate of blood flow (M-->0) the activation of platelets can effectively be terminated. In contrast, for (M-->infinity) an inner layer of activated platelets exists with an infinitesimally thin reaction sheet separating activated and non-activated platelets. This characterisation by the Damkohler number highlights results found clinically, in that thrombus forms in areas of low shear (high M) and in some cases an increased blood flow (low M) can inhibit the activation of platelets completely. The model shows the critical balance that exists between convection, diffusion and reaction. PMID- 12122245 TI - The convergence of haemodynamics, genomics, and endothelial structure in studies of the focal origin of atherosclerosis. AB - The completion of the Human Genome Project and ongoing sequencing of mouse, rat and other genomes has led to an explosion of genetics-related technologies that are finding their way into all areas of biological research; the field of biorheology is no exception. Here we outline how two disparate modern molecular techniques, microarray analyses of gene expression and real-time spatial imaging of living cell structures, are being utilized in studies of endothelial mechanotransduction associated with controlled shear stress in vitro and haemodynamics in vivo. We emphasize the value of such techniques as components of an integrated understanding of vascular rheology. In mechanotransduction, a systems approach is recommended that encompasses fluid dynamics, cell biomechanics, live cell imaging, and the biochemical, cell biology and molecular biology methods that now encompass genomics. Microarrays are a useful and powerful tool for such integration by identifying simultaneous changes in the expression of many genes associated with interconnecting mechanoresponsive cellular pathways. PMID- 12122246 TI - Endothelial control of vascular tone by nitric oxide and gap junctions: a haemodynamic perspective. AB - Local haemodynamic forces acting on the endothelium modulate vascular tone through mechanisms that normalize intimal shear stress. This flow-dependent diameter response contributes to the optimization of circulatory function and is mediated via shear stress-induced release of NO, vasodilator prostanoids and a putative endothelium-derived hyperpolarizing factor or EDHF. There is growing evidence that NO/prostanoid independent relaxations involve direct heterocellular signalling between endothelial and smooth muscle cells via gap junctions. PMID- 12122247 TI - Mechanisms of blood flow-induced vascular enlargement. AB - Chronic changes in wall shear stress lead to vascular remodeling, characterized by increased vascular wall diameter and thickness, to restore wall shear stress values to baseline. Release of nitric oxide from endothelial cells exposed to excessive shear is a fundamental step in the remodeling process, and potentially triggers a cascade of events, including growth factor induction and matrix metalloproteinase activation, that together contribute to restructuralization of the vessel wall. Understanding these processes could help explain how changes in blood vessel wall structure occur in the context of atherosclerosis or aortic aneurisms. PMID- 12122248 TI - Factors influencing the nonuniform localization of monocytes in the arterial wall. AB - Adhesion of monocytes to arterial endothelium may contribute to the asymmetric distribution of atherosclerotic lesions. Possible mechanisms for adhesion in the relatively high shear stress environment found in arteries include greater monocyte deformation and/or more frequent penetration of microvilli through steric and charge barriers. In vivo, secondary flows generate forces acting normal to the endothelial cell surface. These forces may cause compression of the microvilli or enable cells to overcome steric or electrostatic barriers, increasing adhesion. To investigate this, we examined monocyte adhesion to activated endothelium in recirculating flow. Adhesion was characterized by short arrests in a narrow region on either side of the reattachment line. The median arrest time was longer than that observed at comparable shear stresses in a linear shear flow. The lifetimes of adhesion were analyzed using a model for multiple bond formation. For cells adhering near the reattachment line, the bond number per cell was greater than the value found for similar shear stresses under shear flow. Thus, multiple bond formation arising from greater normal forces in recirculating flow permits monocytes to adhere at higher shear stresses. PMID- 12122249 TI - Prediction of LDL concentration at the luminal surface of a vascular endothelium. AB - To find out whether concentration polarization of low-density lipoprotein (LDL) occurs at the surface of a vascular endothelium or not, transport of LDL in flowing blood to an water-permeable endothelium was studied theoretically by means of CFD. Calculations were carried out for an endothelium exposed to a Couette flow by assuming that the surface geometry of the endothelium could be expressed by a cosine function. Two typical cases were considered for the permeability of endothelium to water; one was uniform permeability everywhere in the endothelium, and the other was uneven permeability which was augmented at the intercellular junction. It was found that, in both cases, the surface concentration of LDL increased in going distally from the entrance, taking locally high and low values at the valleys and hills of the endothelium, respectively, and the variation was larger in the case of endothelium with uneven permeability. These results clearly showed that concentration polarization of LDL which might affect the uptake of LDL by the arterial wall certainly occurs at the surface of the endothelium even if the flow is disturbed microscopically by the uneven surface of the endothelium. PMID- 12122250 TI - Parallel-plate flow chamber for studies of 3D flow-endothelium interaction. AB - Flow induced shear stress influences vascular cellular biology and pathophysiology in numerous ways. Previous in vitro studies on interactions between flow and endothelial cells using parallel-plate flow chambers involve two dimensional flows, whereas flows in larger vessels are commonly three dimensional. We have constructed a parallel plate flow chamber with a backward facing step aligned oblique to the axis of the chamber. Flow visualisation by steady injection of ink through a hypodermic tube reveals swirling flow in the recirculation region downstream of the step. At given angles of the step, theta; (to the axis of the chamber), the pitch of the swirl and the width of the separation region, as measured in the direction perpendicular to the step, increase with the Reynolds number (Re). On the other hand, at given values of Re, reduction of theta; results in increases in the swirl pitch but decreases in the width of the separation zone. Furthermore, clearance time of ink from the separation region is shorter with an oblique step than a perpendicular one at given Re. Computer simulation confirms the 3D swirling flow created by the oblique step and provides detailed distribution of wall shear stresses in the flow chamber. PMID- 12122251 TI - Arterial bends: the development and decay of helical flows. AB - The macrocirculation is modelled by incompressible Newtonian flow through a rigid network of pipes for which possible simplifications are discussed. The common assumptions of two-dimensionality or axisymmetry can be generalised to helical symmetry, and in the first part of the paper, the three-dimensionality of arterial bends is considered by varying the curvature and torsion of a section of a helical pipe. The torsion is found to impart a preferential twist to the cross sectional flow. This loss of symmetry ensures that flow separation is less severe for a helical bend than for a toroidal bend. The effects of variations in body size are examined using allometric scaling laws. In the second part of the paper, the approach to "fully developed" Dean or Womersley flow is considered in an attempt to quantify the regions of validity of idealised models. A perturbation approach, akin to hydrodynamic stability theory, is used. It is argued that often potential flows are more suitable for describing the rapid interactions between geometry and pulsatility rather than the eventual fully developed state so that, for example, the first 100 degrees of the aortic arch may be considered irrotational. Helical potential flows are found to develop faster than the corresponding toroidal flows, but slower than those in a straight pipe. The presence of vorticity in the core also retards the development of symmetric flows. It is concluded that while idealised flows can occur at some points in the body, in general experimental observation is needed to justify their use. Particular caution is recommended when interpreting calculations with Poiseuille input. PMID- 12122252 TI - A 3D unsteady flow analysis in a doubly constricted arterial vessel. AB - The aim of this study is to examine the interaction between two mild atherosclerotic proliferations spaced apart by a distance S by analyzing their influence on flow structure, pressure drop and stress field in an arterial vessel under pulsatile flow conditions. This has been achieved numerically by employing a time accurate, cell centered finite volume method in solving the Navier-Stokes equations governing the 3D unsteady flow dynamics in a conceptual model of an multiply constricted arterial vessel. In comparison to the pressure drop across a single stenosis, nearly a 50% increase in the late systolic and early diastolic pressure drops has been observed across the two mild constrictions when they are spaced within a distance of S0.897 (average 0.939) where the velocity profiles were skewed, than along the anterior-posterior axis (average correlation factor 0.876) where the velocity profiles were in M-shape. PMID- 12122269 TI - Adhesion of leukocytes under oscillating stagnation point conditions: a numerical study. AB - Leukocyte recruitment from blood to the endothelium plays an important role in atherosclerotic plaque formation. Cells show a primary and secondary adhesive process with primary bonds responsible for capture and rolling and secondary bonds for arrest. Our objective was to investigate the role played by this process on the adhesion of leukocytes in complex flow. Cells were modelled as rigid spheres with spring like adhesion molecules which formed bonds with endothelial receptors. Models of bond kinetics and Newton's laws of motion were solved numerically to determine cell motion. Fluid force was obtained from the local shear rate obtained from a CFD simulation of the flow over a backward facing step.In stagnation point flow the shear rate near the stagnation point has a large gradient such that adherent cells in this region roll to a high shear region preventing permanent adhesion. This is enhanced if a small time dependent perturbation is imposed upon the stagnation point. For lower shear rates the cell rolling velocity may be such that secondary bonds have time to form. These bonds resist the lower fluid forces and consequently there is a relatively large permanent adhesion region. PMID- 12122270 TI - Experimental assessment of wall shear flow in models. AB - The blood flow immediately adjacent to the wall of a blood vessel or an artificial surface is of great interest. This flow defines the shear stress at the wall and is known to have a great physiological importance. The use of models is a viable method to investigate this flow. However, even in models the shear stress at the wall is difficult to assess. A new optical method is based on transparent models and uses particles in the model fluid, which are only visible near the wall. This is achieved with a model fluid having a defined opacity. This fluid obscures particles in the center of the models, but permits the observation and recording of particles close to the wall. The method has been applied for Hagen-Poiseuille flow and for the likewise well researched flow in a tube with a sudden expansion. PMID- 12122271 TI - Towards the treatment of saphenous vein bypass graft failure--a perspective of the Bristol Heart Institute. AB - Coronary artery bypass graft surgery (CABG) is widely used for the treatment of atheromatous stenosis of coronary arteries. However, as many as 50% of grafts fail within 10 years after CABG due to neointima (NI) formation, a process involving the proliferation and migration of vascular smooth muscle cells (VSMCs). Superimposed on neointima formation is accelerated atherogenesis which ultimately results in late vein graft failure. To date no therapeutic intervention has proved successful in treating late vein graft failure and as such is a matter of some urgency. However, in recent years, several diverse approaches aimed at preventing neointimal formation have been devised which have yielded promising results. These include the use of external stents, gene therapy as well as conventional pharmacological interventions. The objective of this article, therefore, is to review these recent approaches and their potential clinical applications in the treatment of vein graft disease. PMID- 12122272 TI - Effects of stents under asymmetric inflow conditions. AB - Patient-to-patient variations in artery geometry may determine their susceptibility to stenosis formation. These geometrical variations can be linked to variations in flow characteristics such as wall shear stress through stents, which increases the risk of restenosis. This paper considers computer models of stents in non-symmetric flows and their effects on flow characteristics at the wall. This is a fresh approach from the point of view of identifying a stent design whose performance is insensitive to asymmetric flow. Measures of dissipated energy and power are introduced in order to discriminate between competing designs of stents. PMID- 12122273 TI - The geometry of unstented and stented pig common carotid artery bypass grafts. AB - The long-term success of arterial bypass grafting with autologous saphenous veins is limited by neointimal hyperplasia (NIH), which seemingly develops preferentially at sites where hydrodynamic wall shear is low. Placement of a loose-fitting, porous stent around end-to-end, or end-to-side, autologous saphenous vein grafts on the porcine common carotid artery has been found significantly to reduce NIH, but the mechanism is unclear. In a preliminary study, we implanted autologous saphenous vein grafts bilaterally on the common carotid arteries of pigs, placing a stent around one graft and leaving the contralateral graft unstented. At sacrifice 1 month post implantation, the grafts were pressure fixed in situ and resin casts were made. Unstented graft geometry was highly irregular, with non-uniform dilatation, substantial axial lengthening, curvature, kinking, and possible long-pitch helical distortion. In contrast, stented grafts showed no major dilatation, lengthening or curvature, but there was commonly fine corrugation, occasional slight kinking or narrowing of segments, and possible long-pitch helical distortion. Axial growth of grafts against effectively tethered anastomoses could account for these changes. CFD studies are planned, using 3D MR reconstructions, on the effects of graft geometry on the flow. Abnormality of the flow could favour the development of vascular pathology, including NIH. PMID- 12122274 TI - Variability of 3D arterial geometry and dynamics, and its pathologic implications. AB - Geometric parameters and features vary within the vasculature. Furthermore, at any given anatomic site, there are substantial variations in geometry among individuals. These variations can contribute to a corresponding variability in the hemodynamic environment and, to the extent that hemodynamics affects the atherosclerotic process, the progress of vascular disease. Measurements of the geometry and wall morphometry of post-mortem human coronary arteries demonstrate a relationship between these variables that supports the notion that geometric variations can contribute to a corresponding variability in the local rate of progression of arterial disease. The dynamic geometry of the coronary arteries also varies from site to site and among individuals, and this variability too may play a role in the epidemiology of coronary artery disease. PMID- 12122275 TI - Atherosclerosis and flow in carotid arteries with authentic geometries. AB - The influence of blood flow on the depositions and development of atherosclerotic lesions have been observed and described since the 19th century. Observations have shown that depositions correlate with regions of low wall shear stress. However, the exact correlations between depositions, vessel geometry and flow parameters are not yet known. The purpose of this study was the quantification of atherosclerosis risk factors in carotid bifurcation. This artery has attracted particular interest because lesions are often found in this bifurcation. Post mortem, the arteries are excised and vessel casts are produced. Afterwards, the arteries are analyzed morphometrically. The vessel casts are used for the assessment of some geometrical parameters. 31 carotid bifurcations were analyzed in this study. Eight vessel casts were digitized and rendered three-dimensional mathematical models of the arteries. These data were imported by the computational fluid dynamics program FLUENT. Further, the blood flow was reconstructed in a computer model based on the individual vessel geometry. The flow parameters, such as velocity, pressure and wall shear stress were computed. At the same time the geometrical parameters and wall alterations are known. This permits the comparison of the anatomical shape and its flow with the distribution and level of the wall alterations. PMID- 12122276 TI - Combined MR imaging and numerical simulation of flow in realistic arterial bypass graft models. AB - We report methods for (a) transforming a three-dimensional geometry acquired by magnetic resonance angiography (MRA) in vivo, or by imaging a model cast, into a computational surface representation, (b) use of this to construct a three dimensional numerical grid for computational fluid dynamic (CFD) studies, and (c) use of the surface representation to produce a stereo-lithographic replica of the real detailed geometry, at a scale convenient for detailed magnetic resonance imaging (MRI) flow studies. This is applied to assess the local flow field in realistic geometry arterial bypass grafts. Results from a parallel numerical simulation and MRI measurement of flow in an aorto-coronary bypass graft with various inlet flow conditions demonstrate the strong influence of the graft inlet waveform on the perianastomotic flow field. A sinusoidal and a multi harmonic coronary flow waveform both with a mean Reynolds number (Re) of 100 and a Womersley parameter of 2.7 were applied at the graft inlet. A weak axial flow separation region just distal to the toe was found in sinusoidal flow near end deceleration (Re = 25). At the same location and approximately the same point in the cycle (Re = 30) but in coronary flow, the axial flow separation was stronger and more spatially pronounced. No axial flow separation occurred in steady flow for Re = 100. Numerical predictions indicate a region in the vicinity of the suture line (where there is a local narrowing of the graft) with a wall shear magnitude in excess of five times that associated with fully developed flow at the graft inlet. PMID- 12122277 TI - Disease patterns at arterial branches and their relation to flow. AB - The distribution of lesions around arterial branch points is complex and changes with age. Four distinct patterns - here termed the arrowhead pattern, the lateral pattern, the upstream streak and the volcano - have been reported around the origins of intercostal arteries in the human aorta at different ages. The first two patterns also occur in young and old rabbits, the third in minipigs, and the fourth in apolipoprotein E/LDL receptor knockout mice. It is unclear how all four patterns can depend solely on flow; a particular problem is that the prevalence of lipid deposition remains highly nonuniform for several branch diameters upstream of the ostium. Variations in the prevalence of fatty streaks may originate near the branch and then spread by the migration of activated endothelial cells towards the heart. The pattern of raised lesions may reflect a different aetiology. PMID- 12122278 TI - Haemodynamic flow: symmetry and synthesis. AB - This paper is presented as a summary and synthesis of the presentations at the conference entitled "Breaking Symmetry in Haemodynamics". As the accompanying papers will attest, there has been enormous progress in understanding the effects of fluid flow on the arterial endothelium and the consequential effects on the vessel wall. It is now clearly understood that the focal lesions found in atherosclerotic arteries are the product of asymmetrical flow and the resulting disturbed flow that occurs near arterial bifurcations and other selected points around the human vasculature. The flow in large vessels can now be determined accurately with MR and in vitro cast models. Although theory allows arterial flow to be characterized by asymmetry in time and space, our understanding of the processes that act to translate this asymmetry into pathology is becoming much more symmetric, or complete. The new frontiers of research in arterial flow are now translating to smaller scales, at the cellular level and below. PMID- 12122279 TI - Diagnostic DNA-flow- vs. -image-cytometry in effusion cytology. AB - AIMS: To determine the sensitivity and specificity of flow- and image-cytometry for the detection of DNA-aneuploidy as a marker for malignant cells in effusions. METHODS: 200 effusions (80 tumor cell-positive, 74 negative and 46 cytologically equivocal) were stained with DAPI-SR for DNA-flow- and with Feulgen Pararosaniline for -image-cytometry. They were measured using a PAS-flow cytometer and an AutoCyte-QUIC-DNA-workstation according to the ESACP consensus reports for DNA-flow- and -image-cytometry, respectively [7,23,29,49]. RESULTS: Sensitivity of DNA-aneuploidy for the identification of malignant cells was 32.1% for DNA-flow- and 75.0% for -image-cytometry, specificity of -euploidy in benign cells was 100.0% for both methods. Positive predictive value of DNA-aneuploidy for the identification of malignant cells was 100.0% for both techniques, negative predictive value of DNA-euploidy was 48.6% for DNA-flow- and 72.0% for image-cytometry. CONCLUSIONS: Searching for DNA-aneuploidy as a diagnostic marker for neoplastic cells in serous effusions image-cytometry revealed superior sensitivity as compared with monoparametric flow cytometry. PMID- 12122280 TI - Expression of multidrug resistance-associated markers, their relation to quantitative pathologic tumour characteristics and prognosis in advanced ovarian cancer. AB - Mean nuclear area has been consistently shown by different researchers to be a strong and independent prognostic factor in advanced ovarian carcinoma. However, the biological background of the prognostic value of nuclear area remains unclear. Others have found that the multidrug-resistance (MDR) related protein LRP has strong prognostic value. In the present study we have analysed whether the mean nuclear area and LRP are related in tumour tissue of the ovary obtained at the debulking operation before the administration of chemotherapy in 40 patients. The mitotic activity index, volume percentage epithelium, standard deviation of nuclear area and the other MDR-related proteins P-glycoprotein (JSB 1, MRK-16) and MRP have been investigated additionally for correlations and prognostic value. No correlations were found between the morphometrical features and MDR-related proteins. Mean nuclear area tended to be larger in LRP positive tumours, but the correlation was not significant. In multivariate analysis LRP protein expression and mean nuclear area had independent prognostic value. Further studies are required to elucidate the biological background of the strong prognostic value of mean nuclear area in advanced ovarian cancer. PMID- 12122281 TI - Telomerase activity levels in the surgical margin and tumour distant tissue of the squamous cell carcinoma of the head-and-neck. AB - The survival of patients with a head-and-neck squamous cell carcinoma is determined by loco-regional recurrence and second primary carcinomas. As a complement to histopathology, molecular changes of tumour marginal and tumour distant tissue may confirm curative surgical tumour extirpation. We tested telomerase activity with PCR-ELISA kits.20 tumour margin biopsies were chosen by the surgeon from 20 patients. In addition, 3 tissue samples were taken from each of 20 additional patients, one from the carcinoma centre, the tumour margin and one distant from the tumour. 50% of the carcinoma centres were telomerase positive. Thirteen of the 40 tumour margin samples showed increased telomerase levels, and in 3 of these residual carcinoma was histopathologically detected. Six of the 20 tumour distant tissues revealed increased telomerase levels. Telomerase positivity in carcinoma-free tumour margins correlated with a good prognosis. Confirmation of the results in a larger patient group is needed. PMID- 12122308 TI - The validity of animal models of predisposition to depression. AB - Some animal models of depression, including the majority of the more recently introduced models, are better characterized as models of predisposition to depression. In the first part of this paper, we show that the basis for such a model could be either a procedure that increases the ease with which an analogue of major depression may be evoked, or a presentation analogous to dysthymia (chronic mild depression). We then consider how the concepts of predictive, face, and construct validity apply to such models. Next, we review the validity of the available models of predisposition to depression, which derive from genetics, genomics, developmental manipulations, and brain lesioning. Finally, we compare the performance of the different models, using a novel scoring system that formalizes the evaluation of animal models against each of the three sets of validation criteria. PMID- 12122282 TI - CD10 expression in non-small cell lung cancer. AB - CD10 is a cell surface endopeptidase that inactivates various potentially growth stimulatory peptides. In lung cancer cell lines this downregulation has been associated with increased proliferation. Downregulation of CD10 in lung cancer tissue is described, suggesting a potential role in carcinogenesis and a possible use of CD10 as a prognostic marker. We aimed to determine the rate of CD10 expression in our non-small cell lung cancer (NSCLC) collection and to clarify its correlation with clinicopathological parameters and patient survival. 114 NSCLC were analysed immunohistochemically using a monoclonal CD10 antibody (clone NCL-CD10-270) on an NSCLC tissue micro array. The staining was semiquantitatively scored. CD10 expression was observed in 19% of cases, without any significant association with tumour type, -size, -grading, nodal status, clinical stage, and patient survival time. We conclude that a diagnostic use of CD10 immunostaining in NSCLC is unlikely. PMID- 12122309 TI - Apomorphine sensitization: evoking conditions, context dependence, effect persistence and conditioned nature. AB - When repeatedly administered a dose of apomorphine (Apo), pigeons, much like rodents, show behavioural sensitization. In birds this sensitization expresses itself as an increasing pecking response to the drug and is found to be partially dependent on the environmental context in which Apo takes effect. In the first experiment we examined what effect different inter-Apo administration intervals have on the development of Apo sensitization and found that, with some smaller variations, intervals between 3 hours and 5 days all yielded comparable courses of sensitization. In the second experiment we examined how long pigeons had to be exposed to the same distinct cage to reveal a maximal context-dependent sensitization. Pigeons were therefore repeatedly injected with Apo and consistently placed in an experimental cage for different lengths of time (5 to 60 min; the overall drug effect lasted for about 1 h) before being returned to their standard home cages. Subsequent tests in the experimental cage and a standard cage showed that 20-min post-injection exposures were sufficient to yield a maximal response in the experimental cage. After training with 20- and 60 min exposures, the pigeons pecked about three times more in the experimental cage than in the standard cage. This confirmed the marked context dependency of the sensitization effect. In the third experiment, groups of pigeons were injected repeatedly with Apo and directly afterwards placed either consistently into the same experimental cage or into different experimental cages. The same-cage group evidenced a significantly much stronger sensitization than the different-cage group. A cage-habituation group served as a control for the possibility that the weaker sensitization of the different-cage group might be due to a cage novelty effect. This cage-habituation group was run under the same conditions as the different-cage group but with additional exposures to the crucial cage while injected with saline. This extra treatment did not augment the pecking response to Apo in that cage. In the fourth experiment we examined how long the sensitization to Apo lasts and found that, even after 2 years of drug abstinence, it only waned to 50% of the original asymptotic response. The overall results support the hypothesis that a very major part of the sensitization to Apo in pigeons is due to a conditioning to the environmental context and to the drug state itself. PMID- 12122310 TI - Stress hormones enhance retrieval of fear conditioning acquired either one day or many months before. AB - It has been known for years that systemic administration of the stress hormones, adrenocorticotrophin (ACTH), lysine-vasopressin, adrenaline, or beta-endorphin, enhances retrieval of aversive behaviours acquired one or a few days before. Here we show that the pre-test i.p. injection of the hormones in rats can also enhance retrieval when given months after the original training. The effectiveness of the treatments changed with time. When animals were tested 3 months after training the hormones enhanced retrieval only at doses five times higher than those needed 1 day after training. Between 6 and 9 months from training the hormones either lost their effect (vasopressin, beta-endorphin) or actually inhibited retrieval (ACTH, adrenaline). The effects of the hormones cannot be explained by a decrease in locomotor activity: none of the treatments had such an effect, as measured in an open field. However, when the animals were tested between 12 and 19 months after training, the hormones once again became as effective as they had been 1 day after training. This was so in spite of the fact that control retention levels became very low with age, probably as a result of extinction. The oscillation of the sensitivity of retrieval to the hormones does not appear to depend on changes in anxiety levels with ageing or to effects of the hormones on locomotor activity. PMID- 12122311 TI - Bupropion and sertraline enhance retrieval of recent and remote long-term memory in rats. AB - Wistar rats were trained in step-down inhibitory avoidance at the age of 3 months, and tested for retrieval either 1 day later or 3, 6, 9, 12, 15 or 19 months later, when the animals were 6, 9, 12, 15, 18 or 22 months old, respectively. Bupropion (20 or 60 mg/kg) and sertraline (3.3 or 10 mg/kg) given orally 6 or 3 h before retention testing, respectively, enhanced retrieval of this task at all training-test intervals, despite the fact that retrieval at the longest intervals was practically not seen in control animals. The effect cannot be explained by influences of the drugs on locomotor activity; the treatments had no effect on open field behaviour at the age of 3, 8 or 21 months. The findings may be relevant to the use of these drugs as cognitive enhancers in elderly subjects. PMID- 12122312 TI - The effects of morphine on chained and clocked fixed-interval schedule performance in pigeons. AB - The present experiment examined the role of stimulus functions and degree of stimulus control on the effects of drugs on behavior maintained by clocked fixed interval (CFI) schedules. Three pigeons pecked keys under a multiple fixed interval (FI) 1-min, CFI 1-min schedule of food presentation (the FI CFI condition). During the FI component, the key was lit amber. During the CFI component, the key colors changed in a regular manner across the interval. Three other pigeons pecked keys under a multiple schedule in which a three-link chained FI schedule alternated with a yoked-CFI schedule (the chain yoked-CFI condition). During the chain component, three successive FI 20-s links were associated with different key colors. During the yoked-CFI schedule, the clock stimuli were the same duration as, or 'yoked' to, the corresponding link in the chain component. Therefore, key colors changed at the same times as in the chain component, but independently of key pecking. All pigeons received a range of doses of morphine (1.0-17.0 mg/kg) and saline. Baseline rates of responding during the FI and chain components were generally higher than during the CFI and yoked-CFI components. Morphine decreased overall response rates during all components. During the FI component, morphine increased response rates at the beginning of the intervals. Morphine did not disrupt the patterns of responding maintained during the CFI, yoked-CFI, or chain components, despite the fact that the chained schedule required responding early in the intervals. This finding suggests that external stimulus control plays an important role in determining the drug effects on behavior maintained by CFI or chained schedules. PMID- 12122313 TI - Orphanin FQ/nociceptin but not Ro 65-6570 inhibits the expression of cocaine induced conditioned place preference. AB - The present study investigated the effect of orphanin FQ/nociceptin (OFQ/N), the endogenous ligand of the opioid receptor-like 1 (ORL-1) receptor on the expression of cocaine-induced conditioned place preference (CPP) in rats. To extend this study, the new non-peptidic compound Ro 65-6570 (8-acenaphthen-1-yl-1 phenyl-1,3,8-triaza-spiro[4,5]decan-4-one), with agonist activity at ORL-1 receptors, was examined. The influence of both compounds on cocaine-induced hyperactivity was also studied. Our experiments indicated that intracerebroventricular (i.c.v.) injection of OFQ/N, at doses of 10 and 20 microg/rat, significantly suppressed the expression of cocaine-induced place preference. Ro 65-6570 (3 and 6 mg/kg, i.p.) did not change the effect of cocaine, although its acute injection in control rats significantly increased the time spent in the drug-associated compartment of the CPP apparatus. The substances exhibited opposite effects on cocaine-induced hyperactivity (OFQ/N suppressed it but Ro 65-6570 increased it). Our results suggest that the effect of OFQ/N on the expression of cocaine-induced CPP may be a result of its influence on dopamine (DA) neurotransmission in mesolimbic structures. Ro 65-6570 does not share this effect with OFQ/N. PMID- 12122314 TI - Impaired spatial learning in the Morris water maze induced by serotonin reuptake inhibitors in rats. AB - The effects of selective serotonin reuptake inhibitors citalopram and fluoxetine on spatial learning were assessed in rats. Adult male rats were subjected to 4 days of training in the Morris water maze with the invisible platform. Animals received different doses of citalopram (1-8 mg/kg; i.p.) or fluoxetine (1-16 mg/kg; i.p.) or their vehicles (saline or distilled water respectively) 30 minutes before training each day. The results showed that citalopram at doses of 4 and 8 mg/kg and fluoxetine at doses of 8 and 16 mg/kg significantly increased latencies to find the platform and traveled distances compared to the control group. Therefore, it appears that selective serotonin reuptake inhibitors can cause learning deficits in complex spatial tasks such as Morris water maze. PMID- 12122315 TI - Effects of Aframomum melegueta and Piper guineense on sexual behaviour of male rats. AB - The effects of aqueous extracts of Aframomum melegueta and Piper guineense on the sexual behaviour of male rats were studied, considering many criteria, such as penile erection, copulatory behaviour and orientation activities towards themselves (genital grooming) and female rats (ano-genital sniffing, mounting). For 8 days different groups of rats received a daily administration of distilled water (control) or a plant extract: A. melegueta at 115 mg/kg or P. guineense at 122.5 mg/kg. Both plant extracts stimulated male sexual behaviour. In fact, A. melegueta and P. guineense significantly increased penile erection index, and the frequencies of intromission and ejaculation. These plant extracts were found to enhance the orientation of males towards females by increasing mounting and ano genital investigatory behaviour. Results of this study showed that A. melegueta and P. guineense modified the sexual behaviour of male rats by increasing sexual arousal. PMID- 12122316 TI - Memantine does not block antiaggressive effects of morphine in mice. AB - The action of the noncompetitive N-methyl-D-aspartate (NMDA) receptor blocker memantine (5, 10, 20 and 40 mg/kg) was evaluated during social encounters in mice. Although a dose-dependent increase in locomotion was observed, only with the highest dose did it reach statistical significance. Aggressive behavior was decreased with 20 and 40 mg/kg of memantine, social contacts being increased only with 20 mg/kg. Subsequently, the effect of these memantine doses on the antiaggressive actions of morphine (10 mg/kg) was evaluated. None of the doses affected the antiaggressive action of morphine. As memantine administration produced an antiaggressive effect only at doses that affected locomotion, it is unlikely that the glutamatergic system mediates the antiaggressive actions of morphine. PMID- 12122317 TI - [The price of a consensus]. PMID- 12122318 TI - [The management of mucocutaneous herpes in the immunocompetent subject]. PMID- 12122319 TI - [Genital herpes: epidemiology, transmission, clinic, asymptomatic viral excretion, impact on other sexually transmitted diseases, prevention, and treatment]. AB - Genital herpes is one of the most widespread sexually transmitted diseases in the world. HSV2 predominates (60-80 p. 100), but the prevalence of HSV1 genital herpes is rising (20-40 p. 100). Erosive lesions of the genital organs due to HSV infection are a factor favoring HIV contamination and other sexually transmitted diseases. The main factor of transmission is asymptomatic viral excretion. Aciclovir, valaciclovir and famciclovir are effective treatment for genital herpes (primary infection, curative and preventive treatment of recurrence), but none of these compounds alters the natural history of the infection. Aciclovir given as a preventive measure reduces the load of asymptomatic viral excretion. Information and education of patients with genital herpes are key elements for prevention. Use of preservatives appears to be effective. New vaccine strategies favoring humoral and cellular response should be studied. PMID- 12122320 TI - [Orofacial herpes and other localizations (genital herpes and neonatal herpes excluded)]. AB - BACKGROUND: Herpes is a pandemic infection in the human species. The purpose of this work was to conduct a critical analysis of the literature devoted to the pathophysiology and treatment of orofacial herpes an other localizations (genital herpes and neonatal herpes excluded). METHODS: We searched for articles in English and French devoted to orofacial herpes and other herpetic localizations indexed in Medline (1980-July 001), EmBase, and the Cochrane Library (1995-July 2001). Critical analysis was based on level of proof using the ANAES methodology. RESULTS: More than 700 articles were identified. One hundred four were selected for this report. Primary HSV1 infection usually occurred before adulthood and involved acute gingivo-stomatitis in the majority of the cases. Several primary HIV1 infections were asymptomatic or aspecific and non-recognized. After the primary infection, the virus remains in the ganglion of the trigeminal nerve in a latent state and can be reactivated sporadically. Reactivation is associated with viral excretion and can be symptomatic (herpetic recrudescence) or not (recurrence). Recrudescence of orofacial herpes is typically labial. No currently available vaccine can prevent acquisition of HSV1. Treatment of acute gingivostomatitis has been standardized and is based on aciclovir. Inversely, the effectiveness of aciclovir and other nucleoside analogs with anti-herpes activity has not been clearly established for prevention or cure of recrudescence of orofacial herpes. Other localizations (hand, anus, diffuse skin localizations in contact sports, Kaposi-Juliusberg syndrome) are much more exceptional. Treatment has not been standardized. DISCUSSION: Despite the abundance of the literature on orofacial herpes, consistent quality is lacking, particularly concerning therapeutic studies. The quality of these reports is generally inferior to those devoted to genital herpes. There has however been a general trend towards improved methodology over the last years. Very little has been reported on exceptional localizations of orofacial herpes. PMID- 12122321 TI - [Signification and limitations of virology diagnostic tools in herpes facialis, herpes genitalis, herpes gestationis and in neonates at risk]. AB - INTRODUCTION: Cutaneomucous herpes is frequent in humans. Laboratory confirmation of clinical diagnosis is important, as there are atypical presentations in genital herpes. METHODS: The review of the French and English literature on virological diagnosis of herpes simplex, indexed in Medline (1990-2001), Embase and Cochran Library (1995-2001) was carried out. RESULTS: 172 publications were selected, and 125 of them were used for this work. The current methods to diagnose herpes in cutaneous lesions are detection of antigens by immunofluorescence or ELISA, culture and PCR. The sensitivity and accuracy of the diagnostic methods are directly linked to the quality of the sampling. Antigen detection is a rapid method, but lacks sensitivity. Culture (the gold standard) is sensitive but sometimes the detection of the virus requires several days. Most of the PCR procedures developed are individual non standardized "in house methods. However, the rapidity and sensitivity of this latter technic make it very useful. Serology allows determination of the immune status of the patients. The development of type specific serology had lead to numerous seroepidemiological studies demonstrating that genital herpes due to HSV2 has increased in most of the countries. CONCLUSION: Rapid and sensitive methods are available for the diagnosis of cutaneous herpes in virological laboratories. However, questions remain for the acceptance of PCR and type specific serology in routine diagnosis. These two technics still have to be standardised before being widely used. PMID- 12122322 TI - [Cutaneomucosal herpes in immunocompetent pregnant women]. AB - Genital herpes is a common sexually transmitted disease. In pregnant women, the risk of fetal contamination is high, particularly during the per and peripartum period. Certain prevention strategies have been changed with the advent of new antiviral agents and treatment protocols at the end of pregnancy remain to be evaluated. Based on a review of the literature, we discuss a few of these different questions raised at the French consensus conference on the management of cutaneomucosal herpes in immunocompetent pregnant women (ocular manifestations excluded). PMID- 12122324 TI - [National survey of dermatologists]. AB - OBJECTIVE: The purpose of this survey was to ascertain current management practices of French dermatologists treating immunocompetent patients with cutaneomucosal herpes (ocular herpes excluded) as a prelude to the French consensus conference on this topic. METHOD: A random sample of French dermatologists were invited to respond to a telephone interview: 928 dermatologists were contacted. RESULTS: The 216 dermatologists who responded to the telephone interview provided care for five persons per month (pregnancy excluded) who consulted for orofacial or genital herpes. Nearly half of the dermatologists stated they do not talk about herpes spontaneously with their patients. When a suspect lesion is seen for the first time, 48 p. 100 of the dermatologists order one or two complementary exams. Their advice on prevention between partners basically concerns use of preservatives. Therapeutic attitudes vary depending on the type of herpes or the number of recurrences per year: 84 p. 100 of the dermatologists prescribe a specific antiviral treatment for patients with solar herpes. Virological proof of infection is not acquired in 84 p. 100 of the cases before initiating a long-term treatment for recurrence. The most widely used agents are valaciclovir 500 and aciclovir 200. CONCLUSION: This survey demonstrates a certain degree of divergence from the recommendations of the consensus conference. The participation rate appears to be satisfactory, but herpes serology is ordered too often and antiviral agents are not used in compliance with current guidelines. This survey will be redone after diffusion of the guidelines in order to evaluate their impact. PMID- 12122323 TI - [Mother-infant and indirect transmission of HSV infection: treatment and prevention]. AB - INTRODUCTION: Neonatal herpes is a serious condition. The objectives of this critical review of the literature are to: 1) define the modes of mother-infant and indirect transmission of HSV infection; 2) determine current treatments and perspectives for the future. METHOD: We searched for articles published since 1980 in databases using a series of key words. The articles were classed into three categories by level of scientific proof: good (level 1), fair (level 2), poor (level 3, 4 or 5). General reviews were excluded. RESULTS: We selected 153 articles and retained 96. Man to woman contamination was generally reported: 10p.100 of the couples were serodiscordant. Presence of anti-HSV1 antibodies was partially protective against HSV2 infection. Neonates can be contaminated in utero via transplacental hematogenic transmission, at delivery (the most frequent route), or during the postnatal period (indirect transmission). The risk of neonatal contamination is greatest for primary infection (PI) or non-primary infection occurring the last month of pregnancy (50p.100), but transmission is low for maternal recurrence during the week before delivery (5p.100). Cesarean section is mandatory in case of genital PI or non-primary maternal infection during the last month of pregnancy, especially in case of membrane rupture<6 hr, but does not protect the infant in two-thirds of the cases. The decision for cesarean is controversial in case of recurrence. Antiviral treatment of the mother using aciclovir (ACV) is well tolerated. ACV-cesarean combination provides maximal protection for the neonate. A neonate with proven or suspected HSV infection should be isolated from other neonates but not from the mother. Breastfeeding is contraindicated in case of breast lesions. Parenteral ACV 60 g/kg/d is preferred over vidarabine. It should be started immediately after the first virology samples. The risk of recurrence is estimated at 7p.100 for all neonates and warrants treatment using a high oral dose (90-100mg/kg/d) due to the low bioavailability, if the number of recurrences is>3 in 6 months. Antiviral treatment is formally indicated if: 1) neonate viral cultures are positive at day 1 and day 3, 2) clinical lesions suggest herpes, 3) neurological disorders or signs of sepsis with negative bacteriology are present and the mother has a history of herpes or contact with labial herpes; and can be discussed if: 4) PI is proven at delivery or during the last month of pregnancy (irrespective of the delivery route, even if the mother is treated or if the membranes are intact), 5) late cesarean (membrane rupture>4 h) with clinical herpes at delivery, 6) vaginal delivery and recurrent herpes within the last month with associated clinical risk factor(s). CONCLUSION: Many points remain to be clarified concerning optimal management of the mother-infant couple in case of maternal herpes during pregnancy or at delivery. New perspectives concerning diagnosis and prevention of neonatal contamination include: identification of asymptomatic primary infections using rapid identification of genital viral antigen during delivery, identification of women with a risk of asymptomatic excretion using specific serology tests for the pregnant woman and her partner, antiviral treatment for men, topical genital treatments, vaccination of women at risk, monoclonal antibodies, new antiviral agents with mechanisms of action independent of viral thymidine kinase. PMID- 12122325 TI - [National survey of gynecologists-obstetricians and general practitioners]. AB - OBJECTIVE: The purpose of this survey was to ascertain current practices of French gynecologists-obstetricians (GO) and general practitioners (GP) treating immunocompetent patients with cutaneomucosal herpes (ocular lesions excluded) as a prelude to the French consensus conference on this topic. METHODS: A random sample of French GOs and GPs were invited to respond to a telephone interview: 330 GOs and 331 GPs were contacted. RESULTS: Response rate was 24 p. 100 among the GOs who could be contacted (49 responses) and 14 p. 100 among the GPs (45 responses). More than half the GPs cared for 1 to 5 cases of orofacial or genital herpes monthly. When a suspect lesion is seen for the first time, 51 p. 100 of the GOs order one or two tests to achieve virology proof: 65 p. 100 order herpes serology and 47 p. 100 PCR. For the GPs, 56 p. 100 do not order a complementary exam. Advice on prevention between partners basically concerns use of preservatives. The therapeutic attitude varies depending on the type of herpes and the number of recurrences per year. The most widely used agent is valaciclovir. A specific antiviral agent is prescribed for non-ocular herpes by 92 p. 100 of the GOs and 98 p. 100 of the GPs. Virological proof of herpes infection is not obtained by 65 p. 100 of the GOs and 78 p. 100 of the GPs before initiating long-term treatment for recurrence. Genital herpes during pregnancy appears to be rare; 57 p. 100 of the GOs had less than one case over the last five years. Only 33 p. 100 of them discuss herpes spontaneously with their consultants. When a suspect genital lesion is observed during pregnancy, 82 p. 100 seek virological proof. When there is a risk of neonatal herpes, the type of delivery depends on the type of herpes infection and the date of occurrence during the pregnancy. Cesarean section is performed systematically by 63 p. 100 of the GOs when there is a primary herpes infection after 34 weeks gestation. Most practitioners did not respond concerning their attitude for exams and surveillance for the infant. DISCUSSION: The number of responding practitioners was too small in this study to provide a representative sample. This study will be renewed with a larger sample of GOs in order to obtain reliable data. For general practitioners, there is a need for education concerning management of cutaneomucosal herpes which should be provided at the same time as the consensus conference guidelines are distributed. PMID- 12122326 TI - [Natural history of HSV1 and HSV2 transmission modes and epidemiology consequences of HSV infection on HIV infection. Prevention]. AB - Both Herpes simplex viruses HSV1 and HSV2 are transmitted by direct mucosal or cutaneo-mucosal contact between individuals. HSV1 is the leading cause of orofacial herpes and HSV2 the most frequently encountered cause of genital herpes. There are however a number of environmental and behavioral factors that modify the epidemiological pattern in both infections. These factors also affect virus dynamics and spread. In developing countries, HSV1 infections continues to be acquired in early childhood. In developed countries, displacement of this acquisition towards adolescence and adulthood explains, in part, the increase in genital herpes caused by HSV1. HVS2 infection progresses in the sexually active population worldwide. Although the rate of seroprevalance varies greatly from one continent to another, women are still more often infected than men. HSV2 genital infection is a cofactor for transmission and acquisition of HIV, which, in certain African regions where the two infections are highly prevalent, explains in part the progression of the HIV epidemic. Until a vaccine becomes available, the prevention depends on abstention from all oral and genital contact during periods of active disease. For genital herpes, use of a preservative has only a relative protective effect and the contribution of suppressive treatment in potentially contaminated subjects is under evaluation. PMID- 12122327 TI - [Natural history of HSV1 and HSV2 infection. Asymptomatic viral excretion. Mother infant transmission. Indirect transmission]. AB - Recurrent asymptomatic viral shedding accounts for most of the oral, sexual and neonatal transmissions of herpes simplex infections (HSV). In immunocompetent individuals asymptomatic shedding, as determined by culture, occurs during 2 8p.100 of days, and more for persons with HIV infection or if measured by PCR. Antiviral therapy dramatically decrease but does not eliminate asymptomatic shedding. The main risk of HSV transmission to the neonate is during vaginal delivery from infected asymptomatic mothers who acquire HSV genital infection late in pregnancy. Because the survival of HSV out of the oral-genital secretions is weak, indirect and/or nosocomial transmisions of HSV are very rare and should be controlled by common-sense precautions. The prevention of the acquisition of genital or neonatal HSV infection is a challenge because it is based on the understanding and the control of asymptomatic shedding. PMID- 12122328 TI - [Genital herpes in immunocompetent men and non-pregnant women Clinical aspects, accuracy of clinical examination, evolution]. AB - Use of new specific herpes serology tests permits to better study the natural history of herpes infection and define primary infection, primary manifestation, and recurrence The important points to consider are the clinical manifestations of genital herpes PMID- 12122329 TI - [Non-genital herpes simplex virus infections in adults. Clinical aspects, accuracy of clinical signs and evolution]. PMID- 12122330 TI - [Herpes simplex in children. Clinical manifestations, diagnostic value of clinical signs, clinical course]. AB - Herpetic gingivostomatitis (HGS) is the predominant manifestation of cutaneomucosal herpes in children with HSV1 primary infection before the age of 3 years. The infection is self limiting and lasts 10 to 14 days. Pain and dysphagia are particularly important during the first week of infection and may necessitate parenteral rehydratation and administration of antalgesics. HGS in the young child causes substantial morbidity leading to hospital and social costs (work stoppage for parents). The clinical course is generally benign with the exception of forms with important extension, eczema, herpeticum Kaposi-Juliusberg pustulosis observed at this age only in children with atopic dermititis. Other severe forms are observed in the neonate and immunodepressed subject, which can also be caused by HSV1. Forms with little or not clinical manifestation predominate and generally go undiagnosed, explaining the asymptomatic viral excretion observed in the saliva or other secretions (ocular, genital secretions). Despite the sterotypic nature of the clinical expression, HGS is still often undiagnosed both by general practitioners and pediatricians. This lack of diagnosis generally has few consequences due to the benign course in a few days, but the infection can have an important psychological and social leading to significant healthcare costs. Moderate and severe forms require medical care. Aciclovir should be prescribed if the diagnosis is made early (3 days) in combination with symptomatic care. Studies of the medical and economic impact of herpetic gingivostomatis should be conducted. PMID- 12122331 TI - [Management of mucocutaneous herpes simplex virus infections in immunocompetent patients: signification and limits of antigen detection culture methods and antibody detection]. AB - Herpes simplex virus (HSV) infections are very common and may present various manifestations. Mostly asymptomatic, often mild, these infections may become life threatening, specially in neonates. The acute and rapid diagnosis of HSV infections can prevent infections in these patients and is also helpful to confirm clinical diagnosis. The sensitivity of "classical" diagnosis methods, such as culture and antigen detection by immunofluorescence and ELISA, is highly dependent on the quality of the sample. Antigen detection techniques give results in short delays, compatible with the initiation of antiviral treatment; however their sensitivity decreases when lesions get older and they are not convenient for the diagnosis of asymptomatic infections. Isolation of HSV on cell culture remains the gold standard, because it is easy to perform and exhibits good sensitivity whatever the stage of the lesions; the isolation of the virus is required for HSV typing and to determine susceptibility to antiviral agents in case of clinical resistance to the treatment. Serodiagnosis is helpful to determine the immune status of a patient and to distinguish between primary infection and recurrence. It is now possible to differentiate HSV1 and HSV2 antibodies; the benefit of type-specific serology is clear for epidemiological surveys but may be discussed for individual follow-up, specially for genital herpes, both for the diagnosis and the prevention of HSV infections. PMID- 12122332 TI - [Significance and limits of PCR diagnosis in orofacial and genital herpes simplex virus infection in the pregnant woman and neonate]. AB - The availability of methods of detection of DNA by PCR amplification has deeply modified the knowledge on herpes simplex virus infections. These methods have allowed a better understanding of the physiopathogeny of the disease. In particular, PCR has revealed the importance of asymptomatic viral shedding in infected patients. PCR helps diagnosis in many situations where viral isolation by culture proves difficult or impossible, i.e. in treated or atypical lesions, or in newborn central nervous system infections. In addition, PCR has underlined the existence of prolonged viremia in infected newborns. PCR helps sequencing of the viral genome for further epidemiological studies or analysis of resistance to antiviral drugs. Recent PCR-derived techniques have been developed to quantify viral load in real time, and allow to obtain a diagnosis in a few hours. Despite these major advances, the use of PCR in several clinical situations still needs to be further validated. PMID- 12122333 TI - [Antiviral and non-antiviral general treatments for oro-facial and genital herpes (pregnancy and neonates excluded)]. AB - General treatments for immunocompetent individuals with herpes simplex infections are based on the use of antiviral agents which constitute the only treatment with proven efficacy. Antivirals were developed in the 1980s with aciclovir (ACV) as the leading compound and have greatly changed management. However, once the virus has penetrated the organism, it cannot be eradicated, neither by the immune system nor by antiviral agents. This viral resistance is basically related to its capacity to maintain itself in a latent form in the sensorial ganglions. ACV is the first line treatment, used since the 1980s; other antiviral agents are also available. PMID- 12122334 TI - [Local treatments using antiviral and non-antiviral drugs for herpes facialis and genitalis (excluding pregnant females and neonates at risk)]. PMID- 12122335 TI - [Genital herpes and pregnancy. Preventive measures for neonatal HSV infections: 1993 consensus conference and new proposals]. AB - The following document contains four parts The proposals distinguish four maternal situations It is proposed that cesarean section and aciclovir prophylaxis be reserved for women in the first two situations. The proposals also encourage systematic use of condoms during pregnancy and information for the mother before discharge from the maternity ward concerning surveillance of the newborn at risk. Systematic serology tests for HSV type 1 or 2 or PCR are not proposed. PMID- 12122336 TI - [Antiviral and non-antiviral local and general treatments for herpes in the pregnant woman (including prevention of mother-infant transmission): alternative propositions]. AB - Herpes may be manifest during pregnancy as a primary infection, as a recurrent infection, or as asymptomatic excretion. Genital herpes can cause neonatal contamination with rare but very serious consequences for the child if specific treatment is not given rapidly. The gravity of neonatal herpes warrants prophylactic measures including cesarean section if the risk is high. Cesarean section must be considered as a prophylactic measure and by consequence may be performed in some cases when not absolutely necessary. PMID- 12122338 TI - [Perspectives for the treatment and prevention of herpes simplex infections]. PMID- 12122340 TI - [Cancer epidemiology: what next?]. PMID- 12122337 TI - [Modalities of treatment local and general, medicamentous or not, controlling neonate suspected to be infected/contaminated by HSV1 or HSV2]. AB - Neonatal herpes infection is secondary to pre/perinatal viral contamination from mother by HSV2 (70 p. 100) or HSV1 (30 p. 100). Incidence in French population is closed to 3/100,000 live births corresponding to 20 cases per year. Risk for maternal viral transmission to the neonate is 30 p. 100 with genital herpetic primo infection and 3 p. 100 in recurrence. However, in 70 p. 100 of cases, maternal history is not contributive. Three main clinical presentations are described However atypical symptoms - as isolated fever - can be a telltale sign. Mean clinical delay from birth to first clinical symptoms is 6 to 12 days and neonate is usually symptom - free at birth. Viral cultures from pharynx, stools, cutaneous lesions and specific PCR in blood and cerebrospinal fluid confirm the diagnosis. Curative treatment is acyclovir at high dosage - 60 mg/kg/d - during 14 days for localized forms and 21 days for neurological and disseminated diseases. Compared to conventional dosages, this treatment leads to a reduction in mortality which however remains high in disseminated forms, 31 p. 100 and 6 to 11 p. 100 in CNS infection. Morbidity is also high in survived patients, 17 p. 100 and 31 p. 100 respectively. Efficacy of prophylactic viral decontamination by anti-herpetic eye drops and cutaneous polyvidone iodine bath, which is largely used at birth in France, has never been evaluated. PMID- 12122341 TI - [Estimate of regional prevalence of colorectal cancer in France]. AB - BACKGROUND: Colorectal cancer prevalence is an important determinant of the health demand that completes information provided by cancer incidence. Current estimations established from data for the years 1985 and 1995 can be used to establish a precise description of changing healthcare needs for colorectal cancer. METHOD: Prevalence estimates method were based on incidence data computed on the regional scale by the FRANCIM network and mortality data provided by INSERM. We used the relationship that exists between the net risk of cancer, the net risk of dying of the given cancer and the age-specific prevalence of cancer. RESULTS: In 1995, the prevalence of patients who had a diagnosis of colorectal cancer amounted to 200 000 persons. The estimated number of prevalent cases was never lower than 3500 in any region and in 7 regions this number was higher than 10 000. From 1985 to 1995, there has been an increase of 35% in the prevalence rates. CONCLUSION: The evaluation of the number of persons who have had a diagnosis of colorectal cancer provides knowledge for health care planning. Such information on the regional scale is very useful for the health organisation (SROS). This geographical level induces difficulties not encountered at the national level. PMID- 12122342 TI - [Influence of social and occupational class and area of residence on management and survival in patients with digestive tract cancer: a population study in the Calvados area (France)]. AB - Several studies have shown socio-economic differences in cancer survival, low socio-economic level being associated with poor prognosis of cancer. The aim of the study was to investigate the influence of social environment on care procedures for treatment in cancer and to determine to what extent well established socio-economic differences in cancer prognosis can be explained by such an influence. A retrospective analysis was conducted on patients having had a digestive cancer in the department of Calvados (France) between 1978 and 1990 collected by the local digestive cancer registry (1534 males and 1060 females). Jobless male and female farmers visited private specialists and were treated in specialized care centres at a rate two-fold lower than people in higher social classes. However, our results suggest that these variations do not explain all the influence of social environment on cancer survival either in males and females. PMID- 12122343 TI - [School-related injuries: incidence, causes, and consequences]. AB - BACKGROUND: School accidents are frequent but little epidemiological information is available to guide prevention. In this study we examined the incidence, causes, and consequences of school accidents as a function of the pupil's characteristics. METHODS: An epidemiological study was conducted in all 2 396 adolescents attending two secondary school groups. Sociodemographic characteristics of the pupils and data on school accidents during a one-year period were collected using a questionnaire filled out by the school nurse in the presence of the victims. The chi-square independence test, Fisher's exact test and the logistic regression method were used for the statistical analysis. RESULTS: Sports and physical training (SPT) accidents accounted for 52.8% of the accidents, recreation accidents for 12.7% and other accidents for 33.6%. The annual incidence of one accident or more, for all types of accidents combined, was 12.9%, that for two or more accidents 2.3%. The rate of SPT and recreation accidents decreased strongly with age. SPT accidents were more frequent in girls, the other accidents more frequent in boys. Among the SPT accidents, 69.2% occurred under training conditions and 33.7% were caused by another person. Causes mentioned by the victims were: carelessness (26.0%), clumsiness (17.5%), misappreciation of risk (13.8%), tiredness (9.5%), nervous irritation (8.6%), rowdyism (6.0%), disrespect of the teacher's instructions (6.0%). The lesions were: contusions (50.7%), wounds (18.7%), tendinitis (11.7%), wrenches (9.2%), others (7.3%). They differed between age groups, sex, and category of sports. Localizations were mainly: fingers (27.4%), other localizations of the upper limb (20.1%), head (20.6%). A physician was consulted for 19.5% of the accidents and hospitalization followed 2.7%. Absence from school and exemption from SPT were frequent (11.4% and 16.3% respectively). CONCLUSION: The results could be used to inform adolescents so they and their families could become more aware of the risk of school accidents. Prevention should mainly focus on the younger children. An effort must be made regarding risk assessment in order to help the pupils become more careful and responsible during their sports activities. The choice of these activities and the materials used should be made more suitable for adolescents. PMID- 12122344 TI - [Analysis of biases in epidemiological knowledge of road accidents in France]. AB - BACKGROUND: In France, as in many countries, road casualty statistics are mostly based on police reports. It is generally recognized that these data are incomplete but no measurement has been made of the degree of under-reporting and thus of associated biases. This study aims to demonstrate and quantify these biases. METHODS: The study compares, after data linkage, the 10,202 people reported injured or killed in 1996 in the medical road accident victims Register in the Departement du Rhone (France), with the 4,572 victims reported by the police during the same year and in the same area. This Departement was chosen, as it is the only region in France where these two independent data sources coexist. Two types of possible biases are studied: injury severity classification bias and selection bias induced by underreporting. RESULTS: The study shows that the definition of "serious injury" used by the police exaggerates the severity of the victim's condition in over half the cases. This bias depends on road user group. This bias is maximum for pedestrians: compared to a slightly injured car occupant, a pedestrian with the same injury severity level has significantly more chance to be considered as severely injured (RR=1.78; 95% CI: 1.11-2.87). Conversely, significant selection biases are related to data collection by the police. The multivariate analysis shows that the underreporting of victims increases if no third party is involved (i.e. without any other vehicle or pedestrian), and reduces with injury severity. It also varies by road user group (with the largest underreporting for cyclists). Among the most seriously injured in accidents involving third parties, motor cyclists and car users are the most reported category and pedestrians the least (RR=0.80; 95% CI: 0.70-0.92). Biases in Register selection are much more limited and basically concern underreporting of victims of minor accidents who did not require medical care. CONCLUSIONS: This study confirms and quantifies misleading distortions in police statistics used to assess road accidents. These results concern the relevant indicators to be used to define road safety issues. PMID- 12122345 TI - Evolution of blood lead levels in urban French population (1979-1995). AB - BACKGROUND: The aim of the Council Directive of 29 March 1977 of the European Union was to measure non-occupational lead exposure levels in the general adult populations of European countries through biological monitoring. In France, such measurements were carried out during 1979 and 1982 in eight metropolitan areas (having more than 500 000 inhabitants), a period during which the lead content of petrol was decreased. The aim of this study conduct in 1995 was to evaluate the exposure trend to lead. METHODS: In 1995 this measurement was repeated, only in the three largest urban areas (Paris, Marseilles and Lyons). The same sampling method used in the first two campaigns was retained to ensure that the results of 1995 could be compared with those from 1979 and 1982. RESULTS: In these three metropolitan areas, the average blood lead levels decreased by the order of 60 microg/l between the beginning of the 1980's and 1995. This represents a fall of more than 50%. CONCLUSIONS: Certainly car pollution is not the only vector of dissemination of lead in the centre of urban zones, but it is there that the most sustained efforts at eradication have been made. The improvement we have observed is probably due to the policy of eliminating lead from petrol. In conclusion, the blood lead levels in French urban populations seem to have greatly decreased from those of the early 1980s. PMID- 12122346 TI - [Is compliance to current lead regulations safe enough for infants and toddlers?]. AB - BACKGROUND: Neurobehavioural and cognitive impairment may arise from children's exposure to lead. To prevent this risk, a Tolerable Daily Intake (TDI) has been issued by WHO and many regulations addressing lead emission to the environment have been introduced. Taking into account both lead concentrations in soil and the importance of soil in children's exposure, we examined whether compliance to the above current regulations is safe enough for children. METHODS: Exposure scenarios were devised for infants and toddlers, the 2 populations most sensitive to lead toxicity, considering three typical environmental settings, that is, rural, urban, and in the vicinity of lead emitting industrial sites. For all 3 scenarios, we used available data describing the current French levels of both lead contamination in various media (water, air, soil) and lead intake through food. Acute lead intoxication from old lead-containing paints was excluded from this study. RESULTS: We have shown that WHO's TDI is exceeded in several situations for both populations, and thus children may be subject to unacceptable levels of risk. CONCLUSION: We conclude that it may be advisable to take fuller account of the contribution of soil to lead exposure. PMID- 12122347 TI - [Air quality monitoring and personal exposure of children to NO(2) and fine particles]. AB - BACKGROUND: Personal exposure to air pollutants and ambient air measurements are poorly correlated in the short term. Nevertheless, air quality surveillance data are often used to characterize exposure in epidemiological studies. This work explores a method to derive exposure estimates for a population of children, through appropriate usage of surveillance data that allows for heterogeneity of life environments. METHODS: Personal exposure (PE) to PM2.5 and NO(2) of 66 to 184 children was measured in 4 French metropolitan areas (Grenoble, Nice, Toulouse and Paris). The proposed approach provides an estimate of a "translator parameter". This method was applied to subgroups of children who differed in terms of daily time spent in areas more or less influenced by traffic emissions. RESULTS: Ambient air concentrations of NO(2) overestimated personal exposures, on average, but children whose life environments are more influenced by traffic exhausts exhibit, on average, greater PE values; as far as particles are concerned, air quality surveillance and PE values are closer. Hence, translation parameters differ according to pollutants, cities and populations. CONCLUSIONS: These results suggest that ambient air monitors can be used to assess exposure of urban populations living in areas with variable traffic intensities. However, usage of these air quality surveillance data should allow for population and pollutant characteristics. PMID- 12122348 TI - [Participation and medical follow-up in screening of colorectal cancer in France within the SU.VI.MAX study]. PMID- 12122349 TI - [Atmospheric pollution and health: not patently obvious. and yet!]. PMID- 12122350 TI - [Comment: The slow dissolution of the relationship between health and revenue disparity]. PMID- 12122351 TI - [Inserting a percutaneous endoscopic gastrotomy tube in an elderly patient may be a difficult decision]. PMID- 12122352 TI - [Prognosis factors of short and long-term survival in elderly hospitalized patients after percutaneous endoscopic gastrostomy]. AB - OBJECTIVE: Percutaneous endoscopic gastrostomy (PEG) is the procedure of choice to achieve long-term enteral nutrition. The risks and benefits of PEG in elderly hospitalized patients have been poorly documented. The objective of this study was to describe the outcome of elderly patients one-year after insertion of a PEG tube. PATIENTS AND METHODS: Hospital records of 73 patients who underwent PEG for enteral nutrition were reviewed retrospectively. Data on patient age and sex, preexisting medical conditions such as dementia or pressure sores, indication for PEG, concomitant infection, complications of PEG and death were obtained from the hospital charts. RESULTS: The main indication for PEG was anorexia (49%). Before insertion of the gastrostomy tube, 44% of the patients had pressure scores, 30% had concomitant infection, 45% had dementia. PEG complications were observed in 51 patients. The survival rate at 1, 6 and 12 months was 0.68 [95% confidence interval - CI 95%: 0.56-0.78], 0.48 [CI 95%: 0.36-0.59] and 0.37 [CI 95%: 0.26 0.48] respectively. The presence of an infectious disease or of pressure sores at the time of PEG tube insertion were independently associated with mortality. Median survival of patients with these two factors was 32 days [CI 95%: 11-98]. CONCLUSION: According to these results, the PEG tubes should be inserted with a delay from infectious diseases and before the occurrence of pressure sores. PMID- 12122353 TI - [Percutaneous endoscopic gastrostomy in elderly patients. A prospective study in a geriatric hospital]. AB - OBJECTIVES: The purpose of this work was to search for prognostic factors after percutaneous endoscopic gastrostomy (PEG) for enteral nutrition in geriatric patients by studying complications, nutritional benefits, and impact on quality of life. METHODS: In this prospective study, 59 elderly patients referred for PEG were followed for 1 year or until tube removal or death. Complications, tolerance to enteral nutrition, nutritional status, infection, bedsores and quality of life were assessed by a questionnaire at tube insertion, at 1 month and every 3 months. Multivariate analysis was performed to look for factors predictive of early mortality before one month. RESULTS: Insertion of the PEG tube was always successful. Pneumonia in the week before tube insertion was predictive of early mortality (odds-ratio: 8.77 [1.63-47.2], P=0.01). Thirty-day mortality was 25%, but was never related to PEG tube insertion. During follow up, no local complication was observed and enteral feeding was well tolerated. After 3 months, serum albumin and prealbumin levels increased (P<0.001). There were fewer infections (P<0.001) and bedsores remained unchanged. Quality-of-life scores were not modified. At one year, the PEG tube was removed in 16 patients who resumed normal oral nutrition, and 6 other patients were able to return to their home. CONCLUSION: In a cohort of aged institutionalized patients, PEG for enteral nutrition was well tolerated and not definitive in more than one-quarter of them. Active lung infection is a risk factor of early mortality. PMID- 12122354 TI - [Factors predictive of complete resection of operable esophageal cancer: review of 746 patients]. AB - OBJECTIVE: Surgery is the treatment of reference for early-stage esophageal cancer, but 5-year survival is only 20% to 25%. After complete resection (R0), survival is significantly longer than after incomplete resection, with microscopic (R1) or macroscopic (R2) penetration. The purpose of this work was to identify retrospectively the factors predictive of complete resection of operable esophageal cancers. PATIENTS AND METHODS: Between January 1982 and March 2001, 746 patients with esophageal cancer underwent curative surgery. R0 resection was performed in 585 patients (78.4%), R1 in 61 (8.2%) and R2 in 100 (13.4%). Univariate and multivariate analysis included 28 preoperative, clinical, tumor and therapeutic parameters. RESULTS: Multivariate analysis showed that factors predictive of complete resection R0 were: absence of any modification of the esophageal axis on the barium swallow (P=0.054), a partial or complete response to preoperative radio-chemotherapy (P=0.042), tumor height<10 cm (P=0.1) and tumor diameter<30 mm (P=0.01). Three groups of patients were identified from the 2 most significant variables. Group 1: no deviation of the axis on the barium swallow (n=501). Group 2: deviation of the axis on the barium swallow and partial or complete response to radiochemotherapy (n=91). Group 3: deviation of the axis on the barium swallow and no response to radiochemotherapy or no preoperative treatment (n=126). For the three groups, rate of R0 resection was 82.6%, 80.1% and 61.1% and 5-year actuarial survival 36%, 27% and 14%, respectively. These rates were significantly different between groups (P<10(- 4)) and two by two (P<0.04). CONCLUSION: Complete resection of esophageal cancer is predictable. After validation with an independent population the findings presented here could be used to establish stratification criteria for future therapeutic trials. PMID- 12122355 TI - [Positron emission tomography in digestive tract cancer]. PMID- 12122356 TI - [Non alcoholic steatohepatitis: the future in hepatology?]. PMID- 12122357 TI - [Non alcoholic steatohepatitis: a multifactorial, frequent, paucysymptomatic liver disease with a fibrotic outcome]. AB - BACKGROUND: The aim of this study was to evaluate the morphological, clinical and biochemical characteristics of non alcoholic steatohepatitis to understand its pathogenesis. PATIENTS AND METHODS: From January 1993 to June 2000, 44 patients were selected on histological criteria. Alcohol intake, blood pressure, weight, glycaemia, lipid, immune, iron profiles hemochromatosis (HFE) gene mutations were analyzed. Patients were re-examined thereafter or in June 2000. RESULTS: Twenty one women and 10 men were included (mean age=54). Nineteen patients were asymptomatic (61.3%). Patients often presented with an increase in alanine aminotransferase. This was correlated with steatosis (P=0.008). Hypertension, excess weight, abnormal serum glucose levels and dyslipidaemia were respectively observed in 10 (32.2%), 24 (77.4%), 16 (51.6%) and 18 (58.1%) patients. Thirteen of these patients (41.9%) presented abnormal autoantibodies titers without autoimmune hepatitis; 18 (58.1%) presented an iron overload. A mutation of the HFE gene was detected in 14 of 25 patients (51.6%). Liver iron concentrations were not correlated to the extent of fibrosis extension or with mutations. CONCLUSION: Increased alanine aminotransferase levels usually revealed non alcoholic steatohepatitis. A high prevalence of autoantibodies, iron overload and mutation of the HFE gene were detected. Non alcoholic steatohepatitis should be diagnosed because it can be associated with cirrhosis. PMID- 12122358 TI - [Liver repopulation: the selective advantage concept]. PMID- 12122360 TI - [Management of hepatitis B in 2002]. PMID- 12122361 TI - [Chronic hepatitis B: natural history and treatments]. PMID- 12122362 TI - [Questions to professeur Patrick Marcellin]. PMID- 12122363 TI - [Pre-C mutations of the HBV]. PMID- 12122364 TI - [HBV cirrhosis]. PMID- 12122365 TI - [Co-infection HIV/HBV]. PMID- 12122367 TI - [Gastric cancer with lymphoid stroma: a rare form associated with Epstein-Barr virus. Five cases]. AB - We report the cases of 5 patients with gastric cancer with lymphoid stroma, aged from 40 to 66 years. The tumor was located in the upper and the middle third of stomach in four patients. Using in situ hybridization, the tumor was Epstein-Barr virus-positive in all patients whereas immunohistochemical analysis was negative. Four patients had total gastrectomy associated in two cases with splenectomy and caudal pancreatic resection. The last patient had subtotal gastrectomy. One patient was lost to follow-up. After a mean follow-up of 31 months, 4 patients are alive and free of local recurrence. PMID- 12122368 TI - [Gastric interdigitating reticulum cell sarcoma: unusual presentation of a histiocytic tumor]. AB - Histiocytic sarcoma, proliferation arising from immunoregulatory effector system cells, is a very rare and recently recognized tumor. Diagnosis is based on immunohistochemical and molecular genetic study, which allows to distinguish histiocytic sarcoma from lymphocytic proliferation, such as non-Hodgkin's lymphoma. We report the case of a gastric interdigitating reticulum cell sarcoma, treated by resection and chemotherapy. We review here clinical, histological, immunohistochemical and genotypic features of histiocytic tumors of the digestive tract. PMID- 12122369 TI - [Secondary amyloidosis complicating ulcerative colitis]. AB - We report a case of amyloidosis secondary to ulcerative colitis. A 33-year-old man with total ulcerative colitis was treated by proctocolectomy 3 years and 5 months after the onset of the disease. One year later, a nephrotic syndrome revealed a secondary amyloidosis that led to end-stage renal failure 14 years later. Colectomy could have delayed renal failure. During the 27-year time of follow-up, ulcerative colitis was the only detected disease possibly responsible for secondary amyloidosis. PMID- 12122370 TI - [Pneumocystis carinii and cytomegalovirus pneumonia after corticosteroid therapy in acute severe alcoholic hepatitis: 2 case reports]. AB - Most cases of infections described after steroid treatment for severe acute alcoholic hepatitis are of bacterial origin. However, the rate of bacterial infections in these patients is not higher than in those who are not treated by steroids. The opportunistic infections are even more rare. We report two cases of patients with cirrhosis and human immunodeficiency virus, treated for alcoholic hepatitis with steroids and who subsequently developed severe pneumopathy due to Pneumocystis carinii. One patient had a concommitant cytomegalovirus infection and both of them died. Pneumocystis carinii infections usually occur in patients a decreased immune cellular response. Steroid treatments and also alcohol may be responsible for these opportunistic infections. Alcohol may have an immunosuppressive effect by decreasing recruitment of CD4 and CD8 lymphocytes to the lungs. In conclusion, Pneumocystis carinii pneumonia is a potential complication of steroid treatments for acute alcoholic hepatitis and should be suspected in case of unexplained pulmonary infection. PMID- 12122371 TI - [Angioedema in Crohn's disease possibly due to mesalazine]. PMID- 12122372 TI - [Oxetorone-associated lymphocytic colitis]. PMID- 12122373 TI - [Rectal linitis as the first manifestation of a prostatic cancer]. PMID- 12122374 TI - [The management of patients who test positive for hepatitis C in a free and anonymous testing centre is critical for follow-up]. PMID- 12122375 TI - [Hepatobiliary distomatosis: a mistaken cause of cholangitis]. PMID- 12122377 TI - Needed: a summer of healing and learning. PMID- 12122379 TI - Use of directed history and behavioral indicators in the assessment of the child with a developmental disability. AB - It can be very difficult to get a complete history and review of systems for children with developmental disabilities and poor communication skills. In addition, many children with developmental disabilities may engage in self injurious or aggressive behavior. Although the causes of inappropriate behavior are frequently environmental, physiologic components may exist as well, particularly pain or discomfort. History taking must be focused and specific and may need to focus on the child's behavioral patterns, because the child may not have sufficient communication skills to describe his or her problem and parents or guardians may not realize the relevance of certain behaviors. Gastrointestinal problems in particular may be a source of discomfort and should be reviewed with particular care. Referral to a psychologist who is able to perform a functional analysis of behavior may be necessary to treat problem behavior, especially if medical causes have been ruled out. PMID- 12122378 TI - Diagnosis and treatment of the child with a draining ear. AB - Treating a child with a draining ear is a common occurrence in the pediatric primary office. The symptoms of otorrhea are broad, and a multitude of factors must be considered in arriving at a diagnosis. The assessment begins with a thorough history of the frequency, duration, and characteristics of the drainage. Physical examination of the affected ear requires cleansing of the external auditory canal before the tympanic membrane can be accurately assessed. The ear must be adequately visualized for accurate diagnosis and treatment. Differential diagnoses include acute suppurative otitis media, otitis externa, granuloma, and bullous myringitis. The acutely draining ear is frequently an uncomplicated suppurative event that will respond well to a regimen of aural hygiene and topical therapy. Patients with a chronically draining ear should be referred to an otolaryngologist for further diagnostics and aggressive therapy. Children with tympanostomy tubes are especially at high risk for suppurative complications. Avoiding the introduction of water into the ear, protecting the ear from water, and prophylactic topical treatment are options that have been suggested for prevention of otorrhea. PMID- 12122380 TI - Teaching pediatric health assessment: using Internet capabilities. AB - Changes brought about by rapidly expanding information technology are affecting many aspects of life, including nursing education. Graduates must be comfortable and flexible with the use of information technology because they will be required to use it in their practice and continuing education activities. In this article, the authors describe their experience in implementing and refining the use of World Wide Web-based technology to teach pediatric health assessment to pediatric and family nurse practitioner students. The article includes reflections on preparation, implementation, and evaluation. The students' level of confidence in their ability to perform pediatric health assessment rose, as did their test scores, and faculty deemed the revision successful and timesaving for them. PMID- 12122381 TI - Child sexual abuse: psychosocial risk factors. AB - Child sexual abuse is a problem of epidemic proportions in the United States. Child sexual abuse has been recognized as a predictor of many physical and psychological problems. It is important that clinicians have the ability to recognize the psychosocial dynamics present in families in which child sexual abuse takes place. Studies have shown that early detection and treatment of child sexual abuse leads to better outcomes for the victims. The literature discusses psychosocial risk factors present in families of child sexual abuse victims. An understanding of these characteristics may enable professionals to identify children at risk for child sexual abuse and may lead to earlier detection, protection, and treatment for victims. A case study illustrates psychosocial characteristics present in the family of a sexual abuse victim. Implications for practice are discussed, and a plan to assess families for psychosocial risk factors and intervene appropriately is outlined. PMID- 12122382 TI - Understanding parental concerns about immunizations. PMID- 12122383 TI - Roles of maternal folate and calcium use. PMID- 12122384 TI - Infant with feeding difficulties. PMID- 12122385 TI - Lessons learned at the Nurse In Washington Internship (NIWI). PMID- 12122387 TI - Symptoms of acute coronary syndromes: are there gender differences? A review of the literature. AB - Evidence has begun to accumulate that suggests there may be gender differences in the presenting symptoms of acute coronary syndromes (ACS). Identification of gender differences has implications for both health care providers and the general public. Women should be instructed as to the symptoms expected with ACS on the basis of evidence obtained from studies that include both sexes. Twelve studies that identified symptoms of ACS for both women and men were identified through a review of the literature. In several of the studies, which included all types of ACS, women had significantly more back and jaw pain, nausea and/or vomiting, dyspnea, indigestion, and palpitations. In a number of the studies, which solely sampled patients with acute myocardial infarction, women demonstrated more back, jaw, and neck pain; nausea and/or vomiting; dyspnea; palpitations; indigestion; dizziness; fatigue; loss of appetite; and syncope. Men reported more chest pain and diaphoresis in the myocardial infarction sample. Results of these studies showed that women and men experienced the same symptoms with ACS. However, in some studies there were gender differences in the proportion of symptoms. Given the current state of the science, definitive conclusions regarding gender differences in the symptoms of ACS cannot be drawn. Further study is urgently needed to clarify and expand on these findings. PMID- 12122388 TI - Perception of risk for coronary heart disease in women undergoing coronary angiography. AB - BACKGROUND: Even though coronary heart disease (CHD) is the leading cause of death among women in the United States, most women underestimate their risk of developing CHD. DESIGN: Survey to examine the relationship between women's recollection of being told they were at risk for CHD and the presence of risk factors. SETTING/PARTICIPANTS: A convenience sample of 450 women undergoing coronary angiography at 1 university hospital. MAIN OUTCOME MEASURES: Self recollection of being told one was at risk for CHD and presence of CHD risk factors. RESULTS: Most women (83.6%) had 3 or more risk factors, 12.2% had 1 or 2 risk factors, and 0.9% had no risk factors. Only 35% of women recalled being told that they were at risk for CHD. Few relationships were found between being told one was at risk for CHD and the presence of individual risk factors. No difference was found in the mean number of risk factors among women who did and did not recall being told they were at risk. In logistic regression analysis, only 5% of the variance in recollection of being told one was at risk was predicted, with only age, education, and having a high cholesterol level significantly contributing to the equation. CONCLUSIONS: Even though women may not remember conversations with their health care provider about CHD risk, the possibility that risk factors were not adequately assessed cannot be discounted. Patient-provider conversations about CHD risk factors should be encouraged as the first step toward successful risk reduction. PMID- 12122389 TI - Evolution of information on women and heart disease 1957-2000: a review of archival records and secular literature. AB - This project explores the evolution of public information on cardiac disease in women, focusing on warning signs and symptoms. Historical documents available from the American Heart Association of Wisconsin and selected professional and secular literature were reviewed. Analysis revealed little information specific to women's symptoms until the 1980s. During the 1960s, articles and programs focused on how women could take care of their husbands' hearts. The 1980s brought a dramatic surge in literature on women's health, specifically on women's risk for heart disease. The evolution of information about women has mirrored the evolution of women's social roles. Despite the growing body of literature, more information and educational programs are needed to increase public awareness of cardiac warning signs and symptoms in women. PMID- 12122390 TI - Validity and reliability of the NYHA classes for measuring research outcomes in patients with cardiac disease. AB - The New York Heart Association (NYHA) functional classification system was developed to help physicians in clinical practice evaluate the effect of cardiac symptoms on a patient's daily activities. Over time, the role of the NYHA classification system has expanded, and it is now frequently used in clinical research. This review of the literature was undertaken to explore whether the NYHA classes have sufficient validity and reliability to serve as a functional outcome measure in research studies. After exploring its strengths and limitations, we conclude that the NYHA classes are a valid measure of functional status, a concept that is distinct from functional capacity and functional performance. The reproducibility of the NYHA functional classification system has not been established in the literature. Researchers are urged to report the methods for determining NYHA class, the training of raters, and the intra-rater or inter-rater reliability in studies that have multiple raters or measurements. Until the reliability of the NYHA functional classification system is determined, it is prudent to refrain from using the NYHA classes as the sole outcome measure of change in function in research studies of cardiac patients. PMID- 12122392 TI - The effects of prolonged strenuous exercise on left ventricular function: a brief review. PMID- 12122391 TI - Pulmonary veins: preferred site for catheter ablation of atrial fibrillation. AB - Atrial fibrillation (AF) is one of the most challenging arrhythmias to treat. Many patients have to accept this disorder and the medications required. Nonpharmacologic therapies have emerged as alternative methods of treatment. However, technical difficulty, low success rate, high recurrence, and complications still are obstacles. Pulmonary veins as the most common trigger foci of paroxysmal AF are now the most interesting source of curative ablation. With more knowledge, technologies, techniques, and equipment, AF ablation is likely to be more successful. This article introduces some exciting aspects of pulmonary vein ablation, including our hope to cure AF in some selected patients. PMID- 12122393 TI - Illness representations and coping of women with chronic obstructive pulmonary disease: a pilot study. AB - OBJECTIVE: The purpose of this study was to explore how women recognize and respond to symptoms of chronic obstructive pulmonary disease (COPD). FRAMEWORK: The Common Sense Model by Leventhal provided the theoretic framework for the study. DESIGN: A descriptive qualitative design was used. SAMPLE: The study sample included 21 women enrolled in pulmonary rehabilitation programs. FINDINGS: The causes and consequences of COPD were these women's most prominent representations. They struggled not only with dyspnea and fatigue, but with depression, loss of social support and intimacy, and stigma. Breathing techniques, medication, rest, and avoidance measures were the most frequently used coping strategies. PRACTICE IMPLICATIONS: Because dyspnea is the central symptom, nurses must continue to teach methods to control it, as well as to search for new interventions to relieve the symptoms. Optimal levels of rest and the effect on outcomes deserve close attention. Devising ways to deal with social and psychologic isolation would also enhance coping with COPD. PMID- 12122396 TI - The extent, nature, and precipitating factors of nurse assault among three groups of registered nurses in the regional medical center. PMID- 12122394 TI - Use of a pain assessment and intervention notation (P.A.I.N.) tool in critical care nursing practice: nurses' evaluations. AB - BACKGROUND: One of the barriers to effective pain management in critical care is the lack of systematic, comprehensive methods for assessing and treating pain. Use of a printed, standardized pain assessment and intervention tool can stimulate critical thinking and provide a framework for organizing pain assessment and management data. OBJECTIVES: The objectives of this study were to do the following: (1) describe the Pain Assessment and Intervention Notation (P.A.I.N.) tool, (2) detail critical care nurse participants' evaluations of the P.A.I.N. intervention tool when used during care of postoperative patients in pain, and (3) evaluate the tool's usefulness in practice and education. METHODS: Eleven intensive care unit (n = 7) and postanesthesia care unit (n = 4) nurses completed a questionnaire after they had used the pain tool in their clinical practices with 31 postoperative patients. RESULTS: Ten of the 11 nurses who returned an evaluation questionnaire found that the P.A.I.N. tool provided a consistent, systematic method of quantifying their assessment of patient pain and analgesic responsiveness. Five nurse participants believed that the P.A.I.N. tool improved their practice with regard to pain and sedation assessment. Three of the 11 nurses believed that the usefulness of the tool was limited because it was too detailed to be used routinely when caring for critically ill patients. All but 1 of the 11 nurses believed that the tool would have helped them earlier in their practice (ie, when they had less critical care nursing experience). CONCLUSIONS: The assessment and treatment of pain in critically ill patients are highly complex processes. This study identified many advantages of the use of a standardized, systematic approach to pain assessment and treatment by health professionals. PMID- 12122398 TI - Rhabdomyolysis: a late complication of near-drowning. PMID- 12122399 TI - Incidence and nature of farm-related injuries among Pennsylvania Amish children: implications for education. AB - INTRODUCTION: Farming is a dangerous occupation and results in injuries not only to farm workers, but also to families living on farms. Children raised on a farm are at particular risk because the farm is a place where they live, play, and work. Farming is the main Amish occupation, and because Amish farming techniques differ from those used on other us farms, it is important that health care providers recognize the causes of injuries that may occur on Amish farms. However, little is documented about farm-related injuries in Amish children, so we decided to document the incidence and nature of these injuries in Pennsylvania. METHODS: This study was done by reviewing the die botschaft, a weekly newspaper written by the Amish. All references to farm-related injuries among children in Pennsylvania were recorded. RESULTS: During a 5-month period, a total of 89 injuries, including 5 fatalities, were found: male children sustained 64 injuries, and female children sustained 25 injuries. Falls were the most commonly reported mechanism of injury, followed by incidents involving livestock. Both types of events most often resulted in an orthopedic injury. DISCUSSION: Injury to children in the Amish community is a significant problem that needs to be addressed and evaluated by the Amish themselves and in coordination with other health care providers, including emergency nurses. With the information obtained in this study, injury prevention strategies tailored to the Amish way of farming could be developed and implemented. PMID- 12122400 TI - Safety, tolerability, and efficacy of iontophoresis with lidocaine for dermal anesthesia in ED pediatric patients. AB - INTRODUCTION: This study examined the safety, tolerability, and efficacy of iontophoresis with 30 mA of lidocaine for dermal anesthesia in children younger than 84 months in the emergency department and the usefulness of a modified version of the Pre-verbal, Early Verbal Pediatric Pain Scale (M-PEPPS). METHODS: Three expert nurses completed the protocol for iontophoresis and inserted an intravenous catheter. Parents scored pain by using the 10-cm visual analogue scale, nurses used the M-PEPPS, and children, if able, self-reported pain during the procedure and at needle stick. RESULTS: Serum lidocaine levels were within the normal laboratory reference range. Adverse effects were minor and disappeared prior to discharge from the emergency department. Eighty-five percent of the children had M-PEPPS scores < or=6 during the iontophoresis procedure; 42% had scores of < or =6 at needle stick. Eighty-two percent of the parents marked the vas as < or =30 during the procedure; 65% indicated scores of < or =30 at needle stick. Four children self-reported "a lot of pain" at needle stick. Although low to moderate, M-PEPPS scores and parental pain ratings were significantly correlated at both points in time. DISCUSSION: Iontophoresis with lidocaine is safe for use in young children. It does not create any long-term untoward effects and is quite well tolerated. It is not clear if the higher pain scores at needle stick reflect anxiety and fear of a needle or a painful experience. PMID- 12122401 TI - Treating sickle cell pain: an update from the Georgia comprehensive sickle cell center. PMID- 12122403 TI - Knowledge assessment and preparation for the certified emergency nurses examination. PMID- 12122402 TI - Facilitating treatment in patients with stroke or chest pain through a dedicated ED chest pain/stroke unit. PMID- 12122404 TI - Behavioral and medical-surgical restraints: new protocols, forms, and competencies that comply with all regulatory mandates. PMID- 12122405 TI - Reducing use of restraints in the emergency department: one level III community hospital's experience. PMID- 12122406 TI - An informal discussion of emergency nurses' current clinical practice: what's new and what works. PMID- 12122407 TI - Transporting the morbidly obese patient: Framing an EMS challenge. PMID- 12122409 TI - Safe driving and transportation decisions and initiatives that can help ED nurses educate older Americans. PMID- 12122410 TI - The legal use of restraints. PMID- 12122412 TI - Critical care web sites. PMID- 12122414 TI - A cluster of children with lyme meningitis presenting to one emergency department in a Boston suburb. PMID- 12122413 TI - Methods of digital block. PMID- 12122415 TI - Successful management: reflections of one experienced nurse manager. PMID- 12122416 TI - Should only nurses do nursing research? AB - ( RESEARCH: careful investigation or study especially of a scholarly or scientific nature [from French, recercher, "to search closely"])In this column, Jane Koziol-McLain, PhD, RN, CEN, formerly a postdoctoral fellow at Johns Hopkins University School of Nursing and section editor of this Journal's RESEARCH Column, and currently Associate Professor, School of Nursing and Midwifery, and co-director of the Interdisciplinary Trauma RESEARCH Unit, Auckland University of Technology, Auckland, New Zealand (E-mail: Jane.Koziol-McLain@aut.ac.nz), posed a question to 2 nursing leaders: "Should only nurses do nursing research?" PMID- 12122417 TI - The trauma audit group (TAG): Oregon's 13-year experience. PMID- 12122418 TI - An unconscious diaphoretic 46-year-old woman. PMID- 12122419 TI - Never say never. PMID- 12122422 TI - [Stress and psychosis]. AB - An important component of vulnerability and environmental factors, specially stress, are essential for the production of psychoses, in particular schizophrenia. By the other hand, stress impairs clinical features in psychotic and prepsychotic patients. This happens principally because of the activation of Hyppotalamus - Pituitary - Adrenal axis and mesolimbic dopaminergic pathway. Preventive and therapeutical stress management may improve the evolution of psychosis. PMID- 12122421 TI - [Pharmacologic treatment of the psychotic symptoms in schizophrenia]. AB - Psychotic symptoms, among others, are present in Schizophrenia. The most severe developments during the moments of exacerbation, are meliorated after this period, although they do not absolutely disappear in the majority of the cases. Anti psychotic drugs are key in the treatment. As far as it is known, the efficacy of these drugs is due to the dopaminergic blockade. Nevertheless, the rythm, and the universal validity of this treatment actually have an heterogeneous character. At this moment, various medication algorithms are available, which are usefull in the orientation of the medical prescription. The comparative analysis and the practical conclusions are necessary for a correct management of them. PMID- 12122423 TI - [Karl Leonhard's Cycloid Psychosis and its position in the field of the endogenous psychosis]. AB - In this article, the author reviews the description of the cycloid psychosis out of the researches of the German psychiatric Karl Leonhard, placing them inside the wide field of the endogenous psychosis. PMID- 12122424 TI - [The psychotic states in the psychiatric emergency]. AB - The acute psychotic states are often seen in the Psychiatric Services. The syndromes associated with them are: agitation, attempt to commit suicide, aggressivity-violence -all of them make the psychotic states an emergency. The main syndrome to consider the psychotic state as an emergency is the agitation. This article reviews the literature on the pharmacotherapy of the psychotic states in emergency, such as the more used combinations of pharmacotherapies, the dosages, the way and frequency of administration. Likewise, it updates the available data about the uses of atypical antipsychotics in the emergency, such as risperidone syrup, and IM olanzapine and ziprasidone. PMID- 12122425 TI - [Psychosis and bipolarity]. AB - Since Kraepelin s pioneering differentiation, schizophrenia and manic-depressive psychosis have been considered distinct diseases. Research findings from different sources have found several similarities between these two disorders, casting doubts on Kraepelin s distinction. The current concept of spectrum seems to be more adequate to explain the relations between them. At the same time, a group of medicines, atypical antipsychotics, have shown interesting properties for the treatment of both affective and psychotic symptomatology. PMID- 12122426 TI - [Which collateral effects influence in the adherence with antipsychotic medication?]. AB - The collateral effects of antipsychotic medication, specially the mental aspects of extrapyramidal symptoms, sexual disorders and weight gain, constitute an important cause of lack of adherence to treatment. The common characteristic to all these effects is that they produce a high subjective distress. This is more related to the capacity to generate suffering to the patient than to the objective severity of a collateral effect. In this paper these collateral effects are revised. The conclusion is that, in spite of the fact that new antipsychotics show an improvement with regard to their collateral effects profile, these drugs include a new profile of distressing effects that must be evaluated and corrected, as a way to optimize the patient's adherence to antipsychotic treatment. PMID- 12122427 TI - [Psychotic depression]. AB - This paper will bring a brief overview of the clinical aspects of psychotic depression, with emphasis in the differentiation between mood-congruent and mood incongruent delusions. Biological aspects of this disorder will be reviewed. Changes have been found in serotonergic and dopaminergic neurotransmission and also a characteristic shortening of REM latency. Alterations of hypothalamic pituitary-adrenal (HPA) axis are especially prominent and may have specific features in this disorder. Attention will be given to structural brain alterations, cognitive impairments and the relationships of both with the HPA alterations previously described. Finally, a treatment update will be provided. Combination of antidepressants and antipsychotics is still the preferred psychopharmacologic treatment. There is recent research in selective serotonin reuptake inhibitors as monotherapy, atypical antipsychotics and antiglucocorticoid strategies but more investigations are needed. PMID- 12122429 TI - The Opisthorchis felineus paramyosin: cDNA sequence and characterization of its recombinant fragment. AB - The cDNA sequence of Opisthorchis felineus paramyosin (PM) was determined and shown to have 66-70% homology with two schistosomes and two cestodes. Phylogenetic analysis revealed an almost equal distance between O. felineus and these distinct clades. Because of its relatively low conservation, the PM gene may be a convenient genetic marker for studying phylogenetic relationships among platyhelminthes.A 25-kDa recombinant polypeptide corresponding to the central part of the full-length PM was produced. In Western blot analysis, murine hyperimmune serum against recombinant PM (recPM) detected 100-kDa polypeptides in the O. felineusegg and somatic antigens. Interactions of recPM with polyclonal anti-parasite antibodies and anti-recPM sera in ELISA with native antigens demonstrated that recPM carries a B cell epitope identical to the O. felineusnative antigen. Our sequence and immunologic data may be helpful in developing new diagnostic tools and candidate vaccines for O. felineus infection. PMID- 12122428 TI - Maternal malaria: Plasmodium falciparum sequestration in the placenta. AB - The human malarial parasite Plasmodium falciparumis responsible for an estimated 300-500 million clinical cases and 1-3 million deaths annually. At particular risk of developing severe, life-threatening malaria-associated complications are women during their first pregnancy. The observed pathologies, such as premature delivery, intrauterine growth retardation, abortion, and death of the mother and the newborn, are in large parts due to the parasite's ability to render infected erythrocytes adhesive and sequester in the intervillous space of infected placentas. In subsequent pregnancies, women are protected from maternal malaria through antibodies that prevent cytoadhesion of P. falciparum-infected erythrocytes in the placenta. Here, we summarize our current knowledge of the pathophysiological processes underpinning maternal malaria and discuss emerging concepts for intervention. PMID- 12122430 TI - An improved method for whole-mount in situ hybridization of Heterodera schachtii juveniles. AB - An optimized protocol is presented to visualize gene expression in the sedentary beet cyst nematode, Heterodera schachtii, by whole-mount in situ hybridization. Two different probes were used for genes with known expression pattern in other nematodes. Vacuum infiltration of the fixative significantly increased its efficiency and resulted in a nicely preserved morphology. Additional modifications were introduced to simplify and standardize the process. PMID- 12122431 TI - Parasite populations and communities from the shallow littoral of the Orther Bight (Fehmarn, SW Baltic Sea). AB - Parasites of host guilds, such as mud snails (five species), benthic crustaceans (six species) and small-sized fishes ( Pomatoschistus microps, Gasterosteus aculeatus, Pungitius pungitius, young Pleuronectes flesus), were investigated in the Other Bight off West Fehmarn (Kiel Bight, German Baltic Sea). The hosts, especially the herbivorous Hydrobia spp, Gammarus spp and Idothea chelipes, attained extremely high densities in three habitats ( Enteromorpha belt, Fucus belt, sandy bottom), which may be a consequence of high eutrophication. Fish as carnivores and several helminths as parasites can profit from these massive appearances - more than 5,000 Hydrobia mud snails/m(2) or 282 I. chelipes/m(2). Prevalences of mud snails peaked in summer, by up to 30% extra, whereas species of benthic crustaceans attained increases of 47-100%, fish 57-100%. The most abundant helminths were the digeneans Maritrema subdolum, Microphallus claviformis and Asymphylodora demeli in Hydrobia spp, Maritrema subdolum, Microphallus papillorobustum, Levinseniella brachysoma and Podocotyle atomon in benthic crustaceans and Cryptocotyle concavum, Podocotyle atomon and Brachyphallus crenatus in fish. The copepod Thersitina gasterostei was also abundant in sticklebacks. The density of parasites (number/m(2)) peaked in summer, with more than 10,000 Maritrema subdolum metacercariae in I. chelipes from the Fucus belt and more than 1,000 in I. chelipes from the Enteromorpha belt or the sandy bottom. There was a clear seasonality in the appearance of digeneans in G. locusta, G. salinus and I. chelipes infected by M. subdolum and Gammarus spp infected by L. brachysoma and P. atomon. Therefore, the Orther Bight may be a epizootiotope for M. subdolum and C. concavum, i.e. a habitat with an extreme infection rate which can endanger the population of the host. Idothea chelipes also has a high infection potential to the final hosts of M. subdolum, crustacean feeding birds. Similar relationships were found between the bird digenean C. concavum and the common goby Pomatoschistus microps. The infection mechanisms of both benthic crustaceans and small-sized fish follow the rule of predation in two steps: dispersion and accumulation. PMID- 12122432 TI - Agfa morandi sp. n. (Rhabditida, Agfidae) a parasite of Limax sp. (Gastropoda, Limacidae). AB - Agfa morandi sp. n. (Rhabditida, Agfidae), a parasite of Limax sp. (Gastropoda, Limacidae) from Py (Pyrenean mountains, France), is described and illustrated. The present species can be separated from the other two members of the same genus, A. flexilis (Rudolphi, 1819) Morand, 1990 and A. tauricus Korol and Spiridonov, 1991, by size measurements, number and disposition of the male's genital papillae, shape of the spicule and number of eggs in the female. PMID- 12122433 TI - Antiproliferative and leishmanicidal effect of ajoene on various Leishmania species: ultrastructural study. AB - Ajoene [(E,Z)-4,5,9 trithiadodeca 1,6,11 triene 9-oxide], the major bioactive compound derived from garlic, shows a potent trypanolytic and antimicotic activity. In this paper we evaluate its effect on Leishmania mexicana(Lm:MHOM/VE/80/NR), L eishmania mexicana venezuelensis (Lmv: MHOM/VE/80/H16), L eishmania mexicana amazonensis (Lma: M112, IFLA/BR/67/PH8) and L eishmania donovani chagasi (Ldch: MHOM/BR/74/PP75). Ajoene showed a potent leishmanicidal activity in vitro against all species studied. Concentrations higher than 0.3 microM led to total inhibition of growth, and 10 microM induced 100% lysis of Leishmaniaafter 96 h of incubation in a chemically defined culture medium. The 50% inhibitory concentration (IC(50)) for lysis, for all species, was about to 2 microM. The effect was dose-dependent and a threefold increase in concentration (30 microM) produced 100% lysis of cultured forms after 72 h. Ultrastructural studies showed a time- and dose-dependent morphological alteration of the mitochondrial membrane and nuclear envelope, as well as the formation of large autophagic vacuoles. PMID- 12122434 TI - Resistance to arsenite modulates expression of beta- and gamma-tubulin and sensitivity to paclitaxel during differentiation of Leishmania donovani. AB - Differentiation of Leishmania donovani promastigotes into infectious amastigotes is accompanied by differential tubulin gene expression. Tubulin is one of the proposed targets of clinically useful antileishmanial agents and its expression is known to alter due to drug resistance. In this study, beta- and gamma-tubulin expression under various stages of differentiation was measured in an in vitro generated arsenite-resistant L. donovani strain. Results showed higher constitutive expression of beta-tubulin in the arsenite-resistant promastigotes and amastigotes compared with the wild-type. beta-Tubulin expression in the resistant promastigotes increased on paclitaxel treatment. Significant differences in gamma-tubulin expression were observed only between the amastigotes, but not between promastigotes, of wild-type and resistant strains. Paclitaxel did not produce any significant change in the expression profile of gamma-tubulin in either of the strains, neither before nor after differentiation. Data suggest that the beta- and gamma-tubulin expression and the response to paclitaxel is affected due to arsenite resistance. PMID- 12122435 TI - DNA polymerase beta mRNA determination by relative quantitative RT-PCR from Leishmania infantum intracellular amastigotes. AB - Gene expression level is extremely difficult to assess in the intracellular form of Leishmania infantum, the amastigote, due to host mRNA contamination, low supply of parasites and stress degradation problems. We obtained and analyzed L. infantum DNA polymerase beta (Li Pol beta), a suitable enzyme for differential expression studies. The amount of Li Pol beta mRNA was determined in different forms of the parasite (extracellular promastigote, intracellular amastigote) by Northern blot and by relative quantitative reverse transcriptase-polymerase chain reaction (RT-PCR). Amastigote transcript levels were determined from both gradient-purified parasites and directly from a population of infected macrophages. The mRNA, undetectable when obtained from amastigotes by the classic gradient centrifugation method and subsequent Northern analysis, was clearly and specifically detectable by this quantitative RT-PCR method from the mixed macrophage/parasite cell population. Li Pol beta displays a different pattern of expression in the distinct forms of the parasite cycle that correlate with the infectivity of the protozoon. Thus, Li Pol beta mRNA expression is developmentally regulated, being clearly higher in the more infective forms of the parasite. PMID- 12122436 TI - Effect of a short period of desiccation during the patent period on cercarial shedding of Schistosoma haematobium from Planorbarius metidjensis. AB - Planorbarius metidjensis infected with Schistosoma haematobium were subjected, from the first day of shedding, to a 5- or 10-day period of desiccation in order to study its effect on the cercarial release occurring after reactivation of snails in water. These results were compared to those of infected controls that were not subjected to desiccation. Compared to controls, the duration of shedding following reactivation was shorter and the total number of cercariae shed by each infected snail was smaller in the 5- and 10-day groups. The rhythm occurring in the daily cercarial production of the 5-day group was shorter (11 days only) at the beginning of the shedding period and lengthened to 17-18 days at the end of the patent period. In the 10-day group, no rhythm was found. These results could be explained by the length of the desiccation period and its consequences on the development of S. haematobiumsporocysts in the body of snails. PMID- 12122437 TI - Ribosomal and mitochondrial DNA sequence variation in Sarcoptes mites from different hosts and geographical regions. AB - In order to investigate the extent of the genetic variation in the DNA sequences of Sarcoptes scabiei, mite populations collected on Alpine chamois ( Rupicapra rupicapra), Pyrenean chamois ( Rupicapra pyrenaica) and red fox ( Vulpes vulpes) from different localities of Italy and Spain were studied. Sequence analyses were carried out on the second internal transcribed spacer of the nuclear ribosomal DNA and on the 16S mitochondrial rRNA gene. ITS-2 sequences showed a higher degree of genetic polymorphism, mostly randomly distributed in the isolates from different hosts and localities, although both genomic regions are characterised by fixed nucleotide substitutions which were able to discriminate the mites collected on Pyrenean chamois from north-western Spain and on foxes from north eastern Spain and from north-western Italy with respect to the other isolates. These results suggest the existence of a limitation to free gene exchange between the studied populations, probably related to the genetic structuring of local populations rather than to a differential adaptation to host species. PMID- 12122438 TI - Karyotype of Acanthocephalus lucii: the first record of supernumerary chromosomes in thorny-headed worms. AB - The karyotype of Acanthocephalus lucii(Muller, 1776), a parasite of perch, consists of six autosomal biarmed chromosomes and two subtelocentric sex X chromosomes in females or one X in males. The sex determining mechanism is of the XX-X0 type. Most of the acanthocephalans (85%) exhibit small supernumerary metacentric B chromosomes, the number of which varies from one to five. Our data represent the first record of B chromosomes in thorny-headed worms. A possible relationship between the occurrence of B chromosomes and a heavy metal accumulation in acanthocephalan tissue from contaminated aquatic environments is briefly discussed. PMID- 12122439 TI - Existence of phenol oxidase in the argasid tick Ornithodoros moubata. AB - Phenol oxidase (PO, EC 1.10.3.1) activity was detected in the hemolymph of the fourth instar nymphs of the argasid tick, Ornithodoros moubata, with peak levels corresponding to the days before the majority of the nymphs had molted, suggestive of a protective role of PO during the ecdysial phase. Higher PO activity was detected in plasma relative to the hemolymph and was negligible in hemocytes. The concentration of the hemolymph and plasma assayed clearly influenced the level of PO activity, and was significantly reduced ( P<0.005) after treatment with 1-phenyl-2 thiourea, a specific PO inhibitor. This is the first report of the existence of PO in the hemolymph and plasma of a soft tick species. The regulation of PO activity and its precise role in soft tick immunity, particularly during the ecdysial phase, are interesting and need to be examined further. PMID- 12122440 TI - In vitro effects of gliotoxin, a natural proteasome inhibitor, on the infectivity and proteolytic activity of Toxoplasma gondii. AB - We examined the in vitro effect of increasing gliotoxin concentrations on the infectivity of Toxoplasma gondii for NIH-3T3 murine fibroblasts and on Toxoplasma chymotrypsin-like activity, which is specific to the proteasome. Parasite penetration of host cells was not modified by a high gliotoxin concentration (1 microM), but replication was markedly decreased (approximately 50% inhibition by 0.5 microM gliotoxin). Gliotoxin reduced the chymotrypsin-like activity of the Toxoplasma proteasome, but five times less potently than in HeLa cells. PMID- 12122441 TI - The endocytic pathway followed by the keratinocyte growth factor receptor. AB - Keratinocyte growth factor (KGF/FGF7) acts specifically on epithelial cells and regulates their proliferation and differentiation. It binds to and activates a receptor tyrosine kinase, the KGF receptor (KGFR), which is a splicing variant of the fibroblast growth factor receptor 2. The endocytic pathway followed by KGF and its receptor was analyzed here using immunofluorescence and confocal microscopy. After 10 min of internalization at 37 degrees C, both KGF and its receptor were localized in early endosomes, and after 30-60 min of endocytosis ligand and receptor were seen to reach perinuclear late endosomes and not the recycling endosomal compartment. Parallel western blot analysis revealed that KGFRs were tyrosine phosphorylated both at early and late steps of internalization, suggesting that KGF and KGFR remain associated in active complexes through the endocytic pathway. Pulse-chase experiments showed that the internalized KGFRs underwent degradation detectable at 1 h of endocytosis at 37 degrees C, indicating that KGFRs are functionally downregulated. PMID- 12122442 TI - Real-time RT-PCR quantitation of mCLCA1 and mCLCA2 reveals differentially regulated expression in pre- and postnatal murine tissues. AB - Members of the recently discovered chloride channels, calcium-activated (CLCA) gene family are thought to contribute to transmembrane trafficking of anions and other cellular functions. Previous northern blot and in situ hybridization studies revealed expression of the murine putative chloride channel mCLCA1 (alias mCaCC) in numerous epithelia and few other cell types. However, the subsequent cloning of mCLCA2 which shares 96% cDNA sequence identity with mCLCA1 suggested that the distribution pattern proposed for mCLCA1 in fact represented the sum of both mRNA species. In this study, a real-time RT-quantitative PCR assay was established to specifically quantify mCLCA1 and mCLCA2 expression in 19 pre- and 44 postnatal murine tissues. Different expression levels of mCLCA1 and mCLCA2 were found in most tissues analyzed. Particularly strong and virtually exclusive expression was found for mCLCA1 in spleen and bone marrow and for mCLCA2 in lactating and involuting mammary glands. In contrast, other tissues including intestine and trachea were found to express equally moderate levels of both homologues. Moreover, mCLCA2, but not mCLCA1, seems to be involved in stage specific organogenesis in fetal tissues. These results indicate that, in spite of their extremely close sequence homology, mCLCA1 and mCLCA2 are involved in different, yet unidentified pathways. PMID- 12122443 TI - Differentiation of original and regenerated skeletal muscle fibres in mdx dystrophic muscles. AB - The differentiation of both original muscle fibres and the regenerated muscle fibres following necrosis in mdx muscles was investigated using immunoblotting and immunocytochemical procedures. Before the onset of necrosis, postnatal skeletal muscles in mdx mouse differentiated well with only a slight delay in differentiation indicated by the level of developmental isoforms of troponin T. Prior to the onset of apparent myopathic change, both fast and slow skeletal muscle fibre types in mdx leg muscles also differentiated well when investigated by analysis of specific myosin heavy chain expression pattern. While the original muscle fibres in mdx leg muscles developed well, the differentiation of regenerated myotubes into both slow and distinct fast muscle fibre types, however, was markedly delayed or inhibited as indicated by several clusters of homogeneously staining fibres even at 14 weeks of age. The number of slow myosin heavy chain-positive myotubes amongst the regenerated muscle clusters was quite small even in soleus. This study thus established that while muscle fibres initially develop normally with only a slight delay in the differentiation process, the differentiation of regenerated myotubes in mdx muscles is markedly compromised and consequently delayed. PMID- 12122444 TI - Eccentric contractions leading to DOMS do not cause loss of desmin nor fibre necrosis in human muscle. AB - High force eccentric muscle contractions can result in delayed onset muscle soreness (DOMS), prolonged loss of muscle strength, decreased range of motion, muscle swelling and an increase of muscle proteins in the blood. At the ultrastructural level Z-line streaming and myofibrillar disruptions have been taken as evidence for muscle damage. In animal models of eccentric exercise induced injury, disruption of the cytoskeleton and the sarcolemma of muscle fibres occurs within the first hour after the exercise, since a rapid loss of staining of desmin, a cytoskeletal protein, and the presence of fibronectin, a plasma and extracellular protein, are observed within the muscle fibres. In the present study, biopsies from subjects who had performed different eccentric exercises and had developed DOMS were examined. Our aim was to determine whether eccentric exercise leading to DOMS causes sarcolemmal disruption and loss of desmin in humans. Our study shows that even though the subjects had DOMS, muscle fibres had neither lost staining for desmin nor contained plasma fibronectin. This study therefore does not support previous conclusions that there is muscle fibre degeneration and necrosis in human skeletal muscle after eccentric exercise leading to DOMS. Our data are in agreement with the recent findings that there is no inflammatory response in skeletal muscle following eccentric exercise in humans. In combination, these findings should stimulate the search for other mechanisms explaining the functional and structural alterations in human skeletal muscle after eccentric exercise. PMID- 12122445 TI - Golgi-associated filament networks in duct epithelial cells of rabbit submandibular glands: immunohistochemical light and electron microscopic studies. AB - The duct epithelial cells of rabbit submandibular glands expressed keratin 8, keratin 14, and actin in the supranuclear region, and these cytoskeletal proteins formed ring structures, approximately 1-2.5 microm in diameter. Ultrastructurally, these ring structures were observed as a 'Golgi-associated filament network' surrounding Golgi apparatuses. Double immunofluorescent studies showed that keratin 8 and keratin 14 formed keratin 8/14 filaments, and that these filaments and actin filaments colocalized as components of the Golgi associated filament network. Our studies suggested that the Golgi-associated filament network maintains the complex structure and location of the Golgi apparatus of the duct epithelium of rabbit submandibular glands. Tubulin was distributed diffusely throughout the cytoplasm of columnar cells, but no special relationship was found between tubulin and the Golgi apparatus. PMID- 12122446 TI - Mast cells and macrophages in normal C57/BL/6 mice. AB - Mast cells and macrophages have an important role in immunity and inflammation. Because mice are used extensively for experimental studies investigating immunological and inflammatory responses, we examined mast cell and macrophage distribution in normal murine tissues. Mast cells were abundant in the murine dermis, tongue, and skeletal muscle but were rarely found in the heart, lung, spleen, kidney, liver, and the bowel mucosa. In contrast, dogs exhibited large numbers of mast cells in the lung parenchyma, liver, and bowel. Some murine dermal mast cells had long cytoplasmic projections filled with granular content. Mouse mast cells demonstrated intense histamine immunoreactivity and were identified with histochemical enzymatic techniques for tryptase and chymase. Macrophages, identified using the monoclonal antibody F4/80, were abundant in the spleen, lung, liver, kidney, and bowel but relatively rare in the heart, tongue, and dermis. Using a nuclease protection assay we investigated mRNA expression of stem cell factor (SCF), a crucial survival factor for mast cells, and the macrophage growth factors macrophage colony stimulating factor (M-CSF) and granulocyte macrophage colony stimulating factor (GM-CSF). Stem cell factor mRNA was highly expressed in the murine lung. Relatively low levels of SCF mRNA expression were found in the tongue and earlobe, which are tissues containing a high number of mast cells. Macrophage CSF and GM-CSF mRNA was highly expressed in the lung and spleen. The murine heart, an organ with a low macrophage content, expressed high levels of M-CSF but negligible levels of GM-CSF mRNA. Constitutive growth factor mRNA expression in murine tissues without significant populations of mast cells and macrophages may suggest an alternative role for these factors in tissue homeostasis. PMID- 12122448 TI - Delineation of the evolution of compositional changes in atheroma. AB - In a previous manuscript, we described a method combining immunohistochemistry, confocal scanning laser microscopy, and computer-assisted image analysis for the determination of the composition of atherosclerotic plaques [Taatjes et al. (2000) Histochemistry and Cell Biology 113:161-173). We now present an enhanced technique, and its use for age- and gender-related comparative analysis of lesion composition in ApoE knockout mice. Cryosections from aorta were stained with oil red O to detect lipid, SYTOX Green to detect cellularity, and Picrosirius red to delineate collagen fibers. The stained sections were imaged by brightfield light microscopy, epifluorescence microscopy, and polarized light microscopy. Digital images were collected, processed to isolate the lesions, and subjected to computer-assisted image analysis. The average percentage of the vessel wall occupied by lesion increased 1.5-fold in animals from 10 to 20 weeks. Although the amount of lipid in the lesions increased in animals from 10 to 20 weeks, the percentage composition in the lesion remained constant because of the increase in lesion size. Average cellularity showed a modest decrease over the same interval. However, the percentage composition of plaque attributable to collagen increased 2.5-fold in 20-week-old female animals compared with that in males or females of 10 weeks of age and males of 20 weeks of age. PMID- 12122447 TI - Expression of neurotrophins and their receptors in peripheral lung cells of mice. AB - Neurotrophins play an essential role in nerve systems. Recent reports indicated that neurotrophins [nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3), and neurotrophin-4/5 (NT-4/5)] have numerous effects on non-neural cells, especially on immune cells. However, whether lung cells express neurotrophins and/or their receptors (TrkA for NGF, TrkB for BDNF and NT-4/5, and TrkC for NT-3) has never been systematically investigated. We investigated constitutive expression of neurotrophin family and their Trk receptor family in alveolar macrophages and other peripheral lung cells of mice. New findings were: (1) RT-PCR for neurotrophins and their receptors detected NT-3 and NT-4/5 in alveolar macrophages, BDNF, NT-4/5, trkA, the truncated form of trkB, and trkC in lung homogenate, but no trks in alveolar macrophages, (2) immunohistochemistry for neurotrophin receptors detected TrkA in capillary cells, the truncated form of TrkB, and TrkC in interstitial macrophages, (3) immunoelectron microscopy for TrkC revealed expression of TrkC on the surface of interstitial macrophages, and (4) in situ hybridization for neurotrophins detected BDNF in interstitial macrophages and alveolar type I cells, NT-3 in alveolar macrophages, and NT-4/5 in alveolar and interstitial macrophages. These findings indicate that a previously unknown signal trafficking occurs through neurotrophins in peripheral lung. PMID- 12122449 TI - Differential expression of glycosaminoglycans and proteoglycans in the migratory pathway of the primordial germ cells of the mouse. AB - Primordial germ cells are an embryonic cell line that give rise to gametes in vertebrates. They originate outside the embryo proper and migrate by a well defined route to the genital ridges. Proteoglycans and glycosaminoglycans have distinctive properties that affect many of the characteristics of the extracellular microenvironment of migratory pathways in a variety of developmental systems. The purpose of this work was to identify the proteoglycans and glycosaminoglycans that are spatially and temporally expressed in the migratory pathway of primordial germ cells. We showed that the expression of proteoglycans and glycosaminoglycans in the primordial germ cells migratory pathway changes according to the different phases of the migratory process. Some molecules such as chondroitin-0-sulfate, decorin, and biglycan are present only in certain phases of the migratory process of primordial germ cells. Heparan sulfate, chondroitin-6-sulfate, versican, perlecan, and syndecan-4, although exhibiting some variation in expression were detected during all phases of the migratory process. Our results indicate that the successive steps of primordial germ cell migration require a coordinated expression of proteoglycans and glycosaminoglycans, that should be present in appropriate levels and in specific areas of the embryo, and that the sequential expression of these extracellular matrix molecules is under a genetic program that appears to be common to a variety of cell types during embryonic development. PMID- 12122450 TI - Detection of glycosylated sites in embryonic rabbit kidney by lectin chemistry. AB - The reciprocal cell biological interaction between mesenchymal and epithelial tissue plays a critical role during nephrogenesis. It is unknown to date whether the tissues interact during nephron induction by pure diffusion of substances or whether cellular contacts via gap junctions or focal adhesion molecules are involved. In neonatal rabbit kidney the interface between both tissues shows unique features. It consists of a distinct space, which is filled with specific extracellular matrix consisting of glycosylated proteins such as fibronectin, laminin, collagen, and proteoglycans. In the present experiments we tested by histochemistry whether it is possible to detect additional glycosylated proteins using Soybean agglutinin (SBA), Dolichos biflorus agglutinin (DBA), Ulex europaeus I agglutinin (UEA I), and Peanut agglutinin (PNA) as molecular markers. All tested lectins showed distinct labeling patterns in embryonic renal tissue. Within the collecting duct ampulla, DBA and UEA I revealed intensive cellular reaction. In contrast, PNA and SBA reacted at the basal aspect of the collecting duct ampulla tip in addition to a cellular reaction. To identify the individual molecules labeled by the lectins, embryonic tissue was fractionated and separated by electrophoretic methods. For the first time, we were able to show by two dimensional electrophoresis and subsequent western blot experiments that lectins bind to a series of individual protein spots, which have not been identified to date. PMID- 12122451 TI - Reversible depression of transcription during hibernation. AB - Mammalian hibernators downregulate processes of energy production and consumption while maintaining cellular homeostasis. Energetic costs of transcription must be balanced with demands for gene products. Data from nuclear run-on assays indicate transcriptional initiation is reduced two fold in torpid golden-mantled ground squirrels ( Spermophilus lateralis) as compared to euthermic animals between bouts of torpor. In addition, elongation rates across the temperature range experienced by hibernators indicate a virtual arrest of transcription at the low body temperatures of torpor. Finally, there is no seasonal compensation or species-specific adaptation for increased elongational capacity in the cold. Thus, it appears that hibernators are not specifically adapted to continue transcription during torpor. Taken together, these data indicate that transcription arrests during torpor because of a moderate depression of initiation and a more severe inhibition of elongation, largely due to temperature effects. Restoration of euthermic body temperatures during the interbout arousals reverses this transcriptional depression and permits gene expression. PMID- 12122452 TI - Cardiovascular effects of prostaglandin F(2 alpha) and prostaglandin E(2) in Atlantic cod (Gadus morhua). AB - Little is known of the cardiovascular functions of prostaglandins in non mammalian vertebrates. There are indications that prostaglandins may have a function in haemostasis by constricting blood vessels in filament arteries in the fish gill after injury. Our aim was to examine the cardiovascular effect of the prostaglandins F(2 alpha) (PGF(2 alpha)) and E(2) (PGE(2)) with emphasis on branchial circulation. Intra-arterial injections of PGF(2 alpha) (10, 40, 160, 400 nmol kg(-1)) in cod caused a dose-dependent increase in ventral aortic blood pressure, a reduction in cardiac output, and an increase in gill vascular resistance. A contraction of filament arteries was observed with in vivo microscopy only seconds after injection. PGF(2 alpha) may therefore possibly be involved in a haemostatic vasoconstriction. In contrast, the most significant effects of PGE(2) appeared to be on the heart. PGE(2) also reduced dorsal aortic blood pressure. PMID- 12122453 TI - Metabolic rate and thermal conductance of lemmings from high-arctic Canada and Siberia. AB - The arctic climate places high demands on the energy metabolism of its inhabitants. We measured resting (RMR) and basal metabolic rates (BMR), body temperatures, and dry and wet thermal conductances in summer morphs of the lemmings Dicrostonyx groenlandicus and Lemmus trimucronatus in arctic Canada, and the BMR of D. torquatus, D. groenlandicus, L. sibiricus, L. bungei and L. trimucronatus in Siberia. In contrast to previous studies the data were collected on animals that had spent only a limited time in captivity. All parameters were analysed in relation to the variations in body mass (20-90 g). Body temperature and BMR were lower in D. groenlandicus than L. trimucronatus, which coincides with greater longevity in the former species. Wet and dry thermal conductances of both species were similar and comparable with those of other Myomorpha (mouse type rodents), indicating no evidence for a previously claimed lower thermal conductance in lemmings. BMR in lemmings appeared to be higher than in other Arvicolidae (voles, lemmings and muskrats), which could relate to their typically high-latitude distribution. However, the more southerly living Lemmus species had higher BMR than the more northerly living Dicrostonyx species, which may be explained by the former having a relatively low-quality diet. PMID- 12122455 TI - Proton leak in hepatocytes and liver mitochondria from archosaurs (crocodiles) and allometric relationships for ectotherms. AB - It has previously been shown that mitochondrial proton conductance decreases with increasing body mass in mammals and is lower in a 250-g lizard than the laboratory rat. To examine whether mitochondrial proton conductance is extremely low in very large reptiles, hepatocytes and mitochondria were prepared from saltwater crocodiles ( Crocodylus porosus) and freshwater crocodiles ( Crocodylus johnstoni). Respiration rates of hepatocytes and liver mitochondria were measured at 37 degrees C and compared with values obtained for rat or previously measured for other species. Respiration rates of hepatocytes from either species of crocodile were similar to those reported for lizards and approximately one fifth of the rates measured using cells from mammals (rat and sheep). Ten-to-thirty percent of crocodile hepatocyte respiration was used to drive mitochondrial proton leak, similar to the proportion in other species. Respiration rates of crocodile liver mitochondria were similar to those of mammalian species. Proton leak rate in isolated liver mitochondria was measured as a function of membrane potential. Contrary to our prediction, the mitochondrial proton conductance of liver mitochondria from crocodiles was greater than that of liver mitochondria from lizards and was similar to that of rats. The acyl composition of liver mitochondrial phospholipids from the crocodiles was more similar to that in mitochondria from rats than in mitochondria from lizards. The relatively high mitochondrial proton conductance was associated with a relatively small liver, which seems to be characteristic of crocodilians. Comparison of data from a number of diverse ectothermic species suggested that hepatocyte respiration rate may decrease with body mass, with an allometric exponent of about -0.2, similar to the exponent in mammalian hepatocytes. However, unlike mammals, liver mitochondrial proton conductance in ectotherms showed no allometric relationship with body size. PMID- 12122454 TI - Effects of nutritional conditions around weaning on carbonic anhydrase activity in the parotid gland and ruminal and abomasal epithelia of Holstein calves. AB - Thirty-two male Holstein calves were used to investigate the effects of nutritional conditions around weaning and aging on carbonic anhydrase (CA) activity in the parotid gland and epithelium from the rumen and abomasum. We fed calf starter and lucerne hay as well as milk replacer (group N) or fed milk replacer either with (group S) or without (group M) administration of short-chain fatty acids (SCFA) through polypropylene tubing into the forestomach until 13 weeks of age. The diets were fed at 1000 hours and 1600 hours, and SCFA were administrated after milk replacer feeding at 1600 hours. Slaughter and tissue sampling were carried out between 1300 hours and 1430 hours at 1, 3, 7, 13, and 18 weeks of age. Tissue samples from five adult (1.5-2.0 years-old) Holstein steers were obtained from a local abattoir. In group N, CA activity in the parotid gland gradually and significantly increased toward the adult value, whilst in the epithelium from the rumen and abomasum, adult values were reached at 3 and 7 weeks of age, respectively. At 13 weeks, the activity for group N was significantly higher than that for the other two groups in the parotid gland, but there was no significant difference in the epithelium from the rumen and abomasum. The concentration of the carbonic isozyme VI in the parotid gland also changed with age but, in contrast to CA activity, had not reached adult levels by 13 weeks of age. In groups M and S, parotid saliva did not show any change toward an alkaline pH or toward a reciprocal change in the concentrations between Cl(-) and HCO(3)(-), even at 13 weeks of age. From these results we conclude that a concentrate-hay based diet around weaning has a crucial role in CA development in the parotid gland, but not in the epithelium of the rumen and abomasum. PMID- 12122456 TI - Effect of adrenergic antagonists during phenylephrine stimulation of the mandibular gland of red kangaroos, Macropus rufus. AB - Intracarotid infusions of l-phenylephrine at 1.0 nmol.kg(-1).min(-1) or(.)10 nmol.kg(-1).min(-1) were accompanied by increases in salivary protein, urea, magnesium and bicarbonate, and by decreases in osmolality, hydrogen ion activity, sodium, potassium and chloride relative to cholinergically stimulated saliva. Intravenous infusions of phenylephrine at the same dose rates had much less effect on salivary composition with the differences between the routes of administration being greatest for the higher dose rate. Propranolol administered with phenylephrine via the carotid artery, at an antagonist:agonist ratio of 10:1, was much more effective in blocking the phenylephrine-induced changes in salivary composition than equimolar infusion of phentolamine with phenylephrine. Simultaneous intracarotid infusions of either a beta(1)-antagonist (CGP20712A) or a beta(2)-antagonist (ICI118551) with phenylephrine showed that ICI118551 was more potent than CGP20712A at preventing the changes in salivary composition associated with phenylephrine administration. It was concluded that alpha(1) adrenoreceptors were not present in functionally significant numbers in the gland and that the effect of phenylephrine on the kangaroo mandibular was mediated by beta-adrenoreceptors predominantly of the beta(2)-subtype. As the phenylephrine dose rates in the kangaroos were comparable with those used to determine alpha adrenergic responses of eutherian salivary glands and as both propranolol and phentolamine appeared to have minor beta-sympathomimetic activity, at least one subtype of beta-adrenoreceptors in macropods may not be identical to its eutherian counterpart. PMID- 12122457 TI - Plasma and muscle phospholipids are involved in the metabolic response to long distance migration in a shorebird. AB - We studied: (1) concentrations and fatty acid compositions of plasma non esterified fatty acids, neutral lipids, and phospholipids, and (2) fatty acid composition of flight muscle phospholipids in wintering, premigratory, and spring and fall migrating western sandpipers ( Calidris mauri). Plasma neutral lipid and phospholipid levels were elevated in migrants, reflecting high rates of fat deposition. An important role of phospholipids in fattening is suggested by the fact that the amount of fatty acids in plasma phospholipids was similar to, or in spring as much as twice, that of neutral lipids. Changes in the ratio of plasma neutral lipids to phospholipids may indicate seasonal changes in triacylglycerol stores of invertebrate prey. Monounsaturation and total unsaturation of plasma neutral lipids and phospholipids increased during migration. Muscle phospholipids were more monounsaturated in spring and fall, but total unsaturation was reduced in fall. Arachidonic acid [20:4(n-6)] was especially abundant in muscle phospholipids in winter (29%) and declined during migration (19-22%), contributing to a decline in the ratio of n-6 to n-3 fatty acids. The abundance of plasma phospholipids and variability of neutral lipid to phospholipid ratio indicates that measurement of plasma phospholipids will improve methods for assessment of fattening rates of birds. The functional significance of changes in muscle phospholipids is unclear, but may relate to depletion of essential n-6 fatty acids during exercise. PMID- 12122458 TI - Intermittent fasting during winter and spring affects body composition and reproduction of a migratory duck. AB - We compared food intake, body mass and body composition of male and female black ducks (Anas rubripes) during winter (January-March). Birds were fed the same complete diet ad libitum on consecutive days each week without fasting (control; nine male; nine female) or with either short fasts (2 day x week(-1); nine male; nine female), or long fasts (4 day x week(-1); eleven male; twelve female). We continued treatments through spring (March-May) to measure the effect of intermittent fasts on body mass and egg production. Daily food intake of fasted birds was up to four times that of unfasted birds. Weekly food intake of males was similar among treatments (364 g x kg(-1) x week(-1)) but fasted females consumed more than unfasted females in January (363 g x kg(-1) x week(-1) vs. 225 g x kg(-1) x week(-1)). Although both sexes lost 10-14% body mass, fasted females lost less mass and lipid than unfasted females during winter. Total body nitrogen was conserved over winter in both sexes even though the heart and spleen lost mass while the reproductive tract and liver gained mass. Intermittent fasting increased liver, intestinal tissue and digesta mass of females but not of males. Fasting delayed egg production in spring but did not affect size, fertility or hatching of the clutch. Females on long fasts were still heavier than controls after laying eggs. Thus black ducks combine flexibility of food intake with plasticity of digestive tract, liver and adipose tissue when food supply is interrupted during winter. Females modulate body mass for survival and defer reproduction when food supply is interrupted in spring. PMID- 12122459 TI - Calcium ion triggers rapid morphological oscillation of chloride cells in the mudskipper, Periophthalmus modestus. AB - Euryhaline teleosts rapidly regulate their ion flux at the chloride cells on entry to different salinities. The external trigger(s) for the rapid opening and closing of the chloride cell apical surface, the site of salt secretion, were examined in the skin of the mudskipper. With DASPEI (a mitochondrial probe) and Concanavalin-A (an apical surface marker of chloride cells), chloride cells were classified into two groups: those in contact and those not in contact with the external water. The fraction of chloride cells in contact with the water increased dramatically 1.5 h after transfer to seawater, and similarly after transfer to 10 mmol x l(-1) CaCl(2) solution. In comparison, transfer to 1.1 mol x l(-1) mannitol, 0.5 mol x l(-1) NaCl or 50 mmol x l(-1) MgCl(2) resulted in increases of only 40-60% of those in the seawater-transfer group. After return of the fish from 10 mmol x l(-1) CaCl(2) to freshwater, the cells in contact with the water decreased. Environmental Ca(2+) is the trigger for the morphological oscillation of chloride cell apical surface, presumably through modifications in Ca(2+) flux. PMID- 12122460 TI - Effects of nitric oxide synthase inhibitors on the febrile response to muramyl dipeptide and lipopolysaccharide in rats. AB - We have administered aminoguanidine, a relatively specific inhibitor of inducible nitric oxide synthase, and N-nitro-L-arginine methyl ester (L-NAME), an unspecific nitric oxide synthase inhibitor, to rats made febrile with the gram positive pyrogen, muramyl dipeptide and gram-negative pyrogen, lipopolysaccharide. Sprague-Dawley rats, housed individually at approximately 25 degrees C with a 12:12 h light:dark cycle (lights on 0700 hours), were injected (at 0900 hours) intraperitoneally with 50 mg/kg aminoguanidine, 25 mg/kg or 50 mg/kg L-NAME, and intramuscularly with 500 microg/kg muramyl dipeptide or 100 microg/kg lipopolysaccharide. Pyrogen injections were spaced at least 14 days apart. Body temperature was measured throughout the study in unrestrained animals using radio-telemetry. Neither muramyl dipeptide nor lipopolysaccharide-induced fevers were affected by aminoguanidine. However, L-NAME administration inhibited muramyl dipeptide and lipopolysaccharide-induced fevers, but only for the 1st 2-4 h of the fevers (two-way ANOVA, P<0.05). After the initial inhibition, lipopolysaccharide fevers developed normally. Therefore, constitutively expressed nitric oxide synthase appears to be involved in the initial phases of fever genesis of gram-negative and gram-positive fevers in rats. On the other hand, inducible nitric oxide synthase appears not to play a role in these fevers. PMID- 12122461 TI - Differential display of skin mRNAs regulated under varying environmental conditions in a mudskipper. AB - To understand the molecular mechanisms underlying the terrestrial adaptation, as well as adaptation to different salinities, of the euryhaline and amphibious mudskipper ( Periophthalmus modestus), we have looked for the skin mRNAs that change during varying environmental conditions. Using differential mRNA display polymerase chain reaction, we compared skin mRNAs in mudskipper transferred from isotonic 30% seawater to fresh water or to seawater for 1 day and 7 days, as well as those kept out of water for 1 day. At the end of these periods, poly(A(+))RNA was prepared from the Cl(-)-secreting pectoral skins and also from the outer opercular skins where ion transport is negligible, and analyzed by differential display. We identified four cDNA products expressed differently under various environments as homologues of known genes. A further 34 cDNAs were expressed differentially, but they have no significant homology to identified sequences in GenBank. Northern blots demonstrate that mRNA levels of the actin-binding protein and the platelet-activating factor acetylhydrolase increased in the pectoral skins during seawater acclimation. The mRNA of the 90 kDa heat shock protein was down-regulated in water-deprived and freshwater fish, whose plasma cortisol levels were high. The aldolase mRNA was induced in both skins after desiccation. These four genes may be involved in the environmental adaptations. PMID- 12122462 TI - Neural networks in the cockpit of the fly. AB - Flies have been buzzing around on earth for over 300 million years. During this time they have radiated into more than 125,000 different species (Yeates and Wiegmann 1999), so that, by now, roughly every tenth described species is a fly. They thus represent one of the most successful animal groups on our planet. This evolutionary success might, at least in part, be a result of their acrobatic maneuverability, which enables them, for example, to chase mates at turning velocities of more than 3000 degrees s(-1) with delay times of less than 30 ms (Land and Collett 1974; Wagner 1986). It is this fantastic behavior, which has initiated much research during the last decades, both on its sensory control and the biophysical and aerodynamic principles of the flight output (Dickinson et al. 1999, 2000). Here, we review the current state of knowledge about the neural processing of visual motion, which represents one sensory component intimately involved in flight control. Other reviews on this topic have been published with a similar (Hausen 1981, 1984; Hausen and Egelhaaf 1989; Borst 1996) or different emphasis (Frye and Dickinson 2001; Borst and Dickinson 2002). Because of space limitations, we do not review the extensive work that has been done on fly motion sensitive neurons to advance our understanding of neural coding (Bialek et al. 1991; Rieke et al. 1997; de Ruyter et al. 1997, 2000; Haag and Borst 1997, 1998; Borst and Haag 2001). Unless stated otherwise, all data presented in the following were obtained on the blowfly Calliphora vicina which we will often casually refer to as 'the fly'. PMID- 12122463 TI - An early chapter in behavioral physiology and sociobiology: the science of Martin Lindauer. AB - The fields of behavioral physiology and sociobiology enjoyed spectacular success in post World War II Germany. One of the major contributors to this blossoming in behavioral science was Martin Lindauer, who furthered the research approach of his mentor (Karl von Frisch), made numerous seminal discoveries, and nurtured a strong next generation in the area of neurobiology and behavior. We review the scientific development of Martin Lindauer within the German academic system in the years surrounding World War II, examine his research approach and achievements, and discuss his unusually successful methods of scientific pedagogy. PMID- 12122464 TI - Anisotropic imaging in the dragonfly median ocellus: a matched filter for horizon detection. AB - It is suggested that the dragonfly median ocellus is specifically adapted to detect horizontally extended features rather than merely changes in overall intensity. Evidence is presented from the optics, tapetal reflections and retinal ultrastructure. The underfocused ocelli of adult insects are generally incapable of resolving images. However, in the dragonfly median ocellus the geometry of the lens indicates that some image detail is present at the retina in the vertical dimension. Details in the horizontal dimension are blurred by the strongly astigmatic lens. In the excised eye the image of a point source forms a horizontal streak at the level of the retina. Tapetal reflections from the intact eye show that the field of view is not circular as in most other insects but elliptical with the major axis horizontal, and that resolution in the vertical direction is better than in the horizontal. Measurements of tapetal reflections in locust ocelli confirm their visual fields are wide and circular and their optics strongly underfocused. The ultrastructure suggests adaptation for resolution, sensitivity and a high metabolic rate, with long, widely separated rhabdoms, retinulae cupped by reflecting pigment, abundant tracheoles and mitochondria, and convoluted, amplified retinula cell plasma membranes. PMID- 12122465 TI - Retreat site selection and social organization in captive electric fish, Apteronotus leptorhynchus. AB - Gymnotiform fish use their electric organ discharge for electrolocation and communication. They are active nocturnally and seek retreat sites during the day. We examined retreat site selection in Apteronotus leptorhynchus by assessing their preference for retreat tubes that differed in opacity and dimension. Isolated fish preferred opaque to clear tubes, long and narrow diameter tubes to short, wide diameter tubes, and open-ended to closed tubes. We also assessed how groups of fish distributed themselves in tubes according to sex and electric organ discharge frequency under four conditions: (1) unlimited tube availability, (2) limited tube availability, (3) variation in tube opacity, and (4) variation in tube dimension. When tube availability was unlimited, fish generally preferred to occupy tubes alone. However, females, but not males, often cohabited tubes with consexuals. When tube availability was limited, females were more often than males found outside of tubes. When tubes varied by opacity and dimension, fish clustered most commonly in preferred tube types (opaque and long tubes). Males with the highest electric organ discharge frequencies usually occupied the most preferred tube type. Thus, fish have clear preferences in selecting retreat sites and groups of fish reveal their dominance relationships when presented with variation in retreat sites. PMID- 12122466 TI - Brain serotonin and the control of food intake in rainbow trout (Oncorhynchus mykiss): effects of changes in plasma glucose levels. AB - The levels of 5-hydroxytryptamine and its main metabolite 5-hydroxyindoleacetic acid were assessed in two brain regions, hypothalamus and telencephalon, of rainbow trout (Oncorhynchus mykiss) submitted to increases or decreases in plasma glucose levels through different experimental approaches. Thus, intraperitoneal glucose treatment (500 mg kg(-1)) increased 5-hydroxytryptamine telencephalic levels. Long-term food deprivation up to 3 weeks significantly increased hypothalamic (2 weeks and 3 weeks) and telencephalic (1 week, 2 weeks, and 3 weeks) levels of 5-hydroxyindoleacetic acid, whereas the ratio 5 hydroxyindoleacetic acid/5-hydroxytryptamine significantly increased throughout the food-deprivation period assessed. Intraperitoneal treatment with bovine insulin (4 mg kg(-1)) decreased the 5-hydroxyindoleacetic acid/5 hydroxytryptamine ratio in hypothalamus after 1 h. Intraperitoneal administration of fenfluramine (3 mg kg(-1)) caused a depression in food intake coincident with a significant decrease of the hypothalamic 5-hydroxyindoleacetic acid/5 hydroxytryptamine ratio. These data are discussed in the context of the involvement of serotonergic system in the control of food intake in rainbow trout. PMID- 12122467 TI - Different functions of two alarm substances in the honeybee. AB - In the honeybee, isopentyl acetate and 2-heptanone are described as alarm substances. We asked whether both substances have a similar role by testing the effect of their exposure on the appetitive proboscis extension reflex and on the aversive stinging reflex. In the appetitive context of sucrose stimulation no differences were found between isopentyl acetate and 2-heptanone. Small amounts of isopentyl acetate or 2-heptanone (3 microl of 1:9 dilution) yielded a response similar to that of a non-exposed control. Larger amounts of both substances (125 microl of 1:9 dilutions) as well as mixtures led to a decrease of responsiveness to sucrose. In the aversive context of electrical stimulation, significant differences between isopentyl acetate and 2-heptanone were found. Exposure to a small amount of isopentyl acetate (3 microl of 1:9 dilution) or to a large amount of 2-heptanone (125 microl of 1:9 dilution) led to an increase of responsiveness to the electric shock. Larger quantities of isopentyl acetate (125 microl of 1:9 dilution) decreased the responsiveness to the shock. 2-Heptanone never decreased the responsiveness to the shock. Our results indicate that isopentyl acetate and 2-heptanone have different functions even if both are capable of evoking deterrent responses in a defensive context. PMID- 12122468 TI - Evidence for a two pigment visual system in the fiddler crab, Uca thayeri. AB - Intraocular recordings were made from the eyestalks of dark-adapted fiddler crabs (Uca thayeri) during presentation of monochromatic light flashes of different wavelengths and intensities. Two types of signals were recorded in different experiments: slow potentials (electroretinogram) and fast potentials (spikes). The latter were also recorded in the presence of a continuous green or red adapting light. The resulting visual spectral-sensitivity curves, when fitted to rhodopsin-based visual pigment absorption spectra (from Dartnall nomograms), indicated the presence of two visual pigments, one with an absorption maximum near 430 nm, and the other with a peak absorption between 500 nm and 540 nm. The data also provided evidence for some differential bleaching of the pigments in the presence of a colored adapting light, but most of the adaptation effect was probably due to changes in screening pigment and neural desensitization or inhibition. These two observations suggest that an adequate substrate for color vision may exist in this and other species of fiddler crabs. The electroretinogram and spike-recording methods produced similar visual-sensitivity data, suggesting that latter technique, a much more efficient way of collecting data that is physiologically relevant, may be the method of choice for determining spectral sensitivity in crustaceans. PMID- 12122470 TI - Proteins: from chemical to physiological mechanism. PMID- 12122471 TI - Kinetic studies of calcium and cardiac troponin I peptide binding to human cardiac troponin C using NMR spectroscopy. AB - Ca2+ and human cardiac troponin I (cTnI) peptide binding to human cardiac troponin C (cTnC) have been investigated with the use of 2D [1H,15N] HSQC NMR spectroscopy. The spectral intensity, chemical shift, and line-shape changes were analyzed to obtain the dissociation ( K(D)) and off-rate ( k(off)) constants at 30 degrees C. The results show that sites III and IV exhibit 100-fold higher Ca2+ affinity than site II ( K(D(III,IV)) approximately 0.2 microM, K(D(II)) approximately 20 microM), but site II is partially occupied before sites III and IV are saturated. The addition of the first two equivalents of Ca2+ saturates 90% of sites III and IV and 20% of site II. This suggests that the Ca2+ occupancy of all three sites may contribute to the Ca2+-dependent regulation in muscle contraction. We have determined a k(off) of 5000 s(-1) for site II Ca2+ dissociation at 30 degrees C. Such a rapid off-rate had not been previously measured. Three cTnI peptides, cTnI(34-71), cTnI(128-147), and cTnI(147-163), were titrated to Ca2+-saturated cTnC. In each case, the binding occurs with a 1:1 stoichiometry. The determined K(D) and k(off) values are 1 microM and 5 s(-1) for cTnI(34-71), 78+/-10 microM and 5000 s(-1) for cTnI(128-147), and 150+/-10 microM and 5000 s(-1) for cTnI(147-163), respectively. Thus, the dissociation of Ca2+ from site II and cTnI(128-147) and cTnI(147-163) from cTnC are rapid enough to be involved in the contraction/relaxation cycle of cardiac muscle, while that of cTnI(34-71) from cTnC may be too slow for this process. PMID- 12122472 TI - How to get from A to B: strategies for analysing protein motion on DNA. AB - Essentially all genetic events require proteins to move from one location in a DNA polymer to another location in the same chain. A protein will seldom bind to a specific site in the DNA by colliding directly with that site. Instead, the protein will almost always collide first with a random site anywhere in the DNA and then migrate to the specific site by a facilitated-diffusion process that is constrained to the zone of that DNA molecule. Thereafter, many proteins bound to their target sites translocate in a specified direction along the DNA by a energy dependent vectorial mechanism. This review will discuss some of the strategies that have been developed to analyse the motion of proteins on DNA, with respect to both the random diffusion processes involved in target-site location by DNA binding proteins and the vectorial processes involved in unidirectional translocation along DNA. PMID- 12122473 TI - Exchange factors, effectors, GAPs and motor proteins: common thermodynamic and kinetic principles for different functions. AB - In this article, we review the properties of several classes of proteins that interact with ATPases and GTPases involved in energy and signal transduction. We show that certain common basic thermodynamic principles apply to the manner in which the nucleotide hydrolases interact with their partner molecules, and that the principles involved in signal transduction can be quantitatively modified to give systems with the known properties of energy transducing systems. As an example, actin can be described as an exchange factor for myosin, with its exchange activity being specific for ATP or inorganic phosphate in the myosin.ADP.P(i) complex, in contrast to the unspecific exchange activity of guanosine nucleotide exchange factors operating on GTPases involved in signal transduction and regulatory processes. These common aspects are reflected in shared structural features, suggesting an evolutionary relationship between such systems. PMID- 12122474 TI - Oligomerization and kinetic mechanism of the dynamin GTPase. AB - Dynamin is a large molecular weight GTPase. Amongst other biological processes, it is involved in clathrin-dependent endocytosis. It can self-assemble or assemble on other macromolecular structures that result in an increase in its GTPase activity. Its role in endocytosis has been variously attributed to being a force-generating enzyme or a signalling protein. Here we review evidence for the oligomeric state of dynamin at high and low ionic strength conditions. We also review work on the elementary processes of the dynamin GTPase at high ionic strength and compare these to the ATPase of the force-generating protein myosin and the GTPase of the signalling protein Ras. New data on the interaction of dynamin with a fluorescent derivative of GTPgammaS are also presented. The possible mechanism by which assembly of dynamin leads to an increase in its GTPase activity is discussed. PMID- 12122475 TI - Ion channels: recent progress and prospects. AB - Determination of the crystal structure of the KcsA potassium channel and its subsequent refinement at 2 A resolution have stimulated much interest in modelling of ion channels. Here we review the recent developments in ion channels research, focusing especially on the question of structure-function relationships, and discuss how permeation models based on Brownian and molecular dynamics simulations can be used fruitfully in this endeavour. PMID- 12122476 TI - Ion passage pathways and thermodynamics of the amphotericin B membrane channel. AB - Amphotericin B is a polyene macrolide antibiotic used to treat systemic fungal infections. Amphotericin B's chemotherapeutic action requires the formation of transmembrane channels, which are known to transmit monovalent ions. We have investigated the ion passage pathways through the pore of a realistic model structure of the channel and computed the associated thermodynamic properties. Our calculations combined the free energy computations using the Poisson equation with a continuum solvent model and the molecular simulations in which solvent molecules were present explicitly. It was found that there are no substantial structural barriers to a single sodium or chloride ion passage. Thermodynamic free energy calculations showed that the path along which the ions prefer to move is off center from the channel's central axis. In accordance with experiments, Monte Carlo molecular simulations established that sodium ions can pass through the pore. When it encounters a chloride anion in the channel, the sodium cation prefers to form a solvent-bridged pair configuration with the anion. PMID- 12122477 TI - The role of residue 50 and hydration water molecules in homeodomain DNA recognition. AB - We conducted molecular dynamics simulations on several wild-type and mutant homeodomain-DNA complexes to investigate the role of residue 50 in homeodomain DNA interaction and the behavior of interfacial hydration water. Our results suggest that this residue interacts more favorably with its consensus sequence and thus plays a considerable role in DNA recognition. However, residue 50 was not responsible for DNA recognition alone. Other residues in the vicinity could interact with residue 50 in cooperation upon DNA binding. We also found the lifetime for some water in the protein-DNA interface can be as high as nanoseconds and that a few well-conserved sites for water-mediated hydrogen bonds from protein to DNA are occupied by high-mobility hydrating waters. PMID- 12122478 TI - Lateral organization of GM1 in phase-separated monolayers visualized by scanning force microscopy. AB - Phase separation of glycolipids in lipid mono- and bilayers is of great interest for the understanding of membrane function. The distribution of the ganglioside GM1 in sphingomyelin (SM)/1-palmitoyl-2-oleoyl- sn-glycero-3-phosphocholine (POPC), SM/1,2-dipalmitoyl- sn-glycero-3-phosphocholine (DOPC) and SM/cholesterol/POPC Langmuir-Blodgett (LB) monolayers transferred at 36 mN/m has been studied by scanning force microscopy. Besides lateral organization of the glycolipid in LB monolayers as deduced from topography, material properties have been investigated by phase imaging, pulsed force mode and force modulation microscopy. It was shown that GM1 preferentially clusters in an ordered lipid matrix, i.e. the SM phase in the case of the SM/POPC and SM/DOPC mixture or in the ordered phase of POPC/SM/cholesterol monolayers. At higher local concentrations, three-dimensional protrusions enriched in GM1 occur, which may represent a precursor for the formation of micelles budding into the aqueous subphase. Electronic supplementary material to this paper can be obtained by using the Springer Link server located at http://dx.doi.org/10.1007/s00249-002 0232-4. PMID- 12122479 TI - Evidence for dopamine D(1) receptor involvement in the stimulus selection task: overshadowing in the rat. AB - RATIONALE: A number of lines of evidence suggest that dopamine might play a role in stimulus selection, the process whereby specific cues are selected to guide action. OBJECTIVES: In order further to define the potential role for dopamine in stimulus selection, the present series of studies examined whether dopaminergic drugs modulate overshadowing, a paradigm that involves stimulus selection in rats. Overshadowing is where preferential learning occurs to one (usually the more salient) element of a stimulus compound. METHODS: Overshadowing was measured in rats using a thirst motivated conditioned emotional response paradigm (CER). Two simultaneously presented stimuli (light and tone) were paired with an aversive unconditioned stimulus (mild footshock); overshadowing is observed when learning to the less salient stimulus is weaker than learning to the same stimulus when it is conditioned alone. RESULTS: d-Amphetamine sulphate (1 mg/kg, IP) was found selectively to disrupt overshadowing, without affecting the CER in control animals. The dopamine (DA) D(2) receptor antagonists, haloperidol (0.2 mg/kg, IP) or raclopride (0.5 mg/kg, IP), failed to reverse amphetamine-induced disruption of overshadowing. In contrast, the selective DA D(1) antagonist SCH 23390 (0.05 mg/kg, IP) reversed amphetamine-induced disruption of overshadowing. The partial DA D(1) agonist SKF 38393 (5 mg/kg, IP) was found to abolish overshadowing when given alone. CONCLUSION: These data indicate a modulatory role for the DA D(1) receptor in the expression of stimulus selection and suggest that the DA D(1) receptor might play a role in salience allocation aspects of learning. PMID- 12122480 TI - Imipramine restores the long-term impairment of appetitive behavior in socially stressed rats. AB - RATIONALE: Previous observations revealed that defeated and subsequently individually housed rats do not display behavioral anticipatory responses to the conditioned presentation of a bell/light stimulus associated with free access to a 5% sucrose solution reward. The absence of the appetitive responses suggests a decreased sensitivity to reward. This might be homologous to anhedonia, a symptom of human depression. OBJECTIVES: To further validate the inability to anticipate as indicative for a depressionlike state we investigated whether antidepressant treatment restores the impaired anticipatory responses in defeated and subsequently individually housed animals. METHODS: Male rats were defeated and subsequently individually housed or subjected to a control treatment. In the 19 20th weeks after the exposure to defeat rats were either injected daily with imipramine (20 mg/kg per os, dissolved in water) or water. Anticipatory behavior was measured both before and after 3-5 weeks of chronic treatment with imipramine. RESULTS: The long-term impairment of anticipatory behavior in defeated and subsequently individually housed rats was restored by chronic imipramine treatment. Impaired appetitive behavior in socially stressed rats was not accompanied by a decreased consumption of the 5% sucrose solution in the anticipatory tests. The recovery of the appetitive responses was independent of open field activity, body weight, and 5% sucrose preference in the home cage. CONCLUSIONS: Chronic imipramine administration restores the anhedonialike absence of anticipatory behavior in socially stressed rats. Predictive validity of the social stress model of human depression is suggested by the similar action of imipramine on the modeled behavior and on the anhedonia symptoms in depressive human patients. PMID- 12122481 TI - Effects of the nitric oxide donor molsidomine on different memory components as assessed in the object-recognition task in the rat. AB - RATIONALE: Nitric oxide (NO) is sought to be a novel intracellular messenger in the central nervous system. Recently, NO involvement in learning and memory processes has been proposed. Compounds that inhibit NO synthase, the key synthesizing enzyme, may block cognition, while NO donors may facilitate it. OBJECTIVES: The present study was designed to further investigate in the rat the effects on distinct memory processes exerted by the NO donor molsidomine. For this aim, the object-recognition task was chosen. This test is based on the differential exploration of a new and familiar object.METHODS. Object recognition was evaluated in a two-trial nonrewarded paradigm. In a first study, the influence of the retention time (the delay between the two trials) on the performance of 3-month-old male rats was assessed. Subsequently, the effects of molsidomine (2 mg/kg and 4 mg/kg), injected i.p., on acquisition, storage, and retrieval of information were evaluated. For the latter experiments, the delay condition at which recognition memory was extinguished in the normal rat was used (24 h). RESULTS: Under our experimental conditions, object recognition was extinguished in the rat when an intertrial interval (ITI) of 24 h was utilized. Using this ITI, molsidomine at 4 mg/kg but not at 2 mg/kg improved the animal's performance in the object-recognition task, suggesting that molsidomine affected acquisition, storage, and retrieval of information. CONCLUSIONS: These results indicate that the NO donor molsidomine may modulate different aspects of memory. PMID- 12122482 TI - Subchronic caffeine administration sensitizes rats to the motor-activating effects of dopamine D(1) and D(2) receptor agonists. AB - RATIONALE: Dopamine transmission acting at either the D(1) or D(2) receptor level is known to influence the stimulant properties of caffeine, an antagonist of adenosine A(1) and A(2) receptors. In contrast, how caffeine influences the motor stimulant properties of selective D(1) and D(2) receptor agonists is still undefined. OBJECTIVES: In this study the acute motor response to the dopamine D(1) receptor agonist SKF 77434 and the D(2)/D(3) receptor agonist quinpirole was studied in rats treated subchronically with caffeine (15 mg/kg i.p., on alternate days) or vehicle, either in the test cage (paired group) or in the home cage (home group). METHODS AND RESULTS: Repeated caffeine administration did not induce any significant increase in motor activity or in stereotyped behavior during the course of treatment, indicating that the response to caffeine itself did not develop sensitization. Three days after the last caffeine or vehicle administration, rats were challenged with caffeine, SKF 77434, or quinpirole. Caffeine (5 mg/kg i.p.) elicited the same motor stimulant effects in both caffeine- and vehicle-pretreated rats, confirming the presence of neither tolerance nor sensitization to caffeine itself. SKF 77434 (3 mg/kg s.c.) elicited a higher locomotor activation in caffeine- than in vehicle-pretreated rats, whereas quinpirole (0.15 mg/kg s.c.) induced a similar locomotor activation and a higher stereotyped behavior in caffeine-pretreated rats as compared to rats pretreated with vehicle. The sensitized response to SKF 77434 and quinpirole was not due to environmental conditioning since the responses were similar in either paired or home group. CONCLUSIONS: The results provide support for the presence of long-term functional interactions between drugs acting at the adenosine and dopamine receptor levels. Subchronic caffeine, by sensitizing the motor stimulant effects of dopamine D(1) and D(2) receptor agonists, produces adaptive changes which might result in a potentiation of the dopaminergic component of drugs of abuse. PMID- 12122483 TI - Age-related differences in sensitivity to the antinociceptive effects of kappa opioids in adult male rats. AB - RATIONALE: Significant differences in the potency and effectiveness of opioid analgesics have been reported in subject populations differing in age. Although the relationship between aging and sensitivity to the antinociceptive effects of mu opioids has been examined extensively, relatively few studies have examined this relationship in kappa opioids. OBJECTIVES: The purpose of the present investigation was to examine the antinociceptive effects of selected kappa and mixed-action opioids in young (3 months) and aged (21 months) male rats. METHODS: In a warm-water, tail-withdrawal procedure, rats were restrained and the latencies to remove their tails from 50 degrees C (low temperature) and 55 degrees C (high temperature) water were measured. Selected kappa (U69,593, U50,488) and mixed-action (butorphanol, nalbuphine) opioids were tested alone, and in combination with the high-efficacy, kappa-opioid spiradoline. RESULTS: All test drugs were more effective (i.e., produced a greater maximal effect) in aged rats than in young rats at both water temperatures. In drug combination tests, U69,593 and U50,488 enhanced the effects of spiradoline under conditions in which they failed to produce high levels of antinociception when administered alone. In contrast, butorphanol and nalbuphine antagonized the effects of spiradoline under conditions in which they failed to produce high levels of antinociception when administered alone. CONCLUSIONS: These data may be taken as evidence that: (1) aged male rats are more sensitive than young male rats to the antinociceptive effects of kappa opioids, (2) U69,593 and U50,488 display agonist activity in the warm-water, tail-withdrawal procedure under some conditions in which they fail to produce antinociceptive effects, and (3) butorphanol and nalbuphine possess only limited agonist activity at the kappa receptor. PMID- 12122484 TI - Nicotine and its withdrawal alter feeding induced by paraventricular hypothalamic injections of neuropeptide Y in Sprague-Dawley rats. AB - RATIONALE: Cigarette smoking produces feeding and weight suppression in humans that often rebound following cessation. Nicotine (NIC) administration produces similar effects in rats, but the neural mechanisms responsible are not fully known. Recent evidence shows that hypothalamic levels of neuropeptide Y (NPY) change with NIC administration. Infusions of NPY into the paraventricular nucleus of the hypothalamus (PVN), which normally produce robust feeding, were used to investigate changes in the PVN-NPY system that may contribute to NIC's effects on energy balance. OBJECTIVE: To characterize potential differences in PVN-NPY induced feeding during NIC treatment versus withdrawal. METHODS: Three groups of female rats ( n=66) bearing unilateral PVN cannulae were implanted for 14 days with subcutaneous Alzet mini-pumps containing NIC (0, 6, or 12 mg/kg per day). Dark-onset (1800-2000 hours) NPY feeding tests occurred five times: pre-implant, 2 days and 12 days post-implant and 2 days and 8 days after implant removal. Feeding tests consisted of 1 h of pre-feeding prior to lights off, then two 1-h measures of feeding after PVN injections of 0.4 microl saline or NPY (78 pmol, 235 pmol). RESULTS: NIC initially suppressed body weight gain, followed by steady recovery that was briefly exaggerated after withdrawing NIC. Daily feeding was acutely suppressed by NIC but acutely potentiated after NIC cessation. PVN-NPY induced feeding was suppressed by both doses of NIC 2 days after pump implant, elevated 2 days after pump removal, but returned to pre-NIC levels 8 days after pump removal. CONCLUSIONS: These findings provide behavioral support that changes in PVN-NPY neurotransmission may play a functional role in the food intake and weight-modulating effects of NIC. PMID- 12122485 TI - Characterization of the discriminative stimulus effects of N-methyl- D-aspartate ligands under different ethanol training conditions in the cynomolgus monkey ( Macaca fascicularis). AB - RATIONALE: The current study was designed to extend our knowledge of the N-methyl D-aspartate (NMDA) glutamate receptor system in mediating the discriminative stimulus effects of ethanol in non-human primates. OBJECTIVES: To characterize the discriminative stimulus effects of the NMDA uncompetitive antagonists dizocilpine, phencyclidine (PCP) and ketamine in male and female monkeys under different ethanol training conditions. METHODS: Adult male ( n=8) and female ( n=9) cynomolgus monkeys ( Macaca fascicularis) were divided into four groups and trained to discriminate 1.0 g/kg ethanol ( n=8) versus water or 2.0 g/kg ethanol ( n=9) versus water in a 2 x 2 design with training dose and sex as main group factors. Ethanol (20% w/v) solutions were administered intragastrically (IG) and responding was maintained under a fixed ratio schedule of food reinforcement. Dose-response determinations for dizocilpine [IG and intramuscular (IM)], PCP (IM) and ketamine (IM) were made under two training intervals (30 and 60 min). RESULTS: Dizocilpine, PCP and ketamine dose-dependently substituted for ethanol in three of four training conditions, the notable exception being in males trained with 2.0 g/kg ethanol. Ethanol-like discriminative stimulus effects were greater with IM dizocilpine than with IG dizocilpine. At the lower ethanol training dose (1.0 g/kg), there were no sex differences in the ethanol-like discriminative stimulus effects of dizocilpine, PCP or ketamine, nor were there sex differences in the potencies to produce ethanol-like discriminative stimulus effects. Sex differences were readily apparent with the higher ethanol training dose (2.0 g/kg), with the NMDA ligands failing to substitute for ethanol in male monkeys, probably due to the rate-suppressive effects of these compounds. CONCLUSIONS: These data suggest that NMDA receptor-mediated activity is a component to the discriminative stimulus effects of ethanol in male and female nonhuman primates. However, NMDA uncompetitive antagonists were less likely to produce discriminative stimulus effects similar to a high ethanol training dose in male monkeys. In comparison to consistent substitution by GABA(A) positive modulators for ethanol, substitution patterns produced by NMDA uncompetitive antagonists suggest a less robust mediation of the ethanol discriminative stimulus through NMDA receptor systems in nonhuman primates. PMID- 12122486 TI - Multiple 5-HT receptors are involved in the effects of acute MDMA treatment: studies on locomotor activity and responding for conditioned reinforcement. AB - RATIONALE: Responding for conditioned reinforcement is increased by the dopamine releasing agent amphetamine, but reduced by drugs that enhance serotonin (5-HT) function. The amphetamine derivative 3,4-methylenedioxymethamphetamine (MDMA, Ecstasy) releases both monoamines. OBJECTIVES: The primary purpose of this study was to examine the effects of MDMA on responding for conditioned reinforcement as well as on locomotor activity. The roles of several 5-HT receptor sub-types in mediating these behavioural effects of MDMA were also examined. METHODS: Locomotion was measured in photocell activity monitors. For conditioned reinforcement experiments thirsty rats learned to associate a conditioned stimulus (CS) with water in operant chambers. Subsequently, two response levers were available; responding on one lever delivered the CS, while responding on the second lever had no consequences. Drug effects on this operant response were measured. RESULTS: MDMA dose-dependently increased locomotion but reduced responding for conditioned reinforcement. This latter effect differs from that induced by amphetamine, which potentiates conditioned reinforcement responding. The stimulant effect of MDMA was attenuated by GR127935 and ketanserin, indicating facilitatory roles of 5-HT(1B) and 5-HT(2A) receptors in mediating this effect. The 5-HT(2C) antagonist SB242084 enhanced the stimulant effect of MDMA. Only SB242084 attenuated the suppressant effect of MDMA on responding for conditioned reinforcement. CONCLUSIONS: The results show that 5-HT(2A) and 5 HT(1B/1D) receptors play a facilitatory role in mediating the stimulant effect of MDMA, whereas 5-HT(2C) receptors are inhibitory. Activation of 5-HT(2C) receptors also contributes to the deficit in operant responding. Multiple 5-HT receptor sub types appear to contribute to the behavioural effects of MDMA. PMID- 12122487 TI - Memory-related task performance by aged rhesus monkeys administered the muscarinic M(1)-preferring agonist, talsaclidine. AB - RATIONALE: Muscarinic-acetylcholine receptor agonists are yet to be used clinically for the treatment of Alzheimer's disease (AD) even though laboratory evidence continues to support the potential for such an approach. OBJECTIVES: The purpose of this study was to evaluate the M(1)-preferring agonist talsaclidine in aged monkeys for effects on working memory. METHODS: Three doses (0.6, 1.2, and 2.4 mg/kg, PO) of talsaclidine and two time intervals (45 min and 8 h) after drug administration were evaluated in seven aged rhesus macaques trained to perform a computer-assisted delayed matching-to-sample (DMTS) task. The relative effectiveness of talsaclidine was also compared with another M(1)-preferring agonist WAY-132983 that was previously studied in this laboratory. RESULTS: Talsaclidine improved DMTS accuracy only during sessions initiated 8 h after administration of one of the doses (i.e. 0.6 mg/kg). The drug's enhanced effectiveness at the 8-h time point relative to the 45-min time point was surprising in view of the fact that plasma concentrations were highest 45 min after administration. A higher dose of talsaclidine (4.7 mg/kg) resulted in side effects (lethargy and excessive drooling) in some animals. Individualized optimal doses of talsaclidine were associated with 7.4% and 10.6% improvement in overall (all trials averaged) DMTS accuracy during the 45 min and 8 h post-administration sessions, respectively. Under similar experimental conditions WAY-132983 increased DMTS accuracy by up to 15.6% above control levels. CONCLUSION: Both talsaclidine and WAY-132983 provide at least modest improvements in DMTS accuracy in aged monkeys at some doses; however, challenges remain regarding the achievement of an adequate level of efficacy and reliability while minimizing side effects with these compounds. The positive findings do, however, support further study of the potential use of direct muscarinic agonists in the treatment age-related disorders of memory function. PMID- 12122488 TI - Peripheral-type benzodiazepine receptors on platelets are correlated with the degrees of anxiety in normal human subjects. AB - RATIONALE: Anxiety is the one of the main symptoms of psychiatric disorders. Psychosocial stressors have been shown to be related to the onset of anxious episodes. Peripheral-type benzodiazepine receptors (PBR) are involved in regulating stress responses. The sensitivity of PBR to acute or chronic stress has been demonstrated in various situations. The State-Trait Anxiety Inventory (STAI) is one of the longest standing and most frequently used measures of anxiety. The development, evaluation, and use of biological markers with anxious conditions in psychiatry are extremely important. OBJECTIVES: The aims of this survey are to see whether PBR can be used in screening the degrees of anxiety which occur when normal persons are placed in the stressful conditions. METHODS: Twenty-four healthy volunteers (14 men, 10 women; mean age 46 years) participated in this study. We administered the STAI to all the volunteers. The binding of the radioactive PBR antagonist [(3)H]PK 11195 to platelet membranes was determined for these volunteers. RESULTS: The mean STAI scores were 40.3+/-8.0 for trait anxiety and 39.0+/-8.9 for state anxiety. B(max) of the platelet PBR binding was 2845+/-2109 fmol/mg protein. Pearson correlational analyses revealed that B(max) values were significantly and positively correlated with scores for trait anxiety but not significantly correlated with scores for state anxiety. CONCLUSIONS: PBR on platelets are correlated with trait anxiety scales of the STAI in healthy normal subjects. It is therefore suggested that the density of platelet PBR is highly associated with these personality traits for anxiety tolerance. PBR density in platelet could also be used as a promising biological marker of stressful conditions. PMID- 12122489 TI - Actions of alpha-2 noradrenergic agonists on spatial working memory and blood pressure in rhesus monkeys appear to be mediated by the same receptor subtype. AB - RATIONALE: alpha-2 Noradrenergic agonists improve spatial working memory in animals and in humans. Of the three alpha-2 receptor subtypes, evidence has suggested that this cognitive improvement may be mediated by the alpha-2A receptor subtype, but this has not been established. alpha-2 Agonists are also known to decrease blood pressure significantly. Recent evidence using genetically altered mice indicates that the alpha-2A receptor subtype mediates this decrease in blood pressure. OBJECTIVES: The present study examined whether the cognitive improvement and hypotension produced by alpha-2 agonists are mediated by the same receptor subtype in rhesus monkeys. The hypotensive and cognitive-enhancing effects of clonidine and guanfacine were challenged with two alpha-2 antagonists with differing affinities for the three alpha-2 receptor subtypes: MK912, a potent antagonist which shows preferential binding to the alpha-2C receptor subtype, and idazoxan, which slightly prefers the alpha-2A receptor subtype. If alpha-2C receptors contribute to the cognitive enhancement, MK912 should reverse the cognitive-enhancing effects of alpha-2 agonists at lower doses than those needed to reverse the hypotensive effects of these compounds. Conversely, if alpha-2A receptors contribute to cognitive enhancement, MK912 and idazoxan should reverse the cognitive-enhancing effects of alpha-2 agonists at the same doses as those needed to reverse the hypotensive effects of these compounds. RESULTS: MK 912 and idazoxan dose-dependently reversed both clonidine and guanfacine-induced cognitive improvement and hypotension. Both antagonists were equally potent in reversing either the cognitive enhancement or the hypotension. CONCLUSIONS: The identical pattern of dose-dependent reversal of cognitive improvement and hypotension indicates that, in non-human primates, the same receptor subtype mediates both effects. Previous evidence suggests that the most likely candidate is the alpha-2A receptor subtype. PMID- 12122490 TI - Initial sensitivity, tolerance and cross-tolerance to allopregnanolone- and ethanol-induced hypothermia in selected mouse lines. AB - RATIONALE: Acute ethanol administration induces hypothermia in genetically susceptible animals. Tolerance to this effect may develop with repeated administration. Allopregnanolone is an endogenously produced neuroactive steroid that acts at the GABA-A receptor. We postulated that allopregnanolone would induce hypothermia, and that lines of mice selectively bred for high (COLD-1 and COLD-2) or low (HOT-1 and HOT-2) sensitivity to ethanol's hypothermic effects would also be differentially sensitive to allopregnanolone-induced hypothermia. We also postulated that tolerance would develop to these two drugs by similar mechanisms, such that tolerance to one would impart cross-tolerance to the other. OBJECTIVES: To assess sensitivity, tolerance and cross-tolerance to allopregnanolone's and ethanol's hypothermic effects in HOT-1 and 2, and COLD-1 and 2 mice. METHODS: Mice were administered one of several doses of allopregnanolone each day, for 4 days, and initial sensitivity and tolerance to allopregnanolone-induced hypothermia were assessed. On day 5, ethanol was administered to assess cross-tolerance. In a separate experiment, COLD-1 and 2 mice were made tolerant to ethanol's hypothermic effects, and challenged with allopregnanolone to assess cross-tolerance. RESULTS: COLD mice exhibited greater initial sensitivity to the hypothermic effect of allopregnanolone, as compared to HOT mice. Tolerance to allopregnanolone-induced hypothermia was greater in COLD mice than in HOT mice, but only COLD-1 mice showed cross-tolerance to ethanol. Both replicate lines of COLD mice developed tolerance following repeated administration of ethanol, but only COLD-2 mice showed cross-tolerance to allopregnanolone. CONCLUSIONS: These results demonstrate shared genetic influence over allopregnanolone and ethanol's initial hypothermic effects. They also suggest genotype-dependent differences in the mechanisms for tolerance to these two compounds. PMID- 12122491 TI - Neurochemical and behavioural characterization of milnacipran, a serotonin and noradrenaline reuptake inhibitor in rats. AB - RATIONALE: The prefrontal cortex is implicated in the pathophysiology of depression, and hypoactivity of this brain area has been found in depressed patients. Reduced function of the serotonergic and noradrenergic systems is another feature of depression. OBJECTIVES: The present study was aimed at characterizing neurochemically and behaviorally the serotonin and noradrenaline reuptake inhibitor (SNRI), milnacipran, in the prefrontal cortex in comparison with tricyclic antidepressants and selective serotonin reuptake inhibitors. METHODS: Sodium-dependent monoamine uptake measurement, radioligand binding assays, microdialysis procedure, forced swimming test and conditioned fear stress test were carried out in rats. RESULTS: Milnacipran selectively inhibited sodium dependent [(3)H]serotonin (5-hydroxytryptamine, 5-HT) and [(3)H]noradrenaline (NA) uptake into the synaptosomes from rat cerebral cortex (IC(50)=28.0 and 29.6 nM, respectively) without any affinities for various neuroreceptors. In the medial prefrontal cortex, milnacipran (10 and 30 mg/kg, PO) caused a dose-related increase in the extracellular levels of 5-HT and NA with similar potency, whereas imipramine (10 and 30 mg/kg, PO) caused a dominant increase in the output of NA compared to 5-HT. Milnacipran (30 and 60 mg/kg, PO) significantly reduced the duration of both the immobility time in the forced swimming test and the freezing time in the conditioned fear stress test in rats, which are animal behavioral models for depression and anxiety, respectively. Imipramine and maprotiline were active in the former test, but not in the latter. Fluoxetine and fluvoxamine on the other hand were more active in the conditioned fear test. CONCLUSION: These findings show that milnacipran acts as a SNRI in vitro and in vivo and may be useful for the treatment of anxiety as well as depression. PMID- 12122492 TI - Positive relationship between activity in a novel environment and operant ethanol self-administration in rats. AB - RATIONALE: The study of individual differences in drug addiction may have important implications both for understanding the etiology of addiction and for strategies for treatment. Activity of rodents in novel environments, presumably related to the novelty-seeking trait in humans, is the primary behavioral feature that is hypothesized to predict a predisposition for drug self-administration by rodents. OBJECTIVES: The aim of this study was to characterize the relationship between motor activity in a novel environment and operant ethanol self administration using the sucrose-substitution procedure. METHODS: Male Long-Evans rats were exposed to a novel environment for 2 h, and the distance traversed, rearing, and defecation was recorded. After 3 days of forced exposure to ethanol the sucrose-substitution procedure began and lasted for 23 days. Following sucrose substitution the animals were maintained on a schedule of ethanol (10% v/v) self-administration with a fixed ratio 3 (FR3) for 15 days. RESULTS: The activity (distance traversed) in the novel environment was positively correlated with initial ethanol self-administration under the FR3 schedule ( r=0.87) but not with the number of inactive lever presses or active lever presses for ethanol or for sweetened ethanol solutions with lower ratios of response. In contrast, rearing was correlated positively only with the number of inactive lever presses for sucrose. CONCLUSIONS: Motor activity in a novel environment may be related to the acquisition of operant ethanol self-administration only when a given ratio of response is required. PMID- 12122493 TI - Imipramine, but not lithium, induces the serine/threonine phosphatase activity of calcineurin without affecting its mRNA expression in the rat brain. AB - The influence of imipramine and lithium on the expression of calcineurin (CaN) and its serine/threonine phosphatase activity in the rat frontal cortex and hippocampus was examined. Northern blot analysis demonstrated that single or repeated (14 day) administration of imipramine did not affect the levels of CaN A (catalytic subunit) mRNA in rat brain. Similarly, the administration of lithium for 1 or 14 days had no effect on the expression of CaN A mRNA. In contrast, the acute administration of imipramine significantly increased CaN activity in the cortex and hippocampus. In addition, the chronic (14 day) administration also significantly increased CaN activity. However, administration of lithium for either 1 or 14 days did not influence CaN activity. These findings indicate that imipramine, but not lithium, increases the phosphatase activity of CaN, suggesting that the changes in neuronal functions induced by CaN may be involved in the molecular activity of imipramine. PMID- 12122494 TI - M(3) receptor antagonism by the novel antimuscarinic agent solifenacin in the urinary bladder and salivary gland. AB - The antimuscarinic profile of the experimental drug solifenacin/YM905 [(+)-(1 S,3' R)-quinuclidin-3'-yl 1-phenyl-1,2,3,4-tetrahydroisoquinoline-2-carboxylate] for the treatment of overactive bladder was compared with the commonly prescribed agent oxybutynin. In radioligand binding assays, p K(i) values of solifenacin for M(1), M(2), and M(3) receptors were 7.6, 6.9, and 8.0, respectively. These values for oxybutynin were 8.6 (M(1)), 7.7 (M(2)), and 8.9 (M(3)). Solifenacin and oxybutynin antagonized the contractile effect of carbachol (CCh) on isolated guinea pig urinary bladder smooth muscle (detrusor), displaying the negative logarithm of antagonist apparent affinity constant (p K(b) value) of 7.1 for solifenacin and 7.4 for oxybutynin. To study the tissue selectivity between bladders and salivary glands, guinea pig detrusor and mouse submandibular gland cells were stimulated with CCh and monitored for intracellular Ca2+, as determined by Fura 2 fluorescence. Ca2+ mobilization of detrusor cells was inhibited equipotently by solifenacin (p K(i)=8.4) and oxybutynin (p K(i)=8.6), whereas that of the gland cells was antagonized less potently by solifenacin (p K(b)=7.4) than by oxybutynin (p K(b)=8.8), although the M(3) subtype mediated both cell responses. In anesthetized rats, solifenacin (63-2100 nmol kg(-1) or 0.03-1 mg kg(-1)) dose-dependently inhibited CCh-stimulated increases in urinary bladder pressure, while its inhibitory effects on salivation and bradycardia were apparent only at a dose of 2100 nmol kg(-1). In contrast, oxybutynin within a dose range of 77-770 nmol kg(-1) (0.03-0.3 mg kg(-1)) inhibited responses of the bladder and salivary gland slightly more potently than that of the heart. In addition, inhibitory effects of darifenacin indicated a major role of M(3) receptors in the bladder and salivary gland. Therefore, M(3) receptor antagonism by solifenacin could be bladder-selective. This selectivity remains to be elucidated and may provide new approaches to the pharmacotherapy of overactive bladder. PMID- 12122496 TI - Vasodilator and vasoconstrictor responses induced by 5-hydroxytryptamine in the in situ blood autoperfused hindquarters of the anaesthetized rat. AB - In the present study we attempted to characterise the responses and receptors involved in the effects of 5-hydroxytryptamine (5-HT, serotonin) in in situ autoperfused rat hindquarters. Intra-arterial administration of the lowest doses of 5-HT used (0.12-12.5 ng/kg) induced vasodilator responses, whereas the highest doses (25-1000 ng/kg) produced vasoconstriction. The vasodilator effect was inhibited by methiothepin (a non-specific 5-HT(1,2,5,6,7) receptor antagonist) and by a 5-HT(1D/1B) receptor antagonist, i.e., 3-[4-(4-chlorophenyl)piperazin-1 yl]-1,1-diphenyl-2-propanolol (BRL 15572), but not by ritanserin (a selective 5 HT(2) receptor antagonist), 5-methyl-1-(3-pyridylcarbamoyl)-1,2,3,5 tetrahydropyrrolo[2,3-f] indole (SB 206553, a selective 5-HT(2B/2C) receptor antagonist) or mesulergine (a non-specific serotonergic antagonist that shows affinity to the 5-HT(7) receptor). This vasodilator effect was mimicked by administration of a selective 5-HT(1) receptor agonist - 5-carboxamidotryptamine (5-CT) - and by 2-[5-[3-(4-methylsulphonylamino)benzyl-1,2,4-1 H-indol-3 yl]ethanamine (L-694,247, a selective 5-HT(1D/1B) receptor agonist). Methiothepin, but not mesulergine, inhibited 5-CT-induced vasodilatation and the selective 5-HT(1D/1B) receptor antagonist (BRL 15572) inhibited the vasodilator action induced by L-694,247. The vasoconstrictor effect of 5-HT was significantly decreased by methiothepin, ritanserin and SB 206553, and was mimicked by alpha methyl-5-HT (a selective 5-HT(2) receptor agonist) but not by administration of BW 723C86, a selective 5HT(2B) receptor agonist. Ritanserin, SB 206553 and spiperone (a non-specific 5-HT(1/2A) receptor antagonist) inhibited the alpha methyl-5HT-induced vasoconstriction.Our data suggest that the vasodilator response induced by 5-HT in autoperfused rat hindquarters is mainly mediated by 5 HT(1D/1B) receptors, whereas the vasoconstrictor effect is mainly due to the activation of 5-HT(2A) receptors. PMID- 12122495 TI - Noradrenaline-induced contraction of human saphenous vein and human internal mammary artery: involvement of different alpha-adrenoceptor subtypes. AB - Although saphenous veins and internal mammary arteries are commonly used for coronary artery bypass grafting, only a very few comparative studies are available on alpha-adrenoceptor-mediated vasoconstriction in these vessels. Thus, we determined, in isolated rings from human saphenous vein and human internal mammary artery, contractile responses to noradrenaline (10(-8)-10(-4) M) in the absence and presence of the alpha-adrenoceptor antagonists yohimbine (alpha(2) adrenoceptor antagonist, 10(-8)-10(-6) M), prazosin (alpha(1)-adrenoceptor antagonist, 10(-9)-10(-7) M), 5-methyl-urapidil (5-MU, alpha(1A)-adrenoceptor antagonist, 10(-8)-10(-6) M), BMY 7378 (alpha(1D)-adrenoceptor antagonist, 10(-7) 10(-6) M), and chloroethylclonidine (CEC, irreversible alpha(1B)-adrenoceptor antagonist, 3x10(-5) M for 30 min). All experiments were carried out in the presence of 10(-7) M propranolol and 10(-5) M cocaine. In both vessel types noradrenaline evoked concentration-dependent contractions. In saphenous veins yohimbine was a potent antagonist (pA(2)-value 8.32) while prazosin, 5-MU and BMY exhibited only marginal antagonistic effects. CEC, however, significantly decreased noradrenaline-induced contractions. In contrast, in internal mammary arteries prazosin (pA(2)-value 9.65) and 5-MU (pK(B)-values 7.2-7.5) were potent antagonists, while yohimbine and BMY exhibited only weak antagonistic effects. CEC, however, significantly decreased noradrenaline-induced contractions. We conclude that in saphenous vein the contractile response to noradrenaline is mediated predominantly by alpha(2)-adrenoceptors, while in internal mammary artery it is mediated (to a major part) by alpha(1B)- and (to a minor part) by alpha(1A)-adrenoceptors. PMID- 12122497 TI - Spermidine attenuates the inhibitory effect of ethanol on NMDA-induced neurotoxicity. AB - The exact site(s) and the molecular mechanism(s) by which ethanol inhibits the activity of NMDA receptors in the brain have so far not been identified although the involvement of several NMDA receptor modulatory sites activated by glycine, Mg2+, Zn2+, polyamines and red-ox agents has been suggested. In this study we investigated the effects of spermidine, a polyamine site agonist, on NMDA-induced neurotoxicity and its ability to modulate the inhibitory action of ethanol on neurotoxicity produced by the maximal neurotoxic concentration of NMDA as measured by the MTT assay in rat cerebellar granule cells. This assay measures the enzymatic activity in mitochondria and/or endosome/lysosome compartment that closely correlates with the cell viability. Spermidine dramatically potentiated NMDA-induced responses both at nontoxic and maximally neurotoxic NMDA concentrations. Ethanol, as expected, concentration-dependently inhibited the maximal neurotoxicity produced by NMDA. The potentiating effect of spermidine observed at nontoxic concentrations of NMDA was not altered by ethanol evidenced by the fact that the EC(50) value for spermidine was not significantly changed in the presence of ethanol. This suggests that ethanol and spermidine produce their effects by acting at different sites within the NMDA receptor complex. In contrast, the inhibitory effect of ethanol on the maximally neurotoxic action of NMDA was significantly reduced by spermidine in a concentration-dependent manner, suggesting that the spermidine enhancement of NMDA receptor function in this situation is more potent and is able mask the inhibitory action of ethanol on other sites within the NMDA receptor. PMID- 12122499 TI - Relaxation of the mouse isolated aorta and carotid artery in response to adenosine analogues in genetically-modified mice lacking the adenosine A(2A) receptor. AB - The aim of this study was to characterise the receptor(s) mediating responses to adenosine and/or adenosine analogues in mouse isolated aorta and carotid artery. In addition, since mice lacking the A(2A) adenosine receptor are reported to be hypertensive, the possibility that this gene deletion or the altered phenotype results in alteration of responses mediated via adenosine analogues was investigated. This was achieved by comparing results obtained in parallel within single experiments using tissues from A(2A) knock-out animals and their wild-type littermates.In aortic rings, adenosine and 5'- N-ethylcarboxamidoadenosine (NECA) caused relaxations above 10 microM and 30 microM, respectively, which were unaffected by either 8-sulphophenyltheophylline (8-SPT, 100 microM) or A(2A) receptor knockout. 2-[ p-(2-Carbonylethyl)phenylethylamino]-5'- N ethylcarboxamidoadenosine (CGS 21680) was virtually inactive. R- N(6) Phenylisopropyladenosine (R-PIA) induced relaxations which were not inhibited by 8-SPT (100 microM) or altered by A(2A) receptor knockout. No A(1)-mediated contractile responses were observed in wild-type or knock-out tissues in contrast with results in mice of the same strain obtained commercially rather than from our breeding programme. In carotid artery rings NECA contracted at low concentrations (0.1-1 microM) and relaxed at higher concentrations. Curves to NECA were not different in tissues from wild-type and A(2A) receptor knock-out mice and both the contractile and relaxant phases were right-shifted by 8-SPT (100 microM) in tissues from animals of both genotype. 1,3-Dipropyl-8 cyclopentylxanthine (DPCPX, 3 nM) attenuated contractile NECA responses but did not affect relaxant responses. CGS 21680 was inactive in carotid artery rings from both wild-type and A(2A) receptor knock-out mice. In the presence of DPCPX (30 nM) to abolish contractions, R-PIA induced relaxant curves which were not different in tissues from wild-type and A(2A) knock-out mice and were not inhibited by 8-SPT (100 microM). These results confirm the absence of A(2A) or A(2B) receptors in murine aorta and suggest that relaxations to NECA in carotid artery are A(2B) receptor-mediated whilst contractions are A(1) receptor mediated. They also indicate the presence of an antagonist-resistant site activated by R-PIA in both vascular preparations. There is no evidence for compensatory changes in responses mediated by adenosine and its analogues due to the gene deletion or the reported resulting hypertensive phenotype in either aortic or carotid arterial rings obtained from A(2A) knock-out mice. PMID- 12122498 TI - Local administration of ETA (but not ETB) blockers into the PAG area of the brain decreases blood pressure of DOCA-salt rats. AB - The present study investigated the role possibly played by ET-receptor antagonism at periaqueductal grey (PAG) area level in decreasing the arterial blood pressure and heart rate values reached in DOCA-salt hypertensive rats. A 3-week DOCA-salt treatment induced an increase in blood pressure of up to 174+/-3 mmHg in Sprague Dawley (SD) rats. This was paralleled by a significant increase in heart rate (HR), and endothelin-1 (ET-1) levels throughout the brain, as assessed by specific EIA ( P<0.05). In contrast, a 40% reduction of ETA mRNA levels into the brain was detected through RT-PCR. The basal MABP of DOCA rats was significantly modified by PAG injections of FR139317, an ETA receptor antagonist, or SB209670, an ETA/ETB receptor antagonist. BQ-788, an ETB receptor antagonist, was found to have no effect on blood pressure levels, while FR139317 and SB209670 also led to a significantly modified HR. PAG-endothelin ETA antagonism can therefore be said to counteract the cardiovascular changes induced by DOCA-salt treatment in rats. PMID- 12122500 TI - Enhanced stability of wild-type and constitutively active alpha(2A)-adrenoceptors by ligands with agonist, silent and inverse agonist properties. AB - The hypothesis that prolonged treatment of a constitutively active receptor with inverse agonists may lead to increased receptor density was tested for the alpha(2)-adrenoceptor (AR) inverse agonist (+)-RX 811059 at both the wild-type (WT) and Thr(373)Lys alpha(2A) ARs in CHO-K1 cells by monitoring [(3)H]RX 821002 and [(35)S]GTPgammaS binding responses. One-hundred micromolar KCl instead of NaCl in the [(35)S]GTPgammaS membrane binding assay favoured the detection of a high-magnitude constitutive alpha(2A) AR activity. Under this condition, (+)-RX 811059 was an inverse agonist [ E(max) (% vs. basal): Thr(373)Lys alpha(2A) AR ( 52+/-2) > WT alpha(2A) AR (-31+/-6)] while atipamezole was a silent neutral antagonist for both WT and Thr(373)Lys alpha(2A) ARs. The B(max) value of [(3)H]RX 821002 binding sites to membranes of transfected CHO-K1 cells was <90% for the Thr(373)Lys alpha(2A) AR compared with the WT alpha(2A) AR (9.1+/-1.4 pmol/mg protein); K(d) values were similar (1.16+/-0.19 nM and 1.51+/-0.15 nM, respectively). Forty-eight-hours' pre-treatment of cells with either 0.1 microM (+)-RX 811059, 1 microM atipamezole or 1 microM of the efficacious agonist d medetomidine increased the amount of [(3)H]RX 821002 binding sites of both WT (52%-59%) and mutant (306%-447%) Thr(373)Lys alpha(2A) ARs. The same alpha(2) AR ligands also prevented the loss of [(3)H]RX 821002 binding sites as induced by incubation of transfected CHO-K1 cellular membranes at 37 degrees C for 4 h (WT alpha(2A) AR) and 2 h (Thr(373)Lys alpha(2A) AR); 0.1 microM (+)-RX 811059 and 1 microM atipamezole caused an increase compared with the control amount of [(3)H]RX 821002 binding sites to the Thr(373)Lys alpha(2A) AR by 73% and 50%, respectively. In conclusion, no relationship was found between inverse agonism and alpha(2A) AR up-regulation. It is suggested that this is due to structural stabilisation of the alpha(2A) AR, irrespective of the nature of the ligand. PMID- 12122501 TI - Pharmacological characterisation of the somatostatin analogue TT-232: effects on neurogenic and non-neurogenic inflammation and neuropathic hyperalgesia. AB - The putative anti-inflammatory and anti-nociceptive activity of the heptapeptide somatostatin analogue TT-232 ( D-Phe-Cys-Tyr- D-Thr-Lys-Cys-Thr-NH(2)) was investigated in the rat and mouse, as well as its effect on neuropathic hyperalgesia, gastric ulceration and the release of sensory neuropeptides. In the rat, carrageenin-induced paw oedema was inhibited dose dependently by TT-232 (3x2.5-20 microg/kg i.v.). Evans blue accumulation induced by intraarticular bradykinin injection (0.5 nmol in 0.1 ml) was slightly, but significantly inhibited by a single TT-232 dose (5-20 microg/kg). Cutaneous neutrophil accumulation over a 3-h period after intradermal (i.d.) injection of carrageenin (1 mg/site) or interleukin 1beta (IL-1beta, 3 pmol/site) was inhibited significantly by TT-232 (3x80 microg/kg i.v.), while diclofenac (3x10 mg/kg i.v.) elicited significant inhibition only in the IL-1beta test. In the mouse, TT-232 potently decreased oedema formation induced by 2.5% capsaicin applied topically to the ear. Mechano-nociception in the rat hind-paw during neuropathic pain induced by partial sciatic nerve injury (model of Seltzer) was measured using the Randall-Selitto test. TT-232 (5-20 microg/kg i.p. on the 7th day after the operation) dose-dependently inhibited the mechano-nociceptive hyperalgesia. In vitro release of substance P (SP), calcitonin gene-related peptide (CGRP) and somatostatin from the isolated rat trachea in response to electrical field stimulation (40 V, 0.1 ms, 10 Hz, 120 s) of its nervous elements was inhibited significantly by 500 nM TT-232. The role of G protein-coupled receptors in the effect of TT-232 was indicated by the prevention of its inhibitory action on the release of sensory neuropeptides by incubation the tissue for 1 or 6 h with pertussis toxin (100 ng/ml). The release of sensory neuropeptides to in response to electrical nerve stimulation was not inhibited by a potent tyrosine kinase inhibitor, genistein (50 microM). TT-232 (up to 5 mg/kg i.p.) did not induce mucosal lesions in either the stomach or the duodenum. These data suggest that TT 232, a somatostatin analogue devoid of endocrine effects, is a promising lead molecule in the search for novel, broad-spectrum anti-inflammatory and analgesic agents. PMID- 12122502 TI - Uridine nucleotide-induced stimulation of gluconeogenesis in isolated rat proximal tubules. AB - Uridine nucleotides, released into the extracellular environment, influence a variety of metabolic and other cellular activities in a wide range of target tissues. Here we have studied the effects of uridine nucleotides on gluconeogenesis in isolated rat proximal tubules. Gluconeogenesis, from a range of precursors, was stimulated following exposure of isolated proximal tubules to either UTP or UTPgammaS, but not when exposed to other uridine-containing nucleotides. UTP- and UTPgammaS-induced gluconeogenesis was diminished in the presence of purinoceptor antagonists (e.g. suramin, PPADS) indicative of a role for P2Y(2)-like purinoceptors in these effects. Likewise, agents that interfere with either phospholipase C activation or intracellular Ca2+ mobilization decreased UTP- and UTPgammaS-induced stimulation of gluconeogenesis consistent with a role for these secondary messenger systems in the mechanism of action of extracellular UTP and UTPgammaS on proximal tubule metabolism. PMID- 12122503 TI - Block of volume-regulated anion channels by selective serotonin reuptake inhibitors. AB - We have used the whole-cell patch clamp technique to study the effects of the commonly used antidepressants sertraline, paroxetine, citalopram and fluvoxamine on the volume-regulated anion channel (VRAC) in endothelial cells. It was the purpose of the present experiments to investigate whether VRAC block is a general property of this group of selective serotonin reuptake inhibitors (SSRIs). At pH 7.4, all SSRIs induced a fast and reversible block of the volume-sensitive chloride current ( I(Cl,swell)), with an IC(50) value of 2.1+/-0.5 microM for sertraline, 2.7+/-0.2 microM for paroxetine, 12.3+/-1.4 microM for fluvoxamine and 27.7+/-2.8 microM for citalopram. The block was enhanced at more alkaline pH, indicating that it is mediated by the uncharged form. This study describes the effects of a variety of SSRIs on an anion channel. Our data reveal a potent block and suggest a hydrophobic interaction of high affinity between the uncharged SSRI and volume-regulated anion channels. We conclude that VRAC block is a general property of this pharmacological class of selective serotonin reuptake inhibitors. PMID- 12122504 TI - The effect of castration on endothelins, their receptors and endothelin converting enzyme in rat prostate. AB - We previously have shown that experimental diabetes in rats causes prostatic involution, reduces serum testosterone levels, and causes an upregulation in prostatic endothelin (ET) receptors. Furthermore, insulin treatment normalizes these changes (Saito et al., Mol Cell Biochem 210:1-12, 2000). Since experimental diabetes-induced reduction in serum testosterone may be a factor in the alteration of the ET receptors and of prostatic growth, we investigated the effect of castration, another means of involuting the prostate and decreasing serum testosterone levels, on the expression of ET receptors in ventral and dorsolateral regions of the rat prostate.Three-month-old Sprague-Dawley rats were surgically castrated or sham operated, and then killed on the 7th post-operative day. Biochemical and pharmacological properties, and localization of ET receptors in the rat prostate, were determined by performing a series of binding experiments with [(125)I]ET-1 and by light microscopy autoradiography, respectively. The expression levels of ET-1, ET-3, ET receptor subtypes and endothelin converting enzyme-1 (ECE-1) mRNAs were assessed by relative multiplex reverse transcription polymerase chain reaction (RT-PCR). The total density of ET receptors increases 3.7-fold in the ventral and 2.1-fold in the dorsolateral regions of the castrated rat prostate compared to sham operated animals. Castration causes a 2.4-fold increase in the density of alpha(1)-adrenoceptors (alpha(1)-ARs) in the ventral region of the prostate, but no change in the density of alpha(1)-ARs in the dorsolateral region of the rat prostate. The predominant ET receptor subtype in the rat prostate is the ETA subtype, which is mainly located in the prostatic stroma. In addition, RT-PCR data show an upregulation in the expression of ETB receptor subtype, ET-1 and ECE-1 mRNA in both regions, and a downregulation in the expression of ETA receptor subtype mRNA in the dorsolateral region of the castrated rat prostate. There is no change in the expression of ET-3 mRNA in either region. Castration does not cause significant changes in the pharmacological properties of prostatic ET receptors, i.e., the predominance of ETA receptors in either region of the prostate, or the expression of ETA receptor subtype mRNA in the ventral region of the castrated rat prostate. These results suggest the existence of a region/lobe-specific regulatory role for testosterone in the expression of the ET receptor system in the rat prostate. PMID- 12122505 TI - MC(3) receptors are involved in the protective effect of melanocortins in myocardial ischemia/reperfusion-induced arrhythmias. AB - Myocardial ischemia/reperfusion induces ventricular tachycardia (VT), ventricular fibrillation (VF) and a high degree of lethality. Since ACTH-(1-24) (adrenocorticotropin) protects against such injuries in rats, we investigated which melanocortin MC receptor is involved. Ischemia was produced in anesthetized rats by ligature of the left anterior descending coronary artery (5 min), and reperfusion-induced VT, VF, lethality and time-course of arterial blood pressure within the 5 min following reperfusion were evaluated. I.v. administration of the selective MC(3) receptor agonist gamma(1)-melanocyte-stimulating hormone (gamma(1)-MSH), as well as of an equimolar dose (162 nmol/kg) of both the non selective agonist ACTH-(1-24) and alpha-MSH, significantly prevented VT and VF, and increased survival. Coronary reperfusion was followed by an abrupt and massive fall in mean arterial pressure and pulse pressure, in saline-treated rats. Treatment either with ACTH-(1-24) or gamma(1)-MSH completely prevented such fall. The protective effect of ACTH-(1-24) against the occurrence of VT, VF and lethality was neither affected by adrenalectomy, nor by i.v. pretreatment with the selective MC(4) receptor antagonist HS014 and the MC(4)-MC(5) antagonist HS059. On the other hand, the MC(3)-MC(4) receptor antagonist SHU 9119 prevented such protective effect. Moreover, the selective MC(1) receptor agonist MS05 (162 nmol/kg i.v.) failed to reduce the incidence of arrhythmias and lethality. These data demonstrate that MC(3) receptors mediate the protective effect of melanocortins in myocardial ischemia/reperfusion-induced arrhythmias, in rats. PMID- 12122506 TI - Effect of zaleplon on learning and memory in rats. AB - Although structurally not a benzodiazepine, 3'-(3-cyanopyrazolo [1,5-a] pyrimidin 7-yl)- N-ethylacetamide (zaleplon) it acts via the benzodiazepine site of the GABA(A) receptor. In the present study, we investigated the effects of zaleplon on learning and memory in rats in comparison with triazolam and nitrazepam. Oral administration of zaleplon and the reference drugs dose-dependently lessened the step-through latency in the test session of a passive avoidance task and increased the latency for reaching the hidden platform in the Morris water maze task, indicating the amnesic effect of the test drugs. The amnesic liability ratio for zaleplon in the passive avoidance task to sleep inducing activity was 19.6, for triazolam and nitrazepam 4.2 and 5.9, respectively. The liability ratios derived from the Morris water maze task for zaleplon, triazolam and nitrazepam were 10.2, 0.9 and 0.6, respectively. The results may indicate that zaleplon has a preferential sedative effect and that the sedative dose does not interfere with learning and memory. In a binding study, zaleplon displaced bound [(3)H]flunitrazepam from membrane preparations from the rat hippocampus with an IC(50) of 4,454.5 nM. In contrast, triazolam and nitrazepam displaced the binding of [(3)H]flunitrazepam to the membrane with IC(50) values of 15.5 nM and 83.6 nM, respectively. The efficacy of zaleplon for the competitive inhibition of [(3)H]flunitrazepam binding to the membrane preparation from hippocampus was thus less than that of triazolam and nitrazepam. These results suggest that zaleplon is characterized by a reduced amnesic liability, which may be due to its low affinity for the benzodiazepine site of the GABA(A) receptor in the hippocampus. PMID- 12122507 TI - A mitogen-activated protein kinase is involved in the inotropic but not chronotropic actions of adrenoceptor agonists and endothelin-1. AB - The activation of mitogen-activated protein kinase (MAPK) pathways in the heart, for instance by alpha(1)-adrenoceptor agonists and endothelin-1, has primarily been associated with cellular growth regulation. Here we have investigated a possible role of MAPK pathways in the inotropic and chronotropic effects of adrenoceptor agonists and endothelin-1 in isolated rat left and right atria. Inotropic and chronotropic responses of the isolated atria to methoxamine, isoprenaline and endothelin-1 were measured in the absence and presence of inhibitors of MAPK pathways. The MAPK kinase (MKK(mek)) inhibitors PD98059 (100 microM) and U0126 (10 microM) significantly inhibited the inotropic responses to the alpha(1)-adrenoceptor agonist methoxamine (300 microM) and endothelin-1 (50 nM), but not the chronotropic responses to these agonists. U0126 but not PD98059 inhibited the inotropic response to 3 microM isoprenaline. None of the aforementioned inotropic and chronotropic effects were inhibited by the MAPKP(p38) inhibitor SB203580 (2 microM). We conclude that activation of the PD98059/U0126-sensitive MAPK pathway is essential for the inotropic but not chronotropic actions of adrenoceptor agonists and endothelin-1. PMID- 12122508 TI - Selective bowel decontamination in elective liver transplantation: no improvement in endotoxaemia, initial graft function and post-operative morbidity. AB - Peri-operative endotoxaemia during liver transplantation has been linked to compromised graft function and infection. Selective decontamination of the digestive tract (SDD) could prevent endotoxaemia by eradicating Gram-negative bacteria from the intestine. In a randomized placebo controlled study we investigated the effects of endotoxaemia and the efficacy of SDD to prevent its occurrence. Thirty-one patients undergoing elective orthotopic liver transplantation received either SDD ( n=15) or placebo ( n=16), which was started at least 7 days before transplantation. Endotoxin levels were measured in blood peroperatively. Patients were scored daily for signs of liver dysfunction and infection. Endotoxaemia was neither associated with initial poor function nor any routine liver function test. Infections were more prominent in patients without endotoxaemia. SDD did not prevent endotoxaemia. Endotoxaemia does not affect post operative graft function or the incidence of post-operative infections. SDD cannot prevent peri-operative endotoxaemia. Translocation of endotoxin may not be relevant in liver transplantation. PMID- 12122509 TI - sTNF-RII: is it useful for the early diagnosis of rejection and for prognosis after renal transplantation? AB - Changes in soluble tumour necrosis factor receptor II (sTNF-RII) correlate with transplant rejection, and it increases in the course of sepsis. These changes might help to identify rejection early, and thus lead to more effective treatment. Serum and urine sTNF-RII levels were measured in 70 patients during the first 3 weeks after kidney transplantation and correlated with clinical and laboratory findings. Retrospectively, three groups were identified: I. stable transplant function ( n=23), II. at least one rejection episode ( n=38) and III. other complications (infection or reperfusion injury) ( n=9). The pre-operative maximum for serum sTNF-RII was 22.4 +/- 10.7 ng/ml. In group I it decreased to 9.5 +/- 6.7 ng/ml on day 6 after transplantation ( P<0.01), while in group II sTNF-RII serum levels were significantly higher on day 6 (24.9 +/- 15.0 ng/ml, P<0.01). High levels of sTNF-RII in serum (>40 ng/ml for at least 2 days) predicted a higher risk of an unfavourable outcome. High serum levels of sTNF-RII are not specific but seem to be a prognostic indicator of a complicated course; sTNF-RII in urine has no diagnostic value. PMID- 12122511 TI - A preservation solution with polyethylene glycol and calcium: a possible multiorgan liquid. AB - The addition of polyethylene glycol (PEG) to hepatocyte storage medium is known to decrease lipid peroxidation and swelling and to protect the cell cytoskeleton from cold. We therefore decided to investigate the effect of substituting PEG for hydroxyethyl starch (HES) in an extracellular-like UW solution, with and without Ca++, on rat liver preservation. Isolated perfused rat livers were used to assess graft injury after 24h of cold storage. Four groups of preserved livers ( n=6 for each group) were compared to controls (non preserved livers, n=11). For this purpose, Belzer solution (K+-UW, group 1) was stepwise modified. Group 2 (Na+-UW) was treated with the same liquid, however with inverted concentrations of Na+ and K+. Group 3 was preserved in the first experimental solution (EPS-1) with Ca++ (0.5mM) added to the Na+-UW solution. In the EPS-2 (group 4), PEG-35 (0.03mM) was substituted for HES. The last group, EPS-3 (group 5) was treated with the same compounds as EPS-2, but without Ca++. After 24h of cold storage and 120min normothermic reperfusion, there was no statistical difference in transaminases (ALT and AST) release between the control and the Na+-UW groups. Furthermore, rat livers preserved in Na+-UW solution released less ( P<0.05) ALT and AST and excreted more ( P<0.05) indocyanine green (ICG) than livers preserved in K+-UW solution. The addition of 0.5mM Ca++ to Na+-UW solution (EPS-1) dramatically increased ( P<0.05) parenchymal (ALT, AST) and non parenchymal (creatine kinase BB) cellular injury. The substitution of PEG (0.03mM) for HES (EPS-2) reduced ( P<0.05) membrane injuries due to Ca++ while bile flow was statistically increased ( P<0.05). Finally, the omission of Ca++ from EPS-2, that is EPS-3, has no statistically significant effect on the studied parameters. PEG effectively protected the rat liver grafts from the onset of hypothermic ischemia-reperfusion and Ca++ damages and thus may be a valuable additive to preservation solutions. PMID- 12122510 TI - Antifungal prophylaxis in liver transplant recipients: a randomized placebo controlled study. AB - The aim of this study was to evaluate the efficacy of two antifungal prophylaxis regimens in liver transplant recipients. One hundred and twenty-nine consecutive recipients were randomized to receive sequential treatment with intravenous liposomal amphotericin B + oral itraconazole, intravenous fluconazole + oral itraconazole, or intravenous and oral placebo. Frequency and incidence of mycotic colonization, local and systemic infection of mycotic origin, causes of death, and possible risk factors for mycotic infection were evaluated. The incidence of mycotic colonization was higher in the placebo group ( P<0.01), but there was no significant difference in the incidence of infection between the three groups. Pre-transplant colonization, severity of liver disease, and graft rejection were all risk factors for the development of fungal infection. The routine use of antifungal prophylaxis for all liver transplant recipients does not seem to be justified. PMID- 12122512 TI - Hemodynamic interaction between portal vein and hepatic artery flow in small-for size split liver transplantation. AB - In split-liver transplantation, the entire portal flow is redirected through relatively small-for-size grafts. It has been postulated that excessive portal blood flow leads to graft injury. In order to elucidate the mechanisms of this injury, we studied the hemodynamic interactions between portal vein- and hepatic artery flow in an experimental model in pigs. Six whole pig liver grafts were implanted in Group 1 ( n=6) and six whole liver grafts were split into right and left grafts and transplanted to Groups 2 ( n=6) and 3 ( n=6), respectively. The graft-to-recipient liver volume ratio was 1:1, 2:3 and 1:3 in Groups 1, 2 and 3, respectively. Portal vein- and hepatic artery flows were measured with an ultrasonic flow meter at 60,120 and 180 min after graft reperfusion. Portal vein pressure was also recorded at the same time intervals. Graft function was assessed at 3,6h and 12h, and morphological changes at 12h after reperfusion. Following reperfusion, portal vein flow showed an inverse relationship to graft size, while hepatic artery flow was reduced proportionately to graft size. The difference was significant among the three groups ( P<0.05). Portal vein pressure was significantly higher in group 3, compared to groups 1 and 2 ( P<0.05). Hepatic artery buffer response was significantly higher in Group 3, compared to Groups 1 and 2 in relation to pre-occlusion values ( P<0.05). Split-liver transplantation, when resulting in small-for-size grafts, is associated with portal hypertension, diminished arterial flow, and graft dysfunction. Arterial flow impairment appears to be related to increased portal vein flow. PMID- 12122513 TI - Absence of PERV specific humoral immune response in baboons after transplantation of porcine cells or organs. AB - Xenotransplantation of pig organs seems a promising way of overcoming the prevailing limitation on allotransplantation due to donor numbers. However, as porcine endogenous retroviruses (PERVs) can infect human cells in vitro, there is substantial concern regarding the risk of a PERV infection in xenogeneic transplant recipients. Cultured porcine endothelial cells, stimulated peripheral blood mononuclear cells, and pancreatic islet cells can release PERV infectious for human cells in vitro, but it is currently unknown whether PERV is released in vivo, whether these viral particles can infect the transplant recipient, and whether they are pathogenic. In a retrospective study 15 immunosuppressed baboons were tested for a specific immune response against PERV after transplantation of porcine endothelial cells, mononuclear blood cells, and lungs. Anti-PERV antibody expression was analyzed with peptide-based, enzyme-linked immunosorbent assays and highly sensitive Western Blot assays. This xenotransplantation study using nonhuman primates found no evidence of PERV specific humoral immune response. Our data suggest that no productive PERV infection and no continuous PERV release takes place in the nonhuman primates analyzed in this study. PMID- 12122514 TI - Auxiliary partial orthotopic liver transplantation in acute liver failure due to hepatitis B. AB - Auxiliary partial orthotopic liver transplantation is an alternative therapeutic modality in acute liver failure, wherein the capacity of native liver to regenerate is preserved. A case of acute liver failure due to hepatitis B in an 18-year-old male patient treated with an auxiliary left lateral segment graft is described. There was no recurrence of hepatitis B in the auxiliary graft and the patient cleared the virus after 9 months whilst receiving lamivudine. Immunosuppression was withdrawn at 14 months, and the auxiliary graft atrophied secondary to hepatic arterial conduit thrombosis, possibly precipitated by immunosuppression withdrawal. The native liver regenerated completely, and the patient is well and off immunosuppressive and antiviral therapy 3 years after transplantation. Auxiliary partial orthotopic liver transplantation is an attractive treatment option in acute liver failure due to hepatitis B infection and allows a life free of long-term immunosuppression. PMID- 12122516 TI - Bleeding complications precipitated by unrecognized Gingko biloba use after liver transplantation. AB - Because of its neurocognitive enhancing effects, Gingko biloba has emerged as amongst the most commonly used herbal products. We report a liver transplant recipient with potentially life-threatening toxicity resulting from Gingko biloba use. Seven days after a second liver transplantation for recurrent hepatitisB virus infection, subphrenic hematoma was documented in a 59-year-old Korean patient. Failure to control bleeding with CT-guided drainage necessitated exploratory laparotomy for the evacuation of a large subphrenic hematoma. Three weeks later, an episode of vitreous hemorrhage was documented. Unbeknownst to his care providers, the patient had been consuming Gingko biloba throughout the postoperative period. No further bleeding episodes occurred after the cessation of Gingko biloba use. Unrecognized use of herbal products may be associated with serious side effects and adverse clinical sequelae in transplant recipients. Given their increasing popularity, the use of herbal products should be routinely sought as part of the history in transplant recipients. PMID- 12122515 TI - Colchicine myoneuropathy in a renal transplant patient. AB - Colchicine is widely employed for the treatment of gout in renal transplant patients where NSAIDs are contra-indicated and allopurinol prophylaxis is often avoided due to concomitant azathioprine immunosuppression. We report here a case of colchicine-induced myoneuropathy in a renal transplant recipient. Our patient had myalgia, muscle weakness, elevated creatine kinase levels, myopathic changes on electromyography and peripheral neuropathy. Withdrawal of colchicine resulted in recovery within 4 weeks. Renal transplant recipients are likely to be at greater risk of colchicine-induced myoneuropathy due to the unique concurrence of risk factors predisposing to toxicity in such patients. These risk factors include the high incidence of gout in this population, widespread use of colchicine as first-line therapy, impaired renal function and concomitant cyclosporin treatment. The diagnosis should be considered in any renal transplant recipient receiving the drug who develops myopathy. Prompt withdrawal of colchicine therapy should result in rapid clinical and biochemical improvement. PMID- 12122517 TI - Successful outcome of liver transplantation in a patient with hepatitis C and common variable immune deficiency. AB - A 43-year-old man with common variable immune deficiency underwent liver transplantation for cirrhosis caused by hepatitis C virus (HCV). HCV had been acquired from a contaminated batch of immunoglobulin. He developed cirrhosis within 3 years of infection with the virus, then liver failure requiring liver transplantation. The immediate post-transplant course was uncomplicated. Five months after transplantation he developed liver failure, and the histological appearances were those of severe cholestatic hepatitis. Withdrawal of immunosuppression resulted in recovery from liver failure. Clearance of the HCV from serum was also observed and has been sustained during follow-up (despite the subsequent reintroduction of low-dose immunosuppression). The patient is alive and well more than 5 years after transplantation. His post-transplant course has been remarkable for the aggressive recurrence then clearance of the HCV. PMID- 12122518 TI - Clinical implications of antibiotic-induced endotoxin release in septic shock. AB - Antibiotic-induced release of bacterial cell wall components can have immediate adverse effects for the patient. This article reviews the data on endotoxin release after initiation of antibiotic therapy and its role in the pathogenesis of sepsis and septic shock. Antibiotics differ in their potential to liberate endotoxins from bacterial cell walls. When used for treatment of systemic Gram negative infection, some classes of beta-lactam antibiotics lead to markedly increased levels of free endotoxins while treatment with carbapenems and aminoglycosides produces relatively low amounts of endotoxins. Antibiotics that induce the formation of long, aberrant bacterial cells before effectively killing the microorganisms show the highest degree of endotoxin liberation. There is increasing evidence from animal models and clinical studies of sepsis that the antibiotic-mediated release of biologically active cell wall components derived from Gram-positive, Gram-negative or fungal organisms is associated with a rapid clinical deterioration. PMID- 12122519 TI - Kinetic and reversibility of mechanical ventilation-associated pulmonary and systemic inflammatory response in patients with acute lung injury. AB - OBJECTIVE: To investigate the kinetic and reversibility of mechanical ventilation associated pulmonary and systemic inflammatory response in patients with acute lung injury (ALI). DESIGN: Prospective observational cross-over study. SETTING: Intensive care unit of a university hospital. PATIENTS: Twelve mechanically ventilated patients with ALI. INTERVENTIONS: Mechanical ventilation was transiently changed from a lung protective setting with PEEP of 15 cmH(2)O and a V(T) of 5 ml/kg predicted body weight to a more conventional ventilatory setting with PEEP of 5 cmH(2)O and V(T) of 12 ml/kg predicted body weight for a period of 6 h. MEASUREMENTS AND RESULTS: We examined the profile of interleukin (IL)-1beta, IL-1 receptor antagonist, IL-6, IL-10, and tumor necrosis factor in the plasma of all patients, and in the bronchoalveolar lavage (mini-BAL) fluid of six of these patients. Measurements were performed at baseline, 1 h, and 6 h after each change of the ventilatory setting. Switching to conventional mechanical ventilation was associated with a higher PaO(2) ( P < 0.05) and a marked increase ( P < 0.05) of measured plasma cytokines in patients with and without mini-BAL with a maximum after 1 h. Similarly, intraalveolar cytokine concentrations increased with conventional mechanical ventilation. While plasma cytokine levels returned to baseline values, intraalveolar cytokine concentrations further increased when lung protective mechanical ventilation was reestablished. CONCLUSIONS: In patients with ALI, initiation of low PEEP and high V(T) mechanical ventilation is associated with cytokine release into circulation which occurred within 1 h. It is independent from BAL procedures and can be reversed by reinstitution of lung protective mechanical ventilation. PMID- 12122520 TI - The upper inflection point of the pressure-volume curve. Influence of methodology and of different modes of ventilation. AB - OBJECTIVE: The pressure-volume (P/V) curve has been proposed as a tool to adjust the ventilatory settings in cases of acute respiratory distress syndrome (ARDS). The aim of this study was to test the influence of P/V tracing methodology on the presence and value of the upper inflection point (UIP). METHODS: In 13 medical ARDS patients, the interruption and the automated low flow inflation methods were compared while the patients were ventilated at conventional (10-12 ml/kg) and at low (5-6 ml/kg) tidal volume (Vt). Two levels of inspiratory flow and insufflation time were used (3 and 6 s). RESULTS: No significant difference in UIP was found between the static and the dynamic methods, whatever the flow used. At Vt 10-12 ml/kg, the static and dynamic UIPs were 22.4 +/- 4.4 cmH(2)O and 22.1 +/- 4.5 cmH(2)O ( p = 0.86), respectively; at Vt of 5-6 ml/kg, the static and dynamic UIPs were 26.6 +/- 4.1 cmH(2)O and 25.5 +/- 5 cmH(2)O ( p = 0.34), respectively. Significant differences in UIP were found, in the static and dynamic conditions, between the two levels of Vt ( p < 0.005): it was lower with the higher Vt, suggesting that UIP is dependent on previous tidal alveolar recruitment. CONCLUSION: Interruption and continuous flow techniques gave similar results, but the previous Vt influences the pressure value of the UIP. PMID- 12122521 TI - Chest physiotherapy for the prevention of ventilator-associated pneumonia. AB - OBJECTIVE: Pneumonia is an important complication in patients who are intubated and mechanically ventilated, when it is commonly referred to as ventilator associated pneumonia (VAP). Since VAP may be contributed to by impaired sputum clearance, we studied whether chest physiotherapy designed to enhance sputum clearance decreases the occurrence of VAP. DESIGN: Prospective controlled systematic allocation trial. SETTING: Tertiary teaching hospital ICU. PATIENTS AND PARTICIPANTS: Sixty adult patients intubated and mechanically ventilated for at least 48 h. INTERVENTIONS: Chest physiotherapy (intervention group) or sham physiotherapy (control group). MEASUREMENTS AND RESULTS: Control and intervention groups were well matched for age, sex, and admission PaO(2)/FiO(2) ratio, APACHE II score, and Glasgow Coma Score. There were no differences in the duration of mechanical ventilation, length of stay in ICU or mortality. VAP was assessed daily by combined clinical assessment and the clinical pulmonary infection score (CPIS). VAP occurred in 39% (14/36) of the control group and 8% (2/24) of the intervention group (OR = 0.14, 95% CI 0.03 to 0.56, P = 0.02). After adjustment was made by logistic regression for other important variables (APACHE II score, duration of mechanical ventilation, presence of tracheostomy, and GCS score), chest physiotherapy was independently associated with a reduced occurrence of VAP (adjusted OR = 0.16, 95% CI 0.03 to 0.94, P = 0.02). CONCLUSIONS: In this small trial, chest physiotherapy in ventilated patients was independently associated with a reduction in VAP. This suggested benefit of physiotherapy in prevention of VAP requires confirmation with a larger randomised controlled trial. PMID- 12122522 TI - Reproducibility of protected specimen brush and bronchoalveolar lavage conserved at 4 degrees C for 48 hours. AB - OBJECTIVE: Protected specimen brush (PSB) and bronchoalveolar lavage (BAL) are proposed in combination to optimize antimicrobial treatment. Nevertheless, they are only validated for immediate laboratory processing. This study was therefore conducted to determine whether 48 h conservation at a mere 4 degrees C enables good culture reproducibility for both PSB and BAL. DESIGN AND SETTING: Prospective study, evaluation of a conservation procedure for PSB and BAL, from February 1994 to February 1995, in the 12 bed ICU of a general hospital (938 beds). SAMPLES: Ninety-nine PSB and 86 BAL samples, obtained from 100 bronchoscopic procedures, were analyzed. Thresholds were 10(3) and 10(4) cfu/ml for PSB and BAL, respectively. MEASUREMENTS AND RESULTS: Qualitative comparison between the immediate and 48 h procedures were, for PSB, specificity 100%, sensitivity 78%, positive predictive value 100%, negative predictive value 84% and overall accuracy 90%; and for BAL: 100%, 89%, 100%, 89% and 94%. Lowered 10(2) and 10(3) cfu/ml thresholds at the 48 h procedure for PSB and BAL reduce the false negatives from 10 to 3 and 5 to 1, respectively. Microorganism results were comparable for PSB and BAL ( r = 0.63 and 0.67), especially for the most resistant strains: Staphylococcus, Enterobacteriaceae and Pseudomonas. However, there was a decrease in the Neisseria and Haemophilus group ( p < 0.01). CONCLUSION: There is a good culture reproducibility for both PSB and BAL after 48 h conservation at 4 degrees C, especially with lowered thresholds; this technique is therefore appropriate for routine use. PMID- 12122523 TI - The strong ion gap does not have prognostic value in critically ill patients in a mixed medical/surgical adult ICU. AB - OBJECTIVE: To examine whether the strong ion gap (SIG) or standard base excess corrected for abnormalities of serum chloride and albumin (BE(UA)) can predict outcome and to compare the prognostic abilities of these variables with standard base excess (SBE), anion gap (AG), pH, and lactate, the more traditional markers of acid-base disturbance. DESIGN: Prospective, observational study. SETTING: University teaching hospital, general adult ICU. PATIENTS: One hundred consecutive patients on admission to the ICU. MEASUREMENTS AND RESULTS: The anion gap (AG) was calculated and corrected for abnormal serum albumin (AG(corrected)). Serum lactate was measured and SBE, BE(UA), SIG, and APACHE II scores calculated for each patient. 28-day survival was recorded. There was a significant difference between the mean APACHE II (P < 0.001), SBE (P < 0.001), lactate (P = 0.008), AG (P = 0.007), pH (P < 0.001), and BE(UA) (P = 0.009) of survivors and non-survivors. There was no significant difference between the mean SIG (P = 0.088), SIDeff (P = 0.025), and SID app (P = 0.254) between survivors and non survivors. The pH and SBE demonstrated the best ability of the acid-base variables to predict outcome (AUROC curves 0.72 and 0.71, respectively). Neither of these were as good as the APACHE II score (AUROC 0.76) CONCLUSION: Traditional indices of SBE, BE(UA,) lactate, pH, AG, and APACHE II all discriminated well between survivors and non-survivors. In this group of patients the SIG, SIDeff, and SIGapp appear to offer no advantage in prediction of outcome and their use as prognostic markers can therefore not be advocated. PMID- 12122524 TI - Comparison between the Datex-Ohmeda M-COVX metabolic monitor and the Deltatrac II in mechanically ventilated patients. AB - OBJECTIVE: To compare the M-COVX and the Deltatrac II metabolic monitors under clinical conditions. DESIGN: Prospective clinical comparison. SETTING: A general Intensive Care Unit of a university hospital. PATIENTS: Twenty mechanically ventilated critically ill patients. INTERVENTIONS: The monitors were compared at FiO(2) 0.3, 0.5, and 0.7 in each patient where possible. MEASUREMENTS AND RESULTS: Pulmonary gas exchange measurements were recorded using the two monitors sequentially (Deltatrac(before), M-COVX, Deltatrac(after)). Each measurement consisted of five consecutive 1-min readings of VO(2) and VCO(2). We compared the Deltatrac(before) with the Deltatrac(after) and the mean of the Deltatrac with the M-COVX. There was no clinically significant bias between the two monitors for VO(2) or VCO(2) but the limits of agreement (LOA) were wide (bias +/-95% LOA: VCO(2) -13 +/- 30 ml/min, -8 +/- 36 ml/min, 7 +/- 50 ml/min; VO(2) -7 +/- 50 ml/min, -5 +/- 56 ml/min, 6 +/- 64 ml/min, at FiO(2) 0.3, 0.5, and 0.7, respectively). The Deltatrac before and after measurements displayed good agreement for VCO(2) but poorer agreement for VO(2) (bias +/- 95% LOA: VCO(2) 0 +/- 18 ml/min, -6 +/- 16 ml/min, -1 +/- 12 ml/min; VO(2) 2 +/- 12 ml/min, 3 +/- 38 ml/min, 10 +/- 42 ml/min, at FiO(2) 0.3, 0.5, and 0.7, respectively). Using within-patient standard deviation as a measure of reproducibility suggested that for VO(2) the M-COVX performed better than the Deltatrac at high FiO(2), and for VCO(2) Deltatrac was better at lower FiO(2). CONCLUSIONS: The M-COVX is a suitable integrated device for measuring metabolic gas exchange in ventilated patients. PMID- 12122525 TI - Low-dose dopamine: a systematic review. AB - OBJECTIVE: To determine the magnitude of the treatment effect of low-dose dopamine on renal function in patients at risk of and in patients with early renal injury. DATA SOURCES: MEDLINE, citation review of relevant primary and review articles, personal files, and contact with expert informants. STUDY SELECTION: Randomized controlled studies that compared low-dose dopamine with placebo for the prevention or treatment of acute renal dysfunction. From 122 articles screened, 21 met the inclusion criteria for this meta-analysis. Of these six, were excluded. DATA EXTRACTION: Fifteen studies containing 970 subjects were analyzed. Descriptive and outcome data were extracted. The main outcome measure was the absolute change in serum creatinine. In addition the number of patients who developed an acute decline in renal function was recorded. The meta-analysis was performed using the random effects model. DATA SYNTHESIS: The meta-analysis demonstrated no significant difference between the absolute change in serum creatinine (5.1 micromol/l, 95% CI of -6.5 to +16.7) and the incidence of acute renal dysfunction (31% vs 33%, relative risk 1.01, 95% CI of 0.79-1.28) between those patients receiving low-dose dopamine and the control group. In addition, no sub-group of patients showed improved renal function with low-dose dopamine. CONCLUSIONS: The results of this meta-analysis confirms that low-dose dopamine has no reno-protective effect. Considering the potential side-effects of dopamine this agent should not be used for this indication. PMID- 12122526 TI - A randomised controlled trial investigating the effects of dopexamine on gastrointestinal function and organ dysfunction in the critically ill. AB - OBJECTIVE: To determine whether an infusion of dopexamine for up to 7 days has an effect on gastrointestinal (GIT) absorption and permeability, renal function or organ dysfunction in the critically ill. DESIGN AND SETTING: Prospective, randomised controlled clinical trial in two general adult intensive care units. PATIENTS: 102 critically ill adult patients predicted to require organ support for at least 4 days. INTERVENTIONS: After resuscitation patients were randomly assigned to receive an infusion of up to 2 mcg/kg/min [corrected] per minute of dopexamine or control. MEASUREMENTS AND RESULTS: GIT absorption and permeability were measured using the ratio of absorbed rhamnose to 3- O-methyl- D-glucose and the ratio of lactulose to rhamnose on days 1, 4 and 7. Creatinine clearance was measured concurrently. Daily Sequential Organ Failure Assessment scores were calculated. Fifty-two patients received dopexamine. No significant difference between the two groups emerged on any of the measured parameters during the study period. CONCLUSIONS: No benefit was seen from a prolonged infusion of dopexamine in this group of critically ill patients in terms of GIT absorption and permeability, creatinine clearance or organ dysfunction. PMID- 12122527 TI - The effect of propofol on airway pressures generated by magnetic stimulation of the phrenic nerves. AB - OBJECTIVES: To assess the effect of propofol on the change in airway pressure produced by diaphragmatic contraction. DESIGN AND SETTING: Prospective, controlled study in patients anaesthetised with propofol in a university hospital. PATIENTS AND METHODS: We stimulated the phrenic nerves before and immediately after induction of anaesthesia in 11 subjects, using a pair of 43-mm mean diameter double magnetic coils and measured the change in airway pressure at the mouth (TwPmo) produced by the resulting diaphragmatic contraction. Supramaximality of stimulation was confirmed with electromyogram and pressure measurements. We recorded the change in Resting End Expiratory Position (REEP) using a spirometer. We applied an approximate correction for the effect of lung volume on the amplitude of twitch pressure produced by diaphragmatic contraction. INTERVENTION: Following the initial stimulations, the patients were anaesthetised with a propofol infusion. Once stable, repeat measurements were made. MEASUREMENTS AND RESULTS: Following induction, REEP fell by mean 0.3 l standard deviation (SD) 0.2 l. TwPmo fell by mean 14.2% SD 14.0% ( P = 0.01), mean 22.3% SD 11.7% corrected ( P < 0.001). Twitch transdiaphragmatic pressure fell by 18.1% and 20.0% (25.8% and 27.7% corrected) in two further subjects studied with oesophageal and gastric balloon catheters. CONCLUSION: Propofol does reduce the effectiveness with which diaphragmatic contraction produces changes of pressure in the airway. PMID- 12122528 TI - Quality of life after intensive care--evaluation with EQ-5D questionnaire. AB - OBJECTIVE: To evaluate health-related quality of life (HR-QOL) and study its determinants in adult patients discharged from an intensive care unit (ICU). DESIGN: Cohort study. SETTING: Intensive care unit (ICU), tertiary care hospital, Oporto, Portugal. PATIENTS: Of all the patients discharged over a 2 year period, 355 were considered eligible and 275 completed the study. MEASUREMENTS AND RESULTS: Patients were interviewed 6 months after ICU discharge using EuroQol 5-D (EQ-5D). At the interview only 29% reported feeling worse than 6 months before ICU admission. The proportions of those reporting moderate to extreme problems in the five dimensions studied were as follows: mobility (37%), self-care (22%), usual activities (46%), pain/discomfort (45%) and anxiety/depression (54%). Although 77% of patients reported a problem in at least one dimension, 44% referred to no problems or only moderate problems regarding pain or anxiety. EQ visual analogue scale (VAS) and EQ Index medians were 60 and 81, respectively. CONCLUSIONS: Intensive care unit variables (e.g., diagnosis, length of stay and severity of disease) and patient's background data (e.g., age, gender, education, main activity, smoking habits, experience with serious illness and previous health status) may be significant determinants of HR-QOL. However, when adjusted for background data, most ICU variables are no longer associated with EQ-5D. This should cause attention to be paid to the role of a patient's background in the evaluation of HR-QOL and to a careful interpretation of EQ-5D results when comparing ICUs. PMID- 12122529 TI - Survival of mechanically ventilated patients admitted to a specialised weaning centre. AB - OBJECTIVE: Hospital mortality and survival rates of long-term ventilated patients.DESIGN. Retrospective cohort study. SETTING: Specialised national weaning centre. INTERVENTION: Protocol-directed liberation from ventilator. PATIENTS: Four hundred three of 640 patients with prolonged mechanical ventilation (MV) who were admitted to our respiratory intensive care unit (RICU) were studied. MV lasted longer than 2 weeks and patients had failed more than two weaning trials in the referring ICUs. The majority of patients (59.3%) had chronic obstructive pulmonary disease (COPD). RESULTS: After a mean duration of 41 days of MV prior to transfer, 68% of patients were liberated from the ventilator. In total, 98 of 403 patients (24.3%) died during the stay in our hospital, 305 patients (75.7%) were discharged. Compared to the non-survivors, the survivors were characterised by younger age, longer length of stay in our RICU, lower severity of illness scores at admission, fewer cardiac illnesses and a higher rate of weaning success. In 31.5% of the discharged patients non invasive MV (NIV) was initiated during the stay at our unit. We gathered follow up data on 293 patients (96.1%). Post-discharge survival rates were 67.6% at 3 months, 49.4% at 1 year and 38.1% at 3 years. Length of survival was significantly dependent on age, weaning success and main diagnosis (i.e., prognosis in COPD is worse compared to thoracic restriction, neuromuscular disease and others) in the multivariate analysis. CONCLUSIONS: Difficult-to-wean patients have a high hospital mortality rate and poor long-term prognosis. Age, main diagnosis, severity of illness, weaning success and institution of NIV predict survival. PMID- 12122530 TI - Influences on physicians' choices of intravenous colloids. AB - OBJECTIVES: Controversy over the optimal intravenous fluid for volume resuscitation continues unabated. Our objectives were to characterize the demographics of physicians who prescribe intravenous colloids and determine factors that enter into their decision to choose a colloid. DESIGN: Questionnaire with 61 items. PARTICIPANTS AND SETTING: Ten percent ( n = 364) of frequent intravenous fluid prescribers in the province of Ontario, Canada. RESULTS: The response rate was 74%. Colloid use in the past year was reported by 79% of the responding physicians. Important reasons for choosing a colloid included blood loss and manipulation of oncotic pressure. Physicians tended to prefer either albumin or pentastarch, but no important reasons were found for choosing between the two. Albumin with or without crystalloid was preferred in 5/13 scenarios by more than 50% of the respondents, whereas pentastarch was not favored by more than 50% of respondents in any scenario. Physicians practising in critical care areas and teaching hospitals generally preferred pentastarch to albumin. Physicians reporting pentastarch as representing greater than 90% of total colloid use were more likely to have been visited by a drug detailer for pentastarch than those who used less synthetic colloid (54 vs 22%, p < 0.001). CONCLUSIONS: The majority of physicians surveyed prescribe colloid products and the reported use of albumin and pentastarch has a bimodal distribution. Although albumin appeared to be preferred in more clinical niches, most physicians did not state reasons for choosing between products. Marketing, specialty, location of practice and clinical scenario appear to play significant roles in the utilization of colloid products. PMID- 12122531 TI - Conversion of recent-onset atrial fibrillation or flutter with ibutilide after amiodarone has failed. AB - OBJECTIVE: To evaluate whether ibutilide can convert atrial fibrillation or flutter in patients in whom amiodarone has failed. DESIGN AND SETTING: Clinical study in a university hospital intensive care unit (ICU). PATIENTS: Twenty-six patients were studied, in whom atrial fibrillation or flutter persisted for a maximum of 6 h at maximum. Patients were monitored continuously during the arrhythmia. Medical conversion was necessary due to symptomatic or hemodynamic causes. INTERVENTIONS: All patients initially received amiodarone (150 mg i.v.) and after 2 h of persistent arrhythmia ibutilide (1 mg or, without success and body weight > 70 kg, 2 mg i.v.). Before the administration of ibutilide 1 g magnesium was administered, and high normal levels of potassium serum levels were achieved (4.5-5.0 mmol/l). RESULTS. After amiodarone atrial flutter persisted in 73% and atrial fibrillation in 27% of patients. After ibutilide the QT interval was prolonged from 327 +/- 61 to 387 +/- 62 ms. The QTc interval increased from 456 +/-32 to 461 +/- 66 ms. Conversion to normal sinus rhythm was achieved in 22 of 27 of cases. Nonsustained torsade de pointes tachycardia was seen in three patients (11%). No patient showed sustained ventricular tachycardia. Patients with proarrhythmic effects were characterized by a decreased left ventricular function. CONCLUSIONS: In ICU patients ibutilide led to conversion to sinus rhythm in 81.5% of patients in whom amiodarone was unsuccessful. Nonsustained tachycardias were seen in 11%; sustained ventricular tachycardia was not seen. Ibutilide seems to be well suitable for conversion of recent onset atrial fibrillation or flutter and had no severe side effects in this study population. PMID- 12122532 TI - Changes in severe accidental tetanus mortality in the ICU during two decades in Brazil. AB - INTRODUCTION: Tetanus is still a significant health hazard in developing countries, with high associated mortality. OBJECTIVE: Describe the management of patients with severe tetanus in intensive care units (ICUs), in two different periods. SETTING: ICUs of two general hospitals. DESIGN: Concurrent cohort study. METHODS: Follow-up of all patients hospitalized with the diagnosis of severe tetanus in the ICUs from October 1981 to March 2001. We collected data prospectively, regarding the site of injury, clinical features, frequent clinical and infectious complications, concomitant illnesses, and mortality. The patients were divided into two groups according to the treatment protocol used; before 1993 and after 1993. RESULTS: There were 126 patients in group 1 (93 males) with a mean age of 39.0 +/- 18.8 years. There were 110 patients in group 2 (95 males) with a mean age of 48.4+/-17.8 years. Incubation period, onset period, and symptomatic period were higher in group 2 ( P < or = 0.02). The duration of neuromuscular junction blockade, benzodiazepine administration, mechanical ventilation, and ICU stay were longer in group 2, P < 0.001. Infectious complications were more frequent in group 2 ( P < 0.001). The mortality rate in group 1 was 36.5% and in group 2, 18.0% ( P = 0.002). Mortality was directly associated with symptomatic period, acute renal failure cardiac arrest and hypotension, and inversely associated with onset period in the multivariate analyses. CONCLUSIONS: The reduced mortality in severe accidental tetanus patients in group 2 is probably related to advances in ICU management, despite the higher incidence of infectious complications, which are probably related to the longer ICU stay. PMID- 12122533 TI - Experience with a once-daily dosing program of aminoglycosides in critically ill patients. AB - BACKGROUND: As aminoglycosides show concentration-dependent killing, once-daily aminoglycoside (ODA) regimens have been instituted. Data on experience with ODA regimens in critically ill patients are limited. OBJECTIVES: 1) To evaluate the ODA-program in critically ill patients; 2) to describe the pharmacokinetics of aminoglycosides (gentamicin and tobramycin); and 3) to assess the incidence of nephrotoxicity associated with an ODA regimen in this specific of group patients. DESIGN: A prospective, descriptive study. SETTING: Eighteen-bed surgical and 12 bed medical intensive care unit in a referral centre. PATIENTS: Eighty-nine critically ill patients with a suspected or confirmed infection for which gentamicin or tobramycin was indicated and a creatinine clearance > 30 ml/min were monitored. One hundred and nine pharmacokinetic profiles were gathered. INTERVENTIONS: A first dose of 7 mg/kg/24 h of gentamicin or tobramycin was given to every patient independent of renal function. Subsequent doses were chosen on the basis of the pharmacokinetic results of the first dose. MEASUREMENTS: Serum samples were collected 1 h and 6 h after start of the aminoglycoside infusion. All samples were assayed by using immunofluorescence. Pharmacokinetic parameters were estimated using a one-compartment model. RESULTS: The volume of distribution of aminoglycosides was significantly higher in critical ill patients with septic shock than in those without. Consequently, the maximum concentration reached was significantly lower in patients with septic shock. In P. aeruginosa infections the mean (SD) estimated Cmax/MIC ratio was 10.3 (3.3). In n = 17 (49%) of the patients treated > 24 h ( n = 35), a dose adjustment or lengthening of interval was necessary. The recommended dosing interval based on the Hartford Hospital nomogram and one-serum concentration at 6 h was correct in only 62% of all cases. Signs of renal impairment occurred in n = 12 (14%) of the patients; in all survivors renal function recovered completely and no haemofiltration was needed. CONCLUSIONS: An ODA-regimen of 7 mg/kg produced Cmax/MIC ratios > 10 in the majority of critically ill patients in our population. Septic shock and renal dysfunction caused an aberrant pharmacokinetic profile of aminoglycosides in these patients. Therefore, individual therapeutic drug monitoring is warranted. Signs of renal impairment were common in the presence of shock, but appeared to be reversible. PMID- 12122534 TI - Bedside transpyloric tube placement in the pediatric intensive care unit: a modified insufflation air technique. AB - OBJECTIVE: To test air insufflation as an adjunct to placement of enteral feeding tubes and the effectiveness of using a smaller insufflation volume in pediatric patients. DESIGN AND SETTING: A randomized, controlled study in a pediatric intensive care unit in two tertiary hospitals. PATIENTS: A total of 78 children with indication for transpyloric tube feeding were studied. INTERVENTIONS: An unweighted feeding tube was placed into the stomach through the nares; a 20-ml syringe was used to insufflate 10 ml/kg air into the stomach. The tube was advanced an estimated distance into the pylorus or beyond. The control group received the same procedure except for air insufflation. Resident physicians performed all procedures. Abdominal radiography was performed 3 h later. RESULTS: Of 38 tubes in the study group 33 (86.8%) were successfully placed in a single attempt, compared to 18 of 40 tubes (45%) in the control group. Compared with the technique of using 20 ml/kg air for insufflation, no statistically significant difference was observed. No significant complication was observed. CONCLUSIONS: The gastric insufflation technique required no expensive equipment, minimal training, and consistently allowed transpyloric passage of feeding tubes. The use of 10 ml/kg air may significantly improve the rate of success without increasing risks. PMID- 12122535 TI - Cardiac output determination in children: equivalence of the transpulmonary thermodilution method to the direct Fick principle. AB - OBJECTIVE: To show the equivalence of the transpulmonary thermodilution method to the direct Fick principle in children. DESIGN: Prospective single-centre study. SETTING: A 16-bed paediatric cardiac ICU and a cardiac catheterisation laboratory at an university affiliated centre for paediatric cardiology and congenital heart disease. PATIENTS: We consecutively investigated 18 patients (mean age 12.1 +/- 6.4 years) during cardiac catheterisation and after corrective cardiac operation. METHODS AND RESULTS: We prospectively defined limits of equivalence for cardiac index (CI) for both methods of +/- 0.25 l/min x m(2). We measured oxygen consumption for determination of CI by Fick as the clinical "gold standard" and performed a set of three transpulmonary thermodilution measurements. The mean CI(Fick) was 2.88 +/- 1.07 l/min x m(2) (range 1.10-4.62 l/min x m(2)) and CI(TPID)was 2.85 +/- 1.03 l/min x m(2)(range 1.02-4.49 l/min x m(2)). The mean difference between CI(Fick) and CI(TPID)was 0.030 +/- 0.168 l/min x m(2), and limits of agreement -0.306 to 0.366 l/min x m(2)(90% confidence interval -0.040 to 0.099 l/min x m(2)). The regression equation was : CI(Fick)=1.0244 x CI(TPID) 0.040, r(2) = 0.976, P < 0.0001. The intraclass coefficient of reliability for three repeated measurements of CI(TPID) was 0.97, the corresponding lower limit of the 95% confidence interval was 0.94. CONCLUSION: We demonstrated the equivalence of CI measurement by transpulmonary thermodilution and the Fick principle in children. This new method may improve hemodynamic monitoring and management in seriously ill children. PMID- 12122536 TI - Jejunal luminal microdialysate lactate in cardiac tamponade--effect of low systemic blood flow on gut mucosa. AB - OBJECTIVE: To assess gut mucosal metabolic response and susceptibility to dysoxia during low systemic blood flow induced by cardiac tamponade. DESIGN: A randomized, controlled animal experiment. SETTING: National laboratory animal center. INTERVENTIONS: Cardiac tamponade was induced in six pigs, while six additional pigs served as controls. In the tamponade group, fluid was injected into the pericardial space to reduce aortic flow, aiming first at a flow of 50 ml/kg per min and then at 30 ml/kg per min. Each step lasted for 60 min. MEASUREMENTS AND RESULTS: We measured luminal lactate by microdialysis and mucosal PCO(2) by tonometry in the mid-jejunum. Aortic and superior mesenteric artery blood flow, arterial and mesenteric venous lactate, pyruvate and ketone bodies and regional lactate exchange were measured. The distribution of aortic blood flow to superior mesenteric artery remained unchanged (baseline 14 (12 16)%; median (interquartile range), stepwise flow reduction 11 (10-17)% and 13 (12-19)%, NS) during reduction of aortic blood flow from 81 (61-95) ml/kg per min to 49 (47-49) ml/kg per min and 23 (21-27) ml/kg per min. Systemic hyperlactatemia developed early, whereas gut luminal lactate increased only after 60 min of hypoperfusion and could be largely explained by arterial hyperlactatemia. Mesenteric venous lactate-to-pyruvate (L/P) ratio increased after 30 min of tamponade, but both venous-arterial lactate and pyruvate gradients turned negative (gut lactate and pyruvate uptake). Mesenteric venous ss hydroxybutyrate to acetoacetate ratio increased after 60 min. No changes were observed in the controls. CONCLUSIONS: Jejunal mucosal dysoxia and anaerobic metabolism occurs late during low systemic blood flow induced by experimental cardiac tamponade. PMID- 12122537 TI - The effect of dehydroepiandrosterone on hemorrhage-induced suppression of cellular immune function. AB - OBJECTIVE: To determine whether the steroid hormone dehydroepiandrosterone (DHEA) improves cellular immune functions after hemorrhagic shock. DESIGN AND SETTING: Prospective controlled study in a research laboratory at an university medical center. SUBJECTS: Male NMRI mice. INTERVENTIONS: Animals received 0.9% saline or DHEA (20 mg/kg subcutaneously) before induction of a volume-controlled hemorrhagic shock (55% of estimated circulating blood volume) by retro-orbital puncture. One hour after hemorrhage mice underwent fluid resuscitation by intravenous infusion of lactated Ringer's solution (300% of the shed blood). Separate groups of mice were killed to obtain whole blood and spleen 1 h after hemorrhage, 1 h after fluid resuscitation, and 24 h after hemorrhage to determine lymphocyte distribution (CD4(+), CD8(+), NK1.1-AG(+)), splenocyte apoptosis, and plasma concentrations of tumor necrosis factor-alpha and interleukin-10. MEASUREMENTS AND RESULTS: Hemorrhage in control mice was associated with a rapid increase in circulating NK cell numbers. Elevated splenocyte apoptosis, an increased CD4/CD8 ratio, and decreased number of circulating CD8(+) T-cells was observed 24 h after hemorrhagic shock. DHEA administration was accompanied by a normalization of splenocyte apoptosis and lymphocyte migration. Induction of hemorrhagic shock did not affect TNF-alpha or IL-10 plasma concentrations in either treatment group. CONCLUSIONS: DHEA administration improves cellular immune function after hemorrhage and may therefore be beneficial in patients with hemorrhagic shock. PMID- 12122538 TI - The effects of dexmedetomidine on the ventilatory response to hypercapnia in rabbits. AB - OBJECTIVE: Dexmedetomidine is a highly selective alpha(2)-adrenergic agonist that can reduce anesthetic requirements. This study, to assess its effect on respiration, examined the effects of various doses of dexmedetomidine (1, 10, 30 and 50 microg/kg) on the respiratory response to carbon dioxide (CO(2)) breathing in rabbits. DESIGN: Randomized prospective study. SETTING: Animal laboratory at a university school of medicine. INTERVENTION: From 28 animals, four groups of seven were randomly assigned to receive different doses of dexmedetomidine (groups D1, D10, D30 and D50). Under inhalation of sevoflurane, each animal was tracheostomized and intubated with a 4 mm internal diameter (i.d.) endotracheal tube. MEASUREMENTS AND RESULTS: After end-tidal sevoflurane concentration had decreased below 0.03% and during quiet breathing (QB); respiratory rate (RR), tidal volume (V(T)) and inspiratory time (T(I)) were measured, from which minute ventilation (MV) and mean inspiratory flow (V(T)/T(I)) were calculated. After these measurements had been completed, each animal breathed the balloon gas (5% CO(2) and 95% O(2)) until the end-tidal CO(2) (ETCO(2)) reached 10%. The respiratory measurements were repeated during the latter period. After the collection of these data, dexmedetomidine was infused intravenously and the same measurements were repeated 15 and 45 min after dexmedetomidine infusion. The slope of the ventilatory response to hypercapnia in D50 was significantly higher compared with D30 animals. In the range 1-30 microg/kg, during both QB and at 10% ETCO(2), MV was decreased in a dose-dependent manner. Dexmedetomidine depressed both V(T) and RR during QB and at 10% ETCO(2). CONCLUSION: Dexmedetomidine depressed resting ventilation and the respiratory response to CO(2), but it did not induce profound hypoxemia or hypercapnia in rabbits. PMID- 12122539 TI - Successful treatment of a complicated case of neuroleptic malignant syndrome. AB - Neuroleptic malignant syndrome (NMS) is a life-threatening reaction often related to neuroleptic drugs, characterized by rigidity, hyperthermia, altered consciousness, and fluctuating blood pressure. We present a case of NMS that followed a doubled oral dose of a drug compound: tranylcypromine sulfate, a monoamine oxidase inhibitor, and trifluoperazine (neuroleptic). The case was complicated by rhabdomyolisis and disseminated intravascular coagulation. It was treated successfully with dantrolene sodium and generous fluid therapy without using neuromuscular blocking agents or dopamine agonists. PMID- 12122540 TI - Luxatio cordis due to right pericardium tear, a difficult diagnosis: report of a case. AB - BACKGROUND: Dislocation of the heart is a rare complication of thoracic blunt trauma. A high index of suspicion of pericardium rupture is necessary to formulate an early diagnosis to reduce morbidity and mortality. PATIENTS: A 23 year-old man suffered a blunt thoracoabdominal trauma and was admitted 3 days later to a university hospital ICU for right heart luxation due to right pericardial tear. Mechanical ventilation delayed radiological findings. METHODS AND RESULTS: Surgery by repositioning the heart and repairing the pericardial tear allowed restoration of hemodynamic equilibrium. PMID- 12122541 TI - Low plasma granulocyte-macrophage colony stimulating factor is an indicator of poor prognosis in sepsis. AB - OBJECTIVE: Monocyte dysfunction has been shown to be associated with adverse consequences in septic patients. The cytokine growth factor granulocyte macrophage colony stimulating factor (GM-CSF) may be required for optimal monocyte function in these patients. The current study investigates whether plasma GM-CSF levels were significantly different in septic patients and whether there was an association with prognosis. DESIGN: Plasma samples were collected from all septic patients from day 1 of the diagnosis of sepsis for 3 days. Healthy volunteer plasma served as control samples. A novel enzyme-linked immuno adsorbent assay was developed with suitable sensitivity for detection of GM-CSF in patient and normal plasma. APACHE II score, age, sex and outcome were determined for all patients. SETTING: A single centre study at the Royal Liverpool University Hospital in a medico-surgical 13 bed intensive care unit. PATIENTS: All septic patients (n = 53) fulfilling the criteria of the APCC for the diagnosis of sepsis, were recruited for the study with informed consent from day 1 of the diagnosis of sepsis and plasma GM-CSF measured on three consecutive days. Patients were excluded from the study if on immunosuppressive therapy. Normal healthy volunteers (n = 33) were included in the study to serve as controls. RESULTS: Plasma GM-CSF levels were statistically significantly depressed in patients who died compared with those who survived, who had levels comparable with healthy controls. CONCLUSIONS: The results indicate that low plasma GM-CSF is associated with adverse consequences for septic patients. The measurement of GM-CSF in the plasma of septic patients merits further study for use as a prognostic marker and also to identify the type of immunotherapy the patient may benefit from. PMID- 12122542 TI - Inspiratory pressure-volume curves obtained using automated low constant flow inflation and automated occlusion methods in ARDS patients with a new device. AB - OBJECTIVE: To compare the inspiratory volume pressure (VP) curves of the respiratory system (rs) produced by static occlusion (OCC) and dynamic low constant flow inflation (LCFI) methods using a new device in acute respiratory distress syndrome (ARDS) patients. SETTING: A multidisciplinary 24-bed ICU in a tertiary university hospital. PATIENTS: Eleven intubated and mechanically ventilated patients with ARDS. MEASUREMENTS AND RESULTS: OCC and LCFI methods were performed using the same ventilator, which had been specifically implemented for this purpose. LCFI of 5, 10, and 15 l/min and OCC were applied in a random order at zero end-expiratory positive pressure. Airway pressure was measured both proximal (P(ao)) and distal (P(tr)) to the endotracheal tube. Lower inflection point (LIP) and maximal slope (C(max,rs)) were estimated using unbiased iterative linear regressions. LIP(rs) was obtained in all patients under LCFI and in nine patients under OCC. With LCFI of 5, 10, 15 l/min and OCC the average LIP(rs) values were 12.2 +/- 3.9, 12.9 +/- 4, 14.3 +/- 3.4, and 11.9 cm H(2)O for P(ao) and 11.9 +/- 3.9, 11.5 +/- 3.3, 12.5 +/- 3.4 and 11.8 +/- 4.4 for P(tr), respectively. Only the mean values of LIP(rs) for P(ao) with LCFI at 15 l/min were significantly different from those obtained for OCC. The C(max,rs) values found with the two methods were similar. CONCLUSIONS: An LCFI less than or equal to 10 l/min seems to be a quick, safe, and reliable method to determine LIP(rs) and C(max,rs) at the bedside. PMID- 12122544 TI - Predicting the resting metabolic rate of 30-60-year-old Australian males. AB - OBJECTIVES: This study: (a) generated regression equations for predicting the resting metabolic rate (RMR) of 30-60-y-old Australian males from age, height, mass and fat-free mass (FFM); and (b) cross-validated RMR prediction equations which are currently used in Australia against our measured and predicted values. DESIGN: A power analysis demonstrated that 41 subjects would enable the detection of (alpha=0.05, power=0.80) statistically and physiologically significant differences of 8% between predicted/measured RMRs in this study and those predicted from the equations of other investigators. SUBJECTS: Forty-one males ([X]+/-s.d.:, 44.8+/-8.6 y; 83.50+/-11.32 kg; 179.1+/-5.0 cm) were recruited for this study. INTERVENTIONS: The following variables were measured: skinfold thicknesses; RMR using open circuit indirect calorimetry; and FFM via a four compartment (fat mass, total body water, bone mineral mass and residual) body composition model. RESULTS: A multiple regression equation using mass, height and age as predictors correlated 0.745 with RMR and the s.e.e. was 509 kJ/day. Inclusion of FFM as a predictor increased both the correlation and the precision of prediction, but there was no difference between FFM via the four-compartment model (r=0.816, s.e.e.=429 kJ/day) and that predicted from skinfold thicknesses (r=0.805, s.e.e.=441 kJ/day). CONCLUSIONS: Cross-validation analyses emphasised that equations need to be generated from a large database for the prediction of the RMR of 30-60-y-old Australian males. PMID- 12122545 TI - Multiple coronary risk factors in healthy older Kuwaiti males. AB - OBJECTIVE: The objective was to examine the percentage and severity of obesity and some common biochemical coronary heart disease (CHD) risk factors in a sample of healthy Kuwaiti adult males > or =45 y of age. We also sought to determine the percentage of males at increased risk due to the presence of multiple CHD risk factors. DESIGN: The study was a cross-sectional study. SETTING: The study was conducted in all men who underwent a mandatory job related physical examination or who sought to obtain retirement benefits. SUBJECTS: A total of 740 healthy Kuwaiti males between the ages of 45 to 80 y. RESULTS: In all, 37% of the men were obese, 26% had elevated glucose values and 52% had elevated or high cholesterol levels. Some had multiple risk factors for CHD, including age > or =45 y, obesity, male gender, hyperglycemia and hypercholesterolemia. Blood glucose values increased consistently from younger (45-54 y) to older age groups (55-64 and > or =65 y). Blood glucose values varied inversely with education level. CONCLUSIONS: In societies with relatively uniform income levels, educational level may be a better indicator of chronic disease risk than income per se. These results, showing a high percentage of men with several risk factors and high mean values, suggesting more severe risk, suggest that immediate action should be taken to develop a public health intervention strategy to educate Kuwaiti men to become aware of the causes and correlates of CHD and how to decrease their risk for CHD and heart attack, the leading cause of death in Kuwait. PMID- 12122546 TI - Role of fibre and fruit in the Mediterranean diet to protect against myocardial infarction: a case-control study in Spain. AB - OBJECTIVE: To assess the association between a first acute myocardial infarction and the consumption of fibre and fruit. DESIGN: Hospital-based case-control study with incident cases. A validated semi-quantitative food frequency questionnaire (136 items) was used to assess food intake. SETTING: Three third-level university hospitals in Pamplona (Spain). SUBJECTS: Cases were subjects aged under 80, newly diagnosed with acute myocardial infarction. Each case patient (n=171) was matched to a control subject of the same gender and age (5 y bands) admitted to the same hospital. RESULTS: An inverse association was apparent for the three upper quintiles of fibre intake. After adjustment for non-dietary and dietary confounders, an inverse linear trend was clearly significant, showing the highest relative reduction of risk (86%) for the fifth quintile (OR=0.14, 95% confidence interval: 0.03-0.67). An inverse association was also apparent for fruit intake, but not for vegetables or legumes. CONCLUSIONS: Our data suggest that a substantial part of the postulated benefits of the Mediterranean diet on coronary risk might be attributed to a high intake of fibre and fruit. PMID- 12122547 TI - Assessment by bioimpedance of forearm cell mass: a new approach to calibration. AB - OBJECTIVE: Changes in skeletal muscle mass are involved in several important clinical disorders including sarcopenia and obesity. Unlike body fat, skeletal muscle is difficult to quantify in vivo, particularly without highly specialized equipment. The present study had a two-fold aim: to develop a regional (40)K counter for non-invasively estimating cell mass in the arm, mainly skeletal muscle cell mass, without radiation exposure; and to test the hypothesis that cell mass in the arm is highly correlated with electrical impedance after adjusting for the arm's length. METHODS: Forearm cell mass was estimated using a rectangular lead-shielded (40)K counter with 4-NaI crystals; impedance of the arm was measured at multiple frequencies using a segmental bioimpedance analysis (BIA) system. The system's within- and between-day coefficient of variation (CV) for (40)K-derived elemental potassium averaged 1.8+/-1.3 and 5.8+/-1.2%, respectively. The corresponding BIA system's CVs were 1.0+/-0.4 and 2.1+/-1.0%, respectively. SUBJECTS AND RESULTS: Participants in the study were 15 healthy adults (eight females, seven males; age 39+/-2.8 y, BMI 22.9+/-4.5 kg/m(2)). The right arm's K (5.2+/-1.7 g) was highly correlated with length-adjusted impedance (r(2)=0.81, 0.82, and 0.83 for 5, 50 and 300 kHz, respectively; all P<0.001); multiple regression analysis showed no additional improvement by adding age or sex to the prediction models. CONCLUSION: These results demonstrate the feasibility of calibrating BIA-measured electrical properties of the arm against estimates of arm cell mass, mainly of skeletal muscle, obtained by regional (40)K counting. This simple and practical approach should facilitate the development of BIA-based regional cell mass prediction formulas PMID- 12122548 TI - Why healthcare workers give prelacteal feeds. AB - OBJECTIVE: Because prelacteal feeds can adversely affect breastfeeding, UNICEF/WHO discourage their use unless medically indicated. The study was carried out to determine the proportion of healthcare workers who routinely give prelacteal feeds, and their reasons for doing so; further, to determine whether any differences exist between medically and non-medically trained healthcare workers in their administration of prelacteal feeds. DESIGN: Survey. SETTING: Primary, secondary and tertiary health facilities in Kaduna township Nigeria. SUBJECTS: Of 1100 healthcare workers sampled, 747 (68%) responded. Of these 80% had received medical training, 20% had not. METHODS: Use of a pretested validated questionnaire. RESULTS: Large proportions of both medical and non-medically trained healthcare workers stated they routinely give prelacteal feeds (doctors, 68.2%; nurses, 70.2%; and non-medical, 73.6%). However their reasons for doing so differed significantly (P=0.00001). Nurses gave mainly for perceived breast milk insufficiency, doctors for prevention of dehydration, hypoglycaemia and neonatal jaundice and non-medical staff to prepare the gastrointestinal tract for digestion and to quench thirst. CONCLUSIONS: Most healthcare workers (medical and non-medical) routinely and unnecessarily give prelacteal feeds. Therefore training and retraining programmes in lactation management are necessary and must include non-medical staff. These programmes, while emphasizing the danger of giving prelacteal feeds, must deal with the misconceptions of each group. Deliberate efforts have to be made to incorporate clinical training in breastfeeding in curricula of Schools of Medicine and Nursing. PMID- 12122549 TI - A comparison between an in vitro method to determine carbohydrate digestion rate and the glycemic response in young men. AB - OBJECTIVE: This study examines the relationship between the results of in vitro determinations of carbohydrate digestion rates and the glycemic index. SUBJECTS: Ten healthy non smoking men, aged between 21 and 24-y-old, were selected to participate in the study. METHODS: Six different meals with similar levels of carbohydrates were assayed at breakfast in ten subjects and blood samples were obtained at 0, 15, 30, 60 and 120 min to determine glucose levels. The Englyst's enzymatic method was used in the in vitro study. The six meals were based in cereals (rice or spaghetti); legumes (lentil soup and beans with spaghetti); and potato (potato stew with meat and vegetables). RESULTS: The meals showed different glycemic indices: rice and spaghetti based meals had lower values (mean value 31.4 and 42, respectively); the intermediate values corresponded to lentil soup and the bean dish (49.3, and 76.8 respectively) and the higher glycemic index was shown by the potato based meal (82). CONCLUSIONS: A significant correlation was observed when the ratio of rapid carbohydrate digestion rate and the lente carbohydrate digestion rate was correlated with the glycemic index of the meals, but not when only the rapid carbohydrate digestion rate was considered. These results demonstrate a useful, simple and inexpensive method to estimate the biological response of high carbohydrate meals. PMID- 12122550 TI - Meal patterns and obesity in Swedish women-a simple instrument describing usual meal types, frequency and temporal distribution. AB - OBJECTIVE: To characterize meal patterns in relation to obesity in Swedish women using a simple instrument describing meal frequency, meal types and temporal distribution. DESIGN: Cross-sectional parallel group design. SUBJECTS: Eighty three obese women from the Swedish Obese Subjects (SOS) study were compared with 94 reference women, randomly recruited from the population. METHOD: A new, simplified and self-instructing questionnaire was used to assess meal patterns. Usual meal pattern was reported as time and meal type for each intake episode during a typical day. RESULTS: The obese women consumed 6.1 meals/day compared with 5.2 meals/day among the reference women (P<0.0001). All types of meals except 'drink meals' were significantly more frequently consumed in the obese group. The obese women also displayed a different meal pattern across the day, consuming a larger number of meals later in the day. As a result a larger fraction of each obese woman's total meals were consumed in the afternoon and in the evening/night. There was no difference in the number of obese vs reference women consuming breakfast. Snack meals were positively associated with total energy intake in both groups. CONCLUSIONS: A new simplified method assessing meal pattern revealed that the number of reported intake occasions across a usual day was higher in obese women compared with controls and the timing was shifted to later in the day. These findings should be considered in the treatment of obesity. PMID- 12122551 TI - Assessment of three levels of folic acid on serum folate and plasma homocysteine: a randomised placebo-controlled double-blind dietary intervention trial. AB - OBJECTIVE: To determine the minimum effective dose of folic acid required to appreciably increase serum folate and to produce a significant reduction in plasma total homocysteine (tHcy). DESIGN: Double-blind, randomised placebo controlled intervention trial. SETTING: Community-based project in a New Zealand city. SUBJECTS: Seventy free living men and women with tHcy> or =10 micromol/l. Mean age (range) was 58 (29-90) y. INTERVENTIONS: Daily consumption over 4 weeks of 20 g breakfast cereal either unfortified (placebo) or fortified with 100, 200 or 300 microg folic acid. Dietary intake was determined by weighed diet records and consumption of commercially fortified products was avoided. MAIN OUTCOME MEASURES: Plasma tHcy and serum folate concentrations. RESULTS: Average serum folate concentrations (95% CI) increased significantly in the treatment groups relative to the control group by 28(9-51)%, 60(37-87)% and 79(51-114)% for supplementation with 100, 200 and 300 microg folic acid, respectively. A reduction in tHcy was observed, being 16(8-22)%, 12(4-18)% and 17(9-24)% in the three treatment groups, respectively. CONCLUSIONS: A regular intake of as little as 100 microg folic acid per day was sufficient to lower tHcy in persons at the upper end of the normal range for tHcy. Low-level fortification may also be appropriate for lowering the risk of neural tube defects given that, when aggregated from all sources, the total intake of folic acid may be sufficiently high to adequately improve the folate status of young women. PMID- 12122552 TI - The comparative gastrointestinal responses of children and adults following consumption of sweets formulated with sucrose, isomalt and lycasin HBC. AB - OBJECTIVES: To determine the gastrointestinal responses of children and adults following consumption of sucrose, isomalt and lycasin HBC and to compare these at two different dose levels in adults. DESIGN: Both studies were randomised, double blind, cross-over designs. SUBJECTS: Fifty-one children aged 6-9 y were recruited from primary schools in the Salford area of Greater Manchester. Forty-eight children completed the study. Fifty healthy adult volunteers aged 18-24 y were recruited from the student population of the University of Salford. All subjects completed the study. INTERVENTIONS: Children consumed either 25 g of sucrose, isomalt or lycasin HBC and adults 25 and 40 g in hard boiled sweets per day for two consecutive test days. Test periods of 2 days were separated by 7 day washout periods. Children consumed sweets throughout test days and adults in no less than 30 min but no more than 90 min. Subjects reported the prevalence and magnitude of flatulence, borborygmi, bloating, colic, bowel movements and watery faeces. RESULTS: Consumption of 25 g isomalt provoked a mild laxative effect in children but not in adults. Consumption of 25 g isomalt significantly increased the prevalence and magnitude of gastrointestinal responses in both children and adults. Consumption of 25 g lycasin HBC significantly increased borborygml in children and adults but no other gastrointestinal responses. Consumption of 40 g lycasin HBC or isomalt by adults significantly increased the mean frequency of bowel movements and the number of subjects passing watery faeces. In adults, 40 g isomalt and lycasin HBC provoked significantly more gastrointestinal responses compared to 25 g of either product. CONCLUSIONS: Consumption of 25 g lycasin HBC does not provoke an unacceptable laxative effect or gastrointestinal response in children or adults compared to 25 g isomalt, which is associated with a mild laxative effect and increase in gastrointestinal responses. In adults gastrointestinal responses following consumption of products were found to be dose dependent. PMID- 12122553 TI - Assessment of iodine intake in vegans: weighed dietary record vs duplicate portion technique. AB - OBJECTIVE: To compare iodine intakes estimated from weighed dietary records with iodine intakes obtained by direct analysis of duplicate diets in a group of vegans. DESIGN: Cross-sectional study. SETTING: London and the south-east of England. SUBJECTS: Thirty-three vegans consuming their habitual diet were recruited through the UK Vegan Society; 26 subjects (11 males, 15 females), age 21-84 y, completed the study. INTERVENTIONS: Iodine intakes were estimated from 4 day weighed dietary records and compared with iodine intakes obtained by direct analysis of concurrent 4 day duplicate diets. RESULTS: There was wide variation in iodine intakes. Mean daily iodine intake in males was significantly lower (P<0.05) when estimated from dietary records (42 microg) compared with that analysed from duplicate diets (137 microg). Conversely, in females the mean daily iodine intake from dietary records (1448 microg) was higher (P=0.43) than from duplicate diets (216 microg). Variation in iodine intakes determined by the two different methods may be attributed to the absence of iodine content of some foods, in particular foods suitable for vegan consumption, in food composition tables and the variability in iodine content of seaweed. CONCLUSIONS: The use of current food tables to estimate iodine intake in vegans is limited. It is not always practical to determine iodine intake using the duplicate portion technique, therefore more reliable information on iodine content of foods, incorporating the variation within foods, is needed. PMID- 12122555 TI - Use of the scored Patient-Generated Subjective Global Assessment (PG-SGA) as a nutrition assessment tool in patients with cancer. AB - OBJECTIVE: To evaluate the use of the scored Patient-Generated Subjective Global Assessment (PG-SGA) as a nutrition assessment tool in patients with cancer. DESIGN: An observational study assessing the nutritional status of patients with cancer. SETTING: Oncology ward of a private tertiary Australian hospital. SUBJECTS: Seventy-one cancer patients aged 18-92 y. INTERVENTION: Scored PG-SGA questionnaire, comparison of scored PG-SGA with subjective global assessment (SGA), sensitivity, specificity. RESULTS: Some 24% (17) of 71 patients were well nourished, 59% (42) of patients were moderately or suspected of being malnourished and 17% (12) of patients were severely malnourished according to subjective global assessment (SGA). The PG-SGA score had a sensitivity of 98% and a specificity of 82% at predicting SGA classification. There was a significant difference in the median PG-SGA scores for each of the SGA classifications (P<0.001), with the severely malnourished patients having the highest scores. Re admission within 30 days of discharge was significantly different between SGA groups (P=0.037). The mortality rate within 30 days of discharge was not significantly different between SGA groups (P=0.305). The median length of stay of well nourished patients (SGA A) was significantly lower than that of the malnourished (SGA B+C) patients (P=0.024). CONCLUSION: The scored PG-SGA is an easy to use nutrition assessment tool that allows quick identification and prioritisation of malnutrition in hospitalised patients with cancer. PMID- 12122554 TI - The effect of guar gum addition to a semisolid meal on appetite related to blood glucose, in dieting men. AB - OBJECTIVE: To investigate whether addition of modified guar gum (GG) to a low energy semisolid meal might be effective on appetite by modifying the response of blood glucose and other blood parameters. DESIGN: Three intervention periods of 2 weeks each, separated by washout periods of 4 weeks. Randomized and cross-over design. SUBJECTS: Fifteen overweight male subjects (mean+/-s.d.; age, 44+/-9 y; body mass index, 28.6+/-1.8 kg/m(2)). INTERVENTION: Subjects consumed a low energy diet divided over three times a day, consisting of a semisolid meal with (SSM+) or without (SSM) addition of 2.5 g GG, or a solid meal (SM) with the same energy content (947 kJ) and macronutrient composition, plus a dinner of the subject's own choice. At the end of each intervention, time and number of meal initiations, dynamics of blood glucose and other blood parameters, and appetite ratings such as hunger and satiety were determined in a time-blinded situation. RESULTS: The changes in blood glucose from meal initiation to blood glucose peak and from peak to nadir were smaller with SSM+ and SM compared to SSM. Satiety before the third meal was higher with SSM+ and SM compared to SSM (P<0.01). Meal pattern, general appetite and total energy intake were similar for all treatments. CONCLUSIONS: We conclude that, similar to SM, SSM+ resulted in a more moderate change in blood glucose compared to SSM and positively affected satiety before the third meal, while general appetite, total energy intake and meal pattern did not differ. PMID- 12122556 TI - Dietary fats and 16-year coronary heart disease mortality in a cohort of men and women in Great Britain. AB - OBJECTIVE: The paper aims to investigate the relationships of dietary fats to subsequent coronary heart disease (CHD) mortality in men and women while taking account of other CHD-related behaviours. DESIGN: A cohort of randomly selected men and women were interviewed in 1984-85 and monitored subsequently for 16 y for deaths. The interview covered health, health-related behaviours, physical measurements, socio-demographic details and a dietary questionnaire. Appropriate exclusions left 1225 men and 1451 women aged 40-75 with 98 and 57 CHD deaths, respectively. Saturated, polyunsaturated and total fat intakes were estimated. SETTING: The sample was randomly selected from households in Great Britain. The interviews took place in participants' own homes. RESULTS: Not consuming alcohol, smoking, not exercising and being socially disadvantaged were related to high saturated fat intake and CHD death. Cox survival analyses adjusting for these factors found that a level of saturated fat 100 g per week higher corresponded to a relative risk for CHD death for men of 1.00 (0.86-1.18) and 1.40 (1.09-1.79) for women. This difference between the effects of saturated fat in men and women was statistically significant (P=0.019). Results are also reported for total fat and the relative effects of polyunsaturated and saturated fats. CONCLUSIONS: Strong evidence was found for the within cohort relationship of dietary fat and CHD death in women while no evidence was found for a relationship in men. Possible explanations for this are discussed. PMID- 12122557 TI - Munich workshop on evaluation of fiber and particle toxicity: an introduction. PMID- 12122558 TI - Inflammation caused by particles and fibers. PMID- 12122559 TI - Toxicokinetics and effects of fibrous and nonfibrous particles. AB - Chronic inhalation of fibrous and nonfibrous particles by rats at high concentrations results in lung tumor formation if the particles are poorly soluble in the lung. Even rather benign nonfibrous particles such as TiO(2) produce this result. One significant change during a chronic inhalation exposure of poorly soluble particles of low cytotoxicity (PSP) is an impairment of normal clearance mechanisms in the alveolar region of the lung in rats, resulting in a continued buildup to high lung burdens accompanied by chronic alveolar inflammation, fibrosis, and mutational events. Since these are obviously high dose effects, questions about their extrapolation to humans exposed to much lower concentrations have been raised. Results of key studies reported for chronic inhalation of PSP in rats indicate that mechanisms of PSP-induced lung tumors at high doses do not operate at low dose levels. Furthermore, the existence of two thresholds can be postulated: One is a dosimetric threshold for the endpoint alveolar macrophage-mediated clearance, which is related to lung particle overload. The other is a mechanistic threshold for the endpoint mutation, which is determined by the level of antioxidant defenses to counterbalance reactive oxidant species released by activated inflammatory cells. A no-observed-adverse effect level (NOAEL) could therefore be based on avoiding alteration of the toxicokinetic of the particles such that the lung burdens stay below the dosimetric threshold. The suggestion that PSP-associated organic compounds (e.g., diesel particulate matter) contribute to the lung tumor responses in rats observed in chronic inhalation studies is not supported by experimental data from in vivo studies. It can be concluded that high-dose rat lung tumors due to PSP should not be used for low-dose extrapolations, and no significant contribution to human lung cancer risk can be predicted from levels of PSP below lung overload. With respect to the pulmonary toxicokinetics of inhaled fibrous particles, the biopersistence of long fibers (>20 microm) which cannot be phagocytized by alveolar macrophages is a key parameter related to long-term carcinogenic effects. Long fibers with a very low biopersistence should not be considered as carcinogenic. Since the clearance kinetics of fibers can generally be described by a biphasic or multiphasic pattern-fast initial and slow final phase-it is essential that the slow phase of the retention kinetics of fibers longer than 20 microm is considered in a biopersistence assay. Based on the results of such assay, fibers can be classified into one of two categories: a biopersistent fiber that cannot be dissolved in the lung within an acceptable time period; or a biosoluble fiber when even long nonphagocytizable fibers will be disappearing rapidly from the lung. However, in addition to biopersistence, it should be mandatory to evaluate fiber toxicity in an appropriate assay relative to a fiber whose long-term effects are well known. Moreover, for organic fibers it is likely that different rules may have to be established for characterization of their toxic and carcinogenic potential. PMID- 12122560 TI - Mechanisms of genotoxicity of particles and fibers. AB - With regard to genotoxicity testing and cancer risk assessment, particles and fibers form a rather specific group among all toxicants. First, the physicochemical behavior of fibrous and nonfibrous particles is usually very different from that of nonparticulate, chemical carcinogens. Reactive oxygen species (ROS) are believed to play a major role in primary genotoxicity of particles, which may derive from their surface properties, the presence of transition metals, intracellular iron mobilization, and lipid peroxidation. Other aspects relevant to primary genotoxicity are particle size, shape, crystallinity (e.g., silica), and solubility, and may also include particle uptake, interaction with cell division machinery (e.g., asbestos), and the presence of mutagens carried with the particle (e.g., diesel exhaust particles, DEP). Excessive and persistent formation of ROS from inflammatory cells is considered as the hallmark of the secondary genotoxicity of nonfibrous and fibrous particles. Since lung inflammation is known to occur and persist only at sufficient particle dose, this secondary pathway is considered to contain a threshold (Greim et al., 2001). Identification of (mechanisms of) particle genotoxicity has been/can be achieved via (1) acellular assays, (2) in vitro tests, (3) in vivo studies, usually in mice or rats, and finally (4) biomarker studies in humans with (occupational) exposure. The significance of acellular assays and biomarker studies for risk assessment is limited, but has provided some mechanistic insights (e.g., in oxidant generating properties of quartz and asbestos) and may also contribute to hazard identification. In vitro studies have lead to identification of primary genotoxic properties of particles, whereas recent in vivo studies provide further support for the correlation between particle-induced lung inflammation and secondary genotoxicity. Proper risk assessment of particles necessitates identification of the relative impact of primary versus secondary genotoxicity in realistic exposure conditions. However, since it is impossible to discern between primary and secondary genotoxicity with current in vivo tests, concomitant in vitro assays are required to determine primary genotoxicity. In vivo tests should ideally be designed using different doses to allow dose-effect analysis for both inflammation and genotoxicity. PMID- 12122561 TI - Appropriate in vitro test conditions for genotoxicity testing of fibers. AB - With the exception of asbestos fibers, little information is available on genotoxicity testing of fibers (i.e., respirable-sized, fiber-shaped particulates, RFP). In contrast to standard genotoxicity testing of soluble substances, fibers bring about specific problems. Test results can be influenced by fiber dimensions, surface properties, and biopersistence. The mechanisms of fiber-induced genotoxicity are not yet clear, but direct interaction with the genetic material and indirect effects via production of reactive oxygen species (ROS) have been proposed. Asbestos did not significantly induce gene mutations in bacterial and mammalian systems but led to a clear induction of structural and numerical chromosome aberrations in cultured mammalian cells. It is the purpose of this article to critically review positive in vitro genotoxicity data obtained for asbestos in the comet assay, the chromosome aberration test, and the micronucleus test and to identify the test conditions that are specifically required for the detection of asbestos-induced genotoxicity. It is concluded that appropriate cell systems are available for testing fiber-induced genotoxicity. Fiber samples have to be well characterized and phagocytosis and cytotoxic effects have to be determined for the correct interpretation of genotoxicity test results. A combination of the micronucleus test and the comet assay using continuous treatment (without exogenous metabolic activation) seems to be well suited to detect genotoxic activity of asbestos fibers with high accuracy. Further investigations are needed to shown whether this approach can be recommended for genotoxicity testing of fibers in general. PMID- 12122562 TI - Role of DNA repair in particle- and fiber-induced lung injury. AB - Lung injury after particle exposure is associated with the generation of reactive oxygen species (ROS), leading to increased levels of oxidative DNA damage. Furthermore, both stable and unstable DNA lesions may arise after metabolization of polycyclic aromatic hydrocarbons (PAH) frequently bound to particles. Oxidative DNA damage is predominantly removed by base excision repair (BER), which is usually comparatively fast and efficient. Nevertheless, even without exogenous exposure a measurable steady-state level of oxidative DNA lesions exists due to oxygen metabolism, and repair capacities may be exceeded upon particle exposure. Stable DNA adducts induced by PAHs are eliminated by nucleotide excision repair (NER), which is unequally distributed in the genome and incomplete in transcriptionally inactive DNA regions. Thus, even low adduct levels due to environmental exposure lead to persistent lesions in the human lung. Other factors that need to be considered are potential repair inhibitions by associated metal compounds and interindividual differences in repair capacities. Taken together, the extent of DNA repair is an important determinant in particle-induced genotoxicity and carcinogenicity. PMID- 12122563 TI - Antioxidant defense mechanisms and the toxicity of fibrous and nonfibrous particles. PMID- 12122564 TI - 4-week inhalation toxicity study with a mixture of dichloroethylene and perfluorobutylethylene in rats. AB - Inhalation studies were conducted to determine the potential subchronic toxicity of a mixture of trans-1,2-dichloroethylene (70%), cis-1,2-dichloroethylene (5%), and perfluorobutylethylene (25%). Groups of rats were exposed to 0, 400, 2000, or 8000 ppm concentrations of the mixture vapor 6 h/day, 5 days/wk, for a total of 20 exposures. Subgroups of rats were further observed during a 1-mo recovery period. Functional observational battery (FOB) and motor activity (MA) behavioral tests were conducted prior to initiation of the exposures, during exposure wk 4, and after a 1-mo postexposure recovery period. Clinical pathology evaluations were conducted at the end of the exposure period and after a 1-mo recovery period. At the end of the 4-wk exposure period, tissues from rats were collected, histologically processed, and evaluated by light microscopy. Test substance related, biologically significant decreased body weights and body weight gains occurred in male and female rats exposed to 8000 ppm. In addition, test substance related, statistically significant decreases in food consumption and/or food efficiency were observed in male rats exposed to 8000 ppm. During exposures to 8000 ppm, some rats exhibited tremors and ataxia. Usually tremors and ataxia were observed within 1 h after initiation of the daily exposure period and were observed during each exposure day. Tremors were also observed during 1 exposure day in the 2000 ppm animals. In addition to the tremors and ataxia, rats exposed to 2000 ppm or 8000 ppm had a diminished and/or no alerting response to a sharp, sound stimulus during each of the daily exposure periods. These effects were transient since no clinical observations of compromised neurological function were detected when the rats were evaluated upon return to the animal room following exposure. Daily reoccurrence of this apparently acute effect in the 8000 ppm group did not produce enduring neurological changes since there were no test substance-related effects on FOB parameters or on MA conducted the day following the last exposure or during the recovery period. In addition, there were no toxicologically significant changes in hematology, clinical chemistry, or urinalysis parameters in either males or females for any exposure concentration; and there were no test substance-related gross or microscopic morphological changes in males or females administered any exposure concentration. Under the conditions of the study, the no-observed-effect level (NOEL) was 400 ppm in males and females based on clinical signs of toxicity during exposure to 2000 or 8000 ppm. PMID- 12122565 TI - Physiologically based pharmacokinetic modeling of styrene and styrene oxide respiratory-tract dosimetry in rodents and humans. AB - Styrene (ST) is widely used to manufacture resins, glass-reinforced plastics, and a number of commercially important polymers (Miller et al., 1994). Chronic ST inhalation studies in rodents have demonstrated unique species specificity in the resulting pulmonary toxicity and carcinogenicity. Increased incidences of pulmonary bronchioloalveolar tumors have been observed in mice, but not in rats. No other tumor type was increased significantly in either species. Clara cells lining the respiratory epithelium metabolize ST to styrene 7,8-oxide (SO), which is cytotoxic and weakly genotoxic. Rodent species show marked differences in the distribution and regional density of Clara cells within the respiratory tract, as well as in their capacity to produce and eliminate SO. A mode of action-based physiologically based pharmacokinetic (PBPK) model was developed to predict the concentration of ST and SO in blood, liver, and the respiratory-tract tissues, particularly in terminal bronchioles (target tisue), in order to conduct interspecies extrapolations and determine the extent to which there is a pharmacokinetic basis for the observed species specificity. This PBPK model has a multicompartment description of the respiratory tract and incorporates species specific quantitative information on respiratory-tract physiology, cellular composition, and metabolic capacity. The model is validated against multiple data sets, including blood, liver, and whole lung tissue concentration of ST and SO following multiple routes of exposure. The trend in neoplastic incidences in mice correlated well with model-estimated SO concentration in the terminal bronchioles. The PBPK model predicts a 10-fold lower SO concentration in the terminal bronchioles in rats compared to mice, which is consistent with the observed species sensitivity to the development of respiratory-tract neoplasms. The model-based analysis suggests that humans would be expected to be 100-fold less sensitive to ST-inducted lung tumors than mice, based on pharmacokinetic differences. Pharmacodynamic factors are also expected to contribute to species sensitivity, potentially augmenting pharmacokinetics-based differences. PMID- 12122566 TI - Comparing respiratory-tract and hepatic exposure-dose relationships for metabolized inhaled vapors: a pharmacokinetic analysis. AB - Inhaled vapors that are metabolized locally in the respiratory-tract tissues and systemically in the liver and other organs have different dose-response relationships at the portal of entry compared to systemic target organs. For instance, inhaled chloroform and styrene cause cytotoxicity in the nasal cavity at concentrations much lower that those causing hepatic or renal toxicity. Here, we develop a physiologically based pharmacokinetic (PBPK) model that incorporates a multicompartment, unidirectional flow description of the respiratory tract within a whole-body model in order to estimate both respiratory tract and hepatic metabolism. We then use this model to study the difference in exposure-dose relationship between the respiratory-tract tissues and the liver. The integrated PBPK model confirms that for soluble vapors the exposure-dose curve for metabolism in respiratory-tract tissue will be shifted dramatically to lower concentrations compared to the exposure-dose relationship in systemic organs. This behavior is the result of direct air to tissue equilibration at the portal of entry while other systemic tissues only respond to concentrations in the blood. For cases where metabolism/metabolites of inhaled vapors produce local toxicity, portal of entry effects are expected at lower concentrations and, in general, will be the limiting response for setting reference concentrations (RfCs) for many compounds. The difference in dose-response relationships for metabolism in the respiratory tract versus systemic organs depends on blood/air and blood/tissue partition coefficients and on the degree of systemic extraction of the metabolized vapors. PMID- 12122567 TI - Experience from a long-term carcinogenicity study with intraperitoneal injection of biosoluble synthetic mineral fibers. AB - The carcinogenic potential in the intraperitoneal cavity of three newly developed biosoluble insulation glass wool fibers (M, P, and V) and one newly developed biosoluble insulation stone wool fiber (O) was investigated and compared to that of a previously developed soluble glass fiber (B). The in vitro dissolution coefficient of the three glass wool fibers ranged from 450 to 1037 ng/cm(2) x h and was 523 ng/cm(2) x h for the stone wool fiber. The in vitro dissolution coefficient of the B fiber was 580 ng/cm(2) x h. Groups of female Wistar rats (strain Crl: Wi BR) were exposed by repeated injections to doses of 0.5, 2, and 5 x 10(9) WHO fibers, which corresponds to between 41 mg to 724 mg fiber injected. In addition, 2 groups of crocidolite were used as positive controls at doses of 0.1 x 10(9) and 1 x 10(9) WHO fibers (0.5 and 5 mg). The in vitro dissolution coefficient of crocidolite is estimated to be approximately 1 ng/cm(2) x h. The protocol of the study and the size distribution of the test samples conformed to the European Commission Protocol EUR 18748 EN, and the study was executed under Good Laboratory Practice conditions. Two of the new insulation wools, fibers M and 0, showed no statistically significant tumorigenic response even at the very high dose of 5 x 10(9) WHO fibers injected. Fibers P and V showed a small tumorigenic response in the ip cavity similar in magnitude to the B fiber, which has been declared in the German fiber regulations as a noncarcinogenic fiber. The response to the soluble insulation fibers was notably different from that of the known carcinogen crocidolite, which produced 53% tumors at a comparatively low dose of 0.1 x 10(9) WHO fibers. The incidence of mesothelioma was found to be highly correlated to the incidence of intra-abdominal nodules and masses at different sites. The incidence of abdominal nodules and masses was highly correlated to the number of animals with ascites. The incidence of chronic peritonitis with fibrotic nodules at different organs also correlated with the incidence of mesotheliomas. Differences in etiology were observed between the massive doses of the highly soluble insulation wools when injected directly into the ip cavity and the lower doses of the extremely insoluble fiber crocidolite. The variability in this reaction and the impairment of animal health put into question the value of these massive doses in evaluating the carcinogenic response of soluble insulation wools. All of the fibers tested fulfilled the exoneration criteria with respect to carcinogenicity according to the European Directive 97/69/EC ("an appropriate intra-peritoneal test has not expressed signs of excessive carcinogenicity"). The dose as defined in the EC-Protocol EUR 18748 EN was 1 x 10(9) WHO fibers with a defined geometric spectrum. The influence of fiber dimensions on the ip tumor response and the difficulty in assessing the influence of the difference in background levels between this and previous studies make direct application of the German TRGS 905 criteria difficult; however, by comparison to fiber B, which in the TRGS 905 is considered as a noncarcinogenic fiber, all of the synthetic mineral fiber types tested in this study also appear to meet the intended German criteria for exoneration. PMID- 12122568 TI - Attenuation and recovery of pulmonary injury in rats following short-term, repeated daily exposure to ozone. AB - Controlled human and epidemiology studies have demonstrated that during repeated exposure to ozone (O(3)) attenuation of lung function responses may occur. It is yet unknown whether inflammatory and biochemical effects in lower airways of humans, as observed upon single O(3) exposure, also show a diminutive response following repeated exposure to O(3). The aim of this study was to investigate inflammatory, permeability, and histopathological responses in lungs of rats following repeated daily O(3) exposure and to study the time course of attenuation and recovery of these effects using single O(3) challenges at various postexposure times. To aid in animal-to-human extrapolation, this study and a previously reported human study (Devlin et al., 1997) were designed with similar protocols. Wistar rats were exposed for 5 consecutive days to 0.4 ppm O(3) for 12 h/night. Subsequently, the time course of postexposure recovery was determined by a single challenge of 12 h to 0.4 ppm O(3) after a 5-, 10-, 15-, or 20-day recovery period. Bronchoalveolar lavage (BAL) examination and histopathology were performed 12 h after this O(3) challenge. To quantify the magnitude of the O(3) response, results were compared with a group exposed only once for 12 h to 0.4 ppm O(3) and sacrificed simultaneously. The results demonstrate that a single exposure of 0.4 ppm O(3) causes marked permeability and inflammatory responses in lower airways of rats, as evidenced by enhanced BAL fluid levels of proteins, fibronectin, interleukin (IL)-6, and inflammatory cells. However, 5 days of exposure to 0.4 ppm O(3) for 12 h/night resulted in a complete disappearance of these responses, resulting in BAL fluid values that were not different from those observed in unexposed controls. Postexposure analyses of pulmonary response to O(3) challenges demonstrated that these attenuated responses show a gradual recovery. The data indicate that with respect to BAL fluid levels of albumin, IL 6, and number of macrophages and neutrophils, the period for lung tissue to regain its full susceptibility and responsiveness to O(3) following a 5-day preexposure period is approximately 15-20 days. Remarkably, the total protein and fibronectin responses in BAL fluid still exhibited an attenuated response to an O(3) challenge at 20 days postexposure. Morphometry (number of BrdU-labeled cells in terminal bronchiolar epithelium, and number of alveolar macrophages) showed that after a recovery of 5-10 days following a 5-day preexposure the response to a challenge was identical to that after a single exposure. These results suggest that complete repair from lower airway inflammation caused by short-term, repeated exposure to O(3) may take longer than previously assumed. PMID- 12122569 TI - Upper airway and pulmonary effects of oxidation products of (+)-alpha-pinene, d limonene, and isoprene in BALB/c mice. AB - The oxidation products (OPs) of ozone and the unsaturated hydrocarbons d limonene, (+)-alpha-pinene, and isoprene have previously been shown to cause upper airway irritation in mice during 30-min acute exposures. This study evaluated the effects of OPs and the hydrocarbons themselves on the upper airways, the conducting airways, and the lungs over a longer exposure period. The time course of development of effects and the reversibility of effects were investigated; in addition, we assessed possible exacerbation of sensory responses of the airways to the unreacted hydrocarbons. Respiratory parameters in male BALB/c mice were monitored via head-out plethysmography. Exposures to OPs or hydrocarbons were for 60 min, followed by a 30-min challenge period with air or hydrocarbon, and a 15-min recovery period with air only. Experiments were also performed where limonene/ozone exposures were separated 6 h from the challenge period. Ozone concentration in the reaction mixture was 3.4 ppm, and concentrations of hydrocarbons were 47 ppm (alpha-pinene), 51 ppm (d-limonene), and 465 ppm (isoprene). Due to reaction, the ozone concentration at the point of exposure was less than 0.35 ppm; exposure to 0.30 ppm ozone for 60 min did not produce effects different from air-exposed control animals. As previously established, upper airway irritation was a prominent effect of OP exposure. In addition, over the longer exposure period we observed the development of airflow limitation that persisted for at least 45 min postexposure. All effects from limonene/ozone exposures were reversible within 6 h. Exposures to OPs did not cause enhanced upper airway irritation during challenge with the hydrocarbons, indicating that a 1-h exposure to OPs did not increase the sensitivity of the upper respiratory system. However, airflow limitation was exacerbated in animals exposed to d-limonene alone immediately following exposure to limonene OPs. These findings suggest that terpene/ozone reaction products may have moderate-lasting adverse effects on both the upper airways and pulmonary regions. This may be important in the context of the etiology or exacerbation of lower airway symptoms in office workers, or of occupational asthma in workers involved in industrial cleaning operations. PMID- 12122570 TI - Tumorigenicity of cellulose fibers injected into the rat peritoneal cavity. AB - Cellulose fibers, along with many other organic fibers, are durable. Therefore, if inhaled, they have the potential to persist within the lung, and may then cause disease. Here we report the effects of injecting high-purity cellulose fibers into the abdominal cavity of rats. A respirable fraction of cellulose fiber was collected from an aerosol of a thermo-mechanically-processed wood pulp. A sample of respirable crocidolite asbestos, known to produce mesotheliomas in rats, was used as a positive control. Total doses of 10(6), 10(7), 10(8), or 10(9) WHO fibers were injected intraperitoneally as 3 weekly aliquots. A negative control was provided by phosphate-buffered saline used to suspend the fibers for injection. There were 50 rats per treatment group except for the 10(8) and 10(9) fibers crocidolite groups which were reduced to 26 rats because of the expectation of high tumor incidence in these groups. The two higher doses of crocidolite asbestos caused greatly reduced survival compared to the saline controls. With cellulose there was a much less marked effect on survival. In the highest dose cellulose group, multiple large nodules (granulomas) and widespread adhesions (bands of new tissue connecting organs to each other and to the abdominal wall) were present in all animals. Granulomas were not observed in the 10(9) fibers crocidolite group. More than 80% of animals in the 10(8) and 10(9) crocidolite asbestos groups had mesotheliomas, a type of tumor sometimes observed in people exposed to asbestos. In contrast, there were only 2 animals in the cellulose groups with mesothelioma tumors, 1 in the 10(7) and 1 in the 10(8) groups. However, 9 (18%) of the 10(9) cellulose group had malignant tumors that, in contrast to the usual pattern of mesothelioma development following treatment with mineral fibers in rats, showed no obvious involvement of mesothelial tissues, were not associated with blood-stained ascites fluid, and were thus classified as sarcomas. This study has demonstrated that a high dose of cellulose fibers is capable of producing tumors when injected into the abdominal cavity of rats. PMID- 12122571 TI - Computer simulations of particle deposition in the lungs of chronic obstructive pulmonary disease patients. AB - Epidemiology data show that mortality rates for chronic obstructive pulmonary disease (COPD) patients increase with an increase in concentration of ambient particulate matter (PM). This is not seen for normal subjects. Therefore, the U.S. Environmental Protection Agency (EPA) has identified COPD patients as a susceptible subpopulation to be considered in regulatory standards. In the present study, a computer model was used to calculate deposition fractions of PM within the lungs of COPD patients. The morphology of COPD lungs was characterized by two distinct components: obstruction of airways (chronic bronchitis component), and degeneration of alveolar structure (emphysema component). The chronic bronchitis component was modeled by reducing airway diameters using airway resistance measurements in vivo, and the emphysema component was modeled by increasing alveolar volumes. Calculated results were compared with experimental data obtained from COPD patients for controlled breathing trials (tidal volume of 500 ml, respiratory time of 1 s) with a particle size of 1 microm. The model successfully depicts PM deposition patterns and their dependence on the severity of disease. The findings indicate that airway obstructions are the main cause for increased deposition in the COPD lung. PMID- 12122572 TI - Effects of diesel exhaust enriched concentrated PM2.5 in ozone preexposed or monocrotaline-treated rats. AB - Epidemiological studies have observed statistical associations between short-term exposure to increased ambient particulate air pollution and increased hospital admissions, medication use, pulmonary morbidity, and mortality. To examine the effects of particle air pollution in animals, rats with a preexisting pulmonary inflammation (induced by 1600 microg/m(3) ozone) or hypertension (induced by monocrotaline, MCT) were nose-only exposed to concentrated freshly generated diesel exhaust particles (DEP) mixed with ambient air (CDP). It was hypothesized that a single 6-h exposure to PM exacerbates respiratory inflammatory processes, which affects health parameters in the blood. Histopathology of lung and nose, bronchiolar lavage (BAL), and blood analyses were performed at 1, 2, and 4 days after of the CDP exposure. Morphometry of BrdU-labeled cells in lung and nose was performed at 4 days postexposure. One day after ozone exposure, a mild inflammatory reaction in the centriacinar area was present, consisting of an increase in cellularity of septa and in the number of alveolar macrophages, decreasing in time. Additional CDP exposure did not influence this pattern, except for alveolar macrophages that were loaded with CDP. The only effect seen in the nose after ozone exposure was a slight hypertrophy of the septal mucous cells. Additional exposure to CDP did not change this appearance. MCT-treated rats showed hypertrophy of the media of the pulmonary muscular arteries that was not effected by CDP. BrdU labeling of predominantly Clara cells in the terminal bronchioles was significantly increased after ozone exposure as well as after MCT treatment, whereas this labeling index was markedly enhanced after an additional exposure to CDP. However, no increases in Clara cell protein (CC16) levels were measured of Clara cell protein (CC16) in either BAL or blood. BrdU labeling in the nasal epithelium was not influenced by exposure to ozone or ozone + CDP. CDP exposures did not induce significant toxic effects in the lungs. CDP exposures clearly induced an oxidative stress that was indicated by increasing glutathione levels in BAL with time. In addition, blood fibrinogen levels were enhanced in pulmonary hypertensive rats exposed to CDP. The present study demonstrates that very high CDP concentrations are needed to result in pulmonary changes in animal models with a preexisting pulmonary inflammation or hypertension that continue for days after a single exposure. In addition, CDP has the potential to induce changes in blood. It has not yet been determined how the effects seen with CDP would compare to similar levels of ambient particles. PMID- 12122573 TI - Comparison of chamber and face-mask 6.6-hour exposures to ozone on pulmonary function and symptoms responses. AB - Because of increased interest in an 8-h ozone (O(3)) federal air quality standard, acute pulmonary function responses to prolonged square-wave O(3) exposure between 0.08 and 0.12 ppm have been examined in several U.S. Environmental Protection Agency (EPA) chamber studies. A low-cost face-mask O(3) exposure system was developed in this laboratory and found to produce closely similar pulmonary responses to those observed in prolonged exposures by U.S. EPA investigators. The primary purpose of the present study was to investigate the pulmonary function and subjective symptoms effects of 6.6-h square-wave exposure to 0.12 ppm O(3) by these two methods using the same group of subjects. In addition, further investigation of pulmonary function and symptoms responses upon 6.6-h exposures to lower levels of O(3) (0.04-0.08 ppm) were studied with the face-mask inhalation system. Thirty young adult subjects completed five 6.6-h exposures with six 50-min periods of exercise at an intensity requiring a minute ventilation rate (V(E)) of ~20 L/min/m2 of body surface area, each followed by 10 min of rest, except following 3 h when the rest period was lengthened for a lunch break. The total O(3) doses for the chamber and face-mask exposures to 0.12 ppm O(3) were not significantly different from each other, since the additional O(3) dose during the 35 min lunch break in the chamber exposure was offset by a slightly lower average exercise V(E) (i.e., 19.1 L/min/m2). The data convincingly demonstrated that the two methods of exposing young adults to nearly identical total inhaled O(3) doses at 0.12 ppm produce very similar pulmonary function, symptoms, and exercise ventilatory pattern responses. On the other hand, results of the 6.6-h face-mask exposures to 0.08 ppm O(3) in the present study, compared to similar U.S. EPA exposure study results, revealed several incongruities that may be due primarily to high individual subject variability in responses to a relatively low O(3) exposure. Thus, a comparison of chamber exposure responses to those elicited via face-mask exposure to 0.08 ppm O(3) in the same subject group seems warranted. PMID- 12122574 TI - Rapid communication: effect of inhaled chromium on pulmonary A1AT. AB - A major health hazard to coal miners is development of emphysema following long term exposure to coal dust. One mechanism underlying development of emphysema is the oxidation of critical methionine (Met) residues in antiproteolytic factor, alpha1-antitrypsin (A1AT) resulting in a protease-antiprotease imbalance in the lung. Several studies have documented an association between the incidence and severity of emphysema among miners and their exposure to crystalline silica (i.e., SiO(2)). However, what remains unclear is the role of other co-inhaled nonemphysematogenic nonoxidant inorganic constituent in disease pathogenesis. We hypothesize that in miners, inhaled trivalent chromium (Cr(3+), the only form of Cr in coal) may potentially affect lung A1AT activity in situ via Cr complexing with Met residues, and thereby exacerbate any SiO(2)-induced imbalance. To ascertain if Cr(3+) could, in fact, affect A1AT activity, in vitro studies were done to assess elastase inhibitory activity following A1AT incubation with soluble Cr(3+). In addition, to determine if Cr(3+) found in the lungs as detoxification products of inhaled hexavalent Cr (Cr(6+)) could affect A1AT in situ, lavages from the lungs of chromate-exposed rats were also analyzed for elastase inhibitory activity The in vitro results indicate that Cr(3+) ions clearly inhibited A1AT function, with an IC50 of 1.1 mM being estimated under the experimental conditions used. The in vivo results indicate that long-term inhalation (12 wk or longer) of chromate-bearing atmospheres also gave rise to significant (i.e., 50-70%) inhibition of the antielastase activity of A1AT. Together, these results clearly suggest that the Cr(3+) present in coal dusts could potentially act to inhibit A1AT activity in the lungs of miners and thereby promote the emphysematogenicity of SiO(2) or of other emphysematogens present as coconstituents in these dusts. PMID- 12122575 TI - Human exposure to hydrogen fluoride induces acute neutrophilic, eicosanoid, and antioxidant changes in nasal lavage fluid. AB - The development of asthmalike symptoms among aluminum potroom workers has been associated with exposure to fluorides. In the present study, the immediate nasal response in humans was examined subsequent to short-term hydrogen fluoride (HF) exposure. Ten healthy subjects were exposed to HF (3.3-3.9 mg/m(3)) for 1 h. Nasal lavage (NAL) was performed before, immediately after, and 1.5 h after the end of exposure. Control lavages were performed on the same subjects at the same time points but without HF exposure. At the end of HF exposure, 7 of 10 individuals reported upper airway symptoms. A significant increase was observed in the number of neutrophils and total cells, while there was a decrease in cell viability. The changes in neutrophil numbers correlated significantly with the reported airway symptoms. HF also induced a significant increase in tumor necrosis factor-alpha and the total protein content of NAL fluid. Among the eicosanoids, prostaglandin E(2), leukotriene B(4), and peptide leukotrienes were elevated after exposure. Of the antioxidants measured, the concentration of uric acid increased after exposure. In conclusion, exposure to HF induced immediate nasal inflammatory and antioxidant responses in healthy human volunteers. These findings may contribute to a further understanding of the way HF exerts damage to the airways and show that HF could represent an occupational hazard. PMID- 12122576 TI - Rapid decline in sputum IL-10 concentration following occupational smoke exposure. AB - The acute effects of smoke exposure on inflammatory mediators such as interleukin 10 (IL-10), interleukin-8 (IL-8), and tumor necrosis factor-alpha (TNF-alpha) are not well understood. Our study was designed to measure sputum concentrations of these cytokines in firefighters following low-level smoke exposure. At baseline, participating firefighters underwent blood collection, pulmonary function testing, and sputum induction through inhalation of nebulized hypertonic saline. Study participants later performed overhaul of a structural fire, during which time they wore cartridge respirators and were monitored for smoke exposure. Overhaul involves searching for and extinguishing hidden sources of combustion. One hour following overhaul, blood, pulmonary function data, and induced sputum were again collected. IL-10, IL-8, and TNF-alpha concentrations were measured by enzyme-linked immunosorbent assay (ELISA) in sputum supernatant. In 17 firefighters, baseline sputum IL-10 concentrations were 57.0 +/- 56.8 pg/L, and declined to 16.9 +/- 27.2 pg/L following overhaul (p =.02). No significant changes were observed in sputum IL-8 and TNF-alpha concentrations. Forced vital capacity (FVC) declined significantly in study participants following overhaul. Serum concentrations of Clara-cell protein and surfactant-associated protein A increased significantly following overhaul, indicating increased lung permeability. IL-10 concentrations appear to be exquisitely sensitive to smoke, and studies of IL-10 in sputum should control for recent exposure. Reduced suppression of inflammation by IL-10 may be a mechanism by which low-level smoke exposure causes lung injury. PMID- 12122577 TI - Air pollution and birth weight in northern Nevada, 1991-1999. AB - This study examined the association between particulate matter < or =10 microm in aerodynamic size (PM(10)), carbon monoxide (CO), and ozone (O(3)), and birth weight in Washoe County, NV, from 1991 through 1999. In total, 39,338 single births were included in this study. The mean birth weight was 3383 +/- 460 g and prevalence of low birth weight (LBW) was 2.46% for single births with a gestational age of 37-44 wk. After controlling for cofactors including infant sex, maternal residential city, education, medical risk factors, active tobacco use, drug use, alcohol use, prenatal care, mother's age, race and ethnicity of mothers, and weight gain of mothers, we found PM(10) exposure in the third trimester of pregnancy to be a significant predictor of birth weight of newborns. A 10-microg/m(3) increase in the 24-h PM(10) level in the third trimester of pregnancy can be associated with a birth-weight reduction of 11 g (95% CI: 2.3 19.8 g) using multiple linear regression; however, PM(10) was not found to be related with the risk of LBW from logistic regression. CO and O(3) were not found to be associated with birth weight or risk of LBW of newborns by the same modeling procedure. The results suggest PM(10) could be a risk factor associated with birth-weight reduction of newborns in urban northern Nevada; however, the current level of PM(10) is not a risk factor to increase the chance of LBW newborns. PMID- 12122578 TI - Particulate matter inflammation and receptor sensitivity are target cell specific. AB - The complexity of primary source particulate matter (PM) and the various cell types encountered by its inhalation raise the possibility that target cells are differentially activated. Since epithelial cells, which line the nasal-tracheal bronchial airways, and sensory C fibers, which terminate throughout this epithelial layer, are initially targeted by inhaled PM, we compared their relative biological response in vitro to PM originating from volcanic (MSH), anthropogenic (diesel), residential (woodstove), urban ambient (St. Louis, Ottawa), and industrial emission (coal fly ash, CFA; residual oil fly ash, ROFA; oil fly ash, OFA) sources. Increases in intracellular calcium (i.e., [Ca(2+)](i)) are a second-messenger event that indicates cellular activation and signal transduction, in both nerve and epithelial cells. Single-cell calcium imaging recordings were taken of human bronchial epithelial cells (BEAS-2B) exposed to selected PM (50 microg/ml or 30 microg/cm(2)). These cells responded with variable increases in [Ca(2+)](i) ranging from abrupt increases, which returned to baseline upon washing of the cells, to oscillations of the [Ca(2+)](i) that did not wash out. Increases in [Ca(2+)](i) and inflammatory cytokine (i.e., interleukin 6, IL-6) release were measured in populations of BEAS-2B cells exposed to PM (50 microg/ml) and were shown to significantly correlate (r(2) =.80). BEAS-2B cells, stained histochemically with cobalt, displayed a concentration-dependent precipitation in response to acid pH and capsaicin, indicating the presence of acid-sensitive pathways (e.g., VR1 and acid-sensitive receptors). To demonstrate the relevance of these pathways to inflammatory cytokine (i.e., IL-6) release, BEAS-2B cells were pretreated (15 min) with antagonists to the vanilloid (VR1) receptor (i.e., capsazepine, CPZ) or acid sensitive pathways (i.e., amiloride) before their exposure to the selected PM. A significant reduction of IL-6 release occurred in response to all PM, except for MSH and diesel exhaust. Dorsal root ganglia (DRG), which innervate the tracheal airways, were dissociated from fetal mice and pretreated with CPZ or amiloride before exposure (4 h) to the selected PM (50 microg/ml). Overall, significantly higher release occurred in PM-exposed sensory neurons relative to that of BEAS-2B epithelial cells. Although both CPZ and amiloride significantly reduced IL-6 release for all PM, the degree of inhibition was less for the PM-exposed DRG relative to BEAS-2B cells. These data show that differential increases in [Ca(2+)](i) and IL-6 release occur in BEAS-2B epithelial cells and DRG sensory neurons, when exposed to PM derived from different sources. The degree of this activation, however, depends not only on the source of the PM, but also on its cellular target. This differential sensitivity of target cells may contribute to the organism's overall inflammatory response to PM exposure. PMID- 12122580 TI - Electrophysiological and histopathological evaluation of respiratory tract, diaphragm, and phrenic nerve after dichlorvos inhalation in rats. AB - The aim of this study was to investigate the dose-related effects of dichlorvos inhalation on electrophysiological alterations of diaphragm and phrenic nerve and the changes in the histologic structure of respiratory system. This study was performed on 33 rats divided into 5 groups, inhaling 1, 2, 5, 10, and 15 microg/L of dichlorvos, respectively. Electrodiagnostic investigations of diaphragm and phrenic nerve were made before and after inhalations. Aspiration samples were taken from lungs to evaluate the presence of infection agents. The airways, lungs, and diaphragms were dissected out for histologic investigation. Rats exposed to a low concentration of dichlorvos (1-5 microg/L) showed no symptoms of intoxication, but exposure to higher doses (10-15 microg/L) induced dyspnea in several animals. Lower doses of dichlorvos revealed no electromyographic changes on diaphragm, whereas higher doses revealed a clear neuropathic involvement. Delayed phrenic nerve motor conduction velocity was noted for each group (p <.05). Morphologic changes on the tracheal epithelium, hyperplasia, thickening of the blood-air barrier, degeneration in alveoli, and ductus alveolaris were seen in histopathologic investigation. In conclusion, the acute inhalation of dichlorvos caused clear evidence of neuropathic involvement of the diaphragm and the phrenic nerve. Also, toxic pneumonitis and injury to the tracheal epithelial were noticed. PMID- 12122579 TI - 9-nitrocamptothecin liposome aerosol: lack of subacute toxicity in dogs. AB - 9-Nitrocamptothecin (9-NC) dilauroylphosphatidylcholine (DLPC) liposome aerosol was evaluated for potential toxicity in an 8-wk, subacute toxicity study in dogs. Fourteen adult dogs were divided into 2 groups with 10 animals in the 9-NC-DLPC treatment group and 4 animals in the DLPC-only vehicle control group. 9-NC-DLPC was administered to the animals using an Aerotech II nebulizer flowing at 10 L/min. Full-face exposures for 60 min were conducted for 5 consecutive days a week for 8 wk. The estimated deposited aerosol dose was 24.7 microg/kg/day. Animals in the vehicle control group received aerosolized DLPC only. Body weight, food consumption, urinalysis, in-life observations, hematology, plasma chemistry, and necropsy and histopathology were monitored before and during the treatment period and in a subset of animals for 2 wk following the end of treatment. Animals were observed for signs of pharmacologic and/or toxicologic effects three times on days of dosing and once daily on nondosing days. 9-NC-DLPC liposomes administered as a small-particle aerosol were determined to be nontoxic to dogs when given for 5 days/wk, for a duration of 8 wk. DLPC-only liposome also had no toxic effects. PMID- 12122581 TI - [The Shouldice repair for inguinal hernia--technique and results]. AB - The technique of the Shouldice hernioplasty for primary and recurrent inguinal hernia is described on the basis of more than 3 500 procedures at the Surgical University Clinic of Aachen and 700 operations performed at the Surgical Department of the Park-Klinik Berlin-Weissensee. Local anesthesia is the preferred type of anesthesia with a good acceptance by the patients site (96 %). In compliant adults nearly all primary and about 50 % of the recurrent hernias can be repaired under local anesthesia. Essentials of the preparative phase are identification and preservation of the inguinal nerves, resection of the cremaster muscle and exposure of all three hernial sites (lateral, medial and femoral). For all primary hernias and indirect or small recurrent hernias a modified two-layer Shouldice repair of the transversalis fascia using a monofilament running suture (Polypropylene 0) is recommended. In the early postoperative period the physical activity is limited only by the patients complaints. Normal activity is gained back within 2 to 3 weeks. In a non selected group of patients 10-years recurrence rates are 2.6 % in primary hernia and 6.9 % in recurrent repair. The Shouldice repair is recommended as the procedure of choice for all primary and selected recurrent hernia. A mesh prosthesis may be necessary in large medial and combined recurrent hernia. PMID- 12122583 TI - [Meshes in inguinal hernia repair]. AB - Repair of a groin hernia is strongly influenced by prosthetic mesh implantation carried out in nearly 50 % of all operations. Recurrency rates, however, did not decrease by this policy. Many different materials are available. Due to bioinstability on the long-term and elevated infection rates PTFE is not suitable for inguinal hernia repair. Polyester also provides no long-term stability and induces a chronic foreign body reaction. Polypropylene initially leads to an acute inflammatory reaction and often ends in fibrosis. Both reactions are related to the weight of the used mesh. All materials may lead to specific complications. These include seroma formation, infection, migration of the prosthesis with arrosion of organs, damage of the vas deferens, development of recurrency by shrinkage of the fibers around the mesh, formation of adhesions in the preperitoneal position, and chronic inguinal pain. Therefore, meshes should be used only after individual estimation of risks and benefit. This includes the hernia classification, the number of previous operations and the possibility of a defect in collagen metabolism. The unknown long-term risks for the patient may be taken only in strong indications. PMID- 12122582 TI - [Recurrent hernias after previous Shouldice operation]. AB - For all 158 surgical operations performed on hernia recurrences throughout the period from March 2000 until the end of May 2001, we compared the intra-operative findings to the information contained in the operation reports--as far as available--as part of our quality management. In less than 20 % of the patients for whom a Shouldice repair had been documented in the operation reports, we found evidence of the actual performance of a Shouldice repair (typical cicatrised modifications on the rear wall or the fascia transversalis, sutures or residues of sutures). 74 % of the patients were treated with a Marlex(R) Perfix plug, avoiding the resection of stable cicatrisation fractions with incision of the rear wall in the case of an intact fascia. On 26 % of the patients it was possible to perform a Shouldice repair in compliance with the original technique. Meanwhile, mesh techniques have outpaced the Shouldice technique with respect to the recurrence rates in the efficiency statistics. This, however, is not caused by the technique as such, but rather by the fact that in many clinics the anatomical situations are obviously incorrectly assessed and/or that insufficient knowledge about suturing techniques prevails. As a consequence, worse results are reported for the Shouldice technique than for the mesh techniques. It is not the Shouldice technique that is insufficient but its performance suffers in many hospitals from substantial insufficiencies in terms of quality. PMID- 12122584 TI - [PHS--a double-leaved mesh in the open surgery of inguinal hernias]. AB - The double-leaved mesh (Prolene Hernia System) represents a combination of tension-free meshes: the ovoid onlay component lies in position like the patch used in the Lichtenstein - procedure and the circular underlay component in position of techniques with posterior approach. The leaves of the mesh are joined by a central pin (connector), which completely closes the dilated internal ring and the hernial ring, respectively like a plug used in the Rutkow procedure. In a prospective study we investigated the duration of operations, complications, postoperative consumption of analgetics, hospitalisation time, return to normal activity, return to work and patients satisfaction. Between October, 1999 and June, 2001 105 patients with 110 inguinal hernias were treated with a double leaved mesh in an open technique in local or general anesthesia. There were no intraoperative complications. Postoperative complications included 2.7 % seromas, 1.8 % haematomas, 1.8 % infections, 2.7 % neuralgias and 1.8 % testicular atrophies. Follow up examinations over an average time period of 13 months (follow-up-rate 70 %) did not show any recurrences. We conclude that groin hernia repair with this double-leaved mesh is easy to learn and to perform. The procedure is safe and efficient and can be carried out under local anesthesia. PMID- 12122585 TI - [Rectusbanding by polypropylene-mesh--a new method for incisional hernia repair]. AB - We report on a new method of incisional hernia repair applicable to any size. With exception of an area 1 cm medial of the m. rectus abdominis where the strip penetrates the anterior wall of the rectus sheath for a better fixation, the muscle ist looped in a loose way with a 2 cm wide band of polypropylene (Prolene(R)) on both sides at a distance of 2 cm depending on the size of the hernial opening. Different models of strips were used in 27 % of the cases adapted to anatomical particularities of the hernial opening. Anterior and posterior wall of the rectus sheath are closed by a continuous panacryl suture which covers the strip. Because of the wide subcutaneous excavation extending to the lateral margin of the rectus sheath an extensive drainage by Redon-Drainages as well as compression bandages are important therapeutical procedures until formation of seromas has finished. For perioperative antibiotic prophylaxis we used Cefuroxime (3 x 1,5 i. v.). From 07/1999 until 10/2001 75 patients underwent an operation in our department. The direct postoperative complications observed were: Seroma formation up to 300 ml after discharge in 5 patients (6,6 %) and wound infections in 2 patients (2,8 %). In none of the cases the mesh had to be removed. In a follow-up period of 6 to 24 months we found 2 recurrences in 60 patients (3,3 %). These were related to technical failures of the beginning. 64.9 % of the patients were free of complaints after 6 months and almost 96 % after one year. Only 3 patients (4 %) had to take analgetic drugs occasionally. With regard to the mobility of the abdominal wall we found no measurable limitation. The method of Rectusbanding is easy to learn for every surgeon and with little material the mesh-strip can be fixed safely. It can be cut to individual sizes and shapes adapted to the fascial proportions of the hernial opening. PMID- 12122586 TI - [Influence of resterilized polypropylen meshes on growth of human fibroblasts--an experimental in vitro study]. AB - INTRODUCTION: We investigated the influence of resterilized polypropylen meshes (Prolene(R)) on proliferation and apoptosis of human fibroblasts in an experimental in vitro study. METHOD: Human fibroblasts were seeded into six-well culture dishes in a density of 3 x 10 (4) cells/well. After resterilization of meshes (steam autoclave, 121 degrees C, 20 min.) according to the manufacturer's recommendations (Ethicon, Norderstedt) square sheets of 2 x 2 cm were incubated with fibroblasts over a period of 6, 12, 18, 24, 30, 36, 42 and 48 h. Preparations of fibroblasts with non-resterilized meshes and without meshes served as controls. Proliferation index and apoptotic index were estimated by flow cytometry after cell staining with an FITC-conjugated antibody against the Ki-67 antigen or with FITC-conjugated Annexin-V and propidium jodide, respectively. RESULTS: A significant reduction of the proliferation index from 86 % to 42 % was found after 48 h incubation of cells with resterilized meshes, whereas only a slight decrease was found in the group with non-resterilized meshes (75 %) and in controls without meshes (80 %). Apoptotic index increased significantly from 2 % to 48 % after 48 h incubation with resterilized meshes in comparison to both control groups, where only a slight increase could be observed: non-resterilized meshes to 19 % and without meshes to 10 %. CONCLUSION: Resterilized meshes inhibit growth of human fibroblats in vitro significantly, demonstrated by a reduced proliferative activity and an increased apoptotic index. This could be caused by a release of toxic substances from the meshes, which have a negative influence on cell growth. Therefore, resterilization cannot be recommended. PMID- 12122587 TI - [Surgical management of severe peritonitis]. AB - Despite all the efforts made in the areas of intensive care and surgery, severe peritonitis remains a feared condition that is associated with a high mortality rate. Severe abdominal infections are accompanied with a high level of endotoxin production, resulting in the so called systemic inflammatory response syndrome (SIRS), which is often complicated by multiple organ failure. The surgical eradication of the infectious focus is the most important prerequisite for a successful treatment. According to the severity of the local inflammation, different forms of abdominal lavage can be applied. We analysed patient characteristics and the clinical outcome of 180 patients with diffuse peritonitis, including 36 patients with more than 29 MPI-Points. The mean severity of peritonitis (n = 36) was 33 using the Mannheim Peritonitis Index (MPI). The hospital mortality rate was 58 % in this group. PMID- 12122588 TI - [Indications, techniques and results of laparoscopic surgery for diaphragmatic diseases]. AB - Eleven patients with congenital, traumatic and functional extrahiatal diaphragmatic lesions are reported. Since 1991 two patients with acute, two patients with old ruptures of the diaphragm and one patient with a Morgagni Larrey-hernia were successfully treated by laparoscopic direct suturing. In two other patients with Morgagni hernias we used a polypropylene mesh for closure of the defect. One procedure was performed in a patient with symptomatic congenital dysplasia of the diaphragm with aplasia of the pericard. Laparoscopic plication of the diaphragm was performed in three symptomatic patients with phrenic nerve palsy after cardiac surgery. The intra- and postoperative course was uneventful in all cases. During a median follow-up of 60 months there was no recurrence. Therefore the laparoscopic technique is an effective and attractive alternative for treatment of these diseases. PMID- 12122590 TI - [Invited commentary]. PMID- 12122589 TI - [Laparoscopic Collis-Nissen operation for shortened esophagus]. AB - Esophageal shortening as a complication of advanced gastroesophageal reflux disease is seen in 2-4 % of patients with GERD. The Collis gastroplasty with Nissen fundoplication creates an intraabdominal neoesophagus with a fundic wrap and is an effective procedure in the difficult intraoperative situation of esophageal shortening. After own experience with the open Collis gastroplasty and after reports on successful laparoscopic Collis-Nissen operation we performed 2 laparoscopic Collis-Nissen operations. The technique we used is a new modification of the laparoscopic technique described by Johnson and Hunter in 1998. The postoperative course was uncomplicated in both patients. The follow up included endoscopy and barium meal. The patients showed relief from reflux symptoms except a mild dysphagia. There was no persisting esophagitis. The laparoscopic Collis-Nissen gastroplasty is an effective minimal-invasive procedure in patients with shortened esophagus diagnosed intraoperatively. PMID- 12122591 TI - [Laparoscopic adrenalectomy--experiences with transperitoneal approach]. AB - INTRODUCTION: We report our results of laparoscopic anterior transperitoneal adrenalectomy. PATIENTS: Between 4/1996 to 05/2001, a laparoscopic adrenalectomy was performed in 34 patients (median age 48 years). The adrenalectomy was performed transperitoneally (31 unilateral; 3 bilateral). The adrenaline level was measured in 7 patients with a pheochromocytoma. RESULTS: All tumors (mean size 3.5 cm; 0.4 to 8.0 cm) could be extirpated by laparoscopy. 9 pheochromocytomas; 9 cortisol producing tumors (one patient with a Carney's syndrome); 7 Conn's adenomas and 9 incidentalomas constituted these tumors. In the first third of the observation period, the surgery lasted 176 (95-270) minutes, in the last third 82 (50-130) minutes (p < 0,01). We postoperatively observed the following complications: one abdominal wall hematoma at a port-site and one edematous pancreatitis after alteration of the pancreatic tail. The adrenaline level continually rose from the beginning of surgery to the ligature of the suprarenal vein. CONCLUSION: Transperitoneal adrenalectomy in benign tumors (< 8 cm) is our method of choice. The resulting learning curve allowed the performance of adrenalectomy within an acceptable operative time and without significant blood loss. The transperitoneal technique is safe and well reproducible. The cosmetical results are convincing. We recommend an early ligature of the suprarenal vein in a pheochromocytoma. PMID- 12122592 TI - [Which factors are responsible for postoperative mortality in colorectal cancer patients?]. AB - INTRODUCTION: The postoperative decease is the most severe complication in surgery. PATIENTS AND METHOD: Within the framework of a multicentre study between January 1 (st) and December 31, 1999, 3,756 patients, 1,463 of them suffering from rectal cancer and 2 293 from colon carcinoma, from 75 clinics were documented with the help of a standardized questionnaire. We compared data of 211 patients who died postoperatively with data of 3,484 patients who survived after surgical treatment of colorectal cancer. Logistical regressions, under inclusion and exclusion of intra- and postoperative complications, show independent influence factors on the postoperative decease and provide models for the prediction of the postoperative death. RESULTS: Compared to the patients who survived, the postoperative deceased patients were significantly older. They had a poorer risk profile and therefore a higher ASA-score (p < 0.001). 20.4 % of the patients underwent an emergency operation. General and specific postoperative complications occurred significantly more frequently. The model of a logistical regression allowed the prediction of postoperative decease with a sensitivity and specificity of 91 %. General postoperative complications such as pulmonary embolism (relative risk: 30.3), cardiac (relative risk: 24.1), renal (relative risk: 22.1), and pulmonary complications (relative risk: 12.0) are crucial for lethality. DISCUSSION: The postoperative decease is influenced by several factors. It is impossible to reduce the number of influence factors for the prediction of outcome. The general postoperative complications, however, represent a crucial problem. It is important to avoid these problems in order to reduce postoperative lethality. PMID- 12122593 TI - [Sigmoidoperianal fistula associated with diverticulitis and Cul de Sac situation -diagnostics and therapy]. AB - We report on the case of a 64-year-old female patient who presented herself in our outpatient clinic because of a perianal fistula with recurrent abscesses. We describe the step diagnostics and the surgical treatment of the causal sigmoido perianal fistula with diverticulitis and Cul de sac situation. Clinical examination, fistulography, colonoscopy and MRT were part of the precise representation and preparation for the high anterior rectosigmoidal resection with simultaneous rectopexy according to Sudeck which were performed without complications. The sigmoidoperianal fistula must be taken into account as a differential diagnosis of a recalcitrant high perianal fistula. PMID- 12122594 TI - [Inflammatory alterations of the greater omentum--a difficult preoperative diagnosis]. AB - Isolated inflammatory alterations of the greater omentum are rare diseases. Since the preoperative diagnosis is difficult these changes are usually ascertained upon laparotomy. Between 1999 and 2001 four patients (44-88-years-old, 2 males) underwent laparotomy for an inflammatory tumor mass or an acute abdomen, respectively. C-reactive protein was elevated in all cases. In two cases a primary torsion of the greater omentum was found, in the other two a paracolic pseudotumorous omentitis. Resection of the diseased omental parts led to complete recovery. Torsion of the greater omentum and omentitis cause abdominal symptoms with an inflammatory component that often mimics other more common diseases such as acute appendicitis and urges laparotomy. Partial omentectomy is the therapy of choice. PMID- 12122595 TI - [Pneumoperitoneum as occasional finding in chronic dialysis patients--laparotomy or conservative non-operative approach?]. AB - Free air of unknown origin within the abdominal cavity is a serious problem, which in the majority of cases indicates the perforation of a hollow organ. In two cases, we report on i) detection of free air subdiaphragmatically by coincidence during follow-up investigation of an interstitial pulmonary disease (chest X-ray) in a 67-year old patient with chronic renal insufficiency, and ii) diagnostic of pneumoperitoneum (3 times as primary diagnosis) in a 63-year old multimorbid female (with chronic renal insufficiency) with recurrent, but unspecific epigastric symptoms over a time period of 5 years. The following investigations such as endoscopy, contrast enema, and abdominal ultrasound did not detect a perforation as most likely cause. The first patient was discharged after clinical observation, laboratory and ultrasound follow-up for 5 days. In the second case, neither explorative laparoscopy during the second clinical observation period nor laparotomy for required cholecystectomy because of cholecystitis could appropriately clarify the origin. In conclusion, the detection of a pneumoperitoneum in asymptomatic patients or subjects with unspecific abdominal symptoms requires always clinical monitoring and instrumental diagnostic, consisting of endoscopy in the upper gastrointestinal tract, contrast enema of the colon and abdominal and/or thoracal computed tomography, to definitely exclude perforation. In addition, ultrasound as third column detects early low amounts of fluid and is the suitable method for short term follow-up. The cause of pneumoperitoneum, particularly in asymptomatic patients, can not be found in every case. Under these circumstances, non operative treatment is favored. PMID- 12122596 TI - TH1/TH2 serum cytokine profiles and soluble TNF-receptor response in patients with chronic hepatitis C during recombinant human interleukin-12 (rHuIL-12) treatment. AB - Interleukin-12 (IL-12) has many antiviral properties both in vitro and in vivo, such as enhancing cytotoxic lymphocyte reaction, promoting Th1-helper cell response and inducing interferon-gamma - (IFN-gamma) production. The present study investigated possible antiviral effects of recombinant human IL-12 in vivo in 11 chronic hepatitis patients treated with rHuIL-12 subcutaneously in 3 different dosages (0.03, 0.1, 0.5 microg/kg) once weekly for a period of 10 weeks. ELISA was employed to determine before onset of therapy the serum concentrations of IL-12, IL-12p40, IFN-gamma, IL-4, IL-10, tumor mecrosis factor alpha (TNF-alpha), TNFR p55 and p75, as well as on days 3, 5, 8, 10, 12, 15, 17, 22 after onset of therapy and subsequently weekly until the end of the treatment period and at the end of the 12 months' postobservatory period. The HCV-RNS serum concentration was determined by means of an RT-PCR method. Two to three days after the subcutaneous rHuIL-12 injections there was a transient significant rise of n the serum concentrations of IL-12, IL-12p40, IFN-gamma and TNFR p55, followed by a decrease to the original level. The increases of the IL-12 and IFN gamma serum concentrations were dose-dependent. In patients where there was a decline of the HCV-RNS concentration in the serum we confirmed a trend to higher IL-12 serum concentrations. The serum levels of IL-2, TNF-alpha and IL-10 fluctuated only during the treatment period. The present study showed that a once a-week injection of rHuIL-12 results in a merely transient rise of the IL-12, IL 12-p40- and IFN-gamma serum concentrations. This may offer an explanation for the unsatisfactory clinical efficiency of the substance. PMID- 12122597 TI - Polymorphisms of the carcinogen detoxifying UDP-glucuronosyltransferase UGT1A7 in proximal digestive tract cancer. AB - Cancer of the proximal digestive tract is associated with tobacco smoke and ethanol exposure. The UDP-glucuronosyltransferase (UGT) 1A7 is a detoxifying enzyme capable of tobacco-borne carcinogen detoxification and cellular protection and has been implicated as a cancer risk gene. In this study, UGT1A7 expression is demonstrated in oral, esophageal, and gastric tissue, which are the principle sites of proximal digestive tract cancer. Genomic DNA from the blood of 76 patients with esophageal, orolaryngeal and gastric cancer as well as from 210 healthy blood donors was analysed for the presence of UGT1A7 polymorphisms by sequencing and temperature gradient gel electrophoresis. Wild type UGT1A7 alleles were equally distributed between controls (19 %) and cancer patients (22 %). However, the UGT1A7*3 allele combining W208R, N129K and R131K missense mutations and exhibiting substantially reduced carcinogen detoxification activity was significantly associated with proximal gastrointestinal cancer and identified as a risk allele present in 32 % of cancer patients and 19 % of controls (P = 0.0008, OR 2,02 (95 %-CI 1.33-3.07)). We identify the significant association of the UGT1A7*3 allele encoding a low catalytic activity protein as a risk gene in proximal digestive tract cancer and as a potential marker for cancer susceptibility. PMID- 12122598 TI - Metallic stents and plastic endoprostheses in percutaneous treatment of biliary obstruction. AB - BACKGROUND: In the light of the clinical controversy whether metallic stents or plastic endoprostheses should be used in the percutaneous treatment of biliary obstruction we retrospectively evaluated our experience with both drainage systems. METHODS: 71 patients (mean age 68 +/- 12 years) underwent a total of 81 interventions and received either plastic endoprostheses (11.5 or 12 French diameter; N = 57/81) or metallic stents (N = 24/81). RESULTS: Drainage insertion was technically successful in all of the 71 patients. There was no procedure related mortality, but a 30-day mortality of 15 % (N = 11). Overall, 27 complications occurred in 81 interventions with a statistically significant higher complication-rate in plastic endoprostheses (39 %; N = 21/54) compared to metallic stents (22 %; N = 6/27). The average patency of the drainage-systems was 166 +/- 341 days (range 1-2,705 days) and did not differ significantly between the drainage-subtypes. Incidence of complications and a further increase in serum bilirubin following intervention was associated with a higher drainage occlusion rate and reduced survival, irrespective of the drainage-system used. CONCLUSIONS: Percutaneous transhepatic treatment of biliary obstructions with internal drainages is a reliable therapy. The overall complication-rate of metallic stents was lower compared to plastic endoprostheses; however, no significant differences were found with respect to drainage patency or success-rate. PMID- 12122599 TI - [Gastric outlet obstruction in chronic granulomatous disease]. AB - We report the case of a three year old dystrophic boy who suffered from vomiting, loss of weight and fever. In the history there were several episodes of severe infections which had repeatedly lead to hospitalisation. The cause of the actual disorder was a gastric manifestation of chronic granulomatous disease, which led by an inflammatory thickening of the gastric wall to a gastric outlet obstruction. Treatment with prednisone and gamma interferon normalised the thickness of the gastric wall and gastric outlet function. A prophylactic treatment with antibiotic and antifungeal agents was started. During the following 13 months no further severe infections were observed. We discuss incidence, kinds of manifestation, diagnostics and therapeutical options of the disease and give an overview of the literature. PMID- 12122600 TI - Endoscopic therapy for Zenkers's diverticulum by means of argon plasma coagulation. AB - We describe a 80-year-old man who presented with progressive dysphagia because of a Zenker's diverticulum. Barium swallow study revealed a large posterior diverticulum with a distal stenosis of the esophagus caused by compression. Because the patient was a poor candidate for surgery an endoscopic therapy was performed. The Zenker bridge was divided by argon plasma coagulation in two sessions without any complication to allow an overflow. The patient remained asymptomatic to date for a follow-up of 6 months. PMID- 12122601 TI - [Gastrocolic fistula - a rare cause of cachexia and polyneuropathy]. AB - Gastrocolic fistula is a rare clinical disorder which in the past most often occurred after gastric surgery or carcinoma of the gastrointestinal tract. However, during the last decade an increasing number of cases after benign gastric ulcers have been described. Most common symptoms have been weight loss, abdominal pain, diarrhea and copremesis. A 49-year-old cachectic patient presented with a 2-year history of abdominal discomfort and diarrhea. He reported a weight loss of 32 kg during this period and was finally unable to move because of exhaustion. Furthermore, he suffered of burning paresthesia of the legs and the abdomen. His medical history included a Billroth II operation because of recurrent ulcer disease in 1987. Barium enema revealed a gastrocolic fistula which caused small bowel bacterial overgrowth with villous atrophy and malabsorption and development of polyneuropathy. The fistula was surgically resected, and postoperatively, the patient improved and regained his weight. Gastrocolic fistula is a rare cause of diarrhea and should be considered in clinical practice. Barium enema is superior to endoscopy in detecting gastrocolic fistula. PMID- 12122602 TI - Role of the intestinal epithelium in orchestrating innate and adaptive mucosal immunity. AB - The mucosa that lines the human colon and small intestine is a site of chronic regulated "physiologic" inflammation. This contrasts markedly with other mucosal sites in that if the numbers of T and B cells, eosinophils, mast cells, macrophages, and dendritic cells that are present in the human intestinal tract were to be present in other sites, those sites would be considered to be sites of chronic pathological inflammation. This review examines the role of the intestinal epithelium in the development of "physiologic" intestinal mucosal inflammation and focuses on its role in signalling and mediating host innate and adaptive mucosal immune responses. PMID- 12122603 TI - [Reprocessing of flexible endoscopes and endoscopic accessories - an international comparison of guidelines]. AB - Endoscopic examinations and procedures are essential for diagnosis and treatment of gastrointestinal diseases. As a result of poor reprocessing practice microorganisms can be transmitted via endoscope. The majority of infection transmissions is due to insufficient performance of cleaning and disinfection disregarding guidelines of societies of gastrointestinal endoscopy. A review of the literature and a comparison of European and American guidelines for reprocessing flexible endoscopes are given. Differences in the classification of endoscopic devices, on the possibility of prion transmission, recommendations on staff training and protection, quality assurance of reprocessing and evidence based graduation of guidelines are stressed and discussed. With respect to the procedure of endoscope reprocessing, differences concerning the cleaning solution to choose, necessity of thoroughly manual cleaning and brushing of the accessible endoscope channels (even in the case of subsequent automatic reprocessing endoscopes in washers-disinfectors), disinfection solution, microbiological quality of water for final rinsing and rationale for alcohol flush of endoscope channels for better drying are mentioned. The need for experimental investigations of the cleaning and disinfection process is stressed. In contrast to recent guidelines of European and American societies of gastrointestinal endoscopy, the now updated recommendations of the Robert Koch-Institute for reprocessing flexible endoscopes and endoscopic accessories are evidence-based and graduated. PMID- 12122604 TI - [Role of G-protein-coupled adenosine receptors in downregulation of inflammation and protection from tissue damage]. PMID- 12122605 TI - [Eradication of helicobacter pylori: symptomatic treatment of reflux disease?]. PMID- 12122606 TI - [Endoscopic ligation compared with combined treatment with nadolol and isosorbide mononitrate to prevent recurrent variceal bleeding]. PMID- 12122607 TI - Clinical significance of changes in electrocardiographic R-wave voltage on chest leads in patients with acute anterior myocardial infarction. AB - This article aims to clarify the clinical significance of changes in electrocardiographic (ECG) R-wave voltage on chest leads from 1 to 4 weeks in patients with acute anterior myocardial infarction (MI) in combination with echocardiographic findings and dual scintigraphic findings. Seventy-one patients with acute anterior MI who underwent emergency revascularization were subjected to ECG and echocardiography, at both 1 and 4 weeks, and to thallium-201 (TI) and iodine-123-beta-methyl-p-iodophenyl pentadecanoic acid (BMIPP) single-photon emission computed tomography (SPECT) about 1 week after the onset of MI. The total sum of ECG R-wave voltage on each chest lead was calculated. The mean defect ratio on TI and that on BMIPP derived from circumferential profile curve analysis were calculated. The percentage defect-discordant ratio of both SPECT images [(%) discordance on TI/BMIPP] was obtained. The percentage increase ratio of ECG R-wave voltage on chest leads [(%) increase of R wave] and the increase of left ventricular ejection fraction (DeltaEF) from 1 to 4 weeks were obtained. There were significant correlations between the (%) increase of R wave and the DeltaEF as well as between the (%) increase of R wave and the (%) discordance on TI/BMIPP (r =.63, P <.001; r =.74, P <.001). The reversibility of ECG R-wave voltage was related to cardiac functional improvement in addition to the discordance on the 2 images. Monitoring of changes in ECG R-wave voltage on chest leads is useful to detect the presence of myocardial viability and to evaluate functional evolution in patients with acute anterior MI. PMID- 12122608 TI - Dipyridamole body surface potential mapping: noninvasive differentiation of syndrome X from coronary artery disease. AB - Up to 20% to 30% of patients with angina and abnormal stress test have normal coronary arteries at angiography or syndrome X (Sy X). We tested whether body surface potential mapping (BSPM) with intravenous dipyridamole could differentiate patients with Sy X from patients with coronary artery disease (CAD). We compared the effects of intravenous dipyridamole (0.28 mg/kg over 4 min) on BSPM in 17 healthy volunteers (controls) and in 2 groups of patients with angina and abnormal ergometric tests who were referred for angiography: 27 patients with obstructive disease (> or =70% diameter stenosis) in the CAD group, and 17 patients with Sy X. Control subjects were easily differentiated from patients with CAD or Sy X by markedly smaller baseline BSPM DeltaST-T < or = LSD departure areas (P <.001), but the Sy X and CAD groups had similar ST-T departure areas. The average potential integral difference after dipyridamole (APID) differentiated Sy X and CAD patients: the mean APID increased in patients with Sy X and trended negative in the CAD group. The APID(20%-40%) (integrated over 20% to 40% of the ST-T interval) mean value was 0.59 +/- 0.67 microVs in the Sy X group and -0.18 +/- 0.59 microVs in the CAD group (P <.01). At a threshold APID(20%-40%) > 0.17 microVs, the sensitivity and specificity for Sy X was 71% and 78%, respectively; the area under the receiver operating characteristic curve was 0.79 (95% CI 0.64, 0.93). Dipyridamole BSPM is a promising noninvasive diagnostic modality to differentiate patients with Sy X from those with CAD. PMID- 12122609 TI - Myocardial ischemia adjacent to infarction enhances the magnitude of exercise induced ST-segment elevation one month after myocardial infarction. AB - The aims of this study were to compare exercise-induced ST-segment elevation with and without ischemia and to examine the relation between exercise-induced ST segment elevation and the location of myocardial ischemia. Seventy-nine patients with first anterior myocardial infarction underwent thallium-201 exercise myocardial scintigraphy test one month after myocardial infarction. There were 37 patients showing no reversible defect (non ischemia group), 33 with reversible defect in the territory of the left descending coronary artery (homozonal ischemia group) and 9 with a reversible defect in the territory of the left circumflex or right coronary artery (remote ischemia group). There were no significant differences among the three groups with respect to infarct size, presence of dyskinesis and exercise endurance time. Patients with homozonal ischemia had the highest degree of ST-segment elevation (0.22 +/- 0.09 mV) followed by patients without ischemia (0.13 +/- 0.07 mV) and those with remote ischemia (0.09 +/- 0.08 mV, P <.01). In conclusion, Myocardial ischemia adjacent to infarction amplifies exercise-induced ST-segment elevation. PMID- 12122610 TI - QRS aberration during atrial fibrillation at rest and during exercise. Effect of a selective potassium channel blocking agent. AB - This study assesses the occurrence of and identifies clinical characteristics associated with the development of aberrant conduction during infusion of the I(kr)-blocker almokalant. Class III drugs may induce aberrant conduction by prolongation of cardiac repolarization, especially during atrial fibrillation (AF). Ninety-two patients with AF received a 6-hour almokalant infusion, aiming at conversion to sinus rhythm (SR). Fiftyfive of the patients received an identical infusion during SR. During almokalant infusion, the number of patients with intermittent QRS aberration during AF increased, from 21% to 80% at rest, and was further increased to 89% during exercise, with predominantly left, and sequential bilateral, bundle branch aberrancy. Patients with aberrant conduction showed signs of more advanced myocardial disease. Predictors of the development of QRS aberration were female gender, arrhythmia duration, and decreased left ventricular ejection fraction, while use of calcium antagonists decreased the probability. No patient showed aberration during regular SR. Twenty-one patients experienced aberrantly conducted supraventricular premature beats. In conclusion, aberrant conduction is common during infusion of the I(kr)-blocker almokalant during AF, and seems to be more frequent in females and in patients with more advanced myocardial disease. PMID- 12122611 TI - The effect of digoxin on the electrocardiogram of healthy middle-aged and elderly patients at rest and during exercise--a comparison with the ECG reaction induced by myocardial ischemia. AB - The effect of digoxin on electrocardiogram (ECG) at rest and during exercise, and on QRS amplitude variability (variance ECG) was studied in 20 healthy, middle aged men and women. Exercise test and variance ECG were performed before and after pretreatment with digoxin orally. Plots of ST-segment level vergus heart rate (HR) were constructed from the rest and exercise ECG recordings. Thus obtained ST/HR loops were compared with loops from 10 male patients with angiographically verified ischemic heart disease (IHD). Pretreatment with digoxin caused a significant (P <.001) ST depression in precordial leads, which was similar in men and women and returned promptly to the isoelectric level after exercise resulting in a counterclockwise rotation of the ST/HR loop. In IHD patients, the exercise-induced ST-segment depression was significantly more pronounced (P <.01) and the ST-segment recovery slower, resulting in clockwise rotated ST/HR loops. The results of variance ECG were not influenced by digoxin. The digoxin-induced ST-reaction during exercise mimics exercise-induced ischemic ST-reaction in patients with IHD, but can still be discerned by the analysis of ST/HR loops. PMID- 12122612 TI - Biological correlates of QT interval and QT dispersion in 2,265 patients with left ventricular ejection fraction < or =40%. AB - Increased QT interval and QT dispersion have been associated with increased mortality in a variety of cardiovascular disorders including heart failure. However, the biological correlates of these abnormalities are not clear. QTc and QTd were measured in an automated fashion from digitized electrocardiograms in 2,265 patients with an LV ejection fraction 0.05). CONCLUSION: The association between laryngeal cancer and laryngocele requires a detailed evaluation for laryngeal carcinoma in patients in whom an asymptomatic laryngocele had been detected by CT. PMID- 12122626 TI - [Functional outcomes after supracricoid partial laryngectomy]. AB - OBJECTIVES: We evaluated functional outcomes in patients undergoing supracricoid partial laryngectomy (SCPL). PATIENTS AND METHODS: The study included 20 male patients (mean age 61.5 years; range 43 to 76 years) who underwent SCPL for advanced laryngeal carcinoma. Correlations were sought between variables (age, medical history, reconstruction techniques such as cricohyoidopexy or cricohyoidoepiglottopexy, arytenoid resection) and decannulation time, duration for oral feeding, weight change, and complications. The mean follow-up was 20.9 months (range 7 to 39 months). RESULTS: All patients were decannulated in a mean of 19.9 days. Eighty-five percent of patients achieved normal deglutition without aspiration or weight loss within six months postoperatively. The nasogastric feeding tube was removed in a mean of 39.7 days. Voice quality of patients was sufficient for their social communications. CONCLUSION: Our functional results suggest that SCPL is an alternative technique to total laryngectomy in patients in whom other partial laryngectomy techniques are not considered. PMID- 12122627 TI - Fibromatosis of the mandible in a child. AB - Fibromatosis represents a group of fibrous tumors showing clinic and biologic features between benign fibrous lesions and fibrosarcoma. These locally aggressive tumors have high recurrence rates (20% to 70%). A four-year-old boy presented with mandibular fibromatosis occupying the mandible completely and extending to the submandibular gland and soft tissues. Complete hemimandibulectomy and submandibular gland excision were performed followed by reconstruction with a curved Kirschner wire. No signs of recurrence was observed during a follow-up period of 18 months. In addition, no limitations in the functions of the jaw, mastication, and swallowing were noted. PMID- 12122628 TI - [A case of auricular malignant melanoma]. AB - We performed auricular excision, total parathyroidectomy, and extended radical neck dissection in a 36-year-old male patient who developed auricular malignant melanoma. Reconstruction of the surgical defect was made with a split-thickness skin graft. The patient received radiotherapy and chemotherapy after surgery. No evidence of local recurrences or distant metastasis was detected during a follow up period of 14 months. PMID- 12122629 TI - [Four cases of inverted papilloma]. AB - Inverted papilloma is a benign lesion fo the nasal cavity and paranasal sinuses. Its aggressiveness and association with malignancy have been emphasized in the literature. In this paper, four patients (3 females, 1 male) with inverted papilloma are presented. Their ages ranged between 45 and 68 years. Surgery was performed using the degloving method which enables a wide exposure and radical resection with minimal scar on the face. No recurrences were detected in the postoperative period in patients with regular controls. The importance of postoperative histopathologic examination is addressed with a review of the literature. PMID- 12122630 TI - [Efficacy of topical ciprofloxacin and tobramycin in combination with dexamethasone in the treatment of chronic suppurative otitis media]. AB - OBJECTIVES: To evaluate the efficacy of topical ciprofloxacin and tobramycin with and without topical dexamethasone in the treatment of chronic suppurative otitis media without cholesteatoma. PATIENTS AND METHODS: The study included 103 ears of 80 patients (49 males, 31 females; mean age 31 years; range 18 to 60 years) with chronic suppurative otitis media without cholesteatoma. The patients were randomly divided into four groups to receive topical applications of either ciprofloxacin and tobramycin alone, or in combination with dexamethasone. Cultures were obtained from the ears preoperatively and 24 hours after treatment. RESULTS: Aerobic bacteria were isolated in 94.1% of patients before the treatment, the most common being Pseudomonas aeruginosa (38.9%). With dexamethasone, the clinical response for ciprofloxacin and tobramycin increased from 80% to 90% and from 70% to 75%, respectively, but this improvement was not significant (p > 0.03). Addition of dexamethasone to ciprofloxacin decreased the recovery period from 14 days to seven days, whereas no change (7 days) was observed with tobramycin. CONCLUSION: The efficacy of ciprofloxacin and tobramycin were similar in the treatment of chronic suppurative otitis media. Addition of dexamethasone to ciprofloxacin decreased the treatment period. PMID- 12122632 TI - [The incidence of septal deviation in newborns]. AB - OBJECTIVES: To determine the incidence of septum deviation and dislocation in newborns and to investigate relationships between these pathologies and maternal causes and the mode of delivery. PATIENTS AND METHODS: The study included 195 mothers and 200 newborns, of which 10 were twins. Vaginal and cesarean deliveries were performed in 147 (73.5%) and 53 (26.5%) newborns, respectively. All newborns were examined with the use of anterior rhinoscopy. The nasal pyramid, columella, and septum were assessed to detect septal deviation or dislocation. RESULTS: Twenty-three cases (15.6%) of vaginal delivery had septal deviation, while five (3.4%) had septal dislocation. In the newborns delivered by cesarean section, eight (15.1%) had septal deviation, but none had dislocation. Significant correlations were noted between pregnancy, delivery period, the way of delivery and the incidence of septal deviation and columella dislocation (p < 0.05). There was a significant correlation between head circumference and columella dislocation (p < 0.05). CONCLUSION: Since early reconstruction of the potential pathologies may be problem-solving, a careful rhinologic examination should be carried out in the newborns who have prolonged delivery, increased head circumference, and vaginal delivery. PMID- 12122631 TI - [The effect of hyperlipidemia on hearing function]. AB - OBJECTIVES: We evaluated the effects of hyperlipidemia on hearing function. PATIENTS AND METHODS: The study included 274 hyperlipidemic patients (176 females, 98 males; age range 35 to 60 years) who manifested normal tympanic membrane findings, no history of noise exposure, and chronic systemic or ear diseases. Sixty healthy subjects (36 females, 24 males) with normal serum lipid levels were enrolled as controls. Subjects who had elevated serum total cholesterol, triglyceride, and very low density lipoprotein (VLDL) levels, and normal lipid levels were divided into four groups and their hearing functions were analysed with pure-tone audiometry. RESULTS: Evaluation of all groups showed significantly increased hearing levels at three frequencies in female patients, and at six frequencies in male patients (p < 0.05). Significant increases in hearing levels were observed both in females and males at 8000 Hz in three groups, at 6000 Hz in the VLDL group, and in males at 2000 Hz in the total cholesterol and VLDL groups (p < 0.05). When both female and male patients were evaluated together, the largest difference was detected in the VLDL group at five frequencies. CONCLUSION: The findings of our study suggest that hyperlipidemia may have a role on the occurrence of sensorineural hearing loss. PMID- 12122634 TI - [Comparison of palpation, ultrasound and computed tomography in the valuation of lymphatic neck metastasis]. AB - OBJECTIVES: We evaluated the diagnostic value of palpation, ultrasonography (US), and computed tomography (CT) in detecting neck metastasis in head and neck cancers. PATIENTS AND METHODS: The study included 35 patients (34 men, 1 woman; mean age 59 years; range 35 to 72 years) with laryngeal carcinoma. In addition to neck palpation, 17 patients and 27 patients had neck examinations by US and CT, respectively. Histopathologic results of the neck specimens were compared with those obtained from palpation, US, and CT. RESULTS: The accuracy of CT, US, and palpation in the evaluation of lymph nodes of the neck were 85%, 65%, and 80%, respectively. Ultrasonography was found to have the highest sensitivity (100%), but the least specificity (33%). The highest false positive and false negative results were obtained by US (42%) and palpation (10%), respectively. CONCLUSION: Computed tomography proved superior to palpation and US in the evaluation of neck metastasis in patients with head and neck tumors. PMID- 12122633 TI - [Patient selection for near-total laryngectomy and oncologic results]. AB - OBJECTIVES: We reviewed preoperative, perioperative, and postoperative findings and the survival data to determine which patients may be appropriate for near total laryngectomy. PATIENTS AND METHODS: We reviewed hospital records of 20 patients (all males; mean age 56.6 years; range 35 to 73 years) who underwent near-total laryngectomy. Indications for patient selection for near-total laryngectomy and survival data were evaluated in comparison with literature reports. RESULTS: The site of the tumor was the sinus pyriformis in two, and the larynx in 18 patients. Thirteen patients had T3, seven patients had T2 tumors. The lesions were localized in the sinus pyriformis in two patients with T2 tumors. The locoregional control rate at the end of two years was 75%; two- and three-year survival rates were 81.2% and 64.2%, respectively. CONCLUSION: Following a detailed and meticulous investigation in the preoperative period, near-total laryngectomy seems to be appropriate in selected patients with advanced laryngeal and hypopharyngeal tumors in which partial laryngectomy procedures are not considered. It may both provide cure and preserve phonation. It may also be considered for functional purposes in patients whose pulmonary functions are insufficient for partial laryngectomy, in those in whom food aspiration is inevitable after partial laryngectomy, and in those suffering from lifelong food aspiration due to neurologic causes, and for oncologic reasons in patients who develop local recurrences after partial laryngectomy. PMID- 12122635 TI - Laryngotracheal reconstruction of the congenital glotto-subglottic stenosis with autogenous thyroid cartilage interposition: a case report. AB - Surgical correction of grade III glotto-subglottic stenosis in a two-month-old girl was illustrated in a staged manner. Firstly, a silicone keel was placed via anterior thyrotomy following a tracheotomy. Secondly, laryngotracheal reconstruction was performed by interposing an autogenous thyroid cartilage anteriorly between the edges of the longitudinally divided cricoid cartilage and the upper tracheal rings. A stent was maintained for two months. The glottis and subglottis appeared patent and healed following removal of the stent. A meaningful voice and rather comfortable respiration were observed during a 13 month follow-up. The use of thyroid cartilage autograft offers many advantages in laryngotracheal reconstruction with considerably less technical difficulty. PMID- 12122636 TI - [An unusual tracheobronchial aspiration in a patient with total laryngectomy]. AB - Laryngectomized patients use a variety of tools for tracheostomy cleaning, some of which may vary greatly depending on their sociocultural status. The use of inappropriate tools may lead to complications. A seventy-eight-year-old male patient who had a history of total laryngectomy 12 years ago presented with difficulty in breathing and sputum production. Tracheoscopy showed two nails and chest x-rays showed one nail that had been aspirated during stromal care. The three ordinary nails were removed under general anesthesia. This case illustrates the need for detailed and consistent education of laryngectomy patients on proper stomal cleaning and on the associated complications that may arise from the use of inappropriate tools. PMID- 12122637 TI - [Nasolabial cyst: two cases]. AB - Nasolabial cysts are developmental swellings originating from the epithelial remnants of the nasolacrimal ductus. In this report we present two female patients (age 44 and 54 years) with nasolabial cysts. In both cases the leading complaints arose from cosmetic appearance. Surgical excision by sublabial approach was performed. Histopathological diagnoses were in agreement with the clinical diagnoses. The patients had an uneventful postoperative period and no recurrences were seen during a follow-up period of 12 months. PMID- 12122639 TI - The Liversidge Lecture 2001-02. Chemistry amongst the stars: reaction kinetics at a new frontier. AB - Over the past decade, experiments in the Universities of Rennes and Birmingham have provided rate constants for over 40 reactions of molecular and atomic radicals with neutral molecules at temperatures down to 13 K using the CRESU (cinetique de reaction en ecoulement supersonique uniforme) technique. The demonstration that reactions between electrically neutral species can be extremely rapid at these very low temperatures has excited interest both from theoreticians and from those seeking to understand the chemistry that gives rise to the 120 or so molecules that have been identified as being present in dense interstellar clouds. This laboratory work, and its astrochemical and theoretical contexts, are reviewed here. In addition, I deal briefly with the present limitations of the experiments and how they might be overcome in future work. PMID- 12122638 TI - [Genetic sensorineural hearing loss in childhood]. AB - OBJECTIVES: This study aimed to determine the severity, age of presentation, and genetic etiology in hearing handicapped children. PATIENTS AND METHODS: Three groups of children were studied: children who received treatment for hearing loss at the Otolaryngology Department of Ege University Hospital between 1993 and 1999 (1645 children) and throughout 2000 (212 children), and 443 students of Izmir Tulay Aktas Deaf School during 1998 and 1999. All the children underwent otolaryngologic examination and a detailed history taking, audiometric examinations were undertaken in the second and third groups, as well. In children suspected of having a genetic etiology, the pedigrees were drawn and the mode of inheritance was determined. RESULTS: In the first group, awareness of the families to hearing loss and the presentation were at ages two and four, respectively. The most common accompanying anomaly was Down syndrome. Consanguinity was found in 27%, 8%, and 33%, respectively. Moderate to severe hearing loss was detected in 44% of the second group, and in 28% of the third group. Of the pedigrees obtained from 123 families, 86 were nonsyndromic and 27 were syndromic. CONCLUSION: Genetic inheritance plays a substantial role in sensorineural hearing loss in children, with consanguinity being the major culprit. Detection of hearing losses at younger ages will improve the success of hearing rehabilitation programs. PMID- 12122640 TI - Recent developments in the redox-switched binding of organic compounds. AB - This review summarizes and puts into context recent advances in the field of redox-switched binding processes, with particular reference to the interaction between redox-active compounds and organic compounds, both charged and neutral. Simple homogeneous, two-component systems are discussed at first as a precursor to sections on more advanced systems (e.g. multi-component and multi-pole) and binding processes involving components attached to a surface. PMID- 12122641 TI - Towards a fundamental understanding of natural gas hydrates. AB - Gas clathrate hydrates were first identified in 1810 by Sir Humphrey Davy. However, it is believed that other scientists, including Priestley, may have observed their existence before this date. They are solid crystalline inclusion compounds consisting of polyhedral water cavities which enclathrate small gas molecules. Natural gas hydrates are important industrially because the occurrence of these solids in subsea gas pipelines presents high economic loss and ecological risks, as well as potential safety hazards to exploration and transmission personnel. On the other hand, they also have technological importance in separation processes, fuel transportation and storage. They are also a potential fuel resource because natural deposits of predominantly methane hydrate are found in permafrost and continental margins. To progress with understanding and tackling some of the technological challenges relating to natural gas hydrate formation, inhibition and decomposition one needs to develop a fundamental understanding of the molecular mechanisms involved in these processes. This fundamental understanding is also important to the broader field of inclusion chemistry. The present article focuses on the application of a range of physico-chemical techniques and approaches for gaining a fundamental understanding of natural gas hydrate formation, decomposition and inhibition. This article is complementary to other reviews in this field, which have focused more on the applied, engineering and technological aspects of clathrate hydrates. PMID- 12122642 TI - Optical transitions of symmetrical mixed-valence systems in the Class II-III transition regime. AB - In the Robin and Day classification, mixed-valence systems are characterized as Class I, II or III depending on the strength of the electronic interaction between the oxidized and reduced sites, ranging from essentially zero (Class I), to moderate (Class II), to very strong electronic coupling (Class III). The properties of Class I systems are essentially those of the separate sites. Class II systems possess new optical and electronic properties in addition to those of the separate sites. However, the interaction between the sites is sufficiently weak that Class II systems are valence trapped or charge localized and can the be described by a double-well potential. In Class III systems the interaction of the donor and acceptor sites is so great that two separate minima are no longer discernible and the energy surface features a single minimum. The electron is delocalized and the system has its own unique properties. The Robin and Day classification has enjoyed considerable success and most of the redox systems studied to date are readily assigned to Class II. However the situation becomes much more complicated when the system shows borderline Class II/III behavior. Such "almost delocalized" mixed-valence systems are difficult to characterize. In this article spectral band shapes and intensities are calculated utilizing increasingly complex models including two to four states. Free-energy surfaces are constructed for harmonic diabetic surfaces and characterized as a function of increasing electronic coupling to simulate the Class II to III transition. The properties of the charge-transfer absorption bands predicted for borderline mixed valence systems are compared with experimental data. The treatment is restricted to symmetrical (delta G0 = 0) systems. PMID- 12122643 TI - Entropy effects during sorption of alkanes in zeolites. AB - Recent developments in Configurational-Bias Monte Carlo (CBMC) techniques allow the accurate calculation of the sorption isotherms for alkanes, and their mixtures, in various zeolites. The CBMC simulations give new insights into subtle entropy effects affecting mixture adsorption. Three types of entropy effects can be distinguished. (1) Size entropy effects favour the component with the smaller number of C atoms because the smaller molecule finds it easier to fill in the 'gaps' within the zeolite matrix at high molecular loadings. (2) Configurational entropy effects come into play for mixtures of alkanes that differ in the degree of branching. For a mixture of linear and branched alkanes with the same number of C atoms, configurational entropy effects favour the linear isomer because such molecules 'pack' more efficiently within, say, the intersecting channel topology of MFI zeolite. (3) Length entropy effects comes into force for sorption of linear and branched alkanes within the cylindrical channels of say AFI and MOR zeolites; here the double branched alkane has the shortest length and can be packed more efficiently within the channels. We demonstrate that CBMC simulations allow the efficient screening of zeolite structures for a given separation duty and aid the development of novel separation processes exploiting entropy effects. PMID- 12122644 TI - New developments in the Peterson olefination reaction. AB - Noteworthy developments of the Peterson olefination reaction are reviewed. Evidence for both concerted and stepwise mechanisms for the Peterson olefination reaction is presented. The strong affinity of the oxygen anion for the silyl moiety is emphasised when the Peterson olefination reaction takes preference over both the Julia and Wittig reactions in the presence of S- and P-stabilised silyl carbanions. Cerium-mediated Peterson methylenation reactions are discussed. PMID- 12122645 TI - From solid-phase synthesis of pi-conjugated oligomers to combinatorial library construction and screening. AB - A feature article describing concepts for the solid-phase synthesis of pi conjugated oligomers as material-related structures and the translation of the synthetic routes into combinatorial protocols. PMID- 12122646 TI - Synthesis and crystal structure of a novel pentaconta-nuclear silver anionic cluster complex [HNEt3]4[Ag50S7(SC6H4But-4)40].2CS(2).6C3H6O. AB - A silver 4-tert-butylthiophenolate complex generated in situ reacted with CS2 in acetone to give an anionic cluster [Ag50S7(SC6H4But-4)40]4-, the core of which consists of fifty silver atoms and forty-seven sulfur atoms, shaped like a discus with a diameter of ca. 2.0 nm. PMID- 12122647 TI - Preparation of B-free Ti-MWW through reversible structural conversion. AB - B-free titanosilicate with the MWW topology, Ti-MWW, has been successfully prepared from its highly siliceous analogue through structural interconversion and simultaneous titanium incorporation in the presence of piperidine or hexamethyleneimine. PMID- 12122648 TI - Carbon-epoxy electrodes: unambiguous identification of authentic triple-phase (insulator/solution/electrode) processes. AB - A novel electrode consisting of crystals of an electroactive organic solid immobilised into a carbon-epoxy electrode has been subjected to insonation, which facilitates the clear characterisation of triple-phase processes occurring at the insulator/solution/electrode interface. PMID- 12122649 TI - Photosensitization of nanocrystalline TiO2 by self-assembled layers of CdS quantum dots. AB - CdS quantum dots can be self-assembled on high surface area nanocrystalline TiO2 electrodes; spectroscopic and photoelectrochemical studies indicate that the size, and hence the absorption edge, of the CdS particles can be controlled; efficient photosensitization of the TiO2 electrode by the Q-particles has been achieved. PMID- 12122650 TI - A novel [60]fullerene receptor with a Pd(II)-switched bisporphyrin cleft. AB - Porphyrin dimer 1, which does not have an inside cavity and cannot interact with [60]fullerene (C60), becomes an excellent C60-acceptor with a large cavity in the presence of a Pd(II) complex. PMID- 12122651 TI - The regioselective preparation of 1,3-diketones within a micro reactor. AB - We demonstrate a simple method for the regioselective preparation of 1,3 diketones within a micro reactor from silyl enol ethers where the products are free from both competing O-acylation and diacylation products. PMID- 12122652 TI - Approaches to a photocleavable protecting group for alcohols. AB - A new protecting group for the alcohol functionality was devised and shown to be removed photochemically under ultraviolet light in the presence of a radical scavenger in high yields. PMID- 12122653 TI - Discovery of a new family of chromium ethylene polymerisation catalysts using high throughput screening methodology. AB - Following a discovery that salicylaldimines bearing bulky ortho-phenoxy substituents and small imine substituents give very active chromium catalysts for ethylene polymerisation, High Throughput Screening (HTS) methodology has been employed to facilitate a further discovery of exceptionally active catalysts based on tridentate salicylaldimine ligands with bulky triptycenyl groups. PMID- 12122654 TI - Ruthenium catalyzed oxidation of tertiary nitrogen compounds with molecular oxygen: an easy access to N-oxides under mild conditions. AB - A variety of tertiary nitrogen compounds have been efficiently oxidized to their corresponding N-oxides in excellent yields with molecular oxygen as a sole oxidant and ruthenium trichloride as catalyst. PMID- 12122656 TI - Investigation of the morphology-catalytic reactivity relationship for Pt nanoparticles supported on alumina by using the reduction of NO with CH4 as a model reaction. AB - The morphological evolution of large Pt nanoparticles supported on alumina in reaction conditions has a significant impact on the catalytic behaviour for the NO/CH4 reaction. PMID- 12122655 TI - Koilands from thiophiles: mercury(II) clusters from thiacalixarenes. AB - Tetra- and hexa-nuclear mercury(II) complexes have been obtained from tetrathiacalix[4]arene and tetramercaptotetrathiacalix[4]arene, respectively, and structurally characterised in the solid state; the complexes provide new digonal and trigonal receptors of the koiland type. PMID- 12122657 TI - New approaches to high-activity transition-metal catalysts for carbon-carbon bond forming reactions. Rhenium-containing phosphorus donor ligands for palladium catalyzed Suzuki cross-couplings. AB - The rhenium complexes (eta 5-C5H5)Re(NO)(PPh3)((CH2)nPR2:) (n/R = 0/Ph, 0/t-Bu, 0/Me, 1/Ph, 1/t-Bu), which contain electron-rich and sterically congested phosphido moieties, give active catalysts for the title reaction; typical conditions (toluene, 60-100 degrees C): aryl bromide (1.0 equiv.), PhB(OH)2 (1.5 equiv.), K3PO4 (2.0 equiv.), Pd(OAc)2 (1 mol%), and a Re(CH2)nPR2: species or a 1:2 [Re(CH2)nPR2H]+X-/t-BuOK mixture (4 mol% rhenium). PMID- 12122658 TI - Synthesis of Au nanoparticle-conductive polyaniline composite using H2O2 as oxidising as well as reducing agent. AB - We report a new method of synthesis of an Au nanoparticle-conductive polyaniline composite by using H2O2 both for reduction of HAuCl4 and polymerization of aniline in the same aqueous medium: the electrical conductivity of the composite has been measured to be two orders of magnitude higher than the polymer itself. PMID- 12122659 TI - Layered metal organosulfides: hydrothermal synthesis, structure and magnetic behaviour of the spin-canted magnet Co(1,2-(O2C)(S)C6H4). AB - Reaction of cobalt(II) salts with the thiosalicylate dianion under hydrothermal conditions yields green lamellar Co((O2C)(S)C6H4) which displays canted antiferromagnetism. PMID- 12122660 TI - 1,3-Dibromo-2,4,6-trinitrobenzene (DBTNB). Crystal engineering and perfect polar alignment of two-dimensional hyperpolarizable chromophores. AB - 1,3-Dibromo-2,4,6-trinitrobenzene (DBTNB), a two-dimensional charge transfer hyperpolarisable chromophore, crystallizes in the non-centrosymmetric space group C2 in perfect polar order leading to an intense powder SHG signal at 1.06 microns. PMID- 12122661 TI - Engineering of ferrimagnetic Cu12-cluster arrays through supramolecular interactions. AB - Ferrimagnetic oxo-bridged Cu12 aggregates can be organised into a variety of arrays by utilising structure-directing effects of counter ions, solvent molecules and linker units. PMID- 12122662 TI - An evanescent field driven mono-molecular layer photoswitch: coordination and release of metallated macrocycles. AB - Photoswitching of 'on' and 'off' coordination sites in a self assembled monolayer containing the 4-(arylazo)pyridine chromophore has been achieved using the evanescent field and illustrated through coordination and photon induced release of zinc tetraphenylporphyrin. PMID- 12122663 TI - Chiral copper(II) bisoxazoline covalently anchored to silica and mesoporous MCM 41 as a heterogeneous catalyst for the enantioselective Friedel-Crafts hydroxyalkylation. AB - Two solid catalysts in which a chiral copper(II) bisoxazoline has been covalently anchored on silica and MCM-41 have been prepared; the solids are enantioselective catalysts (up to 92% ee) for the Friedel-Crafts hydroxyalkylation of 1,3 dimethoxybenzene with 3,3,3-trifluoropyruvate. PMID- 12122664 TI - Reversible near-infrared fluorescence switch by novel photochromic unsymmetrical phthalocyanine hybrids based on bisthienylethene. AB - A novel family of photochromic hybrids, 2,3-bis(2,5-dimethyl-3-thienyl) unsymmetrical-phthalocyanines (BTE-uPcs), which show a photoregulating fluorescence switch in the near-infrared spectral region were synthesized by a concise route. PMID- 12122665 TI - Impact of Pd-mordenite pretreatment on the heterogeneity of Heck catalysis. AB - In Heck reactions with tributylamine as the base and in toluene, Pd(NH3)4(2+) mordenite (0.4 wt% Pd) and Pd0-mordenite (0.4 and 4 wt% Pd) are not only active and selective, but also truly heterogeneous catalysts, while oxidized PdII species in an all-oxygen environment, i.e. ionic Pd2+ or PdO, are prone to leaching. PMID- 12122666 TI - The versatile conversion of acyclic amides to alpha-alkylated amines. AB - A general and efficient method for the versatile functionalization of acyclic amide via N,O-acetal TMS ether, an excellent precursor for the N-acyliminium ion, has been developed. PMID- 12122667 TI - Stereoselective cycloadditions of chiral acyl-nitroso compounds; selective reactions of ring-cleaved cycloadducts leading to a new approach to polyoxamic acid. AB - Diesters obtained from diacids produced by oxidative ring cleavage of cycloadducts derived from acyl-nitroso compounds and cyclic 1,3-dienes undergo highly regioselective hydrolysis on reaction with lithium hydroperoxide, which allows for easy differentiation of the carboxyl groups leading to a new approach to polyoxamic acid. PMID- 12122668 TI - Catalytic direct 1,4-conjugate addition of aldehydes to vinylketones on secondary amines immobilised in FSM-16 silica. AB - Direct 1,4-conjugate addition of naked aldehydes to vinylketones is catalysed effectively by N-methyl-3-aminopropylated FSM-16 mesoporous silica, which can be regarded as a novel heterogeneous catalysis for a practical C-C bond formation reaction. PMID- 12122669 TI - Ring closing metathesis in protic media by means of a neutral and polar ruthenium benzylidene complex. AB - The ring closing olefine metathesis in protic solvents using a new ruthenium benzylidene complex is described. PMID- 12122670 TI - Using a liquid emulsion membrane system for the encapsulation of organic and inorganic substrates within inorganic microcapsules. AB - We report the development of a novel technique for the encapsulation of molecular and condensed organic and inorganic substrates within hollow calcium carbonate microspheres; the process utilises precipitation at the oil-water interface of a pseudovesicular water-in-oil-in-water emulsion liquid membrane (ELM) system in order to create an inorganic shell around the pre-dispersed media. PMID- 12122671 TI - 12-Tungstophosphoric acid/zirconia--a highly active stable solid acid--comparison with a tungstated zirconia catalyst. AB - A highly active and stable zirconia supported 12-tungstophosphoric acid catalyst is found to be 2-3 times more active in benzylation and acylation reactions than a tungstated zirconia catalyst. PMID- 12122672 TI - Highly reactive heterogeneous Heck and hydrogenation catalysts constructed through 'bottom-up' nanoparticle self-assembly. AB - Polymer-mediated self-assembly of functionalized Pd and SiO2 nanoparticles provides highly active hydrogenation and Heck coupling catalysts. PMID- 12122673 TI - [Cu(tn)]3[Cr(CN)6](2).3H2O: a unique two-dimensional Cu-Cr cyano-bridged ferromagnet (tn = 1,3-diaminopropane). AB - Reaction of the two-coordinate 'assembling complex-ligand' [Cu(tn)]2+ with the building block [Cr(CN)6]3- leads to a unique two-dimensional Cu-Cr cyano-bridged ferromagnet with unusual mu 3- and mu 4-bridging [Cu(tn)]2+ units. PMID- 12122674 TI - Novel hetero-bimetallic metalla-macrocycles based on the bis-1-pyridyl ferrocene [Fe(eta 5-C5H(4)-1-C5H4N)2] ligand. Design, synthesis and structural characterization of the complexes [Fe(eta 5-C5H(4)-1 C5H4N)2](AgI)2(2+)/(CuII)2(4+)/(ZnII)2(4+). AB - The bidentate sandwich ligand [Fe(eta 5-C5H(4)-1-C5H4N)2] has been prepared, structurally characterized and employed in the preparation of the novel supramolecular heterobimetallic metalla-macrocycles [Fe(eta 5-C5H(4)-1 C5H4N)2]Ag2(NO3)(2).1.5H2O, [Fe(eta 5-C5H(4)-1-C5H4N)2]Cu2(CH3COO)(4).3H2O and [Fe(eta 5-C5H(4)-1-C5H4N)2]Zn2Cl4. PMID- 12122675 TI - The direct synthesis of thioesters using an intermolecular radical reaction of aldehydes with dipentafluorophenyl disulfide in water. AB - The combination of the water-soluble radical initiator, 2,2'-azobis[2-(2 imidazolin-2-yl)propane] dihydrochloride (VA-044), and surfactant, cetyltrimethylammonium bromide (CTAB), was found to be the most suitable condition for the effective direct synthesis of useful active thioesters (pentafluorophenyl thioesters) in water. PMID- 12122676 TI - On the electron delocalization in the radical cations formed by oxidation of MM quadruple bonds linked by oxalate and perfluoroterephthalate bridges. AB - Electron paramagnetic resonance, electronic absorption, and resonance Raman spectroscopy reveal that in the oxalate-bridged compounds, [[(tBuCO2)3M2]2(mu O2CCO2)]+[PF6]-, the unpaired electron is delocalized over four metal centers (M = Mo or W) as a result of M2 delta to bridge pi conjugation, but in the related cationic perfluoroterephthalate-bridged species, the tungsten complex is delocalized and the molybdenum analogue valence trapped. PMID- 12122677 TI - In vitro inhibition of gene transcription by novel photo-activated polyazaaromatic ruthenium(II) complexes. AB - Under visible irradiation, [Ru(TAP)2(phen)]2+(Cl-)2, [Ru(TAP)2(POQ-Nmet)]2+(Cl-)2 and [Ru(bpy)2(phen)]2+(Cl-)2 were able to dramatically reduce the in vitro transcription of a plasmid DNA template by a bacteriophage RNA polymerase. This photoactivity is related to two different mechanisms of reactivity towards DNA exhibited by these complexes under illumination. PMID- 12122678 TI - First in vitro norlignan formation with Asparagus officinalis enzyme preparation. AB - We report for the first time that an enzyme preparation from fungal-elicited Asparagus officinalis cultured cells catalyses the formation of a norlignan, (Z) hinokiresinol, from two non-identical phenylpropanoid monomers, 4-coumaryl alcohol and 4-coumaroyl CoA, and from a dimer, 4-coumaryl 4-coumarate, without any additional cofactors. PMID- 12122679 TI - Competing Diels-Alder reactions of activated nitroethylene derivatives and [3,3] sigmatropic rearrangements of the cycloadducts. AB - Diels-Alder reaction of nitroethylene derivatives with cyclohexa-1,3-diene afforded three pericyclic products some of which could be converted to others via a new [3,3]-sigmatropic rearrangement or via a Claisen rearrangement. PMID- 12122680 TI - Muonium addition to DMPO and PBN sorbed in silica-gel. AB - Nitroxyl radicals were formed by adding the light hydrogen isotope, muonium to the spin traps DMPO (5,5-dimethyl-1-pyrroline-N-oxide) and PBN (N-tert-butyl alpha-phenylnitrone) sorbed as 30 wt% ethanol solutions in silica-gel; evidence is presented for a specific hydrogen-bonded interaction between the DMPO adduct and the silica surface; longitudinal-field muon spin relaxation measurements (LF MuSRx) were performed which identified two distinct motional regimes in both samples. PMID- 12122681 TI - Fabrication of two-dimensional layered perovskite [NH3(CH2)12NH3]PbX4 thin films using a self-assembly method. AB - Ultra-thin films of the two-dimensional layered perovskites [NH3(CH2)12NH3]PbX4 (X = Br, I) with the quantum confinement effect have been fabricated by a convenient self-assembly method. PMID- 12122682 TI - Novel ferrocene-containing helical triangular macrocycle achieved via an exchange reaction. AB - A novel ferrocene-containing helical triangular macrocyclic compound has been constructed through an exchange reaction of hydrazone groups which templated and catalyzed by Co(II) ions. PMID- 12122683 TI - Facile fabrication of hollow polystyrene nanocapsules by microemulsion polymerization. AB - The fabrication of polymer nanocapsules with tailored dimensions has been accomplished by microemulsion polymerization using different surfactants. PMID- 12122684 TI - Pyrene covalently anchored on a large external surface area zeolite as a selective heterogeneous sensor for iodide. AB - Pyrene as a fluorophore has been covalently anchored on a delaminated ITQ-2 zeolite having a large external surface area (730 m2 g-1) and the resulting solid found to have a selective response as a heterogeneous sensor for I- in the presence of other halides. PMID- 12122685 TI - Reversible skeletal substitution reactions involving group 13 heterophosphazenes. AB - The cyclic boratophosphazene, N(PCl2NMe)2BCl2 1, reacts with two equivalents of AlMe3 to give the aluminatophosphazene heterocycle, N(PCl2NMe)2AlClMe 4. The unprecedented reverse skeletal substitution (Al for B) was accomplished by treating 4 with Ag[BF4] to form the fluorinated boratophosphazene N(PCl2NMe)2BF2 5. PMID- 12122686 TI - Photocurrent generation system incorporated with antenna function. AB - Porphyrin accumulation onto a self-assembled monolayer (SAM) of imidazole substituted porphyrins by a supramolecular method to form a chain structure leads to significant increase of light absorption in the visible light region and therefore photocurrents. PMID- 12122687 TI - Octanuclear cobalt and nickel cages featuring formate ligands. AB - Two new octanuclear cages are reported which feature formate as a bridging carboxylate which has been formed in situ from decomposition of triphenylacetate used as a ligand. PMID- 12122689 TI - ESI-FTICR investigation of triethylammonium ion-driven resorcin[4]arene dimer formation and structure. AB - In the course of mass spectrometric measurements a self-assembled hydrogen bonded resorcinarene dimer was observed, the formation of which was driven by the binding of triethylammonium ion as a guest and as an ionic label. PMID- 12122688 TI - Molecular batteries: synthesis and characterization of a dendritic 19-electron FeI complex that reduces C60 to its mono-anion. AB - Both redox forms of the dendrimer [dendr-64-NHCOCpFeII(eta 6-C6Me6)]64+/0, 6/7, are synthesized and characterized, and the 19-electron form reduces C60 to [dendr 64-NHCOCpFeII(eta 6-C6Me6)]64+(C60-.)64, 8. PMID- 12122690 TI - Carbon nanohorns grown from ruthenium nanoparticles. AB - A nanoscale ruthenium/gold bimetallic cluster of clusters has been used as a molecular precursor to produce pure ruthenium nanoparticles (seeds) as catalysts for the growth of carbon nanohorns (CNHs). PMID- 12122691 TI - New stereoselective reaction of methylglyoxal with 2-aminopyridine and adenine derivatives: formation of imino acid-nucleic base derivatives in water under mild conditions. AB - A remarkable stereoselective reaction of methylglyoxal with 2-aminopyridine, the nucleic base adenine and adenine nucleosides leads in good yield to heterocycles of a new family in water under mild conditions and should be of interest in the understanding of the biological effects of methylglyoxal which is toxic, mutagenic and involved in diabetic complications. PMID- 12122692 TI - Synthesis of (+)-polyoxamic acid and D-sorbitol from simple achiral allylic halides employing (S,S)-hydrobenzoin as a chiral source. AB - Coupling of cis-1-bromo-2-pentene and (S,S)-hydrobenzoin stannylene acetal followed by regio- and stereoselective transformations of the resulting allylic ether gave (+)-polyoxamic acid and a similar procedure was applied to the synthesis of D-sorbitol from trans-1-iodo-2-hexene. PMID- 12122693 TI - 2-(1,3-Dithiolan-2-ylidene)-5-(1,3-dithian-2-ylidene)-1,3,4,6- tetrathiapentalene(DHDA-TTP), a hybrid of BDH-TTP and BDA-TTP, and its metallic cation-radical salts. AB - The synthesis and electrochemical properties of the DHDA-TTP donor, a hybrid of 2,5-bis(1,3-dithiolan-2-ylidene)-1,3,4,6-tetrathiapentalene (BDH-TTP) and 2,5 bis(1,3-dithian-2-ylidene)-1,3,4,6-tetrathiapentalene (BDA-TTP), has been investigated, and its ability to form metallic cation-radical salts is elucidated. PMID- 12122694 TI - Controlling the variation of axial water exchange rates in macrocyclic lanthanide(III) complexes. AB - The rate of axial water exchange in well-defined series of lanthanide complexes depends on the extent of second sphere hydration which is determined by complex hydrophobicity and the nature of the lanthanide ion and its counter-ion. PMID- 12122695 TI - Laterally-extended porphyrin systems incorporating a switchable unit. AB - p-Quinone units incorporated into the central portion of rigid pi-systems linking either two porphyrin macrocycles or a porphyrin and a phenanthroline group have the potential to function as a chemically and electrochemically controllable switch, thus acting as a means of modulating electronic communication between the two end groups. PMID- 12122696 TI - Synthesis of novel BINOL-derived chiral bisphosphorus ligands and their application in catalytic asymmetric hydrogenation. AB - Some novel ortho-substituted BINOL-derived bisphosphorous ligands (o-BINAPO and o NAPHOS) were synthesis from readily available (S)-BINOL; these ligands showed excellent enantioselectivities (up to 99% ee) in Rh(I)-catalyzed asymmetric hydrogenation of functionalized olefins. PMID- 12122697 TI - Fabrication of vertically and unidirectionally oriented polypeptide assemblies on self-assembled monolayers by stepwise polymerization. AB - A polypeptide assembly prepared by stepwise polymerization on a self-assembled monolayer consisting of amino-alkanethiol and dialkyl disulfide oriented vertically and unidirectionally to the surface. PMID- 12122698 TI - BF3.OEt2-mediated cycloaddition of O-tert-butyldimethylsilyloximes having olefin moieties: intramolecular cycloaddition of N-borano-nitrones. AB - Treatment of O-tert-butyldimethylsilyloximes having olefin moieties in the molecules with BF3.OEt2 results in efficient generation of N-borano-nitrones, which undergo intramolecular cycloaddition at room temperature to afford N nonsubstituted cycloadducts after work-up. PMID- 12122699 TI - Unexpected change of the sense of the enantioselective hydrogenation of ethyl pyruvate catalyzed by a Pt-alumina-cinchona alkaloid system. AB - In the enantioselective hydrogenation of ethyl pyruvate (EtPy) over beta isocinchonine (beta-ICN) modified Pt-alumina catalysts, the major enantiomer was (R)-ethyl lactate ((R)-EtLt (ee 50%)) in toluene, while in AcOH (S)-EtLt (ee 60%) was formed; the (R) configuration is opposite to what is expected from the absolute configuration of the cinchonine backbone. PMID- 12122700 TI - Encapsulation of palladium in polyurea microcapsules. AB - An interfacial polymerisation approach is adopted to encapsulate palladium(II) acetate and palladium nanoparticles in polyurea microcapsules for use in catalysis. PMID- 12122701 TI - Polyurea-encapsulated palladium(II) acetate: a robust and recyclable catalyst for use in conventional and supercritical media. AB - Palladium(II) acetate microencapsulated in polyurea (MC-[Pd]) is an economical and versatile heterogeneous catalyst for a range of phosphine-free cross-coupling reactions in both conventional solvents and supercritical carbon dioxide (scCO2); the catalyst can be recovered by a simple filtration and recycled up to four times. PMID- 12122702 TI - Microfluidic routes to the controlled production of nanoparticles. AB - A microfluidic procedure for the controlled production of cadmium sulfide nanoparticles is described. PMID- 12122703 TI - 2-D soft ferromagnet based on [Wv(CN)8]3- and CuII with a Tc of 34 K. AB - Self-assembly of [Cu(tetren)]2+ (tetren = tetraethylenepentamine) and [W(CN)8]3- in acidic aqueous solution yields the double-layered square grid cyanide-bridged polymer of ((tetrenH5)0.8CuII4[Wv(CN)8](4).7.2H2O)n with Cu(II) centres of square pyramidal geometry coordinatively saturated solely by CN bridges supplied by five [W(CN)8]3- ions; it exhibits soft ferromagnetic behaviour with an ordering temperature Tc of 34 K. PMID- 12122704 TI - Artificial regulation of transcription applying carbohydrate-lectin interaction. AB - A complex of plasmid DNA-lactose conjugate with lectin is proposed as an artificial system to control transcription activity: the in vitro transcription of DNA in the conjugate with T7 RNA polymerase was repressed in the presence of RCA120, and then the transcription ability was recovered by adding lactose or a lactose-carrying polymer to the repression system. PMID- 12122706 TI - Dehydrogenation of aromatic amines to imines via ruthenium-catalyzed hydrogen transfer. AB - An efficient ruthenium-catalyzed transfer dehydrogenation of amines to imines was achieved under mild conditions using 2,6-dimethoxy benzoquinone (2) or cat. 2/MnO2 as oxidant. PMID- 12122705 TI - Conjugation of linoleic acid over a hydrogen pre-activated heterogeneous catalyst. AB - Development of a new heterogeneous catalytic pathway for isomerization of linoleic acid to conjugated linoleic acids at mild reaction conditions over Ni/H MCM-41 in a soluted system. PMID- 12122707 TI - A novel and efficient chiral palladium-phosphinooxazolidine catalyst for the enantioselective Diels-Alder reaction. AB - An efficient ruthenium-catalyzed transfer dehydrogenation of amines to imines was acheived under mild conditions using 2,6-dimethoxy benzoquinone (2) or cat. 2/MnO2 as oxidant. PMID- 12122708 TI - Structure and reactivity studies of the first tungsten cyanoalkylidene complex. AB - Alkylidene complex W(CHCMe2Ph)(NAr)[OCMe(CF3)2]2 (Ar = 2, 6 diisopropylphenyl) (4) reacts with one equivalent of acrylonitrile in methylene chloride to afford the tetrameric, cyanoalkylidene complex [W(CHCN)(NAr)[OCMe(CF3)2]2]4 (5) which reacts with excess acetonitrile to give the tetrameric complex [W(N(H)C(CH3)C(CN)C(CH3)N)(NAr)[OCMe(CF3)2]2]4 (6). PMID- 12122709 TI - Site-selective and stepwise complexation of two M(cod)+ (M = Rh, Ir) fragments with calix[4]arene. AB - The reaction of [(cod)M(mu-OMe)]2 (M = Rh, Ir; cod = cycloocta-1,5-diene) with calix[4]arenes (LH4) in the molar ratio of 0.5-0.6:1 gave the rhodium and iridium pi-arene complexes [(cod)M(eta 6-LH3)], while that in the molar ratio of 1.1 1.5:1 (M = Rh) led to the selective formation of the dinuclear complexes [((cod)Rh)2(eta 6:eta 2-LH2)] in which one of the Rh(cod)+ fragments is coordinated by an eta 6-aryl group and the other by two phenolic oxygen atoms; the stepwise synthesis of the Rh-Ir heterobimetallic analogue of the latter complex was further achieved. PMID- 12122710 TI - Hydrolysis and amine-capping in a glycol solvent as a route to soluble maghemite gamma-Fe2O3 nanoparticles. AB - The preparation of capped metal oxide nanoparticles through the hydrolysis of metal salts is made arduous by the difficulty of dissolving long organic chain capping agents in water; by performing the reaction in propylene glycol under reflux, instead of water, we are able to hydrolyse FeCl3 in the presence of n octylamine to obtain (repeatedly) soluble, monodisperse approximately 5 nm gamma Fe2O3 particles that display a tendency to aggregate into superlattices. PMID- 12122711 TI - The design, synthesis and characterisation of channel-forming peptides. AB - Synthetic peptides designed using criteria derived from statistical analyses of beta-barrel membrane proteins form channels in synthetic membranes at very low peptide:lipid ratios. PMID- 12122712 TI - Phase-transfer catalyst separation by CO2 enhanced aqueous extraction. AB - CO2 is used to enhance the environmentally benign and efficient recovery of phase transfer catalysts with aqueous extraction; this method can alter the distribution of phase transfer catalysts so dramatically that even in dilute organic solutions they can be separated selectively from an organic reaction mixture with only a small fraction of the water required in a traditional aqueous extraction. PMID- 12122713 TI - [Sigmoid diverticulitis: when and to whom should an elective colectomy be offered?]. PMID- 12122714 TI - [Can liver anatomy be changed?]. PMID- 12122715 TI - [Errors in the topographic diagnosis of liver diseases]. AB - To identify the portal pedicles in liver pathology is difficult: anatomical variations are ignored and only the modal disposition is retained, the obliquity of the liver in situ is ignored: strongly inclined to the right, posteriorly and inferiorly (the anterior sector is above and to the right of the posterior sector, their pedicles in an antero-posterior radiogram are superposed); and the sizes of segments IV and VI are quite variable (embryologic result). This study was made with a collection of 111 vasculo-biliary acrylic casts. The main portal fissure containing the middle hepatic vein follows the axis of the cystic fossa. Actually the position of this axis varies from 18 degrees to the right of the vein (gall-bladder under segment V) and 14 degrees to the left (gall-bladder under segment IV); the fissure reaches the inferior vena cava only at the limit of the upper surface of the liver, the vena cava is separated from the right and left livers by the dorsal sector. The anterior half of the right portal fissure is quite variable, it can reach the anterior liver from the main portal fissure up to the anterior portion of the right margin of the liver (segment VI variation); in 41% of the livers (n = 100), the right hepatic vein is in the right portal fissure; occlusion of the anterior or the posterior right arteries indicates the fissure. The left portal fissure is often confused with the left hepatic fissure (limit between academic left and right lobes). Segments breadths are measured in the upper surface of the liver. The largest segments are VIII, V, III and II; their transversal breadth is also the largest (simple to double). In difficult cases, a tri-dimensional reconstruction of the pedicles should be made from an helicoidal tomodensitometry. PMID- 12122716 TI - [Reconstruction after total circular pharyngolaryngectomy: comparison between gastric interposition and free jejunal flap]. AB - AIM OF THE STUDY: To elucidate hospital mortality, morbidity and actuarial survival rates of patients with carcinoma of the hypopharynx and cervical oesophagus and to identify the technique of choice for reconstruction after pharyngolaryngectomy. PATIENTS AND METHODS: We reviewed the records of 209 patients who underwent total pharyngolaryngectomy between May 1982 and January 2000. The majority of patients had advanced cancer: hypopharyngeal in 131 cases and cervical oesophageal in 78 cases. Follow-up was complete for all patients. Chi 2 and log rank tests were used, with a limit of significance of 5%. RESULTS: The postoperative mortality and morbidity rates were 4.8% and 38.3%, respectively. Alimentary continuity was achieved using the stomach (127 patients), colon (5 patients), or free jejunal autograft (77 patients). The 1 year and 5-year survival rates were 62% and 24%, respectively. There was no significant difference with regard to the survival between gastric transposition and free jejunal autograft, but there were fewer complications in the gastric pull-up group with regard to the respiratory complications (33% vs 47.0%, p < 0.05), local recurrences (15.8% vs 33.8%, p = 0.004) and survival without dysphagia (76% vs 89%, p < 10(-5)). CONCLUSION: Surgical ablation is a viable option for advanced hypopharyngeal and cervical oesophageal neoplasms, and stomach interposition is the preferred method of reconstruction. PMID- 12122718 TI - [How to prevent surgical morbidity after a total thyroidectomy for a multinodular euthyroid goiter?]. AB - AIM OF THE STUDY: Total thyroidectomy has been advocated for the treatment of multinodular nontoxic and benign goiter. The aim of this study, based on our experience, was to define the surgical factors which permit to decrease morbidity related to total thyroidectomy for multinodular euthyroid benign goiter. METHODS AND MATERIALS: In a retrospective study performed between January 1996 and September 2000, all records of total thyroidectomy for initial treatment of multinodular euthyroid benign goiter were reviewed. This study allowed to specify recurrent and parathyroid morbidity after surgery. RESULTS: There were 51 women and 13 men with a mean age of 47 years. Recurrent laryngeal nerve injury occurred in 2 patients. It resolved in 1 patient but was permanent in another (1.6%). Transient hypocalcemia was found in 8 patients (12.5%). One patient had permanent hypocalcemia (1.6%). CONCLUSION: The results of our serie are comparable to previous reports. Systematic identification of the recurrent laryngeal nerve, and preservation of the parathyroid blood supply permit to decrease the surgical morbidity. PMID- 12122717 TI - [Total pancreatectomy for mucinous pancreatic tumor]. AB - AIM OF THE STUDY: To report our experience of total pancreatectomy (TP) in ten patients with mucinous pancreatic tumors (MPT), to discuss pre and peroperative investigations in the management of MPT, and operative, functional and carcinologic results after TP. PATIENTS AND METHODS: This retrospective study from January 1985 to January 2001 included ten patients, 5 men and 5 women (mean aged: 64 years). Six patients underwent one step TP for intraductal papillary mucinous tumor of the pancreas (IPMT) in 5 cases, and multifocal mucinous cystadenoma in one case. Four patients underwent a second step TP for tumor recurrence (2 IPMT, and 2 cystadenocarcinomas) which occurred 12 to 121 months post operatively (mean: 49 months). RESULTS: Post TP diabetes was controlled by insulinotherapy (3 injections a day), except in one patient who needed insulin administration through a pump. One patient, with cystadenocarcinoma, died from cancer recurrence 18 months after TP and 140 months after the initial pancreaticoduodenectomy. One patient died from heart disease 34 months postoperatively. The 8 other patients were alive with a mean follow-up of 33 months (range 11-61 months). CONCLUSION: Curative surgery for mucinous tumors of the pancreas may require TP, which is indicated preoperatively according to imaging, or intraoperatively following surgical findings and frozen section of the pancreatic margin. Totalization of a previous partial pancreatectomy is mandatory in case of tumoral persistence or recurrence in the pancreatic remnant. Postoperative diabetes can be managed successfully by a specialized team. PMID- 12122719 TI - [Surgery for intra-bronchial foreign bodies]. AB - OBJECTIVE: The aim of this study was to evaluate the results of surgical treatment of intra-bronchic foreign bodies after unsuccessful endoscopic extraction. MATERIALS AND METHODS: Fifteen patients underwent surgical treatment of intra-bronchic foreign bodies in our department during a period of 10 years. There was 9 males and 6 females, the mean age was 20.27 years (range between 5 and 39 years), there were 8 right localizations and 7 left. The recurrent pulmonary infections were most frequent complaint of patients. RESULTS: Twelve pulmonary resections (including one pneumonectomy) were necessary because of the importance of pulmonary parenchyma destruction. There was one post-operative complication (one empyema) and the peri-operative mortality was nul. CONCLUSION: Surgical treatment of intra-bronchic foreign bodies must be used only after unsuccessful endoscopic extraction. The preventive measures are the best treatment. PMID- 12122720 TI - [Endoscopic, percutaneous and laparoscopic treatment for acute biliary pancreatitis]. AB - AIM OF THE STUDY: The aim of this study is to evaluate the results of acute gallstone pancreatitis treatment and to discuss indications in relation with the different forms of the disease. MATERIAL AND METHOD: From january 1992 to june 2001, 137 patients have been treated for an acute gallstone pancreatitis. Diagnostic criteria were given by the history, clinical examination, biochemical and radiological findings. After exclusion of patients with a systemic disease, a group of 129 patients have been enrolled in a treatment regimen with an endoscopic retrograde cholangiopancreatography (ERCP) and eventual sphincterotomy, a percutaneous US-guided cholecystostomy (PC) when necessary and an elective laparoscopic cholecystectomy. RESULTS: ERCP has been successfully performed in 121/129 patients. A PC has been performed in 5/8 patients of the failed endoscopic procedure and in 14 with acute cholecystitis. Retrograde and percutaneous cholangiographies showed main bile duct stones in 89 patients, a dilatation of the main bile duct without stones in 26 patients and a negative finding in 6 patients. An endoscopic sphincterotomy has been performed in 117 patients. A laparoscopic cholecystectomy has been performed in 118 patients. Mortality and morbidity rates were 1.6 and 10.3%, respectively. CONCLUSION: ERCP and sphincterotomy seem to be indicated in all patients observed during the first 72 hours. Endoscopic treatment and percutaneous procedure make it possible to reduce at a very low rate the cases with an unfavourable course of the disease. A definitive treatment may then be performed by the way of a laparoscopic cholecystectomy. PMID- 12122722 TI - [Recklinghausen's disease and gastrointestinal stromal tumors]. AB - Gastro-intestinal stromal tumors associated with Recklinghausen's disease should be considered in the current concept of the stromal tumors with reference to recent advances in immuno-chemistry. In this setting, there is an high potential of maluignancy. For the treatment of these lesions, surgery is the main tool. Frequency of malignant digestive diseases associated with Recklinghausen disease should be kept in mind. PMID- 12122721 TI - [Natural history of the pancreatic stump after duodenopancreatectomy of the pancreatic head]. AB - Major complications following pancreaticoduodenectomy are thought to be chiefly associated with exocrine secretion of the pancreatic remnant which is not well known. This work aims to assess the exocrine secretion of the pancreatic remnant within the early post-operative period. PATIENTS AND METHODS: Seventy-five patients undergoing pancreaticoduodenectomy for presumed tumour were included in a prospective multicentre study. A tube was inserted in the pancreatic duct at the time of construction of the pancreatic anastomosis. Peripancreatic drainage was routinely used. Pancreatic juice and peripancreatic drainage fluid were collected and measured and pancreatic enzyme monitored. For 7 days patients received total parenteral nutrition and continuous infusion of randomly Somatostatin 14 (S-14) at a dose of 6 mg/24 h (days 1-6) and 3 mg/24 h (day 7) or matching placebo. Pancreatic fistula was defined as a daily drainage of more than 100 cc of amylase-rich fluid after day 3, persisting after day 12 or associated with symptoms or needing specific treatment. RESULTS: Daily output of pancreatic juice was low during the first postoperative day and then increased gradually until day 5. A high enzyme concentration was observed in pancreatic juice on the first post-operative day. S-14 infusion resulted in a significant decrease of both pancreatic fistula rate and enzyme concentration in peripancreatic fluid. CONCLUSIONS: During the first postoperative days, the outflow of the exocrine secretion of the pancreatic remnant is low but contains a high enzyme concentration with significant leaks within the peripancreatic area. S-14 infusion results in a decrease of pancreatic juice leaks from the pancreatic remnant. PMID- 12122723 TI - [Colonic granulocytic sarcoma: a case report]. AB - Granulocytic sarcoma is a rare tumor composed of immature cells of the granulocytic series which usually occurs as a secondary manifestation of acute leukaemia. We report the case of a 60 years old woman without particular previous pathologies who was hospitalised for chronic diarrhea developed in a context of health impairment state. The blood cell count revealed severe leucopenia and thrombopenia; an emergency right colectomy was accomplished. The histologic examination showed granulocytic sarcoma of the ascending colon. The death occurred rapidly as a consequence of a toxic shock. This observation seems to be the sixth case report of the granulocytic large bowel sarcoma in the literature which likely complicated a pre-existant and unknown myeloid leukaemia. PMID- 12122724 TI - [Preparation for Patey incision in mastectomy: technique demonstration]. AB - The authors report a technique of skin resection for mastectomy in order to allow secure, tension free closure using a suture upon the nipple. PMID- 12122726 TI - Epidemiology of inflammatory bowel disease. AB - The incidence of IBD has either continued to increase or has stabilized at a high rate in most developed countries, whereas the incidence continues to rise in regions where IBD had been less common. The prevalence has continued to increase as a result of a combination of previously rising incidence and improved survival. Regardless of the exact prevalence, the burden of disease in North America and Europe is significant. Studying the patterns of geographic variation and age and gender distribution may yield important clues to the cause of IBD. PMID- 12122725 TI - [Acute intraperitoneal rupture of liver hydatid cysts]. PMID- 12122727 TI - Endoscopy in inflammatory bowel disease. AB - With the heterogeneous clinical presentation of IBD, endoscopy plays an integral role in the initial diagnosis of ulcerative colitis and Crohn's disease. Although radiographic tests often are supplemental in the work-up of IBD, colonoscopy with biopsy is the test of choice if IBD is suspected and is more sensitive than radiographic tests. Endoscopic features can be helpful in differentiating ulcerative colitis and Crohn's disease. Currently the only mode for detecting dysplasia and stratifying the risk of developing colorectal cancer is through complete colonoscopy with multiple biopsy specimens. Complications of IBD can be managed effectively with endoscopic therapy. The role of endoscopic ultrasound and future developments in endoscopic therapy need to be defined by future studies. PMID- 12122728 TI - Pathologic features of early inflammatory bowel disease. AB - Often the pathologic changes of IBD are subtle and may not be present in a proportion of biopsy specimens. In cases of early disease, the changes may be missed, and additional specimens should be taken after a period of time. Modifying factors, such as prebiopsy treatment and coexisting disease, should be considered. A forum to review cases and allow for communication between gastroenterologists and pathologists is especially useful for clinicopathologic correlation and assignment of a working diagnosis to each case. Careful attention to the pathologic features of early UC and CD would be most useful when evaluating new therapies for IBD. PMID- 12122729 TI - Medical therapy for ulcerative colitis. AB - Although newer therapeutic agents are being developed for the treatment of inflammatory bowel disease, aminosalicylates and corticosteroids remain the mainstay of treatment for UC (Tables 2-5). Patients who do not respond to these agents or become steroid dependent require immunomodulatory therapy or curative surgery. Cyclosporine represents the greatest treatment advance for UC in 10 years. The role of nicotine, heparin, antibiotics, probiotics, and SCFA in the treatment of UC is less clear, but these agents may offer an alternative therapeutic approach for patients intolerant or nonresponsive to standard therapy. PMID- 12122730 TI - Medical treatment of Crohn's disease. AB - Crohn's disease is not medically (and is rarely surgically) curable. Patients do, however, live a normal life span. The goal of therapy is to optimize the quality of life, minimize disease activity and disease-related complications, and avoid therapeutic toxicity. PMID- 12122731 TI - Inflammatory bowel disease. Medical therapy of specific clinical presentations. AB - Ulcerative colitis and Crohn's disease are chronic relapsing inflammatory disorders of the gastrointestinal tracts. The inflammatory process is restricted to the mucosa and submucosa of the colon in ulcerative colitis and is transmural and may occur anywhere in the gastrointestinal tract in Crohn's disease. Clinical presentation of these inflammatory disorders depends on the segments of digestive tract affected and on the extent and aggressiveness of the disease process. The treatment of specific clinical presentations of these disorders is discussed in this article. PMID- 12122732 TI - Nutritional factors in inflammatory bowel disease. AB - Dietary antigens may act as important stimuli of the mucosal immune system and have led to the study of nutritional therapy for IBD. Patients with active CD respond to bowel rest, along with total enteral nutrition or TPN. Bowel rest and TPN are as effective as corticosteroids at inducing remission for patients with active CD, although benefits are short-lived. Enteral nutrition is consistently less effective than conventional corticosteroids for treatment of active CD. Use of palatable, liquid polymeric diets in active CD is controversial, but these diets are of equal efficacy when compared with elemental diets. UC has not been treated effectively with either elemental diets or TPN. Fish oil contains n-3 PUFA, which inhibits production of proinflammatory cytokines and has some benefit in the treatment of CD. Topical applications of short-chain fatty acids have benefited diversion colitis and distal UC, whereas probiotics hold promise in the treatment of pouchitis. PMID- 12122733 TI - Inflammatory bowel disease and environmental influences. AB - Some environmental factors have been established firmly as influences on the development of IBD, such as smoking and appendectomy. Other behaviors, such as oral contraceptive use and sugar intake, have been suggested as risk factors, but he data conflict sharply. The importance of other behaviors, such as breast feeding, is even murkier with sharply divergent data. Some issues studied may not be factors in themselves but rather markers for other unidentified influences. The conflicting evidence in many of these studies may be clarified as specific genes are identified and the interplay between these environmental factors and genetic subtypes is investigated. PMID- 12122734 TI - Surgery for ulcerative colitis. AB - The type of surgery performed for UC varies from patient to patient and must take into account the nutritional status and health of the patient, the presence of dysplasia or cancer, the desire of the patient to maintain continence, the preoperative anorectal function, the degree of confidence in the diagnosis of UC, and the technical constraint because of certain body habituses. A total proctocolectomy is the surgical procedure of choice for UC. A restorative proctocolectomy is the preferred surgical approach that not only cures the patient of the disease and prevents the development of colorectal cancer, but also maintains continence with an improved quality of life. PMID- 12122735 TI - Cancer in inflammatory bowel disease. An evidence-based analysis and guide for physicians and patients. AB - The risk of cancer in IBD is real and is a cause of anxiety and concern among patients and practitioners. Current modalities for detecting dysplasia in IBD are crude and insensitive and subject to observer and sampling bias. This evidence based review confirms a significant increased risk for colorectal cancer among patients with pancolonic UC and, to a lesser extent, in patients with left-sided disease. Risk increases with longer duration of disease; early age at diagnosis; coexisting PSC; and, perhaps, a family history of colorectal cancer. Physicians must pay greater attention to the manner in which they implement surveillance colonoscopies, including paying heed to the location and number of biopsy specimens required to maximize the benefit. With respect to CD, the evidence suggests that patients with extensive colonic involvement of long duration carry a similar risk of colorectal cancer to patients with UC and should be considered candidates for surveillance colonoscopy. PMID- 12122736 TI - The surgical management of Crohn's disease. AB - The surgical management of patients with CD can be complex and fraught with complications. Thorough preoperative evaluation by a multidisciplinary management team should delineate the indications for surgery and allow formulation of an operative strategy. Although surgery is not a cure for CD, approximately three quarters of patients require an operation. Until better postoperative maintenance strategies are developed, many CD patients eventually require a reoperation. PMID- 12122737 TI - Pregnancy and nursing in inflammatory bowel disease. AB - The peak age of onset for IBD coincides with the peak age for conception and pregnancy. Women with inactive IBD who become pregnant do not have increased complications compared with age-matched controls. Most medications for IBD are safe in pregnancy. The greatest danger to a normal conception and pregnancy is active disease, not the medicine used to treat it. This article outlines fertility in IBD, the effect of IBD on pregnancy, the effect of pregnancy on IBD, and the medical therapy available. PMID- 12122738 TI - Inflammatory bowel disease in the pediatric and adolescent patient. AB - Advances in genetic testing have confirmed the presence of susceptibility loci on chromosomes 12 and 16 for UC and CD. These loci show a strong association with particular disease phenotypes that may explain the clinical heterogeneity of IBD. Whether multiple genotypes will be found to explain these phenotypes remains to be determined. Pharmacogenetic differences in 6-mercaptopurine metabolism can be used clinically to predict patient susceptibility to drug-induced toxicity. Novel treatment strategies are being developed at The Johns Hopkins Medical Center Hospital based on these inherent genetic differences. The aim is to improve treatment efficacy and clinical response times and prevent untoward drug-induced toxicity. PMID- 12122739 TI - The atypical colitides. AB - Collagenous and lymphocytic colitis are atypical colitides that should be considered in elderly patients with unexplained diarrhea. The lack of natural history data and apparent underreporting of these disorders have made the determination of true incidence difficult. Better insight into the pathogenesis and heightened awareness of these conditions will lead to earlier diagnosis and more effective treatment. PMID- 12122740 TI - Extraintestinal manifestations of inflammatory bowel disease. AB - Numerous extraintestinal diseases have been associated with IBD. The role of the gastrointestinal tract in host response to the foreign antigens present in the gut makes the enteric immune system highly susceptible to any external perturbation to the system. Dysregulation of the enteric immune response results in pathology in various organs outside of the gut. The site-specific manifestations of this immune response are not understood fully. Better understanding of the pathogenesis of IBD and the complex interactions between the gut immune system and the extraintestinal systems would provide insights into the development of many of these extraintestinal manifestations. Much is unknown about the presence of cardiac, pulmonary, and hematologic diseases in patients with IBD. True association or coincidental presence of the diseases in these organ systems with IBD requires better delineation. An important consideration in all patients with IBD presenting with extraintestinal manifestations should be a careful search for medication-related complications. PMID- 12122741 TI - Hepatobiliary disease in inflammatory bowel disease. AB - Many hepatobiliary diseases are seen in IBD. PSC is the most common, occurring in 7.5% of patients with UC. The cause of PSC is not well understood, but PSC seems to be associated with genetic susceptibility, sharing some immunologic abnormalities with UC. A characteristic cholangiogram in a patient with abnormal liver function tests usually establishes the diagnosis. Liver biopsy is not essential but can help make the diagnosis of small duct PSC in patients with a normal cholangiogram. There are no medications that treat PSC effectively. Endoscopic dilation of dominant strictures reduces the frequency of cholangitis and may improve survival. OLT remains the only proven treatment of advanced PSC. Cholangiocarcinoma is a feared complication of PSC that is difficult to diagnose. Cholelithiasis, PBC, portal vein thrombosis, and hepatic abscess are hepatobiliary disorders that occur less frequently in IBD patients. PMID- 12122743 TI - Microbial factors in inflammatory bowel disease. AB - An unsolved puzzle in IBD research is whether germs, genes, or a combination of the two with excessive immune responses to gut-associated bacteria explains the pathogenesis of UC and CD. Whatever the answer, there is little doubt that microbial factors are involved intimately in IBD pathogenesis. Although a long search has failed to confirm a direct pathogenic role for a specific infectious agent, compelling evidence suggests that commensal enteric bacteria and their products provide a local environmental trigger that initiates and perpetuates IBD, reactivates quiescent disease, results in the frequent septic complications of CD, and contributes to the development of several extraintesinal manifestations. The most compelling evidence for involvement of the enteric flora in the pathogenesis of IBD has been generated from studies of animal models, which collectively support the view that IBD is due to genetically determined dysregulation of the mucosal immune response to luminal antigens derived from the normal bacterial flora. Although removing or dampening the dominant antigenic stimuli with antibiotics or probiotics is conceptually superior to the current array of immunosuppressive and anti-inflammatory agents that nonspecifically block the inflammatory cascade, more definitive, rigorously designed, controlled trials of treatments directed at the microflora are needed. Future research investigating mechanisms of tolerance to luminal bacteria and an understanding of how probiotics can manipulate the intestinal flora beneficially will bring clinicians closer to identifying potential therapeutic targets and unraveling the bacterial connection to IBD pathogenesis. PMID- 12122742 TI - Functional bowel disorders in inflammatory bowel disease. AB - Patients with IBD in remission often have ongoing gastrointestinal symptoms that are related to active inflammation. It is now apparent that functional gastrointestinal disorders may overlap with IBD and increase morbidity and diminish the quality of life of patients. Recognition and treatment of functional symptoms that may be the result of IBD are crucial in the appropriate medical management of these patients. PMID- 12122744 TI - The genetics of inflammatory bowel disease. AB - The complex genetics of IBD is characterized by more than one susceptibility locus, genetic heterogeneity, incomplete penetrance, and probable gene-gene and gene-environment interactions. Functional candidate gene association studies during the past few decades have revealed only modest associations between IBD and genetic variants in the HLA genes and a limited number of other genes that are involved in immune regulation and the inflammatory response. Important advances in IBD genetics research have come about from systematic genome searches for IBD loci. The identification of Crohn's disease-associated NOD2 genetic variants that appear to alter the innate immune response to bacteria is a seminal finding that perhaps is the greatest advance toward understanding the pathogenesis of IBD in decades. The future discovery of other IBD genetic risk factors, facilitated by the completion of the human genome sequencing and annotation, may allow the development of better therapies, possibly including preventive therapies, for patients with Crohn's disease and ulcerative colitis. PMID- 12122745 TI - The immunology of inflammatory bowel disease. AB - The dogma of systemic immunity can be ignored in relation to mucosal immunity. These distinct immune systems evolved based on specific environmental realities. Given the potentially hostile environment encountered by the mucosal immune system, it is a necessary adaptation that the overall tone is one of suppression. Breaches in this tone may lead to chronic inflammation. Understanding how specific components and processes of the mucosal immune system interact with the environment will elucidate the pathogenesis of IBD and lead to novel therapeutic approaches. PMID- 12122746 TI - Radiographic imaging of inflammatory bowel disease. AB - Radiographic imaging studies have an important role in the workup of patients with suspected IBD and in the differentiation of ulcerative colitis and Crohn's disease. Because of its ability to depict fine mucosal detail, the double contrast barium study is a valuable technique for diagnosing ulcerative colitis and Crohn's disease even in patients with early disease. In contrast, cross sectional imaging studies such as CT, MR, and ultrasound are useful for showing the effects of these conditions on the wall of the bowel and also for demonstrating intra-abdominal abscesses and other extraluminal findings in patients with more advanced disease. Thus, barium studies and cross-sectional imaging studies have complementary roles in the evaluation of these patients. PMID- 12122747 TI - Breakthrough in psychiatric care: pharmacological treatment of obsessive compulsive disorders with implications for nursing care. PMID- 12122748 TI - "Culture matters for the patient in pain". AB - It is important for each nurse to be alert for not only individuals who are experiencing pain but also the impact that cultural phenomena may have on the way pain is experienced, expressed, and the assistance that is desired. The Giger Davidhizar Transcultural assessment model provides a model for assisting in the assessment of pain. It is only by consideration for cultural phenomena that culturally appropriate care can be delivered. PMID- 12122749 TI - [Stenting of the right subclavian artery posttraumatic false aneurysm]. AB - Posttraumatic false aneUrysms can be life threatening and often their management might be a challenge, because of poor general condition as well as because of specific technical difficulties. Open surgery requires wide exposure and dissection of posttraumatic tissues. Endovascular techniques using stent-grafts gains popularity because of high efficiency, safety and good results. We present a case of posttraumatic false aneurysm of right subclavian artery treated with a stent-graft. PMID- 12122750 TI - [Diagnostic and therapeutic strategy by patients with abdominal and thoracoabdominal traumas]. AB - The pointed tendency towards increasing incidence and heaviness of the trauma patients in the last decades led to the emphatic increasement of the hospitalized patients with abdominal and/or thoracoabdominal trauma. For the time-period 1990 2000 total 431 pts with abdominal trauma were admitted. 283 of them underwent operative intervention. Male--221 and female--62, aged from 15 to 83 years (mainly pts under 40 years--188--66.43%). With blunt abdominal trauma were 164 pts (115 male and 49 female); with open wound trauma--119 (107 male and 12 female). According to the injured organ the distributiOn of the patients was: liver--61; spleen--47; pancreas--6; stomach--16; duodenum--2; small intestine- 32; colon--27; diaphragm--18; mesenterium--33; omentum--21; kidney--4; "lucky" penetrating wounds--12. Most valuable for the exact diagnosis were: careful examination of the patient, U S, Plain abdominal X-ray, Abdomenocenthesis (68 pts), Laparoscopy (34 pts), Thoracoscopy (16 pts). Almost all of the patients underwent operative procedure during the first 12 hours (276 pts), and even in 45% of them, operation was performed in the first 2-4 h. Intraoperative autohaemotransfusion was performed in 19 pts with parenchymal lesions (liver and spleen). Reoperations were performed in 9 pts (3.53%). Postoperative morbidity- 13.78%, and postoperative mortality--4 pts (1.41%). PMID- 12122752 TI - [Urological complications of colorectal carcinoma in senile patients]. PMID- 12122751 TI - [Biliary peritonitis: retrospective analysis of the disease]. AB - The authors have done a biliary retrospective analysis of 10 deceased patients with a diagnosis of biliary peritonitis (BP). It is reported that the patients who died of BP are 9.5% of all who died of diffuse purulent peritonitis. All deceased patients were advanced and well advanced in years with prolonged complaints--in over 50% of them over 5 years. The complications set in 50% of the cases are recorded in the postoperative period. The perforation of the gallbladder and the biliopancreatitis are in the second place. Intrabiliary fistulas with 4 patients and 1 with a vesicocolon fistula; tumor of the pancreas with 4 patients; empyema of the gallbladder--3; diabetes with 3 of the patients and others are recorded as accompanying troubles which complicate the operation and the outcome of it. The time between the beginning of the complication and the operation with all the deceased patients is over 72 hours. PMID- 12122753 TI - [Composite rhytidectomy a modo Hamra]. AB - Conventional face lift techniques--subcutaneous, SMAS have always incorporated unopposed lateral vector tissue advancement. In case they are not adequately repositioned, the tissues of the lower eyelid and upper cheek continue to age, which create a "lateral sweep" of the lower face as those malar soft tissues descend at a more rapid rate than the repositioned SMAS. Hollow eyes and the lateral sweep could be prevented with a rhytidectomy technique that includes orbicularis repositioning and preservation of the lower eyelid fat with an arcus marginalis release or with the composite rhytidectomy of Hamra. The technique was applied in 19 cases and the results are discussed. PMID- 12122754 TI - [Contusion traumas with damaged entity of the eye globe]. AB - PURPOSE: Examination of the etiology, the clinical history of the injury, the functional and anatomic result after contusion traumas with damaged entity of the eye globe. MATERIAL AND METHODS: Retrospective examination of the clinical documentation of 35 patients /35 eyes/, treated in Emergency Medical Center during the period 1994-1996. RESULTS AND DISCUSSION: The etiology shows that traumas resulting from housework prevail. According to their localization the ruptures are: scleral--19 eyes (54.28%), corneo-scleral--13 eyes (37.14%) and corneal--3 eyes (8.57%). Prolaps of uvea and vitreous was diagnosed with all the eyes. It was massive in the scleral and corneo-scleral ruptures. No eye underwent primary enucleation. The therapeutical behavior consists in primary reconstructive operation, general and local antibiotic, cortico-steroidal and non steroidal anti-flammatory therapy. The functional and anatomic results are bad: absence of visual acuity with 51.42% of the eyes and vision up to 20/200 with 45.71%, subatrophy of the eye globe during the first 6 months after the trauma with 18 eyes (51.42%). CONCLUSION: The contusion traumas with damaged entity of the eye globe are hard traumatic injuries which usually end with bad functional, anatomic and cosmetic results. The consequence is constant invalidation of the patients. PMID- 12122755 TI - [ND YAG laser in senile cataract microsurgery]. AB - PURPOSE: To enlarge indications for Nd YAG laser anterior capsulotomy in extracapsular extraction of senile cataract and to improve the methods for its implementation. MATERIAL AND METHODS: Nd YAG laser anterior capsulotomy we accomplished in 618 eyes of patients with senile cataract. The eyes we divided in groups according to the matureness of the cataracta. The laser capsulotomy was implemented during the ECCE. We created the methods we used: "symmetric ascending on circumference", "horizontal linerar", "circular", "methods for expholiations". RESULTS AND DISCUSSION: In all 618 eyes we implemented successful Nd YAG laser anterior capsulotomy. It had smooth edges and was precise, sharply shared and executed with maximum saving of the adjacent tissue. The Nd YAG laser anterior capsulotomy as part of the ECCE helped to minimize duration of the surgery intervention and to increase the number of the operations, which can be implemented in the surgical time. We enlarged the indications for Nd YAG laser anterior capsulotomy in extracapsular extraction of senile cataract and opposing to other authors /7/ we agreed that as a method it is extremely appropriate in cases like cataracta senilis, matura and hypermatura in which an instrumental surgical intervention is possible to lead to some complications concerning zonula Zinni damaging. CONCLUSION: The use of Nd YAG laser for capsulotomy during the ECCE gives the opportunity to anticipate the pathological response of the eye tissues and to retain clear cornea and normal IOP. The methods we offered- "symmetric ascending on circumference", "horizontal linerar", "circular" and "methods for expholiations"--support the implementation of precise, sharply shared anterior capsulotomy with maximum preserving of the adjacent tissue. The successful capsulotomy provides for optimum conditions for the next steps in the operation, minimization of the surgical time, tranquil postoperative period. The Nd YAG laser anterior capsulotomy has found its place as a part of the modern ophthalmosurgery. PMID- 12122756 TI - [Contusion eye traumas: etiology, clinical characteristics and final results]. AB - PURPOSE: Examination of the aetiology, clinical characteristics and the final results for contusion traumas of the eyeball. MATERIAL AND METHODS: Retrospective examination of the clinical documentation for 247 patients (260 eyes of which 219 male and 41 female), treated in Emergency Eye Clinic during the period 1994-1996. The patients are separated in groups by their age. DISCUSSION AND RESULTS: Contusion traumas are more frequent in children and men aged up to 50 years old. The traumas are caused by different factors but these having housework and criminal character prevail. The eyeball is with not damaged entity in 225 eyes (86.54%) and with damaged entity in 35 eyes (13.46%). Injuries in both eyes are observed in 11 patients. The clinical characteristics show simultaneous injury of the front protection elements and the eyeball. More often all tissues and areas of the eye are damaged. Serious damages in the area of iris-lens diaphragm, vitreous and retina are observed. The final visual acuity in 178 eyes (68.48%) is between 20/40 and 20/20. CONCLUSION: The contusion traumas are more frequent in children and young men. The traumas caused by factors having housework and criminal character prevail. Usually all tissues and areas of the eye are damaged. The final visual acuity of the most of the eyes is between 20/40 and 20/20. PMID- 12122757 TI - [Implantation microsurgery in traumatic cataract]. AB - ECCE with IOL implantation was applied to 41 eyes. In accordance with the trauma the eyes were divided in two groups: A--traumatic cataract after contusion trauma -5 eyes (12.20%) B--traumatic cataract after perforating trauma--36 eyes (87.80%). Three of the eyes from group B are with intra-ocular foreign body. A decision for implantation microsurgery is taken individually. Primary IOL implantation was applied to 26 of the eyes (63.41%) and secondary IOL implantation--to 15 of the eyes (36.59%). Only in 4 eyes anterior chamber IOL were implanted. In the other 37 eyes the implanted IOL were posterior chamber with fixation in the ciliary sulcus area. The final visual acuity was 0.5 and higher in 30 eyes (73.17%) and 0.8-1.0 in 11 eyes (26.83%). The primary or secondary IOL implantation in traumatic cataract is a reliable and effective method that helps to oppose amblyopy, to restore the binocular function and for the visual rehabilitation of the patients. This method gives the opportunity to achieve high values of the visual acuity with insignificant complications. PMID- 12122758 TI - [Main lymphocyte populations and their subpopulations in patients with acute pancreatitis studied in the course of disease]. AB - The aim of the study was to establish the changes in the main lymphocyte populations and their subpopulations in peripheral blood of patients with acute pancreatitis. Seventy-one patients were studied in the course of disease. They were classified on the basis of severity of pancreatitis--mild and severe form. Percentage and absolute numbers of CD19+ (B lymphocytes), CD2+ (total T cells), CD3+ (immunocompetent T cells), CD3+CD4+ (helper T cells), CD3+CD8+ (suppressor/cytotoxic lymphocytes), CD4+CD29+ and CD4+CD45-RA+ (helper/inducer and suppressor/inducer subpopulations of helper lymphocytes). Immunophenotype analysis was performed using a flowcytometer Epics XL-MCL, Coulter, USA and monoclonal antibodies by the same company. In the early stage of disease the fall of total CD2+, immunocompetent CD3+ T lymphocytes and their subpopulations (more expressed in the severe form) is an indicator of determining the severity of pancreatitis. These results along with the elevated CD4+CD29+ subpopulation are a sign of both activation of these links in the local immune reaction and development of secondary infectious complications. The observed fall in CD4+CD45 RA+ suggests increased risk of development of autoimmune process. The recovery of most of the studied parameter after day 3 shows unimpaired host compensatory capabilities for both forms of disease. PMID- 12122759 TI - [An attempt for the creation a united working scheme for the comparative analysis of peritonitis]. AB - Many attempTs have been made to create an universal classification of peritonitis. The aspirations of many authors to work out a scheme that is able to be in one and the same time thorough, simple and appLicable in practice, have been found to be unrealizable. This blank stimulate us to work up a scheme which will facilitate showing the nature of the pathological process, the analysis of the clinical data, the similarities and the differences in the development of the illness and the most important--to direct to the most rational operation. We offer the following United working scheme for comparative analysis of peritonitis. It includes: Classification of mixed type formulated on the base of anatomical and etiological background; Modified classification of V. Feodorov about dissemination of peritonitis; Classification based on pathophysiology of the peritonitis; Classification system for estimation the seriousness of the illness MPI-modified. PMID- 12122760 TI - [A case of ileus-peritonitis like a complication from small intestine metastatic malignant melanoma]. PMID- 12122761 TI - [Contusion eye trauma caused by explosion of vacuumed coffee tin]. AB - PURPOSE: To report a rare case of eye contusion trauma, resulting from explosion of vacuumed coffee tin. MATERIAL AND METHODS: 7 years old patient underwent injury of the right eye while opening a vacuumed coffee tin. The examination consisted in: history, evaluation of the visual acuity and refraction, examination of intraocular pressure, biomicroscopy, ophthalmoscopy. RESULTS AND DISCUSSION: During the examination an edema has been found in the macula area. This edema lately degenerated. When the patient was admitted his visual acuity was 20/30 and did not change when he was released. The visual acuity of the left eye was 20/20. CONCLUSION: Contusion trauma, caused by vacuumed coffee, can be followed by constant changes in the area of the macula and result in decreasing of the visual acuity. PMID- 12122762 TI - Medication errors and ethical nursing practice. PMID- 12122763 TI - Restraining patients and the practice of the profession: congruent or dissonant. PMID- 12122764 TI - Assessment of older patients in acute care settings. PMID- 12122765 TI - Pain in older persons with cognitive impairment. PMID- 12122766 TI - The missing link. AB - The uniqueness of nursing research is derived from the philosophical view of the individual as a biopsychosocial being. Nurse scientists are prepared to illuminate the linkages among the biophysiological, psychological, and social domains, and this study is much enhanced by the increasing availability of valid and reliable biomarkers. Researchers need to develop expertise in the use of biomarkers and secure appropriate funding for their use. Missing links may be missing no longer. PMID- 12122767 TI - Nurses and the Canadian Institutes of Health Research. PMID- 12122768 TI - Gerontological content in Canadian baccalaureate nursing programs: cause for concern? AB - Over the next several decades the number of older Canadians will rise dramatically. This shift will have implications for the demand for health-care professionals, particularly nurses, educated to work with an aging population. The purpose of this study was to assess the status of gerontological content in Canadian undergraduate nursing curricula. Earthy's Survey of Gerontological Curricula in Canadian Baccalaureate Nursing Programs was used to collect data from a convenience sample of Canadian nursing schools and faculties with baccalaureate programs. Data were analyzed using descriptive and frequency statistics. Gerontological content was found to be integrated into the majority of programs, but only 8% of clinical hours had a focus on the nursing care of older adults and only 5.5% of students chose geriatrics for their final clinical practical prior to graduation. Implications include the need to develop faculty expertise, the potential for interdisciplinary gerontology education in the health sciences, and the need to address ageism in the nursing profession. PMID- 12122769 TI - Coping research: the road ahead. PMID- 12122770 TI - Double agency in clinical research. AB - The current focus on evidence-based practice in nursing may result in nurses playing 2 roles concurrently--that is, acting as researcher and caregiver at the same time and with the same people. Given the fiduciary nature of the patient caregiver relationship, this double agency can give rise to problems, both real and perceived. In this paper, the issues associated with assuming dual roles in research with humans will be examined, particularly in relation to recruitment and informed consent, data collection, and participant withdrawal from a study. In addition, strategies to prevent or minimize problems related to double agency are identified, with attention to the guidance provided by professional codes of ethics and the Tri-Council Policy Statement: Ethical Conduct for Research Involving Humans. PMID- 12122771 TI - The family adaptation model: examination of dimensions and relations. AB - The purpose of this paper is to summarize the theoretical underpinnings and present the model analyses used in the development and evaluation of the Family Adaptation Model. Resilience theory, with its components of protective processes and vulnerability processes, underlies the assumptions of the model. Data analyses are presented from 2 samples in which survey methodology, post-test only experimental designs were implemented. There is moderate support for the linear dimensions of the model. When the paths predicted by the theory were tested, insignificant results were produced. Recent expert reviews of adaptation concepts and research approaches were used to explore the meaning of the null findings when testing the paths of the model in contrast to the success of the model when used to develop practice approaches with families. PMID- 12122772 TI - Envisioning the future: nightingale continues to guide. PMID- 12122773 TI - Employed mothers: stress and balance-focused coping. AB - This critical feminist grounded theory study examined how employed mothers coped with the stress of managing multiple responsibilities in family, health, and paid work. Over a 2-year period, 20 mothers employed as support staff in a large, publicly funded institution participated in repeated individual in-person and telephone interviews and in a focus group. Interviews were transcribed and analyzed using constant comparative methods. The women experienced stress from continuous demands in paid and family health work compounded by time constraints, inflexible expectations, conflicting demands, compromised personal resources, and inadequate support. Most of their coping strategies were individual, such as focusing on priorities, but some women used shared family decision-making. Findings support both individual and family stress and coping theory, yet underscore the need to explicate social-ecological influences such as relational power. Strategies that can enhance coping and reduce stress are described for nurses who work with women and families. PMID- 12122774 TI - Learning to live with early dementia. AB - Much of the literature on early dementia is focused on caregiver perspectives, while little is known about the perspective of persons with early-stage dementia such as what it is like to live with this syndrome. This study was conducted to explore the process of learning to live with early-stage dementia. Interviews were conducted with 6 early-stage participants (3 men and 3 women) ranging in age from 61 to 79 years. Theory construction was facilitated using a qualitative approach and grounded theory. A preliminary theoretical framework was developed from the data which outlines a 5-stage process of learning to live with dementia that begins with various antecedents and proceeds through the stages of anticipation, appearance, assimilation, and acceptance. This process evolved as participants' awareness of themselves and their outer world changed. Ultimately, the findings of this study have several implications for clinicians and researchers working with persons in early-stage dementia. PMID- 12122775 TI - Knowledge in nursing: contemplating life experience. PMID- 12122777 TI - Coping, adaptation, and nursing: what is the future? PMID- 12122776 TI - Mothers' perspectives of an in-home nursing respite service: coping and control. AB - What are mothers' experiences of receiving in-home nursing respite care for their children with medically fragile or complex conditions? This paper highlights selected findings from a comprehensive 2-year study designed to evaluate the impact of a Canadian nursing respite program. Data were collected at 3 time points in the homes of 27 families, using both quantitative and qualitative methods. This paper reports on 1 construct arising from unstructured interview and observational data: learning to manage the system. A specific subset of 10 mothers was chosen for constant comparative analysis of their perspectives of receiving in-home nursing respite, revealing a 4-phase process: taking in, losing control, taking charge, and managing effectively within the constraints of inflexible rules. These findings contribute to nursing knowledge about meeting the in-home respite needs of mothers of children with complex medical conditions. Implications for nursing include how to better support maternal coping, decrease uncertainty, and foster more effective relationships with mothers of children with complex conditions. PMID- 12122778 TI - Using biologic markers to identify legitimate chronic pain. PMID- 12122779 TI - Containing the spread of HIV infection among high-risk groups. PMID- 12122780 TI - Evaluation of the effectiveness of an addiction treatment training program for physicians. PMID- 12122781 TI - Latest discoveries on the infection and coinfection with hepatitis D virus. AB - HDV is an incomplete virus that has HBV infection as a prerequisite. Superinfection by HDV leads to acute hepatitis and causes progression to liver cirrhosis in a significant proportion of HBsAg carriers. The traditional methods for the diagnosis of HDV infection, such as detection of serum anti-HD antibodies, are sufficient for the clinical diagnosis of delta infection. However, such techniques lack the sensitivity and specificity required to more accurately characterize the nature of HDV infection and to assess the efficacy of therapies. Recent improvements in molecular techniques, such as HDV RNA hybridization and RT-PCR, have provided increased diagnostic precision and a more thorough understanding of the natural course of HDV infection. These advances have enhanced the clinician's ability to accurately evaluate the stage of HDV infection, response to therapy, and occurrence of reinfection after orthotopic liver transplant. PMID- 12122782 TI - Balascopy: an emerging tool for diagnostic medicine. PMID- 12122783 TI - LabAutomation 2002: Internet access to health data: elder-care, home-care, and self-care programs--Part 4. PMID- 12122784 TI - [Prof. Dr. Hans Meyer (1877-1964). One of the founders of radiation oncology in Germany and Europe]. PMID- 12122785 TI - [Histomorphological structural changes of head and neck blood vessels after pre- or postoperative radiotherapy]. AB - BACKGROUND: Patients with squamous cell carcinomas of the oral cavity are being increasingly treated by multimodal interdisciplinary regimes using a combination of surgery, chemo- and radiotherapy. Inflammatory alterations of the vascular endothelium following preoperative radiotherapy frequently cause healing delays of free flaps in the irradiated graft bed. The aim of the study was to investigate quantitative and qualitative changes of irradiated neck recipient vessels and transplant vessels used for microsurgical anastomoses in free flaps in patients undergoing preoperative radiotherapy or radiochemotherapy. PATIENTS AND METHODS: In 348 patients (October 1995-March 2002) receiving primarly or secondary 356 microvascular hard- and soft tissue reconstruction, a total of 209 vessels were obtained from neck recipient vessels and transplant vessels during anastomosis. Three groups were analysed: group 1 (27 patients) treated with no radiotherapy or chemotherapy; group 2 (29 patients) treated with preoperative irradiation (40-50 Gy) and chemotherapy (800 mg/m2/day 5-FU and 20 mg/m2/day cisplatin) 1.5 months prior to surgery; group 3 (20 patients) treated with radiotherapy (60-70 Gy) (median interval 78.7 months; IQR: 31.3 months) prior to surgery. From each of the 209 vessel specimens, 3 sections were investigated histomorphometrically, qualitatively and quantitatively (ratio media area/total vessel area) by NIH-Image-digitized measurements. To evaluate these changes as a function of age, radiation dose and chemotherapy, a statistical analysis was performed using an analysis of covariance and chi 2 tests (p > 0.05, SPSS V10). RESULTS: In group 3, qualitative changes (intima dehiscence, hyalinosis) were found in recipient arteries significantly more frequently than in groups 1 and 2. For group 3 recipient arteries, histomorphometry revealed a significant decrease in the ratio media area/total vessel area (median 0.51, IQR 0.10) in comparison with groups 1 (p = 0.02) (median 0.61, IQR 0.29) and 2 (p = 0.046) (median 0.58, IQR 0.19). No significant difference was found between the vessels of groups 1 and 2 (p = 0.48). There were no significant differences in transplant arteries and recipient or transplant veins between the groups. Age and chemotherapy did not appear to have a significant influence on vessel changes in this study (p > 0.05). CONCLUSIONS: Following irradiation with 60-70 Gy, significant qualitative and quantitative histological changes to the recipient arteries, but not to the recipient veins, could be observed. In contrast, irradiation at a dose of 40-50 Gy and chemotherapy given at a median interval of 1.5 months prior to operation did not lead to significant histological changes to the recipient vessels. PMID- 12122786 TI - Microscopic residual disease is a risk factor in the primary treatment of breast cancer. AB - OBJECTIVES: In the primary treatment of breast cancer, postoperative radiotherapy is performed in high-risk patients after mastectomy and in patients who received breast conserving surgery. In a retrospective analysis, our mono-institutional results of postoperative irradiation have been evaluated. PATIENTS AND METHODS: Between 1992 and 1996, 500 patients have been irradiated after surgery for primary breast cancer. Of these, 489 patients had no initial metastases. 89 patients with loco-regional disease had a mastectomy, 400 patients were irradiated after breast conserving surgery. Radiotherapy at the chest wall was performed with 50 Gy and 2 Gy fractions. After microscopically incomplete resection, an electron boost of 10 Gy was given. The ipsilateral lymph nodes were irradiated with 50 Gy when there was extensive lymph node involvement or invasion of tumor in the axillary fat tissue. RESULTS: The 5-year local control rate after mastectomy was 97.4% and 91.2% after breast conserving surgery. The only statistically significant risk factor for local failure was microscopically incomplete resection. The corresponding 5-year local control rates for microscopically incomplete and complete resections were 76.4% and 92.7% (p = 0.01). The risk of local relapse was increased with both marginal invasive and marginal DCIS-tissue. 86.6% of local relapses were in the same quadrant. CONCLUSIONS: High-risk patients after mastectomy and patients with breast conserving surgery achieve a high local control rate with postoperative irradiation. After microscopically incomplete resection, there is an increased risk for local relapse. PMID- 12122787 TI - [The effectiveness of radiation treatment in comparison with extracorporeal shockwave therapy (ESWT) in supraspinatus tendon syndrome]. AB - BACKGROUND AND AIM: Supraspinatus tendinitis is usually treated by antiinflammatoric drugs, local injections, physiotherapy or low-dose irradiation. A novel approach is the use of Extracorporeal Shock Wave Therapy (ESWT) if conservative therapies have failed. So far there has been no controlled study comparing the effectiveness of ESWT with an established conservative method of therapy such as X-ray stimulation radiotherapy. PATIENTS AND METHOD: 30 patients with chronic supraspinatus tendinitis were admitted into the prospective randomized study. After randomization the patients were treated either with X-ray stimulation radiotherapy with 6 x 0.5 Gy on the ICRU reference point (1 fraction/day) with cobalt 60 gamma rays or three times with 2000 pulses (energy flux density ED+ 0.1 mJ/mm2) in 1 week intervals using a Storz Minilith SL1. Primary endpoint was the age-corrected constant score 3 months after intervention. RESULTS: Acute side effects caused by the irradiation were not observed, as expected. One patient described pain and one patient showed a moderate skin irritation after ESWT. In the radiotherapy group average the age corrected constant score improved from 47.6 through 79.5 points to 87.4 points. In the ESWT group it rose from 50.1 points before ESWT to 91.4 points after 12 weeks and 97.8 after 52 weeks. CONCLUSION: No statistically significant differences were proven between ESWT and radiotherapy. ESWT appears to be equivalent but not superior to radiotherapy in treating chronic supraspinatus tendinitis syndrome. A comprehensive randomized study is, however, necessary to ensure the equivalence of ESWT. PMID- 12122788 TI - Intraindividual comparison of two different skin care conceptions in patients undergoing radiotherapy of the head-and-neck region. Creme or powder? AB - OBJECTIVES: This study evaluated for the first time in intraindividual comparison the conception of care with creme or powder. Acute skin reactions on the left and right side of the neck during symmetrically performed radiotherapy and subjective impairment were assessed. PATIENTS AND METHODS: Twelve patients receiving radiotherapy of the head-and-neck region up to 50, 60, 66 and 72 Gy, respectively, were asked to treat one side of the neck with creme, the contralateral side with powder after random assignment. Objective and subjective assessment and photodocumentation were performed at therapy onset and weekly during therapy. The photodocumentation provides an impressive course of acute skin reactions under these care concepts. RESULTS: Altogether we saw no relevant difference in favor of care with creme or powder according to objective as well as subjective assessment criteria (Figure 2 and 3). In this intraindividual comparison the results are independent from interindividual different sensitivity to radiotherapy, total dose or fractionation. Therefore the small patient number is a minor limitation for evidence. Our results are in accordance to trials comparing other care concepts. CONCLUSIONS: A relevant effect concerning onset and degree of acute skin reactions or differences in symptom relief could not be demonstrated. Both conceptions are to be regarded equal in terms of subjective symptom relief and in extent of acute skin reactions. The lack of differences may be explained by the fact that the underlying pathophysiological processes cannot be influenced by topical agents. PMID- 12122789 TI - Radiotherapy in the management of keloids. Clinical experience with electron beam irradiation and comparison with X-ray therapy. AB - BACKGROUND: Aim of this study was to evaluate the advantages of electron beam irradiation compared to kilovoltage X-ray therapy in the treatment of keloids. Furthermore, the risk of developing malignancy following keloid radiotherapy was assessed. PATIENTS AND METHODS: An automatic water phantom was used to evaluate the dose distribution in tissue. Furthermore, a series of measurements was done on the patients using thermoluminescence dosimeters (TLD) to estimate the doses absorbed by the organs at risk. We also report our clinical experience with electron beam radiation of 134 keloids following surgical excision. RESULTS: Electron beam irradiation offers a high control rate (84%) with minimal side effects for keloids. Electron irradiation provides better dose distribution in tissue, and therefore less radiation burden to the organs at risk. After a mean follow-up period of 7.2 years, no severe side effects or malignancies were observed after keloid radiotherapy. CONCLUSIONS: Electron radiation therapy is superior to kilovoltage irradiation for treating keloids due to better dose distribution in tissue. In agreement with the literature, no cases of malignancy were observed after keloid irradiation. PMID- 12122790 TI - [The value of radiotherapy in treatment of meningeal melanocytoma]. AB - BACKGROUND: Meningeal melanocytoma is described as rare benign lesion with a high risk of recurrence. There are no well-substantiated treatment recommendations in the literature. Only case reports have been published by now. PATIENTS AND METHODS: In 1997 a patient was irradiated for a recurrent spinal meningeal melanocytoma and 2 years later for brain metastases indicating malignant transformation. This case gave rise to a literature review for therapeutic options. All sufficiently documented cases published since 1972, when the term meningeal melanocytoma was established, were evaluated. Based on published and on original data recurrence and overall survival rates up to 5 years were calculated for three different therapeutic approaches, namely complete tumor resection, incomplete resection with subsequent radiotherapy, and incomplete resection alone. Statistical evaluation was performed using the chi 2 test and Kaplan-Meier analysis. RESULTS: 53 patients (including our patient) met selection criteria. Complete tumor resection was superior to incomplete resection alone with lower recurrence (4-38% versus 50-92%) and better overall survival rates (86-95% versus 30-58%). After incomplete resection radiotherapy seemed to improve prognosis (recurrence 15-45%; overall survival 91-92%). Between complete resection and incomplete resection plus radiotherapy no significant differences were observed. CONCLUSIONS: For meningeal melanocytoma complete resection must be regarded as the best of the modalities compared. After incomplete resection radiotherapy should be considered, although a specific radiotherapeutic regimen cannot be recommended at present. However, for multiple cranial or spinal lesions total cranial irradiation or craniospinal irradiation is indicated. PMID- 12122791 TI - [Properties of a commercial Hi-pSi detector for dosimetry of stereotactic collimators with very small diameters]. AB - BACKGROUND: Conformal stereotactic radiosurgery and radiotherapy with linear accelerators and hole collimators yield a dose concentration in the target volume by rotation of the gantry. For small target volumes collimators with isocentre diameters of 4-45 mm are used. In this paper dosimetric measurements with a commercial high doped p-type silicon detector are demonstrated and compared to measurements with diamond detector and ionisation chamber. MATERIAL AND METHODS: The properties of the silicon detector SFD from Scanditronix were investigated with the radiation of a Gammatron S and a Varian 2100 CD at 6 MV. The results were compared with those of a calibrated ionisation chamber (0.3 cm3) and a diamond detector. MEASUREMENTS AND RESULTS: At the beginning the reproductibility of the registered dose and dose rate and the temperature dependence of the Si detector were investigated at the Gammatron S. For the comparison the absorbed dose was measured with the ionisation chamber in air. The sensitivity decreases slightly with dose and dose rate. After a period of several days without radiation again higher doses were registered. The temperature dependence causes deviations of 0.25%/K. The signal-to-noise ratio and the spatial resolution were investigated with the linear accelerator. The signal-to-noise ratio is clearly lower compared with that of the diamond detector, whereas the resolution is nearly the same. CONCLUSIONS: The Si-detector is qualified for dosimetry of very small fields because of the insignificant dose and dose rate dependence and in spite of some disadvantages regarding dosimetric properties compared with the diamond detector. The advantage is the availability and the cost. Measurement with ionisation chambers are not useful for collimator diameters below 20 mm. PMID- 12122792 TI - [Comment on the contribution by Hocht S, et al. Treatment of keratitis superficialis chronica of the dog with strontium 90]. PMID- 12122793 TI - Getting the HIPAA consent and notice mix right--for patient and provider. PMID- 12122794 TI - Improved workflow through HIS. Intuitive technology that automates and streamlines hospital workflow contributes to improved treatment outcomes and financial health for the enterprise. PMID- 12122795 TI - The business of surgery. Managing the OR as a profit center requires more than just IT. It requires a profit-making mindset, too. PMID- 12122796 TI - ERP (enterprise resource planning) in the consumer-driven marketplace. Competitive organizations can meet the changing requirements of market-driven healthcare with integrated, Internet-based ERP applications. PMID- 12122797 TI - What works: e-transcription. Talk is cheap. Physician organization implements a Web-based transcription system and saves money, while improving service. PMID- 12122798 TI - What works: clinical information systems. Order entry rules. Healthcare enterprise achieves physician acceptance, reduced medication errors and improved patient outcomes through CIS and CPOE technology. PMID- 12122800 TI - Helping doctors help patients. Interactive pre-procedure education tools increase patient knowledge and enhance patient-provider interaction. PMID- 12122799 TI - What works: physician practice. Using technology as the road map to action. Dallas practice utilizes information management and outsourcing to increase revenues. PMID- 12122801 TI - Healthcare information systems hotlist. PMID- 12122802 TI - Demystifying telehealth. PMID- 12122803 TI - Paresis, historical therapy in the perspective of Caelius Aurelianus, with special reference to the use of hydrotherapy in antiquity. AB - Caelius Aurelianus provides in his work Tardarum sive chronicarum passionum, based on Soranos' famous, but lost, work about acute and chronical illnesses, a remarkably detailed description of the physio-therapy of paresis, which covers the complete therapeutic spectrum of the groundwork of a combined therapy. His view that rehabilitative treatment should be started from the second day of illness sounds almost revolutionary. Also, modern early rehabilitation makes a specific use of combined therapy in a way that is analogous to that described by Caelius Aurelianus. Even today, the view is taken that fast mobilisation of the patient is the top priority of therapy. The three-stage mobilisation therapy involving exercises in rolling-in-bed as well as practice in trying-to-sit-up is quite similar to what is common practice today. PMID- 12122804 TI - Julien Offray de la Mettrie (1709-1751). AB - The year 2001 marks the 250th anniversary of la Mettrie's death. This paper commemorates his stormy life and the contribution he made to the neurosciences. Trained as a physician, la Mattrie soon fell out with both the medical and ecclesiastical authorities and was exiled first to the Netherlands and then to Frederick the Great's circle of intellectuals at Sans Souci (Berlin). The two works which are of greatest interest to the readers of this journal are the 1745 Histoire Naturelle de l'ame (retitled Traite de l'ame in 1751) and the 1747 l'Homme Machine. Both are collected in the 1751 Oeuvres Philosophiques. This paper reviews these two ground-breaking tracts noting that, although they are both materialistic, and hence worthy of the odium theologicum into which they fell, they are not materialistic in the Cartesian sense. La Mettrie is dismissive of the Cartesian concept of inanimate matter and the related notion of the 'beast machine'. Instead, he sees an unbroken continuity between humans and the rest of nature. His vision is in many ways far ahead of its time and prefigures some of the dilemmas and concerns which our evolutionary neuroscience presents today: how is consciousness related to the goings on in the cerebrum? How can we be held responsible for what we do if all is material? His biographer, Raymond Boissier, writes that we can recognise in him an obscure predecessor of Lamarck and a prophet of things to come. PMID- 12122805 TI - Armand Trousseau--some of his contributions to neurology. AB - Trousseau made a remarkably large number of original clinical contributions to medicine and neurology. Best known are Trousseau's syndrome, the combination of venous thrombosis with visceral carcinoma; tache cerebrale, the red streak seen on scratching the skin in acute meningitis; and Trousseau's sign, the cardinal physical sign in tetany. His pioneering work in tracheostomy in diphtheria, haemochromatosis, Parkinson's disease, aphasia and chorea are but a few of his outstanding clinical studies. Based on his famously comprehensive text, Clinique Medicale de l'Hotel Dieu, this paper highlights a few of his discoveries. The name of Armand Trousseau must stand alongside those of Charcot, Oppenheim, Jackson and Gowers in the annals of neurology. PMID- 12122806 TI - The neuroscience of Helmholtz and the theories of Johannes Muller. Part 1: Nerve cell structure, vitalism, and the nerve impulse. AB - Hermann Helmholtz made monumental contributions to the neural sciences in the second half of the nineteenth century. Among his earliest achievements were experiments that challenged vitalism, microscopic studies on the structure of the nerve cell and its processes, and the first reasonable estimates of the speed of nerve transmission based on physiological experiments. In this, the first of a two-part article, we review Helmholtz's early contributions in biographical context and with reference to Johannes Muller's own thoughts. We reveal how Johannes Muller, considered by many to be the greatest physiologist of the first half of the nineteenth century, helped to launch and shape Helmholtz's career. We also show that Helmholtz was only willing to accept some of his mentor's theories, even though he had great admiration for Muller. The point will be made that Helmholtz owed a great debt to Muller, but even from his student days in Berlin he was an independent thinker with his own agenda, and never his strict disciple. PMID- 12122807 TI - Classic articles of 19th-century American neurologists: a critical review. AB - The purpose of this article is to critically review citation classics of 19th century members of the American Neurological Association (ANA), and to elaborate what these works contributed and why they continue to be important. Most classic articles of 19th-century American neurologists were initial or early descriptions of clinical conditions, diseases, or procedures. These include descriptions by Beard of the Jumping Frenchmen of Maine; by Sachs of "amaurotic family idiocy" (Tay-Sachs disease); by Hun of the lateral medullary syndrome; by Mitchell of phantom limbs; and by Dana of familial tremor. Few of these were the initial description, although most were clear and fairly complete by modern standards. Several citation classics were cited mainly as a point of comparison with later events or developments, including those by Corning on spinal anesthesia, Bartholow on electrical stimulation of the brain, Mitchell on the status of American psychiatry, and Starr on childhood brain tumors. The reports of Corning, Bartholow, and Mitchell have been the subjects of continued controversy. The only examples of basic neuroscience among the citation classics are the classic studies by Onuf and Collins involving ablation of portions of the sympathetic chain in cats, and Onuf's description of the nucleus of Onuf in the human spinal cord. Onuf's basic science work was made possible by a unique and short-lived multidisciplinary research environment created at the New York State Pathological Institute for the scientific investigation of insanity and neurologic diseases. PMID- 12122808 TI - The Gordon-Hey reflex. AB - Nowadays, a neurologist, even in many years of practice, rarely sees a patient with Sydenham's chorea. Things were quite different during the period +/- 1850 - +/- 1950, when 'chorea minor' was the subject of hundreds of publications. In those days, the practising neurologist stood a good chance of coming across patients with such a degree of muscular hypotonia (a characteristic feature of severe Sydenham's) that he was inclined to infer the presence of paralysis. Those instances used to be denoted as chorea mollis or chorea paralytica. According to textbooks then considered to be authoritative, any doubt regarding the diagnosis of Sydenham's chorea was largely removed if one succeeded in eliciting Gordon Hey's reflex. This eponym turns out to be as fascinating as the question of its still largely unclarified pathophysiology. In spite of that, mention of the reflex has wholly disappeared from the textbooks of neurology since circa forty years. PMID- 12122809 TI - Neurognostics question 16: the size principle. PMID- 12122810 TI - Lasthenie de Ferjol's syndrome: a tribute paid by Jean Bernard to Jules Amedee Barbey D'Aurevilly. PMID- 12122811 TI - Inclusiveness and coherency in the history of the neurosciences. AB - The International Society for the History of the Neurosciences (ISHN) defines "neurosciences" broadly, and we want to encourage the widest possible range of scholarly approaches to our subject. However, this deliberate inclusiveness could potentially cause problems with internal coherency in our organization and in the scholarship that we are trying to create. In other words, we need to avoid the pitfalls of the internalist-externalist tension without losing the benefits of both perspectives. In fact, I think that there is a large and interesting "gray zone," where the boundary between these supposedly separate approaches is both artificial and porous. This should be one of the most rewarding intellectual domains for the study of neuroscience history, because our subject is always culturally loaded by the mind/body problem and by the assumptions that it entails. Of course, neuroscience history is also influenced by all of the other cultural and scientific aspects of the milieus in which it is conducted. Understanding neuroscience history in all of its multiple historical contexts will require the participation of a wide range of scholarly viewpoints. To keep ourselves coherent in the process, we will have to educate each other. PMID- 12122812 TI - Relative value units and productivity: Part 2 of 4. AB - The initial article in this series (Volume 17, No. 5: 225-228) discussed relative value unit (RVU) basics and touched on some of the more practical applications of RVUs for managing a medical practice. This article addresses how RVUs differ from encounters and fees in terms of measuring provider productivity and resource consumption. RVUs empower practice administrators to objectively measure and quantify a medical practice's physician productivity and performance data versus traditional productivity measures such as office visits, net charges, net collections, etc. The Resource-Based Relative Value Scale (RBRVS) RVU work component is specifically designed to measure physician (and midlevel provider) effort and the degree of independent decision-making skill required for performing a procedure; therefore, productivity is directly linked to provider coding. PMID- 12122813 TI - How good are the internal controls in your group practice? Ten questions to contemplate. AB - Internal controls are the methods and procedures used by any business to prevent or detect errors, safeguard assets (especially cash) from being misappropriated, and encourage staff adherence to prescribed managerial policies. Internal controls in a medical practice differ depending on the size and complexity of the practice. The key, however, is that they prevent or detect errors and efforts to circumvent the established policies and procedures of the organization. How good are the internal controls in your group practice? This article identifies ten questions you should use to evaluate your risk of asset misappropriation. PMID- 12122814 TI - Automated claim and payment verification. AB - Since the start of managed care, there has been steady deterioration in the ability of physicians, hospitals, payors, and patients to understand reimbursement and the contracts and payment policies that drive it. This lack of transparency has generated administrative costs, confusion, and mistrust. It is therefore essential that physicians, hospitals, and payors have rapid access to accurate information on contractual payment terms. This article summarizes problems with contract-based reimbursement and needed responses by medical practices. It describes an innovative, Internet-based claims and payment verification service, Phynance, which automatically verifies the accuracy of all claims and payments by payor, contract and line item. This service enables practices to know and apply the one, true, contractually obligated allowable. The article details implementation costs and processes and anticipated return on investment. The resulting transparency improves business processes throughout health care, increasing efficiency and lowering costs for physicians, hospitals, payors, employers--and patients. PMID- 12122815 TI - Preparing for the brave new world, Part 4: "Minimum necessary" disclosures under HIPAA. AB - "Minimum information given with maximum politeness," Jacqueline Kennedy once directed a White House press secretary. Decades later, a pending federal health privacy rule says nothing about courtesy but does explicitly require a "minimum necessary" standard for most disclosures of personally identifiable information. Physician practices that understand these two key words will have a head start in complying with the privacy regulations, which were published in December 2000 under authority of the Health Insurance Portability and Accountability Act of 1996 (HIPAA). This article, the final installment of a four-part series on how the HIPAA privacy rule will affect the day-to-day lives of physicians and their staffs, focuses on what the U.S. Department of Health and Human Services (DHHS) means by "minimum necessary." PMID- 12122816 TI - Professional liability: informal consults give rise to the physician-patient relationship? PMID- 12122817 TI - Informal consultations: do new risks exist with this age-old tradition? AB - This article cites three court decisions involving physicians who provided an informal or "curbside" consultation. The cases demonstrate the medico-legal implications of physicians engaging in informal consultations. The author then discusses the growing risk of liability for physicians who participate in this traditional practice. PMID- 12122818 TI - Is bigger always better? The optimal size of a group practice. AB - The past decade saw several attempts to consolidate physician practices, but this sector remains one of the last cottage industries in the United States. This article develops a framework for analyzing the optimal size of a physician practice. The framework addresses technological factors (e.g., economies of scale and scope), behavioral factors (e.g., changing physician goals, costs of organizing and operating a practice), and market-driven factors (e.g., managed care contracting). Existing empirical research suggests three "optimal" sizes of practices: 5-10 physicians, based on economies of scale and decision-making; 20 30 physicians, based on economies of scope and initial development of a corporate structure; and 80+ (multi-specialty) physicians, which can create an system of referrals and utilization. The article concludes with observations about the challenges to physician practices as they grow. PMID- 12122819 TI - Becoming a better delegator. AB - Most medical practice managers know that delegation is a useful practice management tool to streamline both personal and practice efficiency. However, delegation is often underused, misused, and misunderstood. What, precisely, should be delegated in a medical practice and to whom? What are some of the obstacles to successful delegation, and how can the astute medical practice manager identify and overcome them? Which tasks should not be delegated? Finally, why do members of the professional practice staff sometimes resist delegation? In this article, the author provides answers to these intriguing questions, as well as a useful self-quiz to rate delegation skills. In addition, the author provides strategies and sample language you can use with your staff to make your own delegated tasks and responsibilities more enthusiastically accepted. PMID- 12122820 TI - Incentives: supercharge your staff. PMID- 12122821 TI - Trends in the consumer price index. PMID- 12122822 TI - Internet visits: a new approach to chronic disease management. AB - First Health has become the first national managed care company to establish reimbursement for Web-based Internet consultations between patients and their physicians. Initially limited to chronic conditions such as diabetes, asthma, and congestive heart failure, the program will reimburse up to 24 consultations per year at a rate of $25 each. Privacy issues have been addressed by utilizing secure network procedures. First Health believes that this initiative will enhance subscribers' well-being and potentially reduce the rates of hospitalization and even lengths of stay. PMID- 12122823 TI - Thoracic imaging in the ICU. Interventional radiology. AB - Radiology in the intensive care unit (ICU) patient is dominated by plain x-rays, with noteworthy findings prompting further imaging and possible intervention. This chapter discusses interventional and minimally invasive techniques used to treat pleural, mediastinal and pulmonary parenchymal problems commonly encountered in the ICU. PMID- 12122824 TI - Nutritional support. AB - Despite the increasing obesity of the American population, many chronically ill patients are malnourished. When this malnutrition is combined with the hypermetabolic response and protein catabolism of an acute event, such as an operation, nutritional support becomes an important facet for optimal critical care. This chapter reviews the basic tenants of nutritional support with special emphasis on patients with pulmonary compromise. Important aspects of caloric and protein support are discussed and enteral nutrition is emphasized because of its numerous advantages and documented improvement in outcome. PMID- 12122825 TI - Postthoracotomy pain management. AB - The following techniques appear efficacious in controlling postthoracotomy pain and reducing the amount of systemic opioids consumed: continuous intercostal blockade, paravertebral blockade, and epidural opioids and/or anesthetics. The combination of thoracic epidural opioid and local anesthetic is very effective at relieving postthoracotomy pain, however, considerable experience is required for insertion of the thoracic epidural catheter and postoperative respiratory monitoring. Intercostal and paravertebral catheters can be inserted intraoperatively under direct visualization, to reduce complications of insertion. One-time intraoperative intercostal blockade may effectively reduce postoperative pain in the first day, but is not a practical long-term method for postthoracotomy pain. The effectiveness of interpleural analgesia, even with proper technique, appears inferior to epidural and other regional techniques. We have incorporated the principles outlined in this review into our general thoracic surgery protocol, as detailed in Fig. 1. Every patient is assessed preoperatively for epidural catheter placement. Contraindications include low platelet count (< 100,000), abnormal coagulation profile, medicinal anticoagulation (aspirin and nonsteroidal anti-inflammatories are not contraindications), bony spinal abnormalities, or neurological disorders. The T5/6 interspace is our preferred level, but T10 can work well with an increased dose of bupivacaine. Upon completion of the muscle sparing, minimal-access thoracotomy, we close the wound and perform a percutaneous intercostal nerve block (two ribs above and three below the incision). We then use patient controlled epidural analgesia, with a basal infusion of bupivacaine and hydromorphone. To supplement inadequate or nonfunctioning epidurals, intravenous patient-controlled opioids are added. When choosing an approach to postthoracotomy pain management, the thoracic surgeon and anesthesiologist must consider the following: (1) the physician's experience, familiarity and personal complication rate with specific techniques; (2) the desired extent of local and systemic pain control; (3) the presence of contraindications to specific analgesic techniques and medications; and (4) availability of appropriate facilities for patient assessment and monitoring postthoracotomy. Refinements in surgical technique including limited or muscle-sparing thoracotomy, video assisted thoracoscopic surgery (VATS) and robotic surgery may lessen the magnitude of postthoracotomy pain. We encourage all thoracic surgeons to be knowledgeable of available techniques and maintain a protocol to generate a database for periodic assessment of safety and efficacy. PMID- 12122826 TI - Mechanical ventilatory support. AB - This chapter reviews the most updated knowledge regarding mechanical ventilation, its' indications and its' features both diagnostic and therapeutic. Further, the various modes of mechanical ventilation are described. The reader will also gain insight into the pathophysiology of various disease processes and the mode of ventilation that may be the most helpful in their treatment. Weaning is also discussed as well as a relatively new type of ventilation, non-invasive. PMID- 12122827 TI - Acute respiratory distress syndrome epidemiology and pathophysiology. AB - Acute respiratory distress syndrome is a devastating syndrome of lung injury following known risk factors, with a persistently high mortality. A consensus conference definition of ARDS has been adopted by clinical researchers, but potential problems remain. ARDS may represent more than one entity, and radiographic and mechanical differences between pulmonary versus extrapulmonary initiated ARDS have been described. There is increasing recognition of inflammatory mediators in the pathophysiology of acute lung injury. Surfactant abnormalities contribute to the associated lung dysfunction. A growing body of evidence supports the presence of VILI and a potential mechanism for developing MOSF, and has led to new management strategies. The importances of apoptosis to the repair process, and mechanisms that may lead to persistent fibrosis, such as the activation of the coagulant pathway with fibrin deposition, are increasingly recognized. PMID- 12122828 TI - Management of the acute respiratory distress syndrome. AB - Significant advances have occurred in the knowledge of the pathogenesis of ARDS. It is now recognized that ARDS is a manifestation of a diffuse process that results from a complicated cascade of events following an initial insult or injury. Mechanical ventilation and PEEP are still important components of supportive therapy. To avoid ventilator-associated lung injury there is emphasis on targeting ventilator management based on measurement of pulmonary mechanics. For those with resistant hypoxia and severe pulmonary hypertension adjunctive modalities, such as prone positioning and low-dose iNO, may provide important benefit. Alternative modes of supporting gas exchange, such as with partial liquid ventilation and extracorporeal gas-exchange, may serve as rescue therapies. Advances in cell and molecular biology have contributed to a better understanding of the role of inflammatory cells and mediators that contribute to the acute lung injury and the pathophysiology of the syndrome that manifests as ARDS. Based on this new understanding, the potential targets for intervention to ameliorate the systemic inflammatory response have proliferated. Examples include the cytokine network and its receptors, antioxidants, and endothelins. Apart from the challenge of testing these agents in experimental models, it seems likely that determination of the optimum combination of agents will become an equally important endeavor. A particular challenge is to develop better methods of predicting which of the many at-risk patients will go on to full-blown ARDS and MODS, thereby targeting subgroups of patients most likely to benefit from anti inflammatory therapies. Similarly, the adverse effects of immunosuppressive therapy may be diminished by improved, perhaps molecular, techniques to detect microbial pathogens and permit differentiation between Systemic inflammatory response syndrome and sepsis. PMID- 12122829 TI - Extracorporeal membrane oxygenation for severe respiratory failure. AB - The use of extracorporeal technology to accomplish gas exchange with or without cardiac support is based on the premise that "lung rest" facilitates repair and avoids the baso- or volutrauma of mechanical ventilator management. Extracorporeal membrane oxygenation (ECMO), a modified form of cardiopulmonary bypass, has been shown to decrease mortality of neonatal, pediatric and adult respiratory failure and is capable of total gas exchange. In neonates, over 20,638 patients have been treated with an overall survival of 77% in a population thought to have 78% mortality. PMID- 12122830 TI - Diagnosis and management of pneumonia in the intensive care unit. AB - Ventilator-associated pneumonia is a significant contributor to excess morbidity and mortality in the ICU. Alteration of physical defenses, bacterial flora, and immune response contribute to the susceptibility of the critically ill ventilated patient. Controversies regarding its definition continue to complicate the understanding of the extent of the disease and evaluation of the outcome of therapies. Traditional parameters, such as clinical suspicion with standard tracheal aspirates, are associated with overuse of antibiotics, a prime risk factor for development of resistant organisms and increased mortality. Global emergence of resistance is also a major concern. Whether invasive methods have any substantial clinical benefit either through improved patient outcomes or lower rates of resistance remains to be proved. In patients who fail to respond to therapy, a search for nonpulmonary infections and noninfectious causes of pulmonary infiltrates is essential. Finally, preventative measures offer additional areas of investigation that could favorably impact on the incidence of infection. PMID- 12122831 TI - Perioperative care of the lung transplant patient. AB - Improvements in the perioperative management of lung transplant recipients have produced a 90% survival in the first 30 days following surgery. Detailed attention to donor organ procurement and preservation of the allograft are important in ensuring an early successful outcome. Early antibacterial administration based on donor or pretransplant cultures and antiviral therapy in CMV-negative recipients assist in avoiding early infectious complications. Development of hypoxemia or hemodynamic instability in the perioperative period requires a rapid, systematic evaluation with attention to mechanical, immunologic, or infectious causes. Nonpulmonary complications are not infrequent in lung transplant recipients. PMID- 12122832 TI - Pulmonary embolism. AB - Pulmonary embolism (PE) is a common problem for which prompt diagnosis and treatment is essential to minimize mortality. The clinical presentation is more variable than sudden dyspnea and chest pain, especially in the critical care patient. Recognition of venous thromboembolic (VTE) risk factors can help develop a good clinical suspicion for PE. A wide range of diagnostic tests are available to the clinician. The ventilation/perfusion scan, pulmonary arteriogram, and lower extremity investigations are still important for diagnosis. Other noninvasive tests such as spiral CT with venography, echocardiography, and D dimers are becoming more accepted. Heparin is the mainstay of PE therapy, but thrombolytic treatment may be lifesaving in the unstable patient. VTE prophylaxis should be considered in all post-operative or critical care patients. PMID- 12122833 TI - Cardiac arrhythmias in cardiothoracic surgery. AB - Most patients with cardiopulmonary disease are predisposed to develop perioperative arrhythmias with the individual patient risk depending upon the type of operative procedure performed, the risk profile of the patient, and the complexity of the post-operative course. There are several management options that may tend to prevent perioperative arrhythmias that should be considered in certain patient subsets. Most important of these is the use of beta-blocker therapy before and after operation in patients with coronary risks factors undergoing non-cardiac thoracic procedures and in patients having coronary artery bypass grafting. The common supraventricular arrhythmias including atrial fibrillation and flutter, multifocal atrial tachycardia, and paroxysmal supraventricular tachycardia must be properly diagnosed and treated appropriately. Placement of atrial pacing wires for use after open cardiac surgery is of great value both for diagnosis, and in some cases, for treatment of arrhythmias. Fortunately, serious life threatening ventricular arrhythmias occurs less commonly but the clinician must recognize and correct important predisposing factors and know how to treat these when they occur. A specific protocol for arrhythmia management that sets guidelines for drug choice and therapies for each of the common arrhythmias is useful for clinicians and adds predictability to patient care. PMID- 12122834 TI - Taking medicine back: Dx and Rx for orthopedic surgeons. AB - The present day practice of medicine is cause of considerable malaise for physicians. This article will invoke the ancient proverb, "Physician Treat Thyself". In other words this article addresses the problems from a diagnostic and therapeutic perspective, working toward a permanent cure. The history of medical practice groups, in-office surgery, and ancillary services is reviewed. The co-morbidity of health maintenance organizations, Evaluation and Management requirements, and cost shifting are mentioned. Finally a list of symptoms and the appropriate therapeutic measures are administered to control costs, increase income, and re-establish some form of authority in the practice of medicine. PMID- 12122835 TI - Improving office operations and maximizing reimbursement for the orthopedic practice. AB - After assessing the efficiency of the medical/surgical practice, the physician should work on implementing goals with the practice manager or administrator. To assist in the implementation of these goals, the physician should be involved in strategic planning. By taking each project one step at a time, and through better communication and practice efficiency, a medical office can survive in today's managed care and complex health care environment. PMID- 12122837 TI - Correct coding for the orthopedic surgeon. AB - Coding accurately is one of the main principles of a successful practice. Some changes that we will see shortly include deletion of the term "separate procedure," deletion of the term "with and/or without," deletion of the term "any method," revision of the criteria for choosing E/M levels, and 52 new and revised Hand Surgery codes. Some other changes to come will be category II and category III codes. More changes are occurring as this is written, and the best advice is to stay tuned. It is obvious to the authors that coding is mainly for reimbursement purposes. The orthopedic surgeon must remain vigilant and must not pass this task on to someone else. Ignorance of coding methods is not an excuse [2]. We must all watch carefully and speak up when necessary. In this day of decreasing reimbursement, we can all increase our revenue stream without working any harder if we code our work properly, completely, and promptly. PMID- 12122836 TI - The computer in office medical practice. AB - There will continue to be change and evolution in the medical office environment. As voice recognition systems continue to improve, instant creation of office notes with the absence of dictation may be commonplace. As medical and computer technology evolves, we must continue to evaluate the many new computer systems that can assist us in our clinical office practice. PMID- 12122838 TI - Making your orthopedic office profitable through the addition of ancillary services. AB - With the precipitous decline of orthopedic reimbursement over the past six years, it will be critical to the success of the majority of orthopedic groups to improve their revenue stream in the future. This will involve improving management, reducing overhead, and adding ancillary services to obtain facility fee revenue. Ancillary service possibilities for the orthopedist include an in office surgicenter with a pain center, MRI, physical therapy with orthotics and braces, occupational health department, pharmacy, and independent medical examination company. PMID- 12122839 TI - In-office MR imaging. AB - The development of new metal alloys, along with more innovative magnet technology, has permitted the construction of smaller magnets for magnetic resonance (MR) systems, which, in turn, has allowed development of MR imaging systems designed to be physically smaller than conventional whole-body MR imaging systems. These specialized devices are commonly referred to as "niche," "dedicated," or "extremity" MR imaging systems. Performing MR imaging procedures with this type of system offers distinct advantages that include reduced overall costs, more convenient installation and siting, and greater patient comfort and safety. Importantly, these critical features permit extremity MR imaging systems to be readily utilized in an "in-office" setting. This article will provide an overview of the technical aspects and clinical applications for extremity MR imaging systems, present patient management issues, and discuss the economic and practical considerations of the use of extremity MR imaging systems in an in office environment. PMID- 12122840 TI - Setting up an in-office independent medical examination company. AB - In a time of declining reimbursement for patient care services, establishing an in-office IME company enables orthopedic practices to generate additional revenue to subsidize clinical activities without compromising the credibility and integrity of their physicians; however, the decision to enter the medical/legal consultation business should be considered carefully. A thorough analysis of the industry, applicable laws, costs-versus-benefits, and the local marketplace is critical in helping the practice to evaluate the feasibility of establishing the new business and develop a product that is well differentiated. The practice should approach the day-to-day management of the IME company with the same careful attention that it pays to the management of its orthopedic service. This includes creating a staffing and information technology infrastructure that supports the new business and allows for its growth. An attitude of continuous learning whereby the physician-reviewer seeks out information about the customer's needs and market shifts enables the practice to respond swiftly to these needs and shifts and further position itself as an innovative provider of medical/legal services. PMID- 12122841 TI - Regulation of ancillary services. AB - Operation of ancillary services is highly regulated. The regulations are not always logical, but they are navigable. Be sure to work with a health care attorney who is familiar with the rules. Be careful to fully disclose all relevant facts. Most importantly, ask a lot of questions. The rules are complicated, but they are understandable. By being inquisitive, you will be able to determine whether your legal or consulting expert has a good understanding of the rules. If you work together, it is usually possible to structure the ancillary service in a way that can be both safe and profitable. PMID- 12122842 TI - The efficient, enjoyable, and profitable orthopedic practice. AB - For the practicing orthopedist to achieve his personal practice goals of developing an efficient, enjoyable, profitable orthopedic practice, he must commit himself to restructuring his practice environment. This restructuring involves bringing in-house as many ancillary services as the practice can support, creating efficiencies in the clinic and surgical practice, and increasing the size of the group to attempt to exert some control of the payers in the marketplace. By doing so, the orthopedist will improve the quality of his practice, increase his income, and dramatically improve the quality of his life. PMID- 12122843 TI - Legislative issues. The federal financing and regulation of physical medicine and rehabilitation services. AB - This article describes major elements in the federal legislative system of the United States affecting the financing and delivery of rehabilitation and related services to persons with disabilities. It deals with existing federal statutory law and regulations, as well as pending legislation dealing with patients rights in health care that has been passed by both bodies in Congress but is not law as of late 2001. The recently revised federal Medicare law financing much of the inpatient hospital rehabilitation and outpatient rehabilitation services is described in detail, as is the Medicare law financing graduate medical education. The article also discusses the Americans with Disabilities Act as it affects PMR professionals and the application of the Stark Physician Referral law as it applies to typical business transactions in the field of physical medicine and rehabilitation services. This information should assist PMR professionals in better understanding the legal regulatory system in which they provide services and support to their patients. PMID- 12122844 TI - Informed consent in physical medicine and rehabilitation. The physician/patient relationship--the doctor as a fiduciary. AB - This article reviews the principle of informed consent and the ethical and legal bases upon which it rests. The process of obtaining an appropriate informed consent is explored, and the elements that make a consent valid are delineated. The principles of substitute decision-making and the special rules applied to circumstances such as emergency, therapeutic privilege, refusal of consent, and medical necessity are discussed. The concept of the relationship between physician and patient as a fiduciary relationship is explored as the fundamental basis for the modern doctrine of informed consent. PMID- 12122845 TI - An introduction to risk management. AB - Risk management is formally defined as the process by which an organization assesses and addresses its risks. Historically, the role of risk management has been associated with insurance-buying, occupational safety and health, and legal liability management. In recent years managers and physicians alike have begun to recognize that organizational risks are pervasive, that these risks are extraordinarily diverse and complex, and that these risks are not just confined to "insurable" or accident-related situations. They may include risks arising from actions of regulatory bodies, third party payers, hospitals, partners, and employees, in addition to the physiatrist's personal or business investment, management and clinical practice. Furthermore, changing customer and patient preferences and/or expectations make the assessment of risk an even more dynamic and continuous process. This article describes the formal risk management process and suggests ways that physiatrists can apply risk management to their business and clinical practice. In developing this description, physiatrists and their office managers will learn about the overall goals and objectives of risk management, the challenge of identifying and analyzing risks, the tools and treatment options available, and the means by which risk management efforts are effectively implemented. PMID- 12122846 TI - Legal issues in alternative health care. AB - This article reviews the use of alternative and complementary health care practices by orthodox medical practitioners. Many alternative modalities such as chiropractic, acupuncture, naturopathic, and homeopathic treatments are available, and modern patients who are increasingly "consumer-oriented" and educated may inquire about these treatments. The article examines issues of informed consent, standards, procedures, and liability as they relate to a medical practice that opts to provide alternative treatments. PMID- 12122847 TI - The independent medical examination. AB - The physiatrist, owing to expertise in impairment and disability analysis, is able to offer the medicolegal process considerable assistance. This chapter describes the scope and process of the independent medical examination (IME) and provides an overview of its component parts. Practical guidelines are provided for performing a physiatric IME of professional standard, and for serving as an impartial, expert witness. Caveats are described regarding testifying and medicolegal ethical issues along with practice management advice. PMID- 12122848 TI - Life care planning. AB - Physicians may be asked to help plan long-term needs of patients with catastrophic injury. It is crucial to know the life expectancy and be intimately familiar with the needs of the disabled person for whom one is planning. This article uses two diagnostic groups as models to illustrate the process: one a spinal cord injured adult and the other a child with cerebral palsy and mental retardation. We provide examples of some of the specific types of needs for these two groups of individuals. PMID- 12122849 TI - Life expectancy determination. AB - The estimate of life expectancy following a personal injury is probably one of the most important factors in determining the final quantum of damages. It is a calculation fraught with difficulties. This article endeavours to outline some general factors that aid prediction of life expectancy, and also discusses the evidence from the few long-term studies currently available. PMID- 12122850 TI - Determination of loss of quality of life. AB - Physiatrists are a valuable resource in legal settings, where assessment of functional capacity to perform work and of future medical needs must be determined. Physiatrists help determine what future medical care is needed to restore and maintain an individual at the maximum level of life function. This article focuses on the use of a quality of life (QOL) rehabilitation model, rather than a medical model, for enhancing functional performance, modifying environments, and facilitating patient coping. We discuss use of the QOL model to describe and influence a patient's physical, psychological, cognitive, vocational/economic, and social/leisure domains. PMID- 12122851 TI - Merits and shortcomings of the American Medical Association Guides to the Evaluation of Permanent Impairment, 5th edition. A physiatric perspective. AB - We review in this article the most recent edition of the American Medical Association Guides to the Evaluation of Permanent Impairment (AMA Guides) from a physiatric perspective. Important general changes within the framework from the 4th to the 5th edition are highlighted. Those sections of the AMA Guides most often consulted by physiatrists are examined in critical detail, including sections dealing with the spine, upper and lower extremities, neurologic impairments, and impairments due to pain. PMID- 12122852 TI - Medicolegal causal analysis. AB - This article reviews basic tort law concepts, which medical evaluators should be familiar with when working in the medicolegal environment. To facilitate medicolegal causal analysis, the concept of the health claim statement or argument is proposed. We describe medicolegal causal analysis model, which should assist the medical evaluator's determination of the relationship between an alleged accident and personal injury. This practical model minimizes bias and ensures that inferences are supported by factual medical evidence. PMID- 12122853 TI - Who are the nurses at the bedside? PMID- 12122854 TI - All stem cells are not alike. PMID- 12122855 TI - ONS supports bedside nurses with many professional resources. PMID- 12122856 TI - One voice against cancer advocates for increased federal funding to defeat cancer. PMID- 12122857 TI - Technology in practice can make the lives of oncology nurses at the bedside easier. PMID- 12122858 TI - Hepatitis B and hepatitis C. AB - Hepatitis B and C are worldwide infectious hepatitides which are distinct in terms of epidemiology and molecular biology, but which may be quite similar in terms of clinical manifestations and histopathology, in both the acute and chronic stages. Hepatitis B virus (HBV), the human prototype of the Hepadnaviridae family of viruses is not directly cytopathic and viral hepatitis is caused by the cellular immune response to HBV. Patients infected with HBV may also have hepatitis D (delta) virus (HDV) infection, either as co-infection or a superinfection. Hepatitis D virus does not infect independently. Better control of HBV has also led to a decline in the incidence of HDV. Hepatitis C virus (HCV) is on of the Flaviviridae family of viruses, and is quite heterogeneous, with six major genotypes and more than 100 subtypes. Hepatitis C virus circulates as quasispecies that result from mutations accumulated over time, which probably enable HCV to replicate efficiently or resist immune mechanisms. Quasispecies have complicated vaccine development. Both HBV and HCV will recur in the transplanted liver. The risk of developing hepatocellular carcinoma is significantly greater in both HBV- and HCV-infected individuals. PMID- 12122859 TI - Scoring of chronic hepatitis. AB - Grading of the severity of chronic hepatitis and staging of its structural consequences are widely used in clinical trials of therapy and in research. Simple and complex methods are available. Intra- and interobserver variation can be reduced but not eliminated, because grading and staging are essentially subjective. The data are categorical rather than numerical and must be treated accordingly. Morphometry of fibrous tissue offers a different approach to biopsy assessment. PMID- 12122860 TI - Autoimmune hepatitis and overlap syndromes. AB - Autoimmune hepatitis (AIH) is an immune-mediated, autodestructive liver disease with hepatocytes as target cells, mostly affecting young women. Primary biliary cirrhosis (PBC) is also regarded as an autoimmune liver disease with bile duct epithelia as the target cells, resulting in a continuous loss of bile ducts. Both diseases may occur simultaneously in their full manifestations in about 10% to 20% of cases, thus constituting an overlap syndrome with PBC directing the course of the disease. AIH may also occur simultaneously with primary sclerosing cholangitis (PSC), with a frequency of between 2% and 8% of patients with PSC. In most cases, AIH precedes manifestation of PSC. In children, the overlap syndrome of AIH and PSC seems to make up an entity of its own: autoimmune sclerosing cholangitis. PMID- 12122861 TI - Primary biliary cirrhosis and other ductopenic diseases. AB - Several distinct conditions are characterized by a reduction in the number of small and medium-sized intrahepatic bile ducts. These diseases are associated with progressive cholestasis, which in turn leads to biliary fibrosis and ultimately cirrhosis. The best-characterized ductopenic condition in adulthood is primary biliary cirrhosis (PBC) for which there is now strong evidence of an autoimmune cause. The antigenic targets are epitopes on proteins of the 2-oxoacid dehydrogenase complex within mitochondria. Some of these proteins appear to be aberrantly expressed at the surface of cholangiocytes in PBC. The basis for the breakdown in tolerance remains uncertain, although there is recent evidence to indicate that apoptosis may play a key role at early stages in the pathogenesis of the disease. Related conditions include autoimmune overlap syndromes and AMA negative PBC (autoimmune cholangitis). Primary sclerosing cholangitis is clinically and histologically distinct, although there is evidence that it also may have an immune-mediated cause. Ductopenia may also arise on the basis of drug induced injury; the best example of this is progressive cholestasis complicating chlorpromazine therapy. PMID- 12122862 TI - Drug hepatotoxicity. AB - Drug-induced liver disease is a relatively common, but often unrecognized, cause of liver injury, primarily because the diagnosis is often not entertained clinically. In addition, drugs are great imitators, capable of producing nearly any clinical scenario and histopathologic lesion. Thus, when dealing with a liver biopsy from a patient with an undiagnosed liver disease, the diagnosis of drug hepatotoxicity is made by first having a high index of suspicion, and then by careful correlation of histopathologic findings with both clinical and laboratory data and with a search for appropriate precedents in the medical literature. PMID- 12122863 TI - Alcoholic and nonalcoholic steatohepatitis. AB - The constellation of histopathologic lesions that characterize alcoholic and nonalcoholic steatohepatitis has been well described and has served as the basis for clinical diagnosis, natural history studies, and experimental models for analyses of etiopathogenesis. The lesions common to both entities include, to varying degrees, steatosis, liver cell ballooning, lobular inflammation with a notable component of polymorphonuclear leukocytes, and a characteristic form of fibrosis that is initially located in the perisinusoidal regions of acinar zone 3. Cirrhosis with or without steatosis or steatohepatitis may occur in both entities. Mallory's hyaline is common but not necessary; megamitochondria and varying amounts of iron may be observed in either process. Hepatocellular carcinoma is a recognized complication of both processes, albeit with greater frequency in the former. Alcoholic hepatitis may present with more severe clinical and histologic manifestations than the nonalcoholic counterpart, including significant morbidity and mortality. The perivenular lesions collectively referred to as sclerosing hyaline necrosis are markers of severity, and are not common in nonalcoholics. In many instances, however, the microscopic lesions of these two processes are similar, likely as a reflection of common pathogenetic pathways, and the distinction between the two is ultimately clinically derived. PMID- 12122864 TI - Common diagnostic problems in pediatric liver pathology. AB - The role of the pathologist in dealing with common problems of liver disease in children is likely to change dramatically as the molecular genetic revolution progresses. For example, microchip arrays for genes involved in bile salt synthesis and transport will pinpoint the specific mutations responsible for infantile cholestasis and similar methods will sort out infectious agents of acute and chronic hepatitis. But even as biochemistry, microbiology, and immunology laboratories already provide essential diagnostic information in such settings, informed histopathologic interpretation will continue to guide investigations of etiology and therapeutics and will remain an important medical necessity [95,96,100,102,104]. PMID- 12122865 TI - Inherited metabolic diseases of the liver. AB - Many inherited metabolic diseases affect the liver in neonates, children, or adults. The histopathologic changes are diverse and may be acute or chronic. They can be considered primary (when the injury is from the cytopathic effect of an accumulated metabolite) or secondary (e.g., an infection caused by an immune deficiency). All forms of liver disease are described: for example, intrahepatic cholestasis, neonatal hepatitis with giant-cell transformation, paucity of bile ducts, steatosis, steatohepatitis, necroinflammatory diseases (acute or chronic), fibrosis, cirrhosis, and neoplasms (benign or malignant). Familiarity with the morphologic changes is important in clinicopathologic correlation, diagnosis, and understanding of pathogenetic mechanisms. PMID- 12122866 TI - Iron overload disorders. AB - Because hepatic siderosis is a frequent finding, there is a risk of making it trite when elaborating the pathology report. Iron is increasingly considered an important cofactor of morbidity. Its finding in hepatic cells must be recognized, indicated, qualified, quantified, and interpreted. A systematic reasoning based on a strict semiological approach allows for guiding the clinician. Iron overload syndromes do not amount to genetic hemochromatosis only. PMID- 12122867 TI - Dysplastic nodules and hepatocarcinogenesis. AB - In the last decade, careful examination of explanted cirrhotic livers in liver transplant centers around the world has confirmed the findings of the earlier Japanese investigators: DNs (by this or any other name) represent hepatic, premalignant lesions in chronic liver disease. Careful examination of their gross and microscopic morphologies has led to the hypothesis of precirrhotic, spreading clonal expansions that are resistent to scarring, and that result in neoplastic islands of hepatic parenchyma. The resultant distinctive nodules, often marked by features suggestive of their clonality (such as increased pigment), are at increased risk for subsequent carcinomatous events, thereby giving rise to HCC. Specialized molecular and immunohistochemical studies confirm many aspects of this hypothesis. In suggesting that some aspects of DN pathophysiology are not integral to the carcinogenetic pathway (i.e., inhibition of HSC inactivation), this hypothesis serves a broader purpose, explaining the various settings in which early HCCs are found in cirrhotic explants and in wedge resections of radiographically defined lesions. Discrepancies between Japanese and non-Japanese investigations regarding dysplasia and early HCCs reflect not different biologic pathways but differences in detection, interpretation, and application of nomenclature. These differences may fade away as more international collaborative work brings investigators of diverse nationalities into regular contact, supporting movement toward a commonly acceptable nomenclature and set of diagnostic criteria. Ultimately, an understanding of the pathophysiology of these lesions, through more detailed molecular and physiologic studies, should lead to more efficient and available early detection, and perhaps chemoprevention approaches to hepatic malignancy. PMID- 12122868 TI - Benign liver tumors. AB - Benign liver tumors are becoming a subject of great interest because increased access to medical care has allowed discovery of many incidental focal lesions. These tumors may be hepatocellular, biliary, or stromal in nature. Several new lesions have been described in recent years causing a need to reassess the pathogenesis and classification of hepatic tumors. Hepatocellular nodules may be neoplastic or a regenerative response to injury. The size and structure of regenerative nodules varies with the distribution and severity of the hepatic injury, leading to a complex classification. Variation in fat content is easily detected on ultrasonography as focal fatty change and focal fatty sparing. Biliary and stromal cells also produce neoplastic or regenerative lesions. Biliary lesions are often cystic in nature. Stromal lesions are varied because of the many nonparenchymal cell types in the liver. PMID- 12122869 TI - Malignant liver tumors. AB - Primary malignant liver tumors can arise from different components of the liver, such as hepatocytes, bile duct epithelium, neuroendocrine cells, and mesenchymal cells. A specific diagnosis frequently can be suggested from imaging studies, but biopsy remains the gold standard for definitive diagnosis of liver tumors. Clinical history of chronic liver disease, known risk factors, or other diseases are of great importance. Patient's age is also an important discriminating feature because several tumors such as hepatoblastoma, mesenchymal hamartoma, and infantile hemangioendothelioma, are found predominantly in pediatric populations, whereas cholangiocarcinoma and hepatocellular carcinoma are rare in pediatric populations. PMID- 12122870 TI - Diagnostic issues in liver transplantation pathology. AB - Liver biopsy is used to determine the pathogenesis of liver dysfunction after liver transplantation. One or more causative factors may be identified on biopsy. The pathologist must be familiar with the histopathology of acute rejection to differentiate it from other potential complications, including biliary obstruction, intercurrent cytomegalovirus hepatitis, or recurrent disease. Consensus documents from the Banff international panel provide useful guidelines for the appropriate grading of acute and chronic rejection. PMID- 12122871 TI - The role of immunohistochemistry in diagnosis. AB - Immunohistochemistry is a strong tool in hepatopathologic diagnosis: the technique is relatively simple and inexpensive. New and very sensitive detection methods have been recently developed (e.g., the EnVision technique and the microwave antigen retrieval method). This article discusses the role of immunohistochemistry in differentiating chronic cholestatic diseases from chronic hepatitis and in characterizing infectious agents. Algorythms for the typing of lymphomas and for the differentiation of primary tumors versus metastases are proposed as well. The immunohistochemical criteria for the diagnosis of premalignant lesions are discussed. PMID- 12122872 TI - Liver histopathology. PMID- 12122873 TI - [Neurologic complications of aortic dissection]. AB - INTRODUCTION: Beside the damages of the cardiovascular system the lesions of the the nervous system are the most common complications of aortic dissection. This is usually an early event, therefore the dissection of the aorta may manifest itself as an acute primary neurologic disease. The aim of this study is to describe the frequency and distribution of acute neurologic symptoms occurring in aortic dissection and the distribution of their clinico-pathologic features and to establish correlations between these and the acute in-hospital mortality as well as to discuss available diagnostic and therapeutic possibilities. PATIENTS AND METHODS: The study was based of 95 cases of acute dissection of aorta (with additional three later events of redissection), observed in a longitudinal study over a period of 29.5 years, in a population of 106,000 (in Western Hungary). RESULTS: Of the 95 patients 20 (21%) died before admission. Neurological complications were observed in 30 of the 75 patients admitted to hospital (40%). Symptoms involving the central nervous system were found in 24 patients, affecting the spinal cord in two and the peripheral nervous system in four cases. The dissection of the aorta was diagnosed in vivo only in 22 out of the 75 patients who died in hospital (29%). 53 patients (71%) without correct diagnosis received supportive therapy only. The average survival time of the 21 patients with proximal dissection of aorta was 48.5 hours. The survival time of 23 patients with the same type of dissection involving the vessels of the aortic arch was 22.2 hours. This difference in survival time was significant (p = 0.0152). 20 of 23 patients (87%) in this group showed signs of neurologic damage confirming earlier experience that neurological complications can seriously worsen the otherwise already catastrophic prognosis of aortic dissection. CONCLUSIONS: The study brought compelling evidence for the need for early diagnosis and rapid transfer of patients to appropriate cardiac surgery centers for definitive diagnosis and therapy. PMID- 12122874 TI - [Investigation of insertion/deletion polymorphism of the ACE gene in stroke patients]. AB - INTRODUCTION: This is the first Hungarian paper on the insertion/deletion polymorphism of ACE gene in stroke patients. According to literature data, the role of this polymorphism is controversial in the pathogenesis of stroke. The aim was to study the prevalence of the polymorphism in healthy persons and in stroke patients. PATIENTS AND METHODS: Blood samples from 173 unrelated healthy donors and 253 stroke patients were investigated by polymerase chain reaction (PCR). PrevIous stroke was documented by CT or MRI and CDS. A routine questionnaire was used to study previous vascular events and the risk profile of patients. RESULTS: I/I allele was found in 20%, I/D 52% and D/D 28% in the healthy group. Prevalence of the pathologic D/D allele did not differ between healthy and patients group (28% and 27%, OR: 0.88, and in subgroup age under 50 years OR: 1.00). No correlation was found between D/D and conventional risk profile but a positivE correlation was found in young patients having D/D and hyperlipidemia (p < 0.05) and hyperfibrinogenemia (p < 0.05). D/D prevalence was found higher in patients with family anamnesis of myocardial infarction (p < 0.05). Very low prevalence of D/D allele was found in cardiogen embolic group (p > 0.05). CONCLUSIONS: The ACE polymorphism does not seem to be an independent risk factor for stroke. However, in young stroke patients with D/D allele, hyperlipidemia and/or hyperfibrinogenemia present very high risk for stroke. PMID- 12122875 TI - [Change of fibroblast calcium levels caused by beta-amyloid peptide in Alzheimer disease]. AB - RATIONALE: beta-amyloid peptides, comprising the major neuropathological lesions of Alzheimer's disease, have been found to form depositions in various peripheral tissues, including the skin. Neurons in the disorder succumb to the altered ionic homeostasis and some other factors caused by this toxic peptide. In line with these findings, our study aimed to find differences in biochemical processes of cultured cutaneous fibroblasts derived from sporadic Alzheimer patients and from age-matched control individuals that may mirror changes in the central nervous system. METHODS: Intracellular ionic homeostasis of Alzheimer and control fibroblasts was measured in Fura-2AM-loaded human fibroblasts by dual wavelength spectrofluorimetry. RESULTS: Cells derived from Alzheimer patients exhibited lower intracellular free calcium levels as compared to the control cultures. Exposure of fibroblasts to beta-amyloid resulted in increased calcium concentrations of the control cells, but not of Alzheimer ones. CONCLUSION: Our findings indicate that Alzheimer's disease is a systemic disorder that, among others, affects the calcium homeostasis of fibroblasts. Even though it is unknown whether the diminished ionic response of Alzheimer fibroblasts is a disease or actual status marker, it could prove to be a useful model for the analysis of Alzheimer specific changes. PMID- 12122876 TI - [MR investigation of spinal cord herniation in the thoracic spine]. AB - Transdural herniation of the spinal cord is thought to be previously extremely rare and very often misdiagnosed. Possible reasons may be iatrogenic and traumatic or in about one third of cases it may be unknown, where the probable origin might be a congenital dural defect. The pathology may show characteristic and misleading MR patterns of the thoracic spine, emphasising the importance of these patterns. This anomaly can lead to progressive Brown-Sequard syndrome. Surgical intervention, consisting the repair of the dural defect may result in improvement or even complete regression of the neurologic deficits. PMID- 12122877 TI - Identification of gliomas by morphological and immunocytochemical analysis in cell cultures. AB - INTRODUCTION: The morphology and immunocytochemical properties of 250 different monolayer cultures derived from various human brain tumor specimens were investigated on purpose to support and complement the neuropathological diagnosis. In this study analyses of 124 glioma cases are presented. METHODS: The tumor samples were mechanically dissociated and seeded on glass coverslips. After the formation of the monolayer cultures were fixed and stained by May-Grunwald Giemsa method for the morphological examination. Semi-quantitative immunocytochemical labeling included several different types of mono- and polyclonal primary antibodies using avidin-biotin visualization system. In nine cases of the glioblastomas the sufficient proliferation made possible to establish cell lines from the primary cultures. RESULTS: The glial origin of the tumors was identified in 124 cases based upon the presence of glial fibrillary acidic protein. A negative correlation between the intensity of glial fibrillary acidic protein immunostaining and the grade of tumor malignancy was found. During long-term cultivation of the higher grade gliomas the incidence and intensity of glial fibrillary acidic protein labeled cells was decreasing. Both the vimentin and the neuron specific enolase labeling were in general stronger than the glial fibrillary acidic protein and almost all the cells were stained. The incidence of Ki-67 positive cells increased with the grade of malignancy. Concerning the tumor classification our immunocytochemical results correlated with the routine histopathological examination. CONCLUSIONS: On the basis of these results we conclude that monolayer cultures obtained from tumor specimens can support and complement the correct diagnosis of the various human brain tumors. PMID- 12122878 TI - [Electrophysiologic investigation of cerebral cortex in the subtypes of Parkinson disease]. AB - OBJECTIVE: Post-movement beta synchronization is an increase in EEG beta power after movement termination. Its characteristics in movement disorders are not well described. Tremor dominant Parkinson's disease shows unique clinical, anatomical and biochemical features. In our study we examined the relation between the laterality of tremor and size of post-movement beta synchronisation in tremor dominant Parkinson's disease. METHODS: In a self-paced movement paradigm we measured movement duration and analyzed EEG power changes at movement reactive beta frequencies. RESULTS: Movement duration was significantly longer in Parkinson-patients than in controls (0.49 +/- 0.170 s, 0.35 +/- 0.087 s, p = 0.013, Mann-Whitney test). There was no difference between the two hands in the control group (0.36 +/- 0.078 s, 0.34 +/- 0.099 s, p = 0.207, Wilcoxon-test), while Parkinson patients performed longer movement with their left hand (0.52 +/- 0.195 s, 0.46 +/- 0.148 s, p = 0.049, Wilcoxon), unrelated to the side of tremor. In controls, post-movement beta synchronisation contralateral to the movement was not significantly different after right and left hand movement (108.1 +/- 68.21% and 92.1 +/- 23.43%, p = 0.78 Wilcoxon). In Parkinson patients post-movement beta synchronisation was significantly smaller contralateral to the tremulous hand movement (36.9 +/- 47.79%, 104.7 +/- 91.42%, p = 0.012, Wilcoxon-test). The post movement beta synchronisation showed anterior shifting in Parkinson-patients. CONCLUSIONS: In tremor dominant Parkinson's disease the asymmetric decrease of post-move beta synchronisation is related to the laterality of tremor rather than bradykinesia. Analysis of this phenomena might provide further insight to the pathophysiology of Parkinson's disease. PMID- 12122879 TI - [Levetiracetam: a molecule with a novel mechanism of action in the pharmaceutical treatment of epilepsy]. PMID- 12122880 TI - [Congress of the American Neuro-Ophthalmology Society (NANOS)]. PMID- 12122881 TI - [Specialty meeting "The future of neurorehabilitation"]. PMID- 12122882 TI - [Rational-emotive therapy in psychogeriatrics: a new application? Case report of a patient in daycare]. AB - Literature indicates that the use of psychotherapeutic techniques in affective problems in patients with cognitive impairment is scarce and often considered to be ineffective. The feasibility of an individual treatment with Rational Emotive Therapy of a psychogeriatric patient was explored. The patient was diagnosed with moderate cognitive impairment, a depressive episode and generalised anxiety disorder. Simplification of the RET principles and integration in an interdisciplinary system were introduced as adjustments in the method. The treatment resulted in a reduction of depressive and anxiety complaints. Treatment limitations and patient variables which seemed to have contributed to the treatment success and which may have predictive value are discussed. PMID- 12122883 TI - [Psychiatric consultation in the nursing home. An inquiry among Amsterdam nursing home physicians]. AB - Psychiatric disorders are common among nursing home residents. However, little is known about psychiatric consultation in Dutch nursing homes. As an exploration of the topic, Amsterdam-based nursing home physicians were asked to rate a number of aspects of psychiatric consultation as performed in their nursing home. Striking differences are reported between 14 participating nursing homes with regard to the intensity of psychiatric consultation and the number of consultation requests, which seems low compared with the perceived psychiatric problems. Somatically ill and psychogeriatric residents are estimated to generate an equal number of consultation requests. Psychiatric consultation appears to be characterized by diagnostic clarification, medication recommendations and behavioral management advice whereas staff-directed activities are scarce. Physicians report shortcomings in psychiatric skills among care staff. Research is necessary concerning the psychiatric care delivered to nursing home residents, as well as with regard to the optimal model for psychiatric consultation services. Integration of psychiatric care in nursing homes with mental health care services appears to be desirable. PMID- 12122884 TI - [The behavior rating scale for intramural psychogeriatric inpatients (GIP-28) in homes for the elderly. A psychometric evaluation]. AB - The GIP-28 is the shortened version of the 82-item Behavior Rating Scale for Psychogeriatric Inpatients. Originally it was meant and psychometrically evaluated for use in psychogeriatric and elderly psychiatric inpatients. We supposed that the GIP-28 might be useful to detect psychosocial and cognitive problems in residents of homes for the elderly. It was therefore tested in 15 residential homes (n = 949). The instrument consists of three, factor analytically derived, scales: 'Apathy', 'Cognition' and 'Affect'. These three principal components were also identified in the data of the inhabitants of the residential homes. Internal consistency of the scales, as measured with Cronbach's Alpha is .75, .66, .80 respectively. Construct validity of the GIP-28 is satisfactory: the correlation between the Affect scale and the GDS was .36 and between the Cognition scale and the MMSE was -.36. The GIP-28 was associated with another observation scale for care needs (r = .54). The GIP-28 can be used to detect mental health problems in the population of residential homes. However, it is neither meant nor suited to replace psychiatric diagnostic procedures. PMID- 12122885 TI - [Orthostatic and postprandial hypotension in patients aged 70 years or older admitted to a medical ward]. AB - The aim of this study was to determine the prevalence of orthostatic (OH) and postprandial hypotension (PPH) in Dutch elderly patients admitted to a medical ward and to explore patient characteristics associated with OH and PPH: symptoms, medications and comorbidity. We studied 50 patients, mean age 78.8 years, 68% female. Orthostatic hypotension (OH) was defined as a decrease of systolic blood pressure (BP) > or = 20 or diastolic > or = 10 mm Hg after 3 minutes of standing. To diagnose postprandial hypotension (PPH) BP was measured sitting before and 30 minutes after the start of patient's noon meal, the same criteria were applied. We registered changes in BP, pulse rate, complaints, reason for hospitalization, medication and comorbidity: hypertension, diabetes mellitus and Parkinson(ism). We found OH in 24% of the patients, PPH was diagnosed more frequently: in 34%. 10% had both, but there were no relations between OH and PPH. In none of the patients OH and PPH were measured before our assessment. Pulse rate increased respectively 8 and 3/min. With OH and PPH. Both subjective and objective complaints were significantly associated with OH, where as only subjective non specific complaints were associated with PPH. Objective complaints were very rare in PPH. OH and PPH were not significantly associated with medication use and comorbidity. PPH and OH are common in hospitalized elderly patients. OH is more often symptomatic than PPH. We found no relationship with medication use or comorbidity. According to research literature, however, OH and PPH are associated with higher morbidity, mortality and possibly cognitive decline. Therapeutic measures must be considered, especially in the presence of comorbidity such as significant carotid artery occlusion. PMID- 12122886 TI - [Aging from a global perspective]. PMID- 12122888 TI - [Good laboratory practice: applications in the plant health sector]. AB - The principles of good laboratory practice (GLP) are also applicable to studies on plant protection products (PPP). In Italy the placing of PPP on the market is authorized by Ministry of Health in accordance with the Decree no. 194/95, which implements Directive 91/414/EEC. The studies listed in Annexes II and III of Directive 91/414/EEC must be conducted in accordance with the principles of GLP, independently of the site of conduct (laboratory, greenhouse, field). Studies on efficacy may be instead conducted in accordance with the principles of good experimental practice. The Ministry of Health grants the compliance status to test facilities which work according to the principles of BPL, with the exception of test facilities which conduct field studies as regards efficacy tests. Such test facilities are under the responsibility of the Ministry of Agriculture and Forestry. PMID- 12122889 TI - [Field studies: legal framework and implementation]. AB - The Decree DL.vo no. 194/95 prescribes compliance with correctness and quality criteria in laboratory and field studies when experimental data on pesticides are to be submitted to the regulatory authority. The Decree requires that also field tests should be conducted in compliance with good experimental practice, by facilities officially authorized by the Ministry of Agriculture and Forestry (MIPAF) and regularly inspected to verify that the prescribed requirements have been implemented and the tests are conducted appropriately. The inter-Ministries Decree DM 37529/96 defines good experimental practice criteria and the requirements for authorization of field studies for the evaluation of pesticide efficacy and the collection of samples for subsequent laboratory tests on residues. The MIPAF has established a technical-scientific Committee named "Experimental field studies" (Decrees DM of 29 January 1997 and DM 36947/97) to deal with the complex issue of field studies and to examine the application for authorization made by test facilities performing field studies. On the basis of the work done by the Committee up to 30 April 2001, the MIPAF has issued 62 certificates of compliance to test facilities conducting field studies. PMID- 12122887 TI - [The Italian inspection system for good laboratory practice: structure and updates]. AB - The Italian inspecting system for compliance with the good laboratory practice (GLP) principles is fully operative since late 1996. Over these past years documents have been issued and specific guidelines have been worked out to properly perform the activities prescribed. This preparatory work has allowed the Italian system to harmonize with those of the other member states of the European Union. In the frame of other international activities, the Italian Inspectorate has participated in the mutual joint visits programme set up by OECD. Much effort has been invested to pinpoint differences in the approaches followed by the various Italian inspecting teams when inspecting test facilities. From this standpoint, the priority was the attainment of reliable information as well as of solutions that would allow the harmonization of the conduct of GLP inspections. It cannot be overlooked, in fact, that there are 24 inspectors, to whom a substantial number of experts has to be added. PMID- 12122891 TI - The OECD pilot project of mutual joint visits: state of the art and need for further action. AB - This paper describes the ongoing pilot project to examine the organisation and procedures of the 33 national good laboratory practice (GLP) compliance monitoring programmes in OECD to determine the extent to which they comply with the recommendations of the 1989 Council Decision-Recommendation on compliance with GLP. The project is based on mutual joint visits (MJV) of peer review teams representing other monitoring authorities. At the time of this paper all 33 MJV had been completed and the reports were being reviewed by the OECD Working group on GLP. A full evaluation of the pilot project will take place in 2002, after which a policy decision will be made regarding the need to continue such a programme and in what form. PMID- 12122890 TI - [Criteria for evaluating deviations from good laboratory practice]. AB - The assessment of compliance with the principles of good laboratory practice (GLP) requires that different observations be integrated in a comprehensive assessment of the adequacy of the test facility inspected. The major types of deviations are examined in detail, since they might jeopardize the scientific quality of data. These are: a) deviations, even though seemingly minor ones, which could signal an inadequate understanding of the GLP principles (e.g., insufficient attention to personnel training); b) inadequate application of the GLP principles to basic procedures and/or structures (e.g., clean-dirty interfaces in the laboratory animal unit); c) inadequacies concerning specific study types. The elaboration of new guidelines leads to the definition of new quality requirements and critical points. One such case is the development of tests to identify endocrine effects which in turn triggers the need of structures and personnel adequate to examine seminological or hormonal parameters. PMID- 12122892 TI - [Harmonization criteria for inspection procedures. Italian Group for Quality Assurance in Research]. AB - In order to achieve higher levels of harmonisation for the application of the principles of good laboratory practice (GLP), the monitoring Authority has set up for the inspectors some guidelines to conduct inspections, as approved by the National co-ordination committee. The extension of these guidelines should allow for an improved application of the GLP principles taking into account actual situations detected during inspections carried out over the past years. In this respect a great number of observations formulated by the inspectors after inspecting several test facilities have been examined without revealing laboratories' and inspectors' identities. In addition to this, the observations made by the inspectors during the annual training course at the Istituto Superiore di Sanita in 2000 have been carefully considered. PMID- 12122893 TI - Critical aspects in the application of the principles of good laboratory practice (GLP). AB - The principles of good laboratory practice (GLP) are very flexible and an accurate interpretation is required in the application phase. Each test facility has to apply the principles of GLP within its own laboratories, taking into account both the cultural and the organisational aspects. Furthermore, the GLP quality system, as all quality systems, is a dynamic, not a static one. Continuous improvement, depending on the evolving state of the art, is essential. As a consequence, there are many difficulties as well as many different ways in applying the principles of GLP. The cultural aspects play an essential role. The knowledge, experience and fully adherence to the GLP principles, combined with the knowledge of the problems linked to the conduct of various types of non clinical laboratory studies as well as continuous discussion with representatives of regulatory authorities are mandatory in order to select the correct and adequate methods of application. PMID- 12122894 TI - [Compatibility of different quality control systems]. AB - Management of the good laboratory practice (GLP) quality system presupposes its linking to a basic recognized and approved quality system, from which it can draw on management procedures common to all quality systems, such as the ISO 9000 set of norms. A quality system organized in this way can also be integrated with other dedicated quality systems, or parts of them, to obtain principles or management procedures for specific topics. The aim of this organization is to set up a reliable, recognized quality system compatible with the principles of GLP and other quality management systems, which provides users with a simplified set of easily accessible management tools and answers. The organization of this quality system is set out in the quality assurance programme, which is actually the document in which the test facility incorporates the GLP principles into its own quality organization. PMID- 12122895 TI - [Criteria for management and use of instrumental techniques]. AB - The measurement of various parameters--be it plain temperature readings or determination of pesticides, drug monitoring or measurement of contaminants in air or water--must be carried out precisely and accurately. After purchase of the instrument, the installation qualification and the operation qualification, both produced by experts in the manufacturing company, are used to ensure correct use of the instrument. The standard operating procedures for instrument management must be sufficiently detailed to allow the operator to use the instrument correctly and, above all, must contain all the information necessary for accurate and precise calibrations. From 2002, the Food and Drug Administration will no longer accept submission of paper dossiers, so that pharmaceutical companies wishing to register a product in the USA will be obliged to submit the dossier electronically. This will involve the validation of all the software used in the management and operation of all instruments. It can be assumed that the EMEA will shortly follow the FDA, so that the validation of instrument software systems will become the rule and create new responsibilities for operators and inspectors. PMID- 12122897 TI - [The Istituto Superiore di Sanita, the organization responsible for the evaluation and accreditation of food product testing laboratories ]. AB - Laboratories in charge of food control play a pivotal role within the actions planned for the safety of food products, the latter identified as a fundamental strategic priority in the European Union Food safety white paper 2000. They are demanded to comply with specific criteria to demonstrate the use of an appropriate quality system, technical competence and production of technically reliable and valid results. Precise guidance is set forth in the Italian law DL. vo 156/97 which requires that official test laboratories should comply with European Standard UNI CEI EN 45001, currently replaced by UNI CEI EN ISO/IEC 17025. and with some of the OECD principles of good laboratory practice. The Istituto Superiore di Sanita, the Italian National Health Institute, was designated by the Ministerial Decree of 12 May 1999 as the organism responsible for evaluation and accreditation of official food product control laboratories. PMID- 12122896 TI - [Computer system validation. The quality assurance procedure]. AB - The use of computerized systems in preclinical studies has increased over the last ten years. This fact has caused dramatic changes in the way raw data are used in studies. Regulatory authorities, in fact, now require that computerized systems used during preclinical studies be validated. Quality assurance was given the task of organizing an approach to this challenge. Above all, this process applies to all company functions, from test facility management to technical staff. Moreover, various systems may be encountered during the process where the operating systems and the technologies used may be different. Another difficulty may be the lack of resources, changes requested by users or regulatory updates of legislation. Last but no least, archiving should not be underestimated. PMID- 12122898 TI - [The network of community and national reference laboratories for residues]. AB - The European Union established in the early 1990's a network of Community reference laboratories (CRL) for residues in living animals and their products. This field is regulated at present by the Council Directive 96/23/CE of 26 April 1996, adopted at the national level through the Decree DL.vo no. 336 of 4 August 1999. The four CRL are based in France, Germany, Italy and The Netherlands, respectively, each of them being responsible for different categories of residues. The CRL provides technical support to the European Commission in this matter as well as to the National reference laboratories (NRL) for residues in the member states. The four CRL bear responsibility also as regards the adoption of quality systems by NRL. In this respect, the revised principles of good laboratory practice recently issued by OECD demand that procedures be developed to inform and assist the NRL to implement them. PMID- 12122899 TI - [Chemical compound safety: typology of competency accreditation for assay centers and analytical laboratories]. AB - The use of chemicals warrants many benefits on which modern society is entirely dependent. On the other hand, the lack of reliable information about the impact of the use of chemicals raises increasing concern. In order to guarantee the safety of chemicals it is mandatory to proceed to risk assessment, which in turn consists of hazard evaluation and exposure estimation. These activities are strictly dependent upon the availability of reliable data and information, produced by, e.g., test facilities, test laboratories and clinical laboratories, the specific competence of which has been properly recognised. All this applies in the pre-marketing phase as well as during the use of chemical substances. In this latter phase it is necessary to carry out an appropriate monitoring of environment, food and, in specific situations, human beings (biological monitoring). In the field of chemical safety, standards, legal instruments and operative instruments are nowadays available. These tools make it possible to assess both the quality of data and the competence of the entities involved in the production of the data themselves. PMID- 12122900 TI - Good laboratory practice in the European Community. Role of the commission and the member states: external aspects. AB - The paper recalls the history of the development of the OECD principles of good laboratory practice (GLP) and explains why the European Community has a role to play in the area of GLP. It presents briefly the current legal framework in the European Community (Directives 87/18/EEC and 88/320/EEC) and describes the role of the Commission and the member states in the practical implementation of the GLP principles within the European Community. Impacts of GLP on the relations of the European Community with third countries, both within the framework of the OECD and through bilateral trade agreements (mutual recognition agreements, MRA) based on article 133 of the treaty establishing the European Community, are then examined in greater detail. PMID- 12122901 TI - Cutaneous and subcutaneous infections in newborns due to anaerobic bacteria. AB - This review describes the microbiology and management of the major cutaneous and subcutaneous infections in newborns where anaerobic bacteria predominate: omphalitis, necrotizing fasciitis, breast abscess, and scalp infection following intrauterine fetal monitoring. The predominant bacteria known to cause these infections are group B streptococcus, group D enterococcus, group A streptococcus, Staphylococcus aureus, Enterobacteriaceae, and anaerobic bacteria. All of these agents can colonize or infect the mother and subsequently colonize or infect the fetus or newborn either intrauterinely or during the passage through the birth canal. Infections due to anaerobes are often polymicrobial, and include also aerobic and facultative bacteria. The anaerobes recovered from these infections are Bacteroided fragilis group, Fusobacterium spp., Peptostreptococcus spp. and Clostridium spp. Early recognition and effective medical and surgical therapy are essential to recovery. Managements of these infections include surgical debridement and drainage when appropriate as well as topical and systemic use of antimicrobial agents effective against both aerobic and anaerobic bacteria. PMID- 12122902 TI - Repeated prenatal corticosteroids reduce glial fibrillary acidic protein in the ovine central nervous system. AB - INTRODUCTION: A single course of corticosteroid reduces intracranial hemorrhage in preterm infants. The mechanism of protection is unclear. Glial fibrillary acidic protein (GFAP), expressed by astrocytes, is regulated by glucosteroids and is an important component of the cells forming the blood brain barrier. We have evaluated the effect of prenatal corticosteroid upon ovine GFAP. METHODS: Date mated ewes were studied in two protocols and lambs delivered on day 125 or 145 (term = 150). In the maternal injection protocol (n = 36) ewes were administered saline, single or repeated injections of corticosteroid. In the fetal injection protocol (n = 48) direct ultrasound-guided fetal injections of saline, single or repeated corticosteroid were administered, and an additional control group did not receive fetal injections. Optic nerve GFAP immunohistochemistry was performed and quantified. RESULTS: At 125 days, repeated, but not single, administration of corticosteroid, by either maternal or fetal route, was associated with a significant reduction in GFAP (both p < 0.002); by 145 days, the deficit had recovered (both p > 0.05). The process of performing repeated fetal injections had an independent effect upon GFAP at 145 days (p = 0.002). CONCLUSION: Repeated administration of corticosteroid results in a reduction in GFAP in the developing ovine optic nerve, with recovery demonstrated by 145 days. PMID- 12122903 TI - Are intrapartum and neonatal deaths in breech delivery at term potentially avoidable?--a blinded controlled audit. AB - The aim of the study was to investigate whether deaths in term breech deliveries could have been avoided with improved care during pregnancy and delivery. All cases of intrapartum/early neonatal death of nonmalformed infants in breech presentation delivered at term in Denmark in the period 1982-92 were studied. For each of the 12 deaths two controls matched by presentation and planned mode of delivery were selected. Eleven obstetricians assessed the care through narratives that ended when the infant was delivered to umbilicus and stated if the infant died, and whether the "possible death" was potentially avoidable. The majority thought that 42% of cases and 9% of the controls had died. Antenatal and intrapartum care was suboptimal respectively in 17% and 25% of cases and 4% and 26% of controls. The assumed death was found to have been potentially avoidable in 58% of cases and 17% of controls. Care in pregnancies with IUGR, pre eclampsia, placental abruption, post-maturity, the time from decision to performance of cesarean section, and compliance between patient and professionals were more often criticized in cases than in controls. Controlled audit seemed to be a valuable tool for quality improvement and for validation of litigation activities. In conclusion, infant death at term breech delivery was to a large extent potentially avoidable. However, even in controls, suboptimal care was not uncommon. PMID- 12122904 TI - A comparison of clinical variables that predict adverse outcome in term infants with severe respiratory failure randomised to a policy of extracorporeal membrane oxygenation or to conventional neonatal intensive care. AB - OBJECTIVE: To identify clinical variables predicting adverse outcome in a group of infants with severe respiratory failure who were randomized either to referral for extra-corporeal membrane oxygenation (ECMO) or to conventional neonatal intensive care within the United Kingdom. METHODS: Adverse outcome was defined by death or disability by four years of age. Receiver operator characteristic (ROC) plots were constructed for variables with continuous data and relative risk (RR) with 95% confidence intervals (CI) calculated for binominal data. RESULTS: Of variables measurable at trial entry, congenital diaphragmatic hernia and lower birthweight was also associated with increased mortality and morbidity. Seizures or supplementary oxygen at discharge were markers of disease course, which predicted a poorer outcome amongst survivors. These variables behaved similarly in the two trial groups. Those infants in the ECMO group with an episode of sepsis, established full sucking feeds after 14 days of age or a hospital stay over 30 days were at increased risk of disability. CONCLUSIONS: This study has identified clinical variables that predict adverse outcome for term infants with severe respiratory failure. The results may assist clinicians caring for these babies, when counseling their families and in the development of guidelines for neonatal ECMO. PMID- 12122905 TI - Experience with first level ultrasound and echocardiography for a selected and an unselected population. AB - OBJECTIVE: To evaluate the difference in prevalence, distribution and prenatal detection rate of congenital heart disease (CHD) in both newborns and second trimester termination of pregnancy (TOP) in two separate time periods. PATIENTS AND METHODS: At the University Hospital of Bari, an observational study was performed, which included all cases of CHD in newborns and second trimester TOP during the periods 1998-99 and 1992-93. Prevalence, distribution and prenatal diagnosis in each group were evaluated, also selecting cases with and without known risk factors. RESULTS: Prevalence of CHD in livebirths and aborted fetuses did not change between the two periods and the same was observed for distribution of CHD spectrum. In the more recent period antenatal detection of CHD significantly increased only in TOP. Moreover, classifying all cases as low or high risk, detection rates were significantly increased in high risk cases while they were unchanged in the general population. CONCLUSION: Our data show that antenatal detection of CHD is ameliorated by concentrating expertise and good equipment on high risk cases, while it remains low in the general population. PMID- 12122906 TI - Fetal superior mesenteric artery blood flow velocimetry in normal and high-risk pregnancy. AB - THE AIM: To record blood flow velocimetry in the fetal superior mesenteric artery in normal pregnancy and to evaluate if blood flow recordings in the vessel might predict adverse outcome in high-risk pregnancy. METHODS: The fetal superior mesenteric artery blood velocimetry was recorded in a cross sectional manner in 75 normal pregnancies between 27 and 41 weeks of gestation. Reference curves were performed for pulsatility and resistance indices. The superior mesenteric artery was also located in 48 singleton pregnancies complicated by pregnancy-induced hypertension and/or intra-uterine growth retardation. Middle cerebral artery, umbilical artery and vein and uterine artery velocimetry were also recorded. RESULTS: Superior mesenteric artery PI and RI values expressed an increase in resistance to blood flow with gestational age after 32 weeks of gestation. In all except eight high-risk pregnancies the fetal mesenteric artery PI values were within normal range. Among the pregnancies with absent or reversed blood flow in the umbilical artery, all had abnormal mesenteric artery pulsatility index (PI) (> 97.5th percentiles), one fetus died intrauterine and two others died after delivery due to prematurity, growth retardation and necrotizing enterocolitis. In the remaining fetuses with increased mesenteric artery PI, necrotizing enterocolitis was diagnosed in three cases. CONCLUSIONS: Increased vascular resistance in the mesenteric artery might be a late sign of fetal circulation redistribution and frequently related to necrotizing enterocolitis in the newborn. PMID- 12122907 TI - The midwife factor in obstetric procedures and neonatal outcome. AB - AIMS: In the face of major tendency towards midwifery-led-care it was our purpose to investigate the extent of the influence of the midwife on the rates of obstetric procedures and perinatal outcome. METHODS: 5384 consecutive deliveries at the Department of Obstetrics and Gynecology, University of Graz, were enrolled in the study. The following data were collected: mode of delivery, pH of umbilical artery, Apgar score. Firstly, data were investigated for interindividual differences and, secondly, for relationship with age of the midwife as a measure of experience. RESULTS: Interindividual differences were significant for episiotomy rates (minimum: 31.6%; maximum 76.9%; p < 0.001), forceps rates (minimum: 1.7%; maximum 11.1%; p = 0.002) and pH of umbilical arteries (minimum: 7.21; maximum: 7.28; p = 0.001) but not for cesarean section rates and Apgar scores. Linear regression analysis was significant between age of midwives and pH of umbilical arteries (p < 0.001; r = 0.055) and for one-minute Apgar score (p = 0.009; r = 0.050) but not for episiotomy rates, cesarean section rates, forceps rates and five-minutes Apgar score. CONCLUSIONS: There are large interindividual differences in obstetric intervention rates which cannot be explained by the midwives' age. Provision of health care should be primarily determined by need and not by the personal characteristics of the health care provider, thus interindividual differences should be reduced and more often taken into account when analyzing any kind of data. PMID- 12122908 TI - Cerebral intracellular calcium concentrations in asphyxiated rat fetuses resuscitated with oxygen. AB - OBJECTIVE: To investigate the effects of resuscitation with three different oxygen concentrations on cerebral intra- and extra-cellular calcium, sodium and potassium changes in asphyxiated rat fetuses. METHODS: Fifty-six fetal rats of gestational age of 20 days were randomly assigned into five study groups: sham operation group (control, n = 11), room-air resuscitation group (n = 10), and 3 oxygen-resuscitated groups (n = 14, 11, and 10 respectively). Different inhaled oxygen concentrations and different timings of oxygen delivery were assigned. Except for control all fetal rats were rendered ischemic and hypoxic in utero by interrupting the placental circulation. After re-circulation, intra- and extra cellular concentrations of calcium, sodium, and potassium in the brains were measured for each individual group. RESULTS: The mean intracellular free calcium concentration of fetal rat brains was similar for the room-air resuscitation group (552.1 +/- 93.5 nmol/L) and the group resuscitated with 92.8% oxygen (520.6 +/- 79.1 nmol/L) and both were significantly higher than in the control (315.3 +/ 86.9 nmol/L) (P < 0.001). After resuscitation with 65% oxygen, be it instituted before or immediately after hypoxia, their mean intracellular free calcium concentrations in the brain cells (441.5 +/- 47.9 and 452.9 +/- 36.4 nmol/L respectively) were significantly lower than those in the room-air resuscitation (P < 0.01) and 92.8% oxygen group (P < 0.05), though still higher than in the control (P > 0.05). There was no difference in the total concentrations of calcium, sodium, or potassium among all groups. CONCLUSIONS: Resuscitation with 92.8% oxygen or room air exerted a similar effect on the parameters measured, indicating that resuscitation of asphyxiated neonates using 100% oxygen might not be superior to using room air. Resuscitation with 65% oxygen resulted in lower cerebral intracellular calcium concentrations and might produce a better outcome than using 100% oxygen or room air. PMID- 12122910 TI - Sudden intractable respiratory failure in extremely low birth weight infants with H-type tracheoesophageal fistula. AB - A fatality from a tracheoesophageal fistula (TEF) in two extremely low birth weight infants is presented. The sudden onset of intractable respiratory failure accompanied by the absence of chest movement and breathing sounds was observed. The typical clinical symptoms were concealed because the infants required mechanical ventilation and nasogastric feedings. When ventilated infants with these symptoms are suspected of the diagnosis of TEF, prompt reintubation under the guidance of a flexible bronchoscopy may be life saving because the endotracheal tube passes through the fistulas into the esophagus with ease. PMID- 12122909 TI - V-shaped deceleration differs in the pattern of carotid blood flow from variable deceleration provoked by cord compression. AB - OBJECTIVE: To investigate whether V-shaped decelerations in fetal heart rate tracing are a physiologic response to fetal movements or secondary to cord compression. STUDY DESIGN: Six pregnant sheep and their fetuses (115-125 days of gestation) were surgically instrumented and studied. Fetal electrocardiogram, carotid blood flow, arterial blood pressure and fetal movement were continuously monitored for 24 hours. Following the undisturbed 24 hour recording, these parameters were monitored during umbilical cord compression (n = 6). Differences in these parameters between V-shaped decelerations and decelerations provoked by cord compressions were examined. RESULTS: Elevation of blood pressure and decreased carotid blood flow were observed coincidentally with the initiation of V-shaped decelerations. In cord compression, elevation of both blood pressure and carotid blood flow were followed by a decreased heart rate. V-shaped decelerations exhibited a different alteration of carotid blood flow compared to decelerations caused by umbilical cord compression. CONCLUSION: V-shaped deceleration is a physiologic response secondary mainly to fetal movements and is not caused by cord compression. PMID- 12122911 TI - Neuropathological features of the brain in acardius acormus. AB - Acardia is the most severe complication in monozygotic twinning. Acardius acormus is an uncommon phenotype among acardias. We report on an acardius acormus using thorough gross and light microscopic examination of the brain. Neuropathological examination revealed a severely disorganized cerebral cortex, agenesis of cerebellum and brain stem, and undeveloped basal ganglia and thalamus. In particular, the cerebral cortex was characterized by no clear lamination, almost complete loss of neurons, extensive gliosis, angiogenesis and focal mineralization. Neurohistology of the acardius acormus clearly demonstrated two types of lesions: developmental arrest of the brain and hypoxic-asphyxic damage to the brain with reactive gliosis and angiogenesis. Both types of lesions may be induced by the same teratogen: early onset of persisting hypoxia due to a reversed arterial perfusion. The quality of the description contributes valuably to a better understanding of the basic mechanism underlying the acardius phenotype. PMID- 12122912 TI - Transient postpartum diabetes insipidus in twin pregnancy associated with HELLP syndrome. AB - Diabetes insipidus during pregnancy is an uncommon medical problem, and its cause is not entirely clear. We present a woman with twin pregnancy associated with HELLP syndrome, who developed diabetes insipidus during postpartum period. A hypertonic saline infusion study with measurement of plasma arginine vasopressin concentrations confirmed the diagnosis. She had mild response to 1-desamino-8-d arginine-vasopressin (dDAVP) during the immediate postpartum period. On the 3rd postpartum day two doses of 100 microliters of dDAVP were administered, and her urinary volume gradually decreased. We could stop dDAVP on the 30th postpartum day. This exacerbation may result from increased vasopressinase activity caused by the excessive production in the placenta due to twin pregnancy, together with the insufficient degradation in the liver due to HELLP syndrome. PMID- 12122913 TI - The rational management of fibromyalgia patients. AB - The exponential increase in pain research over the last 10 years has established fibromyalgia (FM) as a common chronic pain syndrome with similar neurophysiologic aberrations to other chronic pain states. As such, the pathogenesis is considered to involve an interaction of augmented sensory processing (central sensitization) and peripheral pain generators. The notion, that FM symptomatology results from an amplification of incoming sensory impulses, has revolutionized the contemporary understanding of this enigmatic problem and provided a more rational approach to treatment. To date, the management of FM has been mainly palliative, with the aims of reducing pain, improving sleep, maintaining function, treating psychologic distress and diminishing the impact of associated syndromes. The rapidly evolving neurophysiologic, psychophysiologic and molecular biologic basis for chronic pain states has already opened up new avenues for management which should be applicable to this difficult group of patients. Indeed, it is now possible to think about a "rational" approach to managing FM patients that was unthinkable just a few years ago. PMID- 12122914 TI - A comprehensive medical evaluation of patients with fibromyalgia syndrome. AB - Fibromyalgia syndrome (FMS) is a common and distressful condition. It is imperative that all physicians do their best to help these suffering patients with understanding and respect, since the primary responsibility of a physician is to ameliorate suffering of a patient, irrespective of the type of the disease or the illness. (The authors use the terms "disease" and "illness" synonymously, since any distinction between these two terms are really pointless because the word "disease" means lack of ease or presence of suffering.) It is clear that a physician cannot optimize management of a patient with FMS without a thorough medical and psychologic evaluation. A good evaluation helps to make a proper diagnosis, assess severity, recognize aggravating and relieving factors of symptoms, appraise psychologic factors, evaluate relevant associated or concomitant conditions, document individualized problems in a given patient, and subsequently formulate proper and individualized management. This article focuses on the major elements of a comprehensive medical evaluation, with some reference to psychologic aspects--are covered in detail in the article by Turk et al in this issue. PMID- 12122915 TI - Psychological evaluation of patients diagnosed with fibromyalgia syndrome: a comprehensive approach. AB - Symptoms of FMS are extremely distressing, and currently there is no cure or any treatment capable of substantially reducing all symptoms for all patients. Rehabilitation goals include improving emotional functioning, physical functioning, and quality of life. In light of these goals, psychological screening is an essential component of any comprehensive FMS evaluation. In many cases, the high levels of emotional distress, disability, and reduced quality of life noted in these patients warrants a more thorough psychological evaluation [11]. A comprehensive psychological evaluation is complex, involves exploration of a broad range of areas, and should be administered by an experienced health psychologist. The primary objective of this evaluation is to delineate emotional, cognitive, and behavioral factors involved in persistent pain, suffering, and disability, with an emphasis on the prescription of appropriate interventions for altering maladaptive patterns. The results of the psychological evaluation involve a synthesis of information and should assist in developing a list of behavioral problems that contribute to the maintenance and exacerbation of [table: see text] suffering and disability. Information obtained should facilitate treatment planning, specifically the matching of treatment components to the needs of individual patients. PMID- 12122916 TI - The neuropharmacology of centrally-acting analgesic medications in fibromyalgia. AB - As demonstrated above, the anatomy and neuropharmacology of the pain pathways within the CNS, even to the level of the midbrain, are extraordinarily complex. Indeed, discussions of the effects of these agents on the neuropharmacology of the thalamus, hypothalamus, and cortex were excluded from this review owing to their adding further to this complexity. Also, the dearth of data regarding FMS pain pathophysiology necessitated a relatively generic analysis of the pain pathways. As mentioned in the introduction, the current thought is that central sensitization plays an important role in FMS. However, we see in this chapter that the behavioral state of central sensitization may be a result of alterations in either the ascending systems or in one or more descending systems. Studies to assess the presence or relative importance of such changes in FMS are difficult to perform in humans, and to date there are no animal models of FMS. Accepting these limitations, it is apparent that many drugs considered to date for the treatment of FMS do target a number of appropriate sites within both the ascending and descending pain pathways. The data regarding clinical efficacy on some good candidate agents, however, is extremely preliminary. For example, it is evident from the present analysis that SNRIs, alpha 2 agonists, and NK1 antagonists may be particularly well suited to FMS, although current data supporting their use is either anecdotal or from open-label trials [114,149]. Other sites within the pain pathways have not yet been targeted. Examples of these include the use of CCKB antagonists to block on-cell activation or of nitric oxide synthetase antagonists to block the downstream mediators of NMDA activation. Efficacy of such agents may give considerable insight into the pathophysiology of FMS. Finally, as indicated previously, FMS consists of more than just chronic pain, and the question of how sleep abnormalities, depression, fatigues, and so forth tie into disordered pain processing is being researched actively. Future research focusing on how the various manifestations of FMS relate to one another undoubtedly will lead to a more rational targeting of drugs in this complex disorder. PMID- 12122917 TI - Rational and targeted pharmacologic treatment of fibromyalgia. AB - Despite disappointing results when subjected to randomized clinical trials, pharmacologic agents remain an important component of FM management. Addressing the main symptoms of pain, disturbed sleep, mood disturbances, fatigue, and associated conditions is essential to improve patient functioning and enhanced quality of life. However, much work remains to design clinical trials which address the complexity of FM, while satisfying evidence based medicine paradigms. PMID- 12122918 TI - Nonpharmacologic management strategies in fibromyalgia. AB - Clinicians using the results of the extant research base can take an optimistic view of the role of nonpharmacologic treatment strategies for fibromyalgia. There were no negative outcomes in any of the reviewed studies, although in a few studies the experimental treatment did not prove to be more effective than the attention control. Rather than viewing this negatively, one could look more closely at the attention control groups and attempt to better understand what they contained that worked as an active treatment. A number of trials include a follow-up component and all but one of them find maintenance of at least one outcome change. Maintenance of changes is more likely to occur when the patient continues to participate in the experimental activity long-term. Patients especially need strategies that help them continue in exercise regimens. Unlike cognitive skills strategies that once learned are likely to become part of a person's coping repertoire, both exercise and behavioral strategies, like progressive muscle relaxation, need to be performed on a consistent basis in order to have their effect. The goals of increased self-efficacy, symptom reduction, increased functional status and quality of life along with decreased inappropriate use of health care resources are realistic when patients persevere in their use of strategy combinations and receive support from their providers. PMID- 12122919 TI - Management of peripheral pain generators in fibromyalgia. AB - Fibromyalgia is a widespread chronic pain disorder that is characterized in part by central sensitization and increased pain response to peripheral nociceptive and non-nociceptive stimuli. Part of the comprehensive pain management of patients with fibromyalgia should include a thoughtful evaluation and search for peripheral pain generators that either are associated with fibromyalgia or are coincidentally present. The identification and treatment of these pain generators lessens the total pain burden, facilitates rehabilitation and decreases the stimuli for ongoing central sensitization. PMID- 12122920 TI - Current experience with 5-HT3 receptor antagonists in fibromyalgia. AB - Pain is perceived, transmitted, processed and modulated within an extensive network of neurotransmitters and hormones. Despite increasing knowledge about the biologic principles, even on the molecular level, the more we learn about the precise mechanisms of their interactions the more questions arise. It is also pertinent to remember that clinical scientists studying pain modulating pharmacologic agents always have to consider possible placebo effects [57-61]. Most of our knowledge regarding the function of neurotransmitter systems in the CNS has been provided by animal studies. Thus we cannot be sure that they have exactly parallel counterparts in humans. For instance, animal studies suggest an inverse relationship between brain and spinal cord concentrations of substance P. If these observations are converted to an interpretation of human fibromyalgia, low brain-tissue levels of both serotonin and substance P should be expected, while spinal cord serotonin concentrations would be low and spinal cord substance P would be high [1]. There is good evidence that 5-HT, its receptors, and their interactions with other neurotransmitters are essential for nociception and antinociception. The activities of 5-HT receptors can be studied by agonist and in humans especially by antagonist use. But even with a direct spinal application of selective agonists and antagonists, observations may still be confounded by (1) dose, as there can be a dose-dependent activation of different receptor subtypes; (2) type of nociceptive tests (e.g., thermal versus pressure versus chemical models), which may have differences in the way they are regulated; and (3) influences due to effects on temperature, blood flow or motor function. With this potential for variability, it is perhaps not surprising that there is some variability in the results of studies reporting on the effects of various 5-HT agonists and antagonists on nociceptive transmission within the spinal cord [62]. For instance, different 5-HT3 receptor densities could exist in various neuronal systems, one density type being completely inhibited at low concentrations, and the others only at higher concentrations of 5-HT3 receptor antagonists, thus resulting in contrary effects. Finally, the "endogeneous 5-HT tone" may greatly influence agonist and antagonist action. Considering this complexity of serotonin mediated reactions, it is not surprising that treatment of pain by 5-HT3 receptor antagonists appears to yield inconsistent results. As fibromyalgia is now regarded as a pain amplification syndrome with a broad variety of additional nonpain symptoms, the interrelations are complicated even more. Fibromyalgia associated symptoms (e.g., fatigue, insomnia, and irritable bowel syndrome) can be modulated by 5-HT3 receptor antagonists. From the data evaluated so far, there is evidence that 5-HT3 receptor antagonists provide significant benefit in some fibromyalgia patients. In our practice, the data justify a careful application in clinical use according to the study results. The dosage, route of application, long term adverse reactions and duration of therapy still need to be studied in greater detail. Recently reported adverse events from therapy of irritable bowel syndrome with alosetron [63-67] provide a note for caution before hastily using 5 HT3 receptor antagonists without more studies. One can surmise that, much as the biochemistry of depression has been elucidated by the development of the SSRIs, a greater understanding of the role of 5-HT3 receptor antagonists in treating fibromyalgia patients may provide some insights into disease mechanisms of this enigmatic disorder. PMID- 12122921 TI - The promise of substance P inhibitors in fibromyalgia. AB - The discovery of SP and its potent biological activities have lead to the discovery of other tachykinins and to receptors for them, including the NK1 receptor. Blockade of the NK1 receptor has a number of potentially beneficial effects in medical care including the management of drug-induced emesis and the treatment of depression. The analgesic potential of NK1 receptor antagonists that, in theory, seemed so promising has not met early expectations. However, there is still reason to predict valuable clinical uses for more potent NK1 receptor antagonists in a variety of medical conditions, including FMS. PMID- 12122922 TI - The promise of N-methyl-D-aspartate receptor antagonists in fibromyalgia. AB - There is strong evidence that intravenous administration of ketamine following a standardized protocol could be used as a diagnostic test for a central sensitization in the central nervous system in patients with FM. The combination of a weak opioid and an NMDA-receptor antagonist with few side effects is presently a promise for treatment of pain in a subgroup of patients with FM. The response to intravenously administered ketamine may help select patients for this treatment modality. PMID- 12122923 TI - Management of sleep disorders in fibromyalgia. AB - In summary, the treatment of patients with FM requires a proper assessment of the reason for the unrefreshing sleep, which is an important component of the FM syndrome. Sleep laboratory investigations provides a suitable rationale for management where a specific primary sleep disorder is determined. Nonspecific treatments include various behavioral approaches to improve sleep hygiene, fitness, and regular proper nutrition that serve to regularize disturbances in circadian sleep-wake rhythms. As yet, no medication is known to improve the EEG sleep arousal disorders that include phasic (alpha-delta), tonic alpha non-REM sleep disorders, or the periodic K alpha cycling alternating pattern disorder. Traditional hypnotic agents, while helpful in initiating and maintaining sleep and reducing daytime tiredness, do not provide restorative sleep or reduce pain. Tricyclic drugs, such as amitriptyline and cyclobenzaprine, may provide long term benefit for improving sleep but may not have a continuing benefit beyond one month for reducing pain. The use of a biologic agent that facilitates sleep related neuroendocrine functions, for example growth hormone, is reported to improve symptoms but the need for injection and high cost restrict its use. No systematic studies have been reported on the use of remedial measures for the management of PLMS/restless legs syndrome and sleep apnea that occur in some patients with FM. PMID- 12122924 TI - Treatment of fatigue in fibromyalgia. AB - Clearly, fatigue is a large and challenging problem for those suffering from fibromyalgia. It adds greatly to the morbidity and disability associated with the disease. In the management of this specific symptom in fibromyalgia, attention should first be focused on identifying comorbidities that may be present and contribute to fatigue. As with other symptoms of fibromyalgia, education is a critical component of management. This can be done by the practitioner, with available free resources, or with specialized cognitive behavioral programs. This education process can be augmented with a variety of other nonpharmacologic therapies, especially very gradually increasing, low-impact, aerobic exercise programs. Numerous pharmacologic therapies may also be helpful as an adjunct to treatment. Classes of compounds that raise central levels of norepinephrine or dopamine appear to be the most specific for management of fatigue. There are also many medications used to combat fatigue in other disorders that have not yet been adequately explored as to the possible benefits in alleviating the fatigue of fibromyalgia. Advances in the management of fatigue in fibromyalgia are likely to come from a variety of directions. Easier access to well designed nonpharmacologic therapies is essential, because these treatments are underutilized in clinical practice at present. Improvements in pharmacologic therapies will come from new insights into mechanisms, especially those that might only be present in subsets of patients and would respond to more targeted therapies. PMID- 12122925 TI - Management of dysautonomia in fibromyalgia. AB - The realization of dysautonomia in FM has opened the possibility for new and different therapeutic interventions. Much more research is needed to better define the role of ANS in the pathogenesis of FM. If this research supports current hypotheses, therapeutic trials with disciplines and substances intended to correct autonomic dysfunction will be indicated. PMID- 12122927 TI - The management of fibromyalgia-associated syndromes. AB - Most of the six million Americans with fibromyalgia have at least one associated syndrome which mandates specialized attention in addition to traditional therapeutic approaches. These include localized procedures, regional blocks, antiinflammatory or antimicrobial regimens, attention to non soft tissue sources of psychosocial distress, and classes of medicines not usually prescribed for fibromyalgia. The successful treatment of fibromyalgia-associated syndromes improves the symptoms, quality of life, and prognosis of fibromyalgia. PMID- 12122926 TI - Evaluation and management of endocrine dysfunction in fibromyalgia. AB - Fibromyalgia-like symptoms such as muscle pain and tenderness, exhaustion, reduced exercise capacity, and cold intolerance, resemble symptoms associated with endocrine dysfunction like hypothyroidism, and adrenal or growth hormone insufficiency. To investigate the potential of management of endocrine abnormalities for relieve of symptoms of patients with fibromyalgia, we reviewed experimental and clinical studies of endocrine functioning and endocrine treatment. Serum GH, androgen, and 24-hour urinary cortisol levels of patients with fibromyalgia tend to be in the lower part of the normal range, while serum levels of thyroid hormone, female sex hormones, prolactin, and melatonin are normal. With exception of GH, these conclusions are based on studies in small samples. With respect to dynamic responsiveness of the hypothalamic-pituitary adrenal (HPA) axis, the dexamethasone suppression test and stimulation with ACTH show normal results, while patients show marked ACTH hypersecretion in response to severe acute stressors, perhaps indicative of chronic CRH hyposecretion. This finding and slightly altered responsiveness of growth hormone, thyroid hormone, and prolactin in pharmacologic stimulation tests suggest a central rather than peripheral origin of endocrine deviations. Because hormone level deviations were not severe, occurred in subgroups of patients only, and few controlled clinical trials were performed, there is--unless future research shows otherwise--little support for hormone supplementation as a general therapy in the common patient with fibromyalgia. In patients with clinically overt hormone deficiency, hormonal supplementation is an option. In patients with hormone levels that are in the lower part of the normal range, interventions aimed at pain, fatigue, sleep or mood disturbance, and physical deconditioning may indirectly improve endocrine functioning. PMID- 12122928 TI - Individualizing the exercise prescription for persons with fibromyalgia. AB - "Exercise is good for you; you must exercise, and just do it" are common admonitions to fibromyalgia (FM) patients by health professionals. "I can't exercise; I hurt too much to exercise; and, I don't have enough energy to exercise" are equally common responses from patients with FM. Such exchanges can lead to frustration for both patient and provider. The factor that neither participant in the dialogue is addressing is that exercise carries both risks and benefits for persons with FM. Although for decades exercise has been acknowledged to be a key component of the treatment of FM, the majority of FM patients remain aerobically unfit, with poor muscle strength and limited flexibility. Unfit muscle is theoretically more prone to muscle microtrauma, which causes localized pain and may trigger widespread pain through disordered central processing. The purpose of this article is to provide practicing health care providers with guidelines for prescribing exercise to FM patients that take into account the risk/benefit ratio. A sample exercise prescription is included. PMID- 12122929 TI - Office management of fibromyalgia. AB - The office management of fibromyalgia (FM) is best determined by two variables: (1) the severity and complexity of each patient's symptoms, and (2) the specialization and interest of the treating physician. Because there are 6 to 10 million Americans with FM, most patient visits will be to the primary care physician. Rheumatologists, physiatrists, and other musculoskeletal specialists must work with primary care physicians to foster the early diagnosis and appropriate treatment of FM. Primary care physicians are faced with enormous challenges in caring for patients with chronic pain disorders like FM. Our managed health care system insists that patient encounters be brief. Specialty referrals are often discouraged. There is little if any reimbursement for patient education. FM treatment is labor-intensive. Therefore, optimal planning and use of precious office time and resources are most important. Rheumatologists should train our primary care colleagues to recognize FM. Many patients still go months or years before this common syndrome is diagnosed. Rheumatologists should also spearhead teaching primary care physicians the basic treatment principles of FM. If the diagnosis is made early, patients with FM in community practice do very well with simple management techniques. As consultants, rheumatologists should confirm the diagnosis of FM and suggest basic FM management. Some primary care providers or other specialists will be fully capable of bypassing this consultation, especially if the patient responds to simple management suggestions. Manpower surveys have not studied the cost-effectiveness of specialty care in FM. Rheumatologists should also assume the responsibility for the management of FM patients who have not responded to basic FM management. Additionally, some rheumatologists may wish to subspecialize in FM, a major career commitment to this perplexing disorder. These situations constitute advanced FM management. PMID- 12122930 TI - Rational management of fibromyalgia. PMID- 12122931 TI - Occupational skin diseases in nursing. PMID- 12122932 TI - Not just women's business: men in nursing. PMID- 12122933 TI - Aged care violence ignored and trivialized. PMID- 12122934 TI - Will the role of the anaesthetic nurse be lost forever? PMID- 12122936 TI - Time to change the culture of occupational violence. PMID- 12122935 TI - Glutaraldehyde registers long overdue. PMID- 12122938 TI - Funding public provision of private health--the case for a copayment contribution through the tax system. PMID- 12122937 TI - Psychiatric power and informed consent in post-World War II Canada. PMID- 12122939 TI - [Endosonographic anatomy of the ventricular system]. AB - A preclinical cadaver study was performed to test a transendoscopic sonographic probe for neurosurgery. In 25 fresh post-mortem adult human cadaver with a total of 39 endo-sonographic dissections in the ventricular system were carried out. A sonograph with an outer diameter of 6 F was used and radial sonograms were made by a real-time image technique. First results showed precise imaging, comparable to a CT in a neighbouring area of 3 cm. In this publication, the authors describe the endo-neurosonographic anatomy of the ventricular system. The sonographic probe was advanced through the working canal of a ventriculoscope, then the endoscopic and sonographic imaging were compared. Results were documented by parallel sonographic and endoscopic photo and video recordings. Based on the authors experience, it is revealed that the additional sonographic view could also be used as a navigation tool. PMID- 12122940 TI - [Treatment of migraine in patients with hypertension and ischemic heart disease]. AB - Migraine is a common disorder with a prevalence of 9-10% in Hungary. Migraine can be accompanied by hypertension and/or ischemic heart disease sometimes in younger patients, but more frequently in the elderly, which is important for therapeutical considerations. The article reviews the literature with special focus on hypertension and coronary heart disease. In the second part, the authors summarize their experiences on headache patients. PMID- 12122941 TI - [Current view of rehabilitation]. AB - The study selects some changing and developing aspects of the contemporary considerations of rehabilitation and relates them closely to neurological rehabilitation. Functional diagnostics aims at the assessment of impairments and functioning. Several internationally recognized methods of neurological rehabilitation have already been adapted in Hungary. Author emphasises the use of International classification of functioning, disability and health, officially accepted by the Assembly of the World Health Organization in 2001. Its use for functional diagnostics is also possible. The everyday practice of teamwork is still problematic. Its implementation in rehabilitation is inevitable. The neurological rehabilitation is a special field where the cooperation of many teammembers is needed. The attitude to human beings in rehabilitation is different from medicine at large. It considers man not only as biological but also as social beings with own life history, thus helping to achieve compliance and increasing autonomy. The author mentions the possibility for the cooperation with organizers of the independent living movement. The separation of quality of life and health status assessments is suggested. Attention is called to the requirements of the adaptation of assessment methods and an initial account of own experiences is given. PMID- 12122942 TI - [Chemotherapy of recurrent supratentorial malignant gliomas (phase II study)]. AB - At the Hungarian National Institute of Neurosurgery 73 recurrent supratentorial malignant tumours were treated by chemotherapy during the last ten years. Chemotherapy was applied after postoperative radiotherapy but in some cases following reoperation only. All cases were clinically and by CT or MRI verified recurrences. Forty-three patients received BCNU-DBD (dibromodulcitol) treatment (23 anaplastic astrocytoma--AA, and 20 glioblastoma multiforme--GM): day 1. BCNU 150 mg/sq.m. in i.v. infusion, day 2. dibromdulcitol 1000 mg/sq orally was given. This course was repeated every six weeks, altogether 2-8 times. Sixteen patients with AA responded with complete or partial regression but only 6 did with GM. Median survival was 14 and 7 months, the difference proved to be significant, p = 0.0091. PCV combination (procarbazine, CCNU, vincristine) was applied to 16 patients with AA and 14 cases with recurrent oligodendroglioma (O). Treatment started with vincristine 1.5 mg/sq.m. i.v. (2.0 mg maximum), the next day CCNU 100 mg/sq.m. was given, followed by procarbazine 60 mg/sq.m. on days 8-22. and finished by the same dose of vincristine on day 30. The course was repeated after one month, mostly six times. Six patients with AA did not respond; in cases of oligodendroglioma all but one responded with complete or partial improvement. It is remarkable that no significant difference was found between the survivals of BCNU-DBD or PCV treated AA patients. Chemotherapy of supratentorial malignant glioma recurrences with nitroso-ureas and their combination proved to be efficacious. It also seems, that in recurrent cases lower grade gliomas show better response rate than glioblastomas. PMID- 12122943 TI - [Neurology in the era of the Internet]. PMID- 12122944 TI - [Intracranial cholesterol granulomas]. AB - In the development of a cholesterol granuloma both cellular and vascular permeability factors have to be taken into consideration. It may arise as a special degradation product in a chronic cerebral infarct because of the partial insufficient activity of the macrophages. Consequently, the degradation of brain sphingolipids and other compounds does not follow the usual route of degradation and transportation by granular cells to the stage of neutral fat but the necrotic mass transforms into cholesterol esters. Cholesterol crystals produce an irritative effect to neighbouring tissues which may result in the formation of young fibroblasts with proliferative tendency in the vessel wall. Some of the fibroblasts take part in the proliferation of connective tissue, while the rest degenerate, producing more cholesterol or xanthomatous material. Inflammatory changes may also be associated with these lesions. The amount of cholesterol sometimes increases in the inner side of the thickening connective tissue layer. The final result may be an intracranial space occupying mass or it may end as a small cholesterol granuloma, as demonstrated in our incidental cases. By the time a granuloma has developed, the original vessel usually disappears completely, but sometimes remnants of vessels might prove the vascular origin. Other pathomechanisms should also be taken into consideration, such as a cholesterol embolus or anomalous vessel with a large cholesterol plaque in the wall. This also explains why trauma (hemorrhage, granulation), cholesterol embolus, inflammation, metabolic imbalance may predispose to the formation of a granuloma, as well as the hypercholesterolaemia. The nine cases demonstrate the significance of the intracranial granuloma from pathological, clinical and neurosurgical points of view. Such cases have not yet been reported in the national or international literature. PMID- 12122945 TI - [Pregnancy and psychiatric disorders]. PMID- 12122946 TI - [Changes in the post-ischemic glutaminic vascular reactions in newborn swine]. PMID- 12122947 TI - [Reader's letter: Surgical treatment of epilepsy]. PMID- 12122948 TI - The Japanese program of vaccination of schoolchildren against influenza: implications for control of the disease. AB - In 1970, vaccination of the schoolchildren of the town of Tecumseh, MI, against influenza was shown to protect not only the children of the town, but all of its citizens from influenza-derived illness. Subsequently, models suggested that not only illness, but hospitalizations and mortality might be reduced as well. However, influenza control programs in developed countries focused on direct vaccination of the elderly. Only in Japan was a program of schoolchildren vaccination undertaken. Measures used to gauge the effectiveness of that program were insufficiently sensitive to demonstrate value, set against the large social and healthcare gains in that country. The program was discontinued; but this discontinuation revealed that excess mortality had been dramatically reduced. The demonstration of this reduction has prompted expression of several lines of concern. In this review, I have examined these concerns and provided additional detail, bolstering the findings of the hidden success of the Japanese program. In addition, the implications of the vaccination of schoolchildren for augmented control of influenza are explored. PMID- 12122949 TI - Repeated immunization of children with inactivated and live attenuated influenza virus vaccines: safety, immunogenicity, and protective efficacy. AB - Annual immunization of healthy children with live attenuated or inactivated influenza virus vaccine currently is under consideration in an attempt to reduce influenza-associated morbidity and mortality in children and the impact of influenza in the community. However, few studies have assessed the effects of this practice. Available data from studies conducted in children and adults provide reassurance that repeated annual immunization is safe and effective. Although the frequencies of significant responses to immunization (such as significant rise in titer or evidence of vaccine virus infection) are lower among persons who were immunized previously when compared with persons immunized for the first time, no consistent increases or decreases in serum antibody levels achieved after immunization or in the level of protective efficacy have been noted. Additional data regarding the effects of annual immunization are needed, and continued efforts to develop improved vaccines for the prevention of influenza are indicated. PMID- 12122950 TI - Antiviral therapy of influenza. AB - The prevention of influenza virus infections by the use of vaccines remains the most cost-effective and practical method of influenza virus control, but the use of antiviral prophylaxis and treatment in certain populations or high-risk individuals is also possible. Four antiviral drugs are currently licensed in the United States for the treatment and/or prevention of influenza virus infection in children. The M2 blockers, (amantadine and rimantadine) have been licensed for the prophylaxis and treatment of influenza in diverse high-risk populations, including children, for years. Advantages of these agents include the low cost, high oral bioavailability, and relative tolerability of one of these agents (rimantadine) in children. Disadvantages include efficacy against influenza A viruses only (not type B), the relative rapid development of resistance, and adverse effects associated with amantadine in particular (especially in the elderly and those with decreased renal function). Two agents in a new antiviral class, the neuraminidase inhibitors, have been licensed recently for the treatment and prophylaxis of influenza in the United States. Oseltamivir is licensed for the treatment of influenza in children older than 1 year and for the prophylaxis in children older than 13 years. This drug is safe and well tolerated, and is available in capsules or a liquid suspension. Another neuraminidase inhibitor, zanamivir, is administered as an inhaled powder via a special inhaler device and is licensed for the treatment of influenza in children older than 7 years. Both neuraminidase inhibitors appear to be similarly effective and are not associated with the development of antiviral resistance. No direct comparisons of any of these antiviral agents has been performed; all result in clinical improvement approximately 1 to 2 days earlier in otherwise healthy children when therapy is initiated within 48 hours of onset of symptoms. PMID- 12122951 TI - Influenza virus vaccines in children and their impact on the incidence of otitis media. AB - Otitis media has been identified as the most frequent reason for outpatient antibiotic therapy. Several studies have linked viral respiratory infections with bacterial otitis media. In light of rising concerns about antibiotic resistance, the possibility of reducing the incidence of otitis media through vaccination against respiratory viruses has received increasing attention. This article reviews inactivated and live attenuated influenza virus vaccines and their possible impact on the incidence of otitis media. Inactivated and live attenuated influenza virus vaccines are safe and immunogenic in children older than 6 months and are linked to a decrease in the incidence of otitis media. Influenza vaccination of infants younger than 6 months has resulted in less predictable immunogenicity and deserves further investigation. PMID- 12122952 TI - Louis Pasteur: a controversial figure in a debate on scientific ethics. AB - Louis Pasteur long has been heralded as the "father of modern hygiene, public health and much of modern medicine," despite early controversies regarding his findings and methodology. A recently published biography, however, has shed new light on both Pasteur's scientific acumen and integrity. Reactions to this portrayal have been mixed. This article provides an overview of Pasteur's life and the debate regarding his scientific discoveries and honesty that has ensued for more than 100 years, with insights gained from both supporters and critics of the new biography by Gerald L. Geison. PMID- 12122953 TI - Influenza virus in pediatric patients. Introduction. PMID- 12122954 TI - Bilateral lung abscesses in an adolescent. PMID- 12122955 TI - The impact of influenza in children. AB - The highest infection rates with influenza virus occur in young children because of their lack of prior immunity and prior exposure to the virus. Most children with influenza virus infection are healthy otherwise. Previously healthy children who have influenza in their first year of life have a substantial risk for developing serious disease. An underlying chronic medical condition further increases this risk. Infants, young children, and children with underlying conditions have the highest rates of outpatient medical visits and hospitalizations for acute respiratory disease during influenza epidemics. Secondary bacterial infections, antibiotic use, and other complications of influenza are consequences of influenza virus infection in children of all ages. Annual immunization with influenza vaccine is recommended for any child older than 6 months of age in whom prevention of disease is desirable, and particularly in those with underlying medical conditions. The consequences of infection in infants younger than 6 months of age can be modified by maternal immunization currently recommended for women in the second or third trimester of pregnancy during the influenza season. Family members, including siblings, and all other close contacts should receive influenza vaccine to reduce the possibility of transmission to children at risk. Immunization of all children has a positive impact on the occurrence of influenza infection and its complications, both in those at greater risk and in the entire population. PMID- 12122956 TI - Influenza-associated encephalopathy in Japan. AB - Although the clinical entity of influenza-associated encephalopathy (influenza encephalopathy) has not gained universal recognition, it has been reported frequently as a complication of influenza in Japanese children. The influenza type A (H3N2) virus was detected in most cases. Most of the patients have been young children. Influenza encephalopathy typically is associated with a sudden onset of high fever, severe convulsions, rapidly progressive coma, and death within 2 or 3 days. Influenza encephalopathy reported in Japan is distinct from Reye syndrome, and one-fourth of patients exhibit bilateral thalamic necrosis on neuroimaging. Similar encephalopathic symptoms probably occur in North America and European countries. PMID- 12122957 TI - Laboratory diagnosis of influenza: recent advances. AB - The rapid and accurate diagnosis of influenza virus infection now is available to clinicians practicing in both outpatient and inpatient settings. Newly licensed reagents are reliable and "user friendly" and may impact care by providing an immediate diagnosis that allows appropriate antiviral therapy to be given and encourages judicious use of antibiotics. The diagnosis of influenza by viral culture also has become more mainstream, allowing health professionals to confirm diagnoses in individual patients, as well as to track the pattern of each "flu season" in the community. PMID- 12122958 TI - Safety of the trivalent, cold-adapted influenza vaccine (CAIV-T) in children. AB - The trivalent, cold-adapted influenza vaccine (CAIV-T, FluMist, Aviron, Mountain View, CA) is a live attenuated influenza virus vaccine that is administered by nasal spray. CAIV-T is efficacious in preventing influenza virus infection. The vaccine was submitted to the Food and Drug Administration for licensure in healthy children and adults. Universal immunization is being considered in children, and an effective vaccine with minimal adverse reactions is thus required. The published studies on the safety of CAIV-T in children reviewed in this article were clinical trials sponsored by the National Institutes of Health (NIH) conducted in children from 1975 to 1991, clinical trials from 1991 to 1993 sponsored by a cooperative agreement between NIH and Wyeth-Ayerst Research, and clinical trials from 1995 to the present sponsored by a cooperative agreement between NIH and Aviron. Safety assessments included the occurrence of: 1) specific influenza-like symptoms, unexpected symptoms, and use of medications within the first 10 days after vaccination; 2) acute illness and use of medication within 11 to 42 days postvaccination; 3) serious adverse events and rare events within 42 days after vaccination; 4) healthcare utilization within 14 days after vaccination; and 5) acute respiratory symptoms with annual sequential vaccine doses. CAIV-T was safe and well-tolerated. Transient, mild respiratory symptoms were observed in a minority (10%-15%) of children and primarily with the first CAIV-T dose. Vomiting and abdominal pain occurred in fewer than 2 percent of CAIV-T recipients. The gastrointestinal symptoms were mild and of short duration. An excess of illness or use of medication was not observed after the 10th day of vaccination. Sequential annual doses of CAIV-T were well-tolerated and not associated with increased reactogenicity. CAIV-T did not cause an increase in healthcare utilization. Thus CAIV-T is safe in healthy children and should complement the use of inactivated influenza vaccine, trivalent (IIV-T) in children with underlying chronic conditions. PMID- 12122959 TI - Rationale and approach to target children with asthma for annual influenza immunization. AB - Influenza virus infection can cause an acute exacerbation of asthma that may result in hospitalization. Annual influenza vaccination is recommended for children with asthma, but fewer than 10 percent receive it. This review summarizes the influenza-associated morbidity in children with asthma. The risk versus-benefit of influenza vaccination in children with asthma is evaluated. Strategies to improve influenza vaccine uptake in children with asthma are investigated. Influenza virus infection causes substantial morbidity in children with asthma. Data on safety and effectiveness of the trivalent influenza vaccine in these children are limited. However, it is generally safe and effective in preventing influenza infection. The investigational live-attenuated, cold adapted, trivalent nasal spray influenza vaccine needs evaluation for safety and efficacy in children with asthma. An annual, well-organized, computerized multicomponent strategy should be implemented for optimizing influenza immunization in the high-risk population. Children with asthma should be targeted for annual influenza immunization. PMID- 12122960 TI - Advances in diabetic gastroparesis. AB - This review addresses the advances in our understanding of the epidemiology and mechanisms in diabetic gastroparesis and dyspepsia. The mechanisms discussed include: blood glucose levels at the time of presentation, "autovagotomy," and the intrinsic innervation (particularly the interstitial cells of Cajal and nitrergic nerves). In animal models of diabetic gastroparesis, there is evidence that homeostatic mechanisms are activated in the enteric nervous system to compensate for the loss of extrinsic innervation. Understanding these advances is key to the development of novel therapeutic strategies and for making rational choices in the management of diabetic gastroparesis and dyspepsia. PMID- 12122961 TI - A clinician's perspective on chronic pancreatitis--2002. AB - The diagnosis and management of chronic pancreatitis represents a significant challenge to clinicians. Progress is being made in our understanding of this disease, especially in the area of genetics and molecular biology. Despite these advances, diagnosis of early-stage disease depends on indirect tests, and management is limited to relief of obstruction if present. This review will concentrate on mild to moderate chronic pancreatitis and explore the current state of the diagnosis and management of this disease. PMID- 12122962 TI - Gastroesophageal reflux disease: new treatments. AB - The primary therapeutic endpoint for patients with gastroesophageal reflux disease is complete relief of symptoms and improvement in quality of life. The withdrawal of cisapride has created a vacuum in the prokinetic market, with few promising drugs in the pipeline. Reflux inhibitors are being considered for clinical trials, but as of yet are unavailable. Proton pump inhibitors (PPIs) continue to be the backbone of therapy for acid suppression, providing excellent relief of symptoms and healing of erosive esophagitis. Isomeric technology with esomeprazole as the prototype represents an advance in PPI pharmacology. Antireflux surgery is now more patient-friendly, with shorter hospitalization and less major morbidity compared to open fundoplication, but surgery is at best equal to medical therapy when optimal doses of antisecretory therapy are used. Two endoscopic procedures were recently approved by the U.S. Food and Drug Administration for treatment of gastroesophageal reflux disease: radiofrequency energy delivery to the gastroesophageal junction, and transoral flexible endoscopic suturing. These techniques should be used selectively until we have more data and until results are compared to the safe and highly effective medical therapies. PMID- 12122963 TI - Imatinib mesylate. AB - Imatinib is an example of a new group of drugs being developed using the principle of molecular targeting. Imatinib is able to kill the cancer cells and not the body's healthy cells. Imatinib mesylate is indicated for the treatment of patients with Kit (CD117)-positive unresectable and/or metastatic malignant gastrointestinal stromal tumors and patients with Philadelphia chromosome positive chronic myeloid leukemia in blast crisis, accelerated phase, or in chronic phase after failure of interferon-alfa. PMID- 12122964 TI - [Demographic and clinical features of coronary artery patients in the district of Antalya and comparison of them with Turkey's averages]. AB - OBJECTIVES: Many studies have proved high plasma cholesterol and triglyceride levels as determinant major risk factors for coronary artery disease. It is also well known that coronary artery disease incidence and related mortality and morbidity is low in communities applying Mediterranean diet. Turkey, having a high incidence of coronary artery disease, is unique because of the diversity of eating habits in different regions of the country. The inhabitants of Antalya region of interest in our study, are generally kept Mediterranean diet. We thought to determine the clinical and demographic features of the coronary artery disease patients living in the district of Antalya, and to find out if they correlate with Turkey's averages when compared. We also searched for the preventive effect of Mediterranean diet, if there was any. METHODS: 516 patients, who were admitted to the department of cardiology, were investigated in terms of age, sex, smoking habits, hypertension, hyperlipidemia, diabetes, family history, angina class, usage of aspirin and nitrates. RESULTS: The results revealed that clinical and demographical features of the coronary artery disease in the district of Antalya were similar with turkey's averages and that the benefits brought by the preventive effects of Mediterranean diet, might have been comprised by smoking. PMID- 12122965 TI - The effects of amlodipine on cerebral circulatory values in patients with essential hypertension. AB - OBJECTIVE: This study assessed the effects of calcium channel blocker Amlodipine on the cerebral circulation in patients with essential hypertension. METHODS: Cerebral circulation in 37 patients with essential hypertension and 10 healthy subjects was assessed using rheoplethysmography. In patients with essential hypertension cerebral circulation values were also re-estimated after treatment with Amlodipine (5-10 mg daily). RESULTS: The cerebral circulatory parameters in hypertensive patients before treatment with Amlodipine were different from those in healthy persons. Amlodipine along with the reduction in systemic blood pressure caused attenuation of cerebrovascular abnormalities: decrease in spastic signs and increase in cerebral blood supply in patients with essential hypertension. CONCLUSION: Amlodipine causes reduction of cerebral vascular resistance and promotes improvement in arterial blood filling in patients with essential hypertension. These changes in cerebral circulation may be secondary to the increase of cardiac output--known effect of Amlodipine. PMID- 12122966 TI - [Quantitative ultrasonic myocardial texture analysis of the diabetic heart]. AB - OBJECTIVE: Contraction and relaxation of the heart cause decrease and increase in myocardial video intensity (MVI) recorded from echocardiographic images, respectively. The present study was planned to compare this physiological cyclic variations of MVI in patients with type 1 diabetes mellitus and healthy subjects. METHODS: For this purpose, standard echocardiographic examination was performed to 18 young patients (age 23.2+6.4; range: 15-37 years) with insulin dependent type 1 diabetes mellitus (diabetes duration: 7.8+5.6; range: 1-17 years) and 14 age and sex matched controls. In all subjects, end-diastolic and end-systolic 2D echocardiographic images of 3 consecutive beats that had been recorded on videotapes were digitized. The quantitative analysis of digitized imaging was performed with the help of a calibrated digitization system in order to calculate the septum and the posterior wall textural parameters. The cyclic variation index (CVI) of the mean gray level (MGL) was calculated according the formula: (MGL dias- MGL diast x 100. RESULTS: Among the groups, left ventricular diastolic dimension-index, fractional shortening, E/A ratio, and isovolumic relaxation time showed no statistically significant differences, while septum and (8.3+1.1 vs. 7.3+0.9 mm; p=0.016) and posterior wall thickness (8+0.6 vs. 6.8+1.1mm; p=0.004) and E-deceleration time (167+23 vs. 140=19 msec.; p=0.003) were significantly higher in diabetics. The diabetic patients showed significantly lower CVI both for septum (18.2+11.5% vs. 39.3+11.5%; p=0.0001) and posterior wall (16.4+16% vs. 40.5+9.2%; p=0.0001), respectively. CONCLUSIONS: Altered videoensitometric parameters possibly represent a preclinical alteration, conceivably related to the myocardial collagen content increase, which does not necessarily indicate an actual disease but may be considered an early marker of the histopathologic findings of diabetic cardiomyopathy. PMID- 12122968 TI - [The effects of sildenafil citrate on the isolated rat aorta: comparative in vitro study]. AB - OBJECTIVE: Sildenafil, an inhibitor of cGMP-specific phosphodiesterase 5 (PDE5), is currently being used as oral therapy for penile erectile dysfunction. The aim of this study was to investigate the relaxing effect of sildenafil on vascular tissue and compare it with the known vasodilatator agents, sodium nitroprusside and acetylcholine. METHOD: Rat thoracic aorta samples were cut into rings, mounted on steel hooks, and immersed in aerated Krebs solution maintained at 37 degree C. Isometric responses were recorded by strain gauge transducers connected to a polygraph. Graded relaxations were induced using increasing concentrations of acetylcholine sodium nitroprusside and sildenafil. RESULTS: The agents all does-dependently relaxed rat aorta strips. The relaxing potential of sildenafil was found to be similar to sodium nitroprusside, but higher than acetylcholine. CONCLUSIONS: In the absence of regulatory mechanisms, sildenafil citrate has noticeable vasodilatatory effect in vitro. PMID- 12122969 TI - Myocardial repolarization and drugs. Impossibility to predict the dominance of anti-arrhythmic over pro-arrhythmic effects of drugs due to differential and ventricular electrical remodeling. AB - It is known that application of anti-arrhythmic drugs for the acute treatment of arrhythmias can not only result in successful termination or prevention, but also can lead to unwanted pro-arrhythmic effects. On the basis of two arrhythmias, atrial fibrillation and Torsade de Pointes arrhythmias, we will highlight the relevance of differential atrial and ventricular electrical remodeling to explain the delicate and dynamic balance between anti-arrhythmic efficacy and pro arrhythmogenic consequences of class III anti-arrhythmic drugs. PMID- 12122970 TI - [Coronary artery bypass re-operations: basic principles]. AB - Mortality and major complications during primary coronary artery bypass operation has decreased substantially during the past 20 years. However, patients undergoing reoperative myocardial revascularization still face markedly elevated perioperative mortality and morbidity. On the other hand, the incidence of reoperative coronary bypass surgery continues to increase. Aggressive perioperative care and optimal myocardial protection is mandatory in these patients. In this article we reviewed the patient profiles, indications for operation, operative techniques and their impact on the surgical results for patients undergoing reoperative coronary artery bypass surgery. PMID- 12122971 TI - [The relation of coronary artery disease with Doppler flow velocity and resistance index in cases without significant carotid artery stenosis]. AB - OBJECTIVE: Atherosclerosis causes functional vasomotor changes as well as well as atheromatous carotid plaques and luminal stenosis resulting in mechanical effect. The relation between functional vasomotor changes in carotid arteries and extent of coronary artery disease is unknown. In our study, the probable relationship between carotid arterial flow velocities and resistance indexes (RI) with the extent of coronary artery disease (CAD) in patients who do not have significant carotid luminal stenosis was evaluated. METHODS: One hundred and fourteen patients (74 males, mean age 53+/-10 years, range 33-72 years) were studied. All patients underwent color Doppler sonography before coronary angiography. Peak systolic flow velocity, end-diastolic flow velocity and resistance index (RI) of right and left common carotid and internal carotid and internal carotid arteries were measured by color Doppler sonographic technique. Doppler parameters were correlated with the extent of CAD and left ventricular ejection fraction. RESULTS: Patients were classified on the basis of presence of significant CAD and the number of affected coronary arteries. Thirty-three patients did not have (normal group) and 81 patients had significant coronary arterial stenosis (22 patients with one-vessel disease, 27, with two-vessel disease and 32 patients with three-vessel disease). Flow velocities were the highest in normal group but the lowest in CAD patients, especially when 3 coronary arteries were affected. Correlation analysis demonstrated negative relationship of age, ejection fraction and number of affected coronary arteries with end-diastolic flow velocity, but positive and significant correlation with RI value. CONCLUSION: Our study is the first on this object. The results suggest that presence and extent of CAD changes flow velocities and RI values of common and internal carotid arteries. However, further investigations are required before these parameters can be applied as diagnostic criteria. PMID- 12122972 TI - [New and controversial indications for pacemaker implantation]. AB - New technological developments lead to the significant improvement in pacemaker's functions, which has been used for continuation of cardiac rhythm for years. Pacemaker indications have also been changed parallel with these advances and now they are used in the treatment of cardiovascular diseases. PMID- 12122973 TI - [Esteemed Associate Professor Dr. Oguz Tasdemir's correctness, sensitivity and intuition]. PMID- 12122974 TI - Readership survey. Reviews in Gastroenterological Disorders. AB - The readership survey demonstrates that Reviews in Gastroenterological Disorders has been extremely well received by the gastroenterological community. It is through readership surveys that we are able to assess our progress and identify future opportunities for improvement. We are grateful to the 191 gastroenterologists who took the time to complete the survey. The information and insights gained from the survey will help us to deliver an even better product in the future. We hope that our readership will continue providing their feedback in years to come. PMID- 12122975 TI - Nonalcoholic fatty liver disease. AB - Nonalcoholic fatty liver disease (NAFLD) encompasses a broad clinicopathologic spectrum ranging from simple steatosis to nonalcoholic steatohepatitis (NASH), which may advance to cirrhosis and end-stage liver disease. Steatosis alone does not appear to be progressive. The prevalence of NAFLD averages 20% and that of NASH, 2% to 3%, making these conditions the most common liver diseases in the United States. NAFLD is associated with insulin resistance, which may be evident clinically with obesity, type 2 diabetes mellitus, and hypertriglyceridemia. The pathogenesis of NAFLD consists of hepatic fat accumulation and oxidative stress with formation of free radicals. The clinical diagnosis is based on the presence of the insulin resistance syndrome and exclusion of alcohol abuse as well as viral, autoimmune, genetic, and drug-induced liver diseases. Liver biopsy is essential for diagnosis but may not be necessary for clinical management. Treatment is aimed at correcting the risk factors for NAFLD and using potentially hepatoprotective agents. Ursodeoxycholic acid and betaine appear particularly promising in early trials. PMID- 12122976 TI - Alendronate and risedronate: what you need to know about their upper gastrointestinal tract toxicity. AB - Adverse upper gastrointestinal (GI) tract events can occur with alendronate or risedronate therapy. Although the short-term, non-placebo-controlled comparisons of alendronate and risedronate indicated that risedronate therapy may be associated with a lower risk of upper GI toxicity than alendronate therapy, the placebo-controlled comparison shows no difference in the risk of upper GI toxicity between the two drugs. The risk of an adverse upper GI event increases when these drugs are used concurrently with nonsteroidal anti-inflammatory drug (NSAID) therapy, but this incidence is no more than that observed with concurrent placebo and NSAID therapy. Also, the risk of these adverse GI tract events can be decreased by following the dosing instructions (e.g., avoid lying down for 30 minutes after taking the drug and take the drug with a full glass of water) and may be decreased with once-weekly dosing. PMID- 12122977 TI - Update on gastrointestinal imaging. AB - The current status of three gastrointestinal imaging techniques-positron emission tomography (PET) with 18 F-fluorodeoxyglucose (FDG), computed tomographic (CT) colonography, and magnetic resonance cholangiography (MRC)--are reviewed here. FDG-PET should not be used as an initial means to identify patients with primary colorectal malignancy; for the detection of colorectal carcinoma metastases and recurrence, contrast-enhanced CT should be used to monitor patients, with the use of PET reserved for equivocal cases. CT colonography is comparable to colonoscopy for the detection of patients with colorectal polyps > or = 1 cm, and its advantages include its capability of detecting extracolonic abnormalities. MRC is a relatively new application of MR imaging, with utility in multiple clinical settings, including patients with suspected obstructive disease, choledochal cysts, primary sclerosing cholangitis or AIDS cholangiopathy, biliary-enteric anastomoses, and patients with failed or inadequate endoscopic retrograde cholangiograms. PMID- 12122978 TI - Best of the AASLD. Highlights from the 52nd Annual Meeting of the American Association for the Study of Liver Diseases. November 9-13, 2001, Dallas, TX. PMID- 12122979 TI - [Molecular markers of endothelial dysfunction in acute ischemic stroke]. AB - INTRODUCTION: In spite of all similarities, ischemic stroke cases representing 80% of the acute cerebrovascular accidents, different steps of platelet activation, coagulation and fibrinolytic cascade are involved in the pathomechanism of the different stroke subtypes. The differentiation of the atherothrombotic, cardioembolic and lacunar forms of acute ischemic stroke is based on the comprehensive evaluation of clinical signs, neuroimaging technics, and diagnostic ultrasound, but also a significant effort was made to characterize the specificities of the underlying processes of the coagulation system by signal molecules, in order to clarify their possible role and to support the diagnostic and therapeutic decisions. PATIENTS AND METHODS: The von Willebrand factor was studied as the marker of endothelial injury in 34 acute ischemic stroke patients within 24 hours after the onset of their stroke, and repeatedly 2, 4, and 12 weeks thereafter. To determine the probable source of the von Willebrand factor, usually released not only by endothelial cells, but also by platelets, the authors simultaneously measured the levels of an additional endothelial marker, thrombomodulin, and a platelet activation marker, beta-thromboglobulin. RESULTS: The mean of von Willebrand factor levels measured in stroke patients on the first day was 123%, whereas the mean of the control group 72% (p < 0.05). There was no significant difference according to stroke subtype. Von Willebrand values determined two weeks later showed a further 60% increase in stroke patients, and after a gradual fall their level remained above the concentration of the control group. The beta-thromboglobulin level measured in stroke group was significantly higher, than in control individuals (171 IU/ml vs. 32 IU/ml, p < 0.001). This was characteristic for atherothrombotic and cardioembolic stroke, but not for lacunar infarctions. If measured repeatedly, beta-thromboglobulin levels decreased rapidly in the first two weeks, than somewhat slower. Soluble thrombomodulin was slightly elevated in stroke patients (4.24 ng/ml) compared to healthy subjects (3.81 ng/ml), without statistical significance, and without major differences between subgroups. CONCLUSIONS: While early determination of beta-thromboglobulin can contribute to the differential diagnoses of the subtypes of ischemic stroke, the long-lasting elevation of von Willebrand factor may reflect endothelial dysfunction caused by several factors in the microvasculature of the penumbra. PMID- 12122981 TI - [New methods of intensive therapy in stroke: hemicraniectomy in patients with complete middle cerebral artery infarction and treatment of intracerebral and intraventricular hemorrhage with urokinase]. AB - Life-threatening, complete middle cerebral artery infarction occurs in up to 10% of all stroke patients. The "malignant media occlusion" is an infarction occupying more than 50% of middle cerebral artery territory. The malignant, space occupying supratentorial ischemic stroke is characterised by a mortality rate of up to 80%. Several reports indicate, that hemicraniectomy in this situation can be life-saving. Hemicraniectomy increases cerebral perfusion pressure and optimises retrograde perfusion via the leptomeningeal collateral vessels. A case of a patient is presented, having progressive neurological deterioration due to massive cerebral infarctions. The patient rehabilitation was successful. Decompressive surgery is life saving and can also give acceptable functional recovery. Hemorrhagic stroke is due to stroke in 15% of cases and in 10%, it is "spontaneous" intracerebral hematoma. The intracerebral and intraventricular hemorrhage represents one of the most devastating types of stroke associated with high morbidity and mortality. The 30-day mortality rate is 35% to 50% and most survivors are left with a neurological disability. The value of surgical therapy is debatable. The aspiration and urokinase therapy of the hematoma of intracerebral hemorrhage could improve final neurological outcome. Spontaneous, nontraumatic intraventricular hemorrhage frequently carries a grave prognosis. A large part of morbidity after intraventricular hemorrhage is related to intracranial hypertension from hydrocephalus. One patient presented had intracerebral hemorrhage and another had intraventricular hemorrhage treated with urokinase. Rapid and extensive reduction in the amount of intracerebral and intraventricular blood occurred. Urokinase lysis is safe and can be a potentially beneficial intervention in intracerebral and intraventricular hemorrhage. By performing decompressive craniectomy, the neurologists of stroke departments and intensive care units with the neurosurgeons will have to play major role in the management of stroke patients. PMID- 12122980 TI - [Genetics of blood coagulation in young stroke patients]. AB - BACKGROUND AND PURPOSE: The classical risk factors did not explain all the possible etiology of cerebral stroke. Genetic polymorphisms responsible for thrombophilia were implicated recently as risk factors of stroke. In this genetico-epidemiological study the author's aim was to analyse the tendency of genetic polymorphisms to cluster in a cohort of young and elderly stroke patients and in healthy subjects in Hungary. METHODS: 253 patients with stroke were compared with 173 healthy blood donors on the basis of genetic polymorphisms of platelet GP IIb/IIIa receptor (33 LeuPro), prothrombin gene G20210A, Factor V Leiden mutation, ACE I/D, methylenetetrahydrofolate reductase (MTHFR) and beta fibrinogen gene G455A. These data were acquired using PCR. Questionnaires were used to investigate the family history and to determine the risk factor profile. The subtypes of stroke were analysed in a stroke cohort grouped according to different polymorphisms. RESULTS: An increased frequency of GP IIIa heterozygosity was found as compared to a West-European stroke cohort (31% versus 19%). The prothrombin gene variant (2.9% European and 4.8% in Hungary) was also found to increase in frequency. In young stroke patients (age < 50) compared with control subjects the odds ratios were higher: in prothrombin gene (OR: 4.9), in Leiden mutation (OR: 1.67), in fibrinogen gene (OR: 1.64) and in MTHFR(+/+) (OR: 1.58). Clustering of two polymorphisms could only be detected in young patients. These clustering polymorphisms were GP IIb/IIIa with prothrombin G20210A variant (OR: 6.74, 95% CI 1.1-18.2) and prothrombin gene variant with MTHFR (OR: 5.3, CI95 1.2-8.3). CONCLUSION: Selected and clustered genetic polymorphisms of haemostatic factors could be responsible for the high stroke morbidity in Central Europe. The presence and clustering tendency of these factors have been described in young stroke victims. PMID- 12122982 TI - [Neuropsychiatric complications after stroke]. AB - Stroke represents a major public health problem in Hungary, but relatively little attention is directed toward poststroke neuropsychiatric disturbances. Stroke patients frequently represent mood disturbances, cognitive decline, anxiety disorders, and sometimes serious schizophorm or paranoid states. Poststroke depression is the most common and possibly amenable form to therapeutic intervention. Depressive symptoms have negative effect on the rehabilitation process, quality of life and even on long-term survival. Considering drug therapy, in the past decade tricyclic drugs have been replaced by newly developed antidepressants with milder side-effects profile. Our knowledge on the relationship among vascular and other types of dementia has been extended in the recent years. This development also has some therapeutic implications. It seems likely that other psychiatric disorders, psychoses, pathological affect and personality disorders also inhibit recovery and limit long-term quality of life, but available data on this topic is limited. PMID- 12122983 TI - [Issues in the care of stroke patients]. AB - All patients having had stroke or TIA require special post-hospital care, being mainly the task of general practitioners. The number of patients surviving stroke in Hungary is approximately 30,000/year. An important focus of care is secondary prevention: antithrombotic treatment and risk factors reduction. In case of residual signs of stroke, rehabilitation must also be organized and supported by the general practitioner. Medical conditions of cerebrovascular patients requiring special care demand are reviewed by the author. In this respect, some post-stroke conditions like dementia and depression require extra attention. PMID- 12122985 TI - [MR investigations in stroke]. AB - In the article digital imaging methods are presented with special emphasis on the use on diagnostics of cerebral circulation studies. Recently, fundamental changes have happened in this field, concerning especially the MR investigations. These changes have influenced the therapeutic strategies of ischaemic stroke. Authors give the theoretical background on the diffusion and perfusion MR imaging, emphasising the importance of their "mismatch" and its impact in the estimation of the outcome of ischaemic events. More recently, new, controversial facts arose, regarding the reasons of the introduction of the theory of so called "negative" and "positive" mismatches. As a consequence, a level of uncertainty took place in the judgement of prognostics. The leading institutions are searching the way to solve the problem which seems to be the quantitative evaluation of the diffusion, perfusion and mismatch data. The advent of the multislice spiral CT with very fast imaging and the importance of CT investigations increased. With this new kind of equipment, even perfusion studies can be performed using iodinated contrast medium. PMID- 12122984 TI - [Regulatory mechanisms in focal cerebral ischemia. New possibilities in neuroprotective therapy]. AB - Permanent or temporary disruption of cerebral blood flow rapidly depletes brain regions of their limited energy reserves (glycogen, glucose, oxygen, ATP) leading to an energy crisis. Tissue damage occurs due to the energy crisis. The central part of the damage, the ischaemic "core" region is surrounded by zones of the shell-like penumbra. Necrotic, as well as apoptotic cell death could be identified in the penumbra. Going away from the ischaemic core different neurochemical processes are occurring by space and time. "Immediate early response" genes (c-fos, fos-B, c-Jun, krox 20, 24) are activated, heatshock proteins (hsp 70, 72, HSF, HSE, HIF), cytokines (TNF-alpha, IL-1 beta), inflammatory factors (COX), adhesion and glial factors (ICAM-1, ELAM-1, P selectin), vasoactive factors (IL-6, -10, PAF), reactive oxigen radicals and connected factors (O2, OH, NO, NOS, SOD) are produced within minutes and hours. Cell deaths, necrosis and apoptosis due to the activation of calpains, caspases and nucleases occur in days. In parallel, growth factors and plasticity proteins (BDNF, NGF, TGF-beta, VEGF, PDGF, GAP-43) are activated as a basis of functional rehabilitation. PMID- 12122986 TI - [Impairment of vasoreactivity in brainstem and hemispheral small vessel disease: comparative study]. AB - AIMS: Cerebrovascular small vessel disease may lead to an impairment of vasoreactivity (VR). Vasoregulatory impairment in internal carotid artery distribution area has been established. In this study the authors sought the answer to the question if VR of vertebrobasilar (VB) territory was impaired in brainstem small vessel diseases and if vasoregulatory impairment differed between the two distribution territories. METHODS: VR of carotid and VB territory was compared applying different functional tests (ventilation, tilting, acetazolamide) in 25 patients with brainstem lacunar infarcts, 20 patients with periventricular leukoaraiosis and in 35 control subjects. Cerebral blood flow velocity (CBFV) of basilar artery (BA) and middle cerebral artery (MCA) was monitored with transcranial Doppler (TCD), systemic blood pressure and CO2 partial pressure of expired air were also registered. RESULTS: In the BA territory the VR was significantly smaller in the patient than in the control group (3.1 +/- 4.6 cm/sec/kPa vs. 8.2 +/- 6.2 cm/sec/kPa, p = 0.01) during hypercapnia. In a subgroup of patients with mean baseline CBFV < 25 cm/sec, the VR was significantly smaller and Pl non-significantly higher than in patients with baseline CBFV > 25 cm/s (VRCO2 1.5 +/- 2.0 cm/sec/kPa vs. 6.5 +/- 6.5 cm/sec/kPa, p = 0.007; Pl 1.11 +/- 0.30 vs. 1.0 +/- 0.26, p = 0.4) indicating higher vascular resistance in the former group. Results of tilting tests showed similar but nonsignificant changes while acetazolamide tests revealed no differences between the two groups. In the MCA territory the VR was significantly lower in patients than in the controls during hypercapnia (4.7 +/- 3.7 cm/sec/kPa vs. 18.4 +/- 6.8 cm/sec/kPa, p < 0.001) and showed a nonsignificant tendency to be lower in patients than in controls during hypocapnia (14.6 +/- 13.8 cm/sec/kPa vs. 24.7 +/- 21.2 cm/sec/kPa, p = 0.1). Although CBFV measurements during acetazolamide test tended to support these findings, they showed no significant differences between patients and controls. During head-up tilt the CBFV did not differ significantly between the two groups. The VRCO2 is significantly higher in the MCA than in the BA territory (18.4 CI95 2.98 vs. 10.1 CI95 3.01; p < 0.001). The impairment of VRCO2 was more severe in the MCA territory (VR decreased to 26% of baseline in the MCA and to 34% in the BA territory). CONCLUSION: The capacity of carotid territory VR exceeds that of VB territory. The impairment of VR is present in both the carotid and VB territories and is more severe in the former region. The most feasible test to reveal this impairment is the hypercapnic test. There is a strong correlation between the extent of vasoregulatory impairment and baseline CBFV in brainstem small vessel diseases. PMID- 12122987 TI - Auto-organisation of hybrid organic-inorganic materials prepared by sol-gel process. AB - Silica-based hybrid organic-inorganic materials prepared by sol-gel chemistry exhibit chemical and physical properties revealing their anisotropic organisation. Besides the opportunities that this phenomenon opens for the preparation of new materials, it also provides arguments to the chemist looking for a better comprehension and control of the organisation of solids. PMID- 12122988 TI - Studies of surfaces through molecular probe adsorption and solid-state NMR. AB - The interaction between the basic probe trimethylphosphine oxide and the Bronsted acid sites of a silica-alumina has been spectroscopically resolved for the first time using a new solid-state NMR approach that opens the possibilities for the investigation of surfaces. PMID- 12122989 TI - Langmuir and Langmuir-Blodgett films of a novel tryptophan peptide lipid. AB - A novel tryptophan peptide lipid, C18H35O (SA)-Gly-Trp-Gly-OH, was synthesized and studied for its surface chemistry and spectroscopic properties. PMID- 12122990 TI - Structure solution of a novel aluminium methylphosphonate using a new simulated annealing program and powder X-ray diffraction data. AB - The structure of a novel layered aluminium methylphosphonate, formula Al2(CH3PO3)3, has been solved from laboratory X-ray powder diffraction data by simulated annealing of five independent structural sub-units, revealing a combination of four- and five-fold coordinated aluminiums within the inorganic lamellae that is unique for this kind of solid. PMID- 12122991 TI - Synthesis of mixed heterocalixarenes from benzofuranyl methanols and activated indoles. AB - Mixed heterocalix[3]arenes and heterocalix[4]arenes containing indole and benzofuran rings can be synthesised via acid-catalysed reactions of benzofuranyl methanols and activated indoles: these new calixarene types are of interest as potential molecular receptors. PMID- 12122992 TI - Propargyloxycarbonyl (Poc) amino acid chlorides as efficient coupling reagents for the synthesis of 100% diastereopure peptides and resin bound tetrathiomolybdate as an effective deblocking agent for the Poc group. AB - Synthesis of short peptides using propargyloxycarbonyl amino acid chlorides as effective coupling reagents and polymer supported tetrathiomolybdate as an efficient deblocking agent are reported. PMID- 12122993 TI - Synthetic study on CP-263,114 (phomoidride B) by SET-mediated fragmentation. AB - Assembly of the highly functionalized carbocyclic core of CP-263,114 has been accomplished by using radical-mediated fragmentation with lithium naphthalenide as a key step. PMID- 12122994 TI - The oxidation of cytochrome c in nonaqueous solvents. AB - Cytochrome c was adsorbed readily and remained electroactive, with a redox potential of 32 mV, on TiO2 films placed on SnO2; similar behaviour was observed in glycerol, whereas in acetonitrile, irreversible oxidation occurred at a potential of 800 mV. PMID- 12122995 TI - Investigations into the Pd-catalysed cross-coupling of phenylacetylene with aryl chlorides: simple one-pot procedure and the effect of ZnCl2 co-catalysis. AB - [PdCl(C6H3(OPPri2)2-2,6)] 1 catalyses the coupling of electron-rich, electron neutral and electron-deficient aryl chlorides with phenylacetylene in the presence of ZnCl2 as cocatalyst to give the products in modest to excellent yields. PMID- 12122996 TI - Stereoselective one-pot preparation of 1,2,4,5-tetraalkylidene- and 1,4 dialkylidenecyclohexanes. AB - 1,2,4,5-Tetraalkylidene- and 1,4-dialkylidenecyclohexanes were prepared by the regio- and stereoselective homocoupling of skip-type diynes and enynes, and their Diels-Alder reaction to form a polycyclic compound was demonstrated. PMID- 12122997 TI - First example of regiospecific intermolecular C-H insertion reactions of cyclic rhodium carbenoids: novel synthesis of 3-indol-3'-yloxindoles. AB - Facile regiospecific intermolecular C-H insertion reactions of cyclic rhodium carbenoids have been achieved using diazo carbonyl compounds 1 and indole or N substituted indoles to afford 1,3-dihydro-1'H-[3,3']biindolyl-2-ones, 3a-o in an excellent yield. PMID- 12122998 TI - Two chain gallium fluorodiphosphates: synthesis, structure solution, and their transient presence during the hydrothermal crystallisation of a microporous gallium fluorophosphate. AB - Two novel gallium fluorodiphosphates have been isolated and their structures solved ab initio from powder X-ray diffraction data; the materials readily interconvert under hydrothermal conditions, and are metastable with respect to an open-framework zeolitic gallium fluorophosphate, during the synthesis of which they are present as transient intermediates. PMID- 12122999 TI - Albumin-controlled stereoselective reduction of 1,3-diketones to anti-diols. AB - An unprecedented combination of high chemo- and stereoselectivity in the NaBH4 reduction of 1:1 complexes between albumin and aromatic 1,3-diketones results in the formation of anti 1,3-diols with de up to 96%. PMID- 12123000 TI - Confined space and cations enhance the power of a chiral auxiliary: photochemistry of 1,2-diphenylcyclopropane derivatives. AB - Alkali ion-exchanged Y-zeolites significantly enhance asymmetric induction in the photoisomerization of a number of cis-1,2-diphenylcyclopropane derivatives containing a distant chiral auxiliary. PMID- 12123001 TI - Tandem intermolecular Suzuki coupling/intramolecular vinyl triflate-arene coupling. AB - Treatment of a benzyl substituted meso-ditriflate with boronic acids in the presence of palladium acetate, triphenylphosphine and caesium fluoride results in intermolecular Suzuki coupling followed by vinyl triflate-arene cyclisation to provide, in high yields, single regioisomers of tricyclic-carbocycles. PMID- 12123002 TI - Solvent-controlled assembling by hydrogen bridges and halogen-halogen interactions of novel organotin oxo clusters. AB - The crystal structure of the novel methylene-bridged tetraorganodistannoxane ([Ph(HO)SnCH2Sn(I)Ph]O)4 (1) depends on the solvent it is crystallised from and is controlled by hydrogen bridges and interhalogen interactions. PMID- 12123003 TI - New Pd-catalyzed selective reduction of carboxylic acids to aldehydes. AB - A catalyst generated in situ from palladium acetate and tricyclohexylphosphine efficiently catalyzes the reduction of carboxylic acids with sodium hypophosphite in the presence of pivalic anhydride to give aldehydes with high selectivity. The low cost and convenient handling of the reagents makes this process a valuable alternative to hydrogenations and metal hydride reductions. PMID- 12123004 TI - Insertion of an electron-rich alkyne into a molybdenum amido bond. AB - A pseudo-octahedral molybdenum dimethylamido complex that readily inserts 3 hexyne forming a new C-N bond has been characterized. PMID- 12123005 TI - A hydrocarbon anchored peptide that forms a chloride-selective channel in liposomes. AB - The heptapeptide sequence Gly-Gly-Gly-Pro-Gly-Gly-Gly, when anchored to diglycolic acid derived (C18H37)2NCOCH2OCH2COOH, forms chloride-selective ion channels in phospholipid liposomes but the related heptapeptide Gly-Gly-Gly-Leu Gly-Gly-Gly, and tripeptide Gly-Gly-Gly do not. PMID- 12123006 TI - The structures of Cu(I) and Ag(I) coordination polymers using the tricyanofluoroborate anion. AB - The anion BF(CN)3-forms isomorphous network polymers with Cu(I) and Ag(I) that exhibit one-dimensional channels along the b axis and demonstrate stability to air and light respectively. PMID- 12123007 TI - Extreme enantiomeric discrimination of fluoroalkanes using deuterium NMR in chiral liquid crystalline media. AB - The enantiomeric assay of fluoroalkanes using 2H-NMR in a chiral liquid crystalline medium is demonstrated, and at its limit the enantiomers of [5-(2)H] 5-fluorodecane were successfully resolved. PMID- 12123008 TI - Terpyridine Zn (II), Ru (III) and Ir (III) complexex as new asymmetric chromophores for nonlinear optics: first evidence for a shift from positive to negative value of the quadratic hyperpolarizability of a ligand carrying an electron donor substituent upon coordination to different metal centres. AB - The synthesis of 4'-(C6H4-p-NBU2)-2,2':6',2" -terpyridine and the strongly enhanced second-order NLO response of its Zn(II), Ru(III) and Ir(III)complexes are reported, evidencing for the first time a shift from positive to negative value of the ligand quadratic hyperpolarizability by varying the nature of the metal centre. . PMID- 12123009 TI - Castanospermine-trehazolin hybrids: a new family of glycomimetics with tuneable glycosidase inhibitory properties. AB - Bicyclic azasugar glycomimetics related to castanospermine and trehazolin have been prepared from sugar carbodiimides via aminooxazoline intermediates; preliminary enzyme inhibition tests showed a marked dependence of the selectivity and potency towards alpha and beta-glucosidases upon the nature of the exocyclic substituent. PMID- 12123010 TI - Productive trapping of NAD-type radicals. Non-biomimetic reduction of pyridinium salts. AB - One-electron reduction of pyridinium salts (NAD+ analogues) generates dihydropyridyl radicals which may then be engaged in radical addition processes to regioselectively form gamma-substituted dihydropyridines. PMID- 12123011 TI - Catalytic and highly regiospecific carbon-carbon bond formation at alpha-position of Michael acceptor by palladium complex. AB - Activated olefins react with allyl acetates and Bu3SnH in the presence of a catalytic amount of a palladium catalyst to afford the corresponding products which construct a new C-C bond selectively at the alpha-position of Michael acceptors. PMID- 12123012 TI - A modified molecular beacon combining the properties of TaqMan probe. AB - A modified molecular beacon that possesses a stem-hairpin structure as seen in conventional molecular beacons and can be cleaved during PCR in designed, and it can specifically recognize the presence of the target and was obviously more sensitive than conventional molecular beacons. PMID- 12123013 TI - [(eta 5-C5H5)Fe(CO)2]2B(2,4,6-Me3C6H2): synthesis, spectroscopic and structural characterization of a transition metal complex containing an unsupported bridging borylene ligand. AB - The synthesis and characterisation of the dinuclear iron complex [(eta 5 C5H5)Fe(CO)2]2B(2,4,6-Me3C6H2) containing an unsupported bridging borylene ligand are reported. PMID- 12123014 TI - High activity ethylene trimerisation catalysts based on diphosphine ligands. AB - Chromium complexes of ligands of the type Ar2PN(Me)PAr2 (Ar = ortho-methoxy substituted aryl group), on activation with MAO, are extremely active and selective catalysts for the trimerisation of ethylene. PMID- 12123015 TI - Superior performance of nano-Au supported over Co3O4 catalyst in direct N2O decomposition. AB - Au nanoparticles supported over Co3O4 lead to a highly active and stable catalyst for N2O decomposition even in the presence of excess oxygen and steam. PMID- 12123016 TI - Quinoxaline-oligopyrroles: improved pyrrole-based anion receptors. AB - Novel quinoxaline derivatives bearing dipyrromethane or tripyrromethane substituents act as improved anion receptors as compared to the unsubstituted dipyrrolylquinoxaline core from which they are derived. PMID- 12123017 TI - Toward synthetic models for high oxidation state forms of the photosystem II active site metal cluster: the first tetranuclear manganese cluster containing a [Mn4(mu-O)5]6+ core. AB - The first tetrameric high valent manganese complex consisting of a MnIV4(mu-O)5 bridged core, [Mn4(mu-O)5(dmb)4(dmbO)2](ClO4)4, [symbol: see text] was isolated via dimanganese (III,IV) and (IV,IV) intermediates in presence of the oxidant tert-butyl hydroperoxide and was characterized by X-ray crystallography, electrochemistry, infrared, UV-visible, 1H NMR, and mass spectroscopy; the structure found differs greatly from a proposal for the putative Mn4O5 aggregate found in Photosystem II. PMID- 12123018 TI - Direct comparison of the electrocatalytic oxidation of hydrogen by an enzyme and a platinum catalyst. AB - It is shown that for molecules of Allochromatium vinosum [NiFe]-hydrogenase adsorbed on a pyrolytic graphite electrode the nickel-iron active site catalyzes hydrogen oxidation at a diffusion-controlled rate matching that achieved by platinum. PMID- 12123019 TI - Selective liquid phase hydrogenation of citral on Au/Fe2O3 catalysts. AB - Gold supported on iron oxide hydrogenates citral (an alpha,beta-unsaturated aldehyde) to the corresponding alpha,beta-unsaturated alcohols (geraniol and nerol) with a selectivity higher than 95%. PMID- 12123020 TI - Diastereoselective formation of chiral iridium hydrides containing the chiral P,N chelate ligand (4S)-2-(2-(diphenylphosphino)phenyl)-4-isopropyl-1,3-oxazoline. AB - The iridium complex [Ir(mu-Cl)(PN)(PPh3)]2 (1) reacts with H2 affording only the kinetic isomer OC-6-55-C of the dihydride [IrClH2(PN)(PPh3)] (2) and with methanol yielding, also exclusively, the thermodynamic isomer OC-6-53-C (2b) of the same dihydride; complex 2b has been characterised by X-ray diffractometric methods. PMID- 12123021 TI - Revealing the photophysics of fac-[(dppz-12-NO2)Re(CO)3(4-Me2Npy)]+: a picosecond time-resolved IR study. AB - The photophysics of fac-[(dppz-12-NO2)Re(CO)3(4-Me2Npy)]+ in CH3CN have been investigated using picosecond time-resolved IR (ps-TRIR) spectroscopy, to reveal the first example of a Re(I)-dppz complex with a charge separated lowest-lying excited state. PMID- 12123022 TI - Electron-transporting and photopolymerisable liquid crystals. AB - The synthesis of a novel, photopolymerisable liquid crystal (reactive mesogen) with a high mobility of electrons in the smectic C phase at room temperature is reported for the first time as a potential charge transport layer for OLEDs. PMID- 12123023 TI - Polyethylene glycol as a non-ionic liquid solvent for polyoxometalate catalyzed aerobic oxidation. AB - The H5PV2Mo10O40 polyoxometalate in a polyethylene glycol solvent was effective for a series of aerobic oxidation reactions including oxydehydrogenation of alcohols and cyclic dienes, oxidation of sulfides and the Wacker reaction; the solvent-catalyst phase can be recovered and recycled. PMID- 12123024 TI - TS-1 zeolite microengineered reactors for 1-pentene epoxidation. AB - A zeolite-based microengineered reactor was fabricated and tested for 1-pentene epoxidation over titanium silicalite-1 (TS-1) catalyst, which has been selectively incorporated within the microreactor channel using a new synthesis procedure. PMID- 12123025 TI - Differential evolution: crystal structure determination of a triclinic polymorph of adipamide from powder diffraction data. AB - The crystal structure of a previously unknown triclinic polymorph of adipamide has been solved from laboratory X-ray powder diffraction data using a new direct space global optimisation method based on differential evolution. PMID- 12123026 TI - Novel photo-induced coupling reaction of 9-fluorenylidenemalononitrile with 10 methyl-9,10-dihydroacridine. AB - 9-Fluorenylidenemalononitrile reacts with 10-methyl-9,10-dihydroacridine in deaerated acetonitrile under irradiation with lambda > 320 nm to give a coupling product 9-dicyanomethyl-9-(10'-methyl-9'-acridinyl)fluorene, characterized by X ray crystallographic, MS and NMR analyses. PMID- 12123027 TI - Novel family of low molecular weight hydrogelators based on L-lysine derivatives. AB - New L-lysine derivatives with a positively charged terminal can gel water below 1 wt%; particularly, 1a and 2a form a hydrogel at 0.3 wt% corresponding to approximately 12,300 and 12,500 waters/gelator molecule, respectively. PMID- 12123028 TI - Asymmetric epoxidation using a singly-bound supported Katsuki-type (salen)Mn complex. AB - We have successfully prepared an unsymmetrical analogue of a Katsuki-type salen ligand having a single hydroxyalkyl group at its 6-position, and also its Mn(III) complex; attachment of the complex to a polymer gives a highly enantioselective and recoverable catalyst for epoxidation of 1,2-dihydronaphthalene. PMID- 12123029 TI - Large electro-optic activity and low optical loss derived from a highly fluorinated dendritic nonlinear optical chromophore. AB - A 3-D shape nonlinear optical chromophore encapsulated by highly-fluorinated dendrons exhibits significantly improved electro-optic properties and optical attenuation. PMID- 12123031 TI - Electrodeposition of stable and narrowly dispersed germanium nanoclusters from an ionic liquid. AB - Germanium nanoclusters with a narrow height distribution have been electrodeposited from a dilute solution of GeCl4 in the ionic liquid 1-butyl-3 methylimidazolium hexafluorophosphate [BMIm]PF6: under the reported conditions the lateral sizes of most of the clusters range between 20 and 30 nm while their heights vary from 1 to 10 nm with most of them between 1 and 5 nm. PMID- 12123030 TI - Highly emissive, nine-coordinate enantiopure lanthanide complexes incorporating tetraazatriphenylenes as probes for DNA. AB - The interaction of q = 0 delta- and lambda-Tb and Eu complexes with poly(dAdT), poly(dGdC) and calf-thymus DNA has been examined by absorption, emission and chiroptical spectroscopy and is sensitive to complex helicity, base-pair type and the nature of the lanthanide excited state. PMID- 12123032 TI - Organometallic chemistry of carbaporphyrinoids: synthesis and characterization of nickel(II) and palladium(II) azuliporphyrins. AB - Brief treatment of azuliporphyrins 5 with nickel(II) or palladium(II) acetate in refluxing DMF afforded excellent yields of the related chelates; these novel organometallic compounds retain cross-conjugated borderline aromatic structures as judged by UV-Vis and NMR spectroscopy, and X-ray crystallography. PMID- 12123033 TI - The first structurally characterized cationic lanthanide-alkyl complexes. AB - Reaction of rare earth metal-alkyl complexes [Ln(CH2SiMe3)3(THF)2](Ln = Y, Lu) with B(C6X5)3(X = H, F) in the presence of crown ethers gives crystallographically characterized ion pairs [Ln(CH2SiMe3)2(CE)(THF)n]+[B(CH2SiMe3)(C6X5)3]-(CE = [12]-crown-4, n = 1; CE = [15]-crown-5 and [18]-crown-6, n = 0). PMID- 12123034 TI - Steric effects on diffusion of associated molecules in acetone. AB - The effects due to steric hindrance on solute-solvent interactions and on diffusion of associated molecules were found by comparing the diffusion coefficients of different aromatic isomers in acetone at 298.2 K; there exists a correlation between the isomeric effects of intermolecular association on diffusion and the molecular scales of overall hydrogen-bond acidity of the isomers studied. PMID- 12123035 TI - Tethered trinuclear cyclometalated platinum(II) complexes: from crystal engineering to tunable emission energy. AB - The crystal structures and photoluminescence of trinuclear cyclometalated platinum(II) complexes are dependent on the tethering phosphine ligands and sensitive to metal-metal contacts and pi-pi stacking interactions. PMID- 12123036 TI - Vanadium nitride functionalization and denitrogenation by carbon disulfide and dioxide. AB - A dramatic difference in behavior is observed for the dithiocarbamate and carbamate complexes [Ar(But)N]3V(NCE2)Na(THF)2(E = S or O, respectively), prepared from the corresponding nitride species ([Ar(But)N]3V identical to NNa)2 by way of a nucleophilic addition reaction involving carbon disulfide or dioxide, and is rationalized with the aid of DFT calculations. PMID- 12123037 TI - Efficient amination of sulfides with a ketomalonate-derived oxaziridine: application to [2,3]-sigmatropic rearrangements of allylic sulfimides. AB - A novel N-Boc-oxaziridine is reported, derived from diethyl ketomalonate, which effects efficient amination of sulfides to sulfimides. The reagent is applied to the [2,3]-sigmatropic rearrangement of allylic sulfimides. PMID- 12123038 TI - Continuous hydroamination in a liquid-liquid two-phase system. AB - The direct addition of NH across a CC multiple bond (hydroamination) was efficiently catalysed in a liquid-liquid two-phase system. The latter comprised a polar catalyst phase of Zn(CF3SO3)2 in the ionic liquid 1-ethyl-3 methylimidazolium trifuoromethanesulfonate and a substrate mixture in heptane. The possibility of catalysing different hydroamination reactions continuously was demonstrated. PMID- 12123039 TI - Conformation change of the cyclohexanedicarboxylate ligand toward 2D and 3D La(III)-organic coordination networks. AB - 2D and 3D structures of La-1,4-cyclohexanedicarboxylates (chdc) coordination networks, have been constructed using flexible chdc ligands which can have cis and trans conformations of two dicarboxylate groups, controlled by the solution pH in hydrothermal reaction conditions. PMID- 12123040 TI - Formation of A2B2 supramolecular porphyrin co-polymers. AB - New supramolecular A2B2 co-polymers formed in solution from a rigid diporphyrin monomer (the A2 unit) and a short flexible dipyridine monomer (the B2 unit) are reported; NMR experiments show that complete binding occurs at mM concentrations; UV titrations reveal that the dipyridine unit and a monomeric control ligand have identical binding constants, confirming that linear polymers were generated (in preference to small cyclic oligomers); at 2 x 10(-2) M polymers with an average molecular weight of 17,100 g mol-1 and containing approximately 14 porphyrin units have formed. PMID- 12123041 TI - Synthesis and characterisation of the new oxyselenide Bi2YO4Cu2Se2. AB - In this communication we report the synthesis, structure and preliminary characterisation of Bi2LnO4Cu2Se2 (Ln = Y, Gd, Sm, Nd, La) phases; these materials are members of a new family of layered oxychalcogenides. PMID- 12123043 TI - Microwave single walled carbon nanotubes purification. AB - A new single walled carbon nanotubes (SWCNTs) purification procedure has been developed; it consists in a combination of air treatment and acid microwave digestion leading to a high purity SWCNTs material; the procedure reaches high metal removal percentages and the operation time is drastically reduced compared to conventional acid reflux treatments. PMID- 12123042 TI - The rational design of novel chiral oxovanadium(IV) complexes for highly enantioselective oxidative coupling of 2-naphthols. AB - Several novel chiral oxovanadium(IV) complexes have been designed and prepared for the asymmetric catalytic oxidative coupling of 2-naphthols with high enantioselectivities of 83-98% ee. PMID- 12123044 TI - Formation of the first monoanion and dianion of stannole. AB - The first syntheses of mono- and dianions of stannole were accomplished by transmetallation or reduction of the novel bi(1,1-stannole). PMID- 12123045 TI - Alkyne coupling in rhenacarbaborane chemistry. Structure of [1,2-mu-NHBut-2,2 (CO)(2)-3,2-sigma:eta 2-(C(=CHBut)-CH=CHBut)-closo-2,1-ReCB10H9]. AB - The butadienyl moiety in the title compound is bound to both cage-boron and rhenium vertices, and arises from coupling of two alkyne molecules at the rhenium centre, unprecedented in metallacarbaborane chemistry. PMID- 12123046 TI - Influence of the phase composition of titania on catalytic behavior of Co/TiO2 for the dry reforming of methane. AB - The phase transfer of TiO2 from anatase to rutile for a 10 wt% Co/TiO2 catalyst during the reduction causes serious disappearance of activity at 0.1 MPa during CH4/CO2 reforming, whereas the transfer for 0.5 wt% Co/TiO2 brings about relatively stable activity at 2 MPa. PMID- 12123047 TI - Confined supramolecular nanostructures of mesogen-bearing amphiphiles. AB - Stable surface nanostructures with different morphology have been successfully constructed by modifying the chemical structure of synthetic amphiphiles; by introducing mesogenic groups into bolaform amphiphiles, stable spaghetti-like or stripe-like nanostructures can be obtained; it is believed that such a kind of surface structure could be used for templating synthesis and assembly. PMID- 12123048 TI - Plugged hexagonal templated silica: a unique micro- and mesoporous composite material with internal silica nanocapsules. AB - We describe in this paper the development of plugged hexagonal templated silicas (PHTS) which are hexagonally ordered materials, with internal microporous silica nanocapsules; they have a combined micro- and mesoporosity and a tuneable amount of both open and encapsulated mesopores and are much more stable than other tested micellar templated structures. PMID- 12123049 TI - Stepwise, ring-closure synthesis and characterization of a homoleptic palladium(II)-pyrazolato cyclic trimer. AB - A triangular homoleptic complex, [Pd(mu-3-Ph-pz)2]3, is prepared by the stepwise formation of an open-chain trimer and subsequent ring-closure; D3 molecular symmetry results from the one-directional arrangement of the unsymmetrically substituted pyrazolato ligands; the palladium atoms are held in close proximity, with Pd-Pd distances of 2.997(1)-3.087(1) A. PMID- 12123050 TI - Direct synthesis of alkynyl(phenyl)iodonium salts from alk-1-ynes. AB - Alkynyliodonium salts can be directly prepared from alk-1-ynes by the reaction with iodosobenzene, tetrafluoroboric acid, and a catalytic amount of HgO. PMID- 12123051 TI - [Cr(Salen)] as a 'bridge' between asymmetric catalysis, Lewis acids and redox processes. AB - An overview focused on the synthesis, structural features and catalytic applications of chiral [Cr(Salen)] complexes is presented. Key aspects of modern organic chemistry such as Lewis acids, asymmetric catalysis and redox processes are strictly connected with chronium-Schiff base complexes. Among the asymmetric transformations mediated by [Cr(Salen)] complexes, Diels-Alder and hetero-Diels Alder reactions, ARO of epoxides, kinetic resolution of meso-epoxides and Nozki Hyama reactions are taken into account. PMID- 12123052 TI - Fluorinated NAD as an affinity surfactant. AB - Nicatinamide adenine dinucleotide (NAD) with an attached perfluoropolyether tail acts as an affinity surfactant in the extraction of the enzyme horse liver alcohol dehydrogenase (HLADH) from an aqueous medium into a fluorous solvent. PMID- 12123053 TI - The first self-assembled trimetallic lanthanide helicate: different coordination sites in symmetrical molecular architectures. AB - A tris-tridentate segmental ligand has been designed for the self-assembly of homotrimetallic triple-stranded lanthanide helicates possessing different coordination sites along the threefold axis. PMID- 12123054 TI - Synthesis, crystal structure and properties of thiaheterohelicenes containing phenolic hydroxy functions. AB - A convenient method for preparation of dimethoxythiahelicene 4 and dihydroxythiahelicene 5 is described; these helical molecules showed self assembling properties such as enantiomorphic crystal growth and clathrate formation. PMID- 12123055 TI - Synthesis of novel polyurethane polyesters using the enzyme Candida antarctica lipase B. AB - A novel enzymatic route has been used to synthesise standard and unusual polyester polyurethanes without employing the usual highly toxic isocyanate intermediates. PMID- 12123056 TI - Cu(2+)-induced formation of cage-like compounds containing pyrazole macrocycles. AB - The crystal structure of the complex [Cu4(H-2L)2(H2O)2(ClO4)2](ClO4)(2).2H2O where L is a new pyrazole ligand containing 1,5-diaminopentane spacers represents a new form of obtaining metal ion-induced inorganic-organic cages. PMID- 12123057 TI - Simultaneous selection, amplification and isolation of a pseudo-peptide receptor by an immobilised N-methyl ammonium ion template. AB - Immobilised N-methyl ammonium ions have been used to simultaneously select, amplify and isolate a pseudo-peptide receptor from a dynamic library of hydrazones. PMID- 12123058 TI - One pot synthesis of nitrogen-containing polycyclic delta-lactones by double nucleophilic addition of bis(trimethylsilyl)ketene acetals to pyridines. AB - Pyridine and bis(TMS)ketene acetals (TMS = trimethylsilyl) react successively with methylchloroformate and iodine (or peracids) to give, via functionalized dihydropyridines, bicyclic nitrogen-containing lactones which have been characterized by X-ray crystallography. PMID- 12123059 TI - Nickel-catalyzed regioselective carbocyclization of ortho-halophenyl ketones with propiolates: an efficient route to disubstituted indenols. AB - Carbocylization of o-halophenyl ketones with propiolates in the presence of Ni(dppe)Br2 and Zn powder in acetonitrile at 80 degrees C afforded the corresponding 2,3-disubstituted indenols. PMID- 12123060 TI - Allosteric effects in norbadione A. A clue for the accumulation process of 137Cs in mushrooms? AB - A fruitful combination of potentiometry, absorption spectrophotometry, ESMS and 1H NMR enabled the characterisation of two caesium complexes with norbadione A and the determination of the respective stability constants of a mononuclear and a dinuclear caesium complex at pH approximately 6; a preliminary study allowed the assignment of five protonation sites of this pigment; a positively cooperative binding of the second Cs+ cation was observed. PMID- 12123061 TI - 18O incorporation in the oxidation of N-methylcarbazole by lignin peroxidase and a model compound: a mechanistic insight into the oxidative N-demethylation of aromatic tertiary amines. AB - Using 18O labelled reactants and/or solvent, the origin of the oxygen in the products of the oxidation of N-methylcarbazole by H2O2 catalysed by lignin peroxidase and a model compound has been determined, so getting important information about the mechanism of the oxidative N-demethylation of aromatic tertiary amines. PMID- 12123062 TI - Selective catalytic reduction of NOx with propene over CeO2-ferrierite. AB - CeO2-HFerrierite catalysts, prepared by physically mixing the components, showed very high conversions of NO under dry and wet conditions and excellent regeneration properties of the coked catalyst in the presence of water. PMID- 12123063 TI - Photo-induced ligand substitution at a remote site via electron transfer in a porphyrin-appended rhenium carbonyl supermolecule. AB - The photochemical and electrochemical properties of a Zn-porphyrin appended rhenium(I) tricarbonyl bipyridine 3-Me-pyridine complex have been investigated; visible-light sensitisation of electron transfer results in ligand substitution at a site remote from the chromophore. PMID- 12123064 TI - 2,4,6-Tris(4-nitrophenoxy)-1,3,5-triazine: a hexagonal host framework stabilised by the NO2-trimer supramolecular synthon. AB - The title molecule forms a honeycomb network of molecular and nitro-trimer synthons with guest species included in the hexagonal voids. PMID- 12123065 TI - New carbon nanofiber/graphite felt composite for use as a catalyst support for hydrazine catalytic decomposition. AB - Graphite felt supporting 40 nm diameter carbon nanofibers was synthesized and successfully used as a support for a high loaded iridium catalyst (30 wt%) in the decomposition of hydrazine; a strong mechanical resistance and a high thermal conductivity led to a very efficient and stable catalyst as compared to that used industrially, iridium supported on a high surface area alumina. PMID- 12123066 TI - Supramolecular self-assembly of a fluorinated Zn porphyrin. Molecular structure of a two-dimensional network of amine-functionalized, hexacoordinated Zn porphyrins. AB - [5,10,15,20-Tetrakis(4-n-propylamino-2,3,5,6- tetrafluorophenyl)porphyrinato]zinc(II) self-assembles into a two-dimensional supramolecular array in which each porphyrin is linked to four orthogonally oriented chromophores. PMID- 12123067 TI - Binuclear iron(II) complex from a linked-bis(amidinate) ligand: synthesis and its reaction with carbon monoxide. AB - Binuclear iron(II) complexes supported by a cyclohexane-linked bis(amidinate) ligand have been isolated and structurally characterized. PMID- 12123068 TI - Zeolite microtunnels and microchunnels. AB - Using a new fabrication strategy, novel, self-enclosed microporous silicalite-1 zeolite microtunnel and microchunnel architectures were successfully prepared. PMID- 12123069 TI - Self-networking of carbon nanotubes. AB - Carbon nanotube self-assembly into honeycomb-networks via controlling the ratio of the catalyst over hydrocarbon in the vapor phase using a tunable chemical vapor deposition process. PMID- 12123070 TI - The first example of palladium catalysed non-perfectly alternating copolymerisation of ethene and carbon monoxide. AB - Non-alternating ethene-CO copolymerisation is catalysed by a new series of [P O]Pd catalysts based on o-alkoxy derivatives of diphenylphosphinobenzene sulfonic acid. PMID- 12123071 TI - Macrodiscotic triphenylenophthalocyanines. AB - The first heavily substituted triphenylenophthalocyanines have been synthesised and found to be mesogenic. PMID- 12123072 TI - The Baylis-Hillman reaction in supercritical carbon dioxide: enhanced reaction rates, unprecedented ether formation, and a novel phase-dependent 3-component coupling. AB - The Baylis-Hillman reaction can be efficiently carried out in scCO2 with enhanced reaction rates relative to comparable solution phase reactions; at low pressures, a novel dimerisation is observed which has led to the development of a novel one pot three component coupling reaction to form highly functionalized ethers. PMID- 12123073 TI - The contribution of complementary hydrogen bonding to supramolecular structure. AB - Two different types of complementary hydrogen bonding contribute to the formation and temperature dependent behavior of a pseudo[2]rotaxane dimer. PMID- 12123074 TI - A palladium catalysed cyclisation-carbonylation of bromodienes: control in carbonylation over facile beta-hydride elimination. AB - Conditions have been found for the efficient palladium mediated cyclisation carbonylation of bromodienes to give gamma,lambda-unsaturated acids. PMID- 12123075 TI - The in situ oxidation-Wittig reaction of alpha-hydroxyketones. AB - In situ oxidation-Wittig methodology has been applied to alpha-hydroxyketones avoiding the problems normally associated with alpha-ketoaldehydes; the procedure is practically simple, and in many cases gives high yields of the product gamma ketocrotonates; the procedure has also been successfully utilised with ethyl glycolate and glycolaldehyde dimer. PMID- 12123076 TI - Polyoxometalates as pH-sensitive probes in self-assembled multilayers. AB - The polyoxometalate cluster [CoII4(H2O)2P4W30O112]16- (Co-POM) embedded in a self assembled polyelectrolyte matrix shows a remarkable pH dependence of its electrochemical response, opening a route to use Co-POM as a molecular probe or to fabricate pH microelectrodes. PMID- 12123077 TI - A highly enantioselective synthesis of cyclic alpha-amino acids involving a one pot, single catalyst, tandem hydrogenation-hydroformylation sequence. AB - Tandem enantioselective hydrogenation followed by a hydroformylation-cyclisation sequence leading to cyclic alpha-amino acids with ee's > 95% can be achieved in a single pot, one catalyst system by successive reactions of prochiral dienamide esters with H2 followed by H2/CO using Rh(I)-DuPHOS. PMID- 12123078 TI - A novel piezo-optical styrene sensor incorporating polymer-supported tribromide ion. AB - Encapsulation of Amberlyst A-26 supported tribromide in a 10,000 MW polyethylene glycol matrix gives a robust colour-sensitive reagent matrix which can be deposited on indium-tin oxide coated polyvinylidene fluoride (PVDF) piezoelectric film for the detection of styrene (and other alkene) vapours. PMID- 12123079 TI - Ruthenium(II) complexes in polymerised bicontinuous microemulsions. AB - Novel polymerised bicontinuous microemulsions can provide unique microenvironments for some functional molecules of scientific interests and practical applications. PMID- 12123080 TI - The formation of a cyclic diacetal of methyl alpha-D-mannopyranoside with a 16 membered macrocyclic loop. AB - The X-ray structure of a diacetal with a 16-membered macrocyclic loop, which was obtained as a product of the condensation of methyl alpha-D-mannopyranoside and 1,4-bis(2-formylphenoxy)butane is presented together with polymeric compounds resulting from polycondensation; a similar polymer was formed in the reaction of methyl 2,3:4,6-di-O-salicylidene-alpha-D-mannopyranoside with 1,4-dibromobutane. PMID- 12123081 TI - Chiral induction controlled by aggregation of organometallics: trifluoromethylated aminoalcohols for chiral ligands in Et2Zn alkylation of benzaldehyde. AB - Chiral induction in the reaction of Et2Zn with PhCHO by fluorinated chiral amino alcohols was correlated to the extent of aggregation of Et2Zn species, which was estimated by manometric study. PMID- 12123082 TI - Palladium dimethylsilanedithiolato complex: a precursor for Ti-Pd and Ti-Pd2 heterometallic complexes. AB - The silanedithiolato complex Pd(S2SiMe2)(PEt3)2 1 reacted with (C5H5)TiCl3 and TiCl4(thf)2 to produce the heterometallic clusters (C5H5)TiCl(mu-S)2Pd(PEt3)2 2 and TiCl2(S)(mu-S)2Pd2(PEt3)4 3 along with silicon-sulfur bond cleavage, respectively. PMID- 12123083 TI - A superoxide dismutase-modified electrode that detects superoxide ion. AB - A superoxide dismutase (SOD)-modified electrode, in which SOD is oriented on the gold electrode via a self-assembled monolayer of cysteine so as to allow its direct electrode reaction, possesses a bi-directional electrocatalysis for both the oxidation of superoxide ion (O2-) to O2 and the reduction of O2- to H2O2 and functions as a third generation O2- biosensor. PMID- 12123084 TI - Biocatalysis in ionic liquids: batchwise and continuous flow processes using supercritical carbon dioxide as the mobile phase. AB - A new and environmentally benign protocol for enzymatic reactions in ionic liquids is described using supercritical CO2 as the mobile phase; the products are obtained in solvent-free form and the enzyme/ionic liquid mixture can be recycled in batchwise or continuous flow operations. PMID- 12123085 TI - Electrophilic aromatic nitration using perfluorinated rare earth metal salts in fluorous phase. AB - In this paper, we describe a practical, useful electrophilic aromatic nitration process in fluorous phase by using perfluorodecalin (C10F18, cis- and trans mixture) as a fluorous solvent and perfluorinated rare earth metal salt [Yb(OSO2C8F17)3] as a catalyst for the electrophilic aromatic nitration. PMID- 12123086 TI - CeO2-promoted W-Mn/SiO2 catalysts for conversion of methane to C3-C4 hydrocarbons at elevated pressure. AB - A CeO2-doped Na-Mn-W/SiO2 catalyst for oxidative conversion of methane has been studied in a micro-stainless-steel reactor under elevated pressure; a CH4 conversion of 47.2% with a C3-C4 selectivity of 47.3% (C2:C3:C4 = 1:1:3.3) was obtained at 983 K with 1.0 x 10(5) ml g-1 h-1 GHSV, CH4/O2 = 2.5 and P = 0.6 MPa. PMID- 12123087 TI - Morphology controlled growth of arrays of GaN nanopillars and randomly distributed GaN nanowires on sapphire using (N3)2Ga[(CH2)3NMe2] as a single molecule precursor. AB - Controlled growth of oriented GaN nanopillars and randomly distributed nanowires is accomplished by MOCVD using (N3)2Ga[(CH2)3NMe2] as a single molecule precursor. PMID- 12123088 TI - The ascites to serum amylase ratio identifies two distinct populations in acute pancreatitis with ascites. AB - BACKGROUND/AIM: While the characteristics of ascites in the setting of chronic pancreatitis are well established, little has been written about the characteristics of spontaneous, clinically apparent ascites in the setting of acute pancreatitis. Our aim was to define the characteristics of clinically apparent ascites complicating acute pancreatitis, particularly with regard to outcomes. METHODS: We performed a search of our hospital's discharge records for ICD codes 577.0 (acute pancreatitis) and 789.5 (ascites). Clinical and laboratory variables in survivors and nonsurvivors were compared using a Mann-Whitney U test. RESULTS: We identified 59 records of which 25 cases had ascites fluid analysis. Only the ascites amylase (p = 0.033) and the ascites to serum amylase ratio (p = 0.002) correlated with mortality. Setting a cutoff of 1, the ascites to serum amylase ratio achieved a sensitivity of 83% and a specificity of 92% as a predictor of mortality. CONCLUSIONS: The ascites to serum amylase ratio identifies 2 sets of patients with ascites complicating acute pancreatitis. In patients with a high ratio, ascites may result from a localized duct disruption. In patients with a low ratio ascites may be secondary to comorbid conditions or a capillary leak. In acute pancreatitis with clinically apparent ascites, the ascites to serum amylase ratio may be a predictor of mortality. PMID- 12123089 TI - Can fluid resuscitation prevent pancreatic necrosis in severe acute pancreatitis? AB - BACKGROUND/AIMS: In previous studies, we have demonstrated that hemoconcentration was an early marker for necrotizing pancreatitis. The aim of the present study was to determine whether fluid resuscitation could prevent pancreatic necrosis among patients with hemoconcentration at the time of admission. METHODS: Data was pooled from the prior two studies of all patients with necrotizing pancreatitis and interstitial pancreatitis with a hematocrit of > or = 44 on admission. Hematocrit values in necrotizing pancreatitis and interstitial pancreatitis were compared at admission and at 24 h. Statistical analyses were performed using the Wilcoxon rank-sum test. RESULTS: A total of 39 patients satisfied our inclusion criteria, 28 with necrotizing pancreatitis and 11 with interstitial pancreatitis. Patients with necrotizing pancreatitis presented earlier than patients with interstitial pancreatitis (median 18 vs. 38 h, respectively) (p = 0.005). There was no significant difference between the intergroup median hematocrits on admission and at 24 h. All patients with hematocrits that failed to decrease at 24 h developed necrotizing pancreatitis (12/28 with necrotizing pancreatitis vs. 0/11 with interstitial pancreatitis) (p = 0.009). There was no significant difference at 24 h in rehydration among the three groups: 4.0 liters among the 12 patients with necrotizing pancreatitis whose hematocrits increased and 4.5 liters among the 16 whose hematocrits decreased at 24 h, and 4.1 liters among the 11 patients with interstitial pancreatitis (p = 0.81). CONCLUSION: Patients who presented early were more likely to have necrotizing pancreatitis than interstitial pancreatitis. While fluid resuscitation was not shown to prevent pancreatic necrosis, all patients with inadequate fluid resuscitation as evidenced by persistence of hemoconcentration at 24 h developed necrotizing pancreatitis. PMID- 12123090 TI - Increased secretion of the pancreatic secretory trypsin inhibitor (PSTI-I, monitor peptide) during development of chronic pancreatitis in the WBN/Kob rat. AB - BACKGROUND: Recent genetic investigations into cationic trypsinogen and pancreatic secretory trypsin inhibitor (PSTI) led to the conclusion that mutations in either gene can contribute to the development of (hereditary) chronic pancreatitis. Since genetic animal models are not available yet, we have studied the Wistar-Bonn/Kobori (WBN/Kob) rat, a model for chronic pancreatitis (CP). To explore the possibility that PSTI may be secreted at lower levels or contain a mutation in the WBN/Kob rat, we investigated the masses of PSTI-I and II and asked whether the ratio of PSTI/trypsinogen is decreased in animals with CP. METHODS: We collected pancreatic juice from WBN/Kob and Wistar rats aged 6-36 weeks and measured PSTI-I (ELISA) and trypsin. RESULTS: PSTI-I and -II were identified in Wistar and WBN/Kob rats by mass spectrometry and N-terminal sequencing. Using a newly developed PSTI-I ELISA, we can show that the PSTI I/trypsinogen ratio is not decreased but rather increased in WBN/Kob rats compared to healthy Wistar rats. No evidence for a PSTI mutation was found. CONCLUSION: Our data does not support the hypothesis that a dysbalance of PSTI/trypsinogen ratio is a causative factor for CP. PMID- 12123091 TI - Stenting is unnecessary in duct-to-mucosa pancreaticojejunostomy even in the normal pancreas. AB - BACKGROUND: There is a high risk of anastomotic leakage after pancreaticojejunostomy after pancreaticoduodenectomy (PD) in patients with a normal pancreas because of the high degree of exocrine function. These PD are therefore generally performed using a stenting tube (stented method). In recent years, we have performed pancreaticojejunostomy with duct-to-mucosa anastomosis without a stenting tube (nonstented method) and obtained good results. METHODS: The point of this technique is to preserve adequate patency of the pancreatic duct by carefully picking up the pancreatic duct wall with a fine atraumatic needle and monofilament thread. The results of end-to-side pancreaticojejunostomy of the normal pancreas were compared between the nonstented method (n = 109) and the stented method (n = 39). RESULTS: There were no differences in background characteristics between the groups, including age, gender and disease. The mean duration to complete pancreaticojejunostomy was 26.6 min in the nonstented group and 29.2 min in the stented group. The mean durations of surgical procedure and intraoperative blood loss were also similar in the groups. Morbidity rates due to early postoperative complications were 20.2 and 23.1%, with pancreatic leakage occurring in 7.3 and 7.7% of patients, respectively. These differences were not statistically significant. One patient in the stented group died of sepsis following leakage of pancreaticojejunostomy. There were also no significant differences in the mean time to initiation of solid food intake or postoperative hospital stay. CONCLUSION: We conclude that a stenting tube is unnecessary if the duct-to-mucosa anastomosis is completely performed. This operative technique can be considered a basic procedure for pancreaticojejunostomy because of the low risk. PMID- 12123092 TI - Clinical significance of a long common channel. AB - BACKGROUND/AIM: It was reported that in only 19 (11%) of 173 patients was the common channel at the junction of the pancreatic and bile ducts found to be 6 mm or longer. Pancreaticobiliary maljunction (PBM) is defined as an anomaly with a markedly long common channel with the junction located outside the duodenal wall, so the action of the sphincter of Oddi does not functionally affect the junction. We defined high confluence of pancreaticobiliary ducts (HCPBD) as a length of the common channel > or = 6 mm, in which the communication between the pancreatic and bile ducts was occluded when the sphincter was contracted. This study aims at investigating the clinical significance of HCPBD. METHODS: 2,980 consecutive cases with an adequate endoscopic retrograde cholangiopancreatography were reviewed. PBM and HCPBD were diagnosed according to the above definitions. PBM was divided into two groups: with or without biliary dilatation. RESULTS: PBM and HCPBD were detected in 63 (2.1%) and 50 (1.7%) cases, respectively. Biliary dilatation was detected in 30 cases having PBM. The incidences of gallbladder carcinoma associated with PBM with or without biliary dilatation and HCPBD were 13, 67, and 12%, being significantly higher than in controls (p < 0.05, p < 0.01, and p < 0.05). Pancreatic ductal reflux was detected in 11 (85%) of 13 patients with HCPBD in whom postoperative T tube cholangiograms were performed, and acute pancreatitis occurred in 14 (24%) of the 50 patients with HCPBD. CONCLUSIONS: HCPBD may be an intermediate variant of PBM. It is necessary to pay attention to an associated gallbladder carcinoma in patients with HCPBD as well as in those with PBM. PMID- 12123093 TI - Expression of E-cadherin, alpha- and beta-catenins in patients with pancreatic adenocarcinoma. AB - BACKGROUND/AIMS: E-Cadherin and its associated cytoplasmic proteins, catenins, are important mediators of epithelial cell-cell adhesion and intracellular signaling. Much evidence exists suggesting a tumor invasion suppressor role for E cadherin and catenins and loss of expression, as well as mutations, has been described in a number of epithelial cancers. The aim of this study was to evaluate the expression of E-cadherin and catenins in pancreatic adenocarcinoma tissue, and to examine the relationship between these expression and various clinicopathological parameters. METHODS: In this study, we conducted an immunohistochemical investigation of expression of E-cadherin, alpha- and beta catenins in 30 tissue samples obtained from pancreatic ductal adenocarcinoma patients undergoing surgical treatment. RESULTS: In the pancreatic mucosa of noncancerous areas, epithelial cells showed equally strong membranous expression of E-cadherin, alpha- and beta-catenin proteins at the cell-cell boundaries. Reduced expression of E-cadherin, alpha- and beta-catenins was demonstrated in 60.0, 40.0, and 56.7% of cancer tissues, respectively. Reduced expression of E cadherin, alpha- and beta-catenins correlated with tumor dedifferentiation (p = 0.012, 0.013, and 0.033, respectively). Reduced expression of E-cadherin correlated with stage and lymph node involvement (p = 0.031, 0.009, respectively). alpha-Catenin and beta-catenin expression did not correlate with the patient's age and sex, with the tumor size, location, stage and depth of invasion, or lymph node involvement and distant metastasis. CONCLUSION: These results suggest that the E-cadherin and catenins may be a useful marker of differentiation and prognosis in pancreatic adenocarcinoma, although the mechanisms underlying changes in E-cadherin and catenin expression are not fully known. PMID- 12123094 TI - Expression of the G2-M modulators in pancreatic adenocarcinoma. AB - BACKGROUND: Cell-proliferating activity is one of the prominent parameters for evaluating the biological aggressiveness of carcinomas. Pancreatic adenocarcinoma has been studied to identify modulators of the G1-S boundary. In the present study we investigated the modulators of another important checkpoint, the G2-M checkpoint. METHODS: We immunohistochemically studied three representative G2-M modulators, cdc2, cyclin A and cyclin B1 in 62 pancreatic adenocarcinomas and 7 cystadenomas. RESULTS: Overexpression of cdc2, cyclin A and cyclin B1, was observed in 54.8, 54.9 and 56.4%, respectively, of the pancreatic adenocarcinomas. cdc2 overexpression was directly related to lymph node metastasis, Ki-67 labeling index (LI), and cyclin A overexpression which was significantly linked to the stage, carcinoma differentiation, tumor size, and lymphatic invasion. On the other hand, cyclin B1 was not linked to clinicopathological parameters including Ki-67 LI and cdc2 overexpression, except for tumor size. CONCLUSION: The findings suggest that cdc2 and cyclin A play a role in the progression of pancreatic adenocarcinoma, while the clinical significance of cyclin B1 remains to be clarified because of its more random expression. PMID- 12123096 TI - Histological markers for endothelial cells in endogenous and transplanted rodent pancreatic islets. AB - BACKGROUND/AIMS: To obtain a selective marker to identify endothelial cells is difficult, due to the heterogeneity of these cells. Most described markers perform well in some applications, but fail in others. The aim of this study was to identify a selective and specific marker for rodent microvascular endothelial cells, especially for use in studies on the vascular system of pancreatic islets. METHODS: A biotin-labelled form of the lectin Bandeiraea or Griffonia simplicifolia in combination with a streptAB-Complex with alkaline phosphatase was used to stain endothelium in paraffin-embedded tissue sections from C57BL/6 mice, Sprague-Dawley or Wistar-Furth rats. RESULTS: We were consistently able to selectively stain microvascular endothelial cells in lungs, small intestines, white and brown adipose tissue, pancreas and islets of Langerhans with the lectin Bandeiraea simplicifolia. Furthermore, we were able to visualise the vasculature in syngenically transplanted islets of Langerhans in Wistar-Furth rats and C57BL/6 mice. Attempts to stain rodent endothelial cells with antibodies against CD34, CD31, CD200, Ox43, von Willebrand factor and the lectin Ulex europaeus were not uniformly successful. CONCLUSION: The lectin Bandeiraea simplicifolia is a versatile marker for rodent endothelial cells, and may be used to study revascularisation after transplantation of pancreatic islet in rodents. PMID- 12123095 TI - Expression spectrum and methylation-dependent regulation of melanoma antigen encoding gene family members in pancreatic cancer cells. AB - Human MAGE and GAGE genes encode tumor-specific antigens presented by HLA I molecules recognized on tumor cells by cytolytic T lymphocytes. To determine if pancreatic cancer patients would be suitable for MAGE- or GAGE-based immunotherapy, the expression frequency of MAGE-A1, -A2, -A3, -A4, -A6 and GAGE1 8 genes was assessed in 15 pancreatic tumor cell lines and 23 pancreatic tumor specimens using reverse transcription-polymerase chain reaction (RT-PCR). In 67% of the cell lines at least one of the MAGE-A genes was detected, 53% revealed concomitant expression of two or more genes. GAGE1-8 expression was detected in 47% of the cell lines. In the primary pancreatic tumors, MAGE-A analysis revealed exclusive MAGE-A1 and MAGE-A2 gene expression in 26 and 30% of the specimens, respectively, independent from clinicopathologic factors. Treatment of MAGE-A expression-negative pancreatic tumor cells with the demethylating agent 5-aza-2' deoxycytidine could activate MAGE-A1, MAGE-A2, MAGE-A3, MAGE-A4 and GAGE transcription suggesting silencing due to promoter methylation. Interestingly, a metastatic lesion to the liver revealed concomittant expression of MAGE-A1, -A2, A3 and -A6 consistent with a more pronounced genome-wide hypomethylation in metastases. Therefore, a subset of pancreatic cancer patients could be eligible for active, specific immunotherapy directed against MAGE-A antigens and demethylating agents could increase the number of candidate patients. PMID- 12123097 TI - Carcinoid of the pancreas. AB - Pancreatic carcinoids are very rare and usually have a poor prognosis. We describe a case of a pancreatic carcinoid with liver micrometastases in a female of 54 years of age in whom the tumor was without pronounced symptoms apart from rare episodes of flushing. The patient had been treated since November 1995 with the somatostatin analogue octreotide 200 micrograms twice daily for the first 2 years, with the long-acting analogue lanreotide 30 mg every 10 days for the following year, and then with octreotide LAR 20 mg every 28 days until the present. The flushing episodes disappeared completely, and the patient was well. Moreover, the dimensions of the tumor and the liver micrometastases remained stable during the observation period. As far as we known, this is the first case of a pancreatic carcinoid treated successfully with somatostatin analogues and having a satisfactory prognosis. PMID- 12123098 TI - Detection and prognostic impact of disseminated tumor cells in pancreatic carcinoma. AB - BACKGROUND/AIMS: Metastatic disease determines the prognosis of patients with pancreatic cancer. Current routine staging methods often underestimate the tumor stage because they do not include the search for disseminated tumor cells that spread early in different compartments of the body. Immunohistochemical and molecular methods developed recently are able to detect these cells in multiple compartments of the body. METHODS: The current status of the detection and the prognostic impact of disseminated tumor cells detected in lymph nodes, bone marrow, blood and peritoneal lavage of patients with pancreatic carcinoma are reviewed. RESULTS: Disseminated tumor cells can be detected in different compartments of the body even in early tumor stages and when a resection of the primary tumor in curative intention was performed. Furthermore, the detection of these cells has importance for the prognosis and therefore will have therapeutic implications. Standardization of the methods is a prerequisite for further studies. CONCLUSION: The detection of disseminated tumor cells should be included into studies to reveal that this increased staging has an prognostic impact and can be useful for therapeutic decisions in patients with pancreatic carcinoma. PMID- 12123099 TI - Leptin protects the pancreas from damage induced by caerulein overstimulation by modulating cytokine production. AB - BACKGROUND: Recent identification of specific leptin receptors in the pancreas suggests that this peptide may also play some role in this gland. AIM: To examine the effect of intraperitoneal (i.p.) or intracerebroventricular (i.c.v.) administration of leptin in rats on caerulein-induced pancreatitis (CIP), pancreatic gene expression of leptin and inflammatory cytokine production. METHODS: Caerulein (25 micrograms/kg) was infused subcutaneously into conscious rats over 5 h to produce CIP. Leptin (1, 5, or 10 micrograms/kg) was injected i.p. or i.c.v. 30 min prior to the CIP induction. The plasma level of TNF alpha and IL-4 was determined by ELISA, while plasma leptin was measured by RIA and leptin gene expression in pancreas by RT-PCR. RESULTS: CIP was characterized by the usual pancreatic edema, reduction in pancreatic blood flow (PBF) and an increase in serum levels of amylase, TNF alpha and IL-4. Pretreatment with i.p. or i.c.v. leptin of the CIP rats partially reversed the harmful effects of CIP on the pancreas, and reduced pancreatic inflammation and the fall in PBF. This was accompanied by a dose-dependent reduction in serum levels of amylase and TNF alpha, while serum IL-4 in the CIP rats pretreated with leptin rose dose dependently as compared to control rats with CIP alone. Pretreatment with leptin resulted in the dose-dependent rise in plasma leptin level over that observed in vehicle-treated controls. Leptin mRNA expression in the pancreas was dose dependently increased after infusion of caerulein. Leptin content in isolated pancreatic acini was also increased dose-dependently by caerulein added to the incubation medium bathing these acini. CONCLUSIONS: (1) Exogenous leptin protects the pancreas against damage by CIP; (2) endogenous leptin seems to limit the extend of pancreatic damage, and (3) these protective effects of leptin could be attributed to the reduction in TNF alpha and to the increase in IL-4 production. PMID- 12123100 TI - Inhibition of platelet aggregation by rat trophoblasts. AB - OBJECTIVE: To determine the effect of rat trophoblast cell suspensions on adenosine diphosphate (ADP)-induced platelet aggregation. MATERIALS AND METHODS: The trophoblasts were isolated from ectoplacental cones (a preplacental tissue very rich in trophoblasts) developed in rat embryos on day 12 of pregnancy (normal period of gestation: 22-23 days). Platelet-rich plasma (PRP) was obtained from adult male rats. The trophoblasts were preincubated (37 degrees C, 30 min), suspended in the medium, and then re-incubated with PRP for 3-5 min. RESULTS: 5 x 10(4) and 7 x 10(4) trophoblasts inhibited ADP-induced platelet aggregation by 10 and 18%, respectively. When the trophoblast cell concentration was increased to 1 2 x 10(5) cells, a proaggregatory phenomenon was observed, even in the absence of ADP. However, there was no inhibition of aggregation or promotion of aggregation when fixed trophoblasts or live endometrial stromal cells were incubated with PRP. CONCLUSION: The results indicate that the aggregation inhibition response was cell specific and concentration dependent. A 68-kD protein was detected in the medium when it was conditioned with 5-7 x 10(4) but not with 1-2 x 10(5) trophoblasts. However, the inhibitory or stimulatory effect does not seem to be dependent on the presence of this protein. PMID- 12123102 TI - Diagnostic dilemmas in acute intermittent porphyria. A case report. AB - OBJECTIVE: To present the importance of early diagnosis of acute intermittent porphyria (AIP) in patients with atypical presentation and discuss the diagnostic problems encountered in this case. CLINICAL PRESENTATION: A 15-year-old girl presented with upper respiratory tract infection, fever, seizures and abdominal pain. An initial diagnosis of encephalitis was made. She received antiviral drugs and anticonvulsants. Two weeks later, she developed progressive flaccid quadriplegia and facial weakness. She also developed respiratory paralysis and was intubated. Cytoalbuminous dissociation was seen in the cerebrospinal fluid. A diagnosis of severe Guillain-Barre syndrome was made. INTERVENTION: The patient received a course of intravenous immunoglobulins which did not result in any clinical improvement. Plasmapheresis, started after 12 weeks, led to partial improvement. The patient continued to have attacks of seizures, abdominal pain and vomiting with severe quadriparesis. A repeat screening test for urine porphyrins was positive, and AIP was confirmed by specific porphobilinogen deaminase in the blood. The patient was treated with large doses of intravenous glucose, followed by injections of hematin. The patient improved remarkably. She was extubated, discharged from Intensive Care Unit and started on a rehabilitation program. CONCLUSION: This patient was initially diagnosed erroneously with a negative screening test for AIP and consequently treated inappropriately. The proper diagnosis was made after repeating the screening test followed by specific tests of porphobilinogen deaminase. PMID- 12123101 TI - Is laparoscopic cholecystectomy a safe procedure for patients receiving anticoagulant therapy? AB - OBJECTIVE: To evaluate the safety of laparoscopic cholecystectomy (LC) in patients on oral anticoagulant therapy. METHODS: Four patients were involved in this study on whom the procedure was performed in a conventional manner paying meticulous attention to haemostasis. The oral anticoagulant was resumed the night of the operation, and the patient was discharged and acceptable international normalised ratio was achieved postoperatively. RESULTS: There was no thrombo embolic or haemorrhagic complication. The mean hospital stay was 5 days. CONCLUSION: LC can be safely performed in patients on oral anticoagulant therapy. PMID- 12123103 TI - Acute pancreatitis associated with Mycoplasma pneumoniae: a case report of missed diagnosis. AB - OBJECTIVE: Mycoplasma pneumoniae is responsible for approximately 20% of community-acquired pneumonia. A wide variety of extrapulmonary manifestations related to M. pneumoniae infection can occur. The diagnosis of M. pneumoniae and its association with acute pancreatitis is briefly reviewed. CLINICAL PRESENTATION: We describe the case of a 9-year-old boy with clinical features of acute abdomen. INTERVENTION: A perforated appendix was suspected and the patient was accordingly operated on. The clinical, radiological and serological findings that led to the proper diagnosis are described. CONCLUSION: The case is presented to help increasing the awareness of M. pneumoniae infection as a potential cause of acute pancreatitis in children. PMID- 12123104 TI - Acetylcholine-induced vasodilation in the uterine vascular bed of pregnant rats with adriamycin-induced nephrosis. AB - OBJECTIVE: This project was designed to study endothelium-dependent vasodilation in the uterine vascular bed during experimentally induced preeclampsia in rats. METHODS: Uterine vascular beds were isolated from non-pregnant and pregnant rats with or without treatment with adriamycin (ADR) and perfused with physiological solution. Thereafter, vasodilator responses to acetylcholine were recorded. RECORDS: Pregnant ADR-treated rats displayed symptoms of preeclampsia including hypertension and proteinuria. Blood pressure was 110.0 +/- 4.7 mm Hg (n = 5) in control pregnant rats and 136.0 +/- 5.3 mm Hg (n = 5) in ADR-treated pregnant rats, and urinary protein concentrations were 0.35 mg/ml (n = 5) and 13.2 +/- 3.6 mg/ml (n = 9), respectively. Both blood pressure and proteinuria values were significantly (p < 0.05) different between controls and ADR-treated rats. However, acetylcholine-induced dose-dependent vasodilator responses in the vascular beds were not significantly different between the pregnant and nonpregnant rats. Although acetylcholine-induced vasodilation was significantly reduced by N omega-nitro-L-arginine methyl ester hydrochloride (L-NAME) in both groups, the residual response to acetylcholine was not affected by indomethacin, suggesting that prostanoids were not involved in this response. The L-NAME resistant component, endothelium-derived hyperpolarizing factor (EDHF), was greater in ADR-treated uterine beds than in those of the controls, indicating a significant contribution from EDHF in these vessels. In the presence of an elevated external potassium ion concentration, acetylcholine produced similar vasodilator responses, indicating that the release of nitric oxide was not impaired. CONCLUSION: These results indicate that endothelium-dependent vasodilation was not impaired in this model of preeclampsia. PMID- 12123105 TI - Prevalence of primary Helicobacter pylori resistance to several antimicrobials in a Saudi Teaching Hospital. AB - OBJECTIVE: To determine the resistance rate of the most commonly used antimicrobial agents amongst Helicobacter pylori isolates. METHODS: The agar disk diffusion method (Kirby Bauer) was utilized to determine the susceptibility of 223 H. pylori strains isolated before treatment. Isolates were tested against metronidazole (5 micrograms), clarithromycin (15 micrograms), amoxycillin (10 micrograms), and tetracycline (30 micrograms). RESULTS: The resistance rate was 80% for metronidazole and 4% for clarithromycin. Tetracycline and amoxycillin showed very low degree of resistance with 1 (0.4%) and 3 (1.3%) of the strains resistant to these antibiotics, respectively. Age, sex and ethnicity had a remarkable effect on the resistance rate. CONCLUSION: The results indicate that metronidazole and clarithromycin should not be used as the only antimicrobial agents in the treatment of H. pylori infection. Susceptibility testing using the disk diffusion method is cost-effective in the screening of antimicrobial resistance against H. pylori. PMID- 12123106 TI - Apolipoprotein(a) gene polymorphism in the Korean population: is there any relevance to essential hypertension? AB - OBJECTIVE: To investigate the role of the apolipoprotein(a) [Apo(a)] gene in predicting essential hypertension in the Korean population, we undertook a case control study using the TTTTA repeat and the Met/Thr polymorphisms of the Apo(a) gene. METHODS: The study subjects consisted of 74 essential hypertensives and 211 normotensive individuals. For polymorphism analysis, DNA was amplified by polymerase chain reaction (PCR). In the case of the Met/Thr polymorphism, amplified PCR products were digested with the restriction enzyme Kspl. Genotyping was performed by a 10% nondenaturing polyacrylamide gel. RESULTS: There were no significant differences in the allele and genotype frequencies of these polymorphisms between normotensive and essential hypertensive subjects (p > 0.05). Furthermore, levels of lipoprotein(a) in the plasma were not significantly associated with the different genotypes of the APO(a) gene in either group (p > 0.05). CONCLUSION: The polymorphisms of these two loci on the Apo(a) gene are unlikely to contribute to the etiology of essential hypertension in the Korean population. PMID- 12123107 TI - Transport kinetics of alpha-aminoisobutyric acid and water in pre-eclamptic pregnancies: an in vitro study. AB - OBJECTIVE: To investigate the transport kinetics of a model amino acid, alpha aminoisobutyric acid (AIB), in the maternal-fetal direction in placentae from pre eclamptic pregnancies. METHODS: Transport kinetics of the amino acid was assessed in vitro using perfusion of human placental lobules. Control placental lobules were perfused for comparison. National Culture and Tissue Collection--135 medium diluted with Earle's buffered salt solution was used as the perfusate, and tritiated water served as the internal reference marker. AIB along with reference marker was injected as a single bolus into the maternal arterial perfusate, and serial perfusate samples were collected from maternal and fetal venous circuits for a study period of 5 min. RESULTS: The differential transport rate of the amino acid for various efflux fractions differed significantly between control and experimental groups when compared to that of the reference marker. The transport rate (corresponding to 50% of efflux into the fetal vein), transport fraction, absorption and elimination rates of the amino acid differed significantly compared to the reference marker values in study and control groups. CONCLUSION: The results indicate that amino acid transport function is compromised in placentae of pre-eclamptic pregnancies. PMID- 12123108 TI - Prevalence of Type-2 diabetes in patients with hepatitis C and B virus infection in Jeddah, Saudi Arabia. AB - OBJECTIVES: To determine the prevalence of type-2 diabetes mellitus (DM) in patients with hepatitis C virus (HCV) and B virus (HBV) infections. MATERIALS AND METHODS: A cross-sectional study of HCV- and HBV-positive patients admitted to King Abdul Aziz University Hospital, Jeddah, Saudi Arabia, was conducted from January 1999 to September 2000. The following data were collected and analysed: demographic data, the presence and type of DM, details of the treatment, body mass index (BMI), family history of DM, serum transaminases, thrombocytopenia, and presence of liver cirrhosis on liver biopsy. A total of 399 patients were included in the study. RESULTS: 165 (41%) were anti-HCV positive and 234 (59%) were HBsAg positive. Type-2 diabetes was present in 35 of 165 (21.2%) patients with HCV infection, and 33 of 234 (14.1%) with HBV infection. 94% of anti-HCV positive type-2 diabetes were older than 40 years and 6% were younger, while for nondiabetics the corresponding percentages were 55 and 45%, respectively. 76% of HBsAg-positive type-2 diabetics were older than 40 and 24% were younger, while the corresponding percentages for nondiabetics were 27 and 73%, respectively. Anti-HCV-positive type-2 diabetics, when compared to nondiabetics, had a higher BMI, a frequent family history of DM, elevated serum transminases, thrombocytopenia, and liver cirrhosis on biopsy. HBsAg-positive type-2 diabetics had only a more frequent family history of DM than did nondiabetics. CONCLUSION: Our findings indicate that type-2 diabetes is more common in patients with an HCV than with an HBV infection. PMID- 12123109 TI - Recognition of resveratrol by the human estrogen receptor-alpha: a molecular modeling approach to understand its biological actions. AB - OBJECTIVES: Resveratrol (RSVL) is an edible phytoestrogen with multifaceted health benefits that may originate from binding to the estrogen receptors. Despite its structural similarity to the estrogen receptor-alpha (ER alpha) agonist diethylstilbestrol (DES), RSVL showed distinct biological profiles in estrogen-responsive biological systems. The molecular basis of such biological profiles has been undefined. METHODS: We considered possible orientations for RSVL in ER alpha binding pocket. These conformations have been analyzed based on: (i) alignment with the key pharmacophoric elements of DES; (ii) computational energy of interaction, and (iii) pattern of accommodation at the ER alpha binding pocket. The characteristics of the most favored RSVL orientation have been compared with those of DES. RESULTS: Both RSVL and DES interacted with the catalytic amino acid triad of the ER alpha pocket (His524, Arg394 and Glu 353). However, unlike the Er alpha agonists DES and estradiol (E2), RSVL formed three additional hydrogen bonds with Gly521 and Leu525, two paramount ligand recognition residues, and with Met343 at the ER alpha binding cavity. Lastly, RSVL displayed a more favorable energy of interaction with the ER alpha binding cavity. CONCLUSIONS: The present study suggests, for the first time, that RSVL is well recognized by the human ER alpha but in a manner distinct from the pure agonists DES and E2. These variations may well entail the unique biological responses of RSVL in ER-responsive systems. PMID- 12123111 TI - The role of prevention and simple interceptive measures in reducing the need for orthodontic treatment. AB - The purpose of the interceptive orthodontic treatment is to eliminate or reduce the need for treatment in the permanent dentition. The major advantage of such a treatment may also be that it is technically simple and relatively cheap, compared to comprehensive treatment with full fixed appliances. The need for interceptive measures to avoid adverse occlusal and dental consequences, which are effects of the early loss of primary molars and nonnutritive sucking habits, is discussed. The need for prevention and early intervention in patients with anterior open bites, posterior functional crossbites and signs of ectopic erupting canines is also discussed. PMID- 12123110 TI - Gastrointestinal stromal tumors: clinicopathological and immunohistochemical features. AB - OBJECTIVES: To study the clinicopathological and immunohistochemical features of gastrointestinal stromal tumors (GISTs) in Kuwait. MATERIALS AND METHODS: Hematoxylin- and eosin-stained sections of primary gastrointestinal mesenchymal tumors were reviewed. Immunohistochemical staining was performed using a panel of antibodies to determine muscle and neural differentiation, the incidence of CD117 and CD34 expression, as well as bcl-2 and cytokeratin expression. Each stain was interpreted as negative or positive. The staining intensity of positive cases was graded as weak, moderate or strong. RESULTS: The age range was 25-80 with an average age of 54 and a male:female ratio of 3:2. The stomach was the most common site for these tumors, followed by the small intestine. Histologically, 46% were classified as malignant and 54% were benign. Most of the malignant tumors occurred in males, particularly in the stomach or small intestine. There was no significant difference in patient age between malignant and benign tumors. The most sensitive markers were muscle-specific actin for muscle differentiation and glial fibrillary acidic protein for neural differentiation. CD117 expression was seen in 81% and CD34 in 54% of all tumors. CONCLUSIONS: The results of this study show that the stomach is the most common site for these tumors, that malignant tumors are more likely to occur in the small intestine than in the stomach, and that there is no difference between benign and malignant tumors with regard to age. Our findings are comparable to those of other workers, although our male:female ratio was slightly higher. PMID- 12123112 TI - Minimal intervention in dental care. AB - High-quality appropriate dental care should encompass the concepts of effectiveness and efficiency. Many dental procedures are ineffective, and some preventive measures are inefficient. Examples of criteria that are ethically essential to use before carrying out any clinical procedure are outlined based on the concepts included in 'informed consent'. Applying the criteria rigorously will lead to minimal intervention and a more equitable distribution of appropriate dental care. Unnecessary dental care will be reduced. PMID- 12123113 TI - Minimal intervention prosthodontics: current knowledge and societal implications. AB - Minimal intervention prosthodontics can be considered a treatment option for a country's overall dental health care plan. Prosthodontics can cover a range of increasingly aggressive treatment interventions depending on the severity and progression of the disease. The 'shortened dental arch' concept is a minimal treatment intervention approach that has been advocated for a wide range of partially edentulous patients. This concept favors limited prosthodontic intervention to achieve patient-perceived acceptable function levels in the presence of multiple missing teeth. The implementation of minimal interventions should be balanced by considering risk-to-benefit ratios, as well as the consequences of nonintervention of low-level prosthodontic interventions. The 'nonintervention' approach and low-level prosthodontic interventions have inherent consequences and well-documented risks; professional ethics dictate that a practitioner present these risks as well as the known benefits of all treatment options. Developing countries are under significant pressure to effectively utilize limited resources, increase skilled human resources, provide advanced levels of care to very large numbers of patients and plan for the future dental health care of their society. Many developing countries are prime candidates for inadvertent abuse and misappropriation of prosthodontic materials, treatment modalities and human resources in trying to provide cost-effective prosthodontic care. A developing country can learn from the mistakes that developed countries have made in the past and use the evidence from these experiences to plan for a better future state of dental health for their society. PMID- 12123114 TI - Oral cancer: prevention and detection. AB - Researchers in oral cancer agree that the early diagnosis of oral carcinoma greatly increases the probability of cure with minimum impairment and deformity. Primary prevention which involves reducing the exposure to tobacco, alcohol and betel quid has been shown to be effective in reducing the incidence of oral cancer. Secondary prevention involves screening for the early detection of oral cancer. Oral cancer screening can take many forms. Clinical examination and biopsy allow the early detection of premalignant and early oral cancers. Screening can be made more efficient by inspecting high-risk sites--the floor of the mouth, the ventrolateral surface of the tongue and the soft palate. Due to the cost of population screening, it is advisable to initially target high-risk groups, those over 40 years of age, including smokers and heavy drinkers. It is recommended that dentists perform an annual visual oral cancer examination on all their patients and obtain a specialist opinion for suspicious oral lesions. Ora Test with toluidine blue may be used as an adjunct to soft tissue examination to highlight any invisible, asymptomatic lesions. Exfoliative cytology can detect early oral cancer and can be performed by dentally untrained personnel. It is rapid and relatively non-invasive and therefore may be useful in population-based oral cancer screening programmes. Recently, based on various studies, the oral CDx brush biopsy technique has been proposed as a highly accurate method of detecting oral precancerous and cancerous lesions. More frequent oral cancer examinations are recommended for treated oral cancer patients to monitor the development of secondary tumours. Family members of patients with oral cancer are also at high risk and therefore should be examined more frequently. Whatever screening method is used, a positive screening result must be confirmed by biopsy. A public awareness programme that stresses the importance of at least one annual dental examination, identification of warning signs of oral cancer and recognition of the hazards of tobacco and alcohol use is necessary to reverse the high morbidity and mortality rates associated with this disease. In the future, the identification of oncogene and tumour suppressor gene mutations in biopsy specimens may give a clearer indication of the likely behaviour of suspicious oral lesions. PMID- 12123115 TI - Oral health in Kuwait before the Gulf War. AB - OBJECTIVE: The aim of this oral health survey was to determine the oral health status and oral health behavior in the whole population in Kuwait. This survey was part of the Kuwait Health Survey, which was conducted by the Ministry of Public Health. The original data were destroyed during the Gulf War (1990/1991), but the aim of this report is to publish the main findings concerning oral health and related factors. METHODOLOGY: The data were collected between April 1984 and April 1985. The sample consisted of 3,358 households and 26,530 individuals. The survey consisted of interviews and clinical examinations (> 12-year-olds). WHO (1977) criteria were used for examinations. RESULTS: Females were more often brushing their teeth at least once a day than males and non-Kuwaitis slightly more often than Kuwaitis. Over one third (39%) had visited a dentist during the previous 12 months. The proportion of subjects with soft deposit was 66%, calculus 45%, intensive gingivitis 46% and advanced periodontitis 18%. The caries experience was prevalent (52%) in primary dentition (< 8-year-olds) and in permanent dentition, highest (95%) among the 60- to 64-year-olds. CONCLUSION: Preventive programs for periodontal diseases and for dental caries are urgently needed. The coverage of curative care of both diseases also needs to be improved. Health behavior improvement should be targeted by oral health promotion activities. PMID- 12123116 TI - Clinical trial of a new glass ionomer for an atraumatic restorative treatment technique in class I restorations placed in Latvian school children. AB - OBJECTIVE: The aim of this study was to evaluate the performance of a new glass ionomer filling material (Chem-Flex) using the atraumatic restorative treatment (ART) approach in class I cavities in the permanent dentition of Latvian schoolchildren. METHODS: A total of 63 fillings (40 test and 23 control) were placed using the ART technique in 41 schoolchildren in Riga at the Stomatology Institute of the Medical Academy of Latvia. These fillings were then blindly assessed after 2 years. RESULTS: The complete success rate for both the test and the control material fillings were 92.5 and 94.9%, respectively. CONCLUSION: The new glass ionomer filling has shown a good performance in terms of retention, marginal failure and fractures in class I cavities. PMID- 12123117 TI - The role of minimal intervention in orthodontics. AB - Less than 15% of the population develops a normal occlusion as defined by Angle in the permanent dentition. The term 'ideal' may therefore be a more appropriate description, and deviations from this esthetic and functional optimum should not be considered abnormalities in the true sense of the word. Current research indicates that few malocclusions compromise dental, periodontal or temporomandibular health. To determine whether or not protruded, irregular or maloccluded teeth merit orthodontic intervention is therefore a major challenge. Another challenge is to determine to what extent a limited treatment strategy may be successful in correcting the occlusal problems according to the perceived needs of the patients. I discuss these issues in my present communication and conclude that the major reasons for recommending orthodontic treatment are psychosocial in nature. I also conclude that the majority of the orthodontic cases require comprehensive treatment to achieve successful results. Orthodontic patients are typically satisfied with the outcome of the orthodontic intervention, reporting that the esthetic improvements and the increased functional comfort of the dentition have made significant contributions to their quality of life. Technological advances have made orthodontic treatment simpler and safer over the years. Considering the potential for long-lasting results and the low risk of iatrogenic effects if the patients comply with appropriate oral hygiene measures during active appliance therapy, I conclude that treatment of minor occlusal deviations may also be justified in subjects expressing a clearly defined subjective need for treatment. PMID- 12123118 TI - [Etiopathogenicity of primary, secondary, and tertiary hyperparathyroidism: implications of molecular changes in treatment failure]. PMID- 12123119 TI - [Efficacy and effectiveness: a useful distinction for clinical practice and research]. PMID- 12123120 TI - [Do we take full advantage of hemodialysis?]. PMID- 12123121 TI - [Prevention strategies for diabetic nephropathy]. PMID- 12123122 TI - [Conventional and vectorial analysis of bioimpedance in clinical practice]. PMID- 12123123 TI - [Clinico-epidemiologic study of urolithiasis in a Caribbean urban area]. AB - Urolithiasis is a common clinical disorder. Its frequency has risen with the development of humanity and varies wirl the country, geographic area, etc. It poses health problems in most countries. The urolithiasis has some potential risk factors such as intrinsic and extrinsic epidemiological, metabolic, physic chemistry of the urine, mechanics and urinary infection. Our objective in this epidemiological study in a general population was to know the frequency, the potential risk factors, the morbidity, and social and economical impact of the urolithiasis in our subtropical Caribbean country. The prevalence was 4.64% and the annual incidence was 0.1%. Both are with in the estimated range of urolithiasis frequency in the world. It mainly started between 20 and 29 years in both genders. The white (5.2%) and the male (6.36%) patients were the most affected. 40% of all patients had a family history of urolithiasis. It was highly associated with diabetes mellitus, ischaemic cardiopathy, urinary tract infection and arterial hypertension. Stone formation was related to the warmer season. High calcium, protein-purine, carbohydrates and oxalic acid intake together with low fluid intake were closely associated with this disorder. 85% of patients had suffered renal colic and 75% of them more than once. Stone recurrence affected 33.8% of patients and 54.5% of them had more than one recurrence. Procedures for stone removal were needed in 33.8% of subjects. 40% of all patients were admitted to hospital due to urolithiasis morbidity. Non-specific medical treatment had been taken by 49.2% of the patients and specific treatment by none. Urolithiasis in this population was the some as has been reported in others studies. It has shown high frequency, increasing incidence, the same risks factors, high morbidity, and high social and economical impact. The low cost treatment is only taken by half of the patients. PMID- 12123124 TI - [Poor adherence to diet in hemodialysis: role of anxiety and depression symptoms]. AB - OBJECTIVE: To determine whether symptoms of anxiety or depression are factors associated with poor compliance of fluid restriction and dietary selection in chronic renal failure patients in hemodialysis. PATIENTS AND METHODS: A cross sectorial, descriptive, comparative and correlation study was designed between january and march of 2000, patients were selected in 3 hemodialysis centers of Social Security in Lima with Karnofsky index > 80, without an acute failure of treatment or default from therapy in the last 3 months. Depressive symptoms were evaluated with Beck Depression Inventory and Anxiety Symptoms with the Zung Scale. Poor compliance with fluid restriction was defined as an interdialytic weight gain > or = 2.5 kg and dietary restriction as a level of predialysis serum potassium > or = 6 meq/L. The evaluation of risk factors was made with a simple and multiple logistic regression analysis. RESULTS: Eighty eight patients were selected, 47 (53.4%) were men, the average values of age, time on dialysis, level of creatinine and hemoglobin were respectively 55.9 +/- 15.8 years old, 48.8 +/- 38.8 months, 8.5 +/- 1.9 mg/dl and 7.7 +/- 1.4 g/dl. The number of patients with adequacy of dialysis, depressive symptoms, anxiety symptoms, poor compliance with fluid restriction and dietary selection were respectively 50 (62.5%), 54 (61.4%), 46 (52.3%), 47 (53.4%) and 31 (35.2%). The multiple logistic regression analysis showed that depressive symptoms are the only factor associated with poor compliance with fluid restriction (OR = 2.7, p = 0.002) and dietary selection (OR = 2.5, p = 0.0067). Depressive symptoms and them severity had a positive correlation with poor compliance. CONCLUSION: Depressive symptoms and its severity is associated with a higher interdialitytic weight gain and higher predialysis serum potassium in hemodialysis patients. Early diagnosis and therapeutic intervention might benefit these patients. PMID- 12123125 TI - [Analysis of survival in dialysis: hemodialysis versus peritoneal dialysis and the significance of comorbidity]. AB - To choose the best possible dialysis technique for those patients with end-stage renal disease continues to be a matter of debate. Even after putting aside the evident influence that economic and geographic factors as well as the health politics may have in the selection of the technique, different studies comparing survival between hemodialysis (HD) and peritoneal dialysis (PD) have shown contradictory results which could be explained by the differing methodological and statistical methods used together with the different influence assigned to the comorbidity found when starting the treatment, a situation that has increased the confusion about this topic. Based on this we performed a retrospective analysis with a follow-up time of seven years including all those patients who started dialytic treatment in our area, with a final number of 3.106 hemodialysis patients and 542 peritoneal dialysis patients. Those patients who were transferred to another treatment technique during the time of the study were excluded. Age higher than 70 years, cardiovascular disease, liver disease, diabetes mellitus and the presence of dyslipidemia were included as comorbidity factors. Peritoneal dialysis patients were younger than those treated by hemodialysis (54.53 vs 60.1 years), but suffered from higher cardiovascular comorbidity and were more often diabetic. The global survival was the same in both groups up to 32 months of treatment. Although no differences were found when comparing those patients without comorbidity factors, those with comorbidity had better survival on hemodialysis. Age higher than 70 years was the only comorbidity factor with statistically significant difference for a better survival in hemodialysis. PMID- 12123126 TI - [Changes in health-related quality of life in the first year of kidney transplantation]. AB - OBJECTIVE: To study the changes in the health related quality of life (HRQOL) during the first year following renal transplant (RT), comparing azathioprine vs mycophenolate mofetil (MMF) in triple immunosuppressant therapy with prednisone and cyclosporine. METHODS: Prospective, open and random study with 26 patients who received a primary cadaveric renal transplant consecutively. Analysis of clinical variables: delayed graft function (DGF), acute rejection (AR), infections and comorbidity; analytical: haemogram, albumin and serum creatinine, hepatic function, cyclosporin levels; instruments for assessing the HRQOL: Psychological General Well-being Index (PGWBI) and Euroqol-5d (EQ-5d) health questionnaire, which includes a self-assessment scale of the state of health, Visual Analogical Scale (VAS). Controlled collection of data upon discharge following renal transplant, and subsequently 1, 3, 6, and 12 months following the first questionnaire. RESULTS: There were no differences between patients on azathioprine or MMF, except that AR occurred less frequently with MMF (7% vs 42%, p = 0.065). Global AR: 23%, cytomegalovirus infection/illness: 81%/8%, readmissions: 42%. There was an improvement in HRQOL measured by the progressive increase in the scores on the PGWBI, EQ-5d and VAS during the first three months following RT. By the sixth month, in comparison to the third, a decrease in the score was observed (PGWBI, p = 0.011). Later the HRQOL improved, but without reaching the maximum scores achieved. CONCLUSIONS: Patients on MMF showed less frequent incidence of AR. The HRQOL decreases during the third and the sixth month, but with less intensity in patients on MMF, probably related to the lower rate of AR, and excluding the over 60s. PMID- 12123127 TI - [Cost of maintenance immunosuppressive drugs in kidney transplantation]. AB - METHOD: We calculated the cost of maintenance immunosuppression (ISM) in 405 kidney transplant patients under treatment for more than one year, classifying them according to the combination of drugs used and whether the ISM continued to be the same as initially indicated (primary ISM), or a later adaptation. Basic clinical data were also acquired on the through levels of drugs and the use and cost of the associated medication. RESULTS: The mean doses in mg/kg/day and the trough levels in ng/ml were: cyclosporin (Cs), 2.7 +/- 0.9 and 123 +/- 5; tacrolimus (T), 0.09 +/- 0.06 y 7.3 +/- 2.9; Mycophenolate mofetil (MMF), 21.6 +/ 6.8 and 3.3 +/- 2.4. The proportion of patients with doses of steroids (EST) below 5 mg/day among patients treated with MMF was 60% in association with Cs and 73% with T. The cost related to EST and azathioprine (AZA) is very low; the mean cost per patient and day for Cs, T and MMF, was 7.08, 12.59 and 9.53 euros, respectively. The annual cost per patient depending on treatment was, in euros: EST + AZA, 204; Cs or without AZA and/or EST: 2,555; T with or without AZA and/or EST: 4,616; Cs plus MMF with/without EST: 6,136; T plus MMF with/without EST: 7,212. In cases of primary ISM, the difference between the two treatments with MMF drops so, low that its loses statistical significance (7,206 and 6,443 euros). This is because the dose of MMF is 50% higher in cases treated with Cs, even though there is no difference in the trough level of MMF. The use of hypolipidemic agents (with a mean annual cost of 451 euros) is much lower in treatments with T in comparison with Cs (13.8% vs 41.4%, p < 0.001). No significant differences could be established between T and Cs regarding the use of hypotensive agents and the frequency of postransplant insulin dependent diabetes. CONCLUSIONS: The treatment of ISM with T is more expensive than Cs based treatment but the difference is reduced in combination with MMF because of the use of lower doses of MMF in the association of T with MMF. The considerable economic impact of ISM would justify the provision of greater resources to the rationalisation of the treatments currently being applied. PMID- 12123128 TI - [Acute interstitial nephritis caused by clozapine]. AB - We are reporting a case of a 69 years old man developed acute renal failure due to interstitial nephritis during treatment with the drug clozapine, and that improve after to discontinuation of this drug. The clozapine is a new antipsychotic drug that may produce severes adverses reactions, like medullary toxicity. Recently 10 cases of clozapine-induced AIN have been reported. We want to associate to the cases publicated with a new case because it is a severe adverse reaction. PMID- 12123129 TI - [Acute renal failure caused by cholesterol embolism in kidney transplantation from cadaver donor. Is the prognosis more favorable than in native kidneys?]. AB - We present the case of a 47 years old women with a third cadaveric kidney transplant. After surgery, she had effective diuresis reaching a serum creatinine of 2.2 mgs% at 19 postoperative day. In the nest few days, the patient was oliguric with worsening of renal function. The ultrasound examination excluded urinary obstruction. With the suspicion of acute rejection, a renal biopsy was performed. The histopathological record disclosed cholesterol emboli with a widespread multifocal ischemic infarct and eosinophilic tubulointerstitial nephritis. The renal function deteriorated in the seven next days and peritoneal dialysis was carmedow. She then recovered diuresis with improvement of renal function, reaching at discharge a serum creatinine of 1.8 mgs%. The renal function remains stable after 3 years. We analysed the etiopathogenic factors of this disease and the possible beneficial effects of immunosuppresive drugs in better prognosis compared with the same entity in native kidneys. PMID- 12123130 TI - [Foam cell nephropathy in heterozygous female with Fabry's disease]. AB - We describe the clinical and pathological characteristics of a 49 year-old heterozygous female carrier of Anderson-Fabry's disease. Light microscopy and ultrastructural study of a renal biopsy showed the presence of foam cell nephropathy and galactosylceramide deposits affecting podocytes, the parietal epithelium of Bowman's capsule and the distal tubular cells, endothelial cells and medullary interstitial elements. Retrospectively, the presence of storage disease was confirmed in a hysterectomy specimen obtained two years previously. Our observation shows that heterozygotes for this disorder can not only carry and transmit the disease but may also develop pathological deposits in various organs. A renal biopsy from these carriers allows precise identification of the disease, facilitates adequate genetic counseling and gives the option of enzyme replacement therapy in patients who have pathological deposits. PMID- 12123131 TI - [Acute renal failure caused by dihydroergotamine]. PMID- 12123132 TI - [Surgical treatment of aortic aneurysm in patients with kidney transplantation: report of 2 cases]. PMID- 12123133 TI - [Chronic terminal kidney failure: a not very "adequate" expression]. PMID- 12123134 TI - [Kidney transplantation in a patient with HIV infection discovered during transplantation. Initial course]. PMID- 12123135 TI - [Rhabdomyolysis, acute renal failure and use of cerivastatin combined with gemfibrozil. New evidence]. PMID- 12123136 TI - [Risk factors and cardiovascular disease in uremic patients]. PMID- 12123138 TI - [Cardiovascular risk and dyslipidemia after renal transplantation]. PMID- 12123137 TI - [Coronary disease in renal transplantation]. PMID- 12123139 TI - [Cardiovascular risk factors in renal transplantation: clinical markers]. PMID- 12123140 TI - [Prevention of cardiovascular risk in renal transplantation. Consensus document]. PMID- 12123141 TI - [Cardiovascular risk in patients with renal transplantation]. PMID- 12123142 TI - Ligamentous injuries about the ankle and subtalar joints. AB - Ligamentous injuries at the ankle and subtalar joint range from simple sprains to severe talar dislocations. While lateral ankle sprains are among the most frequently encountered injuries and do not pose a greater diagnostic problem, the surgeon must be suspicious not to overlook associated ligamentous injuries at the subtalar and midtarsal level that may result in chronic painful conditions. Syndesmotic instabilities with or without ankle fractures must be assessed carefully and treated properly, since these are prearthrotic conditions. In the treatment of chronic ankle or subtalar instability tenodeses provide mechanical stability while reducing subtalar mobility. Anatomic reconstruction methods therefore should be considered for both conditions. PMID- 12123144 TI - Physical examination of the ankle for ankle pathology. AB - Injury to the ankle joint is very common, but the etiology of the ankle pain can be difficult to determine due to the complexity of the joint. Without a proper diagnosis, treatment is delayed or inappropRiate which can lead to further ankle morbidity. This article is to assist foot and ankle specialists in making an accurate diagnosis of ankle pathology by showing how to perform a thorough ankle examination. Specific tests and examination techniques are described. No exact treatment plan to a particular diagnosis is given, but rather a general approach to treating ankle injuries is shown. PMID- 12123143 TI - Achilles rupture in the athlete. Current science and treatment. AB - Achilles tendon ruptures became increasingly common in the latter half of the 20th century. Once the diagnosis is made, the patient's goals and objectives should be clearly stated. The treatment choice should incorporate the patient's needs, desires, objectives, and functional goals to assure an optimal result. PMID- 12123145 TI - Soft-tissue disorders of the ankle: a comprehensive arthroscopic approach. AB - Ankle arthroscopy has dramatically advanced in the past decade. Many ailments of the ankle joint that were previously treated through open approaches are today treated with arthroscopic techniques. Arthroscopy allows for a more rapid recovery, better visualization of soft tissue lesions within the ankle joint and a more thorough examination of intracapsular ankle pathology. This article describes current arthroscopic treatments of ankle pathology limited to soft tissue structures. Additionally, a perspective is presented for the comprehensive treatment of lateral ankle pain including arthroscopic lateral ankle stabilization. PMID- 12123146 TI - MRI of soft tissue disorders of the ankle. AB - In summary, MRI has become the dominant imaging modality for assessing soft tissue disorders of the ankle. It is useful in evaluating patients with acute or chronic ankle pain or instability, and for diagnosis and staging of soft tissue mass lesions. MRI often provides information that is essential to treatment planning of a variety of traumatic, degenerative, and neoplastic lesions. PMID- 12123147 TI - Conservative treatment of acute lateral ankle sprains. AB - Lateral ankle sprains are among the most common sports injuries. Although ankle sprains are treated conservatively at the present time, for years the treatment was based on acute repair of the ruptured ligaments. Several differing opinions currently exist as to the treatment of lateral ankle sprains. A review of the literature and explanation of the benefits and risks of each treatment protocol is undertaken. PMID- 12123148 TI - Physical therapy rehabilitation of the ankle. AB - Physical therapy intervention following an ankle injury is crucial and essential to returning a patient to his/her prior level of function. Following a physical therapy evaluation, a physical therapy diagnosis is established by relating the physical impairments found (e.g., limitations in range of motion, strength, proprioception, etc.) to functional limitations. The goal of physical therapy intervention is to improve these physical impairments, thereby restoring a patient's normal function. The physical therapist can administer such treatment as joint mobilization, strength training, proprioceptive training, and patient education. Since individuals vary in the extent and severity of physical impairments, physical therapy intervention will also vary on a patient-by-patient basis. Therefore, the purpose of this article is not to serve as a protocol for physical therapy intervention but as a review of evidence-based treatment that is relevant for the impairments found after completing a physical therapy evaluation. PMID- 12123149 TI - A simple algorithm for identifying negated findings and diseases in discharge summaries. AB - Narrative reports in medical records contain a wealth of information that may augment structured data for managing patient information and predicting trends in diseases. Pertinent negatives are evident in text but are not usually indexed in structured databases. The objective of the study reported here was to test a simple algorithm for determining whether a finding or disease mentioned within narrative medical reports is present or absent. We developed a simple regular expression algorithm called NegEx that implements several phrases indicating negation, filters out sentences containing phrases that falsely appear to be negation phrases, and limits the scope of the negation phrases. We compared NegEx against a baseline algorithm that has a limited set of negation phrases and a simpler notion of scope. In a test of 1235 findings and diseases in 1000 sentences taken from discharge summaries indexed by physicians, NegEx had a specificity of 94.5% (versus 85.3% for the baseline), a positive predictive value of 84.5% (versus 68.4% for the baseline) while maintaining a reasonable sensitivity of 77.8% (versus 88.3% for the baseline). We conclude that with little implementation effort a simple regular expression algorithm for determining whether a finding or disease is absent can identify a large portion of the pertinent negatives from discharge summaries. PMID- 12123150 TI - Use of an interactive tool to assess patients' willingness-to-pay. AB - Assessment of willingness to pay (WTP) has become an important issue in health care technology assessment and in providing insight into the risks and benefits of treatment options. We have accordingly explored the use of an interactive method for assessment of WTP. To illustrate our methodology, we describe the development and testing of an interactive tool to administer a WTP survey in a dental setting. The tool was developed to measure patient preference and strength of preference for three dental anesthetic options in a research setting. It delivered written and verbal formats simultaneously, including information about the risks and benefits of treatment options, insurance, and user-based WTP scenarios and questions on previous dental experience. Clinical information was presented using a modified decision aid. Subjects could request additional clinical information and review this information throughout the survey. Information and question algorithms were individualized, depending on the subject's reported clinical status and previous responses. Initial pretesting resulted in substantial modifications to the initial tool: shortening the clinical information (by making more of it optional reading) and personalizing the text to more fully engage the user. In terms of results 196 general population subjects were recruited using random-digit dialing in southwestern Ontario, Canada. Comprehension was tested to ensure the instrument clearly conveyed the clinical information; the average score was 97%. Subjects rated the instrument as easy/very easy to use (99%), interesting/very interesting (91%), and neither long nor short (72.4%). Most subjects were comfortable/very comfortable with a computer (84%). Indirect evaluation revealed most subjects completed the survey in the expected time (30 min). Additional information was requested by 50% of subjects, an average of 2.9 times each. Most subjects wanted this type of information available in the provider's office for use in clinical decision making (92%). Despite extensive pretesting, three "bugs" remained undiscovered until live use. We have demonstrated that the detailed information, complex algorithms, and cognitively challenging questions involved in a WTP survey can be successfully administered using a tailor-made, patient-based, interactive computer tool. Key lessons regarding the use of such tools include allowing the user to set the pace of information flow and tailor the content, engaging the user by personalizing the textual information, inclusion of tests of comprehension and offering opportunities for correction, and pretesting by fully mimicking the live environment. PMID- 12123151 TI - Symbolic representation of anatomical knowledge: concept classification and development strategies. AB - In this paper a novel approach to anatomy knowledge representation is described. The focus of the present research has been on the development of a representational framework where the conceptual level has been implemented by using hierarchical and nonhierarchical conceptual networks. This has allowed handling the demand for multiple views of anatomy (systemic and topographical views). The terminological level of the knowledge representation has been implemented by using a compositional strategy which has avoided the explicit storage of the terms used to express composite concepts. Hierarchical relations and composite concept representations have required supervision of both the inheritance and concept reconstruction. For this purpose heuristic knowledge has been stored in terms of consistency rules in the knowledge base. As proof of the capability of this system, we show how the knowledge base has been used to provide symbolic access to spatial information consisting of a reduced set of images from the Visible Human Dataset. PMID- 12123154 TI - Infected patients win. PMID- 12123153 TI - Analysis of complex decision-making processes in health care: cognitive approaches to health informatics. AB - Decision making by health care professionals is often complicated by the need to integrate ill-structured, uncertain, and potentially conflicting information from various sources. In this paper cognitive approaches to the study of decision making are presented within the context of a variety of complex health care applications. In recent years it has become increasingly accepted that in order to build information systems that can support complex decision making it will be necessary to more fully understand human decision-making processes. Methodological approaches are described that aim to explicate the decision making and reasoning skills of subjects as they perform activities involving the processing of complex information. The paper begins by presenting the theoretical foundations for cognitive analyses of decision making, including discussion of major approaches to the study of decision making in a range of real-world domains, including medicine. Applications of cognitive approaches are then illustrated, including a description of a study in which subjects were asked to "think aloud" in providing treatment decisions for complex medical cases. The resulting protocols were then analyzed for subjects' use of decision strategies and problems in reasoning. Extension of cognitive approaches to the study of group decision-making processes is also described. Recent approaches are discussed which borrow from advances in the study of human-computer interaction and which utilize video analysis of decision-making activities involving information technologies. Using these approaches it has been found that health care information systems, such as computerized patient record systems, may have inadvertent effects on human decision making. Implications of a cognitive approach to improving our understanding of complex decision making are discussed in the context of developing appropriate computer-based decision support for both individuals and groups. PMID- 12123152 TI - An evaluation of extended vs weighted least squares for parameter estimation in physiological modeling. AB - Weighted least squares (WLS) is the technique of choice for parameter estimation from noisy data in physiological modeling. WLS can be derived from maximum likelihood theory, provided that the measurement error variance is known and independent of the model parameters and the weights are calculated as the inverse of the measurement error variance. However, using measured values in lieu of predicted values to quantify the measurement error variance is approximately valid only when the noise in the data is relatively low. This practice may thus introduce sampling variation in the resulting estimates, as weights can be seriously mis-specified. To avoid this, extended least squares (ELS) has been used, especially in pharmacokinetics. ELS uses an augmented objective function where the measurement error variance depends explicitly on the model parameters. Although it is more complex, ELS accounts for the Gaussian maximum likelihood statistical model of the data better than WLS, yet its usage is not as widespread. The use of ELS in high data noise situations will result in more accurate parameter estimates than WLS (when the underlying model is correct). To support this claim, we have undertaken a simulation study using four different models with varying amounts of noise in the data and further assuming that the measurement error standard deviation is proportional to the model prediction. We also motivate this in terms of maximum likelihood and comment on the practical consequences of using WLS and ELS as well as give practical guidelines for choosing one method over the other. PMID- 12123155 TI - UM study: antibiotics may help some periodontal patients. PMID- 12123156 TI - Understanding coordination of benefits--Part One. PMID- 12123158 TI - What are strategic health authorities? PMID- 12123157 TI - Effect of TMJ and masticatory muscle on the range of mandibular motion. PMID- 12123159 TI - Would you care to complain? PMID- 12123160 TI - Here's looking at you. PMID- 12123161 TI - Is the Care Standards Act a missed opportunity? PMID- 12123162 TI - Secrets of successful night nursing. PMID- 12123163 TI - A grief unobserved. PMID- 12123164 TI - Boats, bugs and babies. PMID- 12123165 TI - Part 1. Pressure ulcer prevention. Best practice statements. AB - Although pressure ulcer prevention is an important issue in all areas of health care, a lack of randomised controlled trials has led to the development of clinical guidelines that rely on low-grade evidence. The best practice statement on pressure ulcer prevention consists of five sections: education and audit, risk assessment, skin inspection, positioning, and mattresses, chairs and cushions. It was developed using the best available evidence and the expert opinion of 30 tissue viability nurse specialists. PMID- 12123166 TI - The case for the formal education of HCAs. AB - Recent government policies have legitimised the role development of all health care practitioners as a means of improving patient care and service delivery. This study focuses on the professional development of health care assistants (HCAs) and how this relates to patient care. National inconsistencies in training provision and the need to standardise the education of HCAs through registration are discussed. The implications for nursing practice are also examined. PMID- 12123167 TI - Using research to improve ophthalmic nursing care. AB - This article focuses on the way in which the nursing care of patients undergoing vitreoretinal surgery was developed as a result of an action research project. The research project is described and the following practice developments are discussed: the preoperative preparation of patients; the physiological effects of posturing; communication between patients, staff and the multidisciplinary team; and the knowledge base and clinical skills of nurses. These developments are evaluated and suggestions for future practice are made. PMID- 12123168 TI - Emotional intelligence: its role in training. AB - The ability to manage your own emotions while interpreting other people's is a useful skill in any caring environment, yet emotional intelligence is often overlooked in training programmes. Incorporating it into the curriculum will give nurses greater understanding of themselves and the way they relate to others, enabling more effective interactions with patients. It will also equip them emotionally to deal with highly charged situations. The authors describe a model for nurse education that teaches and promotes emotional intelligence. PMID- 12123169 TI - Professional development. The skin. 3. Managing patients with wounds. PMID- 12123170 TI - Day in the life. PMID- 12123171 TI - Dignity not dependence. PMID- 12123172 TI - Posttransplant viral syndromes in pediatric patients: a review. AB - The success of pediatric solid organ transplantation has been largely due to advancements in surgical techniques, technology, and preoperative and postoperative care. Potent immunosuppression continues to reduce the incidence and severity of rejection, and improve long-term survival. However, there is growing awareness of the role immunosuppression plays in contributing to the incidence of cytomegalovirus, Epstein-Barr virus, and Epstein-Barr virus associated posttransplant lymphoproliferative disease. Herpes viruses such as these present as primary or recurrent disease and continue to be a significant source of morbidity and mortality in transplant recipients. This paper reviews the predictors of disease, clinical features, diagnosis, and methods of treatment of these major posttransplant viral syndromes. As part of the human herpes virus family, varicella-zoster virus will also be discussed. A case study shows the delicate balance of treating concomitant varicella infection at the time of transplantation. PMID- 12123173 TI - Complications following split-liver transplantation: a pediatric case study. AB - Liver transplantation using split and reduced livers has helped to expand the donor pool for pediatric patients in the presence of a severe cadaveric organ donor shortage. However, this technique has been associated with an increased rate of postoperative complications. The purpose of this paper is to review a case report of postoperative complications experienced by a pediatric patient following split-liver transplantation for fulminant hepatic failure of unknown etiology. PMID- 12123174 TI - Pediatric kidney transplantation: growth, development, and nursing implications. AB - The complex issues related to the growth and development of pediatric kidney transplant recipients are explored in this paper. We divide the pediatric population into 3 age groups--toddlers and preschoolers, school age children, and adolescents--and review the literature describing growth and development in kidney transplant recipients and the normal population briefly for each age group. Planning and delivery of nursing care that is based on the implications of growth and development are discussed, and have relevance for all allied healthcare professionals caring for pediatric kidney transplant recipients and their parents. Allied healthcare professionals in adult settings who provide care to recipients who received a transplant before the age of 18 may also benefit from reviewing this article. PMID- 12123175 TI - Pediatric liver transplantation. AB - Pediatric liver transplantation is a fast-growing and challenging field. Healthcare providers must stay informed of advancements in the management of liver transplant candidates and recipients. The goal of this paper is to provide nurses who care for pediatric liver transplant candidates and recipients with a review of the basic medical management of these patients, from the preoperative evaluation to postoperative care. PMID- 12123176 TI - Embracing the uniqueness of children. PMID- 12123177 TI - Pediatric recipients of living donor lobar lung transplants: postoperative care. AB - Bilateral living donor lobar lung transplantation is a treatment option for selected children and adults with end-stage lung disease. Careful donor evaluation, skilled intraoperative management and surgical technique, and diligent immediate postoperative care and follow-up all contribute to better outcomes. Although medical management of whole lung transplant recipients in the immediate postoperative period is similar to that of lobar lung transplant recipients, there are specific differences. Anatomical distinctions, such as the entire cardiac output flowing to 2 lobes instead of 5, and thoracic space issues with simultaneous mechanical ventilation and chest tube suction, contribute to these differences. Early postoperative care, including initial postoperative stabilization, ventilation, fluid management, rejection/infection surveillance and prophylaxis, and beginning rehabilitation, can be adapted to ensure successful outcomes in these patients. PMID- 12123178 TI - Developmental approach to pediatric transplantation. AB - Children who need or who have received an organ transplant endure many hospitalizations, procedures, and surgeries. These experiences are often traumatic for children, repeatedly causing intense fear and anxiety. How stressful situations are perceived and responded to by children depend on their cognitive, physical, emotional, and social development. Alternatively, the introduction and repetition of stressful hospitalizations and medical experiences can delay development in any of these areas. It is important that healthcare professionals recognize potentially stressful situations and know how to introduce appropriate interventions to ease anxiety and facilitate each child to reach his or her developmental potential. Healthcare professionals should also have an understanding of child development and the effect it has on the child's understanding and reaction to transplantation. PMID- 12123179 TI - Retransplantation of multiple organs: how many organs should one individual receive? AB - Every year, the transplant waiting list gets longer, while donor numbers essentially remain the same. This makes the responsibility of being good stewards of this precious and limited resource greater than ever. Transplant teams, who are both committed to their patients and aware of this important responsibility, are left to make the difficult and ethical decisions regarding retransplantation. Retransplantation of organs in pediatric patients presents a different set of issues to consider, and the results are promising. This case study presents a boy who received a kidney transplant for focal segmental glomerulosclerosis at age 5. At age 11, because of the recurrence of focal segmental glomerulosclerosis and severe cardiomyopathy, he required a rare combined kidney-heart transplant. At age 17, he developed chronic renal failure and posttransplant coronary artery disease, most likely related to a period of noncompliance, and required yet another combined kidney-heart transplant. He is currently alive and well 2 years after transplantation. PMID- 12123180 TI - Intestinal transplantation in pediatric patients. AB - Intestine transplantation has evolved into a feasible alternative for children with permanent intestinal failure and life-threatening complications related to total parenteral nutrition. Although the first transplantations were done nearly 40 years ago, long-term survival has only been achieved in the last decade. Nearly 700 intestinal transplantations have been performed internationally since 1985, with an overall patient survival of greater than 50%. Improvements in patient selection, medical management, and assessment and treatment for rejection and infection have contributed to the increased survival. This article will discuss current results and medical management strategies for this innovative type of transplantation for children with end-stage short gut syndrome. PMID- 12123181 TI - What is a 'normal' midwife? AB - When you look through job advertisements these days, there are many different titles for midwifery roles. From Consultant to Research & Development Midwife, breastfeeding specialist to Sure Start ... it makes me wonder what has happened to the 'normal' midwife. Last year (June 2001) we had a focus on Normal Birth- this time we are asking, what is a 'normal' midwife? What skills does she/he have? What does she/he do to keep things 'normal'? A clinical midwife, a researcher, an educationalist and a consumer give us their views. PMID- 12123182 TI - Are midwives a dying breed? PMID- 12123183 TI - Where's the midwifery in midwife-led care? PMID- 12123184 TI - The Oakwood practice. PMID- 12123185 TI - Midwives performing instrumental deliveries. Programme development and personal reflection. PMID- 12123186 TI - Numbers for life. Midwives will shortly be responsible for allocating NHS numbers to neonates. PMID- 12123187 TI - The world wide wonder. PMID- 12123188 TI - Wirral Brook Advisory Centre. Teenage Pregnancy/Midwifery Project 2001. PMID- 12123189 TI - Trust in women.... Don't trust the 'patient'. PMID- 12123190 TI - The genesis of the incubator. PMID- 12123191 TI - Cardiotocography versus intermittent auscultation. Using pinnards and Dopplers in low risk women. PMID- 12123192 TI - Medecins Sans Frontieres: mission impossible? PMID- 12123193 TI - Protected on-duty time for theoretical learning. An experiment. PMID- 12123194 TI - NHS-ConNeCT breastfeeding workshops. PMID- 12123195 TI - Management through enthusiasm. Putting occupational health on the map. PMID- 12123196 TI - Competitive advantage. Implementing strategy in the NHS. PMID- 12123197 TI - Thinking outside the box. Encouraging flexible thinking for problems and decisions. PMID- 12123199 TI - Play your cards right. Mental health care at home. PMID- 12123198 TI - Making things better. Introducing clinical governance in a specialist palliative care unit. PMID- 12123201 TI - Approaches to quality in health and social care. PMID- 12123200 TI - Restoring the wholeness of being. PMID- 12123202 TI - Transformational leadership in nursing. Matching theory to practice. PMID- 12123203 TI - Expression of apoptosis-related genes in chronic cyclosporine nephrotoxicity in mice. AB - To define the mechanism of cyclosporine (CsA)-induced apoptosis, we investigated the expression of apoptosis-related genes in experimental chronic CsA nephrotoxicity. Mice on a low-salt (0.01%) diet were given vehicle (VH, olive oil, 1 mg/kg/day), or CsA (30 mg/kg/day), and sacrificed at 1 and 4 weeks. Apoptosis was detected with deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) stain, and the expressions of apoptosis-related genes were evaluated by reverse transcription-polymerase chain reaction, immunoblot or immunohistochemistry. The activity of caspase 1 and 3 was also evaluated. The CsA group showed increases in apoptotic cells compared with the VH group (54 +/- 41 vs. 3 +/- 3, p < 0.05), and the number of apoptotic cells correlated well with interstitial fibrosis scores (r = 0.83, p < 0.01). The CsA group showed a significant increase in Fas-ligand mRNA (0.20 vs. 0.02 amol/microgram total RNA, p < 0.05) and Fas protein expression (146% vs. 95%, p < 0.05), compared with the VH group. The CsA group showed significant increases in ICE mRNA (0.21 vs. 0.03 amol/microgram total RNA at 4 weeks, p < 0.05) and CPP32 mRNA (0.18 vs. 0.03 amol/microgram total RNA at 4 weeks, p < 0.05), compared with the VH group. The enzymatic activity of ICE (16.6 vs. 7.9 rho mol/microgram/h, p < 0.05) and CPP32 protease (15.6 vs. 2.7 rho mol/microgram/h, p < 0.05) proteases were increased in the CsA group, compared with the VH group. The ratio between bax and bcl-2 protein increased significantly in the CsA group (5.3-fold), compared with the VH group. Levels of p53 protein also increased in the CsA group. Immunohistochemical detection of Fas, Fas-ligand, ICE and CPP32 revealed strong immunoreactivity in renal tubular cells in areas of structural injury. These findings suggest that local activation of the apoptosis-related genes is associated with CsA-induced apoptotic cell death. PMID- 12123204 TI - Cold storage preservation and warm ischaemic injury to isolated arterial segments: endothelial cell injury. AB - Injury to endothelial cells is thought to be important to the development of the vascular lesion of chronic rejection. It was the aim of this study to develop a semiquantitative method to assess endothelial injury in arterial grafts and to document the injury produced by cold storage preservation and additional warm ischaemia. Twelve- and 24-h cold preservation of rat aortic segments, together with an additional 1 h of warm ischaemia, were assessed. Electron micrographs of representative endothelial cells were scored for cytoplasmic, nuclear and mitochondrial injury. The overall injury score was obtained by addition of the individual scores. Storage for up to 24 h in University of Wisconsin (UW) and Terasaki did not produce any injury. Twenty-four hours of storage in Euro-Collins resulted in endothelial cell death. Injury occurred after 12 h of storage in Ross, Collins and normal saline, and the injury increased following 24 h of storage. One hour of warm ischaemia did not increase the injury. Injury to endothelial cells varies with the preservation solution used and the time of cold storage, so that both the type of solution and the storage time should be taken into account in clinical studies looking at the influence of cold ischaemia time and graft outcome. PMID- 12123205 TI - Epithelial kinetics in mouse heterotopic tracheal allografts. AB - Obliterative bronchiolitis (OB) is the most important cause of graft dysfunction post-lung transplantation. It is likely that the small airway epithelium is a target of the alloimmune response, and that epithelial integrity is a crucial determinant of airway patency. Our goals are to elucidate epithelial cell kinetics in the heterotopic mouse trachea model and to determine potential mechanisms of cell death in allografts. Allografts and isografts were obtained by transplanting BALB/c tracheas into C57BL/6 and BALB/c immunosuppressed and non immunosuppressed hosts, respectively and harvested from day 3-20. Morphometry, BrdU and TUNEL labeling, and EM studies were performed. Columnar epithelium in isografts and allografts sloughs during day 0-3, but regenerates in both sets of grafts by day 10. Subsequently, allografts become inflamed and denuded, while isografts retain an intact epithelium. Prior to airway denudation, allografts exhibited significantly increased epithelial cell density, BrdU labeling index (LI), and TUNEL positive cells. Epithelial apoptosis was confirmed by electron microscopy. Allograft percent ciliated columnar epithelium and lumenal circumference were significantly decreased. Cyclosporin delayed airway fibrosis but did not alter the progression of the allograft through the phases of early ischemic injury, airway epithelial cell regeneration, and eventual cell death. These studies quantitatively demonstrate that the allograft epithelium actively regenerates in the alloimmune environment, but succumbs to increased apoptotic cell death, underscoring the importance of the airway epithelium as a self renewing source of alloantigen. PMID- 12123206 TI - Successful 24-h preservation of canine small bowel using the cavitary two-layer (University of Wisconsin solution/perfluorochemical) cold storage method. AB - We previously developed the two-layer cold storage method (TLM), which allows sufficient oxygen delivery to the canine pancreas during preservation, and successfully achieved 96-h preservation. In this study, we applied a modified TLM (cavitary TLM) to small bowel preservation in a canine heterotopic transplant model. Using simple storage in University of Wisconsin solution (UWM, group 1, n = 12) or cavitary TLM (group 2, n = 8), 40 cm segments of the jejunum were preserved for 24 h at 4 degrees C. The nonpreservation group served as the control (group 3, n = 8). The grafts were implanted heterotopically as a Thiry Vella loop. Eleven of 12 dogs in group 1 died within 3 days post-transplant as a result of graft intraluminal hemorrhage, while all dogs in groups 2 and 3 survived until day 7. Histological analyses showed almost normal structures of the graft mucosa in groups 2 and 3 at day 7. Results from maltose and acetaminophen absorption tests in group 2 were comparable to those in group 3. Only one survivor in group 1 showed distinct graft mucosal damage, confirmed by histological and functional analyses. In our transplant model, the canine small bowel was successfully preserved by cavitary TLM for at least 24 h, while this preservation time was beyond the limit with UWM. PMID- 12123207 TI - Treatment of parvovirus B-19 (PV B-19) infection allows for successful kidney transplantation without disease recurrence. AB - Parvovirus B 19 (PV-B19) has emerged as a cause of glomerulopathy in native and transplanted kidneys. Disease recurrence is less well described. The clinical and pathological spectrum of PV-B19 infection can be quite variable and therefore easily missed. There are no data regarding the safety of transplantation in PV B19-infected patients. We diagnosed a kidney transplant patient with recurrent PV B19 infection and collapsing glomerulopathy (CG) on renal biopsy. Retrospective analysis of stored serum showed that PV-B19 DNA was present prior to the transplant. The patient was treated with intravenous immunoglobulin (IVIG), and eventually the virus was successfully eradicated after allograft loss and discontinuation of immunosuppressive medications. The patient subsequently received her fourth kidney transplant. Polymerase chain reaction (PCR) for PV-B19 DNA was negative on multiple occasions for approximately 1 year prior to her transplant. The patient is now 22 months post transplantation, quarterly serum PCRs continue to be negative for PV-B19 DNA despite reinstitution of immunosuppressive therapy. The patient's renal function remains stable with a serum creatinine of 0.9 mg/dL and a serum albumin of 4.2 g/dL. Successful diagnosis and treatment of PV-B19 viremia appears to allow for successful transplantation without evidence of disease recurrence. Since PV-B19 can cause significant morbidity post transplant, it is important to screen potential transplant candidates at risk for PV-B19 infection pre transplant. Careful surveillance post transplantation is necessary to identify disease recurrence so that early treatment can be initiated. PMID- 12123208 TI - Late post-transplant anemia in adult renal transplant recipients. An under recognized problem? AB - Post-transplant anemia (PTA), a frequent complication during the first 3-6 months after transplant, is thought to be uncommon during the late post-transplant period. A study population of adults (> 18 years) transplanted during 1995 at Stanford University (n = 88) and University of North Carolina (n = 40) was selected. Data-collection points were 0, 1, 2, 3, 4 and 5 years post transplant. Anemia was defined as a hematocrit < 33 volume percentage. Thirty percent of patients were anemic at some time during the post-transplant period. The prevalence of PTA increased over time; by 5 years post transplant, 26% of the patients were anemic. Anemia occurred in 62.5% of patients converted from azathioprine to mycophenolate mofetil. A multivariate logistic regression model demonstrated a correlation between anemia and serum total CO2 (p = 0.002), BUN (p = 0.04), and creatinine (p = 0.045) at 1 year post transplant. At 5 years post transplant, only serum total CO2 (p = 0.0004) correlated with anemia. Thus, diminished renal excretory function and metabolic acidosis appear to be the most important correlates of late PTA. These findings should be interpreted in view of the fact that the newer immunosuppressive agents may have an even more profound effect on anemia and its recovery after transplantation. PMID- 12123210 TI - Hepatitis C-positive donors in heart transplantation. AB - Hepatitis C virus (HCV) can be transmitted to heart transplant recipients by donor organs. Mid-term results were reported using HCV-positive donors in patients at risk of imminent death (group I, n = 10), or in patients who otherwise would not have been offered heart transplantation (group II, n = 10) because of age (9/10) or associated medical risk (1/10). Medical records pertaining to patients receiving HCV-positive allografts between July 1994 and December 1999 were reviewed. The recipients consisted of 19 males and one female, with a median age of 54 years for group I and 66 for group II. The HCV RNA level, seroconversion of anti-HCV antibody, biochemical liver dysfunction, and causes of death were examined. Older recipients received reduced immunosuppression. Two patients in group II were HCV positive and were also retransplants. The hospital mortality rate was 10% in group I and 20% in group II; both hepatitis C-positive recipients died postoperatively prior to discharge. All predischarge deaths were related to multi-system organ failure (MSOF). All 17 survivors were HCV negative prior to transplant. Of these, 4/17 seroconverted. HCV RNA was detected in two of them. At a median follow-up of 26.4 months, 2/11 current survivors continue to test anti-HCV positive and are RNA negative. Three-year actual survival was 40% for group I and 70% in group II. Transplant coronary artery disease (TCAD) accounted for one postoperative death in group I. Current data show that four out of 11 survivors had developed TCAD at 3-year follow-up, yielding an actual freedom from TCAD rate of 12/17 (70%) at 3-year follow-up. Hepatitis C transmission using a donor heart as the reservoir is moderate (25%). Limited use of such donors is justified in selected patients. The risk for hepatic disease may be reduced by tailoring immunosuppression specifically for such recipients, particularly if they are at low risk of rejection. Further studies are necessary to define a possible association between HCV and TCAD. PMID- 12123209 TI - Sirolimus does not exhibit nephrotoxicity compared to cyclosporine in renal transplant recipients. AB - Sirolimus and cyclosporine (CsA) prevent acute rejection in man when used as primary therapies in triple drug regimens. Sirolimus does not act via the calcineurin pathway and therefore is not expected to produce the same renal side effects. This paper presents the pooled 2-year data analysis of renal function parameters from two open-label, randomized, multicenter studies. Patients (18-68 years) receiving a primary renal allograft were randomized to receive concentration-controlled sirolimus (n = 81) or CsA (n = 80), in combination with azathioprine and steroids (n = 83), or mycophenolate mofetil (MMF) and steroids (n = 78). From week 10 through year 2, calculated glomerular filtration rate (GFR) was significantly higher in sirolimus--than in CsA-treated patients (69.3 vs. 56.8 mL/min, at 2 years, p = 0.004). Serum uric acid was significantly higher in the CsA-treated patients and magnesium was significantly lower; these parameters were more likely to be within normal limits in the sirolimus group. Mean serum potassium and phosphorus were lower in sirolimus-treated patients. In conclusion, sirolimus, when administered as primary therapy in combination with azathioprine or MMF, has a favorable safety profile compared to CsA with regards to renal function. PMID- 12123211 TI - Expression of inducible and endothelial nitric oxide synthases, formation of peroxynitrite and reactive oxygen species in human chronic renal transplant failure. AB - Nitric oxide (NO.) is produced by NO synthases (NOS) and can interact with reactive oxygen species (ROS) to form peroxynitrite, which induces protein damage by formation of nitrotyrosine. NO. has a promotional effect on acute rejection. To investigate the role of NO. during chronic renal transplant failure (CRTF), we studied the expression of eNOS and iNOS in conjunction with H2O2 production and the formation of nitrotyrosines. Nephrectomy material from 10 patients and 10 control kidneys was used in this study. Expression of iNOS, eNOS, nitrotyrosine and the presence of ROS-producing cells and macrophages were determined using immunohistochemistry. INOS expression in nonsclerosed glomeruli and interstitium was significantly increased in patients with CRTF (p < 0.05). Glomerular eNOS expression was decreased in patients with CRTF compared with glomeruli of control kidneys (p < 0.01). Nitrotyrosine and ROS positive cells were significantly increased in CRTF in the interstitium (p < 0.05), but not in glomeruli. In summary, we found a marked interstitial increase in iNOS protein expression together with a decrease in glomerular eNOS expression in CRTF patients, associated with a significant increment in ROS and nitrotyrosine-positive cells in the interstitium. Our results suggest that loss of NO. production by glomerular eNOS in conjunction with an increased NO. production by interstitial iNOS, together with the formation of ROS and nitrotyrosine, is involved in the pathogenesis of CRTF. PMID- 12123212 TI - Long-term safety, tolerability and efficacy of daclizumab (Zenapax) in a two-dose regimen in liver transplant recipients. AB - A major thrust of transplantation research is to find more effective and less broadly toxic immunosuppressive agents. One potential way is the use of monoclonal antibodies directed to IL-2R alpha. Immunoprophylaxis with daclizumab, a humanized anti-IL-2R alpha monoclonal antibody, has been shown to be effective in the prevention of acute rejection in kidney transplant patients. These results encouraged us to initiate a pilot study in 28 liver transplant patients in 1997. Daclizumab was administered intravenously approximately 6 h after reperfusion (1 mg/kg) and on day 4 post-transplant (0.5 mg/kg). Additional immunosuppression consisted of cyclosporine A as well as of corticosteroids. Administration of daclizumab was not associated with any side-effects. We only experienced one acute rejection in a patient on day 17 post-transplant. It resolved immediately under therapy with prednisolone. The rate of opportunistic infections did not differ from results with conventional immunosuppressive regimens. At 4 years post transplant no lymphoproliferative disease was observed. Patient survival at 12, 24, 36 and 48 months post-transplant was 88.5, 84.6, 80.8 and 73.1%, respectively. Immunoprophylaxis with a two-dose daclizumab regimen is safe, effective and well tolerated, and does not lead to increased opportunistic infections. PMID- 12123213 TI - Interactions between cytomegalovirus, human herpesvirus-6, and the recurrence of hepatitis C after liver transplantation. AB - Recurrence of hepatitis C (HCV) following liver transplantation is common. Herpesvirus reactivation following transplant may have an immunomodulatory effect resulting in increased HCV replication. We studied whether cytomegalovirus (CMV) and human herpesvirus-6 (HHV-6) may be associated with HCV recurrence and viral load after transplant. We prospectively followed 66 HCV liver-transplant recipients with serial viral load testing for CMV and HHV-6. Infection and viral load were correlated with the development of biopsy-proven HCV recurrence and HCV viral loads. Histologic recurrence of HCV occurred in 41/66 (62.1%) patients. In the primary analysis, CMV infection and disease, and HHV-6 infection were not associated with HCV recurrence. Peak CMV and HHV-6 viral loads were not significantly different in patients with and without recurrence. No correlation was observed between HCV viral loads at 1 and 3 months post-transplant and peak HHV-6 or CMV viral loads. In a subgroup analysis, HHV-6 infection was associated with the development of more severe recurrence (hepatitis and/or fibrosis score > or = 2) (p = 0.01). Also, fibrosis scores at last follow up were higher in patients with CMV disease (1.67 vs. 0.56; p = 0.016) and in patients with HHV-6 infection (1.18 vs. 0.55; p = 0.031). In conclusion, HHV-6 and CMV infection and viral load were not associated with increased overall rates of HCV recurrence or HCV viral load after liver transplantation but may be associated with more severe forms of recurrence. PMID- 12123214 TI - Transient bone marrow edema in renal transplantation: a distinct post transplantation syndrome with a characteristic MRI appearance. AB - We describe four patients who developed severe knee pain within 3 months of renal transplantation. Plain radiographs were normal and inflammatory markers (CRP, ESR) were all within normal ranges. Magnetic resonance imaging (MRI) showed a distinctive pattern in all four cases of bone marrow signal changes, extending from the epiphyseal region into the metaphyseal region in two cases. The appearances were different from those of avascular necrosis (AVN) and reflex sympathetic dystrophy and showed no progression to develop AVN during the follow up period of 36 months. In all cases the pain resolved over a period of 3 months without specific therapy. Follow-up MRI scans were obtained in all patients after the pain had subsided, which revealed resolution of the MRI changes. We suggest that MRI be the investigation in such patients and that bone marrow edema changes will regress without the need to withdraw cyclosporin. PMID- 12123215 TI - Use of preserved vascular homografts in liver transplantation: hepatic artery aneurysms and other complications. AB - Hepatic artery aneurysms/pseudoaneurysms (HAAs) are rare but serious complications after orthotopic liver transplantation (OLT). Revascularization should accompany aneurysmectomy if possible and is more feasible if the aneurysm presents late after transplantation. The optimal conduits for revascularization in this situation are not known. Two patients with hepatic artery aneurysms/pseudoaneurysms who had aneurysmectomy and revascularization with third party cadaveric iliac arterial grafts 1 and 4 years after OLT are presented in detail, with an emphasis on the preservation method used for the grafts. Both livers were successfully revascularized with arterial grafts preserved for 21 and 26 days after procurement. Hepatic patency was documented in both 5 and 6 months after repair; graft function has remained normal 13 and 32 months after repair. Third-party vessels preserved for shorter periods have been used successfully in four other situations, including living-donor liver transplantation, and are briefly discussed. In conclusion, properly preserved vascular homografts are useful in LT for purposes other than initial vascular reconstruction. They also provide an excellent vascular conduit in recipients of livers from other (possibly living) donors. PMID- 12123216 TI - Mycobacterium haemophilum infections in heart transplant recipients: case report and review of the literature. AB - Non-tuberculous mycobacteria are becoming increasingly important pathogens among transplant recipients. We report a case of disseminated Mycobacterium haemophilum infection in a heart transplant recipient, manifesting as cellulitis, subcutaneous nodules, septic arthritis, and pneumonitis. Our case illustrates diverse challenges in the identification and treatment of this pathogen, such as its unique culture requirements and variable antimicrobial susceptibilities. Heightened clinical suspicion is necessary to establish a timely diagnosis so that optimal treatment can be administered. PMID- 12123217 TI - Polyomaviruses in kidney transplant recipients. PMID- 12123218 TI - Space life science experiments from Japan 1987-2000. PMID- 12123219 TI - Myrl Lua-Frances Ebert, October 20, 1913-May 5, 2001. PMID- 12123220 TI - NIDDK works on patient education. PMID- 12123221 TI - [Fructosemia]. PMID- 12123222 TI - V-1 Immunitor. PMID- 12123223 TI - Case presentation of a boy referred because of concerns about breathing during sleep. PMID- 12123224 TI - Obtaining a detailed family history for attention-deficit hyperactivity disorder is important. PMID- 12123227 TI - An 8-month old infant discovered to be blue and not breathing during sleep. PMID- 12123225 TI - Cause of abdominal pain in a 4-year old boy with cystic fibrosis. PMID- 12123226 TI - A 12-year old Greek girl with combined immunodeficiency syndrome. PMID- 12123231 TI - B-cell markers in lymphocyte predominance Hodgkin disease. PMID- 12123230 TI - [Congenital ear abnormalities]. PMID- 12123229 TI - [Objective olfactory tests using magnetoencephalography]. PMID- 12123232 TI - New prognostic parameters for chronic myelomonocytic leukemia. PMID- 12123233 TI - A valine deletion of ferroportin 1: a common mutation in hemochromastosis type 4. PMID- 12123235 TI - Association between the methylenetetrahydrofolate reductase 677C>T polymorphism and the risk of secondary lymphoproliferative disease in patients with a first idiopathic thrombosis. PMID- 12123234 TI - Leu208Val and Ile181Leu variants of cytochrome P450 CYP2C9 are not related to the acenocoumarol dose requirement in a Spanish population. PMID- 12123237 TI - JAMA patient page. HIV infection: the basics. PMID- 12123236 TI - Pregnancies after high-dose chemotherapy and autologous stem cell transplantation in aggressive lymphomas. PMID- 12123238 TI - Effectiveness of virtual reality for teaching pedestrian safety. AB - Sixty percent to 70% of pedestrian injuries in children under the age of 10 years are the result of the child either improperly crossing intersections or dashing out in the street between intersections. The purpose of this injury prevention research study was to evaluate a desktop virtual reality (VR) program that was designed to educate and train children to safely cross intersections. Specifically, the objectives were to determine whether children can learn pedestrian safety skills while working in a virtual environment and whether pedestrian safety learning in VR transfers to real world behavior. Following focus groups with a number of key experts, a virtual city with eight interactive intersections was developed. Ninety-five children participated in a community trial from two schools (urban and suburban). Approximately half were assigned to a control group who received an unrelated VR program, and half received the pedestrian safety VR intervention. Children were identified by group and grade by colored tags on their backpacks, and actual street crossing behavior of all children was observed 1 week before and 1 week after the interventions. There was a significant change in performance after three trials with the VR intervention. Children learned safe street crossing within the virtual environment. Learning, identified as improved street-crossing behavior, transferred to real world behavior in the suburban school children but not in the urban school. The results are discussed in relation to possibilities for future VR interventions for injury prevention. PMID- 12123239 TI - Virtual reality system for treatment of the fear of public speaking using image based rendering and moving pictures. AB - The fear of speaking is often cited as the world's most common social phobia. The rapid growth of computer technology enabled us to use virtual reality (VR) for the treatment of the fear of public speaking. There have been two techniques used to construct a virtual environment for the treatment of the fear of public speaking: model-based and movie-based. Virtual audiences and virtual environments made by model-based technique are unrealistic and unnatural. The movie-based technique has a disadvantage in that each virtual audience cannot be controlled respectively, because all virtual audiences are included in one moving picture file. To address this disadvantage, this paper presents a virtual environment made by using image-based rendering (IBR) and chroma keying simultaneously. IBR enables us to make the virtual environment realistic because the images are stitched panoramically with the photos taken from a digital camera. And the use of chroma keying allows a virtual audience to be controlled individually. In addition, a real-time capture technique was applied in constructing the virtual environment to give the subjects more interaction, in that they can talk with a therapist or another subject. PMID- 12123240 TI - Panic and agoraphobia in a virtual world. AB - Virtual reality (VR) offers a great new perspective on what it can offer an individual. These new approaches can give an individual the immersion and cognitive guidance that they need to help overcome his or her disorder. VR differs from the traditional displays in computer graphics as these various displays are integrated to give the user a sense of presence or immersion in the virtual world. To more effectively treat panic and agoraphobic patients using VR, it is necessary to determine the physiological responses of nonphobics when placed in the virtual panic and agoraphobia environments. This study exposed nonphobic participants to virtual panic and agoraphobia worlds with a program entitled "Virtual Medicine." Individuals without a diagnosis of panic and agoraphobia, as confirmed by intake and self-report questionnaires, were exposed to four different VR environments (elevator, supermarket, town square, and beach). During these VR experiences, physiology was measured by noninvasive sensors (peripheral skin temperature, heart rate, heart rate variability, respiration, and skin conductance). These measurements were compared to baseline physiology, which was recorded for five min prior to the VR exposure. These levels of physiological arousal will be useful in comparing against the phobic responses during virtual exposure. It will be useful to explore differences between immersion, physiological responses, and self-report responses in nonphobics versus phobics. PMID- 12123241 TI - Ethical codes and values in a virtual world. AB - Some members of the so-called virtual world deny that technology such as the virtual environment (VE) is value-laden. But there are a variety of complex issues which arise in VE that make it necessary to think about ethics and values in VE applications. Using VE in therapy and psychotherapy research or diagnostics leads to several ethical concerns. VE can impoverish those aspects of life that are essential to social development, interpersonal relations, and emotional growth. This paper will focus on the concept called "virtual world," a question of metaphysics. After reviewing general ethical principles, ethics in applied sciences will be described. Ethical decision-making, code of conduct, and specific issues in VE will be discussed. The VE community is challenged to set guidelines around VE, and its ethics and values. A set of measurement tools around ethical codes and values in a virtual world ought to be discursively gained in this virtual world and in real society as a whole. PMID- 12123242 TI - Virtual reality and haptics as an assessment device in the postacute phase after stroke. AB - Virtual reality (VR) technology is altering the health care environment and is changing the options that are available to therapists. This study describes how a haptic device was used as a cinematic assessment utility. Three chronic stroke inpatients at Sahlgrenska University Hospital with left hemisphere damage were assessed. The patients were administered by the box and block manual dexterity test. For comparisons, a reference group was added to the study. Several parameters, including time, speed, and movement of the right upper extremity, were extracted and evaluated. The results indicate that the system shows potential as an assessment device. The feasibility study setup is working well, as is the assessment method. Further research, testing, refinement of the exercises, and use of VR and haptics within neurological rehabilitation are suggested. PMID- 12123243 TI - Atypical face gaze in autism. AB - An eye-tracking study of face and object recognition was conducted to clarify the character of face gaze in autistic spectrum disorders. Experimental participants were a group of individuals diagnosed with Asperger's disorder or high functioning autistic disorder according to their medical records and confirmed by the Autism Diagnostic Interview-Revised (ADI-R). Controls were selected on the basis of age, gender, and educational level to be comparable to the experimental group. In order to maintain attentional focus, stereoscopic images were presented in a virtual reality (VR) headset in which the eye-tracking system was installed. Preliminary analyses show impairment in face recognition, in contrast with equivalent and even superior performance in object recognition among participants with autism-related diagnoses, relative to controls. Experimental participants displayed less fixation on the central face than did control-group participants. The findings, within the limitations of the small number of subjects and technical difficulties encountered in utilizing the helmet-mounted display, suggest an impairment in face processing on the part of the individuals in the experimental group. This is consistent with the hypothesis of disruption in the first months of life, a period that may be critical to typical social and cognitive development, and has important implications for selection of appropriate targets of intervention. PMID- 12123244 TI - Virtual reality for health care: the status of research. AB - As information technology has advanced and costs have declined over the past decade, there has been a steady growth in the use of virtual reality (VR) in health care. According to the data of the two leading clinical databases--MEDLINE and PSYCINFO--the research in the virtual reality field is moving fast: under the "virtual reality" keyword, there are 739 papers listed in MEDLINE and 569 in PSYCINFO (accessed 6 December 2001). Much of this growth, however, has been in the form of feasibility studies and pilot trials. In fact, many researchers tried to use VR, but only a few were able to deepen their study. According to MEDLINE, only 16 research groups published more than three papers related to health care applications of VR. This number lowers to 12 for papers included in PSYCLIT. Therefore, apart from surgical training and some behavioral treatments, there is little convincing evidence coming from controlled studies of the clinical and economical advantages of this approach. This paper discusses recent evidence and outlines some guidelines for future research in this area. PMID- 12123245 TI - Cultural differences in E-mail use of virtual teams: a critical social theory perspective. AB - This paper provides a perspective on cultural differences in e-mail use of virtual teams from the critical social theory (CST) point of view. CST can be used as a qualitative methodology in information systems research. Among CST concepts, Habermas' theory of communicative action is applied here for the purpose of drawing a plausible explanation for the phenomenon of cultural differences between East Asian countries and the West in the use of e-mail in virtual teams. East Asian countries such as Korea and Japan, which are heavily influenced by the Confucius tradition of emphasizing respect for the social order in all forms of social communication activity, including e-mail, provide different type of patterns of e-mail use in management of virtual teams compared to Western countries such as the United States. Not only can CST provide adequate theoretical support for the claim that e-mail use can be varied due to cultural difference when it is used to manage virtual teams, but it can also identify cultural codes such as respect for seniors as a part of the complete information package that can be illuminated during the use of e-mail, by using a CST concept called "critical reflection." A real-life example of the experience of a Korean firm's virtual team in the use of e-mail is analyzed in order to illustrate the CST perspective on cultural differences in the use of e-mail for virtual teams. PMID- 12123246 TI - Case study: public consensus building on the Internet. AB - Many groups or organizations advocating different special interests or policies now promote and conduct grassroots movements online through their own web sites. On the basic assumption that collective activities on the Internet are another form of social phenomenon, the process of public consensus-building on the Internet, and the ways in which various functions available on the Internet were strategically used to facilitate the public's involvement and participation were examined. The Frame Alignment Process was adopted as a conceptual framework for this study. Based on this framework, four related but not identical processes- "frame bridging," "frame amplification," "frame extension," and "frame transformation"--were elaborated, and various functions on the Internet facilitating this frame alignment process were examined. The Electronic Frontier Foundation's successful online campaign to prevent the passage of the Communication Decency Act of 1996 was chosen as a real-world case to elaborate a frame alignment process and the effective use of various functions on the Internet. PMID- 12123247 TI - Loneliness, self-disclosure, and ICQ ("I seek you") use. AB - This study investigated the relationships between self-disclosure in ICQ ("I seek you") chat, level of loneliness, and ICQ usage. The Revised UCLA Loneliness Scale and the Revised Self-Disclosure Scale (RSDS) were administered to a multistaged stratified random sample of 576 college students. The results indicate that loneliness is not related to level of ICQ use, but inversely related to valence, accuracy, and the amount dimensions of self-disclosure in ICQ chat, and that ICQ usage is significantly related to control of depth and intent of disclosure. Specifically, it was found that the lonelier the student, the more dishonest, more negative, and the less revealing was the quality of the self-disclosure in their ICQ interaction. Conversely, appropriate, honest, positive, and accurate self-disclosure might lead to decreased loneliness when one feels understood, accepted, and cared about on ICQ. More important, as intimate relationships are based on high degrees of depth and intent of self-disclosure, heavy users of ICQ are usually open, personal, and consciously aware of what they are disclosing. PMID- 12123248 TI - Methodological rigor with internet samples: new ways to reach underrepresented populations. AB - We present several rigorous methods for sampling difficult-to-reach and empirically underrepresented populations via the Internet. The methodology's representativeness was tested by comparing the demographics of a small sample of 82 lesbian and bisexual females with a much larger Gallup Organization sample of the general population (n > 1,000) obtained via random digit dialing. Compared to the latter poll, the rigorous sampling designs developed for the Internet were found to be significantly more robust and equally representative of the U.S. general population. The Gallup Organization reached a sample more representative of the age distribution of the United States. The Internet sample reached a sample more representative of the population, with less education, lower incomes, and a broad spectrum of ethnic diversity. The samples were equally effective in representing the distribution of the population with rural and urban residence. PMID- 12123249 TI - Bladder cancer incidence in the world. PMID- 12123250 TI - Gold for the NHS. What exactly is being bought with this gold? PMID- 12123251 TI - Consultants' new contract. So called victory in private practice obscures real contractual problems. PMID- 12123252 TI - Consultants' new contract. Private practice is unlikely to be main cause of long waiting lists. PMID- 12123253 TI - Consultants' new contract. Weak negotiators strike again. PMID- 12123254 TI - Consultants' new contract. New contract means cut in pay for part time consultants. PMID- 12123255 TI - Consultants' new contract. The negotiating committee should come up with a new agreement. PMID- 12123256 TI - Evaluation of studies of assessment and screening tools, and diagnostic tests. PMID- 12123257 TI - Chlorhexidine reduced catheter tip colonisation more than 10% povidone-iodine in critically ill neonates. PMID- 12123258 TI - A semidemand feeding protocol reduced time to full oral feeding in healthy preterm infants. PMID- 12123259 TI - Telephone based peer support increased duration of breast feeding in primiparous mothers. PMID- 12123260 TI - Review: behavioural interventions plus laxatives are effective for defecation disorders in children, but biofeedback does not add benefit. PMID- 12123261 TI - Inactivated influenza vaccine is safe for children and adults with stable asthma. PMID- 12123262 TI - Review: evidence does not support the effectiveness of over the counter preparations for acute cough. PMID- 12123263 TI - Verbal advice plus an information leaflet reduced antibiotic use in patients presenting with acute bronchitis. PMID- 12123264 TI - Review: evidence is incomplete on the benefits and risks of commonly used herbal medicines. PMID- 12123265 TI - Review: primary care counselling improves short term but not long term psychological symptoms in patients with psychological and psychosocial problems. PMID- 12123266 TI - Supportive expressive group therapy did not prolong survival in metastatic breast cancer. PMID- 12123267 TI - Effects of hormone therapy on health related quality of life in postmenopausal women with CAD differed according to presence of menopausal symptoms. PMID- 12123268 TI - A preoperative smoking intervention decreased postoperative complications in elective knee or hip replacement. PMID- 12123269 TI - Intravenous fluid replacement minimised dehydration during sodium picosulphate bowel preparation for colonic surgery. PMID- 12123270 TI - Review: music as a single session intervention reduces anxiety and respiratory rate in patients admitted to hospital. PMID- 12123271 TI - Review: longer bed rest does not prevent more postpuncture headaches than immediate mobilisation or short bed rest. PMID- 12123272 TI - Enteral nutrition reduced postoperative complications but increased minor adverse effects in gastrointestinal cancer. PMID- 12123273 TI - Review: breast cancer is associated with a family history of the disease in first degree relatives. PMID- 12123274 TI - A simple risk index accurately predicted mortality in patients with ST elevation myocardial infarction. PMID- 12123275 TI - Mothers of infants in neonatal nurseries had challenges in establishing feelings of being a good mother. PMID- 12123277 TI - People with enduring mental health problems described the importance of communication, continuity of care, and stigma. PMID- 12123276 TI - People at high risk for STDs used a variety of primary and secondary prevention strategies. PMID- 12123278 TI - Public health nurses used 4 strategies to facilitate client empowerment. PMID- 12123279 TI - Gate keeping and legitimisation were central in the interactions between informal carers of older people and healthcare workers. PMID- 12123280 TI - Older people perceived health as going and doing something meaningful. PMID- 12123281 TI - Kinetic analysis of histone acetylation turnover and Trichostatin A induced hyper and hypoacetylation in alfalfa. AB - Dynamic histone acetylation is a characteristic of chromatin transcription. The first estimates for the rate of acetylation turnover of plants are reported, measured in alfalfa cells by pulse, pulse-chase, and steady-state acetylation labeling. Acetylation turnover half-lives of about 0.5 h were observed by all methods used for histones H3, H4, and H2B. This is consistent with the rate at which changes in gene expression occur in plants. Treatment with histone deacetylase inhibitor Trichostatin A (TSA) induced hyperacetylation at a similar rate. Replacement histone variant H3.2, preferentially localized in highly acetylated chromatin, displayed faster acetyl turnover. Histone H2A with a low level of acetylation was not subject to rapid turnover or hyperacetylation. Patterns of acetate labeling revealed fundamental differences between histone H3 versus histones H4 and H2B. In H3, acetylation of all molecules, limited by lysine methylation, had similar rates, independent of the level of lysine acetylation. Acetylation of histones H4 and H2B was seen in only a fraction of all molecules and involved multiacetylation. Acetylation turnover rates increased from mono- to penta- and hexaacetylated forms, respectively. TSA was an effective inhibitor of alfalfa histone deacetylases in vivo and caused a doubling in steady state acetylation levels by 4-6 h after addition. However, hyperacetylation was transient due to loss of TSA inhibition. TSA-induced overexpression of cellular deacetylase activity produced hypoacetylation by 18 h treatment with enhanced acetate turnover labeling of alfalfa histones. Thus, application of TSA to change gene expression in vivo in plants may have unexpected consequences. PMID- 12123282 TI - Chromatin structure of eukaryotic promoters: a changing perspective. AB - Over the past few years, many studies have attempted to determine the role of nucleosomes as both positive and negative transcription regulators. The emphasis has mostly centered on chromatin remodeling activities and histone modifications, leaving the question of the influence of the higher-order structure out of the spotlight. Recent technical developments allowing direct measurements of size and mechanical properties of in vivo assembled chromatin may shed light on this poorly understood area. This article presents a brief summary of the current knowledge on transcription-dependent chromatin dynamics and how a rather simple agarose electrophoresis method may change the current view on structural changes linked to transcriptional activation of chromatin. PMID- 12123283 TI - The promyelocytic leukemia nuclear body: sites of activity? AB - The promyelocytic leukemia (PML) nuclear body is one of many subnuclear domains in the eukaryotic cell nucleus. It has received much attention in the past few years because it accumulates the promyelocytic leukemia protein called PML. This protein is implicated in many nuclear events and is found as a fusion with the retinoic acid receptor RARalpha in leukemic cells. The importance of PML bodies in cell differentiation and growth is implicated in acute promyelocitic leukemia cells, which do not contain PML bodies. Treatment of patients with drugs that reverse the disease phenotype also causes PML bodies to reform. In this review, we discuss the structure, composition, and dynamics that may provide insights into the function of PML bodies. We also discuss the repsonse of PML bodies to cellular stresses, such as virus infection and heat shock. We interpret the changes that occur as evidence for a role of these structures in gene transcription. We also examine the role of the posttranslational modification. SUMO-1 addition, in directing proteins to this nuclear body. Characterization of the mobility of PML body associated proteins further supports a role in specific nuclear events, rather than the bodies resulting from random accumulations of proteins. PMID- 12123284 TI - The elusive structural role of ubiquitinated histones. AB - It is increasingly apparent that histone posttranslational modifications are important in chromatin structure and dynamics. However, histone ubiquitination has received little attention. Histones H1, H3, H2A, and H2B can be ubiquitinated in vivo, but the most prevalent are uH2A and uH2B. The size of this modification suggests some sort of structural impact. Physiological observations suggest that ubiquitinated histones may have multiple functions and structural effects. Ubiquitinated histones have been correlated with transcriptionally active DNA, implying that it may prevent chromatin folding or help maintain an open conformation. Also, in some organisms during spermiogenesis, a process involving extensive chromatin remodeling, uH2A levels increase just prior to histone replacement by protamines. Determination of chromatin's structural changes resulting from histone ubiquitination is therefore important. Recent work using reconstituted nucleosomes and chromatin fibers containing uH2A indicate that in the absence of linker histones, ubiquitination has little structural impact. DNase I digests and analytical ultracentrifugation of reconstituted ubiquitinated nucleosomes show no structural differences. Solubility assays using reconstituted chromatin fibers in the presence of divalent ions demonstrate that uH2A fibers are slightly more prone to aggregation than controls, and analytical ultracentrifugation results with different MgCl2 and NaCl concentrations determined that chromatin folding is not affected by this modification. Additional work to assess possible synergistic affects with histone acetylation also precludes any structural implications. Protamine displacement experiments concluded that the presence of uH2A does not significantly affect the ability of the protamines to displace histones. In addition, uH2A does not interfere with histone H1 binding to the nucleosome. While work with uH2B remains insufficient to come to any definitive conclusions about its structural impact, current work with uH-2A indicates that, contrary to predictions, this histone modification does not affect either nucleosome or chromatin structure. Consequently, the search for a structural role for ubiquitinated histones continues and their effect on and importance in chromatin dynamics remains elusive. PMID- 12123285 TI - A feel for the template: zinc finger protein transcription factors and chromatin. AB - Transcription factors and chromatin collaborate in bringing the eukaryotic genome to life. An important, and poorly understood, aspect of this collaboration involves targeting the regulators to correct binding sites in vivo. An implicit and insufficiently tested assumption in the field has been that chromatin simply obstructs most sites and leaves only a few functionally relevant ones accessible. The major class of transcription factors in all metazoa, zinc finger proteins (ZFPs), can bind to chromatin in vitro (as clearly shown for Spl, GATA-1 and -4, and the nuclear hormone receptors, for example). Data on the accessibility of DNA within heterochromatin to nonhistone regulators (E.A. Sekinger and D.S. Gross. 2001. Mol. Cell 105: 403-414; C. Jolly et al. 2002. J. Cell. Biol. 156: 775-781) and the ability of the basal transcription machinery to reside within highly condensed chromatin (most recently, R. Christova and T. Oelgeschlaeger. 2002. Nat. Cell Biol. 4: 79-82) further weaken the argument that chromatin acts as an across-the-board deterrent to ZFP binding. These proteins, however, do not bind promiscuously in vivo, and recent data on human cells (C.E. Horak et al. 2002. Proc. Natl. Acad. Sci. U.S.A. 99: 2924-2929) confirm earlier data on budding yeast (B. Ren et al. 2000. Science (Washington, D.C.), 290: 2306-2309) that primary DNA sequence, i.e., density of binding sites per unit DNA length, is not the primary determinant of where a ZFP transcription factor will bind in vivo. This article reviews these data and uses ZFP transcription factors as a model system to compare in vitro binding to chromatin by transcription factors with their in vivo behavior in gene regulation. DNA binding domain structure, nonrandom nucleoprotein organization of chromatin at target promoters, and cooperativity of regulator action may all contribute to target site selection in vivo. PMID- 12123286 TI - The estrogen receptor: more than the average transcription factor. AB - The human estrogen receptor is a steroid nuclear receptor found in breast cancer and a variety of other tissues. Located in the nucleus, it can exist either loosely or tightly associated with the nuclear matrix depending on whether or not it is bound to ligand. When bound to ligand, it is responsible for the transcriptional regulation of estrogen-responsive genes through recruitment of coactivators and corepressors of transcription. The estrogen receptor is also capable of ligand-independent transcriptional activation via the mitogen activated protein kinase pathway. Ligands have been implicated in the regulation of estrogen receptor levels via changing the levels and stability of estrogen receptor mRNA and protein. The resulting levels of estrogen receptor and the type of ligand bound to it have a direct impact on the transcription of estrogen responsive genes. PMID- 12123287 TI - Chromatin remodeling and tissue-selective responses of nuclear hormone receptors. AB - Chromatin structure of eukaryotic genes regulates gene expression by controlling the accessibility of regulatory factors. To overcome the inhibitory nature of chromatin, protein complexes that modify higher order chromatin organization and histone-DNA contacts are critical players in regulating transcription. For example, nuclear hormone receptors regulate transcription by interacting with ATP dependent chromatin-remodeling complexes and coactivators, which include histone acetyltransferases and histone methylases that modify the basic residues of histones. A growing number of tissue-specific nuclear hormone receptor ligands, termed "selective modulators", owe their specificity, at least in part, to the differential recruitment of these chromatin-modifying coactivators. The molecular mechanisms by which these compounds modulate the functions of nuclear hormone receptors are discussed here. PMID- 12123288 TI - Protamine-like proteins: evidence for a novel chromatin structure. AB - Protamine-like (PL) proteins are DNA-condensing proteins that replace somatic type histones during spermatogenesis. Their composition suggests a function intermediate to that of histones and protamines. Although these proteins have been well characterized at the chemical level in a large number of species, particularly in marine invertebrates, little is known about the specific structures arising from their interaction with DNA. Speculation concerning chromatin structure is complicated by the high degree of heterogeneity in both the number and size of these proteins, which can vary considerably even between closely related species. After careful examination and comparison of the protein sequences available to date for the PL proteins, we propose a model for a novel chromatin structure in the sperm of these organisms that is mediated by somatic type histones, which are frequently found associated with these proteins. This structure supports the concept that the PL proteins may represent various evolutionary steps between a sperm-specific histone H1 precursor and true protamines. Potential post-translational modifications and the control of PL protein expression and deposition are also discussed. PMID- 12123289 TI - Dynamics of histone acetylation in vivo. A function for acetylation turnover? AB - Histone acetylation, discovered more than 40 years ago, is a reversible modification of lysines within the amino-terminal domain of core histones. Amino terminal histone domains contribute to the compaction of genes into repressed chromatin fibers. It is thought that their acetylation causes localized relaxation of chromatin as a necessary but not sufficient condition for processes that repackage DNA such as transcription, replication, repair, recombination, and sperm formation. While increased histone acetylation enhances gene transcription and loss of acetylation represses and silences genes, the function of the rapid continuous or repetitive acetylation and deacetylation reactions with half-lives of just a few minutes remains unknown. Thirty years of in vivo measurements of acetylation turnover and rates of change in histone modification levels have been reviewed to identify common chromatin characteristics measured by distinct protocols. It has now become possible to look across a wider spectrum of organisms than ever before and identify common features. The rapid turnover rates in transcriptionally active and competent chromatin are one such feature. While ubiquitously observed, we still do not know whether turnover itself is linked to chromatin transcription beyond its contribution to rapid changes towards hyper- or hypoacetylation of nucleosomes. However, recent experiments suggest that turnover may be linked directly to steps in gene transcription, interacting with nucleosome remodeling complexes. PMID- 12123290 TI - Molecular evolutionary relationships between partulid land snails of the Pacific. AB - Adaptive radiation of partulid land snails in the tropical Pacific has produced an extraordinary array of distinctive morphological, ecological and behavioural types. Here we use part of the nuclear ribosomal RNA gene cluster to investigate the relationships within and between the three partulid genera, Partula, Samoana and Eua. The genera cluster separately, with Samoana and Partula forming monophyletic groups. With one exception, the molecular data generally support the previous generic classification based on genital morphology, even in species that show a number of characteristics otherwise atypical of the genus. Convergent evolution explains morphological similarities between members of different genera. The phylogeny suggests that Samoana has colonized the Pacific from west to east, originating in the area where Eua, believed to be the most ancient partulid genus, is found. An unexplained anomaly is the reported occurrence of a single species of Samoana in the Mariana Islands of the western Pacific. The genus Partula has a disjunct distribution, encompassing islands both to the east and west of the range occupied by Eua. Partula seems to have spread both eastward and westward after the splitting of the Partula lineage. PMID- 12123291 TI - Locomotion through use of the mouth brushes in the larva of Culex pipiens (Diptera: Culicidae). AB - The larva of the mosquito Culex pipiens is a filter-feeder and is able to use the feeding current generated by its mouth brushes to glide slowly through the water. The hydrodynamics of the mouth brushes, and of gliding, were investigated by visualizing the feeding current using dyes. Unlike the mouth brushes of a sessile filter-feeder such as the blackfly larva, those of C. pipiens function more like 'paddles' than 'rakes', a beneficial adaptation to life in still as opposed to running water. Technically, the Froude efficiency of gliding is very low (0.23) because the design of the feeding brushes favours delivery of water into the wake rather than forward momentum to the body. The wider implications of these findings to foraging strategy and other aspects of the behaviour of mosquito larvae are discussed. PMID- 12123292 TI - Mechanical properties of nacre and highly mineralized bone. AB - We compared the mechanical properties of 'ordinary' bovine bone, the highly mineralized bone of the rostrum of the whale Mesoplodon densirostris, and mother of pearl (nacre) of the pearl oyster Pinctada margaritifera. The rostrum and the nacre are similar in having very little organic material. However, the rostral bone is much weaker and more brittle than nacre, which in these properties is close to ordinary bone. The ability of nacre to outperform rostral bone is the result of its extremely well-ordered microstructure, with organic material forming a nearly continuous jacket round all the tiny aragonite plates, a design well adapted to produce toughness. In contrast, in the rostrum the organic material, mainly collagen, is poorly organized and discontinuous, allowing the mineral to join up to form, in effect, a brittle stony material. PMID- 12123294 TI - Agricultural intensification and the collapse of Europe's farmland bird populations. AB - The populations of farmland birds in Europe declined markedly during the last quarter of the 20th century, representing a severe threat to biodiversity. Here, we assess whether declines in the populations and ranges of farmland birds across Europe reflect differences in agricultural intensity, which arise largely through differences in political history. Population and range changes were modelled in terms of a number of indices of agricultural intensity. Population declines and range contractions were significantly greater in countries with more intensive agriculture, and significantly higher in the European Union (EU) than in former communist countries. Cereal yield alone explained over 30% of the variation in population trends. The results suggest that recent trends in agriculture have had deleterious and measurable effects on bird populations on a continental scale. We predict that the introduction of EU agricultural policies into former communist countries hoping to accede to the EU in the near future will result in significant declines in the important bird populations there. PMID- 12123293 TI - The timing of life-history events in a changing climate. AB - Although empirical and theoretical studies suggest that climate influences the timing of life-history events in animals and plants, correlations between climate and the timing of events such as egg-laying, migration or flowering do not reveal the mechanisms by which natural selection operates on life-history events. We present a general autoregressive model of the timing of life-history events in relation to variation in global climate that, like autoregressive models of population dynamics, allows for a more mechanistic understanding of the roles of climate, resources and competition. We applied the model to data on 50 years of annual dates of first flowering by three species of plants in 26 populations covering 4 degrees of latitude in Norway. In agreement with earlier studies, plants in most populations and all three species bloomed earlier following warmer winters. Moreover, our model revealed that earlier blooming reflected increasing influences of resources and density-dependent population limitation under climatic warming. The insights available from the application of this model to phenological data in other taxa will contribute to our understanding of the roles of endogenous versus exogenous processes in the evolution of the timing of life history events in a changing climate. PMID- 12123295 TI - When is now? Perception of simultaneity. AB - We address the following question: Is there a difference (D) between the amount of time for auditory and visual stimuli to be perceived? On each of 1000 trials, observers were presented with a light-sound pair, separated by a stimulus onset asynchrony (SOA) between -250 ms (sound first) and +250 ms. Observers indicated if the light-sound pair came on simultaneously by pressing one of two (yes or no) keys. The SOA most likely to yield affirmative responses was defined as the point of subjective simultaneity (PSS). PSS values were between -21 ms (i.e. sound 21 ms before light) and +150 ms. Evidence is presented that each PSS is observer specific. In a second experiment, each observer was tested using two observer stimulus distances. The resultant PSS values are highly correlated (r = 0.954, p = 0.003), suggesting that each observer's PSS is stable. PSS values were significantly affected by observer-stimulus distance, suggesting that observers do not take account of changes in distance on the resultant difference in arrival times of light and sound. The difference RTd in simple reaction time to single visual and auditory stimuli was also estimated; no evidence that RTd is observer specific or stable was found. The implications of these findings for the perception of multisensory stimuli are discussed. PMID- 12123296 TI - Self-perceived attractiveness influences human female preferences for sexual dimorphism and symmetry in male faces. AB - Exaggerated sexual dimorphism and symmetry in human faces have both been linked to potential 'good-gene' benefits and have also been found to influence the attractiveness of male faces. The current study explores how female self-rated attractiveness influences male face preference in females using faces manipulated with computer graphics. The study demonstrates that there is a relatively increased preference for masculinity and an increased preference for symmetry for women who regard themselves as attractive. This finding may reflect a condition dependent mating strategy analogous to behaviours found in other species. The absence of a preference for proposed markers of good genes may be adaptive in women of low mate value to avoid the costs of decreased parental investment from the owners of such characteristics. PMID- 12123297 TI - Will tuberculosis become resistant to all antibiotics? AB - The discovery of high prevalences of antibiotic resistance in some pathogens, in some parts of the world, has provoked fears of a widespread loss of drug efficacy. Here, we use a mathematical model to investigate the evolution of resistance to four major anti-tuberculosis drugs (isoniazid, rifampicin, ethambutol and streptomycin) in 47 sites around the world. The model provides a new method of estimating the relative risk of treatment failure for patients carrying drug-resistant strains and the proportion of patients who develop resistance after failing treatment. Using estimates of these two quantities together with other published data, we reconstructed the epidemic spread of isoniazid resistance over the past 50 years. The predicted median prevalence of resistance among new cases today was 7.0% (range 0.9-64.3%), close to the 6.3% (range 0-28.1%) observed. Predicted and observed prevalences of resistance to isoniazid plus rifampicin (multidrug-resistant or MDR-TB) after 30 years of combined drug use were also similar, 0.9% (0.1-5.9%) and 1.0% (range 0-14.1%), respectively. With current data, and under prevailing treatment practices, it appears that MDR-TB will remain a localized problem, rather than becoming a global obstacle to tuberculosis control. To substantiate this result, further measurements are needed of the relative fitness of drug-resistant strains. PMID- 12123298 TI - The probability of severe disease in zoonotic and commensal infections. AB - Cross-species transfers of pathogens (zoonoses) cause some of the most virulent diseases, including anthrax, hantavirus and Q fever. Zoonotic infections occur when a pathogen moves from its reservoir host species into a secondary host species. Similarly, commensal infections often have a primary reservoir location within their hosts' bodies from which they rarely cause disease symptoms, but commensals such as Neisseria meningitidis cause severe disease when they cross into a different body compartment from their normal location. Both zoonotic and commensal infections cause either mild symptoms or severe disease, but rarely intermediate symptoms. We develop a mathematical model for studying three factors that affect the probability of severe disease: the size of the inoculum, the route of inoculation and the frequency of naturally occurring infections that do not cause symptoms but do induce protective immunity (vaccinating inoculations). With a single route of infection, increasing pathogen density causes inoculations to develop more often into disease rather than asymptomatic vaccinations that provide protective immunity. With two routes of infection, it may happen that a lower density of a pathogen or of a particular antigenic variant leads to a relatively higher frequency of disease-inducing versus vaccinating inoculations. This reversal occurs when one route of infection tends to vaccinate against relatively common pathogens but less often vaccinates against relatively rare pathogens, whereas the other route of infection is susceptible to disease inducing inoculation even at relatively low pathogen density. PMID- 12123299 TI - The social evolution of bacterial pathogenesis. AB - Many of the genes responsible for the virulence of bacterial pathogens are carried by mobile genetic elements that can be transferred horizontally between different bacterial lineages. Horizontal transfer of virulence-factor genes has played a profound role in the evolution of bacterial pathogens, but it is poorly understood why these genes are so often mobile. Here, I present a hypothetical selective mechanism maintaining virulence-factor genes on horizontally transmissible genetic elements. For virulence factors that are secreted extracellularly, selection within hosts may favour mutant 'cheater' strains of the pathogen that do not produce the virulence factor themselves but still benefit from factors produced by other members of the pathogen population within a host. Using simple mathematical models, I show that if this occurs then selection for infectious transmission between hosts favours pathogen strains that can reintroduce functional copies of virulence-factor genes into cheaters via horizontal transfer, forcing them to produce the virulence factor. Horizontal gene transfer is thus a novel mechanism for the evolution of cooperation. I discuss predictions of this hypothesis that can be tested empirically and its implications for the evolution of pathogen virulence. PMID- 12123300 TI - Indirect fitness consequences of mate choice in sticklebacks: offspring of brighter males grow slowly but resist parasitic infections. AB - 'Good genes' models of sexual selection suggest that elaborate male sexual ornaments have evolved as reliable signals of male quality because only males of high genetic viability are able to develop and maintain them. Females benefit from choosing such individuals if quality is heritable. A key prediction is that the offspring of males with elaborate mating displays will perform better than those of less elaborate males, but it has proved difficult to demonstrate such an effect independently of the effects of differences in parental investment. We tested for 'good genes' linked to male ornamentation in the three-spined stickleback Gasterosteus aculeatus using in vitro fertilization to generate maternal half-siblings, which were raised without parental care. Maternal half siblings sired by brightly coloured males grew less quickly than half-siblings sired by dull males but were more resistant to a controlled disease challenge. Among the offspring that became infected, those with brighter fathers had higher white blood cell counts. This suggests that highly ornamented males confer disease resistance on their offspring. The association with reduced growth suggests a mechanism for the maintenance of heritable variation in both disease resistance and male sexual coloration. PMID- 12123301 TI - Antagonistic pre- and post-copulatory sexual selection on male body size in a water strider (Gerris lacustris). AB - A crucial question in sexual selection theory is whether post-copulatory sexual selection reinforces or counteracts conventional pre-copulatory sexual selection. Male body size is one of the traits most generally favoured by pre-copulatory sexual selection; and recent studies of sperm competition often suggest that large male size is also favoured by post-copulatory sexual selection. In contrast to this general pattern, this study shows that pre- and post-copulatory sexual selection act antagonistically on male body size in Gerris lacustris. One large and one small male were kept together with two females in this experiment. Large males had a significant mating advantage, but small males copulated longer and gained higher fertilization success from each mating. Large and small males, however, gained similar reproductive success, and there was no overall correlation between mating success and reproductive success. These results suggest that estimates of male fitness based solely on mating success should be viewed with caution, because of potentially counteracting post-copulatory selection. PMID- 12123302 TI - Reversible frequency-dependent switches in male mate choice. AB - Current sexual-selection theories predict that mating should occur preferentially with the highest-quality partner, and assume that for distinguishing among potential mates the choosy sex applies an internal representation of the characteristics of the desired mate, i.e. a template. Binary choice experiments were performed to test male mate choice between two different female colour morphs in the damselfly Ischnura elegans. Choice experiments were conducted before and after an habituation period, during which males were exposed to only one female colour morph. Given the choice between the two female morphs, males did exhibit a choice for the most recently experienced female morph. This is the first evidence for a reversible switch in mate choice in a frequency-dependent way. In contrast with previous studies on mate choice, template formation in male I. elegans seems not to be based on quality. Switching mate choice in a frequency dependent manner, choosing the most common morph, probably allows males to minimize their search efforts and to maximize fitness. PMID- 12123303 TI - Males call more from wetter nests: effects of substrate water potential on reproductive behaviours of terrestrial toadlets. AB - Laboratory studies of terrestrial-breeding frogs have demonstrated that wetter substrates produce fitter offspring but the relevance of substrate wetness to adult reproductive strategies is unknown. I hypothesized that male toadlets (Pseudophryne bibronii) would select wetter areas for nesting and would advertise wet nests strongly, and tested these predictions by manipulating water potentials at a breeding site. Males preferred to nest in the wettest areas, and called at greater rates on almost twice as many nights as males occupying drier nests. Overall, males that mated called on significantly more nights than unmated males. Hence, because males occupying wet nests called more, they also mated more and in 19 out of 20 cases, oviposition occurred in wet nests that were suitable for embryonic development. Males occupying drier nests may have risked dehydration by calling, and so were less able to signal to females. Hydration states therefore have the potential to influence the reproductive success of terrestrial male frogs. PMID- 12123304 TI - The effects of diet and physiological stress on the evolutionary dynamics of an enzyme polymorphism. AB - In the northern acorn barnacle Semibalanus balanoides, polymorphism at the mannose-6-phosphate isomerase (Mpi) locus appears to be maintained by distinct selection regimes that vary between intertidal microhabitats. The goal of the present experiment was to elucidate the mechanism of selection at the Mpi locus by examining the relationship between genotype and fitness-related life-history traits in laboratory manipulations. When barnacles were cultured on a mannose supplemented diet and exposed to thermal stress, different Mpi genotypes exhibited differences in the rate of growth that predicted survivorship. In contrast, no such relationship was observed in control or fructose-supplemented dietary treatments either in the presence or in the absence of stress. Similarly, the phenotype and survivorship of genotypes at another allozyme locus and a presumably neutral mitochondrial DNA marker were homogeneous across all treatments and unaffected by experimental manipulations. These results suggest that the differential survivorship of Mpi genotypes in the field and laboratory results from a differential ability to process mannose-6-phosphate through glycolysis. The widespread polymorphism at Mpi observed in marine taxa may reflect the interaction between dietary composition and environmental heterogeneity in intertidal habitats. PMID- 12123305 TI - Dispersal costs set the scene for helping in an atypical avian cooperative breeder. AB - The ecological constraints hypothesis is suggested to explain the evolution of cooperative breeding in birds. This hypothesis predicts that the scene for cooperative breeding is set when ecological factors constrain offspring from dispersal. This prediction was tested in the atypical cooperative breeding system of the long-tailed tit, Aegithalos caudatus, by comparing the degree of philopatry and cooperation in an isolated and a contiguous site whilst experimentally controlling for confounding aspects of reproduction. No difference was found between the two sites in the survival of offspring but a greater proportion were found to remain philopatric in the isolated site. This difference was caused by greater philopatry of normally dispersive females suggesting, as predicted, that dispersal costs were greater from this site. Furthermore, a greater proportion of males and females cooperated following breeding failure in the isolated site than in the contiguous site. Thus, as has been suggested for typical avian cooperative breeders, dispersal costs, relative to philopatric benefits, appear to set the scene for cooperative breeding in long-tailed tits. PMID- 12123306 TI - Assessment of left ventricular diastolic function with Doppler tissue imaging: effects of preload and place of measurements. AB - Mitral inflow velocities are widely used for the evaluation of left ventricular (LV) diastolic function. However, they are closely affected by other factors such as preload. The purpose of this study was to evaluate the usefulness of tissue Doppler velocities obtained from the mitral annulus for the evaluation of ventricular relaxation in patients under different loading conditions. We also evaluated the effect of preload at different sides on the mitral annulus. The study population consisted of 62 consecutive patients (38 male, 24 female with a mean age of 42 +/- 13 years) who have undergone hemodialysis. Both mitral inflow velocities (E wave, A wave, E wave deceleration time and isovolumetric relaxation time) and mitral annulus tissue Doppler velocities (E', A') from the septal, lateral, anterior, posterolateral and inferior sides of the mitral annulus were measured immediately before and after hemodialysis. Mitral inflow E and A wave velocities and E/A ratio decreased significantly (p < 0.001, p = 0.007, p < 0.001, respectively) after hemodialysis. Mitral annulus E' wave velocities and E'/A' ratios obtained from five different sides of the annulus also changed significantly (p < 0.001 for all); however, there was no change in the A' wave velocity (p > 0.05 for all) after hemodialysis. The decrease in E wave and E/A ratio in mitral inflow measurements and E' velocities and E'/A' ratios in tissue Doppler measurements were correlated with the amount of fluid extracted (for mitral inflow E wave, r = 0.392, p = 0.002 and E/A ratio, r = 0.280 and p = 0.027; for lateral side E', r = 0.329, p = 0.009 and E'/A' ratio, r = 0.286, p = 0.04; for septal side E', r = 0.376, p = 0.003 and E'/A' ratio, r = 0.297, p = 0.019; for anterior side E', r = 0.342, p = 0.007 and E'/A' ratio, r = 0.268, p = 0.035; for posterolateral side E', r = 0.423, p = 0.001 and E'/A' ratio, r = 0.343, p = 0.007; and for inferior side E', r = 0.326, p = 0.01 and E'/A' ratio, r = 0.278, p = 0.029). We conclude that mitral annular velocities obtained by tissue Doppler are preload dependent parameters for the evaluation of LV diastolic function. PMID- 12123307 TI - Determination of left ventricular mass by three-dimensional echocardiography: in vitro validation of a novel quantification method using multiple equi-angular rotational planes for rapid measurements. AB - Measuring left ventricular mass by m-mode echocardiography or two-dimensional echocardiography is limited by the fact that calculations are based on assumptions, which describe left ventricular shape by simple geometric figures. The ability of three-dimensional echocardiography (3-DE) to accurately assess left ventricular mass has been shown previously, but 3-DE approaches to quantitative analysis of ventricular mass required multiple tomographic sectioning, manual tracing in various cut planes and were time consuming and laborious. We investigated the accuracy of a novel, rapid method of 3-DE mass quantification using multiple rotational planes in left ventricles in vitro. METHODS: Three-dimensional data sets of 10 fixed pig hearts were obtained using a TomTec 3-DE system. For 3-DE mass calculations, a rotational axis in the center of the ventricle (apical-basal orientation) was defined and 3, 6 and 12 equi angular rotational planes were created. The endocardial and epicardial contour of the left ventricle was traced in each cut plane and the volume of the corresponding myocardial wedge was automatically calculated. Mass was calculated by multiplying the resulting myocardial volume by the specific weight of myocardial tissue. The measurements were performed by two investigators blinded to the anatomic true mass and were analyzed for interobserver and intraobserver variability. RESULTS: The anatomic left ventricular mass was measured 73-219 (168 +/- 50) g. 3-DE mass ranged from 88-247 (207 +/- 51) g (three planes), 84-250 (205 +/- 52) g (six planes) and 86-241 (202 +/- 50) g (12 planes) respectively. The correlation between 3-DE mass and anatomic LV mass measurements (r = 0.92) and between two observers (r = 0.97-0.98) was good. True mass was slightly overestimated by 3-DE measurement (SEE = 22-23 g). The intraobserver and interobserver variabilities were < or = 4 and < or = 7% respectively for all measurements. CONCLUSION: This new 3-DE method of left ventricular mass quantification with rotational approach provides accurate and reproducible measurements. In normal shaped left ventricles even three planes were sufficient to provide accurate mass measurements in vitro. PMID- 12123308 TI - Between observer variation is not eliminated by standardised analysis of dobutamine-atropine stress echocardiography. AB - AIMS: The conventional analysis of dobutamine-atropine stress echocardiography (DASE) is poorly defined and subject to considerable variation. The aim of this study was to investigate the reproducibility of strictly standardised qualitative analysis in DASE. METHODS AND RESULTS: Strict criteria for standardised DASE interpretation were defined through logistic regression analysis on categorical parameters obtained from 20 patients with coronary artery disease (CAD) and 20 healthy controls subjected to DASE. Three expert echocardiographers analysed DASE recordings from 100 consecutive patients referred for coronary angiography. Specificity for CAD and for predicting significant stenosis of a major coronary artery was 94% (95% CI: 83-100%) and 79% (95% CI: 63-96%), whereas sensitivity was 49% in both cases (95% CI: 38-60% and 37-61%). Within and between observer reproducibility was moderate to fair (kappa = 0.56 and 0.38; 95% CI: 0.40-0.72 and 0.24-0.52). In patients without prior myocardial infarction and in echogenic patients within observer reproducibility was good (kappa = 0.72 and 0.74; 95% CI: 0.52-0.92 and 0.56-0.92). CONCLUSIONS: Observer variation was not eliminated in standardised qualitative DASE interpretation based on criteria that predicted the presence of CAD with a high specificity and reproducibility was good only in certain subgroups of patients. PMID- 12123309 TI - Echocardiographic assessment of left atrioventricular plane displacement as a complement to left ventricular regional wall motion evaluation in the detection of myocardial dysfunction. AB - AIM: We aimed to find out if abnormal left atrioventricular plane displacement (AVPD) is a sign of myocardial dysfunction, even in patients with normal left ventricular (LV) regional wall motion (RWM). METHODS: We prospectively performed echocardiography in 1350 consecutive patients referred to our echocardiography laboratory. Left AVPD and LV RWM were evaluated in all patients. We prospectively selected all patients with normal LV RWM but impaired left AVPD for further analysis of clinical parameters. RESULTS: Eighty-eight of the 1350 patients had completely normal LV RWM but impaired left AVPD (< or = 10 mm) in at least one region (septal, lateral, posterior, anterior). Of these, 60.2% had prior and/ or acute myocardial infarction, predominantly non-Q-wave, whereas 33.0% had angina without infarction and 2.3% had hypertension. In 49 (55.7%) patients coronary angiography was performed. All were abnormal. In 4.5% (n = 4) of the patients no obvious reason for the AVPD decrease was found, but was not precluded. CONCLUSION: Almost all patients with abnormal left AVPD and completely normal LV RWM had clinical cardiac disease. Thus, decreased AVPD despite normal LV RWM seems to be a true sign of myocardial dysfunction, predominantly indicating subendocardial dysfunction. In screening for patients with myocardial dysfunction assessment of left AVPD may be useful as a complement to LV RWM evaluation. The prognosis in such patients is currently being evaluated. PMID- 12123310 TI - Reproducibility of the ventricular synchronization parameters assessed by multiharmonic phase analysis of radionuclide angiography in the normal heart. AB - Radionuclide angiography (RNA) permits analysis of contractility and conduction abnormalities. We determined the parameters of normal ventricular synchronization, assessed the reproducibility of the technique, and compared first harmonic (1H) and third harmonic (3H) analysis. Forty-four normal subjects (28 men and 16 women) were studied. RNA was performed in left anterior oblique (LAO) and left lateral (LL) projections. The onset (To), mean time (Tm), total contraction time (Tt) for right ventricle (RV) and left ventricle (LV), interventricular time (T(RV-LV) = Tm(LV - Tm(RV)) in LAO, and the apex-to-base time (T(a-b)) in LL were measured on the histograms of the time-activity curve. Reproducibility (R) was tested by studying 26 consecutive patients with two successive RNAs. RV starts contracting 25 ms before LV (To(RV) = 29 +/- 37 ms; To(LV) = 54 +/- 39 ms; mean +/- SD) with a 37 ms longer total contraction time. T(RV-LV) is 3 +/- 16 ms. In LL projection, apex and base contract synchronously: T(a-b) = 2 +/- 16 ms. 3H analysis enlarges all duration parameters (To, Tm and Tt), but does not alter synchronization (deltaT(a-b) and deltaT(RV-LV) between 1H and 3H <1%, p = NS). Reproducibility of the duration (T(tLV) and T(tRv)) and synchronization parameters (T(a-b) and T(RV-LV)) is high (R < or = 2.2%). In conclusion, the simultaneous contraction of right and left ventricles and of apex and base can be quantified by RNA phase analysis with high reproducibility. These results, consistent with published electrophysiological data, provide the basis for further non-invasive investigations of ventricular resynchronization in patients with basal electrical or mechanical asynchrony. PMID- 12123311 TI - Relationship between exercise-induced ST segmental depression and myocardial ischemia assessed by technetium-99m tetrofosmin SPECT imaging in patients with inferior Q wave myocardial infarction. AB - BACKGROUND: ST segment depression (STD) is a standard electrocardiographic sign of myocardial ischemia. Although STD may represent reciprocal changes in patients with previous myocardial infarction, studies of reciprocal changes during exercise testing are scarce. METHODS: From December 1999 to December 2000, 160 patients (119 males, 41 females, mean age 54 +/- 8 years), undergoing, maximal or symptom-limited exercise treadmill test (Bruce-protocol), myocardial perfusion scintigraphy using technetium-99m tetrofosmin single photon emission computed tomography (SPECT) imaging, within 30 days of an uncomplicated inferior Q wave myocardial infarction. The location of STD at the electrocardiogram (ECG) was defined as anterior (V1-4), high lateral (I, aVL), and lateral (V5-6). Ischemia was defined as reversible perfusion abnormalities. RESULTS: STD occurred in anterior leads in 29 patients (18.1%), in the lateral leads in 41 patients (25.6%), in the high lateral leads in 20 patients (12.5%). In 70 patients (43.8%) no significant STD occurred during the exercise test. ST segment elevation occurred in 28 patients (17.5%) in inferior leads. High lateral STD was associated with inferior ST elevation in 16 patients (80%), whereas only eight patients (19.5%) with lateral STD and nine patients (31%) with anterior STD were associated with inferior ST elevation. Ischemia was detected in 63 of 90 patients (70%) with and in 10 of 70 patients (14.3%) without STD (p < 0.0001). Patients with high lateral STD had a higher prevalence of fixed perfusion defects in the inferior wall (95 vs. 27.8%) and in posterolateral wall (75 vs. 18.9%) compared with other patients (p = 0.003 and 0.002, respectively). Ischemia was more prevalent in patients with lateral STD than without (87.8 vs. 14.3%, p < 0.0001). CONCLUSION: In patients with inferior Q wave, the presence of exercise-induced STD in lateral and anterior leads appears to be a sign of myocardial ischemia, and may require invasive evaluation; on the other hand, the presence of STD in high lateral leads should be recognized as a reciprocal change for ST elevation in the inferior leads, and may not be an indication for invasive evaluation. PMID- 12123312 TI - Magnetic resonance imaging of coronary artery occlusions in the navigator technique. AB - Non-invasive assessment of coronary arteries is possible with magnetic resonance imaging (MRI). Respiratory gated MR coronary angiography is a new imaging technique that permits reconstruction of the coronary arteries based on a three dimensional (3D) data set obtained from the free-breathing patient. In this study, respiratory gated MR angiography (MRA) was performed to assess coronary artery occlusions. MRI was performed in 25 patients who had been referred for conventional coronary angiography because of suspected coronary artery disease. Coronary artery occlusion was evaluated in the proximal and middle vessel segments after multiplanar coronary reconstruction of the MR images. Five patients were excluded from the study; in the remaining 20 patients 120 coronary artery segments were analyzed. Good image quality could be obtained for 85% of the segments. Eighteen of the 24 occlusions were confirmed by MRI, the overall sensitivity was 75% and the specificity was 100%. The best results were found in the proximal left anterior descending (LAD) and descending parts of the right coronary artery (RCA), where all occlusions were confirmed. These results showed that coronary artery occlusions can be detected in the proximal and middle LAD and RCA using 3D respiratory gated MRA. Further technical improvements, especially in spatial resolution, are necessary before MRA can become a reliable diagnostic tool in the non-invasive evaluation of coronary arteries. PMID- 12123314 TI - Can cardiac magnetic resonance imaging reliably differentiate between benign and neoplastic fat? AB - Magnetic resonance imaging is currently frequently used to differentiate lipomatous hypertrophy of the interatrial septum from other cardiac lesions involving abnormal fatty tissue including lipomatous neoplasms. This report discusses potential problems with this approach by emphasizing the variable appearance of lipomatous hypertrophy of the interatrial septum on magnetic resonance imaging. PMID- 12123313 TI - The quantification of pulmonary valve haemodynamics using MRI. AB - The optimum slice location within the pulmonary root to quantify pulmonary valve haemodynamics was examined using magnetic resonance (MR) phase velocity mapping. MRI was carried out on 15 patients with congenital aortic valve disease. Although the patients had aortic valve disease, all measurements were made on the pulmonary valve. Systolic (Q(SYS)) and diastolic (Q(DIAS)) blood flow volume and cardiac index (CI) were determined at four pulmonary artery locations. The change in diastolic flow volume relative to slice 1, closest to the pulmonary valve, was also calculated. For a change in axial position of 1.5 cm, i.e. from 0.5 to 2 cm from the annulus, there was a change in diastolic flow volume of 4.4 ml. There was a significant increase in the mean diastolic flow from 3.4 to 7.7 ml (p = 0.01 between slice positions 0.5 and 2 cm. However, there was no significant change in CI, 3.4-3.7 l/min/m2 (p = 0.14) over the same distance. We believe that two factors are responsible for these results. The first is that of compliance, whose effects can be minimized by placing the MR slice close to the valve, however, this will not account for the second factor, being that of valve motion, and hence diastolic pulmonary valve flow or regurgitant volume will be underestimated. The degree of underestimation may only be important at mild and moderate levels of regurgitation or if changes in regurgitation are to be temporally measured. PMID- 12123315 TI - High prevalence of tetrofosmin reverse redistribution pattern in patients with myocardial infarction and angiographically smooth coronary arteries. Published in voL 18/1, pp. 31-40. PMID- 12123316 TI - Advanced contour detection for three-dimensional intracoronary ultrasound: a validation--in vitro and in vivo. AB - Intracoronary ultrasound (ICUS) provides high-resolution transmural images of the arterial wall. By performing a pullback of the ICUS transducer and three dimensional reconstruction of the images, an advanced assessment of the lumen and vessel wall morphology can be obtained. To reduce the analysis time and the subjectivity of boundary tracing, automated segmentation of the image sequence must be performed. The Quantitative Coronary Ultrasound-Clinical Measurement Solutions (QCU-CMS) (semi)automated analytical software package uses a combination of transversal and longitudinal model- and knowledge-guided contour detection techniques. On multiple longitudinal sections through the pullback stack, the external vessel contours are detected simultaneously, allowing mutual guidance of the detection in difficult areas. Subsequently, luminal contours are detected on these longitudinal sections. Vessel and luminal contour points are transformed to the individual cross-sections, where they guide the vessel and lumen contour detection on these transversal images. The performance of the software was validated stepwise. A set of phantoms was used to determine the systematic and random errors of the contour detection of external vessel and lumen boundaries. Subsequently, the results of the contour detection as obtained in in vivo image sets were compared with expert manual tracing, and finally the contour detection in in vivo image sequences was compared with results obtained from another previously validated ICUS quantification system. The phantom lumen diameters were underestimated by 0.1 mm, equally by the QCU-CMS software and by manual tracing. Comparison of automatically detected contours and expert manual contours, showed that lumen contours correspond very well (systematic and random radius difference: -0.025 +/- 0.067 mm), while automatically detected vessel contours slightly overestimated the expert manual contours (radius difference: 0.061 +/- 0.037 mm). The cross-sectional vessel and lumen areas as detected with our system and with the second computerized system showed a high correlation (r = 0.995 and 0.978, respectively). Thus, use of the new QCU-CMS analytical software is feasible and the validation data suggest its application for the analysis of clinical research. PMID- 12123317 TI - Left ventricular outflow tract presystolic flow velocity--another marker of left ventricular diastolic function. AB - Left ventricular outflow tract (LVOT) presystolic flow velocities were studied using pulse doppler echocardiography in 30 normal persons. Thirty patients of mild hypertension with transmitral flow velocity pattern suggestive of impaired relaxation were also studied. Transmitral flow velocity pattern was correlated with LVOT presystolic flow velocities in the two groups. Hypertensive patients had significantly higher transmitral A wave velocity (p < 0.001) and significantly lower transmitral E wave/A wave velocity ratio (p < 0.001) as compared to normal group. LVOT presystolic flow velocities had significant direct correlation with transmitral A wave velocity (p < 0.01) and significant inverse relation with transmitral E wave/A wave velocity ratio (p < 0.05). Our observations suggest that increased LVOT presystolic flow peak velocity can also be used as another marker of impaired left ventriCular compliance during atrial contraction. More work is needed to establish exact status of this preliminary observation. PMID- 12123318 TI - Myocardial contrast echocardiography for predicting functional recovery after acute myocardial infarction. AB - Myocardial contrast echocardiography (MCE) is a promising diagnostic tool for detecting microvascular integrity. The aim of the study was to investigate the comparative specificity and sensitivity of intravenous MCE, technetium-99m Sestamibi single-photon emission computed tomography (SPECT) and dipyridamole dobutamine (DIDO) stress echocardiography for predicting functional recovery after coronary revascularization in patients with acute myocardial infarction (AMI). METHODS: In a prospective, observational study, 17 consecutive patients short after AMI who received successful treatment with primary percutaneous coronary angioplasty (PTCA) plus stent-implantation were examined with DIDO (dipyridamole with 0.28 mg/kg over 4 min plus dobutamine up to 10 mcg/kg/min), MCE (10 ml 4 g, 400 mg/ml Levovist intravenously; second harmonic power imaging) within 12-24 h and resting perfusion SPECT within 48-72 h after PTCA. Functional recovery of regional contractile function after 6-month follow-up was the gold standard to assess viability. RESULTS: The rate of agreement between SPECT and MCE was 69% and between SPECT and a positive response to stress echo was 76% for combined DIDO. MCE showed a higher sensitivity (96%) in the identification of viability than SPECT (77%) and combined DIDO alone (79%). Specificity was lower for viability recognition with MCE (58%) compared with SPECT (93%) and DIDO (87%). CONCLUSIONS: The wall motion response during DIDO echocardiography is useful in the prediction of recovery of regional and global ventricular function after revascularization in patients after AMI. Combined intravenous MCE and DIDO is more accurate in the diagnosis of stunned myocardium than Tc-99m-MIBI SPECT alone. PMID- 12123319 TI - Adenosine myocardial perfusion single photon emission computed tomographic stress testing 24-72 h after uncomplicated myocardial infarction. AB - Safety of performing adenosine myocardial perfusion stress testing as early as 24 h after acute uncomplicated myocardial infarction is not known. We evaluated 31(14 females and 17 males, average age 72, range 46-89 years) consecutive patients with uncomplicated myocardial infarction, who underwent adenosine myocardial perfusion stress imaging, 24-72 h after infarction for risk stratification. Adenosine was infused at a rate of 140 microg/kg/min for 6 min. Twenty patients were presented with non-ST-elevation myocardial infarction. Eleven patients were admitted with acute ST-elevation myocardial infarction. Patients were monitored for signs of complication during and immediately after the stress test. The average time from admission to performance of stress tests was 51 +/- 19 h, ranging from the minimum of 24 h to maximum 72 h. No complications related to adenosine infusion were detected. In conclusion, our data suggest that a further large study of early adenosine myocardial perfusion SPECT imaging may be safe in a carefully selected group of patients after uncomplicated myocardial infarction. PMID- 12123320 TI - The incidence of discrepant regional myocardial uptake between 201 thallium and 123 I-BMIPP SPECT in patients with coronary heart disease. AB - Myocardial perfusion and fatty acid uptake at rest were assessed by SPECT with 201Tl (Tl) and 123I-BMIPP (BMIPP) in 50 consecutive patients with coronary heart disease. Discrepant regional myocardial uptake was observed in 19 patients and classified into the following two groups: mismatch (MM; Tl uptake > BMIPP uptake, n = 14, mean age, 66 years) and paradoxical mismatch (PM; Tl uptake < BMIPP uptake, n = 5, mean age, 68 years). In the MM group, 77% was single- or zero vessel disease and the artery-perfused region in the mismatched area was almost always ischemia related. Sixty percent of the regions observed with the PM were related to the inferior wall. In the PM group, 80% of cases were associated with multivessel stenoses and 60% of cases was suffered from ischemic attack within a week before scintigraphy. In conclusion, mismatch was related to abnormal fatty acid uptake caused by coronary heart disease. Although the paradoxical mismatch might mainly be related to diaphragmatic attenuation of Tl scans and augmented artifacts of BMIPP scans in the inferior wall, we should not overlook severe coronary heart disease in patients with paradoxical mismatched phenomenon. PMID- 12123321 TI - Perfusion-BMIPP mismatch: specific finding or artifact? PMID- 12123322 TI - Contrast-enhanced three-dimensional MR angiography of the pulmonary vascular tree. AB - Contrast-enhanced three-dimensional MR angiography has evolved into a promising technique in the study of the pulmonary vasculature. Both congenital and acquired entities can be now morphologically demonstrated in a non-invasive manner obviating the need for conventional pulmonary angiography. Due to spatial resolution limitations, however, it is still premature to routinely apply the method in the detection of small subsegmental emboli, in cases of suspected pulmonary embolism, and further technical developments will be required. In this paper we present a spectrum of congenital and acquired disorders affecting the pulmonary vascular tree as demonstrated with contrast-enhanced three-dimensional MR angiography. PMID- 12123323 TI - Determination of coronary calcium with multi-slice spiral computed tomography: a comparative study with electron-beam CT. AB - Electron-beam Computed Tomography (EBCT) has been used for years to quantify coronary artery calcification as a marker of coronary atherosclerosis. The aim of this study was to determine the diagnostic accuracy of a new scanner, the Multi slice Spiral CT (MSCT), for the assessment of coronary calcification and to compare this new technique to EBCT. The study population consisted of 99 male patients, aged 60 +/- 10 years with suspected or known coronary artery disease. With EBCT 40 axial slices, ECG-triggered (scan time = 100 ms, slice thickness = 3 mm), were acquired in one breath-hold (35 +/- 5 s). For MSCT simultaneous acquisition of four axial slices (scan time = 250 ms, slice thickness = 2.5 mm), allowed the entire heart (48 slices) to be covered in one breath-hold of 25 +/- 5 s. For quantification of coronary calcium the Volumetric Calcium Score (VCS) was calculated. There was an excellent correlation for the VCS (r = 0.994, p = 0.01, mean difference = 97 +/- 115) between both scanners. Comparison of low (1-100), moderate (101-400), high (401-1000) and very high score values (>1,000) showed no significant differences. The number of calcified lesions and densities were statistically not different. Mean variability of the two scans was 17%. The MSCT scanner is equivalent to EBCT for the determination and quantification of coronary calcium and can therefore be used for calcium screening. With application of the spiral mode technique further improvement in variability can be expected, thus allowing for follow-up studies to determine progression or regression of atherosclerosis with high accuracy. PMID- 12123324 TI - Visualization of atrial excitation by magnetocardiogram. AB - We tried to visualize normal atrial excitatory process by magnetocardiogram (MCG) measured from both anterior chest and back using our newest multi-channel SQUID system. Twenty normal subjects were studied. After measuring the normal (B(z)) component of the magnetic field, we constructed an isomagnetic and vector arrow map from spatial derivatives of the normal (B(z)) component in the tangential direction. By the MCG measurement from the anterior chest, current arrows were recognized in the right upper portion which were directed to the left lower from the beginning of the P wave to the P-wave peak. By the measurement from the back, current arrows were able to be visualized in the right to middle upper portion which were directed to the left or left lower just before the P-wave peak. We conclude that we successfully recognized the right atrial excitation and its spread to the left atrium and observed the time course of normal atrial excitatory process by the MCG measurement from not only anterior chest but also back using 64-channel SQUID system. PMID- 12123325 TI - Assisted suicide, euthanasia, mercy killing. PMID- 12123326 TI - How important are estimates of cancer prevalence? PMID- 12123327 TI - Pride and judgement: the Annals of Oncology prizes. PMID- 12123328 TI - Frequency and risk factors of subclinical cardiotoxicity after anthracycline therapy in children: a systematic review. AB - BACKGROUND: The aim of this systematic review was to summarise and appraise the published evidence with regard to the frequency and risk factors of subclinical cardiotoxicity in apparently healthy survivors of childhood cancer after anthracycline therapy. PATIENTS AND METHODS: A search was made in Medline for studies published between 1966 and May 2001 that included >50 children and reported on the frequency of measures of subclinical cardiotoxicity. Information about the studies was abstracted by two reviewers and a validity score was calculated for each study. RESULTS: The reported frequency of subclinical cardiotoxicity varied between 0% and 57% in the 25 studies included. Differences in outcome definitions of subclinical cardiotoxicity and differences in study patients with respect to the dose of anthracycline seemed to explain part of the wide variance of the frequency of subclinical cardiotoxicity. Fourteen of the 25 studies showed serious methodological limitations. CONCLUSIONS: The reported frequency of subclinical cardiotoxicity shows a wide variation. Well designed studies with accurate and precise outcome measurements in well described groups of patients, after a sufficiently long follow-up period, are needed to obtain more insight into the frequency and importance of risk factors, and the clinical consequences of anthracycline-related subclinical cardiotoxicity. PMID- 12123329 TI - Measuring cancer prevalence in Europe: the EUROPREVAL project. AB - Cancer prevalence is the proportion of individuals in a population who at some stage during their lifetime have been diagnosed with cancer, irrespective of the date of diagnosis. Cancer prevalence statistics have generally been provided by a limited number of well established cancer registries that have been in existence for several decades. The advent of systematic follow-up of life status of incident cases and the availability of new statistical methodologies, now makes it possible for registries established during the 1970s or 1980s to provide prevalence data. The main problems encountered in the estimation of prevalence are the inclusion of: (i) cases lost to follow-up; (ii) cases known only from their death certificate; (iii) cases diagnosed before the start of registration; and (iv) the treatment of multiple tumours and migrations. The main aim of this paper was to review these problems and discuss, through the experience gained with EUROPREVAL, how they can be overcome. A method is presented for the calculation of prevalence of all cancers combined in the populations covered by the 45 cancer registries participating in EUROPREVAL. Prevalence of cancer is estimated to be 2% on average, with the highest values (3%) in Sweden and the lowest in Eastern Europe, with a minimum of approximately 1% in Poland. PMID- 12123330 TI - Cancer prevalence in European registry areas. AB - BACKGROUND: Information on cancer prevalence is of major importance for health planning and resource allocation. However, systematic information on cancer prevalence is largely unavailable. MATERIALS AND METHODS: Thirty-eight population based cancer registries from 17 European countries, participating in EUROPREVAL, provided data on almost 3 million cancer patients diagnosed from 1970 to 1992. Standardised data collection and validation procedures were used and the whole data set was analysed using proven methodology. The prevalence of stomach, colon, rectum, lung, breast, cervix uteri, corpus uteri and prostate cancer, as well as of melanoma of skin, Hodgkin's disease, leukaemia and all malignant neoplasms combined, were estimated for the end of 1992. RESULTS: There were large differences between countries in the prevalence of all cancers combined; estimates ranged from 1170 per 100000 in the Polish cancer registration areas to 3050 per 100000 in southern Sweden. For most cancers, the Swedish, Swiss, German and Italian areas had high prevalence, and the Polish, Estonian, Slovakian and Slovenian areas had low prevalence. Of the total prevalent cases, 61% were women and 57% were 65 years of age or older. Cases diagnosed within 2 years of the reference date formed 22% of all prevalent cases. Breast cancer accounted for 34% of all prevalent cancers in females and colorectal cancer for 15% in males. Prevalence tended to be high where cancer incidence was high, but the prevalence was highest in countries where survival was also high. Prevalence was low where general mortality was high (correlation between general mortality and the prevalence of all cancers = -0.64) and high where gross domestic product was high (correlation = +0.79). Thus, the richer areas of Europe had higher prevalence, suggesting that prevalence will increase with economic development. CONCLUSIONS: EUROPREVAL is the largest project on prevalence conducted to date. It has provided complete and accurate estimates of cancer prevalence in Europe, constituting essential information for cancer management. The expected increases in prevalence with economic development will require more resources; allocation to primary prevention should therefore be prioritised. PMID- 12123331 TI - Addition of either irinotecan or methotrexate to bolus 5-fluorouracil and high dose folinic acid every 2 weeks in advanced colorectal carcinoma: a randomised study by the Southern Italy Cooperative Oncology Group. AB - PURPOSE: The purpose of this study was to compare the activity and toxicity of the combination of irinotecan (IRI) plus folinic acid (FA)-modulated 5 fluorouracil (5-FU) i.v. bolus with a regimen of double modulation of 5-FU with methotrexate (MTX) and FA in patients with advanced colorectal carcinoma. PATIENTS AND METHODS: Two-hundred and thirty-four patients were enrolled: 118 patients received IRI 200 mg/m2 (90-min i.v. infusion) on day 1, followed by levo FA 250 mg/m2 (2-h i.v. infusion) and 5-FU 850 mg/m2 (i.v. bolus) on day 2 (IRIFAFU), and 116 patients received MTX 750 mg/m2 (2-h i.v. infusion) on day 1, followed by levo-FA 250 mg/m2 (2-h i.v. infusion) and FU 800 mg/m2 (i.v. bolus) on day 2 (MTXFAFU). Both cycles were repeated every 2 weeks until progression or to a maximum of 16 cycles. Response rate (RR) was the main end point of the study; responses were assessed every four cycles and confirmed after 2 additional months of treatment. RESULTS: RR was significantly greater with IRIFAFU (36%) than with MTXFAFU (20%) (P <0.001). Multivariate analysis showed that IRIFAFU was significantly associated with a greater activity (P = 0.028). Median progression free survival was longer with IRIFAFU than with MTXFAFU (7.2 months compared with 4.8 months; P = 0.048). Median survival time (MST) did not differ between the two arms (14.7 months compared with 14.8 months, respectively). Patients not receiving second-line chemotherapy, however, lived longer when treated in the first-line with IRIFAFU (MST 11.9 months compared with 6.4 months; P = 0.038). IRIFAFU caused a significantly greater occurrence of grade 3 or 4 neutropenia (40% compared with 9%; P = 0.001) and diarrhoea (13% compared with 4%; P = 0.024), but a significantly lower incidence of stomatitis (3% compared with 12%; P = 0.007), than the comparative regimen. CONCLUSIONS: IRIFAFU appeared comparable in terms of activity and toxicity with other weekly or biweekly bolus or infusional combination regimens. IRIFAFU, however, seems easier to administer, because it does not require infusional catheter or pump devices, and it is less expensive. It may represent a new option for treating advanced colorectal carcinoma. PMID- 12123332 TI - A three-arm phase III randomised trial comparing combinations of platinum derivatives, ifosfamide and/or gemcitabine in stage IV non-small-cell lung cancer. AB - OBJECTIVE: To determine, in stage IV non-small-cell lung cancer (NSCLC), if the combination of gemcitabine-a new active drug-with ifosfamide (IG) or with the cisplatin-carboplatin association (CCG) will improve survival (primary end point) in comparison with a first-generation regimen, cisplatin-carboplatin-ifosfamide (CCI). PATIENTS AND METHODS: A total of 284 chemotherapy-naive patients with metastatic NSCLC were randomised. Four were ineligible and 16 were not assessable for responses. Cisplatin was given at 60 mg/m2 on day 1, carboplatin AUC 3 mg.min/ml on day 1, ifosfamide 4.5 g/m2 on day 1 and gemcitabine 1 g/m2 on days 1, 8 and 15. Courses were repeated every 4 weeks. Response was assessed after three courses and chemotherapy was continued in responding patients until best response. There were 94 eligible patients in the CCI arm, 92 in CCG and 94 in the IG arm. RESULTS: The objective response rates for CCI, CCG and IG were 23% [95% confidence interval (CI) 15% to 32%], 29% (95% CI 20% to 39%) and 25% (95% CI 16% to 33%), respectively ( P = 0.61). Median survival time was 24, 34 and 30 weeks, respectively (P = 0.20). One-year survival was 23, 33 and 35%, and 2-year survival was 11, 14 and 17%, respectively. In some subgroups (older patients, women), there was a significant survival advantage for CCG and IG compared with CCI. Toxicity was tolerable: severe alopecia was less frequent in the CCG arm, and IG was associated with significantly more thrombopenia while CCG was associated with more leucopenia. CONCLUSION: In stage IV NSCLC, treatment with regimens including the new drug gemcitabine were associated with a better but not statistically significant observed survival compared with a classical first generation cisplatin-containing regimen. The non-platinum combination of gemcitabine was as effective as its combination with platinum. PMID- 12123333 TI - Intramuscular depot medroxyprogesterone versus oral megestrol for the control of postmenopausal hot flashes in breast cancer patients: a randomized study. AB - BACKGROUND: Hot flashes are frequent in postmenopausal breast cancer patients, especially when treated with tamoxifen. Estrogen replacement therapy is the most effective treatment for hot flashes, but its use is controversial in breast cancer survivors. Progestins may offer a good alternative for the control of hot flashes in this setting; in particular, oral megestrol acetate has been proven effective in a randomized, placebo-controlled clinical trial. With the aim of further improving these results, we have designed a randomized study comparing oral megestrol acetate with depot intramuscular (i.m.) medroxyprogesterone acetate (MPA) for the control of hot flashes in postmenopausal patients with a history of breast cancer. PATIENTS AND METHODS: Seventy-one postmenopausal patients were randomized to receive an i.m. injection of depot MPA 500 mg on days 1, 14 and 28, or oral megestrol acetate 40 mg daily for 6 weeks. Patients recorded daily the number and severity of their hot flashes; response was defined as a > or =50% decrease in the number and severity of hot flashes. RESULTS: At week 6, hot flashes were reduced by 86% on average in the whole group of patients, without significant differences between the two progestins. Response was obtained by 75 and 67% of patients receiving MPA or megestrol, respectively (P = 0.5). Responders were followed to assess maintenance of response (without further treatment), which was significantly better with i.m. MPA: in this group, 89% of responders still showed a benefit at week 24, compared with 45% in the megestrol group (P = 0.03). CONCLUSIONS: Our study shows that a short cycle of i.m. depot MPA injections provides significant and long-lasting relief from postmenopausal hot flashes in patients with a history of breast cancer, offering an alternative to estrogen replacement therapy or prolonged administration of oral megestrol. PMID- 12123334 TI - A randomised phase II study of conventional versus accelerated infusional chemotherapy with granulocyte colony-stimulating factor support in advanced breast cancer. AB - BACKGROUND: Granulocyte colony-stimulating factor (G-CSF) allows cycles of conventional bolus chemotherapy to be accelerated with reduction in treatment time and a boost in dose intensity. Theoretically, this approach could be hazardous with infusional 5-fluorouracil (5-FU) chemotherapy, since G-CSF stimulated neutrophil proliferation would be occurring in the face of continuous S-phase active 5-FU. We performed this phase II randomised study to compare the safety, tolerability and efficacy of conventional 3-weekly epirubicin, cyclophosphamide and continuous infusional 5-FU (infusional ECF) to an accelerated 2-weekly schedule with G-CSF support, in patients with advanced breast cancer. PATIENTS AND METHODS: Twenty-seven patients were randomised. with 14 in the accelerated arm. Patients received bolus epirubicin 60 mg/m2 and cyclophosphamide 600 mg/m2 every 3 weeks (conventional arm) or every 2 weeks (accelerated arm) and 5-FU 200 mg/m2/day continuous infusion throughout. G-CSF 300 microg/day s.c. on days 10-12 was given each accelerated cycle. RESULTS: There were no treatment delays secondary to inadequate neutrophil or platelet recovery in either arm, with higher median day 1 neutrophil counts for each cycle in the accelerated arm compared with the conventional arm. Eighty-six per cent of the planned conventional chemotherapy cycles and 82% of the planned accelerated cycles were given. There were no major differences in toxicity between the arms, with the most common grade 3 toxicities being alopecia and stomatitis. Eight patients developed neutropenic sepsis (five in the accelerated arm and three in the conventional arm). Ten patients (77%) responded in the conventional arm and nine (64%) in the accelerated arm. CONCLUSIONS: Accelerated infusional ECF with limited G-CSF support is a feasible and well-tolerated regimen with rapid haematological recovery. A 50% increase in relative dose intensity of epirubicin and cyclophosphamide is achieved, while overall treatment time is reduced by 33%. PMID- 12123335 TI - Recognizing features that are dissimilar in male and female breast cancer: expression of p21Waf1 and p27Kip1 using an immunohistochemical assay. AB - BACKGROUND: Male breast cancer (MBC) is an uncommon disease, and most of our current knowledge of its biology, natural history and treatment has been extrapolated from data on the disease in women. Information is still needed on the molecular biological properties of male breast tumors and their predictive relevance. Kinase inhibitor proteins (KIPs) p27Kip1 and p21Waf1 negatively regulate cell cycle progression by preventing the passage of cycling cells from G1 to S phase through G1 cyclin-dependent kinase activation. No studies exist on the role of these factors in male breast carcinoma. PATIENTS AND METHODS: We have retrospectively analyzed the immunohistochemical expression of p21Waf1 and p27Kip1 protein in 27 primary MBC and in 101 female breast cancers (FBC) treated at the European Institute of Oncology between 1997 and 2000. We also assessed sex hormone receptors status, p53, bcl-2 and c-erb-B2 protein expression, and Ki-67 labeling index. RESULTS: We observed a statistically significant difference in the immunostaining of KIPs p27Kip1 and p21Waf1 in male patients compared with females. Expression of p21Waf1 was observed in 19 of the 27 (70.3%) primary MBCs versus 29 of 101 FBC (29%). Fourteen of 22 negative c-erbB-2 MBCs cases expressed immunostaining for p21Waf1 (P = 0.05). p27Kip1 immunoreactivity was been detected in 26 of 27 (96.2%) male breast patients versus 39 of 101 FBC (39.3%) (P = 0.000). Highly positive staining for P27Kip1 was found in 21 of 25 androgen receptor-expressing samples. Higher levels of p27Kip1 were expressed in bcl-2 positive samples (17 of 20). Eighteen of 22 c-erbB-2-negative cases were strongly immunoreactive for p27Kip1. CONCLUSIONS: p27Kip1 and p21Waf1 immunoreactivity is higher in MBCs compared with FBCs. The findings of higher p27Kip1 and p21Waf1 immunostaining may be an additional predictive factor in MBC. These biological features could be possible indicators for different biological pathways in the tumorigenesis of MBCs. PMID- 12123336 TI - Comparable efficacy and safety profiles of once-per-cycle pegfilgrastim and daily injection filgrastim in chemotherapy-induced neutropenia: a multicenter dose finding study in women with breast cancer. AB - BACKGROUND: Neutropenia is common in patients receiving myelotoxic chemotherapy. Pegfilgrastim, a sustained-duration filgrastim is a once-per-cycle therapy for prophylactic neutrophil support. PATIENTS AND METHODS: Women, treated with four cycles of doxorubicin/docetaxel chemotherapy every 21 days, received pegfilgrastim or filgrastim 24 h after chemotherapy as a single subcutaneous injection per chemotherapy cycle (pegfilgrastim 30, 60 or 100 microg/kg) or daily subcutaneous injections (filgrastim 5 microg/kg/day). Safety, efficacy and pharmacokinetics were analyzed. RESULTS: The incidence of grade 4 neutropenia in cycle 1 was 95, 90 and 74%, in patients who received pegfilgrastim 30, 60 and 100 microg/kg, respectively, and 76% in patients who received filgrastim. Mean duration of grade 4 neutropenia in cycle 1 was 2.7,2 and 1.3 days for doses of pegfilgrastim, and 1.6 days for filgrastim. The pharmacokinetics of pegfilgrastim were non-linear and dependent on both dose and neutrophil count. Pegfilgrastim serum concentration was sustained until the neutrophil nadir occurred then declined rapidly as neutrophils started to recover, consistent with a self regulating neutrophil-mediated clearance mechanism. The safety profiles of pegfilgrastim and filgrastim were similar. CONCLUSIONS: A single subcutaneous injection of pegfilgrastim 100 microg/kg provided neutrophil support and a safety profile comparable to daily subcutaneous injections of filgrastim during multiple chemotherapy cycles. PMID- 12123337 TI - EORTC 10968: a phase I clinical and pharmacokinetic study of polyethylene glycol liposomal doxorubicin (Caelyx, Doxil) at a 6-week interval in patients with metastatic breast cancer. European Organization for Research and Treatment of Cancer. AB - BACKGROUND: We performed a phase I study of polyethylene glycol (pegylated, Stealth) liposomal doxorubicin (Caelyx, Doxil) using a prolonged (6-week) dose interval to reduce the incidence of skin toxicity that was dose-limiting at more conventional dose intervals, and which appeared to be schedule dependent. PATIENTS AND METHODS: Eligible for the study were metastatic breast cancer patients who had received a maximum of one prior therapy for metastatic disease. The defined dose levels were 60, 70, 80 and 90 mg/m2. RESULTS: Twenty patients were assessed at starting doses of 60 mg/m2 (n = 9) or 70 mg/m2 (n = 11). The dose-limiting toxicity was mucositis. Severe skin toxicity was not observed at the 60 mg/m2 dose level, and occurred in only one patient treated at 70 mg/m2. Significant neutropenia, alopecia, and nausea and vomiting were rare events. No clinical cardiac events occurred, despite a median cumulative doxorubicin dose of 323 mg/m2 (range 5-630 mg/m2). Partial responses were documented in five patients. Pharmacokinetics were assessed in 15 patients, and confirmed the long terminal half-life of the agent (median 77 h) demonstrated in earlier studies. CONCLUSIONS: The recommended dose of Caelyx/Doxil using this schedule is 60 mg/m2 every 6 weeks. This is a safe and effective regimen that permits prolonged administration of anthracycline to patients with metastatic breast cancer. PMID- 12123338 TI - Gemcitabine, epirubicin and paclitaxel: pharmacokinetic and pharmacodynamic interactions in advanced breast cancer. AB - BACKGROUND: The objectives of this study were to investigate the disposition of gemcitabine, epirubicin, paclitaxel, 2',2'-difluorodeoxyuridine and epirubicinol, and characterize the pharmacokinetic and pharmacodynamic profile of treatment in patients with breast cancer. PATIENTS AND METHODS: The drug dispostion in 15 patients who received gemcitabine 1000 mg/m2, epirubicin 90 mg/m2 and paclitaxel 175 mg/m2 (GEP) on day 1 of a 21-day cycle, was compared with that of patients treated with epirubicin 90 mg/m2 and paclitaxel 175 mg/m2 (EP, n = 6) and epirubicin 90 mg/m2 alone (n = 6). Drug and metabolite levels in plasma and urine were assessed by high-performance liquid chromatography and parameters of drug exposure were related to hematological toxicity by a sigmoid-maximum effect (Emax) model. RESULTS: Paclitaxel administration significantly increased the epirubicinol area under the concentration-time curve, from 357+/-146 (epirubicin) to 603+/-107 (EP) and 640+/-81 h x ng/ml (GEP), and reduced the renal clearance of epirubicin and epirubicinol by 38 and 52.2% and 34.5 and 53% in GEP- and EP treated patients, respectively, compared with epirubicin alone. Gemcitabine had no apparent effect on paclitaxel and epirubicin pharmacokinetics, and renal clearance of epirubicin and epirubicinol. The only pharmacokinetic/pharmacodynamic relationship observed was between neutropenia and the time spent above the threshold plasma level of 0.1 micromol/l (tC0.1) of paclitaxel, with the time required to obtain a 50% decrease in neutrophil count (Et50) of GEP being 7.8 h, similar to that of EP. CONCLUSIONS: Paclitaxel and/or its vehicle, Cremophor EL, interferes with the disposition and renal excretion of epirubicin and epirubicinol; gemcitabine has no affect on epirubicin and paclitaxel plasma pharmacokinetics and renal excretion of epirubicin, while neutropenia is not enhanced by gemcitabine. PMID- 12123339 TI - Factors affecting toxicity, response and progression-free survival in relapsed patients with indolent B-cell lymphoma and mantle cell lymphoma treated with rituximab: a Japanese phase II study. AB - BACKGROUND: The aim of the study was to determine factors affecting the toxicity and efficacy of rituximab monotherapy in relapsed patients with indolent B-cell lymphoma and mantle cell lymphoma (MCL). PATIENTS AND METHODS: A total of 90 patients were enrolled and treated with rituximab infusions at 375 mg/m2 once weekly for 4 weeks. Central pathology review revealed that histologically, 81 patients had indolent B-cell lymphoma or MCL: 59 with follicular lymphoma, 17 with MCL, four with marginal zone lymphoma and one with lymphoplasmacytoid lymphoma. Of these, four were ineligible due to violation of other eligibility criteria. Pre-treatment variables affecting toxicities were analyzed for all 90 patients, and those affecting response and progression-free survival (PFS) were analyzed for 77 eligible patients with confirmed indolent B-cell lymphoma or MCL. The relationship between serum rituximab levels and efficacy was also analyzed for 66 eligible patients. RESULTS: Hematological toxicities (grade > or =3) occurred more frequently in females (P <0.05), and thrombocytopenia and leukopenia were more frequent in patients with high lactate dehydrogenase (LDH) levels (P <0.05). Non-hematological toxicities (grade > or =2) were more frequent in patients with extranodal disease or bone marrow involvement. The overall response rate (ORR) in patients receiving one prior chemotherapy regimen was higher than those receiving two or more regimens (P <0.05). The median PFS was shorter in MCL patients, in those with extranodal disease, or in those receiving two or more prior chemotherapy regimens (P <0.01). The PFS intervals of patients with higher serum rituximab levels (> or =70 microg/ml) immediately before the third infusion were longer than in other patients (P <0.01). CONCLUSIONS: Several prognostic factors and serum rituximab levels are useful for predicting the toxicity and efficacy of rituximab monotherapy. PMID- 12123341 TI - A comparison of bedside renal function estimates and measured glomerular filtration rate (Tc99mDTPA clearance) in cancer patients. AB - BACKGROUND: The aim of this study was to compare measured glomerular filtration rate (GFR) with estimates of GFR derived from the population pharmacokinetic methods of Martin and Wright, and the creatinine clearance (CrCl) estimates of Cockcroft and Gault, and Jelliffe. PATIENTS AND METHODS: GFR was determined by technetium-99m diethyl triamine penta-acetic acid (Tc99DTPA) clearance in adult cancer patients. Height, actual body weight and serum creatinine were measured, and GFR and CrCl estimates calculated. RESULTS: One hundred and twenty-two patients were included. The mean measured GFR was 87 ml/min (range 30-174 ml/min). The mean bias (mean percentage error) was 2, 1, -10 and -17%, and the mean precision (mean absolute percentage error) was 18, 19, 21 and 23% for the Wright, Martin, Cockcroft and Gault, and Jelliffe formulas, respectively. The Martin formula significantly underestimates GFR for females (mean bias -10%) and overestimates GFR for males (mean bias 8%) (P <0.001 for bias of males versus females). The Wright and Martin formulas significantly overestimate GFR <50 ml/min (mean bias 39 and 30%; P = 0.03 and 0.05, respectively) and all formulas underestimate GFR >100 ml/min (mean bias -18, -16, -24 and -32% for Wright, Martin, Cockcroft and Gault, and Jelliffe formulas, respectively; P <0.001). CONCLUSIONS: All the assessed estimates for renal function were found to have significant limitations. PMID- 12123340 TI - Treatment of childhood Wilms' tumor without radiotherapy in Nicaragua. AB - BACKGROUND: Recent trends in therapeutic strategies for Wilms' tumor are based on an attempt to reduce or omit radiotherapy (RT) in a sizable fraction of patients. We report here the clinical and histological features as well as the results obtained in 37 children (23 males, 14 females; median age at diagnosis 3 years, range 0.8-8 years) diagnosed between 1991 and 1996, and treated with chemotherapy (CT) and surgery at La Mascota Hospital, Managua, Nicaragua. PATIENTS AND METHODS: Patients were grouped as follows: those who underwent surgery at diagnosis (group A, n = 4), patients who received preoperative CT because of large tumor size (group B, n = 27), lung metastases (n = 5) or bilateral disease (n = 1) (group C, n = 6). Treatment consisted of vincristine (VCR) and actinomycin-D (ACTD) for 24 weeks in group A, and of VCR, ACTD and adriamycin for 68 weeks in groups B and C. Histology was classified as favorable in 30 patients (81%), unfavorable in six patients (all of group B) and unknown in one. RESULTS: With a median follow-up time of 6.4 years the event-free survival for the whole group was 80.1%+/-6.8 (SE). No event occurred beyond 5 years of diagnosis. CONCLUSIONS: These results suggest that RT does not appear necessary for the majority of patients, and that an excellent surgical approach associated with an intensive CT schedule can control the disease, even in the absence of adequate information on the intra-abdominal tumor extent. PMID- 12123342 TI - Localized unresectable neuroblastoma: results of treatment based on clinical prognostic factors. AB - BACKGROUND: We previously reported that stage 3 neuroblastoma comprises (i) a low risk group including all infants (age 0-11 months) as well as older children with non-abdominal primaries, and (ii) a high-risk group made up of children >1 year of age with abdominal primaries. Aggressive chemotherapy was effective only in the latter group. PATIENTS AND TREATMENT: On this basis, in 1990 we designed a new protocol by which all low-risk patients received standard-dose chemotherapy, while the high-risk ones received very aggressive chemotherapy. RESULTS: Between November 1990 and December 1997 a total of 95 eligible and evaluable children were enrolled: 47 were low-risk (35 infants and 12>1 year of age at diagnosis and having non-abdominal primaries), and 48 were high-risk (being >1 year of age and having abdominal primaries). Of the 47 low-risk patients, five relapsed and four subsequently died. The 5-year overall survival (OS) was 91%. Of the 48 patients in the high-risk group, 22 relapsed or progressed, 18 of whom died from their disease and two from toxicity, and one was lost to follow-up. The 5-year OS was 60%. Univariate analysis showed that age, site of primary, risk-group, urine vanillylmandelic excretion, plasma levels of lactate dehydrogenase, ferritin and neurone-specific enolase, and MYCN status correlated with outcome. However, multivariate analysis showed that only MYCN status retained prognostic value. CONCLUSIONS: In low-risk stage 3 neuroblastoma, standard-dose chemotherapy is associated with an excellent chance of being cured. Aggressive chemotherapy is effective for high-risk patients, but results are still unsatisfactory. MYCN gene amplification is a prognostic indicator for most, but not all, treatment failures. PMID- 12123343 TI - The impact of hemoglobin levels on fatigue and quality of life in cancer patients. AB - BACKGROUND: Although fatigue is a commonly reported symptom in cancer patients its etiology is still poorly understood. The objective of the present study was to investigate the relationship between hemoglobin (Hb) levels and the subjective experience of fatigue and quality of life in cancer patients with mild or no anemia undergoing chemotherapy. PATIENTS AND METHODS: Sixty-eight cancer patients (25 colorectal, 26 lung and 17 ovarian cancer) presently undergoing chemotherapy participated in the study. Fatigue was measured with the Multidimesional Fatigue Inventory (MFI-20), quality of life with The European Organization for Research and Treatment of Cancer QLQ-C30. In order to provide normative data for fatigue levels, the MFI-20 was also completed by a sex- and age-matched sample of 120 healthy controls. RESULTS: Compared with healthy subjects, cancer patients experienced significantly higher levels of subjective fatigue. Correlations between Hb values and subscales of the MFI-20 were moderate with a tendency to increase during chemotherapy. Hb values alone, however, do not fully account for the observed fatigue. Other symptoms, especially pain, dyspnea and sleep disturbances, also showed an association with perceived fatigue. CONCLUSIONS: Despite significant correlations, these results indicate that Hb values only partially explain subjectively experienced fatigue and quality of life in cancer patients. It is suggested therefore that the treatment of fatigue must be multidimensional and involve all areas which contribute to the syndrome. PMID- 12123344 TI - Sexual precocity and recurrent beta-human chorionic gonadotropin upsurges preceding the diagnosis of a malignant mediastinal germ-cell tumor in a 9-year old boy. AB - Numerous disorders are known to cause sexual precocity. Beta-human chorionic gonadotropin (beta-HCG)-secreting germ-cell tumors are one of the sources that have to be considered in the differential diagnosis of processes inducing a peripheral precocious puberty. Germ-cell tumors might be located in the ovaries or testes, retroperitoneum, mediastinum or the cranium. We present the case of a 9-year-old boy with sexual precocity and a recurrent transient beta-HCG elevation. After an interval of 2 years with repeated radiological examinations including the mediastinum, a mediastinal tumor was identified by magnetic resonance imaging. To our knowledge, this is the first case of a diagnosis of a mediastinal choriocarcinoma with a recurrent serum beta-HCG elevation. So far, factors that might be responsible for the repeated spontaneous beta-HCG decline are unknown. PMID- 12123345 TI - Gastrointestinal toxicity associated with weekly docetaxel treatment. AB - Previous studies have demonstrated a marked reduction of haematological and non haematological toxicity if weekly doses of docetaxel <40 mg/m2 were used. Reviewing the literature, neutropenic enterocolitis is uncommon but not unknown in patients treated with taxane-based chemotherapy. Although this complication occurs rarely, here we report on two patients, one with metastatic breast cancer and one with non-small-cell lung cancer, treated on a weekly schedule with single agent docetaxel. Both patients developed excessive and fatal haemorrhragic gastroduodenitis and enterocolitis associated with grade 2 and 3 neutropenia. We would like to stress the importance of symptoms such as abdominal pain and tenderness, fever, diarrhoea and mucositis, with or without neutropenic fever, in patients treated with docetaxel-based chemotherapy. These symptoms should alert the physician and supportive care management should be started aggressively and immediately. PMID- 12123346 TI - Potential of capecitabine as first-line therapy for metastatic breast cancer: dosing recommendations in patients with diminished renal function. PMID- 12123347 TI - Short- and long-term safety of the 2 x 10(6) CD34+ cells/kg threshold for hematopoietic reconstitution after high-dose chemotherapy and peripheral blood progenitor cell support. PMID- 12123348 TI - Fluorescent sensing of pyrophosphate and bis-carboxylates with charge neutral PET chemosensors. AB - [reaction: see text] The fluorescent photoinduced electron transfer (PET) chemosensors 2 and 3 were designed for the recognition of anions possessing two binding sides such as dicarboxylates and pyrophosphate; the anion recognition in DMSO takes place through the two charge neutral thiourea receptor sites with concomitant PET quenching of the anthracene moiety. The anion binding of acetate, phosphate, and pyrophosphate to 2 and 3 was also evaluated by using 1H NMR in DMSO-d6. PMID- 12123349 TI - High diastereoselectivity in intermolecular carbonyl ylide cycloaddition with aryl aldehyde using methyl diazo(trifluoromethyl)acetate. AB - [reaction: see text] Methyl diazo(trifluoromethyl)acetate undergoes Rh2(OAc)4 catalyzed reaction with aryl aldehyde to form 1,3-dioxolanes bearing a C-4 trifluoromethyl group diastereoselectively in excellent yield. PMID- 12123350 TI - Enantioselective 1,3-dipolar cycloaddition of nitrones catalyzed by optically active cationic cobalt(III) complexes. AB - [reaction: see text] The optically active beta-ketoiminato cationic cobalt(III) complexes were employed as efficient Lewis acid catalysts for the enantioselective 1,3-dipolar cycloaddition reaction of alpha,beta-unsaturated aldehydes with nitrones. Excellent endo selectivities and high enantioselectivities were achieved in the cycloaddition reaction of 1 cyclopentene-1-carbaldehyde and the nitrones derived from 2-halobenzaldehyde. PMID- 12123351 TI - Electrochemical recognition of cations by bis(pyrrolo)tetrathiafulvalene macrocycles. AB - [structure: see text] Tetrathiafulvalene redox-responsive ligands devoid of cis/trans isomerism containing the electroactive bis(pyrrolo[3,4 d])tetrathiafulvalene moiety and polyether subunits have been synthesized. One ligand exhibits high binding affinities for Pb2+ and Ba2+ cations as shown by independent methods (1H NMR, UV-vis spectroscopy, and cyclic voltammetry). The ability of this receptor to electrochemically recognize Pb2+ and Ba2+ is shown by cyclic voltammetry. PMID- 12123352 TI - Synthesis of 2-indanones via [4 + 1] annulation reactions of (trialkylsilyl)arylketenes. AB - [reaction: see text] (Trialkylsilyl)arylketenes combine with (trimethylsilyl)diazomethane in a new [4 + 1] annulation process leading to 2 indanone derivatives. The (trialkylsilyl)arylketene annulation substrates are available via the photochemical Wolff rearrangement of alpha-silyl-alpha-diazo ketones, which are themselves prepared by silylation of the corresponding diazo ketones. The mechanism of the annulation reaction is proposed to involve the formation of a 2,3-bis(silyl)cyclopropanone, which is in equilibrium with an oxyallylic cation. Electrocyclic closure of this intermediate forms the new cyclopentenone ring. PMID- 12123353 TI - Asymmetric synthesis of (-)-adaline. AB - [reaction: see text] An enantioselective total synthesis of (-)-adaline has been achieved starting from a chiral 6,6-disubstituted piperidone derivative previously prepared by diastereoselective allylation of a chiral tricyclic N-acyl N,O-acetal. The key steps include lithium ion-activated SN2-type alkynylation of the tricyclic N,O-acetal leading to exclusive formation of the (6S) ethynylpiperidine and ring-closing olefin metathesis of the (2R,6S)-cis-2,6 dialkenylpiperidine for constructing the bridged azabicyclononane. PMID- 12123354 TI - Toward the combinatorial synthesis of polyketide libraries: asymmetric aldol reactions with alpha-chiral aldehydes on solid support. AB - [reaction: see text] The viability of performing stereocontrolled aldol additions with alpha-chiral aldehydes attached by a silyl linker to a hydroxymethylpolystyrene resin is demonstrated for boron and titanium enolates. Subsequent ketone reduction and manipulation on the solid support leads to elaborate stereopentad sequences, as occur in 6-deoxyerythronolide B and discodermolide. PMID- 12123355 TI - Complete pi-facial diastereoselectivity in Diels-Alder reactions of dissymmetric 2,4-cyclohexadienones. AB - [reaction: see text] The studies in the Diels-Alder reactions of 5-methoxy-masked o-benzoquinone (1a, R = OMe) and simple dissymmetric 2,4-cyclohexadienones 1b-e with methyl vinyl ketone, styrene, and benzyl vinyl ether are described. The dienones 1b-e reacted with dienophiles to form syn adducts (dienophile approach is syn to allylic methoxy group) exclusively. PMID- 12123356 TI - Intramolecular PhI=O mediated copper-catalyzed aziridination of unsaturated sulfamates: a new direct access to polysubstituted amines from simple homoallylic alcohols. AB - [reaction: see text] Olefinic sulfamates derived from primary and secondary alcohols undergo intramolecular copper-catalyzed aziridination in the presence of iodosylbenzene to afford novel bicyclic fused aziridines. The latter were opened by various nucleophiles to give the corresponding substituted cyclic sulfamates, which in turn reacted, after nitrogen activation, with a second nucleophile at the carbon atom bearing the oxygen atom. Concomitant removal of the sulfonyloxy moiety thus gave access to polysubstituted amines. PMID- 12123357 TI - Sonogashira coupling of functionalized trifloyl oxazoles and thiazoles with terminal alkynes: synthesis of disubstituted heterocycles. AB - [reaction: see text] This paper describes Sonogashira cross-coupling of functionalized 2-, 4-, and 5-trifloyl oxazoles and thiazoles with terminal alkynes. This methodology has been extended to 2,4-ditrifloylthiazoles, which results in regioselective cross-coupling at the C2-position of the thiazole. The resulting 2-alkynyl-4-trifloylthiazoles are effective electrophiles in a second palladium(0)-mediated cross-coupling reaction. PMID- 12123358 TI - Electron transfer initiated heterogenerative cascade cyclizations: polyether synthesis under nonacidic conditions. AB - [reaction: see text] Single-electron oxidation has been employed to initiate heterogenerative cascade cyclization reactions that form polyether compounds under essentially neutral conditions. The reactions proceed through mesolytic benzylic carbon-carbon bond cleavages of homobenzylic ether-derived radical cations followed by intramolecular epoxonium ion formation, leading to further cyclizations. Both oligotetrahydrofuran and tetrahydropyran structures can be prepared by altering substrate topography. PMID- 12123359 TI - Alkenenitriles: annulations with omega-chloro Grignard reagents. AB - [reaction: see text] omega-Chloro Grignard reagents chelate with cyclic gamma hydroxy-alpha,beta-alkenenitriles to trigger a conjugate addition-alkylation annulation. The chelation-controlled conjugate addition-alkylation is the first anionic annulation with alpha, beta-alkenenitriles, providing cis bicyclo[3.3.0]octane, hydrindane, and decalin ring systems in a single synthetic operation. PMID- 12123360 TI - Monolithic scavenger resins by amine functionalizations of poly(4-vinylbenzyl chloride-co-divinylbenzene) PolyHIPE materials. AB - [reaction: see text] Monolithic polymer supports and scavengers were prepared via nucleophilic displacement of chlorine in poly(4-vinylbenzyl chloride-co divinylbenzene) PolyHIPE materials. Reactions of monolithic PolyHIPE with tris(2 aminoethyl)amine, 4-aminobutanol, tris(hydroxymethyl)aminomethane, morpholine, and hexamethylenetetramine led to functionalized polymers with amino and hydroxy functionalities with high degrees of conversion. 4-Chlorobenzoyl chloride was efficiently and rapidly scavenged from solution by the tris(2-aminoethyl)amine derivative of monolithic poly(4-vinylbenzyl chloride-co-divinylbenzene) PolyHIPE at ambient temperature. PMID- 12123361 TI - Synthesis and NMR studies of cyclopeptides containing a sugar amino Acid. AB - [structure: see text] Cyclopeptides containing Glucuronic acid methylamine (Gum) alternating with Gly, L-Ala, D-Ala, L-Phe, D-Phe, L-Lys, or D-Lys were synthesized by a combination of solid-phase synthesis and solution chemistry. A more effective pathway to synthesize the sugar amino acid Gum in higher yields and in a shorter period of time was developed. Gum is employed in the benzylated and deprotected form. The cyclopeptides were characterized by NMR and the structure of one deprotected cyclic peptide solved. PMID- 12123362 TI - Synthesis of substituted naphthalenes by the palladium-catalyzed annulation of internal alkynes. AB - [reaction: see text] A variety of substituted naphthalenes have been prepared by the palladium-catalyzed carboannulation of internal alkynes. This method (1) forms two new carbon-carbon bonds in a single step, (2) accommodates a variety of functional groups, and (3) affords excellent yields of highly substituted naphthalenes. PMID- 12123363 TI - Solid-phase synthesis of symmetrical 3,6-bispeptide-acridone conjugates. AB - [reaction: see text] A novel high-yielding method for the solid-phase synthesis of 3,6-bispeptide-acridone conjugates is reported. It involves initial coupling of bifunctionalized acridone to a resin-bound peptide followed by an on-bead site site reaction to couple the second peptide. This method leads to clean symmetrical bispeptide derivatives and appears to be general. This strategy will enable the generation of a library of 3,6-bispeptide-acridones to be screened for selective binding to telomeric G-quadruplex DNA. PMID- 12123364 TI - A novel synthesis of enantiopure octahydropyrrolo[3,4-b]pyrroles by intramolecular [3 + 2] dipolar cycloaddition on chiral perhydro-1,3-benzoxazines. AB - [reaction: see text] Condensation of N-substituted glycines with chiral 3-allyl-2 formyl perhydro-1,3-benzoxazines forms an azomethine ylide that cyclizes to give octahydropyrrolo[3,4-b]pyrrole derivatives. The [3 + 2] dipolar cycloadditions are stereoespecific leading to a single diastereoisomer. The chemical yields are dependent on the reaction temperature and the presence or absence of a base. PMID- 12123365 TI - Total synthesis of (-)-minquartynoic acid: an anti-cancer, anti-HIV natural product. AB - [structure: see text] The tetraacetylenic compound, (S)-minquartynoic acid (1), is synthesized in seven linear steps and 17% overall yield from commercially available azelaic acid monomethyl ester. The key step is a one-pot three component Cadiot-Chodkiewicz reaction to construct the tetrayne unit without using either a diyne or a triyne intermediate. PMID- 12123366 TI - Study of alpha-crustacyanin utilizing halogenated canthaxanthins. AB - [structure: see text] The preparations and spectroscopic characteristics of five all-trans halogenated canthaxanthins are described in this letter. The air/light sensitive halogenated canthaxanthins were used to study alpha-crustacyanin, a blue astaxanthin-protein complex, which is isolated from the carapace of the lobster. Steric and electronegative effects of the halogen substituents on the noncovalent interaction between astaxanthin and the protein in alpha-crustacyanin were observed. PMID- 12123367 TI - An expedient route to the tetrazole analogues of alpha-amino acids. AB - [reaction: see text] Convenient conditions are described for the transformation of alpha-aminonitriles to the tetrazole analogues of alpha-amino acids. Refluxing the starting material in water/2-propanol at 80 degrees C with sodium azide and catalytic zinc bromide affords the tetrazole product in yields generally exceeding 90%. PMID- 12123368 TI - Catalytic asymmetric oxidative couplings of 2-naphthols by tridentate N ketopinidene-based vanadyl dicarboxylates. AB - [reaction: see text] A series of oxovanadium(IV) complexes derived from tridentate N-ketopinidene-alpha-amino acids were synthesized. They serve as efficient catalysts for the enantioselective oxidative couplings of various 3-, 6 , or 7-substituted 2-naphthols. The best scenario involves the use of a vanadyl complex arising from L-tert-leucine in CCl4. The asymmetric couplings of 2 naphthols can be conducted smoothly at 40-45 degrees C under a stream of gaseous oxygen, leading to 2,2'-dihydroxy-1,1'-binaphthyls in good yields (61-99%) and with enantioselectivities of up to 87%. PMID- 12123369 TI - Oligo(octithienylene-diethynylene)s as unprecedentedly long conjugated nanomolecules. AB - [reaction: see text] A series of oligo(octithienylene-diethynylene)s alternately composed of octithiophene and diacetylene units have been prepared by a random Eglinton coupling reaction among mono- and diethynyloctithiophenes. The largest compound isolated in the oligomeric series is comprised of twelve octithiophene units and eleven diacetylene units, and its molecular length reaches ca. 43 nm, which is the longest among single-component conjugated nanomolecules. PMID- 12123370 TI - Stereoselective synthesis of trans beta-lactams through iridium-catalyzed reductive coupling of imines and acrylates. AB - [reaction: see text] Iridium-catalyzed reductive coupling of acrylates and imines provides trans beta-lactams with high diastereoselection. The optimal catalyst allows for the synthesis of trans beta-lactams bearing aromatic, alkenyl, and alkynyl side chains. This reaction appears to proceed through a reductive Mannich addition-cyclization mechanism. Examination of substituent effects reveals a linear Hammett correlation for both the N-aryl group on the imine and the aryloxy group on the acrylate, thereby pointing to rate-determining cyclization in the reaction mechanism. PMID- 12123371 TI - Synthesis, X-ray structure, and properties of the singly bonded C60 dimer having diethoxyphosphorylmethyl groups utilizing the chemistry of C60(2-). AB - [reaction: see text] A singly bonded C60 dimer having a diethoxyphosphorylmethyl group on each C60 cage was obtained by the reaction of C60(2-) dianion with diethyl iodomethylphosphonate followed by the treatment with iodine. The precise structure of the dimer was determined for the first time by X-ray crystallography, and its homolytic dissociation as well as spectroscopic and electrochemical properties were clarified. PMID- 12123372 TI - Highly active Pd(II) catalysts with trans-bidentate pyridine ligands for the Heck reaction. AB - [reaction: see text] The air-, water-, and heat-stable palladium(II) complexes 2a and 2b are prepared by the reaction of palladium(II) salts with the new trans bidentate nitrogen ligands, 1,2-bis(2-pyridylethynyl)benzenes. The structure of complex 2a has been confirmed by X-ray structure analysis. The complexes efficiently catalyze the Heck olefination of aryl iodides and provide a good yield under phosphine-free conditions. The reaction is very sensitive to the nature of the chelating ligand. PMID- 12123373 TI - A new route to sulfonamides via intermolecular radical addition to pentafluorophenyl vinylsulfonate and subsequent aminolysis. AB - [reaction: see text] Various radical species generated from either the corresponding iodo- or bromo- compounds and tri-n-butyltin hydride were added in an intermolecular fashion to the activated acceptor pentafluorophenyl vinylsulfonate. The products of each reaction were then subjected to aminolysis with a variety of different amines. PMID- 12123374 TI - Importance of the methyl group at C10 of squalene for hopene biosynthesis and novel carbocyclic skeletons with 6/5 + 5/5 + (6) ring system(s). AB - [reaction: see text] Incubation of (6E,10E,14E,18E)-2,6,10,19,23-pentamethyl tetracosa-2,6,10,14,18,22-hexaene with Alicyclobacillus acidocaldarius hopene cyclase afforded four products having two types of carbocyclic skeletons, i.e., two hopane products and two products having an unprecedented carbocyclic skeleton of 6/5 + 5/5 +6 pentacyclic and 6/5 + 5/5 tetracyclic ring systems. The former two hopane skeletons were formed from the bioconversion of C15-desmethylsqualene and the latter two skeletons from that of C10-desmethylsqualene. PMID- 12123375 TI - Aromatization of enamines promoted by a catalytic amount of Pd/C. Synthesis of aromatic amines. AB - [reaction: see text] Aromatic amines were obtained efficiently from enamines by using a catalytic amount of Pd/C in the presence of nitrobenzene and 4 A molecular sieves in refluxing toluene. PMID- 12123376 TI - Cyclic carbonate formation from carbon dioxide and oxiranes in tetrabutylammonium halides as solvents and catalysts. AB - [reaction: see text] Epoxides dissolved in molten tetralkylammonium salts bearing halides as counterions are converted into cyclic carbonates under atmospheric pressure of carbon dioxide. The reaction rate depends on the nucleophilicity of the halide ion as well as the structure of the cation. PMID- 12123377 TI - Synthesis of new heterocycles through a cation-driven tandem ring-enlargement annulation reaction. AB - [reaction: see text] The thionium ion, generated through a cyclopropylcarbinyl cyclobutyl ring expansion, is, for the first time, intramolecularly intercepted by activated aromatic rings to generate new versatile 2a-methyl-8b (phenylsulfanyl-1,2a,3,8b-tetrahydro-2H-cyclobuta[c]chromenes. PMID- 12123378 TI - W(CO)5-amine catalyzed exo- and endo-selective cyclizations of omega-alkynyl silyl enol ethers: a highly useful method for the construction of polycyclic compounds. AB - [reaction: see text] A highly useful method for the construction of polycyclic compounds based on the amine-controlled exo- and endo-selective cyclizations of omega-alkynyl silyl enol ethers catalyzed by W(CO)5L is reported. When bis alkynyl silyl enol ethers were treated with a catalytic amount of W(CO)6, DABCO, and water under photoirradiation, synthetically useful tricyclic ketones were obtained in good yield. PMID- 12123379 TI - Generation and trapping of alkene radical cations under nonoxidizing conditions: formation of six-membered rings by exo- and endo-mode cyclizations. AB - [reaction: see text] It is demonstrated that alkene radical cations generated by the radical ionic fragmentation of beta-(phosphatoxy)alkyl radicals undergo efficient nucleophilic capture by amines in either the 6-exo or 6-endo modes, leading to six-membered nitrogen heterocycles. Suitable placement of an alkene enables the juxtaposition of a radical cyclization resulting in the formation of both the indolizidine and 1-azabicyclo[3.2.1]octane skeleta. PMID- 12123380 TI - First asymmetric total synthesis of (+)-sparteine. AB - [reaction: see text] The total synthesis of (+)-sparteine was accomplished from 2,5-norbornadione in 15 steps and 15.7% overall yield. The key steps were two ring-expansion reactions, one involving an intramolecular Schmidt reaction and one using a novel variant of the photo-Beckmann rearrangement. PMID- 12123381 TI - Synthesis of the first 1,3-ditellurole-containing radialene-type TTF derivatives. AB - [reaction: see text] The first synthesis of dendralene-type TTF derivatives bearing a 1,3-ditellurole ring has been achieved. Preliminary electrochemical results are presented. PMID- 12123382 TI - Resolution of 1-(2-naphthyl)ethanol by a combination of an enzyme-catalyzed kinetic resolution with a fluorous triphasic separative reaction. AB - [reaction: see text] Kinetic resolution of a fluorous ester rac-1 with Candida antarctica B lipase provided a mixture of enantioenriched alcohol (R)-2 and fluorous ester (S)-1. The mixture was subjected to a fluorous triphasic reaction to give both enantiomers of 1-(2-naphthyl)ethanol 2 in high ee without chromatographic separation or fluorous-organic liquid-liquid extractive purification. PMID- 12123384 TI - Intramolecular proton transfer. AB - [structure: see text] Reversal of the normal kinetic protonation stereochemistry results as a consequence of intramolecular delivery. PMID- 12123385 TI - Design and development of cyclohexane-based P,N-ligands for transition metal catalysis. AB - [reaction: see text] A new class of cyclohexane-based P,N-ligands is readily obtained through aziridine ring opening with suitable phosphorus nucleophiles under acidic conditions. trans-1-Amino-2-diphenylphosphinocyclohexane is resolved with tartaric acid to give the final product in >99% ee. The new ligands show high stability toward oxidation at the phosphorus atom. PMID- 12123383 TI - BINOLAM, a recoverable chiral ligand for bifunctional enantioselective catalysis: the asymmetric synthesis of cyanohydrins. AB - [reaction: see text] A new bifunctional catalytic system based on a monometallic aluminum complex is used for the efficient enantioselective cyanation of aldehydes. The ligand (S)- or (R)-2,2'-bis(diethylaminomethyl)-substituted binaphthol (BINOLAM) used is recovered for recycling. This methodology is used for the synthesis of a precursor of epothilone A. PMID- 12123387 TI - Efficient enantioselective additions of terminal alkynes and aldehydes under operationally convenient conditions. AB - [reaction: see text] The enantioselective addition reaction of terminal acetylenes and aldehydes mediated by Zn(OTf)2 and N-methyl ephedrine can be conducted with reagent grade solvent containing 84-1000 ppm H2O. The products can be isolated in high yield and useful enantioselectivities (up to 99% ee). PMID- 12123386 TI - An effective new synthesis of 4-aminopyrrole-2-carboxylates. AB - [reaction: see text] A series of 4-amino-1H-pyrrole-2-carboxylic acid benzyl esters has been synthesized in 61-84% yields on treatment of N-PhF-4-oxoproline benzyl ester and its 3-alkyl-substituted derivatives with different primary and secondary amines and a catalytic amount of TsOH in THF. 4-Hydroxy-1H-pyrrole-2 carboxylic acid benzyl esters were prepared in 59 and 70% yields by treatment of N-PhF-4-oxoproline benzyl esters with ammonium hydroxide in THF. PMID- 12123388 TI - Palladium(II)-mediated cascade carbonylative annulation of o-alkynyl-phenols on silyl linker-based macrobeads: a combinatorial synthesis of a 2,3-disubstituted benzo[b]furan library. AB - [reaction: see text] A palladium(II)-mediated cascade carbonylative annulation of o-alkynylphenols was achieved successfully on silyl linker-based macrobeads, which led to an efficient combinatorial synthesis of a 2,3-disubstituted benzo[b]furan library. PMID- 12123391 TI - Extra-individual sources of social support as described by adults with mild intellectual disabilities. AB - The extra-individual social support responses provided by adults with intellectual disabilities on a social support survey were examined. Felton and Berry (1992) argued that extra-individual social support, support from larger than individual entities, is a valid source of social support for older adults. We hypothesized that extra-individual support would also be relevant to adults with intellectual disabilities. Results showed that 43% of participants with intellectual disabilities listed extra-individual support sources in their social networks or as providers of one of several support functions. The most popular forms of extra-individual support were those from "staff" and "work." Findings are discussed in terms of implications for social support measurement for persons with intellectual disabilities. PMID- 12123392 TI - Effects of visual arts instruction on the mental health of adults with mental retardation and mental illness. AB - Single-subject multiple probe designs were employed in two studies with 5 young adults who had a dual diagnosis of mental retardation and mental illness. Our aim was to determine effects of instruction designed to teach visual arts activity skills and promote personal expressiveness on acquisition, maintenance, and generalization of these skills and behaviors associated with these persons' mental health. In Study 1, a 5-second constant time delay procedure was used to teach three chosen art activities. In Study 2, an instructional package was used to promote personally expressive behaviors. After learning the skills in Study 1, participants in Study 2 displayed improvement in occurrence of behaviors associated with mental illness and increases in personally expressive behaviors. PMID- 12123393 TI - Use of visual tools to report sexual abuse for adults with mental retardation. AB - Adults with mental retardation were assessed for their ability to use visual tools for identifying body parts. Participants were shown three representations: anatomical dolls, anatomical drawings, and live models, each of which had a sticker placed on a body part. They were asked to name that body part and place a sticker in the same place on their own body. Results indicated that verbal labeling was easier for participants with mild mental retardation compared to moderate mental retardation. Level of mental retardation affected the participants' ability to correctly place the stickers. Form of representation was also important. Live models were easier to use compared to dolls and drawings. Implications for forensic evaluations are discussed. PMID- 12123394 TI - Comparing the quality of life of school-age children with and without disabilities. AB - Quality of life of 76 school-age children with identified disabilities receiving special education services in public schools was compared to quality of life of 64 students without disabilities enrolled in Grades K-12. The Quality of Student Life Questionnaire (QSLQ) was used. Results indicated that the scores of students with disabilities were lower on all scales. The differences were significant in three of the four quality of life factors of the QSLQ scales: Satisfaction, p <.001, Well-Being, p <.01, Social-Belonging, p <.001, and total QSLQ scores, p <.001. Findings suggest that we have not yet achieved parity in quality of life for children with disabilities. The concept of quality of life is discussed in the context of needs for future intervention. PMID- 12123395 TI - Help or hindrance? Staff perspectives on developmental assessment in multicultural early childhood settings. AB - Staff members' views on developmental assessment in a multicultural early childhood setting are described and analysis of these views used to initiate a critique of current practice in assessment and evaluation of young children. Staff members expressed opinions and beliefs along a range from endorsement to frank rejection of the utility, validity, and ethics of developmental assessment. Those who reject the practice expressed a "theory" of growth and change that is incompatible with current developmentalist orthodoxy. Opinions of "dissenters" (who stated that they do not believe assessment gives meaningful information about children) suggest alternative practice with greater authenticity for families and children who are not from European American mainstream backgrounds. PMID- 12123396 TI - Multidimensional life satisfaction reports of adolescents with mild mental disabilities. AB - Using the Multidimensional Students' Life Satisfaction Scale-MSLSS(Huebner, 1994), we compared life satisfaction reports of 80 high school students with mild mental disabilities with a matched sample of 80 typically achieving students. The results provided preliminary support for the use of the MSLSS in research with secondary school students with mild mental disabilities. Comparisons of mean levels of general and domain-specific satisfaction suggested that the students with mild mental disabilities reported comparable positive levels with two exceptions. They reported lower satisfaction with their friendships and higher satisfaction with school experiences than did their typically achieving counterparts. The school satisfaction reports of students with mild mental disabilities varied as a function of differences in placement in special education. PMID- 12123397 TI - Internists and adolescent medicine. AB - Internists can assume a greater role in the provision of health care to adolescents. Adding an understanding of adolescent development to the skills and knowledge already possessed by internists will allow internists to interact more comfortably and effectively with adolescent patients. Learning about specific areas of importance such as adolescent morbidity and mortality, consent and confidentiality, interviewing techniques, and preventive health care will further enhance an internist's knowledge regarding adolescent health. There are numerous resources available to help intensive care for adolescent patients. PMID- 12123398 TI - Allogeneic hematopoietic stem cell transplantation: complications and results. AB - Acute complications such as veno-occlusive disease of the liver, acute and chronic graft-vs-host disease (GVHD), and infectious conditions remain major obstacles for the success of allogeneic hematopoietic stem cell transplantation (HSCT). Progress in allogeneic HSCT depends on several factors, including the adequate prevention and management of associated complications, advances in the conventional management of diseases currently treated with allogeneic HSCT, expansion of the donor pool, selective control of GVHD, development of more effective preparative regimens to eradicate the neoplastic cell population, characterization of a new generation of hematopoietic growth factors and cytokines, and development of newer techniques for ex vivo manipulation of stem cells. Hematopoietic growth factor-mobilized donor progenitor cells collected from peripheral blood have been shown to be associated with rapid hematopoietic engraftment without an increase in the incidence of acute GVHD compared with allogeneic bone marrow transplantation. Implementation of this approach will enhance donor acceptance, eliminate the risk of general anesthesia, decrease cost, and reduce the risk of infectious complications by reducing the duration of neutropenia. Nonmyeloablative allogeneic stem cell transplantation represents a novel treatment approach that may lead to reduced toxic effects and extended use of this treatment in older patients and in those with malignant and nonmalignant disorders. However, GVHD and disease recurrence remain a challenge. Promising results have been reported in patients with refractory hematologic malignancies and in metastatic renal cell cancer. Because late complications are commonly encountered in patients receiving allogeneic HSCT, lifelong observation is needed. PMID- 12123399 TI - Efficacy, safety, and tolerability of once-daily niacin for the treatment of dyslipidemia associated with type 2 diabetes: results of the assessment of diabetes control and evaluation of the efficacy of niaspan trial. AB - BACKGROUND: Diabetic dyslipidemia is characterized by high triglyceride levels; low high-density lipoprotein cholesterol levels; small, dense low-density lipoprotein particles; and high free fatty acid levels. Niacin reduces concentrations of triglyceride-rich and small low-density lipoprotein particles while increasing high-density lipoprotein cholesterol levels. It also lowers levels of free fatty acids and lipoprotein(a). However, the use of niacin in patients with diabetes has been discouraged because high doses can worsen glycemic control. We evaluated the efficacy and safety of once-daily extended release (ER) niacin in patients with diabetic dyslipidemia. METHODS: During a 16 week, double-blind, placebo-controlled trial, 148 patients were randomized to placebo (n = 49) or 1000 (n = 45) or 1500 mg/d (n = 52) of ER niacin. Sixty-nine patients (47%) were also receiving concomitant therapy with statins. RESULTS: Dose-dependent increases in high-density lipoprotein cholesterol levels (+19% to +24% [P<.05] vs placebo for both niacin dosages) and reductions in triglyceride levels (-13% to -28% [P<.05] vs placebo for the 1500-mg ER niacin) were observed. Baseline and week 16 values for glycosylated hemoglobin levels were 7.13% and 7.11%, respectively, in the placebo group; 7.28% and 7.35%, respectively, in the 1000-mg ER niacin group (P=.16 vs placebo); and 7.2% and 7.5%, respectively, in the 1500-mg ER niacin group (P=.048 vs placebo). Four patients discontinued participation because of inadequate glucose control. Rates of adverse event rates other than flushing were similar for the niacin and placebo groups. Four patients discontinued participation owing to flushing (including 1 receiving placebo). No hepatotoxic effects or myopathy were observed. CONCLUSION: Low doses of ER niacin (1000 or 1500 mg/d) are a treatment option for dyslipidemia in patients with type 2 diabetes. PMID- 12123400 TI - Physician career satisfaction across specialties. AB - BACKGROUND: The career satisfaction and dissatisfaction physicians experience likely influence the quality of medical care. OBJECTIVE: To compare career satisfaction across specialties among US physicians. METHODS: We analyzed data from the Community Tracking Study of 12 474 physicians (response rate, 65%) for the late 1990s. Data are cross-sectional. Two satisfaction variables were created: very satisfied and dissatisfied. Thirty-three specialty categories were analyzed. RESULTS: After adjusting for control variables, the following specialties are significantly more likely than family medicine to be very satisfying: geriatric internal medicine (odds ratio [OR], 2.04); neonatal perinatal medicine (OR, 1.89); dermatology (OR, 1.48); and pediatrics (OR, 1.36). The following are significantly more likely than family medicine to be dissatisfying: otolaryngology (OR, 1.78); obstetrics-gynecology (OR, 1.61); ophthalmology (OR, 1.51); orthopedics (OR, 1.36); and internal medicine (OR, 1.22). Among the control variables, we also found nonlinear relations between age and satisfaction; high satisfaction among physicians in the west north Central and New England states and high dissatisfaction in the south Atlantic, west south Central, Mountain, and Pacific states; positive associations between income and satisfaction; and no differences between women and men. CONCLUSIONS: Career satisfaction and dissatisfaction vary across specialty as well as age, income, and region. These variations are likely to be of interest to residency directors, managed care administrators, students selecting a specialty, and physicians in the groups with high satisfaction and dissatisfaction. PMID- 12123401 TI - The new definition of myocardial infarction: diagnostic and prognostic implications in patients with acute coronary syndromes. AB - BACKGROUND: The clinical implications of the recently revised criteria for diagnosis of acute myocardial infarction (AMI) in patients with suspected acute coronary syndromes are unknown. METHODS: To evaluate the prognostic implications of the new diagnostic criteria for AMI, we studied 493 consecutive patients with suspected acute coronary syndromes admitted to University of Michigan, Ann Arbor, between May 1, 1999, and January 1, 2000. Patients with positive cardiac enzymes and symptoms suggestive of coronary ischemia (n = 275) were divided into 2 groups: group A, with elevated peak creatine kinase-MB fraction and/or new electrocardiographic changes suggestive of AMI regardless of troponin status (diagnosed as AMI by old criteria), and group B, with normal peak creatine kinase MB fraction but elevated troponin I level (additional patients diagnosed as having AMI by new criteria). RESULTS: As compared with group A (n = 224), patients in group B (n = 51) were older women, with increased comorbidities such as previous stroke or aortic stenosis, and had fewer in-hospital procedures. In hospital adverse events (reinfarction, heart failure, shock, and mortality) were similar between the groups, whereas 6-month mortality was higher among group B patients (16.3% vs 5.8%; P =.03). This difference was not statistically significant after adjustment for differences in baseline characteristics between the groups (odds ratio, 1.6; 95% confidence interval, 0.5-5.9). CONCLUSIONS: The new criteria result in a substantial increase in the diagnosis of AMI. Furthermore, they help to identify patients with acute coronary syndromes who have greater comorbidities and worse 6-month outcomes who are otherwise missed by the old criteria. Additional studies are needed to confirm these preliminary findings and to determine the financial implications of the new criteria. PMID- 12123402 TI - Low-dose inhaled corticosteroid therapy and risk of emergency department visits for asthma. AB - BACKGROUND: Patients who visit the emergency department (ED) because of asthma frequently have a relapse. While the use of inhaled corticosteroids has been demonstrated to improve asthma symptoms and lung function, it is not clear whether their use after discharge from the ED reduces asthma relapse rates. OBJECTIVE: To determine whether inhaled corticosteroid therapy reduces ED asthma relapse rates. METHODS: We analyzed ED visit and medication data on patients 5 to 60 years of age who were enrolled in a government-sponsored drug plan and who visited an ED because of asthma between April 1, 1997, and March 31, 1999, in Alberta, Canada (N = 1293). Using a Cox proportional hazards model, we determined the relative risk (RR) of relapse ED visits among users and nonusers of inhaled corticosteroids after discharge from the ED. We also compared the RR of relapse ED visits across different dose categories. RESULTS: Users of inhaled corticosteroids after ED discharge had 45% fewer relapse ED visits than did nonusers (adjusted RR, 0.55; 95% confidence interval [CI], 0.44-0.69). Low-, medium-, and high-dose therapies were associated with similar reductions in the risk of relapse ED visits: low-dose therapy (RR, 0.52; 95% CI, 0.39-0.68), medium dose therapy (RR, 0.51; 95% CI, 0.34-0.76), and high-dose therapy (RR, 0.67; 95% CI, 0.47-0.94). CONCLUSIONS: Inhaled corticosteroid therapy after ED discharge is associated with a significant reduction in the risk of subsequent ED visits. Low dose therapy appears to be as effective as high-dose therapy. However, further studies are needed to determine the optimal dosing regimen for inhaled corticosteroid therapy for asthma. PMID- 12123403 TI - The outcomes and costs of acute myeloid leukemia among the elderly. AB - BACKGROUND: The incidence of acute myeloid leukemia (AML) among the elderly can be expected to grow as the population continues to age. However, data on current treatment practices and costs for this form of cancer are sparse. METHODS: We used a retrospective inception cohort design and data from a linkage between 11 Surveillance, Epidemiology, and End Results cancer registries and Medicare administrative claims. We evaluated survival, use of health care resources, use of chemotherapy, and Medicare payments among adults 65 years and older with an initial diagnosis of AML between January 1, 1991, and December 31, 1996. RESULTS: A total of 2657 elderly patients with AML and complete Medicare claims data were identified. The prognosis for these patients was poor, with median survival estimated to be 2 months and a 2-year survival rate of 6%. Mean +/- SE total Medicare payments were $41,594 +/- $870 (in 1998 US dollars), 84% of which was attributed to inpatient payments. In the 2 years after the AML diagnosis, 790 patients (30%) underwent chemotherapy treatment. These patients had costs almost 3 times higher than those of other patients, and their median survival was 6 months longer. The use of hospice care was rare (17% of patients). CONCLUSIONS: Among the elderly, AML is associated with a poor prognosis and substantial costs during the relatively few remaining months of life. Moreover, most patients do not receive active treatment with chemotherapy or hospice services. Further work is needed to characterize this disease and the patient-related factors that influence treatment decisions and associated health outcomes. PMID- 12123404 TI - Religious involvement and cigarette smoking in young adults: the CARDIA study (Coronary Artery Risk Development in Young Adults)study (. AB - BACKGROUND: Results of previous studies have suggested that involvement in religious activities may be associated with lower rates of smoking. We sought to determine whether frequent attendance at religious services is associated with less smoking among young adults. METHODS: This prospective cohort study of 4569 adults aged 20 to 32 years included approximately equal numbers of blacks and whites and men and women from 4 cities in the United States who attended the 1987/1988 examination of the Coronary Artery Risk Development in Young Adults (CARDIA) study. Frequency of attendance at religious services and denominational affiliation were determined by self-report questionnaire in 1987/1988. Cigarette smoking was determined by interview at this time and again 3 years later. RESULTS: Of 4544 participants who completed the tobacco questionnaire in 1987/1988, 34% (891/2598) who attended religious services less than once per month or never and 23% (451/1946) who attended religious services at least once per month reported current smoking (odds ratio [OR], 1.7; 95% confidence interval [CI], 1.5-2.0; P<.001). This association between less frequent attendance at religious services and current smoking was found in most denominations and remained significant after adjusting for potential confounding variables (OR, 1.5; 95% CI, 1.3-1.8; P<.001). During 3-year follow-up, nonsmokers who reported little or no religious involvement had an increased risk of smoking initiation (adjusted OR, 1.9; 95% CI, 1.3-2.7; P<.001). CONCLUSIONS: Young adults who attend religious services have lower rates of current and subsequent cigarette smoking. The potential health benefits associated with religious involvement deserve further study. PMID- 12123406 TI - Dietary sodium intake and incidence of congestive heart failure in overweight US men and women: first National Health and Nutrition Examination Survey Epidemiologic Follow-up Study. AB - BACKGROUND: Cross-sectional epidemiologic studies suggest that a higher intake of dietary sodium is associated with an increased risk of left ventricular hypertrophy. We studied the relationship between dietary sodium intake and incidence of congestive heart failure (CHF) in the first National Health and Nutrition Examination Survey Epidemiologic Follow-up Study participants. PARTICIPANTS AND METHODS: The study sample consisted of 5233 nonoverweight and 5129 overweight men and women without a history of CHF at their baseline examination. Dietary sodium and other nutrient intake estimates were obtained by a 24-hour dietary recall method at the baseline examination, conducted from 1971 to 1975. The incidence of CHF was assessed using medical records and death certificates obtained in 1982 to 1984, 1986, 1987, and 1992. RESULTS: During an average of 19 years of follow-up, we documented 413 cases of CHF in nonoverweight and 679 cases of CHF in overweight participants. After adjustment for known CHF risk factors, the relative risk of CHF among overweight participants was 1.43 (95% confidence interval, 1.07-1.91) for those whose sodium intake was greater than 113.6 mmol/d compared with those whose intake was less than 50.2 mmol/d. The relative risks of CHF for a 100-mmol/d higher intake of sodium or per 1743 kcal (average energy intake in the study population) were 1.26 (95% confidence interval, 1.03-1.53) and 1.21 (95% confidence interval, 1.04-1.40), respectively. CONCLUSIONS: A higher intake of dietary sodium is a strong independent risk factor for CHF in overweight persons. A reduction in sodium intake may play an important role in the prevention of CHF in overweight individuals and populations. PMID- 12123405 TI - Early intervention in planning end-of-life care with ambulatory geriatric patients: results of a pilot trial. AB - BACKGROUND: A large discrepancy exists between the wishes of dying patients and their actual end-of-life care. However, retrospective clinical experience suggests that early advance care planning (ACP) can markedly reduce this discrepancy. This article describes a randomized trial to evaluate the short-term clinical utility of early ACP. We also assessed the feasibility of performing a larger prospective study to document long-term outcomes. METHODS: Ambulatory geriatric patients (N = 61) were randomized to either a control group, which received only a Massachusetts Health Care Proxy form to complete, or an intervention group, in which each patient and health care agent discussed ACP with a trained nurse facilitator. The benefits and burdens of life-sustaining treatments were discussed, and patient goals and preferences for these treatments were documented. RESULTS: Two-month follow-up revealed that the intervention achieved higher congruence between agents and patients in their understanding of patients' end-of-life care preferences, with 76% (19/25) in complete agreement vs 55% (12/22) of the controls (effect size [ES] = -0.43). There was also a greater increase in patient knowledge about ACP in the intervention group (ES = 0.22). Intervention patients became less willing to undergo life-sustaining treatments for a new serious medical problem (ES = -0.25), more willing to undergo such treatments for an incurable progressive disease (ES = 0.24), and less willing to tolerate poor health states (ES = -0.78). Practical insights were gained about how to conduct a larger study more effectively. CONCLUSION: A facilitated discussion about end-of-life care between patients and their health care agents helps define and document the patient's wishes for both patient and agent. PMID- 12123407 TI - Predictive factors of malaria in travelers to areas where malaria is endemic. AB - BACKGROUND: The differentiation of malaria from other causes of fever is difficult. The development of tools for rapid and specific clinical diagnosis is of paramount importance for the identification of individuals infected with malaria. METHOD: A 4-year prospective study to identify the clinical and biological variables associated with malaria included all patients suspected of having malaria who presented in the emergency department (ED) of a French hospital. RESULTS: Of 783 patients admitted to the ED with suspected malaria, 145 had positive findings of a thick smear for Plasmodium species, mainly Plasmodium falciparum (90.3%). In univariate analysis, the following 12 variables were significantly associated with diagnosis of malaria: older than 30 years, male sex, immigration to France from an area where malaria is endemic, a visit to sub Saharan Africa, insufficient antimalaria prophylaxis, fever, chills, absence of diarrhea, a leukocyte count within the reference range, thrombocytopenia, and increased lactate dehydrogenase and bilirubin levels. In multivariate analysis, the factors predictive of malaria included a visit to sub-Saharan Africa (odds ratio [OR], 7.7; 95% confidence interval [CI], 2.8-21.3), a temperature of at least 38.5 degrees C (OR, 6.2; 95% CI, 2.8-13.3), chills (OR, 3.0; 95% CI, 1.4 6.6), thrombocytopenia (OR, 16.5; 95% CI, 7.1-38.3), and abnormally high total bilirubin levels (OR, 21.5; 95% CI, 6.4-72.5). However, alone or combined, these features had insufficient sensitivity (95.0%) and low specificity (55.0%) for the diagnosis of malaria. CONCLUSIONS: Malaria should be suspected in all patients presenting with complaints after travel to an area where malaria is endemic, and these patients should undergo blood microscopy. PMID- 12123408 TI - Use of a clinical decision rule in combination with D-dimer concentration in diagnostic workup of patients with suspected pulmonary embolism: a prospective management study. AB - BACKGROUND: We designed a diagnostic strategy, based on clinical probability and D-dimer concentration, to select patients who were unlikely to have pulmonary embolism (PE), before further diagnostic workup was performed. The utility and safety of this strategy were evaluated in a prospective management study. METHODS: Consecutive patients with suspected PE had D-dimer testing and clinical probability assessment with a clinical decision rule. Patients with a low probability and a normal D-dimer concentration (<500 ng/mL) were considered not to have PE, and further diagnostic testing and anticoagulant therapy were withheld. In patients with a low probability and elevated D-dimer level or with a moderate or high probability, bilateral compression ultrasonography of the legs was performed. If deep venous thrombosis was detected, venous thromboembolism was diagnosed. If compression ultrasonography was normal, pulmonary angiography was performed. All patients were followed up for 3 months. RESULTS: Of the 234 consecutive patients, 26% had the combination of a low probability and normal D dimer level. During the follow-up period, none of these patients died and 3 patients had recurrent complaints of PE. In these 3 patients, PE was excluded by objective testing. The 3-month thromboembolic risk was therefore 0% (95% confidence interval, 0%-6%). The prevalence of PE in the entire population was 22%. CONCLUSIONS: The combination of a low clinical probability and a normal D dimer concentration appears to be a safe method to exclude PE, with a high clinical utility, and is readily accepted by clinicians. PMID- 12123409 TI - Successful blood pressure control in the African American Study of Kidney Disease and Hypertension. AB - BACKGROUND: The African American Study of Kidney Disease and Hypertension (AASK) is an ongoing trial to evaluate the effect of blood pressure and choice of antihypertensive drug on the rate of decline of renal function. OBJECTIVE: To present the success of the AASK in achieving the trial's rigorous blood pressure goals in an extremely challenging patient population. METHODS: The AASK participants included African American patients with hypertension (n = 1094), aged 18 to 70 years, with glomerular filtration rates between 20 and 65 mL/min per 1.73 m(2) and no other identified causes of renal insufficiency. Participants were randomized to a goal mean arterial blood pressure (MAP) of either 102 to 107 mm Hg (usual MAP goal) or 92 mm Hg or less (low MAP goal). Participants in each of these groups were also randomized (double-blind) to a regimen containing metoprolol succinate, ramipril, or amlodipine besylate. Additional agents were added, if required, in the following recommended order: furosemide, doxazosin mesylate, clonidine hydrochloride, or hydralazine hydrochloride (or minoxidil, if needed). RESULTS: In participants randomized to the low MAP goal, the percentage of participants who achieved a blood pressure of less than 140/90 mm Hg increased from a baseline of 20.0% to 78.9% by 14 months after randomization. For usual MAP goal participants, the corresponding percentages increased from 21.5% to 41.8%. The difference in median levels of MAP between the 2 MAP goal groups increased and remained at approximately 12 mm Hg. Blood pressure reduction was similar regardless of age, sex, body mass index, education, insurance or employment status, income, or marital status. CONCLUSION: The blood pressure goals set and achieved in AASK participants clearly demonstrate that adequate blood pressure control can be achieved even in hypertensive populations whose blood pressure is the most difficult to control. PMID- 12123410 TI - Massive bleeding on a bladder protectant: a case report of pentosan polysulfate sodium-induced coagulopathy. AB - Pentosan polysulfate sodium (Elmiron; Alza Pharmaceuticals, Mountain View, Calif) is an oral preparation of pentosan polysulfate used in the symptomatic management of interstitial cystitis. While pentosan polysulfate has a known heparin-like effect in its parenteral form, there have been no previous reports of coagulopathy with oral use. We present an interesting case of inadvertent systemic anticoagulation resulting in serious bleeding complications in a young woman taking oral pentosan polysulfate for interstitial cystitis. PMID- 12123411 TI - Conflict of interest: authorship issues predominate. PMID- 12123413 TI - Care provided by subspecialists working outside their specialty: the importance of hospitalist training. PMID- 12123414 TI - Acute necrotizing encephalopathy by Mycoplasma pneumoniae infection? PMID- 12123415 TI - Monoamine oxidase deficiency: a cause of symptomatic hyperserotoninemia in the absence of carcinoid. PMID- 12123419 TI - The therapeutic prescription for the transplant patient. PMID- 12123417 TI - A reversibly dissociable ternary complex formed by XpsL, XpsM and XpsN of the Xanthomonas campestris pv. campestris type II secretion apparatus. AB - The cytoplasmic membrane proteins XpsL, XpsM and XpsN are components required for type II secretion in Xanthomonas campestris pv. campestris. We performed metal chelating chromatography to partially purify the His(6)-tagged XpsM (XpsMh) containing complex. Immunoblot analysis revealed that both XpsL and XpsN co eluted with XpsMh. The co-fractionated XpsL and XpsN proteins co-immune precipitated with each other, suggesting the existence of an XpsL-XpsM-XpsN complex. Ternary complex formation does not require other Xps protein components of the type II secretion apparatus. Further purification upon size-exclusion chromatography revealed that XpsN is prone to dissociate from the complex. Reassociation of XpsN with the XpsL-XpsMh complex immobilized on a nickel column is more effective than with XpsMh alone. Membrane-mixing experiments suggested that the XpsL-XpsMh complex and XpsN probably dissociate and reassociate in the membrane vesicles. Comparison of the half-lives of the XpsL-XpsMh-XpsN and XpsL XpsMh complexes revealed that XpsL dissociates from the latter at a faster rate than from the former. Dissociation and reassociation between XpsL and XpsM were also demonstrated with membrane-mixing experiments. A dynamic model is proposed for the XpsL-XpsM-XpsN complex. PMID- 12123420 TI - Risk factors for the development of invasive fungal infections in allogeneic blood and marrow transplant recipients. AB - Blood and marrow transplantation (BMT) is increasingly used to treat malignant and nonmalignant diseases. Despite significant advances in the management of transplant recipients, however, fungal infections remain important life threatening complications of BMT. Over the past two decades, the incidence of fungal infections in this population has continued to rise. Several factors predispose BMT recipients to invasive fungal infections. These include but are not limited to use of intensive myeloablative chemotherapy and radiation therapy combined with prolonged granulocytopenia; development of acute and chronic graft versus-host disease; administration of immunosuppressive therapy, particularly using corticosteroids; use of central venous catheters; and prolonged impairment of cell-mediated immunity secondary to the underlying disease and post-transplant immunodeficiency. Environmental factors also play a key part in the pathogenesis of fungal infections. Therefore, infection-control measures are critical to the prevention of such infections. In addition, although Candida and Aspergillus species are still the major culprits, other opportunistic fungi have emerged in recent years. PMID- 12123418 TI - Endothelial nitric oxide synthase (NOS) deficiency affects energy metabolism pattern in murine oxidative skeletal muscle. AB - Oxidative capacity of muscles correlates with capillary density and with microcirculation, which in turn depend on various regulatory factors, including NO generated by endothelial nitric oxide synthase (eNOS). To determine the role of eNOS in patterns of regulation of energy metabolism in various muscles, we studied mitochondrial respiration in situ in saponin-permeabilized fibres as well as the energy metabolism enzyme profile in the cardiac, soleus (oxidative) and gastrocnemius (glycolytic) muscles isolated from mice lacking eNOS (eNOS(-/-)). In soleus muscle, the absence of eNOS induced a marked decrease in both basal mitochondrial respiration without ADP (-32%; P <0.05) and maximal respiration in the presence of ADP (-29%; P <0.05). Furthermore, the eNOS(-/-) soleus muscle showed a decrease in total creatine kinase (-29%; P <0.05), citrate synthase ( 31%; P <0.01), adenylate kinase (-27%; P <0.05), glyceraldehyde-3-phosphate dehydrogenase (-43%; P <0.01) and pyruvate kinase (-26%; P <0.05) activities. The percentage of myosin heavy chains I (slow isoform) was significantly increased from 24.3+/-1.5% in control to 30.1+/-1.1% in eNOS(-/-) soleus muscle ( P <0.05) at the expense of a slight non-significant decrease in the three other (fast) isoforms. Besides, eNOS(-/-) soleus showed a 28% loss of weight. Interestingly, we did not find differences in any parameters in cardiac and gastrocnemius muscles compared with respective controls. These results show that eNOS knockout has an important effect on muscle oxidative capacity as well on the activities of energy metabolism enzymes in oxidative (soleus) muscle. The absence of such effects in cardiac and glycolytic (gastrocnemius) muscle suggests a specific role for eNOS-produced NO in oxidative skeletal muscle. PMID- 12123421 TI - Antiviral prophylaxis may prevent human herpesvirus-6 reactivation in bone marrow transplant recipients. AB - Human herpesvirus-6 (HHV-6) infects the majority of children under the age of 2 years causing roseola infantum. Following short self-limited disease, the virus enters into a latency phase in peripheral blood lymphocytes (PBL). It has been previously reported that HHV-6 reactivation from latency, in immunocompromised patients undergoing bone marrow transplantation (BMT), could result in febrile illness, pneumonitis, meningitis, and/or encephalitis. In our study, 14 BMT patients received two different antiviral prophylactic therapies: 8 patients received acyclovir, whereas 6 patients received ganciclovir. Clinical manifestations and virus recovery were monitored pre- and post-BMT by polymerase chain reaction tests of cord blood cells cultured with the patients' PBL. No HHV 6 recovery was shown in the 6 patients treated with ganciclovir, whereas 3 of the 8 acyclovir-treated patients experienced virus reactivation 20-21 days post-BMT. One of the 3 patients was asymptomatic but had late engraftment; the second patient had prolonged fever, skin rash, and hemorrhage; the third patient experienced prolonged fever, pneumonitis, marrow rejection, and fatal encephalitis. It is concluded that viral reactivation may be prevented by prophylactic treatment with ganciclovir. Our observation awaits further documentation in prospective randomized trials in high-risk BMT recipients. PMID- 12123422 TI - Effects of changing immunosuppressive regimen on the incidence, duration, and viral load of cytomegalovirus infection in renal transplantation: a single center report. AB - Background. In this retrospective single center study we have evaluated the relation between the immunosuppressive regimen and the incidence and characteristics of cytomegalovirus (CMV) infection in the setting without CMV prophylaxis from 1989 through 1998. Methods. All (470) first cadaveric renal transplantations in nonsensitized (PRA < 60%) patients were analyzed. Immunosuppression consisted of cyclosporine A (Sandimmune) and prednisolone from 1989 through 2-1993 (S; 189 patients), of cyclosporine microemulsion (Neoral) and prednisolone from 3-1993 through 5-1997 (N; 200 patients) and of mycophenolate mofetil, Neoral and prednisolone from 5-1997 until 1998 (M; 81 patients). The CMV pp65-antigenemia was measured routinely at least once weekly from day 10 till 12 weeks after transplantation or until pp65-antigenemia became negative. No CMV prophylaxis was given. Results. By changing from Sandimmune to Neoral and by adding mycophenolate mofetil, respectively, we observed a higher frequency of especially secondary CMV infections (S vs. N vs. M, respectively, 28 vs. 50 vs. 63%, P = 0.026; S vs. N, P = 0.027; S vs. M, P = 0.015; and N vs. M, n.s). The CMV infections lasted longer (median duration antigenemia S vs. N vs. M, respectively, 3 vs. 5 vs. 7 weeks, P = 0.0003; S vs. N, P < 0.002; S vs. M, P < 0.001; and N vs. M, P < 0.05). Viral load was higher in M (median maximal pp65 antigenemia S vs. N vs. M, respectively, 19 vs. 14.5 vs. 73, P < 0.01; S vs. N, n.s.; S vs. M, P < 0.001 and N vs. M, P < 0.01). Conclusions. The use of Neoral and the addition of mycophenolate mofetil caused significant changes in the incidence, duration and viral load of CMV infections. PMID- 12123423 TI - Safety and tolerability of caspofungin acetate in the treatment of fungal infections. AB - Caspofungin acetate is the first member of the novel echinocandin class of antifungal drugs to be marketed in the United States. It has recently been approved for use in patients with invasive aspergillosis who are refractory to or intolerant of conventional therapy. Accordingly, its safety profile is particularly important to review. The safety and tolerability of caspofungin have been examined in 623 persons, including 295 patients who received >/= 50 mg/day for at least one week in clinical studies. In the 263 patients, given caspofungin in randomized double-blind active-control trials to date, there have been no serious clinical or laboratory drug-related adverse events; caspofungin was discontinued in only 2% of these patients because of drug-related adverse experiences. Caspofungin may have potentially important drug interactions with cyclosporine and tacrolimus. PMID- 12123424 TI - Cellular immune responses in transplantation-associated chronic viral infections. AB - Viral pathogens are important causes of morbidity following transplantation. Cytomegalovirus (CMV) and Epstein-Barr virus (EBV) infections represent two major viral complications in transplant recipients. Recent advances in methodology have led to a better understanding of host immune responses directed against chronic viral infections. We review the nature of antiviral immunity involved in control of CMV and EBV. Viral mechanisms of immune evasion and immunotherapeutic strategies in the transplantation setting will also be addressed. PMID- 12123425 TI - Legionellosis in a lung transplant recipient obscured by cytomegalovirus infection and Clostridium difficile colitis. AB - A 52-year-old-white male underwent double lung transplantation for severe emphysema due to alpha-1-antitrypsin deficiency and heavy tobacco use. Following a postoperative course complicated by renal insufficiency, pulmonary emboli, and Clostridium difficile colitis, he was discharged in stable condition. Two months later, he was admitted to a local hospital with a fever, abdominal pain, diarrhea, nausea, and dyspnea. Computerized tomography (CT) of the chest revealed bilateral pleural effusions. Sigmoidoscopy was grossly normal but biopsy demonstrated cytomegalovirus (CMV) colitis, and the patient was placed on intravenous ganciclovir. Over the next week, he became progressively hypoxemic and was transferred to the University of Pittsburgh Medical Center (post transplant day 81) for further evaluation. His medications on transfer included: ganciclovir, prednisone, tacrolimus, dapsone, fluconazole, ondansetron, lansoprazole, digoxin, and coumadin. PMID- 12123427 TI - Rhodococcus equi infection in transplant recipients: a case of mistaken identity and review of the literature. AB - The incidence of Rhodococcus equi infection in solid-organ transplant recipients continues to rise throughout the world. Unfortunately, this opportunistic pathogen is still underestimated and potentially disregarded by physicians and microbiology laboratories due to its morphology on Gram staining. Pulmonary involvement is the most common finding in the immunocompromised host. We report a case of a 63-year-old heart-transplant recipient who presented with increasing fatigue and nonproductive cough for 3 weeks. After full evaluation, a lung abscess was demonstrated by thoracic computerized tomography (CT). Blood and sputum cultures were remarkable for heavy "diphtheroids." Although the Gram-stain result was initially interpreted as a contaminant, a clinical suspicion for Rhodococcus assisted in further investigation. Broncheoalveolar lavage and CT guided biopsy of the lung abscess revealed heavy growth of diphtheroids. However, further evaluation by a reference laboratory demonstrated mycolic acid staining consistent with R. equi. Surgical drainage and prolonged antibiotic therapy resulted in complete remission of the pneumonia and abscess. This represents the fourth reported case of R. equi infection in a heart transplant recipient. It is imperative that all physicians and laboratory staff consider R. equi when an immunocompromised patient has any type of pneumonia, especially with abscess formation. PMID- 12123426 TI - Rhodococcus equi pulmonary infection in a pancreas-alone transplant recipient: consequence of intense immunosuppression. AB - We report the case of a pancreas-alone transplant recipient who developed Rhodococcus equi pneumonia after receiving multiple courses of antilymphocyte therapy for the treatment of recurrent acute pancreas allograft rejection. We also review and discuss the diagnosis, clinical course, and treatment of 18 cases of R. equi infection reported in solid organ transplant recipients. The lung is the most common primary site of infection, but R. equi infection is difficult to diagnose because of the pleomorphic, gram-positive, and partially acid-fast nature of the organism. Treatment usually involves a combination of antibiotics including rifampin, macrolides, vancomycin, and ciprofloxacin. The optimal duration of therapy is unknown, but relapse is common if the duration of treatment is less than 14 days. The duration of therapy should be guided by clinical recovery, culture results, and radiographic findings. Monitoring levels of immunosuppressive agents-such as tacrolimus and cyclosporine-is needed in order to avoid clinically significant drug interactions with rifampin or the macrolides when these agents are used in order to treat R. equi infection in the transplant population. PMID- 12123429 TI - Negative discovery. PMID- 12123431 TI - Is birthweight a good marker for gestational exposures that increase the risk of adult disease? AB - There is much evidence of a link between low birthweight and elevated risk of adult cardiovascular disease, from humans and experimental animals. However, if one relies on data linking birthweight to coronary heart disease to estimate the public health implications of this association, the effects are likely to be modest. The focus on birthweight may be misplaced, because reduced size at birth may not be in the causal pathway linking gestational factors to disease in adult offspring. We need to know more about this before we can estimate the public health implications of gestational factors and assess the potential for intervention. The most studied gestational factor is maternal nutrition. We review here evidence for and against birthweight being in the causal pathways between suboptimal maternal nutrition and increased risk of adult disease in the offspring and provide evidence suggesting that birthweight is not in all of them. From a public health point of view, we suggest that future research in this field should focus on modifiable gestational exposures that may be linked to adult disease, whether or not they influence size at birth. PMID- 12123432 TI - Fetal growth and systolic blood pressure in young adulthood: the Swedish Young Male Twins Study. AB - The objective was to test the fetal origins hypothesis by examining the association between fetal growth and systolic blood pressure in a large cohort of adult male twins of known zygosity. This cohort study is based on a record linkage of routinely collected data from the Swedish Medical Birth Registry, the Swedish Military Conscription Registry and a mailed questionnaire. It comprises 886 pairs of same-sex male twins born in Sweden in 1973-79 for whom data were available on size at birth and systolic blood pressure, height and weight at age 17-19 years. Based on self-reported similarity of appearance in childhood, 384 twin pairs were classified as monozygotic (MZ), 269 as dizygotic (DZ) and 233 as of uncertain zygosity (XZ). Differences in systolic blood pressure within and between twin pairs were analysed in relation to birthweight and birthweight-for gestational-age z-score using random effects linear models. Within twin pairs, a 1 kg difference in birthweight was associated with a difference of -1.30 mmHg in systolic blood pressure [95% confidence interval -4.15, +1.54] for MZ twins and +0.14 [-3.49, +3.76] for DZ twins; for all twins combined it was -0.21 [-2.13, +1.71]. Between twin pairs, a 1 kg difference in birthweight was associated with a difference of -2.68 mmHg in systolic blood pressure [-4.95, -0.42] for MZ twins and +0.28 [-2.35, +2.91] for DZ twins; for all twins combined -1.68 [-3.15, 0.22]. All these estimates included adjustment for age, year and conscription centre of examination, gestational age and height and weight at conscription. These results provide little support for the fetal origins hypothesis. The estimates of the within-pair effect in MZ twins and their wide confidence intervals (including zero) cannot exclude the existence of common genetic mechanisms, but are also not inconsistent with an in utero programming effect. This study highlights some methodological problems with twin studies. PMID- 12123433 TI - Childhood leukaemia in Costa Rica, 1981-96. AB - Childhood leukaemia incidence in Costa Rica during 1981-96, among the highest in the world, was analysed by histology, gender, birth year, time period of diagnosis, age at diagnosis and region. Numbers of cases were extracted from the database of the National Cancer Registry (RNT) of Costa Rica. Person-years at risk were calculated from census data and post-census population estimates. During the follow-up, 918 cases of leukaemia in children under 15 years (510 boys, 408 girls) were reported to the RNT (41% of all childhood malignancies), with an overall age-standardised incidence rate of 56 per million person-years. Acute lymphocytic leukaemia (ALL) represented 79% and acute non-lymphocytic leukaemia (ANLL) 16% of the cases, with rates of 43 and 9 per million person years respectively. There were downward trends in incidence of total leukaemias, ALL and ANLL and 'not otherwise specified' (NOS) combined. Incidence of ALL was highest at 1-4 years of age in boys and girls, whereas ANLL peaked in girls during the first year of life. During 1991-96, the decrease in ALL was significant (P = 0.042). A multivariable Poisson regression model identified significant excesses of ALL for boys, for age groups 1-4 and 5-9 years and for three out of seven regions. Possible reasons for the high rates in Costa Rica are discussed. PMID- 12123434 TI - Central nervous system tumours in children in Costa Rica, 1981-96. AB - Incidence rates of malignant central nervous system (CNS) tumours in children in Costa Rica are presented in an international perspective. For the 16-year period 1981-96, a total of 256 CNS tumours were registered in children below age 15 years by the National Tumour Registry of Costa Rica. The age-standardised incidence rate was 15.2 per million person-years, with a male-to-female ratio of 1.4. The median age-standardised incidence rates of selected registries in other Latin American countries were 19.3, in other developing countries 12.0 and in industrialised countries 29.6 per million person-years. The comparatively low incidence rates in Costa Rica were evident in all diagnostic subgroups, most notably in the youngest age group and for tumours in the brain stem. In the Central Valley, where the capital and the only specialised paediatric hospital are situated, the crude incidence rate was 18.1 [95% CI 15.1, 21.1] compared with 10.5 [95% CI 8.3, 12.8] per million person-years in the rest of the country (RR = 1.7, 95% CI 1.3, 2.3). There was no evidence of any increase over time. The data in this study cannot exclude under-diagnosis and, to a lesser degree, under registration as a partial explanation of the low incidence rates of malignant CNS tumours in children in Costa Rica. PMID- 12123435 TI - West Coast study of childhood brain tumours and maternal use of hair-colouring products. AB - The immature nervous system of the fetus is characterised by rapid cell growth and division and is particularly vulnerable to carcinogens and mutagens. Several epidemiological studies have reported an increased risk for childhood brain tumours (CBT) associated with exposure to N-nitroso compounds (NOC). Hair colouring products (hair "dyes") that contain NOC-related aromatic amines have shown mutagenicity in vitro and carcinogenic properties in vivo. The potential public health impact of the relationship between hair dye use and carcinogenesis has prompted epidemiological research, given that a large proportion of American women have used hair dyes. A large population-based case-control study was conducted on the west coast of the USA to investigate risk factors for CBT including exposure to NOC. Eligible CBT patients (<20 years of age and diagnosed between 1984 and 1991) were identified from cancer registries in Los Angeles County, the San Francisco Bay Area in California and the Seattle area in Washington state. A total of 540 biological mothers of these children were interviewed, and 801 control subjects who were frequency matched to the CBT patients on birth year and sex were obtained using random digit dialling. Mothers were asked details about personal use of hair dyes during the index pregnancy including frequency of use, trimester of use and type of dye used. Results from age- and sex-adjusted unconditional logistic regression analyses showed no association between risk for CBT and use of hair dyes 1 month before and/or during pregnancy nor during specific trimesters. A nearly twofold increased risk for CBT was associated with single-interval use during the 1 month before pregnancy, but the confidence interval (CI) was imprecise and the estimate was not different from unity (OR = 1.9, 95% CI [0.5, 7.0]). Exclusive use of permanent dye, temporary dye or hair darkeners was not associated with risk for CBT. A twofold increased risk (OR = 2.0, 95% CI [0.83, 4.7]) was observed with exclusive use of semi-permanent dye during the month before or during pregnancy. Exclusive use of semi-permanent dye during the month before pregnancy and/or first trimester also was associated with an elevated risk for CBT, again not different from unity and with an imprecise CI (OR = 2.5, 95% CI = [0.58, 10.3]). There was no evidence of an association between risk for CBT by histological subtypes and use of hair dyes during the index pregnancy or the month before conception. Together with results from previous studies, these results provide no consistent evidence of an association between risk for CBT and use of hair dyes during pregnancy. PMID- 12123436 TI - Maternal, paternal and environmental tobacco smoking and breast feeding. AB - The effects of environmental tobacco smoke (ETS) on breast-feeding patterns are poorly understood, while those of parental smoking on breast-feeding initiation vs. duration have not been clearly delineated. We conducted a prospective, population-based birth cohort study to examine the independent effects of maternal, paternal and ETS on breast-feeding initiation and duration. A total of 6747 Hong Kong Chinese infants were recruited and followed up in 1997-8. We obtained detailed household smoking history and recorded breast-feeding patterns in three follow-up interviews over 9 months. We found that both maternal and paternal smoking were associated with not initiating breast feeding (odds ratio [OR] for ever maternal smoking = 2.51, 95% confidence interval [CI] = 1.63, 3.86; OR for ever paternal smoking = 1.22, 95% CI = 1.08, 1.39). Exposure to ETS in utero and post partum were also related to not starting breast feeding (OR(ETS in utero) = 1.10, 95% CI = 0.99,1.24; OR(ETS post partum) = 1.21, 95% CI = 1.08, 1.36). These effects, however, did not persist for breast-feeding duration of < or = 4 months. Cox proportional hazards modelling confirmed the lack of association between any form of smoking and breast-feeding duration. Our findings suggest that smoking of any kind, during or after pregnancy, is a strong risk indicator for not initiating breast feeding. Smoking as a risk indicator for underlying socio-economic, demographic and psychosocial factors is probably responsible for most of the observed adverse effects, although we cannot rule out direct contributions from pathophysiological mechanisms. Public health strategies directed at these underlying factors should be vigorously pursued to reduce the adverse effects of tobacco on breast feeding and infant health in general. PMID- 12123437 TI - Prevalence of cigarette smoking in pregnant women participating in the special supplemental nutrition programme for Women, Infants and Children (WIC) in Minneapolis and Saint Paul, Minnesota, USA. AB - Several adverse birth outcomes are associated with cigarette smoking. It is important to determine the prevalence of cigarette smoking among pregnant low income women and to evaluate their smoking cessation patterns in order to target appropriate interventions. Ethnically diverse pregnant women aged 15-45 years were recruited from Minneapolis or Saint Paul Women, Infants, and Children (WIC) clinics before their third trimester. Serum cotinine levels were assayed for 98 women and compared with self-report. The women were unaware that their smoking status would be validated. Twenty-one (21%) women had a positive serum cotinine value (> or =3 ng/mL); 16 (76%) admitted smoking within the previous 24 h before interview and five denied smoking. Of the five, four had cotinine levels that could suggest passive smoke exposure. Thirty-seven women (38%) admitted cigarette smoking during the pregnancy but before knowing that they were pregnant; 18 (49%) of these denied current smoking at the interview and also presented with negative cotinine levels. These data suggest that some participants in WIC make a concerted effort to quit smoking when they find out they are pregnant, and are generally truthful when reporting their smoking habits during pregnancy. PMID- 12123438 TI - Deprivation and stillbirth risk in rural and urban areas. AB - The objective of the study was to investigate whether stillbirth risk was higher, and the effect of deprivation on inequality in stillbirth risk more marked, in rural than in urban areas. We carried out a cohort study of all 280 757 singleton births to mothers domiciled in Cumbria, north-west England, 1950-92. After allowing for individual social class and community deprivation, the risk of stillbirth was lower outside urban centres both during 1950-65 (OR = 0.91, 95% CI 0.84, 1.00) and during 1966-92 (OR = 0.82, 95% CI 0.73, 0.92). In earlier years, unsupported mothers in rural areas and mothers living in remote areas were particularly at risk. Urban/rural differences in risk were not explained by individual social class, community deprivation or overcrowding and have persisted over a 40-year time period. PMID- 12123439 TI - Socio-demographic determinants of poor infant outcome in north-west Russia. AB - The infant outcome determinants vary in different settings, and there is still a need for analysis within environments. This study was designed to examine the relation between poor infant outcome (PIO), (i.e. any of the following indicators: preterm delivery, low birthweight, perinatal death and low first minute Apgar score) and socio-demographic factors, smoking and alcohol consumption in a Russian setting. The study was conducted in the town of Severodvinsk, north-west Russia. A total of 1404 pregnant women who attended antenatal care clinics in 1999 and delivered at the municipal maternity home comprised the cohort. Data on women and infants were collected from the medical files and a questionnaire on social indicators, smoking and alcohol consumption was administered. The adjusted odds ratio (OR) calculated by multivariable logistic regression was used as the measure of association between PIO and the variables studied. Education was found to be the most significant factor associated with PIO (OR = 1.9, 95% CI 1.2, 3.0 for secondary or less education compared with at least 3 years of university studies). Increased risks of PIO were also found in mothers aged 30 years and older (OR = 1.6, 95% CI 1.1, 2.5 compared with other age groups) and in unmarried mothers (OR = 1.4, 95% CI 1.0, 1.9) after control for the possible confounders. Other socio-demographic determinants studied (smoking, alcohol use, stress, maternal occupation, housing and young age of the mother) could not be found to influence PIO. The findings contribute to the hypothesis that maternal education is one of the most important social factors influencing pregnancy outcomes in countries in transition. PMID- 12123440 TI - Trends and characteristics of induced labour in the United States, 1989-98. AB - Induction of labour is one of the fastest growing medical procedures in the United States. In 1998, 19.2% of all US births were a product of induced labour, more than twice the 9.0% in 1989. Induction of labour has been efficacious in the management of post-term pregnancy and in expediting delivery when the mother or infant is sufficiently ill to make continuation of the pregnancy hazardous. However, the recent rapid increase in induction, and particularly the doubling of the induction rate for preterm pregnancies (from 6.7% in 1989 to 13.4% in 1998), has generated concern among some clinicians. The present study uses vital statistics natality data to examine the epidemiology of induced labour in the US. Multivariable analysis is used to examine the probability of having an induced delivery in relation to a wide variety of socio-demographic and medical characteristics, and also in relation to relative indications and contraindications for induced labour as outlined by the American College of Obstetricians and Gynecologists (ACOG). Induction rates were higher for women who were non-Hispanic white, college educated, born in the US, primaparae and those with intensive prenatal care utilisation. Induction rates were also higher for women with various medical conditions including hypertension, eclampsia and renal disease. For non-Hispanic white women with singleton births, 59% of the increase in the preterm birth rate from 1989 to 1998 can be accounted for by the increase in preterm inductions. The adjusted odds ratio for neonatal mortality among preterm births with induced labour was 1.20 [95% confidence interval 1.11, 1.31]. The rapid increase in induction rates, particularly among preterm births, marks a shift in the obstetric management of pregnancy. More detailed studies are needed to examine physician decision-making protocols, particularly for preterm induction, and to assess the impact of these practice changes on patient outcomes. PMID- 12123441 TI - Growth parameters in newborns with hyperphenylalaninaemia. AB - To understand the effects of hyperphenylalaninaemia on fetal growth, we studied growth parameters (weight, length and head circumference) of 23 phenylketonuric (PKU) and 60 hyperphenylalaninaemic (HPA) newborns from healthy mothers and of 1853 healthy neonates from north-east Italy. A comparison of the growth parameters for both PKU and HPA newborns, as well as for controls, showed a statistically significant higher percentage of PKU and HPA patients with reduced body length and cranial circumference (P < 0.05 for both parameters in affected neonates). The z-scores for all growth parameters regarding both PKU and HPA newborns and controls, and between PKU and HPA newborns according to the Mann Whitney non-parametric test, were statistically significantly lower in PKU newborns than in controls; in contrast, only body length was significantly lower in HPA newborns than in controls (P < 0.01). A comparison of growth parameter z scores using the Kruskal-Wallis test for PKU, HPA and control newborns showed that both body length (P < 0.01) and cranial circumference (P < 0.05) were significantly lower in both groups of affected neonates. Our results showed intrauterine growth retardation for both PKU and HPA newborns. Body length and cranial circumference appeared to be more important than birthweight in evaluating growth of PKU and HPA newborns. PMID- 12123442 TI - Population-based outcomes after acute antenatal transfer. AB - Acute antenatal transfer to specialist centres is an accepted practice but few or no regular data are collected regarding the numbers of transfers performed or subsequent pregnancy outcome. We wished to determine the numbers, and the maternal and fetal outcomes following acute antenatal transfer between consultant obstetric units in the former Northern Region of the UK over a 12-month period (1 January-31 December 99). This is a geographically defined population in terms of provision of perinatal services. All acute antenatal transfers were notified centrally. Data pertaining to each transfer were collected at the time of transfer. Subsequent maternal and fetal outcomes were determined from patient records and neonatal databases. The regional annual acute antenatal transfer rate was 3.7 per 1000 deliveries. Most were for fetal reasons, although transfer rates varied between hospitals. The decision to transfer was influenced by distance and availability of paediatric staff. Even units that have similar characteristics show considerable variation in their transfer rates. No adverse incidents occurred during transfer and no major changes in maternal management occurred following transfer. Twenty-four per cent of women remained undelivered following transfer. Women with preterm labour in the absence of ruptured membranes were less likely to deliver than those transferred for other reasons and if they did deliver, their infants were also less likely to need intensive care. We believe audit of acute antenatal transfers should be routinely undertaken. Numbers of transfers might be reduced if delivery and the need for neonatal intensive care could be predicted with greater accuracy. The psychological and financial costs of transfer to women and healthcare providers need to be addressed. PMID- 12123443 TI - Never say never again: protein glycosylation in pathogenic bacteria. AB - In recent years, accumulating evidence for glycosylated bacterial proteins has overthrown an almost dogmatic belief that prokaryotes are not able to synthesize glycoproteins. Now it is widely accepted that eubacteria express glycoproteins. Although, at present, detailed information about glycosylation and structure function relationships is available for only few eubacterial proteins, the variety of different components and structures observed already indicates that the variations in bacterial glycoproteins seem to exceed the rather limited display found in eukaryotes. Numerous virulence factors of bacterial pathogens have been found to be covalently modified with carbohydrate residues, thereby identifying these factors as true glycoproteins. In several bacterial species, gene clusters suggested to represent a general protein glycosylation system have been identified. In other cases, genes encoding highly specific glycosyltransferases have been found to be directly linked with virulence genes. These findings raise interesting questions concerning a potential role of glycosylation in pathogenesis. In this review, we will therefore focus on protein glycosylation in Gram-negative bacterial pathogens. PMID- 12123444 TI - StcE, a metalloprotease secreted by Escherichia coli O157:H7, specifically cleaves C1 esterase inhibitor. AB - Escherichia coli O157:H7 causes diarrhoea, haemorrhagic colitis, and the haemolytic uraemic syndrome. We have identified a protein of previously unknown function encoded on the pO157 virulence plasmid of E. coli O157:H7, which is the first described protease that specifically cleaves C1 esterase inhibitor (C1 INH), a member of the serine protease inhibitor family. The protein, named StcE for secreted protease of C1 esterase inhibitor from EHEC (formerly Tagn), cleaves C1-INH to produce (unique) approximately 60-65 kDa fragments. StcE does not digest other serine protease inhibitors, extracellular matrix proteins or universal protease targets. We also observed that StcE causes the aggregation of cultured human T cells but not macrophage-like cells or B cells. Substitution of aspartic acid for glutamic acid at StcE position 435 within the consensus metalloprotease active site ablates its abilities to digest C1-INH and to aggregate T cells. StcE is secreted by the etp type II secretion pathway encoded on pO157, and extracellular StcE levels are positively regulated by the LEE encoded regulator, Ler. StcE antigen and activity were detected in the faeces of a child with an E. coli O157:H7 infection, demonstrating the expression of StcE during human disease. Cleavage of C1-INH by StcE could plausibly cause localized pro-inflammatory and coagulation responses resulting in tissue damage, intestinal oedema and thrombotic abnormalities. PMID- 12123445 TI - Gene expression profiling of Escherichia coli growth transitions: an expanded stringent response model. AB - When conditions cause bacterial growth to stop, extensive reprogramming of physiology and gene expression allows for the cell's survival. We used whole genome DNA arrays to determine the system response in Escherichia coli cells experiencing transient growth arrest caused by glucose-lactose diauxie and H2O2 treatment, and also entry into stationary phase. The results show that growth arrested cells induce stringent control of several gene systems. The vast majority of genes encoding the transcription and translation apparatus immediately downregulate, followed by a global return to steady state when growth resumes. Approximately one-half of the amino acid biosynthesis genes downregulate during growth arrest, with the notable exception of the his operon, which transiently upregulates in the diauxie experiment. Nucleotide biosynthesis downregulates, a result that is again consistent with the stringent response. Likewise, aerobic metabolism downregulates during growth arrest, and the results led us to suggest a model for stringent control of the ArcA regulon. The stationary phase stress response fully induces during growth arrest, whether transient or permanent, in a manner consistent with known mechanisms related to stringent control. Cells similarly induce the addiction module anti-toxin and toxin genes during growth arrest; the latter are known to inhibit translation and DNA replication. The results indicate that in all aspects of the response cells do not distinguish between transient and potentially permanent growth arrest (stationary phase). We introduce an expanded model for the stringent response that integrates induction of stationary phase survival genes and inhibition of transcription, translation and DNA replication. Central to the model is the reprogramming of transcription by guanosine tetraphosphate (ppGpp), which provides for the cell's rapid response to growth arrest and, by virtue of its brief half-life, the ability to quickly resume growth as changing conditions allow. PMID- 12123446 TI - A protein kinase specifically associated with proliferative forms of Trypanosoma brucei is functionally related to a yeast kinase involved in the co-ordination of cell shape and division. AB - The life cycle of African trypanosomes is characterized by the alternation of proliferative and quiescent stages but the molecular details of this process remain unknown. Here, we describe a new cytoplasmic protein kinase from Trypanosoma brucei, termed TBPK50, that belongs to a family of protein kinases involved in the regulation of the cell cycle, cell shape and proliferation. TBPK50 is expressed only in proliferative forms but is totally absent in quiescent cells despite the fact that the gene is constitutively transcribed at the same level throughout the life cycle. It is probable that TBPK50 has very specific substrate requirements as it was unable to transphosphorylate a range of classical phosphoacceptor substrates in vitro, although an autophosphorylation activity was readily detectable in the same assays. Complementation studies using a fission yeast mutant demonstrated that TBPK50 is a functional homologue of Orb6, a protein kinase involved in the regulation of cellular morphology and cell cycle progression in yeast. These results link the expression of TBPK50 and the growth status of trypanosomes and support the view that this protein kinase is likely to be involved in the control of life cycle progression and cell division of these parasites. PMID- 12123447 TI - Cyclin-dependent kinase TPK2 is a critical cell cycle regulator in Toxoplasma gondii. AB - The Apicomplexan parasite Toxoplasma gondii replicates by endodyogeny, an unusual form of binary fission. We tested the role of TPK2, a homologue of the CDC2 cyclin-dependent kinases, in cell cycle regulation. TPK2 tagged with HA epitope (TPK2-HA-wt) was expressed in mammalian cells as confirmed by Western blot analysis using HA tag and PSTAIRE antibodies. TPK2-HA-wt phosphorylated a peptide from Histone H1, proving that TPK2 is a functional kinase. TPK2-HA-wt coimmunoprecipitated with mammalian cyclins A, B1, D3 and E. Despite being a functional kinase, TPK2 did not rescue Schizosaccharomyces pombe cdc2 and Saccharomyces cerevisiae cdc28 mutant strains. Overexpression of a dominant negative mutant of TPK2 (TPK2-HA-dn) in T. gondii tachyzoites arrested replication. FACS analysis of tachyzoites expressing TPK2-HA-dn revealed an increase in the fraction of cells in S-phase when compared with TPK2-HA-wt transfected parasites. Expression of TPK2-HA-wt did not arrest tachyzoite replication. No discernable G2 cell cycle block was evident suggesting that cell cycle checkpoints differ in T. gondii from most other eukaryotic cells. These data suggest that TPK2 executes an essential function in T. gondii cell cycle and is likely to be the T. gondii CDC2 orthologue. PMID- 12123448 TI - Molecular characterization of long direct repeat (LDR) sequences expressing a stable mRNA encoding for a 35-amino-acid cell-killing peptide and a cis-encoded small antisense RNA in Escherichia coli. AB - Genome sequence analyses of Escherichia coli K-12 revealed four copies of long repetitive elements. These sequences are designated as long direct repeat (LDR) sequences. Three of the repeats (LDR-A, -B, -C), each approximately 500 bp in length, are located as tandem repeats at 27.4 min on the genetic map. Another copy (LDR-D), 450 bp in length and nearly identical to LDR-A, -B and -C, is located at 79.7 min, a position that is directly opposite the position of LDR-A, B and -C. In this study, we demonstrate that LDR-D encodes a 35-amino-acid peptide, LdrD, the overexpression of which causes rapid cell killing and nucleoid condensation of the host cell. Northern blot and primer extension analysis showed constitutive transcription of a stable mRNA (approximately 370 nucleotides) encoding LdrD and an unstable cis-encoded antisense RNA (approximately 60 nucleotides), which functions as a trans-acting regulator of ldrD translation. We propose that LDR encodes a toxin-antitoxin module. LDR-homologous sequences are not pre-sent on any known plasmids but are conserved in Salmonella and other enterobacterial species. PMID- 12123449 TI - Nac-mediated repression of the serA promoter of Escherichia coli. AB - Escherichia coli and related bacteria contain two paralogous PII-like proteins involved in nitrogen regulation, the glnB product, PII, and the glnK product, GlnK. Previous studies have shown that cells lacking both PII and GlnK have a severe growth defect on minimal media, resulting from elevated expression of the Ntr regulon. Here, we show that this growth defect is caused by activity of the nac product, Nac, a LysR-type transcription factor that is part of the Ntr regulon. Cells with elevated Ntr expression that also contain a null mutation in nac displayed growth rates on minimal medium similar to the wild type. When expressed from high-copy plasmids, Nac imparts a growth defect to wild-type cells in an expression level-dependent manner. Neither expression of Nac nor lack thereof significantly affected Ntr gene expression, suggesting that the activity of Nac at one or more promoters outside the Ntr regulon was responsible for its effects. The growth defect of cells lacking both PII and GlnK was also eliminated upon supplementation of minimal medium with serine or glycine for solid medium or with serine or glycine and glutamine for liquid medium. These observations suggest that high Nac expression results in a reduction in serine biosynthesis. beta-Galactosidase activity expressed from a Mu d1 insertion in serA was reduced approximately 10-fold in cells with high Nac expression. We hypothesize that one role of Nac is to limit serine biosynthesis as part of a cellular mechanism to reduce metabolism in a co-ordinated manner when cells become starved for nitrogen. PMID- 12123450 TI - Role of the extracytoplasmic-function sigma factor sigma(H) in Mycobacterium tuberculosis global gene expression. AB - Like other bacterial species, Mycobacterium tuberculosis has multiple sigma (sigma) factors encoded in its genome. In previously published work, we and others have shown that mutations in some of these transcriptional activators render M. tuberculosis sensitive to various environmental stresses and, in some cases, cause attenuated virulence phenotypes. In this paper, we characterize a M. tuberculosis mutant lacking the ECF sigma factor sigma(H). This mutant was more sensitive than the wild type to heat shock and to various oxidative stresses, but did not show decreased ability to grow inside macrophages. Using quantitative reverse transcription-PCR and microarray technology, we have started to define the sigma(H) regulon and its involvement in the global regulation of the response to heat shock and the thiol-specific oxidizing agent diamide. We identified 48 genes whose expression increased after exposure of M. tuberculosis to diamide; out of these, 39 were not induced in the sigH mutant, showing their direct or indirect dependence on sigma(H). Some of these genes encode proteins whose predicted function is related to thiol metabolism, such as thioredoxin, thioredoxin reductase and enzymes involved in cysteine and molybdopterine biosynthesis. Other genes under sigma(H) control encode transcriptional regulators such as sigB, sigE, and sigH itself. PMID- 12123451 TI - Transfer origins in the conjugative Enterococcus faecalis plasmids pAD1 and pAM373: identification of the pAD1 nic site, a specific relaxase and a possible TraG-like protein. AB - The Enterococcus faecalis conjugative plasmids pAD1 and pAM373 encode a mating response to the peptide sex pheromones cAD1 and cAM373 respectively. Sequence determination of both plasmids has recently been completed with strong similarity evident over many of the structural genes related to conjugation. pAD1 has two origins of transfer, with oriT1 being located within the repA determinant, whereas the more efficiently utilized oriT2 is located between orf53 and orf57, two genes found in the present study to be essential for conjugation. We have found a similarly located oriT to be present in pAM373. oriT2 corresponds to about 285 bp based on its ability to facilitate mobilization by pAD1 when ligated to the shuttle vector pAM401; however, it was not mobilized by pAM373. In contrast, a similarly ligated fragment containing the oriT of pAM373 did not facilitate mobilization by pAD1 but was efficiently mobilized by pAM373. The oriT sites of the two plasmids each contained a homologous large inverted repeat (spanning about 140 bp) adjacent to a series of non-homologous short (6 bp) direct repeats. A hybrid construction containing the inverted repeat of pAM373 and direct repeats of pAD1 was mobilized efficiently by pAD1 but not by pAM373, indicating a significantly greater degree of specificity is associated with the direct repeats. Mutational (deletion) analyses of the pAD1 oriT2 inverted repeat structure suggested its importance in facilitating transfer or perhaps ligation of the ends of the newly transferred DNA strand. Analyses showed that Orf57 (to be called TraX) is the relaxase, which was found to induce a specific nick in the large inverted repeat inside oriT; the protein also facilitated site-specific recombination between two oriT2 sites. Orf53 (to be called TraW) exhibits certain structural similarities to TraG-like proteins, although there is little overall homology. PMID- 12123452 TI - Lon protease functions as a negative regulator of type III protein secretion in Pseudomonas syringae. AB - The central conserved region of the Pseudomonas syringae hrp pathogenicity island encodes a type III protein secretion system (TTSS) that is required for pathogenicity in plants. Expression of the hrp TTSS is controlled by the alternative sigma factor, HrpL, whose expression, in turn, is positively controlled by two truncated enhancer binding proteins, HrpR and HrpS. Although a number of environmental conditions are known to modulate hrp TTSS expression, such as stringent conditions and pathogenesis, the mechanism by which the activities of these transcriptional factors are modulated had not been established. In this study, HrpR and HrpS were found to be constitutively expressed under conditions in which the hrpL promoter was inactive. To identify a postulated negative regulator of hrpL expression, transposome (Tz) mutagenesis was used to isolate hrp constitutive mutants. P. syringae Pss61 and DC3000 hrp constitutive mutants were identified that carried lon::Tz insertions and exhibited increased cell length and UV sensitivity typical of Delta lon mutants. The P. syringae Lon protease retained structural features of its homologues found in other bacteria and was capable of complementing an Escherichia coli Delta lon mutant. P. syringae lon::Tz mutants exhibited enhanced expression of the hrpL promoter, suggesting an effect on HrpR and/or HrpS. HrpR was observed to be unstable in wild-type P. syringae strains grown in non-inductive media. However, the apparent half-life increased more than 10-fold in the P. syringae lon::Tz mutants or upon transfer to an inductive medium. The P. syringae lon mutants elicited rapidly developing plant responses and were shown to hypersecrete effector proteins, such as AvrPto. These results indicate that expression of the hrp regulon and type III secretion are negatively regulated by Lon-mediated degradation of HrpR. PMID- 12123453 TI - Uptake of transforming DNA in Gram-positive bacteria: a view from Streptococcus pneumoniae. AB - In a working model for the uptake of transforming DNA based on evidence taken from both Bacillus subtilis and Streptococcus pneumoniae, the ComG proteins are proposed to form a structure that provides access for DNA to the ComEA receptor through the peptidoglycan. DNA would then be delivered to the ComEC-ComFA transport complex. A DNA strand would be degraded by a nuclease, while its complement is pulled into the cell by ComFA through an aqueous pore formed by ComEC. The nuclease is known in S. pneumoniae only as EndA. We have examined the processing (i.e. binding, degradation and internalization) of DNA in S. pneumoniae strains lacking candidate uptake proteins. Mutants were generated by transposon insertion in endA, comEA/C, comFA/C, comGA and dprA. Processing of DNA was abolished only in a comGA mutant. As significant binding was measured in comEA mutants, we suggest the existence of two stages in binding: surface attachment (abolished in a comGA mutant) required for and preceding deep binding (by ComEA). Abolition of degradation in comGA and comEA mutants indicated that, despite its membrane location, EndA cannot access donor DNA by itself. We propose that ComEA is required to deliver DNA to EndA. DNA was still bound and degraded in comEC and comFA mutants. We conclude that recruitment of EndA can occur in the absence of ComEC or ComFA and that EndA is active even when the single strands it produces are not pulled into the cell. Finally, inactivation of dprA had no effect on the internalization of DNA, indicating that DprA is required at a later stage in transformation. PMID- 12123454 TI - Novel mode of transcription regulation of divergently overlapping promoters by PhoP, the regulator of two-component system sensing external magnesium availability. AB - PhoP is a response regulator of the PhoQ-PhoP two-component system controlling a set of the Mg(II)-response genes in Escherichia coli. Here we demonstrate the mode of transcription regulation by phosphorylated PhoP of divergently transcribed mgtA and treR genes, each encoding a putative Mg(II) transporter and a repressor for the trehalose utilization operon respectively. Under Mg(II) limiting conditions in vivo, two promoters, the upstream constitutive P2 and the downstream inducible P1, were detected for the mgtA gene. Gel-shift analysis in vitro using purified PhoP indicates its binding to a single DNA target, centred between -43 and -24 of the mgtAP1 promoter. This region includes the PhoP box, which consists of a direct repeat of the heptanucleotide sequence (T)G(T)TT(AA). Site-directed mutagenesis studies indicate the critical roles for T (position 3), T (position 4) and A (position 6) for PhoP-dependent transcription from mgtAP1. DNase I footprinting assays reveal weak binding of PhoP to this PhoP box, but the binding becomes stronger in the simultaneous presence of RNA polymerase. Likewise the RNA polymerase binding to the P1 promoter becomes stronger in the presence of PhoP. For the PhoP-assisted formation of open complex at the mgtAP1 promoter, however, the carboxy-terminal domain of alpha subunit (alpha CTD) is not needed. For transcription in vivo of the treR gene, four promoters were identified. The most upstream promoter treRP4 divergently overlaps with the mgtAP1 promoter, sharing the same sequence as the respective -10 signal in the opposite direction. In vitro transcription using mutant promoters support this prediction. In the presence of PhoP, transcription from the promoter treRP3 was repressed with concomitant activation of mgtAP1 transcription. The PhoP box is located between 46 and -30 with respect to treRP3, and the alpha CTD is needed for this repression. PMID- 12123455 TI - Identification of amino acids in the Dr adhesin required for binding to decay accelerating factor. AB - Members of the Dr family of adhesins of Escherichia coli recognize as a receptor the Dr(a) blood-group antigen present on the complement regulatory and signalling molecule, decay-accelerating factor (DAF). One member of this family, the Dr haemagglutinin, also binds to a second receptor, type IV collagen. Structure/function information regarding these adhesins has been limited and domains directly involved in the interaction with DAF have not been determined. We devised a strategy to identify amino acids in the Dr haemagglutinin that are specifically involved in the interaction with DAF. The gene encoding the adhesive subunit, draE, was subjected to random mutagenesis and used to complement a strain defective for its expression. The resulting mutants were enriched and screened to obtain those that do not bind to DAF, but retain binding to type IV collagen. Individual amino acid changes at positions 10, 63, 65, 75, 77, 79 and 131 of the mature DraE sequence significantly reduced the ability of the DraE adhesin to bind DAF, but not collagen. Over half of the mutants obtained had substitutions within amino acids 63-81. Analysis of predicted structures of DraE suggest that these proximal residues may cluster to form a binding domain for DAF. PMID- 12123456 TI - Depression of Saccharomyces cerevisiae invasive growth on non-glucose carbon sources requires the Snf1 kinase. AB - Haploid Saccharomyces cerevisiae cells growing on media lacking glucose but containing high concentrations of carbon sources such as fructose, galactose, raffinose, and ethanol exhibit enhanced agar invasion. These carbon sources also promote diploid filamentous growth in response to nitrogen starvation. The enhanced invasive and filamentous growth phenotypes are suppressed by the addition of glucose to the media and require the Snf1 kinase. Mutations in the PGI1 and GND1 genes encoding carbon source utilization enzymes confer enhanced invasive growth that is unaffected by glucose but requires active Snf1. Carbon source does not modulate FLO11 flocculin expression, but enhanced polarized bud site selection is necessary for invasion on certain carbon sources. Interestingly, deletion of SNF1 blocks invasion without affecting bud site selection. Snf1 is also required for formation of spokes and hubs in multicellular mats. To examine glucose repression of invasive growth more broadly, we performed genome-wide microarray expression analysis in wild-type cells growing on glucose and galactose, and snf1 Delta cells on galactose. SNF1 probably mediates glucose repression of multiple genes potentially involved in invasive and filamentous growth. FLO11-independent cell-cell attachment, cell wall integrity, and/or polarized growth are affected by carbon source metabolism. In addition, derepression of cell cycle genes and signalling via the cAMP-PKA pathway appears to depend upon SNF1 activity during growth on galactose. PMID- 12123458 TI - Genes of non-typeable Haemophilus influenzae expressed during interaction with human epithelial cell lines. AB - Non-typeable Haemophilus influenzae may infect the lower respiratory airways of chronic obstructive pulmonary disease patients. We characterized genes of non typeable H. influenzae expressed during interaction with two human respiratory tract-derived epithelial cell lines. A library of 8000 clones was constructed in H. influenzae Rd (rec1) by cloning chromosomal fragments upstream of a promoterless cat gene. Exposure of this library to NCI-H292 epithelial cell layers in the presence of chloramphenicol (Cam) resulted in survival of bacteria expressing cat. A total of 52 clones were selected that were resistant to Cam in the presence of epithelial cells of cell line NCI-H292. These did not (n = 42) or hardly grow (n = 10) on sBHI plates containing Cam and were sensitive to Cam in cell culture medium alone. All clones, moreover, survived Cam in the presence of Hep2 epithelial cell layers. Sequence analysis showed that four clones contained sequences without homology to Rd or any other sequence, and therefore contained promoters and parts of open reading frames (ORFs) of novel genes. The other 48 clones were homologous to Rd, and characterization was based upon this genome. Six different functional classes were distinguished: (i) metabolic processes; (ii) stress response; (iii) gene expression; (iv) cell envelope biosynthesis; (v) DNA-related processes and cell division; and (vi) ORFs encoding proteins of unknown function. The contribution of identified genes to non-typeable H. influenzae adaptation to the epithelial cell environment is discussed. PMID- 12123459 TI - Rapid induction and reversal of a bacteriostatic condition by controlled expression of toxins and antitoxins. AB - RelE and ChpAK (MazF) toxins of Escherichia coli have previously been described as proteins that mediate efficient cell killing. We show here that induction of relE or chpAK transcription does not confer cell killing but, instead, induces a static condition in which the cells are still viable but unable to proliferate. Later induction of transcription of the antitoxin genes relB or chpAI fully reversed the static condition induced by RelE and ChpAK respectively. We also provide a mechanistic explanation for these findings. Thus, induction of relE transcription severely inhibited translation, whereas induction of chpAK transcription inhibited both translation and replication. Hence, most likely, lack of colony formation is due to inhibition of translation in the case of relE and inhibition of translation and/or replication in the case of chpAK. Consistent with this proposal, later induction of transcription of the cognate antitoxin genes simultaneously reversed cell stasis and the inhibitory effects of RelE and ChpAK on macromolecular syntheses. These results preclude that RelE and ChpAK mediate cell killing during the conditions used here. In vivo and in vitro analyses of a mutant RelE protein supported that inhibition of colony formation was due to inhibition of translation. PMID- 12123457 TI - Vibrio cholerae CytR is a repressor of biofilm development. AB - Vibrio cholerae is both a human pathogen and a natural inhabitant of aquatic environments. In the aquatic environment, microorganisms are found attached to surfaces in structures known as biofilms. We have identified a transcriptional repressor in V. cholerae that inhibits exopolysaccharide synthesis and biofilm development. Our studies show that this repressor is the V. cholerae homologue of Escherichia coli CytR, a protein that represses nucleoside uptake and catabolism when nucleosides are scarce. We propose that the role of CytR in V. cholerae biofilm development is to co-ordinate bacterial biofilm accumulation with the presence of nucleosides. Thus, nucleosides may be a signal to planktonic cells to join the biofilm. PMID- 12123460 TI - A bZIP protein from halophilic archaea: structural features and dimer formation of cGvpE from Halobacterium salinarum. AB - The cGvpE protein of Halobacterium salinarum PHH4 has been identified as transcriptional activator for the promoter of the c-gvpA gene encoding the major gas vesicle structural protein cGvpA. Molecular modelling of the carboxy-terminal region of cGvpE suggests that this protein resembles a basic leucine-zipper protein, and mutations in the putative DNA binding domain DNAB completely abolish the activator function in Haloferax volcanii transformants. Mutations in the key residues of the putative leucine-zipper region AH6 of cGvpE confirmed that the three residues V159, L166 and L173 were essential for the activator function of cGvpE at the c-gvpA promoter, whereas the cysteine residue C180 could be altered to a leucine or an aspartate residue without the loss of this function. Mutations in basic residues of helix AH4 demonstrated the importance of the lysine K104 for the activator function of cGvpE. A cGvpE protein containing a his-tag at the C terminus was still able to activate the expression of c-gvpA in vivo. The cGvpE his-purified from Hf. volcanii formed a dimer in Blue-native polyacrylamide gels that could be resolved into monomers by SDS-polyacrylamide gel electrophoresis (SDS-PAGE). Dimers of cGvpE were already seen using SDS-PAGE, but not with cGvpE mutant proteins containing the alterations L166E or L173E/C180L in the leucine zipper. These results imply that the hydrophobic surface of helix AH6 is indeed required for the establishment of cGvpE dimers. PMID- 12123463 TI - Control of the glycolytic gapA operon by the catabolite control protein A in Bacillus subtilis: a novel mechanism of CcpA-mediated regulation. AB - Glycolysis is one of the main pathways of carbon catabolism in Bacillus subtilis. Expression of the gapA gene encoding glyceraldehyde-3-phosphate dehydrogenase, the key enzyme of glycolysis from an energetic point of view, is induced by glucose and other sugars. Two regulators are involved in induction of the gapA operon, the product of the first gene of the operon, the CggR repressor, and catabolite control protein A (CcpA). CcpA is required for induction of the gapA operon by glucose. Genetic evidence has demonstrated that CcpA does not control the expression of the gapA operon by binding directly to a target in the promoter region. Here, we demonstrate by physiological analysis of the inducer spectrum that CcpA is required only for induction by sugars transported by the phosphotransferase system (PTS). A functional CcpA is needed for efficient transport of these sugars. This interference of CcpA with PTS sugar transport results from an altered phosphorylation pattern of HPr, a phosphotransferase of the PTS. In a ccpA mutant strain, HPr is nearly completely phosphorylated on a regulatory site, Ser-46, and is trapped in this state, resulting in its inactivity in PTS phosphotransfer. A mutation in HPr affecting the regulatory phosphorylation site suppresses both the defect in PTS sugar transport and the induction of the gapA operon. We conclude that a low-molecular effector derived from glucose that acts as an inducer for the repressor CggR is limiting in the ccpA mutant. PMID- 12123461 TI - LrhA as a new transcriptional key regulator of flagella, motility and chemotaxis genes in Escherichia coli. AB - The function of the LysR-type regulator LrhA of Escherichia coli was defined by comparing whole-genome mRNA profiles from wild-type E. coli and an isogenic lrhA mutant on a DNA microarray. In the lrhA mutant, a large number (48) of genes involved in flagellation, motility and chemotaxis showed relative mRNA abundances increased by factors between 3 and 80. When a representative set of five flagellar, motility and chemotaxis genes was tested in lacZ reporter gene fusions, similar factors for derepression were found in the lrhA mutant. In gel retardation experiments, the LrhA protein bound specifically to flhD and lrhA promoter DNA (apparent K(D) approximately 20 nM), whereas the promoters of fliC, fliA and trg were not bound by LrhA. The expression of flhDC (encoding FlhD(2)C(2)) was derepressed by a factor of 3.5 in the lrhA mutant. FlhD(2)C(2) is known as the master regulator for the expression of flagellar and chemotaxis genes. By DNase I footprinting, LrhA binding sites at the flhDC and lrhA promoters were identified. The lrhA gene was under positive autoregulation by LrhA as shown by gel retardation and lrhA expression studies. It is suggested that LrhA is a key regulator controlling the transcription of flagellar, motility and chemotaxis genes by regulating the synthesis and concentration of FlhD(2)C(2). PMID- 12123462 TI - The plastid DNA of the malaria parasite Plasmodium falciparum is replicated by two mechanisms. AB - In common with other apicomplexan parasites, Plasmodium falciparum, a causative organism of human malaria, harbours a residual plastid derived from an ancient secondary endosymbiotic acquisition of an alga. The function of the 35 kb plastid genome is unknown, but its evolutionary origin and genetic content make it a likely target for chemotherapy. Pulsed field gel electrophoresis and ionizing radiation have shown that essentially all the plastid DNA comprises covalently closed circular monomers, together with a tiny minority of linear 35 kb molecules. Using two-dimensional gels and electron microscopy, two replication mechanisms have been revealed. One, sensitive to the topoisomerase inhibitor ciprofloxacin, appears to initiate at twin D-loops located in a large inverted repeat carrying duplicated rRNA and tRNA genes, whereas the second, less drug sensitive, probably involves rolling circles that initiate outside the inverted repeat. PMID- 12123464 TI - The role of heterologous receptors in McpB-mediated signalling in Bacillus subtilis chemotaxis. AB - Asparagine chemotaxis in Bacillus subtilis appears to involve two partially redundant adaptation mechanisms: a receptor methylation-independent process that operates at low attractant concentrations and a receptor methylation-dependent process that is required for optimal responses to high concentrations. In order to elucidate these processes, chemotactic responses were assessed for strains expressing methylation-defective mutations in the asparagine receptor, McpB, in which all 10 putative receptors (10del), five receptors (5del) or only the native copy of mcpB were deleted. This was done in both the presence and the absence of the methylesterase CheB. We found that: (i) only responses to high concentrations of asparagine were impaired; (ii) the presence of all heterologous receptors fully compensated for this defect, whereas responses progressively worsened as more receptors were taken away; (iii) methyl-group turnover occurred on heterologous receptors after the addition of asparagine, and these methylation changes were required for the restoration of normal swimming behaviour; (iv) in the absence of the methyleste-rase, the presence of heterologous receptors in some cases caused impaired chemotaxis; and (v) either a certain threshold number of receptors must be present to promote basal CheA activity, or one or more of the receptors missing in the 10del background (but present in the 5del background) is required for establishing basal CheA activity. Taken together, these findings suggest that many or all chemoreceptors work as an ensemble that constitutes a robust chemotaxis system. We propose that the ability of non-McpB receptors to compensate for the methylation-defective McpB mutations involves lateral transmission of the adapted conformational change across the ensemble. PMID- 12123465 TI - GabR, a member of a novel protein family, regulates the utilization of gamma aminobutyrate in Bacillus subtilis. AB - The Bacillus subtilis ycnG (gabT) and ycnH (gabD) genes were shown to encode gamma-aminobutyrate (GABA) aminotransferase and succinic semi-aldehyde dehydrogenase, respectively, and to form a GABA-inducible operon. Null mutations in gabT, gabD or the divergently transcribed ycnF (gabR) gene blocked the utilization of GABA as sole nitrogen source. GabR proved to be a transcriptional activator of the gabTD operon and a negative autoregulator. The target of GabR action was localized to an 87 bp region that includes both gabR and gabT promoters. GabR is a member of a novel but widespread family of chimeric bacterial proteins that have apparent DNA-binding and aminotransferase domains. Mutations in conserved residues of the putative aminotransferase domain abolished GabR function as a transcriptional activator, but did not affect its activity as a negative autoregulator. PMID- 12123466 TI - Bladder instability: a re-appraisal of classical experimental approaches and development of new therapeutic strategies. AB - 1 Despite the growing social interest in human urinary tract disorders, the aetiology of detrusor instability remains poorly understood. Myogenic and neural impairment of detrusor activity caused by CNS or autonomic injuries can results in dysfunctions of normal voiding of the bladder such as urinary incontinence. 2 The contractility of human detrusor smooth muscle is critically dependent on acetylcholine-induced muscarinic receptor activation. Biochemical and functional in vivo and in vitro studies suggest the presence of an heterogeneous population of muscarinic receptor subtypes (M1-M4) localized at muscular and neutral sites. There is increasing evidence on the prejunctional auto- and hetero-regulation of acetylcholine release from parasympathetic nerve endings in modulating detrusor muscle contraction during micturition. 3 Activation of P2X purinoreceptors closely associated with the parasympathetic varicosities seems to be implicated to varying extent in the contractility in normal or instable human detrusor. Interestingly, P2X(1) subtype expression on smooth muscle increases considerably in the symptomatically obstructed bladder. A striking absence of P2X(3) and P2X(5) subtypes was observed in the cholinergic innervation of detrusor from patients with urgent incontinence. Thus, it is likely that alteration of the neural acetylcholine control can play a critical role in pathological states. 4 If the failures in storage and voiding can be recognized urodynamically, considerable difficulties remain in investigating the underlying functional changes especially because the study of the pathophysiology requires techniques that can be justified in animals but not in humans. 5 Recently, to solve this problem an alternative technique using human smooth muscle cells in culture has been developed. Human cell lines may be relevant in investigating the molecular pathways in physiological and pathological conditions. 6 The potential development of novel molecular therapeutic strategies such as gene therapy and tissue engineering is also discussed. PMID- 12123467 TI - The comparison of vitamin C and vitamin E on the protein oxidation of diabetic rats. AB - 1 We measured the plasma glucose and the glycosylated haemoglobin at the time of sacrifice in streptozotocin-induced diabetic mellitus (DM) rats. 2 In diabetic rats, plasma glucose and glycosylated haemoglobin was increased as compared with normal rats, and vitamin E inhibited the increase of glycosylated haemoglobin level but vitamin C had no effect. 3 The peroxidized proteins and lipids from the diabetic organs such as liver or kidney were measured to assess the oxidative damage. The 2,4-dinitrophenyl-hydrazine (DNPH) incorporation method was used to measure the peroxidized protein. In diabetic rats, DNPH incorporation was increased as compared with normal rats and vitamin E also inhibited the increase of DNPH incorporation but vitamin C had no effect. It suggests that the protein oxidation occurred on the liver in diabetic rats and the oxidative stress is general in the diabetic condition. 4 We measured the systolic arterial pressure and mean arterial pressure in normal rats, nephrectomy (NEPH)-rats, diabetic rats (DM), and NEPH-diabetic rats (NEPH-DM). Blood pressure was significantly increased in DM and NEPH-DM as compared with normal rats. 5 In conclusion, plasma glucose, glycosylated haemoglobin, and the oxidation of proteins or lipid were increased in diabetic rats. Vitamin E decreased the plasma glucose, glycosylated haemoglobin and the oxidation of proteins and lipid, but vitamin C had no effects. PMID- 12123468 TI - Adrenaline hypothesis: effect of formoterol on noradrenaline release. AB - 1 Originally, the so-called 'adrenaline hypothesis' related the release of noradrenaline (NA) to stimulation of presynaptic beta2-receptors in nerve endings; now it confers a possible role to adrenaline taken up then released by nerves endings. It represents a potentially useful therapeutic pathway. The present study aims to investigate the effects of formoterol, a highly selective beta2-adrenoceptor agonist. 2 It was carried out in freely moving rats, the isotope dilution technique being used to measure the NA spill over rate (NA-SOR) and metabolic clearance rate (MCR). 3 A series of three results are reported. (a) When compared with adrenaline on equimolar basis, formoterol (2.3 micro kg-1 min 1) increased NA-SOR while mean arterial blood pressure was decreased and heart rate increased. Thus, it was difficult to separate a direct presynaptic effect from indirect baroreflex-dependent activation of the sympathetic system. (b) When formoterol was infused at 1 ng kg-1 min-1, a dose empirically defined to induce no haemodynamic effect, NA-SOR was significantly increased, while NA-MCR remained unchanged. (c) The NA-SOR response to formoterol was not amplified by the presynaptic alpha2-adrenoceptor blocker, yohimbine, in contrast to the NA-SOR response to adrenaline. 4 In conclusion, formoterol, a beta2-adrenoceptor agonist, is shown to increase the release and plasma concentration of NA while its clearance was not changed. PMID- 12123469 TI - The minor population of M3-receptors mediate contraction of human detrusor muscle in vitro. AB - 1 The objective was to determine the role of muscarinic receptor subtypes in mediating contraction of the human detrusor smooth muscle in vitro. 2 Contractile responses of human detrusor muscle strips to carbachol were obtained in the absence and presence of a range of muscarinic antagonists (pirenzepine, methoctramine, 4-diphenylacetoxy-N-methyl piperidine methiodide (4-DAMP), tropicamide, oxybutynin and tolterodine). Affinity estimates (pKB values) were calculated for the antagonists and correlated with values at the cloned muscarinic receptor subtypes quoted in the literature. 3 Pirenzepine, methoctramine and tropicamide drugs that have high affinities at M1, M2 and M4 receptors, respectively, all had low affinities on the human detrusor (pKB values of 6.8, 6.9 and 6.5, respectively), whilst the M3-selective antagonist 4-DAMP had a high affinity (9.5). Schild plots for all four antagonists had slopes of unity indicating an action at a single receptor. Oxybutynin and tolterodine also acted as competitive antagonists with affinity estimates of 7.6 and 8.1, respectively. 4 When the antagonist affinities obtained on the bladder were plotted against the values published for these antagonists at the cloned muscarinic receptor subtypes, the best correlations were obtained for the m3- and m5-muscarinic receptor subtypes. 5 These data suggest that direct contractile responses of the human detrusor muscle to muscarinic receptor stimulation in vitro are mediated solely via the M3-muscarinic receptor subtype with no contribution from the major M2-receptor population. PMID- 12123470 TI - Modification of cardiovascular responses to adenosine A1 receptor stimulation in the posterior hypothalamus of anaesthetized rats by cAMP and by GABA(B) receptor blockade. AB - 1 Injection of N(6)-cyclohexyladenosine (CHA; 1, 5 and 10 nmol), an adenosine A1 receptor agonist, into the posterior hypothalamus of rats produced a dose dependent decrease in blood pressure (BP) and heart rate (HR). 2 Pretreatment with 8-cyclopentyl-1,3-dimethylxanthine (CPDX; 50 nmol), an adenosine A1 receptor antagonist, blocked the depressor and bradycardic effects of CHA (10 nmol). 3 Pretreatment with 8-bromo-cyclic adenosine monophosphate (AMP) (10 nmol), a cAMP analogue, attenuated the depressor and bradycardic effect of CHA (10 nmol); 8 bromo-cyclic guanosine monophosphate (GMP) (10 nmol), a cGMP analogue, did not modify those effects of CHA. 4 In addition, pretreatment with 5-aminovaleric acid (25 nmol), a gamma-aminobutyric acid (GABA)(B) receptor antagonist, attenuated the depressor and bradycardic effects of CHA (10 nmol). 5 These results suggest that adenosine A1 receptors in the posterior hypothalamus have an inhibitory role in the central cardiovascular regulation and that these vasodepressive and bradycardic actions are modified by raised cAMP and by GABA(B) receptor inhibition. PMID- 12123471 TI - Effects of restraint stress on responsiveness of atria and vas deferens in Sprague-Dawley rats. AB - 1 The effects of stress in rats were evaluated by measuring changes in body weight and in responsiveness to noradrenaline (NA) in the isolated vas deferens and atria after the animals had been exposed to restraint or restraint and isolation. 2 Animals which were subjected to restraint alone (1 h day-1 for 3, 7 or 28 days) had a significantly reduced rate of body weight gain. This effect was not potentiated by the additional stress of isolation. 3 Restraint alone did not produce significant changes in the responsiveness of the vas deferens to NA. However, adding isolation to restraint, as an additional stress, produced a further leftward shift of the NA dose-response curve for the vas deferens, so that there was a significant increase in sensitivity compared with control. 4 There was a significant rightward shift in the NA dose-response curves or reduction in sensitivity in the atria from animals which had been restrained for 7 or 28 days. Isolation did not produce a further rightward shift in the NA dose response curve. 5 The results from this study indicate that the stress associated with repeated restraint reduces the rate of weight gain and reduces the responsiveness of the atria to NA. The responsiveness of the vas deferens to NA was increased by stress, but the combined effect of isolation and restraint was required to produce a significant effect. The differences in the effects of stress on these tissues could be associated with differences in presynaptic receptor populations. PMID- 12123472 TI - Role of endothelium in ischaemia-induced myocardial dysfunction of isolated working hearts: cardioprotection by activation of adenosine A(2A) receptors. AB - 1 This study aimed to determine the role of the vascular endothelium on recovery of contractile function following global low-flow ischaemia of guinea-pig isolated working hearts and the effects of adenosine analogues on this recovery. 2 Guinea-pig isolated spontaneously beating or paced working hearts were set up and coronary flow (CF), aortic output (AO) (as an index of cardiac function), heart rate (HR), left ventricular pressure (LVP) and dP/dt max recorded. The endothelium was either intact or removed by a blast of oxygen. 3 In spontaneously beating hearts, low-flow ischaemia for 30 min reduced CF and cardiac contractility (LVP, dP/dt max) but not AO. On reperfusion, CF, LVP and dP/dt max recovered, while AO fell precipitously followed by a gradual recovery, indicative of myocardial stunning. The effects of ischaemia did not differ between endothelium-intact and -denuded hearts, indicating no role of the endothelium in the changes observed. 4 The adenosine analogues, N6-cyclopentyladenosine (CPA, A1 selective), 5'-N-ethylcarboxamidoadenosine (NECA, two-fold A2 selective over A1) and 2-p-((carboxyethyl)-phenethylamino)-5'carboxamidoadenosine (CGS21680, A2A selective) were infused (3 x 10-7 M) from 10 min into the 30-min low-flow ischaemia of denuded hearts and during reperfusion. 5 CGS21680 increased CF and improved the postischaemic functional recovery, as measured by the AO. NECA and CPA were not cardioprotective. The A2A selective antagonist, ZM241385, attenuated the coronary vasodilatation by CGS21680 and abolished the improved recovery of AO on reperfusion. 6 Reperfusion of paced working hearts caused a dramatic fall in AO which failed to recover. Infusion of CGS21680 from 15 min into the ischaemic period produced vasodilatation but failed to restore AO, presumably because the ischaemic damage was irreversible. 7 Thus, the endothelium plays no role in myocardial dysfunction following low-flow global ischaemia and reperfusion of guinea-pig working hearts. The A2A adenosine receptor-selective agonist but not the non-selective A2 receptor agonist, NECA, attenuated ischaemia- and reperfusion-induced stunning. This was attributed to increased CF and was independent of the endothelium. PMID- 12123474 TI - In situ studies of the phylogeny and physiology of filamentous bacteria with attached growth. AB - Among the filamentous bacteria occasionally causing bulking problems in activated sludge treatment plants, three morphotypes with attached microbial growth are common, Eikelboom Type 0041, Type 1851 and Type 1701. A better knowledge of the phylogeny and physiology of these filamentous bacteria is necessary in order to develop control strategies for bulking. In this study we have used a combination of fluorescence in situ hybridization (FISH) and microautoradiography (MAR) to investigate the identity and in situ physiology of the Type 0041-morphotype and its attached bacteria in two wastewater treatment plants. Identification and enumeration of Type 0041 using group-specific 16S rRNA-targeted FISH probes revealed that approximately 15% of the filaments hybridized with a gene probe specific for the TM7 group, a recently recognized major lineage in the bacterial domain. All other filaments morphologically identified as Type 0041 only hybridized to the general bacterial EUB338-probe, indicating that they probably do not belong to commonly isolated bacterial phyla such as the Proteobacteria, Firmicutes, Actinobacteria and Bacteroidetes, for which group-specific probes were used. The phylogenetic heterogeneity of Type 0041 again highlights the inadequacy of a morphology-based classification system. Like the filaments, most of the attached microbial cells were not identified beyond their affiliation to the Bacteria using the group-specific FISH probes. However, several different bacterial phyla were represented in the identified fraction suggesting that the attached microorganisms are phylogenetically diverse. The study of the in situ physiology of Type 0041 using MAR-FISH revealed that both the filaments and the attached bacteria on Type 0041 were versatile in the use of organic substrates and electron acceptors. It was observed that all Type 0041 could consume glucose, but none of the filaments were able to consume acetate under any conditions tested, in contrast to some of the attached bacteria. No significant physiological differences were found between TM7-positive and TM7-negative Type 0041 filaments, and only minor differences were observed between the two treatment plants tested. These are the first data on the physiology of the almost entirely uncharacterized TM7 phylum and show that TM7 filamentous bacteria can uptake carbon substrates under aerobic and anaerobic conditions. PMID- 12123473 TI - Picobenthic cyanobacterial populations revealed by 16S rRNA-targeted in situ hybridization. AB - We report on the morphological identification of a population of benthic cyanobacteria from microbial mats, known previously only from molecular analyses of field samples, based on the retrieval of environmental 16S rRNA sequences. We used in situ hybridization with horseradish peroxidase-labelled oligonucleotide probes designed to target the 16S rRNA of our unidentified population. Two probes were designed and checked for target binding ability and specificity using membrane hybridization against electroblotted bands from a denaturant gradient gel electrophoresis (DGGE) fingerprint of 16S rDNA gene fragments from the original cyanobacterial community. Under in situ hybridization, these probes bound specifically to extremely small, unicellular, colony-forming cyanobacteria, 0.75-1 microm in diameter, which were embedded in abundant mucilaginous investments. We propose the term picobenthos, by analogy with picoplankton, to describe those unicellular benthic microbes around or less than 1 microm in diameter. Although picoplanktonic cyanobacteria are abundant in ocean and freshwaters, picobenthic (<1 microm) unicellular cyanobacteria are not typically recognized as a major component of microbial mats. The small size and low levels of photopigment autofluorescence from these cells probably rendered them cryptic or indistinguishable from heterotrophic bacteria in routine microscopic observations. It is not known how widespread picobenthic cyanobacteria may be in other environments. PMID- 12123475 TI - Phylogenetic 16S rRNA analysis reveals the presence of complex and partly unknown bacterial communities in Tito Bustillo cave, Spain, and on its Palaeolithic paintings. AB - Tito Bustillo cave (Ribadesella, Spain) contains valuable Palaeolithic paintings, which date back 15 000-20 000 years. Since 1969, the cave has been open to the public. Rock wall surfaces, spelaeothems and soils are covered by apparent biofilms of phototrophic microorganisms, which develop under artificial lighting. In addition, rock surfaces present conspicuous bacterial growth in the form of round colonies of different colours and about 1-2 mm in diameter. Even the famous Paintings Panel shows some evident microbial growth. In the present study, bacterial communities on the paintings and on the rock surfaces near the paintings were analysed by culture-independent techniques, including polymerase chain reaction (PCR) amplification of bacterial 16S rRNA genes (16S rDNA), phylogenetic sequence analyses and genetic community fingerprinting by denaturing gradient gel electrophoresis (DGGE). DGGE fingerprints showed complex bacterial community patterns. Forty-one clones matching DGGE bands of the community fingerprints were sequenced, representing about 39% of DNA fragments in the DGGE patterns. Phylogenetic sequence analyses revealed a high number of phylogenetically novel 16S rDNA sequence types and a high diversity of putatively chemotrophic and heterotrophic bacteria. Sequences were phylogenetically most closely related to the Proteobacteria (20 clones), green non-sulphur bacteria (three clones), Planctomycetales order (one clone), Cytophaga-Flexibacter- Bacteroides division (one clone) and the Actinobacteria (four clones). Furthermore, we report the presence of members of the Acidobacterium division (12 clones) in a karstic hypogean environment. Members of this phylum have not so far been detected in these particular environments. PMID- 12123476 TI - Characterization of the bacterial consortium associated with black band disease in coral using molecular microbiological techniques. AB - The bacterial community associated with black band disease (BBD) of the scleractinian corals Diploria strigosa, Montastrea annularis and Colpophyllia natans was examined using culture-independent techniques. Two complementary molecular screening techniques of 16S rDNA genes [amplified 16S ribosomal DNA restriction analysis (ARDRA) of clone libraries and denaturing gradient gel electrophoresis (DGGE)] were used to give a comprehensive characterization of the community. Findings support previous studies indicating low bacterial abundance and diversity associated with healthy corals. A single cyanobacterial ribotype was present in all the diseased samples, but this was not the same as that identified from Phormidium corallyticum culture isolated from BBD. The study confirms the presence of Desulfovibrio spp. and sulphate-reducing bacteria that have previously been associated with the BBD consortium. However, the species varied between diseased coral samples. We found no evidence of bacteria from terrestrial, freshwater or human sources in any of the samples. We report the presence of previously unrecognized potential pathogens [a Cytophaga sp. and an alpha-proteobacterium identified as the aetiological agent of juvenile oyster disease (JOD)] that were consistently present in all the diseased coral samples. The molecular biological approach described here gives an increasingly comprehensive and more precise picture of the bacterial population associated with BBD. To understand the pathogenesis of BBD, our attention should be focused on the pervasive ribotypes identified in this study (the Cyanobacterium sp., the Cytophaga sp. and the JOD pathogen). PMID- 12123477 TI - Microbial reduction of Fe(III) in the presence of oxygen under low pH conditions. AB - In acidic, coal mining lake sediments, facultatively anaerobic Acidiphilium species are probably involved in the reduction of Fe(III). Previous results indicate that these bacteria can co-respire O2 and Fe(III). In this study, we investigated the capacity of the sediment microbiota to reduce Fe(III) in the presence of O2 at pH 3. In sediment microcosms with 4% O2 in the headspace, the concentration of Fe(II) increased at a rate of 1.03 micromol (g wet sediment)-1 day-1 within the first 7 days of incubation which was similar to the rate obtained with controls incubated under anoxic conditions. However, in microcosms incubated under air, Fe(II) was consumed after a lag phase of 8 h with a rate of 2.66 micromol (g wet sediment)-1 day-1. Acidiphilium cryptum JF-5, isolated from this sediment, reduced soluble Fe(III) with either 4 or 21% O2 in the headspace, and concomitantly consumed O2. However, the rate of Fe(II) formation normalized for cell density decreased under oxic conditions. Schwertmannite, the predominant Fe(III)-mineral of this sediment, was also reduced by A. cryptum JF-5 under oxic conditions. The rate of Fe(II) formation by A. cryptum JF-5 decreased after transfer from preincubation under air in medium lacking Fe(III). Acidiphilium cryptum JF-5 did not form Fe(II) when preincubated under air and transferred to anoxic medium containing Fe(III) and chloramphenicol, an inhibitor of protein synthesis. These results indicate that: (i) the reduction of Fe(III) can occur at low O2 concentrations in acidic sediments; (ii) Fe(II) can be oxidized at O2 concentrations near saturation; and (iii) the enzyme(s) responsible for the reduction of Fe(III) in A. cryptum JF-5 are not constitutive. PMID- 12123478 TI - Development of a novel, bioluminescence-based, fungal bioassay for toxicity testing. AB - Naturally bioluminescent fungi, Armillaria mellea and Mycena citricolor, were used to develop a novel, bioluminescence-based bioassay for toxicity testing. Bioassays were carried out to assess the toxicity of 3,5-dichlorophenol (3,5 DCP), pentachlorophenol (PCP), copper and zinc. The results suggested that 60 min was a suitable exposure time for the bioassay. Light reduction was observed in response to 3,5-DCP, PCP and Cu for both A. mellea and M. citricolor, but to Zn only for A. mellea. Armillaria mellea was significantly less sensitive to 3,5-DCP and PCP than M. citricolor. The EC50 values for A. mellea and M. citricolor were similar to EC50 values for 3,5-DCP, PCP and Cu (but not Zn) of bioluminescence based bacterial biosensors. They were also similar to EC50 values for Cu and Zn of a bioluminescence-based yeast biosensor. The results highlighted the importance of using both prokaryotic and eukaryotic biosensors. The novel bioassay provides a rapid and sensitive method to assess bioavailability of pollutants as well as a method to determine their toxicity to filamentous fungi. It also expands the range of organisms that can be used for bioluminescence-based toxicity testing by complementing existing biosensors. PMID- 12123480 TI - Non-syndromic autosomal-dominant deafness. AB - Non-syndromic deafness is a paradigm of genetic heterogeneity. More than 70 loci have been mapped, and 25 of the nuclear genes responsible for non-syndromic deafness have been identified. Autosomal-dominant genes are responsible for about 20% of the cases of hereditary non-syndromic deafness, with 16 different genes identified to date. In the present article we review these 16 genes, their function and their contribution to deafness in different populations. The complexity is underlined by the fact that several of the genes are involved in both dominant and recessive non-syndromic deafness or in both non-syndromic and syndromic deafness. Mutations in eight of the genes have so far been detected in only single dominant deafness families, and their contribution to deafness on a population base might therefore be limited, or is currently unknown. Identification of all genes involved in hereditary hearing loss will help in the understanding of the basic mechanisms underlying normal hearing, will facilitate early diagnosis and intervention and might offer opportunities for rational therapy. PMID- 12123481 TI - Development and diseases of the pancreas. AB - The pancreas is a vital gland of exocrine and endocrine function. It is the target of two main affections: diabetes and pancreatic cancer. We describe the tissue interactions, signaling pathways and intracellular targets that are involved in the emergence of the pancreas primordium and its proliferation, morphogenesis and differentiation. It appears that several genes of developmental relevance have an adult function and are involved in pancreas affections. Embryological experimentation in animals contributed to provide candidate genes for human disease and holds promise for future treatments. PMID- 12123485 TI - Service provision of complex mutation analysis: a technical and economic appraisal using dystrophin point mutation analysis as an example. AB - Duchenne muscular dystrophy (DMD) results from mutations in the dystrophin gene. One-third of cases arise from point mutations, which are heterogeneous and difficult to detect. The aims of this study of dystrophin point mutation analysis were to assess its technical feasibility in a routine diagnostic laboratory, and to estimate its costs and clinical benefits. The methods used were a laboratory based study using reverse transcription-polymerase chain reaction (RT-PCR) and a protein truncation test, and a mathematical model to estimate costs and clinical benefits. None of the cases analyzed had an identifiable dystrophin deletion or duplication. They were 12 males affected with DMD and two obligate female carriers; two female carriers of known dystrophin point mutations were also analyzed. Point mutations were detected in six out of 12 males, but in none of the female carriers. Assuming a sensitivity of 50% the model predicts significant clinical benefits of point mutation analysis over linkage analysis, including a reduction in the number of prenatal diagnoses (by 0.77 per family), terminations of pregnancy (by 0.18 per family), and terminations of unaffected fetuses (by 0.16 per family). The mean cost of point mutation analysis to prevent the termination of an unaffected fetus is 6220 US dollars. PMID- 12123486 TI - Apolipoprotein E gene polymorphism in cerebrovascular disease: a case-control study. AB - The association between apolipoprotein E (apo E) polymorphism and stroke has been controversial. So far there are no studies reported on the polymorphism of apolipoprotein E in cerebrovascular diseases in the Asian Indians. A blinded case control study was therefore undertaken and the apo E genotypes and lipid profile of a total of 120 subjects (63 stroke patients and 57 healthy controls) were done. The frequency distribution of apo E alleles and genotypes were assessed and their relation with the occurrence of stroke in Asian Indian subjects was determined. A significantly high frequency of apo epsilon4 allele (30%) was observed in the stroke patients than the controls (11%) (p < 0.005), and patients with epsilon4 allele had a fourfold higher odds to develop stroke OR (95%CI) 4.2 (1.8-10.1) (p < 0.005). On multivariate analysis, after adjusting for age, triglycerides and hypertension, the association of epsilon4 allele with stroke was found to be no longer statistically significant, OR (95%CI) 1.2 (0.4-4.5) (p = NS). On multiple logistic regression analysis age, OR (95%CI) 1.1 (1.1-1.2) (p < 0.001), and hypertension OR (95%CI) 15.1 (2.6-89.1) (p < 0.005) were found to be independent risk factors for development of stroke. This is the first report to have examined the association of apo E gene polymorphism with stroke in the Asian Indians. This study suggests that apo epsilon4 allele, triglycerides, age and hypertension are the predictors for stroke development. PMID- 12123487 TI - Influence of the angiotensin-converting enzyme gene insertion/deletion polymorphism on lipoprotein/lipid response to gemfibrozil. AB - Evidence suggests that fibrate therapy reduces the risk of recurrent coronary heart disease among men with low levels of high density lipoprotein cholesterol (HDL-C). Indirect observations and new possible biological pathways suggest that the angiotensin-converting enzyme (ACE) insertion/deletion (I/D) polymorphism might modulate the lipoprotein/lipid profile and its response to fibrate therapy. To assess the possible interaction between fibrate therapy and such variants on plasma lipid and lipoprotein levels, 65 dyslipidemic abdominally obese men were treated for 6 months with or without gemfibrozil (600 mg twice daily). No differences in baseline plasma lipid and lipoprotein levels were found between genotype groups except for the HDL(3)-C subfraction, which was higher in the DD group (p = 0.02). A two-way factorial ANOVA was used to evaluate the effect of the genotype (DD homozygotes vs I allele carriers), the treatment (placebo vs gemfibrozil), and the interaction between these two independent variables on changes observed in lipid and lipoprotein concentrations. A significant genotype by-treatment interaction (p = 0.02) was found for the plasma HDL-C response to the intervention program. In fact, having the DD genotype and being treated with gemfibrozil had a synergical effect on HDL-C levels. The results of this study suggest that the ACE I/D polymorphism influences the effect of gemfibrozil on plasma HDL-C levels. PMID- 12123488 TI - The relationship between plasma t-PA and PAI-1 levels is dependent on epistatic effects of the ACE I/D and PAI-1 4G/5G polymorphisms. AB - Thrombus formation and degradation is partly due to a complex interplay between tissue-type plasminogen activator (t-PA) and plasminogen activator inhibitor 1 (PAI-1). There is accumulating evidence that plasma levels of t-PA and PAI-1 may be influenced by an interaction between the fibrinolytic and renin-angiotensin systems. The goal of this study was to conduct an exploratory data analysis to determine whether there is evidence that the relationship (i.e. correlation) between plasma t-PA and PAI-1 is influenced by interactive effects of the angiotensin converting enzyme (ACE) insertion/deletion (I/D) and plasminogen activator inhibitor 1 (PAI-1) 4G/5G polymorphisms in a sample of 50 unrelated African Americans and 117 unrelated Caucasians. In a single-locus analysis, no evidence for heterogeneity of plasma t-PA and PAI-1 correlations among either ACE I/D or PAI-1 4G/5G genotypes was detected. However, using the combinatorial partitioning method for exploratory data analysis, we identified evidence that is suggestive of heterogeneity of plasma t-PA and PAI-1 correlations among multilocus ACE I/D and PAI-1 4G/5G genotypes in African American females, Caucasian females, Caucasian males, but not African American males. From these results, we propose as a working hypothesis that the correlation between plasma t PA and PAI-1 may be dependent on epistatic effects of the ACE I/D and PAI-1 4G/5G polymorphisms. This study supports the idea that interactions between the fibrinolytic and renin-angiotensin systems play an important role in the genetic architecture of plasma t-PA and PAI-1. PMID- 12123489 TI - Quantification of CFTR splice variants in adults with disseminated bronchiectasis, using the TaqMan fluorogenic detection system. AB - Mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene are associated with different related disorders such as congenital bilateral absence of the vas deferens, chronic idiopathic pancreatitis, or disseminated bronchiectasis (DB). Many different disease-causing mutations are associated with DB, particularly the 5T allele (IVS 8 polyT tract), a variant of the splice acceptor site at the end of intron 8 that affects the efficiency with which the site is used. It therefore affects the accuracy of exon 9 splicing and hence expression of the functional CFTR protein. In this study we quantified transcripts from nasal biopsies of patients with DB compared to normal controls. We developed a real-time quantitative reverse transcription polymerase chain reaction assay (using the TaqMan system) to evaluate the relative amounts of accurately spliced transcript, and transcript lacking exon 9. Patients with the 5T allele had increased amounts of aberrant transcript: in genotypes 5T/7T, 7T/7T and 9T/7T, mean fractions of 38.4%, 3.5% and 0.6%, respectively, of transcripts had been spliced incorrectly. There was also some evidence that nasal biopsies can provide similar information on transcripts to bronchial biopsies. This functional test is of interest for monitoring the amount of CFTR transcript in different clinical situations or to monitor the effect of drugs on CFTR transcription. PMID- 12123490 TI - DGAT1 promoter polymorphism associated with alterations in body mass index, high density lipoprotein levels and blood pressure in Turkish women. AB - Triglyceride synthesis is catalyzed by acyl CoA:diacylglycerol acyltransferases (DGAT), microsomal enzymes that use diacylglycerol and fatty acyl CoAs as substrates. Because DGAT1 expression is up-regulated during adipocyte differentiation and DGAT1 deficiency is associated with leanness in mice, we hypothesized that alterations in DGAT1 expression may affect human body weight. We identified five polymorphisms in the human DGAT1 promoter and 5' non-coding sequence in a random Turkish population. Functional analysis of one common variant, C79T, revealed reduced promoter activity for the 79T allele in cultured cell lines. In 476 Turkish women, the 79T allele was associated with lower body mass index (BMI) (p = 0.004), conferring an odds ratio of 2.0 (95% CI = 1.30 3.07, p = 0.0001) for BMI 1 cm in size. CONCLUSION: Shock-wave treatment in periarthritis of the shoulder is a new and very effective method for symptomatic calcareous tendinopathy. Extracorporeal shock-wave treatment has good prospects of success in any type of calcification. As a non-invasive technique with a high success rate, shock-wave treatment is an alternative to surgical operations in patients who remain symptomatic after exhaustive conservative treatment. PMID- 12123514 TI - Current international developments in population screening for colorectal cancer. AB - Pilot programmes for screening for colorectal cancer in average-risk individuals using faecal occult blood testing are planned to commence in Australia in 2002. The National Cancer Control Initiative was interested to compare Australia's progress in this area with that of other countries. Information on programmes or pilot programmes for colorectal screening in average-risk individuals was sought by letter, fax or email from cancer organizations, government departments and professional bodies in 55 countries worldwide. Based on the responses received (32 replies), no country was identified as having an active programme for colorectal screening for the general population based on the characteristics of identification of eligible subjects, active recruitment and integrated screening and follow up, which would allow determination of the participation rates, detection rate and outcomes such as the stage distribution of cancer. In the United Kingdom a pilot programme is under way to test the feasibility and acceptability of screening for colorectal cancer in the general population using faecal occult blood testing. In Finland, health authorities have agreed that the evidence is sufficient to embark on a pilot programme, and Canada, the Netherlands, Denmark and Switzerland are considering pilot programmes. From our enquiries, in countries using a planned approach, only the UK appears to be ahead of Australia. The British pilots have commenced earlier and are larger and have a shorter time frame than the Australian pilots. PMID- 12123517 TI - Synovial ablation in a rabbit rheumatoid arthritis model using photodynamic therapy. AB - BACKGROUND: At present there is no ideal minimally invasive method for ablating inflamed synovium in joints that has been unresponsive to optimal medical management in patients with rheumatoid arthritis. The aim of this study was to determine whether photo-dynamic therapy could be used for this purpose. METHODS: In a rabbit knee model of rheumatoid arthritis the pharmacokinetics of the photosensitizer Haematoporphyrin Derivative (HpD) into periarticular tissues and blood was measured following intravenous injection of HpD. The second phase of the study was to determine the histological effect of HpD activation by 63 nm light delivered via an intra-articular optic fibre using a dye pumped KTP-YAG laser. The light dose was varied from 0-200 joule/cm2. RESULTS: Pharmacokinetic studies determined that inflamed synovium rapidly accumulated HpD, with peak levels being reached 12 h following intravenous injection. The ratio of HpD uptake into inflamed synovium versus peri-articular quadriceps muscle was found to be 22.8. Histological examination of the treated knees indicated that selective destruction of inflamed synovium was achieved at light doses 100 joules/cm2 and above. No significant effect was observed on normal intra articular tissues. CONCLUSION: We have demonstrated that the first generation photosensitizer HpD selectively accumulates within inflamed -synovium. Activation of HpD by intra-articular light administration resulted in selective ablation of the inflamed synovium. These findings indicate that PDT offers potential as a new selective, minimally invasive synovectomy technique. PMID- 12123520 TI - Comparison of three methods in surgical treatment of pilonidal disease. PMID- 12123518 TI - Technical developments and a team approach leads to an improved outcome: lessons learnt implementing laparoscopic splenectomy. AB - BACKGROUND: To document the technical aspects, outcome and lessons learnt during the learning curve phase of implementing laparoscopic splenectomy, by comparing the results before and after the introduction of a standardized technique. METHODS: We present a retrospective and prospective review of laparoscopic splenectomies over a 4-year period. Two chronological periods were studied, before and after the implementation of a standardized technique of a laparoscopic splenectomy involving: (i) hilar dissection with ultrasonic shears; (ii) two experienced laparoscopic surgeons; and (iii) trained dedicated equipment and staff using a checklist approach in the preparation and conduct of the operation. Two groups of patients were studied relating to the periods before and after the implementation of a standardized technique. Statistical methods used were the Wilcoxon's rank sum test and the two-sample test. RESULTS: Thirty-one laparoscopic splenectomies were attempted. The most common indication was for idiopathic thrombocytopenic purpura. When comparing the early phase (n = 15) with the standardized technique phase (n = 16), there was a significant reduction in conversion rates (40% vs 6%), operating times (218 min vs 171 min), complication rates (6 cases including 1 death vs none) and length of stay (11 days vs 4 days). The results were significant for reduction in hospital stay, conversion rates and complications rates. CONCLUSIONS: A reduction in conversion rates, operating time, morbidity and length of stay was realized during the learning curve of implementing laparoscopic splenectomy by adopting a standardized technique. This technique involved hilar dissection using the ultrasonic shears, two experienced laparoscopic surgeons performing the surgery, dedicated equipment and trained staff using the checklist approach. We recommend such a standardized technique in performing laparoscopic splenectomy. PMID- 12123521 TI - Penetrating orbital injury with wooden foreign body initially diagnosed as an orbital floor blowout fracture. PMID- 12123522 TI - Neuroendocrine tumours of the ampulla of vater. PMID- 12123525 TI - 2001 Annual scientific meeting of the Urological Society of Australasia, Coolum, March 2001. Abstracts. PMID- 12123526 TI - Weather conditions and sudden sensorineural hearing loss. AB - BACKGROUND: Climatic or meteorological condition changes have been implicated in the pathogenesis of Idiopathic Sudden Sensorineural Hearing Loss (ISSHL). We investigated the seasonal distribution of ISSHL and evaluated the influence of meteorological parameters (such as temperature, humidity, and atmospheric pressure), their variation and covariation on the incidence of the disease. METHODS: A total of 82 cases of ISSHL, admitted to our department over a five year period, were enrolled in the study. Seasonal distribution of the disease was investigated by dividing the year in four seasons. Meteorological data included daily values of 13 distinct parameters recorded at the meteorological station of the University of Ioannina during this period. A relationship between each meteorological variable and the incidence of ISSHL was investigated by applying (chi2) test on data from 13 contingency tables as well as by using logistic regression and t-test approaches. In addition, the influence of different weather types on the incidence of ISSHL was investigated using Cluster Analysis in order to create eight clusters (weather types) characteristic for the prefecture of Ioannina. RESULTS: The results of the study could not indicate any seasonal distribution of the disease. The incidence of ISSHL could not be significantly correlated either to any distinct meteorological parameter or to any specific weather type. CONCLUSIONS: Meteorological conditions, such as those dominating in the Northwestern Greece, and/or their changes, have no proven effect on the incidence of ISSHL. PMID- 12123529 TI - Defining signal thresholds in DNA microarrays: exemplary application for invasive cancer. AB - BACKGROUND: Genome-wide or application-targeted microarrays containing a subset of genes of interest have become widely used as a research tool with the prospect of diagnostic application. Intrinsic variability of microarray measurements poses a major problem in defining signal thresholds for absent/present or differentially expressed genes. Most strategies have used fold-change threshold values, but variability at low signal intensities may invalidate this approach and it does not provide information about false-positives and false negatives. RESULTS: We introduce a method to filter false-positives and false-negatives from DNA microarray experiments. This is achieved by evaluating a set of positive and negative controls by receiver operating characteristic (ROC) analysis. As an advantage of this approach, users may define thresholds on the basis of sensitivity and specificity considerations. The area under the ROC curve allows quality control of microarray hybridizations. This method has been applied to custom made microarrays developed for the analysis of invasive melanoma derived tumor cells. It demonstrated that ROC analysis yields a threshold with reduced missclassified genes in microarray experiments. CONCLUSIONS: Provided that a set of appropriate positive and negative controls is included on the microarray, ROC analysis obviates the inherent problem of arbitrarily selecting threshold levels in microarray experiments. The proposed method is applicable to both custom made and commercially available DNA microarrays and will help to improve the reliability of predictions from DNA microarray experiments. PMID- 12123528 TI - High frequency of phenotypic deviations in Physcomitrella patens plants transformed with a gene-disruption library. AB - BACKGROUND: The moss Physcomitrella patens is an attractive model system for plant biology and functional genome analysis. It shares many biological features with higher plants but has the unique advantage of an efficient homologous recombination system for its nuclear DNA. This allows precise genetic manipulations and targeted knockouts to study gene function, an approach that due to the very low frequency of targeted recombination events is not routinely possible in any higher plant. RESULTS: As an important prerequisite for a large scale gene/function correlation study in this plant, we are establishing a collection of Physcomitrella patens transformants with insertion mutations in most expressed genes. A low-redundancy moss cDNA library was mutagenised in E. coli using a derivative of the transposon Tn1000. The resulting gene-disruption library was then used to transform Physcomitrella. Homologous recombination of the mutagenised cDNA with genomic coding sequences is expected to target insertion events preferentially to expressed genes. An immediate phenotypic analysis of transformants is made possible by the predominance of the haploid gametophytic state in the life cycle of the moss. Among the first 16,203 transformants analysed so far, we observed 2636 plants (= 16.2%) that differed from the wild-type in a variety of developmental, morphological and physiological characteristics. CONCLUSIONS: The high proportion of phenotypic deviations and the wide range of abnormalities observed among the transformants suggests that mutagenesis by gene-disruption library transformation is a useful strategy to establish a highly diverse population of Physcomitrella patens mutants for functional genome analysis. PMID- 12123527 TI - Pharmacological and ischemic preconditioning of the human myocardium: mitoK(ATP) channels are upstream and p38MAPK is downstream of PKC. AB - BACKGROUND: These studies investigate the role of mitoK(ATP) channels, protein kinase C (PKC) and Mitogen activated protein kinase (p38MAPK) on the cardioprotection of ischemic (IP) and pharmacological preconditioning (PP) of the human myocardium and their sequence of activation. RESULTS: Right atrial appendages from patients undergoing elective cardiac surgery were equilibrated for 30 min and then subjected to 90 min of simulated ischemia followed by 120 min reoxygenation. At the end of each protocol creatinine kinase leakage (CK U/g wet wt) and the reduction of MTT to formazan dye (mM/g wet wt) were measured. Similar protection was obtained with alpha1 agonist phenylephrine, adenosine and IP and their combination did not afford additional cardioprotection. Blockade of mitoK(ATP) channels with 5-hydroxydecanoate, PKC with chelerythrine, or p38MAPK with SB203580 abolished the protection of IP and of PP. In additional studies, the stimulation of mitoK(ATP) channels with diazoxide or activation of PKC with PMA or p38MAPK with anisomycin induced identical protection to that of IP and PP. The protection induced by diazoxide was abolished by blockade of PKC and by blockade of p38MAPK. Furthermore, the protection induced by PMA was abolished by SB203580 but not by 5-hydroxydecanoate, whereas the protection induced by anisomycin was unaffected by either 5-hydroxydecanoate or chelerythrine. CONCLUSIONS: Opening of mitoK(ATP) channels and activation of PKC and p38MAPK are obligatory steps in the signal transduction cascade of IP and PP of the human myocardium with PKC activation being downstream of the opening of mitoK(ATP) channels and upstream of p38MAPK activation. PMID- 12123531 TI - Can we afford to cure? PMID- 12123530 TI - Screening of the arrestin gene in dogs afflicted with generalized progressive retinal atrophy. AB - BACKGROUND: Intronic DNA sequences of the canine arrestin (SAG) gene was screened to identify potential disease causing mutations in dogs with generalized progressive retinal atrophy (gPRA). The intronic sequences flanking each of the 16 exons were obtained from clones of a canine genomic library. RESULTS: Using polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) and DNA sequence analyses we screened affected and unaffected dogs of 23 breeds with presumed autosomal recessively (ar) transmitted gPRA. In the coding region of the SAG gene 12 nucleotide exchanges were identified, 5 of which lead to amino acid substitutions (H14C; A111V; A113T; D259T; A379E). 7 other exonic substitutions represent silent polymorphisms (C132C; Q199Q; H225H; V247V; P264P; T288T and L293L). 16 additional sequence variations were observed in intronic regions of different dog breeds. CONCLUSIONS: In several breeds, these polymorphisms were found in homozygous state in unaffected and in heterozygous state in affected animals. Consequently these informative substitutions provide evidence to exclude mutations in the SAG gene as causing retinal degeneration in 14 of the 23 dog breeds with presumed ar transmitted gPRA. PMID- 12123535 TI - Breast cancer chemoprevention: current challenges and a look toward the future. AB - There is a need to develop new prevention agents for breast cancer risk reduction that would have fewer side effects than the approved agent, tamoxifen, and/or would be effective in preventing estrogen receptor-negative or tamoxifen resistant, estrogen receptor-positive breast cancers. There also is a need to improve the accuracy of present risk assessment models and to incorporate tissue based biomarkers to supplement risk prediction tools. Candidate risk biomarkers include the serum hormones insulin-like growth factor-1 and its binding protein 3, mammographic breast density, nipple aspirate fluid production, and breast tissue evidence of proliferative breast disease (intraepithelial neoplasia). A variety of techniques have been developed to randomly sample breast tissue to detect precancerous changes and/or detect modulation of proliferation in response to a prevention agent. Based on molecular abnormalities observed in breast intraepithelial neoplasia, a number of drug classes and combinations are suggested as potential chemoprevention approaches. Clinical trial models have been developed to select the appropriate drug dose for subsequent biomarker modulation chemoprevention trials in which the use of surrogate endpoint biomarkers as indicators of efficacy is being explored. If these biomarkers can be validated and shown to reliably predict and monitor response in phase I/II prevention trials, and if favorable modulation is correlated with subsequent decreased cancer incidence, biomarkers may replace cancer incidence as the endpoint in future phase III trials, dramatically reducing the time and expense associated with new prevention drug development. PMID- 12123538 TI - Prognostic value of the urokinase-type plasminogen activator, and its inhibitors and receptor in breast cancer patients. AB - Urokinase-type plasminogen activator (uPA), its inhibitors (PAI-1 and PAI-2), and its receptor (uPAR) play a key role in tumor invasion and metastasis. This study was designed to evaluate the prognostic impact of uPA, PAI-1, PAI-2, and uPAR and the combination of these factors in a group of 460 primary breast cancer patients. Concentrations of all 4 components of the uPA system were measured in tumor extracts using enzyme-linked immunosorbent assays (American Diagnostica, Inc, Greenwich, CT). After a median follow-up of 33 months, 18.5% of the patients had relapsed. The Cox proportional hazards model was applied for both univariate and multivariate analyses of disease-free survival (DFS). PAI-1 and PAI-2 were shown to provide independent prognostic information in breast cancer. Patients with either low levels of PAI-1 or high levels of PAI-2 were found to have better DFS (relative risk was 2.08 and 1.78, respectively). The prognostic value could be even further improved by a combination of both inhibitors. Aside from the uPA inhibitors, only nodal status and hormonal receptor status retained independent prognostic value. The other 2 invasion markers, uPA and uPAR, showed no statistically significant impact on DFS. In our patients, who were mostly treated with adjuvant therapy, uPA was not found to be an independent prognostic marker for DFS; this could be a consequence of the predictive value of uPA for response to adjuvant therapy and should be further investigated. PMID- 12123536 TI - The HER2 extracellular domain as a prognostic and predictive factor in breast cancer. AB - The HER2/neu proto-oncogene encodes a 185-kd transmembrane receptor with tyrosine kinase activity. Amplification of HER2 with overexpression of the p185HER2 receptor occurs in 20%-30% of breast cancers and has been established as an independent prognostic factor in numerous studies. Increasing evidence suggests that HER2 may be a predictive marker for response to chemotherapy and hormonal therapy. HER2 overexpression has provided a new target in breast cancer therapy, as evidenced by the development of trastuzumab (Herceptin(R)), a monoclonal antibody targeted against HER2. Detection of HER2 in the clinical setting is performed by immunohistochemistry or fluorescence in situ hybridization in tissue, and by detection of the shed extracellular domain in serum or plasma. Differences in methodology, reagents, and scoring systems have led to varying results in different patient cohorts, contributing to the debate on the role of HER2 as a prognostic and predictive factor. This review focuses on the prognostic and predictive value of serum HER2 detection in the management of HER2-positive breast cancer. PMID- 12123539 TI - Adjuvant chemotherapy with doxorubicin and cyclophosphamide in women with rapidly proliferating node-negative breast cancer. AB - This prospective clinical trial was designed to assess the impact of adjuvant chemotherapy in women with rapidly proliferating node-negative breast cancer. This group has been predicted to have a 5-year disease-free survival (DFS) of 70% without adjuvant chemotherapy. In this study, 449 women with rapidly proliferating breast cancer (91% measured by S-phase fraction and 9% by histochemistry) received adjuvant chemotherapy with doxorubicin/cyclophosphamide (AC) plus tamoxifen for estrogen receptor-positive or progesterone receptor positive cancer. The 5-year DFS was 90% (+/- 2%) and the 5-year overall survival was 94% (+/- 1%). At a median follow-up of 62 months, the strategy of administering 6 cycles of AC to women with T2 N0 cancer and 3 cycles in those with smaller T1 N0 cancers appeared to eliminate tumor size as a potential prognostic factor. Adjuvant chemotherapy with AC appears effective in reducing recurrence rates for women with rapidly proliferating node-negative breast cancer. PMID- 12123540 TI - The study of tamoxifen and raloxifene: preliminary enrollment data from a randomized breast cancer risk reduction trial. AB - Tamoxifen reduced the risk of invasive breast cancer by 49% among women at increased risk for breast cancer in the Breast Cancer Prevention Trial P-1, and raloxifene reduced breast cancer incidence by more than 70% in the Multiple Outcomes of Raloxifene Evaluation osteoporosis trial. These findings led the National Surgical Adjuvant Breast and Bowel Project to design and launch the Study of Tamoxifen and Raloxifene. Risk-eligible women are = 35 years of age and postmenopausal; they have either lobular carcinoma in situ (LCIS) or a 5-year risk of invasive breast cancer of at least 1.67% as determined by the Gail model. Participants are randomly assigned to receive either tamoxifen 20 mg or raloxifene 60 mg daily. The trial opened for accrual on July 1, 1999. After 32 months of recruitment at 194 clinical centers in North America, risk assessments have been performed in 107,855 women (83.8% white, 9.4% black, 3.8% Hispanic, 3.1% other race/ethnic groups). Of the eligible patients, 12,637 have been randomized (20.9% of risk-eligible women); the median age is 58 years (mean, 58 years), and the median 5-year risk of breast cancer is 3.3% (mean, 4.0%). LCIS was reported in 8.4% of women prior to randomization. Gail model risk was = 3.0% in 5 years for 59.3% of white women, 45.0% of black women, and 44.5% of Hispanic women. The trial will recruit a total of 22,000 postmenopausal women and is powered to demonstrate superior efficacy of either agent or their equivalence in reducing the incidence of primary breast PMID- 12123541 TI - Epstein-Barr virus induces human nasopharyngeal epithelial cells to escape from the replicative senescence. AB - OBJECTIVE: To observe the biological changes of primary human nasopharyngeal epithelial cells in the early stage of immortalization. METHODS: The morphological changes of nasopharyngeal epithelial cells were observed by phase contrast microscopy, and the activity profile of senescence-associated beta galactosidase (SA-beta-Gal) was detected by SA-beta-Gal staining. The expression of p16(INK4a) protein was tested by immunochemical assay, and the life span in vitro of nasopharyngeal epithelial cells was calculated as population doublings. In addition, the expression of Epstein-Barr (EB) virus latent membrane protein 1 (LMP1) was also detected by immunofluorescence staining. RESULTS: Morphologically, cells treated with EB virus and 12-o-tetradecanoyl-phorbol-13 acetate (TPA) formed multi-layer foci, and their cellular life span in vitro was extended (about 155 days of culture). A low percentage of cells (about 4.8%) expressed SA-beta-Gal activity at late primary culture, and did not always express p16(INK4a) protein in the progression of culture. CONCLUSIONS: Nasopharyngeal epithelial cells treated with EB virus in cooperation with TPA can pass through the stage of senescence and enter the early stage of immortalization. Some changes of phenotype occur in these cells. Our results provide data for further studying the mechanism of immortalization and the establishment of a human nasopharyngeal epithelial cell line. PMID- 12123542 TI - Retinoic acid receptor beta is required for anti-activator protein-1 activity by retinoic acid in gastric cancer cells. AB - OBJECTIVE: To investigate the role of retinoic acid receptor beta (RARbeta) in mediating inhibitory effect of all-trans retinoic acid (ATRA) on activator protein-1 (AP-1) activity in gastric cancer cells. METHODS: Transient transfection and chloramphenicol acetyltransferase (CAT) assay, Nort hern blot, gene transfection, 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) assay, and anchorage independent growth assay were used. RESULTS: Transient transfection of RARbeta expression vector into MKN-45 cells resulted in the RARbeta concentration dependent repression of AP-1 activity induced by 12-o tetradecanoylphorbol-13-acetate (TPA), regardless of the presence of ATRA. When the c-jun and c-fos expression vectors were cotransfected with the RARbeta expression vector into MKN-45 cells, AP-1 activity was also obviously repressed. The inhibitory effect, again, was RARbeta-concentration-dependent. The stable transfection of the RARbeta gene into MKN-45 cells led to cell growth inhibition and colony formation inhibition by ATRA. Furthermore, Cotransfection of both RARbeta/DNA binding domain (DBD) and reporter gene could not alter AP-1 activity, even in the presence of ATRA.However, when the cotransfection was substituted with the RARbeta/ligand binding domain (LBD), the inhibition was significantly enhanced by ATRA. CONCLUSION: RARbeta might be required for anti-AP-1 activity, and contribute to growth inhibition of gastric cancer cells by ATRA. PMID- 12123543 TI - Identification of Shiga-like toxin Escherichia coli isolated from children with diarrhea by polymerase chain reaction. AB - OBJECTIVE: To evaluate the etiology of Shiga like toxin producing Escherichia coli (SLTEC) in children with diarrhea. METHODS: We designed and synthesized 3 pairs of primers located in the SLT1, SLT2, and eaeA genes of enterohaemorrhagic Escherichia coli (EHEC), while the virulent genes SLT1, SLT2, and eaeA from E.coli species were amplified by polymerase chain reaction (PCR). RESULTS: One strain of EHEC with SLT1, SLT2, and eaeA in 29 reference strains of diarrhea causing E.coli (DCEC) and 10 strains of other enterobacteria detected by PCR had positive reactions, while all other DCEC and enterobacteria were negative. Of 474 strains of E. coli isolated from 1032 children with diarrhea and detected by PCR, 20 strains of SLT1 producing E. coli (4.2%) positive, and 7 strains of SLT2 producing E. coli (1.5%) positive; while of 74 strains of entero-SLTs-producing and invasive Escherichia coli (ESIEC), 15 strains of SLT1 (20.3%) and 5 strains of SLT2 (6.8%) were positive. CONCLUSION: Shiga-like toxin E. coli has been identified as a major etiologic agent of children with diarrhea in Taiyuan, China. PMID- 12123544 TI - A selective reduction in the relative density of parvalbumin-immunoreactive neurons in the hippocampus in schizophrenia patients. AB - OBJECTIVES: To determine the relative densities of the GABAergic subpopulation defined by calcium-binding proteins and to further study the importance of changes in GABAergic interneurons on neuropathology in the hippocampus in schizophrenia cases. METHODS: The relative densities and neuronal body size of cells immunoreactive for the calcium-binding proteins parvalbumin and calretinin as well as the area size of the hippocampal sub-fields were determined from the hippocampal tissue sections taken from schizophrenic patients and well-matched control subjects (15 per group). RESULTS: No significant difference in the density of calretinin-immunoreactive neurons and the neuronal body size of calretinin-positive neurons was found between subject groups. Relative to normal controls, schizophrenic patients showed a significant and profound deficit in the relative densities of parvalbumin-immunoreactive neurons in all hippocampal sub fields. These reductions were more apparent in male schizophrenic patients and were unrelated to antipsychotic drug treatment, age or duration of illness. CONCLUSION: The findings provide further evidence to support a profound and selective abnormality of a sub- population of GABAergic neurons in the hippocampus in schizophrenia cases, and are consistent with the etiological hypothesis of the neurodevelopment of schizophrenia. PMID- 12123545 TI - Comparative analysis of patients not responding to a single dose of 153Sm-EDTMP palliative treatment for painful skeletal metastases. AB - OBJECTIVE: A certain fraction of patients failed palliative treatment of 153Sm- ethylenediaminetetramethy lenephosphate (153Sm-EDTMP) for painful skeletal metastases were reviewd. A comparative analysis was designed to identify the factors related to therapeutic response. METHODS: From a 3-year multi-center clinical trial, 51 cases were collected who did not respond to an intravenous injection of 153Sm-EDTMP at a dosage of 0.5-1.5 mCi/kg. The therapeutic efficacy was evaluated via changes of symptoms, general condition, consumption of analgesics, sum of effect product, and Karnofsky scores. The age, sex, history of treatment, tumor type, location of bony involvement, uptake ratio and number of metastases, and doses used by the patients were compared to those of the responders. RESULTS: In 51 non-responders, 43 were male, 34 suffered from lung cancer, 41 had bone lesions in the vertebrae, 39 in the pelvis, and 24 had metastases in the lower extremities. Sex distribution, tumor type and location of the lesion differed significantly between responders and non-responders. No other factor showed differences between the two groups. Though patients of younger age, and lesions with lower uptake of radiopharmaceutical seemed to fail the treatment more easily as observed clinically, this was not confirmed by statistical analysis. CONCLUSION: The sex of the patients, certain types of primary tumors and metastases to lower parts of the body were found to influence the patients' response to a single dose of 153Sm-EDTMP palliation. Further exploration of a better way to determine dosage and predict response for each individual case is needed. PMID- 12123546 TI - Roles of protein kinase C and inositol(1,4,5)-trisphosphate in the pathogenesis of hypoxic- ischemic brain injury in neonatal rats. AB - OBJECTIVE: To investigate the effect of second messengers protein kinase C (PKC) and inositol (1,4,5)-trisphosphate (IP(3)) during hypoxic-ischemic brain injury in neonatal rats. METHODS: The protein concentration was determined by Lowry's method. PKC activity was measured by the incorporation of [gamma-32P] into a specific substrate peptide in the cytosol and particulate fraction, respectively. IP(3) was determined by the radioreceptor binding assay. RESULTS: Compared with the control, PKC activities in the particulate fractions in both the cerebral cortex and hippocampus decreased, while increased in the cytosol in cerebral cortex and remained within the normal range in the cytosol in the hippocampus at 0, 4, 12, 24, 48, 72 h, 7, and 14 d after hypoxic-ischemia for 20 min. All these changes restored to normal levels at 21 d post hypoxic-ischemia. Similarly, a decrease in IP(3) in the cerebral cortex and hippocampus and an increase in IP(3) in the thalamus after hypoxic-ischemia were noted, respectively. Changes in cytosolic PKC activity were not related to those of IP(3), as evaluated statistically. CONCLUSION: Hypoxia-ischemia induces disturbance of the second messengers IP(3) and PKC, which may play important roles in the pathogenesis of hypoxic-ischemic brain injury. PMID- 12123548 TI - Assessment of coronary artery disease with second harmonic myocardial perfusion contrast echocardiography. AB - OBJECTIVE: To assess the relationship between myocardial regional perfusion using second harmonic myocardial contrast echocardiography (MCE) by venous injection of Levovist and coronary artery stenosis detected by coronary angiography to determine whe ther MCE can be used to detect coronary artery disease (CAD) and its sensitivity and specificity for detecting CAD. METHODS: Thirty-six patients who underwent coronary artery angiography and MCE formed the study groups. Ten myocardial segments (5 each in the apical two- and four-chamber views) from the images were scored for detecting myocardial perfusion as follows: 1, normal perfusion; 2, decreased perfusion; and 3, perfusion defect. The arteries were classified as normal or diseased. The diseased arteries were classified into three groups according to the perfusion scores. RESULTS: There were significant differences in coronary diameter stenosis among the different perfusion score groups (P < 0.001). There were 10 total occluded arteries, and the myocardial perfusion scores were different because of different collateral circulation. In the normal perfusion group (Group A), the coronary diameter stenosis was 65% +/- 12%, and the myocardial perfusion score index was 1 +/- 0.00. In the decreased perfusion group (Group B), the average coronary diameter stenosis was 82% +/- 8%, and the myocardial perfusion score was 1.93 +/- 0.16. The diameter stenosis was less than 85% in 63% of the coronary arteries (including diameter stenosis < or = 75% in 12% of the vessels). The diameter stenosis was 85%-90% in 22% of the coronary arteries and > 90% in 15% of the arteries. In the perfusion defect group (Group C), the average diameter stenosis was 90% +/- 6%, and the myocardial perfusion score index was 2.89 +/- 0.24. The diameter stenosis was > or = 85% in 94% of the coronary arteries, and the diameter stenosis was < 85% and > 75% only in 6% of the coronary arteries. The overall sensitivity and specificity of MCE in identifying angiographic coronary diameter stenosis was 67% and 100%, respectively. The false negative rate was 32.6% for the 108 coronary arteries. Further subdivided analysis showed the sensitivities in Groups A, B and C were 0, 100%, and 100%, respectively. The sensitivity increased with increased lumen diameter stenosis of coronary arteries. CONCLUSIONS: There is a close relationship between coronary artery stenosis and MCE perfusion scores. MCE with venous injection of new generation contrast can define the presence of CAD and lesion graded classifications. Some disagreements between perfusion score and coronary diameter of stenosis may indicate other factors such as different collateral circulation, which should be further investigated. As artery stenosis increases, the sensitivity of MCE is increased. PMID- 12123547 TI - Two novel mutations of the retinitis pigmentosa GTPase regulator gene in two Chinese families with X-linked retinitis pigmentosa. AB - OBJECTIVE: To detect mutations of the retinitis pigmentosa GTPase regulator (RPGR) gene in two Chinese X-linked retinitis pigmentosa families. METHODS: Fragments of exons 1-19 of the RPGR gene were amplified with intronic primers, using genomic DNA as template. The polymerase chain reaction (PCR) products were analysed by single-strand conformation polymorphism (SSCP) and direct sequencing. Mutations were identified by comparing DNA sequences of the patients with those of the normal controls. RESULTS: Two novel mutations, c1536delC and E332X, were identified in exons 12 and 9 of the RPGR gene in both families. Each mutation was the first mutation found in their respective exons. Both mutations were predicted to cause premature termination, which resulted in truncated proteins without normal functions of the RPGR products. CONCLUSIONS: Both mutations are the genetic basis of the pathogenesis in the respective families. Our data might be helpful in analysing the function of the RPGR protein. PMID- 12123549 TI - Application of obturator to treat velopharyngeal incompetence. AB - OBJECTIVE: To evaluate the effect of a system of velopharyngeal incompetence (VPI) management after the application of obturator. METHODS: Using nasopharyngofiberoscope (NPF) and a computer analysis system, we quantitatively analyzed the improved state of velopharyngeal incompetence in 100 patients with unilateral and/or bilateral cleft palate. RESULTS: The velopharyngeal closure (VPC) can be greatly improved by using a temporary oral prosthesis (obturator) and speech training. An objective quantitative standard was established to evaluate the change of velopharyngeal closure of cleft palate patients after surgery and conservative treatment. CONCLUSIONS: The method used is more succinct, accurate and practical than previous methods. In order to reflect the state of velopharyngeal incompetence, the concept of improvement rate of velopharyngeal incompetence (IRVPI) is put forward. PMID- 12123551 TI - Contraceptive use behavior among never married young women who are seeking pregnancy termination in Beijing. AB - OBJECTIVE: To identify contraceptive use behavior and its determinants among never married young women with an unwanted pregnancy and seeking pregnancy termination in Beijing. METHODS: A cross sectional study, adopting the Lawrence' PRECEDE-PROCEED model was conducted in 1999 in Beijing, China. A total of 306 unmarried young women, aged 18 to 24 years and requesting pregnancy termination, were face to face interviewed. RESULTS: Only 13% of the young women insisted on contraceptive use, and almost an equal proportion occasionally or never used contraceptives (26% and 27%, respectively). Among 224 women who had contraceptive use during the past 12 months, the methods used most often were condom (49%), withdrawal (28%) and the rhythm method (16%). One of the most important reasons cited by 73 percent of women who had never used contraceptives was that they did not realize the risk of getting pregnant. The results of logistic regression analysis revealed that knowledge on contraception, boyfriend's approval of contraceptive use, perceived risk of getting pregnant, perceived availability of contraceptive services and discussion of contraception with boyfriend were important indicators of a young woman's contraceptive use behavior. CONCLUSION: These results indicate an urgent need to develop sex education on conception and contraception among young women and men. It is equally important to enhance young women's perception about the risk of unwanted pregnancy and the complications of induced abortion. Promotinga men's cooperation and participation in contraceptive use as well as strengthening communication on contraception between young women and their partners is essential. PMID- 12123550 TI - Expression of interleukin-12 and its signaling molecules in peripheral blood mononuclear cells in systemic lupus erythematosus patients. AB - OBJECTIVE: To determine the in vitro expression of interleukin-12 (IL-12) and its effect on signal transducers and activators of transcription (STAT) signaling molecules in peripheral blood mononuclear cells (PBMCs) in patients with systemic lupus erythematosus (SLE). METHODS: Peripheral blood mononuclear cells in 39 patients with definite systemic lupus erythematosus and 11 healthy volunteers were collected. Expression of IL-12 P40mRNA in PBMCs was determined with reverse transcription-polymerase chain reaction (RT-PCR). Quantity of IL-12 protein supernatant was measured by enzyme-linked immunosorbent assay (ELISA). The levels of phosphorylated STAT3 and STAT4 signaling molecules in PBMCs were detected by immunoblot. RESULTS: Levels of IL-12 protein and mRNA expression in patients with active or inactive SLE were significantly higher than those in controls. Phytohemagglutinin (PHA ) may promote the expression of IL-12. IL-12 alone induced the phosphorylation of STAT3 and STAT4 in PBMCs from patients with SLE, especially in active SLE. However it had no obvious effect on normal PBMCs. Phosphorylated STAT3 and STAT4 might be observed in normal PBMCs treated with IL 12 plus PHA. CONCLUSION: IL-12 is produced aberrantly in patients with SLE. IL-12 might exert its biological role in SLE via the aberrantly phosphorylated STAT3 and STAT4 signaling molecules. PMID- 12123552 TI - T cell receptor Vbeta gene bias in rheumatoid arthritis. AB - OBJECTIVE: To explore the pathogenesis of rheumatoid arthritis (RA) by studying the expression of T cell receptors (TCRs). METHODS: T cell receptor Vbeta (TCR Vbeta) gene usage and expression were analyzed from synovial membrane and peripheral blood of 8 RA patients, 2 osteoarthritis patients and 2 accident amputees. The complementary determining region 3 (CDR3) of 25 TCR Vbeta subfamily genes in unselected T cell populations were amplified semi-quantitatively by reverse transcription-polymerase chain reaction (RT-PCR). The products were further studied by genescan for frequency of Vbeta usage. RESULTS: The numbers of Vbeta subfamilies expressed by T cells from RA peripheral blood and synovial membrane were not significantly restricted. More importantly, biased Vbeta gene expression in RA synovium was observed and Vbeta6, Vbeta17, and Vbeta22 genes were the predominant subfamilies. It was noteworthy that the expression of Vbeta17 in RA synovium was significantly increased. CONCLUSION: Our data were consistent with the hypothesis that several antigen or superantigen-driven processes may be involved in the pathogenesis of RA. PMID- 12123553 TI - A prospective epidemiological study on tinea pedis and onychomycosis in Hong Kong. AB - OBJECTIVE: To study the epidemiology of foot diseases, including tinea pedis and onychomycosis in clinic attendees in Hong Kong. METHODS: Two groups were included: the institutional group consisted of clinical evaluation and mycological investigations by dermatologists; and the private group consisted of clinical evaluation only by the private physicians. Patients who had a regular visit to the clinics were randomly invited to have a clinical examination of their feet. RESULTS: A total of 1014 patients were studied. The prevalence rate of foot disease, fungal infections, tinea pedis and toe nail onychomycosis were respectively 50.7%, 26.9%, 20.4% and 16.6%. More male and elderly patients were affected except that the sex prevalence in toe nail onychomycosis was not shown to be significant. Vascular disease, diabetes mellitus and obesity were the three most prevalent predisposing factors in foot disease, fungal disease and fungal nail disease. Dermatophytes, in particular Trichophyton rubrum, were shown to be the most common pathogen in both skin and nail infections. CONCLUSIONS: Foot diseases, especially tinea pedis and toe nail onychomycosis, are common in patients attending local clinics in Hong Kong. Both physicians and patients should be more aware of foot problems and have more active approaches and management strategies. PMID- 12123554 TI - Long term outcome of contralateral C7 transfer: a report of 32 cases. AB - OBJECTIVE: To observe long-term functional recovery after contralateral C7 transfer. METHODS: From August 1986 to July 2000, 224 patients with brachial plexus avulsion injuries were treated with contralateral C7 transfer in our department. Thirty-two patients were followed up for over 2 years for evaluation of the following items: 1 influence on healthy limb function; 2 sensory and motor recovery of the recipient nerves in the affected limb; and 3 coordination between the healthy and affected limbs. RESULTS: There was no impairment of healthy limb function. Functional recovery of the recipient area reached > or =M3 in 8 patients (8/10, 80%) after musculocutaneous nerve neurotization, > or =M3 in 4 patients (4/6, 66%) after radial nerve neurotization, > or = M3 in 7 patients (7/14, 50%) and > or = M3 in 12 patients (85.7%) after median nerve neurotization, and > or = M3 in 1 patients (1/2, 50%) after thoracodorsal nerve neurotization. Synchronic contraction of the affected limb with the healthy limb occurred within 2-3 years in 12 patients, within 5 years in 13 patients, and over 5 years in 7 patients. CONCLUSION: Contralateral C7 transfer is an ideal procedure for the treatment of brachial plexus root avulsion injury. Selection of the whole root or the posterior division as neurotizer and a staged operation are the major factors influencing treatment outcome. PMID- 12123555 TI - Cell-specific expression of the diphtheria toxin A-chain coding sequence induces cancer cell suicide. AB - OBJECTIVE: To test whether the diphtheria toxin A (DT-A) chain coding sequence linked to murine immunoglobulin Kappa light chain (IgKappa) promoter and enhancer have selective cytocidal effects on IgKappa producing cells. METHODS: The diphtheria toxin A gene or beta galactosidase (beta-gal) gene were linked to a murine IgKappa promoter and enhancer to construct pcDNA3IgKappaDTA or pcDNA3IgKappaLacZ plasmids. These plasmids were transfected into IgKappa producing or non-producing cells by the liposome coated DNA method. Expression of beta-gal activity and effects on cell growth of transfected cells were assessed. RESULTS: The beta-gal gene, under the control of cytomegalovirus (CMV) promoter, can express in all cell lines. Expression of beta-gal under the control of the IgKappa promoter was detected only in the IgKappa producing cell line, CA46. Expression of beta-gal was greatly suppressed when cotransfected with pcDNA3IgKappaDTA in CA46 cells. Cell growth of CA46 cells transfected with pcDNA3IgKappaDTA plasmid was significantly inhibited compared with CA46 cells transfected with pcDNA3IgKappaLacZ. CONCLUSION: Selective killing of IgKappa producing cells can be attained by introducing the diphtheria toxin A gene under the control of IgKappa promoter and enhancer. PMID- 12123556 TI - Preimplantation gender diagnosis by fluorescence in situ hybridization. AB - OBJECTIVE: To describe the clinical application of fluorescence in situ hybridization (FISH ) in preimplantation gender diagnosis. METHODS: Preimplantation gender diagnosis was performed in 2 female hemophilia A carriers, 1 male patient with glucose-6-phosphate dehydrogenase (G-6-PD) deficiency and 2 male patients with Y chromosome abnormality. Embryo sex was identified by FISH in total of 6 treatment cycles. RESULTS: A total of 123 cumulus-oocytes were retrieved in 6 treatment cycles. Sixty-one embryos were available for embryo biopsy. The success rate of biopsy was 86.9% (53/61), with a further cleavage rate of 62.3% (33/53). In the FISH procedure, one cell was lost during fixation, leading to a 98.1% (52/53) fixation rate. Totally, 16 female embryos and 3 male embryos were transferred to 5 patients in 6 cycles. Three healthy babies were born. The diagnosis was confirmed by subsequent analysis of amniocytes and embryonic buds after embryo reduction. CONCLUSIONS: FISH is an efficient and reliable technique for determining the sex of human preimplantation embryos. Selective abortion and births of affected children can be avoided by preimplantation gender diagnosis. PMID- 12123557 TI - Vascular endothelial growth factor gene transfer improves host endothelialization of xenogeneic biologic heart valve in vivo. AB - OBJECTIVE: To investigate the feasibility of endothelialization of bioprosthesis by transfer of vascular endothelial growth factor (VEGF) gene. METHODS: Bovine pericardium treated with glutaraldehyde and L-glutamic acid was positioned into the pig right atrium. pcD(2)/hVEGF(121) gene (1 mg) was transferred into the right ventricular myocardium using surgical sutures Reverse transcri ption polymerase chain reaction (RT PCR) was employed to evaluate the expression of myocardial VEGF mRNA. The determination of concentrations of VEGF protein in blood from both the right atrium and peripheral vein, and histological and ultrastructural analysis of implanted bovine pericardium were completed simultaneously. RESULTS: The concentration of VEGF derived from the right atrium in pcD(2)/hVEGF(121) group was significantly higher than that in the pcD(2) group 10 days after VEGF gene transfer (P < 0.01). The expression of myocardial VEGF mRNA in pcD(2)/hVEGF(121) group was much higher in comparison with that in the pcD(2) group. The morphological analysis demonstrated that the coverage rate of host endothelium in the pcD(2)/hVEGF(121) group was 2.6 times as fast as that in the pcD(2) group at 16 days after VEGF(121) gene transfer (P < 0.01). Entire endothelialization occurred at 30 days after VEGF gene transfer. In addition, higher expression of myocardial VEGF mRNA was still available. CONCLUSIONS: VEGF gene transfer by surgical suture can remarkably accelerate endothelialization of bioprosthesis, which may provide a new approach for inhibiting biological valve calcification and improve biocompatibility and long-term durability of the bioprosthesis. PMID- 12123558 TI - Injection of bradykinin or cyclosporine A to hippocampus induces Alzheimer-like phosphorylation of Tau and abnormal behavior in rats. AB - OBJECTIVE: To reconstitute an Alzheimer's disease model by administering bradykinin (BK) or cyclosporine A (CSA) to the rat hippocampus. METHODS: BK or CSA was administered to the rat hippocampus using a stereotaxic apparatus. The behavior of the rats was observed with an electronic attack jump platform. The phosphorylation of Tau protein was examined through immunohistochemical assay. RESULTS: Behavior studies showed that an obvious disturbance in learning and memory was seen in BK injected rats.No obvious dysfunction was observed in CSA injected rats. The results obtained by immunohistochemical assay indicated that the staining of M4, 12E8, paired helical filament-1 (PHF-1) and calcium/calmodulin-dependent protein kinase II (CaMKII) was stronger, and that of Tau-1 was weaker in BK injected rats compared with the control group. We also found that the binding of M4 and PHF-1 but not 12E8 to Tau was significantly increased in CSA injected rats. As for BK injection, binding of Tau-1 to Tau was decreased after CSA injection. CONCLUSION: To our knowledge, this is the first data showing in vivo that the activation of CaMKII induces both Alzheimer-like Tau phosphorylation and behavioral disturbances. PMID- 12123559 TI - Two hundred endoscopic extraperitoneal inguinal hernioplasties: cost containment by reusable instruments. AB - OBJECTIVE: To report our experience of 200 endoscopic totally extraperitoneal inguinal hernioplasties utilizing reusable instruments. METHODS: Between August 1999 and June 2000, 200 endoscopic totally extraperitoneal hernioplasties were performed on 163 patients. The mean age of the study population was 63 years with a male to female ratio of 157:6. Perioperative details and postoperative outcomes were prospectively evaluated and analyzed. RESULTS: A total of 196 (98%) endoscopic extraperitoneal inguinal hernioplasties were successfully performed. Conversion rates to transabdominal preperitoneal and open repairs were 1.5% (n = 3) and 0.5% (n = 1), respectively. There were no other intraoperative complications. Postoperative morbidity included retention of urine (n = 7), wound bruising (n = 2), atelectasis (n = 2) and gouty arthritis (n = 1). The mean visual analogue pain scores at rest were 2.3, 1.6 and 1.9 on postoperative days 0, 1 and 2, respectively. The mean length of hospital stay was 1.9 days. 113 patients (69%) returned to normal activities within one week. Of the 35 patients who experienced both open and laparoscopic repair, 80% expressed preference for endoscopic hernioplasty in the event of future recurrence. CONCLUSIONS: Endoscopic extraperitoneal inguinal hernioplasty can be safely performed utilizing reusable trocars. Substantial reduction of operative cost could be achieved by the elimination of disposable instruments. Deficiencies of the reusable metallic trocar, namely peri-cannula air-leak and sliding movements of the trocar, can be overcome by purse-string suture of the fascial opening. PMID- 12123560 TI - Preservation of laryngeal function in treatment of hypopharyngeal carcinoma. AB - OBJECTIVE: To study the surgical technique and results of laryngeal function preservation in treatment of hypopharyngeal carcinoma. METHODS: A retrospective review of 305 patients with malignant neoplasms of the hypopharynx (279 males, 26 females, age ranging from 14 to 77 years) was performed from 1978 to 1996. In the 305 patients (stage I, n = 6; stage II, n = 12; stage III, n = 82; stage IV, n = 205), the sites of origin were pyriform sinus (n = 234), postcricoid (n = 21), posterior pharyngeal wall (n = 35) and superior hypopharynx (n = 15). Of the 305 patients, 206 (67.54%, stage I, n = 6; stage II, n = 12; stage III, n = 65; stage IV, n = 123) were surgically treated with laryngeal function preserved and 99 (32.46%, stage III, n = 17; stage IV, n = 82) had no laryngeal function preserved. All had 55-75 Gy radiotherapy according to their need. RESULTS: A total of 206 patients (67.54%) were surgically treated with laryngeal function preserved, totally (voice, respiration and deglutition) in 139 (67.5%) and partially (voice and deglutition) in 67 (32.5%). 99 patients (32.46%) had no laryngeal function preserved. The overall 5-year survival rate of the 305 patients was 44.8%, which segregated to 83% (stage I), 71% (stage II), 58% (stage III), and 36% (stage IV). The 5-year survival of the laryngeal function preserved group was 48% (n = 66), the rate of complications 28% (n = 58) and the rate of residual tumor 5.8% (n = 12), compared with the no laryngeal function preserved group 37% (n = 20), 31.3% (n = 31), and 6% (n = 6) (P > 0.05). CONCLUSION: Only a small proportion of patients (31/305, 10%) with hypopharyngeal carcinoma who require total laryngectomy and preservation of the laryngeal function is feasible for eradication of tumor and preservation of laryngeal function. PMID- 12123561 TI - Relationship between obesity and cardiovascular risk factors in elderly Chinese subjects. AB - OBJECTIVE: To investigate the relative effects of degree and distribution of body fat with several cardiovascular disease (CVD) risk factors in elderly Chinese subjects. METHODS: One hundred and thirty-five elderly Chinese individuals (age range, 60-65 y) without any history of significant renal, hepatic or cardiac disease were recruited. Seated blood pressure, anthropometric and fasting plasma biochemical parameters were measured. Student's t-test was used to compare the differences in biochemical and anthropometric markers between cohorts. RESULTS: Males were heavier (64.6 +/- 8.6, 57.2 +/- 8.2kg, P < 0.001), taller (1.65 +/- 0.06, 1.51 +/- 0.05 m, P < 0.001) and their greater body fat was predominantly deposited centrally (Waist-to- hip ratio, 0.91 +/- 0.06, 0.88 +/- 0.07, P < 0.05). Females were more generally obese with increased body mass index (BMI, 23.8 +/- 4.6, 25.0 +/- 3.5 kg/m2, P < 0.05) and percentage body fat [26.3% (24.5% 28.1%) vs 37.2% (36.0%-38.9%), P < 0.001] than the males. However, despite an 11% higher proportion of body fat in females, no significant differences were identified in blood pressure, lipid profile, indices of insulin resistance or albumin-to-creatinine ratios. CONCLUSION: It is likely that central adiposity contributes disproportionately to these metabolic disorders in males even though they are much leaner than elderly Chinese females. PMID- 12123562 TI - Prolongation of functional life-span of neutrophils by recombinant verotoxin 2. AB - OBJECTIVE: Verotoxin-producing Escherichia coli (VTEC) strains of serotype O157 : H7 have been implicated in a wide spectrum of diseases, including blood diarrhea, hemorrhagic colitis and hemolytic uremic syndrome (HUS). To further explore the pathological role of verotoxin (VT) in HUS and other VTEC associated diseases, we investigated the effects of recombinant verotoxin 2 (rVT2) on the biological activity of neutrophils. METHODS: The technique of flow cytometry, a fluorescent probe 2,7-bis-(2-carboxyethyl)-5-(and-6)-carboxyfluorescein acetoxymethyl ester (BCECF/AM), and the assay of reduced cytochrome c to detect superoxide production were used in this study. RESULTS: gammaVT2 significantly inhibited spontaneous apoptosis in neutrophils. Neutrophils with prolonged survival due to gammaVT2 maintained various biological functions, such as the expression of adhesion molecules (shading CD62L and raising CD11b/CD18), adherence to human umbilical vein endothelial cells (HUVECs), and generation of superoxide (O(2)(-)). CONCLUSION: Prolongation of the functional life-span of neutrophils by gammaVT2 may accelerate inflammatory responses at sites of inflammation. This may play a crucial role in neutrophil-mediated tissue injury in HUS and other VTEC associated diseases. PMID- 12123563 TI - Clinical features of forty patients with primary biliary cirrhosis. AB - OBJECTIVE: To study the clinical features of patients with primary biliary cirrhosis (PBC) in order to improve the doctors' awareness of the disease. METHODS: General status, clinical manifestations and laboratory findings of 40 patients with PBC were reviewed.Thirty-seven patients were females (37/40), and the mean age at diagnosis was 50.5 +/- 7.8 years. The time interval from initial symptoms or preliminary diagnosis to final diagnosis was 24.0 +/- 23.6 months. RESULTS: The most frequently reported symptoms were fatigue (67.5%, 27/40), jaundice (60%, 24/40) and pruritus (32.5%, 17/40). Eight patients (20%) had associated auto-immune diseases (Sjogren's syndrome and/or rheumatoid a(c)arthritis). Serum alkaline phosphatase (ALP) and gamma glutamyl transpeptidase (gamma-GT) levels were markedly elevated (520.3 +/- 382.3 IU/L and 648.6 +/- 529.1 IU/L, respectively) in all patients, while alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels were mildly elevated (82.6 +/- 54.5 IU/L and 100.7 +/- 47.2 IU/L, respectively). Twenty-four patients (60%) had a total bilirubin level >/= 34.2 micromol/L. Thirty-five patients (87.5%) had elevated serum immunoglobin M,and 97.5% of patients (39/40 ) were anti-mitochondrial antibody (AMA)/AMA-M2 positive. CONCLUSION: Elevated serum ALP and gamma-GT levels, together with a positive AMA/AMA-M2, can help the diagnosis of PBC. Liver biopsy is useful to confirm the diagnosis and to differentiate histopathological stages. PMID- 12123564 TI - Endocardial mapping and ablation of tachycardia guided by noncontact balloon catheter mapping system. AB - OBJECTIVE: To describe a new noncontact balloon catheter mapping system and to assess the clinical utility of this system for guiding endocardial mapping and ablation of tachycardia. METHODS: Five patients with tachycardia underwent endocardial mapping and radiofrequency ablation using the noncontact balloon catheter mapping system. A 9 French, 64-electrode balloon catheter and a conventional 7 French electrode catheter for mapping and ablation were positioned in the same ventricular chamber. Ventricular three-dimensional geometry was established by the computerized mapping system. Using a boundary element inverse solution, 3360 virtual endocardial electrograms were computerized and used to derive isopotential maps. The earliest endocardial activation site, the exit site and the activation sequence of tachycardia or the critical isthmus of the reentry circuit were identified. Radiofrequency ablation with circular or linear lesion was performed at the target sites guided by the locator system. RESULTS: Six clinical types of tachycardia, 5 of which were ventricular tachycardia and one was concealed fasciculoventricular fiber mediated tachycardia, were induced by programmed stimulation. The mean cycle length of these tachycardias was 336.6 +/- 42.69 msec. The earliest activation site and the exit site of 5 mapped tachycardias were all identified using the system. One type of ventricular tachycardia was hemodynamically unstable and difficult to terminate, and could not be mapped. Among the 6 types of tachycardias, radiofrequency ablation was successful in 4. There was no complication during and after the procedure. During the mean follow-up of 6 months, no tachycardia recurred in the patients with a successful ablation. CONCLUSIONS: The noncontact mapping system described in this study has advantage over conventional mapping techniques for refractory tachycardia. It is not only helpful for understanding the electrophysiologic mechanism of a complex case, but also suitable for mapping hemodynamically intolerated and nonsustained ventricular tachycardia. PMID- 12123565 TI - Expression of human VEGF(121) cDNA in mouse bone marrow stromal cells. AB - OBJECTIVE: To construct a retroviral vector carrying human vascular endothelial growth factor (hVEGF (121)) cDNA for evaluation of the possibility of VEGF gene therapy in ischemic bone disease. METHODS: hVEGF(121) cDNA was obtained from the plasmid pCDI/VEGF(121) and cloned into retroviral plasmid pLXSN. Recombinant plasmid was transferred to the retro virus packaging cell, PT-67, by lipofectamine mediated gene transfer. Mouse bone marrow stromal cells (MSCs) were transfected by the retrovirus. The integration of the hVEGF(121) cDNA into MSC genomic DNA and expression of the VEGF gene was detected. Proliferation assays of human umbilical vein endothelial cells (HUVECs) by VEGF(121) in culture medium were performed. RESULTS: Recombinant pLXSN/VEGF(121) was correctly constructed and confirmed by restriction endonuclease analysis and DNA sequencing analysis. hVEGF(121) gene was integrated into MSC genomic DNA after transfection, and the VEGF(121) protein was expressed. Proliferation assays showed VEGF(121) in culture medium was a biologically active protein and had a mitogenic effect on HUVEC. CONCLUSIONS: Recombinant retroviral vector carrying hVEGF(121) cDNA was successfully constructed. VEGF (121) protein expressed by MSCs had mitogenic effect biologically. This provides a further foundation for VEGF gene therapy for bone ischemic disease and bone tissue engineering. PMID- 12123566 TI - Experience in sclerotherapy for esophagogastric variceal bleeding. AB - OBJECTIVE: To evaluate the efficacy of endoscopic variceal sclerotherapy (EVS) for esophago gastric variceal bleeding. METHODS: A retrospective analysis was made on 1010 patients with esophagogastric variceal bleeding who underwent sclerotherapy, among whom there were 834 patients with cirrhosis, 160 with hepatocarcinoma, 12 with Budd-Chiari syndrome and 4 with congenital liver fibrosis. Totally, 3203 sessions of sclerotherapy were performed, including 602 sessions of emergency sclerotherapy and 2601 of selective surgery. The average number of sessions of sclerotherapy for the initial treatment in 710 cirrhosis patients who received continuous sclerotherapy was 3.2 +/- 1.1 times. Follow-up was done in 579 cirrhosis patients for 3-157 months, with an average period of 42.5+/- 32.8 months. RESULTS: The rate of emergency hemostasis in the whole group was 97.0%. The rate of complications was 13.4%, and the mortality rate was 1.8%. The rate of complete eradication and basically complete eradication of esophagogastric varices in cirrhosis patients was 84.1%. The late rebleeding rate was 23.7%, and the survival rates were 95.8% +/- 0.8%, 86.1% +/- 1.6%, 74.5% +/- 2.4%, 53.6% +/- 3.8% at 1, 3, 5 and 10 years, respectively, according to Kaplan Meier analysis. CONCLUSION: EVS is an important method for the treatment of esophagogastric variceal bleeding. PMID- 12123567 TI - Identification of a gene associated with astragalus and angelica's renal protective effects by silver staining mRNA differential display. AB - OBJECTIVE: To identify genes associated with the chronic progression of renal disease and a stragalus and angelica (A&A)'s renal protective effects. METHODS: The technique of silver staining mRNA differential display (DD) was used to investigate changes of gene expression in normal, sclerotic and A&A treated sclerotic kidneys. We isolated genes differentially expressed during the progression of renal disease which could be normalized by A&A. RESULTS: Several genes related to A&A's protective effects were isolated and one of them was confirmed by Northern blot. CONCLUSION: Silver staining mRNA differential display is a simple and effective technique for isolating differentially expressed genes. The isolated new gene may be related to the progression of chronic renal disease and contribute to A&A's protective effects. PMID- 12123568 TI - Effect of retinoid kappa receptor alpha (RXRalpha) transfection on the proliferation and phenotype of rat hepatic stellate cells in vitro. AB - OBJECTIVE: To study the effect of retinoid kappa receptor alpha (RXRalpha) transfection plus treatment with the RXRalpha ligand, 9-cis-RA, on the proliferation and phenotype of platelet-derived growth factor (PDGF) activated hepatic stellate cells (HSCs). METHODS: PDGF activated rat hepatic stellate cells were transfected with eukaryotic expression vector pcDNA3.1- human RXRalpha, and confirmed by Western blot. Proliferation of transfected HSC was assayed by bromodeoxyuridine (BrdU) incorporation as well as MTT, and the phenotype (alpha smooth muscle actin, desmin) was observed by immunocytochemistry with image analysis. RESULTS: Transfection of the RXRalpha gene and treatment with ligand 9 cis-RA of PDGF-activated HSCs extended the increased expression of RXRalpha protein for at least 168 hours. Cell proliferation and expressions of alpha- smooth muscle actin (alpha-SMA) and desmin were blocked, compared with groups of sham-transfected, PDGF-activated, no transfection, no ligand treatment, and irrelevant ligand treated HSCs. CONCLUSION: Transfection with the RXRalpha gene followed by 9-cis-RA ligand treatment will inhibit the proliferation and reverse the phenotype of activated HSC. PMID- 12123569 TI - Analysis of the neuronal selectivity underlying low fMRI signals. AB - BACKGROUND: A prevailing assumption in neuroimaging studies is that relatively low fMRI signals are due to weak neuronal activation, and, therefore, they are commonly ignored. However, lower fMRI signals may also result from intense activation by highly selective, albeit small, subsets of neurons in the imaged voxel. We report on an approach that could form a basis for resolving this ambiguity imposed by the low (mm range) spatial resolution of fMRI. Our approach employs fMR-adaptation as an indicator for highly active neuronal populations even when the measured fMRI signal is low. RESULTS: In this study, we first showed that fMRI-adaptation is diminished when overall neuronal activity is lowered substantially by reducing image contrast. We then applied the same adaptation paradigm, but this time we lowered the fMRI signal by changing object shape. While the overall fMRI signal in category-related regions such as the face related pFs was drastically reduced for non-face stimuli, the adaptation level obtained for these stimuli remained high. We hypothesize that the relatively greater adaptation level following exposure to "nonoptimal" object shapes is indicative of small subsets of neurons responding vigorously to these "nonoptimal" objects even when the overall fMRI activity is low. CONCLUSIONS: Our results show that fMR-adaptation can be used to differentiate between neuronal activation patterns that appear similar in the overall fMRI signal. The results suggest that it may be possible to employ fMR-adaptation to reveal functionally heterogeneous islands of activity, which are too small to image using conventional imaging methods. PMID- 12123570 TI - The dual mechanism of separase regulation by securin. AB - BACKGROUND: Sister chromatid separation and segregation at anaphase onset are triggered by cleavage of the chromosomal cohesin complex by the protease separase. Separase is regulated by its binding partner securin in two ways: securin is required to support separase activity in anaphase; and, at the same time, securin must be destroyed via ubiquitylation before separase becomes active. The molecular mechanisms underlying this dual regulation of separase by securin are unknown. RESULTS: We show that, in budding yeast, securin supports separase localization. Separase enters the nucleus independently of securin, but securin is required and sufficient to cause accumulation of separase in the nucleus, where its known cleavage targets reside. Securin also ensures that separase gains full proteolytic activity in anaphase. We also show that securin, while present, directly inhibits the proteolytic activity of separase. Securin prevents the binding of separase to its substrates. It also hinders the separase N terminus from interacting with and possibly inducing an activating conformational change at the protease active site 150 kDa downstream at the protein's C terminus. CONCLUSIONS: Securin inhibits the proteolytic activity of separase in a 2-fold manner. While inhibiting separase, securin is able to promote nuclear accumulation of separase and help separase to become fully activated after securin's own destruction at anaphase onset. PMID- 12123571 TI - Sonic hedgehog induces the segregation of patched and smoothened in endosomes. AB - BACKGROUND: Sonic hedgehog (Shh) signal transduction involves the ligand binding Patched1 (Ptc1) protein and a signaling component, Smoothened (Smo). A select group of compounds inhibits both Shh signaling, regulated by Ptc1, and late endosomal lipid sorting, regulated by the Ptc-related Niemann-Pick C1 (NPC1) protein. This suggests that Ptc1 regulates Smo activity through a common late endosomal sorting pathway also utilized by NPC1. During signaling, Ptc accumulates in endosomal compartments, but it is unclear if Smo follows Ptc into the endocytic pathway. RESULTS: We characterized the dynamic subcellular distributions of Ptc1, Smo, and activated Smo mutants individually and in combination. Ptc1 and Smo colocalize extensively in the absence of ligand and are internalized together after ligand binding, but Smo becomes segregated from Ptc1/Shh complexes destined for lysosomal degradation. In contrast, activated Smo mutants do not colocalize with nor are cotransported with Ptc1. Agents that block late endosomal transport and protein sorting inhibit the ligand-induced segregation of Ptc1 and Smo. We show that, like NPC1-regulated lipid sorting, Shh signal transduction is blocked by antibodies that specifically disrupt the internal membranes of late endosomes, which provide a platform for protein and lipid sorting. CONCLUSIONS: These data support a model in which Ptc1 inhibits Smo only when in the same compartment. Ligand-induced segregation allows Smo to signal independently of Ptc1 after becoming sorted from Ptc1/Shh complexes in the late endocytic pathway. PMID- 12123572 TI - Inducible expression of double-stranded RNA reveals a role for dFADD in the regulation of the antibacterial response in Drosophila adults. AB - In Drosophila, the immune deficiency (Imd) pathway controls antibacterial peptide gene expression in the fat body in response to Gram-negative bacterial infection. The ultimate target of the Imd pathway is Relish, a transactivator related to mammalian P105 and P100 NF-kappaB precursors. Relish is processed in order to translocate to the nucleus, and this cleavage is dependent on both Dredd, an apical caspase related to caspase-8 of mammals, and the fly Ikappa-B kinase complex (dmIKK). dTAK1, a MAPKKK, functions upstream of the dmIKK complex and downstream of Imd, a protein with a death domain similar to that of mammalian receptor interacting protein (RIP). Finally, the peptidoglycan recognition protein-LC (PGRP-LC) acts upstream of Imd and probably functions as a receptor for the Imd pathway. Using inducible expression of dFADD double-stranded RNA, we demonstrate that dFADD is a novel component of the Imd pathway: dFADD double stranded RNA expression reduces the induction of antibacterial peptide-encoding genes after infection and renders the fly susceptible to Gram-negative bacterial infection. Epistatic studies indicate that dFADD acts between Imd and Dredd. Our results reinforce the parallels between the Imd and the TNF-R1 pathways. PMID- 12123573 TI - Sensory inputs stimulate progenitor cell proliferation in an adult insect brain. AB - Although most brain neurons are produced during embryonic and early postnatal development, recent studies clearly demonstrated in a wide range of species from invertebrates to humans that new neurons are added to specific brain structures throughout adult life. Hormones, neurotransmitters, and growth factors as well as environmental conditions modulate this neurogenesis. In this study, we address the role of sensory inputs in the regulation of adult neural progenitor cell proliferation in an insect model. In some insect species, adult neurogenesis occurs in the mushroom bodies, the main sensory integrative centers of the brain, receiving multimodal information and often considered as the analog of the vertebrate hippocampus. We recently showed that rearing adult crickets in enriched sensory and social conditions enhanced neuroblast proliferation in the mushroom bodies. Here, by manipulating hormonal levels and affecting olfactory and/or visual inputs, we show that environmental regulation of neurogenesis is in direct response to olfactory and visual stimuli rather than being mediated via hormonal control. Experiments of unilateral sensory deprivation reveal that neuroblast proliferation can be inhibited in one brain hemisphere only. These results, obtained in a relatively simple brain, emphasize the role of sensory inputs on stem cell division. PMID- 12123574 TI - Glycogen synthase kinase-3beta modulates notch signaling and stability. AB - Notch receptors modulate transcriptional targets following the proteolytic release of the Notch intracellular domain (NotchIC). Phosphorylated forms of NotchIC have been identified within the nucleus and have been associated with CSL members, as well as correlated with regions of the receptor that are required for activity. Genetic studies have suggested that the Drosophila homolog of glycogen synthase kinase-3beta (GSK3beta), Shaggy, may act as a positive modulator of the Notch signaling. GSK3beta is a serine/threonine kinase and is a component of the Wnt/wingless signaling cascade. Here, we observed that GSK3beta was able to bind and phosphorylate Notch1IC in vitro, and attenuation of GSK3beta activity reduced phosphorylation of NotchIC in vivo. Functionally, ligand-activated signaling through the endogenous Notch1 receptor was reduced in GSK3beta null fibroblasts, implying a positive role for GSK3beta in mammalian Notch signaling. As a possible mechanistic explanation of the effect of GSK3beta on Notch signaling, we observed that inhibition of GSK3beta shortened the half-life of Notch1IC. Conversely, activated GSK3beta reduced the quantity of Notch1IC that was degraded by the proteasome. These studies reveal that GSK3beta modulates Notch1 signaling, possibly through direct phosphorylation of the intracellular domain of Notch, and that the activity of GSK3beta protects the intracellular domain from proteasome degradation. PMID- 12123575 TI - The long-term benefits of human generosity in indirect reciprocity. AB - Among the theories that have been proposed to explain the evolution of altruism are direct reciprocity and indirect reciprocity. The idea of the latter is that helping someone or refusing to do so has an impact on one's reputation within a group. This reputation is constantly assessed and reassessed by others and is taken into account by them in future social interactions. Generosity in indirect reciprocity can evolve if and only if it eventually leads to a net benefit in the long term. Here, we show that this key assumption is met. We let 114 students play for money in an indirect and a subsequent direct reciprocity game. We found that although being generous, i.e., giving something of value to others, had the obvious short-term costs, it paid in the long run because it builds up a reputation that is rewarded by third parties (who thereby themselves increase their reputation). A reputation of being generous also provided an advantage in the subsequent direct reciprocity game, probably because it builds up trust that can lead to more stable cooperation. PMID- 12123576 TI - Mitotically stable association of polycomb group proteins eed and enx1 with the inactive x chromosome in trophoblast stem cells. AB - X inactivation in female mammals is one of the best studied examples of heritable gene silencing and provides an important model for studying maintenance of patterns of gene expression during differentiation and development. The process is initiated by a cis-acting RNA, the X inactive specific transcript (Xist). Xist RNA is thought to recruit silencing complexes to the inactive X, which then serve to establish and maintain the inactive state in all subsequent cell divisions. Most lineages undergo random X inactivation, there being an equal probability of either the maternally (Xm) or paternally (Xp) inherited X chromosome being inactivated in a given cell. In the extraembryonic trophectoderm and primitive endoderm lineages of mouse embryos, however, there is imprinted X inactivation of Xp. This process is also Xist dependent. A recent study has shown that imprinted X inactivation in trophectoderm is not maintained in embryonic ectoderm development (eed) mutant mice. Here we show that Eed and a second Polycomb group protein, Enx1, are directly localized to the inactive X chromosome in XX trophoblast stem (TS) cells. The association of Eed/Enx1 complexes is mitotically stable, suggesting a mechanism for the maintenance of imprinted X inactivation in these cells. PMID- 12123577 TI - The cadherins fat and dachsous regulate dorsal/ventral signaling in the Drosophila eye. AB - The Drosophila eye is a polarized epithelium in which ommatidia of opposing chirality fall on opposite sides of the eye's midline, the equator. The equator is established in at least two steps: photoreceptors R3 and R4 adopt their fates, and then ommatidia rotate clockwise or counterclockwise in accordance with the identity of these photoreceptors. We report the role of two cadherins, Fat (Ft) and Dachsous (Ds), in conveying the polarizing signal from the D/V midline in the Drosophila eye. In eyes lacking Ft, the midline is abolished. In ft and ds mutant clones, wild-type tissue rescues genetically mutant tissue at the clonal borders, giving rise to ectopic equators. These ectopic equators distort a mosaic analysis of these genes and led to the possible misinterpretation that ft and ds are required to specify the R3 and R4 cell fates, respectively. Our interpretation of these data supports a significantly different model in which ft and ds are not necessarily required for fate determination. Rather, they are involved in long range signaling during the formation of the equator, as defined by the presence of an organized arrangement of dorsal and ventral chiral ommatidial forms. PMID- 12123578 TI - The spindle checkpoint kinase bub1 and cyclin e/cdk2 both contribute to the establishment of meiotic metaphase arrest by cytostatic factor. AB - In vertebrate unfertilized eggs, metaphase arrest in Meiosis II is mediated by an activity known as cytostatic factor (CSF). CSF arrest is dependent upon Mos dependent activation of the MAPK/Rsk pathway, and Rsk activates the spindle checkpoint kinase Bub1, leading to inhibition of the anaphase-promoting complex (APC), an E3 ubiquitin ligase required for the metaphase/anaphase transition. However, it is not known whether Bub1 is required for the establishment of CSF arrest or whether other pathways also contribute. Here, we show that immunodepletion of Bub1 from egg extracts blocks the ability of Mos to establish CSF arrest, and arrest can be restored by the addition of wild-type, but not kinase-dead, Bub1. The appearance of CSF arrest at Meiosis II may result from coexpression of cyclin E/Cdk2 with the MAPK/Bub1 pathway. Cyclin E/Cdk2 was able to cause metaphase arrest in egg extracts even in the absence of Mos and could also inhibit cyclin B degradation in oocytes when expressed at anaphase of Meiosis I. Once it has been established, metaphase arrest can be maintained in the absence of MAPK, Bub1, or cyclin E/Cdk2 activity. Both pathways are independent of each other, but each appears to block activation of the APC, which is required for cyclin B degradation and the metaphase/anaphase transition. PMID- 12123579 TI - MAP4 counteracts microtubule catastrophe promotion but not tubulin-sequestering activity in intact cells. AB - Microtubules are polar polymers that continually switch between phases of elongation and shortening, a property referred to as dynamic instability. The ubiquitous microtubule associated protein 4 (MAP4) shows rescue-promoting activity during in vitro assembly of microtubules (i.e., promotes transitions from shortening to elongation), but its regulatory role in intact cells is poorly defined. Here, we demonstrate that ectopic MAP4 promotes outgrowth of extended MTs during beta1-integrin-induced cell spreading. An inducible cotransfection protocol was employed to further analyze the regulatory role of MAP4 in human leukemia cells with microtubules partially destabilized by either ectopic tubulin sequestering proteins or proteins that promote catastrophes (i.e., transitions from elongation to shortening). Coexpression of proteins that sequester free tubulin heterodimers with different efficiencies was found to abolish microtubule stabilization by MAP4. In contrast, however, the microtubule-stabilizing activity of MAP4 was found to suppress the activities of two distinct and specific catastrophe promoters, namely, XKCM1 and a nonsequestering truncation derivative of Op18/stathmin. These observations reveal specificity in the microtubule stabilizing activity of MAP4 that differentiates between two mechanistically distinct types of MT destabilization. PMID- 12123580 TI - Fitness effects of fixed beneficial mutations in microbial populations. AB - Beneficial mutations are intuitively relevant to understanding adaptation, yet not all beneficial mutations are of consequence to the long-term evolutionary outcome of adaptation. Many beneficial mutations-mostly those of small effect-are lost due either to (1) genetic drift or to (2) competition among clones carrying different beneficial mutations, a phenomenon called the "Hill-Robertson effect" for sexual populations and "clonal interference" for asexual populations. Competition among clones becomes more prevalent with increasing genetic linkage and increasing population size, and it is thus generally characteristic of microbial populations. Together, these two phenomena suggest that only those beneficial mutations of large fitness effect should achieve fixation, despite the fact that most beneficial mutations produced are predicted to have very small fitness effects. Here, we confirm this prediction-both empirically and theoretically-by showing that fitness effects of fixed beneficial mutations follow a distribution whose mode is positive. PMID- 12123581 TI - Loss-of-susceptibility mutants of Arabidopsis thaliana reveal an essential role for eIF(iso)4E during potyvirus infection. AB - The Arabidopsis thaliana-potyvirus system was developed to identify compatibility and incompatibility factors involved during infection and disease caused by positive-strand RNA viruses. Several Arabidopsis mutants with increased susceptibility to Tobacco etch potyvirus (TEV) were isolated previously, revealing a virus-specific resistance system in the phloem. In this study, Arabidopsis mutants with decreased susceptibility to Turnip mosaic potyvirus (TuMV) were isolated. Three independent mutants that conferred immunity to TuMV were isolated and assigned to the same complementation group. These mutants were also immune or near-immune to TEV but were susceptible to an unrelated virus. The locus associated with decreased susceptibility was named loss-of-susceptibility to potyviruses 1 (lsp1). The LSP1 locus was isolated by map-based cloning and was identified as the gene encoding translation factor eIF(iso)4E, one of several known Arabidopsis isoforms that has cap binding activity. eIF4E and eIF(iso)4E from different plant species were shown previously to interact with the genome linked protein (VPg) of TEV and TuMV, respectively. Models to explain the roles of eIF(iso)4E during virus infection are presented. PMID- 12123582 TI - Methylation of H3-lysine 79 is mediated by a new family of HMTases without a SET domain. AB - The N-terminal tails of core histones are subjected to multiple covalent modifications, including acetylation, methylation, and phosphorylation. Similar to acetylation, histone methylation has emerged as an important player in regulating chromatin dynamics and gene activity. Histone methylation occurs on arginine and lysine residues and is catalyzed by two families of proteins, the protein arginine methyltransferase family and the SET-domain-containing methyltransferase family. Here, we report that lysine 79 (K79) of H3, located in the globular domain, can be methylated. K79 methylation occurs in a variety of organisms ranging from yeast to human. In budding yeast, K79 methylation is mediated by the silencing protein DOT1. Consistent with conservation of K79 methylation, DOT1 homologs can be found in a variety of eukaryotic organisms. We identified a human DOT1-like (DOT1L) protein and demonstrated that this protein possesses intrinsic H3-K79-specific histone methyltransferase (HMTase) activity in vitro and in vivo. Furthermore, we found that K79 methylation level is regulated throughout the cell cycle. Thus, our studies reveal a new methylation site and define a novel family of histone lysine methyltransferase. PMID- 12123583 TI - Biodiversity challenge to funding priorities. AB - Ten years after the key Earth Summit in Rio, a key group of influential politicians in Britain are flagging up the need to support research into taxonomy as a basis for biodiversity studies ahead of this summer's meeting in South Africa. Nigel Williams reports. PMID- 12123584 TI - Testing notions of global ecology. AB - It's clear that one thing that would boost scientific input to the issue of biodiversity would be evidence that researchers can model effectively the large scale behaviour of ecosystems. A new issue of a prestigious journal aims to explore just that. Nigel Williams reports. PMID- 12123585 TI - UK report calls for protection of genetic privacy. AB - DNA gathered from a used coffee mug could lead to serious breaches of a person's genetic privacy. The UK government's Human Genetics Commission now recommends making theft of genetic information a criminal offence, writes Michael Gross. PMID- 12123586 TI - gamma-Tubulin. PMID- 12123588 TI - Plant epigenetics. PMID- 12123587 TI - Novel human vomeronasal receptor-like genes reveal species-specific families. PMID- 12123589 TI - Genetic architecture: dissecting the genetic basis of phenotypic variation. AB - Identifying genes that influence phenotypic variation within species has proven to be more difficult than expected. A recent study has achieved exceptional success in yeast, and demonstrates how complex genetic architecture can be. PMID- 12123591 TI - Congenital amusia: all the songs sound the same. AB - Recent evidence from individuals born with a profound musical impairment suggests that the ability to process pitch information is normally present from birth. This finding supports the idea that the perception and appreciation of music, both of which critically depend on pitch processing, have a biological basis in the brain. PMID- 12123590 TI - Ena/Vasp: solving a cell motility paradox. AB - Modulating the concentration of the actin-binding protein Ena/Vasp within the lamellipodium of a migrating fibroblast results in marked changes in lamellipodium behaviour and actin network organization at the cell's leading edge. This can explain a cell motility paradox. PMID- 12123592 TI - Ci proteolysis: regulation by a constellation of phosphorylation sites. AB - Proteolytic processing of Ci requires phosphorylation by the kinases Sgg/GSK3 and CK1. The newly identified phosphorylation sites form clusters with previously described PKA sites in which phosphorylation by PKA acts to prime subsequent phosphorylation by Sgg/GSK3 and CK1. PMID- 12123593 TI - Self-incompatibility: how to stay incompatible. AB - Plant self-incompatibility is controlled by different genes for the recognition reactions of pollen and stigmas, yet correct association of the two genes have been maintained in two Brassica species. PMID- 12123594 TI - Bacterial evolution: chromosome arithmetic and geometry. AB - Recent sequencing projects have characterized bacterial genomes that are organized onto elements of various sizes, shapes and numbers. Aside from its biological relevance and curiosity, this diversity calls into question the way that we define bacterial chromosomes. PMID- 12123595 TI - Rac activation: P-Rex1 - a convergence point for PIP(3) and Gbetagamma? AB - P-Rex1, a novel Rac activator, has been identified in the first biochemical purification of a guanine nucleotide exchange factor for GTPases of the Rho family. P-Rex1 is synergistically activated by PIP(3) and Gbetagamma and may act as a coincidence detector for these signaling molecules. PMID- 12123596 TI - Calcium signalling: calcium goes global. AB - Recent evidence suggests that multiple calcium-releasing messengers might be activated simultaneously to regulate patterns of intracellular calcium signalling. In this way, agonists might use different messenger cocktails to encode specific signals and target selected processes. PMID- 12123597 TI - Epigenetics: SUPERMAN dresses up. AB - DNA and histone methylation have been implicated in epigenetic gene regulation. Recent studies in Neurospora and now Arabidopsis indicate that histone methylation can direct DNA methylation, suggesting that these two methylation systems have been functionally linked during evolution. PMID- 12123598 TI - Hedgehog signalling: pulling apart patched and smoothened. AB - Two integral membrane proteins, Patched and Smoothened, were for a long time thought to comprise a preformed receptor complex for secreted Hedgehog signalling proteins. Recent analyses of the subcellular distribution of these proteins argue strongly against this simple model. PMID- 12123599 TI - Cadherins communicate structural plasticity of presynaptic and postsynaptic terminals. AB - Synapse adhesion molecules play a key role in specifying and facilitating the recognition of axodendritic contacts. New studies by and reported in this issue of Neuron reveal multiple functions for the cadherin-catenin complex. This adhesion complex regulates synaptogenesis and coordinates synaptic strength with presynaptic and postsynaptic organization, including the shape of dendritic spines. PMID- 12123600 TI - Head, neck, and spines: a role for LIMK-1 in the hippocampus. AB - LIM kinase 1 regulates actin filament dynamics through inhibition of ADF/cofilins. Surprisingly, nervous system development in LIM kinase 1 knockout mice is grossly normal, but the animals have deficits in spatial learning, alterations in LTP, and abnormalities in hippocampal dendritic spine structure. The findings are consistent with a role for LIMK-1 in synapse formation and function. PMID- 12123601 TI - Sleep and motor skill learning. AB - The improvement of a perceptual or motor skill continues after training has ended. The central question is whether this improvement is just a function of time or whether sleep, a certain circadian phase, or their interaction (sleep occurring in a particular circadian phase) is favorable to the reprocessing of recent memory traces. In this issue of Neuron, provide behavioral evidence that most of the improvement of a motor skill depends on nocturnal sleep. PMID- 12123602 TI - Protein misfolding, amyloid formation, and neurodegeneration: a critical role for molecular chaperones? AB - The most conspicuous feature of many neurodegenerative disorders, including Alzheimer's, Parkinson's, and Huntington's disease, is the occurrence of protein aggregates in ordered fibrillar structures known as amyloid found inside and outside of brain cells. The appearance of aggregates in diseased brains implies an underlying incapacity in the cellular machinery of molecular chaperones that normally functions to prevent the accumulation of misfolded proteins. Here we review recent studies that have revealed a critical role for molecular chaperones in several neurodegenerative disorders. PMID- 12123603 TI - Signaling the pathway to regeneration. AB - Robust axon regeneration occurs after peripheral nerve injury through coordinated activation of a genetic program and local intracellular signaling cascades. Although regeneration-associated genes are being identified with increasing frequency, most aspects of regeneration-associated intracellular signaling remain poorly understood. Two independent studies now report that upregulation of cAMP is a component of the PNS regeneration program that can be exploited to enhance axon regeneration through the normally inhibitory CNS environment. PMID- 12123604 TI - A-to-I editing: new and old sites, functions and speculations. AB - Nuclear pre-mRNA editing by selective adenosine deamination (A-to-I editing) occurs in all organisms from C. elegans to humans. This rare posttranscriptional mechanism can alter codons and hence the structure and function of proteins. New findings report new sites, give evidence that the efficiency of editing can be regulated by neurotransmitter, and reveal that an amino acid substitution introduced by editing into a neurotransmitter-gated ion channel subunit serves as a determinant for controlling the maturation, intracellular trafficking, and assembly with other subunits of this transmembrane protein. PMID- 12123605 TI - Neuroethics for the new millenium. PMID- 12123606 TI - Ataxia and paroxysmal dyskinesia in mice lacking axonally transported FGF14. AB - Fibroblast growth factor 14 (FGF14) belongs to a distinct subclass of FGFs that is expressed in the developing and adult CNS. We disrupted the Fgf14 gene and introduced an Fgf14(N-beta-Gal) allele that abolished Fgf14 expression and generated a fusion protein (FGF14N-beta-gal) containing the first exon of FGF14 and beta-galactosidase. Fgf14-deficient mice were viable, fertile, and anatomically normal, but developed ataxia and a paroxysmal hyperkinetic movement disorder. Neuropharmacological studies showed that Fgf14-deficient mice have reduced responses to dopamine agonists. The paroxysmal hyperkinetic movement disorder phenocopies a form of dystonia, a disease often associated with dysfunction of the putamen. Strikingly, the FGF14N-beta-gal chimeric protein was efficiently transported into neuronal processes in the basal ganglia and cerebellum. Together, these studies identify a novel function for FGF14 in neuronal signaling and implicate FGF14 in axonal trafficking and synaptosomal function. PMID- 12123607 TI - Drosophila homeodomain protein dHb9 directs neuronal fate via crossrepressive and cell-nonautonomous mechanisms. AB - Here we present the identification and characterization of dHb9, the Drosophila homolog of vertebrate Hb9, which encodes a factor central to motorneuron (MN) development. We show that dHb9 regulates neuronal fate by restricting expression of Lim3 and Even-skipped (Eve), two homeodomain (HD) proteins required for development of distinct neuronal classes. Also, dHb9 and Lim3 are activated independently of each other in a virtually identical population of ventrally and laterally projecting MNs. Surprisingly, dHb9 represses Lim3 cell nonautonomously in a subset of dorsally projecting MNs, revealing a novel role for intercellular signaling in the establishment of neuronal fate in Drosophila. Lastly, we provide evidence that dHb9 and Eve regulate each other's expression through Groucho dependent crossrepression. This mutually antagonistic relationship bears similarity to the crossrepressive relationships between pairs of HD proteins that pattern the vertebrate neural tube. PMID- 12123608 TI - Plexin-B1 directly interacts with PDZ-RhoGEF/LARG to regulate RhoA and growth cone morphology. AB - Plexins are widely expressed transmembrane proteins that, in the nervous system, mediate repulsive signals of semaphorins. However, the molecular nature of plexin mediated signal transduction remains poorly understood. Here, we demonstrate that plexin-B family members associate through their C termini with the Rho guanine nucleotide exchange factors PDZ-RhoGEF and LARG. Activation of plexin-B1 by semaphorin 4D regulates PDZ-RhoGEF/LARG activity leading to RhoA activation. In addition, a dominant-negative form of PDZ-RhoGEF blocks semaphorin 4D-induced growth cone collapse in primary hippocampal neurons. Our study indicates that the interaction of mammalian plexin-B family members with the multidomain proteins PDZ-RhoGEF and LARG represents an essential molecular link between plexin-B and localized, Rho-mediated downstream signaling events which underly various plexin mediated cellular phenomena including axonal growth cone collapse. PMID- 12123609 TI - Raf and akt mediate distinct aspects of sensory axon growth. AB - Nerve growth factor (NGF) induces dramatic axon growth from responsive embryonic peripheral neurons. However, the roles of the various NGF-triggered signaling cascades in determining specific axon morphological features remain unknown. Here, we transfected activated and inhibitory mutants of Trk effectors into sensory neurons lacking the proapoptotic protein Bax. This allowed axon growth to be studied in the absence of NGF, enabling us to observe the contributions of individual signaling mediators. While Ras was both necessary and sufficient for NGF-stimulated axon growth, the Ras effectors Raf and Akt induced distinct morphologies. Activated Raf-1 caused axon lengthening comparable to NGF, while active Akt increased axon caliber and branching. Our results suggest that the different Trk effector pathways mediate distinct morphological aspects of developing neurons. PMID- 12123610 TI - Cadherin regulates dendritic spine morphogenesis. AB - Synaptic remodeling has been postulated as a mechanism underlying synaptic plasticity, and cadherin adhesion molecules are thought to be a regulator of such a process. We examined the effects of cadherin blockage on synaptogenesis in cultured hippocampal neurons. This blockade resulted in alterations of dendritic spine morphology, such as filopodia-like elongation of the spine and bifurcation of its head structure, along with concomitant disruption of the distribution of postsynaptic proteins. The accumulation of synapsin at presynaptic sites and synaptic vesicle recycling were also perturbed, although these synaptic responses to the cadherin blockade became less evident upon the maturation of the synapses. These findings suggest that cadherin regulates dendritic spine morphogenesis and related synaptic functions, presumably cooperating with cadherin-independent adhesive mechanisms to maintain spine-axon contacts. PMID- 12123611 TI - Depolarization drives beta-Catenin into neuronal spines promoting changes in synaptic structure and function. AB - Activity-induced changes in adhesion molecules may coordinate presynaptic and postsynaptic plasticity. Here, we demonstrate that beta-catenin, which mediates interactions between cadherins and the actin cytoskeleton, moves from dendritic shafts into spines upon depolarization, increasing its association with cadherins. beta-catenin's redistribution was mimicked or prevented by a tyrosine kinase or phosphatase inhibitor, respectively. Point mutations of beta-catenin's tyrosine 654 altered the shaft/spine distribution: Y654F-beta-catenin-GFP (phosphorylation-prevented) was concentrated in spines, whereas Y654E-beta catenin-GFP (phosphorylation-mimic) accumulated in dendritic shafts. In Y654F expressing neurons, the PSD-95 or associated synapsin-I clusters were larger than those observed in either wild-type-beta-catenin or also Y654E-expressing neurons. Y654F-expressing neurons exhibited a higher minifrequency. Thus, neural activity induces beta-catenin's redistribution into spines, where it interacts with cadherin to influence synaptic size and strength. PMID- 12123612 TI - Ubiquitin and AP180 regulate the abundance of GLR-1 glutamate receptors at postsynaptic elements in C. elegans. AB - Regulated delivery and removal of alpha-amino-3-hydroxy-5-methyl-4 isoxazolepropionic acid (AMPA) glutamate receptors (GluRs) from postsynaptic elements has been proposed as a mechanism for regulating synaptic strength. Here we test the role of ubiquitin in regulating synapses that contain a C. elegans GluR, GLR-1. GLR-1 receptors were ubiquitinated in vivo. Mutations that decreased ubiquitination of GLR-1 increased the abundance of GLR-1 at synapses and altered locomotion behavior in a manner that is consistent with increased synaptic strength. By contrast, overexpression of ubiquitin decreased the abundance of GLR 1 at synapses and decreased the density of GLR-1-containing synapses, and these effects were prevented by mutations in the unc-11 gene, which encodes a clathrin adaptin protein (AP180). These results suggest that ubiquitination of GLR-1 receptors regulates synaptic strength and the formation or stability of GLR-1 containing synapses. PMID- 12123614 TI - Functional expression of AMPA receptors on central terminals of rat dorsal root ganglion neurons and presynaptic inhibition of glutamate release. AB - No direct evidence has been found for expression of functional AMPA receptors by dorsal root ganglion neurons despite immunocytochemical evidence suggesting they are present. Here we report evidence for expression of functional AMPA receptors by a subpopulation of dorsal root ganglion neurons. The AMPA receptors are most prominently located near central terminals of primary afferent fibers. AMPA and kainate receptors were detected by recording receptor-mediated depolarization of the central terminals under selective pharmacological conditions. We demonstrate that activation of presynaptic AMPA receptors by exogenous agonists causes inhibition of glutamate release from the terminals, possibly via primary afferent depolarization (PAD). These results challenge the traditional view that GABA and GABA(A) receptors exclusively mediate PAD, and indicate that PAD is also mediated by glutamate acting on presynaptically localized AMPA and kainate receptors. PMID- 12123613 TI - Abnormal spine morphology and enhanced LTP in LIMK-1 knockout mice. AB - In vitro studies indicate a role for the LIM kinase family in the regulation of cofilin phosphorylation and actin dynamics. In addition, abnormal expression of LIMK-1 is associated with Williams syndrome, a mental disorder with profound deficits in visuospatial cognition. However, the in vivo function of this family of kinases remains elusive. Using LIMK-1 knockout mice, we demonstrate a significant role for LIMK-1 in vivo in regulating cofilin and the actin cytoskeleton. Furthermore, we show that the knockout mice exhibited significant abnormalities in spine morphology and in synaptic function, including enhanced hippocampal long-term potentiation. The knockout mice also showed altered fear responses and spatial learning. These results indicate that LIMK-1 plays a critical role in dendritic spine morphogenesis and brain function. PMID- 12123615 TI - Quantal release of glutamate generates pure kainate and mixed AMPA/kainate EPSCs in hippocampal neurons. AB - The relative contribution of kainate receptors to ongoing glutamatergic activity is at present unknown. We report the presence of spontaneous, miniature, and minimal stimulation-evoked excitatory postsynaptic currents (EPSCs) that are mediated solely by kainate receptors (EPSC(kainate)) or by both AMPA and kainate receptors (EPSC(AMPA/kainate)). EPSC(kainate) and EPSC(AMPA/kainate) are selectively enriched in CA1 interneurons and mossy fibers synapses of CA3 pyramidal neurons, respectively. In CA1 interneurons, the decay time constant of EPSC(kainate) (circa 10 ms) is comparable to values obtained in heterologous expression systems. In both hippocampal neurons, the quantal release of glutamate generates kainate receptor-mediated EPSCs that provide as much as half of the total glutamatergic current. Kainate receptors are, therefore, key players of the ongoing glutamatergic transmission in the hippocampus. PMID- 12123616 TI - Activation of kinetically distinct synaptic conductances on inhibitory interneurons by electrotonically overlapping afferents. AB - Mossy fiber (MF) and CA3 collateral (CL) axons activate common interneurons via synapses comprised of different AMPA receptors to provide feedforward and feedback inhibitory control of the CA3 hippocampal network. Because synapses potentially occur over variable electrotonic distances that distort somatically recorded synaptic currents, it is not known whether the underlying afferent specific synaptic conductances are associated with different time courses. Using a somatic voltage jump technique to alter the driving force at the site of the synapse, we demonstrate that MF and CL synapses overlap in electrotonic location yet differ in conductance time course. Thus, afferent-specific conductance time courses allow single interneurons to differentially integrate feedforward and feedback information without the need to segregate distinct AMPA receptor subunits to different electrotonic domains. PMID- 12123617 TI - Crossmodal processing of object features in human anterior intraparietal cortex: an fMRI study implies equivalencies between humans and monkeys. AB - The organization of macaque posterior parietal cortex (PPC) reflects its functional specialization in integrating polymodal sensory information for object recognition and manipulation. Neuropsychological and recent human imaging studies imply equivalencies between human and macaque PPC, and in particular, the cortex buried in the intraparietal sulcus (IPS). Using functional MRI, we tested the hypothesis that an area in human anterior intraparietal cortex is activated when healthy subjects perform a crossmodal visuo-tactile delayed matching-to-sample task with objects. Tactile or visual object presentation (encoding and recognition) both significantly activated anterior intraparietal cortex. As hypothesized, neural activity in this area was further enhanced when subjects transferred object information between modalities (crossmodal matching). Based on both the observed functional properties and the anatomical location, we suggest that this area in anterior IPS is the human equivalent of macaque area AIP. PMID- 12123618 TI - Tracking the mind's image in the brain I: time-resolved fMRI during visuospatial mental imagery. AB - Mental imagery, the generation and manipulation of mental representations in the absence of sensory stimulation, is a core element of numerous cognitive processes. We investigate the cortical mechanisms underlying imagery and spatial analysis in the visual domain using event-related functional magnetic resonance imaging during the mental clock task. The time-resolved analysis of cortical activation from auditory perception to motor response reveals a sequential activation of the left and right posterior parietal cortex, suggesting that these regions perform distinct functions in this imagery task. This is confirmed by a trial-by-trial analysis of correlations between reaction time and onset, width, and amplitude of the hemodynamic response. These findings pose neurophysiological constraints on cognitive models of mental imagery. PMID- 12123619 TI - Tracking the mind's image in the brain II: transcranial magnetic stimulation reveals parietal asymmetry in visuospatial imagery. AB - The functional relevance of brain activity during visuospatial tasks was investigated by combining functional magnetic resonance imaging with unilateral repetitive transcranial magnetic stimulation (rTMS). The cognitive tasks involved visuospatial operations on visually presented and mentally imagined material ("mental clock task"). While visuospatial operations were associated with activation of the intraparietal sulcus region bilaterally, only the group which received rTMS to the right parietal lobe showed an impairment of performance during and immediately after rTMS. This functional parietal asymmetry might indicate a capacity of the right parietal lobe to compensate for a temporary suppression of the left. This is compatible with current theories of spatial hemineglect and constitutes a constraint for models of distributed information processing in the parietal lobes. PMID- 12123620 TI - Practice with sleep makes perfect: sleep-dependent motor skill learning. AB - Improvement in motor skill performance is known to continue for at least 24 hr following training, yet the relative contributions of time spent awake and asleep are unknown. Here we provide evidence that a night of sleep results in a 20% increase in motor speed without loss of accuracy, while an equivalent period of time during wake provides no significant benefit. Furthermore, a significant correlation exists between the improved performance overnight and the amount of stage 2 NREM sleep, particularly late in the night. This finding of sleep dependent motor skill improvement may have important implications for the efficient learning of all skilled actions in humans. PMID- 12123623 TI - Molecular mechanisms of immunomodulatory activity of glucocorticoids. AB - The development and function of cells in the immune system are regulated by many intrinsic and extrinsic factors. One class of molecule that affects immune cells belongs to the neuroendocrine system and the best-studied mediators in this category are glucocorticoids. These are small lypophilic molecules that participate in a wide number of normal and pathologic processes. This paper concentrates on their physiologic and pharmacologic effects on the immune response. PMID- 12123624 TI - Exploring the variability in antibiotic prescribing profiles among paediatricians from two different areas of Italy. AB - We carried out a multicentre community-based study in order to describe the antibiotic therapeutic approach of paediatricians from two different areas of Italy in the treatment of respiratory tract infection (RTIs), and to assess which factors are involved in a possible variability of prescribing habits. Forty paediatricians participated in the study between October 1998 and April 1999. They had to complete a questionnaire for each therapeutic intervention resulting in an antibiotic prescription. A logistic regression model was used to identify possible predictors in choosing parenteral antibiotics for the treatment of RTIs. In 2 975 questionnaires of antibiotic treatment, RTIs represented 90.2% of the total antibiotics used. Upper respiratory tract infections were the most commonly treated diagnostic group (59.6%), followed by lower respiratory tract infections (20.4%), and middle ear infections (19.8%). Statistically significant differences between northern and southern Italy were reported in the antibiotic prescription profile and the duration of the therapy. Another marked difference was reported in the frequency of laboratory analysis requests. The logistic regression model indicated that the use of parenteral antibiotics appears significantly related to the type of infections [lower RTIs: (OR: 3.99; 95% CI: 2.49-6.37)], the geographic location [northern Italy: (OR: 0.20; 95% CI: 0.20-0.39)], and the presence of concurrent diseases (OR: 3.21; 95% CI: 1.46-7.02). The lack of adherence to clinical guidelines and the marked variability of antibiotic prescription rates between different areas of the country appear to be related to factors other than bacterial resistance, and highlight the importance of carrying out educational programmes targeted at the national level for improving the antibiotic prescription habits for the treatment of RTIs. PMID- 12123625 TI - Role of endogenous reactive oxygen derived species and cyclooxygenase mediators in 5-hydroxytryptamine-induced contractions in rat aorta: relationship to nitric oxide. AB - Endogenous reactive oxygen species (superoxide anion, hydroxyl radical and hydrogen peroxide), endothelium-derived nitric oxide and cyclooxygenase mediators are involved in the regulation of vascular smooth muscle tone. An imbalance of these mediators can have profound implications in various cardiovascular disorders. Involvement of endogenous reactive oxygen species, endothelium-derived nitric oxide (NO) and cyclooxygenase mediators in 5-hydroxytryptamine- (5-HT-) induced contractions of endothelium intact rat aortic rings have been investigated in the present study. The contribution of each of the endogenous reactive oxygen species in mediating 5-HT-induced contractions was studied by pretreating the rings with their respective scavengers. Pretreatment of the rings with superoxide dismutase (superoxide radical scavenger), catalase (H (2)O (2)inactivator), mannitol (extracellular OH. scavenger), or thiourea (intracellular OH. radical scavenger) significantly depressed the 5-HT-induced contractions in the aortic rings. The responses to 5-HT in the presence of SOD or catalase were augmented byL -NAME pretreatment. Though aminotriazole partially inhibited the catalase activity, it inhibited 5-HT-induced contractions significantly. The results obtained thus suggest that endogenous generation of ROS (O(2).(-), H (2)O (2)and OH.) modulates 5-HT-induced rat aortic ring contractions. In addition, H (2)O (2)generated in the endothelium seems to regulate the vascular response and also act as a mediator to release other vasoactive substances. Basal production of NO by the endothelium seems to affect the vascular response due to its interaction with ROS mediators. PMID- 12123626 TI - Ischaemic preconditioning and mast cell histamine release: microdialysis of isolated rat hearts. AB - The aim of this study was to investigate the feasibility of intramyocardium kinetics of histamine release by microdialysis in the isolated rat heart and ascertain if the inhibition of histamine release is implicated in the antiarrhythmic effect of preconditioning. A 30 min normothermic global ischaemia model followed by 30 min reperfusion was carried out in the control group (n= 9). In the preconditioning group (n= 8) there was a 5 min global ischaemia followed by 10 min of reperfusion. A mast cell stabilizing group received the disodium cromoglycate ( 10 micro M, n= 10). The last group received a mast cell degranulator, compound 48/80 (1micro g ml (-1), n= 10). In the control group, the histamine release during reperfusion was significantly different from the basal concentration ( 18.4 +/- 6.5 vs 1.9 +/- 0.5 nM, P< 0.05) and was associated with a maximal period of severe arrhythmias. The ischaemic preconditioning modified the histamine release kinetics with an early mast cell degranulation ( 9.7 +/- 1.5 nM) and a significant decrease in the total period of severe arrhythmias in comparison with the control group ( P< 0.05). In the disodium cromoglycate group, the histamine release during reperfusion decreased ( 3.1 +/- 0.7 nM) and was associated with a maximal period of severe arrhythmias. In the C48/80 group, the increase in the histamine released during reperfusion ( 21.2 +/- 5.0 nM) was associated with a maximal period of severe arrhythmias. These results showed firstly the feasibility of kinetic histamine release in myocardium interstitial fluid on the isolated rat heart and secondly that the inhibition of histamine release did not play a direct role in the antiarrhythmic effect of preconditioning. PMID- 12123628 TI - Comparison of two compartmental models for describing ranitidine's plasmatic profiles. AB - The plasmatic profiles of 12 healthy volunteers after oral administration of ranitidine (150 mg) were studied considering two compartmental models. We observed the presence of two peaks. The proposed mechanism responsible for the existence of secondary peaks includes enterohepatic recirculation and the existence of multiple sites of absorption along the gastrointestinal tract. For characterizing the pharmacokinetic aspect of the drug, both phenomena were described using two compartmental models. We calculated the pharmacokinetic parameters and statistical tests after fitting the data of each volunteer under both models proposed. Statistically significant differences were not found in the statistical test values but existed in the area under the curve (AUC) comparing between models. To decide which of the two proposed models gave the best approximation of the physiological phenomenon undergone, we studied the pharmacokinetic of the drug in the rat, an animal without gallbladder. After oral administration of ranitidine, the plasmatic profile of the animals showed at least two peaks. Less than 0.2% of an oral dose was recovered in bile as ranitidine. Therefore, and considering the rat has no post-absorptive depot from where the drug can be released discontinuously, enterohepatic recycling does not seem to contribute significantly to the occurrence of secondary peaks in the concentration-time profiles of rats. Considering the results, we proposed that the best model able to explain the plasmatic profiles found in man and rats after oral administration of ranitidine is the one that presents multiple sites of absorption along the gastrointestinal tract. It is important to define the correct model in the calculation of the AUC and so in the value of the absolute bioavailability. PMID- 12123627 TI - Effect of age receptor blocker and/or anti-inflammatory coadministration in relation to glycation, oxidative stress and cytokine production in stz diabetic rats. AB - Until now the relation between advanced glycation end products (AGEs) and vascular lesion is still controversial. However, the interaction of the former with a receptor triggers the synthesis of cytokines particularly interleukin 1- beta(IL-1 beta) and tumour necrosis factor- alpha(TNFalpha ). Subsequent release of nitric oxide (NO) may in turn induce certain damage to beta cell islets. Several arguments indicated that AGEs and reactive oxygen intermediates (ROIs) could alter the function of the vessel wall. Therefore, this study was undertaken to investigate the effectiveness of aminoguanidine, AG (inhibitor of AGE formation) joined with omega -3-fatty acids, omega 3FAs (anti-inflammatory immunosuppressive drug) in STZ diabetic rats. Diabetes was induced in 48 female albino Wistar rats by a single intraperitoneal (i.p.) injection of streptozotocin (STZ, 50 mg kg (-1)). Diabetic animals were treated with AG (50 mg kg(-1) ) and/or omega 3FAs (12 mg kg (-1)) daily and orally for 4 weeks. Groups of age matched diabetic rats ( n= 10) and healthy animals ( n= 10) served as positive and negative controls. At the end of the study, plasma glucose, fructosamine, total cholesterol (TC), high density lipoprotein cholesterol (HDLC), low density lipoprotein cholesterol (LDLC), the susceptibility of LDL to copper-catalysed oxidation, catalase activity, NO, IL-1 beta, TNF alpha were measured. Histopathological assessment of pancreatic slices were also determined. Diabetes remarkably increased plasma glucose, fructosamine and dyslipidaemia (increased TC, LDLC and decreased HDLC). Oxidative markers like oxidative susceptibility of LDL, catalase activity and NO levels were greatly enhanced. Finally, it increased the synthesis and release of cytokine (IL-1beta and TNF alpha). Treatment of diabetic rats with AG and omega 3FAs markedly reduced the above mentioned parameters. Combined form therapy has a better effect regarding oxidative cell markers, specifically NO level. Finally, omega 3FAs coadministration with AG nearly restored the atrophy of islets of Langerhan's and the peripheral lymphocytic infiltration compared to diabetic and AG treated groups. In conclusion, there is a direct correlation between glycation, oxidative stress and cytokine production with increased propensity of microvascular disorder in STZ diabetic rats. omega 3FA administration with AGE receptor blocker may represent a possible avenue of research for therapeutics directed for alleviating the complication associated with diabetes. PMID- 12123630 TI - Selective angiotensin II type 1 receptor blockade ameliorates cyclosporine nephrotoxicity. AB - Nephrotoxicity associated with cyclosporine A (CsA) administration is characterized by marked renal vasoconstriction, interstitial fibrosis and arteriolar hypertrophy. The molecular mechanisms of CsA nephrotoxicity are not well characterized, but previous studies have demonstrated that angiotensin II (Ang II), the primary mediator of renin-angiotensin system (RAS) cascade plays a role in its pathogenesis. Recent studies also suggest an involvement of reactive oxygen species (ROS) in CsA nephrotoxicity. There is emerging evidence that Ang II induces oxidative stress in vitro and in vivo. The aims of this study were to investigate the role of Ang II-induced oxidative stress in CsA nephrotoxicity, and to examine the effects of the insurmountable Ang II type 1 (AT (1)) receptor antagonist, candesartan on CsA-induced nephrotoxicity in rats. Candesartan cilexetil (1.0 mg kg (-1), perorally (p.o.), once a day) was administered 24 h before and 21 days concurrently with CsA (20 mg kg(-1), subcutaneously (s.c.)). Tissue lipid peroxidation was measured as thiobarbituric acid reacting substances (TBARS). Renal function was assessed by estimating serum creatinine, blood urea nitrogen (BUN), creatinine and urea clearance. Renal morphological alterations were assessed by histopathological examination of Haematoxylin-Eosin, PAS and Mason's trichome stained sections of the kidneys. CsA (20 mg kg (-1), s.c.) administration for 21 days produced elevated levels of TBARS and deteriorated the renal function as assessed by increased serum creatinine, BUN and decreased creatinine and urea clearance as compared to vehicle treated rats. The kidneys of CsA-treated rats showed severe striped interstitial fibrosis, arteriolopathy, glomerular basement thickening, tubular vacuolisation and hyaline casts. Candesartan cilexetil (1.0 mg kg (-1)) markedly reduced elevated levels of TBARS, significantly attenuated renal dysfunction and morphological changes in CsA treated rats. These results clearly demonstrate the pivotal role of Ang II induced oxidative stress and the therapeutic potential of AT (1)receptor antagonists in ameliorating CsA-induced nephrotoxicity. PMID- 12123629 TI - Noradrenaline release in rat locus coeruleus is regulated by both opioid and alpha(2) -adrenoceptors. AB - Extracellular fluid levels of noradrenaline (NA) in the locus coeruleus (LC) during naloxone-precipitated morphine withdrawal with pretreatment of yohimbine (1 mg kg (-1), s.c.) or clonidine (1 mg kg (-1), s.c.) were measured in rats. There was a significant increase in the NA level after the injection of naloxone (2 mg kg (-1), i.p.) in the morphine-dependent rats. Moreover, the NA levels in the LC markedly increased during the 30-60 min following the naloxone (i.p.) challenge in the morphine-dependent rats pretreated with yohimbine. In contrast, the naloxone challenge in morphine-dependent rats pretreated with clonidine notably decreased the levels of NA in the LC. Behavioral signs of withdrawal were observed following the naloxone challenge in the morphine-dependent rats pretreated with yohimbine, with minimal signs in the morphine-infused rats pretreated with clonidine, and none in the saline-infused controls. These results directly suggest that NA increased within the LC after the naloxone challenge in morphine-dependent animals pretreated with yohimbine may be, at least in part, regulated by alpha(2) -adrenoceptors in the LC. PMID- 12123631 TI - Mechanism of the beneficial effects of dantrolene sodium on ethanol-induced acute gastric mucosal injury in rats. AB - In our study, we examined anti-ulcerogen and antioxidant effects of dantrolene sodium on ethanol-induced gastric lesions in rats. Dantrolene sodium was administered intraperitoneally (i.p.) in several doses, and famotidine was used at a dose of 20 mg kg (-1). It was found that pretreatment with dantrolene sodium at doses of 1, 5 and 10 mg kg(-1) significantly reduced ethanol-induced gastric damage and malondialdehyde levels, and significantly increased antioxidant enzymes superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) activities. We conclude that dantrolene sodium clearly has antioxidant properties and that the protective effect of dantrolene sodium against ethanol-induced gastric mucosal lesion, at least in part, depends upon the reduction in the lipid peroxidation and an increase in the activity of antioxidant enzymes SOD and GSH Px. PMID- 12123632 TI - Characterization of CD95 ligand (CD95L)-induced apoptosis in human tenon fibroblasts. AB - Toxic side effects of cytotoxic agents such as 5-fluorouracil or mitomycin-C in glaucomatous filtering procedures call for alternative approaches to control fibroblast proliferation. CD95L is a death ligand that triggers apoptosis in susceptible target cells. Apoptosis allows for the safe disposal of cells without damaging the surrounding tissue. The goal of this study was to characterize and to evaluate the CD95L induced cell death in cultured Tenon fibroblasts. Human Tenon fibroblasts were treated with different concentrations of CD95L. For comparison, murine NIH 3T3 fibroblasts were used. Immunohistochemistry and Western blot were used to investigate the CD95 and CD95L expression. Cytotoxicity was measured by crystal violet assay. Apoptosis was investigated using in situ DNA end labelling (TUNEL). DEVD-AMC caspase 3 like activity was measured and caspase 3 processing was studied by immunoblot and the use of the caspase inhibitor DEVD-CHO in cell culture assays. Tenon and NIH 3T3 fibroblasts express CD95 and CD95L. The authors found concentration dependent inhibition of proliferation after CD95L treatment. Tenon fibroblasts, but not NIH 3T3 fibroblasts, show synergy when combined with actinomycin D or cyclohexamide. CD95L treatment did not alter total protein or RNA synthesis. Cell death induced by CD95L was apoptotic and activated caspase 3, as TUNEL positive cells and the active fragment of caspase 3 were found. CD95L induced cell death could be inhibited by the caspase-inhibitor.Here, it is demonstrated that the CD95L induced cell death in cultured human Tenon fibroblasts is apoptotic and possibly mediated by the caspase 3 pathway. These results suggest that it may be possible to use CD95L in glaucomatous filtering procedures. In vivo studies are necessary for further evaluation. PMID- 12123633 TI - Molecular profiling and cellular localization of connexin isoforms in the rat ciliary epithelium. AB - The functionally distinct epithelial layers of the ciliary body act as a syncitium to produce the aqueous humour. Ultrastructural studies have shown that the pigmented (PE) and non-pigmented (NPE) cell layers of the ciliary epithelium are connected by gap junctions. However the molecular composition of gap junctions both between and within the two cell layers has not been comprehensively studied. To address this issue the authors have performed an extensive molecular screening of connexin (Cx) expression patterns in ciliary epithelium of the rat. Initially, mRNA was extracted from rat ciliary bodies, reverse-transcribed, and subjected to two rounds of PCR using primer sets designed against each of the 14 Cx isoforms known to be expressed in the rat. This initial screening protocol amplified eight candidate Cx isoforms (Cxs 26, 31, 33, 37, 40, 43, 45 and 46). The Cx isoforms identified in this initial screen were then first assigned to the ciliary epithelium itself (Cxs 26, 31, 40 and 43) or structures outside the epithelium (Cxs 37, 40, and 45) using immunohistochemistry performed on ciliary body whole mounts. No convincing evidence for either Cx 33 or 46 labelling was found in the ciliary body. Then the four Cx isoforms localized to the epithelium were further localized to specific membrane domains within the epithelial cell layers by performing high resolution imaging of the antibody labeling patterns obtained in cryosections. This enabled Cx26 and 31 to be specifically localized to spatially different gap junctions between NPE cells. Cx31 labeled gap junctions associated with an extensive network of membrane interdigitations found between NPE cells at their basal surfaces. In contrast Cx26 labeling in NPE cells was restricted to the basolateral membranes of adjacent NPE cells. Cx40 and Cx43 were both localized to the PE-NPE interface where they formed discrete homomeric/homotypic gap junction plaques. No convincing evidence was found for antibody labeling between PE cells. Thus it appears that intercellular communication, both within the NPE layer and between the PE and NPE cell layers, is mediated by gap junction channels that have distinctive permeability properties. In particular the results raise the possibility that the permeability of PE-NPE gap junctions can be modulated by changing the Cx43 : Cx40 expression ratio. Whether such a change in Cx expression ratios occurs and what effect it has on aqueous humour production and composition remains to be determined. PMID- 12123634 TI - Subretinal transplantation of brain-derived precursor cells to young RCS rats promotes photoreceptor cell survival. AB - The potential use of in vitro-expanded precursor cells or cell lines in brain repair includes transplantation of such cells for cell replacement purposes and the activation of host cells to provide 'self-repair'. Recently, it has been reported that the immortalized brain-derived cell line RN33B (derived from the embryonic rat medullary raphe) survive, integrate and differentiate after subretinal grafting to normal adult rats. Here, it is demonstrated that grafts of these cells survive for at least 6 weeks after implantation into postnatal days 21 and 35 retinas of normal and Royal College of Surgeons rats, a model of retinal degeneration. Implanted cells integrate into the retinal pigment epithelium and the inner retinal layers, and the anterior part of the optic nerve of both normal and Royal College of Surgeons rats. The RN33B cells migrate within the retina, occupying the whole retina from one eccentricity to the other. A significant number of the grafted cells differentiate into glial cells, as shown by the double labelling of the reporter genes LacZ or green fluorescent protein, with several glial markers, including oligodendrocytic markers. Many implanted cells in the host retina were in a proliferative stage judging from proliferative cell nuclear antigen and SV40 large T-antigen immunohistochemistry. Interestingly, there was a promotion of photoreceptor survival, extending over more than 2/3 of the superior hemisphere, in Royal College of Surgeons rats transplanted at postnatal day 21, but not at postnatal day 35. In addition, grafted cells were found in the surviving photoreceptor layer in these rats. PMID- 12123635 TI - Recombinant Sendai virus-mediated gene transfer into adult rat retinal tissue: efficient gene transfer by brief exposure. AB - To determine the usefulness of recombinant Sendai virus (SeV) for ocular gene transfer, the authors characterized SeV-mediated gene transfer to the retinal tissue of adult rats via subretinal injection. Recombinant SeV encoding the lacZ gene achieved frequent transgene expression in the retinal pigment epithelium (RPE) (mean=38.76%), while gene transfer to other retinal cells was rare. These findings are similar to those of previous reports using adenoviruses. Peak reporter gene expression of SeV in cultured RPE cells was similar to that of adenovirus at the same titer; however, SeV achieved high levels of expression after a brief vector-cell contact time, while adenovirus required over 3hr for efficient gene transfer. This finding was also observed in vivo following a brief SeV filling in the subretinal space, and may therefore provide a clinical advantage in avoiding retinal damage due to prolonged detachment. The observed SeV-mediated gene expression in the rat retina was transient. The initial phase of the decrease in luciferase activity could be prevented by daily eye drops of dexamethasone, suggesting that the corticosteroid-sensitive host reaction may affect early clearance of the virus. The late decline of transgene expression (2 weeks) was inhibited by the immunosuppressant, cyclosporin A, in a dose-dependent manner, suggesting that the cytotoxic T-lymphocyte response may be important in this phase. This work represents the first report of SeV-mediated gene transfer to ocular tissue, and identifies recombinant SeV as a new tool for studies of retinal gene transfer and gene therapy. PMID- 12123638 TI - Lipid hydroperoxide stimulates leukocyte-endothelium interaction in the retinal microcirculation. AB - Leukocyte dynamics were evaluatyed in vivo in rat retinal microcirculation following exposure to lipid hydroperoxide (LHP) in the vitreous.Various amounts (1, 5, 10 or 100 microg) of LHP (18:2) dissolved in 5 microl of sodium borate buffer (SBB, 0.02M) were injected into the vitreous of Brown-Norway rats. As a comparative study, 10 microg of linoleic acid (LA) dissolved in 5 microl of SBB was injected in the same way. Rats that did not undergo injection were evaluated as un-treated. At 2 to 48 hr after LHP exposure, the following were examined: (1) the flux of rolling leukocytes along the major retinal veins, (2) the number of leukocytes that accumulated in the retinal microvasculature using acridine orange digital fluorography and (3) the diameter of major retinal vessels. In the LHP treated eyes, leukocyte rolling along the major retinal veins was observed and the number increased in a dose-dependent manner ( 1 to 10 microg). The flux of rolling leukocytes peaked at 6 hr after LHP (10-100 microg) injection. No rolling leukocytes were observed in LA-treated or un-treated eyes. The number of accumulated leukocytes started to increase at 4 hr and peaked at 24 hr after LHP (10 microg) injection. This number was significantly higher than that in LA treated and un-treated eyes. Venous dilation was seen from 4 hr after LHP (10 microg) injection and became significant at 6 and 24 hr as compared with LA treated and un-treated eyes. The results indicate that increased LHP levels in the vitreous due to oxidative stress enhance leukocyte-enothelium interaction in the retinal microcirculation. PMID- 12123637 TI - Differential effects of transforming growth factor-beta2 on corneal endothelial cell proliferation-A role of serum factors. AB - PURPOSE: To determine the influence of serum on transforming growth factor (TGF) beta2 mediated effects on the proliferation of corneal endothelial cells (CE). METHODS: Rat CE were grown in explant culture and the proliferation of CE was measured by [(3)H]thymidine bioassay. Subconfluent cells were synchronized in the G0 (quiescent) phase of the cell cycle by serum starvation for 24hr. Serum and [(3)H]thymidine were then added to the cells in the presence or absence of a physiological concentration (5ngml(-1)) of exogenous active TGF-beta2. Radioactivity was measured at various time points to detect DNA synthesis. These experiments were repeated without adding serum after serum starvation. Preincubation of exogenous TGF-beta2 with neutralizing antibody was used to test the cytokine specificity. RESULTS: Without TGF-beta2, a linear increase in [(3)H]thymidine incorporation, indicating S-phase, began approximately 16hr after serum addition, then plateaued at approximately 24hr. Serum promoted DNA synthesis of CE in a dose-dependent manner at concentrations of 0.5-10%. In cultures with 10% serum, TGF-beta2 (0.5, 1, 5, and 20ngml(-1)) suppressed CE growth dose-dependently. The growth amplitude decreased and the time before S phase entry, G1 phase, was prolonged to 24hr. In culture with 1% serum, TGF-beta2 (5ngml(-1)) suppressed the CE proliferation by delaying S-phase entry without suppressing growth amplitude. In cultures without serum, TGF-beta2 promoted CE growth to a level similar to that of cultures supplemented with 0.5% serum. CONCLUSIONS: Responses of cultured CE to exogenous TGF-beta2 depended on the concentration of serum in the medium. This result implies a possibility that in vivo serum influx through a compromised blood-ocular barrier could influence the CE growth by changing the responses of these cells to TGF-beta2 in aqueous humor. PMID- 12123639 TI - Ca(2+) regulation in differentiating lens cells in culture. AB - In the ocular lens, cataract formation is associated with an elevated intracellular Ca(2+) concentration (Ca(2+)(i)) resulting from the loss of lens cell Ca(2+) regulation. The mechanisms regulating Ca(2+)(i) have been characterized previously in lens epithelial cells, but have not been well characterized in the more differentiated lens fiber cells. The mechanisms regulating Ca(2+)(i) in clusters of fiber-like cells (lentoids) in a sheep lens primary cell culture system in which the epithelial cells differentiate into enlarged fiber-like cells were investigated. Only approximately 50% of the lentoids responded to thapsigargin and/or agonists (ATP and epinephrine), compared to>95% of the epithelial cells. Remarkably, most (90%) lentoids exhibited a resting cytosolic Ca(2+)(i) that was approximately three-fold greater than that in epithelial cells (approximately 100n M). This elevated resting cytosolic Ca(2+)(i) was not affected by thapsigargin treatment, but decreased upon removal of extracellular Ca(2+) or addition of the Ca(2+) channel blocker Gd(3+) (5mM ). These results suggest that a plasma membrane Ca(2+) channel is more active in lentoids than in epithelial cells. Indeed, when plasma membrane cation channel activity was monitored by Mn(2+) influx and quenching of fura-2 fluorescence, quenching was faster in lentoids than epithelial cells. Following thapsigargin treatment, capacitative Ca(2+) entry was activated in epithelial cells but not lentoids. In conclusion, during differentiation in primary cell culture, lens cells lose their ability to respond to agonists and exhibit an elevated resting Ca(2+)(i) that was dependent on the activation of a Ca(2+) influx pathway. The results of this study support the possibility that a sustained elevation in resting Ca(2+)(i) is one of the factors controlling lens cell differentiation, possibly by triggering events such as calpain activation. PMID- 12123636 TI - Innervation of the uvea by galanin and somatostatin immunoreactive axons in macaques and baboons. AB - The neuropeptide galanin has not been localized previously in the primate uvea, and the neuropeptide somatostatin has not been localized in the uvea of any mammal. Here, the distribution of galanin-like and somatostatin-like immunoreactive axons in the iris, ciliary body and choroid of macaques and baboons using double and triple immunofluorescence labeling techniques and confocal microscopy was reported. In the ciliary body, galanin-like immunoreactive axons innervated blood vessels and the ciliary processes, particularly at their bases. In the iris, the majority of these axons was associated with the loose connective tissue in the stroma. Somatostatin-like immunoreactive axons were found in many of the same areas of the uvea supplied by cholinergic nerves. In the ciliary body, there were labelled axons within the ciliary processes and ciliary muscle. They were also found alongside blood vessels in the ciliary stroma. In the iris, somatostatin-like immunoreactive axons were abundant in the sphincter muscle and less so in the dilator muscle. A unilateral sympathectomy had no effect on the distribution of somatostatin-like or galanin-like immunoreactive axons, and these axons did not contain the sympathetic marker tyrosine hydroxylase. They did not contain the parasympathetic marker choline acetyltransferase, either. The galanin-like immunoreactive axons contained other neuropeptides found in sensory nerves, including calcitonin gene related peptide, substance P and cholecystokinin. Somatostatin-like immunoreactive axons did not contain any of these sensory neuropeptides or galanin-like immunoreactivity, and they were neither labelled with an antibody to 200kDa neurofilament protein, nor did they bind isolectin-IB(4). Nevertheless, they are likely to be of sensory origin because somatostatin-like immunoreactive perikarya have previously been localized in the trigeminal ganglion of primates. Taken together, these findings indicate galanin and somatostatin are present in two different subsets of sensory axons in primate uvea. PMID- 12123640 TI - Characterization of Type I and Type II myristoyl-CoA:protein N myristoyltransferases with the Acyl-CoAs found on heterogeneously acylated retinal proteins. AB - Protein myristoylation occurs when the 14 carbon fatty acid, myristic acid, is covalently attached by amide linkage to a protein's N -terminal glycine by an N terminal myristoyltransferase (NMT). A variation of this called heterogeneous acylation occurs in vivo only in retina when specific proteins are modified by myristic acid (14:0), tetradecenoic acid (14:1 n-9), tetradecadienoic acid (14:2n -6), and lauric acid (12:0). Myristic and lauric acids are relatively rare, comprising approximately 1% of the fatty acids in the retina. The unsaturated fatty acids 14:1 n-9 and 14:2 n-6 are less abundant, but can be synthesized in retina by retroconversion of 18:1 n-9 and 18:2 n-6 fatty acids, respectively. A previous quantitative study of acyl-CoA pools in bovine retina, heart, and liver found comparable levels of acyl-CoAs in each tissue, indicating that heterogeneous acylation is not due to limiting amounts of myristoyl-CoA in retina. In this current study the authors have characterized a panel of purified recombinant Type I and II NMTs found in retina and liver by assessing their utilization of the four acyl-CoAs used in vivo to acylate retina proteins. Acceptor peptides used in these assays were derived from the N -termini of src which is only myristoylated in vivo, and the cAMP dependent kinase A catalytic subunit which is heterogeneously acylated in retina, but myristoylated in other tissues. The authors have tested the ability of unlabelled acyl-CoAs to compete with [(3)H] myristoyl-CoA transfer, the efficacy of an NMT inhibitory protein (NIP(71)), and acyl-CoA affinity chromatography was used to isolate endogenous NMT inhibitory factor(s) from bovine heart and retina tissue homogenates. These results provide a basis of kinetic parameters and enzymatic characterization for Type I and Type II NMTs with two acceptor peptides and the four physiologically relevant fatty acid-CoAs found on retinal proteins, but do not indicate that heterogeneous acylation is a specialized function of any of the enzymes tested in this study. PMID- 12123641 TI - Phosphatidylglycerol potently protects human retinal pigment epithelial cells against apoptosis induced by A2E, a compound suspected to cause age-related macula degeneration. AB - Age-related macular degeneration (AMD) affects about one fifth of the population older than 65 years and is one of the main causes of poor vision in the elderly in industrialized nations. The endogenous lipophilic and cationic compound N retinyl-N-retinylidene ethanolamine (A2E) is suspected to cause the dry form of the disease, which currently cannot be treated. The authors recently reported that A2E induces apoptosis in several cell types including porcine retinal pigment epithelial cells, detaches pro-apoptotic proteins from mitochondria, and inhibits cytochrome c oxidase. A2E acts primarily at the level of cardiolipin/cytochrome c oxidase, which in the light becomes permanently inactivated by A2E. The authors now report that A2E at low concentrations causes apoptosis in cultured human retinal pigment epithelial cells. These cells are more sensitive to A2E in the light than in the dark. Phosphatidylglycerol, a negatively charged phospholipid and immediate biosynthetic precursor of cardiolipin readily inhibits apoptosis. Exposure of cells to A2E results in the formation of reactive oxygen and nitrogen species, and exposure of mitochondria to A2E results in oxidative stress. Accordingly, the potent antioxidant coenzyme Q also protects cells against A2E-induced apoptosis. These findings are highly relevant for the treatment and/or prevention of AMD. PMID- 12123642 TI - Sex-linked differences in cataract progression in Emory mice. PMID- 12123643 TI - Aluminum: impacts and disease. AB - Aluminum is the most widely distributed metal in the environment and is extensively used in modern daily life. Aluminum enters into the body from the environment and from diet and medication. However, there is no known physiological role for aluminum within the body and hence this metal may produce adverse physiological effects. The impact of aluminum on neural tissues is well reported but studies on extraneural tissues are not well summarized. In this review, the impacts of aluminum on humans and its impact on major physiological systems are summarized and discussed. The neuropathologies associated with high brain aluminum levels, including structural, biochemical, and neurobehavioral changes, have been summarized. In addition, the impact of aluminum on the musculoskeletal system, respiratory system, cardiovascular system, hepatobiliary system, endocrine system, urinary system, and reproductive system are discussed. PMID- 12123644 TI - Environmental exposure to trace elements and risk of amyotrophic lateral sclerosis: a population-based case-control study. AB - We analyzed the association between the environmental exposure to trace elements and the risk of sporadic amyotrophic lateral sclerosis (ALS) in a population based case-control study in the Emilia-Romagna region in northern Italy. We evaluated exposure to selected trace elements by measuring toenail concentrations of the same by means of inductively coupled plasma optical spectrometry and instrumental neutron activation analysis. The final number enrolled in the study was 22 patients and 40 controls. Disease progression, assessed through a clinical score, was generally unassociated with toenail trace element levels, with the exception of an inverse relation with zinc and selenium content and a direct correlation with copper concentration. In logistic regression analysis, we found no evidence of an association between ALS risk and toenail content of cadmium, lead, copper, zinc, manganese, selenium, chromium, cobalt, iron, and aluminum. This investigation does not suggest a major role in sporadic ALS etiology of environmental exposure to these trace elements, though results for zinc, selenium, and copper should be evaluated with caution due to the potential limitations of toenails as biomarkers of chronic exposure in patients. PMID- 12123645 TI - Chlorination byproducts and nitrate in drinking water and risk for congenital cardiac defects. AB - Drinking water disinfection byproducts have been associated with an increased risk for congenital defects including cardiac defects. Using Swedish health registers linked to information on municipal drinking water composition, individual data on drinking water characteristics were obtained for 58,669 women. Among the infants born, 753 had a cardiac defect. The risk for a cardiac defect was determined for ground water versus surface water, for different chlorination procedures, and for trihalomethane and nitrate concentrations. Ground water was associated with an increased risk for cardiac defect when crude rates were analyzed but after suitable adjustments this excess rate was found to be determined by chlorination procedures including chlorine dioxide. Chlorine dioxide appears itself as an independent risk factor for cardiac defects (adjusted odds ratio 1.61 (95%CI 1.00-2.59)). The risk for cardiac defects increased with increasing trihalomethane concentrations (P=0.0005). There was an indicated but statistically nonsignificant excess risk associated with nitrate concentration. The individual risk for congenital cardiac defect caused by chlorine dioxide and trihalomethanes is small but as a large population is exposed to public drinking water, the attributable risk for cardiac defects may not be negligible. PMID- 12123646 TI - The effects of humic acid-arsenate complexes on human red blood cells. AB - Humic acid (HA) has been proposed as factor in the cause of Blackfoot disease (BFD) among individuals who live along the southwest coast of Taiwan. In this study, the interaction of the synthetic humic acid, made from catechol, with sodium arsenate (As(V)) was investigated and assessed with respect to damage to human red blood cells. HA is characterized as phenolic and phenolic carboxylic polymer structures containing both -COOH and -OH as their main functional groups. HA and As(V) alone are able to hemolyze 60-100 and 5-20% human red blood cells at concentrations of 50-300 microg/ml and 5-100 mM, respectively, after 6 h. HA is shown to be relatively ineffective in causing ATP depletion of red blood cells. For organometallic complexes composed of HA-As(V) the inhibition effect of EDTA was completely abolished and the use of the triple complex HA-As(V)-EDTA resulted in an enhancement of hemolysis. HA caused lipid peroxidation in a concentration- and time-dependent manner. However, HA-As(V) and As(V) decreased lipid peroxidation. These results indicated that HA initiates oxidative stress on red blood cells and this results in their dysfunction. HA-chelated high-concentration metal complexes inhibited the structures containing the main functional groups involved in decreasing hemolysis, and, thus, HA may be a significant factor in the etiology of BFD. PMID- 12123647 TI - Cytotoxicity of heavy metals on primary cultured alveolar type II cells. AB - The lung is the primary target organ of airborne heavy metal-induced toxicity. The aims of this study were to investigate differential acute lung cytotoxicity caused by heavy metals using a primary culture of alveolar type II cells and to establish an in vitro assessment model of lung toxicity. The cytotoxicity of heavy metals was determined by measuring the lactate dehydrogenase release and (51)chromium release from lyzed cells. With respect to the LC(50) values, drug concentrations causing a 50% loss in cell viability, the mean value of Hg was 110 microM and that of Cd was 220 to 250 microM. Cytotoxicity was graded high for Hg and Cd, moderate for Pb and Ni, and negligible for Mn. Additional morphological observations of cell membrane integrity by scanning electron microscopy were compatible with the results of biochemical measurements. In conclusion, we have presented an in vitro assessment model of lung toxicity, which can be used effectively to assess the differential effects of heavy metals on alveolar type II cells. The findings suggests that the potential mechanisms of cytotoxicity are dependent on both the nature and the concentration of the metals. PMID- 12123648 TI - The toxicity of commercial jet oils. AB - Jet oils are specialized synthetic oils used in high-performance jet engines. They have an appreciable hazard due to toxic ingredients, but are safe in use provided that maintenance personnel follow appropriate safety precautions and the oil stays in the engine. Aircraft engines that leak oil may expose others to the oils through uncontrolled exposure. Airplanes that use engines as a source of bleed air for cabin pressurization may have this source contaminated by the oil if an engine leaks. Examination of the ingredients of the oil indicates that at least two ingredients are hazardous: N-phenyl-1-naphthylamine (a skin sensitizer) and tricresyl phosphate (a neurotoxicant, if ortho-cresyl isomers are present). Publicly available information such as labels and MSDS understates the hazards of such ingredients and in the case of ortho-cresyl phosphates by several orders of magnitude. PMID- 12123649 TI - Metals in drinking water from new housing estates in the Sydney area. AB - Metals in drinking water were measured in 95 new houses less than 18 months old in the Sydney metropolitan area. Three samples (first-flush, post-first-flush, and fully flushed water) were collected from each house, and "control" samples from the five Sydney Water points that supplied the houses, a total of 326 samples. They were analyzed for Pb, Cu, Mn, Zn, Cd, and Al. At the supply points, the levels of all metals were at or below Australian Drinking Water Guidelines (ADWG). In the houses, metal levels varied. Of the first-flush samples, Pb was above ADWG in 60% and above US EPA Guidelines in 81%, Cu was above ADWG in 12%, and Cd was above ADWG in 4%. Of the post-first-flush samples, Pb was above ADWG in 24%, Cu was above ADWG in 18%, Cd was above ADWG in 1%, and Zn was above ADWG in 1%. The other metal contaminants (Mn and Al) were within ADWG. In fully flushed water, the levels of all metals were well below ADWG. PMID- 12123650 TI - Evaluation of heavy metals in fish of the Sepetiba and Ilha Grande Bays, Rio de Janeiro, Brazil. AB - Muscles, gonads, and liver tissues of fish caught in Sepetiba and Ilha Grande Bay were analyzed to assess levels of heavy metal concentrations of Cd, Fe, Zn, Pb, Ni, Cu, and Cr. Levels of Cr surpassed the maximum permissible concentration in muscle, followed by Zn in some species. Pb, Cu, and Cd presented concentrations above maximum permissible levels in gonads and liver only. Fe presented concentrations in viscera comparable to those of highly polluted areas, although there are no standards available for this metal. Ni was the only metal that did not present contamination in the fish tissues examined. Overall, fish caught in Sepetiba Bay showed higher metal concentrations than those from Ilha Grande Bay, but the latter also presented high metal concentrations in several samples. PMID- 12123651 TI - Butyl benzyl phthalate affects shoaling behavior and bottom-dwelling behavior in threespine stickleback. AB - In this laboratory experiment, the effects on fish behavior caused by butyl benzyl phthalate (BBP) were of interest. We showed that shoaling behavior and bottom-dwelling behavior in threespine stickleback, Gasterosteus aculeatus, were altered as a result of exposure to 0.1 mg/L BBP. Threespine sticklebacks, collected from a freshwater population in central Norway, were exposed to BBP for 26 days. BBP was administered daily through the water. We found that exposed fish aggregated more into one single shoal than control fish. Further, the exposed fish spent more time at the bottom of the test aquarium than the control fish. From these results we conclude that the behavior traits aggregation and bottom dwelling activity may be suitable and sensitive in detecting effects of BBP in threespine stickleback. PMID- 12123652 TI - Mannose 6-phosphate quantitation in glycoproteins using high-pH anion-exchange chromatography with pulsed amperometric detection. AB - An assay has been developed to quantitate the amount of mannose 6-phosphate in glycoproteins using high-pH anion-exchange chromatography with pulsed amperometric detection (HPAEC-PAD). The method was tested on a recombinant lysosomal enzyme, human alpha-galactosidase A, that contains mannose 6-phosphate. The assay includes two steps: hydrolysis of the glycoprotein in 6.75 M trifluoroacetic acid to release mannose 6-phosphate and quantitation of the released mannose 6-phosphate using HPAEC with PAD. There is a linear relationship between the amount of mannose 6-phosphate measured and the amount of alpha galactosidase hydrolyzed. The assay is also sensitive for as little as 2.5 microg alpha-galactosidase, which contains 117 pmol mannose 6-phosphate. Further, the assay has been shown to have good day-to-day and operator-to-operator consistency. In order to evaluate the assay for glycoprotein in crude extract, the glycoprotein was separated by SDS-PAGE and transferred to polyvinylidene difluoride membrane. The amount of mannose 6-phosphate in the electroblots following hydrolysis was determined using HPAEC-PAD. The assay was also linear when measuring mannose 6-phosphate on electroblots. Therefore, this assay has been shown to be specific, sensitive, and reproducible. PMID- 12123653 TI - 23Na NMR study of Fibrobacter succinogenes S85: comparison of three chemical shift reagents and calculation of sodium concentration using ionophores. AB - In order to measure intracellular sodium concentrations in resting cells of Fibrobacter succinogenes S85 by (23)Na NMR spectrometry, two methodological aspects were studied. First, three different shift reagents (Dy(PPP(i))(7-)(2), Tm(DOTP)(5-), and Dy(TTHA)(3-)) were tested for their ability to separate internal and external (23)Na NMR resonances. Their toxicity toward F. succinogenes cells was evaluated by in vivo(13)C NMR experiments. Tm(DOTP)(5-) was found to be the most efficient shift reagent while being nontoxic. Second, a new methodology was developed to calculate intracellular sodium concentration in F. succinogenes by using ionophores. This approach avoided the problem of intracellular volume measurement and that of sodium visibility determination. PMID- 12123655 TI - Electrochemical detection of nucleic base mismatches with ferrocenyl naphthalene diimide. AB - Electrochemical detection of nucleic base mismatches was attempted successfully with ferrocenyl naphthalene diimide (FND) in a model system with 20-meric double stranded oligonucleotides with or without a mismatch(es). Thus, dA(20) or a 20 meric sequence of the lac Z gene was immobilized on a gold electrode and complementary oligonucleotides with different numbers of mismatches were allowed to hybridize in the presence of FND to give rise to an electrochemical signal. The signal intensity varied depending on the number of unpaired bases on the DNA duplex. From experiments with a quartz crystal microbalance, eight molecules of FND were found to bind to the 20-meric double-stranded oligos and this number decreased as the number of mismatches increased. These findings were further supported by matrix-assisted laser desorption ionization time-of-flight mass spectroscopy. This novel method will be useful for the analysis of single nucleotide polymorphisms present on human genes. PMID- 12123654 TI - Application of isotopic ratio mass spectrometry for the in vitro determination of demethylation activity in human liver microsomes using N-methyl-13C-labeled substrates. AB - The reaction of demethylation mediated by cytochrome P450 (CYP) leads to the equimolar production of demethylated metabolite and formaldehyde. From a 13C substrate labeled on a carbon of the methyl moiety, [13C]formaldehyde (H13CHO) is liberated. A highly sensitive and specific assay involving the oxidation of H13CHO to 13CO(2) by a double-enzymatic-step reaction is reported. The 13CO(2) was quantified by the method of reverse isotopic dilution based on gas chromatography-isotope ratio mass spectrometry analysis. The method first involves the limiting step of the CYP-dependent reaction, which is stopped with a mixture of zinc sulfate 5 mM and trichloroacetic acid 100 mM. Then, the transformation of H13CHO to 13CO(2) is performed with the formaldehyde (0.2 unit) and the formate (0.2 unit) dehydrogenase NAD-dependent enzymes. The recovery of 13CO(2) from the incubation mixture was equal to 91.4 +/- 3.0%. The accuracy and the precision of the present method were within 12 and 10%, respectively. The limit of quantification was set to 25 pmol. The performance of the assay was validated on human liver microsomes with five probes: [13C]erythromycin, [1 13C]caffeine, [3-13C]caffeine, [7-13C]caffeine, and [13C(2)]aminopyrine. This method is useful for the rapid determination of N-demethylase activity of human liver microsomes from methyl-13C-substrates. PMID- 12123656 TI - LC/MS analysis of NAD biosynthesis using stable isotope pyridine precursors. AB - A liquid chromatographic-electrospray ionization ion trap mass spectrometry (LC/MS) method has been developed to measure the biosynthetic incorporation of specific precursors into NAD. The stable isotope-labeled precursors tryptophan, quinolinic acid, nicotinic acid, and nicotinamide were added to the media of human liver tumor cells (SK-HEP) grown in culture. The cells were harvested, the NAD was extracted, and the ratio of labeled to unlabeled NAD was measured using the newly developed LC/MS assay. The quantity of NAD formed from each precursor relative to an internal standard (fully labeled 13C, 15N-labeled NAD prepared from baker's yeast) was measured. The detection limit (signal-to-noise ratio 5:1) of the LC/MS method was 37 fmol (25 pg) of NAD and was linear from 20.0 ng to 25 pg. All reported NAD levels were normalized relative to cellular protein measurements. At 50 microM precursor concentrations, nicotinamide was the dominant precursor and NAD levels in the cell rose well above normal levels. Other precursors were minimally incorporated. The same methods were applied to NAD biosynthesized by macrophages derived from peripheral blood monocytes. However, the NAD concentration in macrophages was about 5% of that in SK-HEP cells and the incorporation of stable isotope-labeled substrates remained below measurable levels. PMID- 12123658 TI - Quantitation of gene-specific DNA damage by competitive PCR. AB - A sensitive assay for quantitating DNA damage within individual genes would be a valuable tool for identifying the molecular mechanisms of disease and the sites of action of various carcinogens and anticancer drugs. This report describes a competitive PCR assay that was used to quantitate DNA damage induced by anticancer drugs within a 683-bp region of the c-myc gene in human CEM leukemia cells. Absolute quantitation of gene-specific DNA damage (attomoles or molecules of damaged DNA sequences) was achieved by coamplification of a homologous internal standard that has the same primer binding sites and PCR amplification efficiency as c-myc. The variability (standard error) associated with four separate determinations of the amount of c-myc sequence in 300 ng of DNA from untreated cells (6.80 +/- 0.05 SE amol) was less than 1% of the mean. The assay was capable of quantitating direct DNA damage that was induced by therapeutic concentrations of VM-26 and cisplatin prior to the onset of cellular apoptosis or necrosis. Both VM-26 (1-10 microM) and cisplatin (25-100 microM) induced a dose dependent decrease in the amount of intact c-myc sequence. This assay should be readily adaptable to current real-time PCR protocols. PMID- 12123657 TI - Bioluminescence immunoassay for cortisol using recombinant aequorin as a label. AB - The analysis of hormones in saliva is a powerful tool in the assessment of a patient's endocrine function, since it allows multiple noninvasive samplings. Since salivary levels of most hormones are 10 to 50 times lower than plasma levels, accurate and highly sensitive assays are needed for saliva measurements. Herein, we describe the development of a solid-phase competitive immunoassay for cortisol in saliva, in which a mutant of the photoprotein aequorin has been used as a label. We have chemically conjugated cortisol to aequorin at different molar ratios. The various cortisol-aequorin conjugates were characterized in terms of bioluminescent activity and affinity for the anti-cortisol antibody. The conjugate that gave the best analytical performance was used for the development of the immunoassay and the analysis of cortisol in saliva samples. The conjugates were stable for at least 6 months when stored at 4 degrees C. The method fulfilled all the standard requirements of precision and accuracy. The optimized immunoassay gave a detection limit of 300 fmol/tube, corresponding to 3 nmol/L, with a linear dynamic range of 10-1000 nmol/L. Therefore, cortisol can be detected down to 0.1 ng in 100 microl of saliva sample using this assay, without any sample pretreatment. This detection limit is almost one order of magnitude lower than the physiological levels of salivary cortisol, which are reported to be 10-25 nmol/L. This allows the quantification of salivary cortisol to be performed in the linear range of the calibration curve, which is most reliable for quantification purposes. PMID- 12123659 TI - Macromolecular assembly of the transition state regulator AbrB in its unbound and complexed states probed by microelectrospray ionization mass spectrometry. AB - The Bacillus subtilis global transition-state regulator AbrB specifically recognizes over 60 different DNA regulatory regions of genes expressed during cellular response to suboptimal environments. Most interestingly the DNA regions recognized by AbrB share no obvious consensus base sequence. To more clearly understand the functional aspects of AbrB activity, microelectrospray ionization mass spectrometry has been employed to resolve the macromolecular assembly of unbound and DNA-bound AbrB. Analysis of the N-terminal DNA binding domain of AbrB (AbrBN53, residues 1-53) demonstrates that AbrBN53 is a stable dimer, showing no apparent exchange with a monomeric form as a function of pH, ionic strength, solvent, or protein concentration. AbrBN53 demonstrates a capacity for DNA binding, underscoring the role of the N-terminal domain in both DNA recognition and dimerization. Full-length AbrB is shown to exist as a homotetramer. An investigation of the binding of AbrBN53 and AbrB to the natural DNA target element sinIR shows that AbrBN53 binds as a dimer and AbrB binds as a tetramer. This study represents the first detailed characterization of the stoichiometry of a transition-state regulator binding to one of its target promoters. PMID- 12123660 TI - Compensation for loss of ligand activity in surface plasmon resonance experiments. AB - The determination of equilibrium binding constants is an important aspect of the analysis of protein-protein interactions. In recent years surface plasmon resonance experiments (e.g., with a BIAcore instrument) have provided a valuable experimental approach to determining such constants. The standard method is based on measuring amounts of analyte bound at equilibrium for different analyte concentrations. During the course of a typical surface plasmon resonance experiment the measured equilibrium levels for a given analyte concentration often decrease. This appears to be due to a loss of activity of the protein coupled to the sensor chip or other phenomena. The loss in signal can lead to an erroneous determination of the equilibrium constant. A data analysis approach is introduced that aims to compensate for the loss of activity so that its influence on the results of the experiments is reduced. PMID- 12123661 TI - A noninvasive cell-based assay for monitoring proteolytic activity within a specific subcellular compartment. AB - A noninvasive cell-based assay has been developed to monitor the proteolytic activity of cathepsin L within a specific subcellular compartment, the lysosome. The green fluorescent protein (GFP) of Aequorea victoria was selected as a substrate. Targeting to lysosomes was achieved by fusing GFP to preprocathepsin L, which also ensures colocalization of the enzyme and the substrate. Stably transfected HeLa-rtTA (reverse tetracycline-controlled transactivator) cells were induced with doxycycline and cultured in the presence of various concentrations of cysteine protease inhibitors for 48 h. In the absence of inhibitor, proteolytic degradation of GFP leads to loss of fluorescence, which is due almost exclusively to the action of recombinant cathepsin L. However, a dose-dependent increase of GFP fluorescence is observed for cells treated with the potent cathepsin L inhibitor benzyloxycarbonyl-LeuLeuTyr-CHN(2). Fluorescence is also observed when GFP is fused to an inactive preprocathepsin L (C25A mutant). Targeting of GFP to an acidic cellular compartment can destabilize the protein and render it susceptible to proteolytic degradation. The approach should be generally applicable for proteases localized in acidic environments. Such an assay can be of great value in validating the participation of a specific enzyme in a given process or in testing the ability of putative inhibitors to reach their intracellular target. PMID- 12123662 TI - Development of peptide substrates for trypsin based on monomer/excimer fluorescence of pyrene. AB - An assay using fluorogenic peptides based on the monomer/excimer fluorescence features of pyrene was developed to measure the proteolytic activity of trypsin, a serine protease. Two pyrene moieties were incorporated into the respective N- and C-terminus of the peptides as (pyrene)-C-Xaa-C-(pyrene), where Xaa represents amino acid residues of 5-, 6-, 7-, or 8-mer containing the cleavage site of trypsin. The proteolytic cleavage of the substrates led to an increase in monomer fluorescence and a decrease in excimer fluorescence of pyrene. Kinetic parameters (k(cat) and K(m)) for the enzymatic hydrolysis of the substrates were successfully determined. The parameters are dependent on the chain length of the substrate and optimal catalytic activity was obtained with substrates that consisted of 9 or 10 amino acid residues. The present assay system is sensitive and the preparation of the substrate is very simple. We suggest that this method may be suitable for high-throughput screening and also applicable to the characterization of other proteases. PMID- 12123663 TI - Determination of the chondroitin sulfate disaccharides in dog and horse plasma by HPLC using chondroitinase digestion, precolumn derivatization, and fluorescence detection. AB - A sensitive and selective HPLC method for the determination of the disaccharides of chondroitin sulfate in horse and dog plasma was validated. Chondroitin sulfate is degraded by chondroitinase ABC to three primary unsaturated disaccharides, (1) 2-acetamido-2-deoxy-3-O-(beta-D-gluco-4-enepyranosyluronic acid)-D-galactose, (2) 2-acetamido-2-deoxy-3-O-(beta-D-gluco-4-enepyranosyluronic acid)-4-O-sulfo-D galactose, and (3) 2-acetamido-2-deoxy-3-O-(beta-D-gluco-4-enepyranosyluronic acid)-6-O-sulfo-D-galactose, when treated with chondroitinase. Plasma samples (0.5 ml) were treated with 50 mU of chondroitinase ABC in 50 microl of 1 mM sodium phosphate buffer (pH 7.0) at 37 degrees C for 6 h. The samples were extracted with 25% trifluoroacetic acid in ethanol. The resultant samples were derivatized with 1% dansylhydrazine in ethanol at 40 degrees C for 3 h. The chromatographic conditions consisted of fluorescence detection (excitation at 350 nm and emission at 530 nm), mu-Bondapack NH(2) (300 x 3.9 mm), and mobile phase of acetonitrile:100 mM acetate buffer, pH 5.6 (76:24), pumped at 1.0 ml/min. The standard curves for each chondroitin disaccharide showed linearity over the selected concentration range (r > or = 0.99). The intraday percentage relative standard deviation was < or =9.5% and the interday precision was < or =6.9% or less. The relative intraday and interday error ranged from -7.3 to 6.6% for each chondroitin disaccharide in the plasma. The extraction recovery was found to be in the range of 90-96%. The validated method accurately quantitated the disaccharides of chondroitin sulfate after administration to dogs and horses. PMID- 12123664 TI - Identification of acetylation and methylation sites of histone H3 from chicken erythrocytes by high-accuracy matrix-assisted laser desorption ionization-time-of flight, matrix-assisted laser desorption ionization-postsource decay, and nanoelectrospray ionization tandem mass spectrometry. AB - A new strategy has been employed for the identification of the covalent modification sites (mainly acetylation and methylation) of histone H3 from chicken erythrocytes using low enzyme/substrate ratios and short digestion times (trypsin used as the protease) with analysis by HPLC separation, matrix-assisted laser desorption ionization-time-of-flight (MALDI-TOF), matrix-assisted laser desorption ionization-postsource decay, and tandem mass spectrometric techniques. High-accuracy MALDI-TOF mass measurements with representative immonium ions (126 for acetylated lysine, 98 for monomethylated lysine, and 84 for di-, tri-, and unmethylated lysine) have been effectively used for differentiating methylated peptides from acetylated peptides. Our results demonstrate that lysines 4, 9, 14, 27, and 36 of the N-terminal of H3 are methylated, while lysines 14, 18, and 23 are acetylated. Surprisingly, a non-N-terminal residue, lysine 79, in the loop region hooking up to the bound DNA, was newly found to be methylated (un-, mono-, and dimethylated isoforms coexist). The reported mass spectrometric method has the advantages of speed, directness, sensitivity, and ease over protein sequencing and Western-blotting methods and holds the promise of an improved method for determining the status of histone modifications in the field of chromosome research. PMID- 12123665 TI - Determination of 4-methylpropranolol in cerebrospinal fluid, serum, and urine by nonprotected fluid room-temperature phosphorescence using simplex optimization. AB - A direct and simple procedure for the determination of 4-methylpropranolol, a specific beta-adrenergic receptor blocking agent, in biological fluids was developed. The method was based on the measurement of the nonprotected fluid room temperature phosphorescence of the drug. This technique enables us to determine analytes in complex matrices without the need for a tedious prior separation process. The appropriate experimental conditions to obtain suitable reproducibility and maximum phosphorescence signal, when sodium sulfite is used to eliminate the oxygen from the solution and when potassium iodide is used as heavy atom, were studied. The optimum concentration of KI was 3.2 M. The optimization of Na(2)SO(3) (7.0 x 10(-3) M) and the accurate value of pH (10.88) were determined using a simplex as the method of optimization. A sodium carbonate hydrogen carbonate buffer solution (5.0 x 10(-2) M) was used to adjust the value of pH. The delay time (124 micros), gate time (206 micros), and time between flashes (5 ms) were also optimized using a simplex. Under the above conditions, the maximum signal of phosphorescence appears instantly once the sample has been prepared, and the intensity was measured at lambda(ex) = 300 nm and lambda(em) = 537 nm, in the concentration range 25-500 ng/ml. Overall least-squares regression was used to find the straight line that fit the experimental data. The detection limit according to the error propagation theory was 6.2 ng/ml and the detection limit calculated as proposed by C. A. Clayton et al. (1987, Anal. Chem. 59, 2506) was 11.7 ng/ml. The repeatability was studied using 10 solutions of 200 ng/ml 4 methylpropranolol; if error propagation theory was assumed, the relative error was 1.78% and the standard deviation for replicate samples was 3.5 ng/ml. This method was successfully applied to the determination of 4-methylpropranolol in urine, serum, and cerebrospinal fluid, with recoveries of 99.3 +/- 0.5% in the case of urine, 99.8 +/- 0.2% for serum, and 101.5 +/- 1.5% for cerebrospinal fluid. PMID- 12123666 TI - Gas chromatography-mass spectrometry assay of the (18)o enrichment of water as trimethyl phosphate. AB - We have developed an assay for determining the 18O enrichment of water in biological fluids. Urine, plasma, or whole blood is reacted with phosphorous pentachloride to yield phosphoric acid. Derivatization of phosphoric acid with diazomethane generates trimethyl phosphate. The enrichment of trimethyl phosphate is nearly four times that of water and is assayed using gas chromatography-mass spectrometry (electron impact ionization). Yang et al. (1998, Anal. Biochem. 258, 315-321) assayed the 2H enrichment of body water after exchange with acetone, by gas chromatography-mass spectrometry. The combination of our 18O method and the 2H method of Yang et al. allows one to measure energy expenditure via "doubly labeled" water (2H(2)O + H(2)18O), using small samples of body fluids. These techniques were used to measure energy expenditure in mice, in which the 18O enrichment of body water can be monitored down to 0.025%. PMID- 12123667 TI - Radiochemical malonyl-CoA decarboxylase assay: activity and subcellular distribution in heart and skeletal muscle. AB - Malonyl-CoA decarboxylase is the main route for the disposal of malonyl-CoA, the key metabolite in the regulation of mitochondrial fatty acid oxidation. We have developed a simple and sensitive radiochemical assay to determine malonyl-CoA decarboxylase activity. The decarboxylation of [2-14C]malonyl-CoA produces [2 14C]acetyl-CoA, which is converted to [2-14C]acetylcarnitine in the presence of excess L-carnitine and carnitine acetyltransferase. The positively charged radiolabeled product, acetylcarnitine, is separated from negatively charged excess radiolabeled substrate and the radioactivity measured by scintillation counting. Measurement of malonyl-CoA decarboxylase activities with this method gives values comparable to those obtained with assays currently in use, but has the advantage of being simpler and less labor intensive. We have applied this assay to rat skeletal muscle of different fiber-type composition and to rat heart. Malonyl-CoA decarboxylase activity (mU/g wet wt) correlates with the oxidative capacity of the muscles, being lowest in type IIb fibers (42.7 +/- 3.0) and highest in heart (1071.4 +/- 260), with intermediate activity in type IIa fibers (150.7 +/- 4.3) and type I fibers (107.8 +/- 7.6). Studies on subcellular distribution of malonyl-CoA decarboxylase activity in rat heart and rat skeletal muscle show that approximately 50 and 65% is localized to mitochondria, while 50 and 35% of the activity is extramitochondrial. PMID- 12123668 TI - Binding stoichiometry of an RNA aptamer and its transcription factor target. AB - RNA molecules serve informational, structural, and catalytic roles in cells. RNA also offers an interesting raw material for the design or genetic selection of modifiers of gene expression. We have been interested in the possibility that natural and/or artificial RNA ligands might be identified for DNA-binding proteins. With these concepts in mind, our laboratory previously isolated a 31 nucleotide RNA aptamer that specifically binds to human transcription factor NF kappaB. This RNA aptamer (alpha-p50) competitively inhibits DNA binding by NF kappaB in vitro. The aptamer may target the DNA-binding groove formed by the junction of the two monomers of NF-kappaB, perhaps mimicking kappaB duplex DNA. This model predicts a binding stoichiometry of one RNA aptamer per NF-kappaB dimer. To test this hypothesis, two complementary biophysical methods were utilized. Both analytical ultracentrifugation and microelectrospray mass spectrometry suggest that 1 mol of alpha-p50 RNA binds per mole of NF-kappaB p50 homodimer. Such a result is consistent with the observed ability of the RNA aptamer to block the access of transcription factor NF-kappaB to its binding site on DNA and highlights the question of how an RNA stem-loop structurally mimics a DNA duplex. This work also demonstrates the successful application of mass spectrometry to characterize noncovalent RNA/protein interactions. PMID- 12123669 TI - Immunoblotting assays for keratan sulfate. AB - The detection of microquantities of glycosaminoglycans (GAGs) in biological samples has been hampered by the lack of sensitive methods. In this paper we describe the modification and development of three sensitive assays capable of detecting nanogram quantities of GAGs in biological samples. The first assay detects total GAGs. It is a modified Alcian blue dye precipitation assay in which the dye binds to the negatively charged GAGs in CsCl-fractionated extracts from chicken tendons. This assay compares favorably with the widely used uronic acid assay in terms of its sensitivity and ability to detect all classes of GAGs, including keratan sulfate (KS). Two other assays, dot-blotting and immunoblotting, detect KS in complex mixtures and can be easily adapted for the detection of other GAGs. Both take advantage of binding of carboxyl and sulfate groups of GAGs to trivalent neodymium. In dot-blotting, samples were directly blotted onto nitrocellulose membrane soaked in Nd(2)(SO(4))(3) buffer, and KS was detected with the monoclonal anti-KS 5-D-4 antibody and an avidin-biotin complex detection system. In immunoblotting, the samples were first separated in 28% polyacrylamide gels, transferred onto a Nd(2)(SO(4))(3)-soaked nitrocellulose membrane using a phosphate buffer system, and stained and developed using the same protocol as in dot-blotting. Whereas dot-blotting allows the use of very low quantities of samples because of its high sensitivity (lower detection limit was 5 ng), immunoblotting provides more specificity. PMID- 12123670 TI - A cell-free electrochemiluminescence assay for measuring beta1-integrin-ligand interactions. AB - We have developed a cell-free assay for binding of solubilized beta1 integrins to their physiologically relevant ligands using an electrochemiluminescent detection method. The method utilizes ruthenium-conjugated monoclonal antibodies for detection of either purified integrins or, more conveniently, integrin-expressing cell lysates, which are captured on beads coated with extracellular matrix or vascular ligand proteins. For the interaction of alpha1beta1 integrin with collagen IV, a signal of 10-fold over background was generated with samples containing only 10 ng (0.05 pmol) of integrin. This interaction is cation dependent and can be inhibited by blocking antibodies to the alpha1 subunit. The method was extended to studies of ligand binding by integrins alpha2beta1, alpha4beta1, alpha5beta1, and alpha6beta1. For each integrin-ligand pair, the specificity of the interaction was verified with neutralizing antibodies against the specific integrin. The specific binding signal correlated with the activating ability of the labeled antibody used for detection, although the ability of divalent cations (Mn2+, Mg2+, Ca2+) to support integrin-ligand binding varied dramatically among the various integrin-ligand pairs. The assay provides a simple method for investigating integrin-ligand interactions without avidity and/or signaling effects which can complicate conventional cell-based assay methods. PMID- 12123671 TI - Bacterial expression of in vivo-biotinylated aequorin for direct application to bioluminometric hybridization assays. AB - We have constructed a plasmid suitable for bacterial expression of in vivo biotinylated photoprotein aequorin. The biotin tag facilitates the isolation of aequorin from crude cell extract and the direct complexation of aequorin with streptavidin for the development of highly sensitive hybridization assays, thereby avoiding the need for chemical crosslinking. The plasmid contains a biotin-acceptor coding sequence fused to an apoaequorin gene. The birA gene, encoding biotin protein ligase (BPL), is inserted downstream of the apoaequorin sequence. BPL biotinylates, posttranslationally, the acceptor domain at a unique position. Functional aequorin is generated by incubating the lysate with coelenterazine and is purified by using a monomeric avidin column that allows elution under nondenaturing conditions. The biotinylated aequorin is complexed with streptavidin and used as a reporter molecule in a hybridization assay. The assay entails immobilization of an oligonucleotide probe on microtiter wells followed by hybridization with a denatured DNA target labeled with biotin through PCR. Streptavidin-biotinylated aequorin is used for quantification of the hybrids. Luminescence is measured in the presence of excess Ca(2+). The analytical range extends from 80 amol of target DNA per well (with a signal-to background ratio of 2.1) up to 40 fmol per well. The coefficient of variation is about 6%. In vivo-biotinylated aequorin produced from 1 liter of culture is sufficient for 300,000 hybridization assays. PMID- 12123673 TI - A biotin capture assay for oligosaccharyltransferase. AB - Oligosaccharyltransferase (OST) catalyzes the en bloc transfer of dolichylpyrophosphate oligosaccharides to an asparagine residue found in the sequon Asn-Xaa-Thr/Ser of newly synthesized proteins. Currently the method most commonly used to monitor this reaction, involving multiple solvent extractions and HPLC, is extremely time consuming and tedious. Herein, we present the use of a biotinylated peptide as the acceptor substrate and dolichylpyrophosphate [3H]chitobiose as the donor substrate for the OST-catalyzed reaction. This allows for separation (avidin-agarose beads) and quantitative analysis (scintillation counting) of only the biotinylated glycopeptide product of the OST-catalyzed reaction. This new assay yields highly reproducible results in a rapid manner. PMID- 12123672 TI - A convenient one-step extraction of cellular ATP using boiling water for the luciferin-luciferase assay of ATP. AB - Cellular ATP is commonly determined as production of bioluminescence using a luciferin-luciferase reaction system. Before the measurement of bioluminescence, cellular ATP must first be extracted. Two commonly used extraction methods are: () Tris-borate buffer (pH 9.2) coupled with a heating process (to inactivate ATPase) and () perchloric acid followed by neutralization. However, we found that both Tris-borate buffer and perchloric acid interfered with the luciferin luciferase system. Here, we report a convenient single-step boiling deionized water (DW) method for extracting cellular ATP to replace perchloric acid and Tris borate buffer. We showed that the boiling DW method did not interfere with the bioluminescence and was effective in inhibiting ATPase. This improved method required no neutralization and dilution and thus was more convenient than the perchloric acid method. Unlike the Tris-borate/heating procedure, our method did not require a separate heating step because boiling DW effectively inhibited ATPase and thus accomplished the two missions in one step for both suspended and attached cells. The improved method was precise for both suspended cells and attached cells, when cell numbers were between 10(3) and 10(6). The method also was more sensitive than other methods because it required much fewer cells (10(4) to 10(5)) than other methods for ATP determination. Thus, this one-step method is suitable for routine assay of cellular ATP for both suspended and attached cells. PMID- 12123674 TI - Zymography of monophenolase and o-diphenolase activities of polyphenol oxidase. PMID- 12123675 TI - Simultaneous detection of methylenetetrahydrofolate reductase gene polymorphisms, C677T and A1298C, by melting curve analysis with LightCycler. PMID- 12123676 TI - Modifying differential display polymerase chain reaction to detect relative changes in gene expression profiles. PMID- 12123677 TI - Site-directed mutagenesis by the megaprimer PCR method: variations on a theme for simultaneous introduction of multiple mutations. PMID- 12123678 TI - A high-throughput microplate method for assessing aggregation of deoxygenated hemoglobin s heterotetramers in vitro. PMID- 12123679 TI - Adaptation to chronic PCP results in hyperfunctional NMDA and hypofunctional GABA(A) synaptic receptors. AB - Schizophrenia is currently thought to be associated with a hypoglutamatergic state that is mimicked by acute phencyclidine (PCP), an antagonist of the N methyl-D-aspartate (NMDA) receptor subtype. In this study we tested the hypothesis that chronic treatment of rats with this antagonist may be a more appropriate animal model than acute exposure since it could result in adaptive synaptic responses that would model certain aspects of the schizophrenic state in humans. In vitro intracellular electrophysiological recordings employing brain slices from rats treated chronically in vivo with PCP demonstrated that chronic PCP caused a substantial increase in synaptic responses mediated by NMDA receptors without any significant changes in alpha-amino-3-hydroxy-5-methyl-4 isoxazolepropionic acid/kainate-mediated synaptic responses. At the same time, GABA(A) receptor-mediated inhibitory responses were depressed significantly. Pharmacological and paired-pulse facilitation experiments demonstrated that these adaptive responses following chronic PCP administration were not the result of altered glutamate or GABA release. Immunoblot analyses suggest that the hyperfunctional NMDA response is at least partially mediated by an increased synthesis of NR1 and NR2A subunits as well as a change in the subunit stoichiometry of the NMDA receptor. This change in receptor composition was also supported by pharmacological experiments with a subunit selective NMDA antagonist. Our data support a reconsideration of NMDA and GABA(A) receptor responsiveness following a chronic, not acute, exposure to PCP and the adaptations that persist after such a regimen. PMID- 12123680 TI - Moderate hypoglycemia aggravates effects of hypoxia in hippocampal slices from diabetic rats. AB - We recorded the effects of hypoxia combined with relative hypoglycemia on pre- and post-synaptic potentials in the CA1 area of slices from 4-month-old control and diabetic (streptozotocin-treated) Wistar rats. In experiments on slices kept in 10 or 4 mM glucose (at 33 degrees C), hypoxia was applied until the pre synaptic afferent volley disappeared--after 12-13 min in most slices, but much earlier (5+/-0.8 min) in diabetic slices kept in 4 mM glucose. When oxygenation was resumed, the afferent volley returned in all slices, for an overall mean recovery of 86.5% (+/-8.8%). Field post-synaptic potentials were fully blocked within 2-3 min of the onset of hypoxia. After the end of hypoxia, they failed to reappear in some slices: overall, their recovery varied between 62 and 68% in control slices, as well as in diabetic slices kept in 10 mM glucose; but recovery was very poor in diabetic slices kept in 4 mM glucose (only 15+/-0.94%). In the latter, hypoxic injury discharges occurred earlier (4.2+/-0.68 min vs. 6.5-8 min for other groups). We conclude that diabetes appears to make hippocampal slices more prone to irreversible loss of synaptic function and early block of axonal conduction when temporary hypoxia is combined with moderate hypoglycemia. PMID- 12123681 TI - Similar ultrastructural distribution of the 5-HT(2A) serotonin receptor and microtubule-associated protein MAP1A in cortical dendrites of adult rat. AB - As visualized by light and electron microscopic immunocytochemistry, the distribution of the neuronal serotonin-2A (5-HT(2A)) receptor is mainly intracellular throughout adult rat brain. This localization is particularly striking in the pyramidal cells of cerebral cortex, the dendrites of which are intensely immunoreactive, but without any labeling of their spines. In view of recent yeast two-hybrid and biochemical results suggesting an association of 5 HT(2A) receptors with the cytoskeletal microtubule-associated protein MAP1A, the respective subcellular distributions of the receptors and of MAP1A were compared by quantitative electron microscopic immunocytochemistry in dendrites of adult rat frontoparietal cortex. Counts of silver-intensified immunogold particles revealed a higher density of 5-HT(2A) receptors in smaller rather than larger dendrites, and an apportionment between pre-defined compartments representing the plasma membrane and the cytoplasm that was proportional to the relative surface area of these compartments. MAP1A immunoreactivity also predominated in smaller versus larger dendrites, but with a slightly lower proportion of labeling in the plasma membrane versus cytoplasmic compartment. The co-localization of 5-HT(2A) receptors and MAP1A protein in the same dendrites could be demonstrated in double immunolabeling experiments. These results confirmed the predominantly somato dendritic, intracellular localization of 5-HT(2A) receptors in cerebral cortex, showed their higher concentration in distal as opposed to proximal dendrites, and suggested their potential association to the cytoskeleton in cortical neurons in vivo. Such a distribution of 5-HT(2A) receptors reinforces our earlier hypothesis that 5-HT(2A) receptors participate in intraneuronal signaling processes involving the cytoskeleton, and raises the possibility that their activation could be dependent upon that of another co-localized, plasma membrane-bound, 5-HT receptor. PMID- 12123682 TI - Pituitary adenylate cyclase-activating polypeptide produces a phase shift associated with induction of mPer expression in the mouse suprachiasmatic nucleus. AB - The main mammalian circadian pacemaker is located in the suprachiasmatic nucleus of the hypothalamus. Clock genes such as the mouse Period gene (mPer) play a role in this core clock mechanism in the mouse. With brief light exposure during the subjective night, the photic information, which is conveyed directly to the suprachiasmatic nucleus via the retinohypothalamic tract, results in mPer1 and mPer2 expression in the suprachiasmatic nucleus. Glutamate and pituitary adenylate cyclase-activating polypeptide (PACAP) are co-stored in the retinohypothalamic tract. Recent studies have suggested that not only glutamate but also PACAP are key players in the phase shift that occurs during subject night; however, research demonstrating a direct association between the PACAP induced phase shift and mPer gene expression has yet to be conducted. In the present study, PACAP (200 pmol) injected into the lateral ventricle during subjective night (circadian time 16; circadian time 12, onset of locomotor activity) caused a moderate phase delay associated with moderate expression of mPer1 and only slight expression of mPer2 in the mouse suprachiasmatic nucleus. PACAP-induced mPer1 expression was also observed in the paraventricular nucleus and periventricular area of the hypothalamus. (+)MK-801 (0.5 mg/kg), an N-methyl D-aspartate (NMDA) receptor antagonist, suppressed both the PACAP-induced phase delay and mPer1 expression. From these results we suggest that PACAP induces phase delays in the mouse circadian rhythm in association with an increase of mPer expression in the suprachiasmatic nucleus via the activation of NMDA receptors. PMID- 12123683 TI - Neuroprotective effects of the antifungal drug clotrimazole. AB - Pretreatment with 10 microM of the antifungal drug clotrimazole potently reduced the death of cultured rat cerebellar granule cells induced by oxygen/glucose deprivation, and the excitotoxic effect of glutamate on cultured hippocampal neurons and cerebellar granule cells. In patch-clamped hippocampal pyramidal neurons, 10-50 microM clotrimazole caused a decrease in the amplitude of N-methyl D-aspartate (NMDA) receptor-mediated currents. Glutamate induced intracellular Ca(2+) overload, as measured by Fluo-3 confocal fluorescence imaging, while clotrimazole reduced Ca(2+) overload and promoted the recovery of intracellular calcium homeostasis after glutamate treatment. Using tetramethylrhodamine ethyl ester fluorescence as a marker of mitochondrial membrane potential we found that clotrimazole prevented the glutamate-induced loss of mitochondrial membrane potential. Our data provide evidence that the protective effect of clotrimazole against oxygen/glucose deprivation and excitotoxicity is due to the ability of this drug to partially block NMDA receptor-gated channel, thus causing both reduced calcium overload and lower probability of the mitochondrial potential collapse. PMID- 12123684 TI - Intrinsic thermal resistance of the canine brain. AB - Hyperthermia above a critical threshold results in multisystemic changes that include neurological manifestations of heat stroke. It is unknown if the latter represents an intrinsic thermal sensitivity of the CNS or whether injury is secondary to physiological responses of non-CNS origin. To address this issue, the present work examined functional, structural, and biochemical changes in the CNS of dogs subjected to a thermal dosage immediately below that which induces disseminated intravascular coagulation with secondary multiple organ injury. The experimental approach is previously reported, inducing a 42.5 degrees C, 90 min, whole body hyperthermia while preventing other physiological responses to treatment, including respiratory alkalosis and significant reductions in mean arterial pressure. Functional analyses included neurologic examinations and brainstem auditory evoked potential recordings in the post-treatment interval in both hyperthermic and euthermic control populations. Biochemical and structural analyses examined the expression of 70-kDa heat shock proteins, cytokines, markers of astroglial and microglial injury/activation, evidence of vascular endothelial damage, and evidence of neuronal and axonal injury in brain between 0.5 h and 8 days from the end of the treatment. The only significant change associated with treatment was induction of the major inducible 70-kDa heat shock protein, this being most prominent in the cerebellum with maximal expression at 6 h and a return to baseline by 8 days.Collectively, from these results we suggest that the canine brain is intrinsically resistant to sublethal hyperthermia such that when CNS lesions occur, they do so in the presence of other physiological derangements. PMID- 12123685 TI - Medullary reticulospinal tract mediating the generalized motor inhibition in cats: II. Functional organization within the medullary reticular formation with respect to postsynaptic inhibition of forelimb and hindlimb motoneurons. AB - We compared postsynaptic inhibitory effects on forelimb motoneurons and those on hindlimb motoneurons during generalized motor inhibition evoked by stimulating the medullary reticular formation in decerebrate cats. Here, we address two questions. First, whether the medullary inhibitory effects upon forelimb motoneurons are equivalent to those upon hindlimb motoneurons. Second, whether there is a somatotopographical organization within the medullary reticular formation in terms of inhibitory connections with motoneurons. Repetitive stimulation (20-50 microA, 50-100 Hz) delivered to the dorsomedial medullary reticular formation bilaterally suppressed muscle tone of both the forelimbs and hindlimbs. The medullary stimulation hyperpolarized the membrane potentials of the forelimb (5.4+/-1.8 mV, n=46) and hindlimb (5.4+/-2.0 mV, n=59) motoneurons together with a decrease in input resistance. The degree of membrane hyperpolarization and input resistance was not different in the forelimb and hindlimb motoneurons. The medullary stimulation also depressed the capability of generating antidromic and orthodromic spikes in the motoneurons. Stimuli with pulse trains (one to three pulses, 5-10-ms intervals, 20-50 microA) applied to the medullary inhibitory region induced a mixture of excitatory and inhibitory postsynaptic potentials in the motoneurons. The most noteworthy potentials were the inhibitory postsynaptic potentials with a late latency. They were observed in most forelimb (n=57/58, 98.3%) and hindlimb (n=63/64, 98.4%) motoneurons. The inhibitory potentials in forelimb motoneurons had a latency of 25-30 ms and a peak latency of 35-40 ms, and those in hindlimb motoneurons had a latency of 30 35 ms and a peak latency of 50-60 ms. A difference was not observed in the location of the effective sites for evoking the inhibitory effects in the forelimb and hindlimb motoneurons. These sites were homogeneously distributed in the dorsomedial part of the medullary reticular formation corresponding to the location of the nucleus reticularis gigantocellularis. From these findings we suggest that there is an equivalent amount of the postsynaptic inhibitory effects exerted on forelimb and hindlimb motoneurons during medullary-induced generalized motor inhibition. In addition, the medullary reticular formation may be functionally organized as a homogeneous or non-specific region in terms of the medullary reticulospinal inhibitory connections with forelimb and hindlimb motoneurons. PMID- 12123686 TI - Kindling induces the mRNA expression of methyl DNA-binding factors in the adult rat hippocampus. AB - We have investigated the gene expression responses of a family of methyl CpG binding domain-containing factors (MeCP2, MBD1, MBD2, and MBD3) in the hippocampus of electrically kindled rats. Expression was examined in both amygdala- and partial perforant-pathway-kindled subjects, 24 h and 28 days following the final stimulation. In general, the responses of MBDs 2 and 3 paralleled each another, both temporally and spatially. The expression of both genes was significantly elevated in all hippocampal subfields at 24 h following either the fifth stage 5 seizure (amygdala kindling) or the 15th stimulation of the perforant pathway. This induced expression was transient, however, as the expression of both genes returned to control levels by 28 days. This pattern of response contrasted to that observed for MeCP2 and MBD1. MeCP2 displayed no change in expression either 24 h or 28 days after amygdala kindling, but did display a late-developing, significant increase in expression in the dentate gyrus at 28 days following perforant-pathway kindling. The expression of MBD1 was unchanged by partial perforant-pathway kindling, but was induced in the dentate gyrus 28 days after amygdala kindling. These results demonstrate that electrical kindling alters the hippocampal expression of methyl DNA-binding factors, but does not affect each factor equivalently. The responsive patterns observed suggest that this family of transcriptional regulators can be differentially altered in the hippocampus by seizure activity. PMID- 12123687 TI - Calcineurin as a potential contributor in estradiol regulation of hippocampal synaptic function. AB - Estradiol influences Ca(2+) regulation and Ca(2+)-dependent synaptic plasticity, suggesting estrogenic effects on Ca(2+)-dependent enzymes that regulate synaptic plasticity may mediate hormonal influences on cognition. In ovariectomized female rats, injections of estradiol benzoate (EB, 10 microg) reduced hippocampal cytosolic activity of serine/threonine protein phosphatases, calcineurin and protein phosphatase 1 (PP1). The decreased activity was rapid and recovered substantially over a 24-h period. Decreased calcineurin activity was associated with a decreased level of calcineurin in the cytosol. In contrast, expression of PP1 was not altered suggesting that the level of calcineurin activity regulated PP1 activity. EB application to hippocampal slices rapidly decreased cytosolic phosphatase activity, which was not blocked by the estrogen receptor antagonist, ICI 182780. Decreased phosphatase activity was associated with an increase in CA3 CA1 synaptic transmission. In addition, EB application shifted synaptic plasticity, blocking the induction of long-term depression and facilitating the establishment of long-term potentiation. The reduction in calcineurin activity and shift in synaptic plasticity were mimicked to a lesser extent by 17-alpha estradiol. From these results we suggest that EB can act to rapidly influence Ca(2+) signaling pathways including the activity of Ca(2+)-regulated phosphatases involved in synaptic plasticity. PMID- 12123688 TI - Inhibition of spinal protein kinase Calpha expression by an antisense oligonucleotide attenuates morphine infusion-induced tolerance. AB - Protein kinase C isoforms including the alpha isozyme have been implicated in morphine tolerance. In the present study, we examined the effect of intrathecal delivery of an antisense oligonucleotide targeting rat protein kinase Calpha mRNA on the expression of spinal protein kinase Calpha isozyme and spinal morphine tolerance. Continuous intrathecal infusion of rats with morphine produced an increase in paw withdrawal threshold to thermal stimulation on day 1, which disappeared by day 5. On day 6, a bolus intrathecal injection of morphine (a probe dose) produced significantly less analgesia in morphine-infused rats than in saline-infused rats, suggesting tolerance. Intrathecal treatment with the protein kinase Calpha antisense concurrent with spinal morphine infusion not only maintained the analgesic effect of morphine during the 5-day infusion, it also significantly increased responsiveness to the probe morphine dose on day 6. In comparison, the missense used in the same treatment paradigm had no effect. The inhibitory effect of protein kinase Calpha antisense on spinal morphine tolerance was dose-dependent, and reversible. Intrathecal treatment with the antisense, but not the missense, in rats decreased expression of spinal protein kinase Calpha mRNA and protein, as revealed by real-time quantitative reverse transcription polymerase chain reaction and western blots. Expression of the gamma isozyme was not affected by the oligonucleotides. The antisense also attenuated protein kinase C-mediated phosphorylation in spinal cord. These results demonstrate that selective reduction in the expression of the spinal protein kinase Calpha isozyme followed by a decrease of local protein kinase C-mediated phosphorylation will reverse spinal morphine infusion-induced tolerance. This finding is consistent with the view that tolerance produced by morphine infusion is dependent upon an increase in phosphorylation by protein kinase C, and also it emphasizes that the protein kinase Calpha isozyme and its activation in spinal cord may specifically participate in the phenomenon of opiate tolerance. PMID- 12123689 TI - GABA release in the medial preoptic area of cyclic female rats. AB - GABA is a potent regulator of gonadotropin-releasing hormone neurons in the hypothalamus. To determine the profile of GABA release in the medial preoptic area where the gonadotropin surge generator resides, an in vivo microdialysis study was performed in cyclic female rats. The microdialysis samples were collected and sequential blood samples (150 microl each) were also obtained, at 1 h intervals. During estrus and diestrus 1, GABA release in the medial preoptic area was relatively low. A small increase in the GABA release began in the afternoon of diestrus 1 and attained its peak in the morning of diestrus 2, but declined in the afternoon of that day. The GABA release markedly increased from late in the night of diestrus 2 through the morning of proestrus, when it attained its peak, and thereafter it declined sharply until the critical period of proestrus. A distinct preovulatory luteinizing hormone surge was observed in the afternoon of proestrus in all proestrous rats. From these results we suggest that the preovulatory elevation of the GABA release from the night through to the morning of proestrus, followed by a sharp decline, is closely associated with the onset of the preovulatory luteinizing hormone surge in cyclic female rats. The present study is the first to report the 4-day profile of GABA release in the medial preoptic area during the estrous cycle. PMID- 12123690 TI - Investigation of the modulation of glutamate release by sodium channels using neurotoxins. AB - The modulation of neurotransmitter release by calcium channels is well established, yet, sodium channels were regarded mainly as charge carriers. Many lines of evidence suggest a more fine-tuning role played by sodium channels. Using rat cerebrocortical isolated nerve endings (synaptosomes) and two toxins that have separate sites of action on sodium channels and provoke distinct changes in channel kinetics, we were able to show that depending on the rate of increase in channel conductance, the outcome in terms of neurotransmitter release and calcium channel types coupled to that event are different. Mainly, our study focused on veratridine, an alkaloid from lilaceous plants that binds to sodium channel toxin site 2, and tityustoxin, a toxin purified from the venom of the Brazilian yellow scorpion Tityus serrulatus that binds to site 3. Veratridine induces a slower increase in intrasynaptosomal sodium and calcium concentrations, slower depolarization, delayed exocytosis and a slower and predominantly calcium independent glutamate release, when compared to tityustoxin.Thus, we have used these two toxins to investigate the events that start with sodium entry and culminate with the release of glutamate in isolated nerve endings (synaptosomes) from rat cerebral cortex. With that in mind we measured intrasynaptosomal free sodium concentration [Na(+)](i), intrasynaptosomal free calcium concentration [Ca(2+)](i), membrane potential, exocytosis and glutamate release using fluorescent probes. PMID- 12123691 TI - Quantal transmitter release by glioma cells: quantification of intramembrane particle changes. AB - Glial cells in situ are able to release neurotransmitters such as glutamate or acetylcholine (ACh). Glioma C6BU-1 cells were used to determine whether the mechanisms of ACh release by a glial cell line are similar or not to quantal release from neurones. Individual C6BU-1 cells, pre-filled with ACh, were moved into contact with a Xenopus myocyte that was used as a real-time ACh detector. Upon electrical stimulation, C6BU-1 cells generated evoked ACh impulses which were Ca(2+)-dependent and quantal (quantal steps of ca. 100 pA). Changes in plasma membrane ultrastructure were investigated by using a freeze-fracture technique designed for obtaining large and flat replicas from monolayer cell cultures. A transient increase in the density of medium and large size intramembrane particles--and a corresponding decrease of small particles- occurred in the plasma membrane of C6BU-1 cells stimulated for ACh release. Changes in interaction forces between adjacent medium and large particles were investigated by computing the radial distribution function and the interaction potential. In resting cells, the radial distribution function revealed a significant increase in the probability to find two particles separated by an interval of 24 nm; the interaction potential suggested repulsive forces for intervals shorter than 24 nm and attractive forces between 24 and 26 nm. In stimulated cells, this interaction was displaced to 21 nm and made weaker, despite of the fact that the overall particle density increased. The nature of this transient change in intramembrane particles is discussed, particularly with regard to the mediatophore proteolipid which is abundant in the membranes C6-BU-1 like in those of cholinergic neurones. In conclusion, evoked ACh release from pre filled C6-BU-1 glioma cells is quantal and Ca(2+)-dependent. It is accompanied by a transient changes in the size distribution and the organisation of intramembrane particles in the plasma membrane. Thus, for the release characteristics, glioma cells do not differ fundamentally from neurones. PMID- 12123692 TI - Neuronal response to local electrical stimulation in rat thalamus: physiological implications for mechanisms of deep brain stimulation. AB - High-frequency deep brain stimulation (DBS) of sensorimotor thalamus containing "tremor cells" leads to tremor arrest in humans with parkinsonian and essential tremor. To examine the possible underlying mechanism(s), we recorded in vitro intracellular responses of rat thalamic neurons to local intrathalamic stimulation. Such simulated DBS (sDBS) induced a sustained membrane depolarization accompanied by an increase in apparent membrane conductance in both motor and sensory neurons. With stimulation frequency above approximately 100 Hz, the sDBS-induced depolarization most typically led to repetitive neuronal firing or less frequently resulted in a complete blockade of action potential genesis. When regular intracellular current pulses were injected into cells to mimic "tremor" activity, such rhythmic discharges were invariably disrupted or abolished by the random spike firing induced during high-frequency sDBS. Low frequency sDBS left rhythmicity unaffected.We conclude that clinical thalamic DBS may lead to a neuronal de-rhythmicity and tremor stoppage through masking and/or blocking rhythmic firing of tremor cells. PMID- 12123693 TI - Single cell laser dissection with molecular beacon polymerase chain reaction identifies 2A as the predominant serotonin receptor subtype in hypoglossal motoneurons. AB - We hypothesize that sleep state-dependent withdrawal of serotonin (5 hydroxytryptamine, 5-HT) at upper airway (UAW) dilator motoneurons contributes significantly to sleep-related suppression of dilator muscle activity in obstructive sleep apnea. Identification of 5-HT receptor subtypes involved in postsynaptic facilitation of UAW motoneuron activity may provide pharmacotherapies for this prevalent disorder. We have adapted two assays to provide semi-quantitative measurements of mRNA copy numbers for 5-HT receptor subtypes in single UAW motoneurons. Specifically, soma of 111 hypoglossal (XII) motoneurons in 10 adult male rats were captured using a laser dissection microscope, and then used individually in single round molecular beacon polymerase chain reaction (PCR) for real-time quantitation of 5-HT(2A), 5-HT(2C), 5-HT(3), 5-HT(4), 5-HT(5A), 5-HT(5B), 5-HT(6) or 5-HT(7) receptor. Receptor mRNA copy numbers from single XII motoneurons were compared to control samples from within the XII nucleus and lateral medulla. All 20 motoneuronal soma assayed for the 5-HT(2A) receptor had measurable copy numbers (7028+/-2656 copies/cell). In contrast, copy numbers for the 5-HT(2A) receptor in XII non-motoneuronal (n=17) and lateral medulla (n=15) samples were 81+/-51 copies and 83+/-35 copies, respectively, P<0.05. Seven of 13 XII motoneurons assayed had measurable 5-HT(2C) receptor copy numbers of mRNA (287+/-112 copies/cell). XII soma had minimal 5 HT(3), 5-HT(4), 5-HT(5A), 5-HT(5B), 5-HT(6) or 5-HT(7) receptor mRNA. 5-HT(2A) receptor mRNA presence within XII motoneurons was confirmed with digoxigenin labeled in situ hybridization. In summary, combined use of laser dissection and molecular beacon PCR revealed 5-HT(2A) receptor as the predominant 5-HT receptor mRNA in XII motoneurons, and identified small quantities of 5-HT(2C) receptor. This information will allow a more complete understanding of serotonergic control of respiratory activity. PMID- 12123695 TI - Estrogen up-regulates estrogen receptor alpha and synaptophysin in slice cultures of rat hippocampus. AB - Previous studies have shown that estrogen application increases the density of synaptic input and the number of spines on CA1 pyramidal neurons. Here, we have investigated whether Schaffer collaterals to CA1 pyramidal cells are involved in this estrogen-induced synaptogenesis on CA1 pyramidal neurons. To this end, we studied estrogen-induced expression of both estrogen receptor (ER) subtypes (ERalpha and ERbeta) together with the presynaptic marker synaptophysin in the rat hippocampus. In tissue sections as well as in slice cultures mRNA expression of ERalpha, ERbeta and synaptophysin was higher in CA3 than in CA1, and mRNA expression and immunoreactivity for both ER subtypes were found in both principal cells and interneurons. By using quantitative image analysis we found stronger nuclear immunoreactivity for ERalpha in CA3 than in CA1. In slice cultures, supplementation of the medium with 10(-8) M estradiol led to an increase of nuclear immunoreactivity for ERalpha, but not for ERbeta, which was accompanied by a dramatic up-regulation of synaptophysin immunoreactivity in stratum radiatum of CA1. Together these findings indicate that estrogen effects on hippocampal neurons are more pronounced in CA3 than in CA1 and that ER activation in CA3 neurons leads to an up-regulation of a presynaptic marker protein in the axons of these cells, the Schaffer collaterals. We conclude that estradiol-induced spine formation on CA1 pyramidal cells may be mediated presynaptically, very likely by activation of ERalpha in CA3 pyramidal cells, followed by an increase in Schaffer collateral synapses. PMID- 12123694 TI - Origins of skeletal pain: sensory and sympathetic innervation of the mouse femur. AB - Although skeletal pain plays a major role in reducing the quality of life in patients suffering from osteoarthritis, Paget's disease, sickle cell anemia and bone cancer, little is known about the mechanisms that generate and maintain this pain. To define the peripheral fibers involved in transmitting and modulating skeletal pain, we used immunohistochemistry with antigen retrieval, confocal microscopy and three-dimensional image reconstruction of the bone to examine the sensory and sympathetic innervation of mineralized bone, bone marrow and periosteum of the normal mouse femur. Thinly myelinated and unmyelinated peptidergic sensory fibers were labeled with antibodies raised against calcitonin gene-related peptide (CGRP) and the unmyelinated, non-peptidergic sensory fibers were labeled with the isolectin B4 (Bandeira simplicifolia). Myelinated sensory fibers were labeled with an antibody raised against 200-kDa neurofilament H (clone RT-97). Sympathetic fibers were labeled with an antibody raised against tyrosine hydroxylase. CGRP, RT-97, and tyrosine hydroxylase immunoreactive fibers, but not isolectin B4 positive fibers, were present throughout the bone marrow, mineralized bone and the periosteum. While the periosteum is the most densely innervated tissue, when the total volume of each tissue is considered, the bone marrow receives the greatest total number of sensory and sympathetic fibers followed by mineralized bone and then periosteum. Understanding the sensory and sympathetic innervation of bone should provide a better understanding of the mechanisms that drive bone pain and aid in developing therapeutic strategies for treating skeletal pain. PMID- 12123696 TI - Selective expression of prion protein in peripheral tissues of the adult mouse. AB - The level of expression of normal cellular prion protein, PrP(c) (cellular prion protein), controls both the rate and the route of neuroinvasive infection, from peripheral entry portal to the CNS. Paradoxically, an overview of the distribution of PrP(c) within tissues outside the CNS is lacking. We have used novel antibodies that recognise cellular prion protein in glutaraldehyde-fixed tissue (in order to optimise immunohistochemical labelling of this conformationally labile protein), in combination with in situ hybridisation, to examine the expression of PrP(c) in peripheral tissues of the adult mouse. We found that although prion protein is expressed in many tissues, it is expressed at high levels only in discrete subpopulations of cells. Prominent amongst these are elements of the "hardwired neuroimmune network" that integrate the body's immune defence and neuroendocrine systems under CNS control. These prion protein expressing elements include small diameter afferent nerves in the skin and the lamina propria of the aerodigestive tract, sympathetic ganglia and nerves, antigen presenting and processing cells (both follicular and non-follicular dendritic cells) and sub-populations of lymphocytes particularly in skin, gut- and bronchus-associated lymphoid tissues. Prion protein is also expressed in the parasympathetic and enteric nervous systems, in the dispersed neuroendocrine system, and in peripheral nervous system axons and their associated Schwann cells. This selective expression of cellular prion protein provides a variety of alternative routes for the propagation and transport of prion infection entering from peripheral sites, either naturally (via the aerodigestive tract or abraded skin) or experimentally (by intraperitoneal injection) to the brain. Key regulatory cells that express prion protein, and in particular enteroendocrine cells in the mucosal wall of the gut, and dendritic cells that convey pathogens from epithelial layers to secondary lymphoid organs, may be particularly important in the transmission of infection in the periphery. PMID- 12123697 TI - Dynamics and diversity in interneurons: a model exploration with slowly inactivating potassium currents. AB - Recent experimental and model work indicates that slowly inactivating potassium currents might play critical roles in generating population rhythms. In particular, slow (<1-4 Hz) rhythms recorded in the hippocampus correlate with oscillatory behaviors in interneurons in this frequency range. Limiting the ion channels to the traditional Hodgkin-Huxley sodium and potassium currents, a persistent sodium current, and a slowly inactivating potassium current, we explore the role of slowly inactivating conductances in a multi-compartmental interneuronal model. We find a rich repertoire of tonic and bursting behaviors depending on the distribution, density and kinetics of this conductance. Specifically, burst frequencies of appropriate frequencies could be obtained for certain distributions and kinetics of this conductance. Robust (with respect to injected currents) regimes of tonic firing and bursting behaviors are uncovered. In addition, we find a bistable tonic firing pattern that depends on the slowly inactivating potassium current. Therefore, this work shows ways in which different channel distributions and heterogeneities could produce variable signal outputs. We suggest that an understanding of the dynamical profiles of inhibitory neurons based on the density and distribution of their currents is helpful in dissecting out the complex roles played by this heterogeneous group of cells. PMID- 12123698 TI - NMDA receptor antagonists attenuate noxious and nonnoxious colorectal distention induced Fos expression in the spinal cord and the visceromotor reflex. AB - In the present three-part study, the effects of intrathecally administered N methyl-D-aspartate (NMDA) receptor antagonists on responses to noxious and innocuous colorectal distention (CRD) were examined. In the first part, a passive avoidance paradigm was used to confirm that 80 mm Hg CRD is a noxious stimulus since it produced avoidance behavior. Acquisition of this behavior was blocked by the NMDA receptor antagonist D(-)-2-amino-5-phosphonopetanoic acid (APV, 60 nmol, intrathecal). In contrast, 20 mm Hg CRD is an innocuous stimulus since there was no difference in the behavior of these animals compared to nondistended controls. In the second part, the effects of the NMDA receptor antagonist dizocilpine maleate (MK-801, 0-100 nmol, intrathecal) on CRD-induced Fos expression in the lumbosacral spinal cord were examined. Noxious and innocuous CRD induced 98+/-4 and 50+/-2 Fos labeled cells per section per side of the spinal cord, respectively. MK-801 dose-dependently attenuated noxious CRD-induced Fos. Compared to saline, the peak attenuation was 55%. Innocuous CRD-induced Fos was attenuated by 36% following 100 nmol MK-801. In the third part, the effects of APV (0-240 nmol, intrathecal) on the visceromotor reflex were examined. APV dose dependently attenuated the visceromotor reflex to graded intensities of CRD that went from the innocuous into the noxious range. In separate animals that only received innocuous stimulation, APV dose-dependently attenuated the visceromotor reflex. The magnitude of attenuation was similar for both stimulus paradigms. These data expand upon our previous dorsal horn neuronal recordings which showed that spinal NMDA receptors partially mediate the processing of both noxious and innocuous colorectal stimuli. They further underscore a difference from somatic tissue in the role of NMDA receptors in processing acute or transient visceral stimuli in the absence of tissue injury. PMID- 12123699 TI - Increases in striatal preproenkephalin gene expression are associated with nigrostriatal damage but not L-DOPA-induced dyskinesias in the squirrel monkey. AB - Changes in preproenkephalin expression in the caudate and putamen have been linked to the development of L-3,4-dihydroxyphenylalanine (L-DOPA)-induced dyskinesias in primate models of Parkinson's disease, although not all investigators have been able to confirm this association. Because nigrostriatal damage per se is associated with increases in striatal preproenkephalin mRNA levels, it is difficult to know if changes in transcript levels are a result of lesioning or concurrent L-DOPA treatment and resulting dyskinesias. To circumvent these difficulties, we measured striatal preproenkephalin mRNA levels in monkeys with L-DOPA-induced dyskinesias both with and without lesions of the nigrostriatal system. The latter model is not confounded by morphological and biochemical changes resulting from nigrostriatal damage. Monkeys were gavaged with L-DOPA (15 mg/kg) twice daily for a 2-week period and killed 3 days after treatment. 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) treatment alone resulted in an increase in preproenkephalin mRNA levels as previously shown. However, striatal transcript levels were similarly elevated in dyskinetic MPTP lesioned animals treated with L-DOPA. In unlesioned animals, preproenkephalin mRNA levels were also similar in control and L-DOPA-treated dyskinetic monkeys. Because drug-induced changes in mRNA may not be sustained for a prolonged period after treatment, a second series of experiments were done in which animals were killed 3-4 h after the last dose of L-DOPA, but the results were similar to those obtained after 3 days. These data show that, while elevations in striatal preproenkephalin mRNA levels are associated with nigrostriatal damage, they are not linked to the development of L-DOPA-induced dyskinesias. These results thus question the importance of preproenkephalin mRNA in the pathogenesis of this disabling complication of L-DOPA therapy in Parkinson's disease. PMID- 12123700 TI - Structural and quantitative analysis of astrocytes in the mouse hippocampus. AB - We revealed the structural features of astrocytes by means of light microscopy, confocal laser scanning microscopy and high voltage electron microscopy, and estimated their numerical densities in the mouse hippocampus. The high voltage electron microscope examinations of Golgi-impregnated astrocytes clearly disclosed their fine leaflet-like processes in the masses occupied by individual astrocytes. The intracellular injection of two different fluorescent tracers into two neighboring astrocytes revealed that each astrocyte occupied a discrete area with a limited overlap only at its peripheral portion. In a quantitative analysis using an optical dissector, the numerical densities of astrocytes identified as S100-immunoreactive cells were only slightly different in their areal and laminar distributions. The numerical densities were higher in the stratum lacunosum moleculare and dentate hilus, while they were slightly lower in the principal cell layers than the average (24.2 x 10(3) mm(-3)) in whole hippocampal regions. As for the dorsoventral difference, the numerical densities were significantly larger at the ventral level in the dentate gyrus, whereas such tendency was not apparent in the hippocampus proper. The projection area of the astrocytes estimated from Golgi-impregnated samples was roughly in inverse relation to the numerical densities; the areas in the stratum lacunosum-moleculare were somewhat smaller than the other layers, where the numerical densities were high. The present study indicates that astrocytes are distributed rather evenly without any prominent areal or laminar differences and that the individual astrocytes have their own domains; the periphery of the domain of a given astrocyte is interdigitated intricately with the processes of adjacent astrocytes whereas its inner core portion is not penetrated by them. PMID- 12123703 TI - Emery-Dreifuss muscular dystrophy, nuclear cell signaling and chromatin remodeling. PMID- 12123701 TI - Activity-dependent expression of calbindin in rabbit floccular Purkinje cells modulated by optokinetic stimulation. AB - Optokinetic stimulation activates visual climbing fiber pathways that synapse upon contralateral floccular Purkinje cells. Long-term horizontal optokinetic stimulation causes a progressive decrease in gain of the optokinetic reflex and leads to the subsequent genesis of a prolonged negative optokinetic afternystagmus. Since the flocculus is involved in adaptation to optokinetic stimulation, we used the technique of differential display reverse transcription polymerase chain reaction to explore transcriptional changes in the flocculus evoked by long-term optokinetically evoked climbing fiber discharge. Several differentially transcribed gene products were isolated and sequenced. One of these, calbindin mRNA, was expressed in relatively decreased abundance in the flocculus that received increased climbing fiber input. Decreased transcription of calbindin mRNA was confirmed by northern blots. Hybridization histochemistry was used to localize calbindin mRNA to Purkinje cells and confirmed decreased transcription of calbindin mRNA in Purkinje cells located in folium 1 of the flocculus. Western blots and immunohistochemistry localized the climbing fiber evoked decreased expression of calbindin to Purkinje cells in folia 1 of the flocculus. The expression of four other calcium-binding proteins in the flocculus was not influenced by optokinetic stimulation. Changes in expression of calbindin could be evoked by decreases in intracellular calcium associated with climbing fiber-evoked decreases in Purkinje cell simple spike activity.The application of differential display reverse transcription-polymerase chain reaction has provided a positive screen for several molecules in addition to calbindin whose expression is affected by naturally evoked activity in a major synaptic pathway to the cerebellum. Further experiments will be required to specify the functional role of each of these molecules. PMID- 12123705 TI - Regulating the mammalian genome: the role of nuclear architecture. PMID- 12123704 TI - In situ detection of phospholipid and phosphoinositide metabolism. PMID- 12123706 TI - Co-operation of phosphatidylinositol transfer protein with phosphoinositide 3 kinase gamma in vitro. PMID- 12123707 TI - Gene expression profiling in RAS oncogene-transformed cell lines and in solid tumors using subtractive suppression hybridization and cDNA arrays. PMID- 12123708 TI - Reciprocal regulation: recognition of pattern of gene expression in cancer cells. PMID- 12123709 TI - Detection of apoptosis in tumors using magnetic resonance imaging and spectroscopy. PMID- 12123710 TI - Mitochondria and ceramide: intertwined roles in regulation of apoptosis. PMID- 12123711 TI - The importance of tumor metabolism in cancer prognosis and therapy; pre-clinical studies on rodent tumors with agents that improve tumor oxygenation. PMID- 12123712 TI - Overexpression of protein disulfide isomerase-like protein in a mouse leukemia L1210 cell line selected for resistance to 4-methyl-5-amino-1-formylisoquinoline thiosemicarbazone, a ribonucleotide reductase inhibitor. PMID- 12123713 TI - Structure-based development of cathepsin L inhibitors and therapeutic applications for prevention of cancer metastasis and cancer-induced osteoporosis. PMID- 12123714 TI - The atypical PKC scaffold protein P62 is a novel target for anti-inflammatory and anti-cancer therapies. PMID- 12123715 TI - Inositides in the nucleus: regulation of nuclear PI-PLCbeta1. PMID- 12123716 TI - Regulation of phospholipase C-gamma1 by protein kinase A-dependent phosphorylation. PMID- 12123717 TI - DGK and nuclear signaling nuclear diacylglycerol kinases in IIC9 cells. PMID- 12123718 TI - The functional efficiency of lipogenic and cholesterogenic gene expression in normal mice and in mice lacking the peroxisomal proliferator-activated receptor alpha (PPAR-alpha). PMID- 12123719 TI - Regulation of the activity of the pyruvate dehydrogenase complex. PMID- 12123720 TI - Regulation of the intracellular localization of phosphoinositide-specific phospholipase Cdelta(1). PMID- 12123721 TI - Lysosomal cathepsins: structure, role in antigen processing and presentation, and cancer. PMID- 12123722 TI - Application of 15N to the study of hepatic nitrogen metabolism. PMID- 12123723 TI - Regulation of glycolysis by Raf protein serine/threonine kinases. PMID- 12123725 TI - If at first you don't succeed em leaderfructose utilization by Escherichia coli. PMID- 12123724 TI - Regulation of the HIF pathway: enzymatic hydroxylation of a conserved prolyl residue in hypoxia-inducible factor alpha subunits governs capture by the pVHL E3 ubiquitin ligase complex. PMID- 12123735 TI - Lysophosphatidylcholine as a ligand for immunoregulation. AB - Despite the recognized effects of lysophosphatidylcholine upon cells of the immune system and its association with inflammatory processes, its mechanism of action has remained poorly characterized. Our recent identification of the first lysophosphatidylcholine receptor as an immunoregulatory G protein-coupled receptor named G2A whose genetic ablation results in the development of inflammatory autoimmune disease has, therefore, provided a new perspective on the role of this lysophospholipid as a modulator of immune responses. This commentary discusses the biological properties of lysophosphatidylcholine as an immunoregulatory ligand for cells of the innate and adaptive arms of the immune system. Although we focus primarily on ligand interactions with G2A, we also discuss the issue of possible functional redundancy with other receptors with recently established ligand specificities towards phosphorylcholine-containing lysolipids including lysophosphatidylcholine. PMID- 12123736 TI - Inhibition of heparin synthesis by methotrexate in rats in vivo. AB - The content and synthesis of heparin and mast cell-dependent skin oedema (as an indirect evaluation of histamine and serotonin content) were investigated in the rat skin after chronic treatment with compound 48/80, a mast cell degranulating substance. The effect of methotrexate, a folic acid analogue that interrupts the synthesis of DNA and RNA, on heparin synthesis and amine storage also was evaluated in rat skin. The heparin content at 6 and 240 hr after treatment with compound 48/80 was reduced markedly (86 and 64%, respectively). At 6 hr, heparin synthesis increased 3.1-fold compared with control animals; maximal synthesis occurred at 24 hr post-treatment (12.8-fold increase), decaying at 240 hr (2.4 fold increase). The dermatan sulfate content and synthesis were not affected by treatment with compound 48/80. Autoradiographic analysis revealed that methotrexate (2.5mg/kg for 3 consecutive days) abolished heparin synthesis at 6, 24, and 72 hr after compound 48/80 treatment, without affecting dermatan sulfate synthesis. The oedema induced by intradermal injection of compound 48/80 (1 microg/site) into the rat skin was decreased significantly at 6 hr after chronic treatment with this compound, but was restored completely 72 hr post-treatment. This pattern of oedematogenic response was also observed in the methotrexate treated rats. In conclusion, our results show that methotrexate suppresses heparin synthesis without affecting the synthesis of either dermatan sulfate or the co-stored amines histamine/serotonin (as evaluated by measuring the mast cell dependent oedema), suggesting that the enzyme system involved in heparin synthesis is inducible. PMID- 12123738 TI - Renal organic cation and nucleoside transport. AB - We previously reported that the rat organic cation transporter rOCT1 could transport the nucleoside analog deoxytubercidin (dTub) (Chen R, Nelson JA. Biochem Pharmacol 2000;60:215-9). The cationic form of dTub (dTub(+)) appeared to be the true substrate of rOCT1. We also reported that although rOCT2 is similar to rOCT1, it does not transport dTub at pH 7.4. In this study, we measured the K(m) and V(max) values of dTub(+) uptake at a reduced pH (pH 5.4) for both rOCT1 and rOCT2. The difference in substrate activity appears due, in large part, to a poor affinity of rOCT2 for dTub(+). The transport efficiency estimated by V(max)/K(m) values for rOCT2 was only 6% that of rOCT1. Chimeras constructed between rOCT1 and rOCT2 revealed that the difference in dTub binding lies within transmembrane domains 2-7. To evaluate the potential of OCT1 in the renal secretion of dTub, tissue distribution and urinary excretion of dTub in OCT1 knockout mice were measured. No significant difference was observed in renal elimination, plasma level, and tissue distribution of dTub between the knockout and the wild-type mice. Therefore, dTub is a good substrate for OCT1; however, OCT1 does not appear to be necessary for its renal secretion. PMID- 12123737 TI - Induction of cyclooxygenase-2 by staurosporine through the activation of nuclear factor for IL-6 (NF-IL6) and activator protein 2 (AP2) in an osteoblast-like cell line. AB - The induction of cyclooxygenase-2 (COX-2) plays a crucial role in many physiological and pathological processes. The expression of the COX-2 gene is regulated by many extracellular stimuli, including growth factors, cytokines, and tumor promoters. Staurosporine, a potential anti-tumor drug, was found recently to up-regulate the expression of the COX-2 gene in the mouse osteoblast-like cell line MC3T3-E1. The ability of staurosporine to induce the expression of the COX-2 gene was investigated using luciferase reporters controlled by various COX-2 core promoter regions. Two cis-acting sites for activator protein 2 (AP2) and nuclear factor for IL-6 (NF-IL6), respectively, were identified as responsible for the staurosporine-mediated COX-2 up-regulation. Mutational analysis further verified that both NF-IL6 and AP2 are involved in this process. Further studies showed the stimulatory effect of staurosporine on luciferase activity to be both time- and concentration-dependent. Luciferase activity could be induced at as low as 5 nM staurosporine and reached a maximum at around 20 nM. At 50 nM, the stimulatory effect of staurosporine on luciferase activity reached a maximum at about 8 hr and fell rapidly following 10 hr of incubation. Interestingly, a selective protein kinase C inhibitor, 2-[1-(3-dimethylaminopropyl)indol-3-yl]-3-(indol-3 yl) maleimide (GF109203X), failed to stimulate luciferase activity under the same conditions. This finding implies that staurosporine-mediated COX-2 gene expression is specific and independent of protein kinase C activity. PMID- 12123739 TI - Mechanism of action of non-cisplatin type DNA-targeted platinum anticancer agents: DNA interactions of novel acridinylthioureas and their platinum conjugates. AB - The DNA binding of two novel acridinylthioureas, ACR-NH-(CH(2))(2)-C(S)-NHCH(3) (1) and ACR-N(CH(3))-C(S)-NHCH(3) (3), and their platinum conjugates 4 and 5 derived from [PtCl(2)(en)]-was studied in cell-free model systems using various physico-chemical and biophysical methods. These included: spectrophotometric drug DNA titrations, ethidium-DNA fluorescence quenching, competitive drug displacement, high-resolution NMR spectroscopy, and unwinding of plasmid DNA monitored by agarose gel electrophoresis. The acridinium cation of 1 showed strong binding to native DNA with K(i)=1.5 x 10(6)M(-1) and an excluded site size (n) of 2bp (McGhee-von Hippel fits of absorbance data). Compound 3 showed no measurable association with DNA. Binding of 1 was an order of magnitude stronger than that of simple 9-methylaminoacridine (2). In alternating copolymers, 1 exhibited slight AT preference. In poly(dA-dT)(2), enhanced association was accompanied by an increased binding site (approximately 3bp), while parameters in poly(dG-dC)(2) were consistent with classical intercalation. Displacement of 1 by distamycin from calf thymus DNA was suggestive of non-intercalating thiourea in 1 being located in the minor groove of the duplex. 1H NMR data of d(GGAGCTCC)(2) modified with 1 indicated intercalative binding of planar acridine, based on upfield shifts of aromatic proton signals relative to those in unbound 1 (Deltadelta approximately equal to -0.5 to -1ppm). Finally, 4 and 5 were found to unwind negatively supercoiled pUC19 plasmid by 21 degrees and 7 degrees per adduct, respectively (electrophoretic gel mobility assays). The difference in DNA binding modes of 4 and 5 is discussed as the ultimate source of the distinctly different biological activities of the conjugates. PMID- 12123740 TI - Inhibition of phosphate transport in rat heart mitochondria by 3'-azido-3' deoxythymidine due to stimulation of superoxide anion mitochondrial production. AB - In order to gain some insight into the mechanism by which 3'-azido-3' deoxythymidine (AZT) damages mitochondria, we investigated whether externally added AZT can stimulate reactive oxygen species (ROS) production by rat heart mitochondria (RHM). An increase in superoxide anion ((O(2)(.-)) production was measured in RHM added with AZT, by using a photometrically method which allows an early O(2)(.-) detection by following the absorbance increase at 550 nm due to the ferricytochrome c reduction. Such an increase was found to be prevented from externally added superoxide dismutase. The stimulation of O(2)(.-) mitochondrial production induced by AZT was found to occur under conditions in which mitochondrial oxygen consumption was prevented by both inhibitors of electron flow and ATP synthesis. Since ROS can cause mitochondrial carrier impairment, we investigated whether AZT can affect mitochondrial permeability in virtue of its capability to stimulate ROS production. In this regard, we studied the transport of phosphate (P(i)), by measuring the mitochondrial shrinkage that takes place as a result of P(i) uptake by RHM previously swollen in a calcium acetate medium. As a result of the AZT-dependent O(2)(.-) production, uncompetitive inhibition of the rate of P(i) transport in RHM was found (K(i) of about 10 microM), consistently, such an inhibition was found to prevent by certain known ROS scavengers, i.e. superoxide dismutase, the antioxidant Vitamin C and reduced gluthatione. PMID- 12123741 TI - Enhanced resistance of HeLa cells to cisplatin by overexpression of gamma glutamyltransferase. AB - Gamma-glutamyltransferase (GGT), which is a key enzyme for the cellular glutathione (GSH) homeostasis, was shown to be overexpressed in human tumor cells selected for resistance to cisplatin and to influence the resistance of experimental tumors in vivo. We first established that cisplatin treatment of HeLa cells was accompanied by an early 3-fold induction of GGT synthesis, enhancing the possibility that this enzyme plays an important role in the cell defenses against this anticancer drug. This role was then studied using a GGT transfected HeLa cell line (HeLa-GGT) exhibiting 10 times the activity of the parental HeLa cells (120-150 and 10-14 mU/mg protein, respectively). Both cell lines showed comparable intracellular GSH levels and cisplatin resistance when cultured in high (250 microM) or low (50 microM) cysteine-containing medium. When 50 microM of GSH were included in the low-cysteine culture medium only HeLa-GGT cells partially recovered their intracellular GSH and exhibited an increased resistance to cisplatin. Cisplatin treatment also inhibited GGT-dependent production of reactive oxygen species, a process depending on the availability of cysteinylglycine produced during GSH catabolism. Furthermore, we showed that cisplatin forms adducts with cysteinylglycine 10 times more rapidly than with GSH, and that these adducts were formed only in the extracellular medium of HeLa GGT cells. This extracellular mechanism could at least partially account for the increased resistance of GGT-rich cells to cisplatin. PMID- 12123743 TI - Inhibition of matrix-proteases by polyphenols: chemical insights for anti inflammatory and anti-invasion drug design. AB - Flavanols--a class of plant polyphenols abundant in tea leaves and grape seeds and skins--have been found to inhibit some matrix-proteases instrumental in inflammation and cancer invasion, such as leukocyte elastase (LE) and gelatinases. In order to establish the relationship between chemical structure and activity, 27 different flavonoids (antocyanidins, dihydrochalcones, dihydroflavonols, flavanolignans, flavanols, flavones, flavonols and isoflavones) and other compounds with anti-oxidant properties were evaluated for their potential in blocking LE and gelatinase activities. LE activity was measured using a chromogenic substrate: from comparison of the different levels of inhibition, it was deduced that a crucial role in inhibition might be played by a galloyl moiety or hydroxyl group at C3, three hydroxyl groups at B ring, one hydroxyl group at C4', and a 2,3-double bond. Gelatinase activity was measured using the gelatin-zymography assay, and its inhibition showed that three hydroxyl groups at the A or B ring, or, for non-planar molecules, a galloyl moiety at C3 could be determinant. This comparative study is proposed as a basis for designing new molecules with enhanced anti-proteolytic activities, and no or reduced side effects, for use in hindering inflammation, cancer invasion and angiogenesis. PMID- 12123742 TI - Effects of polysulfated glycosaminoglycan and triamcinolone acetonid on the production of proteinases and their inhibitors by IL-1alpha treated articular chondrocytes. AB - In this study we determined the in vitro effects of polysulfated glycosaminoglycan (PSGAG) and the glucocorticoid triamcinolone acetonid (TA) on the IL-1 altered expression and activity of matrix metalloproteinases (MMP-1, MMP 3), tissue inhibitor of metalloproteinases-1, the plasminogen activators tPA and uPA and plasminogen activator inhibitor 1 by articular chondrocytes. Bovine chondrocytes were cultured in alginate gel beads. Cells were treated with interleukin-1alpha (IL-1alpha) in the presence of vehicle or drugs at various concentrations. After 48hr mRNA expression of MMP-1, MMP-3, TIMP-1, uPA, tPA and PAI-1 was analyzed by RT-PCR-ELISA. The protein synthesis of TIMP-1 and MMP-3 was determined by immunoprecipitation, PAI-1 protein was quantitated by ELISA. The activity of enzymes and inhibitors was measured by functional assays. Treating chondrocytes with IL-1 induced the expression of MMPs and downregulated TIMP-1 but stimulated both the expression of PAs and PAI-1. Both drugs significantly reduced collagenase and proteoglycanase activities which was accompanied by inhibition of the expression of MMP-1 and MMP-3. The IL-1 decreased expression of TIMP-1 was further reduced by TA, which resulted in a significant loss of TIMP activity. No effects on TIMP activity or TIMP-1 biosynthesis were observed after treatment of chondrocytes with PSGAG. Both drugs inhibited the IL-1-induced mRNA expression of tPA, whereas expression of uPA was only mildly reduced by PSGAG, which also induced PAI-1 above IL-1 stimulated levels. As inhibition of collagenase activities and tPA expression by PSGAG occurred at physiological concentrations it might be of clinical relevance, indicating that PSGAG could help reducing cartilage degradation and has a strong anti-fibrinolytic potential. Due to their co-regulation of MMPs and TIMP(s) glucocorticoids should be carefully studied for their overall effect on extracellular matrix proteolysis. PMID- 12123744 TI - 3'-Azido-2',3'-dideoxythymidine induced deficiency of thymidine kinases 1, 2 and deoxycytidine kinase in H9 T-lymphoid cells. AB - Continuous cultivation of T-lymphoid H9 cells in the presence of 3'-azido-2',3' dideoxythymidine (AZT) resulted in a cell variant cross-resistant to both thymidine and deoxycytidine analogs. Cytotoxic effects of AZT, 2',3'-didehydro-3' deoxythymidine as well as different deoxycytidine analogs such as 2',3' dideoxycytidine, 2',2'-difluoro-2'-deoxycytidine (dFdC) and 1-ss-D arabinofuranosylcytosine (Ara-C) were strongly reduced in H9 cells continuously exposed to AZT when compared to parental cells (>8.3-, >6.6-, >9.1-, 5 x 10(4)-, 5 x 10(3)-fold, respectively). Moreover, anti-HIV-1 effects of AZT, d4T, ddC and 2',3'-dideoxy-3'-thiacytidine (3TC) were significantly diminished (>222-, >25-, >400-, >200-fold, respectively) in AZT-resistant H9 cells. Study of cellular mechanisms responsible for cross-resistance to pyrimidine analogs in AZT resistant H9 cells revealed decreased mRNA levels of thymidine kinase 1 (TK1) and lack of deoxycytidine kinase (dCK) mRNA expression. The loss of dCK gene expression was confirmed by western blot analysis of dCK protein as well as dCK enzyme activity assay. Moreover, enzyme activity of TK1 and TK2 was reduced in AZT-resistant cells. In order to determine whether lack of dCK affected the formation of the active triphosphate of the deoxycytidine analog dFdC, dFdCTP accumulation and retention was measured in H9 parental and AZT-resistant cells after exposure to 1 and 10 microM dFdC. Parental H9 cells accumulated about 30 and 100 pmol dFdCTP/10(6) cells after 4hr, whereas in AZT-resistant cells no dFdCTP accumulation was detected. These results demonstrate that continuous treatment of H9 cells in the presence of AZT selected for a thymidine analog resistant cell variant with cross-resistance to deoxycytidine analogs, due to deficiency in TK1, TK2, and dCK. PMID- 12123745 TI - Relationship between the activation of heat shock factor and the suppression of nuclear factor-kappaB activity in rat hepatocyte cultures treated with cyclosporine A. AB - We investigated on primary cultures of rat hepatocytes the effect of cyclosporine A (CsA) on the activation of nuclear factor-kappaB (NF-kappaB), activator protein 1 (AP-1), and heat shock factor 1 (HSF1), three transcription factors involved in cellular response pathways. Hepatocytes were subjected to a time-course (1, 3, 6, and 22 hr) incubation and CsA treatment in the range 1-50 microM. NF-kappaB, AP 1, and HSF1 binding activities were established through electrophoretic mobility shift assay. Levels of HSP70 mRNA and protein were measured by Northern and Western blot analysis respectively. In cells incubated for 1 and 3 hr, electrophoretic mobility shift assay experiments showed a dose-dependent increase of the NF-kappaB binding activity; while following 22hr of incubation, a suppression of the positive effect of CsA at shorter times was detected. At all periods of incubation assayed, CsA induced the activation of AP-1 which was detected by DNA-binding activity of this transcription factor. A dose-dependent activation of HSF1 was observed at 22 hr of incubation. We conclude that in rat hepatocyte cultures, CsA induces the transcriptional activation of NF-kappaB, AP 1, and HSF1. However, the time point at which activation of each transcription factor occurs is different. Thus, at 22 hr of incubation, the CsA-induced activation of HSF1 is accompanied by the reduction of the positive effect of CsA on NF-kappaB activation at earlier time points. PMID- 12123746 TI - Identification of two C-terminal amino acids, Ser(355) and Thr(357), required for short-term homologous desensitization of mu-opioid receptors. AB - Our recent study suggests that a cluster of Ser/Thr residues (T(354)S(355)S(356)T(357)) at the intracellular carboxyl tail of rat mu-opioid receptor (MOR1) is required for the development of short-term homologous desensitization. To investigate the functional role played by individual serine or threonine residue of this (TSST) cluster in the agonist-induced mu-opioid receptor desensitization, point mutant (T354A), (S355A), (S356A) and (T357A) mu opioid receptors were prepared and stably expressed in human embryonic kidney 293 cells (HEK 293 cells). Similar to wild-type mu-opioid receptors, mutant (T354A) and (S356A) mu-opioid receptors stably expressed in HEK 293 cells developed homologous desensitization after 30 min pretreatment of DAMGO ([D-Ala(2),N-methyl Phe(4),Gly-ol(5)]enkephalin), a specific mu-opioid receptor agonist. Substituting Ser(355)or Thr(357) with alanine resulted in a significant attenuation of agonist induced mu-opioid receptor desensitization. In HEK 293 cells stably expressing double mutant (S355A/T357A) mu-opioid receptors, DAMGO pretreatment failed to significantly affect the efficacy and potency by which DAMGO inhibits forskolin stimulated adenylyl cyclase activity. Consistent with the general belief that agonist-induced phosphorylation of guanine nucleotide binding protein (G protein) coupled receptors is involved in homologous desensitization. Treating HEK 293 cells expressing wild-type mu-opioid receptors with 5 microM DAMGO for 30 min induced the receptor phosphorylation. Mutation of Ser(355) and Thr(357) also greatly impaired DAMGO-induced mu-opioid receptor phosphorylation. These results suggest that two C-terminal amino acids, Ser(355) and Thr(357), are required for short-term homologous desensitization and agonist-induced phosphorylation of mu opioid receptors expressed in HEK 293 cells. PMID- 12123747 TI - Novel mechanism of Vitamin E protection against cyclosporine A cytotoxicity in cultured rat hepatocytes. AB - Cyclosporine A (CsA) is the immunosuppressor most frequently used in transplant surgery and in the treatment of autoimmune diseases. It has been shown that CsA is able to generate reactive oxygen species and lipid peroxidation which are directly involved in the CsA hepatotoxicity. As antioxidant, Vitamin E (VitE) has been used to diminish the toxicity of CsA in vitro. Besides its direct action as the classical antioxidant implicated in preventing lipid peroxidation, we decided to investigate the effect of VitE on the endogenous antioxidant defense system, such as Mn and CuZn superoxide dismutase (MnSOD, CuZnSOD) catalase and glutathione peroxidase (GPx) on CsA cytotoxicity in primary cultures of rat hepatocytes. In cells incubated in the presence of CsA, there was an increase in the expression and activity of MnSOD and CuZnSOD but not in that of catalase and GPx. However, when hepatocytes were coincubated with CsA and VitE, an increase in the expression and activity in all antioxidant enzymes (MnSOD, CuZnSOD, catalase and GPx) was observed. In conclusion, we suggest (a) that the imbalance between SOD and catalase/GPx by the effect of CsA is the main mechanism responsible for peroxide accumulation and cell death in hepatocytes, and (b) that the presence of VitE in culture media reduces the oxidative stress through the inhibition of lipid peroxidation, but also through the increase of the expression and activity of catalase and GPx which allows the restoration of SOD and catalase/GPx coordination, indispensable for the correct cell defense against ROS. PMID- 12123748 TI - Modulation of L-arginine transport and nitric oxide production by gabexate mesylate. AB - Gabexate mesylate, a non-antigenic synthetic inhibitor of trypsin-like serine proteinases, is a drug used efficiently in the treatment of pancreatitis and disseminated intravascular coagulation and as a regional anticoagulant for haemodialysis. Considering the structural similarity between L-arginine and gabexate mesylate, the effect of this drug on L-arginine transport, nitric oxide (NO) formation and constitutive NO synthase activity in human platelets was investigated. Data have shown that gabexate mesylate inhibited competitively L arginine uptake by increasing the K(m) value from 22+/-2 to 86+/-6 microM. The K(i) value was 158 microM at pH 7.4 and 37 degrees. Furthermore, gabexate mesylate decreased dose and time-dependent nitrite and nitrate formation (NO(x) release) and cGMP accumulation in whole cells. In addition, gabexate mesylate inhibited constitutive nitric oxide synthase in a cell-free extract. We concluded that gabexate mesylate could be considered an effective modulator of cellular NO synthesis. PMID- 12123749 TI - Hyperekplexia mutation of glycine receptors: decreased gating efficacy with altered binding thermodynamics. AB - [(3)H]Strychnine binding was studied to recombinant human alpha(1) and the hyperekplexia mutant alpha(1)R271L glycine receptors (GlyRs) transiently expressed in human embryonic kidney (HEK)-293 cell cultures at 0, 18 and 37 degrees. The alpha(1)R271L mutation did not affect the linear van't Hoff plots of the exothermic binding of the antagonist [3H]strychnine while it turned taurine into an antagonist with exothermic binding. The inhibition constants of the agonist glycine showed opposite temperature dependence on alpha(1) GlyRs, corresponding to endothermic binding driven by large entropic increases. The temperature dependence of displacement by the partial agonists taurine on alpha(1) GlyRs and glycine on alpha(1)R271L GlyRs was biphasic reflecting negative heat capacity changes, dehydration changes and/or a complex binding mechanism. The thermodynamic discrimination of efficacy is valid for native rat spinal and recombinant human GlyRs. The alpha(1)R271L mutation impairs the transduction mechanism and distorts gating of GlyRs. Thereby it reduces the potency and efficacy of agonists and affects their thermodynamic parameters of binding. The hyperekplexia mutation offers a model system to demonstrate the correlation among pathophysiology, gating efficacy and binding thermodynamics of GlyRs. PMID- 12123750 TI - Covalent binding of the anticancer drug ellipticine to DNA in V79 cells transfected with human cytochrome P450 enzymes. AB - Ellipticine is a potent antineoplastic agent whose mechanism of action is considered to be based mainly on DNA intercalation and/or inhibition of topoisomerase II. Recently, we found that ellipticine also forms covalent DNA adducts and that the formation of the major adduct is dependent on the activation of ellipticine by cytochrome P450 (CYP). We examined a panel of genetically engineered V79 cell lines including the parental line V79MZ and recombinant cells expressing the human CYP enzymes CYP1A1, CYP1A2 or CYP3A4 for their ability to activate ellipticine. The extent of activation was determined by analysing DNA adducts by 32P-postlabelling. Ellipticine was found to be toxic to all V79 cell lines with IC(50) values ranging from 0.25 to 0.40 microM. The nuclease P1 version of the 32P-postlabelling assay yielded a similar pattern of ellipticine DNA adducts with two major adducts in all cells, the formation of only one of which was dependent on CYP activity. This pattern is identical to that detected in DNA reacted with ellipticine and the reconstituted CYP enzyme system in vitro as confirmed by HPLC of the isolated adducts. Total adduct levels ranged from 2 to 337 adducts per 10(8) nucleotides, in the parental line and in V79 expressing CYP3A4, respectively. As in vitro, human CYP1A2 and CYP1A1 were less active. The results presented here are the first report showing the formation of CYP-mediated covalent DNA adducts by ellipticine in cells in culture, and confirm the formation of covalent DNA adducts as a new mechanism of ellipticine action. PMID- 12123752 TI - Activation of the gamma-glutamyltransferase promoter 2 in the rat colon carcinoma cell line CC531 by histone deacetylase inhibitors is mediated through the Sp1 binding motif. AB - The single-copy gene for rat gamma-glutamyltransferase (GGT) encodes at least seven distinct mRNAs that differ in their 5'-untranslated regions only. Tissue- and developmental-specific expression of GGT is partly achieved by the presence of many transcription factor-binding sites in the promoters of this gene. In an earlier study we found that GGT mRNAs II and IV levels were increased upon butyrate-induced differentiation of the rat colon carcinoma cell line CC531. The mechanism for this butyrate-induced upregulation remains unknown, but may result from altered promoter activity as butyrate is a known histone deacetylase inhibitor. In the present study, we show by transient transfection studies that butyrate enhanced the expression of the luciferase reporter gene driven by the rat GGT promoter 2 (P2). Trichostatine A (TSA), another histone deacetylase inhibitor, also enhanced transcription from this promoter. The role of the transcription factor site Sp1 in butyrate- or TSA-induced activation of the GGT P2 was examined as Sp1 has been previously shown to play a central role in the transcriptional activation of other genes during butyrate and TSA stimulation. A triple sequence-motif of this isolated Sp1 site linked to a minimal promoter was able to mediate butyrate- and TSA-induced expression of the luciferase reporter gene, while no effect was measured using the minimal promoter alone. Deleting the Sp1 site in the context of the rat GGT P2 strongly reduced the basal transcription activity and abrogated butyrate- and TSA-induced activation of the mutated promoter. These results suggest that butyrate- or TSA-induced activation of the rat GGT P2 can be mediated by a Sp1 binding motif. PMID- 12123751 TI - Carbonyl reduction of the potential cytostatic drugs benfluron and 3,9 dimethoxybenfluron in human in vitro. AB - Benfluron (B, [5-(2-N-oxo-2-N',N"-dimethylaminoethoxy)-7-oxo-7H benzo[c]fluorene]) is a potential benzo[c]fluorene antineoplastic agent with high activity against a broad spectrum of experimental tumors in vitro and in vivo. The structure of B has been modified to repress its rapid deactivation through carbonyl reduction on C7. 3,9-Dimethoxybenfluron (D, [3,9-dimethoxy-5-(2-N-oxo-2 N',N"-dimethylaminoethoxy)-7-oxo-7H-benzo[c]fluorene]) is one of the B derivatives developed. The present paper was designed to compare the C7 carbonyl reduction of B and D in microsomes, cytosol and hepatocytes from human liver. Two purified human enzymes, microsomal 11beta-hydroxysteroid dehydrogenase 1 (11beta HSD 1) and cytosolic carbonyl reductase, were tested if they are responsible for B and D carbonyl reduction in the respective fractions. Indeed, carbonyl reduction of D in comparison to that of B was 4 and 6-10 times less extensive in human liver microsomes and cytosol, respectively. Moreover, about 10-20 times higher amounts of dihydro B than dihydro D were detected in primary culture of human hepatocytes. 11beta-HSD 1 was shown to be able to reduce B and D. For this enzyme, about 10 times higher rates of carbonyl reduction were observed for B than for D. Likewise, CR participates in B and D carbonyl reduction, although smaller amounts of both reduced metabolites were detected. In summary, carbonyl reduction of D was significantly less extensive than that of B in all in vitro experiments. This lower rate of D inactivation was especially pronounced in hepatocytes which represent a close to in vivo situation. Our results clearly demonstrate that dimethoxy substitution protects the carbonyl group of the benzo[c]fluorene moiety against the deactivation by microsomal and cytosolic reductases. Detailed knowledge on the participating enzymes may serve as a basis for the co-application of specific inhibitors in chemotherapy to further improve the pharmacokinetics of benzo[c]fluorene derivatives. PMID- 12123753 TI - Induction of interleukin 8 release from the HMC-1 mast cell line: synergy between stem cell factor and activators of the adenosine A(2b) receptor. AB - The HMC-1 mast cell line has both adenosine A(3) and A(2b) receptors on its surface, but only agonists of the A(2b) receptor are effective at releasing interleukin 8. Object of this study was to look for co-factors for adenosine A(2b) receptor activation. There was a powerful and statistically significant synergy for release of IL-8, both at the mRNA level (measured after 4 hr) and protein level (measured after 24 hr), between adenosine A(2b) receptor agonists and stem cell factor (SCF). Suitable concentrations for showing synergy were 100 ng/mL SCF and 3 microM 5'-N-ethylcarboxamidoadenosine (NECA). At these concentrations, the IL-8 released into the culture medium after SCF and NECA together was typically 3-5-fold greater in amount than the sum of the amounts of IL-8 released after exposure to the same concentrations of NECA and SCF separately. Since mast cells may be exposed to both adenosine and stem cell factor in the diseased lung, the synergy observed in this model system may have implications for asthma. PMID- 12123754 TI - GPI 6150, a PARP inhibitor, reduces the colon injury caused by dinitrobenzene sulfonic acid in the rat. AB - Poly (ADP-ribose) polymerase, a nuclear enzyme activated by DNA strand breaks, has been shown to play an important role in the pathogenesis of inflammatory bowel disease. Here we investigate the effects of 1,11b-dihydro-[2H]benzopyrano [4,3,2-de]isoquinolin-3-one (GPI 6150), a new poly (ADP-ribose) polymerase inhibitor, in animal models of experimental colitis. Colitis was induced in rats by intra-colonic instillation of dinitrobenzene sulfonic acid. Rats experienced hemorrhagic diarrhea and weight loss. At 4 days after administration of dinitrobenzensulfonic acid, the mucosa of the colon exhibited large areas of necrosis. Neutrophil infiltration (determined by histology and an increase in myeloperoxidase activity in the mucosa) was associated with up-regulation of ICAM 1. Immunohistochemistry for poly (ADP-ribose) showed an intense staining in the inflamed colon. GPI 6150 (20 or 40 mg/kg daily, i.p.) significantly reduced the degree of hemorrhagic diarrhea and weight loss caused by administration of dinitrobenzensulfonic acid. GPI 6150 also caused a substantial reduction of (i) the degree of colon injury, (ii) the rise in myeloperoxidase activity (mucosa), (iii) the increase in the tissue levels of malondialdehyde, (iv) the increase in staining (immunohistochemistry) for poly (ADP-ribose), as well as (v) the upregulation of ICAM-1 and P-selectin caused by dinitrobenzensulfonic acid in the colon. Thus, GPI 6150 reduces the degree of colitis caused by dinitrobenzensulfonic acid. We propose that GPI 6150 may be useful in the treatment of inflammatory bowel disease. PMID- 12123756 TI - Submissions, impact factor, reviewer's recommendations and geographical bias within the peer review system (1997-2002): focus on Germany. PMID- 12123755 TI - Indomethacin-induced mitochondrial dysfunction and oxidative stress in villus enterocytes. AB - Nonsteroidal anti-inflammatory drugs (NSAIDs) are known to cause small intestinal damage but the pathogenesis of this toxicity is not well established. Intestinal epithelial cells are thought to be affected by these drugs in the course of their absorption. These cells are of different types, viz. villus, middle and crypt cells. There is little information on which of these cells, if any, are particularly vulnerable to the effects of NSAIDs. This paper aimed to study the effects of indomethacin, an NSAID commonly used in toxicity studies, on different populations of enterocytes. Effects of the drug were assessed in terms of oxidative damage, mitotic activity, mitochondrial function and lipid composition in enterocytes isolated from the small intestine of rats that had been orally administered indomethacin. In addition, the effects of arginine and zinc in protecting against such changes were assessed. Cell viability, tetrazolium dye (MTT) reduction and oxygen uptake were significantly reduced in villus tip cells from rats dosed with the drug. Thymidine uptake was higher in the crypt cell fraction from these rats. Similarly, products of lipid peroxidation were elevated in the villus tip cells with a corresponding decrease in the level of the anti oxidant, alpha-tocopherol. In isolated mitochondrial preparations from various enterocyte fractions, significant functional impairment and altered lipid composition were seen mainly in mitochondria from villus cells. Arginine and zinc pre-treatment were found to protect against these effects. These results suggest for the first time that the villus tip cells are more vulnerable to the damaging effects of indomethacin and that oxidative stress is possibly involved in this damage. PMID- 12123757 TI - Defibrillation and cardioversion. PMID- 12123759 TI - Pharmacological rescue of mutant ion channels. PMID- 12123760 TI - Is K(v) channel inhibition a common path to hypoxic pulmonary vasoconstriction? PMID- 12123761 TI - An odyssee of figures or the importance of proofs. PMID- 12123763 TI - Extracellular superoxide dismutase and cardiovascular disease. AB - Excessive production and/or inadequate removal of reactive oxygen species, especially superoxide anion (O(2)(*-)), have been implicated in the pathogenesis of many cardiovascular diseases, including atherosclerosis, hypertension, diabetes, and in endothelial dysfunction by decreasing nitric oxide (NO) bioactivity. Since the vascular levels of O(2)(*-) are regulated by the superoxide dismutase (SOD) enzymes, a role of SOD in the cardiovascular disease is of substantial interest. Particularly, a major form of SOD in the vessel wall is the extracellular SOD (ecSOD). This review will discuss the characteristics of ecSOD and the role of ecSOD in cardiovascular diseases. PMID- 12123764 TI - Vasoprotection by nitric oxide: mechanisms and therapeutic potential. AB - Endothelial production of nitric oxide (nitrogen monoxide, NO) has become a major research area in vascular biology. Some of the most important effects that NO exerts in the vascular wall are potentially vasoprotective, because these effects maintain important physiological functions such as vasodilation, anticoagulation, leucocyte adhesion, smooth muscle proliferation, and the antioxidative capacity. During the last 2 decades it has become apparent that a variety of diseases are associated with an impairment of endothelium-dependent NO activity. One of the major causes is believed to be an increased production of reactive oxygen species, in particular superoxide, which have been shown to interfere with many steps of the NO--cyclic guanosine monophosphate (cGMP) pathway. This phenomenon has been found in diverse conditions such as atherosclerosis, hypertension, diabetes, hypercholesterolemia, heart failure, and cigarette smoking. The aim of this review is to examine the cellular and molecular mechanisms whereby NO exerts potentially vasoprotective effects and to discuss pharmacologic approaches targeting the NO pathway in view of their potential to improve endothelial function and to reduce the progression of atherosclerotic vascular disease. We conclude that there is compelling evidence for vasoprotective actions of NO which are mediated by cGMP-dependent and cGMP-independent mechanisms. These effects may contribute to the beneficial effects of established drugs such as ACE inhibitors or statins. Unfortunately, clinical data on the effect of long-term treatment with nitrates on the progression of coronary artery disease are lacking. Finally, L-arginine or new activators of the NO pathway may become therapeutic options in the future. PMID- 12123765 TI - The pivotal role of lipoprotein lipase in atherosclerosis. PMID- 12123767 TI - A novel SCN5A arrhythmia mutation, M1766L, with expression defect rescued by mexiletine. AB - OBJECTIVE: Mutations in the cardiac sodium channel gene, SCN5A, cause congenital long QT syndrome (LQT3), Brugada syndrome, idiopathic ventricular fibrillation, and conduction disease by distinct cellular and clinical electrophysiological phenotypes. METHODS: Postmortem molecular analysis of SCN5A was conducted on an infant who presented shortly after birth with self-terminating torsades de pointes. The infant was treated with lidocaine, propranolol, and mexiletine and was stable for 16 months manifesting only a prolonged QT interval. The infant collapsed suddenly following presumed viral gastroenteritis, was found in 2:1 AV block, and was subsequently declared brain dead. Genomic DNA was subjected to SCN5A mutational analyses and DNA sequencing revealing a novel, spontaneous germline missense mutation, M1766L. The M1766L mutation was engineered into the hH1a clone by site-directed mutagenesis, transfected into embryonic kidney cells (HEK-293), and studied by voltage clamp. RESULTS: The M1766L mutation caused a significant decrease in the sodium channel expression. Co-expression with beta1 subunit, incubation at low temperature, and most effectively incubation with mexiletine partially 'rescued' the defective expression. In addition to this pronounced loss of function, M1766L also showed a 10-fold increase in the persistent late sodium current. CONCLUSIONS: These findings suggest that M1766L SCN5A channel dysfunction may contribute to the basis of lethal arrhythmias, displays an overlapping electrophysiological phenotype, and represents the first sodium channelopathy rescued by drug. PMID- 12123766 TI - Endomyocardial nitric oxide synthase and the hemodynamic phenotypes of human dilated cardiomyopathy and of athlete's heart. AB - OBJECTIVE: In dilated cardiomyopathy and in athlete's heart, progressive LV dilatation is accompanied by rightward displacement of the diastolic LV pressure volume relation. In dilated cardiomyopathy, an increase in diastolic LV stiffness can limit this rightward displacement thereby decreasing LV systolic performance. Because nitric oxide (NO) reduces diastolic LV stiffness, the present study relates diastolic LV stiffness and LV systolic performance to intensity of endomyocardial NO synthase (NOS) gene expression in dilated cardiomyopathy and in athlete's heart. METHODS: Microtip LV pressures, conductance-catheter or angiographic LV volumes, echocardiographic LV wall thicknesses and snap-frozen LV endomyocardial biopsies were obtained in 33 patients with dilated cardiomyopathy and in three professional cyclists referred for sustained ventricular tachycardia. Intensity of LV endomyocardial inducible NOS (NOS2) and constitutive NOS (NOS3) gene expression was determined using quantitative reverse transcription-polymerase chain reaction (RT-PCR). RESULTS: Dilated cardiomyopathy patients with higher diastolic LV stiffness-modulus and lower LV stroke work had lower NOS2 and NOS3 gene expression at any given level of LV end-diastolic wall stress. The intensity of NOS2 and NOS3 gene expression observed in athlete's heart was similar to dilated cardiomyopathy with low LV diastolic stiffness modulus and preserved LV stroke work. CONCLUSIONS: High LV endomyocardial NOS gene expression is observed in athlete's heart and in dilated cardiomyopathy with low diastolic LV stiffness and preserved LV stroke work. Favourable effects on the hemodynamic phenotype of high LV endomyocardial NOS gene expression could result from a NO-mediated decrease in diastolic LV stiffness and a concomitant rise in LV preload reserve. PMID- 12123768 TI - Effects of sildenafil on cardiac repolarization. AB - OBJECTIVES: Sudden death has occasionally been reported in patients taking sildenafil. The objective of this study was to investigate the effect of sildenafil on cardiac repolarization. METHODS: We used conventional microelectrode recording technique in isolated guinea pig papillary muscles and canine Purkinje fibers, whole-cell patch clamp techniques in guinea pig ventricular myocytes, and in vivo ECG measurements in guinea pigs. RESULTS: Action potential duration at 90% repolarization (APD(90)) was not affected by sildenafil in the therapeutic ranges (< or =1 microM), but shortened by higher concentration (> or =10 microM) in both guinea pig papillary muscles and canine Purkinje fibers. D-Sotalol prolonged APD(90) in the same preparations with concentrations > or =1 microM in a reverse frequency-dependent manner. Co administration of sildenafil (10 and 30 microM) abolished the APD-prolonging effects of D-sotalol (30 microM) and amiodarone (100 microM). Sildenafil, with concentrations up to 30 microM, had no significant effect on both the rapid (I(Kr)) and the slow (I(Ks)) components of the delayed rectifier potassium currents in guinea pig ventricular myocytes. Sildenafil dose-dependently blocked L-type Ca(2+) current (I(Ca,L)), but had no effect on persistent Na(+) current in guinea pig ventricular myocytes. ECG recordings in intact guinea pigs revealed significant shortening of QTc interval by sildenafil (10 and 30 mg/kg orally). The QT-prolonging effects by D,L-sotalol (50 mg/kg) and amiodarone (100 mg/kg) were abolished by sildenafil (30 mg/kg). CONCLUSIONS: Sildenafil does not prolong cardiac repolarization. Instead, in supra-therapeutic concentrations, it accelerates cardiac repolarization, presumably through its blocking effect on I(Ca,L). PMID- 12123769 TI - Intracellular beta-blockade: overexpression of Galpha(i2) depresses the beta adrenergic response in intact myocardium. AB - OBJECTIVE: Increased levels of inhibitory G proteins have been observed in heart failure, but their physiological relevance in mediating the reduced beta adrenergic response is largely unknown. METHODS: To evaluate the functional consequences of Galpha(i2) overexpression, we studied myocardial contraction in intact isometric contracting cardiac rabbit trabeculae and isolated myocytes after adenovirus-mediated gene transfer of Galpha(i2). RESULTS: Neither Galpha(i2) nor lacZ (control) overexpression altered baseline contractile force. After 72 h of continuous contractions, developed force (F(dev)) increased after addition of 1 microM isoproterenol by 28.5+/-9.7 mN/mm(2) in the control group, which was unchanged from the initial response at t=0 h (23.7+/-3.8 mN/mm(2)). In sharp contrast, in preparations transfected with AdGalpha(i2), the response to isoproterenol was significantly attenuated (5.9+/-2.0 vs. 27.6+/-4.2 mN/mm(2), t=72 vs. 0 h, respectively, P<0.01). In a primary culture of transfected isolated myocytes from a nearly identical baseline, isoproterenol increased cell shortening by 3.1+/-0.6% in the lacZ transfected myocytes, but only by 1.3+/-0.5% in Galpha(i2) transfected myocytes (t=72 h, P<0.01). In Galpha(i2) transfected myocytes, pertussis toxin restored beta-adrenergic responsiveness, indicating specificity of attenuation by the transgene. CONCLUSIONS: Overexpression of Galpha(i2) attenuates the positive inotropic effects of beta-adrenergic stimulation in myocardium. In addition, the method we developed allows investigation of a causal link between altered protein expression and subsequent alterations in contractile function in a physiological relevant in vitro manner. PMID- 12123770 TI - Adenovirus-mediated expression of p35 prevents hypoxia/reoxygenation injury by reducing reactive oxygen species and caspase activity. AB - OBJECTIVE: This study aimed to examine the effects of adenovirus-mediated expression of p35, a baculovirus gene, on apoptosis induced by hypoxia/reoxygenation (H/R) in cardiomyocytes. METHODS: Neonatal rat cardiomyocytes were infected with recombinant adenoviral vectors expressing p35 (Ad2/CMVp35) or no transgene (Ad2/CMVEV) and were then subjected to H/R. Separate groups of non-infected cardiomyocytes were treated with pharmacological caspase inhibitors or antioxidants. Cell viability, apoptosis, caspase activity, and cellular reactive oxygen species (ROS) were measured using various assays. RESULTS: H/R decreased cell viability and increased cellular ROS levels, caspase activity, and cell apoptosis. Infection with Ad2/CMVp35 effectively inhibited the increase in cellular ROS levels, the activities of caspases 3 and 8, apoptosis, and cell death following H/R, whereas Ad2/CMVEV had no effect. Despite its ability to abolish the increase in caspase activity and partially inhibit apoptosis, the pan-caspase inhibitor ZVAD-fmk (100 microM) failed to significantly reduce cell death induced by H/R. N-acetyl-L-cysteine, an antioxidant, completely inhibited H/R-induced increase in cellular ROS levels, but reduced apoptosis and cell death by 30% only. CONCLUSIONS: Adenovirus mediated expression of p35 effectively inhibits H/R-induced cardiomyocyte apoptosis by reducing cellular ROS levels and inhibiting caspase activity. PMID- 12123771 TI - Temocapril treatment ameliorates autoimmune myocarditis associated with enhanced cardiomyocyte thioredoxin expression. AB - OBJECTIVE: Thioredoxin (TRX) is a redox regulatory protein that protects cells from various stresses. Angiotensin-converting enzyme (ACE) inhibitor was reported to enhance endogenous antioxidant enzyme activities. This study was carried out to investigate whether temocapril, a novel non-sulfhydryl-containing ACE inhibitor, reduces the severity of myocarditis via redox regulation mechanisms involving TRX. METHODS AND RESULTS: In normal rat myocytes in vitro and in vivo, Western blot showed that temocapril enhanced cytosolic redox regulatory protein TRX expression, but that neither mitochondrial TRX2 nor antioxidant enzymes, such as copper-zinc superoxide dismutase (Cu/Zn-SOD) or manganese superoxide dismutase (Mn-SOD) expression, was up-regulated by the preconditioning treatment. In rats with experimental autoimmune myocarditis (EAM), the severity of myocarditis and the protein carbonyl contents were less increased in temocapril treatment (10 mg/kg/day, orally) from day 1 to day 21, but not in temocapril treatment from day 15 to day 21. An immunohistochemical study showed that TRX stain was enhanced in infiltrating inflammatory cells and in damaged myocytes. Considering the characteristics of this model that myocardial inflammation begins around day 15 and increases until day 21, temocapril treatment for 3 weeks might be thought of as a preconditioning treatment. CONCLUSIONS: The results suggest that TRX and the redox state modified by TRX may play a crucial role in the pathophysiology of EAM. Temocapril ameliorates myocarditis associated with inducing TRX up regulation in a preconditioning manner, although the mechanism of TRX up regulation by temocapril remains to be elucidated. PMID- 12123772 TI - Cardiac cytokine expression is upregulated in the acute phase after myocardial infarction. Experimental studies in rats. AB - OBJECTIVE: The proinflammatory cytokines interleukin (IL)-1beta and IL-6 are supposed to be involved in various cardiovascular diseases including reperfusion injury and cardiac hypertrophy. METHODS AND RESULTS: In the present study, we have examined the cytokine expression from 3 h up to 12 weeks after permanent coronary artery occlusion in rats. In the first 3-12 h, there was a strong induction in IL-1beta and IL-6 mRNA expression in the infarct area (up to 50 fold) as well as in the non-infarcted myocardium (up to 15-fold). From day 3 onwards the cytokine expression was not significantly altered compared to sham operated controls. In addition, the expression of C/AATT-enhancer binding protein beta was about fourfold elevated in the first hours after myocardial infarction, but not thereafter. Furthermore, the expression of gp130 and IL-6 receptor increased significantly in the infarct area. The elevation in cytokine expression preceded the increase in matrix-metalloproteinase-9 in the infarct area as well as the increase in ANP and collagen expression in the non-infarcted myocardium. CONCLUSIONS: We suggest that IL-6 and IL-1beta act synergistically in promoting resorption of the necrotic tissue, matrix remodeling and wound healing. Furthermore, they may be involved in the early induction of fibrosis and compensatory cardiac hypertrophy of the non-infarcted myocardium, but seem not to play a key role in long-term cardiac remodeling in chronic heart failure after myocardial infarction. PMID- 12123773 TI - Increased preload directly induces the activation of heat shock transcription factor 1 in the left ventricular overloaded heart. AB - OBJECTIVES: The rapid induction of heat shock proteins (HSPs) by cardiac overload has been shown using in vivo models and it is assumed that HSPs are involved in myocardial protection against cardiac overload. However, the mechanisms for the induction of heat shock response by cardiac overload remain unclear. We examined whether increased preload as mechanical stress directly induces heat shock gene expression. METHODS: Rat hearts were isolated and perfused with Krebs-Henseleit buffer by the Langendorff method. Whole-cell extracts were prepared for gel mobility shift assay using oligonucleotides containing the heat shock element. We examined the induction of the DNA-binding activity of heat shock transcription factor (HSF), by which the transcription of heat shock genes is mainly regulated, during increased preload of left ventricle (LV) or perfusion with the buffer containing epinephrine, norepinephrine, angiotensin II, or vasopressin. RESULTS: In preloaded hearts, with LVEDP of both 30 and 50 mmHg, the DNA-binding activity of HSF1 was detected at 10 min, and increased at 20 and 60 min. At any time point, the activity with LVEDP of 50 mmHg was stronger than that with LVEDP of 30 mmHg. However, none of these hypertensive agents activated the DNA-binding activities of HSF. In afterloaded hearts, with the perfusion of norepinephrine, the activation of HSF was not induced. CONCLUSION: Our findings demonstrate that increased preload as mechanical stress directly induces the activation of HSF1 in the LV-overloaded heart. PMID- 12123774 TI - K(V)2.1 channels mediate hypoxic inhibition of I(KV) in native pulmonary arterial smooth muscle cells of the rat. AB - OBJECTIVE: To determine whether, in native pulmonary arterial smooth muscle cells (PASMC), K(V)2.1 delayed-rectifying K(+) channels are central to the process of hypoxic pulmonary vasoconstriction. METHODS: In this study, we tested for the presence of K(V)2.1 channel transcripts in rat small pulmonary arteries using RT PCR, and for the protein itself using immunolocalisation. The contribution of K(V)2.1 channels to whole-cell K(V) currents (I(KV)) and their role in hypoxic inhibition of I(KV) in native PASMC was investigated utilising patch-clamp recordings. RESULTS: K(V)2.1 mRNA expression and AbK(V)2.1 (anti-K(V)2.1 antibody) protein immunoreactivity were both present in small pulmonary arteries. Dialysis of PASMC with AbK(V)2.1 significantly attenuated I(KV) by 67% at +50 mV. Hypoxia ( approximately 20-30 mmHg) inhibited I(KV) by approximately 70% at +50 mV. Ablation of currents associated with K(V)2.1 using AbK(V)2.1 caused a marked reduction in the amplitude of I(KV). Hypoxia in the presence of the antibody did not affect the magnitude of I(KV). CONCLUSIONS: These results indicate that K(V)2.1 channel subunits exist within small pulmonary arteries and conduct a significant part of I(KV) within native PASMC. Furthermore, application of AbK(V)2.1 abolishes hypoxic inhibition of I(KV) in native PASMC suggesting that K(V)2.1 channels play a pivotal role in mediating hypoxic pulmonary vasoconstriction. PMID- 12123775 TI - Vascular endothelial growth factor-B-deficient mice show impaired development of hypoxic pulmonary hypertension. AB - OBJECTIVE: To test the hypothesis that Vegf-B contributes to the pulmonary vascular remodelling, and the associated pulmonary hypertension, induced by exposure of mice to chronic hypoxia. METHODS: Right ventricular systolic pressure, the ratio of right ventricle/[left ventricle+septum] (RV/[LV+S]) and the thickness of the media (relative to vessel diameter) of intralobar pulmonary arteries (o.d. 50-150 and 151-420 microm) were determined in Vegfb knockout mice (Vegfb(-/-); n=17) and corresponding wild-type mice (Vegfb(+/+); n=17) exposed to chronic hypoxia (10% oxygen) or housed in room air (normoxia) for 4 weeks. RESULTS: In Vegfb(+/+) mice hypoxia caused (i) pulmonary hypertension (a 70% increase in right ventricular systolic pressure compared with normoxic Vegfb(+/+) mice; P<0.001), (ii) right ventricular hypertrophy (a 66% increase in RV/[LV+S]; P<0.001) and (iii) pulmonary vascular remodelling (a 27-36% increase in pulmonary arterial medial thickness; P<0.05). In contrast, in Vegfb(-/-) mice hypoxia did not cause any increase in either right ventricular systolic pressure or pulmonary arterial medial thickness; also right ventricular hypertrophy (41% increase in RV/[LV+S]; P<0.001) was less pronounced (P<0.05) than in Vegfb(+/+) mice. CONCLUSION: Vegf-B may have a role in the development of chronic hypoxic pulmonary hypertension in mice by contributing to pulmonary vascular remodelling. If so, the effect of Vegf-B appears to be different from that of Vegf-A which is reported to protect against, rather than contribute to, hypoxia-induced pulmonary vascular remodelling. PMID- 12123776 TI - Maternal and postnatal vitamin D ingestion influences rat aortic structure, function and elastin content. AB - OBJECTIVES: Subtle impairment of fetal nutrition appears to predict hypertension and atherosclerosis in adults. It has been hypothesised that impaired aortic elastogenesis is the initiating step in adult hypertension and aortic aneurysms. Vitamin D has been shown to inhibit elastin synthesis by cultured smooth muscle cells. Here we have investigated, in rats, the hypothesis that increased exposure to vitamin D during gestation and in the postnatal period alters aortic elastin content and aortic function. METHODS: Nine breeding pairs of Sprague-Dawley rats were allocated to one of three diets containing 3000 (control group), 6000 (low dose) or 12,000 (high dose) IU/kg vitamin D during pregnancy and lactation. Male pups were continued on the same diet until 6 weeks of age. Aortic elastin content was assessed by measuring desmosine+isodesmosine content using capillary zone electrophoresis. Transverse aortic sections were used for quantification of elastic lamellae and morphometric analysis. The contractility of aortic rings was assessed in an organ bath preparation. RESULTS: The desmosine+isodesmosine content of the abdominal aorta of 6-week-old male pups, was 14.1, 10.0 and 10.1 nmol/mg dry weight in the control (n=20), low- (n=23) and high-dose (n=15) groups, respectively (P=0.007). The median number of elastic lamellae of the distal thoracic aorta was 8.25, 7.13 and 6.88 in the control, low-dose and high dose groups, respectively (P<0.001). There were no significant differences in aortic cross-sectional areas or media:adventitia ratios. The mean peak tension of aortic rings, in response to phenylephrine, was 1.3 g, 1.12 g and 0.87 g in the control, low- and high-dose groups respectively (P=0.002). CONCLUSION: In rats, exposure to increased amounts of vitamin D during gestation and early life results in a reduction of aortic elastin content, number of elastic lamellae in the aorta and force generation in aortic rings. PMID- 12123778 TI - Hypercholesterolemia impairs vascular remodelling after porcine coronary angioplasty. AB - OBJECTIVE: To assess the effect of hypercholesterolemia on neointima formation and vascular remodelling after porcine coronary angioplasty. METHODS: Left anterior descending coronary angioplasty was carried out in five control and 16 age-matched hypercholesterolemic miniature pigs. Vascular remodelling was measured by intravascular ultrasound. Neointima size and composition were assessed by quantitative image analysis. Coronary smooth muscle cells (SMC) from control and diet pigs were collected 1 h after angioplasty for in vitro study of the effect of hypercholesterolemic serum on SMC migration and of macrophage induced matrix degradation on SMC adhesion. RESULTS: Twenty-eight days after angioplasty, lumen increase was 0.08+/-1.7 mm(2) in diet and 2.7+/-2.7 mm(2) (P=0.016) in control pigs. Lumen increase correlated with vascular remodelling (IEL(post)/IEL(pre); R(2)=0.59; P<0.001) and with the circumferential gain relative to the neointima (R(2)=0.32; P<0.01) but not with neointimal area that was similar in control and diet pigs. Circumferential gain correlated with VSMC deposition at the site of the injury (R(2)=0.28; P<0.01) that correlated with organized collagen (R(2)=0.34; P<0.01). The VSMC and collagen content of neointima in diet pigs was lower whereas the macrophage content was higher. Hypercholesterolemic serum and oxidised LDL reduced migration of VSMC from diet pigs. Macrophage-induced degradation of VSMC extracellular matrix reduced VSMC adhesion (P=0.015). CONCLUSION: Hypercholesterolemia impairs vascular remodelling of balloon-treated coronary arteries. It decreases VSMC and collagen accumulation at the site of injury. Our in vitro data suggest that this decrease can be due to macrophage-induced matrix degradation and reduced VSMC adhesion and to impaired VSMC migration. Oxidised LDL mimics the inhibitory effect of hypercholesterolemic serum. PMID- 12123777 TI - Dendroaspis natriuretic peptide relaxes isolated human arteries and veins. AB - BACKGROUND: Dendroaspis natriuretic peptide (DNP) is the newest member of the natriuretic peptide family and is a circulating peptide in humans. The effects of DNP on the human vasculature are unknown. Since other natriuretic peptides are known to cause vasorelaxation, we determined the response to DNP on human blood vessels in vitro. We also investigated the mechanism of DNP mediated vasorelaxation. METHODS: Rings of human internal mammary artery and saphenous vein were suspended in an organ bath. The response to cumulative concentrations of DNP was obtained. Inhibiting agents were used to determine the mechanism of this vasorelaxation. RESULTS: DNP caused dose-dependent relaxation, with a greater effect on the internal mammary arteries (relaxation from 10(-7) mol/l DNP: 80.6+/-4.1%) than the saphenous veins (33.4+/-4.1%). At 10(-7) mol/l, DNP resulted in less arterial relaxation compared with atrial and C-type natriuretic peptides and similar relaxation to brain natriuretic peptide. In veins, DNP caused the greatest relaxation of the natriuretic peptides. DNP increased tissue cyclic guanosine monophosphate (cGMP) determined by radioimmunoassay by over 7 fold. Barium chloride and indomethacin attenuated DNP mediated vasorelaxation. However, glibenclamide, charydotoxin, apamin, tetraethyl-ammonium chloride and diisothiocyanato-stilbene-2,2'-disulfonic acid did not. DNP mediated vasorelaxation was mildly attenuated with removal of the endothelium. DNP immunoreactivity was identified in both arteries and veins. CONCLUSIONS: The current study demonstrates that DNP is an endogenous human natriuretic peptide that relaxes human arteries more than veins. Furthermore, DNP mediated vasorelaxation involves the inward rectifying potassium channels, prostaglandins, and cGMP. This newest member of the natriuretic peptide family may have an important physiologic role in the human vasculature. PMID- 12123779 TI - Carbon monoxide inhibits apoptosis in vascular smooth muscle cells. AB - OBJECTIVE: Carbon monoxide (CO) is generated from vascular smooth muscle cells via the degradation of heme by the enzyme heme oxygenase-1. Since smooth muscle cell apoptosis is associated with numerous vascular disorders, we investigated whether CO regulates apoptosis in vascular smooth muscle. METHODS AND RESULTS: Treatment of cultured rat aortic smooth muscle cells with a combination of cytokines (interleukin-1beta, 5 ng/ml; tumor necrosis factor-alpha, 20 ng/ml; interferon-gamma, 200 U/ml) for 48 h stimulated apoptosis, as demonstrated by DNA laddering, annexin V binding, and caspase-3 activation. However, the exogenous administration of CO inhibited cytokine-mediated apoptosis. The antiapoptotic action of CO was partially dependent on the activation of soluble guanylate cyclase and was associated with the inhibition of mitochondrial cytochrome c release and with the suppression of p53 expression. Incubation of smooth muscle cells with the cytokines also resulted in a pronounced increase in heme oxygenase 1 protein after 24 h of stimulation. The addition of the heme oxygenase inhibitor, zinc protoporphyrin-IX, or the CO scavenger, hemoglobin, stimulated apoptosis following 24 h of cytokine exposure. CONCLUSIONS: These results demonstrate that CO, either administered exogenously or endogenously derived from heme oxygenase-1 activity, inhibits vascular smooth muscle cell apoptosis. The ability of CO to block smooth muscle cell apoptosis may play an important role in blocking lesion formation at sites of vascular injury. PMID- 12123780 TI - Oxidized phospholipids inhibit cyclooxygenase-2 in human macrophages via nuclear factor-kappaB/IkappaB- and ERK2-dependent mechanisms. AB - OBJECTIVE: Oxidized low-density lipoproteins (ox-LDL) or their components suppress macrophage inflammatory response by down-regulating cytokine synthesis, nitric oxide synthase and inducible cyclooxygenase (Cox-2). This event is crucial for the pathophysiological process leading to the formation of atherosclerotic plaque. Our present study focused on the mechanisms through which oxidized phospholipids inhibit LPS-induced Cox-2 expression in human macrophages. METHODS: Macrophages were incubated with a mixture of oxidized fragmented phospholipids (ox-PAPC), present in modified LDL, and then exposed to LPS. Cox-2 was evaluated in terms of protein levels, mRNA and activity. RESULTS: Ox-PAPC dose-dependently inhibited Cox-2 protein, mRNA and activity by preventing NF-kappaB binding to DNA. This effect was consequent to alterations of the degradation pattern of IkappaBalpha. Moreover, ox-PAPC markedly prevented extracellular signal-regulated kinase (ERK2) activation, leading to Cox-2 expression, whereas activation of the transcription factor peroxisome proliferator-activated receptors (PPARs) was not influenced. CONCLUSION: ox-PAPC down-regulates LPS-induced Cox-2 expression in human macrophages by targeting both NF-kappaB/IkappaB and ERK2 pathways. An altered inflammatory response by macrophages within atheromata may contribute to the progression of atherosclerosis. PMID- 12123781 TI - Female gender, menstrual cycle and estradiol affect plasma levels of monocyte chemotactic protein-1 (MCP-1) in humans. PMID- 12123783 TI - Blood pressure and insulin resistance: role for microvascular function?. [Cardiovasc Res 2002;53:271-276]. PMID- 12123785 TI - 26-Nor-25-isopropyl-ergosta-5,7,22E-trien-3beta-ol: a new C(29) sterol from the sponge Agelas sceptrum from Jamaica. AB - In addition to several known sterols, a new C(29) marine sterol with a normal ergosterol nucleus but a modified side chain, named 26-nor-25-isopropyl-ergosta 5,7,22E-trien-3beta-ol, was isolated from the Jamaican sponge Agelas sceptrum. The structures were assigned by spectroscopic methods including high-resolution 2D NMR techniques. PMID- 12123786 TI - Syntheses and advanced NMR structure determination of androsteno-[17,16-d] pyrimidine derivatives. AB - Reactions of 16-hydroxymethylene- and 16-aminomethylene-3beta-hydroxy-5-androsten 17-one with formamide and guanidine were carried out resulting in the formation of [16,17-d]-pyrimidine rings. Advanced two-dimensional NMR methods were used to investigate the structure of the products. Homonuclear-, and heteronuclear chemical shift correlation experiments yielded the complete 1H-, 13C- and 15N signal assignment for these compounds. PMID- 12123787 TI - Conventional estrogen receptors are found in the plasma membrane of vaginal epithelial cells of the rat. AB - Estrogens induce rapid (non-genomic) and delayed (genomic) effects on the target cells. The early effects include induction of signal transduction pathway within seconds, whereas the delayed responses require hours and involve transcription and translation. The rapid effects of estradiol (E) on the vaginal epithelial cells (VEC) involved calcium uptake within seconds via the induction of phosphoinositol lipid metabolism as reported in our earlier studies. In this study, we demonstrate the presence of classical estrogen receptors (ER) on the plasma membrane of VEC of the rats. Immunoreactive bands of 67, 56 and 35 kDa are detectable in the membrane fractions (mf) using antibodies recognizing different epitopes of ER alpha. We have also been able to purify a protein having a mass of 67 kDa from the detergent-soluble fraction of the plasma membrane of VEC, which shows properties identical to the classical receptor purified from the cytosolic fraction of the cells. The membrane receptors get dissociated upon binding to the ligand. Besides a role in signal transduction events induced by estradiol, the membrane estrogen receptors may have an important role to play in translocation of the steroid to the cytosolic compartment. PMID- 12123789 TI - Facile synthesis of 17-formyl steroids via palladium-catalyzed homogeneous carbonylation reaction. AB - 17-formyl-androst-16-ene and its analogues were synthesized from the corresponding 17-iodo-16-ene derivatives in palladium-catalyzed formylation reaction using tributyltin hydride as hydrogen source under mild reaction conditions. The formation of androst-16-ene and its isomerization products, as well as that of analogous steroidal olefins as side-products, was found to be dependent on the reaction conditions. The formylation reaction tolerates various functional groups on the A and B rings of the steroids. PMID- 12123788 TI - 18F-labeled difluoroestradiols: preparation and preclinical evaluation as estrogen receptor-binding radiopharmaceuticals. AB - A-ring fluorination of estradiol (ES) at position 2 or 4 decreases the rate of metabolism by blocking the formation of catechol estrogens, one of the major metabolic pathways of ES. We postulate that adding a 2- or 4-fluoro substituent to 16alpha-[18F]fluoroestradiol (FES), a positron emission tomography (PET) radiopharmaceutical used for estrogen receptor (ER) imaging, should prolong its blood circulation time, and thus, improve its localization in ER-rich target tissues. On such account, we prepared a series of FES derivatives substituted with a fluorine atom at C2 or C4, with or without an 11beta-OMe group, and we tested their binding affinities for the ER and different serum proteins including rat alphafetoprotein (AFP) and human sex hormone-binding globulin (SHBG). Labeling at the 16alpha-position was accomplished via nucleophilic substitution with [18F]F(-) on the reactive 16beta,17beta-cyclic sulfate intermediates. Decay corrected yields varied between 30 and 50% for a total synthesis time of 120 min, providing final products with specific activities >3000 Ci/mmol. The 18F-labeled analogs were evaluated for their biodistribution in immature female rats. Substitutions with the 4-F have little effect on binding affinities. Addition of the 2-F diminishes ER and AFP-binding affinities while augmenting the affinity for the SHBG. Addition of the 11beta-OMe decreases all binding affinities, particularly to AFP and SHBG. In contrast, biodistribution of the corresponding [16alpha-18F]fluoro analogs in immature female rats revealed that the presence of the 11beta-OMe group improves ER-mediated uterus uptake, with the 4,16alpha [16alpha-18F]difluoro-11beta-methoxyestradiol showing the highest uptake values (15% ID at 1-h post-injection). These data suggest that the addition of both a 4 F and 11beta-OMe group onto FES may provide an improved radiopharmaceutical for PET imaging of ER densities in breast cancer patients. PMID- 12123790 TI - Measurement of cortisol secretion rate by stable isotope methodology following oral administration of 13C-labeled cortisol to a human subject. AB - Plasma concentration measurements of 13C-labeled cortisol ([1,2,4,19 13C(4)]cortisol, cortisol-13C(4)) and its metabolite cortisone-13C(4) were made simultaneously with measurements of endogenous cortisol and cortisone by gas chromatography-mass spectrometry (GC-MS). After administering a small amount (3mg) of cortisol-13C(4) to a human subject, changes in cortisol secretion rates were estimated by deconvolution techniques from the measured plasma cortisol and cortisone levels and the rates of elimination and interconversion of cortisol and cortisone were obtained from the plasma concentration-time data of cortisol 13C(4) and cortisone-13C(4). The objective of this study was to look for a novel approach to quantitate rates of minute-to-minute cortisol secretion in man by taking into account the interconversion of cortisol and cortisone by 11beta hydroxysteroid dehydrogenase (11beta-HSD). PMID- 12123791 TI - Synthesis and antiproliferative activity of side-chain unsaturated and homologated analogs of 1,25-dihydroxyvitamin D(2). (24E)-(1S)-24-Dehydro-24a-homo 1,25-dihydroxyergocalciferol and congeners. AB - A series of analogs of 1,25-dihydroxyergocalciferol (1-4) was synthesized and screened for their antiproliferative activity in vitro. The structure of new analogs was designed based on biological activity of the previously obtained side chain modified analogs of vitamin D(2) and D(3). The analogs were obtained by the Julia olefination of C(22)-vitamin D sulfone 11 with side-chain aldehyde 15. The analogs were tested for their antiproliferative activity against the cells of human breast cancer lines T47D and MCF7 as well as human and mouse leukemia lines, HL-60 and WEHI-3, respectively. Analog 2 (PRI-1907) showed the strongest antiproliferative activity out of the present series of analogs of 1,25 dihydroxyvitamin D(2) with the mono homologated and double unsaturated side chain. The activity of 2 was 3-150 times stronger, depending on the cell line, than that of 1,25-dihydroxycholecalciferol (calcitriol), used as standard. PMID- 12123792 TI - Comparison of different antibodies for detection of progesterone receptor in breast cancer. AB - Monoclonal antibodies directed against human estrogen receptor (ER) and progesterone receptor (PR) have been used extensively for biochemical and immunohistochemical detection of receptors independent of hormone-binding assays. These antibodies have been valuable both for experimental work and for detection of receptors in clinical breast cancer specimens. The purpose of this study was to characterize the sensitivity and specificity of different antibodies for detection of PR by immunohistochemistry (IHC) of formalin-fixed paraffin breast carcinoma sections. The panel of twelve antibodies included two new ones (PgR636 and PgR1294) produced prospectively to be resistant to formalin fixation and paraffin embedding. Fifty-nine breast carcinomas, having known PR levels by biochemical ligand-binding assay, were used to prepare multitumor paraffin embedded tissue blocks for characterization of the PR antibodies. Of all the antibodies tested, both PgR636 and PgR1294 stained the highest percentage of breast carcinomas known to be positive by the biochemical assay (95-98%) and they exhibited the highest concordance with the biochemical assay (88-90%). The PgR636 and PgR1294 antibodies, along with one other, PR 88, also gave the highest intensity of nuclear staining, while PgR636 and PgR1294 stained the highest mean percentage of tumor cell nuclei. Antigen retrieval was not necessary for PR immunostaining by PgR636 and PgR1294 in most tumors and other tissues examined, but did slightly increase the staining intensity. The majority of the other antibodies tested were highly dependent on antigen retrieval; only PR 88 and KD 68 antibodies approached the performance of PgR636 and PgR1294 without antigen retrieval. These results indicate that PgR636 and PgR1294 are optimal antibodies for IHC detection of PR in routine paraffin tissue blocks. PMID- 12123794 TI - The F-helix of serpins plays an essential, active role in the proteinase inhibition mechanism. AB - Proteinase inhibition by serpins requires a 70 A translocation of the proteinase, circumvention of the blocking helix F, and a crushing of the proteinase to render it catalytically incompetent. I propose that temporary displacement of the F helix during proteinase transit, and its subsequent return after complete passage of the proteinase, not only allows the proteinase to reach its final location, but provides an absolutely essential coupling mechanism for making the final proteinase crushing step energetically favorable. The F-helix is therefore not a passive impediment to proteinase translocation, but a critical, active element in permitting the serpin inhibition mechanism to operate successfully. PMID- 12123795 TI - What positions nucleosomes?--A model. AB - Here we propose a new determinant for localization of nucleosomes along genomic DNA, in addition to sequence-dependent features. The new specific class of chromatin scaling signals involves curved DNA. According to the observed positional distribution of DNA curvature, the new synchronizing signal occurs once per four nucleosomes on average. This new factor in nucleosome positioning should substantially influence the efficiency of biological reactions through regulatory factors microscopically and the entire chromatin structure through the 30 nm fiber structure macroscopically. Allocation of the new type of signals is found to be fixed evolutionarily although they could be shifted in accordance with the hierarchy of functional genomic structures. PMID- 12123796 TI - Temporal and spatial expression of two isoforms of the Dutt1/Robo1 gene in mouse development. AB - The mammalian homologue of the Drosophila axonal guidance receptor roundabout is expressed in a wide range of tissues. Here we show that alternative splicing of the Dutt1/Robo1 gene results in two mRNA transcripts with different signal peptides, which are differentially expressed throughout mouse embryogenesis. Since mice with a targeted deletion in the Dutt1/Robo1 gene have abnormal lung pathology, immunohistochemistry was used to identify the cellular expression pattern of Dutt1/Robo1 during lung development. Dutt1/Robo1 expression was widespread and diffuse in the lung at embryonic day 17.5 but became increasingly localised to the bronchial epithelium in newborn and adult mice. PMID- 12123797 TI - Iterative database searches demonstrate that glycoside hydrolase families 27, 31, 36 and 66 share a common evolutionary origin with family 13. AB - Classification of glycoside hydrolases (GHs) into families, along with the structure-based grouping together of families into clans, improve our understanding of the evolution of the large natural variety of these enzymes, help rationalise experimental data and guide further studies. Here we identify triose phosphate isomerase (TIM) barrels in GH families 27, 31, 36 and 66. We further show that iterated sequence database searches provide evidence for their sharing a common evolutionary origin with GH family 13. The catalytic, nucleophilic residue common to all these families is thereby determined and candidate catalytic proton donors identified within each family. PMID- 12123798 TI - Impaired maturation of the siderophore pyoverdine chromophore in Pseudomonas fluorescens ATCC 17400 deficient for the cytochrome c biogenesis protein CcmC. AB - Pyoverdines are the main siderophores of fluorescent pseudomonads. They comprise a quinoline chromophore, a peptide chain, and a dicarboxylic acid or a dicarboxylic acid amide side chain. Each Pseudomonas species produces a pyoverdine with a different peptide chain. A cytochrome c biogenesis DeltaccmC mutant of Pseudomonas fluorescens ATCC 17400 produces multiple pyoverdine forms, showing differences at the level of the chromophore or the side chain. When grown in the presence of L-cysteine, DeltaccmC produces only ferribactin, a non fluorescent precursor of pyoverdine, while addition of oxidized glutathione improves pyoverdine production. We suggest that the conversion of ferribactin to pyoverdine does not take place in the DeltaccmC mutant because of lack of oxidizing power in the periplasm. PMID- 12123799 TI - The crystal structure of N(1)-[2-(2-amino-ethylamino)-ethyl]-ethane-1,2-diamine (polyamines) binding to the minor groove of d(CGCGCG)(2), hexamer at room temperature. AB - The crystal structure of a left-handed Z-DNA hexamer, d(CG)(3) in complex with a synthetic polyamine, N(1)-[2-(2-amino-ethylamino)-ethyl]-ethane-1,2-diamine, NH(3)(+)-(CH(2))(2)-NH(2)(+)-(CH(2))(2)-NH(2)(+)-(CH(2))(2)-NH(3)(+) [PA(222)], has been determined by the X-ray diffraction method at 1.0 A resolution. In an orthorhombic crystal, the d(CG)(3) duplex binds two PA(222) molecules, and this synthetic polyamine exhibits dual conformational properties. One of the two PA(222) molecules resides on the floor of the minor groove of a Z-DNA duplex and imino groups bridge the two phosphate chains across a double helix, while the terminal amino groups link the oxygen atoms O2 of four cytosine bases. This PA(222) molecule makes a U-turn like a fishhook at one of its ends to provide a micro-environmental network previously unseen in complexes of DNA with polyamines. The width of the minor groove does not become considerably greater with the looped end of the polyamine, indicating conformational rigidity of the Z DNA backbone imposed by the high stacking energy of the GC base pairs. While polyamine binding to the minor groove has been postulated by theoretical studies for stabilizing the Z-DNA double helical conformation, the finding in the crystal of the looped polyamine chain binding the minor groove of Z-DNA is observed for the first time from the data collected at 10 degrees C (so-called room temperature data). Another PA(222) molecule binds on the convex outer surface of the major groove of the Z-DNA duplex and links three d(CG)(3) duplexes which are symmetrically related to each other. The structure of this PA(222) presents the previously reported zig-zag type conformation [Egli et al., Biochemistry 30 (1991) 11388-11402]. Comparison of this structure with other polyamine-DNA cocrystals reveals structural themes and differences that may relate to the length of the polyamine. PMID- 12123801 TI - Pure antiandrogens disrupt the recruitment of coactivator GRIP1 to colocalize with androgen receptor in nuclei. AB - We have used confocal microscopy to elucidate the effects of antiandrogens on nuclear localization of the androgen receptor (AR) with its transcriptional coactivator GRIP1. We show that the agonist-activated AR recruits GRIP1 to colocalize with the receptor in the nucleoplasm. By contrast, AR complexed to the antiandrogens hydroxyflutamide and bicalutamide fails to influence nuclear distribution of GRIP1. Likewise, the non-steroidal antiandrogens prevent the agonist-induced AR-GRIP1 colocalization from occurring. Androgen antagonists affect nuclear redistribution of AR-GRIP1 in a fashion that parallels their effects on the transcriptional activity of AR, in that the pure antagonists block GRIP1-dependent activation of AR function, whereas the mixed antagonist/agonist cyproterone acetate promotes both AR-driven redistribution of GRIP1 and activation of AR by GRIP1. PMID- 12123800 TI - Identification of the first Rho-GEF inhibitor, TRIPalpha, which targets the RhoA specific GEF domain of Trio. AB - The Rho-guanine nucleotide exchange factors (Rho-GEFs) remodel the actin cytoskeleton via their Rho-GTPase targets and affect numerous physiological processes such as transformation and cell motility. They are therefore attractive targets to design specific inhibitors that may have therapeutic applications. Trio contains two Rho-GEF domains, GEFD1 and GEFD2, which activate the Rac and RhoA pathways, respectively. Here we have used a genetic screen in yeast to select in vivo peptides coupled to thioredoxin, called aptamers, that could inhibit GEFD2 activity. One aptamer, TRIAPalpha (TRio Inhibitory APtamer), specifically blocks GEFD2-exchange activity on RhoA in vitro. The corresponding peptide sequence, TRIPalpha, inhibits TrioGEFD2-mediated activation of RhoA in intact cells and specifically reverts the neurite retraction phenotype induced by TrioGEFD2 in PC12 cells. Thus TRIPalpha is the first Rho-GEF inhibitor isolated so far, and represents an important step in the design of inhibitors for the expanding family of Rho-GEFs. PMID- 12123802 TI - Specific monitoring of Syk protein kinase activity by peptide substrates including constrained analogs of tyrosine. AB - The ability of Syk protein tyrosine kinase (PTK) to phosphorylate peptides, where tyrosine had been replaced by conformationally constrained analogs, has been exploited to develop highly selective substrates suitable for the specific monitoring of Syk activity. In particular we have synthesized a peptidomimetic, RRRAAEDDE(L-Htc)EEV (syktide), with the 3(S)-7-hydroxy-1,2,3,4 tetrahydroisoquinoline-3-carboxyl acid residue (L-Htc) replaced for tyrosine, which is phosphorylated by Syk with remarkable efficiency (K(cat)=73 min(-1), K(m)=11 microM), while it is not affected to any appreciable extent by a number of PTKs tested so far. These properties make syktide the first choice substrate for the specific monitoring of Syk. PMID- 12123803 TI - A thermosensitive mutant of HRV2 2A proteinase: evidence for direct cleavage of eIF4GI and eIF4GII. AB - Infection of mammalian cells with picornaviruses like entero-, rhino-, and aphthoviruses leads to an inhibition of cap-dependent cellular protein synthesis by the cleavage of both translation initiation factors, eIF4GI and eIF4GII. In entero- and rhinovirus infection this cleavage process is mediated by the viral 2A proteinase (2A(pro)). In order to discriminate between a direct mode of eIF4G cleavage and an indirect cleavage via activation of a cellular proteinase, a thermosensitive 2A(pro) mutant (ts-2A(pro)) of human rhinovirus 2 was employed. Temperature shift experiments of cytoplasmic HeLa cell extracts incubated with ts 2A(pro) strongly support a direct mode of cleavage of eIF4GI and eIF4GII by the viral 2A(pro). PMID- 12123804 TI - A survey of Cdk5 activator p35 and p25 levels in Alzheimer's disease brains. AB - P25, a calpain cleavage product of the cyclin-dependent kinase 5 (Cdk5) activator p35, causes prolonged activation of Cdk5. Although p25 has been shown to accumulate in brains of patients with Alzheimer's disease (AD), it is not known whether p25 accumulation in AD is brain region-specific. We analyzed the amounts of p25 and p35 in human autopsy samples from multiple brain regions including frontal cortex, inferior parietal cortex and hippocampus using immunoblotting assays. Our results show that the p25-p35 indices are higher in AD than in the control groups in all three brain regions. The most significant difference in p25 p35 indices between AD and control groups is in the frontal cortex. No significant difference in calpain activity between AD and control groups is observed, indicating that postmortem calpain activation cannot account for the elevation of p25/p35 ratios in AD brains. Our results support the notion that p25 accumulation deregulates Cdk5 activity in AD brains, and the deregulated Cdk5 activity may contribute to the pathogenesis of AD. PMID- 12123805 TI - Mutation of the RGD sequence does not affect plasma membrane association and growth inhibitory effects of elevated IGFBP-2 in vivo. AB - Using insulin-like growth factor-binding protein-2 (IGFBP-2) transgenic mice (D mice) as a model of elevated IGFBP-2 expression, which is often found in unphysiological conditions, we found association of IGFBP-2 to purified plasma membranes of many organs. To determine whether the RGD (Arg-Gly-Asp) motif of IGFBP-2 mediates cell surface binding in vivo, we mutated the RGD motif of IGFBP 2 into an RGE (Arg-Gly-Glu) sequence and produced transgenic mice (E mice) which express elevated amounts of mutated IGFBP-2. Our data demonstrate that in vivo IGFBP-2 cell surface association is not dependent on the RGD motif and that mutation of this sequence does not alter growth inhibitory effects of IGFBP-2. PMID- 12123806 TI - Murine peripherin gene sequences direct Cre recombinase expression to peripheral neurons in transgenic mice. AB - Spatially and temporally regulated somatic mutations can be achieved by using the Cre/loxP recombination system of bacteriophage P1. To develop a cell type specific system of gene targeting in the peripheral nervous system, we generated the transgenic mouse lines expressing Cre recombinase under the control of the mouse peripherin gene promoter. The activity of the Cre recombinase during embryonic development was examined by mating the peripherin-Cre transgenic mice to the knock-in Cre-mediated recombination reporter strain, R26R. Analysis of F1 embryos from this cross showed specific excision of loxP-flanked sequences in the dorsal root ganglia, trigeminal ganglia, and olfactory epithelium, in a pattern very similar to the expression of the endogenous mouse peripherin gene, and the previously reported peripherin-lacZ transgenic mice. Thus, the peripherin-Cre mouse described here will provide a valuable tool for Cre-loxP-mediated conditional expression in the peripheral nervous system. PMID- 12123807 TI - Interaction of hnRNP-C1/C2 proteins with RNA: analysis using the yeast three hybrid system. AB - Three-hybrid assays for the analysis of RNA-protein interactions in vivo are usually used, due to technical limitations, only for RNA baits that do not contain runs of four or more consecutive uridines. The present study provides the first example of a three-hybrid analysis of synthetic and natural uridine-rich RNA sequences. The use of the three-hybrid assay enabled us to demonstrate a functional difference between two closely related proteins, heterogeneous nuclear ribonucleoprotein C1 (hnRNP-C1) and hnRNP-C2. The hnRNP-C2 protein, an alternatively spliced variant of hnRNP-C1, contains an additional 13 amino acids between an RNA binding domain (RBD) and a basic leucine zipper-like motif (bZLM), also implied in RNA binding. This study shows that (i) for efficient binding of hnRNP-C1/C2 to RNA, the context of the U-stretch is more important than the stretch itself; (ii) both the RBD and the bZLM bind RNA independently; and (iii) the C2-related 13-amino acid insert enhances the specificity of either the RBD, the bZLM, or the full-length protein towards its ligand, allowing it to bind only the most high-affinity sequences while discriminating against those that do not perfectly match this category. The three-hybrid system is a powerful tool to work out the functional significance of peptide 'modules' within RNA binding proteins generated by alternative splicing. PMID- 12123808 TI - Influence of hydrophobic mismatch and palmitoylation on the association of transmembrane alpha-helical peptides with detergent-resistant membranes. AB - The aim of this study was to gain insight into the mechanism through which transmembrane proteins are targeted to liquid ordered (L(o)) phase domains or rafts. This was investigated by analyzing the Triton X-100 resistance of designed transmembrane peptides in model membranes of 1,2-dioleoyl-sn-glycero-3 phosphocholine, sphingomyelin and cholesterol (1/1/1, molar ratio), which contain both L(o) phase domains and fluid bilayers. By using peptides with one or two palmitate chains covalently linked to their N-terminus or with variable hydrophobic lengths, the roles of protein palmitoylation and of mismatch between the transmembrane segment of the protein and the bilayer thickness, respectively, were investigated. The results show that neither hydrophobic matching nor palmitoylation is sufficient for partitioning of peptides into L(o) phase domains. It is concluded that the L(o) phase itself, due to the tight packing of the lipids, constitutes an unfavorable environment for accommodation of protein transmembrane segments. PMID- 12123809 TI - Factor Xa induces mitogenesis of coronary artery smooth muscle cell via activation of PAR-2. AB - Factor Xa-induced stimulation of coronary artery smooth muscle cells (CASMC) was investigated by analyzing [(3)H]thymidine incorporation, cell proliferation, and ERK-1/2 activation. Exposure of the cells to factor Xa evoked a time-dependent activation of ERK-1/2 with increased [(3)H]thymidine incorporation and cell proliferation. The factor Xa-induced ERK-1/2 activation was not desensitized by preincubation of the cells with thrombin. However, ERK-1/2 activation was markedly attenuated by prior exposure of the cells to protease-activated receptor 2 (PAR-2) activating peptide, SLIGKV. The mitogenic effect of factor Xa was significantly reduced in the presence of anti-PAR-2 monoclonal antibody. Several lines of experimental evidence indicate that factor Xa-induced mitogenesis of CASMC is a cellular process mediated by PAR-2 activation. PMID- 12123810 TI - PP1/PP2A phosphatases inhibitors okadaic acid and calyculin A block ERK5 activation by growth factors and oxidative stress. AB - Okadaic acid is an inhibitor of the protein Ser/Thr phosphatases PP1 and PP2A, which blocks the activation of extracellular signal-regulated protein kinase 5 (ERK5), a member of the MAP kinase family activated by growth factors and several types of stressors. The blocking of ERK5 activation by okadaic acid was observed in HeLa cells exposed to epidermal growth factor and H(2)O(2) as well as in PC12 cells stimulated by nerve growth factor and H(2)O(2). Calyculin A, another PP1 and PP2A inhibitor, behaved similarly although these compounds are not structurally related. This suggests that either PP1 or PP2A or both are necessary for ERK5 activation. Protein kinase C (PKC) acts as a negative regulator of the ERK5 activation pathway, however our data suggest that the effects of PKC and the phosphatase are unrelated. PMID- 12123811 TI - Monoclonal anti-diuron antibodies prevent inhibition of photosynthesis by diuron. AB - Two monoclonal anti-diuron antibodies were generated that bind to diuron with an extremely low equilibrium dissociation constant. The antibodies prevented and restored in vitro and in vivo the diuron-dependent inhibition of photosynthesis. In isolated thylakoids prepared from spinach leaves (Spinacia oleracea L.) the diuron-inhibited Hill reaction was reconstituted immediately after the addition of the monoclonal antibodies. The antibodies also restored the diuron-dependent inhibition of the photosynthetic oxygen evolution of the cell wall-deficient mutant cw15 of the green alga Chlamydomonas reinhardtii Dangeard. PMID- 12123812 TI - A cytochrome c encoded by the nar operon is required for the synthesis of active respiratory nitrate reductase in Thermus thermophilus. AB - A cytochrome c (NarC) is encoded as the first gene of the operon for nitrate respiration in Thermus thermophilus. NarC is required for anaerobic growth and for the synthesis of active nitrate reductase (NR). The alpha and delta subunits (NarG, NarJ) of the NR were constitutively expressed in narC::kat mutants, but NarG appeared in the soluble fraction instead of associated with the membranes. Our data demonstrate for NarC an essential role in the synthesis of active enzyme and for the attachment to the membrane of the respiratory NR from T. thermophilus. PMID- 12123813 TI - The 1.62 A structure of Thermoascus aurantiacus endoglucanase: completing the structural picture of subfamilies in glycoside hydrolase family 5. AB - The crystal structure of Thermoascus aurantiacus endoglucanase (Cel5A), a family 5 glycoside hydrolase, has been determined to 1.62 A resolution by multiple isomorphous replacement with anomalous scattering. It is the first report of a structure in the subfamily to which Cel5A belongs. Cel5A consists solely of a catalytic module with compact eight-fold beta/alpha barrel architecture. The length of the tryptophan-rich substrate binding groove suggests the presence of substrate binding subsites -4 to +3. Structural comparison shows that two glycines are completely conserved in the family, in addition to the two catalytic glutamates and six other conserved residues previously identified. Gly 44 in particular is part of a type IV C-terminal helix capping motif, whose disruption is likely to affect the position of an essential conserved arginine. One aromatic residue (Trp 170 in Cel5A), not conserved in term of sequence, is nonetheless spatially conserved in the substrate binding groove. Its role might be to force the bend that occurs in the polysaccharide chain on binding, thus favoring substrate distortion at subsite -1. PMID- 12123814 TI - Functional selectivity for glycerol of the nodulin 26 subfamily of plant membrane intrinsic proteins. AB - The nodulin-like intrinsic protein (NIP) subfamily of water and solute channels in plants is named for nodulin 26 of legume nodules. Two NIPs, soybean nodulin 26 and Lotus japonicus LIMP2, show a distinct functional profile with a low intrinsic osmotic water permeability (P(f)) and the ability to flux uncharged polyols such as glycerol. NIPs have a conserved signature sequence within the 'aromatic/arginine' region that forms the selectivity filter for major intrinsic proteins. This sequence is a hybrid of glyceroporin and aquaporin residues as well as exhibiting substitutions unique to the NIP subfamily. Site-directed mutagenesis of a conserved tryptophan in helix 2 of LIMP2 shows that this is a major determinant of glycerol selectivity. PMID- 12123815 TI - Biosynthesis of the major scent components 3,5-dimethoxytoluene and 1,3,5 trimethoxybenzene by novel rose O-methyltransferases. AB - In Chinese rose species and in many modern varieties, two methylated phenolic derivatives, 3,5-dimethoxytoluene and 1,3,5-trimethoxybenzene, are major scent components. We show that cell-free extracts of rose petals catalyse the synthesis of 3,5-dimethoxytoluene and 1,3,5-trimethoxybenzene by methylation of precursor molecules. An expressed sequence tag approach was used to identify four highly similar O-methyltransferase sequences expressed specifically in petals and anthers. Thin layer chromatography analysis showed that the activities of these enzymes with different substrates and the proportions of reaction products produced closely mimicked those observed using cell-free petal extracts, indicating that orcinol O-methyltransferases are responsible for the biosynthesis of 3,5-dimethoxytoluene and 1,3,5-trimethoxybenzene from un-methylated precursors in this organ. PMID- 12123816 TI - 3-Chloro-DL-alanine resistance by L-methionine-alpha-deamino-gamma mercaptomethane-lyase activity. AB - The antibacterial agent 3-chloro-DL-alanine (3CA) is an inhibitor of peptidoglycan synthesis. Fusobacterium nucleatum and Porphyromonas gingivalis, the bacteria responsible for oral malodor, are shown to be resistant to 1 mM 3CA, whereas Streptococcus mutans and Escherichia coli are sensitive to this antibacterial agent at the same concentration. We isolated the 3CA resistance gene from F. nucleatum and showed that the gene encodes an L-methionine-alpha deamino-gamma-mercaptomethane-lyase that catalyzes the alpha,gamma-elimination of L-methionine to produce methyl mercaptan. The enzyme also exhibits 3CA chloride lyase (deaminating) activity. This antibacterial agent is expected to be useful for specific selection of malodorous oral bacteria producing high amounts of methyl mercaptan. PMID- 12123818 TI - Experimental study on photodynamic diagnosis of cancer mediated by chemiluminescence probe. AB - A novel method of photodynamic diagnosis of cancer mediated by chemiluminescence probe is presented. The mechanism for photodynamic therapy involves singlet oxygen ((1)O(2)) generated by energy transfer from photosensitizers. (1)O(2) can react with 3,7-dihydro-6-[4-[2-(N'-(5-fluoresceinyl)thioureido)ethoxy]phenyl]-2 methylimidazo[1,2-a]pyrazin-3-one sodium salt (FCLA), which is a Cypridina luciferin analog and a specific chemiluminescence probe for detecting (1)O(2) and superoxide (O(2)(-)). The reaction of FCLA and (1)O(2) can give emission with peak wavelength at about 532 nm. In the present study, FCLA was chosen as an optical reporter of (1)O(2) produced from the photosensitization reaction of hematoporphyrin derivative in model solution and in nude mice with transplanted mammary cancer. Photosensitized chemiluminescence from the reaction of FCLA with (1)O(2) was detected by a highly sensitive Intensified Charge-Coupled Device detector. The chemiluminescence was markedly inhibited by the addition of 10 mmol/l sodium azide (NaN(3)) to the model solution and minor effects were observed at the addition of 10 micromol/l superoxide dismutase, 20 mmol/l mannitol and 100 microg/ml catalase, respectively, thus indicating that (1)O(2) generation from photosensitization reaction mainly results in light emission. Experiments in vivo with tumor-bearing mice showed a clear chemiluminescence image of tumor. The study suggests that this novel method may be applicable to the diagnosis of superficial tumors. PMID- 12123817 TI - Cell-line specific chromatin acetylation at the Sox10-Pax3 enhancer site modulates the RET proto-oncogene expression. AB - The RET gene is expressed with high tissue and stage specificity during development. Understanding its transcriptional regulation might provide new clues to clarify developmental mechanisms. Here we show that the histone deacetylase inhibitor sodium butyrate (NaB) increases RET transcription in cells displaying low levels of its mRNA, while it has no effect in cells expressing at high levels. Chromatin immunoprecipitation (ChIP) experiments showed increased histone acetylation within the 5' flanking [corrected] region, in particular the Sox10 Pax3 enhancer site, due to NaB. Accordingly, ChIP showed different acetylation levels at the Sox10-Pax3 site associated with cell-line specific RET transcription rates. Concluding, chromatin acetylation targeted to functional sequences in the RET regulatory region may control its transcription. PMID- 12123819 TI - Generation of formate by the formyltransferase/hydrolase complex (Fhc) from Methylobacterium extorquens AM1. AB - Methylobacterium extorquens AM1 possesses a formyltransferase (Ftr) complex that is essential for growth in the presence of methanol and involved in formaldehyde oxidation to CO(2). One of the subunits of the complex carries the catalytic site for transfer of the formyl group from tetrahydromethanopterin to methanofuran (MFR). We now found via nuclear magnetic resonance-based studies that the Ftr complex also catalyzes the hydrolysis of formyl-MFR and generates formate. The enzyme was therefore renamed Ftr/hydrolase complex (Fhc). FhcA shares a sequence pattern with amidohydrolases and is assumed to be the catalytic site where the hydrolysis takes place. PMID- 12123820 TI - Characterization of an Arabidopsis thaliana mutant with impaired psbO, one of two genes encoding extrinsic 33-kDa proteins in photosystem II. AB - A 33-kDa protein component of the oxygen-evolving complex in photosystem II is essential for photosynthesis, and it has been believed that mutants with deletion of this 33-kDa protein are not found in higher plants. We report here the first isolation of an Arabidopsis thaliana mutant with a defect in one of the genes for the 33-kDa proteins, psbO, and an intact gene (psbO2). This mutant showed considerable growth retardation, suggesting that there is a functional difference between psbO and psbO2. PMID- 12123821 TI - The Aspergillus nidulans cyclin PclA accumulates in the nucleus and interacts with the central cell cycle regulator NimX(Cdc2). AB - The filamentous fungus Aspergillus nidulans reproduces asexually through conidiospores, which are continuously generated at morphologically differentiated structures, the conidiophores. In contrast to vegetative, multinucleate cells, spore formation requires a strict coordination of mitosis and cytokinesis. It was shown recently that the key regulator of the cell cycle in A. nidulans NimX(Cdc2) and a G(1)/S cyclin, PclA, are transcriptionally upregulated during development. Here we show that PclA accumulates in the nucleus and interacts with NimX(Cdc2). We propose that PclA modulates the kinase activity of NimX(Cdc2) during spore formation. PMID- 12123823 TI - Direct evidence for alteration of unfolding profile of a helical peptide by far ultraviolet circular dichroism aromatic side-chain contribution. AB - Aromatic side-chains are known to contribute to the far-UV circular dichroism (CD) spectra of peptides and proteins. Among other things, this can significantly affect the measured helix propensities of amino acids [Chakrabartty et al., Biochemistry 32 (1993) 5560-5565]. In order to address how interfering side-chain contributions can affect the backbone unfolding transition of a helical peptide, as monitored by [theta;](222) (molar ellipticity at 222 nm), we have studied the unfolding transition of a short designed (alpha-amino isobutyric acid/alanine based) helical peptide containing an interacting Tyr residue. The guanidinium hydrochloride-induced unfolding of the peptide, as monitored by [theta;](222), showed the presence of a sharp transition superposed over a much broader transition. When the same experiment was performed with a similar peptide that lacked the interacting Tyr residue, the sharp transition disappeared and only the broad transition remained. The sharp transition was assigned to originate from the interacting Tyr side-chain. This demonstrates that conformationally restricted aromatic side-chains that interact with the helical backbone not only can alter the backbone far-UV CD signal, they may also alter the unfolding profiles, monitored by far-UV CD, rendering them unfit for a simple analysis for extracting the appropriate unfolding thermodynamic parameters. PMID- 12123822 TI - Agonist-promoted heteromeric oligomerization between adenosine A(1) and P2Y(1) receptors in living cells. AB - We have explored the process of oligomerization of G protein-coupled purinergic receptors, adenosine A(1) receptor (A(1)R) and P2Y(1) receptor (P2Y(1)R), in intact HEK293T cells by means of modified bioluminescence resonance energy transfer technology (BRET(2)) that offers greatly improved separation of the emission spectra of the donor and acceptor moieties compared to traditional BRET. This approach identified both constitutive and agonist-promoted heteromeric oligomerization between Myc-tagged P2Y(1)R fused to a donor, Renilla luciferase (Myc-P2Y(1)R-Rluc) and HA-tagged A(1)R fused to an acceptor, a different form of green fluorescent protein (HA-A(1)R-GFP(2)). The BRET(2) signal increased in a time-dependent manner in the cells expressing HA-A(1)R-GFP(2)/Myc-P2Y(1)R-Rluc upon addition of agonists for both receptors, which could be inhibited by pretreatment with the P2Y(1)R antagonist MRS2179. A high degree of HA-A(1)R GFP(2) and Myc-P2Y(1)R-Rluc co-localization in the co-transfected HEK293T cells was also observed by confocal laser microscopy. These results indicate that A(1)R and P2Y(1)R can form constitutive hetero-oligomers in living cells and this process is promoted by the simultaneous activation of both receptors. PMID- 12123824 TI - Stress-induced premature senescence in BJ and hTERT-BJ1 human foreskin fibroblasts. AB - To test the involvement of the telomeres in the senescent phenotype, we used telomerase-immortalized human foreskin fibroblasts (hTERT-BJ1). We exposed hTERT BJ1 and parental BJ cells to either UVB or H(2)O(2) subcytotoxic stress(es). Both cell lines developed biomarkers of replicative senescence: loss of replicative potential, increase in senescence-associated beta-galactosidase activity, typical senescence-like morphology, overexpression of p21(WAF-1) and p16(INK-4a), and decreased level of the hyperphosphorylated form of pRb. Telomere shortening was slightly higher under stress for both BJ and hTERT-BJ1 but still much lower than that reported for other cell lines. We conclude that pathways alternative to telomere shortening must cause the appearance of the senescence phenotype. PMID- 12123825 TI - In diatoms, a transthylakoid proton gradient alone is not sufficient to induce a non-photochemical fluorescence quenching. AB - Non-photochemical fluorescence quenching (NPQ) in diatoms is associated with a xanthophyll cycle involving diadinoxanthin (DD) and its de-epoxidized form, diatoxanthin (DT). In higher plants, an obligatory role of de-epoxidized xanthophylls in NPQ remains controversial and the presence of a transthylakoid proton gradient (DeltapH) alone may induce NPQ. We used inhibitors to alter the amplitude of DeltapH and/or DD de-epoxidation, and coupled NPQ. No DeltapH dependent quenching was detected in the absence of DT. In diatoms, both DeltapH and DT are required for NPQ. The binding of DT to protonated antenna sites could be obligatory for energy dissipation. PMID- 12123826 TI - Identification of prolylcarboxypeptidase as the cell matrix-associated prekallikrein activator. AB - Investigations determined that the cell matrix-associated prekallikrein (PK) activator is prolylcarboxypeptidase. PK activation on human umbilical vein endothelial cell (HUVEC) matrix is inhibited by antipain (IC(50)=50 microM) but not anti-factor XIIa antibody, 3 mM benzamidine, 5 mM iodoacetic acid or iodoacetamide, or 3 mM N-ethylmaleimide. Corn trypsin inhibitor (IC(50)=100 nM) or Fmoc-aminoacylpyrrolidine-2-nitrile (IC(50)=100 microM) blocks matrix associated PK activation. Angiotensin II (IC(50)=100 microM) or bradykinin (IC(50)=3 mM), but not angiotensin 1-7 or bradykinin 1-5, inhibits matrix associated PK activation. ECV304 cell matrix PK activator also is blocked by 100 microM angiotensin II, 1 microM corn trypsin inhibitor, and 50 microM antipain, but not angiotensin 1-7. 1 mM angiotensin II or 300 microM Fmoc aminoacylpyrrolidine-2-nitrile indirectly blocks plasminogen activation by inhibiting kallikrein formation for single chain urokinase activation. On immunoblot, prolylcarboxypeptidase antigen is associated with HUVEC matrix. These studies indicate that prolylcarboxypeptidase is the matrix PK activator. PMID- 12123827 TI - The human dynamin-related protein OPA1 is anchored to the mitochondrial inner membrane facing the inter-membrane space. AB - Mutations in the OPA1 gene are associated with autosomal dominant optic atrophy. OPA1 encodes a dynamin-related protein orthologous to Msp1 of Schizosaccharomyces pombe and Mgm1p of Saccharomyces cerevisiae, both involved in mitochondrial morphology and genome maintenance. We present immuno-fluorescence and biochemical evidences showing that OPA1 resides in the mitochondria where it is imported through its highly basic amino-terminal extension. Proteolysis experiments indicate that OPA1 is present in the inter-membrane space and electron microscopy further localizes it close to the cristae. The strong association of OPA1 with membranes suggests its anchoring to the inner membrane. PMID- 12123828 TI - The hydroxyphenylpyruvate dioxygenase from Synechocystis sp. PCC 6803 is not required for plastoquinone biosynthesis. AB - The disruption of the Synechocystis open reading frame Deltaslr0090 encoding a gene with high homology to plant genes encoding 4-hydroxyphenylpyruvate dioxygenase results in an impairment of tocopherol biosynthesis without affecting levels of plastoquinone, carotenoids and chlorophyll as well as cell growth and photosynthesis. Our results indicate that in Synechocystis in contrast to the situation in higher plants the 4-hydroxyphenylpyruvate dioxygenase is not required for the synthesis of plastoquinone. PMID- 12123829 TI - Modification of rRNA as a 'quality control mechanism' in ribosome biogenesis. AB - An efficiently expressed rDNA plasmid was used to quantitatively analyze the effect of base changes in modified positions associated with the peptidyl transferase center of the 25S rRNA from the yeast Schizosaccharomyces pombe. The results show that, unlike normal RNA and relative to a less conserved modified position outside the center, these mutant RNAs are highly unstable and rapidly degraded with little or no effect on cell growth. These results provide direct evidence that the positions of modification can be critical sites for nuclease attack. Taken together with previous genetic analyses of rRNA modification, they raise the possibility that rRNA modification may act, at least in part, as a quality control mechanism to help ensure that only functional RNA is incorporated into active ribosomes. PMID- 12123830 TI - Lysophosphatidic acid (LPA) receptors are activated differentially by biological fluids: possible role of LPA-binding proteins in activation of LPA receptors. AB - Lysophosphatidic acid (LPA) exerts multiple biological functions through G protein-coupled receptors (EDG2/LPA(1), EDG4/LPA(2), and EDG7/LPA(3)) and is present in serum where it is associated with albumin. In this study we examined LPA activity in various biological fluids by measuring the LPA-induced increase in the intracellular concentration of calcium ion in three types of Sf9 insect cells, each expressing one of the LPA receptors. Using this system, we found that EDG2 and EDG4, but not EDG7, were activated strongly by addition of incubated plasma. By contrast, LPA detected in seminal plasma, which contains a low concentration of albumin, selectively activated EDG7. After LPA in these samples was extracted and reconstituted, it activated all three receptors. We also found that serum albumin readily inhibits the activation of EDG7 but not the activation of EDG2 or EDG4. In addition, plasma from Nagase analbuminemic rats but not plasma from control Sprague-Dawley rats was found to strongly activate EDG7, although the plasma of these two types of rats contained equal amounts of LPA and activated both EDG2 and EDG4. The present study shows that serum albumin can negatively regulate EDG7 but not EDG2 or EDG4, and suggests that protein factors are present in seminal plasma and deliver LPA efficiently to EDG7 but not to EDG2 or EDG4. PMID- 12123832 TI - Mitochondria hyperpolarization is an early event in oxidized low-density lipoprotein-induced apoptosis in Caco-2 intestinal cells. AB - We investigated the mechanisms underlying the pro-apoptotic activity exerted by oxidized low-density lipoproteins (oxLDL) in Caco-2 intestinal cells, a cell line which retains many morphological and enzymatic features typical of normal human enterocytes. We found that: (i) oxLDL induced mitochondrial-mediated apoptosis by provoking first an increase in mitochondrial membrane potential, followed, later, by the typical apoptosis-associated depolarization (type II apoptosis); accordingly, (ii) caspase-9 inhibition significantly hindered apoptosis while caspase-8 inhibition did not; and finally (iii) dietary phenolic antioxidizing compounds exerted a significant protective antiapoptotic activity. These results point to mitochondrial hyperpolarization as 'sensitizing feature' in apoptotic proneness of Caco-2 intestinal cells to oxLDL exposure. PMID- 12123831 TI - Structural domains required for channel function of the mouse transient receptor potential protein homologue TRP1beta. AB - Transient receptor potential proteins (TRP) are supposed to participate in the formation of store-operated Ca(2+) influx channels by co-assembly. However, little is known which domains facilitate the interaction of subunits. Contribution of the N-terminal coiled-coil domain and ankyrin-like repeats and the putative pore region of the mouse TRP1beta (mTRP1beta) variant to the formation of functional cation channels were analyzed following overexpression in HEK293 (human embryonic kidney) cells. MTRP1beta expressing cells exhibited enhanced Ca(2+) influx and enhanced whole-cell membrane currents compared to mTRP1beta deletion mutants. Using a yeast two-hybrid assay only the coiled-coil domain facilitated homodimerization of the N-terminus. These results suggest that the N-terminus of mTRP1beta is required for structural organization thus forming functional channels. PMID- 12123833 TI - Magic-angle spinning (31)P NMR spectroscopy of condensed phosphates in parasitic protozoa: visualizing the invisible. AB - We report the results of a solid-state (31)P nuclear magnetic resonance (NMR) spectroscopic investigation of the acidocalcisome organelles from Trypanosoma brucei (bloodstream form), Trypanosoma cruzi and Leishmania major (insect forms). The spectra are characterized by a broad envelope of spinning sidebands having isotropic chemical shifts at approximately 0, -7 and -21 ppm. These resonances are assigned to orthophosphate, terminal (alpha) phosphates of polyphosphates and bridging (beta) phosphates of polyphosphates, respectively. The average polyphosphate chain length is approximately 3.3 phosphates. Similar results were obtained with whole L. major promastigotes. (31)P NMR spectra of living L. major promastigotes recorded under conventional solution NMR conditions had spectral intensities reduced with respect to solution-state NMR spectra of acid extracts, consistent with the invisibility of the solid-state phosphates. These results show that all three parasites contain large stores of condensed phosphates which can be visualized by using magic-angle spinning NMR techniques. PMID- 12123835 TI - Electrophysiological characterization and molecular identification of the Phoneutria nigriventer peptide toxin PnTx2-6. AB - A cDNA with 403 nucleotides encoding the precursor of the toxin PnTx2-6 was cloned and sequenced. Subsequent analysis revealed that the precursor begins with a signal peptide and a glutamate-rich propeptide. The succeeding peptide confirmed the reported sequence of PnTx2-6. The purified toxin exerted complex effects on Na(+) current of frog skeletal muscle. There was a marked decrease of the inactivation kinetics, and a shift to hyperpolarizing potentials of both the Na(+) conductance and the steady-state inactivation voltage dependences, along with a reduction of the current amplitude. The concentration dependence of the modified current suggests a K(D) of 0.8 microM for the toxin-channel complex. PMID- 12123834 TI - Characterisation of a novel mouse liver aldo-keto reductase AKR7A5. AB - We have characterised a novel aldo-keto reductase (AKR7A5) from mouse liver that is 78% identical to rat aflatoxin dialdehyde reductase AKR7A1 and 89% identical to human succinic semialdehyde (SSA) reductase AKR7A2. AKR7A5 can reduce 2 carboxybenzaldehyde (2-CBA) and SSA as well as a range of aldehyde and diketone substrates. Western blots show that it is expressed in liver, kidney, testis and brain, and at lower levels in skeletal muscle, spleen heart and lung. The protein is not inducible in the liver by dietary ethoxyquin. Immunodepletion of AKR7A5 from liver extracts shows that it is one of the major liver 2-CBA reductases but that it is not the main SSA reductase in this tissue. PMID- 12123836 TI - Cytoskeleton-related trafficking of the EAAC1 glutamate transporter after activation of the G(q/11)-coupled neurotensin receptor NTS1. AB - The possible modulation of the glutamate transporter EAAC1 by a class A G protein coupled receptor was studied in transfected C6 glioma cells stably expressing the high-affinity neurotensin receptor NTS1. Brief exposure (5 min) to neurotensin increased Na(+)-dependent D-[(3)H]aspartate uptake by about 70%. The effect of neurotensin was found to result from an increase in cell surface expression of EAAC1 and accordingly, cytochalasin D and colchicine were shown to block the effect of neurotensin on aspartate uptake, suggesting that the cytoskeleton participates in this regulation. Neither protein kinase C nor phosphatidylinositol 3-kinase activities, two intracellular signaling pathways known to modulate EAAC1, was required for EAAC1-mediated aspartate transport regulation by neurotensin. Together, these results provide evidence for an acute regulation of EAAC1 trafficking after activation of a G protein-coupled receptor. PMID- 12123837 TI - Amino-terminally truncated desmin rescues fusion of des(-/-) myoblasts but negatively affects cardiomyogenesis and smooth muscle development. AB - Desmin fulfils important functions in maintenance of muscle cells and mutations in the desmin gene have been linked to a variety of myopathies. To ascertain the role of desmin's amino-terminal domain in muscle cells we generated embryonic stem cells constitutively expressing desmin(Delta1-48) in a null background and investigated muscle cell development in vitro. Desmin(Delta1-48) lacking the first 48 amino acid residues promotes fusion of myoblasts, rescues myogenesis and down-regulates vimentin expression in embryoid bodies, but hampers cardiomyogenesis and blocks smooth muscle development. These results demonstrate that desmin's amino-terminus has different roles in skeletal, cardiac, and smooth muscle cell development and function. PMID- 12123838 TI - Escherichia coli 6-pyruvoyltetrahydropterin synthase ortholog encoded by ygcM has a new catalytic activity for conversion of sepiapterin to 7,8-dihydropterin. AB - The putative gene (ygcM) of Escherichia coli was verified in vitro to encode the ortholog of 6-pyruvoyltetrahydropterin synthase (PTPS). Unexpectedly, the enzyme was found to convert sepiapterin to 7,8-dihydropterin without any cofactors. The enzymatic product 7,8-dihydropterin was identified by HPLC and mass spectrometry analyses, suggesting a novel activity of the enzyme to cleave the C6 side chain of sepiapterin. The optimal activity occurred at pH 6.5-7.0. The reaction rate increased up to 3.2-fold at 60-80 degrees C, reflecting the thermal stability of the enzyme. The reaction required no metal ion and was activated slightly by low concentrations (1-5 mM) of EDTA. The apparent K(m) value for sepiapterin was determined as 0.92 mM and the V(max) value was 151.3 nmol/min/mg. The new catalytic function of E. coli PTPS does not imply any physiological role, because sepiapterin is not an endogenous substrate of the organism. The same activity, however, was also detected in a PTPS ortholog of Synechocystis sp. PCC 6803 but not significant in Drosophila and human enzymes, suggesting that the activity may be prevalent in bacterial PTPS orthologs. PMID- 12123839 TI - The crystal structure of anthranilate phosphoribosyltransferase from the enterobacterium Pectobacterium carotovorum. AB - The structure of anthranilate phosphoribosyltransferase from the enterobacterium Pectobacterium carotovorum has been solved at 2.4 A in complex with Mn(2+) pyrophosphate, and at 1.9 A without ligands. The enzyme structure has a novel phosphoribosyltransferase (PRT) fold and displays close homology to the structures of pyrimidine nucleoside phosphorylases. The enzyme is a homodimer with a monomer of 345 residues. Each monomer consists of two subdomains, alpha and alpha/beta, which form a cleft containing the active site. The nature of the active site is inferred from the trapped MnPPi complex and detailed knowledge of the active sites of nucleoside phosphorylases. With the anthranilate (An)PRT structure solved, the structures of all the enzymes required for tryptophan biosynthesis are now known. PMID- 12123840 TI - First tau repeat domain binding to growing and taxol-stabilized microtubules, and serine 262 residue phosphorylation. AB - Tau phosphorylation plays a crucial role in microtubule stabilization and in Alzheimer's disease. To characterize the molecular mechanisms of tau binding on microtubules, we synthesized the peptide R1 (QTAPVPMPDLKNVKSKIGSTENLKHQPGGGKVQI), reproducing the first tau microtubule binding motif. We thermodynamically characterized the molecular mechanism of tubulin assembly with R1 in vitro, and measured, for the first time, the binding parameters of R1 on both growing and taxol-stabilized microtubules. In addition, we obtained similar binding parameters with R1 phosphorylated on Ser262. These data suggest that the consequences of Ser262 phosphorylation on tau binding to microtubules and on tubulin assembly are due to large intramolecular rearrangements of the tau protein. PMID- 12123841 TI - Transposable elements encoding functional proteins: pitfalls in unprocessed genomic data? PMID- 12123845 TI - Mutation analysis and association studies of the ubiquitin carboxy-terminal hydrolase L1 gene in Huntington's disease. AB - Huntington's disease (HD) is attributed to a triplet CAG repeat mutation, and about 70% of the variance in age-at-onset can be explained by the size of the repeat expansion. Among potential candidates as modifier genes, we investigated the role of ubiquitin carboxy-terminal hydrolase L1 (UCH-L1) gene. We examined the association of HD with the I93M mutation and S18Y polymorphism in 138 HD patients and 136 control subjects, but we did not identify the I93M mutation. The S18Y polymorphism was present in 17% of HD patients. Of the variance in the age at-onset that was not accounted for by the CAG repeat, 13% could be attributed to S18Y polymorphism. We sequenced the entire coding region of the UCH-L1 gene in seven HD patients with unexplained older or younger onset age. The S18Y polymorphism was found in three out of the four patients presenting with a later age-at-onset. We conclude that the UCH-L1 gene may be a genetic factor that influences the variability in age-at-onset of HD. PMID- 12123846 TI - Comparison of the binding of [(3)H]nociceptin/orphaninFQ(1-13)NH(2), [(3)H]nociceptin/orphaninFQ(1-17)OH and [(125)I]Tyr(14)nociceptin/orphaninFQ(1 17)OH to recombinant human and native rat cerebrocortical nociceptin/orphanin FQ receptors. AB - Nociceptin/orphanin FQ (N/OFQ) is a 17-amino acid endogenous neuropeptide ligand for the nociceptin receptor (NOP). We have prepared a [(3)H]-labelled truncated N/OFQ peptide, [(3)H]N/OFQ(1-13)NH(2) and compared its binding characteristics with [(3)H]N/OFQ(1-17)OH and [(125)I]Y(14)N/OFQ(1-17)OH in membranes prepared from Chinese hamster ovary cells expressing the recombinant human NOP (CHO(hNOP)) and the rat cerebrocortex. [(3)H]N/OFQ(1-13)NH(2), [(3)H]N/OFQ(1-17)OH and [(125)I]Y(14)N/OFQ(1-17)OH binding to CHO(hNOP) was concentration dependent and saturable with receptor density (B(max)) and radioligand equilibrium dissociation constant (pK(d)) values (mean +/- SEM) of 1043 +/- 58 fmol/mg protein and 10.35 +/- 0.03, 1348 +/- 44 fmol/mg protein and 10.06 +/- 0.04, and 1169 +/- 76 fmol/mg protein and 10.45 +/- 0.06, respectively. In the rat, B(max) and pK(d) values for [(3)H]N/OFQ(1-13)NH(2) and [(3)H]N/OFQ(1-17)OH were 130 +/- 1 fmol/mg protein and 10.70 +/- 0.03, and 157 +/- 4 fmol/mg protein and 10.34 +/- 0.02, respectively. The binding of all radioligands was displaced by a range of peptide and non peptide ligands. There was a strong correlation (r(2) = 0.92, P = 0.002) between pK(i) values estimated with [(3)H]N/OFQ(1-13)NH(2) and [(3)H]N/OFQ(1-17)OH. No such correlation was observed in comparison with the [(125)I]-labelled peptide (poor agreement with low affinity N/OFQ(1-9)NH(2), Dynorphin-A and Naloxone benzoylhydrazone). We suggest that [(3)H]N/OFQ(1-13)NH(2) may be a useful alternative to [(3)H]N/OFQ(1-17)OH. PMID- 12123847 TI - Genetic variation in the cholesterol 24-hydroxylase (CYP46) gene and the risk of Alzheimer's disease. AB - Alzheimer's disease (AD) is a complex, multifactorial disorder, with many genetic and environmental factors implicated in disease onset and pathology. Increasing evidence points to a link between brain cholesterol turnover and AD. The CYP46 gene encodes for the enzyme, cholesterol 24-hydroxylase, which plays a key role in brain cholesterol turnover. A polymorphism in Intron 2 (T-->C) in the CYP46 gene has recently been reported to be associated with the risk of AD. In the present study, we examined the association of this CYP46 polymorphism with sporadic late-onset AD (LOAD) in American White (434 cases, 401 controls) and African American (54 cases, 61 controls) cohorts. No significant association was observed between the CYP46 polymorphism and LOAD. When the data were stratified by the apolipoprotein E*4 carrier status, no significant difference was observed between cases and controls for the CYP46 single nucleotide polymorphism. In addition, no significant difference in genotype or allele frequency was observed when stratified by the presence or absence of the alpha1-antichymotrypsin*A allele. Our data indicate that the Intron 2 polymorphism of CYP46 does not affect the risk of AD in our sample. PMID- 12123848 TI - Expiratory resistive loaded breathing in humans increases fluctuations of force production in submaximal isometric quadriceps contractions. AB - This study demonstrated that expiratory resistive loading (ERL) induced an increase in force fluctuation during a unilateral, submaximal isometric contraction of the non-dominant left vastus lateralis (VL), but did not effect force fluctuation during complex bilateral contractions. The increase in force fluctuation in the unilateral left VL contraction during ERL was not accompanied by alterations of average force production, motor unit activation (median power frequency) or airflow rate when compared to the bilateral contraction. Inspiratory RL (IRL) did not significantly affect force fluctuation in unilateral or bilateral contractions. The results concur with previous reports of ERL, but not IRL, effecting VL function and suggest that patients with obstructive diseases may also be vulnerable to reduced fine motor control. PMID- 12123849 TI - Delay modulates spectral correlates in the human EEG of non-verbal auditory working memory. AB - Studies using neuroimaging and electro- and magnetoencephalographic techniques have begun to identify the brain structures and dynamics that underlie auditory working memory. However, past research has not clearly characterized how the neural dynamics varies with the delay over which auditory information must be maintained. We used electroencephalogram band power as a measure of relative neuronal synchrony during a non-verbal auditory working memory task. Comparing the working memory task with a control recognition task, the relative synchrony in bilateral theta and alpha bands was unchanged using a two second delay. However, five and ten second delays produced increases and decreases in relative synchrony, respectively. The memory task also induced greater synchronization in beta and gamma bands over the right temporal cortex during the two and five second delays. The results suggest that the cortical dynamics that underlie auditory working memory are highly dependent upon a duration-dependent encoding strategy. PMID- 12123851 TI - Differential effects of novel wasp toxin on rat hippocampal interneurons. AB - We studied the effects of a wasp toxin beta-pompilidotoxin (beta-PMTX) on rat hippocampal CA1 interneurons by the current-clamp technique. The firing patterns of pyramidal neurons and pyramidale interneurons were not affected by beta-PMTX, but in oriens and radiatum interneurons, beta-PMTX converted the action potentials to prolonged depolarizing potentials by slowing the inactivation of Na(+) channels. In lacunosum moleculare interneurons, beta-PMTX induced initial bursting spikes followed by block of succeeding spikes. Comparison of beta-PMTX with a sea anemone toxin, ATX II, revealed that ATX II altered the firing properties of pyramidal neurons and pyramidale interneurons that were unchanged by beta-PMTX. Our results suggest that beta-PMTX modulates Na(+) currents in CA1 interneurons differently in various CA1 neurons and the toxin is useful to classify Na(+) channel subtypes. PMID- 12123850 TI - Cryopreservation and primary culture of cerebral neurons from cynomolgus monkeys (Macaca fascicularis). AB - We established the procedures for cryopreservation and primary culture of fetal cerebral neurons of cynomolgus monkeys (Macaca fascicularis). Three developmental stages of fetuses (80, 93, and 102 days of gestation) were compared to determine the optimal stage of cerebrum development for primary culture. Among the three fetuses, the 80-day-old fetus produced the most process-rich neurons with the highest survival. The number of total recovery cells from the cryopreserved 80 day-old fetus corresponded to 83.4% of that from fresh tissue. Besides, synchronous oscillations of intracellular calcium were first seen in primate cerebral neurons, which suggested the formation of synapse-networks. Cultured neurons expressed synaptophysin protein. Successful cryopreservation and subsequent cell culture of primate neurons would be useful tools for neuroscience research with species specificity. PMID- 12123852 TI - Attentional modulation of human pain processing in the secondary somatosensory cortex: a magnetoencephalographic study. AB - The influence of attention on the processing of pain in the secondary somatosensory cortex (SII) was analyzed using magnetoencephalography in response to painful infra-red heat stimuli applied to the left hand in six male healthy subjects, aged 22-28 years. Three experimental paradigms were chosen to deliver attention dependent results under comparable levels of vigilance. Single moving dipole sources for the pain-evoked responses were calculated in the individual cortex anatomy determined by magnetic resonance imaging. Though pain stimuli followed the same intensity pattern in all paradigms, evoked SII activity increased markedly from the low attention task to the mid-level attention task (P < 0.001). In contrast, further increase of attention from mid-level to high was not accompanied by an additional enhancement of SII activity. It therefore is concluded that activation patterns of SII follow a saturation function which cannot be enlarged by maximizing the relevance of the painful stimuli. PMID- 12123853 TI - Allopregnanolone attenuates N-methyl-D-aspartate-induced excitotoxicity and apoptosis in the human NT2 cell line in culture. AB - Progesterone modulates gamma-aminobutyric acid and excitatory amino acid neurotransmitter systems and has neuroprotective properties in models of hypoxia ischemia. This study examined the in vitro effects of allopregnanolone, the active progesterone metabolite, in models of N-methyl-D-aspartate (NMDA)-induced necrosis and apoptosis. Cultured NT2 neurons were exposed to 1 mM NMDA. Lactate dehydrogenase (LDH) release was measured 24 h later. NMDA at a concentration of 1 mM produced a 39 +/- 19% release of total LDH. Exposure to 10 microM allopregnanolone prior to NMDA exposure reduced LDH release by 51% (P = 0.0028). NMDA stimulated apoptotic cell changes defined by terminal dUTP nick-end labeling (TUNEL) and 5,5', 6,6'-tetrachloro-1,1,3,3'-tetra ethlybenzimidazolycarbocyanide iodide staining were reduced to baseline values by both 10 microM allopregnanolone and 100 microM MK-801. Pretreatment with allopregnanolone (0-10 microM) reduced the percentage of TUNEL-positive cells in a dose-dependent manner (EC(50) = 2.7 +/- 0.1 nM). Physiologic concentrations of allopregnanolone provided protection against both necrotic and apoptotic injury induced by NMDA excitotoxicity. PMID- 12123854 TI - Absence of coherence between cervical and lumbar spinal cord dorsal surface potentials in the anaesthetized cat. AB - Recordings of spontaneous cord dorsum potentials (CDPs) along the longitudinal axis of the spinal cord were made. These recordings were obtained from the surface of the dorsal horn at different points along the spinal cord caudally and cranially in relation to the point giving spontaneous potentials of maximal amplitude. We found two curves (lumbar and cervical) for the longitudinal distribution of the area of the power spectra of these recordings. Each of these curves had a symmetrical decrement on both sides of the position of the point for the maximal area of power. Such points were discovered on the L5-L7 and C3-C4 spinal segments. Spectral analysis of the spontaneous CDPs simultaneously recorded in both regions indicates no evidence of coherence, thus suggesting that the spontaneous CDPs recorded in the lumbar and cervical regions of the pentobarbitone-anaesthetized cat are generated by two independent populations of neurones not functionally interconnected between them. PMID- 12123855 TI - Novel object presentation affects sleep-wake behavior in rats. AB - Sleep is suggested to be crucial for the processing and storage of new information. Several learning tasks have been shown to increase the amount of rapid eye movement sleep (REMS) with its typical theta activity (6-8 Hz) relative to total sleep time. Vice versa, REMS deprivation is able to affect memory consolidation following some, but not all learning tasks. Furthermore, recent studies have shown an increase of spindle activity (12-15 Hz) within the electroencephalogram (EEG) of nonREMS as well. The enhancement of both spindle and theta activity is suggested to serve as background activity for the synchronization of those neuronal pathways that were involved in the registration and, later on, participate in the long-term storage of new information in defined brain regions. In the present study, the presentation of a novel object to rats enhanced the amount of preREMS, an intermediate sleep stage with high spindle activity, within the first 2 h of the subsequent sleeping phase. Four hours later, the amount of REMS was increased as well. However, there were no changes in the EEG power spectra of nonREMS, preREMS and REMS. We therefore hypothesize that the increase of preREMS and REMS amounts and the related spindle and theta activity stand for the processing and storage of new information about the presented novel objects. PMID- 12123856 TI - Heterogeneity of pituitary adenylate cyclase-activating polypeptide and vasoactive intestinal polypeptide receptors in rat intrinsic cardiac neurons. AB - The expression of receptors for pituitary adenylate cyclase-activating polypeptide (PACAP) and vasoactive intestinal polypeptide (VIP) was investigated in isolated parasympathetic neurons of neonatal rat intracardiac ganglia using single-cell reverse transcription-polymerase chain reaction. Individual neurons were shown to express multiple isoforms of the PACAP receptor, PAC1, including PAC1-short, -HOP1 and -HOP2 variants, which differ in the region encoding the G protein-binding domain. The PAC1-HOP1 isoform was the predominant species, being expressed at higher levels and in a greater number of cells than other PAC1 variants. In addition to PAC1, intrinsic cardiac neurons express transcripts for the VIP receptors, VPAC1 and VPAC2, with VPAC2 being found in a greater proportion of the neurons. These findings may explain the complex effects of PACAP and VIP on neuroexcitability in mammalian intracardiac ganglia. PMID- 12123857 TI - Spatial memory testing decreases hippocampal amyloid precursor protein in young, but not aged, female rats. AB - Using young and aged rats, we investigated relationships between amyloid precursor protein (APP) and working or reference memory, as well as assessed whether cognitive testing altered APP levels. In young rats, higher APP levels were related to more working memory errors as a linear function. Aged rats exhibited a curvilinear relationship between APP and working memory, with moderate APP levels associated with better relative performance. A comparison of rats that received cognitive testing with those that did not showed that testing decreased APP levels in young, but not aged, rats. Collectively, the data suggest that young and aged rats exhibit different relationships between APP and working memory, and that aged rats do not maintain the capacity to decrease APP in response to cognitive testing. PMID- 12123858 TI - Ergothioneine treatment protects neurons against N-methyl-D-aspartate excitotoxicity in an in vivo rat retinal model. AB - Injection of the glutamate agonist N-methyl-D-aspartate into the vitreous body of the rat eye resulted in a number of morphological changes in the retina. Most apparent was a dramatic reduction in the density and sizes of neurons accompanied by a decrease in amyloid precursor protein and glial fibrillary acidic protein immunoreactivity. Cell counts revealed that 81% of ganglion cells and 43% of non ganglion cells were lost as a result of the treatment. However, in animals treated with the antioxidant ergothioneine, these figures dropped to 44 and 31%, respectively. Thus, ergothioneine appears to be neuroprotective in this system and the data suggest that antioxidants may provide a useful means of modulating glutamate-based toxicity. PMID- 12123859 TI - Opioid control of prolactin secretion in late pregnant rats is mediated by tuberoinfundibular dopamine neurons. AB - Prolactin (PRL) secretion from the anterior pituitary is tonically inhibited by tuberoinfundibular dopamine (TIDA) neurons in the arcuate nucleus of the hypothalamus. During late pregnancy, TIDA neuronal activity is reduced allowing the expression of an antepartum PRL surge. We show here that continuous infusion of the opioid receptor antagonist naloxone (10 mg/h) during the night preceding parturition completely abolished the antepartum PRL surge and significantly increased TIDA neuronal activity. These data indicate that endogenous opioid neurons facilitate PRL secretion at the end of pregnancy by suppressing TIDA neuronal activity. PMID- 12123860 TI - No evidence of association between the alpha-2 macroglobulin gene and Parkinson's disease in a case-control sample. AB - Alpha-2 macroglobulin (A2M) is a component of Lewy bodies, a hallmark of Parkinson's disease (PD). In 159 PD patients and 190 normal controls, we studied two A2M polymorphisms by the polymerase chain reaction-restriction fragment length polymorphism method: a five-nucleotide deletion at the 5' splice site of exon 18; and a valine to isoleucine exchange in amino acid position 1000 near the thiolester active site. No significant differences in allelic and genotypic distribution were found between cases and controls or between early and late onset PD patients. The present data suggest that these polymorphisms do not represent a risk factor for PD and do not modulate the age at onset of PD. PMID- 12123861 TI - Trait anxiety affects the pupillary light reflex in college students. AB - This study was designed to examine whether anxious personality, i.e. trait anxiety, influences the autonomic nervous functions in humans without manipulation of experimental stressors. The degrees of state and trait anxiety, blood pressure, heart rate, pupillary light reflex (PLR), and body temperature were measured at the same hour on four different days in 14 healthy college students. A multiple regression analysis showed that trait anxiety predominantly influenced state anxiety and the PLR parameters. A single regression analysis showed that trait anxiety positively correlated to the initial pupillary diameter and the constricted diameter of PLR and negatively to the amplitude of PLR. It was concluded that trait anxiety predicts state anxiety and a smaller amplitude of PLR in humans at rest. PMID- 12123862 TI - No synergistic effect between -850 tumor necrosis factor-alpha promoter polymorphism and apolipoprotein E epsilon 4 allele in Alzheimer's disease. AB - In the brains of Alzheimer's disease (AD) patients, microglia cells are activated and produce inflammatory mediators such as tumor necrosis factor-alpha (TNF alpha). A recent study conducted in Northern Ireland showed that a polymorphism in the promotor region (-850) of the TNF-alpha gene increased the risk of AD associated with carriage of the apolipoprotein E (APOE) epsilon 4 allele. In a case-control study restricted to a population from Northern Spain and utilizing 321 sporadic AD patients and 312 control subjects, we have found that the -850 TNF-alpha polymorphism does not interact with the APOE gene to increase the risk associated with the epsilon 4 allele. PMID- 12123864 TI - Abstracts of the 11th World Congress of Psychophysiology. Montreal, Quebec, Canada. July 29-August 3, 2002. PMID- 12123863 TI - Effects of intrathecal injection of tamsulosin and naftopidil, alpha-1A and -1D adrenergic receptor antagonists, on bladder activity in rats. AB - The effects of intrathecal injection of tamsulosin (an alpha-1A adrenergic receptor antagonist) and naftopidil (an alpha-1D adrenergic receptor antagonist) on isovolumetric bladder contraction were investigated in rats under urethane anesthesia. Intrathecal injection of tamsulosin (10-30 microg) or naftopidil (0.1 30 microg) transiently abolished isovolumetric rhythmic bladder contraction. Following the recovery of bladder contraction, the interval between contractions was the same as the control value before the injection. The amplitude of bladder contraction was decreased by intrathecal injection of naftopidil (3-30 microg), but not by tamsulosin. Therefore, in addition to the antagonistic action of these agents on the alpha-1 adrenergic receptors of prostatic smooth muscle, both agents (especially naftopidil) may also act on the lumbosacral cord, and thus may improve collecting disorders in patients with benign prostatic hyperplasia. PMID- 12124166 TI - EGF receptor activation by heterologous mechanisms. AB - A myriad of growth factor-dependent and -independent mechanisms for activating the EGF receptor and the related ErbB receptors underscore the importance of receptor inhibitors in the treatment of some solid tumors. PMID- 12124167 TI - Myc-Is this the oncogene from Hell? AB - A new paper implicates the Myc oncoprotein in the direct induction of DNA damage and consequent genome instability in cultured cells. However, it is less clear whether Myc induces the same genetic pandemonium in vivo. PMID- 12124168 TI - Thinking beyond the tumor cell: Nf1 haploinsufficiency in the tumor environment. AB - Deletion of both copies of the Nf1 gene in Schwann cells combined with Nf1 heterozygosity in the tumor environment promotes neurofibroma formation in mice. PMID- 12124169 TI - Breast cancer susceptibility-A new look at an old model. AB - A polygenic model in which many individually weak genes combine multiplicately to cause a 50-fold range of risk in the population explains several puzzling aspects of familial breast cancer epidemiology, including the very high risk in some families and the failure to identify important new genes since the discovery of BRCA1 and BRCA2. PMID- 12124170 TI - Finding the next Gleevec: FLT3 targeted kinase inhibitor therapy for acute myeloid leukemia. AB - Activating mutations in the FLT3 receptor tyrosine kinase occur in 30% of patients with acute myeloid leukemia. Small molecule FLT3 kinase inhibitors show selective antitumor activity in preclinical models. Clinical studies are underway. PMID- 12124171 TI - Focus on acute leukemias. PMID- 12124172 TI - CT53518, a novel selective FLT3 antagonist for the treatment of acute myelogenous leukemia (AML). AB - Up to 30% of acute myelogenous leukemia (AML) patients harbor an activating internal tandem duplication (ITD) within the juxtamembrane domain of the FLT3 receptor, suggesting that it may be a target for kinase inhibitor therapy. For this purpose we have developed CT53518, a potent antagonist that inhibits FLT3, platelet-derived growth factor receptor (PDGFR), and c-Kit (IC(50) approximately 200 nM), while other tyrosine or serine/threonine kinases were not significantly inhibited. In Ba/F3 cells expressing different FLT3-ITD mutants, CT53518 inhibited IL-3-independent cell growth and FLT3-ITD autophosphorylation with an IC(50) of 10-100 nM. In human FLT3-ITD-positive AML cell lines, CT53518 induced apoptosis and inhibited FLT3-ITD phosphorylation, cellular proliferation, and signaling through the MAP kinase and PI3 kinase pathways. Therapeutic efficacy of CT53518 was demonstrated both in a nude mouse model and in a murine bone marrow transplant model of FLT3-ITD-induced disease. PMID- 12124173 TI - Inhibition of mutant FLT3 receptors in leukemia cells by the small molecule tyrosine kinase inhibitor PKC412. AB - Constitutively activating FLT3 receptor mutations have been found in 35% of patients with acute myeloblastic leukemia (AML). Here we report the identification of a small molecule FLT3 tyrosine kinase inhibitor PKC412, which selectively induced G1 arrest and apoptosis of Ba/F3 cell lines expressing mutant FLT3 (IC(50) < 10 nM) by directly inhibiting the tyrosine kinase. Ba/F3-FLT3 cell lines made resistant to PKC412 demonstrated overexpression of mutant FLT3, confirming that FLT3 is the target of this drug. Finally, progressive leukemia was prevented in PKC412-treated Balb/c mice transplanted with marrow transduced with a FLT3-ITD-expressing retrovirus. PKC412 is a potent inhibitor of mutant FLT3 and is a candidate for testing as an antileukemia agent in AML patients with mutant FLT3 receptors. PMID- 12124174 TI - EGFR is a transducer of the urokinase receptor initiated signal that is required for in vivo growth of a human carcinoma. AB - Urokinase plasminogen activator receptor (uPAR) activates alpha5beta1 integrin and ERK signaling, inducing in vivo proliferation of HEp3 human carcinoma. Here we demonstrate that EGFR mediates the uPAR/integrin/fibronectin (FN) induced growth pathway. Its activation is ligand-independent and does not require high EGFR, but does require high uPAR expression. Only when uPAR level is constitutively elevated does EGFR become alpha5beta1-associated and activated. Domain 1 of uPAR is crucial for EGFR activation, and FAK links integrin and EGFR signaling. Inhibition of EGFR kinase blocks uPAR induced signal to ERK, implicating EGFR as an important effector of the pathway. Disruption of uPAR or EGFR signaling reduces HEp3 proliferation in vivo. These findings unveil a mechanism whereby uPAR subverts ligand-regulated EGFR signaling, providing cancer cells with proliferative advantage. PMID- 12124175 TI - HIF activation identifies early lesions in VHL kidneys: evidence for site specific tumor suppressor function in the nephron. AB - Mutations in the von Hippel-Lindau (VHL) gene are associated with hereditary and sporadic clear cell renal carcinoma. VHL acts in a ubiquitin ligase complex regulating hypoxia-inducible factor-1 (HIF-1), but the link between this function and cancer development is unclear. Here we show that in the kidneys of patients with VHL disease, HIF activation is an early event occurring in morphologically normal single cells within the renal tubules. In comparison, dysplastic lesions, cystic lesions, and tumors showed evidence of additional mechanisms that amplify HIF activation. Detection of cells with constitutive HIF activation identified a large number of previously unrecognized foci of VHL inactivation. In proximal tubules these were almost entirely unicellular, whereas multicellular foci were almost exclusively seen in the distal nephron. PMID- 12124176 TI - HNPCC mutations in hMSH2 result in reduced hMSH2-hMSH6 molecular switch functions. AB - Mutations in the human mismatch repair (MMR) gene hMSH2 have been linked to approximately 40% of hereditary nonpolyposis colorectal cancers (HNPCC). While the consequences of deletion or truncating mutations of hMSH2 would appear clear, the detailed functional defects associated with missense alterations are unknown. We have examined the effect of seven single amino acid substitutions associated with HNPCC that cover the structural subdomains of the hMSH2 protein. We show that alterations which produced a known cancer-causing phenotype affected the mismatch-dependent molecular switch function of the biologically relevant hMSH2 hMSH6 heterodimer. Our observations demonstrate that amino acid substitutions within hMSH2 that are distant from known functional regions significantly alter biochemical activity and the ability of hMSH2-hMSH6 to form a sliding clamp. PMID- 12124178 TI - p53 stabilization is decreased upon NFkappaB activation: a role for NFkappaB in acquisition of resistance to chemotherapy. AB - Chemotherapeutic agents simultaneously induce transcription factors p53 and NFkappaB. p53 induction can activate an apoptotic program, and resistance to chemotherapy correlates with the loss of a functional p53 pathway. By contrast, NFkappaB prevents apoptosis in response to chemotherapeutic agents. We have analyzed the p53 response in IKK1/2(-/-) MEFs, which lack detectable NFkappaB activity. Compared to WT fibroblasts, IKK1/2(-/-) fibroblasts showed increased cell death and p53 induction in response to the chemotherapeutic agent, doxorubicin. Reconstitution of IKK2, but not IKK1, increased Mdm2 levels and decreased doxorubicin-induced p53 stabilization and cell death. IKK2-mediated effects required its kinase function and were abrogated by coexpression of the dominant negative IkappaBalphaM, implying a role for NFkappaB in blocking chemotherapy-induced p53 and cell death. PMID- 12124179 TI - The right-handed slanting of Arabidopsis thaliana roots is due to the combined effects of positive gravitropism, circumnutation and thigmotropism. AB - When primary Arabidopsis roots grow down a tilted agar plate, they do not elongate following the gravitational vector along a straight line, but instead they slant noticeably to the right-hand. This process is seen mostly in the ecotypes Wassilewskjia and Landsberg, whereas it is attenuated in the ecotype Columbia, and in some mutants is even inverted. The origin of the slanting of Arabidopsis roots, that evidently constitutes a form of chirality, has so far not been sufficiently clarified. In the present paper we describe it as the general result of the cumulative effects of positive gravitropism, circumnutation and a thigmotropic obstacle-avoiding movement, and in particular, as the consequence of the alternating movement of circumnutation of the root to the right and to the left of the gravitational vector. This movement, which does not appear symmetrical in its nature, since the waves made to the right-hand are complete whereas those made to the left-hand are reduced or aborted, appears to be the reason for the observed slanting. In addition, evidence is furnished supporting the hypothesis that the strong left-handed cell-files torsion, seen in right handed coiling roots, is not the consequence of a primary process, but of an artifact, and is due to the adjustment of the three dimensional root circumnutating helix to the flat two dimensional agar surface. PMID- 12124177 TI - Critical role for Gab2 in transformation by BCR/ABL. AB - The BCR/ABL oncogene causes chronic myelogenous leukemia (CML) in humans and a CML-like disease, as well as lymphoid leukemia, in mice. p210 BCR/ABL is an activated tyrosine kinase that phosphorylates itself and several cellular signaling proteins. The autophosphorylation site tyrosine 177 binds the adaptor Grb2 and helps determine the lineage and severity of BCR/ABL disease: Tyr177 mutation (BCR/ABL-Y177F) dramatically impairs myeloid leukemogenesis, while diminishing lymphoid leukemogenesis. The critical signal(s) from Tyr177 has remained unclear. We report that Tyr177 recruits the scaffolding adaptor Gab2 via a Grb2/Gab2 complex. Compared to BCR/ABL-expressing Ba/F3 cells, BCR/ABL-Y177F cells exhibit markedly reduced Gab2 tyrosine phosphorylation and association of phosphatidylinositol-3 kinase (PI3K) and Shp2 with Gab2 and BCR/ABL, and decreased PI3K/Akt and Ras/Erk activation, cell proliferation, and spontaneous migration. Remarkably, bone marrow myeloid progenitors from Gab2 (-/-) mice are resistant to transformation by BCR/ABL, whereas lymphoid transformation is diminished as a consequence of markedly increased apoptosis. BCR/ABL-evoked PI3K/Akt and Ras/Erk activation also are impaired in Gab2 (-/-) primary myeloid and lymphoid cells. Our results identify Gab2 and its associated proteins as key determinants of the lineage and severity of BCR/ABL transformation. PMID- 12124180 TI - Bone biochemistry in rat femoral diaphysis after space flight. AB - The aim of this experiment was to identify the location of the biochemical changes associated with depressed mineralization during space flight. We carried out biochemical analysis of 4 sections of the femoral diaphyses from 107 day old male rats flown aboard Cosmos 2044 Biosatellite for 16 days. Control femurs were preflight, vivarium, synchronous for feed, cage and temperature exposure, and a flight simulation model. Distal sections in both the flight and synchronous femurs showed mineral deficits associated with reduced levels of the reducible cross-link product of type I collagen, dehydro-dihydroxylysinonorleucine (deH DHLNL) (p<.05). Unloaded bones in the ground based flight simulation model showed changes in cross-links similar to flight and synchronous controls, but were not associated with the mineral deficit. Mean values of elements measured in each section of all groups revealed significant associations (p<.005) between the non collagenous protein, osteocalcin and calcium (r=0.774), phosphorus (r=-.624) and deH-DHLNL/deH-HLNL (r=.883). The ratio of the nonreducible cross-link, pyridinoline, to its lysl analogue, deoxypyridinoline, was consistently lower in the distal than proximal sections of the groups tested. None of the changes during space flight were unique to flight bone. The most significant and extensive changes in bone composition, i.e. mineral deficits associated with changes in both osteocalcin and reducible cross-links, were located in the distal section of the diaphysis of the femur. PMID- 12124181 TI - Survival and growth of developing rats during centrifugation at 2G. AB - We studied the effects of 2G hypergravity on the survival, body mass and growth of postnatal rats (Rattus norvegicus). Nursing litters comprised of either neonatal (Postnatal day [P]7) or pre-weanling (P14) rats and their mothers were exposed to 16 days of continuous centrifugation. All of the offspring survived and gained body mass, indicating that mothers nursed their young. Following the onset of centrifugation, neonatal and pre-weanling rats showed a reduction in growth relative to age-matched environmental controls (EC). At the completion of testing, body mass of the hypergravity (HG) groups was significantly less than that of controls (p<0.05). Over the course of the test, the HG-exposed P7 group showed an overall 55% gain in body mass as compared to a 71% increase in controls, while the HG-exposed P14 group showed a 62% increase relative to 75% in controls. Neonatal offspring (P7) gained body mass during centrifugation, but at significantly slower rates as compared to EC controls (p<0.05). In contrast, growth rates of pre-weanling (P14) rats were not reduced relative to controls, possibly related to the initiation of weaning, around P18 in the rat. These findings raise key issues relevant to studies of nursing mammals reared in altered gravity. PMID- 12124182 TI - Salt-loading and simulated microgravity on baroreflex responsiveness in rats. AB - Cardiovascular adaptations observed during exposure to microgravity results in impairment of baroreflex activity partially as a result of fluid and electrolyte shifts. The head-down tilt rat model mimics some of the physiological observations that have been made in astronauts. We examined the effects of salt loading on baroreflex activity after 7 day simulated microgravity (30 degrees tail-suspension) and the subsequent 6 hr post-suspension in Sprague-Dawley (SD) rats, using low salt (0.3% NaCl) and high salt (8% NaCl) diets. In suspended animals on a low salt diet, the baroreflex response curve was shifted to the left, while the heart rate (HR) range and MAP50 values were reduced compared to their parallel tethered, non-suspended controls. For non-suspended animals, salt loading shifted the curve to the right with a reduced HR range. In salt-loaded, suspended animals, the curve and its parameters resemble those of non-suspended animals on a low salt diet. In summary, these data have demonstrated that a short term (seven days) simulated weightlessness may elicit cardiovascular deconditioning in rats after release from the simulation manifested as an altered responsiveness in baroreceptor-heart rate reflex and a lowered blood pressure while the rats are tethered and horizontal. Our results also suggest the counteracting effect of salt loading on cardiovascular deconditioning. PMID- 12124183 TI - The effects of hypergravity and substrate vibration on vestibular function in developing chickens. AB - We used linear vestibular evoked potentials (VsEPs) to characterize peripheral and central vestibular function in birds following embryogenesis at 2G centrifugation or at elevated levels of vibration (+20dB re: background levels). Additionally, we characterized peripheral and central vestibular adaptation to 2G centrifugation in early post-hatch birds. Linear VsEP response peak latencies, amplitudes, thresholds and input/output functions were quantified and compared between experimental and control animals. Birds vibrated throughout embryogenesis and up to one-week post-hatch revealed no changes in linear VsEP response components compared to control siblings. Birds centrifuged at 2G throughout embryogenesis also evidenced no changes in the linear VsEP measured at hatch (P0). Significant changes were seen, however, for linear VsEPs of post-hatch birds placed at 2G for 7 days beginning on post-hatch day 5. Linear VsEPs for these animals displayed significant reductions in response amplitudes associated with peaks P2, N2 and P3, response peaks generated by central neural relays of gravity receptors. The earliest response components, generated by the peripheral vestibular nerve (i.e., P1, N1), were not significantly altered with the 7-day exposure to 2G. Thus, there was no evidence of generalized changes in peripheral gravity receptor excitability or in the rate of maturation in developing animals under increased levels of gravity or vibration. If gravity level plays a critical role in shaping peripheral vestibular ontogeny at magnitudes between 1 and 2G, then it may serve to stabilize function under changing G-fields or it may operate on physiological features that can not be resolved by the VsEP. In contrast, exposure to elevated gravity during post-hatch periods does alter central vestibular function thus providing direct evidence for central vestibular adaptation to the gravitational environment. The fact that central functional change was observed in hatchlings and not embryos, raises the possibility that the first 2-weeks post-hatch may be a critical period of "heightened developmental sensitivity" to hypergravity. PMID- 12124184 TI - Neural and neuroendocrine adaptations to microgravity and ground-based models of microgravity. AB - The functional properties of the motor system of humans and non-human primates are readily responsive to microgravity. There is a growing body of evidence that significant adaptations occur in the spinal cord and muscle in response to prolonged exposure to microgravity. Further, there is evidence that the processing of sensory information from the periphery, particularly that input associated with the function of muscle tendons and joints, is significantly altered as a result of prolonged microgravity. We present evidence that the fundamental neural mechanisms that control the relative activity of the motor pools of a slow and fast extensor muscle is changed such that a slow, postural muscle is less readily activated during locomotion following spaceflight. Another type of change observed in mammals exposed to spaceflight relates to the release of a growth factor, called bioassayable growth hormone, which is thought to be released from the pituitary. When an individual generates a series of isometric plantarflexor contractions, the plasma levels of bioassayable growth hormone increases significantly. This response is suppressed after several days of continuous bedrest or spaceflight. These results suggest a unique neuroendocrine control system and demonstrate its sensitivity to chronic patterns of proprioceptive input associated with load-bearing locomotion. PMID- 12124185 TI - Changes in muscle size and architecture following 20 days of bed rest. AB - Five healthy men carried out a program of head-down bed rest (BR) for 20 days. Before and after BR, a series of cross-sectional scans of the thigh were performed using magnetic resonance imaging, from which volumes of the quadriceps muscles were determined and physiological cross-sectional areas (PCSA) were calculated. Muscle thickness and pennation angles of the triceps brachii, vastus lateralis, and triceps surae muscles were also determined by ultrasonography. During BR, subjects performed unilateral isokinetic knee extension exercises every day. The contralateral limb served as a control. Decrease in PCSA after BR was greater in the control (-10.2 +/- 6.3%) than in the trained limb (-5.2 +/- 4.2%). Among the quadriceps, vastus intermedius in the control limb was predominantly atrophied by BR with respect to the volume and PCSA, and the rectus femoris showed the greatest training effect and retained its size in the trained limb. Decreases in muscle thicknesses in leg muscles were not prevented by the present exercise protocol, suggesting a need for specific exercise training for these muscles. Neither trained nor control muscles showed significant changes in pennation angles in any muscles after BR, suggesting that muscle architecture does not change remarkably by muscle atrophy by up to 10%. PMID- 12124187 TI - Fos protein expression in the medulla oblongata and changes in size of spinal lateral horn neurons after 4-wk simulated weightlessness in rats. AB - Responses of the neurons in medulla oblongata and C8-T1 spinal cord lateral horn of rats induced by simulated weightlessness were investigated using anti-Fos protein and anti-tyrosine hydroxylase (TH) double staining immunohistochemical methods, and Nissl-staining technique respectively. After four weeks of tail suspension, many Fos-like positive neurons were localized in the medullary visceral zone (MVZ), predominantly in the nucleus of tractus solitarii and ventrolateral medulla, and some of them showed TH-like immunoreactivity. Sizes of the cell bodies of the lateral horn neurons in C8-T1 segment were significantly increased in 4-wk tail-suspended rats (P<0.05) as compared with that in controls. The results suggest that the neurons in MVZ and the spinal lateral horn may be involved in the adaptation of central cardiovascular regulation during weightlessness. PMID- 12124186 TI - Effects of 2G exposure on lean and genetically obese Zucker rats. AB - Changes in the ambient force environment alter the regulation of adiposity, food intake and energy expenditure (i.e., energy balance). Lean (Fa/Fa) and obese (fa/fa) male Zucker rats were exposed to 2G (twice Earth's normal gravity) for eight weeks via centrifugation to test the hypothesis that the Fa/Fa rats recover to a greater degree from the effects of an increased ambient force environment on body mass and food intake, than do the fa/fa rats which have a dysfunctional leptin regulatory system. The rats (lean and obese exposed to either 1G or 2G) were individually housed in standard vivarium cages with food and water provided ad libitum. The acute response to 2G included a transient hypophagia accompanied by decreased body mass, followed by recovery of feeding to new steady-states. In the lean rats, body mass-independent food intake had returned to 1G control levels six weeks after the onset of centrifugation, and body mass increased towards that of the 1G rats. In contrast, food intake and body mass of the 2G obese rats plateaued at a level lower than that of the 1G controls. Although percent carcass fat was reduced more in the 2G leans vs. 2G obese rats, the latter lost significantly more grams of fat than did the leans. Our data suggest that with respect to food intake and body mass, the lean rats recover from the initial effects of 2G exposure to a greater degree than do the fatty rats, a difference that likely reflects the functionality of the leptin regulatory system in the leans. PMID- 12124188 TI - The effect of 2G on mouse circadian rhythms. AB - This study examined the effect of chronic 2G exposure on the regulation of body temperature (T(b)), activity (ACT), and circadian rhythms of mice. Five mice were implanted with biotelemetry units to record T(b) and ACT. The mice exhibited a stable daily mean of T(b) (37.1 +/- 2.1 degrees C) and ACT and robust circadian rhythms during the control 1G period. Mice exhibited a significant decline in T(b) (30.1 +/- 1.5 degrees C; t(4)=8.32, p<.01) and cessation of ACT within two hours following 2G onset. After 6 hours of continuous 2G exposure there was a recovery in T(b) (34.4 +/- 1.6 degrees C) that remained significantly below that of baseline (t(4)=3.66, p<.05). A similar pattern of recovery was seen following 12 hours of continuous 2G for ACT. A slower pattern of adaptation toward baseline levels occurred steadily over the next 6-13 days. Exposure to 2G also caused an immediate 4 day loss in circadian rhythm amplitude in both T(b) and ACT. Recovery to new steady state levels was achieved by 8 days and 13 days, respectively. These results demonstrate that under chronic 2G, the recovery time for the homeostatic steady-state values and circadian rhythms are shorter for the mouse than for the rat. These differences may be related to the scaling effects of 2G resulting from the mass difference between mice and rats. PMID- 12124189 TI - Lipopolysaccharide induces apoptosis in adult rat ventricular myocytes via cardiac AT(1) receptors. AB - Lipopolysaccharide (LPS) from gram-negative bacteria circulates in acute, subacute, and chronic conditions. It was hypothesized that LPS directly induces cardiac apoptosis. In adult rat ventricular myocytes (isolated with depyrogenated digestive enzymes to minimize tolerance), LPS (10 ng/ml) decreased the ratio of Bcl-2 to Bax at 12 h; increased caspase-3 activity at 16 h; and increased annexin V, propidium iodide, and terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling staining at 24 h. Apoptosis was blocked by the caspase inhibitor benzyloxycarbonyl-valine-alanine-aspartate fluoromethylketone (Z-VAD-fmk), captopril, and angiotensin II type 1 receptor (AT(1)) inhibitor (losartan), but not by inhibitors of AT(2) receptors (PD-123319), tumor necrosis factor-alpha (TNFRII:Fc), or nitric oxide (N(G)-monomethyl-L-arginine). Angiotensin II (100 nmol/l) induced apoptosis similar to LPS without additive effects. LPS in vivo (1 mg/kg iv) increased apoptosis in left ventricular myocytes for 1-3 days, which dissipated after 1-2 wk. Losartan (23 mg. kg(-1). day(-1) in drinking water for 3 days) blocked LPS-induced in vivo apoptosis. In conclusion, low levels of LPS induce cardiac apoptosis in vitro and in vivo by activating AT(1) receptors in myocytes. PMID- 12124190 TI - Improved angiogenic potency by implantation of ex vivo hypoxia prestimulated bone marrow cells in rats. AB - Therapeutic angiogenesis can be induced by local implantation of bone marrow cells. We tried to enhance the angiogenic potential of this treatment by ex vivo hypoxia stimulation of bone marrow cells before implantation. Bone marrow cells were collected and cultured at 33 degrees C under 2% O(2)-5% CO(2)-90% N(2) (hypoxia) or 95% air-5% CO(2) (normoxia). Cells were also injected into the ischemic hindlimb of rats after 24 h of culture. Hypoxia culture increased the mRNA expression of vascular endothelial growth factor (VEGF), vascular endothelial (VE)-cadherin, and fetal liver kinase-1 (Flk-1) from 2.5- to fivefold in bone marrow cells. The levels of VEGF protein in the ischemic hindlimb were significantly higher 1 and 3 days after implantation with hypoxia-cultured cells than with normoxia-cultured or noncultured cells. The microvessel density and blood flow rate in the ischemic hindlimbs were also significantly (P < 0.001) higher 2 wk after implantation with hypoxia-cultured cells (89.7 +/- 5.5%) than with normoxia-cultured cells (67.0 +/- 9.6%) or noncultured cells (70.4 +/- 7.7%). Ex vivo hypoxia stimulation increased the VEGF mRNA expression and endothelial differentiation of bone marrow cells, which together contributed to improved therapeutic angiogenesis in the ischemic hindlimb after implantation. PMID- 12124191 TI - H(2)S-induced vasorelaxation and underlying cellular and molecular mechanisms. AB - H(2)S is endogenously generated in vascular smooth muscle cells. The signal transduction pathways involved in the vascular effects of H(2)S have been unclear and were investigated in the present study. H(2)S induced a concentration dependent relaxation of rat aortic tissues that was not affected by vascular denervation. The vasorelaxant potency of H(2)S was attenuated by the removal of the endothelium. Similarly, the blockade of nitric oxide synthase or the coapplication of the Ca(2+)-dependent K(+) channel blockers apamin and charybdotoxin reduced the H(2)S-induced relaxation of the endothelium-intact aortic tissues. Sodium nitroprusside (SNP)-induced relaxation was completely abolished by either 1H-[1,2,4]oxadiazolo-[4,3-a]quinoxalin-1-one (ODQ) or NS- 2028, two soluble guanylate cyclase inhibitors. Instead of inhibition, ODQ and NS 2028 potentiated the H(2)S-induced vasorelaxation, which was suppressed by superoxide dismutase. The vasorelaxant effect of H(2)S was also significantly attenuated when Ca(2+)-free bath solution was used. Finally, pretreatment of aortic tissues with H(2)S reduced the relaxant response of vascular tissues to SNP. Our results demonstrate that the vascular effect of H(2)S is partially mediated by a functional endothelium and dependent on the extracellular calcium entry but independent of the activation of the cGMP pathway. PMID- 12124192 TI - Ca(2+) loading and adrenergic stimulation reveal male/female differences in susceptibility to ischemia-reperfusion injury. AB - To compare ischemia-reperfusion injury in males versus females under hypercontractile conditions, perfused hearts from 129J mice pretreated with 3 mmol/l Ca(2+) or 10(-8) mol/l isoproterenol +/- 10(-6) mol/l N(omega)-nitro-L arginine methyl ester (L-NAME) were subjected to 20 min of ischemia and 40 min of reperfusion while (31)P NMR spectra were acquired. Basal contractility increased equivalently in female versus male hearts with isoproterenol- or Ca(2+) treatment. Injury was equivalent in untreated male versus female hearts but was greater in isoproterenol or Ca(2+)-treated male than female hearts, as indicated by lower postischemic contractile function, ATP, and PCr. Endothelial nitric oxide (NO) synthase (eNOS) expression was higher in female than male hearts, neuronal NOS (nNOS) did not differ, and inducible NOS (iNOS) was undetectable. Ischemic NO production was higher in female than male hearts, and L-NAME increased injury in female isoproterenol-treated hearts. In summary, isoproterenol or high Ca(2+) pretreatment increased ischemia-reperfusion injury in males more than females. eNOS expression and NO production were higher in female than male hearts, and L-NAME blocked female protection. Females were therefore protected from the detrimental effects of adrenergic stimulation and Ca(2+) loading via a NOS-mediated mechanism. PMID- 12124193 TI - Mechanistic investigation of extracellular K+ accumulation during acute myocardial ischemia: a simulation study. AB - In this study, we have used computer simulations to study the mechanisms of extracellular K+ accumulation during acute ischemia. A modified version of the Luo-Rudy phase II action potential model was used to simulate the electrical behavior of one ventricular myocyte during 14 min of simulated ischemia. Our results show the following: 1) only the integrated effect of activation of ATP dependent K+ current, an ischemic Na+ inward current, and inhibition of Na(+) K(+) pump activity in the absence of coronary flow replicates the biphasic time course of extracellular K+ concentration observed during acute ischemia; 2) the time to onset of the plateau phase and the plateau level value are determined by the rate of stimulation and by the rate of alteration of the three mechanisms. However, acidosis and reduction of extracellular volume produce only a slight anticipation of the plateau phase; and 3) cellular K+ loss is mainly due to an increase of K+ efflux via the time-independent K+ current and ATP-dependent K+ current rather than to a decrease of K+ influx. PMID- 12124194 TI - Sympathoexcitation to intravenous interleukin-1beta is dependent on forebrain neural circuits. AB - We investigated the contributions of forebrain, brain stem, and spinal neural circuits to interleukin (IL)-1beta-induced sympathetic nerve discharge (SND) responses in alpha-chloralose-anesthetized rats. Lumbar and splenic SND responses were determined in spinal cord-transected (first cervical vertebra, C1), midbrain transected (superior colliculus), and sham-transected rats before and for 60 min after intravenous IL-1beta (285 ng/kg). The observations made were the following: 1) lumbar and splenic SND were significantly increased after IL-1beta in sham C1 transected rats but were unchanged after IL-1beta in C1-transected rats; 2) intrathecal administration of DL-homocysteic acid (10 ng) increased SND in C1 transected rats; 3) lumbar and splenic SND were significantly increased after IL 1beta in sham- but not midbrain-transected rats; and 4) midbrain transection did not alter the pattern of lumbar and splenic SND, demonstrating the integrity of brain stem sympathetic neural circuits after decerebration. These results demonstrate that an intact forebrain is required for mediating lumbar and splenic sympathoexcitatory responses to intravenous IL-1beta, thereby providing new information about the organization of neural circuits responsible for mediating sympathetic-immune interactions. PMID- 12124195 TI - Rate-dependent [K+](o) accumulation in canine right atria in vivo: electrophysiological consequences. AB - Sudden increases in heart rate cause accumulation of K+ in the extracellular space. However, the exact relationship between rate and extracellular K+ concentration ([K+](o)) in vivo is unknown. We measured [K+](o) in right atria of anesthetized dogs by using K(+)-sensitive electrodes. Peak increase in [K+](o) ranged from 0.18 +/- 0.04 mM [means +/- SE; cycle length (CL) = 350 ms] to 0.80 +/- 0.09 mM (CL = 250 ms) above baseline (3.50 +/- 0.08 mM at CL = 380 ms; n = 5). During rapid pacing-induced atrial fibrillation, peak increase in [K+](o) averaged 0.80 +/- 0.07 mM (n = 5). Whole cell current-clamp measurements in single right atrial myocytes (n = 5) showed that raising [K+](o) from 3 to 5 mM in 1-mM steps progressively depolarized resting membrane potential and reduced both phase 0 action potential amplitude and maximal upstroke velocity. Multisite epicardial mapping (n = 4) demonstrated that sudden rate increases changed longitudinal conduction velocity (CV(L)) by -3.6 +/- 1.8% to -5.9 +/- 1.2% over a CL range of 330 to 250 ms. Our observations suggest that rate-related [K+](o) accumulation in vivo is of sufficient magnitude to modulate those cellular electrophysiological properties that determine atrial CV(L). PMID- 12124196 TI - Cause and effect relationship between myocardial mast cell number and matrix metalloproteinase activity. AB - The objectives of this study were to investigate the temporal response of left ventricular (LV) matrix metalloproteinase (MMP) activity and collagen volume fraction (CVF) induced by an aortocaval fistula and the role of cardiac mast cells in regulating MMP activity. LV tissue was analyzed for MMP activity, CVF, and mast cell number in rats euthanized at 0.5, 1, 2, 3, 5, 14, 21, 35, and 56 days. Additional rats treated with the mast cell membrane-stabilizing drug cromolyn sodium were euthanized 1, 2, and 3 days postfistula. Marked increases in MMP activity occurred rapidly and remained significantly elevated for 5 days before returning toward normal. A significant decrease in CVF occurred by day 5, but thereafter CVF rebounded to normal or above normal values. The number of myocardial mast cells also significantly increased postfistula, and there was a close association between mast cell density and MMP activity. Cromolyn treatment prevented the increase in mast cell number and MMP activity. Thus it is concluded that cardiac mast cells play a major role in the regulation of MMP activity. PMID- 12124197 TI - Muscle metaboreflex control of coronary blood flow. AB - We investigated the effect of muscle metaboreflex activation on left circumflex coronary blood flow (CBF) and vascular conductance (CVC) in conscious, chronically instrumented dogs during treadmill exercise ranging from mild to severe workloads. Metaboreflex responses were also observed during mild exercise with constant heart rate (HR) of 225 beats/min and beta(1)-adrenergic receptor blockade to attenuate the substantial reflex increases in cardiac work. The muscle metaboreflex was activated via graded partial occlusion of hindlimb blood flow. During mild exercise, with muscle metaboreflex activation, hindlimb ischemia elicited significant reflex increases in mean arterial pressure (MAP), HR, and cardiac output (CO) (+39.0 +/- 5.2 mmHg, +29.9 +/- 7.7 beats/min, and +2.0 +/- 0.4 l/min, respectively; all changes, P < 0.05). CBF increased from 51.9 +/- 4.3 to 88.5 +/- 6.6 ml/min, (P < 0.05), whereas no significant change in CVC occurred (0.56 +/- 0.06 vs. 0.59 +/- 0.05 ml. min(-1). mmHg(-1); P > 0.05). Similar responses were observed during moderate exercise. In contrast, with metaboreflex activation during severe exercise, no further increases in CO or HR occurred, the increases in MAP and CBF were attenuated, and a significant reduction in CVC was observed (1.00 +/- 0.12 vs. 0.90 +/- 0.13 ml. min(-1). mmHg( 1); P < 0.05). Similarly, when the metaboreflex was activated during mild exercise with the rise in cardiac work lessened (via constant HR and beta(1) blockade), no increase in CO occurred, the MAP and CBF responses were attenuated (+15.6 +/- 4.5 mmHg, +8.3 +/- 2 ml/min), and CVC significantly decreased from 0.63 +/- 0.11 to 0.53 +/- 0.10 ml. min(-1). mmHg(-1). We conclude that the muscle metaboreflex induced increases in sympathetic nerve activity to the heart functionally vasoconstricts the coronary vasculature. PMID- 12124198 TI - Direct biomechanical induction of endogenous calcineurin inhibitor Down Syndrome Critical Region-1 in cardiac myocytes. AB - Signaling through the protein phosphatase calcineurin may play a critical role in cardiac hypertrophy. The gene for Down Syndrome Critical Region-1 (DSCR1) encodes a protein that is an endogenous calcineurin inhibitor. This study was designed to test the hypothesis that DSCR1 is directly induced by biomechanical stimuli. Neonatal rat cardiac myocytes were exposed to biaxial cyclic mechanical strain; mechanical strain upregulated DSCR1 mRNA expression in a time- and amplitude dependent manner (3.4 +/- 0.2-fold at 8% strain for 6 h, n = 11, P < 0.01), and this induction was angiotensin II and endothelin I independent. Biomechanical induction of DSCR1 mRNA was partially blocked by calcineurin inhibition with cyclosporine A (30 +/- 5%, n = 3, P < 0.01). DSCR1 promoter-reporter experiments showed that mechanical strain induced DSCR1 promoter activity by 2.3-fold and that this induction was completely inhibited by cyclosporin A. Furthermore, DSCR1 gene expression was increased in the left ventricles of mice with pressure overload hypertrophy induced by transverse aortic banding. These data demonstrate that biomechanical strain directly induces gene expression for the calcineurin inhibitor DSCR1 in cardiac myocytes, indicating that mechanically induced DSCR1 may regulate the hypertrophic response to mechanical overload. PMID- 12124199 TI - Bioenergetic remodeling of heart during treatment of spontaneously hypertensive rats with enalapril. AB - We used spontaneously hypertensive rats to study remodeling of cardiac bioenergetics associated with changes in blood pressure. Blood pressure was manipulated with aggressive antihypertensive treatment combining low dietary salt and the angiotensin-converting enzyme inhibitor enalapril. Successive cycles of 2 wk on, 2 wk off treatment led to rapid, reversible changes in left ventricular (LV) mass (30% change in <10 days). Despite changes in LV mass, specific activities of bioenergetic (cytochrome-c oxidase, citrate synthase, lactate dehydrogenase) and reactive oxygen species (ROS) (total cellular superoxide dismutase) enzymes were actively maintained within relatively narrow ranges regardless of treatment duration, organismal age, or transmural region. Although enalapril led to parallel declines in mitochondrial enzyme content and ventricular mass, total ventricular mtDNA content was unaffected. Altered enzymatic content occurred without significant changes in relevant mRNA and protein levels. Transcript levels of gene products involved in mtDNA maintenance (Tfam), mitochondrial protein degradation (LON protease), fusion (fuzzy onion homolog), and fission (dynamin-like protein, synaptojanin-2alpha) were also unchanged. In contrast, enalapril-mediated ventricular and mitochondrial remodeling was accompanied by a twofold increase in specific activity of catalase, an indicator of oxidative stress, suggesting that rapid cardiac adaptation is accompanied by tight regulation of mitochondrial enzyme activities and increased ROS production. PMID- 12124200 TI - Role of Src protein tyrosine kinases in late preconditioning against myocardial infarction. AB - Although Src protein tyrosine kinases (PTKs) have been shown to be essential in late preconditioning (PC) against myocardial stunning, their role in triggering versus mediating late PC against myocardial infarction remains unclear. Four groups of conscious rabbits were subjected to a 30-min coronary occlusion on day 2, with or without PC ischemia on day 1. Administration of the Src PTK inhibitor lavendustin A (LD-A; 1 mg/kg iv) before the PC ischemia on day 1 (group III, n = 7) failed to block the delayed protective effect against myocardial infarction 24 h later. Late PC against infarction, however, was completely abrogated when LD-A was given 24 h after the PC ischemia, prior to the 30-min occlusion on day 2 (group IV, n = 8). We conclude that, in conscious rabbits, Src PTK activity is necessary for the mediation of late PC protection against myocardial infarction on day 2, but not for the initiation of this phenomenon on day 1. Taken together with previous studies in the setting of stunning, these findings reveal heretofore unrecognized differences in the roles of Src PTKs in late PC against stunning versus late PC against infarction. PMID- 12124201 TI - ecNOS gene polymorphism is associated with endothelium-dependent vasodilation in Type 2 diabetes. AB - The association between endothelial constitutive nitric oxide synthase (ecNOS) gene polymorphism and vascular endothelial function has not been clarified. We investigated the impact of ecNOS gene polymorphism on endothelial function in 95 patients with Type 2 diabetes (ecNOS genotype: 4b/b, n = 62; 4b/a, n = 30; 4a/a, n = 3). Flow-mediated (endothelium dependent, FMD) and nitroglycerin-induced (endothelium independent, NTG) vasodilations of the right brachial artery were studied using a phase-locked echotracking system. There were no significant differences in clinical characteristics among the ecNOS genotypes. The FMD was significantly lower in the patients with ecNOS4a allele than in those without ecNOS4a allele (P < 0.05). Multiple regression analysis showed that ecNOS4a allele and mean blood pressure were significant independent determinants for reduced FMD in all patients (R(2) = 0.122, P = 0.0025). The ecNOS4a allele was an independent determinant for reduced FMD in smokers but not in nonsmokers. These results suggest that ecNOS4a allele is a genetic risk factor for endothelial dysfunction in diabetic patients, especially in smokers. PMID- 12124202 TI - Relationship among hyperinsulinemia, insulin resistance, and hypertension is dependent on sex. AB - Hyperinsulinemia and insulin resistance have been linked to hypertension; however, the influence of sex on this relationship has not been well studied. The purpose of this experiment was to compare the effects of chronic insulin treatment on insulin sensitivity and blood pressure in male and female rats. Male and female Wistar rats were treated with insulin (2 U/day) via subcutaneous sustained release implants for 5 wk. Systolic blood pressure was measured via the tail-cuff method before and after treatment, and insulin sensitivity was assessed with an oral glucose tolerance test. The insulin sensitivity of female rats was 4.5-fold greater than male rats. Chronic insulin treatment impaired insulin sensitivity in both sexes; however, this occurred to a greater degree in male rats. Blood pressure increased in male rats treated with insulin only. The results demonstrate that hyperinsulinemia and insulin resistance are associated with hypertension in male rats only. Therefore, the link between these conditions appears to depend on sex. PMID- 12124203 TI - Contribution of endothelin to pulmonary vascular tone under normoxic and hypoxic conditions. AB - The contribution of endothelin to resting pulmonary vascular tone and hypoxic pulmonary vasoconstriction in humans is unknown. We studied the hemodynamic effects of BQ-123, an endothelin type A receptor antagonist, on healthy volunteers exposed to normoxia and hypoxia. Hemodynamics were measured at room air and after 15 min of exposure to hypoxia (arterial PO(2) 99.8 +/- 1.8 and 49.4 +/- 0.4 mmHg, respectively). Measurements were then repeated in the presence of BQ-123. BQ-123 decreased pulmonary vascular resistance (PVR) 26% and systemic vascular resistance (SVR) 21%, whereas it increased cardiac output (CO) 22% (all P < 0.05). Hypoxia raised CO 28% and PVR 95%, whereas it reduced SVR 23% (all P < 0.01). During BQ-123 infusion, hypoxia increased CO 29% and PVR 97% and decreased SVR 22% (all P < 0.01). The pulmonary vasoconstrictive response to hypoxia was similar in the absence and presence of BQ-123 [P = not significant (NS)]. In vehicle-treated control subjects, hypoxic pulmonary vasoconstriction did not change with repeated exposure to hypoxia (P = NS). Endothelin contributes to basal pulmonary and systemic vascular tone during normoxia, but does not mediate the additional pulmonary vasoconstriction induced by acute hypoxia. PMID- 12124204 TI - Overexpression of phospholemman alters contractility and [Ca(2+)](i) transients in adult rat myocytes. AB - Previous studies showed increased phospholemman (PLM) mRNA after myocardial infarction (MI) in rats (Sehl PD, Tai JTN, Hillan KJ, Brown LA, Goddard A, Yang R, Jin H, and Lowe DG. Circulation 101: 1990-1999, 2000). We tested the hypothesis that, in normal adult rat cardiac myocytes, PLM overexpression alters contractile function and cytosolic Ca(2+) concentration ([Ca(2+)](i)) homeostasis in a manner similar to that observed in post-MI myocytes. Compared with myocytes infected by control adenovirus expressing green fluorescent protein (GFP) alone, Western blots indicated a 41% increase in PLM expression after 72 h (P < 0.001) but no changes in Na(+)/Ca(2+) exchanger, SERCA2, and calsequestrin levels in myocytes infected by adenovirus expressing GFP and PLM. At 5 mM extracellular [Ca(2+)] ([Ca(2+)](o)), maximal contraction amplitudes in PLM-overexpressed myocytes were 24% (P < 0.005) and [Ca(2+)](i) transient amplitudes were 18% (P < 0.05) lower than control myocytes. At 0.6 mM [Ca(2+)](o), however, contraction and [Ca(2+)](i) transient amplitudes were significantly (P < 0.05) higher in PLM overexpressed than control myocytes (18% and 42%, respectively); at 1.8 mM [Ca(2+)](o), the differences in contraction and [Ca(2+)](i) transient amplitudes were narrowed. This pattern of contractile and [Ca(2+)](i) transient abnormalities in PLM-overexpressed myocytes mimics that observed in post-MI rat myocytes. We suggest that PLM overexpression observed in post-MI myocytes may partly account for contractile abnormalities by perturbing Ca(2+) fluxes during excitation-contraction. PMID- 12124206 TI - Reactive carbonyls from tobacco smoke increase arterial endothelial layer injury. AB - We hypothesized that reactive carbonyls generated from smoke exposure cause increased arterial low-density lipoprotein (LDL) accumulation and endothelial layer permeability. In addition, we hypothesized that estrogen supplementation was protective against chronic environmental tobacco smoke (ETS) exposure to the artery wall. Quantitative fluorescence microscopy was used to determine artery injury after exposure. For our chronic studies, ovariectomized rats treated with subcutaneous placebo or 17beta-estradiol pellets were exposed to ETS or filtered air for 6 wk. ETS exposure increased carotid artery LDL accumulation more than fourfold compared with filtered air exposure, an effect largely mediated by increased permeability. No protective effect of estradiol was observed. Acute ETS exposure of a buffer solution containing LDL resulted in a more than sixfold increase in the highly reactive carbonyl glyoxal. Perfusion of this solution through carotid arteries resulted in a 105% increase in permeability. Moreover, perfusion of glyoxal alone caused a 50% increase in carotid artery permeability. This endothelial damage and changes in lipid accumulation may serve as an initiating event in atheroma formation in individuals exposed to ETS. PMID- 12124205 TI - Contractility and ischemic response of hearts from transgenic mice with altered sarcolemmal K(ATP) channels. AB - The functional significance of ATP-sensitive K(+) (K(ATP)) channels is controversial. In the present study, transgenic mice expressing a mutant Kir6.2, with reduced ATP sensitivity, were used to examine the role of sarcolemmal K(ATP) in normal cardiac function and after an ischemic or metabolic challenge. We found left ventricular developed pressure (LVDP) was 15-20% higher in hearts from transgenics in the absence of cardiac hypertrophy. beta-Adrenergic stimulation caused a positive inotropic response from nontransgenic hearts that was not observed in transgenic hearts. Decreasing extracellular Ca(2+) decreased LVDP in hearts from nontransgenics but not in those from transgenics. These data suggest an increase in intracellular [Ca(2+)] in transgenic hearts. Additional studies have demonstrated hearts from nontransgenics and transgenics have a similar postischemic LVDP. However, ischemic preconditioning does not improve postischemic recovery in transgenics. Transgenic hearts also demonstrate a poor recovery after metabolic inhibition. These data are consistent with the hypothesis that sarcolemmal K(ATP) channels are required for development of normal myocardial function, and perturbations of K(ATP) channels lead to hearts that respond poorly to ischemic or metabolic challenges. PMID- 12124207 TI - H(2)O(2)-induced Ca(2+) overload in NRVM involves ERK1/2 MAP kinases: role for an NHE-1-dependent pathway. AB - Generation of reactive oxygen species (ROS) and intracellular Ca(2+) overload are key mechanisms involved in ischemia-reperfusion (I/R)-induced myocardial injury. The relationship between I/R injury and Ca(2+) overload has not been fully characterized. The increase in Na(+)/H(+) exchanger (NHE-1) activity observed during I/R injury is an attractive candidate to link increased ROS production with Ca(2+) overload. We have shown that low doses of H(2)O(2) increase NHE-1 activity in an extracellular signal-regulated kinase (ERK)-dependent manner. In this study, we examined the effect of low doses of H(2)O(2) on intracellular Ca(2+) in fura 2-loaded, spontaneously contracting neonatal rat ventricular myocytes. H(2)O(2) induced a time- and concentration-dependent increase in diastolic intracellular Ca(2+) concentration that was blocked by inhibition of ERK1/2 activation with 5 microM U-0126 (88%) or inhibition of NHE-1 with 5 microM HOE-642 (50%). Increased NHE activity was associated with phosphorylation of the NHE-1 carboxyl tail that was blocked by U-0126. These results suggest that H(2)O(2) induced Ca(2+) overload is partially mediated by NHE-1 activation secondary to phosphorylation of NHE-1 by the ERK1/2 MAP kinase pathway. PMID- 12124208 TI - K(Ca) channel blockers reveal hyperpolarization and relaxation to K+ in rat isolated mesenteric artery. AB - Raising extracellular K+ concentration ([K+](o)) around mesenteric resistance arteries reverses depolarization and contraction to phenylephrine. As smooth muscle depolarizes and intracellular Ca(2+) and tension increase, this effect of K+ is suppressed, whereas efflux of cellular K+ through Ca(2+)-activated K+ (K(Ca)) channels is increased. We investigated whether K+ efflux through K(Ca) suppresses the action of exogenous K+ and whether it prestimulates smooth muscle Na(+)-K(+)-ATPase. Under isometric conditions, 10.8 mM [K+](o) had no effect on arteries contracted >10 mN, unless 100 nM iberiotoxin (IbTX), 100 nM charybdotoxin (ChTX), and/or 50 nM apamin were present. Simultaneous measurements of membrane potential and tension showed that phenylephrine depolarized and contracted arteries to -32.2 +/- 2.3 mV and 13.8 +/- 1.6 mN (n = 5) after blockade of K(Ca), but 10.8 mM K+ reversed fully the responses (107.6 +/- 8.6 and 98.8 +/- 0.6%, respectively). Under isobaric conditions and preconstriction with phenylephrine, 10.7 mM [K+](o) reversed contraction at both 50 mmHg (77.0 +/- 8.5%, n = 9) and 80 mmHg (83.7 +/- 5.5%, n = 5). However, in four additional vessels at 80 mmHg, raising K+ failed to reverse contraction unless ChTX was present. Increases in isometric and decreases in isobaric tension with phenylephrine were augmented by either ChTX or ouabain (100 microM), whereas neither inhibitor altered tension under resting conditions. Inhibition of cellular K+ efflux facilitates hyperpolarization and relaxation to exogenous K+, possibly by indirectly reducing the background activation of Na(+)-K(+)-ATPase. PMID- 12124209 TI - Effects of ACh and adenosine mediated by Kir3.1 and Kir3.4 on ferret ventricular cells. AB - The inotropic effects of ACh and adenosine on ferret ventricular cells were investigated with the action potential-clamp technique. Under current clamp, both agonists resulted in action potential shortening and a decrease in contraction. Under action potential clamp, both agonists failed to decrease contraction substantially. In the absence of agonist, application of the short action potential waveform (recorded previously in the presence of agonist) also resulted in a decrease in contraction. Under action potential clamp, application of ACh resulted in a Ba(2+)-sensitive outward current with the characteristics of muscarinic K+ current (I(K,ACh)); the presence of the muscarinic K+ channel was confirmed by PCR and immunocytochemistry. In the absence of agonist, on application of the short ACh action potential waveform, the decrease in contraction was accompanied by loss of the inward Na(+)/Ca(2+) exchange current (I(NaCa)). ACh also inhibited the background inward K+ current (I(K,1)). It is concluded that ACh activates I(K,ACh), inhibits I(K,1), and indirectly inhibits I(NaCa); this results in action potential shortening, decrease in contraction, and, as a result of the inhibition of I(K,1), minimum decrease in excitability. PMID- 12124210 TI - Cardiac dysfunction in terms of left ventricular mechanical work and energetics in hypothyroid rats. AB - We hypothesized that cardiac dysfunction in hypothyroidism is mainly caused by the impairment of Ca(2+) handling in excitation-contraction coupling. To prove this hypothesis, we investigated left ventricular (LV) mechanical work and energetics without interference of preload and afterload in an excised, blood perfused whole heart preparation from hypothyroid rats. We found that LV inotropism and lusitropism were significantly depressed, and these depressions were causally related to decreased myocardial oxygen consumption for Ca(2+) handling and for basal metabolism. The oxygen costs of LV contractility for Ca(2+) and for dobutamine in the hypothyroid rats did not differ from those in age-matched normal rats. The expression of sarco(endo)plasmic reticulum Ca(2+) ATPase (SERCA2) significantly decreased and that of phospholamban significantly increased. The present results revealed that changes in LV energetics associated with decreased mechanical work in hypothyroid rats are mainly caused by the impairment of Ca(2+) uptake via SERCA2. We conclude that the impairment of Ca(2+) uptake plays an important role in the pathogenesis of cardiac dysfunction in hypothyroidism. PMID- 12124211 TI - Ca(2+) activation and tension cost in myofilaments from mouse hearts ectopically expressing enteric gamma-actin. AB - To determine the significance of actin isoforms in chemomechanical coupling, we compared tension and ATPase rate in heart myofilaments from nontransgenic (NTG) and transgenic (TG) mice in which enteric gamma-actin replaced >95% of the cardiac alpha-actin. Enteric gamma-actin was expressed against three backgrounds: mice expressing cardiac alpha-actin, heterozygous null cardiac alpha-actin mice, and homozygous null cardiac alpha-actin mice. There were no differences in maximum Ca(2+) activated tension or maximum rate of tension redevelopment after a quick release and rapid restretch protocol between TG and NTG skinned fiber bundles. However, compared with NTG controls, Ca(2+) sensitivity of tension was significantly decreased and economy of tension development was significantly increased in myofilaments from all TG hearts. Shifts in myosin isoform population could not fully account for this increase in the economy of force production of TG myofilaments. Our results indicate that an exchange of cardiac alpha-actin with an actin isoform differing in only five amino acids has a significant impact on both Ca(2+) regulation of cardiac myofilaments and the cross-bridge cycling rate. PMID- 12124212 TI - Neutralization of IL-18 attenuates lipopolysaccharide-induced myocardial dysfunction. AB - Tumor necrosis factor-alpha (TNF-alpha) and interleukin-1beta (IL-1beta) have been implicated in cardiac dysfunction during endotoxemia. Because IL-18 is a proinflammatory cytokine known to mediate the production of TNF-alpha and IL 1beta and to induce the expression of intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1), we hypothesized that neutralization of IL-18 would attenuate lipopolysaccharide (LPS)-induced cardiac dysfunction. Mice (C57BL/6) were injected with LPS (0.5 mg/kg ip) or vehicle (normal saline), and left ventricular developed pressure (LVDP) was determined by the Langendorff technique. LVDP was depressed by 38% at 6 h after LPS. LPS induced myocardial dysfunction was associated with increased myocardial levels of TNF-alpha and IL-1beta as well as increased expression of ICAM-1/VCAM-1. Pretreatment with neutralizing anti-mouse IL-18 antibody attenuated LPS-induced myocardial dysfunction (by 92%) and was associated with reduced myocardial IL 1beta production (65% reduction) and ICAM-1/VCAM-1 expression (50% and 35% reduction, respectively). However, myocardial TNF-alpha levels were not influenced by neutralization of IL-18. In conclusion, neutralization of IL-18 protects against LPS-induced myocardial dysfunction. IL-18 may mediate endotoxemic myocardial dysfunction through induction of and/or synergy with IL 1beta, ICAM-1, and VCAM-1. PMID- 12124213 TI - Functional NOS 1 in the rat mesenteric arterial bed. AB - Previously we have demonstrated functional nitric oxide synthase (NOS) 1 in large arteries. Because resistance arteries largely determine blood pressure, this study examined whether functional NOS 1 also exists in resistance arteries. Phenylephrine (PE) contraction was measured in the absence and presence of the NOS 1 inhibitor N(5)-(1-imino-3-butenyl)-L-ornithine (VNIO) in isolated mesenteric resistance arteries (endothelium intact and denuded) from Sprague Dawley rats. For NOS 1 activity and expression, the mesenteric arterial bed was separated into cytosolic and particulate fractions. NOS activity was assayed by measuring the conversion of [(3)H]arginine to [(3)H]citrulline inhibited by a nonselective NOS inhibitor or VNIO. VNIO increased PE sensitivity in endothelium intact and -denuded arteries. In cytosolic and particulate fractions of the arterial bed, approximately 40% of NOS activity was inhibited by VNIO. Immunoprecipitation and Western blot analysis revealed two NOS 1 immunoreactive bands. One band corresponded to the rat brain isoform, whereas the second was of a slightly lower molecular mass. The cytosolic fraction contained both isoforms; however, the particulate fraction had only the lower molecular mass form. These studies demonstrate the existence of functional NOS 1 in resistance arteries. PMID- 12124214 TI - Increasing I(Ks) corrects abnormal repolarization in rabbit models of acquired LQT2 and ventricular hypertrophy. AB - Excessive action potential (AP) prolongation and early afterdepolarizations (EAD) are triggers of malignant ventricular arrhythmias. A slowly activating delayed rectifier K+ current (I(Ks)) is important for repolarization of ventricular AP. We examined the effects of I(Ks) activation by a new benzodiazepine (L3) on the AP of control, dofetilide-treated, and hypertrophied rabbit ventricular myocytes. In both control and hypertrophied myocytes, L3 activated I(Ks) via a negative shift in the voltage dependence of activation and a slowing of deactivation. L3 had no effect on L-type Ca(2+) current or other cardiac K+ currents tested. L3 shortened AP of control, dofetilide-treated, and hypertrophied myocytes more at 0.5 than 2 Hz. Selective activation of I(Ks) by L3 attenuates prolonged AP and eliminated EAD induced by rapidly activating delayed rectifier K+ current inhibition in control myocytes at 0.5 Hz and spontaneous EAD in hypertrophied myocytes at 0.2 Hz. Pharmacological activation of I(Ks) is a promising new strategy to suppress arrhythmias resulting from excessive AP prolongation in patients with certain forms of long QT syndrome or cardiac hypertrophy and failure. PMID- 12124215 TI - Increasing plasmalogen levels protects human endothelial cells during hypoxia. AB - Supplementation of cultured human pulmonary arterial endothelial cells (PAEC) with sn-1-O-hexadecylglycerol (HG) resulted in an approximately twofold increase in cellular levels of plasmalogens, a subclass of phospholipids known to have antioxidant properties; this was due, primarily, to a fourfold increase in the choline plasmalogens. Exposure of unsupplemented human PAEC to hypoxia (PO(2) = 20-25 mmHg) caused an increase in cellular reactive oxygen species (ROS) over a period of 5 days with a coincident decrease in viability. In contrast, HG supplemented cells survived for at least 2 wk under these conditions with no evidence of increased ROS. Hypoxia resulted in a selective increase in the turnover of the plasmalogen plasmenylethanolamine. Human PAEC with elevated plasmalogen levels were also more resistant to H(2)O(2), hyperoxia, and the superoxide generator plumbagin. This protection was seemingly specific to cellular stresses in which significant ROS were generated because the sensitivity to lethal heat shock or glucose deprivation was not altered in HG-treated human PAEC. HG, by itself, was not sufficient for protection; HG supplementation of bovine PAEC had no effect upon plasmalogen levels and did not rescue these cells from the cytotoxic effects of hypoxia. This is the initial demonstration that plasmalogen content can be substantially enhanced in a normal cell. These data also demonstrate that HG can protect cells during hypoxia and other ROS-mediated stress, likely due to the resulting increase in these antioxidant phospholipids. PMID- 12124216 TI - Mitochondrial creatine kinase is critically necessary for normal myocardial high energy phosphate metabolism. AB - The individual functional significance of the various creatine kinase (CK) isoenzymes for myocardial energy homeostasis is poorly understood. Whereas transgenic hearts lacking the M subunit of CK (M-CK) show unaltered cardiac energetics and left ventricular (LV) performance, deletion of M-CK in combination with loss of sarcomeric mitochondrial CK (ScCKmit) leads to significant alterations in myocardial high-energy phosphate metabolites. To address the question as to whether this alteration is due to a decrease in total CK activity below a critical threshold or due to the specific loss of ScCKmit, we studied isolated perfused hearts with selective loss of ScCKmit (ScCKmit(-/-), remaining total CK activity approximately 70%) using (31)P NMR spectroscopy at two different workloads. LV performance in ScCKmit(-/-) hearts (n = 11) was similar compared with wild-type hearts (n = 9). Phosphocreatine/ATP, however, was significantly reduced in ScCKmit(-/-) compared with wild-type hearts (1.02 +/- 0.05 vs. 1.54 +/- 0.07, P < 0.05). In parallel, free [ADP] was higher (144 +/- 11 vs. 67 +/- 7 microM, P < 0.01) and free energy release for ATP hydrolysis (DeltaG(ATP)) was lower (-55.8 +/- 0.5 vs. -58.5 +/- 0.5 kJ/mol, P < 0.01) in ScCKmit(-/-) compared with wild-type hearts. These results demonstrate that M- and B-CK containing isoenzymes are unable to fully substitute for the loss of ScCKmit. We conclude that ScCKmit, in contrast to M-CK, is critically necessary to maintain normal high-energy phosphate metabolite levels in the heart. PMID- 12124217 TI - Cardioselective overexpression of HO-1 prevents I/R-induced cardiac dysfunction and apoptosis. AB - Heme oxygenase (HO)-1 converts heme to bilirubin, carbon monoxide, and iron. Our prior work has suggested a cardioprotective role for HO-1 in heart failure. To test whether HO-1 (heat shock protein 32) prevents cardiomyocyte apoptosis and cardiac dysfunction after ischemia-reperfusion (I/R), we generated transgenic mice overexpressing HO-1 in the heart under the control of the alpha-myosin heavy chain promoter. HO-1 transcript and protein increased markedly in the heart only. In an isolated heart preparation, we observed an enhanced functional recovery during reperfusion after ischemia in the transgenic hearts compared with nontransgenic controls. I/R injury was also performed in intact animals by coronary ligation and reperfusion to assess the protective role of HO-1 overexpression on heart apoptosis. HO-1 overexpression reduced cardiac apoptosis, as evidenced by fewer terminal deoxynucleodidyl transferase-mediated dUTP nick end labeling-positive or in situ oligo ligation-positive myocytes, compared with nontransgenic mice. Our results indicate that cardioselective overexpression of HO-1 exerts a cardioprotective effect after myocardial I/R in mice, and this effect is probably mediated via an antiapoptotic action of HO-1. PMID- 12124218 TI - PYK2 expression and phosphorylation increases in pressure overload-induced left ventricular hypertrophy. AB - Proline-rich tyrosine kinase 2 (PYK2) is a member of the focal adhesion kinase (FAK) family of nonreceptor protein tyrosine kinases. PYK2 has been implicated in linking G protein-coupled receptors to activation of mitogen-activated protein kinase cascades and cellular growth in a variety of cell types. To determine whether PYK2 expression and phosphorylation is altered in left ventricular (LV) myocardium undergoing LV hypertrophy (LVH) and heart failure in vivo, suprarenal abdominal aortic coarctation was performed in 160-g male Sprague-Dawley rats. Immunohistochemistry and Western blotting were performed on LV tissue 1, 8, and 24 wk after aortic banding. Aortic banding produced sustained hypertension and gradually developing LVH. PYK2 levels were increased 1.8 +/- 0.2-, 2.7 +/- 0.6-, and 2.0 +/- 0.2-fold in 1-, 8-, and 24-wk banded animals compared with their respective sham-operated controls. The increase in PYK2 expression was paralleled by an increase in PYK2 phosphorylation, both of which preceded the development of LVH. Immunohistochemistry revealed that enhanced PYK2 expression occurred predominantly in the cardiomyocyte population. Furthermore, there was a high degree of correlation (R = 0.75; P < 0.001) between the level of PYK2 and the degree of LVH in 24-wk sham and banded animals. In contrast, FAK levels and FAK phosphorylation were not increased before the development of LVH. However, there was a high degree of correlation (R = 0.68; P < 0.001) between the level of FAK and the degree of LVH in 24-wk sham and banded rats. There was also a significant increase in the ratio of phosphospecific anti-FAK to FAK at this time point. These data are consistent with a role for PYK2 in the induction of pressure overload-induced cardiomyocyte hypertrophy, and suggest that PYK2 and FAK have distinctly different roles in LVH progression. PMID- 12124220 TI - Electrophysiological response of rat atrial myocytes to acidosis. AB - The effect of acidosis on the electrical activity of isolated rat atrial myocytes was investigated using the patch-clamp technique. Reducing the pH of the bathing solution from 7.4 to 6.5 shortened the action potential. Acidosis had no significant effect on transient outward or inward rectifier currents but increased steady-state outward current. This increase was still present, although reduced, when intracellular Ca(2+) was buffered by 1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid (BAPTA); BAPTA also inhibited acidosis-induced shortening of the action potential. Ni(2+) (5 mM) had no significant effect on the acidosis-induced shortening of the action potential. Acidosis also increased inward current at -80 mV and depolarized the resting membrane potential. Acidosis activated an inwardly rectifying Cl(-) current that was blocked by 4,4' diisothiocyanostilbene-2,2'-disulfonic acid (DIDS), which also inhibited the acidosis-induced depolarization of the resting membrane potential. It is concluded that an acidosis-induced increase in steady-state outward K(+) current underlies the shortening of the action potential and that an acidosis-induced increase in inwardly rectifying Cl(-) current underlies the depolarization of the resting membrane potential during acidosis. PMID- 12124219 TI - Ventricular expression of natriuretic peptides in Npr1(-/-) mice with cardiac hypertrophy and fibrosis. AB - Atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) are cardiac hormones that regulate blood pressure and volume, and exert their biological actions via the natriuretic peptide receptor-A gene (Npr1). Mice lacking Npr1 (Npr(-/-)) have marked cardiac hypertrophy and fibrosis disproportionate to their increased blood pressure. This study examined the relationships between ANP and BNP gene expression, immunoreactivity and fibrosis in cardiac tissue, circulating ANP levels, and ANP and BNP mRNA during embryogenesis in Npr1(-/-) mice. Disruption of the Npr1 signaling pathway resulted in augmented ANP and BNP gene and ANP protein expression in the cardiac ventricles, most pronounced for ANP mRNA in females [414 +/- 57 in Npr1(-/-) ng/mg and 124 +/- 25 ng/mg in wild-type (WT) by Taqman assay, P < 0.001]. This increased expression was highly correlated to the degree of cardiac hypertrophy and was localized to the left ventricle (LV) inner free wall and to areas of ventricular fibrosis. In contrast, plasma ANP was significantly greater than WT in male but not female Npr1(-/-) mice. Increased ANP and BNP gene expression was observed in Npr1(-/-) embryos from 16 days of gestation. Our study suggests that cardiac ventricular expression of ANP and BNP is more closely associated with local hypertrophy and fibrosis than either systemic blood pressure or circulating ANP levels. PMID- 12124221 TI - Differential PDGF secretion by graft and aortic SMC in response to oxidized LDL. AB - Smooth muscle cells (SMC) from prosthetic vascular grafts constitutively secrete higher levels of platelet-derived growth factor-AA (PDGF-AA) than aortic SMC. Lipid oxidation products accumulate in grafts and may stimulate PDGF production. The effect of oxidized low-density lipoprotein (oxLDL) on PDGF-AA secretion by aortic and graft SMC was compared. SMC isolated from canine thoracic aorta or Dacron thoracoabdominal grafts (n = 10) were incubated with native LDL or oxLDL (0-400 microg/ml) for 72 h. PDGF-AA in the conditioned medium was measured with enzyme-linked immunosorbent assay. OxLDL increased PDGF-AA production by graft SMC from 78 +/- 2 to 256 +/- 16 pg PDGF/microg DNA and aortic SMC from 21 +/- 1 to 40 +/- 2 pg PDGF/microg DNA. Native LDL had no effect. N-acetylcysteine inhibited oxLDL-induced PDGF increase. Both superoxide and H(2)O(2) stimulated PDGF secretion by graft SMC had little effect on aortic SMC. Our results suggest that PDGF production by graft (synthetic) SMC is more sensitive to stimulation by oxidative stress than aortic (contractile) SMC. Lipid oxidation products that accumulate in prosthetic vascular grafts can cause an oxidative stress, which stimulates PDGF production by graft SMC. PDGF can induce migration of aortic SMC onto the graft, contributing to the development of intimal hyperplasia. PMID- 12124222 TI - Cardiac modulations of ANG II receptor expression in rats with hypoxic pulmonary hypertension. AB - Right ventricular myocardial hypertrophy during hypoxic pulmonary hypertension is associated with local renin-angiotensin system activation. The expression of angiotensin II type 1 (AT(1)) and type 2 (AT(2)) receptors in this setting has never been investigated. We have therefore examined the chronic hypoxia pattern of AT(1) and AT(2) expression in the right and left cardiac ventricles, using in situ binding and RT-PCR assays. Hypoxia produced right, but not left, ventricular hypertrophy after 7, 14, and 21 days, respectively. Hypoxia for 2 days was associated in each ventricle with a simultaneous and transient increase (P < 0.05) in AT(1) binding and AT(1) mRNA levels in the absence of any significant change in AT(2) expression level. Only after 14 days of hypoxia, AT(2) binding increased (P < 0.05) in the two ventricles, concomitantly with a right ventricular decrease (P < 0.05) in AT(2) mRNA. Along these data, AT(1) and AT(2) binding remained unchanged in both the left and hypertrophied right ventricles from rats treated with monocrotaline for 30 days. These results indicate that chronic hypoxia induces modulations of AT(1) and AT(2) receptors in both cardiac ventricles probably through direct and indirect mechanisms, respectively, which modulations may participate in myogenic (at the level of smooth or striated myocytes) rather than in the growth response of the heart to hypoxia. PMID- 12124223 TI - Initiation and propagation of ectopic waves: insights from an in vitro model of ischemia-reperfusion injury. AB - The objective of the present study was to directly visualize ectopic activity associated with ischemia-reperfusion and its progression to arrhythmia. To accomplish this goal, we employed a two-dimensional network of neonatal rat cardiomyocytes and a recently developed model of localized ischemia-reperfusion. Washout of the ischemia-like solution resulted in tachyarrhythmic episodes lasting 15-200 s. These episodes were preceded by the appearance of multiple ectopic sources and propagation of ectopic activity along the border of the former ischemic zone. The ectopic sources exhibited a slow rise in diastolic calcium, which disappeared upon return to the original pacing pattern. Border zone propagation of ectopic activity was followed by its escape into the surrounding control network, generating arrhythmias. Together, these observations suggest that upon reperfusion, a distinct layer, which consists of ectopically active, poorly coupled cells, is formed transiently over an injured area. Despite being neighbored by a conductive and excitable tissue, this transient functional layer is capable of sustaining autonomous waves and serving as a special conductive medium through which ectopic activity can propagate before spreading into the surrounding healthy tissue. PMID- 12124224 TI - Native LDL and minimally oxidized LDL differentially regulate superoxide anion in vascular endothelium in situ. AB - Low-density lipoprotein (LDL) and its oxidized derivatives are hypothesized to impair vascular function by increasing superoxide anion (O.). To investigate mechanisms in situ, isolated carotid arteries were incubated with native LDL (nLDL) or minimally oxidized LDL (mmLDL). With the use of en face fluorescent confocal microscopy and hydroethidine, an oxidant-sensitive fluorescent probe, we found that nLDL increased O. in vascular endothelium greater than fourfold by an N(omega)-nitro-L-arginine methyl ester (L-NAME)-inhibitable mechanism. In contrast, mmLDL increased O. in vascular endothelium greater than eightfold by mechanisms that were partially inhibited by L-NAME and allopurinol and essentially ablated by diphenyleneiodium. These data indicate that both nLDL and mmLDL uncouple endothelial nitric oxide synthase (eNOS) activity and that mmLDL also activates xanthine oxidase and NADPH oxidoreductase to induce greater increases in O. generation than nLDL. Western analysis revealed that both lipoproteins inhibited A-23187-stimulated association of heat shock protein 90 (HSP90) with eNOS without inhibiting phosphorylation of eNOS at serine-1179 (phospho-eNOS), an immunological index of electron flow through the enzyme. As HSP90 mediates the balance of.NO and O. generation by eNOS, these data provide new insight into the mechanisms by which oxidative stress, induced by nLDL and mmLDL, uncouple eNOS activity to increase endothelial O. generation. PMID- 12124225 TI - Rat carotid artery dilation by PTCA balloon catheter induces neointima formation in presence of IEL rupture. AB - The best animal angioplasty model is the porcine model, which is expensive and not available in all laboratories. The aim of this study was to describe a new rat model of angioplasty. An injury was induced with the use of a standard percutaneous transluminal coronary angioplasty (PTCA) 1.5-mm balloon catheter. The neointimal tissue, arterial dimensions, and the injury index were assessed following angioplasty. Ki-67 expression was detected to evaluate cell turnover after balloon angioplasty. In contrast with the standard Clowes model, a significant neointimal formation was detected only in the presence of ruptured internal elastic lamina (IEL). A positive correlation between the percentage of ruptured IEL and the amount of neointimal tissue was also demonstrated. The percentage of IEL fracture correlates with the proliferation index by anti-Ki-67 immunolabeling 7 and 14 days after the angioplasty. Significant arterial negative remodeling was observed following PTCA balloon dilation. In conclusion, our inexpensive animal model of restenosis after angioplasty may have great relevance toward a better understanding of the mechanisms and toward assessment of new therapeutical strategies for this phenomenon. PMID- 12124226 TI - Simvastatin restores endothelial NO-mediated vasorelaxation in large arteries after myocardial infarction. AB - Congestive heart failure (CHF) after myocardial infarction is associated with diminished endothelial nitric oxide (NO)-mediated vasorelaxation. The 3-hydroxy-3 methylglutaryl-CoA reductase inhibitors have been shown to modulate vascular tone independent of the effects on lipid lowering. We hypothesized that simvastatin restores NO-dependent vasorelaxation with CHF. We found that incubation of the normal rat aorta with 0.1 mM simvastatin for 24 h enhanced ACh-mediated vasorelaxation (P < 0.05). Moreover, simvastatin increased (P < 0.05) endothelial NO synthase (eNOS) protein content by >200% (82.0 +/- 14.0 vs. 21.6 +/- 7.9% II/microg). In cultured endothelial cells, simvastatin (10 and 20 microM) increased eNOS levels by 114.7 +/- 39.9 and 212.0 +/- 75.0% II/microg protein, respectively (both P < 0.05; n = 8). In the rat coronary artery ligation model, oral gavage with 20 mg. kg(-1). day(-1) simvastatin for 3 wk decreased (P < 0.05) mean arterial pressure (121 +/- 20 vs. 96.5 +/- 10.8 mmHg) and left ventricular change in pressure with time (4,500 +/- 700 vs. 4,091 +/- 1,064 mmHg/s, n = 6). Simvastatin reduced (P < 0.05) basal vasoconstriction and improved ACh-mediated vasorelaxation in CHF arterial rings. Inhibition of NO generation by N(G)-nitro-L arginine methyl ester (100 microM) abolished the ACh-induced vasorelaxation in all rats. In conclusion, chronic treatment of CHF with simvastatin restores endothelial NO-dependent dysfunction and upregulates eNOS protein content in arterial tissue. PMID- 12124227 TI - Adenylate kinase AK1 knockout heart: energetics and functional performance under ischemia-reperfusion. AB - Deletion of the major adenylate kinase AK1 isoform, which catalyzes adenine nucleotide exchange, disrupts cellular energetic economy and compromises metabolic signal transduction. However, the consequences of deleting the AK1 gene on cardiac energetic dynamics and performance in the setting of ischemia reperfusion have not been determined. Here, at the onset of ischemia, AK1 knockout mice hearts displayed accelerated loss of contractile force compared with wild-type controls, indicating reduced tolerance to ischemic stress. On reperfusion, AK1 knockout hearts demonstrated reduced nucleotide salvage, resulting in lower ATP, GTP, ADP, and GDP levels and an altered metabolic steady state associated with diminished ATP-to-P(i) and creatine phosphate-to-P(i) ratios. Postischemic AK1 knockout hearts maintained approximately 40% of beta phosphoryl turnover, suggesting increased phosphotransfer flux through remaining adenylate kinase isoforms. This was associated with sustained creatine kinase flux and elevated cellular glucose-6-phosphate levels as the cellular energetic system adapted to deletion of AK1. Such metabolic rearrangements, along with sustained ATP-to-ADP ratio and total ATP turnover rate, maintained postischemic contractile recovery of AK1 knockout hearts at wild-type levels. Thus deletion of the AK1 gene reveals that adenylate kinase phosphotransfer supports myocardial function on initiation of ischemic stress and safeguards intracellular nucleotide pools in postischemic recovery. PMID- 12124228 TI - A highly potent peptide analgesic that protects against ischemia-reperfusion induced myocardial stunning. AB - We recently discovered an opioid peptide analgesic, 2',6'-dimethyltyrosine (Dmt) D-Arg-Phe-Lys-NH(2) ([Dmt(1)]DALDA), that can protect against ischemia-induced myocardial stunning. In buffer-perfused hearts, 30-min global ischemia followed by reperfusion resulted in a significant increase in norepinephrine (NE) overflow immediately upon reperfusion and significant decline in contractile force (45%). Pretreatment with [Dmt(1)]DALDA before ischemia completely abolished myocardial stunning and significantly reduced NE overflow (68%). In contrast, pretreatment with morphine before ischemia only provided brief protection against myocardial stunning and no reduction in NE overflow. [Dmt(1)]DALDA inhibited [(3)H]NE uptake into cardiac synaptosomes in vitro (IC(50) = 3.9 microM), whereas morphine had no effect. Surprisingly, protection against myocardial stunning was apparent even when hearts were perfused with [Dmt(1)]DALDA only upon reperfusion, whereas reperfusion with morphine had no effect. Binding studies with [(3)H][Dmt(1)]DALDA revealed no high-affinity specific binding in cardiac membranes, suggesting that the cardioprotective actions of [Dmt(1)]DALDA are not mediated via opioid receptors. These findings suggest that [Dmt(1)]DALDA is a potent analgesic that may be useful for myocardial stunning resulting from cardiac interventions or myocardial ischemia. PMID- 12124229 TI - Myocardial function defined by strain rate and strain during alterations in inotropic states and heart rate. AB - For porcine myocardium, ultrasonic regional deformation parameters, systolic strain (epsilon(sys)) and peak systolic strain rate (SR(sys)), were compared with stroke volume (SV) and contractility [contractility index (CI)] measured as the ratio of end-systolic strain to end-systolic wall stress. Heart rate (HR) and contractility were varied by atrial pacing (AP = 120-180 beats/min, n = 7), incremental dobutamine infusion (DI = 2.5-20 microg. kg(-1). min(-1), n = 7), or continuous esmolol infusion (0.5 mg. kg(-1). min(-1)) + subsequent pacing (120 180 beats/min) (EI group, n = 6). Baseline SR(sys) and epsilon(sys) averaged 5.0 +/- 0.4 s(-1) and 60 +/- 4%. SR(sys) and CI increased linearly with DI (20 microg. kg(-1). min(-1); SR(sys) = 9.9 +/- 0.7 s(-1), P < 0.0001) and decreased with EI (SR(sys) = 3.4 +/- 0.1 s(-1), P < 0.01). During pacing, SR(sys) and CI remained unchanged in the AP and EI groups. During DI, epsilon(sys) and SV initially increased (5 microg. kg(-1). min(-1); epsilon(sys) = 77 +/- 6%, P < 0.01) and then progressively returned to baseline. During EI, SV and epsilon(sys) decreased (epsilon(sys) = 38 +/- 2%, P < 0.001). Pacing also decreased SV and epsilon(sys) in the AP (180 beats/min; epsilon(sys) = 36 +/- 2%, P < 0.001) and EI groups (180 beats/min; epsilon(sys) = 25 +/- 3%, P < 0.001). Thus, for normal myocardium, SR(sys) reflects regional contractile function (being relatively independent of HR), whereas epsilon(sys) reflects changes in SV. PMID- 12124230 TI - Changes in excitation-contraction coupling in an isolated ventricular myocyte model of cardiac stunning. AB - To investigate cardiac stunning, we recorded intracellular [Ca(2+)], contractions, and electrical activity in isolated guinea pig ventricular myocytes exposed to simulated ischemia and reperfusion. After equilibration, ischemia was simulated by exposing myocytes to hypoxia, acidosis, hyperkalemia, hypercapnia, lactate accumulation, and substrate deprivation for 30 min at 37 degrees C. Reperfusion was simulated by exposure to Tyrode solution. Field-stimulated myocytes exhibited stunning upon reperfusion. By 10 min of reperfusion, contraction amplitude decreased to 43.0 +/- 5.5% of preischemic values (n = 15, P < 0.05), although action potential configuration and sarcoplasmic reticulum Ca(2+) stores, assessed with caffeine, were normal. Diastolic [Ca(2+)] and Ca(2+) transients (fura 2) were also normal in stunned myocytes. In voltage-clamped cells, peak L-type Ca(2+) current was reduced to 47.4 +/- 4.5% of preischemic values at 10 min of reperfusion (n = 21, P < 0.05). Contractions elicited by Ca(2+)-induced Ca(2+) release and the voltage-sensitive release mechanism were both depressed in reperfusion. Our observations suggest that stunning is associated with reduced L-type Ca(2+) current but that alterations in Ca(2+) homeostasis and release are not directly responsible for stunning. PMID- 12124231 TI - Omega-3 fatty acids suppress monocyte adhesion to human endothelial cells: role of endothelial PAF generation. AB - Monocyte-endothelium interaction is a fundamental process in many acute and chronic inflammatory diseases. Eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) are fish oil-derived alternative (omega-3) precursor fatty acids implicated in the suppression of inflammatory events. We investigated their influence on rolling and adhesion of monocytes to human umbilical vein endothelial cells (HUVEC) under laminar flow conditions in vitro. Exposure of HUVEC to tumor necrosis factor (TNF-alpha) strongly increased 1) surface expression of intercellular adhesion molecule (ICAM-1), vascular cell adhesion molecule (VCAM-1), and E-selectin, 2) platelet-activating factor (PAF) synthesis as assessed by thrombin challenge, and 3) rate of rolling and adhesion of monocytes. Preincubation of HUVEC with EPA or DHA markedly suppressed PAF synthesis, monocyte rolling, and adherence, whereas expression of endothelial adhesion molecules was unchanged. Also, PAF receptor antagonists markedly suppressed the adhesion rate of monocytes, and EPA or DHA revealed no additional inhibitory capacity. In contrast, arachidonic acid partially reversed the effect of the antagonist. We conclude that omega-3 fatty acids suppress rolling and adherence of monocytes on activated endothelial cells in vitro by affecting endothelial PAF generation. PMID- 12124232 TI - Role of receptor kinase in long-term desensitization of the cardiac muscarinic receptor-K+ channel system. AB - Desensitization of the cardiac muscarinic K+ channel was studied in cultured neonatal rat atrial cells and in Chinese hamster ovary (CHO) cells transfected with muscarinic receptor (HM(2)), G protein-coupled inward rectifying K+ channels 1 and 4, and G protein-coupled receptor kinase 2. In atrial cells incubated in 10 microM carbachol for 24 h, channel activity in cell-attached patches was substantially reduced as a result of long-term desensitization. The long-term desensitization was also observed in CHO cells transfected with the wild-type receptor and receptor kinase (as well as the channel). However, long-term desensitization was greatly reduced or abolished if the cells were 1) not transfected with the receptor kinase, 2) transfected with a mutant receptor lacking phosphorylation sites (rather than the wild-type receptor), or 3) transfected with a mutant receptor kinase lacking kinase activity (rather than the wild-type receptor kinase). We suggest that long-term desensitization of the cardiac muscarinic receptor-K+ channel system to muscarinic agonist may involve phosphorylation of the receptor by receptor kinase. PMID- 12124233 TI - IRAK contributes to burn-triggered myocardial contractile dysfunction. AB - Major burn injury causes myocardial contractile dysfunction, but the molecular basis of this physiological response is incompletely understood. Previous studies demonstrated a role for the interleukin-1 receptor-associated kinase (IRAK) in the cardiac response to acute lipopolysaccharide administration as well as congestive heart failure. In this study, we examined the contribution of IRAK to burn-mediated cardiac responses. After burn injury, hearts from wild-type and IRAK-deficient mice were compared for intracellular signaling pathway activation and contractile function. IRAK-deficient hearts showed impaired activation of kinases that function downstream of IRAK and were partially protected against burn-induced contractile dysfunction. The findings demonstrate that IRAK and the Toll/interleukin-1 pathways participate in the response to large body surface area burns that leads to impaired cardiac contractility. PMID- 12124234 TI - Involvement of Na(+)/Ca(2+) exchanger in endothelial NO production and endothelium-dependent relaxation. AB - Endothelial nitric oxide (NO) synthase (eNOS) is controlled by Ca(2+)/calmodulin and caveolin-1 in caveolae. It has been recently suggested that Na(+)/Ca(2+) exchanger (NCX), also expressed in endothelial caveolae, is involved in eNOS activation. To investigate the role played by NCX in NO synthesis, we assessed the effects of Na(+) loading (induced by monensin) on rat aortic rings and cultured porcine aortic endothelial cells. Effect of monensin was evaluated by endothelium-dependent relaxation of rat aortic rings in response to acetylcholine and by real-time measurement of NO release from cultured endothelial cells stimulated by A-23187 and bradykinin. Na(+) loading shifted the acetylcholine concentration-response curve to the left. These effects were prevented by pretreatment with the NCX inhibitors benzamil and KB-R7943. Monensin potentiated Ca(2+)-dependent NO release in cultured cells, whereas benzamil and KB-R7943 totally blocked Na(+) loading-induced NO release. These findings confirm the key role of NCX in reverse mode on Ca(2+)-dependent NO production and endothelium dependent relaxation. PMID- 12124235 TI - CT imaging of intrabiliary growth of colorectal liver metastases: a comparison of pathological findings of resected specimens. AB - The objective of this study was to assess the usefulness of CT in the pre operative evaluation of macroscopic intrabiliary tumour growth of colorectal liver metastases. 25 metastatic nodules of 18 patients who underwent an initial hepatectomy for colorectal liver metastasis were retrospectively evaluated. The CT appearance and pathological findings of the resected specimens were correlated. A number of unusual peritumoral features associated with intrabiliary tumour growth were detected by pre-operative CT. These were classified into three patterns: (1) thickened portal tract; (2) intrahepatic bile duct dilatation; and (3) a wedge-shaped area with enhancement. In 8 (32%) of the 25 nodules the portal tract was depicted as thicker than usual and these features were found proximal to the tumour in three instances, distal to the tumour in four instances, and both proximal and distal in one instance. All of the three intrabiliary tumours larger than 30 mm resulted in thickening of the portal tract. Intrahepatic bile duct dilatation was detected in association with 10 (40%) of 25 nodules. Bile duct dilatation was observed in more than one segment when intrabiliary tumour reached the hepatic hilus from the tumour. The presence of bile duct dilatation was not related to either the size of the tumour or the extent of intrabiliary tumour growth. An abnormally high density wedge-shaped area on contrast enhanced CT was another feature indicating intrabiliary tumour growth and was seen in association with four nodules. Such areas were seen in the liver parenchyma distal to the tumour on three occasions, or encompassing the tumour on one accasion. This wedge-shaped area appeared as a well demarcated dark red-brown region in the cut surface of the resected specimen. CT was useful for detecting the presence of intrabiliary tumour growth with these three patterns of radiological findings in patients with liver metastases from colorectal cancer PMID- 12124236 TI - MRI of cochlear otosclerosis. AB - Cochlear otosclerosis is an uncommon cause of mixed and sensorineural hearing loss. This has a characteristic appearance on CT, producing a distinctive pericochlear hypodense double ring. However, its appearance on MRI is not as readily appreciated, producing a ring of intermediate signal in the pericochlear and perilabyrinthine regions on T(1) weighted images, demonstrating mild to moderate enhancement after gadolinium administration. Increased signal on T(2) weighted images may also be seen. Recognition of these MRI features is important as MRI may be the first modality of investigation, especially when patients present with symptoms indicative of sensorineural hearing loss. We review four patients who presented with sensoineural hearing loss, and who were imaged with MRI as the first line of investigation. PMID- 12124237 TI - Serial brain MRI at 3-6 month intervals as a surrogate marker for Alzheimer's disease. AB - A surrogate marker is needed for Alzheimer's disease (AD) both to aid diagnosis and to assess interventions. Despite widespread use, brain imaging markers have largely been confounded by overlap with "normal" ageing. 39 elderly subjects completed up to four serial volumetric brain MRI scans with intervals from 2.5 months to 7 months. By National Institute of Neurological and Communicative Disorders and Stroke (NINCDS) criteria, five subjects had probable AD, two possible AD and 32 were negative for AD, although this group included memory impaired subjects. Total brain and ventricular volumes were measured for each scan, and rates of change for each interval calculated. The rate of change in ventricle-to-brain ratio was 15.6% per year (standard deviation (SD) 2.8%) for probable AD compared with 4.3% per year (SD 1.1%) for negative AD (p<0.001). There was no significant difference between these groups' mean ventricle-to-brain ratios measured at a single time point (p=0.25). Rates of change in brain or ventricular volume over time also differed between the two groups (p<0.001). Power calculations reveal that to detect a 20% reduction in the excess rate of atrophy of a probable AD cohort in just 6 months, with 90% power, 135 subjects would be required in each arm of a randomized placebo controlled trial. For a 30% reduction in the excess rate of atrophy, 61 subjects would be required. Rate of change analysis makes serial brain MRI a valuable surrogate marker for Alzheimer's disease. Since only 6 months or less is required between scans, this procedure has both clinical relevance and potential for monitoring interventions. PMID- 12124238 TI - Can contrast enhanced MRI predict the response of Graves' ophthalmopathy to orbital radiotherapy? AB - The purpose of this study was to try to determine by means of contrast-enhanced MRI, a subset of patients with Graves' ophthalmopathy who will not respond to orbital radiotherapy. 54 patients with Graves' ophthalmopathy were treated with orbital radiotherapy (10 x 2 Gy) and symptom relief was recorded. MRI examinations prior to radiotherapy were retrospectively evaluated for enlargement, contrast enhancement and fibrotic changes in extraocular muscles and surrounding soft tissue. Imaging data were correlated with clinical features and response. Symptom relief was observed in 61% of patients but this could not be predicted by any of the MRI signs investigated. However, there is a trend for a better treatment reponse in patients who show contrast enhancement of extraocular muscles prior to orbital radiotherapy (p=0.08). MRI could not adequately predict the efficacy of orbital radiotherapy in this group of patients. Clinical assessment of disease activity is still the most reliable method. PMID- 12124239 TI - Primary radiation outside the imaged volume of a multislice helical CT scan. AB - Multislice helical CT scanning has advantages of speed and X-ray tube loading, making it possible to image larger volumes in a single exposure. Our aim is to investigate dose implications for short scans from the additional X-ray tube rotations required to reconstruct a given volume in helical scanning. To this end a multislice scanner was compared with a single slice scanner. Two independent methods were used. The first was based on optical density measurements taken from a film exposed free-in-air as it moves with the CT bed along the scan axis. The second used measurements from a pencil ionization chamber supported free-in-air at the centre of the CT aperture for the duration of both a long scan and a short scan. This method assumes the same excess primary radiation at the extremes of both scans and the measurements are incorporated into two simultaneous equations. The dose-length product outside the imaged volume has been compared with the dose length product inside the imaged volume using both methods. For 4 x 5 mm multislice collimation with a 360 degrees interpolation and a pitch of 0.875, the film and simultaneous equations methods show an excess dose-length product at the extremes of the scan volume equivalent to 3.3 cm and 3.5 cm extra scan length, respectively. This represents a large percentage of a short scan and is substantially greater than for a helical scan using the single slice scanner with 5 mm collimation, a 360 degrees interpolation and a pitch of 1. The latter showed an excess dose-length product at the extremes which was equivalent to 0.35 cm scan length by the film method and 0.25 cm using simultaneous equations. Taking the abdominal protocols recommended by the respective manufacturers, however, the multislice scanner could cover a 45 cm scan length in a single exposure, while the single slice scanner needed six exposures to image the same volume. With the multislice scanner set at 4 x 2.5 mm collimation, 360 degrees interpolation and a pitch of 0.875, the dose-length product outside the volume of interest was equivalent to 1.9 cm scan length by the first method and 1.8 cm using the second method. With 4 x 1 mm collimation it was equivalent to 1.0 cm using both methods. Changing the interpolation algorithm from 360 degrees to 180 degrees had no effect on the additional equivalent scan length while doubling the pitch resulted in a 25% increase. We conclude from this study that with the multislice scanner, the axial mode is to be preferred for short CT scans such as those used in patient biopsies and drainage. For paediatric helical scans shorter than 13 cm, dose length product is reduced by using 4 x 2.5 mm rather than 4 x 5 mm collimation. For longer scans, however, the increased CT dose index associated with narrower collimation in the multislice mode offsets the dose reduction at the extremes. PMID- 12124240 TI - Effectiveness and relevance of MR acceptance testing: results of an 8 year audit. AB - The effectiveness and relevance of independent acceptance testing was assessed by means of an audit of acceptance procedures for 17 MRI systems, with field strengths in the range 0.5-1.5 T, acquired over 8 years. Signal-to-noise ratio and geometric linearity were found to be the image quality parameters most likely to fall below acceptable or expected standards. These received confirmed successful corrective action in 69% of instances. Non-uniformity, ghosting and poor fat suppression were the next most common non-compliant parameters, but yielded less satisfactory outcomes. Spatial resolution was not found to be a sensitive parameter in determining acceptability. 49% of all non-compliant parameters received verifiable corrective attention. A schedule of actual acceptance criteria is presented and shown to be reasonable. Parameter failure rates were shown not to have improved with time. A safety audit of 11 of the installations revealed the most common failings to be inadequate suite layout and poor use of signs. The mean number of safety issues per installation identified as requiring attention was 5, from a questionnaire of 100 points. A number of anecdotal errors and omissions are reported. The data support the importance of an appropriate acceptance procedure for new clinical MRI equipment and for the involvement of a suitably qualified safety adviser on the project team from the outset. PMID- 12124241 TI - Idiopathic dilatation of the pulmonary artery. AB - Idiopathic dilatation of the pulmonary artery is an uncommon cause of a large main pulmonary artery whose diagnosis is dependent on the exclusion of other causes of central pulmonary artery dilatation. We present four new cases who have been known to have large central pulmonary arteries for many years, and suggest that a long period of observation should be considered to be a further criterion for diagnosis as, in some patients who appear to have this condition, an underlying pathology will become apparent. PMID- 12124242 TI - Supportive cushions produce no practical reduction in lumbar lordosis. AB - It is common practice to place a pillow or bolster under the knees of patients undergoing lumbar spine CT or MRI. The use of such supportive cushions leads to gentle hip flexion that is thought to ease pain and "reduce lumbar lordosis". It is also thought to facilitate axial imaging through the discs. Observations in seven subjects who underwent lumbar spine MRI with and without such hip flexion showed no appreciable change in the degree of lordosis. As the use of such devices does not produce a practical reduction of lumbar lordosis, the decision to employ them should be made entirely with respect to patient comfort. PMID- 12124243 TI - Malignant haemangiopericytoma of the knee joint: MR findings. AB - The imaging findings of intra-articular haemangiopericytomas have not been described previously. We report on a 35-year-old man with a primary malignant haemangiopericytoma of the left knee joint. MRI demonstrated joint effusion and a heterogeneous mass with haemorrhagic components and multiple peripheral serpentine signal voids, suggesting the presence of vascular channels. Multiple pulmonary metastases were seen on chest CT. These imaging findings, although not pathognomonic, might be useful in suggesting the diagnosis of intra-articular haemangiopericytoma. PMID- 12124244 TI - Peripheral primitive neuroectodermal tumour during pregnancy. AB - The case of a 25-year-old primipara in the second trimester of pregnancy, suffering from a peripheral primitive neuroectodermal tumour (pPNET) diagnosed by bone biopsy, is described. External irradiation was initially performed because of Jacksonian seizures due to a lesion in the right cerebral hemisphere. Appropriate shielding was used to reduce fetal exposure during brain radiotherapy. Caesarian delivery at the 27th week of gestation was performed because of tumour progression. The neonate had no evidence of disease and survived for 1 month. However, the placenta and ovaries showed metastases from the maternal pPNET. The patient died 14 months after initial diagnosis owing to the aggressiveness of the tumour, the rapid and extensive semination (bone marrow, lung, liver, craniospinal axis involvement) and the inability to adequately treat the patient with appropriate doses of chemotherapy. PMID- 12124245 TI - False positive uptake of metaiodobenzylguanidine in hepatocellular carcinoma. AB - We present the case of a patient who showed increased accumulation of (123)I metaiodobenzylguanidine (MIBG) in hepatocellular carcinoma, leading to a false presumptive diagnosis of intrahepatic neuroendocrine tumour. The increase in uptake was not seen on images obtained 4 h after tracer injection but was evident on those taken after 24 h, suggesting slower washout from the liver tumour than from non-tumoural liver parenchyma. Our observations indicate that a non neuroendocrine malignant tumour may exhibit high accumulation of MIBG associated with prolonged retention. PMID- 12124246 TI - MRI of the teeth. AB - The teeth and periapical structures were demonstrated on MRI using an open MRI system. There was good visualization of normal structures including crowns of teeth, pulp chambers and the neurovascular bundle of the inferior dental nerve. Dental and periapical pathology was shown. PMID- 12124247 TI - Intracranial lithography? PMID- 12124248 TI - Do the reports address the questions? PMID- 12124249 TI - Guidelines not directives. PMID- 12124250 TI - Simultaneous measurements of mitochondrial NADH and Ca(2+) during increased work in intact rat heart trabeculae. AB - The main goal of this study is to investigate the role of mitochondrial [Ca(2+)], [Ca(2+)](m), in the possible up-regulation of the NADH production rate during increased workload. Such up-regulation is necessary to support increased flux through the electron transport chain and increased ATP synthesis rates. Intact cardiac trabeculae were loaded with Rhod-2(AM), and [Ca(2+)](m) and mitochondrial [NADH] ([NADH](m)) were simultaneously measured during increased pacing frequency. It was found that 53% of Rhod-2 was localized in mitochondria. Increased pacing frequency caused a fast, followed by a slow rise of the Rhod-2 signal, which could be attributed to an abrupt increase in resting cytosolic [Ca(2+)], and a more gradual rise of [Ca(2+)](m), respectively. When the pacing frequency was increased from 0.25 to 2 Hz, the slow Rhod-2 component and the NADH signal increased by 18 and 11%, respectively. Based on a new calibration method, the 18% increase of the Rhod-2 signal was calculated to correspond to a 43% increase of [Ca(2+)](m). There was also a close temporal relationship between the rise (time constant approximately 25 s) and fall (time constant approximately 65 s) of [Ca(2+)](m) and [NADH](m) when the pacing frequency was increased and decreased, respectively, suggesting that increased workload and [Ca(2+)](c) cause increased [Ca(2+)](m) and consequently up-regulation of the NADH production rate. PMID- 12124251 TI - Fluorescence intensity and lifetime distribution analysis: toward higher accuracy in fluorescence fluctuation spectroscopy. AB - Fluorescence fluctuation methods such as fluorescence correlation spectroscopy and fluorescence intensity distribution analysis (FIDA) have proven to be versatile tools for studying molecular interactions with single molecule sensitivity. Another well-known fluorescence technique is the measurement of the fluorescence lifetime. Here, we introduce a method that combines the benefits of both FIDA and fluorescence lifetime analysis. It is based on fitting the two dimensional histogram of the number of photons detected in counting time intervals of given width and the sum of excitation to detection delay times of these photons. Referred to as fluorescence intensity and lifetime distribution analysis (FILDA), the technique distinguishes fluorescence species on the basis of both their specific molecular brightness and the lifetime of the excited state and is also able to determine absolute fluorophore concentrations. The combined information yielded by FILDA results in significantly increased accuracy compared to that of FIDA or fluorescence lifetime analysis alone. In this paper, the theory of FILDA is elaborated and applied to both simulated and experimental data. The outstanding power of this technique in resolving different species is shown by quantifying the binding of calmodulin to a peptide ligand, thus indicating the potential for application of FILDA to similar problems in the life sciences. PMID- 12124252 TI - Modeling shape and topology of low-resolution density maps of biological macromolecules. AB - In the present work we develop an efficient way of representing the geometry and topology of volumetric datasets of biological structures from medium to low resolution, aiming at storing and querying them in a database framework. We make use of a new vector quantization algorithm to select the points within the macromolecule that best approximate the probability density function of the original volume data. Connectivity among points is obtained with the use of the alpha shapes theory. This novel data representation has a number of interesting characteristics, such as 1) it allows us to automatically segment and quantify a number of important structural features from low-resolution maps, such as cavities and channels, opening the possibility of querying large collections of maps on the basis of these quantitative structural features; 2) it provides a compact representation in terms of size; 3) it contains a subset of three dimensional points that optimally quantify the densities of medium resolution data; and 4) a general model of the geometry and topology of the macromolecule (as opposite to a spatially unrelated bunch of voxels) is easily obtained by the use of the alpha shapes theory. PMID- 12124253 TI - K(+) versus Na(+) ions in a K channel selectivity filter: a simulation study. AB - Molecular dynamics simulations of a bacterial potassium channel (KcsA) embedded in a phospholipid bilayer reveal significant differences in interactions of the selectivity filter with K(+) compared with Na(+) ions. K(+) ions and water molecules within the filter undergo concerted single-file motion in which they translocate between adjacent sites within the filter on a nanosecond timescale. In contrast, Na(+) ions remain bound to sites within the filter and do not exhibit translocation on a nanosecond timescale. Furthermore, entry of a K(+) ion into the filter from the extracellular mouth is observed, whereas this does not occur for a Na(+) ion. Whereas K(+) ions prefer to sit within a cage of eight oxygen atoms of the filter, Na(+) ions prefer to interact with a ring of four oxygen atoms plus two water molecules. These differences in interactions in the selectivity filter may contribute to the selectivity of KcsA for K(+) ions (in addition to the differences in dehydration energy between K(+) and Na(+)) and the block of KcsA by internal Na(+) ions. In our simulations the selectivity filter exhibits significant flexibility in response to changes in ion/protein interactions, with a somewhat greater distortion induced by Na(+) than by K(+) ions. PMID- 12124254 TI - Description and analysis of metabolic connectivity and dynamics in the human red blood cell. AB - The human red blood cell (hRBC) metabolic network is relatively simple compared with other whole cell metabolic networks, yet too complicated to study without the aid of a computer model. Systems science techniques can be used to uncover the key dynamic features of hRBC metabolism. Herein, we have studied a full dynamic hRBC metabolic model and developed several approaches to identify metabolic pools of metabolites. In particular, we have used phase planes, temporal decomposition, and statistical analysis to show hRBC metabolism is characterized by the formation of pseudoequilibrium concentration states. Such equilibria identify metabolic "pools" or aggregates of concentration variables. We proceed to define physiologically meaningful pools, characterize them within the hRBC, and compare them with those derived from systems engineering techniques. In conclusion, systems science methods can decipher detailed information about individual enzymes and metabolites within metabolic networks and provide further understanding of complex biological networks. PMID- 12124255 TI - Relating molecular flexibility to function: a case study of tubulin. AB - Microtubules (MT), along with a variety of associated motor proteins, are involved in a range of cellular functions including vesicle movement, chromosome segregation, and cell motility. MTs are assemblies of heterodimeric proteins, alpha beta-tubulins, the structure of which has been determined by electron crystallography of zinc-induced, pacilitaxel-stabilized tubulin sheets. These data provide a basis for examining relationships between structural features and protein function. Here, we study the fluctuation dynamics of the tubulin dimer with the aim of elucidating its functional motions relevant to substrate binding, polymerization/depolymerization and MT assembly. A coarse-grained model, harmonically constrained according to the crystal structure, is used to explore the global dynamics of the dimer. Our results identify six regions of collective motion, comprised of structurally close but discontinuous sequence fragments, observed only in the dimeric form, dimerization being a prerequisite for domain identification. Boundaries between regions of collective motions appear to act as linkages, found primarily within secondary-structure elements that lack sequence conservation, but are located at minima in the fluctuation curve, at positions of hydrophobic residues. Residue fluctuations within these domains identify the most mobile regions as loops involved in recognition of the adjacent regions. The least mobile regions are associated with nucleotide binding sites where lethal mutations occur. The functional coupling of motions between and within regions identifies three global motions: torsional and wobbling movements, en bloc, between the alpha- and beta-tubulin monomers, and stretching longitudinally. Further analysis finds the antitumor drug pacilitaxel (TaxotereR) to reduce flexibility in the M loop of the beta-tubulin monomer; an effect that may contribute to tightening lateral interactions between protofilaments assembled into MTs. Our analysis provides insights into relationships between intramolecular tubulin movements of MT organization and function. PMID- 12124257 TI - Relaxation kinetics and the glassiness of proteins: the case of bovine pancreatic trypsin inhibitor. AB - Folded proteins may be regarded as soft active matter under physiological conditions. The densely packed hydrophobic interior, the relatively molten hydrophilic exterior, and the spacer connecting these put together a large number of locally homogeneous regions. For the case of the bovine pancreatic trypsin inhibitor, with the aid of molecular dynamics simulations, we have demonstrated that the kinetics of the relaxation of the internal motions is highly concerted, manifesting the protein's heterogeneity, which may arise from variations in density, local packing, or the local energy landscape. This behavior is characterized in a stretched exponential decay described by an exponent of approximately 0.4 at physiological temperatures. Due to the trapped conformations, configurational entropy becomes smaller, and the associated stretch exponent drops to half of its value below the glass transition range. The temperature dependence of the inverse relaxation time closely follows the Vogel Tamman-Fulcher expression when the protein is biologically active. PMID- 12124256 TI - Protein unfolding transitions in an intrinsically unstable annexin domain: molecular dynamics simulation and comparison with nuclear magnetic resonance data. AB - Unfolding transitions of an intrinsically unstable annexin domain and the unfolded state structure have been examined using multiple approximately 10-ns molecular dynamics simulations. Three main basins are observed in the configurational space: native-like state, compact partially unfolded or intermediate compact state, and the unfolded state. In the native-like state fluctuations are observed that are nonproductive for unfolding. During these fluctuations, after an initial loss of approximately 20% of the core residue native contacts, the core of the protein transiently completely refolds to the native state. The transition from the native-like basin to the partially unfolded compact state involves approximately 75% loss of native contacts but little change in the radius of gyration or core hydration properties. The intermediate state adopts for part of the time in one of the trajectories a novel highly compact salt-bridge stabilized structure that can be identified as a conformational trap. The intermediate-to-unfolded state transition is characterized by a large increase in the radius of gyration. After an initial relaxation the unfolded state recovers a native-like topology of the domain. The simulated unfolded state ensemble reproduces in detail experimental nuclear magnetic resonance data and leads to a convincing complete picture of the unfolded domain. PMID- 12124258 TI - Multi-bead-and-spring model to interpret protein detachment studied by AFM force spectroscopy. AB - This article deals with the detachment of molecules (fibrinogen) from a surface studied experimentally with an atomic force microscope. The detachment (or rupture) forces are measured as a function of the retraction velocity and exhibit a clear dependence on this parameter, even though the interaction between the molecules and the surface are nonspecific. To interpret these data, a mechanical multi-bead-and-spring model is developed. It consists of one to several parallel, "molecular" springs connected to an extra spring representing the cantilever that is moved at constant velocity. The free end of each molecular spring terminates with a particle that interacts with the surface through a Lennard-Jones potential. This Brownian dynamics model is used to analyze the experimental findings. In the framework of this model, it appears that the fibrinogen molecule must be ascribed a stiffness much smaller than that of the cantilever. In addition, several bonds between the molecule and the surface must be taken into account for the range of the molecule-surface interaction not to be unrealistically small. In future work, this model will be extended to more complex mechanisms such as the detachment of cells from a surface. PMID- 12124259 TI - Dynamics of proteins in crystals: comparison of experiment with simple models. AB - The dynamic behavior of proteins in crystals is examined by comparing theory and experiments. The Gaussian network model (GNM) and a simplified version of the crystallographic translation libration screw (TLS) model are used to calculate mean square fluctuations of C(alpha) atoms for a set of 113 proteins whose structures have been determined by x-ray crystallography. Correlation coefficients between the theoretical estimations and experiment are calculated and compared. The GNM method gives better correlation with experimental data than the rigid-body libration model and has the added benefit of being able to calculate correlations between the fluctuations of pairs of atoms. By incorporating the effect of neighboring molecules in the crystal the correlation is further improved. PMID- 12124260 TI - A mathematical explanation of an increase in bacterial swimming speed with viscosity in linear-polymer solutions. AB - Bacterial swimming speed is sometimes known to increase with viscosity. This phenomenon is peculiar to bacterial motion. Berg and Turner (Nature. 278:349-351, 1979) indicated that the phenomenon was caused by a loose, quasi-rigid network formed by polymer molecules that were added to increase viscosity. We mathematically developed their concept by introducing two apparent viscosities and obtained results similar to the experimental data reported before. Addition of polymer improved the propulsion efficiency, which surpasses the decline in flagellar rotation rate, and the swimming speed increased with viscosity. PMID- 12124261 TI - Mechanisms for temporal tuning and filtering by postsynaptic signaling pathways. AB - Networks of signaling pathways perform complex temporal decoding functions in diverse biological systems, including the synapse, development, and bacterial chemotaxis. This paper examines temporal filtering and tuning properties of synaptic signaling pathways as a possible substrate for emergent temporal decoding. A mass action kinetic model of 16 synaptic signaling pathways was used to dissect out the contribution of these pathways in linear cascades and when coupled to form a network. The model predicts two primary mechanisms of temporal tuning of pathways: a weighted summation of responses of pathways with different timings and the presence of biochemical feedback loop(s) with emergent dynamics. Regulatory inputs act differently on these two tuning mechanisms. In the first case, regulators act like a gain-control on pathways with different intrinsic tuning. In the case of feedback loops, the temporal properties of the loop itself are changed. These basic tuning mechanisms may underlie specialized temporal tuning functions in more complex signaling systems in biology. PMID- 12124262 TI - Steered molecular dynamics simulations on the "tail helix latch" hypothesis in the gelsolin activation process. AB - The molecular basis of the "tail helix latch" hypothesis in the gelsolin activation process has been studied by using the steered molecular dynamics simulations. In the present nanosecond scale simulations, the tail helix of gelsolin was pulled away from the S2 binding surface, and the required forces were calculated, from which the properties of binding between the tail helix and S2 domain and their dynamic unbinding processes were obtained. The force profile provides a detailed rupture mechanism that includes six major unbinding steps. In particular, the hydrogen bonds formed between Arg-207 and Asp-744 and between Arg 221 and Leu-753 are of the most important interaction pairs. The two hydrogen bond "clamps" stabilize the complex. The subsequent simulation on Arg-207-Ala (R207A) mutation of gelsolin indicated that this mutation facilitates the unbinding of the tail helix and that the contribution of the hydrogen bond between Arg-207 and Asp-744 to the binding is more than 50%, which offers a new clue for further mutagenesis study on the activation mechanism of gelsolin. Surrounding water molecules enhance the stability of the tail helix and facilitate the rupture process. Additionally, temperature also has a significant effect on the conformation of the arginine and arginine-related interactions, which revealed the molecular basis of the temperature dependence in gelsolin activation. PMID- 12124263 TI - OmpA: a pore or not a pore? Simulation and modeling studies. AB - The bacterial outer membrane protein OmpA is composed of an N-terminal 171 residue beta-barrel domain (OmpA(171)) that spans the bilayer and a periplasmic, C-terminal domain of unknown structure. OmpA has been suggested to primarily serve a structural role, as no continuous pore through the center of the barrel can be discerned in the crystal structure of OmpA(171). However, several groups have recorded ionic conductances for bilayer-reconstituted OmpA(171). To resolve this apparent paradox we have used molecular dynamics (MD) simulations on OmpA(171) to explore the conformational dynamics of the protein, in particular the possibility of transient formation of a central pore. A total of 19 ns of MD simulations of OmpA(171) have been run, and the results were analyzed in terms of 1) comparative behavior of OmpA(171) in different bilayer and bilayer-mimetic environments, 2) solvation states of OmpA(171), and 3) pore characteristics in different MD simulations. Significant mobility was observed for residues and water molecules within the beta-barrel. A simulation in which putative gate region side chains of the barrel interior were held in a non-native conformation led to an open pore, with a predicted conductance similar to experimental measurements. The OmpA(171) pore has been shown to be somewhat more dynamic than suggested by the crystal structure. A gating mechanism is proposed to explain its documented channel properties, involving a flickering isomerization of Arg138, forming alternate salt bridges with Glu52 (closed state) and Glu128 (open state). PMID- 12124264 TI - Reaction diffusion model of the enzymatic erosion of insoluble fibrillar matrices. AB - Predicting the time course of in vivo biodegradation is a key issue in the design of an increasing number of biomedical applications such as sutures, tissue analogs and drug-delivery devices. The design of such biodegradable devices is hampered by the absence of quantitative models for the enzymatic erosion of solid protein matrices. In this work, we derive and simulate a reaction diffusion model for the enzymatic erosion of fibrillar gels that successfully reproduces the main qualitative features of this process. A key aspect of the proposed model is the incorporation of steric hindrance into the standard Michaelis-Menten scheme for enzyme kinetics. In the limit of instantaneous diffusion, the model equations are analogous to the standard equations for enzymatic degradation in solution. Invoking this analogy, the total quasi-steady-state approximation is used to derive approximate analytical solutions that are valid for a wide range of in vitro conditions. Using these analytical approximations, an experimental theoretical method is derived to unambiguously estimate all the kinetic model parameters. Moreover, the analytical approximations correctly describe the characteristic hyperbolic dependence of the erosion rate on enzyme concentration and the zero-order erosion of thin fibers. For definiteness, the analysis of published experimental results of enzymatic degradation of fibrillar collagen is demonstrated, and the role of diffusion in these experiments is elucidated. PMID- 12124265 TI - Molecular dynamics simulations of the first steps of the reaction catalyzed by HIV-1 protease. AB - The mechanism of the first steps of the reaction catalyzed by HIV-1 protease was studied through molecular dynamics simulations. The potential energy surface in the active site was generated using the approximate valence bond method. The approximate valence bond (AVB) method was parameterized based on density functional calculations. The surrounding protein and explicit water environment was modeled with conventional, classical force field. The calculations were performed based on HIV-1 protease complexed with the MVT-101 inhibitor that was modified to a model substrate. The protonation state of the catalytic aspartates was determined theoretically. Possible reaction mechanisms involving the lytic water molecule are accounted for in this study. The modeled steps include the dissociation of the lytic water molecule and proton transfer onto Asp-125, the nucleophilic attack followed by a proton transfer onto peptide nitrogen. The simulations show that in the active site most preferable energetically are structures consisting of ionized or polarized molecular fragments that are not accounted for in conventional molecular dynamics. The mobility of the lytic water molecule, the dynamics of the hydrogen bond network, and the conformation of the aspartates in the active center were analyzed. PMID- 12124266 TI - Extreme pathway analysis of human red blood cell metabolism. AB - The development of high-throughput technologies and the resulting large-scale data sets have necessitated a systems approach to the analysis of metabolic networks. One way to approach the issue of complex metabolic function is through the calculation and interpretation of extreme pathways. Extreme pathways are a mathematically defined set of generating vectors that describe the conical steady state solution space for flux distributions through an entire metabolic network. Herein, the extreme pathways of the well-characterized human red blood cell metabolic network were calculated and interpreted in a biochemical and physiological context. These extreme pathways were divided into groups based on such criteria as their cofactor and by-product production, and carbon inputs including those that 1) convert glucose to pyruvate; 2) interchange pyruvate and lactate; 3) produce 2,3-diphosphoglycerate that binds to hemoglobin; 4) convert inosine to pyruvate; 5) induce a change in the total adenosine pool; and 6) dissipate ATP. Additionally, results from a full kinetic model of red blood cell metabolism were predicted based solely on an interpretation of the extreme pathway structure. The extreme pathways for the red blood cell thus give a concise representation of red blood cell metabolism and a way to interpret its metabolic physiology. PMID- 12124267 TI - Protein folding pathways and kinetics: molecular dynamics simulations of beta strand motifs. AB - The folding pathways and the kinetic properties for three different types of off lattice four-strand antiparallel beta-strand protein models interacting via a hybrid Go-type potential have been investigated using discontinuous molecular dynamics simulations. The kinetic study of protein folding was conducted by temperature quenching from a denatured or random coil state to a native state. The progress parameters used in the kinetic study include the squared radius of gyration R(2)(g), the fraction of native contacts within the protein as a whole Q, and between specific strands Q(ab). In the time series of folding, the denatured proteins undergo a conformational change toward the native state. The model proteins exhibit a variety of kinetic folding pathways that include a fast track folding pathway without passing through an intermediate and multiple pathways with trapping into more than one intermediate. The kinetic folding behavior of the beta-strand proteins strongly depends on the native-state geometry of the model proteins and the size of the bias gap g, an artificial measure of a model protein's preference for its native state. PMID- 12124268 TI - Fluid shear regulates the kinetics and molecular mechanisms of activation dependent platelet binding to colon carcinoma cells. AB - This study was undertaken to investigate the kinetics and molecular requirements of platelet binding to tumor cells in bulk suspensions subjected to a uniform linear shear field, using a human colon adenocarcinoma cell line (LS174T) as a model. The effects of shear rate (20-1000 s(-1)), shear exposure time (30-300 s), shear stress (at constant shear rate by adjusting the viscosity of the medium from 1.3-2.6 cP), cell concentration, and platelet activation on platelet-LS174T heteroaggregation were assessed. The results indicate that hydrodynamic shear induced collisions augment platelet-LS174T binding, which is further potentiated by thrombin/GPRP-NH(2). Peak adhesion efficiency occurs at low shear and decreases with increasing shear. Intercellular contact duration is the predominant factor limiting heteroaggregation at shear rates up to 200 s(-1), whereas these interactions become shear stress-sensitive at > or = 400 s(-1). Heteroaggregation increases with platelet concentration due to an elevation of the intercellular collision frequency, whereas adhesion efficiency remains nearly constant. Moreover, hydrodynamic shear affects the receptor specificity of activation-dependent platelet binding to LS174T cells, as evidenced by the transition from a P-selectin-independent/Arg-Gly-Asp (RGD)-dependent process at 100 s(-1) to a P-selectin/alpha(IIb)beta(3)-dependent interaction at 800 s(-1). This study demonstrates that platelet activation and a fluid-mechanical environment representative of the vasculature affect platelet-tumor cell adhesive interactions pertinent to the process of blood-borne metastasis. PMID- 12124269 TI - Exocytosis in bovine chromaffin cells: studies with patch-clamp capacitance and FM1-43 fluorescence. AB - In response to physiological stimuli, neuroendocrine cells secrete neurotransmitters through a Ca(2+)-dependent fusion of secretory granules with the plasma membrane. We studied insertion of granules in bovine chromaffin cells using capacitance as a measure of plasma membrane area and fluorescence of a membrane marker FM1-43 as a measure of exocytosis. Intracellular dialysis with [Ca(2+)] (1.5-100 microM) evoked massive exocytosis that was sufficient to double plasma membrane area but did not swell cells. In principle, in the absence of endocytosis, the addition of granule membrane would be anticipated to produce similar increases in the capacitance and FM1-43 fluorescence responses. However, when endocytosis was minimal, the changes in capacitance were markedly larger than the corresponding changes in FM1-43 fluorescence. Moreover, the apparent differences between capacitance and FM1-43 fluorescence changes increased with larger exocytic responses, as more granules fused with the plasma membrane. In experiments in which exocytosis was suppressed, increasing membrane tension by osmotically induced cell swelling increased FM1-43 fluorescence, suggesting that FM1-43 fluorescence is sensitive to changes in the membrane tension. Thus, increasing membrane area through exocytosis does not swell chromaffin cells but may decrease membrane tension. PMID- 12124271 TI - The role of Trp side chains in tuning single proton conduction through gramicidin channels. AB - We present an extensive set of measurements of proton conduction through gramicidin A (gA), B (gB), and M (gM) homodimer channels which have 4, 3, or 0 Trp residues at each end of the channel, respectively. In gA we find a shoulder separating two domains of conductance increasing with concentration, confirming the results of Eisenman, G., B. Enos, J. Hagglund, and J. Sandblom. 1980. Ann. NY. Acad. Sci. 339:8-20. In gB, the shoulder is shifted by approximately 1/2 pH unit to higher H(+) concentrations and is very sharply defined. No shoulder appears in the gM data, but an associated transition from sublinear to superlinear I-V values occurs at a 100-fold higher [H(+)] in gM than in gA. The data in the low concentration domain are analyzed using a configuration space model of single-proton conduction, assuming that the difference in the proton potential of mean force (PMF) between gA and its analogs is constant, similar to the results of Anderson, D., R. B. Shirts, T. A. Cross, and D. D. Busath. 2001. Biophys. J. 81:1255-1264. Our results suggest that the average amplitudes of the calculated proton PMFs are nearly correct, but that the water reorientation barrier calculated for gA by molecular dynamics using the PM6 water model (Pomes, R., and B. Roux. 1997. Biophys. J. 72:246a) must be reduced in amplitude by 1.5 kcal/mol or more, and is not rate-limiting for gA. PMID- 12124270 TI - Determination of cellular strains by combined atomic force microscopy and finite element modeling. AB - Many organs adapt to their mechanical environment as a result of physiological change or disease. Cells are both the detectors and effectors of this process. Though many studies have been performed in vitro to investigate the mechanisms of detection and adaptation to mechanical strains, the cellular strains remain unknown and results from different stimulation techniques cannot be compared. By combining experimental determination of cell profiles and elasticities by atomic force microscopy with finite element modeling and computational fluid dynamics, we report the cellular strain distributions exerted by common whole-cell straining techniques and from micromanipulation techniques, hence enabling their comparison. Using data from our own analyses and experiments performed by others, we examine the threshold of activation for different signal transduction processes and the strain components that they may detect. We show that modulating cell elasticity, by increasing the F-actin content of the cytoskeleton, or cellular Poisson ratio are good strategies to resist fluid shear or hydrostatic pressure. We report that stray fluid flow in some substrate-stretch systems elicits significant cellular strains. In conclusion, this technique shows promise in furthering our understanding of the interplay among mechanical forces, strain detection, gene expression, and cellular adaptation in physiology and disease. PMID- 12124272 TI - The chloroplast protein import channel Toc75: pore properties and interaction with transit peptides. AB - The channel properties of Toc75 (the protein import pore of the outer chloroplastic membrane) were further characterized by electrophysiological measurements in planar lipid bilayers. After improvement of the Toc75 reconstitution procedure the voltage dependence of the channel open probability resembled those observed for other beta-barrel pores. Studies concerning the pore size of the reconstituted Toc75 indicate the presence of a narrow restriction zone corresponding to the selectivity filter and a wider pore vestibule with diameters of approximately 14 A and 26 A, respectively. Interactions between Toc75 and different peptides (a genuine chloroplastic transit peptide, a synthetic peptide resembling a transit peptide, and a mitochondrial presequence) show that Toc75 itself is able to differentiate between these peptides and the recognition is based on both conformational and electrostatic interactions. PMID- 12124273 TI - Energetic localization of saxitoxin in its channel binding site. AB - Saxitoxin (STX) selectively blocks the voltage-gated sodium channel at the outer vestibule lined by P-loops of the four domains. Neosaxitoxin has an additional OH group at the N1 position of the 1,2,3 guanidinium (N1-OH) that interacts with domains I and IV of the Na(+) channel. Determination of a second toxin interaction with the channel would fix the location of STX. Gonyautoxin 2,3 and Gonyautoxin 1,4 are C-11 sulfated derivatives of saxitoxin and neosaxitoxin, respectively. We used these variants to constrain the STX docking orientation by energetically localizing the C-11 sulfate in the outer vestibule. Interactions between the C-11 sulfate and each of the four domains of the channel were determined by a systematic approach to mutant cycle analysis in which all known carboxyl groups important for site 1 toxin binding were neutralized, allowing energetic triangulation of the toxin sulfate and overcoming some limitations of mutant cycles. Toxin IC(50)s were measured by two-electrode voltage clamp from Xenopus oocytes injected with the channel mRNA. Three unique types of analysis based on the coupling results localized the C-11 sulfate between domains III and IV. Combined with our previous report, the data establish the orientation of STX in the outer vestibule and confirm the clockwise arrangement of the channel domains. PMID- 12124274 TI - Gating of heteromeric retinal rod channels by cyclic AMP: role of the C-terminal and pore domains. AB - Cyclic nucleotide-gated channels are tetramers composed of homologous alpha and beta subunits. C-terminal truncation mutants of the alpha and beta subunits of the retinal rod channel were expressed in Xenopus oocytes, and analyzed for cGMP- and cAMP-induced currents (single-channel records and macroscopic currents). When the alpha subunit truncated downstream of the cGMP-binding site (alpha D608stop) is co-injected with truncated beta subunits, the heteromeric channels present a drastic increase of cAMP sensitivity. A partial effect is observed with heteromeric alpha R656stop-containing channels, while alpha K665stop-containing channels behave like alpha wt/beta wt. The three truncated alpha subunits have wild-type activity when expressed alone. Heteromeric channels composed of alpha wt or truncated alpha subunits and chimeric beta subunits containing the pore domain of the alpha subunit have the same cAMP sensitivity as alpha-only channels. The results disclose the key role of two domains distinct from the nucleotide binding site in the gating of heteromeric channels by cAMP: the pore of the beta subunit, which has an activating effect, and a conserved domain situated downstream of the cGMP-binding site in the alpha subunit (I609-K665), which inhibits this effect. PMID- 12124275 TI - Macroscopic consequences of the action of phospholipase C on giant unilamellar liposomes. AB - Macroscopic consequences of the formation of diacylglycerol by phospholipase C (PC-PLC) in giant 1-stearoyl-2-oleoyl-sn-glycero-3-phosphocholine (SOPC) unilamellar vesicles (GUVs, diameter 10-100 microm) were studied by phase contrast and fluorescence microscopy. PC-PLC caused a series of fast stepwise shrinkages of fluid SOPC GUVs, continuing until the vesicle disappeared beyond the optical resolution of the microscope. The presence of N-palmitoyl sphingomyelin (mole fraction X = 0.25) in the GUVs did not affect the outcome of the PC-PLC reaction. In addition to hydrolysis, PC-PLC induced adhesion of vicinal vesicles. When multilamellar SOPC vesicles were used only a minor decrease in their diameter was evident suggesting that PC-PLC can exert its hydrolytic activity only in the outer monolayer. A series of stepwise shrinkages was observed also for 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) GUVs above their main phase transition temperature, T(m), i.e., when the bilayer is in the liquid crystalline state. However, this process was not observed for DMPC GUVs in the gel state, below T(m). These results are supported by the enhanced activity of PC-PLC upon exceeding T(m) of DMPC large unilamellar vesicles (diameter approximately 0.1 microm) used as a substrate. Studies on SOPC monolayers revealed that PC-PLC can exert its hydrolytic activity only at surface pressures below approximately 30 mN/m. Accordingly, the lack of changes in the gel state DMPC GUVs could be explained by the equilibrium lateral pressure in these vesicles exceeding this critical value. PMID- 12124276 TI - Mechanisms of equinatoxin II-induced transport through the membrane of a giant phospholipid vesicle. AB - Protein equinatoxin II from sea anemone Actinia equina L. was used to form pores in phospholipid membranes. We studied the effect of these pores on the net transmembrane transport of sucrose and glucose by observing single giant (cell size) vesicles under the phase contrast microscope. Sugar composition in the vesicle was determined by measuring the width of the halo, which appears around the vesicle in the phase contrast image. The transport of sugars was induced when a vesicle, filled with the sucrose solution, was transferred into the isomolar environment of a glucose solution with added equinatoxin II. Typically, a vesicle grew to a critical size, then the membrane broke by bursting and the vesicle shrank, started to grow again, and the whole process was repeated. The consecutive membrane breaks occurred in the same spot. The observed behavior was interpreted by the diffusion flow of the glucose molecules through the equinatoxin II-induced pores and the consequent increase of the vesicle water content. The burst relaxed the critically strained membrane, which then apparently resealed. A mathematical model of the described behavior was developed and was used to obtain the equinatoxin II-induced membrane permeability for the glucose molecules. Its dependence on the equinatoxin II concentration is in agreement with the previous reports. PMID- 12124277 TI - Interactions of adriamycin, cytochrome c, and serum albumin with lipid monolayers containing poly(ethylene glycol)-ceramide. AB - Poly(ethylene glycol)(2000)C(20)ceramide (PEG-Cer) containing monolayers at an air/water interface were characterized by measuring their surface pressure versus area/molecule (pi-A) and surface potential versus area/molecule (Delta V-A) isotherms. The behavior of pi-A as well as Delta V versus lipid density (Delta V n) and Delta V-pi isotherms for PEG-Cer are in keeping with two transitions of the lipopolymer, starting at pi approximately equal to 9 and 21 mN/m. We also investigated the effects of PEG-Cer on the binding of adriamycin, cytochrome c and bovine serum albumin to monolayers containing varying mole fractions X of PEG Cer. PEG-Cer impedes the penetration of these ligands into lipid monolayers with similar effects at both X = 0.04 and 0.08. This effect of PEG-Cer depends on the conformation of the lipopolymer and the interactions between the lipid surface and the surface-interacting molecule as well as the size of the latter. PMID- 12124278 TI - Nerve growth factor signals via preexisting TrkA receptor oligomers. AB - Nerve growth factor (NGF) promotes neuronal survival and differentiation by activating TrkA receptors. Similar to other receptor tyrosine kinases, ligand induced dimerization is thought to be required for TrkA receptor activation. To study this process, we expressed TrkA receptors in Xenopus laevis oocytes and analyzed their response to NGF by using a combination of functional, biochemical, and structural approaches. TrkA receptor protein was detected in the membrane fraction of oocytes injected with TrkA receptor cRNA, but not in uninjected or mock-injected oocytes. Application of NGF to TrkA receptor-expressing oocytes promoted tyrosine phosphorylation and activated an oscillating transmembrane inward current, indicating that the TrkA receptors were functional. Freeze fracture electron microscopic analysis demonstrated novel transmembrane particles in the P-face (protoplasmic face) of oocytes injected with TrkA cRNA, but not in uninjected or mock injected oocytes. Incubating TrkA cRNA-injected oocytes with the transcriptional inhibitor actinomycin D did not prevent the appearance of these P-face particles or electrophysiological responses to NGF, demonstrating that they did not arise from de novo transcription of an endogenous Xenopus oocyte gene. The appearance of these particles in the plasma membrane correlated with responsiveness to NGF as detected by electrophysiological analysis and receptor phosphorylation, indicating that these novel P-face particles were TrkA receptors. The dimensions of these particles (8.6 x 10 nm) were too large to be accounted for by TrkA monomers, suggesting the formation of TrkA receptor oligomers. Application of NGF did not lead to a discernible change in the size or shape of these TrkA receptor particles during an active response. These results indicate that in Xenopus oocytes, NGF activates signaling via pre-formed TrkA receptor oligomers. PMID- 12124279 TI - The effects of acyl chain length and saturation of diacylglycerols and phosphatidylcholines on membrane monolayer curvature. AB - The second messenger, diacylglycerol (DAG), introduces negative curvature in phospholipid monolayers and strongly induces the lamellar (L(alpha)) to reverse hexagonal (H(II)) phase transition. The chain lengths and degree of unsaturation of symmetric DAGs influence this effect. Within dioleoylphosphatidylcholine (DOPC) monolayers, the apparent spontaneous radius of curvature (R(0)) of the short, saturated dicaprylglycerol (C10-DCG) itself was determined to be -13.3 A, compared with an R(0) value of -10.1 A for the long, di-monounsaturated dioleoylglycerol (C18-DOG). Such increased length and unsaturation of the DAG acyl chains produces this small change. Di-saturated phosphatidylcholines (PCs) with equal length chains (from C10-C18) with 25 mol % DOG do not form the H(II) phase, even under the unstressed conditions of excess water and alkane. Di unsaturated PCs with equal chain length (from C14-C18) with 25 mol % DOG do form the H(II) phase. Asymmetric chained PCs (position 1 saturated with varying lengths, position 2 differentially unsaturated with varying lengths) all form the H(II) phase in the presence of 25 mol % DOG. As a general rule for PCs, their unsaturation is critical for the induction of the H(II) phase by DOG. The degree of curvature stress induced by the second messenger DOG in membranes, and any protein that might be affected by it, would appear to depend on chain unsaturation of neighboring PCs. PMID- 12124280 TI - Human apolipoprotein H may have various orientations when attached to lipid layer. AB - Apolipoprotein H (ApoH), also known as beta(2)-glycoprotein I, is a plasma glycoprotein with its in vivo physiological and pathogenic roles being closely related to its interaction with negatively charged membranes. Although the three dimensional crystal structure of ApoH has been recently solved, direct evidence about the spatial state of ApoH on the membrane is still lacking. In this work, the interactions of ApoH with the lipid layer are studied by a combination of lipid monolayer approach and surface concentration determination. The spatial state of the orientation of ApoH on the lipid layer is investigated by analyzing the process of membrane-attached ApoH molecules being extruded out from the phospholipid monolayer by compression. The results show that on neutral lipid layer ApoH has an upright orientation, which is not sensitive to the phase state of the lipid layer. However, on acidic lipid layer, ApoH may have two forms of orientation. One is an upright orientation in the liquid phase region, and the other is flat orientation on the condensed domain region. The variation of the spatial state of ApoH on the lipid layer may relate to a variety of its physiological functions. PMID- 12124281 TI - Structural transitions in short-chain lipid assemblies studied by (31)P-NMR spectroscopy. AB - The self-assembled supramolecular structures of diacylphosphatidylcholine (diC(n)PC), diacylphosphatidylethanolamine (diC(n)PE), diacylphosphatidyglycerol (diC(n)PG), and diacylphosphatidylserine (diC(n)PS) were investigated by (31)P nuclear magnetic resonance (NMR) spectroscopy as a function of the hydrophobic acyl chain length. Short-chain homologs of these lipids formed micelles, and longer-chain homologs formed bilayers. The shortest acyl chain lengths that supported bilayer structures depended on the headgroup of the lipids. They increased in the order PE (C(6)) < PC (C(9)) < or = PS (C(9) or C(10)) < PG (C(11) or C(12)). This order correlated with the effective headgroup area, which is a function of the physical size, charge, hydration, and hydrogen-bonding capacity of the four headgroups. Electrostatic screening of the headgroup charge with NaCl reduced the effective headgroup area of PS and PG and thereby decreased the micelle-to-bilayer transition of these lipid classes to shorter chain lengths. The experimentally determined supramolecular structures were compared to the assembly states predicted by packing constraints that were calculated from the hydrocarbon-chain volume and effective headgroup area of each lipid. The model accurately predicted the chain-length threshold for bilayer formation if the relative displacement of the acyl chains of the phospholipid were taken into account. The model also predicted cylindrical rather than spherical micelles for all four diacylphospholipid classes and the (31)P-NMR spectra provided evidence for a tubular network that appeared as an intermediate phase at the micelle-to bilayer transition. The free energy of micellization per methylene group was independent of the structure of the supramolecular assembly, but was -0.95 kJ/mol (-0.23 kcal/mol) for the PGs compared to -2.5 kJ/mol (-0.60 kcal/mol) for the PCs. The integral membrane protein OmpA did not change the bilayer structure of thin (diC(10)PC) bilayers. PMID- 12124282 TI - Membrane composition determines pardaxin's mechanism of lipid bilayer disruption. AB - Pardaxin is a membrane-lysing peptide originally isolated from the fish Pardachirus marmoratus. The effect of the carboxy-amide of pardaxin (P1a) on bilayers of varying composition was studied using (15)N and (31)P solid-state NMR of mechanically aligned samples and differential scanning calorimetry (DSC). (15)N NMR spectroscopy of [(15)N-Leu(19)]P1a found that the orientation of the peptide's C-terminal helix depends on membrane composition. It is located on the surface of lipid bilayers composed of 1-palmitoyl-2-oleoyl-phosphatidylcholine (POPC) and is inserted in lipid bilayers composed of 1,2-dimyristoyl phosphatidylcholine (DMPC). The former suggests a carpet mechanism for bilayer disruption whereas the latter is consistent with a barrel-stave mechanism. The (31)P chemical shift NMR spectra showed that the peptide significantly disrupts lipid bilayers composed solely of zwitterionic lipids, particularly bilayers composed of POPC, in agreement with a carpet mechanism. P1a caused the formation of an isotropic phase in 1-palmitoyl-2-oleoyl-phosphatidylethanolamine (POPE) lipid bilayers. This, combined with DSC data that found P1a reduced the fluid lamellar-to-inverted hexagonal phase transition temperature at very low concentrations (1:50,000), is interpreted as the formation of a cubic phase and not micellization of the membrane. Experiments exploring the effect of P1a on lipid bilayers composed of 4:1 POPC:cholesterol, 4:1 POPE:cholesterol, 3:1 POPC:1 palmitoyl-2-oleoyl-phosphatidylglycerol (POPG), and 3:1 POPE:POPG were also conducted, and the presence of anionic lipids or cholesterol was found to reduce the peptide's ability to disrupt bilayers. Considered together, these data demonstrate that the mechanism of P1a is dependent on membrane composition. PMID- 12124283 TI - Exploration of molecular interactions in cholesterol superlattices: effect of multibody interactions. AB - Experimental evidences have indicated that cholesterol may adapt highly regular lateral distributions (i.e., superlattices) in a phospholipid bilayer. We investigated the formations of superlattices at cholesterol mole fraction of 0.154, 0.25, 0.40, and 0.5 using Monte Carlo simulation. We found that in general, conventional pairwise-additive interactions cannot produce superlattices. Instead, a multibody (nonpairwise) interaction is required. Cholesterol superlattice formation reveals that although the overall interaction between cholesterol and phospholipids is favorable, it contains two large opposing components: an interaction favoring cholesterol-phospholipid mixing and an unfavorable acyl chain multibody interaction that increases nonlinearly with the number of cholesterol contacts. The magnitudes of interactions are in the order of kT. The physical origins of these interactions can be explained by our umbrella model. They most likely come from the requirement for polar phospholipid headgroups to cover the nonpolar cholesterol to avoid the exposure of cholesterol to water and from the sharp decreasing of acyl chain conformation entropy due to cholesterol contact. This study together with our previous work demonstrate that the driving force of cholesterol-phospholipid mixing is a hydrophobic interaction, and multibody interactions dominate others over a wide range of cholesterol concentration. PMID- 12124284 TI - Interfacing molecular dynamics and macro-scale simulations for lipid bilayer vesicles. AB - A continuum-level model for a giant unilamellar vesicle (GUV) is bridged to a corresponding atomistic model of a dimyristoylphosphatidylcholine (DMPC) bilayer at various cholesterol concentrations via computation of the bulk modulus. The bulk modulus and other microscopically determined parameters are passed to a continuum-level model operating in time- and length-scales orders of magnitude beyond that which is accessible by atomistic-level simulation. The continuum level simulation method used is the material point method (MPM), and the particular variation used here takes advantage of the spherical nature of many GUVs. An osmotic pressure gradient due to a solvent concentration change is incorporated into the continuum-level simulation, resulting in osmotic swelling of the vesicle. The model is then extended to treat mixtures of DMPC and cholesterol, where small domains of different composition are considered. PMID- 12124287 TI - Direct x-ray observation of a single hexagonal myofilament lattice in native myofibrils of striated muscle. AB - A striated muscle fiber consists of thousands of myofibrils with crystalline hexagonal myofilament lattices. Because the lattices are randomly oriented, the fiber gives rise to an equatorial x-ray diffraction pattern, which is essentially a rotary-averaged "powder diffraction," carrying only information about the distance between the lattice planes. We were able to record an x-ray diffraction pattern from a single myofilament lattice, very likely originating from a single myofibril from the flight muscle of a bumblebee, by orienting the incident x-ray microbeam along the myofibrillar axis (end-on diffraction). The pattern consisted of a number of hexagonally symmetrical diffraction spots whose originating lattice planes were readily identified. This also held true for some of the weak higher order reflections. The spot-like appearance of reflections implies that the lattice order is extremely well maintained for a distance of millimeters, covering up to a thousand of approximately 2.5-microm-long sarcomeres connected in series. The results open the possibility of applying the x-ray microdiffraction technique to study many other micrometer-sized assemblies of functional biomolecules in the cell. PMID- 12124285 TI - Troponin-tropomyosin: an allosteric switch or a steric blocker? AB - The interaction of myosin subfragment 1 (S1) with actin-tropomyosin-troponin (regulated actin) is highly nucleotide dependent. The binding of S1 or S1-ADP (but not S1-ATP nor N,N'-rho-phenylenedimaleimide-modified S1-ATP) to regulated actin activates ATP hydrolysis even in the absence of Ca(2+). Investigations with S1 and S1-ADP have led to the idea that some actin sites are directly blocked toward the binding of S1 either by tropomyosin or troponin. The blocked state is thought to occur only at ionic strengths greater than 50 mM. The question is whether nonactivating S1 binding is blocked under the same conditions. We show that troponin inhibits binding of the nonactivating state, N,N'-rho phenylenedimaleimide-S1-ATP, to actin but only when tropomyosin is absent. A lag in the rate of binding of activating S1 to actin (an indicator of the blocked state) occurs only in the presence of tropomyosin. Thus, tropomyosin inhibits binding of rigor S1 but not S1-ATP-like states. No evidence for an ionic strength dependent change in the mechanism of regulation was observed either from measurements of the rate of activating S1 binding or from the equilibrium binding of nonactivating S1 to actin. At all conditions examined, N,N'-rho phenylenedimaleimide-S1-ATP bound to regulated actin in the absence of Ca(2+). These results support the view of regulation in which tropomyosin movement is an allosteric switch that is modulated by activating myosin binding but that does not function solely by regulating myosin binding. PMID- 12124286 TI - Polarized fluorescence depletion reports orientation distribution and rotational dynamics of muscle cross-bridges. AB - The method of polarized fluorescence depletion (PFD) has been applied to enhance the resolution of orientational distributions and dynamics obtained from fluorescence polarization (FP) experiments on ordered systems, particularly in muscle fibers. Previous FP data from single fluorescent probes were limited to the 2(nd)- and 4(th)-rank order parameters, and , of the probe angular distribution (beta) relative to the fiber axis and , a coefficient describing the extent of rapid probe motions. We applied intense 12-micros polarized photoselection pulses to transiently populate the triplet state of rhodamine probes and measured the polarization of the ground state depletion using a weak interrogation beam. PFD provides dynamic information describing the extent of motions on the time scale between the fluorescence lifetime (e.g., 4 ns) and the duration of the photoselection pulse and it potentially supplies information about the probe angular distribution corresponding to order parameters above rank 4. Gizzard myosin regulatory light chain (RLC) was labeled with the 6-isomer of iodoacetamidotetramethylrhodamine and exchanged into rabbit psoas muscle fibers. In active contraction, dynamic motions of the RLC on the PFD time scale were intermediate between those observed in relaxation and rigor. The results indicate that previously observed disorder of the light chain region in contraction can be ascribed principally to dynamic motions on the microsecond time scale. PMID- 12124288 TI - Direct modeling of x-ray diffraction pattern from skeletal muscle in rigor. AB - Available high-resolution structures of F-actin, myosin subfragment 1 (S1), and their complex, actin-S1, were used to calculate a 2D x-ray diffraction pattern from skeletal muscle in rigor. Actin sites occupied by myosin heads were chosen using a "principle of minimal elastic distortion energy" so that the 3D actin labeling pattern in the A-band of a sarcomere was determined by a single parameter. Computer calculations demonstrate that the total off-meridional intensity of a layer line does not depend on disorder of the filament lattice. The intensity of the first actin layer A1 line is independent of tilting of the "lever arm" region of the myosin heads. Myosin-based modulation of actin labeling pattern leads not only to the appearance of the myosin and "beating" actin-myosin layer lines in rigor diffraction patterns, but also to changes in the intensities of some actin layer lines compared to random labeling. Results of the modeling were compared to experimental data obtained from small bundles of rabbit muscle fibers. A good fit of the data was obtained without recourse to global parameter search. The approach developed here provides a background for quantitative interpretation of the x-ray diffraction data from contracting muscle and understanding structural changes underlying muscle contraction. PMID- 12124289 TI - Probing protein-DNA interactions by unzipping a single DNA double helix. AB - We present unzipping force analysis of protein association (UFAPA) as a novel and versatile method for detection of the position and dynamic nature of protein-DNA interactions. A single DNA double helix was unzipped in the presence of DNA binding proteins using a feedback-enhanced optical trap. When the unzipping fork in a DNA reached a bound protein molecule we observed a dramatic increase in the tension in the DNA, followed by a sudden tension reduction. Analysis of the unzipping force throughout an unbinding "event" revealed information about the spatial location and dynamic nature of the protein-DNA complex. The capacity of UFAPA to spatially locate protein-DNA interactions is demonstrated by noncatalytic restriction mapping on a 4-kb DNA with three restriction enzymes (BsoBI, XhoI, and EcoRI). A restriction map for a given restriction enzyme was generated with an accuracy of approximately 25 bp. UFAPA also allows direct determination of the site-specific equilibrium association constant (K(A)) for a DNA-binding protein. This capability is demonstrated by measuring the cation concentration dependence of K(A) for EcoRI binding. The measured values are in good agreement with previous measurements of K(A) over an intermediate range of cation concentration. These results demonstrate the potential utility of UFAPA for future studies of site-specific protein-DNA interactions. PMID- 12124290 TI - Dimensions of plectonemically supercoiled DNA. AB - With a view to determine the configuration and regularity of plectonemically supercoiled DNA, we have measured the small angle neutron scattering from pUC18 plasmid in saline solutions. Furthermore, we have derived the mathematical expression for the single chain scattering function (form factor) of a superhelical structure, including the longitudinal and transverse interference over the plectonemic pitch and radius, respectively. It was found that an interwound configuration describes the data well, provided interactions among supercoils are accounted for in the second virial approximation. The opening angle was observed to be relatively constant and close to 58 degrees, but it was necessary to include a significant distribution in radius and pitch. For diluted supercoils with vanishing mutual interaction, the derived structural results agree with independent measurements, including the distribution in linking number deficit as determined by gel electrophoresis. With increasing plasmid concentration, prior and covering the transition to the liquid-crystalline phase, the radius and pitch are seen to decrease significantly. The latter observation shows that compaction of negatively supercoiled DNA by confinement results in a decrease in writhing number at the cost of a positive twist exerted on the DNA duplex. It is our conjecture that the free energy associated with this excess twist is of paramount importance in controlling the critical boundaries pertaining to the transition to the anisotropic, liquid-crystalline phase. PMID- 12124291 TI - Liquid crystal formation in supercoiled DNA solutions. AB - The critical concentrations pertaining to the liquid crystal formation of pUC18 plasmid in saline solutions were obtained from (31)P nuclear magnetic resonance, polarized light microscopy, and phase equilibrium experiments. The transition is strongly first order with a broad gap between the isotropic and anisotropic phase. The critical boundaries are strongly and reversibly dependent on temperature and weakly dependent on ionic strength. With polarized light microscopy on magnetically oriented samples, the liquid crystalline phase is assigned cholesteric with a pitch on the order of 4 microm. Preliminary results show that at higher concentrations a true crystal is formed. The isotropic cholesteric transition is interpreted with lyotropic liquid crystal theory including the effects of charge, orientation entropy, and excluded volume effects. It was found that the molecular free energy associated with the topology of the superhelix is of paramount importance in controlling the width of the phase gap. The theoretical results compare favorably with the critical boundary pertaining to the disappearance of the isotropic phase, but they fail to predict the low concentration at which the anisotropic phase first appears. PMID- 12124292 TI - Role of Asp193 in chromophore-protein interaction of pharaonis phoborhodopsin (sensory rhodopsin II). AB - Pharaonis phoborhodopsin (ppR; also pharaonis sensory rhodopsin II, psRII) is a receptor of the negative phototaxis of Natronobacterium pharaonis. By spectroscopic titration of D193N and D193E mutants, the pK(a) of the Schiff base was evaluated. Asp193 corresponds to Glu204 of bacteriorhodopsin (bR). The pK(a) of the Schiff base (SBH(+)) of D193N was approximately 10.1-10.0 (at XH(+)) and approximately 11.4-11.6 (at X) depending on the protonation state of a certain residue (designated by X) and independent of Cl(-), whereas those of the wild type and D193E were >12. The pK(a) values of XH(+) were approximately 11.8-11.2 at the state of SB, 10.5 at SBH(+) state in the presence of Cl(-), and 9.6 at SBH(+) without Cl(-). These imply the presence of a long-range interaction in the extracellular channel. Asp193 was suggested to be deprotonated in the present dodecyl-maltoside (DDM) solubilized wild-type ppR, which is contrary to Glu204 of bR. In the absence of salts, the irreversible denaturation of D193N (but not the wild type and D193E) occurred via a metastable state, into which the addition of Cl(-) reversed the intact pigment. This suggests that the negative charge at residue 193, which can be substituted by Cl(-), is necessary to maintain the proper conformation in the DDM-solubilized ppR. PMID- 12124293 TI - Energetics and volume changes of the intermediates in the photolysis of octopus rhodopsin at a physiological temperature. AB - Enthalpy changes (Delta H) of the photointermediates that appear in the photolysis of octopus rhodopsin were measured at physiological temperatures by the laser-induced transient grating method. The enthalpy from the initial state, rhodopsin, to bathorhodopsin, lumirhodopsin, mesorhodopsin, transient acid metarhodopsin, and acid metarhodopsin were 146 +/- 15 kJ/mol, 122 +/- 17 kJ/mol, 38 +/- 8 kJ/mol, 12 +/- 5 kJ/mol, and 12 +/- 5 kJ/mol, respectively. These values, except for lumirhodopsin, are similar to those obtained for the cryogenically trapped intermediate species by direct calorimetric measurements. However, the Delta H of lumirhodopsin at physiological temperatures is quite different from that at low temperature. The reaction volume changes of these processes were determined by the pulsed laser-induced photoacoustic method along with the above Delta H values. Initially, in the transformation between rhodopsin and bathorhodopsin, a large volume expansion of +32 +/- 3 ml/mol was obtained. The volume changes of the subsequent reaction steps were rather small. These results are compared with the structural changes of the chromophore, peptide backbone, and water molecules within the membrane helixes reported previously. PMID- 12124294 TI - Intermolecular interactions, nucleation, and thermodynamics of crystallization of hemoglobin C. AB - The mutated hemoglobin HbC (beta 6 Glu-->Lys), in the oxygenated (R) liganded state, forms crystals inside red blood cells of patients with CC and SC diseases. Static and dynamic light scattering characterization of the interactions between the R-state (CO) HbC, HbA, and HbS molecules in low-ionic-strength solutions showed that electrostatics is unimportant and that the interactions are dominated by the specific binding of solutions' ions to the proteins. Microscopic observations and determinations of the nucleation statistics showed that the crystals of HbC nucleate and grow by the attachment of native molecules from the solution and that concurrent amorphous phases, spherulites, and microfibers are not building blocks for the crystal. Using a novel miniaturized light scintillation technique, we quantified a strong retrograde solubility dependence on temperature. Thermodynamic analyses of HbC crystallization yielded a high positive enthalpy of 155 kJ mol(-1), i.e., the specific interactions favor HbC molecules in the solute state. Then, HbC crystallization is only possible because of the huge entropy gain of 610 J mol(-1) K(-1), likely stemming from the release of up to 10 water molecules per protein intermolecular contact-hydrophobic interaction. Thus, the higher crystallization propensity of R-state HbC is attributable to increased hydrophobicity resulting from the conformational changes that accompany the HbC beta 6 mutation. PMID- 12124295 TI - Effect of the environment on the protein dynamical transition: a neutron scattering study. AB - We performed an elastic neutron scattering investigation of the molecular dynamics of lysozyme solvated in glycerol, at different water contents h (grams of water/grams of lysozyme). The marked non-Gaussian behavior of the elastic intensity was studied in a wide experimental momentum transfer range, as a function of the temperature. The internal dynamics is well described in terms of the double-well jump model. At low temperature, the protein total mean square displacements exhibit an almost linear harmonic trend irrespective of the hydration level, whereas at the temperature T(d) a clear changeover toward an anharmonic regime marks a protein dynamical transition. The decrease of T(d) from approximately 238 K to approximately 195 K as a function of h is reminiscent of that found in the glass transition temperature of aqueous solutions of glycerol, thus suggesting that the protein internal dynamics as a whole is slave to the environment properties. Both T(d) and the total mean square displacements indicate that the protein flexibility strongly rises between 0.1 and 0.2h. This hydration-dependent dynamical activation, which is similar to that of hydrated lysozyme powders, is related to the specific interplay of the protein with the surrounding water and glycerol molecules. PMID- 12124296 TI - Fluorescence lifetime spectroscopy in multiply scattering media with dyes exhibiting multiexponential decay kinetics. AB - To investigate fluorescence lifetime spectroscopy in tissue-like scattering, measurements of phase modulation as a function of modulation frequency were made using two fluorescent dyes exhibiting single exponential decay kinetics in a 2% intralipid solution. To experimentally simulate fluorescence multiexponential decay kinetics, we varied the concentration ratios of the two dyes, 3,3 diethylthiatricarbocyanine iodide and indocynanine green (ICG), which exhibit distinctly different lifetimes of 1.33 and 0.57 ns, respectively. The experimental results were then compared with values predicted using the optical diffusion equation incorporating 1) biexponential decay, 2) average of the biexponential decay, as well as 3) stretched exponential decay kinetic models to describe kinetics owing to independent and quenched relaxation of the two dyes. Our results show that while all kinetic models could describe phase-modulation data in nonscattering solution, when incorporated into the diffusion equation, the kinetic parameters failed to likewise predict phase-modulation data in scattering solutions. We attribute the results to the insensitivity of phase modulation measurements in nonscattering solutions and the inaccuracy of the derived kinetic parameters. Our results suggest the high sensitivity of phase modulation measurements in scattering solutions may provide greater opportunities for fluorescence lifetime spectroscopy. PMID- 12124297 TI - Quaternary structure built from subunits combining NMR and small-angle x-ray scattering data. AB - A new principle in constructing molecular complexes from the known high resolution domain structures joining data from NMR and small-angle x-ray scattering (SAXS) measurements is described. Structure of calmodulin in complex with trifluoperazine was built from N- and C-terminal domains oriented based on residual dipolar couplings measured by NMR in a dilute liquid crystal, and the overall shape of the complex was derived from SAXS data. The residual dipolar coupling data serves to reduce angular degrees of freedom, and the small-angle scattering data serves to confine the translational degrees of freedom. The complex built by this method was found to be consistent with the known crystal structure. The study demonstrates how approximate tertiary structures of modular proteins or quaternary structures composed of subunits can be assembled from high resolution structures of domains or subunits using mutually complementary NMR and SAXS data. PMID- 12124298 TI - Probing the endocytic pathway in live cells using dual-color fluorescence cross correlation analysis. AB - Fluorescence (auto)correlation spectroscopy (FCS) has developed into a widely used method for investigating molecular dynamics and mobility of molecules in vitro and in vivo. Dual-color cross-correlation, an extension of this technique, also assesses the concomitant movement of two spectrally distinguishable fluorescent molecules and has therefore proven superior to autocorrelation analysis to study interactions between different molecular species in solution. Here we explore the benefits of cross-correlation analysis when applied to live cells, by demonstrating its potential in analyzing endocytic processes. Bacterial cholera toxin (CTX) was labeled with Cy2 and Cy5 dyes on different subunits of the same holotoxin. Along the endocytic pathway, positive cross-correlation between the A and B subunits was first preserved, later followed by a loss in cross-correlation upon their separation in the Golgi. Furthermore, endocytosis of a mixture of only Cy2- and only Cy5-labeled holotoxins also gave rise to cross correlation. Our results suggest that cross-correlation may be used to recognize whether different cargoes use the same endocytic pathway. Additionally, we show that cross-correlation is applicable to two-dimensional membrane diffusion. CTX bound to GM1-containing artificial giant unilamellar vesicles was diffusible, whereas CTX bound to the plasma membrane was immobile on the FCS time-scale, possibly because of raft-association of GM1. PMID- 12124299 TI - Supramolecular properties of the proline-rich gamma-Zein N-terminal domain. AB - Zeins are maize storages proteins that accumulate inside large vesicles called protein bodies. gamma-Zein lines the inner face of the protein body membrane, and its N-terminal proline-rich repetitive domain with the sequence (VHLPPP)(8) appears to be necessary for the accumulation of the protein within the organelle. Synthetic (VHLPPP)(8) adopts an amphipathic polyproline II conformation. In a preliminary recent work we used atomic force microscopy to study the surface organization of the octamer and transmission electron microscopy to visualize aggregates of the peptide from aqueous solution. We previously envisioned two self-assembly models (i.e., the geometric and the micellar) that take into account the observed features. In the present work we studied in detail the self assembly of the peptide in solution and found that the peptide is able to form cylindrical micelles. Fibrils formed on graphite are generated by assembly of solution micelles. Based on the results of these studies, we focused exclusively on the micellar model. To our knowledge we have characterized for the first time supramolecular aggregates of polyproline structures other than collagen. The spontaneous arrangement of (VHLPPP)(8) suggests a role for the N-terminal domain of gamma-zein in the process of the whole protein deposition in protein bodies. PMID- 12124300 TI - Supramolecular structure in full-length Alzheimer's beta-amyloid fibrils: evidence for a parallel beta-sheet organization from solid-state nuclear magnetic resonance. AB - We report constraints on the supramolecular structure of amyloid fibrils formed by the 40-residue beta-amyloid peptide associated with Alzheimer's disease (A beta(1-40)) obtained from solid-state nuclear magnetic resonance (NMR) measurements of intermolecular dipole-dipole couplings between (13)C labels at 11 carbon sites in residues 2 through 39. The measurements are carried out under magic-angle spinning conditions, using the constant-time finite-pulse radiofrequency-driven recoupling (fpRFDR-CT) technique. We also present one dimensional (13)C magic-angle spinning NMR spectra of the labeled A beta(1-40) samples. The fpRFDR-CT data reveal nearest-neighbor intermolecular distances of 4.8 +/- 0.5 A for carbon sites from residues 12 through 39, indicating a parallel alignment of neighboring peptide chains in the predominantly beta-sheet structure of the amyloid fibrils. The one-dimensional NMR spectra indicate structural order at these sites. The fpRFDR-CT data and NMR spectra also indicate structural disorder in the N-terminal segment of A beta(1-40), including the first nine residues. These results place strong constraints on any molecular-level structural model for full-length beta-amyloid fibrils. PMID- 12124301 TI - Model-based analysis of assembly kinetics for virus capsids or other spherical polymers. AB - The assembly of virus capsids or other spherical polymers--empty, closed structures composed of hundreds of protein subunits--is poorly understood. Assembly of a closed spherical polymer is unlike polymerization of a filament or crystal, examples of open-ended polymers. This must be considered to develop physically meaningful analyses. We have developed a model of capsid assembly, based on a cascade of low-order reactions, that allows us to calculate kinetic simulations. The behavior of this model resembles assembly kinetics observed in solution (Zlotnick, A., J. M. Johnson, P. W. Wingfield, S. J. Stahl, and D. Endres. 1999. Biochemistry. 38:14644-14652). We exhibit two examples of this general model describing assembly of dodecahedral and icosahedral capsids. Using simulations based on these examples, we demonstrate how to extract robust estimates of assembly parameters from accessible experimental data. These parameters, nucleus size, average nucleation rate, and average free energy of association can be determined from measurement of subunit and capsid as time and concentration vary. Mathematical derivations of the analyses, carried out for a general model, are provided in an Appendix. The understanding of capsid assembly developed in this paper is general; the examples provided can be readily modified to reflect different biological systems. This enhanced understanding of virus assembly will allow a more quantitative analysis of virus stability and biological or antiviral factors that affect assembly. PMID- 12124302 TI - Pharmacokinetic-pharmacodynamic analysis of the glucose-lowering effect of metformin in diabetic rats reveals first-pass pharmacodynamic effect. AB - Metformin, a commonly used antidiabetic drug, exerts its glucose-lowering effect due to metabolic activities at several sites of action (biophases), including liver, intestine, muscle cells, and adipocytes. The relative contribution of the individual biophases to the overall glucose-lowering effect is not known. Thus, the aims of this investigation were to study the influence of mode of drug administration on the kinetics of glucose-lowering action of metformin in diabetic rats and identify the contribution of different sites of action to the overall response. Streptozotocin diabetic rats received metformin in crossover fashion via intraduodenal, intravenous, and intraportal routes as bolus dose or infusion regimens designed to yield similar pharmacokinetic profiles. Metformin plasma concentrations and blood glucose levels were measured following each mode of administration. Despite the similarity in the concentration-time profiles obtained for different routes of metformin administration, intraduodenal administration produced larger response than intraportal metformin infusion, and lowest response was observed following intravenous administration. This finding indicates that a significant "first-pass" pharmacodynamic effect, which occurs in the presystemic sites of action (liver and the gastrointestinal wall), contributes to the overall glucose-lowering response of metformin. We applied a combined pharmacokinetic-pharmacodynamic modeling approach to study the nature of the first-pass pharmacodynamic effect. The observed data were successfully described by a novel integrated indirect response pharmacokinetic-pharmacodynamic model that revealed a correlation between the temporal metformin concentrations that transit the portal vein and through the gut wall rather than with drug concentrations that accumulated in the liver and the intestinal wall. PMID- 12124303 TI - Metabolism of tamoxifen by recombinant human cytochrome P450 enzymes: formation of the 4-hydroxy, 4'-hydroxy and N-desmethyl metabolites and isomerization of trans-4-hydroxytamoxifen. AB - The cytochrome P450 (P450)-mediated biotransformation of tamoxifen is important in determining both the clearance of the drug and its conversion to the active metabolite, trans-4-hydroxytamoxifen. Biotransformation by P450 forms expressed extrahepatically, such as in the breast and endometrium, may be particularly important in determining tissue-specific effects of tamoxifen. Moreover, tamoxifen may serve as a useful probe drug to examine the regioselectivity of different forms. Tamoxifen metabolism was investigated in vitro using recombinant human P450s. Forms CYP1A1, 1A2, 1B1, 2A6, 2B6, 2C9, 2C19, 2D6, 2E1, 3A4, 3A5, and 3A7 were coexpressed in Escherichia coli with recombinant human NADPH-cytochrome P450 reductase. Bacterial membranes were harvested and incubated with tamoxifen or trans-4-hydroxytamoxifen under conditions supporting P450-mediated catalysis. CYP2D6 was the major catalyst of 4-hydroxylation at low tamoxifen concentrations (170 +/- 20 pmol/40 min/0.2 nmol P450 using 18 microM tamoxifen), but CYP2B6 showed significant activity at high substrate concentrations (28.1 +/- 0.8 and 3.1 +/- 0.5 nmol/120 min/0.2 nmol P450 for CYP2D6 and CYP2B6, respectively, using 250 microM tamoxifen). These two forms also catalyzed 4'-hydroxylation (13.0 +/- 1.9 and 1.4 +/- 0.1 nmol/120 min/0.2 nmol P450, respectively, for CYP2B6 and CYP2D6 at 250 microM tamoxifen; 0.51 +/- 0.08 pmol/40 min/0.2 nmol P450 for CYP2B6 at 18 microM tamoxifen). Tamoxifen N-demethylation was mediated by CYP2D6, 1A1, 1A2, and 3A4, at low substrate concentrations, with contributions by CYP1B1, 2C9, 2C19 and 3A5 at high concentrations. CYP1B1 was the principal catalyst of 4 hydroxytamoxifen trans-cis isomerization but CYP2B6 and CYP2C19 also contributed. PMID- 12124304 TI - Kinetic analysis of the reactions of 4-hydroperoxycyclophosphamide and acrolein with glutathione, mesna, and WR-1065. AB - The kinetics of the reactions of glutathione (GSH) with 4 hydroperoxycyclophosphamide (4OOH-CP) and acrolein, a metabolite of 4OOH-CP, were investigated in a cell-free medium (pH approximately 7.5) and peripheral blood mononuclear cells. The ability of the thiol drugs, sodium 2-mercaptoethane sulfonate (mesna) and S-2-(3-aminopropylamino)ethanethiol (WR-1065), to affect the reactions of cellular GSH with the alkyalting agents was also studied. The amount of unreacted thiols in the various reactions was determined by derivatization with monobromobimane, followed by separation of fluorescent labeled thioether adducts using high-pressure liquid chromatography. The second order rate constants (k(2)) for reactions of GSH, mesna, and WR-1065 with 4OOH-CP in solution were 38 +/- 5, 25 +/- 5, and 880 +/- 50 M(-1) s(-1), respectively. The corresponding k(2) for reactions of GSH, mesna, and WR-1065 with acrolein were 490 +/- 100, 700 +/- 150, and >2000 M(-1) s(-1), respectively. The apparent rate constants for reactions of cellular GSH with acrolein and 4OOH-CP were smaller than those obtained in solution. Assuming that the k(2) is the same inside and outside cells, we estimate the first-order rate constant (k(1)) for transfer of 4OOH-CP and acrolein across the cell membrane as approximately 0.01 and approximately 0.04 s(-1), respectively. WR-1065 was more effective than mesna in blocking depletion of cellular GSH (because it passes into the cell more quickly and has higher reaction rates with the alkylators than the latter compound). When WR-1065 and mesna were used together, the protection against cellular depletion of GSH was additive. Our results are relevant to the administration of thiol drugs with high-dose alkylating agents. PMID- 12124305 TI - Comparative metabolic capabilities of CYP3A4, CYP3A5, and CYP3A7. AB - The human cytochromes P450 (P450) CYP3A contribute to the biotransformation of 50% of oxidatively metabolized drugs. The predominant hepatic form is CYP3A4, but recent evidence indicates that CYP3A5 contributes more significantly to the total liver CYP3A than was originally thought. CYP3A7 is the major fetal form and is rarely expressed in adults. To compare the metabolic capabilities of CYP3A forms for 10 substrates, incubations were performed using a consistent molar ratio (1:7:9) of recombinant CYP3A, P450 reductase, and cytochrome b5. A wide range of substrate concentrations was examined to determine the best fit to kinetic models for metabolite formation. In general, K(m) or S(50) values for the substrates were 3 to 4 times lower for CYP3A4 than for CYP3A5 or CYP3A7. For a more direct comparison of these P450 forms, clearance to the metabolites was determined as a linear relationship of rate of metabolite formation for the lowest substrate concentrations examined. The clearance for 1'-hydroxy midazolam formation at low substrate concentrations was similar for CYP3A4 and CYP3A5. For CYP3A5 versus CYP3A4, clearance values at low substrate concentrations were 2 to 20 times lower for the other biotransformations. The clearance values for CYP3A7-catalyzed metabolite formation at low substrate concentrations were substantially lower than for CYP3A4 or CYP3A5, except for clarithromycin, 4-OH triazolam, and N desmethyl diltiazem (CYP3A5 - CYP3A7). The CYP3A forms demonstrated regioselective differences in some of the biotransformations. These results demonstrate an equal or reduced metabolic capability for CYP3A5 compared with CYP3A4 and a significantly lower capability for CYP3A7. PMID- 12124306 TI - Prediction of hepatic clearance and availability by cryopreserved human hepatocytes: an application of serum incubation method. AB - A novel and convenient method was established for the prediction of in vivo metabolic clearance in human liver. The present method applied the in vitro-in vivo extrapolation paradigm previously established in rats to the in vitro data obtained from cryopreserved human hepatocytes. Predicted hepatic availability and clearance were compared with the reported oral bioavailability and plasma clearance in humans for 14 clinically used drugs (naloxone, buspirone, verapamil, lidocaine, imipramine, metoprolol, timolol, antipyrine, diazepam, quinidine, caffeine, propranolol, diclofenac, and phenacetin). A large interindividual variation was observed in the intrinsic metabolic clearance among separate cryopreserved preparations from different subjects. The prediction generally resulted in a marked underestimation when the biologically based scaling factor (3.1 x 10(9) cells/kg) was used for the extrapolation of in vitro data (milliliters per minutes per cells) to in vivo value (milliliters per minutes per kilograms). Reasonably good in vitro-in vivo correlations were obtained with empirically calculated scaling factors, 8.5 x 10(9) (cells/kg) from 10 individual preparations and 10.8 x 10(9) (cells/kg) from pooled preparation of two selected lots, which were 3- to 4-fold larger than the biologically based scaling factor. These data suggested that the calibration of inherent interindividual variation of metabolic activities among different cryopreserved preparations of human hepatocytes to obtain the empirical scaling factor, which is applicable only to the preparation used, was an essential step for more reliable and quantitative prediction of in vivo metabolic activity in humans. PMID- 12124307 TI - Glucuronidation and sulfation of the tea flavonoid (-)-epicatechin by the human and rat enzymes. AB - (-)-Epicatechin (EC) is one of the flavonoids present in green tea, suggested to have chemopreventive properties in cancer. However, its bioavailability is not clearly understood. In the present study, we determined the metabolism of EC, focusing on its glucuronic acid and sulfate conjugation using human liver and intestinal microsomes and cytosol as well as recombinant UDP glucuronosyltransferase (UGT) and sulfotransferase (SULT) isoforms in comparison with that occurring in the rat. Surprisingly, EC was not glucuronidated by the human liver and small intestinal microsomes. There was also no evidence of glucuronidation by human colon microsomes or by recombinant UGT1A7, which is not present in the liver or intestine. Interestingly, in the rat liver microsomes EC was efficiently glucuronidated with the formation of two glucuronides. In contrast, the human liver cytosol efficiently sulfated EC mainly through the SULT1A1 isoform. For the intestine, both SULT1A1 and SULT1A3 contributed. Other SULT isoforms contributed little. High-performance liquid chromatography of the sulfate conjugates showed one major sulfatase-sensitive peak with all tissues. An additional minor sulfatase-resistant peak was formed by the liver and intestinal cytosol as well as with SULT1A1 but not by the Caco-2 cytosol and SULT1A3. In the rat, EC sulfation was considerably less efficient than in the human liver. These results indicate that sulfation is the major pathway in EC metabolism in the human liver and intestine with no glucuronidation occurring. There was also a large species difference both in glucuronidation and sulfation of EC between rats and humans. PMID- 12124308 TI - Sex difference in inhibition of in vitro mexazolam metabolism by various 3 hydroxy-3-methylglutaryl-coenzyme a reductase inhibitors in rat liver microsomes. AB - To identify an appropriate animal model for the study of drug interaction via CYP3A4 inhibition, the inhibition of in vitro mexazolam metabolism by various 3 hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase inhibitors [simvastatin (lactone), simvastatin acid, fluvastatin, atorvastatin, cerivastatin, pravastatin lactone, and pravastatin (acid)] in male and female rat liver microsomes was investigated and compared with that by HMG-CoA reductase inhibitors in human liver microsomes reported previously. The metabolism of mexazolam in male and female rat liver microsomes was inhibited by all the HMG-CoA reductase inhibitors examined except pravastatin (acid). The K(i) values in female rats were lower than those in male rats, demonstrating the presence of a sex difference in the inhibition potency of HMG-CoA reductase inhibitors toward mexazolam. Using anti cytochrome P450 (P450) antisera, the main P450 isozyme responsible for the metabolism of mexazolam was identified as CYP3A in female rats and CYP2C11 in male rats. Based on these results, we speculate that the sex difference in the inhibition potency of HMG-CoA reductase inhibitors for mexazolam observed in rats is caused by their different inhibition potencies against CYP2C11 and CYP3A isoforms. For mexazolam metabolism, the results obtained in female rats, rather than those in male rats, seem to be a much better reflection of the results in humans. Since species and sex differences were observed in P450 isozymes in the present study, our results show that establishing appropriate experimental conditions, in particular with respect to the P450 isozymes responsible for the drug metabolism in question, is indispensable for the investigation of drug interactions using rats as a model animal for humans. PMID- 12124309 TI - Cytochrome P450 2E1 (CYP2E1) is essential for acrylonitrile metabolism to cyanide: comparative studies using CYP2E1-null and wild-type mice. AB - Acrylonitrile (AN) is a rodent carcinogen and suspected human carcinogen. Metabolism of AN proceeds via conjugation with glutathione or epoxidation via cytochrome P4502E1 (CYP2E1) to cyanoethylene oxide (CEO). It was hypothesized that CEO metabolism via epoxide hydrolase (EH) is the primary pathway for cyanide formation. The objective of this work is to assess the enzymatic basis of metabolism to cyanide. Male wild-type and CYP2E1-null mice received 0, 2.5, 10, 20, or 40 mg of AN/kg by gavage, and cyanide was measured in blood and tissues. CYP2E1 and EH expression were assessed using Western blot analyses. Present results demonstrated that cyanide concentrations in blood and tissues of AN treated wild-type mice were higher at 1 versus 3 h, increased in a dose-dependent manner, and were significantly higher in AN-treated versus vehicle-treated mice. In contrast, cyanide concentrations in the blood and tissues of AN-treated CYP2E1 null mice were not statistically different from those of vehicle-treated mice. Furthermore, this work showed that EH is expressed in CYP2E1-null and wild-type mice. In conclusion, under the current experimental conditions using CYP2E1-null mice, current work demonstrated for the first time that CYP2E1-mediated oxidation is a prerequisite for AN metabolism to cyanide. Since earlier studies showed that CYP2E1 is the only enzyme responsible for AN epoxidation, it is concluded that AN metabolism to CEO is a prerequisite for cyanide formation, and this pathway is exclusively catalyzed by CYP2E1. Finally, this work confirmed that cyanide plays an essential role in the causation of the acute toxicity/mortality of AN. PMID- 12124310 TI - Role of constitutive androstane receptor in the in vivo induction of Mrp3 and CYP2B1/2 by phenobarbital. AB - Phenobarbital (PB) induces the hepatic organic anion transporter, Mrp3. The present study tested the hypothesis that Mrp3 induction by PB is mediated by the constitutive androstane receptor (CAR). PB induction of Mrp3 and CYP2B was examined in lean and obese Zucker rats, male and female Wistar Kyoto (WKY) rats, HepG2 and mouse CAR-expressing HepG2 (g2car-3) cells; HepG2 and g2car-3 cells also were treated with 1,4-bis[2-(3,5-dichloropyridyloxy)]benzene (TCPOBOP). In obese Zucker rat livers, total and nuclear CAR levels were markedly lower compared with lean rat livers, which correlated with the poor induction of CYP2B1/2 by PB in obese Zucker rats. Mrp3 induction by PB also was impaired in obese Zucker rat livers. Induction of Mrp3 by PB was similar in male and female WKY rat livers, despite the fact that CAR protein levels were significantly lower in female relative to male WKY rat livers. MRP3 levels in both HepG2 and g2car-3 cells were induced to a similar extent in the two cell lines by PB but not by TCPOBOP. In contrast, CYP2B6 levels were measurable and induced by TCPOBOP only in g2car-3 cells. In conclusion, data from WKY rats and HepG2 cells suggest that CAR does not play a key role in PB induction of Mrp3. Impaired induction of Mrp3 by PB in obese Zucker rats is not due solely to CAR deficiency. Interestingly, differences in the constitutive levels of Mrp3 were observed between obese and lean Zucker rats and between male and female WKY rats. PMID- 12124311 TI - Intestinal absorption enhancement of the ester prodrug tenofovir disoproxil fumarate through modulation of the biochemical barrier by defined ester mixtures. AB - The effect of discrete esters and ester mixtures on the intestinal stability and absorption of tenofovir disoproxil fumarate (tenofovir DF, an esterase-sensitive prodrug of the antiviral tenofovir) was compared with the effect of strawberry extract, which has been shown to enhance the absorption of the prodrug across Caco-2 monolayers and in rat ileum. In addition, the mechanism of absorption enhancement was investigated. In rat intestinal homogenates, complete inhibition of the conversion of tenofovir DF (as obtained by strawberry extract) could only be obtained at relatively high concentrations of the discrete esters or by using mixtures of esters (e.g., propyl p-hydroxybenzoate 0.02%, octyl acetate 0.02%, ethyl caprylate 0.01%). Coincubation of tenofovir DF with this mixture also resulted in an enhancement of its absorption in the in vitro Caco-2 system as well as in rat ileum. As tenofovir DF is a substrate for P-glycoprotein (P-gp) related efflux carriers in the Caco-2 model, the modulatory effect of the ester mixtures was studied on the functionality of P-gp using cyclosporin A (CsA) as a model substrate. Strawberry extract as well as the mixture of three esters interfered with the absorptive transport of CsA across Caco-2 monolayers, illustrating that both mixtures interfere with both esterase-activity and P-gp functionality. This concerted barrier was not observed in rat ileum, suggesting differential functional activities of the biochemical barrier toward tenofovir DF in different absorption systems. Overall, our results illustrate that modulation of the biochemical barrier (metabolism and efflux) of tenofovir DF by ester mixtures can be used to increase the intestinal absorption of tenofovir DF in an in vitro and an in situ absorption model; the mechanism of action appears to be a complex interplay of different systems; the differential expression of carriers and enzymes in different systems illustrates the difficulty of extrapolating observations between different systems/species. PMID- 12124312 TI - Amino acid residues affecting the activities of human cytochrome P450 2C9 and 2C19. AB - The amino acid residues affecting the substrate specificity of human cytochrome P450 CYP2C9 and CYP2C19 for their metabolic activities were investigated using chimeras and mutant enzymes, which were constructed by replacing the corresponding residues. Although CYP2C19 showed nearly the same tolbutamide hydroxylase activity as CYP2C9, the activities for the CYP2C19(H99I) mutant and the chimeras that replaced residues 1-212 were much lower than those for CYP2C19. The activities of the CYP2C19(H99I) mutant and the chimeras that replaced residues 228-340 were lower than those for CYP2C19 toward S-mephenytoin, aminopyrine, and testosterone. These results suggest that residues in substrate recognition site (SRS) 3 and 4 are important for the substrate specificity, whereas His99 is important in the substrate binding of CYP2C19. For the 4' hydroxylation of diclofenac, CYP2C9 had a lower K(m) and a higher V(max) than CYP2C19. Although the V(max) values for the CYP2C9(1-288)/CYP2C19(289-490) chimera and the CYP2C9(I99H, V292A, F295L, I331V) mutant were comparable to those for CYP2C9, its K(m) value was comparable to that for CYP2C19. The V(max) and K(m) values for the CYP2C19(1-288)/CYP2C9(289-490) chimera were comparable to those for CYP2C19, and the activity by CYP2C9(1-230)/CYP2C19(231-490) chimera was negligible. These results suggest that the residues 292, 295, and/or 331 of CYP2C9 are essential for the recognition of substrate in CYP2C9 and that the residues of 231-288 including SRS 3 are important for the metabolizing capacity of CYP2C9. PMID- 12124313 TI - Identification of metabolites of a substance P (neurokinin 1 receptor) antagonist in rat hepatocytes and rat plasma. AB - [3R,5R,6S]-3-(2-cyclopropyloxy-5-trifluoromethoxyphenyl)-6-phenyl-1-oxa-7 azaspiro[4.5]decane is a substance P (Neurokinin 1 receptor) antagonist. Substance P antagonists are proven in concept to have excellent potential for the treatment of major depression, and they allow superior and sustained protection from acute and delayed chemotherapy-induced emesis. The metabolism of this compound was investigated in rat hepatocytes, and circulating rat plasma metabolites were identified following oral and intravenous dosing. The turnover in rat hepatocytes within 4 h was about 30%, and the major metabolites were identified as two nitrones and a lactam associated with the piperidine ring. Although these metabolites were also observed in rat plasma, the major circulating metabolite was a keto acid following oxidative de-amination of the piperidine ring. Liquid chromatography/tandem mass spectrometry and nuclear magnetic resonance were used to confirm the structure of the latter metabolite. A mechanism leading to the formation of the keto acid metabolite has been suggested, and most intermediates were observed in rat plasma. PMID- 12124314 TI - Phenol sulfotransferase, ST1A3, as the main enzyme catalyzing sulfation of troglitazone in human liver. AB - Since sulfation is the main metabolic pathway of troglitazone, accounting for about 70% of the metabolites detected in human plasma, we have aimed to identify human cytosolic sulfotransferases catalyzing the sulfation of troglitazone and to examine a possible role of the sulfation in the cytotoxicity observed in cell lines of human origin (HepG2 and Hep3B). Experiments using the recombinant sulfotransferases and human liver cytosols indicated that phenol sulfotransferase (ST1A3) and estrogen sulfotransferase (ST1E4) were the sulfotransferases most active toward troglitazone. Immunoblot analyses indicated that hepatic content of ST1A3 is about 13 times higher than that of ST1E4, suggesting that ST1A3 is mainly responsible for the sulfation of troglitazone in the liver. Lactate dehydrogenase (LDH) leakage was elicited by troglitazone in a concentration dependent manner in the hepatoma cells. The troglitazone metabolites (the sulfate, glucuronide, and quinone forms) caused negligible LDH leakage. These findings suggest that accumulation of unmetabolized troglitazone causes the cytotoxicity in the hepatoma cells and may be responsible for toxicity in human liver. PMID- 12124315 TI - Mechanisms of action of arsenic trioxide. AB - Arsenic trioxide has shown substantial efficacy in treating both newly diagnosed and relapsed patients with acute promyelocytic leukemia (APL). As a single agent, it induces complete remissions, causing few adverse effects and only minimal myelosuppression. These successes have prompted investigations to elucidate the mechanisms of action underlying these clinical responses. Substantial data show that arsenic trioxide produces remissions in patients with APL at least in part through a mechanism that results in the degradation of the aberrant PML-retinoic acid receptor alpha fusion protein. Studies have also investigated concerns about the toxicity and potential carcinogenicity of long-term exposure to environmental arsenic. Arsenic apparently affects numerous intracellular signal transduction pathways and causes many alterations in cellular function. These actions of arsenic may result in the induction of apoptosis, the inhibition of growth and angiogenesis, and the promotion of differentiation. Such effects have been observed in cultured cell lines and animal models, as well as clinical studies. Because arsenic affects so many cellular and physiological pathways, a wide variety of malignancies, including both hematologic cancer and solid tumors derived from several tissue types, may be susceptible to therapy with arsenic trioxide. These multiple actions of arsenic trioxide also highlight the need for additional mechanistic studies to determine which actions mediate the diverse biological effects of this agent. This information will be critical to realizing the potential for synergy between arsenic trioxide and other chemotherapeutic agents, thus providing enhanced benefit in cancer therapy. PMID- 12124316 TI - TEL-AML1, expressed from t(12;21) in human acute lymphocytic leukemia, induces acute leukemia in mice. AB - TEL-AML1 is expressed from the t(12;21) chromosomal translocation inB-precursor acute lymphocytic leukemia (ALL). Creation of the TEL-AML1fusion disrupts one copy of the TEL and AML1 genes, and loss of TEL or AML1 is also associated with cases of acute leukemia without TEL-AML1. To determine whether TEL-AML1 can contribute to leukemogenesis, we transduced marrow from C57BL/6 mice with a retroviral vector expressing TEL-AML1 or with a control vector. Transduced cells were introduced into irradiated syngeneic recipients. Two of 9 TEL-AML1 mice developed ALL (one T-lineage ALL and one B-precursor ALL), whereas 0 of 20 control mice developed leukemia. The B-precursor ALL was retransplantable and expressed TEL-AML1. We similarly transduced marrow from C57BL/6 mice lacking the overlapping p16(INK4a)p19(ARF) genes and transplanted the cells into wild-type recipients. No control mice died, but six of eight TEL-AML1/p16p19 mice died with leukemia. Overall, these findings indicate that TEL-AML1 contributes to leukemogenesis and may cooperate with loss of p16(INK4a)p14(ARF) to transform lymphoid progenitors. PMID- 12124317 TI - Novel therapy for liver cancer: direct intraarterial injection of a potent inhibitor of ATP production. AB - Most types of cancer are difficult to eradicate and some, like liver carcinomas, are almost always fatal. Significantly, we report here that direct intraarterial delivery of 3-bromopyruvate (3-BrPA), a potent inhibitorof cell ATP production, to liver-implanted rabbit tumors, inflicts a rapid, lethal blow to most cancer cells therein. Moreover, systemic delivery of 3-BrPA suppresses "metastatic" tumors that arise in the lungs. In both cases, there is no apparent harm to other organs or to the animals. Thus, intraarterial delivery of agents like 3-BrPA directly to the site of the primary tumor, followed by systemic delivery only when necessary, may represent a powerful new strategy for arresting the growth of liver and other cancers while minimizing toxic side effects. PMID- 12124318 TI - Development of antitumor immune responses in reconstituted lymphopenic hosts. AB - When naive T cells reconstitute lymphopenic hosts, they transiently proliferate and differentiate into memory-like T cells. Here we report that tumor-specific T cells preferentially expand in tumor vaccine-draining lymph nodes after a melanoma vaccine given to RAG1 mice reconstituted with naive T cells from normal mice. The percentage of tumor-specific Tc1 T cells detected by intracellular cytokine staining was increased approximately 4-fold in reconstituted lymphopenic hosts compared with normal hosts. Concomitantly, vaccination-induced Th1 T cells were also dramatically increased in vaccinated, reconstituted RAG1 hosts. T cells derived from reconstituted RAG1 hosts exhibited a higher level of melanoma specific cytotoxicity in vitro. These cells were significantly more potent at mediating tumor regression in vivo after adoptive transfer into mice bearing established pulmonary metastases. Vaccination is best performed concomitantly with reconstitution; delayed vaccination resulted in T cells with less therapeutic activity. PMID- 12124319 TI - Molecular effects of the herbal compound PC-SPES: identification of activity pathways in prostate carcinoma. AB - Clinical trials of the herbal preparation PC-SPES have demonstrated substantial responses in patients with advanced prostate cancer. Biochemical assays and clinical observations suggest that the effects of PC-SPES are mediated at least in part through estrogenic activity, although the mechanism(s) remains largely undefined. In this study, we used cDNA microarray analysis to identify gene expression changes in LNCaP prostate carcinoma cells exposed to PC-SPES and estrogenic agents including diethylstilbestrol. PC-SPES altered the expression of 156 genes after 24 h of exposure. Of particular interest, transcripts encoding cell cycle-regulatory proteins, alpha- and beta-tubulins, and the androgen receptor were down-regulated by PC-SPES. A comparison of gene expression profiles resulting from these treatments indicates that PC-SPES exhibits activities distinct from those attributable to diethylstilbestrol and suggests that alterations in specific genes involved in modulating the cell cycle, cell structure, and androgen response may be responsible for PC-SPES-mediated cytotoxicity. PMID- 12124320 TI - A hereditary nonpolyposis colorectal carcinoma case associated with hypermethylation of the MLH1 gene in normal tissue and loss of heterozygosity of the unmethylated allele in the resulting microsatellite instability-high tumor. AB - Fourteen suspected hereditary nonpolyposis colorectal carcinoma cases with microsatellite unstable(microsatellite instability-high; MSI-H) tumors but no germ-line MSH2, MSH6, or MLH1 mutations were examined for hypermethylation of CpG sites in the critical promoter region of MLH1. The methylation patterns were determined using methylation-specific PCR and by sequence analysis of sodium bisulfite-treated genomic DNA. In one case, DNA hypermethylation of one allele was detected in DNA isolated from blood. In the MSI-H tumor from this case, the unmethylated MLH1 allele was eliminated by loss of heterozygosity, and the methylated allele was retained. This biallelic inactivation resulted in loss of expression of MLH1 in the tumor as confirmed by immunohistochemistry. These results suggest a novel mode of germ-line inactivation of a cancer susceptibility gene. PMID- 12124321 TI - SHPS-1: a budding molecule against cancer dissemination. AB - The expression of SHPS-1 is down-regulated by several oncogene products such as v Src in fibroblasts. In addition, the down-regulation of SHPS-1 is also found inhuman breast cancer tissues compared with the matched normal tissues. On the other hand, forced expression of SHPS-1 suppresses anchorage-independent cell growth of v-Src-transformed cells in vitro as well as peritoneal dissemination of the cells in nude mice. Surprisingly, the extracellular region (EC) of SHPS-1 seems to take part on the inhibitory effect. Because the EC domain interacts with fibronectin and DTT abrogates the inhibitory effects of the EC, we speculate that super-fibronectin may function with the EC as a suppressor of cancer cells. Here we show that SHPS-1 expression provides a unique potential that links suppression of anchorage-independent cell growth and cancer dissemination. PMID- 12124322 TI - Adenovirus-mediated endostatin delivery results in inhibition of mammary gland tumor growth in C3(1)/SV40 T-antigen transgenic mice. AB - We demonstrate the efficacy of systemic administration of a replication-defective adenovirus expressing endostatin (Ad-mEndo) administered during the preinvasive stage of mammary tumor development in C3(1)/T antigen transgenic mice. Mean serum levels of endostatin increased about 8-fold above that of controls and resulted in a significant decrease in tumor growth and an increase in survival. The inhibitory effect of endostatin occurred during or after the progression to invasive carcinoma. Reduced levels of vascular endothelial growth factor mRNA were found in association with high levels of endostatin. Our results demonstrate that the adenoviral induction of high levels of circulating endostatin significantly inhibits mammary tumor growth during the period when the "angiogenic switch" occurs. PMID- 12124323 TI - Comparison of gene expression profiles between hepatitis B virus- and hepatitis C virus-infected hepatocellular carcinoma by oligonucleotide microarray data on the basis of a supervised learning method. AB - Gene expression profiles of hepatocellular carcinomas (HCCs) associated with hepatitis B virus (HBV) and hepatitis C virus (HCV) were analyzed and compared. Oligonucleotide microarrays containing >6000 genes and subsequent gene selection by a supervised learning method yielded 83 genes for which expression differed between the two types of HCCs. Expression levels of 31 of these 83 genes were increased in HBV-associated HCCs, and expression levels of the remaining 52 genes were increased in HCV-associated HCCs. The 31 genes up-regulated in HBV associated HCC included imprinted genes (H19 and IGF2) and genes relating to signal transduction, transcription, and metastasis. The 52 genes up-regulated in HCV-associated HCC included a number of genes responsible for detoxification and immune response. These results suggest that HBV and HCV cause hepatocarcinogenesis by different mechanisms and provide novel tools for diagnosis and treatment of HBV- and HCV-associated HCCs. PMID- 12124324 TI - Loss of retinoic acid receptor beta gene expression is linked to aberrant histone H3 acetylation in lung cancer cell lines. AB - Most lung cancer cell lines do not express retinoic acid receptor (RAR)-beta in response to all-trans retinoic acid (RA) because of a defect in RARbeta gene transcription(RA-refractory cells). Here we investigated mechanisms of RA refractoriness in 14 lung cancer cell lines. Eleven cell lines were found to be RA refractory, and in the other three cell lines, RARbeta levels increased with RA treatment (RA-responsive cells). We observed RARbeta promoter methylation in 7 of 11 RA-refractory cell lines (64%) and in 0 of the 3 RA-responsive cell lines. Treatment with 5-aza-2'-deoxycytidine restored RA response in two of the seven cell lines with RARbeta promoter methylation (29%). RA treatment increased acetylation of histones H3 and H4 on chromatin of the RARbeta promoter in RA responsive cells. Only histone H4 acetylation increased in RA-refractory cells, including refractory cells with and without evidence of promoter methylation. Thus, loss of histone H3 acetylation consistently correlated with RA refractoriness in lung cancer cell lines. RA refractoriness and aberrant histone acetylation were attributable to RARbeta promoter methylation in some cell lines but not in others, suggesting that multiple mechanisms contribute to this transcriptional defect in lung cancer cells. PMID- 12124325 TI - Drg1, a novel target for modulating sensitivity to CPT-11 in colon cancer cells. AB - Treatment of the human colon cancer cells Hct116 with SN-38 (an active metabolite of CPT-11) resulted in G2 cell cycle arrest without induction of apoptosis. However, subsequent treatment of SN-38-treated Hct116 cells with flavopiridol induced apoptosis. One of the genes markedly up-regulated during cell cycle arrest by SN-38 and suppressed during apoptosis by SN-38 followed by flavopiridol in Hct116 cells is Drg1. We found that Drg1 had profound effects on SN-38 sensitivity. Inhibition of endogenous Drg1 expression in Hct116 cells by stable expression of an antisense (AS) Drg1 cDNA increased the sensitivity of cells to undergo apoptosis by SN-38. Clonogenic and apoptosis assays with AS Drg1 expressing cells showed a 2-fold decrease in the IC50 and a 4-5-fold increase in induction of apoptosis with SN-38. Conversely, the forced expression of Drg1 in SW620 cells increased the resistance of these cells to SN-38-induced apoptosis by 2-5-fold. Moreover, when xenografted in mice, AS Drg1-expressing Hct116 cells were 3-fold more sensitive to CPT-11 as compared with vector transfected Hct116 cells. Similarly, tumors established from Drg1 overexpressing SW620 cells were more resistant to CPT-11 as compared with tumors established from vector transfected SW620 cells in mice. Taken together, our data suggest that Drg1 is a novel gene that plays a direct role in resistance to CPT-11. Inhibition of Drg1 may provide a new means to increase the sensitivity of colon cancer cells to CPT 11. PMID- 12124326 TI - Cyclooxygenase-2 inhibitor treatment enhances photodynamic therapy-mediated tumor response. AB - Photodynamic therapy (PDT) continues to be used in the treatment of solid tumors. Clinical results are promising, but the therapy has not been optimized, and tumor recurrences can occur. Recently, it has been shown that inhibitors of cyclooxygenase (COX)-2 can be effective in combination with conventional chemotherapy and radiation therapy. In the current study, we examined the parameters of PDT-mediated activation of COX-2 expression. We also examined the tumoricidal effectiveness of combining PDT with the selective COX-2 inhibitor NS 398. PDT induced the transcriptional activation of COX-2. Prolonged expression of COX-2 protein was observed in PDT-treated mouse sarcoma and carcinoma cell lines, whereas COX-1 was not inducible by PDT. Prostaglandin (PG) E2 synthesis was also increased in PDT-treated cells, and PGE2 levels were attenuated in cells coincubated with NS-398, indicating that PDT induced the expression of biologically active COX-2. Both porphyrin- and chlorin-based photosensitizers were able to elicit PDT-mediated COX-2 expression. COX-2 was also elevated in radiation-induced fibrosarcoma (RIF) tumors after treatment with PDT. We also observed that systemic administration of NS-398 decreased PDT induction of both PGE2 and vascular endothelial growth factor in treated RIF tumors. Additionally, we demonstrated that NS-398 enhanced PDT responsiveness in RIF tumors without increasing toxicity to normal tissue. These results provide strong evidence that combination procedures involving selective COX-2 inhibitors may improve the therapeutic effectiveness of PDT. PMID- 12124327 TI - Hyaluronan-CD44s signaling regulates matrix metalloproteinase-2 secretion in a human lung carcinoma cell line QG90. AB - We investigated the production of matrix metalloproteinase (MMP) by hyaluronan(HA) stimulation in a human cancer cell line, QG90, that expresses a large amount of CD44s, a HA receptor. Treatment of QG90 with HA strongly activated MMP-2 secretion in a time- and dose-dependent manner. We found that expression of antisense CD44s in QG90 cells substantially inhibited the HA dependent secretion of MMP-2, whereas overexpression of full-length CD44s augmented the HA-dependent secretion of MMP-2. In addition, pretreatment of cells with the neutralizing anti-CD44 antibody significantly inhibited both the HA dependent MMP-2 secretion and the HA-dependent activation of mitogen-activated protein kinase in a dose-dependent manner. Similarly, treatment of cells with a Ras farnesyltransferase inhibitor, manumycin A, strongly inhibited the HA dependent MMP-2 secretion. Moreover, in vitro invasiveness of QG90 and its activation by HA were clearly suppressed by the expression of antisense CD44s. In addition, treatment of cells with anti-CD44, a mitogen-activated protein/extracellular signal-regulated kinase kinase 1 inhibitor, PD98059, or phosphatidylinositol 3'-kinase inhibitors, wortmannin and LY294002, effectively blocked the HA-dependent activation of the invasiveness. In contrast, overexpression of full-length CD44 substantially activated the invasiveness of QG90. Taken together, HA-CD44s signaling plays a key role in the HA-dependent secretion of MMP-2 and, hence, in the invasiveness of QG90 cells. PMID- 12124329 TI - Combined use of oligonucleotide and tissue microarrays identifies cancer/testis antigens as biomarkers in lung carcinoma. AB - High density oligonucleotide microarrays (OMAs) have been used recently to profile gene expression in lung carcinoma tissue homogenates. The length of the lists of potentially interesting genes generated by these studies is daunting, and biological and clinical relevance of these lists remains to be validated. Moreover, specific identification of individual biomarkers that might be used for early detection and surveillance has not been the objective of these early studies. We have developed a schema for combining the data derived from the OMA analysis of a few lung cancer cell lines with immunohistochemical testing of tissue microarrays to rapidly identify biomarkers of potential clinical relevance. Initially, we profiled gene expression in lung tumor cell lines using the Affymetrix HG-U95Av2 OMA. RNA from 2 non-small cell lung cancer (NSCLC) cell lines (A549 and H647) and 2 small cell lung cancer (SCLC) cell lines (SHP-77 and UMC-19) were tested. Cells from 1 histologically and cytogenetically normal bronchial epithelial primary culture from a volunteer who had never smoked and 10 samples of histologically unremarkable lung tissue from resection specimens served as normalization controls. Array results were analyzed with Gene Spring software. Results were confirmed by reverse transcription-PCR in an expanded number of cell lines. We then validated the cell line data by immunohistochemical testing for protein using a tissue microarray containing 187 NSCLC clinical samples. Of the 20 most highly expressed genes in the tumor lines, 6 were members of the cancer/testis antigen (CTAG) gene group including 5 MAGE-A subfamily members and NY-ESO-1. SCLC lines strongly expressed all of the MAGE-A genes as well as NY-ESO-1, whereas NSCLC lines expressed a subset of MAGE-A genes at a lower level of intensity and failed to express NY-ESO-1. Reverse transcription PCR of an extended series of 25 lung cancer cell lines including 13 SCLC, 9 NSCLC, and 3 mesothelioma lines indicated that MAGE-A10 and NY-ESO-1 were expressed only by SCLC, and that MAGE-A1, 3, 6, 12, and 4b were expressed by both SCLC and NSCLC. By immunohistochemistry using the monoclonal antibody 6C1 that recognizes several MAGE-A gene subfamily members, 44% of NSCLC clearly expressed MAGE-A proteins in cytoplasm and/or nucleus. Expression of MAGE-A genes did not correlate with survival but did correlate with histological classification with squamous carcinomas more frequently MAGE-A positive than other NSCLC types (P < 0.00002). We conclude that expression of CTAG gene products, whereas apparently not of prognostic importance, may be useful for early detection and surveillance because of a high level of specificity for central airway squamous and small cell carcinomas. PMID- 12124328 TI - Deficient nonhomologous end-joining activity in cell-free extracts from Brca1 null fibroblasts. AB - BRCA1 ensures genomic stability, at least in part, through a functional role in DNA damage repair. BRCA1 interacts with the Rad50/Mre11/Nbs1 complex that occupies a central role in DNA double-strand break repair mediated by homologous recombination and nonhomologous end joining (NHEJ). NHEJ can be catalyzed by mammalian whole cell extract in a reaction dependent upon DNA ligase IV, Xrcc4, Ku70, Ku80, and DNA-PKcs. Here, we show that under identical cell-free reaction conditions, the addition of antibodies specific for BRCA1 and Rad 50 but not Rad51, inhibits end-joining activity. Cell extracts derived from Brca1-deficient mouse embryonic fibroblasts exhibit reduced end-joining activity independent of the endogenous protein amounts of DNA ligase IV, Ku80, and Ku70. The Brca1 dependent NHEJ activity predominates at the lower concentrations of Mg2+ (0.5 mM); elevated Mg2+ or Mn2+ concentrations (10 mM) dramatically increase overall end-joining activity and abrogates the requirement for Brca1, Xrcc4, and Ku70. The addition of partially purified BRCA1, in association with Rad50/Mre11/Nbs1 complex, complements the NHEJ deficiency of Brca1-null fibroblast extracts. These results suggest a role for Brca1 in NHEJ and in the maintenance of genome integrity. PMID- 12124330 TI - Prognostically important chromosomal aberrations in soft tissue sarcomas: a report of the Chromosomes and Morphology (CHAMP) Study Group. AB - Cytogenetic analysis has not only provided important information on the pathogenesis of soft tissue tumors but, by disclosing distinct chromosomal rearrangements in different histopathological entities, has also come to serve as a valuable diagnostic tool. Little is known as yet about the potential prognostic impact of cytogenetic features detected in these tumors. A total of 239 benign and 221 malignant soft tissue tumors with clonal chromosome aberrations were subdivided according to general karyotypic features, such as degree of complexity and ploidy level, and rearrangements of specific chromosomal regions. The cytogenetic variables were analyzed regarding clinical outcome, using time to metastasis as the end point. Selected variables were then compared with established clinicopathological predictors of metastasis development. When the entire material was considered, 167 of 268 investigated cytogenetic variables were associated with clinical outcome. Focusing on the subset of 151 patients with high-grade sarcoma, 17 variables were identified that, besides grade and size, were associated with increased risk of metastasis development. A final Cox regression analysis identified five independent cytogenetic predictors of adverse outcome; breakpoints in chromosome regions 1p1, 1q4, 14q1, and 17q2, and gain of regions 6p1/p2. An increasing effect on metastatic risk was seen with increasing involvement of the selected cytogenetic variables, even when different histopathological types were studied separately. We conclude that cytogenetic data provide independent prognostic information in soft tissue sarcomas. Furthermore, our results point to specific areas of the genome harboring genes that may influence the metastatic potential of sarcoma cells. PMID- 12124331 TI - Antiestrogen ICI 182,780 decreases proliferation of insulin-like growth factor I (IGF-I)-treated MCF-7 cells without inhibiting IGF-I signaling. AB - Previous studies have suggested that antiestrogens inhibit MCF-7 cell proliferation by alteringthe expression or activity of components of the insulin like growth factor I (IGF-I) signaling pathway, including IGF-I receptor, insulin receptor substrate 1, and phosphatidylinositol 3-kinase. In this report, we examine the effects of the pure antiestrogen ICI 182,780 (ICI) on various targets of IGF-I signaling in MCF-7 cells. ICI treatment led to decreases in the absolute levels of cyclin D1 and cyclin A expression, retinoblastoma protein phosphorylation, and DNA synthesis in IGF-I-treated cells. However, IGF-I retained the ability to induce these events in the presence of ICI, suggesting that ICI treatment did not completely block IGF-I signaling. Consistent with this suggestion, IGF-I-induced phosphorylation of extracellular signal-regulated kinase, AKT, and insulin receptor substrate 1 was unaffected by ICI treatment. Finally, transient expression of either constitutively active phosphatidylinositol 3-kinase or AKT was unable to induce proliferation in ICI treated MCF-7 cells. Together, these results indicate that ICI can inhibit proliferation without blocking IGF-I signaling and suggest a model in which both estrogen receptor and IGF-I signaling regulate cell cycle components and are required for MCF-7 cell proliferation. PMID- 12124332 TI - Average midrange ultraviolet radiation flux and time outdoors predict melanoma risk. AB - Sunlight is the major environmental risk factor for melanoma. Descriptive studies have shown latitudinal variation in population incidence and mortality rates [D. C. Whiteman and A. C. Green, Int. J. Dermatol., 38: 481-489, 1999, and B. K. Armstrong, Australian J. Dermatol., 38 (Suppl. 1): 51-56, 1997]. In analytic studies, individual exposure has been particularly difficult to quantify. Lifetime residential history was coupled with levels of midrange UV radiation (UVB flux) to provide a measure of individual exposure to sunlight thought to be less subject to misclassification and recall bias. Data were analyzed from 718 non-Hispanic white patients with invasive cutaneous melanoma from melanoma clinics in Philadelphia and San Francisco. Matched controls were 945 patients from outpatient clinics with similar catchment areas. The association of melanoma risk and history of UVB flux along with the usual outdoor exposure risk factors were studied. A 10% increase in the average annual UVB flux was associated with a 19% [95% confidence interval (CI), 5-35%] increase in individual odds for melanoma for men and 16% (95% CI, 2-32%) for women. In men, a 10% increase in hours outdoors was associated with a 2.8% (95% CI, 1.2-4.5%) increase in odds. Even in women who could develop a deep tan, a 10% increase in hours outdoors was associated with a 5.8% increase in odds (95% CI, 1.4-10.4%). The association between melanoma risk and average annual UVB flux was strong and consistent for men and for women. The association with total adult hours outdoors was notable for men of all skin types and women who develop a suntan. PMID- 12124333 TI - Growth inhibition of human hepatoma cells by acyclic retinoid is associated with induction of p21(CIP1) and inhibition of expression of cyclin D1. AB - Acyclic retinoid (ACR), a novel synthetic retinoid, can prevent the recurrence of human hepatoma after surgical resection of primary tumors, but the molecular mechanisms by which ACR exerts antitumor effects are not known. In this study, we found that ACR inhibited the growth of three human hepatoma cell lines. In HepG2 cells, this inhibition was associated with an arrest of the cell cycle in G(0) G(1), increased cellular levels of p21(CIP1), decreased levels of the hyperphosphorylated form of the retinoblastoma protein, and decreased levels of cyclin D1, but no significant changes were seen in the levels of the p16(INK4a), p27(KIP1), cyclin-dependent kinase 4, cyclin-dependent kinase 6, glycogen synthase kinase 3beta, or beta-catenin proteins. ACR also caused a decrease in the level of cyclin D1 mRNA. Cotreatment of HepG2 human hepatoma cells with the proteasome inhibitor N-acetyl-Leu-Leu-norleu-al did not prevent the ACR-induced decrease in cyclin D1 protein, in contrast to the protective effect of N-acetyl Leu-Leu-norleu-al on the cyclin D1 protein in cells treated with all-trans retinoic acid. In transient transfection reporter assays, ACR, but not all-trans retinoic acid, inhibited transcription from the cyclin D1 promoter. As reported previously in colon carcinoma cells, we found that in hepatoma cells, cyclin D1 promoter activity is markedly stimulated by the beta-catenin/T-cell factor pathway. Nevertheless, even in the presence of excess beta-catenin, ACR markedly inhibited the transcriptional activity of the cyclin D1 promoter. This is the first systematic study of the inhibitory effects of ACR, or any other retinoid compound, on beta-catenin/T-cell factor-stimulated cyclin D1 promoter activity in human tumor cells. These novel effects of ACR provide further evidence that ACR may be a valuable agent in the chemoprevention and therapy of hepatoma and possibly other human malignancies. PMID- 12124334 TI - Hypoestoxide, a natural nonmutagenic diterpenoid with antiangiogenic and antitumor activity: possible mechanisms of action. AB - We have shown previously that hypoestoxide (HE), a natural diterpenoid [a bicyclo (9, 3, 1) pentadecane], is a potent nonsteroidal anti-inflammatory drug. In this report, we demonstrate that HE also inhibits the growth of a variety of human and murine tumor cell lines in vitro at concentrations ranging from 0.3 to 10 microM and was inactive as a mutagen in the Ames test. HE exhibited highly potent (0.3 10 mg/kg dose ranges) activities against B16 melanoma growth in C57BL/6 mice and P388D1 leukemia in C57BL/6 x DBA/2 F(1) mice, respectively. At a low maximal effective dose of 5 mg/kg, HE induced significant in vivo antitumor activities that were better than or comparable with most of the standard chemotherapeutic antiangiogenic agents tested: cortisone acetate, vincristine, bleomycin, Adriamycin, 5-fluorouracil, cyclophosphamide, and etoposide. All of the agents, except vincristine, had much higher maximal effective doses than HE. HE arrested the growth of human Burkitt lymphoma CA46 cells and HeLa (cervical epitheloid carcinoma) cells in the G2-M phase of the cell cycle, which was caused by interference, either direct or indirect, with actin assembly. Thus, the cell cycle arrest occurred at cytokinesis, as demonstrated by an increase in the number of binucleate cells. Moreover, HE inhibited vascular endothelial growth factor-induced cell proliferation in vitro, with an IC(50) of 28.6 microM, and it significantly inhibited basic fibroblast growth factor-induced angiogenesis on the chick chorioallantoic membrane, with an IC50 of 10 microM. Furthermore, HE inhibited endothelial cell migration on vitronectin, collagen, and fibronectin. Besides its activity as a nonsteroidal anti-inflammatory drug, HE also has promise for the chemotherapy of cancer. PMID- 12124335 TI - PTK787/ZK 222584, a specific vascular endothelial growth factor-receptor tyrosine kinase inhibitor, affects the anatomy of the tumor vascular bed and the functional vascular properties as detected by dynamic enhanced magnetic resonance imaging. AB - Antiangiogenic therapy is a promising new strategy of inhibiting tumor growthand formation of metastases. Recently, a number of compounds with different effects on tumor endothelial cells have entered clinical trials and revealed the need for diagnostic methods to detect their biological activity. Dynamic enhanced magnetic resonance imaging (dyMRI) is used in most clinical trials with antiangiogenic active compounds. We evaluated this method by using PTK787/ZK 222584, a specific inhibitor of the VEGF-receptor tyrosine kinases, which showed antitumoral and antiangiogenic activity in a murine renal cell carcinoma (RENCA) model. After intrarenal application of RENCA cells, mice developed a primary tumor and metastases to the lung and abdominal lymph nodes. After daily oral therapy for 21 days with either PTK787/ZK 222584 at a dose of 50 mg/kg or vehicle, primary tumors of all animals were analyzed by dyMRI. Gadolinium-DOTA (Dotarem) was used as a contrast agent to detect vessel permeability and contrast agent extravasation, whereas intravascular iron oxide nanoparticles (Endorem) were used to detect partial tumor blood volume. Additionally, vessel density, architecture, diameter, and blood flow velocity were investigated by appropriate methods. Surprisingly, no changes in extravasation occurred under treatment with PTK787/ZK 222584 as compared with the control group, whereas a significant decrease in vessel permeability occurred. Furthermore, an increase in partial blood volume was found in the PTK787/ZK 222584-treated group, although vessel density was reduced as seen by histology. Using the corrosion cast technique, reduction in vessel density was significant but not very pronounced and predominantly attributable to the loss of microvessels only. This finding correlated with a shift to large vessel diameters in the primary tumors of PTK787/ZK 222584-treated animals and with reduction of blood flow velocity in the tumor feeding renal artery. From these findings, we conclude that the treatment with PTK787/ZK 222584 primarily reduces the number of tumor microvessels, accompanied by a hemodynamic dilation of the remaining vessels. This dilation could influence the result of dyMRI such that no change in extravasation or even an increase in partial tumor blood volume could be observed. PMID- 12124336 TI - Interleukin 2 gene transfer prevents NKG2D suppression and enhances antitumor efficacy in combination with cisplatin for head and neck squamous cell cancer. AB - Cisplatin has been the most promising single chemotherapeutic agent usedagainst head and neck squamous cell cancer to date. However, dose-related toxicity has been one of the major limiting factors in cisplatin-based therapies, because high doses are required for obtaining a significant antitumor effect. To face the challenge of this limiting factor, a novel interleukin 2 (IL-2)-based combination strategy has been developed. Here we show that the strategy of combination of cisplatin with nonviral IL-2 gene therapy resulted in significant antitumor effects while avoiding dose-limiting toxicity in a head and neck squamous cell cancer murine model. Cisplatin systemic therapy alone suppressed NKG2D expression in lymphocytes. The use of local regional IL-2 gene transfer prevented NKG2D suppression. The combination strategy demonstrated a clear synergistic interaction between cisplatin and IL-2, and NKG2D-based cytotoxicity manifested by increased tumor specific lysis from CTLs and natural killer cells. Moreover, the combination of cisplatin and IL-2 gene therapy greatly enhanced apoptosis and growth inhibition in the treated tumors. This novel combination strategy holds promise for the treatment of head and neck cancer, and the mechanism of NKG2D in activating natural killer and CTL receptors provides a foundation for additional investigation, and development of immune modulation and chemotherapy regimens. PMID- 12124337 TI - Identification and characterization of nuclease-stabilized RNA molecules that bind human prostate cancer cells via the prostate-specific membrane antigen. AB - We have identified two synthetic oligonucleotides (aptamers) that bind to prostate cancer cells,with low nanomolar affinity, via the extracellular portion of the prostate-specificmembrane antigen (PSMA). These two specific aptamers were selected from an initial 40mer library of approximately 6 x 10(14) random sequence RNA molecules for their ability to bind to a recombinant protein representing the extracellular 706 amino acids of PSMA, termed xPSM. Six rounds of in vitro selection were performed, enriching for xPSM binding as monitored by aptamer inhibition of xPSM N-acetyl-alpha-linked acid dipeptidase (NAALADase) enzymatic activity. By round six, 95% of the aptamer pool consisted of just two sequences. These two aptamers, termed xPSM-A9 and xPSM-A10, were cloned and found to be unique, sharing no consensus sequences. The affinity of each aptamer for PSMA was quantitated by its ability to inhibit NAALADase activity. Aptamer xPSM A9 inhibits PSMA noncompetitively with an average K(i) of 2.1 nM, whereas aptamer xPSM-A10 inhibits competitively with an average K(i) of 11.9 nM. Distinct modes of inhibition suggest that each aptamer identifies a unique extracellular epitope of xPSM. One aptamer was truncated from 23.4 kDa to 18.5 kDa and specifically binds LNCaP human prostate cancer cells expressing PSMA but not PSMA-devoid PC-3 human prostate cancer cells. These are the first reported RNA aptamers selected to bind a tumor-associated membrane antigen and the first application of RNA aptamers to a prostate specific cell marker. These aptamers may be used clinically as NAALADase inhibitors or be modified to carry imaging agents and therapeutic agents directed to prostate cancer cells. PMID- 12124338 TI - Lymphotoxin-beta receptor immune interaction promotes tumor growth by inducing angiogenesis. AB - Growth of solid fibrosarcoma tumors in mice was inhibited by the release of a solublelymphotoxin-beta receptor inhibitor (LTbetaR-immunoglobulin fusion protein) from the tumor cells. Tumor growth arrest in mice deficient in the ligand LTalpha1beta2 demonstrated the requirement for activation of the LTbetaR on the tumor cells by host cell-derived LTalpha1beta2. Activation of the LTbetaR resulted in enhanced release of macrophage inflammatory protein-2. Blocked angiogenesis was revealed in LTbetaR inhibitor-producing tumor nodules by immunohistochemistry and in vivo microscopy. The growth arrest of LTbetaR inhibitor-producing fibrosarcomas was overcome by forced MIP-2 expression in the tumor cells. Thus, LTbetaR activation on tumor cells by activated host lymphocytes can initiate a novel proangiogenic pathway leading to organized tumor tissue development. PMID- 12124339 TI - Cancer-related serological recognition of human colon cancer: identification of potential diagnostic and immunotherapeutic targets. AB - Monitoring human antibody recognition of tumor antigens could have potential diagnostic and prognostic significance. Serological analysis of recombinant cDNA expression libraries of human cancer has identified a number of immunogenic tumor antigens. To identify colon cancer antigens associated with a cancer-related serum IgG response, serum samples from 74 patients with colon cancer and 75 normal blood donors were screened for antibody reactivity to 77 serologically defined tumor antigens. The following 13 antigens reacted exclusively with sera from the colon cancer patients and not with sera from normal blood donors: p53, MAGEA3, SSX2, NY-ESO-1, HDAC5, MBD2, TRIP4, NY-CO-45, KNSL6, HIP1R, Seb4D, KIAA1416, and LMNA. Serum samples from 34 of 74 (46%) colon cancer patients detected 1 or more of these 13 antigens. Fifty-three of 74 colon cancer patients were of known clinicopathological stage. Analysis of antibody frequency showed that 5 of 7 (71%) stage I colon cancer patients, 4 of 11 (36%) stage II patients, 2 of 14 (14%) stage III patients, and 11 of 21 (52%) stage IV patients had serum IgG antibody that reacted with 1 or more of the 13 antigens. The mRNA expression patterns of 8 of these 13 antigens were altered in cancer. Three of the 13 antigens were cancer/testis antigens (MAGEA3, SSX2, and NY-ESO-1), which are expressed exclusively in normal gametogenic tissues and aberrantly expressed in a broad range of cancer types. Quantitative real-time reverse transcription-PCR showed that the mRNA expression levels of 2 antigens, HDAC5 and Seb4B, were down regulated in colon cancer, 2 other antigens, KNSL6 and KIAA1416, were up regulated, and another antigen, NY-CO-45, showed a variable level of mRNA expression in colon cancer. With regard to KNSL6 mRNA expression, only trace levels were detected in 15 different normal tissues with the exception of testis, which showed a high level of KNSL6 mRNA expression. In contrast, 9 of 9 colon cancer specimens showed overexpression of KNSL6 mRNA, ranging from 5 to 44 times the level detected in normal colon tissue, indicating that this antigen could also be a valuable therapeutic target. PMID- 12124340 TI - The association of death-associated protein kinase hypermethylation with early recurrence in superficial bladder cancers. AB - Mechanisms for bladder carcinogenesis and the development of recurrentbladder cancer remain unclear. Aberrant methylation of the 5' CpG island is thought to play an important role in the inactivation of the tumor suppressor genes in cancer. To study whether specific or bulk hypermethylation predicts intrabladder recurrence, we determined the frequency of aberrant promoter hypermethylation of seven genes, hMLH1, O(6)-methylguanine-DNA-methyltransferase (MGMT), p16, Von Hippel-Lindau (VHL), death-associated protein kinase (DAP-kinase), glutathione S transferase P1 (GST-P1) and E-cadherin in 55 superficial bladder cancers and 5 normal urothelial epithelia by methylation-specific PCR (MSP). These patients of superficial bladder cancer had been followed prospectively by cystoscopy. Simultaneous hypermethylation of three genes or more among the seven genes was observed in 2 (7%) of 30 patients in the nonrecurrence group and 7 (28%) of 25 patients in the recurrence group. There was a significant concordance between the number of methylated genes and the development of recurrence (P = 0.012). In particular, the recurrence rate for 24 months was 88% for hypermethylation of DAP kinase and 28% for nonmethylation of DAP-kinase. Hypermethylation of DAP-kinase is, therefore, a strong indicator of the superficial bladder cancer associated with a high recurrence rate (P < 0.001; hazards ratio, 7.01). Our results suggest that hypermethylation of DAP-kinase might be a useful prognostic marker for disease recurrence in superficial bladder cancers. PMID- 12124341 TI - The fragile histidine triad/common chromosome fragile site 3B locus and repair deficient cancers. AB - In various studies of sporadic breast cancers, 40-70% were strongly positive for fragile histidine triad (Fhit) protein expression, whereas only 18% of BRCA2 mutant breast cancers demonstrated strong Fhit expression, suggesting that the BRCA2 repair function may be necessary to retain intact fragile common chromosome fragile site 3B(FRA3B)/FHITloci. In the current study, 22 breast tumors with deleterious BRCA1 mutations were analyzed for Fhit expression by immunohistochemistry in a case-control matched pair analysis. Loss of Fhit expression was significantly more frequent in the BRCA1 cancers compared with sporadic breast tumors (9% Fhit positive versus 68% Fhit positive), suggesting that the BRCA1 pathway is also important in protecting the FRA3B/FHIT locus from damage. To investigate the relationship between repair gene deficiencies and induction of chromosome fragile sites in vitro, we have analyzed the frequency of aphidicolin induction of chromosome gaps and breaks in PMS2-, BRCA1-, MSH2-, MLH1 , FHIT-, and TP53-deficient cell lines. Each of the repair-deficient cell lines showed elevated expression of chromosome gaps and breaks, consistent with the proposal that proteins involved in mismatch and double-strand break repair are important in maintaining the integrity of common fragile regions. Correspondingly, genes at common fragile sites may sustain elevated levels of DNA damage in cells with deficient DNA repair proteins such as those mutated in several familial cancer syndromes. PMID- 12124342 TI - Gastric cancers overexpress DARPP-32 and a novel isoform, t-DARPP. AB - Gastric carcinoma is the second most common cause of cancer-related death worldwide. Recently, we have demonstrated that expressed sequence tag AA552509 was frequently amplified and the most consistently overexpressed target at 17q in gastric cancers. Herein, we report that DARPP-32 (dopamine and cAMP-regulated phosphoprotein of M(r) 32,000) is the target gene for overexpression of expressed sequence tag AA552509. In addition, we have identified full-length cDNA of DARPP 32 (GenBank accession number AF464196) with 467 bp of additional untranslated mRNA nucleotides upstream of the previously known translation start site in exon 1. Additionally, we have discovered a novel truncated isoform of DARPP-32 that we named t-DARPP (GenBank accession number AY070271), which is also overexpressed in gastric cancers. Using quantitative real-time reverse transcription-PCR, Western blots, and staining of tumor tissue arrays, the two DARPP mRNA transcripts and proteins were overexpressed in gastric cancer cells and exhibited abundant protein overexpression in neoplastic but not normal gastric epithelial cells. DARPP-32 is the only known protein that acts as a protein phosphatase 1 inhibitor or a protein kinase A inhibitor. The novel truncated isoform, t-DARPP, lacks the phosphorylation site related to protein phosphatase 1 inhibition but maintains the phosphorylation site with the protein kinase A inhibitory effect. Our results reveal for the first time the presence of these signaling molecules in human cancer and suggest that they may be important for gastric tumorigenesis. PMID- 12124343 TI - Replication failure, genome instability, and increased cancer susceptibility in mice with a point mutation in the DNA ligase I gene. AB - DNA ligase I has a key role in DNA replication in the joining together of short replication intermediates. We used gene targeting to introduce a point mutation into the mouse DNA ligase I gene that was present in a human cancer patient with immunodeficiency and a cellular accumulation of DNA replication intermediates. Mutant mice grew more slowly and showed hematopoietic defects at critical stages at which the demands for DNA replication were highest. In the spleen and thymus of mutant mice, the accumulation of a sub-G1, but nonapoptotic, population was observed that we believe may represent cells with single-strand DNA breaks. In mutant bone marrow, occasional DNA replication failure was observed. The level of genome instability was significantly elevated in the spleens of DNA ligase I mutant mice and, because we have found no evidence for any DNA repair defect associated with DNA ligase I deficiency, we believe that this may result directly from the accumulation of replication intermediates. Mutant mice showed an increased incidence of spontaneous cancers with a diverse range of epithelial tumors, particularly cutaneous adnexal tumors that are rare in mice. The origin of the tumors from generalized genome instability, rather than the inactivation of one key control gene, should make DNA ligase I mutant mice a useful model to investigate the relationship between genome instability and cancer in humans. PMID- 12124344 TI - LCX, leukemia-associated protein with a CXXC domain, is fused to MLL in acute myeloid leukemia with trilineage dysplasia having t(10;11)(q22;q23). AB - There are a limited number of reports of acute myeloid leukemia (AML) with t(10;11)(q22;q23). We showed that the MLL gene on 11q23 was fused to the LCX (leukemia-associated protein with a CXXC domain) gene on 10q22 in a de novoadult AML-M2 with trilineage dysplasia having t(10;11)(q22;q23). LCX consisted of at least 12 exons and was predicted to encode a 2136-amino-acid protein with an estimated molecular mass of 235.3 kDa. The LCX protein had a zinc-binding CXXC domain that MLL also contains within a methyltransferase domain, three nuclear localization signals, an alpha-helical coiled-coil region, and two homologous regions to CG2083 proteins of Drosophila melanogaster. We found approximately 12 , 9.5-, and 7.5-kb transcripts of LCX. Expression of the 7.5-kb transcript was detected in fetal heart, lung, and brain, and in adult skeletal muscle, thymus, and ovary. Expression of the 9.5-kb transcript was detected in fetal lung and brain and in adult ovary. Expression of the 12-kb transcript was detected in fetal heart and brain and in adult thymus and ovary. LCX was expressed in 8 of 22 leukemic cell lines, but not in EBV-induced normal B-cell lines. The MLL-LCX fusion protein lacked a CXXC domain of LCX, but retained an alpha-helical coiled coil region at the COOH terminus, similar to MLL-SEPTING, MLL-CDCREL1, MLL AF1p/Eps15, and MLL-AF6, which suggests that these fusion proteins are involved in the pathogenesis of 11q23-associated leukemia through similar mechanisms. PMID- 12124345 TI - Late activation of apoptotic pathways plays a negligible role in mediating the cytotoxic effects of discodermolide and epothilone B in non-small cell lung cancer cells. AB - Discodermolide and epothilone B are promising novel chemotherapeutic agentsthat induce cell death through potent stabilization of microtubules. In this study, we investigated the cellular and molecular events underlying the cytotoxicity of these drugs in non-small cell lung carcinoma (NSCLC) cell lines, focusing on apoptotic characteristics. IC80 concentrations of either drug effectively disrupted the microtubule cytoskeleton of H460 cells and induced cell cycle disturbances with early accumulation in the G2-M phase and development of a hypodiploid cell population in both H460 and SW1573 cells. These events were followed by abnormal chromosome segregation during mitosis and subsequent appearance of multinucleated cells. At later time points, the cells displayed several apoptotic features, such as nuclear condensation and fragmentation as well as Annexin V staining, cleavage of poly(ADP-ribose) polymerase and the activation of caspases. To examine the contribution of apoptotic pathways to the cytotoxic effects of these agents, the involvement of the mitochondria and death receptor routes was studied. At 48 h after treatment, both agents disrupted mitochondria of H460 cells, as indicated by cytochrome c release. Nonetheless, H460 cells stably overexpressing antiapoptotic Bcl-2 or Bcl-xL did not show any protective effect from cell death induced by either drug. Possible death receptor dependency was investigated in H460 cells stably overexpressing dominant-negative FADD, which failed to reduce the cytotoxic effects of discodermolide and epothilone B. To study the role of caspases more directly, the effect of stable overexpression of the caspase-8 inhibitor cytokine response modifier A was studied in H460 cells. Furthermore, the effect of the pancaspase inhibitor z-Val Ala-Asp-fluoromethyl ketone was investigated in a panel of lung carcinoma cell lines. Interestingly, caspase inhibition did not rescue cells from discodermolide or epothilone B-induced cell death. In conclusion, these results demonstrate that despite several apoptotic features detected at relatively late time points after drug exposure, apoptosis is not the dominant mode of cell death and induced low but efficacious concentrations of discodermolide and epothilone B. PMID- 12124346 TI - Alternative translocation breakpoint cluster region 5' to BCL-6 in B-cell non Hodgkin's lymphoma. AB - Chromosomal translocations involving band 3q27 with various different partner chromosomes represent a recurrent cytogenetic abnormality in B-cell non-Hodgkin's lymphoma. In a fraction of these translocations, the chromosomal breakpoint is located within the 5' noncoding region of the BCL-6 proto-oncogene where the BCL 6 major breakpoint region (MBR) maps. As a result of the translocation, BCL-6 expression is deregulated by promoter substitution. However, between 30 and 50% of lymphomas with cytogenetically detectable translocations affecting band 3q27 retain a germ-line configuration at the BCL-6 locus. To identify possible additional breakpoint clusters within 3q27, we cloned a t(3;14)(q27;q32) lymphoma without MBR rearrangement and found a novel breakpoint site located between 245 and 285 kb 5' to BCL-6. Breakpoints within this newly described region, which we called the alternative breakpoint region (ABR), were found to be recurrent in lymphomas carrying t(3q27) chromosomal translocations but devoid of BCL-6 MBR rearrangements. Comparative analysis of multiple lymphomas carrying rearrangements within the ABR showed that the breakpoints cluster within a 20-kb distance. Translocations involving the ABR may juxtapose BCL-6 to distantly acting, heterologous transcriptional regulatory elements which cause deregulation of the proto-oncogene. The identification of BCL-6 ABR provides new tools for the diagnosis of lymphomas carrying aberrations at 3q27 and deregulated BCL-6 genes. PMID- 12124347 TI - Genes involved in DNA repair are mutational targets in endometrial cancers with microsatellite instability. AB - Microsatellite instability (MSI) is observed in a subset of endometrial cancers (ECs) and is attributed to defects in mismatch repair. Mismatch repair deficiency allows for accumulation of mutations in the coding repeats of key target genes, which may be involved in the initiation and progression of MSI+ EC. We examined genes implicated in DNA repair pathways in 38 MSI-high (MSI-H), 10 MSI-low, 25 microsatellite stable ECs, and a selected panel of associated premalignant hyperplasias. Genetic alterations were correlated to histopathological data, including tumor grade and stage. Somatic frameshift mutations were observed in hMLH3, hMSH3, hMSH6, CHK1, and BAX genes in MSI-H endometrial hyperplasias and cancers, whereas mutations in ATR and CDC25C were observed only in MSI-H ECs. Increased mutation frequency in DNA damage response pathway genes including ATR, CHK1, and BAX demonstrated a significant trend with advancing tumor grade (P < 0.05). Our observations of the same mutations at short coding mononucleotide repeats in both premalignant lesions and tumors and association of increased frequency of mutation accumulation with advancing tumor grade suggest that these alterations may play a role in the development and progression of MSI+ EC. PMID- 12124348 TI - Transcript map of the 3.7-Mb D19S112-D19S246 candidate tumor suppressor region on the long arm of chromosome 19. AB - Allelic losses of the q13.3 region of chromosome 19 have been documented in malignant gliomas, neuroblastomas, and ovarian carcinomas, strongly suggesting the presence of a 19q13.3 tumor suppressor gene. Deletion mapping in tumors over the past decade has narrowed the candidate region considerably but has produced partially conflicting results, with some small candidate regions defined only by isolated tumors with deletions. Mutation and expression screening of genes from the most likely candidate regions has failed to identify the gene of interest, perhaps because of the conflicting deletion mapping data. The recently increased public availability of human genomic sequence, combined with improved bioinformatics capabilities, has now made it possible to map much larger candidate regions in considerable detail. We have manually generated a transcript map that spans most of the 19q13.3 tumor suppressor candidate region, from D19S219 to D19S246, with a resolution and quality superior to that of computer generated maps. These results are presented in the hope that an improved map of the candidate region will facilitate further widespread screening and eventual identification of the gene or genes deleted in human gliomas, neuroblastomas, and ovarian cancers. PMID- 12124349 TI - Noscapine alters microtubule dynamics in living cells and inhibits the progression of melanoma. AB - Cellular microtubules, polymers of tubulin, alternate relentlessly between phases of growth and shortening. We now show that noscapine, a tubulin-binding agent, increases the time that cellular microtubules spend idle in a paused state. As a result, most mammalian cell types observed arrest in mitosis in the presence of noscapine. We demonstrate that noscapine-treated murine melanoma B16LS9 cells do not arrest in mitosis but rather become polyploid followed by cell death, whereas primary melanocytes reversibly arrest in mitosis and resume a normal cell cycle after noscapine removal. Furthermore, in a syngeneic murine model of established s.c. melanoma, noscapine treatment resulted in an 85% inhibition of tumor volume on day 17 when delivered by gavage compared with untreated animals (P cytosol, thus increasing the ratio of p27:Cdk2 in the nucleus and inhibiting Cdk2 activity and cell proliferation. Antisense p27 oligonucleotides abrogated the increase in p27 induced by Herceptin and prevented the antibody-mediated reduction in S phase. Transduction of BT-474 cells with an adenovirus-encoding active (myristoylated) Akt (Myr-Akt), but not with a beta-galactosidase control adenovirus, prevented the Herceptin- or LY294002-induced down-regulation of cyclin D1 and of phosphorylated GSK-3beta and prevented the accumulation of p27 in the nucleus and cytosol. In addition, Myr-Akt prevented Herceptin-induced inhibition of the cell proliferation of BT-474 cells and Herceptin-induced apoptosis of SKBR-3 cells. These data suggest that (a) changes in cell cycle- and apoptosis-regulatory molecules after HER2 blockade with Herceptin result, at least in part, from the inhibition of Akt; and (b) disabling PI3K and Akt is required for the antitumor effect of HER2 inhibitors. PMID- 12124353 TI - Ras mediates radioresistance through both phosphatidylinositol 3-kinase-dependent and Raf-dependent but mitogen-activated protein kinase/extracellular signal regulated kinase kinase-independent signaling pathways. AB - Cells transformed by the oncogenic small GTPase, Ras, display a radioresistant phenotype in response to ionizing radiation (IR). To determine the mechanisms by which Ras mediates radioresistance in epithelial cells, we assessed the importance of three major survival pathways that can be activated by Ras [phosphatidylinositol 3-kinase (PI3-K)>Akt, nuclear factor kappaB (NF-kappaB), and Raf>mitogen-activated protein kinase/extracellular signal-regulated kinase kinase (MEK)>extracellular signal-regulated kinase] as necessary or sufficient for Ras-mediated radioresistance in matched pairs of RIE-1 rat intestinal epithelial cells expressing oncogenic Ras or empty vector (RIE-Ras and RIE vector). Inhibiting PI3-K with LY294002 sensitized RIE-1 cells to IR in a dose dependent manner, indicating that PI3-K is necessary for radioresistance, whereas inhibition of NF-kappaB with the super-repressor IkappaBalpha had little effect on survival. Expression of either the constitutively active catalytic subunit of PI3-K, p110alpha-CAAX, or the Ras effector domain mutant 12V/40C, which retains binding to PI3-K but is impaired in binding to other Ras effectors, was sufficient to confer partial radioresistance. Expression of either a constitutively active form of the serine/threonine kinase Raf-1 or the Ras effector domain mutant 12V/35S, which retains binding to Raf but is impaired in binding to other Ras effectors, was also sufficient to confer partial radioresistance. Surprisingly, however, even complete inhibition of MEK activity by using U0126 resulted in no change in post-IR survival whatsoever. Thus, whereas Raf contributes to Ras-mediated radioresistance, this is accomplished through a MEK-independent pathway. Taken together, these results indicate that multiple pathways, including both PI3-K-dependent and Raf-dependent but MEK independent signaling, are required for Ras-mediated radioresistance in epithelial cells. Finally, we demonstrate that Ras-mediated radioresistance can be uncoupled from Ras-mediated transformation, in that PI3-K is required for radioresistance but not transformation, whereas MEK and NF-kappaB are required for transformation but not radioresistance in RIE-1 epithelial cells. PMID- 12124354 TI - Epigenetic factors controlling the BRCA1 and BRCA2 genes in sporadic ovarian cancer. AB - Hypermethylation of the BRCA1 promoter has previously been shown to cause reduced mRNA expression in both breast and ovarian cancers. Nothing is yet known of the expression pattern or methylation status of the promoter region of BRCA2 in sporadic ovarian cancer. Whereas our analysis of 30 sporadic ovarian carcinomas showed a statistically significant reduction of BRCA1 mRNA expression (P = 0.001), it also showed, in contrast, overexpression of BRCA2 mRNA (P = 0.002) in tumor compared with nontumor. Hypermethylation of the BRCA1 promoter highly correlated with decreased BRCA1 expression (P = 0.017). Methylated CpGs at the BRCA2 promoter were either absent or at very low levels in tumor DNA, whereas they were present at high levels in nontumor DNA. Such hypomethylation also correlated with elevated levels of BRCA2 mRNA (P = 0.043) and showed a statistically significant correlation with tumor stage (P = 0.037). This supports the role of methylation in BRCA2 contributing to the pathogenesis of sporadic ovarian cancer. Furthermore, 14 (58.2%) and 9 (56.3%) of all of the cases with aberrant BRCA mRNA expression and methylation patterns, respectively, demonstrated opposing mRNA expression and methylation patterns of the BRCA1 and BRCA2 genes within the same cases. Our findings suggest that both genes may be involved in the development of sporadic ovarian cancer. PMID- 12124355 TI - Overexpression of epidermal growth factor receptor in urothelium elicits urothelial hyperplasia and promotes bladder tumor growth. AB - Although urothelium is constantly bathed in high concentrations of epidermal growth factor (EGF) and most urothelial carcinomas overexpress EGF receptor (EGFr), relatively little is known about the role of EGFr signaling pathway in urothelial growth and transformation. In the present study, we used the uroplakin II gene promoter to drive the urothelial overexpression of EGFr in transgenic mice. Three transgenic lines were established, all expressing a higher level of the EGFr mRNA and protein in the urothelium than the nontransgenic controls. The overexpressed EGFr was functionally active because it was autophosphorylated, and its downstream mitogen-activated protein kinases were highly activated. Phenotypically, the urinary bladders of all transgenic lines developed simple urothelial hyperplasia that was strongly positive for proliferative cell nuclear antigen and weakly positive for bromodeoxyuridine incorporation. When coexpressed with the activated Ha-ras oncogene in double transgenic mice, EGFr had no apparent tumor-enhancing effects over the urothelial hyperplastic phenotype induced by Ha-ras oncogene. However, when coexpressed with the SV40 large T antigen, EGFr accelerated tumor growth and converted the carcinoma in situ of the SV40T mice into high-grade bladder carcinomas, without triggering tumor invasion. Our studies indicate that urothelial overexpression of EGFr can induce urothelial proliferation but not frank carcinoma formation. Our results also suggest that, whereas EGFr and Ha-ras, both of which act in the same signal transduction cascade, stimulated urothelial hyperplasia, they were not synergistic in urothelial tumorigenesis, and EGFr overexpression can cooperate with p53 and pRB dysfunction (as occurring in SV40T transgenic mice) to promote bladder tumor growth. PMID- 12124356 TI - Correspondence re: A. H. Ree et al., expression of a novel factor in human breast cancer cells with metastatic potential. Cancer Res., 59: 4675-4680, 1999. PMID- 12124359 TI - The ecological and evolutionary interface of hummingbird flight physiology. AB - The hovering ability, rapidity of maneuvers and upregulated aerobic capacity of hummingbirds have long attracted the interest of flight biologists. The range of intra- and interspecific variation in flight performance among hummingbirds, however, is equally impressive. A dominant theme in hummingbird evolution is progressive invasion of higher-elevation habitats. Hypobaric challenge is met behaviorally through compensatory changes in wingbeat kinematics, particularly in stroke amplitude. Over evolutionary time scales, montane colonization is associated with increases in body mass and relative wing area. Hovering ability has been well-studied in several North American hummingbird taxa, yet the broad range of interspecific variation in hummingbird axial and appendicular anatomy remains to be assessed mechanistically. Such varied features as tail length, molt condition and substantial weight change due to lipid-loading can dramatically alter various features of the flight envelope. Compared with our present knowledge of hovering performance in hummingbirds, the mechanics of forward flight and maneuvers is not well understood. Relationships among flight-related morphology, competitive ability and foraging behavior have been the focus of numerous studies on tropical and temperate hummingbirds. Ecologists have hypothesized that the primary selective agents on hummingbird flight-related morphology are the behaviors involved in floral nectar consumption. However, flight behaviors involved in foraging for insects may also influence the evolution of wing size and shape. Several comparisons of hummingbird communities across elevational gradients suggest that foraging strategies and competitive interactions within and among species vary systematically across elevations as the costs of flight change with body size and wing shape. PMID- 12124360 TI - Fibre-type specific concentration of focal adhesion kinase at the sarcolemma: influence of fibre innervation and regeneration. AB - In skeletal muscles, focal adhesion complexes (FACs) form part of the costamere, a sarcolemmal protein complex that enables lateral transfer of forces and ensures the stability of the sarcolemma. The present investigation tested whether localisation of a major assembly factor of FACs, focal adhesion kinase (FAK), to the sarcolemma parallels the known modulation of FACs by fibre type (innervation pattern) and fibre regeneration. Immunohistochemical experiments indicated that FAK is preferentially associated with the sarcolemma in a high proportion (>74 %) of the (slow-twitch) type I and (fast-twitch) type IIA fibres in normal rat soleus (N-SOL) muscle and of the type IIA fibres in extensor digitorum longus (N EDL) muscle. In contrast, a low proportion (<15 %) of fast-twitch type IIB and type I fibres in N-EDL showed sarcolemmal FAK immunoreactivity. Cross reinnervation of slow-twitch rat SOL muscle with the fast EDL nerve induced slow to-fast fibre transformation and led to a significant reduction in sarcolemmal FAK immunoreactivity in type I and type IIA fibres. Transplantation of the fast EDL into the slow SOL bed with regeneration and reinnervation of the muscle by the SOL nerve (T-EDL) caused a significant increase in sarcolemmal FAK immunoreactivity in new type I and hybrid I/II fibres and a corresponding reduction in sarcolemmal FAK immunoreactivity in 'normal' IIA and IIB fibres. Conversely, sarcolemmal FAK immunoreactivity in small IIB fibres of T-EDL muscle was increased. Correspondingly, the transplanted and regenerated SOL (reinnervated by the fast EDL nerve) maintained the percentage of FAK-positive sarcolemma in the (regenerated) type I and IIA fibres. Thus, the expression and association of FAK with the sarcolemma are regulated (i) by factors that determine the fibre type and (ii) during fibre regeneration. Our data suggest that the integrity of sarcolemmal FACs is dependent on the fibre type and that FAC turnover is increased during regeneration of muscle fibres. PMID- 12124361 TI - Cellularity changes in developing red and white fish muscle at different temperatures: simulating natural environmental conditions for a temperate freshwater cyprinid. AB - Muscle cellularity patterns in teleost fish have normally been investigated using animals reared under constant temperature conditions. In the present study, Danube bleak (Chalcalburnus chalcoides mento) were reared under two different rising temperature regimes (cold, 12-16 degrees C; warm, 18-20 degrees C) designed to mimic the natural conditions experienced by the fish in temperate freshwater environments. Samples were taken from both groups of animals at intervals during their development. Transverse sections at the level of the anal vent were examined using light and electron microscopy, histochemistry and immunohistochemistry techniques. Total cross-sectional area of red and white muscle, as well as fibre numbers and fibre cross-sectional areas of one epaxial quadrant per specimen, were measured. Analysis of fibre numbers and sizes indicated that white and red myotomal muscles each develop in a different manner. In white muscle, the initial growth phase is dominated by fibre hypertrophy, while the later larval growth phase also includes significant hyperplasia. Red muscle growth is mainly due to hypertrophy within the studied developmental period. The temperature regimes applied in the present study may modify the mechanisms of muscle growth in different ways. For white muscle, pre-hatching hyperplasia (i.e. proliferation of somitic white fibre precursor cells) is reduced under the cold regime whereas post-hatching hyperplasia is not. The inverse is true for white fibre hypertrophy. A similar situation is seen with red muscle except that post-hatching hyperplasia is low and refractory to temperature. Rates of increase in relative amount of red muscle appear to depend not only upon species and temperature but also upon whether the fish have been reared under changing or constant thermal regimes. These findings are discussed in relation to 'landmark' events of early ontogeny (hatching, onset of swimming, start of exogeneous feeding) and to their implications for future accurate interpretation of temperature effects on teleost developmental biology and functional ecology. PMID- 12124362 TI - Function of the heterocercal tail in sharks: quantitative wake dynamics during steady horizontal swimming and vertical maneuvering. AB - The function of the heterocercal tail in sharks has long been debated in the literature. Previous kinematic data have supported the classical theory which proposes that the beating of the heterocercal caudal fin during steady horizontal locomotion pushes posteroventrally on the water, generating a reactive force directed anterodorsally and causing rotation around the center of mass. An alternative model suggests that the heterocercal shark tail functions to direct reaction forces through the center of mass. In this paper, we quantify the function of the tail in two species of shark and compare shark tail function with previous hydrodynamic data on the heterocercal tail of sturgeon Acipenser transmontanus. To address the two models of shark heterocercal tail function, we applied the technique of digital particle image velocimetry (DPIV) to quantify the wake of two species of shark swimming in a flow tank. Both steady horizontal locomotion and vertical maneuvering were analyzed. We used DPIV with both horizontal and vertical light sheet orientations to quantify patterns of wake velocity and vorticity behind the heterocercal tail of leopard sharks (Triakis semifasciata) and bamboo sharks (Chiloscyllium punctatum) swimming at 1.0Ls(-1), where L is total body length. Two synchronized high-speed video cameras allowed simultaneous measurement of shark body position and wake structure. We measured the orientation of tail vortices shed into the wake and the orientation of the central jet through the core of these vortices relative to body orientation. Analysis of flow geometry indicates that the tail of both leopard and bamboo shark generates strongly tilted vortex rings with a mean jet angle of approximately 30 degrees below horizontal during steady horizontal swimming. The corresponding angle of the reaction force is much greater than body angle (mean 11 degrees ) and the angle of the path of motion of the center of mass (mean approximately 0 degrees ), thus strongly supporting the classical model of heterocercal tail function for steady horizontal locomotion. Vortex jet angle varies significantly with body angle changes during vertical maneuvering, but sharks show no evidence of active reorientation of jet angle relative to body angle, as was seen in a previous study on the function of sturgeon tail. Vortex jet orientation is significantly more inclined than the relatively horizontal jet generated by sturgeon tail vortex rings, demonstrating substantial differences in function in the heterocercal tails of sharks and sturgeon. We present a summary of forces on a swimming shark integrating data obtained here on the tail with previous data on pectoral fin and body function. Body orientation plays a critical role in the overall force balance and compensates for torques generated by the tail. The pectoral fins do not generate lift during steady horizontal locomotion, but play an important hydrodynamic role during vertical maneuvering. PMID- 12124363 TI - Squeaking with a sliding joint: mechanics and motor control of sound production in palinurid lobsters. AB - The origin of arthropod sound-producing morphology typically involves modification of two translating body surfaces, such as the legs and thorax. In an unusual structural rearrangement, I show that one lineage of palinurid lobsters lost an antennal joint articulation, which transformed this joint from moving with one degree of freedom into a sliding joint with multiple degrees of freedom. With this sliding joint, 'stick-and-slip' sounds are produced by rubbing the base of each antenna against the antennular plate. To understand the musculo-skeletal changes that occurred during the origin and evolutionary variation of this sound producing mechanism, I examined joint morphology and antennal muscle anatomy across sound-producing and non-sound-producing palinurids. Plectrum movement and antennal muscle activity were measured in a sound-producing species, Panulirus argus. The promotor muscle pulls the plectrum over the file during sound producing and non-sound-producing movements; a higher intensity of muscle activity is associated with sound production. The promotor muscle is larger and attaches more medially in sound-producing palinurids than in non-sound producers. In Panulirus argus, each shingle on the file has an additional ridge; in Palinurus elephas, the shingle surfaces are smooth. These differences in shingle surface features suggest variation in the stick-and-slip properties of the system. Translational motion permitted by the sliding joint is necessary for sound production; hence, the construction of a sliding joint is a key modification in the origin of this sound-producing mechanism. PMID- 12124364 TI - Alpha(1)- and beta-adrenoceptor stimulation differentially activate p38-MAPK and atrial natriuretic peptide production in the perfused amphibian heart. AB - We investigated the activation of p38-MAPK by various adrenergic agents in the perfused Rana ridibunda heart. Phenylephrine (50 micromol l(-1)) rapidly induced the differential activation of all three mitogen-activated protein kinase (MAPK) subfamilies (ERK, JNKs and p38-MAPK) in this experimental system. Focusing on p38 MAPK response to phenylephrine, we found that the kinase phosphorylation reached maximal values at 30 s, declining thereafter to basal values at 15 min. p38-MAPK activation by phenylephrine was verified as exclusively alpha(1)-AR-mediated. Furthermore, SB203580 (1 micromol l(-1)) abolished the kinase phosphorylation by phenylephrine. Isoproterenol (50 micromol l(-1)) was also shown to activate p38 MAPK in a time- and temperature-dependent manner. A marked, sustained p38-MAPK activation profile was observed at 25 degrees C, while at 18 degrees C the kinase response to isoproterenol was modest. Isoproterenol effect on p38-MAPK stimulation was beta-AR-mediated. Immunohistochemical studies revealed the enhanced presence of phosphorylated p38-MAPK and atrial natriuretic peptide (ANP) in both phenylephrine- and isoproterenol-stimulated hearts, a reaction completely blocked by the respective specific antagonists, or the specific p38-MAPK inhibitor SB203580. These findings indicate a functional correlation between p38 MAPK activation and ANP accumulation in the perfused amphibian heart. PMID- 12124365 TI - Jumping in a winged stick insect. AB - The Thailand winged stick insect (Sipyloidea sp.) flees rapidly from a disturbance by jumping forwards when stimulated on the abdomen and backwards when stimulated on the head. The mechanisms underlying these fast movements were analysed by measuring movements of the body and legs from images captured at 250 Hz. A forward jump of both adults and nymphs involves movements of the abdomen and the middle and hind pairs of legs. The abdomen is raised and swung forwards by flexion at the joint with the metathorax and at the joint between the meso- and metathorax. At the same time, the tibiae of the hind and middle legs are extended and their femora depressed. The femoro-tibial joints of the legs are not fully flexed before a jump, and no structures in these joints appear to store muscular energy. The whole jumping sequence takes approximately 100 ms and results in take-off angles of 10-35 degrees at velocities of 0.6-0.8 m s(-1) and with an acceleration of 10 m s(-2). The abdominal angular velocity was 2000 degrees s(-1) and the tip of the abdomen moved at linear velocities of some 1 m s(-1), while the maximum rate of tibial extension was 4000 degrees s(-1). Rapid backward movements result either in the collapse of the body onto the ground, with a displacement away from the stimulus of approximately half a body length, or in the propulsion of the insect off its perch. Neither movement involves curling of the abdomen. From a horizontal posture, the forward jumps result in a displacement of a few body lengths. More lift can be generated in adults by elevating the hind wings as the abdomen is swung forwards and depressing them as the legs lose contact with the ground. In this way, jumps can lead directly to flapping flight. Take-off into flight can, however, be achieved without the abdominal movements seen during jumping. From a vertical posture, a forward jump propels the insect upwards and backwards before it falls to the ground horizontally displaced from its perch. Backward movements result in the insect falling with little horizontal displacement from its perch. PMID- 12124366 TI - Lift and power requirements of hovering flight in Drosophila virilis. AB - The lift and power requirements for hovering flight in Drosophila virilis were studied using the method of computational fluid dynamics. The Navier-Stokes equations were solved numerically. The solution provided the flow velocity and pressure fields, from which the unsteady aerodynamic forces and moments were obtained. The inertial torques due to the acceleration of the wing mass were computed analytically. On the basis of the aerodynamic forces and moments and the inertial torques, the lift and power requirements for hovering flight were obtained. For the fruit fly Drosophila virilis in hovering flight (with symmetrical rotation), a midstroke angle of attack of approximately 37 degrees was needed for the mean lift to balance the insect weight, which agreed with observations. The mean drag on the wings over an up- or downstroke was approximately 1.27 times the mean lift or insect weight (i.e. the wings of this tiny insect must overcome a drag that is approximately 27% larger than its weight to produce a lift equal to its weight). The body-mass-specific power was 28.7 W kg(-1), the muscle-mass-specific power was 95.7 W kg(-1) and the muscle efficiency was 17%. With advanced rotation, larger lift was produced than with symmetrical rotation, but it was more energy-demanding, i.e. the power required per unit lift was much larger. With delayed rotation, much less lift was produced than with symmetrical rotation at almost the same power expenditure; again, the power required per unit lift was much larger. On the basis of the calculated results for power expenditure, symmetrical rotation should be used for balanced, long-duration flight and advanced rotation and delayed rotation should be used for flight control and manoeuvring. This agrees with observations. PMID- 12124367 TI - Toward a catalog for the transcripts and proteins (sialome) from the salivary gland of the malaria vector Anopheles gambiae. AB - Hundreds of Anopheles gambiae salivary gland cDNA library clones have been sequenced. A cluster analysis based on sequence similarity at e(-60) grouped the 691 sequences into 251 different clusters that code for proteins with putative secretory, housekeeping, or unknown functions. Among the housekeeping cDNAs, we found sequences predicted to code for novel thioredoxin, tetraspanin, hemopexin, heat shock protein, and TRIO and MBF proteins. Among secreted cDNAs, we found 21 novel A. gambiae salivary sequences including those predicted to encode amylase, calreticulin, selenoprotein, mucin-like protein and 30-kDa allergen, in addition to antigen 5- and D7-related proteins, three novel salivary gland (SG)-like proteins and eight unique putative secreted proteins (Hypothetical Proteins, HP). The electronic version of this paper contains hyperlinks to FASTA-formatted files for each cluster with the best match to the nonredundant (NR) and conserved domain databases (CDD) in addition to CLUSTAL alignments of each cluster. The N terminus of 12 proteins (SG-1, SG-1-like 2, SG-6, HP 8, HP 9-like, 5' nucleotidase, 30-kDa protein, antigen 5- and four D7-related proteins) has been identified by Edman degradation of PVDF-transferred, SDS/PAGE-separated salivary gland proteins. Therefore, we contribute to the generation of a catalog of A. gambiae salivary transcripts and proteins. These data are freely available and will eventually become an invaluable tool to study the role of salivary molecules in parasite-host/vector interactions. PMID- 12124368 TI - Fuel use and metabolic response to endurance exercise: a wind tunnel study of a long-distance migrant shorebird. AB - This study examines fuel use and metabolism in a group of long-distance migrating birds, red knots Calidris canutus (Scolopacidae), flying under controlled conditions in a wind tunnel for up to 10 h. Data are compared with values for resting birds fasting for the same time. Plasma levels of free fatty acids, glycerol and uric acid were elevated during flight, irrespective of flight duration (1-10 h). Triglyceride levels, the estimated concentration of very-low density lipoproteins (VLDLs) and beta-hydroxybutyrate levels were lower during flight, while glucose levels did not change. In flying birds, plasma levels of uric acid and lipid catabolites were positively correlated with the residual variation in body mass loss, and lipid catabolites with energy expenditure (as measured using the doubly labelled water method), after removing the effect of initial body mass. The plasma metabolite levels indicate: (i) that the rates of catabolism of lipids from adipose tissue and of protein are higher during flight; (ii) that low ketone body concentrations probably facilitate fatty acid release from adipose tissue; (iii) that low triglyceride and VLDL levels do not indicate the use of an additional pathway of fatty acid delivery, as found in small birds; and (iv) that the relationships between energy expenditure, body mass loss and metabolic pattern suggest that a higher individual energy expenditure entails a higher rate of catabolism of both lipids and protein and not a shift in fuel substrate. PMID- 12124369 TI - Wingbeat frequency of barn swallows and house martins: a comparison between free flight and wind tunnel experiments. AB - The flight paths and wingbeat patterns of 39 barn swallows (Hirundo rustica) and 26 house martins (Delichon urbica) were recorded by tracking radar during the spring migration. Depending mostly on flight angle, hirundines performed anything from continuous flapping flight during climbing to single pulse-like wing beats during descent. Unlike most other passerines, hirundines rarely showed regular flapping and rest phases, allowing them to be distinguished from other bird migrants by radar echo signatures. Effective wingbeat frequency (F(eff)) was calculated as the mean number of wing beats per second, including non-flapping phases. Under comparable flight conditions, F(eff) was higher in house martins than in barn swallows. Within species, F(eff) values were higher during climbing and slow flying than during descent. Of the variance in F(eff), 71% could be explained by climb rate, air speed and species; similar results were obtained in the wind tunnel. Under comparable flight conditions, barn swallows and house martins in free flight had significantly lower values of F(eff) than individuals in wind tunnel experiments (by 40% and 32%, respectively). This difference may at least partly be due to the shorter wings of the juveniles tested in the wind tunnel during autumn. However, it seems unlikely that this can account for all of the large difference. It is suggested that wind tunnel experiments might overestimate birds' flight costs compared with free flight. PMID- 12124370 TI - Three-dimensional launch kinematics in leaping, parachuting and gliding squirrels. AB - Leaping, parachuting and gliding are the primary means by which arboreal squirrels negotiate gaps in the canopy. There are notable differences among the three locomotor modes with respect to mid-air postures and aerodynamics, yet it is unclear whether variation should also be expected during the launch phase of locomotion. To address this question, launch kinematic profiles were compared in leaping (Tamias striatus), parachuting (Tamiasciurus hudsonicus) and gliding (Glaucomys volans) squirrels. Animals were filmed launching to the ground from a platform using high-speed video. Statistical comparisons among taxa indicated that only six out of 23 variables were significantly different among the three species. Two were associated with tail kinematics and were a consequence of tail morphology. Two were forelimb-related and discriminated gliding from non-gliding taxa. The remaining two variables were performance attributes, indicating significant variation among the species in take-off velocity and horizontal range. The absence of significant differences in hindlimb kinematics indicates that propulsion is essentially identical in leaping, parachuting and gliding squirrels. PMID- 12124371 TI - Roughness-dependent friction force of the tarsal claw system in the beetle Pachnoda marginata (Coleoptera, Scarabaeidae). AB - This paper studies slide-resisting forces generated by claws in the free-walking beetle Pachnoda marginata (Coleoptera, Scarabaeoidea) with emphasis on the relationship between the dimension of the claw tip and the substrate texture. To evaluate the force range by which the claw can interact with a substrate, forces generated by the freely moving legs were measured using a load cell force transducer. To obtain information about material properties of the claw, its mechanical strength was tested in a fracture experiment, and the internal structure of the fractured claw material was studied by scanning electron microscopy. The bending stress of the claw was evaluated as 143.4-684.2 MPa, depending on the cross-section model selected. Data from these different approaches led us to propose a model explaining the saturation of friction force with increased texture roughness. The forces are determined by the relative size of the surface roughness R(a) (or an average particle diameter) and the diameter of the claw tip. When surface roughness is much bigger than the claw tip diameter, the beetle can grasp surface irregularities and generate a high degree of attachment due to mechanical interlocking with substrate texture. When R(a) is lower than or comparable to the claw tip diameter, the frictional properties of the contact between claw and substrate particles play a key role in the generation of the friction force. PMID- 12124372 TI - Respiratory airflow in a wingless dung beetle. AB - The sealed subelytral cavity of many flightless beetle species is widely acknowledged to be an adaptation to water saving in arid-habitat species. However, this hypothesis relies on the acceptance of two largely untested assumptions: (i) that the movement of respiratory gases is unidirectional from anterior to posterior and (ii) that the coordinated action of the spiracles directs this flow. We tested these assumptions by simultaneously measuring CO(2) and O(2) exchange at the anterior mesothorax, independently of gas exchange in the posterior body, which included the subelytral cavity, of a large apterous beetle, Circellium bacchus. Flow-through respirometry revealed a marked discontinuous gas-exchange cycle (DGC) pattern from the anterior half of the body. Very little CO(2) was released from the posterior body, where the DGC was not apparent. Labelled air was shown to flow forwards from the posterior to the anterior body. Individual sampling from the mesothoracic spiracles revealed that the right mesothoracic spiracle, lying outside the elytral cavity, is the primary route for respiratory gas exchange in C. bacchus at rest. This discovery necessitates a reassessment of the currently assumed role of the subelytral cavity in water conservation and is, to our knowledge, the first demonstration of forward airflow associated with the unilateral use of a single thoracic spiracle for respiration in an insect. PMID- 12124373 TI - View-based navigation in insects: how wood ants (Formica rufa L.) look at and are guided by extended landmarks. AB - Bees, wasps and ants learn landmarks as views from particular vantage points, storing the retinal positions of landmark edges. By moving so as to minimise the difference between their stored and current view, they can return to the vantage point from which a view was taken. We have examined what wood ants learn about a laterally placed, extended landmark, a wall, while walking parallel to it to reach a feeder and how they use this stored information to guide their path. Manipulation of the height of the wall and the ant's starting distance from it reveals that ants maintain a desired distance from the wall by keeping the image of the top of the wall at a particular retinal elevation. Ants can thus employ image matching both for returning to a place and for following a fixed route. Unlike many flying insects, an ant's direction of motion while walking is always along its longitudinal body axis and, perhaps for this reason, it favours its frontal retina for viewing discrete landmarks. We find that ants also use their frontal retina for viewing a laterally placed wall. On a coarse scale, the ant's path along the wall is straight, but on a finer scale it is roughly sinusoidal, allowing the ant to scan the surrounding landscape with its frontal retina. The ant's side-to-side scanning means that the wall is viewed with its frontal retina for phases of the scanning cycle throughout its trajectory. Details of the scanning pattern depend on the scene. Ants scan further to the side that is empty of the wall than to the side containing the wall, and they scan further into the wall side when the wall is of a lower apparent height. We conclude that frontal retina is employed for image storage and for path control. PMID- 12124374 TI - The heart rate/oxygen consumption relationship during cold exposure of the king penguin: a comparison with that during exercise. AB - This study investigated whether exposure to low ambient temperature could be used as an alternative to exercise for calibrating heart rate (fH) against rate of oxygen consumption ((O(2))) for subsequent use of fH to estimate (O(2)) in free ranging animals. Using the relationship between the oxygen pulse (OP, the amount of oxygen used per heart beat) and an index of body condition (or nutritional index, NI), a relationship between fH and (O(2)) was established for resting king penguins exposed to a variety of environmental temperatures. Although there was a small but significant increase in the OP above and below the lower critical temperature (-4.9 degrees C), there was no difference in the relationship obtained between the OP and body condition (NI) obtained above or below the lower critical temperature. These results were then compared with those obtained in a previous study in which the relationship between fH and (O(2)) had been established for king penguins during steady-state exercise. The relationship between OP and NI in the present study was not significantly different from the relationship between resting OP and NI in the previous study. However, the relationship was different from that between active OP and NI. We conclude that, at least for king penguins, although thermoregulation does not affect the relationship between resting OP and NI, temperature cannot be used as an alternative to exercise for calibrating fH against (O(2)) for subsequent use of fH to estimate (O(2)) in free-ranging animals. PMID- 12124375 TI - Smelling home: a good solution for burrow-finding in nocturnal petrels? AB - Many burrowing petrels are able to return to their nests in complete darkness. The well-developed anatomy of their olfactory system and the attraction that food related odour cues have for some petrel species suggest that olfaction may be used to recognize the burrow. In contrast, surface-nesting petrels may rely on visual cues to recognise their nest. We performed experiments on nine species of petrel (with different nesting habits) rendered anosmic either by plugging the nostrils or by injecting zinc sulphate onto the nasal epithelium. Compared with shamtreated control birds, we found that anosmia impaired nest recognition only in species that nest in burrows and that return home in darkness. Therefore, petrels showing nocturnal activity on land may rely on their sense of smell to find their burrows, while petrels showing diurnal activity or surface nesters may disregard olfactory cues in favour of visual guidance. PMID- 12124376 TI - Behavioral evidence for post-pause reduced responsiveness in the electrosensory system of Gymnotus carapo. AB - Gymnotiform weakly electric fish find their way in the dark using a continuously operating active sensory system. An electric organ generates a continuous train of discharges (electric organ discharges, EODs), and tuberous high-frequency electroreceptors monitor the pattern of transcutaneous current flow associated with each EOD. Here, I report that a prior interruption to the continuous train of EODs dramatically affects a response shown by many pulse-type gymnotids. In this so-called novelty response, fish normally raise their electrosensory sampling rate in response to novel sensory stimuli. The gymnotid Gymnotus carapo was induced to pause its EODs briefly, and the novelty response to sensory stimuli given post-pause was analyzed. Mechanosensory stimuli given as early as 20 EODs after a pause elicited clear novelty responses, but strong high-frequency electrical stimuli were ineffective at this time. Moreover, high-frequency electrical stimuli remained less efficient in eliciting normal-sized responses until approximately 2000 EODs, or 40s, after a pause. The post-pause inefficiency of high-frequency stimuli was not due to an inappropriate choice of intensity or their temporal patterning and did not result from the stimulation that caused the pausing. Low-frequency stimuli that also recruited ampullary electroreceptors were more efficient than high-frequency stimuli in eliciting post-pause responses. These findings show that continuous activity is required either to maintain sensitivity to high-frequency electrical stimuli or to ensure that such stimuli are able to modulate efficiently the pacemaker that sets the discharge frequency. PMID- 12124377 TI - Behavioural and neuroendocrine effects of environmental background colour and social interaction in Arctic charr (Salvelinus alpinus). AB - In salmonid fish, a darker skin colour has been suggested to signal social subordination. Substratum colour is another factor affecting skin pigmentation in fish; in the present experiment, juvenile Arctic charr (Salvelinus alpinus) were acclimated and allowed to interact in pairs for 5 days on a pale or dark background colour. Skin darkness was quantified prior to and following social interaction. Furthermore, agonistic behaviour and skin darkness were quantified, together with plasma levels of cortisol, adrenocorticotropin (ACTH) and alpha melanocyte-stimulating hormone (alpha-MSH), and brain levels of monoamines and monoamine metabolites. The results show that fish interacting on a white background were more aggressive than those interacting on a black background. Social subordination resulted in skin darkening in fish kept on a white background, but not in fish kept on a black background. Furthermore, subordinate fish on a white background showed an elevation of brain norepinephric activity, an effect not seen in subordinate fish on a black background. Subordinate fish on both white and black backgrounds showed a similar activation of the brain serotonergic system and the hypothalamic-pituitary-interrenal axis. These results support the suggestion that skin darkening in subordinates acts as a social signal announcing social submission. PMID- 12124378 TI - Conserved tyrosine-147 plays a critical role in the ligand-gated current of the epithelial cation/amino acid transporter/channel CAATCH1. AB - CAATCH1 functions both as an amino-acid-gated cation channel and as a cation dependent, proline-preferring, nutrient amino acid transporter in which the two functions are thermodynamically uncoupled. This study focuses on the ionic channel aspect, in which a Tyr(147) (wild type) to Phe(147) (Y147F) site-directed mutation was investigated by steady-state electrophysiological measurements in the Xenopus laevis oocyte expression system. This tyrosine residue is conserved within the third transmembrane domain in members of the Na(+):neurotransmitter transporter family (SNF), where it plays a role in binding pharmacological ligands such as cocaine to the serotonin (SERT), dopamine (DAT) and norepinephrine (NET) transporters. Epithelial CAATCH1 is a member of the SNF family. The results show that amino acid ligand-gating selectivity and current magnitudes in Na(+)- and K(+)-containing media are differentially altered in CAATCH1 Y147F compared with the wild type. In the absence of amino acid ligands, the channel conductance of Na(+), K(+) and Li(+) that is observed in the wild type was reduced to virtually zero in Y147F. In the wild type, proline binding increased conductance strongly in Na(+)-containing medium and moderately in K(+) containing medium, whereas in Y147F proline failed to elicit any cation currents beyond those of N-methyl-D-glucamine- or water-injected oocytes. In the wild type, methionine binding strongly inhibited inward Na(+) currents, whereas in Y147F it strongly stimulated inward currents in both Na(+) and K(+)-containing media. Indeed, in Na(+)-containing medium, the relative potency ranking for inward current inhibition in the wild type (Met>Leu>Gly>Phe>Thr) was similar to the ranking of ligand-permissive gating of large inward currents in Y147F. In Na(+)-containing medium, current/voltage relationships elicited by ligands in the wild type were complex and reversing, whereas in Y147F they were linear and inwardly rectifying. In K(+)-containing medium, current/voltage relationships remained non-linear in Y147F. Both wild-type and Y147F currents were Cl(-) independent. Together, these data demonstrate a critical role for Tyr(147) in ligand-binding selectivity and modulation of the ionic channel conductance in CAATCH1. The results support the argument that inhibition of the CAATCH1 conductance by free methionine shares some properties in common with ligand inhibition of DAT, SERT, NET and the gamma-aminobutyric acid transporter (GAT1). PMID- 12124379 TI - The CYP4A isoforms hydroxylate epoxyeicosatrienoic acids to form high affinity peroxisome proliferator-activated receptor ligands. AB - Cytochromes P450 of the CYP2C and CYP4A gene subfamilies metabolize arachidonic acid to 5,6-, 8,9-, 11,12-, and 14,15-epoxyeicosatrienoic acids (EETs) and to 19- and 20-hydroxyeicosatetraenoic acids (HETEs), respectively. Abundant functional studies indicate that EETs and HETEs display powerful and often opposing biological activities as mediators of ion channel activity and regulators of vascular tone and systemic blood pressures. Incubation of 8,9-, 11,12-, and 14,15 EETs with microsomal and purified forms of rat CYP4A isoforms led to rapid NADPH dependent metabolism to the corresponding 19- and 20-hydroxylated EETs. Comparisons of reaction rates and catalytic efficiency with those of arachidonic and lauric acids showed that EETs are one of the best endogenous substrates so far described for rat CYP4A isoforms. CYP4A1 exhibited a preference for 8,9-EET, whereas CYP4A2, CYP4A3, and CYP4A8 preferred 11,12-EET. In general, the closer the oxido ring is to the carboxylic acid functionality, the higher the rate of EET metabolism and the lower the regiospecificity for the EET omega-carbon. Analysis of cis-parinaric acid displacement from the ligand-binding domain of the human peroxisome proliferator-activated receptor-alpha showed that omega hydroxylated 14,15-EET bound to this receptor with high affinity (K(i) = 3 +/- 1 nm). Moreover, at 1 microm, the omega-alcohol of 14,15-EET or a 1:4 mixture of the omega-alcohols of 8,9- and 11,12-EETs activated human and mouse peroxisome proliferator-activated receptor-alpha in transient transfection assays, suggesting a role for them as endogenous ligands for these orphan nuclear receptors. PMID- 12124380 TI - Perturbation of a very late step of regulated exocytosis by a secretory carrier membrane protein (SCAMP2)-derived peptide. AB - Secretory carrier membrane proteins (SCAMPs) are conserved four transmembrane spanning proteins associated with recycling vesicular carriers. In mast cells, as in other cell types, SCAMPs 1 and 2 are present in secretory granule membranes and other intracellular membranes. We now demonstrate a population of these SCAMPs in plasma membranes. Although small, this population partially colocalizes with SNARE proteins SNAP-23 and syntaxin 4. A fraction of SCAMPs 1 and 2 also coimmunoprecipitates with SNAP-23. An oligopeptide, E peptide, within the cytoplasmic segment linking the second and third transmembrane spans, particularly of SCAMP2, potently inhibits exocytosis in streptolysin O permeabilized mast cells. The E peptide is unique to SCAMPs and highly conserved among SCAMP isoforms, and minor changes in its sequence abrogate inhibition. It blocks fusion beyond the putative docking step where granules contact the cell surface and each other during compound exocytosis. Blockade is also beyond Ca(2+)/ATP-dependent relocation of SNAP-23, which regulates compound exocytosis, and beyond ATP-dependent priming of fusion. Kinetic ordering of exocytotic inhibitors has shown that E peptide acts later than other perturbants at a stage closely associated with membrane fusion. These findings identify a new reagent for analyzing the final stage of exocytosis and point to the likely action of SCAMP2 in this process. PMID- 12124381 TI - Purification and structural characterization of the central hydrophobic domain of oleosin. AB - The oil bodies of rapeseeds contain a triacylglycerol matrix surrounded by a monolayer of phospholipids embedded with abundant structural alkaline proteins termed oleosins and some other minor proteins. Oleosins are unusual proteins because they contain a 70-80-residue uninterrupted nonpolar domain flanked by relatively polar C- and N-terminal domains. Although the hydrophilic N-terminal domain had been studied, the structural feature of the central hydrophobic domain remains unclear due to its high hydrophobicity. In the present study, we reported the generation, purification, and characterization of a 9-kDa central hydrophobic domain from rapeseed oleosin (19 kDa). The 9-kDa central hydrophobic domain was produced by selectively degrading the N and C termini with enzymes and then purifying the digest by SDS-PAGE and electroelution. We have also reconstituted the central domain into liposomes and synthetic oil bodies to determine the secondary structure of the domain using CD and Fourier transform infrared (FTIR) spectroscopy. The spectra obtained from CD and FTIR were analyzed with reference to structural information of the N-terminal domain and the full-length rapeseed oleosin. Both CD and FTIR analysis revealed that 50-63% of the domain was composed of beta-sheet structure. Detailed analysis of the FTIR spectra indicated that 80% of the beta-sheet structure, present in the central domain, was arranged in parallel to the intermolecular beta-sheet structure. Therefore, interactions between adjacent oleosin proteins would give rise to a stable beta-sheet structure that would extend around the surface of the seed oil bodies stabilizing them in emulsion systems. The strategies used in our present study are significant in that it could be generally used to study difficult proteins with different independent structural domains, especially with long hydrophobic domains. PMID- 12124382 TI - The influence of the Cdc27 subunit on the properties of the Schizosaccharomyces pombe DNA polymerase delta. AB - Schizosaccharomyces pombe DNA polymerase (pol) delta contains four subunits, pol 3, Cdc1, Cdc27, and Cdm1. In this report, we examined the role of Cdc27 on the structure and activity of pol delta. We show that the four-subunit complex is monomeric in structure, in contrast to the previous report that it was a dimer (Zuo, S., Bermudez, V., Zhang, G., Kelman, Z., and Hurwitz, J. (2000) J. Biol. Chem. 275, 5153-5162). This discrepancy between the earlier and recent observations was traced to the marked asymmetric shape of Cdc27. Cdc27 contains two critical domains that govern its role in activating pol delta. The N-terminal region (amino acids (aa) 1-160) binds to Cdc1 and its extreme C-terminal end (aa 362-369) interacts with proliferating cell nuclear antigen (PCNA). Mutants of S. pombe pol delta, containing truncated Cdc27 derivatives deficient in binding to PCNA, supported DNA replication less processively than the wild-type complex. Fusion of a minimal PCNA-binding motif (aa 352-372) to C-terminally truncated Cdc27 derivatives restored processive DNA synthesis in vitro. In vivo, the introduction of these fused Cdc27 derivatives into cdc27Delta cells conferred viability. These data support the model in which Cdc27 plays an essential role in DNA replication by recruiting PCNA to the pol delta holoenzyme. PMID- 12124383 TI - PKC-dependent activation of sphingosine kinase 1 and translocation to the plasma membrane. Extracellular release of sphingosine-1-phosphate induced by phorbol 12 myristate 13-acetate (PMA). AB - Sphingosine-1-phosphate (S1P) is a highly bioactive sphingolipid involved in diverse biological processes leading to changes in cell growth, differentiation, motility, and survival. S1P generation is regulated via sphingosine kinase (SK), and many of its effects are mediated through extracelluar action on G-protein coupled receptors. In this study, we have investigated the mechanisms regulating SK, where this occurs in the cell, and whether this leads to release of S1P extracellularly. The protein kinase C (PKC) activator, phorbol 12-myristate 13 acetate (PMA), induced early activation of SK in HEK 293 cells, and this activation was more specific to the membrane-associated SK. Therefore, we next investigated whether PMA induced translocation of SK to the plasma membrane. PMA induced translocation of both endogenous and green fluorescent protein (GFP) tagged human SK1 (hSK1) to the plasma membrane. PMA also induced phosphorylation of GFP-hSK1. The PMA-induced translocation was abrogated by preincubation with known PKC inhibitors (bisindoylmaleimide and calphostin-c) as well as by the indirect inhibitor of PKC, C(6)-ceramide, supporting a role for PKC in mediating translocation of SK to the plasma membrane. SK activity was not necessary for translocation, because a dominant negative G82D mutation also translocated in response to PMA. Importantly, PKC regulation of SK was accompanied by a 4-fold increase in S1P in the media. These results demonstrate a novel mechanism by which PKC regulates SK and increases secretion of S1P, allowing for autocrine/paracrine signaling. PMID- 12124384 TI - The mCpG-binding domain of human MBD3 does not bind to mCpG but interacts with NuRD/Mi2 components HDAC1 and MTA2. AB - Although mammalian MBD3 contains the mCpG-binding domain (MBD) and is highly homologous with the authentic mCpG-binding protein MBD2, it was reported that the protein does not bind to mCpG specifically. Using recombinant human wild type and mutant MBD3 proteins, we demonstrated that atypical amino acids found in MBD3 MBD, namely, His-30 and Phe-34, are responsible for the inability of MBD3 to bind to mCpG. Interestingly, although H30K/F34Y MBD3 mutant protein binds to mCpG efficiently in vitro, it was not localized at the mCpG-rich pericentromeric regions in mouse cells. We also showed that Y34F MBD2b MBD, which possesses not the mCpG-specific DNA-binding activity but the nonspecific DNA-binding activity, was localized at the pericentromeric regions. These results suggested that the mCpG-specific DNA-binding activity is largely dispensable, and another factor(s) is required for the localization of MBD proteins in vivo. MBD3 was identified as a component of the NuRD/Mi2 complex that shows chromatin remodeling and histone deacetylase activities. We demonstrated that MBD3 MBD is necessary and sufficient for binding to HDAC1 and MTA2, two components of the NuRD/Mi2 complex. It was therefore suggested that mCpG-binding-defective MBD3 has evolutionarily conserved its MBD because of the secondary role played by the MBD in protein-protein interactions. PMID- 12124385 TI - Human mitochondrial 5'-deoxyribonucleotidase. Overproduction in cultured cells and functional aspects. AB - Deoxynucleoside triphosphates (dNTPs) used for mitochondrial DNA replication are mainly formed by phosphorylation of deoxynucleosides imported into mitochondria from the cytosol. We earlier obtained evidence for a mitochondrial 5' nucleotidase (dNT2) with a pronounced specificity for dUMP and dTMP and suggested that the enzyme protects mitochondrial DNA replication from excess dTTP. In humans, accumulation of dTTP causes a mitochondrial genetic disease. We now establish that dNT2 in vivo indeed is located in mitochondria. The native enzyme shows the same substrate specificity and affinity for inhibitors as the recombinant dNT2. We constructed ponasterone-inducible cell lines overproducing dNT2 with and without the green fluorescent protein (GFP) linked to its C terminus. The fusion protein occurred in mitochondria mostly in an inactive truncated form, with only a short C-terminal fragment of dNT2 linked to GFP. No truncation occurred when dNT2 and GFP were not linked. The cell mitochondria then contained a large excess of active dNT2 with or without the mitochondrial presequence. After removal of ponasterone overproduced dNT2 disappeared only slowly from the cells, whereas dNT2-mRNA was lost rapidly. Overproduction of dNT2 did not lead to an increased excretion of pyrimidine deoxyribonucleosides, in contrast to overproduction of the corresponding cytosolic deoxynucleotidase, suggesting that the mitochondrial enzyme does not affect overall cellular deoxynucleotide turnover. PMID- 12124386 TI - The role of Asp-462 in regulating Akt activity. AB - Protein kinase Akt, an important downstream target of phosphatidylinositol 3 kinase, is one of the major survival factors in mammalian cells. It has been shown that phosphorylation of the C-terminal hydrophobic motif is required for Akt activation. The activated Akt then phosphorylates several pro-apoptotic proteins and prevents apoptosis mediated by caspases and the mitochondria. Interestingly, Akt has also been implicated to be a direct substrate of caspases in apoptotic cells induced by Fas (Widmann, C., Gibson, S., and Johnson, G. L. (1998) J. Biol. Chem. 273, 7141-7147) and anoikis (Bachelder, R. E., Wendt, M. A., Fujita, N., Tsuruo, T., and Mercurio, A. M. (2001) J. Biol. Chem. 276, 34702 34707). In this study we showed that cytokine withdrawal resulted in Akt degradation by caspases as well. Furthermore, we demonstrated residue Asp-462 of Akt1 which is just upstream of the hydrophobic motif to be the primary cleavage site. The Akt1 mutant (D462N) that prevented caspase cleavage was more stable during factor withdrawal and enhanced cell survival. The Akt truncation mutant mimicking the caspase cleavage product lost its kinase activity and functioned as a dominant negative to promote cell death. Our results suggest that the balance between Akt and caspase activity controls cell survival. In particular, caspases are able to render Akt inactive and dominantly inhibit the Akt pathway by cleaving off the C-terminal hydrophobic motif. Consequently, the survival signal is quickly down-regulated to allow apoptosis to occur. PMID- 12124387 TI - Periodic DNA methylation in maize nucleosomes and demethylation by environmental stress. AB - When maize seedlings were exposed to cold stress, a genome-wide demethylation occurred in root tissues. Screening of genomic DNA identified one particular fragment that was demethylated during chilling. This 1.8-kb fragment, designated ZmMI1, contained part of the coding region of a putative protein and part of a retrotransposon-like sequence. ZmMI1 was transcribed only under cold stress. Direct methylation mapping revealed that hypomethylated regions spanning 150 bases alternated with hypermethylated regions spanning 50 bases. Analysis of nuclear DNA digested with micrococcal nuclease indicated that these regions corresponded to nucleosome cores and linkers, respectively. Cold stress induced severe demethylation in core regions but left linker regions relatively intact. Thus, methylation and demethylation were periodic in nucleosomes. The following biological significance is conceivable. First, because DNA methylation in nucleosomes induces alteration of gene expression by changing chromatin structures, vast demethylation may serve as a common switch for many genes that are simultaneously controlled upon environmental cues. Second, because artificial demethylation induces heritable changes in plant phenotype (Sano, H., Kamada, I., Youssefian, S., Katsumi, M., and Wabilko, H. (1990) Mol. Gen. Genet. 220, 441 447), altered DNA methylation may result in epigenetic inheritance, in which gene expression is modified without changing the nucleotide sequence. PMID- 12124388 TI - Homologous-pairing activity of the Bacillus subtilis bacteriophage SPP1 replication protein G35P. AB - Genetic evidence suggests that the SPP1-encoded gene 35 product (G35P) is essential for phage DNA replication. Purified G35P binds single-strand DNA (ssDNA) and double-strand (dsDNA) and specifically interacts with SPP1-encoded replicative DNA helicase G40P and SSB protein G36P. G35P promotes joint molecule formation between a circular ssDNA and a homologous linear dsDNA with an ssDNA tail. Joint molecule formation requires a metal ion but is independent of a nucleotide cofactor. Joint molecules formed during these reactions contain a displaced linear ssDNA strand. Electron microscopic analysis shows that G35P forms a multimeric ring structure in ssDNA tails of dsDNA molecules and left handed filaments on ssDNA. G35P promotes strand annealing at the AT-rich region of SPP1 oriL on a supercoiled template. These results altogether are consistent with the hypothesis that the homologous pairing catalyzed by G35P is an integral part of SPP1 DNA replication. The loading of G40P at a d-loop (ori DNA or at any stalled replication fork) by G35P could lead to replication fork reactivation. PMID- 12124389 TI - Fidelity of DNA polymerase epsilon holoenzyme from budding yeast Saccharomyces cerevisiae. AB - DNA polymerases delta and epsilon (pol delta and epsilon) are the major replicative polymerases and possess 3'-5' proofreading exonuclease activities that correct errors arising during DNA replication in the yeast Saccharomyces cerevisiae. This study measures the fidelity of the holoenzyme of wild-type pol epsilon, the 3'-5' exonuclease-deficient pol2-4, a +1 frameshift mutator for homonucleotide runs, pol2C1089Y, and pol2C1089Y pol2-4 enzymes using a synthetic 30-mer primer/100-mer template. The nucleotide substitution rate for wild-type pol epsilon was 0.47 x 10(-5) for G:G mismatches, 0.15 x 10(-5) for T:G mismatches, and less than 0.01 x 10(-5) for A:G mismatches. The accuracy for A opposite G was not altered in the exonuclease-deficient pol2-4 pol epsilon; however, G:G and T:G misincorporation rates increased 40- and 73-fold, respectively. The pol2C1089Y pol epsilon mutant also exhibited increased G:G and T:G misincorporation rates, 22- and 10-fold, respectively, whereas A:G misincorporation did not differ from that of wild type. Since the fidelity of the double mutant pol2-4 pol2C1089Y was not greatly decreased, these results suggest that the proofreading 3'-5' exonuclease activity of pol2C1089Y pol epsilon is impaired even though it retains nuclease activity and the mutation is not in the known exonuclease domain. PMID- 12124390 TI - Membrane-protein interactions contribute to efficient 27-hydroxylation of cholesterol by mitochondrial cytochrome P450 27A1. AB - Mitochondrial cytochrome P450 27A1 (P450 27A1) catalyzes 27-hydroxylation of cholesterol, the first step in the alternative bile acid biosynthetic pathway. Although several crystal structures of P450s are known, no structural information is available for the mammalian, membrane-bound enzymes involved in the removal of cholesterol from the body. We prepared a three-dimensional model of P450 27A1 based on the structure of P450 BM-3. Conservative and non-conservative mutations were introduced at hydrophobic and positively charged residues in the putative F G loop and the adjacent helix G (positions 219-237). Subcellular distribution of the mutant P450s expressed in Escherichia coli was used as a measure of membrane protein interactions. Conservative substitutions of residues located on the surface, according to our model, L219V, L219I, Y220F, F223Y, L224I, R229K, V231L, F234Y, K236R, and R237K, weakened the association of the mutant P450s with the membrane and led to the appearance of up to 21% of P450 27A1 in the bacterial cytosol. It is likely that the mutated side chains are involved in binding to membrane phospholipids. Substitutions in the F-G loop did not significantly affect the K(m) value for cholesterol hydroxylation. However, non-conservative mutants, L219N, Y220A, Y220S, F223A, K226R, and R229A, had significantly impaired catalytic properties, indicating strict requirements for the size and polarity of the side chains at these positions for the catalysis. The results provide insight into the membrane topology of mitochondrial P450s and indicate the importance of membrane-protein interactions in the efficiency of reactions catalyzed by P450 27A1. PMID- 12124391 TI - Distinct sites on G protein beta gamma subunits regulate different effector functions. AB - G proteins interact with effectors at multiple sites and regulate their activity. The functional significance of multiple contact points is not well understood. We previously identified three residues on distinct surfaces of Gbetagamma that are crucial for G protein-coupled inward rectifier K(+) (GIRK) channel activation. Here we show that mutations at these sites, S67K, S98T, and T128F, abolished or reduced direct GIRK current activation in inside-out patches, but, surprisingly, all mutants synergized with sodium in activating K(+) currents. Each of the three Gbeta(1) mutants bound the channel indicating that the defects reflected mainly functional impairments. We tested these mutants for functional interactions with effectors other than K(+) channels. With N-type calcium channels, Gbetagamma wild type and mutants all inhibited basal currents. A depolarizing pre-pulse relieved Gbetagamma inhibition of Ca(2+) currents by the wild type and the S98T and T128F mutants but not the S67K mutant. Both wild type and mutant Gbetagamma subunits activated phospholipase C beta(2) with similar potencies; however, the S67K mutant showed reduced maximal activity. These data establish a pattern where mutations can alter the Gbetagamma regulation of a specific effector function without affecting other Gbetagamma-mediated functions. Moreover, Ser-67 showed this pattern in all three effectors tested, suggesting that this residue participates in a common functional domain on Gbeta(1) that regulates several effectors. These data show that distinct domains within Gbetagamma subserve specific functional roles. PMID- 12124392 TI - Group V phospholipase A2 induces leukotriene biosynthesis in human neutrophils through the activation of group IVA phospholipase A2. AB - We reported previously that exogenously added human group V phospholipase A(2) (hVPLA(2)) could elicit leukotriene B(4) (LTB(4)) biosynthesis in human neutrophils (Han, S. K., Kim, K. P., Koduri, R., Bittova, L., Munoz, N. M., Leff, A. R., Wilton, D. C., Gelb, M. H., and Cho, W. (1999) J. Biol. Chem. 274, 11881 11888). To determine the mechanism of the hVPLA(2)-induced LTB(4) biosynthesis in neutrophils, we thoroughly examined the effects of hVPLA(2) and their lipid products on the activity of group IVA cytosolic PLA(2) (cPLA(2)) and LTB(4) biosynthesis under different conditions. As low as 1 nm exogenous hVPLA(2) was able to induce the release of arachidonic acid (AA) and LTB(4). Typically, AA and LTB(4) were released in two phases, which were synchronized with a rise in intracellular calcium concentration ([Ca(2+)](i)) near the perinuclear region and cPLA(2) phosphorylation. A cellular PLA(2) assay showed that hVPLA(2) acted primarily on the outer plasma membrane, liberating fatty acids and lysophosphatidylcholine (lyso-PC), whereas cPLA(2) acted on the perinuclear membrane. Lyso-PC and polyunsaturated fatty acids including AA activated cPLA(2) and 5-lipoxygenase by increasing [Ca(2+)](i) and inducing cPLA(2) phosphorylation, which then led to LTB(4) biosynthesis. The delayed phase was triggered by the binding of secreted LTB(4) to the cell surface LTB(4) receptor, which resulted in a rise in [Ca(2+)](i) and cPLA(2) phosphorylation through the activation of mitogen-activated protein kinase, extracellular signal-regulated kinase 1/2. These results indicate that a main role of exogenous hVPLA(2) in neutrophil activation and LTB(4) biosynthesis is to activate cPLA(2) and 5 lipoxygenase primarily by liberating from the outer plasma membrane lyso-PC that induces [Ca(2+)](i) increase and cPLA(2) phosphorylation and that hVPLA(2) induced LTB(4) production is augmented by the positive feedback activation of cPLA(2) by LTB(4). PMID- 12124393 TI - DNA methylation down-regulates CDX1 gene expression in colorectal cancer cell lines. AB - CDX1 is a homeobox protein that inhibits proliferation of intestinal epithelial cells and regulates intestine-specific genes involved in differentiation. CDX1 expression is developmentally and spatially regulated, and its expression is aberrantly down-regulated in colorectal cancers and colon cancer-derived cell lines. However, very little is known about the molecular mechanism underlying the regulation of CDX1 gene expression. In this study, we characterized the CDX1 gene structure and identified that its gene promoter contained a typical CpG island with a CpG observed/expected ratio of 0.80, suggesting that the CDX1 gene is a target of aberrant methylation. Alterations of DNA methylation in the CDX1 gene promoter were investigated in a series of colorectal cancer cell lines. Combined Bisulfite Restriction Analysis (COBRA) and bisulfite sequencing analysis revealed that the CDX1 promoter is methylated in CDX1 non-expressing colorectal cancer cell lines but not in human normal colon tissue and T84 cells, which express CDX1. Treatment with 5'-aza-2'-deoxycytidine (5-azaC), a DNA methyltransferase inhibitor, induced CDX1 expression in the colorectal cancer cell lines. Furthermore, de novo methylation was determined by establishing stably transfected clones of the CDX1 promoter in SW480 cells and demethylation by 5 azaC-activated reporter gene expression. These results indicate that aberrant methylation of the CpG island in the CDX1 promoter is one of the mechanisms that mediate CDX1 down-regulation in colorectal cancer cell lines. PMID- 12124394 TI - Iron detoxification properties of Escherichia coli bacterioferritin. Attenuation of oxyradical chemistry. AB - Bacterioferritin (EcBFR) of Escherichia coli is an iron-mineralizing hemoprotein composed of 24 identical subunits, each containing a dinuclear metal-binding site known as the "ferroxidase center." The chemistry of Fe(II) binding and oxidation and Fe(III) hydrolysis using H(2)O(2) as oxidant was studied by electrode oximetry, pH-stat, UV-visible spectrophotometry, and electron paramagnetic resonance spin trapping experiments. Absorption spectroscopy data demonstrate the oxidation of two Fe(II) per H(2)O(2) at the ferroxidase center, thus avoiding hydroxyl radical production via Fenton chemistry. The oxidation reaction with H(2)O(2) corresponds to [Fe(II)(2)-P](Z) + H(2)O(2) --> [Fe(III)(2)O-P](Z) + H(2)O, where [Fe(II)(2)-P](Z) represents a diferrous ferroxidase center complex of the protein P with net charge Z and [Fe(III)(2)O-P](Z) a micro-oxo-bridged diferric ferroxidase complex. The mineralization reaction is given by 2Fe(2+) + H(2)O(2) + 2H(2)O --> 2FeOOH((core)) + 4H(+), where two Fe(II) are again oxidized by one H(2)O(2). Hydrogen peroxide is shown to be an intermediate product of dioxygen reduction when O(2) is used as the oxidant in both the ferroxidation and mineralization reactions. Most of the H(2)O(2) produced from O(2) is rapidly consumed in a subsequent ferroxidase reaction with Fe(II) to produce H(2)O. EPR spin trapping experiments show that the presence of EcBFR greatly attenuates the production of hydroxyl radical during Fe(II) oxidation by H(2)O(2), consistent with the ability of the bacterioferritin to facilitate the pairwise oxidation of Fe(II) by H(2)O(2), thus avoiding odd electron reduction products of oxygen and therefore oxidative damage to the protein and cellular components through oxygen radical chemistry. PMID- 12124395 TI - The cystic fibrosis mutation G551D alters the non-Michaelis-Menten behavior of the cystic fibrosis transmembrane conductance regulator (CFTR) channel and abolishes the inhibitory Genistein binding site. AB - Loss of cystic fibrosis transmembrane conductance regulator (CFTR) channel activity explains most of the manifestations of the cystic fibrosis (CF) disease. To understand the consequences of CF mutations on CFTR channel activity, we compared the pharmacological properties of wild-type (wt) and G551D-CFTR. Dose dependent relationships of wt-CFTR activated by genistein follows a non-Michaelis Menten behavior consistent with the presence of two binding sites. With phosphorylated CFTR, a high affinity site for genistein is the activator (K(s) approximately 3 microm), whereas a second site of low affinity (K(i) approximately 75 microm) is the inhibitor. With non-phosphorylated CFTR, K(s) was increased (K(s) approximately 12 microm), but K(i) was not affected (K(i) approximately 70 microm). In G551D-CFTR cells, channel activity was recovered by co-application of forskolin and genistein in a dose-dependent manner. A further stimulation of G551D-CFTR channel activity was measured at concentrations from 30 microm to 1 mm. The dose response is described by a classical Michaelis-Menten kinetics with only a single apparent site (K(m) approximately 11 microm). Our results suggest glycine 551 in NBD1 as an important location within the low affinity inhibitory site for genistein and offers new evidence for pharmacological alteration caused by an NBD1 mutation of CFTR. This study also reveals how a mutation of an ion channel converts a non-Michaelis-Menten behavior (two binding sites) into a classical Michaelis-Menten model (one binding site). PMID- 12124396 TI - Two sequence motifs from HIF-1alpha bind to the DNA-binding site of p53. AB - There is evidence that hypoxia-inducible factor-1alpha (HIF-1alpha) interacts with the tumor suppressor p53. To characterize the putative interaction, we mapped the binding of the core domain of p53 (p53c) to an array of immobilized HIF-1alpha-derived peptides and found two peptide-sequence motifs that bound to p53c with micromolar affinity in solution. One sequence was adjacent to and the other coincided with the two proline residues of the oxygen-dependent degradation domain (P402 and P564) that act as switches for the oxygen-dependent regulation of HIF-1alpha. The binding affinity was independent of the hydroxylation state of P564. We found from NMR spectroscopy that these sequence motifs bind to the DNA binding site of p53c. Because the two sequences are homologous and separated by 120 residues, and one is in a largely unstructured transactivation domain, we speculate that each sequence motif in HIF-1alpha binds to a different subunit of the p53 tetramer, leading to very tight binding. The binding data support the proposal that p53 provides a route for the degradation in hypoxic tumor cells of HIF-1alpha that is not hydroxylated at the two proline residues. PMID- 12124397 TI - Concentration fluctuations in a mesoscopic oscillating chemical reaction system. AB - Under sustained pumping, kinetics of macroscopic nonlinear biochemical reaction systems far from equilibrium either can be in a stationary steady state or can execute sustained oscillations about a fixed mean. For a system of two dynamic species X and Y, the concentrations n(x) and n(y) will be constant or will repetitively trace a closed loop in the (n(x), n(y)) phase plane, respectively. We study a mesoscopic system with n(x) and n(y) very small; hence the occurrence of random fluctuations modifies the deterministic behavior and the law of mass action is replaced by a stochastic model. We show that n(x) and n(y) execute cyclic random walks in the (n(x), n(y)) plane whether or not the deterministic kinetics for the corresponding macroscopic system represents a steady or an oscillating state. Probability distributions and correlation functions for n(x)(t) and n(y)(t) show quantitative but not qualitative differences between states that would appear as either oscillating or steady in the corresponding macroscopic systems. A diffusion-like equation for probability P(n(x), n(y), t) is obtained for the two-dimensional Brownian motion in the (n(x), n(y)) phase plane. In the limit of large n(x), n(y), the deterministic nonlinear kinetics derived from mass action is recovered. The nature of large fluctuations in an oscillating nonequilibrium system and the conceptual difference between "thermal stochasticity" and "temporal complexity" are clarified by this analysis. This result is relevant to fluorescence correlation spectroscopy and metabolic reaction networks. PMID- 12124398 TI - Nitric oxide is consumed, rather than conserved, by reaction with oxyhemoglobin under physiological conditions. AB - Although irreversible reaction of NO with the oxyheme of hemoglobin (producing nitrate and methemoglobin) is extremely rapid, it has been proposed that, under normoxic conditions, NO binds preferentially to the minority deoxyheme to subsequently form S-nitrosohemoglobin (SNOHb). Thus, the primary reaction would be conservation, rather than consumption, of nitrogen oxide. Data supporting this conclusion were generated by using addition of a small volume of a concentrated aqueous solution of NO to a normoxic hemoglobin solution. Under these conditions, however, extremely rapid reactions can occur before mixing. We have thus compared bolus NO addition to NO generated homogeneously throughout solution by using NO donors, a more physiologically relevant condition. With bolus addition, multiple hemoglobin species are formed (as judged by visible spectroscopy) as well as both nitrite and nitrate. With donor, only nitrate and methemoglobin are formed, stoichiometric with the amount of NO liberated from the donor. Studies with increasing hemoglobin concentrations reveal that the nitrite-forming reaction (which may be NO autoxidation under these conditions) competes with reaction with hemoglobin. SNOHb formation is detectable with either bolus or donor; however, the amounts formed are much smaller than the amount of NO added (less than 1%). We conclude that the reaction of NO with hemoglobin under normoxic conditions results in consumption, rather than conservation, of NO. PMID- 12124399 TI - A human embryonic hemoglobin inhibits Hb S polymerization in vitro and restores a normal phenotype to mouse models of sickle cell disease. AB - The principle that developmentally silenced globin genes can be reactivated in adults with defects in beta-globin gene expression has been well established both in vitro and in vivo. In practice, levels of developmental stage-discordant fetal gamma globin that can be achieved by using currently approved therapies are generally insufficient to fully resolve typical clincopathological features of sickle cell disease. The therapeutic potential of another developmentally silenced globin--embryonic epsilon globin--has been difficult to evaluate in the absence of a convenient expression system or an appropriate experimental model. The current work analyzes the antisickling properties of an epsilon -globin containing heterotetramer (Hb Gower-2) both in vitro as well as in vivo in a well established mouse model of sickle cell anemia. These animals, expressing 100% human Hb S, display a chronic hemolytic anemia with compensatory marrow and extramedullary erythropoiesis, abundant circulating sickled erythrocytes, and chronic tissue damage evidenced by parallel histopathological and functional deficits. By comparison, related mice that coexpress Hb S as well as Hb Gower-2 exhibit normal physiological, morphological, histological, and functional attributes. Subsequent in vitro analyses substantiate results from whole-animal studies, indicating that the polymerization of deoxygenated Hb S can be significantly slowed by relatively small quantities of Hb Gower-2. Together, the in vivo and in vitro analyses suggest that reactivation of epsilon-globin gene expression would be therapeutically beneficial to adults with sickle phenotypes, and provide a rationale for detailed investigations into the molecular basis for its developmental silencing. PMID- 12124400 TI - Increasing plant susceptibility to Agrobacterium infection by overexpression of the Arabidopsis nuclear protein VIP1. AB - Agrobacterium is a unique model system as well as a major biotechnological tool for genetic manipulation of plant cells. It is still unknown, however, whether host cellular factors exist that are limiting for infection, and whether their overexpression in plant cells can increase the efficiency of the infection. Here, we examined the effect of overexpression in tobacco plants of an Arabidopsis gene, VIP1, which encodes a recently discovered cellular protein required for Agrobacterium infection. Our results indicate that VIP1 is imported into the plant cell nucleus via the karyopherin alphadependent pathway and that elevated intracellular levels of VIP1 render the host plants significantly more susceptible to transient and stable genetic transformation by Agrobacterium, probably because of the increased nuclear import of the transferred-DNA. PMID- 12124401 TI - Graded contribution of the Gbeta gamma binding domains to GIRK channel activation. AB - G protein coupled inwardly rectifying K(+) channels (GIRK/Kir3.x) are mainly activated by a direct interaction with Gbetagamma subunits, released upon the activation of inhibitory neurotransmitter receptors. Although Gbetagamma binding domains on all four subunits have been found, the relative contribution of each of these binding sites to channel gating has not yet been defined. It is also not known whether GIRK channels open once all Gbetagamma sites are occupied, or whether gating is a graded process. We used a tandem tetrameric approach to enable the selective elimination of specific Gbetagamma binding domains in the tetrameric context. Here, we show that tandem tetramers are fully operational. Tetramers with only one wild-type channel subunit showed receptor-independent high constitutive activity. The presence of two or three wild-type subunits reconstituted receptor activation gradually. Furthermore, a tetramer with no GIRK1 Gbetagamma binding domain displayed slower kinetics of activation. The slowdown in activation was found to be independent of regulator of G protein signaling or receptor coupling, but this slowdown could be reversed once only one Gbetagamma binding domain of GIRK1 was added. These results suggest that partial activation can occur under low Gbetagamma occupancy and that full activation can be accomplished by the interaction with three Gbetagamma binding subunits. PMID- 12124402 TI - Mistargeting hippocampal axons by expression of a truncated Eph receptor. AB - Topographic mapping of axon terminals is a general principle of neural architecture that underlies the interconnections among many neural structures. The Eph family tyrosine kinase receptors and their ligands, the ephrins, have been implicated in the formation of topographic projection maps. We show that multiple Eph receptors and ligands are expressed in the hippocampus and its major subcortical projection target, the lateral septum, and that expression of a truncated Eph receptor in the mouse brain results in a pronounced alteration of the hippocamposeptal topographic map. Our observations provide strong support for a critical role of Eph family guidance factors in regulating ontogeny of hippocampal projections. PMID- 12124403 TI - B-type natriuretic peptide -- a window to the heart. PMID- 12124404 TI - Rapid measurement of B-type natriuretic peptide in the emergency diagnosis of heart failure. AB - BACKGROUND: B-type natriuretic peptide is released from the cardiac ventricles in response to increased wall tension. METHODS: We conducted a prospective study of 1586 patients who came to the emergency department with acute dyspnea and whose B type natriuretic peptide was measured with a bedside assay. The clinical diagnosis of congestive heart failure was adjudicated by two independent cardiologists, who were blinded to the results of the B-type natriuretic peptide assay. RESULTS: The final diagnosis was dyspnea due to congestive heart failure in 744 patients (47 percent), dyspnea due to noncardiac causes in 72 patients with a history of left ventricular dysfunction (5 percent), and no finding of congestive heart failure in 770 patients (49 percent). B-type natriuretic peptide levels by themselves were more accurate than any historical or physical findings or laboratory values in identifying congestive heart failure as the cause of dyspnea. The diagnostic accuracy of B-type natriuretic peptide at a cutoff of 100 pg per milliliter was 83.4 percent. The negative predictive value of B-type natriuretic peptide at levels of less than 50 pg per milliliter was 96 percent. In multiple logistic-regression analysis, measurements of B-type natriuretic peptide added significant independent predictive power to other clinical variables in models predicting which patients had congestive heart failure. CONCLUSIONS: Used in conjunction with other clinical information, rapid measurement of B-type natriuretic peptide is useful in establishing or excluding the diagnosis of congestive heart failure in patients with acute dyspnea. PMID- 12124405 TI - Hepatitis B e antigen and the risk of hepatocellular carcinoma. AB - BACKGROUND: The presence of hepatitis B e antigen (HBeAg) in serum indicates active viral replication in hepatocytes. HBeAg is thus a surrogate marker for the presence of hepatitis B virus DNA. We conducted a prospective study to determine the relation between positivity for hepatitis B surface antigen (HBsAg) and HBeAg and the development of hepatocellular carcinoma. METHODS: In 1991 and 1992, we enrolled 11,893 men without evidence of hepatocellular carcinoma (age range, 30 to 65 years) from seven townships in Taiwan. Serum samples obtained at the time of enrollment were tested for HBsAg and HBeAg by radioimmunoassay. The diagnosis of hepatocellular carcinoma was ascertained through data linkage with the computerized National Cancer Registry in Taiwan and with death certificates. We performed a multiple regression analysis to determine the relative risk of hepatocellular carcinoma among men who were positive for HBsAg alone or for HBsAg and HBeAg, as compared with those who were negative for both. RESULTS: There were 111 cases of newly diagnosed hepatocellular carcinoma during 92,359 person-years of follow-up. The incidence rate of hepatocellular carcinoma was 1169 cases per 100,000 person-years among men who were positive for both HBsAg and HBeAg, 324 per 100,000 person-years for those who were positive for HBsAg only, and 39 per 100,000 person-years for those who were negative for both. After adjustment for age, sex, the presence or absence of antibodies against hepatitis C virus, cigarette-smoking status, and use or nonuse of alcohol, the relative risk of hepatocellular carcinoma was 9.6 (95 percent confidence interval, 6.0 to 15.2) among men who were positive for HBsAg alone and 60.2 (95 percent confidence interval, 35.5 to 102.1) among those who were positive for both HBsAg and HBeAg, as compared with men who were negative for both. CONCLUSIONS: Positivity for HBeAg is associated with an increased risk of hepatocellular carcinoma. PMID- 12124406 TI - Osteoprotegerin deficiency and juvenile Paget's disease. AB - BACKGROUND: Juvenile Paget's disease, an autosomal recessive osteopathy, is characterized by rapidly remodeling woven bone, osteopenia, fractures, and progressive skeletal deformity. The molecular basis is not known. Osteoprotegerin deficiency could explain juvenile Paget's disease because osteoprotegerin suppresses bone turnover by functioning as a decoy receptor for osteoclast differentiation factor (also called RANK ligand). METHODS: We evaluated two apparently unrelated Navajo patients with juvenile Paget's disease for defects in the gene encoding osteoprotegerin (TNFRSF11B) using polymerase-chain-reaction (PCR) amplification followed by direct sequencing and Southern blotting of genomic DNA. Genetic markers near TNFRSF11B were evaluated by both a PCR method that involved sequence-tagged site-content mapping of a deletion of TNFRSF11B and PCR spanning the DNA break points. RESULTS: Both patients had a homozygous deletion of TNFRSF11B, with identical break points, on chromosome 8q24.2. The defect spans approximately 100 kb, but neighboring genes are intact. We found that serum levels of osteoprotegerin and soluble osteoclast differentiation factor were undetectable and markedly increased, respectively. CONCLUSIONS: Juvenile Paget's disease can result from osteoprotegerin deficiency caused by homozygous deletion of TNFRSF11B. PMID- 12124408 TI - Images in clinical medicine. An unusual case of orthopnea. PMID- 12124407 TI - Toll-like receptor 4 polymorphisms and atherogenesis. AB - BACKGROUND: The ability to mount a prominent inflammatory response to bacterial pathogens confers an advantage in innate immune defense but may signal an increased risk of atherosclerosis. We determined whether recently discovered genetic variants of toll-like receptor 4 (TLR4) that confer differences in the inflammatory response elicited by bacterial lipopolysaccharide are related to the development of atherosclerosis. METHODS: As part of the five-year follow-up in the Bruneck (Italy) Study, we screened 810 persons in the study cohort for the TLR4 polymorphisms Asp299Gly and Thr399Ile. The extent and progression of carotid atherosclerosis were assessed by high-resolution duplex ultrasonography. RESULTS: As compared with subjects with wild-type TLR4, the 55 subjects with the Asp299Gly TLR4 allele had lower levels of certain proinflammatory cytokines, acute-phase reactants, and soluble adhesion molecules, such as interleukin-6 and fibrinogen. Although these subjects were found to be more susceptible to severe bacterial infections, they had a lower risk of carotid atherosclerosis (odds ratio, 0.54; 95 percent confidence interval, 0.32 to 0.98; P=0.05) and a smaller intima-media thickness in the common carotid artery (regression coefficient, -0.07; 95 percent confidence interval, -0.12 to -0.02; P=0.01). CONCLUSIONS: The Asp299Gly TLR4 polymorphism, which attenuates receptor signaling and diminishes the inflammatory response to gram-negative pathogens, is associated with a decreased risk of atherosclerosis. This finding is consistent with the hypothesis that innate immunity may play a part in atherogenesis. PMID- 12124409 TI - Clinical practice. Postpartum depression. PMID- 12124410 TI - Case Records of the Massachusetts General Hospital. Weekly clinicopathological exercises. Case 22-2002. A 37-year-old man with unexplained fever after a long trip through South America. PMID- 12124411 TI - Hepatitis B e Antigen--the dangerous endgame of hepatitis B. PMID- 12124412 TI - Genetic control of bone remodeling--insights from a rare disease. PMID- 12124413 TI - Long-term effects of early genetic influences on behavior. PMID- 12124414 TI - Mitochondrial DNA and nucleoside toxicity. PMID- 12124415 TI - Male cells in female recipients of hematopoietic-cell transplants. PMID- 12124416 TI - Chronic urticaria and angioedema. PMID- 12124418 TI - Hypercarotenemia. PMID- 12124417 TI - Prophylaxis against venous thromboembolism after surgery for cancer. PMID- 12124419 TI - Obtaining antibiotics without a prescription. PMID- 12124420 TI - Aortic pseudocoarctation. PMID- 12124421 TI - Mayhem of the multiple mechanisms: modelling neurodegeneration in prion disease. AB - This review examines recent attempts to advance the understanding of the mechanism by which neurones die in prion disease. Prion diseases or transmissible spongiform encephalopathies are characterized by the conversion of a normal glycoprotein, the prion protein, to a protease-resistant form that is suggested to be both the infectious agent and the cause of the rapid neurodegeneration in the disease. Death of the patient results from this widespread neuronal loss. Thus understanding the mechanism by which the abnormal form of the prion protein causes neuronal death might lead to treatments that would prevent the life threatening nature of these diseases. PMID- 12124423 TI - Intracellular distribution of the fluorescent dye nonyl acridine orange responds to the mitochondrial membrane potential: implications for assays of cardiolipin and mitochondrial mass. AB - Cardiolipin, a polyunsaturated acidic phospholipid, is found exclusively in bacterial and mitochondrial membranes where it is intimately associated with the enzyme complexes of the respiratory chain. Cardiolipin structure and concentration are central to the function of these enzyme complexes and damage to the phospholipid may have consequences for mitochondrial function. The fluorescent dye, 10 nonyl acridine orange (NAO), has been shown to bind cardiolipin in vitro and is frequently used as a stain in living cells to assay cardiolipin content. Additionally, NAO staining has been used to measure the mitochondrial content of cells as dye binding to mitochondria is reportedly independent of the membrane potential. We used confocal microscopy to examine the properties of NAO in cortical astrocytes, neonatal cardiomyocytes and in isolated brain mitochondria. We show that NAO, a lipophilic cation, stained mitochondria selectively. However, the accumulation of the dye was clearly dependent upon the mitochondrial membrane potential and depolarisation of mitochondria induced a redistribution of dye. Moreover, depolarisation of mitochondria prior to NAO staining also resulted in a reduced NAO signal. These observations demonstrate that loading and retention of NAO is dependant upon membrane potential, and that the dye cannot be used as an assay of either cardiolipin or mitochondrial mass in living cells. PMID- 12124422 TI - Native group-III metabotropic glutamate receptors are coupled to the mitogen activated protein kinase/phosphatidylinositol-3-kinase pathways. AB - We used cultured cerebellar granule cells to examine whether native group-III metabotropic glutamate (mGlu) receptors are coupled to the mitogen-activated protein kinase (MAPK) and phosphatidylinositol-3-kinase (PI-3-K) pathways. Cultured granule cells responded to the group-III mGlu receptor agonist, L-2 amino-4-phosphonobutanoate (l-AP4), with an increased phosphorylation and activity of MAPKs (ERK-1 and -2) and an increased phosphorylation of the PI-3-K target, protein kinase B (PKB/AKT). These effects were attenuated by the group III antagonists, alpha-methyl-serine-O -phosphate (MSOP) and (R,S )-alpha cyclopropyl-4-phosphonophenylglycine (CPPG), or by pretreatment of the cultures with pertussis toxin. l-AP4 also induced the nuclear translocation of beta catenin, a downstream effector of the PI-3-K pathway. To assess the functional relevance of these mechanisms we examined the ability of l-AP4 to protect granule cells against apoptosis by trophic deprivation, induced by lowering extracellular K(+) from 25 to 10 mm. Neuroprotection by l-AP4 was attenuated by MSOP and abrogated by the compounds PD98059 and UO126, which inhibit the MAPK pathway, or by the compound LY294002, which inhibits the PI-3-K pathway. Taken together, these results show for the first time that native group-III mGlu receptors are coupled to MAPK and PI-3-K, and that activation of both pathways is necessary for neuroprotection mediated by this particular class of receptors. PMID- 12124424 TI - Effects of intrahippocampal CT105, a carboxyl terminal fragment of beta-amyloid precursor protein, alone/with inflammatory cytokines on working memory in rats. AB - In this study, we examined the effects of a 105 amino acid carboxyl terminal fragment of beta-amyloid precursor protein (CT105) and inflammatory cytokines on working memory in rats, by using a three-panel runway set-up. CT105 at 10 nmol/side significantly impaired working memory when it was administered bilaterally into the hippocampus. Furthermore, to elucidate the interaction of CT105 with inflammatory cytokines, we co-administered tumor necrosis factor-alpha (TNF-alpha) and interleukin-1beta (IL-1beta) in combination with CT105. Concurrent injections of CT105 (1.0 nmol/side) and TNF-alpha (100 ng/side) produced a synergistic deficit of working memory, whereas IL-1beta (100 ng/side) combined with CT105 (1.0 nmol/side) did not affect the working memory performance. These results indicate that the CT105-induced impairment of working memory is strongly aggravated by an increase in the level of the inflammatory cytokine TNF-alpha, which may occur in the brains of patients with Alzheimer's disease. PMID- 12124425 TI - An oxidative stress-mediated positive-feedback iron uptake loop in neuronal cells. AB - Intracellular reactive iron is a source of free radicals and a possible cause of cell damage. In this study, we analyzed the changes in iron homeostasis generated by iron accumulation in neuroblastoma (N2A) cells and hippocampal neurons. Increasing concentrations of iron in the culture medium elicited increasing amounts of intracellular iron and of the reactive iron pool. The cells had both IRP1 and IRP2 activities, being IRP1 activity quantitatively predominant. When iron in the culture medium increased from 1 to 40 microm, IRP2 activity decreased to nil. In contrast, IRP1 activity decreased when iron increased up to 20 microm, and then, unexpectedly, increased. IRP1 activity at iron concentrations above 20 microm was functional as it correlated with increased (55) Fe uptake. The increase in IRP1 activity was mediated by oxidative-stress as it was largely abolished by N-acetyl-L-cysteine. Culturing cells with iron resulted in proteins and DNA modifications. In summary, iron uptake by N2A cells and hippocampus neurons did not shut off at high iron concentrations in the culture media. As a consequence, iron accumulated and generated oxidative damage. This behavior is probably a consequence of the paradoxical activation of IRP1 at high iron concentrations, a condition that may underlie some processes associated with neuronal degeneration and death. PMID- 12124427 TI - Effects of systemic and central nervous system localized inflammation on the contributions of metabolic precursors to the L-kynurenine and quinolinic acid pools in brain. AB - L-Kynurenine and quinolinic acid are neuroactive L-tryptophan-kynurenine pathway metabolites of potential importance in pathogenesis and treatment of neurologic disease. To identify precursors of these metabolites in brain, [(2)H(3) ]-L kynurenine was infused subcutaneously by osmotic pump into three groups of gerbils: controls, CNS-localized immune-activated, and systemically immune activated. The specific activity of L-kynurenine and quinolinate in blood, brain and systemic tissues at equilibrium was then quantified by mass spectrometry and the results applied to a model of metabolism to differentiate the relative contributions of various metabolic precursors. In control gerbils, 22% of L kynurenine in brain was derived via local synthesis from L tryptophan/formylkynurenine versus 78% from L-kynurenine from blood. Quinolinate in brain was derived from several sources, including: local tissue L tryptophan/formylkynurenine (10%), blood L-kynurenine (35%), blood 3 hydroxykynurenine/3-hydroxyanthranilate (7%), and blood quinolinate (48%). After systemic immune-activation, however, L-kynurenine in brain was derived exclusively from blood, whereas quinolinate in brain was derived from three sources: blood L-kynurenine (52%), blood 3-hydroxykynurenine or 3 hydroxyanthranilate (8%), and blood quinolinate (40%). During CNS-localized immune activation, > 98% of both L-kynurenine and quinolinate were derived via local synthesis in brain. Thus, immune activation and its site determine the sources from which L-kynurenine and quinolinate are synthesized in brain. Successful therapeutic modulation of their concentrations must take into account the metabolic and compartment sources. PMID- 12124426 TI - Expression of CD47/integrin-associated protein induces death of cultured cerebral cortical neurons. AB - The death and survival of neuronal cells are regulated by various signaling pathways during development of the brain and in neuronal diseases. Previously, we demonstrated that the neuronal adhesion molecule brain immunoglobulin-like molecule with tyrosine-based activation motifs/SHP substrate 1 (BIT/SHPS-1) is involved in brain-derived neurotrophic factor (BDNF)-promoted neuronal cell survival. Here, we report the apoptosis-inducing effect of CD47/integrin associated protein (IAP), the heterophilic binding partner of BIT/SHPS-1, on neuronal cells. We generated a recombinant adenovirus vector expressing a neuronal form of CD47/IAP, and found that the expression of CD47/IAP by infection with CD47/IAP adenovirus induced the death of cultured cerebral cortical neurons. The numbers of TdT-mediated biotin-dUTP nick-end labelling (TUNEL)-positive neurons and of cells displaying apoptotic nuclei increased by expression of CD47/IAP. Neuronal cell death was prevented by the addition of the broad-spectrum caspase inhibitor Z-VAD-fmk. Furthermore, we observed that co-expression of CD47/IAP with BIT/SHPS-1 enhanced neuronal cell death, and that BDNF prevented it. These results suggest that CD47/IAP is involved in a novel pathway which regulates caspase-dependent apoptosis of cultured cerebral cortical neurons. CD47/IAP-induced death of cultured cortical neurons may be regulated by the interaction of CD47/IAP with BIT/SHPS-1 and by BDNF. PMID- 12124428 TI - Different pathways for iNOS-mediated toxicity in vitro dependent on neuronal maturation and NMDA receptor expression. AB - Co-localization of activated microglia and damaged neurones seen in brain injury suggests microglia-induced neurodegeneration. Activated microglia release two potential neurotoxins, excitatory amino acids and nitric oxide (NO), but their contribution to mechanisms of injury is poorly understood. Using co-cultures of rat microglia and embryonic cortical neurones, we show that inducible NO synthase (iNOS)-derived NO aloneis responsible for neuronal death from interferon gamma (IFNgamma) +lipopolysaccharide (LPS)-activated microglia. Neurones remain sensitive to NO irrespective of maturation state but, whereas blocking NMDA receptor activation with MK801 has no effect on NO-mediated toxicity to immature neurones, MK801 rescues 60-70% of neurones matured in culture for 12 days. Neuronal expression of NMDA receptors increases with maturation in culture, accounting for increased susceptibility to excitotoxins seen in more mature cultures. We show that MK801 delays the death of more mature neurones caused by the NO-donor DETA/NO indicating that NO elicits an excitotoxic mechanism, most likely through neuronal glutamate release. Thus, similar concentrations of nitric oxide cause neuronal death by two distinct mechanisms: NO acts directly upon immature neurones but indirectly, via NMDA receptors, on more mature neurones. Our results therefore extend existing evidence for NO-mediated toxicity and show a complex interaction between inflammatory and excitotoxic mechanisms of injury in mature neurones. PMID- 12124429 TI - Caspase cleavage of members of the amyloid precursor family of proteins. AB - The synapse loss and neuronal cell death characteristic of Alzheimer's disease (AD) are believed to result in large part from the neurotoxic effects of beta amyloid peptide (Abeta), a 40-42 amino acid peptide(s) derived proteolytically from beta-amyloid precursor protein (APP). However, APP is also cleaved intracellularly to generate a second cytotoxic peptide, C31, and this cleavage event occurs in vivo as well as in vitro and preferentially in the brains of AD patients (Lu et al. 2000). Here we show that APPC31 is toxic to neurons in primary culture, and that like APP, the APP family members APLP1 and possibly APLP2 are cleaved by caspases at their C-termini. The carboxy-terminal peptide derived from caspase cleavage of APLP1 shows a degree of neurotoxicity comparable to APPC31. Our results suggest that even though APLP1 and APLP2 cannot generate Abeta, they may potentially contribute to the pathology of AD by generating peptide fragments whose toxicity is comparable to that of APPC31. PMID- 12124430 TI - Dynamics of tyrosine hydroxylase promoter activity during midbrain dopaminergic neuron development. AB - Dopamine (DA)-producing neurons in the ventral midbrain are generated from a specified neuronal lineage and form selective axonal pathways that mediate multiple CNS functions. Expression of the gene encoding tyrosine hydroxylase (TH), which is a key enzyme of catecholamine biosynthesis, is regulated during the development of midbrain DA neurons. In the present study, we report the developmental regulation and cell type specificity of TH gene promoter in the ventral midbrain by using a green fluorescent protein (GFP) reporter system. Transgenic mice were generated that express GFP in the majority of midbrain DA neurons under the control of the 9-kb upstream region of the rat TH gene. At an early embryonic stage, GFP expression was induced in the developing DA neurons, and the expression was then markedly down-regulated at later embryonic stages. However, the expression was reactivated and approached the adult levels during early post-natal development. These developmental changes in GFP expression patterns suggest the presence of multistep regulatory mechanisms for TH gene expression during DA neuron development. The TH promoter appears to possess transcriptional elements at least necessary for the induction of TH expression at the early embryonic stage and its reactivation during the post-natal development. PMID- 12124431 TI - Generation of reactive oxygen species and activation of NF-kappaB by non-Abeta component of Alzheimer's disease amyloid. AB - Non-amyloid beta (Abeta) component of Alzheimer's disease (AD) amyloid (NAC) coexists with Abeta protein in senile plaques. After exposure to NAC fibrils, cortical neurons of rat brain primary culture became apoptotic, while astrocytes were activated with extension of their processes. NAC fibrils decreased the activity of reducing 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) in cortical neurons more markedly (IC(50) = 5.6 microm) than in astrocytes (IC(50) approximately 50 microm). The neuron-specific toxicity of NAC fibrils was indicated also by an increased release of lactate dehydrogenase from the cells. Neuronal apoptosis was suppressed by pre-treatment with the antioxidants, propyl gallate (PG) and N-t-butyl-phenylnitrone (BPN), or overexpression of human Bcl-2. Exposure to NAC fibrils enhanced generation of reactive oxygen species (ROS) in neurons and less efficiently in astrocytes, as demonstrated by oxidation of 2',7' dichlorofluorescin. The site of ROS generation was shown to be mitochondria by oxidation of chloromethyl-tetramethyl rosamine. Exposure to NAC fibrils increased also the nuclear translocation of nuclear factor kappa B (NF-kappaB) and enhanced its DNA-binding activity, which was inhibited by PG and BPN more efficiently in neurons than in astrocytes. These results suggest that NAC fibrils increase mitochondrial ROS generation and activate NF-kappaB, thereby causing a differential change in gene expression between neurons and astrocytes in the AD brain. PMID- 12124432 TI - Activation of nuclear Ca(2+)/calmodulin-dependent protein kinase II and brain derived neurotrophic factor gene expression by stimulation of dopamine D2 receptor in transfected NG108-15 cells. AB - We identified the isoforms of Ca(2+) /calmodulin-dependent protein kinase II (CaM kinase II) subunits in rat striatum. All four subunits of CaM kinase II alpha, beta, gamma and delta were detected including the isoforms of alphaB, gammaA, gammaA', gammaA.B, delta3 and delta7 with nuclear localization signal. We established NG108-15 cells with the stably expressed dopamine D2L receptor (D2LR, long form), which is an alternative splicing variant. The cells were termed NGD2L. Immunostaining demonstrated that D2LR was localized in plasma membranes. Calcium imaging with fluo-3 AM revealed that quinpirole, a D2R agonist, increased the intracellular Ca(2+), which was blocked by treatment with sulpiride and pertussis toxin in NGD2L cells, but not in mock cells. Furthermore, stimulation of D2LR with quinpirole in NGD2L cells activated the nuclear isoform of CaM kinase II. Stimulation of D2LR increased the expression of exon III- and IV-BDNF mRNA. Overexpression of CaM kinase II delta3 increased exon IV- but not exon III BDNF mRNA. These results suggest that D2R is involved in the activation of the nuclear isoform of CaM kinase II and thereby in stimulation of gene expression through Ca(2+) signaling. PMID- 12124433 TI - Aspirin provides cyclin-dependent kinase 5-dependent protection against subsequent hypoxia/reoxygenation damage in culture. AB - Aspirin [acetylsalicylic acid (ASA)] is an anti-inflammatory drug that protects against cellular injury by inhibiting cyclooxygenases (COX), inducible nitric oxide synthase (iNOS) and p44/42 mitogen-activated protein kinase (p44/42 MAPK), or by preventing translocation of nuclear factor kappaB (NF-kappaB). We studied the effect of ASA pre-treatment on neuronal survival after hypoxia/reoxygenation damage in rat spinal cord (SC) cultures. In this injury model, COX, iNOS and NF kappaB played no role in the early neuronal death. A 20-h treatment with 3 mm ASA prior to hypoxia/reoxygenation blocked the hypoxia/reoxygenation-induced lactate dehydrogenase (LDH) release from neurons. This neuroprotection was associated with increased phosphorylation of neurofilaments, which are substrates of p44/42 MAPK and cyclin-dependent kinase 5 (Cdk5). PD90859, a p44/42 MAPK inhibitor, had no effect on ASA-induced tolerance, but olomoucine and roscovitine, Cdk5 inhibitors, reduced ASA neuroprotection. Hypoxia/reoxygenation alone reduced both the protein amount and activity of Cdk5, and this reduction was inhibited by pre treatment with ASA. Moreover, the protein amount of a neuronal Cdk5 activator, p35, recovered after reoxygenation only in ASA-treated samples. The prevention of the loss in Cdk5 activity during reoxygenation was crucial for ASA-induced protection, because co-administration of Cdk5 inhibitors at the onset ofreoxygenation abolished the protection. In conclusion, pre-treatment with ASA induces tolerance against hypoxia/reoxygenation damage in spinal cord cultures by restoring Cdk5 and p35 protein expression. PMID- 12124434 TI - Alternative non-coding exons support serotonin transporter mRNA expression in the brain and gut. AB - A number of studies in recent years have linked polymorphisms within the serotonin transporter (5HTT) gene to affective disorders and anxiety traits. The human 5HTT mRNA is alternatively spliced, and the splice variants are equally expressed in the human placental cell line and dorsal raphe. In this study, using 5' rapid amplification of cDNA ends, we show that the rat 5HTT mRNA is alternatively spliced, leading to three distinct mRNAs differing in the 5' untranslated region. To determine whether the three alternatively spliced mRNA species that contain one of the following untranslated regions (i) exon 1A, 63 bp (ii) exon 1A + 1B, 125 bp or (iii) exon 1C, 101 bp, were expressed in a tissue specific manner, we used RT-PCR and exon-specific oligonucleotide hybridization. Our results suggest two of the variants (1A + 1B and 1A) may utilize the same promoter; however, they are not equally expressed. While in the adult CNS and adrenal medulla, the shorter mRNA consisting of exon 1A was considerably more abundant, in the stomach and heart, the two variants were equally expressed. The third splice variant exon 1C is only expressed in the gut and to a lesser extent in the heart. The data from this study suggest the splice variant consisting of exon 1C may utilize a distinct promoter compared to the other two. PMID- 12124435 TI - Mitochondrial voltage-dependent anion channel is involved in dopamine-induced apoptosis. AB - Neuronal NMB cells were used to determine changes in gene expression upon treatment with dopamine. Twelve differentially expressed cDNAs were identified and cloned, one of them having 99.4% sequence homology with isoform 2 of a voltage-dependent anion channel (VDAC-2). The known role of VDAC, a mitochondrial outer-membrane protein, in transport of anions, pore formation, and release of cytochrome C prompted us to investigate the possible role of VDAC gene family in dopamine-induced apoptosis. Semi-quantitative PCR analysis indicated that expression of the three VDAC isoforms was reduced by dopamine. Immunoblotting with anti-VDAC antibodies detected two VDAC protein bands of 33 and 34 kDa. Dopamine decreased differentially the immunoreactivity of the 34 kDa protein. Whether the decrease in VDAC expression influence the mitochondrial membrane potential (Delta(Psi)(m)) was determined with the dye Rhodamine-123. Dopamine indeed decreased the mitochondrial Delta(Psi)(m), but the maximum effect was observed within 3 h, prior to the decrease in VDAC mRNA or protein levels. Cyclosporin A, a blocker of the mitochondrial pore complex, prevented the decrease in Delta(Psi)(m), but did not rescue the cells from dopamine toxicity. To elucidate possible involvement of protease caspases in dopamine-induced apoptosis, the effect of the caspase inhibitor z-Val-Ala-Asp(Ome)-FMK (zVAD) was determined. zVAD decreased dopamine toxicity, yet it did not rescue the mitochondrial Delta(Psi)(m) drop. Dopamine also decreased ATP levels. Finally, transfection of NMB cells with pcDNA-VDAC decreased the cytotoxic effect of dopamine. These findings are in agreement with the notion that the mitochondria, and VDAC, are important participants in dopamine-induced apoptosis. PMID- 12124436 TI - Shp-2 positively regulates brain-derived neurotrophic factor-promoted survival of cultured ventral mesencephalic dopaminergic neurons through a brain immunoglobulin-like molecule with tyrosine-based activation motifs/Shp substrate 1. AB - To examine the roles of Shp-2, a cytoplasmic tyrosine phosphatase, in neuronal survival, we generated and used recombinant adenoviruses expressing wild type and phosphatase-inactive (C/S), phosphatase domain-deficient (delta P) and constitutively active (D61A and E76A) mutants of Shp-2. We found that wild-type Shp-2 enhanced brain-derived neurotrophic factor (BDNF)-promoted survival of cultured ventral mesencephalic dopaminergic neurons. In contrast, the C/S and delta P mutants of Shp-2 did not affect survival. In addition, the constitutively active D61A and E76A mutants mimicked BDNF and promoted survival. Furthermore, to examine the effects of BIT/SHPS-1, a substrate of Shp-2, on the BDNF-promoted survival, we generated adenovirus vectors expressing wild-type BIT/SHPS-1 and its 4F mutant in which all tyrosine residues in the cytoplasmic domain of BIT/SHPS-1 were replaced with phenylalanine. We found that BDNF-promoted survival of cultured mesencephalic dopaminergic neurons was enhanced by expression of the 4F mutant but not of wild-type BIT/SHPS-1. In addition, we found that co-expression of wild-type BIT/SHPS-1 with Shp-2 significantly enhanced the survival-promoting effect of BDNF on cultured mesencephalic dopaminergic neurons. These results indicated that Shp-2 positively regulates the survival-promoting effect of BDNF on cultured ventral mesencephalic dopaminergic neurons. Dephosphorylation of BIT/SHPS-1 by Shp-2 may participate in BDNF-stimulated survival signaling. PMID- 12124437 TI - Temporal patterns of cytokine and apoptosis-related gene expression in spinal cords of the G93A-SOD1 mouse model of amyotrophic lateral sclerosis. AB - Familial amyotrophic lateral sclerosis (FALS) is often caused by gain-of-function mutations in Cu,Zn-superoxide dismutase (SOD1). Multiprobe ribonuclease protection assays (RPAs) were used to investigate expression of 36 different cytokines and apoptosis-related genes in spinal cords of mice that ubiquitously express human SOD1 bearing a glycine (r) alanine substitution at residue 93 (G93A SOD1). Mice were studied at late presymptomatic stage (80 days), and at 120 days when the animals experience severe hindlimb paralysis and accumulation of oxidatively modified proteins. Spinal cord tissue from G93A-SOD1 mice expressed a selective subset of macrophage-typical cytokines (monokines) including interleukin (IL)1alpha, IL1beta and IL1RA at 80 days increasing by 120 days. Contrastingly, T-cell derived cytokines (lymphokines) including IL2, IL3 and IL4 were detected at low levels in non-transgenic mice but these were not elevated in G93A-SOD1 mice even at 120 days. Apoptosis-related genes were generally unaffected at 80 days but multiple caspases and death receptor components were up regulated at 120 days; the only exceptions being FADD and the tumor necrosis factor (TNF)alpha receptor p55 which was up-regulated at 80 days and increased further at 120 days. These data indicate that in the G93A-SOD1 mouse: (i) cytokine expression changes precede bulk protein oxidation and apoptosis gene expression; (ii) lymphocyte contributions to cytokine expression in FALS are likely minor; and (iii) TNFalpha and its receptors may link inflammation to apoptosis in ALS. PMID- 12124438 TI - Oxidative damage increases with age in a canine model of human brain aging. AB - We assayed levels of lipid peroxidation, protein carbonyl formation, glutamine synthetase (GS) activity and both oxidized and reduced glutathione to study the link between oxidative damage, aging and beta-amyloid (Abeta) in the canine brain. The aged canine brain, a model of human brain aging, naturally develops extensive diffuse deposits of human-type Abeta. Abeta was measured in immunostained prefrontal cortex from 19 beagle dogs (4-15 years). Increased malondialdehyde (MDA), which indicates increased lipid peroxidation, was observed in the prefrontal cortex and serum but not in cerebrospinal fluid (CSF). Oxidative damage to proteins (carbonyl formation) also increased in brain. An age dependent decline in GS activity, an enzyme vulnerable to oxidative damage, and in the level of glutathione (GSH) was observed in the prefrontal cortex. MDA level in serum correlated with MDA accumulation in the prefrontal cortex. Although 11/19 animals exhibited Abeta, the extent of deposition did not correlate with any of the oxidative damage measures, suggesting that each form of neuropathology accumulates in parallel with age. This evidence of widespread oxidative damage and Abeta deposition is further justification for using the canine model for studying human brain aging and neurodegenerative diseases. PMID- 12124439 TI - Acute administration of cocaine regulates the phosphorylation of serine-19, -31 and -40 in tyrosine hydroxylase. AB - Acute cocaine can inhibit catecholamine biosynthesis by regulating the enzymatic activity of tyrosine hydroxylase via alterations in the phosphorylation state of the enzyme. The mechanisms underlying acute cocaine-dependent regulation of tyrosine hydroxylase phosphorylation have not been determined. In this study, 0, 15 or 30 mg/kg cocaine was administered intraperitoneally to rats and the phosphorylation state of tyrosine hydroxylase in the brain was examined using antibodies specific for the phosphorylated forms of serine-19, -31 and -40 in tyrosine hydroxylase. In the caudate and nucleus accumbens, cocaine dose dependently decreased the levels of phosphorylated serine-19, -31 and -40. In the ventral tegmental area, the levels of phosphorylated serine-19, but not serine-31 and -40, were decreased by 15 and 30 mg/kg cocaine. In the amygdala, the levels of phosphorylated serine-19, but not serine-31 or -40, were decreased. The functional effects of these alterations in phosphorylation state were assessed by measuring tyrosine hydroxylase activity in vivo (accumulation of DOPA after administration of the decarboxylase inhibitor NSD-1015). Acute administration of 30 mg/kg cocaine significantly decreased l-DOPA production in caudate and accumbens but not in amygdala. These data suggest that the phosphorylation of serine-31 or -40, but not serine-19, is involved in the regulation of tyrosine hydroxylase activity by acute cocaine. PMID- 12124440 TI - Muscarinic receptor-mediated phosphorylation of cyclic AMP response element binding protein in human neuroblastoma cells. AB - This study describes the effect of signalling through muscarinic acetylcholine receptors on two transcription factors implicated in long-term synaptic plasticity and memory formation, EGR1 and the cyclic AMP response element binding protein (CREB). In SK-N-SH neuroblastoma cells, treatment with the cholinergic agonist carbachol led to maximal induction of EGR1 1 h after stimulation. This was preceded by the phosphorylation of CREB, which peaked as early as 5 minutes after carbachol treatment. The levels of both EGR1 and phosphorylated CREB (pCREB) slowly decayed over 4-8 h. CREB phosphorylation and EGR1 induction showed similar sensitivity to carbachol concentration, with EC(50) values in the range of 1-10 microM, and the changes in both transcription factors were blocked by the muscarinic antagonist atropine. As has been described elsewhere, EGR1 induction was dependent on activation of p42/44 MAP kinase, as it was blocked by the MEK inhibitor U0126. However, CREB phosphorylation by carbachol was largely unaffected by MAP kinase blockade. As both CREB phosphorylation and EGR1 induction have been linked to long-term potentiation and some forms of memory consolidation, these results may implicate CREB and EGR1 in independent or partially independent cholinergic signalling pathways involved in memory processes. PMID- 12124441 TI - AMPA receptor-mediated toxicity in oligodendrocyte progenitors involves free radical generation and activation of JNK, calpain and caspase 3. AB - The molecular mechanisms underlying AMPA (alpha-amino-3-hydroxy-5-methylisoxazole 4-propionate) receptor-mediated excitotoxicity were characterized in rat oligodendrocyte progenitor cultures. Activation of AMPA receptors, in the presence of cyclothiazide to selectively block desensitization, produced a massive Ca(2+) influx and cytotoxicity which were blocked by the antagonists CNQX and GYKI 52466. A role for free radical generation in oligodendrocyte progenitor cell death was deduced from three observations: (i) treatment with AMPA agonists decreased intracellular glutathione; (ii) depletion of intracellular glutathione with buthionine sulfoximine potentiated cell death; and (iii) the antioxidant N acetylcysteine replenished intracellular glutathione and protected cultures from AMPA receptor-mediated toxicity. Cell death displayed some characteristics of apoptosis, including DNA fragmentation, chromatin condensation and activation of caspase-3 and c-Jun N-terminal kinase (JNK). A substrate of calpain and caspase 3, alpha-spectrin, was cleaved into characteristic products following treatment with AMPA agonists. In contrast, inhibition of either caspase-3 by DEVD-CHO or calpain by PD 150606 protected cells from excitotoxicity. Our results indicate that overactivation of AMPA receptors causes apoptosis in oligodendrocyte progenitors through mechanisms involving Ca(2+) influx, depletion of glutathione, and activation of JNK, calpain, and caspase-3. PMID- 12124442 TI - Cerebral dicarboxylate transport and metabolism studied with isotopically labelled fumarate, malate and malonate. AB - Transport and metabolism of dicarboxylates may be important in the glial-neuronal metabolic interplay. Further, exogenous dicarboxylates have been suggested as cerebral energy substrates. After intrastriatal injection of [(14) C]fumarate or [(14) C]malate, glutamine attained a specific activity 4.1 and 2.6 times higher than that of glutamate, respectively, indicating predominantly glial uptake of these four-carbon dicarboxylates. In contrast, the three-carbon dicarboxylate [(14) C]malonate gave a specific activity in glutamate which was approximately five times higher than that of glutamine, indicating neuronal uptake of malonate. Therefore, neurones and glia take up different types of dicarboxylates, probably by different transport mechanisms. Labelling of alanine from [(14) C]fumarate and [(14) C]malate demonstrated extensive malate decarboxylation, presumably in glia. Intravenous injection of 75 micromol [U-(13) C]fumarate rapidly led to high concentrations of [U-(13) C]fumarate and [U-(13) C]malate in serum, but neither substrate labelled cerebral metabolites as determined by (13) C NMR spectroscopy. Only after conversion of [U-(13) C]fumarate into serum glucose was there (13) C labelling of cerebral metabolites, and only at <10% of that obtained with 75 micromol [3-(13) C]lactate or [2-(13) C]acetate. These findings suggest a very low transport capacity for four-carbon dicarboxylates across the blood-brain barrier and rule out a role for exogenous fumarate as a cerebral energy substrate. PMID- 12124443 TI - Peroxynitrite affects exocytosis and SNARE complex formation and induces tyrosine nitration of synaptic proteins. AB - The reactive species peroxynitrite, formed via the near diffusion-limited reaction of nitric oxide and superoxide anion, is a potent oxidant that contributes to tissue damage in neurodegenerative disorders. Peroxynitrite readily nitrates tyrosine residues in proteins, producing a permanent modification that can be immunologically detected. We have previously demonstrated that in the nerve terminal, nitrotyrosine immunoreactivity is primarily associated with synaptophysin. Here we identify two other presynaptic proteins nitrated by peroxynitrite, Munc-18 and SNAP25, both of which are involved in sequential steps leading to vesicle exocytosis. To investigate whether peroxynitrite affects vesicle exocytosis, we used the fluorescent dye FM1 43 to label a recycling population of secretory vesicles within the synaptosomes. Bolus addition of peroxynitrite stimulated exocytosis and glutamate release. Notably, these effects were strongly reduced in the presence of NaHCO(3), indicating that peroxynitrite acts mainly intracellularly. Furthermore, peroxynitrite enhanced the formation of the sodium dodecyl sulfate-resistant SNARE complex in a dose-dependent manner (100-1000 microm) and induced the formation of 3-nitrotyrosine in proteins of SNARE complex. These data suggest that modification(s) of synaptic vesicle proteins induced by peroxynitrite may affect protein-protein interactions in the docking/fusion steps, thus promoting exocytosis, and that, under excessive production of superoxide and nitric oxide, neurons may up-regulate neuronal signaling. PMID- 12124444 TI - Insulin-like growth factor (IGF)-I binding to a cell membrane associated IGF binding protein-3 acid-labile subunit complex in human anterior pituitary gland. AB - The binding characteristics of [(125) I]insulin-like growth factor (IGF)-I were studied in human brain and pituitary gland. Competition binding studies with DES(1-3)IGF-I and R(3) -IGF-I, which display high affinity for the IGF-I receptor and low affinity for IGF binding proteins (IGFBPs), were performed to distinguish [(125) I]IGF-I binding to IGF-I receptors and IGFBPs. Specific [(125) I]IGF-I binding in brain regions and the posterior pituitary was completely displaced by DES(1-3)IGF-I and R(3) -IGF-I, indicating binding to IGF-I receptors. In contrast, [(125) I]IGF-I binding in the anterior pituitary was not displaced by DES(1-3)IGF-I and R(3) -IGF-I, suggesting binding to an IGF-binding site that is different from the IGF-I receptor. Binding affinity of IGF-I to this site was about 10-fold lower than for the IGF-I receptor. Using western immunoblotting we were also unable to detect IGF-I receptors in human anterior pituitary. Instead, western immunoblotting and immunoprecipitation experiments showed a 150-kDa IGFBP 3-acid labile subunit (ALS) complex in the anterior pituitary and not in the posterior pituitary and other brain regions. RT-PCR experiments showed the expression of ALS mRNA in human anterior pituitary indicating that the anterior pituitary synthesizes ALS. In the brain regions and posterior pituitary, IGFBP-3 was easily washed away during pre-incubation procedures as used in the [(125) I]IGF-I binding experiments. In contrast, the IGFBP-3 complex in the anterior pituitary could not be removed by these washing procedures. Our results indicate that the human anterior pituitary contains a not previously described tightly cell membrane-bound 150-kDa IGFBP-3-ALS complex that is absent in brain and posterior pituitary. PMID- 12124445 TI - Identification of three cAMP-dependent protein kinase (PKA) phosphorylation sites within the major intracellular domain of neuronal nicotinic receptor alpha4 subunits. AB - This study determined whether all protein kinase A (PKA) and protein kinase C (PKC) phosphorylation sites on the alpha4 subunit of rat alpha4beta2 neuronal nicotinic receptors could be localized to the M3/M4 cytoplasmic domain of the protein, and investigated specific amino acid substrates for the kinases through two-dimensional phosphopeptide mapping and site-directed mutagenesis. Experiments were conducted using alpha4beta2 receptors expressed in Xenopus oocytes and a fusion protein corresponding to the M3/M4 cytoplasmic domain of alpha4 (alpha4(333-594) ). When oocytes expressing alpha4beta2 receptors were incubated with [(32) P]orthophosphate in order to label endogenous ATP stores, phosphorylation of alpha4 subunits was evident. Incubation of either immunoprecipitated receptors or the fusion protein with [(32) P]ATP and either PKA or PKC followed by trypsinization of the samples demonstrated that the kinases phosphorylated alpha4 subunits on multiple phosphopeptides, and that the phosphorylated full-length alpha4 protein and fusion protein produced identical phosphopeptide maps. Site-directed mutagenesis of Ser365, Ser472 and Ser491 to alanines in the fusion protein eliminated phosphopeptides phosphorylated by PKA, but not by PKC. Other mutations investigated, Ser470, Ser493, Ser517 and Ser590, did not alter the phosphopeptide maps. Results indicate that Ser365, Ser472 and Ser491 on neuronal nicotinic receptor alpha4 subunits are phosphorylated by PKA and are likely to represent post-translational regulatory sites on the receptor. PMID- 12124446 TI - Cell surface receptors, virus entry and tropism of primate lentiviruses. AB - Human immunodeficiency virus (HIV) exploits cell surface receptors to attach to and gain entry into cells. The HIV envelope spike glycoprotein on the surface of virus particles binds both CD4 and a seven-transmembrane coreceptor. These interactions trigger conformational changes in the envelope spike that induce fusion of viral and cellular membranes and entry of the viral core into the cell cytoplasm. Other cell surface receptors also interact with gp120 and aid attachment of virus particles. This review describes these receptors, their roles in HIV entry and their influence on cell tropism. PMID- 12124447 TI - Significant differences in nucleocapsid morphology within the Paramyxoviridae. AB - Nucleocapsid (N) proteins from representative viruses of three genera within the Paramyxoviridae were expressed in insect cells using recombinant baculoviruses. RNA-containing structures, which appear morphologically identical to viral nucleocapsids, were isolated and subsequently imaged under a transmission electron microscope. Analysis of these images revealed marked differences in nucleocapsid morphology among the genera investigated, most notably between viruses of the Paramyxovirinae and the Pneumovirinae subfamilies. Helical pitch measurements were made, revealing that measles virus (MV, a Morbillivirus within the subfamily Paramyxovirinae) N protein produces helices that adopt multiple conformations with varying degrees of flexibility, while that of the Rubulavirus simian virus type 5 (SV5, subfamily Paramyxovirinae) produces more rigid structures with a less heterogeneous pitch distribution. Nucleocapsids produced by respiratory syncytial virus (RSV, subfamily Pneumovirinae) appear significantly narrower than those of MV and SV5 and have a longer pitch than the most extended form of MV. In addition to helical nucleocapsids, ring structures were also produced, image analysis of which has demonstrated that rings assembled from MV N protein consist of 13 subunits. This is consistent with previous reports that Sendai virus nucleocapsids have 13.07 subunits per turn. It was determined, however, that SV5 subnucleocapsid rings have 14 subunits, while rings derived from the radically different RSV nucleocapsid have been found to contain predominantly 10 subunits. PMID- 12124448 TI - Respiratory syncytial virus assembly occurs in GM1-rich regions of the host-cell membrane and alters the cellular distribution of tyrosine phosphorylated caveolin 1. AB - We have previously shown that respiratory syncytial virus (RSV) assembly occurs within regions of the host-cell surface membrane that are enriched in the protein caveolin-1 (cav-1). In this report, we have employed immunofluorescence microscopy to further examine the RSV assembly process. Our results show that RSV matures at regions of the cell surface that, in addition to cav-1, are enriched in the lipid-raft ganglioside GM1. Furthermore, a comparison of mock-infected and RSV-infected cells by confocal microscopy revealed a significant change in the cellular distribution of phosphocaveolin-1 (pcav-1). In mock-infected cells, pcav 1 was located at regions of the cell that interact with the extracellular matrix, termed focal adhesions (FA). In contrast, RSV-infected cells showed both a decrease in the levels of pcav-1 associated with FA and the appearance of pcav-1 containing cytoplasmic vesicles, the latter being absent in mock-infected cells. These cytoplasmic vesicles were clearly visible between 9 and 18 h post-infection and coincided with the formation of RSV filaments, although we did not observe a direct association of pcav-1 with mature virus. In addition, we noted a strong colocalization between pcav-1 and growth hormone receptor binding protein-7 (Grb7), within these cytoplasmic vesicles, which was not observed in mock infected cells. Collectively, these findings show that the RSV assembly process occurs within specialized lipid-raft structures on the host-cell plasma membrane, induces the cellular redistribution of pcav-1 and results in the formation of cytoplasmic vesicles that contain both pcav-1 and Grb7. PMID- 12124449 TI - Vaccination of pigs with a DNA construct expressing an influenza virus M2 nucleoprotein fusion protein exacerbates disease after challenge with influenza A virus. AB - In mice, vaccines inducing antibodies to the extracellular domain of the M2 protein (M2e) can confer protection to influenza A virus infection. Unlike the surface glycoproteins, haemagglutinin and neuraminidase, this domain of M2 is highly conserved and is therefore a potential broad-spectrum immunogen. In this study, the protection conferred by vaccines inducing antibodies to M2e was evaluated in a challenge model for swine influenza in pigs. A protein resulting from the fusion between M2e and the hepatitis B virus core protein (M2eHBc), with or without adjuvant, was evaluated. In addition, a DNA construct expressing a fusion protein between M2e and influenza virus nucleoprotein (M2eNP) was evaluated to see if the broad-spectrum protection conferred by antibodies could be further enhanced by T helper cells and cytotoxic T cells. All vaccines induced an antibody response against M2e, and the M2eNP DNA vaccine additionally induced an influenza virus-specific lymphoproliferation response. However, after challenge with a swine influenza virus (H1N1), no protection was observed in the vaccinated groups compared with the non-vaccinated control group. On the contrary, vaccinated pigs showed more severe clinical signs than the control pigs. The M2eNP DNA-vaccinated pigs showed the most severe clinical signs and three out of six pigs died on days 1 and 2 post-challenge. These results indicate that antibodies to M2e, especially in combination with cell-mediated immune responses, exacerbate disease. Thus, clinical signs after infection should be observed closely in further studies using M2e as an immunogen and caution should be exercised in using M2e in humans. PMID- 12124450 TI - Identification of radically different variants of porcine reproductive and respiratory syndrome virus in Eastern Europe: towards a common ancestor for European and American viruses. AB - We determined 22 partial porcine reproductive and respiratory syndrome virus (PRRSV) ORF5 sequences, representing pathogenic field strains mainly from Poland and Lithuania, and two currently available European-type live PRRSV vaccines. Also, the complete ORF7 of two Lithuanian and two Polish strains was sequenced. We found that Polish, and in particular Lithuanian, PRRSV sequences were exceptionally different from the European prototype, the Lelystad virus, and in addition showed a very high national diversity. The most diverse present-day European-type PRRSV sequences were from Poland (2000) and Lithuania (2000), and exhibited only 72.2% nucleotide identity in the investigated ORF5 sequence. While all sequences determined in the present study were clearly of European type, inclusion of the new Lithuanian sequences in the genealogy resulted in a common ancestor for the European type virus significantly closer to the American-type PRRSV than previously seen. In addition, the length of the ORF7 of the Lithuanian strains was 378 nucleotides, and thus intermediate between the sizes of the prototypical EU-type (387 nucleotides) and US-type (372 nucleotides) ORF7 lengths. These findings for the Lithuanian PRRSV sequences provide support for the hypothesis that the EU and US genotypes of PRRSV evolved from a common ancestor. Also, this is the first report of ORF7 protein size polymorphism in field isolates of EU-type PRRSV. PMID- 12124451 TI - Complete genome sequence of Montana Myotis leukoencephalitis virus, phylogenetic analysis and comparative study of the 3' untranslated region of flaviviruses with no known vector. AB - Montana Myotis leukoencephalitis virus (MMLV), a virus isolated from bats, causes an encephalitis in small rodents reminiscent of flavivirus encephalitis in humans. The complete MMLV genome is 10690 nucleotides long and encodes a putative polyprotein of 3374 amino acids. The virus contains the same conserved motifs in genes that are believed to be interesting antiviral targets (NTPase/helicase, serine protease and RNA-dependent RNA polymerase) as flaviviruses of clinical importance. Phylogenetic analysis of the entire coding region has confirmed the classification of MMLV in the clade of the flaviviruses with no known vector (NKV) and within this clade to the Rio Bravo branch (both viruses have the bat as their vertebrate host). We have provided for the first time a comparative analysis of the RNA folding of the 3' UTR of the NKV flaviviruses (Modoc, Rio Bravo and Apoi viruses, in addition to MMLV). Structural elements in the 3' UTR that are preserved among other flaviviruses have been revealed, as well as elements that distinguish the NKV from the mosquito- and tick-borne flaviviruses. In particular, the pentanucleotide sequence 5' CACAG 3', which is conserved in all mosquito- and tick-borne flaviviruses, is replaced by the sequence 5' C(C/U)(C/U)AG 3' in the loop of the 3' long stable hairpin structure of all four NKV flaviviruses. The availability of this latter sequence motif allows us to designate a virus as either an NKV or a vector-borne flavivirus. PMID- 12124452 TI - Infection of SCID mice with Montana Myotis leukoencephalitis virus as a model for flavivirus encephalitis. AB - We have established a convenient animal model for flavivirus encephalitis using Montana Myotis leukoencephalitis virus (MMLV), a bat flavivirus. This virus has the same genomic organization, and contains the same conserved motifs in genes that encode potential antiviral targets, as flaviviruses that cause disease in man (N. Charlier et al., accompanying paper), and has a similar particle size (approximately 40 nm). MMLV replicates well in Vero cells and appears to be equally as sensitive as yellow fever virus and dengue fever virus to a selection of experimental antiviral agents. Cells infected with MMLV show dilation of the endoplasmic reticulum, a characteristic of flavivirus infection. Intraperitoneal, intranasal or direct intracerebral inoculation of SCID mice with MMLV resulted in encephalitis ultimately leading to death, whereas immunocompetent mice were refractory to either intranasal or intraperitoneal infection with MMLV. Viral RNA and/or antigens were detected in the brain and serum of MMLV-infected SCID mice, but not in any other organ examined: MMLV was detected in the olfactory lobes, the cerebral cortex, the limbic structures, the midbrain, cerebellum and medulla oblongata. Infection was confined to neurons. Treatment with the interferon alpha/beta inducer poly(I).poly(C) protected SCID mice against MMLV-induced morbidity and mortality, and this protection correlated with a reduction in infectious virus titre and viral RNA load. This validates the MMLV model for use in antiviral drug studies. The MMLV SCID model may, therefore, be attractive for the study of chemoprophylactic or chemotherapeutic strategies against flavivirus infections causing encephalitis. PMID- 12124453 TI - Suppression of Japanese encephalitis virus infection by non-steroidal anti inflammatory drugs. AB - Japanese encephalitis virus (JEV) infection generates a rapid inflammatory response including peripheral neutrophil leucocytosis and infiltration of neutrophils into extraneural tissue. The level of inflammation correlates well with the clinical outcome in Japanese encephalitis patients. Non-steroidal anti inflammatory drugs (NSAIDs), used medicinally for their analgesic and anti inflammatory properties, are being considered for prevention of cardiovascular disease and cancer, as well as for treatment of human immunodeficiency virus infection. Apart from their ability to inhibit prostaglandin synthesis, the mechanisms underlying the beneficial therapeutic effects are largely unknown. We used aspirin, indomethacin and sodium salicylate to study the role of NSAIDs in JEV propagation in vitro. We found that NSAIDs suppressed JEV propagation in neuronal and non-neuronal cells. Blockade of cyclooxygenase activity by NSAIDs caused decreased production of free radicals and prostaglandins. However, these pharmacological alterations did not seem to correlate well with the antiviral effects. When cells were treated with the mitogen-activated protein kinase (MAPK) inhibitors PD 98059 and SB 203580, salicylate lost its antiviral effect. The activation of MAPK by anisomycin mimicked the action of salicylate in suppressing JEV-induced cytotoxicity. The decreased phosphorylation of extracellular signal regulated kinase (ERK) was induced by JEV infection and the decrease in ERK was reversed by salicylate. Our data suggest that the signalling pathways of MAPK play a role in the antiviral action of salicylate. PMID- 12124454 TI - Serial passage of foot-and-mouth disease virus in sheep reveals declining levels of viraemia over time. AB - If an infectious agent is to maintain itself within a closed population by means of an unbroken serial chain of infections, it must maintain the level of infectiousness of individuals through time, or termination of the transmission chain is inevitable. One possible cause of diminution in infectiousness along serial chains of transmission may be that individuals are unable to amplify and transmit comparable levels of the infectious agent. Here, the results are reported of a novel experiment designed specifically to assess the effects of serial passage of foot-and-mouth disease virus (FMDV) in experimental groups of sheep. A virus isolate taken from an epidemic of foot-and-mouth disease (FMD) characterized by rapid fade-out of infection was passed serially through four groups of sheep housed in an isolation unit. Although it was not possible to measure individual infectiousness directly, blood virus load from infected individuals was quantified using a real-time PCR assay and used as an underlying indicator of the level of infection. The results of this assay concurred well with those of the traditional tissue-culture assay and were shown to be highly repeatable. The level of peak viraemia was shown to fall significantly with the time of infection and with passage group, both in terms of the group mean and regression analysis of individual values, suggesting that this isolate of FMDV may, under certain conditions, be unable to maintain itself indefinitely in susceptible sheep populations. The results of these experiments are discussed in terms of the epidemiology of FMD in sheep. PMID- 12124456 TI - An ex vivo murine model to study poliovirus-induced apoptosis in nerve cells. AB - Paralytic poliomyelitis results from destruction of motor neurons owing to poliovirus (PV) replication. Using a mouse model, we have previously shown that PV kills neurons of the central nervous system (CNS) as a result of apoptosis (Girard et al., Journal of Virology 73, 6066-6072, 1999). We report the development of mixed mouse primary nerve cell cultures from the cerebral cortex of neonatal mice transgenic for the human PV receptor. These cultures contained all three main cell types of the CNS, i.e. neurons, astrocytes and oligodendrocytes. All three cell types were susceptible to PV infection and virus replication in the cultures led to DNA fragmentation characteristic of apoptosis. PV-induced apoptosis was inhibited by the caspase inhibitor benzyloxycarbonyl-Val Ala-Asp(O-Me) fluoromethyl ketone (Z-VAD.FMK), indicating that this process involved caspases. Thus, these mixed mouse primary nerve cell cultures are a new in vitro model for studying the molecular mechanisms of PV-induced apoptosis in nerve cells. PMID- 12124455 TI - Quantities of infectious virus and viral RNA recovered from sheep and cattle experimentally infected with foot-and-mouth disease virus O UK 2001. AB - The profiles of virus production and excretion have been established for sheep experimentally infected with the UK 2001 strain of foot-and-mouth disease (FMD) virus by inoculation and by direct and intensive contact. Virus replicated rapidly in the inoculated sheep, from which a peak infectivity of airborne virus of 10(4.3) TCID(50) per sheep per 24 h was recovered. Around 24 h later, contact infected sheep excreted airborne virus maximally. Similar amounts of airborne virus were recovered from cattle. The excretion of virus by the sheep under these conditions fell into three phases. First, a highly infectious period of around 7 8 days. Second, a period of 1-3 days soon afterwards when trace amounts of viral RNA were recovered in nasal and rectal swabs. Third, at 4 weeks after exposure, the demonstration, by tests on oesophageal-pharyngeal samples, that 50% of the sheep were carriers. The implications of the results and the variable role that sheep may play in the epidemiology of FMD are discussed. PMID- 12124457 TI - Identification of two distinct genotypes of hepatitis E virus in a Japanese patient with acute hepatitis who had not travelled abroad. AB - Two distinct hepatitis E virus (HEV) isolates, designated HE-JI3 and HE-JI4, were identified in a single patient with acute hepatitis in Japan, who had not travelled abroad. The HEV load of HE-JI3 at admission was 10(2) copies/ml, but that of HE-JI4 was tenfold higher at 10(3) copies/ml. The viraemia of HE-JI4 persisted for up to 16 days from admission, whereas HE-JI3 disappeared at 9 days after admission. The entire nucleotide sequence of the HE-JI4 isolate and partial nucleotide sequences of open reading frames (ORFs) 1 and 2 of the HE-JI3 isolate were determined. The full-length nucleotide sequence of HE-JI4 consisted of 7171 nucleotides excluding the poly(A) tail and contained ORF1 encoding 1684 amino acids, ORF2 encoding 671 amino acids and ORF3 encoding 114 amino acids. Sequence and phylogenetic analyses of the HEV genomes indicated that HE-JI4 was most closely related to an HEV isolate (T1) of genotype IV with the same strategy for translation of ORF2 and ORF3, but which differed from it by 16.5% over the entire genome. The HE-JI3 isolate showed the highest nucleotide identity (88.6-95.1%) to the genotype III HEVs, having higher identity to human and swine HEV isolates from the United States (US1, US2 and swUS1) than to those reported thus far from Japan (JRA1 and swJ570). The two co-infecting strains of HE-JI3 and HE-JI4 identified from the single patient shared only 80.1% nucleotide identity. These results indicate that multiple genotypes of HEV co-circulate in Japan, and that genotype IV comprises a remarkably heterogeneous group of HEVs. PMID- 12124458 TI - Common evolutionary origin of aquareoviruses and orthoreoviruses revealed by genome characterization of Golden shiner reovirus, Grass carp reovirus, Striped bass reovirus and golden ide reovirus (genus Aquareovirus, family Reoviridae). AB - Full-length and partial genome sequences of four members of the genus Aquareovirus, family Reoviridae (Golden shiner reovirus, Grass carp reovirus, Striped bass reovirus and golden ide reovirus) were characterized. Based on sequence comparison, the unclassified Grass carp reovirus was shown to be a member of the species Aquareovirus C. The status of golden ide reovirus, another unclassified aquareovirus, was also examined. Sequence analysis showed that it did not belong to the species Aquareovirus A or C, but assessment of its relationship to the species Aquareovirus B, D, E and F was hampered by the absence of genetic data from these species. In agreement with previous reports of ultrastructural resemblance between aquareoviruses and orthoreoviruses, genetic analysis revealed homology in the genes of the two groups. This homology concerned eight of the 11 segments of the aquareovirus genome (amino acid identity 17-42%), and similar genetic organization was observed in two other segments. The conserved terminal sequences in the genomes of members of the two groups were also similar. These data are undoubtedly an indication of the common evolutionary origin of these viruses. This clear genetic relatedness between members of distinct genera is unique within the family Reoviridae. Such a genetic relationship is usually observed between members of a single genus. However, the current taxonomic classification of aquareoviruses and orthoreoviruses in two different genera is supported by a number of characteristics, including their distinct G+C contents, unequal numbers of genome segments, absence of an antigenic relationship, different cytopathic effects and specific econiches. PMID- 12124459 TI - A study of the vaccinia virus interferon-gamma receptor and its contribution to virus virulence. AB - Vaccinia virus (VV) strain Western Reserve gene B8R encodes a 43 kDa glycoprotein that is secreted from infected cells early in infection as a homodimer. This protein has amino acid similarity with the extracellular domain of cellular IFN gamma receptor (IFN-gammaR) and binds and inhibits IFN-gamma from a wide range of species. Here we demonstrate that the B8R protein also inhibits equine IFN-gamma. The 5' end of the B8R mRNA has been mapped by primer extension analysis and the contribution of IFN-gammaRs to VV virulence was studied by the construction of a deletion mutant lacking the B8R gene (vDeltaB8R) and a revertant virus (vB8R-R) in which the B8R gene was re-inserted into the deletion mutant. A recombinant virus that expressed a soluble form of the mouse IFN-gammaR was also constructed and studied. The virulence of these viruses was tested in rodent models of infection. In mice, the loss of the VV IFN-gammaR did not affect virulence compared with WT and revertant viruses, consistent with the low affinity of the VV IFN-gammaR for mouse IFN-gamma. However, expression of the mouse soluble IFN gammaR increased virus virulence slightly. In rabbit skin, loss of the VV IFN gammaR produced lesions with histological differences compared with WT and revertant viruses. Lastly, the affinity constants of the VV IFN-gammaR for human and mouse IFN-gamma were determined by surface plasmon resonance. PMID- 12124460 TI - The vaccinia virus N1L protein is an intracellular homodimer that promotes virulence. AB - The vaccinia virus (VV) N1L gene encodes a protein of 14 kDa that was identified previously in the concentrated supernatant of virus-infected cells. Here we show that the protein is present predominantly (>90%) within cells rather than in the culture supernatant and it exists as a non-glycosylated, non-covalent homodimer. The N1L protein present in the culture supernatant was uncleaved at the N terminus and was released from cells more slowly than the VV A41L gene product, a secreted glycoprotein that has a conventional signal peptide. Bioinformatic analyses predict that the N1L protein is largely alpha-helical and show that it is conserved in many VV strains, in other orthopoxviruses and in members of other chordopoxvirus genera. However, database searches found no non-poxvirus proteins with significant amino acid similarity to N1L. A deletion mutant lacking the N1L gene replicated normally in cell culture, but was attenuated in intranasal and intradermal murine models compared to wild-type and revertant controls. The conservation of the N1L protein and the attenuated phenotype of the deletion mutant indicate an important role in the virus life-cycle. PMID- 12124461 TI - Dermal infection with vaccinia virus reveals roles for virus proteins not seen using other inoculation routes. AB - Previously, we developed a model for testing the virulence and immunogenicity of vaccinia virus (VV) mutants based on the intradermal injection of BALB/c mouse ear pinnae. The model is characterized by a local infection in the inoculated skin without signs of systemic illness, mimicking dermal vaccination with VV. Here a further characterization of this model is presented, including the responses of mice to infectious virus doses as low as 10 p.f.u., a quantification of the infiltrate at the site of infection and use of different virus and mouse strains. The model was then used to compare the pathogenesis of six mutants of VV strain Western Reserve (WR) lacking genes A36R, A40R, A44L, B12R, B13R or B15R with that of appropriate control viruses. All of these genes except B12R and B15R influence the outcome of dermal infection with WR and for A40R and B13R this is the first role that has been reported after infection of mammals. A comparison of new and published results from intradermal and intranasal models is presented, showing that out of 16 gene deletion or insertion mutants of VV, half have phenotypes distinct from controls in only one of these models. Thus, the intranasal and intradermal models are complementary tools for dissecting the genetic basis of VV virulence. PMID- 12124462 TI - Generation of a permanent cell line that supports efficient growth of Marek's disease virus (MDV) by constitutive expression of MDV glycoprotein E. AB - A recombinant cell line (SOgE) was established, which was derived from the permanent quail muscle cell line QM7 and constitutively expressed the glycoprotein E (gE) gene of Marek's disease virus serotype 1 (MDV-1). The SOgE cell line supported growth of virulent (RB-1B) and vaccine (CVI988, 584Ap80C) MDV 1 strains at a level comparable with that of primary chicken embryo cells (CEC). The SOgE cell line was used to produce a vaccine against Marek's disease. Chickens were immunized at 1 day old with 10(3) p.f.u. CVI988 produced on either CEC or SOgE cells. Challenge infection was performed at day 12 with hypervirulent Italian MDV-1 strain EU1. Whereas 7/7 or 6/6 animals, respectively, immunized with SOgE or QM7 cells alone developed Marek's disease, only 1/8 animals from both CVI988-immunized groups exhibited signs of disease, suggesting that SOgE cells are a valuable permanent cell culture system for MDV-1 vaccine production. PMID- 12124463 TI - The attenuated Towne strain of human cytomegalovirus may revert to both endothelial cell tropism and leuko- (neutrophil- and monocyte-) tropism in vitro. AB - The Towne strain of human cytomegalovirus (HCMV), originally recovered from the urine of a congenitally infected newborn, was attenuated through 125 passages in human embryonic lung fibroblast cell cultures. Although reliable markers of attenuation were not identified, the virus was shown to be attenuated by inoculation of both healthy human volunteers and immunocompromised patients. More recently, Towne (like other laboratory-adapted strains) was shown not to have two biological properties typical of recent clinical isolates: endothelial cell tropism and polymorphonuclear leukocyte tropism. These markers of attenuation are lost by all clinical isolates on extensive propagation in cell cultures and are apparently associated with one another. Here, we show that Towne may reacquire both endothelial cell tropism and leuko- (polymorphonuclear- and monocyte-) tropism on adaptation to growth in endothelial cell cultures. However, reversion to endothelial cell tropism is dissociated from reversion to leukotropism, since the latter was reacquired 10-20 passages later. Thus, these two biological properties, which were considered to be encoded by the same viral gene(s), appear to be distinct. Both restriction fragment length polymorphism and Southern blot analysis demonstrated the identity of the attenuated and endothelial cell tropic variants of Towne, thus suggesting that only minor variations (mutations) of the viral genome may be responsible for loss or reacquisition of the two biological properties. Viral genes involved in endothelial cell tropism and leukotropism remain to be identified. However, reversion of attenuated strains to pathogenicity in vivo cannot be excluded a priori. PMID- 12124464 TI - A divalent antibody format is required for neutralization of human cytomegalovirus via antigenic domain 2 on glycoprotein B. AB - Glycoprotein B (gB) of human cytomegalovirus (HCMV) is the dominating protein in the envelope of this virus and gives rise to virus-neutralizing antibodies in most infected individuals. We have previously isolated a neutralizing human antibody specific for antigenic domain 2 (AD-2) on gB, a poorly immunogenic epitope, which nevertheless is capable of eliciting potent neutralizing antibodies. In order to define parameters important for the neutralization of HCMV via gB, we have investigated the virus-neutralizing capacity and the kinetics of the interaction with AD-2 of the monomeric and dimeric forms of a single chain variable fragment (scFv) corresponding to this antibody. We demonstrate here that neutralization of HCMV via AD-2 on gB can be mediated by dimeric scFv, while monomeric fragments cannot mediate neutralization of the virus, despite a slow dissociation from the intact glycoprotein. This finding is discussed in the context of possible mechanisms for antibody-mediated virus neutralization. PMID- 12124465 TI - The Epstein-Barr virus ZEBRA protein activates transcription from the early lytic F promoter by binding to a promoter-proximal AP-1-like site. AB - The ZEBRA protein encoded by the Epstein-Barr virus (EBV) genome activates a switch from the latent to the lytic gene expression programme of the virus. ZEBRA, a member of the basic leucine zipper family of DNA-binding proteins, is a transcriptional activator capable of inducing expression from several virus lytic cycle promoters by binding to activator protein 1 (AP-1)-like sites. The Epstein Barr virus BamHI F promoter, Fp, was for some time believed to initiate EBNA1 specific transcription in EBV-transformed latent cells. More recent data, however, show that Fp is an early lytic promoter and that the dominant EBNA1 gene promoter in latent cells is Qp, located about 200 bp downstream of Fp. In the present investigation we confirm that Fp displays the characteristics of a lytic promoter. Fp is downregulated in latently EBV-infected cells, both in the endogenous virus genome and in reporter plasmids that carry Fp regulatory sequences upstream of position -136 and down to +10 relative to the Fp transcription start site (+1), and is activated on induction of the virus lytic cycle. We show that the repression of Fp in latent stages of infection can be abolished by ZEBRA, and demonstrate that ZEBRA activates Fp through a direct interaction with an AP-1-like site at position -52/-46 in the promoter-proximal Fp region. PMID- 12124466 TI - Characterization of late gene expression factors lef-9 and lef-8 from Bombyx mori nucleopolyhedrovirus. AB - Late gene expression factors, LEF-4, LEF-8, LEF-9 and P47 constitute the primary components of the Autographa californica multinucleocapsid polyhedrovirus (AcMNPV)-encoded RNA polymerase, which initiates transcription from late and very late promoters. Here, characterization of lef-9 and lef-8, which encode their corresponding counterparts, from Bombyx mori NPV is reported. Transcription of lef-9 initiated at two independent sites: from a GCACT sequence located at -38 nt and a CTCTT sequence located at -50 nt, with respect to the +1 ATG of the open reading frame. The 3' end of the transcript was mapped to a site 17 nt downstream of a canonical polyadenylation signal located 7 nt downstream of the first of the two tandem translational termination codons. Maximum synthesis of LEF-9 was seen from 36 h post-infection (p.i.). The transcription of lef-8 initiated early in infection from a GTGCAAT sequence that differed in the corresponding region from its AcMNPV counterpart (GCGCAGT), with consequent elimination of the consensus early transcription start site motif (underlined). Peak levels of lef-8 transcripts were attained by 24 h p.i. Immunocopurification analyses suggested that there was an association between LEF-8 and LEF-9 in vivo. PMID- 12124467 TI - The expansion of a hypervariable, non-hr ori-like region in the genome of Cryptophlebia leucotreta granulovirus provides in vivo evidence for the utilization of baculovirus non-hr oris during replication. AB - In this report a naturally occurring hypervariable region within the genome of different Cryptophlebia leucotreta granulovirus genotypes is characterized. The region consists of a stretch of direct repeats, short palindromes and an unusual AT-rich region. Although the organization of these repeat sequences is unique to baculoviruses, it has the structural features of a 'non-hr' origin of DNA replication (ori). Restriction analysis and Southern hybridization revealed that this region is expanded during virus replication. Sequence comparison of different isolated genotypes indicated that the expansion is caused by concatenation of short repeats within the region or by concatenation of the complete region. These findings indicate that the expansion of non-hr origin-like regions is not restricted to defective-interfering particles, as was found previously for baculoviruses propagated in cell culture. Moreover, it appears that non-hr complexity contributes to the natural heterogeneity and genetic plasticity of baculovirus genomes. Also, circumstantial evidence is discussed that hr oris might have developed from internal rearrangement and multiplication of a non-hr ori during baculovirus evolution. PMID- 12124468 TI - Polydnavirus replication: the EP1 segment of the parasitoid wasp Cotesia congregata is amplified within a larger precursor molecule. AB - Polydnaviruses are unique viruses: they are essential for successful parasitism by tens of thousands of species of parasitoid wasps. These viruses are obligatorily associated with the wasps and are injected into the host during oviposition. Molecular analyses have shown that each virus sequence in the segmented polydnavirus genome is present in the wasp DNA in two forms: a circular form found in the virus particles and an integrated form found in the wasp chromosomes. Recent studies performed on polydnaviruses from braconid wasps suggested that the circular forms were excised from the chromosome. The different forms of the EP1 circle of Cotesia congregata polydnavirus during the pupal-adult development of the parasitoid wasp were analysed. Unexpectedly, an off-size fragment formerly used to diagnose the integration of the EP1 sequence into wasp genomic DNA was found to be amplified in female wasps undergoing virus replication. The EP1 sequence is amplified within a larger molecule comprising at least two virus segments. The amplified molecule is different from the EP1 chromosomally integrated form and is not encapsidated into virus particles. These findings shed light on a new step towards EP1 circle production: the amplification of virus sequences preceding individual circle excision. PMID- 12124469 TI - Adenovirus E1-transformed cells grow despite the continuous presence of transcriptionally active p53. AB - The E1 region of adenovirus (Ad) type 5 is capable of transforming cells. According to current concepts, the Ad E1B 55 kDa (E1B 55K) protein enables transformed cells to grow by constantly binding and inactivating the p53 tumour suppressor protein. To test this model, the transcriptional activity of p53 was determined in Ad E1-transformed cells. Surprisingly, it was found that a p53 responsive promoter is highly active in Ad E1-transformed cells and further activated only 3- to 4-fold (compared to 200-fold in p53(-/-) cells) by exogenously expressed p53 or p53mt24-28, a p53 mutant that is transcriptionally active but unable to bind the E1B 55K. On the other hand, the transient overexpression of E1B 55K led to a strong downregulation of a p53-responsive promoter relative to its baseline activity in Ad E1-transformed cells but not in p53(-/-) cells. COS-7 cells, transformed by simian virus 40 (SV40), also showed constitutive p53 activity, whereas HeLa cells, transformed with oncogenic human papillomavirus, did not. Upon stable transfection, Ad E1-transformed cells but not p53(-/-) cells gave rise to colonies that expressed exogenous p53 or p53mt24 28 but, nonetheless, grew at near-wild-type rates. It is proposed that E1B 55K or the SV40 tumour antigen are saturated by the p53 protein, which accumulates in virus-transformed cells, leaving a proportion of active p53 molecules. The transformation of cells by the Ad E1 genes confers permissiveness for active p53, conceivably by inactivating the relevant products of p53 target genes that would otherwise prevent cell growth. Thus, Ad-transformed cells contain and tolerate active p53. PMID- 12124470 TI - Genotype H: a new Amerindian genotype of hepatitis B virus revealed in Central America. AB - The complete genomes were sequenced for ten hepatitis B virus (HBV) strains. Two of them, from Spain and Sweden, were most similar to genotype D, although encoding d specificity. Five of them were from Central America and belonged to genotype F. Two strains from Nicaragua and one from Los Angeles, USA, showed divergences of 3.1-4.1% within the small S gene from genotype F strains and were recognized previously as a divergent clade within genotype F. The complete genomes of the two genotype D strains were found to differ from published genotype D strains by 2.8-4.6%. Their S genes encoded Lys(122), Thr(127) and Lys(160), corresponding to the putative new subtype adw3 within this genotype, previously known to specify ayw2, ayw3 or, rarely, ayw4. The complete genomes of the three divergent strains diverged by 0.8-2.5% from each other, 7.2-10.2% from genotype F strains and 13.2-15.7% from other HBV strains. Since pairwise comparisons of 82 complete HBV genomes of intratypic and intertypic divergences ranged from 0.1 to 7.4% and 6.8 to 17.1%, respectively, the three sequenced strains should represent a new HBV genotype, for which the designation H is proposed. In the polymerase region, the three strains had 16 unique conserved amino acid residues not present in genotype F strains. So far, genotype H has been encountered in Nicaragua, Mexico and California. Phylogenetic analysis of the complete genomes and subgenomes of the three strains showed them clustering with genotype F but forming a separate branch supported by 100% bootstrap. Being most similar to genotype F, known to be an Amerindian genotype, genotype H has most likely split off from genotype F within the New World. PMID- 12124471 TI - Molecular characterization of three newly recognized rat parvoviruses. AB - Rodent parvoviruses have been documented to interfere with both in vivo and in vitro research. In this study, three rat parvoviruses distinct from previously characterized rodent parvoviruses were identified from naturally infected rats obtained from four discrete sources. These three newly recognized parvoviruses were designated rat minute virus (RMV)-1a, -1b and -1c. In this study, the genomic nucleotide sequence and the predicted amino acid sequences of proteins for each of the three RMV-1 variants and Kilham rat virus (KRV) were determined and compared with previously characterized rodent parvoviruses. The three RMV-1 variants were shown to be closely related to each other, to be distinct from but closely related to KRV and H-1 virus, and to be significantly different from the previously identified rat parvovirus isolate, RPV-1a. PMID- 12124473 TI - Radiotracer characterization of coronary artery lesions. PMID- 12124472 TI - Interaction in vitro between the proteinase of Tomato ringspot virus (genus Nepovirus) and the eukaryotic translation initiation factor iso4E from Arabidopsis thaliana. AB - Eukaryotic initiation factor eIF(iso)4E binds to the cap structure of mRNAs leading to assembly of the translation complex. This factor also interacts with the potyvirus VPg and this interaction has been correlated with virus infectivity. In this study, we show an interaction between eIF(iso)4E and the proteinase (Pro) of a nepovirus (Tomato ringspot virus; ToRSV) in vitro. The ToRSV VPg did not interact with eIF(iso)4E although its presence on the VPg-Pro precursor increased the binding affinity of Pro for the initiation factor. A major determinant of the interaction was mapped to the first 93 residues of Pro. Formation of the complex was inhibited by addition of m(7)GTP (a cap analogue), suggesting that Pro-containing molecules compete with cellular mRNAs for eIF(iso)4E binding. The possible implications of this interaction for translation and/or replication of the virus genome are discussed. PMID- 12124474 TI - Leukoscan for orthopaedic imaging in clinical practice. AB - Studies have suggested that orthopaedic infection can be successfully imaged with 99mTc antigranulocyte antibody Fab' fragments (Leukoscan). This study examined the value of the technique in a UK clinical practice. A retrospective review of the first 55 patients imaged was performed. Patients had three-phase bone scintigraphy followed by Leukoscan imaging. The latter was performed using planar views 2 h and 6 h post-injection of 750 MBq 99mTc Leukoscan. In 47 patients there was adequate clinical information available to compare to the scintigraphy result. There were 13 positive infections. The Leukoscan findings produced 11 true positives, 26 true negatives, eight false positives and two false negatives with resulting sensitivity 85%, specificity 77%, positive predictive accuracy 58%, and negative predictive accuracy 93%. This study suggests that Leukoscan can be used successfully to image orthopaedic infection, with its greatest strength being a high negative predictive accuracy. Positive studies may require further correlative imaging. PMID- 12124475 TI - Assessment of inflammatory bowel disease by using two different (99m)Tc leucocyte labelling methods. AB - The aim of this study was to retrospectively compare the diagnostic accuracy of (99m)Tc hexamethylpropyleneamine oxime (HMPAO) white blood cell scintigraphy in patients with a suspicion of active inflammatory bowel disease by using two different cell suspension media: leukocyte poor plasma (LPP) and Hanks' balanced salt solution (HBSS). Leukocytes from 30 patients were labelled using LPP and in 28 cases using HBSS. In the LPP method the leukocytes were resuspended in 0.5 ml cell-free plasma while in the HBSS method the cells were resuspended in 0.5 ml HBSS. Scintigraphic images were obtained at 30 min and 2 h after injection of 185 200 MBq (99m)Tc-HMPAO leukocytes. The leukocyte labelling efficiency was 65.5% and 89.0%, respectively, for the LPP and HBSS methods. There were 22 true positive, seven true negative and one false negative result in the LPP group, while in the HBSS group results were 18, nine and one, respectively. Diagnostic accuracy was similar with both methods although sensitivity was slightly higher in the HBSS group. In summary, these date indicate that leukocyte scintigraphy labelling using HBSS as the resuspension medium should be used as a first option for white blood cell labelling and diagnosis of inflammatory bowel disease. PMID- 12124476 TI - Biodistribution and dosimetry of [(18)F]fluorodeoxyglucose labelled leukocytes in normal human subjects. AB - SUMMARY: This study was performed in order to assess [(18)F]fluorodeoxyglucose white blood cell ((18)F-FDG WBC) dosimetry in normal human subjects. Using previously reported methods, mixed cell suspensions of autologous leukocytes were prepared from four normal volunteers. Leukocytes were labelled in heparin-saline by incubation with (18)F-FDG at 37 degrees C for 20 min. After washing and resuspension, (18)F-FDG WBCs (225-315 MBq) were administered by intravenous injection. Whole-body imaging was performed at 0.5, 1, 2, 4 and 6 h using a GE Varicam with 511 keV collimation. Blood samples were obtained at corresponding times as well as fractionated urinary collection. Whole-body anterior and posterior images were used for calculation of organ dosimetry. Uptake of (18)F FDG WBCs occurred predominantly within the reticulo-endothelial system. Plasma activity, urinary excretion (9.9+/-2.3% at 6 h), and brain uptake (1.7+/-0.4%) were consistent with partial elution of (18)F-FDG. Positron emission tomography imaging performed at 5-6 h after injection yielded good quality images of reticulo-endothelial uptake. Whole-body and organ dosimetry for (18)F-FDG WBCs in doses of 225-250 MBq are comparable with reported results for conventional doses of (111)In oxine labelled leukocytes. Further studies of (18)F-FDG WBC as an agent for positron emission tomography imaging of inflammatory disease appear warranted. PMID- 12124477 TI - Fluorodeoxyglucose positron emission tomography and somatostatin receptor scintigraphy for diagnosing and staging carcinoid tumours: correlations with the pathological indexes p53 and Ki-67. AB - We performed this study in order to evaluate the diagnostic accuracy of whole body fluorodeoxyglucose positron emission tomography (FDG PET) imaging and somatostatin receptor scintigraphy (SRS) for localizing primary carcinoid tumours and evaluating the extent of the disease. A secondary aim was to correlate those findings with the histological characteristics of the lesions. FDG PET was performed in 17 patients and SRS in 16. All patients had pathologically proven carcinoids. All lesions were verified by histopathological analysis or by follow up. Ki-67 and p53 expression were assessed as an indicator of the tumours' aggressiveness. FDG PET correctly identified 4/7 primary tumours and 8/11 metastatic spreads, as compared to six and 10 respectively, for SRS. Most tumours were typical carcinoids with low Ki-67 expression. No correlation was found between the histological features and the tracer's uptake. We conclude that SRS remains the modality of choice for evaluating patients with carcinoid tumours, regardless of their proliferative activity. FDG PET should be reserved to patients with negative results on SRS. PMID- 12124478 TI - Evaluation of (111)In-pentetreotide, (131)I-MIBG and bone scintigraphy in the detection and clinical management of bone metastases in carcinoid disease. AB - Bone metastases are assumed to be rare in carcinoid disease and to be associated mainly with bronchial primaries. The aim of the present study was to evaluate the occurrence of bone metastases in patients with metastatic carcinoid tumours, and the role of various nuclear medicine modalities (bone scintigraphy, (111)In pentetreotide and (131)I-MIBG) in its detection and clinical management. Nine (2 women, 7 men, median age 65 years) out of 86 consecutive carcinoid patients treated between 1987 and 1998 developed bone metastases (10%) with a median interval of 37 months between the diagnosis of metastatic carcinoid and bone metastases. Seven of them had non-bronchial primaries. (111)In-pentetreotide scintigraphy failed to detect the bone lesions in 50% of the cases, and (131)I meta-iodobenzylguanidine(MIBG) scintigraphy in almost 80% of cases. Standard bone scintigraphy, however, was positive in all. Pain relief of bone metastases by means of radiation therapy was obtained in 5 of 6 patients. In another patient palliation of pain symptoms was obtained with Rhenium-186-hydroxyethylidene diphosphonate. Octreotide, Interferon of MIBG were ineffective for this purpose. It is concluded that bone metastases in carcinoid patients may be missed on (131)I-MIBG and (111)In-pentetreotide scintigraphy. Bone scintigraphy is a sensitive imaging technique. Diagnostic nuclear medicine modalities may be helpful in the clinical management of carcinoid disease. PMID- 12124479 TI - Estimating uptake in liver and abdominal tumours for radionuclide therapy dosimetry. AB - Historically, patients for radionuclide therapy have received fixed activities, sometimes normalized to body weight or body surface area. As radionuclide therapy develops, however, practitioners are becoming interested in individualized doses. This requires individual estimates of uptake to be routinely available. Such estimates need not necessarily be highly accurate but the magnitude of associated errors must be known. This paper looks specifically at simple methods for estimating relative (tumour to organ) uptake and also absolute (organ) uptake, for abdominal tumours and the liver. For relative uptake, the ratio of geometric mean counts gives a rough estimate, to within a factor of 2. Absolute liver uptake can be estimated from the geometric mean of liver counts but a correction must be applied for patient attenuation. Such a correction can be calculated solely from the patient's height and weight. The patient weight alone gives a correlation coefficient of 0.86, when plotted against the measured attenuation. However, this correction does not work well for patients heavier than 100 kg. The attenuation index, which is calculated from a patient's weight and height, gives a correlation coefficient of 0.90, for patients ranging in weight from 41 kg to 120 kg. PMID- 12124480 TI - Dynamic evaluation of absorbed dose to the bladder wall with a balloon-bladder phantom during a study using [(18)F]fluorodeoxyglucose positron emission imaging. AB - An accurate evaluation of the absorbed dose to the bladder wall from 2 [(18)F]fluoro-2-deoxy-d-glucose (FDG) is clinically important because the bladder is considered as a critical organ in most positron emission tomography (PET) studies that cumulate about 20% of the total activity injection during image procedures. In the MIRD calculation, no allowance is made for the inclusion of all the dynamic parameters that affect the actual dose to the bladder wall to be taken in the dose assessment. The goal of the study is to propose a dose evaluation model by using a dynamic bladder phantom and time-activity curves from the bladder PET imaging. The proposed model takes all dynamic parameters into account and provides a much more accurate dose estimation to the bladder. In this study, the lowest dose to the bladder wall was obtained at the conditions of having a larger initial volume for the bladder contents and a higher production rate for urine. It is then advised patients to drink a bulk amount of water before the FDG injection or after urine voiding to facilitate urine production and to enlarge the bladder surface area, which are the most crucial steps in reducing the dose to the bladder wall. In our study, the voiding schedule in dose calculation plays certain roles although it is much more critical in the conventional MIRD calculation. The model estimated that the lowest dose to the bladder would occur at an initial void about 40 min after the FDG injection and the urine voiding was as complete as possible. PMID- 12124481 TI - Bone marrow immunoscintigraphy in haematological patients with pancytopenia: preliminary results. AB - The aim of this study was to assess the clinical value of bone marrow immunoscintigraphy using the (99m)Tc labelled anti-NCA-95 antigranulocyte antibodies (AGAb) and of AGAb bone marrow uptake ratio (UR) in the initial diagnostic work-up of diseases with depression of the bone marrow. Twenty-four whole-body bone marrow scans were performed in 23 patients (11 women, 12 men; median age 46 years, range 17-74 years) 5 h after i.v. injection of 370 MBq of AGAb. The UR was calculated from the posterior view drawing an irregular region of interest around the sacroiliac and a background areas. The mean UR in pancytopenic patients was 2.3+/-1.5 (range 0.3-5.8), thus being significantly lower (P=0.45 x 10(-6)) than the mean UR in a control group of 50 patients (mean UR 7.3+/-2.3; range 4.4-12.6) obtained previously. Considering patient age, there was no overlap between UR of pancytopenic patients and the respective normal ranges. The bone marrow appearance on scans seemed to be characteristic for the different haematological diseases investigated. In six patients with myelofibrosis, bone marrow scans demonstrated diffusely decreased bone marrow activity and prominent splenic uptake, possibly related to extramedullary haematopoiesis. In aplastic anaemia, highly reduced and patchy marrow uptake was observed in four patients (five scans), in one of them persisting even after blood cell counts had recovered to the near-normal range. In another two patients with aplastic anaemia, diffusely decreased bone marrow uptake was obtained. In patients with myeloid leukaemia, bone marrow patterns were almost normal probably because the target antigen is often expressed on neoplastic myeloid cells, too. Bone marrow extension was a common finding in these patients. There is an obvious differentiation between haematological patients with pancytopenia and normal subjects by means of AGAb bone marrow uptake ratio. The distinct patterns of AGAb distribution may be indicative for particular haematological diseases. PMID- 12124482 TI - Predictive value of (99m)Tc sestamibi scintigraphy in the evaluation of doxorubicin based chemotherapy response in patients with advanced breast cancer. AB - Resistance to doxorubicin based chemotherapy is a major therapeutic problem limiting advanced breast cancer treatment. 99mTc hexakis-2 methoxyisobutylisonitrile ((99m)Tc-MIBI) has been reported to be extruded from tumour cells by the P-glycoprotein and multidrug resistance protein encoded by MDR1 and MRP1 genes, respectively. These proteins are involved in the cellular efflux of several chemotherapeutic agents including doxorubicin. The aim of this study was to investigate the clinical value of a standard (99m)Tc-MIBI scintimammography technique in the prediction of response to chemotherapy in advanced breast cancer patients. Fifty-six lesions from 33 female patients with locally advanced (n=27) or recurrent breast cancer (n=6) were included in the study. MIBI scintigraphy was performed 2-8 days prior to chemotherapy (FAC regimen). Images were acquired 10 min and 1 h post-injection of 740-1110 MBq of (99m)Tc-MIBI. Tumour-to-normal background tissue uptake ratios were calculated on each lesion in the early (T/B(e)) and delayed phase of the study (T/B(d)). Both T/B(e) and T/B(d) ratios were significantly higher (P<0.0001) in responders (n=43) than nonresponders (n=13). Diagnostic values of (99m)Tc-MIBI in the prediction of chemotherapy response were evaluated using the arbitrary cut-off values of 1.5 for T/B(e) and 1.4 forT/B(d). Sensitivity, specificity, positive and negative predictive values were 88.4%, 92.3%, 97.4%, 70.6%; and 90.7%, 100.0%, 100.0%, 76.6%, for T/B(e) and T/B(d), respectively. We conclude that (99m)Tc-MIBI scintigraphy may be a clinically valuable tool for guiding chemotherapy in advanced breast cancer patients. PMID- 12124483 TI - Cerebral metabolic changes associated with Lyme disease. AB - There are no positron emission tomography (PET) studies reported in the literature with regards to brain metabolism and function in patients with Lyme disease. These patients frequently present with various neurological symptoms, including memory problems. We used [(18)F]fluorodeoxyglucose (FDG) PET to determine the metabolic landscape in 23 patients with Lyme disease. Images were evaluated for cortical and subcortical abnormalities by two experienced reviewers blinded to the clinical information. The most striking finding was hypometabolism in the temporal lobes in 17/23 (74%) patients. Of these, 12 had bilateral temporal lobe hypometabolism, two had left temporal lobe, and three had right temporal lobe hypometabolism. Seven of the patients with temporal lobe hypometabolism had diffuse cortical hypometabolism that included the frontal and parietal lobes. Lyme disease appears to have two primary patterns of brain involvement on FDG PET scans, specific temporal lobe hypometabolism or a diffuse cortical hypometabolism. The involvement of the temporal lobes in both patterns is likely associated with the memory disturbances described in many of these patients. Although there was no clear diagnostic pattern, and many of the defects were mild, FDG PET imaging may provide important information regarding the areas of the brain affected in patients with neurological symptoms associated with Lyme disease. PMID- 12124484 TI - Relationship between cognitive function and regional cerebral blood flow in Alzheimer's disease. AB - The Mini-Mental State examination (MMSE) is frequently used to assess the cognitive function of neurological patients. The purpose of this study was to investigate associations between regional cerebral blood flow (rCBF) and MMSE scores in Alzheimer's disease (AD) and localization of cognitive functions. Fifty nine patients with probable AD (21 males and 38 females; mean age 74 years) and 12 normal volunteers (five males and seven females; mean age 73 years) were studied. CBF was measured by SPECT using the N-isopropyl-p [(123)I]iodoamphetamine autoradiography method. The CBF images were reconstructed in parallel with the orbitomeatal line and parallel to the long axis of the temporal cortex. Regions of interest were set in the cerebral and cerebellar cortex. Multivariate analysis was performed by the step-wise method, using each section of the MMSE as the dependent variable and the rCBF ratio as the independent variable. The decline in rCBF in the parietal cortex and hippocampus reflected disorientation, and the most significant cortex affecting scores on each section of the MMSE were found to be the anterior temporal cortex for registration, the frontal cortex for attention and calculation, the medial temporal cortex for recall, and the posterior temporal cortex for language. PMID- 12124485 TI - Automatic detection of coronary artery disease in myocardial perfusion SPECT using image registration and voxel to voxel statistical comparisons. AB - The purpose of this study was to compare the performance of automatic detection of coronary artery disease (CAD) with that of expert observers. A male and female normal image template was constructed from normal stress technetium-99m single photon emission computed tomography (SPECT) studies. Mean and standard deviation images for each sex were created by registering normal studies to a standard shape and position. The test group consisted of 104 patients who had been routinely referred for SPECT and angiography. The gold standard for CAD was defined by angiography. The test group studies were registered to the respective templates and the Z-score was calculated for each voxel. Voxels with a Z-score greater than 5 indicated the presence of CAD. The performance of this method and that of three observers were compared by continuous receiver operating characteristic (CROC) analysis. The overall sensitivity and specificity for automatic detection were 73% and 92%, respectively. The area (Az) under the CROC curve (+/-1 SE) for automatic detection of CAD was 0.88+/-0.06. There was no statistically significant difference between the performances of the three observers in terms of Az and that of automatic detection (P> or =0.25, univariate Z-score test). The use of this automated statistical mapping approach shows a performance comparable with experienced observers, but avoids inter-observer and intra-observer variability. PMID- 12124486 TI - Clinical implications of cardiac (123)I-meta-iodobenzylguanidine scintigraphy and cardiac natriuretic peptides in patients with heart disease. AB - The purpose of this study was to evaluate whether or not cardiac sympathetic nerve activity, using (123)I-meta-iodobenzylguanidine ((123)I-MIBG) imaging, and cardiac natriuretic peptides (atrial and brain, ANP and BNP) were independent predictors of cardiac events, and, if so, which was the stronger predictor. Planar (123)I-MIBG images were obtained from 62 patients with heart disease. Plasma ANP and BNP levels, left ventricular ejection fraction (LVEF) by echocardiography, serum total cholesterol and triglyceride were measured. (123)I MIBG was assessed as the heart-to-mediastinum (H/M) ratio of the delayed image and the washout rate (WoR) from the early to the delayed image. Patients were followed up for an average of 16.2 months, and 12 of 62 patients had cardiac events. Patients with events had significantly lower LVEF and H/M ratio compared with those without events. They had significantly higher WoR, ANP and BNP. By multivariate Cox proportional hazard analysis, (123)I-MIBG (H/M or WoR), ANP and BNP were independent predictors for cardiac events. Event-free survival using a Kaplan-Meier model, with a threshold value of 2.0 for H/M and 45% for WoR, showed that patients with H/M<2.0 and/or WoR>45% had a significantly poorer prognosis. These results suggest that (123)I-MIBG imaging and cardiac natriuretic peptides are useful tools for the evaluation of patients with heart disease, and that cardiac sympathetic nerve activity is a stronger predictor of cardiac events. PMID- 12124487 TI - The importance of delayed images in gastrointestinal bleeding scintigraphy. AB - Although radionuclide methods for the detection of gastrointestinal (GI) bleeding have been available for more than 20 years, the value of delayed images in GI bleeding scintigraphy is still regarded controversially. The aim of this study was to determine the value of delayed images in a group of patients with predominantly low-grade intermittent bleeding. Eighty-nine consecutive GI bleeding scintigraphies of 75 patients were analysed retrospectively. All patients were referred to our department after other diagnostic methods had failed to identify the localization of GI bleeding. After the dynamic study, delayed images were acquired for up to 24 h until a bleeding site was identified. Data on the clinical outcome were available in all but five patients. No patient with a negative scan died from GI bleeding. A positive result was found in 41 patients (55%). The scans of 11 of these 41 patients (27%) became positive during dynamic imaging. Four required immediate surgery and, in another patient, surgery was not performed because of diffuse bleeding of the entire GI tract. One patient died without surgical intervention. Thirty-three scans of 30 of these 41 patients (73%) were positive on delayed imaging only, leading to surgery in 12 individuals. Our findings demonstrate the importance of delayed images in GI bleeding scintigraphy. Many of our patients who required surgery had scans that did not become positive for several hours. PMID- 12124488 TI - Triple-head gamma camera PET: system overview and performance characteristics. AB - Positron emission tomography (PET) is currently performed using either a dedicated PET scanner or scintillation gamma camera equipped with electronic circuitry for coincidence detection of 511 keV annihilation quanta (gamma camera PET system). Although the resolution limits of these two instruments are comparable, the sensitivity and count rate performance of the gamma camera PET system are several times lower than that of the PET scanner. Most gamma camera PET systems are manufactured as dual-detector systems capable of performing dual head coincidence imaging. One possible step towards the improvement of the sensitivity of the gamma camera PET system is to add another detector head. This work investigates the characteristics of one such triple-head gamma camera PET system capable of performing triple-head coincidence imaging. The following performance characteristics of the system were assessed: spatial resolution, sensitivity, count rate performance. The spatial resolution, expressed as the full width at half-maximum (FWHM), at 1 cm radius is 5.9 mm; at 10 cm radius, the transverse radial resolution is 5.3 mm, whilst the transverse tangential and axial resolutions are 8.9 mm and 13.3 mm, respectively. The sensitivity for a standard cylindrical phantom is 255 counts.s(-1).MBq*(-1)), using a 30% width photopeak energy window. An increase of 35% in the PET sensitivity is achievable by opening an additional 30% width energy window in the Compton region. The count rate in coincidence mode, at the upper limit of the systems optimal performance, is 45 kc.s(-1) (kc=kilocounts) using the photopeak energy window only, and increases to 60 kc.s(-1) using the photopeak + Compton windows. Sensitivity results are compared with published data for a similar dual-head detector system. PMID- 12124489 TI - [Genotypic analysis and immunohistochemistry in human cancers: conflicting or complementary information?]. PMID- 12124491 TI - [Immunohistochemistry and genotype analysis of tumors. First part: Which future for the immunochemical diagnosis of cancer?]. AB - Immunohistochemistry (IHC) is a rapid morphological method that allows the detection of proteins involved in different mechanisms of cancer development. It is therefore a useful tool in the study of cancerogenesis. The best known example is the product of the p53 gene, a tumour suppressor gene which is altered in 50% of all human tumors. In fact, these p53 gene mutations lead to cell protein accumulation whereas the p53 product is not detectable in normal cells. This method also enables the detection of fusion proteins which result from chimeric transcript like WT1 in desmoplastic small round cell tumors, ALK in anaplastic large-cell lymphomas and FLI-1 in Ewing's sarcomas. On the contrary, gene inactivation can induce loss of immunostaining. hMLH1 and hMSH2, which are committed in DNA mismatch repair, can be altered in familial digestive carcinomas, such as hereditary non polyposis colorectal cancer. Thus IHC, which allows us to focus on the altered gene by loss of its product in tumoral cells, represents a good alternative to molecular analysis. IHC is also useful to detect the product of oncogene overexpression such as HER-2 in some breast carcinomas, which allows appropriate therapeutic protocols. Finally, IHC can be used in diagnostic, prognostic and therapeutic ends. Nevertheless, difficulties can be en countered in the interpretation of the results. Therefore, IHC must be performed in quality control trials. PMID- 12124490 TI - [Epstein-Barr virus in Hodgkin's disease: the example of central Tunisia]. AB - The purpose of this study was to evaluate the prevalence of Epstein-Barr virus in Hodgkin disease in Tunisia through a series of 77 cases. Association with Epstein Barr virus was demonstrated by Epstein-Barr encoded early RNA transcripts (EBER) in situ hybridization in 70% of cases and by latent membrane protein 1 (LMP1) immunohistochemistry in 58.4% of cases. EBER positive cases were more frequent in extreme age classes (<15 and>54 years) there was no correlation with sex, histologic sub-type and clinical stage. Our findings show a high prevalence for EBV infection in Tunisian Hodgkin's disease particularly among extreme ages. PMID- 12124492 TI - [Immunochemistry and genotype analysis of tumors. Second part: What future for the molecular diagnosis of tumors?]. AB - The twentieth century has witnessed the discovery of the various genetic alterations that drive a normal cell toward neoplasia. The challenge of the near future concerns the diagnostic of these alterations and their correlation with clinical parameters such as response to therapy or survival. It will be important to define, for each patient, a specific profile of the various alterations in order to improve and personalize the therapy. It is also essential to understand the various mechanisms of these alterations in order to develop new diagnostic procedures. Furthermore, the relation between genotype and phenotype is not straightforward and can be influence by various parameters such as the localization of the mutation, the alterations of other genes or the expression of modifier genes. PMID- 12124493 TI - [Pulmonary oxalosis with necrotizing pulmonary aspergillosis]. AB - Pulmonary oxalosis is a very rare pseudotumoral lesion; it is often secondary to an aspergillus infection. Oxalic acid (C(2)H(2)O(4)) is a mycotoxin released by Aspergillus niger and sometimes by several other fungi, including A flavus and A fumigatus. We report a case of a 69 year old man, with previous history of pulmonary tuberculosis, followed for recurrent hemoptysis. On the chest radiography, the right upper lobe lung showed a cavitary lesion with thick and irregular walls and a dense material that suggested a pulmonary aspergilloma. Microscopically, it was a pulmonary oxalosis associated with chronic necrotising pulmonary aspergillosis. Our aim is to discuss the epidemiological characteristics, the diagnosis and the histogenesis of this unusual lesion. PMID- 12124494 TI - [Villo-glandular adenoma or polypoid dysplasia of esophagus: a case report]. AB - Gastrointestinal adenomas are neoplasms of glandular epithelium containing dysplasia of varying degrees. They are rare in esophagus. A case of villous tumor of the esophagus in a 71-year-old man is described. Histologically, this polypoid, villous lesion was developed on Barrett's esophagus and contained dysplastic epithelium and small adenocarcinomatous foci. This case illustrates that esophagus adenoma may be a premalignant lesion like other adenomas of the gastrointestinal tract. The term of polypoid dysplasia has therefore been recommended. This lesion, usually developed in gastric heterotopy or Barrett's esophagus, is often associated with high-grade dysplasia or carcinoma. PMID- 12124495 TI - [An unusual mediastinal malignant tumor: thymic carcinoma]. AB - Thymic carcinoma is a relatively rare neoplasm. Current incidence is difficult to assess. There is some controversy regarding its definition and its diagnostic criteria. We report a case of epidermoid carcinoma in a 48-year-old man located in the thymus area of the anterior mediastinum. Clinicopathological features and prognosis are reviewed. PMID- 12124496 TI - [Peritoneal glioblastoma: recurrence of ovarian immature teratoma (report of a case)]. AB - Immature teratomas of the ovary represent less than 1% of all ovarian teratomas. They contain several tissues that derive from the three embryological layers: ectoderm, mesoderm and endoderm. They are rarely associated with peritoneal implants that are essentially composed of mature glial tissue, and of benign evolution. We report the case of a 37-year- old woman who presented an immature teratoma of the right ovary that recurred seven years later as a malignant neuroepithelial peritoneal tumor resembling a glioblastoma. Glioblastoma was diagnosed at a second recurrence six months later. We discuss the histopathogenesis of peritoneal implants secondary to immature teratomas. PMID- 12124497 TI - [Undifferentiated embryonal sarcoma of the liver in an adult: a case report]. AB - Undifferentiated embryonal sarcoma of the liver is rare. It is usually observed in children and adolescents. We report one case of embryonal sarcoma of the liver arising in patient without any antecedent. The only symptom was right scapular pain. The liver scan showed a multicystic lesion suspicious for infectious origin or a tumor. Serologies for ecchinococcus, schistosomiasis and brucellosis were negative. The treatment was a right hepatectomy. On gross examination, the tumor was unencapsulated, multicystic and contained large areas of necrosis admixed with gelatinous areas. Microscopically, there were epithelioid and spindle tumor cells in a myxoid stroma. Lipoblastic-like or rhabdomyoblastic-like, giant cells and PAS positive hyaline globules in the cell cytoplasm were present. The tumor cells expressed vimentin, cytokeratin (KL1), alpha-1-antitrypsin and smooth muscle actin. This observation shows that embryonal sarcoma of the liver may develop in adult patients and should be taken into consideration in any differential diagnosis of cystic hepatic tumor. PMID- 12124498 TI - [An original thymoma]. PMID- 12124499 TI - [Multiple pulmonary nodules]. PMID- 12124500 TI - [Subcutaneous skin nodule of an infant's face]. PMID- 12124501 TI - [Diagnosis with hobnail]. PMID- 12124503 TI - [Immunochemistry evaluation of HER2 status in infiltration breast cancer: technical protocol and interpretation guidelines]. AB - In Europe, patients who may benefit from Herceptin((R)) (an HER2 targeted drug) are currently selected by immunohistochemistry (IHC). Reliable detection of HER2 status is essential to the appropriate usage of Herceptin(R), because its specificity is limited to tumours overexpressing HER2. It is essential that the IHC evaluation of the HER2 status of a mammary carcinoma be optimized and reliable. This technical paper reviews the different steps of the IHC technique, the controls and, the rules for interpretation. The sensitivity of the IHC technique must be adjusted so as not to produce false negatives or false positives. As opposed to other methods, it can be carried out whatever the fixation conditions of the tissues. The interpretation of the immunostains also requires training; it is fraught with problems for intermediate positivities. The ideal score to evaluate HER2 status has not yet been defined. It will thus be necessary to report the percentage of stained cells, the intensity of the staining, and, in respect to Herceptin((R)) treatment, the HercepTest scoring system (recommended in the package insert). Once acquired, this knowledge must be perpetuated by the observation of rules of good technical practice (internal and external controls, quality assurance programs). FISH should be used for complementary assessment of 2+ cases (on condition that they have not been fixed in Bouin's liquid) and for the calibration of the IHC technique. PMID- 12124502 TI - [Interphase FISH analysis on histologic sections of fixed tissues for t(11;14) (q13;q32) detection in mantle cell lymphoma and t(8;14)(q24;q32) in Burkitt's lymphoma]. AB - We describe an interphase FISH analysis for formol-fixed, paraffin-embedded tissue sections with commercial probes detecting the t(11;14)(q13;q32) in mantle cell lymphoma and the t(8;14)(q24;q32) in Burkitt's lymphoma. Staining of an adjacent section allowed identification of tumoral areas. The cut-off value was evaluated in reactive lymph nodes at 5% for both probes. An incomplete hybridization pattern was found in 18 to 26% of the nuclei but was always lower than the percentages of translocated cells in tumoral regions. A t(11;14) was detected in 4/4 mantle cell lymphomas and a t(8;14) in 2/3 Burkitt's lymphomas. Interphase FISH analysis of fixed-tissue sections is a reliable technique for the direct detection of lymphoma-associated translocations on routine histological material. PMID- 12124504 TI - [Diagnosis of a pulmonary coccidioidomycosis on mediastinal sample]. PMID- 12124505 TI - [Comments on the technical note proposed by Ph.Taniere on handling digestive endoscopy mucosectomy specimens]. PMID- 12124506 TI - [Something new for the autopsy room?]. PMID- 12124508 TI - [Morbiliform, rubeoliform and other scarlatiniform rashs: obsolete terminology]. PMID- 12124509 TI - [Pain management and the use of analgesics in dermatology]. AB - OBJECTIVE: To determine pain assessment and management, and the use of analgesics in dermatology. PATIENTS AND METHODS: Two hundred and sixty six patients hospitalised in a dermatology university department (Henri-Mondor, Creteil) between November 1999 and April 2000 were enrolled in a prospective study. Clinical evaluation of pain intensity and evolution were studied using a visual analogic scale (VAS) pain score at presentation, during hospitalisation, and at discharge. Prescription and consumption of analgesics were also studied. RESULTS: Fifty-nine percent of the patients experienced pain. Eighty-four percent of them had mild or moderate to severe pain (VAS<7/10) and were relieved with non or mild opioid analgesics (discharge VAS<4/10). Sixteen percent of them had intense pain requiring morphine (VAS score >=7/10). CONCLUSIONS: Pain management is very important in dermatology. Every physician should know its principles since dermatologists play a crucial role in the patient's care. PMID- 12124510 TI - [Responsibility of food in exercise-induced anaphylaxis: 7 cases]. AB - INTRODUCTION: Exercise induced-anaphylaxis is a rare allergic disease. In most cases, the ingestion of a specific food within a maximum of 4 hours preceding physical activity is necessary to develop the anaphylactic reaction. CASE REPORTS: We report 7 cases of food-dependant exercise induced-anaphylaxis. The responsible foods were wheat (2 cases), corn, barley, shrimp, apple, paprika and mustard. The discovery of the food allergen and its systematic eviction 4 hours before the beginning of a physical effort led to the disappearance of clinical symptoms in all cases. DISCUSSION: The role of food in exercise-induced anaphylaxis is essential. The food in cause varies but our study confirms the predominant role of cereal food which is a less sensitizer in adults, except in a context of effort. The responsibility of this type of food consumed daily can explain partially the frequent delay before diagnosis of this allergic disease. PMID- 12124511 TI - [Male breast cancer in Morocco]. AB - INTRODUCTION: Male breast cancer is a rare disease, often of late presentation and poor prognosis. The aim of this work was to analyze the different clinical and therapeutic features for this disease in men. PATIENTS AND METHODS: This was retrospective study including 12 cases of male breast cancer seen in the Dermatology department of Casablanca between 1988 and 1999. RESULTS: The mean age of the patients was 60 years and the mean delay to consultation was 27 months. The skin was involved by tumor in 11 cases. Ulceration of the skin by tumor was seen in eight patients, and direct extension of the tumor into the nipple without ulceration was seen in three patients. Axillary lymph node involvement was seen in eight patients. Seven patients with invasive disease had metastases at distance. Treatment was usually surgical. Complementary treatment included radiotherapy, chemotherapy and/or hormonotherapy. DISCUSSION: Although breast cancer in men is far less common than breast cancer in women, it is associated with less favorable prognosis because diagnosis is usually made at an advanced stage. Concerted efforts must be made to educate both the public and health professionals, in order to make earlier diagnosis and thereby improve prognosis. PMID- 12124512 TI - [Radiation-induced temporary hair loss after endovascular embolization of the cerebral arteries: six cases]. AB - BACKGROUND: Intraoperative fluoroscopy imaging during coronaroplasty or transjugular intrahepatic portosystemic shunt may induce chronic radiodermatitis. Temporary hair loss is a peculiar form of radiodermatitis following endovascular surgery of the cerebral arteries. CASE REPORTS: Six patients (2 women, 4 men, age range: 27-47 years old) were seen for a solitary plaque of alopecia. In all of the cases, the plaque had appeared two weeks after a neuroradiologically guided embolization procedure. No other skin lesions were seen. Alopecia spontaneously and completely regressed within three to four months. However, it reappeared after a subsequent embolization (one case) but not after arteriographies (five cases). DISCUSSION: Five similar cases have been reported in the literature. Transient alopecia often occurs after neurologically guided endovascular surgery of the cerebral arteries. This side-effect is well known by neurosurgeons and thus, these patients are rarely referred to a dermatologist. Two differential diagnoses must be evoqued: alopecia aerata and anticonvulsant - induced alopecia. The role played as cofactor by carbamazepine which is a photosensitivant drug, is discussed. PMID- 12124513 TI - [Valaciclovir]. AB - Valaciclovir is an aciclovir pro-drug considerably improving its oral availability. Its antiviral activity in vivo is related to that of aciclovir, the principle target of which is the herpes virus. Following digestive absorption, valaciclovir is rapidly transformed into aciclovir. The mean absolute bioavailability of aciclovir is of 54.2% after a single oral dose of 1,000 mg of valaciclovir, i.e., a bioavailability 3 to 5-fold greater than after oral ingestion of aciclovir. The plasma pharmacokinetic profile of valaciclovir and aciclovir observed in volunteers infected by HIV is superimposable on that of healthy subjects. In elderly patients, exposure to aciclovir is enhanced, probably because of the alteration in glomerular filtration. In patients exhibiting agranulocytosis following poly-chemotherapy, the pharmacokinetic parameters are superimposable on those observed in healthy patients. In patients with hepatic failure, there appears no need to adapt the dose, since exposure to aciclovir does not appear altered. However, the dose of valaciclovir must be adapted to renal function. During the first-episode of herpes genitalis, valaciclovir, at the dose of 500 or 1,000 mg twice daily, is as effective as 200 mg of aciclovir five times per day. In recurrent herpes genitalis, 500 mg twice daily of valaciclovir is as effective as 1,000 mg twice daily or 200 mg five times a day of aciclovir. Valaciclovir prevents recurrence herpes genitalis with a dose-dependent effect, and doses of 500 and 1,000 mg/day are as effective as 400 mg twice daily of aciclovir. There are few studies on the efficacy of valaciclovir in the treatment of oro-facial herpes. In the treatment of herpes zoster in patients aged over 50, the principle benefit provided by valaciclovir at the dose of 1,000 mg twice daily, is the decrease in the percentage of patients presenting post-zoster pain and its duration. High doses of valaciclovir (8 capsules/day) provide efficient prevention of infections related to the cyto megalo-virus (CMV) in immunodepressed patients due to HIV infection or following renal transplantation. Tolerance to valaciclovir, like its active metabolite aciclovir, is generally good. Central neurological toxicity is frequently observed with high doses, but regresses on withdrawal. The official indications in France are the curative and preventive treatment of herpes genitalis infections, the prevention of post-zoster pain and the ocular complications of ophthalmologic herpes in immunocompetent adults, and the prevention of CMV infections after organ grafting. PMID- 12124514 TI - [Paget's extramammary disease of the axillae and perineum]. AB - INTRODUCTION: Paget's extramammary disease mostly affects genital, perianal and axillary regions. Whilst triple involvement has been described in Japanese patients, simultaneous lesions of both axillary regions and the inguinal area are exceptional among European patients. We report a case of triple Paget's extramammary disease in a Caucasian patient. CASE-REPORT: A 79-year-old male patient who developed a prostatic adenocarcinoma 3 years ago, was seen for an erythemato-squamous intertrigo of both axillary folds and the pubic area, present for 10 years, not diagnosed and resistant to topical treatments. Triple Paget's extramammary disease was confirmed by both histopathological and immunohistochemical investigations. No recurrence of the prostatic adenocarcinoma was observed. DISCUSSION: Since the first description of triple Paget's extramammary disease, 28 cases have been reported in Japan. To our knowledge, this is the first case observed in a Caucasian patient. The clinical features of axillary lesions are described as pigmented or depigmented plaques, sometimes lichenoid or erosive. For some Japanese authors, a biopsy is mandatory even in the absence of clinical lesions, since typical Paget cells can be found. Immunohistochemical studies reveal CK7 expression, the marker of choice for primary extramammary Paget's disease. CK7 - would suggest underlying regional internal malignancy as well as CK20 +. Despite the fact that the immunophenotype was CK7 +/CK20- the patient developed an evolving prostatic adenocarcinoma. Although various treatments are described in the literature, surgical excision remains the first line treatment whenever possible. PMID- 12124515 TI - [Eosinophilic-like erythema: a clinical subset of Wells' eosinophilic cellulitis responding to antimalarial drugs?]. AB - BACKGROUND: Limits of Wells'syndrome, a nosologically enlarged entity defined after classic eosinophilic cellulitis, are imprecise. We report on a new "frontier" case, remarkable in its clinical pattern and its high sensitivity to antimalarial drugs. CASE REPORT: A 35-year-old woman was referred for evaluation of recurrent inflammatory, figurated and often annular lesions, mainly located on the trunk and the proximal parts of the limbs, associated with significant peripheral eosinophilia and featuring dense perivascular infiltrates throughout the dermis with abundant eosinophils but without any "flame figure". These lesions responded dramatically to protracted treatment with hydroxychloroquine. DISCUSSION: The relationship of this affection to Wells' syndrome is discussed along with its possible similarities to the sparse reports of so-called eosinophilic annular erythema. PMID- 12124516 TI - [Allergic contact dermatitis to the Comfeel hydrocolloid dressing]. AB - INTRODUCTION: Allergic contact dermatitis is frequent in patients with chronic leg ulcers. However, it rarely occurs with modern wound dressings and is exceptional with hydrocolloids. CASE REPORT: A 66-year-old woman was treated for a leg ulcer with the Comfeel plus(R) transparent hydrocolloid dressing for two months. She developed a pruriginous, erythematous and vesiculous dermatitis under the hydrocolloid plaques. Patch tests for the Comfeel plus(R) transparent hydrocolloid, the Comfeel plus(R) hydrocolloid, balsam of Peru and epoxy resin were positive. Only the positive test for the Comfeel plus(R) transparent hydrocolloid was clinically pertinent. The histological examination of the positive test was suggestive of eczema. DISCUSSION: To our knowledge, allergic contact dermatitis to Comfeel plus(R) hydrocolloid dressings has not been reported. Most previous studies which included systematic patch-testings in patients with leg ulcers showed high sensitization rates for various allergens, but no allergy to hydrocolloids. Only isolated cases of allergic contact dermatitis to another hydrocolloid (Duoderm E(R)) have been reported. Our case report shows that allergic contact dermatitis is a possible side-effect of Comfeel plus(R) hydrocolloid dressings. However, it seems exceptional. Since the patch-tests failed to identify the constituent responsible for this allergy in our observation, comprehensive allergologic investigations should be repeated in further cases. PMID- 12124517 TI - [Mycobacterial bovis BCG cutaneous infections following mesotherapy: 2 cases]. AB - INTRODUCTION: Infectious complications following mesotherapy are usually due to ordinary bacteria or atypical mycobacteria. We report two new cases of mycobacterial bovis BCG infections following mesotherapy. To our knowledge only one case has already been reported. CASES REPORTS: A 52 year-old woman developed vaccinal MERIEUX BCG cutaneous abscesses following mesotherapy. Identification was made by a novel class of repeated sequences: Mycobacterial interspersed repetitive units. Despite prolonged anti-tuberculous therapy, complete remission was not obtained and surgical excision was performed. The second case was a 49 year-old man who developed a mycobacterial bovis BCG cutaneous abscess (Connaught) after mesotherapy, the regression of which was obtained with anti tuberculous therapy. DISCUSSION: The severity of these two mycobacterial infections following mesotherapy illustrate the potential risks of mesotherapy. Identification is possible by molecular biology techniques (PCR and sequencing). The origin of this infection is unclear and therapeutic decision is difficult. Some authors recommend anti-tuberculous therapy but surgical excision may be necessary as in our cases. PMID- 12124518 TI - [Cranial fasciitis of childhood]. AB - INTRODUCTION: A nodule of the scalp in a child of less than eleven years should evoke a cranial fasciitis among other serious diagnoses. OBSERVATION: A four month old infant had a firm and pink nodule at the left parietal level, exhibiting a slow growth since two months. It was excised. The pathologic sample showed spindle-shaped cells within a myxoide matrix, with a strong reactivity for smooth muscle actin (immunohistochemical analysis). Diagnosis of cranial fasciitis was made. Due to the results of pathology, it was possible to rule out the diagnosis of sarcoma, therefore, no complementary work-up was performed. Evolution was favorable. DISCUSSION: Cranial fasciitis is a diagnosis to be considered when confronted with a firm nodule of the scalp in a infant or a young child, with or without bone involvement. This is a benign lesion but worrying pathological signs may exist, making diagnosis of benignity difficult. Exeresis of the lesion, even incomplete, protects the child from possible recurrence. Evolution is always good. PMID- 12124519 TI - [Noninvoluting congenital hemangioma: 2 cases]. AB - INTRODUCTION: Hemangiomas (or immature hemangiomas) are characterized by a stereotyped 3-phase evolution: proliferation, stabilization and regression. The rare congenital hemangiomas are present at birth and regress spontaneously more rapidly. However, certain congenital hemangiomas, described recently as "noninvoluting congenital hemangiomas", evolve differently and do not regress. We report two cases. OBSERVATIONS: Two adolescents aged 16 and 17 were born with a congenital hemangioma. Its evolution during childhood was marked by the progressive collapse of the lesion and lightening of the skin. After stabilizing, a round, centrally involuted lesion persisted with numerous telangiectasia on the surface and peripheral varicosities. The lesion was hot but no pulsation or murmur were observed. Doppler sonography revealed a rapid flowing lesion, limited to the cutaneous areas. Magnetic resonance imaging, conducted in one case, showed a hypersignal area limited to the skin. DISCUSSION: Noninvoluting congenital hemangiomas can be differentiated from classical congenital hemangiomas by their partial regression, stereotyped clinical aspect and a certain activity after stabilization. All the clinical, histological and imaging data support a vascular malformation with proliferative component, rather than a true hemangioma. Whenever possible, treatment is surgical. Knowledge of the existence of this type of angioma is important, and the dermatologist should be careful when reassuring parents of children presenting with congenital hemangioma. PMID- 12124520 TI - [S-100B and MIA (melanoma inhibiting activity) serum proteins: a prospective study of their value as melanoma progression markers]. PMID- 12124521 TI - [When a dermatologist and a psychologist share a consultation on atopic dermatitis]. PMID- 12124522 TI - [Dermato-psychiatric consultation]. PMID- 12124523 TI - [Vegetating lesions around a stomy]. PMID- 12124524 TI - [Tumefaction of the back of the hand]. PMID- 12124525 TI - [Multiple perilabial pigmented spots]. PMID- 12124527 TI - [Prevention and treatment of bedsores in adults and the elderly]. PMID- 12124526 TI - [Eczema on contact with molecules of the paraphenylenediamine family in the workshop]. PMID- 12124528 TI - [Tobacco smoking and antimalarials]. PMID- 12124529 TI - [Toxiderma: the origin of the concept]. PMID- 12124530 TI - [Developmental lumbar stenosis in eleven French Antilles patients]. AB - PURPOSE OF THE STUDY: Developmental lumbar stenosis is a rare entity, exceptionally described in the literature. No study has been directly devoted to this condition. The purpose of the present study was to examine specific features, particularly clinical and anatomic expression, observed in a series of operated patients. MATERIAL AND METHODS: Eleven patients from the French Antilles were treated for developmental lumbar stenosis between 1996 and 2000. The Verbiest criteria were used to define canal narrowness. Signs of degeneration and presence of discal herniation were exclusion criteria. Epidemiological and clinical data were collected for the 11 patients. The degree of sagittal stenosis (fixed diameter at the bone level and mobile diameter at the discal level) was measured on computed tomography images. Transverse stenosis was determined by measuring the interpedicular and interapophyseal distances. Lateral stenosis was determined by measuring the depth of the recessus. RESULTS: These patients were young (mean age 42.4 years). Most of the clinical signs were monoradicular. Discal level stenosis predominated, generally at level L4-L5. It was generally central and lateral, sagittal and transverse. The interpedicular distance was the only diameter that remained within normal limits. Soft tissues (yellow ligaments and joint capsules) played an important role in the stenosis. DISCUSSION: The rare reports of developmental lumbar stenosis describe decompensated stenosis due to discal herniation in the adolescent. Developmental lumbar stenosis is considered to be a genetic disease and its particular high frequency in the French Antilles favors this hypothesis. The stenosis results from bony (short pedicles, hypertrophic lateral masses) and ligament (hypertrophy of the yellow ligament and joint capsules) structures. CONCLUSION: Developmental lumbar stenosis produces a global (sagittal, transverse, central, lateral) narrowing of the lumbar canal where soft tissue structures apparently play a greater role than usually thought. A prospective study examining the impact of ethnic origin is required to analyze the genetic hypothesis. PMID- 12124531 TI - [Sagittal alignment of the spine: comparison between soccer players and subjects without sports activities]. AB - PURPOSE OF THE STUDY: The purpose of this study was to compare the sagittal alignment of the spine in a population of soccer players and a population of volunteers with no sports activities in order to assess the effect of sports activities on stress fractures of the isthmic region. MATERIAL AND METHODS: The group of athletes included 31 soccer players and the volunteer group 47 subjects with no sports activities selected from a 131 subjects database. The two groups were matched for sex and age. All athletes were licensed members of a soccer club and participated in regular sports activities at a moderate level (at least 4 hours per week for at least 2 years). Plain radiographs of the entire spine (lateral view) in the same reference position were obtained for all subjects. The following parameters were recorded: thoracic kyphosis, lumbar lordosis, sacral tilt, anteversion of the pelvis, sagittal alignment. The following morphology data were also recorded in this exclusively male population: weight, height, arm spread. RESULTS: Considering the other radiographic parameters, the sagittal alignment of the spine was comparable between the two groups. However, the statistical analysis demonstrated that the distribution of the spinal and pelvic parameters was different between the athletic and the non-athletic subjects. In athletes, spinal alignment was achieved by a less pronounced thoracic kyphosis and a more pronounced angle, sacral tilt and lumbar lordosis. DISCUSSION AND CONCLUSION: The particular spinal morphology observed in soccer players is comparable with that described in the literature for patients with or treated for spondylolisthesis with isthmic lysis. PMID- 12124532 TI - [Pelvic obliquity and scoliosis in non-ambulatory patients with cerebral palsy: a descriptive study of 234 patients over 15 years of age]. AB - PURPOSE OF THE STUDY: Children with cerebral palsy who cannot walk have an oblique pelvis and scoliosis. There is a certain degree of controversy in the literature on the best way to manage this difficult situation. We present a descriptive analysis of a population of non-ambulatory adults with cerebral palsy in order to formulate hypotheses concerning the factors determining scoliosis. MATERIAL AND METHODS: This descriptive cross-sectional study was conducted in 234 patients aged over 15 years who had cerebral palsy and could not walk. Physical examination and an x-ray of the pelvis and spine in the reclining position were obtained for all patients. The following variables were recorded: luxation and subluxation of the hip, spontaneous deviation attitude, ability or not to turn over in bed, pelvic obliquity, history of bone surgery, defective hip abduction. The statistical analysis accounted for laterality and pelvis obliquity to the scoliosis convexity and the laterality of the hip excentration. RESULTS: Scoliosis was observed in 66.2% of the patients; it was more than 60 degrees in 34.5%. Two basic groups were distinguished: thoracolumbar scoliosis (41.6%) and lumbar scoliosis (41.6%). The prevalence of oblique pelvi was 59.9% with important difference by side: 31.6% right oblique and 68.4% left oblique pelvi. We were unable to find any relationship between the side of the pelvic obliquity and the side of the scoliosis convexity, the side of the hip excentration, or the deviation attitude, but the deviation attitude appeared to be a risk factor for pelvic obliquity, which itself was a risk factor for excentration, which was a risk factor for scoliosis. DISCUSSION: Scoliosis is an important problem in this population. Hip luxation is a direct risk factor for scoliosis, but the deviation attitude and pelvic obliquity are intermediary stages. The prevalence of oblique pelvi was greater on the left than the right. This finding should be confirmed in other series before hypotheses can be formulated concerning this difference. PMID- 12124533 TI - [Internal fixation of proximal humerus fracture by "palm tree" pinning]. AB - PURPOSE OF THE STUDY: Fractures of the proximal humerus are increasingly frequent. Conservative treatment is most often proposed, but surgery must be performed when the displacement is significant and/or when the fracture is unstable. Osteoporosois and comminution are two essential elements for deciding on the surgical technique. MATERIAL AND METHODS: This retrospective study included 31 patients who underwent pinning from the deltoid V according to the Kapandji procedure. There were 19 females and 12 males. Mean age was 61 years. There was a fracture of one of the tuberosities in 12 cases and significant metaphyseal comminution in 8. RESULTS: Mean follow-up was 26 months. Outcome was excellent or good in 22 cases (70.9%), fair in 4 (12.9%), and poor in 5 (16.2%). Fifteen complications were noted (48.4%): material displacement 8 cases, reflex sympathic dystrophy 3 cases, radial nerve palsy 2 cases, head osteonecrosis 1 case, and humeral fracture at the site of insertion of the K wires 1 case. DISCUSSION: At the present time, there is no consensus for the surgical management of fractures of the proximal humerus, including proximal metaphyseal fracture with or without fracture of one of the tuberosities. Closed reduction and pinning is not really an invasive procedure, and does not injure the rotator cuff. Surgical approach at the level of the deltoid V according to the Kapandji technique avoids elbow pain and stiffness. The procedure requires a fluoroscan and experience to obtain satisfactory divergence of the K wires in the humeral head, an essential technical point. This procedure cannot be recommended for elderly patients whose bone quality is too poor to obtain good fixation of the K wires. PMID- 12124534 TI - [Painful or unstable shoulder after coracoid transfer: result of surgical treatment]. AB - INTRODUCTION: The purpose of this study was to investigate the results of revision surgery for complications related to previous coracoid transfer for recurrent anterior instability of the shoulder. MATERIALS AND METHODS: Seventeen patients with previous surgery for anterior shoulder instability underwent a new surgical procedure, because of recurrent instability in 10, and painful shoulder with limitation of motion in 7. A soft tissue procedure (Bankart and/or capsuloplasty) was performed in the 10 unstable shoulders, and a joint debridement with removal of the coracoid transfer in the 7 painful shoulders. The subscapularis was found to be normal in only 2 cases, fibrotic in 11, thin in 3, and teared in 1. The interval between the initial procedure and the revision surgery was eleven years on average. RESULTS: At an average of 21 months follow up, the patients were evaluated according to the Duplay scoring system. A radiographic analysis was also performed for all the patients, and a CT examination for fourteen. The results were good or excellent for 11 patients (70% in the soft tissue procedure group, and 57% in the debridement group with removal of the coracoid transfer), fair for 4, and poor for 2. Clinical evaluation of the subscapularis showed a lag of muscle function in 10 patients. Strength in internal rotation was 3.3 kg lesser in the operated shoulder compared to the opposite side. CT-examination showed that 4 patients presented a significantly fatty degeneration of the subscapularis. Finally on radiographic examination, osteoarthritis was present in 9 patients.The most important preoperative factor that affected the final results was the number of previous surgical procedures. DISCUSSION: Recurrent instability, problems related to the bone graft or ostheosynthesis material, osteoarthritis, and neurological damage can complicate a coracoid transfer procedure. Our study shows that this procedure can also induce irreversible damage to the subscapularis muscle. CONCLUSION: Revision surgery for complications related to coracoid transfer for anterior shoulder instability is a challenging procedure. Only 2/3 of patients achieved excellent or satisfactory results. Patients with recurrent instability had better results than those with painful impingement and or osteoarthritis. The high rate of late osteoarthritis and irreversible damage of the subscapularis muscle remain sources of concern. PMID- 12124535 TI - [Reproducibility of CT scan evaluation of muscular fatty degeneration. Intra- and interobserver analysis of 56 shoulders presenting with a ruptured rotator cuff muscles]. AB - PURPOSE OF THE STUDY: This study was undertaken to determine the reproducibility of measurements of fatty degeneration of the rotator cuff using computed tomography (CT). MATERIAL AND METHODS: Fifty-six patients who had undergone surgery for rotator cuff tear were included in this retrospective study. The extent of fatty infiltration was evaluated on CT scans with soft tissue windows in all 56 shoulders using a five-stage scoring system described by Goutallier. Five independent observers made the assessments. The same operation was repeated one month later to test intraobserver agreement. Four parameters were recorded: fatty infiltration of three muscles (supraspinatus, infraspinatus, subscapularis), and overall fatty infiltration grading. Interobserver variability was determined for each parameter using the intercorrelation coefficient (a test of reproducibility of quantitative measurements). RESULTS: The most reproducible measurement was the overall fatty infiltration grade. For this parameter, interobserver agreement was good with an intercorrelation coefficient of 0.75. The interval of confidence was +/- 0.5. Intraobserver agreement depended on the observer's level of experience. It was good for overall fatty infiltration grade assess by three senior observers (r=0.78) and moderate for two junior observers. CONCLUSION: The overall fatty infiltration grade is a reproducible parameter that should be used to evaluate the degree of fatty infiltration as the safety margin of this value (graded 0 to 4) is about 0.5. Fatty infiltration of a torn cuff would not be the only criterion to improve indications for treatment of rotator cuff tears. PMID- 12124536 TI - [Deltoid muscle flap for massive rotator cuff tears: 41 cases with a mean 7-year (minimum 5 year) follow-up]. AB - PURPOSE OF THE STUDY: The aim of this study was to assess outcome after deltoid muscle flap repair of massive rotator cuff tears. We examined functional and radiological results at least five years after surgery. MATERIAL AND METHODS: We reviewed 41 shoulders operated by three senior surgeons (MC, DK, HT). None of the patients were lost to follow-up. The global Constant score was used for pre- and postoperative functional assessment. Three groups were distinguished by preoperative active flexion (group I<90 degrees, group II 90 degrees -120 degrees, group III > 120 degrees ). AP, double oblique (3 rotation views to measure the subacromial space), and Lamy lateral radiographs were obtained in all patients. Shoulder anatomy was evaluated at last follow-up in eight patients: magnetic resonance imaging (MRI) because of persistent pain in one patient and ultrasonography performed by one radiologist (NC) in seven patients. RESULTS: The study population included 26 men and 15 women, mean age at surgery 59 years (42 78, 8). Mean follow-up was 7 years (5-8.5, 0.9). In the coronal plane, there were no distal tears, the stump was in an intermediate position in 7 cases (17%) and retracted to the glenoid in 34 (83%). In the sagittal plane, the supraspinatus exhibited a full thickness tear in all cases. The tear extended anteriorly or posteriorly in all cases. Thirty-eight patients (92%) were satisfied at last follow-up; their global Constant score had improved from 37 to 62 points. Mean anterior flexion improved from 113 degrees to 148 degrees and flexion force from 1.3 kg to 2.9 kg. When preoperative flexion was less than 90 degrees (11 cases), mean gain was + 89 degrees. Inversely, 7 of the 18 patients with flexion > 120 degrees lost a mean 40 degrees at last follow-up. Twenty-seven patient were reviewed at 12 and 89 months: pain relief and force were maintained. The subacromial space, measured in 88% of the cases, was 7.3 mm preoperatively and 5.5 mm at last follow-up. The subacromial space narrowed in 20 patients (56%); none of the patients exhibited an improvement. The flap was explored by ultrasonography in seven patients and by MRI in one: the flap was continuous in 50% and measured more than 4 mm in thickness. Reviews at 12 then 89 months demonstrated good maintenance of pain relief and progression of active flexion and force. DISCUSSION AND CONCLUSION: This long-term study confirms the usefulness of the deltoid flap for the treatment of full thickness massive tears of the rotator cuff. The flap provides persistent pain relief and good function and force. This technique should be discussed for young patients in good physical condition when preoperative imaging demonstrates and irreparable alteration of the tendinomuscular structures (supraspinatus retraction, fatty degeneration, severe amyotrophy). The technique is particularly useful when preoperative flexion is less than 90 degrees. Although the population size is too small for statistical analysis, indications for deltoid flap repair should probably be limited to tears involving at most two tendons and sparing the subscapularis. PMID- 12124537 TI - [Results of an inverted shoulder prosthesis after resection for tumor of the proximal humerus]. AB - PURPOSE OF THE STUDY: Treatment of primary bone tumors of the proximal humerus is a major challenge, both in terms of oncological cure and limitation of functional handicap. Extracorporal radiation and reimplantation have provided satisfactory results for the treatment of bone tumors and the inverted shoulder prosthesis has be found to be effective in treating rotator cuff deficiency. We therefore assessed patients treated with a combination of these two treatments in order to search for a better functional outcome. MATERIAL AND METHODS: Conservative resection of the tumor with reconstruction using an inverted shoulder prosthesis was performed in six patients with a tumor of the proximal humerus of type Ia or Ib in the Malawer classification or type S3-S4-S5 in the Musculoskeletal Tumor Society classification. Patients whose tumor involved the deltoid or the glenohumeral joint were not considered for this procedure. The series included four men and two women (4 left and 2 right shoulders) aged 51 years on the average at the time of surgery. RESULTS: Mean follow-up was 12 months. The mean age- and sex-adjusted Constant score was 74% (70-85%). The patients were generally satisfied and were able to continue their daily activities. There were two displacements, one of which was diagnosed immediately and reduced without anesthesia. An abduction pad was worn for six months after reduction. The post operative period was uneventful. No local recurrence has been observed. DISCUSSION: Our results compared favorably with other therapeutic options for resection-reconstruction reported in the literature: functional outcome was better and more predictable, at least in the short term; oncological results were satisfactory although the follow-up is short, particularly for the three primary malignant tumors. It is known however that the long-term oncology result depends on the quality of the resection and the efficacy of the extracorporeal radiation. Shoulder motion outcome was the least satisfactory in patients who experienced peri-operative complications. We were unable to find any relation between less satisfactory functional outcome and the number of surgical procedures for local recurrence before extracorporeal radiation and arthroplasty (deltoid dystrophy). Normal deltoid function appeared to be crucial for optimal functional outcome with this prosthesis which allows sacrifice of the rotators. The rate of complications was low and recovery of active shoulder movement was more rapid. Despite the short-term nature of these results, and despite the risk of loosening, the inverted shoulder prosthesis appears to be an interesting therapeutic option for these patients. PMID- 12124538 TI - [In vivo measurements of shoulder movements by a goniometer with six degrees of freedom: preliminary study]. AB - PURPOSE OF THE STUDY: The purpose of this work was to develop a goniometer with six degrees of freedom (6DOF) to achieve global measurement of the main movements of the shoulder and to examine the reproducibility of the measurements obtained. MATERIAL AND METHODS: Movement amplitudes, inter- and intra-observer reproducibility of the methodology and the device were tested in a population of ten young subjects with asymptomatic shoulders. RESULTS: Maximal amplitudes obtained for each movement were: abduction 111.8 +/- 11 degrees; flexion 127.9 +/ 12.7; external rotation 86.3 +/- 14 degrees; free elevation 129.3 +/- 9 degrees. The curves traced in the plane of rotation and rotation were plotted against elevation. Comparison between amplitudes on the right and left were not significantly different. DISCUSSION AND CONCLUSION: This preliminary study demonstrated the potential contribution of a 6DOF-goniometer for a 3D assessment of shoulder movement. Measurements made when positioning the goniometer demonstrated good reproducibility. Our data are similar to others reported in the literature. The plane of the scapula and the position in physiological internal rotation, as described in the literature, were also observed. Studying shoulder movements, particularly their association, should provide a new means of assessing function in patients with complex disease of the shoulder joint. The device should be helpful in obtaining an objective measurement of the shoulder kinetics that would be useful for patient follow-up and assessment of corporal damage. PMID- 12124539 TI - [Fresh fractures of the radial head: results with the Judet prosthesis]. AB - PURPOSE OF THE STUDY: Communitive fractures of the radial head are a therapeutic challenge when fixation is not possible. Secondary sequelae including ulnar valgus, ascension of the radius, osteoporosis of the humeral condyle and biomechanical impairment of the elbow cannot be avoided with resection or the Swanson prosthesis. The Judet prosthesis, with its floating cup, is a technically attractive solution, but the question is whether it can avoid the secondary effects observed with resection or the Swanson prosthesis. MATERIAL AND METHODS: We have used the Judet prosthesis since August 1995 in 16 patients. These patients had Mason type IV (Johnston modification) lesions of the radial head in 14 cases and type III lesions in 2. Associated injuries included: Monteggia fracture in 2, open fracture-dislocation in 1, fracture of the radial neck associated with fracture of the lower radius in 1, and dislocation of the elbow in 2. Preoperatively, 7 of the 16 patients had an osteocartilaginous injury of the humeral condyle. The Judet prosthesis was implanted after resection in 3 patients, after osteosynthesis of the radial head in 3 others, and as the first line treatment in 10. Postoperatively, 13 of the 16 patients were given a unique 7 Gy radiation to prevent ossification as well as nonsteroidal antiinflammatory drugs (indometazin 50 mg b.i.d.) for 3 weeks. None of the patients were immobilized. RESULTS: Mean follow-up for the 16 patients was 19 months (12-45). Deficient extension persisted in 5 patients (mean 5 degrees ). Average flexion was 128 degrees; two patients were limited to 100 degrees. Average pronation was 77 degrees, and average supination 79 degrees. Muscle force was 10% weaker than the healthy side. Sagittal and frontal stability in valgus was preserved in 14 patients. Two patients had a frontal instability with minimal valgus related to a minor insufficiency of the medial collateral ligament. According to the Radin and Riseborough classification, outcome was good in 7, fair in 6 and poor in 3. According to the Morrey classification, outcome was excellent in 2, good in 12, fair in 1 and poor in 1. Radiologically, there were no cases with ulnar valgus, humeral condyle osteoporosis, ascension of the radius, or subluxation of the distal radioulnar joint. The prosthesis loosened in one case without clinical expression. DISCUSSION: Our results with the Judet prosthesis were much better than those reported in the literature for resection and Swanson prosthesis. PMID- 12124540 TI - [Kudo non-constrained elbow prosthesis for inflammatory and hemophilic joint disease: analysis in 30 cases]. AB - PURPOSE OF THE STUDY: We analyzed retrospectively 30 Kudo non-constrained elbow prostheses to determine: 1) functional outcome and mobility, 2) frequency of loosening and any complications. MATERIAL AND METHODS: From 1992 to 1998, 30 Kudo total elbow arthroplasties were performed in 29 patients, mean age 55 years. Mean follow-up was 36 months. These patients had severe joint disease: rheumatoid arthritis for 24, psoriatic arthritis for 2, and hemophilic arthritis for 3. The 29 patients experienced severe pain before surgery. RESULTS: At review, 21 elbows were pain free and the 9 others had only occasional pain. Among these 9 elbows, 3 exhibited a rupture of the humeral implant; one had already been revised but remained painful. One patient had a stiff painful elbow after reflex dystrophy and five others had pain but no other complication. Twenty-six patients were satisfied or very satisfied. Three patients were unsatisfied because of the humeral implant fracture. Mean mobility at last follow-up was: 128 degrees flexion, -35 degrees extension, 72 degrees pronation, and 74 degrees supination. Mean gain in flexion-extension was 15 degrees and mean gain in pronosupination was 3 degrees. Pronosupination was greater than 100 degrees except for two patients. There was one immediate post-operative dislocation with failure of prolonged orthopedic treatment after reduction; this patient underwent revision reconstruction with repair of the ulnar collateral ligaments (plasty of the medial collateral ligament with a synthetic ligament). Painful movement of the radial stump was observed with one Kudo prosthesis and required resection to achieve cure. In all, there were 3 fractures of the Kudo I prosthesis at the junction of the trochlea and the humeral stem. Among these patients, one underwent revision due to persistent pain, and two others with currently acceptable symptoms are awaiting revision. At last follow-up, we had: 1 ulnar loosening associated with cortical thinning facing the end of the ulnar implant that had migrated and showed a circular lucent line measuring > 1 mm and progressing; 9 unique ulnar lucent lines measuring<1 mm without progression at the proximal part of the implant (6 at the bone-cement interface and 3 at the bone-implant interface); 3 humeral radiolucent lines (<1 mm without progression) on the distal part of the Kudo II humeral stems corresponding to a zone without surfacing. We also observed 13 cases of incomplete ossification between the humerus and ulna and among these 13, 7 elbows had amplitudes of less than 100 degrees. DISCUSSION AND CONCLUSION: Elbow arthroplasty can restore a painless joint and maintain or improve elbow motion. The procedure is indicated when the joint disease impair daily life activities. Final mobility basically depends on the preoperative mobility. The bone stock remains the greatest problem with these resurfaced prostheses. The GUEPAR elbow prosthesis would appear to be more adapted due to the reconstruction of the trochlea. Resection of the radial head is a source of instability for elbow prostheses and should lead to the design of three-compartment prostheses. PMID- 12124541 TI - [Reposition flap techniques in fingertip amputations: 6 cases]. AB - PURPOSE OF THE STUDY: The purpose of this study was to evaluate an alternative procedure for amputations distal to the distal interphalangeal joint: the reposition flap. MATERIALS AND METHODS: The reposition flap was used for 6 patients who underwent fingertip amputations in an emergency setting. Pulp was excised on the amputated segment and the remaining bone and nail bed were reattached to the proximal stump with a Kirschner wire. The pulp was reconstructed with a local advancement and sensitive flap. The patients were aged 18 to 44 years and had been victims of work accidents. All refused finger shortening. RESULTS: The fingers showed good scarring and good trophicity. Two point discrimination was 6 mm. Bony fusion was constant but all distal interphalangeal joints remained stiff. Cosmetic results were correct except for two cases of claw nail formation. DISCUSSION: Fingertip amputations have been widely reported. Methods have varied from directed scarring to partial toe transfer. These situations present two types of challenge: insensitivity of the volar aspect or an overly sensitive pulp; cosmetic presentation and function of the dorsal aspect due to the complex role of the nail. Distal reimplantation remains the best technique, but the reposition flap offers an interesting alternative in case of failure or for patients who do not accept finger shortening. The advantage of the reposition flap is that it preserves finger length and the nail. Work stoppage and intolerance to cold can be an inconvenience due to the long time required for wound healing. PMID- 12124542 TI - [Mechanical complications of total shoulder inverted prosthesis]. AB - Our series of inverted prosthesis included 5 patients with a mean age of 73 +/- 6 years. In 4 cases, the implant was performed as a surgical revision. The follow up was 81 +/- 15 months. Three shoulders were pain free whereas two caused a dull pain after a free interval due to mechanical complications. The mean active elevation was 72 degrees while external rotation was - 2 degrees. The adjusted Constant score passed from 32 to 60. In case of complications, the score dropped to 32. Mechanical complications were important with in one case, an unscrening of the glenosphere and in two cases, a loosening of the glenoid prosthesis. This last and major complication occurred 6 years after surgery and was promoted by the occurrence of a progressive bone erosion in the scapula. This gap represented an attempt to accomodate the medial part of the humeral prosthesis under the scapula when the arm is at rest or in adduction. The concept of an inverted prosthesis is attractive and this implant remains one of the options in cuff-tear arthropathy. Our results were not as good as those reported by others but most of ours patients had been already operated before. The occurrence of an osseous gap on pilar of scapula may lead to failure of this prosthesis. This gap remains a threath as it can progress and as such warrants a design alteration of the prosthesis. PMID- 12124543 TI - [The superolateral approach for shoulder prosthesis]. AB - We describe a superolateral approach to the shoulder for implantation of total shoulder prostheses or humeral prostheses. The advantages of this approach include preservation of the supraspinatus tendon and an excellent exposure of the posterior part of the glenoid cavity. We illustrate this approach with three clinical examples: total shoulder arthroplasty with reconstruction of the posterior part of the glenoid using a screwed autograft for central degeneration with posterior wear of the glenoid, intermediate arthroplasty for excentric degeneration with irreparable rotator cuff tears, and simple humeral arthroplasty with bone suture of the tuberosities for cephalotuberosity fracture. PMID- 12124545 TI - Peripheral pulmonary atelectasis and oxygentation impairment following coronary artery bypass grafting. AB - BACKGROUND: Severe pulmonary oxygenation impairment occurred in some patients with pleurotomy during the harvest of the internal mammary artery graft followed by coronary artery bypass grafting (CABG). Peripheral pulmonary atelectasis in the postoperative chest X-ray was detected in these patients. We studied the efficacy of intraoperative positive end-expiratory airway pressure (PEEP) therapy for the prevention of postoperative pulmonary oxygenation impairment. METHODS: The pleural cavity was intraoperatively opened in 40 patients with solitary CABG procedure performed during 5 years since January 1992. These patients were divided into two groups. Intraoperative PEEP therapy, which is initiated just after pleurotomy, was not used in 32 patients before May, 1996 (control group) and used for recent 8 patients with pleurotomy (PEEP group). The mean age of patients was 60 years old in the control group and 68 in the PEEP group. RESULTS: Respiratory insufficiency (A-aDO2 >400 mmHg and RI >1.5) was detected in 6 patients in the control group. Three out of these 6 patients required long-term mechanical respiratory support over a week. No respiratory insufficiency occurred in patients of the PEEP group. Values of PaO2, A-aDO2, respiratory index and shunt ratio were significantly worse in the control group than in the PEEP group. CONCLUSIONS: In conclusion, PEEP therapy may prevent pulmonary atelectasis and oxygen impairment after CABG. PMID- 12124546 TI - Clinical experience with Sorin Bicarbon valve in patients with tight mitral valve stenosis and elevated pulmonary hypertension (early and mid-term results). AB - BACKGROUND: The results of mitral valve replacement (MVR) with Sorin mechanical valves in patients who had tight mitral stenosis with high pulmonary artery pressure were reviewed. METHODS: During a period of two years, from August 1998 to May 2000, a mitral valve replacement with a Sorin Bicarbon mechanical valve was performed in 51 patients with a diagnosis of tight mitral stenosis associated with severe pulmonary hypertension (preoperative mean systolic pulmonary artery pressure was 72+/-12 mmHg, range from 60 to 105 mmHg). There were 37 women and 14 men; mean age was 47.2+/-12 years. Forty-eight patients (94.12%) were in NYHA functional class III or IV. All the patients discharged from the hospital were submitted to a clinical follow-up program. A 100% follow-up was obtained with a mean of 12.6+/-6.4 months (range 2 to25 months). RESULTS: Operative mortality was 3.9%, 2 patients who had concomitant CABG died due to low cardiac output. Twelve patients (23.5%) needed an inotropic pharmacological support during the postoperative time. In one patient a re-exploration for bleeding was necessary, and in another one a cerebrovascular accident occurred 3 days after the operation. After 6 months, one patient was reoperated on because of mechanical valve dysfunction due to pannus formation. All survivors underwent a postoperative echocardiographic assessment. The systolic PAP decreased from a mean preoperative value of 72+/-12 mmHg to 39.9+/-12 mmHg. NYHA functional status significantly improved and 86% of the patients were in NYHA functional class I or II. CONCLUSIONS: The mitral valve replacement with Bicarbon mechanical valve prosthesis shows excellent results in patients with mitral valve stenosis associated with a severe pulmonary hypertension. PMID- 12124547 TI - Aprotinin mediated antioxidant effect in Cardiosurgery with mechanical cardiorespiratory support (CMCS). AB - BACKGROUND: Aprotinin has been used in cardiosurgery as a hemostatic agent. Considering the implication of oxygen reactive species and proteases in the pathogenesis of Systemic Inflammatory Response Syndrome, we hypothesized that aprotinin may exert an antioxidant effect. This work was designed to evaluate the antioxidant capacity of aprotinin in vitro and in vivo in child patients undergoing cardiosurgery with mechanical cardiorespiratory support. METHODS: Colorimetric techniques and chemiluminiscent emission assays. A blind controlled clinical trial was performed with a control (G-1, n=14, without aprotinin) and treated with aprotinin (G-2, n=12) groups (both assessed by medical decision) of child patients undergoing cardiosurgery with mechanical cardiorespiratory support. Blood samples were taken at: T-0 (induction of anesthesia), T-1 (10 minutes after begining of perfusion), T-2 (5 minutes after anoxic heart arrest), T-3 (ending operation) and T-4 (24 hours after operation). RESULTS: We proved that aprotinin has no hydroxyl radical, superoxide anion nor H2O2 scavenger capacity as well as its capacity for inhibiting in vitro activated-leukocyte chemiluminiscence. Malonildialdehyde levels were higher in G-1 than G-2 with the greatest difference at T-2 (7.2+/-3.6 nmol/ml in G-1 vs 4+/-1.65 in G-2). Phospholipase A2 activity showed a tendency of higher values in G-1 than G-2 although there was no statistical significance. Uric acid concentration was greater in G-2 at T-1, T-2, T-3 and T-4 than G-1 and catalase activity was higher in G-2 at T-0, T-2 and T-3 than G-1 with noteworthy difference only at 5 minutes after anoxic heart arrest. Low cardiac output, arrhythmias and sudden death in the early postoperative phase were less frequent in the treated group. CONCLUSIONS: These results suggest that aprotinin exerts a primary antioxidant activity and its protective effects in cardiosurgery seem to be associated with reduction of systemic oxidative stress. PMID- 12124548 TI - Effect of a human urinary protease inhibitor (Ulinastatin) on respiratory function in pediatric patients undergoing cardiopulmonary bypass. AB - BACKGROUND: We performed this prospective randomized study to determine the effect of a human urinary protease inhibitor (Ulinastatin) on respiratory function in pediatric patients undergoing cardiopulmonary bypass. METHODS: Twenty two children were included in this study. They were randomly allocated to 1 of the following groups; a control group (n=11) or a Ulinastatin group (n=11) in which patients received 5000 U/kg of Ulinastatin. Arterial blood samples were obtained before cardiopulmonary bypass (CPB), immediately after CPB, and 30 min after protamine administration, as well as 3 hours after and 18 hours after CPB, and Interleukin-8 and neutrophil elastase concentration were evaluated. RESULTS: CPB time and aortic clamp time did not differ between the groups. Interleukin 8 and neutrophil elastase concentrations before CPB increased significantly immediately after CPB, these concentrations did not differ between the groups. However, neutrophil elastase concentrations of a Ulinastatin group were significantly lower than that of a control group at 30 min after protamine administration (a Ulinastatin group: 1011.3+/-539.1 mg/ml, a control group: 1619.7+/-595.7 mg/ml, p<0.05) and A-aDO2 of a Ulinastatin group was significantly lower than that of a control group at 2 hours after CPB (a Ulinastatin group: 67.1+/-70.6 mmHg, a control group: 169.2+/-116.3 g, p<0.05). CONCLUSIONS: These results suggest that Ulinastatin suppresses the increase in neutrophil elastase and protects the respiratory function in pediatric patients undergoing cardiopulmonary bypass. PMID- 12124549 TI - Clinical evaluation of oxidative stress and myocardial reperfusion injury in pediatric cardiac surgery. AB - BACKGROUND: To evaluate oxidative stress and myocardial damage after aortic crossclamping release (ACCR) during cardio pulmonary bypass (CPB) in children two parameters were investigated: total glutathione (GSH) and its oxidoreductive reactions (GSH/GSSG) as expression of oxidative stress, and plasmatic turnover of myocardial taurine (TAU) as expression of cell damage. METHODS: The study was divided in two periods: 1) first period: analysis of oxidative stress and myocardial damage in 18 children. 2) Second period: evaluation of myocardial cell protection by controlled anterograde low oxygen warm reperfusion (ALOWR) before declamping. Twenty-one children were divided in two groups: not receiving (Group 1, 9 patients) and receiving (Group 2, 12 patients) ALOWR. RESULTS: In the first period GSH values increased significantly after onset of mechanical ventilation (MV) in vein, after CPB start in artery and after ACCR in coronary sinus. Moreover TAU turnover in aortic and coronary sinus blood increased significantly after ACCR. In the second period, Group 2 showed a lower oxidative stress after ACCR, while no differences were observed in TAU turnover. CONCLUSIONS: 1) Assessment of TAU and GSH levels can be considered a good method to clinically evaluate myocardial injury during cardiac surgery. 2) MV and CPB can induce oxidative stress before aortic clamping and can decrease the physiologic scavengers. Therefore, to prevent that depletion, the strategy of these techniques must be adapted to the patient and to his cardiac disease. 3) Intramyocardial TAU turnover is not significantly modified by the reperfusion technique. 4) ALOWR can reduce myocardial oxidative stress and can improve heart recovery after the cardioplegic arrest. PMID- 12124551 TI - Early diagnosis of acute mesenteric ischemia after cardiopulmonary bypass. AB - BACKGROUND: The aim of this study is to identify significant risk factors and eventual clinical markers associated with acute mesenteric ischemia (AMI) after cardiopulmonary bypass. METHODS: The study was retrospectively performed on a group of 19 patients (group A) undergoing cardiac surgery between January 1991 and December 1999, who developed AMI within 30 days of their hospitalization. A control group of 48 patients (group B) was compared in order to define preoperative and operative risk factors for AMI. RESULTS: At the abdominal operation, a non-occlusive mesenteric ischemia was found in every case. In hospital mortality was 84.2% (16/19). Compared to the control, there was a significant difference in aortic cross-clamp time (p<0.001) and use of inotropic drugs (p<0.01). Postoperatively, the studied group (group A) had a significantly higher mean value of the enzymatic serum levels at any time. CONCLUSIONS: A high index of suspicion for mesenteric ischemia after cardiopulmonary bypass should be considered in patients with conditions of hypoperfusion. The early laboratory signs of AMI might be searched during the first postoperative hours. PMID- 12124552 TI - Ruptured intra-aortic balloon pump. A case report. AB - The intra-aortic balloon pump (IABP) is the gold standard for the mechanical assistance of failing heart. Balloon rupture is a potential complication that may result in a retained device. The operative removal by direct exposure of the femoral vessels has previously been described. We report the case of a retained IABP requiring laparotomy and direct aortic exposure for removal of the device. PMID- 12124550 TI - Effects of diclofenac in the prevention of pericardial effusion after coronary artery bypass surgery. A prospective, randomized study. AB - BACKGROUND: It is suggested that pericardial effusions after cardiac surgery can be managed with non-steroid anti-inflammatory drugs, but the efficacy of this therapy is not well established. This study was planned to evaluate the efficacy of the prophylactic use of diclofenac in the prevention of pericardial effusion after coronary artery bypass surgery. METHODS: In a prospective, randomized study, diclofenac sodium 50 mg was administered orally every 8 hours to 22 patients in the postoperative period. The control group consisted of 19 patients who were not given postoperatively either steroids or non-steroid anti inflammatory drugs. RESULTS: Twelve patients of the diclofenac-treated group (54.5%) and 7 of the control group (36.8%) experienced supraventricular arrhythmias postoperatively. There was no statistically significant difference in the size of postoperative pericardial effusion as well as in the occurrence of pleural effusion in both groups. However, there was a higher rate of significant pericardial effusion (grade I-III) in the control group as compared with the diclofenac-treated group (52.6% vs 31.8%, p=ns). Based on chest X-ray findings, patients in the control group had higher incidence of pleural effusion either alone (42.1% vs 22.7%, p=ns) or combined with pericardial effusion (21.0% vs 13.6%, p=ns). Patients who received diclofenac had lower median C-reactive protein concentration (76.0+/-45.2 mg/L) than the patients of the control group (99.6+/-47.8 mg/L), (p=ns). CONCLUSIONS: The results of the present study suggest that diclofenac, even if without a striking effect, may lessen the degree of inflammatory reaction after cardiac surgery and may be useful in the prevention and in the management of early pericardial effusion after cardiac surgery. PMID- 12124553 TI - Repair of a postinfarct ventricular septal defect on the beating heart. Surgical considerations. AB - Postinfarct ventricular septal defect (VSD) still remains associated with a high mortality and morbidity. Despite the development of modern surgical techniques and medical care it continues to be a difficult therapeutic challenge. This report describes a case of a 70-year-old female patient, who presented with a postinfarct VSD after having anterior wall infarction. She presented with left heart failure, pulmonary hypertension and left to right shunt of 78% (Qp/Qs=4.3). The patient was operated on using cardiopulmonary bypass on the beating heart. The closure was performed with a Dacron-patch and a single bypass to the diagonal branch using the left internal thoracic artery. Postoperatively the patient did well and was discharged in good condition on the 13th postoperative day. We conclude that postinfarction VSD can be repaired on cardiopulmonary bypass avoiding cross-clamping. This method is helpful for the outcome as well as for the early postoperative recovery of elderly patients. PMID- 12124554 TI - Beware Swan-Ganz complications. Perioperative management. AB - Since the introduction of the pulmonary artery catheter (PAC) in 1970 by Swan et al., various complications are recognized with the insertion and the use of Swan Ganz catheter. We present two different cases with rare but life threatening complications which had been successfully managed. The first case is a carotid cannulation with an 8.5 Fr introducer sheath, in an attempt to insert a pulmonary catheter via the right internal jugular vein. Two weeks later, the patient was re admitted to the hospital and when an arteriovenous fistulae (carotid artery internal jugular vein) was diagnosed, he was treated surgically. The second case presents the rupture of the right atrium in the conjunction with the superior vena cava. This serious cardiac complication was developed during the floatation of the PAC and the lesion was repaired while the mitral valve replacement was in progress. PMID- 12124556 TI - Cystic adventitial disease of the popliteal artery. Etiologic considerations. PMID- 12124555 TI - Coronary subclavian steal syndrome after left internal mammary bypass in a patient with Takayasu's disease. AB - We report the case of a 56-year-old symptomatic woman who underwent 2 coronary bypasses (left internal mammary artery on the left anterior artery and saphenous venous bypass on the circumflex) for a tight stenosis of the left main coronary. An inflammatory syndrome had been explored for 1 year without specific diagnosis. Eight months later, coronary angiography was performed for recurrence of angina: both bypasses were patent without stenosis and the left main stenosis was unchanged, but significant stenosis of the subclavian artery was found just before the LIMA. The diagnosis of Takayasu's disease was suspected in accordance with the ARC criteria and corticosteroids were started. One year later, because of recurrent angina, the patient was surgically treated with subclavian, vertebral and internal mammary endofibrectomy and an inverted saphenous vein graft from the subclavian to the axillary artery for extensive supra-aortic lesions. The patient remains symptom free at 1 year follow-up. PMID- 12124557 TI - Direct minimally invasive approach for aortic root replacement. AB - The author describes a minimally invasive approach for aortic insufficiency associated with severe calcification of the ascending aorta. The Bentall operation was easily performed through an 8 cm skin incision affording excellent exposure of the operating field, and with the use of CPB with vacuum-assisted venous drainage and clamping the aorta with a Casgrove flex clamp. PMID- 12124558 TI - Antiphospholipid antibodies and intracardiac thrombosis. A case report. AB - The antiphospholipid syndrome (APS) has been associated with multiple cardiac abnormalities. The present report describes a case of right ventricle thrombus in a 51-year-old woman with a history of autoimmune haemolytic anemia and antiphospholipid antibodies. Transthoracic echocardiography demonstrated the presence of a right ventricle mass, mimicking a myxoma. She underwent open heart removal of the mass and was started on indefinitely anticoagulant therapy. At 2 years follow-up she was free of symptoms. PMID- 12124559 TI - Can renal dysfunction after infra-renal aortic aneurysm repair be modified by multi-antioxidant supplementation? AB - BACKGROUND: Renal failure after lower torso ischemia is a serious problem, partly caused by hypotension and indirect reperfusion injury. This injury is partly due to the formation of oxygen free radicals by activated neutrophils. This injury results in albuminuria and renal function impairment. There are indications that free radical damage in indirect reperfusion injury can be diminished by administering extra antioxidants before and during reperfusion. METHODS: In this prospective randomised study we have looked at the influence of a multi antioxidant supplementation on renal function in patients undergoing an elective open infrarenal abdominal aneurysm repair. The patients received either standard treatment (n=22) or standard treatment with additional antioxidants perioperatively (Allopurinol, vitamin E and C, N-acetylcysteine and mannitol). For renal function we have looked at the albumin/creatinine ratio in urine and 24 hr creatinine clearance. RESULTS: Despite significantly increased serum total antioxidant capacity, the group receiving extra antioxidants showed no decrease in the albumin/creatinine ratio in urine. There was however a significantly higher creatinine clearance in this group at day 2. CONCLUSIONS: The results indicate that the diminished renal function after infrarenal aneurysm repair may be influenced by antioxidant therapy. PMID- 12124561 TI - Low molecular weight heparins in the long-term treatment of venous thromboembolism. AB - Low molecular weight heparins (LMWHs) have extensively replaced unfractionated heparin (UFH) in both thromboprophylaxis and initial treatment of venous thromboembolism (VTE) and their use for such indications is now well established. This paper reviews the role of LMWHs in the long-term treatment of VTE. Venous thrombosis, although a very frequent occurrence in everyday practice, still remains controversial in its treatment. Available literature comparing different LMWHs with UFH and oral anticoagulants (OAs) is presented. Comparison and evaluation of the effectiveness, safety and costs of alternative treatments are also made. The differences of various LMWHs are discussed and the need for separate clinical trials for every single LMWH is highlighted. PMID- 12124560 TI - Sialic acid is an indicator of inflammation due to cardiopulmonary bypass but not myocardial damage. AB - BACKGROUND: The aim of the present study was to investigate whether serum sialic acid is associated with the inflammatory response of cardiopulmonary bypass (CPB) or not, since cardiopulmonary bypass is known to produce a systemic inflammatory syndrome. METHODS: In 20 patients in whom elective coronary bypass grafting was done, arterial and coronary sinus blood sampling was done simultaneously. The timing of samplings was 8-10 min after the institution of CPB, just before the application of aortic clamping (T1) and 2 min after cross clamp removal (T2), when there was coronary circulation, but no myocardial activation. RESULTS: Sialic acid, CK-MB, lactate levels were significantly higher at T2 than those at T1 for both arterial and coronary sinus samples. Increase at T2 time point for SA, CK-MB and lactic acid at coronary sinus were closely correlated with the systemic increase of these substances also. Actually, increase of systemic and coronary sinus CK-MB levels was also correlated with the duration of aortic cross clamping and cardiopulmonary bypass. Probably due to consumption, a negative correlation with the decrease of fibrinogen at coronary sinus was found with the duration of aortic cross-clamping. CONCLUSIONS: Our study showed a strong and consistent association between serum SA concentration and the inflammatory process. There are previous reports that show sialic acid levels associated with ischemic insult to myocardium. We are able to show that serum TSA correlates well with some of the acute phase proteins but this was not true with ischemic markers after cardioplegic arrest. PMID- 12124563 TI - Thrombosed aneurysm of the infrarenal vena cava: diagnosis and treatment. AB - Aneurysms of the inferior vena cava are rare. Usually they are diagnosed incidentally or due to the patient having suffered thromboembolic complications. We report one case of a patient admitted due to deep vein thrombosis of his left lower limb in whom a thrombosed mass of the infrarenal vena cava and of both proximal common iliac veins was detected by duplex, CT scan and MRI. The additional information obtained by the phlebography showed abundant collateral circulation via ascending lumbar veins, suggesting gradual occlusion rather than sudden thrombosis. Guided biopsy was not contemplated and the patient underwent surgery with a diagnosis of thrombosis of the infrarenal vena cava due to suspected malignancy versus aneurysm. Intraoperative anatomopathological examination revealed no evidence of malignancy and partial resection with infrarenal vena cava ligation was performed. After six months the patient remains well under oral anticoagulation. On the basis of the literature and taking into account this case, the management of aneurysms of the inferior vena cava when they have already suffered thrombosis should include open surgery which allows us to make the diagnosis with certainty and treatment of the patient. PMID- 12124562 TI - Vascular complications after surgical repair of aneurysms in Behcet's disease. AB - The poor prognosis of vasculo Behcet's disease is often due to postoperative vascular complications (false aneurysm and graft occlusion). We report a case of an abdominal aortic aneurysm associated with an aneurysm of the left common femoral artery in a 23-year-old Portuguese man. The primary treatment was surgical (aneurysmectomy and prosthetic revascularization). The early occlusion of the left femoral revascularized artery was treated surgically with a new bypass. The occlusion of the right limb of the aortoiliac graft was asymptomatic and was not treated. Two months after admission to our hospital, the stenosis of the infrarenal aorta successfully treated by angioplasty via the occluded right limb of the graft. The patient was followed up for 18 months. He could only walk a short distance and had rest pain in the left foot. Magnetic resonance angiography showed a false aneurysm of the infrarenal aorta, and an occlusion of the remaining left limb of the aortoiliac graft. The endovascular treatment performed does not avoid the need for surgical treatment, because occlusion and false aneurysm may occur after dilatation. The endovascular approach can also be used during a sudden inflammatory surge, and makes it possible to wait for a quiescent period when surgery can be performed. PMID- 12124564 TI - Acute spontaneous infrarenal aortic dissection. AB - An acute spontaneous rupture of an aortic dissection originating in and limited to the infrarenal aorta associated with an infrarenal aortic aneurysm is reported. The diagnosis was made by ultrasonography and computed tomography and was confirmed intraoperatively followed by a successful graft insertion. PMID- 12124565 TI - A rare case of atherosclerotic popliteal artery aneurysm in a young adult. AB - Popliteal artery aneurysms are not so clinically frequent but are the most common peripheral aneurysms. They usually affect older men aged over 60 and are caused by atherosclerosis. Whenever they occur in younger men, other more unusual etiologies such as trauma, infection, inflammatory arteritis or popliteal entrapment are responsible. In this paper we present the case of a young male with an atherosclerotic popliteal artery aneurysm without a history of penetrating trauma or other possible etiologies. PMID- 12124566 TI - False aneurysm of perforating branch of the profunda femoris artery after external fixation for a complicated femur fracture. AB - False aneurysm of the profunda femoris artery rarely occurs and is a serious complication following femur fracture. A 39-year-old man who developed a false aneurysm arising from the perforating branch of the profunda femoris artery following an external fixation for a complicated femur fracture was presented. Clinical diagnosis was confirmed by selective arterial angiography after occurrence of significant hemorrhage and swelling of the injured thigh. The false aneurysm was treated by ligation of the perforating branch of the profunda femoris artery and excision of the aneurysmal sac via the medial approach. Clinical status of the patient was uneventful postoperatively. The right thigh swelling decreased rapidly following the operation in 1 week. The patient was discharged on the 10th postoperative day with external fixation. False aneurysm in a branch of the profunda femoris artery is a very rare status following application of the external fixator due to complicated femur fracture. Related literatures and interventions were reviewed on the basis of this case. PMID- 12124567 TI - Endovascular treatment of a postlaminectomy arteriovenous fistula. A case report. AB - We report a case of iliac arteriovenous fistula (AVF) following disk surgery. A 51-year-old woman underwent hemilaminectomy for a slipped disk. Two weeks after surgery the patient experienced dyspnea and oedema of the lower limbs. Presence of a systolic murmur on the cardiac floor and on the abdomen was detected and abdomen CT scan which evidenced a AVF between the right common iliac artery and vein. The lesion, confirmed by angiography, was successfully treated with the endovascular technique. The endovascular technique appears to be a valid alternative to the traditional surgical treatment of postlaminectomy AVF. PMID- 12124568 TI - Thoracic venous aneurysms. Clinical observation. AB - Venous aneurysms are infrequent vascular alterations. Their origin is not well known, but pathologic study reveals probably a similar natural history than the arterial aneurysms. Their clinical significance arises from the complications they can originate, specially in certain locations, which include pulmonary embolism, and, on the other hand, differential diagnosis with certain malignant pathologies may be necessary, depending upon the clinical context. A patient is presented whose radiographic findings, in association with clinical factors, led to surgical exploration to rule out malignant lung tumor. A left subclavian vein aneurysm was finally diagnosed using magnetic resonance imaging. PMID- 12124569 TI - Trimodality treatment versus surgery alone for esophageal cancer. A stratified analysis with minimally invasive pretreatment staging. AB - BACKGROUND: Accurate pretreatment staging of esophageal cancer (EC) is important in the evaluation and comparison of results of different treatment modalities. Few studies using minimally invasive staging techniques for this purpose have been reported. We previously demonstrated the usefulness of the thoracoscopic/laparoscopic (Ts/Ls) technique in pretreatment staging of EC. This study was conducted to evaluate the impact of trimodality based on pretreatment Ts/Ls staging diagnosis on EC. METHODS: A retrospective study was performed on 2 groups of EC patients. Group A (44 patients) underwent pretreatment Ts/Ls staging and had trimodality treatment. Preoperative therapy consisted of concurrent chemotherapy (5-FU + cisplatinum) and radiotherapy. Group B (33 patients) underwent surgery alone. The study focused on stratified comparison of patterns of recurrence and survival in different pretreatment surgical T, N, and TNM stage categories. RESULTS: The 3-year disease free survival of Group A was 40.8% with a median survival of 32.0 months, it was 43.6% with a median survival of 23.6 months in Group B. The difference was not significant (p=0.87). There was no difference in recurrence pattern between the 2 groups. Patients with squamous cell carcinoma in Group A had no local recurrence during the follow-up period while those in Group B had a high local recurrence rate of 40% (p<0.005). When stratified by T factor, patients with locally advanced T stage (T3-4) in Group A had a lower distant recurrence rate than their counterpart patients in Group B (9.1 vs 38.5%, p=0.03), they had a better survival but the difference was not significant (3-year disease free survival: 41.7 vs 17.9%, p=0.14). There were no significant differences in recurrence pattern and survival in different N categories and TNM stages between 2 groups. Multivariate analysis showed that only pretreatment surgical N status was an independent prognostic factor for the whole group (p=0.02). CONCLUSIONS: Pretreatment Ts/Ls staging can provide accurate staging information for EC patients. Trimodality treatment was successful in local control for patients with squamous cell carcinoma. It was effective in reducing distant recurrence and might prolong survival in patients with advanced T stages. Pretreatment lymph node status was the most important prognosticator regardless of treatment modality. Pretreatment pathological staging should be included in the future clinical trials on multimodality treatments in EC patients. PMID- 12124570 TI - Postoperative complications of lung resection after induction chemotherapy using Paclitaxel (and radiotherapy) for advanced non-small lung cancer. AB - BACKGROUND: Locally advanced non-small-cell lung carcinoma is currently treated by multidisciplinary protocols using a combination of chemotherapy, radiotherapy and surgery. However the best strategy for applying these therapeutic measures has not yet been established. One of the difficulties of using these forms of treatment is their toxicity. Our aim was to determine whether the postoperative course of the disease can be influenced by preoperative chemotherapy in any way. METHODS: Nineteen patients were surgically treated after receiving induction treatment between October 1996 and October 1998. The indications for giving induction treatment were: stage III disease in 12 patients (1 Pancoast tumor), lung cancer and solitary brain metastasis in 4 patients, double primary lung cancer in 3 patients (1 synchronous and 2 metachronous). Variables were the chemotherapy treatment time interval from the beginning to surgery, the type of surgery, postoperative mortality and morbidity. Mean age was 55.9 years old (range between 25 and 70 years). Predominant gender was male (18 men and 1 woman). Neoadjuvant treatment consisted of chemotherapy in all patients (Paclitaxel, Cysplatin and Vinorelbine in cycles for a mean period of 3 months), and radiotherapy (14 patients). Pulmonary resections were: pneumonectomy (2 patients), lobectomy (16 patients) and wedge resection (1 patient). There were no exploratory thoracotomies. Bronchoplasty procedures were necessary in 5 cases and angioplasty in 5. Cardiopulmonary bypass was necessary in 1 case in order to resect an infiltrated pulmonary vein. Intraoperative radiotherapy (IORT) was used in 9 cases. RESULTS: Complications occurred in the immediate postoperative period in 9 patients: 1 postpneumonectomy respiratory distress syndrome, 2 bronchopleural fistulae, 4 prolonged air leaks, 1 complete dehiscence of the thoracotomy scar and 1 colitis caused by anaerobes. The postoperative mortality (within 30 days) was 2 patients (10.5%): 1 died from bronchopleural fistula and the other from postpneumonectomy respiratory distress syndrome. However, if we take into account the fact that the case of anaerobic colitis also ended with the patient's death on the 48th postoperative day, and we include it in the mortality rate, the final mortality is higher (15.8%). CONCLUSIONS: Surgery for non-small cell lung carcinoma has to be considered a high-risk procedure. But, if patients are selected appropriately and the perioperative management is satisfactory, reasonable rates of morbidity and mortality can be achieved. More studies are needed in order to define the exact role of these therapeutic measures. PMID- 12124571 TI - Biological markers in non-small cell lung cancer. Retrospective study of 10 year follow-up after surgery. AB - BACKGROUND: The biological markers in non-small cell lung cancer (NSCLC) have been widely studied and encouraging results have shown that products of some oncogenes and other molecular markers can predict the aggressiveness of the disease and the outcome of the patients. METHODS: To verify the reliability of these prognostic markers we have studied retrospectively the expression of c-erbB 2 and 67Ki (growth regulation), p53 (cell cycle regulation and apoptosis), bcl-2 (apoptosis) and CD31 and CD34 (angiogenesis) in 78 patients operated on for NSCLC with curative intent between January 1987 and December 1988 and followed up for 10 years. For the determination of the biological markers we have used the ABC (Avidin-Biotin-Peroxidase complex) immunohistochemical method. The Cox regression model was used for the univariate and multivariate analysis. RESULTS: Nineteen patients (24%) were alive after 10 years and 59 (76%) died. The univariate analysis of the relationship between the 10-year survival and the expression of the markers was significant only for p53 (p=0.0097). Stratifying the patients according to the 3 histological subtypes (squamous cell carcinoma, adenocarcinoma and large cell undifferentiated carcinoma) the correlation between markers and survival pointed out that the only significant one was p53 (p=0.0459) in adenocarcinoma. In the same way considering the stages p53 was significant in stage IIIa (p=0.0357). The multivariate analysis emphasized that p53 was the only significant marker with respect to the 10-year survival (p=0.0091). Examining the histological groups significant was only p53 in adenocarcinoma (p=0.0192) and in large cell undifferentiated carcinomas (p=0.0290). This marker is also significant in pathological stage II (p=0.0271) and IIIa (p=0.0402). Apart from histology and staging the 10-year survival was 33% for p53 negative versus 10% for p53 positive. In patients with adenocarcinoma the 10-year survival was 40% for p53 negative and 6% for p53 positive. CONCLUSIONS: In conclusion our results emphasize the importance of p53 as a prognostic factor in 10-year survival in patients with adenocarcinoma and in stage II and IIIa. PMID- 12124572 TI - Necrotizing soft tissue infection of the chest wall. AB - Necrotizing soft tissue infection (NSTI) of the chest wall is a relatively rare but highly lethal surgical infection which has received little attention in the medical and surgical literature. The data reported are based on a literature review, including a Medline database search, and search of existing bibliographies. Twenty well-documented cases of primary chest wall involvement by NSTI were found. Thirteen patients were male. Patients' ages ranged from 10 weeks to 78 years. Thirteen patients were postoperative (65%). The diagnosis was initially considered in only 3 of the postoperative cases, a cause of significant delays in surgical treatment. Among those who lived long enough for their wounds to close, 2 had secondary healing, 5 experienced delayed skin grafting over the granulating wound, and 1 had skin grafting combined with muscle transfer. Mortality was 60%. Chest wall NSTI is a rapidly spreading, highly lethal infection. A high index of suspicion, early diagnosis, and aggressive approach are essential to its successful treatment. PMID- 12124573 TI - The role of gastroesophageal reflux disease in chest pain. AB - Gastroesophageal reflux disease (GERD) is thought to play an important role in the pathogenesis of non-cardiac, unexplained, chest pain. The physiopathological mechanism of this extraesophageal manifestation, remains unclear but it is supposed that the ache could be triggered by the stimulation of acid-sensitive nociceptors of the mucosa. The esophageal origin of the symptom may be identified by an empiric trial of high-dose antisecretory drugs or an abnormal prolonged ambulatory pH monitoring study. Less important is the role of endoscopy especially in subjects without typical symptoms of GERD. The use of manometry or provocative tests can consent to evaluate for esophageal motility abnormalities. It is generally accepted that in clinical practice, in patients with non-cardiac chest pain, the administration of proton pump inhibitors, could serve as a first approach. It is cost-effective in the short-term period, when compared with investigation for gastrointestinal causes, with cost savings persisting beyond a year. In conclusion, patients with non-cardiac chest pain of unknown origin should be carefully screened for the occurrence of GERD, but further research is needed to clarify the role of the latter on the pathogenesis of this symptom. PMID- 12124574 TI - Severe hypoglycaemia associated with a giant solitary fibrous tumor of the pleura. AB - Solitary fibrous tumors (SFT) of the pleura are a rare neoplasm, with benign biological behaviour. Recurrences are rare, and no distant metastases are described in the literature. SFT can secrete hormone-like substances, responsible for paraneoplastic syndromes. The authors describe a case of severe hypoglycaemia due to insulin-like growth factor II (IGF-2)'s secretion by a giant SFT of the pleura. Hypoglycaemia was controlled by the resection of the tumor. Diagnosis and surgical management of these neoplasms are also discussed. PMID- 12124575 TI - Traumatic subarachnoid-pleural fistula in children. Report of 2 cases and review of the literature. AB - Traumatic subarachnoid-pleural fistula (TSAPF) is a rare complication of thoracic trauma. Late diagnosis is a problematic dilemma in these cases. Patients with thoracic injury have persistent pleural leakage, thoracic vertebral injury, pneumocephalus, urinary retention and paraplegia should alert the surgeon for TSAPF. Two cases of TSAPF due to gunshot injury are reported. PMID- 12124576 TI - 4-Dimensional color Doppler echocardiography during mitral valve surgery. PMID- 12124577 TI - Atypically located pericardial cysts. PMID- 12124578 TI - Primary repair and sequential aortocoronary bypass for aortic dissection complicating coronary angioplasty in multivessel disease. PMID- 12124580 TI - New research council needed. PMID- 12124581 TI - California lab fires physicist over retracted finding. PMID- 12124582 TI - Government spending promise offers British research a boost. PMID- 12124583 TI - Senate nod prompts fresh analysis of nuclear waste dump. PMID- 12124584 TI - German council confounds plans for neutron project. PMID- 12124586 TI - Cutbacks 'will cripple space station science'. PMID- 12124587 TI - Publishers soldier on despite electronic bugs. PMID- 12124588 TI - US panel split on research cloning. PMID- 12124589 TI - Poliovirus advance sparks fears of data curbs. PMID- 12124590 TI - Monkey smallpox trial suspended over painkiller use. PMID- 12124591 TI - Medical journal retracts paper over falsified data. PMID- 12124594 TI - Can you keep a secret? PMID- 12124593 TI - They said it couldn't be done... PMID- 12124595 TI - Only vital need justifies primate experiments. PMID- 12124596 TI - Little funding to develop non-animal testing. PMID- 12124597 TI - Are results of primate research worth the suffering it causes? PMID- 12124598 TI - Names: a historical or political perspective? PMID- 12124603 TI - Weaving life's pattern. PMID- 12124604 TI - Survival of the entangled. PMID- 12124605 TI - Developmental biology: decisions, decisions! PMID- 12124606 TI - Population genetics: malaria variorum. PMID- 12124608 TI - Plastic parameter. PMID- 12124609 TI - The extraordinary phagosome. PMID- 12124610 TI - Longevity: don't hold your breath. PMID- 12124611 TI - Cacao usage by the earliest Maya civilization. AB - The Maya archaeological site at Colha in northern Belize, Central America, has yielded several spouted ceramic vessels that contain residues from the preparation of food and beverages. Here we analyse dry residue samples by using high-performance liquid chromatography coupled to atmospheric-pressure chemical ionization mass spectrometry, and show that chocolate (Theobroma cacao) was consumed by the Preclassic Maya as early as 600 bc, pushing back the earliest chemical evidence of cacao use by some 1,000 years. Our application of this new and highly sensitive analytical technique could be extended to the identification of other ancient foods and beverages. PMID- 12124612 TI - Targeted transfection by femtosecond laser. AB - The challenge for successful delivery of foreign DNA into cells in vitro, a key technique in cell and molecular biology with important biomedical implications, is to improve transfection efficiency while leaving the cell's architecture intact. Here we show that a variety of mammalian cells can be directly transfected with DNA without perturbing their structure by first creating a tiny, localized perforation in the membrane using ultrashort (femtosecond), high intensity, near-infrared laser pulses. Not only does this superior optical technique give high transfection efficiency and cell survival, but it also allows simultaneous evaluation of the integration and expression of the introduced gene. PMID- 12124613 TI - Neurodegenerative disease: amyloid pores from pathogenic mutations. AB - Alzheimer's and Parkinson's diseases are associated with the formation in the brain of amyloid fibrils from beta-amyloid and alpha-synuclein proteins, respectively. It is likely that oligomeric fibrillization intermediates (protofibrils), rather than the fibrils themselves, are pathogenic, but the mechanism by which they cause neuronal death remains a mystery. We show here that mutant amyloid proteins associated with familial Alzheimer's and Parkinson's diseases form morphologically indistinguishable annular protofibrils that resemble a class of pore-forming bacterial toxins, suggesting that inappropriate membrane permeabilization might be the cause of cell dysfunction and even cell death in amyloid diseases. PMID- 12124614 TI - Recent temperature trends in the Antarctic. AB - It is important to understand how temperatures across the Antarctic have changed in recent decades because of the huge amount of fresh water locked into the ice sheet and the impact that temperature changes may have on the ice volume. Doran et al. claim that there has been a net cooling of the entire continent between 1966 and 2000, particularly during summer and autumn. We argue that this result has arisen because of an inappropriate extrapolation of station data across large, data-sparse areas of the Antarctic. PMID- 12124617 TI - Hot bubbles from active galactic nuclei as a heat source in cooling-flow clusters. AB - Hot, X-ray-emitting plasma permeates clusters of galaxies. The X-ray surface brightness often shows a peak near the centre of the cluster that is coincident with a drop in the entropy of the gas. This has been taken as evidence for a 'cooling flow', where the gas cools by radiating away its energy, and then falls to the centre. Searches for this cool gas have revealed significantly less than predicted, indicating that the mass deposition rate is much lower than expected. Most clusters with cooling flows, however, also host an active galactic nucleus at their centres. These active galactic nuclei can inflate large bubbles of hot plasma that subsequently rise through the cluster 'atmosphere', thus stirring the cooling gas and adding energy. Here we report highly resolved hydrodynamic simulations which show that buoyant bubbles increase the cooling time in the inner regions of clusters and significantly reduce the deposition of cold gas. PMID- 12124616 TI - A molecular programme for the specification of germ cell fate in mice. AB - Germ cell fate in mice is induced in proximal epiblast cells by the extra embryonic ectoderm, and is not acquired through the inheritance of any preformed germ plasm. To determine precisely how germ cells are specified, we performed a genetic screen between single nascent germ cells and their somatic neighbours that share common ancestry. Here we show that fragilis, an interferon-inducible transmembrane protein, marks the onset of germ cell competence, and we propose that through homotypic association, it demarcates germ cells from somatic neighbours. Using single-cell gene expression profiles, we also show that only those cells with the highest expression of fragilis subsequently express stella, a gene that we detected exclusively in lineage-restricted germ cells. The stella positive nascent germ cells exhibit repression of homeobox genes, which may explain their escape from a somatic cell fate and the retention of pluripotency. PMID- 12124618 TI - Plasmon-assisted transmission of entangled photons. AB - The state of a two-particle system is said to be entangled when its quantum mechanical wavefunction cannot be factorized into two single-particle wavefunctions. This leads to one of the strongest counter-intuitive features of quantum mechanics, namely non-locality. Experimental realization of quantum entanglement is relatively easy for photons; a starting photon can spontaneously split into a pair of entangled photons inside a nonlinear crystal. Here we investigate the effects of nanostructured metal optical elements on the properties of entangled photons. To this end, we place optically thick metal films perforated with a periodic array of subwavelength holes in the paths of the two entangled photons. Such arrays convert photons into surface-plasmon waves- optically excited compressive charge density waves--which tunnel through the holes before reradiating as photons at the far side. We address the question of whether the entanglement survives such a conversion process. Our coincidence counting measurements show that it does, so demonstrating that the surface plasmons have a true quantum nature. Focusing one of the photon beams on its array reduces the quality of the entanglement. The propagation of the surface plasmons makes the array effectively act as a 'which way' detector. PMID- 12124619 TI - Atomistic mechanisms governing elastic limit and incipient plasticity in crystals. AB - Nanometre-scale contact experiments and simulations demonstrate the potential to probe incipient plasticity--the onset of permanent deformation--in crystals. Such studies also point to the need for an understanding of the mechanisms governing defect nucleation in a broad range of fields and applications. Here we present a fundamental framework for describing incipient plasticity that combines results of atomistic and finite-element modelling, theoretical concepts of structural stability at finite strain, and experimental analysis. We quantify two key features of the nucleation and subsequent evolution of defects. A position sensitive criterion based on elastic stability determines the location and character of homogeneously nucleated defects. We validate this stability criterion at both the atomistic and the continuum levels. We then propose a detailed interpretation of the experimentally observed sequence of displacement bursts to elucidate the role of secondary defect sources operating locally at stress levels considerably smaller than the ideal strength required for homogeneous nucleation. These findings provide a self-consistent explanation of the discontinuous elastic plastic response in nanoindentation measurements, and a guide to fundamental studies across many disciplines that seek to quantify and predict the initiation and early stages of plasticity. PMID- 12124620 TI - Dynamic fracture by large extraterrestrial impacts as the origin of shatter cones. AB - A large impact by a comet or meteorite releases an enormous amount of energy, which evaporates, melts and fractures the surrounding rocks. Distinctive features of such impacts are 'shatter cones', deformed rocks characterized by hierarchical striated features. Although such features have been used for decades as unequivocal fingerprints of large-body impacts, the process by which shatter cones form has remained enigmatic. Here we show that the distinctive shatter-cone striations naturally result from nonlinear waves (front waves) that propagate along a fracture front. This explains the observed systematic increase of striation angles with the distance from the impact. Shatter-cone networks, typically spanning many scales, can be understood as hierarchical bifurcations of the fracture front, which is generated by the immense energy flux carried by the initial, impact-generated, shock waves. Our quantitative predictions based on this theory are supported by field measurements at the Kentland and Vredefort impact sites. These measurements indicate that shatter cones near to the impact site were formed by fractures propagating at nearly the Rayleigh wave speed of the host rocks, whereas the furthest shatter cones observed (about 40 km from the impact site) were formed by fronts moving more slowly. These results provide insight into impact dynamics as well as dissipative mechanisms in solids subjected to sudden, extremely intense fluxes of energy. PMID- 12124621 TI - Climate-mediated energetic constraints on the distribution of hibernating mammals. AB - To predict the consequences of human-induced global climate change, we need to understand how climate is linked to biogeography. Energetic constraints are commonly invoked to explain animal distributions, and physiological parameters are known to vary along distributional gradients. But the causal nature of the links between climate and animal biogeography remain largely obscure. Here we develop a bioenergetic model that predicts the feasibility of mammalian hibernation under different climatic conditions. As an example, we use the well quantified hibernation energetics of the little brown bat (Myotis lucifugus) to parameterize the model. Our model predicts pronounced effects of ambient temperature on total winter energy requirements, and a relatively narrow combination of hibernaculum temperatures and winter lengths permitting successful hibernation. Microhabitat and northern distribution limits of M. lucifugus are consistent with model predictions, suggesting that the thermal dependence of hibernation energetics constrains the biogeography of this species. Integrating projections of climate change into our model predicts a pronounced northward range expansion of hibernating bats within the next 80 years. Bioenergetics can provide the simple link between climate and biogeography needed to predict the consequences of climate change. PMID- 12124622 TI - An antioxidant function for DMSP and DMS in marine algae. AB - The algal osmolyte dimethylsulphoniopropionate (DMSP) and its enzymatic cleavage product dimethylsulphide (DMS) contribute significantly to the global sulphur cycle, yet their physiological functions are uncertain. Here we report results that, together with those in the literature, show that DMSP and its breakdown products (DMS, acrylate, dimethylsulphoxide, and methane sulphinic acid) readily scavenge hydroxyl radicals and other reactive oxygen species, and thus may serve as an antioxidant system, regulated in part by enzymatic cleavage of DMSP. In support of this hypothesis, we found that oxidative stressors, solar ultraviolet radiation, CO(2) limitation, Fe limitation, high Cu(2+) (ref. 9) and H(2)O(2) substantially increased cellular DMSP and/or its lysis to DMS in marine algal cultures. Our results indicate direct links between such stressors and the dynamics of DMSP and DMS in marine phytoplankton, which probably influence the production of DMS and its release to the atmosphere. As oxidation of DMS to sulphuric acid in the atmosphere provides a major source of sulphate aerosols and cloud condensation nuclei, oxidative stressors--including solar radiation and Fe limitation--may be involved in complex ocean atmosphere feedback loops that influence global climate and hydrological cycles. PMID- 12124623 TI - Genetic diversity and chloroquine selective sweeps in Plasmodium falciparum. AB - Widespread use of antimalarial agents can profoundly influence the evolution of the human malaria parasite Plasmodium falciparum. Recent selective sweeps for drug-resistant genotypes may have restricted the genetic diversity of this parasite, resembling effects attributed in current debates to a historic population bottleneck. Chloroquine-resistant (CQR) parasites were initially reported about 45 years ago from two foci in southeast Asia and South America, but the number of CQR founder mutations and the impact of chlorquine on parasite genomes worldwide have been difficult to evaluate. Using 342 highly polymorphic microsatellite markers from a genetic map, here we show that the level of genetic diversity varies substantially among different regions of the parasite genome, revealing extensive linkage disequilibrium surrounding the key CQR gene pfcrt and at least four CQR founder events. This disequilibrium and its decay rate in the pfcrt-flanking region are consistent with strong directional selective sweeps occurring over only approximately 20-80 sexual generations, especially a single resistant pfcrt haplotype spreading to very high frequencies throughout most of Asia and Africa. The presence of linkage disequilibrium provides a basis for mapping genes under drug selection in P. falciparum. PMID- 12124624 TI - Chromosome-wide SNPs reveal an ancient origin for Plasmodium falciparum. AB - The Malaria's Eve hypothesis, proposing a severe recent population bottleneck (about 3,000-5,000 years ago) of the human malaria parasite Plasmodium falciparum, has prompted a debate about the origin and evolution of the parasite. The hypothesis implies that the parasite population is relatively homogeneous, favouring malaria control measures. Other studies, however, suggested an ancient origin and large effective population size. To test the hypothesis, we analysed single nucleotide polymorphisms (SNPs) from 204 genes on chromosome 3 of P. falciparum. We have identified 403 polymorphic sites, including 238 SNPs and 165 microsatellites, from five parasite clones, establishing chromosome-wide haplotypes and a dense map with one polymorphic marker per approximately 2.3 kilobases. On the basis of synonymous SNPs and non-coding SNPs, we estimate the time to the most recent common ancestor to be approximately 100,000-180,000 years, significantly older than the proposed bottleneck. Our estimated divergence time coincides approximately with the start of human population expansion, and is consistent with a genetically complex organism able to evade host immunity and other antimalarial efforts. PMID- 12124625 TI - Dendritic spikes as a mechanism for cooperative long-term potentiation. AB - Strengthening of synaptic connections following coincident pre- and postsynaptic activity was proposed by Hebb as a cellular mechanism for learning. Contemporary models assume that multiple synapses must act cooperatively to induce the postsynaptic activity required for hebbian synaptic plasticity. One mechanism for the implementation of this cooperation is action potential firing, which begins in the axon, but which can influence synaptic potentiation following active backpropagation into dendrites. Backpropagation is limited, however, and action potentials often fail to invade the most distal dendrites. Here we show that long term potentiation of synapses on the distal dendrites of hippocampal CA1 pyramidal neurons does require cooperative synaptic inputs, but does not require axonal action potential firing and backpropagation. Rather, locally generated and spatially restricted regenerative potentials (dendritic spikes) contribute to the postsynaptic depolarization and calcium entry necessary to trigger potentiation of distal synapses. We find that this mechanism can also function at proximal synapses, suggesting that dendritic spikes participate generally in a form of synaptic potentiation that does not require postsynaptic action potential firing in the axon. PMID- 12124626 TI - Identification of genes expressed in C. elegans touch receptor neurons. AB - The extent of gene regulation in cell differentiation is poorly understood. We previously used saturation mutagenesis to identify 18 genes that are needed for the development and function of a single type of sensory neuron--the touch receptor neuron for gentle touch in Caenorhabditis elegans. One of these genes, mec-3, encodes a transcription factor that controls touch receptor differentiation. By culturing and isolating wild-type and mec-3 mutant cells from embryos and applying their amplified RNA to DNA microarrays, here we have identified genes that are known to be expressed in touch receptors, a previously uncloned gene (mec-17) that is needed for maintaining touch receptor differentiation, and more than 50 previously unknown mec-3-dependent genes. These genes are randomly distributed in the genome and under-represented both for genes that are co-expressed in operons and for multiple members of gene families. Using regions 5' of the start codon of the first 20 genes, we have also identified an over-represented heptanucleotide, AATGCAT, that is needed for the expression of touch receptor genes. PMID- 12124627 TI - Calorie restriction extends Saccharomyces cerevisiae lifespan by increasing respiration. AB - Calorie restriction (CR) extends lifespan in a wide spectrum of organisms and is the only regimen known to lengthen the lifespan of mammals. We established a model of CR in budding yeast Saccharomyces cerevisiae. In this system, lifespan can be extended by limiting glucose or by reducing the activity of the glucose sensing cyclic-AMP-dependent kinase (PKA). Lifespan extension in a mutant with reduced PKA activity requires Sir2 and NAD (nicotinamide adenine dinucleotide). In this study we explore how CR activates Sir2 to extend lifespan. Here we show that the shunting of carbon metabolism toward the mitochondrial tricarboxylic acid cycle and the concomitant increase in respiration play a central part in this process. We discuss how this metabolic strategy may apply to CR in animals. PMID- 12124628 TI - Adenovirus oncoproteins inactivate the Mre11-Rad50-NBS1 DNA repair complex. AB - In mammalian cells, a conserved multiprotein complex of Mre11, Rad50 and NBS1 (also known as nibrin and p95) is important for double-strand break repair, meiotic recombination and telomere maintenance. This complex forms nuclear foci and may be a sensor of double-strand breaks. In the absence of the early region E4, the double-stranded DNA genome of adenovirus is joined into concatemers too large to be packaged. We have investigated the cellular proteins involved in this concatemer formation and how they are inactivated by E4 products during a wild type infection. Here we show that concatemerization requires functional Mre11 and NBS1, and that these proteins are found at foci adjacent to viral replication centres. Infection with wild-type virus results in both reorganization and degradation of members of the Mre11-Rad50-NBS1 complex. These activities are mediated by three viral oncoproteins that prevent concatemerization. This targeting of cellular proteins involved in genomic stability suggests a mechanism for 'hit-and-run' transformation observed for these viral oncoproteins. PMID- 12124630 TI - Effect of tissue phospholipid composition on synergistic interactions between sympathetic and parasympathetic systems. AB - The content of phospholipids in various tissues was studied during synergistic interaction between two branches of the autonomic nervous system. We found higher myocardial content of choline-containing phospholipids and lower rigidity of erythrocyte membranes in rabbits with pronounced sympathetic potentiation of the cardioinhibitory effect of the vagus nerve compared to rabbits without such potentiation. PMID- 12124631 TI - Respiratory effects of sensorimotor cortex and their mechanisms in rats. AB - Acute experiments on anesthetized rats showed differential effect of various areas of the sensorimotor cortex on activity of the respiratory center. It is hypothesized that GABAergic structures of the solitary tract nucleus play an important role in the mechanisms of respiratory effects of the sensorimotor cortex. PMID- 12124632 TI - Amygdalofugal modulation of Hering-Breuer inspiration-inhibiting reflex. AB - Acute experiments on rats showed that the central nucleus of the amygdaloid complex modulates realization of the Hering-Breuer inspiration-inhibiting reflex. It is hypothesized that amygdalobulbar interrelations are realized via modulation of respiratory reflexes by the amygdala. This interaction is mediated by the GABAergic system of the amygdala. PMID- 12124633 TI - Delayed consequences of protein deficiency during lactation for the development of enzyme systems of digestive and nondigestive organs in offspring. AB - Suboptimal protein nutrition during lactation has a negative impact on the digestive function of the small intestine and trophic barrier functions of the large intestine, liver, and kidneys due to significant enzyme deficiency (disaccharidase, peptidase, alkaline phosphatase) in 6-month-old offspring. Changes in enzyme activity in digestive and nondigestive organs play an important role in metabolic disorders promoting the development of "risk diseases" and reducing lifespan. PMID- 12124634 TI - Effect of electrical stimulation of right stellate ganglion on cardiac function in rats during postnatal ontogeny. AB - In all examined age groups of rats, the threshold amplitude of stellate ganglion stimulation is higher for the positive chronotropic effect than for the inotropic effect. The stimulation produced a more pronounced effect on stroke volume than on heart rate. PMID- 12124635 TI - Effect of low-frequency low-intensity ultrasound on contractile function of isolated heart. AB - Stimulation of the isolated heart with low-intensity low-frequency ultrasound produced a positive inotropic effect (at certain ultrasound intensity). This effect manifested in increased systolic and pulse intraventricular pressures and persisted for tens of minutes after the end of sonication. PMID- 12124636 TI - Immune response and corticosteroid content in different regimens of cooling. AB - The effects of different cooling regimens modulating thermal afferent signal on the immune response were experimentally studied on rats. Immunization at low body temperature changed the immune response to the antigen. These changes depended on the depth and rate of previous cooling. The presence of dynamic activity of peripheral thermosensitive afferents potentiated the immune response after surface cooling and reduced the degree of suppression of the immune response after deep cooling. No clear-cut relationship between changes in the immune response and blood concentration of corticosterone during cooling was found. PMID- 12124637 TI - Protective effect of copper-rutin complex in animals with experimental epilepsy. AB - Copper-rutin complex (2 mg/kg) completely eliminated epileptiform potentials induced by a combination of chlorpromazine and microwave radiation 1-2 min postinjection and suppressed convulsive activity provoked by application of penicillin to the sensorimotor cortex. PMID- 12124638 TI - Receptor specificity of the antiarrhythmic effect produced by opioid peptides Dalargin and DADLE during myocardial reperfusion. AB - Nonselective agonists of mu- and delta-opioid receptors dalargin (D Ala2,Leu5,Arg6-enkephalin) and DADLE (D-Ala2,D-Leu5-enkephalin) administered immediately before coronary reperfusion in a dose of 0.1 mg/kg prevented the development of ventricular arrhythmias. Blockade of mu-opioid receptors abolished the antiarrhythmic effect of these peptides. Hence, antiarrhythmic activity of dalargin and DADLE is primarily associated with activation of mu-opioid receptors. PMID- 12124639 TI - Development of insulin resistance in experimental animals during long-term glucocorticoid treatment. AB - Long-term treatment with glucocorticoids led to the development of insulin resistance in experimental animals, which was confirmed by a progressive increase in blood insulin level and decrease in the glucose/insulin index. The comparative study showed that hydrocortisone produced more pronounced and rapid changes than dexamethasone. However, we found no differences in the glucose/insulin index by the 23rd day of treatment with these hormones. PMID- 12124640 TI - Interaction of human apolipoprotein A-I with rsCD4 receptor. AB - The interaction between recombinant soluble CD4 receptor (rsCD4) with human apolipoprotein A-I was studied by enzyme-linked immunosorbent assay. Our findings suggest that HIV-1 and human apolipoprotein A-I compete for the CD4 receptor and receptor for apoA-I. PMID- 12124641 TI - Possibility of linear dependence between vascular wall tension and blood flow in precortical arterioles. AB - Intravital microscopy was used to determine the diameter and length of pial cortical arterioles in rat brain. Mathematical model yielded a formula showing that regulation of arteriolar wall tension results in autoregulation of the blood flow. PMID- 12124642 TI - Mechanisms of extraribosomal protein biosynthesis in eukaryotes. AB - The appearance of newly synthesized proteins in the total protein fraction of a cell-free protein-synthesizing system not containing ribosomes and mRNA and enriched with polyenzyme complexes of aminoacyl-tRNA synthetases and corresponding proteins is proved experimentally. For modeling of extraribosomal protein biosynthesis we proposed a molecular polyenzyme mechanism of protein biosynthesis on a protein matrix. PMID- 12124643 TI - Changes in the ratio between apolipoprotein (a) and lipids in human plasma during interaction with polyclonal sheep antibodies against lipoprotein (a). AB - Plasma contents of apolipoprotein (a), apolipoprotein B 100, cholesterol, triglycerides, and vitamin E were measured in 2 patients with lipoprotein (a) concentration >100 mg/dl during the interaction with the anti-lipoprotein (a) immunosorbent. Intraindividual heterogeneity of apolipoprotein (a)-containing particles in the plasma was demonstrated. Polyclonal antibodies against lipoprotein (a) immobilized on Sepharose CL-4B more effectively removed free apolipoprotein (a) than complexes containing apolipoproteins B 100, apolipoprotein (a), lipids, and vitamin E. PMID- 12124644 TI - Role of pronase E in ligand binding to Fc receptors in the reaction of cytotoxicity mediated by bovine blood neutrophils. AB - We studied the effects of pronase E on IgG binding to Fcg receptors on bovine blood neutrophils and kinetic parameters of this reaction (association constant and number of binding sites on neutrophils). During in vitro modeling of the cell immune response, we evaluated cytotoxic activity of effector neutrophils against foreign cells. PMID- 12124645 TI - Endogenous dipeptide cycloprolylglycine shows selective anxiolytic activity in animals with manifest fear reaction. AB - Experiments on two mouse strains with opposite reactions to emotional stress showed selectivity of the anxiolytic effect of endogenous dipeptide cycloprolylglycine. In the open field test cycloprolylglycine (0.01-0.10 mg/kg intraperitoneally) dose-dependently (1.8-2.1-fold) increased motor activity of BALB/c mice with manifest fear reaction and had no effect on C57Bl/6 mice with active behavior. The content of endogenous cycloprolylglycine in mouse brain correlated with the type of emotional stress reaction: its content in the brain of C57Bl/6 mice 1.5 times surpassed that in BALB/c mice. It is concluded that cycloprolylglycine is involved in the endogenous regulation of fear reaction. PMID- 12124646 TI - Modification of radiation-induced changes in the reproductive system of male rats with sodium succinate. AB - We studied the effects of sodium succinate on the reproductive system and blood leukocyte count in rats 30 and 60 days after fractionated irradiation in a dose of 1 Gy. Sodium succinate in concentrations of 0.2 and 0.5% was given ad libitum with water before (1 week) and during irradiation (5 days). The preparation promoted morphofunctional recovery of the testes and epididymides and restored the count of peripheral blood leukocytes. PMID- 12124647 TI - Mechanisms of amnestic effect of ergot alkaloid agroclavin. AB - Neurochemical mechanisms of the effect of agroclavin on spatial memory in the Morris water maze in mice were studied by analyzing the effect of neurotransmitter receptor ligands on the amnestic effect of this alkaloid. D1 receptor agonist SKF-38393 and calcium channel blockers verapamil and nimodipine abolished the negative effect of agroclavin on spatial memory. The role of intracellular calcium in the mechanisms of amnestic effect of agroclavin is discussed. PMID- 12124648 TI - Stress-protective effect of glutapyrone belonging to a new type of amino acid containing 1,4-dihydropyridines on periodontal tissues and stomach in rats with different resistance to stress. AB - Glutapyrone belonging to a new type of amino acid-containing 1,4-dihydropyridines produced a protective effect on periodontal tissues and gastric mucosa in rats with different resistance to stress during acute emotional and pain stress. PMID- 12124649 TI - Anxiolytic and anxiogenic effects of diazepam in male mice with different experience of aggression. AB - Effects of diazepam (0.5 mg/kg intraperitoneally) on aggressive and anxious behavior were studied on male C57Bl/6J mice with different experience of aggression. Diazepam reduced aggression in animals with previous 3- and 20-day experience of aggression, but the plus-maze test revealed an anxiogenic effect of this drug in the former group and anxiolytic effect in the latter group. It was hypothesized that previous aggression experience modified animal sensitivity to diazepam. The effect of the drug depends on psychoemotional status of experimental animals, and this status differs in male mice with short- and long term experience of aggression. PMID- 12124650 TI - Photoprotective activity of melanin preparations from black yeast-like fungus during UV irradiation of human skin: dependence on the concentration. AB - The effect of melanin solutions on the skin exposed to UV irradiation (1,050 kJ/m(2)) depended on its dose and varied from photoprotection (0.005 mg/ml) to photosensitization and phototoxicity (burn, 0.1 mg/ml). These results suggest that doses of melanin preparations should be empirically selected to achieve optimum photoprotective effect. PMID- 12124651 TI - Toxic effect of single treatment with bromantane on neurological status of experimental animals. AB - Neurotoxicological profile of actoprotector bromantane was studied on rats using S. Irwin's protocol of multi-test observation. The drug in doses of 30-300 mg/kg stimulated and in doses of 600-9,600 mg/kg suppressed behavioral activity. Spontaneous motor activity increased after single treatment with bromantane in doses of 30-300 mg/kg, did not change after treatment in doses of 600 mg/kg, and was inhibited after treatment in doses above 600 mg/kg. In doses of 300-600 mg/kg the drug reduced pain sensitivity threshold and in doses above 600 mg/kg elevated the pain threshold and tactile sensitivity and reaction to knock. Bromantane induced mydriasis in all studied doses; in doses above 10 g/kg the preparation induced blepharoptosis. In doses above 5 g/kg bromantane slightly increased respiration rate and depth (Kussmaul-like respiration). In some animals bromantane in high doses induced regurgitation, diarrhea, and polyuria. Rectal temperature decreased by 0.5-1 degrees C after virtually all doses. Behavioral effects of bromantane in doses of 30 and 600 mg/kg were associated with stimulation of the central dopamine and suppression of muscarinic and nicotinic cholinergic structures, n-cholinolytic effects of bromantane was more pronounced at a dose of 30 mg/kg than at a dose of 600 mg/kg. PMID- 12124652 TI - Pine resin and biopin ointment: immunotoxic and allergenic activity. AB - We studied properties of ointment Biopin containing pine resin and used for the therapy of burns, wounds, and purulent and inflammatory diseases. Long-term treatment with this preparation in clinical doses had no effect on the nonspecific, but modulated the specific immune response. Biopin ointment inhibited humoral, but stimulated cell immunity. Neither local irritation, nor allergic reactions were found after long-term epicutaneous application of the preparation. PMID- 12124653 TI - Abnormalities in long-term bone culture from mice deficient in tumor necrosis factor: induction of apoptosis and cell proliferation. AB - The total duration of hemopoiesis and the total cell production in long-term bone marrow cultures from mice deficient by tumor necrosis factor are increased, while proliferation of granulocyte-macrophage precursor cells, the main cell populations in long-term bone marrow cultures, did not differ from that in wild type mice. In bone marrow cultures from knockout mice the intensity of apoptosis remained low during 40-week culturing and was similar to that in early wild-type cultures (10 weeks). Then, the intensity of apoptosis in bone marrow cultures from knockout mice did not differ or even surpassed that in wild-type cultures. However, we observed no inhibition of hemopoiesis in bone marrow cultures from knockout mice. The absence of autocrine expression of tumor necrosis factor did not affect proliferation of precursor cells, but modulate the intensity of apoptosis. It remains unclear whether changes in apoptosis are related to intensive cell production. PMID- 12124654 TI - Polymorphism of promotor sites of interleukins-4 and -10 and tumor necrosis factor-alpha genes in HIV-infected patients. AB - The distribution of allele variants of promotor sites of interleukins 4 (C-590T) and 10 (C-597A) and tumor necrosis factor-alpha (G-308A) genes was studied in HIV infected patients and normal subjects of the Europeoid population in Russia. Some deviations in the distribution of genotypes of the studied polymorphism were revealed in HIV-infected patients compared to the control. The distribution of genotypes in these groups is different for men and women, which is significant for inheritance of allele variants of the cytokine gene. PMID- 12124655 TI - Activity and concentration of cathepsin B as prognostic criteria for the development of mouse LS lymphosarcoma and Lewis lung adenocarcinoma. AB - We measured activity and content of cathepsin B in tumor tissues, liver, and spleen in mice with Lewis adenocarcinoma and LS-lymphosarcoma. Cathepsin B activity in Lewis adenocarcinoma cells was lower than in LS-lymphosarcoma cells, which was probably related to differences in their metastatic properties. Antitumor therapy increased activity and content of cathepsin B in tumor tissues. Changes in the content and activity of cathepsin B in tumor tissues can serve as a prognostic criterion for tumor regression during therapy. Cathepsin B is probably involved in apoptosis of tumor cells during chemotherapy of lymphosarcoma-LS with cyclophosphamide. PMID- 12124656 TI - Role of ionic transport in regulation of hemoglobin affinity for oxygen in diabetes mellitus. AB - The rate of Na(+)/H(+) exchange is increased by 24%, activities of Ca-dependent K+ channels is increased by 13%, and activity of erythrocyte Ca(2+)-ATPase decreased by 17% in patients with diabetes mellitus concomitant with essential hypertension in comparison with patients with essential hypertension without disorders of carbohydrate metabolism. Changes in activity of Na(+)/H(+) exchange, Ca-dependent K(+) channels, and erythrocyte Ca(2+)-ATPase and increased oxygen affinity of hemoglobin are due to increased glucose concentration in the plasma and are leveled by olifen. PMID- 12124657 TI - DNA structure in peripheral blood lymphocytes from patients with chronic viral liver damages. AB - We studied DNA damages (single-strand breaks and alkali-labile sites) in peripheral blood lymphocytes from patients with chronic viral hepatitis and cirrhosis of mixed etiology. The structure of DNA was estimated fluorometrically by changes in the intensity of ethidium bromide fluorescence. Monoinfection with hepatitis B and C viruses was not accompanied by considerable changes in DNA structure in peripheral blood lymphocytes from patients with chronic diseases. The incidence of DNA damages in lymphocytes increased in patients with hepatitis G virus and TTV monoinfection. This is probably related to replication of these viruses in nucleated blood cells. Our results suggest that hepatitis C virus potentiates damaging effect of hepatitis G virus on DNA in lymphocytes. PMID- 12124658 TI - Trophic effects of nootropic peptide preparations cerebrolysin and semax on cultured rat pheochromocytoma. AB - Trophic characteristics of neuroprotectors cerebrolysin and semax were evaluated by their capacity to induce differentiation and improve survival of cultured rat pheochromocytoma (PC12) cells. Morphological signs of cell differentiation (enlargement and formation of processes) were seen 24 h after addition of cerebrolysin into culture medium. Cerebrolysin improved survival of PC12 cells in serum-free medium. In a concentration of 100 microg/ml cerebrolysin decreased the content of apoptotic cells from 32% (control) to 10%. Semax produced no trophic effect on PC12 cells. hence, the neuroprotective effect of cerebrolysin in vivo probably results from trophic activity, while the protective effects of semax are mediated by other mechanisms. PMID- 12124659 TI - Protective effect of calcium channel blocker verapamil on morphological and functional state of hair cells of the organ of corti in experimental kanamycin induced ototoxicity. AB - The protective effect of verapamil against acute ototoxic sensorineural damage produced by intraperitoneal injections of kanamycin (50 mg/kg daily for 14 day) was studied on rats. The functional (otoacoustic emission), histological, and physiological methods proved the protective effect of daily injections of calcium channel blocker verapamil (2 mg/kg) on the state of hair cells of the organ of Corti. PMID- 12124660 TI - Cytological analysis of choledochal mucosa in acute cholangitis in patients with mechanical jaundice. AB - Changes in bile ducts in cholestasis are characterized by activation of proliferation and hypertrophy of cholangiocytes and degenerative and necrobiotic changes in ductal epithelium. Long-term cholestasis led to massive necrosis of cholangiocytes. Inflammatory changes in the choledochal mucosa were most pronounced in patients with choledocholithiasis. PMID- 12124662 TI - Method of bioindication for evaluation of the effects of plant extracts on cell monolayer in acute and chronic experiments. AB - Acute and chronic toxicities of plant extracts for external use were studied on a model of cell monolayer. This method detects the toxic dose of the agent during the first stage of the study; the activity of this dose and severity of damaging effect can be further studied on experimental animals. An advantage of the method is high informative value of preliminary investigation in the development of new low-toxic agents. PMID- 12124661 TI - Study of pharmacological properties of new drugs for external use by the method of cell monolayer bioindication. AB - The effects of new plant extracts on phagocytosis-stimulating, antioxidant, antiinflammatory, and mitotic activities were studied by the method of bioindication on cultures of normal human leukocytes, RH cells, and blood neutrophils. The preparations showed dose-dependent antiinflammatory activity and produced wound-healing, antioxidant, and moderate phagocytosis-stimulating effects. PMID- 12124663 TI - A house of cards: women, aging and spinal cord injury. PMID- 12124664 TI - Women with spinal cord injury and the impact of aging. AB - OBJECTIVES: The objectives of this study were to describe what women with longstanding spinal cord injury (SCI) feel they are experiencing as they age, how they are coping and what they require in order to ensure their continued social and economic participation in society. STUDY DESIGN, METHODS AND SETTING: :A naturalistic approach was taken, incorporating three focus groups (n=10) and key informant interviews (n=19) of women with SCI ranging in age from 31 to 70 years and living in rural and urban communities in Ontario, Canada. RESULTS: The women feel isolated and sense many of their key concerns are ignored or dismissed by health care and service providers. The common physical changes and concerns were gynecological/sexual and bowel and bladder issues. Socio-emotional changes with age included impact of their age-related changes on important relationships and re-evaluation of personal priorities. They articulated worries including declining health, increasing dependency and financial stresses. Additional resources they need to age successfully include improved environmental accessibility, assistive devices, more flexible and responsive attendant and household support, access to recreation and fitness opportunities and peer and psychological support. CONCLUSIONS: Many of the issues raised by the women were consistent with the authors' previous examination of aging in men with SCI and women with disabilities. The most striking difference was their profound sense of isolation and perceptions that health care and service providers were unprepared or unwilling to address the unique issues they face as women living and now aging with SCI. PMID- 12124665 TI - Management of immature heterotopic ossification (HO) of the hip. AB - A case of extensive heterotopic ossification involving the left hip in a 16 year old girl who sustained non traumatic spinal paralysis at T4 ASIA scale A. This case demonstrates the practical difficulties facing clinicians involved in the rehabilitation of this paraplegic patient who required intervention before full maturation of her left hip heterotopic ossification (HO). The patient was developing a rapidly progressive fixed scoliosis and severe difficulty in achieving a proper seating posture. In addition there was difficulty with the application of a suitable orthosis to try and limit the progression of scoliosis as a result of the HO. Discussants will comment on heterotopic ossification in general and the course of action in this particular case. PMID- 12124666 TI - Rasch analysis of the Catz-Itzkovich spinal cord independence measure. AB - BACKGROUND: The Spinal Cord Independence Measure (SCIM) is a new disability scale developed specifically for patients with spinal cord lesions (SCL). Its original and second versions (SCIM and SCIM II) were found to be reliable and more sensitive than the Functional Independence Measure (FIM) to functional changes in SCL patients. OBJECTIVE: To further validate the SCIM II, examining its components on a larger population. DESIGN: Retrospective cohort study. SETTING: Two rehabilitation centers in Israel. SUBJECTS: Two hundred and two inpatients with SCL. INTERVENTIONS: Routine SCIM assessments by staff nurses. Rasch and accompanying analyses. MAIN OUTCOME MEASURES: Unidimensionality of subscales (areas of function); goodness of fit of the tasks to the Rasch model; relationship of total-patient and single-task performance-ability; usability of task categories and the order of threshold locations between them; subscale discrimination of ability and difficulty and hierarchical nature; discrimination of task-categories ability, ie, distribution of thresholds along ability levels; and differential task behavior by age, gender and examination subgroups. RESULTS: Four unidimensional subscales were identified, and an acceptable goodness of fit to the Rasch model was demonstrated in most of their tasks (infit mean square=0.8 1.2, outfit mean square=0.6-1.4). However, some tasks showed overfit (bathing lower body) and some showed misfit (wheelchair-car transfer). Additional analyses performed to check for reasons for less than acceptable fit revealed flaws in a minority of the outcome measures. CONCLUSIONS: The findings of this analysis confirm the validity and reliability of the SCIM II. To a large extent we can infer that the SCIM II construct allows for the detection of any level of disability in any patient with SCL. A few item categories, however, should be rephrased or removed. PMID- 12124667 TI - Effects of coping on psychological outcome when controlling for background variables: a study of traumatically spinal cord lesioned persons. AB - STUDY DESIGN: Cross-sectional. OBJECTIVES: In a previous study we found spinal cord lesion (SCL)-related coping factors to be distinctly related to levels of SCL-related psychological outcome. However, we did not control for other potentially confounding variables. In this study we investigated effects of coping strategies on psychological outcome reactions in traumatically spinal cord lesioned persons controlling for sociodemographic, disability-related and social support variables. SETTING: The Gothenburg Spinal Injuries Unit in Sweden. METHODS: The study sample comprised 255 persons and a subsample of 157 persons. A series of stepwise multiple regression analyses were performed. RESULTS: SCL related coping factors clearly predicted psychological outcome even when background variables were controlled. Higher levels of acceptance coping predicted decreased psychological distress and increased positive morale. Elevated social reliance coping predicted heightened distress. Higher levels of social support predicted lower feelings of helplessness. Sociodemographic and disability-related variables were weak predictors of psychological outcome with one exception: higher education predicted less bitterness and brooding. CONCLUSION: SCL-related coping remained the most important predictor of psychological outcome even when a wide range of variables was controlled. Thus we conclude that psychosocial interventions aimed at helping individuals develop their coping strategies might be of substantial value in their adjustment to SCL. PMID- 12124669 TI - Muscle stretching for treatment and prevention of contracture in people with spinal cord injury. PMID- 12124668 TI - A comparison of heparin/warfarin and enoxaparin thromboprophylaxis in spinal cord injury: the Sheffield experience. AB - OBJECTIVES: To compare the safety and effectiveness of two different thromboprophylactic protocols in the management of patients with spinal cord injury - one using heparin/warfarin and the other using enoxaparin. STUDY DESIGN: Retrospective. SETTING: Princess Royal Spinal Injuries Unit, Sheffield, UK. METHODS: Retrospective review of two cohorts of patients with acute spinal injury admitted to a supra-regional spinal injuries centre and treated with different pharmacological agents. One group received heparin/warfarin in combination with antiembolism stockings and mechanical measures for thromboprophylaxis whereas the second group received enoxaparin in combination with the other measures. Patients who developed clinical symptoms suggestive of deep vein thrombosis or pulmonary embolism were investigated as appropriate. RESULTS: Four of the 101 patients on heparin/warfarin developed symptoms of venous thromboembolism compared to 13 of the 72 who were on enoxaparin. Of the 13, three had been on 40 mg of enoxaparin daily and 10 on 20 mg enoxaparin daily. Six patients on enoxaparin and one patient on warfarin developed thromboembolic complications after they had been mobilised and the anticoagulant discontinued. Eight patients on warfarin prophylaxis and three patients on enoxaparin developed haemorrhagic complications necessitating cessation of therapy. CONCLUSION: This study suggests that the traditional protocol of warfarin/heparin for thromboprophylaxis in spinal cord injury patients remains a safer option than enoxaparin. PMID- 12124671 TI - Breathing pattern in tetraplegic patients. PMID- 12124673 TI - Facilitation of ambulatory function in SCI patients by training and electrical stimulation. PMID- 12124674 TI - Spinal cord stimulation facilitates functional walking in a chronic, incomplete spinal cord injured subject. PMID- 12124677 TI - Overview of anticoagulant drugs for the future. AB - More efficacious, safer, and easier to use anticoagulants are under development. Multiple agents have been shown to be effective in ex vivo or animal thrombosis models and several have progressed to clinical studies. Investigators have not yet determined if pharmaceuticals that inhibit coagulation factor activity earlier in the cascade (for example, inhibitors of tissue factor/factor VIIa, factor IXa, or Xa) are superior to those that block the cascade at a later point. Orally bioavailable drugs for the long-term treatment of thrombotic disorders, particularly those that do not require monitoring, are needed and are under development. Local delivery of anticoagulants or genes modulating anticoagulant control at sites of increased thrombogenicity, such as in diseased arteries, is a promising treatment modality that may decrease systemic bleeding problems. Much about the initiating pathophysiologic events leading to venous thrombotic disease needs to be elucidated before such local therapy can be tested in the venous vasculature. While awaiting better anticoagulants to become routinely available, we need to improve patient management with existing drugs by instituting anticoagulation clinics, promoting patient self-monitoring, and improving efforts to educate patients and health care providers about the use of anticoagulant drugs. PMID- 12124678 TI - Pentasaccharides. AB - Fondaparinux sodium is the first in a new class of antithrombotic agents possessing selective inhibitory activity against factor Xa. The agent was designed and developed with the objective of overcoming the limitations of currently available therapies for the prevention and treatment of venous and arterial thromboembolic disease. Extensive data from preclinical and clinical trials demonstrate fondaparinux's favorable pharmacokinetic profile combined with promising efficacy and safety results in the prevention of venous thromboembolism following major orthopedic surgery, in the treatment of deep venous thrombosis, and in acute coronary syndromes. PMID- 12124679 TI - Issues in the utilization of low molecular weight heparins. AB - Low molecular weight heparins (LMWHs) are parenteral anticoagulants that are widely used for the prevention and treatment of thromboembolic disease. These agents possess several advantages compared to standard heparin, including a more predictable anticoagulant response, better bioavailability allowing for subcutaneous therapy, and a longer half-life. Laboratory monitoring of the LMWHs uses an anti-factor Xa assay, but monitoring is generally not necessary for most patients. However, certain patient populations do benefit from an individualized approach to therapy and, in some cases, therapeutic monitoring, because of an increased bleeding risk and/or relative contraindications to anticoagulant therapy. For example, since LMWHs are cleared by the kidney, they must be used with caution in renal insufficiency. LMWHs are safe and effective during pregnancy but may not be the optimal antithrombotic agent in pregnant women with prosthetic valves. In addition, management around the time of delivery can be difficult because of the bleeding risk, particularly associated with epidural anesthesia. Therapeutic monitoring also may be useful for the morbidly obese, in children, and for patients with malignancy. PMID- 12124680 TI - Improving the safety profile of warfarin. AB - Since warfarin remains the predominate drug administered for long-term anticoagulation, optimizing therapy for maximum antithrombotic effect with minimal bleeding risk continues to challenge clinicians. Genetic differences exist that affect an individual's response to warfarin. The clinician can use genetic information in the future, along with current approaches to improved monitoring and dose schedules, to maintain even more successfully the appropriate therapeutic range for anticoagulation. The international normalized ratio (INR) is a good indicator of warfarin effect and addressing a high INR result often prevents bleeding complications. However, even with a therapeutic INR, patients are still at risk for bleeding. This review will discuss optimizing warfarin dosing, reducing an elevated INR, and managing the anticoagulated patient who has a bleeding event. PMID- 12124681 TI - Direct thrombin inhibitors. AB - Direct thrombin inhibitors interact with thrombin and block its catalytic activity on a wide range of substrates. Their action is in contrast to heparin and its derivatives, which inhibit thrombin and other coagulation serine proteases via antithrombin, and to the warfarin-type drugs that interfere with synthesis of the precursors of the coagulation serine proteases. There are three direct thrombin inhibitors approved for clinical use at present (lepirudin, bivalirudin, argatroban) and another in advanced clinical testing (melagatran/ximelagatran). The chemical structure, kinetics of thrombin inhibition, pharmacokinetics, and clinical use of each of these agents are discussed. PMID- 12124682 TI - Activated protein C: potential therapy for severe sepsis, thrombosis, and stroke. AB - Activated protein C (APC) reduced all-cause 28-day mortality by 19% in patients with severe sepsis (sepsis associated with acute organ dysfunction) in the Protein C Evaluation in Severe Sepsis (PROWESS) trial, leading to recent approval of recombinant APC for treatment of this condition in adults. This review summarizes current knowledge derived from studies of a variety of animal models in which infused human APC demonstrated beneficial activities. Based on in vivo and also in vitro data, APC manifests antithrombotic, profibrinolytic, anti inflammatory, and antiapoptotic activities. APC is a normal circulating component of plasma, derived from the protein C zymogen, and is thus a natural endogenous protective homeostatic factor. Because of its multiple activities, APC has a potential role in the treatment of complex and challenging medical disorders, including thrombosis and stroke. PMID- 12124683 TI - Towards safer thrombolytic therapy. AB - Plasminogen activators (PA) are unique agents that are currently applied as thrombolytic therapy to achieve rapid vascular reperfusion. Regimens of PA plus anticoagulants and antiplatelet drugs have attained a high degree of sophistication and predictable rates of positive clinical outcomes for acute myocardial infarction (MI), ischemic stroke, pulmonary embolism (PE), deep vein thrombosis (DVT), and thrombosed catheters. Included in the repertoire are newly approved mutants of tissue plasminogen activator (TPA), which have biochemical advantages that allow for bolus administration. Yet, despite tremendous effort devoted to enormous trials to establish the clinical efficacy of these agents in acute MI, mortality results are not superior to those with native TPA or streptokinase (SK). Furthermore, all PAs have the potential for hemorrhagic complication, most critically intracranial hemorrhage (ICH), occurring in 0.9% of patients treated with native or mutant TPA. It is possible that a limit of clinical effectiveness has been reached, beyond which more potent PAs do not achieve greater benefit without a serious increase in risk of bleeding. A breakthrough is possible, however, if the risk of ICH could be avoided. One solution is the application of the direct-acting thrombolytic enzyme, plasmin. While intravenous plasmin is not effective when administered systemically, regional infusion to a thrombus induces local thrombolysis. Unlike the PAs, plasmin treatment should not cause hemorrhage from vascular trauma sites, as it is neutralized by antiplasmin in the blood. Animal studies are fully consistent with this approach, which offers potential for achieving a truly regional thrombolytic treatment. PMID- 12124684 TI - Prenatally detected trisomy 7 mosaicism in a dysmorphic child. AB - Trisomy 7 mosaicism was detected prenatally in cultured amniocytes but not in fetal lymphocytes. The child that was born had pigmentary changes of the skin and facial asymmetry suggestive of a chromosomal mosaicism. Skin fibroblasts were studied and trisomy 7 mosaicism was confirmed. At 3 years of age the boy had developed mentally within normal limits. However, dysmorphic findings include sparse hair, short left palpebral fissure, ptosis of the left eyelid, strabismus, enamel dysplasia, low-set and posteriorly rotated ears and undescended testes. These findings share some common features with previously reported cases of trisomy 7 mosaicism. PMID- 12124685 TI - Prenatal diagnosis of congenital heart disease in the Naples area during the years 1994-1999 -- the experience of a joint fetal-pediatric cardiology unit. AB - OBJECTIVES: To analyse the spectrum and frequencies of observed malformations; to evaluate associated extracardiac and chromosomal anomalies and outcomes in each diagnostic category; to demonstrate the need for a multidisciplinary approach to the diagnosis of CHD in the fetus. METHODS: From January 1994 to December 1999, 450 cases of CHD were detected among 4052 pregnancies at risk of fetal CHD seen at our combined unit. Confirmation of the diagnosis was not available in 50 cases, leaving 400 cases for analysis. From our computerized database, the following variables were retrieved and analysed: indication, gestational age at diagnosis, associated extracardiac anomalies, karyotype, natural history, pregnancy and feto-neonatal outcome. RESULTS: CHDs most commonly detected were VSD (75 cases), AVSD (40 cases) and HLH (37 cases). The aneuploidy rate was 29.3% in the 355 cases submitted for karyotyping (25.9% in the whole series), with a prevalence of trisomy 21 and 18 (48 and 30 cases, respectively). The aneuploidy rate was highest for AVSD (80%), coarctation (49%), tetralogy of Fallot and VSD (45%). Associated extracardiac anomalies were present in 29.5% of the cases (118/400). As for pregnancy outcome, there were 150 (37.5%) terminations of pregnancy, 16 (4%) intrauterine fetal deaths and 85 (21.3%) neonatal deaths. The remaining 149 neonates are alive (37.3% survival rate). The termination rate for pregnancies in which CHD was detected at a gestational age <25 weeks was 65.2%. Evolutive changes determined progressive prognostic deterioration in 21 cases (5%), consisting of semilunar valve obstructions and development of ventricular hypoplasia. CONCLUSIONS: The high association rate with extracardiac and chromosomal anomalies (29.3% and 25.9%) and the possible progressive prognostic deterioration require a multidisciplinary team for correct management and follow up. Survival of fetuses with certain CHD is severely reduced, in comparison with postnatal figures, for the common association with aneuploidies. PMID- 12124686 TI - Shortened fetal long bones: a possible in utero manifestation of placental function. AB - Shortened fetal long bones (SFLB) are usually indicative of a skeletal dysplasia. Our aim in this observational retrospective study was to describe a new association between SFLB, small for gestational age (SGA) fetuses and placental abnormalities, and to suggest an aetiologic explanation. During the last decade we have evaluated nine cases in which SFLB (more than 2SD below the mean) was associated with SGA, abnormal maternal serum placental hormones and abnormal placental sonography. Six cases had significantly increased second trimester maternal serum beta hCG and four developed toxaemia of pregnancy or had chronic hypertension. On histology, mature placentas with vascular abnormalities, including chorangiosis, large infarcts and slightly increased syncytial knots were noted. The combination of SFLB, SGA fetuses and placental abnormalities (sonographic, as well as histological) suggested a possible common pathway in the aetiology of this association. PMID- 12124687 TI - Intraventricular haemorrhage in a fetus with cerebral cytomegalovirus infection. AB - Cytomegalovirus (CMV) is the leading infectious cause of prenatal neurological damage, which is particularly severe when primary maternal infection occurs during the first 16 weeks of gestation, at the time of organ development and neuronal migration. Vascular involvement has been suggested to be among the possible pathogenic mechanisms of virus-induced pathology, in addition to direct viral effects. We report on a fetus with cerebral CMV infection, which had intraventricular haemorrhage, together with oligohydramnios and hyperechogenic bowel, following maternal primary CMV infection. PMID- 12124689 TI - Prenatal diagnosis of Pierre-Robin sequence as part of Stickler syndrome. AB - Stickler syndrome or hereditary progressive arthro-ophthalmopathy, is an autosomal dominant condition characterized by ocular manifestations, arthritic changes, orofacial features and deafness, in variable degrees.We report the first case of prenatal diagnosis of Stickler syndrome in a child with a Pierre-Robin sequence (PRS) causing a polyhydramnios. When isolated polyhydramnios is not explained by immunological, metabolic or infectious causes, swallowing difficulty due to PRS must be considered. As PRS is aetiologically heterogeneous, the prognosis depends on the cause. Genetic investigations and familial history must be taken into account. Here, in a context of familial Stickler syndrome, making the prenatal diagnosis of PRS as part of Stickler syndrome allowed us to reassure the parents and to anticipate airway trouble at the child's birth. PMID- 12124688 TI - Outcomes of pregnancies diagnosed with Klinefelter syndrome: the possible influence of health professionals. AB - OBJECTIVE: To describe the association between the outcomes of pregnancies diagnosed with Klinefelter syndrome (KS) and the specialty of the health professional providing pre- and post-diagnostic counselling. METHOD: Data were extracted from the case notes of the 111 cases of KS diagnosed prenatally between 1986 and 1997 in eight geographical regions in five European countries. The data extracted included: outcome of pregnancy, maternal age, social class, parity, gestational age at diagnosis, year of diagnosis and specialties of the health professionals conducting pre- and post-diagnosis consultations. RESULTS: The overall termination rate was 44% (49/111: 95% confidence interval: 35 to 54). Using multivariable logistic regression analysis, the only significant predictor of continuation of the pregnancy was the specialties of the health professionals conducting post-diagnosis counselling: the affected pregnancy was more likely to continue when post-diagnosis counselling involved only a genetics specialist (relative risk: 2.42 (1.14 to 5.92)). CONCLUSION: There is an association between whether or not a woman terminates a pregnancy affected by an unfamiliar fetal anomaly and the professional background of the health professional providing post diagnostic counselling. The causal nature of this association remains to be determined. PMID- 12124690 TI - Psychological impact of the detection of soft markers on routine ultrasound scanning: a pilot study investigating the modifying role of information. AB - OBJECTIVES: To determine the impact on maternal anxiety of detecting a soft marker, and the association between anxiety and the information given during the scan. METHODS: Routine 20-week fetal anomaly scans were audiotaped in the obstetric ultrasound unit of a London teaching hospital, across a four month study period. The study sample comprised 28 pregnant women: 14 in whom a soft marker was detected and a comparison group of 14 women in whom no marker was identified. Telephone interviews were conducted within one week of the scan, at 30 weeks' gestation, and one month after the birth of their children. The main outcome was anxiety, assessed using a standardized scale. Information provided during the scan was coded from transcripts. RESULTS: In the week following the scan, women with soft markers had clinically significant levels of anxiety. At 30 weeks' gestation and one month post-partum their levels were within the normal range. Women who were told during their scan that their baby would probably be all right, compared with women not told this, were significantly less anxious and worried about their baby. CONCLUSIONS: Results from this small longitudinal study suggest that the detection of soft markers on routine prenatal ultrasound causes considerable short-term anxiety for women and that providing reassurance during the scan may prevent some of this anxiety. PMID- 12124691 TI - The effect of ethnic origin on nuchal translucency at 10-14 weeks of gestation. AB - INTRODUCTION: Fetal nuchal translucency (NT) increases with gestation and is affected by fetal posture and fetal gender. A recent report suggested that there might also be ethnic differences. We investigated the effect of ethnic origin on NT in an Asian population. METHODS: NT was measured at 10-14 weeks. The measurements were converted into multiples of the median (MoM) for gestational day. The risk of Down syndrome was calculated by combining NT and maternal age. Cases affected by chromosomal and major structural abnormalities were excluded. NT measurements of different ethnic groups were compared. RESULTS: Between January 1997 and October 2001, 16 981 pregnancies with known ethnic origin and normal fetal outcome were analysed. Median NT MoM (95% CI) of the Filipinos was 1.07 (1.04-1.11). This was significantly higher than that of the Chinese, 1.01 (1.01-1.02); other Asians (Indians, Pakistanis and Nepalese), 0.96 (0.94-0.99), and Caucasians, 0.98 (0.93-1.06) (p<0.05, respectively; Mann-Whitney U-test). An NT risk cut-off of 1:180 would classify 5% of the Chinese, 4.6% of the Caucasians, 5.6% of the Filipinos and 4.2% of the other Asians as screen positive. There were no statistically significant differences between these screen-positive rates (p>0.05, Chi-square test). CONCLUSIONS: Although there were statistically significant differences in NT measurements between different ethnic groups, it was clinically insignificant, as reflected by similar screen-positive rates. PMID- 12124692 TI - Mid-trimester triple test levels in early and late onset severe pre-eclampsia. AB - OBJECTIVE: To study whether the degree of elevation of mid-trimester triple test markers differs in patients with early versus late onset severe pre-eclampsia. METHODS: A retrospective study of the medical records of 102 patients with severe pre-eclampsia for whom mid-trimester triple test result data were available was made. None of these patients had fetuses with abnormal karyotype nor delivered infants with malformations. Pre-eclampsia was defined as early onset when it presented before 32 weeks' gestation. The levels of mid-trimester maternal serum alpha-fetoprotein (MSAFP), human chorionic gonadotrophin (hCG) and unconjugated oestriol (MSuE(3)) in patients with early and late onset severe pre-eclampsia were compared. RESULTS: Twenty-five patients had early onset and 77 patients had late onset severe pre-eclampsia. The two groups did not differ significantly with regard to age, weight, parity and severity of pre-eclampsia. The mean MSAFP in patients with early onset was significantly higher than in patients with late onset severe pre-eclampsia (1.46 MoM, SE 0.12 versus 1.16 MoM, SE 0.05; p=0.01). The mean hCG in the early onset group was also significantly higher than in the late onset group (1.71 MoM, SE 0.18 versus 1.21 MoM, SE 0.07; p=0.03). Mean MSuE(3) levels in patients with early onset were significantly lower than in patients with late onset severe pre-eclampsia (0.83 MoM, SE 0.05 versus 1.02 MoM, SE 0.03; p=0.04). CONCLUSIONS: Higher MSAFP and hCG, and lower MSuE(3), may be more significant markers of early rather than late onset severe pre-eclampsia. PMID- 12124693 TI - Prenatal diagnosis of extrahepatic biliary duct atresia. AB - A rare case of extrahepatic biliary atresia was diagnosed by a combination of prenatal ultrasound and measurements of fetal digestive enzymes in amniotic fluid. Ultrasound at 15 and 18 weeks' gestation failed to detect the gall bladder, and amniotic fluid digestive enzyme values were below the fifth percentile. The patient decided to terminate the pregnancy. Post-abortal pathological examination confirmed the diagnosis. PMID- 12124694 TI - Early prenatal diagnosis of major cardiac anomalies in a high-risk population. AB - OBJECTIVE: To examine the accuracy of early fetal echocardiography performed in a high-risk population combining transvaginal and transabdominal routes. METHODS: A series of 330 high-risk pregnancies were screened by transvaginal and transabdominal scan at 12-17 weeks' gestation in a prospective multicentre trial in Spain between September 1999 and May 2001. A total of 334 fetal heart examinations were performed, including four twin pregnancies. Maternal age ranged from 17 to 46 years (mean 33 years with 36% of women over 34 years). The median gestational age at scan was 14.2 weeks (range 12-17 weeks). For each fetus, visualization of the four-chamber view, the origin of the great arteries, aortic and ductal arches and systemic venous return was attempted in a segmental approach. B-mode and colour/pulsed Doppler flow imaging were used in all cases. The duration of complete heart examination was less than 30 minutes. The examinations were performed by three experienced operators. Reliability was assessed by conventional transabdominal echocardiography at 20-22 weeks, by postnatal follow-up in the first three months of life, and/or by autopsy in cases of termination of pregnancy. RESULTS: The rate of successful visualization of the fetal heart was 94.6% (316/334). In 48 out of 334 (14.4%) fetuses the final diagnosis was abnormal. In 38 out of 48 (79.2%) cases with heart defects the diagnosis was suspected at early echocardiography. In the group with congenital heart defects, 27 cases had an abnormal karyotype (56.3%) and 31 cases showed extracardiac anomalies (64.6%). There were 10 false-negative cases at early scan. There were no false-positive diagnoses. CONCLUSIONS: This experience stresses the usefulness of early fetal echocardiography when performed by expert operators on fetuses specifically at risk for cardiac disease. The high rate of successful visualization of the fetal heart provides a reliable diagnosis of major cardiac defects at this early stage of pregnancy. PMID- 12124695 TI - Achondrogenesis type II with normally developed extremities: a case report. AB - We present a case of achondrogenesis type II with normally developed extremities that was confirmed with postmortem ultrasonographic and radiographic examination. The length of the long bones may vary and the diagnosis of achondrogenesis should not be ruled out with normally developed extremities. Intrauterine sonographic examination of the vertebrae is very important and the absence of vertebral body ossification may be the unique finding of achondrogenesis type II. Axial ultrasonographic images and postmortem plain radiographs are useful to clarify the pathology. PMID- 12124696 TI - Long-term follow-up of infants after transcervical chorionic villus sampling and after amniocentesis to compare congenital abnormalities and health status. AB - OBJECTIVES: Next to procedure-related fetal loss, other adverse effects of invasive prenatal diagnosis have been reported: limb defects after chorionic villus sampling (CVS) or early amniocentesis and respiratory distress after amniocentesis (AC). Because minor abnormalities may be overlooked in routine follow-up, we obtained long-term follow-up data after CVS and AC. METHODS: 1509 women with a singleton pregnancy who had transcervical CVS were matched by age and season of conception with 1509 women with singleton pregnancies who had AC. All procedures were performed during 1985-1991 for advanced maternal age >35 years. Data regarding congenital malformations (classified according Eurocat), neonatal and paediatric morbidity and complications of motor development, speech, hearing and visual function were obtained by questionnaire in 1993-1995. RESULTS: Short-term outcome was known in all but ten infants. Questionnaires with a structured design were mailed to all women with a surviving infant (n=2810); 86.7% responded. No difference was detected between infants after CVS compared with infants after AC regarding congenital malformations (7.2% versus 6.3%), neonatal morbidity (15.1% versus 15.9%), paediatric morbidity with clinical treatment (7.7% versus 6.3%) or outpatient treatment only (43.9% versus 40.3%) and evident function disturbance (2.0% versus 2.0%) or doubtful function disturbance (6.3% versus 6.8%). The number of infants with physical growth <10th centile for Dutch infants was equal (10.1%). Sub-analysis for limb abnormalities or respiratory complications did not demonstrate differences between infants after CVS and AC. Only 10% of all congenital malformations were already known through routine post-partum follow-up. CONCLUSIONS: An extensive long-term survey could not demonstrate differences of health status between infants after prenatal diagnosis by transcervical CVS and AC. PMID- 12124697 TI - Second trimester two-step trisomy 18 screening using maternal serum markers. AB - Trisomy 21 maternal serum marker screening has led to screening for other anomalies, including trisomy 18. Trisomy 18 is generally prenatally diagnosed because of major morphological defects. However, in up to 30% of cases ultrasound signs are unclear, and in most cases diagnosis is performed late in pregnancy. Of the different maternal serum markers, PAPP-A is now considered as the best for trisomy 18 screening. However, pregnancy-associated plasma protein A (PAPP-A) is of value in first trimester screening for trisomy 21, but not in the second trimester. We therefore propose a two-step screening strategy. Based on 45 trisomy 18 cases, we confirm the values of alpha-fetoprotein (AFP) (median 0.61 MoM), free beta-human chorionic gonadotrophin (beta-hCG) (median 0.24 MoM) and of PAPP-A (median 0.08 MoM). In the first step, a 0.5 MoM cut-off for AFP or for free beta-hCG resulted in detection of 37/45 trisomy 18 cases (82%) with a 10% false-positive rate. The second step consisted of the measurement of PAPP-A for all these false-positive cases. Using a PAPP-A cut-off of 0.5 MoM, all the 37 trisomy 18 cases were detected, but now with a 0.1-0.2% false-positive rate. Amniocentesis was only offered to these few patients. This two-step second trimester screening will be of value for patients who have not been included in first trimester screening based on nuchal translucency (NT) measurement combined with the first trimester markers, PAPP-A and free beta-hCG. PMID- 12124698 TI - Fetal gender and aneuploidy detection using fetal cells in maternal blood: analysis of NIFTY I data. National Institute of Child Health and Development Fetal Cell Isolation Study. AB - OBJECTIVES: The National Institute of Child Health and Human Development Fetal Cell Isolation Study (NIFTY) is a prospective, multicenter clinical project to develop non-invasive methods of prenatal diagnosis. The initial objective was to assess the utility of fetal cells in the peripheral blood of pregnant women to diagnose or screen for fetal chromosome abnormalities. METHODS: Results of fluorescence in situ hybridization (FISH) analysis on interphase nuclei of fetal cells recovered from maternal blood were compared to metaphase karyotypes of fetal cells obtained by amniocentesis or chorionic villus sampling (CVS). After the first 5 years of the study we performed a planned analysis of the data. We report here the data from 2744 fully processed pre-procedural blood samples; 1292 samples were from women carrying singleton male fetuses. RESULTS: Target cell recovery and fetal cell detection were better using magnetic-based separation systems (MACS) than with flow-sorting (FACS). Blinded FISH assessment of samples from women carrying singleton male fetuses found at least one cell with an X and Y signal in 41.4% of cases (95% CI: 37.4%, 45.5%). The false-positive rate of gender detection was 11.1% (95% CI: 6.1,16.1%). This was higher than expected due to the use of indirectly labeled FISH probes in one center. The detection rate of finding at least one aneuploid cell in cases of fetal aneuploidy was 74.4% (95% CI: 76.0%, 99.0%), with a false-positive rate estimated to be between 0.6% and 4.1%. CONCLUSIONS: The sensitivity of aneuploidy detection using fetal cell analysis from maternal blood is comparable to single marker prenatal serum screening, but technological advances are needed before fetal cell analysis has clinical application as part of a multiple marker method for non-invasive prenatal screening. The limitations of the present study, i.e. multiple processing protocols, are being addressed in the ongoing study. PMID- 12124700 TI - Womens' preference in Down syndrome screening. AB - OBJECTIVE: To determine the knowledge of pregnant women about prenatal tests, and what tests they would choose if offered. Also, the preference of pregnant women for second-trimester or first-trimester screening was assessed. PATIENTS AND METHODS: Pregnant women receiving antenatal care in a decentralized primary care system (n=80), and pregnant women that were offered a prenatal diagnosis at the Academic Medical Centre (n=195), were asked to complete a questionnaire. RESULTS: The response rate was over 80%. Most women in both groups preferred a screening test for Down syndrome to be performed in the first trimester of pregnancy. A combination of nuchal translucency measurement and first-trimester serum screening was the option of choice. The screening possibilities for Down syndrome were less well known to the women in the low-risk group compared with the women in the high-risk group. The offer of a prenatal screening test would have been declined by more than 30% of women at low risk for carrying a fetus with Down syndrome. CONCLUSIONS: Our results show that women prefer screening for Down syndrome to be performed in the first trimester of pregnancy, using both serum and ultrasound tests. In women at low risk for Down syndrome the knowledge of prenatal screening methods was less, as well as the acceptance of prenatal screening being lower. PMID- 12124699 TI - Prenatal ultrasonographic detection of gastrointestinal obstruction: results from 18 European congenital anomaly registries. AB - OBJECTIVES: We evaluated the prenatal detection of gastrointestinal obstruction (GIO, including atresia, stenosis, absence or fistula) by routine ultrasonographic examination in an unselected population all over Europe. METHODS: Data from 18 congenital malformation registries in 11 European countries were analysed. These multisource registries used the same methodology. All fetuses/neonates with GIO confirmed within 1 week after birth who had prenatal sonography and were born during the study period (1 July 1996 to 31 December 1998) were included. RESULTS: There were 670 793 births in the area covered and 349 fetuses/neonates had GIO. The prenatal detection rate of GIO was 34%; of these 40% were detected < or = 24 weeks of gestation (WG). A total of 31% (60/192) of the isolated GIO were detected prenatally, as were 38% (59/157) of the associated GIO (p=0.26). The detection rate was 25% for esophageal obstruction (31/122), 52% for duodenal obstruction (33/64), 40% for small intestine obstruction (27/68) and 29% for large intestine obstruction (28/95) (p=0.002). The detection rate was higher in countries with a policy of routine obstetric ultrasound. Fifteen percent of pregnancies were terminated (51/349). Eleven of these had chromosomal anomalies, 31 multiple malformations, eight non chromosomal recognized syndromes, and one isolated GIO. The participating registries reflect the various national policies for termination of pregnancy (TOP), but TOPs after 24 WG (11/51) do not appear to be performed more frequently in countries with a liberal TOP policy. CONCLUSION: This European study shows that the detection rate of GIO depends on the screening policy and on the sonographic detectability of GIO subgroups. PMID- 12124701 TI - Prenatal diagnosis of Niemann-Pick diseases types A, B and C. AB - Prenatal diagnosis of Niemann-Pick disease types A and B is routinely accomplished by sphingomyelinase assay. For Niemann-Pick type C disease, demonstration of an abnormal intracellular cholesterol trafficking is a complex procedure, and mutational analysis (NPC1 or NPC2/HE1 gene), whenever feasible, represents a major advance. PMID- 12124702 TI - Reporting partial screening results: is it confusing and unsatisfactory? PMID- 12124704 TI - Poland sequence and hyperhomocyst(e)inaemia. PMID- 12124705 TI - Microcephaly with dysgenesis of corpus callosum and colpocephaly in the survivor after the first-trimester death of a monochorionic co-twin. PMID- 12124706 TI - Hyperechogenic bowel loops and meconium ileus in a fetus carrying the D1152H and G542X cystic fibrosis CFTR mutations. PMID- 12124707 TI - Current awareness in prenatal diagnosis. PMID- 12124708 TI - Area ablation: a new lasing concept provides significantly enhanced acute and long-term results for treatment of in-stent restenosis. AB - BACKGROUND AND OBJECTIVES: Debulking is still a technique of choice for in-stent restenosis (ISR). Excimer laser debulking has enabled high procedural success with very low complication rates, but has demonstrated markedly heterogeneous results owing to differences in lasing and laser technology, and selected patient populations. Since new area-ablation technique enables ablation of larger areas than its own device size, we have evaluated their effectiveness and safety in an uncontrolled study. STUDY DESIGN/MATERIALS AND METHODS: Fifty-three patients with diffuse ISR were treated by laser area ablation, followed by adjunctive balloon angioplasty. Primary endpoint was percent of binary stenosis at 6-month follow up; secondary endpoints were procedural success; target lesion revascularization (TLR); major adverse cardiac events (MACE); diameter stenosis (DS); and minimal lumen diameter (MLD) before and after laser debulking, and at 6-month follow-up. RESULTS: Laser debulking was feasible (as defined as < or =30% residual stenosis) in 98.1% of patients. At 6-month follow-up, binary stenosis was 26.4%; angiographic TLR, 20.7%; and MACE, 3.7%. DS decreased from 87+/-17% to 20 +/- 9% after laser debulking, and to 9+/-7% after PTCA; it was 29+/-14% at follow-up (P values in comparison to baseline: 0.0047; 0.0036; 0.0064). MLD increased from 0.6+/-0.3 to 2.4+/-0.5 mm after laser debulking, to 2.8+/- 0.6 mm after adjunctive PTCA, and to 1.9 +/- 0.4 mm at follow-up (P-values in comparison to baseline: 0.0059; 0.0031; 0.0088). CONCLUSIONS: Owing to a significantly greater MLD, area ablation facilitates significantly enhanced immediate and follow-up results for diffuse ISR, including a simpler and more effective laser-debulking procedure than former lasing techniques. PMID- 12124709 TI - A comparative study of retinal effects from continuous wave and femtosecond mode locked lasers. AB - BACKGROUND AND OBJECTIVES: In order to provide a direct comparison of the effects of mode-locked systems to those with continuous-wave (CW) or nonpulsed output, we have performed an experiment with lasers possessing otherwise identical output characteristics. This in vivo minimum visible lesion study compares retinal effects of mode-locked and CW lasers complete with histopathology of the treated areas. STUDY DESIGN/MATERIALS AND METHODS: Titanium:Sapphire lasers produced 800 nm output for either mode-locked (76 MHz repetition rate, 120 femtoseconds) or CW exposures. Alternating CW and mode-locked laser exposures were delivered to the paramacular retinal region of rhesus subjects. Laser exposure duration was set to one-quarter second for both types of exposures. Through ophthalmoscopic examination of the fundus, a minimal visible lesion (MVL) threshold for damage was established. RESULTS: Approximately 75 test sites for each type of exposure were examined. The laser dosage thresholds and 95% confidence intervals for minimal visible damage at 24 hours postexposure were found to be 5.9 mJ (5.23-6.6 mJ) and 5.84 mJ (5.23-6.58 mJ) for mode-locked and CW exposures, respectively. CONCLUSIONS: Results are compared with published studies conducted at similar exposures. These nearly identical damage thresholds indicate a primarily thermal tissue damage mechanism. Comparative histopathology of acute and chronic lesions of both exposure types is also presented. PMID- 12124710 TI - Photoeradication of Helicobacter pylori using 5-aminolevulinic acid: preliminary human studies. AB - BACKGROUND AND OBJECTIVES: Helicobacter pylori (HP) is an endemic pathogenic bacterium causing gastritis and gastroduodenal ulceration in humans and is linked to the development of gastric malignancies. These first human in vivo studies investigated the photoeradication of HP using laser and white light. STUDY DESIGN/MATERIALS AND METHODS: In 13 HP-positive volunteers, a zone of gastric antrum was irradiated with laser (410 nm, 50 J/cm(2)) or endoscopic white light (10 J/cm(2)) 45 minutes after oral 5-aminolevulinic acid (5-ALA) 20 mg/kg. HP eradication was assessed by biopsy urease test and HP-culture from irradiated and control zones 5 minutes, 4 and 48 hours post-irradiation. RESULTS: A maximum eradication effect was achieved at 4 hours post-irradiation when 85% of biopsies in the monochromatic and 66% in the white light exposed zones, and 58 and 33% in the respective control zones were HP-negative. CONCLUSIONS: HP numbers were greatly reduced following exposure to 5-ALA and either laser or white light in vivo. Photoeradication appears feasible, but further light dosimetry and the development of convenient application methods is required. PMID- 12124711 TI - Lichenoid dermatitis--treatment with pulsed dye laser: a case study. AB - BACKGROUND AND OBJECTIVES: Both the diagnosis and the treatment of lichenoid dermatosis are often difficult and can be time-consuming. There are now more and more publications about the use of laser systems--especially the flashlamp-pumped pulsed dye laser--in the treatment of inflammatory dermatoses, although the laser's exact mechanism of action in these cases is not yet clear. STUDY DESIGN/PATIENTS AND METHODS: We report on a female patient with lichenoid dermatitis that was presumably drug-induced (roxatidine, H(2) receptor antagonists). After a 10-month treatment with local corticosteroids, without significant clearance the drug was discontinued and the pulsed dye laser was used (wavelength 585 nm, pulse duration 450 microseconds). RESULTS: Six laser treatments resulted in complete clearance of the lesions. No recurrence occurred during the follow-up period of 54 months. Scars were not observed. A post operative biopsy showed no evidence of lichenoid dermatitis. CONCLUSIONS: The pulsed dye laser seems to accelerate the clearance of presumably drug induced corticosteroid-resistant lichenoid dermatoses. No permanent pigmental changes or scarring were observed. PMID- 12124712 TI - Cryogen spray cooling efficiency: improvement of port wine stain laser therapy through multiple-intermittent cryogen spurts and laser pulses. AB - BACKGROUND AND OBJECTIVES: Cryogen spray cooling (CSC) is used to minimize the risk of epidermal damage during laser treatment of port wine stain (PWS) birthmarks. Unfortunately, CSC may not provide the necessary protection for patients with high concentrations of epidermal melanin. The objectives of this study are to: (1) provide a definition of cooling efficiency (eta) based on the amount of heat removed per unit area of skin for a given cooling time; (2) using this definition, establish the eta of previously reported spray nozzles; (3) identify the maximum benefit expected in PWS laser therapy based solely on improvement of eta; and (4) study the feasibility of using multiple-intermittent cryogen spurts and laser pulses to improve PWS laser therapy. STUDY DESIGN/MATERIALS AND METHODS: A theoretical definition to quantify eta is introduced. Subsequently, finite difference heat diffusion and Monte Carlo light distribution models are used to study the spatial and temporal temperature distributions in PWS skin considering: (1) the current approach to PWS laser therapy consisting of a single cryogen spurt followed by a single pulsed dye laser exposure (SCS-SLP approach); and (2) multiple cryogen spurts and laser pulses (MCS-MLP approach). At the same time, an Arrhenius-type kinetic model is used to compute the epidermal and PWS thermal damages (Omega(E) and Omega(PWS), respectively) for a high epidermal melanin concentration (20%), corresponding to skin types V-VI. RESULTS: The eta corresponding to a wide range of heat transfer coefficients (h) is quantified. For reported CSC nozzle devices eta varies from 40 to 98%. Using the SCS-SLP approach, it is shown that even eta = 100% cannot prevent excessive Omega(E) for a skin types V-VI. In contrast, the MCS-MLP approach provides adequate epidermal protection while permitting PWS photocoagulation for the same skin types. CONCLUSIONS: The new proposed definition allows to compute the cooling efficiency of CSC nozzle devices. Computer models have been developed and used to show that the SCS-SLP approach will not provide adequate epidermal protection for darker skin patients (skin types V-VI), even for eta = 100%. In contrast, the MCS-MLP approach may be a viable solution to improve PWS laser therapy for darker skin patients. PMID- 12124713 TI - Improved microvessel repair: laser welding with an anti-thrombotic solder. AB - BACKGROUND AND OBJECTIVES: Concentrated protein solutions can be used as thermally polymerized solders in laser welding. Solders supplemented with biologically active chemicals may provide in situ drug delivery for localized therapeutics. These studies characterize a serum albumin (SA) solder containing heparin, designed to reduce microvascular thrombosis rates. STUDY DESIGN/MATERIALS AND METHODS: Samples of heparin added to 30% SA to obtain heparin-to-albumin molar ratios (HAMR) of 4:1 and 2:1 were thermally polymerized, and heparin release into saline was measured. Using a rat thrombosis model, patency was determined for suture, and 0 U/ml (control), 2.5 U/ml, 50 U/ml heparin solder repairs. RESULTS: Heparin release was five times higher for 4:1 than 2:1 HAMR solder acutely, but was equivalent after 2 days. Animal patency rates were: 50% suture, 0% control, 50% low heparin, 66% high heparin (P < 0.05 vs. control). CONCLUSIONS: Solders incorporating heparin should provide in situ anti-thrombotic therapy reducing the risk of microvascular thromboses. PMID- 12124715 TI - Measurement of adhesive forces between S. epidermidis and fibronectin-coated surfaces using optical tweezers. AB - BACKGROUND AND OBJECTIVES: Biomaterial-mediated infection, a common cause of medical device failure, is initiated by bacterial adhesion to an adsorbed protein layer on the implant surface. This adhesion is thought to be mediated by specific molecules present on the bacterial cell surface. Optical tweezers can be used to measure the adhesive force between a single bacterium and a protein-coated surface. STUDY DESIGN/MATERIALS AND METHODS: Using optical tweezers, a bacterium was trapped and brought in contact with a 10-microm diameter polystyrene microsphere coated with fibronectin. The minimum force required to detach the cell from the bead was determined over a range of fibronectin concentrations and contact times. RESULTS: The detachment forces were integer multiples of an 18-pN base value that was independent of contact time and coating concentration; we propose that the variation in force is related to the number of bonds formed. CONCLUSIONS: These experiments demonstrate that optical tweezers can be used to investigate the adhesion of individual bacteria to surfaces. The results suggest that S. epidermidis has surface proteins capable of binding fibronectin. PMID- 12124714 TI - Diamond laser scalpel vs. steel scalpel: a side by side comparison of cutaneous wound healing. AB - BACKGROUND AND OBJECTIVES: To compare the cutaneous wound healing using the diamond laser scalpel with wound healing using a steel scalpel and electrocoagulation for hemostasis. STUDY DESIGN/MATERIALS AND METHODS: A prospective and randomized, comparative trial was conducted on eighteen patients. Fusiform excisions were performed using the diamond laser scalpel on one half of each excision and a steel scalpel with electrocoagulation for hemostasis on the other half. The Clinicon SureBlade diamond laser scalpel was used with the Luxar CO(2) attachment at the 6-8-W settings. Blinded assessment of adverse events and photographs were taken at 1 day, 7-10 days, 4 weeks, and 8-12 weeks. The final scar was evaluated at 8-12 weeks for cosmetic outcome and three physicians blinded to the method of excision evaluated photographs of the wounds. Histologic evaluation was performed on all excisions for collateral thermal damage. RESULTS: Investigator assessment showed no statistically significant differences between the diamond laser scalpel side and the steel scalpel side with respect to bleeding, bruising, swelling, pain, dehiscence, or final scar appearance. The mean residual thermal damage was 350.3 microm (95% CI +/- 37 microm, P < 0.001). The diamond laser scalpel scored higher on intra-operative coagulation (P = 0.20) although these differences were not statistically significant. CONCLUSIONS: The cosmetic outcomes of cutaneous excisions performed with the diamond laser scalpel are equivalent to excisions performed with steel scalpels with electrocoagulation for hemostasis. PMID- 12124716 TI - Photothermal time-resolved imaging of living cells. AB - BACKGROUND AND OBJECTIVES: Thermal effects of laser radiation at cell level play very important role in cell functioning and in many laser applications. The aim of this study was to evaluate a new method of photothermal imaging (PTI) for monitoring short-time nano-scale thermal effects in individual living cells. STUDY DESIGN/MATERIALS AND METHODS: PTI is based on the irradiation of a cell with a short laser pump pulse (8 nanoseconds, 532 nm) and on registration of the laser-induced local thermal effects using time-resolved phase-contrast imaging with a pulsed probe laser. RESULTS: PT images of lymphocytes, lympholeukemia cells in vitro were obtained at different laser energies. PTI in time-resolved mode allowed visualizing the structures with size less than diffraction limit (90 nm liposomes). The photodamage process was visualized for a single human leukocyte in suspension. CONCLUSIONS: PTI in non-invasive mode offered better contrast of living cell image than conventional optical phase-contrast microscopy. The data obtained showed that PTI is in perspective for studies of live non-fluorescent cells. PMID- 12124717 TI - Nd:YAG laser therapy for palliation of recurrent squamous cell carcinomas in the oral cavity. AB - BACKGROUND AND OBJECTIVES: The objective of this study was to evaluate the outcome of laser photo-thermoablation for palliation of recurrent squamous cell tumors of the oral cavity. STUDY DESIGN/PATIENTS AND METHODS: Seventeen patients were treated with the Nd:YAG laser (power output was 50 W) delivered through a curved oral handpiece. RESULTS: Ten patients are alive, 7 with tumor remission, and 3 with persistent disease with an average follow-up of 16 months (range = 2 36). A total of 29 tumor sites received laser treatment with 17 (58%) completely ablated. Stratified by tumor site Nd:YAG treatment led to complete local response in 8/10 buccal mucosa, 2/5 retromolar trigone, 2/2 tongue, 2/5 gingiva, 1/2 floor of mouth, 2/4 hard palate. CONCLUSIONS: Nd:YAG laser treatment of recurrent oral cavity squamous cell carcinoma can be performed safely and repeated as needed to achieve tumor palliation. However, extended follow-up may be needed before convincing evidence of long-term therapeutic benefits is obtained. PMID- 12124718 TI - A tribute to Harriet H. Werley, founding editor. PMID- 12124719 TI - Patient-centered interventions. AB - Patient-centered care is valued in nursing. However, until recently, nurse researchers have focused on testing the effects of standardized rather than patient-centered interventions (PCIs). The latter are those interventions that are altered to address selected patient characteristics (e.g., beliefs, habits, or goals). PCIs have been well received, and in some studies they have been associated with improved health outcomes. In this article we describe briefly the concept patient centered, summarize the development of research on PCIs, discuss kinds of PCIs, provide examples of PCIs and how they have been derived and implemented, and raise issues for theory and future research. PMID- 12124721 TI - Trajectories of cognitive recovery following a minor brain injury. AB - Minor brain injury (MBI) is the most frequently diagnosed head trauma in the United States, with treatment costing more than $1.5 billion annually and many patients incapacitated for months following injury. The purpose of this study was to characterize the brain function disruptions associated with MBI and to determine the time trajectory of recovery, using a theoretical model of attention. Distractibility, impulsivity, irritability, and impaired executive functioning were demonstrated in all participants during the 24 hr after injury. Twenty percent of participants continued to complain of distractibility, impulsivity, and/or irritability throughout the 30-day study. Loss of consciousness was shown to confound participants' healing trajectories. These results suggest that standard emergency room treatment following MBI is inadequate and should include discharge directives to reduce cognitive demands for at least 48 hr at a minimum, for 30 days or longer for those with loss of consciousness. PMID- 12124720 TI - Measuring the hospital practice environment: a Canadian context. AB - The primary purpose of this study is to document the psychometric properties of the revised Nursing Work Index (NWI-R) in the context of a large Canadian sample of registered nurses. A self-administered survey containing the NWI-R was completed by 17,965 registered nurses working in 415 hospitals in three Canadian provinces. Using exploratory principal components analysis, with a forced one factor solution, the practice environment index was obtained. In addition, key assumptions were tested from previous work about the rationale for the aggregation of NWI-R responses. In the Canadian context the one-factor solution provides a parsimonious index of the practice environment of registered nurses working in acute care hospitals. Further work is needed to determine the predictive capability of this index and its relevance to cross-national organizational contexts. PMID- 12124722 TI - An analysis of the relationship between job satisfaction and job stress in correctional nurses. AB - Stamps and Piedmonte's Index of Work Satisfaction and Harris's Nurse Stress Index were completed by 287 registered nurses employed in state prison health care facilities in order to assess job satisfaction and job stress among correctional nurses. Correctional nurses' expectations about job satisfaction were influenced by pay and autonomy. This finding was consistent with studies of hospital nurses. Important sources of job satisfaction were professional status and interaction with employees. Analysis of differences between expectations and sources of job satisfaction may provide understanding of career benefits and sources of dissatisfaction. Time pressures and organizational support and involvement were sources of stress. Multivariate analyses showed an inverse relationship between stress and job satisfaction. Information about job satisfaction and work stress and their correlates may be used to develop strategies to improve the recruitment and retention of correctional nurses. PMID- 12124723 TI - Cultural differences in responses to a Likert scale. AB - Cultural differences in responses to a Likert scale were examined. Self identified Chinese, Japanese, and Americans (N=136, 323, and 160, respectively) recruited at ethnic or general supermarkets in Southern California completed a 13 question Sense of Coherence scale with a choice of either four, five, or seven responses in either Chinese, Japanese, or English. The Japanese respondents more frequently reported difficulty with the scale, the Chinese more frequently skipped questions, and both these groups selected the midpoint more frequently on items that involved admitting to a positive emotion than did the Americans, who were more likely to indicate a positive emotion. Construct validity of the scale tended to be better for the Chinese and the Americans when there were four response choices and for the Japanese when there were seven. Although culture affected response patterns, the association of sense of coherence and health was positive in all three cultural groups. PMID- 12124724 TI - Wives giving care to husbands with Alzheimer's disease: a process of interpretive caring. AB - Wives giving care to spouses with dementia are a particularly vulnerable segment of the caregiving population. In this article a grounded theory study of 20 such wives is described, with their experiences explained as a process of interpretive caring. Wives began the process by either seeing changes in their husbands or recognizing changes in their work. Following this, the wives moved on to a phase of drawing inferences about what they observed and then took over their husbands' roles and responsibilities. These changes prompted the wives to rewrite identities for their husbands that incorporated the dementia and to rewrite identities for themselves to reflect their new roles, abilities, and strengths. Finally, the wives set about constructing a new daily life to sustain both partners. This process is neutral and allows for positive aspects of caring to be considered along with grief and frustration. PMID- 12124725 TI - Nursing intervention studies: a descriptive analysis of issues important to clinicians. AB - When reading a report of an intervention study, clinicians are interested in knowing: whether the intervention is effective, with whom it is effective, how much benefit it produces, and whether associated, adverse outcomes occur. Recommendations have been made in the research literature regarding how to conduct and report intervention studies so as to produce knowledge regarding these questions. This descriptive study was conducted to estimate the frequency with which these recommendations are being used in nursing intervention studies. Data pertinent to five research questions were extracted from 84 experimental and quasi-experimental study reports published between 1998 and 2000. Seventeen percent of the studies used a design that could statistically test for variation in intervention effect depending on the level of an individual characteristic. However, a test of interaction was actually conducted in only 8% of the studies. The magnitude of the intervention's effect was addressed in 38% of the study reports. Providing the proportion of persons in the intervention group who attained a discrete outcome was the most frequently used way of showing intervention magnitude. Associated, adverse outcomes were examined in 23% of the studies, and were most often measured as continuous variables. The low level of use of recommended methods leads the author to suggest dialogue between clinicians and researchers to determine if intervention studies are being conducted and reported in ways that produce knowledge that is useful to clinicians. PMID- 12124726 TI - Genome-wide linkage analysis for celiac disease in North American families. AB - Celiac disease (CD) is an autoimmune disease caused by sensitivity to the dietary protein gluten. It has a prevalence of 1 in 250 in the United States. Multiple case families are common with a risk to siblings from 10-12%. Previous linkage studies have found no significant evidence for linkage other than to HLA. In this study, we performed a genome-wide search on 62 families with at least two cases of CD to identify non-HLA loci for CD. Two-point and multipoint parametric and nonparametric analyses were performed on the entire set of families and on sets stratified by the HLA genotype. Accounting for multiple testing, we found genome wide intermediate linkage evidence at 18q (heterogeneity LOD (HLOD) = 3.6) and at 3p (HLOD = 3.2) and suggestive evidence at 5p (HLOD = 2.7). Good consensus between two-point and multipoint evidence was not found, and after genotyping with new markers in these regions, our results were inconclusive. The 18q region had intermediate two-point evidence, but weak multipoint evidence. At 3p and 5p, the addition of follow-up markers added flanking support, yet multipoint evidence was still lacking. Our results indicate that multipoint analyses may be hindered by the complexity of CD. Multipoint analyses are not robust to model misspecification, and further development of models is needed. Additional study of these and other families is necessary to validate or rule out the regions implicated in this study. PMID- 12124728 TI - Supernumerary marker chromosomes 5: confirmation of a critical region and resultant phenotype. AB - A critical region exists at 5p13 for the phenotype associated with duplication 5p. Two unrelated Polynesian children are reported with supernumerary marker chromosomes (SMCs) 5. This brings to seven the total of reported SMCs derived from chromosome 5. The phenotype is clear, and the level of mosaicism of the marker chromosomes is contributory to the clinical severity. PMID- 12124727 TI - Clinical findings and biochemical and molecular analysis of four patients with holocarboxylase synthetase deficiency. AB - Holocarboxylase synthetase (HLCS) deficiency (HLCSD) is a rare autosomal recessive disorder of biotin metabolism. HLCS catalyzes the biotinylation of the four human biotin-dependent carboxylases. Using the newly available human genomic sequence, we report the map of HLCS genomic structure and the predicted exon/intron boundaries. Moreover, the molecular studies of four patients (two Italians, one Iranian, and one Australian) affected by HLCS deficiency are here reported. The clinical findings, the age of onset, and response to biotin treatment differed between our patients. The diagnosis was made by organic acid analysis and confirmed by enzymatic analysis in three patients. Six mutations in the HLCS gene were identified, including two novel (N511K and G582R) and four known missense mutations (L216R, R508W, V550M, and G581S). Five of the mutations are localized within the HLCS biotin-binding domain, whereas the L216R amino acid change is located in the N-terminal region outside of the putative biotin-binding domain. This mutation, previously reported in a heterozygous state, was detected for the first time in a patient with homozygous status. The patient's severe clinical phenotype and partial responsiveness to biotin support a genotype phenotype correlation through the involvement of residues of the N-terminal region in a substrate specificity recognition or regulation of the HLCS enzyme. PMID- 12124729 TI - Exclusion of PITX2 mutations as a major cause of CHARGE association. AB - CHARGE is a nonrandom association of ocular coloboma, congenital heart defects, atresia of the choanae, retarded growth and development, genital hypoplasia, and ear anomalies including deafness. The cause of CHARGE remains unknown; however, there is considerable evidence of an underlying genetic basis, as discussed by Tellier et al. [1996: Clin Genet 50:548-550; 1998: Am J Med Genet 76:402-409] and by Martin et al. [2001: Am J Med Genet 99:115-119]. Based on the ocular, cardiac, and craniofacial expression pattern of Pitx2, a homeodomain transcription factor, and the pleiotropic effects of loss of PITX2 function in both mouse and human, we hypothesized that PITX2 mutations may contribute to the multiple phenotypic anomalies present in CHARGE individuals. By direct sequencing of DNA from 29 individuals with CHARGE, we did not identify any mutations in PITX2. We did, however, identify two PITX2 sequence polymorphisms. Large deletions of PITX2 were excluded in most patients by heterozygosity in at least one of several polymorphic markers near the PITX2 locus. Together, these data indicate that PITX2 mutations are unlikely to be a major contributing cause of the multiple anomalies present in individuals with CHARGE. PMID- 12124730 TI - Frameshift mutation in the cartilage-derived morphogenetic protein 1 (CDMP1) gene and severe acromesomelic chondrodysplasia resembling Grebe-type chondrodysplasia. AB - Grebe-type chondrodysplasia exhibits a severe form of limb shortening and appendicular bone dysmorphogenesis. Here we report a family with seven males and six females who inherited the disorder in an autosomal recessive fashion. While the carrier parents did not exhibit any apparent skeletal abnormalities, all affected patients had a similar phenotype with unaffected axial and craniofacial bones. Since mutations in the cartilage-derived morphogenetic protein 1 (CDMP1) gene have been reported in similar acromesomelic chondrodysplasias, we examined genomic DNA from affected and normal subjects for possible mutations in CDMP1. In affected subjects, an insertion of a C at nucleotide 297 of the coding sequence was discovered. This insertion produced a shift in the reading frame at amino acid residue 99, causing premature termination of the polypeptide six amino acids downstream. DNA samples from 41 control subjects did not show this mutation. The truncated CDMP1 protein in these subjects is predicted to cause a total loss of its signaling function. The present report confirms that CDMP1 plays an important role in the regulation of axial bone growth during development and suggests that its absence does not impair other developmental processes. PMID- 12124731 TI - Maternal isodisomy for 14q21-q24 in a man with diabetes mellitus. AB - We report a 20-year-old man with maternal uniparental disomy for chromosome 14 (UPD14) and maturity-onset diabetes mellitus (DM). He had pre- and postnatal growth retardation, developed DM at age 20 years without any autoimmune antibodies, and had a mosaic 45,XY,der(14;14)(q10;q10)[129]/46,XY,+14,der(14;14)(q10;q10)[1] karyotype. Allelotyping using microsatellite markers covering the entire 14q indicated segmental maternal isodisomy for 14q21-q24 and maternal heterodisomy of the remaining regions of the chromosome. It is thus tempting to speculate that the segmental isodisomy led to reduction to homozygosity for a mutant gene and thus caused his DM, although the possibility of coincidental occurrence of the two events cannot totally be ruled out. Fluorescence in situ hybridization (FISH) analysis using BAC clone probes revealed that the isodisomic segment did not overlap any known IDDM or NIDDM susceptibility loci on chromosome 14, suggesting a novel locus for a subset of DM at the isodisomic segment. PMID- 12124732 TI - Generalized abnormal embryonic development in missed abortion: embryoscopic and cytogenetic findings. AB - A direct view of the embryo by means of transcervical embryoscopy prior to evacuation in 154 cases of missed abortion showed general embryonic maldevelopment in 48 cases (31%). A successful cytogenetic evaluation of these growth-disorganized embryos was performed in 37. Chromosomal abnormalities were found in 26 cases (70%), with autosomal trisomies in 24 cases (92%). Trisomies involved chromosome 3 (one case), 6 (one case), 8 (two cases), 10 (one case), 12 (two cases), 14 (one case), 16 (11 cases), 20 (one case), and 22 (four cases). Most of these chromosome abnormalities represented nonviable defects, and their presence explained the minimal embryonic development observed embryoscopically. An apparently normal karyotype was observed in 11 growth-disorganized embryos whose maldevelopment was similar to that resulting from the trisomies listed above. The factors responsible for embryonic maldevelopment with a normal karyotype are presently unknown and require further study, including investigation of imprinting defects, subtelomeric abnormalities, and cryptic mosaicism. PMID- 12124733 TI - Unexpected retention and concomitant loss of subtelomeric regions in balanced chromosome anomalies by FISH. AB - Florescence in situ hybridization (FISH) using subtelomeric probes has been useful in detecting cryptic telomeric chromosomal rearrangements. We report, for the first time, that cytogenetically visible chromosome rearrangements can occur between the subtelomeric and telomeric region in clinically normal individuals with balanced chromosome anomalies in which one of the breakpoints involves a terminal band region. Using FISH with subtelomeric probes, we observed in three cases with a balanced reciprocal translocations the retention and subsequent loss of subtelomeric regions. In one case with a paracentric inversion, there was a proximal relocation of a subtelomeric region. Because subtelomeric regions serve important roles in chromosome pairing, this retention and concomitant loss or relocation of a subtelomeric region could possibly further disrupt the complex meiotic configurations of these balanced chromosome rearrangements. This may then have an effect on gamete production, placing these individuals at a higher risk for miscarriages and/or abnormal outcomes for individuals with similar chromosome aberrations. PMID- 12124735 TI - Choanal atresia: the result of maternal thyrotoxicosis or fetal carbimazole? AB - We present the fourth published case of a child affected with choanal atresia following maternal treatment with carbimazole. The mother was receiving her highest dose of carbimazole at the crucial period for development of the choanae, between days 35 and 38. PMID- 12124736 TI - Split-cord malformation in a girl with Angelman syndrome: a mere coincidence? AB - We present a case of a girl with both Angelman syndrome and split-cord malformation. The child was initially referred at the age of 2.5 years, for developmental delay and a possible diagnosis of spina bifida occulta, based on the presence of a hair tuft located on the midline of the lumbar area. Magnetic resonance imaging of the spine showed split-cord malformation below L1, whereas a cytogenetically detected deletion of chromosome bands 15q11-q13 (SNRPN) confirmed the clinical diagnosis of Angelman syndrome. Split-cord malformation or diastematomyelia is a rare form of spina bifida occulta that occurs sporadically and is not particularly related to specific syndromes. Hair patches or other distinctive cutaneous stigmata such as those seen in the present case have not, to our knowledge, been reported in other patients with Angelman syndrome; therefore, the association of Angelman syndrome and split-cord malformation in this child is probably coincidental. Spinal cord abnormalities have not been consistently reported in patients with Angelman syndrome; only one adult patient with Angelman syndrome and spina bifida occulta has been reported, and this association was probably considered fortuitous. However, some relatively uncommon clinical features such as deterioration of gait, lower limb malformations, and bladder dysfunction, particularly as the patients age, although nonspecific, are reminiscent of such a cause. We therefore urge clinicians to look for cutaneous stigmata along the spine and consider the evaluation of the spinal cord in children with apparent paraparesis, out of proportion to that usually seen in Angelman syndrome, should our case report not just be a coincidental observation. PMID- 12124738 TI - Severe lymphedema, intestinal lymphangiectasia, seizures and mild mental retardation: further case of Hennekam syndrome with a severe phenotype. AB - We report on an Italian patient affected by severe lymphedema of lower limbs, genitalia and face, intestinal lymphangiectasia, seizures, and moderate mental retardation. He has a flat face, flat nasal bridge, and hypertelorism. We propose that he presents with a severe form of Hennekam syndrome. PMID- 12124737 TI - Mosaic r(13) resulting in large deletion of chromosome 13q in a newborn female with multiple congenital anomalies. AB - A newborn female presented with multiple congenital anomalies including facial dysmorphism, agenesis of the corpus callosum, type I laryngeal cleft, tracheal stenosis, bilaterally small kidneys, segmental vertebral anomalies, extranumerary rib, bilateral hip dislocation, digital anomalies, and growth retardation. Newborn aneuploidy detection (NAD) based on interphase fluorescence in situ hybridization (FISH) indicated monosomy 13 in 47 of 200 (23.5%) peripheral blood cells (normal cutoff 8.5% at 95% CI). The follow-up banded metaphase-based analysis of 20 cells revealed a karyotype of 46,XX. The analysis of 30 additional cells revealed one cell to have monosomy 13 and a small ring chromosome. In the abnormal cell line, the ring was positive for whole chromosome paint (wcp) 13 and negative for Rb1 (13q14.3). The ring was detected in 4% of 80 additional metaphases studied by FISH. Therefore, the ring was present in 4% (5/130) of metaphases from peripheral blood. Analysis of buccal cells by FISH indicated the ring was present in 36% of cells. A higher degree of mosaicism (60%) was detected in fibroblast cultures from a skin biopsy. The low-level mosaicism of ring 13 in metaphase cells from peripheral blood would have been missed if the standard 20 GTL-banded metaphases had been analyzed. In this case, a preliminary interphase FISH study had indicated monosomy 13 resulting from a large 13q deletion that included the Rb1 locus. This finding initiated the analysis of additional metaphases by GTL-banding and the analysis of metaphases and interphases by FISH. The clinical presentation of our patient was consistent with reported cases of 13q deletions. In addition, our patient had airway anomalies, including a type I laryngeal cleft and tracheal stenosis, which are previously unreported. PMID- 12124739 TI - Trisomy 20q caused by der(4) t(4;20) (q35;q13.1): report of a new patient and review of the literature. AB - An infant with multiple congenital anomalies and severe developmental delay was found to have a derivative chromosome 4 by routine karyotypic analysis. Using telomeric FISH analysis, the source of the additional chromatin was determined to be from 20q. The infant, thus, is trisomic for 20q13.1 to 20qter and monosomic for the sub-telomeric region of 4q. Other cases of trisomy 20q13.1to 20qter associated with telomeric deletions are reviewed and compared to the current patient. PMID- 12124741 TI - Cockayne syndrome in three sisters with varying clinical presentation. AB - We report three sisters showing the clinical features and investigational findings of Cockayne syndrome (CS). In the rehabilitation unit of Northwest Armed Forces Hospital (N.W.A.F.H.), Tabuk, Saudi Arabia, there was a 12-year-old girl with typical features of CS. The girl had no apparent problems until the end of the first year when growth and developmental delay prompted medical evaluation. Brain CT, bone X-rays, auditory and ophthalmological evaluation confirmed the clinical impression of Cockayne syndrome. Two of her 13 sibs, both sisters, were later found to have the same syndrome. The sisters varied in clinical severity, as two of them had cataracts and early global delay and died early of inanition and infection. The third showed the disease manifestations at a relatively later age, did not have cataract, exhibited milder manifestations of the disease, and remains alive. The parents are not related by any way and the father is married to two other wives with 11 unaffected children. This report documents variable degrees of manifestations in sibs who presumably have the same gene mutation. PMID- 12124740 TI - Duplication of (2)(q11.1-q13.2) in a boy with mental retardation and cleft lip and palate: another clefting gene locus on proximal 2q? AB - A 4-year-old boy with left cleft lip and cleft palate, multiple minor anomalies and developmental delay revealed an abnormal chromosome 2 with enlarged proximal long arm, de novo, in his karyotype. Fluorescence in situ hybridization with a chromosome 2 library and band-specific YACs confined the duplicated segment to 2q11.1-q13.2 and indicated a direct tandem duplication due to unbalanced crossover between chromatids. PMID- 12124742 TI - Bilateral congenital diaphragmatic hernia: Differentiation between Pallister Killian and Fryns syndromes. PMID- 12124743 TI - Genotype-phenotype correlation in cystic fibrosis: the role of modifier genes. AB - More than 1,000 mutations have been identified in the cystic fibrosis (CF) transmembrane regulator (CFTR) disease gene. The impact of these mutations on the protein and the wide spectrum of CF phenotypes prompted a series of Genotype Phenotype correlation studies. The CFTR genotype is invariably correlated with pancreatic status-in about 85% of cases with pancreatic insufficiency and in about 15% of cases with pancreatic sufficiency. The correlations between the CFTR genotype and pulmonary, liver, and gastrointestinal expression are debatable. The heterogeneous phenotype in CF patients bearing the same genotype or homozygotes for nonsense mutations implicated environmental and/or genetic factors in the disease. However, the discordant phenotype observed in CF siblings argued against a major role of environmental factors and suggested that genes other than CFTR modulate the CF phenotype. A locus that modulates gastrointestinal expression was identified in mice and subsequently in humans. By analyzing nine CF patients discordant for meconium ileus we were able to show that this locus had a dominant effect. Moreover, in a collaborative study we found a higher rate of polymorphisms in beta-defensin genes 1 and 2 in CF patients and in controls. In another multicenter study mutations in alpha-1 antitrypsin (A1AT) and mannose binding lectin genes were found to be independent risk factors for liver disease in CF patients. The body of evidence available suggests that the variegated CF phenotype results from complex interactions between numerous gene products. PMID- 12124744 TI - Cancer genetics. AB - Cancer is a genetic disease of somatic cells. Tumor karyotypes are rarely normal, and most show multiple abnormalities of both number and structure. The first direct evidence for this concept of cancer came from studies of tumor-specific translocations in leukemias and lymphomas, revealing the importance of oncogenes and the regulation of gene transcription in cancer. A second major source of information about human cancer genes is hereditary cancer. Genetic predisposition of the autosomal dominant type imposes a high relative risk for one or more kinds of cancer. In the past decade or so, more than 30 mutant genes for such hereditary cancers have been cloned. Penetrance depends upon additional, somatic, mutations. A few of the genes are oncogenes or DNA repair genes, but most are tumor suppressor genes. Some tumor suppressors regulate transcription, while others operate in signal transduction pathways that are involved in regulating processes of cell birth, differentiation, and death. The knowledge gained is stimulating new approaches to the treatment and prevention of cancer. PMID- 12124745 TI - Abnormal spliceform expression associated with splice acceptor mutations in exon IIIc of FGFR2. PMID- 12124746 TI - A not-so-"new" mental retardation syndrome. PMID- 12124748 TI - Development of depolarization-induced calcium transients in insect glial cells is dependent on the presence of afferent axons. AB - Changes in the intracellular Ca(2+) concentration ([Ca(2+)](i)) induced by depolarization have been measured in glial cells acutely isolated from antennal lobes of the moth Manduca sexta at different postembryonic developmental stages. Depolarization of the glial cell membrane was elicited by increasing the external K(+) concentration from 4 to 25 mM. At midstage 5 and earlier stages, less than 20% of the cells responded to 25 mM K(+) (1 min) with a transient increase in [Ca(2+)](i) of approximately 40 nM. One day later, at late stage 5, 68% of the cells responded to 25 mM K(+), the amplitude of the [Ca(2+)](i) transients averaging 592 nM. At later stages, all cells responded to 25 mM K(+) with [Ca(2+)](i) transients with amplitudes not significantly different from those at late stage 5. In stage 6 glial cells isolated from deafferented antennal lobes, i.e., from antennal lobes chronically deprived of olfactory receptor axons, only 30% of the cells responded with [Ca(2+)](i) transients. The amplitudes of these [Ca(2+)](i) transients averaged 93 nM and were significantly smaller than those in normal stage 6 glial cells. [Ca(2+)](i) transients were greatly reduced in Ca(2+)-free, EGTA-buffered saline, and in the presence of the Ca(2+) channel blockers cadmium and verapamil. The results suggest that depolarization of the cell membrane induces Ca(2+) influx through voltage-activated Ca(2+) channels into antennal lobe glial cells. The development of the depolarization-induced Ca(2+) transients is rapid between midstage 5 and stage 6, and depends on the presence of afferent axons from the olfactory receptor cells in the antenna. PMID- 12124749 TI - Remodeling of an identified motoneuron during metamorphosis: central and peripheral actions of ecdysteroids during regression of dendrites and motor terminals. AB - During metamorphosis of the moth Manduca sexta, an identified leg motoneuron, the femoral depressor motoneuron (FeDe MN), undergoes reorganization of its central and peripheral processes. This remodeling is under the control of two insect hormones: the ecdysteroids and juvenile hormone (JH). Here, we asked whether peripheral or central actions of the ecdysteroids influenced specific regressive aspects of MN remodeling. We used stable hormonal mimics to manipulate the hormonal environment of either the FeDe muscle or the FeDe MN soma. Our results demonstrate that motor-terminal retraction and dendritic regression can be experimentally uncoupled, indicating that central actions of ecdysteroids trigger dendritic regression whereas peripheral actions trigger terminal retraction. Our results further demonstrate that discrete aspects of motor-terminal retraction can also be experimentally uncoupled, suggesting that they also are regulated differently. PMID- 12124750 TI - A new look at an old visual system: structure and development of the compound eyes and optic ganglia of the brine shrimp Artemia salina Linnaeus, 1758 (Branchiopoda, anostraca). AB - Compared to research carried out on decapod crustaceans, the development of the visual system in representatives of the entomostracan crustaceans is poorly understood. However, the structural evolution of the arthropod visual system is an important topic in the new debate on arthropod relationships, and entomostracan crustaceans play a key role in this discussion. Hence, data on structure and ontogeny of the entomostracan visual system are likely to contribute new aspects to our understanding of arthropod phylogeny. Therefore, we explored the proliferation of neuronal stem cells (in vivo incorporation of bromodeoxyuridine) and the developmental expression of synaptic proteins (immunohistochemistry against synapsins) in the developing optic neuropils of the brine shrimp Artemia salina Linnaeus, 1758 (Crustacea, Entomostraca, Branchiopoda, Anostraca) from hatching to adulthood. The morphology of the adult visual system was examined in serial sections of plastic embedded specimens. Our results indicate that the cellular material that gives rise to the visual system (compound eyes and two optic ganglia) is contributed by the mitotic activity of neuronal stem cells that are arranged in three band-shaped proliferation zones. Synapsin-like immunoreactivity in the lamina ganglionaris and the medulla externa initiated only after the anlagen of the compound eyes had already formed, suggesting that the emergence of the two optic neuropils lags behind the proliferative action of these stem cells. Neurogenesis in A. salina is compared to similar processes in malacostracan crustaceans and possible phylogenetic implications are discussed. PMID- 12124751 TI - GABAergic modulation of primary gustatory afferent synaptic efficacy. AB - Modulation of synaptic transmission at the primary sensory afferent synapse is well documented for the somatosensory and olfactory systems. The present study was undertaken to test whether GABA impacts on transmission of gustatory information at the primary afferent synapse. In goldfish, the vagal gustatory input terminates in a laminated structure, the vagal lobes, whose sensory layers are homologous to the mammalian nucleus of the solitary tract. We relied on immunoreactivity for the GABA-transporter, GAT-1, to determine the distribution of GABAergic synapses in the vagal lobe. Immunocytochemistry showed dense, punctate GAT-1 immunoreactivity coincident with the layers of termination of primary afferent fibers. The laminar nature and polarized dendritic structure of the vagal lobe make it amenable to an in vitro slice preparation to study early synaptic events in the transmission of gustatory input. Electrical stimulation of the gustatory nerves in vitro produces synaptic field potentials (fEPSPs) predominantly mediated by ionotropic glutamate receptors. Bath application of either the GABA(A) receptor agonist muscimol or the GABA(B) receptor agonist baclofen caused a nearly complete suppression of the primary fEPSP. Coapplication of the appropriate GABA(A) or GABA(B) receptor antagonist bicuculline or CGP 55845 significantly reversed the effects of the agonists. These data indicate that GABAergic terminals situated in proximity to primary gustatory afferent terminals can modulate primary afferent input via both GABA(A) and GABA(B) receptors. The mechanism of action of GABA(B) receptors suggests a presynaptic locus of action for that receptor. PMID- 12124752 TI - Synaptic inputs onto spiking local interneurons in crayfish are depressed by nitric oxide. AB - We have analyzed the action of nitric oxide on the synaptic inputs of spiking local interneurons that form part of the local circuits in the terminal abdominal ganglion of the crayfish, Pacifastacus leniusculus. Increasing the availability of NO in the ganglion by bath applying the NO donor SNAP, or the substrate for its synthesis, L-arginine, caused a depression of synaptic inputs onto the interneurons evoked by electrically stimulating mechanosensory neurons in nerve 2 of the terminal ganglion. Conversely, reducing the availability of NO by bath application of an NO scavenger, PTIO, and an inhibitor of nitric oxide synthase, L-NAME, increased the amplitude of the evoked potentials. These results suggest that elevated NO concentration causes a depression of the synaptic inputs to spiking local interneurons. To determine whether these effects could be mediated through an NO/cGMP signaling pathway we bath applied a membrane permeable analogue of cGMP, 8-br-cGMP, which decreased the amplitude of the inputs to the interneurons. Bath application of an inhibitor of soluble guanlylyl cyclase, ODQ, produced an increase in the amplitude of the synaptic inputs. Our results suggest that NO causes a depression of synaptic inputs to spiking local interneurons probably by acting through an NO/cGMP signaling pathway. Moreover, application of NO scavengers modulates the inputs to these interneurons, suggesting that NO is continuously providing a powerful and dynamic means of modulating the outputs of local circuits. PMID- 12124753 TI - Role of Nova-1 in regulating alpha2N, a novel glycine receptor splice variant, in developing spinal cord neurons. AB - Inhibitory glycine receptor (GlyR) subunits undergo developmental regulation, but the molecular mechanisms of GlyR regulation in developing neurons are little understood. Using RT-PCR, we investigated the regulation of GlyR alpha-subunit splice forms during the development of the spinal cord of the rat. Experiments to compare the amounts of mRNA for two known splice variants of the GlyR alpha2 subunit, alpha2A and alpha2B, in the developing rat spinal cord revealed the presence of an additional, novel variant that lacked any exon 3, herein named "alpha2N." Examination of the RNA from spinal cords of different-aged rats showed a dramatic down-regulation of alpha2N during prenatal development: alpha2N mRNA formed a significant portion of the alpha2 subunit pool at E14, but its relative level was reduced by 85% by birth and was undetectable in adults. Two proteins previously implicated in regulating the splicing of GlyR alpha2 pre-mRNA, the neurooncological ventral antigen-1 (Nova-1) and the brain isoform of the polypyrimidine tract binding protein (brPTB), underwent small changes over the same period that did not correlate directly with the changes in the level of alpha2N, calling into question their involvement in the developmental regulation of alpha2N. However, treatment of spinal cord neurons in culture with antisense oligonucleotides designed selectively to knock down one of three Nova-1 variants significantly altered the relative level of GlyR alpha2N, showing that Nova-1 isoforms can regulate GlyR alpha2 pre-mRNA splicing in developing neurons. These results provide evidence for a novel splice variant of the GlyR alpha2 subunit that undergoes dramatic developmental regulation, reveal the expression profiles of Nova-1 and brPTB in the developing spinal cord, and suggest that Nova-1 plays a role in regulating GlyR alpha2N in developing neurons. PMID- 12124754 TI - Cloning of the cDNA and mRNA expression of CLRP, a complex leucine repeat protein of the Golgi apparatus expressed by specific neurons of the rat brain. AB - We report the molecular cloning of one novel cDNA isolated from the rat brain. We have named the putative protein CLRP, for complex leucine-repeat protein. The predicted CLRP amino acid sequence shares homology in the amino acid composition with the Galactose, N-Acetylglucosamine, and Sialic acid transporters, and shows 91% identity with the sequence of one human chromosome 5 BAC clone. Expression of the CLRP cDNA tagged with GFP in COS-7 cells was found in cell organelles that resemble the Golgi apparatus of the cytoplasm. In Northern blot, the CLRP probe labels a single band of 2.4 kb in the brain, kidney, lung, testis, and prostate. In the brain, CLRP mRNA is expressed by limited sets of neurons, such as the pyramidal cells of the cortex, the Purkinje cells of the cerebellum, and the motoneurons of the brainstem. In the brain, the CLRP mRNA is expressed at embryonic day 15; levels of expression are maintained until postnatal day 10 and decrease in adults. The results suggest that CLRP codes a novel member of the nucleotide-sugar family of proteins of the Golgi apparatus. PMID- 12124755 TI - Distribution of cholecystokinin, calcitonin gene-related peptide, neuropeptide Y, and galanin in the primary gustatory nuclei of the goldfish. AB - Cholecystokinin (CCK), neuropeptide Y (NPY), calcitonin gene-related peptide (CGRP), and galanin all are known to have central effects on food intake. Immunocytochemistry was used to examine the presence of these substances within the primary gustatory nuclei of the goldfish, including the vagal lobe, which is a large, laminated structure composed of discrete sensory, fiber, and motor layers. The vagal lobes receive primary afferent input from the gustatory portion of the vagus nerve and contain reflex circuitry involved in the ingestion or rejection of potential food items. Immunohistochemistry indicates a heavy concentration of CCK-, CGRP-, NPY-, and galanin-immunoreactive fibers in the capsular fiber layer as well as in deeper sensory layers of the vagal lobe. CGRP immunoreactivity throughout the sensory layers and capsular immunoreactivity for CCK are greatly reduced 1-2 weeks following vagus nerve transection, indicating that the majority of these fibers are primary sensory afferents. In contrast, NPY and galanin immunoreactivity in the capsular fiber layer and reactivity for CCK, NPY, and galanin in the deeper sensory and fiber layers are relatively unaffected by vagus transection. CCK-, NPY-, and galanin-immunoreactive fibers and puncta also were present in the motor layers, as were CGRP-immunoreactive motor somata. CCK-immunoreactive cell bodies are present in layer III and layer VII/VIII of the vagal lobe and in the superficial granular layer of the lateral subnucleus of the commissural nucleus of Cajal, which is caudally contiguous with the vagal lobe. PMID- 12124756 TI - Ontogenetic organization of the FMRFamide immunoreactivity in the nervus terminalis of the lungfish, Neoceratodus forsteri. AB - The development of the nervus terminalis system in the lungfish, Neoceratodus forsteri, was investigated by using FMRFamide as a marker. FMRFamide immunoreactivity appears first within the brain, in the dorsal hypothalamus at a stage around hatching. At a slightly later stage, immunoreactivity appears in the olfactory mucosa. These immunoreactive cells move outside the olfactory organ to form the ganglion of the nervus terminalis. Immunoreactive processes emerge from the ganglion of the nervus terminalis in two directions, one which joins the olfactory nerve to travel to the brain and the other which courses below the brain to enter at the level of the preoptic nucleus. Neither the ganglion of the nervus terminalis nor the two branches of the nervus terminalis form after surgical removal of the olfactory placode at a stage before the development of FMRFamide immunoreactivity external to the brain. Because this study has confirmed that the nervus terminalis in lungfish comprises both an anterior and a posterior branch, it forms the basis for discussion of homology between these branches and the nervus terminalis of other anamniote vertebrates. PMID- 12124757 TI - Three-dimensional cartography of functional territories in the human striatopallidal complex by using calbindin immunoreactivity. AB - This anatomic study presents an analysis of the distribution of calbindin immunohistochemistry in the human striatopallidal complex. Entire brains were sectioned perpendicularly to the mid-commissural line into 70-microm-thick sections. Every tenth section was immunostained for calbindin. Calbindin labeling exhibited a gradient on the basis of which three different regions were defined: poorly labeled, strongly labeled, and intermediate. Corresponding contours were traced in individual sections and reformatted as three-dimensional structures. The poorly labeled region corresponded to the dorsal part of the striatum and to the central part of the pallidum. The strongly labeled region included the ventral part of the striatum, the subcommissural part of the external pallidum but also the adjacent portion of its suscommissural part, and the anterior pole of the internal pallidum. The intermediate region was located between the poorly and strongly labeled regions. As axonal tracing and immunohistochemical studies in monkeys show a similar pattern, poorly, intermediate, and strongly labeled regions were considered as the sensorimotor, associative, and limbic territories of the human striatopallidal complex, respectively. However, the boundaries between these territories were not sharp but formed gradients of labeling, which suggests overlapping between adjacent territories. Similarly, the ventral boundary of the striatopallidal complex was blurred, suggesting a structural intermingling with the substantia innominata. This three-dimensional partitioning of the human striatopallidal complex could help to define functional targets for high-frequency stimulation with greater accuracy and help to identify new stimulation sites. PMID- 12124758 TI - Antagonism of vasoactive intestinal peptide mRNA in the suprachiasmatic nucleus disrupts the rhythm of FRAs expression in neuroendocrine dopaminergic neurons. AB - This study was designed to determine whether there is a functional relationship between cfos expression in vasoactive intestinal peptide (VIP) -containing neurons of the suprachiasmatic nucleus (SCN) and Fos-related antigens (FRAs) expression in neuroendocrine dopaminergic neurons of the arcuate (ARN) and periventricular (PeVN) nuclei of the hypothalamus. Brains were obtained from ovariectomized (OVX) female rats killed at 12:00 AM, 7:00 AM, 9:00 AM, 12:00 PM, and 7:00 PM (12 hours illumination beginning 6:00 AM). Antibodies against FRAs and tyrosine hydroxylase (TH) identified activated neuroendocrine dopaminergic neurons. Antibodies against cfos and VIP identified activated VIP-immunoreactive (IR) neurons in the SCN. The proportion of neuroendocrine dopaminergic neurons in the ARN and PeVN expressing FRAs was greatest and equivalent at 7:00 AM, 9:00 AM, 12:00 PM, and 12:00 AM. At 7:00 PM, the proportion of neuroendocrine dopaminergic neurons expressing FRAs was significantly lower than all other time points. In the SCN, a subpopulation of VIP-IR neurons maximally expressed cfos at 7:00 AM, which decreased through 9:00 AM. cFos was not expressed at 7:00 PM and 12:00 AM in VIP-IR neurons. Antisense VIP oligonucleotides were injected into the SCN to determine whether attenuation of VIP expression disturbs rhythms in neuroendocrine dopaminergic neuronal activity. OVX rats were infused with either antisense VIP oligonucleotides or scrambled sequence oligonucleotides bilaterally (0.5 microg in 0.5 microl of saline per side) in the SCN. Animals were killed 34 hours (7:00 PM) and 46 hours (7:00 AM) after receiving infusions, and brains were recovered. Administration of antisense VIP oligonucleotides decreased VIP protein expression in the SCN and prevented the decrease in the percentage of neuroendocrine dopaminergic neurons expressing FRAs at 7:00 PM but did not affect FRAs expression at 7:00 AM when compared with animals receiving scrambled oligonucleotides. These data suggest that VIP fibers from the SCN may relay time of-day information to neuroendocrine dopaminergic neurons to inhibit their activity and, thus, initiate prolactin release in the evening. PMID- 12124759 TI - Normal dendritic arborization in spinal motoneurons requires neurofilament subunit L. AB - Neurofilaments, composed of three polypeptide subunits, NF-L, NF-M, and NF-H, are major cytoskeletal elements in large neurons with long axons. Neurofilaments play a critical role in the development of axonal diameter; however, their role in the development of dendrites is largely unknown. By overexpressing different neurofilament subunits, we previously demonstrated that alteration of neurofilament subunit composition resulted in dramatic changes in dendritic arborization. To further determine the role of neurofilaments in dendritic growth, we examined and compared the dendritic architecture of spinal cord neurons in young NF-L knockout (-/-), heterozygous (+/-), and wild-type (+/+) mice. We show that an absence or reduction in the expression of NF-L inhibited dendritic growth most dramatically in large motoneurons, mildly in medium neurons, but had no effect on small neurons. We also reveal that a decrease in NF L leads to an increase in NF-M and NF-H subunits in cell bodies and their reduction in dendrites. These results demonstrate that NF-L is a critical intrinsic factor for dendritic growth in large motoneurons. PMID- 12124760 TI - Morphometric analysis of dendritic remodeling in an identified motoneuron during postembryonic development. AB - A detailed quantitative description of modifications in neuronal architecture is an important prerequisite to investigate the signals underlying behaviorally relevant changes in neuronal shape. Extensive morphological remodeling of neurons occurs during the metamorphosis of holometabolous insects, such as Manduca sexta, in which new adult behaviors develop postembryonically. In this study, a morphometric analysis of the structural changes of an identified Manduca motoneuron, MN5, was conducted by sampling its metric parameters at different developmental stages. The remodeling of MN5 is divided into three main phases. The regression of most larval dendrites (1) is followed by the formation of dendritic growth-cones (2), and subsequently, adult dendrite formation (3). In contrast, the cell body and link segment surface increase during dendritic regression and regrowth, indicating that different cell compartments receive different signals, or respond differently to the same signal. During dendritic growth-cone formation, the growth of the cell body and the link segment are arrested. Sholl and branch frequency analysis suggest two different modes of dendritic growth. During a first growth-cone-dependent phase, new branch formation occurs at all dendrites. The maximum path length of the major dendritic tree changes little, whereas branch order increases from 20 to 45. Changes in total dendritic length are correlated with strong changes in the number of nodes but with minor changes in the average dendritic segment length, indicating a mode of growth similar to that induced by steroid hormone application to cultured motoneurons. The second phase is growth-cone-independent, and branching is limited to high order dendrites. PMID- 12124761 TI - Ultrastructural organization of lamprey reticulospinal synapses in three dimensions. AB - The giant reticulospinal synapse in lamprey provides a unique model to study synaptic vesicle traffic. The axon permits microinjections, and the active zones are often separated from each other, which makes it possible to track vesicle cycling at individual release sites. However, the proportion of reticulospinal synapses with individual active zones ("simple synapses") is unknown and a quantitative description of their organization is lacking. Here, we report such data obtained by serial section analysis, intermediate-voltage electron microscopy, and electron tomography. The simple synapse was the most common type (78%). It consisted of one active zone contacting one dendritic process. The remaining synapses were "complex," mostly containing one vesicle cluster and two to three active zones synapsing with distinct dendritic shafts. Occasional axosomatic synapses with multiple active zones forming synapses with the same cell were also observed. The vast majority of active zones in all synapse types contained both chemical and electrotonic synaptic specializations. Quantitative analysis of simple synapses showed that the majority had active zones with a diameter of 0.8-1.8 microm. The number of synaptic vesicles and the height of the vesicle cluster in middle sections of serially cut synapses correlated with the active zone length within, but not above, this size range. Electron tomography of simple synapses revealed small filaments between the clustered synaptic vesicles. A single vesicle could be in contact with up to 12 filaments. Another type of filament, also associated with synaptic vesicles, emerged from dense projections. Up to six filaments could be traced from one dense projection. PMID- 12124762 TI - Premotor circuits controlling eyelid movements in conjunction with vertical saccades in the cat: I. The rostral interstitial nucleus of the medial longitudinal fasciculus. AB - Saccadic eye movements in the vertical plane are controlled by the rostral interstitial nucleus of the medial longitudinal fasciculus (riMLF) and the interstitial nucleus of Cajal. Eye movements in the vertical direction are accompanied by concurrent upper eye lid movements. These gaze-related lid movements are produced by the levator palpebrae superioris muscle, whose motoneurons are located in the caudal central subdivision (CCS) of the oculomotor nucleus. The neural circuits that direct such gaze-related lid movements were examined by use of both conventional and dual neuronal tracing methods in the cat. Injections of wheat germ agglutinin-conjugated horseradish peroxidase (WGA HRP) into the area of the CCS revealed a distinctive subset of retrogradely labeled neurons located in the caudomedial portion of the riMLF. This subset of riMLF neurons was not labeled when injections were localized within the oculomotor nucleus proper, without involving the CCS. Injections of biotinylated dextran amine (BDA) that included this caudomedial riMLF region anterogradely labeled axons that projected profusely throughout the CCS. Labeled terminals were seen in close association with retrogradely labeled levator palpebrae motoneurons, which were primarily found contralateral to WGA-HRP muscle injections. Ultrastructural examination revealed that most BDA-labeled terminals contained clear spherical vesicles and formed asymmetrical synaptic contacts, primarily on the proximal dendrites of WGA-HRP-labeled motoneurons. A few had pleiomorphic vesicles. In summary, these results strongly suggest that the caudomedial part of the cat riMLF is a premotor center that monosynaptically controls lid movements in conjunction with vertical saccades. PMID- 12124763 TI - Common design of mushroom bodies in bees and flies? PMID- 12124764 TI - Organization of the honey bee mushroom body: representation of the calyx within the vertical and gamma lobes. AB - Studies of the mushroom bodies of Drosophila melanogaster have suggested that their gamma lobes specifically support short-term memory, whereas their vertical lobes are essential for long-term memory. Developmental studies have demonstrated that the Drosophila gamma lobe, like its equivalent in the cockroach Periplaneta americana, is supplied by a special class of intrinsic neuron-the clawed Kenyon cells-that are the first to differentiate during early development. To date, however, no account identifies a corresponding lobe in the honey bee, another taxon used extensively for learning and memory research. Received opinion is that, in this taxon, each of the mushroom body lobes comprises three parallel divisions representing one of three concentric zones of the calyces, called the lip, collar, and basal ring. The present account shows that, although these zones are represented in the lobes, they occupy only two thirds of the vertical lobe. Its lowermost third receives the axons of the clawed class II Kenyon cells, which are the first to differentiate during early development and which represent the whole calyx. This component of the lobe is anatomically and developmentally equivalent to the gamma lobe of Drosophila and has been here named the gamma lobe of the honey bee. A new class of intrinsic neurons, originating from perikarya distant from the mushroom body, provides a second system of parallel fibers from the calyx to the gamma lobe. A region immediately beneath the calyces, called the neck, is invaded by these neurons as well as by a third class of intrinsic cell that provides connections within the neck of the pedunculus and the basal ring of the calyces. In the honey bee, the gamma lobe is extensively supplied by afferents from the protocerebrum and gives rise to a distinctive class of efferent neurons. The terminals of these efferents target protocerebral neuropils that are distinct from those receiving efferents from divisions of the vertical lobe that represent the lip, collar, and basal ring. The identification of a gamma lobe unites the mushroom bodies of evolutionarily divergent taxa. The present findings suggest the need for critical reinterpretation of studies that have been predicated on early descriptions of the mushroom body's lobes. PMID- 12124766 TI - Distribution of thyrotropin-releasing hormone (TRH) immunoreactivity in the brain of the zebrafish (Danio rerio). AB - The distribution of thyrotropin-releasing hormone (TRH) in the brain of the adult zebrafish was studied with immunohistochemical techniques. In the telencephalon, abundant TRH-immunoreactive (TRHir) neurons were observed in the central, ventral, and supra- and postcommissural regions of the ventral telencephalic area. In the diencephalon, TRHir neurons were observed in the anterior parvocellular preoptic nucleus, the suprachiasmatic nucleus, the lateral hypothalamic nucleus, the rostral parts of the anterior tuberal nucleus and torus lateralis, and the posterior tuberal nucleus. Some TRHir neurons were also observed in the central posterior thalamic nucleus and in the habenula. The mesencephalon contained TRHir cells in the rostrodorsal tegmentum, the Edinger Westphal nucleus, the torus semicircularis, and the nucleus of the lateral lemniscus. Further TRHir neurons were observed in the interpeduncular nucleus. In the rhombencephalon, TRHir cells were observed in the nucleus isthmi and the locus coeruleus, rostrally, and in the vagal lobe and vagal motor nucleus, caudally. In the forebrain, TRHir fibers were abundant in several regions, including the medial and caudodorsal parts of the dorsal telencephalic area, the ventral and commissural parts of the ventral telencephalic area, the preoptic area, the posterior tubercle, the anterior tuberal nucleus, and the posterior hypothalamic lobe. The dorsal thalamus exhibited moderate TRHir innervation. In the mesencephalon, the optic tectum received a rich TRHir innervation between the periventricular gray zone and the stratum griseum centrale. A conspicuous TRHir longitudinal tract traversed the tegmentum and extended to the rhombencephalon. The medial and lateral mesencephalic reticular areas and the interpeduncular nucleus were richly innervated by TRHir fibers. In the rhombencephalon, the secondary gustatory nucleus received abundant TRHir fibers. TRHir fibers moderately innervated the ventrolateral and ventromedial reticular area and richly innervated the vagal lobe and Cajal's commissural nucleus. Some TRHir fibers coursed in the lateral funiculus of the spinal cord. Some TRHir amacrine cells were observed in the retina. The wide distribution of TRHir neurons and fibers observed in the zebrafish brain suggests that TRH plays different roles. These results in the adult zebrafish reveal a number of differences with respect to the TRHir systems reported in other adult teleosts but were similar to those found during late developmental stages of trout (Diaz et al., 2001). PMID- 12124765 TI - Immunohistochemical localization and biochemical characterization of ghrelin in the brain and stomach of the frog Rana esculenta. AB - Ghrelin is a 28-amino acid n-octanoylated peptide recently isolated from the rat stomach as an endogenous ligand of the growth hormone secretagogue receptor. So far, the occurrence of ghrelin has not been investigated in submammalian vertebrates. In the present work, we have studied the anatomic distribution and biochemical characterization of ghrelin-like immunoreactivity in the brain and stomach of the frog Rana esculenta by using two distinct antisera directed against rat ghrelin. In the brain, sparse ghrelin-positive cells were detected in three nuclei of the diencephalon, namely the suprachiasmatic nucleus and the posterior tuberculum in the hypothalamus, and the posterodorsal aspect of the lateral nucleus in the thalamus. A few ghrelin-immunoreactive neurons were also found in the mesencephalon, i.e., in the pretoral gray and the anterodorsal tegmental nucleus. Ghrelin-containing fibers were widely distributed in the frog brain. In particular, diffuse networks of immunoreactive processes were observed in various regions of the telencephalon, including the medial pallium, the striatum, the nucleus of the diagonal band of Broca, the nucleus accumbens, and the amygdala. In the diencephalon, the magnocellular nucleus, the suprachiasmatic nucleus, the posterior tuberculum, and the ventrolateral and lateral thalamic nuclei were moderately to densely innervated with ghrelin-containing fibers. A moderate density of positive fibers was also found in different areas of the mesencephalon such as the nucleus of the medial longitudinal fasciculus, the pretoral gray, and the tegmentum. In the stomach, a few brightly immunofluorescent cells were detected in the mucosa. The distribution pattern and morphologic characteristics of ghrelin-containing cells in the stomach suggest that they correspond to endocrine cells. Reversed-phase high performance liquid chromatography analysis of frog brain and stomach extracts, combined with RIA detection, revealed that ghrelin-immunoreactive material eluted as a single peak with a retention time slightly shorter than that of synthetic rat ghrelin. The present data provide the first evidence that a ghrelin-related peptide is present in submammalian vertebrates. The occurrence of ghrelin-containing cells in the hypothalamus and the stomach mucosa suggests that, in amphibians, ghrelin may exert both neuroendocrine and endocrine activities. PMID- 12124767 TI - Remodeling of the proximal segment of crayfish motor nerves following transection. AB - Transected crustacean motor axons consist of a soma-endowed proximal segment that regenerates and a soma-less distal segment that survives for up to a year. We report on the anatomical remodeling of the proximal segment of phasic motor nerves innervating the deep flexor muscles in the abdomen of adult crayfish following transection. The intact nerve with 10 phasic axons and its two branches with subsets of 6 and 7 of these 10 axons undergo several remodeling changes. First, the transected nerve displays many more and smaller axon profiles than the 6 and 7 axons of the intact nerve, approximately 100 and 300 profiles in the two branches of a preparation transected 8 weeks previously. Serial images of the transected nerve denote that the proliferation of profiles is due to several orders of axon sprouting primary, secondary, and tertiary branches. The greater proliferation of axon sprouts, their smaller size, and the absence of intervening glia in the one nerve branch compared with the other branch denote that sprouting is more advanced in this branch. Second, the axon sprouts are regionally differentiated; thus, although in most regions the sprouts are basically axon like, with a cytoskeleton of microtubules and peripheral mitochondria, in some regions they appear nerve terminal-like and are characterized by numerous clear synaptic vesicles, a few dense-core vesicles, and dispersed mitochondria. Both regions possess active zone dense bars with clustered synaptic vesicles found opposite other sprouts, glia, hemocytes, and connective tissue, but because the opposing membranes are not differentiated into a synaptic contact, the active zones are extrasynaptic. Third, some of the transected axons display a glial cell nucleus denoting assimilation of an adaxonal glial cell by the transected axons. Fourth, within the nerve trunk are a few myocytes and muscle fibers. These most likely originate from adjoining and intimately connected hemocytes, because such transformation occurs during muscle repair. In a crustacean nerve, however, where muscle is clearly misplaced, its presence implies an instructive role for motor nerves in muscle formation. PMID- 12124768 TI - Conserved and divergent patterns of Reelin expression in the zebrafish central nervous system. AB - The protein Reelin is suggested to function in cell-cell interactions and in mediating neuronal migrations in layered central nervous system structures. With the aim of shedding light on the development of the teleost telencephalon, which forms through the process of eversion and results in the formation of a nonlaminar pallium, we isolated a zebrafish ortholog of the reelin gene and studied its expression in developing and adult brain. The pattern of expression is highly dynamic during the first 24-72 hours of development. By 5 days postfertilization, high amounts of reelin mRNA are found in the dorsal telencephalon, thalamic and hypothalamic regions, pretectal nuclei, optic tectum, cerebellum, hindbrain, reticular formation, and spinal cord, primarily confined to postmitotic neurons. This pattern persists in 1- to 3-month-old zebrafish. This study, together with reports on reelin expression in other vertebrates, shows that reelin mRNA distribution is conserved in many regions of the vertebrate brain. A major exception is that reelin is expressed in the majority of the cells of the dorsal regions of the everted telencephalon in zebrafish embryos, whereas it is restricted to specific neuronal populations in the developing telencephalon of amniotes. To better understand the origin of these differences, we analyzed reelin expression in the telencephalon of an amphibian. Telencephalic reelin expression in Xenopus laevis shows more similarities with the sauropsidian than with the teleostean pattern. Thus, the differences in the telencephalic expression of reelin between teleosts and tetrapods are likely to be due to different roles for Reelin during eversion, a process that is specific for the teleost telencephalon. PMID- 12124769 TI - Elongation of hair cell stereocilia is defective in the mouse mutant whirler. AB - The recessive mouse mutant whirler (wi) shows no response to sound and exhibits circling and head-tossing behaviour, indicative of both auditory and vestibular dysfunction. The wi mutation maps genetically to mouse chromosome 4. We examined the organ of Corti of whirler mutants to explore the possibility that the wi mutation affects sensory hair cells. Scanning electron microscopy (SEM) reveals that the specialised microvilli (stereocilia) that are projected by the sensory hair cells and are vital for sound transduction are abnormal in wi homozygotes. Specifically, wi homozygous inner hair cell (IHC) stereocilia are approximately half the length of equivalent stereocilia in heterozygous littermates. They are arranged normally into ranks, but the gradation in height and width of stereocilia in adjacent ranks is less prominent in wi homozygotes. Analysis of IHC stereocilia during the course of their development shows that, by embryonic day 18.5, mutant stereocilia are already significantly shorter than those in controls. Mutant stereocilia elongate at a normal rate, at least until postnatal day 1, but prematurely stop elongating between postnatal days 1 and 4. Stereocilia length then decreases. At postnatal day 15, outer hair cell (OHC) stereocilia in wi homozygotes appear short and are arranged in a rounded, "U" shape rather than the normal "W" or "V" shape. Eventually, both IHCs and OHCs degenerate. We show that the whirler locus encodes a protein(s) required for the elongation and maintenance of IHC and OHC stereocilia. PMID- 12124770 TI - Physiological functions of imprinted genes. AB - Genomic imprinting in gametogenesis marks a subset of mammalian genes for parent of-origin-dependent monoallelic expression in the offspring. Embryological and classical genetic experiments in mice that uncovered the existence of genomic imprinting nearly two decades ago produced abnormalities of growth or behavior, without severe developmental malformations. Since then, the identification and manipulation of individual imprinted genes has continued to suggest that the diverse products of these genes are largely devoted to controlling pre- and post natal growth, as well as brain function and behavior. Here, we review this evidence, and link our discussion to a website (http://www.otago.ac.nz/IGC) containing a comprehensive database of imprinted genes. Ultimately, these data will answer the long-debated question of whether there is a coherent biological rationale for imprinting. PMID- 12124771 TI - New insights into the role of extracellular matrix during tumor onset and progression. AB - Recently, a view of the tumor as a functional tissue interconnected with the microenvironment has recently been described. For many years, the stroma has been studied in the context of the malignant lesion, and only rarely has its role been considered before carcinogenic lesions appear. Recent studies have provided evidence that stromal cells and their products can cause the transformation of adjacent cells through transient signaling that leads to the disruption of homeostatic regulation, including control of tissue architecture, adhesion, cell death, and proliferation. It is now well established that tumor progression requires a continually evolving network of interactions between neoplastic cells and extracellular matrix. A relevant step of this process is the remodeling of microenvironment which surrounds tumors leading to the release of ECM-associated growth factors which can then stimulate tumor and/or endothelial cells. Finally, tumor cells reorganizing the extracellular matrix to facilitate communications and escape the homeostatic control exerted by the microenvironment modify response to cytotoxic treatments. PMID- 12124773 TI - Structure-activity relationships of heparin-mimicking compounds in induction of bFGF release from extracellular matrix and inhibition of smooth muscle cell proliferation and heparanase activity. AB - A series of nine synthetic polyaromatic compounds were synthesized by polymerization of aromatic ring monomers with formaldehyde, which yield substantially ordered backbones with different functional anionic groups (hydroxyl and carboxyl) on the phenol ring. These compounds were tested for their heparin-mimicking activity: (1) inhibition of heparanase activity; (2) inhibition of SMC proliferation; and (3) release of bFGF from the ECM. We demonstrate that compounds that have two hydroxyl groups para and ortho to the carboxylic group and a carboxylic group at a distance of two carbons from the phenol ring inhibit heparanase activity and SMC proliferation, as well as induced an almost complete release of bFGF from ECM. Addition of a methyl group next to the carboxylic group led to a preferential inhibition of heparanase activity. Similar results were obtained with a compound that contains one hydroxyl group para to the carboxylic group and an ether group near the carboxylic group on the phenol ring. Preferential inhibition of SMC proliferation was best achieved when the position of the hydroxyl group is para and ortho to the carboxylic group and the carboxylic group is at a distance of one carbon from the phenol ring. On the other hand, for maximal release of bFGF from ECM, the position of the carboxylic group should be three carbons away from the phenol ring. These new heparin mimicking compounds may have a potential use in inhibition of tumor metastasis, arteriosclerosis, and inflammation. PMID- 12124772 TI - Antagonistic effect of NK4 on HGF/SF induced changes in the transendothelial resistance (TER) and paracellular permeability of human vascular endothelial cells. AB - Hepatocyte growth factor/scatter factor (HGF/SF) is a multi-function cytokine that has been shown to regulate the expression of cell adhesion molecules in human endothelial cells. It is also a key cytokine in the development and progression of cancer, particularly during metastasis. NK4 is a variant of HGF/SF that has already been shown to be antagonistic to HGF/SF. This study shows that HGF/SF decreased transendothelial resistance (TER) and increased paracellular permeability in human vascular endothelial cells can that such effects can be inhibited by addition of the NK4 variant. In addition, HGF/SF-stimulated invasion of endothelium by breast cancer cells was inhibited by the addition of NK4. Western blotting revealed that HGF/SF decreased the protein level, and increased tyrosine phosphorylation of ZO-1, but did not cause a change in level of occludin or claudin-1, both molecules involved in tight junction function. RT-PCR revealed that addition of HGF/SF caused no change in signal for claudin-5 or junctional adhesion molecule (JAM), but there was a decrease in the signal for claudin-1. NK4 was able to prevent the decrease in levels of ZO-1 protein by HGF/SF. PMID- 12124774 TI - Prognostic significance of p16INK4a alterations and 9p21 loss of heterozygosity in locally advanced laryngeal squamous cell carcinoma. AB - The p16INK4a gene, localized within chromosome 9p21, has been identified as a cyclin-dependent kinase inhibitor and may negatively regulate the cell cycle acting as a tumor suppressor. Genetic alterations involving the 9p21 region are common in human cancers. A consecutive series of 64 untreated patients (median of follow up 53 months) undergoing surgical resection for locally advanced laryngeal squamous-cell carcinomas (LSCCs) has been studied prospectively. Our purpose was to investigate p16 alterations (9p21 allelic loss, hypermethylation and point mutations) and their possible association with clinico-pathological data and flow cytometric variables (DNA-ploidy and S-phase fraction (SPF)), and to determine the possible prognostic role of this gene in these tumors. PCR-based techniques were used for investigating 9p21 loss of heterozygosity (LOH) and methylation promoter status of the p16 gene. p16 mutations were detected by PCR-SSCP (single strand conformation polymorphism) and sequencing. 9p21 LOH was detected in 16/62 (26%) informative tumors, point mutations in 5% (3/64) and hypermethylation in 9% (6/64) of the cases. p16 alterations were significantly associated with high SPF and DNA-aneuploidy. By univariate analysis, poor histologic differentiation, stage IV, DNA-aneuploidy and p16 point mutations proved to be significantly related to quicker relapse, whereas these same factors, and in addition high SPF, 9p21 LOH and any p16 alterations were significantly related to shorter overall survival. By Cox proportional hazards analysis only histologic grade (G3) and p16 point mutations were independently related to both disease relapse and death. Our study has identified p16 point mutations as important biomolecular indicators in LSCCs. PMID- 12124775 TI - Heparan sulfate depletion within pulmonary fibroblasts: implications for elastogenesis and repair. AB - We investigated the role of sulfated proteoglycans in regulating extracellular matrix (ECM) deposition in pulmonary fibroblast cultures. Fibroblast cultures were subject to pharmacologic and enzymatic interventions to modify sulfated proteoglycan levels. Native and proteoglycan-depleted fibroblasts were treated with porcine pancreatic elastase at 2-4-day intervals and the elastase-mediated release of fibroblast growth factor 2 (FGF-2) and glycosaminoglycans was determined. Elastase treatment released significantly less FGF-2 and glycosaminoglycans (GAG) from PG-depleted fibroblasts with respect to native cells. Equilibrium ligand binding studies indicated that 125I FGF-2 binding at both cell surface receptor and heparan sulfate proteoglycan sites was reduced to different extents based on the method of proteoglycan depletion. Quantitation of elastin protein and message levels indicated that biological sulfation is required for the proper incorporation of tropoelastin into the extracellular matrix. These results suggest that sulfated proteoglycans play a central role in modulating pulmonary fibroblast extracellular matrix composition and are important mediators of elastolytic injury. PMID- 12124776 TI - Exendin-4 differentiation of a human pancreatic duct cell line into endocrine cells: involvement of PDX-1 and HNF3beta transcription factors. AB - Exendin-4 (EX-4), a long acting agonist of GLP-1, induces an endocrine phenotype in Capan-1 cells. Under culture conditions which include serum, approximately 10% of the cells contain insulin and glucagon. When exposed to EX-4 (0.1 nM, up to 5 days), the number of cells containing insulin and glucagon increased to approximately 40%. Western blot analysis detected a progressive increase in protein levels of glucokinase and GLUT2 over 3 days of EX-4 treatment. We explored the sequence of activation of certain transcription factors known to be essential for the beta cell phenotype: PDX-1, Beta2/NeuroD, and hepatocyte nuclear factor 3beta (HNF3beta). Double immunostaining showed that PDX-1 coexisted with insulin and glucagon in EX-4-treated cells. Treatment caused an increase in PDX-1 protein levels by 24 h and induced its nuclear translocation. Beta2/NeuroD protein levels also increased progressively over 24 h. HNF3beta protein level increased twofold as early as 6 h after EX-4 treatment. EMSA results indicated that EX-4 caused a 12-fold increase in HNF3beta binding to PDX 1 promoter area II. Beta2/NeuroD protein levels progressively increased after 24 h treatment. Differentiation to insulin-producing cells was also seen when Capan 1 cells were transfected with pdx-1, with 80% of these cells expressing insulin 3 days after transfection. PDX-1 antisense totally inhibited such conversion. During the differentiation of duct cells to endocrine cells, cAMP levels (EX-4 is a ligand for the GLP-1, G-protein coupled receptor) and MAP kinase activity increased. Our results indicate that EX-4 activates adenylyl cyclase and MAP kinase which, in turn, may lead to activation of transcription factors necessary for an endocrine phenotype. PMID- 12124777 TI - Role of leukemia inhibitory factor in the regulation of the proliferation and differentiation of neonatal mouse epidermal melanocytes in culture. AB - Mouse epidermal melanoblasts/melanocytes preferentially proliferated from disaggregated epidermal cell suspensions derived from newborn mouse skin in a serum-free melanoblast/melanocyte-proliferation medium supplemented with dibutyryl adenosine 3':5'-cyclic monophosphate (DBcAMP) and/or basic fibroblast growth factor (bFGF). Leukemia inhibitory factor (LIF) supplemented to the medium from initiation of primary culture increased the proliferation of melanoblasts or melanocytes as well as the differentiation of melanocytes. Pure cultured primary melanoblasts or melanocytes were further cultured with the medium supplemented with LIF from 14 days (keratinocyte depletion). LIF stimulated the proliferation of melanoblasts or melanocytes as well as the differentiation of melanocytes in the absence of keratinocytes. Moreover, anti-LIF antibody supplemented to the medium from initiation of primary culture inhibited the proliferation of melanoblasts or melanocytes as well as the differentiation of melanocytes. These results suggest that LIF is one of the keratinocyte-derived factors involved in regulating the proliferation and differentiation of neonatal mouse epidermal melanocytes in culture in cooperation with cAMP elevator and bFGF. PMID- 12124778 TI - GADD45b and GADD45g are cdc2/cyclinB1 kinase inhibitors with a role in S and G2/M cell cycle checkpoints induced by genotoxic stress. AB - Gadd45a (Gadd45), Gadd45b (MyD118), and Gadd45g (CR6) constitute a family of evolutionarily conserved, small, acidic, nuclear proteins, which have been implicated in terminal differentiation, growth suppression, and apoptosis. How Gadd45 proteins function in negative growth control is not fully understood. Recent evidence has implicated Gadd45a in inhibition of cdc2/cyclinB1 kinase and in G2/M cell cycle arrest. Yet, whether Gadd45b and/or Gadd45g function as inhibitors of cdc2/cyclinB1 kinase and/or play a role in G2/M cell cycle arrest has not been fully established. In this work, we show that Gadd45b and Gadd45g specifically interact with the Cdk1/CyclinB1 complex, but not with other Cdk/Cyclin complexes, in vitro and in vivo. Data also has been obtained that Gadd45b and Gadd45g, as well as GADD45a, interact with both Cdk1 and cyclinB1, resulting in inhibition of the kinase activity of the Cdk1/cyclinB1 complex. Inhibition of Cdk1/cyclinB1 kinase activity by Gadd45b and Gadd45a was found to involve disruption of the complex, whereas Gadd45g did not disrupt the complex. Moreover, using RKO lung carcinoma cell lines, which express antisense Gadd45 RNA, data has been obtained, which indicates that all three Gadd45 proteins are likely to cooperate in activation of S and G2/M checkpoints following exposure of cells to UV irradiation. PMID- 12124779 TI - Novel insulin/GIP co-producing cell lines provide unexpected insights into Gut K cell function in vivo. AB - Enteroendocrine (EE) cells represent complex, rare, and diffusely-distributed intestinal epithelial cells making them difficult to study in vivo. A specific sub-population of EE cells called Gut K-cells produces and secretes glucose dependent insulinotropic peptide (GIP), a hormone important for glucose homeostasis. The factors that regulate hormone production and secretion, as well as the timing of peptide release, are remarkably similar for K-cells and islet beta-cells suggesting engineering insulin production by K-cells is a potential gene therapeutic strategy to treat diabetes. K-cell lines could be used to study the feasibility of this potential therapy and to understand Gut K-cell physiology in general. Heterogeneous STC-1 cells were transfected with a plasmid (pGIP/Neo) encoding neomycin phosphotransferase, driven by the GIP promoter-only cells in which the GIP promoter was active survived genetic selection. Additional clones expressing pGIP/Neo plus a GIP promoter/insulin transgene were isolated-only doubly transfected cells produced preproinsulin mRNA. Bioactive insulin was stored and then released following stimulation with arginine, peptones, and bombesin-physiological GIP secretagogues. Like K-cells in vivo, the GIP/insulin producing cells express the critical glucose sensing enzyme, glucokinase. However, glucose did not regulate insulin or GIP secretion or mRNA levels. Conversely, glyceraldehyde and methyl-pyruvate were secretagogues, indicating cells depolarized in response to changes in intracellular metabolite levels. Potassium channel opening drugs and sulphonylureas had little effect on insulin secretion by K-cells. The K-cell lines also express relatively low levels of Kir 6.1, Kir 6.2, SUR1, and SUR2 suggesting secretion is independent of K(ATP) channels. These results provided unexpected insights into K-cell physiology and our experimental strategy could be easily modified to isolate/characterize additional EE cell populations. PMID- 12124780 TI - Infiltration anesthetic lidocaine inhibits cancer cell invasion by modulating ectodomain shedding of heparin-binding epidermal growth factor-like growth factor (HB-EGF). AB - Although the mechanism is unknown, infiltration anesthetics are believed to have membrane-stabilizing action. We report here that such a most commonly used anesthetic, lidocaine, effectively inhibited the invasive ability of human cancer (HT1080, HOS, and RPMI-7951) cells at concentrations used in surgical operations (5-20 mM). Ectodomain shedding of heparin-binding epidermal growth factor-like growth factor (HB-EGF) from the cell surface plays an important role in invasion by HT1080 cells. Lidocaine reduced the invasion ability of these cells by partly inhibiting the shedding of HB-EGF from the cell surface and modulation of intracellular Ca2+ concentration contributed to this action. The anesthetic action of lidocaine (sodium channel blocking ability) did not contribute to this anti-invasive action. In addition, lidocaine (5-30 mM), infiltrated around the inoculation site, inhibited pulmonary metastases of murine osteosarcoma (LM 8) cells in vivo. These data point to previously unrecognized beneficial actions of lidocaine and suggest that lidocaine might be an ideal infiltration anesthetic for surgical cancer operations. PMID- 12124781 TI - In vivo evaluation of a bioactive scaffold for bone tissue engineering. AB - Revision cases of total hip implants are complicated by the significant amount of bone loss. New materials and/or approaches are needed to provide stability to the site, stimulate bone formation, and ultimately lead to fully functional bone tissue. Porous bioactive glasses (prepared from 45S5 granules, 45% SiO2, 24.5% Na2O, 24.5% CaO, and 6% P2O5) have been developed as scaffolds for bone tissue engineering and have been studied in vitro. In this study, we investigated the incorporation of tissue-engineered constructs utilizing these scaffolds in large, cortical bone defects in the rat simulating revision conditions. With implantation times of 2, 4, and 12 weeks the results were compared to those using the bioactive ceramic scaffold alone. Two tissue-engineered constructs were studied: osteoprogenitor cells that were either seeded onto the scaffold prior to implantation ("primary") or those that were culture expanded to form bonelike tissue on the scaffold prior to implantation ("hybrid"). Defects treated with the hybrid had the greatest amount of bone in the available pore space of the defect over all other groups at 2 weeks (p < 0.05). For both the primary and hybrid groups, woven and lamellar bone was present along the interface of the scaffold and the host cortex and within the porous space of the scaffold at 2 weeks. By 4 weeks, very uniform, lamellar bone was present throughout the scaffold for both tissue-engineered groups. The amount of bone significantly increased over time for all groups while the bioactive ceramic gradually resorbed by 40% at 12 weeks (p < 0.05). Structural properties of the treated long bones improved over time. Long bones treated with the hybrid had an early return in torsional stiffness by 2 weeks. Both tissue-engineered constructs achieved normal torsional strength and stiffness by 4 weeks as compared to the scaffold alone, which achieved this by 12 weeks. Porous, surface modified bioactive ceramic is a promising scaffold material for tissue-engineered bone repair. PMID- 12124782 TI - Poly(glutamic acid) poly(ethylene glycol) hydrogels prepared by photoinduced polymerization: Synthesis, characterization, and preliminary release studies of protein drugs. AB - A class of new biodegradable hydrogels based on poly(ethylene glycol) methacrylate-graft-poly(glutamic acid) and poly(ethylene glycol) dimethacrylate was synthesized by photoinduced polymerization. Because all the polymeric constituents were highly hydrophilic, crosslinking could be performed in aqueous solutions. This type of crosslinked hydrogel was prepared by modifying a select number of acidic side-groups on poly(glutamic acid) with poly(ethylene glycol) methacrylate. These modified chains were then crosslinked in the presence of poly(ethylene glycol) dimethacrylate under a photoinduced polymerization at a wavelength of 365 nm. Swelling experiments were conducted to study the crosslinking density, pH-responsive behavior, and degradation of the hydrogel. Results showed that the degree of swelling of this type of hydrogels increased as the crosslinker concentration (or density) was reduced. Because of the presence of acidic side chains on poly(glutamic acid), swelling behavior was found to be pH-responsive, increasing at high pH in response to the increase in the amount of ionized acidic side chains. The degradation rate of these hydrogels also varied with pH. More rapid degradation was observed under stronger alkaline conditions because of the hydrolysis of the ester bonds between the crosslinker and the polymer backbone. Practically useful degradation rates could be achieved for such hydrogels under physiological conditions. Drug release rates from these hydrogels were found to be proportional to the protein molecular weight and the crosslinker density; increasing at lower protein molecular weight or crosslinker density. The preliminary findings presented in this article suggest that this class of biodegradable hydrogels could be an attractive avenue for drug delivery applications. The specific photoinduced crosslinking chemistry used would permit hydrogels to be synthesized in existence of the entrapped macromolecular drugs including peptides, proteins, and cells. In addition, the rapid feature of this polymerization procedure along with the ability to perform hydrogel synthesis and drug loading in an aqueous environment would offer great advantages in retaining drug activity during hydrogel synthesis. PMID- 12124783 TI - Influence of cosolvents and in situ forming hydroxyapatite on the mechanical characteristics of collagen films. AB - Collagen was processed into films in mixtures containing various ratios of water, propylene glycol, and ethanol. An experimental mixture design was applied to characterize the effects of individual solvents and their interactions on the mechanical properties of collagen films. Scanning electron microscopy (SEM) was used to examine the surface properties of collagen films. The ultimate tensile strength (UTS) and related characteristics of collagen films were also evaluated with dynamic mechanical analysis. The effect of in situ forming hydroxyapatite (HAP) within collagen films at a concentration of 10 mM on the physical characteristics of these films was evaluated by the same methods. With X-ray and SEM examinations, it was confirmed that HAP was formed inside the collagen film. However, the UTS of collagen films without HAP was 4-5 times higher than that with HAP. This was probably due to the discontinuity of the film structure caused by HAP in the collagen films. The results of a statistical analysis of the experimental design revealed the influence of the solvent mixtures on the mechanical properties of the collagen films with and without HAP, showing similar responses for the UTS and modulus of elasticity. Both parameters showed a maximal response in the solvent range containing a lower percentage of ethanol with the desired percentage of propylene glycol to plasticize the collagen films. PMID- 12124784 TI - Behavior of dense and porous hydroxyapatite implants and tissue response in rat femoral defects. AB - Porous and dense hydroxyapatite cylinders (PHA and DHA) were implanted into cavities produced in rat femora and the sites of implantation were examined at different times over a period of 24 weeks by microradiologic and histological techniques. Microradiographs showed the presence of a layer of trabecular bone around the implants, which became more radiopaque and thinner along the experimental time. The microradiologic methodology used was suitable for the evaluation of the interface between hydroxyapatite and newly formed bone in nondecalcified materials. Microscopic observations showed that young bone grew over the surface of both types of implants after 1 and 2 weeks of surgery and that bone also grew inside PHA implants. Progressive bone absorption was observed in both types of implants after the fourth week. A layer of fibrous tissue was formed in the interface between new bone and DHA. Mature bone with haversian systems surrounded DHA implants and filled the pores of PHA implants throughout the experimental period. The pores of PHA implants were smaller than those commonly reported, which should have been a disadvantage, although it was observed that the extra cellular fluid induced disintegration of the ceramic granules, allowing the gradual growth of bone tissue into the spaces among them, without the interposition of fibrous tissue. PMID- 12124785 TI - Raman and IR studies on adsorption behavior of adhesive monomers in a metal primer for Au, Ag, Cu, and Cr surfaces. AB - 6-[N-(4-vinylbenzyl)propylamino]-1,3,5-triazine-2,4-dithione (VBATDT) and 10 methacryloyloxydecyl dihydrogen phosphate (M10P) are functional monomers used for the surface treatment of dental alloys. The aim of our study was to clarify the role of a commercial metal primer containing both the monomers in adhesion between resin and various dental metals on a molecular level. We used surface enhanced Raman scattering (SERS) and infrared reflection absorption (IRA) spectroscopy. An SERS measurement was performed with a 647 nm laser line for a mixture of aqueous Au colloid and the primer. IRA spectra were taken for cast films of the primer on Au, Ag, Cu, and Cr surfaces as a function of rinse time, and for self-assembled monolayer (SAM) films from dilute mixed solution of VBATDT and M10P. These spectra indicate that VBATDT in the primer is mainly chemisorbed on Au, Ag, and Cu surfaces with respect to thickness, whereas only M10P is adsorbed on Cr. We also examined the tensile bond strengths between resin and Au, Ag, Cu, and Cr plates treated by VBATDT, with and without M10P, and found that VBATDT effectively promotes the bond strength between resin and the metals except for Cr, whereas M10P is effective only for Cr. These adhesion characteristics are consistent with the chemisorbed species on each metal surface as shown in the spectroscopic evidence. PMID- 12124786 TI - Glycosaminoglycan mimetics (RGTA) modulate adult skeletal muscle satellite cell proliferation in vitro. AB - Muscle regeneration occurs through the activation of satellite cells, which are stimulated to proliferate and to fuse into myofibers that will reconstitute the damaged muscle. We have previously reported that a family of new compounds called "regenerating agents" (RGTAs), which are polymers engineered to mimic heparan sulfates, stimulate in vivo tissue repair. One of these agents, RG1192, a dextran derivative substituted by CarboxyMethyl, Benzylamide, and Sulfate (noted CMBS, RGTA type), was shown to improve greatly the regeneration of rat skeletal muscle after severe crushing, denervation, and acute ischemia. In vitro, these compounds mimic the protecting and stabilizing properties of heparin or heparan sulfates toward heparin-binding growth factors (HBGFs). We hypothesized that RGTA could act by increasing the bioavailability of some HBGF involved in myoblast growth and thus asked whether RGTA would alter the ability of satellite cells to proliferate. Its effect was tested on primary cultures of rat satellite cells. The RG1192 stimulated the proliferation of satellite cells in vitro in a dose dependent manner. It appeared to be as efficient as natural glycosaminoglycans (GAGs; heparan sulfate, dermatan sulfate, or keratan sulfate) in stimulating satellite cell proliferation but was about 100 times more efficient than heparin. RG1192 stimulated satellite cell proliferation by increasing the potency of fibroblast growth factor 2 and scatter factor-hepatocyte growth factor. It also partially restored myoblast proliferation of satellite cells with chlorate induced hyposulfation. Taken together, our results explain to some extent the improving effect of RGTA with a CMBS structure, such as the RG1192, on muscle regeneration in vivo by providing support for the hypothesis that RGTA may act by increasing the potency of some HBGFs during the proliferation phase of the regenerating muscle. PMID- 12124787 TI - Interactions between MC3T3-E1 cells and textured Ti6Al4V surfaces. AB - This paper presents the results of an experimental study of the interactions between MC3T3-E1 (mouse calvarian) cells and textured Ti6Al4V surfaces, including surfaces produced by laser microgrooving; blasting with alumina particles; and polishing. The multiscale interactions between MC3T3-E1 cells and these textured surfaces are studied using a combination of optical scanning transmission electron microscopy and atomic force microscopy. The potential cytotoxic effects of microchemistry on cell-surface interactions also are considered in studies of cell spreading and orientation over 9-day periods. These studies show that cells on microgrooved Ti6Al4V geometries that are 8 or 12 microm deep undergo contact guidance and limited cell spreading. Similar contact guidance is observed on the surfaces of diamond-polished surfaces on which nanoscale grooves are formed due to the scratching that occurs during polishing. In contrast, random cell orientations are observed on alumina-blasted Ti6Al4V surfaces. The possible effects of surface topography are discussed for scar-tissue formation and improved cell-surface integration. PMID- 12124788 TI - Mechanical behavior of chemically treated ostrich pericardium subjected to uniaxial tensile testing: influence of the suture. AB - The mechanical behavior of sutured ostrich pericardium was studied by uniaxial tensile testing. One hundred forty-four tissue specimens were assessed: 96 sutured samples (48 in which a centrally located suture was placed at an angle of 90 degrees with respect to the longitudinal axis, whereas in the remaining 48, a centrally located suture was placed at a 45 degrees angle to the longitudinal axis, in sets of 12 samples each, sewn with sutures made of Gore-Tex, nylon, Prolene, or silk), and 48 unsutured controls. Each group of 24 samples sewn at one angle or the other with the different suture materials was assayed together with a corresponding control group of 12 unsutured samples. The mean tensile strengths in the unsutured controls ranged between 30.16 MPa and 43.42 MPa, whereas those of the sutured sets ranged from 14.68 MPa to 21.91 MPa. The latter presented a statistically significant loss of resistance (p < 0.01) when compared with the unsutured tissue samples. The angle of the suture with respect to the longitudinal axis influenced the degree of shear stress produced by the suture, as well as the behavior of the different suture materials used. The set of samples sewn with Prolene appeared to be that most sensitive to changes in the angle of the suture, whereas tissue sewn at a 45 degrees angle with Gore-Tex presented lower shear stress values in comparison with samples in which the other three materials were used. A method of tissue selection based on morphological and mechanical criteria was used to ensure the homogeneity of the results in such a way that the coefficients of determination (R2) for the stress/strain curve fitting equation ranged between 0.888 and 0.995. This excellent fit made it possible, applying regression analysis, to predict the mechanical behavior of a specimen by determining that of a contiguous tissue sample. Thus, it should be possible, at least theoretically, to characterize the behavior of a specific region or zone of the biomaterial. In conclusion, ostrich pericardium exhibits strong resistance to rupture, even when sutured. The selection method used ensures the homogeneity of the samples and, thus, of the results. The angle of the suture with respect to the longitudinal axis, where the load is centered, determines the shear stress produced by the suture and the mechanical behavior of each suture material. PMID- 12124789 TI - In situ study of partially crystallized Bioglass and hydroxylapatite in vitro bioactivity using atomic force microscopy. AB - The present work investigates, in situ, the in vitro bioactivity of partially crystallized 45S5 Bioglass (BG) as a function of immersion time in a simulated body fluid (SBF) using atomic force microscopy (AFM). The results obtained for the crystallized BG were compared to those of hydroxyapatite c- and a-faces. The calcium phosphate layer grows on the crystallized 45S5 B by multiple two dimensional nucleation and fusion of these two-dimensional islands, which is essentially the same mode as for the hydroxyapatite c-face. The surface of the crystallized 45S5 BG was almost fully covered with a dense and compact calcium phosphate layer after 24 h. The calcium phosphate formation on the crystallized BG arises from a low surface energy of the surface layer and/or an effect of the layer to lower the resistance when the growth units of calcium phosphate incorporate into the growing island. These results indicate that the crystallized 45S5 BG is suitable to be used as a filler for polymeric matrix bioactive composites, as it maintains a high bioactivity associated with a stiffer behavior (as compared to standard BG). PMID- 12124790 TI - Resorbable defect analog PLGA scaffolds using CO2 as solvent: structural characterization. AB - After tooth extraction, the immediate wound treatment by implanting an exact copy of the root could prevent alveolar bone atrophy. The implant should have an interconnected porosity in order to promote tissue in-growth. This communication reports a novel method to realize such net-shaped porous scaffolds fabricated within a few minutes. Porosity and micro-architecture are evaluated by Hg porosimetry and by image analysis of electron and light microscopy as well as by computed micro-tomography. The total porosity of the scaffold corresponds to (63 +/- 3)%, mainly related to open interconnected porosity. Micro-tomography, as a noninvasive 3D method, is best suited to uncover pores of about 100 microm, a diameter especially important for tissue in-growth. The differentiation between open and closed porosity, however, depends on the method chosen. This effect is attributed to the spherical pores with an orifice only detected in the 3D analysis. Consequently, the closed porosity is overestimated by 8% evaluating 2D images. Finally, the mean pore diameter is found to be 106 and 100 microm for 2D and 3D analysis, respectively. Although the porosity of the scaffold needs to be further optimized for clinical applications, the procedure proposed is a promising route in manufacturing open porous implants without the use of any organic solvent. PMID- 12124791 TI - Synthesis of functionally graded MgCO3 apatite accelerating osteoblast adhesion. AB - As a means of improving the biological properties of materials for use as bone substitutes, functionally graded carbonate apatite containing Mg, FGMgCO3Ap, was synthesized at 60 degrees C and pH 7.4 using a gradient magnesium supply system. X-ray diffraction analysis of FGMgCO3Ap showed a poorly crystallized apatitic pattern, similar to that of human bone. ESCA analysis clearly showed the negative gradient distribution in Mg1s intensity (atomic concentration) of magnesium from the crystal surface toward the inner core. When the FGMgCO3Ap crystals were mixed with collagen, the resulting FGMgCO3Ap-collagen composite, irradiated with UV light for 4 h, retained their features in the saline solution. After washing away the nonadhesive cells, a cell adhesion assay showed that the optical density of the FGMgCO3Ap-collagen composite was higher than that of the CO3Ap-collagen composite. SEM observation showed that the osteoblast-like cells adhered well to the surface of the FGMgCO3Ap-collagen composite. Staining with hematoxylin-eosin and alizarin red confirmed the existence of a great many more cells and a thicker extracellular matrix layer on the FGMgCO3Ap-collagen composite than on the CO3Ap collagen composite. This result demonstrated the acceleration effect of magnesium ions on osteoblast adhesion on the FGMgCO3Ap-collagen composite. PMID- 12124792 TI - Analyses of rampant corrosion in stainless-steel retainers of orthodontic patients. AB - Retainers were collected from private, university, and dental labs. After viewing these corroded and control appliances using scanning electron microscopy, corroded maxillary and mandibular retainers were selected along with a control stainless-steel retainer for in-depth chemical analysis. Using electron spectroscopy for chemical analysis, monochromated Al x-rays were rastered over areas 1.5 x 0.3 mm. After survey spectra were acquired, high-resolution multiplex scans were obtained and binding energy shifts were noted. Using Auger electron spectroscopy, a spot size of approximately 30 nm was analyzed. Photos, survey scans, and depth profiles were acquired using a 3.5kV Ar(+) ion beam that was calibrated using a SiO2 standard. Via electron spectroscopy for chemical analysis, the brown stains contained Fe and Cr decomposition products in which three carbon species were present. Proteinaceous N was found as amines or amides. No Ni was present because it had solubilized. The Cr:Fe ratio indicated severe Cr depletion in the stained regions (0.2) versus the control regions (1.3). The stained regions appeared mottled, having both dark and light areas. Via AES, the dark versus light areas of the stained regions indicated that there was an absence versus a presence of both Cr and Ni. In the dark areas corrosion penetrated 700 nm; in the light areas the depth equaled 30 nm. By comparison, the passivated layer of the control retainer was 10-nm thick. After sputtering away the affected areas, all specimens had similar spectra as the control regions. The bacterial environment created the mottled appearance and induced electrochemical potential differences so that, upon reducing the passivated layer, an otherwise corrosion-resistant alloy became susceptible to rampant corrosion. An integrated biological-biomaterial model is presented for the classic case of an orthodontic acrylic-based stainless steel retainer subject to crevice corrosion. PMID- 12124793 TI - A reporter-cell assay for the detection of BMP-2 immobilized on porous and nonporous materials. AB - Human recombinant bone morphogenetic protein-2 (rhBMP-2) immobilized on the surface of metal implants can facilitate osseointegration. Here, we describe a cell reporter assay useful for quantifying small amounts of immobilized rhBMP-2 on various materials. The peptide was dotted and heat-fixed on titanium, 316L stainless steel, nitrocellulose, or glass, and its distribution was monitored by in situ biotinylation followed by detection with the avidin-biotin method. Bioactivity of rhBMP-2 was demonstrated by means of a confluent layer of osteoblastic MC3T3-E1 cells that evenly covered rhBMP-2-free and rhBMP-2-loaded surface areas, as shown with epifluorescence microscopy of calcein acetoxymethyl (AM)-loaded cells. Expression of osteocalcin, fibronectin, actin, and vimentin increased where cells were located on rhBMP-2 dotted areas, but the signal:noise ratio was too low to bioassay the peptide. However, local pronounced expression of alkaline phosphatase was used to quantify BMP-2 in the range of 5-80 ng/dot by means of a cytochemical color reaction for alkaline phosphatase and image analysis of resulting dots. The lower detection limit was in the order nitrocellulose > glass > titanium > 316L steel. We conclude that the cell reporter assay is useful to assess biological activity of rhBMP-2 even after immobilization on three-dimensional implant materials. PMID- 12124794 TI - Hyaluronate-heparin conjugate gels for the delivery of basic fibroblast growth factor (FGF-2). AB - The stability and activity of recombinant growth factors administered locally for the repair of damaged tissue can be directly influenced by the physical structure and chemical composition of the delivery matrix. This study describes a novel basic fibroblast growth factor-2 (FGF-2) delivery system synthesized by the conjugation of a structure-stabilizing polymer, hyaluronate (HA), with a sulfated glycosaminoglycan, heparin (HP), that has inherent specific binding sites for members of the FGF family. The biopolymers were formed via stable amine or labile imine bonds by coupling amine-modified HA with oxidized heparin. The addition of recombinant human FGF-2 resulted in the rapid binding of FGF-2 to the heparin segment of the hyaluronate-heparin (HAHP) conjugate. The FGF-2 was released in vitro from the imine-bonded (HAHPi) gels in the form of FGF-2-heparin complexes through the hydrolysis of the imine bonds. In contrast, the release of growth factor from the more stable amine-bonded (HAHPa) gels required treatment with free heparin or enzymatic digestion of the hyaluronate segment. Functional analysis of the released FGF-2 showed that the HAHP conjugate gels increased both the stability and activity of the growth factor. PMID- 12124795 TI - Formation of three-dimensional cell/polymer constructs for bone tissue engineering in a spinner flask and a rotating wall vessel bioreactor. AB - The aim of this study is to investigate the effect of the cell culture conditions of three-dimensional polymer scaffolds seeded with rat marrow stromal cells (MSCs) cultured in different bioreactors concerning the ability of these cells to proliferate, differentiate towards the osteoblastic lineage, and generate mineralized extracellular matrix. MSCs harvested from male Sprague-Dawley rats were culture expanded, seeded on three-dimensional porous 75:25 poly(D,L-lactic co-glycolic acid) biodegradable scaffolds, and cultured for 21 days under static conditions or in two model bioreactors (a spinner flask and a rotating wall vessel) that enhance mixing of the media and provide better nutrient transport to the seeded cells. The spinner flask culture demonstrated a 60% enhanced proliferation at the end of the first week when compared to static culture. On day 14, all cell/polymer constructs exhibited their maximum alkaline phosphatase activity (AP). Cell/polymer constructs cultured in the spinner flask had 2.4 times higher AP activity than constructs cultured under static conditions on day 14. The total osteocalcin (OC) secretion in the spinner flask culture was 3.5 times higher than the static culture, with a peak OC secretion occurring on day 18. No considerable AP activity and OC secretion were detected in the rotating wall vessel culture throughout the 21-day culture period. The spinner flask culture had the highest calcium content at day 14. On day 21, the calcium deposition in the spinner flask culture was 6.6 times higher than the static cultured constructs and over 30 times higher than the rotating wall vessel culture. Histological sections showed concentration of cells and mineralization at the exterior of the foams at day 21. This phenomenon may arise from the potential existence of nutrient concentration gradients at the interior of the scaffolds. The better mixing provided in the spinner flask, external to the outer surface of the scaffolds, may explain the accelerated proliferation and differentiation of marrow stromal osteoblasts, and the localization of the enhanced mineralization on the external surface of the scaffolds. PMID- 12124797 TI - Human breast adenocarcinoma cell lines promote angiogenesis by providing cells with uPA-PAI-1 and by enhancing their expression. AB - During angiogenesis, endothelial cells use uPA and PAI-1 to migrate and degrade the basement membrane surrounding capillary blood vessels. Invasive tumor cells produce a large amount of uPA that could bind uPAR present at the endothelial cell surface to facilitate their invasion. To verify this hypothesis, endothelial cells were incubated with conditioned medium (CM) from two breast cancer cell lines (MCF7 and MDA MB 231 cells). Within a short incubation period (30 min) with both CM, an increase of uPA, PAI-1 and uPA-PAI-1 complex was detected in endothelial cell layer as assessed by casein zymography, ELISA and uPA immunostaining. The extent of this enhancement was related to the levels of uPA secreted by tumor cells (high in MDA MB 231 cells and low in MCF7 cells). After 2 hr of incubation, the CM from both tumor cells upregulated uPA and PAI-1 mRNA levels in endothelial cells in a time-dependent manner. The uPA increase in the cell layer could not be attributable to an increase of uPAR level. Only the CM from highly invasive MDA MB 231 cells increased the angiogenic morphotype of endothelial cells assessed in a collagen gel. A single addition of amino-terminal fragment of uPA (ATF) was able to abolish the angiogenic effect induced by MDA MB 231 cell CM. Our data demonstrate that the interactions occurring between breast tumor cells and endothelial cells can modulate tumor angiogenesis at least by two mechanisms: an increase of uPA and PAI-1 cell surface-binding and of their expression by endothelial cells. PMID- 12124796 TI - Osteoblast culture on polished titanium disks modified with phosphonic acids. AB - Titanium is widely used in dental implants due to its suitable physical properties and its good biocompatibility. However, it is integrated into bone only passively, and the resulting fixation in the bone, which is necessary for the function, is mainly mechanical in nature. With the objective of increasing the chemical interaction between the implant and the bone tissue, several phosphonic acids were synthesized and grafted onto titanium disks. Here we report on the proliferation, differentiation, and protein production of rat osteoblastic cells (CRP10/30) on phosphonic-acid-modified titanium surfaces studied in vitro. No statistical differences were found in osteoblast proliferation among the phosphonic-acid-modified titanium, unmodified titanium, and tissue culture plastic (used as a positive control), indicating that the phosphonic acids used were not cytotoxic to the osteoblasts used. For all surfaces (modified or not), the alkaline phosphatase activity was at least as good as it was on tissue culture plastic. However, the total amount of protein, and especially the collagen type I synthesis, was sensitive to surface modification. On titanium modified with ethane-1,1,2-triphosphonic acid, the total amount of synthesized protein was significantly higher than it was on unmodified titanium surfaces. A significant increase (up to 16%) of collagen type I production was observed on titanium surfaces modified with this acid or with methylenediphosphonic acid compared to unmodified titanium surfaces. PMID- 12124798 TI - Characterization of androgen receptor and nuclear receptor co-regulator expression in human breast cancer cell lines exhibiting differential regulation of kallikreins 2 and 3. AB - Accumulating evidence indicates that androgens and the androgen receptor modulate the development and progression of breast adenocarcinoma; however, the precise role and actions remain poorly defined. We examined previously the steroid hormone regulation of 2 known androgen-regulated kallikreins, KLK3 (encoding PSA) and KLK2 (encoding human kallikrein 2 or hK2) in BT-474, T-47D, ZR75-1, MCF-7, MFM-223 and BT-20 human breast cancer cells and found that they were differentially regulated, with the cells showing variable responses to androgen. To determine if this variable response was reflected by differences in androgen receptor, we characterized the expression of androgen receptor in these cells by Western blot analysis and saturation binding analysis. In addition, we sequenced androgen receptor cDNA from each of these cell lines to check whether any androgen receptor mutations were present. The expression of 11 nuclear receptor co-regulatory factors (SRC-1, AIB1, ARA24, ARA54, ARA55, ARA70, ARA160, FHL2, PDEF, NCoR1, SMRT) was compared in these cell lines by semi-quantitative RT-PCR to determine if the pattern of receptor co-activators or -repressors expressed in these cells might explain the differential regulation of KLK2 and KLK3. The levels of androgen receptor varied among the cell lines, but did not correlate with hK2 and PSA secretion determined previously. No mutations within the coding regions of the receptor were detected. With the exception of receptor expressed by MCF-7 cells, the polymorphic CAG repeat length was in the normal range. Every breast cancer cell line exhibited a distinct expression pattern of the nuclear receptor co-regulators examined raising the possibility that the relative levels of these co-activators/-repressors might differentially modulate androgen receptor transcriptional activity within the promoter/enhancer region of KLK2 and KLK3 of these cells. PMID- 12124799 TI - Inhibitory effect of a selective cyclooxygenase-2 inhibitor on liver metastasis of colon cancer. AB - COX-2 overexpression is recognized in various cancers, but the role of COX-2 in the progression of cancer, including the liver metastasis of colon cancer, is not clearly understood. We examined the role of COX-2 in the mechanism of liver metastasis of colon cancer, using a highly metastasizable colon carcinoma cell line, LM-H3. A COX-2 inhibitor, JTE-522, inhibited cell proliferation and invasion of LM-H3 in vitro and clearly reduced the number of metastatic nodules on the surface of nude mouse livers in vivo. We also examined the effects of JTE 522 on the production of growth factors and MMPs through the use of ELISA and gelatin zymography, respectively. JTE-522 downregulated PDGF production by LM-H3 but had no influence on VEGF production. JTE-522 also inhibited MMP-2 secretion by LM-H3. JTE-522 downregulated PGE(2) production, but the associated changes in PGE(2) did not affect PDGF and VEGF production by LM-H3. We conclude that JTE-522 downregulated the cell proliferation and invasive potential of LM-H3 by reducing the production of PDGF and MMP-2 and hypothesize that these inhibitory effects on the production of PDGF and MMP-2 can lead to inhibition of liver metastasis of colon cancer. These data indicate that the COX-2 inhibitor JTE-522 has a high potential for use as a clinical agent for the treatment of liver metastasis of colon cancer. PMID- 12124800 TI - Is growth inhibition and induction of apoptosis in lung cancer cell lines by fenretinide [N-(4-hydroxyphenyl)retinamide] sufficient for cancer therapy? AB - The synthetic retinoid fenretinide [N-(4-hydroxyphenyl)retinamide, 4-HPR] has demonstrated growth inhibition and induction of apoptosis of various malignant cells, including lung cancer cell lines. 4-HPR is now being investigated in several clinical trials. In our study, we show that 4-HPR inhibits growth on a broad panel of lung cancer cell lines (12/12 small cell lung cancer and 9/12 nonsmall cell lung cancer cell lines), including cell lines unresponsive to all trans-retinoic acid (ATRA). 4-HPR revealed a higher potency than ATRA in inhibiting cell growth with IC(50) values ranging from 3.3-8.5 microM. Furthermore, 4-HPR induces apoptosis in lung cancer cell lines as proven by TUNEL and annexin V assay. Despite this, we observed stimulation of growth in 2 SCLC cell lines at 1 microM 4-HPR. In advance to the clinical application of 4-HPR, we demonstrate that growth inhibition is reversible after removal of 4-HPR and that long-term application is necessary. Through long-term stimulation with 4-HPR, we cultivated 3 resistant cell lines that were still inhibited by 4-HPR after several weeks, however, exhibited almost no apoptosis. These cell lines exhibited morphologic changes, which in the case of the SCLC cell lines suggested differentiation. Our data show that 4-HPR inhibits growth in lung cancer cell lines by varying mechanisms including (i) cytostasis, (ii) apoptosis and (iii) presumably, differentiation. In contrast, the observed growth stimulation, reversibility of growth inhibition and development of resistance to apoptosis make successful cancer therapy uncertain and may limit clinical application of 4 HPR in lung cancer patients, although its inhibitory effects last over several weeks. PMID- 12124801 TI - A synthetic matrix metalloproteinase inhibitor prevents squamous carcinoma cell proliferation by interfering with epidermal growth factor receptor autocrine loops. AB - Head and neck squamous cell carcinoma (HNSCC) is characterized by its capacity to invade adjacent tissues and to metastasize locoregionally. Evidence suggests that matrix metalloproteinases (MMPs) may play a causal role in HNSCC progression. While evaluating the role of MMPs in the invasion process, we made the surprising observation that a broad-spectrum MMP inhibitor, (marimastat, BB2516), inhibited the growth in vitro of some HNSCC cell lines. This inhibitory effect was only found in HNSCC cell lines overexpressing epidermal growth factor receptors. The effects of the MMP inhibitor could be reversed by adding exogenous c-erbB ligands, suggesting that the phenomenon may be related to autocrine ligand processing. This hypothesis was supported by the finding that the growth inhibitory effect of marimastat was directly related to its ability to prevent the release of major c-erbB ligands including transforming growth factor-alpha, betacellulin and heregulin beta1 from HNSCC. Marimastat was also found to potentiate the cytotoxic effects of cisplatin both in vitro and in vivo. Our results indicate that the cleavage of several c-erbB ligands from membrane anchored precursors requires MMP activity. We conclude that MMP inhibitors could prevent tumor progression not only by inhibiting invasion and angiogenesis, as previously shown, but also by their ability to inhibit autocrine signaling through the c-erbB receptors. Clinical trials to test this hypothesis in HNSCC should be considered. PMID- 12124802 TI - Critical tumor-suppressor gene regions on chromosome 3P in major human epithelial malignancies: allelotyping and quantitative real-time PCR. AB - To ascertain the involvement of human chromosome 3p and its established critical TSG regions in various epithelial malignancies, 21 polymorphic and 2 nonpolymorphic 3p markers were allelotyped in nonpapillary RCC, NSCLC, CC and BC from a total of 184 patients. LOH was observed with high frequency in all types of cancer studied: RCC (52/57, 91%), BC (41/51, 80%), NSCLC (30/40, 75%) and CC (27/36, 75%). Interstitial deletions, believed to signal TSG inactivation, were verified using the "L-allele rule" and real-time quantitative PCR. Significant correlation was observed between DNA copy numbers for 2 nonpolymorphic STS markers and LOH data for adjacent polymorphic loci. Interstitial deletions in 3p were demonstrated for all cancer types studied. However, the distribution of different types of deletion was characteristic for tumors from various locations. Large terminal deletions were predominantly seen in RCC and NSCLC (51% and 40%, respectively), correlating with clear cell RCC and squamous cell carcinomas of the lung. In addition to the LUCA region at 3p21.3 (centromeric), we found that the AP20 region (3p21.3, telomeric) was frequently affected in all 4 cancers, suggesting that this newly defined critical region contains multiple TSGs. Moreover, at least 3 candidate cancer-specific loci were identified. The telomeric 3p26.1-p25.3 region was predominantly deleted in RCC and NSCLC. The D3S1286 and D3S3047 markers (3p25.2-p24.3) were deleted nonrandomly in NSCLC. High-frequency LOH was detected in a segment mapped closely distal to the LUCA site (3p21.3), around the D3S2409 and D3S2456 markers. PMID- 12124803 TI - Detection of free-circulating tumor-associated DNA in plasma of colorectal cancer patients and its association with prognosis. AB - Tumor cells are characterized by specific genetic alterations. When such genetic alterations are identified in body fluid including plasma, regardless of the presence of detectable tumor cells, it shows the existence of free-circulating tumor-associated DNA. The objective of our study was to assess the prognostic value of free-circulating tumor-associated DNA in colorectal cancer patients' plasma. The first step of our work was to find common genetic alterations in tumors that would subsequently be used for plasma DNA screening. We focused on KRAS2 mutations in codons 12 and 13 by the mutant allele-specific amplification (MASA) method and p16 hypermethylation by the methylation-specific polymerase chain reaction (MSP) method. Patients with a tumor presenting either alteration were selected for plasma screening; 58 tumors were analyzed for KRAS2 mutations and tested for p16 gene promoter methylation. Survival and recurrence rates were assessed in patients with and without free-circulating tumor-associated DNA alterations in plasma. Of the 58 tumors analyzed, 39 (67%) demonstrated either one or both of the studied genetic alterations. Twenty-two (38%) were mutated at KRAS2, and an identical alteration was detected in 10 (45%) of the 22 corresponding plasma samples. Thirty-one (53%) had p16 gene promoter hypermethylation that could also be detected in the plasma in 21 cases (68%). Among the 39 patients who had one or the other alteration in tumor DNA, 37 had at least one reliable plasma test. In 26 (70%) of the 37 patients, free-circulating tumor-associated DNA was detected in plasma. The 2-year overall survival rate was 48% in the group where free-circulating tumor-associated DNA was detected in plasma and 100% in the one where free-circulating tumor-associated DNA was not detected in plasma (p < 0.03). Among these 37 patients, 25 patients had a stage I, II or III disease. In this subgroup of patients, the 2-year recurrence-free survival rate for the 17 patients with free-circulating tumor-associated DNA detected in plasma was 66%, compared to 100% for the 8 patients without free circulating tumor-associated DNA detected in plasma (p = 0.044). The presence of free-circulating tumor-associated DNA in plasma seems to be a relevant prognostic marker for patients with colorectal cancer and may be used to identify patients with a high risk of recurrence. PMID- 12124804 TI - Molecular genetic analysis of malignant melanomas for aberrations of the WNT signaling pathway genes CTNNB1, APC, ICAT and BTRC. AB - Aberrant activation of the Wnt signaling pathway has been reported in different human tumor types, including malignant melanomas. We investigated 37 malignant melanomas (15 primary tumors and 22 metastases) for alterations of 4 genes encoding members of this pathway, i.e., CTNNB1 (beta-catenin gene, 3p22.1), APC (adenomatous polyposis coli gene, 5q22.2), BTRC (beta-transducin repeat containing protein gene, 10q24.3) and ICAT (inhibitor of beta-catenin and Tcf-4, 1p36.2). Mutational analysis of CTNNB1 identified somatic mutations in 1 primary melanoma and 1 melanoma metastasis from 2 different patients (5%). Both mutations affected the N-terminal degradation box of beta-catenin, which is important for the regulation of beta-catenin homeostasis. Another primary melanoma carried a somatic APC missense mutation within the known mutation cluster region in exon 15. Fourteen tumors (40%) showed LOH at microsatellite markers on 1p36. None of the tumors had lost both copies of the ICAT gene, but 1 melanoma metastasis carried a somatic point mutation altering the translation start codon of ICAT. Real-time RT-PCR showed markedly reduced ICAT transcript levels ( 0.12 microg/l with poor overall (p = 0.021 and p = 0.0009) and progression-free survival (p < 0.0005 and p = 0.0025). Multivariate analysis revealed disease stage (p = 0.0093) and tumor burden (p < 0.0005) as independent predictive factors for overall survival, and sHLA-DR (p = 0.0007) and tumor burden (p = 0.0015) for progression-free survival. In contrast to S100 beta, sHLA-DR serum concentrations < 0.3 microg/ml were strongly associated (p = 0.0001) with poor progression-free survival in a subgroup of 60 nonmetastasized patients. In conclusion, our results suggest sHLA-DR as a potent prognostic serum marker in melanoma patients superior to S100-beta in helping to identify early stage patients at high risk of disease progression. PMID- 12124809 TI - Progenipoietin-generated dendritic cells exhibit anti-tumor efficacy in a therapeutic murine tumor model. AB - Progenipoietin (ProGP-4) is a chimeric molecule, exhibiting both Flt-3 and granulocyte-colony stimulating factor (G-CSF) receptor agonist activities. Subcutaneous administration of ProGP-4 to BALB/c mice at a dose of 40-100 microg/day for up to 12 consecutive days induces both CD11c(+) dendritic cells (DCs) and CD11c(-)/CD11b(+) granulocytes in spleen, blood and lymph nodes of treated animals. Peak numbers of all cell populations were observed on day 7 of treatment, with CD11c(+) DCs representing approximately 8% of total splenocytes at that time. Approximately 40-50% of these CD11c(+) cells were also able to endocytose and process the exogenous fluorescent antigen DQ-BSA. As a test of their therapeutic utility, freshly prepared CD11c(+) DCs were pulsed with a defined tumor-associated peptide epitope (murine p53(232-240)) and injected as a vaccine into BALB/c mice bearing day 7 established CMS4 sarcomas. Similarly prepared DCs were injected again 1 week later. Based on our results, we conclude that (i) both peptide-pulsed CD11c(+) DCs (harvested directly from ProGP-4 treated mice) and pulsed bone marrow-derived DCs effectively slow the growth of or mediate the regression (in 25 of 89 [28%] cases) of CMS4 tumors, and (ii) nonpulsed DCs mediated minimal or no therapeutic effect. These data support the ability of ProGP-4 to enhance the peripheral frequencies of DCs that exhibit therapeutic efficacy when applied as a vaccine to treat tumor-bearing animals. PMID- 12124810 TI - Application of differential display to identify genes for lung cancer detection in peripheral blood. AB - A blood assay for detection of lung cancer biomarkers could significantly improve cancer patient prognosis and survival rates. Amplified fragment length polymorphism-differential display (AFLP-DD) was used to identify gene transcripts found in lung cancer tissue and the peripheral blood of lung cancer patients. The clones were evaluated for gene expression in lung cancer tissue, peripheral blood of lung cancer patients and healthy volunteers' blood. The isolated gene transcript clones were found to be from the syndecan 1 gene, collagen 1 gene and 2 novel genes. All 4 transcripts were expressed in normal lung tissue, 4 cultured primary lung cells and 6 lung cancer cell lines. RNA was isolated from peripheral blood samples of 69 lung cancer patients. Reverse transcriptase polymerase chain reaction (RT-PCR) was used to test for the presence of cytokeratin 19 and the 4 gene mRNA transcripts in blood RNA. The positive detection rate of at least 1 of the 5 transcripts was 79% for lung adenocarcinoma and 62% for squamous carcinoma. Using RT-PCR, at least 1 of the markers was found in 53% of stage I patients, 100% of stage II, 71% of stage III and 81% of stage IV lung cancer patients. Blood samples from 20 healthy volunteers were also tested, but only 1 of the 5 transcripts was found in 1 patient. These new molecular markers may aid early detection, staging and follow-up of lung cancer patients by RNA isolated from blood. PMID- 12124811 TI - Polymorphisms of DNA repair genes XRCC1 and XPD and their associations with risk of esophageal squamous cell carcinoma in a Chinese population. AB - Esophageal squamous cell carcinoma (ESCC), which is prevalent in China, is believed to be induced by environmental carcinogens. Accumulating evidence has shown that individual variation in DNA repair capacity resulting from genetic polymorphism influences risk of environmental carcinogenesis. We therefore investigated the associations between genetic polymorphisms in the DNA repair genes XRCC1 (Arg194Trp and Arg399Gln) and XPD (Asp312Asn and Lys751Gln) and risk of ESCC in an at-risk Chinese population. Genotypes were determined by a PCR based approach in 433 patients with ESCC and 524 frequency-matched normal controls. We found that individuals with Trp/Trp genotype at XRCC1 Arg194Trp site had a 2-fold increased risk of this disease compared to Arg/Arg genotype (adjusted OR = 1.98; 95% CI 1.26-3.12). Furthermore, when compared to Arg/Arg and Arg/Trp genotype combined, homozygote for Trp/Trp genotype significantly increased the risk of developing ESCC, with the adjusted OR being 2.07 (95% CI 1.34-3.20). However, the XRCC1 Arg399Gln polymorphism was not significantly associated with risk of ESCC, with the adjusted OR being 0.87 (95% CI 0.55-1.37). Neither Asp312Asn nor Lys751Gln polymorphisms in the XPD gene influenced risk of ESCC in our study. These findings suggest that DNA repair gene XRCC1 but not XPD might play a role in esophageal carcinogenesis and might represent a genetic determinant in the development of the cancer. PMID- 12124812 TI - Prevention of chemotherapy-related toxic side effects by infection with adeno associated virus type 2. AB - Drug resistance and toxic side effects are major limiting factors in the clinical use of antineoplastic chemotherapy. Patients with pancreatic cancer generally do not benefit from chemotherapy. The nonpathogenic adeno-associated virus type 2 (AAV-2) has been shown to sensitize human tumor cells to gamma irradiation and chemotherapeutic drugs. In the present study, we characterized the therapeutic role of AAV-2 infection in combination with 5-fluorouracil (5-FU)-based chemotherapy on pancreatic cancer cells in an animal model. In Lewis rats bearing s.c. implants of syngeneic DSL6A pancreatic cancer cells, intratumoral infection with AAV-2 (MOI 10E8 i.u.) in combination with 5-FU (5 or 50 mg/kg body weight) resulted in significantly reduced tumor growth and prolonged survival time compared with 5-FU single therapy. Most surprisingly, AAV-2-infected rats remained in a much better physical condition compared to their noninfected counterparts. While rats treated with 5-FU single therapy lost weight, were sluggish and died within 4 months after tumor implantation, animals infected with AAV showed much better vigilance, with body weight, leukocyte number and hemoglobin levels similar to healthy rats. In particular, 5-FU-related side effects like thrombocytopenia and leukopenia were significantly reduced in animals treated with the combination regimen. By in vitro analysis, human (Capan 1 and DANG) pancreatic cancer cell lines were shown to be sensitized to 5-FU chemotherapy to an extent similar to DSL6A cells. AAV-2 infection enhanced 5-FU induced apoptosis by a factor of 8 to 14 in both human and rat pancreatic cancer cell lines. The data suggest that infection with the nonpathogenic AAV-2 significantly improves both chemotherapy efficacy and physical appearance and offers a novel strategy in cancer treatment. PMID- 12124813 TI - Effect of O6-(4-bromothenyl)guanine on different temozolomide schedules in a human melanoma xenograft model. AB - The DNA repair protein O(6)-alkylguanine DNA alkyltransferase (ATase) is a major component of resistance to treatment with methylating agents and nitrosoureas. Inactivation of the protein, via the administration of pseudosubstrates, prior to chemotherapy has been shown to improve the latter's therapeutic index in animal models of human tumours. We have also shown that rational scheduling of temozolomide, so that drug is administered at the ATase nadir after the preceding dose, increases tumour growth delay in these models. We now report the results of combining these two approaches. Nude mice bearing A375M human melanoma xenografts were treated with vehicle or 100 mg/kg temozolomide ip for 5 doses spaced 4, 12 or 24 hr apart. Each dose was preceded by the injection of vehicle or 20 mg/kg 4BTG. All treatments resulted in significant delays in tumour quintupling time compared with controls: by 6.2, 5.9 and 16.8 days, respectively, for 24-, 12- and 4-hourly temozolomide alone and by 22.3, 21.3 and 22.1 days, respectively, in combination with 4BTG. Weight loss due to TMZ was unaffected by the presence of 4BTG. This was of the order of 6.2-10.6% with 24- and 12-hourly administration and 17.4-20.1% (p < 0.0001) with 4-hourly treatment. In our model, combining daily temozolomide with 4-BTG confers increased antitumour activity equivalent to that achieved by compressing the temozolomide schedule but with less toxicity. Using temozolomide schedule compression with 4-BTG does not improve on this result, suggesting that ATase inactivation with pseudosubstrates is a more promising means of enhancing the activity of temozolomide than compressed scheduling. PMID- 12124814 TI - Clustering of cancer-related mutations in a subset of BRCA1 alleles: a study in the Spanish population. AB - We have observed that the frequency of D17S855 short alleles (139 bp and 141 bp) in individuals carrying BRCA1 germline mutations is higher than in controls (54% vs. 31%, p = 0.0004). By unambiguously establishing mutation/D17S855 phase in 18 BRCA1-positive families, we find that most (11 of 15 different mutations) BRCA1 defects are linked to chromosomes with short alleles (OR = 8.21, 95% CI 1.97 39.32, p = 0.0007). We suggest that BRCA1 mutations are not randomly distributed but clustered in a subset of BRCA1 alleles that can be identified by D17S855 genotyping. Further analysis involving a larger set of mutations and different populations are needed to clarify the relevance of this unexpected finding. PMID- 12124816 TI - Monotherapy versus combined androgen blockade in patients with advanced prostate cancer. PMID- 12124817 TI - Radical prostatectomy versus radiation therapy for clinically localized prostate carcinoma: the butcher and the baker selling their wares. PMID- 12124818 TI - Radical prostatectomy or external beam radiotherapy: one step forward or two steps back? PMID- 12124819 TI - Factors associated with interval adherence to mammography screening in a population-based sample of New Hampshire women. AB - BACKGROUND: Interval adherence to mammography screening continues to be lower than experts advise. The authors evaluated, using a population-based mammography registry, factors associated with adherence to recommended mammography screening intervals. METHODS: The authors identified and recruited 625 women aged 50 years and older who did and did not adhere to interval mammography screening. Demographic and risk characteristics were ascertained from the registry and were supplemented with responses on a mailed survey to assess knowledge, perceived risk, anxiety regarding breast carcinoma and its detection, and women's experiences with mammography. RESULTS: The authors found no differences in risk factors or psychologic profiles between adhering and nonadhering women. Women who did not adhere had a statistically higher body mass index than women who did adhere (27.6 versus 26.1, P = 0.003). Exploration of mammographic experiences by group found that care taken by technologists in performing or talking women through the exam was higher in adhering women than nonadhering women (75.6% vs 65.71% for performing the exam, and 71.6% vs 60.8% for talking patients through the exam, respectively, P < 0.05). CONCLUSIONS: The authors found that previous negative mammographic experiences, particularly those involving mammography technologists, appear to influence interval adherence to screening and that patient body size may be an important factor in this negative experience. PMID- 12124820 TI - Randomized trial comparing cyclophosphamide, methotrexate, and 5-fluorouracil (CMF) with rotational CMF, epirubicin and vincristine as primary chemotherapy in operable breast carcinoma. AB - BACKGROUND: According to the overview of Early Breast Cancer Trialists' Collaborative Group, anthracycline containing regimens are superior to cyclophosphamide, methotrexate, and 5-fluorouracil (CMF) as adjuvant chemotherapy for breast carcinoma, but no comparative information is available in terms of primary chemotherapy. In the current randomized controlled trial, the authors compared CMF with a chemotherapy regimen including CMF, epirubicin, and vincristine (CMFEV). METHODS: Two hundred eleven patients with Stages I and II palpable breast carcinoma and tumor diameter > 2.5 cm or < or = 2.5 cm with cytologically proven axillary lymph node involvement were randomized to receive CMF (arm A) or CMFEV regimen (arm B) for four cycles before surgery. After surgery, patients in both arms received adjuvant CMF for three cycles; the postmenopausal patients also received tamoxifen for two years. RESULTS: There were no significant differences in the complete response (CR) and in the CR plus partial response (PR) rates between the two arms. In the subset analysis, among premenopausal patients, significantly higher rates of CR (26% vs 4%, P = 0.004) and of CR + PR rates (80% vs 54%, P = 0.007) were observed in the CMFEV, as compared to the CMF arm. Multivariate analysis confirmed the presence of a significant interaction between menopausal status and type of treatment on the probability of achieving CR (P = 0.02) or CR + PR (P = 0.01). There were no major differences in the side effects of the two treatments, with the exception of more frequent alopecia in the experimental arm. CONCLUSIONS: The results of the current study are in line with those of previous published randomized clinical trials comparing regimens without and with anthracycline as adjuvant treatment, indicating an agreement between the short term response to primary chemotherapy and the long term results observed in the adjuvant setting. PMID- 12124821 TI - Prophylactic bilateral mastectomy: patterns of practice. AB - BACKGROUND: Many women who are at an elevated risk of developing breast carcinoma choose prophylactic mastectomy to decrease their risk. We conducted a population based study to review the indications for, and patterns of practice of prophylactic mastectomy in Ontario, Canada, since 1991. METHODS: A medical chart review was conducted at 33 hospitals that were identified as having conducted at least one prophylactic mastectomy. All bilateral mastectomy patients with no diagnosis of invasive or in situ breast carcinoma were eligible. RESULTS: The number of prophylactic bilateral mastectomies performed varied from 6 to 19. The mean age of women undergoing prophylactic mastectomy was 43.5 years. Eighty percent of the women had prophylactic mastectomy performed because of a family history of breast carcinoma (89 of 99) or because of a known BRCA1 or BRCA2 mutation (10 of 99). Twenty percent of the women had no family history, but had the surgery for other benign breast conditions. Women with a family history of breast carcinoma were much more likely to have a total mastectomy (89%) than a subcutaneous mastectomy (11%). Sixty percent of the women had reconstructive surgery following mastectomy. CONCLUSIONS: Prophylactic mastectomy is not performed on a large scale. The introduction of genetic testing for BRCA1 and BRCA2 has the potential to change the patterns of practice for prophylactic mastectomy. PMID- 12124822 TI - Treatment of intracranial metastatic esthesioneuroblastoma. AB - BACKGROUND: Esthesioneuroblastoma (ENB) is an uncommon tumor that may metastasize to the central nervous system (CNS). To the authors' knowledge there is no current consensus regarding treatment. The current study was conducted to determine the toxicity and response rate of combined modality therapy in the treatment of patients with ENB metastatic to the brain. METHODS: Six patients (2 men and 4 women) ranging in age from 47-61 years (median age, 53.5 years) with ENB had clinical and neuroradiographic evidence of CNS metastases (brain parenchyma in 6 patients and leptomeningeal metastases in 3 patients). CNS directed therapy included radiotherapy (to the brain in three patients and to the spine in two patients) and chemotherapy (systemic in six patients and regional in three patients). Systemic chemotherapy was comprised of carboplatin, lomustine, and vincristine administered every 2 months. Patients were evaluated by neuroradiographic and neurologic examination every other month. RESULTS: Between 1-6 cycles of systemic chemotherapy were administered (median, 4.5 cycles). and 6 19 cycles of regional chemotherapy were administered (median, 17 cycles). Toxicity included aseptic meningitis (three patients), radiation enteritis (one patient), and > or = Grade 3 (according to the National Cancer Institute Common toxicity Criteria) myelosuppression (four patients). Two patients required hospitalization for neutropenic fever and two patients required a transfusion (platelets in two patients and red blood cells in one patient). No treatment related deaths were reported. A partial response was achieved in four patients, and two patients demonstrated progressive disease. The median duration of response was 9 months (range, 2-12 months). Overall survival ranged from 3-13 months (median, 10.5 months). The cause of death was progressive parenchymal brain disease in three patients, systemic disease in two patients, and leptomeningeal disease in one patient. CONCLUSIONS: In the small cohort of patients in the current study, combined modality therapy was found to have modest toxicity and palliative efficacy. PMID- 12124823 TI - Influence of p53 mutations on prognosis of patients with glioblastoma. AB - BACKGROUND: The influence of p53 mutations on the biology of astrocytic tumors is controversial. p53 is thought to be inactivated in the early stage of gliomagenesis; however, what role its inactivation plays in the malignancy of gliomas remains unknown. To understand the significance of p53 inactivation, the authors identified the locus of p53 gene mutation in glioma samples at different stages of progression and studied the correlation between the mutation and clinical behavior. METHODS: Samples from newly diagnosed gliomas, including pure and mixed astrocytomas, were analyzed for p53 mutations using a yeast functional assay. To determine the locus of the gene mutations, DNA sequencing was performed. RESULTS: The incidence of p53 mutations was higher in anaplastic astrocytomas (AA, 48%) than glioblastomas (GBM, 31%). There was no significant difference in the average ages of GBM patients with and without p53 mutations (54.9 years +/- 2.3 and 53.2 years +/- 4.6, respectively). In GBM patients, the mutation did not affect progression free survival or overall survival. Astrocytomas and GBM differed in the distribution of p53 mutation loci. CONCLUSIONS: The p53 gene mutation does not markedly affect the survival of GBM patients. The difference in the location of p53 mutations between AA and GBM suggests that in gliomas, the p53 mutation may contribute not only to tumorigenesis (as an early event) but also to progression to malignancy (as a late event). PMID- 12124824 TI - Clinicopathologic study of 123 cases of prolactin-secreting pituitary adenomas with special reference to multihormone production and clonality of the adenomas. AB - BACKGROUND: Prolactinoma is the most invasive type of pituitary adenoma and is generally believed to be well-differentiated adenoma and to produce only prolactin (PRL). The factors related to the various biologic behaviors occurring in patients of different ages and sexes await clarification. Since different immunophenotypes of adenoma may show different biologic behaviors and responses to medical agents, the authors examined hormone production and tried to clarify the clonality of plurihormonal prolactinoma. METHODS: Clinicopathologic factors were studied in 123 patients with prolactinomas (40 males and 83 females). The specimens were fixed in either 10% neutral buffered formalin or 70% alcohol and used for light microscopy. Alcohol-fixed tissue was used to extract DNA from 26 samples obtained from female patients for human androgen receptor gene (HUMARA) assay. RESULTS: Sixty one cases (50%) were pure prolactinoma and 62 cases (50%) were plurihormonal prolactinoma. Spearman rank correlation analysis revealed a significant relationship between age and serum PRL level (P = 0.0002), age and tumor volume (P < 0.0001), and tumor volume and serum PRL level (P < 0.0001). Multiple regression analysis showed a significant correlation only between tumor volume and serum PRL level. The Mann-Whitney U test revealed that prolactinomas associated with higher PRL levels, larger adenomas, and higher ages were significantly more invasive to the cavernous sinus and that male patients had significantly higher PRL levels and larger adenomas. The HUMARA assay disclosed that 11 of 13 plurihormonal prolactinomas (85%) were compatible with monoclonal origin. CONCLUSIONS: The current results suggest that not only can various hormones other than PRL be secreted by prolactinoma, but also that most multihormone-producing prolactinomas are monoclonal in origin. PMID- 12124825 TI - High serum YKL-40 level after surgery for colorectal carcinoma is related to short survival. AB - BACKGROUND: YKL-40 is a member of family 18 glycosyl hydrolases. YKL-40 is a growth factor and may stimulate migration of endothelial cells. YKL-40 may also play a role in inflammation and degradation of connective tissue. Elevated preoperative serum YKL-40 levels in patients with colorectal carcinoma are associated with a significantly poorer prognosis compared to patients with normal serum YKL-40. In the current study the authors evaluated the value of serum YKL 40 in monitoring patients with colorectal carcinoma. METHODS: YKL-40 was determined by an in-house radioimmunoassay method in serum obtained pre- and postoperatively from 324 patients who underwent curative resection (Dukes Stage A: 47; B: 148; C: 119; and D: 10). The patients were followed with serum YKL-40 levels every 6 months postoperatively, and the median followup time was 82 months (range, 68-95). In that period 146 patients died. RESULTS: Serum YKL-40 was significantly decreased in the first postoperative blood sample in 62% of patients with high preoperative levels. In addition, patients with high serum YKL 40 (adjusted for age) six months after curative operation had significantly shorter survival times (P = 0.0002) and shorter relapse free intervals (P = 0.004) than patients with normal postoperative serum YKL-40. This result was independent of simultaneous serum carcinoembryonic antigen levels at six months. Analysis of survival by scoring serum YKL-40 as a time-dependent covariate in a Cox regression analysis showed that patients exhibiting elevated serum YKL-40 had an increased hazard for death within the following six months compared to those patients with normal serum YKL-40 level (hazard ratio [HR] = 9.6, 95% confidence interval [CI]: 6.0-15.5, P < 0.0001). Multivariate analysis including Dukes stage, age, gender, and tumor location as well as the time-dependent serum YKL-40 showed that high serum YKL-40 was an independent prognostic variable of survival (HR = 8.5, 95% CI: 5.3-13.7, P < 0.0001). CONCLUSIONS: These results suggest that determination of serum YKL-40 during followup of patients operated on for colorectal carcinoma might be useful for monitoring curatively resected patients. PMID- 12124826 TI - Initial decline in hemoglobin during neoadjuvant hormonal therapy predicts for early prostate specific antigen failure following radiation and hormonal therapy for patients with intermediate and high-risk prostate cancer. AB - BACKGROUND: Declines in serum hemoglobin (Hgb) levels occur from the use of androgen suppression therapy (AST) in the treatment of prostate cancer patients. We studied whether time to prostate specific antigen (PSA) failure following external beam radiation therapy (RT) and AST could be predicted by the rate of decline in the Hgb level following the administration of neoadjuvant AST or by the Hgb level at presentation or at the start of RT. METHODS: The study cohort comprised 110 intermediate or high-risk prostate cancer patients who were managed using three-dimensional conformal RT (70 Gy) and 6 months of AST (2 months neoadjuvant, concurrent, and adjuvant). A Cox regression multivariable analysis was performed to evaluate the ability of the rate of decline of the Hgb from baseline to the start of RT, baseline PSA level, Gleason score, percent positive biopsies, and T-category to predict time to PSA failure. RESULTS: A decline in the Hgb level of 1 g/dL or more during the first month of AST was the only significant predictor of time to PSA failure (P = 0.02) on multivariable analysis. The relative risk of PSA failure (95% confidence interval) for patients with a decline in Hgb level during the first month (> or = 1 g/dL vs. < 1 g/dL) was 6.3 (2.4, 8.3) and the 3-year estimate of PSA outcome was 66% versus 82% (P = 0.04), respectively. There were no imbalances in the pretreatment prognostic factors or length of follow-up in each of these groups. CONCLUSION: A decline of 1 g/dL or more in Hgb level during the first month of neoadjuvant AST was a predictor of early PSA failure following RT and AST in intermediate and high-risk prostate cancer patients. PMID- 12124827 TI - Biochemical outcome after radical prostatectomy or external beam radiation therapy for patients with clinically localized prostate carcinoma in the prostate specific antigen era. AB - BACKGROUND: To the authors' knowledge, consensus is lacking regarding the relative long-term efficacy of radical prostatectomy (RP) versus conventional dose external beam radiation therapy (RT) in the treatment of patients with clinically localized prostate carcinoma. METHODS: A retrospective cohort study of 2635 men treated with RP (n = 2254) or conventional-dose RT (n = 381) between 1988-2000 was performed. The primary endpoint was prostate specific antigen (PSA) survival stratified by treatment received and high-risk, intermediate-risk, or low-risk group based on the serum PSA level, biopsy Gleason score, 1992 American Joint Commission on Cancer clinical tumor category, and percent positive prostate biopsies. RESULTS: Estimates of 8-year PSA survival (95% confidence interval [95% CI]) for low-risk patients (T1c,T2a, a PSA level < or = 10 ng/mL, and a Gleason score < or = 6) were 88% (95% CI, 85, 90) versus 78% (95% CI, 72, 83) for RP versus patients treated with RT, respectively. Eight-year estimates of PSA survival also favored RP for intermediate-risk patients (T2b or Gleason score 7 or a PSA level > 10 and < or = 20 ng/mL) with < 34% positive prostate biopsies, being 79% (95% CI, 73, 85) versus 65% (95% CI, 58, 72), respectively. Estimates of PSA survival in high-risk (T2c or PSA level > 20 ng/mL or Gleason score > or = 8) and intermediate-risk patients with at least 34% positive prostate biopsies initially favored RT, but were not significantly different after 8 years. CONCLUSIONS: Intermediate-risk and low-risk patients with a low biopsy tumor volume who were treated with RP appeared to fare significantly better compared with patients who were treated using conventional-dose RT. Intermediate-risk and high-risk patients with a high biopsy tumor volume who were treated with RP or RT had long-term estimates of PSA survival that were not found to be significantly different. PMID- 12124828 TI - The impact of androgen deprivation on quality of life after radical prostatectomy for prostate carcinoma. AB - BACKGROUND: Androgen deprivation is commonly prescribed for men with a rising prostate specific antigen level after radical prostatectomy, despite scant evidence regarding its efficacy and side effects. In the current study, the authors compared measures of health-related quality of life (HRQOL) in men who were treated with androgen deprivation after radical prostatectomy with those for men who underwent surgery but were not treated with androgen deprivation. METHODS: Medicare Provider and Analysis and Review (MedPAR) files were used to identify men who had undergone radical prostatectomies between 1991-1992. Medicare Part B data then were used to select two samples: men who subsequently were androgen deprived and those who were not. In 1999, a mail survey was administered that addressed a range of disease-related and treatment-related issues, including HRQOL. Age-adjusted comparisons of responses to seven multiitem measures of HRQOL were performed. RESULTS: The overall response rate was 82%. On all seven HRQOL measures (impact of cancer and treatment, concern regarding body image, mental health, general health, activity, worries about cancer and dying, and energy), there were statistically significant decrements associated with androgen deprivation. CONCLUSIONS: Patients and physicians must weigh the price patients pay with regard to HRQOL against the uncertain benefits of early androgen deprivation. PMID- 12124829 TI - Fas and Fas ligand expression is elevated in prostatic intraepithelial neoplasia and prostatic adenocarcinoma. AB - BACKGROUND: Fas is a Type I membrane receptor of the tumor necrosis factor/nerve growth factor family. On binding to Fas ligand, a Type II transmembrane protein, the Fas/Fas ligand complex, induces apoptosis in target cells. Dysregulation of Fas and Fas ligand expression has been found in some malignant neoplasms. METHODS: Using immunohistochemical analysis, the authors studied the expression of Fas and Fas ligand in prostatic adenocarcinoma, high-grade prostatic intraepithelial neoplasia (PIN), and adjacent benign prostate tissue from 95 radical prostatectomy specimens. RESULTS: The percentage of cells that stained positively with Fas in benign prostate tissue (mean, 2%) was statistically significantly lower compared with that in prostatic intraepithelial neoplasia (mean, 13%; P = 0.0014) and prostatic adenocarcinoma (mean, 31%; P = 0.0001). The staining intensity of Fas was significantly less in benign prostate tissue compared with the staining intensity in PIN and prostatic adenocarcinoma. The percentage of cells that stained positively with Fas ligand in benign prostate tissue (mean, 13%) was statistically significantly lower compared with that in PIN (mean, 47%; P = 0.0001) and in prostatic adenocarcinoma (mean, 53%; P = 0.0001). The staining intensity of Fas ligand was significantly less in benign prostate tissue compared with that in PIN and prostatic adenocarcinoma. CONCLUSIONS: Data from the current study indicate that Fas/Fas ligand is elevated in prostatic malignancy, suggesting that Fas-mediated apoptosis may be a potential target for therapeutic intervention. PMID- 12124831 TI - Salvage high-dose chemotherapy in patients with germ cell tumors: an Italian experience with 84 patients. AB - BACKGROUND: High-dose chemotherapy (HDCT) followed by hematopoietic stem cell support (HSCS) potentially may be curative in patients with germ cell tumor (GCT) who develop recurrent tumors or who have an inadequate response after receiving standard-dose chemotherapy. The authors report their experience with HDCT as salvage therapy for patients with GCT. METHODS: Between 1986 and 2000, 84 patients with GCT, with a median age 29 years (range, 15-50 years), were treated with 105 courses of HDCT with HSCS. Patients were stratified into good, intermediate, and poor risk categories according to a validated prognostic index. RESULTS: Overall, 28 patients (33%) have been continuously disease free. In the good risk group, 24 patients (69%) have been continuously disease free compared with 4 patients (13%) in the intermediate risk group (P < 0.001) and 0 patients in the poor risk group (P < 0.001). Treatment-related mortality occurred only among patients in the poor risk (n = 6 patients) and the intermediate risk groups (n = 4 patients). CONCLUSIONS: In the authors' experience, HDCT induced impressive long-term remissions as salvage treatment among patients in the good risk group. Moreover, the use of validated prognostic classifications may contribute to a better definition of the role of HDCT other than improving the outcome of patients with GCT. The definitive statement on the possible role of HDCT in patients with GCT will derive from the ongoing Phase III randomized studies. PMID- 12124830 TI - Absence of c-KIT and members of the epidermal growth factor receptor family in refractory germ cell cancer. AB - BACKGROUND: Germ cell tumors (GCTs) in adolescent and young males are very sensitive to cisplatin-based chemotherapy. However, 10-20% of the patients cannot be cured by currently available therapeutic options. Once a tumor does not respond to cisplatin, current therapeutic modalities offer only a chance for short palliation. Recently, new treatment options that interfere with various receptor tyrosine kinases, including c-KIT and members of the epidermal growth factor receptor (EGFR) family, have been used successfully in chemotherapy resistant tumors overexpressing c-KIT, ERB-B2, or EGFR. METHODS: We studied the presence of c-KIT and the four members of the EGFR family by immunohistochemistry, as well as by ERB-B2 gene amplification using fluorescent in situ hybridization, in a series of 22 patients with cisplatin-resistant GCTs in search of new treatment targets. The results in these refractory tumors were compared with those of 12 patients with chemosensitive GCTs diagnosed in an advanced metastatic stage. RESULTS: The data obtained in both groups did not differ in any of the investigated biologic markers. c-KIT was detected in the one case of pure seminoma studied and in the seminomatous components of combined tumors. The presence of EGFR was restricted to trophoblastic giant cells and the syncytiotrophoblastic elements of four nonseminomas including one pure choriocarcinoma and to a secondary non-germ cell malignancy, which had developed most likely from a mature teratoma. ERB-B2 was moderately positive in the secondary non-germ cell malignancy, in one mature teratoma component of a mixed nonseminoma, and together with EGFR in the syncytiotrophoblastic cells of a pure choriocarcinoma. Of all samples investigated, this latter case was the only one showing an amplification of the ERB-B2 gene in the syncytiotrophoblasts. ERB-B3 and ERB-B4 were detected rarely. CONCLUSION: The majority of refractory GCTs do not qualify for treatment with new biologic agents targeting the receptor tyrosine kinases EGFR, ERB-B2, or c-KIT. The lack of differences between the tumors of refractory and the responsive patients indicates that overexpression of any of these receptor tyrosine kinases does not contribute to a resistant phenotype in GCTs. PMID- 12124832 TI - The significance of the amount of myometrial invasion in patients with Stage IB endometrial carcinoma. AB - BACKGROUND: The 1988 International Federation of Gynecology and Obstetrics (FIGO) staging system for endometrial carcinoma defined Stage IB as disease with invasion of less than one-half of the myometrium, although most of the data on prognostic factors are based on invasion of the inner one-third, middle one third, or outer one-third of the myometrium. The objective of this study was to determine whether the depth of myometrial invasion is correlated with outcome in patients with Stage IB endometrial carcinoma. METHODS: Between November, 1987 and June, 1998, 251 patients with Stage IB endometrioid adenocarcinoma of the uterus underwent simple hysterectomy followed by intravaginal brachytherapy. The depth of myometrial invasion was less than or equal to one-third (Group I) in 191 of 251 patients (79%) and greater than one-third to less than one-half (Group II) in 52 of 251 patients (21%). Comprehensive surgical staging (CSS) was done in 12% of patients. The two groups were balanced with regard to age (< 60 years vs. > or = 60 years), FIGO grade, lower uterine segment involvement (LUSI), CSS, and the use of postoperative external-beam radiation. The rate of capillary-like space invasion (CLSI), however, was 9% in Group I compared with 25% in Group II (P = 0.002). The median follow-up was 58 months. RESULTS: The overall 5-year actuarial local-regional control (LRC), disease free (DFS) survival, and overall survival (OS) rates were 95%, 92%, and 92%, respectively. These end points, however, did not vary significantly between the two groups. The 5-year LRC, DFS, and OS rates in Groups I and II were 96% versus 95%, respectively (P = 0.9); 92% versus 94%, respectively (P = 0.7); and 92% versus 90%, respectively (P = 0.5). On multivariate analysis, the influence on outcome of age, grade, amount of myometrial invasion, LUSI, and CLSI was evaluated. Only age > or = 60 years and FIGO Grade 3 were correlated with poor DFS (P = 0.02 and P = 0.03, respectively) and OS (P = 0.001 and P = 0.01, respectively). CONCLUSIONS: Based on this study, in patients with Stage IB endometrial carcinoma, the amount of myometrial invasion, defined as invasion less than or equal to one-third of the myometrium versus invasion greater than one-third and less than one-half of the myometrium, did not appear to influence outcome. Age > or = 60 years and FIGO Grade 3, however, emerged as independent prognostic factors for poor DFS and OS. PMID- 12124833 TI - Phase II study of induction chemotherapy with paclitaxel, ifosfamide, and carboplatin (TIC) for patients with locally advanced squamous cell carcinoma of the head and neck. AB - BACKGROUND: This Phase II trial was conducted to determine the response rate, particularly of the primary sites, tolerability, and toxicity of induction chemotherapy of paclitaxel, ifosfamide, and carboplatin for patients with previously untreated locally advanced squamous cell carcinoma of the head and neck (SCCHN). We also hypothesized that improved complete response (CR) rates with the induction chemotherapy may render better survival rates with subsequently delivered definitive local treatment. METHODS: All eligible patients with locally advanced SCCHN received two courses of induction chemotherapy and underwent repeated head and neck examination and computed tomography or magnetic resonance imaging scans. If the patients achieved responses (CR or partial [PR]), they received two more courses of chemotherapy before undergoing definitive local treatment. Induction chemotherapy consisted of paclitaxel (T; 175 mg/m(2) in a 3 hour infusion) on Day 1, ifosfamide (I; 1000 mg/m(2) in a 2-hour infusion) on Days 1-3 with intravenous mesna (200 mg/m(2) before and 400 mg/m(2) after ifosfamide), and carboplatin (C) using the Calvert formula for the area under the plasma concentration-versus-time curve of 6 on Day 1, repeated every 3-4 weeks. Prophylactic hematopoietic growth factors or antibiotics were not used in this study. Definitive local treatment was given based on the investigators' preference. RESULTS: Fifty-four patients were registered and 52 patients were assessable for response to induction chemotherapy; 2 were not evaluable. After four courses of induction chemotherapy, the CR rates of the primary and lymph node sites were 60%, and 41%, respectively. For both primary and lymph node sites, there were 31% CRs and 50% PRs with an overall response rate of 81%. Five (9%) patients developed neutropenic fever, all of whom recovered with antibiotic therapy. Two (4%) patients had infection without neutropenia and recovered without any complication. Grade 3/4 thrombocytopenia and anemia occurred in three (6%) and four (7%) patients, respectively. Grade 3/4 fatigue developed in four (7%), arthralgia/myalgia in two (4%), peripheral neuropathy in two (4%), and orthostatic hypotension in two (4%) patients. One patient died of severe anaphylaxis although a maximized resuscitation effort was made. With a median follow-up of 22 months, the organ preservation rate was about 81% (42 of 52 patients). Although survival rates were not primary end points in this study, with a median follow-up of 22 months, 43 (83%) patients are still alive. Overall 1 and 2-year survival rates were 88% and 82%, respectively. Disease-free 1 and 2 year survival rates were 88% and 77% respectively. CONCLUSIONS: TIC induction chemotherapy is associated with a high CR rate at the primary sites and with excellent survival and organ preservations rates with subsequently delivered definitive local therapy. The regimen was also well tolerated in the majority of patients. The TIC regimen should be developed further in the context of induction chemotherapy followed by concomitant chemoradiotherapy or with specific molecular targeted agents. PMID- 12124834 TI - Impact of aging on the development of hepatocellular carcinoma in patients with posttransfusion chronic hepatitis C. AB - BACKGROUND: Hepatitis C virus (HCV) infection is a heterogeneous disease, the natural history of which remains controversial. There is solid evidence that chronic HCV infection is responsible for the occurrence of hepatocellular carcinoma (HCC). The aim of the current cohort study was to determine the rate of the development of HCC from the time of primary HCV infection and to assess the risk factors for the development of HCC in chronic posttransfusion hepatitis C patients. METHODS: Four hundred sixty-nine patients with clinically compensated HCV, who had undergone a single blood transfusion comprised the current study cohort. Patients with other risk factors for chronic liver disease were excluded. All patients were referred to the liver center at the National Nagasaki Medical Center between December 1980 and December 1998 and were followed prospectively until the end of the analysis (June 2000). RESULTS: Follow-up data were obtained for 445 patients. The mean duration from HCV infection to the end of the observation was 28 years. Fifty-two patients (11.1%) progressed to HCC. The mean duration from the time of blood transfusion to the diagnosis of HCC was 31 years. Multivariate Cox regression analyses revealed age, fibrosis, duration from HCV infection to study entry, and alcohol consumption to be the independent factors affecting the development of HCC. The risk of developing HCC in patients age > or = 56 years was increased 7.8-fold compared with that in patients age < 56 years. The mean age of patients at the time of HCC diagnosis was 65 years (range, 58-79 years). CONCLUSIONS: At the time of diagnosis, 92% of the 52 HCC patients were age > 60 years and 38 of the HCC patients (73%) were in their 60s. There was a significantly negative correlation between the duration from HCV infection to the development of HCC and the age of the patient at the time of infection (correlation coefficient = 0.702; P < 0.0001; Y = 61.1-0.82X), indicating that the age of patients, rather than the duration of HCV infection, is more significant for HCC development in patients with posttransfusion HCV. Moreover, these data may contribute to the design of an optimal follow-up schedule for patients with posttransfusion HCV. PMID- 12124835 TI - The novel and effective nonplatinum, nontaxane combination of gemcitabine and vinorelbine in advanced nonsmall cell lung carcinoma: potential for decreased toxicity and combination with biological therapy. AB - BACKGROUND: Gemcitabine and vinorelbine are two of the most active third generation agents for the treatment of advanced nonsmall cell lung carcinoma (NSCLC). The authors conducted a formal Phase II trial to evaluate the efficacy of this combination in both untreated and previously treated patients with Stage IIIB (with pleural effusion) or Stage IV NSCLC. METHODS: A total of 78 patients were treated on the current Phase II trial of front-line or second/third-line therapy with gemcitabine and vinorelbine in NSCLC. Eligible patients manifested either untreated disease (n = 42) or had received at least one but not more than two prior chemotherapy regimens (n = 36). The median age was 57.5 years (range, 33-79) with 57 men (73%) and 21 women (27%). The median performance status was one (range, one to two). The initial eight patients (four untreated and four previously treated) were treated at a previously established maximum tolerated dose of vinorelbine (30 mg/m(2)) and gemcitabine (1000 mg/m(2)) on Days 1, 8, and 15, with significant myelosuppression seen in five out of eight patients requiring dose omission in the first cycle. The next 70 patients received a reduced dose of vinorelbine (25 mg/m(2)) followed by gemcitabine (900 mg/m(2)) on Days 1, 8, and 15. RESULTS: Seventy eight patients were treated. Fifteen (36%) of the 42 evaluable patients who received front-line therapy had objective responses and 14 (33%) had stable disease. In the patients with prior treatment, 6 (17%) of 36 patients had partial response and 18 patients (50%) had stable disease. Median survival time for the previously untreated patient group was 10.1 months, with a one year survival of 43% and a two year survival rate of 32%. For the previously treated patients, the median survival time was 8.5 months, with a one year survival rate of 30%. Toxic effects were notable for significant myelosuppression, with > or =Grade 3 granulocytopenia seen in 55% of the patients on the untreated arm and 67% of the patients on the previously treated arm. Additionally, 9.5% and 13.9% (untreated and previously treated), respectively, of these patients experienced Grades 3 and 4 thrombocytopenia at some point in their treatment. A full dose delivery analysis showed that this myelosuppression resulted in Course 1, Day 15 skipped doses (even at the reduced dose level) in 42% of previously untreated patients and 47% of pretreated patients. Other side effects seen at Grades 3 and 4 in previously untreated and treated patients included anemia (9.5% and 2.8%), asthenia (4.8% and 5.5%), infection (14.3% and 5.6%), pain (9.5% and 19.4%), and pulmonary complications (4.8% and 13.8%). CONCLUSIONS: Gemcitabine/vinorelbine is an active, well-tolerated combination in both front-line and second/third-line therapy for Stage IIIB/IV NSCLC. The response rate, median survival rate, and one year survival rate compare favorably with platinum-based regimens. The toxicity profile of the gemcitabine/vinorelbine combination was quite favorable, with minimal Grade 3 and 4 toxic effects aside from granulocytopenia, which resulted in numerous Day 15 skipped doses but no significant febrile neutropenia or infection. The combination of gemcitabine and vinorelbine could be a useful regimen in standard clinical practice and has the potential for efficient combination with biologic/targeted therapy. Multiple randomized trials of this combination versus platinum combinations are now ongoing [corrected]. PMID- 12124836 TI - Implications of lymphatic drainage to unusual sentinel lymph node sites in patients with primary cutaneous melanoma. AB - BACKGROUND: Sentinel lymphadenectomy reliably identifies the first site(s) of regional lymphatic drainage and, therefore, the most likely lymph nodes to contain occult metastasis in patients with primary cutaneous melanoma. Although in most patients lymphatic drainage from the primary melanoma first reaches a standard lymph node basin, a sentinel lymph node (SLN) may be identified in an unusual location. The objective of this study was to determine the frequency and significance of unusual sentinel lymph node drainage patterns in a large cohort of patients with primary melanoma. METHODS: The records of 1145 consecutive primary melanoma patients who underwent SLN biopsy were reviewed. Preoperative lymphoscintigraphy was performed in all patients with truncal melanoma and in many patients with distal extremity lesions. Unusual lymph node sites were defined as epitrochlear, popliteal, or ectopic/interval (in-transit or any other nonstandard lymph node-bearing area). RESULTS: At least one SLN was harvested in 1117 patients (98%). SLN biopsy of an unusual lymph node site was attempted in 59 patients (5%). Successful intraoperative localization and biopsy was performed in 54 (92%) of 59 patients for a total of 56 unusual sites. Of these, 7 (13%) were popliteal, 8 (14%) were epitrochlear, and 41 (73%) were ectopic/interval. Preoperative lymphoscintigraphy was performed in 41 of these 54 patients and correctly identified unusual SLN locations in 12 (29%); the majority of unusual SLNs were identified only with the assistance of the intraoperative gamma probe. In four patients (7%), the unusual lymph node site was the only site from which SLNs were harvested. In the remaining 50 patients (93%), biopsies were performed on SLNs from both unusual sites and from a standard lymph node basin. Among the 54 patients who underwent a SLN biopsy of an unusual nodal site, 7 (13%) had lymph node metastases in that location. In four of the seven patients, the only positive SLN was from the unusual site. CONCLUSIONS: Sentinel lymphatic drainage patterns include lymph node-bearing areas that may be outside established standard lymph node basins and may represent the only site of regional lymph node metastases. Although preoperative lymphoscintigraphy may assist in the identification of unusual SLN drainage patterns, intraoperative use of the gamma probe is recommended to identify accurately and completely all sites of regional lymph node drainage. PMID- 12124837 TI - Systematic review and meta-analysis of monotherapy compared with combined androgen blockade for patients with advanced prostate carcinoma. AB - BACKGROUND: The current systematic review and meta-analysis compared monotherapy and combined androgen blockade in the treatment of men with advanced prostate carcinoma. Outcomes of interest included overall, cancer specific, and progression-free survival; time to treatment failure; adverse events; and quality of life. METHODS: The literature search identified randomized trials comparing monotherapy (orchiectomy and luteinizing hormone-releasing hormone [LHRH] agonists) with combination therapy using orchiectomy or a LHRH agonist plus a nonsteroidal or steroidal antiandrogen. Dual independent review occurred. The meta-analysis used a random effects model. RESULTS: Twenty-one trials compared survival after monotherapy with survival after combined androgen blockade (n = 6871 patients). The meta-analysis found no statistically significant difference in survival at 2 years between patients treated with combined androgen blockade and those treated with monotherapy (20 trials; hazard ratio [HR] = 0.970; 95% confidence interval [95% CI], 0.866-1.087). The authors determined a statistically significant difference in survival at 5 years that favored combined androgen blockade (10 trials; HR = 0.871; 95% CI, 0.805-0.942). For the subgroup of patients with a good prognosis, there was no statistically significant difference in survival. Adverse effects leading to withdrawal from therapy occurred more often with combined androgen blockade. To the authors' knowledge there is little evidence published to date comparing the effects of combined androgen blockade and monotherapy on quality of life, but the single randomized trial that adequately addressed this outcome reported an advantage for monotherapy over combined androgen blockade. CONCLUSIONS: A thorough examination of the usefulness of combined androgen blockade must balance the modest increase in expected survival observed at 5 years against the increased risk of adverse effects and the potential for adversely affecting the patient's overall quality of life. PMID- 12124838 TI - Adult rhabdomyosarcoma: outcome following multimodality treatment. AB - BACKGROUND: Childhood rhabdomyosarcoma (RMS) has a relatively good prognosis. Outcome for adults with this disease is poorly documented due to its rarity. METHODS: The clinicopathologic features, treatment methods, and disease outcome were reviewed retrospectively for 82 adults with locoregional RMS treated between 1960 and 1998. Patients with distant metastasis at diagnosis were excluded. Actuarial univariate and multivariate statistical methods were used to evaluate outcome. RESULTS: Patient ages ranged from 17 to 84 years (median, 27 years). Histologic subtypes were embryonal (34%), pleomorphic (43%), and alveolar (23%). Anatomic sites of origin were head and neck (52%), trunk (26%), and extremity (7%). Tumor size was 5 cm or smaller in 51% of patients. Regional lymph node metastasis was present in 33% of patients at presentation. Treatment consisted of radiation alone in 11%, radiation and surgery in 18%, radiation and chemotherapy in 34%, and all three modalities in 37%. With a median follow-up of 10.5 years, the 10-year actuarial disease-free and overall survival rates were 41% and 40%, respectively. The 10-year actuarial local, lymph node, and metastatic control rates were 75%, 82%, and 53%, respectively. The major determinant of metastatic control and survival was primary tumor size (< or = 5 vs. > 5 cm). Local control was satisfactory (10-year rate of 87%) for sites other than parameningeal (50% at 10 years). Patients whose disease responded to chemotherapy had a significantly better metastasis free period (72% at 10 years) than those whose disease failed to respond (19% at 10 years). CONCLUSIONS: Adult RMS is a highly malignant tumor with a significant incidence of metastatic recurrence. Continuing investigation of new and potentially more effective chemotherapy is crucial. Local control is satisfactory for sites other than parameningeal where new radiation technologies such as intensity-modulated therapy may be necessary to safely deliver adequate doses. PMID- 12124839 TI - Treating cancer with PEG Intron: pharmacokinetic profile and dosing guidelines for an improved interferon-alpha-2b formulation. AB - BACKGROUND: PEG Intron (pegylated interferon-alpha-2b [IFN-alpha-2b]; Schering Plough, Kenilworth, NJ) has demonstrated delayed clearance and increased area under the curve compared with native IFN-alpha-2b. Studies in patients with chronic hepatitis C infection and malignancies have demonstrated both biologic and clinical activity of PEG Intron and have provided empiric data to compare the pharmacokinetics (PK) and pharmacodynamics of PEG Intron and IFN-alpha-2b. METHODS: The authors conducted a review of the available data comparing the PK and pharmacodynamic effects of PEG Intron and IFN-alpha-2b. Safety and efficacy data from Phase I/II studies of PEG Intron in patients with chronic myelogenous leukemia (CML) and solid tumors were also reviewed. RESULTS: Data from patients with chronic hepatitis C infection suggest that exposure to IFN at a PEG Intron dose of 0.25 microg/kg per week is similar to that observed after administration of IFN-alpha-2b at a dose of 3 million International Units, three times per week. PEG Intron at doses up to 6 microg/kg per week was well tolerated and demonstrated clinical activity in patients with CML and solid tumors, including metastatic melanoma and renal cell carcinoma. CONCLUSIONS: Dose intensification can be achieved safely in patients with CML and solid tumors using PEG Intron, which could improve efficacy. These results provide useful dosing guidelines to clinicians investigating the antitumor activity of PEG Intron in patients with malignancies. More data are needed to determine the optimal dose in various oncologic indications. However, these results provide a sound rationale for further investigation of PEG Intron. PMID- 12124840 TI - A possible association between ionizing radiation and pituitary adenoma: a descriptive study. AB - BACKGROUND: Despite the recognition of ionizing radiation as a causal risk factor for a variety of solid tumors (including brain tumors), to date, such an association with pituitary adenoma (PA) has not been demonstrated. METHODS: To evaluate a possible association between past exposure to radiation and the occurrence of PA, the authors reviewed about 4900 medical records of patients who had been irradiated in childhood for tinea capitis. An additional search for patients was performed using the Israel Cancer Registry. The average radiation dose to the pituitary gland was estimated as 0.56 grays, and, for all patients, a meticulous validation of the irradiation was performed. RESULTS: A group of 16 patients who developed symptomatic PA after childhood exposure to radiotherapy were identified. Overall, the clinical and demographic characteristics of these patients were similar to other series reported in the literature. There was an apparently high rate of second primary tumors (25%), all of them in the irradiated area, diagnosed among this group. The methodologic issues that limit the demonstration of a possible association between radiation and PA and the epidemiologic and experimental findings in the literature are discussed. CONCLUSIONS: In view of the ample amount of evidence identifying low-dose ionizing radiation as a risk factor for a number of intracranial tumors as well as for tumors arising in endocrine organs, a radiation immunity of the pituitary gland is difficult to accept. Hence, the authors suggest that this series should be considered as preliminary observation that supports the role of ionizing radiation in the development of this tumor. PMID- 12124841 TI - Radiation-related cranial nerve palsy in patients with nasopharyngeal carcinoma. AB - BACKGROUND: Cranial nerve palsy is a rare complication after patients with nasopharyngeal carcinoma (NPC) receive radiotherapy using a technique that delivers 180-200 centigrays (cGy) per day. Cranial neuropathy is of particular clinical interest in terms of making a differential diagnosis, because it is also a common presenting manifestation in patients with NPC. Cranial neuropathy may lead to distressing signs and symptoms in these patients, and their treatment has not been addressed in previous reports. This article presents the authors' experience with radiotherapy-related cranial nerve palsy in patients with NPC. METHODS: Nineteen patients were diagnosed with radiation-related neuropathy. Patients with recurrent tumors or with a suspicion of persistent or recurrent tumors were excluded. Most patients were treated using 180 cGy or 200 cGy per fraction per day. The total dose was 7000-13,000 cGy to the nasopharynx and 5000 9000 cGy to the neck. Unilateral vocal cord paralysis alone and hearing loss were not included in the analysis. RESULTS: There were 15 male patients and 4 female patients. The latency before palsy occurred was 12-240 months. Single nerve palsy developed in four patients, including two patients with hypoglossal palsy and two patients with recurrent laryngeal palsy. Two patients had three nerve palsies each. The other 13 patients presented with 2 nerve palsies each. Vagus and hypoglossal palsy appeared to be a frequent combination and occurred in 11 patients. Overall, there were 17 patients with hypoglossal palsy (7 bilateral, 8 left-sided, and 2 right-sided), 11 patients with vagus palsy (2 bilateral, 7 left sided, and 2 right-sided), 6 patients with recurrent laryngeal nerve palsy (5 bilateral), and 2 patients with accessory palsies (all bilateral). Marked neck fibrosis was present in 12 patients. Patients who had vocal cord paralysis suffered from easy choking and hoarseness. Severe respiratory difficulty occurred in two patients who had bilateral vocal cord palsy. Surgical procedures included laryngoplasty, tracheostomy, and gastrostomy. Quality of life improved considerably after patients underwent surgery. CONCLUSIONS: Radiotherapy-related cranial nerve palsy may occur in patients with NPC after they receive conventional radiotherapy. Hypoglossal nerve palsy was found the most frequently in this series, followed by vagus nerve palsy and recurrent laryngeal nerve palsy. Neck fibrosis and the course of the three nerves through the neck may be important risk factors for the development of palsy. The diagnosis must be made only after the possibilities of tumor-induced palsy and idiopathic palsy are excluded. Surgery is helpful in improving the quality of life in many patients. PMID- 12124842 TI - Calbindin-d(28k): a marker of recurrence for medulloblastomas. AB - BACKGROUND: The expression of the Ca(2+)-binding protein calbindin-D(28k) was analyzed in medulloblastomas in relation to clinical features and other biologic markers related to cell proliferation, differentiation, p53, and cerebellar developmental regulated gene expression. METHODS: Immunohistochemistry was carried out on histologic slides from a first retrospective series of 29 nonmetastatic and 10 metastatic medulloblastoma formalin-fixed, paraffin-embedded tissues, using specific antibodies against calbindin-D(28k), calretinin, alpha parvalbumin and beta-parvalbumin, and S100 proteins. Informed consent was obtained from the subjects and/or guardians. Other biologic markers for differentiation, cell proliferation, the expression of the p53 tumor suppressor gene protein, and cerebellar developmental regulated genes were similarly investigated. A second series of 16 medulloblastomas from young patients (younger than 15 years) was added in order to validate the results obtained in the first series. RESULTS: Of all the markers investigated, only calbindin-D(28k) was significantly associated with prognosis. Survival and remission (i.e. recurrence free) time analysis performed on all the cases (n = 55) confirmed a high risk of death (P = 0.004) and recurrence (P = 0.003) associated with calbindin positivity. As calbindin-positivity was predominantly observed in tumors from young patients, the authors confirmed its prognostic value in the subgroup of patients younger than 15 years (n = 37). Cox regression analysis showed a significant and independent prognostic value for calbindin expression and, to a lesser extent, the type of surgery (total or subtotal). Three risk groups were thus identified, distinguishing among the cases characterized by a total resection and calbindin-negativity (good prognosis), by a subtotal resection and calbindin-negativity (intermediary), and by calbindin-positivity (bad prognosis). CONCLUSIONS: The current study suggests that calbindin-positive medulloblastomas represent a subclass of aggressive tumors more frequently seen in younger patients. PMID- 12124843 TI - New WHO histologic classification predicts prognosis of thymic epithelial tumors: a clinicopathologic study of 200 thymoma cases from China. AB - BACKGROUND: In 1999, a World Health Organization (WHO) committee published histologic criteria for distinct thymoma entities (labeled as Type A, AB, B1, B2, B3 thymomas) and for the heterogeneous group of thymic carcinomas, collectively called Type C thymomas. Whether WHO-defined histologic thymoma subtypes are of independent prognostic relevance has yet to be proved. METHODS: Two hundred thymomas from the Shanghai Chest Hospital with a mean follow-up time of 15 years (range, 1-246 months) were studied for the relevance of WHO histologic subtype and other factors (stage, therapy, and myasthenia gravis [MG]) for survival. RESULTS: In order of frequency, 68 patients (34.0%) had Type AB, 39 (19.5%) had Type B2, 36 (18.0%) had Type C, 27 (13.5%) had Type B3, 17 (8.5%) had Type B1, and 8 (4.0%) had Type A thymoma. Five cases (2.5%) were rare thymomas not mentioned in the WHO classification. Survival data showed significant differences among the histologic subtypes (log rank test: P < 0.001). Among patients with Type A and AB thymomas, none died of tumor; of the Type B1 thymoma patients, only one (5.9%) died at 22 months. Type B2, B3, and C thymomas had a significantly worse prognosis with 5-year survival rates of 75.0%, 70.0%, and 48.0%, respectively. Ninety-six patients (48.0%) were in Masaoka Stage I, 26 (13.0%) were in Stage II, 65 (32.5%) were in Stage III, and 13 (6.5%) were in Stage IV. Stage was highly significant in predicting survival (log rank, test P < 0.001). The association between histologic subtype and invasive behavior (stage) was statistically significant (P < 0.001). However, histology was an independent predictive factor of survival in Stage I and II thymomas: Type B2, B3, and C thymomas had a worse prognosis than Type A, AB, and B1 thymomas (log rank test: P < 0.003). Thirty patients (15.0%) presented with MG. MG was significantly more frequent in Type B2 and B3 than in Type A, AB, and B1 thymomas (P < 0.01). On multivariate analysis, MG had no adverse effect on survival (P = 0.17). Radiation or chemotherapy improved patients' survival at 5 and 10 years in Type B2, B3, and C thymomas (log rank test: P < 0.003). CONCLUSIONS: Tumor stage is the most important determinant of survival in thymoma patients, but the WHO histologic subtype is an independent prognostic factor in Stage I and II thymomas, among which WHO Type A, AB, and B1 thymomas form a low-risk group. Patients with high risk thymomas might profit from novel adjuvant radiochemotherapy regimens. PMID- 12124844 TI - Contrasts in cancer prevalence in Connecticut, Iowa, and Utah. AB - BACKGROUND: Cancer prevalence--the proportion of a population with cancer, including those recently diagnosed, those in treatment, and survivors--is an important indicator of future health care requirements. Only limited information on cancer prevalence is available for the United States. In particular, comparative interstate studies are not available. In this study, we estimate and analyze the prevalence of seven major cancers in Connecticut, lowa, and Utah using the tried and tested PREVAL method applied to National Cancer Institute registry data. METHODS: We analyzed data on 242,851 carcinomas of the stomach, colorectum, pancreas, breast, uterus (corpus), ovary, and non-Hodgkin lymphoma (NHL), diagnosed in white Americans from 1973 through 1992. Observed prevalence was estimated by applying the PREVAL method to incidence and life status data from the cancer registries. Complete prevalence was estimated by applying correction factors obtained by modeling incidence and survival rates. RESULTS: The ratio of the highest to the lowest prevalence (as proportions) ranged from 1.69 for uterine carcinoma to 2.73 for stomach carcinoma, showing that marked differences in cancer prevalence exist within the United States. Utah had the lowest prevalence for each carcinoma. Connecticut and lowa had similar prevalence levels for carcinomas of the colorectum, pancreas, and ovary and for NHL. Breast carcinoma was the most prevalent, with 826 cases per 100,000 of population in Utah, 1518 per 100,000 in lowa, and 1619 per 100,000 in Connecticut. Cancer survival did not differ greatly among the three registry populations. The major determinants of prevalence differences were incidence and the population age distribution. CONCLUSIONS: PREVAL provides reliable estimates of the numbers of living people in a population who have had a cancer diagnosis. Prevalence depends on incidence and survival and on the age structure of population. All these factors have changed markedly in recent years and will continue to do so in the future. Cancer prevalence should be monitored over time to evaluate changes by area, sex, age, and cancer site. The prevalence figures presented are directly comparable with those from European cancer registries. PMID- 12124845 TI - Reduced focal adhesion kinase and paxillin phosphorylation in BCR-ABL-transfected cells. AB - BACKGROUND: BCR-ABL formation is critical to oncogenic transformation in chronic myelogenous leukemia and has been implicated as a key event leading to alterations in cytoskeletal structures and adhesion in the leukemic cells. The authors therefore investigated the effect of p210(BCR-ABL) on actin polymerization as well as on the expression and phosphorylation state of the adhesion proteins paxillin and focal adhesion kinase (FAK). METHODS: Transfection with BCR-ABL constructs abrogated the ability of NIH 3T3 fibroblasts to adhere and the cells underwent striking morphologic changes. RESULTS: Scanning electron microscopy revealed that the cells lost their elongated appearance and became rounded. This alteration was associated with significantly reduced actin polymerization. In addition, steady-state levels of paxillin and FAK protein were increased. However, while the overall level of phosphotyrosines was also increased, the amount of tyrosine phosphorylated paxillin and FAK was reduced in the BCR-ABL-transfected cells as compared to the parental cells. Culture on extracellular fibronectin matrix partially reversed the morphologic changes and resulted in a return, albeit incomplete, of filamentous actin in BCR-ABL transfected 3T3 fibroblasts. In addition, phosphorylation of paxillin and FAK in the BCR-ABL-transfected NIH 3T3 cells was restored. CONCLUSIONS: The authors conclude that, in the current system, transfection of BCR-ABL attenuates FAK and paxillin phosphorylation and reduces actin polymerization, events accompanied by significant alterations in cellular morphology. The observation that exposure of the cells to fibronectin partially reverses all these changes suggests that the focal adhesion proteins and actin structures nevertheless remain responsive to signaling from the outside. PMID- 12124846 TI - Solution structure of a pentasaccharide representing the repeating unit of the O antigen polysaccharide from Escherichia coli O142: NMR spectroscopy and molecular simulation studies. AB - Conformational studies have been performed of a pentasaccharide derived from the O-polysaccharide from Escherichia coli O142. The polymer was selectively degraded by anhydrous hydrogen fluoride and reduced to yield an oligosaccharide model of its repeating unit, which in the branching region consists of four aminosugars. A comparison of (1)H and (13)C chemical shifts between the pentasaccharide and the polymer showed only minor differences, except where the cleavage had taken place, indicating that the oligomer is a good model of the repeating unit. Langevin dynamics and molecular dynamics simulations with explicit water molecules were carried out to sample the conformational space of the pentasaccharide. For the glycosidic linkages between the hexopyranoside residues, small but significant changes were observed between the simulation techniques. One-dimensional (1D) (1)H,(1)H double pulsed field gradient spin echo (DPFGSE) transverse rotating frame Overhauser effect spectroscopy (T-ROESY) experiments were performed, and homonuclear cross-relaxation rates were obtained. Subsequently, a comparison of interproton distances from NMR experiment and the two simulation approaches showed that in all cases the use of explicit water in the simulations resulted in better agreement. Hydrogen-bond analysis of the trajectories from the molecular dynamics simulation revealed interresidue interactions to be important as a cluster of different hydrogen bonds and as a distinct highly populated hydrogen bond. NMR data are consistent with the presence of hydrogen bonding within the model of the repeating unit. PMID- 12124847 TI - In vitro protein-polysaccharide conjugation: tyrosinase-catalyzed conjugation of gelatin and chitosan. AB - The enzyme tyrosinase was used for the in vitro conjugation of the protein gelatin to the polysaccharide chitosan. Tyrosinases are oxidative enzymes that convert accessible tyrosine residues of proteins into reactive o-quinone moieties. Spectrophotometric and dissolved oxygen studies indicate that tyrosinase can oxidize gelatin and we estimate that 1 in 5 gelatin chains undergo reaction. Oxidized tyrosyl residues (i.e., quinone residues) can undergo nonenzymatic reactions with available nucleophiles such as the nucleophilic amino groups of chitosan. Ultraviolet/visible, (1)H-NMR, and ir provided chemical evidence for the conjugation of oxidized gelatin with chitosan. Physical evidence for conjugation was provided by dynamic viscometry, which indicated that tyrosinase catalyzes the sol-to-gel conversion of gelatin/chitosan mixtures. The gels formed from tyrosinase-catalyzed reactions were observed to differ from gels formed by cooling gelatin. In contrast to gelatin gels, tyrosinase-generated gels had different thermal behavior and were broken by the chitosan-hydrolyzing enzyme chitosanase. These results demonstrate that tyrosinase can be exploited for the in vitro formation of protein-polysaccharide conjugates that offer interesting mechanical properties. PMID- 12124848 TI - Conformational dynamics of helix S6 from Shaker potassium channel: simulation studies. AB - Prolines in transmembrane (TM) alpha-helices are believed to play an important structural and/or functional role in membrane proteins. At a structural level a proline residue distorts alpha-helical structure due to the loss of at least one stabilizing backbone hydrogen bond, and introduces flexibility in the helix that may result in substantial kink and swivel motions about the effective "hinge." At a functional level, for example in Kv channels, it is believed that proline induced molecular hinges may have a direct role in gating, i.e., the conformational change linked to opening/closing the channel to movement of ions. In this article we study the conformational dynamics of the S6 TM helix from of the Kv channel Shaker, which possesses the motif PVP--a motif that is conserved in Kv channels. We perform multiple molecular dynamics simulations of single S6 helices in a membrane-mimetic environment in order to effectively map the kink swivel conformational space of the protein, exploiting the ability of multiple simulations to achieve greater sampling. We show that the presence of proline locally perturbs the helix, disrupting local dihedral angles and producing local twist and unwinding in the region of the hinge--an effect that is relaxed with distance from the PVP motif. We furthermore show that motions about the hinge are highly anisotropic, reflecting a preferred region of kink-swivel conformation space that may have implications for the gating process. PMID- 12124849 TI - Dimer structure of magainin 2 bound to phospholipid vesicles. AB - Magainin 2 from African clawed frog Xenopus laevis is an antimicrobial peptide with broad spectra and action mechanisms considered to permeabilize bacterial membranes. CD, vibration, and solid-state NMR spectroscopies indicate the peptide adopts an alpha-helical conformation on binding to phospholipid bilayers, and its micelle-bound conformation, being monomeric and alpha-helical, is well detailed. We showed, however, that the peptide dimerizes on binding to phospholipid bilayers. This difference in the conformation and aggregation state between micelle- and bilayer-bound states prompted us to analyze the conformation of an equipotent analog of magainin 2 (F5Y,F16W magainin 2) bound to phosphatidylcholine vesicles using transferred nuclear Overhauser enhancement (TRNOE) spectroscopy. While observed medium-range TRNOE cross peaks were characteristic of alpha-helix, many long-range cross peaks were not compatible with the peptide's monomeric state. Simulated annealing calculations generated dimer structures indicating (1) two peptide molecules have a largely helical conformation in antiparallel orientation forming a short coiled-coil structure, (2) residues 4-20 are well converged and residues 9-20 are in an alpha-helical conformation, and (3) the interface of the two peptide molecules is formed by well-defined side chains of hydrophobic residues. Finally, determined structures are compatible with numerous investigations examining magainin-phospholipid interactions. PMID- 12124850 TI - Self-aggregation properties of spin-labeled zervamicin IIA as studied by PELDOR spectroscopy. AB - In this article, the pulsed double electron-electron resonance in electron spin echo (PELDOR) technique is applied to study the self-aggregation of spin-labeled zervamicin IIA, a hexadecapeptide antibiotic of fungal origin, which is known to form ion channels in a phospholipid double layer. Measurements of the ion channel forming properties and the antibiotic activity of the analog indicate that replacement of the C-terminal phenylalaninol by the amino-2,2,6,6 tetramethylpiperidinyloxy (TEMPO) residue does not influence the biophysical and biological properties. The dipole-dipole interaction between the spin labels of the fully biologically active peptide analog was studied in frozen (77 K) glassy solutions in different ratios of toluene-methanol. The spin-labeled zervamicin IIA molecules were shown to form aggregates. An average distance between the spin labels in the aggregates was estimated to be in the range of 25-35 A (depending on the solvent composition), indicating that the amphiphilic helical peptide molecules are oriented in an antiparallel fashion. Increasing of methanol content in the solution results in a loosening of the aggregate structure. It was shown that the fraction of aggregated zervamicin IIA molecules is less than 44-67% depending on the solvent composition. The general usefulness of the method to obtain structural long-range information in a range of several tens of angstroms is demonstrated by comparison with the peptide cluster of trichogin GA IV. PMID- 12124851 TI - Cell death, autoantigen cleavage, and autoimmunity. PMID- 12124852 TI - Scleroderma and Smads: dysfunctional Smad family dynamics culminating in fibrosis. PMID- 12124853 TI - Atherosclerosis in rheumatoid arthritis: morphologic evidence obtained by carotid ultrasound. AB - OBJECTIVE: Recent studies have suggested increased cardiovascular disease among patients with rheumatoid arthritis (RA). We undertook this study to obtain morphologic evidence of subclinical atherosclerosis in RA patients. METHODS: We used high-resolution B-mode ultrasound to compare carotid artery intima-media wall thickness (IMT) between 53 postmenopausal women with RA and 53 controls matched by age, sex, and menopause status. No subject in either group had a history of atherosclerosis or its complications. We investigated the association between IMT and relevant clinical and therapeutic variables, including the impact of low-dose corticosteroid therapy (< or = 10 mg/day prednisolone). RESULTS: The mean +/- SD IMT of the left and right common carotid arteries in RA patients was significantly greater than that in controls (0.77 +/- 0.09 mm versus 0.68 +/- 0.14 mm; P < 0.001). Early RA (duration < or = 1 year) was associated with lesser IMT than was RA of longer duration (0.72 +/- 0.03 mm versus 0.78 +/- 0.01 mm; P < 0.04). Prednisolone use was not associated with increased IMT (0.78 +/- 0.02 mm in nonusers versus 0.76 +/- 0.01 mm in users; P = 0.38). CONCLUSION: Our data indicate that RA patients have an ultrasonic marker of early atherosclerosis consistent with an increased risk for atherosclerosis. PMID- 12124854 TI - Bone loss in patients with rheumatoid arthritis: results from a population-based cohort of 366 patients followed up for two years. AB - OBJECTIVE: To evaluate the extent of and risk factors for bone loss in a population-based cohort of patients with rheumatoid arthritis (RA) receiving conventional health care. METHODS: In a longitudinal study, clinical data were collected and bone mineral density (BMD) measurements were performed at baseline and after 2 years. Dual-energy x-ray absorptiometry was used for hip and spine BMD measurements. At baseline, patients received advice about lifestyle adjustments and calcium and vitamin D supplementation; during the followup period they were treated with antirheumatic and bone-sparing drugs, according to clinical judgment. RESULTS: After a mean +/- SD of 2.2 +/- 0.2 years, 366 (298 women, 68 men) of the 488 patients who were examined at baseline were reexamined. At that time, 47.9% were current users of corticosteroids and 37.0% were using antiresorptive drugs (hormone replacement therapy, bisphosphonates, or calcitonin). The mean BMD reduction was -0.64% in the femoral neck, -0.77% in the total hip, and -0.29% in the spine at L2-4. BMD was increased at all measurement sites in current users of antiresorptive drugs (0.16-1.64%) but was decreased in patients using calcium and vitamin D alone (-1.99% to -1.39%) and in patients not using any osteoporosis treatment (-1.20% to -0.43%). Current use of corticosteroids was independently associated with increased risk for BMD loss in the total hip (odds ratio [OR] 2.63, 95% confidence interval [95% CI] 1.38-5.00) and spine at L2-4 (OR 2.70, 95% CI 1.30-5.63), whereas current use of antiresorptive drugs was associated with decreased risk for bone loss in the total hip (OR 0.43, 95% CI 0.20-0.89). CONCLUSION: Results of this population based, 2-year followup study indicate that adequate management of patients with RA, addressing both the rheumatic disease and osteoporosis, protects against bone loss. PMID- 12124855 TI - An epidemiologic study of trends in prevalence of rheumatoid factor seropositivity in Pima Indians: evidence of a decline due to both secular and birth-cohort influences. AB - OBJECTIVE: Previous population studies have suggested that both rheumatoid factor (RF) production and rheumatoid arthritis (RA) may be declining in occurrence, and both secular and birth-cohort influences have been implicated. Since Pima Indians have a very high incidence of RA and also have shown recent evidence of a decline in RA, this study evaluated the relative contributions of age, secular, and birth cohort influences on RF seropositivity in the Pima Indian population. METHODS: RF data, as assayed by both the bentonite flocculation test (BFT) and the sheep cell agglutination test (SCAT), were available on 5,345 Pima Indians born between 1886 and 1975, who were surveyed at biennial intervals between 1966 and 1995. An age period-cohort analysis was conducted using data on 18,295 examinations undertaken during the period of study. RESULTS: There was a decline in the proportion of positive test results for RF (titer > or = 1:32) by both BFT and SCAT, in both male and female subjects from 1966-1975 to the later decades of the study (1976 1985 and 1986-1995). Across all periods, by both assays, the crude proportion of positive titers increased with increasing age of the subjects. There was a very clear birth-cohort effect: the highest likelihood of seropositivity was in those individuals born around the end of the nineteenth century, with continuing decline in seropositivity up to the most recent birth cohort. A logistic regression analysis, adjusting for Pima heritage and sex, demonstrated a substantially greater influence of birth cohort than of calendar year on the frequency of RF positivity. CONCLUSION: In the Pima Indian population, environmental influences in early life are important predictors of the lifelong likelihood of RF positivity. This may have implications for understanding the epidemiology and etiology of RA. PMID- 12124856 TI - Benefit of an extract of Tripterygium Wilfordii Hook F in patients with rheumatoid arthritis: a double-blind, placebo-controlled study. AB - OBJECTIVE: To examine the safety and efficacy of an extract of Tripterygium wilfordii Hook F (TWHF) in the treatment of patients with rheumatoid arthritis (RA). METHODS: An ethanol/ethyl acetate extract from the roots of TWHF was prepared and used in a prospective, double-blind, placebo-controlled study in patients with longstanding RA in whom conventional therapy had failed. Patients were randomly assigned to receive either placebo or low-dose (180 mg/day) or high dose (360 mg/day) extract for 20 weeks, followed by an open-label extension period. Clinical responses were defined as 20% improvement in disease activity according to the American College of Rheumatology criteria. Side effects were actively queried and recorded at each visit. RESULTS: A total of 35 patients were enrolled in the trial; 21 patients completed the 20-week study. One patient from each group withdrew because of side effects. Twelve, 10, and 10 patients in the placebo, low-dose, and high-dose groups, respectively, completed at least 4 weeks of treatment. Of these patients, 8 and 4 in the high-dose and low-dose groups, but none in the placebo group, met criteria for clinical response. Four, 4, and 7 patients in the placebo, low-dose, and high-dose groups, respectively, were enrolled in the open-label extension; of these, 2, 4, and 5 patients, respectively, met criteria for clinical response. The most common side effect was diarrhea, which caused 1 patient in the high-dose group to withdraw from the trial. No patients withdrew because of adverse events during the open-label extension. CONCLUSION: The ethanol/ethyl acetate extract of TWHF shows therapeutic benefit in patients with treatment-refractory RA. At therapeutic dosages, the TWHF extract was well tolerated by most patients in this study. PMID- 12124857 TI - High levels of osteoprotegerin and soluble receptor activator of nuclear factor kappa B ligand in serum of rheumatoid arthritis patients and their normalization after anti-tumor necrosis factor alpha treatment. AB - OBJECTIVE: To test the hypotheses that 1) proinflammatory cytokines affect osteoprotegerin (OPG) and soluble receptor activator of nuclear factor kappa B ligand (sRANKL) production and therefore the OPG and sRANKL levels differ in rheumatoid arthritis (RA) patients in comparison with healthy individuals; and 2) anti-tumor necrosis factor alpha (anti-TNF alpha) therapy influences OPG and sRANKL levels. METHODS: Sera were obtained from healthy individuals or RA patients receiving the combination of infliximab and methotrexate. Peripheral blood mononuclear cells (PBMCs) and synovial fluid mononuclear cells (SFMCs) were isolated from RA patients. Fibroblast-like synoviocytes (FLS) were isolated from synovial tissue obtained at total knee replacement in RA patients. Supernatants from cells stimulated with cytokines were collected after culture in vitro. Concentrations of OPG and sRANKL were determined by enzyme-linked immunosorbent assays. RESULTS: A strong positive correlation between OPG concentration and age was observed in healthy individuals but not in RA patients. The OPG and sRANKL levels were higher in RA patients than in healthy controls. Cultured FLS spontaneously secreted much higher amounts of OPG than PBMCs or SFMCs. Proinflammatory cytokines enhanced OPG production. Anti-TNF alpha treatment resulted in the normalization of serum OPG and sRANKL levels in RA patients without influencing the OPG:sRANKL ratio. CONCLUSION: Although higher serum levels of OPG and sRANKL are present in RA patients than in healthy individuals, the ratio of OPG:sRANKL is similar. There is an age-dependent increase of OPG but not sRANKL levels in healthy subjects. Anti-TNF alpha treatment results in the normalization of elevated levels of OPG and sRANKL in RA patients. PMID- 12124858 TI - Stromelysin 1 (matrix metalloproteinase 3) and HLA-DRB1 gene polymorphisms: Association with severity and progression of rheumatoid arthritis in a prospective study. AB - OBJECTIVE: To test the hypothesis of an association between a polymorphism in the matrix metalloproteinase 3 (MMP-3) gene promoter and the susceptibility, severity, and progression of rheumatoid arthritis (RA), and to further document the association between HLA-DRB1 alleles encoding the shared epitope (SE) and the severity and progression of RA. METHODS: Patients with early RA (n = 103) were included in this prospective study. A total radiographic damage score (TDS; by the Sharp/van der Heijde method) was used to quantify RA severity at baseline and after 4 years of followup. The 5A/6A biallelic polymorphism in the MMP-3 gene promoter was analyzed using fluorescence-based polymerase chain reaction (PCR). HLA-DRB1 genotyping was performed using PCR methods. Control subjects (n = 127) were unrelated healthy individuals. RESULTS: MMP-3 allele carriage rates and allele and genotype frequencies did not differ between patients and controls. The MMP-3 6A/6A genotype was associated with the highest TDS both at baseline and after a 4-year followup and with the highest progression of the TDS over the 4 years of followup. The DRB1 SE+/+ genotype was associated with the highest TDS after a 4-year followup and with the highest progression of the TDS over the 4 years of followup. Patients homozygous for MMP-3 6A and DRB1 SE had the highest progression of the TDS. CONCLUSION: This study provides the first evidence of an association between a polymorphism in the MMP-3 gene promoter and the severity and progression of RA, but not RA susceptibility. Investigation of this polymorphism could be combined with that of DRB1 gene polymorphism to improve the predictive accuracy and management strategy in early RA. PMID- 12124859 TI - A subset of natural killer cells is greatly expanded within inflamed joints. AB - OBJECTIVE: To determine whether natural killer (NK) cells are present within inflamed joints and whether they might play a role in amplifying the inflammatory process. METHODS: Paired samples of peripheral blood and synovial fluid were obtained from 22 patients with inflammatory arthritis. The frequency and phenotype of the peripheral and synovial NK cells were analyzed using a panel of monoclonal antibodies. Further experiments were performed to investigate the functional capacity of the synovial NK cells. RESULTS: The study showed that the CD3-, CD56(bright) subset of NK cells was greatly expanded within inflamed joints. Our experiments suggested that this subset of cells was preferentially recruited from the periphery and that NK cells may be further activated by cytokines present within the joint. Furthermore, synovial NK cells responded to a combination of interleukin-12 (IL-12) and IL-15, cytokines that are secreted by cells of the monocyte/macrophage lineage, by rapidly secreting interferon-gamma, a cytokine that can, in turn, activate macrophages. CONCLUSION: A subset of NK cells was expanded within inflamed joints. The functional properties of these NK cells rendered them good candidates for a role in interacting with the macrophage/monocyte population within the joint, thus amplifying the production of proinflammatory cytokines. PMID- 12124860 TI - Very low prevalence of hip osteoarthritis among Chinese elderly in Beijing, China, compared with whites in the United States: the Beijing osteoarthritis study. AB - OBJECTIVE: To compare the prevalence of osteoarthritis (OA) of the hip among elderly persons in China and the US. METHODS: We recruited a population-based sample of 1,506 persons (82% of those enumerated) ages > or = 60 years living in Beijing, China. Subjects answered questions about joint symptoms and underwent radiography of the pelvis. Radiographs of the Beijing subjects were intermingled with hip radiographs of white women ages > or = 65 years from the Study of Osteoporotic Fractures (SOF) and white men and women ages 60-74 years from the First National Health and Nutrition Examination Survey (NHANES-I) and were then interpreted. Radiographic hip OA was defined as the presence of 1 of the following 3 findings in either hip: minimum joint space of < or = 1.5 mm, definite osteophytes and joint space narrowing, or > or = 3 radiographic features of OA. Symptomatic hip OA was defined as both radiographic OA and hip pain. RESULTS: The crude prevalence of radiographic hip OA in Chinese ages 60-89 years was 0.9% in women and 1.1% in men; it did not increase with age. Chinese women had a lower age-standardized prevalence of radiographic hip OA compared with white women in the SOF (age-standardized prevalence ratio 0.07) and the NHANES-I (prevalence ratio 0.22). Chinese men had a lower prevalence of radiographic hip OA compared with white men of the same age in the NHANES-I (prevalence ratio 0.19). There were no cases of symptomatic hip OA in the Chinese men and only 1 case in the Chinese women; 35 cases were expected in both sexes. CONCLUSION: This is the first population-based study of hip OA in China to use standardized radiographic methods and definitions. We found that hip OA was 80-90% less frequent than in white persons in the US. Identification of the genetic and environmental factors that underlie these differences may help elucidate the etiology and prevention of hip OA. PMID- 12124861 TI - Finer linkage mapping of primary hip osteoarthritis susceptibility on chromosome 11q in a cohort of affected female sibling pairs. AB - OBJECTIVE: To finer linkage-map a primary osteoarthritis (OA) susceptibility locus as a prerequisite to linkage disequilibrium/association analysis. METHODS: A 50-cM interval of chromosome 11q that we had previously identified as harboring susceptibility to hip OA in a female sibling-pair cohort was subjected to finer linkage mapping. Thirty-five microsatellite markers with a mean marker interval of 1.4 cM were genotyped in 146 families containing female sibling pairs who were concordant for hip OA, as ascertained by total hip replacement. RESULTS: Two point and multipoint linkage analysis revealed 2 distinct regions of linkage within the 50-cM interval. The first locus had a linkage interval of 11.9 cM and was centered at 81.5 cM from the 11p telomere, with a maximum multipoint logarithm of odds (LOD) score of 2.4. The second region had a linkage interval of 6.5 cM and was centered at 93.1 cM from the 11p telomere, with a maximum multipoint LOD score of 1.8. CONCLUSION: Dense linkage mapping has highlighted the presence of 2 loci on chromosome 11q, each conferring susceptibility to hip OA. Both loci are sufficiently narrow for association analysis to be undertaken. PMID- 12124862 TI - Avoidance of activity and disability in patients with osteoarthritis of the knee: the mediating role of muscle strength. AB - OBJECTIVE: Avoidance of activity is hypothesized to lead to muscle weakness and consequently, to physical disability. This study was undertaken to validate the avoidance model by providing evidence for the mediating role of muscle weakness in the relationship between avoidance of activity and physical disability in patients with osteoarthritis (OA) of the knee. METHODS: Data on avoidance of activity, observed physical disability, and muscle strength of the knee in 107 patients with knee OA were analyzed. A series of regression analyses was performed to establish the mediating role of muscle weakness. First, the effect of avoidance of activity on the level of disability was assessed. Next, the relationship between avoidance of activity and muscle strength was established. Finally, the mediating role of muscle strength could be established if the effect of avoidance of activity on disability decreased when muscle strength was taken into account. RESULTS: Initially, avoidance of activity accounted for 21.5% of variance in disability. Avoidance of activity also accounted for 3.9% of variance in muscle strength. After muscle strength was taken into account, the variance in disability accounted for by avoidance of activity was reduced from 21.5% to 15.7%. Thus, the criteria for establishing the mediating role of muscle strength were met. CONCLUSION: The results of this study provide evidence for the mediating role of muscle weakness in the relationship between avoidance of activity and disability in patients with knee OA. PMID- 12124863 TI - Cyclooxygenase 2-dependent prostaglandin E2 modulates cartilage proteoglycan degradation in human osteoarthritis explants. AB - OBJECTIVE: To examine cyclooxygenase-2 (COX-2) enzyme expression, its regulation by interleukin-1 beta (IL-1 beta), and the role of prostaglandin E(2) (PGE(2)) in proteoglycan degradation in human osteoarthritic (OA) cartilage. METHODS: Samples of human OA articular cartilage, meniscus, synovial membrane, and osteophytic fibrocartilage were obtained at knee arthroplasty and cultured ex vivo with or without IL-1 beta and COX inhibitors. COX expression was evaluated by immunohistochemistry and Western blot analysis. The enzymatic activity of COX was measured by conversion of arachidonic acid to PGE(2). Cartilage degradation was evaluated by measuring the accumulation of sulfated glycosaminoglycans in the medium. RESULTS: IL-1 beta induced robust expression of COX-2 and PGE(2) in OA meniscus, synovial membrane, and osteophytic fibrocartilage explants, whereas low levels were produced in OA articular cartilage. IL-1 beta also induced cartilage proteoglycan degradation in OA synovial membrane-cartilage cocultures. Increased proteoglycan degradation corresponded to the induction of COX-2 protein expression in, and PGE(2) production from, the synovial membrane. Dexamethasone, neutralizing IL-1 beta antibody, or the selective COX-2 inhibitor, SC-236, attenuated both the IL-1 beta-induced PGE(2) production and cartilage proteoglycan degradation in these cocultures. The addition of PGE(2) reversed the inhibition of proteoglycan degradation caused by SC-236. CONCLUSION: IL-1 beta induced production of COX-2 protein and PGE(2) was low in OA articular cartilage compared with that in the other OA tissues examined. IL-1 beta-mediated degradation of cartilage proteoglycans in OA synovial membrane-cartilage cocultures was blocked by the selective COX-2 inhibitor, SC-236, and the effect of SC-236 was reversed by the addition of exogenous PGE(2). Our data suggest that induction of synovial COX-2-produced PGE(2) is one mechanism by which IL-1 beta modulates cartilage proteoglycan degradation in OA. PMID- 12124864 TI - Study of the role of leukotriene B()4 in abnormal function of human subchondral osteoarthritis osteoblasts: effects of cyclooxygenase and/or 5-lipoxygenase inhibition. AB - OBJECTIVE: To compare the effect of licofelone, NS-398 (an inhibitor of cyclooxygenase 2 [COX-2]), and BayX-1005 (an inhibitor of 5-lipoxygenase activating protein) on the production of leukotriene B(4) (LTB(4)) and prostaglandin E(2) (PGE(2)), and on cell biomarkers by human osteoarthritis (OA) subchondral osteoblasts. METHODS: Primary in vitro osteoblasts were prepared from subchondral bone specimens obtained from OA patients and autopsy subjects. LTB(4) and PGE(2) levels were measured by enzyme-linked immunosorbent assay in conditioned media of osteoblasts incubated in the presence or absence of licofelone, NS-398, or BayX-1005. The effect of these drugs or of the addition of LTB(4) on alkaline phosphatase (AP) activity and osteocalcin release by OA and normal osteoblasts was determined. The presence of LTB(4) receptors in normal and OA osteoblasts was evaluated by Western blot analysis. RESULTS: OA osteoblasts produced variable levels of PGE(2) and LTB(4) compared with normal osteoblasts. Licofelone, at the maximal dose used, inhibited production of PGE(2) and LTB(4) by OA osteoblasts by a mean +/- SEM of 61.2 +/- 6.4% and 67.0 +/- 7.6%, respectively. NS-398 reduced PGE(2) production by 75.8 +/- 5.3%. BayX-1005 inhibited LTB(4) production in OA osteoblasts by 38.7 +/- 14.5% and marginally affected PGE(2) levels (reduction of 14.8 +/- 5.3%). Licofelone dose-dependently stimulated 1,25-dihydroxyvitamin D-induced AP activity while inhibiting osteocalcin release. BayX-1005 partly reproduced these effects, but NS-398 failed to affect them. LTB(4) dose-dependently inhibited AP activity in OA osteoblasts, while its effect on osteocalcin depended on endogenous LTB(4) levels in these cells. In normal osteoblasts, LTB(4) dose-dependently stimulated osteocalcin, whereas it failed to influence AP. LTB(4) receptors BLT1 and BLT2 were present in normal and OA osteoblasts. CONCLUSION: Licofelone inhibits the production of PGE(2) and LTB(4). Selective effects of licofelone on AP and osteocalcin occur via its role on LTB(4) production. Because LTB(4) can modify cell biomarkers in OA and normal osteoblasts, our results suggest licofelone could modify abnormal bone remodeling in OA. PMID- 12124865 TI - Use of joint fluid analysis for determining cartilage damage in osteonecrosis of the knee. AB - OBJECTIVE: To assess the value of joint fluid analysis for determining cartilage degradation and prognosis in spontaneous osteonecrosis (ON) of the knee. METHODS: Synovial fluid was obtained from 30 knees with spontaneous ON (26 medial femoral condyles, 4 medial tibial plateaus) as well as from 50 knees with medial compartmental osteoarthritis (OA) as a control. Levels of chondroitin 6-sulfate (C6S), C4S, and hyaluronic acid were measured with high-performance liquid chromatography. The lesion size, appearance of the articular cartilage, and results of magnetic resonance imaging (MRI) were compared with the results of joint fluid analysis. RESULTS: The mean +/- SD level of C6S was 82.2 +/- 36.6 nmoles/ml in joint fluid from ON knees, which was significantly higher than the levels in knees with grade 2 (47.2 +/- 20.0 nmoles/ml) and grade 3 (55.8 +/- 29.2 nmoles/ml) OA. The C6S:C4S ratio was highest in lesions with mild articular changes and reflected the macroscopic alteration of cartilage overlying the ON lesion. The concentration of C6S in the 9 knees with lesions that covered > or = 40% of the condyle (99.0 +/- 32.9 nmoles/ml) was higher than that in the 17 knees with lesions that covered <40% of the condyle (67.2 +/- 31.7 nmoles/ml). Knees with bone marrow edema on MRI had a higher level of C6S than did knees with a fibrous-like appearance. CONCLUSION: While radiologic staging was useful for indicating the size of the ON lesion, it was less valuable for determining articular cartilage damage. Joint fluid analysis may provide more precise information about articular cartilage degradation in ON, and the findings may also be of prognostic significance. PMID- 12124866 TI - Effects of prasterone on corticosteroid requirements of women with systemic lupus erythematosus: a double-blind, randomized, placebo-controlled trial. AB - OBJECTIVE: To evaluate whether treatment with prasterone (dehydroepiandrosterone [DHEA]) would allow the dosage of prednisone (or an equivalent corticosteroid) to be reduced to < or = 7.5 mg/day for 2 months or longer while maintaining stable or reduced disease activity in steroid-dependent women with systemic lupus erythematosus (SLE). METHODS: In a double-blind, randomized trial, 191 female SLE patients receiving prednisone (10-30 mg/day) were treated daily with either placebo, 100 mg of oral prasterone (an adrenal androgen), or 200 mg of oral prasterone for 7-9-months. At monthly intervals, corticosteroid dosages were reduced by algorithm in patients whose SLE Disease Activity Index (SLEDAI) score was stable or improved. Patients for whom a sustained reduction in the dosage of prednisone (< or = 7.5 mg/day) was achieved for at least the last 2 months of the 7-9-month treatment period were classified as responders. RESULTS: Response rates were 41% in the placebo group, 44% in the 100-mg prasterone group, and 55% in the 200-mg group (P = 0.110, 200 mg versus placebo). Among the 137 subjects (45 in the placebo group, 47 in the 100-mg group, and 45 in the 200-mg group) who had active disease at baseline (defined as SLEDAI score >2), 29%, 38%, and 51%, respectively, were responders (P = 0.031 for 200 mg prasterone versus placebo). Acne was the most common adverse event but was generally mild. Clinical and laboratory changes primarily reflected androgenic effects of prasterone. CONCLUSION: Among women with lupus disease activity, reducing the dosage of prednisone to < or = 7.5 mg/day for a sustained period of time while maintaining stabilization or a reduction of disease activity was possible in a significantly greater proportion of patients treated with oral prasterone, 200 mg once daily, compared with patients treated with placebo. PMID- 12124867 TI - Hormonal and reproductive risk factors for development of systemic lupus erythematosus: results of a population-based, case-control study. AB - OBJECTIVE: Estrogen and prolactin may accelerate the progression of murine systemic lupus erythematosus (SLE). In humans, 85% of lupus patients are women, which also suggests the importance of hormonal factors in disease pathogenesis. The purpose of this study was to examine hormonal and reproductive risk factors for lupus among women. METHODS: This population-based, case-control study included 240 female SLE patients diagnosed between January 1, 1995 and July 31, 1999 who fulfilled the American College of Rheumatology classification criteria. Female controls (n = 321) were identified through driver's license records. Logistic regression was used to estimate odds ratios (ORs) and 95% confidence intervals (95% CIs) as measures of association, adjusting for age, state, race, and education. Analyses were limited to exposures before diagnosis. RESULTS: Breast-feeding was associated with a decreased risk of developing SLE (OR 0.6, 95% CI 0.4-0.9), with a statistically significant trend for number of babies breast-fed and total weeks of breast-feeding. There were no associations with number of pregnancies or live births. Natural menopause occurred earlier in women with subsequent development of SLE compared with controls (P < 0.001). There was little association between SLE and current use or duration of use of hormone replacement therapy or oral contraceptives, and no association with previous use of fertility drugs. CONCLUSION: We found little evidence that estrogen- or prolactin-related exposures are associated with an increased risk of lupus. The reduced risk observed among women who had breast-fed one or more babies should be examined in other studies. Early natural menopause, rather than decreasing risk of SLE because of reduced estrogen exposure, may be a marker of susceptibility to development of SLE. PMID- 12124868 TI - Occupational exposure to crystalline silica and risk of systemic lupus erythematosus: a population-based, case-control study in the southeastern United States. AB - OBJECTIVE: Crystalline silica may act as an immune adjuvant to increase inflammation and antibody production, and findings of occupational cohort studies suggest that silica exposure may be a risk factor for systemic lupus erythematosus (SLE). We undertook this population-based study to examine the association between occupational silica exposure and SLE in the southeastern US. METHODS: SLE patients (n = 265; diagnosed between January 1, 1995 and July 31, 1999) were recruited from 4 university rheumatology practices and 30 community based rheumatologists in 60 contiguous counties. Controls (n = 355), frequency matched to patients by age, sex, and state of residence, were randomly selected from driver's license registries. The mean age of the patients at diagnosis was 39 years; 91% were women and 60% were African American. Detailed occupational and farming histories were collected by in-person interviews. Silica exposure was determined through blinded assessment of job histories by 3 industrial hygienists, and potential medium- or high-level exposures were confirmed through followup telephone interviews. Odds ratios (ORs) and 95% confidence intervals (95% CIs) were estimated by logistic regression. RESULTS: More patients (19%) than controls (8%) had a history of medium- or high-level silica exposure from farming or trades. We observed an association between silica and SLE (medium exposure OR 2.1 [95% CI 1.1-4.0], high exposure OR 4.6 [95% CI 1.4-15.4]) that was seen in separate analyses by sex, race, and at different levels of education. CONCLUSION: These results suggest that crystalline silica exposure may promote the development of SLE in some individuals. Additional research is recommended in other populations, using study designs that minimize potential selection bias and maximize the quality of exposure assessment. PMID- 12124869 TI - Increased prevalence of familial autoimmunity in simplex and multiplex families with juvenile rheumatoid arthritis. AB - OBJECTIVE: To determine if the prevalence of autoimmunity among relatives of patients with juvenile rheumatoid arthritis (JRA) is greater than that among relatives of healthy volunteer control subjects. METHODS: Interviews were used to obtain histories of the following disorders among living first- and second-degree relatives of 110 patients and 45 controls: alopecia areata, ankylosing spondylitis, dermatomyositis, Graves' disease, Hashimoto thyroiditis, insulin dependent diabetes mellitus, inflammatory bowel disease, iritis, JRA, multiple sclerosis, psoriasis, RA, systemic lupus erythematosus, and vitiligo. Chi squares, odds ratios (ORs), and 95% confidence intervals (95% CIs) were calculated. Families of 23 JRA affected sibpairs were interviewed subsequently. RESULTS: There were no significant differences between patients and controls with regard to age, sex, ethnicity, or family size. Patients had 1,228 relatives and controls had 496 relatives. Of all the relatives of the patients, 155 had at least 1 autoimmune disorder, compared with 20 relatives of the controls (12.6% versus 4.0%; OR 3.4 [95% CI 2.1-5.7], P < 0.000001). The prevalence of autoimmunity was increased in first-degree and in second-degree relatives of patients (16.1% and 10.6%, respectively). The prevalence of Hashimoto thyroiditis was significantly higher in the relatives of patients (OR 3.5 [95% CI 1.6-7.9], P = 0.0008). The prevalences of other disorders were not significantly different. JRA affected sibpair families had an increased prevalence of autoimmunity (15.0%). A history of arthritis was found significantly more frequently in the JRA affected sibpair families, but not in the simplex families. CONCLUSION: These data demonstrate that the prevalence of autoimmunity is significantly higher among first- and second-degree relatives of JRA patients. This suggests that clinically different autoimmune phenotypes may share common susceptibility genes, which may act as risk factors for autoimmunity. PMID- 12124870 TI - Dysregulation of transforming growth factor beta signaling in scleroderma: overexpression of endoglin in cutaneous scleroderma fibroblasts. AB - OBJECTIVE: As an initial approach to understanding the basis of the systemic sclerosis (SSc; scleroderma) phenotype, we sought to identify genes in the transforming growth factor beta (TGF beta) signaling pathway that are up regulated in lesional SSc fibroblasts relative to their normal counterparts. METHODS: We used gene chip, differential display, fluorescence-activated cell sorter, and overexpression analyses to assess the potential role of TGF beta signaling components in fibrosis. Fibroblasts were obtained by punch biopsy from patients with diffuse cutaneous SSc of 2-14 months' duration (mean 8 months) and from age- and sex-matched healthy control subjects. RESULTS: Unexpectedly, we found that fibroblasts from SSc patients showed elevated expression of the endothelial cell-enriched TGF beta receptor endoglin. Endoglin is a member of the nonsignaling high-affinity TGF beta receptor type III family. The expression of endoglin increased with progression of disease. Transfection of endoglin in fibroblasts suppressed the TGF beta-mediated induction of connective tissue growth factor promoter activity. CONCLUSION: SSc is characterized by overproduction of matrix; that is, genes that are targets of TGF beta signaling in normal fibroblasts. Our findings suggest that lesional SSc fibroblasts may overexpress endoglin as a negative feedback mechanism in an attempt to block further induction of profibrotic genes by TGF beta. PMID- 12124871 TI - Development of a CENP-A/CENP-B-specific immune response in a patient with systemic sclerosis. AB - Antibodies directed against an epitope motif on CENP-A have been shown to cross react with mimotopes on other autoantigens and on Epstein-Barr nuclear antigen 1 (EBNA-1), suggesting a molecular mimicry. We describe here the gradual development of an anticentromere immune response in a patient with systemic sclerosis, which started from an antihistone response and was not mediated by molecular mimicry. Via an epitope on histone H3, the antibody response spread to a homologous epitope in the H3 homology domain of CENP-A. This was followed by an intramolecular epitope spreading to N-terminal peptides of CENP-A containing the known epitope motif G-P-X(1)-R-X(2). From there it spread to corresponding epitopes on CENP-B and to mimotopes of the major CENP-A epitope motif on other autoantigens including EBNA-1. Whether the D-penicillamine treatment received by this patient was involved in the triggering of this cascade remains a matter of speculation. PMID- 12124872 TI - Recognition of Granzyme B-generated autoantigen fragments in scleroderma patients with ischemic digital loss. AB - OBJECTIVE: To examine whether autoantibodies recognizing granzyme B (GB)-cleaved autoantigens are associated with ischemic digital loss (IDL) in limited systemic sclerosis (SSc). METHODS: Fifteen of 19 patients with limited SSc and IDL were matched by age, sex, race, and duration of disease to controls with limited SSc but without IDL. The sera were used to immunoblot HeLa cell lysates and chromosome preparations that had been incubated in vitro in the absence or presence of GB. Anticentromere antibodies (ACAs) were assayed by immunofluorescence and immunoprecipitation of in vitro-translated centromere proteins (CENP-B and CENP-C). Immunoprecipitation of GB-cleaved CENPs was also performed. RESULTS: GB-cleaved autoantigens were immunoblotted by 16 of 19 IDL sera (84.2%) compared with 6 of 15 non-IDL sera (40.0%) (odds ratio 8.0, 95% confidence interval 1.6-40.0). This association persisted after adjustment for ACA status. Furthermore, the presence of antibodies to centromere proteins as well as to GB-cleaved antigens was highly specific for IDL, occurring in 12 of 19 IDL patients (63.2%) and in none of 15 controls (P < 0.0001). An identical 60-kd GB-generated fragment was recognized by 5 of 16 IDL sera (31.3%) and was demonstrated to arise through GB-mediated cleavage of CENP-C. GB-cleaved CENP-C fragments were recognized preferentially over the intact CENP-C molecule by antibodies from patients with IDL. CONCLUSION: The striking recognition of GB generated autoantigen fragments by sera from patients with limited SSc and IDL constitutes the first in vivo evidence that antibodies against GB-generated centromeric peptide fragments identify a distinct clinical subset. PMID- 12124873 TI - Differences in idiopathic inflammatory myopathy phenotypes and genotypes between Mesoamerican Mestizos and North American Caucasians: ethnogeographic influences in the genetics and clinical expression of myositis. AB - OBJECTIVE: As part of a larger, worldwide study of the ethnogeography of myositis, we evaluated the clinical, serologic, and immunogenetic features of Mestizo (Mexican and Guatemalan) and North American Caucasian patients with idiopathic inflammatory myopathy (IIM). METHODS: Clinical manifestations, autoantibodies, HLA-DRB1 and DQA1 alleles, and immunoglobulin Gm/Km allotypes were compared between 138 Mestizos with IIM and 287 Caucasians with IIM, using the same classification criteria and standardized questionnaires. RESULTS: IIM in Mestizo patients was characterized by a higher proportion of dermatomyositis (69% of adult Mestizos versus 35% of adult Caucasians; P < 0.001) and anti-Mi-2 autoantibodies (30% versus 7% of adults, respectively, and 32% versus 4% of children, respectively; P < 0.01). Genetic risk factors also differed in these populations. Whereas Mestizos had no HLA risk factors for IIM, HLA-DRB1*0301, the linked allele DQA1*0501, and DRB1 alleles sharing the first hypervariable region motif (9)EYSTS(13) were major risk factors in Caucasian patients with IIM. Furthermore, different HLA-DRB1 and DQA1 alleles were associated with anti-Mi-2 autoantibodies (DRB1*04 and DQA1*03 in Mestizos and DRB1*07 and DQA1*02 in Caucasians). Immunoglobulin gamma-chain allotypes Gm(1), Gm(17) (odds ratio for both 11.3, P = 0.008), and Gm(21) (odds ratio 7.3, P = 0.005) and kappa-chain allotype Km(3) (odds ratio 7.3, P = 0.005) were risk factors for IIM in Mestizos; however, no Gm or Km allotypes were risk or protective factors in Caucasians. In addition, Gm and Km phenotypes were unique risk factors (Gm 1,3,17 5,13,21 and Gm 1,17 23 21 and Km 3,3) or protective factors (Km 1,1) for the development of myositis and anti-Mi-2 autoantibodies (Gm 1,2,3,17 23 5,13,21) in adult Mestizos. CONCLUSION: IIM in Mesoamerican Mestizos differs from IIM in North American Caucasians in the frequency of phenotypic features and in the immune-response genes predisposing to and protecting from myositis and anti-Mi-2 autoantibodies at 4 chromosomal loci. These and other data suggest the likelihood that the expression of IIM is modulated by different genes and environmental exposures around the world. PMID- 12124874 TI - In vitro cytokine production and proliferation of T cells from patients with anti proteinase 3- and antimyeloperoxidase-associated vasculitis, in response to proteinase 3 and myeloperoxidase. AB - OBJECTIVE: To investigate in vitro proliferative responses of CD4+ T cells and generation of specific cytokines induced by stimulation of peripheral blood mononuclear cells (PBMCs) from patients with antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis with the autoantigens proteinase 3 (PR3) and myeloperoxidase (MPO). METHODS: PBMCs from vasculitis patients with PR3 ANCA or MPO ANCA and from healthy controls were stimulated for 7 days with PR3, MPO, or control stimuli. Proliferation of CD4+ T cells was assessed by flow cytometry, using the proliferation marker Ki-67. Levels of the pro-proliferative cytokines interleukin-2 (IL-2) and IL-6 and of the Th1 and Th2 cytokines interferon-gamma (IFN gamma) and IL-10 in culture supernatants were determined. RESULTS: PR3 and MPO induced proliferative responses in CD4+ T cells from individual patients with ANCA-associated vasculitides and healthy controls in vitro. Neither PR3 nor MPO elicited significant IL-2 production. Levels of IL-6 were highest after stimulation with PR3 but low after stimulation with MPO, independent of study group. Stimulation with PR3, and to a lesser extent with MPO, induced a Th2 cytokine milieu, characterized by high production of IL-6 and IL-10 and low production of IFN gamma in patients and controls. CONCLUSION: PR3 and MPO promote proliferation of CD4+ T cells from patients with ANCA-associated vasculitides, but also cross-stimulate T cells from healthy individuals. Strong IL-10 production elicited by PR3 in vitro may act as an inhibitory signal for T cell proliferation and may have an important immunoregulatory function in vivo. PMID- 12124876 TI - Discriminatory sonographic criteria for the diagnosis of carpal tunnel syndrome. AB - OBJECTIVE: Sonographic examination of the median nerve has been suggested as a useful alternative to electrophysiologic study in the diagnosis of carpal tunnel syndrome. To determine its usefulness and the best diagnostic criterion, sonograms of patients with the disease were compared with sonograms of healthy subjects in a case-control study. METHODS: Patients with carpal tunnel syndrome and asymptomatic controls who were matched for age and sex were enrolled and underwent sonography of the wrists. Eight separate sonographic criteria were analyzed in each wrist. Data from the patient group and the control group were compared to establish optimal diagnostic criteria for carpal tunnel syndrome, using receiver operating characteristic analytic techniques. RESULTS: Thirty-five patients with carpal tunnel syndrome and 35 asymptomatic controls were examined. Increased cross-sectional area of the median nerve was found to be the most predictive measure of carpal tunnel syndrome, proximal to the tunnel inlet, at the tunnel inlet, and at the tunnel outlet, with significant differences between patients and controls. Using a receiver operating characteristic curve, a cut-off value >0.098 cm(2) at the tunnel inlet provided a diagnostic sensitivity of 89% and a specificity of 83%. CONCLUSION: Sonographic measurement of the median nerve cross-sectional area is both sensitive and specific for the diagnosis of carpal tunnel syndrome. PMID- 12124875 TI - A pulse-chase study tracking the conversion of macrophage-endocytosed serum amyloid A into extracellular amyloid. AB - OBJECTIVE: To determine whether serum amyloid A (SAA) is internalized by and processed in macrophages en route to deposition as extracellular amyloid. METHODS: SAA was tracked in cultures of peritoneal macrophages, using a pulse chase protocol. Macrophages were pulsed with either fluorescently (with Texas Red) tagged SAA (TxR-SAA) or iodinated SAA ((125)I-SAA). Cells were then rinsed and shifted to chase medium containing unlabeled SAA and amyloid-enhancing factor (AEF) to induce amyloid formation. At selected times, TxR-SAA in living cells was observed by confocal scanning microscopy. (125)I-SAA was visualized and quantified in cell lysates and medium by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and phosphorimaging. The presence of amyloid was confirmed by Congo red staining. RESULTS: Confocal microscopy immediately after the pulse revealed TxR-SAA in endosomal vesicles, with no extracellular or cell surface accumulation. After 24 hours and 72 hours of chase, virtually all TxR-SAA remained intracellular. By 10 days, extracellular fluorescence was very strong, indicating that SAA had moved out of cells. Congo red staining revealed amyloid colocalized with areas of extracellular fluorescence. Experiments using (125)I SAA showed that while 90-95% of internalized (125)I-SAA was degraded within 24 hours, 5-10% persisted as intact SAA or SAA peptides. Immediately after the pulse, SAA was full-length, but within 24 hours, discrete (125)I-SAA peptides were seen. Each peptide had an intact SAA amino-terminus, as expected for AA protein. Amyloid was detected in cultures as early as 24 hours after initiation of treatment with SAA and AEF and appeared to be intracellular. CONCLUSION: The results of this study provide direct evidence that SAA internalized by and processed in macrophages forms extracellular amyloid. Based on the presence of (125)I-AA protein in macrophage lysates prior to the appearance of extracellular TxR-labeled amyloid, it was concluded that cleavage of SAA to AA occurs intracellularly. PMID- 12124877 TI - Avascular necrosis of the femoral head in a patient with Fabry's disease: identification of ceramide trihexoside in the bone by delayed-extraction matrix assisted laser desorption ionization-time-of-flight mass spectrometry. AB - Fabry's disease is a lipid storage disease caused by an X-linked hereditary deficiency of alpha-galactosidase. The enzymatic defect causes progressive deposition of ceramide trihexoside (CTH) in various tissues, leading to renal failure, premature myocardial infarction, and stroke, with a high rate of mortality in younger patients. Among the complications associated with Fabry's disease, a few cases involving avascular necrosis (AVN) of the femoral head have been reported. However, direct evidence of deposition of CTH in bone marrow in the femoral head has not been demonstrated. This report describes a 58-year-old man who underwent total hip arthroplasty for femoral head AVN associated with Fabry's disease. The accumulation of CTH was examined by chemical analysis of the sphingolipid extracted from the femoral head, using delayed-extraction matrix assisted laser desorption ionization-time-of-flight mass spectrometry. This is the first report confirming the presence of CTH in the sphingolipid fraction from normal and necrotic bone of a patient with Fabry's disease. PMID- 12124878 TI - Kinetics of bone protection by recombinant osteoprotegerin therapy in Lewis rats with adjuvant arthritis. AB - OBJECTIVE: To assess the effect of different dosages and treatment schedules of osteoprotegerin (OPG) on joint preservation in an experimental model of adjuvant induced arthritis (AIA). METHODS: Male Lewis rats with AIA (6-8 per group) were treated with a subcutaneous bolus of recombinant human OPG according to one of the following schedules: daily OPG (an efficacious regimen) starting at disease onset (days 9-15), early intervention (days 9-11), delayed intervention (days 13 15), and extended therapy (days 9-22). Inflammation (hind paw swelling) was quantified throughout the clinical course; osteoporosis (bone mineral density [BMD], by quantitative dual x-ray absorptiometry) and morphologic appraisals of inflammation, bone damage, intralesional osteoclasts (by semiquantitative histopathologic scoring), and integrity of the articular cartilage matrix (by retention of toluidine blue stain) were determined in histology sections of arthritic hind paws. RESULTS: OPG provided dose- and schedule-dependent preservation of BMD and periarticular bone while essentially eliminating intralesional osteoclasts. Dosages > or = 2.5 mg/kg/day preserved or enhanced BMD and prevented essentially all erosions. A dosage of 4 mg/kg/day protected joint integrity to a comparable degree when given for 7 (days 9-15) or 14 (days 9-22) consecutive days. At this dosage, early intervention (days 9-11) was twice as effective as delayed intervention (days 13-15) at preventing joint dissolution. Erosions and osteoclast scores were greatly decreased for 26 days (measured from the first treatment) after 7 or 14 daily doses of OPG (4 mg/kg/day). OPG treatment also prevented loss of cartilage matrix proteoglycans, an indirect consequence of protecting the subchondral bone. No OPG dosage or regimen alleviated weight loss, inflammation, or periosteal osteophyte production. CONCLUSION: These data indicate that OPG preserves articular bone and (indirectly) articular cartilage in arthritic joints in a dose- and schedule dependent manner, halts bone erosion when given at any point during the course of arthritis, produces sustained antierosive activity after a short course, and is most effective when initiated early in the disease. PMID- 12124879 TI - Inhibition of adjuvant-induced arthritis by interleukin-10-driven regulatory cells induced via nasal administration of a peptide analog of an arthritis related heat-shock protein 60 T cell epitope. AB - OBJECTIVE: To prevent and treat experimental arthritis via nasal administration of an altered peptide ligand (APL) from the major arthritogenic epitope in adjuvant-induced arthritis (AIA) and to explore the mechanisms involved. METHODS: Peptides were administered nasally before and after induction of arthritis. Splenocytes and lymph node cells draining both the site of inflammation and the site of tolerance induction were used for cell transfer and were studied for antigen-specific T cell characteristics. In addition, attempts were made to stop T cell tolerance in vitro, using anticytokine antibodies. RESULTS: Nasal administration of a modulatory APL of the heat-shock protein 60 (Hsp60) 180-188 T cell epitope, alanine 183, had a suppressive effect in AIA that far exceeded that of the wild-type epitope. In addition to its effectiveness in preventing AIA, alanine 183 may be effective in the treatment of ongoing AIA. The protective effect of alanine 183 can be passively transferred using activated splenocytes. Nasal administration of alanine 183 did not lead to detectable T cell proliferation or interleukin-2 (IL-2) production in mandibular lymph node cells, while transforming growth factor beta (TGF beta), IL-10, and IL-4 were readily produced. Likewise, after nasally induced tolerance, followed by induction of arthritis, inguinal lymph node cells produced IL-4, TGF beta, and IL-10. After neutralizing in vitro the individual cytokines with anticytokine antibodies, only blocking of IL-10 production led to reversal of tolerance, at the site of tolerance induction and the site of inflammation. CONCLUSION: Nasal administration of an APL of Hsp60 180-188 induces highly effective protection against AIA through generation of regulatory cells that produce IL-4, TGF beta, and IL-10, whereas the induced tolerance is driven mainly by production of IL-10. PMID- 12124880 TI - Folate-targeted imaging of activated macrophages in rats with adjuvant-induced arthritis. AB - OBJECTIVE: To determine whether overexpression of the high-affinity folate receptor (FR) on activated macrophages can be exploited to selectively target imaging agents to sites of inflammation in rats with adjuvant-induced arthritis (AIA). METHODS: Folic acid was conjugated to a (99m)Tc chelator (the complex termed EC20), and its distribution was visualized using gamma scintigraphy in healthy rats, rats with AIA, and arthritic rats that had been depleted of macrophages. To confirm that uptake was mediated by the FR, excess folic acid competition studies were conducted, and tissue FR levels were quantitated using a radioligand binding assay. Flow cytometry was also used to investigate uptake of folate conjugates into macrophages of both arthritic and healthy rats. RESULTS: EC20 concentrated in the arthritic extremities of diseased rats but not in the extremities of healthy rats. The intensity of images of affected tissues was greatly reduced in the presence of excess competing folic acid. The livers and spleens of arthritic animals also showed enhanced uptake of EC20 and increased levels of FR. Depletion of macrophages from arthritic animals reduced tissue FR content and concomitantly abolished uptake of EC20. In addition, macrophages isolated from livers of rats with AIA exhibited a significantly higher binding capacity for folate conjugates than did macrophages obtained from healthy rats. CONCLUSION: Although EC20 is currently undergoing clinical evaluation for use in the imaging of ovarian carcinomas, the present results suggest that it may also be useful for assaying the participation of activated macrophages in inflammatory processes such as rheumatoid arthritis. PMID- 12124881 TI - Effects of long-term estrogen replacement therapy on osteoarthritis severity in cynomolgus monkeys. AB - OBJECTIVE: To determine the effects of long-term estrogen replacement therapy (ERT) on the severity of osteoarthritis (OA) of the knee joint in surgically postmenopausal (bilaterally ovariectomized) female monkeys. A secondary aim was to evaluate the effect of soy phytoestrogen (SPE) treatment on the severity of OA. METHODS: Feral adult female cynomolgus macaques were ovariectomized bilaterally and then randomly divided into 3 age- and weight-matched treatment groups. For 3 years, the first group received ERT with conjugated equine estrogens, the second group received SPE, and the third group received no treatment (controls). At necropsy, histologic lesions of OA were graded, and the area and thickness of cartilage and subchondral bone were measured. The data were summarized by principal components analysis, and the resulting factors and individual variables were compared using analysis of variance and analysis of covariance (age and weight as covariates). RESULTS: Cartilage lesions of OA were significantly less severe in the animals given ERT compared with those in the control group. This treatment effect remained significant when adjusted for age and weight. The factor representing subchondral bone was significantly higher, but the number of osteophytes was lower, in the ERT group compared with the control group. SPE treatment had no significant effect on cartilage or bone lesions of OA. CONCLUSION: These results demonstrate that long-term ERT significantly reduces the severity of OA lesions in this animal model. PMID- 12124882 TI - Sustained response to tumor necrosis factor alpha-blocking agents in two patients with SAPHO syndrome. PMID- 12124883 TI - Clinical images: Osteosclerotic myeloma associated with the POEMS syndrome. PMID- 12124884 TI - High-resolution ultrasound for the study of target joints in rheumatoid arthritis. PMID- 12124886 TI - Can very high-dose anti-tumor necrosis factor blockade at onset of rheumatoid arthritis produce long-term remission? PMID- 12124889 TI - DNASE I mutation and systemic lupus erythematosus in a Spanish population: comment on the article by Tew et al. PMID- 12124887 TI - Morbidity not increased in rheumatoid arthritis patient with profound lymphopenia following CD4 monoclonal antibody therapy: comment on the article by Isaacs et al. PMID- 12124892 TI - Call for a trial of Lyprinol, an over-the-counter 5-lipoxygenase inhibitor: comment on the article by Kowal-Bielecka et al. PMID- 12124894 TI - Congenital fascial dystrophy, a new scleroderma-like genetic disease with limitation of joint mobility: comment on the clinical image presented by Di Rocco. PMID- 12124896 TI - Cognitive function in fibromyalgia: comment on the article by Park et al. PMID- 12124900 TI - Common patterns of facial ontogeny in the hominid lineage. AB - Recent evaluation of Neanderthal and modern human ontogeny suggests that taxon specific features arose very early in development in both lineages, with early, possibly prenatal, morphological divergence followed by parallel postnatal developmental patterns. Here we use morphometric techniques to compare hominoid facial growth patterns, and show that this developmental phenomenon is, in fact, not unique to comparisons between Neanderthals and modern humans but extends to Australopithecus africanus and to the hominoid lineage more broadly. This finding suggests that a common pattern of juvenile facial development may be more widespread and that the roots of ontogenetically early developmental differentiation are deep-perhaps predating the ape/human split of 6+ million years ago. PMID- 12124901 TI - A closer look at Neanderthal postcanine dental morphology: the mandibular dentition. AB - Neanderthals are known to exhibit unique incisor morphology as well as enlarged pulp chambers in postcanine teeth (taurodontism). Recent studies suggest that their overall dental pattern (i.e., in morphologic trait frequencies) is also unique. However, what this means in a phylogenetic sense is not known. Although exploring the polarity of dental morphologic characters is essential to understanding the phylogenetic implications of unique patterns of variation, few have undertaken this task. This study moves beyond standard scoring methods, which are based on modern humans, to include several postcanine traits that have not been considered previously. In addition, Homo erectus is used as an outgroup to Neanderthals and modern humans to explore the polarity of these traits. The findings of this study suggest that Neanderthals are not only unique in their pattern of dental trait frequencies (as found in previous studies) but that they present several dental autapomorphies, as well. These include a high frequency of the mid-trigonid crest in lower molars and unique morphology of the lower premolars. Interestingly, these characters are not observed in the Mauer mandible, which some have claimed to be a member of a chronospecies that is a unique ancestor to Neanderthals. PMID- 12124902 TI - Trust, but verify: the necessity of empirical verification in quantitative neurobiology. AB - The preferred procedure for estimating neuron number has been the subject of intense discussion. Sampling error, systematic bias, and the physical properties of the tissue examined all contribute to the possibility that estimates may not reflect actual neuron number. Even when restricting analysis to a particular, well-defined structure such as dorsal root ganglia, these factors act in a manner that varies with maturity of the animal, species, fixation, processing, and neuron size. These considerations reinforce the necessity of comparing estimates with the actual number of neurons as determined by three-dimensional reconstructions. PMID- 12124903 TI - Midline corpus callosum is a neuroanatomical focus of fetal alcohol damage. AB - Prenatal exposure to high levels of alcohol often induces birth defects that combine morphological stigmata with neurological or neuropsychological deficits. But it has proved problematic to diagnose these syndromes in adolescents and adults, in whom the morphological signs are absent or attenuated, the behavioral deficits nonspecific, and the exposure history often difficult to reconstruct. Localizing the associated brain abnormalities might circumvent most of these difficulties. To this end, three-dimensional (3D) locations were recorded for 67 homologous points on or near the corpus callosum in magnetic resonance (MR) brain images from 60 adolescents and adults who were normal, 60 diagnosed with fetal alcohol syndrome, and 60 diagnosed with fetal alcohol effects. We combined the standard statistical approach to this type of geometric data, Procrustes analysis, with a multivariate strategy focusing on differences in variability. In this data set, the shape of the corpus callosum and its vicinity proves systematically much more variable in the alcohol-affected brains than in those of the normal subjects. From this excess variability follows a promising classification rule, having both high sensitivity (100 out of 117) and high specificity (49 out of 60) in this sample. The discrimination uses four landmark points and two summary scores of callosal outline shape. The information from the corpus callosum and vicinity, as viewed in MR brain images of full-grown subjects, may serve as a permanent record of the prenatal effects of alcohol, even in patients who are first suspected of these syndromes relatively late in life or who lack the facial signs of prenatal alcohol damage. The statistical pattern underlying the callosal diagnosis also leads to speculations on mechanisms of the prenatal damage. PMID- 12124904 TI - Proliferation and apoptosis in the developing human neocortex. AB - The cell kinetics of the developing central nervous system (CNS) is determined by both proliferation and apoptosis. In the human neocortex at week 6 of gestation, proliferation is confined to the ventricular zone, where mitotic figures and nuclear immunoreactivity for proliferating cell nuclear antigen (PCNA) are detectable. Cell division is symmetric, with both daughter cells reentering mitosis. At week 7, the subventricular zone, a secondary proliferative zone, appears. It mainly gives rise to local circuit neurons and glial cells. Around week 12, the ventricular and subventricular zones are thickest, and the nuclear PCNA label is strongest, indicating that proliferation peaks at this stage. Thereafter, asymmetric division becomes the predominant mode of proliferation, with one daughter cell reentering mitosis and the other one migrating out. Towards late gestation, the ventricular and subventricular zones almost completely disappear and proliferation shifts towards the intermediate and subplate zones, where mainly glial cells are generated. A remnant of the subventricular zone with proliferative activity persists into adulthood. In general, proliferation follows a latero-medial gradient in the neocortex lasting longer in its lateral parts. Apoptotic nuclei have been detected around week 5, occurring in low numbers in the ventricular zone at this stage. Apoptotic cell death increases around midgestation and then spreads throughout all cortical layers, with most dying cells located in the ventricular and subventricular zones. This spatial distribution of apoptosis extends into late gestation. During the early postnatal period, most apoptotic cells are still located in the subcortical layers. During early embryonic development, proliferation and apoptosis are closely related, and are probably regulated by common regulators. In the late fetal and early postnatal periods, when proliferation has considerably declined in all cortical layers, apoptosis may occur in neurons whose sprouting axons do not find their targets. PMID- 12124905 TI - Intraorbital anatomy of the koala (Phascolarctos cinereus). AB - This is the first documented study of the anatomical details of the contents of the normal koala orbit, excluding the bulbus oculi. Baseline data were established which are necessary for understanding and treating ocular disease in the koala (Phascolarctos cinereus). The anatomy of the orbital contents of the koala were examined and described from animals that presented dead or were euthanized for humane reasons. Dissections of the orbital cavity were performed under magnification. Polymethyl methacrylate (PMMA) casts of the nasolacrimal system and the vascular supply of the orbit were also made in order to study these systems. The superficial lymphatic drainage of the conjunctival tissues was studied by subcutaneous injection of Evan's Blue into the palpebral conjunctiva of a freshly deceased animal, and by Microfil casts of the efferent lymphatics. In general, the orbital contents of the koala are consistent with those of other carnivorous polyprotodont and herbivorous diprotodont marsupials. PMID- 12124906 TI - Bony orbital anatomy of the koala (Phascolarctos cinereus). AB - This is the first documented study of the anatomical details of the normal koala orbit. Baseline data are established which are necessary for understanding and treating ocular disease in the koala (Phascolarctos cinereus). The bony anatomy of the orbit of the koala was examined and described from animals that presented dead or had been euthanized for humane reasons. Dissections of the orbital cavity were performed under magnification, and descriptions of the orbital bones were from macerated skulls that had been boiled and cleaned. In general, the orbital bones of the koala, and their respective foramina, are consistent with those of other carnivorous polyprotodont and herbivorous diprotodont marsupials. PMID- 12124907 TI - Determination of bone volume by osteocyte population. AB - During development and growth, biological tissues and organisms can control their size and mass by regulating cell number (Raff, 1992; Conlon and Raff, 1999). Later in life both cell number and organ mass decrease (Buetow, 1985). We demonstrate that the number density of bone cells buried in the calcified matrix (osteocyte lacunar density) predicts extracellular matrix volume for both cancellous and cortical bone in a broad cross-section of the population (males and females, age range 23-91 years, r(2) = 0.98). Our hypothesis is that bone mass is determined by the control of osteocyte number, and that this is a particular instance of the control of organ size through the social controls on cell survival and death (Raff, 1992; Conlon and Raff, 1999). PMID- 12124908 TI - Glial fibrillary acidic protein-positive cells of the kidney are capable of raising a protective biochemical barrier similar to astrocytes: expression of metallothionein in podocytes. AB - Blood-tissue exchange and homeostasis within the organs depend on various interactions between endothelial and perivascular cells (Buniatian, 2001). Podocytes possess anatomical and cellular features intermediate between those of astrocytes and hepatic stellate cells (HSCs). Podocytes, like HSCs, are associated with fenestrated capillaries and, similar to astrocytes, interact with the capillaries via the basement membrane and participate in permeability limiting ultrafiltration. The fact that podocytes come in direct contact with xenobiotics prompted us to investigate whether they express metallothionein (MT), an anticytotoxic system characteristic of astrocytes. In comparative studies, cryosections of 1- and 3-month-old rat kidney and adult rat brain, as well as podocytes and astrocytes from early and prolonged primary cultures of glomerular explants and newborn rat brain, respectively, were investigated. The cells were double-labeled with antiserum against glial fibrillary acidic protein (GFAP) and monoclonal antibody (MAb) against the lysine-containing epitope of Cd/Zn-MT-I (MAb MT) or MAb against alpha-actin. In kidney sections, MT immunoreactivity was detected in GFAP-positive glomerular cells and in interstitial fibroblasts. The pattern of staining for MT and GFAP in glomerular cells was similar to that of astrocytes in vivo. In glomerular cell cultures, MT was expressed in cobblestone like podocytes which contained Wilms' tumor protein and lacked desmin. MT was upregulated at later culture periods, during which podocytes acquired features typical of undifferentiated astrocytes. This study hints at the existence of common regulatory mechanisms of blood-tissue interactions by neural and non neural perivascular cells. These mechanisms appear to be used in an organ specific manner. PMID- 12124909 TI - Size and shape changes during late fetal growth (137-157 gestational days) in the pigtailed macaque (Macaca nemestrina) craniofacial complex: an application using three-dimensional coordinate data and finite element scaling analysis. AB - Form changes within the fetal pigtailed macaque (Macaca nemestrina) craniofacial complex was documented using finite element scaling analysis (FESA) and three dimensional (3D) coordinate data for 35 craniofacial landmarks. Coordinate data were digitized from 3D reconstructions of computed tomography (CT) images and 2D axial slices. Twenty-two fetal pigtailed macaques ranging in age from 137 to 157 gestational days were included (in this species, birth is estimated at 170 gestational days). The null hypothesis that the craniofacial complex grows with isometry during late fetal growth of the craniofacial complex was tested (P < 0.05), and the prediction that morphological change along an anteroposterior axis dominates late fetal growth was also investigated. The null hypothesis was rejected, indicating that allometric growth is present during late fetal growth. Growth along an anteroposterior axis is localized in the palate and mandible. The neurocranium grows along a superoinferior axis, while the neurofacial junction displays growth along both the anteroposterior and superoinferior axes. Mediolateral changes are localized between asterions, the external auditory meati, and maxillary and mandibular alveolar points. Finally, a 3D model of craniofacial growth for this species was created, localizing size and shape changes that occur during late fetal growth for each of the 35 craniofacial landmarks defined in this study. PMID- 12124910 TI - Immunohistochemical localization of heat shock protein 25 (HSP 25) during root formation of the rat molar. AB - The present study investigated the immunohistochemical localization of heat shock protein 25 (HSP 25) of rat molar teeth during root formation. Most, probably all, cells of the epithelial rest of Malassez (ERM cells) had immunoreaction for laminin, a marker protein for basement membrane. During root formation, HSP 25 immunoreactivity was observed in odontoblasts, cells at the subodontoblastic layer, and those in close proximity to the acellular cementum. HSP 25 immunopositive cells at the subodontoblastic layer were present only at the apical region. Most HSP 25-immunoreactive cells in close proximity to the cementum lacked laminin immunoreactivity. However, at postnatal day 28 a small number of cells showed immunoreaction for both HSP 25 and laminin at the cervical and bifurcational regions. Under the electron microscope, most HSP 25 immunoreactive cells along the surface of the cementum were round and contained rich organelles such as mitochondria and rough endoplasmic reticulum. They lay between fiber bundles of the periodontal ligament. The localization and morphological features of these HSP 25-immunoreactive cells resemble those of cementoblasts. On the other hand, HSP 25-immunoreactive cells at the cervical region were oval and contained few cell organelles. They were closely apposed to each other, and separated from the surrounding tissues with basal lamina. These features were similar to those of mature ERM cells. In contrast, cells with microvillus-like processes and relatively rich mitochondria, which were similar to immature ERM cells, had no immunoreaction for HSP 25. These results suggest that HSP 25 may be involved in shape alterations of ERM cells, cementoblasts, and odontoblasts during differentiation. PMID- 12124911 TI - Rapid technique for imaging the blood vascular system using stereoangiography. AB - Data on vascular anatomy traditionally have been derived from time-consuming gross dissection and histology, which has prevented the assembly of large sample sizes. Vascular injection of radiopaque medium (angiography) is a rapid technique, but, as typically performed, it has limitations (e.g., superimposition and poor subsequent dissectibility). We present a novel angiographic technique comprised of two elements: 1) a new, dissectible injection medium; and (2) stereoradiography. The injection medium consists of liquid barium (providing radiopacity) and latex (providing dissectibility). Domestic duck heads were the study system. The relative concentrations of barium and latex were varied, and the resulting radiographs were assayed for vessel radiopacity and the number of observable vessels. A wide range of barium percentages yielded excellent results, suggesting that preparation of the medium can be "eyeballed" for most applications, which enhances processing speed. The stereoradiographic element solved the superimposition problem, allowing stunning resolution of the spatial relationships of vessels to each other and to other tissues. Stereoangiography is a fast and easy technique that permits the acquisition of detailed anatomical data from many specimens, thereby enabling something rarely achieved: population level anatomical studies. PMID- 12124912 TI - Evolutionary significance of cranial variation in Asian Homo erectus. AB - Homo erectus inhabited a wide geographic area of Asia, ranging from 40 degrees north latitude in China to 8 degrees south latitude in island Southeast Asia. Yet variation within Asian H. erectus and its relation to ecological and temporal parameters have been little studied. I synthesize the revised radiometric chronologies for hominid sites in Asia and their relation to new oxygen isotope curves (proxies for climatic fluctuations and landbridge connections). These data suggest substantial opportunities in the later Pleistocene for both regional isolation and gene flow between hominids in mainland and Southeast Asia. They also suggest that the most northerly located Chinese sites (Zhoukoudian and Nanjing) may have been occupied during sequential, interglacial periods. Probably reflecting these periods of isolation, nonmetric features and principal components analysis (PCA) of calvarial shape suggest regional differentiation between northern Asian and Southeast Asian H. erectus. The most recent Southeast Asian fossils (e.g., Ngandong) conform to the Southeast Asian pattern. Except perhaps in brain size, there is no evidence that the temporally intermediate Chinese fossils are intermediate in morphology between older and younger Indonesian fossils. In fact, northern Chinese calvaria are easier to exclude from the larger Asian H. erectus hypodigm than are the Ngandong fossils. The Chinese specimens differ from the others based on their narrower occipitals and frontals for their cranial size. The Chinese sample from Zhoukoudian alone is thus not a good proxy for the morphology and variation seen within Asian H. erectus. Both the Chinese and late Indonesian samples exhibit less variation than does the early Indonesian sample; this along with their shared morphological bauplan suggests a common origin and no more than subspecific differentiation. This shared morphology, despite regional differences, was likely maintained by the increasing intensity of multiple glaciations (and longer-lasting land bridge connections) between mainland and island Southeast Asia during the last million years. PMID- 12124913 TI - Brain size and the human cranial base: a prenatal perspective. AB - Pivotally positioned as the interface between the neurocranium and the face, the cranial base has long been recognized as a key area to our understanding of the origins of modern human skull form. Compared with other primates, modern humans have more coronally orientated petrous bones and a higher degree of basicranial flexion, resulting in a deeper and wider posterior cranial fossa. It has been argued that this derived condition results from a phylogenetic increase in the size of the brain and its subcomponents (infra- and supratentorial volumes) relative to corresponding lengths of the cranial base (posterior and anterior, respectively). The purpose of this study was to test such evolutionary hypotheses in a prenatal ontogenetic context. We measured the degree of basicranial flexion, petrous reorientation, base lengths, and endocranial volumes from high-resolution magnetic resonance images (hrMRI) of 46 human fetuses ranging from 10-29 weeks of gestation. Bivariate comparisons with age revealed a number of temporal trends during the period investigated, most notable of which were coronal rotation of the petrous bones and basicranial retroflexion (flattening). Importantly, the results reveal significant increases of relative endocranial sizes across the sample, and the hypotheses therefore predict correlated variations of cranial base flexion and petrous orientation in accordance with these increases. Statistical analyses did not yield results as predicted by the hypotheses. Thus, the propositions that base flexion and petrous reorientation are due to increases of relative endocranial sizes were not corroborated by the findings of this study, at least for the period investigated. PMID- 12124914 TI - Normal neuroanatomical variation in the human brain: an MRI-volumetric study. AB - Normative data on the in vivo size of the human brain and its major anatomically defined subdivisions are not readily available. In this study, high-resolution magnetic resonance imaging was used to measure regional brain volumes in 46 normal, right-handed adults (23 men, 23 women) between the ages of 22-49 years. Parcellation of the brain was based on neuroanatomical landmarks. The following brain regions were measured: the cerebral hemispheres, frontal lobe, temporal lobe, parietal lobe, occipital lobe, cingulate gyrus, insula, cerebellum, corpus callosum, and lateral ventricles. Males tend to be significantly larger than females, for the whole brain and for nearly all of its major subdivisions, including the corpus callosum. However, the proportional sizes of regions relative to total volume of the hemisphere are remarkably similar in males and females. Variation in size of region is always greater than variation in proportional representation. Asymmetries in brain regions are not profound, with the exception of the cingulate gyrus, which is larger in the left hemisphere. Brain regions are highly correlated in size, with the exception of the lateral ventricles. After controlling for hemisphere size, the volumes of the frontal and parietal lobes are significantly negatively correlated. The occipital lobe tends to be less sexually dimorphic than other major lobes, and less correlated with other brain regions for volume. These results have implications for understanding whether or not certain sectors of the brain have shown relative expansion over the course of hominid and hominoid evolution. PMID- 12124915 TI - Late pleistocene human femoral diaphyseal curvature. AB - Anterior femoral curvature is a consistent characteristic of Pleistocene and recent humans, although variation exists in the degree of curvature among individuals and across populations. In particular, one group, the Neandertals, has been characterized for a century as having marked femoral curvature. To evaluate the degree of anterior femoral curvature in both Neandertals and other Late Pleistocene humans, their curvature subtenses and proximodistal positions were evaluated in the context of recent human variation. Recent human comparisons show little relationship between subtense (absolute curvature) and femoral length, suggesting that an index that incorporates subtense relative to the length of the femur is inappropriate for between-group assessments. Neandertals were statistically indistinguishable from Middle or earlier Upper Paleolithic modern humans in the degree of absolute curvature, all of whom had greater curvature on average than all later humans. Additionally, Neandertals and Qafzeh Skhul early modern humans had a more distal point of maximum curvature than any other group. Curvature was not strongly correlated with functional considerations including body mass estimates, surrogate variables for body size, proximal femoral articular orientation, or knee anteroposterior dimensions. The functional role of femoral anterior curvature is unknown; however, the general decrease in curvature subtense closely parallels the between-group changes in inferred levels of mobility from femoral diaphyseal robusticity and shape, suggesting that femoral curvature may reflect mobility levels and patterns among Late Pleistocene and recent humans. PMID- 12124916 TI - Probable case of Binder syndrome in a skeleton from Quarai, New Mexico. AB - Binder syndrome (maxillonasal dysplasia) is a not uncommon disorder reported in the clinical literature and is characterized by hypoplastic development of the midface. An extensive review of the paleopathology literature did not reveal any examples of Binder syndrome. In this paper, a probable case of Binder syndrome in a female skeleton, 16-17 years at age of death, from Quarai, New Mexico (ca. AD 1375-1450) is presented. This case was identified during standard documentation prior to repatriation at the National Museum of Natural History, Smithsonian Institution. The skull of this individual (381243) exhibits unusual facial features, including an underdeveloped midface, flattened glabella, absent nasal spine, and apparent alveolar prognathism, in addition to a vertebral anomaly. All of these characteristics are consistent with skeletal dysmorphologies associated with Binder syndrome. Measurements of the Quarai skull are compared with published data on Binder patients and normal control groups in order to quantify the nature of the observed morphology. Univariate analysis of craniometric/cephalometric data provides further support for a diagnosis of Binder syndrome, as critical measurements on the Quarai skull are consistent with those reported in Binder patients and significantly different from those reported for normal control groups. In addition to presenting a probable prehistoric case of Binder syndrome, this paper demonstrates the applicability of using direct comparisons of clinical data to help identify unusual conditions in skeletal remains. PMID- 12124917 TI - Population variation in second metacarpal sexual size dimorphism. AB - This paper contrasts levels of sexual size dimorphism in second metacarpal osteometric and geometric morphology in two bioculturally distinctive populations: 19th century Euro-Canadian settlers, and proto/historic central Canadian Inuit. Significant within-group sexual size dimorphism is found for all variables, though few show significant interpopulation differences. However, in every case the Euro-Canadian sample is more dimorphic than the Inuit sample. Notably, differences reside in geometric measures (total area, Imax) sensitive to variation in functional strain, and thus are interpretable in light of proximate causal models, i.e., activity profiles distinct from generalized mode of subsistence. Other proximate factors, such as nutritional stress acting to diminish Inuit sexual size dimorphism, may also play a role. However, models often cited to explain dimorphism, such as marriage practice (e.g., polygyny) or division of labor situated in mode of subsistence, do not. The higher sexual size dimorphism in the 19th century settler sample belies the notion that technological progress inevitably leads to reduced dimorphism. PMID- 12124918 TI - Scoring system for estimating age in the foot skeleton. AB - This study assesses chronological age of immature individuals from the degree of ossification evident in the foot skeleton. We considered all tarsal and ray bones in various combinations to determine the most sensitive indicators of chronological age. Skeletal maturity was rated according to a subjective but simple scoring system applied to radiographs of normal feet of children of known chronological age. Scales for assessing the primary center of ossification, secondary center of ossification, and state of fusion are defined. In general, as expected, females show earlier onset of skeletal maturity than males; in particular, females in our sample are skeletally mature in most elements beginning 48 months prior to the earliest incidence of skeletal maturity in males for the same bones. Females in our sample show a marked tendency toward skeletal maturity of all elements by 150 months of age, while males do not show the same tendency until approximately 200 months of age. In general within each sex, consecutive states of fusion show broad overlap in range of chronological age within each bone. Combining scores from several different bones enables a reasonable estimate of potential age in a test application of the model. PMID- 12124919 TI - Apportionment of global human genetic diversity based on craniometrics and skin color. AB - A number of analyses of classical genetic markers and DNA polymorphisms have shown that the majority of human genetic diversity exists within local populations (approximately 85%), with much less among local populations (approximately 5%) or between major geographic regions or "races" (approximately 10%). Previous analysis of craniometric variation (Relethford [1994] Am J Phys Anthropol 95:53-62) found that between 11-14% of global diversity exists among geographic regions, with the remaining diversity existing within regions. The methods used in this earlier paper are extended to a hierarchical partitioning of genetic diversity in quantitative traits, allowing for assessment of diversity among regions, among local populations within regions, and within local populations. These methods are applied to global data on craniometric variation (57 traits) and skin color. Multivariate analysis of craniometric variation shows results similar to those obtained from genetic markers and DNA polymorphisms: roughly 13% of the total diversity is among regions, 6% among local populations within regions, and 81% within local populations. This distribution is concordant with neutral genetic markers. Skin color shows the opposite pattern, with 88% of total variation among regions, 3% among local populations within regions, and 9% within local populations, a pattern shaped by natural selection. The apportionment of genetic diversity in skin color is atypical, and cannot be used for purposes of classification. If racial groups are based on skin color, it appears unlikely that other genetic and quantitative traits will show the same patterns of variation. PMID- 12124920 TI - Brief communication: how much larger is the relative volume of area 10 of the prefrontal cortex in humans? AB - It has long been thought that the prefrontal cerebral cortex has been greatly expanded in the human brain. Semendeferi et al. ([2001] Am. J. Phys. Anthropol. 114:224-241) showed that Brodmann's area 10 is relatively larger in the human compared to pongid brains. The question is: how much larger relatively is it? Using their data, it can be shown that the relative increase for human prefrontal area 10 is only 6% larger. Looking at the data base of neural structures provided by Stephan et al. ([1981] Folia Primatol. (Basel) 35:1-29), it is apparent that 6% is a relatively low residual value from a predicted value based on allometric considerations between total brain weight and any given neural structure. When this small increase is combined with their earlier findings on area 13 of prefrontal cortex (Semendeferi et al. [1997] J. Hum. Evol. 32:375-388), it appears that the prefrontal cortex in humans is not some 200% larger as claimed by some researchers (Deacon [1997] Symbolic Species, New York: W.W. Norton; cf. Holloway [1998] Am Sci 86:184-186), and that the findings of Semendeferi et al. ([2001] Am. J. Phys. Anthropol. 114:224-241) are in agreement with the earlier work (Semendeferi and Damasio [2000] J. Hum. Evol. 38:317-332; Semendeferi et al. [1997] J. Hum. Evol. 32:375-388), showing that the human frontal lobe volume is what would be expected for a primate of its brain size. While the prefrontal cortex may have increased relatively in Homo sapiens, the increase is likely to have been far less than currently believed. PMID- 12124921 TI - Thyroid hormone production in chimpanzees and humans: implications for the origins of human intelligence. PMID- 12124923 TI - Proceedings of the 1st Annual Conference of the Swiss Proteomics Society. Geneva, Switzerland. November 21-22, 2001. PMID- 12124925 TI - Proteomics and its trends facing nature's complexity. AB - The complexity of nature is tremendous, particularly at the epigenetic level. Proteomic studies must therefore complement genomic discoveries to better understand biological processes. Because of the very large number of modified proteins and their great variability in physico-chemical properties, no single method can be used to analyze all of them. Mass spectrometry has demonstrated its superior ability to rapidly identify and partially characterize numerous proteins in low abundance and has become a central element in most proteomic projects. Studies of protein function are necessary to understand biological pathways and this is being tackled using several approaches such as two hybrid systems, phage technology or affinity methods. Finally, mathematical and bioinformatic developments will be essential to study nature's complex systems. PMID- 12124926 TI - The immunoglobulinopathies: from physiopathology to diagnosis. AB - The diversity of immunoglobulins (Igs) results mainly from recombinations of numerous genes within the heavy (V(Heavy), D, and J(Heavy)) and within the light (V(Light), J(Light)) chain gene loci, and from somatic hypermutations occurring during the immune response of B-cells. Igs production is controlled by complex cellular and humoral mechanisms. Plasma of healthy individuals contains polyclonal Igs. Clonal expansion of cells producing antigen-specific Igs may result from physiological as well as from pathological immune events. Exquisitely sensitive and specific molecular biology techniques have been used to evaluate the clonal diversity of cells producing antigen-specific Ig. However, the application of such techniques is hampered by the necessity to collect the totality of antigen-specific B-cells for subsequent analysis, which is impossible to perform routinely in humans. In addition, these techniques do not provide quantitative information about the concentration of the circulating Igs. It is therefore necessary to use tools allowing study of the quantity of the circulating Igs, and more particularly to detect overproduction of a single homogeneous Ig resulting from the expansion of a B-cell clone secreting Igs. Here, we review the mechanisms of B-cell differentiation and Ig synthesis, discuss the diseases associated with clonal Ig production and review the methods available in the clinical laboratory for Ig analysis. PMID- 12124927 TI - Challenges in mass spectrometry. AB - Report of the round table discussionThe reported MS round table took place within the ProteomeValley Technology Fair during the Swiss Proteomics Society 2001 Congress. It was organised by the Swiss Proteomics Society (SPS) in collaboration with FontisMedia (http://www.fontismedia. com). It was scheduled November 22(nd), 2001 from 16:40 to 18:00 at the Geneva University Hospital. The objective of the MS round table was to bring together two kinds of specialists in mass spectrometry: those who develop and commercialise MS equipment and those who use it as a tool in proteomic sciences. Six expert users and six representatives of leading MS instrument companies were thus invited to actively participate in this round table and share their points of view. The main objectives of this round table were not only to share experiences, but also to share dreams regarding MS technology and discuss the actual limits. It also focused on what MS users presently expect from MS producers and what are the current priorities in terms of development, for the benefit of all parties. Although it was one of the final events of the meeting, this MS round table attracted an audience of some fifty participants of the congress who were actively involved in the debate. PMID- 12124928 TI - Subproteomics analysis of phosphorylated proteins: application to the study of B lymphoblasts from a patient with Scott syndrome. AB - Proteomics based approaches, which examine the expressed proteins of a tissue or cell type, complement the genome initiatives and are increasingly used to address biomedical questions. Proteins are the main functional output, and post translational modifications such as phosphorylation are very important in determining protein function. To address this question, we developed a method for specific immunoprecipitation using anti-phosphotyrosine antibodies. This method is directly compatible with two-dimensional gel electrophoresis (2-DE). In this report data are presented on B-lymphoblasts from a patient suffering of Scott syndrome. Scott syndrome is an orphan inherited hemorrhagic disorder due to a lack of exposure of procoagulant phosphatidylserine at the exoplasmic leaflet of plasma membrane of blood cells. We hypothesized that a consequence of the mutation is to alter phosphorylation of proteins involved in signal transduction leading to breakdown in cellular signaling pathways mediating phosphatidylserine exposure. An immunoprecipitation method combined with 2-DE was applied to search for modifications in the expression of phosphorylated polypeptides related to Scott syndrome phenotype. We report here the construction of a B-lymphoblast subproteomic map comprising of polypeptides observed after immunoprecipitation using antibodies to phosphotyrosine. The polypeptides were identified either by mass fingerprinting, by liquid chromatography-tandem mass spectrometry (LC-MS/MS) and/or by matching with various lymphoid cell 2-DE maps included in the Laboratoire de Biochimie des Proteines et Proteomique 2-DE database. A differential analysis was further performed to explore several hundred proteins in Scott B-lymphoblasts in comparison with control B-lymphoblasts. Then, image analysis allowed detection of variations between control and Scott syndrome phenotype lymphoblasts. Five spots were specifically found on 2-DE from Scott syndrome phenotype lymphoblasts, and four only appeared on 2-DE from control cells. Protein identification was achieved using a combination of mass fingerprinting and peptide identification using LC-MS/MS. PMID- 12124929 TI - Automatic amide I frequency selection for rapid quantification of protein secondary structure from Fourier transform infrared spectra of proteins. AB - Here we report the development of a new neural network based approach for rapid quantification of protein secondary structure from Fourier transform infrared (FTIR) spectra of proteins. A technique for efficiently reducing the amount of spectral data by almost 90% is suggested to facilitate faster neural network analysis. Additionally, an automatic procedure is introduced for selecting only those regions within the amide I band of protein FTIR spectra, which can be best related to secondary structure contents by subsequent neural network analysis. Based on a given reference set of FTIR spectra from proteins with known secondary structure, a subset of merely 29 out of 101 amide I absorbance values could be identified, which lead to an improved prediction accuracy. The average prediction accuracy achieved for helix, sheet, turn, bend, and other is 4.96% which is better than that achieved by alternative methods that have been previously reported indicating the significant potential of this approach. Our suggested automatic amide I frequency selection procedure may be easily extended to identify promising regions from spectral data recorded by other spectroscopic techniques, like for example circular dichroism spectroscopy. PMID- 12124930 TI - A proteomic approach to evaluate the butyrophilin gene family expression in human milk fat globule membrane. AB - Human butyrophilin (BTN) expression in milk fat globule (MFGM) was evaluated using two dimensional electrophoresis (2-DE) as the separation technique, and peptide mass mapping by matrix-assisted laser desorption/ionisation-mass spectrometry (MALDI-MS) as the identification tool. Since milk composition changes throughout lactation time, 2-DE maps in the pH range 4-7 of colostral MFGM and mature MFGM were compared, showing only slight differences in BTN spot distribution. The BTN gene family codes for seven proteins (BTN, BTN2A1, BTN2A2, BTN2A3, BTN3A1, BTN3A2, BTN3A3), their presence in human tissues has to date been evaluated only at a transcriptional level. Among 70 spots, analyzed and identified by MALDI-MS, 13 spots were identified as BTN spots and only one as a fragment of BTN2A1. BTN was present in multiple glycoforms, and two smaller BTN forms of about 45 kDa were also identified. We propose an array of BTNs on human MFGM, which could provide breast-fed infants with immune molecules during the neonatal period. PMID- 12124931 TI - Mapping of immunoreactive antigens of Francisella tularensis live vaccine strain. AB - Francisella tularensis subsp. holarctica is the common causal agent of tularemia in Europe. Besides clinical signs, the diagnosis of the disease mostly depends on serological tests. To date, there is a lack of information about the F. tularensis antigens that induce antibody response. Therefore, we have started comprehensive mapping of immunoreactive antigens using the attenuated live vaccine strain of F. tularensis LVS originating from the European virulent strain. For this purpose, the immunoreactivity of sera collected from patients suffering from tularemia, together with the control sera of patients with Lyme disease and healthy blood donors, were examined by means of one-dimensional and two-dimensional immunoblotting. Furthermore, whole cell bacterial lysates, isolated integral membrane proteins and basic proteins were exploited as antigens. By this approach more than 80 different immunorelevant antigens were detected. Most of them came from whole cell bacterial lysate and integral membrane proteins. Conversely, only a negligible reaction was found in the case of basic proteins. Forty-five spots were further selected for mass spectrometric analyses and 22 of them were annotated. Among the spots that provided characteristic reactions with sera from patients with tularemia, 60 kDa and 10 kDa chaperonins that occurred in several charge and mass variants, predominated. PMID- 12124932 TI - Matrix-assisted laser desorption/ionization-tandem mass spectrometry with high resolution and sensitivity for identification and characterization of proteins. AB - Although peptide mass fingerprinting is currently the method of choice to identify proteins, the number of proteins available in databases is increasing constantly, and hence, the advantage of having sequence data on a selected peptide, in order to increase the effectiveness of database searching, is more crucial. Until recently, the ability to identify proteins based on the peptide sequence was essentially limited to the use of electrospray ionization tandem mass spectrometry (MS) methods. The recent development of new instruments with matrix-assisted laser desorption/ionization (MALDI) sources and true tandem mass spectrometry (MS/MS) capabilities creates the capacity to obtain high quality tandem mass spectra of peptides. In this work, using the new high resolution tandem time of flight MALDI-(TOF/TOF) mass spectrometer from Applied Biosystems, examples of successful identification and characterization of bovine heart proteins (SWISS-PROT entries: P02192, Q9XSC6, P13620) separated by two dimensional electrophoresis and blotted onto polyvinylidene difluoride membrane are described. Tryptic protein digests were analyzed by MALDI-TOF to identify peptide masses afterward used for MS/MS. Subsequent high energy MALDI-TOF/TOF collision-induced dissociation spectra were recorded on selected ions. All data, both MS and MS/MS, were recorded on the same instrument. Tandem mass spectra were submitted to database searching using MS-Tag or were manually de novo sequenced. An interesting modification of a tryptophan residue, a "double oxidation", came to light during these analyses. PMID- 12124933 TI - Establishing two-dimensional gels for the analysis of Leishmania proteomes. AB - Several different sample preparation methods for two-dimensional electrophoresis (2-DE) analysis of Leishmania parasites were compared. From this work, we were able to identify a solubilization method using Nonidet P-40 as detergent, which was simple to follow, and which produced 2-DE gels of high resolution and reproducibility. PMID- 12124934 TI - A survey of the plant mitochondrial proteome in relation to development. AB - To expand the functional analysis of plant mitochondria, we have undertaken the building of the proteome of pea mitochondria purified from leaves (green and etiolated), roots and seeds. In the first stage, we focused our proteomic exploration on the soluble protein complement of the green leaf mitochondria. We used traditional two-dimensional polyacrylamide gel electrophoresis, in combination with size exclusion chromatography as a third dimension, to identify the major proteins and further resolve their macromolecular complexity. The two dimensional map of soluble proteins of green leaf mitochondria revealed 433 spots (with Coomassie blue staining) and around 73% of the proteins (in mass) were identified using three different approaches: Edman degradation, matrix-assisted laser desorption/ionization mass spectrometry and electrospray ionization tandem mass spectrometry. Quite a lot of the polypeptides were present in multiforms which indicated the presence of isoforms or the occurrence of post-translational modifications. Among these proteins, we uncovered an abundant family that was identified as aldehyde dehydrogenases, representing approximately 7.5% of the soluble proteins. The comparative analysis of soluble mitochondrial proteomes led to the identification of a number of proteins which were specifically present in root or in seed mitochondria, thus revealing the impact of tissue differentiation at the mitochondrial level. PMID- 12124935 TI - Profiling the alkaline membrane proteome of Caulobacter crescentus with two dimensional electrophoresis and mass spectrometry. AB - Attempts at protein profiling in the alkaline pH region using isoelectric focusing have often proved difficult, greatly limiting the scope of proteome analysis. We investigated several parameters using custom pH 8-11 immobilized pH gradients to separate a Caulobacter crescentus membrane preparation. These included sample application, quenching endoosomotic flow and gel matrix composition. Among these factors, the sample application position was the predominant parameter to affect two-dimensional gel quality. Separated proteins were silver stained and profiled using matrix-assisted laser desorption/ionization (MALDI) mass spectrometry. The use of a prototype MALDI-Q Tof mass spectrometer assisted identification of several proteins by providing highly informative peptide fragmentation data from the sample digests. Thirty-two unique alkaline proteins were identified in this study, which complements our previously described C. crescentus membrane proteome. Our experiments point towards new options for proteomic researchers aiming to both extend the scope of analysis, and simplify methods of identifying proteins with high confidence. PMID- 12124936 TI - Identification of protein phosphorylation sites by combination of elastase digestion, immobilized metal affinity chromatography, and quadrupole-time of flight tandem mass spectrometry. AB - Using the combination of in-gel elastase digestion, immobilized metal affinity chromatography and high resolution electrospray tandem mass spectrometry, the phosphorylation sites of two phosphoproteins were determined. Complete coverage of all phosphorylation sites (Ser10, Ser139, Thr197, Ser338) of the model phosphoprotein protein kinase A C(alpha)-subunit could be achieved by this strategy in the low picomole range. In addition, three previously unknown phosphorylation sites of the human transcription initiation factor TIF-IA (Ser44, Ser170, Ser172) were determined in this way. Both phosphoproteins could be identified in a protein database on the basis of their elastase generated phosphopeptides alone. The data of seven phosphopeptides were used for identification of protein kinase A, and those of two phosphopeptides for TIF-IA, respectively. The accurate mass data of the electrospray mass spectra recorded at high resolution are extremely useful for sequencing of the elastase generated phosphopeptides and for protein identification by database searching. PMID- 12124937 TI - A contribution to breast cancer cell proteomics: detection of new sequences. AB - Ductal infiltrating carcinoma (DIC) of the breast is the most common and potentially aggressive form of cancer. Knowledge of proteomic profiles, attained both in vivo and in vitro, is fundamental to acquire as much information as possible on the proteins expressed in these pathologic conditions. We used the breast cancer cell line 8701-BC, established from a primary DIC, with the aim of contributing to the databases on mammary cancer cells, which in turn will be very useful for the identification of differentially expressed proteins in normal and neoplastic cells. Within an analysis window comprising about 1750 discernible spots, we have at present catalogued 84 protein spots. The proteins for which an identity was assigned were identified essentially using gel comparison, N terminal (Nt) microseqencing and immune detection. Among the protein spots Nt microsequenced, sixteen corresponded to known proteins, four resulted as modified, relative to matching sequences deposited on databases, and seven were unknown. These modified or novel sequences are thus of potential interest to the knowledge of breast cancer proteomics and its applications. PMID- 12124938 TI - Identification of proteins from two-dimensional polyacrylamide gels using a novel acid-labile surfactant. AB - Protein identification by peptide mass mapping usually involves digestion of gel separated proteins with trypsin, followed by mass measurement of the resulting peptides by matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS). Positive identification requires measurement of enough peptide masses to obtain a definitive match with sequence information recorded in protein or DNA sequence databases. However, competitive binding and ionization of residual surfactant introduced during polyacrylamide gel electrophoresis (PAGE) can inhibit solid-phase extraction and MS analysis of tryptic peptides. We have evaluated a novel, acid-labile surfactant (ALS) as an alternative to sodium dodecylsulfate (SDS) for two-dimensional (2-D) PAGE separation and MALDI-MS mapping of proteins. ALS was substituted for SDS at the same concentration in buffers and gels used for 2-D PAGE. Manual and automated procedures for spot cutting and in-gel digestion were used to process Coomassie stained proteins for MS analysis. Results indicate that substituting ALS for SDS during PAGE can significantly increase the number of peptides detected by MALDI-MS, especially for proteins of relatively low abundance. This effect is attributed to decomposition of ALS under acidic conditions during gel staining, destaining, peptide extraction and MS sample preparation. Automated excision and digestion procedures reduce contamination by keratin and other impurities, further enhancing MS identification of gel separated proteins. PMID- 12124942 TI - Isolation of differentially expressed genes from wild-type and Twist mutant mouse limb buds. AB - In the mouse, Twist is required for normal limb and craniofacial development. We show that the aristaless-like transcription factors, Alx3 and Alx4 are downregulated in the Twist(-/-) mutant and may be potential targets of Twist. By suppression subtractive hybridization we isolated 31 and 18 unique clones representing mRNAs that are putatively downregulated and upregulated respectively in Twist(-/-) forelimb buds. These included genes encoding cytoskeletal components, metabolic enzymes, hemoglobin molecules, membrane transport proteins, components of transcription and translation complexes, protein modification enzymes and proteins related to cell proliferation and apoptosis. Differential expression of selected clones was validated by whole mount in situ hybridization to E10.5 wild-type and Twist(-/-) embryos. We show that four novel clones are expressed in the Twist-expressing craniofacial tissues and paraxial mesoderm and downregulated in Twist(-/-) embryos, raising the possibility that they are, in addition to genes of the Alx family, downstream targets of Twist. PMID- 12124943 TI - Sertoli and granulosa cell-specific Cre recombinase activity in transgenic mice. AB - We have established transgenic mice expressing the Cre recombinase under the control of the anti-Mullerian hormone (AMH) gene promoter. Cre activity and specificity were evaluated by different means. In AMH-Cre mice, expression of the Cre recombinase mRNA was confined to the testis and ovary. AMH-Cre mice were crossed with reporter transgenic lines and the offspring exhibited Cre-mediated recombination only in the testis and the ovary. In male, histochemical analysis indicated that recombination occurred in every Sertoli cells. In female, Cre mediated recombination was restricted to granulosa cells, but the protein was not evenly active in every cells. From these results, we conclude that potentially, this transgenic line possessing AMH promoter-driven expression of the Cre recombinase is a powerful tool to delete genes in Sertoli cells only, in order to study Sertoli cell gene function during mammalian spermatogenesis. PMID- 12124944 TI - Efficient in vitro and in vivo excision of floxed sequences with a high-capacity adenoviral vector expressing Cre recombinase. AB - Conditional gene expression or gene disruption using Cre/loxP- or FLP/frt-based recombination systems are valuable tools for studying gene function in development and disease. Recombinant adenoviral vectors expressing Cre recombinase have been suggested as an alternative for deletion of floxed sequences. To further improve this approach we generated a high-capacity adenoviral (HC-Ad) vector expressing Cre (HC-Adcre). In this vector all viral coding sequences are deleted resulting in decreased toxicity. In the present study HC-Adcre efficiently mediated recombination between two loxP sites located in the genome of a reporter cell line. When intravenously injected into ROSA26 reporter mice, a floxed sequence was excised in hepatocytes resulting in expression of the beta-gal reporter. Our data indicate that HC-Ad vectors expressing Cre effectively delete floxed sequences in vivo and have a significant potential as a tool for functional studies in mice. PMID- 12124945 TI - Targeting mammary epithelial cells using a bacterial artificial chromosome. AB - We describe the generation of transgenic mouse lines expressing Cre recombinase in epithelial cells of the lactating mammary gland. As an expression vector, we used a P1-derived bacterial artificial chromosome (PAC) which harbors the gene for the secretory milk protein, whey acidic protein (Wap). Using homologous recombination in E. coli, the PAC was modified to carry the improved coding sequence of Cre recombinase (iCre). Transgenic lines carrying the WAPiCre PAC express Cre recombinase efficiently in the majority of mammary epithelial cells upon lactation. Of only four transgenic lines produced, three express Cre recombinase to a high efficiency. LoxP-flanked DNA sequences are recombined in virtually all epithelial cells of WAPiCre transgenic mice at lactation day 3. PMID- 12124947 TI - Functional genomics: a rose by another name. PMID- 12124946 TI - SOX9 has both conserved and novel roles in marsupial sexual differentiation. AB - In addition to an essential role in chondrogenesis, SOX9 is a highly conserved and integral part of the testis determining pathway in human and mouse. To determine whether SOX9 is involved in sex determination in noneutherian mammals we cloned a marsupial orthologue and studied its expression. The tammar wallaby SOX9 gene proved to be highly conserved, and maps to a region of the tammar genome syntenic to human chromosome 17. Marsupial SOX9 transcripts were detected by RT-PCR in the developing limb buds and both the developing ovary and testis from the first sign of gonadal development through to adulthood. Northern blot, in situ hybridisation, and immunohistochemical analyses showed that SOX9 reaches high levels of expression in the developing testis, where it is confined to the Sertoli cell nuclei, and the brain. This is similar to the expression pattern seen in human and mouse embryos and is consistent with a conserved role for SOX9 in vertebrate brain, skeletal, and gonadal development. In addition, SOX9 was expressed in the developing scrotum and mammary gland primordium regions of the tammar up to the time of birth. SOX9 protein was also detected in the developing Wolffian duct epithelium in the male mesonephros. These previously undescribed locations of SOX9 expression suggest that SOX9 may play additional roles in the differentiation of the marsupial reproductive system. PMID- 12124948 TI - Genetic screen for modifiers of the rough eye phenotype resulting from overexpression of the Notch antagonist hairless in Drosophila. AB - Hairless was identified as antagonist in the Notch signaling pathway based on genetic interactions. Molecularly, Hairless inhibits Notch target gene activation by directly binding to the Notch signal transducer Su(H). Additional functional domains apart from the Su(H) binding domain, however, suggest additional roles for the Hairless protein. To further our understanding of Hairless functions, we have performed a genetic screen for modifiers of a rough eye phenotype caused by overexpression of Hairless during eye development. A number of enhancers were identified that comprise mutations in components of Notch- and EGFR-signaling pathways, some unknown genes and the gene rugose. Mutant alleles of rugose display manifold genetic interactions with mutants in Notch and EGFR signaling pathway components. Accordingly, the rugose eye phenotype is rescued by Hairless and enhanced by Delta. Molecularly, interactions might occur at the protein level because rugose appears not to be a direct transcriptional target of Notch. PMID- 12124949 TI - Clinical results for the training-phase roll-in patients in the Intracoil femoralpopliteal stent trial. AB - The purpose of this study was to report the results of the roll-in patients for the multicenter IntraCoil trial in the femoropopliteal arteries at 9-month follow up. Ninety-three roll-in patients (mean age, 67.8 +/- 10.5 years; 62.4% male gender) constituted the learning phase for the 22 clinical sites. Obstructive femoropopliteal artery disease up to 15 cm was treated with stenting. Clinical patency was measured over a 9-month period by clinical and hemodynamic data as well as the Rutherford scale. Diabetes mellitus was present in 35.5%. Twenty-nine percent of lesions treated were occlusions. The mean reference diameter for treated lesions was 4.27 +/- 1.11 mm, while the mean lesion length was 3.83 +/- 3.69 cm. Acute angiographic success by operator evaluation was obtained in 98.9% of patients. Major complications occurred in 3.2%. No patient experienced abrupt or subacute closure. There were two reports of failure to deliver assigned stent. In both incidences, the stent was removed without surgical intervention. There were also three reports of stent misplacement (moving during delivery), one report of stent migration, one report of delivery system failure. At 30-day and 9 month follow-up of successfully treated patients, 100%/77.9% remained free of major adverse clinical events (MACE) and 100%/81.8% target lesion revascularization (TLR), respectively. At 9-month follow-up, ankle-brachial index increased from 0.66 +/- 0.22 to 0.83 +/- 0.20 while mean maximum walking time increased form 4.47 +/- 3.02 to 5.91 +/- 3.97 min. The use of the IntraCoil stent appears to have a short learning curve. Excellent clinical and hemodynamic patency is seen at 9 months. The randomized comparison trial comparing the IntraCoil to angioplasty is pending. PMID- 12124950 TI - Hope springs eternal: the unfulfilled promise of femoropopliteal stenting. PMID- 12124951 TI - Effective plaque removal with a new 8 French-compatible atherectomy catheter. AB - The purpose of this study was to evaluate the safety and efficacy of the new 8 Fr guide catheter-compatible Flexicut directional atherectomy device and to compare it with the conventional Atherocath GTO catheter. The 6 Fr Flexicut catheter has a larger cutting window and a titanium nitride-coated cutter to effect more tissue removal as well as treat mildly calcified lesions. A group of 143 lesions in 117 consecutive patients treated with the Flexicut catheter in four centers were compared with a control group of 277 lesions in 212 consecutive patients treated with the GTO device. Postatherectomy luminal diameters were larger (2.92 +/- 0.79 vs. 2.52 +/- 0.64 mm; P < 0.0001), with more luminal gain (relative gain: 0.58 +/- 0.24 vs. 0.48 +/- 0.25; P = 0.0007) using fewer directional coronary atherectomy (DCA) cuts (12 +/- 7 vs. 16 +/- 9; P = 0.0001) in the Flexicut group. A residual diameter stenosis < 20% immediately after DCA was obtained in 77% of the lesions in the Flexicut group vs. 45% in the GTO group (P < 0.0001). Histology in the former group revealed large calcium speckles in the retrieved specimens. In the Flexicut group, there was a lower incidence of access site complications and damage to the coronary ostium (2.5% vs. 7.5%; P = 0.08). The new Flexicut catheter is more effective than the conventional GTO catheter with a trend for reduced guiding catheter-related complications. PMID- 12124952 TI - Intra-atrial thrombolytic therapy for dissolution of chronic left atrial thrombi in patients undergoing balloon mitral commissurotomy. AB - The purpose of this study was to evaluate the solubility of left atrial thrombi to thrombolytics after failure of long-term anticoagulant therapy in patient with mitral stenosis. One hundred and eighty-one consecutive patients with mitral valve area < or = 1.5 cm(2) and without severe mitral regurgitation were screened with echocardiography; 30 were found to have left atrial thrombi. Follow-up echocardiography performed 7.4 +/- 5.6 months after warfarin therapy revealed that 8/30 of patients had complete dissolution and 3/30 had partial dissolution of the thrombi. Thirteen patients with residual isolated appendageal thrombi underwent balloon mitral commissurotomy and were randomized into four groups at the end of balloon mitral commissurotomy: group A, receiving intra-atrial infusion of heparin and tissue plasminogen activator (t-PA; n = 4); group B, heparin and streptokinase (n = 3); group C, heparin (n = 3); and group D, acting as control (n = 3). It was found that only two patients in the t-PA group had their thrombi either completely or partially dissolved within 48 hr. Thus, this study suggests that t-PA may have the potential of dissolving chronic left atrial thrombi. PMID- 12124953 TI - Elderly patients have a favorable outcome after intracoronary radiation for in stent restenosis. AB - Intracoronary radiation therapy (IRT) reduces recurrent in-stent restenosis (ISR) by inhibition of smooth muscle cell proliferation. The ability of these cells to replicate is limited with age due to changes in the telomeres. The purpose of this study was to assess the effect of age on outcomes following IRT for ISR. We evaluated 1,088 patients with 6-month clinical follow-up who were enrolled in radiation trials for ISR using gamma- and beta-emitters. Patients were analyzed within and between IRT (n = 861) or placebo therapy (n = 227) in four age groups (< 55, 55-65, 66-75, > 75 years). Baseline characteristics were similar within each age group of IRT patients, except elderly patients (> 75 years) had a lower rate of diabetes (28% in patients > 75 years; P = 0.008) and a higher rate of previous CABG (59% in patients > 75 years; P < 0.001). The rate of target lesion revascularization (TLR) was reduced in the elderly. TLR at 6 months was 18% in patients < 55 years, 21% in 55-65 years, 12% in 66-75 years, and 10% in patients > 75 (P = 0.009). The MACE rate at 6 months was 21% in patients < 55 years, 29% in 55-65 years, 26% in 66-75 years, and 17% in patients > 75 (P = 0.03). No effect of age was seen in placebo patients. IRT-treated patients had reduced MACE compared to placebo in all age groups, driven by reduced target vessel revascularization. Age was an independent predictor of MACE at 6 months (OR = 0.8; CI = 0.70-0.93; P = 0.004). Angiographic restenosis was not clearly associated with need for TLR in patients > 75 years. In elderly patients (> 75 years) treated with IRT for ISR, the rate of TLR was significantly reduced compared to younger patients. However, this reduction in TLR was not associated with a reduction in angiographic restenosis, suggesting that TLR should not be used as a surrogate for angiographic evaluation. PMID- 12124954 TI - Age paradox with intracoronary radiation: rays of hope for the elderly. PMID- 12124955 TI - Platelet inhibition with tirofiban early during percutaneous coronary intervention: dosing revisited. AB - Platelet inhibition is central to the efficacy of the intravenous glycoprotein IIb/IIIa receptor inhibitors. Differences in the degree of platelet inhibition achieved with these agents may account for the disparity in clinical efficacy noted in recently completed clinical trials. The purpose of this study was to evaluate ex vivo platelet inhibition with tirofiban in patients admitted with acute coronary syndrome and who were referred for percutaneous coronary interventions. Twenty-five patients were studied. Ten patients received tirofiban 10 microg/kg bolus and 0.15 microg/kg infusion for 16 hr. Platelet inhibition was determined at 5, 15, 30, 45, 60, and 120 min after tirofiban, by light transmission aggregometry (LTA), rapid platelet function assay (RPFA), and platelet works (PW). The average platelet inhibition using RPFA with PPACK, was 87% at 5 min, then decreased to a nadir of 72% at 30 min and recovered back to > 80% at 60 min and onward. Similar trends were noted with RPFA-citrate, PW, and LTA. Ca-chelating anticoagulants (EDTA and citrate) overestimated platelet inhibition at all time points. Dose adjustment was done by increasing the bolus (15 microg/kg) in five patients, increasing the maintenance dose (0.2 microg/kg/min) in five patients, and increasing both the bolus and the maintenance dose in another five patients. Platelet inhibition tested by all the above methods was consistently over 90% when both the bolus and maintenance doses were increased. No increased incidence of major bleeding was noted at this adjusted dose. The current dosing of tirofiban may be inadequate to achieve appropriate platelet inhibition during PCI in patients admitted with acute coronary syndrome and receiving tirofiban immediately before the procedure in the cardiac catheterization laboratory. Dose adjustment may be needed to maximize platelet inhibition early during PCI. PMID- 12124956 TI - Stent design-related coronary artery remodeling and patterns of neointima formation following self-expanding and balloon-expandable stent implantation. AB - The self-expanding Wallstent (WS) and balloon-expandable Palmaz-Schatz stents (PS) display different mechanical and dynamical stent properties. We analyzed the impact of the respective stent design on coronary wall geometry using quantitative coronary angiography (QCA) and intracoronary ultrasound (ICUS) measurements. Serial measurements were performed within the stent and within reference segments of 50 patients (25 WS, 25 PS). Relative changes for each parameter in both stent designs were calculated (Mann-Whitney U-test; 95% CI). The luminal net gain in WS was not significantly higher in WS compared with PS (1.63 +/- 1.11 vs. 1.44 +/- 0.63 mm; P = 0.2554). The respective loss indexes were also similar (0.38 +/- 0.42 vs. 0.36 +/- 0.23; P = 0.8578). The WS segments showed significant postinterventional stent expansion with positive vessel remodeling. The neointima formation was significantly higher in WS segments (4.23 +/- 2.07 vs. 2.22 +/- 2.22 mm(2)). The coronary wall morphology and stent geometry after 6.5 +/- 1.2 months are related to the stent design. In WS segments, the neointima formation was balanced by postinterventional stent expansion, resulting in a comparable relative lumen loss in both stent types. The respective stent design had no impact on the vessel reference segments. PMID- 12124957 TI - Self-expanding stent deployment strategies may be the key to reducing in-stent restenosis. PMID- 12124958 TI - Use of a pigtail catheter to engage a difficult internal mammary artery. AB - Increasing use of bilateral internal mammary arteries for coronary surgery will increase the number of interventions in these grafts. Such interventions may be technically challenging because of often tortuous and angulated vessels. We describe a technique to intubate an acutely angulated right internal mammary artery that was inaccessible with conventional catheters. PMID- 12124959 TI - Wag the pig. PMID- 12124960 TI - Less invasive PTCA of a gastroepiploic artery combining the transradial approach and 5 Fr guiding catheter: a case report. AB - The gastroepiploic artery has been successfully used as an arterial conduit in selected patients undergoing CABG with acceptable immediate and long-term results. Myocardial ischemia may occur during the follow-up period as a result of spasm, occlusion, or stenosis at the anastomosis site. Because of tortuosity and in order to avoid graft spasm and to obtain good extra backup support, we require low-profile wide-lumen guiding catheters for deep intubation and increased procedural success. In the case presented here, a gastroepiploic graft stenosis was treated by balloon angioplasty performed through a less invasive approach combining transradial access and use of 5 Fr guiding catheter. PMID- 12124961 TI - Carotid stent-supported angioplasty in a patient with symptomatic and critical aortic stenosis. AB - We report a successful carotid angioplasty in a patient with symptomatic and critical aortic stenosis. The safety and feasibility of this technique in this scenario have not been well established and reported. With careful planning and technique, we believe the procedure should be safe and feasible. PMID- 12124962 TI - Delayed occurrence of complete heart block without warning after alcohol septal ablation for hypertrophic obstructive cardiomyopathy. PMID- 12124963 TI - Catheter closure of perimembranous ventricular septal defects using the new Amplatzer membranous VSD occluder: initial clinical experience. AB - The surgical closure of membranous ventricular septal defects (VSDs) is associated with morbidity and low mortality. Six patients with VSDs located in the membranous part of the ventricular septum underwent an attempt of catheter closure using a new device specifically designed for the membranous septum. Patients ranged in age from 3.5 to 19 years (median, 10.5 years) and in weight from 15 to 45 kg (median, 29 kg). One patient with associated pulmonary valve stenosis had shortness of breath. The median Qp/Qs ratio was 1.6 (range, 1.1-3) and the median left ventricle end-diastolic dimension (LVEDD) was 44 mm (range, 38-52 mm). The devices were deployed via the femoral vein using 7-8 Fr sheaths. There was immediate complete closure in all patients. One patient developed trivial aortic regurgitation. There were no other complications. The median fluoroscopy time was 15.5 min (range, 10.3-53.4 min). At 24 hr, all patients were doing well. The median LVEDD decreased to 38 mm (range, 34-47 mm). One patient continued to have trace aortic regurgitation. All patients were discharged home after 24 hr. Transcatheter occlusion of membranous VSDs is safe and effective. Further clinical trials are underway to assess the long-term safety and results. PMID- 12124964 TI - Anterograde balloon valvuloplasty for the treatment of neonatal critical valvar aortic stenosis. AB - We report our experience with anterograde balloon valvuloplasty in 17 neonates treated between November 1996 and June 2001 for critical aortic stenosis. Patients with hypoplastic left heart syndrome were excluded. Anterograde balloon valvoplasty of the aortic valve was possible in all 17 patients. The mean peak systolic gradient prior to cardiac catheterization was 73 mm Hg (range, 30-117 mm Hg) and decreased to 37 mm Hg (range, 21-60 mm Hg) after the dilation. Aortic regurgitation after balloon valvoplasty was absent or mild in 14/17 patients, moderate in 2 patients, and severe in 1 patient. There was no mortality or echocardiographic evidence for aortic cusp perforation or mitral regurgitation associated with the procedure. Redilation was necessary in 3/17 patients. Two patients are awaiting elective Ross operation. One patient with endocardial fibroelastosis died at 11 months of age. Anterograde balloon valvoplasty can be safely and effectively performed to palliate neonates with critical aortic valve stenosis. PMID- 12124967 TI - Should PFO closure devices be used "off label"? PMID- 12124966 TI - Thrombosis of a patent foramen ovale closure device: thrombolytic management. AB - Transcatheter closure of a patent foramen ovale (PFO) offers a valuable alternative to surgery in selected patients with presumed paradoxical embolism. Like for all other devices, several complications have been described. Although the majority of these complications are caused by technical problems of the device itself, thrombosis is a relatively infrequent but feared finding, as it may result in recurrent embolic events and persistent neurological deficits. The majority of the complications after implantation are dealt with by surgical removal of the device and subsequent surgical closure of the PFO. The present case report describes biatrial thrombosis of a PFO device that was successfully treated with thrombolytics and GP IIb/IIIa receptor blockers. In stable patients, this pharmaco-therapeutic approach should be attempted to save the thrombosed PFO device and could therefore offer a valuable alternative to surgical removal of the device. PMID- 12124968 TI - Nonobstructive ASD creation to qualify patients for the Fontan operation: effects on pulmonary hypertension due to restrictive left atrioventricular valve and interatrial communication. AB - Pulmonary hypertension (PH) due to elevated left atrial pressure (LAp) caused by restrictive left atrioventricular valve and interatrial communication sometimes precludes patients with univentricular heart from undergoing the Fontan operation. We have created atrial septal defect (ASD) for such patients to reduce LAp and then pulmonary arterial pressure (PAp) and pulmonary vascular resistance (Rp) in an attempt to qualify the patients for the Fontan repair. This study was performed to clarify the efficacy and limitation of this approach. Twelve patients with PH (mean PAp > 20 mm Hg and/or Rp > 3.0 Wood's unit/m(2)) due to obstruction at the LA exit underwent ASD creation at the age of 4.0 +/- 4.0 (mean +/- SD) years. Follow-up catheterization 14.9 +/- 15.6 months after the ASD creation demonstrated marked reductions in mean LAp (from 21 +/- 5 to 8 +/- 2 mm Hg), mean PAp (from 39 +/- 13 to 17 +/- 4 mm Hg), and Rp (from 6.2 +/- 4.5 to 2.7 +/- 1.4 Wood's unit/m(2)) compared with those before the procedure (all P < 0.0001). Seven of the 12 patients (58%) qualified for the Fontan operation (mean PAp < 20 mm Hg, Rp < 3.0 Wood's unit/m(2), and Nakata's PA index > or = 250) after the ASD creation. Final surgical outcomes of the seven patients included successful Fontan operation in five and biventricular repair in two. Patients with severe PH [mean PAp > or = 45 mm Hg (n = 6) or Rp > or = 4.5 Wood's unit/m(2) (n = 5)] before the ASD creation qualified less frequently for the Fontan repair than those with mild PH [mean PAp < 45 mm Hg (n = 6; 17% vs. 100%; P = 0.008) or Rp < 4.5 Wood's unit/m(2) (n = 7; 20 vs. 86%; P = 0.045)]. Patients with PA banded after 6 months of age (n = 3) qualified less frequently for the Fontan operation than those banded before 6 months of age (n = 6; 0 vs. 83%; P = 0.048). These data suggest that ASD creation is an effective approach to qualify patients for the Fontan operation in the presence of restrictive left atrioventricular valve and interatrial obstruction, except those with longstanding severe PH following delayed PA banding. PMID- 12124969 TI - Patterns of late diameter change after balloon angioplasty of branch pulmonary artery stenosis: evidence for vascular remodeling. AB - Angiographic diameters of 36 pulmonary artery stenoses (26 patients; median age, 3.3 years) before and after balloon angioplasty and at repeat angiography after 2 64 months were compared to diameters of 31 untreated pulmonary artery stenoses (20 patients) at a median age of 3.6 years and after 4-76 months. In the treatment group, an acute diameter gain of > 50% was achieved in 58%. On follow up, 16 lesions remained unchanged, 6 lesions had > 20% late loss, and 12 lesions had > 20% late gain. The three patterns of vascular response were confirmed when compared to the control group. In the late gain group, overall diameter increase was 125% compared to 41% initial increase. The net result was a long-term success rate of 57%. Patterns of late diameter change appear to suggest vascular remodeling after balloon angioplasty of pulmonary arteries. PMID- 12124970 TI - Initial experience with intratherapeutics Intrastent Doublestrut LD stents in patients with congenital heart defects. AB - Limitations of the currently available balloon-expandable stainless steel (BE-SS) stents for use in patients with congenital heart defects (CHD) include inflexibility, significant shortening with expansion, sharp ends, and limited sizes. These limitations increase risk and greatly effect success of stent placement in these patients. The recently approved IntraStent DoubleStrut LD (IS LD) stents are BE-SS stents designed to address such limitations. We report our initial experience with IS-LD stents in patients with CHD. Using standard techniques, 36 stents were implanted in 22 patients whose median age was 11 years (range, 1.4-35 years) and weight was 33 kg (range, 9-96.8 kg). Lesions stented included aortic coarctation (4), branch pulmonary arteries (19), inferior or superior vena cava (11), Mustard baffle (1), and Fontan baffle (1). All attempts at stent placement were successful. Hemodynamic assessment and angiography was performed pre- and poststent placement in all patients. Intravascular ultrasound was performed in four patients. Stenosis diameter increased from 5.7 +/- 0.6 mm (mean +/- SD) to 11.6 +/- 0.5 mm (P < 0.001). Stents did not shorten significantly with dilation (median percent shortening, 3.8%; P > 0.1). Complications included stent recoil (2), stent migration (1), and stent distortion (1). IS-LD stents are a safe and effective treatment for the majority of vascular stenoses in patients with CHD. Stent recoil at large diameters may limit its usefulness in selected lesions. Follow-up studies are required to determine the long-term performance of IS-LD stents. PMID- 12124971 TI - Rotational ablation and stent placement for severe calcific coronary artery stenosis after Kawasaki disease. AB - We report on a 5-year-old child who had an episode of Kawasaki disease with giant coronary artery aneurysms at the age of 4 months. Surveillance coronary angiography showed severe calcific stenosis in the proximal left anterior descending artery. Balloon angioplasty failed to resolve the obstruction. Rotational ablation was therefore performed. Surveillance angiogram performed 6 months after rotational ablation showed critical restenosis. Rotational ablation was therefore repeated, followed by stent placement. To the best of our knowledge, this is the youngest child who has undergone coronary stenting after Kawasaki disease. PMID- 12124972 TI - Caught between a rock and a hard place? It is good to have some help. PMID- 12124973 TI - Femoropopliteal occlusive disease: diagnosis, indications for treatment, and results of interventional therapy. AB - The relative role of percutaneous therapy versus bypass surgery for femoropopliteal occlusive disease (FPD) represents an area of great controversy in the field of vascular medicine. This controversy is complicated by the very dynamic therapeutic options that continue to be introduced, particularly on the endovascular side. The purpose of this review is to provide an overview of the current diagnostic modalities available to characterize the level and degree of vascular insufficiency and summarize the currently available endovascular therapeutic options with their clinical role and outcomes. Finally, future therapeutic advances that will soon be evaluated in clinical trials will be discussed. PMID- 12124974 TI - Coronary artery stents: evaluating new designs for contemporary percutaneous intervention. AB - Intracoronary stents have markedly improved the short- and long-term safety and efficacy of percutaneous coronary intervention by improving acute gains in luminal dimensions, decreasing abrupt vessel occlusion, and reducing restenosis. At present, nearly 90% of all coronary interventions involve stenting. A variety of advances in stent technology and design have expanded the clinical application of stenting to include complex coronary lesions, multivessel disease, and small diameter vessels. In addition, the development of stents as drug delivery systems for antithrombotic or antiproliferative agents has the potential to expand the role of coronary stenting, and early clinical experience appears promising. The purpose of this review is to describe recent developments in stent design, examine the results of clinical trials of contemporary stents, and present future directions for investigation of new stent technologies. PMID- 12124977 TI - Development and characterization of a minimal inducible packaging cell line for simian immunodeficiency virus-based lentiviral vectors. AB - BACKGROUND: Lentiviral vectors allow gene transfer into non-dividing cells. Further development of these vector systems requires stable packaging cell lines that enable adequate safety testing. METHODS: To generate a packaging cell line for vectors based on simian immunodeficiency virus (SIV), expression plasmids were constructed that contain the codon-optimized gag-pol gene of SIV and the gene for the G protein of vesicular stomatitis virus (VSV-G) under the control of an ponasterone-inducible promoter. Stable cell lines expressing these packaging constructs were established and characterized. RESULTS: The RT activity and vector titers of cell clones stably transfected with the inducible gag-pol expession plasmid could be induced by ponasterone by more than a factor of 1000. One of these clones was subsequently transfected with the ponasterone-inducible VSV-G expression plasmid to generate packaging cells. Clones of the packaging cells were screened for vector production by infection with an SIV vector and subsequent induction by ponasterone. In the supernatant of selected ponasterone induced producer clones vector titers of more than 1x10(5) transducing units/ml were obtained. Producer cell clones were stable for at least five months, as tested by vector production. CONCLUSIONS: The packaging cells described should be suitable for most preclinical applications of SIV-based vectors. By avoiding regions of high homology between the vector and the packaging constructs, the design of the SIV packaging cell line should reduce the risk of transfer of packaging genes to target cells and at the same time provide flexibility with respect to the SIV vector constructs that can be packaged. PMID- 12124978 TI - Genetic retargeting of adenovirus vectors: functionality of targeting ligands and their influence on virus viability. AB - BACKGROUND: We studied the ability of adenovirus type 5 (Ad5) to encapsidate new cellular ligands carried by their fibers to yield functional retargeted vectors for gene therapy. Recombinant Ad5 fibers containing shaft repeats 1 to 7 and an extrinsic trimerization motif, and terminated by its native knob or amino acid motifs containing RGD, have been rescued into infectious virions. METHODS: Polypeptide ligands of cell surface molecules, including single-chain antibodies or epidermal growth factor, were cloned into recombinant fibers. Phenotypic analysis of fiber constructs and rescuing into the Ad5 genome were performed. Recombinant viruses were characterized with reference to fiber content, growth rate and infectivity. RESULTS: A major limiting factor for recovering viable recombinant Ad5 carrying fiber-fused polypeptide ligands was apparently the ability of the ligand to fold correctly within the cellular cytoplasm. This constraint has previously not been systematically evaluated in the literature. Phenotypic analysis of the fiber-ligand fusions showed that their degree of cytoplasmic solubility correlated with their ability to yield viable Ad5 vectors. Our results suggested that the fiber manipulations diminish virus growth rate, probably through different, opposing effects: (i) the reduced shaft length increases fiber solubility in the absence of the knob but (ii) diminishes virus entry, and (iii) the absence of the knob alters the overall protein composition of the virion and decreases its fiber copy number. CONCLUSIONS: Based on our findings, cytoplasmic solubility and cytoplasmic ligand reactivity of fiber ligand fusion proteins are the best prediction criterion for viability and recovery of genetically retargeted Ad vectors. PMID- 12124979 TI - Evaluation of angiogenesis and side effects in ischemic rabbit hindlimbs after intramuscular injection of adenoviral vectors encoding VEGF and LacZ. AB - BACKGROUND: Recent studies have suggested the therapeutic potential of vascular endothelial growth factor (VEGF) gene therapy in ischemic skeletal muscle. However, only limited information is available about the effects of VEGF gene therapy in different regions of ischemic limbs, effects of control adenoviruses, and biodistribution of the transgenes after intramuscular (i.m.) administration. Here we studied angiogenesis and side effects of adenovirus-mediated VEGF and beta-galactosidase (LacZ) gene transfers in ischemic rabbit hindlimbs. METHODS AND RESULTS: Ten days after induction of ischemia, rabbits were treated with i.m. injections of saline, LacZ adenovirus (AdLacZ; 2x10(10) pfu) or adenovirus encoding mouse VEGF(164) (AdVEGF; 2x10(10) pfu). In rabbits treated with AdVEGF an increase in serum VEGF(164) levels was detected by ELISA three and seven days after the gene transfer. 30 days after the gene transfer a positive effect on capillary density was observed in the thigh region both in rabbits treated with AdVEGF and AdLacZ compared with animals that received saline. On the other hand, AdVEGF and AdLacZ gene transfers had no effect on the capillary density in the calf region on day 30. A positive correlation between the capillary density and the number of collateral arteries was observed in the thigh. Hindlimb and testis edema and excess non-physiological growth of capillaries were detected as adverse effects of the AdVEGF gene therapy. Biodistribution analysis showed that the transgene was present not only in the target muscle, but also in ectopic tissues seven days after i.m. gene transfer. CONCLUSIONS: The results suggest that a high dose of adenoviral vector encoding either AdVEGF or AdLacZ induces angiogenesis in the rabbit hindlimb ischemia model; i.m. injection of adenovirus leads to the transfection of ectopic organs; and AdVEGF gene transfer induces edema in ischemic skeletal muscle. PMID- 12124980 TI - Antisense MBD2 gene therapy inhibits tumorigenesis. AB - BACKGROUND: Aberration in the pattern of DNA methylation is one of the hallmarks of cancer. We present data suggesting that dysregulation of MBD2, a recently characterized member of a novel family of methylated DNA binding proteins, is involved in tumorigenesis. Two functions were ascribed to MBD2, DNA demethylase activity and repression of methylated genes. METHODS: Multiple antisense expression and delivery systems, transfection, electrotransfer and adenoviral were employed to demonstrate that MBD2 is essential in tumorigenesis, both ex vivo and in vivo. RESULTS: Inhibition of MBD2 by antisense expression resulted in inhibition of anchorage-independent growth of antisense transfected cancer cells or cells infected with an adenoviral vector expressing MBD2 antisense. Xenograft tumors treated with an adenoviral vector expressing MBD2 antisense or xenografts treated with electrotransferred plasmids expressing MBD2 antisense showed reduced growth. CONCLUSIONS: These results support the hypothesis that one or both of the functions described for MBD2 are critical in tumorigenesis and that MBD2 is a potential anticancer target. PMID- 12124981 TI - Prenatal human ocular degeneration occurs in Leber's congenital amaurosis (LCA2). AB - BACKGROUND: Leber's congenital amaurosis (LCA) encompasses the most precocious and severe forms of inherited retinal dystrophy, displaying very significant visual handicap at or soon after birth. Among the currently identified mutations, alterations in the gene coding for retinal pigment epithelium 65-kDa protein (RPE65) lead to LCA2. Existing animal models for LCA2 (RPE65(-/-) null mice and naturally occurring RPE65(-/-) Briard dogs) exhibit near normal retinal histology at birth, although no recordable photofunction can be detected. Structural degeneration in both cases occurs with delayed onset, cone death generally preceding that of rods. METHODS: We obtained retinal tissue from a voluntarily aborted embryo of an LCA2 carrier in order to compare histopathology and immunohistochemistry with age-matched normal foetal retina. RESULTS: Compared to normal retinas, affected retina displayed cell loss and thinning of the outer nuclear (photoreceptor) layer, decreased immunoreactivity for key phototransduction proteins, and aberrant synaptic and inner retinal organisation. The gene mutation abolished detectable expression of RPE65 within the retinal pigment epithelium (RPE) of affected eyes, and ultrastructural examination revealed the presence of lipid and vesicular inclusions not seen in normal RPE. In addition, mutant eyes demonstrated thickening, detachment and collagen fibril disorganisation in the underlying Bruch's membrane, and the choroid was distended and abnormally vascularised, in comparison with controls. CONCLUSIONS: Such data contrast with the late-onset ocular changes observed in animal models, indicating caution should be exercised when inferring human retinal pathophysiology from information based on other species. PMID- 12124983 TI - Cyanide bystander effect of the linamarase/linamarin killer-suicide gene therapy system. AB - BACKGROUND: The killer-suicide system linamarase/linamarin (lis/lin) uses the plant gene linamarase (beta-glucosidase) to convert the cyanogenic glucoside substrate, linamarin, into glucose and cyanide. We have studied the bystander effect associated with this new system mediated by the production of the cyanide ion that diffuses freely across membranes. METHODS: Immunofluorescent staining of cells treated with an anti-linamarase antibody allowed us to localize the enzyme within the cells. Flow cytometry was used to determine the sensitivity of different mixtures of cells, C6lis and C6gfp (green), to linamarin as a percentage of cell survival. RESULTS: We demonstrate here that rat glioblastoma C6 cells carrying the linamarase gene (lis), mixed with naive C6 cells and exposed to linamarin, induce generalized cell death. Cells expressing lis efficiently export linamarase, whereas linamarin enters cells poorly by endocytosis; as a result most of the cyanide is produced outside the cells. The study was facilitated by the presence of the green fluorescent protein (gfp) gene in the bystander population. As few as 10% C6lis-positive cells are sufficient to eliminate the entire cell culture in 96 h. CONCLUSIONS: This bystander mechanism does not preferentially kill toxic metabolite producer cells compared with bystander cells, thus allowing production of sufficient cyanide to cause tumor regression. In this report we confirm the potential of the lis/lin gene therapy system as a powerful tool to eliminate tumors in vivo. PMID- 12124984 TI - Cationic lipids derived from glycine betaine promote efficient and non-toxic gene transfection in cultured hepatocytes. AB - BACKGROUND: The low efficiency and toxicity of transfection in a primary culture of hepatocytes using cationic lipids remains a limiting step to the study of gene function and the setting up of non-viral gene therapy. METHODS: A novel class of cationic lipids (GBs) derived from natural glycine betaine compounds covalently linked to acyl chains by enzymatically hydrolysable peptide and ester bonds, a structure designed to reduce cytotoxicity, was used to improve transfection efficiency in a primary culture of rat hepatocytes. The relationship between lipid structure, lipoplex formulation and transfection efficiency was studied using six GBs (12-14-16, 22-24-26) varying in their spacer and acyl chains. RESULTS: GB12, characterized by short [(CH(2))(10)] acyl chains and spacer, allowed plasmid uptake in all cells and reporter gene expression in up to 40% of hepatocytes with a low cytotoxicity, a much higher efficiency compared with transfections using other reagents including Fugene6 and Lipofectin. We also showed that numerous cells accumulated high amounts of plasmids demonstrating that GB12 promoted a very efficient DNA transfer through plasma membrane leading to an increase in nuclear plasmid translocation, allowing a much higher gene expression. Moreover, GB12-transfected hepatocytes survived to injection in normal livers and were found to express the LacZ reporter gene. CONCLUSIONS: The non-toxic GB12 formulation is a powerful vehicle for plasmid delivery in cultured hepatocytes with relevance in liver gene therapy. PMID- 12124982 TI - Human cell lines engineered for tetracycline-regulated expression of tumor suppressor candidate genes from a frequently affected chromosomal region, 3p21. AB - BACKGROUND: We modified a tetracycline-regulated system that can control the activity of individual genes quantitatively and reversibly in transgenic mammals. Despite these advances, there remained one problem in the intensive use of the tet-system: the limited range of acceptor cell lines, expressing a tetracycline controlled transcriptional activator (tTA). This study describes in detail new vectors and a unifying strategy to generate tTA-expressing cell lines. METHOD: Two retroviral vectors pLNCtTA-hCMV and pLNCtTA-EF1alpha coding for the tTA were used to engineer cell lines to constitutively express tTA. New expression vectors pETE-Hyg and pETE-Bsd were also created that replicate in episomal form in human cells and facilitate tetracycline-regulated expression of targeted genes. RESULTS: The primate-tropic retroviruses efficiently delivered the regulatory tTA gene into 12 selected human cancer cell lines. Two candidate tumor suppressor genes from the human 3p21-p22 region MAPKAPK3 (3pK) and MLH1 were cloned into the episomal vector and transfected into engineered A9 and KRC/Y cells. The transfectants were subcutaneously grown in SCID mice, and the expression of the transgene was successfully controlled in vivo by tetracycline administered ad libitum in drinking water. The experiments demonstrated that both transgenes did not antagonize the tumorous growth of these cells. CONCLUSIONS: New retroviral and episomal vectors appear particularly suited for tight regulation of genes that cause suppression of cell growth. The generated cell lines can be used in various applications to study the effect of an inducible transgene in human cancer cells. PMID- 12124985 TI - Preparation of dry powder dispersions for non-viral gene delivery by freeze drying and spray-drying. AB - BACKGROUND: Dry powder dispersion devices offer potential for delivering therapeutic macromolecules to the pulmonary epithelia. Previously, freeze-drying (lyophilisation) has been the accepted method for preparing dried formulations of proteins and non-viral gene vectors despite the respirability of such powders being inadequate without further processing. In this study we compare the utility of freeze-drying and spray-drying, a one-step process for producing dry and respirable powders, as methods for preparing non-viral respiratory gene delivery systems. METHODS: Lipid:polycation:pDNA (LPD) vectors comprising 1,2-dioleoyl-3 trimethylammoniumpropane (DOTAP), protamine sulphate and pEGFP-N1 in 3% lactose solution were either snap-frozen and lyophilised or spray-dried. Lyophilised powder was used as recovered or following coarse grinding. Structural integrity of dehydrated pDNA was assessed by agarose gel electrophoresis and powder particle size determined by laser diffraction. The apparent structure of the systems was visualised by scanning and transmission electron microscopy with the biological functionality quantified in vitro (A549 human lung epithelial cell line) by Green Fluorescent Protein (GFP) associated fluorescence. RESULTS: Lyophilisation produced large, irregularly shaped particles prior to (mean diameter approximately 21 microm) and following (mean diameter approximately 18 microm) coarse grinding. Spray-drying produced uniformly shaped spherical particles (mean diameter approximately 4 microm). All dehydrated formulations mediated reporter gene expression in A549 cells with the spray-dried formulation generally proving superior even when compared with freshly prepared LPD complexes. Biological functionality of the LPD dry powders was not adversely affected following 3 months storage. CONCLUSIONS: Spray-drying has utility for producing stable, efficient and potentially respirable non-viral dry powder systems for respiratory gene delivery. PMID- 12124986 TI - Evaluation of a porcine model for pulmonary gene transfer using a novel synthetic vector. AB - BACKGROUND: The pig lung, given its gross anatomical, histological and physiological similarities to the human lung, may be useful as a large animal model, in addition to rodents, in which to assess the potential of vectors for pulmonary airway gene transfer. The aim of this study was to assess the utility of the pig lung as a model of gene transfer to the human lung with a synthetic vector system. METHODS: The LID vector system consists of a complex of lipofectin (L), integrin-binding peptide (I) and plasmid DNA (D). LID complexes containing a beta-galactosidase reporter gene under a CMV promoter or a control plasmid at1 mg/3 ml PBS, or 3 ml buffer, was administered to the right lower lobe of the pig lung through a bronchoscope. Pigs were culled at 48 h and lung sections prepared for immunohistochemical and histological analysis. Bronchoalveolar lavage fluid was collected and analysed for TNF-alpha by ELISA. RESULTS: Immunohistochemical staining for the beta-galactosidase reporter gene indicated high efficiency of gene transfer by the LID vector to pig bronchial epithelium with 46% of large bronchi staining positively. There was no evidence for vector-specific inflammation assessed by leukocytosis and cytokine production. CONCLUSIONS: This study demonstrates the use of the pig for studies of gene transfer in the lung and confirms in a second species the potential of the LID vector for gene therapy of pulmonary diseases such as cystic fibrosis. PMID- 12124987 TI - Human liver-derived cells stably modified for regulated proinsulin secretion function as bioimplants in vivo. AB - BACKGROUND: Insulin deficiency is currently treated with pharmacological insulin secretagogues, insulin injections or islet transplants. Secondary failure of pharmacological agents is common; insulin injections often fail to achieve euglycemic control; and islet transplants are rare. Non-beta cells capable of regulated insulin secretion in vivo could be a functional cure for diabetes. Hepatocytes are good candidates, being naturally glucose-responsive, protein secreting cells, while the liver is positioned to receive direct nutrient signals that regulate insulin production. METHODS: Human liver-derived Chang cells were modified with a plasmid construct in which a bifunctional promoter comprising carbohydrate response elements and the human metallothionein IIA promoter controlled human proinsulin cDNA expression. Secretory responses of stable cell clones were characterized in vitro and in vivo by proinsulin radioimmunoassay. RESULTS: Transfected Chang cells secreted 5-8 pmol proinsulin/10(6) cells per 24 h in continuous passage for at least a year in response to 5-25 mM glucose and 10 90 microM zinc in vitro. Glucose and zinc synergistically increased proinsulin production by up to 30-fold. Non-glucose secretagogues were also active. Glucose transporter 2 (GLUT2) and glucokinase cDNA co-transfection enhanced glucose responsiveness. Intraperitoneally implanted Chang cells secreted proinsulin in scid and Balb/c mice. Serum proinsulin levels were further increased 1.3-fold (p<0.05) after glucose and 1.4- to 1.6-fold (p<0.005) after zinc administration in vivo. CONCLUSIONS: These results are the first to demonstrate stable proinsulin production in a human liver-derived cell line with activity in vitro and in vivo and provide a basis for engineering hepatocytes as in vivo bioimplants for future diabetes treatment. PMID- 12124988 TI - The UMD-LDLR database: additions to the software and 490 new entries to the database. AB - Mutations in the LDL receptor gene (LDLR) cause familial hypercholesterolemia (FH), one of the most frequent hereditary dominant disorders. The protein defect was identified in 1973, the gene was localized by in situ hybridization in 1985, and since, a growing number of mutations have been reported. The UMD-LDLR database is customized software that has been developed to list all mutations, and also to provide means to analyze them at the nucleotide and protein levels. The database has been recently modified to fulfill the recommendations of the Nomenclature Working Group for human gene mutations. However, in the current version, both the nomenclature and usual LDLR gene mutation names are reported since the latter are more commonly used. The software has also been modified to accommodate the splicing mutations and alleles that carry two nucleotide variations. The current version of UMD-LDLR contains 840 entries, of which 490 are new entries. Point mutations account for 90% of all mutations in the LDLR gene; the remaining are mostly major rearrangements, due to the presence of Alu sequences. Three new routines have been implemented in the software, thus giving users access to 13 sorting tools. In addition to the database, a Web site containing information about polymorphisms, major rearrangements, and promoter mutations is available. Both are accessible to the scientific community (www.umd.necker.fr) and should help groups working on LDLR to check their mutations and identify new ones, and greatly facilitate the understanding of functional classes/genotype relationships and of genotype/phenotype correlations. PMID- 12124989 TI - RYR1 mutations causing central core disease are associated with more severe malignant hyperthermia in vitro contracture test phenotypes. AB - Malignant hyperthermia (MH) and central core disease (CCD) are autosomal dominant disorders of skeletal muscle. Susceptibility to MH is only apparent after exposure to volatile anesthetics and/or depolarizing muscle relaxants. CCD patients present with diffuse muscular weakness but are also at risk of MH. Mutations in RYR1 (19q13.1), encoding a skeletal muscle calcium release channel (ryanodine receptor), account for the majority of MH and CCD cases. Fifteen RYR1 N-terminal mutations are considered causative of MH susceptibility, five of which are also associated with CCD. In the first extensive UK population survey, eight of 15 mutations were detected in 85 out of 297 (29%) unrelated MH susceptible cases, with G2434R detected in 53 cases (18%). Mutation type was shown to affect significantly MH phenotypes (in vitro contracture test (IVCT) response to caffeine, halothane, and ryanodine). RYR1 mutations associated with both CCD and MH (R163C, R2163H, R2435H) had more severe caffeine and halothane response phenotypes than those associated with MH alone. Mutations near the amino terminal (R163C, G341R) had a relatively greater effect on responses to caffeine than halothane, with a significantly increased caffeine:halothane tension ratio compared to G2434R of the central domain. All phenotypes were more severe in males than females, and were also affected by muscle specimen size and viability. Discordance between RYR1 genotype and IVCT phenotype was observed in seven families (nine individuals), with five false-positives and four false-negatives. This represents the most extensive study of MH patient clinical and genetic data to date and demonstrates that RYR1 mutations involved in CCD are those associated with one end of the spectrum of MH IVCT phenotypes. PMID- 12124990 TI - Assessing the relative importance of the biophysical properties of amino acid substitutions associated with human genetic disease. AB - The inclusion of a mutation in a pathology-based database such as the Human Gene Mutation Database (HGMD) is a two-stage process: first, the mutation must occur at the DNA level, then it must cause a clinically detectable disease state. The likelihood of the latter step, termed the relative clinical observation likelihood (RCOL), can be regarded as a function of the structural/functional consequences of a mutation at the protein level. Following this paradigm, we modeled in silico all amino acid replacements that could potentially have arisen from an inherited single base pair substitution in five human genes encoding arylsulphatase A (ARSA), antithrombin III (SERPINC1), protein C (PROC), phenylalanine hydroxylase (PAH), and transthyretin (TTR). These proteins were chosen on the basis of 1) the availability of a crystallographic structure, and 2) a sufficiently large number of amino acid replacements being logged in HGMD. A total of 9,795 possible mutant structures were modeled and 20 different biophysical parameters assessed. Together with the HGMD-derived spectra of clinically detected mutations, these data allowed maximum likelihood estimation of RCOL profiles for the 20 parameters studied. Nine parameters (including energy difference between wild-type and mutant structures, accessibility of the mutated residue, and distance from the binding/active site) exhibited statistically significant variability in their RCOL profiles, indicating that mutation associated changes affected protein function. As yet, however, a biological meaning could only be attributed to the RCOL profiles of solvent accessibility and, for three proteins, local energy change, disturbed geometry, and distance from the active center. The limited ability of the biophysical properties of mutations to explain clinical consequences is probably due to our current lack of understanding as to which amino acid residues are critical for protein folding. However, since the proteins examined here were unrelated, and our findings consistent, it may nevertheless prove possible to extrapolate to other proteins whose dysfunction underlies inherited disease. PMID- 12124991 TI - Novel mutations of APOB cause ApoB truncations undetectable in plasma and familial hypobetalipoproteinemia. AB - Familial hypobetalipoproteinemia (FHBL) is a genetic disorder characterized by low levels of apoB-100 and LDL cholesterol. Truncation-producing mutations of apoB (chromosome 2) are among several potential causes of FHBL in patients. Ten new families with FHBL linked to chromosome 2 were identified. In Family 8, a 4432delT in exon 26 produces a frame-shift and a premature stop codon predicted to produce a truncated apoB-30.9. Even though this truncation is just 10 amino acid shorter than the well-documented apoB-31, which is readily detectable in plasma, apoB-30.9 is undetectable. Most truncations shorter than apoB-30 are not detectable in plasma. In Family 34, an acceptor splicing mutation at position -1 of exon 14 changes the acceptor splice site AG to AA. Two families (Family 50 and 52) had mutations (apoB-9 and apoB-29) reported previously. In Family 98, a novel point mutation in exon 26 (11163T>G) causes a premature stop codon, and produces a truncated apoB-80.5 readily detectable in plasma. Sequencing of the ApoB gene in families 1, 5, 18, 58, and 59 did not reveal mutations. PMID- 12124992 TI - The molecular basis of cystathionine beta-synthase deficiency in Australian patients: genotype-phenotype correlations and response to treatment. AB - Cystathionine beta-synthase (CBS) deficiency is the most common cause of homocystinuria. It is inherited as an autosomal recessive trait and common clinical features are: dislocation of the optic lens, osteoporosis, mental retardation, and thromboembolism. We determined the molecular basis of CBS deficiency in 36 Australian patients from 28 unrelated families, using direct sequencing of the entire coding region of the CBS gene. The G307S and I278T mutations were the most common mutations. They were present in 19% and 18% of independent alleles, respectively. In total, seven novel and 20 known mutations were detected. Of those, the two novel missense mutations (C109R and G347S), as well as two known missense mutations (L101P and N228K), were expressed in E. Coli. All mutant proteins completely lacked catalytic activity. Furthermore, we studied the correlation between genotype and the biochemical response to pyridoxine treatment in the patients of whom 13 were pyridoxine responsive, 21 were non-responsive, and two were partially responsive. The G307S mutation always resulted in a severe non-responsive phenotype, whereas I278T resulted in a milder B6 responsive phenotype. From our results, we were also able to establish three other mild mutations: P49L, R369C, and V371M. PMID- 12124993 TI - Mutation analysis of the spastin gene (SPG4) in patients in Germany with autosomal dominant hereditary spastic paraplegia. AB - Hereditary spastic paraplegias (HSP) comprise a genetically and clinically heterogeneous group of neurodegenerative disorders characterized by progressive spasticity and hyperreflexia of the lower limbs. Autosomal dominant hereditary spastic paraplegia 4 linked to chromosome 2p (SPG4) is the most common form of autosomal dominant hereditary spastic paraplegia. It is caused by mutations in the SPG4 gene encoding spastin, a member of the AAA protein family of ATPases. In this study the spastin gene of HSP patients from 161 apparently unrelated families in Germany was analyzed. The authors identified mutations in 27 out of the 161 HSP families; 23 of these mutations have not been described before and only one mutation was found in two families. Among the detected mutations are 14 frameshift, four nonsense, and four missense mutations, one large deletion spanning several exons, as well as four mutations that affect splicing. Most of the novel mutations are located in the conserved AAA cassette-encoding region of the spastin gene. The relative frequency of spastin gene mutations in an unselected group of German HSP patients is approximately 17%. Frameshift mutations account for the majority of SPG4 mutations in this population. The proportion of splice mutations is considerably lower than reported elsewhere. PMID- 12124994 TI - Highly multiplexed genotyping of coronary artery disease-associated SNPs using MALDI-TOF mass spectrometry. AB - Highly multiplexed genotyping methods are needed to support a comprehensive analysis of single nucleotide polymorphisms (SNPs) in coronary artery disease (CAD)-related genes. In this study we evaluated chip-based MALDI-TOF mass spectrometry for multiplexed genotyping of SNPs associated with CAD. Our analysis included 14 healthy Japanese individuals and 19 Japanese patients with myocardial infarction whose first attack occurred before age 50. We selected 29 candidate genes involved in 1) the renin-angiotensin system, 2) lipid metabolism, 3) cytokines and adhesion molecules, 4) growth factors, and 5) the coagulation fibrinolysis system. Genotyping of candidate SNPs was performed by MALDI-TOF MS using a MassARRAY system, and 4-plex analysis was achieved at a maximum. All 39 SNPs determined by the fluorescent dye-terminator cycle sequencing method from four randomly selected patients were found to be in complete agreement with the results obtained from MassARRAY system. Significant differences were observed in the -1965delG of PAI1 (SERPINE1) with respect to allelic frequency, the G>A in the promoter region SNP in SM22 (TAGLN) for dominant genotype, and in two other SNPs (C>T in intron 1 of HGF, and -1965delG of PAI1) for recessive genotype. Three SNPs (803T>C of AGT, 677CT of MTHFR, 190T>C of ADRB3) showed weak differences in allelic frequency. MALDI-TOF-MS provided high performance with a multiplex assay design for analysis of CAD-related SNPs by increasing the throughput while maintaining a high level of accuracy. PMID- 12124995 TI - An amplification and ligation-based method to scan for unknown mutations in DNA. AB - A new approach is presented for the sensitive and selective scanning for unknown DNA mutations, based on ligation-mediated PCR and the use of the glycosylases TDG and MutY. These two highly selective enzymes together can detect about 70% of commonly observed polymorphisms and mutations in human tumors. DNA is cross hybridized to form mismatches at the positions of point mutations, de phosphorylated to eliminate any pre-existing phosphorylated DNA ends, and then exposed to enzymatic treatment to remove mismatched thymidine (TDG) or adenine (MutY). The resulting apurinic/apyrimidinic sites at the position of the mismatches are heat-converted to 5'-phosphate-containing strand breaks, the DNA is denatured, and an oligonucleotide is ligated at the position of the newly created 5'-phosphate-containing DNA ends. The ligated oligonucleotide then participates in a PCR reaction that amplifies exponentially only the mutation containing fragments. Using this method, A-->G mutations in a p53 (TP53) containing system, T-->G, G-->A, and C-->A, mutations in the Ku gene (XRCC5), and ATM, gene for a number of patient-derived genomic DNA samples have been successfully screened. This PCR-based assay is capable of detecting one mutated allele in 100 normal alleles and requires 5 to 100 ng of genomic DNA as starting material. The assay allows final visualization of the mutated fragments on a common ethidium gel or biotinylation and use in a capture format, potentially allowing the isolation of diverse mutated DNA fragments simultaneously. This versatile new approach should allow high throughput detection of DNA alterations and application in diverse areas of human mutation research. PMID- 12124996 TI - I591T MEFV mutation in a Spanish kindred: is it a mild mutation, a benign polymorphism, or a variant influenced by another modifier? PMID- 12124997 TI - Identification of five novel WASP mutations in Chinese families with Wiskott Aldrich syndrome. AB - The Wiskott-Aldrich Syndrome (WAS) is an X-linked recessive immunodeficiency caused by mutation in the gene encoding WAS protein (WASP). The disease is characterized by eczema, thrombocytopenia and severe immunodeificency and is associated with extensive clinical heterogeneity. Mutation studies indicated that the mutated genotypes are also highly variable. In this study, we performed PCR direct sequencing analysis of the WAS gene in six unrelated Chinese families. Five novel mutations identified, included two nonsense mutations (506C-->T, 1388- >T), a small insertion (685-686insCGCA) and two single-base deletions (384delT, 984delC). All of the mutations are predicted to lead to premature translational termination of WASP. PMID- 12124998 TI - Mutations in the human ATP-binding cassette transporters ABCG5 and ABCG8 in sitosterolemia. AB - Phytosterolemia or Sitosterolemia is a rare autosomal recessive disorder characterized by highly elevated plasma levels of plant sterols and cholesterol as a consequence of hyperabsorption and impaired biliary secretion of sterols. The disease is caused by mutations in two half size ATP-binding cassette transporters, ABCG5 and ABCG8. We have analyzed the genomic sequence of ABCG5 and ABCG8 in five well-characterized patients with Sitosterolemia. In the first patient we found a heterozygous mutation in exon 8 of the ABCG5 gene leading to a premature termination of the protein (Arg408Ter). This German patient is the first European showing a mutation of the ABCG5 gene. In a second patient we found a novel heterozygous mutation in exon 5 of ABCG8 (c.584T>A; Leu195Gln). Both patients were heterozygous for the identified mutation, but no mutation could be identified on the other chromosome. In three further analyzed patients we found mutations in exons 7, 9 and 11 of the ABCG8 gene, respectively, of which two result in a premature termination signal for translation products. One of these patients was compound heterozygous (Trp361Ter and Arg412Ter), the other was homozygous for Trp361Ter. The third patient was homozygous for an amino acid exchange (Gly574Arg). In conclusion this report describes one novel mutation affecting a highly conserved amino acid and two previously identified mutations in the ABCG8 gene. In addition, we identified for the first time a mutation in the ABCG5 gene of a European Sitosterolemia patient. PMID- 12124999 TI - 'Top down' protein characterization via tandem mass spectrometry. AB - Technological and scientific advances over the past decade have enabled protein identification and characterization strategies to be developed that are based on subjecting intact protein ions and large protein fragments directly to tandem mass spectrometry. These approaches are referred to collectively as 'top down' to contrast them with 'bottom up' approaches whereby protein identification is based on mass spectrometric analysis of peptides derived from proteolytic digestion, usually with trypsin. A key step in enabling top down approaches has been the ability to assign tandem mass spectrometer product ion identities, which can be done either via high resolving power or through product ion charge state manipulation. The ability to determine product ion charge states has permitted studies of the reactions, including dissociation, ion-molecule reactions, ion electron reactions and ion-ion reactions of high-mass, multiply charged protein ions. Electrospray ionization combined with high magnetic field strength Fourier transform ion cyclotron resonance has proven to be particularly powerful for detailed protein characterization owing to its high mass resolution and mass accuracy and its ability to effect electron capture-induced dissociation. Other types of tandem mass spectrometers are also beginning to find increasing use in top down protein identification/characterization studies. Charge state manipulation via ion-ion reactions in electrodynamic ion traps, for example, enables top down strategies to be considered using instruments with relatively modest mass resolution capabilities. Precursor ion charge state manipulation techniques have also recently been demonstrated to be capable of concentrating and charge-state purifying proteins in the gas phase. Advances in technologies applied to the structural analysis of whole protein ions and in understanding their reactions, such as those described here, are providing new options for the study of complex protein mixtures. PMID- 12125000 TI - Clustering of nucleobases with alkali metals studied by electrospray ionization tandem mass spectrometry: implications for mechanisms of multistrand DNA stabilization. AB - Self-clustering of the five common nucleobases was investigated by electrospray ionization tandem mass spectrometry and shown to provide insight into the non covalent interactions between identical bases. Alkali and ammonium cations significantly increase self-aggregation of the nucleobases and lead to the formation of uniquely stable magic number clusters. Sodium adducts of guanine, thymine and uracil preferentially take the form of tetrameric (quartet) clusters. This gas-phase result correlates with previously reported solution-phase data on sodium cation stabilized guanosine, thymine and uracil quartet structures believed to be responsible for telomere stabilization. In the presence of potassium, cesium or ammonium cations, pentameric magic number clusters are formed from thymine and uracil, while in solution the nucleoside isoguanosine yields clusters of this favored size. The formation of magic number metaclusters occurs for thymine and uracil in the presence of ammonium cations. These doubly charged 10- and 15-mers are tentatively attributed to the formation of pentamer/ammonium cation/ pentamer sandwich structures. PMID- 12125002 TI - Liquid chromatography/mass spectrometry in anabolic steroid analysis- optimization and comparison of three ionization techniques: electrospray ionization, atmospheric pressure chemical ionization and atmospheric pressure photoionization. AB - The applicability of liquid chromatography/tandem mass spectrometry (LC/MS/MS) for the detection of the free anabolic steroid fraction in human urine was examined. Electrospray ionization (ESI), atmospheric pressure chemical ionization and atmospheric pressure photoionization methods were optimized regarding eluent composition, ion source parameters and fragmentation. The methods were compared with respect to specificity and detection limit. Although all methods proved suitable, LC/ESI-MS/MS with a methanol-water gradient including 5 mM ammonium acetate and 0.01% acetic acid was found best for the purpose. Multiple reaction monitoring allowed the determination of steroids in urine at low nanogram per milliliter levels. LC/MS/MS exhibited high sensitivity and specificity for the detection of free steroids and may be a suitable technique for screening for the abuse of anabolic steroids in sports. PMID- 12125001 TI - Validated assay for quantification of oxcarbazepine and its active dihydro metabolite 10-hydroxycarbazepine in plasma by atmospheric pressure chemical ionization liquid chromatography/mass spectrometry. AB - Oxcarbazepine (OX), a new antiepileptic, may lead to unwanted side-effects or even life-threatening intoxications after overdose. Therefore, a validated liquid chromatographic/mass spectrometric (LC/MS) assay was developed for the quantification of OX and its pharmacologically active dihydro metabolite (dihydrooxcarbazepine, DOX, often named 10-hydroxycarbazepine). OX and DOX were extracted from plasma by the authors' standard liquid/liquid extraction and were separated on a Merck LiChroCART column with Superspher 60 RP Select B as the stationary phase. Gradient elution was performed using aqueous ammonium formate and acetonitrile. The compounds were quantified in the selected-ion monitoring mode using atmospheric pressure chemical ionization electrospray LC/MS. The assay was fully validated. It was found to be selective. The calibration curves were linear from 0.1 to 50 mg l(-1) for OX and DOX. Limits of quantification were 0.1 mg l(-1) for OX and DOX. The absolute recoveries were between 60 and 86%. The accuracy and precision data were within the required limits. The analytes in frozen plasma samples were stable for at least 1 month. The method was successfully applied to several authentic plasma samples from patients treated or intoxicated with OX. The measured therapeutic plasma levels ranged from 1 to 2 mg l(-1) for OX and from 10 to 40 mg l(-1) for DOX. The validated LC/MS assay proved to be appropriate for quantification of OX and DOX in plasma for clinical toxicology and therapeutic drug monitoring purposes. The assay is part of a general analysis procedure for the isolation, separation and quantification of various drugs and for their full-scan screening and identification. PMID- 12125003 TI - Quantification of grafted poly(ethylene glycol)-silanes on silicon by time-of flight secondary ion mass spectrometry. AB - Silicon grafted monodisperse poly(ethylene glycol) (PEG) silanes with various PEG chain lengths and mixtures of these were systematically analyzed with static time of-flight secondary ion mass spectrometry (TOF-SIMS). The mass spectra show differences in the various relative signal intensities, an observation that was used to elucidate important aspects of the grafting process. The relationship between PEG-silane fragment ion abundances and Si(+) ion abundances were used to (i) qualitatively describe layer thicknesses of grafted mixtures of PEG-silanes on silicon, (ii) construct a calibration curve from which PEG chain length (or molecular mass) can be determined and (iii) quantitatively determine surface mixture compositions of grafted monodisperse PEG-silanes of different chain lengths (3, 7 and 11 PEG units). The results suggest that discrimination does take place in the adsorption process. The PEG-silane with the shorter PEG chain is discriminated for mixtures containing PEG3-silane, whereas the PEG-silane with the longer PEG chain is discriminated in PEG7/PEG11-silane mixtures. The origin of this difference in adsorption behavior is not well understood. Aspects of the grafting process and the TOF-SIMS analyses are discussed. PMID- 12125004 TI - Mass spectrometric characterization of substituted 2-thiazolin-4-one derivatives. AB - Electron ionization mass spectrometry was used for the structural characterization of substituted 2-thiazolin-4-one derivatives in the gas phase. The compounds follow common fragmentation pathways, producing ions whose abundances are dependent on the chemical nature of the substituent at position 2. Collision-induced dissociation tandem mass spectrometric experiments, carried out on both molecular ions and fragment ions produced in the source, allowed the elucidation of gas-phase decompositions. The presence of tautomeric forms is suggested for some ionic species. Rapid identification of a primary or secondary amine moiety at position 2 of the thiazoline ring can be achieved by the detection of characteristic fragmentations occurring both in the ion source and under the collision-induced dissociation regime. PMID- 12125005 TI - Detection of water-soluble vitamins by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry using porphyrin matrices. AB - The detection of water-soluble vitamins B(1), B(2), B(6), B(12) and C by matrix assisted laser desorption/ionization (MALDI) time-of-flight mass spectrometry (TOFMS) was attempted by studying 17 porphyrin matrices. Comparative studies of porphyrin matrices, useful mass spectral window, matrix/analyte molar ratio (M/A), ultraviolet-visible absorption characteristics and quantitative results were made. Most porphyrin matrices provide a useful mass spectral window in the low-mass range. The optimal M/A increases with increasing molecular mass of the vitamin. Vitamin B(12) possesses the highest molecular mass and requires a higher M/A. The presence of hydroxyl or carboxyl groups in the porphyrin is an indicator of a useful MALDI matrix. Vitamins B(2) and B(6) were readily ionized upon irradiation with a 337 nm laser without the use of any porphyrin matrix. Improved linearity and sensitivity of the calibration curves were obtained with samples prepared with a constant M/A. The limits of detection and quantitation are at the picomole level. The results indicate that MALDI-TOFMS with porphyrin matrices is a rapid and viable technique for the detection of low molecular mass water soluble vitamins. PMID- 12125006 TI - Mass spectrometry of cis-diamminedichloroplatinum(II) adducts with the dinucleosidemonophosphates d(ApG), d(GpG) and d(TpC) in an ion trap. AB - The detection and fragmentation behaviour of adducts of the chemotherapeutic cis diamminedichloroplatinum(II) (cisplatin) with the dinucleosidemonophosphates d(ApG), d(GpG) and d(TpC) as model compounds for DNA adducts in an ion trap with electrospray ionization were studied. Mainly the monofunctional adduct, the bifunctional adduct and the bifunctional adduct with platinum bridging two dinucleosidemonophosphates were detected. In addition, several more complex adducts were seen resulting from reactions among these species. Adduct formation was low in the case of d(TpC). Fragmentation could be controlled strongly by varying the temperature of the transfer capillary; furthermore, tandem mass spectrometric (MS/MS) experiments on both the monofunctional and the bifunctional adducts were performed. For the adducts of d(ApG) and d(GpG) losses of NH(3) and HCl were the most dominant reactions, followed by the losses of one, then another two units of 98 amu from the sugar-phosphate backbone, whereas d(TpC)-Pt predominantly forms the dinucleosidemonophosphate. In the gas phase, the conversion of the monofunctional into the bifunctional adducts through binding to another site in the dinucleotide accompanied by loss of NH(3) or HCl could also be observed. The removal of a ligand from the coordination sphere of the square planar platinum complexes appeared to be the crucial step for the induction of further fragmentation of the dinucleotide ligand. MS(n) experiments of the bifunctional adducts of d(ApG) and d(GpG) revealed different fragmentation pathways involving the loss of phosphoric acid, metaphosphoric acid, deoxyribose units (intact or dehydrated) and the nucleobases in different orders, leaving characteristic binding site-determining fragments. Fragmentation of these ions was also performed, mainly resulting in fragmentation of the bases. The study confirmed the remarkable stability of the platinum-guanine bond compared with other nucleobases. PMID- 12125007 TI - Decomposition of multicomponent mass spectra using Bayesian probability theory. AB - We present a method for the decomposition of the mass spectra of mixed gases using Bayesian probability theory. The method works without any calibration measurement and therefore applies also to the analysis of spectra containing unstable species. For the example of mixtures of three different hydrocarbon gases the algorithm provides concentrations and cracking coefficients of each mixture component and also their confidence intervals. The amount of information needed to obtain reliable results and its relation to the accuracy of our analysis are discussed. PMID- 12125008 TI - Qualitative characterization of biomolecular zinc complexes by collisionally induced dissociation. AB - Nanospray and collisionally induced dissociation (CID) on a quadrupole/time-of flight mass spectrometer were used to examine the complexes formed between the zinc ion binding protein metallothionein and a series of peptides related to glutathione. The objective of the study was to determine if CID could be used to distinguish complexes that are stabilized by co-chelation of a zinc ion from non covalent complexes that were formed in some other way. Differences in the collision energy required for dissociation and, more importantly, differences in the distribution of zinc ions between the pairs of dissociation products suggest that mass spectrometry can provide qualitative information about the bimolecular chelation of metal ions. The potential application to zinc chelates is particularly important, since biological chelates do not provide signals directly detectable by NMR, Mossbauer or other spectroscopies. The observations reported here also allowed a molecular mechanism to be proposed to explain the differences observed by others in the physiological interactions of reduced and oxidized glutathione with metallothionein. PMID- 12125009 TI - Electrospray mass spectral studies on molybdenum complexes with 2,3- and 2,5 dihydroxybenzoic acid and their degradation products. PMID- 12125010 TI - Current literature in mass spectrometry. PMID- 12125011 TI - Interactions between viral and human genes. PMID- 12125012 TI - Studies in smallpox and vaccination. 1913. PMID- 12125013 TI - CMV and blood transfusions. AB - Among the human herpesviruses, cytomegalovirus (CMV) is the only one that has assumed significant importance in blood transfusion. Transfusion transmission of CMV (TT-CMV) to seronegative immunocompromised patients can lead to lethal CMV disease. Studies over the past 30 years have demonstrated that monocytes latently infected with CMV represent the primary vector for TT-CMV, and that TT-CMV can be largely abrogated by transfusing at-risk patients with either seronegative units or blood filtered to remove white blood cells. However, the small number of cases of breakthrough TT-CMV that follow transfusion of either seronegative or filtered blood still produce morbidity and mortality. These circumstances have motivated ongoing efforts to provide improved protection from TT-CMV, including the use of CMV DNA amplification for blood screening, and pathogen inactivation to sterilise all blood components prior to transfusion. PMID- 12125014 TI - The application of molecular phylogenetics to the analysis of viral genome diversity and evolution. AB - DNA sequencing and molecular phylogenetics are increasingly being used in virology laboratories to study the transmission of viruses. By reconstructing the evolutionary history of viral genomes the behaviour of viral populations can be modelled, and the future of epidemics may be forecast. The manner in which such viral DNA sequences are analysed is the focus of this review. Many researchers resort to the often-quoted 'black box' approach because phylogenetics theory can be daunting, and phylogenetics software packages can appear to be difficult to use. However, because phylogenetic analyses are often used in important and sensitive arenas, for example to provide evidence indicating transmission between persons, it is vital that appropriate care is taken to estimate reliably true relationships. In this review, we discuss how a molecular phylogenetics study should be approached, give an overview of the methods and programs for analysing DNA sequence data, and point readers to appropriate texts for further details. The aim of this review, therefore, is to provide researchers with an easy to understand guide to molecular phylogenetics, with special reference to viral genomes. PMID- 12125015 TI - Protein-protein interactions as targets for antiviral chemotherapy. AB - Most cellular and viral processes depend on the coordinated formation of protein protein interactions. With a better understanding of the molecular biology and biochemistry of human viruses it has become possible to screen for and detect inhibitors with activity against specific viral functions and to develop new approaches for the treatment of viral infections. A novel strategy to inhibit viral replication is based on the disruption of viral protein-protein complexes by peptides that mimic either face of the interaction between subunits. Peptides and peptide mimetics capable of dissociating protein-protein interactions have such exquisite specificity that they hold great promise as the next generation of therapeutic agents. This review is focused on recent developments using peptides and small molecules to inhibit protein-protein interactions between cellular and/or viral proteins with comments on the practicalities of transforming chemical leads into derivatives with the characteristics desired of medicinal compounds. PMID- 12125016 TI - Identification of omega hydroxy fatty acids in biological samples as their pentafluoropropyl derivatives by gas chromatography/mass spectrometry with positive and negative ion detection. AB - A simple one-step procedure for derivatization of the omega hydroxy fatty acids 20-hydroxyeicotetraeonic acid and 12-hydroxylauric acid is presented. The procedure involves acylation of the terminal hydroxy group and esterification of the carboxylic acid with a mixture of pentafluoropropionic anhydride and pentafluoropropanol. Positive and negative ion spectra for the derivatives are presented. The procedure was used to demonstrate conversion of arachidonic acid to 20-hydroxyeicosatetraeonic acid and lauric acid to 12-hydroxylauric acid in kidney microsomal incubations. The reaction appears to be specific, since derivatives of subterminal fatty acids (secondary alcohols) could not be detected. PMID- 12125017 TI - A method to identify and simultaneously determine the relative quantities of proteins isolated by gel electrophoresis. AB - Gel electrophoresis is often used for the primary analysis and purification of proteins, and peptide mapping by matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) is a widely used technique for the rapid identification of unknown proteins. The identification is usually obtained by digesting the protein with an enzyme and matching the masses of the proteolytic peptides with those of each protein in a sequence database. Another important aspect in many proteomic experiments is the determination of the relative protein quantities (e.g. comparison between control and altered states). Usually, this is obtained by comparing the spot intensities of two independent gels. This procedure is time consuming and not very accurate. Recently, several methodologies using isotope labeling of proteins for quantitative proteomic studies have been introduced (e.g. using ICAT reagents or growing cells in isotopically enriched nutrients). However, none of these methodologies is foolproof and there is still the need for simple and inexpensive alternatives for determining the relative quantities of proteins. Previously, we showed that a mixture of acrylamide and deuterated acrylamide could be used as cysteine alkylating reagent prior to electrophoresis, improving the coverage and the confidence of the protein identification procedure (Sechi S, Chait BT. Anal. Chem. 1998; 70: 5150). Here we show that a similar approach can be used to obtain relative quantitation at the femtomole level of proteins isolated by gel electrophoresis. Deuterated acrylamide is used to alkylate the cysteines in one sample and regular acrylamide is used to alkylate the cysteines in the second sample. The two samples are then mixed together in a 1:1 ratio and the relative protein quantities are determined from the ion intensity ratios of the two cysteine-containing peptides isotopic envelopes (regular/deuterated). The analysis of several proteins mixed in different ratios is reported showing that this approach can reliably be used for protein identification and quantification. Briefly, a simple and inexpensive method for quantifying and simultaneously identifying proteins isolated by gel electrophoresis using MALDI-MS is presented. PMID- 12125018 TI - Influence of the 6th and 10th spatial harmonics on the peak shape of a quadrupole mass filter with round rods. AB - The influence of the ratio of the rod radius, r, to field radius, r(0), on the peak shape for a linear quadrupole mass filter constructed with round rods has been investigated. The expansion of the potential in multipoles, phi(N),Phi(x, y) = sum(infinity)(N=0)A(N)phi(N)/r(N)(0) has been considered, and the peak shape and resolution have been determined by numerical calculation of ion trajectories in quadrupoles with different ratios, r/r(0). Geometries that make the dodecapole term (A(6)) zero (r/r(0) = 1.14511) do not give the best performance because the contribution of the 20-pole term, A(10), must be considered. The optimum ratio is r/r(0) approximately 1.13. With this ratio the dodecapole term (A(6)) is ca. 1 x 10(-3), but its effects are partially compensated by the A(10) term which has similar magnitude, but opposite sign. PMID- 12125020 TI - High-throughput liquid chromatography ultraviolet/mass spectrometric analysis of combinatorial libraries using an eight-channel multiplexed electrospray time-of flight mass spectrometer. AB - We report a high-throughput liquid chromatography/mass spectrometry (LC/MS) protocol for analyzing large combinatorial libraries using an eight-channel parallel LC/UV/MS (MUX-LCT) system. System configuration, linear response range in UV absorbance, LC column selection, and flow rate were optimized for 24 h/7 day unattended operations. Combinatorial libraries were analyzed on this system at a rate of 3200 compounds per day for a 3.5 min cycle time per injection. This parallel system is compared with a single-channel system in terms of performance and operation. PMID- 12125019 TI - Determination of total urinary mercury by on-line sample microwave digestion followed by flow injection cold vapour inductively coupled plasma mass spectrometry or atomic absorption spectrometry. AB - The total mercury content in urine was determined by inductively coupled plasma mass spectrometry with the so-called cold vapour method after on-line oxidative treatment of the sample in a microwave oven (FI-MW-CV-ICPMS). Use of a KBr/KBrO(3) mixture, microwave digestion, and the final oxidation with KMnO(4), assure the complete recovery of the organic forms of Hg which would be difficult to determine otherwise if using only the CV-ICPMS apparatus. Quantitative recoveries were obtained for phenyl Hg chloride (PMC), dimethyl Hg (DMM), Hg acetate (MA) and methyl Hg chloride (MMC). Use of automatic flow injection microwave systems (FI-MW) for sample treatment reduces environmental contamination and allows detection limits suitable for the determination of reference values. Since no certified reference materials were commercially available in the concentration ranges of interest, the accuracy of the proposed procedure has been assessed by analysing a series of urine samples with two independent techniques, ICP-MS and AAS. When using the FI-MW-CV-ICP-MS technique, the detection limit was assessed at 0.03microg/L Hg, while with FI-MW-CV-AAS it was 0.2microg/L Hg. The precision of the method was less than 2-3% for FI-MW-CV ICP-MS and about 3-5% for FI-MV-CV-AAS at concentrations below 1microg/L Hg. These results show that ICP-MS can be considered as a "reference technique" for the determination of total urinary Hg at very low concentrations, such as are present in non-exposed subjects. PMID- 12125021 TI - Improved peptide detection with matrix-assisted laser desorption/ionization mass spectrometry by trimethylation of amino groups. AB - Trimethyl alpha-amino derivatives of peptides (penta to deca) with a permanent positive charge on their alpha-amino groups were prepared by in vacuo reaction with iodomethane and subjected to matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS). Compared to the unmodified peptide, the signal intensity of the trimethyl alpha-amino derivative in MALDI-MS is increased by at least an order of magnitude. Similarly, an octapeptide with a trimethylated epsilon-amino group derived from the solitary lysine residue of the B-chain of insulin also shows the same relative increase in signal intensity. Another advantage of the in vacuo methylation procedure is that trimethylation of a peptide amino group can be carried out readily with a combination of isotopes (13)CH(3)I and (12)CH(3)I or CD(3)I and CH(3)I, yielding a doublet signal either 3 or 9 units apart, respectively. The presence or absence of such a doublet signal can be used as a criterion to discriminate between peptide and non-peptide signals in the mass spectrum. PMID- 12125022 TI - Taylor dispersion monitored by electrospray mass spectrometry: a novel approach for studying diffusion in solution. AB - This work describes a novel approach for monitoring analyte diffusion in solution that is based on electrospray ionization mass spectrometry (ESI-MS). A mass spectrometer at the end of a laminar flow tube is used to measure the Taylor dispersion of an initially sharp boundary between two solutions of different analyte concentration. This boundary is dispersed by the laminar flow profile in the tube. However, this effect is diminished by analyte diffusion that continuously changes the radial position, and hence the flow velocity of individual analyte molecules. The steepness of the resulting dispersion profile therefore increases with increasing diffusion coefficient of the analyte. A theoretical framework is developed to adapt the equations governing the dispersion process to the case of mass spectrometric detection. This novel technique is applied to determine the diffusion coefficients of choline and cytochrome c. The measured diffusion coefficients, (11.9 +/- 1.0) x 10(-10) m(2) s(-1) and (1.35 +/- 0.08) x 10(-10) m(2) s(-1), respectively, are in agreement with the results of control experiments where the Taylor dispersion of these two analytes was monitored optically. Due to the inherent selectivity and sensitivity of ESI-MS, it appears that the approach described in this work could become a valuable alternative to existing methods for studying diffusion processes, especially for experiments on multicomponent systems. PMID- 12125023 TI - Quantitative analysis of diclazuril in animal plasma by liquid chromatography/electrospray ionization mass spectrometry. AB - A novel, sensitive and specific method for the quantitative determination of diclazuril in animal plasma using liquid chromatography combined with electrospray ionization tandem mass spectrometry (LC/ESI-MS/MS) with negative ion detection is presented. Extraction of the samples was performed with a rapid deproteinization step using acetonitrile. Chromatography of diclazuril and the internal standard was achieved on a Nucleosil ODS 5-microm column, using a gradient elution with 0.01 M ammonium acetate and acetonitrile. To obtain the highest sensitivity and specificity possible, the mass spectrometer was operated in the multiple reaction monitoring (MRM) mode. The method was validated according to the requirements defined by the European Community. Calibration curves using plasma fortified between 1-100 ng/mL and 100-2000 ng/mL showed good linear correlation (r >or= 0.9991, goodness-of-fit coefficient 200 A that represent hundreds of aggregated chains. Samples of mass fraction 1% DPS (6050 g mol(-1)) in 2,5 dihydroxybenzoic acid, all-trans-retinoic acid, and sinapinic acid also have large zero angle scattering characteristic of large aggregates. The morphological trend obtained from the SANS measurements of the DPS aggregate size in the four matrixes is dithranol > 2,5-dihydroxybenzoic acid > all-trans-retinoic acid > sinapinic acid. These measurements indicate that DPS in dithranol exhibits the most strong phase separation, while DPS in sinapinic acid shows considerable domain mixing. All of these matrixes produce MALDI signal strength under appropriate conditions, suggesting that strong phase separation does not diminish the signal-to-noise ratio. DPS (188,000 g mol(-1)) in biphenyl was used as a model system of a matrix that can be either crystalline or amorphous. SANS data shows that above the biphenyl melting point, a conventional solution is formed that has molecularly dispersed polymers. Upon crystallization, there is strong aggregation of the DPS into large domains. Therefore, the crystalline matrixes commonly used in MALDI measurements probably cause large aggregations of polymers to be present during the MALDI process. PMID- 12125028 TI - Generic fast gradient liquid chromatography/tandem mass spectrometry techniques for the assessment of the in vitro permeability across the blood-brain barrier in drug discovery. AB - Rapid, generic gradient liquid chromatography/tandem mass spectrometry (LC/MS/MS) assays, designed to accelerate sample analyses, have been developed to keep pace with the productivity of advanced synthetic procedures. In this study, LC/MS/MS was combined with an in vitro, cell-based, blood-brain barrier (BBB) model to evaluate the potential of new chemical entities (NCEs) to cross the BBB. This in vitro assay provides the permeability of discovery compounds across a monolayer of a primary culture of bovine brain microvessel endothelial cells in a fraction of the time that is required for in vivo studies (brain/plasma concentrations), using only 2 mg of the compound. The results are consistent with in vivo brain/plasma concentration ratio data. PMID- 12125029 TI - Assignment of absolute configuration of chiral carboxylic acids via exciton coupled CD treatment: 4-phenylthioproline as a case study. AB - The 4-hydroxybenzoate chromophore was used as an exciton reporter group for the assignment of the absolute configuration of a chiral carboxylic acid of pharmaceutical interest, (-)-(2S,4S)-4-phenylthioproline, precursor of the ACE inhibitor zofenopril. The nondegenerate coupling with the preexisting phenylthio chromophore was observed. A detailed conformational analysis, accomplished by means of (1)H-NMR NOESY and semiempirical PM3 computational methods, and quantitative CD calculations were necessary to substantiate the assignment based on a weak experimental couplet. PMID- 12125030 TI - Absolute configuration and conformational analysis of a degradation product of inhalation anesthetic Sevoflurane: A vibrational circular dichroism study. AB - 1,1,1,3,3-pentafluoro-2-(fluoromethoxy)-3-methoxypropane, compound B, is a product obtained in the degradation of the anesthetic Sevoflurane. Enantiopure (+)-B was investigated using vibrational circular dichroism (VCD). Experimental absorption and VCD spectra of (+)-B in CDCl(3) solution in the 2,000-900 cm(-1) region are compared with the ab initio predictions of absorption and VCD spectra obtained from density functional theory using B3LYP/6-31G* basis set for different conformers of (S)-1,1,1,3,3-pentafluoro-2-(fluoromethoxy)-3 methoxypropane. This comparison indicates that (+)-B is of the (S)-configuration in CDCl(3) solution, in agreement with previous literature results. Our results also indicate that this compound adopts six predominant conformations in CDCl(3) solution. PMID- 12125031 TI - Chemoenzymatic synthesis and properties of Schiff bases containing (R)-1-(9 anthryl)ethylamine. AB - Racemic 1-(9-anthryl)ethylamine (10), obtained in 70% overall yield from commercial 9-cyanoanthracene, was kinetically resolved by the Candida antarctica A lipase-catalyzed acetylation with isopropyl acetate as acyl donor, affording (R)-(+)-10 with 95.8% enantiomeric excess (e.e.) (E-value 43.5), which afforded Schiff bases (R)-4 and(R)-8. (1)H-NMR, CD, and MM2 calculations offer a consistent picture of the conformational properties of these potential ligands and an explanation for the limited enhancement of enantioselectivity in cyclopropanation of styrene by their Cu(I) complexes, as compared with previously studied ligands in this series. PMID- 12125032 TI - Enantioselective chromatography of alkyl derivatives of 5-ethyl-5-phenyl-2 thiobarbituric acid studied by semiempirical AM1 method. AB - Complexation of alkyl derivatives of 5-ethyl-5-phenyl-2-thiobarbituric acid (2 thiophenobarbital) enantiomers by beta-cyclodextrin was investigated by the AM1 method. The inclusion complexes of beta-cyclodextrin with neutral and anionic forms of these enantiomers have been modeled and energetically optimized. The chiral discrimination of enantiomers was analyzed in terms of differences in the interaction energies. The calculated interaction energies between each enantiomer of the investigated 2-thiobarbiturates and beta-cyclodextrin confirm the ability of beta-cyclodextrin to act as a mobile phase additive in reversed-phase HPLC to separate enantiomers by liquid chromatography and rationalize their order of elution. PMID- 12125033 TI - Enantioselective plasma protein binding of bimoclomol. AB - The binding of bimoclomol enantiomers to human plasma, its components, as well as to plasma from monkey, dog, rat, and mouse was investigated by ultrafiltration and equilibrium dialysis. The considerably stronger binding of the (-)-(S) enantiomer found in human plasma is due to the alpha(1)-acid glycoprotein (AAG) component. The binding parameters for AAG (n(R)K(R) = 1.3 x 10(4) M(-1) and n(S)K(S) = 1.0 x 10(5) M(-1)) revealed high enantioselectivity, while the binding to human serum albumin was found to be weak (nK = 5 x 10(3) M(-1)) and not stereoselective. (-)-(S)-Bimoclomol was extensively displaced in the presence of specific marker ligands for the "FIS" subfraction of human AAG. Comparative binding studies indicated considerable differences between plasma of the five species investigated. PMID- 12125034 TI - Stereoselective metabolism of pentoxifylline in vitro and in vivo in humans. AB - Pentoxifylline increases erythrocyte flexibility, reduces blood viscosity, and inhibits platelet aggregation and is thus used in the treatment of peripheral vascular disease. It is transformed into at least seven phase I metabolites, of which two, M1 and M5, are active. The reduction of the keto group of pentoxifylline to a secondary alcohol in M1 takes place chiefly in erythrocytes, is rapidly reversible, and creates a chiral center. The aims of this study were: to develop HPLC methods to separate the enantiomers of M1, to investigate the kinetics of the reversible biotransformation of pentoxifylline to (R)- and (S)-M1 in hemolysed erythrocyte suspension, and to quantify the formation of the enantiomers of M1 (as well as M4 and M5) after intravenous and oral administration of pentoxifylline to human volunteers. (R)- and (S)-M1 could be separated preparatively on a cellobiohydrolase column, while determination in blood or plasma was by HPLC after chiral derivatization with diacetyl-L-tartaric acid anhydride. The metabolism of pentoxifylline to (R)-M1 in suspensions of hemolysed erythrocytes followed simple Michaelis-Menten kinetics (K(m) = 11 mM), while that to (S)-M1 was best described by a two-enzyme model (K(m) = 1.1 and 132 mM). Studies with inhibitors indicated that the enzymes were of the carbonyl reductase type. At a therapeutic blood concentration of pentoxifylline, the calculated rate of formation of (S)-M1 is 15 times higher than that of the (R) enantiomer. Back-conversion of M1 to pentoxifylline was 3-4 times faster for the (S)- than for the (R)-enantiomer. In vivo, the R:S plasma concentration ratio of M1 ranged from 0.010-0.025 after intravenous infusion of 300 or 600 mg of pentoxifylline, and from 0.019-0.037 after oral administration of 600 mg. The biotransformation of pentoxifylline to M1 was thus highly stereoselective in favor of the (S)-enantiomer both in vitro and in vivo. PMID- 12125035 TI - Use of an adsorbed chiral selector in capillary LC-MS. AB - Lasalocid was utilized as a chiral selector adsorbed on porous graphitic carbon. Important parameters were identified for the use of the chiral selector in capillary liquid chromatography combined with MS detection. The influence on both retention and enantioselectivity as well as mass spectrometric performance was studied at lasalocid concentrations of up to 12 mg/l (20 microM). Moreover, the stability of retention factors and enantioselectivities was also evaluated at low selector concentration of 1 mg/l (2 microM). The results show that in order to optimize the MS performance the selector concentration should be kept at low microM. The chromatographic performance at reduced selector concentration (1 mg/l = 2 microM) was studied over a 10-day period, showing satisfactory enantioselectivity and stable performance concerning enantioselectivity and retention. PMID- 12125036 TI - Development of a nonchiral HPLC method with circular dichroism detection for chiral analysis of molybdenum-catalyzed enantioselective synthesis products. AB - The combination of a circular dichroism detector and nonchiral liquid chromatography was used for the chiral analysis of unresolved enantiomers to determine the enantioselectivity of a molybdenum-catalyzed asymmetric allylic alkylation reaction. The CD/UV peak area ratio of the unresolved enantiomers was calculated and compared with that of a reference standard to determine the enantiomeric purity. The limit of quantitation (LOQ) is 20 ng for the chiral ligand and 1 microg for the branched chiral product. The viability of the system depends on the limit of quantitation of the CD response and the linearity range of the CD and UV response. PMID- 12125037 TI - Contribution of chiral HPLC in tandem with polarimetric detection in the determination of absolute configuration by chemical interconversion method: Example in 1-(thi)oxothiazolinyl-3-(thi)oxothiazolinyl toluene atropisomer series. AB - We report the determination of the absolute configuration of eight stereoisomers in the series of chiral 1-(thi)oxothiazolinyl-3-(thi)oxothiazolinyl toluene atropisomers 1-3, from the known absolute configuration of one stereoisomer, determined by X-ray crystallography. The method uses the affiliation between signs of rotation of polarised light during chemical transformations which preserve the absolute configuration and also during rotation around a single pivot bond producing a compound of known configuration. The use of chiral HPLC in tandem with a chirality detector gives a decisive advantage since such correlation can be performed on a mixture of a very limited quantity of compounds, without tedious purification steps. The method shown as an example in this article, which uses chiral HPLC with chirality detection, may prove useful in many other cases where the determination of the absolute configuration is necessary and where a chemical interconversion method can be used on a microscale. PMID- 12125038 TI - Nonlinearity in optical resolution via distillation applying mixtures of resolving agents. AB - During an optical resolution it is the resolving agent that has the strongest influence on the outcome of the process. Applying a mixture of resolving agents can result either in antagonism or in synergy. We found that using mixtures of tartaric acid and its derivatives chiral selectivity is at least the same, but in several cases markedly better (synergistic effect), than the sum of the effect of the individual resolving agents. Thus, the "Dutch method," reported for the crystallization method, also works for distillation. A calculation method is applied for measuring the synergistic effect. Interestingly, an individually inactive resolving agent can be a useful contributor to the mixture of the resolving agents. PMID- 12125039 TI - Ligand distortion modes leading to increased chirality content of Katsuki Jacobsen catalysts. AB - The "chirality content" of Katsuki-Jacobsen epoxidation catalysts are computed with the Avnir continuous chirality measure (CCM). An assessment of Mn(salen) molecules from the Cambridge Structural Database shows there exist some variation in CCM and the chirality content for several triplet state complexes of these catalysts purported in the literature to be the active species show even larger CCM values. Several deformation modes were analyzed to examine how chirality content changes as catalyst distortion is induced. The deformations studied include in-plane deformations, cup-shaped puckering, ligand twisting motions, and step-like deformations. Some distortions lead to increases of chirality while others lead to a decrease in chirality content. The most influential distortion modes that can be used for ligand design are twisting and step induction. PMID- 12125040 TI - Enantioselectivity in the steady-state pharmacokinetics and transplacental distribution of pindolol at delivery in pregnancy-induced hypertension. AB - Nine patients taking oral doses of 10 mg/12 h rac-pindolol as part of their treatment for hypertension in pregnancy were recruited for the study. Maternal and fetal gestational age ranged from 20-38 years and 28-41 weeks, respectively. Blood was collected from the umbilical cord vein and from the mother from zero to 12 h after drug administration. Urine was collected for 12 h after rac-pindolol administration at the following intervals: 0-3, 3-6, 6-9, and 9-12 h. Plasma and urine concentrations of the pindolol enantiomers were determined by HPLC using a Chiralpak AD chiral column and fluorescence detection. The data were fitted to a one-compartment model and differences between (+)-R and (-)-S enantiomers were compared by the paired t-test (P < 0.05). Mean results are reported. The disposition of pindolol in maternal plasma was stereoselective, with higher AUC(SS)0-12 (84.34 vs. 95.69 ng.h/ml) and Cl(R) values (9.16 vs. 10.85 L/h) and lower Vd/f (251.38 vs. 225.17 L) and Cl/f (62.48 vs. 55.74 L/h) for the (+)-R pindolol. The transplacental distribution of pindolol was not stereoselective. Cord, plasma, and presumably fetal, concentrations of the pindolol enantiomers were 56% of the maternal plasma concentrations up to 6 h after the last dose. PMID- 12125041 TI - Comparative effects of methamphetamine and nicotine on the striatal [(11)C]raclopride binding in unanesthetized monkeys. AB - Although a very large literature exists on the in vitro, ex vivo, and in vivo effects of nicotine on dopamine release in rodents, similar data in primates are scant. This study was initiated to compare methamphetamine to nicotine given i.v. to normal unanesthetized monkeys using positron emission tomography (PET) techniques. Release of dopamine in the striatum using [(11)C]raclopride was determined indirectly in four nicotine-naive adult Macaca mulatta monkeys under conscious and isoflurane-anesthetized conditions using high-resolution PET. [(11)C]Raclopride was given i.v. as a bolus injection followed by continuous infusion with steady state over 30-45 min. Nicotine bitartrate was then given as a bolus plus infusion for 30 min in doses of 32 microg/kg + 0.8 microg/kg/min or 100 microg/kg + 2.53 microg/kg/min as base. The larger doses of nicotine caused significant cardiovascular effects; these doses did not displace [(11)C]raclopride binding in either dorsal or ventral striatum under the anesthetized conscious condition. In contrast, isoflurane-anesthesia induced a slight but significant dose-dependent reduction of [(11)C]raclopride binding by nicotine even at the same doses used in the anesthetized condition. Methamphetamine in bolus doses of 0.1, 0.3, and 1.0 mg/kg i.v. under conscious condition caused a significant displacement of [(11)C]raclopride and isoflurane anesthesia facilitated the displacement induced by nicotine. These results indicate that nicotine, in high tobacco-smoking-related doses, does not release sufficient brain dopamine to displace [(11)C]raclopride in the striatum in the awake and fully conscious state, in contrast to small doses of methamphetamine. PMID- 12125042 TI - LTP in the mouse nucleus accumbens is developmentally regulated. AB - Glutamatergic transmission in the nucleus accumbens (NAc) has been shown to be important for behavioral adaptations in response to drugs of abuse. NMDA-receptor dependent long-term potentiation (LTP) of glutamatergic synaptic transmission has been hypothesized to underlie many lasting alterations in behavior. Thus, we examined LTP in NAc core and find that it is developmentally regulated. Specifically, tetanus-evoked, NMDA receptor-dependent LTP is observed in the NAc of "adolescent" (3-week-old) mice more frequently than in adult (6-20-week-old) mice. In contrast, cAMP-dependent enhancement of transmission is not developmentally regulated. Removal of extracellular Mg(2+) restores LTP in adult NAc core, suggesting developmental regulation of induction processes rather than maintenance mechanisms. These findings are discussed in the context of behavioral changes elicited in response to drugs of abuse, which differ in adolescent vs. adult rodents and humans. PMID- 12125043 TI - Effects of acute "binge" cocaine on preprodynorphin, preproenkephalin, proopiomelanocortin, and corticotropin-releasing hormone receptor mRNA levels in the striatum and hypothalamic-pituitary-adrenal axis of mu-opioid receptor knockout mice. AB - Cocaine administration increases activity at dopamine receptors, increases preprodynorphin (ppDyn) gene expression in the caudate-putamen (CPu), and activates the stress responsive hypothalamic-pituitary-adrenal (HPA) axis. To examine the hypothesis that mu-opioid receptors (MOR) may play roles in these cocaine effects, we tested the effects of acute "binge" pattern cocaine administration in mice with targeted disruption of the MOR gene. Wild-type (+/+) and homozygous MOR-deficient (-/-) mice received three injections of 15 mg/kg cocaine at 1-h intervals. Mice were sacrificed 30 min after the last injection and mRNAs for ppDyn and preproenkephalin (ppEnk) in the CPu and nucleus accumbens (NAc), and for type I corticotropin-releasing hormone receptor (CRH(1) receptor) and pro-opiomelanocortin (POMC) in the hypothalamus and pituitary, were measured by solution hybridization RNase protection assays. Cocaine elevated ppDyn mRNA in the CPu, but not NAc, of both the MOR -/- and wild-type mice. ppEnk mRNA in the CPu, but not NAc, was lower in MOR -/- mice than in wild-type mice following cocaine administration. Hypothalamic CRH(1) receptor and POMC mRNAs were expressed at similar levels in untreated and in cocaine-treated mice of each genotype. However, there were lower basal levels of CRH(1) receptor mRNA in the anterior pituitary of the MOR -/- mice than in wild-type mice and the MOR -/- mice failed to show the cocaine-induced decreases in CRH(1) receptor mRNA found in the wild-type mice. Cocaine activated the HPA axis similarly in MOR -/- and wild-type mice, as reflected in similar increases in plasma corticosterone levels in both genotypes. These results support a specific role for MORs in acute cocaine effects on striatal ppEnk gene expression and fail to support critical roles for these receptors in acute cocaine's effects on either ppDyn gene expression or HPA activation. MOR -/- mice are useful models for studying cocaine effects on ppEnk gene expression that could aid interpretation of the similar postmortem phenomena found in human cocaine addicts. PMID- 12125045 TI - Flip and flop splice variants of AMPA receptor subunits in the spinal cord of amyotrophic lateral sclerosis. AB - Excitotoxicity mediated by AMPA receptors has been suggested to be implicated in the pathogenesis of amyotrophic lateral sclerosis (ALS). To investigate the relevance of AMPA receptors to motor neuron degeneration in ALS, we evaluated the expression of mRNAs coding for flip and flop splice variants of AMPA receptor subunits (GluR-A to GluR-D) in the cervical segment of the spinal cord from control individuals and patients with ALS using in situ hybridization histochemistry. Transcript mRNAs coding for flop variants were significantly decreased in the ventral horn of the spinal cord from patients with ALS, whereas the mRNAs for flip variants were preserved. These findings suggest that the relative abundance of flip variants vs. flop variants is increased in spinal motor neurons of ALS patients when compared to that of control individuals. Flip variants promote assemblies of slowly desensitizing AMPA receptors. These results imply that spinal motor neurons of ALS patients possess more slowly desensitizing AMPA receptors than those of control individuals. This expression change of AMPA receptors in ALS may account for vulnerability of motor neurons in this disease. PMID- 12125044 TI - Spatial and temporal profile of haloperidol-induced immediate-early gene expression and phosphoCREB binding in the dorsal and ventral striatum of amphetamine-sensitized rats. AB - To determine if D(2) dopamine receptor-mediated nuclear signaling is altered during the development of amphetamine sensitization, we examined the expression of immediate-early gene (IEG) products, Fos, Jun, and Fos-related antigen (FRA), in both controls and amphetamine-sensitized rats after a challenge with the D(2) antagonist haloperidol. When chronic saline- or amphetamine (5 mg/kg, i.p. for 14 days)-treated rats were challenged with 2 mg/kg haloperidol at withdrawal day 3 (w3), more 35-kDa FRA was induced in the ventral striatum of the control group than in the amphetamine-treated rats. In contrast, more Jun and 35-kDa FRA were expressed in the ventral striatum of the amphetamine-treated group than in the controls when haloperidol was given at w10. Topographical analyses indicate that the decrease in FRA immunoreactive neuronal density in amphetamine-treated rats at w3 were located in the dorsolateral caudate/putamen and the nucleus accumbens shell and core subregions. Conversely, the increase in Jun-immunoreactive neurons in amphetamine-treated rats at w10 was observed in the dorsolateral caudate/putamen; in the case of the FRAs, the increase was observed in the nucleus accumbens shell. In addition, the time-dependent profile of IEG expression paralleled the activation of an upstream regulator, cAMP-response element binding protein, in the ventral striatum after haloperidol treatment. These neurochemical changes may be associated with behavioral plasticity, since amphetamine-treated rats displayed a lower amount of locomotor activity when exposed to a novel environment at w3, but had recovered at w10. Overall, the current study reveals that there is a distinct temporal and spatial profile of haloperidol-induced IEG expression and/or CREB phosphorylation in amphetamine treated rats, suggesting that there is a critical transition between the early and late withdrawal periods. PMID- 12125046 TI - Further postmortem autoradiographic studies of AMPA receptor binding in schizophrenia. AB - Previous research indicated an increased binding of AMPA receptor ligand [(3)H]CNQX at 50 nM in the caudate nucleus of schizophrenics and suicides relative to normal and neuroleptic-treated controls. The current work aimed to replicate this finding in a larger, independent sample of schizophrenics and controls. In addition to neostriatal structures, the hippocampal region and amygdala were also studied. Postmortem frozen sections from 15 schizophrenics (four suicides), 15 normal controls, 15 bipolars (eight suicides), and 15 unipolar nonpsychotic depressed (seven suicides) subjects were studied with quantitative autoradiographic procedures at 5, 20, and 50 nM [(3)H]CNQX in the striatum and at 20 nM in medial temporal structures. [(3)H]KA (kainic acid) binding was also examined. Instead of an expected increase, schizophrenics in this sample have a lower degree of [(3)H]CNQX binding in caudate and nucleus accumbens at 20 nM and in the nucleus accumbens at 50 nM, with suicided schizophrenics having higher binding than nonsuicided schizophrenics at the 20 nM concentration of [(3)H]CNQX in the caudate. [(3)H]CNQX binding was uniform across diagnostic categories at 5 nM in the striatum and at 20 nM in amygdala and hippocampal structures. KA receptor binding also did not differ among groups in any structures examined. These assays in a larger sample at three different concentrations do not support the previously reported increase in binding to AMPA receptors in schizophrenia but rather indicate an abnormal decrease in binding to this receptor in this sample. Possible explanations for the disparity in results between the two studies are considered. The data continue to indicate pathology of AMPA glutamate receptors in striatal structures in schizophrenia; however, it may be variable and its precise nature remains to be clarified. PMID- 12125047 TI - Differential distribution of PDE4D splice variant mRNAs in rat brain suggests association with specific pathways and presynaptical localization. AB - cAMP plays an important role as a second-messenger molecule controlling multiple cellular processes. Its hydrolysis provides an important mechanism by which cAMP levels are regulated. This is performed by a large multigene family of cyclic nucleotide phosphodiesterases (PDEs). Members of the PDE4 enzyme family are selectively inhibited by rolipram. Five different mRNA splice forms for PDE4D have been isolated. Here, we analyzed the regional distribution of the mRNAs coding for the splice variants PDE4D1, PDE4D2, PDE4D3, PDE4D4, and PDE4D5 in the rat brain by in situ hybridization histochemistry using specific radiolabeled oligonucleotides. We found that all five splice variants showed a distinct distribution pattern and, in some cases, in association with specific brain pathways. The most relevant differences were in hippocampal formation, medial habenula, basal ganglia, and area postrema, at both the regional and cellular level. The dorsal and median raphe nuclei exclusively contained PDE4D2 mRNA transcripts, probably located on serotonergic cells. PDE4D1 mRNA was expressed in some white matter cells. PDE4D1 and PDE4D2 mRNA splice forms presented a similar distribution in the area postrema, whereas for PDE4D4 and PDE4D5 the cellular distribution presented a complementary pattern. The differential expression of PDE4D mRNA splice variants in the area postrema is consistent with their possible involvement in emesis control and suggests new molecular targets for a more selective drug design. PMID- 12125048 TI - Molecular characterization of Saccharomyces cerevisiae Sco2p reveals a high degree of redundancy with Sco1p. AB - The Saccharomyces cerevisiae gene SCO1 has been shown to play an essential role in the transfer of copper to the Cu(A)-centre of the mitochondrial cytochrome c oxidase subunit Cox2p. By contrast, the function of Sco2p, the gene product of the highly homologous SCO2 gene, remains to be elucidated. Deletion of the SCO2 gene does not affect growth on a variety of carbon sources, including glycerol, lactate and ethanol. We report here, that Sco2p is anchored in the mitochondrial membrane by a single transmembrane segment and displays a similar tripartite structure as Sco1p. Most parts of Sco1p can be replaced by the homologous parts of Sco2p without loss of function. A short stretch of 13 amino acids, immediately adjacent to the transmembrane region, is crucial for Sco1p function and cannot be replaced by its Sco2p counterpart. We propose that this region is relevant for the correct spatial orientation of the C-terminal part of the protein. Immunoprecipitation and in vitro binding assays show that Sco2p interacts with the C-terminal portion of Cox2p. This interaction is neither dependent on bound copper ions nor on the presence of Sco1p. Furthermore we report on in vitro binding assays which show that Sco2p can form homomeric complexes, but also heteromeric complexes with Sco1p. Our data suggest that Sco2p is involved in the transfer of copper to Cox2p, but that this activity is insufficient for oxidative growth and not able to substitute for Sco1p activity. PMID- 12125049 TI - Dissection of the promoter of the HAP4 gene in S. cerevisiae unveils a complex regulatory framework of transcriptional regulation. AB - In S. cerevisiae, the heteromeric Hap2/3/4/5 complex is necessary for induced transcription of a large number of genes involved in oxidative metabolism on non fermentable carbon sources. The Hap4p subunit is the activator subunit and at the same time also the regulatory part of the complex, since it is the only one whose level is regulated by carbon source itself. HAP4 promoter analysis shows a 265 bp activating region at position -1006/-741 bp upstream of the ATG start codon. Specific and differential protein-binding to a 30 nt CSRE-like sequence within this region was observed with extracts from repressing and inducing carbon sources. Carbon source-dependent activation mediated by the 265 bp fragment, as well as protein binding to the 30 nt CSRE-like region, is dependent on the presence of CAT8 function, unveiling a complex framework by which the expression of the HAP4 gene is coordinated. PMID- 12125050 TI - 6-phosphofructokinase from Pichia pastoris: purification, kinetic and molecular characterization of the enzyme. AB - 6-Phosphofructokinase from Pichia pastoris was purified for the first time to homogeneity applying seven steps, including pseudo-affinity dye-ligand chromatography on Procion Blue H-5R-Sepharose. The specific activity of the purified enzyme was about 80 U/mg. It behaves as a typically allosteric 6 phosphofructokinase exhibiting activation by AMP and fructose 2,6-bis(phosphate), inhibition by ATP and cooperativity to fructose 6-phosphate. However, in comparison with the enzymes from Saccharomyces cerevisiae and Kluyveromyces lactis, the activation ratio of 6-phosphofructokinase from Pichia pastoris by AMP is several times higher, the ATP inhibition is stronger and the apparent affinity to fructose 6-phosphate is significantly lower. Aqueous two-phase affinity partitioning with Cibacron Blue F3G-A did not reflect remarkable structural differences of the nucleotide binding sites of the Pfks from Pichia pastoris and Saccharomyces cerevisiae. The structural organisation of the active enzyme seems to be different in comparison with hetero-octameric 6-phosphofructokinases from other yeast species. The enzyme was found to be a hetero-oligomer with an molecular mass of 975 kDa (sedimentation equilibrium measurements) consisting of two distinct types of subunits in an equimolar ratio with molecular masses of 113 kDa and 98 kDa (SDS-PAGE), respectively, and a third non-covalently complexed protein component (34 kDa, SDS-PAGE). The latter seems to be necessary for the catalytic activity of the enzyme. Sequencing of the N-terminus (VTKDSIXRDLEXENXGXXFF) and of peptide fragments by applying MALDI-TOF PSD, m/z 1517.3 (DAMNVVNH) and m/z 2177.2 [AQNCNVC(L/I)SVHEAHTM] gave no relevant information about the identity of this protein. PMID- 12125051 TI - Molecular genetics of 6-phosphofructokinase in Pichia pastoris. AB - Previously, studies on glucose-induced microautophagy in the methylotrophic yeast Pichia pastoris provided evidence that the glucose-induced selective autophagy-1 protein is the alpha-subunit of 6-phosphofructokinase (Pfk), a key enzyme in the glycolytic pathway. In our work, we could clearly demonstrate that two types of subunits of Pfk exist in P. pastoris. Investigating the yeast cell-free extract by Western blot analysis, two distinct signals of Pfk were obtained. In addition, we isolated a DNA sequence containing the complete ORF of PpPFK2 encoding the beta-subunit of Pfk from P. pastoris with a deduced molecular mass of 103.7 kDa. On the basis of these results, a hetero-oligomeric structure of Pfk in P. pastoris became obvious. Because the molecular and kinetic properties of a homo oligomeric yeast Pfk appear to be more similar to those of mammalian Pfk, as described in the literature, our results are of interest for the growing number of studies on P. pastoris as a heterologous production system. Furthermore, the 3'- and 5'-non-coding regions of PpPFK2 were isolated and several putative binding sites for regulatory factors could be identified in the promoter region. PMID- 12125052 TI - Rapid differentiation of Candida albicans from other Candida species using its unique germ tube formation at 39 degrees C. AB - In this study, we found that no Candida species other than C. albicans is able to form germ tubes at 39 degrees C in serum-free YEPD (1% (w/v) yeast extract, 2% (w/v) peptone and 2% (w/v) dextrose) media, which makes it easy to identify C. albicans from other Candida species. When cultivated in rabbit serum for at least 2 h at 37 degrees C, more than 60% of C. albicans cells generated germ tubes. In YEPD, however, germ tubes began to appear from C. albicans cells within 30 min at 39 degrees C, and more than 60% of C. albicans cells formed the germ tubes after 1 h at 39 degrees C. Standard Candida strains (ATCC, CBS), three C. albicans and two C. dubliniensis strains were cultured in serum at 37 degrees C for 2 h and in YEPD at 39 degrees C for 1 h. All of the three C. albicans formed germ tubes at 39 degrees C. The two C. dubliniensis strains formed germ tubes in serum at 37 degrees C, but grew as a yeast form in YEPD at 39 degrees C. All of the clinically isolated C. albicans strains in our laboratory formed germ tubes in YEPD at 39 degrees C for 1 h, and none of the clinically isolated Candida species other than C. albicans generated germ tubes in YEPD at 39 degrees C. Thus, the unique germ tube formation of C. albicans induced by high temperature (39 degrees C) in YEPD could be applied to a protocol for the rapid and convenient identification of C. albicans in clinical laboratories. PMID- 12125053 TI - Characterization of a chaperone ClpB homologue of Paracoccidioides brasiliensis. AB - We report the cloning and sequence analysis of a genomic clone encoding a Paracoccidioides brasiliensis ClpB chaperone homologue (PbClpB). The clpb gene was identified in a lambda Dash II library. Sequencing of Pbclpb revealed a long open reading frame capable of encoding a 792 amino acid, 87.9 kDa protein, pI of 5.34. The predicted polypeptide contains several consensus motifs of the ClpB proteins. Canonical sequences such as two putative nucleotide-binding sites, chaperonins ClpA/B signatures and highly conserved casein kinase phosphorylation domains are present. ClpB is 69% to 49% identical to members of the ClpB family from several organisms from prokaryotes to eukaryotes. The transcript of PbclpB was detected as a mRNA species of 3.0 kb, preferentially expressed in the yeast parasitic phase of the fungus. A 89 kDa protein was also detected in yeast cells of P. brasiliensis. PMID- 12125054 TI - PCR-based identification of pathogenic Candida species using primer mixes specific to Candida DNA topoisomerase II genes. AB - For rapid identification of Candida to the species level, degenerated primers and specific primers based on the genomic sequences of DNA topoisomerase II of C. albicans, C. dubliniensis, C. tropicalis (genotypes I and II), C. parapsilosis (genotypes I and II), C. krusei, C. kefyr, C. guilliermondii, C. glabrata, C. lusitaniae and Y. lipolytica were designed and their specificities tested in PCR based identifications. Each of the specific primers selectively and exclusively amplified its own DNA fragment, not only from the corresponding genomic DNA of the Candida sp. but also from DNA mixtures containing other DNAs from several fungal species. For a simpler PCR-based identification, the specific primers were divided into three groups (PsI, PsII and PsIII), each of which contained four specific primer pairs. PCR with the primer mixes yielded four different sizes of PCR product, corresponding to each Candida sp. in the sample DNA. To obtain higher sensitivity of PCR amplification, sample DNAs were preamplified by the degenerated primer pair (CDF28/CDR148), followed by the main amplification using the primer mixes. By including this nested PCR step, 40 fg yeast genomic DNA was detected in the sample. Furthermore, we applied this nested PCR to a clinical diagnosis, using splenic tissues from experimentally infected mice and several clinical materials from patients. In all cases, the nested PCR amplifications detected proper DNA fragments of Candida spp., which were also identified by the standard identification tests. These results suggest that nested PCR, using primer mixes of the Candida DNA topoisomerase II genes, is simple and feasible for the rapid detection/identification of Candida to species level in clinical materials. PMID- 12125056 TI - Current awareness on yeast. PMID- 12125055 TI - Genomic differences between Candida glabrata and Saccharomyces cerevisiae around the MRPL28 and GCN3 loci. AB - We report the sequences of two genomic regions from the pathogenic yeast Candida glabrata and their comparison to Saccharomyces cerevisiae. A 3 kb region from C. glabrata was sequenced that contains homologues of the S. cerevisiae genes TFB3, MRPL28 and STP1. The equivalent region in S. cerevisiae includes a fourth gene, MFA1, coding for mating factor a. The absence of MFA1 is consistent with C. glabrata's asexual life cycle, although we cannot exclude the possibility that a factor gene(s) are located somewhere else in its genome. We also report the sequence of a 16 kb region from C. glabrata that contains a five-gene cluster similar to S. cerevisiae chromosome XI (including GCN3) followed by a four-gene cluster similar to chromosome XV (including HIS3). A small-scale rearrangement of gene order has occurred in the chromosome XI-like section. PMID- 12125057 TI - Two juvenile hormone suppressible storage proteins may play different roles in Hyphantria cunea Drury. AB - We isolated and sequenced cDNA clones corresponding to two storage proteins (HcSP 1 and HcSP-2) from fall webworm, Hyphantria cunea. The cDNAs for HcSP-1 (2,337 bp) and HcSP-2 (2,572 bp) code for 753 and 747 residue proteins with predicted molecular masses of 88.3 and 88.5 kDa, respectively. The calculated isoelectric points are pI = 8.4 (HcSP-1) and 7.6 (HcSP-2). Multiple alignment analysis of the amino acid sequence revealed that HcSP-1 is most similar to SL-1 from S. litura (73.8% identity) and other methionine-rich hexamers, whereas HcSP-2 is most similar to the SL-2 alpha subunit from S. litura (74.8% identity) and other moderately methionine-rich hexamers. The two storage proteins from H. cunea shared only 38.4% identity with one another. According to both phylogenetic analyses and the criteria of amino acid composition, HcSP-1 belongs to the subfamily of Met-rich storage proteins (6% methionine, 10% aromatic amino acid), and HcSP-2 belongs to the subfamily of moderately methionine-rich storage proteins (3.2% methionine, 12.9% aromatic amino acid). Topical application of the JH analog, methoprene, after head ligation of larvae, suppressed transcription of the SP genes, indicating hormonal effects at the transcriptional level. The HcSP 1 transcript was detected by Northern blot analysis in Malpighian tubule, testis, and ovary, in addition to fat body where it was most abundant. The HcSP-2 transcript was detected only in fat body and Malpighian tubule. The accumulation of HcSP-1 in ovary and HcSP-2 in Malpighian tubule might be related to differential functions in both organs. PMID- 12125058 TI - Characterisation of aphid myrosinase and degradation studies of glucosinolates. AB - Myrosinase from Brevicoryne brassicae was purified by ammonium sulfate fractionation, dialysis, and chromatography on a DEAE column. The chromatography yielded a single peak and a 115.6-fold purification. Further FPLC gel filtration gave a single peak at 120 kDa. Denaturing SDS/PAGE of the protein revealed a single band at 60 kDa, indicating that the native B. brassicae myrosinase is a dimer. Kinetic parameters towards 8 glucosinolates were calculated. Strong differences of V(max) and K(m) were observed depending on the substrate. Degradation products of each glucosinolate were identified and quantified by GC MS and GLC-FID, respectively. Using both crude aphid homogenates and purified myrosinase, two unique hydroxyglucosinolates, 3-butenyl- and benzyl isothiocyanates were identified from progoitrin ((2S)-2-hydroxybut-3-enyl glucosinolate) and sinalbin (4-hydroxybenzyl-glucosinolate) degradation respectively. Addition of ascorbic acid to the reaction mixtures containing sinalbin and progoitrin caused the production of hydroxylated degradation products usually associated with plant myrosinase metabolisation. The occurrence of the myrosinase system in B. brassicae is discussed in terms of similar allelochemical adaptation between the herbivore and its host plant. PMID- 12125059 TI - Cloning and characterization of Acmar1, a mariner-like element in the asiatic honey bee, Apis cerana japonica (Hymenoptera, Apocrita). AB - A mariner-like element was cloned from the genome of the Asiatic honey bee, Apis cerana japonica (Hymenoptera, Apocrita). The (composite) clone, named Acmar1, was 1,378 bp long, and encoded 336 amino acids corresponding to a transposase-like putative polypeptide in a single open reading frame. The D,D(34)D motif, the catalytic domain of the mariner transposase, was present, although there was a deletion of five amino acid residues within it as compared with the active transposase in Drosophila mauritiana. Nineteen-bp-long imperfect inverted terminal repeat-like sequences flanked by TA dinucleotides, the typical target site for mariner insertion, were observed. Southern blot analysis using a fragment covering two-thirds of the Acmar1 transposase coding sequence as a probe indicated the presence of multiple Acmar1-like elements in the genome. Maximum parsimony phylogenetic analysis based on the transposase amino acid sequences of insect mariner-like elements revealed that Acmar1 is a member of the mellifera subfamily. PMID- 12125060 TI - Cloning, partial purification and in vivo developmental profile of expression of the juvenile hormone epoxide hydrolase of Ctenocephalides felis. AB - cDNAs encoding two different epoxide hydrolases (nCfEH1 and nCfEH2) were cloned from a cDNA library prepared from the wandering larval stage of the cat flea, Ctenocephalides felis. Predicted translations of the open reading frames indicated the clones encoded proteins of 464 (CfEH1) and 465 (CfEH2) amino acids. These proteins have a predicted molecular weight of 53 kDa and a putative 22 amino acid N-terminal hydrophobic membrane anchor. The amino acid sequences are 77% identical, and both are homologous to previously isolated epoxide hydrolases from Manduca sexta, Trichoplusia ni, and Rattus norvegicus. Purification of native juvenile hormone epoxide hydrolase (JHEH) from unfed adult cat fleas generated a partially pure protein that hydrolyzed juvenile hormone III to juvenile hormone III-diol. The amino terminal sequence of this;50-kDa protein is identical to the deduced amino terminus of the protein encoded by the nCfEH1 clone. Affinity-purified rabbit polyclonal antibodies raised against Escherichia coli-expressed HisCfEH1 recognized a approximately 50-kDa protein present in the partially purified fraction containing JHEH activity. Immunohistochemistry experiments using the same affinity-purified rabbit polyclonal antibodies localized the epoxide hydrolase in developing oocytes, fat body, and midgut epithelium of the adult flea. The presence of JHEH in various flea life stages and tissues was assessed by Northern blot and enzymatic activity assays. JHEH mRNA expression remained relatively constant throughout the different flea larval stages and was slightly elevated in the unfed adult flea. JHEH enzymatic activity was highest in the late larval, pupal, and adult stages. In all stages and tissues examined, JHEH activity was significantly lower than juvenile hormone esterase (JHE) activity, the other enzyme responsible for JH catalysis. PMID- 12125061 TI - Pelvic and hindlimb musculature of Tyrannosaurus rex (Dinosauria: Theropoda). AB - In this article, we develop a new reconstruction of the pelvic and hindlimb muscles of the large theropod dinosaur Tyrannosaurus rex. Our new reconstruction relies primarily on direct examination of both extant and fossil turtles, lepidosaurs, and archosaurs. These observations are placed into a phylogenetic context and data from extant taxa are used to constrain inferences concerning the soft-tissue structures in T. rex. Using this extant phylogenetic bracket, we are able to offer well-supported inferences concerning most of the hindlimb musculature in this taxon. We also refrain from making any inferences for certain muscles where the resulting optimizations are ambiguous. This reconstruction differs from several previous attempts and we evaluate these discrepancies. In addition to providing a new and more detailed understanding of the hindlimb morphology of T. rex--the largest known terrestrial biped--this reconstruction also helps to clarify the sequence of character-state change along the line to extant birds. PMID- 12125062 TI - Osteology and myology of the cephalic region and pectoral girdle of the Chinese catfish Cranoglanis bouderius, with a discussion on the autapomorphies and phylogenetic relationships of the Cranoglanididae (Teleostei: Siluriformes). AB - The cephalic and pectoral girdle structures of the Chinese catfish Cranoglanis bouderius are described and compared with those of other catfishes as the foundation for an analysis on the Cranoglanididae autapomorphies and also for a discussion on the phylogenetic relationships between the cranoglanidids and the other catfishes. Our observations and comparisons indicate that cranoglanidids are defined, at least, by four autapomorphies, namely: 1) the cartilages associated with the mandibular barbels are broad, somewhat circular; 2) epioccipital with a well-developed posterodorsal process, which presents a large, deep, circular posterior concavity; 3) a well-defined, deep, anteroposteriorly elongated concavity formed by both the frontal and the lateral ethmoid to receive the anteromedial surface of the metapterygoid; 4) the adductor mandibulae A3" is dorsally divided into two bundles and partially inserted on the posterior portion of the primordial ligament. With respect to the phylogenetic relationships of the Cranoglanididae, this study strongly suggests that these fishes are probably closely related to the Ariidae and the Claroteidae. PMID- 12125063 TI - Ultrastructural observations of eu- and paraspermiogenesis in the cottid fish Hemilepidotus gilberti (Teleostei: Scorpaeniformes: Cottidae). AB - The developmental process of eu- and paraspermatozoa in the cottid fish, Hemilepidotus gilberti, was observed by electron microscopy. Euspermatozoa of H. gilberti consist of a thin disk-like sperm head (about 3 microm in length), a short middle piece, and a long flagellum, but lack an acrosome. On the other hand, during spermiogenesis, aberrant spermatids, rich in cytoplasm and possessing binuclei, develop into cysts containing spermatids. The developing aberrant spermatids connect with normal spermatids and euspermatozoa by intercellular bridges. The early phase of chromatin condensation in aberrant spermatids is almost identical to that in normal spermatids, but the nuclei in the later phase develop into a mass of highly electron-dense globules. Since the aberrant spermatids complete karyokinesis but not cytokinesis at telophase of the second meiotic division, they are considered to develop into hyperpyrenic cells due to incomplete cytokinesis of the second meiotic division. These spermatids are oval in shape (5-7 microm in diameter) and lack a flagellum. The aberrant spermatids of H. gilberti are shed along with euspermatozoa and amount to about 50% of semen in volume. Judging from their form and developmental process, aberrant spermatids produced in H. gilberti are considered hyperpyrenic paraspermatozoa. PMID- 12125064 TI - Fine structure and probable function of ring organs in the mite Histiostoma feroniarum (Acari: Actinotrichida: Acaridida: Histiostomatidae). AB - Histiostoma feroniarum, like other histiostomatid mites, possesses peculiar ring organs that are visible under the light microscope as ventrally located, characteristic rings of sclerotized cuticle. The ring organ is composed of three elements: a disc of modified cuticle, ring organ cells located underneath the disc, and an "empty" chamber frequently visible between the cuticular disc and the cells. The cuticle of the disc is not perforated and differs from the surrounding unmodified cuticle as revealed by special staining developed for light microscopy and by electron microscopy. The ring organ cells show a polarity, with a practically smooth apical surface and an extremely folded basal membrane. The basal invaginations reach the apical cell portion, where they form tubular canaliculi distributed beneath the apical cell membrane. The cytoplasm contains many mitochondria, which are usually in contact with the cell membrane invaginations. Structurally, the ring organ cells closely resemble the transport cells described in osmoregulatory organs both in water-inhabiting and terrestrial arthropods. Thus, our results support earlier suggestions of an osmoregulatory function performed by sclerotized rings (=ring organs), as an adaptation to aqueous environments. A possible homology with similar organs of other mites is discussed. PMID- 12125065 TI - Ultrastructure of spermatozoa of the lizard Ameiva ameiva, with considerations on polymorphism within the family Teiidae (Squamata). AB - A detailed description of sperm ultrastructure of the lizard Ameiva ameiva (Teiidae) is provided. Mature spermatozoa are characterized by: a depressed acrosome at the anterior portion; a unilateral ridge at the anterolateral portion; an acrosome vesicle divided into cortex and medulla; medulla divided into two regions with different electron-densities; paracrystalline subacrosomal material with radial organization in transverse section; a pointed prenuclear perforatorium; a stopper-like perforatorium base plate that appears embedded in the subacrosomal material; the presence of an epinuclear lucent zone surrounded by its own membrane; a large nuclear rostrum; round nuclear shoulders; a nuclear space at the nucleus tip; a bilateral stratified laminar structure; a central dense body within the proximal centriole; a short midpiece; an axonemal midpiece axial component; peripheral fibers 3 and 8 grossly enlarged at the anterior portion of axoneme; columnar mitochondria with linear cristae; solid dense bodies arranged as rings or spirals; a triangular-shaped annulus in transverse section; a fibrous sheath into the midpiece; a thin zone of cytoplasm at the anterior portion of the principal piece; and a slight decrease in diameter of the principal piece immediately after the annulus. Comparisons with Cnemidophorus sexlineatus and Micrablepharus maximiliani failed to identify unique sperm ultrastructure traits of Teiidae or Teiioidea (Teiidae + Gymnophthalmidae). High levels of polymorphism between Ameiva and Cnemidophorus, two closely related genera of the family Teiidae, were detected, suggesting that extensive sampling within squamate families is essential if sperm ultrastructure data are to be used in phylogenetic analyses at this taxonomic level. PMID- 12125066 TI - Morphology and evolution of the respiratory apparatus in the family Eubelidae (Crustacea, Isopoda, Oniscidea). AB - The morphology of the respiratory apparatus in the pleopodal lungs of the family Eubelidae was investigated. The family is a monophyletic group including more than 240 species in 53 genera (three of which are nomina dubia), mostly distributed in the Afrotropical Region (tropical Africa and Arabian Peninsula). In all the Eubelidae, except for the monospecific genus Parelumoides and two species of the genus Elumoides, the exopods of pleopods have lungs. All the pulmonary morphologies present in the entire suborder Oniscidea are found: 1) uncovered lungs, composed of a pleated respiratory surface, directly exposed to the air (Atracheodillo-type) or partially enclosed within the appendage (Synarmadilloides-type); 2) covered lungs with several spiracles and respiratory trees, housed within the appendages, with spiracles surrounded by a specialized, nonrespiratory, structure (perispiracular area) (Eubelum- and Somaloniscus types); 3) covered lungs with only one spiracle, with or without perispiracular area, and one respiratory tree (Aethiopopactes- and Periscyphis-types), which in taxa with Periscyphis-type lung crosses the insertion of the appendage and penetrates into the pleon with bundles of respiratory tubules. The evolution of the various types of lungs is discussed. It is concluded that the two main evolutionary lines, i.e., uncovered lungs and covered lungs, originated independently from an ancestral respiratory structure-the semilunar area. A first mechanism of development of the semilunar area by folding of its surface produced the Atracheodillo-type (all folds coplanar with the surface of the exopod) and Synarmadilloides-type (folds partly coplanar and partly intraflexed inside the exopod) uncovered lungs. A second mechanism of development by tubular invagination of the cuticle of the semilunar area produced the polyspiracular Eubelum-type lungs (numerous arborescent invaginations) and the monospiracular Aethiopopactes-type lungs (only one arborescent invagination), probably passing through a common intermediate pattern. From the common pattern, both the poly- and monospiracular types would have inherited the characteristic concave cell arrangement of the perispiracular area. The Somaloniscus-type and Periscyphis type lungs are forms specialized for arid environments, directly derived from the Eubelum-type and Aethiopopactes-type, respectively. PMID- 12125067 TI - Architecture of the integument in lower teleostomes: functional morphology and evolutionary implications. AB - A bony ganoid squamation is the plesiomorphic type in actinopterygians. During evolution, it was replaced by weak and more flexible elasmoid scales. We provide a comparative description of the integument of "ganoid" fishes and "nonganoid" fishes that considers all dermal components of mechanical significance (stratum compactum, morphology of ganoid scales, and their regional differences) in order to develop a functional understanding of the ganoid integument as a whole. Data were obtained for the extant "ganoid" fishes (Polypteridae and Lepisosteidae) and for closely related "lower" actinopterygians (Acipenser ruthenus, Amia calva) and "lower" sarcopterygians (Latimeria chalumnae, Neoceratodus forsteri). Body curvatures during steady undulatory locomotion, sharp turns, prey-strikes, and fast starts in "ganoid" fishes were measured from videotapes. Extreme body curvatures as measured in anesthetized specimens are never reached during steady swimming, but are sometimes closely approached in certain situations (sharp turns, prey-strike). During extreme body curvatures we measured high values of lateral strain on the convex and on the concave side of the body. Scale overlap changes considerably (66-127% in Lepisosteus, 42-140% in Polypterus). The ganoid squamation forms a protective coat, but at the same time it permits extreme body curvatures. This is reflected in characteristic morphological features of the ganoid scales, such as an anterior process, concave anterior margin, and peg-and socket articulation. These characters are most pronounced in the anterior body region, where maximum changes in scale overlap are required. The anterior processes and anterior concave margin, together with the attached stratum compactum, guide movements in a horizontal plane during bending. Displacements of scales relative to each other are possible for scales of different scale rows, but are impeded in scales of the same scale row due to the peg-and-socket articulation. Furthermore, ganoid scale rows, fibers of collagen layers of the stratum compactum, and the lateral myoseptal structures follow the same oblique orientation, which is needed to achieve extreme body curvatures. There is no evidence that body curvatures are limited by the ganoid squamation in Polypterus or Lepisosteus to any larger extent than by a type of integument devoid of ganoid scales in teleostomes of similar body shape. Our results essentially contradict former functional interpretations: 1) Ganoid scales do not especially limit body curvature during steady undulatory locomotion; 2) They do not act as torsion resisting devices, but may be able to damp torsion together with the stratum compactum and internal body pressure. PMID- 12125068 TI - New characters in the Gnathostomulid mouth parts revealed by scanning electron microscopy. AB - An analysis with SEM of the mouth parts of 16 species belonging to 10 genera of Gnathostomulida resulted in the following new characters and conclusions: 1) At least in the genus Haplognathia, jaw teeth that are visible by conventional light microscopy are composed of the same aggregated needle-like denticles that are found, often in large numbers, on the basal plates of many filospermoid species. 2) Other new ultrastructural tooth features include the tricuspid basal plate teeth in Problognathia minima, tripartite teeth in Austrognathia and Austrognatharia, and the clear separation, in the Gnathostomula basal plate, of a mediodorsal set of teeth from a more extensive rostroventral set. 3) Three rows of teeth, as typical for Gnathostomulidae and Austrognathiidae, are also present in the filospermoid Haplognathia filum. 4) The wide range of geographic variation in Haplognathia ruberrima is confirmed by significant differences in jaw teeth between specimens from Belize and Bermuda. 5) A compartmentalized involucrum is present in Labidgonathia longicollis. 6) A pair of lamellae addentales, until now only known from Valvognathia pogonostoma, was found in Tenuignathia rikerae, Problognathia minima, and probably also Labidognathia rikerae. 7) In all gnathostomulids, the lamella symphysis is composed of identical rods that are considered homologous with those in the mouth parts of Rotifera and Micrognathozoa. PMID- 12125069 TI - Ill-fated amyloid-beta vaccine. PMID- 12125070 TI - The integrin family of cell adhesion molecules has multiple functions within the CNS. AB - Integrins comprise a large family of cell adhesion molecules that mediate interactions between the extracellular environment and the cytoplasm. During the last decade, analysis of the expression and function of these molecules has revealed that integrins regulate many aspects of cell behavior including cell death, proliferation, migration, and differentiation. Within the central nervous system (CNS), most of the early studies focused on the role of integrins in mediating adhesive and migratory events in two distinct processes: neural development and CNS inflammation. Interestingly, recent analysis of transgenic mice has provided some surprising results regarding the role of integrins in neural development. Furthermore, a large body of evidence now supports the idea that in addition to these well-described functions, integrins play multiple roles in the CNS, both during development and in the adult in areas as diverse as synaptogenesis, activation of microglia, and stabilization of the endothelium and blood-brain barrier. Many excellent reviews have addressed the contribution of integrins in mediating leukocyte extravasation during CNS inflammation. This review will focus on recently emerging evidence of novel and diverse roles of integrins and their ligands in the CNS during development and in the adult, in health and disease. PMID- 12125071 TI - Neural (N-) cadherin, a synaptic adhesion molecule, is induced in hippocampal mossy fiber axonal sprouts by seizure. AB - Aberrant mossy fiber sprouting and synaptic reorganization are plastic responses in human temporal lobe epilepsy, and in pilocarpine-induced epilepsy in rodents. Although the morphological features of the hippocampal epileptic reaction have been well documented, the molecular mechanisms underlying these structural changes are not understood. The classic cadherins, calcium-dependent cell adhesion molecules, are known to function in development in neurite outgrowth, synapse formation, and stabilization. In pilocarpine-induced status epilepticus, the expression of N-cadherin mRNA was sharply upregulated and reached a maximum level (1- to 2.5-fold) at 1- to 4 weeks postseizure in the granule cell layer and the pyramidal cell layer of CA3. N-cadherin protein was correspondingly increased and became concentrated in the inner molecular layer of the dentate gyrus, consistent with the position of mossy fiber axonal sprouts. Moreover, N-cadherin labeling was punctate; colocalized with definitive synaptic markers, and partially localized on polysialated forms of neural cell adhesion molecule (PSA NCAM)-positive dendrites of granule cells in the inner molecular layer. Our findings show that N-cadherin is likely to be a key factor in responsive synaptogenesis following status epilepticus, where it functions as a mediator of de novo synapse formation. PMID- 12125073 TI - Glutathione release from cultured brain cells: multidrug resistance protein 1 mediates the release of GSH from rat astroglial cells. AB - To investigate the release of glutathione (GSH) from brain cells, cultures enriched for astroglial cells, neurons, oligodendroglial cells, and microglial cells derived from rat brain were studied. During incubation of astroglial cultures, GSH accumulated in the medium with a rate of 3.1 +/- 0.6 nmol x h(-1) x mg protein(-1). In contrast, only marginal amounts of extracellular GSH were detectable in the media of the other brain cell cultures investigated. The mechanism of GSH release from astroglial cells, as yet, has not been reported. Multidrug resistance protein 1 (Mrp1), a transport protein known to mediate cellular export of glutathione disulfide and glutathione conjugates, is expressed in astroglial cultures. Inhibitors of Mrp1 were used to test for a function of this transporter in mediating GSH release from astroglial cells. The presence of the competitive Mrp1 inhibitor MK571 at a concentration of 50 microM inhibited the rate of GSH release by 63%. In contrast, the low concentration of 1 microM of MK571 increased the rate of GSH release by 83%. This bimodal concentration dependent effect of MK571 is in accord with literature data for the effects of Mrp1 substrates on GSH release from cells. In addition, the presence of cyclosporin A (10 microM) reduced the GSH release rate significantly and completely blocked the stimulating effect of 1 microM MK571 on the release of GSH from astroglial cells. In conclusion, the data presented are a strong indication that Mrp1 participates in the release of GSH from astroglial cells. PMID- 12125072 TI - CNS myelinogenesis in vitro: myelin basic protein deficient shiverer oligodendrocytes. AB - The shiverer mutant mouse is an autosomal recessive mutant characterized by incomplete myelin sheath formation in the central nervous system (CNS). Such mice contain a deletion in the MBP gene, do not produce MBP proteins, and have little or no compact myelin in the CNS. To investigate the myelin sheath formation in shiverer mutant mice resulting from the absence of compact myelin, firstly we developed new methods for generating oligodendrocyte precursor cells (OPCs) from an E17 mouse brain, and examined homozygous shiverer (shi/shi) OPCs with respect to myelinogenesis in vitro. After treatment of shi/shi OPCs in vitro with PDGF or bFGF, proliferation of shi/shi OPCs was enhanced similar to that observed in wild type OPCs. The majority of cells from the shiverer mutant mouse, however, remained as A2B5-immunoreactive early OPCs. To determine which molecular events affect the differentiation of shi/shi OPCs, we determined the signaling pathway that could be responsible for activating myelin sheath-specific proteins. We found that the developmental schedule of shi/shi OPCs in vitro was accelerated by the addition of cyclic AMP analogs, dibutyryl cAMP (dbcAMP). Treatment of shi/shi OPCs with dbcAMP had significant effect on the differentiation of OPCs that became MAG-expressing oligodendrocytes. To further determine the possible mechanism involved in the activation of MAG by dbcAMP, we examined the cAMP dependent signaling cascades. The activation of JNK was markedly stimulated by treatment with dbcAMP, and the phosphorylation of transcription factor ATF-2 was also stimulated by dbcAMP. We demonstrated that the MAG-positive shi/shi oligodendrocytes extend processes around axons and finally covered the axon, this was clearly observed by immunocytochemistry of shi/shi oligodendrocyte-DRG cocultures. These results suggest that ATF-2 coupled to specific signal transduction cascades plays an important regulatory role in MAG expression at a specific stage of shi/shi oligodendrocyte differentiation, and OPCs grow to become myelin-forming cells with numerous cell processes that wraps around an axon to form a thin myelin sheath. PMID- 12125074 TI - Neuronal nitric oxide synthase modulates basal catecholamine secretion in bovine chromaffin cells. AB - The role of endogenously produced nitric oxide (NO) in the regulation of basal catecholamine (CA) secretion was studied in chromaffin cells. Treatment of chromaffin cells with nitric oxide synthase (NOS) inhibitors produced a dose dependent increase in basal catecholamine secretion, which paralleled their ability to inhibit NOS activity. This inhibitory profile was similar to that found in neurons, suggesting the constitutive expression of neuronal NOS (nNOS) in these cells, which was confirmed by Western blot analysis. A study of the kinetics and pharmacology of nNOS activity expressed in chromaffin cells in culture indicated that NOS activity is calcium-dependent, increases with time, and is highly dependent on both intracellular concentrations of L-arginine (K(m) approximately 4 microM, V(max) = 908 +/- 60 pmol/hr x 10(6) cells) and transport of L-arginine into the cells (exhibiting two affinity constants of k(1) = 3.2 +/- 0.3 microM and k(2) = 126 +/- 5.5 microM). The effects of NOS inhibitors on CA secretion were mediated by the L-arginine-NO-cGMP pathway, insofar as exogenous L arginine was able to partially block the increase in CA secretion evoked by them, and 1H-[1,2,4]oxadiazolo[4,3-a]quinoxaline-1-one (ODQ), a specific inhibitor of guanylate cyclase, and zaprinast, an inhibitor of the cGMP phosphodiesterase, were able to increase and inhibit, respectively, basal CA secretion in a dose dependent manner. These results suggest that chromaffin cells exhibit a tonic production of NO by nNOS that keeps the basal CA secretion at low levels, and this could be necessary for maintaining a normotensive state. PMID- 12125075 TI - Transgenic expression of the protein-L-isoaspartyl methyltransferase (PIMT) gene in the brain rescues mice from the fatal epilepsy of PIMT deficiency. AB - Protein-L-isoaspartyl methyltransfearase (PIMT) plays a physiological role in the repair of damaged proteins containing isoaspartyl residues. In previous studies, we showed that PIMT-deficient mice developed a fatal epileptic seizure associated with the accumulation of damaged proteins in the brain. The mutant mice also showed a neurodegenerative pathology in hippocampi and impaired spatial memory. Still undefined, however, is how the accumulation of isoaspartates leads to the death of PIMT-deficient mice. In the present study, we generated PIMT transgenic (Tg) mice to investigate whether the exogenous expression of PIMT could improve the symptoms associated with PIMT deficiency. Rescue experiments showed that Tg expression of PIMT driven by a prion promoter effectively cured the PIMT deficient mice. Biochemically, a higher expression level of transgene led to the effective repair of damaged proteins in vivo. Although a lower level of expression caused an accumulation of damaged proteins in a partially rescued line, the mice survived. Interestingly, synapsin I, which was extensively modified posttranslationally in PIMT-deficient mice, was specifically repaired in a partially rescued, but symptom-improved, Tg line. Our results suggest that an overall accumulation of damaged proteins does not necessarily lead to a fatal epileptic seizure, whereas certain modifications, such as changes in synapsin I, may play a pivotal pathological role in epilepsy. PMID- 12125076 TI - Adenoviral expression of protein-L-isoaspartyl methyltransferase (PIMT) partially attenuates the biochemical changes in PIMT-deficient mice. AB - Protein-L-isoaspartyl methyltransferase (PIMT) is a putative protein repair enzyme, which methylates the alpha-carboxyl group of atypical L-isoaspartyl residues in aged proteins and converts them to normal L-aspartyl residues. Two splicing variants, PIMT-I and PIMT-II, have been reported, although their biological functions and specific subcellular substrates are still to be defined. We and another group have previously showed that PIMT-deficient mice succumbed to fatal epileptic seizures associated with an abnormal accumulation of isoaspartate (IsoAsp) in the brain. In the present study, we prepared two recombinant adenovirus vectors that contained PIMT-I or PIMT-II, respectively, in order to investigate the differential biological roles of PIMT-I and PIMT-II. These recombinant viruses differentially conferred PIMT-I or PIMT-II expressions in cultured neurons. Biochemical analyses showed that either of PIMT-I or PIMT-II effectively repaired the damaged proteins in PIMT-deficient neurons, but the concomitant expression failed to show an additive effect in the repair of IsoAsp. These results suggested that PIMT-I and PIMT-II might share a common biological function and/or subcellular substrates. In addition, we administered an adeno PIMT-I vector into the brain of PIMT-deficient mice at embryonic day 14.5 by an exo-utero method to assess the biological effects in vivo. The result showed that recombinant adeno-PIMT improved the symptoms of PIMT-deficient mice in vivo, but only partially repaired IsoAsp in damaged proteins. The gene therapy presented in this report provided a better prognosis for the survival of PIMT-deficient mice than the previously reported anti-epileptic drug therapy. The results suggested a new reagent for gene therapy applicable to ageing-associated neurodegenerative disorders. PMID- 12125077 TI - p35/Cdk5 pathway mediates soluble amyloid-beta peptide-induced tau phosphorylation in vitro. AB - Alzheimer's disease (AD) is pathologically characterized by deposition of amyloid beta peptides (Abeta) as senile plaques and by the occurrence of neurofibrillary tangles (NFTs) composed primarily of hyperphosphorylated tau protein. Activation of cyclin-dependent kinase 5 (Cdk5) via its potent activator p25 has recently been shown to promote phosphorylation of tau at AD-specific phosphoepitopes, and increased cleavage of p35 to p25 has been demonstrated in AD patients, suggesting that Cdk5 may represent a pathogenic tau protein kinase. We were interested in the potential effect of soluble forms of Abeta on Cdk5-mediated AD-like tau phosphorylation, insofar as previous studies of human biopsies and aged canine and primate brains have shown that dystrophic neurites appear before the formation of neuritic plaques. We transfected N2a cells with a p35 vector (N2a/p35 cells) and, after differentiation, challenged these cells with Abeta(1 42) peptide in soluble form (sAbeta(1-42)). Results show that sAbeta(1-42) at relatively low levels (1-5 microM) dose-dependently increases tau phosphorylation at AD-specific phosphoepitopes in differentiated N2a/p35 cells compared with controls, an effect that is blocked by antisense oligonucleotides against p35. sAbeta(1-42)-induced tau phosphorylation is concomitant with an increase in both p25 to p35 ratio and Cdk5 activity (but not protein levels). Additionally, blockade of L-type calcium channels or inhibition of calpain completely abolishes this effect. Taken together, these data indicate that sAbeta is a potent activator of the p25/Cdk5 pathway, resulting in promotion of AD-like tau phosphorylation in vitro. PMID- 12125078 TI - Effect of geranylgeranylaceton on cellular damage induced by proteasome inhibition in cultured spinal neurons. AB - We investigated the effect of two proteasome inhibitors, lactacystin and epoxomicin, on cultured spinal cord neurons. The incubation of spinal neurons with proteasome inhibitors for 24 hr induced neurotoxicity in a dose-dependent manner. We found motor neurons to be more vulnerable to proteasome-induced neurotoxicity than nonmotor neurons. The staining of cell bodies in treated motor neurons was markedly disrupted and showed characteristic granular patterns. Proteasome-induced neurotoxicity is accompanied by apoptotic nuclear changes, posttranslational modification of the cellular proteins, generation of intracellular free radicals, reduction in the amount of reduced glutathione, and mitochondrial dysfunction. Neurotoxicity was reduced by the administration of low concentrations (1-100 nM) of geranylgeranylacetone (GGA), which is widely used as an antiulcer drug, although higher concentrations of this drug produced neurotoxicity in spinal cord neurons. GGA was found to induce the expression of heat shock protein 70 as well as thioredoxin, which may partly contribute to the protective effect of GGA. These data suggest that the inhibition of proteasome may play a role in the mechanism of neurodegenerative diseases of the spinal cord, such as amyotrophic lateral sclerosis, and that the use of GGA may be effective in the treatment of these conditions. PMID- 12125079 TI - Comparison of fresh and cryopreserved porcine ventral mesencephalon cells transplanted in A rat model of Parkinson's disease. AB - To evaluate whether cryopreservation of porcine ventral mesencephalon cells influences graft survival and function in vivo, we have transplanted either freshly prepared or cryopreserved cells into the striatum of 6-hydroxydopamine lesioned rats. A single cell suspension of porcine ventral mesencephalon cells from the same isolation either was stored at 4 degrees C and transplanted the next day or was cryopreserved for 4 weeks in liquid nitrogen vapor. The cryopreserved cells were then rapidly thawed, rinsed, and transplanted in the same manner as the fresh cells, with the same dose of viable cells. All animals received daily injections of cyclosporin A to prevent xenograft rejection. To monitor graft function, amphetamine-induced rotation was measured every 3 weeks between 6 and 15 weeks posttransplantation. After sacrifice at 15 weeks posttransplantation, histological methods were used to compare fresh cell and cryopreserved cell transplants with respect to graft survival, differentiation and integration, and host immune response. Cryopreserved cells were found to be either equivalent or in some cases superior to fresh cells with respect to rotational correction, graft survival, graft volume, numbers of graft-derived dopaminergic neurons, and host immune responses. In conclusion, the results indicate that it is feasible to cryopreserve porcine ventral mesencephalon cells for long-term storage of cells prior to transplantation in an animal model of Parkinson's disease. PMID- 12125080 TI - Neuroprotection of glial cell line-derived neurotrophic factor in damaged spinal cords following contusive injury. AB - Glial cell line-derived neurotrophic factor (GDNF) acts as a potent survival factor for many neuronal populations, including spinal motoneurons, indicating the therapeutic promise of GDNF for neurological disorders. Injury to spinal cord (SCI) triggers processes destructive to ascending sensory and descending motor conduction and extends tissue loss, thereby leading to permanent behavioral dysfunction. In this study, we attempted to examine whether GDNF protects neurons from SCI and subsequently lessens locomotor deficit in SCI rats. We utilized the NYU weight-drop device developed at New York University to induce spinal cord contusion at the T9-10 spinal segment. After SCI, GDNF was administrated into the cord 1-2 mm rostral and caudal to the epicenter. Animals receiving GDNF treatment showed significant improvement over phosphate-buffered saline (PBS)-treated controls on the Basso Beattie Bresnahan (BBB) locomotor rating scale (P < 0.01 0.001). GDNF treatment increased the remaining neuronal fibers with calcitonin gene-related peptide, neurofilament, and growth-associated protein 43 immunoreactivity in injured spinal tissues compared with PBS-treated controls. Moreover, treatment with GDNF caused approximately 50% cell survival in the contused spinal cord tissues. Examination of signal transduction triggered by GDNF indicated that GDNF injection transiently induced activation of the mitogen activated protein (MAP) kinase pathway in the spinal cord. Additionally, an up regulation of anti-apoptotic Bcl-2 levels in the contusive center of the damaged spinal cord was observed 24 hr post-GDNF injection. Together our results show that GDNF exerts behavioral and anatomic neuroprotection following SCI. Additionally, GDNF-activated MAP kinase and Bcl-2 signaling may contribute to neuronal survival after spinal cord contusion. PMID- 12125082 TI - Fractalkine and fractalkine receptors in human neurons and glial cells. AB - Fractalkine has been identified as a novel chemokine that exhibits cell adhesion and chemoattractive properties in the central nervous system (CNS), and the fractalkine receptors, CX3CR1, are also expressed in the CNS. In the present study, the expression of fractalkine and fractalkine receptors was investigated in enriched populations of human CNS neurons, astrocytes, and microglia. In addition, the regulatory role played by protein kinase C (PKC) in fractalkine secretion in neurons was determined in A1 human hybrid neuronal cell line produced between a human cerebral neuron and a human neuroblastoma cell. Human neurons and astrocytes expressed fractalkine mRNA as determined by the revserse transcriptase-polymerase chain reaction (RT-PCR) analysis, while human microglia preparation did not express the fractalkine message. Human neurons and microglia expressed CX3CR1 mRNA, but astrocytes did not. These results suggest that fractalkine secreted by CNS neurons and astrocytes produce biological effects in neurons and microglia. Although phorbol ester did not change the expression of fractalkine mRNA level in A1 hybrid neurons, it did upregulate fractalkine secretion over unstimulated controls. This upregulation of fractalkine production was suppressed by the treatment with Ro32-0432, a PKC inhibitor. These results indicate that intracellular signals transduced by PKC play an important role in the regulation of soluble fractalkine at the post-transcriptional level in human neurons. As for the biological function of fractalkine, extracellularly applied fractalkine increased the number of bromodeoxyuridine-labeled microglia 3-fold over the untreated controls, indicating fractalkine induces proliferation of human microglia. These observations suggest that fractalkine released by injured neurons could induce proliferation, activation and/or migration of microglia at the injured brain sites. PMID- 12125081 TI - Molecular analysis of the vagal motoneuronal degeneration after right vagotomy. AB - The aim of this study was to investigate the vagal motoneuronal degeneration after right vagotomy using in situ hybridization, RT-PCR, and immunohistochemistry methods. The morphology of the vagal motoneurons in dorsal motor nucleus of the vagus nerve (DMV) and nucleus of ambiguus (NA) after right vagotomy was examined by using Nissl staing and TUNEL. The expression of inducible nitric oxide synthase (iNOS), bcl-2, bax, and caspase-3 in DMV and NA of rats after right vagotomy was studied. Additionally, the involvement of the N methyl-D-aspartate (NMDA) receptor-calcium-neuronal nitric oxide synthase (nNOS) pathway in the vagal motoneuronal degeneration was addressed by double immunolabeling analysis of nNOS with NMDAR1 and calbindin D28K in right vagotomized rats. The neurons in right DMV and NA displayed a darkly stained, shrunken morphology at 1 day and 5 days following right vagotomy as shown by Nissl staining. Quantitative analysis revealed that, at 1 day and 5 days following right vagotomy, the number of neurons in right DMV, but not NA, was significantly reduced in comparison with that of control rats. Occasional TUNEL positive neurons were detected in right DMV of rat at 1 day after right vagotomy. The expression of iNOS protein and mRNA was absent in DMV and NA of control rats. However, the iNOS mRNA expression was induced bilaterally in DMV and NA at 1 day postoperation and continued to be up-regulated until 5 days after vagotomy as shown by in situ hybridization. Immunohistochemistry analysis also showed the increased expression of iNOS in bilateral DMV and NA of vagotomized rats. RT-PCR analysis revealed the enhanced bcl-2 and reduced bax mRNA levels and subsequent up-regulation of both bcl-2 and bax mRNA in right sides of the vagotomized brainstems at 1 day and 5 days postoperation, respectively. In situ hybridization analysis confirmed the up-regulation of bcl-2 and bax mRNA in right DMV and NA of the rats at 5 days following operation. Immunohistochemistry analysis showed up regulated Bcl-2 immunoreactivity and undetectable changes in Bax immunoreactivity in DMV and NA of rats at 1 day after vagotomy, whereas enhancement of both Bcl-2 and Bax immunoreactivity was observed at 5 days postoperation. In addition, the caspase-3 mRNA level was elevated ipsilaterally in DMV and NA at 1 day and 5 days following right vagotomy. Double-immunofluorescence analysis showed complete colocalization of nNOS with NMDAR1 and with calbindin in ipsilateral DMV and NA at 10 days following right vagotomy. This study suggests that the signal pathway for NMDAR1-calcium-nNOS and the up-regulation of iNOS in DMV and NA may be involved in the vagal motor neurodgeneration after right vagotomy. Furthermore, our results imply that the apoptosis pathway mediated by Bcl-2, Bax, and caspase 3 may be activated in vagal motoneurons after right vagotomy. PMID- 12125083 TI - Development of quantitative exposure data for a pooled exposure-response analysis of 10 silica cohorts. AB - BACKGROUND: Comprehensive quantitative silica exposure estimates over time, measured in the same units across a number of cohorts, would make possible a pooled exposure-response analysis for lung cancer. Such an analysis would help clarify the continuing controversy regarding whether silica causes lung cancer. METHODS: Existing quantitative exposure data for 10 silica-exposed cohorts were retrieved from the original investigators. Occupation- and time-specific exposure estimates were either adopted/adapted or developed for each cohort, and converted to milligram per cubic meter (mg/m(3)) respirable crystalline silica. RESULTS: Quantitative exposure assignments were typically based on a large number (thousands) of raw measurements, or otherwise consisted of exposure estimates by experts (for two cohorts). Median exposure level of the cohorts ranged between 0.04 and 0.59 mg/m(3) respirable crystalline silica. Exposure estimates were partially validated via their successful prediction of silicosis in these cohorts. CONCLUSIONS: Existing data were successfully adopted or modified to create comparable quantitative exposure estimates over time for 10 silica-exposed cohorts, permitting a pooled exposure-response analysis. The difficulties encountered in deriving common exposure estimates across cohorts are discussed. PMID- 12125084 TI - Leukemia in relation to occupational exposures to benzene and other agents: a case-control study nested in a cohort of gas and electric utility workers. AB - BACKGROUND: Many occupational and environmental exposures have been implicated in the etiology of leukemia, but only a few, such as benzene, are well-established leukemogens. The risk of leukemia in a large cohort of gas and electricity utility workers with exposures to several suspected or confirmed carcinogens was investigated. METHODS: A case-control study nested within the cohort was conducted, with 72 leukemia cases identified among male workers, and 285 controls matched to the cases by year of birth. Only cases, and their matched controls, active in the company at the date of diagnosis were included. Exposure assessment was based on a job-exposure matrix (JEM) developed from expert judgment using a standardized procedure. RESULTS: The risk of leukemia was increased in workers with an estimated cumulative exposure to benzene > or = 16.8 ppm-years (OR = 3.6; 95% CI 1.1-11.7), and there was an indication of a dose-response relation (OR = 1.2; 95% CI 1.0-1.5 per 10 ppm-years increase in exposure). The link with benzene was more pronounced for acute leukemia than for chronic leukemia, but no association with a particular leukemia cell type was apparent. The risk of leukemia remained elevated for latency periods of 2, 5, or 10 years. CONCLUSIONS: From our evaluation, it could be estimated that the median TWA exposure to benzene among exposed workers was 0.16 ppm, i.e., within concentration ranges where an increased leukemia risk was usually not apparent in previous epidemiological studies. Although an increased leukemia risk may be real, it may also be related to other occupational factors not totally controlled for in the analysis, or to benzene exposures actually higher than expected. PMID- 12125085 TI - Association between bone lead concentration and blood pressure among young adults. AB - BACKGROUND: Occupational and environmental exposure to lead has been examined for its effect on blood pressure (BP) in adults with varying results. The present analyses assessed the association between bone lead concentration and BP in early adult life in persons exposed during childhood. METHODS: Study participants included young adult members of two cohorts with different past histories of lead exposure. Lead exposure was assessed using noninvasive K-X-ray fluorescence spectroscopy to quantify bone lead concentration, an index of long-term lead exposure superior to current blood lead concentration. Systolic and diastolic BP measurements were obtained using conventional clinical methods. Multiple linear regression models were constructed to allow for control of covariates of BP identified a priori. RESULTS: Analyses were performed on 508 participants. While controlling for potential confounders, systolic BP was 4.3 mm Hg greater among members of the highest of four bone lead concentration groups (> 10 microg Pb/g bone) when compared with the lowest bone lead concentration group (< 1 microg Pb/g bone; P = 0.004), and diastolic BP was 2.8 mm Hg greater among members of the highest bone lead concentration group when compared with the lowest bone lead concentration group (P = 0.03). CONCLUSIONS: These results suggest that substantial lead exposure during childhood can increase BP during young adulthood. PMID- 12125086 TI - Carpal tunnel syndrome among apprentice construction workers. AB - BACKGROUND: In terms of lost-work time and restricted workdays, surgery, and rehabilitation, one of the most costly occupational musculoskeletal disorders is carpal tunnel syndrome (CTS). The purpose of this study was to determine the prevalence of CTS among apprentice construction workers. METHODS: This cross sectional study included apprentices from four construction trades. Apprentices completed a self-administered questionnaire and received electrophysiologic studies assessing median nerve function across the carpal tunnel. A surveillance case definition for CTS was based on characteristic hand symptoms and the presence of median mononeuropathy across the carpal tunnel. RESULTS: Of the 1,325 eligible apprentices, 1,142 (86.2%) participated in the study. The prevalence of CTS among apprentices was 8.2%; sheet metal workers had the highest rate (9.2%). In operating engineers, the prevalence of CTS was significantly higher (OR = 6.9; 95% CI = 2.6-18.2) among the heavy equipment mechanics than the drivers of those vehicles. Body mass index, age, and self-reports of working overhead were associated with prevalent CTS. Less than 15% of the apprentices with CTS sought medical attention for their disorder. CONCLUSIONS: Many construction workers begin developing CTS before or during their apprenticeship. Few apprentices seek medical attention for hand symptoms characteristic of CTS. The results of this study indicate a public health need for the implementation of prevention strategies for CTS in the construction industry. PMID- 12125087 TI - Occupational risks and injuries in non-agricultural immigrant Latino workers. AB - BACKGROUND: To investigate occupational health in urban immigrant Latino workers, using a community-based method. METHODS: A survey was administered through consecutively selected door-to-door interviews. RESULTS: Response rate was 80% (n = 427). Average time in the US was 7.6 years, and average job tenure was 2.8 years. Twenty-five reported exposures to over 10 different hazards, and 18% thought these hazards had harmed their health. Only 31% received any job safety training; 55% had no workers' compensation coverage. Of the 47 (11%) with a work injury in the past 3 years, 27% reported difficulty obtaining treatment, 91% lost time from work (median = 13 days) and 29% had to change jobs because of the injury. The annual occupational injury rate was 12.2/100 full-time workers, compared to an expected rate of 7.1. CONCLUSIONS: Urban immigrant workers have increased risk of occupational injuries, with adverse outcomes. PMID- 12125088 TI - Injury risks in children of California migrant Hispanic farm worker families. AB - BACKGROUND: Few data are available addressing occupational and other injury risks among children of migrant Hispanic farm workers. METHODS: We conducted the U.C. Davis Farm Worker Injury Study (UCD-FWIS), a longitudinal follow-up study of injury among migrant Hispanic farm worker families living in six Northern California Migrant Housing Centers (MHCs). Nine hundred forty-one children (age < 18 years) were interviewed through parental proxy. RESULTS: Fifty-one injuries resulting in medical care or at least one-half day of lost or restricted work or school time occurred among 49 children (3.8 injuries/100 person-years). Open wounds (31.4%) and fractures (29.4%) were most common. Falls comprised over one third of the cases, followed by being struck and bicycle injuries. Over three quarters of subjects never use a helmet when riding a bicycle. Seventy-eight (8.3%) children reported employment in the preceding year, typically involving manual agricultural tasks. Two injury cases were occupational and involved agricultural work. CONCLUSIONS: Occupational injury was uncommon in this group of children in migrant Hispanic farm worker families. Injury prevention in this population should include a focus on the home and surrounding environment as well as the work place. PMID- 12125089 TI - Adolescent work patterns and work-related injury incidence in rural Minnesota. AB - BACKGROUND: Although there have been many studies on working youth in the United States, we have noted none which have provided a broad picture of adolescent work practices in a rural community. METHODS: Six high schools in rural Minnesota were evaluated for adolescent work practices. Schools ranged in size from 173 to 525 students in grades 9 through 12. A 20 page self-administered survey examining work practices was administered to students. RESULTS: A total of 2,250 students completed the survey, representing 92% of the student body. Twenty-eight percent of students lived on a farm. Approximately 45% of the male students and slightly more than 21% of the females were involved in farm work. Only 2.6% of students were injured during this 8-month time period in farm-related activities, and 5.1% were injured doing non-farm work. Many students reported working long hours. CONCLUSIONS: Work represents a serious problem for rural youth. These data are significant in the context of national policy discussion concerning the failure of the Fair Labor Standards Act to regulate the agricultural environment. PMID- 12125090 TI - Causes, nature, and outcomes of work-related injuries to adolescents working at farm and non-farm jobs in rural Minnesota. AB - BACKGROUND: Although there are many studies on working adolescents, information on youth who simultaneously hold jobs on both a farm and in other sectors of the economy is lacking. METHODS: Six high schools in rural Minnesota were evaluated for adolescent work practices and injury incidence. A 20-page self-administered survey was administered to students. RESULTS: A total of 2,250 students completed the survey, representing 92% of the student body. Students that simultaneously hold both farm and non-farm jobs have a significantly higher proportion of injuries. However, annual injury rates are highest for those working in non-farm only (26.7/100 full-time equivalents, FTEs) or farm only (25.9/100 FTEs) employment when compared with those working simultaneously in farm and non-farm jobs (21.9/100 FTEs). CONCLUSIONS: Many rural students employed simultaneously on farm and non-farm jobs work long hours and are at significant risk of work related injury. The annual injury rates we estimated are higher than those reported in previous studies. PMID- 12125092 TI - Asbestos screenings. PMID- 12125091 TI - The hypersensitivity pneumonitis diagnostic index: use of non-invasive testing to diagnose hypersensitivity pneumonitis in metalworkers. AB - BACKGROUND: Since 1993, several outbreaks of hypersensitivity pneumonitis (HP) have been reported in metalworkers. We report the largest outbreak of HP in metalworkers yet known. It occurred in a Connecticut factory that produces precision parts for the aerospace industry. The workers typically presented with systemic and respiratory problems ("sick fatigue," myalgias, cough, dyspnea, wheezing, and chest tightness). They had variable findings on clinical tests, which complicated diagnosis. An HP diagnostic index was developed to help reduce the uncertainties in case identification. METHODS: Data from 16 biopsy-confirmed cases and 14 non-HP patients were compared, and the HP diagnostic index was derived using variables that best discriminated between the two groups. The index is based on (a) work-related symptoms, (b) dry crackles on auscultation, (c) restrictive spirometry, (d) decreased diffusion capacity and/or increased A-a oxygen gradient, (e) elevated erythrocyte sedimentation rate, (f) abnormal radiographic images, and (g) abnormal gallium scans. We then applied the HP diagnostic index and, for comparison, the "Kenosha epidemiological case criteria" (developed during a recent HP outbreak in an automobile factory) to our data set. RESULTS: The HP diagnostic index and the Kenosha criteria confirmed HP in overlapping sets of 36 and 34 patients, respectively, that were both in good agreement with the clinical diagnoses. CONCLUSIONS: The HP diagnostic index relies less heavily on symptoms, subjective evaluations, and invasive tests than the Kenosha criteria, but both identified similar subsets of the 61 patients as having HP. The HP diagnostic index could provide a useful tool in future HP outbreaks, which are increasingly being recognized in metalworking facilities. PMID- 12125093 TI - Sub-lethal concentrations of activated complement increase rat lymphocyte glutamine utilization and oxidation while lethal concentrations cause death by a mechanism involving ATP depletion. AB - Nucleated cells are more resistant to complement-mediated cell death than anucleated cells such as erythrocytes. There are few reports concerning the metabolic response of nucleated cells subjected to sub-lethal complement attack. It is possible that the rate of utilization of specific metabolic fuels by the cell is increased to enhance cell defence. We have measured the maximum activity of hexokinase, citrate synthase, glucose 6-phosphate dehydrogenase and glutaminase in rat mesenteric lymphocytes exposed to sub-lethal concentrations of activated complement (present in zymosan-activated serum, ZAS). These enzymes were carefully selected as they indicate changes of flux in glycolysis, TCA cycle, pentose phosphate pathway and glutaminolysis, respectively. The only enzyme activity to change on exposure of lymphocytes to ZAS was glutaminase, which was enhanced approximately by two-fold. Although rates of both glutamine and glucose utilization were enhanced by exposure to ZAS, only the rate of oxidation of glutamine was increased. Complement kills anucleated cells by simple osmotic lysis. However, it is likely that some nucleated cells will display characteristics of an ordered death mechanism and we have demonstrated that the concentration of lymphocyte ATP is dramatically decreased by activated complement. Nevertheless, the extent of cell death could be significantly reduced by the addition of inhibitors of the nuclear enzyme poly (ADP-ribose) polymerase (PARP). We conclude that glutamine metabolism is not only important for lymphocyte proliferative responses but is also important for cell defence from sub-lethal concentrations of activated complement. The rapid rate of complement induced lymphocyte death reported here is suggested to be a consequence of over activation of the nuclear enzyme PARP and ATP depletion. PMID- 12125094 TI - Cytokine detection in HIV-1/HHV-8 co-infected subjects. AB - In a previous work we have evaluated some immunologic and haematologic parameters of HIV-1 positive subjects co-infected with HHV-8. A worsening of these values were generally described in these patients as compared with those HIV-1 positive, but negative for HHV-8. Now we have studied the influence of HHV-8 co-infection of HIV-1 positive subjects on the production of some cytokines to make clear the question of its role in the immuno-deregulation of the above-mentioned subjects. In particular we have analysed serum levels of IL-4 and IL-10, Th2 type T cells cytokines, IFN-gamma, an indirect marker of Th1 cells activation and IL-18, a cytokine produced by monocytic-macrophagic cells, which is able to induce IFN gamma production and Th1 T lymphocytes activation. No significant differences were found as regards the IFN-gamma serum levels (92.1 +/- 24.3 pg ml(-1) in the case of HIV-1 positive/HHV-8 negative subjects and 96.0 +/- 17.4 pg ml(-1) in those HIV-1 positive/HHV-8 positive). In healthy subjects the mean level of this cytokine was 17.6 +/- 5.2 pg ml(-1) (significant difference with both the former values at p < 0.001). Moreover IL-4 and IL-10, which were undetectable in healthy individuals, showed the following values in HIV-1 positive/HHV-8 negative subjects: 31.9 +/- 2.7 pg ml(-1) and 119.8 +/- 85.1 pg ml(-1) respectively and in HIV-1 positive/HHV-8 positive subjects: 30.4 +/- 4.8 pg ml(-1) and 69.4 +/- 65.3 pg ml(-1) (not significant differences). In contrast IL-18 reached a mean level of 1001.2 +/- 360.5 pg ml(-1) in HIV-1 positive/HHV-8 negative subjects, but showed a significant reduction in HIV-1 positive/HHV-8 positive subjects (737.6 +/- 284.3 pg ml(-1) --> p < 0.05) and presented very low levels in healthy individuals (21.3 +/- 30.3 pg ml(-1)). Moreover a significant correlation (-0.984 --> p < 0.001) was noticed between IL-18 reduction in HIV-1 positive subjects co infected with HHV-8 and the degree of positivity of HHV-8. These data suggest that HHV-8 co-infection has no influence on the switch Th1 --> Th2 in HIV-1 positive subjects, but is able to reduce IL-18 production, useful for Th1 subset restoration. PMID- 12125095 TI - 1alpha,25-Dihydroxyvitamin D(3) inhibits rat liver ultrastructural changes and the development of gamma-glutamyltranspeptidase-positive foci in diethylnitrosamine-initiated and streptozotocin-induced diabetes-promoted hepatocarcinogenesis. AB - In the present study, the chemopreventive effect of the active metabolite of vitamin D, 1alpha,25-dihydroxyvitamin D(3) (VD(3)), against chemically-induced and diabetes-promoted rat liver carcinogenesis was investigated. Hepatocarcinogenesis was initiated with a single intraperitoneal (i.p.) injection of diethylnitrosamine (DEN) (125 mg kg(-1) body weight) at week 4 followed by promotion with streptozotocin (STZ) (65 mg kg(-1) body weight with a single i.p. injection) at week 7. With this basic experimental regimen, the effect of VD(3) (0.3 microg (0.1 ml)(-1) propylene glycol per os twice a week) was investigated with effect from 4 weeks prior to the exposure of DEN. The results showed that VD(3) supplementation throughout the experimental period reduced the incidence, total number and multiplicity and altered the size of visible persistent nodules (PNs) in DEN- or DEN + STZ-treated rats as compared with their respective controls. In these two groups, it also caused a significant decrease in the number (p < 0.002 and 0.001 respectively) and focal area (p < 0.05) of gamma glutamyltranspeptidase (GGT)-positive hepatic foci. Moreover, continuous supplementation of VD(3) exhibits a protective effect in maintaining the normal cellular architecture of the hepatocytes in DEN- or DEN + STZ-treated rats. Our results thus strongly suggest that VD(3) is very effective in the inhibition of DEN-initiated and STZ-induced diabetes-promoted rat liver carcinogenesis. PMID- 12125096 TI - Azaanthraquinone inhibits respiration and in vitro growth of long slender bloodstream forms of Trypanosoma congolense. AB - An ethanolic extract of Mitracarpus scaber was found to possess in vitro and in vivo trypanocidal activity against Trypanosoma congolense. At a dosage of 50 mg kg(-1) day(-1) in normal saline for 5 days, the extract cured Balbc mice infected with T. congolense without any relapse. The isolated active component benz(g)isoquinoline 5,10 dione (Azaanthraquinone) (AQ) purified from the extract was found to inhibit glucose-dependent cellular respiration and glycerol-3 phosphate-dependent mitochondrial O(2) assimilation of the long bloodstream forms of Trypanosoma congolense. On account of the pattern of inhibition, the target could be the mitochondrial electron transport system composed of glyceraldehyde 3 phosphate dehydrogenase (G3PDH). The azaanthraquinone specifically inhibited the reduced coenzyme Q(1)-dependent O(2) uptake of the mitochondria with respect to ubiquinone. The susceptible site could be due to ubiquinone redox system which links the two enzyme activities. PMID- 12125097 TI - The difference in the stimulation by putrescine of DNA synthesis using DNA polymerase extracts of normal rat liver or of tumour tissue or host liver from tumour-bearing rats. AB - Putrescine biosynthesis is elevated before DNA replication, and a stimulation of DNA synthesis by 20 mM putrescine has been found using an in vitro DNA synthesizing system. Furthermore, this stimulation of DNA synthesis by putrescine involves a particular factor (factor PA). This factor PA stimulates DNA polymerases alpha, beta, and gamma, and is present in nuclei and mitochondria but not in cytoplasm. Factor PA loses about 80% of its activity by heating at 45 degrees C for 15 min or by hydrolysis with 100 mg ml(-1) Enzygel trypsin. These properties indicate that factor PA is a protein. Its size is estimated to be about 2.1 S. DNA synthesis in nuclear and mitochondrial DNA polymerase extracts from tumour tissues and host livers of tumour-bearing rats are not stimulated by 20 mM putrescine. However, the addition of excess factor PA to DNA synthesizing systems using DNA polymerase extracts from proliferative tissues again results in a stimulation of DNA synthesis by exogenous putrescine. These findings indicate that the stimulatory effect of DNA synthesis in vitro by exogenous putrescine is controlled by the ratio between factor PA and endogenously synthesized putrescine in proliferative tissues or that sent by the bloodstream from proliferative tissues. These results suggest that a non-stimulatory effect of putrescine on DNA synthesis may be diagnostic in tumour-bearing patients. PMID- 12125098 TI - Glyceraldehyde metabolism in human erythrocytes in comparison with that of glucose and dihydroxyacetone. AB - Metabolism of D-glyceraldehyde in human erythrocytes in comparison with that of glucose and dihydroxyacetone was studied. Both trioses were metabolized to produce L-lactate at rates comparable to that of L-lactate formation from glucose. Almost complete inactivation of glyceraldehyde-3-phosphate dehydrogenase by treatment of cells with iodoacetate resulted in a 95% decrease in L-lactate formation from the ketotriose as well as from glucose, whereas L-lactate formation from the aldotriose was only partially reduced (60%). D-Lactate was produced faster from either the aldotriose or the ketotriose than from glucose, but the ability of the two trioses to produce D-lactate was far lower than that to produce L-lactate. Almost complete inhibition of aldehyde dehydrogenase by disulfiram and of both aldose reductase and aldehyde reductase II by sorbinil, had no effect on L-lactate formation from D-glyceraldehyde. The present study suggests that D-glyceraldehyde is metabolized via two or more pathways including the glycolytic pathway after its phosphorylation by triokinase, and that neither oxidation to D-glyceric acid nor reduction to glycerol is a prerequisite for D glyceraldehyde metabolism. PMID- 12125099 TI - Transcriptional activation of the human insulin receptor gene by 1,25 dihydroxyvitamin D(3). AB - Treatment with 10(-8) M 1,25-dihydroxyvitamin D(3) for 24 h causes transcriptional activation of the human insulin receptor gene in U-937 human promonocytic cells. The activation seems to potentiate the response to insulin in terms of glucose oxidation. Wortmannin, a phosphatidylinositol 3-kinase inhibitor, causes a greater inhibition of insulin-stimulated glucose oxidation in 1,25-dihydroxyvitamin D(3)-treated cells than in untreated cells. This suggests a stimulation of phosphatidylinositol 3-kinase activity by 1,25-dihydroxyvitamin D(3), which could mediate, at least in part, the potentiation of the insulin response. PMID- 12125100 TI - Cervico-vaginal dysplasia--papillomavirus-induced and HIV-1 infection: role of correlated markers for prognostic evaluation. AB - Sexually-transmitted diseases (STD) can facilitate the progression of HIV-1 infection. Among them, as we have previously demonstrated, cervico-vaginal dysplasia-papillomavirus (HPV)-induced, together with HSV-2 co-infection, seems to be correlated with a more evident immunodepression in HIV-positive women, compared with other sexually transmitted diseases. Here we have analysed some of the main correlated markers of HIV-1 infection progression: CD4 + T lymphocyte concentration, CD4 +/CD8 + T cells ratio, HIV-1 RNA loads and haemoglobin (Hb) concentration in 30 HIV-1 positive women co-infected with HPV, and suffering from cervico-vaginal dysplasia, in different stages. In particular, we noticed a positive correlation, evaluated by Spearman's test, between the degree of progression of dysplastic stages (CIN1 --> 3) until invasive carcinoma (IC) and HIV-1 RNA loads (C(s) = +0.78; p < 0.001), and in contrast, a negative correlation between the same stages of progression and respectively CD4 + T cell concentration (C(s) = -0.54; p = 0.01), ratio (C(s) = - 0.63; p = 0.002) and Hb concentration (C(s) = -0.85; p < 0.001). In conclusion, it is important to underline that low levels of Hb generally paralleled the degree of immunodepression. In fact CD4 + T cell levels and ratio positively correlated with Hb concentrations respectively, with C(s) = + 0.83; p < 0.001 and C(s) = + 0.90; p < 0.001. Finally, the most efficacious antiretroviral combined therapy (HAART = Highly Active Antiretroviral Therapy) can improve the above described laboratory parameters in HIV-1/HPV co-infected women and seems to prevent the progression of CIN1 to the following stages of the dysplastic disease. PMID- 12125101 TI - Aldosterone impairs insulin responsiveness in U-937 human promonocytic cells via the downregulation of its own receptor. AB - In an earlier study, we have reported an inhibition of insulin receptor (IR) mRNA levels and insulin binding by aldosterone in U-937 human promonocytic cells. In the present extension of our studies, we demonstrate that this inhibition by aldosterone had no effects on basal glucose transport or on basal thymidine incorporation into DNA, while the cell responsiveness reflected by the maximal response to insulin was decreased by 23% for glucose transport and by 31% for DNA synthesis after the aldosterone treatment. We also prove that this inhibition of the insulin response by aldosterone is mediated by a downregulation of the levels of mineralocorticoid receptors (MRs) (50% decrease) and their mRNA (50% decrease). In addition, the mineralocorticoid antagonist spironolactone reversed the decrease in MR mRNA levels elicited by aldosterone, which suggests the involvement of this receptor in the process. PMID- 12125102 TI - Cross-reactivity of some antibodies to human epitopes with shrimp Pandalus borealis proteins: a possible aid in validation and characterization of crustacean cells in vitro. AB - Cell characterization of primary cultures in vertebrates is well established but not in marine invertebrates. This fact is hampering advances in the development of tissue cultures from this species. In the present study, a panel of antibodies to structural proteins, stress proteins, oncogenes and proliferation antigens, developed against mammalian antigens, were tested in paraffin sections of the crustacean Pandalus borealis tissues. Several tissues were analysed: hepatopancreas, gills, ovaries, epithelium under the cuticle and abdominal muscle. Specific antibodies to crustacean proteins are not commercially available. The immunocytochemical results show that antibodies to human epitopes cross-react with antigens in the crustacean Pandalus borealis indicating that some cellular proteins are highly conserved in evolution. Cytokeratin, proliferating cell nuclear antigen, ras and p-glycoprotein were detected by immunocytochemistry in Pandalus borealis. No immunoreactivity for Ki-67 and metallothionein was observed. This system can help in validation and characterization of invertebrate cultures. PMID- 12125103 TI - A 45-kDa midgut glycoprotein from Anopheles albimanus mosquito mediates the killing of trypanosomes. AB - Trypanosomes do not inhabit or grow in anopheles mosquitoes, the vector for the transmission of Plasmodium parasites the causative agent for malaria. The possession of lytic factors by the anopheline mosquito was thus considered. Head and midgut sections prepared in phosphate buffered saline were tested for trypanocidal action against T. congolense. While the head section was inactive towards the trypanosomes, the midgut extract at 0.2 mg ml(-1) diminished the motility of the parasites within 2 min of incubation; killing 50% of the population after 5 min. At 0.5 mg ml(-1) of the extract, about 90% of the parasites were killed within 2 min of incubation. The midgut fraction was subjected to a purification protocol involving successive chromatography on: octyl-sepharose, reactive brown agarose and fetuin-agarose columns. A final trypanocidal active fraction (gp45), which moved homogeneously during electrophoresis as a 45-kDa protein, was recovered from the fetuin-agarose column. The protein reacted positively with thiobarbituric acid, which suggests it is a sialoglycoprotein. Desialylation of the glycoprotein nullified its trypanocidal activity on T. congolense. Similarly, when the saccharides, lactose, methyl-beta-galactoside, lactulose, methyl-umbelliferyl-beta-galactoside (MU Gal), were included in the culture medium, they inhibited the gp45 trypanocidal activity. Asialo-fetuin and asialo-RBC also inhibited the gp45-induced killing of T. congolense cells. The potential use of anopheline 45 kDa protein in the production of transgenic tsetse flies (Glossina spp.) in the control of trypanosomiasis is discussed. PMID- 12125104 TI - Apoptosis induction by S-allylcysteine, a garlic constituent, during 7,12 dimethylbenz[a]anthracene-induced hamster buccal pouch carcinogenesis. AB - The apoptosis-inducing capacity of S-allylcysteine (SAC), a water-soluble garlic constituent, during 7,12-dimethylbenz[a]anthracene-induced hamster buccal pouch (HBP) carcinogenesis was investigated in male Syrian hamsters using DNA fragmentation and the apoptosis-associated proteins, tissue transglutaminase (tTG) and Bcl-2. Hamsters were divided into four groups of six animals each. Animals in group 1 were painted with a 0.5% solution of DMBA in liquid paraffin on the right buccal pouches three times a week for 14 weeks. Group 2 animals painted with DMBA as in group 1, in addition received 200 mg kg(-1) body weight SAC orally on days alternate to DMBA application. Group 3 animals received SAC as in group 2. Group 4 animals received neither DMBA nor SAC and served as the control. The experiment was terminated at the end of 14 weeks. Administration of SAC (200 mg kg(-1) body weight) to animals painted with DMBA inhibited DMBA induced HBP carcinogenesis as revealed by the absence of neoplasms, induction of tTG and inhibition of Bcl-2 expression. The results of the present study suggest that SAC may exert its chemopreventive effect by inducing apoptosis. PMID- 12125105 TI - Melatonin in the unicellular Tetrahymena pyriformis: effects of different lighting conditions. AB - Melatonin content in the cellular fraction and medium of Tetrahymena pyriformis GL cultures was measured at different time points of light and dark exposures. Tetrahymena produced, stored and secreted immunoreactive melatonin, which in displacement and HPLC studies, behaved like synthetic melatonin. There was not a continuous secretion of melatonin produced by the cells. In contrast to this, storage of melatonin was observed, which was more expressed in dark conditions. Prolonged light exposure suppressed melatonin production and secretion alike, however it did not block it completely. PMID- 12125106 TI - Genotyping of Plasmodium spp. Nested PCR. PMID- 12125107 TI - Genotyping of Plasmodium falciparum. PCR-RFLP analysis. PMID- 12125108 TI - Epidemiological measures of risk of malaria. PMID- 12125110 TI - Quantitation of liver-stage parasites by automated TaqMan real-time PCR. PMID- 12125109 TI - Microsatellite analysis in Plasmodium falciparum. PMID- 12125111 TI - Quantitation of liver-stage parasites by competitive RT-PCR. PMID- 12125112 TI - Extraction and purification of Plasmodium total RNA. PMID- 12125113 TI - Extraction and purification of Plasmodium parasite DNA. PMID- 12125114 TI - Southern blotting of parasite DNA. PMID- 12125115 TI - SDS-PAGE and western blotting of Plasmodium falciparum proteins. PMID- 12125116 TI - Nested PCR analysis of Plasmodium parasites. PMID- 12125117 TI - RFLP analysis. PMID- 12125118 TI - Analysis of gene expression by RT-PCR. PMID- 12125119 TI - In situ detection of RNA in blood- and mosquito-stage malaria parasites. PMID- 12125120 TI - Purification of chromosomes from Plasmodium falciparum. PMID- 12125121 TI - Construction of genomic libraries from the DNA of Plasmodium species. PMID- 12125122 TI - Maintenance of the Plasmodium berghei life cycle. PMID- 12125123 TI - Construction of a gene library with mung bean nuclease-treated genomic DNA. PMID- 12125124 TI - Construction of Plasmodium falciparum lambda cDNA libraries. PMID- 12125125 TI - Production of stage-specific Plasmodium falciparum cDNA libraries using subtractive hybridization. PMID- 12125126 TI - Construction and screening of YAC libraries. PMID- 12125127 TI - Vector incrimination and entomological inoculation rates. PMID- 12125128 TI - Episomal transformation of Plasmodium berghei. PMID- 12125129 TI - Gene targeting in Plasmodium berghei. PMID- 12125130 TI - Peptide vaccination. PMID- 12125131 TI - DNA vaccination. PMID- 12125132 TI - Assessing antigen-specific CD8+ and CD4+ T-cell responses in mice after immunization with recombinant viruses. PMID- 12125133 TI - Assessing CD4+ helper T-lymphocyte responses by lymphoproliferation. PMID- 12125134 TI - Limiting dilution analysis of antigen-specific CD4+ T-cell responses in mice. PMID- 12125135 TI - Cell trafficking. Malaria blood-stage parasite-specific CD4+ T cells after adoptive transfer into mice. PMID- 12125136 TI - Assessing antigen-specific proliferation and cytokine responses using flow cytometry. PMID- 12125137 TI - Mouse models for pre-erythrocytic-stage malaria. PMID- 12125138 TI - Cytokine analysis by intracellular staining. PMID- 12125139 TI - Assessment of antigen-specific CTL- and CD8(+)-dependent IFN-gamma responses in mice. PMID- 12125141 TI - Human antibody subclass ELISA. PMID- 12125140 TI - Assessing antigen-specific CD8+ CTL responses in humans. PMID- 12125142 TI - Systemic nitric oxide production in human malaria. I. Analysis of NO metabolites in biological fluids. PMID- 12125143 TI - Systemic nitric oxide production in human malaria. II. Analysis of mononuclear cell nitric oxide synthase type 2 antigen expression. PMID- 12125144 TI - In vitro culture of Plasmodium parasites. PMID- 12125145 TI - Automated synchronization of Plasmodium falciparum parasites by culture in a temperature-cycling incubator. PMID- 12125146 TI - Hepatic portal branch inoculation. PMID- 12125147 TI - Hepatocyte perfusion, isolation, and culture. PMID- 12125148 TI - Inhibition of sporozoite invasion. The double-staining assay. PMID- 12125149 TI - Inhibition of liver-stage development assay. PMID- 12125150 TI - T-cell mediated inhibition of liver-stage development assay. PMID- 12125151 TI - Antibody-dependent cellular inhibition assay. PMID- 12125152 TI - Erythrocytic malaria growth or invasion inhibition assays with emphasis on suspension culture GIA. AB - Erythrocytic cycle malaria parasite growth or invasion inhibition assays (GIA) compare the effects of various test and control substances on malaria parasite growth in erythrocytes or invasion into erythrocytes in vitro. Although inhibitions by antimalarial drugs in vitro correlate well with drug protective levels required in vivo, as yet there are too few data to know how well inhibitions by antibodies in vitro correlate with the types and degrees of immune protection in vivo. Antibody-mediated GIA is frequently complicated by parasite strain-specific inhibitions, as well as nonspecific inhibitory factors generated in sera collected or stored under nonoptimal conditions. In this chapter, we describe methods for collecting and processing sera, for using different strains of parasite, and a simplified method for staining parasite DNA with Hoechst dye 33342 before quantitating parasites using ultraviolet (UV)-excited flow cytometry. We also describe a new type of GIA using suspension cultures in a 48 well plate. Critical to this method is enclosing the plate in a gassed, heat sealed plastic bag, which, being low mass, can easily be rested at a 13.5 degrees angle on a rotor platform (114 rpm with 1-in. displacement) to produce gentle pulsatile waves of media in each well. The suspension GIA, which, relative to the static GIA, increased inhibition by one antibody and decreased inhibition by another (Table 1), may better simulate in vivo blood flow and may thus better predict in vivo efficacy. PMID- 12125153 TI - Analysis of CSA-binding parasites and antiadhesion antibodies. PMID- 12125154 TI - Analysis of the adhesive properties of Plasmodium falciparum-infected red blood cells under conditions of flow. PMID- 12125155 TI - Mouse models for erythrocytic-stage malaria. PMID- 12125156 TI - Preparation of adhesive targets for flow-based cytoadhesion assays. PMID- 12125157 TI - Triton X-114 phase partitioning for antigen characterization. PMID- 12125158 TI - Immunoprecipitation for antigen localization. PMID- 12125159 TI - Field trials. PMID- 12125160 TI - Nonhuman primate models. I. Nonhuman primate host-parasite combinations. PMID- 12125161 TI - Nonhuman primate models. II. Infection of Saimiri and Aotus monkeys with Plasmodium vivax. PMID- 12125162 TI - Vector analysis. PMID- 12125163 TI - [The role of virus infections in bovine mastitis]. AB - Mastitis is an often occurring disease in dairy cattle with an enormous economic impact for milk producers worldwide. Despite intensive research, which is historically based on the detection of bacterial udder pathogens, still around 20 35% of clinical cases of bovine mastitis have an unknown aetiology. Due to the high number of unknown causes of clinical mastitis, studies were undertaken to gain more insight into the role of viruses in this important disease. This review deals with the role of viruses in the aetiology of bovine mastitis, including the results of the recently performed study on the role of bovine herpesvirus 4 (BHV4) in this aetiology. We conclude that viral infections can play a direct or indirect role in the aetiology of bovine mastitis; therefore their importance and their economic impact needs further attention. PMID- 12125164 TI - [Botulism poisoning in cattle, a case report, diagnosis and prevention]. AB - On a dairy farm 22 animals die in 14 days. After 10 days the clinical diagnosis is confirmed: clostridium botulinum type D intoxication. The clinical, diagnosis, therapy and prevention are discussed. PMID- 12125165 TI - [Actinobacillosis, an old friend found in an unusual place]. PMID- 12125166 TI - [Serologic test in rabbits with Encephalitozoon cuniculi infection]. PMID- 12125167 TI - ['Madam, collies don't have hip dysplasia']. PMID- 12125168 TI - [Pure water, a necessity for life, but not such an abundant phenomenon]. PMID- 12125169 TI - [From veterinarian student to university student]. PMID- 12125170 TI - [About type and incidence of heart disease in psittacines kept in captivity in Germany]. AB - It was the aim of the study to get an overview about spectrum and incidence of pathological aberrations in hearts and proximal vessels of psittacines which are kept in captivity in Germany. 107 psittacines which had been sent for routine necroscopy were examined especially for evidence of pathological alterations of the heart and/or the proximal vessels. In 36.4% of these birds macroscopic alterations could be verified. 99.0% of all hearts examined resp. proximal vessels showed at least mild histological alterations. This study allocates that especially noninfectious heart disease is of markedly importance in psittacines and that their evidence is much higher than it might be suspected on the basis of publications. PMID- 12125171 TI - [Basic ethological considerations concerning the evaluation of the welfare of farm animals]. AB - In the assessment of husbandry conditions with regard to farm animal welfare the probability or risk is evaluated to which extent the animals are feeling well or are prone to pain, suffering or physical harm under the specific husbandry conditions. It is emphasised that well-being is more than merely the absence of pain, suffering and physical harm. Well-being is defined as the experience of the extent of being able to successfully cope with the environment. Consequently, any prevention to actively and successfully interact with the environment may impair animal well-being. This situation often arises because of conflicts between husbandry conditions and innate species-specific behaviour programs, regardless of any domestication effects on the reactivity of the farm animals to their environment. Based on these presumptions, four broad categories of influence on the well-being of animals are identified and exemplary explained. On the side of the environment these are the extent of (1) physical opportunity to perform species-specific behaviour, (2) availability of adequate stimuli and substrates for this behaviour and (3) adequate learning opportunities, especially during rearing. On the animals' side it is the extent of (4) their genetically based bodily capacity to perform species-specific behaviour. Less behavioural restriction is associated with the likelihood that better well-being is safeguarded under the aspect of behaviour. For a full assessment with respect to animal welfare also health aspects and other variables as appropriate must be taken into account. The assessment is comparative by nature and does not in itself include any conclusion about the acceptability of certain conditions. PMID- 12125172 TI - Influence of diet on the occurrence of some bacteria in the intestinal flora of wild and pet birds. AB - Enterobacteriaceae, E. coli in particular, do not belong to the intestinal flora of granivorous pet birds. This is indicated by the fact that the feces of only 9% of healthy budgerigars and 17% of finches tested were positive for enterobacteria. Stressful situations such as overcrowding in small cages coincident with increased noise and low light levels can enhance the colonization of the gut with E. coli. On the other hand it seems nearly impossible to colonize the intestine of budgerigars with E. coli or Klebsiella spp. even under favourable conditions. The frequent findings of enterobacteria in deceased granivorous birds suggest that E. coli and other enterobacteria are involved in the course of diseases with predisposing factors. Nutritional experiments with young chickens suggest that a diet consisting exclusively of seeds has an inhibitory effect on intestinal colonization with E. coli. Determination of Aeromonas hydrophila in nearly 3500 wild and pet birds provides statistically significant evidence that the composition of the intestinal flora may depend on dietary habits: infection was found in 1.9% of the granivorous and herbivorous species, in 7.1% of the omnivorous and in 12.4% of the carnivorous and insectivorous birds. The occurrence of enterobacteria and Aeromonas hydrophila in the digestive tract is obviously influenced by the composition of the nutrients. PMID- 12125173 TI - The effect of a homologous bacterin given to sows prefarrowing on the development of Glasser's disease in postweaning pigs after i.v. challenge with Haemophilus parasuis serotype 5. AB - The trial was carried out to evaluate the impact of maternal antibodies on the development of Glasser's disease after i.v. exposure of weaned pigs with a homologous serovar of Haemophilus parasuis (HPS). Two groups of weaned pigs were formed. Group one VI (n = 10): born to vaccinated sows, weaners i.v. challenged one week postweaning and euthanatized 14 days postweaning. Group two NVI (n = 10 wearners): born to non vaccinated sows, i.v. challenged one week postweaning euthanatized 14 days postweaning. One week postweaning all weaners were i.v. inoculated with HPS serovar 5. The following parameters were evaluated: clinical signs (depression, centralnervous signs, fever, lameness), macroscopic lung, pleura, peritoneum, liver and joint changes, and mortality. All trial sows were HPS seronegative prior to vaccination. The HPS vaccinated sows were proven seropositive on day 3 p.p. (values > 0.24), the non vaccinated ones were tested seronegative (values < 0.23). The progeny of sows vaccinated prefarrowing with two doses of HPS serovar 5 bacterin were partially protected against HPS caused clinical and pathological signs. The majority of clinical signs as fever, depression, recumbency, lack of response to verbal stimuli and lameness showed significant (P < 0.05) milder clinical symptoms in VI than in NVI animals. Respiratory signs (P = .169) and involvement of the central nervous system as ataxia, muscular tremor, incoordination of hind legs and convulsions (P = 1) showed no significant differences between the groups. Except lesions of pericard (VI vs. NVI, P = .14) and pleura (VI vs. NVI, P = .14) there were significant (P < 0.05) macroscopic differences at necropsy in lung, liver, joints and cerebrospinal fluid between the offspring of vaccinated sows and the ones of non vaccinated dams. No HPS were isolated from the nasal mucosa of the pigs prior to inoculation. HPS serovar 5 was recovered at necropsy from the nasal mucosa of all pigs in both groups. One pig from group VI presented in all examined organs the presence of HPS serovar 5. The remaining animals in group VI revealed in lung, pericard, pleura, liver, joints and cerebrospinal fluid no presence of HPS. The rate of isolation between VI and NVI groups revealed a significant (P < 0.05) difference. All the survived piglets of group NVI showed positive ELISA titres against HPS serovar 5 (values > .24). The piglets that died or were euthanatized before the end of the study have not been subjected to ELISA serological testing. One piglet died in group VI before the end of the study. Non of the remaining animals in group VI showed seroconversion to HPS serovar 5. IMPLICATIONS: Vaccination of sows did not influence the colonisation of nasal mucosa, but progeny of sows vaccinated prefarrowing with two doses of HPS serovar 5 bacterin were partially protected against HPS caused diseases. PMID- 12125174 TI - [Effects of housing conditions of farrowing and nursing sows on development of piglets: our own studies and an evaluation of the literature]. AB - Results of rearing and growing 3250 piglets derived from 287 litters of 99 German Landrace sows in parities 1-11, from parturition throughout 5 weeks post-weaning, were compared in two farrowing systems. The Group-Farrowing System (GFS) was designed according to natural behaviour of wild pigs and contained 8 farrowing boxes allowing sows and their litters to move freely on an area of 85.2 m2. In contrast, the Single Farrowing System (SFS) was a pen in which sows and their litters were housed individually and could move freely on a restricted area of 6.7 m2. Sows were only fixed during farrowing. The first experiment with a suckling period of 35 days comprised 98 litters from 60 sows (parities 1-4). The second experiment with a suckling period of 28 days involved 189 litters of 70 sows (parities 1-11). In experiment 1, body weight of GFS-piglets was significantly higher by 4.57 kg (19%) at the age of 70 d compared with SFS piglets. In the second experiment, the proportion of stillborns was significantly lower by 50% in GFS than in SFS. Furthermore, body weight of GFS-piglets showed a greater homogeneity during the first 35 d of life than that of SFS-piglets. In SFS, a positive correlation between parity of the sow and stillborn rate was observed. Behavioural studies in both experiments revealed that GFS-piglets, reared during the suckling period in close contact to other sows and piglets from other litters, were in a better condition at weaning than SFS-piglets. Comparisons with other GFS-types show that the success of farrowing in group housing is dependent on the type of housing conditions, in particular on the possibility for sows and piglets to live according to their specific behavioural repertoire. The factors critical in this context are discussed. PMID- 12125175 TI - [Requirements for quality of approved plant medicines]. AB - Herbal medicinal products differ substantially from drugs with synthetic active ingredients. Whereas the active ingredients of synthetic drugs are chemically well defined and pure substances, those of phytopharmaceuticals are plants, parts of plants or extracts. Therefore starting material of high quality and a standardised manufacturing procedure are particularly important for phytopharmaceuticals. Thus the documentation of the quality of a herbal medicinal product is crucial. The quality criteria aim to guarantee a phytopharmaceutical of high and constant quality and as well as a successful and reproducible therapy. The new Swiss Federal Law on Medicinal Products and Medical Devices (Law on Therapeutic Products--LTP) came into effect on 1 January 2002. The Intercantonal Office for the Control of Medicines IOCM which previously was in charge of herbal medicinal products was closed down and together with the Therapeutic Products Section of the Swiss Federal Office of Public Health SFOPH merged to create Swissmedic, the Swiss Agency for Therapeutic Products. Swissmedic is now responsible for affairs related to herbal medicinal products. The following article gives an overview of the current quality requirements of phytopharmaceuticals. These are now based on the LTP, but the essence remains largely unchanged. PMID- 12125176 TI - [From medicinal plant to phytotherapeutic drug]. AB - The effects of phytotherapy are generally attributed to a specific plant as a whole. However, plants only provide the raw material from which the active ingredients are collected by special procedures. Most of the herbal medicinal preparations as the active ingredients in herbal medicinal products are obtained by extraction using water or organic solvents such as ethanol or methanol. This yields a large variety of preparations with diverse properties. Particular to phytotherapeutic preparations in contrast to synthesized drugs is that they contain a complex mixture of active principles that is difficult to characterize chemically and biologically. Nevertheless, the quality of medicinal preparations from plants can be guaranteed if the raw material, the mode of preparation and the end product are well defined by the manufacturer. PMID- 12125177 TI - [Adverse effects and interactions of phytotherapeutic drugs]. AB - The significantly increased sales figures many phytopharmaceuticals have achieved during the last years prove the confidence that a great part of the population has in herbal remedies. This is primarily due to the wide-spread opinion that herbal remedies are free from side-effects. The long tradition and presumed 'natural' origin are no guarantee for safety in the treatment with herbal remedies. Even if a large proportion of the undesirable events is traceable to falsifications, impurities and lacking quality controls, herbal drugs with controlled quality should not be generally classified as harmless. In the meantime it has been possible to prove the presence of active substances with toxic and cancerogenic properties in various phytopharmaceuticals. Interactions with other drugs have been documented in a number of notes, where phytopharmaceuticals could influence the blood plasma level of various drugs, presumably by activating or inhibiting the cytochrom-P450-system. At present, especially data about adverse effects during long-term administration of herbal remedies are under-represented. Particularly because of their presumed harmlessness they often are prescribed in the case of chronic diseases and then taken over a longer period of time. The frequency of undesirable effects of phytopharmaceuticals is remarkably low, even if the present lack of data about side-effects is considered. PMID- 12125178 TI - [Interactions of phytotherapeutic drugs, foods and drinks with medicines]. AB - Phytotherapeutic preparations contain a large number of pharmacologically active components. Protective systems have evolved to detoxify and eliminate these xenobiotics. Among them is the cytochrome P450 system and the transporter p glycoprotein in intestine and liver that control the absorption, biotransformation and elimination of drugs. Components of phytotherapeutic preparations can interfere with the function of these systems and lead to interactions with drugs. St John's wort, for example, induces the expression of p glycoprotein and CYP3A4 in liver and intestine and thereby decreases the activity of other drugs. Garlic extracts as well may decrease the activity of drugs that are substrates for CYP3A4. In contrast, grapefruit juice inhibits intestinal CYP3A4. This results in a higher bioavailability of some drugs and possibly more adverse effects. Some relevant interactions were only detected after many years of widespread use, indicating that the treating physician should not only inquire about a change in co-medication but also about the use of alternative medicines or a change in dietary habits when a patient presents with unexpected and unusual adverse effects or a sudden loss of drug efficacy. It would be desirable if more information regarding the potential for interactions with commonly used drugs was available prior to registration of new phytotherapeutic preparations in order to document their safety for patients who require continuous treatment with a drug because of a chronic disease. PMID- 12125179 TI - [Phytotherapy in cardiovascular medicine]. AB - There is widespread use of herbal medicine in patients suffering from cardiovascular diseases. The discussion about the benefit of these drugs is still controversial because of lack of scientific evidence. Ginkgo biloba, Crataegus and Garlic are often recommended substances for patients with cardiovascular diseases. For these substances there is a lot of data available from experimental and clinical studies, unfortunately not always adhering to the criteria of evidence based medicine. Extracts from ginkgo biloba contain several active constituents, mainly flavonoids and terpens, which have antioxidative properties and an inhibitory effect on platelet aggregation by inhibiting platelet activation factor PAF. Ginkgo is mainly used in vascular dementia and peripheral vascular disease. Garlic shows a modest lipid-lowering effect in the same range as a low-cholesterol diet. Effect on blood pressure seems to be at best minor. Crataegus is often used in patients with heart failure because of its positive inotropic effect. Additionally, crataegus acts as an antiarrhythmic substance by prolonging refractory period of the action potential. PMID- 12125180 TI - [Phytotherapeutic drugs in psychiatry]. AB - During recent years, the use of phytotherapeutics in psychiatry has gained enormous significance. The present paper reviews current scientific data on the most important herbal substances including St. John's wort, kava, valerian, and gingko. Although psychotropic phytotherapeutics have been shown to be partly effective in some psychiatric disorders, they cannot generally be recommended as an alternative to conventional medicaments because sufficient data concerning their efficacy, differentiated indication, and safety profile are still missing. Psychiatric patients treated with herbal drugs need intensive medical advice and supervision. PMID- 12125181 TI - [Phytotherapeutic drugs in gastroenterology and hepatology]. AB - More and more patients are trying out herbal medicine. It is estimated that half of the population have used alternative products at least once in their live. Gastrointestinal diseases often require long-lasting treatments involving many side-effects that can impair the patient's motivation. The majority of persons with symptoms of the irritable bowel syndrome or chronic liver disease resort to non-conventional therapies. However, potential hepatotoxicity of herbal products should not be underestimated. In this article, we discuss herbal preparations in specific gastrointestinal and hepatological indications, concentrating on products that have been tested in randomized, controlled clinical trials. Effective symptomatic treatment of obstipation, irritable bowel and inflammatory bowel disease has been demonstrated with plant-derived preparations. On the other hand phytotherapeutic preparations can not be recommended at present for the treatment of cirrhosis or chronic viral hepatitis based on the available data. PMID- 12125182 TI - Conflict of interest. PMID- 12125183 TI - Bacterial contamination of colostrum fed to newborn calves in Quebec dairy herds. AB - A convenience sample of 234 colostral specimens, collected directly from the nursing bottle immediately prior to the first feeding, was studied. Samples originated from 6 farms and were collected over 24 months. Routine bacteriologic techniques were used to quantify the bacterial load of the colostrum, as well as to identify the bacteria. Overall, at least 1 microorganism was cultured from 221 colostral samples (94.4%). By using the upper tolerance level of 100,000 bacteria/mL, 84 samples (35.9%) were considered contaminated. Staphylococcus spp. (57.7%), gram-negative rods (47.9%), coliforms (44.0%), and Streptococcus uberis (20.5%) were among the most frequently isolated bacteria. The relative risk (RR) of contamination with more than 100,000 bacteria/mL was significantly greater in warm months [RR = 2.55, 95% confidence interval (CI): 1.63 to 4.02] than in cool months and in colostrum offered to male calves (RR = 1.55, 95% CI: 1.09 to 2.20). Bacterial load was also associated with the farm of origin (P < 0.0001). When assessing colostrum management, one should consider bacterial contamination. Multiple factors are likely associated with the degree of contamination, and farm specific factors may be important. Further studies are necessary to evaluate the impact of bacterial contamination of colostrum on neonatal health. PMID- 12125184 TI - Complication rate and factors affecting outcome of olecranon osteotomies repaired with pin and tension-band fixation in dogs. AB - The objective of this study was to determine the complication rate and evaluate factors affecting the outcome of olecranon osteotomy in dogs. Medical records were searched to identify dogs that had undergone olecranon osteotomy (stabilized with 2 Kirschner wires and a figure-of-8 wire) during internal fixation of a supracondylar or condylar humeral fracture. Signalment, description of the fracture, parameters regarding the osteotomy and its repair, and radiographic outcome were recorded. A logistic regression model compared patient and technical parameters with the osteotomy outcome. Of the 19 dogs, 7 (37%) had complications of the osteotomy, including osteomyelitis, loss of reduction, and improper placement and migration of the Kirschner wires. Olecranon osteotomy is associated with a high complication rate in dogs; however, there was no correlation between patient-related or technical parameters and the development of complications. Further clinical and biomechanical investigations are warranted to improve the results of olecranon osteotomy and its repair. PMID- 12125186 TI - Survey of Mycobacterium avium subspecies paratuberculosis serological status in beef herds on community pastures in Saskatchewan. AB - Johne's disease is a well recognized problem in dairy herds. Relatively little information is available on either the prevalence or the control of Johne's disease in commercial cow-calf operations. In the fall of 1999, blood samples were collected during pregnancy testing from cows on community pastures in Saskatchewan. Sera from these cows were analyzed using a commercial ELISA for antibodies to Mycoplasma avium subspecies paratuberculosis. All cows from each herd examined at the community pastures were sampled. Of the 1799 samples tested, 15 had sample to positive (S/P) ratios greater than 0.25 and were considered positive (apparent sample prevalence, 0.8%; 95% CI, 0.4% to 1.5%). If we assume test sensitivity of 25% and specificity of 98% as recommended by the National Johne's Working Group, the true sample prevalence is not significantly different from 0.0%. The ELISA S/P results for the antibody test-positive animals ranged from 0.27 to 2.5. If a herd was classified as positive based on one test-positive animal, the average herd apparent prevalence was 15.2% (95% CI, 7.1% to 28.6%). If the potential for false-positive results was considered with 2 or more positive animals being required for positive herd status, the herd prevalence was 3.0% (95% CI, 0.4% to 13.4%). Because of the very low prevalence in cow-calf herds, future research to identify risk factors and control points should target problem herds and utilize a case-control study design. PMID- 12125187 TI - Priapism secondary to penile metastasis in a dog. AB - A 4-year-old, male Newfoundland cross was presented for lethargy, anorexia, and dysuria. The main clinical finding was an enlarged and painful prostate gland. While the dog was hospitalized, priapism developed. Following euthanasia, microscopic examination revealed that a carcinoma involving both bladder and prostate gland had widely metastasized to the penile vasculature. PMID- 12125185 TI - The ischiorectal fossa: an alternative route for the administration of prostaglandin in cattle. AB - Three experiments were conducted to investigate the ischiorectal fossa (IRF) as a route for the administration of prostaglandin F2 alpha (dinoprost) in cattle. In Experiment 1, 21 nonlactating Holstein cows were given 100 micrograms of gonadotropin releasing hormone (GnRH), intramuscularly (i.m.), and, 7 d later, 25 mg of dinoprost into the IRF. Sixteen cows had serum progesterone concentrations > or = 1.0 ng/mL at the time of dinoprost treatment, and all of these had rapid and complete luteolysis; the other 5 cows were not considered to have a functional corpus luteum (CL) at the time of treatment. There were minimal adverse behavioral reactions to the IRF injections and no visible or palpable tissue reactions at the injection site. In Experiment 2, 74 Holstein heifers were given 25 mg of dinoprost by IRF injection. Luteolysis occurred in 84.3% of the heifers with a functional CL (as determined by the serum progesterone concentration at the time of treatment). Of the heifers bred by either natural service or artificial insemination, 61.8% became pregnant. In Experiment 3, 48 beef heifers received dinoprost 7 d after ovulation, as follows: 25 mg, i.m. (n = 9); 25 mg, IRF (n = 10); 10 mg, IRF (n = 10); 10 mg, subcutaneously (s.c.) (n = 10); or 10 mg, intravulvosubmucosally (IVSM) (n = 9). Fewer heifers (P < 0.05) were found to be in estrus or ovulating in the 10 mg IVSM group (0% and 11%, respectively) than in the 25 mg i.m. group (100% and 100%), the 25 mg IRF group (90% and 100%, respectively), or the 10 mg IRF group (80% and 80%). The rates of estrus (50%) and ovulation (50%) were intermediate in the 10 mg s.c. group. In summary, 25 mg of dinoprost injected into the IRF caused minimal behavioral or tissue response and induced luteolysis and fertile estrus. In addition, 10 mg of dinoprost injected into the IRF was as efficacious as 25 mg given either i.m. or into the IRF in inducing estrus and ovulation. PMID- 12125188 TI - Disseminated blastomycosis in a German shepherd dog. AB - A 5-year-old German shepherd was evaluated after collapsing at home following a week of lethargy and anorexia. Systemic blastomycosis was diagnosed histologically at necropsy. Diagnosis and treatment were difficult due to unusual neurological symptoms, the absence of abnormalities on diagnostic tests, and the advanced stage of the disease at presentation. PMID- 12125189 TI - Hypothyroidism in a boxer dog. AB - A 6-year-old boxer was presented with head tilt and facial nerve paralysis. Hypothyroidism was diagnosed and treated appropriately. Hypothyroidism can have an affect on almost any organ system, so the practitioner must be familiar with all clinical signs in order to select appropriate tests and treatment. PMID- 12125191 TI - Diagnosis of blindness in a horse. PMID- 12125190 TI - Darwinian medicine: applications of evolutionary biology for veterinarians. AB - Every medical phenomenon has both a mechanistic explanation and an evolutionary explanation. Veterinarians are accustomed to dealing with the mechanistic, the "what" or the "how", of various disease conditions, and applying treatment accordingly. Darwinian medicine is a field that addresses the evolutionary explanation, the "why" for various medical conditions. This review focuses on these Darwinian explanations and is divided into 4 main categories--host defenses, virulence, genetic conflict, and incomplete adaptation to a changing environment. Each of these areas is reviewed, with examples of evolutionary reasons for disease conditions. Consideration of adaptationist reasons for many of these disease phenomena should make veterinarians better clinicians, educators, and researchers. PMID- 12125192 TI - Neonatal sepsis: evaluation and management. AB - Bacterial antigenic challenge presents a difficult fight for the neonatal immune system, and they have a smaller arsenal of weapons to fight bacterial infections than adults and older children. The baby's own systemic inflammatory response may have detrimental effects on several organs and longer lasting effects on the developing brain. Neurodevelopmental outcomes after maternal chorioamnionitis are worse than neonates without a contaminated intrauterine environment, regardless of gestation age and the baby's culture results. Successes with intrapartum antibiotic prophylaxis decreasing rates of GBS sepsis and maternal chorioamnionitis, have heartened care providers and parents. These results demonstrate the advances possible when specific diseases are made a national health priority, and good clinical trial work is applied to clinical practice. PMID- 12125193 TI - Update on neonatal resuscitation. AB - Pediatricians and other practitioners who attend deliveries maintain resuscitation skills throughout their careers. NRP certification and continuing education programs (PCEP) have done much to standardize the care a newborn receives in the first minutes after birth. Neonates have benefited tremendously from strong national programs (NRP and PCEP) with great local execution of training modules. PMID- 12125194 TI - Breastfeeding: what the primary care provider should know. PMID- 12125195 TI - Preparation of the critically ill neonate for transport. AB - Transport of a critically ill neonate is stressful for all involved. Adequate communication and stabilization will reduce stresses and improve outcomes. Periodic review of the stabilization and care provided to neonates prior to transport can help in further improving the process. Such reviews can be done in conjunction with the Regional Perinatal Center. PMID- 12125196 TI - The care of the back-transported neonate. PMID- 12125197 TI - Discharge planning for very low birthweight infants. AB - As discharge approaches, usually indicated by feeding progression, thermoregulation and other events, it is important for the medical team to develop a plan of action and stick with it, assuming that nothing untoward happens. Experience tells us that families of infants with special needs at discharge cope better and maintain a more positive attitude if plans are clearly defined and followed consistently. A coordinated team approach is also helpful for personnel responsible for arranging equipment, training, and home health services. PMID- 12125198 TI - Neurodevelopmental outcome of neonatal intensive care graduates: a practical approach to developmental follow-up and intervention strategies for primary care physicians. PMID- 12125199 TI - Hyperbilirubinemia in the newborn infant born at term. PMID- 12125200 TI - Neonatal hypoglycemia. PMID- 12125201 TI - Hemoglobin affinity in Andean rodents. AB - Blood hemoglobin oxygen affinity (P50) was measured in three Andean species and in the laboratory rat (control), all raised near sea level. Chinchilla lanigera (Molina, 1792) has an altitudinal habitat range from low Andean slopes up to 3000 m., while Chinchilla brevicaudata (Waterhouse, 1848) has an altitudinal range from 3000 to 5000 m. The laboratory type guinea pig, wild type guinea pig (Cavia porcellus), (Waterhouse, 1748), and laboratory rat (Rattus norvegicus) were also raised at sea level. The Andean species had high hemoglobin oxygen affinities (low P50) compared with the rat. Chinchilla brevicaudata had a higher affinity than Chinchilla lanigera. The wild type guinea pig had a higher affinity than the laboratory type. As has been shown in other species, this is another example of an inverse correlation between the altitude level and the P50 values. This is the first hemoglobin oxygen affinity study in Chinchilla brevicaudata. PMID- 12125202 TI - Vincristine induces somatic segregation, via mitotic crossing-over, in diploid cells of Aspergillus nidulans. AB - Vincristine is an alkaloid widely used as an antineoplastic agent. In eukaryotic cells the drug causes blockage in the G2 phase of the cell cycle and an increase in the frequency of sister chromatid exchanges. Due to the fact that germinating Aspergillus nidulans cells spend most of their cycle in G2 phase, they provide an excellent system for the study of mitotic crossing-over. Taking into account that mitotic crossing-over occurs during G2 period, the evaluation of recombinagenic and aneugenic potential of vincristine is provided with regard to two diploid strains of A. nidulans: a wild strain (uvsH+//uvsH+) and a defective one in DNA repair (uvsH//uvsH). Drug toxicity and its effect on the asexual cycle of A. nidulans has been evaluated as well. Treatment of both strains with vincristine did not change colony growth in the culture, however cytological analyses showed aberrant conidiophores. Recombinagenic potential of vincristine was evaluated by induction of gene homozygosis originally present in heterozygosity diploid strains (Homozygotization Index). Results show that vincristine induces mitotic crossing-over and higher frequency of aneuploid mitotic segregants. The results also show the recombinagenic and aneuploidogenic potential of vincristine and suggest its participation in the induction of secondary malignancies. PMID- 12125203 TI - Cytotoxicity and trypanocidal activity of nifurtimox encapsulated in ethylcyanoacrylate nanoparticles. AB - The aim of the present study was to study the trypanocidal activity of nanoparticles loaded with nifurtimox in comparison with the free drug against Trypanosoma cruzi, responsible for Chagas' disease. Ethylcyanoacrylate nanoparticles acted as the delivery system into cells. As the obligate replicative intracellular form is amastigote, in vitro studies were performed on this form of parasite as well as on cell culture derived trypomastigotes. The fluorescence method used here was very useful as it allowed for the simultaneous study of trypanocide activity and cytotoxicity by determining living or dead parasites within living or dead host cells. According to these results, the greatest trypanocide activity on cell culture-derived trypomastigotes was recorded for nifurtimox-loaded nanoparticles with a 50% inhibitory concentration (IC50) twenty times less than that of the free drug. The cytotoxicity of unloaded nanoparticles at low concentrations was similar to that obtained by free drug when evaluated on Vero cells. Furthermore, nifurtimox-loaded nanoparticles showed increased trypanocide activity on intracellular amastigotes with an IC50 thirteen times less than that of nifurtimox. We also observed that the unloaded nanoparticles possess the previously-described trypanocide activity, similar to the standard solution of nifurtimox, although the mechanism for this has not yet been elucidated. In conclusion, it was possible to establish in vitro conditions using nifurtimox encapsulated nanoparticles in order to decrease the doses of the drug and thus to obtain high trypanocidal activity on both free trypomastigotes and intracellular amastigotes with low cytotoxicity for the host cell. PMID- 12125204 TI - Early and late molecular and morphologic changes that occur during the in vitro transformation of Trypanosoma cruzi metacyclic trypomastigotes to amastigotes. AB - The amastigogenesis primary of T. cruzi occurs naturally when metacyclic trypomastigotes transform into amastigotes within the cells of the mammalian host. The in vitro study of the macromolecular changes that occur over several days during the transformation process should provide significant indications of how the parasite adapts to the mammalian host environment. We show here that metacyclic trypomastigotes pre-incubated at 37 degrees C in a protein-rich medium reach a high degree of transformation to amastigotes when re-incubated in the fresh medium. Giemsa-stained smears show that during the pre-incubation phase, the metacyclic trypomastigotes undergo lengthening at the posterior end and a thinning out of the entire body. SDS-PAGE analysis of polypeptides and glycopeptides or Western blot with stage-specific antisera analyses indicate that the in vitro primary amastigogenesis is associated with abrupt changes in protein, glycoprotein, and stage-specific antigens that occur simultaneously during the first 24 hours of pre-incubation. Since the differentiating system consists of a rich media at 37 degrees C, temperature and medium constitution must trigger a macromolecular differentiation to amastigotes that precedes the morphological transformation by several days. This transformation is associated with the rearrangement of stage-specific antigens and takes place when the culture medium is changed. PMID- 12125207 TI - [Human capital in Sciences]. PMID- 12125206 TI - Peroxidase and phenylalanine ammonia-lyase activities, phenolic acid contents, and allelochemicals-inhibited root growth of soybean. AB - The influence of the allelochemicals ferulic (FA) and vanillic (VA) acids on peroxidase (POD, EC 1.11.1.7) and phenylalanine ammonia-lyase (PAL, EC 4.3.1.5) activities and their relationships with phenolic acid (PhAs) contents and root growth of soybean (Glycine max (L.) Merr.) were examined. Three-day-old seedlings were cultivated in nutrient solution containing FA or VA (0.1 to 1 mM) for 48 h. Both compounds (at 0.5 and 1 mM) decreased root length (RL), fresh weight (FW) and dry weight (DW) and increased PhAs contents. At 0.5 and 1 mM, FA increased soluble POD activity (18% and 47%, respectively) and cell wall (CW)-bound POD activity (61% and 34%), while VA increased soluble POD activity (33% and 17%) but did not affect CW-bound POD activity. At 1 mM, FA increased (82%) while VA reduced (32%) PAL activities. The results are discussed on the basis of the role of these compounds on phenylpropanoid metabolism and root growth and suggest that the effects caused on POD and PAL activities are some of the many mechanisms by which allelochemicals influence plant growth. PMID- 12125208 TI - Cloning and comparison of ten gene sequences of a Chilean H. pylori strain with other H. pylori strains revealed higher variability for VacA and CagA virulence factors. AB - We have cloned and sequenced ten Helicobacter pylori genes from a Chilean strain (CH-CTX1) including: a cytotoxin VacA fragment, a CagA fragment (A17), a species specific protein (TsaA), urease subunits (UreA, UreB), a flagellin subunit (FlaB), heat shock proteins (HspA and HspB), adhesin (HpaA) and a lipoprotein (Lpp20). We compared their deduced amino acid sequences with the corresponding sequences from three unrelated H. pylori strains, including fully sequenced strains 26695(UK) and J99(USA), and found that eight of them (UreA, UreB, FlaB, HspA, HspB, Lpp20, TsaA and HpaA) presented more than 97.3% identity. In contrast, VacA partial sequence showed lower identity values (93.2-94.9%). Moreover, we found major differences in the A17 region respect to the number and arrangement of the internal repeated elements when sequences from different strains were aligned. The A17 regions from strains CH-CTX1 and 26695 are very similar (91.8% identity) but lacked 6 repeated elements when compared to the Australian strains ATCC 43526 and NCTC 11637. The CCUG 17874 A17 region showed the largest deletion involving 9 repeats. A17 size differences between strains CCUG 17874 and CH-CTX1 were verified by PCR and polypeptide size. Such differences may explain variations in virulence among H. pylori strains as well as diversity in serum immunoreactivity. PMID- 12125210 TI - Analysis of 5382insC (BRCA1) and 6174delT (BRCA2) mutations in 382 healthy Chilean women with a family history of breast cancer. AB - Breast cancer is the most common malignancy among women. Chilean studies reveal that this cancer presents the third highest mortality rate. A family history of breast cancer is one of the major risk factors for the development of this disease. BRCA1 and BRCA2 are the two main hereditary breast cancer susceptibility genes, and mutations in these genes are related to inherited breast cancer. In specific populations only some mutations have been found to be associated with susceptibility. The purpose of this study was to establish the frequency of 5382insC (BRCA1) and 6174delT (BRCA2) germline mutations in 382 healthy Chilean women with at least two relatives affected with breast cancer and in probands and their relatives from 8 high risk families for breast cancer, using mismatch PCR assay. The results obtained showed that 5382insC and 6174delT mutations were not found in either of the groups studied. The ethnic origin of the contemporary Chilean population and the data reported in the literature suggest that these mutations may be absent or have a very low frequency in this population.. This genetic study is part of a breast cancer screening program that also includes annual mammography and clinical breast examination over a five-year period. Strategies to reduce morbidity and mortality associated with breast cancer lie in early detection in women with genetic risk. PMID- 12125211 TI - Glucose transporters: expression, regulation and cancer. AB - Mammalian cells depend on glucose as a major substrate for energy production. Glucose is transported into the cell via facilitative glucose transporters (GLUT) present in all cell types. Many GLUT isoforms have been described and their expression is cell-specific and subject to hormonal and environmental control. The kinetic properties and substrate specificities of the different isoforms are specifically suited to the energy requirements of the particular cell types. Due to the ubiquitousness of these transporters, their differential expression is involved in various disease states such as diabetes, ischemia and cancer. The majority of cancers and isolated cancer cell lines over-express the GLUT family members which are present in the respective tissue of origin under non-cancerous conditions. Moreover, due to the requirement of energy to feed uncontrolled proliferation, cancer cells often express GLUTs which under normal conditions would not be present in these tissues. This over-expression is predominantly associated with the likelihood of metastasis and hence poor patient prognosis. This article presents a review of the current literature on the regulation and expression of GLUT family members and has compiled clinical and research data on GLUT expression in human cancers and in isolated human cancer cell lines. PMID- 12125212 TI - [A epistemometric view of some biological disciplines in Chile]. AB - During the last decade the articles published by Chilean Research Centers grew 1,73 which compares to the 2.34 fold increase of mainstream research articles registered as a whole in Latin America. However, the relative impact of the Chilean publications surpassed that of Latin America. In Biological Sciences, traditionally the strongest research area within Chile, Latin America also shows a steeper slope of growth. Qualitatively, biological disciplines in Chile are comparable to those published in Latin America although in Chile there are specialties as Physiology that surpass the average world's impact. The scientometric data is consistent with the fall in individual grants that the Chilean Research Fund (FONDECYT) has been allocating during the last decade. PMID- 12125213 TI - PSNA partners in development of Girl Scout Nursing Patch. PMID- 12125214 TI - PSNA budget summary 2002-2003. PMID- 12125215 TI - [Primary osteosynthesis of the odontoid process: a multicenter study]. AB - PURPOSE OF THE STUDY: Direct osteosynthesis is a method of choice for the treatment of odontoid process fractures. It is based on insertion, from the anterior approach, of one or two screws from the C2 body into the apex of the odontoid across the fracture line. The tensile action of screws results in compression of fragments and stabilization of the fracture. The aim of the study was to evaluate a group of patients treated by this method and to compare our results with those reported in the foreign literature. MATERIAL: A total of 99 patients were treated by direct osteosynthesis of the odontoid in the departments involved in the study between 1994 and 2001. METHODS: Patients indicated for this surgery were those with fractures of type II according to Anderson and D'Alonzo and those with type III fractures but only when the fracture line went across the articulation surfaces of C1-C2, when closed reduction was not possible or the patients were not indicated for halo fixation. Direct osteosynthesis was not applied to fractures with comminution at the base of the odontoid, irreducible fractures, odontoid fractures combined with dislocated fractures of the atlas or pathological fractures. Severe kyphosis of the cervical spine or a large thoracic cage was also regarded as a contraindication. RESULTS: All the 99 patients were followed up from 3 up to 102 months, with an average of 28.5 months; only in seven patients, the follow-up period was shorter than 6 months. The most frequent subjective complaint was a painful operation wound. This usually resolved within two weeks of surgery. Except for four patients, alle were satisfied with the outcome. Type II fractures were diagnosed in 84 and type III fractures in 15 cases. A total of 174 screws were inserted into the odontoid processes of 99 patients. A single screw was used in 25 and two screws in 73 patients. In one case, three screws had to be inserted. Screw lenght ranged from 36 to 44 mm, diameter was 40.9 mm. Three months after surgery, X-ray examination, both in flexion and extension, did not reveal any instability in any of the patients. No morphological change in the C2-C3 intervertebral space was observed Of 92 (92.9%) paitents under longterm follow-up, 84 (91.3%) showed complete healing of the fracture, three died and five patients eventually developed pseudoarthrosis, which was due to a broken screw in three of them. This condition was treated by dorsal fixation of C1-C2 according to Magerl or by one of the dorsal cerclage techniques. The group was free of any perioperative complications related to the anterior approach or injury to nerve structures by screws. DISCUSSION: The most frequent subjective complaint was a painful operation wound. Treatment of odontoid fractures varies according to the type of injury, bone quality and also practice at each department. Type II injuries are highly unstable and, because of the small fracture surface, their healing ability is much lower than in type III fractures. Previously, most of the patients with odontoid injuries were treated conservatively by immobilization in a plaster cast or a brace or, later, by a halo device. In the long term, however, they showed a high proportion of pseudoarthroses (10 to 100%). Direct osteosynthesis of the odontoid by screws permits the maintenance of rotation of the C1-C2 mobile segment. We followed the scheme of indications used abroad but did not perform osteosynthesis to correct pseudoarthrosis. The number of osteosyntheses healed (91.3%) was also in agreement with the literature data. Earlier, we used two screws for all types of fractures. Recently, we have preferred insertion of a single screw in type II and III fractures in narrow odontoids. In the later, there is no danger of rotational dislocation during screw insertion; to insert one screw from the centre of the C2 base is easy and speeds up the procedure. However, in displaced type II and type II T fractures, two screws are a necessity. Similarly to other authors, we recorded a slight limitation of cervical spine rotation in patients at long-term follow-up, particularly in elderly subjects with advanced osteochondrosis. No complications leading to deterioration of the patient's state were recorded. CONCLUSIONS: Direct osteosynthesis is a method of choice for most of the type II and indicated cases of type III fractures of the odontoid process of the axis. This surgical procedure facilitates restoration of anatomical conditions of the spine and its immediate stability. Consequently, patients can be readily mobilized and rehabilitated. PMID- 12125216 TI - [Meniscoids of the intervertebral joints]. AB - PURPOSE OF THE STUDY: A large amount of material was used to study the distribution, location and shape of meniscoids in intervertebral joints of the human spine, from the atlanto-occipital joint to the sacrum, in order to find out how many of intervertebral joints had mobile meniscoids. These might be regarded as possible causes of spinal blockade or other vertebrogenous complaints. MATERIAL: The materials provided by the Department of Anatomy and Department of Forensic Medicine at the Faculty of Medicine of Charles University in Pilznen included 20 cadaverous spines from humans aged 20 to 80 years. METHODS: Access to each joint was provided by dissection of the articular capsule from the lower articular processes of the vertebra situated above. In the orthograde view, all meniscoids were described in terms of shape, size, consistence and location. Their structure was ascertained by histological examination of cross sections stained with haematoxylin and eosin. RESULTS: Meniscoids varying in shape and size were found in all of the intervertebral joints. They were classified by their histological structure as synovial, fat and fibrous meniscoids. The first category was observed frequently, the last only rarely. A total of 29 mobile meniscoids were recorded, most of them in the lumbar spine. Most of the meniscoids present in the cervical spine were of synovial and less frequently of fat types. Meniscoids found in the thoracic spine were poorly developed synovial ones and those present in the lumbar spine were of all types and were also largest in size. The most conspicuous meniscoids were seen in the spines that showed degenerative changes in intervertebral joints. Large fat pads were found in atlanto-occipital and atlanto-axial joints. Mobile meniscoids, most of them present in the lumbar spine (6.4% of all joints.), were connected with the capsule by a thin pedicle and it was possible to move them over a half of the articular surface. Some inter-individual changes were also found; in some spines, the most developed meniscoids were fat pads, in the others, these were synovial meniscoids. Spines of younger individuals showed a predominance of synovial meniscoids with smooth surfaces that arched against the articular cavity. In spines of elderly individuals, meniscoids were rough, in some cases fibrous in structure, and had a lobulated or frayed edge. DISCUSSION: The shape, location of meniscoids and their presence in every joint indicate their definite role for the spine: they compensate the incongruence of articular surfaces, fill in empty spaces and facilitate spread of synovial fluid during translation movements. Variability in shape, size and location of meniscoids give support to the view that meniscoids developed secondarily in relation to the morphogenesis of articular surfaces and that they are fully adapted to the shape and function of the joint. Mobile meniscoids, particularly fibrous ones, can get wedged between articular surfaces due to a sudden, rush movement (entrapment theory) or can be caught between the edge of an articular surface and the articular capsule attachment (extrapment theory). This situation may result in either mechanical or functional blockade of the spine and a subsequent painful condition due to compression of nerves and reflex contraction of muscles. Direct evidence of such blockade and the validity of either hypothesis can today be provided by magnetic resonance imaging. CONCLUSIONS: All intervertebral joints, along the length of spine, possess capsule processes, i.e., meniscoids, which can be classified as synovial, fat and fibrous. Meniscoids are most developed in the lumbar and cervical spine. They serve to compensate for the incongruence of articular surfaces and to fill in empty spaces. Mobile, peduncular meniscoids can, at sudden or non-physiological movements, be caught between articular surfaces and cause spinal blockade and painful conditions. Manipulative treatment is, therefore, justified in indicated cases. PMID- 12125217 TI - [Replacement of the vertebral body with an expansion implant (Synex)]. AB - PURPOSE OF THE STUDY: This paper describes replacement of the vertebral body with the expansion implant Synex. Usually, autologous bone graft is used to replace the vertebral body. In patients with bone cancer or multiple injuries to the spine, cement filling is preferred whereas, in other indicated cases, implants are inserted, of which Harms' titanium cage has been the most common one. However, this needs filling with a large amount of bone tissue and it is often difficult to adjust its size into the space available. Telescopic devices, on the other hand, are easier to implant and their application requires only a minimum amount of autologous bone tissue. MATERIAL: In the period from January 2000 to June 2001, we used telescopic implants Synex to replace vertebral bodies in 34 patients. Indications for treatment were: vertebral fractures in 14, post traumatic kyphosis in six, vertebral metastatic tumours in eight and a primary tumour in six patients. METHODS: In 25 cases, the vertebral body replacement was completed by posterior stabilization using internal fixation and, in nine cases, by anterior stabilization with a Ventrofix fixator. In 32 patients, the implant was inserted from the anterior approach and, in two, from the posterior approach following complete spondylectomy. RESULTS: The L1 vertebra was replaced most frequently (nine patients), then T 12 (seven patients) and L2 (six patients). For treatment of fresh fractures, the Synex implant was used in 14 cases. Of these one was inserted from the posterior approach in the L1 region where trauma had caused severe injury to the spinal cord. In spinal tumours. Synex was used in 14 patients, i.e., in six with diagnosed plasmacytoma, in two with metastatic dissemination from prostate carcinoma, in four with vertebral metastases from breast cancer and in two patients with non-differentiated metastases. The anterior approach was performed by conventional thoracotomy or combined thoracotomy and lumbotomy in 20 patients and a less invasive retroperitoneal approach was used in 12 patients. One patient died of multiple metastases at 7 months after surgery and one patient had relapse of a local tumour resulting in paraparesis that required a repeat decompression of the spinal canal. The operation took 1 h and 50 min when the anterior approach and anterior stabilization with a Ventrofix fixator were used; the operation lasted from 3 h 20 min to 6 h 10 min when complementary posterior stabilization was involved. The patients were followed up for 2 to 24 months. No failure of the implant in terms of migration, change in position or penetration into adjacent vertebral bodies occurred. DISCUSSION: The replacement of a vertebral body has conventionally been performed with the use of a massive bone graft. However, collection of an autologous bone graft large enough to suit this purpose is not always possible. Complications at the donor site have been described. A homologous bone graft carries a risk of disease transmission and the reconstruction ability of a massive graft has not been confirmed for certain. Cement filling augmented with Kirschner's wires is usually used in cancer patients. Titanium cages require application of a large amount of spongiose bone tissue into their interior. Consequently, bone in the centre fails to remodel. A sharp edge of the mesh may induce migration of the cage towards the vertebral body and failure of the implant. Mechanical failure and collapse of cages have also been described. Telescopic cylindrical implants, on the other hand, need only a small amount of spongiose bone tissue to fill. They can be adapted directly to the implantation site by means of a special distractor and, therefore, before adjusting its final length, the exact position and orientation of the implant can be achieved in the space prepared. This facilitates close contact with the endplates of adjacent vertebral bodies and the development of osteointegration. The use of telescopic implants enabled us to avoid the force that is often necessary to apply during insertion of Harms' cages in the patients whose spines had already been stabilized with posterior fixation or to avoid the need of a triple surgical procedure in order to achieve better stability of the implant. In two patients, Synex was inserted from a non-standard posterior approach. Indications for Synex implantation should be evaluated in view of disease prognosis in each patient. If only limited survival is expected, cement filling with K-wires should be preferred. CONCLUSIONS: Synex is a sophisticated implant to replace severely damaged vertebral bodies regardless of the nature of lesion. Its application required additional stabilization by either posterior or anterior fixation (internal transpedicular fixator and Ventrofix or Kaneda, respectively). Its use is indicated in post-traumatic defects of vertebrae in acute or poorly healed scervical. PMID- 12125218 TI - [Somatometric and impedance measurements of the lower extremities in orthopedics]. AB - PURPOSE OF THE STUDY: The objective was to demonstrate the contribution of somatometry and vascular impedance measurement to the evaluation of somatic status and nutrition, particularly the state and function of the vascular system of lower extremities, in patients with arthropathy of the weight-bearing joints. MATERIAL: Impedance plethysmography and impedance phlebography appeared to be the best methods for assessment of vascular hemodynamics; pulse oxymetry, carried out on toes, was most convenient for measurement of haemoglobin saturation with oxygen (HbO2). The state of vessels was determined according to the CEAP classification. Somatometric methods included measurement of the circumference and volume of calves, and assessment of body height, at the anthropometric wall, and weight to calculate the body mass index (BMI). RESULTS: The lower extremities affected by arthropathy were compared with the unaffected ones by means of CEAP classes and related to the body mass values of patients. It was found that only 15 men and 26 women had normal body mass. The rest of the patients were overweight or obese (grade I and II). Only 14 men and 17 women were free of signs of vascular insufficiency, the others were classified in the range of C1 to C4 of the CEAP system. Impedance measurements showed lower mean values for affected extremities and for overweight or obese patients. A comparison of the parameters of arterial and venous hemodynamics between the affected and unaffected extremity by means of the paried t-test gave statistically significant results in patients with higher BMI grades and higher classes of vascular insufficiency, regardless of their gender. The results of HbO2 saturation measurement showed normal values in both men and women. DISCUSSION: To meet the requirements of orthopaedists, the Biophysical Laboratory carried out non-invasive measurements of somatometric parameters and vascular impedance values in patients with arthropathy of the weight-bearing joints of lower extremities. The results were similar to those of a large Framingham cohort study on knee arthropathy and obesity. The data derived from measurement of basic impedance, specific impedance and pulse-related impedance changes show that it is important to record and evaluate skin lesions and oedema because they are related to vascular disease. The finding that an increasing volume the calf was indirectly related to a decreasing basic impedance of calf vessels was confirmed by our results. This information is important for the evaluation of impedance measurement results in order to make diagnosis in individual patients. CONCLUSIONS: Our results showed that, in the group investigated, (i) there was a high proportion of overweight and obese patients; (ii) assessment by CEAP classification revealed higher grades of vascular insufficiency; (iii) affected extremities showed a significant decrease in arterial and venous haemodynamics, particularly in women with higher-grade vascular insufficiency; (iv) there was no difference in HbO2 saturation measured on toes. PMID- 12125219 TI - [Arthroscopic reconstruction of the anterior cruciate ligament using the transtibial technique and a graft from the patellar ligament--results after 5-6 years]. AB - PURPOSE OF THE STUDY: Arthroscopic reconstruction of the anterior cruciate ligament (ACL) by transtibial technique with free graft from the patellar tendon fixed by interference screws is a commonly used procedure. The aim of this study is to evaluate the results 5 to 6 years after the surgery. MATERIAL: Fifty-two of 79 patients operated on in 1995 were checked in the period of 2000-2001. The average age of patients at the time of surgery was 26 years (range, 15-20 years), there were more men (62%), prevailing were injuries resulting from sports activities (92%). METHOD: We have used evaluation according to IKDC. We focused on the problems at the site from which the graft was harvested, on the difficulties in the kneeling position and walking on knees. Evaluation of the activity was based on the Tegner score. On radiographs we assessed the length of the patellar tendon, dilatation of the tibial canal and the incidence and degree of osteoarthritis. RESULTS: General evaluation according to IKDC was normal or nearly normal in 79% of patients. The difference of the anterior displacement of tibia between the operated on and non-operated on knee measured by arthrometer (KT 1000, 89N) was < 3 mm in 58% of patients and < or = mm in 95% of patients. Significant difficulties on the operated on side in the kneeling position and walking on knees were reported by 3 patients, slight difficulties were reported by 28 patients. Twenty patients reported slight difficulties also on the non operated on side. Prior to the injury the average level of activity after Tegner was 7.9 (range, 6-10), prior to the operation 2.7 (range, 0-6) and at the time of check examination 7.0 (range, 4-10). The same level of activity as before the injury was reported by 61.5% of patients. Shortening of the patellar tendon on the operated on side > 5% (6 mm) was found out in 1 patient, shortening of < or = 5% (2-3 mm) in 30 patients. Tibial canal on the lateral projection 1 cm beneath the joint line was wider than 10 mm (dilated) in 6 patients. On the operated on knee osteoarthritis was found of the medial compartment of degree B and C in 42% of patients and on the non-operated knee in 19% patients. Aggravation of osteoarhritis of the medial compartment by one degree occurred after 5 to 6 years after the surgery in 4 out of 20 patients. All of them underwent meniscectomy. Eighty-six per cent (19 of 22) of patients with osteoarthritis of the medial compartment underwent medial meniscectomy. DISCUSSION: General results according to IKDC are compared with results published by Aglietti, Jomha and Patel. Dilatation of the tibial canal was found out in 6 patients and it was associated with the graft-tunnel mismatch. The relationship between meniscectomy and arthritis is well-know. In 31% of patients after medial meniscectomy there was found out narrowing of the joint line to 2-4 mm (degree C). In case of partial medial meniscectomy the posterior horn, i.e. the most important part of the meniscus, was in most cases removed. Worsening of the arthritis after resection of the minor radial lesions of the lateral meniscus or after leaving incomplete lesions of the posterior horn of the lateral meniscus untreated has not been observed. CONCLUSION: The surgery allows to improve the stability and extent of activity in most of the patients. The most frequent problem consists in the difficulties at the site of graft harvesting which may contribute to the decrease of subjective satisfaction. The relation between shortening of the patellar tendon and patellar difficulties was not proved. In the patients after medial meniscectomy osteoarthritis progresses even after ACL reconstruction. PMID- 12125220 TI - [Infections in surgery of idiopathic scoliosis]. AB - PURPOSE OF THE STUDY: In this retrospective study, we evaluated infectious complications in the patients undergoing surgical treatment for idiopathic scolions in order to identify risk factors for postoperative infections. MATERIAL: A total of 786 patients with idiopathic scoliosis were operated on during 24 years. In 754 (96%) cases, we used the posterior approach, involving posterior fusion and internal fixation, and subsequent immobilization in a brace. During that period, we recorded 15 (1.9%) deep wound infections in the area of fusion. Early infections were treated by debridement and lavage, with targeted administration of antibiotics, while instrumentation was kept in place. In late und recurrent infections, instrumentation was always removed. METHODS: We investigated a relationship between the infectious agent and the device used, the length of period between surgery and the onset of infection, the effect of device removal on curve progression, the agent causing infection and the effect of allergy to metal or infectious lesions at other body sites on the outbreak of infection. RESULTS: Early infections (within 6 weeks) were observed in six, late in nine patients. Repeat operations were necessary on average after 487 days. Staphylococcus aureus, the most frequent infectious agent, was isolated from four patients; on four occasions, cultivation was negative. Allergy to nickel was found in four patients. Infection was most often associated with the most frequently used Harrington Instrumentation (six cases, 1.1%). However, in relation to the number of patients treated, infection frequency was highest in TSRH (5.0%) and Isola (4.8%) devices. When Miami Moss fixation or the anterior approach was used, no infection was recorded. In comparison with the non-infected cases, the patients with infectious complications showed the same average values for the curve before and after surgery. At a check up, however, the loss of correction increased to 6 degrees and, after instrumentation removal, to 10 degrees as against 3 degrees in the non-infected patients. Pseudoarthrosis developed in two cases. DISCUSSION: The incidence of deep wound infections in patients who had surgery for idiopathic scoliosis was comparable with the data in the relevant literature. A higher number of infections, particularly late ones, in patients treated with the use of modern instrumentation is probably related to a higher volume of these implants. Early infections are a rare feature and their cause is known (allergy, sepsis). Treatment involves surgical intervention; in early infections, instrumentation is retained but is removed in late infections. CONCLUSIONS: Even though our group included a low number of patients with infections, we can conclude that risk factors for the development of infectious complications associated with surgical treatment of idiopathic scoliosis are as follows: allergy, higher age, large volume of metal used for stabilization and the presence of another infectious lesion. PMID- 12125221 TI - [Experience with the Walter-Motorlet/Poldi hybrid hip joint endoprosthesis in our department]. AB - PURPOSE OF THE STUDY: Patients with total hip replacement, in whom the hybrid prosthesis consisting of a Walter-Motorlet socket and a Poldi shaft was used, were followed up and the results were evaluated. MATERIAL: Eighty eight total hip replacements were carried out in 85 patients in the period from 1995 to 1997. Of these, 64 were evaluated at the follow-up lasting on a average 67 months; 14 patients who failed to come for a check-up were excluded from the study. METHODS: All patients were operated on from the Watson-Jones approach. They underwent clinical examination, outcome assessment by the Harris classification and X-ray examination in anterior-lateral and axial projections. RESULTS: The average age of patients at the time of surgery was 58.3 years. Indications for the operation included primary arthritis in 45.3%, post-dysplastic arthritis in 31.3%, secondary arthritis in 10.9% and trauma in 12.5% of the patients. The results evaluated according to the Harris classification system were excellent or very good in 83%, satisfactory in 6% and poor in 11% of the replacements. On subjective evaluation, the joint free from pain had 37.5% and slight paint was experienced by 48.5% of the patients. Para-articular ossification developed in 39% of the patients. A total of five sockets (7.8%) and three shafts (4.7%) showed signs of aseptic loosening. DISCUSSION: This study showed worse outcomes of total hip replacement, as classified by the Harris system, that those reported in the relevant Czech or foreign literature. Our results of pain evaluation were also worse than those reported by the Czech an foreign authors. The diagnoses leading to total hip replacement and their frequency were in agreement with the foreign literature data and this was also true for the occurrence of para articular ossification. In the reports by the Czech authors cited, however, the reasons for hip replacement, in terms of diagnosis frequency, were different and the development of para-articular ossification was recorded in fewer cases. CONCLUSIONS: The occurrence of soft linings of the socket was high (in 51% of the patients) and became a prerequisite for subsequent socket loosening. This finding was also in accordance with the results reported in the literature quoted. PMID- 12125222 TI - [Nitric oxide and its role in orthopedics]. PMID- 12125223 TI - [Purulent Salmonella arthritis of the ankle joint]. AB - In the case study the authors focus on Salmonella enteritidis which caused purulent osteoarthritis of the ankle and sepsis. Salmonella osteoarthritis is one of the nidal forms of the salmonella infection. The authors briefly present etiology, pathogenesis, diagnosis and principles of the treatment of infectious osteoarthritis. They analyze a case of a 56 years old immunocompromised patient who was treated at the authors' department for a late diagnosis with an already developed septicemia. They describe the course of the disease, complications and the result of the treatment and discuss the pitfalls of the diagnosis and treatment. Only a timely diagnosis followed by a sufficiently vigorous surgical intervention with drainage of the joint and combined with a timely, targeted and long-term antibiotic therapy leads to the management of this severe nosologic unit. PMID- 12125224 TI - Kinetics of in vitro lipolysis of human very low-density lipoprotein by lipoprotein lipase. AB - BACKGROUND AND AIM: An initial step in the catabolism of triglyceride-rich lipoprotein involves the hydrolysis of the triglyceride moiety by lipoprotein lipase (LPL). As differences in the lipolytic behaviour of very low-density lipoprotein (VLDL) particles have been observed, it is possible that different VLDL particles have a different affinity to the enzyme, which means that their fate may partially depend on the LPL-mediated hydrolysis of their triglyceride content. Our aim was to determine whether variation in VLDL chemical composition affects their properties as a substrate for LPL. METHODS AND RESULTS: Isolated VLDL was incubated in vitro with bovine LPL to determine substrate affinity. Under optimal assay conditions, free fatty acids were measured and the kinetic indicators for in vitro triglyceride hydrolysis (Km and Vmax) were calculated. VLDL cholesterol (VLDL-C), VLDL-apoB and the cholesterol/triglyceride ratio were assessed and the triglyceride/protein and triglyceride/apoB ratios were calculated as lipoprotein size estimators. VLDL-C, VLDL-apoB and the VLDL C/triglyceride ratio positively correlated with Km: r = 0.52, p < 0.01; r = 0.52, p < 0.03; r = 0.69, p < 0.001 respectively. No correlation was found between the VLDL-triglyceride/protein or the VLDL-triglyceride/apoB ratios and Km (r = -0.20, and -0.06 respectively, p = not significant). Of the subjects' anthropometric characteristics, only the waist/hip ratio significantly correlated with Km: r = 0.63, p < 0.01. CONCLUSIONS: In the present study, we investigated the substrate function of VLDL particles in vitro. Enzyme affinity seems to be associated with cholesterol-triglyceride content or the number of VLDL particles rather than particle size. It may be expected that VLDL with a low cholesterol/triglyceride ratio will be efficiently lypolised by LPL, thus leading to the formation of a smaller particle with atherogenic potential. PMID- 12125226 TI - Atorvastatin increases HDL cholesterol in hypercholesterolemic patients. Evidence of a relationship with baseline HDL cholesterol. AB - BACKGROUND AND AIM: It has been reported that atorvastatin increases high-density lipoprotein cholesterol (HDL-C) more in patients with low than in those with high baseline HDL-C levels. This may have a biological explanation, but also suggests a statistical artifact known as the regression to the mean. METHODS AND RESULTS: Atorvastatin 10 mg/day led to a 4% increase in HDL-C after two months in 67/121 patients with hypercholesterolemia (55%), who had lower baseline HDL-C levels than the patients in whom HDL-C did not increase. In the patients with baseline HDL-C below the median, HDL-C significantly increased whereas no change was observed in patients with baseline HDL-C above the median. The correlation coefficient between pre- and post-treatment HDL-C was 0.84, thus suggesting a regression to the mean. However, the regression artifact did not entirely explain the increase in HDL-C in patients with low baseline HDL-C or the lack of an increase in those with high baseline HDL-C. The adjusted mean increase was 5.4% in patients with low pretreatment HDL-C, and 2.4% in the patients with high pretreatment HDL-C. Multiple regression analysis with the changes in HDL-C as the dependent variable showed that baseline HDL-C and the changes in serum triglycerides independently contributed to the change in HDL-C levels. CONCLUSIONS: Atorvastatin 10 mg/day increases HDL-C more in patients with low pretreatment HDL-C levels, an effect that seems to be related to the hypotriglyceridemic activity of the drug. PMID- 12125225 TI - LDL cholesterol lowering by bile acid malabsorption during inhibited synthesis and absorption of cholesterol in hypercholesterolemic coronary subjects. AB - BACKGROUND AND AIMS: Recent large-scale trials have consistently documented the fact that a 25-35% reduction in low-density lipoprotein cholesterol (LDL-C) can delay the progression of atherosclerosis. This raises the question as to how much it is possible to reduce serum cholesterol using feasible therapies. The aim of this study was to investigate the cholesterol-lowering efficacy of a triple therapy combining bile acid malabsorption with the inhibition of cholesterol synthesis and absorption. METHODS AND RESULTS: Eleven consecutive hypercholesterolemic coronary patients from Lipid Clinics on a low-fat, low cholesterol baseline diet added simvastatin (20 mg/day) for three months, and then dietary plant stanol ester margarine (2.25 g of stanols/day) for eight weeks; finally, cholestyramine 8 g/day was added for another eight weeks. This was a before-after trial, in which the results of each period were compared with baseline and those of the previous period. Serum lipids were quantitated using commercial kits, and serum sterols by means of gas-liquid chromatography. Simvastatin lowered LDL-C by 39% (p < 0.001), and additional stanol ester margarine by a further 13% (p < 0.05). The triple treatment led to 67% reduction from baseline (p < 0.001), with all LDL-C values being < 2.6 mmol/L, and increased high-density lipoprotein cholesterol (HDL-C) by 15% (p < 0.01). It also increased the serum lathosterol/cholesterol ratio (p < 0.01), thus indicating an upregulation of cholesterol synthesis, and increased the serum sitosterol ratio (p < 0.01) despite the simultaneous consumption of plant stanols. CONCLUSIONS: The massive reduction in LDL and increase in HDL-C obtained using our triple therapy suggests that the combination of stanol ester with only moderate doses of statin and resin makes it possible to control LDL-C levels effectively in hypercholesterolemic subjects. PMID- 12125227 TI - Long-term (18-month) efficacy of atorvastatin therapy in type 2 diabetics at cardiovascular risk. AB - BACKGROUND AND AIM: Dyslipidemia (increased triglyceride and low high-density lipoprotein [HDL] levels, with normal or slightly increased total cholesterol levels) is a common characteristic of type 2 diabetics and a major risk factor for cardiovascular diseases. The aim of this study was to evaluate the long-term efficacy of atorvastatin in a cohort of type 2 diabetics. METHODS AND RESULTS: Participants were divided into 3 groups on the basis of whether they had evidence of myocardial infarction or coronary lesions (group A), a family history of hypercholesterolemia (and/or cardiovascular diseases) and total cholesterol levels constantly above 270 mg/dL in blood samples taken at regular 4-month intervals and previously never at target level (group B), or clinical and/or instrumental (electrocardiogram) evidence of cardiovascular risk (group C). Their mean age was 64 +/- 7 years, known disease duration 0.5 +/- 3 years, body mass index (BMI) 27.7 +/- 1.3 Kg/m2, and haemoglobin A1c 8 +/- 0.6%. Total cholesterol was 256 +/- 24 mg/dL in group A, 298 +/- 25 mg/dL in group B and 244 +/- 31 mg/dL in group C (p < 0.05: group B vs groups A and C). HDL-cholesterol (HDL-C) was 45 +/- 7 mg/dL, triglycerides 225 +/- 20 mg/dL, systolic and diastolic blood pressure (DBP) respectively 144 +/- 7 and 85 +/- 8 mmHg, fibrinogen 330 +/- 23 mg/dL and microalbuminuria 58 +/- 9 mg/L. Eighteen months' atorvastatin treatment (10 mg/day in 106 subjects, 20 mg in 14 subjects, 30 mg in 5 subjects, and 40 mg in 30 subjects) led to a significant decrease in total and low-density lipoprotein (LDL)-cholesterol and triglyceride levels (p < 0.01), with about 86% of the patients achieving target levels, and a significant (p < 0.05) increase in HDL-C. There was a significant decrease in fibrinogen, microalbuminuria and DBP (p < 0.01), without any change in diet, BMI, physical activity or antihypertensive treatment. No new cardiovascular events or hospital admissions due to cardiovascular diseases were recorded during the 18 months of the study. CONCLUSIONS: These long-term treatment findings confirm and validate previous medium-term results, and suggest that atorvastatin therapy is effective and safe in the primary and secondary prevention of cardiovascular complications in type 2 diabetes. PMID- 12125228 TI - Macrophage function and stability of the atherosclerotic plaque: progress report of a European project. PMID- 12125229 TI - High-density lipoprotein metabolism in familial hypercholesterolaemia: significance, mechanisms, therapy. AB - AIM: To assess the significance, mechanisms and therapy of impaired high-density lipoprotein (HDL) metabolism in patients with heterozygous familial hypercholesterolaemia (FH). METHODS: Review of epidemiological, metabolic and clinical literature with synthetic analysis of data referring to HDL. PRINCIPAL FINDINGS AND SYNTHESIS: Low HDL is a powerful risk factor for coronary artery disease (CAD) in FH. Low HDL in FH is a metabolic sequel of insulin resistance, which is the central feature of the visceral accumulation of adipose tissue acquired in later life. Insulin resistance increases hepatic secretion of triglycerides that subsequently results in remodelling of holo-HDL particles and enhanced catabolism of apolipoprotein A-I. Gene-gene and gene-environment interactions may contribute to the pathogenesis of low HDL in FH. Low HDL may also point to other cardiovascular risk factors, such as hyper-remnantaemia, increased small dense low-density lipoproteins (LDL), procoagulopathy and vascular insulin resistance. Visceral obesity should be vigorously identified and treated in FH. Judicious addition of fish oils, niacin and fibrates to a statin may be required to optimise HDL metabolism. Extracorporeal removal of LDL cholesterol should aim not to extract HDL from plasma. CONCLUSIONS: With the increasing incidence of obesity in the community, the metabolic syndrome will overmanifest in patients with FH. Impaired HDL metabolism is an important consequence of this syndrome for FH patients, because it will per se or in collusion with other associated risk factors accelerate the progression of atherosclerosis and CAD. Dysregulation of HDL metabolism principally results from hepatic insulin resistance and remodelling of HDL particles. Obesity should be prevented and aggressively treated in FH and use of a safe combination drug regimen considered to correct the dysmetabolism of HDL in these patients. PMID- 12125230 TI - Vasculoprotective effects of oleic acid: epidemiological background and direct vascular antiatherogenic properties. AB - BACKGROUND AND AIMS: The use of the "Mediterranean diet" as a means of preventing atherosclerotic vascular disease is gaining increasing acceptance. As early as in the late 1950s, it was found that the inhabitants of Greece and Southern Italy had a very low incidence of coronary artery disease and that, among other components, their diets were very rich in oleic acid, the main constituent of the olive oil, making up about 29% of their daily caloric intake. It is now clear that, in addition to its relatively minor effects on cholesterol levels, oleic acid directly interferes with the inflammatory response characterising early atherogenesis due to the endothelial expression of adhesion molecules for circulating monocytes. RESULTS: In in vitro models of early atherogenesis based on cytokine-stimulated cultured endothelial cells, we have observed that the incorporation of oleic acid in total cell lipids is accompanied by decreased expression of a number of major pro-inflammatory proteins, such as endothelial leukocyte adhesion molecules. CONCLUSIONS: The results of our investigations indicate that oleic acid has a direct vascular atheroprotective effect, and suggest that it may be possible to prevent atherosclerosis by modulating the vascular response to classical triggers (high levels of cholesterol and the advanced glycation end-products of diabetes) using a strategy that is fundamentally different from, and therefore complementary to, drug-based therapy. PMID- 12125231 TI - The value of partnering with patients. PMID- 12125233 TI - You are up to bat! Second base: identify the needs of our patients. PMID- 12125234 TI - Slow food. PMID- 12125235 TI - Top 10 deficiencies in long-term care. PMID- 12125236 TI - Care and treatment of residents (TX). Part I: The care planning process. PMID- 12125237 TI - [The necrotizing enteritis by Clostridium perfringens type C in piglets: practical observations, control and epidemiology]. AB - Necrotizing enteritis in piglets is caused by the spore-forming anaerobe Clostridium perfringens type C. The pathology is believed to be due to the production of beta-toxin by the agent and the infection tends to persist in affected herds despite appropriate hygiene measures. During the period 1989 to 2001, 35 outbreaks were observed in 15 herds in a limited geographic region in northwestern Switzerland in the canton of Fribourg. Initial outbreaks of acute disease were followed by chronic manifestations of necrotizing enteritis in eleven herds. Since clinical symptoms in case of chronic necrotizing enteritis and of mixed infections were rather non-specific, diagnosis in all herds was confirmed by gross pathological analysis, intestinal histology and bacteriological culture. After initial evaluation in 135 litters (1500 piglets), metaphylaxis consisted of penicillin for outbreaks of acute disease or of amoxycillin-clavulanic acid pending results of laboratory confirmation. Vaccination with a C. perfringens toxoid vaccine (Gletvax 6, also containing E. coli pilus antigens) together with penicillin chemoprophylaxis was used in 2400 litters (27,000 piglets). In 12 to 17% of litters disease recurred during this combined prophylaxis. Necrotizing enteritis persisted in all affected herds throughout the follow-up period. PMID- 12125238 TI - [The necrotizing enteritis by Clostridium perfringens type C in piglets: II. Molecular epidemiology study]. AB - Investigations were performed on shedding of C. perfringens in sows from four different pig farms. In two farms where no outbreaks of necrotizing enteritis had been observed, no strains of C. perfringens producing beta-toxin were detected in the faeces of sows. In contrast, C. perfringens strains producing beta-toxin were detected in sows on both farms suffering outbreaks of acute necrotizing enteritis. Strains of C. perfringens producing beta-toxin were invariably positive for the beta 2-toxin gene. However, strains carrying the beta 2-toxin gene only (i.e. negative for beta-toxin) were present in animals on all farms with roughly similar frequencies (mean 28.2% carriers). Some sows carried C. perfringens strains of both toxin genotypes simultaneously. Whereas these data further support the role of betatoxin as a cause of necrotizing enteritis, the role of beta 2-toxin in intestinal disease of piglets remains unclear. To establish the role of faecal shedding vs. environmental contamination as reservoirs of C. perfringens type C, strains were isolated from teats and feedlot trough swabs (toxin genotype beta/beta 2), as well as from fodder (genotype beta 2). However, sows carried this pathogen intermittently and in small numbers. This renders an individual, reliable diagnosis of carrier sows very difficult. Ribotyping of 34 C. perfringens isolates of different toxin genotypes showed five distinct profiles. Different toxin genotypes can belong to the same ribotype, and the same toxin genotype can be present in different ribotypes. Thus, even if a majority (79.4%) of strains investigated in a limited geographic region belonged to ribotype 1, ribotyping offered discrimination of strains beyond toxin typing. PMID- 12125240 TI - [Polymelia in a Holstein Friesian calf]. PMID- 12125239 TI - [Reference values in the cerebrospinal fluid of calves between four and eight weeks of age]. AB - Reference values for the following parameters were established in the cerebrospinal fluid of 27 calves between four and eight weeks of age: specific weight, protein concentration, erythrocyte count, total leucocyte count with cell differentiation, creatin kinase activity, glucose and sodium. If possible, the findings were compared with those of other authors in calves and adult bovines. With 24.3 cells per microliter the 90% quantile of the total leucocyte count was seated significantly above comparable values for adult bovines. Hence, in individual cases markedly higher leucocyte counts can be expected in the cerebrospinal fluid of calves. In agreement with other authors, the protein concentration in calves was lower than in adult bovines. The reference range for creatin kinase activity was increased whereas the one for sodium was only slightly increased compared to earlier investigations in calves and in adult bovines. PMID- 12125241 TI - Certification of poliomyelitis eradication. PMID- 12125242 TI - WHO-UNICEF joint statement on strategies to reduce measles mortality worldwide. PMID- 12125244 TI - [Pathomorphosis of infiltrative pulmonary tuberculosis: clinical aspects]. AB - Based on the data available in the literature and their own findings, the authors have established that the pathomorphism of infiltrative pulmonary tuberculosis in the preantibiotic age was characterized by negative features, then underwent phases of positive changes and relative stabilization, thereafter it again acquired obviously marked negative peculiarities. At present, the clinical features of new cases of infiltrative pulmonary tuberculosis in Perm residents are the predominance of acute onset (67.3%), disseminated (55.8%) and destructive (91.6%) pattern of specific lesion accompanied by bacterial isolation (84.6%). PMID- 12125243 TI - [Adverse effects of multi-drug therapy in patients with tuberculosis and ways of their correction]. AB - Thirty and seventy six patients with tuberculosis receiving multidrug therapy and 107 healthy persons were examined. There is a high rate (38.5%) of adverse responses to antibiotics, deteriorated baseline vitamin deficiency, PLO-AOA imbalance, poorer liver tests, preserved protein deficiency, and secondary immunodeficiency. The use of Trofosan, an enterosorbent-antioxidant complex, and MRT as pathogenetic drugs corrects the disorders found. With this, the incidence of adverse decreased on the average by 2 times and the efficiency of inpatient treatment increased. PMID- 12125245 TI - [The detection rate and clinical and diagnostic values of L forms of the pathogen in patients with pulmonary or extrapulmonary tuberculosis]. AB - The detection rate and clinical and diagnostic values of L-forms of pathogens were determined in patients with pulmonary and extrapulmonary tuberculosis. Simultaneous culturing the specimens for typical and L forms of Mycobacterium tuberculosis (MBT) increased the number of positive results by isolating only L forms by 10.3% in patients with pulmonary tuberculosis and by 27.7% in those with extrapulmonary tuberculosis. No bacterial isolation in tests only for typical forms of MBT is shown not to be true and a purposeful search for L-forms of MBT enhances the efficiency of a bacteriological test. This is of great significance in confirming the specific nature of the disease, its progression, in choosing a treatment policy, in evaluating is efficiency, in defining prognosis, and in correcting preventive measures in the focus of tuberculous infection. With extrapulmonary tuberculosis, the tuberculous nature of isolated L-forms has been evidenced by the polymerase chain reaction. PMID- 12125246 TI - [Surgical treatment of bilateral forms of pulmonary tuberculosis]. PMID- 12125247 TI - [Specific features of spontaneous pneumothorax in patients with pulmonary tuberculosis]. AB - To study the impact of pulmonary tuberculosis on the natural development of spontaneous pneumothorax and on the outcomes of its treatment, data on 589 patients were retrospectively analyzed. Two hundred and eighty patients had primary spontaneous pneumothorax and 170 had secondary nontuberculosis pneumothorax. Tuberculous pneumothorax was in 139 patients, of them 71 had decay cavities. The authors defined the causes of pneumothorax, the severity of respiratory diseases, assessed admission X-ray data, evaluated the efficiency of the first 24 hours of treatment, complications and mortality rates in these groups of patients. The efficiency of treatment for spontaneous pneumothorax has been ascertained to be determined by the degree of lung decay, by the adequacy of pleural cavity drainage, and by the rationality of a tactic algorithm rather than by the type of an infectious agent (Mycobacterium tuberculosis, nonspecific microflora). PMID- 12125248 TI - [Current reconstructive surgery for tuberculosis of the vertebral column and joints]. AB - There has been recently a rise in the incidence of complicated forms of bone and joint tuberculosis, one of its causes is its late diagnosis. The outcomes of surgical treatment in 390 patients with tuberculosis of the spine and large joints are presented. Vascularization of bone tissues and a graft has been found to substantially increase the efficiency of an intervention used and the use of carbon-carbonic implants and posterior clamps accelerates the time of formation of a bony trochlea and diminishes deformity of a vertebral column part undergone an operation. To improve the functional results of mobilizing surgery for tuberculous arthritis and its sequelae, low-traumatic operations using the arthroscopic devices, perichondrioplasty and endoprosthesis of large joints are being practically introduced. PMID- 12125249 TI - [New approaches to diagnosis and prevention of childhood tuberculosis]. AB - The concept that tuberculosis is a disappearing disease has turned out to be untrue not only for Russia, but also for many countries of the world. The last good year was, in terms of tuberculosis morbidity among the children of Vladivostok, 1992 (15.5 per 100,000 children). In the past years, the morbidity has substantially increased. The basic method for detecting tuberculosis is tuberculin diagnosis in children and fluorography in adolescents. Nonspecific vaccinations were found to affect the inversion of early negative results to questionable results. To cover the maximum number of children with revaccinations I and II, it is advisable to repeat the tuberculin test following 3-6 months in case of questionable Mantoux test with 2 TE in the decreed period in previously non-infected persons. PMID- 12125250 TI - [Use of the plant hepatoprotector Galstena tuberculostatics-induced hepatic lesions: experimental and clinical study]. AB - The efficiency of the use of the natural drug Galstena (Richard Bittner GmbH) was experimentally and clinically studied on a model of damage induced by toxic doses of tuberculostatics (rifampicin, isoniazid, pyrazinamide) in laboratory rats and in patients with different forms of pulmonary tuberculosis and with hepatitis caused by specific antituberculous drug therapy, who were treated at the Clinic of Phthiziology. Galstena was found to have marked hepatoprotective properties and to be able to prevent renal and pancreatic disorders. The drug also showed an antioxidative activity. The use of Galstena in patients with pulmonary tuberculosis substantially reduced the magnitude of clinical and laboratory signs of drug-induced hepatic damage. PMID- 12125251 TI - [Efficiency of the use of peppermint (Mentha piperita L) essential oil inhalations in the combined multi-drug therapy for pulmonary tuberculosis]. AB - The essential oil of peppermint (Mentha piperita L.) has been found to have an in vitro pronounced and equal antimycobacterial effect in doses of 300 and 600 micrograms/ml, respectively. The use of its inhalations (upon 20-min heat evaporation into the room atmosphere for 2 months) as a supplement to combined multidrug therapy for pulmonary tuberculosis has indicated their significantly high positive effect in terms of abacillation (by 26.8 and 58.5% with doses of 0.01 and 0.005 ml/m3, respectively). This was followed by earlier positive X-ray changes in the lung and by attenuation of the intoxication syndrome. The findings suggest that peppermint essential oil may be used in combined multidrug therapy in patients with disseminated and infiltrative pulmonary tuberculosis. PMID- 12125252 TI - [Respiratory and renal functions and blood electrolytic composition in patients with pulmonary tuberculosis in the treatment of heart failure]. AB - The authors studied respiratory and renal functions and blood electrolytic composition in 94 patients with pulmonary tuberculosis in the treatment of heart failure with angiotensin-converting enzyme inhibitors (ACEI) and an angiotensin II blocker (AIIB). At the same time they examined a control group of 24 patients untreated with ACEI and AIIB. All the patients received combined antituberculous therapy. Cardiac glycosides and diuctics were given in some cases. During 1-2 months, the patients from the experimental group additionally took captopril, 12.5-25.0 mg, twice a day or prestarium, 2.0-4.0 mg, once a day, or cossar, 50 mg, once a day. After the treatment, ACEI and AIIB were found to have no effect on blood gas and electrolyte composition, and respiratory and renal functions. With captopril and ramipril there was a higher patency of minor bronchi in the experimental groups than in the controls. PMID- 12125253 TI - [Red blood cell antibodies and anemia in pulmonary tuberculosis]. AB - To examine the association of anemia with the formation of antibodies to intrinsic red blood cells, 56 phthisiosurgical patients were examined. Red blood cell antibodies were detected in 66.7 of patients with anemia and in 36.9% without anemia. The results were processed by the chi 2 Pierson test, which confirmed a 95% probability of association of hemolytic anemia with the formation of antibodies to red blood cells. The direct antiglobulin test and the enzyme assay may be recommended for examination of patients with pulmonary tuberculosis in the preoperative period to reveal risk factors of progressive clinical manifestations of anemia. PMID- 12125254 TI - [Characterization of Mycobacterium tuberculosis strains prevalent in North-West of Russia by spoligotyping]. AB - Spoligotyping was used for genotyping of 238 M. tuberculosis cultures isolated from 302 patients with pulmonary tuberculosis living in the north-west of Russia, including those in Saint Petersburg, in 1998-2001, and the M. tuberculosis strains H37Rv, M. bovis, and M. bovic BCG and M. tuberculosis in the specimens painted for microscopy. The quality of spoligotyping was high and similar to that when DNA from the cultured mycobacteria and slide scrapes. The findings were compared with the data available in the International database. There was heterogenicity in the microbial population: the clinical strains showed 58 (RO R57) types of profiles, of them 56% were revealed in single cases. The spoligotype RO(1; S1) was found in 153 (51%) patients. This spoligotype is predominant in the world and characteristic of polyresistant high-transmissive strains of the genetic family Beijing. The spoligotypes R24 (251), R14 (252), and R8 (253) first described by the authors are now encountered only in Russia. To extend the Russian database that contains M. tuberculosis spologotyping profiles from different regions will promote the improvement of an epidemiological surveillance system and tuberculosis control programmes not only within administrative territorial entities, but also within the whole country. PMID- 12125255 TI - [Quantitative characteristics of the cellular link of immunity and a lymphocytic response to tumor-associated antigens in patients with tuberculosis and malignant neoplasms of the lung]. AB - The relative quantities of basic populations and subpopulations of lymphocytes and their response to tumor-associated antigens were estimated in the peripheral blood patients with tuberculosis and malignant neoplasms of the lung. In both diseases, the changes in the count of immunocompetent cells are similar and they characterize immunodeficiency typical of these conditions. A response to tumor associated antigens was found only in patients with lung cancer, the pattern of this response depending on the grade of a tumor. PMID- 12125256 TI - [Epidemiological value of the efficiency of treatment in patients with tuberculosis]. AB - The main tuberculosis epidemiological parameters in the Ukraine are analyzed. The present epidemic of tuberculosis has been found to be characterized by the predominance of mortality rates over morbidity ones along less efficiency of treatment and abnormal stabilization of the incidence of recurrent tuberculosis. The efficiency of treatment (ceased bacterial isolation and healed decay cavities and caverns) is of epidemiological value. The range of these parameters, whose achievement may significantly affect immunological parameters is to be defined. It is concluded that it is necessary to make a large-scale epidemiological study of tuberculosis to establish many causal correlations of the present-days epidemic of tuberculosis. PMID- 12125257 TI - [Current view of the problem of spontaneous pneumothorax]. PMID- 12125258 TI - [Methodology for the assessment of epidemiological indices of pulmonary tuberculosis and the efficiency of tuberculosis control measures]. PMID- 12125259 TI - [Efficiency of locoregional and lymphotropic intermittent chemo-immunotherapy for pulmonary tuberculosis]. PMID- 12125261 TI - [Regularities in forming chromosome aberrations in cattle lymphocytes from irradiation in vitro]. AB - Aberrations in lymphocytes of cattle blood exposed to gamma-radiation with doses from 1 to 7 Gy were studied. The rate of variable cells depended linearly on the irradiation dose, whereas the total frequency of aberrations, as well as that of dicentric and annular chromosomes followed a linear-quadratic dependence. PMID- 12125260 TI - [Cytogenetic effects of low doses of radiation in mammalian cells: analysis of the hypersensitivity phenomenon and induced resistance]. AB - The induction of cytogenetic damage after irradiation of chinese hamster cells and human melanoma cells within a dose range 1-200 cGy was studied. The anaphase and metaphase analysis of chromosome damage and micronuclei test were applied. The hypersensitivity (HRS) at doses below 20 cGy and the increased radio resistence at higher doses (IR) were shown with all cytogenetic critheria for both cell lines. The phenomenon of HRS/IR was reproduced in synchronic as well as in a synchronic population of chinese hamster cells. This fact shows that HRS was caused by high radiosensitivity of all cells and can not be explained by any differential sensitivity of cells in different phase of the cell cycle. So it was supposed that the increasing radio-resistence is determined by the inclusion of the inducible repair processes in all cells. This conclusion consents with the facts, that there was no evidence of HRS on dose-effect curves and that some parts of pre-existent damage was repaired after preliminary irradiation with low doses (1-20 cGy) which induce repair processes. It can be concluded that same inducible repair processes an analogous in mechanisms underlying in the base of HRS/IR phenomenon and adaptive response. PMID- 12125262 TI - [Regularities in forming chromosome aberrations in cattle lymphocytes in external and combined radiation exposure]. AB - Aberrations in lymphocytes of cattle blood exposed to gamma-radiation and combined radiation were found. It was shown that seven days after the exposure to doses from 64.5 to 103.3 mC/kg a number of aberration varied in the range from 16.1 to 37.7 per 100 cells, whereas a frequency of aberrated cells was from 13.5 to 29.8. After the exposure to 77.3 mC/h and the following inclusion of fused radioactive particles into the fodder (48.1-762.2 Mbq/kg of the live weight), the number of aberrations increased from 25.5 up to 55.0 per 100 cells and the number of aberrated cells increased from 21 up to 43%. PMID- 12125263 TI - [Prediction of the severity of damage and disruption of work ability in reaction of the body to alcohol load prior to radiation exposure in the superlethal range]. AB - In experiments on 121 white non-linear rats, 44 Papio hamadryas and 29 Macaca fascicularis, animals' reactions on the alcohol impact (AI) and following exposure to supralethal doses were compared. The animals were intravenously injected with 5% ethanol in the glucose solution, 2.1 g/kg for rats and 0.46-0.51 g/kg for monkeys. Monkeys' response to AI was scored in four-point scale by estimating of abnormalities in motor activity, coordination of motion and changes in conditioned reflex activity. It was shown that changes in the ability of alcohol-injected rats to perform the learnt exercises in the "jump box" could be used for prediction of their response to the exposure to supralethal doses of ionizing radiation. Observing the AI-response in monkeys along with a method "function of spying for moving object" made possible to predict not only a general degree of loss of working ability but also to estimate individual impairments of spying functions. In 65% monkeys high similarity of the reactions to AI and ionizing radiation was observed. PMID- 12125265 TI - [Evaluation of cell membrane permeability for Ca2+ and adenylate cyclase in sheep peripheral blood cells, exposed to low doses of radiation]. AB - Chronic irradiation of sheep with doses of 2.6 and 12.9 mC.kg was characterized by the modification of the adenylatecyclase activity and Ca2+ permeability of plasma membrane in cells of the peripheric blood, with no changes in the clinical and hematological indicators. The observed effects are assumed to result from structural and dynamic variations in the lipids of membranes. PMID- 12125264 TI - [Change in heparin-binding activity of rat brain proteins after single exposure to ionizing radiation at a dose of 0.25 Gr]. AB - Sex-dependent description of the total protein heparin-binding activity in different rat brain regions was established in experiment on white rats. In norm named parameter above in males, than in females on a 22%. The specific alteration of the total heparine-binding activity was found in cortex, cerebellum, middle brain, hippocampus, striatum and pons 1, 12, 24, 120 and 168 hours after whole body single exposure to ionizing radiation with a dose of 0.25 Gy. The most pronounced changes were typical for limbic system. In the cortex and cerebellum of males no significant changes of the activity were found. The data suggest that modification of the system of heparin/heparin-binding proteins, which takes part in regulation of the intercellular brain communication, occurs at early hours after exposure to ionizing radiation. PMID- 12125266 TI - [Interphase death of irradiated thymocytes--result of the "bystander effect"]. AB - It was shown that interphase death of thymocytes, which mechanism was established by author's laboratory already over 10 years ago belongs to phenomena which recently was named as caused by "bystander effect". PMID- 12125267 TI - [Morphological types of radiation-induced hematopoietic tissue cell death, its biological essence and significance at various stages of developing acute radiation injury]. AB - The results of morphologic investigation of radiation-induced cell death types in human and animal (dogs) hematopoietic tissue within an acute radiation injury are presented. It has been shown that early and late necrobiosis of myelokaryocytes occurs via apoptosis. An attempt to designate the pathogenic role of apoptosis in hematologic syndrome of acute radiation sickness was performed. PMID- 12125268 TI - [Theoretical analysis of the reason for unsatisfactory approximation of experimental data on fractionated effect of gamma-radiation and (or) neutrons on lymphocytes at the G0 and G1 stage]. AB - The causes were analyzed of unsatisfactory approximation, based on a mathematical model of cytogenetic effect of interaction of lesions induced by two doses of gamma-radiation and/or neutron in human blood lymphocytes at the stage G0 and G1. For the analysis the published data were used concerning the effect of inhibitors of protein and DNA synthesis on aberration yield in G0- and G1-cell cycle stages at different periods of time after their irradiation with different doses of gamma-rays and neutrons. It is assumed that the cause of the unsatisfactory approximation is the effect of the combined repair of lesions caused by two dose fractions under the process of replication repair caused by the first dose fraction. PMID- 12125269 TI - [Role of superoxide dismutase in maintaining cellular homeostasis upon exposure to gamma-rays and nickel sulfate]. AB - It was shown that the inhibition of one of the main enzymes of antioxidant system -superoxide dismutasw (SOD)--increased the level of damage in gamma-irradiated and NiSO4-treated human cells. The viability of cells pretreated with inhibitor of SOD (TRIEN) decreased in the experiments with gamma-radiation and with NiSO4. DNA repair and replication involved in cell homeostasis also were suppressed in the cells with inhibited SOD. These data are served as proof of participation of SOD in the defence of human cells against oxidative radicals. PMID- 12125270 TI - [Specificity of remote influence of gamma-irradiation of plant seeds on non irradiated ones]. AB - In laboratory and field conditions the consequences of remote influence of gamma irradiated plant seeds on the non-irradiated ones were studied. It was found the the consequences of the influence depended on the time of storage in conditions ensuring gaseous exchange as well as on radiation susceptibility. PMID- 12125271 TI - [Phenomenology and genesis of changes in the total bioelectrical activity of the brain in response to electromagnetic radiation]. AB - A review of the date on phenomenology and genesis of changes in overall activity of brain in response to electromagnetic radiation is presented. It is validated the nonspecific synchronizing influence of the considered factor. This influence can be a reason of epileptoid class convulsions under growth of its biological significance as an irritant of central nervous system. PMID- 12125272 TI - [Histogenetic, metabolic, and immunologic aspects of the effect of infrared laser radiation on injured skeletal muscles from irradiated and nonirradiated rats]. AB - Using biochemical, histological, morphometric and cytogenetic methods, it was shown that low-intensive infrared laser radiation (total dose 3.6 J/cm2), applied to the injured rat skeletal muscles, stimulated metabolism and regeneration more efficiently in the muscles locally exposed to 20 Gy X-rays compared to the unexposed muscles. The laser irradiation promoted postradiative recovery in bone marrow cells, but did not provide normalization in thymus lymphocyte activity. PMID- 12125273 TI - [Enzymatic activity of some tissues and blood serum from animals and humans exposed to microwaves and hypothesis on the possible role of free radical processes in the nonlinear effects and modification of emotional behavior of animals]. AB - The dependence of activities of actomyosin ATPase, alkaline phosphatase, aspartataminotranspherase, monoaminoxidase and that of affective rat behavior on frequency of modulation of microwaves (0.8-10 microW/cm2) was explored at short time actions. Series of nonlinear phenomenons, inexplicable from positions of the energy approaches are revealed, The working hypothesis explaining opportunity of high performance of weak and super-weak microwaves and other revealed phenomena by resonance interaction of such electromagnetic radiofrequency radiation with paramagnetic molecules of biological tissues was proposed. This resonance interaction activate free radicals and initiate auto-supporting and auto intensifying of chain chemical reactions. The spontaneous autocatalytic oxidation of catecholamines enlarges a common pool of free radicals, capable to participate in such enhanced generating. The protective role of monoaminoxidase is postulated. Monoaminoxidase is basically located on an outer surface of mitochondrias and it is deaminating monoamines. The deaminating prevents penetration of catecholamines inside of mitochondrias and their quinoid oxidation there with formation of free-radical semi-quinons, capable to destroy system of ATP synthesis. These inferences are obliquely confirmed by the experimentally revealed correlation between activity of monoaminoxidase and integrative activity of the rat brain. PMID- 12125274 TI - [Effect of low intensity pulsed laser radiation of basic parameters of aging in Drosophila melanogaster]. AB - The effect of the low-intensive impulse laser radiation (LILR) on the life span of Drosophila melanogaster has been studied. The flies at various stages of their life (larvae and imago) were exposed to LILR. The estimation of the effect of LILR was carried out on the basis of the analysis of the basis parameters of aging. We found out increasing as well as shortening effects of the life span. The direction of the effect depends on parameters of radiation, stage of development of irradiated individuals and their sex. PMID- 12125275 TI - [Change in kidney function upon administration of (131)I to the rat]. PMID- 12125276 TI - [Evaluation of the bioavailability of radionuclides from soil in cattle by an in vitro method]. AB - Factors have been examined which influence the radionuclides sorption from soil particles by fluid imitating rumen liquid of the cattle. It is noted that the extent of extractability of 137Cs from soil is influenced mainly by presence of potassium ions in modelling liquid. Major factor influencing the release of 90Sr from soil is pH value of the medium. The presence of heavy metals salts affected the release of radionuclides from soil particles. The data observed make it possible to use the modelling solutions to predict bioavailability of radionuclides from soil as well as to predict possible contamination of animal products (milk and meat). PMID- 12125277 TI - [Accumulation of 90Sr in field crops in conditions of radioactive pollution of agricultural lands]. AB - In the field experiment the influence of manure, peat, lake silt and their combinations on the uptake of 90Sr by barley, corn and potatoes was studied. Saturation of soil with organic substance, the decrease in soil acidity resulted in the lower accumulation of 90Sr by plants. In the beginning of the experiment, the specific activity of barley grain was from 75 to 132 Bq/kg. After three years of application of fertilizers the content of 90Sr in the barley grain decreased to 39 Bq/kg. There were no increase in 90Sr accumulation by the basic field crops with time. PMID- 12125278 TI - [Effect of microcirculation on the morphofunctional state of skin]. AB - Peculiarities of the morphofunctional state of skin, to a large extent, depend on the state of microcirculation. As the skin nutrition is provided by the microvessels, the question arises if the disturbances in microcirculation affect the parametres of the morphofunctional state of the skin. The main clinical investigations on microcirculation are represented in this review: the influences of systemic and local microcirculatory disturbances on the physiological parameters of skin, as well as the age changes of microcirculation and their influences on the dermatosis have been elucidated. The main questions on physiology and pathology of the skin microcirculation and methods of their study have been described. PMID- 12125279 TI - [Effects of hypogeomagnetic fields on the structural-functional activity of rat cerebral cortex]. AB - Structural functional organization state of brain cortex has been determined by histological, hystochemical and morfometrical methods from 104 animals, which were exposed by long stay of hypogeomagnetic field. It is found the disturbance of brain cortex micro-circulation; the disturbance of the enzymes activity of oxidate-reductive types, analogous to hypoxic; monthly chronobiological cycle of neurons content of basic structural functional types replace by changes of this index. The authors suppose that as a result the desynchronization of brain cortex activity as functional system appear. PMID- 12125280 TI - [Effect of irbesartan, an antagonist of AT-1 receptors for angiotensin II, on L arginine metabolism in arterial hypertension]. AB - Effects of an antagonist of AT-1 receptors for angiotensin-II (Ang-II) irbezantane on the NO-synthase and arginase ways of the metabolism of L-arginine were studied in plasma and erythrocytes of the patients with arterial hypertension. The intensity of the non-oxidative arginase way of L-arginine metabolism in plasma and erythrocytes has been shown to be inhanced at hypertension versus the normotensive patients, while the activity of the alternative oxidative NO-synthase way was reduced. Inhibiting AT-1 receptors for Ang-II with high-affinity antagonist irbezantane normalized the ratio between two alternative ways of L-arginine metabolism through inhibiting the arginase way and reciprocal activating the NO-synthase way both in human plasma and erythrocytes. PMID- 12125281 TI - [Effect of dexamethasone on mast cell reaction in inflammation]. AB - Injections of dexamethasone to rats with carrageenan-induced acute aseptic peritonitis have been shown to cause more expressed mast cell reaction in the inflammatory focus, as compared to those with natural development of the process. More expressed degranulation of the cells, a decrease in their number and a release of histamine were observed. It corresponds to the ability of dexamethasone to increase an accumulation and degranulation of leukocytes in an inflammatory focus. The results testify to the role of an increased activation of mast cells in the mechanisms of anti-inflammatory action of glucocorticoids and support the significance of leukocytes in the mast cell reaction in inflammation. PMID- 12125282 TI - [Effect of acetylcholine and adenosine triphosphate on membrane potential of intact rat aorta endothelium in aging]. AB - Changes in the electric reactions of intact endothelium of isolated rat aorta were studied in conditions of aging. A value of resting membrane potential of endothelium of old rats (24-26 mo) was significantly greater, than in control group of animals (6 mo). In responses to acetylcholine and ATP the old rats did not show a typical course of reactions, which was peculiar to control animals. Inability of old rats to produce a typical response to acetylcholine and ATP may signify a disturbance of links between endothelial cells, which resulted in functional clusterization of the latter. The data obtained suggest a possible mechanism of decrease in production and release of potent vasodilator--nitric oxide--due to changes that occur in the electric properties of endothelium in aging. PMID- 12125283 TI - [Effect of exogenous leukotrienes and lipoxygenase inhibitors on apoptosis and necrosis in cultured rat hepatocytes]. AB - Liver cell death by apoptosis and necrosis occurs upon the liver injury. Lipoxygenase pathway of arachidonic acid metabolism is known to regulate the viability and apoptosis in some cell types, but its role in hepatocyte cell death is not fully understood. We studied the influence of leukotrienes (LT) and lipoxygenase inhibitors on apoptosis and necrosis in rat hepatocyte primary culture by double staining with Hoechst 33342 and propidium iodide and electron microscopy. Treatment with general lipoxygenase inhibitor nordihydoguaiaretic acid and 5-lipoxygenase inhibitor caffeic acid (2. 10(-5) M) for 4 and 24 h induced hepatocyte apoptosis. LTB4 and LTC4 (10(-8) M) decreased the number of living cells and increased the number of necrotic cells. LTs exerted the same necrotic effect on hepatocytes, treated with lipoxygenase inhibitors. It is important that LTs decreased apoptosis induced by inhibitors treatment. These data suggest that lipoxygenase pathway of arachidonic acid metabolism is important regulator of hepatocytes viability and apoptosis The increase of lipoxygenase product formation, in particular LTs, may diminish apoptosis and increase necrosis in hepatocytes upon the liver injury. PMID- 12125284 TI - [Effect of intracellular trypsin on characteristics of voltage-dependent inactivation of chloride currents in prostatic cancer cells]. AB - By means of the patch-clamp technique we have studied the effects of intracellular applied trypsin, a known modulator of membrane channel function, on the properties of the Cl- current induced by hypotonicity-obliged cell swelling (ICl, swell) in human prostate cancer epithelial cells, LNCaP. Intracellular infusion of 1 mg/ml of trypsin into LNCaP cells via the patch pipette shortened the delay for the onset and the time of development of ICl, swell in response to hypotonicity as well as accelerated the rate of current diminution following the return to isotonic conditions. The maximal density of ICl, swell in the presence of intracellular trypsin was 2-fold higher while the current voltage-dependent inactivation at high depolarizing potentials was virtually eliminated. Intracellular co-application of the trypsin inhibitor together with trypsin abolished all effects of trypsin. We conclude that VRACs share a great degree of functional and structural homology to voltage-gated Na+, K+ and Cl- channels by having intracellular inactivation domain subjected to proteolytic cleavage that function in conformity with "ball-and-chain" inactivation model. PMID- 12125285 TI - [Physiological activity of peptide complex from heart tissues]. AB - It was studied an influence of heart peptide complex on myocardial metabolism of intact animals, animals with immune deficiency of heart peptides and acute immobilizing stress. There was no significant changes of biochemical, fibrinolitic and thromboplastine properties of heart tissues by 0.1 and 1.0 mg/kg administration of peptide complex to animals. Treatment with peptide complex leads to a restoration of these properties by immune deficiency. Moreover, peptide complex has stress protective effect by acute immobilization. PMID- 12125286 TI - [Hormonal stimulation of female sexual maturation]. AB - The mixture of the sexual steroids: estradiol (E2) and dihydrotestosterone (DHT) was injected to the female rats at prepuberty. This hormonal combination has been established to accelerate the different stages of the female rats' puberty--the vaginal opening and first ovulation. The puberty was accompanied with an increase in the estrogen level in the blood and a reduce of the relation between testosterone and E2. The stimulating effect of the hormonal combination on the ovarian folliculogenesis was observed and post-ovulatory corpus luteum were found in the experimental rats. We suggest that DHT blocked overripening the ovarian follicles at estrogenizating, and E2 reduces the negative effect of DHT on the hypophysis. The steroid combination (E2 + DHT) is effective for stimulating female puberty. PMID- 12125287 TI - [Effect of prenatal hyperandrogenism during pregnancy and on postnatal development of rats]. AB - Hyperandrogenia was modelled in the female rats within the whole term of pregnancy by means of testosterone or dihydrotestosterone. Both androgens have been established to induce significant prenatal mortality and high percentage of mortinatus and postnatal mortality of progeny. Hyperandrogenization influenced negatively the sexual development and reproductive function of the female progeny versus the male one. PMID- 12125288 TI - [Effect of various antisecretory drugs on morphological changes in rat gastric mucosa in experimental stomach ulcers]. AB - In experiments with white rats of Wistar linea there was reproduced the model of acetate gastric ulcer by Okabe. Since 7 day after ulcerogenous trauma the animals received one of the antisecretory drugs (sandostatine, famotidine, lansoprazole) for 14 days. There was studied the influence of these drugs on microcirculation and epithelium of gastric mucosa using morphometry. Using sandostatine and famotidine showed to decrease for sure the volumetric density of capillaries comparing with intact animals. Taking lansoprazole did not influenced on microcirculation state but markedly activated the regeneration of gastric epithelium. PMID- 12125289 TI - [Characteristics of visual acuity and contrast sensitivity in students of different ages]. AB - To investigate the peculiarities of the visual acuity and contrastive sensitivity 60 healthy pupils (120 eyes) of different ages (7-16) were examined with a help of the computer diagnostic program "Oculus-W" V95. The increase in both the visual acuity and the contrastive sensitivity with age have been shown, as well as the parametres of their normal levels for different ages have been determined. It makes estimating the functional state of the visual analyzer more exact. The processes of making the visual acuity and the contrast sensitivity in the schoolchildren have been proved to interdependent. PMID- 12125290 TI - [Comparative study of electrophysiological properties of arterial endothelial cells]. AB - The electrophysiological properties of intact endothelial cells of isolated arteries from mice, rats, guinea pigs and rabbits have been studied. Endothelial cells in all studied arteries were electrically coupled and had similar membrane potentials. The electrical responses to acetylcholine and ATP in the arteries of both elastic and muscular type were qualitatively similar, but varied among species in the length and amplitude of different stages of the responses. PMID- 12125291 TI - [Effect of polarized light on the immune status and cytokine levels of patients with bronchial asthma during immunotherapy with bronchomunal]. AB - The influence of polarized polychromatic light on immunocompetent cells in complex with immunomodulated bronchomunal is studied. Data of content of the main cytokines taking part in development of inflammation are presented. It is cleared up that polarized light increases the number of T-lymphocyties, normalizes ratio of subpopulation of T-lymphocyties and level of serum FNO-alpha and level of interleukin-4 reaches the level of healthy people. It is ascertained that complex use bronchomunal and polarized polychromatic increases level of serum interferon gamma. PMID- 12125292 TI - [Effect of anosmia on sex-related differences in conditioned avoidance in rats]. AB - An influence of peripheral anosmia on sex-related differences in training and maintaining the conditioned avoidance (CA) behaviour in a shuttlle box was studied. Intact males have been shown to manifest conditioned behaviour earlier than intact females, whereas the degree of CA maintenance did not depend on an animal's sex. Prolonged anosmia facilitated training of conditional avoidance in both males and females, but it reversed the ratio of the amount of intersignal responses at anosmatic males to females versus the intact ones. Observed effects of anosmia are supposed to be realized due to the appropriate changes in the functional activity of the nuclei of the amygdaloid complex. PMID- 12125293 TI - [Relationship between human emotional status and parameters of central hemodynamics during physical and psychological stress]. AB - Changes in central hemodynamics (CHD) to postural changes, psychoemotional stress (mental arithmetic subtraction test) and combined isometric and psychoemotional stress were studied in 21 healthy males aged 18-23, and correlation between CHD parameters and 23 psyhoemotional traits was determined. Negative correlation of the arterial pressure level with parametres of physical aggression, the total level of aggression, and rigidity, as well as its positive correlation with the excitation processes in cerebral cortex have been found. Cardiac output index just after postural loading and at the combined stress correlated with sense of fault. Heart rate at isometric loading correlated with the sense of fault, either. Thus, the arterial pressure is the parameter of CHD that correlates with the human psychoemotional status most closely. PMID- 12125294 TI - Working with HIV/AIDS sufferers: "when good enough is not enough". AB - The authors begin by examining the intrapsychic implications that HIV/AIDS presents after knowledge of infection. Using examples drawn from two cases, they explore how knowledge of infection precipitates an insidious traumatizing process that comprises a number of key defensive strategies and dynamic processes. Particular kinds of defensive splitting, projective dynamics, and key identifications, as well as the collapse of the symbolic function, are isolated as being central to understanding the traumatizing process. With this in mind, the role and aim of the insight-oriented therapist is considered. The authors argue that much of the therapeutic work in this area revolves around a central organizing fantasy about the limitations of "good enough" objects in helping them with their diagnosis and its implications. This is linked to a number of technical dilemmas that the therapist will inevitability have to face if he or she chooses to work analytically. Particular technical problems explored include: 1. the management of frame deviations, 2. the therapist's role/s, 3. the use of interpretation, and 4. countertransference experience and enactment. PMID- 12125295 TI - Overcoming therapeutic pessimism in hypochondriasis. AB - The treatment of hypochondriasis continues to be challenging because of the nature of hypochondriasis and reactions that it often elicits in therapists. In this paper, it is argued that due to complex interactions between hypochondriacal patients and their therapists, there should be a radically different approach to treating hypochondriacal patients. Within such a framework, it is crucial to determine whether therapists are "eligible" to treat hypochondriacal patients, rather than whether hypochondriacal patients fulfill criteria for particular types of treatment. Regardless of the specific treatment modality used, it is suggested that treatment of hypochondriasis should always entail empathy, acceptance, understanding, and explanation and education. In addition, good therapeutic results can be achieved and therapeutic pessimism about hypochondriasis overcome if therapists are accessible, reliable, and flexible, if their behavior is consistent and predictable, and if they express themselves clearly and in unequivocal terms. PMID- 12125296 TI - The secret that guilty confessions fail to disclose. PMID- 12125297 TI - Under the influence of unconscious process: countertransference in the treatment of PTSD and substance abuse in women. AB - PTSD and addiction are a marriage made in the avoidance of unbearable affect; an avoidance that is costly in the resulting traumatic reenactments experienced by patients whose attempts to escape the past keep them evermore tightly bound to it. Rather than "difficult patients" a more dynamic and intersubjective conceptualization emphasizes the notion of a "difficult treatment dyad." Vicarious traumatization, unconscious affects about addiction, and pressures within the treatment surround conspire to pull the therapist out of connection with the patient at critical points, and toward sadistic abandonment or collusive indulgence. The concomitant desires to rescue and desert patients create forces for action in the therapist, precisely when what is needed most is the ability to tolerate and contain one's own and the patient's affective experience. The pull for action is also felt by treatment systems, eager for "action" that can be measured in "behavioral observables." Support for the therapist in the form of process supervision can assist the therapist to contain, identify, and acknowledge his/her affective responses evoked in treatment. The therapist is called upon to "grow one's own heart" through a confrontation with the undeveloped parts of self that are vulnerable to the dynamics of the treatment. PMID- 12125298 TI - Complementary dreams: a window to the subconscious processes of countertransference and subjectivity. AB - The British psychoanalysts were the first to be interested in reciprocal and interpersonal interactions of psychotherapy. The Freudian mirror model was progressively questioned in the 1940s and 1950s. Throughout the 1950s, positions and terms were created that either defended or attacked the use of subjectivity and countertransference in psychodynamic psychotherapy. The objective of this article is to discuss the participation of the therapist's subconscious mind, as it is involved in communication with the patient's subconscious mind during psychodynamic treatment. Specifically, this takes the form of complementary dreams, a clinical phenomenon that I will describe as secondary to the therapist identification with the patient infantile object relations. Complementary dreams will be discussed as a helpful therapeutic tool used to understand the subjective communication that happened between patient and therapist in two separate cases. Complementary dreams will be presented as a helpful therapeutic instrument in containing countertransference enactment. PMID- 12125299 TI - Intensive Short-term Dynamic Psychotherapy in a private psychiatric office: clinical and cost effectiveness. AB - OBJECTIVE: To assess the cost and clinical effectiveness of psychiatrist-provided Intensive Short-term Dynamic Psychotherapy (ISTDP) of patients referred to a private office. METHODS: ISTDP was provided to 89 patients referred to a private psychiatric office in Vancouver, Canada. Pretherapy self-report scores, (BDI, BAI, BSI, IIP [see full names in text]) medication costs, disability costs and healthcare costs were compared with posttherapy values and normative values. RESULTS: Patients' mean self-report scores went from the abnormal to normal range after an average of 14.9 hours of therapy. Returns to work, reduced healthcare utilization, and medication stopping accounted for a cost reduction of over Cdn$400,000 at one year after therapy. CONCLUSIONS: ISTDP appears to be an effective and cost-effective form of intervention when provided by a psychiatrist in a private office. Randomized controlled studies are warranted to further examine the cost benefits and efficacy of ISTDP. PMID- 12125300 TI - Content analysis of social phobics' discourse in cognitive-behavioral therapy. AB - The present study utilized an empirically derived coding system to identify content categories in the spontaneous verbalizations of social phobics in the context of a short-term cognitive-behavioral group therapy. The coding system was applied successfully to segments of transcriptions of eleven group sessions. Evaluation of changes in the content categories utilized by two subjects over the course of the life of the group indicates the usefulness of this coding system in identifying meaningful shifts in cognitive processes during treatment. PMID- 12125301 TI - Iatrogenic symptoms in psychotherapy. A theoretical exploration of the potential impact of labels, language, and belief systems. AB - Although the mental health professions are effective in ameliorating personal distress, treatment can sometimes have negative consequences. The authors explore causal mechanisms for iatrogenic symptoms in therapy by discussing the processes by which clients may be socialized into therapy and the potential impact that psychiatric labels and language may have in influencing clients' self perceptions. The authors review research that has examined possible negative effects of psychiatric labels and then examine other forms of language, categorization, and conceptualizations that may contribute to negative effects in therapy. Iatrogenic symptoms may originate through the over-reliance on a belief system within which therapists interpret, reinterpret, or label clients' characteristics or distress as pathological. Therapeutic communication that emphasizes pejorative language may introduce clients to this belief system. Iatrogenic symptoms may also provide clients and therapists with secondary gains. Possible approaches for minimizing iatrogenic symptoms are explored. PMID- 12125303 TI - Psychotherapy of children with pervasive developmental disorders. AB - In recent years, the role of psychotherapy in the treatment of children with pervasive developmental disorders, such as autism and Asperger's Syndrome, has been questioned. Advances in neuropsychiatry, neuropsychology, and genetics, as well as the refinement of behavioral and educational techniques, have relegated discussions about psychotherapy mostly to reviews about the history of treatment for autism. Even when psychotherapeutic work is suggested, it is typically only very structured supportive counseling for older and high functioning children and adolescents that is considered. This paper argues that there is a central role for psychotherapy for children with pervasive developmental disorders, and that psychotherapeutic interventions should begin at a younger, rather than older age. PMID- 12125302 TI - Why no sex? PMID- 12125304 TI - [Pain perception: a dynamic process...]. PMID- 12125305 TI - [Pain and the immune system: friend or foe?]. AB - When tissue is destroyed, pain arises. Tissue destruction as well as wound healing are associated with an inflammatory reaction. This leads to activation of nociceptors ("pain receptors") which can cross-communicate with the inflammatory infiltrate. The following review will concentrate on pain-exaggerating (hyperalgesic) and pain-ameliorating (analgesic) mediators which arise from immune cells or the circulation during the inflammation. In the early stages of inflammation endogenous hyperalgesic mediators are produced, including the proinflammatory cytokines IL-1, IL-6 and TNF-alpha, nerve growth factor as well as bradykinin and prostaglandins. Simultaneously, analgesic mechanisms are activated. Opioid peptides such as endorphins, enkephalins and dynorphins are produced by immune cells and can be released locally in the inflamed tissue on stimulation with IL-1 or corticotropin releasing factor. Analgesia is elicited by binding of the opioid peptides to receptors on peripheral sensory neurons. During the course of an inflammatory process, peripheral opioid-mediated analgesia increases. In parallel, antiinflammatory cytokines such as IL-4, IL-10, IL-13 and IL-1ra are produced and reduce hyperalgesic effects of the proinflammatory cytokines initially produced. Inflammatory pain, therefore, is the result of an interplay between hyperalgesic and analgesic mediators. Drugs such as immunosuppressants influencing this interplay may also impair endogenous hyperalgesic and analgesic mechanisms. PMID- 12125306 TI - Perioperative monitoring of indocyanine green clearance and plasma disappearance rate in patients undergoing liver transplantation. AB - INTRODUCTION: Indocyanine green (ICG) elimination tests have been repeatedly suggested as an early predictor of graft function in patients with liver transplantation. Conventionally, ICG clearance (ClICG) is measured by a series of blood samples with subsequent laboratory analysis. More recently bedside techniques have become available to measure ICG concentrations in vivo and in addition to ClICG, the plasma disappearance rate of ICG (PDRICG) is increasingly being used. The aim of this study was to assess and to compare the normal time courses of ClICG and PDRICG in liver transplant recipients. METHODS: ClICG and PDRICG were measured perioperatively and at various times up to 24 h after liver transplantation. The bedside transpulmonary indicator dilution technique with an arterial fiberoptic-thermistor catheter was used to assess the ICG concentration time curve together with total circulating blood volume (Vd circ). RESULTS: Similar patterns of the time courses of ClICG and PDRICG with a fast recovery of ICG elimination in the early reperfusion period were observed. Compared to healthy subjects, ClICG was supranormal and PDRICG was slightly subnormal. In this study, Vd circ was increased at baseline and remained increased during surgery. CONCLUSIONS: PDRICG and ClICG are well suited to monitor onset and maintenance of graft function in patients undergoing liver transplantation. The PDRICG values measured tend to be relatively lower than ClICG because of an increased blood volume in these patients. By knowing these differences it is justified to monitor liver function in a very simple manner with PDRICG. PMID- 12125307 TI - [Equipment of physician-staffed ambulance systems in the state of Baden Wuerttemberg]. AB - BACKGROUND: Laws regulating emergency medical systems in the federal state of Baden-Wuerttemberg call for equipment of physician-staffed ambulances that is based on current knowledge in emergency medicine. The grade of implementation is determined using single issue complexes. METHODS: A total of 127 emergency physician bases were located and each received a questionnaire regarding the equipment of the physician-staffed ambulances and helicopters and planned supplementation of the equipment, deadline was 30 June 2001. RESULTS: Of the 127 stations 116 (91.3%) participated. A 12-lead ECG is available in 52.6%, out-of hospital fibrinolysis is possible in 15 bases (12.9%). Alternatives to endotracheal intubation are carried in 53.3% (cricothyroidotomy: 83.3%) and 31 bases provide capnometry or other devices for verifying correct tube placement. A mobile phone is available in 88 bases (75.9%). CONCLUSIONS: When comparing equipment of physician-staffed ambulances statewide, striking differences can be found. PMID- 12125308 TI - [Neuromuscular blockade with cisatracurium in infants andchildren. Its course under sevoflurane anesthesia]. AB - OBJECTIVE: To compare the onset, duration and maximum effect of 0.1 mg/kg cisatracurium during balanced anesthesia with sevoflurane and remifentanil between infants and children. METHODS: We measured the time course of the neuromuscular blockade in 15 infants and 15 children by electromyography. Anesthesia was induced with propofol/remifentanil and maintained with sevoflurane (constant 2% endtidal) and remifentanil according to the patients individual requirements. After injection of 0.1 mg/kg cisatracurium we measured the following parameters: onset time: time between the beginning of injection of cisatracurium and maximum T1 depression, clinical duration: time between injection of the drug and recovery of T1 to 25%, recovery index: time between recovery of T1 from 25% to 75%. TOFR 0.9: time between injection of cisatracurium and recovery of the train-of-four ratio to 90%. In addition, we determined the maximum neuromuscular blockade Tmax after 0.1 mg/kg cistracurium. RESULTS: Both groups differed significantly with regard to onset time and clinical duration. In the infants, the onset time was shorter (74 s vs. 198 s) and the clinical duration longer (55 min vs. 41 min) compared to the older children. The TOFR 0.9 was 73 min (range 56-86 min) in the group of the infants and 59 min (range 43-72 min) in the group of the older children (p < 0.001). Tmax was 100% (range 97 100%) in the infants and 98% (range 92-100%) in the children (p < 0.01). However, the recovery index was comparable in both groups (21 vs. 16 min). CONCLUSIONS: Infants are substantially more sensitive to cisatracurium than children, which can be demonstrated in a significantly shorter onset time, a prolonged clinical duration and a delayed neuromuscular recovery. As there exist large interindividual differences, we recommend the use of neuromuscular monitoring in the routine practice of pediatric anesthesia. PMID- 12125309 TI - [40 years after the last polio epidemic. Postpolio syndrome as a cause of "weaning failure"]. AB - About 80,000 polio survivors are still living 40 years after the last polio epidemics in Germany. Of these 40-70% have developed the so called post-polio syndrome (PPS) decades after the infection. The main symptoms of PPS are decreasing strength in voluntary muscles, pain and fatigue which occur spontaneously but may also be induced by physical stress and general illness. We report the case of a 79-year-old male who developed hypercapnia due to ventilatory failure which necessitated reintubation several times after cholecystectomy. The medical history revealed that he had had poliomyelitis at the age of 8 years. There was only a slight residual handicap from this infection which included mild pareses of the left limbs but had remained stable for about 70 years. Electromyography revealed signs of chronic neurogenic changes in muscles of the left upper limb as well as in the pectoralis major. The diagnostic criteria of a post-polio syndrome were fulfilled and other neuromuscular diseases were excluded. The patient could be discharged from intensive care only after treatment by intermittent positive pressure ventilation via a facial mask. This case report shows that even patients who have a mild handicap after poliomyelitis can develop weaning problems. A PPS can exacerbate with inclusion of respiratory muscles in critically ill patients. PMID- 12125310 TI - [Emergency management of an acute rupture of a perforated thoracic aortic aneurysm]. AB - The emergency medical service was called to a 60-year-old woman with intensive chest pain, signs of shock, dyspnoea, intermittent paraesthesia of the right leg and disturbance of consciousness. With the diagnosis of an acute rupture of an aneurysm of the thoracic aorta, the patient was stabilised with volume, catecholamines, intubation and mechanical ventilation before being rushed to the preinformed department of cardiovascular surgery. The diagnosis was verified by transesophageal echocardiography immediately and the patient underwent surgery 2 h after onset of symptoms. Despite the rupture of the aorta and a short period of cardiac arrest, the patient recovered totally and could be discharged without any residual problems. This case shows that a ruptured thoracic aortic aneurysm can be survived although the overall mortality of this incident is more than 97%. The essentials of a good outcome are: 1. perfectly coordinated rescue operation which means an emergency medical service which includes the rupture of an aortic aneurysm in the differential diagnosis of acute chest pain, 2. an early verification of the diagnosis by means of transesophageal echocardiography which should also be carried out by anaesthesiologists due to its importance in the differential diagnosis in haemodynamic unstable patients. PMID- 12125311 TI - [Recombinant factor VIIa (NovoSeven). A review of current and possible future indications]. AB - Recombinant activated coagulation factor VII (rFVIIa, NovoSeven) was originally developed for the treatment of bleeding complications in haemophilia patients with allo-antibodies (inhibitors) against exogenous factor VIII or IX. In 1988, rFVIIa was used successfully in such patients for the first time. Subsequently, the efficacy and safety of rFVIIa in haemophilia patients with inhibitors has been proven in several prospective trials. A large number of case reports and results from initial clinical trials suggest that rFVIIa may also be effective in the prevention and treatment of bleeding in patients under oral anticoagulation, with liver diseases, and in patients without any pre-existing haemorrhagic diathesis. However, further clinical studies will be necessary to specify the future potential of rFVIIa. PMID- 12125312 TI - [Guidelines of the European Resuscitation Council 2000 for basic pediatric life support. A statement of the Pediatric Life Support Working Group following approval by the executive committee of the European Resuscitation Council]. PMID- 12125313 TI - [Guidelines of the European Resuscitation Council 2000 for advanced pediatric life support. A statement of the Pediatric Life Support Working Group following approval by the executive committee of the European Resuscitation Council]. PMID- 12125314 TI - [Placental passage of anesthetics and adjuvants]. AB - In obstetric anaesthesia almost all anaesthetic agents are capable of traversing the fetomaternal blood barrier. They all carry potential side-effects putting the unborn or newborn child at risk. Of major relevance is their potential for embryotoxicity, teratogenicity, postpartal cardiorespiratory or neuromuscular depression and the disturbance of thermoregulation. This can possibly lead to fetal malformation, asphyxia or floppy infant syndrome. Furthermore compromisation of uterine blood flow or contractility of the mature uterus plays an important role for the incidence of intrauterine asphyxia and premature labour or birth. Considering the physiological and pathophysiological alterations during pregnancy regarding all organ systems, the overall goal is to find an ideal choice of anaesthetic drugs and techniques in order to minimise an increased anaesthetic risk during pregnancy. PMID- 12125315 TI - [General H1-H2 blockade for anesthetic induction]. PMID- 12125316 TI - [Congenital coagulopathy caused by missing coagulation factors]. PMID- 12125317 TI - [Retransfusion of autologous blood]. PMID- 12125318 TI - [Care-based learning in continuing medical training: its use in anesthesiology, emergency and intensive care medicine]. PMID- 12125319 TI - [Acute pulmonary artery embolism]. PMID- 12125320 TI - What can I do to ensure I work in a safe and healthy environment? PMID- 12125321 TI - [Parathyroid gland, calcium receptor and calcimimetics]. AB - Parathyroid glands, bones, intestines and kidneys closely regulate ionized extracellular calcium concentration and thereby bone mineral density. This regulation is accomplished by a membrane associated receptor that responds to small changes in the extracellular calcium concentration (Ca2+)ec. The activation of this receptor regulates the secretion of PTH and the urinary excretion of calcium. The cloning of this calcium sensing receptor (RCa) in 1993 has allowed the identification of several hereditary disorders characterized by either a hyperparathyroidism or a hypoparathyroidism, as well as the development of pharmaceutical compounds potentially of interest for the treatment of these diseases. The calcimimetics are able either to directly stimulate the RCa or to make the RCa more sensitive to the effects of (Ca2+)ec. By this way, they decrease the secretion of PTH, but they also stimulate the secretion of calcitonin. The first clinical studies with a first-generation calcimimetic have demonstrated their efficacy lowering plasma intact PTH concentration in uremic patients with secondary hyperparathyroidism. However, the low biodisponibility of these first calcimimetics predict a difficult clinical utilization. The preliminary results obtained with a second-generation calcimimetic, the AMG-073, are very promising and await long term evaluation. The goal of this manuscript is to review the physiological, biological and clinical bases for the development of calcimimetic. Moreover, to try to replace the potential use of these drugs in the context of chronic renal failure and in the treatment of secondary hyperparathyroidism. PMID- 12125322 TI - [Hemolytic and uremic syndrome in the adult]. AB - Hemolytic-uremic syndrome (HUS) is a rare disease but may be fatal in the absence of appropriate treatment. It is the first cause of acute renal failure (ARF) in children and is less frequently observed in adults. We report the case of a 50 years old woman who was first admitted in the surgery department for intestinal occlusion. Haematological and renal disorders were not recognised initially. Specific treatment with plasma exchanges was beneficial. This case was idiopathic but in 50% of HUS in adults, an aetiology can be found and sometimes treated: drugs, pregnancy, systemic diseases, hypertension or cancer. Although still debated, treatment is based on fresh frozen plasma infusion, associated or not to plasma exchanges. Some other therapies can be added: steroids, polyvalent immunoglobulin infusion, vincristin or splenectomy. In all cases, blood pressure control must be obtained, especially with ACE inhibitors. Response to treatment is often excellent, but chronic renal failure persists in about 25% of cases in adults. Recurrences are rare and occur mainly in patients with an hereditary factor H deficiency. The hemolytic-uremic syndrome should be diagnosed and treated immediately. PMID- 12125323 TI - [Compensatory hyperfunction in living kidney donors]. AB - Renal transplantation using living donors still remains of interest, given the shortage of cadaveric donors. Using reference methods for measuring kidney function, we studied the adaptation to nephrectomy in 99 living donors. The glomerular filtration rate and renal plasma flow showed long lasting increase (by 40 and 33% respectively). Age and the glomerular filtration rate at surgery had a clear-cut effect on these changes. The spontaneous changes in protein intake further influence the value of post-nephrectomy glomerular filtration rate. The analysis of serial changes in serum creatinine or creatinine clearance would falsely have suggested a late increase in renal function. Microalbuminuria increased in few patients, pointing to the need for careful long term follow-up of such donors. PMID- 12125324 TI - [Epidemiology of end-stage renal failure: comparison between 2 Swiss cantons]. AB - Since a previous survey in the Valais Canton had shown a disparity in the prevalence of chronic dialysis patients compared to the national mean, an epidemiological survey was conducted in the cantons of Valais (VS) and Vaud (VD) among all the dialysis units (VS 6, VD 6) and all adult dialysis patients (VS 130, VD 187). The survey confirms the disparity in prevalence: VS 476, VD 340 and Switzerland 329 per million inhabitants (p.m.h.). This disparity also exists among different parts of the VS canton: Haut-Valais 316, Valais Central 650 and Bas-Valais 409 per million d'habitants. The mean age at start of therapy and at the time of the survey are comparable: 57 +/- 17 vs 55 +/- 15 years and 62 +/- 15 vs 61 +/- 14 respectively. Among the different etiological diseases, vascular nephropathies are less frequent and hereditary diseases more frequent in the VS canton. Co-morbidity factors were comparable. In VS there are less patients on peritoneal dialysis (6 vs 19%) and on the transplant waiting list (12 vs 16%). Globally, disparities exist in the prevalence of dialysis patients between the 2 cantons; they can be explained by both medical factors (type of nephropathies) and different options in the treatment modalities. PMID- 12125325 TI - [Intraregional disparities in Reunion]. PMID- 12125326 TI - Evidence-based practice in dentistry. AB - The importance of evidence in teaching and in support of clinical decisions is well established in health care, including dentistry. Defence of clinical decisions increasingly requires reliable data or evidence to support the stance taken. Assistance in finding the best evidence comes from a variety of sources, including computerised databases, journals, continuing education meetings, and study clubs. The randomised controlled trial heads the hierarchy of research designs on which evaluation of evidence is based; anecdotally based evidence and individual case studies are the least preferred study designs. Evaluation of a study requires a number of questions to be asked to determine how the study was performed, and whether it applies to a clinical situation. These questions relate to how the study was carried out, whether controls were used, were the results likely to be valid, and was statistical and clinical significance present. Quackery, pressure from consumers, and legal considerations have contributed to an increase in the importance of evidence-based practice. The benefits, however, of evidence-based practice are that treatment decisions are easier to justify, especially when there is a complaint or a dento-legal issue, and the personal satisfaction that patients are being offered the best treatment. PMID- 12125327 TI - Antifungal drug susceptibilities of commensal Candida isolates. AB - Two hundred and forty-two oral commensal yeast isolates were obtained from a convenience sample of 134 healthy 7- and 8-year-old children (65 males and 69 females). The isolates were initially tested for their susceptibilities to the antifungal azole drug fluconazole, using an agar diffusion method (Etest), which was suitable for screening large numbers of yeast isolates, and confirmed as equivalent to the broth microdilution reference method. Eighteen isolates from 7 children were found to have low fluconazole susceptibility according to guidelines published by the United States National Committee for Clinical Laboratory Standards (NCCLS). The isolates with low susceptibility were identified as either Candida tropicalis (n = 9 of 34 strains tested) or Candida glabrata (n = 9 of 13 strains tested). Selected isolates (6 susceptible and 7 with lower susceptibility to fluconazole) were also tested by a reference broth microdilution method for susceptibility to fluconazole and to a related over-the counter azole antifungal, miconazole. A positive correlation between susceptibility to fluconazole and to miconazole was observed. The high rate (38 percent) of reduced susceptibility in commensal C tropicalis and C glabrata strains may represent a future treatment problem if the use of over-the-counter azole drugs increase. PMID- 12125328 TI - Betel chewing creeps into the New World. AB - Areca nut has been chewed either alone or as a component of the betel quid since ancient times. It has been estimated that more than 10 percent of the world's population chew it for its mild psychoactive effects. Betel is chewed in New Zealand and Australia by immigrants from India now dwelling in these countries. Various forms of areca nut preparations are available in Asian groceries throughout New Zealand. The regular use of betel will, in time, stain the mucosa, gums, and teeth. This habit is discouraged in many countries because of its oncogenic, addictive, and dysaesthetic properties. Dentists and other health professionals should be aware of the effects of this habit. PMID- 12125329 TI - Xerostomia and you. AB - The aims of management of patients with xerostomia include the provision of a definitive aetiology and diagnosis, and a prognosis for the salivary dysfunction. Salivary flow assessment and function testing, other relevant laboratory tests, and even referrals should be performed as necessary. Management of the condition depends on diagnosis and severity, and includes enhancement of salivary flow, oral lubrication, control of soft-tissue infections and discomfort, and prevention of dental caries. PMID- 12125330 TI - [General surgery in Europe. General surgery--specialized surgery]. PMID- 12125331 TI - Does Switzerland train tomorrow's surgeons today? Today's reflections on tomorrow's surgeons. AB - Various factors are influencing the Swiss health care system, and therefore, surgeons' actual and future profession. Initiated by the current president of the Swiss Society of Surgery, a group of eight young Swiss surgeons constituted the ad hoc committee "Chirurgie 2020". The goal was to develop several scenarios of the surgeon's professional situation in 20 years. The future direction of surgery will be markedly influenced by medical innovation, political and economic relationships between Switzerland and Europe. However, the development of health care costs represents the most powerful factor predicting all further changes. The current situation of today's surgeons is best characterised as "hamsters in the treadmill" who try to fulfill a multitude of diverging tasks causing major demotivation. In order to retain a leading role in the health care system, surgeons must take part in social and political activities. To provide an excellent curriculum, university and county hospitals must join together to form clinical and educational networks. A clearly defined and structured curriculum must be introduced by the Swiss Society of Surgery. From our current point of view, only everyone's strong personal commitment will help to find solutions and to improve the current and future situation. In particular, today's surgical residents must actively take part in the development of tomorrow's surgery. PMID- 12125332 TI - [The learning curve in surgery: possibilities and limits of this method]. AB - The learning curve is a graphic representation of the relationship between the experience of a surgeon and one or more performance indicators. The operation time alone is an insufficient indicator to assess the performance of a surgeon. The procedure time has to be set in relation to the complication rate and in laparoscopic surgery to the conversion rate. Techniques to visualize the changes over time are the moving average method for the operation time and the Cusum method for dichotomous outcomes, like the conversion and the complication rates. At the time when the learning curve reaches the plateau phase this representation can be used to assess the quality of a surgeon or a team in one hospital. As far as there is no validated complexity scale for laparoscopic procedures available it is difficult to compare patient populations between different hospitals. Out of this reason the learning curve is no legitimate instrument to rank surgeons or different hospitals. PMID- 12125333 TI - Helicobacter pylori infection in patients undergoing appendectomy. AB - AIMS: Helicobacter pylori has been found in the upper gastrointestinal tract; it is incriminated as aetiological factor in various pathological conditions. This prospective study assesses the presence of this microorganism in the appendix flora and the possible role of its infection in the pathogenesis of acute appendicitis. METHODS: H. pylori was investigated in 46 consecutive patients undergoing emergent appendectomy for presumed acute appendicitis. Blood sample for serological test of H. pylori infection was drawn before operation. The removed appendix specimen was stained for H. pylori; confirmation was made by PCR (Polymerase Chain Reaction) analysis. The intensity of inflammation was determined pathologically grading from no inflammation to gangrenous appendicitis. Statistical analysis was made using the chi-square test. RESULTS: Seropositivity for H. pylori infection was found in 18 patients (39%), but the microbe was detected in just two appendix specimens (4%). In all seropositive patients acute appendicitis was confirmed by the pathology study; serous (33%) and purulent or gangrenous (67%). The latter incidence in the seronegative patients was 50%. There were found eight specimens (17%) negative for inflammation dealing all with seronegative patients. CONCLUSIONS: It seems that H. pylori colonizes the appendix in small proportion and is unlikely to be associated in direct correlation with acute appendicitis. However, seropositive patients with acute inflammation are likely to suffer from purulent or gangrenous form. PMID- 12125334 TI - Medial meniscectomy in patients over the age of fifty: a six year follow-up study. AB - PURPOSE: Meniscectomy in the older patient remains a controversial topic. The aim of our study is to assess the mid-term outcome of arthroscopic partial medial meniscectomy in patients over fifty years of age and attempt to retrospectively identify symptoms and/or findings on examination which can differentiate between non-degenerative medial meniscal tears versus degenerative meniscal changes. MATERIALS AND METHODS: Thirty-two patients over the age of fifty who had undergone arthroscopic medial partial meniscectomy, were reviewed. The average age was 60 (51-74 yrs) and the average follow-up was six years (3-7 yrs). Based upon the intra-operative findings, patients were divided into two groups: (1) non degenerative meniscal tears (NDM; n = 12) and (2) degenerative meniscal changes (DM; n = 20). Our outcome measurements were with the HSS knee score, a satisfaction score, and weight-bearing X-rays. RESULTS: In the NDM group, eleven patients were rated excellent or good, and one was rated poor. In the DM group, three patients were rated as excellent or good, eight as fair, and nine as poor. The HSS score was 97 +/- 4.6 for the NDM group and 85 +/- 9.5 for the DM group. The average satisfaction score was 9.2 +/- 0.7 (very satisfied) for the NDM group and 5.8 +/- 2.6 (fairly satisfied) for the DM group. There was no significant difference between the NDM and the DM groups with regards to pre-operative symptoms and signs, except for the McMurray sign, which was found to be positive in 83% of NDM cases versus 25% of DM cases (sensitivity = 83%). Using only these data, the McMurray sign was 67% specific for NDM. CONCLUSIONS: Arthroscopic medial meniscectomy in older patients provides 90% good results six years after a non-degenerative meniscal tear, but only 20% of good results after a degenerative meniscal tear. However, based on this study, neither symptoms nor physical examination are able to differentiate between traumatic meniscal tears and degenerative meniscal changes in older patients. A positive McMurray's sign favors the diagnosis of a traumatic tear. However, a specificity of this test of only 67% as shown in our data questions its utility in clinical decision-making. PMID- 12125335 TI - Schistosomal appendicitis in pregnancy. AB - Acute appendicitis is the most common acute surgical infection during pregnancy. Although usually pyogenic in origin, parasitic infections account for a small percentage of cases. Despite the relatively high prevalence of acute appendicitis in our environment, it is not commonly associated with schistosomiasis. We report here the association of pregnancy and appendicitis caused by Schistosoma haematobium. Schistosomiasis is very common complication of pregnancy in hyperendemic areas. Schistosome egg masses can lodge throughout the body and cause acute inflammation of the appendix, liver and spleen. Congestion of pelvic vessels during pregnancy facilitates passage of eggs into the villi and intervillous spaces, causing an inflammatory reaction. Tourism and immigration make this disease a potential challenge for practitioners everywhere. PMID- 12125337 TI - [The clinician in surgery of tomorrow]. PMID- 12125336 TI - [Craniocerebral trauma--new pathophysiological and therapeutic viewpoints. 1: Management of the acute phase]. AB - AIM: As there is only few evidence-based knowledge in the treatment of head trauma, the aim of this work was to demonstrate the close relationship between pathophysiology and clinical treatment modalities during the primary phase of head trauma. METHODS: Review article. DISCUSSION: Cerebral hypoxia represents the main source of secondary brain injury. Depending on severity and duration, there results an irreversible brain damage. Under clinical conditions, the continuous monitoring of cerebral perfusion pressure (CPP) allows an estimation of the risk for cerebral hypoxia and the beginning of causal therapy. The priority of treatment lies in normalization of arterial O2 and increase in cerebral blood flow (CBF). In case of unchanged autoregulation of CBF, a decrease of CPP is followed by vasodilation with increase of cerebral blood volume (CBV) and a increase of CPP leads to a decrease of CBV by vasoconstriction. The usual shift of autoregulation on the right side after trauma requires CPP-values > 70 mmHg in the initial posttraumatic phase as there exists the most probable risk of hypoxia. The therapy of elevated ICP > 20 mmHg is oriented on CPP. A decrease in ICP under load of CPP has to be avoided. Beyond it, there is a need of origin of ICP-increase. CONCLUSIONS: The priority of initial treatment after head trauma lies in the individual optimization of CBF by maintenance of adequate CPP and cerebrovascular resistance. Afterwards, there will be performed a goal-directed treatment being based on the differentiation of the varying subgroups of head trauma and their related pathophysiology, what leads to the required treatment strategy. For this reason, it is necessary to put in the traditional treatment option by means of the multimodal monitoring as soon as possible. PMID- 12125338 TI - Does the EBSQ adequately evaluate tomorrow's surgeons? PMID- 12125339 TI - Identification of redox-active proteins on cell surface. PMID- 12125340 TI - Probing redox activity of human breast cells by scanning electrochemical microscopy. PMID- 12125341 TI - Determining influence of oxidants on nuclear transport using digitonin permeabilized cell assay. PMID- 12125342 TI - Functional imaging of mitochondrial redox state. PMID- 12125343 TI - Hydrogen peroxide-induced apoptosis: oxidative or reductive stress? PMID- 12125344 TI - Peroxidation of phosphatidylserine in mechanisms of apoptotic signaling. PMID- 12125345 TI - Quantitative high throughput endothelial cell migration and invasion assay system. AB - We have developed a novel assay system combining fluorescent signal-blocking microporous PET membrane inserts and ECM to study dynamic endothelial cell migration and invasion. The assay described here may be applicable to other cell types for analogous assay. The currently used method for analyzing migration and invasion requires the removal of nonmigratory cells from the top of a clear microporous membrane. This laborious step is required to quantitate the invaded or migrated cells on the bottom of the membrane. When the fluorescent signal blocking the microporous membrane is used, the fluorescent signal from noninvaded or nonmigratory cells from the top of the membrane is virtually eliminated and allows for rapid and efficient measurement of the migrated or invaded cells on the underside of the insert. Unlike conventional low throughput migration assays, this assay is easy to perform and provides a quantitative invasion/migration profile of the chemoattractant of choice. This robust automation compatible assay will serve as a powerful tool to screen potential antiangiogenic compounds for drug discovery. PMID- 12125346 TI - In vitro model of oxidative stress in cortical neurons. PMID- 12125347 TI - Glutamate-induced c-Src activation in neuronal cells. PMID- 12125348 TI - Measurement of inflammatory properties of fatty acids in human endothelial cells. PMID- 12125349 TI - Redox control of tissue factor expression in smooth muscle cells and other vascular cells. PMID- 12125350 TI - Redox processes regulate intestinal lamina propria T lymphocytes. PMID- 12125351 TI - Linker for activation of T cells: sensing redox imbalance. PMID- 12125352 TI - Generation of prooxidant conditions in intact cells to induce modifications of cell cycle regulatory proteins. PMID- 12125353 TI - Analysis of transmembrane redox reactions: interaction of intra- and extracellular ascorbate species. PMID- 12125354 TI - Regulation of endothelial cell proliferation by nitric oxide. PMID- 12125355 TI - Fluorescent imaging of mitochondrial nitric oxide in living cells. PMID- 12125356 TI - Measurement of absolute oxygen levels in cells and tissues using oxygen sensors and 2-nitroimidazole EF5. AB - We have established basic methods, using quantitative measures of EF5 binding, to estimate the actual pO2 of cells and tissues. In situations where the tissue can be dissociated into single cells, or for cell cultures, we can measure the distribution of cellular binding rates using flow cytometry and these can be compared with cells treated under pO2S controlled by the spinner vial or thin film methods in vitro. The flow cytometer is calibrated by staining V79 cells treated with EF5 under "standard" conditions. For intact tissues treated with EF5 in vivo, we need to correct for possible variations in drug exposure (AUC). Frozen sections are stained for EF5 binding and are analyzed by a sensitive (cooled) CCD camera with linear output vs fluorescence [figure: see text] input. The camera has very consistent sensitivity, but the entire optical system, including the camera, can be calibrated by an absolute fluorescence standard (dye in hemocytometer). This system can also be used to measure the fluorescence of the flow cytometer standards, providing a direct link between the two assays. We can measure the maximum binding rate using the tissue cube method, but need to assume an "average" oxygen dependence of binding for intact tissues. The best-fit approximation for existing data is an inverse relationship between binding and pO2, with binding decreasing 50-fold between 0.1 and 10% oxygen. Using these methods, we routinely estimate the minimum pO2 (maximum binding) in experimental rodent and human tumors. In normal tissue models, an excellent correlation is found between near-maximal binding (severe hypoxia) and apoptosis (heart infarct and ductus arteriosus). Some normal tissues (e.g., skeletal muscle) are refractory to both cellular disaggregation and cube calibration methods. To extend the tissue imaging measurements to a complete two- or three-dimensional analysis of the distribution of tissue pO2s requires a substantial additional investment of imaging methods, which are currently being implemented. PMID- 12125357 TI - Detection of reactive oxygen and nitrogen species in tissues using redox sensitive fluorescent probes. AB - The take-home message of this chapter is that the fluorescent probes for ROS and RNS have great potential in improving our understanding of redox behavior within cells and tissues. However, data obtained from studies using these probes must be expressed in the context of the limitations of the chemistry of the probes in the cellular microenvironment, which may change under different conditions, such as cell stress or injury. In most cases, as suggested, results should be described in a general context of reflecting an increase in oxidizing reactions within the cell and not as a quantitative measure of the production of a specific oxidant species. It is highly recommended that results be verified, when possible, with alternative fluorescent probes or preferably using alternative methods, such as electron spin resonance or other newly emerging technology. PMID- 12125358 TI - Detection of oxygen-sensing properties of mitochondria. PMID- 12125359 TI - Simultaneous detection of tocopherols and tocotrienols in biological samples using HPLC-coulometric electrode array. PMID- 12125360 TI - In vivo measurement of oxidative stress status in human skin. PMID- 12125361 TI - Localization of oxidation-specific epitopes in tissue. PMID- 12125362 TI - Quantitation of S-nitrosothiols in cells and biological fluids. PMID- 12125363 TI - Peroxisomal fatty acid oxidation and cellular redox. PMID- 12125364 TI - Ultrastructural localization and relative quantification of 4-hydroxynonenal modified proteins in tissues and cell compartments. PMID- 12125365 TI - Ultrastructural localization of light-induced lipid peroxides. PMID- 12125366 TI - A survival model for the study of myocardial angiogenesis. PMID- 12125367 TI - Determination of angiogenesis-regulating properties of NO. PMID- 12125368 TI - Direct detection of singlet oxygen via its phosphorescence from cellular and fungal cultures. AB - In summary, we have developed methods to detect 1O2 spectrally from viable keratinocytes and fungal mycelial cultures. Both time-resolved and steady-state spectrophotometers were used, providing complementary information on optimal conditions for the unambiguous detection of 1O2 phosphorescence. By using our techniques, we were able to confirm that photosensitizers in contact with living cells indeed generate 1O2. The model systems and the procedures described can be adopted for other studies involving 1O2 and oxidative stress in living cells. PMID- 12125369 TI - Hemangioma model for in vivo angiogenesis: inducible oxidative stress and MCP-1 expression in EOMA cells. PMID- 12125370 TI - Redox aspects of vascular response to injury. PMID- 12125371 TI - Involvement of superoxide in pathogenic action of mutations that cause Alzheimer's disease. PMID- 12125372 TI - Three-dimensional redox imaging of frozen-quenched brain and other organs. PMID- 12125373 TI - In vivo fluorescent imaging of NADH redox state in brain. PMID- 12125374 TI - Nitroxyl probes for brain research and their application to brain imaging. PMID- 12125375 TI - Analytical implications of iron dithiocarbamates for measurement of nitric oxide. PMID- 12125376 TI - Molecular analysis of mitogen-activated protein kinase signaling pathways induced by reactive oxygen intermediates. PMID- 12125377 TI - Detection of intracellular reactive oxygen species in cultured cells using fluorescent probes. PMID- 12125378 TI - Flow cytometric determination of cytoplasmic oxidants and mitochondrial membrane potential in neuronal cells. PMID- 12125380 TI - Determination of intracellular reactive oxygen species as function of cell density. PMID- 12125379 TI - Flow cytometric determination of lipid peroxidation using fluoresceinated phosphoethanolamine. AB - In conclusion, we describe the use of fluor-DHPE as a flow cytometric probe to assess lipid peroxidation in erythrocytes. The stability and nonexchangeability of this probe make it suitable for monitoring lipid peroxidation in a particular cell type via flow cytometry, which can be done in the presence of other cells. Application of the probe in erythrocytes from rats fed a vitamin E-deficient diet demonstrated a higher susceptibility to lipid peroxidation among these cells than among erythrocytes from rats fed a normal diet. The current flow cytometric lipid peroxidation detection method can be interfaced directly with several standard techniques that are available to measure specific blood cell populations via flow cytometry. PMID- 12125381 TI - [Less medicalization, more prevention]. PMID- 12125382 TI - [The epidemiologist: who is he?]. PMID- 12125383 TI - [The difficult birth of a discipline]. PMID- 12125384 TI - [Schematization and forcing of the analysis of mammographic screening effectiveness]. PMID- 12125386 TI - [The National Guideline Program, 2 years later]. PMID- 12125385 TI - [Globalization and global health: a prologue (1492), 3 acts (XIX and XX centuries) and an uncertain epilogue]. PMID- 12125387 TI - [Mortality in a cohort of asbestos cement workers in Bari]. AB - The study describes the mortality of 417 workers employed in a asbestos-cement plant, located in Bari, Puglia, Southern Italy. Follow up started on February 1st 1972. The vital status and cause of death were ascertained at 1995. The mortality experience of the Apulian population was used as comparison. Using 90% confidence limits (CLs), a significant increase in mortality was observed in our cohort from: all causes of death (SMR 118, CL 100-139), pneumoconiosis (SMR 14810, CL 10298-20683), all types of cancer (SMR 139, CL 105-181), lung (SMR 191, CL 126 277), pleural (SMR 1560 CL 431-4081) and peritoneum (SMR 1705, CL 303-5367) malignant neoplasms. In our cohort, the discrepancy between observed and expected mortality for lung and pleural cancer occurred 30 years after the first exposure, after 40 years for all neoplasms and peritoneum cancer. Under the Cox regression model, lung cancer SMR showed a curvilinear trend along time since first exposure, the peak being detected at 35 years. Finally, SMRs from our cohort were compared to a previously described cohort including workers from the same plant compensated for asbestosis by INAIL. PMID- 12125388 TI - [Mortality study of a cohort of insulation workers]. AB - Cause specific mortality was investigated in a cohort of insulators employed by a company which operated in various parts of Italy. Follow-up covered the years 1960-1996. The cohort, which included 893 subjects, was derived from company files of relatively poor quality, which resulted in a high rate of lost to follow up (10.1%) and of deaths with unknown cause (12.4%). The mortality experience of the cohort was contrasted with that of the Italian population. Overall mortality (SMR 141, 90% CI 118-167, 97 observed), and cancer mortality (SMR 165, 90% CI 123 216, 38 observed) were significantly increased. Among neoplasms, significant increases were observed for lung cancer (SMR 202, 90% CI 124-311, 15 observed), pleural neoplasms (SMR 2667, CI 90% 911-6103, 4 observed), and peritoneal neoplasms (SMR 1853, 90% CI 329-5832, 2 observed). The excess mortality for lung cancer was especially pronounced in subjects with latency time longer than ten years (SMR 237.1, 90% CI 140-377, 13 observed). PMID- 12125389 TI - [Testicular tumors in Italy: historical trends, geographic differences, and etiological hypotheses]. AB - Testicular cancer is a rare tumour. Its incidence has been increasing in many parts of the world during the last decades. This cancer has an unusual age distribution with one peak in incidence in young adults (aged 20-39) and a second peak in over 60. On the basis of the Italian Bureau of Statistics, mortality rate was higher in northern regions of the country compared to southern ones, up to the beginning of the 1980's. However, the progressive decline in mortality in all regions eliminated the geographical differences in recent years. Incidence rates obtained from the existing cancer Registries, confirm a strong geographical gradient, with higher rates in the North (ex. 5.1/100,000 in Parma in 1988-92) than in the South (ex. 1.1/100,000 in Ragusa in 1988-92). The comparison between incidence rates of two recent periods (1983-87 and 1988-92), showed an increase in most of the provinces for which data were available. The age distribution curve showed that testicular cancer occurs after puberty, with a peak in incidence among 30-34 year olds, and after the age of 60. These observations suggest the possible causative role of some environmental exposures. Epidemiological studies have found an association between some occupational exposures in industrial and agricultural settings and testicular cancer. Moreover some studies have found an association between parental occupation and testicular cancer in the offspring. However, the aetiology of testicular cancer is still poorly understood. PMID- 12125391 TI - [Number needed to treat]. PMID- 12125390 TI - [Personality traits of women participating in a breast cancer prevention trial]. AB - We have evaluated the psycho-social factors in women--during menopause with different biological characteristics--who participated in two extensive trials of breast cancer prevention: Diana1 and Tamoxifen. Through the use of a recognized personality test (MMPI, Minnesota Multiphasic Personality Inventory), we observed 500 healthy women who agreed to or refused the health care proposal. The findings show that the women who accept chemical preparations or to modify their dietary habits present different personality traits from those who refuse to adhere. One should ask oneself if the lack of homogeneity of the samples with a different concentration of psycho-social factors can alter the efficacy of a cancer prevention program. During chemoprevention studies, in which a high compliance could bring about a redundancy of experience of sickness, in coherence with our goal of health protection, we think it is necessary to supply psycho-social support which tempers any experience of physical, psychological and inter personal discomfort in the healthy women. The cognitive model of the personality traits could be programmed also for the compliance of mammographical screening. This model requires the training of health care professionals. PMID- 12125392 TI - [Health risk communication: between uncertainty and opportunity]. PMID- 12125393 TI - [Lateral effects of emotional stimulation on verbal memory processes in men and women]. AB - Effects of short-term changes in emotional state of men and women on hemispheric organization of verbal memory processes were studied in the model of emotional electrodermal stimulation. Retrieval of dichotically presented lists of words was analyzed. The lists of words included pairs of emotionally neutral word and pairs combined of a neutral and emotional or both emotional words. There were no gender differences in the immediate retrieval of the words. Electrodermal stimulation prior to presentation of list of word resulted in a lateralized reduction of the immediate report of neutral words presented to the right ear and of emotional words presented to the left ear. Gender-related differences were shown in the delayed retrieval. In women, the stimulation applied before word presentation induced a decrease in delayed retrieval of left-ear emotional words. In men, the stimulation applied after the word presentation improved the retrieval of word lists independently of the side of presentation. Mechanisms of gender-related differences in interhemispheric interaction in verbal memory processes are discussed. PMID- 12125394 TI - [Chaotic component of human high-frequency EEG in the state of quiet wakefulness]. AB - Chaotic component of human EEG oscillations in the high-frequency band (14.7-100 Hz) was investigated. EEG was recorded from four points in symmetrical frontal and occipital scalp areas. The results of the non-linear analysis of the high frequency EEG indicate the existence of the deterministic chaotic component with a high attractor correlation dimension. It was significantly different from the respective values of the Gaussian noise filtered in the same frequency band. In the state of quiet wakefulness (eyes closed), the dimensions of chaotic components of the EEG in all derivations did not differ from each other. Analysis of correlation pairs between the ensembles of correlation dimensions of the high frequency EEG revealed reliable patterns of significant connections between the neocortical areas with individual features in different subjects. When the functional state of the brain was changed by hyperventilation, both the values of the correlation dimensions and the structure of inter-area connection patterns changed. We believe that the nonlinear component of high-frequency EEG is a sensitive and local characteristic of the functional state of the human brain. PMID- 12125395 TI - [Activation of glutamate ionotropic connections of sensorimotor cortex neurons during conditioning]. AB - Changes in conditioned impulse reactions of neurons in sensorimotor cortex were studied during microiontophoretic application of glutamatergic and GABA ergic agonistic and antagonistic drugs. It was shown that ionotropic glutamate receptors (AMPA and NMDA) are activated by a conditioned stimulus. Not only large pyramidal neurons of deep cortical layers but surrounding short-axon inhibitory interneurons are involved in the reaction. It was shown that the activity of pyramidal neurons is under a constant inhibitory control from surrounding interneurons. This inhibition is involved in organization of excitatory cortical responses during conditioning. PMID- 12125396 TI - [Dynamics of reorganization of instrumental food getting behavior of monkey under conditions of reinforcement delay]. AB - Six adult monkeys (Macaca fascicularis) were trained to switch on a lamp by pressing the lever, to hold the lever for not less than 1 s (the lamp being switched on), and get a portion of food. After reaching a learning criterion, the delay between the lever pressing and food reinforcement was increased to 2.5 s. The experimental procedure was repeated in three experimental sessions with 2 month between-session intervals. It was shown that the retraining process after the uneven change in the delay duration developed in three stages: (1) stage of emotional hyperreactivity that reflected a mismatch between the cation and absence of the expected result; (2) stage of intermediate stabilization, when the percent of efficient attempts was the same as under conditions of 1-s delay; (3) stage of purposive instrumental lever holding till the moment of reinforcement presentation. PMID- 12125397 TI - [Activity of neurons in the pedunculopontine nucleus during defensive instrumental conditioning]. AB - The activity of 109 neurons in the compact and diffuse parts of the pedunculopontine tegmental nucleus (PPTg) was recorded in freely moving rabbits during the acquisition and actualization of the defensive instrumental conditioned reflex. It was found that 47% of the recorded neurons responded to a conditioned stimulus (CS). This finding suggests the involvement of the PPTg in the instrumental conditioning. A significant prevalence of the excitatory conditioned responses to the CS suggests the predominantly activating influence of the PPTg on its projection structures during conditioning. The neuronal responses to the CS were classified into several basic patterns reflecting stimulus effects, the structure of the behavioral act, and the nature of the reinforcement. They indicated the involvement of the PPTg in attention, motor learning, and reinforcement. A significant decrease in the reactivity of the CS of the PPTg neurons as a result of learning specialization was shown. The revealed differences in the associative and reactive properties (with respect to CS) between the neurons of the compact and diffuse PPTg parts testify to the functional heterogeneity of this structure and suggest the leading role of the cholinergic compact part of the PPTg in the instrumental defensive conditioning. Thus, the obtained evidence suggests the involvement of the PPTg in the mechanisms of attention and acquisition of the active defensive motor conditioning. PMID- 12125398 TI - [Dopamine antibodies as neuromodulators of behavioral reactions in mice of different genotypes]. AB - A possibility of the long-term modification of inborn behavioral features of different mice genotypes (C57B1/6 and BALB/c) by active immunization with dopamine-bovine serum albumin conjugate was investigated. Significant interstrain differences were found in the effects of dopamine antibodies on the open-field behavior and the content of neurotransmitters in the brain cortex and striatum. It was shown that the active immunization of mice to dopamine produces an increase in the functional activity of brain dopamine receptors. The extent, to which this increase is pronounced, is genotype-dependent. PMID- 12125400 TI - [Reactivity of brain benzodiazepine receptors and individual sensitivity of rats to pentylenetetrazole]. AB - The individual sensitivity of the male Wistar rats to acute pentylenetetrazole injection was studied, the density and the affinity of benzodiazepine receptors in the cerebellar cortex for 3H-diazepam was measured. It was demonstrated that the reactivity of benzodiazepine receptors underlies the individual sensitivity to pentylenetetrazole. The animals with higher sensitivity were characterized by more intensive reaction than the control and resistant animals, i.e., by an decrease in the receptors density (the initial receptors density being equal in the sensitive, resistant, and control animals). Daily injections of a subconvulsive dose of pentylenetetrazole during 24 days increase the animal sensitivity to this substance, and this was accompanied by an increase in the reactivity of benzodiasepine receptors. Later on, the produced high sensitivity became somewhat lower but persisted for 6 months. The receptors density in this period reduced almost by half. In sensitive rats, a single low dose of pentylenetetrazole injected 6 months after treatment increased the density of benzodiazepine receptors. The age-matched controls, the same acute dose of pentylenetetrazole decreased both the receptor density and affinity of their binding. It is suggested that the increase in reactivity of benzodiazepine receptors is actualized via the intracellular metabotropic feedback mechanism. PMID- 12125399 TI - [Critical role of intracellular calcium in plasticity mechanisms of defense behavior command neurons LPl1 and PPl1 of Helix lucorum during nociceptive sensitization]. AB - The role of intracellular calcium in changes in excitability and responses of defense behavior command neurons LP11 and PP11 of Helix lucorum to sensory stimulation was investigated in semi-intact preparation of a snail during nociceptive sensitization. It was found that application of sensitizing stimuli onto the snail's head initiated membrane depolarization, increase in its excitability as well as depression of neural responses evoked by sensory stimuli in short-term period of sensitization and significant facilitation of neural responses in long-term period of sensitization. To elucidate the contribution of LP11 and PP11 neurons in plasticity rearrangements involved in the mechanisms of sensitization, we applied sensitizing stimuli during strong hyperpolarization of the neurons or after intracellular injection of calcium chelators. Application of sensitizing stimuli during hyperpolarization of the neurons suppressed the increase in membrane excitability and depressed the neural responses evoked by chemical stimulation of snail's head i.m. short- and long-term periods of sensitization. At the same time, synaptic facilitation of neural responses evoked by tactile stimulation of snail's head and foot was observed, which was similar to synaptic facilitation in the control sensitized snail. Intracellular injection of EGTA or BARTA (calcium chelators) before sensitization suppressed synaptic facilitation in neural responses evoked by sensory stimulation. Under these conditions, the increase in excitability was more pronounced then in the control snail neurons. The experimental results suggest the changes in neural responses evoked by sensory stimulation in sensitized snails involve postsynaptic calcium dependent mechanisms of plasticity in LP11 and PP11 neurons. PMID- 12125401 TI - [Intercentral reticulo-cortical relations of brain electrical activity during "animal hypnosis" in rabbits]. AB - The electrical activity of the right and left sensorimotor and premotor cortical areas and right and left medulary reticular formation was recorded during "animal hypnosis" in rabbits. In this state, the spectral power of potentials (predominantly, in the delta-range) recorded from the left reticular formation was higher than that recorded from the right side. The value of the function of coherence between the right and left reticular recordings was decreased to 0.1 0.2 in the whole frequency range. The tight-side intrahemispheric coherence between the activities recorded from the sensorimotor cortex and reticular formation was higher than respective left-side values, whereas the coherent relations between the activities recorded from the reticular formation and premotor cortex were not changed (as compared to nonhypnotic state). PMID- 12125402 TI - [Modulation of neuronal evoked responses to the medial septal area of hibernating ground squirrels by some neuropeptides after chronic disconnection between septum and preoptico-hypothalamic structures]. AB - Effects of some neuropeptides identified in the brain of hibernators (TSKYR, TSKY, DY) and of monoaminergic neurotransmitters (noradrenaline and serotonin) on responses of the medial septal neurons evoked by intraseptal electrical stimulation were analyzed in slices taken from the ground squirrels with chronic basal undercutting of the septum. Despite the elimination of direct contacts with the preoptic area and afferents ascending in the medial forebrain bundle, the neurons retained almost normal level of reactivity and distribution of the reaction types. The neuropeptides effectively modulated neuronal responses of various types, including oligosynaptic short-latency single-spike responses. The latter were strongly facilitated by the neuropeptides. As a rule, changes in the responses to electrical stimulation were independent of the spontaneous activity shifts (in 78% of the tests). It was suggested that the neuropeptides exert a double influence on the septal neurons: direct nonsynaptic effects on the pacemaker potential responsible for the background activity and modulation of synaptic processes. Our experiments showed that descending influences of the septo-hippocampal system are not crucial for the entrance into the hibernation state and its tonic maintenance. The influences of the thermoregulatory- circadian structures of the preoptico-hypothalamic area determine the paradoxically increased latent excitability of septal neurons that allows the septo-hippocampal system to gate external stimuli and organize arousal of the forebrain during hibernation in case of emergency. PMID- 12125403 TI - [Behavioral impairments in Morris water maze in rats selectively bred for audiogenic seizure susceptibility (Krushinsky-Molodkina strain)]. AB - Krushinsky-Molodkina rats (KM strain) with genetically determined seizure susceptibility (clonic and tonic seizures in response to the sound of an electric bell, Krushinsky, 1960) were tested in two versions of Morris water maze and compared with normal albino rats (Sprague-Dawley and Wistar). The tests revealed a learning deficit in KM rats. They showed slow acquisition in both the spatial version of the test and the version with the platform, less efficient strategy of searching for target platform, and high scores of floating and thigmotaxis. However, males of KM rats (not females) did not differ significantly from Wistar strain in the probe trial in the spatial variant of the Morris test. No preference for searching for the platform at the place of its previous localization was observed in KM females. Together with our previous findings of the low scores in Revecz-Krushinsky test and data of other authors (Batuev et al., 1983) concerning a working memory deficit in the radial maze, the results suggest the of complex cognitive deficit combined with possible increased stress reactivity in KM rats. PMID- 12125404 TI - [Impairment of latent inhibition in OXYS rats with hereditary syndrome of premature aging]. AB - The effect of latent inhibition has been tested in OXYS (strain selected for accelerated aging) and Wistar (control) rats at the age of 4 months using a conditioned passive avoidance reaction. The OXYS rats displayed a higher level of anxiety in the elevated plus-maze as compared to Wistar strain. The experimental procedure consisted of preexposure stage (presentation of non-reinforced environmental stimulus), conditioning (presentation of a preexposed stimulus paired with shock), and testing the effect of latent inhibition. The effect of latent inhibition was revealed only in Wistar rats, in which the latency of transition to the dark compartment of the experimental chamber was reduced. Contrastingly, in OXYS rats the conditioned performance was not inhibited. It is suggested that aging-associated factors such as the low rate of habituation to experimental situation at the preexposure stage and the high general anxiety level of OXYS rats can interfere with inhibition of the attention to irrelevant information. PMID- 12125405 TI - [Effects of dehydroepiandrosterone on passive avoidance in ovariectomized female rats of different age]. AB - The aim of the present work was to study the influence of long-term treatment with dehydroepiandrosterone (DHEA) in doses of 0.1 and 0.7 mg/kg, i.p. on the passive avoidance performance in the ovariectomized female rats of 5- and 18 month old. The results obtained indicated that DHEA administration during 7 days in dose of 0.1 mg/kg normalized the passive avoidance performance in the ovariectomized rats of 5-month old while DHEA administration during 7 days in dose of 0.7 mg/kg restored passive avoidance performance in the ovariectomized rats of 18-month old. PMID- 12125406 TI - [Serotonin in brain regions of rats differing in their inborn locomotor activity]. AB - Serotonin contents in the paraventricular hypothalamic nucleus (PVN) and dorsal hippocampus of rats with different levels of inborn motor activity were studied by microdialysis in basal and stimulated conditions. Rats were exposed to elevated platform and forced swimming stress. In basal conditions, differences in serotonin contents between rats with different levels of inborn motor activity were found neither in hippocampus nor in PVN. In both kinds of stress conditions, serotonin content in hippocampus increased only in rats with higher level of inborn motor activity. Serotonin content in PVN dramatically increased during forced swimming in both rat groups. This increase was significantly more pronounced in rats with low activity. The data suggest that serotonin release in stress depends on inborn motor activity, brain area dialyzed, and the stressor the animals were exposed to. PMID- 12125407 TI - [Corneal dystrophies and molecular genetics. Results of current research reveal prospects for new therapeutic possibilities]. PMID- 12125408 TI - [Corneal dystrophies in the light of modern molecular genetic research]. AB - Modern methods of molecular genetics have resulted in new insights into the pathogenesis of corneal dystrophies which now require a new classification. Clinical evidence, the results of histopathological and electron-microscopical examinations, especially in cases of recurrence which reflect early disease, and the immunohistochemical analysis of the deposits have already aroused the suspicion that the "old" classification dividing the dystrophies into those of the epithelium, of the so-called anterior membrane, of the stroma and of the endothelium may no longer be adequate. The detection of the BIGH 3 gene which is mainly expressed in the corneal epithelium, and its gene product keratoepithelin, have led to the insight that the so-called anterior membrane dystrophies (Reis Bucklers, Thiel-Behnke) as well as the more common "classical" stromal dystrophies (granular dystrophies Types I and II, lattice dystrophies Types I and IIIA) are caused by different mutations of the above mentioned BIGH3 gene and are thus to be regarded as epithelial in origin. Lattice dystrophy Type II is part of the Meretoja syndrome, a systemic amyloidosis, and is caused by a mutation of the gelsoline gene on chromosome 9 (9q34). Gelsoline is also predominantly expressed in the corneal epithelium. In addition, the responsible genes, their gene products and the mutations are known for Meesmann's epithelial dystrophy and for the so-called gelatinous drop-like dystrophy, while in other dystrophies only the location on a certain chromosome can be given, namely: 16q22 for the macular dystrophy, 1p36 for the central crystalline dystrophy of Schnyder and 20p11.2 q11.2 for the congenital hereditary endothelial and for Schlichting's posterior polymorphous dystrophies. As the production rate of new results in molecular genetics is very fast, the proposed new classification can only be of preliminary character. PMID- 12125409 TI - [Possibilities and prospects of therapy and prevention of corneal dystrophies]. AB - Therapeutic approaches in corneal dystrophies should aim at long-term avoidance of recurrences. At the moment, this goal can only be achieved by transplantation of healthy corneal cells. In dystrophies of keratocytes and/or endothelial cells this can be realized by conventional penetrating keratoplasty. In epithelial dystrophies, however, simultaneous transplantation of limbal stem cells is necessary. Optimal HLA-matching strategies and immunosuppressive medication are major means to prevent immunological destruction of these. PMID- 12125410 TI - [Multifocal ERG with 30 Hz flicker stimulation in glaucoma patients and normal probands]. AB - BACKGROUND: It was the aim of the present study to investigate the value of multifocal ERG with 30 Hz flicker stimulation for glaucoma diagnosis. METHODS: Multifocal ERGs with 30 Hz flicker stimulation were recorded with a mean luminance of 50 cd/m2 and a contrast of 99% from the central 60 degrees of the retina which were stimulated by 61 hexagons. From 30 patients with primary and secondary open-angle and low-tension glaucomas with reproducible visual field losses and glaucomatous optic disc atrophy and 21 normal subjects, one eye was included in the study. The first harmonic wave, the basewave and the ratio of these two parameters were ana-lysed. The responses of seven neighbouring hexagons were averaged for the intraocular comparison between areas with and without perimetric defects and for the comparison between glaucomas and normal subjects. RESULTS: In the glaucoma group, the ratio of the first harmonic wave and the basewave was significantly lower in an area within a perimetric defect than outside the defect (p < 0.001) when compared intraocularly. The extent of ERG changes however did not correlate with the extent of perimetric defects. Comparing the glaucoma with the normal group, the ratio of the first harmonic and the basewave was significantly lower in an area within the perimetric defect in the glaucoma group than in corresponding areas of the normal group (p = 0.002). The first harmonic and the basewave did not differ significantly. DISCUSSION: These results show that the method is not appropriate to objectively indicate visual field losses although it statistically separates glaucomas from normal subjects. PMID- 12125411 TI - [Complications with foldable intraocular lenses with subsequent explantation in 1998 and 1999. Results of a questionnaire evaluation]. AB - PURPOSE: A questionnaire was sent to all members of the German Society of Ophthalmic Surgeons to evaluate complications of foldable intraocular lenses (IOLs) that required explanation. METHODS: Information on preoperative visual acuity, foldable IOL design and material as well as the reason for IOL explantation was obtained and analysed. We received 167 completed questionnaires for 1998 and 1999. RESULTS: In 1998 and 1999 the most common reasons for IOL explantation were as follows: incorrect lens power for 56% of the 3-piece hydrophobic acrylic IOLs, 16% because of glare or other photic phenomena, 40% of the 1-piece hydrophilic acrylic IOLs were explanted because of incorrect lens power and 30% for IOL damage. For the 3-piece monofocal silicone IOLs, 41% were explanted because of incorrect IOL power and 32% because of IOL decentration. For the 1-piece Hydrogel IOL, 76% were explanted because of opacification of the optic and 14% because of incorrect lens power. Most multifocal IOLs were explanted because of photic phenomena. CONCLUSION: In addition to the most common complications such as decentration and incorrect IOL power observed in rigid IOLs, new complications associated with foldable IOLs occurred such as optic opacification, glare and photic phenomena. Some complications seemed to appear in particular IOL types (opacification: SC-60BOUV, MDR), while others were observed in all types of foldable lenses. Accurate calculation of the IOL power and further improvement of the IOL material and design seem to be necessary to minimise the rate of explantations. PMID- 12125412 TI - [Perforating keratoplasty with transplants from pseudophakic donors]. AB - PURPOSE: Pseudophacia is considered to be a relative exclusion criterion for cornea donation. The aim of this retrospective study was to investigate the clinical outcome and prognosis of penetrating keratoplasty with grafts from pseudophacic donors. PATIENTS AND METHODS: From April 1996 to July 2000 16 patients received grafts from pseudophacic donors (group I). The control group (group II) comprised 26 patients with grafts from donors without a history of intraocular surgery. Graft survival, ratio of grafts without immune reactions and chronic endothelial cell loss were compared between both groups. RESULTS: During a follow-up of almost 1 year, no graft failures and no graft rejections were observed in group I. In group II 90% of the grafts remained clear and 91% of the grafts remained free of immune reactions in the same follow-up period (Kaplan Meier estimation). The differences between the study groups were not statistically significant. Chronic endothelial cell loss was 0.86 +/- 0.82 cells per mm2 and day in group I and 1.4 +/- 1.8 cells per mm2 and day in group II. This difference was not statistically significant. CONCLUSION: Our clinical results reveal no reason to regard pseudophacia as a principal contraindication for cornea donation, provided a critical endothelial evaluation is carried out. PMID- 12125413 TI - [In vivo measurement of temperature during phacoemulsification]. AB - BACKGROUND: Based on our experience of in vitro measurements of temperature in the anterior chamber in 1994 we designed a new spatula with an integrated probe that enabled us to measure temperature continuously during phacoemulsification without any surgical effort. METHODS: We measured the temperature in the anterior chamber during 50 cases of phacoemulsification. Depending on the indications, we used different energy modes (0-100%) and pulse modes ranging from 3 to 10 pulses per second or continuous ultrasonic output. Different viscoelastic substances were also used. RESULTS: The average temperature in the anterior chamber was 30 degrees C and adapted to the temperature of the irrigation fluid within seconds as soon as irrigation was started. During phacoemulsification each ultrasonic output causes an increase of temperature. In the case of continuous mode, the maximum rise in temperature was 3.5 degrees C. When the pulse mode was used the temperature increased up to 1.7 degrees C. It is important to note that one single pulse may increase the temperature by 0.8 degree C close to the phacotip. The rise of temperature correlated with the viscosity of the viscoelastic substance used. CONCLUSIONS: The new probe proved to be a practical device to monitor temperature in the anterior chamber during phacoemulsification. Up to now we did not encounter any critical degrees of temperature but it should be pointed out that a functional irrigation unit, the use of pulse mode or permanent ultrasonic output and in particular the type of viscoelastic material used are all important parameters influencing the rise of temperature during phacoemulsification. Further experience and results obtained with the new probe will be reported in the future. PMID- 12125414 TI - [Multifocal ERG with confocal scanning laser ophthalmoscope. Comparison with monitor simulation]. AB - BACKGROUND: Multifocal electroretinograms (mfERG) were recorded using a confocal scanning laser ophthalmoscope (cSLO) and compared to the results from conventional monitor stimulation. METHODS: Single and repeated measurements were recorded from 23 normal subjects using the cSLO (Heidelberg Retina Angiograph, Heidelberg Engineering, Heidelberg) as well as a conventional monitor as stimulation devices. Laser power output was modified by various optical filters. The reliability of the method and agreement with the conventional monitor stimulation were determined. RESULTS: CSLO recordings showed a high degree of variability. Reduction of laser power output improved the retinal response topography and characteristically modified response variations with each filter. Differences in amplitude size between cSLO and monitor recordings decreased with increasing amplitude levels. The results of repeated measurements showed considerable variation. CONCLUSION: It is possible to use a cSLO as a stimulator for mfERG recordings. However, a relatively high degree of variability represents a significant limitation of this method. Appropriate reduction of laser power decreases variations and serves to obtain photopic response topographies. PMID- 12125415 TI - [Implantation of toric intraocular lenses for correction of high post keratoplasty astigmatism]. AB - BACKGROUND: High postkeratoplasty astigmatism remains a challenge for the surgeon. First experiences after implantation of a toric PMMA IOL in three eyes from patients with cataracts are reported. MATERIALS AND METHODS: After routine phacoemulsification we implanted an individually manufactured toric PMMA posterior chamber IOL via a sclerocorneal 6-mm tunnel incision in three postkeratoplasty eyes with high, topographically relatively regular astigmatism. One eye with the intraoperative aspect of circular zonular instability also received a capsular tension ring. A complete standard ocular examination was performed pre- and postoperatively including corneal topography, evaluation of anterior chamber depth, ultrasonic biomicroscopy and perimetry. RESULTS: Implantation and intraoperative alignment of the toric IOL were uneventful. The refractive astigmatisms of 5.0, 6.0 and 7.5 cyl D preoperatively, were reduced to 2.0, 1.75 and 3.0 cyl D, 10 and 12 months postoperation, respectively. The eye with the capsular tension ring showed no morphological or refractive changes during follow-up. In the other two eyes we observed IOL rotation of 20 and 30, respectively after 6 months. There were no posterior capsule opacification or capsular folds in the optical centre. CONCLUSION: Toric IOL technology allows enhancement of IOL surgery with improved refractive outcome. Simultaneous implantation of a capsular tension ring may improve long-term stability. PMID- 12125416 TI - [First unilateral, later bilateral optic neuropathy. Amiodarone as the cause?]. AB - CASE REPORT: Amiodarone has many severe side-effects and can cause keratopathy and neuropathy. We report the case of a 79-year-old male patient with progressive visual loss and papilloedema of the right eye during therapy with amiodarone. To reduce papilloedema he was treated with corticosteroids. Amiodarone therapy was then discontinued, but there was no considerable improvement and after 6 weeks visual loss occurred in the left eye as well. CONCLUSION: With regard to the findings we consider this case to be an amiodarone-induced optic neuropathy. According to our knowledge only 19 cases have been reported in the literature but the results of one study indicate that the incidence of optic neuropathy is as high as 1.79%. The pathophysiology of the optic nerve damage remains unclear and an effective treatment is not available. It might be possible that the accelerated elimination of amiodarone with colestyramine could be suitable to prevent optic neuropathy of the second eye. PMID- 12125417 TI - [Treatment of primary retinal detachment. Minimal extraocular or intraocular?]. AB - The developments in treatment modalities for a primary retinal detachment over the last 70 years have been reviewed. There was a change from a surgery limited to the area of the break to a form of prophylactic surgery including the extent of the detachment. In between Rosengren had limited the treatment to the break with an intraocular gas bubble. A change was brought about by Custodis in 1953 who limited surgery to the break and omitted drainage. This procedure had serious postoperative complications which were eliminated by Lincoff by developing the cryosurgical detachment operation which was subsequently refined to extraocular minimal surgery. The ultimate realization of a minimal extraocular approach was the operation with a temporary balloon. Two additional intraocular procedures evolved, pneumatic retinopexy and primary vitrectomy, following one or the other pattern of treatment. With all four methods reattachment can result in 94-99% of the cases but differences can be seen in the morbidity and rate of reoperations. PMID- 12125418 TI - [Papilledema associated with visual field defects and motility disorders]. PMID- 12125420 TI - [Differential eyelid tumors diagnosis. II]. PMID- 12125419 TI - [Glass orbital foreign body 15 years after windshield injury]. PMID- 12125421 TI - [An original initiative at the Nantes Information Center for Human Immunodeficiency Care]. PMID- 12125424 TI - [Nurses in a home nursing care service for elderly persons]. PMID- 12125425 TI - [Other modes of contraception]. PMID- 12125426 TI - [Nurses' education in medical device surveillance]. PMID- 12125427 TI - [The tortured child]. PMID- 12125428 TI - [Parents of an autistic child]. PMID- 12125429 TI - [Primary prevention, a State responsibility?]. PMID- 12125430 TI - [Registered nurses in the center of the world campaign against diabetes]. PMID- 12125431 TI - [Malnutrition in elderly patients: from the hospital to home]. PMID- 12125432 TI - [Female condoms. An valid contraception method and a solid defense against sexually transmitted diseases]. PMID- 12125433 TI - [The diabetic child and sports]. PMID- 12125434 TI - [Syphilis: the return of the "great imitator"]. PMID- 12125437 TI - [Geriatric care]. PMID- 12125436 TI - [Home care services. An opportunity for our health care system]. PMID- 12125438 TI - [Nurses' words]. PMID- 12125439 TI - [60 minutes to persuade... yes, but about what?]. PMID- 12125440 TI - [Evaluation of the risk of nosocomial infection in home hospitalization]. PMID- 12125441 TI - [Being a liberal nurse in autodialysis]. PMID- 12125442 TI - [Care strategy. Communication in the treatment of pain]. PMID- 12125443 TI - [Care knowledge (1/2). History and theory]. PMID- 12125444 TI - [Care strategy. From nurses' knowledge to nursing knowledge]. PMID- 12125445 TI - [Research. Theories of care and care knowledge, from concepts to tools]. PMID- 12125446 TI - [Care strategy. Nursing knowledge and standardized language]. PMID- 12125447 TI - [Evolutions of languages and evolving languages]. PMID- 12125448 TI - [Organization. Nursing knowledge and professionalization]. PMID- 12125449 TI - [Care strategy. Index knowledge, nursing, and narrative]. PMID- 12125450 TI - [Care strategy. Care knowledge, between art and science]. PMID- 12125451 TI - [Good utilization of medications. A. Classification of antibiotics. 1/12 -- Overview]. PMID- 12125452 TI - [Emergencies, an indispensable identification... and urgent!]. PMID- 12125453 TI - [Elaboration of competence and in the area of communication]. PMID- 12125454 TI - Determination of traces of iron in indium phosphide by electrothermal atomic absorption spectrometry combined with solvent extraction. AB - An electrothermal atomic absorption (ETAAS) method for the determination of traces of iron (0.1-1.0 microgram g-1) in Fe-doped indium phosphide (InP) has been developed. In order to overcome the indium matrix-effect and to achieve a useful detection limit, a preliminary solvent-extraction of Fe(III) with acetylacetone (HAA) is necessary. After sample dissolution with hydrochloric acid (1 + 1) the digest is evaporated to dryness, Fe(II) is oxidized to Fe(III) with nitric acid, the residue is dissolved in 0.01 mol L-1 HCl and the iron is extracted at pH 2.0 with 0.5 mol L-1 HAA in toluene. The organic phase is injected into the graphite furnace and the iron is directly evaluated by external organic standard calibration. The limit of detection (3SB) resulting from further in-situ preconcentration is 0.03 microgram g-1. When the method was applied to the analysis of real samples containing 0.2-0.7 microgram g-1 Fe, the RSD was in the range 8-21%. Results were compared with those independently obtained on the decomposed sample solution with inductively coupled atomic emission spectrometry (ICP-AES). The detection limit of the ICP-AES method, that needs matrix-matched standards, is 0.20 microgram g-1. PMID- 12125455 TI - Determination of the acid dissociation constant of bromocresol green and cresol red in water/AOT/isooctane reverse micelles by multiple linear regression and extended principal component analysis. AB - The pKa of 3',3",5',5"tetrabromo-m-cresolsulfonephtalein (Bromocresol Green) and o-cresolsulphonephtalein (Cresol Red) was spectrophotometrically measured in a water/AOT/isooctane microemulsion in the presence of a series of buffers carrying different charges at different water/surfactant ratios. Extended Principal Component Analysis was used for a precise determination of the apparent pKa and of the spectra of the acid and base forms of the dye. The apparent pKa of dyes in water-in-oil microemulsions depends on the charge of the acid and base forms of the buffers present in the water pool. Combination with multiple linear regression increases the precision. Results are discussed taking into account the profile of the electrostatic potential in the water pool and the possible partition of the indicator between the aqueous core and the surfactant. The pKa corrected for these effects are independent of w0 and are close to the value of the pKa in bulk water. On the basis of a tentative hypothesis it is possible to calculate the true pKa of the buffer in the pool. PMID- 12125456 TI - Stability constants of iron(II) sulfate complexes. AB - The complex formation equilibria between iron(II) and sulfate ions have been studied at 25 degrees C in 3 M NaClO4 ionic medium by measuring with a glass electrode the competition of Fe2+ and H+ ions for the sulfate ion. The concentrations of the metal and of the ligand were varied in the ranges 0.01 to 0.125 and 0.01 to 0.250 M, respectively. The analytical concentration of strong acid was chosen to be 0.01 or 0.03 M. The potentials of the glass electrode, corrected for the effect of replacement of medium ions with reagent species, have been interpreted with the equilibria [formula: see text] Stability constants valid in the infinite dilution reference state, logK zero = 1.98 +/- 0.16, log beta 1 zero = 2.1(5) +/- 0.2 and log beta 2 = 2.5 +/- 0.2, have been estimated by assuming the validity of the specific interaction theory. PMID- 12125457 TI - The effect of ionic strength on the complexation of copper (II) with salicylate ion. AB - The complexation of Cu2+ with the hydrogen salicylate, HL-, ion has been studied, at 25 degrees C, by potentiometric measurements with a glass electrode in NaClO4 media for an ionic strength range from 0.5 to 3 M. The complexes found were CuL, CuL2(2-) and CuHL+. Stability constants, extrapolated to zero ionic strength by using the Specific Interaction Theory were: [formula: see text] PMID- 12125458 TI - Treatment options for tannery wastewater II: integrated chemical and biological oxidation. AB - This is the second of two papers each dealing with a specific technological option for replacing the Fenton's reagent with simpler processes for treating industrial wastewater. In particular, the paper reports the results of an investigation aimed to check, at lab scale, the effectiveness of an alternative wastewater treatment combining biological degradation and chemical oxidation with ozone. The treatment was carried out in a lab scale hybrid reactor fed with the biological stage effluent of a plant treating the wastewater of a large tanning district in Central Italy whose residual COD result still higher than the Italian COD Maximum Allowable Concentration (MAC) value (i.e., 160 mgO2/L) The results are very promising, considering that a removal efficiency of 41% (as COD) has been achieved by treating an influent characterized by a COD content fully biorefractory. In addition, the proposed treatment presents the significant advantage of no additional sludge production, as happens with commonly utilized tertiary processes (i.e. Fenton), that is characterized by high chemical sludge production. PMID- 12125460 TI - Protonation and complex formation of 5-sulfosalicylate in NaCl, CaCl2 and MgCl2 aqueous media. Speciation in synthetic seawater. AB - The binding capacity of 5-sulfosalicylic acid (ssa) towards cationic macro components of natural waters has been investigated in different ionic media (NaCl, MgCl2 and CaCl2 aqueous solutions) and in the ionic strength range 0 < or = I < or = 1 mol dm-3. In order to contribute to the speciation of this multi sites ligand, measurements have been carried out also in a synthetic seawater (SSWE) containing the major components of seawater (Na+, K+, Ca2+, Mg2+, Cl- and SO4(2-)). Measurements have been performed by potentiometry ([H+]-glass electrode), at t = 25 degrees C. A critical analysis on the experimental and literature data is also given. PMID- 12125459 TI - Chemometric investigations on environmental data from Carlo Alberto irrigation canal (Alessandria, Piedmont, Italy). AB - The Carlo Alberto Canal connects Bormida and Tanaro rivers in Piedmont (ITALY). It was created for irrigation purposes but since its waters are suspected to be polluted, a sampling campaign was performed by the ARPA of Alessandria. The physico-chemical parameters analysed along 3 years (1998-2000) were investigated by multivariate chemometric methods. PCA showed that the waters situation depends heavily on the sampling period. Also a Kohonen self-organising map confirmed the clustering observed, providing insights into the causes of the clusterisation. New samplings are now being performed and a larger set of environmental variables is determined on each sample. PMID- 12125461 TI - Simultaneous determination of nanomole amounts of sulphur dioxide and hydrogen sulphide by flow injection analysis with on-line preconcentration by means of capillary denuder tubes. AB - A simple and rapid method for trace determination of SO2 and H2S in gaseous samples by using a flow injection system with on line preconcentration on capillary denuder is described. The gaseous samples are led through a 0.4 M sulphamic acid solution, retaining nitrogen dioxide, ammonia and hydrogen chloride. The sulphur dioxide is collected from the carrier gas stream (250 cm3 min-1) as sulphuric acid in a capillary denuder tube coated with a thin layer of 0.01-0.03 M hydrogen peroxide solution of 0.05 mM sulphuric acid; hydrogen sulphide passes into a second tube coated with 0.075 mM sodium sulphide solution of 0.1 M aqueous sodium hydroxide. The films containing the sulphuric acid and the sodium sulphide, respectively, are eluted with the corresponding circulating absorbent streams and pass through the detectors. Sulphuric acid is detected by conductimetry and sulphide is determined spectrophotometrically at 230 nm. If nanoequivalent amounts of H2S are present in the sample containing a large concentration of SO2 (SO2/H2S concentration ratio > 20), the sulphur dioxide is filtered out of the sample gas stream by solid sodium hydrogen carbonate. A limit of detection of 3.5 micrograms m-3 is obtained. PMID- 12125462 TI - Effect of the medium on the ionization constants of some triazole compounds. AB - The deprotonation and acid ionization constants of some triazole derivatives in various aqueous-organic solvent mixtures were determined potentiometrically at 20 degrees C. The organic solvents used were methanol, ethanol, DMF, DMSO, acetonitrile, acetone and dioxane. The high stabilization of both the non protonated form by dispersion forces and of the proton by its interaction with the organic solvent are the main factors influencing the deprotonation constant in aqueous mixtures of methanol, ethanol, DMF or DMSO. On the other hand, the hydrogen bonding interactions and the solvent basicity, in addition to the electrostatic effect, contribute to the major effects in the deprotonation process (in solutions enriched with acetonitrile, acetone or dioxane) and the acid ionization process in different aqueous-organic solvent mixtures. Some thermodynamic parameters (delta H, delta G, delta S) of the ionization processes in a pure aqueous medium are also determined and discussed. PMID- 12125463 TI - Preconcentration of trace silver with yeast for river water analysis. AB - A new preconcentration method with yeast is presented. The method was evaluated for the determination of trace silver in river waters by graphite furnace atomic absorption spectrometry (GFAAS). A suitable cultivation bed for preconcentration of silver was 1.75 mg ml-1 2-ammonium hydrogen phosphate. The optimal cultivation time and temperature were 2 h and 25 degrees C. Under optimal conditions, silver in aqueous sample was concentrated to 6.9-fold by yeast. The detection limit was 4.6 pg ml-1 (3S/N) for silver in river water. The yeast preconcentration method was applied to the determination of silver in river waters. The recovery of spiked silver was in the range of 89 to 110%. By the preconcentration, it was found that ultra trace silver in river waters could be determined without interferences of matrix elements, after only the cultivation and with no chemical treatment. PMID- 12125464 TI - Electrothermal atomization of arsenic, antimony and thallium using a graphite atomizer with refractory metal platforms. AB - The electrothermal atomization of the volatile elements arsenic, antimony and thallium from a refractory metal platform consisting of a tungsten coil and/or a refractory metal foil with the dimensions of a conventional graphite platform was studied. Several combinations of refractory metal platforms were investigated, as follows: W platform; Ta platform; W coil; W coil on a W platform and W coil on a Ta platform. The best combination for these elements as regards both thermal stabilization and sensitivity is the W coil on a Ta platform. Thermal stabilization is also achieved with a W coil on a W platform. The presence of Pd containing chemical modifier favors the thermal stabilization of the analytes. The sufficient amount is 2 micrograms of Pd. The maximal temperatures of pyrolysis are higher (arsenic, antimony) or equal (thallium) to those when using different chemical modifiers, added as solutions. It may be concluded, that the refractory metal platforms act as "built-in modifiers". They are suitable for the determination of arsenic, antimony and thallium in samples of complex matrix composition where high thermal stability of the analytes during the pyrolysis step is required. PMID- 12125465 TI - Spectroscopic, spectrophotometric and potentiometric studies on 3-(naphthylazo)-5 phenylpyrazole dye and its metal chelates. AB - The electronic absorption spectra of 3-(naphthylazo)-5-phenylpyrazole dye (L) are studied in some organic solvents of different polarities. The spectra comprise three absorption bands, which are assigned to the corresponding electronic transitions. The pKa value of (L) in ethanol-water mixtures have been determined pH-metrically at different temperatures. The thermodynamic parameters delta G zero, delta H zero and delta S zero have also been evaluated. The successive stability constants of some d-transition elements have been determined pH metrically at 25 degrees C and mu = 0.1 in 60% (v/v) ethanol-water mixture. A 1:1 and 1:2 (M: L) complexes are formed and evidenced by conductometric studies. The solid complexes of Cu(II) have been isolated and characterized on the basis of elemental analysis, TGA and IR data. The complexes formed between (L) and Cu(II) were studied spectrophotometrically in solutions. PMID- 12125466 TI - Liquid anion exchanger study of gold (III) from aqueous halide media with N-n octylaniline. AB - N-n-octylaniline in xylene is used for the extractive separation of gold(III) from halide media. Gold(III) was extracted quantitatively with 10 ml of 2% reagent in xylene from 0.5-10 M and 0.5-8 M hydrochloric acid and hydrobromic acid, respectively. It was stripped from the organic phase with ammonia buffer solution (pH 10.1) and estimated spectrophotometrically with stannous chloride. The effect of metal ion, acids, reagent concentration and of various foreign ions has been investigated. Method is applicable to the analysis of synthetic mixtures containing platinum metals and alloy samples. The method is fast, accurate and precise. PMID- 12125467 TI - [Vitamin status of patients with respiratory tract diseases]. AB - Of the with infringement of the acidity--basic condition the tendency to Decrease of a level of vitamins B6 and C is marked in comparison wits those at the patients without infringement of the specified condition. PMID- 12125468 TI - [Soybean proteins in correction of coronary risk factors in patients with cardiovascular diseases]. AB - At 95 patients with coronary artery disease and high blood pressure studied influence of a diet with low lipid by a directedness including yields of processing soy, on dynamics(changes) of clinical symptoms, indexes lipid of the status and coronary risk factors. The application of a diet, 50% of fiber in which was replaced on soy protein isolate, promoted positive dynamics(changes) of clinical manifestations of disease on a hum noise of improving of indexes lipid of a spectrum and coronary risk factors. The research has shown, that the use in the treat-preventive purposes soy protein isolate is rather perspective. PMID- 12125469 TI - [Experience of nutritional support in late period of traumatic disease of wounded patients]. AB - There are no standards of nutritional support of wounded patients in different periods of disease. Influence of supplementation with Nutrition on dynamic of anthropometric indexes, some biochemical and laboratory parameters in wounded malnourished patients in the last period of disease was examined. The results of investigation indicated that supplementation with Nutrizon increased all tianthropometric indexes, normalized albumin blood level, nitrogen balance, decreased nitrogen of urine. PMID- 12125470 TI - [Comparative study of postprandial glycaemia in type 2 diabetic patients after consumption of mono- and disaccharides and sweeteners]. AB - It was investigated the influence of mono- and disaccharides, sugar alcohols, honey, corn patoka and products with nutritive and nonnutritive sweeteners on dynamic of postprandial glycaemia in type 2 diabetic patients. After ingestion of 30 g fructose, blood glucose did not show a marked increase in comparison with sucrose or honey. After ingestion of 30 g sorbitol or isomalt, blood glucose curve was not significantly different. It was indicated that corn patoka in chewing candies with isomalt has a high hyperglycaemic effect whereas drink with nonnutritive sweeteners did not change blood glucose from fasting levels. PMID- 12125471 TI - [Food dietary supplement efficacy in complex therapy of patients with type 2 diabetes with obesity]. AB - The dynamic of glycemia, insulin, C peptide, glycosylated hemoglobin, fructosamine, parameters of serum lipids, lipid peroxidation and system of antioxidant defense in 28 patients with diabetes mellitus type 2 associated with obesity was studied of influence of hypoglycemic hypocaloric diet. Besides favorable influence to a clinical picture of the disease universal normalizing influence of hypoglycemic hypocaloric diet with food dietary supplement "Cogent DB+" on parameters of carbohydrate, lipid and oxidative metabolism. The scheme of investigation of efficacy of food dietary supplements in complex treatment of patients with diabetes mellitus type 2 associated with obesity was offered on basis of study of influence "Cogent DB+" on mechanisms of metabolic disorders in these patients. PMID- 12125472 TI - [Diet in low-income families: adult working population]. AB - The epidemiological research of the nutrition and health status of the low-income people was held in five regions of the Russian Federation. The analyze of the nutrients, energy, micronutrients and of the main groups of foods consumption in the families with the different level of income was executed. The estimation of the nutrition status was made with using body mass index. The results of this work showed the need of the development of nutrition support in social service and using the medical criteria for this support in low-income people. PMID- 12125473 TI - [Effects of biologically active pectin-containing dietary supplement on gastroduodenal motility in patients with a functional dyspepsia]. AB - For 21 patients with a functional dyspepsia the influencing biologically active additives to nutrition "Pekcecom" on dynamics of clinical symptoms and parameters gastroduodenal motility under the data gastroduodenoscintigraphy was studied. The usage of biologically active additives during 4 weeks was accompanied by deboosting of accelerated gastric emptying for want of statistically significant influencing on a normal and delayed gastric emptying and parameters of duodenal transit. PMID- 12125474 TI - [Clinical study of calcium-containing bioactive food additive in postmenopausal osteoporosis]. AB - The purpose of the investigation: to study the biological active supplement (BAS) Calcimax influence on the pain syndrome (PS) dynamics and the psycho-emotional breaches, on the ionized calcium level in the blood plasma of the osteoporosis patients. The Calcimax has been produced by the "Artlife" company (Tomsk, Russia). The calcium hydroxiapatit (200 mg per 1 capsule) It is balanced with the microelements and vitamins. 31 women, who suffer from the osteoporosis disease, at the age of 47 to 64, took place in the prospective randomized investigation. All the patients were subdivided in two groups (16 and 15 patients). The basic group patients, exempt prophylaxis and following the recommendations took two capsules after light supper within 6 months. The comparative analysis of the monthly monitoring of the average number of the PS intensity has shown the trustworthy (p < 0.05) lowering of them. The Calcimax rises the (IC) level in the blood plasma also. Thus the Calcimax may be used as the common preventive remedy. PMID- 12125475 TI - [Protective influence of bioactive food supplement "Rekicen RD" in alimentary T-2 mycotoxicosis in rats]. AB - Inclusion of a biologically active food supplement (BAS) "Rekicen RD" in rat Wistar ration in percentage 5% result in reduction of toxic action of a trichothecene mycotoxin T-2 toxin. The changes of daily increase in body weight of animals, of relative internal weight, of serum enzyme activity, of not sedimentated activity of lysosomal enzymes and of activity of enzymes of a xenobiotic metabolism in a liver were expressed in a lesser degree. It is possible that a protective action of BAS is related with high adsorption capacity of wheat bran in BAS composition and with their possible influence on activity of enzymes participating in detoxification T-2 toxin (UDP-glucuronosyltransferase). PMID- 12125476 TI - [Physiology of nutrition]. AB - The article presents the data about scientific theories of nutrition. It is showed the necessity of gastro-intestinal tract participation in metabolic substances, discovered by I.P. Razenkov. PMID- 12125477 TI - [Nutrition assessment of patients with cataract]. PMID- 12125478 TI - [Influence of bioactive dietary supplement--dietary unsaturated fats--on immune status of patients with myocardial ischemia and impaired glucose tolerance]. AB - Immune status measured by the flow cytrometric analysis under the ischemic heart disease and IHD with impaired glucose tolerance has been found to be altered in B cells as well as in T lymphocytes in some cases. Having received the diet supplemented by PUFA "Eicolen", IHD patients present with the restored immune status. Patients with IHD and impaired glucose tolerance did not present with any significant modifications in immune status after PUFA "Eicolen" supplementation. PMID- 12125479 TI - Clinical study on the treatment of male immune infertility with sheng jing zhong zi tang. PMID- 12125480 TI - Study on cytokines IL-2, IL-6, IL-10 in patients of chronic allergic rhinitis treated with acupuncture. AB - OBJECTIVES: To observe the plasmatic concentration of IL-6, IL-10 and IL-2 in the patient of chronic allergic rhinitis before and after acupuncture therapy. METHODS: Cytokine levels were determined before and after treatment in 30 healthy volunteers (Group A) and 90 patients of chronic allergic rhinitis (Group B) with an increased plasma IL-10 level. Group B was then divided into 3 subgroups: 30 patients treated with real acupuncture (Group B1); 30 patients treated with sham acupuncture (Group B2); 30 non-treated patients (Group B3). RESULTS: The allergic subjects of group B1, compared with controls, showed a significant reduction of IL-10 after a specific treatment with acupuncture (P < 0.05). On the other hand, in those patients treated with sham acupuncture (B2) as well as in non-treated patients (B3), the IL-10 values remained high and unchanged. There was a statistically significant change in IL-2 values at 24 hours (P < 0.05) after real acupuncture (Groups A, B1), however the values remained within normal ranges. The IL-6 do not change after therapy. CONCLUSION: The acupuncture treatment can reduce plasmatic level of IL-10 in chronic allergic rhinitis. PMID- 12125481 TI - A variant maneuver of acupuncture in treating cervical spondylopathy. AB - Lifting and thrusting with reducing method at acupoint Huatuojiaji (Extra 21) of the affected side was performed to make the limb of the affected side contract involuntarily 1-2 times in 53 cases with cervical spondylopathy. The maneuver produced a much better total effective rate than the routine maneuver (P < 0.01). The instant and long-term analgesic effects were also found superior (P < 0.01). Probably this is due to the increased serum content of morphine-like substance. PMID- 12125482 TI - Point injection for treating nephritic colic. PMID- 12125483 TI - Fifty-six cases of acne treated by auricular needle-embedding. PMID- 12125484 TI - Acupuncture treatment of superficial pain by subcutaneous needling. PMID- 12125485 TI - Clinical application of the point yongquan. PMID- 12125486 TI - TCM treatment of fundus hemorrhage due to obstruction of retinal vein. PMID- 12125487 TI - Cupping therapy for 103 cases of high fever due to infection of the upper respiratory tract. PMID- 12125488 TI - Treatment of protrusion of the lumbar intervertebral disc by TCM massage. PMID- 12125489 TI - Examples of clinical application of fengchi. PMID- 12125490 TI - Clinical experience in application of the acupoint futu. PMID- 12125491 TI - Effects of electroacupuncture at weiwanxiashu and zusanli points on blood glucose and plasma pancreatic glucagon contents in diabetic rabbits. AB - The Effects of electroacupuncture (EA) at Weiwanxiashu (EX-B3) and Zusanli (ST 36) points on blood glucose (BG) and plasma pancreatic glucagon (PG) contents were dynamically observed in diabetic rabbits induced by Alloxan. It is found that acupuncture at Weiwanxiashu point can significantly lower the BG content and inhibit release of PG; no significant changes in BG and PG are found when acupuncture is given at Zusanli (ST 36) point alone, however BG and PG contents decrease more obviously when acupuncture employed at both Zusanli and Weiwanxiashu, suggesting that Zusanli has a marked synergetic action with Weiwanxiashu. PMID- 12125493 TI - Application of healthy-side needling to treatment of apoplectic hemiplegia. PMID- 12125494 TI - Acupuncture treatment of hiccup. PMID- 12125492 TI - Advances in TCM symptomatology of rheumatoid arthritis. PMID- 12125495 TI - How to treat cervical spondylopathy with acupuncture? PMID- 12125497 TI - Brief introduction to application of dosage of Chinese herbs. PMID- 12125498 TI - A comparison between the Chinese and European patients in acupuncture treatment. PMID- 12125499 TI - Long-term effect of TCM decoctions in treatment of nephrotic syndrome. AB - Fifty-seven cases of nephrotic syndrome were treated with TCM decoctions as accessory treatment for prednisone and cyclophosphamide, and the effects were observed in a follow-up period of 5-15 years. The long-term complete remission rate of 68.4% and recurrence rate of 26.3% in the treatment group were respectively higher and lower than those in the control group (P < 0.01, and P < 0.01). The results suggested that the TCM decoctions were very helpful in treating this condition. PMID- 12125501 TI - Clinical observation on the therapeutic effects of zhongyan-2 recipe in treating 29 HIV-infected and AIDS patients. AB - OBJECTIVE: To investigate the efficacy of Zhongyan-2 Recipe (ZY-2), a recipe consisting of Chinese herbal medicine. METHODS: Twenty-nine HIV-infected patients were treated with ZY-2 for 3 months as one treatment course; and viral load, immune function and symptoms-signs were determined before and after treatment. RESULTS: After treatment, the absolute values of CD4 were elevated in 8 cases, and lowered in 14 cases, 7 cases were not changed; the ratio of CD4/CD8 were elevated in 2 cases, lowered in 1 case, 16 of them were not changed obviously. The CD4 and CD4/CD8 ratio were elevated simultaneously in 2 cases. Neopterin lowered in 4 cases, elevated in 11 cases, and not changed in 1 case. Viral load (VL) was lowered in 6 cases, elevated in 8 cases and not changed in 2 cases. The total effective rate was 42.28% on the basis of comprehensive assessment of VL, immune function, body weight and symptoms-signs. CONCLUSION: ZY-2 is an effective Chinese recipe in treating HIV-infected patients. PMID- 12125500 TI - A clinical and experimental study on the protective effects of jiangzhi tongmai fang on endothelial injury. AB - 31 cases of atherosclerosis (AS) were treated with Jiang Zhi Tong Mai Fang ([symbol: see text], formula of JZTMF), and its effect was compared with 30 cases treated with lovastatin in the control group. Clinically, the JZTMF formula showed an effect of regulating blood lipids, and therefore it was antiatherosclerotic. The mechanism is, probably, restoration of the function of endothelial cells (EC) by increasing the synthesis of 6-keto-PGF1 alpha and decreasing the release of endothelin (ET) as evidenced in the experimental study. PMID- 12125502 TI - Application of bu zhong yi qi tang according to TCM differentiation of syndromes. PMID- 12125504 TI - Interleukin-5 messenger RNA expression in peripheral blood mononuclear cells from patients with bronchial asthma and eosinophilia. AB - Interleukin-5 (IL-5) plays an important role in allergic diseases accompanied by eosinophilia. We examined IL-5 messenger RNA (mRNA) expression in peripheral blood mononuclear cells (PBMC) isolated from 13 patients with bronchial asthma, 1 patient with chronic eosinophilic pneumonia, 2 patients with idiopathic eosinophilia, and 5 control subjects. IL-5 mRNA was expressed in PBMC from 2 of 13 patients with asthma, 1 patient with chronic eosinophilic pneumonia, and 1 of 2 patients with idiopathic eosinophilia. IL-5 mRNA expression was not detected in PBMC from control subjects. PBMC from a patient with chronic eosinophilic pneumonia expressed IL-5 mRNA spontaneously. Prednisolone treatment decreased his clinical symptoms and IL-5 mRNA expression. IL-5 mRNA expression preceded revival of the disease. These observations show that IL-5 plays a role in the condition, accompanied by eosinophilia, and IL-5 mRNa examination in PBMC by means of reverse transcription-polymerase chain reaction may be useful to predict the activity of eosinophilia. PMID- 12125503 TI - Nasal polyps in patients with rhinitis and asthma. AB - The objective of this study was to investigate the prevalence of nasal polyposis in Greek patients with chronic rhinitis and asthma. We studied 3817 patients (2342 men and 1385 women) who were referred for allergy evaluation during 1990 1998 and diagnosed as having chronic rhinitis and asthma. Skin-prick tests with allergens common in Greece and controls were used in all subjects. A wheal with a mean diameter > or = 3 mm was considered as positive. According to the history of symptoms and the results of skin tests, patients were divided into the following groups: patients who have allergic rhinitis (seasonal, perennial), patients who have allergic asthma (seasonal, perennial), patients who have nonallergic rhinitis, and patients who have nonallergic asthma. All patients were examined for nasal polyps by anterior rhinoscopy and endoscopic investigation with a rigid or/and flexible endoscope. We found that 4.2% of the patients with chronic rhinitis and asthma (4.4% of the men and 3.8% of the women; p > 0.05) had nasal polyps. The prevalence of nasal polyps increased with age (p < 0.001) in both sexes. The prevalence of nasal polyps was 13% in patients with nonallergic asthma, 2.4% in patients with allergic asthma, 8.9% in patients with nonallergic rhinitis, and 1.7% in patients with allergic rhinitis. Nasal polyps were found in 3.6% of the patients with rhinitis and in 4.8% of the patients with asthma (p > 0.05). Nasal polyps were present more frequently (1) in patients with nonallergic respiratory disease (rhinitis, asthma) than in patients with allergic respiratory disease (10.8% versus 2.1%; p < 0.001) and (2) in patients with perennial respiratory allergy (rhinitis, asthma) than in patients with seasonal respiratory allergy (4.8% versus 0.4%; p < 0.001). We found that 4.2% of patients with chronic rhinitis and asthma had nasal polyps. Nasal polyps were present more frequently in nonallergic patients than in allergic patients and in patients with perennial allergy than in patients with seasonal allergy. PMID- 12125505 TI - Furosemide plus albuterol compared with albuterol alone in children with acute asthma. AB - Several reports have shown that inhaled furosemide protects patients with asthma from different bronchoconstrictor agents. However, the effect of this widely used diuretic in acute exacerbation in adults is unproven. There are no reports of furosemide's therapeutic effect in acute asthma in children; thus, the objective of this study was to determine the effectiveness of the combined treatment of furosemide and albuterol in pediatric patients. Using a double-blind design, 20 emergency room patients with an asthmatic exacerbation were studied and randomly assigned to one of the following treatments: (1) furosemide + albuterol (1 and 0.15 mg/kg, respectively) or (2) albuterol (0.15 mg/kg). The forced expiratory volume in one second (FEV1) was measured in each patient before medication and then 30 and 60 minutes after inhalation of the individual drug or drug combination. Neither group differed in age or baseline FEV1. An increase in FEV1 of 22.8 +/- 4.3% (mean +/- SE) in the drug combination group was noted at 60 minutes, and an increase in FEV1 of 18.0 +/- 2.6% in the albuterol group was obtained at the same time. Although the increase in FEV1 was greater in the first group after 1 hour of treatment, this was not significant. These results suggest that inhaled furosemide does not have a synergistic effect with albuterol in the treatment of asthmatic exacerbations in children. PMID- 12125507 TI - Tacrolimus ointment for the treatment of atopic dermatitis: clinical and pharmacologic effects. AB - The topical immunomodulator tacrolimus ointment has been shown to be safe and effective in the treatment of atopic dermatitis in clinical trials involving over 16,000 patients. Clinical trial results focusing on tacrolimus' safety and efficacy are summarized. Minimal systemic absorption results from topical application in patients with atopic dermatitis. Although the exact mechanism of action of tacrolimus ointment in atopic dermatitis is unknown, tacrolimus is known to inhibit up-regulation of cytokine production following T cell activation and to decrease Fc epsilon RI expression on dendritic antigen-presenting cells in skin. Additional mechanisms of action of tacrolimus relevant in the pathogenesis of inflammatory skin disorders are discussed. PMID- 12125506 TI - Cytokines in nasal lavage fluids from acute sinusitis, allergic rhinitis, and chronic fatigue syndrome subjects. AB - The aim of this study was to compare the degree of inflammation present in acute sinusitis, allergic rhinitis, chronic Fatigue Syndrome (CFS), and non-CFS control subjects by measuring cytokine concentrations in nasal lavage fluids. The concentrations of total protein (TP; Lowry assay), nerve growth factor (NGF), tumor necrosis factor (TNF) alpha, and interleukin (IL)-8 were measured by ELISA in nasal lavage fluids from acute sinusitis (n = 13), active allergic rhinitis (n = 16), CFS (n = 95), and non-CFS (n = 89) subjects. CFS and non-CFS groups were subdivided further using allergy skin test and rhinitis score results. Acute sinusitis subjects had significantly higher TP (p = 0.011, ANOVA), TNF-alpha (p = 0.00071), and IL-8 (p = 0.0000027) concentrations and IL-8/TP ratios (p = 0.0030) than the other three patient groups. There were no differences based on skin test or rhinitis score severity within either the CFS or non-CFS groups. The mucopurulent discharge of acute sinusitis contained significantly higher TNF alpha and IL-8. Neutrophils were a likely source for these cytokines. There were no differences between CFS and non-CFS subjects, making it unlikely that the rhinitis of CFS has an inflammatory component. PMID- 12125508 TI - Identification of immunoglobulin E binding components of the two-spotted spider mite Tetranychus urticae: allergenic relationships with the citrus red mite and house-dust mite. AB - The two-spotted mite (TSM) is commonly found on fruit trees, herbaceous plants, and greenhouse flowers. However, a recent investigation indicated that the sensitization rate to TSM was as high as that of house-dust mites (HDMs) in nonfarmers as well as in farmers working in orchards in this country. The aim of this study was to identify immunoglobulin E (IgE)-binding components within TSM and to evaluate the allergenic relationship with the citrus red mite (CRM) and HDM. Sera were collected from eight patients who were not farmers and who had asthma with high serum-specific IgE to the TSM and from unexposed controls showing negative responses to the TSM on skin-prick test. Twelve percent sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblot analysis were applied. To evaluate allergenic relationships with HDMs and CRMs, two kinds of sera pools were used: one (A) showing positive responses to both TSMs and HDMs and the other (B) showing an isolated positive response to TSMs. ELISA inhibition tests using A and B pooled sera were conducted. The TSM-ELISA inhibition test using sera A showed significant inhibition with addition of TSMs and CRMs, partial inhibition with HDMs, and minimal inhibition with other inhalant allergens. The ELISA inhibition test using sera B showed significant inhibition with TSMs and CRMs and minimal inhibition was noted with HDMs as well as other inhalant allergens. Immunoblot analysis using individual sera showed seven IgE binding components (75, 56, 47, 41, 37, 28, and 14 kDa) and two (75 and 14 kDa) of them were bound to IgE in > 50% of the sera tested. Seven IgE-binding components were identified within the TSM extract and two (75 and 14 kDa) could be considered major allergens. It is suggested that the TSM contains species specific allergen in addition to shared allergens with CRMs and HDMs. PMID- 12125510 TI - Asthma among the famous: series index. PMID- 12125509 TI - Taxol reactions. AB - Paclitaxel (Taxol) a taxane antineoplastic agent causing irreversible microtubule aggregation with activity against breast, ovarian, lung, head and neck, bladder, testicular, esophageal, endometrial and other less common tumors was derived from the bark of the Pacific yew (Taxus brevifolia). Phase I trials conducted in the late 1980s were almost halted because of the high frequency of hypersensitivity like reactions. Respiratory distress (dyspnea and/or bronchospasm), hypotension, and angioedema were the major manifestations, but flushing, urticaria, chest, abdomen, and extremity pains were described also. Reactions occurred on first exposure in the majority of cases raising etiologic questions. The vehicle for paclitaxel Cremophor EL (polyoxyethylated castor oil in 50% ethanol) was strongly suspect as a direct (non-immunoglobulin E dependent) histamine releaser. Premedication regimens and longer infusion times lowered the incidence of reactivity allowing phase II and III trials to progress through the early 1990s. The mechanism(s) underlying paclitaxel hypersensitivity-like reactions is still unknown, and clinical data on probable complement and mast cell activation are lacking. The original clinical trial protocols for paclitaxel required discontinuation of therapy for patients who experienced hypersensitivity-like reactions. Here, we review the current etiologic knowledge of these reactions and describe our clinical approach to allow completion of chemotherapy with this powerful plant-derived agent. PMID- 12125511 TI - Public health. As good as new. AB - The current reorganisation of public health risks the specialty being marginalised. There is not sufficient public health expertise to go round 300 primary care trusts. To be effective, networks will need clear structures and accountability. PMID- 12125512 TI - Public health. Too much to handle? AB - Organisations involved in public health welcome primary care trusts' role in the reorganised service and the establishment of public health networks. But there are concerns about shortage of specialist expertise. They believe PCTs have not yet developed their public health role. The public health role of strategic health authorities is seen as unclear. There are concerns about inequities in the distribution of public health resources to PCTs. PMID- 12125514 TI - Open space. Three's is a crowd. PMID- 12125513 TI - US healthcare. United straits. PMID- 12125515 TI - Data briefing. Nurse vacancies. PMID- 12125516 TI - Serological profile following malaria outbreak in Mewat region of Haryana, India. PMID- 12125517 TI - Malarial mitochondrial genome: the 6 kb element. PMID- 12125518 TI - Impact of urbanization on bionomics of An. culicifacies and An. stephensi in Delhi. AB - Study on bionomics of malaria vectors was carried out in riverine and non riverine areas, on account of tremendous ecological changes in the topography of Delhi. The densities of adult anophelines were estimated by two techniques, hand catch and total catch index. Percentage of An. stephensi (15.68) collected by both the techniques was more than An. culicifacies (3.16) in both the areas. Day time resting preferences of vector species in human dwellings and cattlesheds did not differ significantly. Preferred larval habitats of An. culicifacies in riverine area shifted to large lakes, channels and ponds. In malaria transmission, An. culicifacies played a role only in the northern part of the riverine area where water pollution was at minimal level, while An. stephensi played an equal role in the malaria transmission in both the areas. High sporozoite rates were found in type form of An. stephensi in localities where its proportion was high, thus confirming its active role in malaria transmission. The overall sporozoite rate of vectors was 0.7 per cent and P. falciparum sporozoite infections of the vectors were detected in An. stephensi only. P. vivax and P. falciparum infections were found in the ratio of 68:32. The non-riverine area was more malarious than the riverine area. PMID- 12125519 TI - Plasmodium falciparum dihydrofolate reductase Val-16 and Thr-108 mutation associated with in vivo resistance to antifolate drug: a case study. AB - Due to increasing trend in chloroquine resistance, the antifolate (sulpha pyrimethamine combination) drugs are gaining more importance in the treatment of uncomplicated falciparum malaria. The efficacy of sulpha-pyrimethamine combinations in the treatment is compromised by the development of resistance in parasite. The occurrence of mutations at active sites in Plasmodium falciparum gene sequences coding for dihydrofolate reductase (DHFR) and dihydropteroate synthetase (DHPS) confer resistance to pyrimethamine and sulphadoxine. This study presents the characterization of a P. falciparum sample from a patient who did not respond to standard doses of a pyrimethamine/sulpha regimen. Although parasitaemia fell rapidly, the infection had not resolved six days later as because the response to treatment selected resistant sub-population. The in vitro drug sensitivity assays demonstrated resistance to pyrimethamine, sulphadoxine and cycloguanil; while polymerase chain reaction (PCR) and restriction digest based methods indicated that at known drug resistant loci the isolate had a genotype of DHFR Val-16 and Thr-108 previously only associated with cycloguanil resistance. As per the published reports this type of paired mutations in natural isolates are rare. It is of considerable interest to carry out studies on alleles on alleles of this gene in relation to resistance at epidemiological level. PMID- 12125520 TI - Serological appraisal of malaria status in tribal area of Orissa, India. AB - A cross-sectional seroepidemiological study conducted in 173 tribal residents (including all age groups) from three villages of Sundergarh district of Orissa using ELISA as a tool revealed high levels of antibody titres against both AR1 and Pf antigens. The mean +/- S.D. ELISA O.D. obtained for AR1 were 0.73 +/- 0.2, 1.037 +/- 0.196, 1.05 +/- 0.42 and for Pf, 0.70 +/- 0.2, 1.0 +/- 0.28, 0.94 +/- 0.4 respectively for Badramoli, Jharbeda and Boneikela population. Parasitological results showed high incidence of malaria in < 5 years age group in Badramoli and Boneikela villages when compared to other age groups. An. fluviatilis was found to be the principle malaria vector as found in the HBI results. High ELISA O.D. and high equivalent transmission index (ETI) indicate a high malaria transmission in the area. Seroepidemiological studies could be used for effective surveillance and stratification of endemicity in a given area which can help in executing intervention strategy for malaria control. PMID- 12125522 TI - Prevalence of G-6-PD deficiency and sickle-cell haemoglobin carriers in malaria endemic tribal dominated districts--Mandla and Jabalpur, Madhya Pradesh. PMID- 12125523 TI - Diagnostic imaging has a bright future. PMID- 12125521 TI - Dynamics of malaria transmission under changing ecological scenario in and around Nanak Matta Dam, Uttaranchal, India. AB - To understand the transmission dynamics of malaria in three different ecotypes, namely watershed (forest), seepage (Nanak Matta Dam) and plain (non-forest, non dam) areas of Nainital and Udham Singh Nagar districts of Uttaranchal, entomological and parasitological investigations were carried out from July 1996 to June 1997. In the three ecotypes, average per man hour densities of adult vector species in human dwellings and cattlesheds recorded were high for Anopheles culicifacies from April to September and October to March for An. fluviatilis. Prevalence of both An. culicifacies and An. fluviatilis was higher in the forest area as compared to other two areas. Observations on gonotrophic condition revealed endophilic tendency of both vector species. Higher number of both vector species were found in outdoor than indoor during night human bait collections. Out of 864 specimens of An. fluviatilis dissected, one showed natural infection of sporozoites in salivary glands in the month of November from the forest area only. Sibling species study of An. fluviatilis revealed the presence of species S for the first time in the forest area. Parasitological investigations also depicted high incidence of malaria in the forest area as compared to other two areas. Overall results from the study indicated active malaria transmission in the forest area. PMID- 12125524 TI - Mammography is a case study in the economics of radiology. PMID- 12125525 TI - Imaging technology outlook: promising future for new imaging technologies. PMID- 12125526 TI - Words, words, words: meanings, meanings, meanings! PMID- 12125527 TI - Critical thinking and nursing science: judgment, or vision? AB - For decades, critical thinking has been widely regarded as a concept and process of central importance in the practice of nurses and in their education. Numerous nursing textbooks heavily emphasize the development of critical thinking skills in students. The attention given to critical thinking in nursing parallels a reform movement that has had an impact on virtually all fields. Close attention to the ways in which critical thinking is addressed in nursing, however, reveals a starkly delimited view of its meaning within the discipline. This article questions whether the vision of critical thinking as a cornerstone of practice in nursing may need to be expanded. The discussion centers on three points: nursing theories and frameworks as ways to enrich critical thinking in nursing practice; the inadequacy of the model of critical thinking as an individual, analytical process; and possibilities that are cocreated when critical thinking is conceived as a creative/constructive, relational/dialogical process. PMID- 12125528 TI - Research evaluating human becoming in practice. AB - In this column the author synthesizes and discusses the findings of six studies conducted to examine what happens for nurses and patients when human becoming is the guide for practice. In all of the studies, nurse participants' descriptions led to three main themes: transforming intent, unburdening joy, and struggling with change. Patient and family participants' descriptions of nursing care guided by human becoming are also summarized. This column concludes with a presentation of the universal and overarching values for knowledge development in nursing that emerged with the synthesis of the findings and that have the potential to ensure personalized, meaningful, and dignified nursing service delivery. PMID- 12125529 TI - Scientific merit and research ethics. AB - It is widely accepted that scientific merit and research ethics are closely related. And yet, while the ethical principles that guide research are uncontested, the meaning of scientific merit is open to interpretation based on one's philosophical perspective and research tradition. When adherents of the natural science tradition use the criteria of that tradition to evaluate research situated in the human science tradition, misunderstanding and misjudgment will result. This unfortunate situation has negative implications for the flourishing of human science nursing and, ultimately, may deprive society of opportunities to benefit from humanistic practice aimed at enhancing the quality of life for recipients of healthcare. In such circumstances, it behooves nurses committed to human science nursing to continue to vigorously articulate and pursue their values and ethics concerning nursing research and healthcare. PMID- 12125530 TI - Doctoral education in nursing: seeking clarity. Interview by Sandra Schmidt Bunkers. PMID- 12125531 TI - Learning to practice the discipline of nursing. AB - This column addresses the issues of teaching and growing the discipline of nursing. Experiences of two undergraduate nursing students highlight the benefits and opportunities that accompany the teaching of nursing theories in undergraduate programs. The 3rd-year baccalaureate students, Kendra Fitzsimmons and Elspeth Ferguson, submitted papers from their York University nursing program to help demonstrate key points in this column. Their papers show thoughtful critique and passionate belief in nursing as a discipline and a basic science. Kudos to them and the faculty who inspired them. PMID- 12125532 TI - The nurse theorists: 21st-century updates--Jean Watson. Interview by Jacqueline Fawcett. PMID- 12125533 TI - Having courage: a lived experience of human becoming. AB - The purposes of this research were to discover the structure of the experience of having courage and to contribute to knowledge about human becoming. Participants were 10 persons with spinal cord injuries. The Parse research method was used to answer the research question, What is the structure of the lived experience of having courage? The central finding of this study is the following structure: The lived experience of having courage is a fortifying tenacity arising with triumph amid the burdensome, while guarded confidence emerges with the treasured. The findings are discussed in relation to human becoming, relevant literature, and future research. PMID- 12125534 TI - Living with multiple sclerosis: a Roy adaptation model-based study. AB - This Roy adaptation model-based study examined how people make sense of the experience of living with multiple sclerosis. Eighteen persons diagnosed with multiple sclerosis participated in three interviews over a period of 1 year. Content analysis of the interview transcripts and the researcher's logs and analytic memos yielded five recurring themes that reflect the Roy model adaptive modes: we're not completely the same (physiological mode), how I view my future (self-concept mode), let me tell you about my feelings (self-concept mode), how I see work (role function mode), and let me tell you about my life (interdependence mode). PMID- 12125535 TI - King's theory as foundation for an advance directive decision-making model. AB - This article discusses the utilization of King's conceptual system, transaction process model, and theory of goal attainment as foundations for an advance directive decision-making model. Research has shown nurses may be educationally unprepared, experience conflicts between beliefs and actions, or resist the responsibility to address advance directives and end-of-life issues. Nurses, especially nurse practitioners providing primary care, are in positions to facilitate the process. By understanding and incorporating this model into practice, both the nurse and the client may achieve mutual goal attainment resulting in both increased client autonomy and Patient Self-Determination Act compliance. PMID- 12125536 TI - Family reflections on living through sudden death of a child. AB - The author describes a method of cooperative inquiry into a family's experience of the sudden death of a child. Through the praxis method based on Newman's health as expanding consciousness, family members had an opportunity, individually and together, to explore the impact of this experience on subsequent family life over three generations. Members were given transcripts of all dialogues for further reflection. All writings were shared with all family members. Both process and content of the inquiry will illustrate the shared and unique ways in which this loss was expressed over time. Implications of the mode of inquiry will be discussed. PMID- 12125537 TI - Dilemmas of long-term care. PMID- 12125538 TI - Long-term care for the elderly in Taiwan. AB - An aging population, the prevalence of chronic disease and functional disabilities, and changes in society have caused a greater demand for long-term care in Taiwan. The development of long-term care has gone through four stages: undeveloped, early beginning, expanding, and establishment. Problems in several areas including lack of care providers and facilities, expenses of care, and difficulty coordinating health and social services are issues that must be explored. Well-planned and managed long-term care services are an important task for the Taiwanese government to address so holistic care of elders can be provided in this new century. PMID- 12125539 TI - Parse's view of nursing science and art and about the human becoming theory (HBT). PMID- 12125540 TI - What is nursing science? PMID- 12125541 TI - Borrowed theories, shared theories and the advancement of nursing knowledge. PMID- 12125546 TI - An extremely stable open-framework metal-organic polymer with expandable structure and selective adsorption capability. AB - A novel porous framework polymer with expandable structure, over 8% volume expansion upon inclusion, has been characterized. PMID- 12125547 TI - A mixed-valence copper coordination polymer generated by hydrothermal metal/ligand redox reactions. AB - A novel coordination polymer of mixed-valence copper(I,II) with 4,4'-bipyridine and in situ oxidized isophthalate, [Cu2(ipO)(4,4'-bpy)] (ipOH = 2 hydroxyisophthalate), was hydrothermally synthesized and crystallographically characterized to be a laminated structure via weak copper(II)--oxygen interactions. PMID- 12125557 TI - A reversible resistivity-based nitric oxide sensor. AB - A sensor for nitric oxide is reported that uses a novel redox matching mechanism to induce a resistance change upon binding this important ligand to cobalt. PMID- 12125559 TI - Conventional free radical polymerization in room temperature ionic liquids: a green approach to commodity polymers with practical advantages. AB - Free-radical polymerization of methyl methacrylate and styrene using conventional organic initiators in the room temperature ionic liquid, 1-butyl-3 methylimidazolium hexafluorophosphate ([C4mim][PF6]) is rapid and produces polymers with molecular weights up to 10x higher than from benzene; both polymerization and isolation of products were achieved without using VOCs, offering economic as well as environmental advantages. PMID- 12125560 TI - Application of ionic liquids as plasticizers for poly(methyl methacrylate). AB - The room temperature ionic liquid, 1-butyl-3-methylimidazolium hexafluorophosphate, [C4mim][PF6] was found to be an efficient plasticizer for poly(methyl methacrylate), prepared by in situ radical polymerization in the ionic liquid medium; the polymers have physical characteristics comparable with those containing traditional plasticizers and retain greater thermal stability. PMID- 12125562 TI - Synthesis of carbamoylphosphonate silanes for the selective sequestration of actinides. AB - The synthesis of carbamoylphosphonate silanes (CMPO analogs) designed for sequestering actinide cations in self-assembled monolayers on mesoporous supports (SAMMS) is described. PMID- 12125565 TI - CsPb3Bi3Te8 and CsPb4Bi3Te9: low-dimensional compounds and the homologous series CsPbmBi3Te5 + m. AB - Two new thermoelectric materials of quaternary bismuth telluride CsPb3Bi3Te8 and CsPb4Bi3Te9 are reported, which are members of a homologous series featuring anionic slabs [PbmBi3Te5 + m]- (m = 1-4) of monotonically increasing thickness. PMID- 12125567 TI - Ustalic acid as a toxin and related compounds from the mushroom Tricholoma ustale. AB - A toxin and related compounds were isolated from a poisonous mushroom Tricholoma ustale. Their structures were determined by analyses of the spectral data and synthesis. PMID- 12125572 TI - Polar, non-coordinating ionic liquids as solvents for the alternating copolymerization of styrene and CO catalyzed by cationic palladium catalysts. AB - The palladium-catalyzed copolymerization of styrene and CO in an ionic liquid solvent, 1-hexylpyridinium bis(trifluoromethanesulfonyl)imide, gave improved yields and increased molecular weights compared to polymerizations run in methanol. PMID- 12125577 TI - Labelling of [Leu5]-enkephalin with organometallic Mo complexes by solid-phase synthesis. AB - The neuropeptide leucine-enkephalin ([Leu5]-enkephalin) has been covalently labelled with the two organometallic Mo carbonyl complexes Mo(R-His)(allyl)(CO)2 (His = N delta,N,O-L-histidinate) and Mo(R-bpa)(CO)3 (bpa = bis(2-picolyl)a-mine) on the resin as the last step of a solid-phase synthesis scheme and in solution. PMID- 12125580 TI - Stereochemical control of Zn(II)/Cu(II) selectivity in piperidine tripod ligands. AB - Stereochemistry plays a major role in the selectivity toward zinc ion over copper(II) of some tripodal ligands with a central piperidine scaffold, one of which acts as a fluorescent zinc sensor with nanomolar sensitivity. PMID- 12125583 TI - Crystal engineering of metalloporphyrin assemblies. New supramolecular architectures mediated by bipyridyl ligands. AB - Targeted synthesis of new supramolecular motifs of metalloporphyrins in crystals by a concerted mechanism of molecular recognition in three dimensions, aided by organic ligands, is presented; it involves induced assembly of [tetrakis(4 hydroxyphenyl)porphyrinato]zinc species by a combination of axial coordination through bridging bipyridyls and of lateral hydrogen bonding. PMID- 12125585 TI - Novel zinc fluorescent probe bearing dansyl and aminoquinoline groups. AB - A novel fluorescent chemosensor (L) demonstrates a remarkable selectivity and sensitivity for zinc(II) ion as evidenced from the solution characterisations and in vitro experiments using Hela cell lines. PMID- 12125587 TI - A screening system for enantioselective enzymes based on differential cell growth. AB - The esterase-catalyzed enantioselective hydrolysis of the fluoroacetate of pantolactone leads to fluoroacetic acid, a toxic compound which inhibits the growth of esterase-producing yeast; this forms the basis of an ee-assay. PMID- 12125590 TI - Proceedings of the meeting New Insights and Therapeutic Strategies in Cardiovascular Disease: Focus on Endothelial Targets. Venice, Italy, April 28-29, 2000. PMID- 12125591 TI - The 4th international six-day symposium. Prague, Czech Republic: 16-21 September 2001. PMID- 12125592 TI - Proceedings of the Symposium on Scientific Integration of Western Medicine and Complementary, Alternative, Mind Body Medicine. Seoul, Korea, July 1999. PMID- 12125593 TI - Proceedings of the 31st International Symposium on Growth Hormone and Growth Factors in Endocrinology and Metabolism. Valletta, Malta, 27-28 April 2001. PMID- 12125594 TI - Thrombosis and Haemostasis Issues in Cancer. International Conference. Bergamo, Italy, 2-4 November 2001. Lectures and abstracts. PMID- 12125595 TI - Day in the life. PMID- 12125596 TI - Proceedings of the 6th International Symposium on Inorganic Nitrogen Assimilation. Reims, France, 8-12 July 2001. PMID- 12125597 TI - Assess carefully before starting training programs. PMID- 12125598 TI - President Bush maintains focus on health care priorities. PMID- 12125599 TI - Day in the life. PMID- 12125601 TI - 2002 Membership list of the Belgian Royal Academy of Medicine. PMID- 12125600 TI - [Idiopathic hemosiderosis of the lung, synonym is Ceelen-Gellerstedt syndrome]. PMID- 12125602 TI - Proceedings of the 10th International Vasculitis and ANCA Workshop. April 25-28, 2002. Cleveland, Ohio, USA. PMID- 12125603 TI - Clinical laboratory reference 2002-2003. PMID- 12125604 TI - Side effects. Protease inhibitors and loss of fat. PMID- 12125605 TI - Side effects. Some notes on energy. PMID- 12125606 TI - Side effects. Nukes and fat cell damage. PMID- 12125607 TI - Side effects. Helping to repair damaged nerves. PMID- 12125608 TI - Side effects. Lactic acidosis. PMID- 12125609 TI - Side effects. Growth hormone helps get rid of fat. PMID- 12125611 TI - Anti-HIV agents. Antibiotic holidays may be OK for some. PMID- 12125610 TI - Side effects. Bone problems and anti-HIV therapy. PMID- 12125612 TI - Anti-HIV agents. AMD-3100--chemokine blocker under study. PMID- 12125613 TI - Immune boosters. CD8+ cells--from suppressors to saviours. PMID- 12125614 TI - Immune boosters. CD8+ cell infusions. PMID- 12125615 TI - Interferon and ribavirin for hepatitis C. PMID- 12125616 TI - Testosterone and depression in men. AB - Researchers at Harvard University recently conducted a study to compare levels of testosterone among HIV-positive men who had HIV-related weight loss. The researchers also gave some subjects injections of testosterone to find out if supplements of this hormone had an impact on feelings of depression. The researchers found that men who had low levels of testosterone were more likely to be depressed than men who had normal levels of this hormone. Moreover, when the depressed men received regular injections of testosterone their mood significantly improved. PMID- 12125617 TI - Anti-HIV agents. Early versus delayed therapy--results from Switzerland. PMID- 12125618 TI - Anti-HIV agents. Questions about the early and aggressive use of therapy. PMID- 12125619 TI - Anti-HIV agents. Infections after HAART--who is at risk? PMID- 12125620 TI - Anti-HIV agents. Rescue therapy. PMID- 12125621 TI - The risk of cervical dysplasia. PMID- 12125622 TI - Toxicity. Protecting the power plant. PMID- 12125623 TI - Vitamins C and E for nuke toxicity? PMID- 12125624 TI - Toxicity. St. John's wort--interactions with indinavir and other drugs. PMID- 12125625 TI - Anti-HIV agents. Trizivir--three drugs in one pill. PMID- 12125626 TI - Anti-HIV agents. TXU-PAP--early results. PMID- 12125627 TI - Anti-HIV agents. Hitting hard, hitting early--one year later. PMID- 12125628 TI - Anti-HIV agents. Efavirenz versus indinavir: who really wins? PMID- 12125629 TI - Anti-cancer agents. Cream reduces reappearance of cervical lesions. PMID- 12125630 TI - Toxicity. Study finds that HAART is OK for some people co-infected with hepatitis B and C virus. PMID- 12125631 TI - Anti-HIV agents. Immunotoxins to the rescue. PMID- 12125633 TI - Immune boosters. Cytokines can help fight HIV. PMID- 12125632 TI - Predicting lymphoma with soluble CD27. PMID- 12125634 TI - HAART as an immune booster. PMID- 12125636 TI - Immune boosters. Curcumin--some issues to consider. PMID- 12125635 TI - Immune boosters. A placebo-controlled study of Chinese herbs. PMID- 12125637 TI - Immune boosters. Curcumin and the immune system. PMID- 12125638 TI - Immune boosters. Mistletoe extract for HIV/AIDS? PMID- 12125639 TI - Anti-cancer agents. Curry for cancer? PMID- 12125640 TI - [II Guidelines on Ergometric Tests of the Brazilian Society of Cardiology]. PMID- 12125641 TI - [Guidelines for cardiologists on obesity and cardiovascular disease from the Departments of Atherosclerosis, Clinical Cardiology and FUNCOR of the Brazilian Society of Cardiology]. PMID- 12125642 TI - Abstracts of the 33rd International Danube Symposium. Lublin, August 29-September 1, 2001. PMID- 12125643 TI - A catalyst for me. PMID- 12125644 TI - Prognostic value of the erythrocyte sedimentation rate in patients with unstable angina. PMID- 12125645 TI - Diagnosis of chondroid lipoma by fine-needle aspiration biopsy. PMID- 12125646 TI - Coming of age of melanogenesis-related proteins. PMID- 12125647 TI - Pathologic quiz case. An incidental endometrial tumor masked by large leiomyoma. PMID- 12125648 TI - Pathologic quiz case. A colonic mass in a 53-year-old woman. PMID- 12125649 TI - Pathologic quiz case. A 30-year-old woman with severe disseminated intravascular coagulation during delivery. PMID- 12125650 TI - Pathologic quiz case. Multicystic ovarian tumor in a 46-year-old female patient. PMID- 12125651 TI - An unusual finding of plasma cell iron. PMID- 12125652 TI - A rare presentation of biphasic pulmonary blastoma. PMID- 12125653 TI - [XL Polish and VI International Medical Student Conference. Krakow, 2002. Abstracts]. PMID- 12125654 TI - Abstracts of the 17th European Congress of Obstetrics and Gynaecology. Prague, Czech Republic. May 22-25, 2002. PMID- 12125655 TI - Abstracts of the 35th Annual Meeting of the Society for the Study of Reproduction. July 28-31, 2002. Baltimore, Maryland, USA. PMID- 12125656 TI - Abstracts of the Annual Meeting of the Society for Experimental Biology. 8-12 April 2002, Swansea, United Kingdom. PMID- 12125657 TI - Cumulative author, subject, compound and special issue indexes, volumes 902-950. PMID- 12125658 TI - Author and subject indexes 1972-2000. PMID- 12125659 TI - Current practices in the validation of aseptic processing 2001. Technical report no. 36. PDA. PMID- 12125660 TI - [Even if fasting blood glucose value is quite normal, proceed with a glucose tolerance test]. PMID- 12125661 TI - Abstracts of the XXIV Endocrinology Meeting of Pisa. Italy, June 5-7, 2002. PMID- 12125662 TI - Abstracts of the International Conference 2002: Basic and Therapeutic Aspects of Botulinum and Tetanus Toxins. Hannover, Germany, June 8-12, 2002. PMID- 12125663 TI - The relation between alcohol consumption and smoking abstinence: results from the Working Well Trial. AB - The current study examined the relation between drinking and smoking abstinence in a community-based sample from the Working Well Trial (WWT). At baseline, drinking level was related to smoking history (never, former, or current smoker; P < .0001) and abstinence history. Mean monthly alcohol consumption increased linearly with decreases in duration of recent abstinence (i.e., longest period quit in the past year among current smokers; P < .05) and current abstinence (i.e., time since quitting among former smokers; P < .0001), even controlling for relevant demographic factors. Among baseline smokers, lower beer consumption predicted smoking abstinence at 4-year follow-up (P< .01). A trend towards significance was found for total alcohol consumption (P = .06). The results suggest (a) a dose-response relation between baseline drinking and duration of smoking abstinence, and (b) that heavier drinkers are less likely to quit smoking over a 4-year period. PMID- 12125664 TI - Totally endoscopic atrial septal defect closure with a robotic system: experience with seven cases. AB - BACKGROUND: The development of minimally invasive cardiac surgery has shown good clinical results with shorter recovery time and better cosmetic results. The introduction of the robotic systems can further reduce the surgical trauma and improve the surgical dexterity. We report seven cases of complete closed chest atrial septal defect closure using the "da Vinci" Surgical System (Intuitive Surgical, Mountain View, CA). METHODS: Following peripheral cannulation for cardiopulmonary bypass (CPB), aortic occlusion and cardioplegia delivery, five patients with atrial septal defect (ASD) and two patients with patent forame ovale (PFO) with atrial septal aneurysm (ASA) were successfully treated using the robotic system. Two robotic arms and an endoscopic camera were inserted through ports in the right hemithorax and an accessory port was placed for blood suction and ancillary instruments insertion. The defect closure was carried out with interrupted stitches in one patient and with a continuous suture in the others. RESULTS: Mean cardiopulmonary bypass and cross clamp time were 101.8 +/- 39.6 and 63.4 +/- 21.9 minutes respectively. Extubation was carried out within the seventh postoperative hour. All patients returned to normal lifestyle in one week. CONCLUSION: Complete closed chest ASD closure can be carried out using robotic technique with rapid postoperative recovery and excellent cosmetic result. PMID- 12125665 TI - Five years of less invasive mitral valve surgery: from experimental to routine approach. AB - BACKGROUND: In the last five years, mitral valve surgery has changed fundamentally. This study reviews our experience in less invasive mitral valve surgery (LIMS) during that time. METHODS: LIMS was performed in 449 patients (age 59 +/- 14 years, 237 female) via a right lateral minithoracotomy. The operations included 42 "redo" procedures. After initially experiencing a high number of complications, we have modified and simplified the procedure. After using the Port- Access? Technique (PAT) in the earlier stages of our series, in the last 226 patients the aorta was clamped directly using the transthoracic clamping (Chitwood) technique (TTC). In our most recent cases, PAT was only employed in redo procedures. In 336 patients, the procedure was completed with robotic assistance, and in 23 of these we used the da Vinci telemanipulation system. RESULTS: The mitral valve was repaired in 327 patients and replaced in 122 patients. In 100 patients, additional surgical procedures (TVR n = 13, ASD closure n = 16, left atrial ablation n = 65, left atrial reduction plasty n = 6) were performed. Bypass and clamp time, including times for both additional and redo procedures, were 124 +/- 44 min. and 65 +/- 29 min. in the overall series. Complications, which were mainly neurological, were fewer in the TTC group than in the PAT group (n = 4 vs. n = 17; p <0.04). Hospital mortality was 3.1% and 5.2%, respectively, for the two groups. There were no additional costs associated with using the TTC technique compared to conventional procedures. Mean survival rate was 96.3% at a mean follow-up of 727 +/- 451 days (95% CI, 677 to 779). CONCLUSIONS: Less invasive mitral valve surgery enables the patient to avoid the surgical trauma associated with sternotomy. It has developed into a reliable technique with reproducible results for primary, redo, and additional procedures. LIMS has become the standard approach for mitral valve operations at our institution. PMID- 12125667 TI - Why is "off-pump" coronary artery bypass grafting better? AB - The objective of this contribution is to review the results of operations we have performed upon patients using the off-pump coronary artery bypass grafting (OPCAB) technique. The OPCAB technique was examined as a cause of death in 2495 cases from September 1981 to September 1999. Our results indicated four deaths due to stroke and a total hospital mortality of 1.9% (48/2495). When myocardial revascularization without cardiopulmonary bypass (CPB) is appropriately chosen, it is a treatment to be highly recommended for patients with coronary insufficiency. PMID- 12125666 TI - Comparison of port access to sternotomy in tricuspid or mitral/tricuspid operations. AB - BACKGROUND: Outcomes for a port-access (PA) approach for tricuspid valve operations have not been reported or compared to those using median sternotomy (MS). METHODS: Retrospective analysis was performed for 88 consecutive patients undergoing tricuspid valve repair or replacement using port-access techniques (n = 27, 1997-2000) versus sternotomy (n = 61, 1990-1997). PA procedures were performed through a 6 cm right fourth interspace thoracotomy. RESULTS: PA patients had lower ejection fractions (46% +/- 11% vs. 54% +/- 10%, p = 0.02), but had a similar incidence of previous surgery (17/27 (63%) vs. 33/61 (54%), p = 0.4). PA patients had more frequent concurrent mitral valve operations (22/27 (82%) vs. 37/61 (61%), p <0.05) and more tricuspid repairs versus replacement (24/27 (89%) vs. 29/61 (48%), p <0.01). PA patients had longer pump times (254 min. +/- 82 vs. 162 min. +/- 61, p = 0.001) but comparable clamp times (65 min. +/- 15 vs. 63 min. +/- 41, p = 0.9), lengths of stay (14 days +/- 14 vs. 16 days +/- 16, p = 0.6), mortality (2/27 (7%) vs. 9/61 (15%), p = 0.3), strokes (3/27 (11%) vs. 4/59 (7%), p = 0.9), and need for new pacemaker implantation (5/27 (19%) vs. 12/61 (20%), p = 0.9). CONCLUSIONS: PA provided excellent short-term results comparable to MS in relatively high-risk tricuspid valve patients. For tricuspid operations, PA may have the advantage of avoiding sternotomy or reoperative sternotomy at the expense of longer pump times. PMID- 12125668 TI - How much myocardial revascularization can we do without extracorporeal circulation? AB - OBJECTIVE: The main goal of this study is to present a technical alternative that allows myocardial revascularization to be performed on the marginal branches of the circumflex coronary artery without the use of extracorporeal circulation (ECC). METHODS: The technique for exposing the coronary vessels is performed by placing a stitch in the posterior pericardium. The surgeon lifts the heart using his or her left hand with the aim of exposing the posterior pericardium. A single polypropylene "O" stitch on a #4 needle is threaded into a cardiac ribbon folded back on itself and run twice through the region between the inferior pulmonary vein and the inferior vena cava. The suture thread is run along a tourniquet-type rubber tube and the latter is then directed down to the pericardium, bringing the cardiac ribbon with it. Handling the thread by means of such a tourniquet and the two parts of the ribbon makes it possible to maneuver the heart into different positions in order to expose the coronary arteries: anterior interventricular, diagonal, circumflex, and right coronary. In the present study, the position exposing the coronary arteries was adopted. The circumflex artery was exposed by separating the two legs of the ribbon and pulling one of them +/- 90 degrees to the right and the other +/- 90 degrees to the left of the patient's main axis, with the polypropylene thread being pulled in the direction of the patient's main axis and fixed in the inferior angle of the surgical wound. RESULTS: From August 1981 to June 1999, 609 patients had their arteries revascularized without the use of ECC. Among this group, 147 patients (24.14%) had the circumflex artery revascularized. Of the 609 patients, 48 (7.88%) presented serious complications and 21 (3.44%) died. CONCLUSIONS: It was possible to modify the anatomical position of the heart by exposing the circumflex artery with its marginal obtuse branches and performing anastomosis in a simple manner with no loss of quality or hemodynamic involvement, permitting a complete revascularization without the aid of extracorporeal circulation. PMID- 12125670 TI - Aortic and mitral valve surgery on the beating heart is lowering cardiopulmonary bypass and aortic cross clamp time. AB - OBJECTIVE: The concept of cardiac surgery on the beating heart is acceptable rationale for the cardiac surgery in the next millenium. Beating heart (off-pump) coronary artery bypass grafting (CABG) techniques have led us to consider the possibility for performing the aortic and mitral valve surgery (mitral valve repairs and replacements - with or without CABG) on the beating heart with the technique of retrograde oxygenated coronary sinus perfusion. METHODS: We used the technique of retrograde oxygenated blood coronary sinus perfusion in 78 patients (Group All) - (36 patients were with extremely low ejection fraction (Group X) - 62% of whom were in New York Heart Association (NYHA) class 4 and 34% of whom were in NYHA class 3). The procedures for the patients were: aortic, mitral and tricuspid valve surgery, in combination with CABG in ischemic patients. CABG was done in all the cases off-pump. In addition, we performed a case match study for 37 patients with good ejection fraction (51.65 +/- 11.88) (Beating Heart Group) operated on the beating heart with most appropriate group of patients (No. 37) operated in our institutions on arrested heart (ejection fraction 51.07 +/- 12.93) (Arrested Heart Group). The case match selection criteria were: gender, left ventricular ejection fraction, atrial fibrillation, hypertension, pulmonary hypertension, and diabetes. The selected beating heart group and selected arrested heart groups were without statistically significant differences for the mentioned criteria. RESULTS: There were statistically significant differences between Beating Heart Group and Arrested Heart Group in the duration of Cardiopulmonary Bypass Time (69.35 +/- 13.52 min. versus 93.59 +/- 28.54 min.), p<0.001, and statistically significant differences in Aortic Cross Clamp Time (46.5 +/- 8.95 min. versus 61.5 +/- 18.34 min.), p<0.001. The values for Creatinin Kinase (CK) and LDH were not statistically different, however the absolute values for Beating Heart Group were lower. There was no statistical difference in complication rate for both the groups for: sternal infection, bleeding, death, atrial fibrillation, AV block and neurological complications. The total early mortality for all the patients was 5.1% (4 out of 78) - for the group X 8.3 % (3 of 36 patients). Two were in-hospital deaths. One patient with triple-vessel disease and acute mitral insufficiency on intra aortic balloon pump (IABP) had been operated on 6 days after acute myocardial infarction (AMI). The cause of the death was systemic meticillin resistant staphylococus aureus (MRSA) infection - (eight days prior to our operation, arthrodesis of the talocrural joint was performed by an orthopedic surgeon). The other death was a female patient who was operated on after previous multiple cerebrovascular infarctions (CVI) (cause of the death was CVI). In addition, one patient died one month after the operation because of prosthetic valve endocarditis (PVE) on aortic and mitral valves (silver-coated silzone aortic and mitral valves were implanted because of chronic latent asymptomatic tibial osteitis). None of these deaths were cardiac related. CONCLUSIONS: We conclude that beating heart valve surgery (any combination) with or without CABG significantly lower the cardiopulmonary bypass and aortic cross clamp time. In addition, the advantages of beating-heart surgery are 1) the perfused myocardial muscle, 2) the heart is not doing any work, 3) no reperfusion injury, 4) the possibility for ablation of atrial fibrillation on the beating heart, and 5) testing of the mitral valve repair is done in real physiologic conditions in the state of left ventricle beating tonus. The procedure could be the procedure of choice for the valve operation or combined operations (valve operation and CABG) in high-risk patients with low ejection fractions. There is no doubt that at present day in cardiac surgery exist at least two major factors for mortality and morbidity after cardiac surgery, which are operation - related, namely cardiopulmonary bypass time and its duration and aortic cross clamp time (ischemic time of myocardium). In the last few years a number of different techniques emerged in the field of cardiac surgery, which were directed toward better results in the selected high risk patients or to minimize the deleterious effects of cardiopulmonary bypass (CPB) on the overall postoperative performance [Calafiore 1996, Tasdemir 1998]. Due to the fact, that the cardiac muscle should be protected at most during the cardiac arrest, retrograde blood cardioplegia was successfully introduced [Buckberg 1990], and more - the warm cardioplegia is being used recently [Kawasuji 1997]. The natural status of the human heart is the beating status, so it is reasonable to try to perform the operations on the beating heart. This has been done recently with the MID - CAB and OP - CAB (off-pump CABG) operations [Tasdemir 1998]. The retrograde warm blood cardioplegia has therefore led us to the premise, that with retrograde oxygenated blood perfusion it would be possible to achieve the operations on the beating heart even in the open heart surgery, such as aortic and/or mitral valve surgery. All will agree that the most damaging effect of the cardioplegia is the reperfusion injury [Allen 1997], and it is obvious that with the technique of retrograde continuous oxygenated blood perfusion this effect will be canceled. In this article, we would like to show the how-to technique for the operations on the beating heart in the case of operations on the aortic valve replacement (AVR) with mitral valve repair (MVR) or replacement MVR and with/without concomitant coronary artery bypass (CABG) surgery. The tricuspid valve repair (PTV) is normally done on the beating heart and there it is realized what problems or technical difficulties may arise during procedures on the mitral valve: the walls of the ventricles are not flattened and the exposure of the mitral valve is challenging task. Furthermore, the free walls of the ventricles with interventricular septum are in the state of the tonus, so every force applied to better expose the aortic or mitral valve is not acceptable PMID- 12125669 TI - Early- and long-term comparison of the on- and off-pump bypass surgery in patients with left ventricular dysfunction. AB - OBJECTIVE: The adverse effects of extracorporeal circulation increase the morbidity and mortality risk of coronary bypass surgery, especially in patients with left ventricular dysfunction. The purpose of this study was to provide a comparison of the early and long-term outcome between patient groups with left ventricular dysfunction (LVEF<40% or LVPS>or=15) operated with or without using cardiopulmonary bypass. METHODS: Fifty-one patients with left ventricular dysfunction, who were operated on between October 1992 and March 1994, were investigated retrospectively. They were divided into two groups: BH-group included 26 patients and cardiopulmonary bypass group had 25 patients. Mean age and risk factors were identical. All patients received one vessel bypass left internal mammary artery to left descending artery. RESULTS: There was no early mortality and perioperative myocardial infarction in either group. In the early postoperative period the need of cardiac support therapy was significantly higher in the cardiopulmonary-bypass group than in the beating heart-group: 32% versus 7.7% (p<0.05). The need for blood products (for fresh frozen plasma 3.63 +/- 2.15u versus 2.5 +/- 1.34u; p = 0.023; for packed red blood cells 1.8 +/- 0.75u versus 1.25 +/- 0.46u; p = 0.048), the extubation time (18.2 +/- 5.5 hours versus 15.3 +/- 3.8 hours; p = 0.03) and the hospital stay (10.64 +/- 3.2 days versus 7.92 +/- 2.25 days; p = 0.001) were higher in the cardiopulmonary bypass -group than in the beating heart-group. Actuarial survival for the beating heart-group was 92.3 +/- 5.2% at 6 years, and for the cardiopulmonary bypass group was 92 +/- 5.4% at 6 years (p = 0.67). CONCLUSIONS: In spite of more than four times as many patients in the cardiopulmonary bypass group requiring inotropic support after surgery, survival and cardiac death rates were similar for both groups. Off-pump bypass surgery conserves the blood constituents. The benefits of both techniques to improve the left ventricular performance score and ejection fraction were similar, but postoperative extubation time, length of intensive care unit and hospital stay were reduced significantly in the beating heart group. With these good results of the beating heart coronary bypass surgery and considering its cost effectiveness, we concluded that coronary bypass on a beating heart can be an alternative to cardiopulmonary bypass technique in selective patient groups. PMID- 12125671 TI - Body surface mapping after partial left ventriculotomy. AB - OBJECTIVE: To demonstrate cardiac electrophysiological changes in patients where partial left ventriculotomy was performed and multichannel electrocardiographical measurements and body surface potential mapping were used. METHODS: Body surface ECG signals were recorded during sinus rhythm for one minute. Six patients were operated on with partial left ventriculotomy were monitored. All patients had normal coronary angiography data. The data were acquired prior to the partial left ventriculotomy, and on the second, third, fourth, and fifth postoperative day using 32-body surface leads. The recorded data were analysed by determining ST-40 and QRS integral maps. The analysis was done on a set of selected beats during the sinus rhythm and on the averaged beats. RESULTS: Before the operation, ST-40 maps typically showed an area of strong positive potentials (elevation) over the anterior aspect of the torso and a region of strong negative potentials (depression) over the lateral, and posterior aspects of the torso. After the operation, the ST elevation over the anterior, lateral and posterior aspects of the torso was reduced. An area of marked positive potentials remained in the precordial area (overlying the excised area of the heart), even during the postoperative monitoring interval (day two through day five). We also noticed that the amplitude of cardiac signals decreased by approximately 30% after the partial left ventriculotomy. Qualitative map changes were substantiated by statistical parameters. CONCLUSIONS: Results of our study demonstrate that noninvasive acquisition of body-surface electrocardiographs may detect changes in the cardiac activity of patients undergoing partial left ventriculotomy. This finding suggests that body surface mapping may also be useful in assessing the arrhythmia vulnerability. PMID- 12125672 TI - Fatal granulomatous disease of childhood: presenting as acute abdomen. AB - Fatal granulomatous disease of childhood is a rare disorder of phagocytic function. We report a 6-year-old boy who presented with acute abdomen. The diagnosis was established by mesenteric lymph node biopsy obtained at laparotomy. The boy succumbed within hours of surgery. PMID- 12125673 TI - Report of the International Leprosy Association Technical Forum. Paris, France, 22-28 February 2002. PMID- 12125674 TI - Biodegradation of polycarbonate-based polyurethanes by the human monocytes derived macrophage and U937 cell systems. AB - The prominent cell type found on implanted medical devices during the chronic inflammatory response is the monocyte-derived macrophage (MDM). Using an activated in vitro cell system, it was possible to show that MDMs possess esterolytic activities that may contribute to the degradation of polyurethanes. In the present study, the U937 cell line was paralleled to the MDM cell system in order to validate the use of a cell line that could expedite studies on biomaterial biocompatibility and biostability. Using 12-o-tetradecanoylphorbol 13 acetate (PMA), the optimum differentiation time for the U937 cells was 72 h based on biodegradation, degradative potential, and (35)S-methionine uptake. After activation of the cells by resuspending from tissue culture polystyrene plates and reseeding onto a (14)C-labeled polycarbonate-based polyurethane(PCNU), both U937 cells and the MDMs elicited comparable radiolabel release (measure of polymer breakdown) and esterase activity (measure of degradative potential) at 48 h. There was no difference in the effect on radiolabel release and esterase activity elicited by both cell types with inhibitors of protein synthesis, esterase activity, and phospholipase A(2). This established that both cell types likely used similar hydrolytic activities and signaling pathways to cause degradation of the PCNU. Immunoblotting demonstrated that both cell systems secreted monocyte-specific esterase and cholesterol esterase enzymes previously shown to degrade PCNUs. The U937 cell system is more convenient and reproducible than MDMs for pursuing possible biological pathways elucidating the mechanism of polyurethane biodegradation. Once established with U937s, the pathways can then be validated with the more physiologically relevant human MDM cell system. PMID- 12125675 TI - Biological warfare -- a growing healthcare concern. PMID- 12125676 TI - Severe, acute asthma as therapeutic orphan. PMID- 12125678 TI - Comment: a need for more research. . PMID- 12125677 TI - The impact of chemotherapy on advanced neuroblastoma. Survival of patients diagnosed in 1956, 1962, and 1966-68 in Children's Cancer Study Group A. AB - Survival statistics from 50 children with disseminated neuroblastoma, treated according to two CCSGA protocols between 1966 and 1968, are compared to those collected by Sutow and associates for 1956 and 1962. Little improvement occurred in the median survival rates of the children in these groups. Survival at 24 months is 13, 22, and 13 per cent, respectively, for the three groups. The lack of improvement in the median survival rate following treatment with a combination of VCR and CPM is attributed to an initial response rate of less than 40 per cent in the treated patients. The survival of patients who did respond, when compared to those who did not, was twice as long (15 vs. 7 months). Clearly, improved chemotherapy or other modes of treatment are needed for patients with advanced neuroblastoma at the time of diagnosis. PMID- 12125679 TI - Abstracts of the 4th International Symposium of NeuroVirology and 10th Conference on Neuroscience of HIV Infection. 18-22 June 2002. Dusseldorf, Germany. PMID- 12125681 TI - Abstracts of the European Conference on Viral Diseases. May 10-12, 2002. Munich, Germany. PMID- 12125682 TI - European Association of Nuclear Medicine Congress. 14-18 September 1996, Copenhagen, Denmark. Abstracts. PMID- 12125680 TI - Recurrences of rectal cancers: results of a multimodal approach with intraoperative radiation therapy. French Group of IORT. Intraoperative Radiation Therapy. AB - PURPOSE: Prognosis of recurrent rectal cancer remains poor, mainly because of the difficulties of achieving a satisfactory local control. Intraoperative radiation therapy (IORT) allows for the delivery of a complementary single dose to the tumor residues or to the tumor bed and could be useful jn a multimodal treatment. In an attempt to evaluate this interest, a retrospective analysis of patients treated with IORT in six French hospitals has been performed. METHODS AND MATERIALS: Data have been collected in 73 patients (41 men), with a mean age of 62 years, treated with IORT. Initial rectal tumors were large (mean diameter: 45 mm), partially or totally fixed to the contiguous structures in 39%, and with nodal involvement in 50% of the cases. Initial surgery had been a sphincter sparing surgery in 67%; external radiation therapy had been delivered in 52%, and a chemotherapy had been given in 10% of the patients. Recurrences were isolated (without metastases) in 86%, and were posterior or posterolateral in 55% of the cases. Surgery allowed for a complete macroscopical resection in 57%, a partial resection with gross residual disease in 29%, and no resection in 14% of the recurrences. Intraoperative radiation therapy was delivered in a dose of 10 to 25 Gy (mean 18.5) through localizators of a mean diameter of 75 mm (60 to 110). External radiation therapy, either preoperative or postoperatively was given to 30 patients without prior radiation therapy. Ten patients received additional chemotherapy with 5-fluorouracil. RESULTS: Four postoperative deaths occurred. Postoperative morbidity occurred in 16 patients and some complications were probably related to the IORT procedure. Four long-term complications were observed. Overall actuarial survival occurred in 72.4% of the patients at 1 year, in 44.6% at 2 years, and in 30.6% at 3 years. Twenty-one local failures have been observed. Actuarial local control occurred in 71.3% of the patients at 1 year, 47.7% at 2 years, and 31.3% at 3 years. CONCLUSION: Intraoperative radiation therapy is a complementary treatment for recurrences of rectal cancer. It provides encouraging results, particularly in some selected situations, when patients have not previously been treated with external radiation therapy. Further studies of multimodal treatments are necessary. PMID- 12125683 TI - vCJD: the epidemic that never was. Creutzfeldt's patient did not have Creutzfeldt Jakob disease. PMID- 12125684 TI - vCJD: the epidemic that never was. Possibility of BSE being cause of variant CJD is indeed biologically plausible. PMID- 12125685 TI - vCJD: the epidemic that never was. Author has overlooked several findings that support his argument. PMID- 12125686 TI - Has medicalisation of childbirth gone too far? Timely intervention is the key. PMID- 12125687 TI - Has medicalisation of childbirth gone too far? Childbirth in Taiwan is certainly overmedicalised. PMID- 12125688 TI - Evolving general practice consultation in Britain. Longer consultations might necessitate redeployment of pharmacists. PMID- 12125689 TI - Abstracts of the 7th Meeting of the European Society of Neurosonology and Cerebral Hemodynamics. Bern, Switzerland, May 26-28, 2002. PMID- 12125690 TI - Abstracts of the 11th European Stroke Conference. Geneva, Switzerland, May 29 June 1, 2002. PMID- 12125691 TI - From the Centers for Disease Control and Prevention. Trends in cigarette smoking among high school students--United States, 1991-2001. PMID- 12125692 TI - From the Centers for Disease Control and Prevention. State-specific mortality from stroke and distribution of place of death--United States, 1999. PMID- 12125693 TI - Gerrit Dou: 17th century Dutch artist portraying dentists at work. AB - The Dutch painter, Gerrit Dou (1613-1675), was, during his lifetime, a prestigious and highly paid artist. Critics and collectors of that era marveled at his extraordinary technical skill, his masterful illusionism and his keen portrayal of everyday life. He was the first artist to perfect the technique of "fine painting"--the production of small, detailed pictures wherein subjects are painted in a photographically realistic style. Because of his exceptional skill in this technique, the brushwork in his painting was almost invisible. Additionally, he masterfully used light and shade to achieve the effect of a third dimension. In many of his portraits, subjects stare ahead from inside a window (or niche), whose sill is overflowing with small, familiar objects. Dou's focus was on 17th century Holland and his work included: portraits, genre (scenes of everyday life) and religious themes. Between 1628 and 1675, he painted about three hundred pictures: six of these portrayed dentists at work. Most artists of those days depicted dentists as plying their trade in theatrical, circus-like environments, as in crowded county fairs. However, Dou's dentists interact with their patients in a realistic, professional manner. In this article, his most famous dental painting, "The Dentist," created in 1672, is analyzed in detail. Over the centuries, Dou's public recognition has faded into obscurity. However, his exceptional talents are currently being rediscovered. PMID- 12125694 TI - Correspondence between Paul N. Baer and Sigurd P. Ramford. PMID- 12125695 TI - Gleanings about dentistry from the world of literature (Twenty-seventh in a series). PMID- 12125696 TI - Oliver Wendell Holmes, Sr., MD: champion of dentistry. PMID- 12125697 TI - A brief history of aerospace dentistry. AB - In April 2000, the National Academy of Sciences Institute of Medicine (NAS/IOM) Committee on Space Medicine held a workshop under contract with the National Aeronautics and Space Administration (NASA) to explore "innovative terrestrial medical care." There was also a NAS/IOM panel held on "Space Dentistry: Maintaining Astronauts' Oral Health on Long Missions." Air Force Dental Officer Col. Shannon E. Mills chaired the dental committee. Many questions were raised but few answers were available. Prevention was emphasized with the hope that within twenty to thirty years there may be a number of astronaut candidates with no existing dental restorations and with optimum oral health. However, there remains the concern that trauma to teeth could occur within the confines of a zero gravity space capsule as crew members carry out their daily responsibilities. The possibility is evident considering the duration of a space flight to Mars and back could require up to three years. The dental concerns of a space mission are only a small part of a much larger team effort, however, it is one not to be overlooked. An historical review of dentistry's involvement with America's flight and space programs of the 20th Century would be prudent. Many of same questions asked today were addressed in the early days of aviation dentistry as it transitioned into aerospace dentistry. Any past research and experiences would help serve as a foundation to build upon. PMID- 12125698 TI - Dental postcards XXIV. PMID- 12125699 TI - The hidden poetry of Solyman Brown, the "poet laureate of dentistry". AB - One of the most important figures in the development of the dental profession was Solyman Brown, in whose home the leading dentists of his day met and organized the first national dental organization in the world as well as the first dental journal in the world. He was named the first secretary of the organization and served, for two years, as the first editor of the journal. A true Renaissance man, Brown was also a consummate, and widely published, poet, and his epic poem, Dentologia, was hailed by the critics of his day as a literary masterpiece, earning him the sobriquet, "Poet Laureate of Dentistry." Recently Solyman Brown's great-great grandson, cleaning out an attic, discovered a trunk full of material relating to Brown's life - his daughters' memoirs, many of his letters and other of his writings, and many of his unpublished poetical works. This author was fortunate to be able to study all this material, and the poems published herein give a much fuller and rounded picture of this monumental figure in American, and world, dentistry. PMID- 12125700 TI - 19th Century dentistry advertising trade cards. PMID- 12125701 TI - Chapin A. Harris' forceps: the Arnold Ruby set. PMID- 12125702 TI - Sources for literature searches in dentistry. PMID- 12125703 TI - Dentistry on stamps. PMID- 12125704 TI - Ischemia/reperfusion injury. PMID- 12125705 TI - [Alfred Eduard Heinrich Bruggemann (1882 - 1971)]. PMID- 12125706 TI - [Medical history and politics: careers of Fritz Lejeune during the Weimar Republic and national socialism]. PMID- 12125707 TI - [Orthopedic technology in Switzerland]. PMID- 12125708 TI - [The care of the sick monk in medieval monasteries]. PMID- 12125709 TI - Downstreamers: public health and relationships on the Missouri River. PMID- 12125710 TI - Mitochondrial DNA and nucleoside toxicity. PMID- 12125711 TI - Mitochondrial DNA and nucleoside toxicity. PMID- 12125712 TI - Male cells in female recipients of hematopoietic-cell transplants. PMID- 12125713 TI - Male cells in female recipients of hematopoietic-cell transplants. PMID- 12125714 TI - Male cells in female recipients of hematopoietic-cell transplants. PMID- 12125715 TI - Medical devices; apnea monitor; special controls. Final rule. AB - The Food and Drug Administration (FDA) is issuing a final rule to create a separate classification for the apnea monitor. The device currently is included in the generic type of device called breathing frequency monitors. The apnea monitor will remain in class II, but will be subject to a special control. The special control is an FDA guidance document that identifies minimum performance, testing, and labeling recommendations for the device. Following the effective date of this final classification rule, any firm submitting a 510(k) premarket notification for a "new" apnea monitor will need to address the issues covered in the special control guidance. However, the firm need only show that its device meets the recommendations of the guidance or in some other way provides equivalent assurances of safety and effectiveness. Elsewhere in this issue of the Federal Register, FDA is announcing the availability of the guidance document that will serve as the special control. FDA is taking these actions because it believes that they are necessary to provide reasonable assurance of the safety and effectiveness of the apnea monitor. PMID- 12125716 TI - Medical devices; reclassification of polymethylmethacrylate (PMMA) bone cement. Final rule. AB - The Food and Drug Administration (FDA) is announcing that it has reclassified the polymethylmethacrylate (PMMA) bone cement intended for use in arthroplastic procedures of the hip, knee, and other joints for the fixation of polymer or metallic prosthetic implants to living bone from class III to class II (special controls). The agency is also announcing that it has issued an order in the form of a letter to the Orthopedic Surgical Manufacturers Association (OSMA) reclassifying the device. The special control for the device is a guidance document entitled "Class II Special Controls Guidance Document: Polymethylmethacrylate (PMMA) Bone Cement." The agency is reclassifying this device into class II because special controls, in addition to general controls, would provide reasonable assurance of the safety and effectiveness of the device, and there is sufficient information to establish special controls. PMID- 12125717 TI - Assumptions and principles about psychosocial aspects of disasters. PMID- 12125718 TI - Grief work versus continuing bonds: a call for paradigm integration or replacement? AB - In this paper we compare grief work and continuing bond models of grief to determine if one explains the data better than the other. Sixty individuals in active grief completed a questionnaire in which they were asked to rate their grief status, perceived similarity to the deceased along 7 dimensions, and closeness of relationship to the deceased. A matched control group was also asked to rate the closeness of relationship and perceived similarity to a living person of the same relationship as the deceased to the griever. In line with grief work, we found that perceived similarity was directly related to severity of grief and that perceived closeness of relationship declined over time. In support of continuing bonds, however, perceived similarity did not decline over time, nor was overall perception of similarity among grievers different from their matched controls. PMID- 12125719 TI - Current bibliography in the history of technology (1999). PMID- 12125720 TI - Pathology quiz. Asymptomatic bacteriuria in the elderly. PMID- 12125721 TI - Radiology quiz. Soft tissue mass arising in the thigh. PMID- 12125722 TI - An accidental acid burn. Providing emergency care. PMID- 12125723 TI - Inhibition of retinoid activity by components of a paper mill effluent. AB - A cell line stably transfected with reporter genes activated by retinoic acid was used to test a paper mill effluent for the presence of retinoids or components that interfere with retinoic acid-stimulated gene transcription. No retinoids were detected in effluent or control water. However, effluent water significantly decreased reporter activity stimulated by all-trans-retinoic acid, while activity stimulated by 9-cis-retinoic acid was unaffected. In a limited fractionation through a C-18 solid phase-exchange column the inhibitory activity was retained in the aqueous fraction, indicating that the activity was polar. PMID- 12125724 TI - Correlations between toxic Pb effects and production of Pb-induced thiol peptides in the microalga Stichococcus bacillaris. AB - Comparing Pb toxicity to the green alga Stichococcus bacillaris and production of Pb-induced thiol peptides in 24-h exposed cells made it possible to show the level of these peptides to reflect Pb availability to algal cells. LC-ESI/MS analysis of these peptides confirmed that they are phytochelatins PC2-PC4 and their des-Gly derivatives. Studies were carried out at environmentally relevant pH values (5-8.5) and various concentrations of hardness cations (Ca, Mg) and orthophosphate: (0-0.1 mM), chloride (0-100 mM), citrate (0-1 mM), and humic acids (0-10 mg/l). It has been stated, on the basis of the level of Pb-induced thiol peptides, that Pb availability to algae changed significantly within the range of the pH values studied, and was limited in the presence of calcium and of some complexing ligands like orthophosphate, chloride and citrate; it did not decrease in the presence of magnesium or humic acid. PMID- 12125725 TI - Comparative study of the suitability of three lichen species to trace-element air monitoring. AB - To investigate the suitability of three lichen species (Cetraria islandica, Evernia prunastri, and Ramalina farinacea) as transplants to trace-element air biomonitoring, they were exposed on substratum-free supports, from July 1996 until July 1997, in three European countries with different climates (Germany, Italy, Romania), at six sites with different types of air pollutants (two in each country). After 2, 4, 6, and 12 months of exposure, some portions of thallus were collected, prepared, and measured by instrumental neutron activation analysis (INAA) at the Institute of Physics and Nuclear Engineering in Bucharest and by energy dispersive X-ray fluorescence analysis (EDXRFA) at the University of Hohenheim in Stuttgart. Fifteen environmentally relevant elements: As, Br, Ca, Co, Cr, Cu, Fe, K, Mn, Ni, Pb, S, Sb, V, and Zn were determined. The analytical results were compared statistically. To study the distribution of the trace elements between the lichens and the lichen throughfall water inside a virtual column, the throughfall water was collected under the lichen transplants during 6 and 12 months. The dried residues were analysed by INAA at Bucharest. The accumulating capacity for all investigated species is evident. For a comparative evaluation, the initial element contents, the "accumulation factors" relative to the bulk deposition, the interspecies "calibration factors", and the "retention efficiencies", defined as ratios of the lichen enrichment to the sum of this enrichment and the content in the lichen throughfall water, were considered. These criteria attest the best suitability for Evernia prunastri, followed by Ramalina farinacea and Cetraria islandica. PMID- 12125726 TI - Bioremediation of diesel-contaminated soil with composting. AB - The major objective of this research was to find the appropriate mix ratio of organic amendments for enhancing diesel oil degradation during contaminated soil composting. Sewage sludge or compost was added as an amendment for supplementing organic matter for composting of contaminated soil. The ratios of contaminated soil to organic amendments were 1:0.1, 1:0.3, 1:0.5, and 1:1 as wet weight basis. Target contaminant of this research was diesel oil, which was spiked at 10,000 mg/kg sample on a dry weight basis. The degradation of diesel oil was significantly enhanced by the addition of these organic amendments relative to straight soil. Degradation rates of total petroleum hydrocarbons (TPH) and n alkanes were the greatest at the ratio of 1:0.5 of contaminated soil to organic amendments on wet weight basis. Preferential degradation of n-alkanes over TPH was observed regardless of the kind and the amount of organic amendments. The first order degradation constant of n-alkanes was about twice TPH degradation constant. Normal alkanes could be divided in two groups (C10-C15 versus C16-C20) based on the first order kinetic constant. Volatilization loss of TPH was only about 2% of initial TPH. Normal alkanes lost by volatilization were mainly by the compounds of C10 to C16. High correlations (r=0.80-0.86) were found among TPH degradation rate, amount of CO2 evolved, and dehydrogenase activity. PMID- 12125727 TI - Heavy metals in agricultural soils of the Pearl River Delta, South China. AB - There is a growing public concern over the potential accumulation of heavy metals in agricultural soils in China owing to rapid urban and industrial development and increasing reliance on agrochemicals in the last several decades. Excessive accumulation of heavy metals in agricultural soils may not only result in environmental contamination, but elevated heavy metal uptake by crops may also affect food quality and safety. The present study is aimed at studying heavy metal concentrations of crop, paddy and natural soils in the Pearl River Delta, one of the most developed regions in China. In addition, some selected soil samples were analyzed for chemical partitioning of Co, Cu, Pb and Zn. The Pb isotopic composition of the extracted solutions was also determined. The analytical results indicated that the crop, paddy and natural soils in many sampling sites were enriched with Cd and Pb. Furthermore, heavy metal enrichment was most significant in the crop soils, which might be attributed to the use of agrochemicals. Flooding of the paddy soils and subsequent dissolution of Mn oxides may cause the loss of Cd and Co through leaching and percolation, resulting in low Cd and Co concentrations of the paddy soils. The chemical partitioning patterns of Pb, Zn and Cu indicated that Pb was largely associated with the Fe-Mn oxide and residual fractions, while Zn was predominantly found in the residual phase. A significant percent fraction of Cu was bound in the organic/sulphide and residual phases. Based on the 206Pb/207Pb ratios of the five fractions, it was evident that some of the soils were enriched with anthropogenic Pb, such as industrial and automobile Pb. The strong associations between anthropogenic Pb and the Fe-Mn oxide and organic/sulphide phases suggested that anthropogenic Pb was relatively stable after deposition in soils. PMID- 12125728 TI - Phytoremediation potential of Spirulina (Arthrospira) platensis: biosorption and toxicity studies of cadmium. AB - This study examines the possibility of using Spirulina (Arthrospira) platensis TISTR 8217 to remove low concentrations of cadmium (less than 100 mg/l) from wastewater. The cyanobacteria were exposed to six different cadmium concentrations for 96 h, and the growth rate was determined using an optical density at 560 nm. The inhibiting concentration (IC50) was estimated using probit analysis. The IC50 at 24, 48, 72, and 96 h were 13.15, 16.68, 17.28, and 18.35 mg/l Cd, respectively. Cellular damage was studied under a light microscope and a transmission electron microscope. Swollen cells and fragmented filaments were observed. Cell injury increased with increasing concentrations of cadmium. Ultrastructural changes were observed in the algae exposed to cadmium concentrations both close to IC50 (14.68 mg/l) and at IC50 (18.35 mg/l). The alterations induced by cadmium were disintegration and disorganization of thylakoid membranes, presence of large intrathylakoidal space, increase of polyphosphate bodies, and cell lysis. In addition, the cadmium adsorption by algal cells was studied. Environmental factors were found to have an effect on biosorption. The uptake of cadmium was not affected by the temperature of the solution, but the sorption was pH dependent. The optimum pH for biosorption of algal cells was 7. The cadmium uptake process was rapid, with 78% of metal sorption completed within 5 min. The sorption data fit well to the Langmuir isotherm. The maximum adsorption capacity for S. platensis was 98.04 mg Cd per g biomass. PMID- 12125729 TI - Relations between crown condition and ozone and its dependence on environmental factors. AB - It is now widely accepted that stomatal ozone uptake provides a more reliable indicator for the assessment of risk of ozone damage to forest trees than ozone exposure. Thus, environmental factors influencing stomatal opening strongly affect the sensitivity of forest trees towards high ozone concentrations. In this study the hydrological model WAWAHAMO is used to simulate stomatal opening dependent on air temperature, solar radiation, vapor pressure deficit and soil water potential. Then, a statistical analysis was carried out to investigate relations between crown defoliation and ozone concentrations dependent on stomatal opening. The results support the hypothesis that tropospheric ozone contributes to the observed crown defoliation. However, environmental factors strongly affect tree response to high ozone concentrations. PMID- 12125730 TI - Polychlorinated naphthalenes and coplanar polychlorinated biphenyls in beluga whale (Delphinapterus leucas) and ringed seal (Phoca hispida) from the eastern Canadian Arctic. AB - Blubber collected from beluga whales and ringed seals during subsistence hunts in the southern Baffin Island region of the Canadian Arctic were analysed for polychlorinated naphthalenes and eight planar PCB congeners (mono-ortho PCBs: 105, 114, 118 and 156; non-ortho PCBs: 77, 81, 126, 169). SigmaPCN (3-7 Cl) concentrations in blubber ranged from 35.9-383 pg/g (lipid weight; lw) in beluga and 35.4-71.3 pg/g (lw) in ringed seal. These represent the first measurements of PCNs in marine mammals in the Canadian Arctic, mammals which are an important part of the traditional diet of the indigenous population. SigmaCoplPCB concentrations were much higher, ranging from 15.5-317 ng/g (lw) in beluga whale blubber and 16.5-40.9 ng/g (lw) in ringed seal blubber. PCNs and coplanar PCBs both exhibit dioxin-like toxicity. Although average sigmaPCN concentrations were less than 1% of sigmaCoplPCBs, PCNs contribute up to 11% of TEQ relative to the coplanar PCBs based on TEFs determined by H4IIE enzyme assays. PMID- 12125731 TI - Introduction: Fluxes and impacts of atmospheric ammonia on national, landscape and farm scales. PMID- 12125732 TI - Seasonal variation and background levels of heavy metals in two green seaweeds. AB - Seasonal variation in the contents of different metals (Al, Cr, Cu, Fe, Mn, Ni and Zn) in two genera of macroalgae, Ulva and Enteromorpha was studied at 22 sites on the northwest coast of Spain. The seasonal variation in the different metals followed a similar pattern in both seaweeds and appeared to be caused by dilution during the period of maximum growth and concentration during periods of slow growth. Fluvial inputs of Al, Fe and Mn in autumn and winter appeared to accentuate the latter effect: the concentrations of these three metals in both macroalgae and of Cr in Enteromorpha were highest at those sites most influenced by inputs from rivers. The background levels of Cr, Cu, Ni and Zn in the algae in summer and winter were established. PMID- 12125733 TI - Defining the spatial impacts of poultry farm ammonia emissions on species composition of adjacent woodland groundflora using Ellenberg Nitrogen Index, nitrous oxide and nitric oxide emissions and foliar nitrogen as marker variables. AB - The marker variables, Ellenberg Nitrogen Index, nitrous oxide and nitric oxide fluxes and foliar nitrogen, were used to define the impacts of NH3 deposition from nearby livestock buildings on species composition of woodland ground flora, using a woodland site close to a major poultry complex in the UK. The study centred on 2 units in close proximity to each other, containing 350,000 birds, and estimated to emit around 140,000 kg N year(-1) as NH3. Annual mean concentrations of NH3 close to the buildings were very large (60 microg m(-3)) and declined to 3 microg m(-3) at a distance of 650 m from the buildings. Estimated total N deposition ranged from 80 kg N ha(-1) year(-1) at a distance of 30 m to 14 kg N ha(-1) year(-1) at 650 m downwind. Emissions of N2O and NO were 56 and 131 microg N m(-2) h(-1), respectively at 30 m and 13 and 80 microg N m( 2) h(-1), respectively at 250 m downwind of the livestock buildings. Species number in woodland ground flora downwind of the buildings remained fairly constant for a distance of 200 m from the units then increased considerably, doubling at a distance of 650 m. Within the first 200 m downwind, trends in plant species composition were hard to discern because of variations in tree canopy composition and cover. The mean Ellenberg N Index ranged from 6.0 immediately downwind of the livestock buildings to 4.8 at 650 m downwind. The mean abundance weighted Ellenberg N Index also declined with distance from the buildings. Tissue N concentrations in trees, herbs and mosses were all large, reflecting the substantial ammonia emissions at this site. Tissue N content of ectohydric mosses ranged from approximately 4% at 30 m downwind to 1.6% at 650 m downwind. An assessment of the relative merits of the three marker variables concludes, that while Ellenberg Index and trace gas fluxes of N2O and NO give broad indications of impacts of ammonia emissions on woodland vegetation, the application of a critical foliar N content for ectohydric mosses is the most useful method for providing spatial information which could be of value to policy developers and planners. PMID- 12125734 TI - Distribution of polycyclic aromatic hydrocarbons in the sediments of the Adriatic Sea. AB - Distribution of the sixteen polycyclic aromatic hydrocarbons (PAHs) indicated from USEPA as priority pollutants was studied in surface sediments of two coastal areas of the Adriatic Sea. PAHs were recovered from the sediments by solvent extraction and then analyzed by means of gas chromatography-mass spectrometry. Total concentrations of the analytes in the range 24.1-501.1 ng/g were detected. The observed distribution has been discussed taking into account different aspects, such as the fluvial inputs and the grain size of the sediments. By using a molecular marker approach and characteristic compositional patterns it was possible to ascribe to combustion processes the main source of PAHs. Furthermore a good correlation between benzo[a]pyrene and the total concentration of PAHs (r=0.953) has been pointed out. PMID- 12125735 TI - Predictive mapping of air pollution involving sparse spatial observations. AB - A limited number of sample points greatly reduces the availability of appropriate spatial interpolation methods. This is a common problem when one attempts to accurately predict air pollution levels across a metropolitan area. Using ground level ozone concentrations in the Tucson, Arizona, region as an example, this paper discusses the above problem and its solution, which involves the use of linear regression. A large range of temporal variability is used to compensate for sparse spatial observations (i.e. few ozone monitors). Gridded estimates of emissions of ozone precursor chemicals, which are developed, stored, and manipulated within a geographic information system, are the core predictor variables in multiple linear regression models. Cross-validation of the pooled models reveals an overall R2 of 0.90 and approximately 7% error. Composite ozone maps predict that the highest ozone concentrations occur in a monitor-less area on the eastern edge of Tucson. The maps also reveal the need for ozone monitors in industrialized areas and in rural, forested areas. PMID- 12125736 TI - Gangliogliomas: an intriguing tumor entity associated with focal epilepsies. AB - Gangliogliomas represent the most frequent tumor entity in young patients suffering from chronic focal epilepsies. In a series of 326 gangliogliomas collected from the University of Bonn Epilepsy Surgery Program and other departments of neuropathology in Germany, Austria, and Switzerland, epidemiological findings and histopathological hallmarks of gangliogliomas are systematically reviewed. The majority of these tumors occur within the temporal lobe and reveal a biphasic histological architecture characterized by a combination of dysplastic neurons and neoplastic glial cell elements. However, gangliogliomas exhibit a considerable variability in their histopathological appearance. Immunohistochemical studies are an important tool to discriminate these neoplasms from other tumor entities. Almost 80% of gangliogliomas reveal immunoreactivity for CD34, a stem cell epitope not expressed in normal brain. Immunohistochemical reactions for MAP2 or NeuN can be employed to characterize the dysplastic nature of neurons in those areas difficult to discriminate from pre-existing brain parenchyma. Less than 50% of the cases display binucleated neurons. With the frequent finding of "satellite" tumor clusters in adjacent brain regions, gangliogliomas are microscopically less circumscribed than previously assumed. The distinction from diffusely infiltrating gliomas is of considerable importance since tumor recurrence or malignant progression are rare events in gangliogliomas. Only little is known about the molecular pathogenesis of these glioneuronal tumors. Our findings support a dysontogenic origin from a glioneuronal precursor lesion with neoplastic, clonal proliferation of the glial cell population. Candidate genes appear to associate with neurodevelopmental signaling cascades rather than cell cycle control or DNA repair mechanisms. The reelin signaling and tuberin/insulin growth receptor pathways have recently been implicated in ganglioglioma development. Powerful new molecular genetic and biological tools can now be employed to unravel the pathogenesis of these intriguing lesions. PMID- 12125737 TI - Galectin-1 modulates human glioblastoma cell migration into the brain through modifications to the actin cytoskeleton and levels of expression of small GTPases. AB - We show that high-grade astrocytic tumors with high levels of galectin-1 expression are associated with dismal prognoses. The immunohistochemical analysis of galectin-1 expression of human U87 and U373 glioblastoma xenografts from the brains of nude mice revealed a higher level of galectin-1 expression in invasive areas rather than non-invasive areas of the xenografts. Nude mice intracranially grafted with U87 or U373 cells constitutively expressing low levels of galectin-1 (by stable transfection of an expression vector containing the antisense mRNA of galectin-1) had longer survival periods than those grafted with U87 or U373 cells expressing normal levels of galectin-1. Galectin-1 added to the culture media markedly and specifically increased cell motility levels in human neoplastic astrocytes. These effects are related to marked modifications in the organization of the actin cytoskeleton and the increase in small GTPase RhoA expression. All the data obtained indicate that galectin-1 enhances the migratory capabilities of tumor astrocytes and, therefore, their biological aggressiveness. PMID- 12125738 TI - Subtractive expression cloning reveals high expression of CD46 at the blood-brain barrier. AB - A subtractive expression cloning methodology was used to identify proteins having enriched expression at the blood-brain barrier (BBB) in comparison to liver and kidney tissues. A bovine brain capillary COS-1 cell cDNA expression library was screened with a BBB-specific antiserum. This strategy revealed that the membrane cofactor protein CD46, which is a regulator of complement activation in vivo and is also a potential measles virus receptor, is highly expressed at the BBB. The selective CD46 expression in brain at the BBB was confirmed by Northern blot analysis and confocal microscopy. The finding of selective expression of CD46 at the BBB is consistent with an important role played by the microvasculature in the immune surveillance of the brain. PMID- 12125739 TI - Cerebral vasculature is the major target of oxidative protein alterations in bacterial meningitis. AB - We have previously shown that antioxidants such as a-phenyl-tert-butyl nitrone or N-acetylcysteine attenuate cortical neuronal injury in infant rats with bacterial meningitis, suggesting that oxidative alterations play an important role in this disease. However, the precise mechanism(s) by which antioxidants inhibit this injury remain(s) unclear. We therefore studied the extent and location of protein oxidation in the brain using various biochemical and immunochemical methods. In cortical parenchyma, a trend for increased protein carbonyls was not evident until 21 hours after infection and the activity of glutamine synthetase (another index of protein oxidation) remained unchanged. Consistent with these results, there was no evidence for oxidative alterations in the cortex by various immunohistochemical methods even in cortical lesions. In contrast, there was a marked increase in carbonyls, 4-hydroxynonenal protein adducts and manganese superoxide dismutase in the cerebral vasculature. Elevated lipid peroxidation was also observed in cerebrospinal fluid and occasionally in the hippocampus. All of these oxidative alterations were inhibited by treatment of infected animals with N-acetylcysteine or alpha-phenyl-tert-butyl nitrone. Because N-acetylcysteine does not readily cross the blood-brain barrier and has no effect on the loss of endogenous brain antioxidants, its neuroprotective effect is likely based on extraparenchymal action such as inhibition of vascular oxidative alterations. PMID- 12125740 TI - Neutralizing antibodies to IL-18 ameliorate experimental autoimmune neuritis by counter-regulation of autoreactive Th1 responses to peripheral myelin antigen. AB - Experimental autoimmune neuritis (EAN) is a demyelinating disease of the peripheral nervous system (PNS). This acute inflammatory disease is mediated by CD4+ T cells and bears significant similarities to the Guillain-Barre syndrome of humans. In the present study, we investigated the function of IL-18 in T cell mediated autoimmunity of EAN in mice induced by P0 peptide 180-199 and Freund's complete adjuvant. Our data indicate that in 2 different therapeutic regimens, anti-IL-18 monoclonal antibody (mAb) effectively ameliorates the clinical and pathological signs of EAN. The suppression is associated with reduced inflammatory cell infiltration into the PNS and an insufficiency of autoreactive Th1 cells, as reflected by a reduced mononuclear cell proliferation and IFN-gamma secretion in the spleen. Increased numbers of IL-4 expressing cells and decreased numbers of IFN-gamma and TNF-alpha expressing cells were found in the PNS. Our results suggest that shifting the Th1/Th2 balance towards Th2 cells may be one mechanism underlying EAN suppression by anti-IL-18 mAb. In addition, anti-IL-18 mAb treatment reduced anti-P0 peptide 180-199 autoantibody responses, which may also contribute to EAN suppression. We conclude that endogenous IL-18 plays a critical role in the pathogenesis of autoimmune demyelinating disease of the PNS and that IL-18 antagonists may provide a new therapy for these diseases. PMID- 12125742 TI - Mitochondrial DNA deletions/rearrangements in parkinson disease and related neurodegenerative disorders. AB - Inhibition of mitochondrial respiratory chain function may contribute to dopaminergic neurodegeneration in the substantia nigra (SN) of patients with Parkinson disease (PD). Since large-scale structural changes (e.g. deletions and rearrangements in mitochondrial DNA [mtDNA]) have been associated with mitochondrial dysfunction, we tested the hypothesis that increased total mtDNA deletions/rearrangements are associated with neurodegeneration in PD. This study employed a well-established technique, long-extension polymerase chain reaction (LX-PCR), to detect the multiple mtDNA deletions/rearrangements in the SN of patients with PD, multiple system atrophy (MSA), dementia with Lewy bodies (DLB), Alzheimer disease (AD), and age-matched controls. We also compared the total mtDNA deletions/rearrangements in different brain regions of PD patients. The results demonstrated that both the number and variety of mtDNA deletions/rearrangements were selectively increased in the SN of PD patients compared to patients with other movement disorders as well as patients with AD and age-matched controls. In addition, increased mtDNA deletions/rearrangements were observed in other brain regions in PD patients, indicating that mitochondrial dysfunction is not just limited to the SN of PD patients. These data suggest that accumulation of total mtDNA deletions/rearrangements is a relatively specific characteristic of PD and may be one of the contributing factors leading to mitochondrial dysfunction and neurodegeneration in PD. PMID- 12125741 TI - The neuropathological and behavioral consequences of intraspinal microglial/macrophage activation. AB - Activated microglia and macrophages (CNS macrophages) have been implicated in the secondary or "bystander" pathology (e.g. axon injury, demyelination) that accompanies traumatic or autoimmune injury to the brain and spinal cord. These cells also can provide neurotrophic support and promote axonal regeneration. Studying the divergent functional potential of CNS macrophages in trauma models is especially difficult due to the various degradative mechanisms that are initiated prior to or concomitant with microglial/macrophage activation (e.g. hemorrhage, edema, excitotoxicity, lipid peroxidation). To study the potential impact of activated CNS macrophages on the spinal cord parenchyma, we have characterized an in vivo model of non-traumatic spinal cord neuroinflammation. Specifically, focal activation of CNS macrophages was achieved using stereotaxic microinjections of zymosan. Although microinjection does not cause direct mechanical trauma, localized activation of macrophages with zymosan acts as an "inflammatory scalpel" causing tissue injury at and nearby the injection site. The present data reveal that activation of CNS macrophages in vivo can result in permanent axonal injury and demyelination. Moreover, the pathology can be graded and localized to specific white matter tracts to produce quantifiable behavioral deficits. Further development of this model will help to clarify the biological potential of microglia and macrophages and the molecular signals that control their function within the spinal cord. PMID- 12125743 TI - Intracellular deposition, microtubule destabilization, and transport failure: an "early" pathogenic cascade leading to synaptic decline. AB - Protein deposition is a common event in age-related neurological diseases that are characterized by neuronal dysfunction and eventual cell death. Here, cultured hippocampal slices were infused with the lysosomal disrupter chloroquine to examine the link between abnormal protein processing/deposition and early synaptopathogenesis. Tau species of 55 to 69 kDa increased over several days of treatment with chloroquine, while the protein and message levels of synaptic markers were selectively reduced. Neurons of subfields CA1, CA3, and dentate gyrus accumulated protein deposits recognized by antibodies against paired helical filaments and ubiquitin, and this was accompanied by tubulin fragmentation and deacetylation. The deposition filled the basal pole of pyramidal neurons, encompassing the area of the axon hillock and initial dendritic branching but without causing overt neuronal atrophy. Neurons containing the polar aggregates exhibited severely impaired transport along basal dendrites. Transport capability was also lost along apical dendrites, the opposite direction of deposited material in the basal pole; thus, perpetuating the problem beyond physical blockage must be the associated loss of microtubule integrity. These data indicate that transport failure forms a link between tau deposition and synaptic decline, thus shedding light on how protein aggregation events disrupt synaptic and cognitive functions before the ensuing cellular destruction. PMID- 12125744 TI - Planning an adaptive management process for biodiversity conservation and resource development in the Camisea River Basin. AB - The Smithsonian Institution's Monitoring and Assessment of Biodiversity Program joined Shell Prospecting and Development Peru (SPDP) to protect biodiversity during a natural gas exploration project. Emphasis was on long-term societal and environmental benefits in addition to financial gain for the company. The systematic, cyclical adaptive management process was used to generate feedback for SPDP managers. Adaptive management enables ongoing improvement of management policies and practices based on lessons learned from operational activities. Previous to this study, very little information about the local biodiversity was available. Over a 2-year period, the team conducted biological assessments of six taxonomic groups at five sites located within 600 km2. A broad range of management options such as location, timing and technology were developed from the beginning of the project. They were considered in conjunction with emerging lessons from the biodiversity assessments. Critical decisions included location of a gas plant and the cost of helicopter access versus roads to service the full field development. Both of these decisions were evaluated to ensure that they were economically and environmentally feasible. Project design changes, addressed in the planning stage, were accepted once consensus was achieved. Stakeholders were apprised of the implications of the baseline biodiversity assessments. PMID- 12125745 TI - Indices for assessment and monitoring of large mammals within an adaptive management framework. AB - Many development projects intended to exploit natural resources are occurring in fragile ecosystems, and therefore the need for sound biodiversity assessment and monitoring programs is growing. Large mammals are important components of these fragile ecosystems, yet there are few strategies that attempt to assess and monitor entire large mammal communities in relation to development projects. We propose the use of two indices applied within a framework of adaptive management. An occurrence index assesses the composition and distribution of large mammals at a site, and an abundance index monitors the abundance of large mammals over time in relation to development. We discuss the design, applicability and effectiveness of these indices based on our experience with a natural gas development project in the Amazon forests of southeastern Peru. PMID- 12125746 TI - A corporate approach to social monitoring and assessment for development in a fragile environment. AB - The prevailing corporate trend regarding development of energy resources in the tropics emphasizes financial gain over long-term societal benefits. Some corporations are beginning to find a competitive advantage linked to proactive relations with host communities and adequate protection of fragile ecosystems. Herein, we describe a case study where an international energy production company worked with stakeholders to achieve social capital and sustainable development. The strategies aimed to strengthen local capacity to improve social welfare and to ensure conservation and wise use of biodiversity. We provide examples, discuss lessons learned and make recommendations for future development projects. PMID- 12125747 TI - The corporate impact of addressing social issues: a financial case study of a project in Peru. AB - Large, multinational resource development projects can affect many aspects, including social, economic and ecological realities, in the regions where they operate. Social and environmental issues that are usually ignored in such projects are increasingly affecting the financial future of multinational corporations in negative ways. In this article, we advance the argument that corporations can successfully manage these issues and that if they choose to view these management efforts as an investment rather than an expense, they may well acquire a competitive advantage over companies that do not. We describe as a case study the Camisea natural gas and condensates development project in Peru, operated by Shell Prospecting and Development Peru (SPDP). Camisea is one of the first projects anywhere in the world to conduct a detailed analysis of key industry-related social issues and the processes, required investment and financial impact of managing them. The Camisea example supports the argument that addressing social and environmental concerns makes financial sense. In present value terms, the benefit of managing these concerns was expected to surpass the cost investment by approximately US$50 million. PMID- 12125748 TI - Modified Whittaker plots as an assessment and monitoring tool for vegetation in a lowland tropical rainforest. AB - Resource exploitation in lowland tropical forests is increasing and causing loss of biodiversity. Effective evaluation and management of the impacts of development on tropical forests requires appropriate assessment and monitoring tools. We propose the use of 0.1-ha multi-scale, modified Whittaker plots (MWPs) to assess and monitor vegetation in lowland tropical rainforests. We established MWPs at 4 sites to: (1) describe and compare composition and structure of the sites using MWPs, (2) compare these results to those of 1-ha permanent vegetation plots (BDPs), and (3) evaluate the ability of MWPs to detect changes in populations (statistical power). We recorded more than 400 species at each site. Species composition among the sites was distinctive, while mean abundance and basal area was similar. Comparisons between MWPs and BDPs show that they record similar species composition and abundance and that both perform equally well at detecting rare species. However, MWPs tend to record more species, and power analysis studies show that MWPs were more effective at detecting changes in the mean number of species of trees > or = 10 cm in diameter at breast height (dbh) and in herbaceous plants. Ten MWPs were sufficient to detect a change of 11% in the mean number of herb species, and they were able to detect a 14% change in the mean number of species of trees > or =10 cm dbh. The value of MWPs for assessment and monitoring is discussed, along with recommendations for improving the sampling design to increase power. PMID- 12125749 TI - A framework for assessment and monitoring of arthropods in a lowland tropical forest. AB - By applying principles of adaptive management, and by using the valuable information that arthropods provide from assessment and monitoring programs, managers can identify and reduce possible impacts on biodiversity in development projects. In 1996, the Smithsonian Institution's Monitoring and Assessment of Biodiversity program worked together with Shell Prospecting and Development Peru to establish an adaptive management program to protect biodiversity in a natural gas exploration project in a Peruvian rainforest. In this paper, we outlined the conceptual steps involved in establishing an assessment and monitoring program for arthropods, including setting objectives, evaluating the results and making decisions. We also present the results of the assessment using some of groups of arthropods, and summarize the steps taken to identify appropriate groups for monitoring. PMID- 12125750 TI - Framework for assessment and monitoring of amphibians and reptiles in the Lower Urubamba region, Peru. AB - Populations of amphibians and reptiles are experiencing new or increasing threats to their survival. Many of these threats are directly attributable to human activity and resource development. This presents the increasing need for worldwide amphibian and reptile assessments and effective, standardized monitoring protocols. Adaptive management techniques can assist managers in identifying and mitigating threats to amphibian and reptile populations. In 1996, Shell Prospecting and Development, Peru initiated a natural gas exploration project in the rainforest of southeastern Peru. The Smithsonian Institution's Monitoring and Assessment of Biodiversity Program worked closely with Shell engineers and managers to establish an adaptive management program to protect the region's biodiversity. In this manuscript, we discuss the steps we took to establish an adaptive management program for amphibian and reptile communities in the region. We define and outline the conceptual issues involved in establishing an assessment and monitoring program, including setting objectives, evaluating the results and making appropriate decisions. We also provide results from the assessment and discuss the appropriateness and effectiveness of protocols and criteria used for selecting species to monitor. PMID- 12125751 TI - An assessment and monitoring program for birds in the Lower Urubamba region, Peru. AB - We developed an assessment and monitoring plan for birds in connection with the exploration and potential development of a large natural gas field in the Lower Urubamba drainage of Peru, a project of Shell Prospecting and Development Peru (SPDP). Our objectives were to: (1) inventory the birds in the area, including information on habitat use and abundance, and (2) devise long-term monitoring protocols for birds. We sampled birds through a combination of visual and auditory surveys and mist-netting at 4 well sites and 3 sites along the Urubamba and Camisea rivers. We recorded 420 species during 135 days of field work. We consider the highest priorities for a future monitoring program to be: (1) establish whether edge effects are occurring at well sites, along roads and along the planned pipeline route and determine the significance and extent of these effects and (2) assess the impact of increased human access to the area on game and other exploited species. The remoteness of the area, its rugged terrain and dense vegetation and the lack of trained personnel limit the choice of survey and monitoring methods. We recommend use of mist-netting and transects for monitoring edge effects and use of transects for monitoring game and other exploited species. PMID- 12125753 TI - Polyadenylated transcripts containing random gene fragments are expressed in dinoflagellate mitochondria. AB - An AT-rich cDNA isolated from a Gonyaulax library was identified as a putative mitochondrial cytochrome oxidase subunit 3 (cox3) by sequence comparisons. Transcripts hybridizing to this cox3 probe are abundant, are found in poly(A) enriched RNA fractions and have a variable length when tested by Northern blot analysis. A PCR analysis of 21 cox3 clones, designed to measure the length of 5' and 3' ends separately, indicated that the greatest variability in length was found at the 3' end. The variable length of the 3' end may be due to polyadenylation at random sites or to a reduced stringency for polyadenylation signals. Eight cox3 cDNAs were completely sequenced, and were found to differ by up to thirteen bases in the regions of overlap. Curiously, the longest cDNA contained a 350 base pair fragment of a dinoflagellate cox1 gene and a 600 bp fragment of a cytochrome b (cob) gene in the region 3' to the cox3 sequence. Gene fragments or multiple copies of the cox3 and cob sequences may be abundant in the mitochondrial genome since Southern blots, using these sequences separately as probes, show multiple restriction fragments with several enzymes. PMID- 12125752 TI - A framework for assessment and monitoring of small mammals in a lowland tropical forest. AB - Development projects in tropical forests can impact biodiversity. Assessment and monitoring programs based on the principles of adaptive management assist managers to identify and reduce such impacts. The small mammal community is one important component of a forest ecosystem that may be impacted by development projects. In 1996, a natural gas exploration project was initiated in a Peruvian rainforest. The Smithsonian Institution's Monitoring and Assessment of Biodiversity program cooperated with Shell Prospecting and Development Peru to establish an adaptive management program to protect the region's biodiversity. In this article, we discuss the role of assessing and monitoring small mammals in relation to the natural gas project. We outline the conceptual issues involved in establishing an assessment and monitoring program, including setting objectives, evaluating the results and making appropriate decisions. We also summarize the steps taken to implement the small mammal assessment, provide results from the assessment and discuss protocols to identify appropriate species for monitoring. PMID- 12125754 TI - Evolutionary relationships among heterokont algae (the autotrophic stramenopiles) based on combined analyses of small and large subunit ribosomal RNA. AB - In order to study the phylogenetic relationships within the stramenopiles, and particularly among the heterokont algae, we have determined complete or nearly complete large-subunit ribosomal RNA sequences for different species of raphidophytes, phaeophytes, xanthophytes, chrysophytes, synurophytes and pinguiophytes. With the small- and large-subunit ribosomal RNA sequences of representatives for nearly all known groups of heterokont algae, phylogenetic trees were constructed from a concatenated alignment of both ribosomal RNAs, including more than 5,000 positions. By using different tree construction methods, inferred phylogenies showed phaeophytes and xanthophytes as sister taxa, as well as the pelagophytes and dictyochophytes, and the chrysophytes/synurophytes and eustigmatophytes. All these relationships are highly supported by bootstrap analysis. However, apart from these sister group relationships, very few other internodes are well resolved and most groups of heterokont algae seem to have diverged within a relatively short time frame. PMID- 12125755 TI - A simple and rapid PCR-based method to isolate complete small macronuclear minichromosomes from hypotrich ciliates: 5S rDNA and S26 ribosomal protein gene of Oxytricha (Sterkiella) nova. AB - Hypotrich ciliates present a macronuclear genome consisting of gene-sized instead of chromosome-sized DNA molecules. Exploiting this unique eukaryotic genome feature, we introduce, for the first time in ciliates, a rapid and easy PCR method using telomeric primers to isolate small complete macronuclear DNA molecules or minichromosomes. Two presumably abundant macronuclear DNA molecules, containing ribosomal genes, were amplified from the Oxytricha (Sterkiella) nova complete genome after using this method, and then were cloned and sequenced. The 5S rDNA sequence of O. (S.) nova is the third one reported among hypotrich ciliates; its primary and secondary structure is compared with other eukaryotic 5S rRNAs. The ribosomal protein S26 gene is the first one reported among ciliates. This "End-End-PCR" method might be useful to obtain similar gene-sized macronuclear molecules from other hypotrich ciliates, and, therefore, to increase our knowledge on ribosomal genes in these eukaryotic microorganisms. PMID- 12125756 TI - A new class of the stramenopiles, Placididea Classis nova: description of Placidia cafeteriopsis gen. et sp. nov. AB - A marine flagellate resembling Cafeteria roenbergensis (bicosoecids, stramenopiles) in cell shape and behavior of the cell while attached to substratum was collected from the coast of Japan. The flagellate was examined by light and electron microscopy, and the 18S rDNA was sequenced to elucidate its taxonomic and phylogenetic position. Ultrastructural features suggested that the flagellate is not a bicosoecid, but a relative of the recently described stramenopile, Wobblia lunata. 18S rDNA phylogenetic trees also revealed that the flagellate forms a monophyletic clade with W. lunata and that it is distantly related to Cafeteria and other bicosoecids. The flagellate differs from W. lunata due to its lack of wobbling motion as well as intracellular features such as the number of mitochondria, flagellar apparatus architecture, the presence of a paranuclear body and cytoplasmic microtubules. The similarity of 18S rDNA sequences was 81% between the flagellate and W. lunata. This new flagellate was described as Placidia cafeteriopsis gen. et sp. nov. Because the phylogenetic lineage comprised of W. lunata and P. cafeteriopsis was one of the major, deep branching clades of the stramenopiles, the class Placididea (= Placidiophyceae) classis nova was proposed. PMID- 12125757 TI - Vestigial chloroplasts in heterotrophic stramenopiles Pteridomonas danica and Ciliophrys infusionum (Dictyochophyceae). AB - Two heterotrophic members of the Dictyochophyceae (stramenopiles), Pteridomonas danica and Ciliophrys infusionum, were investigated. An undescribed organelle bounded by four membranes and closely associated with the nucleus was detected in P. danica. The outermost membrane was continuous with the outer nuclear membrane. These features strongly suggested that this organelle was a vestigial chloroplast. A photosynthetic gene, rbcL, was successfully amplified by polymerase chain reaction (PCR) from P. danica and C. infusionum. These sequences were readily and well aligned with those of photosynthetic stramenopiles. Phylogenetic trees of 18S rDNA and rbcL were constructed. In all the trees obtained, P. danica and C. infusionum appeared in two different clades, the Pedinellales clade and the Ciliophryales/Rhizochromulinales clade, each of which contained photosynthetic members as well as heterotrophic members. The results indicated that the loss of photosynthetic ability occurred independently in P. danica and C. infusionum. This is the first report of the presence of a vestigial chloroplast (leucoplast) in colorless dictyochophytes. PMID- 12125758 TI - mRNA processing in Dictyostelium: sequence requirements for termination and splicing. AB - Criteria for the identification of termination regions in Dictyostelium discoideum genes have been established and the sequence requirements for termination in 33 genes have been analyzed. A canonical hexamer signal AATAAA was present 15-30 nucleotides upstream of the cleavage site, usually a TA, and was embedded in a particularly A-rich environment. T- or GT-rich downstream elements characteristic of animal cells could not be identified. In a sample of 102 introns we have established the consensus AG/GTAAGT and ATAG/ for the 5' and 3' splice sites, respectively. Most introns are 75-150 nucleotides long and the A+T content is high (90%). A putative branch point was identified in half of the introns 20-60 nucleotides upstream of the 3' splice site and the consensus TACTAAY was derived. A polypyrimidine tract required for branching in vertebrates was not identified, but weak preference for pyrimidine was found 10-45 nucleotides upstream of the 3' splice site. PMID- 12125759 TI - A salute to Antony van Leeuwenhoek of Delft, most versatile 17th century founding father of protistology. PMID- 12125760 TI - Gene expression patterns in Dictyostelium using microarrays. PMID- 12125761 TI - Genetic control of germ-soma differentiation in Volvox carteri. PMID- 12125762 TI - Histological analysis of the maturation of native and wound periderm in potato (Solanum tuberosum L.) Tuber. AB - Maturation of potato (Solanum tuberosum L.) tuber native and wound periderm and development of resistance to periderm abrasion were investigated utilizing cytological and histochemical techniques. Both native and wound periderm consist of three different tissues: phellem, phellogen and phelloderm. It was previously determined that the phellogen walls of immature native periderm are thin and prone to fracture during harvest, leading to periderm abrasion (excoriation). Phellogen walls thicken and become less susceptible to fracture upon maturation of the periderm, leading to resistance to excoriation. We now demonstrate that phellogen cells of immature wound periderm also have thin radial walls and that wound periderm abrasion is due to fracture of these walls. Maturation of the wound periderm is also associated with an increase in the thickness of the phellogen radial walls. Histological analysis with ruthenium red and hydroxylamine-FeCI2, which stain unesterified and highly methyl-esterified pectins, respectively, indicates that the phellogen cell walls of native and wound periderm differ significantly regardless of the stage of maturity. Results obtained by staining with ruthenium red and hydroxylamine-FeCI2 imply that phellogen cell walls of immature native periderm contain methyl-esterified pectin, but are lacking in unesterified (acidic) pectins. Maturation of native periderm is accompanied by an apparent increase in unesterified pectins in the walls of phellogen cells, which may allow for the strengthening of phellogen cell walls via calcium pectate formation. Histological staining of the phellogen walls of wound periderm, on the other hand, implies that these walls are deficient in pectins. Moreover, maturation of wound periderm is not accompanied by an increase in unesterified pectins in these walls. Since peroxidase is known to catalyse the cross-linking of cell wall polymers, we stained native and wound periderm for the presence of peroxidase utilizing guaiacol as a substrate. Peroxidase staining was strong in the phellogen walls of both immature and mature native periderm and we could not detect any differences in staining between them. Peroxidase staining was weak in the phellogen walls of immature wound periderm and was not detectably different in mature wound periderm. Peroxidase data imply that there are distinct differences between native and wound periderm, though our data do not indicate that changes in peroxidase activity are involved in the development of resistance to periderm abrasion that occurs upon maturation of the periderm. However, we cannot rule out the involvement in this process of peroxidase isozymes that have low affinity for the substrates utilized here. PMID- 12125763 TI - Photoautotrophic culture of Coffea arabusta somatic embryos: photosynthetic ability and growth of different stage embryos. AB - Coffea arabusta somatic embryos were cultured and development of stomata, rate of CO2 fixation or production, chlorophyll content and chlorophyll fluorescence were studied in embryos at different stages of development. Cotyledonary and germinated embryos have photosynthetic capacity, although pretreatment at a high photosynthetic photon flux (PPF) (100 micromol m(-2) s(-1)) for 14 d increased photosynthetic ability. Except in a very small number of cases, stomata did not develop fully in precotyledonary stage embryos and were absent in torpedo stage embryos. Low chlorophyll content (90-130 microg g(-1) fresh mass) was noted in torpedo and precotyledonary stage embryos compared with cotyledonary and germinated embryos (300-500 microg g(-1) fresh mass). Due to the absence of stomata and low chlorophyll content in the torpedo and precotyledonary stage embryos, the photosynthetic rate was low and, in some cases, CO2 production was observed. These data suggest that the cotyledonary stage is the earliest stage that can be cultured photoautotrophically to ensure plantlet development. When grown photoautotrophically (in a sugar-free medium with CO2 enrichment in the culture headspace and high photosynthetic photon flux), torpedo and precotyledonary stage embryos lost 20-25% of their initial dry mass after 60 d of culture. However, in cotyledonary and germinated embryos, the dry mass of each embryo increased by 10 and 50%, respectively. By using a porous supporting material, growth (especially root growth) was increased in cotyledonary stage embryos. In addition, photoautotrophic conditions, high PPF (100-150 micromol m( 2) s(-1)) and increased CO2 concentration (1100 micromol mol(-1)) were found to be necessary for the development of plantlets from cotyledonary stage embryos. PMID- 12125764 TI - Effect of day and night temperature on internode and stem length in chrysanthemum: is everything explained by DIF? AB - In many plant species, including chrysanthemum, a strong positive correlation between internode length and DIF [difference between day (DT) and night (NT) temperature] has been observed. However, Langton and Cockshull (1997. Scientia Horticulturae 69: 229-237) reported no such relationship and showed that absolute DT and NT explained internode length rather than DIF. To investigate these conflicting results and to clarify the validity of the DIF concept, cut chrysanthemums (Chrysanthemum 'Reagan Improved') were grown in growth chambers at all 16 combinations of four DT and four NT (16, 20, 24 and 28 degrees C) with a 12 h day length. Length of internode 10, number of internodes and stem length were measured on days 5, 10, 17, 22 and 27 after starting the temperature treatments. Internode length on day 10 showed a positive linear relationship with DIF (R2 = 0.64). However, when internodes had reached their final length in all treatments (day 27), a much stronger positive linear relation was observed (R2 = 0.81). A model to predict final internode length was developed based on the absolute DT and NT responses: both responses were optimum curves and no significant interaction between DT and NT occurred [final internode length (mm) = -32.23 + 3.56DT + 1.08NT - 0.0687DT2 - 0.0371NT2; R2 = 0.91, where TD is day temperature and TN is night temperature]. It is shown that DIF can predict final internode length only within a temperature range where effects of DT and NT are equal in magnitude and opposite in sign (18-24 degrees C). Internode appearance rate, as well as stem length formed during the experiment, showed an optimum response to DT. PMID- 12125765 TI - Allocation to floral structures in Thalictrum pubescens (Ranunculaceae), a cryptically dioecious species. AB - Females of Thalictrum pubescens produce stamens that contain sterile pollen, whereas males are both functionally and morphologically unisexual. This study examines the investment in stamen production by females of T. pubescens by comparing the female structures with those of their fully functional male counterparts. Stamens from females had the same biomass and contained the same amount of nitrogen and phosphorus as stamens from males. Anther size was the same in males and females, but filaments were longer in stamens from males. Females produced more pollen per anther than males, and pollen size was the same in both sexes. Within flowers, there was a positive correlation between the amount of pollen per anther and the length of anthers in males, but not in females. This would be expected if males growing in better environmental conditions or with greater vigour invested more resources in pollen production, thereby increasing fitness. Females, who receive no fitness benefits from increased pollen production, did not show this pattern. There was also evidence of a trade-off within female flowers between the number of stamens and the number of pistils. This trade-off was noted in conditions when variance among plants was reduced, namely in the field during a year when flower size was particularly small and in a previous glasshouse study. Therefore, it appears that when environmental variance is low, stamens are produced at the expense of producing more pistils, and hence seeds. In conclusion, stamen production does not appear to be inconsequential to females of Thalictrum pubescens. PMID- 12125766 TI - Effects of elevated ultraviolet-B radiation on native and cultivated plants of southern Africa. AB - Seventeen herb, shrub and tree species of commercial and ecological importance in southern Africa were exposed at one location to ultraviolet-B (UV-B, 280-315 nm) radiation approx. 35 % above clear-sky background (control). The aims were to assess how UV-B affects canopy area, dry mass, and some biochemical and morphological properties of leaves, and to investigate whether differences between species are related to growth form of the plants. There was no pattern of response to UV-B related to growth form. Leaves of trees had altered chlorophyll a and b, carotenoid and flavonoid concentrations, but those of shrubs or herbs did not. Non-structural carbohydrates were unaffected. Smaller canopy areas and dry masses were observed under enhanced UV-B, but these were not statistically different among growth forms. There was a general insensitivity of species to elevated UV-B. Only five species had significantly altered leaf biochemical and morphological properties, canopy area and dry mass, the changes differing in magnitude. There was no consistent pattern of change in leaf thickness or biochemical composition with increased UV-B. Correlation analyses did not support the view that growth is less negatively affected in species with thick leaves or in those where leaf thickness increases, or in species with naturally high leaf flavonoid contents or that are able to synthesize additional flavonoids in response to UV-B enhancement. The analyses did not support the hypothesis that growth was inhibited by starch accumulation in leaves under elevated UV-B. However, changes in leaf shape did correlate with canopy area and dry mass, showing the importance of photomorphogenetic changes caused by UV-B which affect species' performance. We conclude that generalizations on plant sensitivity to UV B based on growth form and functional type could be misleading, and that the great majority of economically important species of the region are likely to be insensitive to future UV-B increases. Notable exceptions include the Colophospermum mopane tree ecotypes chota and leslie and the arable annual Vigna unguiculata, both of which are traditional sources of livelihood to rural African populations and of importance to African industry and agriculture. PMID- 12125767 TI - A simple model of feedback regulation for nitrate uptake and N2 fixation in contrasting phenotypes of white clover. AB - A simple three equation model is proposed for the feedback regulation of nitrate uptake and N2 fixation, based on the concentration of the organic N substrate pool within the plant and two parameters denoting the N substrate concentrations at which half-maximal inhibition occurs. This model simulated three contrasting phenotypes of white clover (Trifolium repens L.) inbred lines with (1) normal rates of nitrate uptake and N2 fixation (NNU); (2) low rates of nitrate uptake (LNU); and (3) very low rates of N2 fixation (VLF). The LNU phenotype was simulated by a decrease in the value of the inhibition parameter for nitrate uptake and the VLF phenotype was simulated by a decrease in the value of the N2 fixation inhibition parameter. The model was tested against nitrate uptake data obtained from white clover plants growing in flowing nutrient culture. There was an accurate prediction of the increase in nitrate uptake caused by N2 fixation activity of the NNU and LNU inbred lines being interrupted by a switch in gas phase from air to Ar : O2. The model was also tested against data for nitrate uptake, N2 fixation and %N from fixation for the three inbred clover lines grown in flowing nutrient culture at 0, 5 or 20 mmol m(-3) N(3-). Again there was accurate prediction of nitrate uptake, although simulated values for N2 fixation were more variable. The simple model has potential use as a sub-routine in larger models of legume growth under field conditions. PMID- 12125768 TI - Silica accumulation in long-lived leaves of Sasa veitchii (Carriere) Rehder (Poaceae-Bambusoideae). AB - Silica accumulation in long-lived leaves of Sasa veitchii was investigated using the molybdenum blue method. In general, silica accumulation was rapid during spring and summer, and slow during winter. The leaves continuously accumulate silica throughout their life. The significance of these observations is discussed in relation to two hypotheses for silica accumulation in plants: (1) that silica accumulation is a result of water consumption by the plant: and (2) that silica is actively accumulated to protect the plant. The results of the present study support the former hypothesis. PMID- 12125769 TI - Photoautotrophic culture of Coffea arabusta somatic embryos: development of a bioreactor for large-scale plantlet conversion from cotyledonary embryos. AB - Somatic embryos were developed from in vitro-grown leaf discs of Coffea arabusta in modified Murashige and Skoog medium under 30 micromol m(-2) s(-1) photosynthetic photon flux (PPF). Cotyledonary stage embryos were selected from the 14-week-old cultures and were placed under a high (100 micromol m(-2) s(-1) PPF for 14 d. These pretreated embryos were grown photoautotrophically in three different types of culture systems: Magenta vessel; RITA-bioreactor (modified to improve air exchange); and a specially designed temporary root zone immersion bioreactor system (TRI-bioreactor) with forced ventilation. The aims of the study were to achieve large-scale embryo-to-plantlet conversion, and to optimize growth of plantlets under photoautotrophic conditions. The plantlet conversion percentage was highest (84 %) in the TRI-bioreactor and lowest in the modified RITA-bioreactor (20 %). Growth and survival of converted plantlets following 45 d of photoautotrophic culture in each of the three culture systems were studied. Fresh and dry masses of leaves and roots of plantlets developed in the TRI bioreactor were significantly greater than those of plantlets developed in the modified RITA-bioreactor or Magenta vessel. The net photosynthetic rate, chlorophyll fluorescence and chlorophyll contents were also highest in plantlets grown in the TRI-bioreactor. Normal stomata were observed in leaves of plantlets grown in the TRI-bioreactor, whereas they could be abnormal in plantlets from the modified RITA-bioreactor. Survival of the plants after transfer from culture followed a similar pattern and was highest in the group grown in the TRI bioreactor, followed by plants grown in the modified RITA-bioreactor and Magenta vessel. In addition, ex vitro growth of plants transferred from the TRI bioreactor was faster than that of plants from the other culture systems. PMID- 12125771 TI - Avoiding bias in calculations of relative growth rate. AB - In classical growth analysis, relative growth rate (RGR) is calculated as RGR = (ln W2 - ln W1)/(t2 - t1), where W1 and W2 are plant dry weights at times t1 and t2. Since RGR is usually calculated using destructive harvests of several individuals, an obvious approach is to substitute W1 and W2 with sample means W1 and W2. Here we demonstrate that this approach yields a biased estimate of RGR whenever the variance of the natural logarithm-transformed plant weight changes through time. This bias increases with an increase in the variance in RGR, in the length of the interval between harvests, or in sample size. The bias can be avoided by using the formula RGR = (ln W2 - ln W1)/(t2 - t1),where ln W1 and ln W2 are the means of the natural logarithm-transformed plant weights. PMID- 12125770 TI - Fluorescence in situ hybridization analysis of alien genes in Agrobacterium mediated Cry1A(b)-transformed rice. AB - The transgene in Agrobacterium-mediated Cry1A(b)-transgenic rice plants has been detected and its chromosomal location determined by fluorescence in situ hybridization (FISH). Eight of the nine transgenic lines tested showed hybridization signals. Signals were located on regions of the chromosome in which fraction length (FL) values varied from 26.2 (near the centromere) to 95.2 (distal regions). No signal was found on regions where the fraction length was less than 26.2, while six of the nine signals detected were located on regions with FL values of 75.3 or over. This demonstrates that Agrobacterium-mediated genes can integrate into multiple sites distributed in different parts of the chromosome, but that distal regions are the preferred sites and regions near the centromeres are colder for T-DNA integration. The donor DNA of the transformation was divided into two parts, labelled separately as probes for two-colour FISH. Results show that the transformed DNA sequences remained linked in the recipient genome. The relationship between integration position and transgene silencing, known as the 'position effect', is discussed. PMID- 12125772 TI - Number, position, diameter and initial direction of growth of primary roots in Musa. AB - To understand soil colonization by a root system, information is needed on the architecture of the root system. In monocotyledons, soil exploration is mainly due to the growth of adventitious primary roots. Primary root emergence in banana was quantified in relation to shoot and corm development. Root emergence kinetics were closely related to the development of aerial organs. Root position at emergence on the corm followed an asymptotic function of corm dry weight, so that the age of each root at a given time could be deduced from its position. Root diameter at emergence was related to the position of the roots on the corm, with younger roots being thicker than older ones. However, root diameters were not constant along a given root, but instead decreased with the distance to the base; roots appear to be conical in their basal and apical parts. Root growth directions at emergence were variable, but a high proportion of the primary roots emerged with a low angle to the horizontal. Further research is needed to evaluate whether these initial trajectories are conserved during root development. Results presented in this study are in good agreement with those reported for other monocotyledons such as maize and rice. They give quantitative information that will facilitate the development of models of root system architecture in banana. PMID- 12125773 TI - Aluminium accumulation in leaves of 127 species in Melastomataceae, with comments on the order Myrtales. AB - The distribution and systematic significance of aluminium accumulation is surveyed based on semi-quantitative tests of 166 species, representing all tribes and subfamilies of the Melastomataceae as well as a few members of related families within the Myrtales. The character is strongly present in nearly all members of the Memecylaceae and in most primitive taxa of the Melastomataceae, while non-accumulating taxa are widespread in the more derived tribes of the Melastomataceae. The variable distribution of Al accumulation in advanced clades of the latter family is probably associated with the tendency to herbaceousness, although it is unclear whether the more herbaceous representatives have developed more specialized Al-response mechanisms that may exclude high Al levels from the shoot. It is hypothesized that Al accumulation is symplesiomorphic for Melastomataceae and Memecylaceae, and that the feature characterizes the most primitive families in the Myrtales. Indeed, Al accumulation is also characteristic of Crypteroniaceae, Rhynchocalycaceae and Vochysiaceae. Crypteroniaceae and Rhynchocalycaceae probably take a basal position in a sister clade of the Memecylaceae and Melastomataceae, while Al accumulation suggests a basal position for Vochysiaceae in the Myrtaceae clade. PMID- 12125774 TI - A generic model to describe the dynamics of nutrient concentrations within stemwood across an age series of a eucalyptus hybrid. AB - Nutrient concentrations (N, P, K) were determined within stemwood in an age series of eucalyptus stands. Four trees per stand were selected according to their size to represent the whole range of basal areas in 1-, 2-, 3-, 4-, 5-, 6- and 7-year-old stands. Cross-sections were sampled every 4 m from the ground to the top of the tree, and chemical analyses were performed for each annual ring in the cross-sections. We constructed a new and generic model to describe the dynamics of nutrient concentrations within the stemwood. Three main parameters were used: (1) the initial concentration of the ring, Ic; (2) the final concentration of the ring at harvest, Fc; and (3) the rate of change in concentration, k. The model is very flexible and was adapted to describe N, P and K concentrations within the stems, and their dynamics over time. An analysis of the parameters showed that k was constant for a given nutrient. Ic varied with height within the tree for P, whereas for N and K it was a function of: (1) the age of the tree when the ring was initiated: and (2) height within the tree. Fc was constant for N, and dependent on the age of the tree when the ring was initiated for K and P. The final models showed a low Root Mean Square Error for a limited number of parameters (less than seven). When validated on an independent sample, the models were shown to have high predictive quality. PMID- 12125775 TI - Comparison of somatic embryogenesis-derived coffee (Coffea arabica L.) plantlets regenerated in vitro or ex vitro: morphological, mineral and water characteristics. AB - Coffea arabica L. plantlets obtained ex vitro after sowing somatic embryos produced in a bioreactor in horticultural substrate were compared with those obtained in vitro from the same embryo population under conventional culturing conditions on semi-solid media. The intensity and quality of aerial and root system development were compared. Shoot emergence was more efficient in vitro but rooting frequencies were low. In contrast, all ex vitro-regenerated embryos rooted. The cotyledon area of mature embryos produced in a bioreactor positively affected plantlet development when regeneration was carried out ex vitro. Embryos with an intermediate cotyledon area (0.86 cm2) had the highest rates of plant conversion ex vitro (63%), and also resulted in vigorous plantlets. Mortality was higher in nursery conditions, but better plant development was obtained. The quality of plantlets produced under ex vitro conditions was reflected in better growth of the aerial and root systems, and also by similar morphological, mineral and water status characteristics to seedlings. Unlike roots formed on semi-solid media, those produced in soil were branched, fine (30-50% had a diameter of less than 0-5 mm) and they bore root hairs. Leaves of plantlets regenerated ex vitro had a histological structure similar to that of seedling leaves, and a lower stomatal density (100 vs. 233 mm-2). Moreover, they were more turgid, as indicated by higher pressure potential (psiP) (0.91 s. 0.30 MPa) and relative water content values (97 vs. 93%). Furthermore, under in vitro conditions, leaves had larger stomata which were abnormally round and raised. Direct sowing of germinated somatic embryos resulted in the rapid production of vigorous plantlets under ex vitro conditions, whilst removing the need for problematical and costly conventional acclimatization procedures. PMID- 12125776 TI - Tillering in grain sorghum over a wide range of population densities: identification of a common hierarchy for tiller emergence, leaf area development and fertility. AB - Most studies of tiller development have not related the physiological and morphological features of each culm to its subsequent fertility. This introduces problems when trying to account for the effects of tillering on yield in crop models. The objective of this study was to detect the most likely early determinants of tiller fertility in sorghum by identifying hierarchies for emergence, fertility and grain number of tillers over a wide range of assimilate availabilities. Emergence, phenology, leaf area development and dry weight partitioning were quantified weekly for individual tillers and main culms of tillering and uniculm plants grown at one of four densities, from two to 16 plants m(-2). For a given plant in any given density, the same tiller hierarchy applied for emergence of tillers, fertility of the emerged tillers and their subsequent grain number. These results were observed over a range of tiller fertility rates (from 7 to 91%), fertile tiller number per plant at maturity (from 0.2 to 4.7), and tiller contribution to grain yield (from 5 to 78 %). Tiller emergence was most probably related to assimilate supply and light quality. Development, fertility and contribution to yield of a specific tiller were highly dependent on growing conditions at the time of tiller emergence, particularly via early leaf area development of the tiller, which affected its subsequent leaf area accumulation. Assimilate availability in the main culm at the time of tiller emergence was the most likely early determinant of subsequent tiller fertility in this study. PMID- 12125777 TI - Tillering in grain sorghum over a wiide range of population densities: modelling dynamics of tiller fertility. AB - The prediction of tillering is poor or absent in existing sorghum crop models even though fertile tillers contribute significantly to grain yield. The objective of this study was to identify general quantitative relationships underpinning tiller dynamics of sorghum for a broad range of assimilate availabilities. Emergence, phenology, leaf area development and fertility of individual main culms and tillers were quantified weekly in plants grown at one of four plant densities ranging from two to 16 plants m(-2). On any given day, a tiller was considered potentially fertile (a posteriori) if its number of leaves continued to increase thereafter. The dynamics of potentially fertile tiller number per plant varied greatly with plant density, but could generally be described by three determinants, stable across plant densities: tiller emergence rate aligned with leaf ligule appearance rate; cessation of tiller emergence occurred at a stable leaf area index; and rate of decrease in potentially fertile tillers was linearly related to the ratio of realized to potential leaf area growth. Realized leaf area growth is the measured increase in leaf area, whereas potential leaf area growth is the estimated increase in leaf area if all potentially fertile tillers were to continue to develop. Procedures to predict this ratio, by estimating realized leaf area per plant from intercepted radiation and potential leaf area per plant from the number and type of developing axes, are presented. While it is suitable for modelling tiller dynamics in grain sorghum, this general framework needs to be validated by testing it in different environments and for other cultivars. PMID- 12125778 TI - Concepts of social justice in community psychology: toward a social ecological epistemology. AB - In this paper we address the pervasive tendency in community psychology to treat values like social justice only as general objectives rather than contested theoretical concepts possessing identifiable empirical content. First we discuss how distinctive concepts of social justice have figured in three major intellectual traditions within community psychology: (1) the prevention and health promotion tradition, (2) the empowerment tradition, and most recently, (3) the critical tradition. We point out the epistemological gains and limitations of these respective concepts and argue for greater sensitivity to the context dependency of normative concepts like social justice. More specifically, we point to a pressing need in community psychology for an epistemology that: (1) subsumes both descriptive and evaluative concepts, and (2) acknowledges its own embeddedness in history and culture without thereby reducing all knowledge claims to the status of ideology. Finally, we describe and demonstrate the promise of what we are calling a social ecological epistemology for fulfilling this need. PMID- 12125779 TI - The evolution of epistemology and concepts in an iterative-generative reflective practice: the importance of small differences. AB - Using a contextualist epistemology, it would be expected that regional differences in community psychology would develop over time. It is argued that the epistemology and theory of Western Australian community psychology, while largely based on North American approaches, has developed its own idiosyncracies. These developed through the integration of practice and theory in an "iterative generative" fashion. The process of development is conceptualized in terms of Schon's and Altman's distinctions between foundational knowledge, and professional and socially responsive knowledge (I. Altman, 1996; D. A. Schon, 1983). It has also been characterized as an incremental development based on the reflection on tacit and conceptual knowledge. From the small differences that have developed between regions, a dialogue can emerge that will better allow understanding of how social forces shape people's actions. PMID- 12125780 TI - Ideology and community social psychology: theoretical considerations and practical implications. AB - This paper addresses the importance of the concept of ideology in community work. The implications of a Marxist approach to ideology in community practice are analyzed in terms of the concepts of problematization (P. Freire, 1979) and consciousness-raising (J. Barreiro, 1976), illustrating the point with some examples. The traditional Marxist perspective is also examined in relation to the perspectives of social constructionism (I. Ibanez, 1996), cultural studies (A. McRobbie, 1992), post-Marxism (E. Laclau & C. Mouffe, 1985), and feminism (D. Haraway, 1991). It is argued that the concepts of hegemony and habitus (P. Bourdieu, 1985) can be useful to community social psychology theory and practice. A "situated perspective"--in which it is possible to dialogue from different "subject positions," and articulate transformation and political action--is argued. The implications of this shifting in the concept of ideology by means of theoretical developments outside social communitypsychology can help to define the external (outside) agent's position in community practice. PMID- 12125781 TI - Community, subjectivity, and intersubjectivity. AB - This paper deals with the notions of "subjectivity," "intersubjectivity," and "community" from several different points of view that include subjective and intersubjective agency, a sense of community, the community as a social institution, and the idea of social justice. The context of these considerations can be found in the Community-Social-Psychological approach to social action as it is often practiced in Latin America. A review of these themes is considered important because different models of community intervention and practice may lead to different expressions of community interaction. PMID- 12125782 TI - The institutionalization of community interventions in Chile: characteristics and contradictions. AB - A critical analysis of the consequences of successive institutionalization of community intervention in Chile is presented. This analysis is based on research of current community interventions in Chile, whose results are compared with Chilean community practice from previous historical periods. Chilean community intervention was formerly practiced out of governmental institutions and universities and was sustained primarily by foreign agencies and ecclesiastical institutions. Nowadays, with the return to democracy, the vast majority of community intervention programs arefinanced, partially or totally, by means of governmental resources. This institutionalization of community interventions has had consequences worthy of critical analysis: an increase in the number and stability of intervention programs; and also negative consequences for intervention goals definition of target groups, and intervention practices. Dilemmas, such as assistance vs. promotion or adaptation vs. social change, have arisen. These dilemmas show the contradictions of a Community Psychology, which has left behind the position of institutional marginality to form part of Chilean society's "normal" psychosocial care. They also show the need for a paradigm shift from a "critical" view of the social world to a constructionist one. PMID- 12125783 TI - On the construction of reality and truth. Towards an epistemology of community social psychology. AB - An expression of community socialpsychology based on the need to transform social reality, and to considerpeople as the constructors ofthat reality, is examined from an epistemologicalpoint of view. Dualism, the position considering that object and subject are separate entities, and monism, the perspective stating that there is only one substance, are discussed The consequences of both conceptions for community social psychology, and their incompatibility, as well as the notions of reality and truth are analyzed. That analysis deals with the problems of defining reality, of separating subject and object of knowledge, of language's role, and of relativism and truth. Finally, a constructionist view of monism based on relatedness, and action, is proposed, stating the mutually influencing union of subject and object in the construction of reality. PMID- 12125784 TI - One size fits all. PMID- 12125785 TI - A respected profession. PMID- 12125786 TI - Professional responsibility. PMID- 12125787 TI - Carious primary teeth. PMID- 12125788 TI - Carious primary teeth. PMID- 12125789 TI - Carious primary teeth. PMID- 12125790 TI - Carious primary teeth. PMID- 12125791 TI - Carious primary teeth. PMID- 12125792 TI - Carious primary teeth. PMID- 12125793 TI - Genes 'n' greens: the future of oral medicine? PMID- 12125794 TI - Crowns and other extra-coronal restorations: impression materials and technique. AB - Well-fitting indirect restorations can only be made if there are accurate models of the oral tissues available, made from high quality impressions. Waiting for an impression to set may be more stressful for the dentist than the patient. Should the impression need to be repeated there is the embarrassment of having to explain this to the patient, the cost implications of material and time wasted and the aggravation of running late for the next appointment. Yet, if a 'Nelsonian' eye is turned to a defective impression we can only expect a substandard restoration in return. PMID- 12125796 TI - The relationship between oral health status and Body Mass Index among older people: a national survey of older people in Great Britain. AB - AIMS: To assess the relationship between oral health status and Body Mass Index. MATERIAL AND METHODS: This paper relates to the free-living sample (participants who lived in their own home, rather than an institution) of the National Diet and Nutrition Survey: people aged 65 years and older. SUBJECTS: 629 adults. DATA COLLECTION: A probability random national sample of adults who had a dental examination, an interview and an anthropometric examination. DATA ANALYSIS: Fisher's exact test and multivariate logistic modeling. FINDINGS: Being underweight was relatively uncommon in this population. People without teeth were significantly (P=0.05) more likely to be underweight than those with 11 or more teeth; 12.3% and 2.9%. A highly statistically significant (P=0.001) difference was observed in BMI between dentate people with 1-10 teeth and with more than 10 teeth; 24% and 2.9% were underweight. Dentate people with less than 21 natural teeth were on average more than 3 times more likely to be obese than those with 21-32 teeth (P=0.036). There was no significant difference in both the proportion of overweight and obese adults between those who were edentulous and dentate with 21 or more teeth. A similar pattern was observed when the number of posterior occluding pairs was compared with BMI categories. Results of multiple logistic regression were adjusted for the confounding effects of age, social class, region of origin and partial denture wearing. CONCLUSIONS: Older people in Britain with more than 20 teeth are more likely to have a normal Body Mass Index. PMID- 12125795 TI - Influence of the method of funding on the age of failed restorations in general dental practice in the UK. AB - OBJECTIVE: This study examined the effect of the method of funding treatment on the age of restorations at the time of replacement. METHOD: A group of general dental practitioners were recruited to take part in the study. Each participant was asked to record the reason for placement and replacement of restorations. The age and class of the restoration being replaced was also recorded, together with details of the material being used and the material being replaced. Details of the method of payment of the failed restoration were recorded. RESULTS: Details of the reason for placement/replacement were received for 3,196 restorations from 32 GDPs. Of the restorations placed, 54% were amalgam, 32% composite, 8% compomer and 7% glass ionomer. The age of restorations at the time of replacement was significantly associated with the method of payment for the restoration, with restorations placed in the Armed Forces having been in service significantly longer at the time of their replacement than restorations placed under NHS regulations. CONCLUSION: Statistical analysis indicated that restorations placed within the NHS regulations were replaced at a significantly lower age than restorations placed under the other funding arrangements investigated. PMID- 12125797 TI - Toothache tales: Part 2. AB - The second part of the selection of literary excerpts on toothache deals with extraction as the ultimate curse. In common with killing a cat, there are many ways to do it. PMID- 12125798 TI - 2002 British and Irish Dental Associations' Annual Conference. AB - This year's annual conference held at Belfast's Waterfront Hall proved a resounding success. The three-day conference focused on 'Quality Partnership'- the importance of the dentist/team partnership, keeping-up-to-date and ensuring patients are well informed. PMID- 12125800 TI - Clinical performance review for diabetic care comparing medical record versus claims and administrative data. PMID- 12125799 TI - Focus Awards 2001. AB - In the last of a series of five articles in which we look at each of the practice finalists from the Focus Awards 2001, we visit 68 The Dental Practice in Leeds, West Yorkshire. PMID- 12125801 TI - The economic benefit for family/general medicine practices employing physician assistants. AB - OBJECTIVE: To measure the economic benefit of a family/general medicine physician assistant (PA) practice. STUDY DESIGN: Qualitative description of a model PA practice in a family/general medicine practice office setting, and comparison of the financial productivity of a PA practice with that of a non-PA (physician only) practice. METHODS: The study site was a family/general medicine practice office in southwestern Pennsylvania. The description of PA practice was obtained through direct observation and semistructured interviews during site visits in 1998. Comparison of site practice characteristics with published national statistics was performed to confirm the site's usefulness as a model practice. Data used for PA productivity analyses were obtained from site visits, interviews, office billing records, office appointment logs, and national organizations. RESULTS: The PA in the model practice had a same-task substitution ratio of 0.86 compared with the supervising physician. The PA was economically beneficial for the practice, with a compensation-to-production ratio of 0.36. Compared with a practice employing a full-time physician, the annual financial differential of a practice employing a full-time PA was $52,592. Sensitivity analyses illustrated the economic benefit of a PA practice in a variety of theoretical family/general medicine practice office settings. CONCLUSIONS: Family/general medicine PAs are of significant economic benefit to practices that employ them. PMID- 12125802 TI - Patient-physician communication as organizational innovation in the managed care setting. AB - Despite changes in the healthcare system, the relationship between patients and physicians remains fundamental to high-quality care. Managed care rules and restrictions, such as constraints on choice of providers, review processes, and decreasing length of visits, are creating potential conflicts between patients and their physicians. To strengthen the patient-physician relationship, some managed care organizations are implementing communication skills training for physicians. This article provides case studies describing how 2 large managed care organizations successfully incorporated communication skills training into their environments. An organizational perspective is used to delineate the 3 stages--adoption, implementation, and institutionalization--that managed care organizations generally traverse in incorporating communication skills programs and making them an integral part of their organizational culture. Specific suggestions are provided for physician leaders and administrators who are considering similar programs in their settings. PMID- 12125803 TI - Disease management in healthcare organizations: results of in-depth interviews with disease management decision makers. AB - BACKGROUND: Despite the widening use of disease management (DM) programs throughout the country, little is understood about the "state of DM" in healthcare systems and managed care organizations. OBJECTIVE: To better characterize the range of users of DM in healthcare and to identify critical issues, both present and future, for DM. STUDY DESIGN: Qualitative survey. PARTICIPANTS AND METHODS: Forty-seven healthcare systems (n = 22) and managed care organizations (n = 25) were randomly selected. Decision makers were identified and interviewed between January 1, 2000, and March 31, 2000. We limited quantitative analysis to tabulations of suitable responses, without statistical testing. Responses were organized around 3 themes: models for DM, implementation strategies, and measurements of success. RESULTS: Of 47 decision makers surveyed, 42 (89%) reported that their organizations currently have (75%) or are working to develop (14%) DM programs. Although the goals of DM programs were similar, organizations took a variety of approaches to achieving these ends. There were typically 3 steps in implementing a DM program: analysis of patient data, external analysis, and organizational analysis. Decision makers believed that DM programs had only achieved partial success in reaching the 2 main goals of improved quality of care and cost savings. CONCLUSIONS: Given the variety of DM programs, there is a need to develop a classification scheme to allow for better comparison between programs. Further quantitative studies of decision makers' opinions would be helpful in developing programs and in designing necessary studies of patient management strategies. PMID- 12125804 TI - Frequency of negative coronary arteriographic findings in patients with chest pain is related to community practice patterns. AB - OBJECTIVE: To determine factors contributing to the relatively high frequency and variability (10% to 30%) of finding no significant coronary disease by coronary angiography in patients with chest pain. STUDY DESIGN: Retrospective, comparative analysis of practice patterns at 3 southeastern Michigan hospitals and a composite sample from New York State. PATIENTS AND METHODS: Medical records for 7668 patients were reviewed to determine the frequency of negative coronary arteriographic findings in patients undergoing chest pain evaluation. A private practice allopathic community hospital with interventional cardiologists and a private practice osteopathic community hospital with diagnostic facilities (DiagCommunity) were compared with a university hospital with full-time salaried interventional cardiologists and a sample of 17 New York hospitals. RESULTS: Of the 7668 coronary angiograms at all centers, 39.7% were performed to assess patients with stable chest pain. There was no significant obstruction found in 16.5%, and the frequency was not different between the Michigan (17.8%+/-3.8%) and New York (14.2%) hospitals. The DiagCommunity had the highest proportion (22%; P < .001 vs others). On review of the negative coronary arteriographic findings, normal or near normal coronary arteriographic findings were infrequent (range, 2.4%-6.6%) but higher in the DiagCommunity (6.6% vs 2.9%+/-1.6%; P < .0001). CONCLUSIONS: The frequency of finding no significant coronary disease by arteriography in patients with chest pain is similar in southeastern Michigan hospitals and comparable to an established external database. Cardiology self referral and personal gain does not seem to be a major factor in selection of patients for invasive studies. PMID- 12125805 TI - Managed care for uninsured adults: the rise and fall of a university-based program. AB - OBJECTIVE: To assess the impact of CU CARE, a managed care program for medically indigent adults developed by University Hospital (UH) in Denver and Kaiser Permanente, on outpatient and inpatient utilization. STUDY DESIGN: Pre-post study with concurrent comparison groups. PATIENTS AND METHODS: Administrative claims from 1994-1996 were analyzed for all enrollees in a state-funded medically indigent program (intervention group) compared with Medicaid patients and uninsured adults rated as "self-pay" who were ineligible for the medically indigent program. RESULTS: In 1994, before initiation of CU CARE, UH provided care to 10,118 medically indigent, 5330 Medicaid, and 7626 self-pay patients; similar numbers received care in 1995-1996, but only 12% of medically indigent patients received care in both time periods. The proportion of medically indigent patients with 1 or more primary care visits increased by 185% (from 10.9% in 1994 to 31.1% in 1995-1996). Medically indigent patients had relative declines of 36% in specialty clinic visits, 25% in emergency department visits, 40% in hospital visits, and 31% in visit costs between 1994 and 1995-1996. All these changes were significant compared with Medicaid and self-pay patients. The impact on acute care utilization was greater for medically indigent patients who used UH in both 1994 and 1995-1996. CONCLUSIONS: This managed care program increased utilization of primary care and reduced specialty and acute care utilization. However, the program was scaled back in 1997 and terminated in 2000 because of problems with care coordination across institutions, increasing costs (particularly pharmacy costs), and competitive pressures. PMID- 12125806 TI - Hormone replacement therapy: current concerns and considerations. AB - AUDIENCE: This activity is designed for pharmacists and other healthcare professionals who evaluate and treat perimenopausal and postmenopausal women. GOALS: To understand the benefits, risks, and adverse effects associated with estrogen replacement therapy (ERT) and hormone replacement therapy (HRT) and their influence on a postmenopausal woman's initiation, adherence, and satisfaction with therapy. OBJECTIVES: 1. Discuss menopause and its effects. 2. Identify ERT/HRT's potential benefits and risks. 3. Discuss ERT/HRT's adverse effects and management approaches. 4. Identify various administration routes for ERT/H RT. 5. Identify currently available ERT/HRT products. 6. Recognize potential reasons for lack of initiation and continuation as well as ways to improve adherence in patients. PMID- 12125807 TI - Pycnodysostosis: role and regulation of cathepsin K in osteoclast function and human disease. AB - Patients with pycnodysostosis, a rare skeletal dysplasia, present with bone abnormalities such as short stature, acroosteolysis of distal phalanges, and skull deformities. The disease is caused by a deficiency of the cysteine protease cathepsin K which is responsible for degradation of collagen type I and other bone proteins. Osteoclasts, bone cells of hematopoietic origin responsible for bone mineral as well as protein matrix degradation, are dysfunctional in patients with pycnodysostosis due to mutations in the cathepsin K gene. Cathepsin K deficient osteoclasts can demineralize bone but cannot degrade the protein matrix. Mutations in the cathepsin K gene disrupting wild type cathepsin K activity have been described in patients with pycnodysostosis. Animal models of cathepsin K deficiency have been created and provide a valuable tool to study osteoclast function and treatment for cathepsin K deficiency. Understanding the regulation and role of cathepsin K in osteoclast function is important for designing future therapies for pycnodysostosis. Cathepsin K inhibitors will be useful in pathological processes involving excess osteoclast activation and bone resorption such as osteoporosis, bone metastasis and multiple myeloma. This review will discuss the bone remodeling cycle, the human disease pycnodysostosis caused by cathepsin K deficiency and cathepsin K activity and regulation. PMID- 12125808 TI - Neuronal ceroid lipofuscinoses caused by defects in soluble lysosomal enzymes (CLN1 and CLN2). AB - Infantile and classical late infantile neuronal ceroid lipofuscinoses (NCL) are two recent additions to the expanding spectrum of lysosomal storage disorders caused by deficiencies in lysosomal hydrolases. They are latecomers to the lysosomal storage disorders, probably because of the heterogeneous nature of the storage material, which precluded meaningful biochemical analysis. Infantile NCL is caused by deficiency in palmitoyl-protein thioesterase, an enzyme that hydrolyzes fatty acids from cysteine residues in lipid-modified proteins. Classical late-infantile NCL is caused by a deficiency in tripeptidyl amino peptidase-I, a lysosomal peptidase that removes three amino acids from the free amino terminus of peptides or small proteins. Late-onset forms of these disorders have been described. The clinical, biochemical, and molecular genetic aspects of these two latest lysosomal storage disorders are discussed in this review. In addition, approaches to treatment and future directions for research are examined. PMID- 12125809 TI - Mutated genes in juvenile and variant late infantile neuronal ceroid lipofuscinoses encode lysosomal proteins. AB - Positional cloning efforts of genes mutated in Batten disease and in the Finnish type of variant late infantile neuronal ceroid lipofuscinosis resulted in the identification of two novel genes, CLN3 and CLN5, and corresponding gene products that proved to be residents of lysosomes. Although the clinical phenotype of these NCL subtypes differs in the age of onset, average life span and EEG findings, the major component of material accumulating in patients' lysosomes is subunit c of mitochondrial ATPase in both these diseases. The CLN3 and CLN5 genes show ubiquitous expression patterns and are targeted to lysosomes in vitro, but the observed synaptosomal localization of the CLN3 protein in neurons would suggest some cell specificity in targeting and function of these proteins. So far, 31 different mutations of the CLN3 gene have been described in Batten patients, with one deletion of 1.02 kb accounting for 75% of disease alleles worldwide. Four CLN5 mutations are known, with one premature stop representing the major founder mutation in the isolated Finnish population. Functional studies of the yeast homolog of CLN3 and increased pH in patients' lysosomes would suggest an involvement of this protein in lysosomal pH homeostasis. Knock-out mouse models for CLN3 have been produced and the histopathology bears a close resemblance to human counterparts with characteristic lysosomal accumulations. Both CLN3 and CLN5 mouse models will provide experimental tools to resolve the pathological cascade in these neurodegenerative diseases. PMID- 12125810 TI - The molecular basis of mucolipidosis type IV. AB - Mucolipidosis Type IV (MLIV) is a lysosomal storage disorder that is characterized by severe neurologic and ophthalmologic abnormalities. It is a progressive disease that usually presents during the first year of life with mental retardation, corneal opacities, and delayed motor milestones. First described in 1974, MLIV is a rare autosomal recessive disease and the majority of patients diagnosed to date are of Ashkenazi Jewish descent. MLIV was originally classified as a lysosomal storage disorder due to the abnormal accumulation of mucopolysaccharides and lipids. Extensive studies in MLIV cells, however, have shown that the abnormal storage is due to a defect in the late endocytic pathway. Positional cloning led to the recent discovery of a novel gene on human chromosome 19, MCOLN1, that is mutated in MLIV. To date 14 independent mutations have been reported in MCOLN1, with two mutations accounting for 95% of the Ashkenazi Jewish MLIV alleles. The identification of the MLIV gene has led to a simple tool for definitive diagnosis and will permit carrier screening in the Ashkenazi Jewish population. MCOLN1 is a new member of the transient receptor potential (TRP) cation channel gene family. The protein encoded by MCOLN1, mucolipin-1, has six predicted transmembrane domains and a putative channel pore. The identification of mutations in MCOLN1 represents the first example of a neurological disease caused by a TRP-related channel. While the function of mucolipin-1 is currently unknown, homology to the TRP superfamily and the recent description of the C. elegans mucolipin-1 homolog allow us to begin to speculate about the role of mucolipin-1 in diverse cellular processes. PMID- 12125811 TI - Disorders of vesicles of lysosomal lineage: the Hermansky-Pudlak syndromes. AB - Hermansky-Pudlak syndrome (HPS) has evolved into a group of genetically distinct disorders characterized by oculocutaneous albinism, a storage pool deficiency, and impaired formation or trafficking of intracellular vesicles. HPS-1 results from mutations in the HPS1 gene and affects approximately 400 individuals in northwest Puerto Rico due to a 16-bp duplication in exon 15. Another 13 mutations have been reported in non-Puerto Ricans. HPS1 codes for a 79.3 kDa cytoplasmic protein of unknown function. HPS-1 patients typically develop fatal pulmonary fibrosis in their fourth decade. HPS-2 is caused by mutations in ADTB3A, which codes for the beta3A subunit of the adaptor protein-3 complex, AP3. This coat protein complex has been localized to the TGN as well as to a peripheral endosomal compartment. Evidence indicates that AP3 plays a role in the stepwise process of vesicular trafficking which leads to formation of the melanosomal, platelet dense body and lysosomal compartments. All three known HPS-2 patients had childhood neutropenia and infections. HPS-3 results from mutations in HPS3 and affects central Puerto Ricans homozygous for a 3904-bp deletion removing exon 1. At least 8 non-Puerto Rican patients have other HPS3 mutations, including an IVS5+1G->A splicing mutation in five Ashkenazi Jewish patients. HPS3 codes for a 113.7 kDa protein of unknown function. HPS-3 manifests with mild hypopigmentation and bleeding. All types of HPS are diagnosed by whole mount electron microscopic demonstration of absent platelet dense bodies, and molecular diagnoses are available for the Puerto Rican HPS1 and HPS3 founder mutations. Mouse and Drosophila models provide candidates for new genes causing HPS in humans. These genes will reveal the pathways by which specialized vesicles of lysosomal lineage arise within cells. PMID- 12125812 TI - Chediak-Higashi syndrome: a clinical and molecular view of a rare lysosomal storage disorder. AB - Chediak Higashi syndrome (CHS) is a rare, autosomal recessive disorder that affects multiple systems of the body. Patients with CHS exhibit hypopigmentation of the skin, eyes and hair, prolonged bleeding times, easy bruisability, recurrent infections, abnormal NK cell function and peripheral neuropathy. Morbidity results from patients succumbing to frequent bacterial infections or to an "accelerated phase" lymphoproliferation into the major organs of the body. Current treatment for the disorder is bone marrow transplant, which alleviates the immune problems and the accelerated phase, but does not inhibit the development of neurologic disorders that grow increasingly worse with age. There are several animal models of CHS, the beige mouse being the most characterized. Positional cloning and YAC complementation resulted in the identification of the Beige and CHS1/LYST genes. These genes encode a cytosolic protein of 430,000 Da. Sequence analysis identified three conserved regions in the protein: a HEAT repeat motif at the amino-terminus that contains several a helices, a BEACH domain containing the amino acid sequence WIDL, and a WD40 repeat motif, which is described as a protein-protein interaction domain. The presence of the BEACH and WD40 domains defines a family of genes that encode extremely large proteins. PMID- 12125813 TI - Albinism and immunity: what's the link? AB - A small number of inherited diseases show a combination of immunological and pigmentation defects. Chediak-Higashi, Griscellis and Hermansky-Pudlak syndromes are all autosomal recessive diseases with these characteristics. Recent advances in both the identification of the genes giving rise to these diseases and the cell biology of immune cells and melanocytes have begun to reveal the molecular links between immunodeficiencies and albinism. These studies identify key proteins, such as Rab27a, which are critical for secretion of specialised granules found in melanocytes and immune cells. The granules of these cells are modified lysosomes termed 'secretory lysosomes'. These studies reveal that secretory lysosomes use specialised mechanisms of secretion, not found in other cell types, which explains the selective defects in these diseases. PMID- 12125814 TI - The Niemann-Pick C proteins and trafficking of cholesterol through the late endosomal/lysosomal system. AB - To maintain proper cellular function, the amount and distribution of cholesterol residing within cellular membranes must be regulated. The principal disorder affecting transport of cholesterol through the late endosomal/lysosomal system and intracellular cholesterol homeostasis is Niemann-Pick type C (NPC) disease. The genes responsible for NPC disease have been identified, and the encoded Niemann-Pick C1 (NPC1) and Niemann-Pick C2 (HE1/NPC2) proteins are currently the subject of intense investigation. This review provides a detailed examination of NPC1 and HE1/NPC2 in regulating the transport of cholesterol through the late endosomal/lysosomal system to other cellular compartments responsible for maintaining intracellular cholesterol homeostasis, and how defective function of these proteins may be responsible for the pathophysiology associated with NPC disease. PMID- 12125815 TI - The bacteriocins of ruminal bacteria and their potential as an alternative to antibiotics. AB - Beef cattle have been fed ionophores and other antibiotics for more than 20 years to decrease ruminal fermentation losses (e.g methane and ammonia) and increase feed efficiency, and these improvements have been explained by an inhibition of gram-positive ruminal bacteria. Ionophores are not used to treat human disease, but there has been an increased perception that antibiotics should not be used as feed additives. Some bacteria produce small peptides (bacteriocins) that inhibit gram-positive bacteria. In vitro experiments indicated that the bacteriocin, nisin, and the ionophore, monensin, had similar effects on ruminal fermentation. However, preliminary results indicated that mixed ruminal bacteria degraded nisin, and the ruminal bacterium, Streptococcus bovis, became highly nisin resistant. A variety of ruminal bacteria produce bacteriocins, and bacteriocin production has, in some cases, been correlated with changes in ruminal ecology. Some ruminal bacteriocins are as potent as nisin in vitro, and resistance can be circumvented. Based on these results, ruminal bacteriocins may provide an alternative to antibiotics in cattle rations. PMID- 12125816 TI - Bacterial bioinformatics: pathogenesis and the genome. AB - As the number of completed microbial genome sequences continues to grow, there is a pressing need for the exploitation of this wealth of data through a synergistic interaction between the well-established science of bacteriology and the emergent discipline of bioinformatics. Antibiotic resistance and pathogenicity in virulent bacteria has become an increasing problem, with even the strongest drugs useless against some species, such as multi-drug resistant Enterococcus faecium and Mycobacterium tuberculosis. The global spread of Human Immunodeficiency Virus (HIV) and Acquired Immune Deficiency Syndrome (AIDS) has contributed to the re emergence of tuberculosis and the threat from new and emergent diseases. To address these problems, bacterial pathogenicity requires redefinition as Koch's postulates become obsolete. This review discusses how the use of bacterial genomic information, and the in silico tools available at present, may aid in determining the definition of a current pathogen. The combination of both fields should provide a rapid and efficient way of assisting in the future development of antimicrobial therapies. PMID- 12125818 TI - A simple sensitive program for detecting internal repeats in sets of multiply aligned homologous proteins. AB - We designed a simple but sensitive program, IntraCompare, for identifying internal repeats in families of homologous proteins. The protein sequences are aligned (Clustal X), the regions to be compared are selected, and all potential repeat sequences are compared with all others. The output provides comparison scores (GAP program) expressed in standard deviations. PMID- 12125817 TI - Enhancer-dependent transcription in Salmonella enterica Typhimurium: new members of the sigmaN regulon inferred from protein sequence homology and predicted promoter sites. AB - DNA-looping mediated by regulatory proteins is a ubiquitous mode of transcriptional control that allows interactions between genetic elements separated over long distances in DNA. In prokaryotes, one of the best-studied examples of regulatory proteins that use DNA-looping is the NtrC family of enhancer-binding proteins (EBPs), which activate transcription from sigmaN (sigma N, sigma-54) dependent promoters. The completely sequenced genome of food-borne pathogen Salmonella enterica serovar Typhimurium LT2 contains seven novel EBPs of unknown function. Four of these EBPs have a similar domain organisation to NtrC whilst surprisingly the remaining three resemble LevR in Bacillus subtilis. Probable transcriptional targets are identified for each of the EBPs, including novel homologues of phosphotransferase system Enzyme II (EII) and several virulence-associated functions. Comparisons are made with the related enteric bacteria Salmonella Typhi, Escherichia coli and Yersinia pestis. PMID- 12125819 TI - Specific growth inhibition by acetate of an Escherichia coli strain expressing Era-dE, a dominant negative Era mutant. AB - Escherichia coli Era is a GTP binding protein and essential for cell growth. We have previously reported that an Era mutant, designated Era-dE, causes a dominant negative effect on the growth and the loss of the ability to utilize TCA cycle metabolites as carbon source when overproduced. To investigate the role of Era, the gene expression in the cells overproducing Era-dE was examined by DNA microarray analysis. The expression of lipA and nadAB, which are involved in lipoic acid synthesis and NAD synthesis, respectively, was found to be reduced in the cells overproducing Era-dE. Lipoic acid and NAD are essential cofactors for the activities of pyruvate dehydrogenase complex, 2-oxoglutarate dehydrogenase complex and glycine cleavage enzyme complex. The expression of numerous genes involved in dissimilatory carbon metabolism and carbon source transport was increased. This set of genes partially overlaps with the set of genes controlled by cAMP-CAP in E coli. Moreover, the growth defect of Era-dE overproduction was specifically enhanced by acetate but not by TCA cycle metabolites both in rich and synthetic media. Intracellular serine pool in Era-dE overproducing cells was found to be increased significantly compared to that of the cells overproducing wild-type Era. It was further found that even the wild-type E. coli cells not overproducing Era-dE became sensitive to acetate in the presence of serine in a medium. We propose that when Era-dE is overproduced, carbon fluxes to the TCA cycle and to C1 units become impaired, resulting in a higher cellular serine concentration. We demonstrated that such cells with a high serine concentration became sensitive to acetate, however the reason for this acetate sensitivity is not known at the present. PMID- 12125820 TI - Multiple operons connected with catabolism of aromatic compounds in Acinetobacter sp. strain ADP1 are under carbon catabolite repression. AB - Repression of enzymes contributing to degradation of aromatic compounds via the beta-ketoadipate pathway in the presence of additional carbon sources (carbon catabolite repression) in the bacterium Acinetobacter sp. strain ADP1 is described. The phenomenon was investigated on the level of specific activity of protocatechuate 3,4-dioxygenase and p-hydroxybenzoate hydroxylase participating in catabolism of protocatechuate and p-hydroxybenzoate. Strong repression (90%) was found in cells grown on succinate and acetate in addition to the aromatic carbon source; partial derepression occurred towards the end of the logarithmic growth phase. Glucose, pyruvate, or lactate as secondary carbon sources had no repressing effect. The consumption of the aromatic substrate from the medium was delayed in the presence of acetate and succinate. The differences in specific enzyme activities were reflected at the transcript level for three operons connected to catabolism of aromatic compounds (pob, pca, van) as shown by Northern blot hybridization. Transcriptional fusions between the promoters of the pob and the pca operon identified the transcriptional level as the regulatory one. A mechanism of global regulation is postulated, which enables the organism to consume the offered carbon sources hierarchically in the most efficient manner. PMID- 12125821 TI - Bioinformatic analyses of the tRNA: (guanine 26, N2,N2)-dimethyltransferase (Trm1) family. AB - Functional analyses of the tRNA:(guanine 26, N2,N2)-dimethyltransferase (Trm1) have been hampered by a lack of structural information about the enzyme and by low sequence similarity to better studied methyltransferases. Here we used computational methods to detect novel homologs of Trm1, infer the evolutionary relationships of the family, and predict the structure of the Trm1 methyltransferase. The N-terminal region of the protein is predicted to form an S adenosylmethionine-binding domain, which harbors the active site. The C-terminal region is rich in predicted alpha-helices and, in analogy to other nucleic acid methyltransferases, may constitute the target recognition domain of the enzyme. Interposing these two domains, most Trm1 homologs possess a highly variable inserted sequence that is delimited by a Cys4 cluster, likely forming a Zn-finger structure. The residues of Trm1 predicted to participate in cofactor binding, target recognition, and catalysis, were mapped onto a preliminary structural model, providing a platform for designing new experiments to better understand the molecular functions of this protein family. In addition, identification of novel, atypical Trm1 homologs suggests candidates for cloning and biochemical characterization. PMID- 12125822 TI - Increasing the efficiency of heterologous promoters in actinomycetes. AB - An Escherichia coli-actinomycete shuttle vector, pCJW93, was constructed which places cloned genes under the control of the thiostrepton-inducible tip promoter from Streptomyces lividans. We also constructed expression vectors bearing the actII-ORF4/PactI activator-promoter system of the actinorhodin biosynthetic pathway of Streptomyces coelicolor. With both types of vector, levels of expression varied widely in different actinomycete strains, indicating different levels of the host factors needed for optimal expression. Deletion of the actII ORF4 activator gene from one such plasmid in Saccharopolyspora erythraea drastically reduced expression from the cognate actI promoter, showing that host factors are required for optimal production of the activator protein itself. However, a low copy number expression vector pWIZ1 for the polyketide synthase DEBS1-TE, in which the promoter for the activator gene was replaced by the strong heterologous ermE* promoter of S. erythraea directed highly efficient production of polyketide synthase protein in Streptomyces cinnamonensis; and the levels of triketide lactone product found were up to 100-fold greater than were produced by the same plasmid in which actII-ORF4 was expressed from its own promoter. Ensuring appropriate expression of a specific activator protein should enable more convenient and consistent heterologous expression of genes in a broad range of actinomycete hosts. PMID- 12125823 TI - A phylogenomic study of the general stress response sigma factor sigmaB of Bacillus subtilis and its regulatory proteins. AB - Regulation of expression of the general stress regulon of Bacillus subtilis is mediated by the activation of the alternative sigma factor sigmaB. Activation of sigmaB is accomplished by a complex regulatory network involving protein-protein interactions and reversible protein phosphorylation. PSI-BLAST searches were performed and phylogenetic trees for sigmaB and its regulatory proteins were constructed. Occurrence of sigmaB is restricted to a small group of gram-positive bacteria (Bacillus, Staphylococcus, Listeria). Related sigma factors also involved in stress responses are present in Mycobacterium tuberculosis, Streptomyces species and even in cyanobacteria (Synechocystis species). Putative regulatory proteins found in several other bacterial species can be broadly catagorized into three categories: Anti sigma factors, anti-anti sigma factors and phosphatases. Anti sigma factors are able to bind to sigma factors and are also kinases of anti sigma factor antagonists. Only in their nonphosphorylated state, these antagonists are able to bind to the anti sigma factor. Phosphorylated antagonists can be dephosphorylated by PP2C phosphatases. These phosphatases are of pivotal importance for activation of the sigma factor. Different phosphatases identified in this search contain a wide variety of domains found in signal transducing proteins (PAS/PAC, GAF, REC, HATase_c, HAMP). The HATPase_c domain found in several phosphatases most probably constitutes a serine/threonine kinase domain of anti sigma factors. Such proteins are most probably bifunctional anti-anti sigma factor kinases and phosphatases. The regulatory network of anti-anti sigma factors anti sigma factors and phosphatases is probably ancient and most likely evolved from a structurally similar network found in the Deinococcus radiodurans genome. In completely sequenced genomes of several bacterial species, some elements of the network are missing. The N terminus of RsbU, a phosphatase activated in response to environmental stress exhibits similarities to a region in the beta chain of phenylalanyl-tRNA synthetases. PMID- 12125824 TI - The genome of Methanosarcina mazei: evidence for lateral gene transfer between bacteria and archaea. AB - The Archaeon Methanosarcina mazei and related species are of great ecological importance as they are the only organisms fermenting acetate, methylamines and methanol to methane, carbon dioxide and ammonia (in case of methylamines). Since acetate is the precursor of 60% of the methane produced on earth these organisms contribute significantly to the production of this greenhouse gas, e.g. in rice paddies. The 4,096,345 base pairs circular chromosome of M. mazei is more than twice as large as the genomes of the methanogenic Archaea currently completely sequenced (Bult et al., 1996; Smith et al., 1997). 3,371 open reading frames (ORFs) were identified. Based on currently available sequence data 376 of these ORFs are Methanosarcina-specific and 1,043 ORFs find their closest homologue in the bacterial domain. 544 of these ORFs reach significant similarity values only in the bacterial domain. They include 56 of the 102 transposases, and proteins involved in gluconeogenesis, proline biosynthesis, transport processes, DNA repair, environmental sensing, gene regulation, and stress response. Striking examples are the occurrence of the bacterial GroEL/GroES chaperone system and the presence of tetrahydrofolate-dependent enzymes. These findings might indicate that lateral gene transfer has played an important evolutionary role in forging the physiology of this metabolically versatile methanogen. PMID- 12125825 TI - Keeping body and soul together. PMID- 12125826 TI - Independence as a practice issue in occupational therapy: the safety clause. AB - This article reports findings from interviews that explored the meanings occupational therapists attach to independence as a value and a therapeutic goal in interactions with elderly clients. Through a historical review of the literature, we trace the changing use of this term and identify two analytically distinct concepts associated with it: independence as self-reliance in activity and independence as autonomy, self-determination, or choice. We show how the latter has emerged in contemporary service contexts to represent an ideal of client-centered practice for persons with chronic disabilities, such as frail elderly clients. Using a "critical incident" interview approach with 12 Australian occupational therapists, we identified the therapists' explicit and implicit understandings of independence as a value concept and practice issue. Our findings suggest that a mismatch often exists between idealized and practice based talk about independence and that therapists narrativize this opposition around what we call "the safety clause." That is, therapists invoke concerns about safety and duty of care as a caveat to implementing their independence ideals and justifying the retention of professional control. We identify key issues that therapists need to address if the rhetoric of independence-related client-centered practice is to be achieved in reality. PMID- 12125827 TI - Effect of instructions on functional reach in persons with and without cerebrovascular accident. AB - OBJECTIVE: Verbal instructions comprise an important element of clinical practice, however, their effectiveness in promoting movement organization in persons with cerebrovascular accident (CVA) has not been well investigated. METHOD: A counterbalanced, repeated-measures design was used to examine the effects of externally focused (task-related) versus internally focused (movement related) instructions on movement kinematics during three functional reaching tasks. Participants included 16 persons with stroke who were able to perform the tasks with their affected arm and 17 age-matched adults without neurological impairments. RESULTS: Significantly shorter movement time and greater peak velocity were evident when reaching under the external-focus condition of all tasks than for the internal-focus condition. CONCLUSION: One clinical implication is that internally focused instructions can contribute to slower and less forceful reach in adults with and without CVA. This research reinforces the need for therapists to consider their use of instruction during the evaluation and treatment of movement disorders. PMID- 12125828 TI - Manual asymmetry during object release under varying task constraints. AB - OBJECTIVE: This study examined differences between the dominant and nondominant hands of children during an object release task under various accuracy and speed constraints. METHOD: Fifteen children 7 to 14 years of age who were typically developing released an instrumented object held with a precision grip onto a stable or unstable surface at a self-paced or quick speed while temporal and force measures were recorded. RESULTS: Few differences were found between the two hands under stable, self-paced conditions. However, given accuracy constraints, object replacement with the nondominant hand took longer than the dominant hand. The nondominant hand exhibited different force coordination patterns than the dominant hand when both accuracy and speed constraints were imposed. CONCLUSION: Task constraints differentially affected the performance of the dominant and nondominant hands during this object release task. The results suggest that clinicians should incorporate various task constraints into hand preference evaluations and during "dominance retraining". PMID- 12125829 TI - Outcomes of an occupational therapy program for mothers of children with disabilities: impact on satisfaction with time use and occupational performance. AB - OBJECTIVE: This experimental research study evaluated the impact of an 8-week psychosocial occupational therapy intervention program for mothers who have children with disabilities. METHOD: Thirty-eight mothers of children with disabilities were randomly assigned to participate either in the treatment or the control group (19 in each). The occupational therapy intervention was designed to facilitate increased perceptions of satisfaction with time use and occupational performance, thereby positively affecting maternal and family well-being. The Canadian Occupational Performance Measure (COPM) was administered to measure self perceptions of occupational performance and satisfaction over time. RESULTS: No significant differences were found between the two groups on time use perceptions. Although no significant differences were found between the two groups on the COPM Performance subscale, the treatment group demonstrated significantly greater score increases (p < .05) on the COPM Satisfaction subscale. CONCLUSION: This preliminary study suggests that attending to the time use and occupational concerns of mothers of children with disabilities can have a positive impact on their satisfaction with occupational performance. PMID- 12125831 TI - Stigma and its management: a pilot study of parental perceptions of the experiences of children with developmental coordination disorder. AB - The findings of a small qualitative interview study with 8 parents of 6 children with developmental coordination disorder are reported. The parents discussed the social consequences of their children's motor difficulties. The new International Classification of Functioning, Disability and Health was used as a framework for the analysis of the interview transcripts. The analysis revealed that the parents believed that their children's impairments restrict their participation in society. The interactions between impairment and participation are interpreted in the context of stigma and its management. The significance of occupational therapy interventions in the area of physical activity play to children's social life is discussed. PMID- 12125830 TI - Therapists' perceptions about making a difference in parent-child relationships in early intervention occupational therapy services. AB - OBJECTIVE: The purpose of this study was to better understand occupational therapists' experiences of making a difference in parent-child relationships. METHOD: In this qualitative, instrumental case study, occupational therapists working in early intervention were asked to reflect on and describe occasions in which they believed that they made a real difference in parent-child relationships. The primary investigator interviewed nine experienced pediatric occupational therapists. RESULTS: All nine therapists highly valued the parent child relationship and focused on these relationships in therapy. Eight themes emerged that described the therapists' practice insights and methods by which the therapists facilitated the parent-child relationship. CONCLUSION: The occupational therapists in this study reflected insights that resonate with the literature regarding the role of the parent-child relationship in the development of children. The authors raise the question about the adequacy of instruction at the pre-service level that prepares therapists to both assess and facilitate the parent-child relationship in early intervention. PMID- 12125832 TI - Implicit learning in children with and without developmental coordination disorder. AB - OBJECTIVE: The ability to use perceptual cues within the environment to guide movement accurately can be acquired implicitly in that skill may increase while the learner is not consciously aware of what cues are being used. In this study, the implicit learning capabilities of children with and without developmental coordination disorder were compared. METHOD: Twenty-two children (11 with developmental coordination disorder, 11 without the disorder) played a computer game where they "caught" a descending ball image with a paddle on the screen. The dependent variable was accuracy of catch, as measured by the score on the computer game. On training trials, a visual cue appeared 50% of the time that signaled the direction of the ball. On probe trials, the visual cue was false. After completing the task, the children were interviewed about their conscious awareness of the cue. A mixed factorial analysis of variance (ANOVA) was used to analyze the data for group comparisons. RESULTS: Of the 22 children, only 6 indicated that they were aware of the cue. A mixed factorial ANOVA was significant for greater error when the visual cue was false, F(11.12, 22.49) = 27.27, p = .000, indicating that the children responded to the cue. No difference was found between groups in game performance. CONCLUSION: Children with developmental coordination disorder in this study were able to implicitly recognize and use a perceptual cue to enhance their performance on a computer game. Strategies that foster implicit learning may be relevant to occupational therapy intervention. PMID- 12125833 TI - The importance of leisure in the lives of persons with congenital physical disabilities. AB - Although occupational therapists emphasize a balance among the three occupational areas of self-care, productivity, and leisure in people's lives, leisure often is focused on less than the other areas in both the research literature and clinical practice. Very little research has been conducted on the benefits of leisure activities in adults with congenital disabilities. The information contained in this article is a secondary analysis of the interview protocols of nine adults (30-50 years of age) with either cerebral palsy or spina bifida. The primary purpose of the interview was to determine protective processes surrounding turning points in the lives of persons with disabilities. This secondary analysis allowed us to determine the benefits and meaning of leisure for this population. Consistent with literature that focused on either persons without disabilities or persons with acquired disabilities, the participants in the present study reported that involvement in leisure activity provides mental and physical health benefits, enjoyment, opportunity to develop a self-concept and increase self esteem, and opportunities to build and enhance social relationships. All these benefits enable people to find meaning in life through doing, belonging, and understanding self in the context of their worlds. PMID- 12125834 TI - A validity study of the Evaluation Tool of Children's Handwriting-Cursive. AB - OBJECTIVES: This study examined concurrent validity of the Evaluation Tool of Children's Handwriting-Cursive (ETCH-C) by comparing ETCH-C total legibility percentage scores with handwriting grades from teachers. The study also identified the legibility percentage score discriminating between satisfactory and unsatisfactory handwriting. METHOD: The participants were 101 fourth graders who completed two handwriting instruments. Three teachers sorted the Cursive Practice and Review work sheets into A, B, C, and unsatisfactory groups to establish handwriting grade. The study compared ETCH-C total legibility percentage scores with teacher grading, using personal judgment of handwriting on the Cursive Practice and Review work sheet. RESULTS: Mean legibility percentage scores increased significantly as handwriting grade increased. The concurrent validity coefficients were .61 for ETCH-C total words and .65 for ETCH-C total letters and handwriting grade. ETCH-C legibility percentage scores discriminating satisfactory from unsatisfactory handwriting ranged from 73% to 82% on ROC (receiver operating characteristic) curves. CONCLUSION: Results support the concurrent validity of the ETCH-C with handwriting grades. A 75% word legibility percentage score discriminated satisfactory and unsatisfactory handwriting. PMID- 12125835 TI - Attaining cultural competence, critical thinking, and intellectual development: a challenge for occupational therapists. PMID- 12125836 TI - The ADL ability and use of technical aids in persons with late effects of polio. AB - OBJECTIVE: The purpose of this study was to describe functional performance in activities of daily living (ADL) and the use of technical aids among persons with late effects of polio. METHOD: Abilities in ADL of 150 participants 20 to 82 years of age were assessed with the Sunnaas Index of ADL, and the participants' use of technical aids was recorded. RESULTS: The activities in which most participants were independent were eating, daily hygiene, and communication. Many needed technical aids, adaptation of their homes, or both to perform mobility related activities and to dress or undress, take a bath or shower, cook, or manage toilet visits. In total, 86 (57%) used mobility aids such as canes, crutches, and walkers. Thirty-one (21%) used wheelchairs within or outside the home. Bath and shower aids were the most commonly used technical aids other than mobility aids. The activity where most participants depended on others was housework. CONCLUSION: In spite of their disabilities, most participants performed well in many ADL, functioning independently by using technical aids and by living in an adapted environment. PMID- 12125837 TI - Improving mobility and community access in an adult with ataxia. AB - This case report summarizes the evaluation and treatment used to provide occupational therapy services to a man living with multiple sclerosis. Primary impairments included ataxia, paraparesis, and decreased endurance. The focus of this case study was on improving the client's ability to use powered mobility to access the community despite severe ataxia. A task-oriented approach was used as a frame of reference to guide the evaluation and intervention process. The primary goals of intervention were to control the degrees of freedom required for task participation and simultaneously increase postural stability, resulting in independent control of a power wheelchair. A combination of occupational therapy interventions is illustrated, including assistive technology, positioning, orthotic prescription, and adaptation of movement patterns. PMID- 12125838 TI - Occupational therapy education in a technological world. AB - As occupational therapy curricula prepare students to meet the needs of clients living in a technological world, at SHU we have chosen to emphasize professional survival and practice in a technology learning context. The curriculum is guided throughout by the pedagogy of learning through doing (Dewey, 1916), using technology-based learning that increases in complexity and application as the program advances. This education results in the professional preparation of therapists who are equipped to apply their skills in the treatment of clients living in a modern world. Aspects of this approach, which ties technology to occupation, might be considered by other training programs in different settings. PMID- 12125839 TI - Client-centered practice, therapeutic relationship, and the use of research evidence. PMID- 12125840 TI - EBP in psychiatric OT should be strengthened. PMID- 12125841 TI - The influence of military service on outpatient care use among racial/ethnic groups in Department of Veterans Affairs medical centers. AB - This study examines race-specific military service effects on outpatient care utilization in the Department of Veterans Affairs (VA) using data from the 1992 National Survey of Veterans. The study population consisted of 4,791 male veterans. After controlling for predisposing, enabling, and need variables, black veterans were 3.7 times more likely than white veterans to use VA outpatient care. Veterans discharged from the military for medical release were less likely to use VA outpatient care (odds ratio = 0.76) than veterans discharged at the end of their normal terms. Hispanic veterans discharged for medical release were 5.3 times more likely than white veterans discharged for the same reason to use VA outpatient care. Korean conflict and mixed war period veterans were more likely to use VA outpatient care than World War II veterans. Racial/ethnic differences in military service characteristics influence the use of VA outpatient care and should be understood in delivering outpatient care to veterans. PMID- 12125842 TI - Vision readiness in Operation Restore Hope. AB - Ensuring that our forces are vision ready for their mission is essential on today's battlefield. Vision readiness considers optical readiness (appropriate correcting eyewear) and visual readiness (adequate job-required visual acuity). A study of vision readiness among deploying personnel for Operation Restore Hope in Bosnia from December 1995 to September 1997 was conducted at Fort Benning, Georgia. Of the 10,063 personnel screened, 3,554 (35.3%) were not optically ready for deployment and 406 (4.0%) were not visually ready for deployment. Analyses indicated a statistically significant difference between the active duty and reserve components in optical and visual readiness. A more effective vision readiness process should be implemented before deployment to ensure that all personnel are deployment ready. Optometry personnel, commanders, and deploying soldiers, sailors, airmen, and marines must take a more active role in ensuring that our forces have the appropriate visual acuity and optical devices to deploy. PMID- 12125843 TI - Patellar dislocation in army conscripts. AB - Between 1990 and 1996, 119 Finnish male conscripts underwent operative treatment for patellar dislocation. There were 68 conscripts (58%) with primary and 51 conscripts (42%) with recurrent patellar dislocation. Sixty-five (55%) dislocations occurred during military service, 40 (34%) occurred during sports activities, and 14 (11%) occurred during leisure time. The most common cause of injury was military training at battle exercises (n = 30). The typical injury mechanism was knee valgus rotation on fixed foot and tibia (97 conscripts, 82%). Surgical procedures performed were open in 75 conscripts (63%) and arthroscopically assisted in 44 conscripts (37%). Twenty-three (19%) redislocations occurred during follow-up (mean, 6 years; range, 3-9 years). The subjective outcome of treatment was excellent in 23 (19%), good in 42 (35%), moderate in 44 (37%), and poor in 10 (9%) conscripts. The most common residual complaint was patellofemoral pain (25 conscripts, 21%). Only 42 (35%) conscripts were able to finish their military service normally; fitness classification was decreased in 16 conscripts (13%), and 61 (52%) were temporarily exempted from military service (class E). The results of operative treatment did not differ significantly in conscripts with primary and recurrent dislocation, except for the time of first recurrence, which was significantly longer in conscripts with primary than with recurrent dislocation (27 versus 9 months). Patellar dislocation is the most common form of severe knee injury among conscripts and significantly hampers military service. Operative treatment yields only satisfactory results. Preventive measurements should be considered. A suggested algorithm for the treatment and classification of acute and recurrent patellar dislocation among military conscripts is presented. PMID- 12125844 TI - A retrospective analysis of long-term survival in severe aplastic anemia patients treated with allogeneic bone marrow transplantation or immunosuppressive therapy with antithymocyte globulin and cyclosporin A at a single institution. AB - Severe aplastic anemia can be treated with either bone marrow transplantation (BMT) or immunosuppressive therapy (IST). A retrospective review of patients with severe aplastic anemia treated with both of these modalities was conducted. Fifteen BMT and 16 IST patients were available for analysis, and follow-up of 22 and 15 years was available for the BMT and IST groups, respectively. Median survival was limited to 4.3 months in BMT patients vs. 135.2 months in IST patients, despite the older median age of the latter (22 vs. 55 years). Actuarial survival at 1 and 5 years was 87% and 78% for the IST patients and 40% and 33% for the BMT patients. Hematologic response rates, as defined by achievement of transfusion independence, were similar for the two groups. Long-term responses and survival are possible with antithymocyte globulin/cyclosporin A. PMID- 12125846 TI - Healthy People 2000 and population health improvement in the Department of Defense Military Health System. AB - The historical review of the literature presented in this article traces the current emphasis on population health improvement in the Department of Defense (DoD) Military Health System (MHS) to the emphasis placed on health promotion, disease prevention, and population health in the landmark document, Healthy People 2000. This review of the literature is presented to provide DoD MHS health care professionals with an overview of the impact of Healthy People 2000 on health promotion and disease prevention policy and on generating population health improvement initiatives within the DoD MHS. DoD MHS health care professionals who understand the evolution of population health-oriented initiatives will be better equipped to facilitate the organizational transformation necessary to embed population health improvement in the culture of the DoD MHS. PMID- 12125845 TI - The cause of death in smallpox: an examination of the pathology record. AB - OBJECTIVE: Because the cause of death in smallpox remains controversial, the human pathology record was examined. METHODS: The surviving case series of smallpox pathology in humans as well as other review articles from English language journals written during the last 200 years were reviewed. RESULTS: The skin lesions in smallpox developed as a result of viral damage and inflammation. Secondary bacterial infection did not occur until the scabs started shedding. During the papular stage of skin eruption, a secondary viremia caused focal lesions in the pharynx, larynx, tongue, trachea, and esophagus in descending frequency. The virus also caused potentially lethal interstitial pneumonitis as well as tubulointerstitial nephritis. CONCLUSIONS: The cytopathic effects of smallpox cause death. The data did not support previously promulgated theories attributing death to a bacterial sepsis syndrome seeded from the pustules or immune complex deposition. In a future outbreak, antibiotic therapy would minimally influence mortality. PMID- 12125847 TI - Veterans' care preference for coronary artery bypass grafting in a rural setting. AB - We wanted to determine what factors were associated with rural veterans' use of Department of Veterans Affairs (VA) facilities over the private sector for coronary artery bypass grafting (CABG) surgery. We reviewed the charts of 137 veterans who were referred for CABG by their VA cardiologists. Most veterans (69%) obtained CABG through the VA system. Although patients who had to drive fewer additional miles to obtain VA care were somewhat more likely to use the VA system, patients who lacked insurance or faced high out-of-pocket cost estimates for care in the private sector obtained care through the VA at dramatically higher rates. Although patients using the VA system were younger and more likely to have significant coronary artery disease, clinical outcomes did not significantly differ across systems of care. As the VA begins to understand veterans' use of multiple systems of care, it will be important to understand what influences veterans' choice of VA or private sector care. PMID- 12125848 TI - Stress fractures in Royal Marines recruits. AB - Data are presented on stress fracture patterns at the Commando Training Center Royal Marines. To reduce the high number of training injuries, including stress fractures, a complete review of the training syllabus was undertaken. Following recommendations from this review, a newly designed, physiologically progressive training program was introduced. The result of this revised training syllabus was a statistically significant reduction in stress fracture incidence. The causes of stress fractures are discussed with particular reference to skeletal adaptation to physical loading and the role of muscular support of the axial skeleton. PMID- 12125849 TI - The Combat Trauma Life Support course: resource-constrained first responder trauma care for Special Forces medics. AB - OBJECTIVE: In response to advances in civilian trauma care and changing military priorities, the United Kingdom Special Forces (SF) community undertook a complete review of its medical support in 1992 and developed a mission-orientated prehospital trauma care program known as the Combat Trauma Life Support (CTLS) program. METHODS: The course was developed dynamically, using a faculty of civilian trauma experts and military doctors to allow both medical and military doctrine to be included. RESULTS: Three scenarios were developed to cover all aspects of SF operations and civilian hospital practice. CONCLUSIONS: The CTLS course provides an evidence-based adaptable model to teach trauma care to SF soldiers operating in austere environments with limited medical equipment and prolonged evacuation times. It allows military and medical priorities to be balanced in a structured format. We believe that the development process may provide the basis for other specific needs-based prehospital trauma care. PMID- 12125851 TI - Using general population data to project idiopathic physical symptoms in the U.S. Army. AB - Many personnel deployed to the 1991 Persian Gulf War complained of symptoms that sometimes remained unexplained after comprehensive clinical examinations. We used existing population-based data from the Epidemiologic Catchment Area survey to estimate the expected incidence, prevalence, resolution, and mortality for multiple idiopathic physical symptoms (MIPS) among Army active duty personnel. Multinomial logistic regression was used to model the relationship between sociodemographic characteristics and the distribution of MIPS in U.S. workers. We used logit coefficients and known sociodemographic characteristics of U.S. Army active duty personnel to predict MIPS status in the Army. High, medium, and low estimates of MIPS incidence, prevalence, resolution, and MIPS-related mortality were obtained by altering logit intercepts. Among workers, the estimated MIPS prevalence and annual incidence, resolution, and mortality were 4.62%, 2.11%, 63.85%, and 0.26%, respectively. In contrast, the same predicted rates among Army active duty personnel were 3.89%, 1.64%, 71.33%, and 0.15%. PMID- 12125850 TI - Sulfur mustard intoxication, oxidative stress, and antioxidants. AB - Sulfur Mustard (SM) is a potent alkylating agent with electrophilic property which has been used as a chemical warfare agent in at least 12 conflicts. It has reemerged as a major threat in recent years. Medical attention is primarily concerned with its action on the skin, eyes, and respiratory tract which may be complicated by damage to ophthalmic, pulmonary, and gasterointestinal systems, followed by bone marrow depression. The cytotoxicity of SM and production of reactive oxygen substances (ROS) has been proposed to result from electrophilic or oxidative stress with depletion of cellular detoxifying thiol levels including glutathione. Also, ROS are transformed by iron-requiring reactions into highly toxic oxidants that cause a chain reaction with membrane phospholipids to form lipid peroxides, leading to loss of membrane function, membrane fluidity, and finally membrane integrity. Provision and availability of scavengers of ROS and electrophilic compounds such as glutathione, sulfhydryls compounds, antioxidants, and substances that will increase production of endogenous scavengers may be considered protective and useful. Thereby, the role of substances such as selenium, copper, zinc, and antioxidants including vitamin E, vitamin C, and compounds like beta-carotene against SM cytotoxicity and lipid peroxidation might be interesting to be investigated in experimental animal models. PMID- 12125852 TI - Prevalence of seasonal affective disorder among U.S. Army soldiers in Alaska. AB - OBJECTIVES: The goal of this study was to examine the prevalence of seasonal affective disorder among 1,116 active duty men and 191 active duty women stationed at an Army installation in Alaska. METHOD: Prevalence estimates of seasonal affective disorder (SAD) were calculated in two different ways. The first was based on the "classic" criteria in which only January and February are regarded as the winter months. The second was based on the "arctic" criteria in which November and December are included as winter months. RESULTS: Prevalence estimates of classic SAD were 5.2% for men and 8.9% for women. Prevalence estimates of arctic SAD were 6.5% for men and 13.1% for women. Subjects who met arctic criteria but not classic criteria for SAD self-reported significantly more current depression than those who met classic criteria. CONCLUSION: The question of whether individuals meeting arctic criteria have a more severe form of SAD may be an area worth exploring. PMID- 12125853 TI - Are overweight and obese airmen at greater risk of discharge from the United States Air Force? AB - This study examined whether overweight or obesity results in greater discharge risk from basic military training (BMT) or during the first year of service in the U.S. Air Force (USAF). Participants were 32,144 male and female airmen who underwent BMT from August 1995 to August 1996 and were classified as underweight (body mass index [BMI] < 18), normal range (BMI = 18-24.9), overweight (BMI = 25 29.9), or obese (BMI > or = 30). Underweight airmen were 63% more likely and overweight airmen were 24% more likely to be medically discharged during BMT compared with individuals in the normal range. Underweight airmen were 22% more likely to be discharged within the first year of service compared with those in the normal range, whereas overweight airmen were 15% less likely to be discharged. These findings do not support overweight or obesity as being major causes for discharge from the USAF within the first year of active duty. PMID- 12125854 TI - Attitudes to smoking on submarines: results of a questionnaire study. AB - An anonymous survey to assess the attitudes to smoking of men serving on two Trident Nuclear submarines was conducted by questionnaire. A total of 244 questionnaires were completed, representing 87% of the two crews. Thirty-two percent of respondents declared themselves to be smokers, 69% were nonsmokers, and of these, 31% were ex-smokers. Attitudes of all respondents to an enforced ban of smoking on submarines indicated that 55% felt that it would be justifiable, 46% felt that it would be unfair, 42% felt that it was uncalled for, 46% thought that it would be illegal, and 47% thought that a ban was about time. The separate opinions of smokers and nonsmokers were polarized, whereas the overall results indicate indifferent attitudes of crew members. Further research into the atmospheric effects of environmental tobacco smoke on a submarine is required. PMID- 12125855 TI - Injuries and injury prevention among senior military officers at the Army War College. AB - Injuries and activities associated with injuries were extracted from a retrospective review of the medical records of officers attending the U.S. Army War College during academic years 1999 and 2000 (AY99 and AY00). In AY99, cumulative injury incidence (officers with one or more injuries) was 56%. The next year (AY00), there was command emphasis on injury reduction and education of students on injury prevention strategies. Cumulative injury incidence in AY00 was 44%, significantly lower than in AY99 (p = 0.01, risk ratio [AY99/AY99] = 1.3, 95% confidence interval = 1.1-1.5). Among activities that could be linked to injuries, sports were associated with 41% in AY99 and 45% in AY00. Recommendations for ongoing injury reduction include the following: (1) continued command emphasis and instruction on injury reduction techniques; (2) encouraging the use of semirigid ankle braces to reduce ankle sprains; (3) reducing the number of practice and game sessions in sports activities; (4) encouraging overrunning of second and third base in softball; (5) prohibiting contact with the center line below the net in volleyball; and (6) encouraging medical care providers to record the activity associated with each injury in the medical records. PMID- 12125856 TI - The prevalence of asymptomatic Chlamydia trachomatis in military dependent adolescents. AB - Chlamydia trachomatis infection is common among sexually active populations and often asymptomatic. Infection is associated with complications, including pelvic inflammatory disease and infertility. Using a noninvasive urine ligase chain reaction assay, we determined the prevalence of asymptomatic C. trachomatis infection among sexually active military dependent adolescents and young adults attending clinics at military facilities in San Antonio. The overall prevalence rate was 14%, higher than that reported in many high-risk settings, including sexually transmitted disease clinics. Gender-specific rates were 15% for females and 11% for males. Given a clear cost benefit to screening and treating at-risk populations, we conclude that screening of all sexually active military dependents, both male and female, using this noninvasive test should be performed routinely. PMID- 12125857 TI - The impact of commuter war on military personnel. AB - BACKGROUND: The U.S. armed forces recently experienced a reduction in forces along with an increase in operational tempo. The Air Force and other branches supporting its mission share this common military burden and also experience a unique stressor. The Air Force has developed a military force that can fight by night and return home by day. The relatively new phenomenon of "commuter war" was especially evident during Operation Allied Force over Kosovo. METHODS: Military personnel (N = 540) participating in Operation Allied Force were administered a survey measuring morale, wellness behaviors, and work-family conflict. RESULTS: The deployment had adverse effects on wellness behaviors of permanent party and temporary duty assignment populations. Additionally, levels of morale and motivation varied between the two groups. Permanent party personnel also reported increased rates of work-family conflict. CONCLUSIONS: Commuter war affects wellness behaviors, morale, and work-family conflicts of military personnel. PMID- 12125858 TI - A 55-year-old mechanically ventilated male requiring aeromedical evacuation. AB - OBJECTIVE: To present a case that illustrates the problems unique to transporting a mechanically ventilated patient by air. A 55-year-old mechanically ventilated male with Guillain-Barre Syndrome, a condition with respiratory effects often similar to those of traumatic brain injury, required air transport from Walter Reed Army Medical Center in Washington, DC, to a hospital in Nevada. A medical team, including one physician, one nurse, and one respiratory therapist, accompanied the patient. This team was not trained in air travel and its unique risks. To complete the mission they had to rapidly familiarize themselves with the specific risks of air travel and the precautions that should be taken. This case is presented to illustrate these risks and what can be done during flight to minimize them. PMID- 12125859 TI - Good Lord deliver us--revisited. PMID- 12125860 TI - Primary care outcomes and provider practice styles. PMID- 12125861 TI - Evaluation of various models of propane-powered mosquito traps. AB - Large cage and field studies were conducted to determine the efficacy of various models of propane-powered mosquito traps. These traps utilized counterflow technology in conjunction with catalytic combustion to produce attractants (carbon dioxide, water vapor, and heat) and a thermoelectric generator that converted excess heat into electricity for stand-alone operation. The cage studies showed that large numbers of Aedes aegypti and Ochlerotatus taeniorhynchus were captured and that each progressive model resulted in increased trapping efficiency. In several field studies against natural populations of mosquitoes two different propane traps were compared against two other trap systems, the professional (PRO) and counterflow geometry (CFG) traps. In these studies the propane traps consistently caught more mosquitoes than the PRO trap and significantly fewer mosquitoes than the CFG traps. The difference in collection size between the CFG and propane traps was due mostly to Anopheles crucians. In spring 1997 the CFG trap captured 3.6X more An. crucians than the Portable Propane (PP) model and in spring 1998 it captured 6.3X more An. crucians than the Mosquito Magnet Beta-1 (MMB-1) trap. Both the PP and MMB-1 captured slightly more Culex spp. than the CFG trap. PMID- 12125862 TI - Malaria vectors in Bioko Island (Equatorial Guinea): PCR determination of the members of Anopheles gambiae Giles complex (Diptera: Culicidae) and pyrethroid knockdown resistance (kdr) in An. gambiae sensu stricto. AB - Anopheles gambiae sensu lato Giles, 1902 and Anophelesfunestus Giles, 1900 are the main malaria vectors on the island of Bioko (Equatorial Guinea). This study was carried out to determine: a) members of the An. gambiae complex that may be present on the island of Bioko and, b) the sensitivity of An. gambiae sensu stricto to pyrethroids. The analysis by PCR detected the presence of An. gambiae s.s. as the major vector of the complex and the "forest chromosomal form" was demonstrated by cytogenetic analysis. The presence of Anopheles melas in the southwest, north and southeast of the island justifies its study as a vector. The molecular characterization of pyrethroid knockdown resistance (kdr) showed that the populations of An. gambiae s.s. were sensitive and no mutations were found. This fact justifies the implementation on a large scale of pyrethroid-impregnated bednets within the framework of the Malaria Control Program of Equatorial Guinea. PMID- 12125864 TI - Spatial occurrence and hatch of field eggs of the tadpole shrimp Triops newberryi (Notostraca: Triopsidae), a potential biological control agent of immature mosquitoes. AB - The tadpole shrimp (TPS), Triops newberryi (Packard) (Notostraca: Triopsidae) is a potential biological control agent for immature mosquitoes breeding in ephemeral habitats. The occurrence of TPS eggs in soil and their hatch were investigated in 11 flood-irrigated date gardens in the Coachella Valley of southern California in 1999. Each garden was sampled several times after the rows were recently irrigated. All these date gardens harbored from very few to a large number of eggs in the soil. Overall, the average density of total eggs on ranches with clay loam soil was significantly higher than that on ranches with silt loam soil. The average densities of total eggs were significantly lower on the ranches that were disked compared to those on the ranches that were undisked before sampling. Two types of eggs were found and designated as "fresh" (yellowish to brownish) and "old" (blackish) eggs. This is the first time that these dimorphic eggs have been reported. The density of fresh eggs was lower than that of old eggs in most soil samples. The date gardens with high egg densities were sampled for determination of vertical occurrence, where soil was sampled up to 38.5 cm deep. Fresh eggs were recovered from soil in depths up to 25.6 cm, but the densities progressively declined with depth. The old eggs, however, were recovered from all soil depths studied, and there was no obvious relationship between soil depth and their density. This pattern of vertical occurrence of TPS eggs is the result of frequent disking for weed control and fruit harvest. Hatch of TPS eggs in surface soil samples ranged from 0 to 7.2 per 100 g dried soil. Hatch of viable eggs had an inverse relationship with soil depth. No TPS hatched out from the soil samples taken deeper than 15.4 cm. Fresh and old eggs distinguished by color were subjected to hatching tests. Fresh eggs exhibited high hatch, with hatching rates of 35.5-45.0% and 40.2-60.3% for the 1st and 1st plus the 2nd hydrations respectively. The old eggs, however, did not hatch at all. These findings provide quantitative information with regard to occurrence of natural TPS populations in flood irrigated agricultural fields, which could serve as a potential regulatory force of immature mosquito populations sharing ephemeral habitats with TPS. PMID- 12125863 TI - Flea, rodent, and plague ecology at Chuchupate Campground, Ventura County, California. AB - Chuchupate Campground in Ventura County, California, was closed to the public for 18 years (1982 to 2000) because of uncontrolled vector fleas and persistent plague antibody titers in rodents. The primary purpose of this study was to clarify the plague ecology of Chuchupate Campground by identifying involved rodents and their vector fleas and by determining many of their ecological parameters: abundance, flea and host preferences and diversities, and flea seasonality. Rodents and fleas were identified to species, some fleas were tested for Yersinia pestis, and rodent bloods were analyzed for the presence of antibodies to Y. pestis. During this study, 20 flea species were identified from 10 rodent and one lagomorph species collected. Five species of rodents were seropositive for plague during 13 of the 17 years in which plague testing was conducted. A likely reservoir species was not determined, but evidence of plague resistance was discovered in Merriam's chipmunks (Tamias merriami) and dusky footed woodrats (Neotoma fuscipes). The "susceptible" rodent and flea complexes at Chuchupate are the California ground squirrel (Spermophilus beecheyi) and its fleas, Oropsylla montana and Hoplopsyllus anomalus, Merriam's chipmunk and its flea, Eumolpianusfornacis, and the dusky-footed woodrat and its flea, Orchopeas sexdentatus. Host preference, diversity, and seasonality of fleas are discussed, as well as the pivotal role of woodrat houses and nests as foci for hosts, fleas, and plague. PMID- 12125865 TI - Introduction and establishment of tadpole shrimp Triops newberryi (Notostraca: Triopsidae) in a date garden for biological control of mosquitoes in the Coachella Valley, Southern California. AB - Tadpole shrimp (TPS), Triops newberryi (Packard), has been reported to have a potential as a biocontrol agent for larval mosquitoes breeding in intermittently flooded habitats. To develop and promote this predator for controlling mosquitoes, a date garden devoid of preexisting TPS populations was chosen in the Coachella Valley, southern California in 2000 to receive introductions of TPS eggs and mature TPS. Mosquito control by TPS was assessed in the plots one year after their introductions. In a selected block on this ranch, 2 rows were stocked with TPS eggs, where soil containing approximately 2,000 eggs was spread over the surface of dry ground in each row before flooding. Another 2 rows were used for mature TPS introduction, where about 400 mature TPS were released into standing water in each row 1 day after flooding. After a single egg or mature TPS introduction, active TPS in water and viable eggs in dry surface soil were noted in increasing numbers during the 3-4 subsequent irrigations. Disking before irrigation, which turned the eggs over and mixed them into the soil column, reduced TPS egg populations at the soil surface and subsequent active TPS populations in standing water after irrigation. After one or two irrigations, viable eggs and active shrimp were found in centers adjacent to the introduced plots in the stocked rows. Ample evidence is presented to show that TPS populations were established after a single introduction of eggs or mature TPS. TPS eggs and/or newly hatched TPS were also carried into the neighboring rows across the borders by the overflowing irrigation water, and TPS populations became established there too, as active TPS were noted after each irrigation in the adjacent unstocked rows. Considering the ease and economical storage, transportation and handling, dessication--resistant eggs have advantages over mature TPS for field introductions. Mosquito control by TPS was assessed in rows with and without TPS in July 2001, one year after TPS introductions. Production of Psorophora columbiae Dyar and Knab and TPS populations were determined 4-5 days after each of 2 irrigations, when there was no or little vegetation inside the rows. As compared with the row without TPS, the presence of relatively high numbers of TPS reduced Ps. columbiae by 73 to 99% as based on the average numbers of larvae, pupae and exuviae per dip in the rows with versus without TPS. PMID- 12125866 TI - Introduction of Aedes albopictus (Skuse) in southern California and potential for its establishment. AB - Isolated incidences of Aedes albopictus in the continental U.S. were reported as early as 1946 and the first incidence in California was reported in 1972. These introductions were referred to as "isolated incidences" because very few immatures were observed in used tires shipped from Southeast Asia. The first major discovery of a large population and subsequent establishment of Ae. albopictus in the U.S. was reported in 1986 from Houston, TX, in a shipment of used tires from Japan. In early June 2001, infestations of this species associated with containerized oceanic shipments of "lucky bamboo" (Dracaena spp.) packaged in standing water were introduced into southern California from mainland south China. Focal points of infestation are currently at the wholesale nurseries in southern as well as northern California. A control protocol for adulticiding and larviciding has been implemented by the Greater Los Angeles County Vector Control District. Surveys are presently being conducted by local vector control agencies in southern and northern California to determine the extent of infestation. Potential forAe. albopictus establishment is discussed. PMID- 12125867 TI - Seasonal abundance of Culex nigripalpus Theobald and Culex salinarius Coquillett in north Florida, USA. AB - North Florida is a transition zone between widespread Culex nigripalpus populations to the south and focal Culex salinarius populations to the north. Culex nigripalpus is a vector of St. Louis encephalitis (SLE) and eastern equine encephalitis (EEE) viruses in south Florida, while Cx. salinarius is a suspected New World vector of West Nile (WN) virus. Abundant vector populations are often a prerequisite for epidemic and epizootic transmission of arboviruses. Extensive SLE transmission has never been reported from north Florida, but sporadic WN transmission was reported there during the summer of 2001. The disparate flavivirus transmission patterns observed in north and south Florida may be due, in part, to the local geographical and seasonal distribution of Culex vectors. Here we report that from May 1991 to April 1994, Cx. salinarius was most commonly observed during the winter and spring in northeast Florida (Duval County), whereas Cx. nigripalpus was most abundant during the summer and autumn. An unusually mild spring in 1991 allowed Cx. nigripalpus to reproduce early in the year, resulting in a summer population that emerged more than 8 wks earlier than in 1992 and 1993. The 1991 Cx. nigripalpus population persisted through October, when SLE transmission was detected by sentinel chickens. Transmission of SLE was not detected in Duval County during 1992 or 1993. These data indicate that mild winter and spring conditions in north Florida may favor increased abundance and survival of Cx. nigripalpus in a region where this species is normally not abundant. A seasonal shift in population structure may increase the transmission risk of arboviruses for which Cx. nigripalpus is a competent vector, including SLE, WN, and EEE. PMID- 12125868 TI - Selected literature references to vector ecology--October 2001-March 2002. PMID- 12125869 TI - Spatial distribution of adult Anopheles darlingi and Anopheles albimanus in relation to riparian habitats in Belize, Central America. AB - Collections of Anopheles darlingi Root and An. albimanus Wiedemann from central and northern Belize were conducted as landing captures from 6:30 to 8:00 p.m. to define spatial distributions and outdoor:indoor ratios of biting during the early evening. In central Belize, collections were made at 31 houses in riparian zones (> or = 1 km from rivers) and 14 houses in upland zones (>1 km from rivers) during the dry and wet seasons of 1993 and 1994. Females of both species were abundant in houses < or = 1 km from rivers. Females were not present in houses located in upland areas during the dry season, but were present in the wet season. A total of 63 paired collections (representing 130 individual captures) from 42 houses showed An. darlingi females were more endophagic (ratio of 1:0.6) during the early evening than were An. albimanus females (ratio of 1:0.21). Paired landing collections from 22 houses in riparian zones in April-May were analyzed in an index of species abundance (ISA). ISA values rated An. darlingi as the dominant Anopheles mosquito indoors and An. albimanus was dominant outside. Although An. darlingi and An. albimanus were abundant in riparian zones, there was no association in their numerical abundance, suggesting that different environmental factors influenced their abundance. In northern Belize, one house for each of 16 villages was sampled during April and May 1994. Large numbers ofAn. albimanus were captured outdoors in houses located in riparian and marshland areas (means of 217.5 and 247.5/1.5 personhours outdoors, respectively). Numbers of An. albimanus were low at houses located away from rivers and marshes (12.2 per collection). Anopheles darlingi was uncommon at sites in northern Belize. Proportionally fewer An. albimanus females entered houses in the north (outdoor:indoor ratio of 1:0.16) compared to the central region (ratio of 1:0.21), which probably reflects differences in house construction, anti-mosquito behavior (i.e., closing windows and doors at sunset), and insecticide treatments. The ISA gave a quantitative assessment of vector dominance in relation to the parameters of spatial distribution and numerical abundance. The index was also sensitive to the variables of indoor and outdoor biting behaviors. PMID- 12125870 TI - Leaf litter decay process and the growth performance of Aedes albopictus larvae (Diptera: Culicidae). AB - Larvae of the mosquito Ae. albopictus typically develop in small aquatic sites such as tree holes and artificial containers. Organic detritus, in particular decaying leaves, is therefore their major carbon source. Here we demonstrate the importance of leaf characteristics, and in particular their rates of decay, in determining the development and survivorship of larvae. We compared the effects of a rapidly decaying leaf, the maple Acer buergerianum (Angiospermae: Aceraceae) and a slowly decaying leaf, the camphor Cinnamomum japonicum (Angiospermae: Lauraceae), on the larval development of Ae. albopictus at different larval densities in laboratory microcosms. Overall, the maple leaves provided a better substrate and the observed growth patterns could be explained on the basis of a difference in nutritive and chemical contents of the two leaf types. At the highest population density, the duration of the larval period was much shorter in maple litter microcosms. Larval mortality gradually increased with population density in the camphor treatment. In contrast in the rapidly decaying leaf litter microcosms, mortality remained low even as densities increased. Mean pupal size was greater in the individuals fed on the rapidly decaying leaf litter as well as at lower density. Size is likely to be correlated with fitness in the field. In general, rapidly decaying leaf litter will favor mosquito growth resulting in quicker development and higher population sizes. This work emphasizes the importance of the local environment on the development of vector mosquitoes and has important implications for control. PMID- 12125871 TI - The effects of diet upon pupal development and cocoon formation by the cat flea (Siphonaptera: Pulicidae). AB - Cocoon formation by cat flea larvae was directly related to the quantity of eggs or yeast consumed. Larvae consuming either 1-3 eggs or 0.25-1.0 mg of yeast developed as naked pupae or formed incomplete cocoons. Third instar cat flea larvae fed upon naked pupae and pupae within partial cocoons, but complete cocoons protected pupae from cannibalism. First and second instars did not attack pupae. When larvae were provided with a carpet fiber for protection, a greater number of fleas successfully developed to the adult stage. PMID- 12125872 TI - Studies on the bionomics of Anopheles dirus (Culicidae: Diptera) in Mudon, Mon State, Myanmar. AB - This study examined some environmental factors influencing the larval habitats of Anopheles dirus (breeding in wells) in Mudon, Myanmar, from May 1998 to March 2000. The larval/pupal density was found to be directly proportional to rainfall and indirectly proportional to the well water level. Shade, vegetation and debris on the surface of well water were important factors influencing the abundance of the aquatic stages of An. dirus. Salinity had an inverse correlation with the larval and pupal density. Other mosquito species associated with An. dirus were identified. Important predators of the mosquito larvae were larvivorous fish, damselfly and dragonfly nymphs. All wells examined were lined with lateritic rocks. Chemical analysis of water samples from wells was conducted. PMID- 12125873 TI - Use of liquid deltamethrin in modified, host-targeted bait tubes for control of fleas on sciurid rodents in northern California. AB - The efficacy of liquid deltamethrin was evaluated for controlling fleas on golden mantled ground squirrels, Spermophilus lateralis, and chipmunks, Tamias amoenus and T. senex. A host-targeted bait tube was modified to deliver insecticide to visiting rodents during a seven-week trial conducted in the Southern Cascade Mountains of northern California. A single deltamethrin treatment with one replenishment of bait provided effective flea control on rodents within one week of deployment of bait tubes. A second treatment accompanied by two bait replenishments during the following two-week period maintained effective control throughout the seven-week period, even though all bait tubes were removed from the site after three weeks. Our results suggest that the use of modified bait tubes treated with liquid deltamethrin can provide an effective, economical, and efficient means of controlling vector fleas on the primary disease amplifying rodent hosts in this plague endemic region of California. PMID- 12125874 TI - Role of residual spraying for malaria control in Belize. AB - We studied the impact of reduced residual spraying in Belize by developing a logistic regression model on relationships between numbers of houses sprayed (mostly with DDT) and numbers of malaria cases. We defined the "minimum effective house spray rate" (MEHSR) as the level of spraying that will prevent increases in malaria rates for a defined population. Under the total coverage approach (all houses sprayed), the MEHSR for Belize was 134.6. The model also showed that the odds for decreasing malaria is 1.086 for each increase of 10 houses sprayed per 1,000 population. In further testing, highly significant and differential changes in malaria rates were documented for paired groups of years with house spray rates that were either above or below the MEHSR. Numbers of malaria cases since 1995 are used to show how stratification methods are used in Belize to spray fewer houses (at levels below the MEHSR of 134.6). PMID- 12125875 TI - Time of survival under starvation in two flea species (Siphonaptera: Pulicidae) at different air temperatures and relative humidities. AB - We studied the effect of air temperature and RH on the survival time of adult Xenopsylla conformis Wagner, 1903 and Xenopsylla ramesis Rothschild, 1904 fleas during starvation to explain the paratopic habitat distribution of these species on opposite ends of a precipitation and temperature gradient in the Negev Highlands, Israel. We hypothesized that the pattern of distribution of the two flea species is caused by differential microclimatic preferences of imagoes and predicted that (1) the resistance to starvation would differ between the two flea species at different air temperatures (10 degrees C - 38 degrees C) and relative humidities (RH) (40%-92%) and (2) survival time of starving X conformis would be longer than that of starving X ramesis at high air temperatures and low RHs. Survival time of newly emerged X conformis was dependent on air temperature but not on RH, whereas in newly emerged X ramesis it was affected by both air temperature and RH. Generally, survival time of newly emerged fleas was longer at lower air temperatures and higher humidities than at higher air temperatures and lower humidities. No significant difference in survival time between species in dependence on either air temperature or RH were found for newly emerged fleas. Fed fleas of both species responded similarly to changes in air temperature and RH in terms of survival time. Survival time at lower temperatures was longer than that at higher temperatures. Females survived longer than males at all air temperatures except for the highest temperature when the survival time of both sexes was similarly low. In both species, the effect of RH on survival time was significant at the highest RH only, with a decrease in survival time at 92% RH. In contrast, survival times at lower RHs did not differ. Males of both species survived for less time than females at all RHs. X conformis generally survived for less time than X ramesis, all else being equal. The only regime at which X conformis survived longer than X. ramesis was 38 degrees C and 40% RH. Newly emerged fleas survived for significantly longer time than fed fleas. PMID- 12125876 TI - Habitat size: a factor determining the opportunity for encounters between mosquito larvae and aquatic predators. AB - Occurrence patterns of mosquito immatures and insect predators in containers of various sizes were surveyed in summer (June-July) and autumn (September) of 1998 in a rural area of Saga, southwestern Japan. Mosquitoes were categorized into three types in relation to habitat size. First, Aedes (Stegomyia) spp. and Tripteroides bambusa occurred mostly in small containers of < 0.1 m2. Second, Ae. japonicus and Culex kyotoensis occurred in larger container sizes, compared with the first group. Third, Cx. tritaeniorhynchus and Anopheles sinensis occurred in rice fields in summer and in large containers in the autumn. Predators such as Notonectidae, Anisoptera nymphs, and Chaoborus sp. and a predaceous mosquito Cx. halifaxii occurred mainly in large (> or = 0.1 m2) containers. The mosquitoes of the third group showed similarities with predators in the occurrence of each habitat type, and they frequently co-occurred with predators. The mosquitoes of the first group showed less similarity with predators in habitat type preference, and they rarely co-occurred with predators. The second group mosquitoes showed intermediate patterns of the first and the third groups. PMID- 12125877 TI - Isolation of Salmonella from muscoid flies at commercial animal establishments in San Bernardino County, California. AB - Adult muscoid flies collected from three dairies and eight commercial poultry ranches in San Bernardino County were tested for Salmonella enteritidis. Of the total 2,686 flies tested, 92.3% were Musca domestica, 5.5% Fannia canicularis, 1.9% Ophyra leucostoma, 0.2% Phaenicia sericata, and <0.1% Muscina stabulans. Whereas flies at the dairies belonged exclusively to M. domestica, faunal composition at the poultry ranches was more diverse, including all five species. Among the five fly species, only M. domestica tested positive for serotypes of S. enteritidis. The six isolates from fly pools (25-50 flies per pool) collected from dairies included five S. enteritidis serotype Kentucky and one S. enteritidis ser. Muenster. Isolates from flies collected at poultry ranches were three S. enteritidis ser. Braenderup, S. enteritidis ser. Kottbus, and S. enteritidis ser. Montevideo. The male to female ratio in M. domestica testing positive for S. enteritidis serotypes, was 3 to 8 pools, showing more female than male flies carrying the organism. PMID- 12125878 TI - Does temperature affect the outcome of larval competition between Aedes aegypti and Aedes albopictus? AB - The superior larval competitive ability of Aedes albopictus has been proposed to explain the recent displacement of Aedes aegyptiby the former species inparts of the southeastern U.S. Ae. aegypti persists, however, in sympatry with Ae. albopictus in urban areas of southern Louisiana, Florida, and Texas, and the impact of larval competition between these species has not been investigated at higher temperatures that may be characteristic of these urban environments. We compared growth and survivorship of the two species at controlled temperatures of 24 degrees and 30 degrees C in water-containing tires under conditions of intra- and interspecific competition and with or without leaf litter. When other variables were controlled statistically, the estimated finite rate of increase (lambda') was significantly higher for both species at the higher temperature, and the proportional increases in lambda' did not differ between species. Therefore, our experiment predicts that by itself, temperatures between 24 degrees and 30 degrees C would not alter the outcome of larval competition. Overall, response measures of Ae. albopictus were more sensitive than those of Ae. aegypti to the litter and species/density variables, although the development ofAe. aegypti females was uniquely retarded by a high density of its own species. PMID- 12125879 TI - Vertical transmission of Rickettsia felis in the cat flea (Ctenocephalides felis Bouche). AB - Rickettsia felis can be maintained in cat fleas by vertical transmission for up to 12 generations without the benefit of an infected host. Horizontal transmission or the acquisition of R. felis by fleas feeding on cats or artificially infected meals were not demonstrated in this study. Horizontal transmission of R. felis by the ingestion of feces or eggs by flea larvae was not detected. We also tested for and found no evidence to support horizontal transmission by contact among positive fleas and negative fleas. PMID- 12125880 TI - Safety issues as exemplified by the activities of the Joint Commission on Accreditation of Healthcare Organizations: context for the safety and disclosure standards. AB - The author, the General Counsel of the joint Commission on Accreditation of Healthcare Organizations, discusses the concerns that have been raised about the Commission's Patient Safety standards and Sentinel Event policy. Mr. Bressler understands the view that compliance with the Commission's standards and procedures (which are designed to help advance safety in our healthcare system) could, in particular cases and under certain circumstances, heighten litigation risks, but emphasizes the benefits attainable from systematic analysis of errors. Moreover, the article concludes that thoughtful lawyering by counsel for healthcare organizations can help minimize any risks, pending legislation establishing the needed certainty in the law. PMID- 12125881 TI - The new JCAHO patient safety standards and the disclosure of unanticipated outcomes. Joint Commission on Accreditation of Healthcare Organizations. AB - This article looks at the newly-issued JCAHO standards and their increased focus on patient safety in performance standards for healthcare organizations. As part of these standards, hospitals are required to inform patients of outcomes of care, including unanticipated outcomes. This article examines this requirement and suggests varying interpretations of it. After looking at the current legal and ethical standards requiring disclosure of errors or negligent acts, the article suggests that hospitals are faced with many difficulties in implementing the standard. Specifically, the article argues that more details are necessary regarding what events must be reported and what hospitals are required to do when members of the medical staff refuse to inform patients of medical error. PMID- 12125882 TI - Unanticipated consequences of unanticipated outcomes disclosures. AB - This article discusses the complex legal and operational issues related to the Joint Commission on Accreditation of Healthcare Organizations' (JCAHO) unanticipated outcomes disclosure standard. The author examines case law on the discoverability of hospital quality improvement efforts and information submitted to the JCAHO, and considers whether submitting this information to JCAHO amounts to a waiver of privilege. The author concludes disclosure of an unanticipated outcome should not give rise to a waiver if the hospital and physician are careful in making the disclosure. PMID- 12125883 TI - The fiduciary duties of healthcare directors in the "zone of insolvency". AB - Directors of healthcare organizations normally owe fiduciary duties to their shareholders or, in the case of nonprofits, to the charitable mission of the organization. As an organization descends to bankruptcy, however, the board's duties may shift. At some point, the board may be imposed with different and often conflicting obligations to the corporate enterprise as a whole, with a primary criterion being the interests of creditors. In this article, the authors analyze the murky areas of the Zone and give guidance as to when the board's duty may shift-and as to how directors should proceed both in determining their duties and in working to fulfill them. PMID- 12125884 TI - The implementation of quality and safety measures: from rhetoric to reality. AB - The evolution of American healthcare is the history of recurrent attempts to enhance quality of care. The latest wave of debate in this area was sparked by the 1999 Institute of Medicine Report, To Err is Human. In the current debate over how best to improve safety and quality, however, there has been a disconnect between the theoretical optimum and the practical possibilities. The author examines the history of our nation's efforts at healthcare improvement over the years, and concludes that the present debate will likely lead to improvements- but only after a messy (and necessary) political struggle over what price Americans are willing to pay for improvement, and where that money will come from. When reform occurs, the author believes that it will be on multiple levels, with accreditation organizations and various levels of government acting in the manner (and for the constituencies) appropriate to each. Ultimately, the political processes will yield a variety of approaches to be watched, studied, and amended as the healthcare system evolves to provide safer, more effective care. PMID- 12125885 TI - Checklist of federal and state privacy issues. PMID- 12125886 TI - Three creatures named 'forward model'. AB - It has recently been suggested that the nervous system employs forward models for the purpose of motor control. The evidence for this hypothetical computational structure comes from various sources, theoretical and experimental. The purpose of this commentary is to distinguish between three different structures that are used to support the forward model hypothesis: (1) output predictor, (2) state estimator, and (3) distal teacher. It is possible that the brain employs all these structures for control. However, the general term forward model could be misleading since evidence for one structure could be misinterpreted as supporting evidence for the other structures. PMID- 12125887 TI - A control model of the movement of attention. AB - A control model of the movement of the focus of attention is developed and applied to explain its observed effects on single cell activity and to various quantitative features of the Posner benefit paradigm. This supports the presence of an inverse controller and a rules component in the control model. The ability of the control model to explain a range of deficits is then analyzed, as is its relation to other modeling approaches. PMID- 12125888 TI - A local and neurobiologically plausible method of learning correlated patterns. AB - The problem of learning correlated patterns in a neurobiologically plausible way without multiple iterations is discussed. Three guiding principles are outlined for solving this problem in a manner suggestive of how the memory modules of the human brain do it. The first, already known, principle of minimum disturbance is applied quantitatively to minimise the number of learning iterations. The second guiding principle involves a self-induced remedy to undo any damage caused to the stabilities of the old stored patterns after a new one has been learnt. The third involves localising connectivities between neurons depending on the structure of the input patterns. A neurobiologically plausible network and a learning rule are constructed based on these principles. Satisfactory use of this network in learning English words as sequences of their letters is demonstrated by means of computer simulation. PMID- 12125889 TI - Learning generative models of natural images. AB - This work proposes an unsupervised learning process for analysis of natural images. The derivation is based on a generative model, a stochastic coin-flip process directly operating on many disjoint multivariate Gaussian distributions. Following the maximal likelihood principle and using the Potts encoding, the goodness-of-fit of the generative model to tremendous patches randomly sampled from natural images is quantitatively expressed by an objective function subject to a set of constraints. By further combination of the objective function and the minimal wiring criterion, we achieve a mixed integer and linear programming. A hybrid of the mean field annealing and the gradient descent method is applied to the mathematical framework and produces three sets of interactive dynamics for the learning process. Numerical simulations show that the learning process is effective for extraction of orientation, localization and bandpass features and the generative model can make an ensemble of a sparse code for natural images. PMID- 12125890 TI - Optimal design of regularization term and regularization parameter by subspace information criterion. AB - The problem of designing the regularization term and regularization parameter for linear regression models is discussed. Previously, we derived an approximation to the generalization error called the subspace information criterion (SIC), which is an unbiased estimator of the generalization error with finite samples under certain conditions. In this paper, we apply SIC to regularization learning and use it for: (a) choosing the optimal regularization term and regularization parameter from the given candidates; (b) obtaining the closed form of the optimal regularization parameter for a fixed regularization term. The effectiveness of SIC is demonstrated through computer simulations with artificial and real data. PMID- 12125892 TI - Transformations of sigma-pi nets: obtaining reflected functions by reflecting weight matrices. AB - This paper presents a methodology that reflected functions by reflecting the weight matrices of an artificial neural network. One of the major problems with the connectionist approach is that trained neural networks can only associate fixed sets of input-output mappings. We provide a methodology which allows the post-trained net to associate different input-output mappings. The different mappings are reflected in a horizontal axis, reflected in a vertical axis and scaling of the initial mapping. The methodology does not train the net on the different mappings but it transforms the weight matrix of the neural network. This paper describes a novel way of utilising sigma-pi neural networks. Our new methodology manipulates sigma-pi unit's weight matrices which transform the unit's output. The weights are cast in a matrix formulation, and then transformations can be performed on the weight matrix of the sigma-pi net. To test the new methodology, the following three steps were carried out on a neural network: (1) the network was trained to perform a mapping function, f; (2) the weights of the network were transformed; and (3) the network was tested to evaluate whether it performs the reflection in the vertical axis,f(ref-vert)(x) = a - f(x). This reflects the function in one dimension. A reflection transformation was used to manipulate the network's weight matrices to obtain a reflection in the vertical axis. Note that the network was not trained to perform the reflection in the vertical axis. The transformation of the weight matrix transformed the function the output performs. This article explains the theory which enables us to perform transformations of sigma-pi networks and obtain reflections of the output by reflecting the weight matrices. These transforms empower the network to perform related mapping tasks once one mapping task has been learnt. This article explains how each transformation is performed and it considers whether a set of 'standard' transformations can indeed be derived. PMID- 12125891 TI - Parameter setting of the Hopfield network applied to TSP. AB - The major drawbacks of the continuous Hopfield network (CHN) model when it is used to solve some combinatorial problems, for instance, the traveling salesman problem (TSP), are the non feasibility of the obtained solutions and the trial and-error setting values process of the model parameters. In this paper, both drawbacks are avoided by introducing a set of analytical conditions guaranteeing that any equilibrium point of the CHN characterizes a tour for the TSP. In this way, any instance of the TSP can be solved with this parameter setting. Some computational experiences are also included, allowing the solution of instances with sizes of up to 1000 cities. PMID- 12125893 TI - On the capabilities of neural networks using limited precision weights. AB - This paper analyzes some aspects of the computational power of neural networks using integer weights in a very restricted range. Using limited range integer values opens the road for efficient VLSI implementations because: (i) a limited range for the weights can be translated into reduced storage requirements and (ii) integer computation can be implemented in a more efficient way than the floating point one. The paper concentrates on classification problems and shows that, if the weights are restricted in a drastic way (both range and precision), the existence of a solution is not to be taken for granted anymore. The paper presents an existence result which relates the difficulty of the problem as characterized by the minimum distance between patterns of different classes to the weight range necessary to ensure that a solution exists. This result allows us to calculate a weight range for a given category of problems and be confident that the network has the capability to solve the given problems with integer weights in that range. Worst-case lower bounds are given for the number of entropy bits and weights necessary to solve a given problem. Various practical issues such as the relationship between the information entropy bits and storage bits are also discussed. The approach presented here uses a worst-case analysis. Therefore, the approach tends to overestimate the values obtained for the weight range, the number of bits and the number of weights. The paper also presents some statistical considerations that can be used to give up the absolute confidence of a successful training in exchange for values more appropriate for practical use. The approach presented is also discussed in the context of the VC-complexity. PMID- 12125894 TI - Exponential stability of Cohen-Grossberg neural networks. AB - Exponential stabilities of the Cohen-Grossberg neural network with and without delays are analyzed. By Liapunov functions/functionals, sufficient conditions are obtained for general exponential stability, while by using a comparison result from the theory of monotone dynamical systems, componentwise exponential stability is also discussed. All results are established without assuming any symmetry of the connection matrix, and the differentiability and monotonicity of the activation functions. PMID- 12125896 TI - A deterministic annealing algorithm for approximating a solution of the max bisection problem. AB - The max-bisection problem is an NP-hard combinatorial optimization problem. In this paper an equivalent linearly constrained continuous optimization problem is formulated and a deterministic annealing algorithm is proposed for approximating its solution. The algorithm is derived from the introduction of a square-root barrier function, where the barrier parameter behaves as temperature in an annealing procedure and decreases from a sufficiently large positive number to 0. The algorithm searches for a better solution in a feasible descent direction, which has a desired property that lower and upper bounds on variables are always satisfied automatically if the step length is a number between 0 and 1. We prove that the algorithm converges to at least an integral local minimum point of the continuous problem if a local minimum point of the barrier problem is generated for a sequence of descending values of the barrier parameter with zero limit. Numerical results show that the algorithm is much faster than one of the best existing approximation algorithms while they produce more or less the same quality solution. PMID- 12125895 TI - A dynamically coupled neural oscillator network for image segmentation. AB - We propose a dynamically coupled neural oscillator network for image segmentation. Instead of pair-wise coupling, an ensemble of oscillators coupled in a local region is used for grouping. We introduce a set of neighborhoods to generate dynamical coupling structures associated with a specific oscillator. Based on the proximity and similarity principles, two grouping rules are proposed to explicitly consider the distinct cases of whether an oscillator is inside a homogeneous image region or near a boundary between different regions. The use of dynamical coupling makes our segmentation network robust to noise on an image, and unlike image processing algorithms no iterative operation is needed for noise removal. For fast computation, a segmentation algorithm is abstracted from the underlying oscillatory dynamics, and has been applied to synthetic and real images. Simulation results demonstrate the effectiveness of our oscillator network in image segmentation. PMID- 12125897 TI - AANN: an alternative to GMM for pattern recognition. AB - The objective in any pattern recognition problem is to capture the characteristics common to each class from feature vectors of the training data. While Gaussian mixture models appear to be general enough to characterize the distribution of the given data, the model is constrained by the fact that the shape of the components of the distribution is assumed to be Gaussian, and the number of mixtures are fixed a priori. In this context, we investigate the potential of non-linear models such as autoassociative neural network (AANN) models, which perform identity mapping of the input space. We show that the training error surface realized by the neural network model in the feature space is useful to study the characteristics of the distribution of the input data. We also propose a method of obtaining an error surface to match the distribution of the given data. The distribution capturing ability of AANN models is illustrated in the context of speaker verification. PMID- 12125899 TI - Evidence-based practice and heat loss prevention in trauma patients. AB - This article describes a quality management process used to develop an evidence based protocol for heat loss prevention in trauma patients. The quality management tool applied was the ADDIE process (Assess the problem, Determine the root cause, Design a solution, Implement the solution, and Evaluate the outcome). Use of this process required multiple applications and resulted in the identification of a multifactorial root cause. Ongoing changes and additions to the action plans were implemented. PMID- 12125898 TI - An interdisciplinary approach to addressing patient activity and mobility in the medical-surgical patient. AB - Patient functional activity and mobility are essential to recovery and minimization of the risks associated with immobility in hospitalized patients. In practice, there is inconsistency in attending to this aspect of patient care and limited information in the literature to guide clinicians caring for medical surgical patients. An interdisciplinary quality improvement team of nurses, physical therapists, occupational therapists, and patient care assistants developed a programmatic approach to the assessment, planning, and implementation of patient activity criteria in this patient population. Increased awareness and application of the patient activity criteria have improved the consistency with which patient activity is addressed and reduced the incidence of immobility associated complications. PMID- 12125900 TI - "The less-than-perfect medication system": a systems approach to improvement. AB - The 1999 Institute of Medicine (IOM) report increased the focus by health care providers, regulators, and the public on the cause and effect of medical errors. The IOM report recognizes the complexity of the problem of medical errors and advocates a systematic approach to error reduction. Medication delivery systems in health care facilities are an excellent example of the complexity that exists. General Systems Theory (GST) provides a framework to evaluate system design and effectiveness. This article presents an example of the use of GST in the design and evaluation of medication systems with a focus on error reduction. PMID- 12125901 TI - The measurement of patient satisfaction. AB - Patient satisfaction is an important quality outcome indicator of health care in the hospital setting. The measurement of patients' satisfaction with nursing is particularly important since nursing service is often a primary determinant of overall satisfaction during a hospital stay. This article reports on a study designed to update and revise the definition of patient satisfaction for application with ambulatory surgical patients and to develop a questionnaire that captures this definition. The Patient Satisfaction Scale, which specifically focuses on patient satisfaction with nursing care and is used extensively by nursing researchers, was selected for factor analytical examination. Psychometric testing resulted in a 15-item scale with three underlying dimensions. PMID- 12125902 TI - Work site change and psychosocial well-being among health care personnel in geriatric wards--effects of an intervention program. AB - The study evaluated the effect of a change of work site and organization on work environment and psychosocial parameters: the change involved health care personnel at a geriatric hospital. Another aim of this study was to evaluate the effects of a structured psychoeducational intervention program. The study found few changes in the indices of interest on the experimental and control wards. There were, however, significant improvements in social climate, goal quality, and independence of work on the control ward. The investigators postulated that too much external support hampers a group's ability to actively cope with change and might actually lower a group's ability and self-esteem. In order to achieve successful organizational change, psychosocial intervention programs for personnel must be performed by a well-informed, well-chosen, and experienced counsellor who is well tailored to the local organization. PMID- 12125903 TI - Robbing Peter to pay Paul: breaking the RN "recruitment cycle". AB - As health care organizations struggle to fill their share of core vacant positions from a dwindling supply of registered nurses, there has been a focus on recruitment of nurses like never before. Investment in strategies to retain valuable and scarce nurse resources is necessary in order to create a sense of stability on the acute patient care unit (an attractive, but now seldom encountered, hospital environment characteristic). Once an atmosphere of work environment stability is established, nurses can be liberated and moved to focus on the meaning and value of the profession to the community, the nation, and the world. PMID- 12125904 TI - Family planning patterns and acculturation level of high-risk, pregnant Hispanic women. AB - This article describes acculturation level and family planning patterns among a convenience sample of 100 Hispanic women experiencing high-risk pregnancies. The majority were having a second pregnancy and had complications secondary to gestational diabetes or pregnancy-induced hypertension. Most were from Mexico, married, had little formal education, and were very Mexican oriented in their beliefs and values. Almost 75 percent returned for a postpartum visit compared with 14 percent who returned for the family planning visit at one year after giving birth. Age and gravidity were inversely correlated with return for family planning visits: gravidity was a significant predictor of number of post-birth visits. PMID- 12125905 TI - Patient satisfaction and nurses' perceptions of quality in an inpatient cardiology population. AB - Cardiology patients have important learning needs. A 21-item patient satisfaction questionnaire was mailed to 384 cardiology patients 1 week after discharge. Satisfaction ratings indicated that the 161 respondents were satisfied with their care: however, they wanted more information regarding the management of their symptoms and activity level at home. Nurses' interviews revealed that they assessed the availability of follow-up care upon discharge. The results suggest that patients are not satisfied with the information they receive before discharge. Also, nurses and patients have different perceptions about the information patients need. These differences need to be taken into consideration when designing discharge teaching interventions. PMID- 12125906 TI - A case of impairment of mitochondrial fatty acid beta-oxidation. AB - We describe a patient with impairment of mitochondrial fatty acid P-oxidation. A Japanese baby boy was delivered in the 38th week of gestation by emergency cesarean section due to fetal asphyxia. His birth weight was 1,985 g (<10th percentile), length 44.8 cm (<10th percentile), and head circumference 31.0 cm (10th percentile). His Apgar scores were 3 and 5 at 1 min and 5 min, respectively. Blood glucose was 12 mg/dl at 1 hour after birth, requiring glucose administration. On day 1 his serum CK was 20,780 IU/l, which was thought to be due to asphyxia. His serum CK levels gradually began to decrease. At 3 months of age, he sucked poorly, had poor body weight gain, and muscle hypotonia was observed. On day 117 his general condition was impaired, and marked hepatomegaly was observed. The blood glucose level was 43 mg/dl. The patient's urine was negative for ketone bodies. His serum triglyceride level was 3,670 mg/dl. Abdominal CT scan revealed a fatty liver. Serum levels of acyl carnitine from very-long chain fatty acid increased. On day 118 he died due to ventricular fibrillation. On necropsy, massive lipid deposition was observed in the liver, cardiac muscle, kidney, skeletal muscle, and intestinal mucosa. The ratio of very long chain acyl-CoA dehydrogenase (VLCAD) activity for C16/C8 fatty acid was 0.50 (normal control 1.29), suggesting abnormal VLCAD. He was diagnosed as having impairment of mitochondrial fatty acid beta-oxidation, presumably due to the VLCAD deficiency. PMID- 12125907 TI - Tyrosine phosphorylation in cell signaling and disease. PMID- 12125908 TI - Mechanisms of morphogenetic cell assembly. PMID- 12125909 TI - The 22q11.2 deletion syndrome. AB - The 22q11.2 deletion syndrome (22q11DS) encompasses DiGeorge syndrome, velo cardio-facial syndrome and conotruncal anomaly face syndrome and is due to a microdeletion of chromosome 22q11.2. This is the most frequent known interstitial deletion found in human with an incidence of 1 in 4,000 live births. A large number of clinical findings have been reported in affected patients, including cardiac defects, characteristic facial features, thymic hypoplasia, cleft palate, hypoparathyroidism, learning difficulties and psychiatric disorders. A comprehensive evaluation and follow-up program is necessary for patients with 22q11DS. A striking aspect of the 22q11DS phenotype is its variability, the basis of which remains unclear, and no phenotype-genotype correlation has been made. The structures primarily affected in patients with 22q11DS are derivatives of the embryonic pharyngeal arches and pouches suggesting that haploinsufficiency of the gene(s) on the deleted region, spanning 2-3 Mb, is important in pharyngeal arch/pouch development. Extensive gene searches have been successful in identifying more than 30 genes in the deleted segment. Although standard positional cloning has failed to demonstrate a role for any of these genes in the syndrome, the use of experimental animal models and advanced genome manipulation technologies in mice have been providing an insight into the developmental role of some of these genes, including TBXI. In this review, the clinical features and management of patients with 22q11DS are integrated with our current understanding of the embryological and molecular basis of this syndrome, as presented at the 1235th Meeting of The Keio Medical Society. PMID- 12125910 TI - Splenectomy and ligation of the left gastric vein in schistosomiasis mansoni: the effect on esophageal variceal pressure measured by a non-invasive technique. AB - The treatment of choice, in the Northeast of Brazil, of patients with a history of upper GI bleeding from ruptured esophageal varices (EV) and with hepatosplenomegaly secondary to schistosomiasis (HSS), is splenectomy and left gastric vein ligation (SLGL). However, the effect of this procedure on the EV pressure, the parameter that best correlates to re-bleeding risk, has not yet been evaluated. With the introduction of a minimally invasive technique to measure the EV pressure, it has become possible to assess the effect of this surgery without an increased risk to the patient. SLGL was performed in twenty two patients with a history of HSS and upper GI Bleeding secondary to esophageal varices. The non-invasive endoscopic pneumatic balloon was used to measure the EV pressure before surgery and the results were then compared with measurements made between five and eight days post-operatively. The pre-operative EV pressure ranged from 20.0 mmHg to 28.7 mmHg (mean 24.35 +/- 2.36 mmHg), with no correlation between the pressure and the calibre of the varices. In the post operative period, a significant decrease in EV pressure was observed, ranging from 14.6 mmHg to 21.5 mmHg (mean 17.29 +/- 1.75 mmHg, p < 0.001). These results support the use of SLGL in patients with HSS and a history of variceal bleeding. The operation results in, at least for the short term and in the majority of cases, a reduction in the EV pressure, and therefore a reduced risk of repeating upper GI Bleeding. PMID- 12125911 TI - Protein tyrosine phosphorylation signaling in the differentiation of human endometrial stromal cells. AB - Reversible protein tyrosine phosphorylation, coordinately controlled by protein tyrosine kinases (PTKs) and protein tyrosine phosphatases (PTPs), is a critical element in signal transduction pathways regulating cell growth, differentiation, apoptosis, and tumorigenesis. The differentiation of human endometrial stromal cells (decidualization) is crucial for successful embryo implantation and maintenance of pregnancy; however, little is known about the molecular events involving tyrosine phosphorylation, PTKs, and PTPs in the process of decidualization. We have previously reported that the tyrosine kinase activity of c-Src belonging to the Src family kinase is increased together with altered tyrosine phosphorylation of several cellular proteins in the in vitro model of decidualization. Focal adhesion kinase (FAK) and paxillin are known to form a complex with c-Src at the focal contacts and to participate in the integrin mediated signal transduction as c-Src substrates. Those focal adhesion proteins, however, are not hyperphosphorylated on tyrosine during decidualization. Moreover, the loss of focal adhesions and the disorganization of the actin-based cytoskeleton were observed in decidualized stromal cells, suggesting that the escape from regulation by c-Src may be in part due to the decidualization-induced disruption of the interaction between the focal adhesion proteins and c-Src. These findings collectively indicate that decidual c-Src may activate signaling pathway(s) different from the integrin-mediated signaling cascade involving FAK and paxillin. This review summarizes our recent studies on the tyrosine phosphorylation signaling pathway(s) in decidualization. PMID- 12125912 TI - The prevalence of asthma among secondary school students in Afyon, Turkey. AB - Ethnic origin has been reported to affect the prevalence of atopic diseases in several studies in different parts of the world. The aim of this study was to determine the current and cumulative prevalence of asthma, allergic rhinitis, atopic dermatitis and reactivities to allergen skin prick tests (SPT) among secondary school students in Afyon, Turkey. The data of the first stage were collated through the application of the European Community Respiratory Health Survey Questionnaire on 1,400 students (1,366 were properly completed) registering at various schools in Afyon-Turkey, in the academic year 2000-2001. At the second stage, a physical examination and SPT were performed on 121 students. Of the students within the study group, 53 (3.9%) had experienced an asthma attack within the previous 12 months, 49 (3.6%) had a past history of asthma attacks and 167 (12.2%) reported wheezing attacks within the previous 12 months. The cumulative prevalence of asthma, rhinoconjunctivitis and dermatitis were found to be 7.46%, 8.10% and 3.5%, respectively. A positive SPT reaction to at least one allergen was found in 15.7% of the students. SPT positivity was 11.13% in symptomless students and 20.3% in asthmatic students. In analysis of logistic regression, a history of atopy, as determined in the questionnaire, was seen to be significantly correlated with SPT positivity. The prevalence of self reported asthma and/or asthma-like symptoms was shown to be lower in students living in Afyon than other countries. Asthma and asthma-like symptoms were found to be significantly more prevalent among students who smoked. PMID- 12125913 TI - Estrogen receptor-alpha mRNA in primary breast cancer: relationship to estrogen and progesterone receptor proteins and other prognostic factors. AB - The estrogen receptor (ER)-alpha protein and ER mRNA were measured in 314 primary breast cancer patients by enzyme immunoassay (EIA) and reverse-transcription polymerase chain reaction (RT-PCR) assay, respectively. The positivity of ER protein was 53% while of ER mRNA was 37.6%. A significant positive association between ER phenotype and ER mRNA was observed (r = 0.40, p < 0.0001) with a positive-negative agreement between them of 71.8%. The percentage of ER-negative, progesterone receptor (PR)-positive breast tumors was 1.9% by EIA and 7% by RT PCR assay. This may indicate a difference in ER variants in these studied patients. The ER protein and ER mRNA status were inversely related to tumor size and p53 positivity. Also, ER protein was frequently positive in patients with a higher number of lymph node invasions, well to moderate nuclear differentiated tumor cells and negative c-erbB-2 status. The difference of the ER or ER mRNA status regarding ages, menopausal status, tumor stages and histological types was not shown. In the present study, ER mRNA did not demonstrate a closer relationship to prognostic indicators of breast cancer than ER protein. Before including the ER mRNA assessment in routine investigations of breast cancer, its relationship to prognostic factors and survival outcome should be further assessed with a higher number of patients and a longer follow-up time. PMID- 12125914 TI - The effect of cigarette smoking on ceruloplasmin and C3 complement: risk of cardiovascular disease (atherosclerosis). AB - Serum ceruloplasmin, C3 complement and albumin in 119 male smokers and 65 male non-smoker; from a military unit in Bangkok were investigated in this study. The serum ceruloplasmin concentration was found to be significantly higher in smokers than in non-smokers. However, the serum albumin concentration in smokers was statistically significantly lower than in non-smokers. Significant associations were also found between ages, albumin levels and the quantity of cigarettes smoked. There was a significant positive correlation between serum ceruloplasmin and C3 complement concentrations. An association between the quantity of cigarettes smoked and albumin was also found, as well as a significant relationship between smoking and the quantities of cigarettes smoked to serum ceruloplasmin levels when smoking and the quantity of cigarettes smoked were taken as independent variables, and the serum ceruloplasmin levels as a dependent variable. This might suggest that high concentrations of the acute-phase protein, i.e. ceruloplasmin, might constitute a risk of developing atherosclerosis or cardiovascular disease in smokers. PMID- 12125915 TI - Co-existence of HIV-1 subtypes B' and E infections among Thai injecting drug users. AB - Subtypes B' and E are the two major subtypes of HIV-1 among injecting drug users (IDU) in Thailand. However, there are not many reports on subtype distribution during the early epidemic. Random blood specimens collected during 1994-2000 from 3,286 IDU at the Thanyarak Hospital were tested for HIV antibody and subtyped by using peptide binding enzyme immunoassay. The prevalence rate of HIV infection was 36.8%. All HIV-seropositive IDU were ascertained for "year of first HIV seropositivity" from their medical records. Of 1,512 HIV-seropositive samples, 1,408 (93.1%) were typeable. During 1987-1988, the proportion of subtype B' was as high as 80% but decreased rapidly to 27.6% during 1999-2000. At the same time, the proportions of subtype E increased correspondingly (Chi-square test for the trend, p < 0.05). The relatively high proportion of subtype E among IDU since an early stage of the epidemic suggests early co-existence of both subtypes and needs further investigation. PMID- 12125916 TI - Flow cytometry for the analysis of T cells expressing CD69 after stimulation with glutamic acid decarboxylase. AB - Type 1 diabetes mellitus is a T-cell mediated autoimmune disease in which the insulin-producing pancreatic beta cells are selectively destroyed. We recently found that the detection of cell-mediated immune response to glutamic acid decarboxylase (GAD) was more useful than the detection of specific autoantibodies for the diagnosis of type 1 diabetes mellitus. In this study, we established a flow cytometric analysis for the detection of activated T cells in whole venous blood, obtained from diabetic patients and normal controls after stimulation by GAD. Two millitiers of peripheral venous blood and 6 hours incubation time were used for performing the test. It was found that 33% (3/9) type 1 diabetic patients, 7.7% (1/13) type 2 diabetic patients and neither patients with fibrocalculous pancreatopathy nor normal controls had > or = 20% CD8+ T cells expressing CD69. The results suggest that flow cytometry may be a useful tool for the detection of surrogate markers of type 1 diabetes mellitus. PMID- 12125917 TI - Allergic rhinitis and its impact on asthma: an evidence-based treatment strategy for allergic rhinitis. AB - The overall pathogenic view of respiratory allergy has deeply changed and evolved during the last ten years. Much emphasis has been laid to the relationship between rhinitis and asthma, which is between the upper and the lower respiratory airways. This strict link has been evidenced through clinical observations and epidemiological studies and also on the basis of immunological observations and outcomes of therapy. Furthermore, the frequent co-existence of rhinitis and asthma (up to 80 percent of asthmatic patients have co-existing allergic rhinitis, while up to 40 percent of allergic rhinitis patients have asthma, the coexistence of sinusitis and asthma, the presence of rhinitis as a risk factor for developing asthma, further emphasize this link and together lead to the operative definition of Allergic Rhinobronchitis or, United Airways Disease (UAD). The strict link existing between upper and lower respiratory tract can be also regarded from the viewpoint of therapeutical outcomes. The more detailed knowledge of the intricate mechanisms sustaining allergic inflammation in the respiratory tract (i.e. antigen presentation, cytokines, chemokines and adhesion molecules) has clarified the functional relationships between nose and lung. Thus allergic rhinitis or asthma is not a disease confined to a specific target organ, but rather a disorder of the whole respiratory tract, with a range of clinical manifestations, leading to relevant diagnostic and therapeutic implications as indicated in the WHO Initiative ARIA, the first evidence-based guideline emphasizing the impact of allergic rhinitis on asthma and where a step-wise treatment strategy targeting both the upper and lower airway effectively has been proposed. Moreover, the use of novel potential therapies that target both rhinitis and asthma like antileukotrienes or anti-IgE are indeed a future strategy. PMID- 12125918 TI - Declining hepatitis A seroprevalence among medical students in Bangkok, Thailand, 1981-2001. AB - The severity of clinical symptoms following hepatitis A virus (HAV) infection is age dependent. Hepatitis A in children is mostly an asymptomatic disease while adolescents and adults usually show symptoms of clinical hepatitis. Improved personal hygiene and environmental sanitation has led to a decline in natural immunity acquired in childhood, creating a population of susceptible adults. In the past decade, the incidence and prevalence of hepatitis A disease in Thailand have decreased significantly. In this study, we used enzyme-linked immunosorbent assay to determine the prevalence of anti-HAV antibodies among medical students at two different time points in 1996 and 2001. We then compared these results with data from previous studies in 1981 and 1992. The seroprevalence was 73.01%, 30.23%, 16.67% and 6.67% in 1981, 1992, 1996 and 2001, respectively. A significant decline has happened over the past two decades (p < 0.001). Considering the decreasing immunity to HAV in the younger generations, more cases of symptomatic HAV infection could be anticipated. Further seroprevalence studies in other adolescence groups from different socioeconomic status are needed to elucidate the current situation of HAV infection in the young generation more comprehensively and to develop an appropriate prevention program. PMID- 12125919 TI - Cyclins D1, E, A expression and synergistic cytotoxicity to a cholangiocarcinoma cell line from recombinant TNF-alpha and PHA supernate. AB - We studied the cytotoxic effects of recombinant TNF-alpha and supernate of phytohemagglutinin stimulated peripheral blood mononuclear cells individually and in combination against a cholangiocarcinoma cell line. Levels of cyclins D1, E and A in the cell line were detected by immunoblotting, and the cell cycle stage was assayed by propidium iodide staining followed by flow cytometry analysis. Viable and apoptotic cells were assessed by trypan blue dye exclusion, DAPI staining, agarose DNA laddering and propidium iodide staining. At the beginning of each experiment, the majority of cholangiocarcinoma cells expressed cyclin A and were in S phase as determined by propidium iodide staining. Treatment of such cells with recombinant TNF-alpha resulted in cytotoxic effects clearly evident at 36 hours post exposure. There was a synergistic killing effect when recombinant TNF-alpha was combined with PHA supernate and this effect could be partly neutralized by monoclonal anti TNF-alpha, interleukin (IL)-2, IL-12 and IFN gamma. PMID- 12125920 TI - Stromal cell-derived factor (SDF) 1-3'A polymorphism and sequences in Thais. AB - Stromal cell derived factor (SDF) 1-3'A polymorphism in Thai subjects was determined by restriction fragment length polymorphism (RFLP) of polymerase chain reaction (PCR) products using a restriction enzyme Msp 1 cleavage. The allelic frequency of SDF1-3'A in this population was 0.289. SDF1-3'A genotyping showed 9.52% SDF1-3'A/3'A, 38.89% SDF1-wt/3'A, and 51.59% SDF1-wt/wt. Two clones of Thai SDF genes, BD41 and BD42, were isolated and sequenced. Wild type Thai-SDF1 (BD42), a 278 bp sequence was identically aligned to human pre B cell stimulating factor homoloque (SDF1-b) (GenBank accession number L-36033) from nucleotides 788 to 1,065. Homologous Thai SDF1-3'A (BD41), a 277 bp sequence differed by single nucleotide with adenine substitution to guanine at position 880 of the SDF1-b, is the first evidence of SDF1-3'A polymorphism in Thais. PMID- 12125921 TI - Clinical utility of telangiectasia of hands in scleroderma and other rheumatic disorders. AB - Telangiectasia of the hands were observed in 76% of patients with scleroderma (n = 53) as compared with 12% of patients with other rheumatic disorders (n = 100) and in 13% of healthy subjects (n = 30). In scleroderma, telangiectasia of the hands were commonly multiple (mean number +/- SD = 22.9+/-30.1) with 7.3% being >1 mm in size. They were found in greater numbers in those patients with the limited subtype. The distribution of telangiectasia was observed on all but 4 of 158 sectors of the hand with significant higher numbers on the ventral surface of the digits. Significant associations of telangiectasia of the hands were also observed with numbers of telangiectasia on face and lips (p = 0.001), disease duration (p = 0.002), surface area of digital calcinosis (p = 0.03) and with the presence of the centromere antibody (p = 0.005). Possible associations were observed with prior gastrointestinal bleeds (particularly with telangiectasia >1 mm) and with isolated pulmonary hypertension. No significant correlation was found between number of telangiectasia and with nailfold capillary size or density. In conclusion, multiple telangiectasia of the hands were most commonly observed in patients with the centromere positive, limited subtype of scleroderma of long duration. Their pathogenesis is unknown. PMID- 12125922 TI - Biomineralization of the spicules of sea urchin embryos. AB - The formation of calcareous skeletal elements by various echinoderms, especially sea urchins, offers a splendid opportunity to learn more about some processes involved in the formation of biominerals. The spicules of larvae of euechinoids have been the focus of considerable work, including their developmental origins. The spicules are composed of a single optical crystal of high magnesium calcite and variable amounts of amorphous calcium carbonate. Occluded within the spicule is a proteinaceous matrix, most of which is soluble; this matrix constitutes about 0.1% of the mass. The spicules are also enclosed by an extracellular matrix and are almost completely surrounded by cytoplasmic cords. The spicules are deposited by primary mesenchyme cells (PMCs), which accumulate calcium and secrete calcium carbonate. A number of proteins specific, or highly enriched, in PMCs, have been cloned and studied. Recent work supports the hypothesis that proteins found in the extracellular matrix of the spicule are important for biomineralization. PMID- 12125924 TI - In vitro cultivation of cells from ovotestis tissue of pigmented Biomphalaria glabrata. AB - Cells derived from ovotestis tissue of pigmented Biomphalaria glabrata, Puerto Rican strain were cultured in double diluted GIT medium supplemented with modification of amino acids components of pigmented B. glabrata, ovotestis and mid-gut region and 3% inactivated fetal calf serum. As a result, two types of cells, epithelial and fibroblastic like cells increased in number during the cultivation. It seem that the medium used in this study is a suitable medium for cultivation of cells from ovotestis of pigemeted B. glabrata. These two types of cells have been maintained by successive transplantation for over 3 passages. PMID- 12125923 TI - Mutual and directional allogeneic cytotoxic reaction of hemocytes in the solitary ascidian Halocynthia roretzi revealed by one-step quantitative fluorimetric assay. AB - A novel one-step microplate cytotoxicity assay using the cytoplasmic fluorescent viability dye calcein AM was established for simple, rapid, sensitive, and quantitative measurements of the allogeneic cytotoxic reaction (ACR) mediated by hemocytes in the ascidian Halocynthia roretzi. The mutual and directional ACR was distinguishable by the assay using the hemocytes from pairs of animals with different alloreactivities. The ACR assay may allow more precise genetic analysis of the gene that controls alloreactivity of hemocytes, since the mutual and directional ACR may be related to levels of expression or numbers of the gene product or products on the target cells. The directional ACR will be useful in elucidating the cellular and molecular mechanisms of self-recognition in H. roretzi, since it allowed us to equate hemocytes from one animal with "effector cells" and those from the other animal of the pair with "target cells". In addition, the quantitative ACR assay in a large number of samples is possible and it will allow production of monoclonal antibodies that may recognize receptors or ligands functioning in self-recognition processes by the H. roretzi hemocytes. PMID- 12125925 TI - Biochemical and immunohistochemical studies on tropomyosin and glutamate dehydrogenase in the chicken liver. AB - Recently, we have reported a novel tropomyosin (TM) -binding protein, glutamate dehydrogenase (GDH) and demonstrated by affinity column chromatography that chicken liver TM interacts with GDH in an ATP-dependent manner. To elucidate the physiological roles of the interaction between TM and GDH, we performed co sedimentation assays of TM and GDH with F-actin, because it is known that TM exerts its physiological functions by associating with actin filaments. The results showed that TM and GDH co-pelleted with F-actin. GDH alone also co precipitated with F-actin, but the amount of GDH sedimenting with F-actin was increased in the presence of chicken liver TM, suggesting that GDH is involved in the regulation of the actin cytoskeleton. We also prepared crude GDH from the nuclear and mitochondrial fractions obtained by subcellular fractionation of the chicken liver cells. Semi-nondenaturing 2D-PAGE revealed that partially purified GDH from the nuclear fraction was associated with TM, but not GDH from the mitochondrial fraction, suggesting preferential binding of TM to GDH. We determined the nucleotide sequence of chicken GDH cDNA and showed that the GDH transcript was widely expressed in the chicken organs. We examined the localization of TM and GDH by immunohistochemistry and revealed that they were distributed in the cytoplasm of the adult chicken liver. From these results, we propose two hypotheses on the physiological roles of the interaction between TM and GDH in nonmuscle cells. PMID- 12125926 TI - Molecular characterization of a cDNA from the silk moth Bombyx mori encoding Manduca sexta allatotropin peptide. AB - Allatotropin is a 13-residue amidated neuropeptide isolated from pharate adult heads of the tobacco hornworm, Manduca sexta and strongly stimulates biosynthesis of juvenile hormones in adults, but not larval, lepidopteran corpora allata. From a Bombyx mori midgut cDNA library, a cDNA that encodes a 130-amino-acid polypeptide containing M. sexta allatotropin sequence was isolated. The B. mori allatotropin cDNA consists of 1196 nucleotides. The encoded allatotropin peptide is identical to that isolated from M. sexta and that predicted from Pseudaletia unipuncta, with 84% and 81% identity in the amino acid sequence of the allatotropin peptide precursor, respectively. M. sexta allatotropin is flanked by two different endoproteolytic cleavage sites within the precursor of the B. mori allatotropin peptide. Evidence from northern blotting of B. mori tissues showed that the allatotropin gene is expressed in the cells of midgut, head and integument with different transcription amount, but not in the fat body and silk gland. Midgut has also a number of allatotropin-immunoreactive cells and nerve fibers. These results will provide valuable information in understanding the AT gene of insects. PMID- 12125927 TI - Molecular cloning of a putative gastric chitinase in the toad Bufo japonicus. AB - On the basis of our preliminary observation that a crude extract of the stomach of the toad Bufo japonicus exhibited a chitinase activity with its optimum pH around 3.0, we undertook molecular cloning of a cDNA encoding this putative gastric chitinase. By use of 2 degenerate oligonucleotide primers derived from the 2 conserved regions of the vertebrate chitinases, a reverse transcription-PCR product was obtained. This product was used as a probe to screen a cDNA library constructed from the toad stomach. The longest positive clone was revealed to contain an open reading frame for a putative chitinase protein of 484 amino acids, which protein exhibited sequence similarity to the known vertebrate chitinases. Our data also revealed this putative gastric chitinase to be distinct from the chitinase that we had previously isolated from the pancreas of the same species. In this putative gastric chitinase, both the N-terminal catalytic domain and the C-terminal chitin-binding domain were perfectly conserved, suggesting this protein to function as chitinase in the toad stomach. PMID- 12125928 TI - Role of cell contact in the specification process of pigment founder cells in the sea urchin embryo. AB - Effects of LiCl on the specification process of pigment founder cells were examined in the sea urchin Hemicentrotus pulcherrimus. If embryos were treated with 30 mM LiCl during 4-7 or 7-10 hours postfertilization, pigment cells increased significantly. Aphidicolin treatment indicated that this increase was due to the increase in the pigment founder cells. Interestingly, if the embryos were treated sequentially with LiCl and Ca2+-free seawater during 4-7 and 7-10 hr, respectively, they differentiated only about the same number of pigment cells as control embryos. Further, the increase was scarcely discerned when the embryos were treated with LiCl in the absence of Ca2+ during 7-10 hr. These results suggested that effect of LiCl would be ascribed to the increase in cell adhesiveness. In fact, LiCl-treated embryos were more difficult to be dissociated into single cells. Cell electrophoresis showed that the amount of the negative cell surface charges decreased considerably in LiCl-treated embryos. It was also found that the number of pigment cells seldom exceeded 100, even if embryos were exposed to a higher concentration of LiCl. This suggested that only a subpopulation of the descendants of veg2 blastomeres received the inductive signal emanated from the micromere progeny. PMID- 12125929 TI - Aryl hydrocarbon receptor is required for prevention of blood clotting and for the development of vasculature and bone in the embryos of medaka fish, Oryzias latipes. AB - The aryl hydrocarbon receptor (AHR) is a member of ligand-activated transcription factors and conserved among vertebrates. To investigate the role of AHR in fish development, medaka embryos were treated with agonist (2,3,7,8-tetrachlorodibenzo p-dioxin), antagonists (alpha-naphthoflavone and resveratrol), and inhibitor (piperonyl butoxide) of cytochromes (Cyts) P450 encoded by a battery of target genes. These embryos were found to have similar abnormal phenotypes. Among the most consistent phenotypes were blood clotting and malformation of bone that were associated with vascular damages. These results thus indicate that control of AHR is important for proper development of fish embryos. AHR may control levels of Cyts P450 that are responsible for synthesis and metabolism of a toxic compound that caused the abnormal phenotypes. Complementary DNA fragments encoding AHR homologs were cloned from medaka embryos. AHR-specific mRNA was ubiquitously expressed in embryos and adult tissues. PMID- 12125930 TI - Comparative studies between in vivo and in vitro spermatogenesis of Japanese eel (Anguilla japonica). AB - In order to check the quality of in vitro spermatogenesis of Japanese eel, in vitrol 1-ketotestosterone (11-KT) induced spermatogenesis was compared with in vivo spermatogenesis induced by a single injection of human chorionic gonadotropin (hCG) in detail. DNA contents of germ cells from in vitro and in vivo testicular fragments were compared using flow cytometry. Since the in vitro result of flow cytometry showed prominent 1C peak including spermatozoa and spermatids, the reduction of DNA by meiosis was assumed to progress normally, (i.e., haploid spermatozoa were produced in this in vitro system). In the testes of in vitro culture, however, spermatozoa were not released into lumen. Furthermore, the number of mitotic divisions of the in vitro experiment (6 divisions) was fewer than that of in vivo (10 divisions). In electron microscopy observations, both of in vivo and in vitro spermatozoon had a crescent-shaped nucleus with a flagellum, and a single large spherical mitochondrion. However, the elongation of the sperm head was not sufficient and the mitochondrion was not always located at the anterior end as is observed for the spermatozoa obtained from hCG injected eels. Eel spermatogenesis related substance-11 (eSRS11) is homologue of histone H1 which is up-regulated during spermatogenesis. Using this probe, in vitro spermatogenesis was also evaluated in molecular levels. In Northern blot analysis, eSRS11 mRNA was detected in both in vivo and in vitro testes. However, the expression of in vitro was much weaker than that of in vivo. These differences indicate that the stimulation of 11-KT is not sufficient, and another factors are needed to induce complete spermatogenesis in vitro. PMID- 12125932 TI - Fluctuation in abundance and age structure of a breeding population of the Japanese brown frog, Rana japonica gunther (Amphibia, anura). AB - A breeding population of Rana japonica was studied at a marsh on the campus of Hiroshima University in Higashi-Hiroshima during the five years 1995-1999. The mark-recapture study showed that the size of the breeding population varied from year to year, and increased more than twofold in 1999 in comparison with the preceding years. The sex ratio of the breeding population (male/female) was from nearly 1.0 to 1.6. Frogs of both sexes were estimated to breed for the first time at the age of one or two years, and their maximum age was four years according to skeletochronology using phalanges and mark-recapture. Modes of the estimated ages were one year for males during the study years except 1997, but one or two years for females. Two thirds of breeding frogs, irrespective of their sex, were estimated to breed only once throughout their lives. PMID- 12125931 TI - Effects of starvation on gonadotropin and thyrotropin subunit mRNA levels and plasma hormone levels in the male Japanese quail (Coturnix coturnix japonica). AB - Contents of mRNAs encoding LHbeta-, FSHbeta-, TSHbeta- and common a-subunit precursor molecules were measured in male Japanese quail deprived of food for three days. Plasma LH, FSH, thyroxine and triiodothyronine levels were also measured in the same birds. Plasma LH levels declined during the period of food deprivation. Levels in starved birds were not different from those in control birds after one day of starvation but were significantly lower after three days. Plasma FSH levels showed a similar decline, although the changes were not significant. Plasma thyroxine levels did not decrease during starvation, whilst plasma triiodothyronine levels decreased drastically and significantly soon after the start of starvation. All the hormone subunit mRNA contents in starved birds also decreased, with differences from control birds significant 3 days after the start of starvation. Plasma FSH levels showed a strong positive correlation with pituitary FSHbeta mRNA levels, while plasma LH levels had a strong positive correlation with common a mRNA levels and practically no correlation or even a negative correlation with LHbeta mRNA levels. These results suggest that starvation suppresses not only gonadotropin and thyrotropin secretion but also their synthesis in the pituitary gland. Furthermore, these results showed that FSH and LH have different synthesis and secretion dynamics in the Japanese quail. Contradicting results with TSHbeta mRNA and thyroid hormones lead us to assume that starvation affects thyroid hormone metabolism in peripheral tissue, presumably in the liver. PMID- 12125933 TI - A quantitative analysis of geographic color variation in two Geotrupes dung beetles. AB - We conducted a quantitative analysis of geographic color variation in two species of dung beetles: Geotrupes auratus and G. laevistriatus. The reflectance of the dorsal surfaces was measured from 300 to 700 nm using a spectrophotometer. The reflectance curves for both beetles were bimodal; there were two distinct peaks, namely, the a peak, between 400 and 700 nm, and the beta peak at around 300 nm. A stepwise discriminant analysis indicated that geographic color variation in Geotrupes beetles was primarily characterized by a shift of the a peak. Using beetles from three locations, we compared the wavelength (nm) of the a peak (lambdamax(alpha)) and its reflectance intensity (R(a)) to investigate sex and population differences. Intraspecific geographic variation in coloration was effectively detected by discriminant analysis of spectral reflectance curves. Our results showed that G. auratus and G. laevistriatus had similar coloration within each sampling location. Our study also revealed hidden sex differences in R(a); R(a) of males were significantly higher than those of females in both species. Since the dorsal surface of the beetles shows remarkable color variation, and coloration can be assessed objectively using reflectance spectra, Geotrupes beetles may be good model organisms to investigate geographic color variation. PMID- 12125934 TI - Loxocorone, a new genus of the family loxosomatidae (Entoprocta: Solitaria), with descriptions of two new Loxomitra (sensu stricto) and a new Loxocorone from Okinawa, the Ryukyu Archipelago, Japan. AB - The type species of Loxomitra Nielsen, 1964, L. kefersteinii (Claparede, 1867), has following characters: A) the buds have a pair of terminal wings at the base of the stalk, and B) the liberated buds crawl on the substratum by attaching the two terminal wings to the substratum and twisting the whole body. By contrast, some other members of the genus have following characters: A') the buds lack the terminal wings but have a foot with foot groove, and B') the liberated buds glide over the substratum using the foot with foot groove. I thus propose to divide Loxomitra (sensu lato) into two genera- Loxomitra (sensu stricto) characterized by A) and B), and Loxocorone gen. nov. by A') and B'). I also describe two new Loxomitra (sensu stricto), L. mizugamaensis sp. nov., and L. tetraorganon sp. nov., and one new Loxocorone, L. allax sp. nov. from the Ryukyu Archipelago, Japan. All of the currently recognized species of the Loxomitra (sensu lato) are reviewed to specify their generic allocations in response to the above change. PMID- 12125935 TI - Genetic relationships among Korean brown frog species (Anura, Ranidae), with special reference to evolutionary divergences between two allied species Rana dybowskii and R. huanrenensis. AB - Allozyme analysis for 41 populations of brown frog species, Rana dybowskii, R. huanrenensis, and R. amurensis from Korea and three reference species (Chinese R. chensinensis and Japanese R. dybowskii and R. tsushimensis), were performed to clarify taxonomic status of Korean brown frogs. The level of average genetic differentiation (Nei's D) among local populations of each species in Korea was very low (D<0.01 2) and Korean and Japanese R. dybowskii also showed conspecific level of differentiation (D=0.070). Whereas, much larger, discrete genetic differences were detected in the interspecific comparisons (D>0.370). In the genetic relationships among five species examined, the 24 chromosome brown frogs (R. dybowskii, R. huanrenensis, and R. chensinensis) did not form a monophyletic group. Rana dybowskii with the chromosome number of 2n=24 was grouped together with R. amurensis with the chromosome number of 2n=26. The hypothesis of reversal change from 24 to 26 in Korean R. amurensis seems to better explain the phylogenetic relationships of east Asian brown frogs than the assumption of parallel reduction in chromosome number from 2n=26 to 24 in R. dybowskii and in the common ancestor of R. huanrenensis and R. chensinensis. The genetic, morphological, and reproductive divergences between Korean R. dybowskii and R. huanrenensis were compared. PMID- 12125936 TI - A community-based epidemiological study of acne vulgaris in Hong Kong adolescents. AB - Using a questionnaire survey, the prevalence and severity of acne were assessed in a randomized sample of 522 persons (aged 15-25 years) out of 5,522 telephone interviews in Hong Kong. The prevalence of self-reported acne was 91.3%. At the time of interview, 52.2% had acne. More acne scars and pigmentation were present (52.6%) than in a Western population; 26.6% were disturbed psychologically by acne and 82.9% by its physical appearance. Only 2.4% had sought the advice of a doctor for managing acne, while 41.5% had tried some form of medical treatment. Topical treatment comprised 94.7% of medications used for acne. The results show that acne and its complications are common problems. The treatment of acne scars and pigmentation is difficult and complicated by Asian skin phototypes. The findings suggest the need for refined educational programmes to ensure that adolescents know what effective treatments are available so that complications can be reduced. PMID- 12125937 TI - Quality of life and prevalence of arthritis reported by 5,795 members of the Nordic Psoriasis Associations. Data from the Nordic Quality of Life Study. AB - Quality of life measures are widely used in dermatology as well as in rheumatology, but there are no large studies taking arthritis into consideration when studying quality of life in psoriasis. The aim of this study was to investigate psoriasis-related quality of life in a large sample of members of the psoriasis associations from the Nordic countries including an arthritis-related evaluation. The prevalence of reported arthritis within the groups was also estimated. An Arthritis Disability Index suitable for parallel use together with Finlay's Psoriasis Disability Index was constructed. A total of 5,795 members and 702 patients seen by Nordic dermatologists rated the severity of their disease and completed the Psoriasis Disability Index formula and a Psoriasis Life Stress Inventory, and if arthritis had been diagnosed, the Arthritis Disability Index formula. Approximately 30% of all psoriatic patients, irrespective of group, received a diagnosis of arthritis either by their dermatologist or a rheumatologist. Members previously hospitalized for their disease had a higher frequency of arthritis (41%) than those without a history of hospitalization (23%). The highest prevalence of arthritis was found in Norway (33.8%). Members with arthritis exhibited greater impairment of psoriasis-related quality of life, longer disease duration, and greater self-reported disease severity for psoriasis. Important predictors for impairment of arthritis-related quality of life were pain, number of affected joints, and restriction of joint mobility. These data show, that the prevalence of arthritis in psoriasis may be significantly higher than the previously accepted average of 7%. The results demonstrate that when studying quality of life in psoriasis, arthritis and arthralgia are important independent factors to be included in the evaluation. PMID- 12125938 TI - Insulin-induced drug eruptions and reliability of skin tests. AB - Allergic reaction to insulin preparations seemed to have decreased since the introduction of contaminant-free, human preparations. The role of protamine sulfate in decreasing the prevalence of allergy is unclear. This study examines the causative components of insulin allergy along with the value of skin tests for diagnosis. Eleven patients with insulin allergy and 53 patients receiving insulin but without an insulin allergy were included as controls. Intradermal skin tests were conducted using preparations containing various concentrations of insulin [Neutral protamine Hagedorn (NPH) insulin, regular insulin (RI)] and protamine sulfate. Of the 11 patients studied, 3 had anaphylaxis and 8 displayed localized reactions. All of the patients reacted positively during skin testing. Five patients showed positive intradermal skin test reactions to protamine sulfate, and 4 reacted to insulin. Two patients that were not tested with protamine sulfate reacted positively to NPH insulin. In the case of protamine sulfate, 4 patients with localized symptoms displayed positive reactions at concentrations of 10 microg/ml, 3 microg/ml or 0.3 microg/ml. One patient with anaphylaxis reacted positively to a concentration as low as 0.03 ng/ml. In the case of insulin protein, 3 patients reacted positively to a 100-fold dilution (1 UI/ml). Eight of the 53 controls experienced pruritus and/or skin lesions. However, none of the controls reacted at a concentration of NPH insulin of less than 10 U/ml or to protamine sulfate at less than 30 microg/ml. Allergic reactions to protamine sulfate are common and should not be ignored. This study shows a good correlation between clinical manifestations and skin test reactions for insulin allergy. PMID- 12125939 TI - Proportion of herpes simplex virus (HSV) type 1 and type 2 among genital and extragenital HSV isolates. AB - Herpes simplex virus type 1 (HSV-1) has been associated with orofacial infections and HSV type 2 (HSV-2) with genital infections. This tropism of the virus seems to have changed and in clinical reports an increasing number of genital herpes infections caused by HSV-1 have been recognized. The aim of this study was to estimate the proportion of HSV-1 and HSV-2, respectively, among isolates from different anatomical sites typed in our laboratory during the years 1994-1998. Out of a total of 3,085 anogenital isolates, 29% were typed as HSV-1 and 71% as HSV-2. The highest prevalence of HSV-1 was registered among isolates from young women. Of 631 orofacial isolates, 4% were typed as HSV-2 and 96% as HSV-1. Of 69 finger/hand isolates, 54% were typed as HSV-1 and 46% as HSV-2, and of 95 isolates from other regions (abdomen, foot, etc.), 60% were typed as HSV-1 and 40% as HSV-2. It was found that HSV-2 was as common as HSV-1 in the extra-genital regions with the exception of the orofacial area, in which HSV-2 was seldom detected. Furthermore, the study showed an increasing proportion of HSV-1 among anogenital isolates during the study period. Taken together, these results suggest that a clear HSV type-related tropism might be limited to the permissiveness of the orofacial region for HSV-1, and that both serotypes may readily establish infections below the neck. PMID- 12125940 TI - Acute allergic reactions to Anisakis simplex after ingestion of anchovies. AB - Anisakis simplex is a seafish nematode, which is responsible for the well-known human infection (anisakiasis) and can induce IgE-mediated reactions. IgE sensitization to Anisakis simplex can be frequent in particular countries and should be suspected in patients with acute allergic symptoms after ingestion of fish. The etiological role of Anisakis simplex was evaluated in 49 adult subjects with acute allergic symptoms after ingestion of anchovies. Serum-specific IgE and prick tests to anchovy were negative in each patient. Specific IgE reactions to Anisakis were positive in 45 patients and skin tests in 43. Only 3 patients with allergy to the nematode were atopic. However, IgE responses to Anisakis were also observed in habitual consumers of raw fish, without any clinical manifestations, suggesting that the relevance of results of conventional tests has to be interpreted on the basis of clinical aspects. PMID- 12125941 TI - Benign neonatal hemangiomatosis with conjunctival involvement. Report of a case and review of the literature. AB - Hemangiomas are the most common tumours of infancy. When limited to the skin, multiple lesions have a benign course and excellent prognosis but in cases of visceral involvement, the morbidity and mortality rates are high. We report a rare case of a female infant with benign neonatal hemangiomatosis who had dramatic conjunctival involvement. The spectrum of neonatal hemangiomatosis is reviewed, highlighting the importance of differentiation of the two extremes of this disorder. PMID- 12125942 TI - Vascular variant of keratosis lichenoides chronica associated with hypothyroidism and response to tacalcitol and acitretin. AB - Keratosis lichenoides chronica (KLC) is a rare chronic progressive cutaneous disease that is part of the heterogeneous group of lichenoid dermatoses. The typical clinical presentation is characterized by lichenoid hyperkeratotic papules and nodules arranged in a linear and reticular pattern on the trunk and extremities. Our case confirms the existence of a vascular variant of KLC. There is no consensus about its treatment, since it is refractory to many different treatment modalities. We report the effectiveness of acitretin in KLC in combination with tacalcitol. KLC is of unknown aetiology, but is perhaps associated with systemic diseases, most importantly glomerulonephritis and lymphoma. This is the second case associated with hypothyroidism. PMID- 12125943 TI - A new calcipotriol/betamethasone formulation with rapid onset of action was superior to monotherapy with betamethasone dipropionate or calcipotriol in psoriasis vulgaris. AB - In this study, we compared a new combination ointment containing both calcipotriol and betamethasone dipropionate with betamethasone dipropionate ointment (Diprosone) and calcipotriol ointment (Daivonex) in patients with psoriasis vulgaris; 1106 patients were randomized to twice daily double-blind treatment with combination, betamethasone dipropionate or calcipotriol for 4 weeks. Patients then received twice daily calcipotriol, unblinded, for a further 4 weeks. Mean percentage change in PASI at end of the double-blind phase was 74.4 (combination group), -61.3 (betamethasone group) and -55.3 (calcipotriol group). Mean difference (95% Cl) combination-betamethasone was -13.1 (-16.9 to 9.3, p < 0.001) and for combination-calcipotriol -19.0 (-22.8 to -15.2, p <0.001). The differences in PASI were also statistically significant after 1 week. In the double-blind phase, 8.1% of patients (combination) reported lesional/ perilesional adverse reactions compared to 4.7% (betamethasone) and 12.0% (calcipotriol). In the combination group, mean PASI at the end of the double-blind phase was 2.5, and at end of the unblinded phase 3.6, compared with 3.9 and 4.1 (betamethasone) and 4.4 and 3.7 (calcipotriol). Calcipotriol/betamethasone combination is more effective and has a more rapid onset of action than either active constituent used alone, and is well tolerated. It is safe to transfer patients from combination to calcipotriol, with maintenance of clinical effect. PMID- 12125944 TI - Systemic immunotherapy with topical dinitrochlorobenzene as additional treatment of alopecia areata. PMID- 12125945 TI - Two unusual cases of Kaposi's varicelliform eruption. PMID- 12125947 TI - Tinea caused by Trichophyton soudanense in non-African subjects in Spain. PMID- 12125948 TI - Sporotrichoid spread of Mycobacterium chelonae in a presumably immunocompetent patient. PMID- 12125946 TI - Nail psoriasis: a combined treatment using calcipotriol cream and clobetasol propionate cream. PMID- 12125949 TI - A case of primary cutaneous actinomycosis. PMID- 12125950 TI - Self-healing juvenile cutaneous mucinosis in an infant. PMID- 12125951 TI - Fixed erythrodysaesthesia plaque due to gemcitabine and epirubicin treatment. PMID- 12125952 TI - Cold urticaria and angiotensin converting enzyme inhibitor. PMID- 12125953 TI - Association of HIV infection, pyoderma gangrenosum and psoriasis. PMID- 12125954 TI - Pellagra-like skin lesions associated with Wernicke's encephalopathy in a heavy wine drinker. PMID- 12125955 TI - Eccrine naevus: case report and literature review. PMID- 12125956 TI - Cutaneous Mycobacterium chelonae infection in a presumably immunocompetent host. PMID- 12125957 TI - Analysis of the expression of cutaneous lymphocyte-associated antigen on the peripheral blood and cutaneous lymphocytes of alopecia areata patients. AB - Alopecia areata has been reported to be accompanied by abnormal autoimmune dysfunction. We examined the expression of cutaneous lymphocyte-associated antigen (CLA), which is a skin-specific lymphocyte homing receptor, in the peripheral blood lymphocytes and skin of patients with alopecia areata. In the patients' peripheral blood, the percentage of CLA-positive CD4+ or CD8+ lymphocytes, was significantly higher than that of normal controls. The patients with severe or progressive alopecia areata showed a much higher CLA-positivity compared to patients recovering from the disease. A chronological study showed that the percentage of CLA-positive peripheral blood lymphocytes, CD4 + or CD8 + lymphocytes decreased in parallel with the patients' good clinical course. The CLA-positivity in peripheral blood lymphocytes, CD4+ or CD8+ lymphocytes of patients with alopecia areata who did not respond to oral corticosteroid therapy remained higher than in those who responded well to the treatment. In the affected scalp skin, many infiltrating lymphocytes around the hair follicles, which were CD4+ or CD8+ lymphocytes, expressed CLA. These findings suggest that the CLA-positivity correlates with clinical activity and that CLA-positive CD4+ or CD8+ lymphocytes may play an important role in alopecia areata. PMID- 12125958 TI - Iodine and gliadin challenge on oral mucosa in dermatitis herpetiformis. AB - Oral lesions and mucosal inflammatory changes may appear in dermatitis herpetiformis (DH). We examined whether potassium iodine, known to initiate blisters in the DH skin, or wheat gliadin, responsible for T-cell-dependent intestinal damage, can induce visible or microscopic changes in oral mucosa. Six patients with active DH were challenged with crude gliadin and 50% potassium iodine applied in patch test chambers on buccal mucosa for 12 h. After reading, biopsies were taken from the challenged and non-challenged mucosa. No macroscopic or microscopic vesicles were seen. However, gliadin- but not iodine-challenged epithelium showed increased numbers of CD4+ lymphocytes in all 5 patients with representative specimens (p = 0.06). No marked changes were found in the numbers of CD8+ or TcR alpha/beta+ lymphocytes, and the numbers of TcR gamma/delta+ cells remained at a low level. The results show that oral mucosa is resistant to production of macroscopic or microscopic DH lesions. It is, however, capable of reacting to locally applied gliadin by a T-cell response consisting of CD4+ lymphocytes. PMID- 12125959 TI - Molecular genetic analysis of HLA-DR and -DQ alleles in Spanish patients with melanoma. AB - Controversial data have been reported about HLA alleles and susceptibility to melanoma. The relationship between distribution of HLA alleles in patients with melanoma and susceptibility to tumour was analysed, to study the possible correlation between HLA class II DQA1, DQB1 and DRBI genes and melanoma in a Spanish population. Genomic DNA from 82 patients with melanoma and 367 random healthy donors, from the same geographic area, were typed by PCR-SSP (sequence specific primers). The patients were also divided into different groups according to the age and presence of cancer relatives, and compared with the controls. None of these HLA class II alleles showed significant positive or negative associations with either the overall population of patients with melanoma or the considered subgroups. Moreover, values for relative risk of DQB1*0301, DQB1*0302, DQB1*0303, DQB*05, DQA1*0401, DQA1*0101/0104 and DRB*08, which have been reported to be increased or decreased in patients with melanoma, were very low and of no statistical significance. Our results indicate that HLA class II alleles may not contribute to a strong susceptibility to melanoma in the Spanish population, although further studies on larger series are needed to corroborate this. Key words: PMID- 12125960 TI - The tandem repeated irritation test: a new method to assess prevention of irritant combination damage to the skin. AB - The effect of a protective cream was tested in a new tandem repeated irritation test with tandem application of 0.5% sodium lauryl sulphate (SLS) and undiluted toluene. The irritants were applied twice daily for 30 min to the ventral forearms of 20 volunteers. Irritant cutaneous reactions were quantified by a visual score, transepidermal water loss, chromametry and skin capacitance. Concurrent application of SLS/toluene induced stronger reactions than those caused by twice daily application of each irritant on its own. A protective effect of the protective cream was obtained against all treatment combinations and was significant for SLS/SLS (p < or = 0.01) and SLS/ toluene (p < or = 0.05). Our results indicate that the tandem repetitive irritation test has great potential in the evaluation of skin care products to prevent irritant contact dermatitis. PMID- 12125961 TI - Atopic dermatitis and concomitant disease patterns in children up to two years of age. AB - There are few prospective studies of atopic dermatitis and co-existing diseases such as respiratory infections in children up to 2 years of age. Using annual questionnaires, we studied the cumulative incidence of atopic dermatitis and concomitant symptoms indicating other atopic diseases and respiratory infections in 0-2-year-old children in a prospective birth cohort of 4089 children. We found associations between atopic dermatitis and asthma (ratio of proportion 1.45, 95% CI 1.16-1.80), allergic rhinoconjunctivitis (RP 2.25, CI 1.77-2.85), adverse reactions to foods (RP 3.20, CI 2.83-3.62), urticaria (RP 2.04, CI 1.80-2.31), acute otitis media (RP 1.13, CI 1.05-1.21), more than one pneumonia during the first and/or second year of life (RP 2.17, CI 1.14-4.15), and use of antibiotics at least twice yearly (RP 1.29, CI 1.07-1.56). The association between atopic dermatitis and respiratory infections persisted after stratification for asthma. There was a higher proportion of atopic disease manifestations, but not respiratory infections, in children with onset of atopic dermatitis during the first year of life than during the second. The study shows that during the first 2 years of life there is a significant association not only between atopic dermatitis and other atopic disease manifestations, but also between atopic dermatitis and respiratory infections manifested in an increased rate of acute otitis media, pneumonia and use of antibiotics. PMID- 12125962 TI - ALA- and ALA-ester-mediated photodynamic therapy of human glioma spheroids. AB - The effects of photodynamic therapy (PDT) in human glioma spheroids incubated in 5-aminolevulinic acid (ALA), or ALA esters, are investigated. Spheroid survival and growth are monitored following PDT at representative drug concentrations, light doses, and dose rates. The primary finding of this study is that the response of human glioma spheroids to PDT with lipophilic ester derivatives, such as benzyl-ALA and hexyl-ALA, is equivalent to that observed with ALA, however, this equivalency is obtained for ester concentrations 10-20 times lower than the parent compound. The enhanced efficiency of the esters is likely due to their increased membrane penetrance. Potential clinical advantages of using lipophilic esters in PDT of gliomas are discussed. PMID- 12125963 TI - Expression of IGF-II, IGFBP-2, -5, and -6 in meningiomas with different brain invasiveness. AB - Meningiomas show clinical characteristics that vary from very benign to clearly malignant with rapid invasive growth and metastases. This study was undertaken to analyze the expression of members in the insulin-like growth factor (IGF) system in meningiomas showing different degrees of brain invasion. Tissue samples from 16 meningiomas were analyzed for members in the IGF family by mRNA in situ hybridization. The meningiomas comprised three groups: I. Benign meningiomas that did not interfere with the arachnoid plane and showed no edema. II. Benign meningiomas that did not respect the arachnoid plane and tumors that caused edema. III. Aggressive and malignant meningiomas that caused edema and showed brain invasion. IGF-II mRNA was identified in all tumors analyzed, and with a clear increase in expression observed in group III tumors. IGFBP-2 mRNA was detected in equal levels in all tumors. IGFBP-5 mRNA levels were highest in the benign group without edema (I) of meningiomas whereas IGFBP-6 mRNA levels were highest in the group with brain invasion (III). Brain invasiveness in degrees from respect of the arachnoid membrane progressing to frank brain invasion correlated with increases in IGF-II and IGFBP-6 expression. High levels of IGFBP 6 may not inhibit IGF-II actions, which is known to be growth promoting in tumors. Instead, IGFBP-6 appears to have an importance for the characteristics of edema and brain invasion in meningiomas. PMID- 12125965 TI - Orbital apex syndrome: a rare presentation of extramedullary hematopoiesis: case report and review of literature. AB - We describe a case of compressive neuropathy in the orbital apex due to extramedullary hematopoiesis (EMH). A 64-year-old man with Polycythemia Rubra Vera developed unilateral visual loss, proptosis, complete ophthalmoplegia, and facial paresis. Bone marrow biopsy showed myelofibrosis. Magnetic resonance imaging demonstrated enhancement at the orbital apex and subtle optic canal narrowing. Decompression of the optic nerve with biopsy of surrounding bone showed EMH. The patient received a course of radiation without benefit. We suggest including the diagnosis of EMH of the orbital apex bones in the differential diagnosis of patients with myeloproliferative disorders who develop an orbital apex syndrome. PMID- 12125964 TI - Expression and functional activity of the ABC-transporter proteins P-glycoprotein and multidrug-resistance protein 1 in human brain tumor cells and astrocytes. AB - The poor prognosis of glioma patients is partly based on the minor success obtained from chemotherapeutic treatments. Resistance mechanisms at the tumor cell level may be, in addition to the blood-brain barrier, involved in the intrinsic chemo-insensitivity of brain tumors. We investigated the expression of the drug-transporter proteins P-glycoprotein (P-gp) and multidrug-resistance protein 1 (MRP1) in cell lines (N = 24) and primary cell cultures (N = 36) from neuroectodermal tumors, as well as in brain tumor extracts (N = 18) and normal human astrocytes (N = 1). We found that a considerable expression of P-gp was relatively rare in glioma cells, in contrast to MRP1, which was constitutively overexpressed in cells derived from astrocytomas as well as glioblastomas. Also, normal astrocytes cultured in vitro expressed high amounts of MRPI but no detectable P-gp. Meningioma cells frequently co-expressed P-gp and MRP1, while, most of the neuroblastoma cell lines express higher P-gp but lower MRP1 levels as compared to the other tumor types. Both, a drug-exporting and a chemoprotective function of P-gp as well as MRP1 could be demonstrated in selected tumor cells by a significant upregulation of cellular 3H-daunomycin accumulation and daunomycin cytotoxicity via administration of transporter antagonists. Summing up, our data suggest that P-gp contributes to cellular resistance merely in a small subgroup of gliomas, but frequently in neuroblastomas and meningiomas. In contrast, MRP1 is demonstrated to play a constitutive role in the intrinsic chemoresistance of gliomas and their normal cell counterpart. PMID- 12125966 TI - Quality of life in brain tumor patients: the relative contributions of depression, fatigue, emotional distress, and existential issues. AB - Neuropsychiatric problems, and how they interact to impact on quality of life (QOL) in brain tumor patients, are generally poorly understood. The objectives of this study were: (1) to document the prevalence of depression, fatigue, emotional distress, and existential issues in a sample of brain tumor patients (2) to examine the interconnectedness of these problems, and (3) to explore their relationship with disease-related variables and QOL. This is a cross-sectional, questionnaire-based survey of 73 patients with primary brain tumors who presented to a neurological clinic at a tertiary cancer centre for ongoing care. Data for 60 participants (29 women, 31 men) who completed validated questionnaires were retained for analysis. Results showed that there was a high burden of depressive symptoms as measured by the Beck Depression Inventory-II (mean score 11.1, SD 7.4), with 38% of the sample scoring in the clinically depressed range. Overall QOL scores for this sample were similar to a reference sample of brain tumor patients. The scores on the existential subscale of the McGill Quality of Life questionnaire were comparable to those of a reference sample of cancer patients receiving ongoing care (mean score 7.2; SD 1.7). Fifty per cent of the sample could be classified as struggling with existential issues. Although scores reflecting depression, fatigue, emotional distress, and existential problems were interrelated, the presence of depressive symptoms was the single most important independent predictor of QOL in this cohort of brain fumor patients. Implications for treatment are discussed. PMID- 12125967 TI - Maternal smoking during pregnancy and the risk of childhood brain tumors: a meta analysis of 6566 subjects from twelve epidemiological studies. AB - OBJECTIVE: Prior epidemiological studies suggest a possible association between maternal smoking during pregnancy and risk of childhood brain tumors. A meta analysis was performed statistically pooling all available observational studies on this topic in order to evaluate this suspected association. METHODS: Using previously described methods, a protocol was developed for a meta-analysis examining the association between maternal smoking during pregnancy and subsequent development of primary brain tumors in their offspring. Literature search techniques, study inclusion criteria and statistical procedures were prospectively defined. Data from epidemiological studies were pooled using a general variance-based meta-analytic method employing confidence intervals previously described by Greenland. The outcome of interest was a summary relative risk (RRs) reflecting the risk of childhood brain tumor development associated with mother's smoking during the index pregnancy. Sensitivity analyses were performed when necessary to explain any observed statistical heterogeneity and/or to evaluate the impact of demographic or study characteristics on the summary estimate of effect. RESULTS: Twelve observational studies meeting protocol specified inclusion criteria were obtained via a comprehensive literature search. These studies enrolled a total of 6566 patients. Analysis for homogeneity demonstrated that the data were homogeneous (P > 0.50) and could be statistically combined. Pooling all twelve reports yielded an RRs of 1.05 (0.90-1.21), a non statistically significant result suggesting no clear association between maternal smoking during pregnancy and risk of childhood brain tumor development. Numerous sensitivity analyses examining the possible effect of study design and various patient characteristics failed to show any influence on the RRs further supporting the observed lack of association. CONCLUSION: The available epidemiological data do not support a clear association between maternal smoking during pregnancy and pediatric brain tumor development. Although it appears likely that no association exists, limitations in study designs limit definitive conclusions based on available data. PMID- 12125968 TI - Supratentorial ganglioglioma and epilepsy: postoperative seizure outcome. AB - Ganglioglioma is increasingly recognized as being a cause of epilepsy. However, controversy still exists regarding the type of surgery required for optimal seizure control. To find out the factors associated with seizure outcome and to clarify treatment guidelines, a retrospective analysis of 29 epileptic patients who had pathologically proven gangliogliomas that were operated on at our institute during a 13-year period, was performed. There were 23 temporal and six extratemporal gangliogliomas, with a mean age of 16.5 years at surgery. Epilepsy was medically intractable in 14 (48.3%) patients. There were 26 temporal and six extratemporal resections including three re-operations. Three patients underwent intracranial electroencephalography (EEG) recordings (invasive monitoring). Intraoperative electrocorticography (ECoG) was performed in four patients. The mean duration of follow-up after surgery was 36.5 months. The Fisher's Exact test (two-tailed) was used to assess the association between the seizure outcome and several preoperative and operative parameters. Surgical treatment rendered most patients (75.9%, 22/29) seizure free. All three patients who underwent re operation were seizure-free postoperatively. Eleven (47.8%) of the 23 temporal lobe gangliogliomas were associated with cortical dysplasia. Postoperative incomplete seizure control was associated with the presence of cortical dysplasia (p < 0.001). Good seizure outcome is expected in patients with gangliogliomas, despite years of medically intractable epilepsy, once the tumor is resected. Invasive monitoring/intraoperative electrocorticography is recommended for patients with suspected cortical dysplasia on MRI or for the patients with persistent epilepsy after ganglioglioma resection. PMID- 12125969 TI - The combination of X-ray-mediated radiosurgery and gene-mediated immunoprophylaxis for advanced intracerebral gliosarcomas in rats. AB - Rats with advanced, imminently lethal, approximately 4 mm diameter, left-sided intracerebral 9L gliosarcoma (9LGS), a well characterized malignant tumor with some similarities to human high-grade astrocytomas, were used as a therapy model 14 days post-implantation of 10(4) cells. Such tumor-bearing rats die within two weeks (median, 6 days) thereafter if untreated. However, if these tumors are exposed on day 14 to 12-25 Gy of an electron-equilibrated 6 MV photon beam (radiosurgery), survival is extended about 5-6 fold to a median of 34 days, but long-term survival (> 1 year) is increased only to approximately 18%. Multiple subcutaneous inoculations of radiation-disabled 9LGS cells post-radiosurgery (immunoprophylaxis) extended lifespan and long-term (> 1 year) survival minimally (median, 37 days; 25%, respectively). In sharp contrast, radiosurgery followed by multiple subcutaneous inoculations of radiation-disabled 9LGS cells that had been transfected with granulocyte macrophage colony stimulating factor (GMCSF), a cytokine with demonstrated immune-enhancing properties (i.e. gene-mediated immunoprophylaxis, GMIMPR) increased long-term survival to approximately 67%. To our knowledge, these results are the first to show that the combination of photon radiosurgery and GMIMPR is effective for an advanced, imminently lethal brain tumor in a mammal. These data raise the possibility that GMIMPR following radiation therapy might prove effective for the treatment of some human malignant gliomas. PMID- 12125971 TI - PCNA and Ki-67 in central nervous system tumors: correlation with the histological type and grade. AB - Determination of proteins in the control of proliferation in normal cells helps a better understanding of cellular transformation and proliferation mechanisms. Measurement of proliferative activity is important in determining the tumor grade, recurrence span and malignancy. Proliferating cell nuclear antigen (PCNA) and Ki-67 are two of the nuclear markers used to demonstrate the proliferative phase of the cell cycle. In the present study, 63 central nervous system (CNS) tumors of various histologic types, diagnosed in Cukurova University Medical Faculty. Department of Pathology and graded according to WHO grading system were examined for PCNA and Ki-67 monoclonal antibodies using immunohistochemistry. Results were analyzed with statistical methods. Distribution of PCNA and Ki-67 LI (labeling index) values were determined for different tumor types. The highest PCNA and Ki-67 LI values were detected in medulloblastoma, malignant meningioma, primitive neuroectodermal tumor (PNET) and glioblastoma (GBM) groups, while pilocytic astrocytoma, meningioma, craniopharyngioma and oligodendroglioma showed the lowest values. In such tumors, the correlation between the increasing grade and PCNA and Ki-67 LI values were statistically significant. A correlation between the clinical outcome and Ki-67 and PCNA LI values was also detected. Conclusively, both markers can be used to evaluate the tumor grade and to asses the possibility of recurrence and malignancy in CNS tumors. PMID- 12125970 TI - Enhanced apoptosis in pilocytic astrocytoma: a comparative study of apoptosis and proliferation in astrocytic tumors. AB - Both cell proliferation and cell death occur simultaneously in tumor tissue, and extent of tumor growth reflects the net balance of these events. We correlated cell proliferation, spontaneous cell death, and alterations in tumor suppressor proteins with one another and with survival of patients with primary astrocytic tumors. In 39 astrocytic tumor specimens (6 pilocytic astrocytomas, 14 fibrillary astrocytomas, 9 anaplastic astrocytomas, and 10 glioblastomas), we determined the MIB-1 labeling index, the apoptotic ratio according to nick end labeling with morphologic confirmation, the p53 labeling index, and the presence of p53 or PTEN mutations. MIB- I labeling indices of pilocytic astrocytomas, fibrillary astrocytomas, anaplastic astrocytomas, and glioblastomas were 0.30+/-0.32; 1.84+/ 1.87; 19.3+/-6.42; and 28.0+/-14.5 (mean +/- SD), respectively. Corresponding apoptotic ratios were 17.9+/-5.16; 3.96+/-3.57; 1.18+/-0.93; and 2.11+/-1.60 (mean +/- SD). The apoptotic ratio in pilocytic astrocytomas was significantly higher than in other astrocytic tumors (fibrillary astrocytomas, p < 0.05; anaplastic astrocytomas and glioblastomas, p < 0.01). MIB-1 showed a significant negative correlation with apoptosis (p < 0.01). MIB- I and apoptosis showed significant negative and positive correlations with patient survival (p < 0.01). Mutations of p53 and PTEN show no correlation with survival and apoptotic ratio. The apoptotic ratio can clearly distinguish pilocytic astrocytomas from other tumors, and this biological feature may reflect less aggressive growth of pilocytic astrocytomas. PMID- 12125972 TI - Is there a requirement for adjuvant therapy for choroid plexus carcinoma that has been completely resected? AB - Choroid plexus carcinomas (CPC) are rare central nervous system malignancies. While attempted surgical resection is imperative, the benefit of adjuvant therapy, particularly in the setting of a gross total resection (GTR), is unclear. We reviewed all pediatric cases of CPC reported in the literature between 1985 and 2000. Seventy-five cases of CPC were identified. Mean age at the time of diagnosis was 26 months. Thirty-seven patients had a GTR and 38 patients had a subtotal resection (STR). Thirty-eight cases (51%) were alive and 37 cases (49%) were deceased at time of publication. For cases with a GTR, survival was 84% compared to an 18% survival for patients with a STR. For patients with a GTR, all forms of adjuvant therapy were statistically equivalent. Our retrospective literature review confirms the importance of GTR in the therapy of CPC, with GTR alone being the single most important predictor of survival. The prognosis is poor for any patient with a STR, with the exception of those patients for whom adjuvant therapy allowed for an eventual GTR. The small number of patients receiving a GTR and no further therapy precluded a statistical comparison of no therapy in the setting of a GTR versus any form of adjuvant therapy. However, all four of these patients are alive, raising the possibility that adjuvant therapy in the setting of a GTR may not be required. PMID- 12125973 TI - Effect of breathing a hyperoxic hypercapnic gas mixture on the oxygenation of meningiomas; preliminary results. AB - For meningiomas in which complete resection is impossible stereotactic radiosurgery and radiotherapy are increasingly important therapeutical options. The radiosensitivity of meningiomas may be improved by increasing tumor oxygen levels. Hyperoxygenating agents, like breathing a hyperoxic hypercapnic gas mixture, have already been applied successfully in the treatment of other tumors. The aim of this study was to explore the effect of breathing a hyperoxic hypercapnic gas mixture on tumor blood oxygenation of meningiomas using magnetic resonance imaging (MRI) methods. Three patients with convexity meningiomas were each measured twice; with and without breathing the hyperoxic hypercapnic gas mixture. Tumor blood oxygenation changes were measured using blood oxygen level dependent MR imaging. Dynamic contrast enhanced MRI was used to assess functional changes of tumor vasculature. A significant increase in tumor blood oxygenation was observed under hypercapnic hyperoxic conditions in all patients, exceeding the increase in normal brain tissue. It was concluded that the oxygenation status of meningiomas can be improved by breathing a hyperoxic hypercapnic gas mixture. PMID- 12125974 TI - Surgical outcome and prognostic factors of spinal intramedullary ependymomas in adults. AB - The goal of treatment for spinal ependymoma is complete removal with minimal postoperative neurological deficit. The authors correlated the results of surgical management for spinal cord ependymoma with the rate of postoperative disease progression and the prognostic factors. Thirty-one cases of spinal ependymomas, surgically treated between 1979 and 1998, were retrospectively analyzed. The authors reviewed clinical features, radiological characteristics and operative findings for the surgical outcome analysis. Thirty-five percent of patients with preoperative Nurick's grade better than grade 4 showed improvement in functional status, whereas no improvement was observed in patients with preoperatively poorer functional status (P = 0.05). The proportion of complete surgical removals was influenced by tumor location (40% in cases around the conus versus 97% in other regions, P = 0.003) and histology (42% in the myxopapillary subtype versus 97% in the non-myxopapillary subtype, P = 0.001). Disease progression was observed in six cases, the mean progression free interval after surgical removal was 83 months and the 5-year progression free rate was 70%. Disease progression was found in two out of 23 cases from the complete removal group and in four out of eight cases from the incomplete removal group (P = 0.008). In the aspect of disease progression, the only statistically significant factor by multivariate analysis was the surgical extent of removal (P = 0.010). Of those patients where there was incomplete removal, radiation therapy lead to improved clinical results, which were not statistically significant (P = 0.27). In the surgical treatment of spinal cord ependymoma, preoperative functional status and the extent of removal were the significant prognostic factors influencing postoperative outcome. Early diagnosis is vital and complete removal of the tumor should be attempted in all surgical treatment of spinal ependymoma. PMID- 12125975 TI - Gemcitabine and radiotherapy in the treatment of brain metastases from transitional cell carcinoma of the bladder: a case report. AB - Hematogenous metastases occur in over 50% of muscle-invasive transitional cell carcinomas (TCC) of the bladder. Despite treatment (mostly surgery and radiotherapy), patients with brain metastases have an especially poor prognosis (median survival 2-5 months), making palliative treatment an important consideration. We followed a 60-year-old man with multiple brain metastases who was ultimately treated with gemcitabine chemotherapy. He underwent a cystectomy in 1997 because of a T3a N0 M0 G3 TCC of the bladder. Two years later, he developed one brain metastasis and one lung metastasis. Both metastases were resected and adjuvant chemotherapy was planned. Before chemotherapy, the patient suffered from headaches and symptoms of hemiparesis. A magnetic resonance imaging (MRI) showed multiple brain metastases of up to 2 cm, particularly in the brain stem. The patient underwent whole-brain radiotherapy with 30 Gy, followed by four cycles of a 3-week gemcitabine (800 mg/m2 on days 1 and 8) schedule. Another MRI showed a nearly complete response after four cycles of chemotherapy, with only small residual tumors remaining in the brain stem. This impressive activity was accomplished without adverse side effects, suggesting that radiotherapy combined with gemcitabine monotherapy may be an excellent choice for palliative treatment of TCC of the bladder. PMID- 12125976 TI - The alteration of prostaglandin E2 levels in patients with brain tumors before and after tumor removal. AB - BACKGROUND: Both experimental and human tumors often synthesize high levels of prostaglandins, most notably prostaglandin E2 (PGE2). This compound may play an important role in tumor growth and immunosuppression. Little is known of the production of PGE2 by brain tumors. The present study was designed to investigate the levels of PGE2 in the plasma of human brain tumors before and after tumor removal. METHODS: The plasma PGE2 levels of brain tumors before and after tumor removal were measured by high-performance liquid chromatography (HPLC). RESULTS: There is a significantly high concentration of PGE2 in malignant brain tumor before tumor removal. Significantly decrease of PGE2 concentration after total removal of the tumor was found both in the malignant and benign brain tumor groups (P = 0.0001 and P = 0.0039 respectively). However, compared to the control group, only malignant brain tumor showed a significant decrease of PGE2 concentration after tumor removal (P = 0.0009). CONCLUSION: Our study demonstrates the malignant brain tumor synthesized higher relative proportions of PGE2 and surgical removal of the brain tumor can reduce the production of PGE2. PMID- 12125977 TI - Radiosurgical boost for primary high-grade gliomas. AB - The purpose of this study was to retrospectively evaluate the survival of patients with high-grade gliomas treated with external beam radiotherapy with or without radiosurgical boost. From July 1993 to April 1998, 32 patients were selected, 15 of which received radiosurgery. Inclusion criteria were age > 18 years, histological confirmation of high-grade glioma, primary tumor treatment with curative intent, unifocal tumor and supratentorial location. All patients were found to be in classes III-VI, according to the recursive partitioning analysis proposed by the Radiation Therapy Oncology Group. The median interval between radiotherapy and radiosurgery was 5 weeks (range 1-13). Treatment volumes ranged from 2.9 to 70.3 cc (median 15.0 cc). Prescribed radiosurgery doses varied from 8.0 to 12.5 Gy (median 10.0 Gy). Radiosurgery and control groups were well balanced with respect to prognostic factor distributions. Median actuarial survival time in radiosurgery and control groups was 21.4 months and 11.6 months, respectively (p = 0.0254). Among patients with KPS > 80, median survival time was 11.0 months and 53.9 months in the control and radiosurgery groups, respectively (p = 0.0103). Radiosurgery was the single factor correlated with survival on Cox model analysis (p = 0.0362) and was associated with a 2.76 relative reduction in the risk of cancer death (95% confidence interval (CI) 1.07-7.13). Our results suggest that radiosurgery may confer a survival advantage for patients in RPA classes III-VI, especially for those with Karnofsky performance status >80. The definitive role of radiosurgical boost for patients with high-grade gliomas awaits the results of randomized trials. PMID- 12125978 TI - Determination of the subcellular distribution of mercaptoundecahydro-closo dodecaborate (BSH) in human glioblastoma multiforme by electron microscopy. AB - The subcellular distribution of mercaptoundecahydro-closo-dodecaborate (BSH) in glioblastoma multiforme tissue sections of several patients having received BSH prior to surgery was investigated by transmission electron microscopy (TEM) using antibodies against BSH and electron energy loss spectroscopy (EELS) and electron spectroscopic imaging (ESI). These microscopic techniques show that BSH is associated with extracellular structures, the cell membrane as well as with the chromatin in the nucleus. PMID- 12125979 TI - Expression of Nedd5, a mammalian septin, in human brain tumors. AB - The septins are a family of cytoskeletal GTPases that play an essential role in cytokinesis in yeast and mammalian cells. Nedd5 is a mammalian septin known to associate with actin-based structures such as the contractile ring and stress fibers. In the present study, we examined the expression of Nedd5 in a series of human brain tumor cell lines and surgical specimens by northern and western analyses. The temporal expression of Nedd5 in U373 astrocytoma cells at various timepoints throughout the cell cycle was determined by an analysis of lovastatin- and nocodazole-treated, synchronized cell populations. The intracellular localization of Nedd5 was determined by immunocytochemistry of steady state cultures and nocodazole-treated cultures enriched in M phase cells. The effects of inhibiting Nedd5 expression in human brain tumors was determined by stably transfecting U373 astrocytoma cells with an antisense-Nedd5 cDNA expression vector and by analyzing clones for Nedd5 expression by immunocytochemistry, morphological changes, cell growth and nuclear content. All human brain tumor cell lines and surgical specimens expressed Nedd5 transcript and protein. Synchronized U373 astrocytoma cells showed a relative increase in Nedd5 transcript levels from late Gl to G2M phases; and an increase in Nedd5 protein levels from S to G2M phases. Maximum expression of both transcript and protein levels was observed at the G2M phase. By immunocytochemistry, Nedd5 was concentrated at the cleavage furrow of mitotic cells. Double staining with Nedd5 and F-actin showed co-localization of Nedd5 with actin filaments except during cytokinesis. Antisense-Nedd5 expression led to an accumulation of nuclear content. These data suggest that Nedd5 is involved in the process of cytokinesis in human brain tumours. Nedd5 expression may be cell cycle-dependent with increased levels found at G2M phase. Blocking Nedd5 expression in astrocytoma cells by antisense interferes with the process of cytokinesis during cell division. PMID- 12125980 TI - Induction of apoptosis in glioma cells by molecules released from activated macrophages. AB - Macrophages play an important role in the regulation of malignant tumors. Although glioma contains abundance of macrophages, their role in apoptosis of glioma is not known. We stimulated macrophages with lipopolysaccharide and culture supernatants of activated macrophages were collected to treat glioma cells. The results showed that molecules released from activated macrophages significantly increased apoptosis of glioma via Fas/FasL and caspase-3 pathways. The level of soluble Fas did not appear to be involved in the mechanism responsible for apoptosis seen in this study, as its level was barely detected in both experimental and control groups. Two cytokines, TNFalpha and IFNgamma, were significantly elevated in the supernatant obtained from the activated macrophages. Considering an important role of these two molecules in the induction of apoptosis mediated by the Fas/FasL system, the present data suggested that TNFalpha and IFNgamma were the main molecules to trigger the cascade of apoptotic reactions in glioma cells. In conclusion, the present study indicates that molecules released from the activated macrophages provide significant signals to stimulate the expression of Fas/FasL and caspase-3, which function to induce apoptosis in glioma cells. PMID- 12125981 TI - Relationship between the expression of E-, N-cadherins and beta-catenin and tumor grade in astrocytomas. AB - Cadherins are cell-surface glycoproteins that mediate Ca2+-dependent, homophilic cell-cell adhesion. The classical cadherins, E- and N-cadherins, connect to beta catenin, the lining protein. There appears to be a relationship between their dysfunction and tumor invasion and metastasis. The aim of our study was to examine the possibility of a relationship between alterations in the E- and N cadherin and catenin expression and malignancy in astrocytomas. Forty-five astrocytomas (18 glioblastomas, 16 anaplastic astrocytomas, and 11 diffuse astrocytomas) were collected and stained immunohistochemically for cadherins and beta-catenin. None of the astrocytomas were immunoreactive for E-cadherin. N cadherin and beta-catenin were present at cell-cell borders in 61% of glioblastomas and 31% of anaplastic astrocytomas. The incidence of immunoreactivity for N-cadherin and beta-catenin increased significantly with the histological grade of astrocytomas (p = 0.001, by Kruskal-Wallis test). Moreover, in anaplastic astrocytomas and glioblastomas, the Ki-67 labeling indices in both N-cadherin-positive and beta-catenin-positive cases were higher than that in negative cases (p = 0.05 and 0.03, respectively, by Fisher's exact test). These results suggest that the expression of N-cadherin or beta-catenin may be related to the biological behavior of astrocytomas. PMID- 12125982 TI - Synergistic effect of genistein and BCNU on growth inhibition and cytotoxicity of glioblastoma cells. AB - OBJECTIVE: Recent experiments have shown that dietary soy isoflavones such as genistein can significantly suppress invasiveness and growth of a number of human malignancies. This study examined whether genistein, at a concentration typical of plasma levels following soy diet intake, in combination with 1,3-bis(2 chloroethyl)-1-nitrosourea (BCNU, carmustine) exhibited an additive or synergistic inhibitory effect on the growth of glioma cells. METHODS: The human glioblastoma multiforme (GBM) cell line U87 and the rodent C6 glioma were treated with genistein at 4 microM, combined with BCNU (0-50 microM). Monolayer cell growth and cytotoxicity, as measured by colonigenic survival in soft agarose, were then compared in control and drug-treated cultures. Presence of apoptosis, using the DNA ladder assay and laser scanning cytometry (LSC), was investigated in all cell lines at those concentrations where an enhancement of antiproliferative effect of BCNU in presence of genistein was observed. RESULTS: A 32-41% increase in monolayer growth inhibition and a 28-42% increase in colony cytotoxicity in the U87 cell line were observed when genistein (4 microM) was added to BCNU in the 0-10 microM dose range. In the C6 cell line, a 30-36% increase in monolayer growth inhibition and a 39-54% increase in colony cytotoxicity were observed with the BCNU dose range of 0-50 microM. All experiments showed a significant increase in growth inhibition and a decrease in colonogenic survival (P < 0.05). We were unable to detect apoptosis in any of the lines when genistein was combined with BCNU. CONCLUSION: These results indicate that genistein at typical adult dietary plasma levels can significantly enhance the antiproliferative and cytotoxic action of BCNU. The implication for treatment of GBM may be a reduction in the chemotherapeutic dose recommendations of these agents and subsequently a decrease in the risk of treatment sequelae for these patients. PMID- 12125983 TI - Differential expression of SOX4 and SOX11 in medulloblastoma. AB - Primitive neuroectodermal tumors (PNETs) are composed of immature neuronal precursor cells and sometimes more mature neuronal cell types. Medulloblastomas, occuring in the cerebellum, represent the most common PNET and are broadly classified into two subgroups: classical and desmoplastic. Desmoplastic medulloblastomas exhibit a slightly better prognosis than classical medulloblastomas. However, there are currently no good molecular markers available to distinguish clinical outcome and similar treatment is used for most patients with associated complications. It has been shown that neoplastic cells in these tumors recapitulate stages in maturation of normal human neuroblasts; therefore, embryological studies of the earliest events in the development of the cerebellum may provide useful information about the molecular behavior of the tumor. Transcription factors such as Sox proteins involved in neural development may also play a role in the etiology of brain tumors. Sox4 in particular has been implicated in the biology of several other types of cancer. We have studied the expression of Sox4, and the closely related Sox11 gene, in medulloblastomas. Sox4 and Sox11 were strongly expressed in most classical medulloblastomas but only weakly in desmoplastic medulloblastomas. The expression profile of these two genes in developing cerebellum was also analyzed. Our results suggest that strong Sox4 and Sox11 expression in classical medulloblastomas reflects their maturation dependent expression during normal cerebellum development, and that they may therefore provide markers to divide tumors into clinically relevant subgroups. PMID- 12125984 TI - Concurrent modified PCV chemotherapy and radiotherapy in newly diagnosed grade IV astrocytoma. AB - Adjuvant chemotherapy (CT) in the treatment of grade IV astrocytoma is at best modestly effective. The radiosensitizing effect of CT may confer an advantage to concurrent radiotherapy (RT) and CT. This study investigated concurrent procarbazine, lomustine, and vincristine (PCV) CT in newly diagnosed grade IV astrocytoma patients. METHODS: From 1992 to 1997, patients diagnosed with grade IV astrocytoma (Daumas-Duport criteria), Karnofsky performance score (KPS) > or =70 and age <65 were offered CT. Twenty-seven study patients received concurrent modified PCV plus partial brain RT. Twenty-seven controls treated at the same institution with cranial RT alone were matched for histology, age and KPS. RESULTS: Median age was 49 years and mean KPS was 80 for both groups. Debulking operations were more frequent in study patients than controls (p = 0.034). One year survival was 70% and 56%, while median survival was 82 weeks and 53 weeks for the study and control groups, respectively (p = 0.1554). CT complications were predominantly hematologic, grades II and III. Two patients developed febrile neutropenia; one patient died from Pneumocystis carinii pneumonia. Nausea, vomiting and allergic reactions were all grade I. CONCLUSION: While a trend to increased survival was seen in the study, patients treated with concurrent PCV CT, this did not reach statistical significance. A phase III trial would help delineate the true effectiveness of concurrent CT in this population. Modified PCV is safe and reasonably well tolerated. PMID- 12125985 TI - Nuclear accumulation of basic fibroblast growth factor as a predictor for the recurrence of pituitary adenomas. AB - Although pituitary adenomas often recur, a reliable predictor for their recurrences has not yet been established. The aim of this study is to assess the utility of the nuclear accumulation of basic fibroblast growth factor (bFGF) as a predictor for the recurrence of pituitary adenomas. We studied 64 patients who had primary pituitary adenomas and underwent operations. The immunohistochemistry for bFGF and MIB-1 was retrospectively examined in paraffin-embedded tissues. The bFGF immunoreactivity in the cytoplasm was assigned one of four grades and the bFGF immunoreactivity in the nucleus was recorded as the bFGF nuclear index (NI), which was calculated as a percentage of tumor cells with the bFGF immunoreactivity in the nuclei in more than 1000 tumor cells. Recurrent adenomas were found in 7 patients during follow-up periods ranging from 8 to 134 months (mean: 57.3). Kaplan-Meier analysis demonstrated that high bFGF NI (>30%) correlated with poor recurrence free rate (p < 0.02). We assessed the relative contribution of bFGF NI to recurrence free by using multivariate (Cox's proportional hazards model) analyses with variable factors. Multivariate analysis showed that only bFGF NI was a potential predictor of recurrence free, independent of all other variables. High bFGF NI (>30%) had a relative risk of 8.9, with a 95% confidence interval of 1.0-74.9 (p < 0.05). We suggest that the bFGF NI may be a potentially useful predictor for the recurrence of pituitary adenomas. PMID- 12125987 TI - Imaging response to chemotherapy with RMP-7 and carboplatin in malignant glioma: size matters but speed does not. AB - INTRODUCTION: In recurrent malignant glioma, early imaging response to two courses of chemotherapy is generally considered to be predictive of good survival. We studied the relationships between initial tumour volume and speed of imaging response to chemotherapy in malignant glioma. METHODS: In 43 chemotherapy naive patients, 26 glioblastoma multiforme (GBM) + 17 anaplastic astrocytoma (AA), median age 45 years, MRI responses to intravenous cereport and carboplatin were assessed at baseline, then at 2 monthly intervals. Patients were classified as fast responders if they had reached a partial response (PR) after two courses, and slow responders if PR was achieved after three or more courses of chemotherapy. RESULTS: PR occurred in four patients with GBM (15%) and nine patients with AA (53%). Likelihood of response was related to initial tumour enhancing volume in GBM but not in AA. PR occurred in four of five GBM patients (80%) with initial volume < 15,000 mm3 and none of the 21 cases with an initial volume > 15,000 mm3. In patients achieving a PR, there was no association between speed or duration of response and eventual survival. Fast responders with AA were significantly older than slow responders (p = 0.033). CONCLUSIONS: Initial enhancing volume of GBMs may be an important predictor of imaging response. This has implications where response rates of phase II studies are reported and in stratification for phase III trials. Further work is necessary to confirm these findings with other types of chemotherapy and examine the relationship between proliferation markers and speed of response. PMID- 12125986 TI - An open label trial of sustained-release cytarabine (DepoCyt) for the intrathecal treatment of solid tumor neoplastic meningitis. AB - Drugs currently available for intrathecal administration are cleared rapidly from the CSF. DepoCyt is a slow-release formulation of cytarabine that maintains cytotoxic concentrations of free cytarabine in the CSF for >14 days following a single injection. DepoCyt was administered to 110 patients with a diagnosis of neoplastic meningitis based on either a positive CSF cytology (76) or neurologic and CT or MRI scan findings sufficient to document neoplastic meningitis (34). Patients were treated with DepoCyt 50mg every 2 weeks for 1 month of induction therapy by either lumbar puncture (LP) or intraventricular (IVT) injection. Patients without neurologic progression were candidates to receive an additional 3 months of consolidation therapy. All patients received dexamethasone 4 mg BID on days 1-5 of each cycle. Median time to neurologic progression was 55 days; median overall survival was 95 days. Among the 76 patients with a positive CSF cytology at baseline, 70 were evaluable for response, and of this group 19 (27%) attained the criteria for response (cytologic response in the absence of neurologic progression). The most important adverse events were headache and arachnoiditis. When drug-related, these were largely low grade, transient, and reversible. Drug-related grade 3 headache occurred on 4% of cycles; grade 3 or 4 arachnoiditis occurred on 6% of cycles. No cumulative toxicity was observed. DepoCyt injected once every 2 weeks produced a response-rate comparable to that previously reported for methotrexate given twice a week. The once in every 2-week dosing interval offers an advantage over conventional schedules (2-3 doses/week) used for other agents available for intrathecal injection. PMID- 12125989 TI - Cochlear implants in children--a review. AB - Over the past two decades, cochlear implantation has become a widely accepted treatment of deafness in children. Over 20,000 children have received cochlear implants worldwide. Hearing, language and social development outcomes have been positive. We review current issues in cochlear implantation, candidacy, evaluation, surgery, habilitation, ethics and outcomes. PMID- 12125988 TI - Temozolomide as second-line chemotherapy for relapsed gliomas. AB - BACKGROUND: Temozolomide, an imidazotetrazine prodrug has shown activity in phase II studies in patients with high-grade glioma at first recurrence. We assessed the efficacy of temozolomide as second-line therapy following failure of PCV chemotherapy in patients with recurrent/progressive gliomas. PATIENTS AND METHODS: Between September 1994 and November 2000, 32 patients with high-grade gliomas at second recurrence/progression received temozolomide as salvage therapy and results were reviewed retrospectively. RESULTS: Of 32 assessable patients 7 had clinical improvement; there were no imaging responses. Median survival of the cohort was 4 months, with 28% alive at 6 months. Age, performance status, histology and previous response to PCV chemotherapy did not predict for clinical response to temozolomide. CONCLUSION: In the small cohort of patients with recurrent malignant glioma who failed PCV chemotherapy temozolomide demonstrated limited activity as second-line treatment although this remains within the confidence intervals of response seen in patients with glioblastoma. PMID- 12125990 TI - Modiolar proximity of three perimodiolar cochlear implant electrodes. AB - A new generation of cochlear implant electrodes has been designed to position the stimulating contacts close to the modiolus in order to reduce power consumption and increase stimulation selectivity. The purpose of this study was to assess electrode position in the cochlea for three recently designed electrodes. Fifteen cadaveric temporal bones were implanted with one of three perimodiolar electrode arrays: Nucleus Contour; Med-El Combi40+ PM (developmental version); and Clarion HiFocus II. Image-enhanced videofluoroscopy and computer morphometrics were used to assess stimulating contact position relative to the modiolus. The mean distance (+/- 1 standard deviation) to the modiolus for all electrode contacts was 0.33 (+/-0.24), 0.30 (+/-0.27) and 0.16 mm (+/-0.19) for the Contour, Combi40+ PM and HiFocus II arrays, respectively. In addition, dynamic videofluoroscopy was used to correlate device-specific insertion characteristics with contact-to-modiolus distance. All three devices were successful in terms of locating electrode contacts very close to the modiolar wall. PMID- 12125991 TI - Protective effect of basic fibroblast growth factor on auditory hair cells after noise exposure. AB - The purpose of this study was to observe the protective effects of basic fibroblast growth factor (bFGF) on the cells of the inner ear using in vivo experiments. The studies were carried out using guinea pigs in which bFGF or artificial perilymph was perfused into the cochlea. The compound action potential (CAP) was measured before and after exposure to a sound simulating an explosion. The difference in CAP was significant between the bFGF-perfused group and the control group (p < 0.01, t = 3.896) and between the bFGF-perfused group and the artificial perilymph-perfused group (p < 0.05, t = 2.520). The cochleae were removed and hair cell loss estimated from surface preparations. Acoustic trauma caused loss of outer hair cells in the first and second turns of the cochlea in the bFGF-perfused group and the artificial perilymph-perfused group and partial loss of inner hair cells in the control group. Treatment with bFGF reduced the loss of inner hair cells compared to that of control animals. Our results demonstrate that treatment with bFGF protects the hair cells from acoustic trauma and may facilitate the recovery of hearing. PMID- 12125992 TI - A model of active Eustachian tube function in the rat: effect of modulating parasympathetic innervation. AB - The effect of altering secretion into the Eustachian tube by modulating cholinergic innervation was studied in the anaesthetized rat. Active properties of the Eustachian tube were determined by measuring the ability of reflex-induced swallowing to equalize against an increased pressure level in the bulla. Reflex induced swallowing was initiated by electrically stimulating the superior laryngeal nerve. Passive properties of the Eustachian tube were determined by increasing middle ear pressure until the Eustachian tube spontaneously opened. Blocking cholinergic neurotransmission with atropine had no effect on active or passive properties of the Eustachian tube. Potentiating cholinergic neurotransmission with neostigmine significantly impaired the ability of active swallowing to equilibrate middle ear pressure, but had no effect on passive properties of the Eustachian tube. The findings show that cholinergic nerve transmission, most likely from the parasympathetic division of the autonomic nervous system, can influence Eustachian tube function. We hypothesize that this effect is due to changes in surface tension in the Eustachian tube as a result of changes in secretion. PMID- 12125993 TI - Usefulness of virtual endoscopic three-dimensional reconstructions of the middle ear. AB - This work uses a new programme for producing 3D radiological images acquired by means of CT which enables the internal surfaces of the examined structures to be visualized. This new method, which is able to navigate inside organs in a similar way to fibreoptic endoscopy, is known as virtual endoscopy. CT examinations of the temporal bone were carried out using spiral equipment and endoscopic 3D processing was carried out on a separate workstation equipped with a volume rendering programme. Once the technical parameters necessary for obtaining a representation of the internal surfaces had been defined, a simulation of a virtual otoscopy was conducted by moving the virtual endoscope from the external auditory canal through the annulus to the tympanic cavity. The simulation can be obtained either by moving the endoscope by hand, using the mouse, or by defining a path along which the software automatically creates an endoscopic 3D reconstruction. The images thus obtained are projected sequentially to give a "movie" effect, i.e. a continuous progression of the endoscope. The average time required to conduct the procedure ranges from 20 to 30 min. A virtual endoscopic visualization of the middle ear was obtained which, in particular, generated images of the tympanic cavity with the ossicular chain. In our experience, virtual otoscopy shows the anatomy of the structures of the tympanic cavity in excellent detail and may be considered complementary to CT, providing useful images enabling better visual representation and understanding of this complex structure. Although clinical applications of the technique remain to be defined it may have a role to play in presurgical diagnostic evaluation of the ossicular chain, epitympanum and retrotympanum. PMID- 12125994 TI - Altered cupular mechanics: a cause of peripheral vestibular disorders? AB - It has taken many decades to arrive at today's concept of cupula mechanics in the stimulation of endolymphatic flows on the hair cells in the ampullae of the semicircular canal. While Steinhausen assumed free swing-door movement of the cupula in the 1930s, Hillman was the first to demonstrate firm cupula attachment to the ampulla wall as a physiological necessity in the 1970s. In contrast to the present clinical concepts of acute peripheral vestibular functional disorders (circulatory disturbances, viral or bacterial infection, altered electrolytes in the endolymph), this study examines the extent to which an impaired attachment mechanism can trigger peripheral vestibular disorders. For this purpose, we used a pigeon model (n = 8), in which mechanical detachment of the cupula from the ampulla wall was achieved by means of a targeted pressure increase in the ampulla of the lateral semicircular canal. In two additional animals the labyrinth was completely destroyed on one side in order to directly compare partial and complete vestibular disorders. In this way partial damage to the lateral semicircular canal ampulla presents a clinical picture whose symptoms are very similar to those of an idiopathic vestibular disorder in humans. Their intensity and course of compensation differ markedly from the symptoms of complete vestibular destruction. Subsequent histological examination revealed that the hair cells remained intact during the experimental detachment of the cupula. Our results thus show that only altered cupula mechanics seem to trigger the clinical picture of a peripheral vestibular disorder. This may result in completely new approaches to differential diagnosis and the therapy of vestibular neuronitis. PMID- 12125995 TI - Visual influence on postural control, with and without visual motion feedback. AB - Body sway was investigated in 20 healthy subjects to determine whether visual input must contain motion feedback information from the surroundings in order to influence postural control. Posturography was used to record body sway under the following visual conditions: eyes open with or without a restricted visual field; eyes open in ganzfield white light; eyes open in darkness with a head-fixed visual target; eyes open in darkness; and eyes closed in darkness. Stance was perturbed by means of a pseudorandomly applied vibratory stimulation to the calf muscles. Least sway was found with eyes open in an unrestricted visual field but increased in a restricted visual field. Greatest sway was found without visual motion feedback, i.e. under the following conditions: eyes closed; eyes open in darkness; eyes open in ganzfield white light; and with a head-mounted fixation point. Sway was significantly (p < 0.05) greater with eyes open in darkness compared with eyes closed during the initial 50 s with perturbations. After 150 s, sway was almost identical under the four test conditions without visual motion feedback. Standing with eyes open in darkness was initially a disadvantage compared with having the eyes closed. The postural control system may be programmed to expect visual feedback information when the eyes are open, which may delay changes in postural strategy. PMID- 12125996 TI - Respiratory epithelial adenomatoid hamartoma in the nasal cavity. AB - We report a 65-year-old male with a hamartoma in the left nasal cavity. A mass was found in the left nasal cavity and was diagnosed as a benign tumor on the basis of preoperative findings. Left lateral rhinotomy was performed to completely remove it and the lesion was found to have arisen from the inferior turbinate. The pathological diagnosis was respiratory epithelial adenomatoid hamartoma. We discuss the pathological features of this disease. PMID- 12125997 TI - Upregulation of MUC8 and downregulation of MUC5AC by inflammatory mediators in human nasal polyps and cultured nasal epithelium. AB - Polyps are believed to be the source of mucus hypersecretion in chronic inflammation of the sinus. However, it is not clear which mucins are responsible for the hypersecretion of mucus by nasal polyps. We describe the over-expression of MUC8 mRNA in nasal polyps and the upregulation of MUC8 mRNA expression and downregulation of MUC5AC mRNA expression by inflammatory mediators. We found that the level of MUC8 mRNA, but not the level of MUC5AC mRNA, increased in nasal polyps. We also found that there was an increase in intracellular mucin in nasal polyps, compared to the normal nasal inferior turbinate. A mixture of inflammatory mediators increased MUC8 mRNA expression and decreased MUC5AC mRNA expression in cultured normal human nasal epithelial cells. Among inflammatory mediators, IL-4 is responsible for the decrease in MUC5AC mRNA and MUC5AC mucin secretion. These results indicate that MUC8 may be one of the major mucins secreted from the polyp epithelium and that it may play an important role in the pathogenesis of mucus hypersecretion in chronic sinusitis with polyps. PMID- 12125998 TI - Progesterone receptor expression in angioleiomyoma of the nasal cavity. AB - In recent years sex steroid receptors have been identified in several types of human neoplasia. Angioleiomyoma of the nasal cavity and paranasal sinus is an extremely rare benign neoplasm which is more common in females (3.75:1 female:male ratio). To the best of our knowledge, only approximately 25 cases have been described. We describe a new case of angioleiomyoma of the nasal cavity that was found to be progesterone-receptor positive and oestrogen-receptor negative on immunohistochemical analysis. We suggest that the growth of this tumour may be hormone-dependent. PMID- 12125999 TI - Effect of neuraminidase on receptor-mediated adherence of Streptococcus pneumoniae to chinchilla tracheal epithelium. AB - The trachea whole organ perfusion technique was used to study the effect of the disruption of the Streptococcus pneumoniae neuraminidase nanA gene on bacterial adherence and alteration of the carbohydrate surface structures of respiratory epithelial cells. Six different lectin probes were used to examine alterations of the cell surface carbohydrates in chinchilla tracheal epithelium incubated in vitro with S. pneumoniae deltaNA1, a neuraminidase-deficient mutant, or its D39 parent strain. The labeling pattern revealed that the binding of wheat germ agglutinin (WGA), Erythrina cristagalli lectin (ECL), peanut agglutinin (PNA), Bandeiraea simplicifolia lectin II (BSL II) and succinylated WGA was significantly increased in the luminal surface of the trachea in the D39 incubated cohort compared with the uninfected control, which indicated that GlcNAc and D-galactose residues were exposed. Concurrently, decreased labeling with Sambucus nigra agglutinin (SNA) indicated that there were few sialic acid residues remaining in the tracheal epithelium subsequent to incubation with D39. The deltaNA1 neuraminidase-deficient mutant, however, did not induce any significant changes in the lectin labeling patterns, which were comparable to those of the control cohort. Moreover, adherence data expressed as colony-forming units (CFU) of S. pneumoniae per millimeter of trachea indicated a significant decline in the ability of deltaNA1 to adhere in vitro. We propose that products of the nanA gene have a significant impact on changes in the carbohydrate moieties in the tracheal epithelium, and may be responsible for the previously reported increased ability of the D39 parent to colonize the nasopharynx and invade the middle ear. PMID- 12126000 TI - Direction of lateral traction in Ejnell's technique: an experimental study and case report of bilateral vocal cord paralysis. AB - In order to develop the method of laterofixation of the vocal cord (Ejnell's method) in cases of bilateral vocal cord paralysis, six autopsy specimens of normal larynx were obtained. We inserted traction sutures under conditions of direct visualization and studied the relationship between the direction of the traction exerted by the suture and subsequent enlargement of the glottis. When the vocal cord was pulled perpendicular to the thyroid cartilage wing, the mean glottal area was 106.2% of its area before traction. When the vocal cord was pulled perpendicular to the median line of the glottis, the mean glottal area was 112.7% of its area before traction. The latter angle of traction therefore produced more efficient enlargement of the glottis (p < 0.05; paired t-test). Based on an anatomic study of cadaveric laryngeal regions, a formula was developed to predict at which point the needle should be placed in order to produce optimal results using Ejnell's technique. Further clinical studies will be carried out in patients to test the validity of this formula. PMID- 12126001 TI - Basaloid squamous cell carcinoma of the maxilla: a case report and immunohistochemical analysis. AB - Basaloid squamous cell carcinoma (BSCC) is a recently recognized high-grade tumor with a propensity for nodal as well as systemic metastasis and can arise from different anatomic locations. The differential diagnosis includes adenoid cystic carcinoma, small cell neuroendocrine carcinoma and squamous cell carcinoma. Monoclonal antibodies reactive with cytokeratin (34betaE12, AE3, pancytokeratin), as well as other cellular antigens (vimentin [VIM]; synaptophysin [SYNF]; chromogranin A [ChA]; neuron-specific enolase [NSE]; S-100, desmin, smooth-muscle actin [SMA]), were used in an immunoperoxidase method with paraffin-embedded tissue to phenotypically characterize a case with features of BSCC arising in the maxillary sinus. Neoplastic cells reacted with the high-molecular-weight cytokeratin antibody 34betaE12, as well as with other antikeratin antibodies, but failed to react with the antibodies VIM, desmin and SMA and showed variable immunoreactivity for NSE, SYNF and S-100. The staining pattern for NSE was diffuse and intense and reactivity for ChA was inconsistent. PMID- 12126002 TI - Lymphoepithelial carcinoma of the larynx. AB - Lymphoepithelial carcinoma is a very rare tumour of the larynx, with an exhaustive review of the literature having disclosed only 33 documented cases. The relationship between lymphoepithelial carcinoma of the larynx and Epstein Barr virus is still controversial. We describe one new case of this tumour involving the supraglottis. The patient was treated with supraglottic laryngectomy and left modified neck dissection. Three years and 4 months later, the right side of the neck was found positive for metastatic disease and a right modified neck dissection was performed. No evidence of disease was exhibited 4 years after the diagnosis of metastatic disease. The diagnostic problems and therapy associated with this rare tumour are discussed. PMID- 12126003 TI - Extracranial head and neck schwannomas: a review of 8 years experience. AB - Schwannomas of the head and neck are uncommon tumors that arise from cranial, peripheral or autonomic nerves. In this study we review a series of 52 cases of schwannoma originating in the head and neck region over an 8-year period. All the tumors were benign, with the exception of one malignant schwannoma. The age range of the patients studied was 13-76 years and there was a predilection for males. Twenty-five schwannomas occurred in the scalp, face and external ear canal, 9 in the oral or nasal cavity and 18 in the neck. Seven cases of neck schwannoma originating from the major nerve system were found in the parapharyngeal space, all of which were located in the post-styloid compartment. Cervical plexus schwannomas originated either in the peripheral nerves or in an unidentified area of the nervous system; seven tumors were found in the posterior triangle of the neck and two in the anterior triangle. Two of the tumors originating in the brachial plexus were located in the posterior neck and one in the anterior neck. Tumors originating in the vagus nerve or sympathetic chain were all located in the anterior triangle of the neck. PMID- 12126004 TI - Organ and voice preservation in advanced laryngeal cancer. PMID- 12126005 TI - Is elective neck treatment indicated in patients with squamous cell carcinoma of the maxillary sinus? PMID- 12126006 TI - Management of cancer of the parathyroid. PMID- 12126007 TI - Status of carotid resection in head and neck cancer. PMID- 12126008 TI - Prognostic value of Delphian lymph node metastasis from laryngeal and hypopharyngeal cancer. PMID- 12126009 TI - Cellular schwannoma in the oral mucosa. AB - A case of cellular schwannoma of the oral mucosa in a 34-year-old Japanese man is described. Cellular schwannoma commonly affects soft tissues such as the retroperitoneum and posterior mediastinum, and also bone, but is extremely rare in the oral region. To our knowledge, this is only the second report of oral cellular schwannoma. Histologically, the tumor parenchyma consisted of hypercellular spindle cells with nuclear and cytoplasmic pleomorphism and nuclear palisading resembling Antoni A-type conventional schwannoma, without evidence of Verocay bodies. These features were indicative of cellular schwannoma. Immunohistochemically, the tumor cells were positive for S-100, S-100alpha, S 100beta and vimentin, suggesting that they were of peripheral nervous origin. Furthermore, it is speculated that the tumor was intermediate between a benign and a malignant state, based on the histological features and positivity for S 100alpha. PMID- 12126010 TI - Dendritic cell influx differs between the subglottic and glottic mucosae during acute laryngotracheitis induced by a broad spectrum of stimuli. AB - Clinically, the subglottic and glottic mucosae may react differently, eg, during acute laryngotracheitis. In healthy rats, we showed previously that the composition of the mucosal immune system of the larynx also differs between these areas. Neutrophils, lymphocytes, and dendritic cells (DCs) are part of this mucosal immune system. In particular, DCs occupy a key function. They migrate into inflamed mucosae during the early phase of the immune response, which is normally characterized by an influx of neutrophils. Thus, they help to overcome the time lag between the innate and the adaptive immune responses. In the present study, the influx of DCs, neutrophils, and T lymphocytes into the subglottic and glottic mucosae of rats was examined at different time points after challenge with a broad spectrum of stimuli such as dead Moraxella catarrhalis, viable Bordetella pertussis, viable Sendai virus, and the soluble protein ovalbumin. The number of DCs increased rapidly after the application of the antigens. This increase was as rapid as the increase in neutrophils. Depending on the kind of antigen, their number in the mucosa increased up to 1,000 cells per 0.1 mm2 (Sendai virus). The comparison of different mucosal areas shows that an overwhelming number of immunocompetent cells entered the subglottic mucosa, whereas only a few cells migrated into the adjacent glottic mucosa. In conclusion, after inhalation of different kinds of antigens, the subset of immunocompetent cells investigated in this study entered the laryngeal mucosa in high numbers. The number of DCs entering the laryngeal mucosa was higher than the numbers of the other immune cells investigated. This finding underlines their function as first-line sentinels of the mucosal immune system of the larynx. The observation that the number of cells entering the laryngeal mucosa is location dependent indicates the ability of adjacent laryngeal regions to react differently. This is similar to the clinical observation of a selective subglottic reaction during acute laryngotracheitis. PMID- 12126011 TI - Morphology of the cricopharyngeal muscle in Zenker and control specimens. AB - The cricopharyngeal muscle (CPM) is essential for normal deglutition. Pharyngeal dysphagia commonly results from impaired or uncoordinated CPM dilation. Dysfunction of the CPM has also been implicated in the genesis of Zenker's (pharyngoesophageal) diverticulum. Despite the CPM's significance, little is understood about its morphology. We studied CPM biopsy specimens from 20 patients with Zenker's diverticulum and from 5 fresh cadaver patients with detailed histologic techniques to include fiber size and shape and adenosine triphosphatase, reduced nicotinamide adenine dinucleotide, trichrome, succinate dehydrogenase, cytochrome C oxidase, periodic acid-Schiff reaction, oil red O, acid phosphatase, Congo red, crystal violet, and monoadenylate deaminase stains. The normal CPM has unique morphological characteristics, with some myofibers having staining properties that are a hybrid between striated muscle and muscle spindle. The variable orientation of the muscle fibers is also different from that of most other striated musculature. Of the 20 Zenker CPM specimens, 4 specimens did not reveal any significant differences from controls (2 of which had insufficient amounts of tissue for complete analysis). In the remaining 16 specimens, several abnormalities existed, including excessive size variation (16/16), grouping of atrophic fibers (9/16), target or targetoid formations (4/16), cores (2/16), and ragged red fibers (2/16). The final pathological pattern of the 16 specimens was neurogenic in 7, myopathic in 4, and mixed (with neurogenic predominance) in the remaining 5. Two specimens contained significant lymphocytic inflammatory infiltrates. We conclude that the unique neuromuscular function of the CPM in deglutition is likely due to its fiber orientation and the hybrid nature of some of the myofibers. Morphological disturbances of the CPM impair its dilation and may account for the development of Zenker's diverticulum. This disturbance is most often due to progressive denervation of the CPM. PMID- 12126012 TI - Neuromuscular junction changes in aged rat thyroarytenoid muscle. AB - Dynamic remodeling of neuromuscular junction (NMJ) structure is postulated as a cause of age-related muscular atrophy. Direct study of NMJ morphology in laryngeal muscles is important to our understanding of age-related decrements in voice and swallowing. The morphology of NMJs was studied in a rat model to compare young and old specimens of thyroarytenoid muscle--a muscle critical to airway protection and phonation. Fluorescent, triple-label immunohistochemical analysis and confocal microscopic visualization were used to analyze the structure of NMJs. We found that laryngeal NMJs underwent significant changes that were similar to those observed after denervation. Specifically, the axon terminal area was significantly reduced, there were a number of postsynaptic acetylcholine receptor areas unoccupied by nerve terminals, and there was increased variability in end plate architecture in the old muscles. The results of this study increase our understanding of the age-related morphological changes in the larynx, and may serve as a baseline to test the effectiveness of future interventions. PMID- 12126014 TI - Immunohistochemical localization of subtypes of muscarinic receptors in human inferior turbinate mucosa. AB - The regulation of glandular secretions and vasomotor tone in human nasal mucosa implicates muscarinic receptors. There are 5 recognized classes (m1 through m5) of muscarinic receptor subtypes, and the aim of our study was to localize muscarinic receptor subtypes (m1 through m5) in human inferior turbinate mucosa by an immunohistochemical method. We found m1 and m2 receptors distributed on glands, arteries, veins, and epithelia; m4 receptors were found around arteries; and m5 receptors were identified on glands and arteries. We found m3 receptors to be the most extensively distributed on glands, arteries, and veins of all of the muscarinic receptor subtypes. The m3 receptor is probably important in the physiology of the human inferior turbinate. This study may help identify the best target for more selective muscarinic drugs and guide the treatment of allergic and nonallergic rhinitis. PMID- 12126013 TI - Induction of endolymphatic hydrops by directly infused monoclonal antibody against type II collagen CB11 peptide. AB - Autoimmune ear disease induced by type II collagen has been investigated by Yoo et al. In the present study, we investigated the effects of direct chronic infusion of type II collagen-specific monoclonal antibodies into guinea pig cochlea. Type II collagen fragment CB11 peptide-specific monoclonal antibodies (anti-CB11 Mabs) were infused directly into the scala tympani of guinea pigs with an Alzet mini-osmotic pump (anti-CB11 Mab group). As a control, normal mouse serum was infused by the same method (control group). To evaluate the auditory function, we recorded brain stem auditory-evoked potentials (BAEPs). In the anti CB11 Mab group, 80% of the animals showed an increased hearing threshold of more than 25 dB at 7 days after infusion. The hearing threshold shift observed in the guinea pigs of the control group was minimal (15 dB or less). To detect the structural changes, we performed histopathologic studies using hematoxylin and eosin staining. Inflammatory cell migration was detected mainly in the scala tympani of the guinea pigs of both groups. In the anti-CB11 Mab group, endolymphatic hydrops was also observed. The results of this experiment suggest that type II collagen autoimmunity is responsible for the production of hearing loss and endolymphatic hydrops. PMID- 12126015 TI - Otologic facial palsy: etiology, onset, and symptom duration. AB - To estimate the occurrence of otogenic facial palsy, we performed a retrospective case record study of all patients hospitalized for otogenic facial palsy in the period 1989 to 1999 at Ulleval University Hospital, which is the only referral hospital for patients with otologic sequelae in Oslo. The facial palsy was a complication of acute otitis media in 10 patients (56%), of acute mastoiditis in 3 patients (17%), of secretory otitis media in 3 patients (17%), and of chronic otitis media in 2 patients (11%). In half of the patients, complete facial palsy was found at the time of diagnosis. Sixteen patients (89%) reported a gradual onset of the facial palsy. The mean duration of otologic symptoms before the onset of facial palsy was 3 days (range, 1 to 9 days), and the median time to remission was 9 weeks (range, 2 to 96 weeks). Total remission was achieved in all patients who received follow-up. Although most patients recover within a few weeks, some patients have long-lasting facial palsy. Multicenter studies are needed to increase the sample size and to identify predictors of facial palsy duration. PMID- 12126016 TI - Digital photography of the larynx. AB - Digital photographic technology has ushered in a new era in medical documentation. The author's method of still digital photography for direct and indirect laryngeal photodocumentation using a single-lens-reflex camera and rigid rod-lens telescopes is described, and the results are presented. The advantages and disadvantages of different systems using still or video photography are discussed, and the importance of storing the archival image securely is emphasized. It is concluded that digital photography is convenient and versatile and gives good-quality images. It is the way of the future. PMID- 12126017 TI - Histopathologic study of the temporal bones and Eustachian tubes of children with cholesterol granuloma. AB - Six temporal bone-eustachian tube (ET) specimens with cholesterol granuloma (CG), obtained from 6 children 6 months to 15 years of age, were studied histopathologically to obtain further information about the pathogenesis of CG. We observed CG in the mastoid air cells in 5 ears, the mastoid antrum in 1 ear, the aditus ad antrum in 2 ears, and the epitympanum in 1 ear. All 6 cases exhibited a large amount of remaining mesenchyme that was in continuity with the hematopoietic bone marrow in the locations in which CG was present. All cases demonstrated otitis media with effusion and inflammation of the ET. Apparent morphological abnormalities of the ET and its associated structures, including hypoplastic ET cartilage and an abnormal tensor veli palatini muscle, were noted in 3 of the 6 cases. Furthermore, the posterior cartilaginous portion of the ET that includes its narrowest portion was completely filled with effusion in 2 of the 3 cases with the ET anomaly. The findings obtained were compared with data from age-matched control cases. Our results suggest that the source of erythrocytes in the remaining mesenchyme is the hematopoietic bone marrow. The pathogenesis of CG in children is likely promoted by ET dysfunction resulting in failure of ventilation of the middle ear. PMID- 12126018 TI - Pathophysiology of facial nerve paralysis induced by herpes simplex virus type 1 infection. AB - Herpes simplex virus type 1 (HSV-1) has been proven to be a cause of Bell's palsy; however, the underlying pathophysiology of the facial nerve paralysis is not fully understood. We established a mouse model with facial nerve paralysis induced by HSV-1 infection simulating Bell's palsy and investigated the pathophysiology of the facial nerve paralysis. The time course of the R1 latency in the blink reflex tests paralleled the recovery of the facial nerve paralysis well, whereas electroneurographic recovery tended to be delayed, compared to that of the paralysis; these responses are usually seen in Bell's palsy. On histopathologic analysis, intact, demyelinated, and degenerated nerves were intermingled in the facial nerve in the model. The similarity of the time course of facial nerve paralysis and the electrophysiological results in Bell's palsy and the model strongly suggest that the pathophysiological basis of Bell's palsy is a mixed lesion of various nerve injuries. PMID- 12126019 TI - Acoustic signature of the normal swallow: characterization by age, gender, and bolus volume. AB - Despite growing clinical use, cervical auscultation suffers from a lack of research-based data. One of the strongest criticisms of cervical auscultation is that there has been little research to demonstrate how dysphagic swallowing sounds are different from normal swallowing sounds. In order to answer this question, however, one first needs to document the acoustic characteristics of "normal," nondysphagic swallowing sounds. This article provides the first normative database of normal swallowing sounds for the adult population. The current investigation documents the acoustic characteristics of normal swallowing sounds for individuals from 18 to more than 60 years of age over a range of thin liquid volumes. Previous research has shown the normal swallow to be a dynamic event. The normal swallow is sensitive to aging of the oropharyngeal system, and also to the volume of bolus swallowed. The current investigation found that the acoustic signals generated during swallowing were sensitive to an individual's age and to the volume of the bolus swallowed. There were also some gender specific differences in the acoustic profile of the swallowing sound. It is anticipated that the results will provide a catalyst for further research into cervical auscultation. PMID- 12126020 TI - Exposure to 200 ppm of methanol increases the concentrations of interleukin-1beta and interleukin-8 in nasal secretions of healthy volunteers. AB - This study was designed to investigate subclinical irritating effects of methanol on functional and immunologic parameters in human respiratory epithelia. Twelve healthy, nonsmoking individuals were exposed to concentrations of 20 and 200 ppm of methanol in an exposure chamber. The concentrations of interleukin (IL)-8, IL 1beta, IL-6, and prostaglandin E2 (PGE2) were monitored. The saccharin transport time test was used to evaluate mucociliary transport. Video interference contrast microscopy was used to determine the ciliary beat frequency of nasal epithelial cells. Subjective symptoms were assessed with a questionnaire. The median concentrations of IL-8 and IL-1beta were significantly elevated after exposure to 200 ppm of methanol as compared to exposure to 20 ppm (IL-1beta, 21.4 versus 8.3 pg/mL, p = .001; IL-8, 424 versus 356 pg/mL, p = .02). The release of IL-6 and PGE2 did not change significantly (IL-6, 10.3 versus 6.5 pg/mL, p = .13; PGE2, 13.6 versus 13.4 pg/mL), nor did the ciliary beat frequency or the saccharin transport time. Both IL-8 and IL-1beta proved to be sensitive indicators for subclinical irritating effects of methanol in vivo. The German threshold limit of 200 ppm of methanol does not prevent subclinical inflammatory reactions of the nasal respiratory mucosa. PMID- 12126021 TI - Further delineation of the DFNA5 phenotype: results of speech recognition tests. AB - Speech recognition scores were analyzed in 34 carriers of a DFNA5 mutation. Cross sectional linear regression analysis (last visit, maximum recognition score in %Correct on age or PTA1,2,4 kHz) established onset age (score 90%) at 16 years and onset PTA1,2,4 kHz level (score 90%) at 41 dB hearing level. The deterioration rate was 0.7%/y in the plot of maximum score against age, whereas the deterioration gradient was 0.4%/dB in the plot of maximum score against PTA1,2,4 kHz. Given the previously demonstrated rapid progression of hearing impairment, speech recognition was relatively good: at age 70, the score was still >50%. PMID- 12126023 TI - Perilymphatic pressure measurement in Meniere's disease. AB - The aim of this study was to evaluate the perilymphatic pressure, by means of the MMS-10 Tympanic Displacement Analyzer (TDA), in patients with Meniere's disease (MD). Measurements were performed in 37 patients with MD and in 14 normal-hearing subjects. Data were collected from 3 groups: healthy ears of normal-hearing subjects, unaffected ears of patients with MD, and affected ears of patients with MD. Analysis of the results obtained with the TDA showed no significant differences between the 3 groups. Several hypotheses could explain this lack of difference: 1) perhaps indirect measurement of perilymphatic pressure with the TDA is not relevant; 2) perhaps hyperpressure of the inner ear is not the physiological basis for the clinical triad of MD; or 3) perhaps endolymphatic hydrops does not produce changes in perilymphatic pressure. The results of this series indicate that the TDA is not useful in the evaluation of patients with MD. PMID- 12126024 TI - Isolated congenital agenesis of the olfactory bulbs and tracts in a child without Kallmann's syndrome. PMID- 12126022 TI - Lemon-flavored cod liver oil and a multivitamin-mineral supplement for the secondary prevention of otitis media in young children: pilot research. AB - We measured blood levels of fatty acids, vitamin A, and trace metals in children undergoing ambulatory surgery for placement of tympanostomy tubes and a comparison group having other ambulatory surgical procedures. We then performed a small, outpatient, secondary prevention study using nutritional supplements chosen on the basis of those blood levels. The study subjects had lower levels of red blood cell eicosapentaenoic acid (EPA) than did adult controls. Consistent with previous reports, the levels of vitamin A were < or = 40 microg/dL for 69% of our subjects, and the plasma selenium levels for children were lower than published values for adults. We then studied one otitis media (OM) season; 8 children (0.8 to 4.4 years of age) received 1 teaspoon of lemon-flavored cod liver oil (containing both EPA and vitamin A) and 1 half-tablet of a selenium containing children's chewable multivitamin-mineral tablet per day. During this OM season, study subjects received antibiotics for OM for 12.3% +/- 13.4% (SD; p < .05) fewer days during supplementation than before supplementation. Larger, controlled trials are warranted to assess the utility of cod liver oil (of acceptable purity and taste) and a children's multivitamin-mineral preparation containing selenium, both for the prevention of OM and for the acceptance of delayed prescription of antibiotics for this disorder. PMID- 12126025 TI - Virtual endoscopy: a promising new technology. AB - Growing evidence shows that early detection of cancer can substantially reduce mortality, necessitating screening programs that encourage patient compliance. Radiology is already established as a screening tool, as in mammography for breast cancer and ultrasonography for congenital anomalies. Advanced processing of helical computed tomographic data sets permits three-dimensional and virtual endoscopic models. Such models are noninvasive and require minimal patient preparation, making them ideal for screening. Virtual endoscopy has been used to evaluate the colon, bronchi, stomach, blood vessels, bladder, kidney, larynx, and paranasal sinuses. The most promising role for virtual endoscopy is in screening patients for colorectal cancer. The technique has also been used to evaluate the tracheobronchial tree for bronchogenic carcinoma. Three-dimensional and virtual endoscopy can screen, diagnose, evaluate and assist determination of surgical approach, and provide surveillance of certain malignancies. PMID- 12126026 TI - Common bacterial skin infections. AB - Family physicians frequently treat bacterial skin infections in the office and in the hospital. Common skin infections include cellulitis, erysipelas, impetigo, folliculitis, and furuncles and carbuncles. Cellulitis is an infection of the dermis and subcutaneous tissue that has poorly demarcated borders and is usually caused by Streptococcus or Staphylococcus species. Erysipelas is a superficial form of cellulitis with sharply demarcated borders and is caused almost exclusively by Streptococcus. Impetigo is also caused by Streptococcus or Staphylococcus and can lead to lifting of the stratum corneum resulting in the commonly seen bullous effect. Folliculitis is an inflammation of the hair follicles. When the infection is bacterial rather than mechanical in nature, it is most commonly caused by Staphylococcus. If the infection of the follicle is deeper and involves more follicles, it moves into the furuncle and carbuncle stages and usually requires incision and drainage. All of these infections are typically diagnosed by clinical presentation and treated empirically. If antibiotics are required, one that is active against gram-positive organisms such as penicillinase-resistant penicillins, cephalosporins, macrolides, or fluoroquinolones should be chosen. Children, patients who have diabetes, or patients who have immunodeficiencies are more susceptible to gram-negative infections and may require treatment with a second- or third-generation cephalosporin. PMID- 12126027 TI - Alternative therapies for traditional disease states: menopause. AB - With growing concern about the use of hormone replacement therapy, some women are looking for alternative treatments for menopausal symptoms and preventing postmenopausal cardiovascular disease and osteoporosis. In observational trials, exercise has been associated with decreased vasomotor symptoms. One trial suggested that black cohosh may reduce menopausal symptoms. Soy has been shown to decrease vasomotor symptoms, lower lipid levels, and increase bone density. However, large amounts of soy must be consumed, and it is not clear whether soy consumption causes a decrease in cardiovascular events or fractures. The evidence for St. John's wort is equivocal. Fish oil is helpful for secondary prevention of coronary artery disease. PMID- 12126028 TI - Community-acquired pneumonia. PMID- 12126029 TI - Screening for chlamydial infection. PMID- 12126030 TI - Photo quiz. Red ear. PMID- 12126032 TI - Alternatives to rehydration during hypodermoclysis. PMID- 12126031 TI - NIH releases statement on osteoporosis prevention, diagnosis, and therapy. PMID- 12126033 TI - Managing chronic pain in the primary care setting. PMID- 12126035 TI - Information from your family doctor. BPH--a problem with your prostate. PMID- 12126034 TI - Managing benign prostatic hyperplasia. AB - Medical and surgical options for the treatment of benign prostatic hyperplasia have expanded in recent years. Saw palmetto, the most widely used complementary medication, is less effective than standard medical therapy but has fewer side effects. Although non-selective alpha blockers provide rapid relief of symptoms and are relatively inexpensive, they can cause dizziness and orthostatic hypotension. These effects occur less often with tamsulosin, a more selective alpha blocker. Finasteride, a 5alpha-reductase inhibitor, slowly reduces prostatic volume but is not as effective as alpha blockers, especially in men with a smaller prostate. Dutasteride, a new 5alpha-reductase inhibitor, has recently been labeled for the treatment of benign prostatic hyperplasia. Surgery may be appropriate initial treatment in patients with severe symptoms who are not at high risk for complications. Surgery may also be indicated in patients who have failed medical therapy or have recurrent infection, hematuria, or renal insufficiency. Transurethral resection of the prostate is effective in most patients, but it carries some risk of sexual dysfunction, incontinence, and bleeding. Surgical procedures that use thermal microwave or laser energy to reduce hyperplastic prostate tissue have recently become available. In general, the newer procedures are less expensive than transurethral resection of the prostate and have fewer complications; however, the need for retreatment is somewhat greater with these less invasive techniques. PMID- 12126036 TI - Principles of office anesthesia: part I. Infiltrative anesthesia. AB - The use of effective analgesia is vital for any office procedure in which pain may be inflicted. The ideal anesthetic achieves 100 percent analgesia in a short period of time, works on intact or nonintact skin without systemic side effects, and invokes neither pain nor toxicity. Because no single agent meets all of these criteria, the physician must choose from the available armamentarium based on the anesthetic properties that are most desired. Infiltrative anesthetics are frequently chosen because of their proven safety record, low cost, ease of storage, widespread availability, and rapid onset of action. Allergy to local injectable anesthetics is rare, and when it occurs it is often secondary to the preservative in multidose vials. Anesthesia can be prolonged with the addition of epinephrine or the use of longer-acting agents. Buffering the local anesthetic with bicarbonate, warming the solution, and injecting slowly can minimize the pain of anesthetic injection. Complications are rare but include central nervous system and cardiovascular toxicity, or extreme vasoconstriction in an end organ, if epinephrine is used. PMID- 12126037 TI - Principles of office anesthesia: part II. Topical anesthesia. AB - The development of topical anesthetics has provided the family physician with multiple options in anesthetizing open and intact skin. The combination of tetracaine, adrenaline (epinephrine), and cocaine, better known as TAC, was the first topical agent available for analgesia of lacerations to the face and scalp. Cocaine has been replaced with lidocaine in a newer formulation called LET (lidocaine, epinephrine, and tetracaine). For analgesia to nonintact skin, LET gel is generally preferred over TAC because of its superior safety record and cost-effectiveness. EMLA (eutectic mixture of local anesthetics) is perhaps the most well-known topical anesthetic for use on intact skin. EMLA can be used to anesthetize the skin before intramuscular injections, venipuncture, and simple skin procedures such as curettage or biopsy. To be fully effective, EMLA should be applied at least 90 minutes before the procedure. ELA-Max is a new, rapidly acting topical agent for intact skin that works by way of a liposomal delivery system and is available over the counter. Other delivery vehicles for topical anesthesia currently in development, including iontophoresis and anesthetic patches, may one day give patients and physicians even more flexibility. PMID- 12126039 TI - Smooth muscle changes in varicose veins: an ultrastructural study. AB - In order to understand the pathology of varicose veins, we prospectively collected a total of 23 vein specimens both from the normal proximal thigh long saphenous vein (LSV) in 3 young trauma patients and from the unstripped proximal LSV near the sapheno-femoral junction and the distal calf blowouts in 10 primary varicose veins patients. Ultra-thin sections were examined under the transmission electron microscope (TEM). Compared with the normal control LSV, varicose vein sections showed increase in the diameter of the lumen, hypertrophy of the wall and elongation and invagination of the intima. Smooth muscle cells (SMCS) lost their normal fusiform shape and were widely separated by increased amounts of extra-cellular collagen fibers. The cells underwent marked degeneration, vacuolization and disintegration into fiber-like material and small separated fragments. SMCs were seen in the subintimal tissue and some of them were lost into tile lumen. SMCs also showed marked phagocytic activity, engulfing not only collagen and elastic fibers, but also other smooth muscle cells. Although these changes were more marked and advanced in the distal calf blowouts, they were also present in the proximal, clinically non-dilated LSV. In conclusion, SMCs of varicose veins show severe degeneration in both the distal calf blowouts and the proximal, clinically non-varicose LSV. It appears that they both form and phagocytose collagen and elastic fibers and play a major role in the pathogenesis of varicose veins. PMID- 12126038 TI - Myosin light chain phosphorylation is correlated with cold-induced changes in platelet shape. AB - Chilling induces shape changes in platelets from disks to spheres with abundant filopodia. Such changes were time-dependent and correlated well with the phosphorylation of 20-kDa myosin light chain (LC20). Both the shape changes and the phosphorylation were reversible. After the platelets had been chilled, myosin became incorporated into the Triton X-insoluble fraction. When the chilled platelets were immunocytochemically stained, anti-myosin antibody was localized with filamentous structures inside the filopodia. These results suggest that LC20 phosphorylation and subsequent interactions with actin filaments play a crucial role in the cold-induced changes in platelet shape and in the formation of filopodia. PMID- 12126040 TI - Bone metabolic markers as gauges of metastasis to bone: a review. AB - Currently, imaging techniques are the leading methods used to diagnose of metastasis to bone. However, these techniques are expensive, expose patients to toxic and radioactive compounds, and monitor response to treatment poorly; these drawbacks have prompted the search for alternative screening methods. Therefore, bone metabolic markers have been evaluated as possible methods to diagnose and monitor the development and progression of metastatic bone disease. Although bone metabolic markers are often grouped as either resorption or formation markers, studies have revealed that each marker has its own biologic meaning and clinical relevance. Recent milestones in the use of bone metabolic markers as screening methods for metastatic bone disease and as evaluation methods for treatment response are shown in the following lists. 1. Bone metabolic marker measurements provide insight into mechanisms of metastasis to bone. 2. Although promising data have been reported, bone metabolic markers are not yet considered to be reliable screening methods for metastasis to bone. 3. Bone metabolic markers are reliable indicators of response to both conventional and bisphosphonate therapies. 4. Preliminary results indicate bone metabolic markers might be an independent prognostic factor in patients whose tumors metastasize to bone. 5. New or refined assays for bone metabolic markers are expected to improve the sensitivity and specificity of bone metabolic marker use in diagnosing and monitoring metastasis to bone. PMID- 12126041 TI - The usefulness of 18F-FDG PET images obtained 2 hours after intravenous injection in liver tumor. AB - Liver tumors, especially hepatocellular carcinomas (HCCs), often exhibit no contrast with surrounding non-tumorous liver tissue in F-18-fluoro-2-deoxy-2 fluoro-D-glucose (FDG) positron emission tomography (PET) images obtained at the usual interval of one hour after intravenous FDG injection. We evaluated the usefulness of FDG PET studies of liver tumors performed 2 hours after intravenous injection. METHODS AND MATERIALS: Fifteen pretherapeutic patients with 33 liver tumors were studied, including 11 patients with 18 HCCs, and 4 patients with 15 metastatic liver tumors (METAs) from 3 colorectal carcinomas and 1 esophageal carcinoma. After transmission scans, emission scans were obtained 45-55 minutes and 115-125 minutes after intravenous injection of 185-370 MBq FDG as early images and delayed FDG PET images, respectively. Visual analysis of early and delayed images was performed, and the FDG uptake in the tumor to that in nontumorous liver ratio (T/N ratio), the FDG uptake in tumor to that in soft tissue ratio (T/S ratio) and the FDG uptake in non-tumorous liver to that in soft tissue ratio (N/S ratio) were calculated for each image. RESULTS: In visual analysis, visual improvement seen in images was observed in 6 of 18 HCC lesions and all 15 META lesions. In quantitative analysis, the mean T/S ratio and T/N ratio of HCCs in early images were 4.97 and 1.90, respectively, and those in delayed images were 6.24 and 2.20, respectively. The mean T/S ratio and T/N ratio of METAs in early images were 5.97 and 2.21, respectively, and those in delayed images were 6.99 and 3.80, respectively. The T/S ratio of HCCs and T/S ratio and T/N ratio of METAs were significantly higher in delayed images than in early images. The mean N/S ratios of HCC cases were 2.58 in the early images and 2.57 in the delayed images, but the ratio showed no constant tendency in the images. All N/S ratios of META cases were decreased in delayed images, although the significance of the difference between early and delayed images in N/S ratios was not analyzed because of the small number of cases. CONCLUSION: FDG PET studies performed 2 hours after intravenous injection were useful for clear visualization of liver tumors, especially metastatic liver tumors. PMID- 12126042 TI - Gastric emptying in patients with chronic liver diseases. AB - There have been a number of reports of gastric emptying in cirrhosis, all with unconfirmed results. Moreover, the mechanism for delayed emptying in cirrhotic patients is unclear. We evaluated gastric emptying in patients with chronic hepatitis and cirrhosis by means of gastric emptying scintigraphy. METHODS: The subjects were 18 normal controls and 75 patients with chronic viral hepatitis (50 patients had chronic hepatitis and 25 patients had cirrhosis). Tc-99m diethyltriamine pentaacetic acid labeled solid meals were used to evaluate gastric emptying; the half-time (T 1/2) of which was calculated. Digestive symptom scores were determined at the time of gastric emptying tests. RESULTS: Fourteen (28%) of 50 patients with chronic hepatitis and 16 (64%) of 25 patients with cirrhosis had delayed gastric emptying. T 1/2 in patients with cirrhosis was significantly higher than that in normal controls and patients with chronic hepatitis (p = 0.0001 and 0.0003, respectively). The difference between T 1/2 in patients with chronic hepatitis and that in normal controls was not significant. On regression analysis, two indices-the serum albumin level and platelet count were found to be significantly related to delayed gastric emptying. CONCLUSIONS: Gastric emptying was more delayed in cirrhotic patients than in those with chronic hepatitis and normal controls. Delayed gastric emptying may be related to liver function and portal hypertension. PMID- 12126043 TI - Reversible defect of 123I-15-(p-iodophenyl)-9-(R,S)-methylpentadecanoic acid indicates residual viability within infarct-related area. AB - To evaluate the relationship between the reversible defect of 123I-15-(p iodophenyl)-9-(R,S)-methylpentadecanoic acid (9MPA) and residual viability within an infarct-related area, we performed resting single photon emission computed tomography (SPECT) with 9MPA and positron emission tomography (PET) with 18F deoxyglucose (FDG) and 13N-ammonia (NH3) in 7 patients with prior myocardial infarction. 9MPA-SPECT was obtained 2 min (early) and 50 min (delayed) after tracer injection. Tomographic images of the left ventricle were divided into 13 segments to correlate the regional uptake of each tracer. Residual viability within an infarct-related segment was confirmed by NH3- and FDG-PET. Twenty-six infarct-related segments, confirmed by NH3-PET, showed reduced uptake of 9MPA on early images. In these 26 segments, 6 showed reversible defect of 9MPA and 20 showed fixed defect on delayed images. Residual viability was present in all segments exhibiting reversible 9MPA defect and 7 segments (35%) exhibiting fixed defect (p < 0.05). The sensitivity, specificity and accuracy of reversible 9MPA defect for the detection of myocardial viability were 46%, 100%, and 73%, respectively. Myocardial clearance of 9MPA was significantly slower in non-viable segments than in ischemic but viable segments (4.9+/-5.1% vs. 10.1+/-5.3%; p < 0.05). These data suggest that a reversible 9MPA defect indicates residual viability within the infarct-related area. PMID- 12126044 TI - Evaluation of radioiodinated 5-iodo-3-(2(S)-azetidinylmethoxy)pyridine as a ligand for SPECT investigations of brain nicotinic acetylcholine receptors. AB - 5-Iodo-3-(2(S)-azetidinylmethoxy)pyridine (5IA), an A-85380 analog iodinated at the 5-position of the pyridine ring, was evaluated as a radiopharmaceutical for investigating brain nicotinic acethylcholine receptors (nAChRs) by single photon emission computed tomography (SPECT). [123/125I]5IA was synthesized by the iododestannylation reaction under no-carrier-added conditions and purified by high-performance liquid chromatography (HPLC) with high radiochemical yield (50%), high radiochemical purity (> 98%), and high specific radioactivity (> 55 GBq/micromol). The binding affinity of 5IA for brain nAChRs was measured in terms of displacement of [3H]cytisine and [125I]5IA from binding sites in rat cortical membranes. The binding data revealed that the affinity of 5IA was the same as that of A-85380 and more than seven fold higher than that of (-)-nicotine, and that 5IA bound selectively to the alpha4beta2 nAChR subtype. Biodistribution studies in rats indicated that the brain uptake of [125I]51A was rapid and profound. Regional cerebral distribution studies in rats demonstrated that the accumulation of [125I]5IA was consistent with the density of high affinity nAChRs with highest uptake observed in the nAChR-rich thalamus, moderate uptake in the cortex and lowest uptake in the cerebellum. Administration of the nAChR agonists (-)-cytisine and (-)-nicotine reduced the uptake of [125I]5IA in all regions studied with most pronounced reduction in the thalamus, and resulted in similar levels of radioactivity throughout the brain. [125I]5IA binding sites were shown to be saturable with unlabeled 5IA. Behavioral studies in mice demonstrated that 5IA did not show signs of behavioral toxicity. Furthermore, SPECT studies with [123I]5IA in the common marmoset demonstrated appropriate brain uptake and regional localization for a high-affinity nAChR imaging radiopharmaceutical. These results suggested that [123I]5IA is a promising radiopharmaceutical for SPECT studies of central nAChRs in human subjects. PMID- 12126045 TI - Comparison of brain perfusion SPECT and MRI findings in children with neuronal ceroid-lipofuscinosis and in their families. AB - PURPOSE: Neuronal ceroid-lipofuscinoses (NCL) are among the progressive encephalopathies of childhood that are inherited in an autosomal recessive manner. In this study we specifically aimed to investigate any white-matter changes in the carriers (parents) and the healthy siblings of individuals with neuronal ceroid lipofuscinosis disease and whether we may be able to predict the occurrence of any neurological symptoms in healthy children in the future thus enabling early management. MATERIALS AND METHODS: Since the NCLs are genetically determined diseases, we investigated fifteen individuals in three families that had diseased children of the juvenile type, with brain perfusion SPECT and MRI. Brain perfusion SPECT was performed after administering 222-555 MBq (6-15 mCi) Tc 99m HMPAO intravenously in a dimmed and quiet room. Imaging was performed at least one hour after injection, with a three headed gamma camera equipped with high resolution collimators. A Metz filter (FWHM: 11 mm) was used for processing. Cranial MRI was performed with an imager operating at 1.5 Tesla. Spin-echo T1- and T2-weighted and FLAIR slices were obtained for each individual. RESULTS: In all of the five diseased children we observed pathologic findings both on MRI and Tc-99m HMPAO SPECT. The findings on MRI were mainly features of cerebral and cerebellar atrophy and the observations on Tc-99m HMPAO SPECT were regional perfusion abnormalities. We observed some structural abnormalities on MRI in four of the parents and two of the four healthy siblings. We also noted perfusion abnormalities on Tc-99m HMPAO SPECT in two of the parents and two of the healthy siblings. CONCLUSION: Because the disease is inherited in an autosomal recessive manner, the parents and the healthy siblings were not supposed to exhibit any demonstrable brain lesions, but the brain perfusion SPECT and MRI examinations clearly revealed multiple lesions in some of the parents and healthy siblings. Detailed neurological examinations of these individuals were normal except for one apparently healthy sibling (EY). Follow-up imaging of these families is being undertaken and further studies are essential in understanding the pathogenesis and genetics of neuronal ceroid-lipofuscinoses. PMID- 12126046 TI - 11C-methionine uptake in cerebrovascular disease: a comparison with 18F-fDG PET and 99mTc-HMPAO SPECT. AB - OBJECTIVES: Carbon-11-L-methyl-methionine (11C-methionine) has been reported to be useful for evaluating brain tumors, but several other brain disorders have also shown signs of high methionine uptake. We retrospectively evaluated the significance of 11C-methionine uptake in cerebrovascular diseases, and also compared our results with those for 18F-FDG PET and 99mTc-HMPAO SPECT. METHODS: Seven patients, including 3 patients with a cerebral hematoma and 4 patients with a cerebral infarction, were examined. All 7 patients underwent both 11C methionine PET and 99mTc-HMPAO SPECT, and 6 of them underwent 18F-FDG PET. RESULTS: A high 11C-methionine uptake was observed in all 3 patients with cerebral hematoma. Increased 99mTc-HMPAO uptake was observed in 2 out of 3 patients, and all 3 patients had decreased 18F-FDG uptake. Of 4 patients with a cerebral infarction, high 11C-methionine uptake was observed in 3. Increased 99mTc-HMPAO uptake was also observed in one patient, whereas 3 patients had decreased 18F-FDG uptake. CONCLUSIONS: We should keep in mind that high 11C methionine uptake is frequently observed in cerebrovascular diseases. CVD should therefore be included in the differential diagnosis when encounting patients with a high 11C-methionine uptake. PMID- 12126047 TI - Biodistribution on Tc-99m labeled somatostatin receptor-binding peptide (Depreotide, NeoTec) planar and SPECT studies. AB - PURPOSE: To determine the biodistribution of Tc-99m labeled somatostatin receptor binding peptide (Depreotide) on planar and SPECT studies of the thorax and upper abdomen in order to improve diagnostic accuracy. METHODS AND MATERIALS: Retrospectively 29 planar and SPECT studies from 28 patients (all males, average age of 65.79) were reviewed. All the patients had been referred for evaluation of solitary pulmonary nodules. Two to four hours after IV injection of 555- to 740 MBq (15-20 mCi) Tc-99m Depreotide, anterior and posterior total body images, and anterior, posterior, right lateral and left lateral planar images were obtained, and thoracic SPECT was acquired with a three-head gamma camera. The degree of uptake in the lungs, thoracic cage, and organs of the upper abdomen was rated from "0" to "++++". RESULTS: The range of normal activity in the thorax includes cardiac, "0"; pulmonary, "+"; rib, "+/++"; sternum, "++"; vertebrae, "++". The degree of normal activity seen in the upper abdominal organs includes liver and spleen, "+++", and kidneys, "+++/++++". Eight patients with emphysema had diffuse pulmonary uptake graded as "+/++". One patient with left pneumonectomy and radiation therapy to the left hemithorax had photon-deficiency in the left hemithorax and decreased to absent uptake including the vertebrae and ribs. Although some cases had background pulmonary uptake of Tc-99m Depreotide, the bone/ bone marrow activity of the thoracic cage including the ribs, sternum, and thoracic spine is sufficiently great enough to produce a clear distinction between bone and lung in the thoracic cavity that gives high-contrast resolution on SPECT. CONCLUSIONS: The intensity of radioactivity in the sub-diaphragmatic organs such as the liver, spleen, and kidneys provides useful guidance for the categorization of pulmonary lesions. The uptake of land marks such as the sternum, which is anteriorly located, and the thoracic vertebrae, which are posteriorly located in the thoracic cage, can be used in the localization of a Depreotide avid tumor. PMID- 12126048 TI - Study on the usefulness of whole body SPECT coronal image, MIP image in 67Ga scintigraphy. AB - In this study, we examined the usefulness of whole body coronal images and whole body cine display MIP images (CMIP) upon which image processing was carried out after whole body SPECT in comparison to the usefulness of whole body images (WB/SC) compensated by scattered radiation in tumor/inflammation scintigraphy with 67Ga-citrate (67Ga). Image interpretation was performed for the 120 patients with confirmed diagnoses, and the accuracy of their diagnoses was studied by three nuclear medical physicians and two clinical radiological technologists by means of sensitivity, specificity and ROC analysis. The resultant data show that sensitivity, specificity, accuracy and the area under the ROC curve Az in the WB/SC were approximately 65%, 86%, 74% and 0.724, respectively, whereas sensitivity, specificity, accuracy and Az of the image reading system in which CMIP is combined with whole body coronal images reconstructed by the OS-EM method were approximately 93%, 95%, 94% and 0.860, respectively. Furthermore, coronal images reconstructed by the OS-EM method tended to be superior to those produced by the FBP method in both diagnostic accuracy and ROC analysis. In conclusion, the image reading system in which CMIP is combined with whole body coronal images reconstructed by the OS-EM method was shown to be superior in diagnostic accuracy and ROC analysis. Our data suggest that whole body SPECT is an excellent technique as an alternative to WB/SC. PMID- 12126049 TI - A case of gastric duplication evaluated by gastric emptying scintigraphy. AB - Gastric duplications are relatively rare, and communication with the gastric lumen is extremely rare. A 67-year-old man was referred to our hospital because of recurrence of epigastric pain and fullness. An upper gastrointestinal contrast study revealed a double compartment stomach, with gastric duplication starting at the esophagogastric junction outside the greater curvature. Computed tomography of the stomach with gastrografin as contrast demonstrated complete communication of the gastric duplication and primary stomach. The patient was diagnosed with complete gastric duplication. Gastric emptying scintigraphy with Tc-99m diethyltriamine pentaacetic acid was performed. Test meal entered the primary stomach and duplication cyst simultaneously, and radioactivity in the primary stomach decreased linearly and gastric emptying was not delayed. In the duplication cyst, about 70% of the food that entered the cyst once was immediately evacuated from it, but the remaining 30% remained in the cyst for a long time. Gastric emptying of the primary stomach was not affected by formation of the duplication cyst. PMID- 12126050 TI - 125I-iomazenil-benzodiazepine receptor binding during psychological stress in rats. AB - OBJECTIVE: We investigated the changes in 125I-iomazenil (125I-IMZ) benzodiazepine receptor (BZR) binding with psychological stress in a rat model. METHODS: Six male Wistar rats were placed under psychological stress for 1 hour by using a communication box. No physical stress was not received. 1.85 MBq of 125I-IMZ was injected into the lateral tail vein and the rat was killed 3 hours later. Twenty-micrometer-thick sections of the brain were collected and % injected dose per body weight (%ID/BW) of eleven regions (frontal, parietal, temporal, occipital cortices, caudate putamen, accumubens nuclei, globus pallidus, amygdala, thalamus, hippocampus and hypothalamus) were calculated by autoradiography. The %ID/BW of rats which were placed under psychological stress was compared with that of 6 control rats. RESULTS: The %ID/BW of rats which were placed under psychological stress diffusely tended to show a reduction in 125I IMZ-BZR binding. A significant decrease in BZR binding was observed in the hippocampus of the rats which were placed under psychological stress. CONCLUSION: 125I-IMZ-BZR binding tended to decrease throughout the brain. PMID- 12126051 TI - Diagnosis: pulmonary tumor thrombotic microangiopathy developing cor pulmonale. PMID- 12126052 TI - Distal end locus of the load axis of femora for two different periods in Japan. AB - The loci of the load axis at the femoral distal end were compared for specimens of Neolithic Jomon and modern periods, in which mean oblique length of the femur has been shown to differ significantly. In the present study, mean oblique length of the Neolithic Jomon exceeded that for the modern Japanese. However, mean distal end locus in either period was near the median of the lateral condyle within the bi-epicondylar width for both males and females. Differences in other morphological features of the femur were noted at the site of the condylo diaphyseal angle. Mean condylo-diaphyseal angle for the modern Japanese was greater than for the Neolithic Jomon. For the same period specimens, mean oblique length of males was significantly longer than in females but mean condylo diaphyseal angles and distal end loci were essentially the same. Distal end locus of the load axis within the width of the lateral condyle may be determined by the condylo-diaphyseal angle, regardless of oblique length. PMID- 12126054 TI - Comparative anatomical study of the m. retractor bulbi with special reference to the nerve innervations in rabbits and dogs. AB - Detailed dissection was performed on eight head halves of four rabbits and six head halves of three dogs in order to investigate the nerve distributions to the m. retractor bulbi. Our observations indicated that the muscle was innervated by the branches of the abducens nerve in all sides which were examined in the present study. In addition, we observed that the ventral ramus of the oculomotor nerve gave rise to branches to the m. retractor bulbi in three sides of the rabbits, and in two sides of the dogs, which suggests that the innervation pattern of the muscle is variable even in the identical species. Considering these observations, we propose that the anlage of the m. retractor bulbi is mainly composed of the anlage innervated by the abducens nerve, and occasionally the anlage innervated by the ventral ramus of the oculomotor nerve is added to it, because the anlage of the m. retractor bulbi may be formed near the border region between the anlages innervated by the oculomotor and abducens nerves. PMID- 12126053 TI - Expression of S-100 protein in intercalated duct cells of bovine pancreas. AB - Although Mutoh et al. have found intercalated ducts in the pancreatic islets of avians, including the chicken, moorhen (Gallinula chloorpus), and Japanese quail (Coturnix coturnix japonica), and even demonstrated the functions of intercalated duct cells in pancreatic islets, to our knowledge, there have been only a few reports on the relationship between intercalated ducts and islets in mammalia. Accordingly, in this study, we investigated whether intercalated ducts are morphologically related to the islets of cattle, by using S-100 protein as a ductal cell marker for immunocytochemistry and examining the ultrastructure of intercalated ducts and islets electron-microscopically. The results revealed intercalated ducts that reacted positively for S-100 protein within and near islets, with approximately 12% of the islets having intercalated ducts in the vicinity and approximately 1.5% containing intercalated ducts within them. Ultrastructurally, intercalated ducts were also seem to be closely related to the islets. In conclusion, the results of the present study indicate that a close relationship exists between intercalated ducts and islets. PMID- 12126055 TI - Morphology of accessory sex organs from neonatally DES- and DES-dp-injected mouse. AB - The neonatal estrogen induces morphological changes in accessory sex organs. We have reported that papillary proliferation in prostates and squamous metaplasia of the epithelium in seminal vesicle occurred and inflammatory cells have emigrated to the lumen through the stroma and the epithelium of organs from neonatal mice treated with beta-estradiol 17-cypionate. In this study, we observed the different effect between neonatal DES and DES-dp on morphological changes in accessory sex organs of mice. After 25 weeks, neonatal estrogen injections induced the infiltration of inflammatory cells in the ventral prostate and squamous metaplasia in the epithelium of seminal vesicles. It was observed that the inflammatory cells have already infiltrated into prostates from DES-dp injected mice after 5 weeks. But DES did not cause the changes in prostates. DES induced organs from a half of mice to involute and inflammatory cell to infiltrate into the epithelium. But these were not seen in organs from another half of mice. DES-dp occurred similar effect of beta-estradiol 17-cypionate on the male accessory sex organs. It remained to be seen whether DES could have estrogen action on accessory sex organ. PMID- 12126056 TI - Distributions of lesions in hanging suicide brains. AB - We found a morphological similarity in the distribution of vascular lesions in five hanging suicide brains. The overall findings on the lesions remind us of the venous origin but not of the arterial origin of the blood supply. Morphometric evaluations did not reveal any valuable conclusion. The results of this pathological research may be of clinical importance for the treatment of hanging patients. PMID- 12126057 TI - Morphometric analysis of the human femoral nerve and its ageing process. AB - We analysed the sizes of nerve fibres in the human femoral nerve which innervates the quadriceps femoris muscles. The material was taken from 14 cadavers aged from 61 to 97 years. A linear regression analysis disclosed a significant age-related decrease of the mean transverse area of axons. Such decrease with age may be an indication of motoneuron atrophy. Our results could help in the understanding of the correlation between morphology and function during the ageing process. PMID- 12126058 TI - Organization and distribution of the upper and lower esophageal motoneurons in the medulla and the spinal cord of the rat. AB - The upper cervical esophagus is exerted on swallowing and peristalsis by somatic and visceral motoneurons, whereas the lower esophagus is exerted on only peristalsis by visceral motoneurons. We examined the origin of the esophageal motoneurons and whether there were any differences between the distributions of the upper and the lower esophageal motoneurons in the medulla and the spinal cord using cholera toxin subunit b (CTb) as the retrograde tracer. Following injection of CTb into the cervical esophagus resulted in heavy labeling of the neurons in the nucleus ambiguus including the compact (AmC), semicompact (AmS) and loose (AmL) formations, and the medial column of lamina IX at the C1-C5 levels of the cervical spinal cord corresponding to the spinal accessory nucleus. A few labeled neurons were found in the inferior salivatory nucleus, the rostral division of the dorsal motor nucleus of the vagus (DMX), the accessory facial nucleus and the lateral column of lamina IX at the C2 and C3 levels. All these labeled neurons showed ChAT immunoreactivity. When CTb was injected into the cut end of the unilateral recurrent laryngeal nerve, many labeled neurons were found in the ipsilateral AmC, the AmL, and the bilateral medial column at the C1 and C2 levels. Following injection of CTb into the subdiaphragmatic esophagus resulted in heavy labeling of the neurons only in the AmC and the DMX. When CTb was injected into the sternomastoid muscle, many labeled neurons were found in the medullary reticular formation, the facial nucleus, the medial column at the C1 C3, C5 and C6 levels, and the lateral column at the C2, C3, C5 and C6 levels. Injections of a Fluoro-Gold into the cervical esophagus and a CTb into the sternomastoid muscle or the subdiaphragmatic esophagus in the same animal showed many double labeled neurons in the medial column of the accessory nucleus at the C1 and C2 levels, but no double labeled neurons in the AmC. These results indicated that the upper cervical esophagus is innervated by the visceral medullary vagal motoneurons as well as the somatic spinal accessory motoneurons. The lower esophagus is innervated only by the visceral medullary vagal motoneurons. PMID- 12126059 TI - Use of fluorescently labeled caspase inhibitors as affinity labels to detect activated caspases. AB - Activation of caspases is the key event of apoptosis and different approaches were developed to assay it. To detect their activation in situ, we applied fluorochrome labeled inhibitors of caspases (FLICA) as affinity labels of active centers of these enzymes. The FLICA ligands are fluorescein or sulforhodamine conjugated peptide-fluoromethyl ketones that covalently bind to enzymatic centers of caspases with 1:1 stoichiometry. The specificity of FLICA towards individual caspases is provided by the peptide sequence of amino acids. Exposure of live cells to FLICA results in uptake of these ligands and their binding to activated caspases; unbound FLICA is removed by cell rinse. Cells labeled with FLICA can be examined by fluorescence microscopy or subjected to quantitative analysis by cytometry. Intracellular binding sites of FLICA are consistent with known localization of caspases. Covalent binding of FLICA allowed us to identify the labeled proteins by immunoblotting: the proteins that bound individual FLICAs had molecular weight between 17 and 22 kDa, which corresponds to large subunits of the caspases. Detection of caspases activation by FLICA can be combined with other markers of apoptosis or cell cycle for multiparametric analysis. Because FLICA are caspase inhibitors they arrest the process of apoptosis preventing cell disintegration. The stathmo-apoptotic method was developed, therefore, that allows one to assay cumulative apoptotic index over long period of time and estimate the rate of cell entry into apoptosis for large cell populations. FLICA offers a rapid and convenient assay of caspases activation and can also be used to accurately estimate the incidence of apoptosis. PMID- 12126060 TI - Cyclooxygenase-2 and adenylate cyclase/protein kinase A signaling pathway enhances angiogenesis through induction of vascular endothelial growth factor in rat sponge implants. AB - Angiogenesis is reportedly enhanced by prostaglandins (PGs). In the present experiment, we tested whether or not COX-2 and adenylate cyclase/protein kinase A (AC/PKA)-dependent VEGF induction enhanced angiogenesis in this model. Angiogenesis was enhanced by topical injection of human recombinant basic fibroblast growth factor (bFGF). The enhanced angiogenesis by bFGF was inhibited by indomethacin or selective COX-2 inhibitors, NS398, nimesulide, and JTE-522. Topical daily injections of 8-bromo-cAMP enhanced angiogenesis in a dose dependent manner. Forskolin, an activator of AC, also facilitated angiogenesis in a dose-dependent manner, as did amrinone, an inhibitor of phosphodiesterase. VEGF induction was confirmed by the increased levels in the fluids in the sponge matrix after topical injection of 8-bromo-cAMP. Immunohistochemical investigation further revealed the VEGF-expressed cells in the sponge granulation tissues to be fibroblasts, and the intensity of positive reactions was enhanced by bFGF, 8 bromo-cAMP, forskolin, and amrinone. Angiogenesis was inhibited by indometacin or selective COX-2 inhibitors, NS-398, nimesulide, and JTE-522. In addition, angiogenesis without topical injections of the above compounds was also suppressed by SQ22,536, an inhibitor for AC. or H-89, an inhibitor for PKA, with concomitant reductions in VEGF levels. Daily topical injections of neutralizing antibody or anti-sense oligonucleotide against VEGF significantly suppressed angiogenesis. These results suggested that COX-2 and AC/PKA-dependent induction of VEGF certainly enhanced angiogenesis, and that pharmacological tools for controlling this signaling pathway may be able to facilitate the management of conditions involving angiogenesis. PMID- 12126061 TI - The role of CD98 in astrocytic neoplasms. AB - The high expression of CD98 was reported in some normal tissues, including blood brain barrier, activated lymphocytes, the basal layer of skin, proximal tubles of kidney, placenta, testis and a wide variety of tumors. The CD98 complex consists of an 80-85kD heavy chain (4F2hc/FRP-1) and a 40-45kD light chain. CD98hc, 4F2hc, and FRP-1 are the same glycosylated protein each other and define antigenicity of CD98. LAT1, the sodium-independent L-type amino acid transporter 1, has been identified as a light chain of the CD98 heterodimer from C6 glioma cells. LAT1 also corresponds to TA1, an oncofetal antigen that is expressed primarily in fetal tissues and cancer cells such as glioma cells. Increased LAT1 expression was found in various malignancies including human gliomas. Several studies implicated the important role of LAT1 and 4F2hc in malignant transformation and carcinogenesis. The LAT1-CD98 pathway may represent a unique therapeutic target for cancer intervention. PMID- 12126062 TI - Combined use of hyperthermia and irradiation cause antiproliferative activity and cell death to human esophageal cell carcinoma cells--mainly cell cycle examination. AB - In clinically hyperthermia and irradiation therapy for malignant neoplasms are known that they have antiproliferative activity and cell death (including apoptosis) inducing activity. However not only mechanisms of cell death induction but treatment effects of them still have been unclear. In this time we showed that cell cycles from G0/G1 phase to S-G2/M phase were delayed by hyperthermia and G2/M phase accumulation were caused immediately by irradiation. And we also demonstrated that the combination treatments of hyperthermia and irradiation induced synergistic antiproliferative effects and strong effects of cell death to human esophageal carcinoma cell lines. Although treatments of hyperthermia and irradiation were mild individually, combination treatment of hyperthermia and irradiation were useful for esophageal carcinoma treatment. PMID- 12126063 TI - A possible intermediate step during apoptotic execution. AB - Many proteases are known to be involved in apoptosis. Among them, interleukin 1beta converting enzyme (ICE) and its family proteases, which are called caspases, play critical roles in the execution stage of apoptosis. We previously reported that a proteasome-inhibitor, benzyloxycarbonyl Leu-Leu-leucinal (ZLLLal), induced apoptosis in MOLT-4 cells. In the present study, in order to analyze the detailed mechanism of ZLLLal-induced apoptosis, we examined the effect of a caspase-inhibitor, acetyl(Ac)-Tyr-Val-Ala-Asp-chloromethyl ketone (AcYVADcmk), on ZLLLal-induced apoptosis in the cells. Agarose gel electrophoresis revealed that low concentrations of AcYVADcmk efficiently suppressed apoptotic DNA fragmentation. However, the cells presented morphology different from normal, apoptotic or necrotic cells, although DNA fragmentation was suppressed. The same examination was performed on the cells with anti-Fas antibody-induced apoptosis, and the same results were obtained. Some cells with a similar morphology were found even without the caspase-inhibitor in the early stage of anti-Fas antibody-induced physiological apoptosis. In addition, apoptotic cascade was reactivated by washing out the caspase inhibitor from the DNA degradation-suppressed cells. Therefore, this newly found morphological feature shows the presence of a step prior to caspase activation in the cells, and this is the first report presenting the pre-caspase-activated step in the apoptotic cascade. PMID- 12126066 TI - Papillon-Lefevre syndrome: a case report. AB - We report the first diagnosed case of Papillon-Lefevre syndrome in Thailand. The patient is the youngest child of consanguinous parents, and she has had symmetrical hyperkeratotic plaques on both plantar surfaces since birth with a history of chronic gingivitis, periodontitis, and premature loss of primary dentition. The histologic study revealed compact hyperkeratosis with epidermal acanthosis. Radiologic studies of the skull were normal. The radiographic panoramic view of the oral cavity revealed generalized severe vertical and horizontal alveolar bone loss. The immunologic analysis of polymorphonuclear leukocyte phagocytic function by nitrobluetetrazolium test (NBT test) showed decreasing response to latex stimulation. Serum parathyroid hormone, calcium, phosphate, and alkaline phosphatase levels were within normal limits. The skin lesions were temporary relieved with topical keratolytic agents. The oral lesions were improved by the extraction of hopeless teeth and conventional periodontal treatments. PMID- 12126065 TI - Fas-Fas ligand system as a possible mediator of spermatogenic cell apoptosis in human maturation-arrested testes. AB - To elucidate the mechanism of maturation arrest, known as one of the male infertility, we addressed whether germ cell apoptosis occurs during maturation arrest, and if so, whether Fas and Fas ligand expressions are involved in the apoptosis. By electron microscopy and terminal deoxynucleotidyl transferase mediated dUTP-biotin nick end labeling (TUNEL), typical apoptotic features were frequently found around the spermatocytic stage in maturation arrest, compared to that in normal testes. When paraffin-embedded sections reacted with anti-Fas antiserum, staining for Fas was found in the plasma membranes of spermatocytes in the maturation-arrested testes, while no positive spermatogenic cells were seen in the normal testes. On the other hand, positive immunostaining for Fas ligand was restricted to Sertoli cells in the maturation-arrested testes as well as in the normal testes, although the intensity of staining for Fas ligand in normal testicular Sertoli cells was much weaker than that of maturation-arrested ones. Thus, these findings demonstrate that "maturation arrest" is characterized by frequent apoptosis of spermatocytes, and that Fas and Fas ligand staining are associated with a high frequency of apoptosis. PMID- 12126064 TI - Involvement of Fas/Fas ligand in the induction of apoptosis in chronic sialadenitis of minor salivary glands including Sjogren's syndrome. AB - The role of apoptosis and contribution of Fas/FasL systems in the pathogenesis of Sjogren's syndrome (SS) are still controversial. With serial sections, we explored apoptosis assessed by the dUTP nick end labeling (TUNEL) method and expression of Fas and FasL by immunohistochemistry, and compared their distribution in minor salivary gland (MSG) of SS and sialolithiasis (SIL) patient tissues. Fas and FasL were co-localized in ductular and acinar cells of SS and SIL TUNEL+ cells co-distributed with the Fas and FasL expressing cells in ductular and acinar cells of SS in the vicinity of lymphocytic infiltration, while not in those of SIL Moreover, to morphologically confirm apoptosis, we identified TUNEL-positive(+) cells in the MSGs of SS at the ultra structural level by applying an inversion method to paraffin-embedded sections stained by TUNEL method. Surprisingly, these cells did not show characteristic apoptotic figures although TUNEL products were deposited on the hyperchromatin of acinar and ductular cells. On the other hand, acinar and ductular cells of SIL included clusters of TUNEL+ apoptotic bodies as did those cells by phagocytosis or having fallen into the ductular lumen. These findings suggest that Fas and FasL expressed in ducts and acini of chronic sialadenitis in SS patients induce apoptosis, possibily in an autocrine and/or paracrine manner. PMID- 12126067 TI - Severe angioedema induced by angiotensin converting enzyme inhibitors: role of precipitating factors. AB - Angiotensin converting enzyme inhibitors like captopril, enalapril, lisinopril, trandopril and ramipril may rarely induce a life threatening angioedema. We present two cases of severe angioedema induced by enalapril and ramipril along with possible precipitating factors observed in these patients. The importance of prompt recognition and early management of such cases is emphasized. PMID- 12126068 TI - Wolf's isotopic response: a case of zosteriform lichen planus. AB - Lichen planus is a lichenoid disorder characterized by shiny, flat papules. In addition to the classical appearance, there are several variants. Zonal or zosteriform lesions have been described. A 25-year-old male with a complaint of increasing numbers of erythematous swellings on his left groin for twenty days was admitted to our out-patient clinic. He had a history of herpes zoster in the same localization which had been treated with topical acyclovir two weeks prior to his admission. Dermatological examination revealed multiple, shiny, erythematous, umblicated papules localized to the left inguinal region in a linear pattern. A biopsy was taken from the lesions. According to the clinical and pathological findings the diagnosis was zosteriform lichen planus. Zosteriform lichen planus is a rare variant of lichen planus; its differentiation from zona zoster and other linear dermatoses is difficult. We presented our case because of its rarity as a variant of lichen planus and its appearance in the area of healed herpes zoster as an isotopic response. PMID- 12126069 TI - Onion juice (Allium cepa L.), a new topical treatment for alopecia areata. AB - Alopecia areata is a patchy, non-scarring hair loss condition. Any hair-bearing surface may be involved, and different modalities of treatment have been used to induce hair regrowth. This study was designed to test the effectiveness of topical crude onion juice in the treatment of patchy alopecia areata in comparison with tap water. The patients were divided into two groups. The first group [onion juice treated] consisted of 23 patients, 16 males (69.5%) and 7 females (30.5%). Their ages ranged between 5-42 years with a mean of 22.7 years. The second group [control; tap-water-treated] consisted of 15 patients, 8 males (53.3%) and 7 females (46.6%). Their ages ranged between 3-35 years with a mean of 18.3 years. The two groups were advised to apply the treatment twice daily for two months. Re-growth of terminal coarse hairs started after two weeks of treatment with crude onion juice. At four weeks, hair re-growth was seen in 17 patients (73.9%), and, at six weeks, the hair re-growth was observed in 20 patients (86.9%) and was significantly higher among males (93.7%) compared to females (71.4%) P<0.0001. In the tap-water treated-control group, hair re-growth was apparent in only 2 patients (13%) at 8 weeks of treatment with no sex difference. The present study showed that the use of crude onion juice gave significantly higher results with regard to hair re-growth than did tap water (P<0.0001), and that it can be an effective topical therapy for patchy alopecia areata. PMID- 12126070 TI - Erythema annulare centrifugum following pancreatico-biliary surgery. AB - Annular erythemas are distinct cutaneous eruptions associated with a variety of causes. Occurrence of erythema annulare centrifugum, a variant of annular erythemas, in the immediate post-operative period in a patient with surgical intervention in the gall bladder and pancreas region is being described for the first time. Its probable association with surgical trauma is discussed. PMID- 12126071 TI - Segmentally distributed neurofibromatosis associated with adenocarcinoma of the colon. AB - Neurofibromatosis can be associated with various malignancies, but an association with adenocarcinoma is extremely rare. A 61-year-old man who had been diagnosed with adenocarcinoma of the hepatic flexure of the colon was referred for segmentally located, multiple cafe-au-lait spots and tumors on his back and axillary freckles for 40 years. Histopathologic examination of the tumor was consistent with the neurofibromatosis. We report an unusual case of type I neurofibromatosis associated with adenocarcinoma of the colon that was clinically similar to segmental neurofibromatosis. PMID- 12126072 TI - Diffuse cutaneous mastocytosis: pseudoxanthomatous variant. AB - Diffuse cutaneous mastocytosis is one of the extremely rare benign forms of mastocytosis that have varied clinical presentations. The pseudoxanthomatous variant is an extremely rare multinodular nonpigmented entity. Only two cases have been reported so far in the literature. We report a 21 year old female patient who had diffusely infiltrated skin with multiple papulo-nodular lesions resembling xanthomas and the classical histopathological features of mastocytosis. PMID- 12126073 TI - Malignant melanoma of the buttock presenting as a Buruli ulcer. AB - We report an unusual case of malignant melanoma clinically diagnosed as Buruli ulcer, that arose in a 13-year-old boy and presented as an ulcerated, fungating 2 cm mass on the right buttock. The tumor showed the histology and immunohistology of a malignant melanoma. We present this interesting case of malignant melanoma of soft tissue, arising in an unusual location of the body. The tumor presented with clinical features of Buruli ulcer in an area endemic for this disease as well as other tropical ulcerations. Neoplasms, including melanoma, should be considered in the differential diagnosis of Buruli ulcer in endemic areas. PMID- 12126074 TI - Colon carcinoma with synchronous subcutaneous and osseous metastasis: a case report. AB - Colon cancer usually metastasizes initially to regional lymphatics and later through the bloodstream. Hematogenous metastasis usually includes the liver, lungs, and brain. In colorectal cancer, osseous and/or subcutaneous metastasis without liver metastasis is a very uncommon event. We present here a case of colon adenocarcinoma, which synchronously metastasized to facial and other subcutaneous tissue and to bone within a short period after definitive therapy. Although such a pattern is uncommon, diagnostic biopsy for any new or suspicious lesion of the skin and bone scintigraphy for symptomatic patients should be done for patients with a colorectal cancer history. PMID- 12126075 TI - Perianal ulceration in a "healthy" Chinese man with disseminated tuberculosis. AB - Orificial tuberculosis (OTB) is a rare form of cutaneous Mycobacterium tuberculosis infection affecting the mucosa and skin around orifices in patients with advanced internal tuberculosis and poor general health. We report a 72-year old Chinese man who had a 10-year history of OTB with disseminated tuberculosis infection of the lungs and urinary tract. He appeared surprisingly healthy and had been free from systemic symptoms all along despite widespread tuberculosis. The diagnosis of OTB was established by the microscopic presence of acid-fast bacilli (AFB) in the tissue section and was rapidly confirmed by polymerase chain reaction (PCR) to be Mycobacterium tuberculosis. PCR shortens the time of diagnosing rare presentations of cutaneous tuberculosis and prevents delays in treatment. Conventional culture is still important in confirming the diagnosis and screening for drug resistance. PMID- 12126076 TI - Papular-purpuric gloves-and-socks syndrome with bloody bullae. AB - Papular-purpuric gloves-and-socks syndrome (PPGSS) is a disease characterized by itchy, painful acral erythema with edema, confluent papules, and purpura in a gloves-and-socks distribution and is associated with fever and mucosal lesions. Parvovirus B19 and other viral infections have been proven to be causative agents of this syndrome. Its histological findings have been the non-specific ones of interface dermatitis. Here, we report a case of PPGSS in a 44-year-old man that we believe to be the first such case in Japan. He developed, within one day, a painful edematous eruption with confluent papules and purpura on his hands and feet accompanied by high fever. A unique clinical manifestation in this case was multiple bloody bullae on the toes, which have not been previously described. Serological tests were negative for parvovirus B19, cytomegalovirus, and measles virus. PMID- 12126077 TI - Our experience with aplasia cutis congenita. AB - Aplasia cutis congenita is a rare disorder characterized by developmental absence of skin on the scalp as multiple or solitary, noninflammatory, well demarcated, oval or circular 1- to 2-cm ulcers. The disease may be isolated or associated with anomalies of the skin, eyes, ear-nose-neck and limbs, developmental defects of the cardiovascular, gastrointestinal, genitourinary and central nervous systems, and malformation syndromes such as chromosomal abnormalities, Adams Oliver syndrome, Bart's syndrome, and Johanson-Bilzzard syndrome. In this article, five newborn infants with aplasia cutis congenita (one associated with Adams-Oliver syndrome and another concomitant with Bart's syndrome) are reported because of their rare presentation in the literature. PMID- 12126078 TI - A case of multiple disseminated eruptive Spitz nevi. AB - Spitz nevus is usually a solitary lesion. However, multiple Spitz nevi do arise and can be divided into two variants, agminated (grouped) or disseminated. We report an interesting case of multiple disseminated eruptive Spitz nevi with an unusual feature. The appearance of most of the nevi resembled that of melanocytic nevi, and clinically it was impossible to distinguish these multiple disseminated eruptive Spitz nevi from eruptive melanocytic one. PMID- 12126079 TI - A case of pretibial mucinosis without thyroid disease. PMID- 12126080 TI - Acropigmentation of Dohi in an Indian family. PMID- 12126081 TI - A case of erythema type drug eruption due to domperidone. PMID- 12126082 TI - IL-12 in varicella zoster and herpes simplex virus infection. PMID- 12126083 TI - Optimal multi-phase three-dimensional fast imaging with steady-state free precession dynamic MRI and its clinical application to the diagnosis of hepatocellular carcinoma. AB - PURPOSE: To determine the appropriate acquisition parameters for three dimensional fast imaging with steady-state free precession (3D-FISP), to clarify the superiority of 3D-FISP to two-dimensional fast low-angle shot (2D-FLASH) on phantom study, and to clarify the clinical usefulness of 3D-FISP in diagnosing hepatocellular carcinoma (HCC). MATERIALS AND METHODS: 3D-FISP images with varying flip angles were compared by using a phantom. Signal-to-noise ratios (SNRs) and contrast-enhancement ratios (CERs) were compared for the four two dimensional fast low-angle shot (2D-FLASH) sequences and 3D-FISP sequences in a phantom. The optimal 3D-FISP dynamic study was compared with plain, postcontrast MR sequences used to study 78 HCC cases and analyzed according to histological grade. The 3D-FISP image obtained 30 sec after gadopentetate dimeglumine (Gd) administration was also compared with CT hepatic angiography (CTHA). RESULTS: A 25 degrees flip angle and double-dose Gd administration were appropriate for 3D FISP dynamic study. CER was the highest with 3D-FISP, and SNR was higher in 3D FISP than in 2D-FLASH images in a phantom with high Gd concentration. Among the 105 lesions, 103 (98%) were depicted on 3D-FISP images. The detection rate of HCC on 3D-FISP was higher than 95% for each histological grade. The vascularity of the tumors as determined by CTHA findings was correctly diagnosed on 3D-FISP in 80% of cases. CONCLUSION: In phantom study, 3D-FISP with double-dose Gd injection showed higher contrast than 2D-FLASH as a sequence for liver dynamic study. In clinical study, 3D-FISP is useful in the detection of HCC, regardless of tumor vascularity and histological grade. PMID- 12126084 TI - Loss of TRF2 by radiation-induced apoptosis in HL60 cells. AB - PURPOSE: A telomere consists of the short tandem DNA repeats of (T2AG3)n localized to the distal ends of chromosomes. Telomere repeat binding factor 2 (TRF2) has been implicated in the protection of chromosome ends. Recently, it has been reported that the loss of TRF2 induces apoptosis by various stimuli or genetic technique, however, the effects of radiation are not known. Therefore, this study investigated the interaction between TRF2 and radiation. MATERIALS AND METHODS: Western blot analysis, immunohistochemistry, and a DNA fragmentation assay for the detection of apoptosis were performed. The interaction between elastase and TRF2 was also investigated in vitro. RESULTS: Western blot analyses and immunohistochemistry showed that gamma-rays induce the temporary accumulation and subsequent loss of TRF2 protein in the nuclei of irradiated HL60 cells. Following DNA fragmentation, the loss of TRF2 could be detected. TRF2 was broken down by elastase, which was translocated into the nucleus before the loss of TRF2. CONCLUSIONS: The results of the study showed that irradiation first induces activation of TRF2, consequently protecting the end of the chromosome. Subsequently, translocation of elastase into the nucleus results in the breakdown of TRF2 after DNA fragmentation has occurred. PMID- 12126085 TI - Thoracolithiasis--a mobile calcified nodule in the intrathoracic space: radiographic, CT, and MRI findings. AB - We present a 74-year-old male patient with a mobile thoracolithiasis of 1x1 cm in diameter in the left intrathoracic space that was visualized by chest radiography and CT examinations. This calcified nodule was thought to have originated from a lipoma that had arisen from the pleura or the peripheral region of the lung, or from pericardial fat, then degenerated, dropped, and entered into the intrathoracic space, where it became calcified. The central core of this nodule showed high intensity on T1- and T2-weighted MR images, suggesting that the core was a soft tissue component, probably fat. PMID- 12126086 TI - A case of venous thoracic outlet syndrome recognized by arm swelling. AB - Most cases of thoracic outlet syndrome are detected by neulogical symptoms, and most of the other symptoms are caused by arterial stenosis. It is rare for the syndrome to be recognized by venous symptoms. We report a 56-year-old woman with thoracic outlet syndrome recognized by arm swelling. She was admitted for radiation therapy of a recurrent tumor of lung cancer at the left apex. Her right arm gradually became swollen. We performed venography from the right median cubital vein because of suspected venous thrombosis. Venography revealed stenosis of the right subclavian vein at the costoclavicular space, and this finding was confirmed by helical CT. These findings strongly support our diagnosis of thoracic outlet syndrome. PMID- 12126087 TI - Granular cell tumor of the spinal nerve root: MR findings. AB - We report granular cell tumor in the right SI nerve root, a location which has not been reported previously. The tumor showed heterogeneous hypointensity on T2 weighted images and heterogeneous enhancement. MRI also demonstrated the precise relation between the tumor and the nerve root. PMID- 12126089 TI - A case of Mycobacterium avium complex infection showing solitary pulmonary mass. AB - We report a case of Mycobacterium avium complex (MAC) infection showing a solitary pulmonary mass. High-resolution computed tomography (CT) revealed a well defined, lobulated mass (32x25 mm) without calcification, cavitation, or bronchiectasis on the right upper lobe. There were no abnormalities in other sites of the lung even on high-resolution CT. The diagnosis was established by open lung needle biopsy. The patient exhibited a large solitary pulmonary mass caused by MAC infection, suggesting that, although the incidence is rare, MAC infection can show a solitary mass that is radiographically indistinguishable from lung cancer. PMID- 12126088 TI - A case of malignant fibrous histiocytoma (MFH) in the skull bone presenting extensive permeative osteolysis. AB - Malignant fibrous histiocytoma (MFH) of the skull is rare. This case showed an extremely unusual growth pattern for MFH. The radiographs and magnetic resonance imaging (MRI) demonstrated an osteolytic growth pattern in the skull base and temporal bone, without forming a mass lesion. We considered the possibility of a malignant tumor with a myxoid component based on its growth pattern and MRI intensity. Pathological examination revealed dense proliferation of the neoplastic spindle cells with storiform pattern and focal myxoid or fibrotic areas. Only one other case of MFH with myxoid change of the skull bone has been documented in the English literature. PMID- 12126090 TI - Bronchiolitis obliterans organizing pneumonia after tangential beam irradiation to the breast: discrimination from radiation pneumonitis. AB - We report a case of bronchiolitis obliterans organizing pneumonia (BOOP) secondary to tangential beam irradiation to the breast, which occurred seven months after the completion of radiotherapy. Although radiation pneumonitis is an alternative consideration, BOOP could be differentiated from it by its relatively late onset and extensive distribution, which did not respect the radiation field. This disease should always be kept in mind in patients with a history of tangential beam irradiation to the breast. PMID- 12126091 TI - A case of metastatic bone tumor of missed lung cancer showing unusual manifestations on chest radiogram. AB - We encountered a patient who had a primary unknown carcinoma with lumbago caused by lumbar vertebral metastasis as the initial chief complaint. Autopsy revealed primary pulmonary adenocarcinoma. Abnormal findings noted on plain chest radiogram and chest CT led to misdiagnosis as inflammatory changes. The primary lesion was poorly differentiated pulmonary adenocarcinoma that could not be diagnosed correctly using the diagnostic criteria of these diagnostic modalities. PMID- 12126092 TI - Assessment of myocardial washout of Tc-99m-sestamibi in patients with chronic heart failure: comparison with normal control. AB - BACKGROUND: In contrast to 201TlCl, 99mTc-sestamibi shows very slow myocardial clearance after its initial myocardial uptake. In the present study, myocardial washout of 99mTc-sestamibi was calculated in patients with non-ischemic chronic heart failure (CHF) and compared with biventricular parameters obtained from first-pass and ECG-gated myocardial perfusion SPECT data. METHODS AND RESULTS: After administration of 99mTc-sestamibi, 25 patients with CHF and 8 normal controls (NC) were examined by ECG-gated myocardial perfusion SPECT and planar data acquisition in the early and delayed (interval of 3 hours) phase. Left ventricular ejection fraction (LVEF, %), peak filling rate (PFR, sec(-1)), end diastolic volume (LVEDV, ml) and end-systolic volume (LVESV, ml) were automatically calculated from the ECG-gated SPECT data. Myocardial washout rates over 3 hours were calculated from the early and delayed planar images. Myocardial washout rates in the CHF group (39.6+/-5.2%) were significantly higher than those in the NC group (31.2+/-5.5%, p < 0.01). The myocardial washout rates for the 33 subjects showed significant correlations with LVEF (r = -0.61, p < 0.001), PFR (r = -0.47, p < 0.01), LVEDV (r = 0.45, p < 0.01) and LVESV (r = 0.48, p < 0.01). CONCLUSION: The myocardial washout rate of 99Tc-sestamibi is considered to be a novel marker for the diagnosis of myocardial damage in patients with chronic heart failure. PMID- 12126093 TI - Technetium-99m human immunoglobulin scintigraphy in patients with adhesive capsulitis: a correlative study with bone scintigraphy. AB - Adhesive capsulitis (AC) is a disorder that is characterized by shoulder pain and progressive limitation of both active and passive shoulder motion. Although the underlying pathological mechanisms of the disease are not well understood, the inflammatory reactions depending on the stage have been demonstrated histologically. The purpose of the study is to investigate the inflammatory changes that can be demonstrated with Tc-99m HIG in AC, and to determine the presence of correlations between scintigraphic findings and the clinical assessment. Twenty-one patients (12 females and 9 males) with a mean age of 50.57+/-8.49 were included in the study. AC was diagnosed according to recognized criteria. The planar X-ray images of the affected shoulders of all patients were normal. The patients were evaluated with the Constant Scoring System, and the functional and pain assessment parts of the American Shoulder and Elbow Surgeons' Form (ASES). Three phase bone scans and Tc-99m HIG scintigraphy were performed at least two days apart. Bone scan and Tc-99m HIG scintigraphy were evaluated visually and HIG uptake was evaluated in comparison with the contralateral normal shoulder. Bone scan demonstrated hypervascularity in 9 of the 21 patients (43%), whereas increased osteoblastic activity was detected in 19 (90%) in the affected shoulder. Tc-99m HIG uptake was positive in 12 (57%), and negative in 9 (43%) patients. All patients with increased Tc-99m HIG accumulation in the affected shoulder, also had increased osteoblastic activity on Tc-99m bone scintigraphy. A significant correlation was found between HIG uptake and constant, functional and pain scores. The difference between these scores was also statistically significant in patients with HIG positive and negative uptake. This study indicates that there is a good correlation between Tc-99m HIG scan findings and clinical scores. Tc-99m HIG accumulation in the affected shoulder may be related to continuing inflammatory reaction to AC. Tc-99m HIG scan may be a noninvasive, complementary method for demonstrating continuing inflammatory changes and may help in staging the disease. PMID- 12126094 TI - Hemodynamic changes during neural deactivation in human brain: a positron emission tomography study of crossed cerebellar diaschisis. AB - The mechanism of crossed cerebellar diaschisis (CCD) is considered to be secondary hypoperfusion due to neural deactivation. To elucidate the hemodynamics during neural deactivation, the hemodynamics of CCD was investigated. The cerebral blood flow (CBF), cerebral blood volume (CBV), cerebral oxygen extraction fraction (OEF), cerebral metabolic rate of oxygen (CMRO2), and vascular responses to hypercapnia and acetazolamide stress for CCD were measured in 20 patients with cerebrovascular disease by positron emission tomography with H2(15O), C15O, and 15O2. Vascular responses to hypercapnia and acetazolamide stress were almost the same between CCD side and unaffected side of the cerebellum, a finding that supports the idea that the mechanism of CCD is secondary hypoperfusion due to neural deactivation. The degree of decrease in CBF on the CCD side was almost the same as that in CBV, indicating that vascular blood velocity does not change during neural deactivation. The relation between CBF and CBV of the CCD and unaffected sides was CBV = 0.29 CBF0.56. On the CCD side, the degree of deerease in CMRO2 was less than that in CBF, resulting in a significantly increased OEF. The increased OEF along with the decreased CBV on the CCD side might indicate that neural deactivation primarily causes vasoconstriction rather than a reduction of oxygen metabolism. PMID- 12126095 TI - Correction of nonuniform attenuation and image fusion in SPECT imaging by means of separate X-ray CT. AB - Improvements in image quality and quantitation measurement, and the addition of detailed anatomical structures are important topics for single-photon emission tomography (SPECT). The goal of this study was to develop a practical system enabling both nonuniform attenuation correction and image fusion of SPECT images by means of high-performance X-ray computed tomography (CT). A SPECT system and a helical X-ray CT system were placed next to each other and linked with Ethernet. To avoid positional differences between the SPECT and X-ray CT studies, identical flat patient tables were used for both scans; body distortion was minimized with laser beams from the upper and lateral directions to detect the position of the skin surface. For the raw projection data of SPECT, a scatter correction was performed with the triple energy window method. Image fusion of the X-ray CT and SPECT images was performed automatically by auto-registration of fiducial markers attached to the skin surface. After registration of the X-ray CT and SPECT images, an X-ray CT-derived attenuation map was created with the calibration curve for 99mTc. The SPECT images were then reconstructed with scatter and attenuation correction by means of a maximum likelihood expectation maximization algorithm. This system was evaluated in torso and cylindlical phantoms and in 4 patients referred for myocardial SPECT imaging with Tc-99m tetrofosmin. In the torso phantom study, the SPECT and X-ray CT images overlapped exactly on the computer display. After scatter and attenuation correction, the artifactual activity reduction in the inferior wall of the myocardium improved. Conversely, the incresed activity around the torso surface and the lungs was reduced. In the abdomen, the liver activity, which was originally uniform, had recovered after scatter and attenuation correction processing. The clinical study also showed good overlapping of cardiac and skin surface outlines on the fused SPECT and X ray CT images. The effectiveness of the scatter and attenuation correction process was similar to that observed in the phantom study. Because the total time required for computer processing was less than 10 minutes, this method of attenuation correction and image fusion for SPECT images is expected to become popular in clinical practice. PMID- 12126096 TI - Decision-tree sensitivity analysis for cost-effectiveness of whole-body FDG PET in the management of patients with non-small-cell lung carcinoma in Japan. AB - BACKGROUND: Whole-body 2-fluoro-2-D-[18F]deoxyglucose [FDG] positron emission tomography (WB-PET) may be more cost-effective than chest PET because WB-PET does not require conventional imaging (CI) for extrathoracic staging. METHODS: The cost-effectiveness of WB-PET for the management of Japanese patients with non small-cell lung carcinoma (NSCLC) was assessed. Decision-tree sensitivity analysis was designed, based on the two competing strategies of WB-PET vs. CI. WB PET was assumed to have a sensitivity and specificity for detecting metastases, of 90% to 100% and CI of 80% to 90%. The prevalences of M1 disease were 34% and 20%. One thousand patients suspected of having NSCLC were simulated in each strategy. We surveyed the relevant literature for the choice of variables. Expected cost saving (CS) and expected life expectancy (LE) for NSCLC patients were calculated. RESULTS: The WB-PET strategy yielded an expected CS of $951 US to $1,493US per patient and an expected LE of minus 0.0246 years to minus 0.0136 years per patient for the 71.4% NSCLC and 34% M1 disease prevalence at our hospital. PET avoided unnecessary bronchoscopies and thoracotomies for incurable and benign diseases. Overall, the CS for each patient was $833US to $2,010US at NSCLC prevalences ranging from 10% to 90%. The LE of the WB-PET strategy was similar to that of the CI strategy. The CS and LE minimally varied in the two situations of 34% and 20% M1 disease prevalence. CONCLUSIONS: The introduction of a WB-PET strategy in place of CI for managing NSCLC patients is potentially cost effective in Japan. PMID- 12126097 TI - Changes in regional cerebral blood flow in irradiated regions and normal brain after stereotactic radiosurgery. AB - OBJECTIVE: To elucidate the radiation effect on the normal brain after stereotactic radiosurgery (SRS), we evaluated the change in regional cerebral blood flow (CBF) in targeted and extra-targeted areas according to the radiation dose given. METHODS: Thirteen patients who underwent SRS for brain tumors or arteriovenous malformations were included in this study. Maximum radiation doses to the lesion ranged from 24 to 37 Gy. Mean and regional CBF were measured by 99mTc-HMPAO scintigraphy with graphic analysis, performed at before, 2 weeks and 3 months (5 patients) after SRS. Under the co-registration with the CT with superimposed isodose distribution, ROIs were set on target (37-20 Gy), peri target (20-5 Gy) and out-of-field (5-2 Gy and less than 2 Gy) areas on the quantitative SPECT images. RESULTS: Significant reductions in mean CBF (by 7%) and regional CBF in the peri-target areas (by 5-7%) and out-of-field areas (by 6 22%) were recognized at 2 weeks and 3 months after SRS. Regional CBF in the target and peri-target areas did not significantly change, presumably because there was little or no normal tissue in these areas. CONCLUSION: These results suggest that subclinical regional CBF reduction occurs after SRS in the normal brain in out-of-field of radiation. PMID- 12126099 TI - Scintigraphic findings of MALT lymphoma of the thyroid. AB - Mucosa-associated lymphoid tissue (MALT) lymphoma has been established as a distinct entity among non-Hodgkin's lymphomas, and the most common primary site is the stomach. We describe scintigraphic findings in a patient with MALT lymphoma of the thyroid. A 71-year-old woman with Hashimoto's thyroiditis suffered from rapid cervical swelling, and ultrasonography and CT revealed a thyroid nodule. The nodule showed accumulation of 99mTc pertechnetate comparable to the surrounding thyroid tissue, mimicking a benign nodule. Both 67Ga and 201Tl imaging visualized the lesion as an increased uptake area. After radiotherapy, abnormally increased uptake disappeared on 67Ga images, which predicted a favorable outcome. MALT lymphoma of the thyroid may be visualized as a warm nodule on 99mTc pertechnetate scintigraphy. PMID- 12126098 TI - Causes of appearance of scintigraphic hot areas on thyroid scintigraphy analyzed with clinical features and comparative ultrasonographic findings. AB - This study was done retrospectively to analyze the ultrasonographic (US) findings in thyroid scintigraphic hot areas (HA). Three-thousand, eight-hundred and thirty nine consecutive patients who underwent 99mTc-pertechnetate (n = 3435) or 123I (n = 457) scintigraphy were analyzed. HA were regarded as present when the tracer concentration was greater than the remaining thyroid tissue, or when hemilobar uptake was observed. High-resolution US examinations were performed with a real time electronic linear scanner with a 7.5 or 10 MHz transducer. One hundred and four (2.7%) were found to be scintigraphic HA (n = 120). US revealed a nodular lesion or well-demarcated thyroid tissue corresponding to the HA in 94 areas (78.4%, Category 1), an ill-defined region with different echogenicity in 13 areas (10.8%, Category 2), and no correlating lesion in 13 areas (10.8%, Category 3). These 104 patients included 43 with adenomatous goiter (59 areas), 33 with adenoma, 11 with Hashimoto's thyroiditis, 5 with primary thyroid cancer, 4 with euthyroid ophthalmic Graves' disease (EOG), 3 with hemilobar atrophy or hypogenesis, 2 with hemilobar agenesis, 2 with hypothyroidism with blocking-type TSH-receptor antibodies (TSHRAb), I with acute suppurative thyroiditis. Among the 59 adenomatous nodules and 33 adenomas, 51 (86.4%) and 32 (97.0%), respectively, belonged to Category 1. A solitary toxic nodule was significantly larger and occurs more often in older patients than in younger patients. On the other hand, all 17 patients with known autoimmune thyroid diseases including Hashimoto's thyroiditis, EOG and hypothyroidism with blocking TSHRAb belonged to Category 2 or 3. Possible underlying mechanisms are 1) hyperfunctioning tumors or nodules, 2) localized functioning thyroid tissue freed from autoimmune destruction, inflammation or tumor invasion, 3) congenital abnormality, 4) clusters of hyperactive follicular cells caused by long-term TSH and/or TSHRAb stimulation, 5) asymmetry, etc. Scintigraphic HA are observed in patients with various thyroid diseases and high-resolution US appears to be helpful clinically for the differential diagnosis of the above mentioned disorders. PMID- 12126100 TI - Tc-99m RBC perfusion and blood-pool scintigraphy in the evaluation of vascular leiomyoma of the hand. AB - A 62-year-old man with a soft, non-tender, movable mass 2.5 x 2.5-cm in diameter in the volar surface of the right index finger over the proximal phalanx underwent Tc-99m RBC perfusion and blood-pool scintigraphy to evaluate the vascular nature and extent of the mass. Highly increased activity on early and delayed blood-pool images with increased perfusion was demonstrated in the mass. The lesion with high flow rates and large blood pool spaces was considered highly suggestive of one of the various types of peripheral hemangioma. Angiography revealed a vascular neoplasm with tumor vessels. Microscopic examination of the resected tumor revealed vascular leiomyoma containing numerous dilated vascular channels. These scintigraphic abnormalities were regarded as resulting from hypervascularity demonstrated angiographically and blood pooling within the dilated vascular channels demonstrated histologically. It is concluded that Tc 99m RBC perfusion and blood pool scintigraphy may be an important non-invasive approach to demonstrate vascular leiomyoma prior to surgical biopsy or resection. PMID- 12126101 TI - A case of non-Hodgkin's lymphoma infiltrating the brachial plexus detected by Ga 67 scintigraphy. AB - The author reports a rare case of brachial plexus infiltration by non-Hodgkin's lymphoma. T2-weighted MRI showed high signal intensity along the right brachial plexus and soft tissue masses. Ga-67 scintigraphy showed abnormal tracer uptake along the course of the right brachial plexus, and was superior to MRI in detecting the abnormality. PMID- 12126102 TI - Variation in FDG uptakes in different regions in normal human brain as a function of the time (30 and 60 minutes) after injection of FDG. AB - OBJECTIVE: The authors' goal was to determine whether FDG uptakes in various regions of the brain are different for early and late scanning time in positron emission tomography (PET). METHOD: F-18 fluorodeoxyglucose (FDG) PET was performed on 15 healthy normal subjects to obtain early and late acquisition glucose metabolic images (30 and 60 min after FDG injection), respectively. The two sets of images were compared in a voxel-by-voxel analysis. RESULTS: In the bilateral posterior cingulate gyrus, parietal and frontal association cortices, and subcallosal cortices, the FDG uptakes were larger on the late scan image than on the early scan image, and the FDG uptakes were larger in the cerebellar hemisphere, vermis and frontal basis on the early scan image than on the late scan image. CONCLUSIONS: These results suggest that there are different regional FDG uptakes depending on the scanning time after FDG injection and we must be careful in replacing conventional FDG PET scanning with early scanning in FDG PET study. PMID- 12126103 TI - I-123 MIBG scintigraphy. Pheochromocytoma. PMID- 12126104 TI - Adenylate kinase from plant tissues. Influence of ribonuclease on binding properties on Mono Q. AB - Adenylate kinases modulate the three adenine nucleotide pools and were found to be localized as isoenzymes in different tissues and organelles in animals and plants. For investigations of adenylate kinase isoenzymes from plant tissues different plant extracts were examined by anion-exchange chromatography. During investigations with the strong anion exchanger Mono Q, adenylate kinase activity eluted in the void volume. This void volume activity did not always occur, but depended on the age of the plants and light treatment. The nature of the factors affecting void volume activity could only be partially resolved. It could be shown that RNase treatment at the beginning of extraction led to the disappearance of void volume activity, whereas an untreated extract still showed this activity. PMID- 12126105 TI - Immobilized metal affinity chromatography for the separation of photosystems I and II from the thermophilic cyanobacterium Synechococcus elongatus. AB - Immobilized metal affinity chromatography (IMAC) of solubilized, photosystem II (PS II) enriched particles from the thermophilic cyanobacterium Synechococcus elongatus was studied. A chelating Sepharose Fast Flow column was charged with various metal ions (Mn2+, Fe2+, Fe3+, Ni2+, Co2+, Ca2+, Sr2+, Zn2+ and Cu2+) and their affinity to photosystem I (PS I) and PS II was examined. Among all the metal ions tested, only copper was able to bind the two protein complexes. For elution of the column, a pH gradient, a pH step gradient and gradients of imidazole, amino acids, organic acids and various other eluents were tested; only the pH step gradient, which selectively eluted PS II at a pH between 6 and 5, was useful for the separation of PS I and PS II. All other gradients proved to be inappropriate for the separation of these two photosystems. Mechanisms of protein elution by these compounds are discussed. Alternatively, a separation of PS I and PS II at pH 7.5 could be achieved when an IMAC column was used on which the free coordination positions of the bound copper ions were occupied by imidazole. When solubilized photosystems were loaded on to this column, PS I replaced imidazole and remained bound on the column, whereas PS II was highly enriched in the effluent. PMID- 12126106 TI - Application of cross-flow filtration to the purification of biologically active peptides in human plasma after incubation with a protease-rich extract. AB - The aim of this study was to find an experimental procedure to purify biologically active peptides from a complex biological matrix (plasma), which was incubated with a protease-rich extract (submandibular gland extract). Special interest was focused on the practicability of cross-flow filtration for this purpose. Therefore, peptides in the incubation mixture were purified with a combination of high-performance liquid chromatographic steps. Purification of biologically active peptides was monitored by a sensitive bioassay and by laser desorption/ionization mass spectrometry. This permitted not only purity control at each purification step but also identification of one of the peptides with vasoconstrictor properties as angiotensin II. This result demonstrates the practicability of cross-flow filtration for extracting enzymatic reaction products from complex substrate-enzyme mixtures during the incubation. PMID- 12126107 TI - Scale-up of recombinant protein purification by hydrophobic interaction chromatography. AB - The scale-up of hydrophobic interaction chromatography is described. Human recombinant superoxide dismutase was used as a model. The scale-up was performed by keeping the height to diameter (H/D) ratio of the column constant. The success of scale-up was evaluated by reversed-phase high-performance liquid chromatography of the eluted material. The wrong H/D ratio causes decreased resolution. PMID- 12126108 TI - Purification of the C1 repressor of bacteriophage P1 by fast protein liquid chromatography. AB - A fast protein liquid chromatographic method is described for the purification of the C1 repressor of bacteriophage P1 and its truncated form C1*. By using one crude extract, both repressor proteins were purified in parallel to homogeneity and were shown to interact specifically with P1 operator DNA in vitro. The method involves an affinity chromatographic step on heparin-Sepharose, followed by a combination of ion-exchange chromatography on Q Sepharose and S Sepharose. The availability of a homogeneous preparation of the phage repressor is a prerequisite for studies on its structure-function relationship. PMID- 12126109 TI - Fast high-performance liquid chromatographic purification of Saccharomyces cerevisiae phosphoenolpyruvate carboxykinase. AB - A procedure was established for the rapid isolation of Saccharomyces cerevisiae phosphoenolpyruvate carboxykinase (PEPCK) from an overproducing strain. Overexpression was achieved by the transformation of yeast cells with the multicopy plasmid YEp352 harbouring the PEPCK structural gene. The enzyme was purified to homogeneity using first anion-exchange chromatography on Q-Sepharose followed by hydrophobic interaction chromatography on phenyl-Sepharose and gel filtration on Sephacryl S200. The purified phosphoenolpyruvate carboxykinase was further characterized with respect to the molecular mass, displaying an apparent molecular mass corresponding to a tetrameric form. PMID- 12126110 TI - Purification of cytochromes P-450 derived from liver microsomes of untreated and 2,3,7,8-tetrachlorodibenzo-p-dioxin-treated marmoset monkeys. AB - The purification of multiple forms of cytochrome P-450 (P450) from 2,3,7,8 tetrachlorodibenzo-p-dioxin (TCDD)-treated marmosets using fast protein liquid chromatography (FPLC) is described. The main aim was to achieve a better separation of certain closely related P450 sub-forms from each other than that previously obtained using conventional chromatography. An 8-aminooctyl-Sepharose fraction of cholate-solubilized microsomes was obtained first and, after fast desalting on Sephadex G-25, loaded on to a preparative Mono Q column. Five of the six gradient peaks contained P450 and were each rechromatographed on an analytical Mono Q column. The pass-through peak was fractionated further using a Mono S column. Other HPLC-quality anion- and cation-exchange gels were compared. For removal of excess of non-ionic detergent, five types of hydroxyapatite gels were compared. Seven purified forms of P450 and cytochrome b5 and P420 were isolated and characterized according to PHAST sodium dodecylsulphate polyacrylamide gel electrophoretic apparent molecular masses, catalytic, spectral and magnetic properties and also TCDD-binding capacity (molar ratio of [14C]TCDD to P450). There are at least two sub-forms which appear to be TCDD inducible, one showing a substantial ethoxyresorufin-O-deethylase activity and the other having a high TCDD-binding molar ratio. Two other forms appear to be constitutive, as deduced from comparisons with forms purified from untreated animals. PMID- 12126111 TI - Activation of topoisomerase II during partial purification by heparin-Sepharose chromatography. AB - Partial purification of topoisomerase II from small samples (10(7)-10(8) cells) of human leukaemic cells was achieved by isolation of cell nuclei, hyper-osmotic extraction of nuclear proteins, sorption of nuclear proteins by heparin-Sepharose and elution with potassium phosphate. Similar results were obtained by gradient and batchwise elution. The calatylic activity of topoisomerase increased ca. eightfold after removal of ca. 95% of the contaminating nuclear proteins. The conserved enzymatic activity after partial purification indicates that the enzyme was not damaged. The half-life of enzymatic activity is increased by the chromatographic procedure. Owing to its high yield and technical simplicity, this could be a candidate procedure for the study of topoisomerase II in patient derived blood samples. PMID- 12126112 TI - Angiotensinase A (aminopeptidase A): properties of chromatographically purified isoforms from human kidney. AB - Angiotensin-II-cleaving angiotensinase A (aminopeptidase A, E.C. 3.4.11.7, ATA) plays an important role in glomerular haemodynamics. the pathophysiology of essential arterial hypertension and the induction of vascular disorders. In order to study biochemical and immunological properties of ATA, two isoforms (I and II) of the glycoprotein were isolated for the first time from human kidney cortex. Kidney cortex homogenate, digested with bromelain, was fractionated by ammonium sulphate precipitation and subsequent hydrophobic interaction chromatography, using a fast protein liquid chromatographic (FPLC) system. By anion-exchange FPLC (Mono Q column), the isoforms of ATA were eluted in two distinct peaks and were further purified by size-exclusion FPLC and preparative polyacrylamide gel electrophoresis. Biochemical, immunological and immunohistological characterization disclosed differences in the intrarenal localization, glycosylation Michaelis constant and apparent molecular mass (native and sodium dodecyl sulphate gel electrophoresis) but similar properties in the double immunodiffusion technique. Polyclonal rabbit antibodies, raised against ATA isoforms I and II, precipitated an analogous antigen in urine from patients with renal tubular damage. PMID- 12126113 TI - Board funds NCVEI core competency projects. PMID- 12126114 TI - HSUS says ban reptiles as pets; few agree. Humane Society of the United States. PMID- 12126115 TI - Euthanatized animals can poison wildlife: veterinarians receive fines. PMID- 12126116 TI - Genetically engineering a pet? History shows it won't be easy. PMID- 12126117 TI - Mentoring, public education, thanks to Minnesota new moms. PMID- 12126118 TI - Praise for outcomes assessment program. PMID- 12126119 TI - Thinks calf roping, steer tripping inhumane. PMID- 12126120 TI - More information on blood component therapy. PMID- 12126121 TI - Agrees with treatment for urine spraying, marking. PMID- 12126122 TI - Disagrees with classification of a treatment for digital dermatitis. PMID- 12126123 TI - What is your diagnosis? An ill-defined soft-tissue opacity in the caudal abdominal region and numerous metallic opacities in the soft tissues of the lumbar area. PMID- 12126124 TI - ECG of the month. Second-degree atrioventricular (AV) block, type II advanced (3:1), with ventricular escape complexes. PMID- 12126125 TI - Thenogenology question of the month. Pyometra. PMID- 12126126 TI - Employment, starting salaries, and educational indebtedness of year-2001 graduates of US veterinary medical colleges. PMID- 12126127 TI - Clinical and hematologic effects of experimental infection of dogs with recently identified Babesia gibsoni-like isolates from Oklahoma. AB - OBJECTIVE: To characterize clinical and hematologic responses in dogs following experimental inoculation with Babesia gibsoni-like isolates from infected dogs in Oklahoma. DESIGN: Prospective study. ANIMALS: 6 mixed-breed dogs. PROCEDURE: 2 dogs were inoculated with organisms from a naturally infected dog, and 3 were inoculated with organisms from a second naturally infected dog (1 of these 3 dogs was splenectomized 1 week prior to inoculation). One dog was not inoculated. Complete blood counts were performed weekly. RESULTS: In the 5 dogs inoculated with organisms, parasites were initially detected 1 to 5 weeks after inoculation, and severity of parasitemia peaked with 1.9 to 6.0% of RBC infected by 4 to 6 weeks after inoculation. Parasitemia was easily detectable (> 0.1% of RBC infected) for 3 to 4 weeks. Clinical abnormalities included lethargy, fever, and pale mucous membranes but were mild to nearly inapparent in 2 dogs. All dogs developed regenerative anemia and marked thrombocytopenia. Thrombocytopenia developed before and lasted longer than the parasitemia. Profound but transient neutropenia was detected in some dogs. The splenectomized dog developed more severe parasitemia and anemia and more pronounced clinical abnormalities. Three dogs with intact spleens recovered without treatment. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that 2 or more genotypically distinct, but morphologically identical, small Babesia parasites can infect dogs in the United States. Compared with infection with small Babesia parasites from California, infection with these isolates resulted in less severe parasitemia and clinical abnormalities. Parasitemia was transient, indicating that identification of organisms in blood smears may be difficult in some dogs. PMID- 12126128 TI - Enrofloxacin resistance in Escherichia coli isolated from dogs with urinary tract infections. AB - OBJECTIVE: To assess the strain heterogeneity of enrofloxacin-resistant Escherichia coli associated with urinary tract infections in dogs at a veterinary medical teaching hospital (VMTH). In addition, strains from other veterinary hospitals in California were compared with the VMTH strains to assess the geographic distribution of specific enrofloxacin-resistant E. coli isolates. DESIGN: Bacteriologic study. SAMPLE POPULATION: 56 isolates of E. coli from urine samples (43 isolates from dogs at the VMTH, 13 isolates from dogs from other veterinary clinics in California). PROCEDURES: Pulsed field gel electrophoresis was performed on 56 isolates of E. coli from urine samples from 56 dogs. All 56 isolates were tested for susceptibility to amoxicillin, chloramphenicol, enrofloxacin, tetracycline, trimethoprim-sulphamethoxazole, cephalexin, and ampicillin. Enrofloxacin usage data from 1994 to 1998 were obtained from the VMTH pharmacy. RESULTS: Several strains of enrofloxacin-resistant E. coli were collected from urine samples from the VMTH, and strains identical to those from the VMTH were collected from other veterinary clinics in California. For the isolates that did share similar DNA banding patterns, variable antibiotic resistance profiles were observed. CONCLUSIONS AND CLINICAL RELEVANCE: The increased occurrence of enrofloxacin-resistant E. coli from urine samples from dogs at the VMTH was not likely attributable to a single enrofloxacin-resistant clone but may be attributed to a collective increase in enrofloxacin resistance among uropathogenic E. coli in dogs in general. PMID- 12126129 TI - Heterobilharzia americana infection in a dog. AB - A 7-year-old castrated male Golden Retriever cross was evaluated because of intermittent blood-tinged diarrhea, severe weight loss, anorexia, and lethargy of 2 months' duration; the dog was unresponsive to antimicrobial and standard anthelmintic treatment. Results of fecal flotations for parasite ova were negative. Alkaline phosphatase, aspartate aminotransferase, and alanine aminotransferase activities and total protein and globulin conentrations were greater than reference ranges. Biopsy specimens were obtained during laparotomy and examination revealed multiple granulomatous lesions with helminth ova nidi in the intestine, pancreas, liver, and mesenteric lymph node. Saline solution direct smear and saline solution sedimentation of feces yielded trematode ova that were morphologically consistent with Heterobilharzia americana. Identification was confirmed when miracidia were hatched from these ova and produced characteristic cercariae from infected snails. An antigen capture ELISA, typically used for the diagnosis of schistosomiasis in humans, was performed, and schistosome circulating anodic antigen was detected. Treatment with 30 mg of praziquantel/kg (14 mg/lb) of body weight stopped ova shedding, removed detectable circulating antigens, and caused the dog's body weight and attitude to return to normal. Although this is the first report of canine heterobilharziasis in North Carolina, it suggests that heterobilharziasis is underdiagnosed in dogs that have contact with water frequented by raccoons. Inappropriate diagnostic procedures can foil accurate detection of this parasitic disease. PMID- 12126130 TI - Use of human immunoglobulin for treatment of severe erythema multiforme in a cat. AB - A 5-month old female domestic shorthair cat developed lethargy and severe ulcerative skin lesions that covered more than half of its body after routine administration of rabies vaccine, anthelmintic, and ear medication. Clinical and histologic findings were consistent with a severe cutaneous drug reaction or erythema multiforme. The cat's condition continued to deteriorate despite drug withdrawal and supportive care. Administration of human intravenous immunoglobulin was well tolerated by the cat and led to rapid resolution of ulcerative cutaneous lesions, accompanied by substantial improvement in the cat's demeanor within 8 days. Human intravenous immunoglobulin appears to be a novel promising treatment for life-threatening cutaneous drug reactions. PMID- 12126132 TI - Detection of antibodies against Sarcocystis neurona in cerebrospinal fluid from clinically normal neonatal foals. AB - OBJECTIVE: To determine whether antibodies against Sarcocystis neurona could be detected in CSF from clinically normal neonatal (2 to 7 days old) and young (2 to 3 months old) foals. DESIGN: Prospective study. ANIMALS: 15 clinically normal neonatal Thoroughbred foals. PROCEDURE: Serum and CSF samples were obtained from foals at 2 to 7 days of age and tested for antibodies against S. neurona by means of western blotting. Serum samples from the mares were also tested for antibodies against S. neurona. Additional CSF and blood samples were obtained from 5 foals between 13 and 41 days after birth and between 62 and 90 days after birth. RESULTS: Antibodies against S. neurona were detected in serum from 13 mares and their foals; antibodies against S. neurona were detected in CSF from 12 of these 13 foals. Degree of immunoreactivity in serum and CSF decreased over time, and antibodies against S. neurona were no longer detected in CSF from 2 foals 83 and 84 days after birth. However, antibodies could still be detected in CSF from the other 3 foals between 62 and 90 days after birth. CONCLUSIONS AND CLINICAL RELEVANCE: Results indicate that antibodies against S. neurona can be detected in CSF from clinically normal neonatal (2 to 7 days old) foals born to seropositive mares. This suggests that western blotting of CSF cannot be reliably used to diagnose equine protozoal myeloencephalitis in foals < 3 months of age born to seropositive mares. PMID- 12126131 TI - Population and survival characteristics of cats with hypertrophic cardiomyopathy: 260 cases (1990-1999). AB - OBJECTIVE: To determine current population characteristics of, clinical findings in, and survival times for cats with hypertrophic cardiomyopathy (HCM). DESIGN: Retrospective study. ANIMALS: 260 cats with HCM. PROCEDURE: Information was obtained from the medical records. Cats were classified into 1 of 4 clinical groups (congestive heart failure [CHF] group, arterial thromboembolism [ATE] group, syncope group, or cats without clinical signs [subclinical group]) on the basis of the primary clinical signs at the initial examination. RESULTS: 120 cats were classified in the CHF group, 43 in the ATE group, 10 in the syncope group, and 87 in the subclinical group. Antecedent events that may have precipitated CHF included i.v. fluid administration, anesthesia, surgery, and recent corticosteroid administration. Median survival time was 709 days (range, 2 to 4,418 days) for cats that survived > 24 hours. Cats in the subclinical group lived the longest (median survival time, 1,129 days; range, 2 to 3,778 days), followed by cats in the syncope group (654 days; range, 28 to 1,505 days), cats in the CHF group (563 days; range, 2 to 4,418 days), and cats in the ATE group (184 days; range, 2 to 2,278 days). Causes of death included ATE (n = 56), CHF (49), sudden death (13), and noncardiac causes (27). In univariate analyses, survival time was negatively correlated with left atrial size, age, right ventricular enlargement, and thoracentesis. Cats with systolic anterior motion of the mitral valve lived longer than cats without this echocardiographic finding. In multivariate analyses, only age and left atrial size remained significant predictors of survival time. CONCLUSIONS AND CLINICAL RELEVANCE: Although overall survival time for cats with HCM was similar to earlier reports, survival times for cats with CHF or ATE were longer than previously reported. PMID- 12126133 TI - Percutaneous retrieval of a jugular catheter fragment from the pulmonary artery of a foal. AB - A 49-kg (107.8-lb) sexually intact male Arabian foal was evaluated at 3 days of age because of profuse watery diarrhea, anorexia, and signs of abdominal pain. Physical examination findings were unremarkable except for evidence of diarrhea. A catheter was placed in the right jugular vein for administration of antimicrobials and lactated Ringer's solution. The foal was discharged with instructions to the owner to continue antimicrobial administration and fluid therapy; at home, the owner inadvertently cut the catheter at the level of the hub during attempted removal, and the catheter fragment migrated distally in the jugular vein and subsequently lodged in the pulmonary artery. The foal was readmitted to the hospital for retrieval of the fragment, using a percutaneous retrieval technique. Catheter fragmentation is a well-recognized risk of catheterization in horses. Catheter fragments can be retrieved somewhat easily from the jugular vein; however, if the fragment migrates to the heart or pulmonary artery, imaging the fragment to locate and retrieve it can be difficult. Complications associated with catheter fragmentation include septicemia, endocarditis, lung abscesses, pulmonary embolism, dysrhythmias, cardiac perforation, pulmonary or caval thrombosis, and death. To our knowledge, this is the first report of successful retrieval of a catheter fragment from the pulmonary artery in a horse. PMID- 12126134 TI - Clinical assessment and outcome of three techniques for jejunal resection and anastomosis in horses: 59 cases (1989-2000). AB - OBJECTIVE: To compare postoperative complications, short- and long-term survival, and surgical times for hand-sewn end-to-end (EE), stapled functional end-to-end (FEE), and stapled side-to-side (SS) anastomotic techniques for jejunal resection in horses. DESIGN: Retrospective study. ANIMALS: 59 horses. PROCEDURE: Medical records were reviewed to obtain signalment, diagnosis, treatment, and outcome for horses that underwent jejunojejunostomy in our hospital. Only horses that recovered from anesthesia were included in the study. RESULTS: Among the 59 horses, there were 33 EE, 15 FEE, and 11 SS anastomoses. No difference was found in duration of surgery among the 3 techniques. The most common postoperative complications were colic episodes (56%), ileus (53%), diarrhea (20%), and adhesions (15%). Horses with SS anastomosis had a significantly shorter duration of postoperative ileus than the EE group did. No significant difference in duration of postoperative ileus was found among the other groups. No difference was found among the 3 anastomotic techniques in regard to survival rate at the time of discharge, 6 months after surgery, or 1 year after surgery. Overall survival rates after jejunal anastomosis were 88% at the time of discharge, 65% at 6 months after surgery, and 57% at > or = 1 year after surgery. CONCLUSIONS AND CLINICAL RELEVANCE: The hand-sewn EE, stapled FEE, and stapled SS anastomotic techniques should be considered equivalent methods for small intestinal anastomosis in the horse. However, the stapled SS technique may be preferred because of possible decreased duration of postoperative ileus. PMID- 12126135 TI - The influence of short-term anion salt exposure on urine pH and on resistance to experimentally induced hypocalcaemia in cows. AB - The objective of this experiment was to induce acidification by anion salt supplementation for 2 days or 10 days and to study the prophylactic effects of such supplementation in preventing hypocalcaemia in cows. It was further attempted to monitor the extent to which any effect on the calcium-regulating mechanisms would persist following a 10-day period of acidification with anion salts. Study animals were three untreated control cows and three cows supplemented with ammonium chloride and ammonium sulphate in their ration for 2 days or 10 days through the rumen cannula. The basic ration of hay was dominated by Urochloa spp. The pH of the urine of the control cows was around 8.00 throughout the experiment and was considered normal. Anion-supplemented cows produced urine with a daily mean pH between 5.5 and 7.0, possibly due to anion salt exposure. The ability to withstand hypocalcaemic challenges was tested by a standardized intravenous infusion of disodium ethylenediaminetetraacetate (Na2EDTA). The calcium regaining time (CRT), expressed as time spent to reach 1.00 mmol/l of ionized calcium during recovery from the EDTA-induced hypocalcaemia, was used to compare cow responses. In the control cows the unexpectedly short CRT, especially during the weekly EDTA tests, could be a result of the repeated induced episodes of hypocalcaemia caused by the EDTA infusions. The improved CRT in the anion-supplemented cows may thus be interpreted as the combined effect of the repeated hypocalcaemic episodes due to EDTA infusions and probably the effect of anion-induced metabolic acidosis on endocrine-regulated calcium homeostatic mechanisms. The effect of anion salt exposure for 10 days on the improvement of calcium-regulating mechanisms was not clear due to the unexpected improvement of CRT that was exhibited by the untreated control cows as well. An on-farm trial of the effect of a 2-day or 10 day anion exposure of dry cows on calcium-regulating mechanisms is suggested. PMID- 12126137 TI - Primary evaluation of methenamine as a NPN compound with probable effects on increasing ruminal escaped protein. AB - The effects of methenamine as a non-protein nitrogenous compound on protein and health status of feedlot lambs were studied in three groups of lambs receiving 5, 10, or 15 g of the substance daily in their feed for 100 days. The results were compared with data obtained from a control group receiving a diet low in crude protein without methenamine. Serum total protein, serum urea nitrogen and serum creatinine were measured every 10 days as indicators of protein metabolism and kidney function. Urine samples were also examined on the same days for possible side-effects of methenamine on the urinary tract. Following slaughter, various internal organs, including the brain and various parts of the gastrointestinal and urinary tracts, were examined both grossly and microscopically to detect any lesions. All groups receiving methenamine had serum total protein and serum urea nitrogen levels higher than those in the control group. The serum creatinine level was normal in all the groups throughout the experiment. No gross or microscopic lesion attributed to the toxic effects of methenamine was detected in any of the internal organs. Therefore, it is concluded that methenamine can be used as a non-protein nitrogenous compound without serious side-effects. PMID- 12126136 TI - Influence of a vegetarian diet versus a diet with fishmeal on bone in growing pigs. AB - This study was conducted to examine if substantial bone loss occurs in growing pigs fed a vegetarian diet in comparison with a diet containing fishmeal. Twelve 6-week-old weaned pigs were assigned to two groups: group V [vegetarian diet; 0.61% phosphorus (P) in dry matter until 25 kg and 0.46% P until the end of the experiment] and group F (fishmeal diet; 0.61% P in dry matter until 25 kg and 0.46% P until the end of the experiment). Phytase was added to both diets. These two diets were fed to the two groups for a period of 6 weeks. Blood samples were collected weekly, faeces were collected three times a week. Concentrations of osteocalcin (OC) and carboxyterminal telopeptide of type I collagen (ICTP) were measured in serum, using a radioimmunoassay, and bone-specific alkaline phosphatase (bAP) was measured using an enzyme immunoassay. Bone mineral density (BMD) and content (BMC) were determined by peripheral quantitative computer tomography (pQCT) in the tibia and phalanx. In addition, 1,25-dihydroxyvitamin D (VitD) and parathyroid hormone (PTH) were measured in serum. The digestibility of P was significantly decreased in group V. Significant changes in bAP activities and OC concentrations occurred with time during the 6 weeks. ICTP concentrations were significantly higher in group V. Total BMC and BMD in the tibia and BMD in the phalanx significantly decreased in group V. The results show that a vegetarian diet induces a significant loss of bone and a higher bone formation in group V compared with group F, although phytase was added to both diets. The dietary requirements for P in pigs, especially in the context of feeding vegetarian diets and adding an appropriate amount of phytase, should be investigated further. PMID- 12126138 TI - A study on the pathogenesis of equine sesamoiditis: the effects of experimental occlusion of the sesamoidean artery. AB - In this study the potential role of circulatory disturbances in the pathogenesis of sesamoiditis was investigated by studying the clinical and histological effects of experimental occlusion of the sesamoidean artery, which is the main nutrient artery of the proximal sesamoid bone (PSB). For this purpose, five adult Dutch Warmblood horses were used in which the sesamoidean artery was occluded with polyvinyl alcohol foam particles. Bone labelling was carried out with oxytetracycline and calcein. All animal were checked clinically three times a week and radiographically at days 14, 21, 28 and 35. At day 35 the animals were killed and the fetlock was dissected and macroscopically evaluated. The PSBs were isolated and radiographed and the soft tissues adjacent to the abaxial side of the PSBs were histologically examined [routine histology and for the neuropeptides substance P (SP) and calcitonin gene-related peptide (CGRP)]. The PSBs were divided into bony slices which were radiographed and evaluated histologically (routine, SP, CGRP, fluorescence). All horses showed a slight lameness that gradually diminished. Radiographically enlarged vascular channels were seen in only one horse. Histological data showed that on average 60% (range 37-89%) of the total area had been deprived of vascularization. In two horses an increase in the extent of the arterial network from the basal side of the PSB was seen. Only in the horse with the greatest extent of occlusion were bone necrosis and a reactively increased uptake of fluorochromes adjacent to the occluded arteries found. Bone density did not change. The distribution of neuropeptides in the surrounding soft tissues was not affected by the occlusion. From this study, it can be concluded that a large part of the arterial supply to the PSB can be interrupted without provoking histological and/or radiographic changes that are consistent with clinical sesamoiditis. Therefore, it seems very improbable that circulatory disturbances are a primary aetiopathogenic factor in the disease. PMID- 12126139 TI - Effect of vitamin E on ruminal fermentation in vitro. AB - The effects of vitamin E on pH value, total protozoa counts, volatile fatty acid (VFA), ammonia nitrogen and lactate levels were examined using an in vitro ruminal incubation system. The ruminal fluid (100 ml) of the first and second group was supplemented with 0.4 mg or 0.8 mg of vitamin E, respectively. Samples were taken immediately before and following 3, 6, 12 and 24 h of incubation at 39 degrees C and analysed for the total protozoa counts, the pH and the levels of ammonia nitrogen, lactate and VFA. Levels of propionate at 24 h and ammonia nitrogen at 12 and 24 h were significantly higher in the second group than in the control. In contrast, the levels of butyrate at 6, 12 and 24 h and lactate at 6, 12 and 24 h were lower in the second group than in the control. Propionate at 24 h, acetate levels at 6, 12 and 24 hand ammonia nitrogen levels at 6, 12 and 24 h and total rumen protozoa counts at 6, 12 and 24 h were significantly higher in the second group as compared with control. In contrary, butyrate levels at 6, 12 and 24 h, lactate levels at 6, 12 and 24 h were lower in second group than in control. There was no statistically significant difference among the groups in the pH values. In conclusion, the addition of vitamin E to in vitro ruminal fluid was found to increase the concentrations of acetate and propionate, total counts of protozoa, levels of ammonia nitrogen, but to decrease the butyrate and lactate levels of the ruminal aliquots in in vitro ruminal fermentation. PMID- 12126140 TI - Rats with a glucocorticoid-induced catabolic state show symptoms of oxidative stress and spleen atrophy: the effects of age and recovery. AB - In this study we wanted to determine whether changes in antioxidant profile could follow the catabolic effects of glucocorticoids. We also wanted to compare resistance to glucocorticoid overload in young and old rats. To address these questions, whole body catabolism was induced by the administration of dexamethasone (Dex) at either 2 mg/kg bodyweight/day to young (6 weeks old) or 0.5 mg/kg body-weight/day to old (94 weeks old) rats. Bodyweight loss of pair-fed rats not given Dex was only 2% in the young rats and 8% in the old rats, whereas in Dex-treated rats the decrease in bodyweight was 22% in the young rats and 13% in the old rats after 5 days of treatment. Spleen weight decreased by 65% in the young rats and by 52% in the old rats. Additionally, in the young rats there was a 46% reduction in glutathione (GSH) in erythrocytes as well as a 36% reduction in GSH/tissue wet weight in the soleus muscle. The corresponding figures for the old rats were 35 and 26%, respectively. Taken together, these results suggest that Dex directly and/or indirectly impaired the antioxidant reactions. This was further confirmed by a significant (50%) decline in Cu-Zn superoxide dismutase (SOD-1) activity in erythrocytes isolated from the young rats treated with Dex but not the old rats as they showed a significant elevation in SOD-1 activity (by 101%). Thiobarbituric acid reactant substances were significantly higher in both young and old rats. Activity of blood plasma creatine kinase increased by 73% in the young rats and by 307% in the old rats treated with Dex. Although both the young and the old rats could recover from oxidative stress, the old rats in contrast to the young rats remained catabolic until the end of the experiment. In conclusion, we suggest that old rats are more vulnerable to the catabolic action of Dex, whereas young rats are more susceptible to the oxidative stress induced by Dex. PMID- 12126141 TI - Correlations among body temperature, plasma progesterone, cortisol and prostaglandin F2alpha of the periparturient bitch. AB - The results of this study suggest that, besides the irrelevant role of body temperature measurement to predict the impending parturition in the bitch, progesterone and 15-ketodihydroprostaglandin F2alpha plasma level records could be more suitable to detect the approaching whelping in this species. More interesting was the statistically significant substantial increase in body temperature beginning 12 h after the onset of parturition. Therefore, if any significant increase in body temperature is recorded at the end of pregnancy without the beginning of the expulsion of fetuses, it could indicate problems at parturition. In this study, cortisol levels increased significantly at the time of delivery and remained high 12 h after the beginning of parturition, decreasing within 36 h after the onset of whelping. 15-ketodihydro-prostaglandin F2alpha levels increased significantly 24 h before parturition and again at the onset of whelping. Progesterone levels decreased significantly, starting 24 h before the onset of whelping and remained low after delivery. PMID- 12126142 TI - Carbon fibres and plasma-preserved tendon allografts for gap repair of flexor tendon in bovines: gross, microscopic and scanning electron microscopic observations. AB - The efficacy of carbon fibres and plasma-preserved tendon allografts for gap repair in the superficial digital flexor tendon in the mid-metatarsal region was evaluated in 12 crossbred calves. Experimental tenectomies were performed, followed by implantation of carbon fibres in group I (12 legs) and plasma preserved tendon allografts in group II (12 legs). Gross observations in group I showed filling of the defect with granulation tissue with more vascularity on day 7, which was less prominent at day 14. On day 30, the neotendon formed was slightly thicker and comparable to normal tendon in appearance and texture. On day 90, it exhibited all the characteristics of a fully developed tendon. Whereas, in group II increased vascularity at the site and encapsulation of the graft with connective tissue in early periods was observed. The gap between graft and host was filled with fibrous connective tissue. Peritendinous adhesions were maximum on day 7 which were gradually reduced in both groups. Microscopically, an acute inflammatory reaction in the periphery of carbon fibres was observed on day 7. Immature fibroblasts were arranged in a haphazard pattern at this stage. By day 14, numerous newly formed capillaries and comparatively more mature fibroblasts were present in between and around the carbon fibres which were aligning parallel to the longitudinal axis of the tendon. By day 30 the healing tissue exhibited longitudinal orientation of collagen fibres and was at a more advance stage of maturation. By day 90, the neotendon formed simulated the picture of normal tendon. In the grafted tendon group, there was normal healing tissue at the functional sites between host and grafted tendon. The fibroblastic activity appeared to be both extrinsic and intrinsic in origin. The connective tissue had invaded the graft to a variable distance and there was resorption of graft which was replaced by newly formed connective tissue on day 90. Scanning electron microscopic observation revealed formation of neotendon between carbon fibre strands, resulting in thickening of the implant. In later stages parallel collagen fibres resembling normal tendon were observed in both groups. PMID- 12126143 TI - Caudomedial approach for removal of an ununited anconeal process and assessment of the medial coronoid process of the ulna. AB - A caudomedial approach that allows the removal of an ununited anconeal process (UAP) is described. Intraoperative examination of the medial part of the joint to diagnose a fragmented coronoid process (FCP) is also possible using the same approach. Sixty-two joints (53 dogs) with UAP and various degrees of arthrosis were included in the study. In all joints the UAP could be removed via the caudomedial approach and the medial coronoid process could be inspected after extending the incision of the joint capsule distally. In eight cases a FCP was diagnosed and removed either by the caudomedial or by a medial approach. The results of the present study show that an UAP can occur simultaneously with a FCP in the same joint and that inspection of the medial part of the joint is useful. PMID- 12126144 TI - Endometrial hyperplasia and mineralization in zoo felids treated with melengestrol acetate contraceptives. AB - Melengestrol acetate (MGA) contraceptives are widely used in zoo felids to regulate fertility and may have deleterious effects on endometrial health. To determine whether MGA exposure was associated with endometrial disease, the genital tracts of 212 zoo felids (99 MGA treated and 113 control) representing 23 species were evaluated. Adenomatous and cystic hyperplasia were prevalent in both MGA-treated (85%) and control (61%) groups, and the risk of developing these lesions increased with age. Treatment with MGA further increased the risk of developing advanced hyperplasia regardless of dose, and treatment for >72 months significantly elevated that risk, whereas parous animals had a lower risk. Endometrial polyps, fibrosis, adenomyosis, and hydrometra occurred in both MGA treated and control animals. MGA treatment was associated with an increased risk of hydrometra and mineralization but not of adenomyosis, polyps, or fibrosis after adjusting for advanced hyperplasia. Acute or chronic endometritis were associated with advanced hyperplasia but not with MGA treatment. These results indicate that proliferative and inflammatory endometrial lesions are common spontaneous diseases in zoo cats, and MGA contraceptives increase the risk of some diseases. The association of MGA with endometrial lesions that could impair fertility should be considered when using this contraceptive in genetically valuable felids. PMID- 12126145 TI - Cyclooxygenase-2 expression in spontaneous intestinal neoplasia of domestic dogs. AB - Cyclooxygenase-2 (Cox-2) is commonly upregulated during human colorectal tumorigenesis, and its contribution to this process has been clearly demonstrated in genetic mouse models. The only other species that naturally develops intestinal cancer with any frequency is the domestic dog. Intestinal carcinogenesis in humans has been strongly linked to environmental factors such as diet, which may be shared by household pets. We have previously reported that beta-catenin is overexpressed in the neoplastic epithelium of canine colorectal polyps, as it is in humans and rodents. We now show that Cox-2 is also upregulated in the majority of these lesions. Thirteen out of 20 colorectal adenomas (65%) contained immunohistochemically detectable Cox-2 protein restricted to the nonneoplastic tumor stroma, including myofibroblasts and a smooth muscle actin-negative mesenchymal cells morphologically consistent with macrophages and/or fibroblasts. In contrast to benign polyps, seven of 15 adenocarcinomas (47%) also expressed Cox-2 in the neoplastic epithelium. These changes duplicate molecular changes in human intestinal tumorigenesis and substantiate a fundamental role for both beta-catenin and Cox-2 in intestinal neoplasia. PMID- 12126146 TI - Primary infection, latency, and reactivation of bovine herpesvirus type 5 in the bovine nervous system. AB - Bovine herpesvirus type 5 (BHV-5) infection in calves causes meningoencephalitis, a fatal disease highly prevalent in South America. To study the pathogenesis of BHV-5 infection in cattle, 12 calves (group 1: acute infection) and 11 calves (group 2: latent infection) were intranasally inoculated with an Argentinean BHV 5 isolate at 10(8) and 10(4.7) tissue culture infective doses, respectively; six calves (control group) were mock infected. At 3 months postinoculation, all of the calves in group 2 and three calves in group 3 were given dexamethasone to reactivate the virus. The animals were euthanatized between days 6 and 17 postinoculation (group 1) and between days 6 and 16 postreactivation (group 2). Seventy-five percent and 91% of animals in groups 1 and 2, respectively, excreted BHV-5 in nasal and ocular discharges. Following dexamethasone administration, 45% of calves shed virus in both types of secretions. Spontaneous virus reactivation and shedding was observed in one calf. Neurologic signs consisting of circling, teeth grinding, ptyalism, jaw chomping, tongue protrusion, and apathy were observed in two animals in group 1 and, during the reactivation period, in four animals in group 2. Macroscopic findings consisted of softening of the cerebral tissue, meningeal hemorrhages and swelling, and edema and hemorrhages of prescapular, retropharyngeal and submandibular lymph nodes. Histologic lesions consisted of meningitis, mononuclear perivascular cuffing, neuronophagia, satellitosis, gliosis, hemorrhage, and necrosis and edema. Lesions in anterior cerebral cortex, medulla, and pons were consistently seen in all the animals of group 1. In the acutely infected animals, lesions in the diencephalon appeared at day 10 postinoculation, whereas in the latently infected calves these lesions were observed as early as at day 6 postreactivation. Latently infected animals developed lesions simultaneously in anterior cortex, medulla, pons, and diencephalon, showing a remarkable difference from the acutely infected group. Trigeminal ganglionitis appeared relatively early in animals of both groups (day 7 postinoculation in group 1 and day 8 postreactivation in group 2). PMID- 12126147 TI - Immunohistochemical characterization of tumor cells and inflammatory infiltrate associated with cutaneous melanocytic tumors of Duroc and Iberian swine. AB - The immunophenotype of tumor cells and inflammatory infiltrate associated with cutaneous melanocytic lesions (29 melanocytomas, two malignant melanomas, and 23 residual lesions) from 54 adult Iberian and Iberian x Duroc pigs were examined using a panel of nine antibodies. All neoplastic cells were vimentin+, cytokeratin-, and alpha-1-antitrypsin- and the majority were S100+, whereas all pigmented macrophages were vimentin+, cytokeratin-, and S100- and most expressed alpha-1-antitrypsin. Regressing tumors were characterized by zones with low density of neoplastic cells accompanied by heavy infiltration of CD3+ T lymphocytes, whereas zones with high density of neoplastic cells showed very low numbers of CD3+ T lymphocytes. The infiltrate of CD79a+ B cells and IgG, IgM, and IgA plasma cells was low. The majority of lymphocytes of the peri- and intratumoral infiltrate were major histocompatibility complex class II+, but neoplastic cells did not express class II antigen. The 17 residual lesions examined were composed of macrophages containing abundant melanin pigment and low to moderate numbers of CD3+ T lymphocytes. The results of the present study suggest that the local cellular immune response plays a crucial role in the host response that induces regression of cutaneous melanomas and melanocytomas of the Iberian and crossbred Iberian x Duroc pigs. PMID- 12126148 TI - Histopathology and biologic behavior of pleomorphic cutaneous mast cell tumors in fifteen cats. AB - Most feline cutaneous mast cell tumors (CMCT) are behaviorally benign; however, there is a subset of these tumors with, marked pleomorphism (previously termed poorly differentiated) that have been reported to be more aggressive. In this study, pleomorphic CMCT from 15 cats were identified from surgical biopsy submissions, and follow-up clinical data were obtained for 14 of these cats. Pleomorphic CMCT were discrete dermal nodules composed of sheets of pleomorphic round cells. Tumors from all 15 cats contained markedly cytomegalic and karyomegalic cells; 9/15 tumors (60%) contained multinucleated tumor giant cells. Typical mast cell granules were easily identified in sections stained with hematoxylin and eosin and with metachromatic stains and based on ultrastructural evaluation in cytomegalic as well as smaller tumor cells, indicating that the tumors were not poorly differentiated. The mitotic rate was very low (<1 mitosis per 10 high-power fields [hpf]) in 14 of 15 tumors (93%). Affected cats were 6-19 years old (mean age = 11.5 years), and there was no breed or sex predilection. Two cats had local recurrence. The only cat that had a pleomorphic CMCT with a high mitotic rate (1-2 mitoses/hpf) subsequently developed numerous other dermal neoplasms and was euthanatized. In this study, the large majority of feline pleomorphic CMCT were behaviorally benign. Mitotic rate is likely an important prognostic indicator of CMCT behavior. PMID- 12126149 TI - Expression and significance of p53, rb, p21/waf-1, p16/ink-4a, and PTEN tumor suppressors in canine melanoma. AB - The role of tumor suppressor genes in the pathogenesis of canine melanoma is incompletely understood. The genes encoding the tumor suppressors p53, Rb, p21 (waf-1), p16 (ink-4a), and PTEN have been postulated to contribute to the pathogenesis of melanoma in humans and experimental animal models. To assess whether inactivation of these genes similarly contributes to the origin and progression of canine melanoma, we examined their expression in seven distinct canine melanoma cell lines and in 31 retrospective samples (representing 29 dogs) of spontaneous canine melanoma. Various patterns suggestive of loss of tumor suppressor function emerged in these cell lines. The most frequently observed abnormality was loss or significant reduction of p16 expression in six of seven cell lines and in 21 of 26 tumor samples. Loss or significant reduction of PTEN expression was seen in four of seven cell lines and in 13 of 27 tumor samples. Although p53 was detectable in all the cell lines and in 24 of 30 tumors, exclusion of p53 from the nuclear compartment was observed in each of the cell lines and in 18 of 25 tumor samples. These results indicate that loss of function of these tumor suppressor proteins is a common occurrence that may contribute to the origin of canine melanoma. In our sample population, abnormalities in the expression or localization of one or more tumor suppressor proteins occurred with similar frequency in malignant and benign tumors; thus, additional work is necessary to determine how these proteins may impact disease progression and response to therapy. PMID- 12126150 TI - Immunohistochemical and histopathologic features of 14 malignant fibrous histiocytomas from Flat-Coated Retrievers. AB - Flat-Coated Retrievers seem to be at increased risk of developing soft-tissue sarcomas, and undifferentiated round cell or spindle cell sarcomas account for approximately 59% of sarcomas in the breed. In an attempt to classify these tumors further, formalin-fixed, paraffin-embedded sections from 14 undifferentiated sarcomas from Flat-Coated Retrievers were reviewed and examined with a panel of histologic and immunohistochemical stains. The panel included vimentin, desmin, Myo D1, smooth muscle actin, cytokeratin, S100, von Willebrand factor (factor VIII), Mac 387, CD3, major histocompatibility complex (MHC) class II, and CD79a. The majority of the sarcomas showed greater than 70% staining for MHC class II. We conclude that these undifferentiated sarcomas in Flat-Coated Retrievers belong to a spectrum of tumors with varying proportions of characteristic cell types and morphologic features, some of which fit the diagnostic criteria for malignant fibrous histiocytoma. Many of these sarcomas seem to have a significant myofibroblast component and a mild or moderate T cell infiltrate but the precise cell lineage is still uncertain. PMID- 12126151 TI - Immunohistochemical detection of thyroid transcription factor-1, thyroglobulin, and calcitonin in canine normal, hyperplastic, and neoplastic thyroid gland. AB - Immunohistochemistry for thyroid transcription factor-1 (TTF-1), thyroglobulin, and calcitonin was done in normal, hyperplastic, and neoplastic canine thyroid glands that had been fixed in formalin and embedded in paraffin. Prolonged fixation did not significantly alter the immunostaining for TTF-1. Staining for TTF-1 was always nuclear and usually strong. One of two C-cell adenomas, five of five follicular cell adenomas, 5 of 11 C-cell carcinomas, 38 of 42 follicular cell carcinomas, two of five cases of C-cell hyperplasia, two of two cases of follicular epithelial hyperplasia, one of two metastatic C-cell carcinomas, and three of four metastatic follicular carcinomas were positive for TTF-1. One follicular carcinoma that was positive for TTF-1 was negative for thyroglobulin. Thirty-nine of 42 follicular cell carcinomas were positive for thyroglobulin, including two cases that were negative for TTF-1. All C-cell lesions were positive for calcitonin. Of a variety of normal canine tissues and 278 nonthyroid tumors, only normal lung (airway and alveolar epithelial cells) and four of five pulmonary carcinomas were positive for TTF-1. TTF-1 is a good marker of thyroid differentiation and can be used in conjunction with thyroglobulin and calcitonin to increase the detection and differentiation of thyroid tumors and their metastases. PMID- 12126152 TI - Expression of vascular endothelial growth factor in canine mammary tumors. AB - Vascular endothelial growth factor (VEGF) is a dimeric protein that stimulates angiogenesis in vitro and in vivo by inducing endothelial cell proliferation and migration. In this immunohistochemical study, VEGF-immunolabeled cells were counted in a series of 10 benign and 40 malignant canine mammary tumors. The morphologic pattern of VEGF positivity (intensity of immunolabeling and VEGF granule size and distribution) was also evaluated. A low number of cells weakly positive for VEGF with few and small granules polarized to the luminal pole was detected in benign neoplasms. In contrast, in malignancies a high number of VEGF positive cells had strong immunolabeling, often with large granules found diffusely in the cytoplasm. This level of immunolabeling was more pronounced in the less differentiated, more malignant phenotypes (grade 3). Macrophages, which can synthesize VEGF, were strongly positive. Stromal and myoepithelial cells were negative. VEGF data were correlated statistically with intratumoral microvessel density (number of newly formed microvessels) and both measures were greater in less differentiated malignant neoplasms, demonstrating that angiogenesis and malignancy increase together. VEGF appears to be a powerful angiogenic factor in canine mammary tumors. PMID- 12126153 TI - Globoid cell-like leukodystrophy in a domestic longhaired cat. AB - Globoid cell leukodystrophy (GLD; Krabbe disease), is a rare heritable metabolic disorder in humans, dogs, mutant twitcher mice, and rhesus monkeys that is caused by a deficiency in the lysosomal enzyme galactocerebrosidase (GALC). GALC deficiency results in the accumulation of psychosine, which is toxic to oligodendrocytes and Schwann cells of the central and peripheral nervous systems. Clinical signs include hypotonia, mental regression, and death by 2 years of age in most human patients. Here we describe a domestic longhaired kitten with rapidly progressive neurologic disease and brain and spinal cord lesions characteristic of GLD. Pathologic hallmarks of the disease reflect the loss of oligodendrocytes and include myelin loss, gliosis, and the perivascular accumulation of large mononuclear cells with fine cytoplasmic vacuoles (globoid cells) in the peripheral and central nervous systems. Globoid cells were CD68 and ferritin positive, confirming their monocytic origin, and cytoplasmic contents were nonmetachromatic and periodic acid-Schiff positive. PMID- 12126154 TI - Neuroendocrine carcinoma of the nasopharynx in a dog. AB - Neuroendocrine carcinoma of the nasopharynx was diagnosed in a 9-year-old male Golden Retriever. The mass was identified by computed tomography of the nasal cavity and nasopharyngoscopy, and it was surgically excised. Histologic, cytochemical, and electromicroscopic examination of specimens confirmed the type of tumor. The dog was clincally improved for 150 days but was then reexamined because of respiratory difficulty and poor appetite. Thoracic radiographs revealed multiple nodules in all lung lobes, and ultrasonography revealed a mass in the spleen. The dog died the next day. PMID- 12126155 TI - Bronchitis and bronchiolitis in a cat with cilia-associated respiratory bacillus like organisms. AB - A 10-year-old male domestic shorthair cat died during anesthesia. Grossly, the lungs had multiple nodules corresponding to inflamed airways (bronchitis and bronchiolitis). Microscopically, cuffs and nodular aggregates of lymphocytes and plasma cells surrounded airways. Peribronchiolar fibrosis was also common. Globule leukocytes infiltrated the respiratory epithelium of noninflamed or mildly inflamed bronchi. Argyrophilic and filamentous organisms, consistent with cilia-associated respiratory bacillus-like organisms (CLO), were intermixed with cilia of respiratory epithelium. Ultrastructurally, CLO were longer and thinner than cilia and had a distinct trilaminar membrane, central electron-lucent areas, and no specialized external structures. Silver stained lung sections from 18 additional feline airways revealed similar bacilli in 2/9 normal lungs, 1/7 lungs with bronchitis and bronchiolitis, and 1/2 lungs with pneumonia. The significance of CLO in the pulmonary lesions was not determined. PMID- 12126156 TI - Eimeria organisms develop in the epithelial cells of equine small intestine. AB - Histopathologic and immunohistochemical examinations were performed to determine the origin of host cells parasitized by Eimeria in the small intestines collected from five foals. Eimeria organisms at various stages (mainly microgametes and macrogametes) were frequently found in the cytoplasm of hypertrophied host cells in the lamina propria at the tips of villi of the jejunum and ileum. The cytoplasm of the host cell was immunohistochemically positive for cytokeratin AE1/AE3 and cytokeratin 13 and was negative for vimentin, desmin, alpha-smooth muscle actin, chromogranin A, neuron-specific enolase, and factor VIII. The host cells parasitized by Eimeria species had the immunostaining characteristics of epithelial cells but not of mesenchymal cells, endothelial cells of lacteals or capillaries, smooth muscle cells or neuroendocrine cells. These results suggest that the host cell of Eimeria species is possibly derived from intestinal epithelial cells and then displaced into the lamina propria of the small intestine. PMID- 12126157 TI - Hepatosplenic T-cell lymphoma in a mare. AB - A 21-year-old Dutch crossbred mare was presented with a 1-month history of weight loss and weakness. Clinical evaluation revealed severe anemia and thrombocytopenia with evidence of hepatic disease and muscular damage. Necropsy findings included diaphragmatic rupture with an extensive retroperitoneal hematoma and severe hepatosplenomegaly. Microscopic findings were characterized by hepatic sinusoidal and splenic red pulp infiltration by atypical CD3-positive lymphocytes. No other nodal or extranodal sites were affected. Hepatosplenic lymphoma with a probable T-cell origin was diagnosed based on gross and histologic findings. PMID- 12126158 TI - Myofiber expression of class I major histocompatibility complex accompanied by CD8+ T-cell-associated myofiber injury in a case of canine polymyositis. AB - A 7-year-old female Labrador Retriever dog showed extreme muscular weakness, muscle wasting, dysbasia, and mild dysphagia. An elevated value of creatine kinase (335 IU/liter) in the serum was detected. Electromyographic findings included increased insertional activity, fibrillation potentials, and bizarre high-frequency repetitive potentials. Histopathologic examination of skeletal muscles revealed myofiber necrosis and phagocytosis, regeneration of myofibers, and perivascular, perimysial, and endomysial infiltrations of lymphocytes, macrophages and plasma cells. Immunohistochemical evaluation demonstrated that infiltrative cells in the early stage of myositis were CD8+ T-cells and that an increased expression of major histocompatibility complex (MHC) class I was apparent on the surface of nonnecrotic muscle fibers. In contrast, many CD3+ cells (T cells) and HLA-DR-positive macrophages and B lymphocytes were found in the severely affected areas. These results suggest that both expression of MHC class I and CD8+ T-cell infiltration may play an important role in initiation of myositis. These histopathologic findings resemble those reported in naturally occurring polymyositis in humans. PMID- 12126159 TI - Tertoma in a feline unilateral cryptochid testis. PMID- 12126160 TI - Eye movement desensitization and reprocessing psychotherapy: a model for integrative medicine. PMID- 12126161 TI - The springs of Clifton: water and wellness. PMID- 12126162 TI - Brief case reports of medically supervised, water-only fasting associated with remission of autoimmune disease. PMID- 12126163 TI - Maggots and leeches: when science and aesthetics collide. PMID- 12126164 TI - Reasons and recipes for longevity. PMID- 12126165 TI - Research ethics questioned in Qigong study. PMID- 12126166 TI - Does longevity research marginalize mortality? PMID- 12126167 TI - Effects of guggul in a rat model of stroke. PMID- 12126168 TI - Study ethics, patients' rights highlight 11th NACCAM meeting. National Advisory Council for Complementary and Alternative Medicine. PMID- 12126169 TI - New codes for CAM: HHS review could make them a reality. PMID- 12126170 TI - Survey of naturopathic physicians: implications and recommendations. PMID- 12126171 TI - Tribulations and trials: acupuncture study designs affect outcome. PMID- 12126172 TI - New model guidelines for the use of complementary and alternative therapies in medical practice. PMID- 12126173 TI - Effects of static magnets on chronic knee pain and physical function: a double blind study. AB - CONTEXT: Static magnets have become an increasingly popular alternative therapy for individuals with musculoskeletal pain despite limited scientific evidence to support their efficacy or safety. OBJECTIVE: To determine the effects of static magnets on the pain and functional limitations associated with chronic knee pain due to degenerative joint disease. DESIGN: Double-blind, randomized, controlled clinical trial. SETTING: Pretests and posttests were conducted in an academic health science center. PARTICIPANTS: Forty-three ambulatory subjects with chronic pain in 1 or both knee joints who were recruited from outpatient clinics or who volunteered to participate. INTERVENTION: Subjects wore pads containing magnets or placebos over their painful knee joints for 2 weeks. MAIN OUTCOME MEASURES: Self-administered ratings of pain and physical function using the Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC) and a timed 15-m (50-ft) walk. RESULTS: Multivariate analysis of covariance revealed significantly greater improvements in the group wearing magnets (P=.002). Univariate analyses indicated that comparative changes in self-rated pain and physical function (P=.002 and .001, respectively) were greater than changes in gait speed (P=.042). CONCLUSIONS: The application of static magnets over painful knee joints appears to reduce pain and enhance functional movement. However, further study is needed to determine the physiological mechanisms responsible for this analgesic effect. PMID- 12126174 TI - Acustimulation wristbands for the relief of chemotherapy-induced nausea. AB - CONTEXT: Substantial evidence suggests that acupuncture-point stimulation may be effective in controlling side effects of chemotherapy. OBJECTIVE: To examine the efficacy of an acustimulation wristband for the relief of chemotherapy-induced nausea. DESIGN: Randomized clinical trial using a 3-level crossover design. SETTING: Three outpatient oncology clinics in the northeastern United States. PARTICIPANTS: Twenty-five women and 2 men who experienced moderate or more severe nausea following their first chemotherapy treatment. INTERVENTION: We compared active acustimulation of the Pericardium 6 (PC-6) point on the ventral surface of the wrist with sham acustimulation (a corresponding point on the posterior surface of the wrist). A control group received no acustimulation. OUTCOME MEASURES: Severity of nausea and quantity of antiemetic medication used. RESULTS: No statistically significant differences in average severity of nausea were observed between the 3 interventions. However, the data showed a difference close to statistical significance in the severity of delayed nausea reported during active acustimulation compared to no acustimulation (P <.06). In addition, patients took fewer antinausea pills during the active-acustimulation cycle of this experiment compared to the no-acustimulation phase (P < .05). CONCLUSION: Findings on the efficacy of an acustimulation band for the control of chemotherapy-induced nausea are positive but not conclusive. These findings provide ample justification for further study of acustimulation in clinical oncology. PMID- 12126175 TI - Changes in spirituality and well-being in a retreat program for cardiac patients. AB - CONTEXT: Many epidemiological studies indicate that spirituality or religion are positively correlated with health measures, but research is needed on interventions that change spirituality to verify that it actually affects health and to justify suggestions that changes in spiritual practices or beliefs may have health benefits. However, it is not clear that health interventions can influence spirituality or which techniques are effective. OBJECTIVE: To evaluate whether participation in a retreat program for cardiac patients and their partners resulted in changes in spirituality and whether changes in spirituality were related to changes in well-being meaning in life, anger, and confidence in handling problems. DESIGN: Participants filled out questionnaires before and after participating in the retreat. SETTING: Retreats were sponsored by the Health Promotion and Wellness Program, University of Wisconsin-Stevens Point, and were held in a remote training center. PARTICIPANTS: Notices were sent to cardiac rehabilitation programs and directly to heart patients, resulting in the enrollment of 72 first-time participants. INTERVENTION: The 2.5-day educational retreats included discussion and opportunities to experience healthy lifestyle options. Exercise, nutrition, stress management techniques, communication skills that enhance social support, and spiritual principles of healing were incorporated. Experiential practices included yoga, meditation, visualization, and prayer. RESULTS: Of the participants, 78% reported increased spirituality after the retreat. Changes in spirituality were positively associated with increased well-being meaning in life, confidence in handling problems, and decreased tendency to become angry. CONCLUSIONS: Programs that explore spirituality in a health context can result in increased spirituality that is associated with increased well-being and related measures. Many patients and their families want to integrate the spiritual and health dimensions of their lives. Further work is needed to develop healthcare settings that can support this integration. PMID- 12126176 TI - Complementary and alternative medical treatment of breast cancer: a survey of licensed North American naturopathic physicians. AB - CONTEXT: Complementary and alternative medicine (CAM) use is on the rise in the United States, especially for breast cancer patients. Many CAM therapies are delivered by licensed naturopathic physicians using individualized treatment plans. OBJECTIVE: To describe naturopathic treatment for women with breast cancer. DESIGN: Cross-sectional mail survey in 2 parts: screening form and 13 page survey. SETTING: Bastyr University Cancer Research Center, Kenmore, Wash. PARTICIPANTS: All licensed naturopathic physicians in the United States and Canada (N=1,356) received screening forms; 642 (47%) completed the form. Of the respondents, 333 (52%) were eligible, and 161 completed the survey (48%). MAIN OUTCOME MEASURES: Demographics of naturopathic physicians, development of treatment plans, CAM therapies used, perceived efficacy of therapeutic interventions. RESULTS: Of those respondents screened, 497 (77%) had provided naturopathic care to women with breast cancer, and 402 (63%) had treated women with breast cancer in the previous 12 months. Naturopaths who were women were more likely than men to treat breast cancer (P < or = .004). Of the survey respondents, 104 (65%) practiced in the United States, and 57 (35%) practiced in Canada; 107 (66.5%) were women, and 54 (33.5%) were men. To develop naturopathic treatment plans, naturopathic physicians most often considered the stage of cancer, the patient's emotional constitution, and the conventional therapies used. To monitor patients clinically, 64% of the naturopathic physicians used diagnostic imaging, 57% considered the patient's quality of life, and 51% used physical examinations. The most common general CAM therapies used were dietary counseling (94%), botanical medicines (88%), antioxidants (84%), and supplemental nutrition (84%). The most common specific treatments were vitamin C (39%), coenzyme Q-10 (34%), and Hoxsey formula (29%). PMID- 12126177 TI - Thomas Rau, MD: biological medicine and the dynamics of regulation. Interview by Bonnie Horrigan. PMID- 12126178 TI - Progress notes: Maine Medical Center family practice residency program. PMID- 12126180 TI - Interview with Wallace D. Winters. AB - Wallace Winters exemplifies the model of the basic scientist/clinical toxicologist. His extensive research interests have led to a better understanding of central nervous system excitation and depression, and have included pioneering studies on the neuropharmacology of gammahydroxy butyrate that date back to the 1960s. Dr. Winters was born in New York, NY on June 20, 1929. He received his undergraduate degree at George Washington University, his Ph.D. in Pharmacology/Toxicology at the University of Wisconsin, and his M.D. at the Medical College of Wisconsin. From 1959 to 1962 Dr. Winters pursued postdoctoral studies at the Brain Research Institute at University of California, Los Angeles where he remained on the faculty rising to full Professor. In 1971, Dr. Winters relocated to University of California at Davis where he served as a Professor of Pharmacology/Toxicology for the next 20 years. During this time, Dr. Winters founded the University of California Davis Poison Control Center in Sacramento in 1977 and served as its first medical director until 1983. From 1979 to 1984 he served on the American Academy of Clinical Toxicology Board of Trustees. After retiring from University of California, Davis, Dr. Winters worked as a Medical Officer for the U.S. Food and Drug Administration from 1991 to 1997, and continues to serve as a consultant and medical expert in clinical pharmacology and toxicology. PMID- 12126179 TI - Traditional use of sakau (kava) in Pohnpei: lessons for integrative medicine. PMID- 12126181 TI - Acute ingestion poisoning with insecticide formulations containing the pyrethroid permethrin, xylene, and surfactant: a review of 48 cases. AB - BACKGROUND: Forty-eight patients poisoned with insecticide formulations containing permethrin (a Type I pyrethroid insecticide), xylene, and surfactants are reported here. These patients were diagnosed and treated in the Chang Gung Memorial Hospital in Taiwan from January 1987 to June 1999. Ten patients ingested permethrin in error and 38 patients attempted suicide. Gastrointestinal tract signs and symptoms were most common (35/48; 73%), and included sore throat, mouth ulcerations, dysphagia, epigastric pain, and vomiting. Pulmonary abnormalities were documented in 29% (14/48) of patients. Aspiration pneumonitis occurred in eight patients, including onefatal case. Pulmonary edema was observed in two patients. Sixteen patients (33%) had central nervous system involvement including confusion (6/48; 13%), coma (10/48; 21%), and seizures (4/48; 8%). Cardiovascular symptoms in 3/48 (7%) patients were limited to arrhythmias and shock. Mild renal and hepatic dysfunction was found in 5/48 (10%) and 3/48 (6%) of patients, respectively. Leukocytosis occurred in 16 patients (33%) but was not associated with infection. Only one death occurred during this 12.5-year period. CONCLUSION: Poisoning caused by ingesting insecticides containing permethrin, xylene, and surfactant manifests primarily gastrointestinal tract symptoms and signs. The involvement of the central nervous system and lungs were less common, but clinically more significant. The relative contributions of the 20% permethrin, 70% xylene, and 10% surfactant to these toxic manifestations, however, is uncharacterized. PMID- 12126182 TI - Acute wood or coal exposure with carbon monoxide intoxication induces sister chromatid exchange. AB - OBJECTIVE: The object of this study was to investigate the genotoxic effect of acute overexposure to combustion products originating from coal or wood stoves in patients presenting with acute carbon monoxide intoxication. STUDY DESIGN: In a prospective study, we analyzed the frequency of sister chromatid exchange and the carboxyhemoglobin concentration in 20 consecutive patients without a history of smoking or drug use who had been treated in the Emergency Care Unit of Istanbul Medical Faculty due to acute carbon monoxide intoxication. All of these cases were domestic accidents due to dysfunctioning coal or wood stoves. The results were compared with a control group of 20 nonsmoking, nondrug-using healthy individuals matched for age, sex, and absence of other chemical exposure. RESULTS: The mean sister chromatid exchange frequency per metaphase was significantly higher in the study group compared to the control group: 8.11 +/- 2.39 vs. 6.33 +/- 1.60 (p = 0.008). We found that there was no positive correlation between the blood carboxyhemoglobin concentration and sister chromatid exchange frequency. CONCLUSIONS: These results suggest that acute exposure to combustion products of wood or coal is genotoxic to DNA. Potential causes of genotoxicity include known mutagenic compounds present in coal or wood smoke and ash, oxygen radicals formed during combustion, as well as hypoxic and reperfusion injury mechanisms initiated by carbon monoxide intoxication. Additional studies on separate carbon monoxide exposure from smoke and ash are needed to understand individual genotoxic contributions and mechanisms. PMID- 12126183 TI - The effect of the nitric oxide synthesis inhibitor L-NAME on amitriptyline induced hypotension in rats. AB - OBJECTIVE: Hypotension induced by tricyclic antidepressants is multifactorial. Previous animal experiments suggest a contribution from nitric oxide production. Our study aimed to evaluate the role of nitric oxide in amitriptyline-induced hypotension using N-nitro-L-arginine methyl ester, a nitric oxide synthesis inhibitor, and 3-morpholino sydnonimine, a nitric oxide donor, in anesthetized rats. METHODS: Amitriptyline intoxication was induced by the continuous infusion of amitriptyline 0.625 mg/kg/min throughout the experiment in anesthetized rats. Fifteen and 25 minutes after amitriptyline infusion began, two bolus doses of 10 mg/kg of N-nitro-L-arginine methyl ester (n = 8) or an equivalent volume of 5% dextrose solution (n = 8) was administered to each rat (Protocol 1). To investigate whether the effect of N-nitro-L-arginine methyl ester on blood pressure is counteracted by 3-morpholino sydnonimine, after the same protocol of amitriptyline infusion and 5 minutes after an N-nitro-L-arginine methyl ester bolus, a bolus of 3000 nmol/kg of 3-morpholino sydnonimine was administered (n = 8) to each rat (Protocol 2). To investigate the effect of N-nitro-L-arginine methyl ester on 3-morpholino sydnonimine induced hypotension, a group of rats received a continuous infusion of 0.54 mg/kg/h of 3-morpholino sydnonimine until 50% reduction was observed in mean arterial blood pressure followed by a bolus dose of 10 mg/kg of N-nitro-L-arginine methyl ester (n = 6) or 5% dextrose solution (n = 6) (Protocol 3). Outcome measures included mean arterial blood pressure, heart rate, and QRS duration in electrocardiogram. Student's t test and survival analysis were used for selected comparisons. RESULTS: For all parameters, the treatment groups were similar at baseline and at postamitriptyline periods before therapy was rendered. Amitriptyline infusion significantly reduced mean arterial blood pressure by 50.8 +/- 2.2% and prolonged QRS by 23.9 +/- 7.2% after 15 minutes. In Protocol 1, N-nitro-L-arginine methyl ester significantly increased mean arterial blood pressure compared to dextrose treated control animals within 30 minutes (77.9 +/- 8.5% vs. 49.7 +/- 5.0% mmHg, p < 0.01, 95% CI 57.1-98.7%). QRS duration progressively increased during the amitriptyline infusion; however, there was no significant difference in QRS width between N-nitro-L-arginine methyl ester and control groups at any time point. N nitro-L-arginine methyl ester increased survival time compared to controls (33.4 +/- 4.1 vs. 19.9 +/- 2.7 minutes, p < 0.01, 95% CI 25.4-41.3) but did not affect mortality. In Protocol 2 of continuous infusion of amitriptyline, 3-morpholino sydnonimine counteracted the N-nitro-L-arginine methyl ester-induced increase in mean arterial blood pressure. In both protocols, heart rate decreased significantly during amitriptyline infusion but there was no difference between treatment and control groups. In Protocol 3, N-nitro-L-arginine methyl ester bolus reversed 3-morpholino sydnonimine-induced hypotension compared to dextrose bolus. (83.8 +/- 5.7% vs. 54.6 +/- 4.8%, p < 0.01, 95% CI 69.2-98.4). CONCLUSION: N-nitro-L-arginine methyl ester is found to be effective in temporarily improving hypotension and prolonging survival time but does not affect overall mortality. Because this effect was antagonized by 3-morpholino sydnonimine, nitric oxide production appears to contribute to the pathophysiology of amitriptyline-induced hypotension. PMID- 12126184 TI - Trends in illicit drug emergencies: the emerging role of gamma-hydroxybutyrate. AB - BACKGROUND: Previously used as a general anesthetic, gamma-hydroxybutyrate is now used as a recreational drug. Not surprisingly, an increasing number of acute overdose cases requiring emergency medical care have been reported and described, especially in the United States. OBJECTIVES: To determine the number and percentage of gamma-hydroxybutyrate overdoses over a 15-month period and to describe the clinical hallmarks and course of this new drug in overdose. METHODS: All toxicological emergencies, including those caused by illicit drug consumption, were recorded for 15 months in an urban public hospital emergency department. Accurate toxicological history was obtained from the patients and, if gamma-hydroxybutyrate was suspected, confirmation was performed by urine mass spectrometry. The study data were compared with data recorded in the same emergency department in 1989. RESULTS: The total number of toxicological emergencies attended in our emergency department have remained unchanged during the last decade, with a significant decrease in number of opiate overdoses and an increase in the number of cocaine, amphetamine, and gamma-hydroxybutyrate overdoses. During the study period, 104 gamma-hydroxybutyrate overdoses presented to the emergency department (3.1% of all toxicological emergencies), ranking second in illicit drugs requiring emergency consultation. The profile of a patient with gamma-hydroxybutyrate intoxication is well defined: a young individual (23 +/- 5 years), male (64%), emergency department presentation on weekends (90%), with simultaneous ethanol consumption (73%) and ingestion of additional illicit drugs (86%), decrease of consciousness being the main complaint in all cases [16% with Glasgow Coma Scale (GCS) = 3]. Complete recovery without sequelae occurred in all cases. CONCLUSION: Health authorities must be aware of the hazards of recreational gamma-hydroxybutyrate, and physicians must be cognizant of this recent cause of coma among youths presenting to the emergency departments. PMID- 12126185 TI - Formate kinetics in methanol poisoning. AB - OBJECTIVE: We sought to describe the kinetics, dialysis clearance, and laboratory markers of formate (FA), the toxic metabolite of methanol (meOH). METHODS: Data were obtained from a prospective, multicenter study of fomepizole +/- dialysis for methanol poisoning. Inclusion criteria confirmed methanol exposure or suspicion of exposure plus either acidemia or abnormal osmolar gap. Dialysis indications were [meOH] > 50 mg/dL, pH < 7.1, refractory acidosis, or visual toxicity. Serial plasma formate, methanol, pH, and electrolyte measurements were made. Formate was determined by gas chromatography. Endogenous and dialysis elimination half-lives were calculated as t(1/2) = 0.693/Ke, with Ke (elimination constant) derived from the slope of log (FA) vs. time. Half-lives were compared with an unpaired Student's t-test. Dialysis clearance was calculated using the Fick Principle. Pearson correlation analysis compared initial formate with initial pH, serum bicarbonate, and anion gap. RESULTS: Eleven patients were treated in the study. Eight had detectable formate with mean [FA] of 15.1 mmol/L (range 0.5-34.8). Endogenous elimination half-life was 205 +/- 90 minutes. Elimination half-life during dialysis (n = 5) was 150 +/- 37 minutes, which was not different (t = 0.22; NS). The overall dialysis formate clearance rate was 223 +/- 25 mL/min. Correlation coefficients were: pH vs. formate r2 = 0.93; bicarbonate vs. formate r2 = 0.81; and anion gap vs. formate r2 = 0.76 (all p < 0.05). CONCLUSIONS: Although dialysis clears formate, it did not significantly enhance endogenous elimination in our series of patients. Low pH, low bicarbonate, and elevated anion gap correlate independently with formate presence. PMID- 12126186 TI - Clonidine toxicity revisited. AB - The incidence of clonidine overdose is increasing, yet there is a paucity of new information regarding treatment options for clonidine toxicity. Reported treatment approaches vary widely, demonstrating the lack of science on which current treatment is based. Available research needs to be reassessed. Neurotransmitters, receptors, endogenous opioids, and baseline sympathetic tone determine the clinical response to clonidine as well as the potential response to drug therapy following clonidine overdose. This article reviews aspects of clonidine toxicity that need to be further investigated. Multicenter research trials will be required to evaluate new treatment options. PMID- 12126187 TI - Development of a Chinese herbal medicine toxicology database. AB - BACKGROUND: Use of Chinese herbal medicine has increased steadily in the West and adverse reactions have been reported. However, information is not readily available on the toxicity of the herbs and the management of poisoning. The goals of this project were to retrieve and evaluate scientific evidence on the toxicity of Chinese herbal medicine, to grade the toxicity of individual herbs, and to summarize relevant herb data via a searchable electronic database. METHODS: English and Chinese primary studies were systematically retrieved via journal abstracting databases and key toxicity texts. Partial translation of published research was achieved via an audited process utilizing data extraction forms. Methods for grading herb toxicity (in therapeutic use or overdose) were developed based on a combination of the quality of reports, severity of adverse reaction, supporting animal studies, extrapolation from pharmacology and empirical evidence. RESULTS: Good quality studies on the toxicity of Chinese herbs are lacking. In some cases there is insufficient scientific evidence to create an evidence-based grading of the toxicity of individual herbs. Available data have been summarized into detailed monographs. Twelve herb monographs (with a full toxicity profile and grading) have been completed and summary data for all adequate quality papers used in the grading are linked to the monographs. CONCLUSIONS: The resultant database and monographs represent the first reviews specifically on the toxicity of individual Chinese herbs with toxicological grades based on the evidence of published studies. The database and monographs should assist in promoting the safe and effective use of Chinese herbal medicine. PMID- 12126188 TI - Chinese herbal medicine toxicology database: monograph on Herba Asari, "xi xin". PMID- 12126189 TI - Chinese herbal medicine toxicology database: monograph on Radix Sophorae Flavescentis, "ku shen". PMID- 12126190 TI - Clearance of metformin by hemofiltration in overdose. AB - BACKGROUND: Metformin is prescribed with an increasing frequency for patients with Type II diabetes mellitus; the increasing availability increases the risk of intentional overdoses. Metformin may cause severe lactic acidosis in overdose, especially when accompanied by co-ingestants or other medical conditions that alter lactate handling or metformin elimination. Though the clearance of therapeutic metformin by hemodialysis is known, the clearance in the setting of a large overdose has not been reported. CASE REPORT: A 58-year-old man with a history of Type II diabetes, hypertension, bipolar disease, and decreased renal function presented after ingestion of approximately 40 500-mg metformin tablets and 20 240-mg diltiazem sustained-release tablets. Clinical manifestations of poisoning included somnolence, hypotension, bradycardia, severe lactic acidosis, and ultimately death. Gastric decontamination was attempted with gastric lavage, multiple dose activated charcoal, and whole bowel irrigation. Hemodynamic support was provided with pressors, glucagon, insulin, and intra-aortic balloon pump. Due to hypotension, continuous renal replacement therapy, rather than hemodialysis, was initiated. Continuous veno-venous hemodialysis was performed with a blood flow of 180 mL/min and dialysate flow of 2.5 L/h. A Multiflow 60 kidney (Cobe) on a Prisma (Cobe) continuous renal replacement therapy machine was used. The initial metformin level was 110 microg/mL (therapeutic range 1-2 microg/mL). By continuous veno-venous hemodialysis, an absolute clearance of 50.4 mL/min was obtained. CONCLUSION: Metformin was cleared by the continuous veno-venous hemodialysis modality of continuous renal replacement therapy in this metformin overdose. Although a fatal outcome occurred in this patient, its utility in other patients with metformin overdose should be investigated. PMID- 12126191 TI - Acute cyanide intoxication treated with a combination of hydroxycobalamin, sodium nitrite, and sodium thiosulfate. AB - An 80-year-old diabetic patient was admitted to the hospital because of sudden unconsciousness and severe metabolic acidosis. His son reported the possibility of cyanide poisoning. Clinical data and the detection of cyanide in blood and gastric material confirmed this possibility. Supportive therapy and the following antidotes--sodium nitrite two doses 300 mg i.v., sodium thiosulfate 3 g i.v., and hydroxocobalamin 4 g in 24 hours--were administered immediately and the patient completely recovered in 48 hours. Our observations suggest that timely and appropriate use of antidotes for cyanide intoxication may prevent death, even in aged diabetic patients. PMID- 12126192 TI - Medicinal herb Erycibe henri Prain ("Ting Kung Teng") resulting in acute cholinergic syndrome. AB - A 73-year-old man consumed a decoction of the medicinal herb Erycibe henri Prain ("Ting Kung Teng"), as recommended in traditional Chinese medicine for arthritis. Shortly, he developed a cholinergic syndrome that included dizziness, diaphoresis, chills, lacrimation, salivation, rhinorrhea, nausea, and vomiting. He was also hypothermic and hypotensive. Notable laboratory values included a normal serum cholinesterase and transiently elevated blood urea nitrogen, creatinine, and glucose. There is no previous report on the toxicity due to this herb in the literature. Active constituents of the herb include a number of tropane alkaloids, one of which possesses cholinergic rather than anticholinergic activities. A study conducted on mice, with a related herb, has demonstrated renal, hepatic, and erythrocyte toxicity. PMID- 12126193 TI - The Delphic oracle: a multidisciplinary defense of the gaseous vent theory. AB - Ancient historical references consistently describe an intoxicating gas, produced by a cavern in the ground, as the source of the power at the oracle of Delphi. These ancient writings are supported by a series of associated geological findings. Chemical analysis of the spring waters and travertine deposits at the site show these gases to be the light hydrocarbon gases methane, ethane, and ethylene. The effects of inhaling ethylene, a major anesthetic gas in the mid 20th century, are similar to those described in the ancient writings. We believe the probable cause of the trancelike state of the Priestess (the Pythia) at the oracle of Delphi during her mantic sessions was produced by inhaling ethylene gas or a mixture of ethylene and ethane from a naturally occurring vent of geological origin. PMID- 12126194 TI - Faults suggest a high calling for Delphi priestesses. PMID- 12126195 TI - Nosocomial poisoning: is third party cholinesterase poisoning anecdotal? PMID- 12126197 TI - Resurgence of a lethal drug: paramethoxyamphetamine deaths in Belgium. PMID- 12126196 TI - Nosocomial poisoning: organophosphate contamination of emergency room staff is real. PMID- 12126198 TI - Acute tacrolimus overdose without significant toxicity. PMID- 12126199 TI - Platelet function testing in apheresis products: flow cytometric, resonance thrombographic (RTG) and rotational thrombelastographic (roTEG) analyses. AB - During storage of platelet concentrates, quality control of the units is mandatory. This includes the important testing of the hemostatic function of platelets. So far, mostly platelet aggregation analyses have been performed. In this study, new approaches were tested to evaluate the applicability of modern techniques for quality monitoring. Plateletpheresis was performed with two different cell separators (AMICUS cell separator, Fenwal, Baxter Healthcare, Deerfield, USA; COBE Spectra, COBE BCT, Lakewood, USA). In each procedure split products (n = 22) were prepared and stored for 1-2 days (n = 22) or 3 5 days (n = 22). Platelet hemostatic capacity was tested by applying flow cytometry. platelet aggregation (platelet-rich-plasma [PRP]+agonist), resonance thrombography (RTG; PRP, no agonist) and rotational thrombelastography (roTEG; PRP+agonist). Flow cytometric analyses did not reveal significant changes in structural (CD41a. CD42b) or activation-dependent antigens (CD62p, CD63, LIBS, RIBS). Also, differences in the data from the flow cytometric reactivity tests were not significant between the two groups. In platelet aggregation assays, shape change (p = 0.8), maximum aggregation (p = 0.4), and maximum gradient (p = 0.8) did not show significant differences between the two groups. In the RTG test, differences between r-time (reaction time; p = 0.4), and f-time (clot formation time [fibrin influence]; p = 0.3), and in roTEG r-time (coagulation time; p = 0.1) and k-time (clot formation time; p = 1.0) were not significant. P-time (clot formation time [platelet influence]) and M (maximum amplitude) in RTG, and k-time and MA (maximum amplitude) in roTEG showed a slight decrease in platelet function (p < or = 0.05). We conclude that platelet function is well maintained during storage. This is reflected by the results of immunological and platelet function assays. Rotational thrombelastography (in the case of PRP) and especially resonance thrombography represent promising methods for quality control of platelet concentrates and rapidly provide information about the status of platelet function and the whole clotting process. PMID- 12126200 TI - Investigation of transfusion transmitted viruses in cases clinically suspected of posttransfusion hepatitis with undetermined ethiology. AB - Transfusion transmitted viruses (TTV) were investigated in cardiac surgery cases who were previously transfused with blood and/or blood products and were suspected of having posttransfusion hepatitis (PTH) based on the results of physical examination, clinical findings, biochemical blood test results and in a smaller number, on radiological results. They were identified as having non-A-C hepatitis based on serological or molecular test methods. In this study, out of 90 cases suspected for PTH and non-A-C, 78 (86.7%) were male, 12 (13.3%) were female and their ages were between 17 and 67. Ninety healthy blood donors, who donated blood for the first time and had never had a transfusion, were selected as the control group. They had alanine aminotransferase (ALT) levels < 40 U, were seronegative for hepatitis B virus (HBV) and hepatitis C virus (HCV). Seventy seven were immune, and 13 were seronegative for hepatitis A virus (HAV). In this study, TTV-deoxyribonucleic acid (DNA) investigation was performed by the polymerase chain reaction (PCR) method suggested by Takahashi et al. with 5' GCT ACG TCA CTA ACC ACG TG 3' (T801) and 5' CTG CGG TGT GTA AAC TCA CC 3' (T935) primers. TTV-DNA was found to be positive in 21 (23.3%) of the patient group and 4 (4.4%) of the control group (p < 0.05). In the patients determined to be TTV DNA positive, the admission time following transfusion was a minimum of 3, and a maximum of 15 (average 7) weeks. The average ALT levels detected at the time of admission did not show a difference between TTV-DNA positive and negative cases (p > 0.05). However the ALT levels had a tendency to rise and reached their highest level nine weeks after transfusion in the TTV-DNA positive cases, although in two cases the ALT levels decreased to normal value after the 13th week. During the 24 month follow up of the TTV-DNA positives all cases except one were positive at the end of this period. The results of this study are the same as those reported in the literature suggesting that TTV-DNA, excluding the main viral agents which are known to cause PTH, can be determined in transfused PTH or non-transfused asymptomatic patients in varying ratios. In order to define the epidemiological properties and hepatic-extrahepatic pathologies more clearly we have looked for evidence of the viral agent, which probably contaminates both by transfusion and non-transfusion routes. It is suggested that, in addition to the case groups in this study, new clinical studies are necessary including transfused but non-PTH patients. PMID- 12126202 TI - Evaluating pediatric apheresis toxicity. PMID- 12126201 TI - Lymphocyte subsets in inline filtered packed red blood cell units: comparison between low and high spin procedures. AB - Lymphocyte subsets were determined in 20 packed red blood cell units (PRC) before and after filtration (FPRC) with the Pall Leukotrap RC inline filter system; 10 units were prepared by low spin and platelet rich plasma (PRP) removal (Group A) and 10 with high spin, plasma and buffy-coat (BC) removal (Group B). Flow cytometry was employed for white blood cell (WBC) enumeration and phenotype analysis. Median WBCs in prefiltered units was 2.08 x 10(9) (Group A) vs. 0.8 x 10(9) (Group B) (p < 0.0001). Five Group A and three Group B filtered units had WBC counts above the limit of detection (LD), median values being 25.59 and 3.08 x 10(3), respectively. Whereas CD3+, CD3+CD4+ and CD3+CD8+ lymphocyte subsets were assessable in 20-40% of Group A units, inline filtration of Group B units lowered lymphocytes below the LD of the present study. Post-filtration CD19+ lymphocytes were below the LD in all the 20 units. PMID- 12126203 TI - Transimmunization: the science catches up to the clinical success. PMID- 12126204 TI - Transimmunization and the evolution of extracorporeal photochemotherapy. AB - We are now aware that extracorporeal photopheresis (ECP) - in which a patient's leukocytes are isolated, passed through an ultrathin clear plastic plate, and exposed to 8-methoxypsoralen (8-MOP) and ultraviolet A light prior to reinfusion is a simple and efficient dendritic cell (DC) therapy and the first FDA approved selective immunotherapy for cancer. DCs, as the most effective antigen presenting cells (APCs), are central to many ongoing efforts to stimulate immune responses to cancer cells. Moreover, ECP has not only demonstrated efficacy in the treatment of a T cell malignancy--namely cutaneous T-cell lymphoma (CTCL)--but also in treatment of oligoclonal T-cell-mediated diseases such as graft-versus host-disease (GVHD) and organ transplant rejection. Recent advances in the understanding of DC/T-cell interactions provide insight into how ECP-induced DCs (EI-DCs) can be utilized to stimulate specific T-cell (i.e. anti-tumor) responses, or down-regulate a pre-existing potent T-cell response. The mechanism of this apparent paradox of EI-DC functionality is likely dependent on several fundamental principles: (1) the status of existing in vivo T-cell reactions, (2) the temporal stage of EI-DC differentiation, and (3) the affinity of the available repertoire of T-cell receptors (TCRs) for the antigen(s) in question. Further investigation into DC/T-cell interactions will help to shape the future of ECP and the ability to optimize this therapy for the desired immune effect. To this end, we are developing and testing Transimmunization to replace conventional ECP. PMID- 12126205 TI - T cell clonality and the effect of photopheresis in systemic sclerosis and graft versus host disease. AB - Photopheresis, a leukapheresis-based therapy that combines 8-methoxypsoralen (8 MOP) and ultraviolet-A irradiation, has been shown to be an effective treatment for advanced cutaneous T cell lymphoma (CTCL), but also appears to be effective in certain patients with systemic sclerosis (SSc) and chronic graft versus host disease (cGVHD). In CTCL, clinical responsiveness to photopheresis has been shown to be dependent on the presence of detectable circulating clonal T cells in peripheral blood. Herein we review recent work from our group demonstrating that circulating clonal populations of T cells are detectable in a subgroup of patients with SSc and cGVHD. Furthermore, we provide initial evidence that the presence of a T cell clone in such patients may be associated with responsiveness to photopheresis. PMID- 12126206 TI - Extracorporeal photochemotherapy: a new therapeutic approach for allograft rejection. AB - Photopheresis (ECP) is a new immunomodulatory therapy in which recipient lymphocytes are treated extracorporeally with 8-methoxypsoralen (8-MOP) and ultraviolet light. The treatment seems to induce an inhibition of both umoral and cellular rejections after transplantation. More than 160 transplanted patients have been treated with ECP (107 heart, 30 kidney, 24 lung and I liver) in different studies. Indication for ECP included acute rejection, recurrent/refractory rejection, prophilaxis of rejection, need of reducing standard immunosuppression. Patient survival is satisfactory. Only one study where ECP was used as the last therapeutic resource in very compromised patients shows a high rate of mortality. On the contrary, when ECP was used earlier after the failure of a first immunosuppressive line the outcome was better with a very low mortality. An hystological resolution of acute rejection is reported in 89% of cardiac transplant patients. The rate of response is similar even in the other transplanted patients treated with ECP. A better control of alloreactivity has been also reported in both cardiac and renal transplant patients with recurrent rejection. In renal allograft the treatment induces a reduction of both lymphocytes and monocytes infiltrate and downregulates the expression of HLA-DR and integrins ICAM-1 and VCAM-1 on tubular cells. Markers of fibrogenesis such as TGFbeta1 and ASMA are only moderately reduced with a more focal pattern of distribution in the post-ECP specimens. The optimal schedule and the length of treatment are still unclear and probably a patient-tailored treatment is needed at least in responder patients. ECP is effective for patients resistant to conventional treatments, particularly when it is started early. This beneficial effect is obtained without the complications typically encountered with immunosuppressive regimens used to control rejection. PMID- 12126207 TI - Transimmunization, a novel approach for tumor immunotherapy. AB - This review describes our experience with the development of a novel form of immunotherapy that may represent the first practical and effective means of performing tumor-loaded dendritic cell (DC) immunotherapy. We have modified the highly successful extracorporeal photopheresis (ECP) treatment that has been used in the therapy of cutaneous T cell lymphoma (CTCL). autoimmune disease, transplantation rejection episodes and graft-versus-host disease to enhance its efficacy by the addition of an overnight incubation period. This adaption of ECP is termed "transimmunization (TI)" since the new therapy permits transfer of tumor antigens that have been previously poorly recognized to potent antigen presenting cells where the tumor epitopes can be displayed in the full context of major histocompatibility, co-stimulatory and adhesion molecules. The TI modification of ECP is a practical and safe means of rapidly inducing DC differentiation from peripheral blood monocytes in the presence of apoptotic tumor cells. Uptake of the apoptotic CTCL cells by the immature DC, in the presence of inflammatory cytokines, further drives their maturation into potent antigen presenting cells. Reinfusion of these tumor-loaded DC, that have access to the full spectrum of tumor antigens, has the potential to invoke an anti-tumor immune response in the recipient. Standard ECP has been a very useful form of immunotherapy and a modification of this approach that can enhance its ellicacy and utility should broaden its application to a larger variety of disorders including potentially the treatment of solid tumors and the modulation of the immune response in graft-versus-leukemia and graft-versus-host transplantation regimens. An understanding of the mechanism of ECP and TI will provide the physician with the ability to more finely tune the desired immune response and thereby, provide an enhanced immunotherapy for malignancy and other disorders of immunocompetence. PMID- 12126208 TI - Assessment of the future resources and needs for hospitalization in Hong Kong SAR (Special Administrative District). AB - OBJECTIVES: To study the 'compression of morbidity' theory in the Hong Kong SAR by analysing the age and gender-specific hospitalization rates and the expected length of stay per patient for the period 1996-2000. 'Compression of morbidity' refers to the hypothesis that medical progress will reduce the duration of morbidity during life more significantly than increasing morbidity by extending life. DESIGN: This is a retrospective study based on hospital admissions data from the Hospital Authority of Hong Kong which covers 93% of the patient population. SETTING: Age and gender-specific hospitalization rates, expected length of stay and hospitalization needs for each specific age group in Hong Kong from 1996 to 2000, are estimated. MAIN RESULTS: There is no empirical support for compression theory; and there is no significant change in the hospital admission rates for the period 1996-2000. The total number of patient days is expected to increase by 80% because of the ageing effect alone. It is projected that the geriatric service will account for more than 60% of the hospital patient days utilization in 2029. The elderly dependency ratio will increase and the social burden for the next generation will be increased, as the working populations size continues to decrease due to low fertility in the Hong Kong SAR. CONCLUSION: The health care burden on the government is large and increasing. It is therefore essential to make plans to deal with the ageing population, which is predicted to be at its highest in 2020. The rising effect of public expectations on hospital services exerts further pressure on demand. PMID- 12126209 TI - Decentralization, health care and policy process in the Punjab, Pakistan in the 1990s. AB - The Province of the Punjab underwent a number of attempts to decentralize the health sector in the 1990s. Among the most important were the decentralization of financial management within the district, the Sheikhupura PHC Pilot Project, the establishment of the District Health Authorities and District Health Management Teams, the creation of semi-autonomous hospitals and the programme of District Health Government (DHG). These usually received donor support and promotion, and emerged from within the provincial Department of Health, and more specifically the Secretariat and the internationally supported Second Family Health Project (FH2). Of particular significance was the DHG change, which involved a decentralization to the district, the appointment of powerful Chief Executives, the formation of District Management Committees and purchaser-provider separation. The paper reviews these proposals, focusing on the need to build on experiences and learning lessons from pilot projects, reform continuity, developing consultation and involvement and policy analysis. The latter indicates the importance of developing more in-depth policy analysis around the role of the central organization, the form of decentralization and the purchaser-provider separation. The paper concludes by underlining the need to ensure that political strategy and in-depth policy are appropriately coordinated in the policy process. PMID- 12126210 TI - Informal risk-sharing arrangements (IRSAs) in rural Burkina Faso: lessons for the development of community-based insurance (CBI). AB - In resource-poor environments, community-based insurance (CBI) is increasingly being propagated as a strategy to improve access of poor rural populations to modern health care. It has been repeatedly hypothesized that CBI schemes need to be grounded in national as well as local traditions of solidarity. This paper presents a typology of informal risk sharing arrangements (IRSAs) in a rural area of North-Western Burkina Faso and discusses their modus operandi as well as the underlying concepts of solidarity and reciprocity. The research was explicitly multi-disciplinary, combining anthropological and economic as well as qualitative and quantitative data collection methods. Focus group and interview data were complemented by a census of existing IRSAs. In addition to presenting the main features of existing institutions, the paper discusses whether IRSAs can serve as entry points for CBI schemes. In spite of the fact that existing IRSAs fulfil important solidarity functions in the rural Burkinian context, we conclude that they cannot serve as institutional models for more formalized CBI schemes. Community participation in a future CBI scheme will need to tap into existing notions of solidarity and mutuality. The CBI scheme itself, however, needs to be newly tailored. PMID- 12126211 TI - A development framework for promoting evidence-based policy action: drawing on experiences in Sri Lanka. AB - Most developing countries have embarked on one form or another of 'health sector reform' as a result of the global trend for health and health care reform that has emerged during the past decade. One consequence is that the issue of health sector performance is moving higher on the agenda of many developing countries, and particularly that of the corporate performance of health sector staff. Along with this movement has come increased attention to strengthening evidence-based management decision-making. To date, studies on measuring health sector performance, have had little impact on developing country health systems and have been limited to explorations primarily at an operational level. However, there is a growing recognition that there is a need to strengthen the policy function of ministries and their ability to monitor policy impact. Sri Lanka is one country that has identified the need to strengthen policy at national level. Many developing countries, like Sri Lanka, are familiar with input, process and output dimensions of operational performance. However, most are not ready to engage in routine performance assessment that can strengthen policy processes at national level. This paper explores (1) the implication and the use of indicators to support evidence based policy decision-making, and (2) the complexity of doing so in Ministries of Health that are undergoing some form of health sector reform. The paper emphasizes that new forms of organizational support are required for performance management at policy level. A conceptual framework for managing the collection and use of performance evidence is developed, including proposals for the introduction of outcome indicators into that process. The paper concludes with recommendations on initiatives required to develop appropriate organizational and technical capacity to engage with performance management at policy level and for further research towards creating ministries of health as 'learning organizations' that can change and adapt with informed decisions. PMID- 12126212 TI - Survey of the medical financial assistance schemes of the urban poor in Shanghai. AB - After 1990, being a 'pilot city' for health care reform, Shanghai constructed modernized medical financial assistance schemes (MFAS), which were progressively applied by the central government rate in the 1990s. As the pioneer of social security reform in China, Shanghai again is the place with most experience of the problems of the transition from 'Public-Relief' to 'Social-Assistance'. From the 'user' perspective of MFAS, the study reported has attempted to evaluate MFASs. Furthermore, the research seeks to point to future policy and programme options for implementing MFAS and optimizing the allocation of urban health care resources. PMID- 12126213 TI - Risk markers for thrombocytopenia in critically ill patients: a prospective analysis. AB - STUDY OBJECTIVE: To identify independent risk markers for thrombocytopenia in critically ill patients. DESIGN: Prospective, observational study. SETTING: Eleven-bed intensive care unit-coronary care unit (ICU-CCU) in a community hospital. PATIENTS: Three hundred sixty-two consecutive patients meeting inclusion criteria during 1 year. INTERVENTION: Potential risk marker data were collected on admission to the ICU-CCU and for the period before development of thrombocytopenia (defined as two or more consecutive platelet counts < 150 x 10(3)/mm3 obtained at least 12 hours apart), or for the duration of ICU-CCU stay if thrombocytopenia did not develop. MEASUREMENTS AND MAIN RESULTS: Thrombocytopenia developed in 68 patients (18.8%). Multivariate logistic regression analyses identified patients at risk on admission, but the predictive, potential of the regression model improved when all risk marker exposures during the ICU-CCU stay were considered. Independent risk markers included fresh frozen plasma administration, sepsis, musculoskeletal diagnosis, pulmonary artery catheter insertion, gastrointestinal diagnosis, packed red blood cell administration, and nonsurgical respiratory diagnosis. Higher admission platelet count and aspirin administration were associated with a lower risk of thrombocytopenia. Heparin administration was not identified as a risk marker, and no patient developed heparin-induced thrombocytopenia with thrombosis. Patients with thrombocytopenia had longer ICU-CCU and hospital stays, and higher ICU-CCU and hospital mortality than those without thrombocytopenia. CONCLUSIONS: Development of thrombocytopenia in critically ill patients is associated with specific diagnoses, packed red cell and fresh frozen plasma transfusions, pulmonary artery catheter insertion, and admission platelet count. PMID- 12126214 TI - Effects of orlistat on fat-soluble vitamins in obese adolescents. AB - STUDY OBJECTIVES: To determine whether orlistat causes fat-soluble vitamin deficiencies in African-American and Caucasian adolescents. DESIGN: Prospective, open-label pilot study. SETTING: Warren Grant Magnuson Clinical Center of the National Institutes of Health. PATIENTS: Seventeen adolescents with body mass indexes above the 95th percentile for age, race, and gender who also had at least one obesity-related comorbid condition. INTERVENTION: Subjects received orlistat 120 mg 3 times/day and a daily multivitamin supplement containing vitamin A 5000 IU, vitamin D 400 IU, vitamin E 300 IU, and vitamin K 25 microg. MEASUREMENTS AND MAIN RESULTS: During 3-6 months of orlistat treatment, acute absorption of retinol (vitamin A) was not significantly altered, but absorption of alpha tocopherol (vitamin E) was significantly reduced compared with baseline levels (p<0.001). Serum levels of vitamins A and E did not change significantly; however, there was a nonsignificant decrease in vitamin K. Mean vitamin D levels were significantly reduced compared with baseline (p<0.02) after 1 month of orlistat, despite multivitamin supplementation. CONCLUSION: It may be prudent to monitor vitamin D concentrations in adolescents who take orlistat, even when a multivitamin is prescribed. PMID- 12126215 TI - Tachyphylaxis associated with continuous cisatracurium versus pancuronium therapy. AB - STUDY OBJECTIVES: To compare dosing requirements over time among patients receiving continuous cisatracurium versus pancuronium therapy, and to identify factors that may account for changes in pancuronium versus cisatracurium infusion requirements over time. DESIGN: Retrospective, comparative cohort analysis. SETTING: A tertiary level 1 trauma center. PATIENTS: Forty-five consecutive adult patients who were admitted to intensive care units at our institution from January 1998-August 2000 and received continuous cisatracurium or pancuronium therapy for at least 48 hours. MEASUREMENTS AND MAIN RESULTS: Dosing requirements of patients treated with pancuronium or cisatracurium were recorded over time throughout the treatment period. Factors that could affect dosing requirements of a neuromuscular blocking agent (NMBA) were stratified as time invariant (admitting service, acute physiology and chronic health evaluation II score, duration of mechanical ventilation, pressure control ventilation, baseline hepatic or renal insufficiency, thermal injury, train-of-four assessment, and concurrent drug administration or disorders affecting neuromuscular transmission) or time variant (concurrent sedation and narcotic analgesia therapy; serum magnesium, potassium, and creatinine concentrations; arterial pH level; temperature; peak airway pressure; and partial pressure of oxygen:fraction of inspired oxygen ratio). Hierarchical linear modeling was used to compare the dosing requirements and to identify confounders affecting the relationship. The infusion rate escalation for the cisatracurium group was greater (0.39 microg/kg/min; 95% confidence interval [CI] 0.22-0.56; 23 patients) than for the pancuronium group (-0.06 microg/kg/min; 95% CI -0.24-0.12; 22 patients; p<0.001) and was associated with an average daily cost/patient significantly higher (p<0.001) with cisatracurium ($258+/-$114) than pancuronium ($11+/-$5). Confounder analysis revealed that only the admitting service and the number of times the NMBA infusion was suspended because no twitch was detected differed between groups. Neither of these confounders significantly affected the temporal relationship between cisatracurium and pancuronium infusion rates. CONCLUSION: Dosing requirements increase over time at a significantly greater rate for cisatracurium than pancuronium infusions. Tachyphylaxis with cisatracurium is associated with substantial drug-related costs and is not accounted for by various disease-, patient-, and therapy-related factors. Further investigation is required to elucidate the mechanisms and risk factors underlying this phenomenon. PMID- 12126217 TI - Nonsteroidal antiinflammatory drug starter packs for chronic musculoskeletal pain. AB - STUDY OBJECTIVE: To determine whether prescribing a nonsteroidal antiinflammatory drug (NSAID) starter pack for chronic musculoskeletal pain expedites the process of finding an appropriate drug for a given patient. DESIGN: Prospective patient interviews. SETTING: Veterans Affairs Medical Center. PATIENTS: Sixty-four patients with chronic musculoskeletal pain were prescribed NSAID starter packs. Of those, 42% were interviewed and their data evaluated. INTERVENTION: Between March and June 2001, patients received starter packs containing 1-week supplies of the following NSAIDs: ibuprofen, salsalate, etodolac, naproxen, sulindac, and piroxicam. The patients took one drug each week, then returned to their providers to receive a prescription for the agent that was considered most effective and tolerable. MEASUREMENTS AND MAIN RESULTS: Patients assessed pain each day based on a numeric pain-rating scale. During telephone interviews, seven patients reported better pain control when they were able to select a drug from the starter pack than when they were prescribed a specific drug by their providers. Providers rated the starter pack as easy to use by patients and generally effective for finding the best NSAID for a particular patient. Drugs prescribed after completing the starter pack were salsalate 25.9%, piroxicam 22.2%, etodolac 14.8%, ibuprofen 14.8%, naproxen 11.1%, celecoxib 7.4%, and an opiate 3.7%. CONCLUSION: The NSAID starter pack appears to be a successful method for quickly and easily finding an NSAID that is effective and tolerated. PMID- 12126216 TI - Topiramate and weight loss in patients with neurodevelopmental disabilities. AB - STUDY OBJECTIVE: To characterize weight changes in patients with neurodevelopmental disabilities who received topiramate. DESIGN: Retrospective, observational study. SETTING: State-supported developmental center. PATIENTS: Fifteen patients with neurodevelopmental disabilities who received topiramate therapy MEASUREMENTS AND MAIN RESULTS: Monthly weights for 1 year after the start of topiramate therapy were recorded. Mean body weight at drug initiation differed significantly compared with the nadir weight during the next 12 months (52.2+/ 7.5 vs 49.9+/-7.2 kg, p<0.02). Twelve patients who were receiving ad libitum oral diets demonstrated a significant decrease in body weight (52.1+/-7.5 vs 49.0+/ 7.3 kg, p<0.01), whereas the three patients who received enteral nutrition through enterostomy did not experience significant weight loss. CONCLUSION: Weight loss is common in patients with neurodevelopmental disabilities who receive topiramate. Since patients who received oral diets lost weight whereas those receiving enteral nutrition did not, decreased nutrient intake is the likely cause of weight loss. PMID- 12126218 TI - Olanzapine-associated diabetes mellitus. AB - STUDY OBJECTIVE: To explore the clinical characteristics of hyperglycemia in patients treated with olanzapine. DESIGN: Retrospective, epidemiologic survey of spontaneously reported adverse events related to olanzapine therapy SETTING: Government-affiliated drug evaluation center. PATIENTS: Two hundred thirty-seven patients with olanzapine-associated diabetes or hyperglycemia. INTERVENTION: One hundred ninety-six cases from January 1994-May 15, 2001, were identified with the United States Food and Drug Administration's MedWatch Drug Surveillance System, and 41 cases published through May 15, 2001, were identified with MEDLINE or through meeting abstracts. MEASUREMENTS AND MAIN RESULTS: Of the 237 cases, 188 were new-onset diabetes, 44 were exacerbations of preexistent disease, and 5 could not be classified. Mean patient age for newly diagnosed cases was 40.7+/ 12.9 years and male:female ratio was 1.8. Seventy-three percent of all cases of hyperglycemia appeared within 6 months of start of olanzapine therapy. Eighty patients had metabolic acidosis or ketosis, 41 had glucose levels of 1000 mg/dl or greater, and 15 patients died. When olanzapine was discontinued or the dosage decreased, 78% of patients had improved glycemic control. Hyperglycemia recurred in 8 of 10 cases with rechallenge. CONCLUSIONS: Number of reports, temporal relationship to start of olanzapine therapy, relatively young age, and improvement on drug withdrawal suggest that olanzapine may precipitate or unmask diabetes in susceptible patients. PMID- 12126219 TI - New cholesterol guidelines, new treatment challenges. AB - Coronary heart disease (CHD) is the largest single killer of Americans. Epidemiologic trials have indicted low-density lipoprotein cholesterol (LDL) as directly correlating with CHD events, and clinical trials confirmed that lipid lowering therapy decreases the risk of CHD events. The literature also indicates that only about 18% of these patients are treated to their goal LDL levels. New guidelines from the National Cholesterol Education Program extend the LDL lowering recommendations, add a new non-high-density lipoprotein cholesterol (non HDL) goal for patients with hypertriglyceridemia, and increase the number of drug eligible patients from about 15 to 36 million. Most of those who are eligible for lipid-altering drug therapy have the highest CHD risk and require the most aggressive treatment to achieve goals. This presents a challenge to clinicians: how best to achieve LDL and non-HDL goals. Statins are the most effective agents for lowering LDL and one of the most effective for lowering non-HDL. This efficacy and the ability to reduce CHD risk were well documented in a number of randomized clinical trials. When statin monotherapy fails to achieve goals, a bile acid resin, niacin, or both may be added to lower LDL further, or a fibrate, niacin, or fish oils may be added to lower non-HDL further. Drugs are under development that may enhance our ability to reach LDL and non-HDL goals. Included are a group of so-called super statins, rosuvastatin and pitavastatin; agents that interfere with cholesterol or bile acid transport in the gut, such as ezetimibe; and dual peroxisome proliferator-activated receptor agonists that have both fibrate and thiazolidinedione-like effects. PMID- 12126220 TI - Administration of glycoprotein IIb-IIIa inhibitors in patients with ST-segment elevation myocardial infarction. AB - Patients with ST-segment elevation acute myocardial infarction require immediate reperfusion therapy. Reperfusion therapy can be provided by either pharmacologic or mechanical means. Pharmacologic reperfusion therapy consists of administering fibrinolytics, whereas mechanical reperfusion consists of performing percutaneous intervention, usually with stent placement. Each approach has been shown to decrease mortality, but each has disadvantages in establishing flow in the infarct-related artery. Regardless of the approach, during an acute myocardial infarction, activation and externalization of glycoprotein (GP) IIb-IIIa receptors occur on the surface of platelets. The GP IIb-IIIa inhibitors block the binding of fibrinogen to these platelet receptors. These inhibitors have been investigated in combination with both reperfusion strategies. The goal of adding GP IIb-IIIa inhibitor therapy to either reperfusion approach is to obtain better early, complete, and sustained reperfusion. Subsequently, this should lead to better clinical outcomes for patients with ST-segment elevation acute myocardial infarction. Although no mortality benefit has been seen with the addition of GP IIb-IIIa inhibitor therapy, ischemic complications have been reduced significantly. PMID- 12126221 TI - Darbepoetin-alpha: a review of the literature. AB - Anemia is a chronic condition that affects many patients with chronic kidney disease (CKD). These patients often require recombinant human erythropoietin (rHuEPO) to stimulate bone marrow to produce red blood cells. The agent often has to be administered 2-3 times/week for maximum efficacy A new product, darbepoetin alpha, is a hyperglycosylated erythropoiesis-stimulating protein that has a longer terminal half-life than rHuEPO (25.3 vs 8.5 hrs), which allows for less frequent dosing. At an equivalent dosage as rHuEPO, darbepoetin-alpha maintains hemoglobin values within target range and has a similar adverse effect profile. It is safe and effective for treatment of anemia of CKD. Pharmacoeconomic and quality-of-life studies are warranted to determine the compound's overall benefit. PMID- 12126222 TI - Safety and efficacy of controlled-release oxycodone: a systematic literature review. AB - Prescriptions for controlled-release oxycodone, a narcotic analgesic, recently contributed to a dramatic increase in pharmacy costs for a large private insurance company. To determine whether this agent offered clinical benefits over other available drugs that would justify its significantly greater cost, a systematic review of 16 clinical trials was undertaken. The review suggested that immediate-release and controlled-release preparations of oxycodone have similar efficacy and comparable side effect profiles. Controlled-release oxycodone has the advantage of less frequent dosing than immediate-release oxycodone; however, other agents may be dosed infrequently at much lower costs. For patients requiring a controlled-release opioid treatment, controlled-release morphine and methadone should be considered because they appear to be as effective as oxycodone and cost considerably less. Controlled-release oxycodone may be appropriate for some patients, particularly if they cannot tolerate other controlled-release or long-acting opioid analgesics. PMID- 12126223 TI - Secretin treatment for autistic disorder: a critical analysis. AB - We assessed evidence of the effects of secretin on behavior in individuals with autistic disorder. Articles were obtained through a MEDLINE search of the English language literature from January 1966-November 2001; all investigations and case reports on the topic were included. Press releases obtained from the World Wide Web also were included. Secretin, a gastrointestinal hormone, is suggested to improve autistic symptoms, particularly social function and communication. Two formulations, porcine and synthetic human secretin, were evaluated in humans. A small body of literature and popular belief in autistic disorder communities supported the agent's efficacy. A number of controlled clinical trials did not show improvement in autistic symptoms with secretin compared with placebo, possibly indicating no role for the drug in autistic disorder. PMID- 12126224 TI - Drug-related visits to the emergency department: how big is the problem? AB - OBJECTIVES: To review the literature concerning drug-related problems that result in emergency department visits, estimate the frequency of these problems and the rates of hospital admissions, and identify patient risk factors and drugs that are associated with the greatest risk. METHODS: A systematic search of MEDLINE (January 1966-December 2001), EMBASE (January 1980-December 2001), and PubMed (January 1966-December 2001) databases for full reports published in English was performed. The Ottawa Valley Regional Drug Information Service database of nonindexed pharmacy journals also was searched. RESULTS: Data from eight retrospective and four prospective trials retrieved indicated that as many as 28% of all emergency department visits were drug related. Of these, 70% were preventable, and as many as 24% resulted in hospital admission. Drug classes often implicated in drug-related visits to an emergency department were nonsteroidal antiinflammatory drugs, anticonvulsants, antidiabetic drugs, antibiotics, respiratory drugs, hormones, central nervous system drugs, and cardiovascular drugs. Common drug-related problems resulting in emergency department visits were adverse drug reactions, noncompliance, and inappropriate prescribing. CONCLUSION: Drug-related problems are a significant cause of emergency department visits and subsequent resource use. Primary caregivers, such as family physicians and pharmacists, should collaborate more closely to provide and reinforce care plans and monitor patients to prevent drug-related visits to the emergency department and subsequent morbidity and mortality. PMID- 12126225 TI - Thiazolidinedione-induced edema. AB - Edema is an adverse event associated with thiazolidinedione therapy The potential for mild-to-moderate peripheral edema with thiazolidinedione is known, especially in patients who have heart failure or use insulin. Our experience reveals that patients who do not have heart failure or do not use insulin also can develop moderate-to-severe edema that necessitates discontinuation of the thiazolidinedione. A 77-year-old man developed ankle, hand, and facial swelling 2 weeks after starting rosiglitazone. After discontinuing the drug, his edema resolved. A 75-year-old man developed bilateral lower leg edema 6 months after switching from troglitazone to pioglitazone. He was hospitalized for 51 days and, after aggressive diuresis and discontinuation of pioglitazone, was discharged with a weight loss of 30 pounds. A 53-year-old man developed lower leg edema 4 weeks after rosiglitazone was increased from 4 mg once/day to 4 mg twice/day. Rosiglitazone was discontinued and the edema resolved. Until the mechanism of action responsible for fluid overload is known, we suggest that thiazolidinediones be administered with caution in all patients. PMID- 12126226 TI - Central nervous system adverse effects with efavirenz: case report and review. AB - Efavirenz is a nonnucleoside reverse transcriptase inhibitor that can be given with other antiretroviral agents for the treatment of human immunodeficiency virus infection. A 47-year-old man with acquired immunodeficiency syndrome developed severe depression and suicidal ideation necessitating psychiatric hospitalization and antidepressant therapy. The symptoms occurred in temporal relation to the introduction of efavirenz into his highly active antiretroviral therapy regimen. Similar serious psychiatric adverse effects have been associated with this agent. Clinicians should monitor for central nervous system adverse effects in all patients taking efavirenz. PMID- 12126227 TI - A casting call from industry: reel in and retain appropriate information, release the rest. PMID- 12126228 TI - Molecular complexity of vertebrate tight junctions (Review). AB - Separation of distinct body, organ and tissue compartments, and maintenance of epithelial cell polarity require tight junctions (TJ)-cell-cell junctions located in the apicolateral regions of epithelial and endothelial cells. Studies on the protein components of vertebrate TJ have revealed an intricate network of membrane, sub-membrane, cytoskeletal, and signalling molecules. How these molecules functionally interact to provide TJ with their functions, and what roles these molecules play in control of cell growth and differentiation is a fundamental problem in cell biology. PMID- 12126230 TI - Gap junctions: structure and function (Review). AB - Gap junctions are plasma membrane spatial microdomains constructed of assemblies of channel proteins called connexins in vertebrates and innexins in invertebrates. The channels provide direct intercellular communication pathways allowing rapid exchange of ions and metabolites up to approximately 1 kD in size. Approximately 20 connexins are identified in the human or mouse genome, and orthologues are increasingly characterized in other vertebrates. Most cell types express multiple connexin isoforms, making likely the construction of a spectrum of heteromeric hemichannels and heterotypic gap junctions that could provide a structural basis for the charge and size selectivity of these intercellular channels. The precise nature of the potential signalling information traversing junctions in physiologically defined situations remains elusive, but extensive progress has been made in elucidating how connexins are assembled into gap junctions. Also, participation of gap junction hemichannels in the propagation of calcium waves via an extracellular purinergic pathway is emerging. Connexin mutations have been identified in a number of genetically inherited channel communication-opathies. These are detected in connexin 32 in Charcot Marie Tooth X linked disease, in connexins 26 and 30 in deafness and skin diseases, and in connexins 46 and 50 in hereditary cataracts. Biochemical approaches indicate that many of the mutated connexins are mistargeted to gap junctions and/or fail to oligomerize correctly into hemichannels. Genetic ablation approaches are helping to map out a connexin code and point to specific connexins being required for cell growth and differentiation as well as underwriting basic intercellular communication. PMID- 12126229 TI - Regulation of epithelial cell shape and polarity by cell-cell adhesion (Review). AB - Among all cell types that exhibit a polarized phenotype, epithelial cells are unique in that their polarity depends on the integration of the cell into a tissue, the epithelium. In recent years, the analysis of epithelial cell polarity in different epithelia and organisms has contributed to an understanding of the components involved and has further demonstrated that cell polarity and cell adhesion are intimately related to each other. Therefore, processes that mediate and modulate cell adhesion and coordinate adhesion and cell shape are fundamental for the function of epithelia. Recent results obtained in Drosophila melanogaster and Caenorhabditis elegans have provided further insight into the complex circuits regulating these processes, and have laid the direction for future analysis. PMID- 12126231 TI - Voltage-gated H+ channels associated with human phagocyte superoxide-generating NADPH oxidases: sequence comparisons, structural predictions, and phylogenetic analyses. AB - The N-terminal domain of the human phagocyte flavocytochrome b558 NADPH oxidase, gp91phox, is believed to be a heme-containing voltage-gated H+ channel. The authors have conducted structural, sequence and phylogenetic analyses of the putative transmembrane channel/heme-binding domains of all homologous proteins in the NCBI GenBank database as of May 2001, as well as of the full-length proteins. Fifty-six homologues were identified, including 26 from animals, 19 from plants, seven from yeast, one from a slime mould and three from bacteria. Six well defined sub-families were revealed by phylogenetic tree construction, two consisting of animal proteins, two of plant proteins, and one each of yeast and bacterial homologues, with the slime mould protein clustering loosely with one of the animal clusters. Signature sequences for the entire family as well as for the sub-families were determined. Most proteins have six putative TMSs, four of which may comprise the heme-binding H+ channel. The hydrophobic and amphipathic characteristics of each of the putative alpha-helical transmembrane segments were defined, and conserved residues that may be involved in heme binding, channel formation, and/or conformational changes were identified. The analyses lead to the suggestion that the oxidase domain became associated with the channel/heme binding domain to form a single polypeptide chain early in evolutionary history, before eukaryotes diverged from prokaryotes, and that genetic transmission to present day organisms occurred primarily by vertical descent. PMID- 12126232 TI - The role of human red blood cell membrane skeletal proteins in repetitive membrane strain and rupture. AB - The influence of the membrane skeleton on cell membrane deformability, elasticity, and rupture after repetitive cycles of membrane strain, release and rupture was investigated. Human red blood cells were electrofused to doublets, which showed the post-fusion oscillation-movement. Geometrical developments of heat-treated cells were measured and compared to control cells. Alterations of cluster length and fusion zone diameter during repetitive colloidosmotic swelling period grow with heat treatment, and the number of precedent swell phases has minor influence on these values. Irrespective of the treatment, the geometrical doublet configuration at which a membrane rupture is initiated has an almost constant roundness index of 0.89. Increasing heat treatment temperature was shown to affect both deformability and elasticity of the membrane, such that doublets start each swell phase of the oscillation cycle from decreased roundness values. Evidence is given that there is a difference in the mechanical properties between the membrane at the fusion zone and the membrane of native red blood cells. PMID- 12126233 TI - Plasma membrane components of adherens junctions (Review). AB - This review focuses on the three known plasma membrane components of adherens junctions: E-cadherin, nectin-2 and vezatin. The structures of these three components are discussed, with particular emphasis on the molecular mechanisms by which E-cadherin and nectin-2 promote cell adhesion. PMID- 12126234 TI - Desmosomal adhesion: structural basis, molecular mechanism and regulation (Review). AB - Desmosomes are adhesive intercellular junctions of epithelia and cardiac muscle. They have an essential function in maintaining the integrity of tissues, which is compromised in human genetic and autoimmune disease that targets desmosomal components. Recent evidence (1) suggests new roles for the function of desmosomal adhesion in tissue morphogenesis, (2) gives new insights into the molecular mechanism of adhesion, (3) indicates that the desmosomal adhesion molecules, desmocollin and desmoglein, may contribute to the regulation of epidermal diffentiation, and (4) shows that the affinity of desmosomal adhesion is regulated by protein kinase C. PMID- 12126235 TI - Cellular junctions of myelinated nerves (Review). AB - Myelinated nerves are specifically designed to allow the efficient and rapid propagation of action potentials. Myelinating glial cells contain several types of cellular junctions that are found between the myelin lamellas themselves in specialized regions of non-compact myelin and between the myelin membrane and the underlying axon. These include most of the junctional specializations found in epithelial cells, including tight, gap and adherens junctions. However, whereas in epithelial cells these junctions are formed between different cells, in myelinating glia these so called autotypic junctions are found between membrane lamellae of the same cell. In addition, myelinating glial cells form a heterotypic septate-like junction with the axon around the nodes of Ranvier and, in the peripheral nerve system, contact the basal lamina, which surrounds myelinating Schwann cells. This short review discusses the structure, molecular composition and function of the junctions present in myelinating cells, concentrating on the axo-glial junction. PMID- 12126236 TI - Gut inflammation modulated by the enteric nervous system and neurotrophic factors. PMID- 12126237 TI - Body mass and reflux oesophagitis: an oestrogen-dependent association? AB - BACKGROUND: There is widespread belief that obesity is associated with gastro oesophageal reflux disease, but the scientific evidence is weak and contradictory. Our aim is to evaluate the relation between body mass and reflux oesophagitis. METHODS: A population-based case-control study of endoscopically verified case subjects with reflux oesophagitis, and of randomly selected, control subjects matched for age, sex and area of residence. Subjects were classified within three body mass index (BMI) categories: BMI <25 (normal in the WHO classification), BMI 25-30 (overweight) and BMI >30 (obese). Odds ratios (OR) with 95% confidence intervals (CI) were the measures of association. RESULTS: Of 179 matched case-control pairs included in the study, 71 pairs were female. In males, no association between overweight and/or obesity and the risk of reflux oesophagitis was found. In females, there was a strong association between increasing BMI and the risk of reflux oesophagitis, with an OR of 2.9 (95% CI: 1.1-7.6) in the BMI 25-30 group and 14.6 (95% CI: 2.6-80.9) in the BMI >30 group (P value for trend = 0.0007). The association between obesity and oesophagitis was further strengthened by the use of oestrogen replacement medication. CONCLUSIONS: The study discloses a strong and dose-dependent association between body mass and reflux oesophagitis in women as opposed to no association among men. This association might be caused by increased oestrogen activity in overweight and obese females. PMID- 12126238 TI - Sources of intra-oesophageal nitric oxide production following intraluminal acid exposure. AB - BACKGROUND: The aim of the present study was to assess luminal nitric oxide (NO) levels in the oesophagus during baseline and acidic conditions and to clarify the sources of such oesophageal NO formation. METHODS: Healthy volunteers received an intra-oesophageal infusion of either HCl (100 mM) or NaCl (50 mM) on two separate study days. After a low nitrate diet, nitrate load or no dietary restrictions/pretreatment, direct intraluminal measurements of NO formation were performed using a tonometric technique. Endoscopy was performed and mucosal biopsies were taken and analysed by means of immunohistochemistry, Western blot and RT-PCR. RESULTS: No intra-oesophageal NO was detected during baseline conditions with pH neutrality. During the infusion of HCI the NO levels rose dramatically to around 12000 ppb. This high rate of NO formation fell by 95% following deviation of saliva. NO formation after an acute nitrate load was almost doubled during acid perfusion compared to control. Immunohistochemistry demonstrated distinct staining for iNOS in the oesophageal squamous epithelial cells, and Western blot and RT-PCR confirmed the presence of iNOS. CONCLUSION: Two sources exist for intra-oesophageal NO formation, both dependent on the luminal acidity: 1) chemical reduction of salivary nitrite, a mechanism related to dietary intake of nitrate, and 2) NO formation within the oesophageal mucosal epithelium by enzymatic degradation of L-arginine. In the latter case, the NO synthase has antigenic characteristics, indicating the inducible isoform, although a functional behaviour suggests an unconventional subtype. PMID- 12126239 TI - The natural course of gastroesophageal reflux disease in children. AB - BACKGROUND: The consequences of chronic gastroesophageal reflux disease (GERD) starting in childhood have not been widely studied. Our aim was to evaluate the usefulness of endoscopy in the primary diagnosis of GERD and to investigate the long-term course of this disease in children. METHODS: Between 1989 and 1999, 136 children had been endoscoped because of persisting symptoms of GER. After exclusions (neurological impairment, infant GER), 96 subjects were included, and files from 76 were available for the final evaluation. Twenty-four hour pH monitoring had been performed primarily on 67 children and at follow-up on 28, and endoscopy to 69 subjects and at follow-up to 33, respectively. Medical therapy as well as symptoms prior to the therapy were registered. Clinical outcome was assessed at the end of the follow-up period. RESULTS: Presenting symptoms were recurrent abdominal pain, heartburn, regurgitation and vomiting. Twenty-two patients had respiratory symptoms in addition to the gastrointestinal complaints. PH-recording was normal in 17/67 subjects, slightly pathological in 33 and severe reflux was diagnosed in 13 patients. Histologically, minimal changes associated with GER were diagnosed in 22 and mild esophagitis in 7. Thirty-six patients had been treated with prokinetic drugs. H2-blockers had been used in 24 children and proton-pump inhibitors in 4. After a mean follow-up period of 28 months, only 24% of patients had become symptom-free. Control endoscopy showed no progression of the esophageal inflammation in any of the subjects. CONCLUSIONS: Pathological reflux in children is associated with no or mild esophageal inflammation, which is unlikely to deteriorate. Therefore endoscopic control could be limited to cases with severe esophagitis. PMID- 12126240 TI - On-demand treatment in patients with oesophagitis and reflux symptoms: comparison of lansoprazole and omeprazole. AB - BACKGROUND: There are few data on how patients on maintenance treatment of reflux oesophagitis take their medication. This study was designed to investigate the dosing patterns of patients on on-demand treatment and to compare lansoprazole with omeprazole in this regard. METHODS: Patients with reflux oesophagitis, initially treated until absence of symptoms, took capsules of either lansoprazole (30 mg) or omeprazole (20 mg) for 6 months; they were instructed to take the medication only when reflux symptoms occurred. In order to document dosing patterns, the medication was dispensed in bottles supplied with a Medication Event Monitoring System recording date and time the bottles were opened. There were regular follow-up visits with assessment of symptoms. RESULTS: Three-hundred patients were eligible for analysis according to 'all patients treated'. A dosing pattern was found of an increased intake mornings and evenings and constant intervals between intakes. Although there was no correlation between oesophagitis grade or initial symptoms and the amount of medication consumed, the patients had significantly fewer reflux symptoms the more medication they consumed. There was no difference in the number of capsules consumed between the lansoprazole (0.73 capsules/day) and omeprazole groups (0.71 capsules/day). Nor was there any difference between the groups in reflux symptoms during the course of the study. CONCLUSION: Despite rigorous instructions to take medication on demand, the results suggest that it is patient habits more so than symptoms that determine the frequency and interval of medication intake. Symptoms are not therefore decisive for the amount of medication consumed. PMID- 12126241 TI - Ethanol-derived microbial production of carcinogenic acetaldehyde in achlorhydric atrophic gastritis. AB - BACKGROUND: Acetaldehyde is a local carcinogen in the digestive tract in humans. Atrophic gastritis leads to microbial colonization of the stomach, which could enhance microbial production of acetaldehyde from ethanol. The aim of the study was to study microbial ethanol metabolism and acetaldehyde production in the stomach of achlorhydric atrophic gastritis patients. METHODS: For the in vivo study, glucose or ethanol was infused via a nasogastric tube to the stomach of seven achlorhydric atrophic gastritis patients and five healthy controls. Gastric juice samples for ethanol and acetaldehyde determinations and microbial analysis were obtained at 30 and 60 min after the infusions. For the in vitro study, gastric juice samples from 14 atrophic gastritis patients and 16 controls were obtained during gastroscopy, whereafter the samples were incubated for 2 h with 1% ethanol at 37 degrees C and acetaldehyde was determined. RESULTS: Minor endogenous ethanol and acetaldehyde concentrations were detected after glucose infusion in the gastric juice of four atrophic gastritis patients. After ethanol infusion, the mean intragastric acetaldehyde level of the atrophic gastritis patients was 4.5-fold at 30 min and 6.5-fold at 60 min compared to controls. In vitro, the difference between the study groups was even higher, 7.6-fold. A vast selection of oral bacterial species and some Enterobacteriaceae and yeasts were presented in the gastric juice of atrophic gastritis patients. CONCLUSIONS: Microbial ethanol metabolism leads to high intragastric acetaldehyde levels after ethanol drinking in achlorhydric atrophic gastritis patients. This could be one of the factors responsible for enhanced gastric cancer risk among atrophic gastritis patients. PMID- 12126242 TI - A multinational comparison of racecadotril and loperamide in the treatment of acute watery diarrhoea in adults. AB - BACKGROUND: Racecadotril (acetorphan) is an orally active, potent inhibitor of enkephalinase, which exerts an antihypersecretory effect without increasing intestinal transit time. The aim of this study was to compare the efficacy, safety and tolerability of racecadotril with those of loperamide by assessing their effects on the resolution of the signs and symptoms of diarrhoea in patients in developing countries who had acute watery diarrhoea of less than 5 days' duration. METHODS: 945 outpatients from 21 centres in 14 countries received racecadotril (100 mg) or loperamide (2 mg) three times daily in a single-blind study. Duration of diarrhoea was the primary measure of efficacy; secondary criteria were overall clinical response, occurrence and duration of abdominal pain and distension, and occurrence of other associated signs and symptoms. Occurrence of constipation and adverse events were the main safety assessments. RESULTS: Diarrhoea resolved rapidly with both racecadotril and loperamide (55.0 h in both groups), 92% of patients on racecadotril and 93% on loperamide being treatment successes. Racecadotril produced a significantly greater reduction in abdominal pain and distension than loperamide (P = 0.024 and 0.03, respectively). The duration of abdominal distension was significantly shorter with racecadotril (5.4 versus 24.4 h; P = 0.0001), and constipation was also significantly less frequent (16% versus 25%; P = 0.001). One-hundred-and-eighty patients (19%) experienced one or more adverse event during the study: 67 (14.2%) in the racecadotril group and 113 (23.9%) in the loperamide group (P = 0.001). CONCLUSIONS: Racecadotril resolved the symptoms of acute diarrhoea rapidly and effectively, and produced more rapid resolution of abdominal symptoms and less constipation than loperamide. PMID- 12126243 TI - Evaluation of gastric emptying measured with the 13C-octanoic acid breath test in patients with functional dyspepsia: comparison with ultrasonography. AB - BACKGROUND: As a non-invasive modality by which to evaluate the gastric emptying of a solid meal, the 13C-octanoic acid breath test has recently become more widely used. Previously, we reported that ultrasonography was another non invasive and reliable method for assessing gastric motility. The aim of this study was to compare the reliability of these two methods. METHODS: Seventeen patients with functional dyspepsia and 10 healthy volunteers were studied. The solid test meal consisted of a scrambled egg labeled with 13C-octanoic acid (100 mg) and served with a bowl of rice and boiled chicken (total 424 kcal). After ingestion of the test meal, all subjects were examined in the sitting position. Ultrasonography images were obtained every 15 min for 3 h. Breath sampling followed the same time schedule as for the ultrasonography, with an additional 3 h of sampling at 30-min intervals. We investigated the half emptying time (T1/2) and the lag phase with both methods. RESULTS: The T1/2 by the ultrasonography method and the breath test were positively correlated (r2 = 0.638); however, there was no significant agreement between the study groups. Both the T1/2 and the lag phase were prolonged in the functional dyspepsia patients compared with the healthy volunteers, regardless of the method of measurement. The lag phase was significantly correlated (r2=0.864) with the T1/2 by the breath test. CONCLUSIONS: Although the 13C-octanoic acid breath test cannot assess the gastric emptying of solids as reliably as ultrasonography, both tests are useful for evaluating functional dyspepsia patients with delayed gastric emptying. PMID- 12126244 TI - Mechanism of the prorelaxing effect of thyroxine on the sphincter of Oddi. AB - BACKGROUND: Disturbances in the function of sphincter of Oddi (SO) may prevent normal bile flow and thus enhance the probability of common bile duct stone (CBDS) formation. Previously, we have shown increased prevalence of hypothyroidism in CBDS patients. METHODS: In animal (pig) experiments, thyroxine (T4) and triiodothyronine have a specific inhibitory effect on SO contractility, which raises the possibility that the lack of this prorelaxing effect in hypothyroidism could, at least in part, explain the increased prevalence of CBDS. The aims of the present study were to investigate, whether human SO reacts similarly to T4, and to study the mechanisms of the T4 prorelaxing effect. RESULTS: We found that T4 had similar inhibitory effects on both human and pig SO contractions. The T4 effect was dose-dependent, and maximum was observed in 30 min. The maximal prorelaxing effect was achieved with 0.1 nM T4 concentration, the effect of the physiological T4 concentration (0.01 nM) being about half of the maximal effect. Addition of alpha-adrenoceptor antagonist phentolamine, beta adrenoceptor antagonist propranolol, nitric oxide (NO)-synthesis inhibitor L NAME, nerve conductance blocker tetrodotoxin, or cyclooxygenase inhibitor diclofenac did not affect the T4-induced inhibition of contraction. Addition of transcription inhibitor actinomycin D or translation inhibitor cyclophosphamide partially reversed the T4-induced inhibition of contraction. Addition of K+ channel blocker glibenclamide totally reversed the T4-induced inhibition of contraction. In Western blotting, the thyroid hormone receptor (TR) antibody recognized 53 kDa and 58 kDa proteins, corresponding to beta1 and beta2 isoforms of TR, in the human SO tissue. CONCLUSIONS: We conclude that T4 has a direct prorelaxing effect on human SO that expresses TR beta1 and beta2. This effect is mediated through a transcriptional mechanism that requires new mRNA and protein synthesis and subsequently leads to the activation of K+ channels. PMID- 12126245 TI - Evaluation of esophageal motility by endosonography using a miniature ultrasonographic probe in patients with reflux esophagitis. AB - BACKGROUND: Although there are several established methods used to evaluate esophageal motility, none allows for direct observation of esophageal wall motion. Esophageal dysmotility is thought to contribute to reflux esophagitis (RE). The aim of this study was to evaluate esophageal wall motility by endosonography using a miniature ultrasonographic probe (MUP) in patients with RE. METHODS: The subjects consisted of 10 healthy controls (10 men with a mean age of 31.5 years) and 9 patients with RE (4 men and 5 women with a mean age of 51.5 years). High-frequency endoluminal sonography was performed using a 20-MHz transducer through a 16F gastric tube to evaluate esophageal wall motion. Four sonographic phases of an esophageal peristaltic sequence were identified. In the resting phase, the esophageal wall was in direct contact with the transducer. In the passive distention phase, the esophageal lumen was stretched maximally; in the contraction phase, it contracted; and in the relaxation phase, it returned to baseline. The baseline thickness of the muscle layers of the esophageal wall was measured at rest. The width decreased during the passive distention phase, increased and reached a maximum during the contraction phase, and returned to baseline during the relaxation phase. RESULTS: The contractility index of the circular smooth muscle (CSM) in the distal esophagus and of the longitudinal smooth muscle (LSM) in the proximal esophagus were significantly lower in patients with RE. The duration of contraction in the distal esophagus was significantly longer in RE. CONCLUSIONS: We used a MUP to demonstrate abnormalities in esophageal wall motility in patients with RE. We conclude that the MUP is a potentially useful technique for evaluating esophageal dysmotility. PMID- 12126246 TI - A reliable screening procedure for coeliac disease in clinical practice. AB - BACKGROUND: The main autoantigen recognized by the sera of patients with coeliac disease (CD) is tissue transglutaminase (tTG). A human-recombinant form of tTG was used to develop an ELISA to measure anti-tTG serum antibodies for the diagnosis of CD. Preliminary retrospective reports suggest that the human tTG based ELISA could identify coeliac patients missed by the IgA-anti-endomysium antibody test (AEA). Whether the human recombinant tTG ELISA is sufficiently accurate to become the main diagnostic CD tool in everyday clinical practice is unknown. The objective was to determine, in a prospective study, the sensitivity and specificity of an ELISA test based on the use of human tTG compared with AEA, to analyse the discordant cases for HLA DQ2-8 and for clinical and intestinal biopsy characteristics. METHODS: 1106 patients referred to a gastrointestinal outpatient clinic for symptoms attributable to CD, 52 first-degree relatives of CD patients and 200 healthy controls were tested for both anti-human tTG and AEA antibodies. RESULTS: Out of 1158 subjects, 146 were tested positive for anti-tTG antibodies and 140 were biopsy-proven coeliacs. The AEA test identified 126/1158 coeliacs who also tested positive for anti-tTG antibodies. The 14 patients missed by the AEA test carried the typical HLA-DQ for CD; they had normal levels of total serum IgA and had milder pathology than those with both anti-tTG and AEA positivity (P < 0001). CONCLUSIONS: These results prove that human tTG-based ELISA is an excellent diagnostic tool for CD, for mass screening by both the specialist and the general clinic. PMID- 12126247 TI - Antibodies against human tissue transglutaminase and endomysium in diagnosing and monitoring coeliac disease. AB - BACKGROUND: Coeliac disease (CD) patients often present a variety of uncharacteristic symptoms and therefore sensitive and specific screening tests are needed as an aid in making an accurate diagnosis. A recently developed ELISA, using human recombinant tissue transglutaminase (tTG) as antigen, was evaluated for its significance in the diagnosis of CD. The patient's compliance to a gluten free diet and the serological reaction during gluten challenge were also monitored. The results were compared with IgA-endomysium antibody (EMA) results. METHODS: Sera previously collected from 365 patients (0.4-76 years) with jejunal biopsy on a gluten-containing diet and from 41 patients on a gluten-free diet or challenge were tested for IgA anti-human tTG antibodies (IgA tTG ab) with Celikey (Pharmacia Diagnostics). The study population comprised 208 CD patients and 157 controls. The diagnostic performance and cut-off for the assay were estimated with ROC analysis. EMA was analysed by indirect immunofluorescence microscopy on cryostat sections of monkey oesophagus. RESULTS: 200/208 patients with CD had positive IgA tTG ab (median >100 U/ml), while only 1/157 of the control patients were positive (median 1.67 U/ml). The area under the ROC curve was 98.3% and the sensitivity and specificity of the test were 96% and 99% for the study population. Only 4/365 patients (1%) presented discordant IgA tTG ab and EMA results, 2 of them had only IgA tTG ab and 2 only EMA. The IgA tTG ab levels and the EMA titres were closely correlated to the duration of gluten-free diet and gluten challenge, respectively. CONCLUSION: IgA tTG ab can be used as an accurate observer-independent alternative to EMA in diagnosing or monitoring CD. PMID- 12126248 TI - Familial and sporadic inflammatory bowel disease: comparison of clinical features and serological markers in a genetically homogeneous population. AB - BACKGROUND: The familial occurrence of inflammatory bowel disease (IBD) and the clinical features of familial and sporadic IBD in the genetically homogeneous Finnish population are evaluated. METHODS: 257 patients with Crohn disease (CD) and 436 with ulcerative colitis (UC) participated in the study. They were asked whether IBD was present (familial IBD) or absent (sporadic IBD) in their first degree relatives. Data on the clinical course of the disease were collected from the patient records. Antibodies to Saccharomyces cerevisiae (ASCA) and anti neutrophil cytoplasmic antibodies (ANCA) were determined from serum samples. RESULTS: Affected first-degree relatives were found in 15.6% of patients with CD and in 13.8% of patients with UC. In familial cases CD was more often located in the ileum (38% versus 21%) and less often in the ileocolon (35% versus 50%) (P< 0.05) than in sporadic cases. A greater percentage of CD patients than UC patients were smokers (47% versus 13%; P < 0.01). An elevated level of IgA and/or IgG antibodies for ASCA was found more often in CD patients than in UC patients (59% versus 14%; P < 0.01), while pANCA were found more often in UC than in CD patients (48% versus 12%; P < 0.01). The combination of pANCA-ASCA+ yielded a sensitivity, specificity and positive predictive value of 48%, 92% and 90%, respectively, for CD, and the combination of pANCA + ASCA- of 55%, 94% and 90%, respectively, for UC. CONCLUSIONS: The percentage of familial IBD cases in Finland is comparable to that reported elsewhere in Europe. No important clinical differences between patients with familial and sporadic forms of the disease were found. ASCA is associated with both familial and sporadic CD and pANCA with UC, but low sensitivity diminishes their value as a serological marker of IBD or as a differential diagnostic test between CD and UC. PMID- 12126249 TI - Ulcerative colitis is associated with a promoter polymorphism of lipopolysaccharide receptor gene, CD14. AB - BACKGROUND: Inflammatory bowel disease (IBD) is a multifactorial disease with a significant genetic background. Evidence is accumulating that molecules such as CD14, which interact with luminal bacterial constituents, are involved in the pathogenesis. It has recently been shown that the T allele of the 5'-flanking region of the CD14 gene at position -159 is related to high expression of CD14. In further exploring the genetic background of IBD, we investigated this novel polymorphism of CD14 gene in patients with ulcerative colitis or Crohn disease. METHODS: DNA was obtained from 101 patients with ulcerative colitis, 82 with Crohn disease and 123 healthy controls. All were typed for the promoter polymorphism of the CD14 gene at position -159 by restriction fragment length polymorphism analysis. Serum samples were obtained from 105 healthy controls and serum sCD14 levels were measured. RESULTS: T allele frequencies were 57.4%, 48.2% and 44.7% in ulcerative colitis, Crohn disease and healthy controls, respectively. The T allele and T/T genotype frequencies were significantly higher in ulcerative colitis patients than in healthy controls (P = 0.0074, OR = 1.67, 95% CI = 1.15-2.42, P = 0.022, OR= 1.96 95% CI: 1.10-3.48, respectively). The sCD14 level was significantly higher in TT genotype populations than CC (P = 0.0205). CONCLUSIONS: The promoter polymorphism of the CD14 gene at -159T plays a significant role in regulating the CD14 expression and is positively associated with ulcerative colitis, and this polymorphism may confer a genetic predisposition to ulcerative colitis. The results also support the concept that bacterial constituents may be involved in the pathogenesis of ulcerative colitis. PMID- 12126250 TI - Effect of budesonide enema on remission and relapse rate in distal ulcerative colitis and proctitis. AB - BACKGROUND: Glucocorticosteroid enemas are equally effective as 5-ASA enemas in the treatment of active distal ulcerative colitis (UC). With the introduction of budesonide, the risk of systemic side effects may be reduced. We investigated whether budesonide enema, 2 mg/100 ml, administered twice daily (b.i.d.) could increase the remission rate in comparison with the once daily (o.d.) standard regimen. Furthermore, we evaluated whether 2 mg budesonide enema, given twice weekly, could have a relapse preventing effect. METHODS: 149 patients with active distal UC were treated in a controlled, double-blind multicentre study with two parallel groups: placebo enema in the morning and budesonide enema in the evening (i.e. 2 mg/day) or budesonide enema b.i.d. (i.e. 4 mg/day) until remission (absence of clinical symptoms and endoscopic healing) or at most 8 weeks. Patients in remission were randomized to either budesonide enema or placebo enema twice weekly for 24 weeks or until relapse. RESULTS: The remission rates at 4 weeks were 33% for o.d. and 41% for b.i.d. regimens (NS) and correspondingly 51% and 54% at 8 weeks (NS). The b.i.d. group had an increased frequency of impaired adrenal function, 32% versus 4.8% (P = 0.001). The relapse rates during maintenance treatment with budesonide enema and placebo were 15% versus 24% after 8 weeks, 31% versus 27% after 16 weeks and 41% versus 51% after 24 weeks (NS). CONCLUSION: Budesonide enema 2 mg o.d. appears to be the optimal dosage in active distal UC. We could not show that budesonide enema twice weekly is sufficient to maintain remission. PMID- 12126251 TI - Yersinia species in collagenous colitis: a serologic study. AB - BACKGROUND: The etiology of collagenous colitis is unknown. An infectious cause seems a possibility, and in a recent report three out of six patients with collagenous colitis were shown to have had an infection with Yersinia enterocolitica. The aim was to investigate the occurrence of Yersinia antibodies in collagenous colitis. METHODS: Sera from 32 collagenous colitis patients and 17 healthy controls were analysed for antibodies against Yersinia virulence proteins. RESULTS: Collagenous colitis patients had Yersinia antibodies more often than the controls, 9 having a positive and 4 an intermediate antibody score of the 32 patients. In comparison, I out of 17 controls had a positive and 2 an intermediate antibody score, which represents a strong, although not significant, trend (P = 0.078). CONCLUSION: The data showed that Yersinia antibodies are more common in collagenous colitis patients than in healthy controls. In some cases, Yersinia might have been the triggering event in the development of collagenous colitis. PMID- 12126252 TI - K-ras mutations in sera of patients with colorectal neoplasias and long-standing inflammatory bowel disease. AB - BACKGROUND: About 50% of colorectal carcinomas and adenomas display K-ras mutations, which have also been described in stool or colonic lavage fluid. Moreover, the presence of K-ras mutations in plasma samples originating from patients with colorectal cancer has been reported recently. METHODS: DNA was extracted from sera of 16 patients with colorectal carcinomas, 6 with large adenomas, 3 with Crohn disease and 4 with ulcerative colitis. Sera of 20 healthy blood donors served as negative controls. K-ras mutations at the first or second position of codon 12 were detected by an enriched RFLP-PCR method and confirmed by sequencing. RESULTS: Mutations were found in sera of 5 patients with colorectal carcinomas (31%) and 2 patients with long-standing ulcerative pancolitis (50%), but not in patients with adenomas, Crohn disease or the controls. CONCLUSIONS: K-ras mutations can be detected in serum samples from patients with manifest colorectal cancer and in patients who display an increased risk for malignant transformation of the colonic mucosa. This observation may have clinical application concerning noninvasive surveillance of these patients. Because of the low sensitivity of this approach it may be useful to combine it with other molecular markers. PMID- 12126254 TI - Iron overload decreases the protective effect of tumour necrosis factor-alpha on rat hepatocytes exposed to oxidative stress. AB - BACKGROUND: Intracellular iron can participate in the formation of free radicals, leading to liver cell damage. This may be prevented by the ability of ferritin to oxidize and store iron. Tumour necrosis factor alpha (TNF-alpha) has been shown to increase the ferritin synthesis. In the liver, cytokines are secreted by activated Kupffer cells and T-lymphocytes. The aim of this study was to investigate the effects of TNF-alpha on normal and iron-loaded rat hepatocytes exposed to oxidative stress. METHODS: Primary cultures of hepatocytes from rats fed a normal rat chow or a carbonyl iron-enriched diet were incubated with TNF alpha before incubation with tert-butyl hydroperoxide. Malondialdehyde concentrations, activities of lactate dehydrogenase, ferritin H and manganese superoxide dismutase mRNA and ferritin H protein were analysed. The total amounts of glutathione and chelatable iron were measured. RESULTS: TNF-alpha diminished the concentrations of malondialdehyde and activities of lactate dehydrogenase in hepatocytes exposed to tert-butyl hydroperoxide. This was seen in hepatocytes from normal but not iron-loaded animals. The transcription of manganese superoxide dismutase mRNA was increased in both cell types, whereas total glutathione contents of cells were unaffected. The transcription and translation of ferritin H was induced in cells from normal but not from iron-loaded animals. The amount of chelatable iron was significantly lowered only in hepatocytes from normal rats. CONCLUSIONS: TNF-alpha protects rat hepatocytes from normal but not iron-loaded rats from oxidative stress. The protection may be due to an induction of the ferritin synthesis. PMID- 12126253 TI - Persons with screening-detected haemochromatosis: as healthy as the general population? AB - BACKGROUND: Hereditary haemochromatosis (HH) is a common genetic disease leading to iron deposition in the liver and other organs. Early treatment will prevent clinical disease and population-based screening for HH has been advocated. However, the benefit of screening depends on the morbidity of HH. We have compared the morbidity in HH persons detected by screening with the morbidity in the rest of the population. METHODS: All inhabitants 20 years or older in a Norwegian county (94,191 persons) were invited to participate in a health survey programme. Of 65,717 participating persons, a blood specimen for transferrin saturation was obtained from 65,238. After repeated laboratory testing and clinical examination, 269 persons were found to have phenotypic HH, while 297 had genotypic HH (the C282/C282Y mutation). Using self-reported data, clinical examinations and analysis of non-fasting blood samples, the morbidity in phenotypic and genotypic HH persons was compared with the morbidity in the rest of the population. All data were collected before subjects were diagnosed with HH, and all comparisons were corrected for age and gender. RESULTS: Compared to control persons, phenotypic and genotypic HH men and women had a higher score on 1 of 17 questions dealing with joint complaints. Phenotypic and genotypic HH women below 50 years of age had a higher prevalence of hypothyroidism (15.2% and 12.5%, respectively, compared to 3.0% in the control population). Phenotypic HH women below 50 years of age had higher diastolic blood pressure than control women. Phenotypic HH men above 50 years of age and genotypic HH men scored lower than control men on a compound myocardial infarction risk score variable, in part due to lower serum cholesterol concentration. Fewer phenotypic HH men above 50 years of age reported having angina pectoris. Otherwise, the health of phenotypic and genotypic HH persons was not different from the health of control persons. CONCLUSION: When corrected for age and gender, the morbidity in persons with screening-detected HH was not very different from the morbidity in the control group, indicating that population-based screening may not be as beneficial as anticipated. PMID- 12126255 TI - Endoscopic ultrasonography for preoperative diagnosis and localization of insulinomas. AB - BACKGROUND: Endoscopic ultrasonography (EUS) is generally accepted as a sensitive method for the detection of small pancreatic tumors. We report our experience with EUS for preoperative imaging of insulinomas. METHODS: Nine patients with clinical and biochemical signs of insulinoma were examined by EUS using a 7.5/12 MHz radial-scanning ultrasound endoscope prior to surgery. EUS outcome was evaluated on the basis of surgery (open or laparoscopic) and examination of the resected specimens. RESULTS: Two EUS-negative patients appeared, by reassessment of clinical and biochemical data, not to have an insulinoma and were not operated on. EUS correctly imaged and localized five of seven insulinomas that were surgically removed. One isoechoic tumor in the pancreatic head and one pedunculated tumor connected to the caudal side of the pancreatic body were missed by EUS. EUS could demonstrate the size and shape of the imaged tumors, as well as their relationship to adjacent structures, such as the pancreatic duct, bile duct, and large vessels. CONCLUSIONS: Our experience with seven insulinomas accords with previous reports claiming EUS to be the method of choice for preoperative imaging and localization of pancreatic islet cell tumors. PMID- 12126256 TI - PH-activated phospholipase A2: an important mucosal barrier breaker in peptic ulcer disease. PMID- 12126257 TI - Rectal muciphages are rich in lysozymes: a novel source of antimicrobial mucosal defense? PMID- 12126258 TI - Researchers receive record compensation from TV company for media insult linked to the calcium antagonist controversy. PMID- 12126259 TI - Systolic hypertension as a cardiovascular risk factor. PMID- 12126260 TI - Diagnostic criteria of white coat hypertension (WCH): consequences for the implications of WCH for target organs. AB - In a sample comprising 51 normotensive subjects and 51 subjects with in-clinic arterial hypertension [blood pressures (BPs) > or = 140/90 mmHg), we investigated the prevalence of target organ damage [left ventricular hypertrophy (LVH) and retinal vasculopathy] in white coat hypertension (WCH) groups defined using: (a) the "optimal ambulatory BP" criterion of the Seventh International Consensus Conference (in-clinic BPs >140/90 mmHg, daytime mean BPs < 130/80 mmHg) and (b) the "normal ambulatory BP" criterion proposed in 1997 by Verdecchia and co workers (in-clinic BPs >140/ 90 mmHg, daytime mean BPs < 135/85 mmHg), and we compared the results with those obtained for the normotensive group and for a WCH group defined as in a 1996 study of the same data. We found that the newer criteria did not alter the conclusions reached in 1996: namely, that WCH constitutes a state of risk intermediate between normotension and sustained hypertension, which demands in-depth evaluation and active monitoring, if not immediate therapy. We also found that when the WCH group was defined as those patients with in-clinic BPs > or = 140/90 mmHg and 24-h mean BPs < 121/78 mmHg, the prevalence of target organ damage was similar to that found in the control group. We conclude that if WCH status is to imply absence of elevated risk of target organ damage, then the ambulatory BP threshold defining WCH should be lower than the upper limit of ambulatory BPs among subjects who are normotensive in the clinic. The desirability of predicting target organ damage in both hypertensive and normotensive subjects using criteria combining in-clinic BPs, daytime mean ambulatory BPs and night-time mean ambulatory BPs is suggested. PMID- 12126261 TI - The pandora project: cost of hypertension from a general practitioner database. AB - AIM: The Pandora Project is a longitudinal database--implemented by general practitioners since June 1997 in the Ravenna area (Italy)--providing information on patients with hypertension. Data from 1,651 patients were followed up for I year in order to investigate the cost of hypertension. Only direct medical costs were considered in the perspective of the National Healthcare System. FINDINGS: At enrollment, 552 patients were classified as normotensive, 1,099 as hypertensive. After 1 year, among normotensive group, 352 patients remained normotensive and 200 became hypertensive; among hypertensive group, 323 patients became normotensive and 776 remained hypertensive. The average total cost per patient at follow-up was 779.59 Euros. About 46% of total cost was due to anti hypertensive therapy, irrespective of the evolution of blood pressure levels registered, whilst other direct costs represented 54% of total patient cost in all cohorts. It is possible that co-morbidities play a significant role in this situation. Patient aged 80-89 years generate higher costs. Even if further investigation is needed on the burden of comorbidity on a per-patient cost of hypertension, this work provides evidence that the average total cost per patient is likely to increase with age and co-morbidities. Key words: cost-of-illness, costs, economics, hypertension. PMID- 12126263 TI - Placebo-controlled comparison of the efficacy and tolerability of once-daily moxonidine and enalapril in mild to moderate essential hypertension. AB - OBJECTIVES: To compare the antihypertensive efficacy and tolerability of the imidazoline I1-receptor agonist moxonidine, administered as a single daily dose of 0.6 mg, with the angiotensin-converting enzyme inhibitor enalapril (20 mg o.d.) in patients with mild to moderate essential hypertension. METHODS: A total of 154 patients were enrolled and randomized to placebo (n = 50), moxonidine (0.2 mg o.d.; n = 51) or enalapril (5 mg o.d.; n = 53) for 2 weeks. Dosages were then increased to moxonidine 0.6 mg o.d. or enalapril 20 mg o.d. for a further 6 weeks. Blood pressure responses to therapy were measured using conventional office techniques and by 24-h ambulatory blood pressure monitoring. RESULTS: The average reduction in sitting blood pressure with moxonidine was similar to that achieved with enalapril (24.9 +/- 20.7/13.2 +/- 8.4 mmHg vs 21.9 +/- 17.1/11.9 +/ 7.5 mmHg, respectively) and significantly superior to that seen with placebo (1.2 +/- 14.4/2.3 +/- 7.0 mmHg; p < 0.001). Reductions in blood pressure after 8 weeks of treatment were as follows: moxonidine, from 166.2 +/- 15.5/101.3 +/- 4.0 to 141.2 +/- 15.7/88.1 +/- 7.7mmHg; enalapril, from 165.4 +/- 14.3/101.1 +/- 4.4 to 143.5 +/- 12.7/89.2 +/- 7.4 mmHg; and placebo, from 162.5 +/- 14.4/99.9 +/- 3.9 to 161.3 +/- 17.9/97.6 +/- 6.6 mmHg. Both moxonidine and enalapril produced sustained reductions in blood pressure during 24-h recording (p < 0.01 for overall effect of either drug vs placebo). Average 24-h blood pressure was reduced from 149.8 +/- 14.3/92.2 +/- 7.0 to 134.0 +/- 13.1/82.3 +/- 8.9 mmHg with moxonidine and from 146.5 +/- 13.0/ 92.5 +/- 7.2 to 131.1 +/- 11.0/82.1 +/- 8.8 mmHg with enalapril; the change with placebo was small (from 147.3 +/- 13.3/90.0 +/- 6.2 to 144.8 +/- 12.9/89.5 +/- 8.0 mmHg). Both drugs were generally well tolerated. No patients withdrew from the study because of drug-attributed adverse events. CONCLUSION: Moxonidine 0.6 mg o.d. and enalapril 20 mg o.d. have similar antihypertensive efficacy. PMID- 12126262 TI - Cognitive function and health-related quality of life in elderly patients with hypertension--baseline data from the study on cognition and prognosis in the elderly (SCOPE). AB - The Study on COgnition and Prognosis in the Elderly (SCOPE) study is a multi centre, prospective, randomized, double-blind, and parallel-group study aiming at comparing the effects of candesartan cilexetil and placebo on cardiovascular events and cognitive function in elderly patients with hypertension. The aim of this sub-analysis was to present data on cognitive function and health-related quality of life (HRQL) at baseline (randomization), and to investigate whether cognitive function was related to HRQL. More specifically, the main aim was to investigate the possible relationship between the Mini-Mental State Examination on one hand, and the Psychological General Well-Being (PGWB) index, the Subjective Symptom Assessment (SSA-P) Profile and the EuroQoL Health Utility Index (EQ-5D) on the other. All the instruments are extensively validated. A general finding was that cognitive function was positively associated with higher well-being (PGWB total score and self-control) and higher utility value (EQ-SD current health) but was unrelated to the occurrence of subjective adverse symptoms (SSA-P). Age and the use of psychotropic drugs, but not gender and education level, influenced this relationship significantly. PMID- 12126264 TI - Long-term exposure to telmisartan as monotherapy or combination therapy: efficacy and safety. AB - This multicentre, open-label extension study to four controlled trials involved 888 patients with mild-to-moderate primary hypertension. Patients received telmisartan 40-80 mg once daily with add-on hydrochlorothiazide (HCTZ; 12.5-25 mg) if necessary and/or other antihypertensives to achieve diastolic blood pressure (DBP) control (<90 mmHg). Treatment continued for up to 4 years. At treatment end, 578 (65.1%) patients were on telmisartan monotherapy, 106 (11.9%) were on telmisartan + HCTZ 12.5 mg, 101 (11.4%) were on telmisartan + HCTZ 25 mg, and 103 (11.6%) were on telmisartan + another antihypertensive + HCTZ. Overall, 84.4% (746/884) patients achieved DBP control. The highest proportion of responders was in the telmisartan monotherapy (40 or 80 mg) treatment category (89.0% 1,511/574 patients]). The mean change in systolic blood pressure (SBP)/DBP from the previous trial baseline to last available trough was -21.2/-17.3 mmHg with telmisartan alone, -24.6/-16.7 mmHg with telmisartan + HCTZ, and - 18.7/ 14.9 mmHg with telmisartan + another antihypertensive +/- HCTZ. Most adverse events were of mild-to-moderate intensity and unrelated to treatment. The proportions of patients discontinuing the study due to adverse events, by treatment at onset, were 7.3% (65/888) with telmisartan monotherapy, 6.6% (20/304) with telmisartan + HCTZ and 2.9% (3/103) with telmisartan + another antihypertensive +/- HCTZ. There were 15 deaths during the study, but none was considered drug related. Thus, telmisartan alone or in combination with other antihypertensives achieved and maintained clinically relevant reductions in DBP and SBP. This long-term analysis supports the favourable efficacy and safety profile of telmisartan both as monotherapy and in combination with other antihypertensive drugs. PMID- 12126265 TI - Nebivolol vs amlodipine as first-line treatment of essential arterial hypertension in the elderly. AB - The antihypertensive efficacy of nebivolol and amlodipine and their tolerability were compared in a multicentre, randomized, active-controlled, double-blind parallel-group trial in elderly patients with mild to moderate essential arterial hypertension. One hundred and eighty-four subjects aged > or = 65 years were screened. After a run-in phase of 4 weeks, only 168 of these were randomized with either nebivolol 2.5-5 mg daily (n = 81) or amlodipine 5-10 mg daily (n = 87) over a period of 12 weeks. The response rate to treatment and the changes of sitting diastolic blood pressure (BP) at week 12 were similar between the two groups. A lower sitting systolic BP (SBP) was detected with amlodipine at week 4 (p < 0.05) and at week 8 (p < 0.05). Standing BP showed no changes between the two groups; only SBP was lower with amlodipine at week 8 (p < 0.05). Heart rate was lower at all treatment visits with nebivolol (p < 0.001). The incidence of adverse events was no different between the two groups; however the incidence of headache and ankle oedema was significantly higher with amlodipine (p < 0.05). In elderly subjects with essential hypertension, the antihypertensive efficacy of nebivolol and amlodipine was similar. Both drugs were well tolerated, although amlodipine was accompanied by higher incidence of drug-related adverse events. PMID- 12126267 TI - Molecularly imprinted materials--receptors more durable than nature can provide. AB - The chapter describes the concept of molecular imprinting. This technology allows the fabrication of artificial polymeric receptors applicable in many areas of biotechnology. Polymers imprinted with selected template molecules can be used as specific recognition elements in sensors or as selective stationary phases in affinity chromatography or in capillary electrochromatography. However, also in solid phase extraction or immunoassays these polymers (MIP) are able to compete with traditional materials such as biological antibodies. Furthermore, polymers molecularly imprinted with so-called transition state analogue templates can be applied as catalysts. In other words, these kind of polymers may be used as artificial antibodies (plastibodies) or biomimicking enzymes (plastizymes). Compared to their biological counterparts, MIP offer different advantages such as simplicity in manufacturing and durability. Thus, the author expects MIP to have a major impact on the whole area of biotechnology. PMID- 12126266 TI - Capillary electrochromatography: a rapidly emerging separation method. AB - This overview concerns the new chromatographic method--capillary electrochromatography (CEC)--that is recently receiving remarkable attention. The principles of this method based on a combination of electroosmotic flow and analyte-stationary phase interactions, CEC instrumentation, capillary column technology, separation conditions, and examples of a variety of applications are discussed in detail. PMID- 12126268 TI - Chromatographic reactors based on biological activity. AB - In the last decade there were many papers published on the study of enzyme catalyzed reactions performed in so-called chromatographic reactors. The attractive feature of such systems is that during the course of the reaction the compounds are already separated, which can drive the reaction beyond the thermodynamic equilibrium as well as remove putative inhibitors. In this chapter, an overview of such chromatographic bioreactor systems is given. Besides, some immobilization techniques to improve enzyme activity are discussed together with modern chromatographic supports with improved hydrodynamic characteristics to be used in this context. PMID- 12126270 TI - Continuous annular chromatography. AB - In recent years the demand for process scale chromatography systems in the industrial downstream process has been increasing steadily. Chromatography seems to be the method of choice when biological active compounds must be recovered from a mixture containing dozens of side products and contaminants as it is for example the case when processing fermentation broths. Since chromatography can solve almost any separation problem under mild operating conditions, a continuous chromatography system represents an extremely attractive and powerful option for such large-scale applications. The increasing number of biotechnological products forces system suppliers of the downstream processing side to develop new and improved high throughput purification technologies. Continuous Annular Chromatography (CAC) has been shown to be the only continuous chromatography technique to fulfill the high demands raised by modern biotechnological productions. In recent years Prior Separation Technology has transferred the principle of Continuous annular chromatography from the research laboratories to the fully developed industrial downstream process scale. The technology is now called Preparative Continuous Annular Chromatography--P-CAC. It can be placed at any stage in the downstream line starting at the very early stages where capturing and concentration of the desired product is required down to the polishing steps, which assure a sufficient final purity of the end product. PMID- 12126269 TI - Simulated moving bed chromatography (SMB) for application in bioseparation. AB - Simulated Moving Bed (SMB) technology is of rising interest in the field of bioseparation. This is particularly due to its advantages such as reduction of solvent consumption, high productivity and final purities as well as low investment costs in comparison to eluent chromatography. SMB units can operate under high productivity overloaded conditions. This leads to non-linear competitive adsorption behavior, which has to be accounted for when designing and optimizing new SMB separations. The so called "Triangle Theory", which is briefly reviewed in this chapter, provides explicit criteria for the choice of the operating conditions of SMB units to achieve the prescribed separation of a mixture characterized by Langmuir, modified Langmuir and bi-Langmuir isotherms. The application of the SMB-technique to the downstream processing of biotechnological products requires some specific changes to meet the special demands of bioproduct isolation. Some exemplary applications are given including separations of sugars, proteins, monoclonal antibodies, ionic molecules and optical isomers and for desalting. PMID- 12126271 TI - Short monolithic columns as stationary phases for biochromatography. AB - Monolithic supports represent a novel type of stationary phases for liquid and gas chromatography, for capillary electrochromatography, and as supports for bioconversion and solid phase synthesis. As opposed to individual particles packed into chromatographic columns, monolithic supports are cast as continuous homogeneous phases. They represent an approach that provides high rates of mass transfer at lower pressure drops as well as high efficiencies even at elevated flow rates. Therefore, much faster separations are possible and the productivity of chromatographic processes can be increased by at least one order of magnitude as compared to traditional chromatographic columns packed with porous particles. Besides the speed, the nature of the pores allows easy access even in the case of large molecules, which make monolithic supports a method of choice for the separation of nanoparticles like pDNA and viruses. Finally, for the optimal purification of larger biomolecules, the chromatographic column needs to be short. This enhances the speed of the separation process and reduces backpressure, unspecific binding, product degradation and minor changes in the structure of the biomolecule, without sacrificing resolution. Short Monolithic Columns (SMC) were engineered to combine both features and have the potential of becoming the method of choice for the purification of larger biomolecules and nanopartides on the semi-preparative scale. PMID- 12126272 TI - Porous polymer monoliths: an alternative to classical beads. AB - Porous polymer monoliths are a new category of materials developed during the last decade. These materials are prepared using a simple molding process carried out within the confines of a closed mold. Polymerization of a mixture that typically contains monomers, free-radical initiator, and porogenic solvent affords macroporous materials with large through-pores that enable flow-through applications. The versatility of the preparation technique is demonstrated by its use with hydrophobic, hydrophilic, ionizable, and zwitterionic monomers. The porous properties of the monolith can be controlled over a broad range. These, in turn, determine the hydrodynamic properties of the devices that contain the molded media. Since all the mobile phase must flow through the monolith, the mass transport within the molded material is dominated very much by convection, and the monolithic devices perform well even at very high flow rates. The applications of monolithic materials are demonstrated on the chromatographic separation of biological compounds and synthetic polymers, electrochromatography, gas chromatography, enzyme immobilization, molecular recognition, and in advanced detection systems. Grafting of the pore walls with selected polymers leads to materials with completely changed surface chemistries. PMID- 12126273 TI - A heroin epidemic at the intersection of histories: the 1960s epidemic among African Americans in Baltimore. AB - In the drug field the fundamental epidemiological question-why illicit drug use, here, now, among these people-has still not been adequately answered. Drawing on the work of colleagues in medical anthropology, we attempt to move closer to an answer by developing a "trend theory." In this article we analyze a single case: the increase in heroin use and addiction among African Americans in the City of Baltimore in the 1960s. We found that the two most important historical processes behind the epidemic were (1) a changing distribution/supply system and (2) the mix of hope and despair that was part of the early civil rights movement. PMID- 12126274 TI - Theorizing health in the context of transition: the dynamics of perceived morbidity among women in peri-urban Maharashtra, India. AB - In international health, transition models that project global trends in fertility, morbidity, and mortality have been challenged for their underlying assumptions about modernization and the inability to capture inequities in the distribution of health and health services, the resurgence of infectious diseases and an apparent rise in perceived morbidity. The evidence that individuals' perceived morbidity rarely corresponds to biomedical measures of health status prompts questions regarding the relationship of epidemiologic transitions to shifts in ways of recognizing, representing, and responding to health and illness. In both international and national health policy and research, the discursive parameters of "women's health" in India largely reflect the logic of transition models and rarely consider how women themselves see "health" and its determinants, the dynamics of women's perceived morbidity, and women's response to changes in local medical discourse and practice. Drawing on survey and ethnographic data collected during 1996-97 in a peri-urban community located near the city of Pune in Maharashtra, Western India, I examine the context and content of intergenerational differences in perceived morbidity among women. Among these women, perceived morbidity is an important gauge of the conceptual health transition taking place in urbanizing areas of India, and, potentially, it tells us far more about social change and its impact on health than do statistical representations and models. PMID- 12126275 TI - Breastfeeding, wage labor, and insufficient milk in peri-urban Kathmandu, Nepal. AB - This article presents a case study of breastfeeding mothers who are working as carpet-makers in peri-urban Kathmandu, Nepal. A sample of women surveyed about their current infant feeding practices revealed that half of the infants aged three to four months had been introduced to non-breast milk foods and liquids. During in-depth interviews some mothers explained that they supplemented breastfeeding with either milk or solids if they felt that they did not have enough breast milk for their infants. Reports of insufficient milk (IM) among these Nepali women is discussed within the larger context of IM as a worldwide phenomenon that is often associated with the cessation of breastfeeding and the switch to bottle-feeding based on commercial milk products. On average, the women in this study breastfed their infants until the latter were approximately three years of age. A status quo method for determining median duration of breastfeeding indicates that there is no significant difference in the duration of breastfeeding between mothers who work in carpet-making factories and those who spin wool at home. It is argued that reports of IM in this setting are not associated with the abandonment of breastfeeding, for a number of reasons including: the cultural approbation of breastfeeding; the low usage of baby bottles among peri-urban mothers, and the flexible labor practices of the carpet making industry. PMID- 12126276 TI - Adaptation, punctuation and information: a rate-distortion approach to non cognitive 'learning plateaus' in evolutionary process. AB - We extend recent information-theoretic phase transition approaches to evolutionary and cognitive process via the Rate Distortion and Joint Asymptotic Equipartition Theorems, in the circumstance of interaction with a highly structured environment. This suggests that learning plateaus in cognitive systems and punctuated equilibria in evolutionary process are formally analogous, even though evolution is not cognitive. Extending arguments by Adami et al. (2000), we argue that 'adaptation' is the process by which a distorted genetic image of a coherently structured environment is imposed upon a species. PMID- 12126278 TI - Methodology going astray in population biology. AB - This paper analyses the broad methodological structure of population-biological theorising. In it, I show that the distinction between initial exploratory, hypothesis-generating research and the subsequent process-reconstructing, hypothesis-testing type of research is not being made. Rather, the hypotheses generated in population biology are elaborated in such detail that students confound the initial research phase with the subsequent hypotheses-testing phase of research. In this context, I therefore analyse some testing procedures within the exploration phase and show that, as an extreme form of confusion, statistical null-models are mistakenly given the status of causal population-biological theory. PMID- 12126277 TI - Recent philosophy of biology: a review. AB - Academia is subdivided into separate disciplines, most of which are quite discrete. In this review I trace the interactions between two of these disciplines: biology and philosophy of biology. I concentrate on those topics that have the most extensive biological content: function, species, systematics, selection, reduction and development. In the final section of this paper I touch briefly on those issues that biologists and philosophers have addressed that do not have much in the way of biological content. PMID- 12126279 TI - Similarity among nucleotide sequences. AB - In this work we report a simple way to measure the similarity between two nucleotide sequences by using graph theory and information theory. This method reported allows for theoretical comparisons of naturally occurring nucleotide sequences. PMID- 12126281 TI - The Jamaican hypertension prevalence study. AB - This study was designed to investigate the prevalence of hypertension in Jamaica. Jamaica has an area of 4,411 square miles and is divided into 14 parishes. The visited districts were randomly selected. The sample population was selected based upon a two-stage stratified random sampling design. Each dwelling in the "Sampling Universe" had an equal probability of being selected. The survey team spent a week in the districts in each parish selected. Employing the Statistical Institute of Jamaica's (STATIN) two-stage stratified random sampling design, preselected house-holds were visited. Non-response was documented and considered in the final analysis. Only individuals 15 years and older were allowed to participate in the study. The 2,064 subjects who participated were the basis for estimates of hypertension. Following logistic regression analysis, the main risk factors for hypertension are being female, advancing age, obesity, having diabetes and having a family history of hypertension. Jamaica has a point prevalence of hypertension of 30.8% in the 15-and-over age group. These findings would greatly assist in formulating policies to deal with this scourge of society. PMID- 12126280 TI - Preventing diabetes-related morbidity and mortality in the primary care setting. AB - Diabetes is the leading cause of blindness, end-stage renal failure, non traumatic limb amputations, and cardiovascular morbidity and mortality. The vast majority of patients with diabetes receive routine care from primary care providers who are not endocrinologists. Primary care providers, including internists, family practice physicians, and physician extenders with advanced skills, face the important task of implementing standards of care recommendations for persons with diabetes. These recommendations draw upon an emerging body of compelling evidence regarding the prevention and management diabetes and its complications. The challenge of diabetes must be tackled on three fronts: Primary prevention, secondary prevention (of diabetes complications), and tertiary prevention (of morbidity and mortality from established complications). There is now abundant evidence that type 2 diabetes, which accounts for greater than 90% of diabetes world-wide, is preventable. Moreover, the complications of diabetes are preventable by a policy of tight glycemic control and comprehensive risk reduction. Even after complications have set in, intensive glucose control dramatically reduces the risk of progression of complications. The challenge, therefore, is the identification of strategies that enable translation of existing scientific data to pragmatic benefits. This article proposes 10 strategies for preventing or reducing diabetes-related morbidity and mortality at the primary care level. These strategies include provider education; patient empowerment through promotion of lifestyle and self-care practices; surveillance for microvascular complications; cardiovascular risk reduction; efficient use of medications; goal setting; and stratification of patients and triaging of those with poor glycemic control for more intensive management. PMID- 12126282 TI - Trend in the prevalence of overweight and obesity among urban African American hospital employees and public housing residents. AB - INTRODUCTION: To help understand the impact of socioeconomic status, we examined the current prevalence and age-specific trend in overweight and obesity among two socioeconomically diverse groups of African Americans in the Washington, DC, area. MATERIALS AND METHODS: Data on height and weight were collected between March 1995 and December 1996 as a part of nutrition survey to develop a food frequency questionnaire. Gender-stratified multiple logistic regression analyses were used to examine factors related to the current prevalence of overweight and obesity. RESULTS: Three hundred nine African American public housing residents and 293 African American hospital employees participated in this survey. Overall, hospital workers and public housing residents differed significantly in the distribution of BMI (p = 0.003). Among men, the prevalence of overweight and obesity were 34.9% and 29.4% for hospital workers and 27.0% and 18.2% for public housing residents, respectively. For females these rates were 31.3% and 46.3% for hospital employees and 26.1% and 42.9% for public housing residents, respectively. CONCLUSION: Overweight and obesity were highly prevalent among all age and socioeconomic groups. Future research should focus on a more in-depth study of the relationship between socioeconomic status and the correlates of obesity among African-Americans, particularly women. PMID- 12126283 TI - Urotherapy recommendations for bedwetting. AB - OBJECTIVE: To assess the efficacy of urotherapy recommendations prior to pharmacological or moisture alarm treatment in the management of bedwetting in children. METHODS: Children assessed for bedwetting at a voiding dysfunction clinic were admitted to a prospective, uncontrolled pilot study. The families were instructed to follow specific urotherapy recommendations. RESULTS: Of the 23 children who completed the study, sixteen (70%) improved with at least one less wet night per week, nine (39%) with at least a 50% reduction, and five (22%) resolved. CONCLUSION: Urotherapy recommendations prior to pharmacological or moisture alarm treatment shows promise and potential for the management of children with bedwetting. Further studies are necessary to determine if the improvement is sustained. PMID- 12126284 TI - The association of access to medical care with regular source of care and sociodemographic characteristics in patients with HIV and tuberculosis. AB - PURPOSE: To examine satisfaction with access to health care in two populations, one with HIV and one with TB, and examine the effect of having a regular doctor and sociodemographic characteristics. DESIGN: Cross-Sectional survey. PATIENTS: A sample of HIV inpatients hospitalized at seven Los Angeles sites (N = 217) and TB outpatients chosen randomly from the Los Angeles County TB Registry Census (N = 313). ANALYSIS: We performed bivariate and multivariate regression analyses of satisfaction with access to care on gender, race/ethnicity, age, education, income, insurance, and having a regular doctor. MAIN OUTCOME MEASURES: A six-item scale of satisfaction with access to care (range 0-100; Cronbach's alpha 0.87). RESULTS: The mean satisfaction with access score for the HIV sample was significantly lower than the TB sample (53.5 vs. 61.2, p<0.001). The HIV sample multivariate analysis (including all the variables) showed that increasing age (p<0.021 and having a regular doctor (p<0.002) were associated with better access, and that low income (p<0.005) was associated with poor access. In the TB sample analysis, only increasing age was associated with better satisfaction with access to care (p< 0.01). CONCLUSION: HIV patients receiving care in the private sector reported less satisfaction with access to care compared to TB patients receiving care in the public health sector. The traditional factors of socio economic status and having a regular doctor were associated with satisfaction with access-to-care in the HIV sample but not the TB sample. Our findings suggest that certain characteristics of the TB public health programs may explain these differences and suggests that, perhaps, the existence of a similar public health program for vulnerable low-income populations with HIV would improve their satisfaction with access, as well. PMID- 12126285 TI - Caveats in the neuropsychological assessment of African Americans. AB - This preliminary investigation examined the predictive accuracy of six neuropsychological tests in a population of non-brain-injured African Americans. False positives were unacceptably high on five of the neuropsychological tests administered. These pilot data raise important questions about the utility of neuropsychological test norms with groups dissimilar in sociocultural background to the normative population. These findings are examined in terms of the relative merits of the race-homogenous and race-comparative paradigms and underscore the importance of conducting normative studies that involve ethnic minority populations. PMID- 12126286 TI - The prevalence of victimization and perceptions of job neighborhood safety in a social service agency and the need for screening. AB - Because staff attitudes and affects are impediments or strengths in appropriately assessing and managing traumatized or violent patients, the authors discuss the need to explore staff's experience with trauma and violence. A survey of social service field and administrative office-based staff revealed that numerous staff had experienced traumatic events. Compared with staff based in a downtown administrative office, significantly more field-based staff were dissatisfied with their job's neighborhood and perceived their job's neighborhood as a dangerous place to work. Further, significantly more field staff perceived that their job's neighbors would be "unlikely" to intervene in observed antisocial behaviors. In our sample of primarily female, African-American, social service staff, the ratio of victimization was higher than rates in other studies. Screening social service staff for experiences of trauma and victimization and asking about their perceptions about their job's neighborhood will help leadership shape efforts to address these dynamics. By addressing staff's perceptions of their job's neighborhood, leadership can influence staff's willingness to participate in proactive community organizing and prevention activities designed to reduce violence and increase safety. Several recommendations to reduce violence in the workplace are presented. PMID- 12126287 TI - Participation of African Americans in a smoking cessation trial: a quantitative and qualitative study. AB - African Americans (AAs) have been considered a hard to reach research population. In a clinical trial of bupropion for smoking cessation, failure to return for randomization was concerning during the initial phase of recruitment. There were three study goals: to review the research on clinical trial participation barriers, to use quantitative analyses to identify differences between randomized (n = 66) and non-randomized (n = 54) participants, and to use focus groups and interviews (2 groups and 2 interviews, 17 participants) to elicit participation barriers and suggestions for participation enhancement. Randomized participants were older (44.1 vs. 37.6; p = 0.0019), predominately female (81.8% vs. 63.0%; p = 0.0201), more likely to have some college (33.3% vs. 16.7%; p = 0.0380), and less likely to be employed full time (32.4% vs. 53.7%; p = 0.0347). Randomized participants reported higher motivation to quit smoking (8.3 vs. 7.3; p = 0.0083) and higher confidence to quit (8.2 vs. 7.3; p = 0.0357). A logistic regression model specified age, gender, and motivation to quit, as predictors of randomization. Focus groups identified transportation, lack of readiness to quit, inadequate reminders, and employment conflicts as barriers to participation. Knowledge of differences between those who return for enrollment and those who do not may be used to increase AA enrollment in clinical trials. PMID- 12126288 TI - The changing pattern of prostate cancer in Nigerians: current status in the southeastern states. AB - This was a ten-year, hospital-based retrospective study for the incidence and clinical pattern of prostate cancer in southeastern Nigeria. Clinical information extracted from the files included the TNM stage, histo-pathological grading, level of prostatic acid phosphatase (PAP), mode of presentation and clinical and biochemical response to intravenous and oral diethylstilboestrol diphosphate (Honvan)/ orchidectomy. There were 145 patients, mean age 66.6 + 9.8 years, giving an incidence of 61.3 per 10(5), with 54% under 70 years. Most patients (81.4%) presented late, with 62% metastatic. Over 98% were adenocarcinomas, 77% of which were moderate to well-differentiated cancers. PAP was elevated in 109 patients (75%), (representing 92% of all advanced tumours), and normal in 36 (25%). Forty-two percent of poorly differentiated cancers had normal levels of PAP. Most patients presented with urinary retention (56%), prostatism (44%), anaemia (41%), recurrent UTI (35%), bone pains (20%), haematuria (18%), backache (16%) and paraplegia (6%). Nearly 79% responded to treatment with lowered PAP levels and improved quality of life, within a mean of 26.3+/-13.8 months (range 5 78); objective 81 (58%), subjective 32 (23%), no response 27 (19%). Among paraplegics, 78% had full, and 22% had partial motor recovery. Patients with poorly differentiated cancers had only a 33% two-year survival rate. This study confirmed an upward, though moderate trend in the incidence of prostate cancer in Nigeria. The use of PAP instead of PSA as the tumor marker, a local diet with high fish content but lower animal fat, and poor hospital access may account for the lower incidence in the southeast. Poor health education may account for the high rate of late presentations. PMID- 12126289 TI - Understanding insulin-resistance in type 2 diabetes. PMID- 12126290 TI - SAT review course. PMID- 12126291 TI - Epiphany in the doctor's lounge. PMID- 12126292 TI - Albumin, cancer & pregnancy. PMID- 12126293 TI - Mad scientists, compassionate healers, and greedy egotists: the portrayal of physicians in the movies. AB - Cinematic depictions of physicians potentially can affect public expectations and the patient-physician relationship, but little attention has been devoted to portrayals of physicians in movies. The objective of the study was the analysis of cinematic depictions of physicians to determine common demographic attributes of movie physicians, major themes, and whether portrayals have changed over time. All movies released on videotape with physicians as main characters and readily available to the public were viewed in their entirety. Data were collected on physician characteristics, diagnoses, and medical accuracy, and dialogue concerning physicians was transcribed. The results showed that in the 131 films, movie physicians were significantly more likely to be male (p < 0.00001), White (p < 0.00001), and < 40 years of age (p < 0.009). The proportion of women and minority film physicians has declined steadily in recent decades. Movie physicians are most commonly surgeons (33%), psychiatrists (26%), and family practitioners (18%). Physicians were portrayed negatively in 44% of movies, and since the 1960s positive portrayals declined while negative portrayals increased. Physicians frequently are depicted as greedy, egotistical, uncaring, and unethical, especially in recent films. Medical inaccuracies occurred in 27% of films. Compassion and idealism were common in early physician movies but are increasingly scarce in recent decades. A recurrent theme is the "mad scientist," the physician-researcher that values research more than patients' welfare. Portrayals of physicians as egotistical and materialistic have increased, whereas sexism and racism have waned. Movies from the past two decades have explored critical issues surrounding medical ethics and managed care. We conclude that negative cinematic portrayals of physicians are on the rise, which may adversely affect patient expectations and the patient-physician relationship. Nevertheless, films about physicians can serve as useful gauges of public opinion about the medical profession, as tools for medical education, and as instruments of positive social change in efforts to reform managed care. PMID- 12126294 TI - Theoretical mechanisms in targeted and random integration of transgene DNA. AB - The genetic manipulation of mammalian cells and animals would be greatly expedited if gene targeting could be reliably achieved in the widest possible range of host cell types. This paper considers empirical evidence and theoretical considerations associated with transgene integration, and concludes that utilisation of gene targeting in non-selective systems awaits further progress in modelling homologous recombination. PMID- 12126296 TI - Heterogeneity of circulating prolactin in the bitch. AB - Different molecular forms of circulating prolactin (PRL) are known to occur in several species. As no such information was available in dogs, we assessed the molecular profile of circulating PRL in bitches. Pooled sera from covertly (CTRL) and overtly pseudopregnant (PSPT) diestrous bitches with high or low (> 10 or < 10 ng x mL(-1), respectively) serum PRL (measured by ELISA) were analyzed by Sephadex G-100 and Concanavalin A-Sepharose column chromatography. Four serum PRL fractions were identified and termed big-big, big (> 67 kDa), native (23 kDa) and fragmented (< 20) kDa) PRL. The percentages of these fractions were roughly similar in CTRL and PSPT animals, irrespective of their serum PRL levels (higher in PSPT than in CTRL bitches). A large proportion of glycosylated PRL (between 69 and 100%) was also detected in these sera. We conclude that in dogs, circulating PRL occurs in multiple molecular forms, whose relative abundance is comparable in covertly and overtly pseudopregnant bitches. PMID- 12126295 TI - Relation between dietary lipid level and voluntary feed intake, growth, nutrient gain, lipid deposition and hepatic lipogenesis in rainbow trout. AB - Four diets with differing lipid contents (15, 20, 25 or 30% DM) were tested on small (initial body weight: 27 g) and larger (IBW: 93 g) rainbow trout (Oncorhynchus mykiss) fed on demand or by hand, respectively. In both trials, voluntary feed intake was inversely related to dietary lipid levels. Protein efficiency increased when dietary fat content increased. Final whole-body lipid content was positively related to dietary lipid levels. The main sites of lipid storage were visceral adipose tissue and to a lesser extent muscle. Increased fat deposition in the visceral cavity of young trout was due to both hyperplasic and hypertrophic responses and in larger trout mostly due to a hypertrophic response. Liver activities of glucose-6-phosphate dehydrogenase and fatty acid synthetase were negatively correlated with fat intake and positively with starch intake, whereas malic enzyme was little affected by dietary treatments. PMID- 12126297 TI - Correlation between microbial enzyme activities in the rumen fluid of sheep under different treatments. AB - Five total mixed rations prepared from finger millet (Eleusine Coracana) straw as a roughage (48%) and mixed concentrate (52%), supplemented with a 1% isoacid mixture (i-C4, i-C5, C5 and phenylacetic acid in equal proportions) or oil (groundnut oil, 5% more than the control) or urea (5% more nitrogen than the control), and protein (groundnut cake, 5% more nitrogen than the control) were given in a Latin square experiment to sheep. Enzymatic activities were estimated for urease, cellulase, protease, amylase, and lipase in various fractions of rumen fluid on the one hand and rumen microbial biomass on the other hand. Rumen samples were taken 3-4 hours after feeding and mixed rumen bacteria were separated as a strained rumen fluid without protozoa (SRFWP), cell free rumen fluid (CFRF) and enzymes associated with the bacteria cell (EABC). Samples of SRFWP and EABC contained higher enzyme activities than CFRF. Depending on the type of enzymes in each fraction, some significant coefficient of determination (r2) was seen. These values showed very close cooperative action between proteolytic and amylolytic enzymes under the experimental conditions, or perhaps the presence of some species of bacteria with both activities. Lipolytic bacteria are completely specialized for lipase production only (P < 0.05). The results showed oil, isoacid and crude protein enhanced microbial production (P < 0.05) and this can change the pattern of enzymes in the rumen of sheep. PMID- 12126298 TI - Controlled ovarian stimulation and ultrasound guided follicular aspiration in the baboon (Papio cynocephalus anubis). AB - The objective of this study was to investigate whether baboon females respond to an ovarian stimulation protocol incorporating pituitary suppression with a GnRH agonist (GnRHa) and highly purified human FSH (hphFSH) with follicular development and oocyte maturation. An ovulation induction protocol was applied to 5 adult female baboons with a history of regular menstrual cycles (33-34 days). A long-acting GnRHa implant containing goserelin acetate was placed s.c. on days 22 24 of their menstrual cycle. Daily hphFSH (75 IU im) treatments were started approximately 10 days following menses. When the majority of the follicles were > or = 5 mm in diameter and the E2 levels had reached a maximum, hCG (2000 IU i.m.) was administered to induce final maturation of the oocytes and ovulation. 30 to 34 h after hCG administration, transabdominal follicular aspiration was performed using a variable frequency transvaginal transducer with ultrasound. A total of 71 oocytes were collected (average: 17). 91% of the oocytes were morphologically normal indicating that they were appropriate for in vitro insemination. PMID- 12126299 TI - Coexistence of domains with distinct order and polarity in fluid bacterial membranes. AB - In this study we sought the detection and characterization of bacterial membrane domains. Fluorescence generalized polarization (GP) spectra of laurdan-labeled Escherichia coli and temperature dependencies of both laurdan's GP and fluorescence anisotropy of 1,3-diphenyl-1,3,5-hexatriene (DPH) (rDPH) affirmed that at physiological temperatures, the E. coli membrane is in a liquid crystalline phase. However, the strong excitation wavelength dependence of rlaurdan at 37 degrees C reflects membrane heterogeneity. Time-resolved fluorescence emission spectra, which display distinct biphasic redshift kinetics, verified the coexistence of two subpopulations of laurdan. In the initial phase, <50 ps, the redshift in the spectral mass center is much faster for laurdan excited at the blue edge (350 nm), whereas at longer time intervals, similar kinetics is observed upon excitation at either blue or red edge (400 nm). Excitation in the blue region selects laurdan molecules presumably located in a lipid domain in which fast intramolecular relaxation and low anisotropy characterize laurdan's emission. In the proteo-lipid domain, laurdan motion and conformation are restricted as exhibited by a slower relaxation rate, higher anisotropy and a lower GP value. Triple-Gaussian decomposition of laurdan emission spectra showed a sharp phase transition in the temperature dependence of individual components when excited in the blue but not in the red region. At least two kinds of domains of distinct polarity and order are suggested to coexist in the liquid-crystalline bacterial membrane: a lipid-enriched and a proteolipid domain. In bacteria with chloramphenicol (Cam)-inhibited protein synthesis, laurdan showed reduced polarity and restoration of an isoemissive point in the temperature-dependent spectra. These results suggest a decrease in membrane heterogeneity caused by Cam-induced domain dissipation. PMID- 12126300 TI - Circadian gating of photoinduction of commitment to cell-cycle transitions in relation to photoperiodic control of cell reproduction in Euglena. AB - A novel type of circadian and photoperiodic control of the cell division cycle was found in photoautotrophic Euglena gracilis. When algae entrained to 24 h light-dark (LD) cycles (14 h L) were transferred to continuous darkness (DD) at the eighth hour of the final LD photoperiod, cell-cycle transition was arrested in phase G1, S or G2. The subsequent exposure of these dark-arrested cells to a 6 h light-break allowed the dark-arrested cells to undergo cell-cycle progression in DD, in a manner dependent on the circadian phase; maximum photoinduction occurred around dusk. Inhibitor experiments suggested that the photoinduced commitment of G2 cells to cell division required light for a signal originating in noncyclic photosynthetic electron transport (PET), particularly cytochrome b6 f but not for the metabolic energy required by the process. The fact that the circadian rhythm of photoinduction ran out-of-phase from that of noncyclic PET signaling suggests that the site of regulation by the former rhythm is downstream of noncyclic PET. The occurrence of maximum photoinduction around dusk suggests that the 'external coincidence' model of photoperiodic induction describes the activation of the photoinductive phase. Further evidence supporting this hypothesis is the relationship between cell reproduction and day length; the resulting sigmoidal curve indicates a combined effect of photosynthesizing period and circadian stimulation around dusk. Circadian control is shown to be an integral part of the mechanism for 24 h LD cycle-induced synchronous cell division. PMID- 12126301 TI - High-efficiency energy transfer from carotenoids to a phthalocyanine in an artificial photosynthetic antenna. AB - Two carotenoid pigments have been linked as axial ligands to the central silicon atom of a phthalocyanine derivative, forming molecular triad 1. Laser flash studies on the femtosecond and picosecond time scales show that both the carotenoid S1 and S2 excited states act as donor states in 1, resulting in highly efficient singlet energy transfer from the carotenoids to the phthalocyanine. Triplet energy transfer in the opposite direction was also observed. In polar solvents efficient electron transfer from a carotenoid to the phthalocyanine excited singlet state yields a charge-separated state that recombines to the ground state of 1. PMID- 12126302 TI - Excitation pulse deconvolution in luminescence lifetime analysis for oxygen measurements in vivo. AB - Oxygen-dependent quenching of phosphorescence has been proven to be a valuable tool for the measurement of oxygen concentrations both in vitro and in vivo. For biological measurements the relatively long lifetimes of phosphorescence have promoted time-domain-based devices using xenon arc flashlamps as the most common excitation light source. The resulting complex form of the excitation pulse leads to complications in the analysis of phosphorescence lifetimes and ultimately to errors in the recovered pO2 values. Although the problem has been recognized, the consequences on in vivo phosphorescence lifetime measurements have been neglected so far. In this study, the consequences of finite excitation flash duration are analyzed using computer simulations, and a method for the recovery of phosphorescence decay times from complex photometric signals is presented. The analysis provides an explanation as to why different calibration constants are reported in the literature and presents a unified explanation whereby calibration constants are not solely a property of the dye but also of the measuring device. It is concluded that complex excitation pulse patterns without appropriate analysis methods lead to device-specific calibration constants and nonlinearity and can be a potent source of errors when applied in vivo. The method of analysis presented in this article allows reliable phosphorescence lifetime measurements to be made for oxygen pressure measurements and can easily be applied to existing phosphorimeters. PMID- 12126303 TI - Frequency-domain measurement of the photodegradation process of fluorescein. AB - The frequency-domain technique is applied to measure the photodegradation rate of fluorescein in aqueous solutions. The illuminating light is modulated, and the changes in fluorescence from the illuminated region are detected synchronously. A constant flow rate is imposed on the fluorescein solution to control the mass transport of fluorescein into the illuminated region. The fluorescence response is described by a model that assumes that photodegradation occurs from the triplet excited state. The predictions of the model are consistent with the observed variations in the fluorescence response with flow rate, modulation frequency and incident power. We discuss in this article how the dependence of the model parameters on experimental conditions can be used to infer the photodegradation rate as well as some of the details of the photodegradation mechanism. The results are consistent with the known mechanism of photodegradation of fluorescein. The frequency-domain technique gives a photodegradation rate of 53 s(-1) in an air-saturated solution and 37 s(-1) in solutions purged with argon gas. PMID- 12126304 TI - Ionic photodissociation in arylallyl acetates. AB - We have synthesized several 3-arylallyl acetates 1, 2, 3, 5 and 6, and E-3-(1 naphthyl)-2-propene-1-ol 4 for studying ionic photodissociation. Compounds 1, 2 and 3 underwent an efficient ionic photodissociation in polar solvents like acetonitrile and methanol leading to the formation of rearranged acetate and methyl ether products, as well as undergoing an E-Z isomerization. The arylallyl alcohol 4 and the two arylallyl acetates 5 and 6 did not undergo ionic photodissociation. Quantum yields of product formation, quantum yields of fluorescence, solvent polarity effects and triplet-sensitization studies indicated that a highly polarized excited singlet state is responsible for the ionic photodissociation. Both the singlet- and triplet-excited states are effective in displaying E-Z isomerization in 1, 2, 3 and 4. Compounds 5 and 6 are highly fluorescent, and the fluorescence may be the excited state deactivation pathway along with internal conversion. PMID- 12126305 TI - Singlet oxygen-mediated protein oxidation: evidence for the formation of reactive side chain peroxides on tyrosine residues. AB - Singlet oxygen (1O2) is generated by a number of enzymes as well as by UV or visible light in the presence of a sensitizer and has been proposed as a damaging agent in a number of pathologies including cataract, sunburn, and skin cancers. Proteins, and Cys, Met, Trp, Tyr and His side chains in particular, are major targets for 1O2 as a result of their abundance and high rate constants for reaction. In this study it is shown that long-lived peroxides are formed on free Tyr, Tyr residues in peptides and proteins, and model compounds on exposure to 1O2 generated by both photochemical and chemical methods. The yield of these species is significantly enhanced in D2O and decreased by azide. Nuclear magnetic resonance and mass spectroscopic analysis of reaction mixtures, or materials separated by high-performance liquid chromatography, are consistent with the initial formation of an (undetected) endoperoxide that undergoes rapid ring opening to give a hydroperoxide situated at the C1 ring-position (i.e. para to the phenolic group). In the presence of a free alpha-amino group (e.g. with free Tyr), rapid ring-closure occurs to give an indolic hydroperoxide that decays into the corresponding alcohol, 3a-hydroxy-6-oxo-2,3,3a,6,7,7a-hexahydro-1H-indole-2 carboxylic acid. Hydroperoxides that lack a free alpha-amino group (e.g. those formed on 3-(4-hydroxyphenyl)propionic acid, N-Ac-Tyr and Tyr-containing peptides) are longer-lived, with half-lives of hours to days. These species undergo slow decay at low temperatures to give the corresponding alcohol. Their rate of decay is enhanced at 37 degrees C, or on exposure to UV light or metal ions, and gives rise to reactive radicals, via cleavage of the peroxide bond. These radicals have been characterized by electron paramagnetic resonance spin trapping. These studies demonstrate that long-lived Tyr-derived peroxides are formed on proteins exposed to 1O2 and that these may promote damage to other targets via further radical generation. PMID- 12126306 TI - The gold porphyrin first excited singlet state. AB - Gold porphyrins are often used as electron-accepting chromophores in artificial photosynthetic constructs. Because of the heavy atom effect, the gold porphyrin first-excited singlet state undergoes rapid intersystem crossing to form the triplet state. The lowest triplet state can undergo a reduction by electron donation from a nearby porphyrin or another moiety. In addition, it can be involved in triplet-triplet energy transfer interactions with other chromophores. In contrast, little has been known about the short-lived singlet excited state. In this work, ultrafast time-resolved absorption spectroscopy has been used to investigate the singlet excited state of Au(III) 5,15-bis(3,5-di-t-butylphenyl) 2,8,12,18,-tetraethyl-3,7,13,17-tetramethylporphyrin in ethanol solution. The excited singlet state is found to form with the laser pulse and decay with a time constant of 240 fs to give the triplet state. The triplet returns to the ground state with a life-time of 400 ps. The lifetime of the singlet state is comparable with the time constants for energy and photoinduced electron transfer in some model and natural photosynthetic systems. Thus, it is kinetically competent to take part in such processes in suitably designed supermolecular systems. PMID- 12126307 TI - Antimicrobial DNA-binding photosensitizers from the common rush, Juncus effusus. AB - Our continuing survey of phototoxins from higher plants has led to the isolation and identification from the common rush, Juncus effusus L., of the phenanthrene, dehydroeffusol (1), and the dihydrophenanthrene, juncusol (2), compounds that display enhanced antimicrobial activities in light. The antimicrobial activities (minimum inhibitory concentrations) for these compounds against methicillin resistant and -sensitive Staphylococcus aureus and Candida albicans were increased 16- and two-fold, respectively, by irradiation with ultraviolet A (UVA). Photosensitized DNA-binding activities (as possible covalent bond formation) of these compounds were determined by using restriction enzymes and a specially prepared 1.5 kb DNA fragment. Under UVA irradiation, dehydroeffusol strongly inhibited all the restriction enzymes (KpnI, XbaI, PmeI, DraI, PacI and BciVI) that have at least one 5'-TpA sequence in their recognition sites. Weak inhibitions were found for the restriction enzymes EcoRI, SacI, BamHI, SalI, PstI and HindIII, which do not possess a 5'-TpA sequence at their restriction sites and the restriction site sequences of which consist of all bases, A, T, G and C. Trace or no inhibition was found for AscI and SmaI, the restriction site sequences of which are composed of only C and G. The results indicate the necessity of thymine (adenine) for the photosensitized DNA-binding activity of dehydroeffusol. A strong inhibition against SphI, which does not have a 5'-TpA sequence in the restriction sequence, indicates that there are possibly other binding sequence(s) for dehydroeffusol. With juncusol and UVA, strong inhibitions for KpnI and BciVI and trace inhibitions for PacI, XbaI, PmeI and DraI were found. This result also showed a preference of juncusol for 5'-TpA, but the preference could be more selective than that of dehydroeffusol depending on the surrounding sequences of 5'-TpA in the respective restriction sites. A strong inhibition of SphI by juncusol with UVA also indicated the existence of an unknown binding sequence for this compound. Generally, the DNA-binding activity of this compound was weaker than that of dehydroeffusol. PMID- 12126308 TI - Observation and quantification of ultraviolet-induced reactive oxygen species in ex vivo human skin. AB - Two-photon fluorescence imaging is used to detect UV-induced reactive oxygen species (ROS) in ex vivo human skin in this study. ROS (potentially H202, singlet oxygen or peroxynitrite [or all]) are detected after reaction with nonfluorescent dihydrorhodamine-123 (DHR) and the consequent formation of fluorescent rhodamine 123 (R123). The cellular regions at each epidermal stratum that generate ROS are identified. R-123 fluorescence is detected predominately in the lipid matrix of the stratum corneum. In contrast, the strongest R123 fluorescence signal is detected in the intracellular cytoplasm of the viable epidermal keratinocytes. A simple bimolecular one-step kinetic model is used for estimating the upper bound of the number of ROS that are generated in the skin and that react with DHR. After ultraviolet-B radiation (280-320 nm) (UVB) equivalent to 2 h of noonday summer North American solar exposure (1600 J m(-2) UVB), the model finds that 14.70 x 10(-3) mol of ROS that react with DHR are generated in the stratum corneum of an average adult-size face (258 cm(-2)). Approximately 10(-4) mol are potentially generated in the lower epidermal strata. The data show that two photon fluorescence imaging can be used to detect ROS in UV-irradiated skin. PMID- 12126309 TI - On the importance of spectral responsivity of Robertson-Berger-type ultraviolet radiometers for long-term observations. AB - A system to determine the spectral responsivity of ultraviolet (UV) radiometers has been developed and is routinely operated at the Central Ultraviolet Calibration Facility, at the National Oceanic and Atmospheric Administration. The instrument and the measurement methodologies are described. Results of measurements from thermally controlled broadband UV radiometers of the Robertson Berger (R-B)-type are described. Systematic differences in the spectral response curves for these instruments have been detected. The effect of these differences on the field operation of UV-B radiometers has been studied by calculating the instrumental response from modeled UV spectra. The differences of the weighted spectral UV irradiances, measured by two radiometers with different spectral response functions, caused by the daily variation in the position of the sun were estimated for fixed values of total ozone, altitude and albedo, and for cloud free conditions. These differences increase with the solar zenith angle and are as large as 8%. Larger differences in the instrumental response may be produced by ozone variations. Thus, care must be taken when analyzing data from R-B radiometers and comparing results from different instruments. Routine cycling of UV-B radiometers in operative networks without a careful determination of the spectral responsivity, or small drifts of the spectral responsivity, may strongly affect the accuracy of UV radiation measurements and produce an erroneous trend. Because of the possible differences among radiometers, it would not be practical to derive the long-term behavior of UV radiation without routine and thorough characterization of the spectral responsivities of the instruments. PMID- 12126310 TI - Cyclooxygenase-2 expression in murine and human nonmelanoma skin cancers: implications for therapeutic approaches. AB - Inflammatory stimuli result in the production of cutaneous eicosanoids, which are known to contribute to the process of tumor promotion. Cyclooxygenase (COX), the rate-limiting enzyme for the production of prostaglandins (PG) from arachidonic acid, exists in at least two isoforms, COX-1 and COX-2. COX-1 is constitutively expressed in most tissues and plays various physiological roles, whereas increased COX-2 expression is known to occur in several types of epithelial neoplasms. Enhanced PG synthesis is a potential contributing factor in UVB induced nonmelanoma skin cancers (NMSC). Increased COX-2 staining occurs in murine skin neoplasms after chronic exposure to carcinogenic doses of UVB. In this study, immunohistochemical and Western blot analyses were employed to assess longitudinally COX-2 expression in a standard mouse UVB complete carcinogenesis protocol and in human basal cell carcinomas (BCC) and squamous cell carcinomas (SCC). During UVB irradiation of mice, COX-2 expression consistently increased in the hyperplastic skin, the benign papillomas and the SCC. COX-2 expression was also increased in human actinic keratoses, SCC and BCC as well as in murine SCC and BCC. The pattern of COX-2 expression was quite variable, occurring in a patchy distribution in some lesions with staining confined mainly to suprabasal cell layers. In general, COX-2 expression progressively became more extensive in benign papillomas and well-differentiated murine SCC. The staining was predominantly cytoplasmic and perinuclear in some focal areas in tissue stroma around both murine and human tumors. Western blot analysis confirmed negative COX 2 expression in normal skin, whereas acute UVB exposure resulted in increased enzyme expression, which continued to increase in developing papillomas and SCC. Because of the evidence indicating a pathogenic role for eicosanoids in murine and human skin neoplasms, we performed studies to assess the anti-inflammatory and anticarcinogenic effects of green tea extracts, which are potent antioxidants. Acute exposure of the human skin to UVB (minimum erythema dose x 4) caused a transient enhancement of the COX-2 expression, which reverted to baseline within hours; however, in murine skin the expression persisted for several days. Pretreatment with the topically applied green tea extract (1 mg/cm2) largely abrogated the acute COX-2 response to UVB in mice or humans. In summary, enhanced COX-2 expression serves as a marker of epidermal UVB exposure for murine and human NMSC. These results suggest that COX-2 inhibitors could have potent anticarcinogenic effects in UVB-induced skin cancer. PMID- 12126311 TI - In vivo pharmacokinetics of 8-aminolevulinic acid-induced protoporphyrin IX during pre- and post-photodynamic therapy in 7,12-dimethylbenz(a)nthracene treated skin carcinogenesis in Swiss mice: a comparison by three-compartment model. AB - Delta-aminolevulinic acid-photodynamic therapy (ALA-PDT) has emerged as a useful technique in the treatment of superficial basal cell carcinoma, actinic keratosis, squamous cell carcinoma and tumors of other organs. Earlier reports mention that there is reappearance of protoporphyrin IX (PpIX) after photoirradiation of tumors. This property of reappearance of PpIX is being utilized to treat nodular tumors by fractionated light dose delivery. However, there is still no unanimously accepted reason for this reappearance phenomenon and the rate of resynthesis after PDT. On account of this, studies are carried out on the estimation of the pharmacokinetics of the ALA-induced PpIX in mice tumor models and the surrounding normal tissues before and after PDT. Further, a mathematical model based on a multiple compartment system is proposed to estimate the rate parameter for the diffusion of PpIX from the surrounding normal tissues into the tumor tissue (km) caused by photobleaching during PDT with irradiating fluences of 36.0 and 57.6 J/cm2. The km value at two different fluences, 36.0 and 57.6 J/cm2, are estimated as 3.0636+/-0.7083 h(-1) and 6.9231+/-2.17651 h(-1), respectively. Further, the rate parameter for the cleavage and efflux of ALA (k1) and the rate parameter for the evasion of PpIX from the tumor tissues after PDT (kt) were also estimated by fitting the experimental data to the developed mathematical model. The statistical significance of the estimated parameters was determined using Student's t-test. The experimental results and the rate parameters obtained using the proposed compartment model suggest that in addition to the earlier reported reasons, the invasion or diffusion of PpIX from the surrounding tissues to the tumor tissues after photoirradiation might also contribute to the reappearance of PpIX after PDT. PMID- 12126312 TI - The role of the peripheral benzodiazepine receptor in photodynamic activity of certain pyropheophorbide ether photosensitizers: albumin site II as a surrogate marker for activity. AB - A study has been carried out to define the importance of the peripheral benzodiazepine receptor (PBR) as a binding site for a series of chlorin-type photosensitizers, pyropheophorbide-a ethers, the subject of a previous quantitative structure-activity relationship study by us. The effects of the PBR ligand PK11195 on the photodynamic activity have been determined in vivo for certain members of this series of alkyl-substituted ethers: two of the most active derivatives (hexyl and heptyl), the least active derivative (dodecyl [C12]) and one of intermediate activity (octyl [C8]). The photodynamic therapy (PDT) effect was inhibited by PK11195 for both of the most active derivatives, but no effect on PDT activity was found for the less active C12 or C8 ethers. The inhibitory effects of PK11195 were predicted by the binding of only the active derivatives to the benzodiazepine site on albumin, ie. human serum albumin (HSA) Site II. Thus, as with certain other types of photosensitizers, it has been demonstrated with this series of pyropheophorbide ethers that in vitro binding to HSA-Site II is a predictor of both optimal in vivo activity and binding to the PBR in vivo. PMID- 12126314 TI - Failure to thrive: aid and Africa. PMID- 12126313 TI - Up-regulation of Hsp27 plays a role in the resistance of human colon carcinoma HT29 cells to photooxidative stress. AB - The Photofrin-resistant cell line (HT29-P14) was used in the present study to investigate the mechanism(s) involved in Photofrin-mediated photodynamic therapy (PDT). We compared gene expression profiles between the resistant cell line and its parental cell line (HT29) using DNA microarray analysis. A significant up regulation of small heat shock protein 27 (Hsp27) was found in HT29-P14 cells. The elevated Hsp27 level may play an important role in the resistance of HT29-P14 to Photofrin-PDT. To test this hypothesis, we stably transfected HT29 cells with human Hsp27 complementary DNA. The potential role of Hsp27 in the resistance to PDT was examined in Hsp27-overexpressing cells. Stable trasnfected cells (H13) showed an increased survival after Photofrin-PDT, suggesting that the up regulation of Hsp27 is related to the induced resistance to Photofrin-PDT. Phosphorylation of Hsp27 has been suggested to play an important role in cytoprotection. We have examined the phosphorylation activity of Hsp27 among the parental and resistant cells, as well as the overexpression cells. An elevated level of Hsp27 resulted in an increased ability of phosphorylation in both resistant and overexpressing cells after PDT. The activation of the phosphorylation of Hsp27 induced by PDT was not mediated by the p38 mitogen activated protein kinase. These data suggest that Hsp27 may play an important role in mediating the adaptive response to Photofrin-PDT-induced oxidative stress and that the pathways leading to Hsp27 phosphorylation may contribute to the resistance of the cells to photooxidative damage. PMID- 12126315 TI - Digging for data from the COX-2 trials. PMID- 12126316 TI - Safe drug prescribing. PMID- 12126317 TI - tPA for acute stroke: balancing baseline imbalances. PMID- 12126318 TI - Safe drug prescribing. PMID- 12126319 TI - Impact of reference-based pricing for histamine-2 receptor antagonists and restricted access for proton pump inhibitors in British Columbia. AB - BACKGROUND: Two programs to reduce expenditures for common gastrointestinal drugs were introduced simultaneously by British Columbia (BC) Pharmacare in 1995. Reference-based pricing restricted reimbursement for all histamine-2 receptor antagonists (H2RAs) to the cost of the least expensive H2RA available, generic cimetidine. Special authority restricted reimbursement for proton pump inhibitors (PPIs) to patients who met certain eligibility criteria. We evaluated the effect of reference-based pricing for H2RAs and special authority for PPIs on dispensing and reimbursement for senior citizen beneficiaries of BC Pharmacare. METHODS: Itemized monthly claims data for upper gastrointestinal drugs were obtained from BC Pharmacare for all beneficiaries 65 years of age or older. Periods before and after implementation of reference-based pricing and special authority were compared with respect to defined daily doses dispensed per 100,000 beneficiaries, BC Pharmacare reimbursement per 100,000 beneficiaries, BC Pharmacare reimbursement per defined daily dose and beneficiary contributions per defined daily dose. We used regression models to project forward trends in expenditures observed before implementation of the new policies and hence to estimate accrued cost savings. RESULTS: Before reference-based pricing and special authority, the numbers of defined daily doses that were dispensed and total BC Pharmacare reimbursements for H2RAs appeared to be declining gradually, whereas those for PPIs were rising. With reference-based pricing, the monthly defined daily dose of cimetidine dispensed increased more than 4-fold, to 116,257 per 100,000 beneficiaries, while those of other restricted H2RAs decreased by more than half, to 50,927 per 100,000 beneficiaries. Special authority immediately reduced the dispensed volumes of PPIs by one-fourth, but growth in volume then appeared to resume at its previous rate. The estimated annualized cost savings achieved by reference-based pricing and special authority were $1.8 million to $3.2 million for H2RAs (depending on the estimation method used) and $5.5 million for PPIs. However, beneficiary contributions for H2RAs increased from negligible amounts to approximately 16% of total drug expenditures. INTERPRETATION: Reference-based pricing and special authority appear to have been successful in altering prescribing habits and reducing provincial expenditures for upper gastrointestinal drugs, but they have increased the financial burden on senior citizen beneficiaries. PMID- 12126321 TI - Risk of death or readmission among people discharged from hospital on Fridays. PMID- 12126320 TI - Long-term results of pediatric liver transplantation in a combined pediatric and adult transplant program. AB - BACKGROUND: Liver transplantation is now routine therapy for a variety of childhood liver diseases; however, there are no detailed reports of long-term results from a Canadian centre. We reviewed data from the first 16 years of a pediatric liver transplantation program to determine survival, complications and long-term outcomes. METHODS: The outcomes to December 2000 for all children (age less than 18 years) who received a liver transplant at the London Health Sciences Centre between April 1984 and December 1999 were reviewed. The recipients were grouped according to the period in which they received the transplant (period 1, April 1984 to July 1988; period 2, August 1988 to December 1993; or period 3, January 1994 to December 1999). Data were obtained from medical charts; in-person interviews, questionnaires or telephone contact with patients and their families; contact with referring physicians; and school records. Outcome measures included patient survival, retransplantation, complications and long-term outcomes (specifically steroid withdrawal and growth and development). RESULTS: A total of 116 children (29 in period 1, 46 in period 2 and 41 in period 3) (median age 5.6 years at the time of the procedure) received a total of 140 liver grafts (32 in period 1, 57 in period 2 and 51 in period 3). Of the 116 patients, 23 (20%) were less than 1 year old at the time of transplantation. Biliary atresia was the most common indication for liver transplantation (57 [49%] of the 116 patients). The number of patients surviving to 1 year after transplantation was 20 (69%) of the 29 patients from period 1, 40 (87%) of the 46 patients from period 2 and 38 (93%) of the 41 patients from period 3. The percentage of patients receiving reduced size grafts from adult donors, including live donors, increased from 2/32 (6%) in period 1 to 22/51 (43%) in period 3. Retransplantation was required for 9 (31%) of the 29 patients from period 1, 6 (13%) of the 46 patients from period 2 and 7 (17%) of the 41 patients from period 3. Among these patients, 1-year survival was 33% (3/9) for period 1, 83% (5/6) for period 2 and 100% (7/7) for period 3. Eighteen of the 22 deaths occurred within 4 months after surgery. Only 3 (3%) of the 116 patients experienced post-transplant lymphoproliferative disease. Steroids were discontinued (usually within 2 years after surgery) for the following proportions of surviving transplant recipients: 17 (89%) of the 19 patients from period 1, 29 (78%) of the 37 patients from period 2 and 21 (55%) of the 38 patients from period 3. Most of the surviving patients had normal growth (82/94 or 87%) and development (73/94 or 78%), and these outcomes were consistent across the 3 periods of study. INTERPRETATION: Survival and long-term outcomes continue to improve for most children who receive liver transplants. These improvements may be due to improved surgical technique, perioperative care and, to a lesser extent, immunosuppressive therapy. PMID- 12126322 TI - Potential solutions to the problem of conducting systematic reviews of new health technologies. PMID- 12126323 TI - Missing the point (estimate)? Confidence intervals for the number needed to treat. PMID- 12126324 TI - Expanding the health care debate. PMID- 12126325 TI - Problems for clinical judgement: 5. Principles of influence in medical practice. AB - The basic science of psychology has identified specific ingrained responses that are fundamental elements of human nature, underpin common influence strategies and may apply in medical settings. People feel a sense of obligation to repay a perceived debt. A request becomes more attractive when preceded by a marginally worse request. The drive to act consistently will persist even if demands escalate. Peer pressure is intense when people face uncertainty. The image of the requester influences the attractiveness of a request. Authorities have power beyond their expertise. Opportunities appear more valuable when they appear less available. These 7 responses were discovered decades ago in psychology research and seem intuitively understood in the business world, but they are rarely discussed in medical texts. An awareness of these principles can provide a framework for physicians to help patients change their behaviour and to understand how others in society sometime alter patients' choices. PMID- 12126326 TI - Identifying and managing adverse environmental health effects: 6. Carbon monoxide poisoning. AB - Carbon monoxide poisoning is an enigmatic illness. The symptoms are often non specific or masked by an exacerbation of an underlying illness, such as congestive heart failure, that has been triggered by carbon monoxide inhalation. The effects can range from mild, annoying symptoms relieved by removal of the source to severe morbidity with profound central nervous system dysfunction, acute complications and delayed sequelae. Estimates suggest that about one-third of nonfatal cases of carbon monoxide poisoning go undetected and undiagnosed. We present a case of residential carbon monoxide poisoning to illustrate these points and to demonstrate the usefulness of a simple tool based on the CH2OPD2 mnemonic (Community, Home, Hobbies, Occupation, Personal habits, Diet and Drugs) that physicians can use to obtain an environmental exposure history. We outline the clinical management of carbon monoxide poisoning and provide strategies and resources to prevent exposure. PMID- 12126327 TI - Buruli ulcer: the third most common mycobacterial infection. PMID- 12126329 TI - Atopic eczema in children. PMID- 12126328 TI - What's all the fuss? Safety concerns about COX-2 inhibitors rofecoxib (Vioxx) and celecoxib (Celebrex). PMID- 12126330 TI - Red hemiscrotum in a child with abdominal pain. PMID- 12126331 TI - Romanow heard it all: spend more, spend less; privatize, don't privatize... PMID- 12126332 TI - Stem cell controversy continues as Ottawa tables bill. PMID- 12126333 TI - Post-traumatic stress an occupational hazard for journalists? PMID- 12126334 TI - Israel criticized after Palestinian MDs shot, killed by soldiers in West Bank. PMID- 12126335 TI - Health care's declining years. PMID- 12126336 TI - Pressure mounting to curb MD poaching by rich nations. PMID- 12126337 TI - Chronic tension-type headache--therapeutic options. PMID- 12126338 TI - Sertraline in chronic tension-type headache. AB - OBJECTIVES: To study the effect of Sertraline, a selective serotonin reuptake inhibitor in chronic tension type headache (CTTH) in a double blind placebo controlled randomized trial. MATERIAL AND METHODS: The study design was a double blind placebo controlled randomized study of 50 patients, over a period of 10 weeks including run-in period of two weeks, treatment period four weeks, and follow-up four weeks. The diagnosis of CTTH was based on criteria given by International Headache Society (IHS). The overall response was graded in terms of headache index, analgesic intake per week and percentage reduction in headache frequency using chi-square and 't' test. Anxiety and depression was assessed using Hamilton Rating Scale. RESULTS: The mean analgesic intake per week declined from 4.34 +/- 0.736 tab/week to 1.07 +/- 0.592 tab/week in sertraline group (p < 0.01) while reduction in placebo group was not significant (3.98 +/- 0.729 tab/week to 2.94 +/- 0.665 tab/week) (p > 0.05). The reduction in headache index and percentage reduction in frequency of headache was not significant in drug treatment group. Side effects were seen in 24.4% of patients nausea, nervousness and dizziness being the commonest. CONCLUSION: The drug treatment group showed a significant decline in analgesic intake per week. The study result shows that sertraline can be a useful alternative to amitryptiline in those patients who fail to respond or cannot tolerate it. PMID- 12126339 TI - Changing scenario of transfusion-related viral infections. AB - OBJECTIVE: To study the changing incidence of blood transfusion-related viral infections consequent to compulsory screening of blood and greater awareness of the problem, over the last five years. METHODS: This is a cross-sectional study carried out at Medical College, Calcutta. Three groups each consisting of 100 subjects were selected for this study. Group A comprised multiple transfused patients who have also received transfusion before 1995. Group B comprised patients who had received transfusions only since 1995. Group C comprised of control patients who have never been transfused. The incidence of HBsAg +ve, anti HCV +ve and HIV +ve cases were calculated and expressed as percentages and compared using the chi square test. RESULTS: The incidences of HBsAg +ve and anti HCV +ve cases in the three groups were 20% and 16% in Group A, 7% and 6% in Group B and 4% and 2% in Group C. The difference between Group A and Group B were statistically significant. CONCLUSION: The incidence of HBsAg and anti-HCV positive cases among the multi-transfused has decreased over the last five years. PMID- 12126340 TI - Kidney functions in newly diagnosed type 2 diabetic subjects: role of glycemic control. AB - BACKGROUND: The increased urinary albumin excretion (UAE) found at the diagnosis in type 2 diabetes may at least be functional and, thus reversible, as in type 1 diabetes. Furthermore, a positive correlation has been found between the glycemic control and UAE in type 2 diabetic subjects. MATERIAL AND METHODS: The impact of glycemic control after achieving three months of continuous normo-glycemia was studied in 26 (including 16 males and 10 females) newly diagnosed type 2 diabetic patients to assess renal size/volumes, various renal functions and UAE. RESULTS: A significant reduction was seen in glomerular filtration rate (GFR), UAE and fractional albumin clearance after achieving near normoglycemia (p < 0.01). A decrease in the kidney size/volume, though statistically insignificant, was also observed. Besides a decline in GFR of 4.8% was also noted after three months of stable normoglycemia. UAE normalized in six of these subjects. A statistically significant correlation was also found between the changes in mean fasting plasma glucose and the decline in UAE (r=0.56 and p < 0.01). CONCLUSION: The study emphasizes the necessity to correct the apparently reversible increase in microabuminuria that accompanies hyperglycemia prior to patients recruitment for intervention therapy. PMID- 12126341 TI - Gallbladder disorders and type 2 diabetes mellitus--a clinic-based study. AB - AIMS OF THE STUDY: To study the prevalence of gallbladder disorders in type 2 diabetic patients and their correlation with patient factors like age, sex, weight, duration of diabetes and autonomic neuropathy. METHODOLOGY: Fifty type 2 diabetic patients and 30 healthy controls underwent realtime ultrasonography to study the prevalence of gallbladder disorders. The fasting gallbladder volume and contraction 60 minutes after a fatty meal of the diabetic subjects were compared with 30 age and sex matched healthy volunteers. The age, sex, weight, duration of diabetes, autonomic neuropathy, control of diabetes were correlated to the prevalence of gallbladder disorders in diabetic patients. RESULTS: 32% of the diabetic patients had ultrasonographic evidence of gallstones, as compared to 6.7% in healthy subjects. 73.7% of the diabetic patients with gallbladder disorders were females. Mean fasting gallbladder volume was significantly increased in diabetic patients (26.2 cm3) as compared to non-diabetic healthy subjects (15.8 cm3). Further mean fasting gallbladder volume of diabetic patients with gallbladder disorder (28.1 cm3) was found to be significantly larger than that of those patients without gallbladder disorder (24.6 cm3). Mean percentage of contractions of gallbladder 60 min after fatty meal was reduced in diabetic patients (53.1%) and it was observed to be further reduced in the patients with gallbladder disorder (41.8%). Mean fasting gallbladder volume was larger in diabetic subjects with autonomic neuropathy, than those without. However, difference in mean percentage contraction of gallbladder 60 min after fatty meal was not statistically significant. Mean duration of diabetes was significantly longer in diabetic patients with gallbladder disorder. CONCLUSIONS: We conclude that type 2 diabetic patients have increased prevalence of gallbladder disorder which can only partially be explained by autonomic neuropathy leading, to increased fasting volume. Factors like decreased cholecystokinin or decreased sensitivity of the smooth muscle of gallbladder to normal level of cholecystokinin need to be studied. PMID- 12126342 TI - Bone marrow trephine biopsy as an adjunct to bone marrow aspiration. AB - OBJECTIVE: To determine the relative efficacy of bone marrow aspiration as compared to that of trephine biopsy. METHODS: Bone marrow aspiration and bilateral trephine biopsies were performed in 420 consecutive cases. The diagnosis and findings made on bone marrow aspiration were compared with that made on trephine biopsy in each case. RESULTS: Aspiration alone was sufficient in making a diagnosis in 372 (88.6%) cases as it correlated well with the diagnosis made on trephine sections. In the remaining 48 (11.4%) cases trephine biopsy was necessary for making a diagnosis due to incomplete information provided by aspiration or its inability to give a correct diagnosis. These cases were mostly hypoplastic/aplastic marrow, myelofibrosis and marrow involvement by metastatic tumour and lymphomatous infiltration. Often a bilateral marrow biopsy picked up the diagnostic lesion. CONCLUSION: The decision to perform a marrow aspiration alone or in combination with marrow biopsy depends on the diagnosis being considered. In nutritional anaemias, most hematologic malignancies and immune thrombocytopenias, marrow aspiration alone is sufficient, but for detection of disorders with focal marrow involvement bilateral marrow biopsies are a must. PMID- 12126343 TI - Cognitive behavioural intervention in bronchial asthma. AB - OBJECTIVE: The aim of the present study was to find out the efficacy of cognitive behaviour therapy, as an adjunct to standard pharmacotherapy in bronchial asthma. DESIGN: An experimental design with pre- and post-therapy assessments was adopted. SETTING: The Medicine Out-patient Department of St. John's Medical College and Hospital, and Department of Clinical Psychology, NIMHANS, Bangalore. PATIENTS: Ten asthma patients who fulfilled the inclusion and exclusion criteria, matched for use of drugs, were sequentially allotted to two groups: a) experimental group, who were exposed to cognitive behaviour therapy along with standard pharmacotherapy, b) control group, who were exposed to standard pharmacotherapy alone. INTERVENTION: Cognitive behaviour therapy included 15 individual sessions consisting of asthma education, Jacobson progressive muscle relaxation (JPMR), behavioural techniques, cognitive restructuring, cognitive coping skills and behavioural counseling to significant others. MEASUREMENTS: The measures used for pre- and post-therapy assessments were--Asthma symptom checklist, asthma diary, state trait anxiety inventory-Y1 and Y2, Beck depression inventory, asthma quality of life questionnaire and peak expiratory flow rate. RESULTS: There was significant decrease in asthma symptoms, anxiety and depression; and significant increase in quality of life in the experimental group (p < 0.05) at the post-assessment. The control group did not show any significant change at the post-assessment. CONCLUSION: Cognitive behaviour therapy helps in improving the management of asthma. PMID- 12126344 TI - Relationship of tissue plasminogen activator, plasminogen activator inhibitor-1 and fibrinogen with coronary artery disease in South Indian male subjects. AB - AIM: Prevalence rates of coronary artery disease (CAD) are reported to be very high in Asian Indians. Conventional risk factors do not explain the high rates of CAD among Indians. Recently, several newer risk factors have been reported to be associated with CAD. We measured tissue plasminogen activator (tPA) antigen, plasminogen activator inhibitor-1 (PAI-1) and fibrinogen levels in South Indian diabetic and non-diabetic subjects with and without CAD. METHODS: Four groups of subjects were studied (all males); Group 1 comprised of non-diabetic subjects without CAD (n=50). Non-diabetic subjects with CAD formed group 2 (n=50); group 3 comprised of type 2 diabetic patients without CAD (n=50) and group 4 consisted of type 2 diabetic patients with CAD (n=50). CAD was diagnosed based on coronary angiographic evidence of severe double or triple vessel disease. RESULTS: Both diabetic and non-diabetic patients with CAD had significantly higher levels of tPA, PAI-1 and fibrinogen compared to non-diabetic without CAD (p < 0.05). Patients with CAD were distributed more in the upper tertiles of these risk factors compared to those without CAD. A strong association between tPA and PAI-1 was noted in the Pearson's correlation analysis (p < 0.001). Univariate regression analysis showed tPA (Odds ratio--1.12, p = 0.03), PAI-1 (Odds ratio- 1.03, p = 0.008), fibrinogen (Odds ratio--1.01, p < 0.0001), serum cholesterol (Odds ratio--1.008, p = 0.04) and hypertension (Odds ratio--3.7, p = 0.0001) to be associated with CAD. Multiple logistic regression analysis revealed hypertension (Odds ratio--4.6, 95% confidence interval--2.113-9.950, p = 0.0001) and fibrinogen (Odds ratio--1.012, 95% confidence interval--1.007-1.018, p = 0.0001) as risk factors for CAD. CONCLUSION: Our study suggests that prothrombogenic risk factors particularly fibrinogen may be associated with CAD in South Indians. PMID- 12126345 TI - Treatment of AA amyloidosis in rheumatoid arthritis. AB - Four patients of rheumatoid arthritis (RA) with biopsy confirmed AA amyloidosis were treated with chlorambucil. All had established but uncontrolled RA with a persistently raised ESR. Moderate (> 1 gm, < 3.5 gm/d) to nephrotic range (> 3.5 gm/d) proteinuria and a relatively well preserved renal function was noted in three patients. One patient had deranged renal function and required dialysis. On chlorambucil, there was complete recovery, partial improvement and no improvement in one patient each. The fourth patient required haemodialysis, did not tolerate chlorambucil and succumbed to the illness. Therapy with chlorambucil can benefit some patients of RA with AA amyloidosis. Leucopenia is the most important dose limiting side effect. PMID- 12126346 TI - Bacteriological infections during the first hundred days of allogenic bone marrow transplantation--experience from Oman. AB - BACKGROUND: Allogeneic bone marrow transplantation (ABMT) produces severe neutropenia lasting for days. During the first 100 days of allogeneic BMT bacterial infection is a major cause of mortality and morbidity. This is specially so till the patient engrafts with good neutrophil count (> 500/microl). Nature of bacterial infection and its sensitivity pattern partly reflects the patient's own flora, hospital flora and the antibiotic usage pattern in the hospital and the community. Hence although we know quite a lot about the nature of bacterial sepsis following ABMT quite well, its details vary from centre to centre. Hence the present study was undertaken at a newly developed BMT unit at Muscat, Oman. MATERIAL AND METHODS: Twenty one patients receiving allogeneic BMT for different indications were included in this study. These patients had blood culture from central and peripheral line whenever they developed fever > 38 degrees C lasting more than two hours and the relevant investigators like X-ray chest, USG, CRP levels, urine culture was done as and when required. RESULTS: All patients in BMT unit developed fever > 38 degrees C during first 100 days but only nine patients showed positive blood culture on 12 occasions. 42% of these isolates were gram-positive organisms. No fungal infections were noticed. Twenty percent of the gram-negative isolates from adjoining pardiatric oncology ward were resistant to ciprofloxacin though this is a reserved drug in this hospital. One out of 21 BMT patient in the present study died due to sepsis with resistant Klebsiella organism. Imipenem as a single drug did not cover all the organisms isolated. Listeria monocytogenes was an unusual organism in our BMT patient. CONCLUSION: The present result of less than 5% mortality in the ABMT patients due to sepsis is excellent and this is related to early marrow recovery and efficient microbiological surveillance in this centre. Cautions should be exercised while using imipenem as a single antibiotic in ABMT situations. Though ciprofloxacin is a reserved drug in this hospital, increased incidence of resistance to this antibiotic probably represents its wide usage in the community. PMID- 12126347 TI - Problem of timely diagnosis in anthrax meningitis. AB - Anthrax continues to remain a problem in parts of India. Meningitis is often a complication encountered among cases with cutaneous anthrax. We have encountered a dozen cases of anthrax meningitis in our hosptal in the past decade. A sudden unexplained rise in cases in the past two years with hundred percent mortality stresses the need for rapid confirmatory diagnosis. Most of the cases admitted with central nervous system involvement had a provisional diagnosis of conditions other than anthrax meningitis. A strong clinical suspicion with a simple Gram stain smear of the CSF will help confirm anthrax meningitis in endemic areas. PMID- 12126348 TI - The human leucocyte antigen (HLA) system. PMID- 12126349 TI - Postal stamps released on John Gregor Mendel (1822-1884). PMID- 12126350 TI - Pictorial CME. A break in the lead density below the clavicle suggestive of fractured lead. PMID- 12126351 TI - Critical evaluation of available medical literature--a general guideline for a busy physician. PMID- 12126352 TI - HLA and bone marrow transplant donor search strategies. AB - Searching for a family donor other than an HLA identical sibling can be successful. The chance of finding an identical or one antigen mismatched family donor can be calculated with a computer program, so this may help in the decision making whether to perform extended family typing or not. Extended family donors are often better matched rather than unrelated donors. The reason is that they have at least one haplotype genetically identical to the patient, and that there may be some higher match grade for the minor histocompatibility antigens. Sometimes extended family donor could be the only donor for the patient. Ethical considerations have to be taken into account and extended family donors should be volunteer donors. PMID- 12126353 TI - Dysrexia nervosa. PMID- 12126354 TI - New frontiers of therapy in hemato-oncology. PMID- 12126355 TI - Isolated duodenal metastasis from breast carcinoma. AB - Small bowel neoplasms comprise 0.1% of all malignancies. Of these one-third are duodenal, mainly primary. Isolated duodenal secondaries arising on account of haematogenous spread is rare. Our case of duodenal obstruction due to an isolated duodenal metastasis as a result of haematogenous spread from breast carcinoma is unique in the literature. PMID- 12126356 TI - Renal failure in a patient with primary antiphospholipid syndrome. AB - Antiphospholipid antibody syndrome (APLA) is a syndrome complex characterized by recurrent arterial or venous thrombosis, recurrent fetal wastage, thrombocytopenia and presence in serum of antibodies against negatively charged phospholipids like lupus anticoagulant (LA), anticardiolipin antibody (ACLA) and subgroups. These are classified further as primary (wherein this occurs in isolation) and secondary (associated with infection, drugs and malignancies). It is uncommon to find both LA and ACLA in primary APLA syndrome (unlike as in the secondary form). Renal manifestations which include renal arterial and venous occlusion and infarction and thrombotic microangiopathy have also been infrequently described. We hereby present a case of primary APLA syndrome with unusual features of LA and ACLA occurring together and also the presence of renal failure due to left renal artery thrombosis and right renal artery occlusion. PMID- 12126357 TI - Isolated necrotising arteritis of the cervix and myometrium: a much neglected but puzzling entity. AB - Arteritis of the uterine cervix and corpus described here was an incidental finding at the routine histopathological examination of the hysterectomy specimen resected from a 62 year old female who underwent laparotomy for twisted ovarian cyst. Investigations and eleven months of follow up without any specific treatment for arteritis, have shown no systemic involvement. This case highlights that a knowledge of such isolated arteritis is of importance to the physician to avoid misdiagnosing it as polyarteritis nodosa and treat with systemic steroids. PMID- 12126358 TI - Renal amyloidosis in a pentazocine addict. PMID- 12126359 TI - Bronchobiliary fistula: an anaesthetic point of view. AB - Bronchobiliary fistula is defined as the passage of bile in the bronchi and in the sputum (bilioptysis). This rare disorder is associated with significant morbidity. Authors review the anaesthetic management of bronchobiliary fistula and recommend the use of double lumen endotracheal tube even in cases with a closed/sealed bronchobiliary fistula. PMID- 12126360 TI - An unusual presentation of the Papillon-Lefevre syndrome as recurrent liver abscesses. AB - The Papillon-Lefevre syndrome is a rare genetic disorder with a predisposition to severe infections. We describe Papillon-Lefevre syndrome in a 17 year old boy from a family where four out of eight siblings were affected with this disease and who presented with recurrent pyogenic liver abscesses over a period of 9 years, an association never previously reported. PMID- 12126361 TI - Granulomatous angiitis of the central nervous system associated with herpes zoster. AB - Granulomatous angiitis of central nervous system (CNS) is a rare inflammatory disease of blood vessels mostly confined to CNS. We describe a case which presented with right sided hemiplegia with aphasia, after herpes zoster ophthalmicus. CT scan and MRI brain showed a large left sided infarct in the left middle cerebral artery (MCA) territory. MRI angiography revealed narrowing and thinning of left internal carotid artery (ICA) and to a lesser extent, left MCA suggestive of granulomatous vasculitis. Herpes zoster is often associated with major CNS involvement and a vascular etiology was previously postulated. Recent pathological reports suggest that cerebral angiitis secondary to herpes virus infection may be more common than realised. PMID- 12126362 TI - Visceral leishmaniasis: a rare cause of post-transplant fever and pancytopenia. AB - Despite the endemic distribution of visceral leishmaniasis in certain parts of our country, there are only a few reports of this infection in renal transplant recipients. We report one renal transplant recipient from non-endemic area with visceral leishmaniasis and graft dysfunction that responded to treatment with stibogluconate. The infection should be considered in the differential diagnosis of a febrile transplant recipient with pancytopenia and allograft dysfunction. PMID- 12126363 TI - IgA myeloma presenting as diabetes mellitus with refractory anaemia. PMID- 12126364 TI - Malignant insulinoma. PMID- 12126365 TI - Pregnancy with functioning adrenocortical carcinoma. PMID- 12126366 TI - High resolution computed tomography (HRCT) of the lungs of people living near the garbage dumping grounds in Deonar, Mumbai--a pilot study. PMID- 12126367 TI - Pre-hospital issues in acute myocardial infarction. PMID- 12126368 TI - Role of API in promoting medical education. PMID- 12126369 TI - Value of tuberculin skin test to diagnose tuberculosis among patients with respiratory systems in a chest clinic. PMID- 12126370 TI - Gulliane Barre syndrome following vaccination with hepatitis B vaccine. PMID- 12126371 TI - Bringing mental health into the mainstream. PMID- 12126372 TI - CME programme of Indian College of Physicians at Shirdi. PMID- 12126373 TI - Depositions of nitrogen on NiTi instruments. AB - This study was designed to obtain nitrogen-rich layers on the surfaces of endodontic files made of nickel-titanium (NiTi) alloy by chemical vapor deposition. Experimental samples (GT rotary instruments) were deposited by using two different methods. The first one was based on the reaction of wet NH3 with NiTi under high temperatures (300 degrees C). The second technique is a typical MOCVD (metal organic chemical vapor deposition) procedure that uses Ti(Et2N)4 as a titanium and nitrogen precursor. Control samples were not exposed to any process. The chemical composition of the surface layers of each sample was determined by means of X-ray photoelectron spectroscopy and X-ray diffraction measurements. The experimental instruments showed surface chemical composition that was different from that seen in the control group; samples treated with gaseous NH3 showed a surface nitrogen/titanium (N/Ti) ratio = 0.9; MOCVD instruments showed a surface N/Ti ratio of 2; control samples showed a N/Ti ratio = 0.2; MOCVD of nitrogen ion of nickel-titanium files produced a higher concentration of nitrogen on the surface. PMID- 12126374 TI - The demineralizing effects of EDTA at different concentrations and pH. AB - The purpose of this study was to compare the effects of concentration and pH variations of EDTA on dentin demineralization. Twenty extracted, human permanent teeth with single canals were used in this study. Demineralizing effects of EDTA solutions at 10% and 17% concentrations at pH 7.5 and 9.0 were determined by measuring the amount of liberated phosphorus 1, 3, 5,10, and 15 min after exposure. The results showed that the amount of phosphorus liberated from dentin was greater with increased EDTA concentration and increased time of exposure, and it was more effective at neutral pH than pH 9.0. The pH of the EDTA solutions did not display any significant alterations during the demineralization process. PMID- 12126375 TI - The effect of a bioresorbable matrix barrier in endodontic surgery on the rate of periapical healing: an in vivo study. AB - The purpose of this prospective clinical study was to assess the rate of healing of periapical bony defects created at the time of endodontic periapical surgery by measuring the densitometric ratio change in periapical osseous tissues, after periapical endodontic surgery was performed, by using or not using Guidor bioresorbable membrane material. Periapical surgeries were performed on 25 patients where nonsurgical root canal therapy had failed and a periapical lesion was present. For inclusion in the study, the osseous defect to be analyzed had to be confined to the apical area, with bone covering the entire root surface coronally, and an intact lingual cortical plate had to be present. A series of radiographs at the 3-, 6-, and 12-month recalls were compared with the radiograph taken immediately after surgery by using digital imaging and a densitometric ratio that gave a numerical estimation of osseous healing. Using repeated-measure ANOVA, it was shown that there was no statistical difference between the rate of healing in those cases where a Guidor resorbable membrane was used and those cases where no membrane was used. The results suggest that placement of a guided tissue membrane over the bony opening created during an endodontic periapical surgical procedure has no beneficial effect on the rate of healing and the added expense to the patient would not be warranted in these cases. PMID- 12126376 TI - Observation of depth and incidence of the mesial groove between the mesiobuccal and mesiolingual orifices in mandibular molars. AB - The purpose of this study was to analyze the mesiolingual-mesiobuccal groove indentations at the pulpal floor in mandibular first and second molars. The depth and incidence of occurrence were assessed. Sixty freshly extracted, mature teeth were kept in a 2.5% sodium hypochlorite for 5 to 7 days. The teeth were accessed without touching the floor of the chamber. The dental debris was washed thoroughly and then the patency and the presence of two mesial canals were established with a #10 file. The teeth were placed in 2.5% sodium hypochlorite for 3 to 5 days and later rinsed and air-dried before placement in centrifuge tubes. A vinyl polysiloxane impression material was injected into the chambers, and the teeth were centrifuged. The impressions were carefully removed and then measured by using a dissecting microscope and a transparent millimeter ruler. The ruler, at zero, placed at the mid-floor area of the impression and viewed from the mesial, measured (to the nearest 0.5 mm) the depth of the mesial groove between the mesiolingual and mesiobuccal canals. Due to imperfect impressions, 50 teeth were included in the study. The recorded average in depth was 1.0 mm. Some of the impressions had depths measuring 3.5 mm. This could be a significant space when considering the limitations of instrumentation techniques. There is the question of whether this area may remain untouched, thereby effecting the prognosis of treatment in both vital and nonvital teeth. Modifications in access preparation and/or an increased emphasis on irrigation and intracanal medication may be needed. PMID- 12126377 TI - Patency file and apical transportation: an in vitro study. AB - The purpose of this study was to evaluate the transportation produced at the apical foramen by the use of stainless steel and nickel-titanium K-files #10, #15, #20, and #25 as a patency file. Thirty human maxillary lateral incisors were transversally cut at 4 mm from the apex and mounted in square pieces of silicone with special marks to ensure repositioning of the specimens. The specimens were randomly divided in two equal groups of 15. In group A, stainless steel K-files #10, #15, #20, and #25 were used as a patency file. In group B, a stainless steel K-file #10 followed by nickel-titanium K-files #15, #20, and #25 were used as in group A. Sodium hypochlorite was used as the irrigating solution. Photographic slides of the apical foramen of each specimen before instrumentation and after the use of each file size were taken. The photographic slides of each specimen were mounted in slide mounts, projected onto the same drawing paper, and superimposed according to the peripheral shape of the root. Transportation of the apical foramen was determined comparing the five drawings of each specimen. Transportation was detected in 18 of the 30 specimens; 9 in group A and 9 in group B. No statistically significant differences were seen between groups (p = 1.00). PMID- 12126378 TI - In vitro evaluation of furcal perforation repair using mineral trioxide aggregate or resin modified glass lonomer cement with and without the use of the operating microscope. AB - This study evaluated in vitro the effect of using the operating microscope on repairing furcation perforations using Vitrebond or mineral trioxide aggregate. Forty-six human molar teeth were mounted into a jig attached to a simulated jaw. The teeth were allocated randomly to four groups (n = 10). Furcal perforations were made in the teeth using an ISO 012 round bur in a slow-speed hand-piece. Each material was used to repair a group of teeth with and without the use of the operating microscope. The remaining six teeth provided positive and negative controls. All groups were stored in 100% humidity, and the repair materials were allowed to set for 72 h at room temperature before being assessed for the quality of placement under x26 magnification. Leakage at the repair was then tested using India ink; the teeth were demineralized, dehydrated in alcohol, and rendered transparent in methyl salicylate. Dye penetration into the furcation repair was evaluated at x26 magnification. There was no difference in the acceptability of the repair with either material whether or not the operating microscope was used. The perforations repaired with mineral trioxide aggregate leaked significantly less to the tracer dye than those repaired with Vitrebond (p < 0.001). PMID- 12126379 TI - Lateral condensation in canals prepared with nickel titanium rotary instruments: an evaluation of the use of three different master cones. AB - Forty-five single-rooted teeth were instrumented with ProFile ISO .06 nickel titanium rotary instruments and obturated with lateral condensation using three different master cones: an ISO-standardized gutta-percha cone (group A), a Dia ISO(GT).06 gutta-percha cone (group B), and a size medium gutta-percha cone (group C). Obturation efficiency in each group was evaluated by recording the number of accessory cones utilized. To evaluate obturation quality in the three groups, a cross-sectional observation of the ability of the gutta-percha in each group to obliterate the prepared canal space was carried out. Results showed that obturation efficiency was significantly greater in groups B and C than in group A. There was no significant difference in obturation quality between any of the three groups. PMID- 12126380 TI - New core and sealer materials for root canal obturation and retrofilling. AB - A new endodontic obturation and retrofilling material has been invented to satisfy Grossman's requirements for an ideal material. Methylene blue dye leakage studies were carried out to compare the new material with conventional products, for both obturation and retrofilling. The experimental material when used in root canal obturation had a mean apical leakage of 0.49+/-0.27 mm, compared with 3.75+/-2.81 mm for the conventional material (p < 0.01, t test). Retrofilling leakage was not significantly different from that of a glass-ionomer (p < 0.01, t test) but was substantially lower than that of MTA and Super EBA (p < 0.01, Chi square test). It was concluded that leakage in root canal obturation was reduced by an order of magnitude and in retrofilling was significantly better than clinically advocated materials. It was inferred that the adhesion mechanism used has contributed to the reduction in leakage. PMID- 12126381 TI - The effect of an apex locator on exposure to radiation during endodontic therapy. AB - The objective of this study was to investigate whether the use of an electronic apex locator results in a reduction in X-ray exposure during endodontic therapy. Fifty, sound, extracted, single-rooted canine and incisor teeth were selected and randomly divided into two groups (A and B) of 25 teeth. The working length was determined radiographically with (group B) and without (group A) adjunctive use of an apex locator by one operator. After working length determination, the teeth were sectioned and the actual working length measured for comparison. For group A, 14 retake radiographs were required to determine the working length, whereas group B required no retake radiographs; this difference was highly significant (p < 0.001). The electronic apex locator was extremely accurate in locating the apical foramen with all teeth tested within 0.5 mm of the anatomical apex and 11 (44%) teeth at the apical foramen. In contrast, 15 (60%) teeth tested using radiographs alone were within 0.5 mm of the anatomical apex and only 4 (16%) teeth were actually at the anatomical apex. It was concluded that using an electronic apex locator as an aid to endodontic therapy could potentially reduce the number of diagnostic radiographs required for working length determination. Location of the apical foramen using a combination of an electronic apex locator and radiographs to determine working length is more accurate than using radiographs alone. PMID- 12126383 TI - Comparison of carbon dioxide versus refrigerant spray to determine pulpal responsiveness. AB - There is little evidence for which cold delivery method is most accurate in determining pulp responsiveness. This study compared carbon dioxide dry ice sticks (CO2) versus refrigerant spray (RS) to generate a patient response from different types of teeth restored to varying degrees. Fifteen human patients were selected. In each quadrant, three teeth were identified and the restoration type recorded. Blindfolded patients were randomly tested with either CO2 or RS in a crossover design at two different sessions. Responsiveness was recorded as yes/no and the interval (in seconds) from application to response was determined. Results showed that (a) CO2 and RS were equivalent in producing a pulpal response regardless of tooth and presence of restoration, and (b) CO2 took significantly (p < 0.05) longer to evoke a response than RS using paired t tests. In conclusion, RS and CO2 were equivalent in determining pulpal responsiveness, but the elicited response from RS was faster. PMID- 12126382 TI - Case selection for apical surgery: a retrospective evaluation of associated factors and rational. AB - Endodontic failures associated with poor quality of endodontics respond favorably to retreatment. Nevertheless, under certain clinical conditions, apicoectomy should be the preferred procedure. A retrospective survey of 200 roots that were referred for apical surgery revealed that 83% of the roots were inadequately obturated, including 8.5% with no root canal filling at all. In 49 of the roots in this group (24.5% of the referred cases) nonsurgical retreatment was judged by an endodontist as either impossible or improbable because it might jeopardize the root integrity. Retreatment should have been the preferred treatment modality for the rest of the group, provided that coronal restorations could be safely bypassed or removed. Posts were found in 63 of these teeth, however 35 of them were either short or loosely fitting and could safely be removed. The rest of the posts were longer than 5 mm, which might have presented a problem if their removal was attempted. In 45% of the 200 cases in the present study, surgical intervention was justified. The rest of the cases (55%) should have either been subjected to a follow-up (10.5%) or retreated nonsurgically by a skilled endodontist (44.5%). These results indicate that referring dentists may not appreciate the retreatment possibilities offered by modern endodontics, and they emphasize the need for a shift of concept: endodontists should be involved in the decision making before referring a patient to surgery. PMID- 12126384 TI - Obturation of accessory canals after four different final irrigation regimes. AB - The aim of this study was to evaluate accessory canal obturation after four different final irrigation regimes. Sixty-four extracted human lower premolars were instrumented, divided randomly into four groups, and finally irrigated as follows: no irrigation (NO), distilled water (WA), sodium hypochlorite ([SH] 6%, 20 ml for 15 min), and EDTA (15%, 8 ml for 3 min) combined with the SH regimen (ES). The System B and the Obtura II were used to fill the canals. The teeth were cleared and observed under a digital microscope. Obturation material penetration rates in length into the accessory canals were measured and the following values were obtained: 22.3% in group NO, 21.8% in group WA, 53.5% and 68.1% in groups SH and ES, respectively. Statistically significant differences were found when comparing each of the first two groups (NO and WA) with SH and ES groups (p < 0.05). The use of sodium hypochlorite alone or in combination with EDTA in final irrigation flushes may enhance a better obturation material penetration into the accessory canals. PMID- 12126385 TI - Effect of narcotic pain reliever on pulp tests in women. AB - The purpose of this study was to determine the effect of one dose of a common narcotic-based pain reliever (Vicodin) on a battery of oral sensitivity tests across time in women. Fifteen Caucasian women randomly were given an oral dose of 10 mg of hydrocodone/1000 mg of acetaminophen or placebo in a double-blind, cross over design. At baseline (before drug) and after 2, 4, and 8 h each subject was evaluated for sensitivity thresholds with four tests around an experimental tooth: (a) electric pulp tester applied to exposed root; (b) electric pulp tester on adjacent mucosa; (c) increasing probe pressure (grams) on adjacent mucosa; and (d) decreasing cold probe (degrees C) on the exposed root. The outcomes of all tests were not statistically different between drug and placebo treatments at any time point (p > 0.05). These results suggest that a systemic dose of hydrocodone/acetaminophen has little impact on healthy pulp or mucosa sensitivity in women as measured by common diagnostic tests. PMID- 12126386 TI - The effect of rotational speed and the curvature of root canals on the breakage of rotary endodontic instruments. AB - One of the biggest concerns when working with rotary endodontic instruments is that the instrument might break in the root canal thus compromising the treatment. This study evaluated the effect of rotational speed and the curvature of root canals on the breakage of ProFile rotary instruments to determine whether either causes an increase in instrument breakage. One hundred twenty root canals of extracted molars were divided into two groups according to whether the curvature of the canal was greater or lesser than 30 degrees. Three different rotational speeds (150, 250, and 350 rpm) were then used to carry out the instrumentation of the canals. The curvature of the canals would seem to be the most important factor that increases the risk of instrument breakage. PMID- 12126387 TI - The effect of preflaring on the rates of separation for 0.04 taper nickel titanium rotary instruments. AB - The purpose of this study was to compare the rates of separation of 0.04 taper nickel titanium (NiTi) rotary instruments using two different instrumentation techniques. Twenty sets of 0.04 taper Profile Series 29 rotary instruments, sizes 2 to 6 were used in the mesial (mandibular) or buccal (maxillary) canals of extracted human molars with a 20 to 30 degree root curvature according to the Schneider classification. The rotary instruments were used up to 20 times either with the crown-down technique recommended by the manufacturer or with a combination of preflaring with hand files in a passive step-back technique followed by rotary instrumentation. Statistical analysis of the data showed that the combination technique allowed more uses before separation compared with the crown-down technique recommended by the manufacturer p < 0.0001. PMID- 12126388 TI - Failure of treatment--or failure of the patient to heal? PMID- 12126389 TI - Catalytic oxidations of steroid substrates by artificial cytochrome p-450 enzymes. AB - Catalysts comprising manganese-porphyrins carrying cyclodextrin binding groups are able to perform hydroxylations with substrate selectivity and regio- and stereoselectivity and high catalytic turnovers. The geometries of the catalyst/substrate complexes override intrinsic substrate reactivities, permitting attack on geometrically accessible saturated carbons of steroids in the presence of secondary carbinol groups and carbon-carbon double bonds, as in enzymatic reactions. Selective hydroxylations of steroid carbon 9 positions are of particular practical interest. PMID- 12126390 TI - Theoretical study of factors controlling rates of cyclization of radical intermediates from diallylamine and diallylammonium monomers in radical polymerizations. AB - The radical cyclization reactions of models for the growing radical chains formed from N,N-diallylamine (1), N-methyl-N,N-diallylamine (2), N,N-diallylammonium (3), N-methyl-N,N-diallylammonium (4) and N,N-dimethyl-N,N-diallylammonium (5) have been investigated computationally by DFT theory, using the B3LYP functional. Models formed by hydrogen atom addition to dienes 1-5 undergo five-membered ring cyclization reactions with activation energies predicted to be 7.2, 5.0, 8.6, 6.4, and 6.2 kcal/mol, respectively. Methyl substitution on nitrogen decreases the barrier to cyclization. One methyl has a larger effect on the cyclization rate than the second methyl. This rate enhancement is attributed to a decrease in gauche interactions in the transition state as compared to the initial structure and to different destabilizing effects when an H is replaced by a methyl group. These predicted rate effects are in agreement with the experimental data on polymerization efficiencies. PMID- 12126391 TI - Mechanistic study of stereoselective gas-phase reactions of phosphenium ions with cis- and trans-1,2-diaminocyclohexanes. AB - The 1,3-dioxolane-2-phosphenium ion, 1,3-benzodioxole-2-phosphenium ion, and o biphenylenephosphenium ion are reported to react in a stereoselective manner with cis- and trans-1,2-diaminocyclohexanes in the gas phase in a Fourier transform ion cyclotron resonance mass spectrometer. Elimination of NH3 from an addition product was observed only for the trans isomer. Several reaction mechanisms were experimentally and computationally examined (B3LYP/6-31G(d)//HF/6-31G(d) + ZPVE level of theory). The most plausible mechanism is initiated by addition of one of the amino groups to the electrophilic phosphorus atom followed by proton transfer between the amino groups. A change to a diaxial conformation for the trans isomer facilitates anchimeric assistance by the now nucleophilic phosphorus atom as the C-N bond breaks to release NH3. Intramolecular proton transfer competes with the conformational change and ultimately leads to ethylene glycol elimination. The transition states for the critical steps of these two reactions are calculated to be nearly equal in magnitude, which rationalizes the observation of both reactions for the trans-diamine. In contrast, the adduct of the cis isomer can eliminate NH3 via a concerted 1,2-hydride shift without a need for a conformational change. However, the barrier associated with this reaction was found to be substantially greater than for proton transfer between the N- and O atoms. The latter reaction dominates and ultimately leads to ethylene glycol elimination. PMID- 12126392 TI - Ni-to-Ni+ 3-ethylene-bridged partially modified retro-inverso tetrapeptide beta turn mimetic: design, synthesis, and structural characterization. AB - A 10-membered heterocyclic ring system 1,3,8-trisubstituted 2,5,7-trioxo-1,4,8 triazadecane that represents a Ni-to-Ni+ 3-ethylene-bridged partially modified retro-inverso tetrapeptide beta-turn mimetic (EBRIT-BTM) has been designed, synthesized, and structurally analyzed. These compounds utilize an ethylene bridge to replace the COi...HNi + 3 10-membered hydrogen bond of standard beta turns. The N,N'-ethylene-bridged dimer was obtained in 90% yield by reductive alkylation of phenylalanylamide with a tert-butyl N-(9 fluorenylmethyloxycarbonyl),N-(2-formylmethyl)-glycinate. An orthogonal protection strategy and HATU-mediated couplings allowed efficient stepwise additions of monomeric building blocks leading to a N(i)-to-N(i+3)-ethylene bridged linear precursor: Further elaboration of the linear precursor generated the ethylene-bridged model compounds (16) and (18) (g, gem-diaminoalkyl; m, malonyl; and r, direction-reversed amino acid residue) in 44 and 39% yields, respectively. The structural features of the two EBRIT-BTM compounds were determined using 1H NMR and extensive computer simulations. The results indicate that the 10-membered rings are conformationally constrained with well-defined structural features and that the three amide bonds in the ring are in the trans orientation. The topological arrangement of the residues in the ring system closely resembles a type II' beta-turn. Transformation of CONH(2) in the N terminal amino acid residue of 16 into NHCOCH3 in 18 resulted in the formation of a hydrogen bond between the NH of gPhe-COCH3 and the C-terminal carboxyl of Gly, initiating an antiparallel beta-sheet. The formulation of the concept applying a minimalistic structural elaboration approach and the synthetic exploration, together with the conformational analysis, offer a new molecular scaffolding system and a true tetrapeptide secondary structure mimetic that can be used to generate peptidomimetics of biological interest. PMID- 12126393 TI - Computational studies of aliphatic amine basicity. AB - Computational studies have been used to examine the structural and energetic effects of adding small numbers of water molecules to ammonia, methylamine, dimethylamine, and trimethylamine, and their respective ammoniums ions using the effective fragment potential method. Distinct structural effects with only a few fragment water molecules are revealed. The complexity of structures increases with the number of water fragments with the water fragments forming complex networks. Structural and energetic effects are used to probe the so-called anomalous basicity effect of ammonia and the methylamines on going from the gas phase to aqueous solution. PMID- 12126394 TI - Steric hindrance as a mechanistic probe for olefin reactivity: variability of the hydrogenic canopy over the isomeric adamantylideneadamantane/sesquihomoadamantene pair (a combined experimental and theoretical study). AB - Access to each C=C face of adamantylideneadamantane (AA) and sesquihomoadamantene (SA) is hindered by the hydrogenic canopy consisting of four beta-hydrogens; otherwise, these olefins have quite normal environments. X-ray crystallography and density functional (DFT) calculations show a 0.5 A larger annular opening in the protective cover of AA than that in SA. This contributes to the remarkable differences in reactivity toward various reagents, not only by limiting access to the olefin site in SA but also by inhibiting reactions which force these hydrogens closer together. Thus, AA is subject to typical olefin-addition reactions with bromine, sulfuryl chloride, m-chloroperbenzoic acid, dioxygen, and so forth, albeit sometimes at attenuated rates. On the other hand, SA is singularly unreactive under identical reaction conditions, except for the notable exceptions that include Bronsted (protonic) acids, a nitrosonium cation, and dichlorine. The exceptions are characterized as three sterically limited (electrophilic) reagents whose unique reactivity patterns are shown to be strongly influenced by steric access to the C=C center. As such, the different degrees of steric encumbrance in the isomeric donors AA and SA shed considerable light on the diverse nature of olefinic reactions. In particular, they evoke mechanistic features in electrophilic addition versus electron transfer, which are otherwise not readily discernible with other less hindered olefinic donors. Transient structures of the olefinic-reaction intermediates such as the protonated carbocations AA-H+ and SA-H+ as well as the cation radicals AA*+ and SA*+ are probed by the combination of X-ray crystallographic analyses and density functional theoretical computations. PMID- 12126395 TI - Cyclobutenbriarein A, the first diterpene with a tricyclo[8.4.0.0(3,6)]tetradec-4 ene ring system isolated from the Gorgonian Briareum asbestinum. AB - Cyclobutenbriarein A (2), a novel class of C20-rearranged diterpene briarane possessing an unprecedented tricyclo[8.4.0.0(3,6)]tetradec-4-ene ring system was isolated, along with five new briarane diterpenoids (1 and 3-6), from the organic extracts of the Bahamian gorgonian Briareum asbestinum. The structures of these secondary metabolites were established on the basis of extensive NMR studies and accurate mass measurements (HRFABMS). PMID- 12126396 TI - Lewis acid-activated chiral leaving group: enantioselective electrophilic addition to prochiral olefins. AB - A new strategy using a BINOL derivative as a chiral leaving group and Lewis acid has been developed for enantioselective alkylation of prochiral olefins. (R)-2,2' Bis[2-(trimethylsilyl)ethoxymethyl]-1,1'-binaphthol is demonstrated to be an effective reagent for enantioselective hydroxymethylation of silyl enol ethers and trisubstituted alkenes. Electrophilic addition to prochiral olefins is accompanied by cleavage of an acetal that is dual activated by SnCl4 and the delta-effect of silicon through the S(N)2 substitution process. Enantioselective synthesis of cyclic terpenes is also described using this strategy. PMID- 12126397 TI - Synthesis of 6H-dibenzo[b,d]pyran-6-ones from aryl 3-bromopropenoates via a sequential one-pot procedure using the Sonogashira coupling-benzannulation reaction. AB - Various kinds of 6H-dibenzo[b,d]pyran-6-ones 4 were synthesized via a sequential one-pot procedure using the Sonogashira coupling-benzannulation reaction of aryl 3-bromopropenoates 1, in which the ortho-position of aryl group is substituted with enynes or iodine, with acetylenes 2 in the presence of palladium and copper catalysts. The Sonogashira coupling between the aryl 3-bromopropenoates 1a and 1b, bearing enynes at the ortho-position of aryl group, and alkynes 2a-g gave the enyne intermediates 3 in situ, which subsequently underwent the palladium catalyzed benzannulation reaction to afford the 6H-dibenzo[b,d]pyran-6-ones 5a-g and 6. The Sonogashira coupling between the aryl 3-bromopropenoate 1c, bearing iodine at the ortho-position of aryl group, and diynes 2f and 2h produced the diyne intermediates 13, which underwent the benzannulation reaction to afford the 6H-dibenzo[b,d]pyran-6-ones 14f and 14h. PMID- 12126399 TI - Mild and highly selective formyl protection of primary hydroxyl groups. AB - Efficient conversion of primary alcohols to the corresponding formate esters can be carried out at room temperature in methylene chloride, using 2,4,6-trichloro 1,3,5-triazine and N,N-dimethylformamide in the presence of lithium fluoride. This procedure appears as a valid method for selectively protecting primary hydroxyl groups. PMID- 12126398 TI - Synthesis of [5'-13C]ribonucleosides and 2'-deoxy[5'-13C]ribonucleosides. AB - The present efficient synthesis of [5'-13C]ribonucleosides and 2'-deoxy[5' 13C]ribonucleosides is characterized by the synthesis of the D-[5-13C]ribose derivative as an intermediate via the Wittig reaction of 4-aldehydo-D-erythrose dialkyl acetals with Ph3P13CH3I-BuLi to introduce the 13C label at the 5-position of a pentose. This was followed by the highly diastereoselective osmium dihydroxylation for the preparation of 2,3-di-O-benzyl-D-[5-13C]ribose dialkyl acetal and the cyclization from D-[5-13C]ribose dialkyl acetal derivatives to the alkyl D-[5-13C]ribofuranoside derivative by the use of LiBF(4). The obtained D-[5 13C]ribose derivative was converted into [5'-13C]ribonucleosides and subsequently into the corresponding 2'-deoxynucleosides. PMID- 12126400 TI - Chiral nonracemic alpha-alkylidene and alpha-silylidene cyclopentenones from chiral allenes using an intramolecular allenic Pauson-Khand-type cycloaddition. AB - We have successfully effected a transfer of chirality from a chiral nonracemic allene to an alpha-alkylidene and an alpha-silylidene cyclopentenone. The molybdenum-mediated examples possessing a silyl group on the terminus of the allene show good facial selectivities, but isomerization of the (E)-silylidene cyclopentenone to the (Z)-silylidene cyclopentenone occurs upon purification of these products. Alternatively, an alkyl group on the terminus of the allene undergoes cycloaddition with moderate selectivities but gives products that undergo an isomerization of the (Z)-alkylidene cyclopentenone to the (E) alkylidene cyclopentenone when exposed to acidic conditions. Thus, erosion of the enantiomeric excesses is observed for one of the two products as a result of this isomerization. The allenic Pauson-Khand-type cycloaddition has also been effected by first isolation the (eta(6)-arene)molybdenum tricarbonyl complex, demonstrating for the first time that this is most likely the active complex in the molybdenum-mediated reactions. Attempts to increase the facial selectivity by increasing the size of the arene moiety resulted in a marginal increase in the selectivity at the expense of the yield. Based upon these results, we have concluded that altering the approach for the preparation of nonracemic alpha alkylidene cyclopentenones is necessary in order to obtain synthetically useful levels of stereocontrol. PMID- 12126401 TI - New routes to N-alkylated cyclic sulfamidates. AB - BOC- and dibenzosuberyl-protected chiral and hindered cyclic sulfamidates ([1,2,3]-oxathiazolidine-2,2-dioxides) were synthesized and subsequently deprotected using trifluoroacetic acid. The resulting crystalline sulfamidates were then used in several alkylation reactions involving benzyl bromide and alcohols in a versatile route to cyclic sulfamidates with differing N-alkyl substituents. PMID- 12126402 TI - Ring-opening reactions of cyclic acetals and 1,3-oxazolidines with halosilane equivalents. AB - Reactions of acetal and 1,3-oxazolidine rings were examined using two kinds of iodosilane equivalent reagents, a 1:2 mixture of Me3SiNEt2 and MeI (reagent 1a) and a 1:1 mixture of Et3SiH and MeI containing a catalytic amount of PdCl2 (reagent 1b). In the reactions of alkanone ethylene acetals with reagent 1a, a C O bond in the acetal ring readily cleaved to give 2-(trimethylsiloxy)ethyl enol ethers. Similarly, the C-O bond of 1,3-oxazolidine rings cleaved to give ring opened imine or enamine derivatives. The reactions of aromatic ketone ethylene acetals and cyclohexanone trimethylene acetal led to deprotection of the acetal unit to liberate free ketones. With reagent 1b, cycloalkanone ethylene acetal afforded a dimeric product with 2-iodoethyl alkenoate moieties, while aromatic ketone ethylene or trimethylene acetals produced deprotected ketones. PMID- 12126403 TI - Approach toward the total synthesis of orevactaene. 2. Convergent and stereoselective synthesis of the C18-C31 domain of orevactaene. Evidence for the relative configuration of the side chain. AB - The synthesis of the C18-C31 subunit of orevactaene (1) in an enantioselective and convergent manner is reported. Four chiral centers in the structure (i.e., carbons 23, 25, 32, and 33) have unknown configuration; thus, a modular approach has been devised to link the two stereocenter-containing ends of the structure together via a trisubstituted olefin template to ultimately produce all possible diastereomers of the target. Keys to the success of this approach include (i) an efficient synthesis of four diastereomeric hydrophobic tails (C22-C29) of the molecule with two stereogenic centers at C23 and C25; (ii) the synthesis of three stereodefined trisubstituted olefins 37, 38, and 43 using palladium(0)-catalyzed hydrometalation and metallometalation; and (iii) the convergent assembly of the aforementioned sections by a 'one-pot' lithium/halogen exchange, boron/lithium exchange, borate ester saponification, and Suzuki cross-coupling followed by oxidative deprotection. The sequence provided the desired aldehydes 49 and 50 as single isomers in good yields. Compiled spectroscopic data from the literature and present work provides evidence that the relative configuration of the methyl groups in the side chain of orevactaene may be 1,3-syn, which will be confirmed when the total synthesis has been completed. These results have paved the way for a parallel synthesis approach to prepare all 16 possible stereoisomers of orevactaene so that the relative and absolute stereochemistry of this compound can be determined. PMID- 12126404 TI - Chiral N-alkyl-2,4,6-triphenylpyridiniums as enantioselective triplet photosensitizers. laser flash photolysis and preparative studies. AB - Three N-alkylpyridinium photosensitizers having chiral alkyl groups have been prepared by reacting 2,4,6-triphenylpyrylium tetrafluoroborate ion with (1R,2S)-( )-norephedrine, (S)-(+)-2-(aminomethyl)pyrrolidine, and (R)-(-)-1 cyclohexylethylamine. Laser flash photolysis allows detection of the corresponding triplet excited states that are quenched by hydrogen atom donors and electron donors. Asymmetric quenching of the chiral triplet excited state was observed using enantiomerically pure 1,2-diamino cyclohexane as quencher. Low enantiomeric excess values (up to 7%) were measured for the photochemical cyclization of 5-methyl-4-hexenoic acid to its corresponding gamma-lactone using these chiral N-alkylpyridinium as photosensitizers. PMID- 12126406 TI - Reaction of allylsilanes and allylstannanes with alkynes catalyzed by electrophilic late transition metal chlorides. AB - The intramolecular reaction of allylsilanes and allylstannanes with alkynes proceeds catalytically in the presence of Pt(II), Pd(II), Ru(II), and Au(III) chlorides. Although more limited, AgOTf also catalyzes the cyclization. Usually, PtCl2 as the catalyst in methanol or acetone gives the best results. The reaction proceeds by exo attack of the allyl nucleophile on the alkyne to form five- or six-membered ring carbocycles. The reaction generally proceeds with anti stereoselectivity. However, a terminally substituted trimethylsilyl derivative reacts by a syn-type addition. The intermediate alkenylpalladium complex has been trapped with allyl chloride to form an allylated derivative with an additional carbon-carbon bond. PMID- 12126405 TI - Naphthalene diols: a new class of antioxidants intramolecular hydrogen bonding in catechols, naphthalene diols, and their aryloxyl radicals. AB - 1,8-Naphthalenediol, 5, and its 4-methoxy derivative, 6, were found to be potent H-atom transfer (HAT) compounds on the basis of their rate constants for H-atom transfer to the 2,2-di(4-t-octylphenyl)-1-picrylhydrazyl radical (DOPPH*), k(ArOH/DOPPH)*, or as antioxidants during inhibited styrene autoxidation, k(ArOH/ROO)*, initiated with AIBN. The rate constants showed that 5 and 6 are more active HAT compounds than the ortho-diols, catechol, 1, 2,3-naphthalenediol, 2, and 3,5-di-tert-butylcatechol, 3. Compound 6 has almost twice the antioxidant activity, k(ArOH/ROO)* = 6.0 x 10(6) M(-)(1) s(-1), of that of the vitamin E model compound, 2,2,5,7,8-pentamethyl-6-chromanol, 4. Calculations of the O-H bond dissociation enthalpies compared to those of phenols, (deltaBDEs), of 1-6 predict a HAT order of reactivity of 2 < 1 < 3 approximately 4 < 5 < 6 in general agreement with kinetic results. Calculations on the diols show that intramolecular H-bonding stabilizes the radicals formed on H-atom transfer more than it does the parent diols, and this effect contributes to the increased HAT activity of 5 and 6 compared to the activities of the catechols. For example, the increased stabilization due to the intramolecular H-bond of 5 radical over 5 parent of 8.6 kcal/mol was about double that of 2 radical over 2 parent of 4.6 kcal/mol. Linear free energy plots of log k(ArOH/DOPPH)* and log k(ArOH/ROO)* versus deltaBDEs for compounds 1-6 along with available literature values for nonsterically hindered monophenols placed the compounds on common scales. The derived Evans-Polanyi constants from the plots for the two reactions, alpha(DOPPH)* = 0.48 > alpha(ROO)* = 0.32, gave the expected order, since the ROO* reaction is more exothermic than the DOPPH* reaction. Compound 6 is sufficiently reactive to react directly with oxygen, and it lies off the log k(ArOH/ROO)* versus deltaBDE plot. PMID- 12126407 TI - A simple and versatile method for the synthesis of acetals from aldehydes and ketones using bismuth triflate. AB - Acetals are obtained in good yields by treatment of aldehydes and ketones with trialkyl orthoformate and the corresponding alcohol in the presence of 0.1 mol % Bi(OTf)3.4H2O. A simple procedure for the formation of acetals of diaryl ketones has also been developed. The conversion of carbonyl compounds to the corresponding 1,3-dioxolane using ethylene glycol is also catalyzed by Bi(OTf)3.4H2O (1 mol %). Two methods, both of which avoid the use of benzene, have been developed. PMID- 12126408 TI - Synthesis of polysubstituted benzothiophenes and sulfur-containing polycyclic aromatic compounds via samarium diiodide promoted three-component coupling reactions of thiophene-2-carboxylate. AB - By the promotion of samarium diiodide, thiophene-2-carboxylate reacted with 2 equiv of ketones at the C-4 and C-5 positions to give diols such as 2 and 9. Because the intermediary organosamarium species were oxophilic but not too basic, the double hydroxyalkylations with various ketone substrates, including alkyl aryl ketones, acetylthiophenes, cyclohexanone, alpha-tetralone, and alpha phenylacetophenones, were realized without complication of side reactions. The diol products underwent an acid-catalyzed dehydration to give dienes such as 3 and 10, which were treated with DDQ to give either polysubstituted thiophenes (e.g., 4 and 11) or benzothiophenes (e.g., 5, 13, and 14) depending on the reaction conditions. Oxidative annulations of 4,5-diarylthiophenes 11 and 4,5,6,7 tetraphenylbenzothiophenes 14 were carried out by photochemical or chemical methods to give the sulfur-containing polycyclic aromatic compounds, such as phenanthro[9,10-b]thiophene-2-carboxylate, piceno[13,14-b]thiophene-2 carboxylate, and tribenzo[fg,ij,rst]pentapheno[15,16-b]thiophene-2-carboxylates. This method is applicable to the preparation of polysubstituted thiophenes, benzothiophenes, and the related compounds possessing liquid crystalline, photochromic, and other functional properties. PMID- 12126409 TI - Synthetic applications of o- and p-halobenzyl sulfones as zwitterionic synthons: preparation of ortho-substituted cinnamates and biarylacetic acids. AB - The synthetic applications of o-halobenzyl and p-halobenzyl sulfones as precursors of 1,3- and 1,5-zwitterionic synthons, respectively, are described. Their alpha-sulfonyl carbanions, generated by means of the phosphazene base P2-Et or BuLi or K2CO3 under PTC conditions, reacted with different electrophiles such as alkyl halides, aldehydes, and electrophilic olefins. Palladium-catalyzed cross coupling processes such as Heck, Suzuki, and Sonogashira reactions can be efficiently performed at the halogen atom. These two sequential functionalization processes are applied to the synthesis of ortho-substituted cinnamates and pharmaceuticals belonging to the family of p-biarylacetic acids such as 4 biphenylacetic acid, namoxyrate, xenyhexenic acid, and biphenylpropionic acid. PMID- 12126410 TI - A one-step procedure for the monoacylation of symmetrical 1,2-diols. AB - A series of lanthanide (III) salts have been shown to catalyze the monoacylation of symmetrical 1,2-diols by carboxylic acid anhydrides with surprisingly high selectivity. PMID- 12126411 TI - Ketene-forming eliminations from aryl phenylacetates promoted by R2NH/R2NH2+ in aqueous MeCN. Mechanistic borderline between E2 and E1cb. AB - Elimination reactions of 2-X-4-NO2C6H3CH2C(O)OC6H3-2-Y-4-NO2 [X = H (1), NO2 (2)] promoted by R2NH/R2NH2+ in 70 mol % MeCN(aq) have been studied kinetically. The base-promoted eliminations from 1 proceeded by the E2 mechanism when Y = Cl, CF3, and NO2. The mechanism changed to the competing E2 and E1cb mechanisms by the poorer leaving groups (Y = H, OMe) and to the E1cb extreme by the strongly electron-withdrawing beta-aryl group (2, X = NO2). The values of beta = 0.14 and beta(lg) = 0.10-0.21 calculated for elimination from 1 (Y = NO2) indicate a reactant-like transition state with small extents of proton transfer and C(alpha) OAr bond cleavage. The extent of proton transfer increased with a poorer leaving group, and the degree of leaving group bond cleavage increased with a weaker base. Also, the changes in the k(1) and k(-1)/k(2) values with the reactant structure variation are consistent with the E1cb mechanism. From these results, a plausible pathway of the change of the mechanism from E2 to the E1cb extreme is proposed. PMID- 12126412 TI - Exploring stereogenic phosphorus: synthetic strategies for diphosphines containing bulky, highly symmetric substituents. AB - Diphosphine ligands bearing highly symmetric, bulky substituents at a stereogenic P atom were prepared, exploiting established protocols, which include the use of chiral synthons such as 3,4-dimethyl-2,5-diphenyl-1,3,2-oxazaphospholidine-2 borane (3a) and phenylmethylchlorophosphine borane (10) and the enantioselective deprotonation of dimethylarylphosphine boranes. However, only (Bu(t)())(Me)PCH(2)CH(2)P(Bu(t)Me (8a) could be prepared from 3a. The diphosphines (S,S)-1,2-bis(mesitylmethylphosphino)ethane, ((S,S)-8b) and (S,S) 1,2-bis(9-anthrylmethylphosphino)ethane ((S,S)-8c), which contain 2,6 disubstituted aryl P-substituents, were prepared by Evans' sparteine-assisted enantioselective deprotonation of P(Ar)(Me)(2)(BH(3)) (Ar = mesityl or 9 anthryl), but the enantioselectivity did not exceed 37% ee. The asymmetrically substituted, methylene-bridged diphosphine (2R,4R)-(Ph)(CH(3))PCH(2)P(Mes)(CH(3)) ((2R,4R)-12) (Mes = mesityl) was prepared by the newly developed stereospecific reaction of the enantiomerically pure chlorophosphine borane PCl(Ph)(Me)(BH(3)) (10) with the racemic, monolithiated dimethylmesitylphosphine borane P(Mes)(Me)(CH(2)Li)(BH(3)). Diastereomerically pure (2R,4R)-12 was obtained with 86% ee. The rhodium(I) derivatives [Rh(COD)(P-P)]BF(4) containing the diphosphine ligands 8a, 8b, and 12, as well as the previously reported (S,S)-1,2-bis(1 naphthylphenylphosphino)ethane ((S,S)-8d), were prepared and tested in the enantioselective catalytic hydrogenation of acetamidocinnamates. The best catalytic result (98.6% ee) was obtained with [Rh(COD)(8d)](+) as catalyst and methyl Z-alpha-acetamidocinnamate as substrate. Some of the catalytic results are discussed in terms of the preferred conformations of the substituents at phosphorus, as calculated by molecular modeling. PMID- 12126413 TI - Boron-mediated aldol reaction of carboxylic esters: complementary anti- and syn selective asymmetric aldol reactions. AB - The boron-mediated aldol reaction of carboxylic esters is described in detail. Contrary to the general belief that carboxylic esters are inert under the condition of the boron enolate formation, propionate esters are enolized with certain combinations of a boron triflate and an amine. More importantly, the stereochemical course of the aldol reaction can be controlled by the judicious selection of the enolization reagents. Treatment of propionate esters with c Hex2BOTf and triethylamine produces anti-aldol products, and that with Bu2BOTf and diisopropylethylamine gives syn-aldol products selectively after reaction with aldehydes. Complementary anti- and syn-selective asymmetric aldol reactions with structurally related, readily available chiral norephedrine-derived propionate esters are developed. PMID- 12126414 TI - Iron-assisted nucleophilic aromatic substitution on solid phase. AB - Iron-assisted S(N)Ar reactions were performed for the first time on solid phase, and a library of 36 unsymmetrically substituted phenylpiperazines and phenyl-1,4 diazepanes was synthesized with this novel strategy. The scope of iron-assisted S(N)Ar reactions on solid phase was investigated, and reactions of representative nucleophiles from groups VI (O, S, and Se) and V (N and P) of the periodic table were examined. Decomplexation of resin-bound iron complexes was achieved with 1,10-phenanthroline under irradiation, thereby overcoming the notorious disadvantages of decomplexation observed in solution-phase chemistry. PMID- 12126415 TI - Total synthesis of (+/-)-momilactone A. AB - The first total synthesis of (+/-)-momilactone A was accomplished using a highly diastereoselective transannular Diels-Alder reaction on a trans-trans-cis macrocyclic triene. PMID- 12126416 TI - An efficient convergent synthesis of novel anisotropic adsorbates based on nanometer-sized and tripod-shaped oligophenylenes end-capped with triallylsilyl groups. AB - This article describes an efficient and convergent synthesis of a novel tripod shaped oligophenylene compound. The compound consists of three oligophenylene heptamers as the tripod legs and one bromophenyl group as the functional arm, joining at a tetrahedron silicon atom. Each tripod leg is end-capped with a triallylsilyl group for covalently anchoring the molecules on hydrogen-terminated silicon surfaces via hydrosilylation. The compound may represent a new type of anisotropic adsorbates for controlling the orientation of and spacing between functional groups in organic thin films and nanostructures grown on silicon surfaces. An oligophenylene hexamer end-capped with a triallylsilyl group and a pinacol arylboronate group was readily derived from diiodoterphenyl with a bromophenyl boronic acid by selective Suzuki coupling. Reaction conditions for highly selective Suzuki coupling of sterically hindered pinacol boronate with aryl iodide in the presence of multiple ethenyl groups for the competing Heck reaction were developed. Under the optimal conditions (Pd(PPh3)4/KOH/Bu4NBr/toluene/H2O, 85 degrees C, overnight), p-bromophenyl-tris(p iodo-phenyl)silane reacted with 3 equiv of the hexaphenylene boronate to provide the tripod-shaped oligophenylene, composed of 22 phenylene and 9 allyl groups, in 78% yield. PMID- 12126417 TI - Origins of regioselectivity in the reactions of alpha-lactams with nucleophiles. Two distinct acid-catalyzed pathways involving O- and N-protonation. AB - Sterically stabilized alpha-lactams react by two distinct acid-catalyzed pathways. Protonation on oxygen results in nucleophilic attack at the acyl carbon and gives C-2 products. Protonation on nitrogen leads to nucleophilic attack at the C-3 carbon and yields C-3 products. The mechanism thus developed is very useful for understanding the changes in rates and product distributions in the reactions of sterically stabilized alpha-lactams with nucleophiles. It can also be extrapolated to other alpha-lactams so that a more coherent picture of alpha lactam reactivity can be developed. PMID- 12126418 TI - Tributylphosphine: a remarkable promoting reagent for the ring-opening reaction of aziridines. AB - Tributylphosphine was found to be an effective promoting reagent for ring opening of a variety of aziridines and nucleophiles to produce anti-bifunctional products in good to excellent yield. The study showed that the reaction is initiated through the attack of tributylphosphine as a nucleophile at the carbon atom of the aziridine ring. PMID- 12126419 TI - Multiple dendritic catalysts for asymmetric transfer hydrogenation. AB - The first and second generation multiple dendritic ligands based on chiral diamine were synthesized in a convergent approach and were well-characterized by NMR and MS techniques. Their ruthenium complexes prepared in situ had good solubility in the reaction medium (azeotrope of formic acid and triethylamine) and demonstrated high catalytic activity and enantioselectivity comparable to monomeric catalysts in the asymmetric transfer hydrogenation of ketones and imines. Quantitative yields and for some cases a slightly higher enantioselectivity (up to 98.7% ee) were obtained in the dendritic catalysis. Considering the high local catalyst concentrations at the periphery, diones were tested for the possible synergic reactivity between catalytic units at the surface, while no apparent differences were noted. PMID- 12126420 TI - 1,2-dioxines as masked cis gamma-hydroxy enones and their versatility in the synthesis of highly substituted gamma-lactones. AB - Addition of highly stabilized ester nucleophiles to 1,2-dioxines affords good to high yields of gamma-lactones with high diastereoselectivity. Heterolytic or homolytic cleavage of the 1,2-dioxines under appropriate conditions generates the key reactive cis gamma-hydroxy enones, which ultimately afford the observed gamma lactones. Diastereoselectivity is installed as a result of anti 1,4-addition by the ester enolate to the cis enones followed by intramolecular cyclization. The reaction is tolerant of a range of substitution patterns on the 1,2-dioxine while a broad range of esters are also accommodated. In addition to the synthesis of racemic gamma-lactones, highly enantioenriched gamma-lactones can also be synthesized when chiral cobalt(II) catalysts are employed for the initial homolytic ring-opening of the 1,2-dioxine. PMID- 12126421 TI - Selective reductions. 59. Effective intramolecular asymmetric reductions of alpha , beta-, and gamma-keto acids with diisopinocampheylborane and intermolecular asymmetric reductions of the corresponding esters with B chlorodiisopinocampheylborane. AB - A comparison of the stereochemistry of the products obtained from the intramolecular asymmetric reduction of a series of keto acids with (-) diisopinocampheylborane and intermolecular asymmetric reduction of the corresponding series of keto esters with (-)-B-chlorodiisopinocampheylborane ((-) DIP-Chloride) has been made. The stereochemistry of the hydroxy acids from the reduction of keto acids is dependent only on the enantiomer of the reagent used. The stereochemistry of the products from the reduction of keto esters is also consistent, except those of aliphatic alpha-keto esters. alpha-, beta-, and gamma keto acids provide the corresponding hydroxy acids in 77-98% ee, and the alpha- and gamma- keto esters afford the hydroxy esters in 82->or=99% ee. beta-Keto esters do not undergo reduction. Although the reduction of delta-keto acids does not proceed under the same reaction conditions, the reduction of delta-keto esters is facile. All of the products from the reduction of gamma-keto acids and esters and delta-keto esters were converted to the corresponding lactones. This study revealed that DIP-Chloride is an efficient reagent for the reduction of alpha-keto esters at low temperatures. PMID- 12126422 TI - Unexpected catalyst for Wittig-type and dehalogenation reactions. AB - A novel catalyst 2 for Wittig-type and dehalogenation reactions was developed. In the presence of triphenyl phosphite, a wide variety of aldehydes could react with alpha-bromoacetates to afford alpha,beta-unsaturated esters or ketones in high yields with excellent E-stereoselectivity when 1-2 mol % of compound 2 was used. Compound 2 was also an effective catalyst for reductive dehalogenation of alpha bromocarbonyl compounds. The mechanisms for the above reactions were also proposed. PMID- 12126423 TI - Synthesis of low-generation, aryl-/alkyl-type, nonpolar dendrons carrying protected hydroxyalkyl groups in the periphery. AB - An efficient convergent synthesis of first- and second-generation aryl-/alkyl type nonpolar dendrons via Suzuki cross-coupling is described. The dendrons carry either one or two benzyl-protected hydroxyalkyl groups/terminus. Iododesilylation reactions of aryltrimethylsilanes with iodo chloride are used as a tool for the incorporation of iodo, an important functionality for transition-metal-catalyzed cross-coupling reactions. In the case of sensitive aromatics, the addition of some donor solvent like diethyl ether proved effective in suppressing side reactions through electrophilic aromatic iodination. PMID- 12126424 TI - Synthesis of [n]- and [n.n]cyclophanes by using Suzuki-Miyaura coupling. AB - Reaction of the bis-9-BBN adduct of several dienes with 1,3-dibromobenzene via Suzuki coupling leads to a series of [n]metacyclophanes ranging in size from 10 to 17 atom members. In each case, two carbon-carbon bonds are formed in one reaction vessel. However, when the bis-9-BBN adduct of 1,5-hexadiene is coupled with a variety of aryl dihalides, larger [n.n]cyclophanes were formed in preference to the [n]cyclophanes. Four carbon-carbon bonds are formed in this instance. Single-crystal X-ray analyses of these [n.n]cyclophanes reveal interestingly shaped molecules with large cavities. PMID- 12126425 TI - Variable strategy toward carbasugars and relatives. 4. Viable access to (4a carbapentofuranosyl)amines, (5a-carbahexopyranosyl)amines, and amino acids thereof. AB - A chiral, divergent synthesis of two carbafuranosylamines, 1 and 2, two carbapyranosylamines, 3 and 4, two carbafuranosylamino acids, 5 and 6, and two carbapyranosylamino acids, 7 and 8, has been achieved. Highlights of the procedure include the following: a diastereoselective crossed vinylogous Mukaiyama aldol coupling between N-(tert-butoxycarbonyl)-2-[(tert butyldimethylsilyl)oxy]pyrrole (TBSOP, 9) and 2,3-O-isopropylidene-D glyceraldehyde (10) for the assembly of the target compound carbon backbone; a high-yielding silylative cycloaldolization that gives the cyclopentanoid and cyclohexanoid motifs; and a reductive or hydrolytic breakage of the lactam C(O)-N link to liberate the carbasugar and install the desired pseudo-anomeric amine and the hydroxymethyl or carboxyl functionalities. The sequences leading to trans configured carbafuranosyl compounds 1 and 5 and carbapyranosyl compounds 3 and 7 were 12- and 13-step processes, with overall yields of 34%, 35%, 17%, and 16%. Cis-configured isomers 2, 4, 6, and 8 were obtained only in minor yields. PMID- 12126426 TI - Dynamic kinetic resolution of racemic gamma-aryl-delta-oxoesters. Enantioselective synthesis of 3-arylpiperidines. AB - Cyclodehydration of racemic gamma-aryl-delta-oxoesters with (R)- or (S) phenylglycinol stereoselectively affords bicyclic delta-lactams, in a process that involves a dynamic kinetic resolution. Subsequent reduction of these lactams leads to enantiopure 3-arylpiperidines. Starting from racemic aldehyde esters, this short sequence has been applied to the synthesis of (R)-3-phenylpiperidine and the antipsychotic drug (-)-3-PPP (an (S)-3-arylpiperidine), whereas starting from racemic ketone esters enantiopure cis-2-alkyl-3-arylpiperidines are prepared. PMID- 12126427 TI - Synthesis of the tRNA(Lys,3) anticodon stem-loop domain containing the hypermodified ms2t6A nucleoside. AB - The synthesis of a protected form of the hypermodified nucleoside, N-[(9-beta-D ribofuranosyl-2-methylthiopurin-6-yl)carbamoyl]threonine, (ms2t6A) is reported. The hypermodified nucleoside was subsequently elaborated to the protected nucleoside phosphophoramidite using a protecting group strategy compatible with standard RNA oligonucleotide chemistry. The phosphoramidite reagent was then used to synthesize the 17-nucleotide RNA hairpin having the sequence of the anticodon stem-loop (ASL) domain of human tRNA(Lys,3), the primer for HIV-1 reverse transcriptase. Introduction of the modification at position 37 of the tRNA ASL modestly decreases the thermodynamic stability of the RNA hairpin as has been seen previously for the prokaryotic t6A nucleoside lacking the 2-methylthio substituent. 2D NOESY NMR spectra of the ms2t6A containing tRNA ASL indicate that the threonyl side chain adopts a conformation similar to that seen in the solution structure of the analogous t6A containing E. coli tRNA(Lys), despite the presence of the bulky methylthio group. This synthetic approach allows for site specific incorporation of the hypermodified nucleoside and should facilitate future studies directed at understanding the roles of nucleoside modification in modulating the stability and specificity of biologically important RNA-RNA interactions. Our synthesis of the ms2t6A containing RNAs demonstrates that this methodology is suitable for obtaining quantities of RNA required for structural studies of the HIV primer tRNA. PMID- 12126428 TI - Stereoselective synthesis of both syn- and anti-N-tert-alkylamines using highly stereospecific crotylation of ketone-derived acylhydrazones with crotyltrichlorosilanes. AB - Crotyltrichlorosilanes reacted with ketone-derived N-benzoylhydrazones in DMF without any catalyst. This is the first example of highly stereospecific crotylation of ketimine analogues leading to both syn- and anti-N'-tert-alkyl-N benzoylhydrazines. Different reactivities between (Z)- and (E)-crotylsilanes in terms of yields and selectivities were observed. A kinetic study with both geometrically pure (Z)- and (E)-crotylsilanes was performed. These reactions are most likely to proceed via a cyclic chairlike transition state where the aromatic group of hydrazones takes an axial position. Both diastereomers of allylation products can be converted to the corresponding alpha,alpha-disubstituted homoallylic amines without epimerization. PMID- 12126429 TI - Dipole-stabilized carbanions: a computational study of N-methylformamide anion and methyl N-methylcarbamate anion. AB - The relative energies of the rotamers of the carbanions derived from N methylformamide and methyl N-methylcarbamate have been studied at the MP2/6-311+G and B3LYP/6-311+G theoretical levels. There results are in good agreement but differ substantially from previously reported HF/6-31G calculations for the N methylformamide anions. Transition states for rotation and inversion of the anions were located. The methoxy group in methyl N-methylcarbamate has a large effect on the relative energies of the anions. The results are compared with those previously reported for the N-(methoxycarbonyl)piperidine anions, and it is found that the decreased conformational flexibility has an effect on the relative energies of these ions. PMID- 12126430 TI - Halo-enediynes: probing the electronic and stereoelectronic contributions to the Bergman cycloaromatization. AB - A series of halogen-substituted cyclic enediynes were prepared with use of carbenoid coupling strategy. DFT analysis, initially used to identify synthesis candidates, was also employed to rationalize the propensity for cycloaromatization of the compounds. In all cases studied the halogen atom had a strongly retardative effect on the thermal Bergman cycloaromatization reaction. The isolation of the first C-9 monochloroenediyne is noteworthy, and may find application in prodrug design. PMID- 12126431 TI - Novel synthesis of cis-nickel(II) 3-alkylimino-3-alkyl(or aryl)thio-1 arylpropenethiolates and their application to the preparation of 5-aryl-3 (arylthio)isothiazoles. AB - Treatment of (E)-3-alkylamino-3-alkylthio-1-(thioaroyl)propenes 2 with Ni(OAc)2.4H2O in EtOH at room temperature gave cis-Ni(II)-3-alkylimino-3 alkylthio-1-arylpropenethiolates 3 in excellent yields. Heating a mixture of 3 and alkyl- or arylthiols in 1,2-dichloroethane gave new ketene S,N-acetals 4 having new alkyl- or arylthio groups depending on the thiols employed. For the first time, 5-aryl-3-(arylthio)isothiazoles were prepared from 2 with an arylthio group according to a known procedure. PMID- 12126432 TI - A biomimetic transformation of serratinine into serratezomine A through a modified Polonovski reaction. AB - Application of a modified Polonovski reaction for serratinine (1) resulted in generation of serratezomine A (2) with a novel seco-serratinine-type skeleton recently isolated from the club moss Lycopodium serratum var. serratum. This biomimetic transformation supports a biogenetic pathway proposed for serratezomine A (2). The absolute stereochemistry of serratezomine A (2) was established by an induced exciton chirality and modified Mosher methods. PMID- 12126433 TI - Study of fluorocarbonyls for the Baylis-Hillman reaction. AB - A study of the effect of fluorine substitution in Baylis-Hillman reactions of various fluorocarbonyl partners with acrolein, methyl vinyl ketone, ethyl acrylate, and acrylonitrile has been made. PMID- 12126434 TI - InBr3-catalyzed Friedel-Crafts addition of indoles to chiral aromatic epoxides: a facile route to enantiopure indolyl derivatives. AB - Aromatic optically active epoxides can be opened in a regioselective and clean way with indoles in the presence of catalytic amount of InBr3 (1 mol %). The reaction takes place with a SN2 pathway affording the 2-aryl-2-(3'-indolyl)ethan 1-ols with excellent enantioselectivity (ee up to 99%). PMID- 12126435 TI - Preparation of 7-alkylamino-2-methylquinoline-5,8-diones. AB - Several novel 7-alkylamino-2-methylquinoline-5,8-diones (2) were synthesized from 2,5-dimethoxyaniline in five steps via the Skraup reaction followed by demethylations, oxidative bromination, amination, and debromination. We have achieved an unusual hydrobromic acid catalyzed debromination reactions of several 6-bromo-7-alkylamino-2-methylquinoline-5,8-diones, giving 7-alkylamino-2 methylquinoline-5,8-diones in good yields. PMID- 12126436 TI - An improved protocol for the preparation of 3-pyridyl- and some arylboronic acids. AB - 3-Pyridylboronic acid was prepared in high yield and bulk quantity from 3 bromopyridine via a protocol of lithium-halogen exchange and "in situ quench". This technique was further studied and evaluated on other aryl halides in the preparation of arylboronic acids. PMID- 12126437 TI - [4 + 2] cycloaromatization of 4-bis(methylthio)-3-buten-2-one with active methylene ketones: a simple and facile phenol annulation. AB - A new method for beta-phenol annulation involving base-induced [4C + 2C] cycloaromatization of readily available 4-bis(methylthio)-3-buten-2-one with a variety of cyclic and acyclic active methylene ketones has been reported. Appropriate choice of ketones allows synthesis of diverse frameworks such as dihydroindan, tetrahydronaphthalenes, their higher homologues, dihydro/octahydrophenanthrenes, anthracene, and heteroannulated analogue in high yields with regiocontrol of phenolic functionality. PMID- 12126438 TI - Kottamides A-D: novel bioactive imidazolone-containing alkaloids from the New Zealand ascidian Pycnoclavella kottae. AB - Kottamides A-D (1-4), novel 2,2,5-trisubstituted imidazolone-containing alkaloids, were isolated from the New Zealand endemic ascidian Pycnoclavella kottae and structurally characterized using 15N natural abundance 2-D NMR in addition to standard spectroscopic methods. The kottamides exhibited anti inflammatory and anti-metabolic activity as well as cytotoxicity toward tumor cell lines. PMID- 12126440 TI - Toward synthesis of alpha-alkyl amino glycines (A3G), new amino acid surrogates. AB - A general method giving access to protected alpha-alkyl amino glycines (A3G) 4 from the previously described precursor alpha-isopropylthioglycine 1 is described. In the presence of N-bromosuccinimide, displacement of the thiol by a large variety of amines afforded the corresponding racemic amino acid mimics. The efficiency of the reaction was strongly dependent on the protective groups of the nucleophile used in the condensation. PMID- 12126439 TI - Total synthesis of two naturally occurring bicyclo[3.2.1]octanoid neolignans. AB - The first total syntheses of the racemates of naturally occurring macrophyllin type bicyclo[3.2.1]octanoid neolignans, kadsurenin C 3 and kadsurenin L 4, were accomplished starting from vanillin and resorcinol. The acid-catalyzed rearrangement of hydrobenzofuranoid neolignans into bicyclo[3.2.1]octanoid neolignans was used as the key step. PMID- 12126441 TI - Synthesis of novel heteroaromatics structurally related to ellipticine alkaloids via thermolysis of pyridannulated enyne-carbodiimides. AB - New synthetic pathways to the 5H-pyrido[3',4':4,5]pyrrolo[2,3-b]quinolines 6, the 6H-pyrido[3',4':4,5]pyrrolo[2,3-b][1,6]naphthyridines 7, and the 11H pyrido[4',3':4,5]pyrrolo[2,3-b]quinolines 8 via thermolysis of the pyridannulated enyne-carbodiimides 14, 19, and 23 were established. These novel heteroaromatic systems are structurally related to ellipticine alkaloids and could serve as DNA intercalating agents. PMID- 12126442 TI - Synthesis of 4-diphosphocytidyl-2-C-methyl-D-erythritol and 2-C-methyl-D erythritol-4-phosphate. AB - 2-C-Methyl-D-erythritol 4-phosphate (MEP, 2) and 4-diphosphocytidyl-2-C-methyl-D erythritol (CDPME, 3) are metabolites in the MEP pathway for biosynthesis of isoprenoid compounds in bacteria, plant chloroplasts, and algae. The free phosphoacid of 2 was prepared from benzyloxyacetone in five steps with an overall yield of 27% and an enantiomeric ratio (er) of 75:25. Following titration to the corresponding tributylammonium salt, 2 was coupled to cytidine 5'-monophosphate using a protocol originally developed for synthesis of base-sensitive nucleoside diphosphate sugars to give 3 in 40% yield, following purification by size exclusion chromatography. PMID- 12126443 TI - First stereoselective synthesis of 2-deoxy-alpha-D-ribosyl-1-phosphate: novel application of crystallization-induced asymmetric transformation. AB - A first stereoselective synthesis of bis(cyclohexylamine) 2-deoxy-alpha-D-ribosyl 1-phosphate has been achieved. The synthesis features a key crystallization induced asymmetric transformation (AT) to generate a desired alpha-anomer in 99% yield at a 98.8:1.2 ratio of alpha/beta. PMID- 12126444 TI - Synthesis of new tricyclic phosphines and phosphinites by intramolecular Diels Alder reactions of trivalent phospholes. AB - Phospholes bearing an allyl-X substituent at phosphorus tend to undergo an intramolecular Diels-Alder cycloaddition (IMDA) leading to the corresponding tricyclic derivative. When X = O or NR, the IMDA easily takes place at room temperature. When X = CH2, the IMDA slowly takes place around 110-140 degrees C, as a function to the substitution pattern of the dienic system. Two tricyclic derivatives (X = O and CH2) have been characterized by X-ray crystal structure analysis of the P-sulfides. PMID- 12126445 TI - Boronic acid receptors for alpha-hydroxycarboxylates: high affinity of Shinkai's glucose receptor for tartrate. AB - The glucose receptor 1 developed by Shinkai was synthesized by known methods and with modifications involving the final synthetic step, installation of the phenylboronic acid moieties. Binding of the bis(alpha-hydroxycarbolxylate), tartrate, was assessed and compared to the corresponding bis(diol), erythritol, as well as the corresponding mono(alpha-hydoxycarboxylate), malate. These results suggest that bisboronate/bis(alpha-hydoxycarboxylate) interactions are stronger than the corresponding bisboronate/bis(diol) interactions. Furthermore, we report that the receptor is an order of magnitude more selective for tartrate than malate. PMID- 12126446 TI - Sebastianines A and B, novel biologically active pyridoacridine alkaloids from the Brazilian ascidian Cystodytes dellechiajei. AB - Fractionation of the crude methanol extract of the ascidian Cystodytes dellechiajei collected in Brazil yielded two novel alkaloids, sebastianine A (1) and sebastianine B (2). The structures of both 1 and 2 were established by analysis of spectroscopic data, indicating an unprecedented ring system for both compounds, comprising a pyridoacridine system fused with a pyrrole unit in sebastianine A (1) and a pyridoacridine system fused with a pyrrolidine system condensed with alpha-hydroxyisovaleric acid in sebastianine B (2). Both alkaloids displayed a cytotoxic profile against a panel of HCT-116 colon carcinoma cells indicative of a p53 dependent mechanism. PMID- 12126447 TI - Stereoselective syntheses of the three isomers of ethylene glycol bis(tropane-3 carboxylate). AB - Three epimers of ethylene glycol bis(tropane-3-carboxylate) (3alpha,3alpha'-, 3alpha,3beta'-, 3beta,3beta'-) have been synthesized by starting from 3 tropinone. 3-Tropinone was converted into the corresponding enol triflate and then subjected to palladium-catalyzed alkoxycarbonylation to provide the key intermediate methyl trop-2-ene-3-carboxylate in good yield. Stereoselective routes were developed to afford the three stereoisomers of ethylene glycol bis(tropane-3-carboxylate). PMID- 12126448 TI - A new bis(2,2,2-trifluoroethyl)phosphonate for the synthesis of Z-unsaturated N methoxy-N-methylamides. AB - The N-methoxy-N-methyl bis(2,2,2-trifluoroethyl)phosphonamide was easily obtained via the Arbuzov reaction with use of commercially available tris(2,2,2 trifluoroethyl)phosphite, 2-bromo-N-methoxy-N-methylacetamide, and KF/alumina. The reaction of bis(2,2,2-trifluoroethyl)phosphonate with several aldehydes demonstrates the versatility of the method, which gives Z-unsaturated amides in good yields. PMID- 12126449 TI - Rearrangement of epoxynitriles: a convenient homologation of acyclic and cyclic ketones to carboxylic acids. AB - A convenient two-step homologation of both aliphatic and aromatic ketones to the corresponding carboxylic acid has been developed. First ketones were converted to epoxynitriles with the Darzens reaction. Second, a Lewis acid mediated rearrangement of these epoxynitriles with lithium bromide was achieved to give homologated secondary alkanoic acids (as well as aryl-alkanoic) in good yields. The mechanism and the scope of the rearrangement reaction were investigated. This strategy constitutes a two-step homologation of ketones to secondary carboxylic acids. PMID- 12126452 TI - Lung surfactants for neonatal respiratory distress syndrome: animal-derived or synthetic agents? AB - Exogenous surfactant therapy is widely used in the management of neonatal respiratory distress syndrome. Two types of surfactants are available: synthetic surfactants, and those derived from animal sources ("natural" surfactants). Both of these surfactants have been shown to be effective. In this article, we review the evidence to compare the two types of surfactants in terms of their physical properties, physiologic effects, and clinical outcomes. Natural surfactants have been shown to have advantages over synthetic surfactants in their physical properties and physiologic effects in animals, as well as in humans. A systematic review of 11 randomized clinical trials comparing natural and synthetic surfactants demonstrated that the use of natural surfactant preparations results in greater clinical benefits compared with synthetic surfactants. These benefits include a more rapid improvement in oxygenation and lung compliance after surfactant therapy, a decrease in the risk of mortality (typical relative risk 0.87; typical risk difference -0.02), and a decrease in the risk of pneumothorax (typical relative risk 0.63; typical risk difference -0.04). Although the use of natural surfactants results in a slightly increased risk of intraventricular hemorrhage (typical relative risk 1.09; typical risk difference 0.03), there is no increase in the risk of grade 3 or 4 intraventricular hemorrhage. There are theoretical but unproven risks of natural surfactants, such as transmission of infectious agents, immunogenicity and impurities in composition. The use of natural surfactants is preferred in most situations. In addition, clinicians should determine the costs of different types of surfactants in their individual practice settings and use this information in decision-making. PMID- 12126453 TI - Alcohol use disorders in adolescents: epidemiology, diagnosis, psychosocial interventions, and pharmacological treatment. AB - Alcohol (ethanol) abuse and dependence are the most common substance use disorders among adolescents. Binge drinking occurs in up to one-third of adolescents, and alcohol use disorders occur in about 6% of this age group. Adolescents with alcohol use disorders also typically have problems with other substances and comorbid mental disorders. Validated measures are available for the clinical detection and diagnosis of adolescent alcohol use disorders and related problems. Psychosocial interventions promoting abstinence are the most common treatments for alcohol use disorders, with empirical support particularly strong for family-based approaches. Pharmacological interventions may diminish the effects of alcohol withdrawal, prevent a return to alcohol consumption, or treat comorbid mental disorders. In this population, pharmacological interventions require further investigation and, where indicated, are generally considered to be supplementary to psychosocial approaches. PMID- 12126454 TI - Drug compliance in adolescents: assessing and managing modifiable risk factors. AB - Many studies have found that adolescence represents a problem in compliance with prescribed drug regimens. Multiple factors contribute to this problem, including the developmental evolution taking place in the adolescent physique and psyche. Health belief and patient demographic factors, inherent disease and regimen factors, as well as the dynamics between patient and provider may also contribute to problems with compliance to treatment. Simple interventions such as working with the teen to construct a tolerable treatment regimen, assessing anticipated compliance, discussing potential adverse effects, and establishing cues from the adolescent's daily routine can positively impact treatment compliance. Healthcare providers should recognize the fact that psychosocial changes in an adolescent's life can impact upon compliance with medications and enlist the help of their patients in constructing treatment regimens taking into account the individual's lifestyle that may impact upon compliance. In particular, the healthcare provider should ask the adolescents what they anticipate their success with compliance to treatment might be, adverse effects they are concerned about and what cues could best aid the treatment plan. The healthcare provider should then synthesize this information to create the best treatment plan for that patient. PMID- 12126456 TI - Distribution of cyclosporin in organ transplant recipients. AB - Cyclosporin is an immunosuppressive agent with a narrow therapeutic index. The total concentration of cyclosporin in blood is usually monitored to guide dosage adjustment and to compensate for substantial interindividual and intraindividual variability in cyclosporin pharmacokinetics. Cyclosporin is a highly lipophilic molecule and widely distributes into blood, plasma and tissue components. It mainly accumulates in fat-rich organs, including adipose tissue and liver. In blood, it binds to erythrocytes in a saturable fashion that is dependent on haematocrit, temperature and the concentration of plasma proteins. In plasma, it binds primarily to lipoproteins, including high-density, low-density and very-low density lipoprotein, and, to a lesser extent, albumin. The unbound fraction of cyclosporin in plasma (CsA(fu)) expressed as a percentage is approximately 2%. It has been shown that both the pharmacokinetic and pharmacodynamic properties of cyclosporin are related to its binding characteristics in plasma. Furthermore, there is some evidence to indicate that the unbound concentration of cyclosporin (CsA(U)) has a closer association with both kidney and heart allograft rejection than the total (bound + unbound) concentration. However, the measurement of CsA(fu) is inherently complex and cannot easily be performed in a clinical setting. Mathematical models that calculate CsA(fu), and hence CsA(U), from the concentration of plasma lipoproteins may be a more practical option, and should provide a more accurate correlate of effectiveness and toxicity of this drug in transplant recipients than do conventional monitoring procedures. In conclusion, the distribution characteristics of cyclosporin in blood, plasma and various tissues are clinically important. Further investigations are needed to verify whether determination of CsA(U) improves the clinical management of transplant recipients. PMID- 12126455 TI - Zidovudine: a review of its use in the management of vertically-acquired pediatric HIV infection. AB - Zidovudine is a thymidine analog that, after intracellular phosphorylation to zidovudine triphosphate metabolite, inhibits HIV-specific reverse transcriptase and terminates proviral DNA. Zidovudine administered to mildly symptomatic women with HIV infection in the antepartum (100mg orally 5 times/day), intrapartum (2 mg/kg intravenously over 1 hour then 1 mg/kg/h) and then to the neonate for 6 weeks (2 mg/kg), significantly reduced the rate of vertical HIV transmission by about two thirds, in the absence of breast-feeding (The Pediatric AIDS Clinical Trials Group 076 trial, standard protocol). Shorter zidovudine regimens, reduced the risk of transmission of HIV by 50% in a non-breast-feeding population and by about 37% in breast-feeding populations. Zidovudine (standard protocol) in combination with lamivudine was superior to zidovudine alone. A short oral zidovudine regimen was not as effective as a two-dose oral nevirapine regime, although the combination of short-course zidovudine plus lamivudine was as effective. Suppression of viral replication in neonates, infants and children has been achieved with zidovudine when used in triple-therapy regimens that include other antiviral drugs. Results from a trial of treatment-naive children indicate that the antiviral efficacy of combinations of zidovudine and lamivudine or abacavir, given with the protease inhibitor nelfinavir, is superior to treatment with this combination minus nelfinavir. When zidovudine was used in other highly active antiretroviral therapy regimens significant improvements in surrogate markers were consistently seen. Changing to ritonavir-containing regimens was superior to changing to treatment with two new nucleoside reverse transcriptase inhibitors. Short- and long-term (up to 5.6 years) outcomes from clinical trials showed that prenatal and neonatal exposure to zidovudine was generally well tolerated with the exception of mild anemia that resolved spontaneously after treatment cessation. Zidovudine was generally well tolerated as monotherapy in clinical trials of pediatric patients with HIV infection, and adverse events were similar to those reported in adults, with anemia and neutropenia being the most common. CONCLUSION: Zidovudine, as monotherapy or in combination with other antiretroviral agents, remains a first-choice therapy for the prophylaxis of mother-to-child HIV transmission as shown by substantial reductions in transmission rates. Where feasible, the optimal strategy to prevent vertical transmission is to combine drug therapy with Cesarean section delivery and no breast-feeding. In addition, zidovudine in combination with another nucleoside analogue and a protease inhibitor is a first- or second-choice therapy for the treatment of pediatric HIV infection as significant and sustained reductions in viral load have been shown in both plasma and cerebrospinal fluid. PMID- 12126459 TI - Pharmacokinetic interactions between efavirenz and rifampicin in HIV-infected patients with tuberculosis. AB - OBJECTIVE: To evaluate the pharmacokinetic interactions between efavirenz and rifampicin (rifampin) in patients with HIV infection and tuberculosis. DESIGN: Nonblind, randomised, pharmacokinetic study. PATIENTS: 24 patients (21 male, 3 female; mean age 37 years) with HIV infection and tuberculosis. INTERVENTIONS: Patients were randomised to one of the following treatments: group A (n = 16) received antituberculosis drugs without rifampicin, plus highly active antiretroviral therapy (HAART) including efavirenz 600 mg once daily, on days 1 to 7. Patients were then switched to rifampicin in bodyweight-adjusted fixed-dose combination plus HAART including efavirenz 600 mg once daily (group A-1; n = 8) or efavirenz 800 mg once daily (group A-2; n = 8). Group B (n = 8) received rifampicin in bodyweight-adjusted fixed-dose combination on days 1 to 7; on day 8, HAART including efavirenz 800 mg once daily was added. Blood samples were obtained on days 7 and 14. METHODS: Plasma concentrations of efavirenz and rifampicin were quantified by using validated high performance liquid chromatography assays, and pharmacokinetic parameter values were determined by noncompartmental methods. The differences between pharmacokinetic parameters on days 7 and 14 were used to assess interactions. RESULTS: There was a correlation between the pharmacokinetic parameters of efavirenz and the dose/kg administered. For efavirenz, mean (median) peak concentration, trough concentration and area under the concentration-time curve over the administration interval decreased 24% (24%), 25% (18%) and 22% (10%), respectively, in the presence of rifampicin. Large interpatient variability was observed, suggesting that plasma concentration monitoring of efavirenz may be advisable. Overall, the pharmacokinetics of efavirenz 800 mg plus rifampicin were similar to those of efavirenz 600 mg without rifampicin. The pharmacokinetics of rifampicin did not change substantially in the presence of efavirenz. Differences in patients' bodyweight appeared to cause further differences in exposure to efavirenz. Plasma concentrations of efavirenz in patients weighing <50 kg were similar to those previously described in HIV-infected patients without concomitant tuberculosis. However, plasma concentrations in patients weighing >or=50 kg were almost halved compared with those in patients weighing <50 kg. CONCLUSIONS: Although the minimal effective efavirenz plasma concentration that assures virological success is not currently known, it may be advisable to increase the dosage of efavirenz to 800 mg once daily when it is coadministered with rifampicin. Rifampicin can be used with efavirenz without dosage modification. PMID- 12126460 TI - Seeing red. PMID- 12126458 TI - Oral mucosal drug delivery: clinical pharmacokinetics and therapeutic applications. AB - Oral mucosal drug delivery is an alternative method of systemic drug delivery that offers several advantages over both injectable and enteral methods. Because the oral mucosa is highly vascularised, drugs that are absorbed through the oral mucosa directly enter the systemic circulation, bypassing the gastrointestinal tract and first-pass metabolism in the liver. For some drugs, this results in rapid onset of action via a more comfortable and convenient delivery route than the intravenous route. Not all drugs, however, can be administered through the oral mucosa because of the characteristics of the oral mucosa and the physicochemical properties of the drug. Several cardiovascular drugs administered transmucosally have been studied extensively. Nitroglycerin is one of the most common drugs delivered through the oral mucosa. Research on other cardiovascular drugs, such as captopril, verapamil and propafenone, has proven promising. Oral transmucosal delivery of analgesics has received considerable attention. Oral transmucosal fentanyl is designed to deliver rapid analgesia for breakthrough pain, providing patients with a noninvasive, easy to use and nonintimidating option. For analgesics that are used to treat mild to moderate pain, rapid onset has relatively little benefit and oral mucosal delivery is a poor option. Oral mucosal delivery of sedatives such as midazolam, triazolam and etomidate has shown favourable results with clinical advantages over other routes of administration. Oral mucosal delivery of the antinausea drugs scopolamine and prochlorperazine has received some attention, as has oral mucosal delivery of drugs for erectile dysfunction. Oral transmucosal formulations of testosterone and estrogen have been developed. In clinical studies, sublingual testosterone has been shown to result in increases in lean muscle mass and muscle strength, improvement in positive mood parameters, and increases in genital responsiveness in women. Short-term administration of estrogen to menopausal women with cardiovascular disease has been shown to produce coronary and peripheral vasodilation, reduction of vascular resistance and improvement in endothelial function. Studies of sublingual administration of estrogen are needed to clarify the most beneficial regimen. Although many drugs have been evaluated for oral transmucosal delivery, few are commercially available. The clinical need for oral transmucosal delivery of a drug must be high enough to offset the high costs associated with developing this type of product. Drugs considered for oral transmucosal delivery are limited to existing products, and until there is a change in the selection and development process for new drugs, candidates for oral transmucosal delivery will be limited. PMID- 12126457 TI - Lithium in bipolar disorder: can drug concentrations predict therapeutic effect? AB - The evidence is reviewed for effective serum lithium concentrations for the acute and prophylactic treatment of mania and depression in patients with bipolar disorder. The efficacy of lithium in the treatment of acute manic episodes has been recognised for several decades, primarily using concentrations in the range of 0.8 to 2 mmol/L. The number of patients responding increases as the serum lithium concentration increases, although individual patients may respond at lower concentrations (<0.8 mmol/L). Lithium doses and serum concentrations similar to those used to treat acute mania have been studied in bipolar depression, with no evaluation of a relationship between concentration and clinical response. Several prospective controlled trials have evaluated this relationship in the prophylactic treatment of bipolar disorder. Maintaining higher serum lithium concentrations (0.8 to 1 mmol/L) improves the likelihood of good effect in prophylactic treatment, although individual patients may do well on lower concentrations. Despite the paucity of evidence to specifically support the efficacy of lithium at lower serum lithium concentrations in the elderly, lower target ranges (0.5 to 0.8 mmol/L) are commonly recommended due to an increased sensitivity to adverse effects, particularly neurotoxicity. The serum lithium concentrations recommended in adults have been applied to children; however, this has not been studied. Overall, the evidence suggests a relationship between serum lithium concentration and therapeutic effect, although the exact nature of this relationship is not clear. For example, it is not known why some people respond to lower concentrations and others do not. There are many factors that influence studies trying to elucidate this relationship. Many of these factors are related to the interpretation of the serum lithium concentration. In summary, patients have an increased chance of responding to lithium if 12-hour serum lithium concentrations at steady state are above 0.8 mmol/L. Many patients will respond to lower concentrations (0.4 to 0.7 mmol/L), but we are unable to identify these patients a priori. The relationship between serum lithium concentrations and adverse effects is also very important in determining appropriate target lithium concentrations. The current best advice is to individualise the target serum lithium concentrations based on efficacy and tolerability and to optimise the interpretation of these concentrations by ensuring within-patient consistency with respect to dosage schedule, lithium preparation and the timing of blood sampling. PMID- 12126461 TI - Eating disorders: a review of the literature with emphasis on medical complications and clinical nutrition. AB - Eating disorders, including anorexia nervosa, bulimia nervosa, binge-eating disorder, and atypical eating disorder (eating disorder not otherwise specified or NOS), are estimated to occur in 5-10 million young and adult women and one million males in the United States. The etiology of eating disorders is complex and appears to include predisposing genetic factors and serotonin dysregulation, as well as psychological factors that include a history of trauma and childhood sexual abuse. Both anorexia nervosa and bulimia nervosa are medical conditions complicated by multiple neuroendocrine dysfunctions, nutritional deficiencies, and psychiatric diagnoses. Medical complications, specific nutritional deficiencies, and research involving the therapeutic use of inositol and zinc are reviewed. PMID- 12126462 TI - Policosanol: a new treatment for cardiovascular disease? AB - Policosanol is a mixture of alcohols isolated and purified from sugar cane. Recently, Cuban researchers found 5-20 mg daily of policosanol to be effective at improving serum lipid profiles. Policosanol is believed to decrease total cholesterol (TC), low-density lipoprotein (LDL), and increase high-density lipoprotein (HDL) by inhibiting cholesterol synthesis and increasing LDL processing. Lipid profile improvements are seen in healthy volunteers, patients with type II hypercholesterolemia, type 2 diabetics with hypercholesterolemia, postmenopausal women with hypercholesterolemia, and patients with combined hypercholesterolemia and abnormal liver function tests. Additionally, policosanol has performed equal to or better than simvastatin, pravastatin, lovastatin, probucol, or acipimox with fewer side effects in patients with type II hypercholesterolemia. Policosanol also decreases several other risk factors of cardiovascular disease by decreasing LDL oxidation, platelet aggregation, endothelial damage, and smooth muscle cell proliferation. Furthermore, policosanol decreases progression and increases regression of cardiovascular disease assessed by thallium-labeled myocardial perfusion scintigraphy (TL-MPS) and Doppler-ultrasound, and decreases symptoms of cardiovascular disease assessed by the Specific Activity Scale. In post-marketing studies, only 0.31 percent of patients have had adverse events. Furthermore, in animal toxicity studies doses up to 1500 times normal human doses (on the basis of body weight) have shown no negative effects on carcinogenesis, reproduction, growth, and development. However, despite the positive research on policosanol on Cubans, policosanol produced in Cuba is not available in the United States, and only Cuban subjects have been studied. Further research is needed to determine if the same effects will be obtained in U.S. populations with non-Cuban produced policosanol. PMID- 12126463 TI - The safety and efficacy of high-dose chromium. AB - The data on the standards for chromium requirements and the safety of various chromium compounds and doses are reviewed. The 350-fold difference between the acceptable daily intake and the calculated reference dose for humans of 70 mg per day seems without precedent with respect to other nutritional minerals. Previous claims of mutagenic effects of chromium are of questionable relevance. While studies have found DNA fragmentation (clastogenic effects) by chromium picolinate, anecdotal reports of high-dose chromium picolinate toxicity are few and ambiguous. The beneficial effects of chromium on serum glucose and lipids and insulin resistance occur even in the healthy. Serum glucose can be improved by chromium supplementation in both types 1 and 2 diabetes, and the effect appears dose dependent. Relative absorption of various chromium compounds is summarized and the mechanism of low molecular weight chromium binding substance (LMWCr) in up-regulating the insulin effect eight-fold is discussed. There is evidence of hormonal effects of supplemental chromium besides the effect on insulin. Chromium supplementation does result in tissue retention, especially in the kidney, although no pathogenic effect has been demonstrated despite considerable study. PMID- 12126464 TI - Can maitake MD-fraction aid cancer patients? AB - Maitake mushroom (Grifola frondosa) MD-fraction containing beta-1,6 glucan with beta-1,3 branched chains has previously exhibited strong anticancer activity by increasing immune-competent cell activity.1,2 In this non-random case series, a combination of MD-fraction and whole maitake powder was investigated to determine its effectiveness for 22- to 57-year-old cancer patients in stages II-IV. Cancer regression or significant symptom improvement was observed in 58.3 percent of liver cancer patients, 68.8 percent of breast cancer patients, and 62.5 percent of lung cancer patients. The trial found a less than 10-20 percent improvement for leukemia, stomach cancer, and brain cancer patients. Furthermore, when maitake was taken in addition to chemotherapy, immune-competent cell activities were enhanced 1.2-1.4 times, compared with chemotherapy alone. Animal studies have supported the use of maitake MD-fraction for cancer. PMID- 12126465 TI - Vinpocetine. Monograph. PMID- 12126466 TI - Inositol hexaphosphate. Monograph. PMID- 12126467 TI - VPg unlinkase, the phosphodiesterase that hydrolyzes the bond between VPg and picornavirus RNA: a minimal nucleic moiety of the substrate. AB - VPg unlinkase is an unusual eukaryotic enzyme that catalyzes hydrolysis of the phosphodiester bond between residues of the unique tyrosine of VPg (viral protein genome-linked) and the 5;-terminal uridylic acid of picornavirus RNA. Cellular targets of the VPg unlinking enzyme are yet unknown. To determine an essential nucleic part of the covalent linkage unit that is necessary for the VPg unlinkase reaction, the following derivatives of the encephalomyocarditis virus (EMCV) VPg RNA complex were used: [125I]Kp-pUpUpGp, [125I]Kp-pUp, and [125I]Kp-pU (Kp is residual peptides bound to RNA after proteinase K treatment of VPg-RNA). [125I]K peptides were unlinked from [125I]Kp-pUpUpGp and [125I]Kp-RNA with similar velocity, but [125I]Kp-pUp was split much slower. Under the same conditions [125I]Kp-pU was not dissociated at all. Thus, pUp is a minimal part of picornavirus RNA that is necessary for VPg unlinkase. We speculate that cellular substrates of the enzyme are phosphodiesters of oligo(poly)ribonucleotides and tyrosine or tyrosine peptides. In no case [125I]VPg-pU, [125I]VPg-pUp, and [125I]VPg-pUpUpGp were hydrolyzed by VPg unlinkase, in contrast with [125I]VPg RNA and [125I]VPg-pUpUpGpApApApGp. We conclude that the whole VPg, when bound to trinucleotide (but not to heptanucleotide), protects the inter-polymeric phosphodiester bond against hydrolysis of the covalent linkage unit. We speculate that VPg unlinkase might repair covalent complexes of RNA and topoisomerases and trigger degradation process of the picornavirus RNA. PMID- 12126468 TI - Operation of the cbb3-type terminal oxidase in Azotobacter vinelandii. AB - A part of the gene encoding cbb3-type cytochrome oxidase CcoN subunit was cloned from Azotobacter vinelandii and a mutant strain of this bacterium with disrupted ccoN gene was constructed. In contrast to the wild type strain, this one is unable to oxidize cytochromes c4 and c5. Thus, the A. vinelandii respiratory chain is shown to contain cbb3-type cytochrome c oxidase. It is also shown that the activity of this enzyme is not necessary for diazotrophic growth of A. vinelandii at high oxygen concentrations. PMID- 12126469 TI - Involvement of chloroplasts in the programmed death of plant cells. AB - The effect of cyanide, an apoptosis inducer, on pea leaf epidermal peels was investigated. Illumination stimulated the CN--induced destruction of guard cells (containing chloroplasts and mitochondria) but not of epidermal cells (containing mitochondria only). The process was prevented by antioxidants (alpha-tocopherol, 2,5-di-tret-butyl-4-hydroxytoluene, and mannitol), by anaerobiosis, by the protein kinase C inhibitor staurosporine, and by cysteine and serine protease inhibitors. Electron acceptors (menadione, p-benzoquinone, diaminodurene, TMPD, DCPIP, and methyl viologen) suppressed CN--induced apoptosis of guard cells, but not epidermal cells. Methyl viologen had no influence on the removal of CN- induced nucleus destruction in guard cells under anaerobic conditions. The light activation of CN--induced apoptosis of guard cells was suppressed by DCMU (an inhibitor of the electron transfer in Photosystem II) and by DNP-INT (an antagonist of plastoquinol at the Qo site of the chloroplast cytochrome b6f complex). It is concluded that apoptosis initiation in guard cells depends on the simultaneous availability of two factors, ROS and reduced quinones of the electron transfer chain. The conditions for manifestation of programmed cell death in guard and epidermal cells of the pea leaf were significantly different. PMID- 12126470 TI - Determination of Mn(II) and Co(II) with Arsenazo III. AB - Complex formation between Arsenazo III and Mn2+ and Co2+ at equilibrium has been investigated at pH 7.2, and the stoichiometry and stability of the complexes have been determined. The data indicate that Arsenazo III is suitable for determination of Mn2+ and Co2+ on the micromolar scale. The dissociation constants of the phosphate complexes of Mn2+ and Co2+ at pH 7.2 were estimated with Arsenazo III as 3.6 and 10 mM, respectively. PMID- 12126471 TI - Epitope mapping of the outer surface protein A (OspA) of the spirochete Borrelia burgdorferi using a panel of monoclonal antibodies and lanthanide competition fluoroimmunoassay. AB - A panel of fourteen different monoclonal antibodies was used for detection and analysis of antigenic determinants located on the outer surface protein A (OspA) of the spirochete Borrelia burgdorferi, which is a causative agent of tick-borne borreliosis (Lyme disease). Two main and several minor partially overlapping antigenic determinants have been found on the surface of the OspA protein of Borrelia burgdorferi sensu stricto (strain 297) by lanthanide competition fluoroimmunoassay. One of the main antigenic determinants is located in the N- and the other in the C-half of the OspA molecule. The involvement of the OspA protein in intact Borrelia burgdorferi sensu stricto (four bacterial strains have been analyzed: 297, B31, FR90-594, and CA90-742) is associated with retention of the above-mentioned two major antigenic determinants, but unlike the case of the isolated OspA they are partially overlapping with each other and with other antigenic determinants. The protein of the spirochete Borrelia afzelii (two bacterial strains have been analyzed: Ip-21 and Pko) contains only one antigenic determinant, which is the same as the main determinant of the OspA protein of Borrelia burgdorferi sensu stricto located in the N-half of the OspA molecule. PMID- 12126472 TI - Kinetics of the transhydrogenase reaction catalyzed by mitochondrial NADH:ubiquinone oxidoreductase (complex I). AB - The kinetics of the NADH-->3;-acetylpyridine adenine dinucleotide (APAD+) transhydrogenase reaction (DD-reaction) catalyzed by different preparations of mitochondrial NADH-dehydrogenase (submitochondrial particles (SMP), purified Complex I, and three-subunit fragment of Complex I (FP)) have been studied. Complex I (in SMP or in purified preparation) catalyzes two NADH-->APAD+ reactions with different rates and nucleotide affinities. Reaction 1 has high affinity to APAD+ (Km = 7 microM, for SMP) and low rate (Vm = 0.2 micromol/min per mg protein, for SMP) and occurs with formation of a ternary complex. Reaction 2 has much higher rate and considerably lower affinity for oxidized nucleotide (Vm = 1.7 micromol/min per mg protein and Km = 160 microM, for SMP). FP catalyzes only reaction 1. ADP-ribose inhibits reaction 1 with mixed type inhibition (competitive with non-competitive) with respect to NADH and APAD+. Rhein competes with both substrates. The results suggest that at least two nucleotide-binding sites exist in Complex I. PMID- 12126473 TI - Influence of platelet-activating factor, its cell analogs, and antagonist on the production of superoxide radicals by blood leukocytes of healthy and hypercholesterolemic individuals. AB - The influence of the phospholipid platelet-activating factor (PAF), its cell analogs, and lipid PAF antagonist on the production of superoxide radicals by leukocytes isolated from the blood of healthy and hypercholesterolemia IIA individuals was studied. It was found that endogenous superoxide production level in the leukocytes of hypercholesterolemic individuals more than 4-5 times higher than in the leukocytes of healthy individuals. Exogenous PAF stimulates the superoxide production in the leukocytes of healthy individuals but significantly inhibits the superoxide production in the leukocytes of hypercholesterolemic individuals. The compounds 1-acyl-2-acetyl-sn-glycero-3-phosphocholine (1-acyl PAF) and 1-alk-1;-enyl-2-acetyl-sn-glycero-3-phosphocholine (1-alkenyl-PAF) only slightly inhibited the endogenous superoxide production in the leukocytes of hypercholesterolemia individuals. However, pretreatment of leukocytes by 1 alkenyl-PAF or PAF-antagonist (1-O-alk-1;-enyl-2-(2;-acetoxybenzoyl)-sn-glycero-3 phosphocholine) results in a 50% inhibition of the PAF-induced superoxide production by leukocytes of healthy individuals. This PAF-antagonist alone or in combination with PAF induces a substantial (65-70%) inhibition of superoxide production in the leukocytes of hypercholesterolemic individuals. It is concluded that superoxide production by leukocytes of healthy individuals and especially by leukocytes of hypercholesterolemic individuals is process that depends on PAF or PAF-like lipids. PMID- 12126474 TI - Isolation and properties of noncovalent complex of transketolase with RNA. AB - A method for isolation of homogenous transketolase from baker's yeast using immunoaffinity chromatography was significantly simplified. It was demonstrated that transketolase could be isolated from fresh yeast in the form of a complex with a high molecular weight RNA. Storage of yeast led to the dissociation of the complex to a low molecular weight complex and then to the free enzyme. Conditions were chosen for complex dissociation and free enzyme isolation. In comparison to the free enzyme, the specific activities of the high and low molecular weight complexes were decreased 20-25- and 3-5.5-fold, respectively. The affinity to the cofactor thiamine diphosphate and to xylulose-5-phosphate (donor substrate) did not change for the low molecular weight complex, while the time of binding to calcium increased. The latter was necessary for the complete manifestation of the enzymatic activity. Changes in the circular dichroism spectrum between 300 and 360 nm after the addition of thiamine diphosphate, which characterize the formation of the catalytically active holoenzyme, were significantly lower for the low molecular weight complex than for the free enzyme. PMID- 12126475 TI - Structure of O-specific polysaccharide from Pseudoalteromonas nigrifaciens strain KMM 161. AB - On mild acid degradation of the lipopolysaccharide of the marine microorganism Pseudoalteromonas nigrifaciens KMM 161 an O-specific polysaccharide containing D galactose, 2-acetamido-2-deoxy-D-glucose, 3,6-dideoxy-3-(4-hydroxybutyramido)-D galactose, and 2-acetamido-2-deoxy-L-guluronic acid residues was obtained. From the results of Smith degradation, O-deacetylation of the polysaccharide, and NMR spectroscopy the following structure of the tetrasaccharide repeating unit of the O-specific polysaccharide was established [see reaction]. It should be noted that the same structure occurs in the antigenic polysaccharide of Pseudoalteromonas nigrifaciens KMM 158 described earlier as Alteromonas macleodii 2MM6. PMID- 12126476 TI - Viscometric method for assaying of total endodepolymerase activity of pectinases. AB - An improved method for assaying of the total endodepolymerase activity of pectinases has been developed. The method is based on the determination of the viscosity of a citrus pectin solution in the presence of the enzyme using an Ostwald viscometer. The depolymerizing activity of different pectinases can be detected including polygalacturonase, polymethylgalacturonase, pectin lyase, and pectate lyase. One unit of the endodepolymerase activity corresponds to the activity resulting in 50% decrease in the relative viscosity of 0.5% citrus pectin solution for 5 min at 40 degrees C and the appropriate pH. Depending on the pH-optima of the enzymes, two modifications of the method are described: 1) for acid pectinases at pH 5.0, and 2) for neutral (mildly alkaline) pectinases at pH 8.0. The modifications differed in the control and in the calculation of the activity. Six enzyme preparations were used to demonstrate the applicability of the method. The parameter used for the calculation of the enzymatic activity was directly proportional to the enzyme concentration (the dependence was linear in the range of at least 10-fold change in the enzyme concentration). The relative error of the method did not exceed 10%. PMID- 12126477 TI - Photoconsumption of oxygen in photosystem II preparations under impairment of the water-oxidizing complex. AB - Oxygen consumption in photosystem II (PSII) preparations in the light was 2 micromol O2/h per mg Chl at weakly acidic and at neutral pH values. It increased fourfold to fivefold at pH 8.5-9.0. The addition of either artificial electron donors for PSII such as MnCl2 or diphenylcarbazide, or diuron as an inhibitor of electron transfer from QA, the primary bound quinone acceptor, to QB, the secondary bound quinone acceptor of PSII, resulted in a decrease in oxygen consumption rate at basic pH to value close to ones measured at pH 6.5. Such additions did not affect oxygen consumption at lower pH values. The induction of variable chlorophyll fluorescence yield in the light differed greatly at pH 6.5 and 8.5. While at pH 6.5 the fluorescence yield, after an initial fast rise almost to Fmax, only slightly decreased, at pH 8.5 after such a rise it dropped promptly to a low value. The additions of the artificial electron donors at pH 8.5 resulted in the induction kinetics close to that observed at pH 6.5. These data indicate impairment of electron donation to P680+ that could be caused by damage to the water oxidation system at basic pH values. In experiments with PSII preparations treated with Tris to destroy the water-oxidizing complex, photoconsumption of oxygen in the entire pH region was close to the values in untreated preparations at basic pH. In untreated preparations the rate of light induced oxygen consumption decreased in the presence of catalase, which decomposes H2O2, as well as in the presence of electron acceptor potassium ferricyanide. From these data it is suggested that the light-induced oxygen consumption in PSII is caused by two processes, by an interaction of O2 with organic radicals, which were formed due to oxidation of components of the donor side of this photosystem (proteins, lipids, pigments) by cation-radical P680+, as well as by oxygen reduction by still unidentified components of PSII. PMID- 12126478 TI - Alpha-N-acetylgalactosaminidase from marine bacterium Arenibacter latericius KMM 426T removing blood type specificity of A-erythrocytes. AB - An alpha-N-acetylgalactosaminidase IV able to remove blood type specificity of human A(II)-erythrocytes and not effecting B(III)-erythrocytes was isolated from the marine bacterium Arenibacter latericius KMM 426T. The alpha-N acetylgalactosaminidase IV preparation exhibits high activity during inhibition of hemagglutination with blood group substance A containing determinants analogous to A-erythrocytes. The enzyme has a pH optimum from 7.0 to 8.0 and completely retains its activity during 30-min heating at 50 degrees C and for a week at 20 degrees C. The enzyme can be stored under the sterile conditions for any length of time at 4 degrees C, but it does not withstand freezing. The alpha N-acetylgalactosaminidase is resistant to NaCl; for p-nitrophenyl-alpha-N-acetyl D-galactosaminide, the Km is 0.38 mM. The molecular mass of the enzyme determined by gel filtration is 84 kD. PMID- 12126479 TI - Intensity of free radical processes and regulation of cytoplasmic NADP-isocitrate dehydrogenase in rat cardiomyocytes under normal and ischemic conditions. AB - The intensity of free radical processes and the regulation of NADP-isocitrate dehydrogenase (EC 1.1.1.42; NADP-IDH) activity have been studied in the cytoplasmic fraction of normal and ischemized rat myocardium. Chemiluminescence parameters, such as the light sum (S) of slow flash and the tangent of the kinetic curve slope angle (tanalpha1), which characterize the intensity of free radical processes, were increased in ischemia 2.1- and 20.0-fold, respectively. The slow flash intensity (Imax) was increased 22-fold. The contents of lipid peroxidation products--diene conjugates and malonic dialdehyde--were increased 11.9- and 4.7-fold, respectively, suggesting pronounced oxidative stress. Using homogenous enzyme preparations of NADP-IDH isolated from the normal and experimentally ischemized rat myocardium, a number of catalytic properties of the enzyme were characterized for normal and pathologic conditions. NADP-IDH from the normal and ischemized myocardium had the same electrophoretic mobility and was regulated similarly by Fe2+, Cu2+, Zn2+, and also with succinate and fumarate. However, under normal and pathologic conditions NADP-IDH was different in the affinity for substrates and in the sensitivity to inhibitory effects of hydrogen peroxide, reduced glutathione, and of Ca2+. The degree of synergy in the enzyme inhibition with Fe2+ and H2O2 was less pronounced in ischemia. The inhibitory effect of the reaction product 2-oxoglutarate was higher under normal conditions than in ischemia (the Ki values were 0.22 and 0.75 mM, respectively). The specific features of the NADP-IDH regulation in ischemia are suggested to promote the stimulation of the enzyme functioning during increased level of free radical processes, and this seems to be important for NADPH supplying for the glutathione reductase/glutathione peroxidase antioxidant system of cardiomyocytes. PMID- 12126480 TI - DNA aptamers as radically new recognition elements for biosensors. PMID- 12126482 TI - Priority setting for new technologies in medicine: a transdisciplinary study. AB - BACKGROUND: Decision makers in health care organizations struggle with how to set priorities for new technologies in medicine. Traditional approaches to priority setting for new technologies in medicine are insufficient and there is no widely accepted model that can guide decision makers. DISCUSSION: Daniels and Sabin have developed an ethically based account about how priority setting decisions should be made. We have developed an empirically based account of how priority setting decisions are made. In this paper, we integrate these two accounts into a transdisciplinary model of priority setting for new technologies in medicine that is both ethically and empirically based. SUMMARY: We have developed a transdisciplinary model of priority setting that provides guidance to decision makers that they can operationalize to help address priority setting problems in their institution. PMID- 12126481 TI - Growth inhibition by the muscarinic M(3) acetylcholine receptor: evidence for p21(Cip1/Waf1) involvement in G(1) arrest. AB - We have assessed the growth response of Chinese-hamster ovary (CHO) cells to activation of recombinantly expressed G-protein-coupled muscarinic M(2) or M(3) acetylcholine receptors (AChRs). We show that activation of these receptors leads to divergent growth responses: M(2) AChR activation causes an increase in DNA synthesis, whereas M(3) AChR activation causes a dramatic decrease in DNA synthesis. We have characterized the M(3) AChR-mediated growth inhibition and show that it involves a G(1) phase cell-cycle arrest. Further analysis of this arrest indicates that it involves an increase in expression of the cyclin dependent kinase (CDK) inhibitor, p21(Cip1/Waf1) (where Cip1 is CDK-interacting protein 1 and Waf1 is wild-type p53-associated fragment 1), in response to M(3) AChR activation. This increase in protein expression leads to an increase in p21(Cip1/Waf1) association with CDK2, a decrease in CDK2 activity and an accumulation of hypophosphorylated retinoblastoma protein. The increased p21(Cip1/Waf1) expression is due, at least in part, to an increase in p21(Cip1/Waf1) mRNA, and receptor-mediated changes in phosphorylation of c-Jun provide a mechanism to account for this p21(Cip1/Waf1) transcriptional regulation. Evaluation of the extracellular signal-regulated protein kinase and c Jun N-terminal kinase activities has shown striking differences in the profiles of activation of these mitogen-activated protein kinases by the M(2) and M(3) AChRs, and their potential involvement in mediating growth arrest by the M(3) AChR is discussed. PMID- 12126483 TI - Signal and noise in bridging PCR. AB - BACKGROUND: In a variant of the standard PCR reaction termed bridging, or jumping, PCR the primer-bound sequences are originally on separate template molecules. Bridging can occur if, and only if, the templates contain a region of sequence similarity. A 3' end of synthesis in one round of synthesis that terminates in this region of similarity can prime on the other. In principle, Bridging PCR (BPCR) can detect a subpopulation of one template that terminates synthesis in the region of sequence shared by the other template. This study considers the sensitivity and noise of BPCR as a quantitative assay for backbone interruptions. Bridging synthesis is also important to some methods for computing with DNA. RESULTS: In this study, BPCR was tested over a 328 base pair segment of the E. coli lac operon and a signal to noise ratio (S/N) of approximately 10 was obtained under normal PCR conditions with Taq polymerase. With special precautions in the case of Taq or by using the Stoffel fragment the S/N was improved to 100, i.e. 1 part of cut input DNA yielded the same output as 100 parts of intact input DNA. CONCLUSIONS: In the E. coli lac operator region studied here, depending on details of protocol, between 3 and 30% per kilobase of final PCR product resulted from bridging. Other systems are expected to differ in the proportion of product that is bridged consequent to PCR protocol and the sequence analyzed. In many cases physical bridging during PCR will have no informational consequence because the bridged templates are of identical sequence, but in a number of special cases bridging creates, or, destroys, information. PMID- 12126485 TI - Prescribing to drug misusers in practice--often effective, but rarely straightforward. AB - Many reviews describe the effectiveness of methadone treatment in reducing illicit drug use and associated behaviours among opiate misusers. The strongest evidence includes social outcomes such as reduced debt and crime, and relates overwhelmingly to maintenance rather than detoxification treatment. Drug clinics are often dominated by individuals unable to withdraw fully from methadone, while the "harm reduction" model accepts some ongoing drug use, with attendant risks. Security measures are necessary to avoid abuse of treatments, but these may be undermined by the agenda of "partnerships with patients" in decision-making. Buprenorphine appears both safer and less addictive than methadone, and lofexidine is effective as a non-substitute detoxification method. Naltrexone can clearly reduce relapse rates, provided consumption is assured, while for individuals unable to detoxify or avoid euphoriant opiates, morphine and diamorphine are sometimes used. In non-opiate misuse, clinical studies of a wide range of medications have produced relatively few positive findings. PMID- 12126484 TI - The identity and distribution of neural cells expressing the mesodermal determinant spadetail. AB - BACKGROUND: The spadetail (spt) gene of zebrafish is expressed in presomitic mesoderm and in neural cells previously suggested to be Rohon-Beard neurons. The mechanism(s) generating the apparently irregular rostrocaudal distribution of spt expressing cells in the developing CNS is unknown. RESULTS: spt-expressing neural cells co-express huC, a marker of neurons. These cells also co-express the genes islet-1, -2 and -3 but not valentino. The islet-1 gene expression, irregular distribution and dorsolateral position of spt-expressing cells in the developing CNS are characteristic of dorsal longitudinal ascending (DoLA) interneurons. Shortly after their birth, these neurons extend processes rostrally into which spt mRNA is transported. At 24 hours post fertilisation(hpf), spt-expressing neurons occur most frequently at rostral levels caudal of the 5th-formed somite pair. There is no apparent bias in the number of spt-expressing cells on the left or right sides of embryos. Extended staining for spt-transcription reveals expression in the dorsocaudal cells of somites at the same dorsoventral level as the spt-expressing neurons. There is frequent juxtaposition of spt-expression in newly formed somites and in neurons. This suggests that both types of spt expressing cell respond to a common positional cue or that neurons expressing spt are patterned irregularly by flanking somitic mesoderm. CONCLUSIONS: spt expressing cells in the developing CNS appear to be DoLA interneurons. The irregular distribution of these cells along the rostrocaudal axis of the spinal cord may be due to "inefficient" patterning of neural spt expression by a signal(s) from flanking, regularly distributed somites also expressing spt. PMID- 12126486 TI - Effect of a dose of ethanol on acute tolerance and ethanol consumption in alcohol drinker(UChB) and non-drinker (UChA) rats. AB - Acute tolerance that develops within minutes of ethanol exposure appears to influence the apparent acute behavioral sensitivity of laboratory animals to ethanol actions. The existence of a correlation between voluntary ethanol consumption and the speed of acquiring acute tolerance has been proposed. In the present paper we investigated the effect of an acute dose of ethanol on tolerance development and on ethanol voluntary consumption in our two selected bred strains, UChA (low ethanol drinker) and UChB (high ethanol drinker) rats. Acute tolerance developed to motor impairment induced by a dose of ethanol of 2.3 g/kg. administered intraperitoneally was evaluated by the tilting plane test. Voluntary ethanol consumption was compared in rats receiving the ethanol dose, to rats receiving a saline intraperitoneal (i.p.) injection. The results show that UChB rats receiving an intoxicating dose of ethanol develop more tolerance and they significantly increased their ethanol consumption compared to the same line that received a saline injection, while no change in acute tolerance and voluntary ethanol consumption were obtained in UChA rats. In conclusion, a possible mechanism by which UChB rats drink high amounts of ethanol appears to be the development of tolerance to the pharmacological effects of ethanol. PMID- 12126487 TI - Does paradoxical sleep deprivation and cocaine induce penile erection and ejaculation in old rats? AB - A recent study has established that paradoxical sleep deprivation (PSD) and cocaine administration elicit genital reflexes (penile erection and ejaculation) in young rats. To discover whether the same effects occurred in old animals submitted to PSD, we administered cocaine (15 mg/kg) to young (3-month) and old (22-month) male rats after a 4-day period of PSD or at the equivalent time-point in control animals. We then evaluated erections and ejaculations. Sixty per cent of the old-PSD group displayed erection, although ejaculation was not observed. Genital reflexes were absent in young and old control groups. We found that PSD reduced testosterone and increased progesterone levels in both young and old PSD groups. In conclusion, our results suggest that although genital reflexes usually decrease with age, testosterone levels alone cannot account for these changes. The interaction of PSD and cocaine probably enhances dopamine transmission in the brain and may elicit penile erection in old rats. PMID- 12126488 TI - Differential propensity to ethanol sensitization is not associated with altered binding to D1 receptors or dopamine transporters in mouse brain. AB - Behavioral sensitization to ethanol's stimulant effect has been proposed as a marker for individual abuse liability. In previous work we have demonstrated that mice showing an increased propensity to EtOH sensitization had higher levels of dopamine (DA) D2 receptor binding in localized brain areas compared to mice showing less sensitization. In the present study we examined whether altered binding to D1 or the DA transporter (DAT) might also be associated with differential propensity to develop EtOH sensitization. Male Swiss mice received 2.4 g/kg EtOH or saline intraperitoneally (i.p.) daily for 21 days, were tested weekly for locomotor activity, and then sacrificed. D1 and DAT binding were assessed by quantitative autoradiography using [(3)H]SCH-23390 and [(3)H]WIN 35,428, respectively. EtOH-treated mice were subdivided into sensitized and non sensitized subgroups according to their locomotor activity during treatment. Analyses of brain D1 (19 regions) and DAT (12 regions) binding densities revealed no significant differences among EtOH-sensitized, -non-sensitized or saline groups in any of the regions measured (all p values > 0.32 for D1 and > 0.16 for DAT). These results suggest that brain D1 and DAT binding, unlike the recently reported changes in D2 binding, do not differentiate mice that develop behavioral sensitization to ethanol from those that do not. PMID- 12126489 TI - A specific anti-aggressive effect of repeatedly administered lobeline. AB - The effects of chronic treatments with nicotinic agonists on agonistic encounters have received little attention. The effects of repeated (for 10 days) SC administration of (-)-lobeline (9.3, 18.6 and 37.2 micromol/kg) and (-)-nicotine (0.93, 1.86 and 3.72 micromol/kg) were evaluated using the mouse isolation induced aggression model. Individually housed OF1 male mice served as experimental animals and were confronted by 'standard opponents'. Each mouse was tested only once on the last day of the repeated drug treatment. Videotaped agonistic encounters were analysed estimating the times allocated to 11 behavioural categories. Repeated treatment with the highest dose of lobeline diminished attack behaviour without significantly increasing immobility or changing any other behavioural category involving motor activity. In contrast, nicotine did not significantly alter time allocated to any behavioural category. PMID- 12126490 TI - Green tea as a potent antioxidant in alcohol intoxication. AB - Ethanol oxidation to acetaldehyde and next to acetate is accompanied by free radical generation. Free radicals can affect cell integrity when antioxidant mechanisms are no longer able to cope with the free radical generation observed in ethanol intoxication. Natural antioxidants are particularly useful in such a situation. The present study was designed to investigate the efficacy of green tea as a source of water-soluble antioxidants (catechins) on the liver and blood serum antioxidative potential of rats chronically (28 days) intoxicated with ethanol. Alcohol caused a decrease in liver superoxide dismutase, glutathione peroxidase and catalase activities and an increase in activity of glutathione reductase. Moreover, a decrease in the level of reduced glutathione, ascorbic acid, vitamins A and E and beta-carotene were observed. The activity of serum glutathione peroxidase decreased while glutathione reductase activity increased. The level of serum non-enzymatic antioxidants was also decreased in the liver. Alcohol administration caused an increase in the liver and serum lipid peroxidation products, measured as thiobarbituric acid-reactive substances. However, green tea prevents the changes observed after ethanol intoxication. Green tea also protects membrane phospholipids from enhanced peroxidation. These results indicate a beneficial effect of green tea in alcohol intoxication. PMID- 12126491 TI - T-wave response: a sensitive test for latent alcoholic polyneuropathy. AB - To date, the H-reflex is the most sensitive test to measure nerve conduction velocity in alcoholic polyneuropathy. Analogous to the H-reflex, we investigated the T-wave response from the soleus muscle using a hand-held reflex hammer. Twenty-four inpatients suffering from chronic alcoholism and 24 healthy volunteers were recruited. All probands had a careful neurological examination and were graded (PNP-classifications). The T- and H-reflexes were measured. In the clinical examination, only a few patients exhibited symptoms of alcoholic PNP. However, when the autonomic nervous system was also tested, 50% exhibited signs of alcoholic PNP. Both the T- and H-reflex responses were pathologically retarded, indicating latent alcoholic PNP in 60% of the patients. Thus the main finding in our study is the difference between clinical and electrophysiological examinations: only a few of the patients had neurological symptoms for alcoholic PNP but 14 patients (60%) exhibited a so-called latent, subclinical alcoholic PNP by showing delayed reflex latencies. Measuring the T-wave proved to be a simple and painless screening method for diagnosis and monitoring of alcoholic PNP. Among the clinical tests the best indicator for alcoholic PNP was the test for autonomous alcoholic PNP. PMID- 12126492 TI - Serum naltrexone and 6-beta-naltrexol levels from naltrexone implants can block very large amounts of heroin: a report of two cases. AB - The maximum dose of heroin that is blocked by customary doses of oral naltrexone (NTX) and its active metabolite 6-beta-naltrexol (6BNT) is unknown, particularly at trough serum levels which show much interindividual variation and can be low. NTX has only once been tested formally against opiate equivalents of more than 25 mg of diamorphine. Increasing interest in long-acting implantable NTX preparations makes it important to have objective information about blocking activity at various blood levels of NTX and 6BNT. Two cases are described in which NTX and 6BNT levels as low as 2.8 and 9.0 ng/ml, respectively, were sufficient to block doses of pure diamorphine as high as 500 mg. PMID- 12126493 TI - Bromocriptine use is associated with decreased smoking rates. AB - Dopaminergic transmission in the central nervous system is thought to underlie addictive behaviours, including smoking. One effective smoking cessation drug, bupropion, enhances dopaminergic transmission; conversely, antipsychotic drugs, which are dopamine antagonists, are associated with increased smoking. Thus we hypothesized that subfertile women treated with the potent dopamine agonist bromocriptine might smoke less as a consequence of their treatment. Among 4,608 subfertile women those conceiving on bromocriptine were half as likely to smoke as those taking other drugs or those conceiving without medication (p < 0.0001). This observation supports the role of dopamine in nicotine addiction, and suggests that bromocriptine-like drugs could be used effectively by pregnant women to aid cessation. PMID- 12126495 TI - Figure-ground segregation in a recurrent network architecture. AB - Here we propose a model of how the visual brain segregates textured scenes into figures and background. During texture segregation, locations where the properties of texture elements change abruptly are assigned to boundaries, whereas image regions that are relatively homogeneous are grouped together. Boundary detection and grouping of image regions require different connection schemes, which are accommodated in a single network architecture by implementing them in different layers. As a result, all units carry signals related to boundary detection as well as grouping of image regions, in accordance with cortical physiology. Boundaries yield an early enhancement of network responses, but at a later point, an entire figural region is grouped together, because units that respond to it are labeled with enhanced activity. The model predicts which image regions are preferentially perceived as figure or as background and reproduces the spatio-temporal profile of neuronal activity in the visual cortex during texture segregation in intact animals, as well as in animals with cortical lesions. PMID- 12126496 TI - A neural model of perceptual-motor alignment. AB - Sensorimotor systems face complex and frequent discrepancies among spatial modalities, for example, growth, optical distortion, and telemanipulation. Adaptive mechanisms must act continuously to restore perceptual-motor alignments necessary for perception of a coherent world. Experimental manipulations that exposed participants to localized discrepancies showed that adaptation is revealed by the acquisition of a constrained relation between entire modalities rather than associations between individual exemplars within these modalities. The computational problem faced by the human nervous system can thus be conceived as having to induce constrained relations between continuous stimulus and response dimensions from ambiguous or incomplete training sets, that is, performing interpolation and extrapolation. How biological neuronal networks solve this problem is unknown. Here we show that neural processing based on linear collective computation and least-square (LS) error learning in populations of frequency-coded neurons (i.e., whose discharge varies in a monotonic fashion with a parameter) has built-in interpolation and extrapolation capacities. This model can account for the properties of perceptual-motor adaptations in sensorimotor systems. PMID- 12126497 TI - The response of left temporal cortex to sentences. AB - The meaning of a sentence differs from the sum of the meanings of its constituents. Left anterior temporal cortex responds to sentences more strongly than to unconnected words. We hypothesized that the anterior temporal response to sentences is due to this difference in meaning (compositional semantics). Using positron emission tomography (PET), we studied four experimental conditions (2 x 2 factorial design): In one condition, subjects read normal sentences. In a second condition, they read grammatically correct sentences containing numerous semantic violations (semantically random sentences). In a third condition, we scrambled the word order within the normal sentences, and, in a fourth condition, the word order was scrambled within the semantically random sentences. The left anterior temporal pole responded strongly to sentences compared to scrambled versions of sentences. A similar although weaker response occurred in the left anterior superior temporal sulcus and the left posterior middle temporal gyrus. A subset of voxels within the left anterior temporal pole responded more to semantically random sentences and their scrambled versions than to normal sentences and the corresponding scrambled versions (main effect of semantic randomness). Finally, the grammatical and the semantic factor interacted in a subset of voxels within the anterior temporal pole: Activity was higher when subjects read normal sentences compared to their scrambled versions but not for semantically random sentences compared to their corresponding scrambled versions. The effects of grammar and meaning and, most importantly, the interaction between grammatical and semantic factors are compatible with the hypothesis that the left anterior temporal pole contributes to the composition of sentence meaning. PMID- 12126498 TI - Neural differentiation of lexico-syntactic categories or semantic features? event related potential evidence for both. AB - Event-related potentials (ERPs) were used to investigate whether processing differences between nouns and verbs can be accounted for by the differential salience of visual-perceptual and motor attributes in their semantic specifications. Three subclasses of nouns and verbs were selected, which differed in their semantic attribute composition (abstract, high visual, high visual and motor). Single visual word presentation with a recognition memory task was used. While multiple robust and parallel ERP effects were observed for both grammatical class and attribute type, there were no interactions between these. This pattern of effects provides support for lexical-semantic knowledge being organized in a manner that takes account both of category-based (grammatical class) and attribute-based distinctions. PMID- 12126499 TI - Perceptual priming versus explicit memory: dissociable neural correlates at encoding. AB - We addressed the hypothesis that perceptual priming and explicit memory have distinct neural correlates at encoding. Event-related potentials (ERPs) were recorded while participants studied visually presented words at deep versus shallow levels of processing (LOPs). The ERPs were sorted by whether or not participants later used studied words as completions to three-letter word stems in an intentional memory test, and by whether or not they indicated that these completions were remembered from the study list. Study trials from which words were later used and not remembered (primed trials) and study trials from which words were later used and remembered (remembered trials) were compared to study trials from which words were later not used (forgotten trials), in order to measure the ERP difference associated with later memory (DM effect). Primed trials involved an early (200-450 msec) centroparietal negative-going DM effect. Remembered trials involved a late (900-1200 msec) right frontal, positive-going DM effect regardless of LOP, as well as an earlier (600-800 msec) central, positive-going DM effect during shallow study processing only. All three DM effects differed topographically, and, in terms of their onset or duration, from the extended (600-1200 msec) fronto-central, positive-going shift for deep compared with shallow study processing. The results provide the first clear evidence that perceptual priming and explicit memory have distinct neural correlates at encoding, consistent with Tulving and Schacter's (1990) distinction between brain systems concerned with perceptual representation versus semantic and episodic memory. They also shed additional light on encoding processes associated with later explicit memory, by suggesting that brain processes influenced by LOP set the stage for other, at least partially separable, brain processes that are more directly related to encoding success. PMID- 12126500 TI - The timing of action-monitoring processes in the anterior cingulate cortex. AB - The anterior cingulate cortex (ACC) has been shown to respond to conflict between simultaneously active, incompatible response tendencies. This area is active during high-conflict correct trials and also when participants make errors. Here, we use the temporal resolution of high-density event-related potentials (ERPs) in combination with source localization to investigate the timing of ACC activity during conflict and error detection. We predicted that the same area of the ACC is active prior to high-conflict correct responses and following erroneous responses. Dipole modeling supported this prediction: The frontocentral N2, occurring prior to the response on correct conflict trials, and the ERN, occurring immediately following error responses, could both be modeled as having a generator in the caudal ACC, suggesting the same process to underlie both peaks. Modeling further suggested that the rostral area of the ACC was also active following errors, but later in time, contributing to the error positivity (P(E)), and peaking at 200-250 msec following the ERN peak. Despite the inherent limitations of source localization, these data may begin to shed light on the timing of action-monitoring processes. First, the time course of caudal ACC activity follows the time course as predicted by the conflict theory of this region. Second, caudal ACC activity might be temporally dissociated from rostral ACC activity during error trials, which possibly reflects a separate, affective component of the evaluative functions of the ACC. PMID- 12126501 TI - Abnormal auditory cortical activation in dyslexia 100 msec after speech onset. AB - Reading difficulties are associated with problems in processing and manipulating speech sounds. Dyslexic individuals seem to have, for instance, difficulties in perceiving the length and identity of consonants. Using magnetoencephalography (MEG), we characterized the spatio-temporal pattern of auditory cortical activation in dyslexia evoked by three types of natural bisyllabic pseudowords (/ata/, /atta/, and /a a/), complex nonspeech sound pairs (corresponding to /atta/ and /a a/) and simple 1-kHz tones. The most robust difference between dyslexic and non-reading-impaired adults was seen in the left supratemporal auditory cortex 100 msec after the onset of the vowel /a/. This N100m response was abnormally strong in dyslexic individuals. For the complex nonspeech sounds and tone, the N100m response amplitudes were similar in dyslexic and nonimpaired individuals. The responses evoked by syllable /ta/ of the pseudoword /atta/ also showed modest latency differences between the two subject groups. The responses evoked by the corresponding nonspeech sounds did not differ between the two subject groups. Further, when the initial formant transition, that is, the consonant, was removed from the syllable /ta/, the N100m latency was normal in dyslexic individuals. Thus, it appears that dyslexia is reflected as abnormal activation of the auditory cortex already 100 msec after speech onset, manifested as abnormal response strengths for natural speech and as delays for speech sounds containing rapid frequency transition. These differences between the dyslexic and nonimpaired individuals also imply that the N100m response codes stimulus specific features likely to be critical for speech perception. Which features of speech (or nonspeech stimuli) are critical in eliciting the abnormally strong N100m response in dyslexic individuals should be resolved in future studies. PMID- 12126502 TI - The neural correlates of grammatical gender: an fMRI investigation. AB - In an fMRI experiment, subjects saw a written noun and made three distinct decisions in separate sessions: Is its grammatical gender masculine or feminine (grammatical feature task)? Is it an animal or an artifact (semantic task)? Does it contain a /tch/ or a /k/ sound (phonological task)? Relative to the other experimental conditions, the grammatical feature task activated areas of the left middle and inferior frontal gyrus and of the left middle and inferior temporal gyrus. These activations fit in well with neuropsychological studies that document the correlation between left frontal lesions and damage to morphological processes in agrammatism, and the correlation between left temporal lesions and failure to access lexical representations in anomia. Taken together, these data suggest that grammatical gender is processed in a left frontotemporal network. In addition, the observation that the grammatical feature task and the phonology task activated neighboring but distinct regions of the left frontal lobe provides a plausible neuroanatomical basis for the systematic occurrence of phonological errors in aphasic subjects with morphological deficits. PMID- 12126503 TI - Visualizing the neural bases of a disconnection syndrome with diffusion tensor imaging. AB - Disconnection syndromes are often conceptualized exclusively within cognitive box and-arrow diagrams unrelated to brain anatomy. In a patient with alexia in his left visual field resulting from a posterior callosal lesion, we illustrate how diffusion tensor imaging can reveal the anatomical bases of a disconnection syndrome by tracking the degeneration of neural pathways and relating it to impaired fMRI activations and behavior. Compared to controls, an abnormal pattern of brain activity was observed in the patient during word reading, with a lack of activation of the left visual word form area (VWFA) by left hemifield words. Statistical analyses of diffusion images revealed a damaged fiber tract linking the left ventral occipito-temporal region to its right homolog across the lesioned area of corpus callosum and stopping close to the areas found active in fMRI. The behavioral disconnection syndrome could, thus, be related functionally to abnormal fMRI activations and anatomically to the absence of a connection between those activations. The present approach, based on the "negative tracking" of degenerated bundles, provides new perspectives on the understanding of human brain connections and disconnections. PMID- 12126504 TI - Covert auditory attention generates activation in the rostral/dorsal anterior cingulate cortex. AB - The anterior cingulate cortex (ACC) is believed to mediate conscious information processing or high-capacity attention. However, previous functional imaging studies have largely relied on tasks that involve motor function as well as attention. The work from our group utilizing an auditory continuous performance task demonstrated increased activity in a caudal division of the ACC that borders the supplementary motor area (SMA). Activity in this region was attributed to motor responding as well as attention. In the present study, we used (15)O H(2)O positron emission tomography (PET) to map brain activation during nonmotor, covert auditory attention. Our hypothesis was that a different region within the ACC, anterior to the SMA, would be active during covert attention (CA). Six men and six women were asked to monitor aurally presented syllables presented at a 1 sec interstimulus interval. During the CA condition, subjects were asked to continuously discriminate target (.19 probability) from nontarget stimuli. Simultaneous recording of event-related potentials (ERPs) confirmed the discrimination of target and nontarget stimuli and the allocation of attention capacity. Comparison of the monitored versus nonmonitored presentation of stimuli demonstrated significant activity in a rostral/dorsal division of the right ACC, anterior to SMA. Other regions of activation included the lateral prefrontal cortex and posterior superior temporal gyrus in the left hemisphere, consistent with neurocognitive models of language and vigilance. We conclude that a rostral/dorsal subdivision of the right ACC is specific for conscious attention during auditory processing, in contrast to premotor response formation. PMID- 12126505 TI - The quantitative nature of a visual task differentiates between ventral and dorsal stream. AB - The aim of the present positron emission tomography (PET) study was to investigate how visual processing in dorsal and ventral streams depends on the quantitative nature of the task. In the same-different task, participants identified the presence of an orientation difference between two gratings, presented centrally in succession. In the quantification task, participants estimated the magnitude of the difference and compared it to a fixed standard. Detection of dimming of the fixation point was used as a control task. Visual input, motor responses, and performance were equated across tasks. Subtracting same-different from quantification yielded significant activation in the left superior parietal lobule and left ventral premotor cortex, consistent with results obtained in number-processing tasks. The reverse subtraction yielded activation in the right inferior temporal gyrus, in agreement with earlier results. These results demonstrate that a single attribute can be processed either in the ventral or dorsal stream, depending on the cognitive operations required by the tasks. PMID- 12126506 TI - Sustained mnemonic response in the human middle frontal gyrus during on-line storage of spatial memoranda. AB - The mapping of cognitive functions to neural systems is a central goal of cognitive neuroscience. On the basis of homology with lesion and physiological studies in nonhuman primates, Brodmann's area (BA) 46/9 in the middle frontal gyrus (MFG) has been proposed as the cortical focus for both the storage as well as processing components of working memory in the human brain, but the evidence on the segregation of these components and their exact areal localization has been inconsistent. In order to study this issue and increase the temporal resolution of functional mapping, we disambiguated the storage component of working memory from sensory and motor responses by employing functional magnetic resonance imaging (fMRI) in spatial delayed-response (DR) tasks with long delay intervals and different conditions of demand. We here show that BA 46 can support a sustained mnemonic response for as long as 24 sec in a high-demand task and the signal change in this area exceeded that in the other prefrontal areas examined. Our findings support a conservation of functional architecture between human and nonhuman primate in showing that the MFG is prominently engaged in the storage of spatial information. PMID- 12126507 TI - [Association between Helicobacter pylori cagA strain infection and expression of cyclooxygenase 2 in gastric carcinoma]. AB - OBJECTIVE: To observe the expression of cyclooxygenase 2 (COX-2) in tissues of human gastric cancer and explore the association between Helicobacter pylori cagA strain infection and the expression of COX-2 so as to provide a theoretical basis for early prevention of gastric cancer. METHODS: Flow cytometry was adopted to quantitatively determine and analyze the expression of COX-2 in 31 specimens of gastric cancer and normal juxta cancerous tissues. PCR was used to detect the cagA gene in gastric cancer. RESULTS: COX-2 was over-expressed in 26 of the 31 (84%) specimens of gastric cancer. The expression of COX-2 in the 18 specimens of gastric cancer with cagA positive strain infection was significantly higher than that in the 13 specimens with cagA negative strain infection. CONCLUSION: COX-2 is over-expressed in gastric cancer and cagA positive strain infection up regulates the expression of COX-2 in gastric cancer. There may be another way or channel to regulate the expression of COX-2 in gastric cancer besides cagA positive strain infection. Therefore, applying COX-2 selective inhibitors to prevent gastric cancer can be an effective and promising way. PMID- 12126508 TI - [The difference of the bismuth absorption from a single colloidal bismuth pectin therapy and quadruple therapy for eradicating Helicobacter pylori infection]. AB - OBJECTIVE: To investigate whether the quadruple therapy influences the absorption of bismuth from Colloidal Bismuth Pectin (CBP). METHODS: 24 male Wistar rats weighing from 400 g to 500 g were randomly divided into two groups: CBP group (group A(1)) and CBP+AMO+MTR+ LAN group (group A(2)). The serum bismuth concentration was observed at 0,1,2,3,4,5,6 hour after these medicines were fed. Other 24 male Wistar rats weighing from 200 g to 250 g were randomly divided into two groups: CBP group (group B( 1)), CBP+AMO+MTR+LAN group (group B(2)). These medicines had been taken every day for 14 days. The accumulation of bismuth in the heart, brain, kidney and liver of rats were detected eight weeks later. RESULTS: The serum peak value of bismuth is not significantly different between group A(1) and group A(2) (P > 0.05), but the peak time of bismuth of the group A(2) is one hour earlier than that of group A(1). The amounts of the accumulation of bismuth in the kidney are 154 +/- 15 ng/g and 212 +/- 20 ng/g in the group B(1) and B(2) respectively at the eighth week after the medicines were fed in rats with a significant difference (P < 0.05). CONCLUSION: The quadruple therapy causes an increase of the absorption and the accumulation of bismuth from CBP in the kidney in rats. PMID- 12126509 TI - [Distribution characteristics of types of stroke in three urban area in China]. AB - OBJECTIVE: To analyze the distribution characteristics of types of stroke in china. METHODS: A prospective study and comprehensive prevention were conducted in urban communities with 300 000 people in Changsha, Beijing and Shanghai, and 2 570 case of stroke at their first occurance were selected with established typing diagnosis (r). RESULTS: Hemmorrhagic, ischemic, and unclassified stroke accounted for 37.1%, 62.3%, and 0.6% respectively. The proportion of hemmorrhagic stroke was more than 50 percent among persons under 50. The average yearly incidence of stroke was 142.3/100 thousand in the intervention areas and was 187.0/100 thousand in the control areas. CONCLUSION: Prevention should be focused on hemorrhagic stroke for those aged over 50 and on ischemic stroke for those aged less than 50. The average incidence rates of hemmorrhagic and of ischemic stroke were lower in the intervention area than in the control area. PMID- 12126510 TI - [Modification and application of anterior skull base microsurgery with navigation system]. AB - OBJECTIVE: To investigate the accuracy and safety of the modified techniques in anterior skull base microsurgery with navigation system. METHODS: Operation was performed upon 42 patients with benign and malignant tumor of anterior skull base involving both cerebral cranium and visceral cranium with the help of modified navigation system. A new arrangement was designed for stable maintenance of the headset with dynamic reference frame to avoid disturbance during emission of encoded infrared signal. Six reference points were used, distributed in the parietal, frontal, zygomatic, maxillary, and mastoid regions. The STN navigation system and SMN surgical microscope navigator were used to provide virtual pointer, virtual approach route and virtual contour as an orientation for stereotaxic guidance during surgery. RESULTS: The modified technique could keep the accuracy of surgical guidance within 1 approximately 1.5 mm in comparison with the accuracy of 4 mm or more by the original design, thus guaranteeing the safety of surgery. CONCLUSION: The modified techniques raise the accuracy and safety of anterior skull base microsurgery. However, the cost remains high. PMID- 12126511 TI - [Comparative study of laparoscopically assisted myomectomy and mini- laparotomy for uterine intramural fibroids]. AB - OBJECTIVE: To evaluate the feasibility and safety of laparoscopically assisted myomectomy (LAM) for uterine intramural fibroids. METHODS: Thirty-seven selected patients with uterine intramural fibroids with the diameters > 5 cm and < 9 cm were subjected to LAM technique, the intramural fibroid of 2 among which penetrated into the uterine cavity (< 50%). LAM was completed with a laparoscopically assisted enucleation. After the fibroid was half-scripped, the incision of puncture point at the middle of abdominal wall was expanded to 4 approximately 5 cm in length. The scripping of the fibroid and the suture of the uterine wound were completed outside the abdominal incision. The duration of operation, blood loss, intra- and post-operation complications and time to full recovery were evaluated and comparison with those in 630 patients with hysteromyoama who underwent myomectomy by laparotomy during the same period. RESULTS: The operative time in LAM group ranged from 50 min to 240 min (101 min +/- 56 min), without a significant difference in comparison with that in the group of myomectomy by laparotomy (89 min +/- 38 min, P > 0.05). However, the median of blood loss in LAM group was 50 ml, significantly less than that in the group of myomectomy by laparotomy (80 ml, P < 0.05). The incidence of pyrexia was lower and the time needed for recovery was shorter after LAM in comparison with those in the group of myomectomy by laparotomy (P < 0.001). Neither intra operative nor post- operative complication was observed in the LAM group. Eight of the 11 cases with infertility in the LAM group were conceived at least one year after LAM. The pregnancy was uneventful and proceeded to cesarean section at the 37 approximately 38 th week. CONCLUSION: LAM operation is feasible and safe, and offers a easy-to-perform minimally invasive myomectomy technique for intramural fibroid removal with all the advantages of laparoscopic surgery and a good reconstructive outcome. PMID- 12126512 TI - [Functions of cultured dendritic cells from patients with chronic hepatitis Beta decreased]. AB - OBJECTIVE: To compare the phenotype and function of dendritic cells (DCs) of patients with chronic hepatitis Beta with those of DCs from healthy persons. METHOD: Peripheral blood monocytes (PBMCs) were isolated from 10 patients with chronic hepatitis Beta and 10 healthy blood donors. DCs were generated by culturing PBMCs in 1 000 U/ml granulocyte-macrophage stimulating factor (GM-CSF) and 500 U/ml IL-4 with AIM-V in vitro. DCs and supernatant were collected 3, 5, 7, 9, 11, 13 and 15 days after culture. TNF- alpha 50 ng/ml was added into the culture of DCs from patients with hepatitis Beta and DCs were collected 48 hours after. FACS was used to detect the phenotype. The changes of cell phenotype on different days with or without TNF-alpha were tested by immunolabelling and flow cytometry analysis. IL-12 ELISA kit was applied to investigate IL-12 secretion of DCs. To perform mixed leucocyte reaction (MLR) test, DCs were cocultured with allogeneic T cells at different stimulatory ratio. RESULTS: The expression of CD83 was higher in the group with TNF-alpha than in the group without TNF-alpha. The positive rates of CD80 and CD54 were not different between the two groups. DCs from patients and normal donors with typical morphology were harvested successfully in vitro. The IL-12 level was 103.93 +/- 12.59 pg/ml in the group with TNF-alpha, higher than that in the group without TNF-alpha (11.45 +/- 2.58 pg/ml, P < 0.05). MLR test showed that when the stimulatory ratio of allogeneic T cells was 1 : 20, 1 : 50, and 1 : 100 respectively, the results were 40 835 +/- 3 480, 37 525 +/- 6 398, and 31 814 +/- 5 521 respectively in group of patient; and 49 097 +/- 2 273, 43 049 +/- 2 833, and 39 091 +/- 6 469 respectively in the group of healthy blood donors. CONCLUSION: 1.DCs of patients and of healthy persons can be cultured success fully with AIM-V when induced by GM-CSF, IL-4 and TNF-alpha in vitro. 2. Functions of DCs from patients are lower than those from healthy donors. However, they are not different in amount and morphology. PMID- 12126513 TI - [Clinical and polysomnographic features of rapid eye movement sleep behavior disorder]. AB - OBJECTIVE: To evaluate the clinical and polysomnographic features of rapid eye movement sleep behavior disorder (RBD) in Chinese patients. METHODS: Six parasomnic patients, 4 males and 2 females, with the mean onset age of 58 years (range 50 approximately 66 years) and 6 age and sex matched controls, were video monitored for successive two nights, to record the electroencephalography, electrooculography, electromyography, electrocardiography and nasal airflow. RESULTS: Polysomnographic recordings disclosed an augmented muscle tone, which appeared intermittently or continuously in REM sleep, accompanied by complex behaviors correlated to dream contents observed by video-monitoring, and without seizure activity. All the 6 patients presented with a long history of parasomnia. The average total-sleep time of the patients was 353 +/- 42 minutes, the stage II sleep time was 139 +/- 76 minutes, and sleep efficiency was 74.3% +/- 12.3% lower compared with that of the controls. Pakinsonism occurred in 1 patient 9 years after parasomnia and dementia in 2 patients 8 and 18 years respectively after symptom of RBD. Clonazepam was administered for parasomnia in 3 cases with a favorable response. CONCLUSION: REM sleep without atonia is demonstrated in association with violent movements by video-monitored polysomnography in 6 Chinese patients. PMID- 12126514 TI - [The expression of human specific proteins in liver tissue of chimeric goats engrafted with human hematopoitic stem cells]. AB - OBJECTIVE: To investigate the expression of human specific proteins in liver tissue obtained from goats engrafted with human hematopoietic stem cells (hHSC). METHODS: hHSCs derived from cord blood were transplanted into fetal goats for production of chimerism. Liver specimens were obtained from four 10-month-old chimeric goats and examined for their reactivity with monoclonal antibodies against human antigens: proliferating cell nuclear antigen (PCNA) and hepatocyte specific antigen (HSA). The presence of human HSA positive cells in the goat liver tissue was determined by fluorescence assisted cell sorting (FACS). The expression of human hepatocyte nuclear factor (hHNF-3beta) and human serum albumin (hALB) mRNAs was determined by RT-PCR. Meanwhile, FISH experiment was also performed to detect the human cells in the chimeric livers with the probe of human p17H8. RESULTS: Successful engraftment was confirmed by the detection of human blood cell chimerism in the circulation of the transplanted goats. Human PCNA and HSA were found in liver specimens of transplanted goats but not of normals. FACS analysis showed the presence of human HSA positive cells in the liver of chimeric goats. RT-PCR results demonstrated that hHNF-3beta and hALB mRNAs were specifically expressed in liver tissues of chimeric goats. FISH experiment showed two positive hybridized signals in some liver cells of the chimeric goats, indicating that human liver-like cells were present there. CONCLUSION: Goats engrafted with hHSC are capable to produce chimeric livers. Growth and propagation of human cells in the liver tissue of such transplanted goats were possible. Particularly, the human liver-like cells in chimeric goat livers had transcriptional activity specific to human hepatocytes. PMID- 12126515 TI - [A study of DNA polymerase beta mutation in human esophageal cancer]. AB - OBJECTIVE: To investigate whether DNA polymerase beta (POLB) gene mutations occur in esophageal cancer. METHODS: Thirty specimens of esophageal squamous epithelial cancer were resected during operation. A piece of cancer tissue and a piece of juxtacancerous tissue were taken from each specimen. Fourteen specimens of preinvasive esophageal carcer were obtained by esophagoscopy. Reverse transcription polymerase chain reaction (RT-PCR), single-strand conformation polymorphism (SSCP) and sequence analysis were used to examine the DNA polymerase beta genes. DNASIS and OMIGA softwares were used for sequencing. RESULTS: Obvious mutation was detected by SSCP in 13 of the 30 esophageal infiltrative cancer tissues (with a mutation rate of 43.3%) and in only 1 of the 30 juxtacancerous normal tissues from the preinvasive cancer group, obvious mutation was detected by SSCP in 5 of the 14 preinvasive esophageal cancer tissues (with the mutation rate of 35.7%). In addition, obvious mutation was also detected in one specimen of hyperplasia of squamous epithelium. Within 7 esophageal cancer specimens, 58 bp (177nt to 234nt) deleted mutation of POLB was found. There were 8 point mutation forms: (1) A --> G at 375nt (Ile --> Val) (2) T --> C at 454nt (Phe --> Ser) (3) G --> T at 462nt (Glu --> terminal code) (4) G --> A at 466nt (Gly --> Glu) (5) A --> T at 613nt (Lys --> Ile) (6) G --> C at 648nt (Gly --> Arg) (7) A --> G at 660nt (Arg --> Gly), and (8) A --> G at 670nt (Glu --> Gly). CONCLUSION: DNA polymerase beta gene mutations are discovered in human esophageal carcinoma for the first time. It may be related to the development of esophageal cancer, suggesting that POLB activity may be changed. PMID- 12126516 TI - [Association of tumor necrosis factor microsatellites TNF with the susceptibility to and outcome of postoperative severe sepsis]. AB - OBJECTIVE: To investigate whether tumor necrosis factor microsatellites TNFa and TNFb were associated with the susceptibility to and outcome of post operative severe sepsis. METHODS: 122 postoperative patients suffering from severe sepsis and 138 ethnically matched healthy individuals (controls) were included in this study. The genotypes of microsatellites TNFa and TNFb were analyzed using modified nested-PCR followed by polyacrylamide gel electrophoresis with silver staining. RESULTS: Microsatellites TNFa and TNFb consisted of 14 alleles (TNFa1 14) and 5 alleles (TNFb1, 3-5, 7) respectively. The frequency of TNFa6 microsatellite allele was significantly higher in patients with severe sepsis than in controls (20.1% versus 10.5%, P < 0.05). The frequency of TNFa2 microsatellite allele was significantly lower in patients with severe sepsis than in controls (21.3% versus 31.5%, P < 0.05). Furthermore, among patients with severe sepsis, the frequency of TNFa10 microsatellite allele was significantly higher in non-survivors than in survivors (19.5% versus 9.5%, P < 0.05). Whereas, there was no significant difference in the distribution of TNFb microsatellite alleles between patients and controls, and non-survivors and survivors (both P > 0.05). CONCLUSION: Microsatellite TNFa is significantly associated with both the susceptibility to and outcome of severe sepsis. In contrast, microsatellite TNFb is neither associated with the susceptibility to severe sepsis nor with the outcome of severe sepsis. PMID- 12126517 TI - [Effects of HOXB2 antisense oligodeoxynucleotides on the biological properties of primary human umbilical vein endothelial cells]. AB - OBJECTIVE: To explore the effects of HOXB2 antisense oligodeoxynuc leotides (Asodn) on the biological properties of primary human umbilical vein endothelial cells (ECs). METHODS: Fluorescent labelled Asodn was transfected into the endothelial cells of human unbilical vein mediated liposome and its distribution within endothelia was observed. (3)H-TdR incorporation test was employed to determine its effects on the DNA synthesis. Flow cytometry was applied to determine the change of the cell cycle. In the same time, RT-PCR was adopted to study the influence of Asodn on the expression of target genes. RESULTS: Fifteen minutes after the transfection, weak nucleic staining was observed. The fluorescent staining was the strongest 4 approximately 8 hours after the transfection and began to weaken in 16 hours. The proportion of cells in G1/0 phase in Asodn group was 53.4 +/- 3.1, significantly higher than that in control group (35.8 +/- 7.3, P < 0.01), and the proportion of cells in S phase in Asodn group was 42.2 +/- 3.5, significantly lower than that in control group (60.8 +/- 6.2, P < 0.01). The expression of HOXB2 mRNA was remarkably decreased during 24 to 48 hours. CONCLUSION: HOXB2 Asodn exerts inhibitory effects on EC proliferation dose-dependently, delays the conversion of G1 phase to S Phase, and inhibits the expression of HOXB2 mRNA. HOXB2 gene plays an important role in proliferation of endothelial cells and the mechanism is related to cell cycle. PMID- 12126518 TI - [Apoptosis of drug-resistant human ovarian carcinoma cell line 3AO/cDDP induced by arsenic trioxide and its mechanism]. AB - OBJECTIVE: To explore the effects of arsenic trioxide (As(2)O(3)) on the growth of drug-resistant human epithelial ovarian carcinoma cell line 3AO/cDDP and its possible mechanism. METHODS: Human epithelial ovarian carcinoma cell line 3AO and drug- resistant human epithelial ovarian carcinoma cell line 3AO/cDDP were cultured As(2)O(3) of different concentrations was added into the media. Cell culture without addition of As(2)O(3) was used as control. The growth inhibiting rates of 3AO/cDDP cells with various concentrations of As(2)O(3) in different time course (24, 48, 72, and 96 hours after) were studied by methyl thiazolyl tetrazolium (MTT) method. The apoptosis percentage, cell cycle phase distribution and expression of Fas/FasL gene were estimated by flow cytometry (FCM). The apoptosis phenotype of 3AO/cDDP cells was observed by cytoskeleton dying, and the apoptosis phenotype of 3AO cells was observed by acridine dying under fluorescent microscopy. RESULTS: 3AO/cDDP cell growing inhibiting rates by As(2)O(3) were different significantly in dose-dependent and time-dependent manners (P < 0.05). Within a certain concentration range, 3AO/cDDP apoptosis inducing rates by As(2)O(3) were dose -and time- dependent, and the most appropriate concentration was 3.0 micromol/L; lower concentrations of As(2)O(3) perturbed the cells to progress through S/G(2) phase, while higher concentrations of As(2)O(3) selectively induced apoptosis of S phase cells. As(2)O(3) up-regulated Fas gene expression, but did not affect FasL gene expression in both cell lines without significant difference (P > 0.05). Morphological observation indicated that As(2)O(3) induced typical apoptotic bodies in 3AO/cDDP and 3AO cells. CONCLUSION: As(2)O(3) effectively inhibits the proliferation of drug-resistant human ovarian carcinoma cell line through up-regulating Fas gene expression and inducing apoptosis of cells in S phase. PMID- 12126519 TI - [Association between alpha-1-antichymotrypsin gene polymorphism and cerebral hemorrhage]. AB - OBJECTIVE: To explore the association between alpha-1-antichymotrypsin (ACT) gene A/T polymorphism and cerebral hemorrhage among Chinese people. METHODS: The DNA of peripheral leucocytes was extracted from 220 patients with cerebral hemorrhage and 276 controls. PCR- RELP was used to determine the ACT gene polymorphism. Multiple Logistic regression was performed to explore the risk factors for cerebral hemorrhage. RESULTS: After adjusting age, gender, alcohol drinking, smoking, education,history of diabetes mellitus and primary hypertension, the analysis showed: (1) the ACT AT and TT genotypes increased the risk of cerebral hemorrhage (OR = 2.33 and 2.17 respectively, P < 0.05), but were not associated with primary hypertension. Such association was more prevalent in hypertensive cerebral hemorrhage. (2) Primary hypertension significantly increased the risk of cerebral hemorrhage (OR = 8.17 P = 0.000). (3) A significant gene environment interaction, super multiplicative type 4 interaction, was shown between both ACT AT and TT genotypes and primary hypertension with an interaction index (r) value of 2.84 and an OR value of 15.92. A significant additive type 3 gene-environment interaction was shown between both ACT AT and TT genotypes and diabetes mellitus with an OR value of 2.89. CONCLUSION: ACT gene polymorphism and primary hypertension are both independent risk factors of cerebral hemorrhage. The ACT AT and TT genotypes show a significant gene-environment interaction, however, of different types, with both primary hypertension and diabetes mellitus in the occurrence of cerebral hemorrhage. PMID- 12126520 TI - [Identification of nonmuscle mysin heavy chain 9 gene mutation in a May-Hegglin anomaly family]. AB - OBJECTIVE: To describe the clinical phenotype and identify the nonmuscle myosin heavy chain 9 (MYH9) gene mutation in the first May-Hegglin anomaly family in China. METHODS: The exons 25, 31 approximately 32 38 and 40 in the MYH9 gene of the proband and her affected father were amplified with polymerase chain reaction, and the PCR products were sequenced. After the specific point mutation in exon 38 was identified in these two cases, the corresponding region of the MYH9 gene was amplified and nuclear acid sequence analysis was conducted among 30 healthy persons, one patient with idiopathic thrombocytopenic purpura and one patient with thrombotic thrombocytopenic purpura. The CpoI restriction endonuclease map from the PCR products of exon 38 of MYH9 gene was analysed among the proband, her father and other family members, 30 normal controls, 1 ITP patient, and one TTP patient. RESULTS: The proband and her affected father manifested a typical triad of thrombocytopenia, giant platelets, and inclusion bodies in leukocytes. The patient showed mild hemorrhagic tendency since infancy, while the platelet aggregation function was normal. A 5521G --> A mutation (GAG - > AAG) in the exon 38 of the MYH9 gene existed in the proband and her affected father, resulting in a characteristic change in CpoI restriction endonuclease map. CONCLUSION: The cases of May-Heggelin anomaly in China show typical triad of thrombocytopenia, giant platelets, and inclusion bodies in leukocytes too. Mutation of MYH9 gene exists in cases of May-Hegglin anomaly in China. The point mutation is located in exon 38 (G5521A) in this family. PMID- 12126521 TI - [Differentially expressed genes in vascular endothelial cell line ECV 304 induced by high density lipoprotein]. AB - OBJECTIVE: To isolate and clone the differentially expressed genes in vascular endothelial cell induced by high density lipoprotein (HDL) so as to study the anti- atherogenesis molecular mechanism of HDL. METHODS: Differential display reverse transcription PCR method was used to analyze the differentially expressed cDNA in vascular endothelial cell line ECV 304 induced by HDL. After sequencing and homology research, several differentially expressed cDNA fragments were confirmed by Northern blot analysis. RESULTS: Up-regulated and down-regulated cDNA fragments in ECV 304 induced by HDL were isolated. Nine cDNA fragments were highly homologous to the known human genes and four were fragments of novel genes. The known up-regulated genes included genes of human STE20-like kinase, PBK1 protein, transglutaminase, myosin alkali light chain, apobec-1 binding protein 1, and death- associated protein (DAP). The known down-regulated genes included genes of ribosomal protein L7a (RPL7A), voltage-dependent anion channel isoform 2, and glycinamide ribonucleotide transformylase. Northern blot analysis revealed that the expression levels of transglutaminase and apobec-1 binding protein 1 were upregulated by 50% and 70% respectively. CONCLUSION: HDL upregulates the expression of apobec-1 binding protein 1 and transglutaminase in endothelial cells. The high level expression of transglutaminase and apobec-1 binding protein 1 in endothelial cells induced by HDL may be related to anti atherogenesis function of HDL. PMID- 12126522 TI - [A randomized control trial of mycophenolate mofeil treatment in severe IgA nephropathy]. AB - OBJECTIVE: To investigate the effectiveness safety and tolerance of mycophenolate mofeil(MMF) in severe IgA nephropathy and evaluate the dosage adjustment and course for clinical treatment. METHODS: 62 patients with IgA nephropathy diagnosed by renal biopsy as Lee's grade IV and V with urinary protein > 2.0 g/d were enrolled randomly in the trial. The initial dosage of MMF was 1.0 g/d (body weight < 50 kg) or 1.5 g/d (body weight > 50 kg). The dosage was reduced to 0.75 approximately 1.0 g/d after 6 months treatment, the maintaining dosage was 0.5 approximately 0.75 g/d after 12 months. The total course of treatment lasted at least 12 months. Another 31 patients matched with age gender and severity of renal damage were given prednisone orally (0.8mg(;)kg(;)d) (control group).Blood and urinary tests hepatic and renal function plasma albumin serum triglyceride and cholesterol 24 h protein excretion urinary NAG enzyme, creatinine clearance(Ccr) were performed before and 3 6 12 18 months after treatments in both groups 5 patients in MMF group received repeated renal biopsy. RESULTS: (1) After 3 months treatment, decrease of urinary protein (1.9 g/24 h +/- 1.6 g/24 h vs 3.2 g/24 h +/- 1.7 g/24 h, P < 0.01) and improvement of plasma albumin (41 g/L +/- 6 g/L vs 37 g/L +/- 6 g/L, P < 0.01) were observed in MMF groups while in control group, no significant changes were found in uinary protein (2.3 g/24 h +/ 1.8 g/24 h vs 2.9 g/24 h +/- 1.5 g/24 h, P < 0.05) and plasma albumin (40 g/L +/ 6 g/L vs 37 g/L +/- 6 g/L, P < 0.05). After treatment for 6, 12 and 18 months, both group showed obvious alleviation of proteinuria and albumin. At the 12th and 18th month, the proteinuria in MMF group was significantly improved than that in control group (0.8 g/24 h +/- 0.8 g/24 h vs 1.4 g/24 h +/- 1.6 g/24 h and 0.6 g/24 h +/- 0.7 g/24 h vs 1.4 g/24 h +/- 1.3 g/24 h, P < 0.05 respectively). The remission rate and total effective rate of MMF group were higher than those of the control group (44.4% vs 19.1% and 88.9% vs 61.9%, P < 0.05 respectively). Patients were administered with MMF for 13.8 +/- 6.3 months (6 approximately 30 m). (2) Serum cholesterol and triglyceride were remarkably reduced after 6,12 and 18 months treatment in MMF group, no significant difference was found in control group(P < 0.05). (3) For the 6 patients with renal insufficiency in MMF group, MMF treatment was significantly effective in 1 patient, effective in 2 patients, not effective in 3 patients with an overall effective rate of 50%. For the 7 patients with renal insufficiency in control group, the treatment was significantly effective in 1 patient, effective in 1 patient, not effective in 5 patients and total effective rate is 28.6%. (4) 5 patients in MMF group received repeated renal biopsy after 7 approximately 12 months treatment (mean 9.8 +/- 2.3 m). The results showed that the interstitial lesions were alleviated. No special drug-induced renal damage was obtained. (5) Side effects: 3 patients in MMF group suffered from slight diarrhea, 1 patient herpes zoster, all of them got remission without drug withdrawal. 1 patient suffered nausea in the first weeks. No significant change was found in hepatic function (P > 0.05). CONCLUSIONS: MMF is more effective in reducing proteinuria and serum lipid than the currently widespread use of prednisone therapy in IgA nephropathy patients with Lee SMK's grade IV approximately V and urinary protein > 2.0 g/d. Treatment with MMF associates with less adverse effect and good tolerance. PMID- 12126523 TI - [Acquired mutation of type II transforming growth factor-beta receptor gene in glomerulosclerosis]. AB - OBJECTIVE: To investigate the acquired mutation of type II transforming growth factor beta receptor TbetaR II gene plays a role in the process of glumerulosclerosis. METHODS: Biopsy specimens of 32 patients with primary focal segmental glomerulosclerosis (FSGS) were examined. Microdissection was used to isolate the morphologically normal glumeruli, segmental sclerotic glomeruli (SSG) global sclerotic glomeruli (GSG), and in the same specimen, representing the sequence of glomerulosclerosis. DNA was extracted by one-step method. The A10 microsatellite instability (709 approximately 718) in TbetaRII gene was detected by high-fidelity PCR and strand length polymorphism. Scanning technique was used to calculate the mutation rate of TBRII gene A10 sequence. RESULTS: A total of 192 sites of glomeruli from 32 patients were analyzed. The standard curve for mutant integrated pixel density (IPD) percentages (10% 32% 55% 75% 88%) versus DNA mutant ratios (17% 24% 53% 79% 95%) showed excellent linear correlation (r = 0.990 8). DNA from the glomeruli of NG, SSG, and GSG of all cases showed very low ratios of mutant allele (1 approximately 5%). CONCLUSION: Mutation of TbetaRII gene A10 microsatellite sequence can not been detected during the progress of glomerulosclerosis. PMID- 12126524 TI - [Distraction osteogenesis for treatment of temporomandibular joint ankylosis]. AB - OBJECTIVE: To study the application of distraction osteogenesis in temporomandibular joint (TMJ) ankylosis patients. METHODS: 11 temporomandibular joint ankylosis patients (13 sides of TMJ) were treated by this technique. After removal of bony mass on the ankylosis area, a L-shape osteotomy of the ramus was performed to create the distraction transport disc which connected the mandible by distractor. Distractions were started on the 4 approximately 8th postoperative day. The distraction rhythm and rate were 0.25 mm 4 times a day. Distraction was stopped when contact was made between the transport disc and glenoid fossa. The patients underwent active mouth opening postoperatively. Distractor was kept in place for 3 approximately 4 months after completion of distraction and then removed under local anesthesia. RESULTS: Postoperatively, the range of mouth opening of the 11 patients (13 sides) was increased to normal (33 approximately 45 mm), bone formation in the gaps were perfect and the neocondyle was remodeling and approximated normal shape. No complications were found. CONCLUSION: Distraction osteogenesis for arthroplasty has many advantages compared with traditional methods. It is valuable in clinical practice as a new method for treatment of TMJ ankylosis. PMID- 12126526 TI - [Effect of P(ET)CO(2) monitoring on cardiac function during hepatectomy]. AB - OBJECTIVE: To investigate the relationship between end-tidal Pco2 (P(ET)CO(2)) and cardiac output (CO) during hepatectomy. METHOD: Swan-Ganz catheter was inserted into the right jugular vein of twenty-one patients undergoing hepatectomy to observe the hemodynamic parameters. Cardiac output (CO) was examined by thermodilution technique. Mechanical ventilation was performed after anaesthesia induction, and P(ET) CO(2) was monitored continuously. The relationship between CO and P(ET)CO(2) was analyzed. RESULTS: A strong positive correlation was found between P(ET)CO(2) and CO, r =0.90, P < 0.05. CONCLUSION: P(ET)CO(2) can be used as a predictor of CO. This technique is safe, simple and can be used in hemodynamic unstable patients during the major surgical procedure and in patient with poor cardiac function. PMID- 12126525 TI - [Treatment of hyperthyroid atrial fibrillation associated with Graves disease by prednisone]. AB - OBJECTIVE: To investigate the clinical effect of prednisone on hyper thyroid atrial fibrillation associated with Graves disease (HAFGD). METHODS: Twenty-four patients with hyperthyroid atrial fibrillation associated with Graves disease were divided into two groups: traditional antithyroid group (10 cases, with 2 males and 8 females, treated by methimazole 15 approximately 30 mg/d and propranolol 15 approximately 30 mg/d) and prednisone group (14 cases with 3 males and 11 females, treated by methimazole 15 approximately 30 mg/d, propranolol 15 approximately 30 mg/d, and prednisone 30 mg/d). The effects of these two remedies on reversion from atrial fibrillation to sinus rhythm were compared. RESULTS: The cardiac rhythm reverted to sinus rhythm in 12 out of the 14 cases in prednisone group with a reversion rate of 86% and the mean reversion time of 3.8 months (range 3.8 +/- 2.6 months), and in 4 out of the 10 cases in traditional antithyroid remedy group with the mean reversion rate of 40% and mean reversion time of 2.8 months (range 2.8 +/- 1.0 months). The reversion rate was significantly higher in prednisone group than in the traditional remedy group (P < 0.05). CONCLUSION: Treatment of prednisone is more beneficial to reversion of atrial fibrillation into sinus rhythm among patients with HARGD. PMID- 12126527 TI - [Cox/maze III procedure combined with mitral valve replacement in treatment of rheumatic mitral valve disease with atrial fibrilation]. AB - OBJECTIVE: To compare the curative effect of Cox/maze III procedure combined with mitral replacement and that of mitral valve replacement (MVR). METHODS: Fifty-six patients suffering from rheumatic heart disease with atrial fibrillation (AF) were treated by Cox/maze III procedure combined with MVR (maze group). Another 56 age, sex, and heart function-matched patients with the same diagnosis underwent MVR alone during the same period. Warfarin was administered after operation in both groups. Comparison of operative complication and curative effects was made. RESULTS: The aortic cross-clamp time and cardio pulmonary bypass time (CPB) were longer in maze group than in MVT group (75 +/- 22 min vs 41 +/- 11 min, P < 0.05 and 124 +/- 40 min VS 68 +/- 19 min, P < 0.05). Bleeding happened after the heart reatored beating in 2 patients in maze group and in one patient in MVT group, all these 3 patients responding satisfactorily to hemostasis. The early post operative mortality was 1.79% (1/56) in both groups. In maze group, AF disappeared in all patients but one who had node rhythm. Normal sinus rhythm was restored in 98.18% of the patients (54/55). Atrial contractility was restored in all patients with sinus rhythm. One year after operation, 98.18% patients' cardiac function changed to grade and 1.82% changed to grade II. In MVR group AF disappeared after operation temporarily for 24 hours in 7 patients and re appeared, and AF disappeared in one patients for 2 years so far. One year after operation, the cardiac function of 94.6% patients in MVR group changed to grade I, of 3.6% patients to grade II, and of 1.8% patients to grade III. No serious hemorrhage relate d to anticoagulant therapy happened. One patient in MVR group suffered from hemiplegia due to cerebral embolism. The late mortality was 1.8% on maze group amd 3.6% in MVR group. CONCLUSION: Cox/maze III procedure combined with NVR is safe and effective in treating rheumatic heart disease with AF. PMID- 12126528 TI - [Management of ruptured aneurysm of anterior communicating artery by emergency operation]. AB - OBJECTIVE: To investigate the effectiveness of emergency operation on ruptured anterior communicating artery. METHODS: Twenty-one cases of ruptured anterior communicating aneurysms, 9 of grade III, 10 of grade IV and 2 grade V according to Hunt-Hess classification, were treated by emergency clipping. RESULTS: By the time of being discharged from hospital, 16 patients were perfectly recovered, 2 suffered from moderated disability, 1 suffered from severe disability and 2 were dead. No patients of grade III before operation died. CONCLUSION: Emergency clipping of ruptured aneurysm of anterior communicating artery prevents death and deterioration of condition caused by aneurysmal re-rupture and relieves the secondary cerebral vasospasm, thus decreasing the mortality and improving the prognosis. PMID- 12126529 TI - [Association between pre-excitation syndrome and 7q3 D7S505 pseudonym gene]. AB - OBJECTIVE: Pre-excitation syndrome is considered to be autosomal dominant hereditary disease. The objective of this study was to search the genetic foundation of the pre-excitation syndrome. METHODS: Genomic DNA was isolated from peripheral lymphocytes obtained from 44 cases of patients with pre-excitation syndrome and 53 normal persons. Polymorphic short tandem repeats(STR) were amplified using polymerase chain reaction (PCR) and analyzed by polyacrylamide gel electrophoresis. The genotype of each individual was determined by polymorphic STR including D7S505,D7S688,D7S483. Association analysis between the pre-excitation syndrome and the 3 STR (D7S505,D7S483 and D7S688) was tested by genotyping. RESULTS: The relative risk(RR) of alleles A2 A3 A4 and A6 of D7S505 were 1.051 6, 3.432,1.563 1 and 1.714 3 respectively all of which were more than 1, but only the RR of A3 had statistic significant difference, P < 0.05 after tested by kappa(2). It supposed that the distribution of allele A3(266 bp) of D7S505 in patients with pre-excitation syndrome was much higher than that in normal controls, which suggested that pre-excitation syndrome is associated with D7S505. Whereas, there were no significant difference in every allele of D7S483 between the pre-excitation syndrome and normal persons, which suggested that pre excitation syndrome is not associated with D7S483. CONCLUSION: The pre-excitation syndrome is associated with D7S505,the result is the foundation of the molecular genetics of the disease. PMID- 12126530 TI - [Correlation of the uptake of technetium-99m methoxyisobutyl isonitrile with expression of multidrug resistance genes mdr-1 and MRP in human lung cancer]. AB - OBJECTIVE: To investigate the relationship between the uptake of technetium-99m methoxyisobutyl isonitrile ((99m)Tc-MIBI) and the expression of multidrug resistance genes mdr-1 and MRP in lung cancer so as to guide chemotherapy. METHODS: Eighteen lung cancer patients were given 1 110 MBq(99m)Tc-MIBI intravenously. Chest single-photon emission-computed tomography was performed 60 and 120 min after the injection. The early and delay uptake rates and retain index of (99m)Tc-MIBI were calculated by the count ratio of the lung lesion to contralateral normal lung tissue. The expressions of mdr-1 and MRP genes in specimens of lung cancer and normal lung tissues near the cancers resected during operation among the 27 patients were detected with RT-PCR method. RESULTS: The positive rate of (99m)Tc-MIBI in the chest of lung cancer patients was 83.3% (15/18). The early uptake ratio of imaging in lung cancer patients was 1.99 +/- 0.64. After time decay correction, the rate of target to background and the retain index in 13 delay imagings were 2.06 +/- 0.69 and -45% approximately 33% respectively. In specimens of lung cancer, the positive expression rates of mdr-1 and MRP were 22.2%(6/27) and 63%(17/27) with the expression amount of 0.39 +/- 0.1 and 0.23 +/- 0.17 respectively. In specimens of normal lung tissues near lung cancer, the positive expression rates of mdr-1 and MRP genes were 35%(7/20) and 45.0%(9/20) with the expression amount of 0.44 +/- 0.14 and 0.17 +/- 0.18 respectively. In lung cancer tissues, the positive rate of MRP was higher than that of mdr-1 (P < 0.05), however, in the tissues near lung cancer, no significant difference was found between the positive rate of MRP and that of mdr 1 (P >< 0.05). There was no marked correlation (P >< 0.05) among the co expression or expression of mdr-1 and/or MRP and the early uptake or retain index in lung cancer. CONCLUSION: The primary chemoresistance of lung cancer has not significant correlation to the expression of multidrug resistance genes mdr-1 and MRP. Other resistance mechanism may exist. (99m)Tc-MIBI imaging has not much clinical value in predicting the expression of mdr-1 and MRP genes. PMID- 12126531 TI - [Computer-assisted planning for neurosurgery: a clinical study]. AB - OBJECTIVE: To investigate the clinical effect of computer-assisted planning for neurosurgery (CAPN). METHODS: 100 groups of directional targets were randomly sampled. The targets' directional three-dimension coordinates measured with CAPN and those measured by traditional reading device were analyzed statistically. CAPN was actualized in 137 cases of stereotactic operation. RESULTS: The data measured with CAPN and those with traditional reading device were analyzed with two samples. The paired t test showed no statistically significantly difference between the data of targets measured by both methods (t < t(0.05/2,99)=1.984, P >< 0.05). Different kinds of operation based on CAPN were performed upon 137 cases success fully. CAPN was proved reliable. All operations were successful and the clinic effects satisfactory. CONCLUSION: CAPN is accurate and reliable, and has positive effect on the improvement of stereotactic neurosurgery. PMID- 12126532 TI - [Study on natural killer cell subset and activity in patients with depression]. AB - OBJECTIVE: The study was to investigate natural killer cells subset and activity in patients with depression. METHODS: 46 outpatients with unipolar depression and 46 normal controls were recruited. Peripheral blood samples were drawn for natural killer cells subset and activity. And the results in patients with depression before and after therapy were compared with each other. Flow cytometer was used for natural killer cells subset and enzyme-released assay was used for their activity. RESULTS: Natural killer cells subset and activities (effect target cell ratio: 20:1,10:1)in patients with depression were significantly lower than normal controls (P < 0.01). Natural killer cell activities were increased significantly after therapy (P < 0.01)but subset was not (P > 0.05). CONCLUSION: The study suggested that unipolar depression may impact natural killer cells count and function. PMID- 12126533 TI - [Isolation, cultivation and identification of stem cells from cerebral cortex of mouse embryo]. AB - OBJECTIVE: To isolate, cultivate and identify the neural stem cells from the cortex of embryo mouse in vitro for the further related research. METHODS: Cerebral hemisphere of mouse fetuses were taken out. Suspension of cerebral cells was made. Neural stem cells were identified by immunofluorescent cytochemistry to the primary culture. Neural stem cells were isolated and cultured in special culture medium using single-cell clone culture technique. After the stem cells were stably cultured successively for 20 generations, nestin, glial fibrillary acidic protein, beta-tublin, and galactosidase (Galc) were detected by indirect immunofluorescence cytochemistry. RESULTS: The stem cells isolated from the cerebral hemisphere of mouse embryo with positive expression of nestin could be passaged in vitro for at least 20 generations. Inoculated on the coverglass with poly-1-ornithine and cultured in medium containing serum, the stem cells were induced to differentiate. The cells differentiated from these stem cells showed green fluorescence by GFAP fluorescent staining, red fluorescence by beta-tublin fluorescent staining, or green fluorescence by Galc fluorescent staining. CONCLUSION: The stem cells from cortex of mouse embryo can be successfully isolated and cultured in vitro. The stem cells can be passaged in series and possessed the potential of multi-directional differentiation and express the specific antigens of neuron, astrocytes, and oligodendrocytes. Thus they can be used for further experimental study. PMID- 12126534 TI - [Regulation of human progesterone receptor isoforms A and B in uterine endometrial carcinoma by estrogen and insulin-like growth factor-1]. AB - OBJECTIVE: To investigate the regulation of human progesterone receptor isoforms A and B in uterine endometrial carcinoma by estrogen and insulin-like growth factor-1 (IGF-I) so as to provide theoretical basis for clinical hormone treatment of uterine endothelium cancer. METHODS: The uterine endometrial adenocarcinoma cell line HEC-IB and the breast cancer cell line MCF-7 were cultured in vitro. The HEC-IB cells were stimulated by 10 nmol/L estrogen and 20 ng/ml IGF-I respectively for 72 h. The MCF-7 cells were stimulated by 10 nmol/L estrogen for 72 h. Western blotting was used to examine the protein levels of the two isoforms of receptors of progesterone. RNA was extracted from the cells. Reverse transcription-polymerase chain reaction (RT-PCR) was used to detect the mRNA levels of the isoform B. RESULTS: (1) Western blotting showed that the photodensity values of hPRA and hPRB did not change significantly after the HEC IB cells were stimulated by estrogen for 72 hours (P = 0.716, and 0.391 respectively), however, the hPRA and hPRB were significantly down-regulated after IGF-I had been given for 72h (P = 0.008 and 0.002 respectively). After MCF-7 cell was given estrogen for 72 h hPR-A tended to be up- regulated but this change had no significant difference (P = 0.074) however, hPRB was significantly up- regulated (P = 0.044). (2) RT-PCR showed that hPRB mRNA was not expressed in HEC IB cells originally, and became positive after stimulation by estrogen and IGF-1 for 72 hours respectively, however, the expression level being higher in the HEC IB cells stimulated by estrogen than in those stimulated by IGF-1. hPRB was not expressed in MCF-7 cells originally too, and became positive after simulation by estrogen and IGF-1 for 72 hours respectively, however, the expression level being higher in the MCF-7 cells stimulated by estrogen than in those stimulated by IGF 1. CONCLUSION: (1) Estrogen and IGF-I regulate hPR isoforms and have cell specialty. (2) Estrogen and IGF-I up- regulate hPRB mRNA at gene level or transcription level, but there are some inhibitory factors which make the protein production become less after transcription. PMID- 12126535 TI - [Effect of superoxide dismutase on adhesion molecules expression in skeletal muscle ischemia/reperfusion injury in rats]. AB - OBJECTIVE: To study the effect of SOD on the expression of CD11b/CD18 and ICAM-1 after skeletal muscle ischemia / reperfusion injury in rats. METHODS: Establish the rat model of hind limb ischemia / reperfusion. One hundred and two rats were divided into 5 groups: sham operation group (Group I n = 6), ischemia group (GroupII n = 6), ischemia / reperfusion injury group (Group III n = 30), normal saline treatment group (Group IV n = 30), superoxide dismutase treatment group (Group V n = 30). Flow cytometric analysis and immunohistochemistry were used to assess the expression of CD11b/CD18 on the leukocytes, ICAM-1 in skeletal muscle at the end of ischemia and 1h, 2h, 4h,8h,12h after reperfusion. The changes of MDA in the plasma,MPO in the skeletal muscle and the tissue morphology were studied too. RESULTS: In group III,the expression of CD11b/CD18, ICAM-1,MDA,MPO were increased evidently. The longer time of reperfusion,the more serious the skeletal muscle's injury was. In group V, the changes were ameliorated as compared with group III (P < 0.05), group IV has no difference with group III. CONCLUSION: These data indicate that the expression of CD11b/CD18 and ICAM-1 were significantly correlated with skeletal muscle ischemia/reperfusion injury. SOD can inhibit the expression of free radicals and the adhesion molecules,consequently skeletal muscle ischemia/reperfusion injury is prevented. PMID- 12126537 TI - [Transfection of MDR1-mRNA into human mononuclear cells to improve their resistance to anticancer agents, an in vitro study]. AB - OBJECTIVE: To investigate the expression and function of P-glycoprotein (P-gp), an efflux pump encoded by multidrug resistance complementary DNA (MDR1), in human mononuclear cells (MNCs) transfected with MDR1 mRNA. METHOD: Two kinds of human MDR1 mRNA, pT7TS-MDR1 and pGEM5Zf (+)-MDR1 with difference in the 5' and 3' untranslated regions only, were constructed and modified and then were transfected into human MNCs rich in hematopoietic progenitor cells from a patient with small cell lung cancer. The expression efficiency and pump function of P-gp were measured with FACS. Rhodomine 123 efflux test was used to examine the function of P-gp. Un- transfected mononuclear cells from the same person were used as controls. RESULT: The expression of P-gp 12 hours after transfection was 2.16 % in control group, 8.94% in pGEM5Zf (+)-MDR1 group, and 19.14% in pT7TS MDR1 group. The expression of P-gp 72 hours after transfection was 2.12% in control group, 6.12% in pGEM5Zf(+)-MDR1 group, and 10.71% in pT7TS-MDR1 group, significantly higher in the third group. Rhodamine-123 efflux test showed that the efflux-pump function of P-gp 12 hours after the trransfection was 19.20% in control group, 25.59% in GEM5Zf (+)- MDR1 group, and 35.02% in pT7TS-MDR1 group (all P < 0.01). The expression and presence of P-gp by transfected cells lasted at least 72 hours. CONCLUSION: Transfection of human mononuclear cells with MDR1 mRNA significantly increases their resistance to anticancer agents. PMID- 12126536 TI - [Therapeutic effects of human interleukin 10 gene transfer on severe acute pancreatitis in rats, an experimental study]. AB - OBJECTIVE: To study the therapeutic effects of human interleukin 10 (IL-10) gene transfer on severe acute pancreatitis (SAP) in rats. METHODS: Twenty healthy SD rats were injected intraperitoneally with SA liposome, SA liposome/pcDNA3 or SA liposome/pcDNA3-IL-10. Another twenty SD rats were randomly divided into five groups: rats in one group underwent laparotomy only (normal control), and SAP was induced in the other 4 groups induced by homogeneous injection of sodium taurocholate beneath the pancreatic capsule. Among the 4 SAP groups, one group did not receive any drugs, and liposomes, pcDNA3 or pcDNA3-IL-10 complexed with cationic liposomes were administered to the other groups. Drugs were administered by a single intraperitoneal injection thirty minutes after SAP had been induced. The levels of IL-10 in pancreas, liver and lungs were determined by ELISA kits. The level of serum amylase, histology, and tissue tumor necrosis factor (TNF) were assessed and mortality rate was observed in different groups for one week. RESULTS: The levels of IL-10 in the pancreas, liver and lung 24 hours after IL-10 gene transfer, increased significantly (all > 350 pg/g), and then gradually decreased, however, the levels of IL-10 were still significantly higher that those in the control groups (P < 0.05) 96 hours later and decreased to normal in one week. The levels of IL-10 of transfer control group were not significantly different from those of the normal control group. The levels of IL-10 expression in pancreas, liver and lungs were increased significantly in the gene therapy group, compared with the SAP group. The serum amylase level was (4 300 +/- 700) U/L in normal control group, increased to (20 300 +/- 1 100) U/L 24 hour after SAP induction without a difference between the therapy control group and SAP group, and decreased to (6 800 +/- 700) U/L after IL-10 gene therapy (P < 0.05). The histological score of pancreas was 4.1 +/- 0.2 24 hours after the induction of SAP, and was 3.2 +/- 0.3 in the IL-10 therapy group. The level of TNF in pancreas, liver, and lungs 24 hours after the induction of SAP was significantly higher than that in normal control group (P < 0.05) and was not different from that in therapeutic control group. However, it was decreased markedly in IL-10 therapy group (P < 0.05). No rat in any group died within 2 days after onset. There was no difference of mortality between SAP group and therapeutic control group. The one-week mortality was 90% in the whole SAP group. The one-week mortality of IL-10 gene therapy group was 30 %, significantly lower than that in SAP group (P < 0.05). There was no significant difference in the therapeutic control groups and the SAP group. The values of relative risk of SAP group, SA liposome group, and pcDNA3 group were 12, 8, and 11 times higher than that of gene therapy group (P < 0.05). CONCLUSION: Cationic liposome mediated pcDNA3-IL 10 gene therapy decreases significantly the severity and mortality of SAP. PMID- 12126538 TI - [The abnormal reaction to post methionine loading test in type diabetes with or without retinopathy]. AB - OBJECTIVE: To investigate the reaction of patients with type 2 diabetes mellitus (DM) with retinopathy (DR) or without retinopathy (NDC) to post methionine loading (PML) test, the value of PML test to identify hyperhomocystenemia (Hhcy), and the possible mechanism of vascular damages caused by Hcy. METHODS: Solution of methione was given orally to 17DR patients, 15 NDC patients, and 28 normal controls. The plasma fasting homocysteine (Fhcy) level and the Hcy levels 4 hours after PML test (Phcy) of these subjects were measured by fluorescence polarization immunoassay (FPIA) before and 4 hours after PML test. The levels of fasting plasma folic acid and vitamin B(12) were tested by radioimmunoassay. PCR RFLP was used to test the mutation of methelene tetrahydrofolate reductase (MTHFR) C6671. BMI, HbA1c, FBG, PBG, BUN, Cr, Ch and TG were also detected. The final metabolites of NO were determined by Griess method. RESULTS: The levels of BMI, HbA1c, FBG and PBG in DM group and NDC group were higher than those in the control group (P < 0.05). The Fhcy levels were 13.87 micro mol/L and 11.6 +/- 1.3 micro mol/L in DR and NDC groups respectively, both higher than that in control group (8.85 micro mol/L, both P < 0.05). The Phcy was 37.3 +/- 1.3 micro mol/L in DR group and was 25.03 micro mol /L in NDC group, both higher than that in the control group (22.65 micro mol/ L, both P < 0.05). The difference between Fhcy and Phcy (Dhcy) was 24.36 micro mol/L in DR group and was 14.26 micro mol/L in NDC group, both significantly higher than that in the control group (15.07 micro mol/L, both P < 0.05). The proportion of BB genetype of MTHFR in DR group (9/17) was higher than those in NDC group (5/28) and control group (2/15). The number of Hhcy patients diagnosed by the level of Fhcy was 9 before PML test and became 15 after PML test diagnosed by the level of Phcy (P < 0.05). The whole prevalence of Hhcy in DR group was higher than that in NDC group, both the prevalence of Hhcy in DR group and the prevalence of Hhcy in NDC group were higher than those in CON group (P < 0.001). The levels of fasting metabolite of NO were 5.4 +/- 1.8 micro mol/L, 5.4 +/- 1.8 micro mol/L, and 3.4 +/- 1.7 micro mol/L in DR group, NDC group, and control group respectively. The levels of metabolits of NO after meal were 25.0 +/- 2.3 micro mol/L, 15.0 +/- 2.3 micro mol/L, and 10.4 +/- 2.0 micro mol/L in DR group, NDC group, and control group respectively. The differences between DR group and control group were statistically significant (P < 0.05). The correlation coefficient of metabolites of NO to Hcy was 0.744 (P < 0.001). CONCLUSION: Patients with diabetes mellitus, especially those with retinopathy, had abnormal reaction to PML test, which may be related to higher BB genetype of MTHFR and dysfunction of insulin. PML test can be used to identify more Hhcy patients. Disturbance of NO metabolism may be one of the mechanisms of the vascular damages caused by Hcy. PMID- 12126539 TI - [Effect of cilostazol on adhesion molecules of STZ-induced diabetic rats]. AB - OBJECTIVE: To investigate the effect of cilostazol on the expression of CD(54), CD(106), CD(62P) in kidney, aorta, surface of platelet, and mononuclear cell of blood in STZ-induced diabetic rats. METHODS: Rats were randomly divided into four groups: normal control group (n = 8), diabetes control group (n = 8), insulin group (n = 7 subcutaneous injection of insulin, 1 approximately 4 U * d(-1)), and cilostazol group (n = 7, gastric infusion of cilostazol, 18 mg * kg(-1) * d(-1)). Sciatic nerve conductive velocity and level of CD(54)/CD(62p) on the surface of mononuclear cell/platelet were examined by flow cytometry. Immunohistochemistry was used to examine the expression of CD(54) and/or CD(106) in kidney and aorta. RESULTS: The sciatic nerve conductive velocity was 20.3 +/- 2.2 m * s(-1) and 28.9 +/- 7.9 m * s(-1) in diabetic rats and cilostazol group respectively (P < 0.05). The mononuclear cell surface expression of CDa-a(c)54 was 69.1 +/- 14.9% and 25.9 +/- 8.6% in diabetic group and cilostazol group respectively (P < 0.05). The expression of CD(54)/CD(106) in kidney and aorta was decreased. The expression of CD(62p) on the surface of platelets was 40.4 +/- 8.7% in diabetic group and 28.5 +/- 3.7% in cilostazol group (P > 0.05). CONCLUSION: Cilostazol treatment delays the development of diabetic pathological change in kidney, aorta and sciatic nerve and decreases the expression of CD(54) and CD(106). PMID- 12126540 TI - [Associations of insulin resistance and pancreatic beta-cell function with plasma glucose level in type 2 diabetes]. AB - OBJECTIVE: To investigate the influence of insulin resistance and pancreatic beta cell function on plasma glucose level in type 2 diabetes so as to provide theoretical basis for reasonable selection of hypoglycemic agents. METHODS: The plasma non-specific insulin (NSINS), true insulin (TI) and glucose in eight-one type 2 diabetics, 38 males and 43 females, with a mean age of 53 years, were examined 0, 30, 60 and 120 minutes after they had 75 grams of instant noodles. The patients were divided into two groups according to their fasting plasma glucose (FPG): group A (FPG < 8.89 mmol/L) and group B (FPG> = 8.89 mmol/L). The insulin resistance was evaluated by HOMA-IR, the beta-cell function was evaluated by HOMA-beta formula and the formula deltaI(30)/deltaG(30) = (deltaI(30) deltaI(0))/(deltaG(30)-deltaG(0)). The insulin area under curve (INSAUC) was evaluated by the formula INSAUC=FINS/2+INS(30)+INS(60)+INS(120)/2. RESULTS: The mean FPG was 6.23 mmol/L in group A and 12.6 mmol/L in group B. PG2H was 11.7 mmol/L in group A and 19.2 mmol/L in group B. The TI levels in group B at 0, 30, 60, 120 min during standard meal test were significantly higher than those in group A: 6.15 +/- 1.06 vs 4.77 +/- 1.06, 9.76 +/- 1.1 vs 5.88 +/- 1.1,14.68 +/- 1.11 vs 6.87 +/- 1.1 and 17.13 +/- 1.12 vs 8.0 +/- 1.1 microU/dl (all P< 0.01). The NSINS showed the same trend. The insulin resistance in group B was 1.5 times that in group A. With the insulin resistance adjusted, the beta cell function in group A was 5 to 6 times that in group B. The INSAUC in group A was 1.66 times larger than that in group B, especially the INSAUC for true insulin (2 times larger). The contribution of insulin resistance and beta cell function to PG2H was half by half in group A and 1:8 in group B. beta cell function calculated by insulin (Homa-beta) explained 41% of the plasma glucose changes in group A and 54% of the plasma glucose changes in group B. The contribution of insulin deficiency to plasma glocose was 3.3.times that of insulin resistance in group A and was 9.5 times that of insulin resistance in group B. Insulin sensitivity explained 12% of the plasma glucose changes in group A, and only 5.7% of the plasma glucose changes in group B. CONCLUSION: Diabetics with FPG greater than 8.89 mmol/L have both higher insulin resistance and poorer beta-cell function, their hyperglycemia being caused mainly by beta-cell failure, The combined use of insulin sensitizer and insulin or insulintropic agents during the initial stage of treatment is effective. PMID- 12126541 TI - [Sleep time of children from 1 month to 5 years of age in Shanghai, an epidemiological study]. AB - OBJECTIVE: To assess the age specific sleep time among children 1 month to 5 years of age in Shanghai, China, and to identify the related factors. METHODS: A questionnaire survey was conducted among the parents of 3 266 children 1 month to 5 years of age, randomly selected in five urban districts of Shanghai in August and September of 1999 and 2000, with a response rate of 97.83%. RESULTS: The total sleep time among children less than 48 months of age in Shanghai was less than that among children the same age in Western countries. The factors related to children's inadequacy of total time included parental insufficient sleep time, children's difficult temperament parental presence at sleep onset of children, and cosleeping. CONCLUSIONS: The total sleep time among children at tender age in Shanghai is on the low side. The main influencing factor of insufficiency of sleep time among children is the family environmental factor, especially parental improper response to children's sleep behaviors. PMID- 12126542 TI - [Fine deletion mapping on chromosome 8p21 - 8p22 in adenocarcinoma of lung]. AB - OBJECTIVE: To determine the common deletion region of loss of heterozygosity (LOH) on chromosome 8p21-8p22 in adenocarcinomas of lung, and facilitate identification of candidate tumor suppressor genes associated with adenocarcinoma of lung. METHODS: PCR and microsatellite analysis were used to examine the LOH frequency of 17 microsatellite loci at the 8p21-8p22 in the samples resected from 32 patients with lung adenocarcinoma and adenosquamous carcinoma. The relationship of LOH for each marker to pathological grade and that to clinical stage are investigated. RESULTS: Thirty-one out of the 32 (96.67%) samples showed allelic loss in at least one of the 17 markers. The most frequent LOH loci were mainly located in the three regions: D8S254-261, D8S1827-1731, and D8S1135 loci. Among them, the LOH frequency of D8S261 locus was related to the stage of tumor (P < 0.05). CONCLUSION: An interval of common deletion on chromosome 8p22, encompassing D8S254-261, D8S1827-1731 and D8S1135, might harbor candidate tumor suppressor gene(s) associated with pathogenesis of adenocarcinoma of lung. PMID- 12126544 TI - [Study of treatment of refractory rheumatoid arthritis with autologous peripheral blood stem cell transplantation]. AB - OBJECTIVE: To explore the efficacy, safety and immune reconstitution of autologous peripheral blood stem cell transplantation (APBSCT) using T cell depleted grafts in the treatment of refractory rheumatoid arthritis (RA). METHODS: One patient with RA was treated with APBSCT. The method included mobilization with 2 g/m(2) cyclophosphamide (CY) and subcutaneous injection of granulocyte-colony stimulating factor (G-CSF). Immunomagnetic selection of CD34(+) cells from the leukapheresis products was performed to deplete potentially autoreative lymphocytes. The conditioning regimen consisted of intravenous administration of 2 g/m2 CY and 90 mg/kg ATG, with subsequent reinfusion of the graft. G-CSF was used to help hematopoietic and immunologic reconstitution. Phenotype of the peripheral blood lymphocytes was analyzed to observe the immunologic reconstitution after transplantation. RESULTS: The patient completed the mobilization, conditioning regimen and transplantation successfully. The hematologic recovery was rapid and the patient achieved clinical remission. The erythrocyte sedimentation rate (ESR), C-reactive protein (CRP) decreased to normal level and the rheumatoid factor (RF) turned negative after a follow-up of 12 months. An ongoing course of immunologic reconstitution was observed. CONCLUSION: APBSCT is effective and safey for refractory RA, and can induce improvement of disease activity. The course of immunologic reconstitution after transplantation remains to be observed in long-term followup. PMID- 12126543 TI - [Study on single nucleotide polymorphism of TIGR gene in primary open-angle glaucoma patients]. AB - OBJECTIVE: To detect the single nucleotide polymorphism (SNP) of trabecular meshwork-induced glucocorticoid response protein (TIGR) gene and to investigate the association between SNP and primary open-angle glaucoma (POAG) in Chinese Han population. METHODS: High throughput conformation sensitive gel electrophoresis (HTCSGE) and fluorescent labeling automated sequencing method were used to screen and identify the SNPs of TIGR in 82 unrelated patients with POAG. Restriction endonuclease analysis was used to detect the SNPs of TIGR in 150 sex-and age matched unrelated control subjects without POAG. The frequencies of genotypes and alleles of each SNP between the two groups were compared by chi-square test. RESULTS: A total of six SNPs were identified in TIGR gene with a length of 2172 bp, including all coding regions and part of promoter, introns and 3'UTR region: 1-83(G --> A), G12R, R46X, R76K, IVS2 + 35(A --> G), and T353I. Significant differences were found in frequencies of the genotype TC and the allele T of T353I between POAG patients and control subjects. The frequency of heterozygote TC was 12.20% in POAG patients, significantly higher than that in control subjects (3.33%, kappa(2) = 6.885, P = 0.009, OR = 4.028, OR95% CI = 1.327 approximately 12.223). The frequency of the allele T was 6.10% in POAG patients, significantly higher than that in control subjects (1.67%, kappa(2) = 6.655, P = 0.010). No significant difference was found between the two groups in frequencies of genotypes and alleles of the other SNPs. CONCLUSION: The T353I polymorphism of TIGR gene is associated with POAG in Chinese Han population. PMID- 12126545 TI - [Influence of aging, menopause and knee osteoarthritis on women's femoral neck strength and muscle force ]. AB - OBJECTIVE: To study the influence of aging, menopause and knee retrograde osteoarthritis on femoral neck strength (FS) and muscle force (MF) of women. METHODS: A total of 306 healthy women aged 15 - 74 were stratified according to age and menstruation status, among which 76 menopausal women suffered from knee osteoarthritis. The bone mineral density (BMD) of lumbar vertebrae (L2 - 4) and femoral neck was measured by DEXA among all subjects. The FS and MF were measured by the Mobility Evaluation System (MES). RESULTS: (1) Influence of aging: FS and MF were at the peak in the group aged 15 - 19 and then decreased with the increase of age. Comparing with those in 15 -19 age group, the FS and MF values of the group aged 60 -74 decreased by 26.2% and 46.9% with the annual decrease rates of 0.55% and 0.99% respectively. (2) Influence of menopause: In the group aged 40 - 55, the FS and MF values of the postmenopausal ones were 9.4% and 21.7% lower than those in the women with normal menstruation respectively. ( 3) Influence of osteoarthritis: The women with knee osteoarthritis had lower FS and MF than those with normal knees; however, only the difference in MF was statistically significant (t = 5.33, P < 0.001). (4) Multiple regression suggested that MF and femoral neck BMD have positive correlation with FS, whereas age and BMI have negative correlation with FS. CONCLUSION: FS and MF decrease annually with the increase of age and such decrease, especially that of MF, is aggravated with the onset of menopause. The menopausal women with knee osteoarthritis have lower FS and MF in comparison with normal controls. It is critical to maintain muscle mass and to enhance muscle force for the prevention and treatment of knee retrograde osteoarthritis. PMID- 12126546 TI - [Effects of preincisional epidural administration of lidocaine and fentanyl on postoperative pain management following hysterectomy]. AB - OBJECTIVE: To compare the effect of preincisional administration different combination of epidural lidocaine and fentanyl on postoperative analgesia after hysterectomy. METHODS: 75 ASA I-II patients undergoing hysterectomy who were matched in age, weight, and duration of surgical procedure, were randomly allocated to one of the three treatment groups, 25 in each: group I receiving epidural saline as control, group II receiving epidural 2% lidocaine 10 ml combined with 2.5 microg/ml of fentanyl 30 minutes before skin incision followed by 0.125% ropivacaine and 2 microg/ml fentanyl infusion at 9ml/h to the end of surgery, and group III receiving epidural 2% lidocaine 10 ml combined with 2.5 microg/ml of fentanyl and 0.15 mg/ml of droperidol 30 min before incision followed by 0.125% ropivacaine and 2 microg/ml fentanyl infusion at 9ml/h to the end of surgery. Patient-controlled epidural analgesia (0.125% ropivacaine and 2 microg/ml fentanyl.) was given for postoperative pain relief in all groups (5 ml bolus followed by 3 ml/h infusion, PCA dose: 2 ml, lock out time: 10 min.). Postoperative pain intensity at rest and during movement was assessed on a visual analogue scale (VAS) by a blinded observer for 48 h after surgery. Analgesic requirements and side effects were compared among the three groups. RESULTS: In the first 24 hours after surgery, the VAS score at rest reported by the patients in group I was 2.2 +/- 1.5, significantly higher than that in group II (1.4 +/- 0.8, P < 0.05,) and that in group III (1.1 +/- 1.1, P < 0.01), while the VAS score during coughing in group I was 3.0 +/- 1.1, higher than that in group II (2.1 +/- 0.8) and that in group III (1.5 + 1.3,P < 0.01 ). Analgesic requirements in the first 24 hours were 101 +/- 17 ml in group I, 82 +/-9 ml in group II, and 78 ml +/- 9 ml in group III, without significant difference among the groups. However, the analgesic requirement in the second 24 hours was 82 +/- 11 ml in group I, much more than that in group II (76 +/- 5 ml, P < 0.05) and in group III (73 ml +/- 2 ml, P < 0.01). The incidence of nausea, was significantly higher in Group I than in group III (P < 0.05). The incidence rates of pruritus and motor block were higher in group I too. CONCLUSION: The preoperative administration of epidural lidocaine and small dose of fentanyl with droperidol, compared with saline administration, improves the postoperative pain management and reduces postoperative analgesic consumption and side effects. PMID- 12126547 TI - [Carotid endarterectomy, clinical analysis of 43 cases]. AB - OBJECTIVE: To evaluate the important points in carotid endarterectomy. METHODS: Carotid endarterectomy was performed upon 43 cases of carotid artery stenosis under cervical plexus anaesthesia. The operation was performed with shunting between the common carotid artery and the internal carotid artery in 2 cases. RESULTS: The average clamping time of carotid artery was 27 min +/- 7 min. There were one case suffering from transient ischemic attack (TIA), 7 cases suffering from hypertension and 2 cases suffering from light edema of the wound. All of the patients recovered uneventfully. Twenty-six patients who had suffered from TIA before operation felt much better after operation. The follow-up of one month to one year showed a better condition among all patients. CONCLUSION: Cervial plexus anaesthesia helps diagnose stroke during operation. Shunting between the common carotid artery and internal carotid artery helps avoid ischemic brain injury. The effect of operation depends upon complete removal of the residual tunica intima, complete exsufflation, and control of hypertension postoperationally. PMID- 12126548 TI - [Effect of recombinant human growth hormone on postoperative fatigue syndrome in patients after cardiac operations]. AB - OBJECTIVE: To observe the effect of recombinant human growth hormone (rhGH) treatment on postoperative fatigue syndrome (POFS) in patients after cardiac surgery under cardiaopulmonary bypass (CPB). METHODS: 60 patients undergoing open heart surgery were randomly divided (double blind trial) into two groups : group A (injected with rhGH, 4.5 U Bid for 5-7 days) and group B (administrated with placebo). The patients were asked to score their sense of fatigue (Christensen scores). Their 24-hour urine was collected to calculate the daily nitrogen balance. The serum growth hormone (GH) and plasma proteins were detected. RESULTS: In the 4(th) day after operation, the level of fatigue was increased significantly in both groups, however, being lower in group A then in group B, and returned to the pre- operative level in the 8(th) postoperative day in group A (P < 0.01 ). The serum GH was similar in both groups before operation and was significantly increased on the first, fourth, and eighth days, especially the first day, postoperatively in group A (P < 0.01). The plasma total protein and albumin were similar in both groups before operation and on the first and fourth days postoperatively. On the 8(th) day postoperatively the plasma TP and albumin were significantly higher in group A than in group B (P < 0.05 and P < 0.01 respectively). The plasma retinol binding protein (RBP), prealbumin (PA), and transferrin (TFN) were similar in both groups before operation, on the first day after operation the plasma PA and TFN were lower than before operation in both groups. On the first postoperative day, the plasma RBP was higher in group A than in group B (P < 0.05), on the fourth day after operation the plasma PA and TEN were significantly higher in group A than in group B (P < 0.01). The time needed to wean from mechanical ventilation was 15.44 +/- 5.66 hr and 22.66 +/- 10.66 hr in groups A and B respectively (P < 0.01). In group A four patients established positive nitrogen balance on the second postoperative day and all patients in group B failed to obtain positiver nitrogen balance till the seventh postoperative day (P < 0.01). CONCLUSION: 1) Application of exogenous rhGH improves the symptoms of POFS, reduce the duration of POFS, and shorten the time needed to wean from mechanical ventilation, elevate plasma albumin, PA, TFN and RBP levels, and improve negative nitrogen balance. 2) The acute phase plasma proteins (such as PA, TFN, and RBP) are reliable biochemical indicators of POFS. PMID- 12126549 TI - [Specific humoral immune response against B-cell lymphoma elicited by mixed immunoglobulin fragments]. AB - OBJECTIVE: To explore the feasibility of construction of the universal nucleic acid vaccine against B-cell lymphoma. METHODS: RT-PCR was employed to obtain the immuncy losulin heavy chain variable region (IgHV1) gene fragments of the Namalwa cell, normal fetal umbilical cord blood and adult peripheral blood. Then these RT PCR products were cloned into the eukaryotic expression vector pcDNA3.0 and sequenced. Six plasmids that had high identities with the germ line genes were mixed to serve as vaccines. Eighteen Balb/c mice were divided randomly into three groups and were immunized respectively with the six mixed plasmids, the specific IgHV1 fragment of the Namalwa cell and the blank plasmid pcDNA3.0. 100 microg plasmid and 5 microg hIL-6 per mouse were injected into the muscle, 3 times in 4 weeks. Western blotting and indirect immunofluorescence staining were used to assess the humoral immune responses against the Namalwa cell. RESULTS: The specific humoral immune responses against the Namalwa lymphoma cell could be induced in both the IgHV1-mix group and the Na-IgHV1 group, and the antibodies could recognize the nature antigenic determinants in the surface of the Namalwa cell. The antibodies could be detected since the fourth week after the first immunization, and reached the climax at the sixth week. There was no significant difference of the titers of the antibodies between the two groups. The antibody couldn't be found in the negative control group. CONCLUSION: The mixed IgHV plasmids can be used as the universal nucleic acid vaccine against the B-cell lymphoma which expresses the same IgHV1 family genes. PMID- 12126550 TI - [Expressions of Ca(2+)/calmodulin-dependent protein kinases cascades in hippocampi of rats with posttraumatic stress disorder-like behavior]. AB - OBJECTIVE: To explore the neurobiological bases in the pathogenesis of lasting emotional behavioral disorders following posttraumatic stress disorder (PTSD). METHODS: 224 male Wistar rats were divided randomly into 3 groups. Group SE included 88 experimental rats the hippocampi of which underwent electrode implantation and subconvulsive stimulation by constant pulsating current of 100 microA with intratrain frequency of 16 Hz, pulsating duration of 1 ms, train duration of 10 s, and interstimulus interval of 7 min 8 times per day for 5 days so as to induce PTSD-like behavior. Group CE included 88 rats the hippocampi of which underwent electrode implantation and without stimulation to be used as controls. Group NC included 48 rat used as normal controls. Twelve, twenty-four, forty-eight, and seventy two hours after subconvulsive stimulation a certain number of rats in the three groups were killed and their hippocampi were taken. The intracellular free calcium, free calmodulin (CaM), and total CaM, Ca(2+)/CaM dependent kinase II alpha(CaMKIIalpha) and IV (CaMKIV) in hippocampi were examined by fluorescence spectrophotometer, flow cytometry, and Western blotting respectively. RESULTS: The intracellular free calcium level in the experimental rats reached the peak 24 hours after subconvulsive stimulation (487.34 +/- 117.93 nmol/L, P < 0.01), and was still elevated 72 hours after the stimulation (289.46 +/- 69.45 nmol/L, P < 0.05). while the mean channel fluorescence of intracellular free CaM decreased remarkably synchronously (1.46 +/- 0.36, P < 0.01, and 2.53 +/ 0.62, P < 0.05, 24 hours and 72 hours after the stimulation respectively). The expression of total CaM 48 hours after the last stimulation in hippocampi of Group SE rats was significantly elevated (P < 0.01), as well as the expression of CaMKIV (P < 0.05). However the expression of CaMKIIalpha was markedly decreased (P < 0.01) 48 hours after-stimulation. CONCLUSION: The lasting dysfunction of Ca(2+)-CaM-CaMKIIalpha/CaMKIV signaling cascades in hippocampus may play an important role in the long-term neuropsychological sequelae in rats with PTSD like behavior. PMID- 12126551 TI - [Gene transfer of murine Flt3 ligand mediated by adenoviral vector efficiently induces growth inhibition of murine liver cancer]. AB - OBJECTIVE: To observe the in vivo therapeutic effects of murine Flt3 ligand (mFL) mediated by recombinant adenoviral vector on murine liver cancer. METHODS: Murine liver cancer cell line Hepal-6 was infected with adenovirus in vitro. The infection efficacy was measured by green fluorescence protein (GFP) expression and the amount of mFL in supernatant was measured by ELISA 48 hrs following infection of Hepal-6. AdmFL, Ad-null, and PBS were added into the culture of Hepal-6 cells, the number of cells was counted every other day for 14 days. A murine liver cancer model was established by subcutaneous inoculation of Hepal-6 cells. A single dosage of 1 x 10(9) expression forming unit (efu) of Ad-mFL, Ad null or PBS was injected intratumorally. The tumor volume and survival rate were measured twice a week. Twenty days after treatment of adenoviral vectors, three treated mice with their tumor disappearing were killed and their spleen was taken. The splenocytes from these tumor free mice were adoptively transferred to naive mice to whom Hepal-6 cells were inoculated 3 days thereaftter and then the tumor volume was measured once a week.for 4 weeks. 38 days after administration of adenoviral vectors, tumor free animals were rechallenged by parental Hepal-6 cells or syngenic EL4 lymphoma cells at the opposite sites of the original inoculation sites, and the tumor volume was also measured once a week for 4 weeks. RESULTS: Adenoviral vector efficiently infected Hepal-6 cells in intro, and lead to the secretion of high levels of mFL protein (80.5 +/- 7.3 ng/10(6)/24 h) in the supernatant. The growth of Hepal-6 tumor was significantly inhibited by one single intratumoral administration of Ad-mFL in 90% of the treated mice, and the tumor gradually grew in the two other groups. Two of the 7 mice (30%) in PBS group died in 17 days, 14% still lived in 37 days, and all died within 45 days. The mice in the Ad-null group began to die since the 21(st) day, only 11% of them were still alive in 60 days. All mice in the Ad-mFL group were alive at the 60(th) day after tumor implantation. Adoptive transfer of splenocytes to the animals receiving Ad-mFL treatment protected effectively them against a subsequent challenge with the identical tumor cells. Rechallenge of the Ad-mFL cured mice with the parental Hepal-6 cells resulted in complete inhibition of tumor growth. The growth of inoculated EL4 lymphoma cells gradually grew equally in the controls and the experimental mice. CONCLUSION: FL gene transfer mediated by recombinant adenoviral vector has potent therapeutic effects on Hepal-6 liver cancer, and develops long-lasting specific antitumor immunity, which may become a potent cancer gene therapy candidate for further clinic application. PMID- 12126552 TI - [Regulated expression of insulin gene in non-beta cell]. AB - OBJECTIVE: To engineer 293 cells to an artificial beta cell line which secrets insulin in response to stimulation by doxycycline. METHODS: A recombined expression vector, pTRE2mINS, which contained both of the tetracycline response element and proinsulin gene, was constructed. This vector was co-transfected with plasmid pTK-Hyg encoding hygromycin in the tet293 cells, which express the reverse tetracycline-controlled transactivator stably. Following hygromycin screening, the survived cells expressing insulin were selected and enriched. Then, doxycycline one of tetracycline derivatives, was used to control the expression of insulin in these cells. At the levels of mRNA and protein, the regulating effect of doxycycline in culture medium on the expression of proinsulin gene was estimated respectively with Northern blotting, RT-PCR, and radioimmunoassay. RESULTS: Of the 28 hygromycin resistant cell strains, we selected one cell strain secreting insulin not only automatically, but in response to stimulation by doxycycline was selected. The cells secreted insulin at the rate of 9.7 U/24 h/1.0 x10(6) cells without doxycy cline treatment, and the amount of secreted insulin increased to 241.0 U/24 h/1.0 x 10(6) cells (25 fold) in the presence of doxycycline(1 000 ng/ml). I insulin secretion was induced by doxycycline in a dose dependent manner. CONCLUSION: Human proinsulin gene was transfected successfully and expressed efficiently in 293 cells, and the expression was modulated by tetracycline and its derivatives, improving the accuracy, safety, and reliability for the gene therapy of diabetes mellitus. PMID- 12126553 TI - [Changes of proinflammatory cytokines and their receptors in serum from patients with pulmonary tuberculosis]. AB - OBJECTIVE To investigate the characteristics and clinical value of serum tumor necrosis factor-alpha (TNF-alpha) and its receptor (sTNF-R), interleukin 1beta(IL-1beta) and its receptor(IL-1R), interleukin-6(IL-6) and its receptor(IL 6R) in patients with pulmonary tuberculosis, and to evaluate their role in the immunopathogenesis of tuberculosis. METHODS The serum levels of TNF-alpha, sTNF-R Iota IL-1beta,IL-1R, IL-6 and IL-6R were measured using the sandwich ABC-ELISA method in 41 cases of active tuberculosis, 21 cases of inactive tuberculosis and 20 normal controls. The serum levels of the cytokines in 17 cases of active tuberculos is were followed. RESULTS The serum levels of TNF-alpha sTNF-RIota IL-1beta,IL-1R, IL-6 IL-6R and the TNF-alpha/sTNF-RIota ratio were significantly higher in both the active and the inactive tuberculosis groups than those in normal controls (P <0.01 approximately 0.05). The TNF-alpha sTNF-R Iota IL-1 beta, IL-1R, IL-6 IL-6R levels and the TNF-alpha/sTNF-R Iota ratio in the active tuberculosis group were significantly higher than those in the inactive tuberculosis(P <0.01 approximately 0.05). The serum levels of TNF-alpha sTNF-R Iota, IL-1beta and IL-6 and the TNF-alpha,/sTNF-R Iota ratio were significantly lower in cavernous tuberculosis than those in non- cavernous tuberculosis (P < 0.01 approximately 0.05). After 2 months' antituberculosis treatment, the serum levels of TNF-alpha,sTNF-R Iota IL-1 beta, IL-1R,IL-6, IL-6R and the TNF alpha/sTNF-R Iota ratio in 15(15/17) cases were significantly lower than those before treatment(P < 0.01 approximately 0.05). CONCLUSIONS TNF-alpha, IL-1 beta, IL-6 and their receptors may be involved in the immunopathogenesis of tuberculosis. Measuring the serum levels of proinflammatory cytokines and their receptors may be useful in evaluating the activity, the clinical pattern, and the prognosis of the disease and monitoring the clinical effect of antituberculous therapy. PMID- 12126554 TI - [Transbronchial needle aspiration in diagnosing mediastinal and hilar tuberculous lymphadenopathy]. AB - OBJECTIVE: To study the application of transbronchial needle aspiration(TBNA) in the diagnosis of mediastinal and hilar tuberculous lymphadenopathy. METHODS: Brochoscopic examination was carried out in 24 patients with computed tomography confirmed mediastinal and hilar lymphadenopathy, but without signs of lung or airway tuberculosis. TBNA was performed and at least two prominent lesions were biopsied. Cytological needle aspiration was done for all diseased sites, while some lesions were also sampled by histological needles. RESULTS: Pathologic diagnosis was made in 18 patients showing tissue necrosis and granuloma, cases being acid fast positive. There were no serious complications. CONCLUSION: TBNA is a useful and convenient technique in the diagnosis of mediastinal and hilar tuberculous lymphadenopathy. PMID- 12126555 TI - [Pharmacodynamics and pharmacokinetics of domestic fixed-dose combination of antituberculosis drugs]. AB - OBJECTIVE To study the pharmacodynamics and pharmacokinetics of domestic fixed dose of antituberculosis drugs and to evaluate its quality and activity against Mycobacterium tuberculosis both in vitro and in vivo. METHODS The MIC was determined by the tube doubling dilution method, and the effect of the drugs was assessed by half survival time of the mice. A single oral dose of domestic and imported fixed-dose combination of antituberculosis drugs was given to healthy volunteers, and the drug concentration in serum was determined by HPLC. The pharmacokinetic parameters and the relative bioavailability were calculated. RESULTS The MIC of each composition in the compound (INH, RFP, PZA) against Mycobacterium tuberculosis was lower than that of each composition used by single dose. In a murine tuberculosis model, the antituberculosis activity of this compound drug was superior to that of each agent used alone with the same dose. No significant difference was found as compared to the imported drug, Refater; The major pharmacokinetic parameters of the domestic and the imported drugs, t (1/2), C (max), AUC, and t(max), were not significantly different. Statistical analysis showed the two formulations were bioequivalent. CONCLUSION The three compositions in the combination had synergistic effect, and the domestic and the imported drugs were bioequivalent. PMID- 12126556 TI - [The pathogenic role of macrophage migration inhibitory factor in acute respiratory distress syndrome]. AB - OBJECTIVE To investigate the expression and pathogenic role of macrophage migration inhibitory factor( MIF) in human acute respiratory distress syndrome(ARDS) METHODS The serum level of MIF in ARDS patients and normal persons were measured by ELISA method. Peripheral blood mononuclear cell (PBMC) MIF expression was determined by flow- cytometry. The expression of MIF mRNA and protein in the lung tissues were detected by using double immuno histochemistry labeling and in situ hybridization. RESULTS The serum level of MIF increased significantly in ARDS patients as compared with normal persons (P < 0.01). The percentage of PBMC MIF expression was higher in ARDS patients than in normal controls (P < 0.01). In situ hybridization and immunohistochemistry showed undetectable or weak MIF mRNA and protein expression in normal lungs. In contrast, there was marked upregulation of MIF mRNA and protein expression in the ARDS lungs. In ARDS macro phages infiltrated the alveolar space and interstitium, most of which also expressed MIF. Infiltrating macrophages were almost restricted to the areas of severe tissue damage. The MIF expression level showed a strong correlation with the number of infiltrating macrophages. CONCLUSIONS The serum level of MIF and PBMC MIF expression increased in ARDS patients with enhanced pulmonary MIF expression and macrophage infiltration, which suggests that MIF plays a pivotal role in the pathogenesis of ARDS. PMID- 12126557 TI - [A study on viral infections in patients with chronic obstructive pulmonary disease]. AB - OBJECTIVE: To study the relation between viral infection and chronic obstructive pulmonary disease (COPD), and the role of viral infection in the pathogenesis of COPD. METHODS: (1)Serum samples from 91 patients with acute exacerbation of COPD (Group A), 42 patients with stable COPD (Group B) and 25 healthy subjects (Group C) were tested for specific IgM and IgG for respiratory syncytial virus (RSV) herpes simplex virus type 1 (HSV-1) parainfluenza virus(PIV),adenovirus (ADV) and cytomegalovirus (CMV) with an indirect enzyme linked immunosorbent assay (ELISA). (2) T lymphocyte subsets (CD(3) CD(4) and CD(8)) were studied by flow cytometry in patients with IgM positivity (Group D n = 28) and patients in Group C. RESULTS: (1) The positive rates of IgM were significantly different (P < 0.001) in group A (12%, 8%, 6%, 2%, 3% for RSV HSV-1 PIV ADV and CMV) as compared to group B (0%, 0%, 0%, 0%, 0%) and group C (0%, 0%, 0%, 0%, 0%). There was no significant difference in the positive rate of IgG between group A (47%, 41%, 12%, 14%, 10%) and group B (43%, 33%, 12%, 7%, 7%, P > 0.05), but the positive rates of IgG in group A and group B were significantly different (P < 0.01) as compared to group C (0%, 8%, 0%, 0%, 0%) (2)In group D the expression of CD(4) and CD(4)/CD(8) were significantly lower, the expression of CD(8) was significantly higher (P < 0.01) than those in group C but the expression of CD(3) was not significantly different (P > 0.05). CONCLUSION: Viral infection was common in acute exacerbation of COPD, and related to the pathogenic mechanism of COPD. PMID- 12126558 TI - [Relationship between airway remodeling and inflammatory mediators and cytokines in sputum in patients with bronchial asthma]. AB - OBJECTIVE: To compare the thickness of subepithelial reticular basement membrane (BM) in asthmatic patients and healthy subjects and to compare the BM thickness to inflammatory mediators and cytokines. METHODS: Fiberoptic bronchoscopy with bronchial biopsy was performed on 20 stable asthmatics and 10 normal subjects. The subepithelial reticular BM thickness was measured. Levels of eosinophil cationic protein (ECP), interleukin 5 (IL-5 ), and tumor necrosis factor alpha (TNF alpha ) in sputum were measured by immunoassay. The relationship between ECP, IL-5, TNF alpha and BM thickness was estimated by the spearman rank correlation. RESULTS: The mean BM thickness was significantly higher in asthmatic patients (10.1 +/- 2.6) microm than in normal subjects (4.4 +/- 1.2) microm (P < 0.005). In asthmatic patients, the ECP (144 +/- 80) microgram/L and the IL-5 (17 +/- 4) microgram/L levels in sputum were positively correlated with BM thickness (r = 0.569, P < 0.005 and r = 0.466 P < 0.005, respectively ). Sputum TNF-alpha (53 +/- 36) microgram/L showed no significant correlation with BM thickness (r = 0.254 P > 0.05). CONCLUSIONS: Airway remodeling is a distinctive and characteristic pathologic finding of asthma. Airway remodeling is related to levels of sputum ECP and IL-5, but not that of TNF-alpha. PMID- 12126559 TI - [Regulation of the expression of pulmonary arterial collagen by protein kinase C and breviscapine in chronic hypoxic rats]. AB - OBJECTIVE: To investigate the effects of protein kinase C and breviscapine on the expression of pulmonary arterial collagen in chronic hypoxic rats. METHODS: Thirty-six rats were randomly divided into three groups: control group (A), hypoxic group(B),hypoxic + breviscapine(bre.)group (C). The ultrastructure of pulmonary arterioles was observed by electron microscope; the PKC activities of lung tissues were measured by radioactivity; the expression of PKC and collagen I and III in arterioles was observed by immunohistochemistry; the expression of procollagen I and III mRNA in arterioles was observed by in situ hybridization. The averages of integral light density( A )of PKC, collagen I and III and procollagen I and III mRNA in pulmonary arterioles were detected by image analysor and their relative contents were calculated. RESULTS: (1) The mean pulmonary arterial pressure (mPAP) and the weight ratio of RV to LV + S in group B were higher than those in group A(P < 0.01); the mPAP and the weight ratio of RV to LV + S in group C were lower than those in group B (P < 0.01). (2)Electron microscopy showed breviscapine could inhibit the deposition of collagenous fibers in pulmonary arterioles induced by hypoxia.(3) The total,cytosolic and particulate fractions of PKC activity and the ratio of particulate fraction to total PKC activity in group B were higher than those in group A(P < 0.01); the total,cytosolic and particulate fractions of PKC activity and the ratio of particulate fraction to total PKC activity in group C were lower than those in group B (P < 0.05). (4) The A values of PKC, collagen I and procollagen I mRNA in pulmonary arterioles were higher in group B than in group A(P < 0.01),and the A values of PKC, collagen I and procollagen I mRNA in pulmonary arterioles were lower in group C than in group B(P < 0.01); the differences in the A values of collagen III and procollagen III mRNA in pulmonary arterioles were not significant among the three groups (P > 0.05).(5)There were good correlations between the PKC activity of the lung tissues and the A values of collagen I and procollagen I mRNA in pulmonary arterioles(P < 0.05),and between the A values of PKC in pulmonary arterioles and those of collagen I and procollagen I mRNA in pulmonary arterioles(P < 0.01). CONCLUSIONS: The PKC signal pathway regulates the expression of pulmonary arterial collagen in chronic hypoxic rats which may play an important role in the pathogenesis of pulmonary hypertension and structural remodeling of pulmonary arterials breviscapine can lower hypoxic pulmonary hypertension by inhibiting the effect of PKC and decreasing the expression of pulmonary arterial collagen. PMID- 12126560 TI - [Protective effect of Ginkgo biloba extract on acute lung injury induced by lipopolysaccharide in D-galactose aging rats]. AB - OBJECTIVE: To investigate the effect of Ginkgo biloba extract (GBE) on acute lung injury(ALI) induced by lipopolysaccharide (LPS) in aging rats. METHODS: The rats were randomly divided into two parts: six rats served as normal controls; 18 rats were used for producing the aging animal model ( D-gal 50 mg/kg body weight was injected intraperitoneally, once a day for 6 weeks). The aging rats were then randomly divided into 3 groups: 6 rats as the aging control group; another 6 as the LPS treated aging group (LPS,5 mg/kg body weight intravenous injection); and the third as the GBE+LPS group (6 rats, GBE was started 7 days before the experiment,given once a day via the esophagus, the amount of flavone glycosides administered being 8 mg/kg body weight, and on the day of experiment, one dose of GBE was given 2 hours before LPS administration ). The samples were collected 2 hours after LPS or saline administration. RESULTS: Compared with normal controls, the SOD activity in red cells and the Na(+) -K(+) -ATPase activity in the lung tissue decreased markedly (all P < 0.05), but the LDH activity increased (P < 0.05) in the aging rats. ALI was observed in the aging rats 2 hours after LPS administration. Compared with the aging control, in the LPS treated rats, there were more inflammatory cells in the lung tissue; protein content in bronchial alveolar lavage fluid (BALF) and the pulmonary permeability index (PPI) increased significantly (all P < 0.001); LD, MDA, NO(2) (-)/NO(3)(-), ET-1 and TNF-alpha content and LDH activity in blood, and MPO activity in lung tissue all increased significantly. On the contrary, SOD activity in red cells and Na(+) -K(+) -ATPase activity in lung tissue decreased markedly (P < 0.05, P < 0.01). These changes, except SOD, were markedly attenuated in the GBE+LPS rats. CONCLUSIONS: The anti oxidant activity was decreased in D-gal-induced aging rats. Intravenous administration of LPS caused acute lung injury in aging rats. GBE had protective effect on ALI induced by LPS. PMID- 12126561 TI - [Prophylactic effect of retinoic acid on chronic obstructive pulmonary emphysema in rats]. AB - OBJECTIVE: To evaluate the effect and molecular mechanism of retinoic acid (RA) in preventing experimental pulmonary emphysema in rats. METHODS: Thirty-six Wistar rats were randomly divided into three groups, the normal group(A), the model group (B) and the RA treated group (C). Rats in group B group were exposed to cigarette smoke, and group C was treated with RA. Pulmonary function indices and histopathological changes were evaluated. The enzymatic activity of gelatinases was measured by ELISA, and the expression of gelatinases and the proliferating cell nuclear antigen (PCNA) was examined by immunohistochemical method. RESULTS: Compared with group A, group B presented significant differences in FEV(0.3),FEV(0.3)/FVC and FRC [(5.1 +/- 0.4) ml vs (6.0 +/- 0.3) ml, (71 +/- 10) ml/s vs (87 +/- 3) ml/s, (7.2 +/- 2.2) ml vs (2.9 +/- 1.1) ml (P < 0.01)];the enzymatic activity [(1.06 +/- 0.23)ng/ml vs (0.53 +/- 0.17)ng/ml, (0.960 +/- 0.230)ng/ml vs (0.300 +/- 0.090)ng/ml] and the expression of gelatinases were also markedly elevated(P < 0.01). In comparison with group B, the pulmonary function indices in group C [(5.2 +/- 0.4)ml, (81 +/- 5) ml/s, (6.1 +/- 2.7)ml/s] were improved (P < 0.05); the enzymatic activity [(0.83 +/- 0.23)ng/ml, (0.570 +/ 0.010)ng/ml] and the expression of gelatinases were decreased, while the PCNA index staining (93 +/- 4 vs 53 +/- 6) was increased (P < 0.01). CONCLUSION: RA can prevent the development of pulmonary emphysema in rats. PMID- 12126562 TI - [Effect of adrenomedullin on the regulation of pulmonary arterial pressure in hypoxic rats]. AB - OBJECTIVE: To evaluate the effect of adrenomedullin (ADM) on pulmonary circulation and the change of ADM in plasma and lung tissue from rats with hypoxic pulmonary hypertension. METHODS: Fifty Wistar rats were divided into the control group (10 rats) and the hypoxic group (40 rats). The animal model of pulmonary hypertension was established by exposing the rats to normobaric hypoxic conditions for 3 weeks; Mean pulmonary arterial pressure (mPAP) was measured by right cardiac catheterization, and mean systemic blood pressure (mSBP) was measured by left femoral catheterization. The thickness of pulmonary arterioles was measured by a computerized image analyser. The level of ADM in plasma and lung tissue was measured by radioimmunoassay. We ADM was administered in doses of 0.5, 1.0, 2.0 nmol/kg respectively to 30 hypoxic rats and the changes in mPAP and mSBP to ADM was evaluated. RESULTS: Rats exposed to hypoxia developed pulmonary hypertension, the mPAP in the control group being (16 +/- 3) mm Hg and in hypoxic group being (30 +/- 4) mm Hg, and the difference was significant (P < 0.01). The hypoxic rats developed significantly thickened pulmonary arterioles. The plasma level of ADM was (288 +/- 24) pg/ml in the hypoxic rats and (168 +/- 25) pg/ml in the control group the difference being significant (P < 0.01). The level of ADM in the lung homogenates from the hypoxic group was (2 319 +/- 238) pg/g and that from the control group was (1 153 +/- 127) pg/g and the difference was significant (P < 0.01). The ADM levels had a positive correlation with the mPAP (gamma = 0.567 and 0.612 P < 0.01, respectively). Administration of exogenous ADM reduced the mPAP in a dose-dependent manner in hypoxic rats, and the effect lasted 5 approximately 15 minutes. CONCLUSION: ADM has a relaxing effect on pulmonary circulation. The change of ADM in plasma and lung tissue may serve as a compensatory mechanism in maintaining the stability of pulmonary circulation in hypoxic condition. PMID- 12126563 TI - [Relationship between human parvovirus B19 infection and spontaneous abortion and congenital heart disease in China]. AB - OBJECTIVE: To explore the status of human parvovirus B19(HPV B19) infection to the fetus in China and the relationship between HPV B19 infection and spontaneous abortion,and congenital heart disease (CHD). METHODS: Nest polymerase chain reaction (PCR) and in situ hybridization (ISH) were used to detect HPV B19-DNA in the tissues of spontaneous abortion and cardiac muscle of CHD, enzyme linked immunosorbent assay (ELISA) to detect HPV B19-IgM. RESULTS: (1) The positive rate of HPV B19-DNA was 25.8%(47/182) in spontaneous abortion, comparing with artificial abortion 5.0%(2/40). (2) The positive rate in cardiac muscle tissues of CHD in autopsy and biopsy were 17.2%(5/29) and 18.9%(7/37), respectively. The total rate was 18.2% (12/66), but the positive case was not found in control group (0/30). The HPV B19 gene were found locating in the nucleus of cardiac cell by ISH. The HPV B19-IgM was negative in the blood in all patients when the HPV B19-DNA positive in the cardiac cell of biopsy. CONCLUSION: (1)There may be a relationship between HPV B19 infection and spontaneous abortion and CHD. (2) Two spontaneous abortion specimens from one person in the different period were both positive. It indicated that, there was a persistent infection. It may be one cause in frequently abortion. PMID- 12126564 TI - [Perinatal outcome of twin pregnancies conceived by assisted reproductive techniques and those conceived spontaneously]. AB - OBJECTIVE: To investigate perinatal outcomes of twin pregnancies conceived by assisted reproductive techniques and those conceived spontaneously. METHODS: The pregnant complications, delivery situation and neonatal outcomes of 104 twin pregnancies conceived by assisted reproductive techniques were compared to 173 spontaneous ones. RESULTS: (1) The women of twin pregnancies conceived by assisted reproductive techniques were (31.2 +/- 3.7) years old while the spontaneous ones were (27.8 +/- 3.5) years old. There was significant difference between two groups (P < 0.05). (2) There were also significant differences in the incidence of premature delivery and gestational diabetes mellitus or impaired glucose tolerance(P < 0.01) between two groups. There were 70 cases (67.3%) premature delivery in assisted reproduction group, while 78 cases (45.1%) in spontaneous group. There were 16 cases (15.4%) gestational diabetes mellitus or impaired glucose tolerance in assisted reproduction group, while 4 cases (2.3%) in spontaneous group. (3) Cesarean sections were more frequently performed in assisted reproduction group (79 cases, 76.0%) than in spontaneous group (113 cases 65.3%). (4) There were no statistically significant differences in perinatal deaths, congenital malformations, neonatal asphyxia between two groups (P < 0.05). CONCLUSION: The women of twin pregnancies conceived by assisted reproductive techniques were older than those conceived spontaneously. The incidence of premature delivery, gestational diabetes mellitus or impaired glucose tolerance and cesarean section were higher in assisted reproduction group. However, no statistically significant differences in perinatal morbidity were found between the two groups. PMID- 12126565 TI - [Expression of inducible nitric oxide synthase and macrophage in human placenta of pregnancy induced hypertension syndrome]. AB - OBJECTIVE: To investigate the expression of inducible nitric oxide synthase (iNOS) and macrophage (CD(68)) in the placenta of the patients with pregnancy induced hypertension syndrome(PIH) and the role of them in the pathogenesis of PIH. METHODS: Immunohistochemical technique (SABC) was used for immunostaining and detecting inducible nitric oxide synthase and CD(68) in the placentas of thirty patients of PIH (two patients of mild PIH; seven moderate PIH; twenty-one severe PIH) and twenty-two normal pregnancies (the control group). RESULTS: The level of inducible nitric oxide synthase and CD(68) in the placenta of patients of PIH were (2.9 +/- 0.7) % and (3.0 +/- 0.8) %, respectively and (2.1 +/- 0.8) % and (2.1 +/- 0.7) % in the control group. They were both significantly different (P < 0.01). The levels of inducible nitric oxide synthase in the mild moderate and severe PIH were (2.8 +/- 0.7) % and (3.0 +/- 0.7) %, and the CD(68) were (2.8 +/- 1.0) % and (3.1 +/- 0.8) %, which were not significantly different (P > 0.05). CONCLUSION: The obvious increase of iNOS and macrophage in the placenta of patients of PIH can be observed and iNOS and CD(68) are involved in the immunization and inflammatory reaction. This may be one of the causes for human placental dysfunction and high impedance of fetoplacental circulation in PIH. It may play an important role in the genesis of pregnancy induced hypertension. PMID- 12126566 TI - [Management of prolactinomas during pregnancy]. AB - OBJECTIVE: To evaluate the safety of discontinuing drugs for prolactinoma during pregnancy, and the effect of prolactinoma on pregnancy. METHODS: Thirty-two cases of prolactinoma in pregnancy were enrolled, 22 microadenoma and 10 macroadenoma. RESULTS: Adenoma growing appeared in 6 cases. Among them 5 were macroadenoma, dopamine agonists were then reused and surgery were carried out in 2 of them. The other was one microadenoma, dopamine agonists was reinstituted. No side effect on fetus's weight and appearance was seen. Gestation complication was as normal. CONCLUSIONS: Discontinuing dopamine agonists is safe during pregnancy for microadenoma. The risk of macroadenoma enlargement during pregnancy is high. Regular visal field testing is necessary during pregnancy. Prolactinoma doesn't influence gestation complication and fetus. PMID- 12126567 TI - [Effects of macrophage colony-stimulating factor on production of estradiol and progesterone by human luteinized granulosa cells in vitro and detection of macrophage colony-stimulating factor receptor on human luteinized granulosa cells]. AB - OBJECTIVE: To investigate the effects of macrophage colony-stimulating factor (M CSF) on estradiol and progesterone production by human luteinized granulosa cells in vitro and to detect M-CSF receptor in human granulosa cells. METHODS: Human luteinized granulosa cells (LGC) were isolated from follicular fluid of superovulated infertile patients undergoing intracytoplasmatic sperm injection. Some of the LGC were used for detecting M-CSF receptor by immunocytochemical staining (ABC method). Most of them were cultured with HAM's F-10 medium plus various concentration of M-CSF (0, 10, 25, 50, 100, 250 ng/ml) in the presence or absence of follicle stimulating hormone (FSH). Media were collected at 72 hours after culture and estradiol (E(2)) and progesterone (P) in media were measured by enzyme immunoassays. RESULTS: About 80% of the LGC presented a positive M-CSF receptor staining with the immunochemical signal on cell membrane. After 72 hours culture of LGC, in absence of FSH, the baseline concentration of E(2) and P were (2,185 +/- 189) pmol/L, (3,157 +/- 401) nmol/L respectively; with the increasing dose of M-CSF from 10 to 100 ng/ml, the concentrations of E(2) increased from (2,789 +/- 365)pmol/L to (4,282 +/- 318) pmol/L, and those of P increased from (4,256 +/- 595) nmol/L to (7,789 +/- 828) nmol/L. In presence of FSH (75 IU/ml) the baseline concentrations of E(2) and P were (5,045 +/- 486) pmol/L and (8,667 +/- 923) nmol/L respectively; while the 10 - 100 ng/ml M-CSF was added the concentrations of E(2) and P were from (6,567 +/- 673) to (8,373 +/- 935) pmol/L and from (10,999 +/- 985) to (14,990 +/- 1,158) nmol/L respectively. Thus M-CSF caused a significant dose dependent increase of estradiol and progesterone production (P < 0.05), and M-CSF+ FSH further stimulated higher production of E(2) and P by LGC than M-CSF or FSH alone (P < 0.05). CONCLUSIONS: M-CSF receptor was expressed on membrane of human LGC. M-CSF enhances estradiol and progesterone production by LGC and FSH has an additional or synergetic effect with M-CSF. PMID- 12126568 TI - [Clinical study on 235 cases performed under hysteroscopy combined with laparoscopy]. AB - OBJECTIVE: To evaluate the clinical value of using hysteroscopy combined with laparoscopy in gynecologic disease. METHODS: From January 1995 to January 2001, two hundred thirty five operations were performed under hysteroscopy combined with laparoscopy. This paper focuses on their indications, method and complications as well as management. RESULTS: Among the 235 cases, all of them were performed more than two procedures inside uterine and pelvic cavity. Hysteroscopy underwent on all cases for the lesion of uterine cavity except one changed laparotomy due to heavy bleeding. These hysteroscopy included some complicated operations such as transcervical resection of septum, transcervical resection of adhesion, transcervical resection of embryo bones and IUD pieces embedded endometrium as well as transcervical resection of myoma that the size was bigger than 4.5 centimeter or intramural and broad peduncle one. Seven cases with incomplete uterine perforation and three cases with uterine perforation were found by laparoscopy during hysteroscopic procedures and treated under laparoscopy. In addition, other laparoscopic procedures such as cystectomy, adhesiolysis, myomectomy, hydrotubation, salpingostomy and oophorectomy etc. Were also performed. There were no complications caused by hysteroscopy combined with laparoscopy. CONCLUSIONS: Hysteroscopy combined with laparoscopy can be performed under one anesthesia and managed the lesions in both uterine and pelvic cavity that could not be treated previously with single hysteroscopy or laparoscopy. Hysteroscopy combined with laparoscopy can find out and treat uterine perforation timely and reduce complications. PMID- 12126569 TI - [Study on polymorphism of human leukocyte antigen-DRB1 allele in patients with endometriosis]. AB - OBJECTIVE: To study the possibility of a correlation between occurrence of endometriosis (EM) and polymorphism of human leukocyte antigen (HLA) DRB1 allele. METHODS: Polymorphism of HLA-DRB1 allele were detected by polymerase chain reaction- sequence specific primers (PCR-SSP) testing 40 surgically proven EM patients and 50 normal control. RESULTS: At HLA-DRB1 15 allele, positive frequency of EM group was significant higher as compared with those of the normal control group [45.0% vs 26.0%, P < 0.05, relative risk (RR) = 2.1]. Other HLA DRB1 allele positive frequency of EM group and control group had no significant difference. But for Polymorphism of DRB1 * 15 allele, positive frequency of EM group and control group had no significant difference [1501 : 35.0% vs 20.0%; 1502 : 10.0% vs 6.0%; P > 0.05]. CONCLUSIONS: The occurrence of EM may be associated with the presence of HLA-DRB1 allele, but hasn't associated about polymorphism of DRB1* 15 allele. HLA-DRB1 * 15 maybe one of some factors for the pathogenesis of endometriosis. PMID- 12126570 TI - [BRCA2 gene mutation detection in hereditary ovarian cancer tissues]. AB - OBJECTIVE: The aim of this study was to detect the mutation of BRCA2 gene in hereditary ovarian cancer tissues to probe the clinical significance of it. METHODS: DNA was abstract from paraffin embedded tissues archived in pathology department. A pair of primer located on 11 exon was used to detect the 6174 delT mutation,BRCA2 gene mutation in 12 hereditary ovarian cancer patients and 15 sporadic ovarian cancer patients were screened by polymerase chain reaction single strand conformation polymorphism analysis with DNA non- isotopic silver staining methods. RESULTS: Two of 12 hereditary ovarian cancer victims were found carrying mutation of BRCA2 gene. The mutation type was 6174del T in the 11 exon of BRCA2 gene. No mutation of BRCA2 gene on this site was found in 15 sporadic ovarian cancer patients. CONCLUSION: BRCA2 gene mutation was closely associated with the carcinogenesis and development of hereditary ovarian cancer, but had no relationship with sporadic ovarian cancer. PMID- 12126571 TI - [Experimental study on adenovirus-mediated transfer of p53 or adenovirus- mediated transfer of p53 combined with radiotherapy for human cervical cancer cell line HeLa cells]. AB - OBJECTIVE: To evaluate the therapeutic efficiency of adenovirus-mediated transfer of p53(Ad-p53) or Ad-p53 combined with radiotherapy for human cervical cancer cell line HeLa cells. METHODS: HeLa cells were either transfected with Ad-p53 alone or with Ad-p53 combining with radiotherapy. Reverse transcription polymerase chain reaction(RT-PCR), the cell growth, morphological changes, cell cycle, apoptosis and molecular changes were measured using cell counting, microscopy, flow cytometry, DNA ladder, and in vivo therapy experiments to evaluate the therapeutic efficiency of the regimens. RESULTS: Ad-p53 could be transfected into HeLa cells and p53 mRNA could be expressed more in the targeted HeLa cells. Ad-p53 transfer had strong therapeutic effects to HeLa cells in vitro and in vivo (the inhibition rates were 61.8% and 54.8%, respectively), moreover, the combined administration of Ad-p53 and radiotherapy had stronger therapeutic effects in vitro and in vivo (the inhibition rate was 85.5% in vitro, and the inhibition rate was 77.4% in vivo). The inhibition rate by radiotherapy alone was 44.6% in vitro and 58.4% in vivo. Massive apoptosis of HeLa cells was induced by these regimens. Cell cycle analysis demonstrated that all these regimens could arrested HeLa cells in G(2)-M. CONCLUSIONS: After introduced into HeLa cells, Ad p53 shows therapeutic efficiency for HeLa cells both in vitro and in vivo. Moreover, Ad-p53 could enhance therapeutic efficiency for HeLa cells when combined with radiotherapy. PMID- 12126572 TI - [Role of positron emission tomography in detecting recurrent epithelial ovarian carcinoma]. AB - OBJECTIVE: To investigate the efficiency of positron emission tomography (PET) with (fluorine-18)-2-deoxyglucose ((18)FDG) for detecting recurrent epithelial ovarian carcinoma. METHODS: Fifteen (18)FDG-PET scanning were performed on 13 patients, who was clinically free of disease after optimal cytoreductive surgery and sufficient chemotherapy, with suspicion of recurrence. Ten second-look or re debulking operation were given after PET scanning. RESULTS: In all 8 cases with and 1 without elevated serum CA(125), PET demonstrated sites of increased (18)FDG uptake, which correlated well with surgical-pathologic findings (100%). Computed tomography scan and B ultrasonography each detected 1 recurrence. One of the 6 patients with negative PET scan results underwent a second-look operation, which found no malignancy. The other 5 were closely followed up for 11 to 13 months. No sign of recurrence was noticed within 8 months. One patient received her second PET scanning after the serum CA(125) began to elevate in the 9th month. Recurrent tumor was found by the second PET scan and confirmed by the operation after that. CONCLUSIONS: PET is accurate in detecting the recurrent epithelial ovarian carcinoma. As a non-invasive method, PET might be a fairly effective tool for monitoring and locating the recurrence of ovarian carcinoma. PMID- 12126573 TI - [Clinicopathologic analysis of aggressive angiomyxoma in the pelvis and vulva of 8 cases]. AB - OBJECTIVE: To study the clinical and pathologic features of aggressive angiomyxoma (AAM) and its occurrence, treatment and prognosis. METHODS: The 8 cases of AAM were analyzed retrospectively, among which 6 cases in vulva, 2 cases in perineum, vagina and pelvis. As well we performed immunohistochemical studies. RESULTS: The neoplastic cells were at least focally immunoreactive for vimentin (7/7), smooth muscle actin (7/7), factor 8 (7/7), proliferating cell nuclear antigen (4/7), estrogen (3/7) and progesterone (4/7) receptors. All of the examined tumors were negative for S100 protein and cluster designation 68. CONCLUSIONS: AAM is an uncommon mesenchymal tumor that preferentially involves the pelvic and perineal regions of females. In immunohistochemical investigation the neoplastic cells of AAM exhibit fibroblastic and myofibroblastic features. They are characterized by slow growth, infiltration of adjacent structures, absence of distant metastases, a tendency to recur locally and a fairly good prognosis. Treatment should consist of wide surgical excision, as complete as technically possible. PMID- 12126574 TI - Relaxins: lessons and limitations. PMID- 12126575 TI - A realistic approach to managing patients with fibromyalgia. AB - Fibromyalgia syndrome is common and variable in impact, with some patients having a milder and shorter duration of symptoms and others suffering significant and prolonged pain. Disability also varies. It is thought that the syndrome arises from a disordered neurophysiology that, through links to central control inputs, involves emotions, thoughts, and cognitions. Social and psychological sequelae contribute to and result from this process. The biopsychosocial model of disease epitomizes fibromyalgia. Although management may be difficult at times, and much needs to be done, the growing appreciation of strategies that use this described model and the knowledge of the potential reversibility of the syndrome are resulting in improved outcomes. PMID- 12126580 TI - An association of silicone-gel breast implant rupture and fibromyalgia. AB - Silicone-gel breast implant rupture is common. Silicone-gel from ruptured implants may escape the scar capsule that forms around breast implants and become "extracapsular silicone." Our previously published study found that women with extracapsular silicone gel were at higher risk of reporting that they were diagnosed with fibromyalgia. There has been a limited number of studies addressing this association in the literature. Some studies addressing the issue of silicone breast implants and connective tissue disease specifically exclude patients with fibromyalgia from the sample or do not include the syndrome in the analysis. Case series describing fibromyalgia in patients with implants have been published, but many of these papers lack information on extracapsular silicone and are not representative because the patients are typically from referral populations. In addition, most studies do not have control groups of women without implants for comparison or do not distinguish between saline and silicone implants. Additional observational studies of women from nonreferral populations are necessary to validate an association. These studies should provide information on how the rupture is diagnosed, state whether the rupture extended beyond the capsule, and provide an appropriate control group for comparison. The findings from such studies may be important to physicians as they describe potential risks associated with implants to their patients. These findings should also be important for regulatory decision making on silicone-gel breast implants. PMID- 12126581 TI - Evidence of involvement of central neural mechanisms in generating fibromyalgia pain. AB - Fibromyalgia syndrome (FMS) is characterized by widespread pain, fatigue, sleep abnormalities, and distress. Because FMS lacks consistent evidence of tissue abnormalities, recent investigations have focused on central nervous system mechanisms of pain. Abnormal temporal summation of second pain (wind-up) and central sensitization have been described recently in patients with FMS. Wind-up and central sensitization, which rely on central pain mechanisms, occur after prolonged C-nociceptor input and depend on activation of nociceptor-specific neurons and wide dynamic range neurons in the dorsal horn of the spinal cord. Other abnormal central pain mechanisms recently detected in patients with FMS include diffuse noxious inhibitory controls. These pain inhibitory mechanisms rely on spinal cord and supraspinal systems involving pain facilitatory and pain inhibitory pathways. Brain-imaging techniques that can detect neuronal activation after nociceptive stimuli have provided additional evidence for abnormal central pain mechanisms in FMS. Brain images have corroborated the augmented reported pain experience of patients with fibromyalgia during experimental pain stimuli. In addition, thalamic activity, which contributes significantly to pain processing, was decreased in fibromyalgia. However, central pain mechanisms of fibromyalgia may not depend exclusively on neuronal activation. Neuroglial activation has been found to play an important role in the induction and maintenance of chronic pain. These findings may have important implications for future research and the treatment of fibromyalgia pain. PMID- 12126582 TI - Adult growth hormone deficiency in patients with fibromyalgia. AB - Adult growth hormone (GH) deficiency is a well-described clinical syndrome with many features reminiscent of fibromyalgia. There is evidence that GH deficiency as defined in terms of a low insulin-like growth factor-1 (IGF-1) level occurs in approximately 30% of patients with fibromyalgia and is probably the cause of some morbidity. It seems most likely that impaired GH secretion in fibromyalgia is related to a physiologic dysregulation of the hypothalamic-pituitary-adrenal axis (HPA) with a resulting increase in hypothalamic somatostatin tone. It is postulated that impaired GH secretion is secondary to chronic physical and psychological stressors. It appears that impaired GH secretion is more common than clinically significant GH deficiency with low IGF-1 levels. The severe GH deficiency that occurs in a subset of patients with fibromyalgia is of clinical relevance because it is a treatable disorder with demonstrated benefits to patients. PMID- 12126583 TI - Peripheral and central sensitization in musculoskeletal pain disorders: an experimental approach. AB - This report provides a brief introduction to the manifestations of peripheral and central sensitization involved in musculoskeletal pain disorders. It has become increasingly evident that muscle hyperalgesia, referred pain, referred hyperalgesia, and widespread hyperalgesia play an important role in chronic musculoskeletal pain. A better understanding of the involved basic mechanisms and better methods to assess muscle pain in the clinic may provide new possibilities for designing rational therapies and for targeting the pharmacologic intervention optimally. Peripheral sensitization plays an important role for increased sensitivity of deep tissue. However, central sensitization may be equally important but less addressed. Quantitative sensory testing provides the possibility to evaluate these manifestations in a standardized way in patients with musculoskeletal pain or in healthy volunteers (eg, experimentally induced referred pain can be used to assess the potential involvement of central sensitization in musculoskeletal pain conditions). Central sensitization may play a role in the persistence, amplification, and spread of pain. Interventions should take this aspect into consideration. PMID- 12126584 TI - Irritable bowel syndrome as a common precipitant of central sensitization. AB - Animal models of neuropathic pain have significantly advanced our knowledge of abnormalities in central pain processing mechanisms in chronic pain disorders. New neuroimaging techniques using functional magnetic resonance imaging and positron emission tomography scanning are beginning to provide insight into cortical participation in the processing of pain. Irritable bowel syndrome (IBS) is one of the most common gastrointestinal disorders seen by physicians. Visceral hypersensitivity or decreased pain thresholds to distension of the gut is considered to be a biologic marker for IBS and is present in most patients with this gastrointestinal disorder. Patients with IBS also have many extraintestinal symptoms consistent with a central hyperalgesic state. Recent studies suggest that patients with IBS may also have cutaneous hyperalgesia similar to that seen in other chronic pain disorders such as fibromyalgia. This suggests that abnormalities of central nociceptive processing are present in IBS. PMID- 12126585 TI - Functional imaging of pain perception. AB - The application of functional imaging techniques has revolutionized the field of human pain physiology and has elaborated the understanding of mechanisms involved in pain processing at the cortical and subcortical levels. With these insights, new therapeutic interventions are being developed in the treatment of acute and chronic pain conditions. PMID- 12126586 TI - Neuropsychiatric involvement in systemic lupus erythematosus. AB - The correct diagnosis of central and peripheral nervous system manifestations in patients with systemic lupus erythematosus (SLE) can be challenging because of many SLE-related and non-SLE-related processes that can be present in a patient. The American College of Rheumatology (ACR) has published case definitions for 19 neuropsychiatric SLE (NPSLE) syndromes, with careful attention to important exclusion criteria. These criteria were developed for research purposes but can be helpful to clinicians with a patient who has nervous system dysfunction. This report reviews the data regarding the application of the ACR NPSLE criteria and the influence of ethnicity and disease duration on the development of NPSLE syndromes. Cognitive dysfunction and psychiatric disorders are the two most common manifestations. The work-up and treatment of nervous system syndromes are also discussed. PMID- 12126587 TI - B lymphocyte stimulator protein levels in systemic lupus erythematosus and other diseases. AB - The size of the known members of the tumor necrosis factor ligand and receptor superfamilies has burgeoned in the past few years. Among the novel tumor necrosis factor ligand and receptor superfamily members recently described is B lymphocyte stimulator (BLyS; Human Genome Sciences, Rockville, MD) protein. By virtue of its ability to promote B-cell survival, expansion, and differentiation, it likely plays an important contributory role in systemic lupus erythematosus pathogenesis and propagation. In addition, it may play a similar role in other systemic immune based rheumatic diseases, and becomes a legitimate candidate target for antagonist biologic agents. PMID- 12126588 TI - Malignancy and systemic lupus erythematosus. AB - The association of malignancy with systemic lupus erythematosus (SLE) has been investigated for years. The findings of cohort studies lend support for an increased risk of malignancy in SLE but are difficult to interpret definitively. In addition, several cohort studies have suggested an increased risk of non Hodgkin's lymphoma but with imprecise estimation. There is inadequate evidence for any conclusions about the risk of solid tumors in these patients. A multicenter international research effort is in progress to elucidate these issues and to establish the role of exposures such as cytotoxic or immunomodulatory therapy. The recommendations advocated for cancer screening policies and for minimizing known risk factors for cancer in the general population should not be neglected in persons with SLE. PMID- 12126590 TI - Issues and perspectives on brain tissue banking. PMID- 12126591 TI - Botanical medicines for anxiety: do they have a place? PMID- 12126589 TI - An update on genetic studies of systemic lupus erythematosus. AB - Systemic lupus erythematosus (SLE) is a complex, multifactorial autoimmune disease. Genetic factors are thought to contribute to its pathogenesis. There have been numerous recent advances in the study of murine and human lupus genetics. In well-defined experimental transgenic or gene-knockout mouse models, the development of lupus-like disease has implicated specific genes and pathways in the disease pathogenesis. Linkage analyses have mapped multiple susceptibility loci and disease suppressive loci using inbred strains of mice that spontaneously develop lupus-like disease. Elegant genetic dissection and function studies have led to the recent identification of two murine candidate susceptibility genes, Ifi202 (encoding an interferon-inducible protein) and Cr2 (encoding complement receptors 1 and 2). In human lupus, case- control studies have established associations of SLE with certain major histocompatibility class II alleles, complement deficiencies, and polymorphisms of Fc gamma receptor genes, a complement-related gene, and cytokine genes. During the past several years, linkage analyses using SLE multiplex families have provided many chromosomal regions for further exploration of susceptibility genes. Six regions exhibiting significant linkage to SLE are promising. Studies are underway to fine map these linked regions and to identify the genes in the susceptibility regions. An understanding of the genes involved in the development of lupus should provide targets for more focused therapy in lupus. PMID- 12126592 TI - Managing acute stress response to major trauma. AB - In this article, the authors review the current empiric literature on early interventions. Findings on the effects, course, help-seeking, and recovery from disasters are first reviewed, with recommendations given that are pertinent to intervention following mass casualties. In reviewing the most commonly used interventions, it is clear that evidence from well-controlled studies showing that early intervention can help prevent longer-term problems is limited. The authors discuss the approaches that have received the most attention or empiric support as early interventions following trauma, which include psychologic debriefing, cognitive-behavioral interventions, eye movement desensitization and processing (EMDR) and other neoteric approaches, and psychopharmacology. At this time, the most promising results for prevention of psychopathology have been achieved with brief four- or five-session cognitive-behavioral therapy. In contrast, randomized clinical trials on psychologic debriefing currently suggest that this approach is either ineffective at preventing psychopathology, or contributive to post-traumatic stress disorder symptoms. Research support is currently lacking for EMDR and pharmacotherapy as early interventions. A major challenge to the field is to integrate the practical experience and knowledge of professional responders with well-controlled, timely intervention research, and to effectively disseminate these findings to practitioners in the field. PMID- 12126593 TI - Neuroimaging studies in post-traumatic stress disorder. AB - The past decade has seen a rapid advance in understanding of the neural circuits of post-traumatic stress disorder (PTSD), which has largely been due to the application of neuroimaging to the study of this disorder. Based on studies in animals of the effects of stress on the brain, dysfunction of the medial prefrontal cortex, hippocampus, and amygdala have been hypothesized to underlie symptoms of PTSD. Neuroimaging studies in PTSD have been consistent with these hypotheses, with the most replicated findings showing decreased medial prefrontal cortical function in PTSD. Other replicated findings include decreased inferior frontal gyrus function, decreased hippocampal function, increased posterior cingulate function, and, in some behavioral paradigms, increased amygdala function. Several studies have now shown changes in structure (smaller volume) of the hippocampus in PTSD. These studies are beginning to map out a neural circuitry of PTSD that may have future implications for diagnosis and treatment. PMID- 12126594 TI - Pharmacotherapy of childhood anxiety disorders. AB - Anxiety disorders are among the most common psychiatric disorders of childhood, yet limited data is available regarding the use of psychotropic medications for these conditions. Until recently, much of the data on the pharmacologic treatment of pediatric anxiety disorders has consisted of case reports and small open-label studies, with the exception of pediatric obsessive-compulsive disorder (OCD), which has had a comparatively rich literature consisting of several double blind trials. This void has been lessening, however, with four double blind, placebo controlled studies published in the past year alone. Although the majority of pharmacologic studies of pediatric anxiety continue to focus on the treatment of OCD, additional reports on treatment of generalized anxiety disorder, panic disorder, social anxiety disorder, and separation anxiety disorder have recently been published. This article will review significant pharmacologic studies published in the prior year, and the role of pharmacotherapy in the treatment of pediatric anxiety disorders. PMID- 12126595 TI - Postpsychotic depression in schizophrenia patients. AB - Depression is a frequent comorbidity in the course of schizophrenia and is associated with increased mortality from suicide. Postpsychotic depression is defined as the syndrome of major depression occurring following remission of psychotic symptoms in a person with schizophrenia. Various proposed causes, differential diagnosis, and issues regarding management of postpsychotic depression are discussed. PMID- 12126596 TI - Suicidality in schizophrenia: a review of the evidence for risk factors and treatment options. AB - Suicide is a major contributor to the morbidity and mortality of schizophrenia, accounting for approximately 10% of deaths in these patients. The known risk factors for suicide in schizophrenia include prior suicide attempts, substance abuse, male sex, onset during first decade of illness, social isolation, depression, and feelings of hopelessness. There is significant evidence suggesting that clozapine reduces the suicide rate in patients with schizophrenia and schizoaffective disorder. Possible factors that lead to a decrease in suicidality with clozapine include the following: a direct antidepressant action, improved cognitive function and insight, diminished negative symptoms, reduced substance abuse, and improved compliance. These effects may converge or lessen feelings of hopelessness and more of its converse optimism. The International Suicide Prevention Trial (InterSePT) is a large prospective, 2-year randomized trial of the comparative effects of clozapine and olanzapine involving 980 patients at high risk for suicide in 11 countries in 56 sites. The study included complete freedom to augment these treatments if needed, blinded ratings, a blinded Suicide Monitoring Board, and equivalent clinical contact. The results support the superiority of clozapine over olanzapine to reduce the risk of suicidality and suggest its use should be considered for all patients with schizophrenia with high risk for suicide. PMID- 12126597 TI - A clinical review of cognitive therapy for schizophrenia. AB - Major advances have been made in the cognitive understanding and treatment of the symptoms of schizophrenia, including delusions, hallucinations, and emotional withdrawal. Experimental studies on the psychologic aspects of schizophrenia demonstrate the importance of information processing biases, such as cognitive biases and distortions, that are functionally related to the maintenance of symptoms. Understanding the aspects of schizophrenia in cognitive terms provides a framework for psychotherapeutic intervention with the adaptation of the cognitive strategies proven effective in the treatment of mood and anxiety disorders. The authors of this paper first outline the cognitive conceptualization and strategies employed by cognitive therapists to treat positive and negative symptoms, and conclude with a summary of the empiric status of cognitive therapy for schizophrenia. Cognitive therapy has been shown to be an important adjunct to standard treatments of schizophrenia. PMID- 12126598 TI - The development of schizophrenia in late adolescence. AB - Adolescence is an unusual psychologic time. A recent psychodevelopmental approach to psychosis attempted to show how psychotic signs might arise from typical features of adolescence; for example, delusions appear to reflect common adolescent themes of attachment and autonomy. This psychodevelopmental approach emphasizes how normal adolescents grow out of a natural egocentricity and idealism through learning about others; this theory converges with more recent neurologic theories. (Such neurologic theories agree that mentalizing-for others abilities are a crucial mechanism whereby the suspected neurologic problems of psychosis translate into the symptoms.) A psychodevelopmental account implies a therapeutic priority would be reattaching psychosis sufferers with their peer group, perhaps through work placement schemes. It also recommends cognitive work directing self-consciousness into understanding other people. Psychodevelopmental approaches offer a useful theoretic background for psychologic interventions with young "at risk" people. PMID- 12126599 TI - Antipsychotic treatment for late-life schizophrenia. AB - The clinical characteristics of schizophrenia in older persons vary to some extent, depending on whether the onset of illness was earlier or later in life. Regardless of age of onset, antipsychotic medications are the mainstay of treatment. Age-related physiologic changes make older persons more sensitive to the therapeutic and toxic effects of antipsychotics. There is a paucity of controlled studies on the efficacy of antipsychotic medications in older persons with schizophrenia. Existing data suggest that atypical antipsychotics are at least as efficacious as and better tolerated than the conventional agents. In late-life schizophrenia, important adverse effects of antipsychotics include sedating, anticholinergic and cardiovascular effects, extrapyramidal symptoms, and tardive dyskinesia. Certain atypical antipsychotics are associated with a risk of metabolic changes as well as agranulocytosis. Clinical recommendations include a thorough diagnostic evaluation followed by treatment with low doses of atypical antipsychotics. Medication alone is likely to be less effective than when it is combined with an appropriate psychosocial intervention. PMID- 12126602 TI - Short-course therapy for right-sided endocarditis due to Staphylococcus aureus in drug abusers: cloxacillin versus glycopeptides in combination with gentamicin. PMID- 12126601 TI - West Nile Virus: An Overview for the Primary Care Provider. PMID- 12126600 TI - Schizophrenia and genetics: new insights. AB - There is consistent evidence that the principal etiology of schizophrenia involves predisposing genetic factors. Recent years have seen several new insights in the genetics of schizophrenia. Several chromosomal regions show significant evidence that they contain schizophrenia susceptibility genes. A clinically relevant genetic subtype of schizophrenia (22q deletion syndrome) has been identified. There is new evidence that spontaneous mutations may play a role. There are new recommendations for genetic counseling. The progress to date suggests that understanding of a neurodevelopmental pathway from genetic susceptibility to schizophrenia will soon be fundamentally altered by molecular genetic advances in this complex disease. PMID- 12126603 TI - Quinolone treatment for pediatric bacterial meningitis: a comparative study of trovafloxaxin and ceftriaxone with or without vancomycin. PMID- 12126605 TI - Nonvalvular Intravascular Device-related Infections. AB - Nonvalvular intravascular devices are in widespread use in developed countries and are now more commonly employed than are prosthetic cardiac valves. Although the variety of devices that are included in the category of "nonvalvular" represent a heterogenous collection, there is commonality among several of the devices. This includes their requirement to sustain life and the need for removal for cure if they become infected, which is often due to multidrug-resistant microorganisms. Thus, infection of these devices often presents difficult treatment scenarios. This update will address the pathogenesis and immunobiology of nonvalvular intravascular device-related infections and the epidemiology and treatment of infections of several of these devices that include pacemakers, implantable cardioverter defibrillators, intravascular catheters, vascular grafts and stents, and left ventricular assist devices. PMID- 12126604 TI - Cardiac Transplantation: Pre-transplant Infectious Diseases Evaluation and Post transplant Prophylaxis. AB - Screening of recipients and donors of cardiac allografts for infectious pathogens, and the use of appropriate immunization and antimicrobial prophylaxis strategies, remain important for the control of infection following heart transplantation. However, the risk of infectious complications in a particular patient must often be weighed against the risk of delaying or denying allograft transplantation. In addition, the ongoing degree of immunosuppression and its contribution to the risk for infectious complications should also be considered to guide the length of prophylactic antimicrobial therapy and provide optimal patient care. PMID- 12126606 TI - Platelets and Platelet Inhibitors in Infective Endocarditis. AB - The pathogenesis of infective endocarditis depends on complex interactions between the causative pathogen, plasma proteins, platelets, and vascular endothelial cells. In addition to being the main target of bacteria in the initial stage of bacterial adherence to the endocardium, platelets now appear to play an important role in antimicrobial host defense against endocarditis through the secretion of so-called platelet microbicidal proteins. In animal models of endocarditis, low-dose aspirin was shown to significantly reduce the vegetation weight, the bacterial density of vegetation, the hematogenous bacterial dissemination, and the frequency of embolic events. However, these facts cannot be extrapolated to clinical care in humans, since to date, there is no definitive proof of the adjunctive benefit of aspirin in human infective endocarditis. PMID- 12126607 TI - Diagnosis and Management of Enteroviral Infections of the Central Nervous System. AB - Even with the approaching worldwide eradication of the polioviruses, the nonpolio enteroviruses remain major pathogens of the central nervous system. Our understanding of this important group of viral pathogens has increased dramatically in the past decade. The advent of molecular virology has yielded information that has been vital to the development of molecular diagnostic techniques for the detection of the enteroviruses and for the design of novel antienteroviral drugs. Advances in molecular diagnostics have allowed for a better definition of the diseases they cause and have impacted on patient care. This review discusses recent developments in the diagnosis and treatment of enterovirus infections of the central nervous system, including an overview of the molecular virology of the enteroviruses as it pertains to taxonomy, diagnosis, and treatment. PMID- 12126608 TI - Prevention of Pneumococcal Meningitis. AB - With the success of the conjugated Haemophilus influenzae type b vaccines, Streptococcus pneumoniae has become one of the most important causes of bacterial meningitis worldwide, causing significant morbidity and mortality. Additionally, the increasing amount of resistance that this organism is developing to multiple classes of antimicrobial agents has made the treatment of pneumococcal infections, especially meningitis, much more difficult. Immunization has been shown to be one of most effective methods for preventing pneumococcal meningitis, resulting not only in a decrease in disease burden, but also a decrease in antimicrobial resistance. Currently, a 23-valent pneumococcal polysaccharide vaccine and a heptavalent protein conjugate vaccine are licensed for use. However, the 23-valent polysaccharide vaccine is poorly immunogenic in infants and young children. The continued development, licensing, and use of pneumococcal conjugate vaccines have the best potential to both prevent disease and decrease the prevalence of pneumococcal meningitis. PMID- 12126609 TI - Approach to the Diagnosis and Management of Tuberculous Meningitis. AB - Meningitis caused by Mycobacterium tuberculosis remains an important cause of morbidity and mortality worldwide, and presents specific challenges in terms of diagnosis and management. The nonspecific clinical presentation of tuberculous meningitis has led researchers to develop newer biochemical and molecular methods of making the diagnosis. Several of these methods have excellent sensitivity and specificity, although are not yet available for clinical use. Successful therapy for tuberculous meningitis requires a combination of antimicrobial agents, with vigilance towards the possibility of disease caused by resistant organisms. Adjunctive corticosteroids also have a role in treating this potentially devastating infection. With proper therapy, morbidity and mortality can be minimized in patients with tuberculous meningitis. PMID- 12126610 TI - Management of Increased Intracranial Pressure in Cryptococcal Meningitis. AB - Therapy of cryptococcal meningitis has been focused primarily on optimizing the antifungal regimen to improve the previously high treatment failure rates. Until recently, relatively little attention has been paid to the impact of increased intracranial pressure, which is a frequent problem that complicates management of patients with cryptococcal meningitis. Patients with elevated baseline opening pressures have higher titers of cerebrospinal fluid cryptococcal capsular polysaccharide antigen, more frequent headaches, meningismus, papilledema, hearing loss, and pathologic reflexes, as well as increased morbidity and mortality compared with those patients with normal opening pressures. Optimal therapy has not yet been firmly established, but the diagnostic evaluation and available treatment options are reviewed here, including frequent high volume lumbar punctures, lumbar drains, ventriculoperitoneal shunting, and corticosteroids. PMID- 12126612 TI - The Role of Therapeutic Drug Monitoring in the Management of HIV-infected Patients. AB - The concept of targeting drug therapy based on plasma concentrations, also called therapeutic drug monitoring (TDM), has been used in the treatment of infectious diseases and other illnesses for decades. A number of clinical trials have demonstrated that drug concentrations are an important factor in response to therapy for HIV, but whether TDM will become a tool for the routine management of HIV infection remains to be determined. The concept of the inhibitory quotient, which integrates drug concentrations and resistance testing, also shows promise in a number of retrospective analyses. Logistical problems still remain with regard to its feasibility, and theoretical issues such as protein binding, variability, and the appropriate time of sampling continue to be debated. A growing body of literature supports the concept of TDM in HIV, but it is important that it be used with other interventions such as resistance testing, adherence monitoring, and patient counseling to be an effective tool in patient management. PMID- 12126613 TI - HIV Vaccines: Biological and Clinical Considerations. AB - The discovery of an HIV-1 vaccine is a high priority. Recent advances in HIV vaccine development include an improved understanding about virus biology and structure, and the development of quantitative techniques that enable a detailed analysis of vaccine-induced immune responses in humans. The preclinical vaccine pipeline looks healthy, and a common feature of the new vaccine strategies is their ability to attenuate clinical disease rather than prevent HIV infection in nonhuman primates. Human clinical trials to evaluate the safety and immunogenicity of these vaccine candidates and strategies are being actively pursued. PMID- 12126611 TI - Molecular Techniques in the Diagnosis of Central Nervous System Infections. AB - Development of polymerase chain reaction (PCR)-based molecular techniques has initiated a revolution in the field of diagnostic microbiology. These techniques have not only provided rapid, noninvasive detection of microorganisms that cause central nervous system (CNS) infections, but have also demonstrated that several neurologic disorders are linked to infectious agents. While PCR-based techniques are predicted to be widely used in diagnosing and monitoring CNS infections, the limitations, as well as strengths, of these techniques must be clearly understood by both clinicians and laboratory personnel to ensure proper utilization. PMID- 12126614 TI - The solution structure of d(G(4)T(4)G(3))(2): a bimolecular G-quadruplex with a novel fold. AB - The G-rich 11-mer oligonucleotide d(G(4)T(4)G(3)) forms a bimolecular G quadruplex in the presence of sodium ions with a topology that is distinct from the folds of the closely related and well-characterized sequences d(G(4)T(4)G(4)) and d(G(3)T(4)G(3)). The solution structure of d(G(4)T(4)G(3))(2) has been determined using a combination of NMR spectroscopy and restrained molecular dynamics calculations. d(G(4)T(4)G(3))(2) forms an asymmetric dimeric fold-back structure consisting of three stacked G-quartets. The two T(4) loops that span diagonally across the outer faces of the G-quartets assume different conformations. The glycosidic torsion angle conformations of the guanine bases are 5'-syn-anti-syn-anti-(T(4) loop)-anti-syn-anti in one strand and 5'-syn-anti syn-anti-(T(4) loop)-syn-anti-syn in the other strand. The guanine bases of the two outer G-quartets exhibit a clockwise donor-acceptor hydrogen-bonding directionality, while those of the middle G-quartet exhibit the anti-clockwise directionality. The topology of this G-quadruplex, like other bimolecular fold back structures with diagonal loops, places each strand of the G-quartet region next to a neighboring parallel and an anti-parallel strand. The two guanine residues not involved in G-quartet formation, G4 and G12 (i.e. the fourth guanine base of one strand and the first guanine base of the other strand), adopt distinct conformations. G4 is stacked on top of an adjacent G-quartet, and this base-stacking continues along with the bases of the loop residues T5 and T6. G12 is orientated away from the core of G-quartets; stacked on the T7 base and apparently involved in hydrogen-bonding interactions with the phosphodiester group of this same residue. The cation-dependent folding of the d(G(4)T(4)G(3))(2) quadruplex structure is distinct from that observed for similar sequences. While both d(G(4)T(4)G(4)) and d(G(3)T(4)G(3)) form bimolecular, diagonally looped G-quadruplex structures in the presence of Na(+), K(+) and NH(4)(+), we have observed this folding to be favored for d(G(4)T(4)G(3)) in the presence of Na(+), but not in the presence of K(+) or NH(4)(+). The structure of d(G(4)T(4)G(3))(2) exhibits a "slipped-loop" element that is similar to what has been proposed for structural intermediates in the folding pathway of some G-quadruplexes, and therefore provides support for the feasibility of these proposed transient structures in G-quadruplex formation. PMID- 12126615 TI - A bimolecular mechanism of HIV-1 Tat protein interaction with RNA polymerase II transcription elongation complexes. AB - Transcriptional activation of the human immunodeficiency virus type 1 (HIV-1) long terminal repeat (LTR) promoter element is regulated by the essential viral Tat protein that binds to the viral TAR RNA target and recruits a positive transcription elongation complex (P-TEFb). We have used a stepwise transcription approach and a highly sensitive assay to determine the dynamics of interactions between HIV-1 Tat and the transcription complexes actively engaged in elongation. Our results demonstrate that Tat protein associates with RNA polymerase II complexes during early transcription elongation after the promoter clearance and before the synthesis of full-length TAR RNA transcript. This interaction of Tat with RNA polymerase II elongation complexes is P-TEFb-independent. Our results also show that there are two Tat binding sites on each transcription elongation complex; one is located on TAR RNA and the other one on RNA polymerase II near the exit site for nascent mRNA transcripts. These findings suggest that two Tat molecules are involved in performing various functions during a single round of HIV-1 mRNA synthesis. PMID- 12126616 TI - Functional importance of nucleotide identities within the pea atp9 mitochondrial promoter sequence. AB - Sequences ranging from nucleotide positions -14 to +4 relative to the transcription start site constitute an in vitro functional pea atp9 promoter. A comparison of respective sequence segments surrounding 11 unambiguously identified transcription initiation sites of various dicotyledoneous plant species revealed the highest level of evolutionary fidelity of nucleotide identities within the conserved nonanucleotide motif (CNM), suggesting their importance for promoter function. Using a mitochondrial in vitro transcription system, a detailed analysis by site-directed mutagenesis now reveals that the alteration of nucleotides -6 to -2 and +1 within the CNM indeed reduces promoter activity by more than 80%. Changes of nucleotide identities at the less conserved positions -12 to -9 within the AT-rich region reduced the initiation efficiency by about 70%. The alteration of the highly conserved position -7 has little influence on promoter function, indicating that evolutionary conservation does not always correlate with the functional importance of certain nucleotides. Mutagenesis of nucleotides at positions +3 or +4 reveals a minimal requirement of at least one purine for wild-type transcription initiation efficiency. The assignment of functionally important nucleotide identities should now facilitate an efficient and reliable prediction of other promoters in mitochondria of dicotyledon plants. PMID- 12126617 TI - The crystal structures of four peptide deformylases bound to the antibiotic actinonin reveal two distinct types: a platform for the structure-based design of antibacterial agents. AB - Bacterial peptide deformylase (PDF) belongs to a sub-family of metalloproteases that catalyse the removal of the N-terminal formyl group from newly synthesised proteins. PDF is essential in prokaryotes and conserved throughout the eubacteria. It is therefore considered an attractive target for developing new antibacterial agents. Here, we report the crystal structures of four bacterial deformylases, free or bound to the naturally occurring antibiotic actinonin, including two from the major bacterial pathogens Pseudomonas aeruginosa and Staphylococcus aureus. The overall tertiary structure is essentially conserved but shows significant differences, namely at the C terminus, which are directly related to the deformylase type (i.e. I or II) they belong to. The geometry around the catalytic metal ion exhibits a high level of similarity within the different enzymes, as does the binding mode of actinonin to the various deformylases. However, some significant structural differences are found in the vicinity of the active site, highlighting the structural and molecular requirements for the design of a deformylase inhibitor active against a broad spectrum of bacterial strains. PMID- 12126618 TI - Do mRNA and rRNA binding sites of E.coli ribosomal protein S15 share common structural determinants? AB - Escherichia coli ribosomal protein S15 recognizes two RNA targets: a three-way junction in 16S rRNA and a pseudoknot structure on its own mRNA. Binding to mRNA occurs when S15 is expressed in excess over its rRNA target, resulting in an inhibition of translation start. The sole apparent similarity between the rRNA and mRNA targets is the presence of a G-U/G-C motif that contributes only modestly to rRNA binding but is essential for mRNA. To get more information on the structural determinants used by S15 to bind its mRNA target as compared to its rRNA site, we used site-directed mutagenesis, substitution by nucleotide analogs, footprinting experiments on both RNA and protein, and graphic modeling. The size of the mRNA-binding site could be reduced to 45 nucleotides, without loss of affinity. This short RNA preferentially folds into a pseudoknot, the formation of which depends on magnesium concentration and temperature. The size of the loop L2 that bridges the two stems of the pseudoknot through the minor groove could not be reduced below nine nucleotides. Then we showed that the pseudoknot recognizes the same side of S15 as 16S rRNA, although shielding a smaller surface area. It turned out that the G-U/G-C motif is recognized from the minor groove in both cases, and that the G-C pair is recognized in a very similar manner. However, the wobble G-U pair of the mRNA is not directly contacted by S15, as in rRNA, but is most likely involved in building a precise conformation of the RNA, essential for binding. Otherwise, unique specific features are utilized, such as the three-way junction in the case of 16S rRNA and the looped out A(-46) for the mRNA pseudoknot. PMID- 12126619 TI - Functional studies of the 900 tetraloop capping helix 27 of 16S ribosomal RNA. AB - The 900 tetraloop (positions 898-901) of Escherichia coli 16S rRNA caps helix 27, which is involved in a conformational switch crucial for the decoding function of the ribosome. This tetraloop forms a GNRA motif involved in intramolecular RNA RNA interactions with its receptor in helix 24 of 16S rRNA. It is involved also in an intersubunit bridge, via an interaction with helix 67 in domain IV of 23S rRNA. Using a specialized ribosome system and an instant-evolution procedure, the four nucleotides of this loop were randomized and 15 functional mutants were selected in vivo. Positions 899 and 900, responsible for most of the tetraloop/receptor interactions, were found to be the most critical for ribosome activity. Functional studies showed that mutations in the 900 tetraloop impair subunit association and decrease translational fidelity. Computer modeling of the mutations allows correlation of the effect of mutations with perturbations of the tetraloop/receptor interactions. PMID- 12126620 TI - Protein-DNA interactions: amino acid conservation and the effects of mutations on binding specificity. AB - We investigate the conservation of amino acid residue sequences in 21 DNA-binding protein families and study the effects that mutations have on DNA-sequence recognition. The observations are best understood by assigning each protein family to one of three classes: (i) non-specific, where binding is independent of DNA sequence; (ii) highly specific, where binding is specific and all members of the family target the same DNA sequence; and (iii) multi-specific, where binding is also specific, but individual family members target different DNA sequences. Overall, protein residues in contact with the DNA are better conserved than the rest of the protein surface, but there is a complex underlying trend of conservation for individual residue positions. Amino acid residues that interact with the DNA backbone are well conserved across all protein families and provide a core of stabilising contacts for homologous protein-DNA complexes. In contrast, amino acid residues that interact with DNA bases have variable levels of conservation depending on the family classification. In non-specific families, base-contacting residues are well conserved and interactions are always found in the minor groove where there is little discrimination between base types. In highly specific families, base-contacting residues are highly conserved and allow member proteins to recognise the same target sequence. In multi-specific families, base-contacting residues undergo frequent mutations and enable different proteins to recognise distinct target sequences. Finally, we report that interactions with bases in the target sequence often follow (though not always) a universal code of amino acid-base recognition and the effects of amino acid mutations can be most easily understood for these interactions. PMID- 12126621 TI - Simplified normal mode analysis of conformational transitions in DNA-dependent polymerases: the elastic network model. AB - The Elastic Network Model is used to investigate the open/closed transition in all DNA-dependent polymerases whose structure is known in both forms. For each structure the model accounts well for experimental crystallographic B-factors. It is found in all cases that the transition can be well described with just a handful of the normal modes. Usually, only the lowest and/or the second lowest frequency normal modes deduced from the open form give rise to calculated displacement vectors that have a correlation coefficient larger than 0.50 with the observed difference vectors between the two forms. This is true for every structural class of DNA-dependent polymerases where a direct comparison with experimental structural data is available. In cases where only one form has been observed by X-ray crystallography, it is possible to make predictions concerning the possible existence of another form in solution by carefully examining the vector displacements predicted for the lowest frequency normal modes. This simple model, which has the advantage to be computationally inexpensive, could be used to design novel kind of drugs directed against polymerases, namely drugs preventing the open/closed transition from occurring in bacterial or viral DNA dependent polymerases. PMID- 12126622 TI - A conserved tyrosine residue aids ternary complex formation, but not catalysis, in phage T5 flap endonuclease. AB - The flap endonucleases, or 5' nucleases, are involved in DNA replication and repair. They possess both 5'-3' exonucleolytic activity and the ability to cleave bifurcated, or branched DNA, in an endonucleolytic, structure-specific manner. These enzymes share a great degree of structural and sequence similarity. Conserved acidic amino acids, whose primary role appears to be chelation of essential divalent cation cofactors, lie at the base of the active site. A loop, or helical archway, is located above the active site. A conserved tyrosine residue lies at the base of the archway in phage T5 flap endonuclease. This residue is conserved in the structures of all flap endonucleases analysed to date. We mutated the tyrosine 82 codon in the cloned T5 5' nuclease to one encoding phenylalanine. Detailed analysis of the purified Y82F protein revealed only a modest (3.5-fold) decrease in binding affinity for DNA compared with wild type in the absence of cofactor. The modified nuclease retains both structure specific endonuclease and exonuclease activities. Kinetic analysis was performed using a newly developed single-cleavage assay based on hydrolysis of a fluorescently labelled oligonucleotide substrate. Substrate and products were resolved by denaturing HPLC. Steady-state kinetic analysis revealed that loss of the tyrosine hydroxyl function did not significantly impair k(cat). Pre-steady state analysis under single-turnover conditions also demonstrated little change in the rate of reaction compared to the wild-type protein. The pH dependence of the kinetic parameters for the Y82F enzyme-catalysed reaction was bell-shaped as for the wild-type protein. Thus, Y82 does not play a role in catalysis. However, steady-state analysis did detect a large (approximately 300-fold) defect in K(M). These results imply that this conserved tyrosine plays a key role in ternary complex formation (protein-DNA-metal ion), a prerequisite for catalysis. PMID- 12126623 TI - The inherent properties of DNA four-way junctions: comparing the crystal structures of holliday junctions. AB - Holliday junctions are four-stranded DNA complexes that are formed during recombination and related DNA repair events. Much work has focused on the overall structure and properties of four-way junctions in solution, but we are just now beginning to understand these complexes at the atomic level. The crystal structures of two all-DNA Holliday junctions have been determined recently from the sequences d(CCGGGACCGG) and d(CCGGTACCGG). A detailed comparison of the two structures helps to distinguish distortions of the DNA conformation that are inherent to the cross-overs of the junctions in this crystal system from those that are consequences of the mismatched dG.dA base-pair in the d(CCGGGACCGG) structure. This analysis shows that the junction itself perturbs the sequence dependent conformational features of the B-DNA duplexes and the associated patterns of hydration in the major and minor grooves only minimally. This supports the idea that a DNA four-way junction can be assembled at relatively low energetic cost. Both structures show a concerted rotation of the adjacent duplex arms relative to B-DNA, and this is discussed in terms of the conserved interactions between the duplexes at the junctions and further down the helical arms. The interactions distant from the strand cross-overs of the junction appear to be significant in defining its macroscopic properties, including the angle relating the stacked duplexes across the junction. PMID- 12126624 TI - Transactivation of involucrin, a marker of differentiation in keratinocytes, by lens epithelium-derived growth factor (LEDGF). AB - Human involucrin (hINV), first appears in the cytosol of keratinocytes and ultimately cross-linked to membrane proteins via transglutaminase and forms a protective barrier as an insoluble envelope beneath the plasma membrane. Although the function and evolution of involucrin is known, the regulation of its gene expression is not well understood. An analysis of the hINV gene sequence, upstream of the transcription start site (-534 to +1 nt) revealed the presence of potential sites for binding of lens epithelium-derived growth factor (LEDGF); stress response element (STRE; A/TGGGGA/T) and heat shock element (HSE; nGAAn). We reported earlier that LEDGF activates stress-associated genes by binding to these elements and elevates cellular resistance to various stresses. Here, gel shift and super-shift assays confirm the binding of LEDGF to the DNA fragments containing HSEs and STREs that are present in the involucrin gene promoter. Furthermore, hINV promoter linked to CAT reporter gene, cotransfected in human corneal simian virus 40-transformed keratinocytes (HCK), was transactivated by LEDGF significantly. In contrast, the activity of hINV promoter bearing mutations at the WT1 (containing HSE and STRE), WT2 (containing STRE) and WT3 (containing STRE) binding sites was diminished. In addition, in HCK cell over-expressing LEDGF, the levels of hINV mRNA and hINV protein are increased by four to five fold. LEDGF is inducible to oxidants. Cells treated with 12-O-tetradecanoyl phorbol-13-acetate (TPA), known to stimulate production of H(2)O(2), showed higher levels of LEDGF mRNA. Furthermore, our immunohistochemical studies revealed that hINV protein is found in the cytoplasm of HCK cells over-expressing LEDGF, but not detectable in the normal HCK cells or HCK cells transfected with vector. This regulation appears to be physiologically important, as over expression of HCK with LEDGF increases the expression of the endogenous hINV gene and may provide new insight to understand the molecular mechanism of transcriptional regulation of this gene. LEDGF may play an important role in establishing an important barrier in corneal keratinocytes by maintaining epidermal turn-over rate, and protecting HCKs against stress. PMID- 12126625 TI - Structural basis of the adaptive molecular recognition by MMP9. AB - Matrix metalloproteinase (MMPs) are critical for the degradation of extracellular matrix components and, therefore, need to be regulated tightly. Almost all MMPs share a homologous C-terminal haemopexin-like domain (PEX). Besides its role in macromolecular substrate processing, the PEX domains appear to play a major role in regulating MMP activation, localisation and inhibition. One intriguing property of MMP9 is its competence to bind different proteins, involved in these regulatory processes, with high affinity at an overlapping recognition site on its PEX domain. With the crystal structure of the PEX9 dimer, we present the first example of how PEX domains accomplish these diverse roles. Blade IV of PEX9 mediates the non-covalent and predominantly hydrophobic dimerisation contact. Large shifts of blade III and, in particular, blade IV, accompany the dimerisation, resulting in a remarkably asymmetric homodimeric structure. The asymmetry provides a novel mechanism of adaptive protein recognition, where different proteins (PEX9, PEX1, and TIMP1) can bind with high affinity to PEX9 at an overlapping site. Finally, the structure illustrates how the dimerisation generates new properties on both a physico-chemical and functional level. PMID- 12126626 TI - The structures of crystalline complexes of human serum amyloid P component with its carbohydrate ligand, the cyclic pyruvate acetal of galactose. AB - Two monoclinic (P2(1)) crystal forms of human serum amyloid P component (SAP) in complex with the 4,6-pyruvate acetal of beta-D-galactose (MObetaDG) were prepared. Structure analysis by molecular replacement and refinement at 2.2A resolution revealed that crystal form 1 (a=95.76A, b=70.53A, c=103.41A, beta=96.80 degrees) contained a pentamer in the asymmetric unit with a structure very similar to that of the published search model. The mode of ligand co ordination was also similar except that four of the five subunits showed bound ligand with an additional H-bond between O1 of the galactose and the side-chain of Lys79. One sub-unit showed no bound ligand and a vacant calcium site close to a crystal contact. The 2.6A resolution structure of crystal form 2 (a=118.60A, b=109.10A, c=120.80A and beta=95.16 degrees ) showed ten sub-units in the asymmetric unit, all with two bound calcium ions and ligand. The most extensive protein-protein interactions between pentamers describe an AB face-to-face interaction involving 15 ion pairs that sandwiches five molecules of bound MObetaDG at the interface. PMID- 12126627 TI - Structural enzymology of Li(+)-sensitive/Mg(2+)-dependent phosphatases. AB - Li(+)-sensitive/Mg(2+)-dependent phosphatases have attracted considerable attention since they have been proposed as targets for lithium therapy in the treatment of manic-depressive patients. The members of this enzyme superfamily display low levels of sequence identity while possessing a common fold and active site. Extensive structural and biochemical data demonstrate the direct involvement of two metal ions in catalysis, and show that lithium exerts its inhibitory action by blocking the products at the active site. By exploiting the different inhibitory properties of magnesium and calcium, we have been able to solve the X-ray structures of the Li(+)-sensitive/Mg(2+)-dependent 3' phosphoadenosine-5'-phosphatase in complex with its substrate and with its products. The structural comparison of these complexes provides a 3D picture of the different stages of the catalytic cycle. This gives new insights into the understanding of the biological function of this group of enzymes and their lithium inhibition, and should assist in the design of improved inhibitors of therapeutic value. PMID- 12126628 TI - Crystal structure of the ternary complex of the catalytic domain of human phenylalanine hydroxylase with tetrahydrobiopterin and 3-(2-thienyl)-L-alanine, and its implications for the mechanism of catalysis and substrate activation. AB - Phenylalanine hydroxylase catalyzes the stereospecific hydroxylation of L phenylalanine, the committed step in the degradation of this amino acid. We have solved the crystal structure of the ternary complex (hPheOH-Fe(II).BH(4).THA) of the catalytically active Fe(II) form of a truncated form (DeltaN1-102/DeltaC428 452) of human phenylalanine hydroxylase (hPheOH), using the catalytically active reduced cofactor 6(R)-L-erythro-5,6,7,8-tetrahydrobiopterin (BH(4)) and 3-(2 thienyl)-L-alanine (THA) as a substrate analogue. The analogue is bound in the second coordination sphere of the catalytic iron atom with the thiophene ring stacking against the imidazole group of His285 (average interplanar distance 3.8A) and with a network of hydrogen bonds and hydrophobic contacts. Binding of the analogue to the binary complex hPheOH-Fe(II).BH(4) triggers structural changes throughout the entire molecule, which adopts a slightly more compact structure. The largest change occurs in the loop region comprising residues 131 155, where the maximum r.m.s. displacement (9.6A) is at Tyr138. This loop is refolded, bringing the hydroxyl oxygen atom of Tyr138 18.5A closer to the iron atom and into the active site. The iron geometry is highly distorted square pyramidal, and Glu330 adopts a conformation different from that observed in the hPheOH-Fe(II).BH(4) structure, with bidentate iron coordination. BH(4) binds in the second coordination sphere of the catalytic iron atom, and is displaced 2.6A in the direction of Glu286 and the iron atom, relative to the hPheOH-Fe(II).BH(4) structure, thus changing its hydrogen bonding network. The active-site structure of the ternary complex gives new insight into the substrate specificity of the enzyme, notably the low affinity for L-tyrosine. Furthermore, the structure has implications both for the catalytic mechanism and the molecular basis for the activation of the full-length tetrameric enzyme by its substrate. The large conformational change, moving Tyr138 from a surface position into the active site, may reflect a possible functional role for this residue. PMID- 12126629 TI - A structure for the trimeric MHC class II-associated invariant chain transmembrane domain. AB - The major histocompatibility complex (MHC)-associated invariant chain (Ii) contains a single transmembrane domain that forms trimers. Ii is involved in the assembly of the MHC and antigen presentation, and is thus central to the function of the immune system. Here, we show by attenuated total reflectance, Fourier transform infrared (ATR-FTIR) spectroscopy that the transmembrane domain is alpha helical and we provide a structural model of the transmembrane domain obtained by a combination of site-specific infrared dichroism and molecular dynamics (MD) simulations. This work resolves the backbone structure of a transmembrane peptide by multiple (13)C=(18)O labelling at ten different residues. A second purely computational approach, based on MD simulations of Ii transmembrane homologous sequences, yields a similar structure that is consistent with our experimental results. The structure presented forms a left-handed coiled coil with an average helix tilt of 13(+/-6) degrees; the residue Gln47 implicated in trimer formation forms strong interhelical contacts, Thr50 points to the inside of the trimeric coil and forms a network of hydrogen bonds. PMID- 12126630 TI - Folding mechanism of indole-3-glycerol phosphate synthase from Sulfolobus solfataricus: a test of the conservation of folding mechanisms hypothesis in (beta(alpha))(8) barrels. AB - As a test of the hypothesis that folding mechanisms are better conserved than sequences in TIM barrels, the equilibrium and kinetic folding mechanisms of indole-3-glycerol phosphate synthase (sIGPS) from the thermoacidophilic archaebacterium Sulfolobus solfataricus were compared to the well-characterized models of the alpha subunit of tryptophan synthase (alphaTS) from Escherichia coli. A multifaceted approach combining urea denaturation and far-UV circular dichroism, tyrosine fluorescence total intensity, and tyrosine fluorescence anisotropy was employed. Despite a sequence identity of only 13%, a stable intermediate (I) in sIGPS was found to be similar to a stable intermediate in alphaTS in terms of its thermodynamic properties and secondary structure. Kinetic experiments revealed that the fastest detectable folding event for sIGPS involves a burst-phase (<5ms) reaction that leads directly to the stable intermediate. The slower of two subsequent phases reflects the formation/disruption of an off pathway dimeric form of I. The faster phase reflects the conversion of I to the native state and is limited by folding under marginally stable conditions and by isomerization or rearrangement under strongly folding conditions. By contrast, alphaTS is thought to fold via an off-pathway burst-phase intermediate whose unfolding controls access to a set of four on-pathway intermediates that comprise the stable equilibrium intermediate. At least three proline isomerization reactions are known to limit their interconversions and lead to a parallel channel mechanism. The simple sequential mechanism deduced for sIGPS reflects the dominance of the on-pathway burst-phase intermediate and the absence of prolyl residues that partition the stable intermediate into kinetically distinguishable species. Comparison of the results for sIGPS and alphaTS demonstrates that the thermodynamic properties and the final steps of the folding reaction are better conserved than the early events. The initial events in folding appear to be more sensitive to the sequence differences between the two TIM barrel proteins. PMID- 12126631 TI - The natively helical chain segment 169-188 of Escherichia coli adenylate kinase is formed in the latest phase of the refolding transition. AB - The refolding transition of Escherichia coli adenylate kinase (AK) was investigated by monitoring the refolding kinetics of a selected 20 residue helical segment in the CORE domain of the protein. Residues 169 and 188 were labeled by 1-acetamido-methyl-pyrene, and by bimane, respectively. The experiment combines double-jump stopped-flow fast mixing initiation of refolding and time resolved Forster energy transfer spectroscopy for monitoring the conformational transitions (double-kinetics experiment). Two kinetic phases were found in the denaturant-induced unfolding of AK. In the first phase, the fluorescence quantum yields of both probes decreased. The distribution of the distances between them transformed from the native state's narrow distribution with the mean distance corresponding to the distance in the crystal structure, to a distribution compatible with an unordered structure. In the second, slow step of denaturation, neither the fluorescence parameters of the probes nor the distance distribution between them changed. This step appeared to be a transformation of the fast folding species formed in the first phase, to the slow-folding species. Refolding of the fast-folding species of the denatured state of AK was also a two-phase process. During the first fast phase, within less than 5ms, the fluorescence emission of both probes increased, but the distance distribution between the labeled sites was unchanged. Only during the second slow refolding step did the intramolecular distance distribution change from the characteristic of the denatured state to the narrow distribution of the native state. This experiment shows that for the case of the CORE domain of AK, the large helical segment of residues 169-188 was not formed in the first compaction step of refolding. The helical conformation of this segment is established only in the second, much slower, refolding phase, simultaneously with the completion of the native structure. PMID- 12126632 TI - Allosteric inhibition of 3-deoxy-D-arabino-heptulosonate-7-phosphate synthase alters the coordination of both substrates. AB - 3-Deoxy-D-arabino-heptulosonate-7-phosphate synthase (DAHPS), the first enzyme of the aromatic biosynthetic pathway in microorganisms and plants, catalyzes the aldol-like condensation of phosphoenolpyruvate and D-erythrose-4-phosphate with the formation of 3-deoxy-D-arabino-heptulosonate-7-phosphate. In Escherichia coli, there are three isoforms of DAHPS, each specifically feedback-regulated by one of the three aromatic amino acid end products. The crystal structure of the phenylalanine-regulated DAHPS from E.coli in complex with its inhibitor, L phenylalanine, phosphoenolpyruvate, and metal cofactor, Mn(2+), has been determined to 2.8A resolution. Phe binds in a cavity formed by residues of two adjacent subunits and is located about 20A from the closest active site. A model for the mechanism of allosteric inhibition has been derived from conformational differences between the Phe-bound and previously determined Phe-free structures. Two interrelated paths of conformational changes transmit the inhibitory signal from the Phe-binding site to the active site of DAHPS. The first path involves transmission within a single subunit due to the movement of adjacent segments of the protein. The second involves alterations in the contacts between subunits. The combination of these two paths changes the conformation of one of the active site loops significantly and shifts the other slightly. This alters the interaction of DAHPS with both of its substrates. Upon binding of Phe, the enzyme loses the ability to bind D-erythrose-4-phosphate and binds phosphoenolpyruvate in a flipped orientation. PMID- 12126633 TI - Immunization of rats with homologous type XI collagen leads to chronic and relapsing arthritis with different genetics and joint pathology than arthritis induced with homologous type II collagen. AB - The most commonly used animal model for rheumatoid arthritis (RA) is collagen induced arthritis (CIA), induced by immunization with type II collagen (CII), a cartilage restricted protein. In this work we show that type XI collagen (CXI), which is a minor component in cartilage, induces a different form of erosive and chronic relapsing polyarthritis in rats. Using a series of inbred rat strains involving various genetic backgrounds (DA, LEW, E3), and congenic MHC regions (a, u, f, n, c, d), we found that CXI induced arthritis (C(XI)IA) is associated with the RT1f haplotype in contrast to CII induced arthritis (C(II)IA), which is associated with the RT1a and RT1u haplotypes. The C(XI)IA follows a chronic disease course affecting peripheral joints with both progression and relapses, which appear not to cease (occurring >800 days). Susceptible strains showed a sustained antibody response to CXI with time indicating that the autoimmune response was self-perpetuated. Microscopic analysis of the joints at different stages demonstrated the severe destruction of bone and cartilage by pannus tissue consisting of activated macrophages and T cells. The main difference to joints from rats with C(II)IA was larger numbers of infiltrating lymphocytes and these tended to form follicle-like aggregates. Surprisingly, males were more susceptible to C(XI)IA than females whereas the opposite has been observed in other rat arthritis models, including C(II)IA. Taken together, C(XI)IA is a chronic relapsing and erosive polyarthritis that is MHC associated, which in fact fulfills the criteria for diagnosis of RA. Thus the C(XI)IA model will be useful as a novel and relevant animal model for RA. PMID- 12126634 TI - Coexpression of susceptible and resistant HLA class II transgenes in murine experimental autoimmune thyroiditis: DQ8 molecules downregulate DR3-mediated thyroiditis. AB - Experimental autoimmune thyroiditis (EAT) can be induced in genetically susceptible mice by immunization with the self antigen, thyroglobulin (Tg). Since susceptibility is linked to H2 class II molecules, we have generated human leukocyte antigen (HLA) class II transgenic mice to study potential HLA associations with Hashimoto's thyroiditis. DR3 (HLA-DRA/DRB1*0301) and DQ8 (HLA DQA1*0301/DQB1*0302) transgenes were introduced into class II-negative Ab(0)/B10 and Ab(0) nonobese diabetic (Ab(0)/NOD) mice. Previous work had shown that DR3 transgenic mice were susceptible to both mouse Tg and human Tg-induced EAT, whereas DQ8 transgenic mice were moderately susceptible only to human Tg induction. In this report, we examined the effect of DQ8 transgene on mouse Tg- and human Tg-induced EAT in double transgenic DR3/DQ8 mice. After mouse Tg induction, thyroiditis in DR3(+)DQ8(+) Ab(0)/B10 mice was significantly less severe than in DR3(+) mice but more severe than in DQ8(+) mice. No difference in thyroiditis was observed between DR3(+) and DR3(+)DQ8(+) mice in another background strain, Ab(0)/NOD. However, after immunization with human Tg, DQ8 coexpression downregulated thyroiditis severity, compared to DR3(+) mice, whereas thyroiditis was more extensive than in DQ8(+) mice. Thus, depending on the background strain and the Tg used to induce disease, the presence of the DQ8 transgene can reduce thyroiditis mediated by DR3 molecules. PMID- 12126635 TI - Autoimmune antibodies to hnRNPG protein in dogs with systemic lupus erythematosus: epitope mapping of the antigen. AB - We previously reported the presence of circulating autoantibodies to hnRNPG protein in dogs with systemic lupus erythematosus (Soulard et al. 1993, 1994). These antibodies appeared to be specifically limited to German shepherd dog species. In the present report, we have analysed the nature of the hnRNPG epitopes responsible for autoantibody specificity. By using a set of 11 dog sera selected for their strong reactivity to hnRNPG protein, we have found that these sera had the ability to recognize two epitopes: (1) within a stretch of 33 amino acids located around the central part of protein, that is readily detected by immunoblotting; (2) a N-terminal conformation of the protein close to the RNA binding domain (RBD), that is revealed by immunoprecipitation. Our data strongly support the notion that the dog autoimmune response against hnRNPG protein is antigen-driven. PMID- 12126636 TI - Endothelial cell binding by systemic lupus antibodies: functional properties and relationship with anti-DNA activity. AB - Anti-DNA antibodies and anti-endothelial cell antibodies (AECA) are often detected in systemic lupus erythematosus (SLE). Anti-DNA antibodies can also bind the membrane of human umbilical vein endothelial cells (HUVEC), but little is known about the presence of AECA in the population of immunoglobulins from SLE sera that do not bind DNA. The aim of this study is to analyse the ability of anti-DNA and non-anti-DNA antibodies from SLE sera to bind endothelial cell antigens and to investigate their pathogenic potential. Both anti-DNA and non anti-DNA antibodies display AECA activity by immunoprecipitation and flow cytometry and in some patients recognize antigens of identical molecular weight. Complement-dependent cytotoxicity on HUVEC was not detected with either anti-DNA or non-anti-DNA antibodies. Similarly, apoptosis was not induced in HUVEC and HL60 incubated with anti-DNA or non-anti-DNA antibodies, as shown by the DNA hypodiploid content. These data indicate that AECA are highly heterogeneous, as they recognize a wide variety of surface molecules on HUVEC and equally present in anti-DNA and non-anti-DNA antibodies from SLE patients. PMID- 12126637 TI - Vitronectin- and fibronectin-containing immune complexes in primary systemic vasculitis. AB - In primary systemic vasculitis anti endothelial cell autoantibodies (AECA) have been described frequently. They represent a heterogeneous group of autoantibodies whose target antigens are mostly unknown. We tried to find AECA-antigens by a co operative binding assay with a panel of monoclonal antibodies (mAb) directed to human umbilical vein endothelial cells (HUVEC) and extracellular matrix proteins. The mAb were used to bind antigens from lysate of endothelial cells, and binding of human antibodies to these antigens was measured. mAb directed to Vitronectin (VN) and Fibronectin (FN) resulted in enhanced binding of antibodies in sera from patients with Churg Strauss Syndrome (CSS) and Wegener's Granulomatosis (WG) compared to normal sera. Neither free autoantibodies against VN or FN could be detected nor did the addition of endothelial cell lysate influence the binding activity from the patients' sera. This suggests that preformed VN and FN containing immune complexes (IC) are present in the patient sera. The amount of IC was decreased by incubation with HUVEC, demonstrating that these IC can bind to endothelial cells. However, their involvement in the pathogenesis of the disease is not clearly defined. Our data suggest that there are preformed IC present in sera of patients with CSS and WG that contain VN and FN and bind to endothelial cells. PMID- 12126638 TI - Observations on recent studies showing increased co-occurrence of autoimmune diseases. AB - The co-occurrence of autoimmune diseases has been epidemiologically studied and has aided in our understanding of autoimmunity. However, as new perspectives develop on the pathogenesis and natural history of autoimmune diseases, a refinement in the methodology for the study of the co-occurrence of disease is warranted in order to maximize the information that one may realize from such studies. This paper presents some recent results of co-occurrence studies and then proposes several refinements in the design of epidemiological studies in light of current understanding of the natural history of autoimmune diseases. It also suggests an historical perspective on the results of past studies as to the type of information that can be inferred from them. PMID- 12126639 TI - Reactive oxygen species induce cardiomyocyte apoptosis partly through TNF-alpha. AB - Many studies have indicated that oxidative stress induces apoptosis in cardiomyocytes, but its mechanism remains unknown. We examined whether tumor necrosis factor-alpha (TNF-alpha) is involved in oxidative stress-induced cardiomyocyte apoptosis. Pretreatment with anti-TNF-alpha antibody significantly decreased the number of H(2)O(2)-induced TUNEL-positive cardiomyocytes. Expression of TNF-alpha gene was upregulated by H(2)O(2), and H(2)O(2) mildly but significantly increased the concentration of TNF-alpha in the culture medium. Although neither low dose of H(2)O(2) nor TNF-alpha induced apoptosis, stimulation with H(2)O(2) and TNF-alpha synergistically increased apoptosis. These results suggest that oxidative stress induces apoptosis of cardiac myocytes partly through TNF-alpha. PMID- 12126640 TI - Ccr2 regulates the level of MCP-1/CCL2 in vitro and at inflammatory sites and controls T cell activation in response to alloantigen. AB - CCR2, and its principle ligand MCP-1/CCL2, have been well documented for their ability to induce monocyte infiltration and promote the pathogenesis of rheumatoid arthritis and atherosclerosis. In order to assess additional roles for CCR2, we inserted allogeneic implants into CCR2-/- and MCP-1-/- mice and characterized T cell responses and the regulatory role of CCR2 on MCP-1 expression. The results demonstrate a marked decrease in lymphocyte infiltration in both CCR2-/- and MCP-1-/- animals. In contrast, IL-12 and CTL function were only suppressed in CCR2-/- animals. Further, whereas MCP-1 was only transiently elevated in the inflammatory fluid of WT animals, levels were sustained within the implants (5000pg/ml; >8 days) and serum (243pg/ml) of CCR2-/- mice. Higher levels of MCP-1 were also observed in the culture supernatants of CCR2-/- macrophages as compared to WT cells despite no difference in mRNA levels. Evidence that MCP-1 levels are regulated by receptor binding and internalization was suggested by its rapid decline when added to WT macrophages at 37 degrees C but not 4 degrees C. These studies indicate that CCR2 plays an important role in regulating T cell responses and controlling the level of MCP-1 at inflammatory sites. PMID- 12126641 TI - Human lactiferous mammary gland cells produce vascular endothelial growth factor (VEGF) and express the VEGF receptors, Flt-1 AND KDR/Flk-1. AB - Human milk contains a variety of growth factors. Recently, it was reported that vascular endothelial growth factor (VEGF) was one of them. We investigated milk VEGF isoforms, their functions, and VEGF receptors on mammary gland epithelial cells (MEC). The VEGF concentration in human milk was 74.3+/-34.9ng/ml on the first day after delivery, and rapidly decreased in a couple of days to 6.2+/ 2.3ng/ml on the fifth day, and matured milk maintained about 4ng/ml. In an MTT assay, human milk accelerated HUVEC proliferation and MV303, a neutralizing antibody of VEGF, blocked 17.3 % of the effect. Immunoprecipitation and Western blotting showed that VEGF121 and VEGF165 were contained in human colostrums, and RT-PCR of human MEC confirmed that VEGF121, VEGF165 and VEGF189 were present. By immunostaining of human breast tissues, RT-PCR of MEC from human colostrum and measurement of the VEGF concentrations of conditioned media of cultured human MEC, it was confirmed that VEGF was produced by MEC. MEC was also expressed VEGF receptors, flt-1 and Flk-1/KDR. These results speculate us that the existence of autocrine or paracrine system within breast tissue via VEGF receptors on MEC and have a role in lactation. PMID- 12126642 TI - Secretion of IL-6, IL-11 and LIF by human cardiomyocytes in primary culture. AB - Interleukin (IL)-6-type cytokines are multifunctional proteins involved in cardiac hypertrophy and myocardial protection. Recent studies, performed on animal models, report the production of these cytokines by heart. The aim of this study was to analyse the capacity of myocytes and fibroblasts isolated from human atrium to secrete IL-6, leukaemia inhibitory factor (LIF), cardiotrophin-1 (CT 1), IL-11, oncostatin M (OSM), ciliary neurotrophic factor (CNTF) and the soluble receptor subunits sIL-6R and sgp130 during primary culture. We detected LIF, IL 11, sgp130 and a large amount of IL-6, but not OSM, CT-1, CNTF nor IL-6R in these culture supernatants. Both cardiomyocytes and fibroblasts are able to spontaneously produce IL-6. The increase of IL-6 production all along the culture period appears to be the consequence of fibroblast proliferation and gp130 stimulation. This is the first demonstration that human cardiac cells are able to secrete IL-6, but also LIF and IL-11 in vitro. These cytokines could be involved in an autocrine and/or a paracrine networks regulating myocardial cyto protection, hypertrophy and fibrosis. PMID- 12126644 TI - Site-specific interaction of bone morphogenetic protein 2 with procollagen II. AB - Bone morphogenetic proteins (BMPs) play a critical role in embryo development, organogenesis, and regeneration of damaged tissues. Biological activity of BMPs depends on their local concentration, which is regulated by intracellular enzymatic processing of pro-BMPs, and then the binding of secreted BMPs to antagonizing extracellular proteins. It has been suggested that BMPs interact with structural proteins of the extracellular matrix, but this process is poorly understood. To study interactions of BMPs with fibrillar collagens in detail we expressed recombinant procollagen II variants in which specific domains that correspond to the D-periods were deleted. Subsequently, the procollagen II variants were used in biosensor and immuno-precipitation binding assays to map the regions of procollagen II with a high affinity for the BMP-2. Our data suggest that interaction of BMP-2 with procollagen II is site-specific, and that the high-affinity binding site is located in the D4-period of the collagen triple helix. We hypothesize that the binding of BMP-2 to collagen II reflects a general mechanism of interaction between the fibrillar collagens and morphogens that belong to the transforming growth factor (TGF)-beta superfamily. PMID- 12126643 TI - Role of P38 MAPK, AP-1, and NF-kappaB in interleukin-1beta-induced IL-8 expression in human vascular smooth muscle cells. AB - Interleukin (IL)-1 modulates the expression of various genes in normal and tumor cells. We investigated the molecular mechanisms underlying IL-1beta-induced expression of IL-8 mRNA and protein in human vascular smooth muscle cells (hVSMCs). P38 mitogen-activated protein kinase (MAPK) was activated after 5min of IL-1beta treatment, whereas the extracellular signal-regulated kinases, the c-jun amino-terminal kinases, and protein kinase B/Akt were not activated by IL-1beta. IL-1beta induced activation of a full-length IL-8 promoter-reporter construct. Deletional mutagenesis localized the IL-1beta-responsive domains to two regions ( 133 to -98 and -85 to -50) that contain consensus binding sites for activator protein-1 (AP-1) and nuclear factor-kappaB (NF-kappaB), respectively. Site directed mutagenesis of the 133-bp minimal promoter confirmed that these sites were required for promoter activity. Electrophoretic mobility shift assays confirmed that IL-1beta increased AP-1 and NF-kappaB DNA-binding activities in a time-dependent manner. SB203580, a specific P38 MAPK inhibitor, partially blocked IL-1beta induction of IL-8 mRNA, IL-8 promoter activity, and AP-1 nuclear extract binding but not NF-kappaB DNA binding. Our data demonstrate that AP-1 and NF kappaB are essential transcription factors for IL-1beta-induced IL-8 gene expression in hVSMCs. P38 MAPK is involved in inducing IL-8 gene transcription via AP-1 activation in hVSMCs. PMID- 12126645 TI - Activation of platelet caspases by TNF and its consequences for kinetics. AB - TNF is known to induce a thrombocytopenia, due to a reduced platelet life span. Injection of TNF (10 microg) to mice did markedly increase the number of platelet derived microparticles in plasma, most pronounced 1h after injection. Injection of TNF induced a transient activation of platelet caspases, -1, -3, -6, -8, -9, as seen by the binding of caspases probes detected by flow cytometry, most pronounced 1h after injection. Activation of caspase-3 was also evidenced by antibodies. Injection of the caspases inhibitor ZVAD-fmk decreased TNF-induced generation of microparticles and thrombocytopenia, indicating a causal role of caspases in platelet fragmentation. Activation of platelet caspases was also evident in platelets exposed to TNF in vitro, indicating that TNF acts on platelets directly. Comparison of platelets from +/+, TNFR1 -/- and TNFR2 -/- mice showed that caspases are activated mainly by the TNFR1. These observations indicate that TNF activates platelet caspases via the TNFR1, which results in platelet fragmentation and thrombocytopenia. PMID- 12126646 TI - HMGB-1, a DNA-binding protein with cytokine activity, induces brain TNF and IL-6 production, and mediates anorexia and taste aversion. AB - High-mobility group protein-1 (HMG-1 also termed HMGB-1), a DNA-binding protein, regulates gene transcription and stabilizes nucleosome formation. HMG-1 was recently implicated as a cytokine, because it is a late-acting mediator of endotoxin lethality that induces the release of pro-inflammatory cytokines from monocytes. Here it is shown that administration of HMG-1 into the cerebral ventricles decreases food intake (food intake=4.6g/mouse in controls vs 1.6g/mouse after 1 microg HMG-1 i.c.v.; P <0.05). Intracerebroventricular HMG-1 induced an increased in TNF and IL-6 expression in the brain, and mediated taste aversion with potencies equivalent to LPS. In a model of endotoxemia, passive immunization with anti-HMG-1 antibodies attenuated the development of hypophagia, indicating that HMG-1 is a mediator of sickness behaviour associated with endotoxemia. PMID- 12126647 TI - The presence of cytokine (IL-8, IL-1alpha, IL-1beta)-producing cells in inflamed gingival tissue from a patient manifesting Papillon-Lefevre syndrome(PLS). AB - The point of this study was to examine the presence or absence of cytokine positive cells by means of immunohistochemical methods in the samples of inflamed gingival tissues obtained from an 11-year-old girl with Papillon-Lefevre syndrome (PLS). Interleukin-8 (IL-8)-positive cells were found to be present. In addition, IL-1alpha-and IL-1beta-positive cells were detected. No dysfunction in the phagocytosis and the bacterial killing of peripheral blood polymorphonuclear neutrophils (PMNs) was observed in this patient. Our findings suggest that these cytokines may be members responsible for modulating the process of rapidly progressive periodontitis for patient with PLS. PMID- 12126648 TI - Trauma-haemorrhage-induced alterations in thymic prolactin receptor expression: implications in immune dysfunction. AB - Male gender and age appear to be causative factors in development of immunodepression and septic complications following trauma-haemorrhage. Studies have demonstrated that administration of the sex hormone prolactin following trauma-haemorrhage in male mice prevents immunodepression. Since the thymus is the primary location of the T-cell-lymphopoiesis, we investigated the effect of trauma-haemorrhage to thymic prolactin-receptor (PRLr)-expression in male and proestrus female mice in three different age groups (young, adult, aged) by flow cytometry and PCR. C3H/HeN mice were subjected to laparotomy (i.e., soft-tissue trauma) and hemorrhagic shock (35+/-5mmHg for 90min, then resuscitated) or sham operation. Twenty-four hours later thymocytes were isolated. Trauma-haemorrhage upregulated PRLr expression in young and mature mice of both genders, however, the increase was attenuated in females. In contrast, in aged mice PRLr expression was elevated in both genders, independent of trauma-haemorrhage and was not further increased under such conditions. These findings suggest that the gender dimorphism in the immune response to trauma-haemorrhage may in part be related to differences in thymic PRLr expression under such conditions. PMID- 12126649 TI - Osteoprotegerin levels increased by interleukin-1beta in human periodontal ligament cells are suppressed through prostaglandin E(2) synthesized de novo. AB - Osteoprotegerin (OPG) is a novel tumor necrosis factor receptor superfamily that inhibits osteoclast differentiation, activity, and survival. Interleukin-1beta (IL-1beta) increases OPG expression. IL-1beta also increases prostaglandin E(2) (PGE(2)) production and stimulates bone resorption. In the present study, we examined the involvement of PGE(2) in IL-1beta-induced increases in OPG levels in human periodontal ligament cells (HPL cells) in an effort to clarify apparently conflicting IL-1beta actions on bone resorption and understand IL-1beta-induced increases in secretion of OPG and PGE(2) in HPL cells. 5,6-dichloro-1-beta-D ribofuranosyl-benzimidazole, a mRNA synthesis inhibitor, partly inhibited the increase in OPG mRNA levels induced by IL-1beta. Cycloheximide, a protein synthesis inhibitor, enhanced the stimulatory effect of IL-1beta. Etodolac, a selective cyclooxygenase-2 inhibitor, suppressed the increase in PGE(2) levels. Furthermore, etodolac reinforced the promotion of OPG expression by IL-1beta at the mRNA and protein levels. PGE(2) added to cultures of HPL cells decreased OPG mRNA levels in a dose- and time- dependent manner. These findings suggest that the increase in OPG levels induced by IL-1beta in HPL cells is suppressed through PGE(2) synthesized de novo. PMID- 12126650 TI - Molecular cloning and sequencing of 25 different rhesus macaque chemokine cDNAs reveals evolutionary conservation among C, CC, CXC, AND CX3C families of chemokines. AB - Chemokines are small chemoattractant cytokines involved in normal and pathological immune processes. Although extensive nucleotide sequence data are available for human and murine chemokine cDNA sequences, very few data are currently available regarding rhesus macaque sequences. To increase our understanding of immune function in nonhuman primates, we have used reverse transcription polymerase chain reaction (RT-PCR) to clone and sequence rhesus macaque cDNAs from each of the C, CC, CXC, and CX3C groups of chemokines. Relative to the respective human chemokines, these 25 chemokine cDNA sequences were from 77% to 98% identical. Of the amino acid differences between the rhesus macaque and human chemokines, 51% were species-specific when compared together with the respective murine chemokine sequences. These studies of rhesus macaque chemokine sequences demonstrate that chemokine genes are highly conserved across species, and provide a large foundation for the study of chemokine biology and genetics in nonhuman primates. PMID- 12126651 TI - IL-12 and neutralization of endogenous IL-10 revert the in vitro antigen-specific cellular immunosuppression of paracoccidioidomycosis patients. AB - Treatment of patients with paracoccidioidomycosis is still a challenge. Patients present defective lymphoproliferation and IFN-gamma responses to the main Paracoccidioides brasiliensis antigen (gp43), which correlates with disease severity. Here, we demonstrated that the patients show also a defective synthesis of interleukin (IL)-12. Therefore, we attempted to revert this immune disfunction by adding IL-12 and neutralizing anti-IL-10 antibody to gp-43-stimulated peripheral blood mononuclear cell cultures. Both treatments increased IFN-gamma secretion to levels observed with healthy sensitized individuals, but affected proliferation only modestly. When combined, the treatments further increased IFN gamma synthesis and cell proliferation. The addition of suboptimal concentrations of IL-2 also further increased the IL-12-mediated secretion of IFN-gamma. Interestingly, the immune modulation was mostly antigen-specific, since the responses to Candida albicans' antigen were not affected. These results suggest that appropriate immune intervention with cytokines and/or anti-cytokines may help in the treatment of PCM. PMID- 12126652 TI - Characterization of the interaction between murine tumour necrosis factor and monoclonal antibodies. AB - We characterized the interaction between murine TNF (mTNF) and the neutralizing monoclonal antibodies TN3 and 1F3F3. The epitopes were localized by comparing the detection efficiency for a panel of TNF chimaeric proteins and site-specific muteins in ELISA. Mutation of mTNF amino acid Q131 inhibited the interaction with 1F3F3, whereas mutation of D71/Y72 inhibited the binding to TN3. As D71/Y72 are located in an exposed loop near the TNF intersubunit groove, binding of TN3 promoted the dissociation and/or interfered with the reassociation of subunits into mTNF trimers. 1F3F3, on the other hand, prevented the spontaneous dissociation of bound (hetero)trimeric mTNF. PMID- 12126653 TI - Downregulation of both interleukin-12 and interleukin-2 in heart allografts by pretransplant host treatment with granulocyte colony-stimulating factor and tacrolimus. AB - Since recombinant human granulocyte colony-stimulating factor (rhG-CSF) has been reported to induce immune deviation, we examined the effects of pretransplant treatment of recipients with rhG-CSF on heart allograft survival. Before heterotopic heart transplantation from DA to Lewis rats, recipients were given rhG-CSF (125microg/kg s.c. twice a day from day -5 to 0) and/or tacrolimus (2mg/kg i.m. on day 0). Combined treatment with both rhG-CSF and tacrolimus prolonged graft survival significantly (P<0.05), while rhG-CSF or tacrolimus alone did not. At 24h after transplantation, cytokine mRNA levels in the grafts were measured by reverse transcription and real-time polymerase chain reaction. IL-12 p35 expression was highly inhibited by rhG-CSF treatment, but tacrolimus did not change the levels. Conversely, rhG-CSF treatment did not affect IL-2 levels, while tacrolimus completely blocked its expression. Combined pretreatment was effective for suppressing acute rejection reaction by downregulating these two key type-1 cytokines, IL-12 and IL-2, with unchanged levels of IL-10. PMID- 12126654 TI - Relative contribution of NF-kappaB and AP-1 in the modulation by curcumin and pyrrolidine dithiocarbamate of the UVB-induced cytokine expression by keratinocytes. AB - Following ultraviolet B treatment, expression of tumour necrosis factor (TNF) alpha, interleukin (IL)-6, and IL-8 by NCTC 2544 keratinocyte cell line was significantly enhanced both at the mRNA and protein level. The UVB also increased the IL-10 steady-state mRNAs level. Radiation-induced cytokine overexpression was accompanied by NF-kappaB and AP-1 transcription factors activation as assessed by electrophoretic mobility shift assays. To investigate in keratinocytes the relative contributions of those transcription factors on UVB-mediated cytokine induction, cell cultures were supplemented with curcumin and pyrrolidine dithiocarbamate (PDTC), agents known to modulate NF-kappaB and AP-1 activation. Both compounds significantly inhibited NF-kappaB activation by UVB, but AP-1 activation was unaffected by curcumin while PDTC further stimulated its activation. In parallel, curcumin decreased, in a dose-dependent manner, the UVB mediated overexpression of all three pro-inflammatory cytokines and only exhibited a moderate enhancing influence on IL-10 expression. In turn, the inhibitory influence of PDTC on radiation-induced TNF-alpha and IL-6 expression is much lower and in contrast to curcumin, it stimulated IL-8. Taken together, our data indicated that control of proinflammatory cytokine expression induced by UVB in keratinocytes required the selective inhibition of NF-kappaB activation. Simultaneous AP-1 activation by agents like PDTC might, partially or totally, depending on cytokine-type, counterbalanced the inhibitory effect exerted on UVB induced NF-kappaB activation in keratinocytes. PMID- 12126655 TI - The motor system in neuroscience: a history and analysis of conceptual developments. AB - Neuroscientific reflection on the integrative action of the nervous system was dominated by consideration of the motor system from the time of Aristotle in the 4th century B.C. to that of Sherrington, his contemporaries and proteges in the first-half of the 20th century. We describe the significant discoveries concerning the action of the spinal cord and cortex in motor phenomena during this period. This provides a vivid account of how great neuroscientists, over a period of more than 2000 years, have endeavoured to clarify notions concerning the integrative action of the nervous system in the context of the prevailing philosophical traditions of their times. We examine these traditions as well as the conceptual schemes offered by neuroscientists, especially in relation to the workings of the cortex. It is shown that neuroscientists cleave to this day to a tradition that goes back to Descartes, and that this is the case even for those who explicitly claim to reject such a tradition. The review concludes with what we take to be an appropriate basis for rejecting the Cartesian paradigm that we hope will assist neuroscientists in understanding the integrative action of the nervous system. PMID- 12126657 TI - A brain perspective on language mechanisms: from discrete neuronal ensembles to serial order. AB - Language is constituted by discrete building blocks, sounds and words, which can be concatenated according to serial order principles. The neurobiological organization of these building blocks, in particular words, has been illuminated by recent metabolic and neurophysiological imaging studies. When humans process words of different kinds, various sets of cortical areas have been found to become active differentially. The old concept of two language centers processing all words alike must therefore be replaced by a model according to which words are organized as discrete distributed neuron ensembles that differ in their cortical topographies. The meaning of a word, more precisely, aspects of its reference, may be crucial for determining which set of cortical areas becomes involved in its processing. Whereas the serial order of sounds constituting a word may be established by serially aligned sets of neurons called synfire chains, different mechanisms are necessary for establishing word order in sentences. The serial order of words may be organized by higher-order neuronal sets, called sequence detectors here, which are being activated by sequential excitation of neuronal sets representing words. Sets of sequence detectors are proposed to process aspects of the syntactic information contained in a sentence. Other syntactic rules can be related to general features of the dynamics of cortical activation and deactivation. These postulates about the brain mechanisms of language, which are rooted in principles known from neuroanatomy and neurophysiology, may provide a framework for theory-driven neuroscientific research on language. PMID- 12126656 TI - Dopamine and the regulation of cognition and attention. AB - Dopamine (DA) acts as a key neurotransmitter in the brain. Numerous studies have shown its regulatory role for motor and limbic functions. However, in the early stages of Parkinson's disease (PD), alterations of executive functions also suggest a role for DA in regulating cognitive functions. Some other diseases, which can also involve DA dysfunction, such as schizophrenia or attention deficit hyperactivity disorder (ADHD) in children, as shown from the ameliorative action of dopaminergic antagonists and agonists, respectively, also show alteration of cognitive functions. Experimental studies showed that selective lesions of the dopaminergic neurons in rats or primates can actually provide cognitive deficits, especially when the mesocorticolimbic component of the dopaminergic systems is altered. Data from the experiments also showed significant alteration in attentional processes, thus raising the question of direct involvement of DA in regulating attention. Since the dopaminergic influence is mainly exerted over the frontal lobe and basal ganglia, it has been suggested that cognitive deficits express alteration in these subcortical brain structures closely linked to cortical areas, more than simple deficit in dopaminergic transmission. This point is still a matter of debate but, undoubtedly, DA acts as a powerful regulator of different aspects of cognitive brain functions. In this respect, normalizing DA transmission will contribute to improve the cognitive deficits not only related to neurologic or psychiatric diseases, but also in normal aging. Ontogenic and phylogenetic analysis of dopaminergic systems can provide evidences for a role of DA in the development of cognitive general capacities. DA can have a trophic action during maturation, which may influence the later cortical specification, particularly of pre-frontal cortical areas. Moreover, the characteristic extension of the dopaminergic cortical innervation in the rostro-caudal direction during the last stages of evolution in mammals can also be related to the appearance of progressively more developed cognitive capacities. Such an extension of cortical DA innervation could be related to increased processing of cortical information through basal ganglia, either during the course of evolution or development. DA has thus to be considered as a key neuroregulator which contributes to behavioral adaptation and to anticipatory processes necessary for preparing voluntary action consequent upon intention. All together, it can be suggested that a correlation exists between DA innervation and expression of cognitive capacities. Altering the dopaminergic transmission could, therefore, contribute to cognitive impairment. PMID- 12126659 TI - Involvement of inflammatory cytokines in central nervous system injury. AB - Pro-inflammatory cytokines, interleukin (IL) 1 and tumor necrosis factor (TNF), possess a wide range of biological actions in various tissues. In recent years, there has been increasing evidence that these cytokines are involved in inflammatory reactions in central nervous system (CNS) diseases. Although many studies have demonstrated that IL-1, TNF, and their mRNA are up-regulated in the CNS after injury, the functional roles of these cytokines in the injury are far from completely understood. Overexpression of these cytokines, such as observed during the early stage of injury, can be harmful for the injured tissue. However, low levels of these cytokines, observed during the recovery stage after injury, can enhance repair processes of the injured tissues. PMID- 12126658 TI - Drug interactions at GABA(A) receptors. AB - Neurotransmitter receptor systems have been the focus of intensive pharmacological research for more than 20 years for basic and applied scientific reasons, but only recently has there been a better understanding of their key features. One of these systems includes the type A receptor for the gamma aminobutyric acid (GABA), which forms an integral anion channel from a pentameric subunit assembly and mediates most of the fast inhibitory neurotransmission in the adult vertebrate central nervous system. Up to now, depending on the definition, 16-19 mammalian subunits have been cloned and localized on different genes. Their assembly into proteins in a poorly defined stoichiometry forms the basis of functional and pharmacological GABA(A) receptor diversity, i.e. the receptor subtypes. The latter has been well documented in autoradiographic studies using ligands that label some of the receptors' various binding sites, corroborated by recombinant expression studies using the same tools. Significantly less heterogeneity has been found at the physiological level in native receptors, where the subunit combinations have been difficult to dissect. This review focuses on the characteristics, use and usefulness of various ligands and their binding sites to probe GABA(A) receptor properties and to gain insight into the biological function from fish to man and into evolutionary conserved GABA(A) receptor heterogeneity. We also summarize the properties of the novel mouse models created for the study of various brain functions and review the state-of-the-art imaging of brain GABA(A) receptors in various human neuropsychiatric conditions. The data indicate that the present ligands are only partly satisfactory tools and further ligands with subtype-selective properties are needed for imaging purposes and for confirming the behavioral and functional results of the studies presently carried out in gene-targeted mice with other species, including man. PMID- 12126660 TI - Understanding muscle coordination of the human leg with dynamical simulations. AB - Muscles coordinate multijoint motion by generating forces that cause reaction forces throughout the body. Thus, a muscle can redistribute existing segmental energy by accelerating some segments and decelerating others. In the process, a muscle may also produce or absorb energy, in which case its summed energetic effect on the segments is positive or negative, respectively. This Borelli Lecture shows how dynamical simulations derived from musculoskeletal models reveal muscle-induced segmental energy redistribution and muscle co-functions and synergies. Synergy occurs when co-excited muscles distribute segmental energy differently to execute the motor task. In maximum height jumping, high vertical velocity at lift-off occurs desirably at full body extension because biarticular leg muscles redistribute the energy produced by the uniarticular leg muscles. In pedaling, synergistic ankle plantarflexor force generation during leg extension allows the high energy produced by the uniarticular hip and knee extensors to be delivered to the crank. An analogous less-powerful flexor synergy exists during leg flexion. Hamstrings reduce crank deceleration during the leg extension-to flexion transition by not only producing energy but delivering it to the crank through its contribution to the tangential (accelerating) crank force, though this hamstrings function occurs at the opposite (flexion-extension) transition when pedaling backwards. In walking, the eccentric quadriceps activity in early stance not only decelerates the leg but also accelerates the trunk. In mid stance, the uni- and biarticular plantarflexors, by having opposite segmental energetic effects, act in synergy to support the whole body, so segmental potential and kinetic energy exchange can occur. To conclude, the extraction of unmeasurable variables from dynamical simulations emulating task kinematics, kinetics, and EMGs shows how the production of force and energy by individual muscles contribute to the energy flow among the individual segments during task execution. PMID- 12126661 TI - Tensile properties of in vivo human tendinous tissue. AB - The biomechanical properties of tendinous structures have traditionally been studied using excised material. Limitations associated with displacement measurements and clamping, and uncertainties as to whether in vitro testing represents physiological function, necessitate developing a method for assessing the mechanical properties of tendinous tissue in the in vivo state. This paper reviews recent results taken with an in vivo and noninvasive protocol using ultrasound as a means of measuring tendon-aponeurosis elongation during tensile loading applied by contraction of the in-series muscle. The results obtained indicate that: (1) the Young's modulus and mechanical hysteresis of in vivo tendons is independent of physiological function and loading, (2) there is a strain variation along the tendon-aponeurosis, and (3) in vivo tendons may exhibit creep. These findings agree with reports from experiments on isolated material and have important biological implications for both the tendon and the in-series muscle. The method described here allows designing longitudinal studies on tendon adaptability, but it also has direct clinical applications. PMID- 12126662 TI - In vivo motion of the rectus femoris muscle after tendon transfer surgery. AB - Rectus femoris transfer surgery is performed to convert the rectus femoris muscle from a knee extensor to a knee flexor. In this surgery, the distal tendon of the rectus femoris is detached from the patella and reattached to one of the knee flexor tendons. The outcomes of this procedure are variable, and it is not known if the surgery successfully converts the muscle to a knee flexor. We measured the motion of muscle tissue within the rectus femoris and vastus intermedius during knee extension in 10 unimpaired control subjects (10 limbs) and 6 subjects (10 limbs) after rectus femoris transfer using cine phase-contrast magnetic resonance imaging. Displacements of the vastus intermedius during knee extension were similar between control and tendon transfer subjects. In the control subjects, the rectus femoris muscle consistently moved in the direction of the knee extensors and displaced more than the vastus intermedius. The rectus femoris also moved in the direction of the knee extensors in the tendon transfer subjects; however, the transferred rectus femoris displaced less than the vastus intermedius. These results suggest that the rectus femoris is not converted to a knee flexor after its distal tendon is transferred to the posterior side of the knee, but its capacity for knee extension is diminished by the surgery. PMID- 12126663 TI - Mechanical considerations in the design of surgical reconstructive procedures. AB - Tendon transfers are used to restore arm and hand function after injury to the peripheral nerves or after spinal cord injury. Traditional guidelines to choose the length at which the transferred muscle should be attached have a poor scientific foundation. We postulate that passive tension only becomes significant at relatively long lengths and that passive tension as the major factor in intra operative decision making may result in overstretch of the muscle-tendon unit (MTU) and accompanying low-active force generation. It appears unwise to rely on unknown factors, such as slippage or stress relaxation, to correct an overstretched transfer. Instead, we suggest the use of intra-operative sarcomere length measurements to predict and set the optimal MTU length during reconstructive upper limb surgery. PMID- 12126664 TI - Kinematic simulation of fracture reduction and bone deformity correction under unilateral external fixation. AB - Combined kinematic analysis and graphic models of two unilateral external fixators are presented to simulate and visualize the correction of bone fracture deformities through systematic adjustments of the fixator joints. The models were developed as rigid linkage systems, and the analysis utilized the 4x4 transformation matrices and the kinematic chain theory to obtain the necessary rotations and translations at each joint of the fixator to correct bone deformities at the fracture site. Three-dimensional malalignments with fracture gaps were simulated to correct the deformities. Due to the redundant pair variables in the fixator joints and other problems in obtaining unique solutions, an optimization technique was used to solve the governing linkage loop equations. For each adjustment solution, the bone correction paths were infinite but a unique and optimal reduction path was obtained by applying corrections to all joints simultaneously and in small increments. When the deformity exceeded a certain range, no admissible solution could be obtained, partially due to the limitation of the unilateral fixator configuration and partially due to the restricted joint rotation and translation in the fixator design. The present models and analysis technique can be used to investigate a fixator's adjustability to correct a 3-D bone deformity at a fracture or lengthening site facilitating patient care planning and medical personnel training. PMID- 12126665 TI - The importance of being light: aerodynamic forces and weight in ski jumping. AB - Many contemporary world class ski jumpers are alarmingly underweight and several cases of anorexia nervosa have come to light. Athletes strive for low body weight because it gives them a major competitive advantage. In order to stop this hazardous development, changes to the regulations are being discussed, and the International Ski Federation and the International Olympic Committee wish to be proactive in safe guarding the interest of the athletes and their health. This study of ski jumping uses field studies conducted during World Cup competitions, large-scale wind tunnel measurements with 1:1 models of ski jumpers in current equipment and highly accurate computer simulations of the flight phase that include the effects due to the athlete's position changes. Particular attention has been directed to the design of a reference jump that mirrors current flight style and equipment regulations (2001), and to the investigation of effects associated with variation in body mass, air density, and wind gusts during the simulated flight. The detailed analysis of the physics of ski jumping described here can be used for the investigation of all initial value and parameter variations that determine the flight path of a ski jumper and will form a reliable basis for setting regulations that will make it less attractive or even disadvantageous for the athlete to be extremely light. PMID- 12126666 TI - In vitro biomechanical analysis of glenoids before and after implantation of prosthetic components. AB - Biomechanical investigations are yet necessary to better understand the origin of the glenoid loosening which is the main reason for revision surgery. The aim of this study was to analyse the behaviour of cadaveric glenoids before and after implantation of glenoid prostheses. For that, we developed a new experimental protocol allowing measurement of bone strains and implants displacements under various loading cases. Ten pairs of fresh cadaveric scapulae were tested. Two kinds of loads were applied on the intact glenoid: physiological loads corresponding to a 0-180 degrees abduction and anteflexion movements, and 500N loads. The glenoids were then implanted with a keeled or pegged cemented polyethylene implant. The same previous 500N loads were then applied on the implanted glenoids. Strains were measured using six strain gages placed on precise points around the peripheral cortex of the glenoid. Displacements of implants under loading were measured using two CCD cameras. Maximum strains were obtained between 60 degrees and 120 degrees of abduction or anteflexion. They were located at the anterior and antero-superior parts of the glenoid during abduction and at the posterior and postero-superior parts during anteflexion. Implantation of a prosthetic component generally seemed to increase strains, but tensile strains were decreased at the postero-inferior part with the antero inferior loading point.Some differences were observed between the implants, but they have to be confirmed by further experiments. The great number of data obtained for intact scapulae could be used for a better understanding of glenoid behaviour and for validation of finite element models. PMID- 12126667 TI - A one-dimensional model for the propagation of transient pressure waves through the lung. AB - The propagation of pressure waves in the lung has been investigated by many authors concerned with respiratory physiology, ultrasound medical techniques or thoracic impact injuries. In most of the theoretical studies, the lung has been modeled as an isotropic and homogeneous medium, and by using Hooke's constitutive law (see e.g. Ganesan et al. Respir. Physiol. 110 (1997) 19; Jahed et al. J. Appl. Physiol. 66 (1989) 2675; Grimal et al. C.R. Acad. Sci., Paris 329 (IIb) (2001) 655-662), or more elaborated material laws (see, e.g. Bush and Challener (Proceedings of the International Research Council on Biokinetics Impacts (IRCOBI), Bergish-gladbach, 1988); Stuhmiller et al. J. Trauma 28 (1988) S132; Yang and Wang, Finite element modeling of the human thorax. Web page: http://wwwils.nlm.nih.gov/research/visible/vhpconf98/AUTHORS/YANG/YANG.HTM.). The hypothesis of homogeneous medium may be inappropriate for certain problems. Because of its foam-like structure, the behavior of the lung-even if the air and the soft tissue are assumed to behave like linearly elastic materials-is susceptible to be frequency dependent. In the present study, the lung is viewed as a one-dimensional stack of air and soft tissue layers; wave propagation in such a stack can be investigated in an equivalent mass-spring chain (El-Raheb (J. Acoust. Soc. Am. 94 (1993) 172; Int. J. Solids Struct. 34 (1997) 2969), where the masses and springs, respectively, represent the alveolar walls and alveolar gas. Results are presented in the time and frequency domains. The frequency dependence (cutoff frequency, variations in phase velocity) of the lung model is found to be highly dependent on the mean alveolar size. We found that short pulses induced by high velocity impacts (bullet stopped by a bulletproof jacket) can be highly distorted during the propagation. The pressure differential between two alveoli is discussed as a possible injury criterion. PMID- 12126668 TI - Combination of topological parameters and bone volume fraction better predicts the mechanical properties of trabecular bone. AB - Trabecular bone structure may complement bone volume/total volume fraction (BV/TV) in the prediction of the mechanical properties. Nonetheless, the direct in vivo use of information pertaining to trabecular bone structure necessitates some predictive analytical model linking structure measures to mechanical properties. In this context, the purpose of this study was to combine BV/TV and topological parameters so as to better estimate the mechanical properties of trabecular bone. Thirteen trabecular bone mid-sagittal sections were imaged by magnetic resonance (MR) imaging at the resolution of 117 x 117x 300 microm(3). Topological parameters were evaluated in applying the 3D-line skeleton graph analysis (LSGA) technique to the binary MR images. The same images were used to estimate the elastic moduli by finite element analysis (FEA). In addition to the mid-sagittal section, two cylindrical samples were cored from each vertebra along vertical and horizontal directions. Monotonic compression tests were applied to these samples to measure both vertical and horizontal ultimate stresses. BV/TV was found as a strong predictor of the mechanical properties, accounting for 89 94% of the variability of the elastic moduli and for 69-86% of the variability of the ultimate stresses. Topological parameters and BV/TV were combined following two analytical formulations, based on: (1) the normalization of the topological parameters; and on (2) an exponential fit-model. The normalized parameters accounted for 96-98% of the variability of the elastic moduli, and the exponential model accounted for 80-95% of the variability of the ultimate stresses. Such formulations could potentially be used to increase the prediction of the mechanical properties of trabecular bone. PMID- 12126669 TI - A morphological model of vertebral trabecular bone. AB - In their micro-structures, typical natural cellular materials such as vertebral trabecular bone have a network of doubly tapered struts, thickening near the strut joints. However, past analytical models for vertebral trabecular bone do not take account of the effect of strut taper on the mechanical properties. This paper presents an analytical cell model comprised of doubly tapered struts to predict the global mechanical properties of vertebral trabecular bone. The predicted results for male, female, and both sexes fit the experimental data well. By considering several strut taper geometries, it is shown that the horizontal Young's modulus and the horizontal uniaxial collapse stress are, in some cases, approximately 1.8- and 2.2-fold higher, respectively, than those of the uniform strut model. This finding illustrates the importance of increased trabecular thickening near the strut joints (i) for improving the accuracy of calculating the mechanical properties and (ii) for the effective treatment of aged bone using drug therapy. It also highlights the need to combine trabecular architecture measurements with information about the morphology near the strut joints. PMID- 12126670 TI - Numerical simulation of steady flow fields in a model of abdominal aorta with its peripheral branches. AB - In the present study, a numerical calculation procedure based on a finite volume method was developed to simulate steady flow fields in a model of abdominal aorta with its peripheral branches. The study focused on the steady baseline flow fields and the wall shear stress (WSS) distribution as well as the localization of the reversed flow regions and results were compared to those obtained by other investigators. In the case of resting conditions, the existence of a region of reversed flow of about one to two diameters in size and next to the renal arteries and along the posterior wall as observed by other researchers was confirmed. However, under the exercise conditions this region could be wiped out. The flow reversal along the lateral walls proximal to the bifurcation persisted in both rest and exercise conditions. The WSS distribution and the wall shear stress gradient distribution were obtained. The lowest WSS occurred near the ostia of the renal arteries and the lateral walls of the iliac arteries. And the highest is always at the turn to the branch. The results were generally consistent with those obtained experimentally and numerically by other investigators. It was also shown that the steady flow might be used to depict the averaged behavior of pulsatile flow. The present computer code provides a platform for the future more realistic simulations. PMID- 12126671 TI - Hierarchical genetic algorithm versus static optimization-investigation of elbow flexion and extension movements. AB - The applicability of static optimization (and, respectively, frequently used objective functions) for prediction of individual muscle forces for dynamic conditions has often been discussed. Some of the problems are whether time independent objective functions are suitable, and how to incorporate muscle physiology in models. The present paper deals with a twofold task: (1) implementation of hierarchical genetic algorithm (HGA) based on the properties of the motor units (MUs) twitches, and using multi-objective, time-dependent optimization functions; and (2) comparison of the results of the HGA application with those obtained through static optimization with a criterion "minimum of a weighted sum of the muscle forces raised to the power of n". HGA and its software implementation are presented. The moments of neural stimulation of all MUs are design variables coding the problem in the terms of HGA. The main idea is in using genetic operations to find these moments, so that the sum of MUs twitches satisfies the imposed goals (required joint moments, minimal sum of muscle forces, etc.). Elbow flexion and extension movements with different velocities are considered as proper illustration. It is supposed that they are performed by two extensor muscles and three flexor muscles. The results show that HGA is a suitable means for precise investigation of motor control. Many experimentally observed phenomena (such as antagonistic co-contraction, three-phasic behavior of the muscles during fast movements) can find their explanation by the properties of the MUs twitches. Static optimization is also able to predict three-phasic behavior and could be used as practicable and computationally inexpensive method for total estimation of the muscle forces. PMID- 12126672 TI - Predicting the kinetics of cell spreading. AB - We apply the wetting theory to predict the kinetics of fibroblast spreading onto an adhesive substrate, under simplifying assumptions on the cell structure and geometry. Three main parameters are used: cytoplasmic viscosity, cortical tension, and cell-to-substrate adhesion energy. The viscosity and tension values are taken from previous micromechanical studies. The adhesion energy, ill known, is adjusted by fitting the model predictions to available experimental data of contact radius versus time. The agreement is quite good, justifying such a "macroscopic" view of cell morphology. PMID- 12126673 TI - Reconstruction of shoulder function using a reflected long head biceps: a moment arm study. AB - A tendon transfer technique is proposed for the reconstruction of the paralyzed shoulders secondary to brachial plexus injury. This innovative technique does not require bone-to-bone or tendon to-bone fixation, and attempts to overcome other clinical limitations such as those due to insufficient length of donor muscle. The approach is referred to as the reflected long head biceps (RLHB) technique. The long head of biceps tendons is utilized as a bridging tendon graft. Two surgical alternatives, namely the through-deltoid (TD) pathway and the sub deltoid (SD) pathway, were studied. The moment arms of the transferred tendons were assessed and reported. The TD technique yielded a larger moment than the SD technique. In the plane 30 degrees anterior to the scapular plane, the average moment arms were 3.8cm TD and 3.0cm SD at zero elevation. Such differences tended to further widen with increasing elevation. At 80 degrees elevation, the moment arms became 3.2cm TD and 1.2cm SD. The results supported the clinical feasibility of this RLHB tendon transfer approach. PMID- 12126674 TI - Technique and first clinical results of robot-assisted total knee replacement. AB - Total knee replacement (TKR) is a common procedure for treatment of severe gonarthrosis, but the outcome may be unsatisfactory due to primary malalignment of the prosthetic components. In order to improve precision and accuracy of this surgical procedure, a commercial robotic surgical system (CASPAR) has been adapted to assist the surgeon in the preoperative planning and intraoperative execution of TKR. So far, 70 patients with idiopathic gonarthrosis were successfully treated with a robot-assisted technique in our institution. No major adverse events related to the use of the robotic system have been observed. The mean difference between preoperatively planned and postoperatively achieved tibiofemoral alignment was 0.8 degrees (0-4.1 degrees ) in the robotic group vs. 2.6 degrees (0-7 degrees ) in a manually operated historical control group of 50 patients. A clear advantage of robot-assisted TKR seems to be the ability to execute a highly precise preoperative plan based on computed tomography (CT) scans. Due to better alignment of the prosthetic components and improved bone implant fit, implant loosening is anticipated to be diminished which may be most evident in non-cemented prostheses. Current disadvantages such as the need for placement of fiducial markers, increased operating times and higher overall costs have to be resolved in the future. PMID- 12126675 TI - Long-term assessment of arthroscopic meniscus repair: a 13-year follow-up study. AB - A prospective study was set up to evaluate meniscal suturing using an inside-out technique. Of an initial group of 20 patients who underwent closed meniscus repair between 1985 and 1988 using an inside-out technique, 13 were studied. All patients were subjected to a clinical examination and a magnetic resonance imaging (MRI) investigation. The findings were compared with those of their previous follow-up examination (1994). The Hospital for Special Surgery (HSS) knee rating system (R.G. Stone et al. Athroscopy 1990; 73-78) was used. The study included seven men and six women, ranging in age from 29 years to 50 years (mean age: 35 years 6 months). The mean follow-up was 13 years 2 months (11 years 11 months-15 years 4 months). Six left and seven right knees were involved. Seven patients also had an anterior cruciate ligament (ACL) injury of which one was repaired 6 years after meniscal repair. All patients obtained an HSS score of more than 75%. In all patients, the site of the previous suture was still visible on MRI mainly by small metal artefacts in the meniscus. Patients with an unrepaired ACL lesion had an early onset of arthrosis and cartilage degeneration. Meniscal suturing gives good clinical long-term results. Magnetic resonance imaging, however, showed signs of mucoid degeneration or scar tissue in 46% of the patients. PMID- 12126676 TI - Increase in range of knee motion to obtain floor sitting after high tibial osteotomy for osteoarthritis. AB - In order to obtain better range of motion in knees with osteoarthritis, medial and lateral parapatellar retinaculo-capsular release operations were performed at the time of high tibial osteotomy, with fixation using dual plating on medial compartmental knee osteoarthritis (29 knees) or spontaneous osteonecrosis of the medial femoral condyle (15 knees). At removal of the blade plate, 1 to 2 years after the initial osteotomy, the same release procedures were performed together with resection of adhesive soft tissue and resection of osteophytes, which were obstacles to full flexion. A grading system (Grade 0-3) was proposed to evaluate the duration (min) of formal floor sitting. After these procedures, the patients were able to sit on the floor with 155-165 degrees of flexion for more than 30 min (Grade 3) in 20 knees, for 10-29 min (Grade 2) in seven, for less than 10 min (Grade 1) in nine and were unable to sit on the floor (Grade 0) in eight knees. Maximum knee flexion and total range of motion were 142+/-8.4 degrees and 137+/ 11 degrees before and 152+/-6.6 degrees and 151+/-7.4 degrees after surgery, respectively. The American Knee Society Knee Score and Function Score were 61+/ 17 and 46+/-16 before, and 97+/-5 (P<0.0001) and 91+/-13 (P<0.0001) after surgery at the final follow-up, respectively. The femoro-tibial angle in standing with one leg was 183+/-6 degrees (3 degrees of anatomical varus angulation) before and 170+/-3 degrees (P<0.0001) (10 degrees of anatomical valgus angulation) after surgery. PMID- 12126677 TI - Arthroscopic ACL reconstruction: a 5-9 year follow-up. AB - Arthroscopic ACL reconstruction has a satisfactory functional outcome of up to 90%, but there are few long-term prospective studies. This prospective study presents the outcomes of ACL reconstruction in terms of laxity, function and degenerative change, after a mean follow-up of 7 years. Function was assessed using the Lysholm and Tegner Activity Scores, laxity using the Stryker Knee Laxity Tester, employing maximum manual effort, and degenerative change was assessed as joint line narrowing on standardised radiographs. At latest follow up, the mean Lysholm score improved from 70 to 87 and the Tegner from 4 to 7 (P<0.001). AP translation also improved (P<0.001). The incidence of early degenerative change was 50% and although this appeared to be associated with a previous meniscectomy, the correlation was not significant (P=0.06). In conclusion, the improved functional scores and laxity are sustained beyond 7 years but the 50% incidence of early degenerative change may be a cause for concern. PMID- 12126678 TI - Osteochondritis dissecans of the weight-bearing surface of the medial femoral condyle in adults. AB - Osteochondritis dissecans (OD) is a syndrome that can be characterized as a non infectious disturbance of enchondral ossification or as a post-traumatic event. OD occurs in the joint cartilage and physis of long bones, as well as in the talus. The medial femoral condyle is the most commonly affected site. OD of the weight-bearing, inferocentral portion of medial femoral condyle is an uncommon, but still challenging issue in knee surgery. This study reports one surgeon's experience in the treatment of OD of the weight-bearing surface of the medial femoral condyle in adult patients. A total of 29 knees in 28 patients with OD were reviewed as the basis of this study. Four patients were women and 24 were men. The average age was 29.5 years. Patients were observed for an average of 2 years after surgery. Medial joint line tenderness, anterior knee pain, stiffness and locking were the major complaints in 60% of cases. Arthroscopic excision of loose bodies with or without drilling of the crater and fixation of the lesions with 2 K-wires, with or without bone grafting, was undertaken for the patients. In our study, the clinical outcome was excellent in 11 patients, good in 13, fair in four and poor in one. On subjective questioning, all patients reported marked improvement and satisfaction with the surgery. PMID- 12126679 TI - A potential animal model for creating a controlled and reversible anterior cruciate ligament insufficiency. AB - We developed and tested a device to manipulate the axial position of the tibial anterior cruciate ligament (ACL) insertion in vitro to create a potential animal model that could simulate both ACL insufficiency and 'optimal' ACL reconstruction. This model is based on the concept that controlled incremental proximal displacement of the tibial ACL insertion simulates ACL insufficiency. Replacing the insertion at the joint level and then adjusting its position until the forces recorded equaled those in the ACL-intact knee can simulate 'optimal' ACL reconstruction. Anterior tibial translation (ATT) was quantified in vitro in 24 sheep cadaver knees with the ACL intact and after the ACL was cut or detached (ACL insufficiency). In 8 knees, a bone plug including the tibial ACL insertion was detached, mounted in a specially designed device, and adjusted to reproduce ATT of the ACL-intact knee. ATT was then measured after proximal displacement of the tibial ACL insertion in calibrated 1 mm increments. The results revealed that detaching the ACL increased ATT by 132-700%. Controlled 3 mm proximal displacement of the insertion using this device increased ATT by more than 100%. Comparing the increase in ATT due to controlled displacement of the ACL insertion to that due to detaching the ACL, in only one case was the same magnitude of ACL insufficiency reproduced. Despite the variability between knees, the device was able to reproduce ATT of the ACL-intact knee and to substantially increase ATT with controlled proximal displacement of the tibial ACL insertion. Use of this device, if successful in an in vivo ACL reconstruction model, could help define any quantitative association between altered joint kinematics and degenerative changes in the joint. PMID- 12126680 TI - Reconstruction of soft tissue defects following total knee arthroplasty. AB - Soft tissue defects following total knee arthroplasty can represent serious problems for the patient and the surgeon. Perioperative soft tissue complications can result in loss of the prosthesis or limb. In this study, we present 17 cases with complex wounds following total knee arthroplasty who had surgery between May 1994 and July-2001. Patient-related factors, wound factors, surgical operation, secondary procedures, and duration of follow-up have been analysed for each patient. After local wound care and debridement, soft tissue defects have been covered with either a fasciocutaneous or gastrocnemius myocutaneous flap. All the knees (100%) have been salvaged although in 1 patient (6%) replacement of the prosthesis was necessary. In 5 patients (30%) secondary surgical procedures have been performed. Even though there is no consensus in the management of soft tissue defects following total knee arthroplasty, adequate wound care, including identification of infection, debridement, and early appropriate defect coverage should be the main points to consider. PMID- 12126681 TI - Accelerated recovery for unicompartmental knee replacement--a feasibility study. AB - A pilot study assessed the feasibility of discharging NHS patients undergoing knee replacement within a day of surgery. Seven patients with medial compartment osteoarthritis were recruited after fulfilling strict exclusion criteria. Pre operative assessment revealed that all patients had significant dysfunction and pain before operation. They had medial unicompartmental replacement through a short incision without dislocation of the patella. Each patient underwent an accelerated recovery program that included pain control, physiotherapy and self assessment. Patients were mobilised immediately after operation. Follow-up assessment was performed at 1, 2 and 6 weeks after surgery. All patients, except one (who failed to go home because of an administrative error), returned home the day after surgery. The average pain score for the first 2 weeks after surgery was 2/10. At 6 weeks, knee flexion averaged 125 degrees and all patients were walking independently and painfree. The new protocol allows for early, safe discharge of patients undergoing unicompartmental knee replacement. PMID- 12126682 TI - The use of knee splints after total knee replacements. AB - The aim of this randomised prospective study was to establish whether the use of knee splints following total knee replacement is necessary. The study included 81 patients undergoing total knee replacement who were randomised into a 'splint' and a 'no splint' group post-operatively. The following parameters were recorded: The range of movement pre-operatively, 5 days post-operatively and 6 weeks post operatively; the length of time to straight leg raise; the blood drained from the wound; and the amount of post-operative analgesia required. We found that patients in the 'no splint' group achieved significantly greater flexion at 5 days and 6 weeks post-operatively but drained significantly more blood from the wound. Transfusion requirements were similar in the two groups. There was no other significant difference in the parameters measured between the two groups. In conclusion we found no evidence to advocate the use of knee splints following total knee arthroplasty. PMID- 12126683 TI - Is routine splintage following primary total knee replacement necessary? A prospective randomised trial. AB - It was hypothesised that routine splintage following primary total knee replacement has no affect on flexion deformity and offers no benefit over simple wool and crepe. Fifty-five patients undergoing primary total knee replacement were entered into a prospective study. The patients were randomly assigned to two groups: The first group was rehabilitated without a splint and the second received an adjustable semi-rigid extension splint (Richards splint) for the first 48 h after surgery. Range of motion measurements were recorded pre operatively and at 2 days, 1 week and 3 months post-operation by a research nurse blinded to the allocation. No statistically significant difference in flexion deformity was found at any stage (P>0.5). No difference was found in general or wound complications, or requirement for blood transfusion, and the post-operative stay was equal in the two groups. We conclude that routine use of a semi-rigid splint following primary total knee replacement has no advantage over simple wound dressings. PMID- 12126684 TI - The importance of early diagnosis in the management of proximal tibiofibular dislocation: a 9- and 5-year follow-up of a bilateral case. AB - Unilateral fibula head dislocation is an uncommon injury. Ogden in 1974 reviewed the literature and detected 108 cases [J. Bone Joint Surg. 56(A) (1974) 145-154]. There since have been few reported cases. Bilateral fibula head dislocation only has been reported once [Rheumatol. Int. 5(1) (1984) 45-47]. We report a patient who had developed bilateral fibula head dislocations, each side independently over a 5-year period. We emphasise the rarity of the injury, the necessity of prompt recognition and reduction, and the uncertainty for best management of this injury. PMID- 12126685 TI - A kinematic study of lateral unicompartmental arthroplasty. AB - When the Oxford unicompartmental meniscal bearing arthroplasty (UCA) is used in the lateral compartment 10% of the bearings dislocate. A fluoroscopic study was performed to investigate if abnormal mid-sagittal plane kinematics was related to bearing dislocation. Video fluoroscopy is an accepted means of determining in vivo knee kinematics in the sagittal plane. Video fluoroscopy was obtained of 5 Oxford lateral UCAs 10 years post-operatively and of five normal knees. Patellar tendon angle (PTA), derived from dynamic fluoroscopic images, was used to describe the joint kinematics. This in-vivo experiment demonstrated that the PTA/knee relationship for the Oxford lateral UCA is similar to the normal knee. Both the normal knee (r(2)=0.99) and the Oxford lateral UCA (r(2)=0.98) demonstrated a linear relationship between flexion angle and PTA. No significant difference in PTA was found between the normal knee and the Oxford lateral UCA. This study demonstrated normal kinematics, as indicated by PTA, ten years after implantation of the Oxford lateral UCA. It is therefore reasonable to suggest that abnormal kinematics is not a significant factor relating to meniscal bearing dislocation in the lateral compartment. PMID- 12126686 TI - Transient osteoporosis of the femoral condyle: a case report. AB - In this paper, we describe a patient with transient osteoporosis of the femoral condyle. A differential diagnosis should be made for osteonecrosis, infectious disorders, and infiltrative neoplasms based on the normal laboratory findings and diffuse bone edema pattern in MRI. Since this disorder is self-limiting, both the surgeon and clinician should be aware of this condition and must avoid unnecessary intervention. PMID- 12126687 TI - Failure of anterior cruciate ligament reconstruction following calcification of the graft. AB - Anterior cruciate ligament (ACL) reconstruction is a commonplace orthopaedic procedure. It is most commonly performed using an autologous bone-patella tendon bone graft, although other donor sites are also used. Failure of the graft can occur for a variety of reasons. We describe here failure of such an ACL reconstruction in a young man secondary to ectopic calcification of the graft, which led to excessive rigidity and its subsequent avulsion from the proximal attachment. This case illustrates the benefits of being able to reconstruct the ACL by more than one technique, which can be used if another method fails. PMID- 12126688 TI - Osteophyte impingement of the popliteus tendon as a cause of lateral knee joint pain. AB - Three cases of symptomatic popliteus tendon impingement on an osteophyte are presented. In only one case did the patient complain of a painful 'snapping' on flexing and extending the knee. Diagnosis was made when tenderness was elicited on palpating the osteophyte during flexion and extension with varus stress on the knee. It may be difficult to determine whether such a prominence represents an osteophyte or an exaggerated normal lower border of the popliteus groove. Two cases were successfully treated by arthroscopic excision of the osteophyte and the remaining case by resection of the popliteus tendon. PMID- 12126689 TI - Bucket handle tear of medial plica. AB - A case of locking of the knee due to a 'bucket handle tear' of the medial plica is described. The sign of clunking could be relieved by pushing the patella medially, so reducing the plica from the trochlea into the medial gutter. Arthroscopic resection cured the condition. There may be association with patellar instability, possibly the lesion is more common than previously recognised. PMID- 12126690 TI - Nottingham knee symposium 2001. PMID- 12126693 TI - Significance of ROS in oxygen sensing in cell systems with sensitivity to physiological hypoxia. AB - Reactive oxygen species (ROS) are oxygen-containing molecular entities which are more potent and effective oxidizing agents than is molecular oxygen itself. With the exception of phagocytic cells, where ROS play an important physiological role in defense reactions, ROS have classically been considered undesirable byproducts of cell metabolism, existing several cellular mechanisms aimed to dispose them. Recently, however, ROS have been considered important intracellular signaling molecules, which may act as mediators or second messengers in many cell functions. This is the proposed role for ROS in oxygen sensing in systems, such as carotid body chemoreceptor cells, pulmonary artery smooth muscle cells, and erythropoietin-producing cells. These unique cells comprise essential parts of homeostatic loops directed to maintain oxygen levels in multicellular organisms in situations of hypoxia. The present article examines the possible significance of ROS in these three cell systems, and proposes a set of criteria that ROS should satisfy for their consideration as mediators in hypoxic transduction cascades. In none of the three cell types do ROS satisfy these criteria, and thus it appears that alternative mechanisms are responsible for the transduction cascades linking hypoxia to the release of neurotransmitters in chemoreceptor cells, contraction in pulmonary artery smooth muscle cells and erythropoietin secretion in erythropoietin producing cells. PMID- 12126692 TI - Hypoxic pulmonary vasoconstriction: cyclic adenosine diphosphate-ribose, smooth muscle Ca(2+) stores and the endothelium. AB - Hypoxic pulmonary vasoconstriction (HPV) is unique to pulmonary arteries, and supports ventilation/perfusion matching. However, in diseases such as emphysema, HPV can promote hypoxic pulmonary hypertension (HPH), which ultimately leads to right heart failure. Since it was first described, the mechanisms underpinning HPV have remained obscure, and current therapies for HPH are poor. Previous investigations have suggested that HPV may be mediated by processes intrinsic to the pulmonary artery smooth muscle, and by the release of a vasoconstrictor(s) from the endothelium. It was thought that oxygen-sensitive ion channels in the smooth muscle cell membrane triggered HPV, and it has been argued that the endothelium-derived vasoconstrictor is endothelin-1. However, these proposals remain controversial. This review discusses the regulation by hypoxia of cyclic adenosine diphosphate-ribose production and Ca(2+) release from the sarcoplasmic reticulum in pulmonary artery smooth muscle. The role of these processes in triggering maintained HPV is then related to its subsequent progression due to vasoconstrictor(s) release from the endothelium. PMID- 12126694 TI - Multiple sites of oxygen sensing and their contributions to hypoxic pulmonary vasoconstriction. AB - Oxygen sensing by the pulmonary vasculature is important for the regulation of vessel tone and the matching of lung perfusion to ventilation. Airways hypoxia is a major stimulus for vasoconstriction, which diverts blood from hypoxic alveoli to better ventilated areas of the lung. Several hypotheses have emerged to explain how pulmonary arteries sense a decrease in oxygen and mediate hypoxic pulmonary vasoconstriction (HPV). They differ mainly in where they place the main site of HPV: in the endothelial or smooth muscle cells of the artery wall. HPV probably results from synergistic actions on both cell types, but it can proceed in the absence of endothelium, suggesting that the primary oxygen sensor is the smooth muscle cell and endothelium-derived agents modulate the muscle response. Several oxygen-sensing targets have been identified in smooth muscle, including potassium channels, Ca(2+) stores in the sarcoplasmic reticulum (SR) and the Ca(2+) sensitivity of the contractile proteins. The evidence for different oxygen sensing mechanisms in pulmonary vessels is discussed. PMID- 12126695 TI - Hypoxia, energy state and pulmonary vasomotor tone. AB - Vasomotor responses to hypoxia constitute a fundamental adaptation to a commonly encountered stress. It has long been suspected that changes in cellular energetics may modulate both hypoxic systemic artery vasodilatation (HSV) and hypoxic pulmonary artery vasoconstriction (HPV). Although limitation of energy has been shown to underlie hypoxic relaxation in some smooth muscles, the response to hypoxia in vascular smooth muscle does not appear to be a simple function of energy stores, but instead may involve perturbations of ATP or energy delivery to mechanisms controlling muscle force, and/or changes associated with anaerobic metabolism. Recent work in pulmonary vascular smooth muscle has demonstrated that energy stores are maintained during hypoxic pulmonary vasoconstriction, and that this is dependent on glucose availability and up regulation of glycolysis. There is increasing evidence that glycolysis is preferentially coupled to a variety of membrane associated ATP dependent processes, including the Na(+) pump, Ca(2+)-ATPase, and possibly some protein kinases. These and other mechanisms may influence excitation-contraction coupling in both systemic and pulmonary arteries by effects on intracellular Ca(2+) and/or Ca(2+) sensitivity. Hypoxia has also been postulated to have major effects on other cytosolic second messenger systems including phosphatidylinositol pathways, cell redox state and mitochondrial reactive oxygen species production. This review examines the relationship between energy state, anaerobic respiration and hypoxic vasomotor tone, with a particular emphasis on hypoxic pulmonary vasoconstriction. PMID- 12126696 TI - Carotid body thin slices: responses of glomus cells to hypoxia and K(+)-channel blockers. AB - We describe the rat carotid body thin slice preparation, which allows to perform patch-clamp recording of membrane ionic currents and to monitor catecholamine secretion by amperometry in single glomus cells under direct visual control. We observed several electrophysiologically distinct cell classes within the same glomerulus. A voltage- and Ca(2+)-dependent component of the whole cell K(+) current was reversibly inhibited by low P(O(2)) (20 mmHg). Exposure of the cells to hypoxia elicited the appearance of spike-like exocytotic events. This response to hypoxia was reversible and required extracellular Ca(2+) influx. Addition of tetraethylammonium (TEA, 2-5 mM) to the extracellular solution induced in most (>95%) cells tested a secretory response similar to that elicited by low P(O(2)). Cells non-responsive to hypoxia but activated by exposure to high external K(+) were also stimulated by TEA. A secretory response similar to that of hypoxia or TEA was also observed after treatment of the cells with iberiotoxin to block selectively maxi-K(+) channels. Our data further support the view that membrane ion channels are critically involved in sensory transduction in the carotid body. We demonstrate that in intact glomus cells inhibition of voltage-dependent K(+) channels can contribute to initiate the secretory response to low P(O(2)). PMID- 12126697 TI - O(2) sensing in hypoxic pulmonary vasoconstriction: the mitochondrial door re opens. AB - The identity of the O(2) sensor underlying the hypoxic pulmonary vasoconstriction (HPV) response has been sought for more than 50 years. Recently, the mitochondria have again come into sharp focus as the cellular organelle responsible for triggering the events that culminate in pulmonary artery constriction. Studies from different laboratories propose two disparate models to explain how mitochondria react to a decrease in P(O(2)). One model proposes that hypoxia slows or inhibits mitochondrial electron transport resulting in the accumulation of reducing equivalents and a decrease in the generation of reactive oxygen species (ROS). This is proposed to activate a redox-sensitive pathway leading to pulmonary vasoconstriction. A second and opposing model suggests that hypoxia triggers a paradoxical increase in mitochondrial ROS generation. This increase would then lead to the activation of an oxidant-sensitive signaling transduction pathway leading to HPV. This article summarizes the potential involvement of mitochondria in these two very different models. PMID- 12126698 TI - Acute and chronic hypoxic pulmonary vasoconstriction: a central role for endothelin-1? AB - In the pulmonary circulation, a decrease in oxygen tension results in the development of hypoxic pulmonary vasoconstriction (HPV), although the exact mechanism by which HPV occurs remains unclear. Evidence gathered from many laboratories suggests that while pulmonary arterial smooth muscle cells (PASMCs) can sense and respond to changes in oxygen tension, full expression of HPV requires modulating influences from the endothelium. In this review, we propose a model of HPV, based on recent studies from our laboratory, in which endothelin-1 (ET-1), a vasoactive peptide released from the endothelium, plays a central role and discuss how this model may be involved in the long-term adaptation to hypoxia. PMID- 12126699 TI - Endothelium-derived mediators and hypoxic pulmonary vasoconstriction. AB - The vascular endothelium synthesises, metabolises or converts a multitude of vasoactive mediators, and plays a vital role in the regulation of pulmonary vascular resistance. Its role in hypoxic pulmonary vasoconstriction (HPV) is however controversial. Although HPV has been demonstrated in both pulmonary arteries where the endothelium has been removed and isolated pulmonary artery smooth muscle cells, many reports have shown either partial or complete dependence on an intact endothelium for sustained HPV (> approximately 20 min). However, despite many years of study no known endothelium-derived mediator has yet been unequivocally shown to be essential for HPV, although several may either facilitate the response or act as physiological brakes to limit the extent of HPV. In this article we review the evidence for and against the role of specific endothelium-derived mediators in HPV. We make the case for a facilitatory or permissive function of the endothelium, that in conjunction with a rise in smooth muscle intracellular Ca(2+) initiated by a mechanism intrinsic to smooth muscle, allows the development of sustained HPV. In particular, we propose that in response to hypoxia the pulmonary vascular endothelium releases an as yet unidentified agent that causes Ca(2+) sensitisation in the smooth muscle. PMID- 12126700 TI - Redox signaling in oxygen sensing by vessels. AB - In response to the increase in oxygen tension at birth, the resistance pulmonary arteries dilate, while the ductus arteriosus constricts. Although modulated by the endothelium, these opposite responses are intrinsic to the vascular smooth muscle. While still controversial, it seems likely that during normoxia the production of reactive oxygen species (ROS) increases and the smooth muscle cell cytoplasm is more oxidized in both pulmonary arteries and ductus, compared to hypoxia. However, the effect of changes in the endogenous redox status or the addition of a redox agent, oxidizing or reducing, is exactly opposite in the two vessels. A reducing agent, dithiothreitol, like hypoxia, in the pulmonary artery will inhibit potassium current, cause depolarization, increase cytosolic calcium and lead to contraction. Responses to dithiothreitol in the ductus are opposite and removal of endogenous H(2)O(2) by intracellular catalase in the ductus increases potassium current. Oxygen sensing in both vessels is probably mediated by redox effects on both calcium influx and calcium release from the sarcoplasmic reticulum (SR). PMID- 12126701 TI - Laccase: new functions for an old enzyme. AB - Laccases occur widely in fungi; they have been characterized less frequently in higher plants. Here we have focused on more recent reports on the occurrence of laccase and its functions in physiological development and industrial utility. The reports of molecular weights, pH optima, and substrate specificity are extremely diverse. Conclusive proof of the occurrence of laccase in a tissue must demonstrate that the enzyme be able to oxidize quinol with concomitant uptake of oxygen. Laccase is involved in the pigmentation process of fungal spores, the regeneration of tobacco protoplasts, as fungal virulence factors, and in lignification of cell walls and delignification during white rot of wood. Commercially, laccases have been used to delignify woody tissues, produce ethanol, and to distinguish between morphine and codeine. A very wide variety of bioremediation processes employ laccase in order to protect the environment from damage caused by industrial effluents. Research in recent years has been intense, much of it elicited by the wide diversity of laccases, their utility and their very interesting enzymology. PMID- 12126702 TI - Peroxidase-mediated transformation of hydroxy-9,10-anthraquinones. AB - A peroxidase (EC 1.11.1.7) has been isolated and purified from Senna angustifolia. The enzyme was purified by ion-exchange chromatography on high Q and high S columns. SDS-PAGE electrophoresis showed that the protein has a molecular mass of approximately 70 kDa. Hydroxy-anthraquinones and hydroxy anthracenones were evaluated as substrate of S. angustifolia and horseradish peroxidases. Both peroxidases catalyzed the oxidation of alizarin and purpurin anthraquinones to the corresponding 3,3'-bializarin and the new compound 3,3' bipurpurin, respectively, as well as the formation of 2,2'-biquinizarin from quinizarin anthracenone. The K(Mapp) and V(max) values for alizarin and purpurin were 97 and 95 microM, and 1.5 and 2.1 microM min(-1) mg prot(-1), respectively. The results suggest that peroxidase may participate in the biogenesis of anthraquinones. PMID- 12126703 TI - Ascorbic acid and flavonoid-peroxidase reaction as a detoxifying system of H(2)O(2) in grapevine leaves. AB - Biosynthesis of both ascorbic acid (AsA) and peroxidase activity were induced by light in cv. Sultana grapevine leaves. Induced peroxidase activity mainly involved basic isoenzymes of pI 9.8 and 9.6 and catalyzed the oxidation of flavonoids like quercetin and kaempferol and derivatives of hydroxycinnamic acids such as ferulic and p-coumaric acids, but not AsA. However, the peroxidase dependent oxidation of ferulic acid and quercetin was temporarily suppressed by AsA as long as it remained in the reaction medium. Kinetics and spectroscopic results indicated that AsA was oxidized to dehydroascorbic acid only in the presence of phenols or flavonoids, and did not interfere with the catalytic activity of the peroxidase. Ascorbate peroxidase isoenzymes (APx), whose activities are widely considered central for detoxification of H(2)O(2) in most plant cells, were not detected in grape leaves extracts. The significance of light stimulus on peroxidase activity and leaf AsA content is discussed in terms of a flavonoid-redox cycle proposed as an alternative system to detoxify H(2)O(2) in grapevine leaves. PMID- 12126704 TI - Formation of Bowman-Birk inhibitors during the germination of horsegram (Dolichos biflorus). AB - Three Bowman-Birk type inhibitors (HGGI-I, II and III), which appear in the cotyledons of 120 h germinated horsegram (Dolichos biflorus) seeds have been purified to homogeneity by size-exclusion chromatography and ion-exchange chromatography. HGGI-I, HGGI-II and HGGI-III differ from each other and from the dormant seed inhibitors in amino acid composition, molecular size and charge. The amino-terminal sequence analyses indicate that these inhibitors are derived from the isoinhibitors of the dormant seed by a limited proteolysis and not by de novo synthesis. These inhibitors differ from each other at their amino-terminus. HGGI II identical to HGGI-I except for the loss of a single amino-terminal aspartyl residue, where as HGGI-III shows the loss of a pentapeptide. All the three inhibitors are potent competitive inhibitors of trypsin and chymotrypsin. The dissociation constants (K(i)s) for trypsin inhibition indicate that amino terminal tail of the inhibitors play a role in trypsin binding probably through electrostatic interaction. PMID- 12126705 TI - In vitro properties of a recombinant flavonol synthase from Arabidopsis thaliana. AB - cDNA corresponding to a flavonol synthase gene from Arabidopsis thaliana was cloned and expressed in Escherichia coli. The recombinant protein was purified to near-homogeneity and the catalytic properties of the enzyme were studied in vitro. Together with kaempferol and apigenin the recombinant protein synthesised the (2R,3S)-cis- and (2S,3S)-trans-isomers of dihydrokaempferol from the (2S)- and (2R)-isomers of naringenin, respectively. Flavanones and dihydroflavanols differing in degree of A- or B-ring hydroxylation were also accepted as substrates. PMID- 12126706 TI - Expression of the vanadium-dependent bromoperoxidase gene from a marine macro alga Corallina pilulifera in Saccharomyces cerevisiae and characterization of the recombinant enzyme. AB - The vanadium-dependent bromoperoxidase from the marine macro-alga Corallina pilulifera was heterologously expressed in Saccharomyces cerevisiae. The enzyme was purified and crystals in "tear drop" form were obtained. The catalytic properties of the recombinant enzyme were studied and compared with those of the native enzyme purified from C. pilulifera. Differences in thermal stability and chloroperoxidase activity were observed. The recombinant enzyme retained full activity after preincubation at 65 degrees C for 20 min, but the native enzyme was completely inactivated under the same conditions. The chlorinating activity of the native enzyme was more than ten times higher than that of the recombinant enzyme. Other properties, such as K(m) values for KBr and H(2)O(2), and optimal temperature and pH, were similar for each source of C. pilulifera bromoperoxidase. PMID- 12126707 TI - Dynamic changes in cell wall polysaccharides during wheat seedling development. AB - Changes in arabinoxylan content and composition during development of wheat seedlings were investigated. The cell walls isolated from the seedlings showed an increasing content of arabinoxylan during development, which could be correlated to increased activity of xylan synthase and arabinoxylan arabinosyltransferase. Arabinoxylan changed from initially having a high degree of arabinose substitution to a much lower degree of substitution. beta-Glucan was present in the walls at the early stages of development, but was actively degraded after day 4. Increased deposition of arabinoxylan did not take place until beta-glucan had been fully degraded. Ferulic and p-coumaric acid esters were present at all points but increased significantly from day 3 to 6, where lignification began. Ferulic acid dimers did not appear in the cell wall until day three and the different ferulic acid dimers varied in the course of accumulation. The ratio of ferulic acid dimers to free ferulic acid was maximal at the time when the wall had been depleted for beta-glucan, which had not yet been fully replaced by arabinoxylan. This pattern suggests a role for ferulic acid dimers in stabilizing the wall during the transition from a flexible to a more rigid structure. To investigate if the same changes could be observed within a single seedling, 7 day old seedlings were divided into four sections and the walls were analyzed. Some of the changes observed during the seedling development could also be observed within a single seedling, when analyzing the segments from the elongation zone at the base to the top of the leaf. However, the expanding region of older seedlings was much richer in hydroxycinnamates than the expanding region of younger seedlings. Diferulic acids are stabilizing the wall in the transition phase from an expanding to a mature wall. This transition can take place in different manners depending on the cell and tissue type. PMID- 12126708 TI - Emission of floral volatiles from Mahonia japonica (Berberidaceae). AB - Flowering Mahonia japonica plants were subjected to controlled environments and the floral volatiles emitted from whole racemes (laterals) were trapped by Porapak Q adsorbent and analysed by GC-FID. An experiment with photoperiods of 6 and 9 h at constant temperature (10+/-1 degrees C) demonstrated that photoperiod was the stimulus for enhanced emission of most volatiles. Small quantitative differences in emitted fragrance composition were observed between light and dark periods and between plants acclimatised to different photoperiods. Maximum rates of emission occurred in the middle of the light period; aromatic compounds (benzaldehyde, benzyl alcohol and indole) displayed a more rapid increase and subsequent decline compared with monoterpenes (cis- and trans-ocimene and linalool). When the photoperiod was extended from 6 to 9 h, maximum rates of emission continued throughout the additional 3 h. Total emission (microg/h) of volatiles was 2-fold greater in the day-time (DT) (39.7 microg/h) compared with the night-time (NT) (19.8 microgg/h) under a 6 h photoperiod and was not significantly different from total emission under a 9 h photoperiod. PMID- 12126710 TI - Taxol content in the bark of Himalayan Yew in relation to tree age and sex. AB - Taxol content in the bark of Taxus baccata trees growing in a homogenous (uniform) environment at Jageshwar, District Almora in Central Himalaya has been quantified. The average taxol concentration in the bark of sampled trees was 0.0558+/-0.008% (of dry wt.) and was about 64% higher for male plants (averaged across tree age) in comparison to female trees. Maximum taxol content was recorded in the bark samples collected from trees of >110 yrs age. ANOVA indicates a significant difference in the taxol content of bark from trees of different ages, however, differences were not significant between sexes. Taxol was quantified by HPLC using a standard curve prepared with authentic taxol; the identification of bark taxol was confirmed by UV and mass spectrometry. The total taxol content of the bark of Taxus trees across an age series was found to range between 0.064 to 8.032 g/tree, and a tree of about 100 yrs age can yield 5.74 kg dry bark. PMID- 12126709 TI - Accumulation of chloroplast-targeted lipoxygenase in passion fruit leaves in response to methyl jasmonate. AB - Wounding caused local and systemic induction of lipoxygenase (LOX) activity in passion fruit (Passiflora edulis f. flavicarpa) leaves, while exposing intact plants to methyl jasmonate (MJ) vapor provoked a much stronger response. Western blot analysis of these leaf protein extracts using polyclonal antibodies against cucumber LOX, revealed an accumulation of a 90 kDa protein, consistent with LOX enzymatic assays. The inducible LOX was purified to apparent homogeneity, and in vitro analysis of LOXactivity using linoleic acid as substrate showed that it possesses C-13 specificity. Immunocytochemical localization studies using leaf tissue from MJ-treated plants demonstrated that the inducible LOX was compartmented in large quantities in the chloroplasts of mesophyll cells, associated with the stroma. The results suggest that the wound response in passion fruit plants may be mediated by a chloroplast 13-LOX, a key enzyme of the octadecanoid defense-signaling pathway. PMID- 12126711 TI - Phenanthrenoids from the wetland Juncus acutus. AB - Nine 9,10-dihydrophenanthrenes, three phenanthrenes and a related pyrene have been isolated from the wetland plant Juncus acutus. The structures have been attributed by means of their spectral data and chemical correlation. 5-(1-Ethoxy ethyl)-2-hydroxy-7-methoxy-1,8-dimethyl-9,10-dihydrophenanthrene and 5-(1-phytoxy ethyl)-2-hydroxy-7-methoxy-1,8-dimethyl-9,10-dihydrophenanthrene, 2,7-dihydroxy-1 methyl-5-vinylphenanthrene, 2,7-dimethoxy-1,6-dimethyl-5-vinylphenanthrene and 2,7-dihydroxy-1,6-dimethylpyrene are described for the first time. Many of the compounds showed in vitro phytotoxicity against Selenastrum capricornutum, a microalga used in aquatic tests. PMID- 12126712 TI - Glycosides of polyenoic branched fatty acids from myxomycetes. AB - The determination of chemical structures of five novel compounds, i.e. one multibranched polyunsaturated fatty acid ((2E,4E,7S,8E,10E,12E,14S)-7,9,13,17 tetramethyl-7,14-dihydroxy-2,4,8,10,12,16-octadecahexaenoic acid) and its four glycosides from seven different myxomycetes is described. The absolute configuration of both hydroxyl groups was determined. The glycosides containing glucose, mannose and rhamnose. These compounds were identified by means of 1H and 13C NMR, MS, UV and IR spectra. Three of them were identified in Arcyria cinerea (Bull.) Pers., two in A. denudata (L.) Wetts., and A. nutans (Bull.) Grev., Fuligo septica (L.) Wigg., Lycogala epidendrum (L.) Fries, Physarum polycephalum Schwein., and Trichia varia Pers. contained one of the identified glycosides each. PMID- 12126716 TI - The role of the virology laboratory in the management of hepatitis C virus infection. PMID- 12126717 TI - Treat or test first? Decision analysis of empirical antiviral treatment of influenza virus infection versus treatment based on rapid test results. AB - BACKGROUND: neuraminidase (NA) inhibitors have recently become available for treatment of influenza. Rapid antigen detection assays at 'point-of-care' may improve the accuracy of clinical diagnosis, but the value of these techniques in assisting with the appropriate use of antivirals remains controversial. OBJECTIVE: to compare the diagnostic utilities of two management strategies for influenza, empirical antiviral therapy versus therapy based on a positive rapid test result in pre-epidemic and epidemic periods. STUDY DESIGN: a threshold decision analytic model was designed to compare these competing strategies and sensitivity analysis performed to examine the impact of diagnostic variables on the expected utility of the decision with a range of prior probabilities of infection between 1 and 50%. RESULTS: on the basis of the calculated sensitivity (77%) and specificity (95%) of a point-of-care test for influenza, pre-treatment testing was preferred and cost-effective in non-epidemic stage of the influenza cycle. The alternative strategy of empirical treatment produces a higher utility value during epidemics, but may result in overuse of antivirals for low-risk populations. The two strategies had equivalent efficacy when the probability of influenza was 42%. CONCLUSIONS: Patients with flu-like illness, who present outside the influenza outbreak and are considered to be at low risk for influenza related complications, should be tested to confirm the diagnosis before starting antiviral treatment with a NA inhibitor. The most important variables in the model were the accuracy of the clinical diagnosis and the pre-test probability of influenza. A threshold probability of influenza of 42% would dictate changing from the rapid testing strategy to a 'treat regardless' strategy. PMID- 12126718 TI - Prevalence of tick-borne-encephalitis virus antibodies in Lithuania. AB - BACKGROUND: Clinical infections caused by tick-borne encephalitis virus (TBEV) are quite common in Lithuania and cause significant disease burden not only as acute cases but as chronic sequealeae as well. In order to evaluate the spread of the disease and risk factors, a population based seroprevalence study was done. MATERIAL AND METHODS: about 1488 serum samples collected from healthy people from different parts of the country during the year 2000 were studied by hemagglutination inhibition (HI) method. For risk factor analysis detailed information was collected by a questionnaire. RESULTS: 44 samples (2.96%) were positive. This indicates that at least 1500 infections occur in the country annually. Seropositivity did not increase with increasing age. In certain areas, seropositivity was clearly higher than the average. Other living conditions or outdoor habits correlated poorly with seropositivity. Certain groups of people such as farmers, cattle breeders, or those having a summer cottage or spending time in the nature daily had increased risk. Seropositivity was significantly linked with meningoencephalitis without laboratory confirmation for TBE in the anamnesis, and drinking of goat milk. CONCLUSION: The study suggests that TBEV is prevalent in Lithuania. The data also supports the view that an increase in the incidence has occurred in the 1990s. The correlation between seropositivity and presumed risk factors do not seem strong enough to warrant a selective vaccination policy based on risk factors. PMID- 12126719 TI - Detection of late pp67-mRNA by NASBA in peripheral blood for the diagnosis of human cytomegalovirus disease in AIDS patients. AB - OBJECTIVE: To evaluate the value of late-pp67-mRNA nucleic acid sequence-based amplification (NASBA) in comparison to DNA-PCR, blood culture and pp65 antigenemia assay for the detection of human cytomegalovirus (HCMV) disease in HIV-infected patients. METHODS: The results of pp67-mRNA NASBA, DNA-PCR, culture and pp65-antigenemia assay were compared in 402 whole blood specimens of 98 HIV infected patients with a low CD4 lymphocyte count who had not yet received highly active antiretroviral therapy (HAART). Thirty-seven samples were obtained from 30 patients with a diagnosis of HCMV disease and 365 samples from 68 patients without HCMV disease. RESULTS: The highest agreement of test results was observed between pp67-mRNA NASBA and quantitative pp65-antigenemia, with a threshold of nine antigen-positive cells/10(5) leukocytes (kappa-value 0.70, 95% CI=0.58 0.82). The sensitivity of pp67-mRNA NASBA for the diagnosis of HCMV disease (59.3%) was identical to that of the quantitative pp65-antigenemia assay, higher than that of the blood culture (48.2%) but lower than that of the DNA-PCR (77.8%). Pp67-mRNA NASBA (92.3%), quantitative pp65-antigenemia assay (92.3%) and blood culture (93.9%) were highly specific for the diagnosis of HCMV disease and as a result, had a higher positive predictive value (76.2, 76.2 and 76.5%, respectively) than the qualitative DNA-PCR (58.3%) and the qualitative pp65 antigenemia assay (47.6%). CONCLUSION: pp67-mRNA NASBA, an easy and rapid to perform assay, well-standardised by virtue of co-amplified internal system control RNA, provides a high specificity and positive predictive value for the diagnosis of HCMV disease in HIV-infected patients, comparable to that of the quantitative pp65-antigenemia assay and blood culture. PMID- 12126720 TI - Prevalence of mutations related to HIV-1 antiretroviral resistance in Brazilian patients failing HAART. AB - BACKGROUND: Current guidelines for antiretroviral (ARV) therapy recommend at least triple-drug combination, the so-called highly active antiretroviral therapy (HAART). Not all patients respond to HAART and the development of drug resistance remains one of the most serious obstacles to sustained suppression of HIV. OBJECTIVE: In an attempt to correlate the HIV therapeutic failure with reverse transcriptase (RT) and protease resistance mutations, we describe the ARV resistance profile in patients failing HAART in Brazil. We studied 267 Brazilian HIV-1 infected patients failing HAART looking for mutations in RT and protease genes. The mutation profile of the viruses infecting these individuals were deduced and correlated to laboratorial parameters. STUDY DESIGN: Two different HIV-1 genomic regions were targeted for PCR amplification, the protease (pro) and pol RT (palm finger region) genes. The mutations related to drug resistance in RT gene was analyzed using a line probe assay (LIPA(R)) and pro amino acids positions 82 and 90 were screened through RFLP using HincII restriction digestion. RESULTS: There was strong correlation between the mutation in the pro and RT genes and therapeutic failure. The main mutation found in RT gene was the M184V (48%) followed by T69D/N (47%), T215Y/F (46%), M41L (39%), and L74V (7%). In the pro gene the main mutation found was L90M (26%) followed by dual substitution in L90M and V82A (6%). All mutations profiles matched very well with the patients drug regimen. CONCLUSIONS: This study has shown that 84.7% of HIV infected subjects failing HAART for more than 3 months presented viral genomic mutations associated with drug resistance. PMID- 12126721 TI - Effect of perinatal short-course zidovudine on the clinical and virological manifestations of HIV-1 subtype E infection in infants. AB - BACKGROUND: The perinatal short-course zidovudine (ZDV) chemoprophylaxis that can reduce HIV-1 vertical transmission by 51% has been widely practiced in developing countries such as Thailand because of its simpler and less cost. OBJECTIVES: To investigate the effects of short-course regimen of oral ZDV for prophylaxis of HIV-1 subtype E vertical transmission among 'break-through' HIV-1 infected infants. STUDY DESIGN: The study analyzed clinical and virological outcomes of 80 infants, whose mothers received ZDV prophylaxis starting at 36 weeks gestation (group Z) and 37 infants whose mothers never received anti-retroviral drugs (group C), at the ages of 1-2, 4-6, and 12 months. RESULTS: Of the 12 HIV-1 infected infants, 5/7 (71.4%) from group Z and 1/5 (20%) from group C progressed to a symptomatic clinical stage by the age 4-6 months. The intersample nucleotide distance of HIV-1 pol reverse transcriptase (RT) sequences of isolates collected at age of 1-2 months from group Z was significantly higher than that from group C (3.34 and 2.92%, P=0.02). All twelve virus isolates from infected infants were non syncytium inducing (NSI) and macrophage tropic strains; and 5/6 (83.3%) viruses from symptomatic infants were also T-tropic viruses. The symptomatic infants also had significantly higher HIV-1 nucleic acid quantitation than asymptomatic infants. CONCLUSION: Our results preliminary suggested that infected infants who were perinatally exposed to ZDV may have a more rapid early disease progression with unfavorable viral manifestations than those without exposure to antiretroviral drug. PMID- 12126722 TI - Performance of an in-house genotypic antiretroviral resistance assay in patients pretreated with multiple human immunodeficiency virus type 1 protease and reverse transcriptase inhibitors. AB - An in-house genotypic antiretroviral resistance assay was evaluated by testing 32 plasma samples obtained from heavily pretreated human immunodeficiency virus type 1 (HIV-1)-infected patients failing multiple antiretroviral regimens. The same samples were also sent to Virco Laboratories for genotypic (VircoGEN) and phenotypic (Antivirogram) resistance analysis. Sequencing results obtained by in house (HG) and VircoGEN (VG) genotyping were concordant for 387 of 400 (96.75%) drug resistance mutations. Genotype-based prediction of drug susceptibility for 13 currently licensed antiretroviral compounds were in agreement in 336 (80.78%) cases, partially concordant in 73 (17.54%) cases and discordant in only seven (1.68%) cases. VG indicated 'possible resistance' twice as much as HG. When genotype interpretation was compared with the Antivirogram phenotypic data, there were 27 (6.49%) and 23 (5.52%) wrong calls by HG and by VG, respectively. Both assays were more sensitive in detecting drug resistance than drug susceptibility (94.61 vs. 65.19% for HG, 80.84 vs. 56.91% for VG) and more specific in detecting drug susceptibility than drug resistance (93.62 vs. 73.49% for HG, 93.62 vs. 80.32% for VG). Rule-based algorithms can reliably interpret genotypic data obtained from most heavily pretreated patients. However, occasional genotypic patterns may be erroneously interpreted without resistance phenotyping. PMID- 12126723 TI - The dynamics of mutations in the YMDD motif of the hepatitis B virus polymerase gene during and after lamivudine treatment as determined by reverse hybridisation. AB - We have analysed the dynamics of HBV variants related to Lamivudine resistance in 22 chronically infected patients during and after the end of Lamivudine therapy. Thirteen patients had a confirmed methionine mutation in the YMDD region of the reverse transcriptase domain determined by sequence analysis. They responded to therapy having a mean reduction of HBV DNA of 4.55 log (range 2.93-8.91). Nine patients partly responded to therapy, with a small decline of HBV DNA (mean reduction of 3.39 log, range 1.72-5.12) and no indication for an YMDD variant. Samples were re-analysed with a HBV Drug Resistance Line Probe assay (InnoLipa HBV-DR), which detected as low as 10% of a variant HBV population. With this assay, changes in the YMDD region were detected with a mean of 2 weeks (range -8 to 10) earlier than by an increase of HBV DNA levels. Increase of ALT was observed with a mean of 31 weeks (range 29-51) later than the methionine changes in the YMDD motif. Indications for a methionine into valine change could be determined in only one of the partial responders. An unexpected observation was the predominant presence of variant virus populations after end of therapy. In ten patients, we detected the wild type virus with a mean of 14 weeks after end of therapy (range 1-42 weeks). In three patients, no variant virus population could be detected at 25, 36 and 37 weeks, respectively, after cessation of treatment. This observation is important for the inclusion of the so-called naive patients in clinical trials. PMID- 12126724 TI - Guidelines for laboratory monitoring of treatment of persistent virus infections. AB - A growing number of antiviral agents are available for treatment of persistent viral infections. This has increased the requirement for virology laboratories to undertake sophisticated assays for monitoring the efficacy of treatment and identifying drug failure at an early stage. The consensus guidelines within this article address the laboratory requirements for monitoring treatment of the herpes viruses, HIV-1, Hepatitis B and Hepatitis C. PMID- 12126725 TI - Detection of HBV DNA using real time analysis. PMID- 12126726 TI - A nested primer set targeting the cytomegalovirus glycoprotein B gene. PMID- 12126727 TI - Isolation and characterization of luffacylin, a ribosome inactivating peptide with anti-fungal activity from sponge gourd (Luffa cylindrica) seeds. AB - A purification scheme involving ion exchange chromatography on DEAE-cellulose, affinity chromatography on Affi-gel blue gel, and ion exchange chromatography on CM-Sepharose and Mono S was employed to isolate a peptide with a molecular weight of 7.8kDa from sponge gourd seeds. The peptide, which was designated luffacylin, exhibited an N-terminal sequence with pronounced resemblance to that of the 6.5kDa arginine-glutamate rich polypeptide previously isolated from sponge gourd seeds. Luffacylin inhibited translation in a rabbit reticulocyte lysate system with an IC(50) of 140pM and reacted positively in the N-glycosidase assay for ribosome inactivating proteins. Luffacylin exerted anti-fungal activity against Mycosphaerella arachidicola and Fusarium oxysporum. PMID- 12126728 TI - Ascalin, a new anti-fungal peptide with human immunodeficiency virus type 1 reverse transcriptase-inhibiting activity from shallot bulbs. AB - An isolation procedure comprising ion exchange chromatography on DEAE-cellulose, affinity chromatography on Affi-gel blue gel, ion exchange chromatography on SP Sepharose and gel filtration on Superdex 75 was used to isolate an anti-fungal peptide from the bulbs of the shallot Allium ascalonicum. The peptide demonstrated a molecular weight of 9.5kDa, and possessed an N-terminal sequence YQCGQGG somewhat similar to chitinases from other Allium species which are however much larger in molecular weight. The peptide designated ascalin manifested a unique specific anti-fungal activity. It inhibited mycelial growth in the fungus Botrytis cinerea but not in the fungi Mycosphaerella arachidicola and Fusarium oxysporum. Ascalin inhibited HIV-1 reverse transcriptase with an IC(50) of 10 microM, much more potently than Allium tuberosum anti-fungal protein and other anti-fungal proteins. PMID- 12126729 TI - Identification and tissue mapping of APGWamide-related peptides in Sepia officinalis using LC-ESI-MS/MS. AB - This paper demonstrates for the first time the occurrence of tetrapeptides related to APGWamide in the mollusk cephalopod Sepia officinalis. LC-ESI-MS/MS analysis allowed the identification of the APGWamide-related peptides predicted by the two genes cloned previously in Lymnaea stagnalis and in Mytilus edulis, as well as the dipeptide GWamide released from the processing of the tetrapeptides by a dipeptidyl aminopeptidase (DPAP). TPGWamide and GWamide appeared to be exclusively located in the CNS, and the APGWamide in both the CNS and the nerve endings. The RPGWamide and the KPGWamide were not detected by LC-ESI-MS/MS suggesting they could be totally processed into GWamide. The in vitro processing of the tetrapeptides into GWamide by optic lobe extract revealed a differential processing for each, with APGWamide (44.7%)>RPGWamide(24.3%)>KPGWamide(19.3%)>TPGWamide (11.7%). The tissue mapping results, together with the processing efficiency data suggest that the GWamide is mainly produced from the M. edulis gene products RPGWamide and KPGWamide. PMID- 12126730 TI - Allatostatins of the tiger prawn, Penaeus monodon (Crustacea: Penaeidea). AB - More than 40 peptides belonging to the -Y/FXFGL-NH(2) allatostatin superfamily have been isolated and identified from the central nervous system (CNS) of the tiger prawn, Penaeus monodon (Crustacea: Penaeidea). The peptides can be arranged in seven sub-groups according to the variable post-tyrosyl residue represented by Ala, Gly, Ser, Thr, Asn, Asp, and Glu. Two of the residues (Thr and Glu) have not been observed in this position previously in either insects or crustaceans. Also reported for the first time for allatostatins, two of the peptides are N terminally blocked by a pyroglutamic acid residue. The yields of certain peptides with similar amino acid sequences to each other were, in some instances, very different. As an example, the yield of ANQYTFGL-NH(2) was 2pmol, compared with ASQYTFGL-NH(2), with a yield of 156 pmol. There are several possibilities to account for this. If, as in all species so far investigated, there is a single allatostatin gene in P. monodon, then it would appear that different sub populations have contributed mutant forms of particular peptides to the extract. Another, less likely possibility is that this species has more than one allatostatin gene, producing a variable array of peptides albeit in different molar ratios. Several peptides were present apparently as a result of the loss of one or more residues at the N-terminus of a larger form, either due to N-terminal degradation or specific post-translational processing. The number of peptides identified exceeds that for any other insect or crustacean species previously investigated. None is identical to any of the 60-70 insect allatostatins so far identified, and only three are common to other crustaceans. Immunohistochemical study of the CNS of P. monodon, with the same antisera as used to monitor the purification, confirms the widespread nature and complexity of allatostatinergic neural pathways in arthropods. Thus, all neuromeres of the brain, and all except one of the ventral cord ganglia, possess allatostatin neurons and extensive areas of allatostatin-innervated neuropile. In addition to the cytological evidence that the allatostatins act as neurotransmitters, associated with tissues as varied as eyes and legs, their presence in neurohemal areas such as the sinus gland and the perineural sheath of the thoracic ganglia suggests a neuroendocrine function. As well as posing a challenge to physiologists assigning specific functions to the allatostatins, their extensive intra-species multiplicity, linked to their inter-species variability, also presents a complex problem to geneticists and evolutionists. PMID- 12126731 TI - Corticotropin-releasing hormone (CRH) in the teleost fish Oreochromis mossambicus (tilapia): in vitro release and brain distribution determined by a novel radioimmunoassay. AB - The quantitative distribution of corticotropin-releasing hormone (CRH) in the brain and pituitary of the fish Oreochromis mossambicus (tilapia) was studied following the validation of a radioimmunoassay. Compared to the pituitary content, the brain contained 20 times more CRH. Eighty percent of the total brain content was located outside the hypothalamus, particularly in the telencephalon. Substantial amounts of CRH were also present in other regions devoid of hypophysiotropic neurons, such as the vagal lobe and optic tectum. Telencephalic and pituitary CRH co-eluted with the tilapia CRH(1-41)standard on reverse phase HPLC. In vitro CRH release by the telencephalon amounted to 5% of its content per hour, whereas release from the pituitary was negligible. We conclude that CRH in the brain of tilapia regulates pituitary and non-pituitary related functions, probably as a neurotransmitter or neuromodulator. PMID- 12126732 TI - Diet-induced changes in hypothalamic pro-opio-melanocortin mRNA in the rat hypothalamus. AB - Hypothalamic mRNA and peptide levels of pro-opio-melanocortin (POMC) and other neuropeptides were studied in rats that either develop obesity (diet-induced obese, DIO), when fed a palatable and hypercaloric diet (cafeteria diet, caf) or do not develop obesity (diet resistant, DR), when fed the same diet. cafDIO rats showed a significant increase in POMC, but not in melanin concentrating hormone, mRNA levels as determined by semiquantitative in situ hybridization. cafDR and cafDIO rats showed no change in POMC-derived peptide levels, whereas neuropeptide Y immunoreactivity was significantly increased in cafDR rats. POMC mRNA levels were also studied in high-fat diet-fed rats but no significant change was observed. Altered hypothalamic transmission by POMC-derived peptides may contribute to the susceptibility of cafDIO rats to the weight promoting action of caf diet. PMID- 12126733 TI - Sustained orexigenic effect of Agouti related protein may be not mediated by the melanocortin 4 receptor. AB - Intracerebroventricular (ICV) injection of Agouti related protein (AgRP), an endogenous melanocortin 3 and 4 receptor (MC3/4-R) antagonist, produces a prolonged increase in food intake. To clarify the roles of the MC3-R and MC4-R in AgRP-induced hyperphagia, the feeding effect of AgRP (83-132) was compared with that of the selective MC4-R antagonist, JKC-363 (cyclic [Mpr11, D-Nal14, Cys18, Asp22-NH2]-beta-MSH11-22). Single ICV administration of AgRP (83-132) increased food intake for 48 h whilst ICV JKC-363 increased food intake for 8h. An increase in body weight at 24 and 48 h was observed following AgRP (83-132) but not JKC 363 treatment. These data suggest that the sustained orexigenic action of AgRP (83-132) may not be through MC4-R antagonism. PMID- 12126735 TI - Attenuation of hypercholesterolemia and hyperglycemia in ob/ob mice by NPY Y2 receptor ablation. AB - Neuropeptide Y (NPY) is a 36 amino acid peptide well known for its role in regulating food intake and energy homeostasis. It has previously been shown that the NPY Y2 receptor is required for a full biological response to leptin in the central nervous system. We have examined the impact of this receptor on plasma levels of lipid and cholesterol in wild type and obese (ob/ob) mice. The results show that an absence of Y2 in female mice has no effect on cholesterol level in normal lean mice but profoundly decreases serum cholesterol and glucose levels in ob/ob mice. We conclude that NPY, interacting with the Y2 receptor, participates in cholesterol and glucose homeostasis of obese mice. PMID- 12126734 TI - Structure-activity relationship and signal transduction of gamma-MSH peptides in GH3 cells: further evidence for a new melanocortin receptor. AB - The structure-activity relationship and signal transduction properties of the pro opiomelanocortin (POMC)-derived gamma-MSH peptides in the GH3 cell line was compared with that described for the known melanocortin receptors (MCRs). Single alanine replacements showed that, unlike the classical MCRs, the His(5)-Phe(6) Arg(7)-Trp(8) sequence in gamma2-MSH is not a core sequence for activating the gamma-MSH receptor in GH3 cells, whereas Met(3) is essential. gamma2-MSH increased binding of [35S]GTPgammaS to membrane preparations of GH3 cells. Blockade of protein kinase A abolished the [Ca(2+)](i) responses to gamma3-MSH, and low nanomolar doses of gamma3-MSH increased intracellular cAMP levels, which could be blocked by pertussis toxin (PTX). We conclude that the putative novel gamma-MSH receptor in GH3 cells is a GPCR, but with structure-activity and signal transduction features different from those of the classical MCRs. PMID- 12126736 TI - Immunotoxic catecholamine lesions attenuate 2DG-induced increase of AGRP mRNA. AB - Injections of the immunotoxin, saporin conjugated to anti-dopamine-beta hydroxylase (DSAP), into the hypothalamic paraventricular nucleus (PVH) selectively destroy norepinephrine (NE) and epinephrine (E) terminals in the medial hypothalamus and abolish glucoprivic feeding. We utilized PVH DSAP injections to examine the role of NE/E neurons in the previously reported 2-deoxy D-glucose (2DG)-induced increases in mRNA levels for the orexigenic peptides, AGRP and NPY. Northern blot analysis revealed that DSAP lesions elevated basal but blocked 2DG-induced increases in AGRP mRNA levels. Changes in NPY mRNA were not detectable. AGRP neurons may contribute to circuitry activated by NE/E neurons for elicitation of glucoregulatory responses. PMID- 12126737 TI - Pharmacological characterization of beta-endorphin- and dynorphin A(1-17)-induced feeding using G-protein alpha-subunit antisense probes in rats. AB - Antisense (AS) oligodeoxynucleotides targeting G-protein alpha-subunits distinguish feeding responses of morphine and its metabolite, as well as nocturnal and deprivation-induced feeding. The present study examined whether feeding elicited by beta-endorphin (betaEND) or dynorphin A(1-17) was altered by ventricularly-applied G(i)alpha(1), G(i)alpha(2), G(i)alpha(3), G(s)alpha, G(o)alpha, G(q)alpha or G(x/z)alpha AS probes, or a nonsense (NS) control. The betaEND-induced feeding was reduced by the G(i)alpha(1) and G(x/z)alpha AS probes, and increased by G(i)alpha(2) or G(i)alpha(3) AS treatment. Dynorphin induced feeding was attenuated by G(i)alpha(1) and G(o)alpha AS treatment. Yet, G(s)alpha or G(q)alpha AS and NS treatments failed to alter opioid agonist induced feeding. These data provide initial characterization of potential effector signaling pathways mediating betaEND and dynorphin-induced feeding. PMID- 12126738 TI - Peptide nucleic acids targeted to the mouse proNPFF(A) reveal an endogenous opioid tonus. AB - Pharmacological studies have implicated the anti-opioid neuropeptide FF (NPFF) in the modulation of pain transmission. Since its physiological role has not yet been fully elucidated, the present study examined whether antisense peptide nucleic acid (PNA) complementary to the NPFF precursor (proNPFF(A)) modified pain sensitivity. Mice received three intraperitoneal (i.p.) injections (10mg/kg) of antisense PNA (As-proNPFF(A)) over a period of 24h. As-proNPFF(A) treatment significantly increased the basal tail withdrawal latency in the tail-flick test. This analgesia persisted during 2 days and was completely reversed by naloxone. Thus, antisense PNAs, by decreasing anti-opioid effects, revealed a basal endogenous opioid activity. Our results evidence a physiological interplay between NPFF and opioid systems and further support the use of PNA as effective antisense agents, for studying gene function in vivo. PMID- 12126739 TI - Synthetic peptide SLTCLVKGFY competes with beta-endorphin for naloxone insensitive binding sites on rat brain membranes. AB - The synthetic decapeptide Ser-Leu-Thr-Cys-Leu-Val-Lys-Gly-Phe-Tyr (termed immunorphin) corresponding to the sequence 364-373 of the CH3 domain of human immunoglobulin G heavy chain and its synthetic fragment VKGFY were found to compete with 125I-labeled beta-endorphin for high-affinity naloxone-insensitive binding sites on membranes isolated from the rat brain cortex (K(i)=1.18+/-0.09 and 1.58+/-0.11 nM, respectively). The binding specificity study revealed that these binding sites were insensitive not only to naloxone but to [Met(5)]enkephalin and [Leu(5)]enkephalin as well. The K(d) values characterizing the specific binding of 125I-labeled immunorphin and its fragment Val-Lys-Gly-Phe Tyr to these binding sites were determined to be 2.93+/-0.27 nM and 3.17+/-0.29 nM, respectively. PMID- 12126741 TI - Expression of growth hormone-releasing hormone (GHRH) and splice variants of GHRH receptors in human experimental prostate cancers. AB - The expression of mRNA for GHRH and splice variants (SVs) of GHRH receptors in LNCaP, MDA-PCa-2b and PC-3 human prostate cancers grown in nude mice was investigated by RT-PCR. The expression of mRNA for GHRH was detected in LNCaP and PC-3, but not in MDA-PCa-2b prostatic carcinoma. RT-PCR analyses of mRNA isolated from LNCaP, MDA-PCa-2b and PC-3 cancers, revealed the presence of 720 and 566 bp products, corresponding to SV(1) and SV(2) isoforms of GHRH receptors. In PC-3 tumor membranes a radiolabeled GHRH antagonist [125I]-JV-1-42 was bound to one class of high-affinity binding sites (K(d)=1.81+/-0.47 nM) and maximum binding capacity of 332.7+/-27.8 fmol/mg membrane protein. The in vivo uptake of [125I] JV-1-42 was observed in all xenografts of human prostate cancer, the tracer accumulation being the highest in PC-3 tumors. These results indicate that GHRH and SVs of its receptors, different from those found in the pituitary, are present in experimental human prostate cancers and may form a local mitogenic loop. The antiproliferative effects of GHRH antagonists on growth of prostate cancer could be exerted in part by an interference with this local GHRH system. PMID- 12126740 TI - Oxytocin stimulates proliferation of human osteoblast-like cells. AB - Oxytocin receptors have recently been demonstrated in human osteoblast-like (hOB) cells. In this study, oxytocin 100-1000 pmol/l increased cell proliferation of primary cultures of hOB cells, measured by [3H]thymidine incorporation, (P<0.01). In human osteosarcoma cell-line (SaOS-2), oxytocin 100 pmol/l increased cell proliferation (measured by [3H]thymidine incorporation and a commercially available kit) and protein synthesis ([3H]proline incorporation) (P<0.05). The increase in cell proliferation was abolished when SaOS-2 cells were incubated with an oxytocin antagonist and oxytocin. Oxytocin 100 pmol/l decreased interleukin-6 (IL-6) production of the hOB cells (23.4+/-1.96 versus 33.4+/-2.65 pg/well; P<0.001). These findings indicate that oxytocin may affect bone metabolism in humans. PMID- 12126742 TI - Expression of prolactin-releasing peptide and its receptor in the human adrenal glands and tumor tissues of adrenocortical tumors, pheochromocytomas and neuroblastomas. AB - Adrenal tumors, such as pheochromocytomas, are known to express various peptides and their receptors. Prolactin-releasing peptide (PrRP) is a novel neuropeptide isolated from bovine hypothalamic tissues. In the present study, expression of PrRP receptor was studied in the human brain, pituitaries, adrenal glands and tumor tissues of adrenocortical tumors, pheochromocytomas, a ganglioneuroblastoma and neuroblastomas by reverse transcriptase polymerase chain reaction (RT-PCR) and Northern blot analysis. The presence of immunoreactive-PrRP in the adrenal glands and in these tumor tissues was studied by radioimmunoassay. Human brain tissues and pituitaries were obtained at autopsy. Normal portions of adrenal glands and tumor tissues were obtained at surgery. RT-PCR analysis showed expression of PrRP receptor in the human brain, pituitaries, normal portions of adrenal glands and various tumor tissues. Northern blot analysis showed high expression of PrRP receptor only in tumor tissues of pheochromocytomas, indicating that PrRP receptor expression is high in pheochromocytomas. Immunoreactive-PrRP was detected in normal portions of adrenal glands (0.162+/ 0.024 pmol/g wet weight, n=4, mean+/-S.E.M.), four out of six cases of pheochromocytomas (0.050-7.9 pmol/g wet weight), one neuroblastoma and some adrenocortical tumors. The present study has shown that PrRP receptor mRNA was widely expressed in the brain tissues, pituitaries, adrenal glands and various tumors. The high expression of PrRP receptor in pheochromocytomas suggests potential pathophysiological roles of PrRP in these tumors. PMID- 12126743 TI - Effects of different peptide fragments derived from proadrenomedullin on gene expression of adrenomedullin gene. AB - Primary culture of vascular smooth muscle cells (VSMC) from rat aorta was used for the study of the effect of different peptides derived from proadrenomedullin on the expression of adrenomedullin (ADM) gene. ADM and preproADM(22-41) (PAMP) secreted by VSMC were measured by radioimmunoassay, and ADM mRNA in VSMC was determined by quantitative RT-PCR. After the incubation of VSMC in 10(-7)M ADM for 24h, PAMP in the medium and ADM mRNA in the VSMC were decreased by 34 and 41.3%, respectively, and cAMP concentration in the VSMC was increased by 385%. After the incubation of VSMC in 10(-7)M PAMP for 24h, ADM in the medium and ADM mRNA in the VSMC were decreased by 12.2 and 39.1%, respectively, and cAMP concentration in the VSMC was increased by 67%. The decreased ADM mRNA in VSMC induced by the ADM and PAMP treatment was completely reversed by the pre treatment of the cells in 10(-7)M protein kinase inhibitor for 30 min. After the incubation of VSMC in 10(-7)M preproADM(153-185) (ADT) for 24h, however, ADM in the medium and ADM mRNA in the VSMC were increased by 21 and 35.2%. The increased ADM mRNA in VSMC induced by the ADT treatment was partially blocked by the co incubation in ADM and ADT, and was totally blocked by the co-incubation in PAMP+ADM and ADT, but was not blocked by the co-incubation in PAMP and ADT. Our results suggest that the four peptides derived from proadrenomedullin may have different effects, possibly through a cAMP-dependent pathway, on the expression of ADM gene. PMID- 12126744 TI - Gastroprotective effect of adrenomedullin administered subcutaneously in the rat. AB - Subcutaneous injections of adrenomedullin prevented reserpine-induced gastric mucosal damage in a dose-dependent manner (1-1000 ng/kg), but did not interfere with the lesions produced by ethanol administration. In pylorus-ligated rats adrenomedullin significantly reduced gastric volume, total and free acid output as well as ulcer formation. The gastroprotective activity of adrenomedullin was not present in rats pretreated with cysteamine. These results suggest that adrenomedullin exerts its antiulcer effect, when it is administered subcutaneously (s.c.), probably by a mechanism which involves somatostatin related transmission. PMID- 12126746 TI - The mechanism of the angiotensin-converting enzyme inhibitor quinapril is not related to bradykinin level in heart tissue. AB - In order to examine the effect of the angiotensin-converting enzyme inhibitor (ACEi) quinapril, we performed a sensitive and specific radioimmunoassay (RIA) to quantify bradykinin, BK-(1-9), in heart and kidney tissues. The BK-(1-9) level was unaffected in the heart of sham and water-deprived rats treated for 2h with quinapril (10mg/kg), but was significantly higher in the kidneys in the two groups. In these conditions, circulating and tissue angiotensin II (Ang II) levels were significantly decreased by quinapril. Moreover, our results indicated that acute treatment with this dose of quinapril induced kinin-mediated effects which were not related to its action on bradykinin degradation in rat hearts. PMID- 12126745 TI - The tachykinin NK-2 receptor antagonist SR48968 does not block noncholinergic contractions in unstable human bladder. AB - Concentration-response curves to acetylcholine, and responses to electrical field stimulation (EFS) were compared in detrusor muscle strips, from control patients and those with idiopathic detrusor instability (IDI). Responses were similar in both groups. However, atropine abolished responses to EFS in 80% of control but only 33% of IDI patients (P>0.05), with the residual atropine-resistant response in most IDI patients abolished by tetrodotoxin. The post-atropine residual response was unaffected by the tachykinin NK-2 receptor antagonist SR48968. Despite the known existence of NK-2 receptors in the human detrusor, there was no evidence for tachykinin contribution to EFS-induced contractions. PMID- 12126747 TI - Metabolism of angiotensin I in the coronary circulation of normal and diabetic rats. AB - Formation of metabolites from angiotensin I that passed the coronary vessels in the isolated working rat hearts of normal and streptozotocin-induced diabetes was evaluated. HPLC analysis showed that the levels of angiotensin II and angiotensin 1-7 were unaltered in the diabetic hearts, but the perfusates of the diabetic hearts contained smaller amounts of angiotensin 1-9. It is not clear why the perfusates of diabetic hearts contain less amount of angiotensin 1-9. It is possible that the peptide is metabolized faster or greater internalization takes place in the diabetic heart. The amount of angiotensin II in the perfusates of normal hearts was 5.8 times greater at the perfusion rate of 2 than at 10 ml/min/g wet heart weight. At such conditions, the amount of angiotensin 1-9 and angiotensin 1-7 in the perfusates were increased 2.4 and 1.5 times, respectively. A higher amount of angiotensin II during myocardial hypoperfusion may lead to constriction of the coronary vessels. As a result, myocardial damage may be facilitated. PMID- 12126748 TI - Sexually dimorphic expression of prepro-orexin mRNA in the rat hypothalamus. AB - The neuropeptides orexin A and B are expressed in the lateral hypothalamic area and are involved in the regulation of energy homeostasis and arousal. Recent results showed gender differences in the expression of orexin receptor subtypes in rats. In the present study, we analyzed the mRNA expression of prepro-orexin (PPO) in the hypothalamus of male and female rats using quantitative real-time PCR. We found significantly higher levels of PPO mRNA in the hypothalamus of female rats compared to male rats. Our study indicates a sex-dependent regulation of hypothalamic PPO expression and suggests gender-specific functions of orexins. PMID- 12126750 TI - Characterization of a growth hormone-releasing hormone binding site in the rat renal medulla. AB - Receptor binding analysis was performed in the renal medulla from 2-month-old rats, an extrapituitary tissue containing the highest level of GHRH receptor mRNA. At 4 degrees C, in the presence of a cocktail of protease inhibitors, binding of [125I-Tyr(10)]hGHRH (1-44)NH(2) to medullary homogenates was specific, time-dependent, reversible and saturable (K(d): 28 nM; B(max): 30 fmol/mgprot.). In these experimental conditions, no change of binding parameters could be detected in the course of aging. The structure-affinity profile was different in the two tissues and chemical cross-linking revealed the presence of 65-, 55- and 38-kDa 125I-GHRH-labeled complexes in the renal medulla compared to 65-, 47- and 28-kDa radioactive complexes in the anterior pituitary. It is suggested that GHRH binding sites, and possibly the receptor, may be different in the two tissues. PMID- 12126749 TI - The PAR-1-activating peptide attenuates carrageenan-induced hyperalgesia in rats. AB - We examined if thrombin or a receptor-activating peptide for protease-activated receptor-1 (PAR-1), a thrombin receptor, could modulate nociception at peripheral levels. Intraplantar administration of PAR-1 activators, thrombin or TFLLR-NH(2), but not its inactive control FTLLR-NH(2) or a PAR-2 activator SLIGRL-NH(2), significantly attenuated the hyperalgesia in rats treated with carrageenan, although they had no effect on nociception in naive rats. The thrombin-PAR-1 system might thus act to attenuate nociception during inflammatory hyperalgesia. PMID- 12126751 TI - Introduction--serial review: reactive oxygen and nitrogen in inflammation(1,2). PMID- 12126752 TI - Mediators and regulation of neutrophil accumulation in inflammatory responses in lung: insights from the IgG immune complex model. AB - Neutrophil trafficking in lung involves transendothelial migration, migration in tissue interstitium, and transepithelial migration. In a rat model of IgG immune complex-induced lung injury, it was demonstrated that neutrophil emigration involves regulatory mechanisms including complement activation, cytokine regulation, chemokine production, activation of adhesion molecules, and their respective counter receptors. The process is presumably initiated and modulated by the production of early response cytokines and chemokines from lung cells, especially from alveolar macrophages. TNF-alpha and IL-1 up-regulate intracellular adhesion molecule-1 (ICAM-1) and E-selectin, setting the stage for neutrophil migration through endothelium. The CXC chemokines, such as macrophage inflammatory protein (MIP)-2 and cytokine-inducible neutrophil chemoattractant (CINC), constitute chemokine gradient to orchestrate neutrophil migration in lung. Complement activation induced by IgG immune complex deposition is another important event leading to neutrophil accumulation in lung. Complement activation product C5a not only plays an important role in chemoattracting neutrophils into lung, but regulates adhesion molecules, chemokines, and cytokines expression. In addition, oxidative stress may regulate neutrophil accumulation in lung by modulation of adhesion molecule activation and chemokine production. In this review, we focus on the current knowledge of the mechanisms leading to accumulation of neutrophils during acute lung injury. PMID- 12126753 TI - Role of reactive metabolites of oxygen and nitrogen in inflammatory bowel disease. AB - The inflammatory bowel diseases (IBD; Crohn's disease, ulcerative colitis) are a collection of chronic idiopathic inflammatory disorders of the intestine and/or colon. Although the pathophysiology of IBD is not known with certainty, a growing body of experimental and clinical data suggests that chronic gut inflammation may result from a dysregulated immune response to normal bacterial antigens. This uncontrolled immune system activation results in the sustained overproduction of reactive metabolites of oxygen and nitrogen. It is thought that some of the intestinal and/or colonic injury and dysfunction observed in IBD is due to elaboration of these reactive species. This review summarizes the current state of-knowledge of the role of reactive oxygen species and nitric oxide in the pathophysiology of IBD. PMID- 12126754 TI - Molecular and cellular mechanisms involved in Helicobacter pylori-induced inflammation and oxidative stress. AB - Helicobacter pylori (H. pylori)-infection leads to different clinical and pathological outcomes in humans, including chronic gastritis, peptic ulcer disease, and gastric neoplasia. The key determinants of these outcomes are the severity and distribution of the H. pylori-induced inflammation. Antral-type gastritis is associated with excessive acid secretion and a high risk of duodenal ulcer. In contrast, gastritis that involves the acid-secreting corpus region leads to hypochlorhydria, progressive gastric atrophy, and an increased risk of gastric cancer. The key pathophysiological event in H. pylori infection is initiation and continuance of an inflammatory response. Bacteria or their products trigger this inflammatory process and the main mediators are cytokines. Identification of both host- and bacterial-factors that mediate is an intense area of interest in current researches. Recent data indicates that the cag pathogenicity island plays a crucial role in H. pylori-induced gastric inflammation via the activation of gene transcription. It has been demonstrated that oxidative and nitrosative stress associated with inflammation plays an important role in gastric carcinogenesis as a mediator of carcinogenic compound formation, DNA damage, and cell proliferation. Genetic information regulating such stress would be one of the host factors determining the outcome- particularly when the outcome is gastric cancer--of H. pylori infection, and the compound that attenuates such stress may be a candidate for use in chemoprevention. This review highlights recent advances in understanding of the mechanisms underlying chronic inflammation following infection with H. pylori. PMID- 12126755 TI - Superoxide dismutase multigene family: a comparison of the CuZn-SOD (SOD1), Mn SOD (SOD2), and EC-SOD (SOD3) gene structures, evolution, and expression. AB - Superoxide dismutases are an ubiquitous family of enzymes that function to efficiently catalyze the dismutation of superoxide anions. Three unique and highly compartmentalized mammalian superoxide dismutases have been biochemically and molecularly characterized to date. SOD1, or CuZn-SOD (EC 1.15.1.1), was the first enzyme to be characterized and is a copper and zinc-containing homodimer that is found almost exclusively in intracellular cytoplasmic spaces. SOD2, or Mn SOD (EC 1.15.1.1), exists as a tetramer and is initially synthesized containing a leader peptide, which targets this manganese-containing enzyme exclusively to the mitochondrial spaces. SOD3, or EC-SOD (EC 1.15.1.1), is the most recently characterized SOD, exists as a copper and zinc-containing tetramer, and is synthesized containing a signal peptide that directs this enzyme exclusively to extracellular spaces. What role(s) these SODs play in both normal and disease states is only slowly beginning to be understood. A molecular understanding of each of these genes has proven useful toward the deciphering of their biological roles. For example, a variety of single amino acid mutations in SOD1 have been linked to familial amyotrophic lateral sclerosis. Knocking out the SOD2 gene in mice results in a lethal cardiomyopathy. A single amino acid mutation in human SOD3 is associated with 10 to 30-fold increases in serum SOD3 levels. As more information is obtained, further insights will be gained. PMID- 12126756 TI - Effect of Kupffer cell inactivation on ethanol-induced protein adducts in the liver. AB - Tissue deposition of protein adducts derived from ethanol metabolism and lipid peroxidation, has been suggested to play a role in the initiation of alcoholic liver disease. The mechanisms modulating adduct formation have, however, remained unclear. We used immunohistochemical methods to examine acetaldehyde (AA) and malondialdehyde (MDA) adducts and cytochrome P4502E1 and P4503A2 expression in rats after administration of (i) an ethanol-diet (n = 6), (ii) ethanol-diet plus gadolinium chloride (GdCl(3)), a selective Kupffer cell toxicant (n = 7), or (iii) control diet (n = 6). A 4 week ethanol treatment resulted in liver steatosis, necrosis, and inflammation and deposition of protein adducts with both AA and MDA, which colocalized with areas of fatty change. The intensities (mean +/- SD) of the immunohistochemical reactions for both AA (2.50 +/- 1.23) and MDA (3.00 +/- 1.10) adducts were significantly higher in the ethanol-fed animals than in the controls (0.083 +/- 0.20) (0.16 +/- 0.25) (p <.001). GdCl(3) prevented adduct accumulation, the mean immunohistochemistry scores being 0.86 +/- 1.07 for AA and 1.64 +/- 0.63 for MDA, the former showing a more striking reduction (p <.01). The hepatic cytochrome enzymes were not different in the ethanol-fed groups with or without GdCl(3). The data indicates that Kupffer cells are involved in the generation of protein adducts with both acetaldehyde and ethanol induced lipid peroxidation products in alcoholic liver disease. PMID- 12126757 TI - A novel iron chelator that does not induce HIF-1 activity. AB - Human skin cells (FEK-4) have been shown to undergo an immediate and transient release of low molecular mass iron (LMrFe) when subjected to UVA (320-380 nm) irradiation and this iron may act as a pro-oxidant and increase tissue injury. In order to decrease this transient release of LMrFe, cells were treated with the iron chelators desferrioxamine (DFO) and salicylaldehyde isonicotinoyl hydrazone (SIH). However, although the iron pool decreased, an increase in the DNA binding activity of the hypoxia inducible factor-1 (HIF-1) was observed when DFO and SIH were administered to normal growing FEK-4 cells. The induction of HIF-1 activates the expression of several genes associated with hypoxia and iron homeostasis. HIF 1 induction has also been associated with protection against certain forms of oxidative stress. Therefore, it is difficult to use a conventional HIF-1 activating iron chelator (such as DFO) for mechanistic studies of protection against iron-mediated oxidative stress since any protection observed could be a consequence of either the chelation of LMrFe or the induction of protective genes associated with the hypoxic response. In order to observe the effect of iron chelation on cell function without the induction of hypoxia responsive genes, cells were treated with the novel iron chelator N-(2-hydroxybenzyl)-L-serine (HBSer). Although this compound is an effective iron chelator under the conditions employed in this experiment, it does have a lower iron-binding constant than either DFO or SIH. This may be the major determinant of the observation that the compound does not induce HIF-1 binding or activate HIF-1 responsive transcriptional promoters. PMID- 12126758 TI - Immunological identification of the heart myoglobin radical formed by hydrogen peroxide. AB - This study reports the detection of protein free radicals using the specific free radical reactivity of nitrone spin traps in conjunction with nitrone-antibody specificity. Polyclonal antibodies were developed that bind to protein adducts of the nitrone spin-trap 5,5-dimethyl-1-pyrroline N-oxide (DMPO). The antibodies were used to detect DMPO protein adducts produced on horse myoglobin resulting from self-peroxidation. Western blot analysis demonstrates that myoglobin forms the predominant radical-derived nitrone adduct in rat heart supernatant. PMID- 12126759 TI - Multidrug resistance-associated protein2 (MRP2) plays an important role in the biliary excretion of glutathione conjugates of 4-hydroxynonenal. AB - Glutathione (GSH) conjugates of 4-hydroxy-trans-2,3-nonenal (HNE) are the final products of lipid peroxidation. In the present study, the role of multidrug resistance-associated protein 2 (MRP2) in biliary excretion of GSH conjugates of HNE (HNE-SG) was studied in vitro by using Madin-Darby canine kidney II (MDCK II) cells expressing human MRP2 and in vivo using a mutant rat strain whose MRP2 expression is defective (Eisai-hyperbilirubinemic rats [EHBR]). A high performance liquid chromatography method was developed to assay HNE-SG conjugates. Four diastereomeric HNE-SG conjugates could be separated with this method. Three of four HNE-SG conjugates were detectable after incubation of the cell monolayers with HNE. Expression of human MRP2 resulted in a 10-fold increase in HNE-SG conjugates excretion across the apical membrane of MDCK II cells. The four HNE-SG conjugates appeared swiftly in bile from Sprague Dawley rats after intravenous administration of HNE, whereas no detectable HNE-SG conjugates were observed in the bile of EHBR. These results demonstrate the role of MRP2 in the biliary excretion of HNE-SG conjugates. PMID- 12126760 TI - Inhibition of airway inflammation and hyperreactivity by an antioxidant mimetic. AB - A catalytic antioxidant, AEOL 10113, was used in a murine model of asthma to test the hypothesis that oxidants contribute to the pathogenesis of asthma. Balb/c mice were immunized and challenged with ovalbumin. AEOL 10113 was administered to the mice by intratracheal instillation during ovalbumin challenges. Enhanced pause as an indicator of airway reactivity and differential cell count of lavage cells as an indicator of airway inflammation were assessed. Lung expressions of the adhesion molecules VCAM-1 and ICAM-1 were measured. We found that treatment of ovalbumin-challenged mice with AEOL 10113 drastically reduced the severity of airway inflammation as evidenced by the reduced numbers of eosinophils, neutrophils, and lymphocytes found in bronchoalveolar lavage fluid. Inhibition of ovalbumin-induced airway inflammation is associated with inhibited expression of VCAM-1, which is a key adhesion molecule responsible for the recruitment of inflammatory cells into the lungs of ovalbumin-challenged mice. In addition, treatment with AEOL 10113 reduced the magnitude of ovalbumin-induced airway hyperreactivity to methacholine. These results suggest that oxidative stress is an important factor in the pathogenesis of asthma and that a synthetic catalytic antioxidant could be effective in the treatment of asthma. PMID- 12126761 TI - Inhibition of cancer growth by resveratrol is related to its low bioavailability. AB - The relationship between resveratrol (RES) bioavalability and its effect on tumor growth was investigated. Tissue levels of RES were studied after i.v. and oral administration of trans-resveratrol (t-RES) to rabbits, rats, and mice. Half-life of RES in plasma, after i.v. administration of 20 mg t-RES/kg b.wt., was very short (e.g., 14.4 min in rabbits). The highest concentration of RES in plasma, either after i.v. or oral administration (e.g., 2.6 +/- 1.0 microM in mice 2.5 min after receiving 20 mg t-RES/kg orally), was reached within the first 5 min in all animals studied. Extravascular levels (brain, lung, liver, and kidney) of RES, which paralleled those in plasma, were always < 1 nmol/g fresh tissue. RES measured in plasma or tissues was in the trans form (at least 99%). Hepatocytes metabolized t-RES in a dose-dependent fashion (e.g., 43 nmol of t-RES/g x min in the presence of 20 microM tRES), which means that the liver can remove circulating RES very rapidly. In vitro B16 melanoma (B16M) cell proliferation and generation of reactive oxygen species (ROS) was inhibited by t-RES in a concentration-dependent fashion (100% inhibition of tumor growth was found in the presence of 5 microM t-RES). Addition of 10 microM H(2)O(2) to B16M cells, cultured in the presence of 5 microM t-RES, reactivated cell growth. Oral administration of t-RES (20 mg/kg twice per day; or included in the drinking water at 23 mg/l) did not inhibit growth of B16M inoculated into the footpad of mice (solid growth). However, oral administration of t-RES (as above) decreased hepatic metastatic invasion of B16M cells inoculated intrasplenically. The antimetastatic mechanism involves a t-RES (1 microM)-induced inhibition of vascular adhesion molecule 1 (VCAM-1) expression in the hepatic sinusoidal endothelium (HSE), which consequently decreased in vitro B16M cell adhesion to the endothelium via very late activation antigen 4 (VLA-4). PMID- 12126762 TI - The role of cysteine residues in the oxidation of ferritin. AB - We have shown that ferritin is oxidized during iron loading using its own ferroxidase activity and that this oxidation results in its aggregation (Welch et al., Free Radic. Biol. Med. 31:999-1006; 2001). In this study we determined the role of cysteine residues in the oxidation of ferritin. Loading iron into recombinant human ferritin by its own ferroxidase activity decreased its conjugation by a cysteine specific spin label, indicating that cysteine residues were altered during iron loading. Using LC/MS, we demonstrated that tryptic peptides of ferritin that contained cysteine residues were susceptible to modification as a result of iron loading. To assess the role of cysteine residues in the oxidation of ferritin, we used site-directed mutagenesis to engineer variants of human ferritin H chain homomers where the cysteines were substituted with other amino acids. The cysteine at position 90, which is located at the end of the BC-loop, appeared to be critical for the formation of ferritin aggregates during iron loading. We also provide evidence that dityrosine moieties are formed during iron loading into ferritin by its own ferroxidase activity and that the dityrosine formation is dependent upon the oxidation of cysteine residues, especially cysteine 90. In conclusion, cysteine residues play an integral role in the oxidation of ferritin and are essential for the formation of ferritin aggregates. PMID- 12126763 TI - Renal damage mediated by oxidative stress: a hypothesis of protective effects of red wine. AB - Over the last decade, oxidative stress has been implicated in the pathogenesis of a wide variety of seemingly unrelated renal diseases. Epidemiological studies have documented an association of moderate wine consumption with a decreased risk of cardiovascular and neurological diseases; however, similar studies in the kidney are still lacking. The kidney is an organ highly vulnerable to damage caused by reactive oxygen species (ROS), likely due to the abundance of polyunsaturated fatty acids in the composition of renal lipids. ROS are involved in the pathogenic mechanism of conditions such as glomerulosclerosis and tubulointerstitial fibrosis. The health benefits of moderate consumption of red wine can be partly attributed to its antioxidant properties. Indeed, the kidney antioxidant defense system is enhanced after chronic exposure to moderate amounts of wine, a response arising from the combined effects of ethanol and the nonalcoholic components, mainly polyphenols. Polyphenols behave as potent ROS scavengers and metal chelators; ethanol, in turn, modulates the activity of antioxidant enzymes. Therefore, a hypothesis that red wine causes a decreased vulnerability of the kidney to the oxidative challenges could be proposed. This view is partly supported by direct evidences indicating that wine and antioxidants isolated from red wine, as well as other antioxidants, significantly attenuate or prevent the oxidative damage to the kidney. The present hypothesis paper provides a collective body of evidence suggesting a protective role of moderate wine consumption against the production and progression of renal diseases, based on the existing concepts on the pathophysiology of kidney injury mediated by oxidative stress. PMID- 12126764 TI - Beraprost sodium, prostacyclin analogue, attenuates glomerular hyperfiltration and glomerular macrophage infiltration by modulating ecNOS expression in diabetic rats. AB - Stable prostacyclin analogue, beraprost sodium (BPS) has recently been reported to attenuate glomerular hyperfiltration in diabetic rats, however, the mechanism has been still unknown. We previously reported that overexpression of endothelial cell nitric oxide synthase (ecNOS) in afferent arterioles and glomeruli induce inappropriate dilatation of afferent arterioles and glomerular hyperfiltration through overproduction of nitric oxide in early stage of diabetic nephropathy. In this study, we tested the hypothesis that BPS ameliorates glomerular hyperfiltration through modulating ecNOS expression in diabetic nephropathy. Furthermore, we examined the effects of BPS on the expression of intercellular adhesion molecule-1 (ICAM-1) and macrophage infiltration in diabetic glomeruli, because glomerular hyperfiltration induces the expression of ICAM-1 resulting in macrophage infiltration. Male Sprague-Dawley (SD) rats were administered continuously with BPS for 4 weeks after induction of diabetes by streptozotocin. In diabetic rats, the diameters of afferent arterioles, glomerular volume, creatinine clearance and urinary excretion of albumin and NO2/NO3 were increased as compared with non-diabetic control rats. Treatment with BPS improved these changes. The expression of ecNOS was increased in afferent arterioles and glomeruli in diabetic rats and suppressed by BPS. Prostacyclin receptor was expressed along afferent arterioles. Our results suggest that BPS attenuates glomerular hyperfiltration by modulating ecNOS expression in early stage of diabetic nephropathy. Moreover, BPS may inhibit ICAM-1-dependent infiltration of macrophages in diabetic glomeruli. PMID- 12126765 TI - The effect of high carbohydrate diet on glucose tolerance in patients with type 2 diabetes mellitus. AB - The effect of high carbohydrate (hc) diet on glucose tolerance and on lipid profiles in patients with type 2 diabetes mellitus is contradicted. Japanese patients with mild type 2 diabetes mellitus were allocated either 55% standard carbohydrate (sc) or 80% high carbohydrate diets for 1 week, and OGTT and lipid profiles were examined. Then the diet was crossed over for another week, and OGTT and other identical parameters were re-evaluated. High carbohydrate diet improved the area under the glucose concentration-time curve (AUG) in 16/24 patients, and significantly increased and decreased 1,5-anhydroglucitol and homeostasis model assessment insulin resistance (HOMA-R) as a whole, respectively. Fasting plasma glucose (FPG) hc/sc ratio was inversely correlated with HOMA-R on a standard carbohydrate diet. High carbohydrate diet significantly decreased LDL- and HDL cholesterol, whereas it significantly increased triglyceride. Furthermore, hc/sc ratios of the lipid parameters were inversely correlated with the respective parameters on standard carbohydrate diet. The present study indicates that high carbohydrate diet improved glucose tolerance depending on patients and the improvement in FPG was predicted by HOMA-R on a standard carbohydrate diet. The effect of high carbohydrate diet on glucose tolerance and lipid profiles should be investigated through a long-term study in the future. PMID- 12126766 TI - Evidence of autoantibodies to glutamic acid decarboxylase in oral fluid of type 1 diabetic patients. AB - Circulating antibodies to glutamic acid decarboxylase (GADab) are a major indicator for autoimmune destruction of pancreatic islet cells (type 1 diabetes). Previously reported detection of GADab in oral fluid, however, was assayed by enzyme-linked immunosorbent assay (ELISA) with low diagnostic sensitivity and high non-specific binding. We re-assessed oral fluid GADab detection using a different sampling technique and a more robust assay. Type 1 diabetic subjects (n = 32; mean age +/- SD: 13.9 +/- 3.7 years) provided Orasure oral fluid and venous blood samples. Orasure collections were assayed for total immunoglobulin G (IgG), then concentrated to 1/10 of their volume using mini-centrifugal protein concentrators. All samples were assayed by a GAD65 antibody radio immunoprecipitation method. Oral fluid antibodies were detected ( > 99th percentile of radio-binding (%counts per min (%cpm)) for seronegatives) in 10/16 seropositive subjects, with %cpm (median: 6.4%; range: 4.6-25.8) significantly greater (P < 0.001) than for seronegatives (median: 4.7%; range: 3.4-5.7). A highly significant correlation (Spearman's rho: 0.85; P<0.001) was demonstrated between %cpm of concentrates and respective serum titres for seropositive diabetics. Median IgG concentration of Orasure collections was 22.8 mg/l (range: 9.4-168.0). GADab recovery from Orasure collectors was estimated at 90%. This is the first confirmatory detection of diabetes-specific autoimmune markers in oral fluid. Acceptable correlation between concentrated oral fluid radio-binding and serum titre was achieved. Improved antibody recovery and assay re-optimisation could provide a basis for more extensive studies that may lead to an alternative non-invasive screening method for pre-clinical autoimmune diabetes. PMID- 12126768 TI - Prevalence of overweight in urban Indian adolescent school children. AB - The prevalence of diabetes mellitus (DM) and cardiovascular disease (CVD) is increasing in urban India. Overweight in adolescence is a marker of overweight in adult age, and it shows an association with the above diseases. There have been meagre data from India on the prevalence of childhood obesity. The objective of the study was to quantify the prevalence of overweight and its risk factors in adolescent children in urban India. School students in the age group of 13-18 years (n = 4700, M:F 2382:2318) were studied. Body mass index (BMI) was measured. Data on physical activity, food habits, occupation of parents and their economic status, birth weight of the children and age at menarche in girls were obtained by questionnaire. Age-adjusted prevalence of overweight was 17.8% for boys and 15.8% for girls. It increased with age and was higher in lower tertiles of physical activity and in higher socio-economic group. Birth weight and current BMI were positively associated. The study highlighted the high prevalence of overweight in adolescent children in urban India. Life style factors influenced BMI in adolescent age. PMID- 12126767 TI - Renoprotective effects of low-dose valsartan in type 2 diabetic patients with diabetic nephropathy. AB - It is unknown whether the angiotensin receptor antagonist valsartan exerts a renoprotective effect on patients with type 2 diabetes and diabetic nephropathy independent of its hypotensive effects. Forty patients with type 2 diabetes participated in this study. All patients received valsartan 40 mg, a dose with no clinical effect on blood pressure levels. Blood pressure, urinary albumin excretion (UAE), and creatinine clearance were determined at baseline and at the end of the 6-month treatment period. Antihypertensive and/or antidiabetic drugs, including insulin, were permitted throughout the study. After 6 months of valsartan therapy, mean UAE decreased from 86.8 +/- 196 to 46.9 +/- 97 microg/min (n = 37). In addition, a significant decrease was observed in the UAE of the subgroup of patients displaying diabetic nephropathy (UAE > 20 microg/min, n = 14), from 219.4 +/- 275 to 102.7 +/- 141 microg/min, (P < 0.01). Changes in UAE for valsartan correlated significantly with UAE at baseline (r = -0.935, P < 0.0001). Serum creatinine levels and creatinine clearance remained stable before and after treatment with valsartan. No significant differences were observed between pre- and post-treatment body mass index, glycosylated hemoglobin, or systolic and diastolic blood pressure. In type 2 diabetic patients with diabetic nephropathy, 6 months of treatment with low dose valsartan, an angiotensin-II receptor antagonist, thus reduced UAE with no reduction in systemic blood pressure. The drug may be safely administered in this subset of type 2 diabetic patients. The long-term benefits in terms of risk reduction must still be evaluated in further trials. PMID- 12126769 TI - Alcohol consumption and the risk of diabetes by body mass index levels in a cohort of 5,636 Japanese. AB - The association between alcohol consumption and the risk of diabetes in Japanese with a low-body mass index (BMI) (< or = 22.0 kg/m(2)), middle-BMI (22.1-24.9 kg/m(2)) and high-BMI (> or =25.0 kg/m(2)) was investigated among a cohort of 5,636 employees of a Japanese insurance company. Participants were free of diabetes at baseline and were followed up for a mean of 5.7 years with annual assessments of fasting plasma glucose (FPG). The outcome was a clinical diagnosis of diabetes on the basis of a questionnaire administered at each follow-up assessment or a follow-up FPG level of 7.8 mmol/l or more. Relative risks and 95% confidence intervals (95% CIs) were estimated by fitting pooled logistic regression models, which included age, gender, BMI, baseline FPG level, current tobacco use and current alcohol consumption. A total of 264 outcome events were recorded. The relative risk of diabetes associated with current alcohol consumption was 3.19 (95% CI 1.09-9.37) among low-BMI individuals, 0.41 (0.23 0.73) among middle-BMI individuals and 0.74 (0.44-1.25) among high-BMI individuals. In this study, current alcohol consumption was associated with an increased risk of diabetes among low-BMI individuals and a decreased risk of diabetes among middle-BMI individuals. A tendency for an association of alcohol consumption with a decreased risk of diabetes among high-BMI individuals was noted, although without statistical significance. PMID- 12126771 TI - Pulmonary function parameters in patients with diabetes mellitus. PMID- 12126770 TI - Relationships of apolipoprotein B(100) with the metabolic syndrome in Type 2 diabetes mellitus. AB - The current study assessed whether features of the metabolic syndrome are associated with higher apolipoprotein B(100) (apoB(100)) levels in people with Type 2 diabetes (n = 298) not taking lipid-lowering drugs. Body-mass index (BMI), waist:hip ratio (WHR), urinary albumin excretion rate, presence or absence of hypertension, uric acid levels, and apoB(100) levels were assessed. Both higher BMI and urinary albumin excretion rate were associated with higher apoB(100) levels (1.02 +/- 0.25 ( +/- S.D.) g/l in normal weight, 1.07 +/- 0.22 g/l in overweight and 1.14 +/- 0.25 g/l in obese individuals; P < 0.01; 1.09 +/- 0.23 g/l in normoalbuminuric patients, 1.06 +/- 0.22 g/l if urinary albumin excretion rate 20-50 microg/min and 1.17 +/- 0.27 g/l if urinary albumin excretion rate > 50 microg/min; P < 0.05). An association between the number of features of the metabolic syndrome and higher apoB(100) levels was found (1.03 +/- 0.22 g/l if no features, 1.08 +/- 0.25 g/l if one feature, 1.11 +/- 0.20 g/l if two features and 1.15 +/- 0.27 g/l if > 2 features; P for trend < 0.01). Thus apoB(100) levels show an association with the metabolic syndrome and, hypothetically, to insulin insensitivity in Type 2 diabetes. BMI (but not WHR) and urinary albumin excretion rate accounted for most of the power of this relationship. PMID- 12126772 TI - Arylethylamine psychotropic recreational drugs: a chemical perspective. AB - The arylethylamines substituted in the aryl ring, side-chain carbons and on the terminal amine, comprise a large number of human mood and behaviour altering chemicals. Some of these psychotropic drugs have been used since pre-history, but in many states are proscribed and are consequently subject to clandestine synthesis and illegal traffic world-wide in the forms particularly of amphetamines and to a lesser extent tryptamines. The chemistry employed in the synthesis of these compounds is dictated often by the available precursors and relies usually on relatively simple, unsophisticated conversion reactions to a suitable product. The internet web sites and documentation of the recreational drug culture have been studied alongside the professional scientific and regulatory literature. The review demonstrates the great complexity of the chemistry and neuro-pharmacology of these chemicals and the challenge faced by legislative bodies to control their traffic and use for the sake of social welfare. PMID- 12126773 TI - Synthesis and antitumour activity of new dendritic polyamines-(imide-DNA intercalator) conjugates: potent Lck inhibitors. AB - A series of dendritic polyamines-(imide-DNA-intercalators) conjugates with different connectivity in their basic chain were synthesised and evaluated as antitumour compounds. Although their antiproliferative activity against HT-29 was not significant, conjugates 13 and 16 showed a promising profile as inhibitors of Lck. PMID- 12126774 TI - Hydrazones of 1,2-benzisothiazole hydrazides: synthesis, antimicrobial activity and QSAR investigations. AB - A series of hydrazones of 1,2-benzisothiazole hydrazides 1a-m, 2a-m, 3a-m, 4a-m, 5a-m as well as their cyclic (1 and 4) and acyclic (2, 3 and 5) 1,2 benzisothiazole parent hydrazides, were synthesised and evaluated as antibacterial and antifungal agents. All of the 2-amino-1,2-benzisothiazol-3(2H) one derivatives, belonging to series I and IV, showed a good antibacterial activity against Gram positive bacteria. Most of them were active against yeasts too. Compounds 1 and 4, together with 1l, proved to be the most effective compounds. Quantitative structure-activity relationship (QSAR) investigation with a 2D-QSAR analysis was applied to find a correlation between different experimental or calculated physicochemical parameters of the compounds studied. A 3D-QSAR study was performed, applying Comparative Molecular Similarity Indices Analysis (CoMSIA) method, to derive quantitative models relating the structural features of 1,2-benzisothiazole derivatives 1, 1a-m and 4, 4a-m and their antimicrobial activity against Bacillus subtilis, resulted the most sensitive micro-organism. PMID- 12126775 TI - New 1,2,3-triazolo[1,5-a]quinoxalines: synthesis and binding to benzodiazepine and adenosine receptors. II. AB - On pursuing research about 1,2,3-triazolo[1,5-a]quinoxalines, in this paper we report synthesis and binding assays toward the benzodiazepine and A(1) and A(2A) adenosine receptors, of a new series of derivatives, bearing some structural changes (introduction of fluorine and trifluoromethyl in the seventh position, amino substituents in the fourth position, benzyl group in the fifth position and aroyl substituents in the third position). The biological tests have shown that only the 7-fluorosubstituted compounds 3a and 4a and the N-benzyl derivative 7 have a good affinity toward the benzodiazepine receptors, while only the 7 trifluoromethyl substituted compound 3b presents a moderate affinity with low selectivity toward the A(1) adenosine receptors. The other structural modifications strongly decreased biological activity. PMID- 12126776 TI - Modelling, synthesis and biological evaluation of an ethidium-arginine conjugate linked to a ribonuclease mimic directed against TAR RNA of HIV-1. AB - Using molecular modelling studies, an active anti-HIV ethidium-arginine conjugate targeted against the viral TAR RNA sequence has been linked to an artificial ribonuclease, with the aim to obtain an irreversible inhibitor. The ribonuclease moiety consists of an N-[N-(3-aminopropyl)-3-aminopropyl] glycine and has been constructed via two successive N-alkylations following the Fukuyama procedure. PMID- 12126777 TI - Aryl-substituted methyleneaminoxymethyl (MAOM) analogues of diarylcyclopentenyl cyclooxygenase-2 inhibitors: effects of some structural modifications on their biological properties. AB - The (E)-[2-(4-aminosulfonylphenyl)-1-cyclopentenyl-1-methyliden] (arylmethyloxy)amines (6a,b), which are the sulfonamidic analogues of the previously described methylsulfonyl derivatives 5a,b, and their corresponding sulfides (7a,b) and sulfoxides (8a,b) were synthesised in order to obtain information about the role played by these different sulphur-containing groups in the cyclooxygenase-2 inhibitory activity of this class of compounds. In addition, other chemical manipulations concerning the oxime-ether substituent of this type of derivatives were affected by preparing compounds 9a,b, which present a methyl group on the oximic carbon of the oxime-ether chain of 5a,b, and compounds 10 and 11, in which the atomic sequence (C=NOCH(2)) of the MAOMM of 8b and 5b, respectively, is inverted. Compounds 6-11 were tested in vitro for their inhibitory activity towards COX-1 and COX-2 by measuring prostaglandin E2 (PGE2) production in U937 cell lines and activated J774.2 macrophages, respectively. Some of the new compounds showed an appreciable in vitro COX-2 inhibitory activity, with IC(50) values in the microM (7a,b, 8a and 9b) or sub-microM (8b) range. This last compound was also assayed in vivo for its antiinflammatory activity by means of the carrageenan-induced paw edema test in rats. No inhibitory effects were detected up to dose of 30 mg kg(-1) orally administered. PMID- 12126778 TI - On the mechanism of human intestinal absorption. AB - In order to investigate whether the main step in intestinal absorption in humans is dominated by partition or by diffusion, we have transformed % human intestinal absorption into a first-order rate constant, and have regressed the latter, as logk, against our solvation parameters. The obtained regression coefficients are compared with those for diffusion and partition processes. The coefficients in the logk equation are completely different to those for water/solvent partitions, but are very similar to those for processes (not involving transport through membranes) in which diffusion is the major step. It is suggested that the main step in the absorption process is diffusion through a stagnant mucus layer, together with transfer across the mucusmid R:membrane interface. It is further shown that for strong Bronsted acids and bases, the rate constant for absorption of ionic species is close to that for absorption of the corresponding neutral species, so that to a first approximation the % intestinal absorption can be calculated from properties of the neutral species. PMID- 12126779 TI - Cinnamic acid derived oxazolinium ions as novel cytotoxic agents. AB - Substituted cinnamoyl chlorides, 11, were converted into (2-hydroxyethyl) oxazolinium chlorides 14, N,N-bis-(2-chloroethyl)amides 16 and (2-chloroethyl) oxazolinium chlorides 17. Although derivatives 14 which possess electron-donating substituents (Me or MeO) were more potent than those substituted by electron withdrawing groups (NO(2), Cl or CF(3)), the difference in cytotoxic actin was not significant. Modification of the lipophilic character in a series of alkoxy substituted derivatives 14 led to more active compounds, where 14t that possesses a 4-octyloxy-phenyl-substituent was the most potent and displayed cytotoxic activity in the microM range. It is assumed that the oxazolinium salts act as alkylating agents, and undergo nucleophilic attack on the methylene adjacent to the ring oxygen where the oxazolinium ring parallels the aziridinium ring intermediate found in classical alkylating agents. PMID- 12126783 TI - ACE and PC-1 gene polymorphisms in normoalbuminuric Type 1 diabetic patients: a 10-year prospective study. AB - The aim of this study was to analyze the role of ACE gene insertion/deletion (I/D) and PC-1 gene K121Q polymorphisms in the changes of glomerular filtration rate (GFR), urinary albumin excretion rate (UAER), and blood pressure (BP) levels in a cohort of normoalbuminuric Type 1 diabetic patients. This is a 10.2+/-2.0 year prospective study of 30 normotensive normoalbuminuric Type 1 diabetic patients. UAER (immunoturbidimetry), GFR ((51)Cr-EDTA single injection technique), GHb (ion exchange chromatography), and BP levels were measured at baseline and at 1.7+/-0.6-year intervals. The presence of ACE gene I/D and PC-1 gene K121Q polymorphisms was determined by polymerase chain reaction (PCR) and restriction enzyme techniques. Three patients developed diabetic nephropathy (DN), all carriers of allele D. The presence of allele D was the only predictor (R(2)=.15, F=4.92, P=.035) of the observed GFR decline (-0.29+/-0.34 ml/min/month, P<.05). UAER increased during the study (log UAER=0.0275+/-0.042 microg/min/month, P=.002) and was associated with baseline UAER levels only (R(2)=.17, F=5.72, P=.024). A significant increase (P<.05) in cases of hypertension and retinopathy were observed in ID/DD (n=19) and not in II patients (n=11). Patients with the KQ/QQ genotype (n=8) presented a significant increase (P=.045) in new cases of retinopathy. In conclusion, the presence of the ACE gene D allele in this sample of normoalbuminuric normotensive Type 1 diabetic patients was associated with a higher proportion of microvascular complications and hypertension. PMID- 12126784 TI - Late potentials: are they related to cardiovascular complications in children with type 1 diabetes? AB - INTRODUCTION: The aim of our study was to assess the influence of cardiovascular complications on the occurrence of late ventricular potentials (LP) in children with diabetes mellitus type 1. MATERIALS AND METHODS: 72 children (36 boys and 36 girls), with average course of diabetes type 1 of 6.5+/-2.8 years, were included into the study. Standard physical examination, blood pressure measurements, signal-averaged electrocardiogram (SAECG), autonomic test, 24-h Holter monitoring, and Doppler echo investigations were performed. The control group consisted of 55 sex- and age-matched healthy children. We utilised nonlinear logistic regression analysis to assess the effect of disease duration, albuminuria, insulin demand, cardiac autonomic neuropathy (CAN), heart rate variability (HRV) indices, diabetes complication score, metabolic control, systolic and diastolic blood pressure, left ventricular parameters, and ventricular arrhythmias on LP occurrence. RESULTS: LP was discovered in 12 patients with diabetes and in 1 from the control group (P<.014). Diabetic children with LPs had thicker left ventricular posterior wall (LVPW) and longer diabetes duration time than children without LP (P<.045 and.031, respectively). Nonlinear regression model shows that duration of diabetes, CAN, and LVPW were the strongest independent parameters of LP occurrence (P<.001,.01 and.005, respectively). CONCLUSIONS: Diabetes type 1 is associated not only with increased occurrence of abnormal SAECG but also with LP presence. The disease duration, posterior wall thickness, and CAN are independent predictors of LP appearance in diabetes type 1 children. The presence of cardiovascular complication has no influence on LP occurrence in diabetic children. PMID- 12126785 TI - Risk of macrovascular and microvascular complications in Type 2 diabetes: results of longitudinal study design. AB - Type 2 diabetes is associated with the increased risk of microvascular and macrovascular complications. The aim of this study was to determine risk factors for the development of long-term complications of Type 2 diabetes. We analyzed medical records of all patients, who came with newly diagnosed Type 2 diabetes to one regional outpatient diabetes clinic from 1980 to 1994 (n=2175). The data, such as fasting plasma glucose, total cholesterol, triglyceride, blood pressure and body mass index (BMI), were assessed. Also, the time from the diagnosis of Type 2 diabetes to the occurrence of complications was obtained. Using the regression model in the survival analysis, we examined which of the risk factors determined the rate of the development of nephropathy, proliferative retinopathy, cardiovascular disease and stroke. Patients with higher fasting plasma glucose and higher mean blood pressure had higher risk of developing nephropathy and proliferative retinopathy. Higher mean arterial blood pressure was associated with higher rate of stroke and cardiovascular disease. High total cholesterol increased the hazard of coronary artery disease and proliferative retinopathy. In conclusion, blood pressure and fasting plasma glucose are major risk factors for microvascular complications in Type 2 diabetes. An increased blood pressure determined the macrovascular complications in Type 2 diabetes, but the effect of increased fasting plasma glucose could not be proved. PMID- 12126786 TI - Determinants of plantar pressures in the diabetic foot. AB - The aim of this study was to determine, by the use of regression analysis, the factors that are associated with the increased plantar pressure in the diabetic foot. In-shoe plantar pressure measurements using the Novel Pedar were carried out on 50 subjects with diabetes. Variables measured were age, body weight, duration of diabetes, a number of selected structural radiographic angles, soft tissue thickness, plantarflexion, and dorsiflexion strength at the ankle and first metatarsophalangeal joint, Neuropathy Symptom Score, and the Michigan Neuropathy Disability Score. Stepwise regression modelling indicates that 28% of the variability in hallux peak pressure could be explained by the first metatarsophalangeal joint range of motion and the Michigan Neuropathy Disability Score (P=.0004). The Michigan Neuropathy Disability Score explained 17% of the peak pressure under the first metatarsal head (P=.002). None of the measured variables could explain any of the variation in peak pressure plantar to the lateral forefoot. Thirty-two percent (32%) of the variability in peak pressure under the heel was explained by the Michigan Neuropathy Disability Score and age (P<.0001). Very little of the variation in the pressure time integrals could be explained by the measured variables except for 10.3% of the variation in the pressure time integral for the heel being explained by body weight. This study has shown that neuropathy-related variables play an important role in the plantar pressure under the diabetic foot. The range of motion of the first metatarsophalangeal joint is also important in determining pressures under the hallux. PMID- 12126787 TI - Advanced glycation end products increase, through a protein kinase C-dependent pathway, vascular endothelial growth factor expression in retinal endothelial cells. Inhibitory effect of gliclazide. AB - Accumulating evidence points to a causal role for advanced glycation end products (AGEs) in the development of diabetic vascular complications, including retinopathy. Possible pathogenic mechanisms linking AGEs to diabetic retinopathy include protein kinase C (PKC) activation, oxidative stress, and vascular endothelial growth factor (VEGF) expression. In the present study, we investigated the effect of AGEs on VEGF expression in bovine retinal endothelial cells (BRECs) and determined the role of PKC and oxidative stress in this effect. Incubation of BRECs with AGEs led to enhanced VEGF mRNA and protein expression. This treatment also induced PKC translocation in these cells. The AGE-induced increases in VEGF expression and PKC activation were inhibited by the pan specific PKC inhibitor, calphostin C, and by the antioxidant drug and compounds, gliclazide, N-acetylcysteine, and vitamin E. In contrast, glyburide which does not exhibit antioxidant properties, did not affect the AGE-induced VEGF expression. Exposure of BRECs to AGEs resulted in a significant increase of nuclear protein binding to the NF-kappa B consensus sequence of the VEGF promoter region. Induction of DNA binding activity for NF-kappa B by AGEs was prevented by gliclazide. Treatment of BRECs with AGEs also increased the proliferation of these cells. This effect was abrogated by incubating the cells with an anti-VEGF antibody and was inhibited in the presence of gliclazide. Overall, these data demonstrate that AGEs increase VEGF expression in retinal endothelial cells through generation of oxidative stress and downstream activation of the PKC pathway. Targeting VEGF expression with specific pharmacological agents, such as antioxidants and PKC inhibitors, may prove efficacious for the treatment of diabetic retinopathy. PMID- 12126788 TI - Lipid peroxidation and antioxidant vitamins prior to, during, and after correction of diabetic ketoacidosis. AB - Oxidative stress plays an important role in the chronic complications of insulin dependent diabetes mellitus (IDDM). Hyperketonemia, as well as hyperglycemia, is involved in the generation of oxygen-free radicals. We have studied the degree of oxidative stress in six patients before, during, and after correction of diabetic ketoacidosis (DKA) by determining the plasma ratios of C20 and C18 fatty acids to C16 fatty acids in the cholesteryl esters of the lipoproteins as well as in the plasma concentrations of the antioxidant vitamins A, C, and E. Lipid peroxidation was slightly increased prior to treatment. However, the C20/C16 ratio at 120 h was significantly decreased in comparison to the ratio at pretreatment (P<.025), at 6-8 h (P<.005), and at 24 h (P<.025). The C18/16 ratio at 120 h was also decreased in comparison to the ratio at 6-8 h (P<.025), indicating an increase in lipid peroxidation after correction of DKA. Vitamin A was below normal at pretreatment and was increased at 120 h (P<.05). Vitamin C was normal at pretreatment and decreased to low normal at 24 h (P<.005). Vitamin E was normal at pretreatment and decreased to below normal at 24 and 120 h, although the changes were not statistically significant. These data demonstrate that there is an increase in lipid peroxidation after the correction of DKA and therefore support the position that administering antioxidant vitamins during the treatment of DKA could be beneficial in minimizing oxidative stress and possibly both the acute and chronic complications of IDDM. PMID- 12126790 TI - Effect of oral fluid intake on urinary albumin excretion in diabetes mellitus. AB - The importance of routine screening for microalbuminuria in patients with diabetes has gained widespread acceptance. Since shortened collection times and random urine sampling facilitate outpatient testing for patients, it is necessary to establish whether a recent fluid intake might alter test results. Specifically, does an oral fluid load (15 ml/kg) effect urinary albumin excretion in Type I diabetes? Indeed, we show that fluid intake had a significant impact: albumin concentration decreased, whereas the albumin/creatinine ratio (A/C) and albumin excretion rate increased (P<.001) significantly. PMID- 12126789 TI - The effects of aminoguanidine on renal changes in a baboon model of Type 1 diabetes. AB - BACKGROUND: The efficacy of aminoguanidine (AG) on primary prevention of diabetic nephropathy was investigated in a nonhuman primate model of Type 1 diabetes over a period of 4 years. METHODS: Adolescent male baboons (Papio hamadryas) were assigned to four groups: control, diabetic, and control and diabetic treated with AG. Diabetes was induced with streptozocin (60 mg/kg) and treated with insulin to maintain a mean HbA1c level of about 9%. AG was given subcutaneously (10 mg/kg) each day. All animals had annual renal biopsies and 24-h urine collections for measurements of glomerular basement membrane (GBM) thickness, fractional mesangium volume (FMV), albumin excretion rate (AER), and creatinine clearance. Glomerular filtration rate (GFR) and renal plasma flow (RPF) were also determined. RESULT: The diabetic animals had increased GBM after 2 years of diabetes, but there was no increase in FMV over the study period. AG prevented the thickening of GBM at the 3- and 4-year time points. AG and diabetes synergistically increased the GFR. All diabetic animals developed increased albuminuria during the study although lower than the conventionally accepted microalbuminuria range. AG was not able to prevent this and, in fact, led to the nondiabetic animals also developing albuminuria. CONCLUSION: This is the first study to investigate the early use of AG in ameliorating renal damage in a primate model of Type 1 diabetes. The structural and functional changes in the kidney of these animals resemble those seen in the early stages of the human disease. AG was able to significantly reduce the thickening of GBM due to diabetes. This may suggest a potential role for this in primary prevention of diabetic nephropathy in the future. PMID- 12126791 TI - Nitric oxide trapping of the tyrosyl radical-chemistry and biochemistry. AB - The quenching of the Y(D) tyrosyl radical in photosystem II by nitric oxide was reported to result from the formation of a weak tyrosyl radical-nitric oxide complex. This radical/radical reaction is expected to generate an electron spin resonance (ESR)-silent nitrosocyclohexadienone species that can reversibly regenerate the tyrosyl radical and nitric oxide or undergo rearrangement to form 3-nitrosotyrosine. It has been proposed that 3-nitrosotyrosine can be oxidized by one electron to form the tyrosine iminoxyl radical (>C=N-O.). This proposal was put forth as a result of ESR detection of the iminoxyl radical intermediate when photosystem II was exposed to nitric oxide. Although the detection of the iminoxyl radical in photosystem II strongly suggested a mechanism involving 3 nitrosotyrosine, the iminoxyl radical ESR spectrum was not unequivocally identified as originating from tyrosine. Subsequently, non-protein L-tyrosine iminoxyl radical was generated by two methods: (1) peroxidase oxidation of synthetic 3-nitroso-N-acetyl-L-tyrosine; and (2) peroxidase oxidation of free L tyrosine in the presence of nitric oxide. The determination of protein nitrotyrosine content has become a frequently used technique for the detection of nitrosative tissue damage. Protein nitration has been suggested to be a final product of the production of highly reactive nitrogen oxide intermediates (e.g. peroxynitrite) formed in reactions between nitric oxide (NO.) and oxygen-derived species such as superoxide. The enzyme prostaglandin H synthase-2 also forms a tyrosyl radical during its enzymatic catalysis of prostaglandin formation. In the presence of the NO.-generator diethylamine nonoate, the tyrosyl radical of prostaglandin H synthase-2 also changes to that of an iminoxyl radical. Western blot analysis of prostaglandin H synthase-2 after exposure to the NO.-generator revealed nitrotyrosine formation. The results provide a mechanism for nitric oxide-dependent tyrosine nitration that does not require formation of more highly reactive nitrogen oxide intermediates such as peroxynitrite or nitrogen dioxide. PMID- 12126792 TI - Tyrosyl radical production by myeloperoxidase: a phagocyte pathway for lipid peroxidation and dityrosine cross-linking of proteins. AB - To kill invading bacteria, viruses, and fungi, phagocytes secrete hydrogen peroxide (H(2)O(2)) and the heme enzyme myeloperoxidase. We have explored the possibility that myeloperoxidase might use H(2)O(2) to convert L-tyrosine to tyrosyl radical. Activated human neutrophils and monocytes used the system to oxidize free L-tyrosine to o,o'-dityrosine, a stable product of tyrosyl radical. Protein-bound tyrosyl residues exposed to myeloperoxidase, H(2)O(2), and L tyrosine were also oxidized to o,o'-dityrosine. The cross-linking reaction required free L-tyrosine, suggesting that myeloperoxidase converts the amino acid to a diffusible radical catalyst that promotes protein oxidation. We used electron paramagnetic resonance to provide direct evidence that the oxidizing intermediate is free tyrosyl radical. Myeloperoxidase-generated tyrosyl radical also initiates lipid peroxidation, suggesting that activated phagocytes might also be able to oxidize lipids in host tissues. Moreover, myeloperoxidase is present and active in human atherosclerotic tissue, and levels of protein-bound dityrosine are elevated in such lesions. Our recent studies indicate that activated neutrophils use oxidants generated by the phagocyte NADPH oxidase to produce protein-bound dityrosine during acute inflammation. Collectively, these findings suggest that generation of tyrosyl radical by myeloperoxidase allows activated phagocytes to damage both proteins and lipids. Elevated levels of o,o' dityrosine have been detected in inflammatory lung disease, neurodegenerative disorders, and aging. Thus, oxidation of tyrosine to tyrosyl radical might play a role in the pathogenesis of many diseases. PMID- 12126793 TI - Oxidation of DNA, proteins and lipids by DOPA, protein-bound DOPA, and related catechol(amine)s. AB - Incubation of free 3,4-dihydroxyphenylalanine (DOPA), protein-bound DOPA (PB DOPA) and related catechols with DNA, proteins and lipids has been shown to result in oxidative damage to the target molecule. This article reviews these reactions with particular emphasis on those that occur in the presence of molecular O(2) and redox-active metal ions (e.g. Fe(3+), Cu(2+), Cr(6+)), which are known to increase the rate of DOPA oxidation. The majority of oxidative damage appears to be mediated by reactive oxygen species (ROS) such as superoxide and HO(.) radicals, though other DOPA oxidation products, including semiquinone radicals, quinones, and metal ion-DOPA complexes have also been implicated in some cases. Non-radical reactions of DOPA with suitable nucleophiles (e.g. thiol groups) can also result in modification of the target, with this process being particularly prevalent with proteins. The exacerbation of damage observed on addition of H(2)O(2) is in accord with a key role for ROS in many of these reactions. PMID- 12126794 TI - Antioxidant and prooxidant action of eugenol-related compounds and their cytotoxicity. AB - To clarify the possible link between radicals and the cytotoxicity of eugenol related compounds, 2-allyl-4-X-phenols (2-allyl-4-chlorophenol (1), 2-allyl-4 phenylphenol (2), 2-allyl-4-methoxyphenol (3), 2-allyl-4-acetylphenol (4), 2 allyl-4-nitrophenol (5), 2-allyl-4-t-butylphenol (6), 2-allyl-4-methyphenol (7), 2-allyl-4-bromophenol (8), 2,4-dimethoxyphenol (9)), and dimeric compounds from eugenol (4-allyl-2-methoxyphenol), BHA (2-t-butyl-4-methoxyphenol) or MMP (2 methoxy-4-methylphenol); bis-EUG (3,3'-dimethoxy-5,5'-di-2-propenyl-1, 1' biphenyl-2,2'-diol) (10), bis-MMP (3,3'-dimethoxy-5,5'-dimethyl-1,1'-biphenyl 2,2'-diol) (11) bis-BHA (3,3'-di-t-butyl-5,5'-dimethoxy-1,1'-biphenyl-2,2'-diol) (12) were synthesized. The radical production, radical-scavenging activity and the cytotoxicity of these synthetic compounds and conventional antioxidants (i.e. butylhydroxytoluine, BHT; butylhydroxyanisole, BHA; alpha-tocopherol (alpha-Toc); eugenol, phenol) were studied. Erectron spin resonance (ESR) spectroscopy suggested that compounds of 3, 6, 9, eugenol and BHA, but not compounds of 10, 11, and 12 produced radicals in alkaline solutions (pH>9.5) and compounds, 3, eugenol and 9 most efficiently scavenged reactive oxygen species (ROS, O(2)(-)). The cytotoxic activity of 6 toward human submandibular gland carcinoma (HSG) cells was the highest and was 1000-fold greater than that of eugenol and 100-fold greater than that of BHA, possibly due to the high hydrophobicity and stable phenoxy radicals of this compound. The kinetic polymerization method in the presence of methyl methacrylate (MMA), an antioxidant, and 2,2' azobisisobutyronitrile (AIBN) was developed for the measurements of the number of moles of peroxy radicals trapped by moles of the relative phenols (stoichiometric factors, n), the inhibition rate of polymerization (R(inh)), and the inhibition rate constants (k(inh), the rate constants for scavenging of radicals by an antioxidant). The n values of conventional phenolic antioxidants decreased in the order: alpha-Toc>BHT>eugenol>phenol. Those for eugenol and phenol, less hindered phenols, were much less than two, whereas those for alpha-Toc and BHT, hindered phenols, were approximately two. The R(inh) of alpha-Toc significantly increased tcompared with that of BHT, eugenol and phenol. The k(inh) of the polymer radicals of the MMA reaction with conventional phenolic antioxidants was a low value of 1-2x10(2) M(-1) s(-1), suggesting that the antioxidants trapped radicals quickly. The comparative cytotoxicity of methoxyphenols against HSG cells was investigated. The cytotoxic activity of dimers of 10 and 12 was markedly lower than that of their corresponding monomers, whereas that of the dimer of MMP, 11 was not reduced even after the dimerization. In particular, visible-light (VL) exposure enhanced the cytotoxicity of 11 similar to the monomers of eugenol, BHA and MMP. Changes in BDE (ph(O-H)) (homolytic bond dissociation energy) for phenols is well known to be associated with the n and k(inh) values, and consequently the cytotoxic activity. Thus, the BDE was calculated using a PM3 semiempirical method. The n and k(inh) values for monophenols, but not for dimers were correlated to the BDE, possibly due to the steric hindrance of orthosubstituents of dimers. The quantitative structure-activity relationship (QSAR) of eugenol-related compounds was investigated, indicating that logP (octanol-water partition coefficients), the redox potential measured in culture medium, was effective as a term for QSAR. A parabolic relation between the cytotoxic activity and the logP or the redox potential, but not the BDE was observed with an optimum value. In conclusion, the cytotocity of eugenol-related compounds was significantly associated with the activity of the production of phenoxyl radicals, their stability of the subsequent quinonemethide (QM) and the hydrophobicity. PMID- 12126795 TI - Quinoids, quinoid radicals, and phenoxyl radicals formed from estrogens and antiestrogens. AB - Estrogens have a variety of beneficial effects in vivo including protection against osteoporosis, coronary heart disease, Alzheimer's disease, and stroke. Similarly, antiestrogens have been shown to be effective both in treating breast cancer as well as preventing this disease. Despite the health benefits of these drugs adverse side effects have been reported including increased risk for developing certain hormone dependent cancers. Although an estrogenic mechanism likely contributes to mechanism of estrogen/antiestrogen carcinogenesis there is substantial evidence to suggest that metabolism to reactive intermediates is also involved. Both estrogens and antiestrogens can be metabolized to phenoxyl radicals, o-quinones, and semiquinone radicals, all of which could cause damage in cells either through alkylation or oxidation of cellular macromolecules including DNA. In contrast, there are several reports that estrogens and antiestrogens can act as antioxidants which could protect cells against free radical mediated damage and contribute to the beneficial effects of these compounds discussed above. The focus of this review is the role of quinoids, quinoid radicals, and phenoxyl radicals in the biological effects of estrogens and antiestrogens. PMID- 12126796 TI - Plant phenolic antioxidant and prooxidant activities: phenolics-induced oxidative damage mediated by metals in plants. AB - Plant phenolic compounds such as flavonoids and lignin precursors are important constituents of the human diet. These dietary phytophenolics have been recognized largely as beneficial antioxidants that can scavenge harmful active oxygen species including O(2)(.-), H(2)O(2), .OH, and (1)O(2). Here we review our current understanding of the antioxidant and prooxidant actions of phenolics in plant cells. In plant systems, phytophenolics can act as antioxidants by donating electrons to guaiacol-type peroxidases (GuPXs) for the detoxification of H(2)O(2) produced under stress conditions. As a result of such enzymatic as well as non enzymatic antioxidant reactions, phenoxyl radicals are formed as the primary oxidized products. Until recently, phenoxyl radicals had been difficult to detect by static electron spin resonance (ESR) because they rapidly change to non radical products. Application of Zn exerts spin-stabilizing effects on phenoxyl radicals that enables us to analyze the formation and decay kinetics of the radicals. The ESR signals of phenoxyl radicals are eliminated by monodehydroascorbate radical (MDA) reductase, suggesting that phenoxyl radicals, like the ascorbate radical, are enzymatically recycled to parent phenolics. Thus, phenolics in plant cells can form an antioxidant system equivalent to that of ascorbate. In contrast to their antioxidant activity, phytophenolics also have the potential to act as prooxidants under certain conditions. For example, flavonoids and dihydroxycinnamic acids can nick DNA via the production of radicals in the presence of Cu and O(2). Phenoxyl radicals can also initiate lipid peroxidation. Recently, Al, Zn, Ca, Mg and Cd have been found to stimulate phenoxyl radical-induced lipid peroxidation. We discuss the mechanism of phenoxyl radical prooxidant activity in terms of lifetime prolongation by spin-stabilizing agents. PMID- 12126797 TI - Prooxidant activity of free radicals derived from phenol-containing neurotransmitters. AB - It has been suggested that biogenic amines may partake in neurodegenerative disease processes by causing oxidative stress. In the following, we present evidence showing for the first time that biogenic amines can form prooxidant radicals when metabolized. The order of prooxidant activity of neurotransmitter phenols or hydroxyindoles in catalyzing beta-nicontinamide adenine dinucleotide (reduced) (NADH) or cysteine cooxidation found when metabolically activated by peroxidase/H(2)O(2) was tyramine>N-acetyltyrosine>tyrosine>serotonin>N acetylserotonin, 5-hydroxyindoleacetic acid (5-HIAA). This order likely reflects the reactivity of the phenoxyl radicals (for phenols) as extensive oxygen activation accompanied the NADH oxidation and only catalytic amounts of H(2)O(2) were required. The low reactivity of the hydroxyindoles suggests that the redox potential of the radical (semiquinone-imine radical?) was too low to oxidize NADH and/or that the radical dimerization rate was too rapid. The order of catalytic effectiveness for phenolic or hydroxyindole neurotransmitters in catalyzing ascorbate cooxidation on the otherhand, was N-acetylserotonin>serotonin>5 HIAA>>tyramine>N-acetyltyrosine>tyrosine. The first formed hydroxyindole radical product was likely the active cooxidizing species formed from hydroxyindoles. The order for catecholamine catalytic effectiveness in catalyzing NADH or ascorbate cooxidation rate was N-acetyldopamine>3,4-dihydroxyphenylacetic acid (DOPAC)>dopamine>norepinephrine>(-)-3,4-dihydroxyphenylalanine (L DOPA)>epinephrine which correlated with the second-order rate constant for the peroxidase/H(2)O(2) catalyzed oxidation of the catecholamines. However, the total amount of NADH oxidized was proportional to the amount of H(2)O(2) added and was not accompanied by oxygen uptake, suggesting that NADH was oxidized by the o quinone metabolite formed by semiquinone radical disproportionation. These results show that biogenic amines form prooxidant radicals, when metabolized by peroxidase, cooxidize cellular antioxidants (ascorbate, NADH, or cysteine). PMID- 12126798 TI - Prooxidant activity and cellular effects of the phenoxyl radicals of dietary flavonoids and other polyphenolics. AB - Dietary polyphenolics in fruits, vegetables, wines, spices and herbal medicines have beneficial antioxidant, anti-inflammatory and anticancer effects. However, we have observed that dietary polyphenolics with phenol rings were metabolized by peroxidase to form prooxidant phenoxyl radicals which, in some cases were sufficiently reactive to cooxidize GSH or NADH accompanied by extensive oxygen uptake and reactive oxygen species formation. The order of catalytic effectiveness found for oxygen activation when polyphenolics were metabolized by peroxidase in the presence of GSH was phloretin>phloridzin>4,2'-dihydroxy chalcone>p-coumaric acid>naringenin>apigenin>curcumin>resveratrol>isoliquiritigenin>capsaicin>kaempfe ol. Ascorbate was also cooxidized by the phenoxyl radicals but without oxygen activation. Polyphenolics with catechol rings also cooxidized ascorbate, likely mediated by semiquinone radicals. The order of catalytic effectiveness found for ascorbate cooxidation was fisetin luteolin, quercetin, >eriodictyol, caffeic acid, nordihydroguaiaretic acid>catechin>taxifolin, catechol. NADH was stoichiometrically oxidized without oxygen uptake which, suggests that o-quinone metabolites were responsible. GSH was not cooxidized and GSH conjugates were formed, likely mediated by the o-quinone metabolites. Incubation of hepatocytes with dietary polyphenolics containing phenol rings was found to partially oxidize hepatocyte GSH to GSSG while polyphenolics with a catechol ring were found to deplete GSH through formation of GSH conjugates. Dietary polyphenolics with phenol rings also oxidized human erythrocyte oxyhemoglobin and caused erythrocyte hemolysis more readily than polyphenolics with catechol rings. It is concluded that polyphenolics containing a phenol ring are generally more prooxidant than polyphenolics containing a catechol ring. PMID- 12126800 TI - Highlights from ECB10-Biotechnology and Society. PMID- 12126799 TI - Anti-/pro-oxidant effects of phenolic compounds in cells: are colchicine metabolites chain-breaking antioxidants? AB - Effective scavenging of reactive radicals and low reactivity of generated secondary antioxidant radicals towards vital intracellular components are two critical requirements for a chain-breaking antioxidant. Tubulin-binding properties aside, colchicine metabolites remain largely untested for other possible biological activities, including antioxidant activity. Mourelle et al. [Life Sci. 45 (1989) 891] proposed that colchiceine (EIN) acts as an antioxidant and protective agent against lipid peroxidation in a rat model of liver injury. Since EIN as well as two other colchicine metabolites, 2-demethylcolchicine (2DM) and 3-demethylcolchicine (3DM), possess a hydroxy-group on their carbon ring that could participate in radical scavenging, we tested whether they can act as chain breaking antioxidants. Using our fluorescence-HPLC assay with metabolically incorporated oxidation-sensitive cis-parinaric acid (PnA) we studied the effects of colchicine metabolites on peroxidation of different classes of membrane phospholipids in HL-60 cells. None of the colchicine metabolites in concentrations ranging from 10(-6) to 10(-4) M was able to protect phospholipids against peroxidation induced by either azo-initiators of peroxyl radicals or via myeloperoxidase (MPO)-catalyzed reactions in the presence of hydrogen peroxide. However, the metabolites did exhibit dose-dependent depletion of glutathione, resembling the behavior of etoposide, a hindered phenol with antioxidant properties against lipid peroxidation. Electron spin resonance (ESR) experiments demonstrated that in a catalytic system containing horseradish peroxidase (HRP)/H(2)O(2), colchicine metabolites undergo one-electron oxidation to form phenoxyl radicals that, in turn, cause ESR-detectable ascorbate radicals by oxidation of ascorbate. Phenoxyl radicals of colchicine metabolites formed through MPO-catalyzed H(2)O(2)-dependent reactions in HL-60 cells and via HRP/H(2)O(2) in model systems can also oxidize GSH. Thus, colchicine metabolites possess the prerequisites of many antioxidants, i.e. a nucleophilic hydroxy-group on a carbon ring and the ability to scavenge reactive radicals and form a secondary radical. However, the latter retain high reactivity towards critical biomolecules in cells such as lipids, thiols, ascorbate, thereby, rendering colchicine metabolites effective radical scavengers but not effective chain breaking antioxidants. PMID- 12126801 TI - People's concerns about biotechnology: some problems and some solutions. AB - Pharmaceuticals and vaccines made by genetic engineering are well accepted all over the world. In contrast, there are many people, particularly in Europe, who are worried that food, made by the same new technology, may harm their health or cause damage to the environment. This is despite the growing evidence that genetically modified crops have the potential to improve world food security and the fact that there have, as yet, been no adverse results of their use in the food chain. Because of these worries and the mechanisms of politics, agricultural biotechnology has become the target of concerns about food safety (BSE, Foot & Mouth Disease), along with globalisation and the power of multinational companies. These concerns will, hopefully, be overcome by a more open and well informed dialogue between scientists, opinion leaders, educators and the public. If judiciously applied, genetically modified crops will help increase sustainability and the fight against hunger in the world. PMID- 12126802 TI - Attitudes toward biotechnology in the European Union. AB - Public attitudes toward biotechnology in the European Union have been characterized as negative using Eurobarometer data, but so far little attention has been paid to building a robust metric appropriate for emerging public opinion issues which combine high salience with very limited knowledge by the public. On the basis of the general literature about the formation and structure of attitudes and about public perceptions of science, this article presents a new metric and analysis: first, for estimating the level of awareness and knowledge of biotechnology in Europe; second, for assessing the stability and depth of these evaluative perceptions; and third, for exploring the roles of canonical socio-demographic variables, the knowledge variable and general attitudinal schemas for understanding the perceptions of both benefits and risks of biotech applications. The results show the importance of general value orientations or "worldviews" in shaping positive attitudes, and more of these general cognitive schemas should be measured in future research. The same multivariate model was unable to account for a significant percentage of the total variance in the perception of risks, suggesting that new measures are needed to tap this critical area in the acceptance of biotech in Europe. PMID- 12126804 TI - The biotech equipment and supplies sector in Europe-is it European? AB - Socio-economic research on biotechnology is dealing mainly with the sectors of biopharmaceuticals, agro-food or environmental technologies. In contrast, the equipment and supplies sector seems to be largely ignored. This is surprising because this sector provides important input in terms of technology and material for the development of biotechnology in general. Our comparative analysis of the sector in eight countries indicates that there exists no specific science base for the sector and that it is largely neglected by public research funding. Commercial activities are concentrated in countries with a large general science base in biotechnology and strong multinational pharmaceutical or chemical companies. There is a rather broad diversity in the way the sector has developed in the eight countries. Our data support the notion that national peculiarities seem dominant for explaining this picture. We anticipate growing business opportunities for European firms to step into large markets of equipment and supplies for functional genomics and protein analyses where Europe maintains a strong science base. PMID- 12126803 TI - The socio-economic landscape of biotechnology in Spain. A comparative study using the innovation system concept. AB - Biotechnology is becoming a crucial factor for the innovation strategies of the industrialised countries. Thus, the analysis of the sector is gaining relevance for the identification of the technological strength and potential of a country (or region) in a context of globalisation. A specific national case study may serve for more general comparative analyses. We have selected the case of Spain as illustrative of the complexity and differences existing in Europe. By using the analytical framework of the "national systems of innovation" concept, we have performed a multistep analysis of the biotechnology sector in Spain, focussing first on regions of Madrid and Cataluna which together account for more than 50% of the sector in Spain. The firms in both regions have followed a common strategy based on diversification and investment in R&D and innovation, so as to be able to compete in an international and competitive environment. There are, however, some interesting differences between the two subsectors; the one from Cataluna being more based on industrial traditions, and the one from Madrid characterised by the emergence of more specialised firms. The study has been extended to the remainder of Spanish firms for comparative purposes. The case of Spain is illustrative of the divergences existing in the biotechnology sector in Europe. A comparison is made with the structural and organisational characteristics of the biotechnology sector in several European countries. It shows that there is diversity in the pattern of commercialisation between countries and within regions of countries. Understanding these differences may assist the design of appropriate policies to promote the development of biotechnology in Europe. PMID- 12126805 TI - The pros and cons of human therapeutic cloning in the public debate. AB - Few issues linked to genetic research have raised as much controversial debate as the use of somatic cell nuclear transfer technology to create embryos specifically for stem cell research. Whereas European countries unanimously agree that reproductive cloning should be prohibited there is no agreement to be found on whether or not research into therapeutic cloning should be permitted. Since the UK took the lead and voted in favour of regulations allowing therapeutic cloning the public debate has intensified on the Continent. This debate reflects the wide spectrum of diverse religious and secular moralities that are prevalent in modern multicultural European democratic societies. Arguments range from putting forward strictly utilitarian views that weight the moral issues involved against the potential benefits that embryonic stem cell research may harbour to considering the embryo as a human being, endowed with human dignity and human rights from the moment of its creation, concluding that its use for research is unethical and should be strictly prohibited. Given the current state of dissension among the various European states, it is difficult to predict whether 'non-harmonisation' will prevail or whether in the long run 'harmonisation' of legislation that will allow stem cell research will evolve in the EU. PMID- 12126806 TI - Ethical dimensions of therapeutic human cloning. AB - Therapeutic human cloning has the potential significantly to reduce human suffering and enhance human happiness. This is the main ethical argument in its favour. The main ethical arguments against it centre on questions to do with the moral status of the human embryo. A subsidiary set of arguments arises from the connections between therapeutic human cloning and reproductive cloning. Most of the ethical questions concerning the status of the human embryo have long been examined in the context of abortion, though they are being re-examined in the context of genetic screening and embryo research. A consensus on such matters seems extremely unlikely to result in the near future. The current role of ethicists may not, therefore, be so much to attempt to produce a definitive answer to the question of the status of the human embryo at the very early developmental stages at which therapeutic human cloning would take place, but more to help clarify arguments and indicate the implications of particular approaches. That is what this paper seeks to do. PMID- 12126807 TI - Lessons we can learn from ecological biosafety research. AB - The last decade has seen an increasing number of biosafety related publications focusing on transgenic organisms. Recent extensive field studies suggest that harmful laboratory effects on non-target organisms rarely occur in the environment. Moreover, biosafety studies typically show no difference in hybridisation between genetically modified plants (GMPs) or non-GMPs and related wild species. Since risk is a product of both exposure and hazard, biosafety research should clearly not only target gene flow exposure but specifically concentrate on expected hazards emerging from successful transgene flow to wild relatives of GMPs. Generally, transgenic plants behave in an ecologically similar manner to non-GMPs if the modified trait confers a neutral advantage under environmental or experimental conditions. However, GMPs perform better than non GMPs if the new phenotype is challenged by conditions ecologically advantageous for the modified trait. Since biosafety research is a laborious process it will have to concentrate resources on thoughtful, thorough experiments, and target ecologically 'riskier' organisms. So far, we have no evidence that the use of GMPs contradicts sustainable agriculture and nature conservation per se. PMID- 12126808 TI - Assuring the safety of genetically modified (GM) foods: the importance of an holistic, integrative approach. AB - Genes change continuously by natural mutation and recombination enabling man to select and breed crops having the most desirable traits such as yield or flavour. Genetic modification (GM) is a recent development which allows specific genes to be identified, isolated, copied and inserted into other plants with a high level of specificity. The food safety considerations for GM crops are basically the same as those arising from conventionally bred crops, very few of which have been subject to any testing yet are generally regarded as being safe to eat. In contrast a rigorous safety testing paradigm has been developed for GM crops, which utilises a systematic, stepwise and holistic approach. The resultant science based process, focuses on a classical evaluation of the toxic potential of the introduced novel trait and the wholesomeness of the transformed crop. In addition, detailed consideration is given to the history and safe use of the parent crop as well as that of the gene donor. The overall safety evaluation is conducted under the concept known as substantial equivalence which is enshrined in all international crop biotechnology guidelines. This provides the framework for a comparative approach to identify the similarities and differences between the GM product and its comparator which has a known history of safe use. By building a detailed profile on each step in the transformation process, from parent to new crop, and by thoroughly evaluating the significance from a safety perspective, of any differences that may be detected, a very comprehensive matrix of information is constructed which enables the conclusion as to whether the GM crop, derived food or feed is as safe as its traditional counterpart. Using this approach in the evaluation of more than 50 GM crops which have been approved worldwide, the conclusion has been that foods and feeds derived from genetically modified crops are as safe and nutritious as those derived from traditional crops. The lack of any adverse effects resulting from the production and consumption of GM crops grown on more than 300 million cumulative acres over the last 5 years supports these safety conclusions. PMID- 12126809 TI - The application of biotechnological methods in authenticity testing. AB - By counterfeiting brand names in the food and drink industry as well as fraudulently labelling and selling low quality products as premium products, this sector of the industry has lost significant amounts of money and the consumer has been deceived. While it was difficult to establish certain types of fraud before the advent of modern biotechnology, DNA-based methods make an important contribution to protect high-quality brand names and protect the consumer. Several years ago, DNA technologies were considered as methods used in universities, primarily for research purpose, not so much for 'real-life' applications. However, this has changed and a number of laboratories have specialised in offering such services to the industry. This article will review DNA-based techniques commonly used for authenticity testing. PMID- 12126810 TI - Environmental biotechnology: the ongoing quest. AB - Environmental biotechnology, until now, has primarily focused on the development of technologies to treat aqueous, solid and gaseous wastes. At present, the basic knowledge on how biotechnology can handle these wastes has been acquired and the focus is now on the implementation of these processes as 'best available technology not entailing excessive costs' (BATNEEC) in the framework of strict and transparent environmental legislation. New environmental challenges continue to evolve, as it becomes clear that waste streams should be tackled in an overall holistic way. New technologies to reach this goal are currently under development. Novel aspects with respect to the domain of water treatment are, for example, the biomembrane reactor technology and the newly discovered processes to remove nitrogen by means of anaerobic ammonium oxidation. Also, most challenging is the continuing strive for re-use of treated wastewater. Indeed, water shortage is emerging in an increasing number of countries all over the world and necessitates the short cycling of water. Finally, biotechnology has a key role to play in the novel approaches to design wastewater treatment based on decentralised sanitation and reuse (DESAR). Solid waste is a major challenge worldwide. The implementation of anaerobic digestion to treat biowastes has become a grown-up technology. New approaches in which biotechnological processes are linked to physical processes, such as plasma technology, certainly deserve special attention for the coming decades. Soil and sediment clean up by means of biostimulation/remediation/augmentation is now well established. Certainly, a number of prospects need to be further explored, such as the use of special energy sources to stimulate in situ the microbial community and the seeding of knowledge to the in situ community by means of horizontal gene transfer mechanisms. A number of waste gases can be handled by biofilter systems. Biological treatment of wastegases is also evolving, inasmuch as that besides conventional chemical pollutants, now also highly problematic chemicals (even dioxins) can be dealt with through proper biotechnological approaches. A remarkable new potential is the use of well designed probiotics to upgrade aquaculture and together with conventional biological water treatment processes, to guarantee the overall water quality of this domain of food production. PMID- 12126811 TI - Heavy metals and arsenic uptake by wild vegetation in the Guadiamar river area after the toxic spill of the Aznalcollar mine. AB - On 25 April 1998, approximately 4.5 hm(3) of pyritic sludge, containing 5000 mg of As kg(-1) among other pollutants, was spilled into the Agrio and Guadiamar rivers and the surrounding agricultural areas (Aznalcollar, Seville, Southern Spain). Many trace metals such as Pb, Cu, Zn, Cd, Tl, Sb and As reached the Donana National Park, the largest wetland area in Europe, affecting soils, different plant and animal species. In order to recuperate the affected lands by employing plants capable of accumulating high levels of contaminants in shoots, periodical field surveys have been made to identify the metal-tolerant species that are spontaneously growing in the polluted soils, and are able to uptake one or various of the contaminants. Among the 99 different plant species studied, Anchusa azurea, Beta vulgaris, Chamaemelum fuscatum, Convolvulus arvensis, Cynodon dactylon, Diplotaxis virgata, Erodium aethiopicum, Lavatera cretica, Malva nicaeensis, Silybum marianum and, above all, Amaranthus blitoides highlight as the most promising to be used in the remediation of the affected area. PMID- 12126812 TI - Abstinence, monogamy, and sex. PMID- 12126813 TI - Haemopoietic stem-cell transplantation: improving immune reconstitution, avoiding graft-versus-host disease. PMID- 12126814 TI - Doping in sport. PMID- 12126815 TI - Seminal pharmaceutical trials: maintaining masking in analysis. PMID- 12126816 TI - Treatment for bronchiolitis: the story continues. PMID- 12126817 TI - Rapid reviews in The Lancet--and beyond. PMID- 12126818 TI - Long-term results with immediate androgen suppression and external irradiation in patients with locally advanced prostate cancer (an EORTC study): a phase III randomised trial. AB - BACKGROUND: We did a randomised phase III trial comparing external irradiation alone and external irradiation combined with an analogue of luteinising-hormone releasing hormone (LHRH) to investigate the added value of long-term androgen suppression in locally advanced prostate cancer. METHODS: Between 1987 and 1995, 415 patients were randomly assigned radiotherapy alone or radiotherapy plus immediate androgen suppression. Eligible patients had T1-2 tumours of WHO grade 3 or T3-4 N0-1 M0 tumours; the median age of participants was 71 years (range 51 80). In both treatment groups, 50 Gy radiation was delivered to the pelvis over 5 weeks, and 20 Gy over 2 weeks as a prostatic boost. Goserelin (3.6 mg subcutaneously every 4 weeks) was started on the first day of irradiation and continued for 3 years; cyproterone acetate (150 mg orally) was given for 1 month starting 1 week before the first goserelin injection. The primary endpoint was clinical disease-free survival. Analyses were by intention to treat. FINDINGS: 412 patients had evaluable data, with median follow-up of 66 months (range 1 126). 5-year clinical disease-free survival was 40% (95% CI 32-48) in the radiotherapy-alone group and 74% (67-81) in the combined-treatment group (p=0.0001). 5-year overall survival was 62% (52-72) and 78% (72-84), respectively (p=0.0002) and 5-year specific survival 79% (72-86) and 94% (90-98). INTERPRETATION: Immediate androgen suppression with an LHRH analogue given during and for 3 years after external irradiation improves disease-free and overall survival of patients with locally advanced prostate cancer. PMID- 12126819 TI - Aspirin and coumadin after acute coronary syndromes (the ASPECT-2 study): a randomised controlled trial. AB - BACKGROUND: Antiplatelet treatment with aspirin and oral anticoagulants reduces recurrence of ischaemic events after myocardial infarction. We aimed to investigate which of these drugs is more effective in the long term after acute coronary events, and whether the combination of aspirin and oral anticoagulants offers greater benefit than either of these agents alone, without excessive risk of bleeding. METHODS: In a randomised open-label trial in 53 sites, we randomly assigned 999 patients to low-dose aspirin, high-intensity oral anticoagulation, or combined low-dose aspirin and moderate intensity oral anticoagulation. Patients were followed up for a maximum of 26 months. The primary composite endpoint was first occurrence of myocardial infarction, stroke, or death. FINDINGS: The primary endpoint was reached in 31 (9%) of 336 patients on aspirin, in 17 (5%) of 325 on anticoagulants (hazard ratio 0.55 [95% CI 0.30-1.00], p=0.0479), and in 16 (5%) of 332 on combination therapy (0.50 [0.27-0.92], p=0.03). Major bleeding was recorded in three (1%) patients on aspirin, three (1%) on anticoagulants (1.03 [0.21-5.08], p=1.0), and seven (2%) on combination therapy (2.35 [0.61-9.10], p=0.2). Frequency of minor bleeding was 5%, 8% (1.68 [0.92-3.07], p=0.20), and 15% (3.13 [1.82-5.37], p=<0.0001), in the three groups, respectively. 164 patients permanently discontinued the study drug. Analyses were done by intention to treat. INTERPRETATION: In patients recently admitted with acute coronary events, treatment with high-intensity oral anticoagulants or aspirin with medium-intensity oral anticoagulants was more effective than aspirin on its own in reduction of subsequent cardiovascular events and death. PMID- 12126820 TI - Assessment of guidelines for good practice in psychosocial care of mothers after stillbirth: a cohort study. AB - BACKGROUND: Most maternity units have good practice protocols, advising that after stillbirth parents should be encouraged to see and hold their dead infant. Our aim was to assess whether adherence to these protocols is associated with measurably beneficial effects on the psychological health of mother and next-born child. This study forms part of a wider case-control study of the psychological effects of stillbirth. METHODS: We identified 65 women in the pregnancy after stillbirth, and enrolled matched controls for 60 of them. Outcome measures included depression, anxiety, and post-traumatic-stress disorder (PTSD) in pregnancy and 1 year after the next birth, and disorganised attachment behaviour in the next-born infant. Comparison variables included seeing and holding the stillborn infant, having a funeral, and keeping mementoes. FINDINGS: Behaviours that promote contact with the stillborn infant were associated with worse outcome. Women who had held their stillborn infant were more depressed than those who only saw the infant, while those who did not see the infant were least likely to be depressed (13 of 33, 39%, vs three of 14, 21%, vs one of 17, 6%; p=0.03). Women who had seen their stillborn infant had greater anxiety (p=0.02) and higher symptoms of PTSD than those who had not (p=0.02), and their next-born infants were more likely to show disorganised attachment behaviour (18 of 43, 42%, vs one of 12, 8%, p=0 x 04). Having a funeral and keeping mementoes were not associated with further adverse outcomes, but small numbers limited interpretation. INTERPRETATION: Our findings do not support good-practice guidelines, which state that failure to see and hold the dead child could have adverse effects on parents' mourning. PMID- 12126821 TI - Prognosis of HIV-1-infected patients starting highly active antiretroviral therapy: a collaborative analysis of prospective studies. AB - BACKGROUND: Insufficient data are available from single cohort studies to allow estimation of the prognosis of HIV-1 infected, treatment-naive patients who start highly active antiretroviral therapy (HAART). The ART Cohort Collaboration, which includes 13 cohort studies from Europe and North America, was established to fill this knowledge gap. METHODS: We analysed data on 12,574 adult patients starting HAART with a combination of at least three drugs. Data were analysed by intention to-continue-treatment, ignoring treatment changes and interruptions. We considered progression to a combined endpoint of a new AIDS-defining disease or death, and to death alone. The prognostic model that generalised best was a Weibull model, stratified by baseline CD4 cell count and transmission group. FINDINGS During 24,310 person-years of follow up, 1094 patients developed AIDS or died and 344 patients died. Baseline CD4 cell count was strongly associated with the probability of progression to AIDS or death: compared with patients starting HAART with less than 50 CD4 cells/microL, adjusted hazard ratios were 0.74 (95% CI 0.62-0.89) for 50-99 cells/microL, 0.52 (0.44-0.63) for 100-199 cells/microL, 0.24 (0.20-0.30) for 200-349 cells/microL, and 0.18 (0.14-0.22) for 350 or more CD4 cells/microL. Baseline HIV-1 viral load was associated with a higher probability of progression only if 100,000 copies/microL or above. Other independent predictors of poorer outcome were advanced age, infection through injection-drug use, and a previous diagnosis of AIDS. The probability of progression to AIDS or death at 3 years ranged from 3.4% (2.8-4.1) in patients in the lowest-risk stratum for each prognostic variable, to 50% (43-58) in patients in the highest-risk strata. INTERPRETATION: The CD4 cell count at initiation was the dominant prognostic factor in patients starting HAART. Our findings have important implications for clinical management and should be taken into account in future treatment guidelines. PMID- 12126822 TI - Unilateral anhidrosis of the leg. PMID- 12126823 TI - Immune reconstitution without graft-versus-host disease after haemopoietic stem cell transplantation: a phase 1/2 study. AB - BACKGROUND: Allogeneic haemopoietic stem-cell transplantation (HSCT) is the treatment of choice for many haematological malignancies and inherited disorders. When stem cells for transplantation come from a human leucocyte antigen matched unrelated donor, or from a partly mismatched related donor, ex-vivo T-cell depletion of the graft can prevent development of graft-versus-host disease, but lead in turn to a delay in immune reconstitution and a concordant increase in incidence of opportunistic infections and leukaemic relapses. We aimed to infuse T cells selectively depleted in allogeneic T cells that cause graft-versus-host disease using an ex-vivo procedure designed to eliminate alloactivated donor T cells, with an immunotoxin that reacts with a cell surface activation antigen, CD25. METHODS: We did a phase 1/2 study, in which 1-8 x 10(5) allodepleted T cells/kg were infused between days 15 and 47 into 15 paediatric patients who had acquired or congenital haemopoietic disorders and who received HSCT on day 0. Occurrence of graft-versus-host disease and time to immune reconstitution were assessed. No treatment for graft-versus-host disease was given. FINDINGS: Less than 1% residual anti-host alloreactivity was recorded in 12 of 16 procedures. Other immune responses were preserved by the allodepletion procedure in 12 cases. No cases of severe (greater than grade II) graft-versus-host disease arose. Evidence for early T-cell expansion was shown in three patients with continuing viral infections. Specific antiviral responses, such as strong cytolytic activity, were noted. INTERPRETATION: Our results show that ex-vivo selective depletion of T cells that cause graft-versus-host disease is efficient and feasible, even in haploidentical settings. PMID- 12126824 TI - All that glitters is not gold. PMID- 12126825 TI - Plummeting lenses in the TB clinic. PMID- 12126826 TI - Incidence of Creutzfeldt-Jakob disease in Switzerland. AB - The incidence of Creutzfeldt-Jakob disease (CJD) in Switzerland increased two fold in 2001, and figures from the first quarter of 2002 indicate that it continues to rise. Neither age at onset nor duration of disease were different from previous years. Genetic analysis of the 27 reported cases revealed only one disease-associated mutation in the prion gene. None of the recognised risk factors for acquired CJD were reported on the official notification forms. Glycotype profiling, histopathology, and immunohistochemistry indicate that none of the cases fulfilled the definition of variant CJD, which is thought to be caused by bovine prions. Several scenarios could account for the increase in CJD, including improved reporting, iatrogenic transmission, and transmission of a prion zoonosis. PMID- 12126827 TI - Inhaled ethyl nitrite gas for persistent pulmonary hypertension of the newborn. AB - Inhaled nitric oxide is used to alleviate pulmonary hypertension and hypoxaemia, but generates toxic free radicals and oxides of nitrogen (NO(x)), which can cause rebound-hypoxia and additional pulmonary and other morbidity. To address these problems, we assessed the efficacy of inhaled O-nitrosoethanol gas (ENO) as a novel alternative means of providing nitric oxide bioactivity in the treatment of persistent pulmonary hypertension of newborns. We administered ENO over 4 h to seven neonates who required assisted ventilation, and who had an oxygenation index of 25 or more. ENO was then shut off for 15 min before start of treatment with inhaled nitric oxide. Our results show that ENO produced sustained improvements in postductal arterial oxygenation and systemic haemodynamics, which were maintained during the off-drug observation period. Increases in methaemoglobinaemia were modest and toxic NO(x) were not detected. Thus, ENO can improve oxygenation and systemic haemodynamics in neonates, and seems to reduce rebound hypoxaemia and production of toxic byproducts. PMID- 12126829 TI - Experts predict global devastation due to HIV/AIDS. PMID- 12126828 TI - Number of boys born to men exposed to polychlorinated byphenyls. AB - We studied the sex of children born to individuals involved in the Yucheng oil disaster, Taiwan, who were exposed to polychlorinated byphenyls (PCBs) after an oil contamination accident in 1979. Men exposed to PCBs before age 20 years had a lower chance of having a baby boy than did age-matched and neighbourhood-matched controls (odds ratio 0.65, 95% CI 0.45-0.93). The male-to-female sex ratio of children born to men exposed to PCBs after age 20 years, however, approached that seen in controls (0.90, 0.59-1.35). We noted no significant difference in the birth ratio of infants born to exposed and unexposed mothers (0.93, 0.77-1.12). Our findings suggest that paternal exposure to PCBs before age 20 years affects the sex of a subsequently born child. PMID- 12126832 TI - Can flossing teeth foil heart disease? PMID- 12126833 TI - Indian government reduces charges against Union Carbide. PMID- 12126835 TI - New Serbian health minister appointed at last. PMID- 12126837 TI - US health chiefs balk at homeland security plan. PMID- 12126838 TI - Female subfertility. AB - With an average monthly fecundity rate of only 20%, human beings are not fertile mammals. 10-15% of couples have difficulties conceiving, or conceiving the number of children they want, and seek specialist fertility care at least once during their reproductive lifetime. Dependent on the two main factors that determine subfertility, duration of childlessness and age of the woman, three questions need to be addressed before treatment is offered. Is it time to start the routine fertility investigation?--ie, has sufficient exposure to the chance of conception taken place? Are cost-effective, safe, and reliable treatments available for the disorder diagnosed? And, should the couple be referred straightaway for assisted reproduction? PMID- 12126839 TI - Minimal residual disease evaluation in acute myeloid leukaemia. PMID- 12126840 TI - Alzheimer's disease. PMID- 12126841 TI - Communication of risk: choice, consent, and trust. PMID- 12126842 TI - Proteomic patterns in serum and identification of ovarian cancer. PMID- 12126843 TI - Proteomic patterns in serum and identification of ovarian cancer. PMID- 12126844 TI - Proteomic patterns in serum and identification of ovarian cancer. PMID- 12126845 TI - Proteomic patterns in serum and identification of ovarian cancer. PMID- 12126847 TI - Screening mammography: but how do women decide? PMID- 12126848 TI - Effect of supplementation with folic-acid on relation between plasma homocysteine, folate, and vitamin B12. PMID- 12126850 TI - Effect of supplementation with folic-acid on relation between plasma homocysteine, folate, and vitamin B12. PMID- 12126851 TI - Gene-expression profiling and identification of patients at high risk of breast cancer. PMID- 12126852 TI - Gene-expression profiling and identification of patients at high risk of breast cancer. PMID- 12126855 TI - GAVI, the first steps: lessons for the Global Fund. PMID- 12126854 TI - Detection of a previously uncommon salmonella phage in tourists returning from Europe. PMID- 12126859 TI - Condoms and contraception. PMID- 12126856 TI - GAVI, the first steps: lessons for the Global Fund. PMID- 12126865 TI - 24-Hour motor activity after treatment with imipramine or fluvoxamine in major depressive disorder. AB - Psychomotor dysfunction in depression is related to alterations in the 24-h pattern of motor activity. After antidepressant treatment the diurnal pattern may be changed due to improvement of clinical state or pharmacological actions. The purpose of this study was to evaluate in 52 depressed in-patients the effects of imipramine (tricyclic antidepressant) and fluvoxamine (SSRI) on the 24-h motor activity. Motor activity was monitored by wrist-actigraphy during a medication free period and after 4 weeks of treatment. Clinical improvement was not different after imipramine or fluvoxamine treatment. The Hamilton depression score decreased in patients treated with imipramine, as well as in patients treated with fluvoxamine. The clinical retardation score was also reduced in both treatment groups. However, patients treated with imipramine showed higher motor activity levels during the wake period in comparison to the medication-free period, and more fragmentation of motor activity during sleep. Treatment with fluvoxamine did not result in alterations in the 24-h pattern of motor activity. The improvement of depressive mood and retardation seems to play a minor role in the change of the pattern of motor activity after imipramine. PMID- 12126866 TI - Valproate acidifies hippocampal CA3-neurons--a novel mode of action. AB - Various hypotheses try to explain the anticonvulsive and mood stabilizing effects of valproate. Among them, amplification of GABAergic inhibition and reduction of membrane excitability is favored. Here we show that superfusion with 0.1-1 mM valproate induced a moderate intracellular acidification of BCECF-AM-loaded CA3 neurons (hippocampal slices, guinea pig) which was measured as the difference between intracellular pH before (baseline pH(i)) and during valproate treatment (deltapH(i)). In two groups of neurons treated with 1 mM and 0.1-0.5 mM, deltapH(i) values amounted to 0.20 +/- 0.10 and 0.10 +/- 0.04 (deltapH(i) +/- S.D.), respectively, suggesting a dependence on the used valproate-concentration. DeltapH(i) did not correlate with the baseline pH(i). Furthermore, the acidification seems to be independent from an activation of postsynaptic GABA-A receptors, as it was not influenced by 0.1 mM picrotoxin. Since our previous studies clearly demonstrated a reduction of membrane excitability during moderate intracellular acidification, we suggest that the valproate-mediated intracellular acidification may substantially contribute to its anticonvulsive and mood stabilizing properties. PMID- 12126867 TI - Neural degeneration following chronic stimulant abuse reveals a weak link in brain, fasciculus retroflexus, implying the loss of forebrain control circuitry. AB - There is increasing evidence that the fasciculus retroflexus (FR) represents a 'weak link' following the continuous administration of drugs of abuse. A variety of drugs which predominantly potentiate dopamine, including D-amphetamine, methamphetamine, MDMA, cocaine, and cathinone, all induce degeneration in axons from lateral habenula, through the sheath of FR, to midbrain cells such as SN, VTA, and raphe. For some drugs, such as cocaine, this is virtually the only degeneration induced in brain. Continuous nicotine also selectively induces degeneration in FR, but in the other half of the tract, i.e. in axons from medial habenula through the core of the tract to interpeduncular nucleus. This phylogenetically primitive tract carries much of the negative feedback from forebrain back onto midbrain reward cells, and the finding that these descending control pathways are compromised following simulated drug binges has implications for theories of drug addiction but also psychosis in general. PMID- 12126868 TI - Functional catechol-O-methyltransferase gene polymorphism and susceptibility to schizophrenia. AB - Genetic polymorphism of catechol-O-methyltransferase (COMT), involved in the degradation of catecholamine neurotransmitters, has been investigated as a candidate for modifier of susceptibility to development of schizophrenia. To address this issue further, we carried out a study in Korean schizophrenic patients and controls. The study population consisted of 103 Korean inpatients diagnosed as schizophrenic and their 103 age and sex matched controls. The patients were divided into two groups on the basis of history of aggressive behavior, family history of schizophrenia and related disorders, and age at onset of the disease. The COMT genotypes were determined by a PCR based method. No statistically significant overall associations between the COMT genotypes and risk of schizophrenia were observed. However, subjects with at least one low activity associated COMT-L allele showed a tendency of elevated risk for schizophrenia (OR=1.7, 95% CI=0.9-3.1) compared with those homozygous for the high activity associated COMT-H alleles. Moreover, when cases were stratified by family history of schizophrenia, a significant combined effect was seen: the cases with concurrent family history of schizophrenia and the COMT-L allele containing genotypes had an almost 4-fold (OR=3.9, 95% CI=1.1-14.3) higher risk of schizophrenia compared to controls with the COMT-HH genotypes. Future studies with larger sample sizes are, however, needed to confirm this novel finding. PMID- 12126869 TI - Amisulpride improves depressive symptoms in acute exacerbations of schizophrenia: comparison with haloperidol and risperidone. AB - The Brief Psychiatric Rating Scale (BPRS) anxiety/depression subscore has been used to assess affective symptoms in three studies (n = 612) comparing amisulpride (400-800 mg/day, n = 339) with haloperidol (15-20 mg/day, n = 160) and risperidone (8 mg/day, n = 113) in the treatment of acute exacerbations of schizophrenia. At endpoint, the mean improvement in the anxiety/depression subscore showed a significant (P = 0.011) difference in favour of amisulpride (5.6+/-6.1) compared with haloperidol (4.4+/-5.5) and risperidone (3.7+/-4.7). Amisulpride provided a significantly greater improvement compared both to haloperidol and risperidone in more severely depressed patients (BPRS anxiety/depression subscore > or = 16 at baseline, P = 0.001). This significant advantage in favour of amisulpride is seen from the 2nd week of treatment. PMID- 12126870 TI - Influence of agents affecting voltage-dependent calcium channels and dantrolene on the anticonvulsant action of the AMPA/kainate receptor antagonist LY 300164 in mice. AB - It was previously documented that calcium (Ca(2+)) channel inhibitors intensified the protective effects of conventional antiepileptics against electroconvulsions in mice. The aim of this study was to evaluate the effects of Ca(2+) channel inhibitors (nifedipine, nicardipine and flunarizine) on the anticonvulsant action of the new AMPA/kainate receptor antagonist, 7-acetyl-3-(4-aminophenyl)-8,9 dihydro-8-methyl-7H-1,3-dioxazolo[4,5-h][2,3]-benzodiazepine (LY 300164), against maximal electroshock (MES)-induced seizures in mice. Dantrolene (an inhibitor of Ca(2+)release from intracellular stores) was also included. Nifedipine (30 mg/kg) and flunarizine (15 mg/kg) raised the threshold for electroconvulsions, being ineffective at lower doses. Nicardipine (up to 30 mg/kg) and dantrolene (up to 20 mg/kg) did not affect this parameter. Flunarizine (10 mg/kg), nicardipine (20 mg/kg) and dantrolene (20 mg/kg) potentiated the efficacy of LY 300164 against MES. However, nicardipine (at 20 mg/kg) raised the free plasma concentration of LY 300164. Nifedipine (30 mg/kg), given even in a dose raising the electroconvulsive threshold, did not significantly alter the protective effect of LY 300164 against MES. Furthermore, the Ca(2+) channel agonist-BAY k-8644 (at 5 mg/kg) did not influence the protection offered by LY 300164 against MES. Finally, this Ca(2+) channel activator did not affect the enhanced efficacy of LY 300164 by Ca(2+) channel modulators. The only exception was the combination of LY 300164 with flunarizine. Combined treatment with LY 300164 and dantrolene (20 mg/kg), compared to LY 300164 alone, resulted in an impairment of motor performance in mice. Ca(2+) channel inhibitors were without effect upon this parameter evaluated in the chimney test. As shown in the passive avoidance task, LY 300164 alone (at its ED(50)) or combined with agents affecting neuronal Ca(2+) concentration did not disturb long-term memory. The present results suggest that agents preventing influx of Ca(2+) ions into neurons may enhance the protective action of LY 300164. PMID- 12126871 TI - Molsidomine potentiates the protective activity of GYKI 52466, a non-NMDA antagonist, MK-801, a non-competitive NMDA antagonist, and riluzole against electroconvulsions in mice. AB - The influence of molsidomine, a donor of nitric oxide (NO), L-arginine, a substrate for NO synthesis, and N(G)-nitro-L-arginine (NNA), an inhibitor of NO synthase, on the protective activity of CGP 40116, GYKI 52466, MK-801, and riluzole against electroconvulsions was studied in mice. Molsidomine (100 mg kg( 1); i.p.) potentiated the protective activity of GYKI 52466, MK-801, and riluzole but did not influence the protection offered by CGP 40116. In contrast to molsidomine, L-arginine (500 mg kg(-1); i.p.) did not impair the protective activity of any anticonvulsant. In a dose of 40 mg kg(-1), NNA administered i.p. did not affect the protection offered by any excitatory amino acid antagonists and riluzole. Combinations of molsidomine with either GYKI 52466 or MK-801 as well as riluzole did not cause a memory deficit in the passive avoidance task. However, the combined treatment of molsidomine with these anticonvulsants resulted in a motor impairment quantified by the chimney test. The lack of effect of L-arginine and NNA on the protective activity of excitatory amino acid antagonists suggests that molsidomine-evoked alterations in the protection provided by some excitatory amino acid antagonists against electroconvulsions are independent of the NO pathway. PMID- 12126872 TI - Effects on drug disposition, brain monoamines and behavior after chronic treatment with the antidepressant venlafaxine in rats with experimental hepatic encephalopathy. AB - Patients with chronic hepatic encephalopathy (HE) may present affective symptoms and antidepressant drug treatment in this condition is not uncommon. The present microdialysis study investigated treatment with the chronic antidepressant venlafaxine (VEN) in experimental HE with regard to tentative changes in pharmacokinetic and/or pharmacodynamic parameters. Three weeks after portacaval shunt (PCS) or sham operation in rats, VEN (10 mg/kg daily) was administered by implanted osmotic minipumps. VEN treatment for 14 days resulted in higher concentrations of VEN in PCS rats than in sham controls in serum and brain compartments, and the VEN levels in serum and brain were strongly inter correlated. The serum N-desmethylvenlafaxine concentration did not differ between the groups, but correlated with the serum VEN levels. The other VEN metabolites were below the quantification limits. VEN treatment for 9-12 days significantly stimulated locomotion and rearing in the open field in sham controls, but failed to do so in the PCS rats. The concentrations of noradrenaline, dopamine, 5-HT, and 5-HIAA in neocortical dialysates were higher in PCS than in sham rats after 14 days of VEN treatment, but the elevations reached statistical significance only in the case of dopamine and 5-HIAA. In summary, there were significant pharmacokinetic and pharmacodynamic alterations in rats with experimental HE as compared to controls. The described experimental HE model may be useful for continued pharmacokinetic/pharmacodynamic interaction studies to unravel the pathophysiological consequences of HE on the CNS. PMID- 12126873 TI - Painful ejaculation and urinary hesitancy in association with antidepressant therapy: relief with tamsulosin. AB - Painful ejaculation has been reported in association with a variety of antidepressants such as the tricyclic antidepressants (TCAs, e.g. clomipramine, imipramine, desipramine, protriptyline, amoxapine), the selective serotonin reuptake inhibitors (SSRIs, e.g. fluoxetine), venlafaxine and the MAOIs. Apart from lowering the dose and changing the antidepressant, no strategies are available to treat this side effect. In this paper, painful ejaculation following the administration of reboxetine is described in two patients. Both patients were treated concomitantly with the selective alpha(1A)-adrenoceptor antagonist, tamsulosin. A re-challenge was performed in one patient. The Hamilton Depression Rating Scale (HAM-D), the American Urological Association symptom index, a (dis)satisfaction item score and the Udvalg for Kliniske Undersoegelser (UKU-side effect rating scale) were used to assess the treatment. Tamsulosin rapidly and completely resolved the painful ejaculation and urinary hesitancy in both patients. A re-challenge in one patient resulted in a prompt reappearance of both side effects. Tamsulosin resolved the problem of painful ejaculation in these patients; however, larger studies are needed to confirm these results. PMID- 12126874 TI - Increased platelet vesicular monoamine transporter density in adult schizophrenia patients. AB - Vesicular monoamine transporter (VMAT2) plays a major role in the synaptic accumulation and release of monoamines. In the present study, we assessed high affinity [(3)H]dihydrotetrabenazine binding to platelet VMAT2 in a group of treated (n = 9) and untreated (n = 4) patients with schizophrenia and age- and sex-matched controls. Significantly elevated platelet VMAT2 density (B(max)) (53%, P<0.0001) was observed in the mixed population of schizophrenia patients. The affinity of the ligand (K(d)) to platelet VMAT2 was similar in both groups. The increased platelet VMAT2 density may indicate a schizophrenia-related hyperactivity of the monoaminergic system or an adaptive response to chronic drug treatment. PMID- 12126875 TI - A preliminary study of buspirone stimulated prolactin release in generalised social phobia: evidence for enhanced serotonergic responsivity? AB - Serotonergic dysfunction has been postulated to play a role in the aetiology of social phobia. Buspirone, which is a partial agonist at 5HT1A receptors, increases prolactin release and may be used as a probe to examine serotonergic responses, dysfunction of which may be relevant to the pathophysiology of social phobia. We compared buspirone stimulated prolactin release in 14 patients with generalised social phobia and 14 healthy controls. Buspirone 30 mg was administered orally and prolactin release over 180 min was monitored. Overall, patients with generalised social phobia had greater prolactin release in response to buspirone challenge than healthy comparison subjects. There was no correlation between prolactin response and measures of severity of social phobia. Patients with generalised social phobia had enhanced central serotonergic responses, an abnormality shared with some other anxiety disorders and which may be of aetiological significance. PMID- 12126876 TI - Differential effects of repeated imipramine on hippocampal responsiveness to adenosine and serotonin. AB - Imipramine-induced enhancement of the inhibitory action of 5-HT(lA) receptor activation in hippocampal pyramidal neurons has been attributed to alterations in the transduction mechanism that involves G protein-dependent opening of K(+) channels. Postsynaptic 5-HT(lA) and adenosine Al receptors may share that transduction pathway. We investigated the influence of repeated imipramine administration on 5-HT(lA) and adenosine A1 receptor-mediated effects in rat hippocampal slices. Repeated imipramine selectively enhanced the postsynaptic effects of 5-HT(1A) receptor activation, including hyperpolarization and reduction of input resistance of neurons and reduction of the population spike amplitude. In contrast, after imipramine treatment only the presynaptic effect of adenosine receptor agonists, a decrease of the field excitatory postsynaptic potential, was enhanced. The data demonstrate that alterations in the presumed common transduction mechanism that was postulated for the 5-HT(lA) and adenosine A1 receptor-mediated activation of K(+) channels are not involved in the effect of repeated imipramine administration. PMID- 12126877 TI - A 6-hydroxydopamine lesion of the mesostriatal dopamine system decreases the expression of corticotropin releasing hormone and neurotensin mRNAs in the amygdala and bed nucleus of the stria terminalis. AB - The mesostriatal dopamine (DA) system is known to play a vital role in extrapyramidal motor responses, and animals with a unilateral 6-hydroxydopamine (6-OHDA) lesion of this system have proved useful in studying the behavioral and neurobiological effects of DA depletion. Less is known about the role of this system in modulating emotional responses, although a number of lines of evidence suggest that dopamine influences emotional behavior. During the course of a study involving rats that had a unilateral 6-OHDA lesion, we discovered a hemispheric asymmetry in the levels of corticotropin releasing hormone (CRH) mRNA in the central nucleus of the amygdala (CEA). The present study was performed in order to determine (1) if the lesion resulted in a decrease in CRH mRNA, and/or if there was upregulation on the intact side, (2) if a similar imbalance in CRH mRNA was observed in other brain regions and (3) if levels of other neuropeptide mRNAs were affected by the lesion. Adult male Sprague-Dawley rats were left unoperated or were pretreated with desipramine and then injected unilaterally with 6-OHDA into the medial forebrain bundle to lesion the ascending mesostriatal DA neurons. Animals were killed 15-31 days following surgery and brain sections processed for CRH, neurotensin and enkephalin mRNAs by in situ hybridization. Levels of CRH and neurotensin mRNAs were decreased on the lesioned side in the CEA and oval nucleus of the BST (BSTov) relative to the intact side and to unoperated controls. Levels of enkephalin mRNA in these regions were not affected by the lesion. These effects appeared specific, because the lesion did not alter CRH mRNA expression in the ventral BST, paraventricular nucleus of the hypothalamus or cortex or neurotensin mRNA expression in the CA1 region of the hippocampus. In contrast, and consistent with previous reports, levels of neurotensin and enkephalin mRNAs were upregulated on the lesioned side of the striatum. This study provides evidence that the mesostriatal DA system regulates CRH and neurotensin mRNA in the BSTov and CEA, suggesting that dopamine may be an important modulator of CRH and neurotensin function within these nuclei. Although the precise mechanisms are not clear, and the involvement of noradrenergic systems cannot be precluded, data are consistent with the idea that dopamine, released in response to a stressful experience for example, interacts with CRH and neurotensin in the extended amygdala to affect emotional responsiveness. PMID- 12126878 TI - Survival motor neuron protein in the nucleolus of mammalian neurons. AB - Spinal muscular atrophy (SMA) is an inherited motor neuron disease caused by mutations in the survival motor neuron gene (SMN1). While it has been shown that the SMN protein is involved in spliceosome biogenesis and pre-mRNA splicing, there is increasing evidence indicating that SMN may also perform important functions in the nucleolus. We demonstrate here through the use of a previously characterized polyclonal anti-SMN antibody, abSMN, that the SMN protein shows a striking colocalization with the nucleolar protein, fibrillarin, in both nucleoli and Cajal bodies/gems of primary neurons. Immunoblot analysis with antifibrillarin and two different anti-SMN antibodies reveals that SMN and fibrillarin also cofractionate in the insoluble protein fraction of cultured cell lysates. Immunoprecipitation experiments using whole cell extracts of HeLa cells and cultured neurons revealed that abSMN coprecipitated small amounts of the U3 small nucleolar RNA (snoRNA) previously shown to be associated with fibrillarin in vivo. These studies raise the possibility that SMN may serve a function in rRNA maturation/ribosome synthesis similar to its role in spliceosome biogenesis. PMID- 12126879 TI - Comparison of spontaneous and evoked epileptiform activity in three in vitro epilepsy models. AB - Rat neocortical slices express spontaneous epileptiform activity after incubation with GABA(A) receptor blocker bicuculline (BIC, 20 microM), with potassium channel blocker 4-aminopyridine (4-AP, 50 microM) or in Mg(2+)-free medium (LMG). Various parameters of spontaneous and evoked epileptiform discharges and their pharmacological sensitivity were analysed using extracellular field potential recordings in this comparative in vitro study. All types of convulsant solution induced spontaneous epileptiform activity, however, the analysed parameters showed that characteristics of epileptiform discharges are rather different in the three models. The longest duration of discharges was recorded in LMG, while the highest frequency of spontaneous events was detected in 4-AP. The epileptiform field responses elicited by electrical stimulation appeared in an all-or-none manner in BIC. On the contrary, in 4-AP and in LMG the amplitude of the responses increased gradually with increasing stimulation intensities. The NMDA receptor antagonist D,L-2-amino-5-phosphonovaleric acid (APV, 25 microM) abolished the LMG induced spontaneous epileptiform activity and significantly reduced the frequency of the epileptiform discharges in BIC and 4-AP. Blocking the AMPA type of glutamate transmission with 1-(aminophenyl)-4-methyl-7,8 methylenedioxy-5H-2,3-benzodiazepine (GYKI 52466, 40 microM) rapidly abolished BIC-induced spontaneous epileptiform activity and caused a significant decrease in the frequency of 4-AP induced spontaneous epileptiform discharges. However, it had only a weak effect on the LMG-induced epileptiform activity. We conclude that the contribution of NMDA and AMPA types of glutamate receptors to the development and maintenance of epileptiform activity in cortical cell assemblies is different in the three models. There are significant alterations in contribution of NMDA and AMPA types of glutamate receptors to the above-mentioned processes in the different convulsants. In BIC the synchronisation is mainly due to the altered network properties, namely inhibition is reduced in the local circuits. Although inhibition is reduced in the local circuits, the AMPA receptor antagonist relatively easily blocked the synchronised excitation. In 4-AP, and especially in LMG, changes in the membrane characteristics of neurones play a crucial role in the increased excitability. In this case the AMPA antagonist was less effective. PMID- 12126880 TI - Calretinin and calbindin D-28k, but not parvalbumin protect against glutamate induced delayed excitotoxicity in transfected N18-RE 105 neuroblastoma-retina hybrid cells. AB - Excitotoxic effects leading to neuronal cell degeneration are often accompanied by a prolonged increase in the intracellular level of Ca(2+) ions and L-glutamate induced toxicity is assumed to be mediated via a Ca(2+)-dependent mechanism. Due to their buffering properties, EF-hand Ca(2+)-binding proteins (CaBPs) can affect intracellular Ca(2+) homeostasis and a neuroprotective role has been attributed to some of the family members including calretinin, calbindin D-28k and parvalbumin. We have stably transfected N18-RE 105 neuroblastoma-retina hybrid cells with the cDNAs for the three CaBPs and investigated the effect of these proteins on the L-glutamate-induced, Ca(2+)-dependent cytotoxicity. Several clones for each CaBP were selected according to immunocytochemical staining and characterization of the overexpressed proteins by Western blot analysis. In calretinin- and parvalbumin-expressing clones, expression levels were quantitatively determined by ELISA techniques. Cytotoxicity of transfected clones was quantified by measurement of the activity of lactate dehydrogenase (LDH) that was released into the medium after L-glutamate (10 mM) exposure as a result of necrotic cell death. In untransfected and parvalbumin-transfected cells, LDH released into the medium progressively increased (starting from the 20th hour) reaching maximum levels after 28-30 h of glutamate application. In contrast, LDH release in both, calretinin and calbindin D-28k-transfected clones, was not significantly different from unstimulated transfected or untransfected cells over the same period of time. The results indicate that the 'fast' Ca(2+)-buffers calretinin and calbindin D-28k, but not the 'slow' buffer parvalbumin can protect N18-RE 105 cells from this type of Ca(2+)-dependent L-glutamate-induced delayed cytotoxicity. PMID- 12126881 TI - Factors regulating the influence of melatonin on hippocampal evoked potentials: comparative studies on different strains of mice. AB - Factors regulating the influence of melatonin on the hippocampal glutamergic system in mouse hippocampal slices were evaluated. The sensitivity of hippocampal pyramidal neurons to melatonin (Sigma) was highest at 2 h following slice preparation and then declined with time. This pattern of sensitivity to melatonin correlated well with a reduced binding of melatonin to its receptors. The slices obtained from older animals remained sensitive to melatonin through the entire incubation period. Most of the experiments evaluating the influence of melatonin on hippocampal evoked potentials were performed within 2 h following slice preparation. The effect of melatonin was biphasic: an initial depression of the potential was followed by a recovery/amplification phase. The recovery phase was not a result of melatonin decomposition. The effect of melatonin was similar in three different strains of mice tested: CD-1, C57J/B6, and Swiss Webster. While the melatonin from another vendor (Regis) gave similar results, it was effective at much lower concentrations. In slices obtained from CD-1 light-deprived mice, the sensitivity to melatonin was significantly reduced. Thus, it appears that melatonin may control the hippocampal glutamergic system in a complex manner, which may be regulated by the circadian rhythm. This may influence memory formation in the hippocampus. PMID- 12126882 TI - Glutamine-, glutamine synthetase-, glutamate dehydrogenase- and pyruvate carboxylase-immunoreactivities in the rat dorsal root ganglion and peripheral nerve. AB - Supporting glial cells of the peripheral nervous system include satellite cells of dorsal root ganglia and Schwann cells of peripheral nerves. In the central nervous system, glial cells contain enzymes related to the tricarboxylic acid and glutamine cycles: pyruvate carboxylase, glutamate dehydrogenase, and glutamine synthetase. The present study used immunohistochemistry in the rat peripheral nervous system to determine the cellular distribution of these enzymes along with glutamine. In dorsal root ganglia and peripheral nerves, glutamine and glutamine related enzymes were enriched in satellite and Schwann cells. In the dorsal root ganglia, immunoreactive satellite cells surrounded neurons of all sizes. In peripheral nerve, immunoreactive Schwann cells were most easily observed surrounding large diameter, myelinated axons. These Schwann cells contained immunoreactivity in their cell bodies, nodes of Ranvier, and the rim of cytoplasm outside the myelin sheath. Myelin sheaths were non-immunoreactive. The peripheral glial tricarboxylic and glutamine cycles may be used to produce glutamine for neuronal cell uptake and conversion to glutamate for synaptic transmission. Alternatively, these cycles may function in peripheral glia similar to central nervous system astrocytes for supporting the energy demands of neurons. PMID- 12126883 TI - 2-deoxy-D-glucose attenuates harmaline induced tremors in rats. AB - Neuronal hyperactivity in essential tremor is accompanied by high energy demand in cerebellum, medulla and the thalamus. It has been suggested that brain regions that have increased metabolic demands are highly vulnerable to interruptions in glucose metabolism. In the present investigation attempt was made to study the effect of 2-deoxyglucose (2DG) a glycolytic pathway inhibitor on harmaline induced tremor in rats. Wistar rats of either sex weighing 100+/-3 g were given harmaline (10 mg/kg, i.p.) alone or along with 2DG (15 min before harmaline) in doses of 300, 600 and 900 mg/kg, respectively. The latency of onset, intensity and duration of tremor following harmaline administration were recorded. Neurobehavioral responses, electromyography (EMG) and levels of blood glucose and cerebellar serotonin (5HT) were determined after 40 min of harmaline administration. 2DG significantly and dose dependently attenuated severity of harmaline induced tremors and amplitude of EMG. Treatment of rats with 2DG alone reduced the locomotor activity, however, no significant change was observed in grip strength, landing foot splay, air righting reflex and response to tactile stimuli. Harmaline alone and along with 2DG had no effect on behavioral parameters except a decrease in landing foot splay. 2DG produced a dose-dependent hyperglycemia and attenuated harmaline induced increase in cerebellar 5HT levels. Our results clearly suggest the protective effect of 2DG in harmaline induced tremor. Further studies are warranted to assess the role of glucoprivation in the suppression of neuronal excitability in tremors. PMID- 12126884 TI - Immunocytochemical characterization of hippocamposeptal projecting GABAergic nonprincipal neurons in the mouse brain: a retrograde labeling study. AB - The neurochemical contents of hippocamposeptal projecting nonprincipal neurons were examined in the mouse brain by using retrograde labeling techniques. We used the immunofluorescent multiple labeling method with a confocal laser-scanning microscope. First of all, the hippocamposeptal projecting nonprincipal neurons were glutamic acid decarboxylase 67-immunoreactive (IR), i.e., these hippocamposeptal projecting nonprincipal neurons were immunocytochemically GABAergic in the mouse brain. Next, most (93.0%) of the hippocamposeptal projecting GABAergic neurons were somatostatin-like immunoreactive (SS-LIR). The SS-LIR hippocamposeptal projecting neurons were frequently found in the stratum oriens of the CA1 and CA3 regions, and were also occasionally found in the stratum radiatum, stratum lucidum, and stratum pyramidale of the CA3 region. They were also frequently found in the dentate hilus. On the other hand, at least 40.6% of SS-LIR neurons in the hippocampus projected to the medial septum. Next, 38.0% of hippocamposeptal projecting GABAergic neurons were calbindin D28K (CB) IR. Although the distribution of the CB-IR hippocamposeptal projecting neurons was generally similar to that of the SS-LIR projecting neurons in Ammon's horn, they were never seen in the dentate hilus. At least 22.1% of CB-IR GABAergic neurons in the hippocampus projected to the medial septum. Furthermore, 5.8% of hippocamposeptal projecting GABAergic neurons were parvalbumin-IR, which were most always found in Ammon's horn. Finally, no hippocamposeptal projecting GABAergic neurons were neuronal nitric oxide synthase-IR nor calretinin-IR. These results indicate that the SS-LIR neurons play a crucial role in the hippocamposeptal projection of the mouse brain, and they are also assumed to be involved in the theta oscillation of the mouse hippocampus. PMID- 12126885 TI - Expression of TGF-beta type II receptors in the olfactory epithelium and their regulation in TGF-alpha transgenic mice. AB - Numerous in vitro studies of neurogenesis of olfactory receptor neurons (ORNs) suggest that transforming growth factor (TGF)-beta promotes the maturation/differentiation of olfactory progenitors. We demonstrate that in vivo both mature and immature ORNs, and possibly a basal neuronal progenitor cell, express the TGF-beta type II receptor (TGF-betaRII), suggesting that these cells are targets for TGF-beta signaling. In a previous study of neurogenesis in the OE of TGF-alpha overexpressing transgenic (T) mice, we observed an apparent reduction in the expression of olfactory marker protein (OMP), a marker of terminal differentiation in ORNs in T mice compared to nontransgenic (NT) littermate controls; this was confirmed by Western blotting and immunohistochemistry. In contrast, there was no apparent difference between T and NT mice in the intensity of immunoreactivity for a neuronal marker, protein gene product 9.5. Because TGF-alpha overexpression has been reported to affect TGF beta signaling in other epithelia, we compared the expression of the TGF-beta type II receptor (TGF-betaRII) in T and NT mice. The intensity of TGF-betaRII immunoreactivity on ORNs was substantially reduced in T compared to NT mice. Similar reductions in TGF-betaRII expression in vomeronasal receptor neurons and in other epithelia in the nasal cavity of T mice were also observed. Taken together, these results indicate that TGF-beta signaling regulates terminal differentiation of ORNs in vivo and suggest ways in which interactions between TGF-alpha and TGF-beta signaling pathways may interact in the OE. PMID- 12126886 TI - SM-20220, a Na(+)/H(+) exchanger inhibitor: effects on ischemic brain damage through edema and neutrophil accumulation in a rat middle cerebral artery occlusion model. AB - The Na(+)/H(+) exchanger (NHE) is activated during ischemia-reperfusion in an effort to restore intracellular pH to normal levels. The NHE is recognized to exist as a distinct protein in the plasma membranes of a variety of cells. We investigated the pharmacological effects of a Na(+)/H(+) exchanger inhibitor, SM 20220 (N-(aminoiminomethyl)-1-methyl-1-H-indole-2-carboxamide methanesulfonate), on ischemic brain damage, edema and neutrophil accumulation at 72 h after middle cerebral artery (MCA) occlusion in a rat MCA occlusion model. SM-20220 was intravenously administered as a bolus injection immediately after occlusion, followed by a continuous infusion over 2.5 h. At 72 h after occlusion, the infract area was measured using hematoxylin-eosin staining and, using the same slices, neutrophils in the brain were immuno-stained with anti-myeloperoxidase (n=11). In a separate study, rat behavior was scored and scaled, and brains removed for the determination of water content by the dry-weight method. SM-20220 significantly (P<0.05) attenuated cerebral infarct volume, water content, and the neutrophil accumulation at 72 h after the MCA occlusion, and ameliorated neurological deficits. SM-20220, an NHE inhibitor prevented the progress of cerebral ischemic damage and edema following MCA occlusion in rats though a possible mechanism that may be due to the inhibition of neutrophil accumulation. The NHE in neutrophils may enhance the progress of cerebral damage following cerebral ischemia-reperfusion. PMID- 12126887 TI - Serotonergic projections to the rostroventrolateral medulla from midbrain and raphe nuclei. AB - Double-label fluoresence immunohistochemistry was performed to define serotonergic projections from the raphe and midbrain to the sympathoexcitatory region of the rostroventrolateral medulla (RVLM). Immunolabelling of cholera toxin B subunit retrogradely transported from the pressor region of the RVLM was combined with serotonin (5-HT) immunohistochemistry. Major sources of serotonergic input to the RVLM were shown to include the raphe obscurus, raphe pallidus and raphe magnus with a minor contribution from the ventrolateral, lateral and ventral regions of the periaqueductal gray matter, and the dorsal raphe nucleus. Serotonergic modulation of sympathoexcitatory neurons may establish patterns of sympathetic nerve activity evident in many aspects of cardiovascular regulation. PMID- 12126888 TI - Antiamnesic effects of azaindolizinone derivative ZSET845 on impaired learning and decreased ChAT activity induced by amyloid-beta 25-35 in the rat. AB - Antiamnesic effects of a newly synthesized azaindolizinone derivative ZSET845 were assessed in rats made learning ability deficient by amyloid-beta (Abeta)25 35 treatment. Intracerebroventricular injection of Abeta25-35 induced a marked decrease in step-through latency in passive avoidance task and reduction in choline acetyltransferase (ChAT) activity in the medial septum and hippocampus, but not in the basal forebrain and cortex. The number of ChAT-immunoreactive cells was decreased in the medial septum. Oral administration of ZSET845 at a dose of 1 or 10 mg/kg ameliorated learning impairment in passive avoidance task and enhanced ChAT activity in the basal forebrain, medial septum and hippocampus, and increased in the number of ChAT-immunoreactive cells in the medial septum in Abeta-treated rats to the levels of vehicle-injected control rats. These results suggest that ZSET845 is worth testing for further preclinical study aimed for the treatment of senile dementia such as Alzheimer's disease. PMID- 12126889 TI - Expression of orphanin FQ/nociceptin and its receptor in rat peripheral ganglia and spinal cord. AB - Expression of the neuropeptide orphanin FQ/nociceptin (OFQ/N) and its receptor, the opioid receptor-like receptor (ORL1), have been found to have a wide distribution in the central nervous system, and in brain areas involved in sensory perception in particular. The effects of OFQ/N on, e.g., sensory transmission are very complex, and a modulatory effect on pain perception has been suggested. We therefore wanted to investigate the distribution of OFQ/N and ORL1 in the spinal cord and DRG, and also in SCG and some other peripheral tissues. The methods used were in situ hybridization, immunohistochemistry and ligand binding. We found that OFQ/N and ORL1 mRNA are expressed in DRG; primarily in small and large neurons, respectively. In spinal cord, mRNA for OFQ/N and ORL1 is expressed in neurons in laminae I, II and X, and in ventral horn neurons. Further, immunoreactivity for OFQ/N is observed in fibers and neurons in the superficial laminae of the dorsal horn and around the central canal, and also in neurons in the ventral horn of the spinal cord. Receptor ligand binding to the spinal cord grey matter is demonstrated, primarily concentrated to the dorsal horn and around the central canal, and also to medium and large size DRG neurons. These findings on the morphological distribution pattern of OFQ/N and ORL1 at the cellular level may support the notion that OFQ/N is involved in modulating pain transmission. Further, expression of OFQ/N and ORL1 mRNA was also found in SCG, whereas expression was undetectable in skin. PMID- 12126891 TI - Abstracts of the British Toxicology Society Annual Congress. Kent, Canterbury, United Kingdom, 7-10 April, 2002. PMID- 12126892 TI - On the 2002 measles vaccination furore in the UK. PMID- 12126890 TI - Phasic but not tonic REM-selective discharge of periaqueductal gray neurons in freely behaving animals: relevance to postulates of GABAergic inhibition of monoaminergic neurons. AB - To determine if ventrolateral periaqueductal gray contains neurons that selectively increase their discharge activity before and during rapid eye movement (REM) sleep, and hence might furnish GABAergic inhibition of monoaminergic neurons, we recorded the extracellular activity of 33 neurons across sleep-wakefulness in freely behaving cats. Several types of state-specific neuronal populations were found in the periaqueductal gray, although we did not find any neurons that had a tonic discharge increase before and during REM. Thus, these data suggest that, although periaqueductal gray neurons may regulate phasic components of REM sleep, they do not have the requisite tonic pre-REM and REM activity to be a source of GABAergic inhibition of monoaminergic neurons. PMID- 12126893 TI - The cost of not implementing routine neonates immunization programmes in HBsAg high prevalence countries. PMID- 12126894 TI - Anticapsular polysaccharide meningococcal antibodies in Israeli military recruits: immune status and the effect of simultaneous administration of immune globulin on the response to polysaccharide vaccine. AB - The effect of the administration of immune globulin (Ig), given during summer months to prevent hepatitis A, on the immune response to a simultaneously administered quadrivalent meningococcal polysaccharide vaccine (QMPV) was studied in Israeli military recruits. Data were obtained for the first time regarding the immune status of an Israeli population. Anticapsular polysaccharide antibodies were determined using a standardized ELISA assay before, 2 weeks and 3 months after QMPV immunization with or without Ig in two groups of recruits chosen to span the date determining seasonal administration or non-administration of Ig. Pre-vaccination antibody concentrations were > or = 2 microg/ml in 98.4 and 38.9% of subjects against serogroups A and C meningococci, respectively. These levels are consistent with the extremely low incidence of disease due to serogroup A in Israel, and with the previously documented occurrence of serogroup C disease in servicemen and women. Co-administration of Ig was associated with some reduction in antibody concentrations 3 months after vaccination, especially against serogroup A meningococci (P<0.05), but not to an extent likely to be of clinical significance. PMID- 12126895 TI - Adjuvant effect of multi-CpG motifs on an HIV-1 DNA vaccine. AB - Synthetic oligodeoxynucleotides (ODN) containing unmethylated CpG motifs trigger an immune response characterized by the activation of B cells, NK cells and monocytes/macrophages. Based on evidence that the immunogenicity of DNA vaccines can be augmented by the addition of CpG motifs, 5-20 additional CpG motifs were cloned into a pUC-derived plasmid. Treating bone-marrow derived dendritic cells (BM-DCs) with CpG-enriched plasmids in vitro boosted their expressions of MHC class II molecules, the CD40 and CD86 activation markers. Co-administering the CpG-enriched plasmids with a DNA vaccine encoding the envelope glycoprotein of HIV to BALB/c mice significantly increased HIV-specific cell mediated and humoral immunity. A significant boost was observed when the CpG plasmid was administered either 2 or 4 days after DNA vaccination. Plasmids containing 20 CpG copies were the most effective immune enhancers both in vitro and in vivo. These results suggest that plasmids containing multiple CpG motifs may improve the immunogenicity of DNA vaccines. PMID- 12126896 TI - Protection against oronasal challenge with virulent feline leukaemia virus lasts for at least 12 months following a primary course of immunisation with Leukocell 2 vaccine. AB - The duration of immunity provided by a feline leukemia virus (FeLV) vaccine, Leukocell 2, was determined. Kittens were vaccinated when 9 and 12 weeks of age and were challenged 12 months later with FeLV-A/Glasgow-1. An oronasal challenge protocol without corticosteroid enhancement was developed in order to induce a persistent viraemia in a high proportion of adult cats. Fourteen of 18 (80%) of the vaccinated cats challenged in this way remained non-viraemic while 9/15 (60%) of age-matched controls became persistently infected, a preventable fraction of 63%. This difference was statistically significant (P=0.038). For comparison, 10 of 12 (83%) 15-17-week-old kittens challenged in the same way became persistently infected, confirming the relative resistance of adult animals to FeLV. Tests for virus neutralising and anti-feline oncornavirus-associated cell membrane antigen (FOCMA) antibodies suggested that the former were more important than the latter in protection. Thus, Leukocell 2 protected a significant proportion of cats from FeLV challenge 1 year after primary vaccination as kittens. PMID- 12126897 TI - Pig chromosome aberrations after vaccination against classical swine fever in field trials. AB - Chromosomal aberrations were observed after vaccination against classical swine fever (CSF) in a previous study done on experimental pigs. To determine if the same effect occurs in farm animals, field trials were done with immunized pigs. The cytogenetic analysis was made from lymphocyte cultures of pigs sampled in three farms of Rio Cuarto region on six different periods: one, pre-vaccination (control) and in five post-vaccination (days 3, 7, 10, 15 and 22). Vaccine inoculation induced significant increase of cell frequency with chromosomal aberrations from days 3 to 10, when maximal frequencies of 6.78, 10.36 and 7.21% were observed in farms A, B and C, respectively. Afterwards, a decrease was recorded, reaching values not significantly different from controls. Mean frequencies of cells with chromosomal aberrations were 2.8+/-0.3, 5.9+/-0.4, 3.1+/-0.3%, for A, B and C farms, respectively. Cytogenetic effect was clastogenic, being chromosome breaks the first type of alteration to increase from the day 3 post-vaccination onwards, followed by chromatid exchanges and cells with multiple abnormalities. Chromosome pulverization was the last type to show increment, reaching a top value on day 10, after that it started to diminish gradually. There was no difference in the frequency of polyploid cells among sampling dates meaning that this type of alteration would not be induced by vaccination. Results of this field study confirm the mutagenic capacity of live virus vaccines against CSF and indicate that the evolution of chromosome alterations on the five evaluated post-vaccination periods was similar at different environments. PMID- 12126898 TI - DNA immunization using a secreted cell wall antigen Mp1p is protective against Penicillium marneffei infection. AB - None of the vaccines used in dimorphic fungal infections utilized the mucosal route for immunization, whereas only one utilized a secreted protein as antigen, despite knowing that infections caused by dimorphic fungi are usually acquired through inhalation. In this study, we investigated the usefulness of Mp1p (a secreted cell wall antigen encoded by MP1)-based vaccines for generation of protective immune responses against Penicillium marneffei infection using a mouse model, and compared the relative effectiveness of intramuscular MP1 DNA vaccine, oral mucosal MP1 DNA vaccine delivered by live-attenuated Salmonella typhimurium, and intraperitoneal recombinant Mp1p protein vaccine. The serum IgM level of the Mp1p protein vaccine group at day 7 and the serum IgG levels of the Mp1p protein vaccine group at days 7 and 21 were significantly higher than those of the other groups (P<0.0001). The serum IgG level of the MP1 DNA vaccine group was significantly higher than that of the corresponding control group and oral mucosal MP1 DNA vaccine group (one dose) at day 21 (P<0.0001 and <0.05, respectively). The groups of mice immunized with intramuscular MP1 DNA vaccine, oral mucosal MP1 DNA vaccine, and intraperitoneal Mp1p protein vaccine showed significantly higher Mp1p-specific lymphocyte proliferation index (LPI) than the control groups. The interferon-gamma (IF-gamma) levels of supernatant of splenic cell cultures obtained from mice after intramuscular MP1 DNA vaccine, mucosal MP1 DNA vaccine (three doses), or intraperitoneal Mp1p protein vaccine administration were higher than that which occurred after mucosal MP1 DNA vaccine (one dose) administration or those of controls. Interleukin-4 (IL-4) was not detectable in the supernatant of splenic cell cultures obtained from all groups of mice. The percentage survival of the mice immunized with intramuscular MP1 DNA vaccine, oral mucosal MP1 DNA vaccine (three doses), oral mucosal MP1 DNA vaccine (one dose), intraperitoneal recombinant Mp1p protein, oral live-attenuated S. typhimurium control, and intramuscular pJW4303 DNA control at day 60 after wild type P. marneffei challenge were 100, 60, 40, 40, 40, and 0%, respectively. The survival of mice in the MP1 DNA vaccine group was significantly better than those of the oral mucosal MP1 DNA vaccine (three doses) group (P<0.05), oral mucosal MP1 DNA vaccine (one dose) group (P<0.005), recombinant Mp1p protein group (P<0.005), S. typhimurium aroA strain group (P<0.05), and pJW4303 group (P<0.00001). Although, the mechanism by which intramuscular MP1 DNA vaccine offered the best protection against P. marneffei infection remains to be elucidated, the present observation prompted further clinical trials on the use of MP1 DNA immunization on asymptomatic human immunodeficiency virus carriers in P. marneffei endemic areas. PMID- 12126899 TI - Enhanced immunogenicity of BCG vaccine by using a viral-based GM-CSF transgene adjuvant formulation. AB - The failure of current BCG vaccine in controlling the global tuberculosis (TB) epidemic highlights an urgent need for improved TB vaccine formulations. In this study, we have investigated the effect of a novel adenoviral granulocyte macrophage-colony stimulating factor (GM-CSF) transgene-based adjuvant formulation (AdGM-CSF) on BCG vaccination in a mouse strain that is genetically weak responders to BCG vaccine. BALB/c mice were immunized subcutaneously (s.c.) with PBS, BCG, or BCG plus AdGM-CSF or control vector Addl70-3, the immunogenicity of BCG vaccine was evaluated by type 1 IFN-gamma production from lymphocytes of various lymphoid tissues upon mycobacterial antigen stimulation ex vivo. While mycobacterial antigen-specific IFN-gamma production was slightly enhanced by co-immunization BCG with Addl70-3 as compared to BCG immunization alone, a marked increase both in the magnitude and longevity of anti mycobacterial type 1 immunity was observed in the local draining lymph nodes and spleens by immunization with AdGM-CSF-adjuvanted BCG. Furthermore, there was a significant increase in the number of mycobacterial antigen-specific IFN-gamma releasing CD4 T cells in mice immunized with AdGM-CSF-adjuvanted BCG vaccine. Consistent with these enhanced T-cell immunity and memory responses, AdGM-CSF adjuvanted BCG vaccine significantly improved immune protection against secondary mycobacterial challenge. Our results suggest that GM-CSF transgene-based adjuvant formulation is an effective way to improve the immunogenicity of BCG vaccine. PMID- 12126900 TI - Anti-HBs levels after hepatitis B immunisation depend on test reagents: routinely determined 10 and 100 IU/l seroprotection levels unreliable. AB - In a large series of post-vaccination samples we compared the result of three commercially available anti-HBs assays (AxSYM, Architect and Access) on the quantitation of anti-HBs after immunisation with Engerix-B (HBsAg/ad) and GenHevacB (HBsAg/ay) vaccine. Two of the assays (AxSYM, Architect: Abbott Laboratories) gave related but not identical results with HBsAg from different sources. The result of the third assay (Access, Beckman Coulter) was related to that of AxSYM and Architect only for GenHevacB anti-HBs but differed for Engerix B anti-HBs (P<0.001). This vaccine dependent discrepancy was also observed with the Vidas anti-HBs assay (BioMerieux). An external WHO reference panel could harmonise geometric mean anti-HBs levels from the four assays for GenHevacB but not for Engerix-B vaccination sera. We conclude that the individually determined anti-HBs level (IU/l) strongly depend on the test reagents and the vaccine under study. PMID- 12126901 TI - Enteric administration of a live attenuated measles vaccine does not induce protective immunity in a macaque model. AB - To test the option of oral vaccination with a live attenuated measles vaccine (LAV), we have evaluated the potential of an orally administered enteric-coated tablet containing a candidate LAV (strain Leningrad-16, MV-L16). To this end three groups of two cynomolgus macaques each were vaccinated via different routes with 10(3.8) TCID(50) MV-L16 vaccine: intramuscularly (i.m.), intraintestinally (i.i.) upon laparotomy and via enteric-coated tablets. Upon vaccination, MV-L16 could only be isolated from one of the i.m.-vaccinated monkeys and not from any of the other five. Both the i.m.-infected monkeys and one of the i.i.-infected monkeys developed a MV-specific serum antibody response. Also, MV-specific CD8(+) IFN gamma-producing T cells could be demonstrated in all three monkeys that had seroconverted. Upon challenge with wild-type MV 1 year after vaccination, only these three monkeys proved to be protected. These data do not support the viability of the concept of oral vaccination with LAVs. PMID- 12126902 TI - Recombinant Lactobacillus johnsonii as a mucosal vaccine delivery vehicle. AB - Lactobacilli are considered to be safe organisms making them attractive as vehicles for oral vaccination. We report that Lactobacillus johnsonii (Lj) partially survived simulated gastric conditions in vitro, suggesting that it could be used as an oral vaccine delivery vehicle. In order to test this approach, we used the cell wall anchored proteinase PrtB, isolated from Lactobacillus delbrueckii subsp. bulgaricus as a model antigen. Using a new vector system, we demonstrated expression of both proteinase PrtB alone and a mimotope peptide derived from tetanus toxin integrated in the sequence of proteinase PrtB (TTmim-PrtB fusion protein) on the surface of Lj. Oral immunisation of mice with recombinant Lj, expressing the TTmim-PrtB fusion protein induced a systemic IgG response against Lj and recombinantly expressed proteinase PrtB but no antibody response against the tetanus toxin mimotope suggesting that the mimotope was not sufficiently immunogenic to induce an immune response. Interestingly, a proteinase PrtB specific fecal IgA response was also induced, indicating that the proteinase PrtB fusion protein expressed as a cell surface protein on Lj can induce both systemic and local mucosal immune responses. PMID- 12126903 TI - Comparison of the efficacy of early versus late viral proteins in vaccination against SIV. AB - The immune response against early regulatory proteins of simian- and human immunodeficiency virus (SIV, HIV) has been associated with a milder course of infection. Here, we directly compared vaccination with Tat/Rev versus Pol/Gag. Challenge infection with SIVmac32H (pJ5) suggested that vaccination with Tat/Rev induced cellular immune responses that enabled cynomolgus macaques to more efficiently control SIV replication than the vaccine-induced immune responses against Pol/Gag. Vaccination with Tat/Rev resulted in reduced plasma SIV loads compared with control (P=0.058) or Pol/Gag-vaccinated (P=0.089) animals, with undetectable plasma viral loads in two of the four Tat/Rev-vaccinated animals. Therefore, the results warrant further investigation of the early regulatory proteins and their potential for vaccination against HIV. PMID- 12126904 TI - Long-term pertussis-specific immune responses to a combined diphtheria, tetanus, tricomponent acellular pertussis and hepatitis B vaccine in pre-term infants. AB - Immunoglobulin G (IgG) antibody titres against pertussis antigens, Bordetella pertussis-specific proliferation and cytokine production by peripheral blood mononuclear cells (PBMCs) were evaluated at the age of 5-6 years in 13 children who had been pre-term infants with a gestational age (GA) of < or = 31 weeks, 10 who had been pre-term infants with a GA of 32-37 weeks, and 15 who had been term infants with a GA of 38-42 weeks. All of the infants had been immunised with a combined diphtheria, tetanus, tricomponent acellular pertussis and hepatitis B vaccine (DTaP-HBV) at 3, 5, and 11 months of post-natal age. Our results show that the long-term immune responses induced by primary pertussis vaccination in the pre-term infants (especially those with a GA of < or = 31 weeks) were qualitatively and quantitatively lower than those observed in the term infants. In order to maintain an adequate pertussis-specific immune response, in term children a booster at 5-6 years of age may be suggested, whereas in pre-term an earlier booster should be useful. PMID- 12126905 TI - Co-immunisation with a plasmid DNA cocktail primes mice against anthrax and plague. AB - The protective antigen (PA) of Bacillus anthracis and the V antigen of Yersinia pestis are potent immunogens and candidate vaccine sub-units. When plasmid DNA encoding either PA or V antigen was used to immunise the Balb/c mouse, a low serum IgG titre was detected (log (10)1.0 or less) which was slightly increased by boosting with plasmid DNA. However, when mice immunised with plasmid DNA were later boosted with the respective recombinant protein, a significant increase in titre (up to 100-fold) was observed. Mice primed with a combination of each plasmid and boosted with a combination of the recombinant proteins, were fully protected (6/6) against challenge with Y. pestis. This compared favourably with mice primed only with plasmid DNA encoding the V antigen and boosted with rV, which were partially protected (3/6) against homologous challenge or with mice primed and boosted with plasmid DNA encoding the V antigen which were poorly protected (1/6). Combined immunisation with the two plasmid DNA constructs followed by boosting with a combination of the encoded recombinant proteins enhanced the protective immune response to Y. pestis compared with priming only with plasmid DNA encoding the V antigen and boosting with rV. This enhancement may be due to the effect of CpG motifs known to be present in the plasmid DNA construct encoding PA. PMID- 12126906 TI - A comparison of safety, tolerability and immunogenicity of Oka/Merck varicella vaccine and VARILRIX in healthy children. AB - This study compared safety, tolerability, and immunogenicity of the Oka/Merck varicella vaccine and VARILRIX [Oka-RIT strain SmithKline Beecham Biologicals] in healthy children 12-24 months of age. Subjects were randomized in this double blind study to receive either a single dose of Oka/Merck varicella vaccine, (approximately 50,000 plaque forming units (PFU), Group A or approximately 16,000 PFU, Group B) or 1 dose of VARILRIX, (approximately 40,000 PFU/dose, Group C). Safety profiles in each treatment group were similar. The proportions of subjects achieving a 6-week postvaccination titer> or = 5 gpELISA units in Groups A, B or C were 97.1, 95.2 and 85.6%, respectively. PMID- 12126907 TI - Early phase II trial of human rotavirus vaccine candidate RV3. AB - A naturally attenuated, human neonatal strain, rotavirus vaccine candidate RV3, was tested in a limited phase II randomized double-blind controlled trial. Doses of 1 ml, containing placebo or 6.5 x 10(5) fluorescent cell forming units (fcfu) of virus in AGMK cells, were given at 3, 5 and 7 months of age. Limited replication in the small intestine is implied by the lack of virus excretion, and by the occurrence of an immune response in only 46% of the infants. However, those who developed an immune response were partially protected against rotavirus disease during the subsequent winter epidemic (protective efficacy 54%), supporting observations of protection induced by natural infection by this strain. Protection appeared to be heterotypic. Further trials are warranted, employing strategies to increase immunogenicity of this human rotavirus candidate vaccine. PMID- 12126908 TI - In vitro induction of memory-driven responses against Neisseria meningitidis by priming with Neisseria lactamica. AB - Natural immunity against Neisseria meningitidis is acquired during childhood and youth through successive colonizations by commensal Neisseria, carrier N. meningitidis, and other bacterial genera sharing cross-reactive antigens with the meningococci. We have analyzed in mice the ability of Neisseria lactamica strains to induce immunological memory so that, upon a later contact with N. meningitidis, quickly raise protective responses against antigens that show cross reactivity with meningococcal surface proteins. Sera obtained from mice immunized with N. lactamica and boosted with N. meningitidis were able to kill meningococci, with bactericidal activities variable depending on the immunizing strains used in the assays. Different mixtures of those sera resulted in higher killing activities, which agrees with the idea that successive colonizations by N. lactamica enhance the anti-meningococcal response. The existence of such outer membrane cross-reactive antigens has to be kept in mind when using outer membrane vesicle (OMV)-based anti-meningococcal vaccines because their use can affect colonization by N. lactamica and other species, hampering the natural mechanisms of acquisition of immunity to the meningococci, and leaving its ecological niche free for colonization by undesirable microorganisms. PMID- 12126909 TI - Interspecific neisserial high molecular weight proteins able to induce natural immunity responses are strongly correlated with in vitro bactericidal activity. AB - Human sera from healthy volunteers and patients convalescent from meningitis were used to search for widely cross-reactive antigens implicated in vivo protective responses. Using the type strain Neisseria meningitidis B16B6 and a wide variety of both N. meningitidis and N. lactamica strains, several cross-reactive antigens and bactericidal sera were found, although the cross-reactivity patterns did not correlate with bactericidal activity, a total correlation was found between bactericidal activity and reaction with one or two high molecular weight proteins (162 and 138 kDa), and a mouse serum against the purified proteins showed a high bactericidal activity. Our results suggest that the high molecular weight proteins found are immunogenic and cross-reactive, eliciting bactericidal responses during infection and as a result of natural immunity. These proteins should be taken into account in studies for future vaccine formulations. PMID- 12126910 TI - Nasal vaccination induces the ability to eliminate Candida colonization without influencing the pre-existing antigen-specific IgE Abs: a possibility for the control of Candida-related atopic dermatitis. AB - In some cases of atopic dermatitis (AD), a possible pathological contribution to disease development by Candida albicans (C. albicans) has been suggested. AD patients with severe symptoms showing positive capsulated hydrolic carrier polymer radioallergosorbent test (CAP-RAST) against C. albicans demonstrated significantly higher levels of serum IgE Abs than did AD patients with mild symptoms. Based on the clinical facts, we have postulated that elimination of C. albicans by mucosal vaccination may lead to the restoration of severe symptoms in AD patients. For this purpose, we have developed an allergic murine model. Mice which were systemically challenged with C. albicans-associated antigen, manganese superoxide dismutase (MnSOD) or secreted aspartic proteases 2 (SAP2), together with alum, exhibited hyper IgE Abs. Systemically primed mice were then immunized with MnSOD or SAP2 plus cholera toxin (CT) as mucosal adjuvant through the nasal route. Interestingly, nasally immunized mice showed increased levels of Candida Ag-specific IgA Ab in fecal and nasal washes as well as in saliva samples but unchanged levels in Ag-specific IgE responses. Consistent with the Ab levels, high numbers of Candida Ag-specific IgA Ab-forming cells were induced in mononuclear cells isolated from intestinal lamina propria, nasal passages and salivary glands of nasally vaccinated mice with Ag plus CT. Furthermore, nasal immunization using MnSOD or SAP2 together with CT resulted in the elimination of colonized C. albicans from the intestinal tract. These results also suggest a potential role of mucosal vaccination in the control of C. albicans in patients with allergic diseases, including AD, although more research is needed to establish this therapeutic approach for mucosal vaccination. PMID- 12126911 TI - Serum-derived IgG1-mediated immune exclusion as a mechanism of protection against H. pylori infection. AB - The induction of protective immunity against Helicobacter challenge in a murine model was found to correlate with the magnitude of IgG (serum and gastric lavage) responsiveness to intra-nasal (i.n.) immunisation. IgG1-secreting hybridoma backpacks in Helicobacter pylori (H. pylori)-infected mice revealed serum transudation into the stomach. A Lpp20-specific monoclonal antibody was associated with significantly reduced H. pylori colonisation. Histology revealed aggregates of the remaining H. pylori in these mice, suggesting a role for IgG1 mediated immune exclusion of the bacteria. In vitro immunogold electron microscopy supported this hypothesis, but also suggested that a threshold of H. pylori-specific antibody needs to be maintained if immune exclusion by the host is to overcome immune evasion by the bacteria. PMID- 12126912 TI - The epidemiology of invasive Streptococcus pneumoniae disease in Catalonia (Spain). A hospital-based study. AB - The aim of this study was to investigate the incidence of invasive pneumococcal disease (IPD) in Catalonia. A hospital-based incidence study of the period 1997 1999 was carried out by reviewing the isolations of Streptococcus pneumoniae obtained from normally sterile sites reported by the hospitals that voluntarily participate in the Microbiological Reporting System of Catalonia (MRSC), and those obtained by active retrospective surveillance of cases recorded by microbiology laboratories of the remaining hospitals. Cases of pneumonia were included only if the blood culture was positive. The age, sex and clinical presentation of each patient were recorded. The global incidence of hospital based IPD was 10.5 per 100,000 persons-year, and was much higher in subjects <2 years of age (59.6) and in those aged > or = 65 years (27.9). In subjects > or = 65 years, bacteremic pneumococcal pneumonias were more frequent in the months from December to March than they were in children <2 years of age (P<0.0001). The global incidence of IPD (10.5 per 100,000 persons-year) is high in Catalonia, greater than that of meningococcal or Haemophilus influenzae invasive disease. In children <2 years, the incidence is nearly six times higher (59.6 per 100,000 persons-year) with pneumonias (rate of 26.2 per 100,000 persons-year) and non focal bacteremias (rate of 22.1 per 100,000 persons-year) being especially frequent. PMID- 12126913 TI - Different kinetic of antibody responses following infection of newly weaned pigs with an F4 enterotoxigenic Escherichia coli strain or an F18 verotoxigenic Escherichia coli strain. AB - To develop a vaccine against Escherichia coli-induced post-weaning diarrhea and edema disease, insights in the induction of the protective immune response following infection with these pathogenic E. coli is needed. Therefore, the fimbriae-specific antibody response of newly weaned pigs following infection with the Shiga-like toxin type II variant (SLT-IIv) producing F18(+) verotoxigenic E. coli (VTEC) (strain 107/86) was compared with the response following an infection with LT producing F4(+) enterotoxigenic E. coli (ETEC) (strain GIS 26). F4(+) ETEC were able to colonize the gut very soon after infection, as peak excretion of F4(+) E. coli bacteria was seen 2 days post-infection (dpi), but had already disappeared 7dpi. On the other hand, F18(+) VTEC infection resulted in a slower colonization of the gut as the peak excretion of F18(+) E. coli was observed between 3 and 5dpi, but this colonization remained longer as F18(+) E. coli were detected till 9dpi in feces. Furthermore, this fast colonization pattern of F4(+) ETEC is accompanied with the presence of F4-specific antibodies in mucosal tissues and serum from 4dpi onward, with maximal amounts of F4-specific IgA in the jejunal lamina propria and serum 7dpi. In contrast, F18-specific IgA was only readily detected in the jejunal lamina propria 15dpi and showed a maximum serum titer 21dpi. Besides this faster induction and higher antibody response, the switch from IgM to IgA and IgG was also earlier following the F4(+) ETEC infection. PMID- 12126914 TI - Influence of maternal antibodies on efficacy of a subunit vaccine: transmission of classical swine fever virus between pigs vaccinated at 2 weeks of age. AB - This study shows the effectiveness of vaccination with an E2 subunit vaccine against classical swine fever (CSF) in 2-week-old piglets. Half of the piglets were carrying maternally derived antibodies (MDAs) at the time of vaccination. Three and 6 months later, antibody levels were compared between the two treatments. Moreover, reduction of virus transmission was investigated at 3 and 6 months by doing transmission experiments. The vaccine was found to be capable of reducing virus transmission significantly at both time intervals. Maternal immunity reduced vaccination-induced antibody levels after 3 and 6 months and possibly led to a less effective protection against virus transmission after 6 months. PMID- 12126915 TI - CpG-containing oligodeoxynucleotides augment and switch the immune responses of cattle to bovine herpesvirus-1 glycoprotein D. AB - The adjuvanticity of a synthetic oligodeoxynucleotide containing unmethylated CpG motifs (CpG ODN) was determined in cattle. Calves were immunized with a truncated secreted version of glycoprotein D (tgD) of bovine herpes virus-1 (BHV-1) formulated with alum, CpG ODN, or a combination of both. BHV-1 tgD formulated with CpG ODN or with alum and CpG ODN induced a stronger and more balanced immune response than tgD in alum. This level of immunity was of sufficient magnitude to minimize weight loss and significantly reduce the duration of virus shedding after intranasal viral challenge. Local tissue reactions generated by CpG ODN were very mild and transient, whereas reactions induced by alum or a combination of CpG ODN and alum were moderate in severity and duration. These data demonstrate that CpG ODN causes minimal injection site reactions and yet acts as an effective adjuvant in cattle. PMID- 12126916 TI - Plasmid DNA encoding the respiratory syncytial virus G protein protects against RSV-induced airway hyperresponsiveness. AB - Respiratory syncytial virus (RSV) is an important cause of childhood respiratory disease as well as exacerbations of asthma. Although previous studies have demonstrated that a DNA vaccine encoding the RSV G protein can inhibit RSV replication in mouse models of RSV infection, studies have not been performed to determine whether a DNA vaccine encoding the RSV G protein can protect against RSV induced mucus expression and airway hyperresponsiveness which was the focus of this study. The DNA-G vaccine we constructed significantly inhibited RSV viral replication, mucus expression, and importantly was associated with inhibition of RSV induced airway responsiveness. PMID- 12126917 TI - The effect of vaccination with a Salmonella enteritidis aroA mutant on early cellular responses in caecal lamina propria of newly-hatched chickens. AB - When newly hatched chicks are inoculated with a Salmonella strain, they induce a rapid onset of resistance to intestinal colonization by other Salmonella strains. The exact mechanism of this early colonization-inhibition is not known. To study host-related contributions to this phenomenon, the kinetics of immune cell infiltration in the caecal wall was analyzed during the first 10 days after vaccination of newly hatched chickens with a Salmonella enterica serovar Enteritidis aroA mutant, and infection 1 day later with a virulent S. enterica serovar Enteritidis strain. These data were correlated with bacterial colonization and clearance of the Salmonella Enteritidis challenge strain. Bacteriological data showed that vaccinated animals had a much lower number of challenge bacteria in their organs and caecal contents the first days post challenge, relative to unvaccinated animals. Immunohistochemical analysis of the caecal lamina propria revealed that heterophils started infiltrating the caecal lamina propria from 12 h post-vaccination. Macrophages and T-lymphocytes started infiltrating from 20 h and B-lymphocytes from 24 h post-vaccination. These data imply that immune cells already colonized the caecal wall at the time of challenge in vaccinated animals. The presence and activity of these cells in the caecal wall shortly after administration of a Salmonella Enteritidis aroA mutant might contribute to the inhibition of colonization of a virulent Salmonella strain, subsequently administered. PMID- 12126918 TI - Gap junctions in CO(2)-chemoreception and respiratory control. AB - Recent evidence indicates that gap junctions play a more prominent role in normal functioning of the mammalian central nervous system (CNS) than was once believed. Accumulating evidence from both neonatal and adult rodents indicates that gap junctions participate in multiple aspects of respiratory control, including central CO(2) chemoreception, respiratory rhythmogenesis, and respiratory motoneuron output. This review provides an overview of gap junction neurobiology in the mammalian CNS and presents the anatomical and electrophysiological evidence for gap junctions in CO(2) chemoreception and respiratory control. PMID- 12126919 TI - Carotid body chemosensory excitation induced by nitric oxide: involvement of oxidative metabolism. AB - Nitric oxide (NO) produces a dual effect on carotid body (CB) oxygen chemoreception. At low concentration, NO inhibits chemosensory response to hypoxia, while in normoxia, medium and high [NO] increases the frequency of carotid chemosensory discharges (f(x)). Since NO and peroxynitrite inhibit mitochondrial respiration, it is plausible that the NO-induced excitation may depend on the mitochondrial oxidative metabolism. To test this hypothesis, we studied the effects of oligomycin, FCCP and antimycin A that produce selective blockade of hypoxic and NaCN-induced chemosensory responses, leaving nicotinic response less affected. CBs excised from pentobarbitone-anaesthetised cats were perfused in vitro with Tyrode (P(O(2)) approximately 125 Torr, pH 7.40 at 38 degrees C). Hypoxia (P(O(2)) approximately equal 30 Torr), NaCN and nicotine (1 100 microg) and S-nitroso-N-acetylpenicillamide (SNAP, 300-600 microg) increased f(x). Oligomycin (12.5-25 microg), antimycin A (10 microg) and FCCP (5 microM) transiently increased f(x). Subsequently, chemosensory responses to hypoxia, NaCN and SNAP were reduced or abolished, while the response to nicotine was less affected. The electron donor system tetramethyl-p-phenylene diamide and ascorbate that bypasses the electron chain blockade produced by antimycin A, restores the excitatory responses to NaCN and SNAP. Present results suggest that the chemoexcitatory effect of NO depends on the integrity of mitochondrial metabolism. PMID- 12126920 TI - Apnoeic response to stimulation of peripheral GABA receptors in rats. AB - Respiratory effects of intracarotid injection of gamma-amino-butyric acid (GABA) were investigated in two groups of rats. In the first group of 12 rats the effects of GABA were checked in the intact state, following bilateral vagotomy and GABA receptor blockade. The second group consisted of five initially vagotomized rats, challenged with GABA prior to and after bilateral carotid chemodenervation (CSN-cut). All rats were urethane and chloralose anaesthetized and spontaneously breathing. Injection of 39 micromol/kg GABA prior to and after vagotomy induced an expiratory apnoea of, respectively 5.5+/-0.84 sec and 3.9+/ 0.6 sec duration (mean+/-S.E.M.), P>0.05 in all 12 rats. In breaths that followed the apnoea tidal volume increased above the control level by 23.3% (P<0.01) and 25.6% (P<0.01) pre- and post-vagotomy, respectively. Blockade of GABA receptors with bicuculline and picrotoxin abolished the inhibition of breathing. In five vagotomized rats with intact carotid sinus nerves (CSNs) intracarotid GABA challenge increased tidal volume by 39% compared with baseline breathing (P<0.05). Section of the CSNs precluded the occurrence of apnoea and undergoing respiratory changes evoked by GABA. Intracarotid GABA caused significant decrease in the mean blood pressure independent of the neural state, but the fall was delayed by CSNs neurotomy. Results of this study indicate that GABA given systemically induces apnoea followed by post-apnoeic hyperventilation. Carotid bodies are required for the ventilatory response to GABA; vagal afferents are not involved in this response. PMID- 12126921 TI - Lack of involvement of mu(1) opioid receptors in dermorphin-induced inhibition of hypoxic and hypercapnic ventilation in rat pups. AB - The effects of dermorphin, a mu-selective opioid agonist, on respiratory responses to altered O(2) and CO(2) during postnatal development were investigated in conscious, unrestrained Wistar rats aged 2-21 days. Respiration was recorded by barometric plethysmography. Dermorphin (4 mg kg(-1)) was administered subcutaneously, and the ventilatory responses to hypoxia (11% O(2), 89% N(2)) in 2-21-day-old pups and hyperoxia (100% O(2)), and hypercapnia (8% CO(2), 92% O(2)) in 2-13-day-old pups were assessed in the presence and absence of the mu(1) receptor antagonist naloxonazine (10 mg kg(-1) s.c.) administered 1 day before testing. Six minutes of hypoxia increased ventilation in all age groups, largely via an increase in frequency. Dermorphin inhibited the ventilatory response to hypoxia, and this inhibition was insensitive to naloxonazine. After 5 min of hyperoxia, ventilation was the same as with air breathing except in the presence of dermorphin, when hyperoxic ventilation was depressed by a naloxonazine-insensitive decrease in frequency. Following this 5 min 100% O(2) exposure, pups were exposed to hypercapnia, and respiratory parameters were measured 5 min later. The ventilatory response to CO(2) was inhibited by dermorphin in a naloxonazine-insensitive manner. There was no evidence for endogenous mu(1) receptor modulation of the ventilatory responses to altered gases in rat pups of any age. Thus, mu opioid-induced inhibition of the hypoxic and hypercapnic responses in young rats does not occur via activation of mu(1) opioid receptors. PMID- 12126922 TI - Ventilatory responses to hypercapnia and hypoxia in heterozygous c-ret newborn mice. AB - The c-ret proto-oncogene encodes a tyrosine-kinase receptor involved in survival and differentiation of neural crest cell lineages. Previous studies have shown that homozygous c-ret-/- mice die soon after birth and have impaired ventilatory responses to hypercapnia. Heterozygous c-ret +/- mice develop normally, but their respiratory phenotype has not been described in detail. We used whole-body flow plethysmography to compare baseline breathing and ventilatory and arousal responses to chemical stimuli in unrestrained heterozygous c-ret +/- newborn mice and their wild-type c-ret +/+ littermates at 10-12 h of postnatal age. The hyperpnoeic and arousal responses to hypoxia and hypercapnia were not significantly different in these two groups. However, the number and total duration of apnoeas and periodic breathing episodes were significantly higher in c-ret +/- than in c-ret +/+ pups during hypoxia and post-hypoxic normoxia. These results are further evidence that respiratory control at birth is heavily dependent on genes involved in the neural determination of neural crest cells. PMID- 12126923 TI - Cardioventilatory effects of acclimatization to aquatic hypoxia in channel catfish. AB - The mechanisms responsible for altering cardioventilatory control in vertebrates in response to chronic hypoxia are not well understood but appear to be mediated through the oxygen-sensitive chemoreceptor pathway. Little is known about the effects of chronic hypoxia on cardioventilatory control in vertebrates other than mammals. The purpose of this study was to determine how cardioventilatory control and the pattern of response is altered in channel catfish (Ictalurus punctatus) by 1 week of moderate hypoxia. Fish were acclimatized for 7 days in either normoxia (P(O(2)) approximately 150 Torr) or hypoxia (P(O(2)) approximately 75 Torr). After acclimatization, cardioventilatory, blood-gas and acid/base variables were measured during normoxia (P(O(2)) 148+/-1 Torr) then at two levels of acute (5 min) hypoxia, (P(O(2)) 72.6+/-1 and 50.4+/-0.4 Torr). Ventilation was significantly greater in hypoxic acclimatized fish as was the ventilatory sensitivity to hypoxia (Delta ventilation/Delta P(O(2))). The increase in ventilation and hypoxic sensitivity was due to increases in opercular pressure amplitude, gill ventilation frequency did not change. Heart rate was greater in hypoxic acclimatized fish but decreased in both acclimatization groups in response to acute hypoxia. Heart rate sensitivity to hypoxia (Delta heart rate/Delta P(O(2))) was not affected by hypoxic acclimatization. The ventilatory effects of hypoxic acclimatization can be explained by increased sensitivity to oxygen but the effects on heart rate cannot. PMID- 12126924 TI - Relative motion of lung and chest wall promotes uniform pleural space thickness. AB - The pleural space is modeled in two dimensions as a thin layer of fluid separating a deformable membrane and a rigid surface containing a bump. We computed the steady-state membrane configuration and fluid pressure distribution during relative sliding of the two surfaces. For physiologically relevant values of membrane tension, shear flow-induced pressures near the bump and far-field pressure gradients are similar to those measured in vivo within the pleural space (e.g. Lai-Fook et al.) [J. Appl. Physiol.: Respirat. Environ. Exercise Physiol. 56 (1984) 1633-1639]. Deformation of the membrane over the bump suggests that the pressure field generated by the sliding motion promotes an even layer of fluid in the pleural space, preventing asperities from touching. Results also suggest a possible mechanism for pleural fluid redistribution during breathing, whereby irreversible fluid motion is associated with the deformability of the membrane. PMID- 12126925 TI - Inhaled sodium fluoride decreases airway responsiveness to acetylcholine analogs in vivo. AB - The study was conducted to characterize the action of NaF, which had relaxing property in carbachol precontracted isolated bovine bronchus, on airway responsiveness challenged by acetylcholine receptor agonists in rats and asthmatic humans. Tracheal flow rate and airway resistance were measured in anaesthetized rats. NaF was delivered either before carbachol challenge or together with carbachol. Patients with mild asthma were challenged with methacholine aerosol, and NaF was delivered when FEV1 fell by more than 20%. The results indicated that: (1) in rats NaF significantly inhibited carbachol-induced bronchial constriction when inhaled prior to carbachol challenge as airway resistances in the NaF and NaF+verapamil groups were significantly lower than those in the control group; (2) NaF significantly reversed carbachol or methacholine-induced bronchial constriction in asthmatic patients. In conclusion, NaF, delivered in form of aerosol, reduced bronchial responsiveness to carbachol in rats and had a bronchodilating effect on rat and human airways precontracted by inhalation of acetylcholine analogs. PMID- 12126927 TI - The effects of ventilation pattern on carbon dioxide transfer in three computer models of the airways. AB - We investigate the effects on arterial P(CO(2)) and on arterial-end tidal P(CO(2)) difference of six different ventilation patterns of equal tidal volume, and also of various combinations of tidal volume and respiratory rate that maintain a constant alveolar ventilation. We use predictions from three different mathematical models. Models 1 (distributed) and 2 (compartmental) include combined convection and diffusion effects. Model 3 incorporates a single well mixed alveolar compartment and an anatomical dead-space in which plug flow occurs. We found that: (i) breathing patterns with longer inspiratory times yield lower arterial P(CO(2)); (ii) varying tidal volume and respiratory rate so that alveolar ventilation is kept constant may change both PA(CO(2)) and the PA(CO(2)) PET(CO(2)) difference; (iii) the distributed model predicts higher end-tidal and arterial P(CO(2)) than the compartmental models under similar conditions; and (iv) P(CO(2)) capnograms predicted by the distributed model exhibit longer phase I and steeper phase II than other models. PMID- 12126926 TI - Acute hypervolemia lengthens red cell pulmonary transit time during exercise in endurance athletes. AB - The purpose was to determine if acute plasma volume expansion (PVE) changed red cell pulmonary transit time (PTT) during severe exercise. Twelve endurance athletes performed 6.5 min of severe cycling exercise on different days. Pentaspan [(500 ml, infusion condition, I] or placebo [(60 ml saline), non infusion condition, N] were infused prior to exercise. Blood gas tensions, PTT, multigated acquisition (MUGA) derived cardiac output, and oxygen uptake were measured during exercise. PTT was measured during minute 3 of exercise by radionuclide cardiography. Arterial P(O(2)) (Pa(O(2))), and alveolar-arterial oxygen pressure difference (AaD(O(2))) at minute 3 of exercise did not differ between conditions. Mean PTT at minute 3 of exercise was 0.3 sec longer in the I condition (P=0.002). However, the change in PTT between conditions was not correlated to the change in either Pa(O(2)) or AaD(O(2)). We conclude that PVE slows (lengthens) PTT without affecting pulmonary gas exchange. Therefore, rapid PTT may not be related to hypoxemia during exercise. PMID- 12126928 TI - CO-induced K(+) currents in rat glomus cells are insensitive to light unlike carotid body neural discharge and Vo(O(2)). AB - The hypothesis that the light sensitive properties of CO-induced chemosensory nerve (CSN) discharge and oxygen consumption of the carotid body (CB) were shared by the pre-synaptic glomus cells was tested. The light effect on K(+) currents were measured before and during perfusion of the isolated rat glomus cells with high P(CO) of 550 Torr during nomoxia (P(O(2)approximately equal 100 Torr) at extra-cellular pH 7.0 and intracellular pH 6.8 with HEPES buffer. CO increased the K(+) currents with a left ward shift of the reversal potential, which showed no light effect. Thus the K(+) permeability of the glomus cell membrane were not shared by the light-sensitive CSN discharge of the CB and oxygen consumption in the presence of high P(CO.) PMID- 12126929 TI - Immune consequences of mutations in the human common gamma-chain gene. AB - A mutation (c.878T>A) in the common gamma chain (gamma(c)) causes an X-linked combined immunodeficiency (XCID) in a large kindred of British origin. In the disease, gamma(c) is expressed, but its binding to Jak3 is reduced. The immune deficiencies and clinical course were less marked in toddlers and school age children with XCID(L293Q) than in severe combined immunodeficiency (SCID). However, affected newborns were profoundly deficient in thymic size and T cells. In some affected infants, thymic size and numbers of T cells gradually increased during the first year. Their clinical course was relatively benign. In affected infants of one lineage, the number of blood T cells failed to increase substantially. They succumbed to opportunistic infections. T cell deficiencies in XCID(L293Q) progressively worsened during adolescence. Decreased thymic function, failure to rescue T cells from apoptosis, and replication senescence were possible causes. Blood T cells with the phenotype CD45RA(+)CD62L(+) (unstimulated T cells) were most depressed. CD4(+) T cells were also deficient in a specific marker of recent thymic emigrants, episomal DNA deletion circles created during TcR gene rearrangements. Apoptosis of T cells was increased, but neither apoptosis nor cell death was age-related. In contrast, telomere shortening in T cells increased with age. Unlike murine gamma(c) gene deletions, gamma delta T cells were prominent in affected adolescents and young adults. Furthermore, T cells with a V delta 2/V gamma 9 specificity declined with age and were replaced in the oldest male with a V delta 1 specificity. Thus, the mutation provides many insights concerning the role of gamma(c) in the biology of T cells. PMID- 12126930 TI - Characterization of the human gene encoding alpha-aminoadipate aminotransferase (AADAT). AB - In mammals, the conversion of alpha-aminoadipate to alpha-ketoadipate by alpha aminoadipate aminotransferase (AADAT) is an intermediate step in lysine degradation. A gene encoding for alpha-aminoadipate aminotransferase and kynurenine aminotransferase activities had been previously identified in the rat (KAT/AadAT). We identified the human gene (AADAT) encoding for AADAT. It has a 2329 bp cDNA, a 1278 bp open-reading frame, and is predicted to encode 425 amino acids with a mitochondrial cleavage signal and a pyridoxal-phosphate binding site. AADAT is 73% and 72% identical to the mouse and rat orthologs, respectively. The genomic structure spans 30 kb and consists of 13 exons. FISH studies localized the gene to 4q32.2. Two transcripts (approximately 2.9 and approximately 4.7 kb) were identified, with expression highest in liver. Bacterial expression studies confirm that the gene encodes for AADAT activity. The availability of the DNA sequence and enzyme assay will allow further evaluation of individuals suspected to have defects in this enzyme. PMID- 12126931 TI - Detachment of glycolytic enzymes from cytoskeleton of Lewis lung carcinoma and colon adenocarcinoma cells induced by clotrimazole and its correlation to cell viability and morphology. AB - Cancer cells are characterized by a high rate of glycolysis, which is their primary energy source. Glycolysis is known to be controlled by allosteric regulators, as well as by reversible binding of glycolytic enzymes to cytoskeleton. We report here that clotrimazole (l-(alpha-2 chlorotrityl)imidazole), the antifungal azole derivative, which was recently recognized as calmodulin antagonist, induced a dose-dependent detachment of the glycolytic enzymes, phosphofructokinase (ATP: D-fructose-6-phosphate 1 phosphotransferase, EC 2.7.1.11) and aldolase (D-fructose-l,6-bisphosphate D glyceraldehyde-3-phosphate-lyase, EC 4.1.2.13), from cytoskeleton of LL/2 Lewis lung carcinoma cells and CT-26 colon adenocarcinoma cells. The detachment of glycolytic enzymes from cytoskeleton would reduce the provision of local ATP, in the vicinity of the cytoskeleton membrane, and would also affect cytoskeleton structure and cell shape. We show here that clotrimazole decreased the viability of LL/2 Lewis lung carcinoma cells and CT-26 colon adenocarcinoma cells. After 3h of incubation with clotrimazole, complete cell destruction was detected. Ultrastructural cell damage was manifested by disintegration of the outer membrane by scanning electron microscopy (SEM). The detachment of glycolytic enzymes from cytoskeleton, induced by clotrimazole, preceded the decrease in cell viability, which indicates that this is an early effect and not a result of cell death. Since the cytoskeleton is being recognized as an important modulator of cell function, proliferation, differentiation, and neoplasia, detachment of the glycolytic enzymes from cytoskeleton induced by clotrimazole, as well as its reported inhibitory action on cell proliferation, makes this drug the most promising agent in the treatment of cancer. PMID- 12126933 TI - 3-Methylglutaconic aciduria type III in a non-Iraqi-Jewish kindred: clinical and molecular findings. AB - Type III 3-methylglutaconic aciduria (MGA) (MIM 258501) consists of early bilateral optic atrophy, later development of spasticity, extrapyramidal dysfunction and occasionally cognitive deficits, and urinary excretion of 3 methylglutaconic acid and 3-methylglutaric acid. The presence of the disorder in an Iraqi-Jewish genetic isolate led to mapping of the OPA3 gene to chromosome 19q13.2-q13.3, followed by isolation of the gene itself. OPA3 consists of two exons and codes for a peptide of 179 amino acids. Iraqi-Jewish patients with type III MGA are homozygous for a splice site founder mutation in OPA3 (IVS1-1G>C) which abolishes mRNA expression in fibroblasts. Here we report a novel mutation in OPA3 (320-337del) in a Kurdish-Turkish patient with optic atrophy and 3 methylglutaconic and 3-methylglutaric aciduria, previously carrying the diagnosis of type IV MGA. We conclude that type III MGA occurs in patients of non-Iraqi Jewish ancestry, and should be considered in patients with type IV MGA that have optic atrophy and ataxia. PMID- 12126932 TI - Ex vivo localization of the mouse saposin C activation region for acid beta glucosidase. AB - Saposin C is a biological activator of acid beta-glucosidase (GCase), the lysosomal hydrolase with activity towards glucosylceramide (GC). In addition, saposin C possesses a functional domain that determines the in vitro and ex vivo neuritogenic effects of prosaposin, the precursor of saposins A, B, C, and D. The domains for enzymatic activation and neuritogenic function segregate in vitro, respectively, to the carboxyl- and amino-terminal halves of human and mouse saposin C. A chimeric mouse saposin C(1-8)B(8-28)C(30-80) was created to obliterate the neuritogenic region by substituting amino acids 9-29 of saposin C with amino acids 8-28 of saposin B. This saposin showed normal in vitro enzymatic activation effects toward GCase, but no neuritogenic activity. An altered prosaposin was made to contain the chimeric saposin C region. Expression of this altered or wild-type prosaposin was driven by the PGK-1 promoter as a transgene in prosaposin knock-out mice. In cultured fibroblasts from such mice, expressed saposins localized to the lysosomal compartments. Metabolic lipid labeling using L-[3-(14)C]serine showed retention or clearance of GC in prosaposin deficient or transgene reconstituted cells, respectively. In addition, sulfatide catabolism, that requires saposin B and arylsulfatase, was also normalized in prosaposin KO cells reconstituted with the transgenes. These data show that the transgenic prosaposins were expressed and processed to functional saposins in fibroblasts. These results also show that the enzymatic activation domain is located at carboxyl-terminal half of saposin C and functions only in the context of the general saposin structure. PMID- 12126935 TI - The kyphoscoliotic type of Ehlers-Danlos syndrome (type VI): differential effects on the hydroxylation of lysine in collagens I and II revealed by analysis of cross-linked telopeptides from urine. AB - The kyphoscoliotic type of Ehlers-Danlos syndrome (EDS type VIA) (OMIM 225400) is an autosomal recessive connective tissue disorder that results from mutations in the lysyl hydroxylase 1 gene (PLOD1) causing underhydroxylation of lysine residues in tissue collagens, particularly of skin. Previous studies have shown that the pool of collagen cross-linking amino acids, hydroxylysyl pyridinoline (HP) and lysyl pyridinoline (LP) excreted in urine has an abnormally low HP/LP ratio, which is diagnostic of the condition. Here we isolated cross-linked peptides containing these residues from the urine of a child with EDS VIA homozygous for a mutation that results in a stop codon and effective null expression of PLOD1 enzyme activity. Peptides that had originated from bone type I collagen and cartilage type II collagen were identified. A cross-linked N telopeptide fraction that is derived from bone type I collagen contained only LP, no HP, which means that the helical lysines at residues 930 of alpha 1(I) and 933 of alpha 2(I) of the collagen triple-helix had not been hydroxylated. The equivalent peptide fraction from a normal child's urine gave a ratio of HP to LP of 1.5:1 typical for normal bone collagen. A second cross-linked peptide that is derived from the C-telopeptide domain of cartilage type II collagen showed both HP and LP in a 2:1 ratio, compared with 18:1 for the equivalent peptide from a normal child's urine. The results show that in EDS VIA, bone type I collagen is more markedly underhydroxylated than cartilage type II collagen, at least at those helical sites that form cross-links. The residual fraction of HP found in the urine of EDS VI patients therefore appears to be contributed in significant part by the degradation products of cartilage. Since PLOD1 is null, other PLOD genes must be responsible for the helical hydroxylation activity that results in HP. The presented approach of analyzing urinary cross-linked C-telopeptide fragments of type II collagen may allow the detection of chondrodysplasias due to genetic defects in lysyl hydroxylase isoforms active in cartilage. PMID- 12126934 TI - Two newly identified genomic mutations in a Japanese female patient with fructose 1,6-bisphosphatase (FBPase) deficiency. AB - Fructose-1,6-bisphosphatase (FBPase) (EC 3.1.3.11) catalyzes the splitting of fructose-1,6-bisphosphate into fructose 6-phosphate and inorganic phosphate. FBPase deficiency is an autosomal recessive inherited disorder caused by distraction of the fructose-1,6-bisphosphatase 1 gene (FBP1) and features severely impaired gluconeogenesis. We studied a female patient with typical FBPase deficiency symptoms. The FBPase activity of her peripheral white blood cells was undetectable. Genetic analyses of FBP1 revealed her to be a compound heterozygote of two new mutations F194S and P284R. Gene tracking in the family revealed the mother to be a heterozygote of F194S, and the father and a sister to be heterozygotes of P284R. As both Phe194 and Pro284 of FBPase are highly conserved in many species and close to crucial amino acid residues to FBPase functions, these mutations could be responsible for the loss of FBPase activities. PMID- 12126936 TI - Hearing loss in the laminin-deficient dy mouse model of congenital muscular dystrophy. AB - Sensorineural hearing loss is found in many inherited forms of muscular dystrophy. We investigated the dy mouse model, which has congenital muscular dystrophy due to a defect in laminin alpha 2, for evidence of cochlear dysfunction. Auditory brainstem response (ABR) audiometry to pure tones was used to evaluate 3-month-old homozygous dy/dy and age-matched C57 control mice. The average ABR thresholds to tone-burst stimuli for four frequencies (4, 8, 16, and 32 kHz) were determined and statistically compared by ANOVA. The dy/dy mice demonstrated elevated auditory thresholds ranging from 25 to 27 dB at each frequency tested (p<0.0001). Anatomic evaluations of the ears revealed pathology ranging from extensive connective tissue infiltration within the inner ear to possible minor defects in the cells of the organ of Corti. These anatomic and physiologic observations suggest that the extracellular matrix protein laminin plays a crucial role in normal cochlear function. Furthermore, the dy congenital muscular dystrophy mouse offers a novel model for evaluation of sensorineural hearing loss associated with muscular dystrophy. PMID- 12126937 TI - Effects of selective estrogen receptor modulators (SERMs) on coactivator nuclear receptor (NR) box binding to estrogen receptors. AB - Coactivators are required for activation of target genes by nuclear receptors. A well-studied class of coactivators, the p160 proteins, use short nuclear receptor interaction domains (NR boxes) to bind to the activated ligand-binding domain of a nuclear receptor. To investigate how selective estrogen receptor modulators (SERMs) affect NR box recruitment, we compared the recruitment of p160 NR box peptides to the estrogen receptor (ER)alpha and ER beta in the presence of 17beta estradiol (E2), 4-OH tamoxifen (4-OH Tam), LY 117018 (a raloxifene analog), and ICI 182780 (ICI, an ER antagonist). Our coactivator interaction assay utilizes time-resolved fluorescence technology to assess the binding of the 10 NR boxes derived from the three known p160 coactivators (SRC-1, -2, -3) to the ER subtypes in the presence of each ligand. The SERMs we studied did not increase NR box binding to either ER alpha or ER beta, but instead were potent antagonists decreasing estradiol-dependent NR box binding. We also demonstrated inverse agonism for all of the SERMs tested as they dose-dependently decreased hormone independent NR box binding to ER beta. Therefore, the SERMs studied behave as antagonists of ER alpha and ER beta NR box binding and do not increase coactivator NR box binding to either ER subtype. In addition, we examined the preference of E2-bound ER alpha and ER beta for various naturally occurring NR boxes including the 10 SRC boxes as well as the motifs from PGC-1, TRBP, TRAP220, and CBP. Interestingly, a clear preferential pattern of interaction was noted that was receptor specific. PMID- 12126938 TI - Effect of pirfenidone on the pulmonary fibrosis of Hermansky-Pudlak syndrome. AB - Hermansky-Pudlak syndrome (HPS) consists of oculocutaneous albinism, a platelet storage pool deficiency and, in patients with HPS1 gene mutations, a progressive, fatal pulmonary fibrosis. We investigated the safety and efficacy of an antifibrotic agent, pirfenidone (800 mg, t.i.d.), in treating 21 adult Puerto Rican HPS patients, including 20 homozygous for the same HPS1 mutation. Patients were examined every 4 months for up to 44 months in a randomized, placebo controlled trial, with rate of change in pulmonary function values as outcome parameters. Using the complete data set of 130 patient admissions, a repeated measures model showed that 11 pirfenidone-treated patients lost FVC at a rate 5% of predicted ( approximately 400 mL) per year slower than 10 placebo-treated patients (p=0.001). A random coefficients model showed no significant difference. However, using data restricted to patients with an initial FVC >50% of predicted, both models showed the pirfenidone group losing FVC (p<0.022), FEV(1) (p<0.0007), TLC (p<0.001), and DL(CO) (p<0.122) at a rate approximately 8%/year slower than the placebo group. Clinical and laboratory side effects were similar in the two groups. Pirfenidone appears to slow the progression of pulmonary fibrosis in HPS patients who have significant residual lung function. PMID- 12126939 TI - Novel mutations in the P-protein (glycine decarboxylase) gene in patients with glycine encephalopathy (non-ketotic hyperglycinemia). AB - Eight novel mutations were found in the P-protein (glycine decarboxylase) gene (GLDC) of the glycine cleavage system (EC 2.1.1.10) by screening five exons of the gene in patients with glycine encephalopathy (NKH). The mutations identified were of eight single base changes: a one-base deletion 1054del A, a splice site mutation IVS18-2A-->G and six amino acid substitutions A283P, A313P, P329T, R410K, P700A, and G762R. PMID- 12126941 TI - Fish oil (polyunsaturated fatty acid) prevents ischemic-induced injury in the mammalian retina. AB - The long-chain polyunsaturated omega-3 fatty acid docosahexaenoic (22:6n-3) acid (DHA) accumulates in rod outer segment disks and synaptic terminals. It has been thought to play an important role in disordering disk membranes and in providing an adequate environment for conformational rhodopsin changes and in modifying the activity of retinal enzymes. The decrease of DHA content in the retina has been shown to affect visual function in monkey. In rat retina, prolonged light exposure has produced reduction of DHA content in rod outer segments. The authors found that when DHA was administered before ischemia, it diminished pressure induced retinal damage. The recovery of electroretinographic amplitudes in DHA pretreated eyes was significantly greater than those in the control eyes after 4 hr of reperfusion. The histopathologic study of control eyes showed cell swelling and cell nuclei pyknosis in the inner nuclear layer after 4 hr of reperfusion and in TUNEL-positive cells in the inner and outer nuclear layers after 24-72 hr of reperfusion. The DHA pre-treated eyes had fewer pyknotic nuclei and vacuolated spaces in the inner nuclear layer and no TUNEL-positive cells for up to 72 hr of reperfusion. The precise role of the polyunsaturated n-3 fatty acid has not been identified in the retina and other tissues. Our findings showed that DHA probably prevented sensory retina from ischemic-reperfusion cell damage not only by inhibiting the formation of hydroxyl radicals, but also by reducing the non-NMDA responses or the inflammatory responses. PMID- 12126940 TI - Bcl-2 overexpression increases survival in human retinal pigment epithelial cells exposed to H(2)O(2). AB - The integrity of the retinal pigment epithelium, especially that of the macula is essential for the preservation of vision into old age. The chronic exposure to sunlight and peroxidized lipids from phagocytized photoreceptor outer segments imposes a high level of oxidative stress on the retinal tissues, which increases with age as antioxidant protection declines and therefore could accelerate apoptosis. Bcl-2 known to facilitate mitochondrial DNA repair and cellular survival in other tissues was overexpressed in a single clone of human retinal pigment epithelium cells after stable transfection with humanbcl-2 in rhoSFV neoexpression factor. Near confluent cells (2nd-4th generation permanently bcl-2 transfected) were protected from mitochondrial dysfunction after exposure to H(2)O(2) up to 150 microM. With 200 microM H(2)O(2), function in transfected cells declined by only 25% control activity as determined by MTT reduction assays, compared to wild type and vector only transfected cells expressing normal bcl-2 levels. Similarly the bcl-2 -transfected cells were more resistant to mitochondrial DNA damage after H(2)O(2) treatment than the other groups and suffered 50% less damage after exposure to 200 microM H(2)O(2), as assayed by quantitative polymerase chain reaction assays. These data suggest that bcl-2 overexpression protects human RPE cells from mitochondrial respiratory dysfuction, mitochondrial DNA damage and promotes cellular survival in response to oxidative stress induced by H(2)O(2). PMID- 12126942 TI - Anisotropic water transport in the human eye lens studied by diffusion tensor NMR micro-imaging. AB - We report in vitro measurements of effective diffusion tensors characterising the anisotropic transport of water in human eye lenses ranging in age from 13 to 86 years. The measurements were obtained by means of a pulsed field gradient spin echo (PFGSE) magnetic resonance imaging (MRI) technique at a spatial resolution of 218 x 218 x 1000 microm(3). The results show that water diffusion is both spatially inhomogeneous and highly anisotropic on the timescale of the measurements (approximately 15 msec). Diffusion parallel to the long axes of the lens fibre cells is relatively unrestricted, whereas that between cells is substantially inhibited by the cell membranes, particularly in the inner cortex region of the lens. The data confirm the presence of a diffusion barrier surrounding the lens nucleus, which inhibits transport of water and other small molecules into and out of the nucleus. The results shed light on factors that may influence the onset of presbyopia and senile cataract. They also have implications for delivery of drugs to the lens nucleus. PMID- 12126943 TI - Fourier analysis of cytoplasmic texture in nuclear fiber cells from transparent and cataractous human and animal lenses. AB - Comparisons were made of the cytoplasmic textures in electron microscope images of nuclear fiber cells from a variety of human and animal lenses. The goals were to establish the optimal conditions for quantifying the textural features and for relating the extent of roughness with the observed extent of nuclear opacification. Freshly cut Vibratome sections were fixed and processed for thin section electron microscopy. Normal human donor lenses, human age-related cataracts from surgery, and rat, guinea pig, and canine lenses were analyzed using density linescans, Fourier transforms, and autocorrelation analysis. Normal and control lenses were compared to lenses with varying degrees of scattering including fully opaque nuclear cataract. Images were recorded at 21,000 x, giving structural information in the critical range of 2-300 nm. Human normal and nuclear cataractous lens cytoplasm produce Fourier transforms with relatively high intensity in the range 10-50 nm (equivalent spacing) and relatively low intensity greater than 100 nm. This is consistent with the smooth image appearance, linescans with small fluctuations and autocorrelation functions indicating that the images are nearly homogeneous. Images of the transparent animal lenses were very smooth and produced Fourier transforms that showed less intensity in the range 10-50 nm and less intensity greater than 100 nm compared to the human lenses. Animal lenses with progressively enhanced light scattering showed a strong correlation between increased textural roughness and increased Fourier intensity greater than 100 nm. These analytical image analysis techniques readily documented the wide range of cytoplasmic textural variations in human and animal lenses and cataracts. Consistent comparisons were possible only when well preserved tissues were examined with high-resolution images. The cytoplasm with the greatest roughness correlated with the greatest light scattering suggests that redistribution and/or loss of cytoplasmic proteins contribute to cataract formation. PMID- 12126944 TI - Studies of the mitogen-activated protein kinases and phosphatidylinositol-3 kinase in the lens. 1. The mitogenic and stress responses. AB - The current understanding of the cellular signal transduction system is that cells initially respond to outside stimuli, such as growth factors or neurotransmitters, through ligand binding to the respective growth factor receptors or the G-protein-coupled receptors, to initiate transduction of the stimulus. This is followed by a series of association-dissociation and phosphorylation-dephosphorylation processes among the components of a well defined and intricate infrastructure between the cell membrane and the cytosolic protein kinases to activate and initiate nuclear target genes for cell proliferation, differentiation and other cellular functions. Although some past reports have indicated this signaling machinery is present in the lens, certain pathways, namely the mitogen-response pathway (Raf-MEK-ERK cascade), the stress response pathways (p38 and SAPK/JNK cascades) and the survival pathway (PI-3K Akt), have not been thoroughly explored in an intact lens. These pathways were studied using porcine lenses cultured under mitogenic (10 ngml(-1) growth factor) or osmotic stress (30 mM galactose) conditions to examine the cellular response in the epithelial layer, using unstimulated lenses as controls. It was found that all the key members in the Raf-MEK-ERK cascade and PI-3K-Akt cascade were present and that growth factors had a differential stimulatory effect on them. Basic-FGF was the most potent stimulator for ERK followed by EGF and IGF-1, while PDGFab and VEGF were less active. The opposite was true for their stimulatory effect on PI-3K. Hyperglycemic-induced osmotic stress stimulated p38 but not SAPK/JNK, while bFGF could stimulate SAPK/JNK but not p38. Both stimuli activated the Raf MEK-ERK and PI-3K-Akt pathways. Osmotic-induced activation could be normalized using an aldose reductase inhibitor. PMID- 12126945 TI - Effects of adeno-associated virus-vectored ciliary neurotrophic factor on retinal structure and function in mice with a P216L rds/peripherin mutation. AB - Past studies have shown that acute administration of ciliary neurotrophic factor (CNTF) can prolong the survival of retinal photoreceptor cells that have undergone phototoxic injury or that express gene mutations. Adenovirus-vectored CNTF has also been effective but for all of these treatments, the effect has been transient. On the other hand, adeno-associated virus-vectored minigenes offer considerable promise for long-term survival. The authors sought to provide long term, CNTF-based protection of mouse photoreceptors expressing a dominant negative point mutation in the rds gene by using recombinant adeno-associated virus (rAAV) to deliver minigenes that code for a secreted form of CNTF.Secreted CNTF, under control of a cytomegalovirus (CMV) or chick beta actin (CBA) promoter provided long-term, panretinal rescue of photoreceptors following single injections of rAAV vectors into the subretinal compartment. Rescue was much less effective and less reproducible when the vectors were placed in the vitreous compartment. However, there were unexpected side effects that appeared to be dose related. One side effect was a change in rod photoreceptor nucleus phenotype, featuring an increase in euchromatin and an increase in nuclear size following subretinal injections but not intravitreal injections. These nuclear changes were panretinal when the putatively stronger CBA promoter was used but not panretinal when the CMV promoter was used. In the latter case, the nuclear changes were much more pronounced at the site of injection. Thus, chronic hyperstimulation of retinal cells with CNTF may up-regulate gene expression in photoreceptors. Based on current knowledge of retinal cell targets for CNTF, this effect may be indirect and may not represent direct stimulation of photoreceptors by CNTF.A second side effect was a paradoxical decrease in scotopic a- and b-wave amplitudes and a decrease in photopic b-wave amplitudes in the injected, rescued retina when compared to its contralateral, uninjected counterpart, in spite of the fact that these retinas had more photoreceptors than their untreated mates. The basis for these decreased ERG amplitudes may be related to changes in gene expression. The mechanisms for these side effects and proper doses of CNTF administration should be determined before human clinical trials are considered for the amelioration of inherited retinal degenerations with CNTF. PMID- 12126946 TI - Spinocerebellar ataxia type 7 (SCA7) shows a cone-rod dystrophy phenotype. AB - Autosomal dominant spinocerebellar ataxia 7 is associated with retinal degeneration. SCA7, the causative gene, encodes ataxin-7, a ubiquitous 892 amino acid protein of variable sub-cellular localization, and the disease is due to expansion of an unstable CAG repeat in the coding region of the gene. Recent increases in understanding of the mechanisms ofSCA7 -related retinopathy from in vitro and murine model studies prompted us to perform a detailed study of the retinal phenotype of affected members of a family with SCA7 mutation (45-47 CAG repeats). There was a spectrum of severity from mild to severe dysfunction. Early functional abnormalities were at both photoreceptor and post-receptoral levels. When cone and rod photoreceptor dysfunction was present, it was approximately equal. Regional retinal dysfunction was evident: there was more dysfunction centrally than peripherally with least effect in the midperiphery. In vivo cross sectional retinal images with optical coherence tomography showed an early disease stage of altered foveal lamination (abnormal area of low reflectivity splitting the outer retina-choroidal complex) accompanied in the parafovea by reduced retinal thickness. Later disease stages showed foveal and parafoveal retinal thinning. The phenotype in this family with SCA7 is that of a cone-rod dystrophy. These observations increase interest in a recent hypothesis that ataxin-7 may interfere with the function of CRX (cone-rod homeobox), a transcription factor regulating photoreceptor genes and a cause of a cone-rod dystrophy phenotype in man. PMID- 12126947 TI - Phenytoin blocks retinal ganglion cell death after partial optic nerve crush. AB - Phenytoin is a well-characterized sodium channel blocker in widespread use as an anticonvulsant. In 1972, Becker and co-workers reported that phenytoin could reverse visual field loss from glaucoma. The authors therefore explored whether phenytoin could protect retinal ganglion cells from optic nerve crush. The optic nerve of Long-Evans rats was partially crushed; animals were given a single dose of either intraperitoneal phenytoin or vehicle. A third group underwent sham optic nerve crush. In a second set of experiments, the effect of phenytoin was compared to the N -methyl- D -receptor antagonist, memantine. Retinal ganglion survival was evaluated 1 week later. In addition, the effect of memantine and phenytoin on glutamate-induced intracellular calcium fluxes was evaluated.Phenytoin and memantine significantly reduced ganglion cell loss after optic nerve crush, and blunted the rise in intracellular calcium seen after administration of glutamate. Co-administration of the two agents, however, did not increase ganglion cell survival, and had no effect on ganglion cell calcium fluxes. Phenytoin can preserve retinal ganglion cells after partial optic nerve crush. This effect was not additive with a glutamate antagonist, suggesting that both agents alone are equally protective at saving the same population of ganglion cells at risk. In fact, the neuroprotective effect of the combined administration of phenytoin and memantine was significantly less than either of the two drugs alone. Phenytoin is known to decrease neuronal firing and neurotransmitter release; this may underlie its ability to serve as a neuro protectant in this experimental paradigm. PMID- 12126948 TI - Systematic analysis of E-, N- and P-cadherin expression in mouse eye development. AB - Cadherins are a family of Ca(2+)-dependent cell adhesion molecules. Through their homophilic binding interactions, cadherins play important roles in tissue formation and maintenance during development. Here the authors compare the expression patterns of the three classical cadherins, E-, N- and P-cadherin, during mouse eye development from embryonic day 9.5 (E9.5) to adult. It was found that: (1) The expression patterns of N- and P-cadherin are mutually exclusive in most ocular tissues during development. N-cadherin mRNA is detected specifically in the lens placode during lens induction at E9.5, and is absent in the rest of the surface ectodermal tissues. In contrast, P-cadherin is expressed in the surface ectoderm but not in the lens vesicle. N-cadherin is expressed continuously in the lens pit, lens vesicle, and in the epithelial cells and newly differentiating fiber cells of the mature lens. P-cadherin is expressed in the epithelial cells of the cornea, eyelids and Harderian gland. Reciprocal expression patterns of N-and P-cadherins are also seen during retinal development. N-cadherin is initially expressed in both the inner and outer layers of the optic cup at E9.5. N-cadherin expression persists in the inner layer as it develops into neural retina, but is turned off in the outer layer where the cells differentiate into retinal pigment epithelial (RPE) cells and express P-cadherin. Reciprocal patterns of expression are also seen in the ciliary epithelium. N cadherin is expressed in the inner layer and P-cadherin in the outer layer of the ciliary epithelium. (2) E- and P-cadherins are epithelial cadherins. Their expression patterns in the eye are not identical. Both cadherins are found in the epithelia of the cornea, eyelid and Harderian gland. In contrast, lens epithelial cells express E- but not P-cadherin, and RPE cells express P- but not E-cadherin. (3) In addition to its high expression in surface ectoderm-derived tissues, E cadherin mRNA was also detected in some of the retinal ganglion neurons at postnatal day 14 (P14). E-cadherin expression in the neural retina has not been reported before. This study shows that cell fate determination in the eye occurs in conjunction with distinct changes in the patterns of cadherin gene expression. PMID- 12126949 TI - Increased ocular blood vessel numbers and sizes following chronic sympathectomy in rat. AB - Disease states characterized by ocular vascular pathology are often associated with impaired sympathetic function. This study examined the effect of sympathetic denervation on ocular vasculature of the adult rat. Uveal perfusion and choroidal and retinal blood vessel sizes and numbers were assessed in rats with intact innervation and after short- (2 days) or long-term (6 weeks) sympathetic denervation induced by ipsilateral superior cervical ganglion excision. In rats with intact innervation and after short-term sympathectomy, blood flow in both eyes was comparable. However, after long-term sympathectomy, blood flow was four fold greater in the denervated than in the innervated eye, but was unaltered in lacrimal gland, cerebral cortex, and masseter muscle. Choroid surface area was not affected by long-term sympathectomy, but choroidal thickness was increased and choroidal cross-sectional area occupied by vascular lumina was greater. Arteriolar number per unit cross-sectional area of choroid was not altered although arteriolar diameters were enlarged. Choroidal venules were larger and more abundant. Choroidal capillary numbers were unchanged, but retinal capillaries of the outer plexiform layer were increased. To determine if these changes result from loss of sympathetic activity, sympathetic preganglionic innervation was excised chronically. This produced significant increases in choroidal thickness and vascular luminal area, and in numbers of arterioles, small venules, and capillaries in the outer plexiform layer. These findings show that sympathetic innervation is critical in regulating choroidal and retinal vascularity, and that chronic loss of sympathetic activity may contribute to abnormal vascular proliferation in diseases such as age-related macular degeneration and diabetic retinopathy. PMID- 12126950 TI - Distribution of acetylcholine-sensitive currents around the rabbit crystalline lens. AB - The relative distribution of acetylcholine (ACh) receptors on the surface of the isolated ocular lens of the rabbit was determined from induced changes in translens short-circuit current (I(SC)) and the translenticular resistance (R(t)) at seven delineated, parallel zones from the anterior to the posterior pole. For this, one O-ring (from among several having different diameters) was used to separate two zones in a vertically arranged Ussing-type chamber. Different O rings separated different zone pairs. Earlier experiments from this laboratory used a conventional divided chamber, which occluded the equatorial surface, to demonstrate that anterior applications of ACh transiently decreased the I(SC) due to an intracellular Ca(2+) release and inhibition of anteriorly located K(+) channels. Measurements obtained with the newly designed zonal arrangement determined that the entire epithelial surface from its anterior-most aspect to the equatorial region responds electrically to ACh exposure, while the posterior most region does not. Furthermore, lens-mounting positions that resulted in separation of the epithelium so that portions of its surface were present in each hemichamber resulted in inverse current changes upon bilateral ACh addition to the bathing solutions. Reductions in outward cationic current across the anterior surface into the anterior bath upon ACh treatment were accompanied by an increase in translens resistance consistent with a closure of basolateral K(+) channels. Overall, these results suggest that the posterior fiber cells may lack ACh receptors, which are clearly present in the lens epithelium that covers about two thirds of the rabbit lens surface area, and indicate that an ACh-evoked Ca(2+) signal does not spread throughout the epithelial layer. A functional role for lens acetylcholine receptors remains to be determined. PMID- 12126951 TI - Cellular and cytoskeletal dynamics within organ cultures of porcine neuroretina. AB - The aim of this study was to establish a retinal organ culture and to follow the cellular and cytoskeletal changes. For this purpose the authors detached porcine neuroretinas from the underlying pigment epithelium, and incubated them for 2 weeks under standardized conditions. After 3, 6, 10, and 13 days in culture the retinal tissues were fixed, embedded in LR-White resin or paraffin wax and processed for electron-, light-, immunofluorescence- and confocal laser scanning microscopy. Antibodies directed against alpha-tubulin, actin, glial fibrillary acidic protein (GFAP), vimentin, neurofilament(200) and beta-catenin were used to investigate the cytoskeletal changes over a certain period of time. After experimental detachment Muller cells quickly started to hypertrophy showing increased levels of intermediate filaments (i.e. vimentin and GFAP). The actin labelling of photoreceptor cells decreased concomitantly with a rapid degeneration of the outer segments. After 1 week of detachment the Muller cell cytoplasm revealed increasing amounts of actin and tubulin staining. Actin filaments appeared frequently organized in thick bundles across the full width of the retina, whereas increasing levels of tubulin shifted into the outer nuclear region especially concentrated near the outer limiting membrane. A prolonged time of explant culturing resulted in a discontinuous staining of beta-catenin along the adherent junctions of the outer limiting membrane, followed by an outgrowth of Muller cell extensions into the subretinal space. Double staining of tubulin and cones showed that this outgrowth predominantly occurred between cone inner segments. The outer limiting membrane was penetrated by stalk-like structures, highly enriched with tubulin and associated with swollen tips, reaching into the subretina. Electron microscopy demonstrated in detail the focal disruption of the outer limiting membrane by Muller cell extensions and subsequent subretinal gliosis. The cytoskeletal reactions described here were compared with degenerative changes observed after induced retinal detachments. PMID- 12126952 TI - Chromatin structure and transcriptional regulation of the beta-globin locus. AB - Chromatin structure plays a critical role in eukaryotic gene transcriptional regulation. The beta-globin locus provides an ideal system within which to study the interplay between chromatin structure and transcriptional regulation. The process of beta-globin locus activation is remarkably intricate and involves at least two distinct events: chromatin opening and gene activation. Great progress has been made in recent years in understanding how locus control regions confer high-level expression to linked genes. Current interest focuses on some special events, including formation of locus control region hypersensitivity sites, ATP dependent chromatin remodeling, localized H3 hyperacetylation, and intergenic transcription, which link chromatin and beta-globin locus regulation. These events, and their possible molecular bases, are summarized together with speculations concerning their connections. PMID- 12126953 TI - Oxygen consumption and expression of the adenine nucleotide translocator in cells lacking mitochondrial DNA. AB - It has been shown previously that human rho degrees cells, deprived of mitochondrial DNA and consequently of functional oxidative phosphorylation, maintain a mitochondrial membrane potential, which is necessary for their growth. The goal of our study was to determine the precise origin of this membrane potential in three rho degrees cell lines originating from the human HepG2, 143B, and HeLa S3 cell lines. Residual cyanide-sensitive oxygen consumption suggests the persistence of residual mitochondrial respiratory chain activity, about 8% of that of the corresponding parental cells. The fluorescence emitted by the three rho degrees cell lines in the presence of a mitochondrial specific fluorochrome was partially reduced by a protonophore, suggesting the existence of a proton gradient. The mitochondrial membrane potential is maintained both by a residual proton gradient (up to 45 to 50% of the potential) and by other ion movements such as the glycolytic ATP(4-) to mitochondrial ADP(3-) exchange. The ANT2 gene, encoding isoform 2 of the adenine nucleotide translocator, is overexpressed in rho degrees HepG2 and 143B cells strongly dependent on glycolytic ATP synthesis, as compared to the corresponding parental cells, which present a more oxidative metabolism. In rho degrees HeLa S3 cells, originating from the HeLa S3 cell line, which already displays a glycolytic energy status, ANT2 gene expression was not higher as in parental cells. Mitochondrial oxygen consumption and ANT2 gene overexpression vary in opposite ways and this suggests that these two parameters have complementary roles in the maintenance of the mitochondrial membrane potential in rho degrees cells. PMID- 12126954 TI - E-cadherin and the differentiation of mammalian vestibular hair cells. AB - E-cadherin is expressed in vestibular, mechanosensory epithelia during early embryonic development. During late embryonic and neonatal stages it is expressed in supporting cells but down-regulated in differentiating sensory hair cells. We used a conditionally immortal cell line (UB/UE-1) from the neonatal mouse utricle to test the hypothesis that constitutive expression of E-cadherin inhibits the progression of hair cell differentiation. Under differentiating culture conditions, transfected E-cadherin inhibited expression of the cytoskeletal protein myosin VIIa and functional expression of both acetylcholine receptors and potassium channels, which are normally expressed by neonatal hair cells. However, it had no effect on the expression of the transcription factor Brn3c or the cytoskeletal protein fimbrin, which are also expressed by neonatal hair cells. The number of adherens junctions increased significantly under differentiating conditions but there was no detectable change in formation of tight junctions or gap junctions. However, E-cadherin expression led to density-dependent cell death under differentiating conditions. We have shown that E-cadherin is expressed in vestibular supporting cells, which form the basis of the sensory epithelium, but that constitutive expression inhibits the full differentiation of hair cells. Down-regulation of E-cadherin is thus likely to be a key element in the regeneration of hair cells. PMID- 12126955 TI - Mouse Notch 3 expression in the pre- and postnatal brain: relationship to the stroke and dementia syndrome CADASIL. AB - Mutations in the human Notch 3 gene cause the vascular stroke and dementia syndrome CADASIL (Cerebral Autosomal Dominant Arteriopathy with Subcortical Infarcts and Leukoencephalopathy) characterized by degeneration of vascular smooth muscle cells and multiple small infarcts in the white and deep gray matter of the brain. Here we have analyzed the expression pattern of the Notch 3 gene in the pre- and postnatal mouse brain. Prenatal Notch 3 expression is restricted to a scattered population of cells within the vessel wall of all major blood vessels in the developing embryo, including those that form the perineural vascular plexus. Expression in the postnatal brain is confined to a scattered cell population within the vessel wall of small to medium-sized penetrating arteries, which are the vessel type primarily affected in CADASIL patients. In contrast, no expression was observed in capillaries and veins. Notch 3 is most likely expressed in a subset of vascular smooth muscle cells, and the expression pattern of one of the Notch ligands, Serrate 1, was very similar to that observed for Notch 3. The Notch 3 expressing pattern was not significantly altered in platelet derived growth factor B- (PDGF-B) deficient mouse embryos, demonstrating that Notch 3 expression is not under direct control of PDGF-B. These data show that Notch 3 expression is conserved between mouse and human and suggest that the mouse is a valid system for analysis of CADASIL. PMID- 12126956 TI - ROCK-II-induced membrane blebbing and chromatin condensation require actin cytoskeleton. AB - Ectopic expression of ROCK II (Rho kinase II or ROKalpha), an effector of Rho GTPase, induces membrane blebbing and chromatin condensation. ROCK II can induce membrane blebbing in the presence of the caspase inhibitor z-VAD-fmk or in caspase-3-deficient MCF-7 cells, indicating that the activation of caspases is not required. ROCK-II-induced membrane blebbing, however, is reversed by the myosin light chain kinase inhibitor ML-7 or cytochalasin D. In addition, the expression of a constitutively activated form of cofilin (S3A-cofilin) suppresses both membrane blebbing and chromatin condensation in ROCK II expressing cells. These findings suggest that the activation of actin-myosin contractility is responsible for membrane blebbing and chromatin condensation and implicate ROCK II as a potential mediator of the morphological changes associated with apoptosis. PMID- 12126957 TI - Hypoxia stimulates p16 expression and association with cdk4. AB - Exposure of CV-1P cells to hypoxic conditions causes cell proliferation inhibition concomitant with the accumulation of pRb in the hypophosphorylated, growth suppressive form. This is in part due to inhibition of pRb-directed cdk4 and cdk2 activity. In this study we attempted to elucidate the mechanism by which cdk4 is inactivated under hypoxic conditions. After 18 h of hypoxia, CV-1P cells are inhibited from progressing from G(1) phase into the S phase of the cell cycle. This occurs in conjunction with dephosphorylation of serine-795, which is a putative substrate of cdk4. The amounts of cdk4, cdk6, and the D type cyclins are not affected by 18 h of hypoxia. The levels of cdki p16, p18, p19, and p57 under aerobic or hypoxic conditions were analyzed and although the levels of most cdki are unaffected by hypoxic conditions, the level of p16 increases significantly by 18 h of hypoxia. The mechanism by which cdk4 activity is inhibited under hypoxic conditions may be mediated through p16 association with cdk4. Immunoprecipitation analysis shows that p16 binds to cdk4 under hypoxic conditions but does not in cells maintained under aerobic conditions. Thus p16 may be involved in hypoxia-induced growth inhibition. PMID- 12126958 TI - Activation of caspases and serine proteases during apoptosis induced by onconase (Ranpirnase). AB - Onconase (ONC) is a ribonuclease isolated from amphibian oocytes that is cytostatic and cytotoxic to numerous tumor lines. ONC shows in vivo anti-tumor activity in mouse tumor models and is currently in Phase III clinical trials. Previous studies indicated that ONC induces apoptosis of the target cells most likely along the mitochondrial pathway involving caspase-9 as the initiator caspase. We have recently developed an approach to detect the activation of serine (Ser) proteases during apoptosis. The method is based on affinity labeling of Ser protease active centers with fluorochrome-tagged inhibitors. The aim of the present study was to reveal whether Ser proteases are activated during apoptosis induced by ONC. Human leukemic HL-60 cells were treated with ONC for up to 72 h and then exposed to 5(6)-carboxyfluoresceinyl-L-phenylalanylchloromethyl ketone (FFCK) or 5(6)-carboxyfluoresceinyl-L-leucylchloromethyl ketone (FLCK), the fluorescing green reagents reactive with active centers of the chymotrypsin like enzymes that cleave proteins at the Phe (FFCK) or Leu (FLCK) site. Activation of caspases was assayed in the same cells using sulforhodamine-labeled (fluorescing red) pan-caspases inhibitor (SR-VAD-FMK). Administration of 1.67 microM ONC into cultures of HL-60 cells led to the appearance of cells that bound SR-VAD-FMK as well as FFCK and FLCK. Most labeled cells had features characteristic of apoptosis. We interpret the binding of these ligands, which was irreversible and withstood cell fixation, as revealing activation of caspases and chymotrypsin-like Ser proteases. Because the induction of binding of each of the three ligands occurred at approximately the same time, the data suggest that during apoptosis caspases and Ser proteases may transactivate each other. The intercellular and subcellular pattern of binding SR-VAD-FMK vs FFCK or vs FLCK was different indicating a variability in abundance and localization of these enzymes within individual apoptotic cells. The FFCK- and FLCK-reactive proteins were of similar molecular mass, approximately 59 and approximately 57 kDa, respectively. PMID- 12126959 TI - Regulation of smooth muscle actin expression and contraction in adult human mesenchymal stem cells. AB - Prior studies have demonstrated the expression of a contractile actin isoform, alpha-smooth muscle actin, in bone marrow stromal cells. One objective of the current study was to correlate contractility with alpha-smooth muscle actin expression in human bone marrow stroma-derived mesenchymal stem cells. A second objective was to determine the effects of transforming growth factor-beta1, platelet derived growth factor-BB, and a microfilament-modifying agent on alpha smooth muscle actin expression and alpha-smooth muscle actin-enabled contraction. Adult human bone marrow stromal cells were passaged in monolayer and their inducibility to chondrocytic, osteoblastic, and adipogenic phenotypes was demonstrated. Western blot analysis was employed along with densitometry to quantify the alpha-smooth muscle actin content of the cells and their contractility was evaluated by their contraction of a type I collagen glycosaminoglycan sponge-like matrix into which they were seeded. Transforming growth factor-beta1 (1 ng/ml) significantly increased and platelet-derived growth factor-BB (10 ng/ml) decreased alpha-smooth muscle actin expression and the contractility of the cells. Cytochalasin D also blocked cell contraction. There was a notably high correlation of cell-mediated contraction normalized to the DNA content of the matrices with alpha-smooth muscle actin content of the cells by linear regression analysis (R(2) = 0.88). These findings lay the groundwork for considering the role of alpha-smooth muscle actin-enabled contraction in mesenchymal stem cells and in their connective tissue cell progeny. PMID- 12126960 TI - Properties of primary mouse myoblasts expanded in culture. AB - Implantation of myoblasts is a strategy used to enhance the regeneration of skeletal muscle tissue in vivo. In mouse models, myogenic cell lines and primary cells have been employed with different yields of adult muscle tissue formed. The present work is a study of some developmental features of expanded primary mouse myoblasts (i28), which have been shown to form muscle tissue. i28 myoblasts were differentiated in vitro and the expression of acetylcholine receptor channels and maturation of the excitation-contraction coupling mechanism were investigated using patch clamp and videoimaging techniques. In all the developing cells the embryonic isoform of the acetylcholine receptors was present. Skeletal muscle type excitation-contraction coupling (i.e., a mechanical link between voltage dependent calcium channels and ryanodine receptor channels) was detected in about 75% of differentiating i28 myotubes. Only these cells showed spontaneous changes in cytosolic free calcium concentration associated with twitches. Our findings are the first description of the physiological properties of expanded primary myoblasts which are used for implantation and confirm that they are a heterogeneous cell population. In comparison to permanent cell lines, the Ca(2+) signaling is more similar to that described in mature nonexpanded muscle fibers. This suggests that cultured primary cells are, so far, the most suitable cell type for muscle regeneration. PMID- 12126961 TI - Extracellular matrix- and cytoskeleton-dependent changes in cell shape and stiffness. AB - Cell spreading is correlated with changes in important cell functions including DNA synthesis, motility, and differentiation. Spreading is accompanied by a complex reorganization of the cytoskeleton that can be related to changes in cell stiffness. While cytoskeletal organization and the resulting cell stiffness have been studied in motile cells such as fibroblasts, less is known of these events in nonmigratory, epithelial cells. Hence, we examined the relationship between cell function, spreading, and stiffness, as measured by atomic force microscopy. Cell stiffness increased with spreading on a high density of fibronectin (1000 ng/cm(2)) but remained low in cells that stayed rounded on a low fibronectin density (1 ng/cm(2)). Disrupting actin or myosin had the same effect of inhibiting spreading, but had different effects on stiffness. Disrupting f-actin assembly lowered both stiffness and spreading, while inhibiting myosin light chain kinase inhibited spreading but increased cell stiffness. However, disrupting either actin or myosin inhibited DNA synthesis. These results demonstrate the relationship between cell stiffness and spreading in hepatocytes. They specifically show that normal actin and myosin function is required for hepatocyte spreading and DNA synthesis and demonstrate opposing effects on cell stiffness upon disruption of actin and myosin. PMID- 12126962 TI - Bcl-2 phosphorylation and proteasome-dependent degradation induced by paclitaxel treatment: consequences on sensitivity of isolated mitochondria to Bid. AB - Several studies have suggested that Bcl-2 phosphorylation, which occurs during mitotic arrest induced by paclitaxel, inhibits its antiapoptotic function. In the present study, we demonstrated that the level of phosphorylated Bcl-2 was threefold higher in mitochondria than in the nuclear membrane or endoplasmic reticulum. Our results show, in isolated mitochondria, that phosphorylation of Bcl-2 in mitosis does not modify either its integration into the mitochondrial membrane or the ability to release cytochrome c in response to Bid, a cytochrome c releasing agent. In HeLa cells, in which paclitaxel induces apoptosis, the nonphosphorylated form of Bcl-2 is degraded by a proteasome-dependent degradation pathway, whereas the phosphorylated forms of mitochondrial Bcl-2 appear to be resistant to proteasome-induced degradation. We found that low concentrations of recombinant Bid triggered a greater release of cytochrome c from mitochondria isolated from paclitaxel-treated HeLa cells than from mitochondria isolated from control HeLa cells. Taken together, these results show that Bcl-2 phosphorylation does not inhibit its function. On the contrary, Bcl-2 phosphorylation indirectly regulated its antiapoptotic action via protection against degradation. Indeed, in response to paclitaxel treatment, the level of Bcl-2 expression in mitochondria rather than its phosphorylation state could regulate the sensitivity of mitochondria to cytochrome c releasing agents in vitro. PMID- 12126963 TI - Spatiotemporal regulation of moesin phosphorylation and rear release by Rho and serine/threonine phosphatase during neutrophil migration. AB - Neutrophil motility is crucial to effective host defenses against microorganisms. While uropod retraction is a critical step in the migration of neutrophils, the underlying molecular mechanism is not well understood. Here, we show that inhibition of the Rho small GTPase with C3 exoenzyme prevented the retraction of trailing uropods, indicating that the process of rear release is mediated by a Rho signaling pathway. C3 exoenzyme caused marked elongation of directionally migrating neutrophils, suggesting an additional role for Rho in the maintenance of functional polarized cell shape. We also show that phosphorylation and dephosphorylation of the plasma membrane-actin filament cross-linker moesin are spatiotemporally controlled in migrating neutrophils. In particular, phosphorylation of moesin at threonine 558 depended on Rho activity. Videomicroscopy showed that dephosphorylation of this carboxy-terminal threonine preceded uropod retraction. Calyculin A, an inhibitor of type 1 and type 2A serine/threonine phosphatases, suppressed the moesin dephosphorylation and impaired uropod retraction in a dose-dependent manner. Cypermethrin, an inhibitor of type 2B serine/threonine phosphatase, had no such effects. The finding that Rho small GTPase and type 1/type 2A phosphatases are involved in rear release yields novel insights into the biochemical mechanisms of neutrophil migration. PMID- 12126964 TI - Angiotensin II and cAMP regulate AT(1)-mRNA expression in rat cardiomyocytes by transcriptional mechanism. AB - Mechanisms of angiotensin II and cAMP regulating the expression of angiotensin II type 1 (AT(1)) receptor mRNA were studied in neonatal rat cardiomyocytes. Angiotensin II induced a transient decrease of AT(1)-mRNA expression in time- and dose-dependent manner. Maximal decrease (49.2 +/- 9.5% of control) occurred at 6 h of angiotensin II (10 nmol/l) treatment. AT(1) receptor antagonists 4-ethyl-2-n propyl-1-[2'-(1H-tetrazol-5-yl)biphenyl-4-yl)methyl]imidazole-5-carboxylic acid (DMP811) and losartan as well as 1-(5-isoquinolinesulfonyl)-2-methylpiperazine dihydrochloride (H-7) reversed the down-regulation of AT(1)-mRNA expression. 6 h of phorbol 12-myristate 13-acetate (PMA) stimulation caused a decrease of AT(1) mRNA level. Treatment by angiotensin II plus actinomycin D for 6 h produced the same effect as actinomycin D alone. These results suggest that angiotensin II down-regulates AT(1)-mRNA level of rat cardiomyocytes by inhibiting the transcription of AT(1) gene, which is mediated by AT(1) receptor and related to the activation of protein kinase C. Stimulation by forskolin plus 3-isobutyl-1 methyl-xanthine (IBMX) decreased the expression of AT(1)-mRNA to 68.1 +/- 21.5% of control at 6 h treatment; while increased to 207.9 +/- 27.1% of control at 48 h treatment. A series of 5'-upstream deletion mutants of AT(1A) promoter were produced and then were recombined with pGL(3) basic vector utilizing luciferase as reporter gene. Among all the constructors, p(-201/+ 74)Luc was of the highest luciferase activity (5.9 times higher than control) after stimulation by forskolin for 48 h. Further deletion from -201 to -61 resulted in a large decrease of activity. These results indicate that cAMP induces a time-dependent bi-directional regulation of AT(1)-mRNA expression. The cAMP responsible element (CRE) cis-element located in the region -201/-61 of rat AT(1A) promoter is forskolin inducible, which may mediate the up-regulation of AT(1)-mRNA expression induced by cAMP long-lasting stimulation. PMID- 12126965 TI - Arsenite-induced reactive oxygen species and the repression of alpha-tocopherol in the MGC-803 cells. AB - We have investigated the action of oxidative stress in arsenite-induced apoptosis of human gastric cancer MGC-803 cells. Cells exhibited obvious characteristic of apoptosis following the treatment with 1.0 microM arsenite for 24 h. During the process, low concentration of arsenite significantly increased superoxide formation and lipid peroxidation, which was dose-dependent and was related to cell apoptosis induced by arsenite. The oxidant-dependent increase in intracellular [Ca(2+)] level and p53 gene expression were also observed at the same time. A phospholipase C inhibitor, 1-[6-([(17 beta)-3-methoxyestra 1,3,5,(10)-trien-17-yl]-amino)hexyl]-2,5-dione (U73122), could block the rapid transient increase in intracellular Ca(2+) levels, as well as the subsequent fragmentation of nuclear DNA. Addition of alpha-tocopherol before arsenite treatment abolished the transient increase in superoxide formation, lipid peroxidation, intracellular [Ca(2+)] levels and p53 gene expression, and furthermore could significantly inhibited the arsenite-induced apoptosis of MGC 803 cells. These results indicate that arsenite-induced oxidative stress, which stimulate cellular signaling systems, are involved in apoptosis of MGC-803 cells. PMID- 12126966 TI - Fibrates suppress chenodeoxycholic acid-induced RANTES expression through inhibition of NF-kappaB activation. AB - Fibrates, hypolipidemic agents, are reported to be effective in treatment of primary biliary cirrhosis. However, the mechanism involved in therapeutic benefits of fibrates in primary biliary cirrhosis remains unknown. In contrast, hepatic regulated upon activation, normal T-cell expressed and secreted (RANTES) is increased in patients with primary biliary cirrhosis and bile acids up regulate RANTES expression in hepatocytes. The role of fibrates in bile acid induced RANTES expression was investigated in human hepatoma cells; 100 microM of bezafibrate and fenofibrate decreased expression of chenodeoxycholic acid-induced RANTES mRNA and protein. In addition, luciferase enzyme assay using RANTES promoter-luciferase reporter plasmid revealed that 100 microM of bezafibrate and fenofibrate transcriptionally reduced chenodeoxycholic acid-induced RANTES gene expression. Moreover, bezafibrate clearly repressed DNA-binding activity of nuclear factor-kappaB (NF-kappaB) induced by chenodeoxycholic acid. Therefore, fibrates might be inhibitory agents of inflammatory cell migration by RANTES to the liver in patients with primary biliary cirrhosis, possibly indicating that fibrates are therapeutic agents in primary biliary cirrhosis. PMID- 12126967 TI - Nitric oxide scavenger carboxy-PTIO potentiates the inhibition of dopamine uptake by nitric oxide donors. AB - 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (carboxy-PTIO) has been increasingly used as nitric oxide (NO) scavenger. Carboxy-PTIO reacts with NO to form nitric dioxide and 2-(4-carboxyphenyl)-4,4,5,5 tetramethylimidazoline-1-oxyl (carboxy-PTI). In rat C6 glioma cells expressing human dopamine transporter, carboxy-PTIO paradoxically potentiated the inhibition of [(3)H]dopamine uptake by two NO donors, diethylamine/NO and (Z)-1-[N-(3 ammoniopropyl)-N-(n-propyl)-amino]/NO. Further examinations revealed that carboxy PTI concentration-dependently reduced dopamine uptake, indicating that the formation of carboxy-PTI may account for the failure of carboxy-PTIO to abolish NO elicited effects. These results suggest that caution should be taken in interpreting data obtained using carboxy-PTIO and probably other NO scavengers. PMID- 12126968 TI - Acute and carryover effects in mice of MDMA ("ecstasy") administration during periadolescence. AB - In spite of the increasing evidence concerning its neurotoxicity, young human individuals are often involved in the recreational use of amphetamine-type stimulants such as 3,4-methylenedioxymethamphetamine (MDMA or "ecstasy"). A study aimed to investigate short- and long-term consequences of a repeated and intermittent MDMA administration (0, 5 or 10 mg/kg i.p., 3 days treatment history) was conducted in mice. Mice were injected at different phases in development, namely at early (28 days old), middle (38 days old) or late (52 days old) adolescence. When assessed for nociceptive response, a dose-dependent analgesia was found in middle and late adolescent mice. Carryover consequences of previous MDMA treatment were then investigated at adulthood (80 days old). In a social interaction test, levels of environment exploration and social behaviour resulted markedly increased in drug-free state as a function of drug exposure during development, whereas others behaviours were reduced. MDMA challenge (5 mg/kg dose) produced the expected hyperactivity, as well as a marked increment of hypothalamic serotonin (5-hydroxyhyptamine, 5-HT) levels. Mice treated chronically with MDMA during middle and late adolescence were associated with important reductions of the indoleamine. As a whole, these results indicate a differential long-term vulnerability to behavioural and neurotoxicant effects of MDMA as a function of the developmental stage of exposure. PMID- 12126969 TI - Effect of MK-801 on the antinociceptive effect of [D-Ala(2),N-MePhe(4), Gly ol(5)]enkephalin in diabetic mice. AB - The role of N-methyl-D-aspartate (NMDA) receptors in supraspinal and spinal sites on the reduction of supraspinal micro-opioid receptor-induced antinociception in diabetic mice was examined. The antinociceptive effect of i.c.v. [D-Ala(2), N MePhe(4), Gly-ol(5)]enkephalin (DAMGO, 20 pmol) in diabetic mice was significantly less than that in non-diabetic mice. The antinociceptive effect of i.c.v. DAMGO (20 pmol) was significantly and dose dependently reduced by i.c.v. (+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine maleate (MK-801) in both non-diabetic (0.03-0.3 nmol) and diabetic mice (0.1-3.0 nmol). While the antinociceptive effect of i.c.v. DAMGO (10 pmol) was significantly enhanced by i.c.v NMDA (0.01-0.1 nmol) in non-diabetic mice, the same doses of i.c.v. NMDA had no significant effect on the antinociceptive effect of i.c.v. DAMGO (20 pmol) in diabetic mice. In non-diabetic mice, the antinociceptive effect of DAMGO (20 pmol, i.c.v.) was dose dependently reduced by intrathecal administration of MK 801 (0.1-1.0 nmol). The antinociceptive effect of DAMGO (20 pmol, i.c.v.) was dose-dependently enhanced by MK-801 (0.1-1.0 nmol, i.t.) in diabetic mice. Furthermore, NMDA (0.1 nmol, i.t.) significantly enhanced the antinociceptive effect of DAMGO (10 pmol, i.c.v.) in non-diabetic mice. However, in non-diabetic mice, the antinociceptive effect of DAMGO (40 pmol, i.c.v.) was dose dependently reduced by NMDA (0.03-0.3 nmol, i.t.). These results suggest that NMDA receptor function in supraspinal and spinal sites appear to be modulated differently by the diabetic state, and this functional modulation may play an important role in the reduction of supraspinal micro-opioid receptor-induced antinociception in diabetic animals. PMID- 12126970 TI - Facilitation of noradrenaline release by adenosine A(2A) receptors in the epididymal portion and adenosine A(2B) receptors in the prostatic portion of the rat vas deferens. AB - The adenosine-receptor modulation of noradrenaline release was compared in prostatic and epididymal portions of rat vas deferens. In both portions, tritium overflow elicited by electrical stimulation (100 pulses/8 Hz) was reduced by the adenosine A(1) receptor agonist, N(6)-cyclopentyladenosine, and enhanced by the nonselective receptor agonist, 5'-N-ethylcarboxamidoadenosine, in the presence of the adenosine A(1) receptor antagonist, 1,3-dipropyl-8-cyclopentyl-1,3 dipropylxanthine (DPCPX; 20 and 100 nM). The adenosine A(2A) receptor agonist, 2 p-(2-carboxyethyl)phenethyl-amino-5'-N-ethylcarboxamidoadenosine, increased tritium overflow, but only in the epididymal portion. The enhancement caused by NECA was prevented by the adenosine A(2A) receptor antagonist, 4-(2-[7-amino-2-(2 furyl)[1,2,4]triazolo-[2,3-a][1,3,5]triazin-5-ylamino]ethyl)phenol (ZM 241385; 20 nM), in the epididymal and by the adenosine A(2B) receptor antagonist, alloxazine (1 microM), in the prostatic portion. Inhibition of adenosine uptake enhanced tritium overflow in both portions, an effect blocked by ZM 241385 in the epididymal and by alloxazine in the prostatic portion. The results indicate that adenosine exerts an adenosine A(1) receptor-mediated inhibition, in both portions, and facilitation mediated by adenosine A(2A) receptors in the epididymal and by A(2B) receptors in the prostatic portion. PMID- 12126971 TI - Cyclooxygenase inhibitors attenuate endothelin ET(B) receptor-mediated contraction in human temporal artery. AB - It is well documented that endothelin ET(B) receptor-mediated contraction develops in artery segments incubated in culture and that the reaction is augmented by proinflammatory cytokines, but little is known of the mechanisms involved. Segments of human temporal artery were incubated in organ culture for 2 days in the absence or presence of interleukin-1 beta (IL-1 beta), with or without nonsteroidal anti-inflammatory drugs, glucocorticoids or a nitric oxide synthase inhibitor. Thereafter, contractions were induced by the selective endothelin ET(B) receptor agonist, sarafotoxin S6c. Acetylsalicylic acid, indomethacin, nimesulide and rofecoxib were all effective in eliminating the increase in endothelin ET(B) receptor-mediated contraction induced by interleukin 1 beta, but only indomethacin and rofecoxib significantly reduced the spontaneous development of this reaction in cultured arteries. Dexamethasone and methylprednisolone augmented the reaction, and the nitric oxide synthase inhibitor had no effect. The results clearly indicate a role for cyclooxygenase, most likely cyclooxygenase-2, in endothelin ET(B) receptor-mediated contraction in this preparation. PMID- 12126972 TI - A comparative study of the effects of Cl(-) channel blockers on mesenteric vascular conductance in anaesthetized rat. AB - There is evidence to suggest that niflumic acid is capable of selectively inhibiting Ca(2+)-dependent Cl(-) channels. Furthermore, it has been demonstrated that niflumic acid is capable of antagonizing contractile responses due to activation of alpha(1)-adrenoceptor in mesenteric vasculature. Here, we have examined the effects of three Cl(-) channel blockers, niflumic acid, indanyloxyacetic acid 94 (IAA-94) and diphenylamine-2-carboxylic acid (DPC) on cirazoline-mediated vasoconstriction in mesenteric blood vessel in vivo. Infusion of cirazoline produced a dose-dependent increase in blood pressure, decrease in superior mesenteric blood flow, mesenteric vascular conductance and heart rate. While niflumic acid and IAA-94 did not have any impact on cirazoline-induced changes in blood pressure, DPC accentuated the pressor effect of cirazoline. Neither agent affected cirazoline-mediated reflex reduction in the heart rate. Niflumic acid, IAA-94 and DPC attenuated alpha(1)-adrenoceptor mediated decrease in mesenteric blood flow and vascular conductance. Based on the profile of the actions of these compounds, it may be suggested that IAA-94 did not appear to act as selective inhibitor of Ca(2+)-activated Cl(-) channels when compared to niflumic acid in the mesenteric blood vessels. In addition, while DPC seems to be as effective as niflumic acid in its effects on mesenteric blood vessels, its actions may be attributed to other pharmacological effects. PMID- 12126973 TI - Inhibitory effects of GABA(B) receptor agonists on swallowing in the dog. AB - The effects of the GABA(B) receptor agonists baclofen (1.4 and 7 micromol/kg i.v.) and CGP 44532 ([(2S)-3-amino-2-hydroxypropyl]methyl phosphinic acid], 0.2 and 0.7 micromol/kg i.v.) on transient lower esophageal sphincter relaxations and spontaneous and pharyngeally stimulated swallowing were investigated in conscious dogs. Both compounds inhibited transient lower esophageal sphincter relaxations dose-dependently, CGP 44532 being approximately fivefold more potent. In experiments designed to measure transient lower esophageal sphincter relaxations, spontaneous swallowing was suppressed by both compounds. When swallowing was evoked by intrapharyngeal water injection, both baclofen and CGP 44532 reduced the occurrence of primary peristalsis. It is concluded that centrally acting GABA(B) receptor agonists inhibit spontaneous and stimulated swallowing probably through an action in the central pattern generator for swallowing. PMID- 12126974 TI - Modeling and biological evaluation of 3,3'-(1,2-ethanediyl)bis[2-(4 methoxyphenyl)-thiazolidin-4-one], a new synthetic cyclooxygenase-2 inhibitor. AB - Within the series of chiral 3,3'-(1,2-ethanediyl)bis[2-arylthiazolidin-4-ones], the 3,4-dimethoxyphenyl substituted derivative was found in the primary anti inflammatory screening to be endowed with superior in vivo properties and good safety profile. Such a lead compound was modified by eliminating 3-methoxy group while retaining 4-methoxy group on the aryl rings at 2 and 2' stereogenic carbons. The 2R,2'S-meso isomer (VIG3b) of the resulting bisthiazolidinone has been widely investigated. The inhibitory effects on cyclo-oxygenase-1 and cyclo oxygenase-2 isoenzymes were measured in a human whole blood assay. VIG3b was almost 50 times more selective on the inducible isoform. The cyclo-oxygenase-2 preferential selectivity has been confirmed by modeling VIG3b into the cyclo oxygenase-1 and cyclo-oxygenase-2 active sites. Furthermore, VIG3b was assayed in the experimental model of carrageenan-induced lung injury by evaluating its ability to inhibit: (1) fluid accumulation in the pleural cavity, (2) neutrophil infiltration, (3) prostaglandin E(2) production and (4) lung injury. VIG3b exhibited interesting activity in all these tests. PMID- 12126975 TI - Pharmacological studies of diacerein in animal models of inflammation, arthritis and bone resorption. AB - Diacerein has proved to be effective in the treatment of osteoarthritis. We investigated the effects of diacerein in animal models of carrageenin-, zymosan-, or dextran-induced paw edema and adjuvant-induced arthritis and in ovariectomized rats. In acute inflammatory models, unlike classical nonsteroidal anti inflammatory drugs such as naproxen and ibuprofen, diacerein inhibited the rat paw edema induced by various agents. In the adjuvant-induced arthritic rats, diacerein at 100 mg/kg/day significantly suppressed the paw edema and the increase in serum mucoprotein. Addition of 3 mg/kg/day naproxen to each diacerein (3, 10, 30 mg/kg/day) dose resulted in significantly greater anti-inflammatory activity than with naproxen alone. In the ovariectomized rats, diacerein (10, 100 mg/kg/day) also significantly prevented bone loss and reduced the serum alkaline phosphatase and decreased the excretion of urinary hydroxyproline. In addition, rhein (10, 30 microM) inhibited calcium release from mouse calvaria induced by interleukin-1 beta, prostaglandin E(2) and parathyroid hormone 1-34 human fragment. These findings indicate that diacerein is a novel anti-inflammatory drug with pharmacological properties different from those of classical nonsteroidal anti-inflammatory drugs and support the clinical investigation of the use of combination therapy with diacerein and nonsteroidal anti-inflammatory drugs in patients with not only osteoarthritis but also rheumatoid arthritis. PMID- 12126976 TI - Role of leukocytes in ethanol-induced microvascular injury in the rat brain in situ: potential role in alcohol brain pathology and stroke. AB - Effects of acute and chronic alcohol ethanol administration on in vivo microvascular-leukocyte dynamics was studied in brains of naive and leukocyte depleted rats by direct, quantitative intravital high-resolution TV microscopy, fluorescence microscopy and myeloperoxidase staining. Administration of alcohol produced dose-dependent venular vasospasm, and rolling and adherence of leukocytes to venular walls; leukocyte velocity concomitantly decreased. Intermediate to high doses of ethanol resulted in infiltration of leukocytes and macrophages across venular walls, and concentration-dependent increases in myeloperoxidase staining in parenchyma, and rupture of postcapillary venules with focal hemorrhages. Use of phosphorus 31-nuclear magnetic resonance spectroscopy on intact animals revealed that the latter were associated with whole brain losses in intracellular levels of ATP and phosphocreatine with concomitant rises in intracellular inorganic phosphate and hydrogen ion concentration. Vinblastine depletion of circulating leukocytes prevented or ameliorated greatly the alcohol induced microvascular damage and proinflammatory-like reactions. These new results, when viewed in light of other recent findings, suggest that alcohol induced cerebral vascular and brain damage is dependent, to a large extent, on recruitment of leukocytes. PMID- 12126977 TI - Role of mu-opioid receptors in insulin release in the presence of inhibitory and excitatory secretagogues. AB - In mouse pancreatic islets incubated under static conditions, the inhibitory effects on glucose-evoked insulin release induced by adrenaline (1 microM), clonidine (2 microM) and UK 14,304 (brimonidine, 0.001-1 microM) were abolished by naloxone (30 nM). Only CTOP (D-Phe-Cys-Tyr-D-Trp-Orn-Thr-Phe-Thr-NH(2), 0.1 microM), a very selective mu-opioid receptor antagonist, blocked the response to UK 14,304. Glucose-induced insulin secretion was attenuated by both beta endorphin (0.01 microM) and endomorphin-1 (0.1 microM). Naloxone and CTOP prevented these inhibitory responses. The stimulatory effect of glibenclamide (1 microM) was also reduced by endomorphin-1. However, when islets were incubated in the presence of K(+) (30 mM), carbachol (100 microM) or forskolin (0.1 microM), neither the inhibitory effect induced by UK 14,304 was reversed by naloxone, nor endomorphin-1 altered the responses promoted by the excitatory agents. Thus, alpha(2)-adrenoceptor stimulation might inhibit glucose-induced insulin secretion by releasing endogenous opioids. Mu-Opioid receptor activation and opening of K(ATP) channels could be involved in the response. PMID- 12126978 TI - Disparate effects of small medial amygdala lesions on noncontact erection, copulation, and partner preference. AB - Male rats with radiofrequency lesions in the anterior medial amygdala (MeAa) or the posterior medial amygdala (MeAp), respectively, were tested for copulation and for noncontact erection (NCE; evoked by inaccessible estrous females) in a chamber in which the male was located between estrous and anestrous females. Barriers allowed only olfactory and auditory interaction between animals. With conscious females as stimuli, MeAp lesions virtually eliminated NCEs, and MeAa lesions moderately impaired them, without affecting the normal preference for estrous over anestrous females. When tested with anesthetized females to remove auditory stimulation, few males with lesions had NCEs. Only the males with MeAp lesions had a significant reduction in preference for estrous over anestrous anesthetized females. Neither MeAa nor MeAp lesions had an effect on copulatory behavior. MeAp lesions may have caused a reduced sensitivity to--or impaired processing of--estrous odors, thereby preventing NCE without disrupting copulatory behavior. PMID- 12126979 TI - Specific neuronal protein: a new tool for histological evaluation of excitotoxic lesions. AB - An important issue in the interpretation of behavioral data obtained from animals with excitotoxic lesions is evaluation of the extent of the lesions. Animals often have to be excluded from the behavioral analysis because the lesions are either not at the intended location or extend beyond it. Therefore, a clear cut histological evaluation is imperative for a meaningful interpretation of the behavioral results. Although Nissl staining is the most commonly used histological method for the evaluation of lesions, it is very difficult, if not impossible, to obtain a clear delineation of the lesioned area in Nissl-stained sections in some regions of the brain, such as the nucleus accumbens. This is especially the case when long survival times are used. In the present study, we introduce a simple and reliable immunohistochemical marker for the evaluation of excitotoxic lesions in the brain, the neuronal nuclei (NeuN) protein. With this staining, we have been able to delineate the lesions in problematic areas, such as the shell territory of the nucleus accumbens, with far greater accuracy than conventional Nissl staining. PMID- 12126980 TI - Social stress does not alter the expression of sensitization to cocaine. AB - The effects of chronic social stress on behavioral sensitization to cocaine were investigated in the Syrian hamster. Adolescent animals received either 15 mg/kg i.p. of cocaine or saline twice per day for 7 consecutive days. Two weeks following the last injection (young adulthood), they were given a challenge dose of 5 mg/kg i.p. of cocaine and scored for locomotion. Motor activity was significantly greater in cocaine-treated animals, demonstrating sensitization to this psychostimulant. Following the results of the first study, another group of adolescent animals was exposed to either a novel clean cage (control) or an aggressive resident male hamster (social stress) for 15 min following an injection of cocaine (20 mg/kg i.p. once daily) or saline for 7 consecutive days. The groups were as follows: Social Stress/Cocaine (SSC), No Social Stress/Cocaine (NSSC), Social Stress/Saline (SSS) and No Social Stress/Saline (NSSS). Two weeks following the last injection (Day 21), all animals were given a challenge dose of cocaine (5 mg/kg i.p.) and were rescored for locomotion. At that time, the suppressive effect of stress on locomotion was no longer detectable, as the expression of sensitization was observed in the NSSC but not in the SSC group. These results suggest that chronic social stress administered during adolescence does not cross-sensitize with cocaine in young adult hamsters. PMID- 12126981 TI - GABAergic control of food intake in the meat-type chickens. AB - This study examined the effects of intracerebroventricular injections of gamma aminobutyric acid (GABA) agonists on short-term food intake in meat-type cockerels. In Experiment 1, birds were injected with various doses of muscimol, a GABA(A) agonist. In Experiment 2, the birds received bicuculline, a GABA(A) antagonist, prior to injection of muscimol. In Experiment 3, the effect of varying doses of baclofen, a GABA(B) agonist, on food intake was determined. The intracerebroventricular injection of muscimol caused a dose-dependent increase in food intake. This effect was significantly attenuated by pretreatment with bicuculline. Food intake was not affected by the intracerebroventricular injection of baclofen. These results suggest that GABA acts within the brain of broilers at a GABA(A), but not GABA(B), receptor to increase voluntary food intake. PMID- 12126983 TI - The influence of heredity on self-reported sleep patterns in free-living humans. AB - The heritability of self-reported sleep patterns was investigated with 86 identical and 78 fraternal same-sex and 51 fraternal mixed-gender adult twin pairs who were paid to maintain 7-day diaries. Linear structural modeling was applied to investigate the nature and degree of genetic and environmental influences and revealed significant genetic influences on the time that individuals went to sleep and woke up, how often the individual woke up during the night, the duration of sleep and wakefulness, and how alert the individual felt upon waking and over the day, accounting for 21% to 41% of the variance. These influences of heredity were present for sleep-wake behavior over the entire week and also when the sleep-wake pattern was analyzed separately for weekdays and weekends. Further, it was demonstrated that there were multiple independent influences of heredity on sleep-wake behavior. The results suggest that sleep wake patterns are not learned but result in part from multiple heritable influences. PMID- 12126982 TI - Split circadian rhythms of female Syrian hamsters and their offspring. AB - In several mammalian species, circadian pacemakers of breeding females synchronize the developing clocks of offspring by as of yet unspecified mechanisms. The present study assessed whether maternal communication of circadian rhythms extends beyond setting pacemaker phase to include transfer of a fundamental reorganization of component circadian oscillators from dams to pups. In Experiment 1, a regimen of daily novel wheelrunning previously demonstrated to split activity rhythms of adult male hamsters into two discrete components was shown to similarly reorganize female hamster rhythms. In Experiment 2, females split by this method and unsplit controls exposed to similar light environments were mated with males. Split and unsplit females were equally fecund, but the former weaned pups of lower body weight. After weaning into running wheel cages, offspring of split dams were more likely to exhibit split activity rhythms than were offspring of unsplit females. Among pups not categorized as split, moreover, maternal entrainment nonetheless influenced distribution of pup activity across the 24-h cycle. Entrainment patterns of split and unsplit pups resembled those of adults. Thus, split and unsplit hamster dams provide different entraining signals to their developing offspring. The influence of maternal rhythms extends beyond entraining phase to alter interactions between component circadian oscillators that underlie split activity bouts. Maternal effects did not persist beyond the second week postweaning in split or unsplit hamsters, however, and rhythms of many split pups later joined. Thus, the maternal influence on the pup's circadian pacemaker may be transient. PMID- 12126984 TI - Effects of aging on food intake and body composition in rats. AB - Alterations in the ability to adjust energy intake when optional dietary foods are available may contribute to aging-related changes in body composition. Ingestive behavior, however, has not been extensively studied in aging models. The present research used young, middle-aged and old rats to examine food intake under several different schedules of optional fat availability. All rats were provided with continuous access to a nutritionally complete diet throughout the 6 week study. In addition, different subgroups within each age had access to an optional source of vegetable shortening under schedules in which the frequency of access was manipulated: controls (C)-- no shortening access; MWF--2-h shortening access on Monday, Wednesday and Friday; D2--2-h shortening access every day; D24- 24-h shortening access every day. Energy intake was significantly greater in groups with more frequent access to shortening regardless of age. The length of time the rats remained hyperphagic, however, increased with age. Body weight increased significantly in the D24 group in middle-aged and old rats, but not in young rats. Total body fat was also significantly higher in middle-aged and old groups with more frequent access to shortening, but not in young rats. Finally, body ash mass was significantly reduced in old rats on the D24 diet. These results suggest that alterations in responses to an optional source of dietary fat may contribute to aging-associated body fat accretion and body mineral loss. PMID- 12126985 TI - Isotonic NaCl intake by cell-dehydrated rats. AB - Male adult rats that received an intragastric load of 2 ml of 12% NaCl (n=13) ingested both water (4.0+/-0.2 ml/2 h) and 0.9% NaCl (3.7+/-1.0 ml/2 h) when compared with rats that received intragastric load of 2 ml of water (water: 0.1+/ 0.1; 0.9% NaCl: 0.5+/-0.3 ml/2 h, n=12) in a two-bottle test. Intragastric sodium load increased plasma sodium concentration and osmolality by 5% and reduced plasma renin activity by half compared to rats that received intragastric load of water. Intravenous infusion of 1.5 ml/10 min of 10% NaCl (n=16) also induced ingestion of water (6.2+/-0.8 ml/2 h) and 0.9% NaCl (2.9+/-0.8 ml/2 h) compared with intravenous infusion of 1.5 ml/10 min of 0.9% NaCl (water: 0.9+/-0.4; 0.9% NaCl: 0.5+/-0.2 ml/2 h, n=14). Therefore, a sodium load that raises natremia and plasma osmolality, and therefore induces cell dehydration, results in both 0.9% NaCl and water ingestion when the rats have a two-bottle choice. PMID- 12126986 TI - Efficacy of elevated body swing test in the early model of Parkinson's disease in rat. AB - Animal models of Parkinson's disease (PD) with partial damage of the dopaminergic nigrostriatal system are very suitable for the development of neuroprotective and neurotrophic treatment strategies. Although drug-induced rotational behavior has conventionally been used for the analysis of lesioned animals, a pure behavioral test that can evaluate such animals in a drug-free state may better reflect a more natural response following lesion. In this study, an early model of PD was developed by intrastriatal injection of 12.5 microg of 6-hydroxydopamine (6-OHDA) into the left striatum. Apomorphine-induced rotational and drug-free elevated body swing behaviors were evaluated. The results of the rotational test revealed a very significant contralateral turning in the rats of the lesion group (L+V) compared with the sham-operated group (SH) (P<.0001). In addition, the results of elevated body swing test (EBST) showed a significant difference between the L+V and SH groups in the second (P<.01) and fourth weeks (P<.05) after surgery. Further analysis of correlation for the net number of rotations versus the net number of swings revealed a significant and positive correlation (r=.52) in the second week in the L+V group, but no such correlation was observed in the fourth week (r=.24). Taken together, it is concluded that despite a poor correlation at fourth week postlesion, EBST itself may be appropriate and sensitive for the evaluation of motor asymmetry in the unilateral model of early PD in rats. PMID- 12126987 TI - Some failures of intragastric NaCl infusions to support flavor preference learning. AB - Rats can acquire conditioned flavor preferences based on the postingestive delivery of energy-rich nutrients. Here, we tried to condition flavor preferences to the postingestive delivery of the nutrient, sodium. Three experiments were conducted. In one, sodium-depleted rats were allowed to choose between two arbitrary flavors for 8 days. Ingestion of one of the flavors initiated infusions of one of two volumes of 150 mM NaCl, whereas ingestion of the other flavor had no effect. The rats showed no preference for the flavor paired with NaCl infusion. In another, sodium-depleted rats were given two pairs of 24-h one bottle training trials. Ingestion of the flavor presented during one trial initiated infusions of water or one of four concentrations of NaCl (75, 150, 300 or 450 mM NaCl). Ingestion of the flavor presented on the other trial in each pair had no effect. In a subsequent two-bottle choice between the two flavors, the rats were either indifferent to (water, 75 and 150 mM) or avoided (300 or 450 mM) the infusion-paired flavor. In the third study, replete rats were trained as in the previous experiment with 150 mM NaCl as the infusate and tested after sodium deficiency was induced by furosemide. These animals avoided the flavor paired with NaCl infusion. We conclude that rats cannot associate an arbitrary taste with the beneficial effects of postingestive delivery of NaCl, or if they can, this occurs only under rare circumstances. PMID- 12126988 TI - Intragastric calcium infusions support flavor preference learning by calcium deprived rats. AB - We investigated whether rats can associate the flavor of ingested solutions with the postingestive delivery of calcium. In one series of experiments using the "electronic esophagus" preparation, calcium-deprived rats received pairs of daily one-bottle training trials in which they received intragastric infusions whenever they ingested an arbitrary flavor of Kool Aid. Rats later preferred the flavor associated with infusions of 50 mM CaCl(2) or 50 mM calcium lactate (CaLa), but not with water, 10, 100 or 250 mM CaCl(2) or 100 mM sodium lactate (NaLa). In another experiment, rats had simultaneous access to two arbitrary flavors, ingestion of one of which produced intragastric infusion of 50 mM CaCl(2), 75 mM NaCl or water. Only the rats given 50 mM CaCl(2) developed a preference for the flavor associated with the infusion. The preference for calcium infusions was not as large as that seen for orally ingested calcium. Nevertheless, these results show that 50 mM calcium infusions are rewarding to calcium-deprived rats. They thus suggest that rats can associate flavor ingestion with the postingestive benefits of consuming calcium. PMID- 12126989 TI - Blind mice are not impaired in T-maze footshock avoidance acquisition and retention. AB - The processing of visual information during learning and memory is considered to be a vital function of the hippocampus. Some researchers believe that the sole purpose of the hippocampus is to process visuo-spatial information, whereas other investigators believe that the hippocampus integrates cues from multiple sources. In the current studies, we tested the effects of vision loss on a hippocampal task, acquisition and retention with T-maze footshock avoidance conditioning. Acquisition and retention, in adult-blinded mice, were not significantly impaired in T-maze footshock avoidance. Blindness did not affect activity, footshock startle or motivation to avoid shock. The same doses of memory enhancing drugs that improve memory in sighted mice improved memory in blind mice. Electrolytic lesions in blind mice, which destroyed 31+/-4% of the hippocampus, significantly impaired acquisition and retention for T-maze footshock avoidance and so demonstrated that the hippocampus retained its integrative role in blind mice. The current findings show that blind mice are as capable of learning T-maze footshock avoidance as sighted mice and that the hippocampus retains its important role in blind mice in learning and memory processing. It is concluded that the T-maze footshock avoidance conditioning task is a spatially but not visually dependent task that is hippocampally dependent. PMID- 12126990 TI - Evidence that transient nicotine lowers the body weight set point. AB - OBJECTIVE: Smokers usually gain weight when they quit smoking. The present work explores the hypothesis according to which such a rise is a behavioral response to a raised body weight set point taking place when nicotine is eliminated from the body. RESEARCH METHODS AND PROCEDURES: The human body weight set point was assessed with classical behavioral and psychophysical methods, from the delay to experience negative alliesthesia when repeatedly ingesting sweet stimuli. Seven habitual smokers were tested once before lunch, after smoking (nonabstinent) as usual and once again after refraining from smoking (abstinent). Three additional nicotine-naive subjects were tested under the same procedure after receiving at 0730 h in the morning a transdermal nicotine patch (14 mg) or a placebo patch. Two of the subjects also received nicotine (7 mg) for a third session. RESULTS: Oral and transdermal administration of nicotine did not decrease the initial pleasure or modify the initial palatability of eating sweet stimuli, but significantly accelerated the following onset of self-reported displeasure (negative alliesthesia) aroused by repeated ingestion of sweet stimuli. DISCUSSION: These results are understood as an acute lowering of the body weight set point by nicotine. The body weight gain taking place after quitting smoking may, therefore, be explained by the removal of the lowering of the body weight set point induced by nicotine. PMID- 12126991 TI - Rats fed only during the light period are resistant to stress-induced weight loss. AB - Repeated restraint stress (3 h/day for 3 days) causes a chronic down-regulation of body weight in rats. This study determined whether weight loss was influenced by the time of day that rats had access to food or that stress was applied. Groups of male Sprague-Dawley rats were fed a 40% kcal fat diet with food given ad libitum, only during the light phase or only during the dark phase. After 2 weeks of adaptation, rats within each feeding treatment were divided into four groups. One was exposed to repeated restraint at the start of the light phase, another was restrained at the start of the dark phase and the remaining groups were nonstressed controls for restrained rats. Body weight was significantly reduced in ad libitum- and dark-fed restrained rats, compared with nonstressed controls, from Day 2 of restraint, regardless of the time of day that they were stressed. There was no significant effect of restraint on weight change of light fed rats. Food intake was inhibited by stress in ad libitum- and dark-fed rats, but it was not changed in light-fed rats. Serum corticosterone was increased by restraint in all rats irrespective of feeding schedule. This study demonstrates that stress-induced weight loss only occurs when rats have food available during their normal feeding period (dark phase) and is not determined by increased corticosterone release. PMID- 12126992 TI - The effects of manipulated arousal on children's willingness to taste novel foods. AB - We examined the effects of manipulated arousal on willingness to taste moderately novel and extremely novel foods in children ranging from 7 to 12 years of age. Children were assigned at random to one of three arousal conditions (low, moderate, and high). Twice during the 30-min manipulation period, the children rated their willingness to taste the foods, with the understanding that these ratings would be used to determine which foods they would taste later in the session. Results of an Age x Gender x Arousal condition analysis on willingness to try the novel foods revealed a significant effect of arousal condition; willingness increased with decreasing arousal. Separate analyses for the moderately and extremely novel foods yielded significant condition and age effects for the former and no significant effects for the latter. The results were discussed in the context of optimal level of arousal theories. PMID- 12126993 TI - Social factors regulate female-female aggression and affiliation in prairie voles. AB - Although patterns of aggression and affiliation may play a major role in social organization, the mechanisms underlying these behaviors are not well understood. The purpose of this study was to examine the effects of social and hormonal experience on female-female aggression and affiliation in prairie voles, Microtus ochrogaster. Prairie voles exhibit the traits of social monogamy and tend to live in communal families structured around a male-female pair. It is rare for two unrelated females within a family to successfully reproduce. In this study, the social and/or hormonal experiences of female prairie voles were varied and female female aggression and affiliation were measured during dyadic encounters with unfamiliar, nonaggressive females. An increase in aggression and decline in affiliative behaviors toward a stimulus female was observed during pregnancy and following male-cohabitation, with or without mating. Pairing with another female did not result in changes in either aggressive or affiliative behaviors toward the stimulus female. Female-female aggression increased and affiliative behaviors declined, with a maximal effect following approximately 8-12 days of male cohabitation. Similar patterns of change were seen in both ovariectomized and gonadally intact females, and treatment with estradiol and subsequent sexual experience did not significantly alter the tendency of females to show aggression or affiliative contact. Social experiences associated with prolonged cohabitation with a male facilitate the induction of female-female aggression; however, ovarian hormones, pregnancy or mating are not essential to induce aggression. PMID- 12126994 TI - The effects of exercise intensity on thermoregulatory responses to exercise in women. AB - We investigated the influence of altering exercise intensity (150, 300, and 450 kpm/min) on the resetting of the core temperature threshold for the onset of the sweating rate (M(sw)) and the alteration of sweating sensitivity during the menstrual cycle in women. Five women underwent cycling exercise for 30 min in both the luteal and follicular phases under controlled neutral environmental conditions (T: 25 degrees C, RH: 55%). A significantly higher rectal temperature (T(re)) was seen in the luteal phase at all exercise intensities, and the same time course of the T(re) response with a constant difference of approximately 0.2 degrees C was shown between the follicular phase and the luteal phase. The T(re) threshold for M(sw) was also apparently shifted rightward a constant value of 0.2 degrees C from the follicular phase to the luteal phase, independent of the alteration of exercise intensity. The slope of the M(sw)-T(re) relationship in the follicular phase did not differ from that in the luteal phase. These results indicate that (1) a rightward shift in the T(re) threshold from the follicular phase to the luteal phase can be observed independent of any alteration of the exercise intensity; and (2) the sensitivity of M(sw) is also not physiologically influenced by exercise intensity. Thus, alterative thermoregulation during the menstrual cycle was fundamentally unaffected by the change of exercise intensity. PMID- 12126995 TI - Intracerebroventricular administration of octopamine stimulates food intake of chicks through alpha(2)-adrenoceptor. AB - Octopamine, known to be an important neurotransmitter in invertebrates, has been noted to have several similarities to noradrenaline (NA) in mammals. The present study was done to elucidate whether central injection of octopamine enhances the feeding behavior of chicks and to investigate the interaction of octopamine with both alpha(1)- and alpha(2)-adrenoceptors. We found that the intracerebroventricular injection of octopamine significantly stimulated food intake of neonatal chicks during 30 min postinjection, but not thereafter. Moreover, this octopamine-induced eating response was attenuated by the alpha(2) antagonist yohimbine, but not by the alpha(1)-antagonist prazosin. These results suggest that the action of octopamine on the feeding behavior of the neonatal chick is similar to that of NA, since octopamine regulates food intake through the alpha(2)-adrenoceptor. PMID- 12126996 TI - Brain opioid receptor density reflects behavioral and heart rate responses in pigs. AB - Results from our previous research indicate that long-term tether-housed pigs with high and low levels of stereotypies show differences in the density of endogenous opioid receptors in the hippocampus and hypothalamus. It was not clear whether differences in opioid receptor density were induced by the chronic stress of tether housing or stereotypy performance, or were already present before the animals were tethered. The latter possibility was tested in the present experiment. We used a group of 18 nonstereotyping pigs that had no experience with tether housing and investigated whether the animals differed in the density of endogenous opioid receptors in the brain and, if so, whether these differences were related to the animals' reactions to acute challenges. The pigs were subjected to two tests: an open field test and a tethering test. Behavioral reactions as well as heart rate responses were measured. Opioid receptor densities were determined postmortem in the hippocampus and hypothalamus using a membrane binding assay with [(3)H]naloxone as a ligand. Animals differed widely in their responses to the two tests. In support of our hypothesis, we found a relationship between behavioral and heart rate responses and densities of naloxone binding sites in the hippocampus and hypothalamus. The data suggest that endogenous opioid systems in the brain contribute to differences in stress responding between individual pigs. PMID- 12126997 TI - Effects of exposure to extremely low-frequency magnetic field of 2 G intensity on memory and corticosterone level in rats. AB - In the present study, we examined the effects of chronic exposure (1 and 2 weeks) to an extremely low-frequency magnetic field (ELFMF) of 2 G intensity on memory in rats using an object recognition task. Comparable groups of rats were exposed for 1, 2 or 4 weeks to ELFMF and the following day blood samples were collected from each rat for the measurement of corticosterone level. Our results demonstrate that exposure to ELFMF induces a significant increase in the level of corticosterone in blood plasma and is associated with impairment in discrimination between familiar and novel objects. PMID- 12126999 TI - Cocaine-induced locomotor activity is enhanced by exogenous testosterone. AB - Anabolic-androgenic steroids are synthetic derivatives of testosterone, which are increasingly abused by adolescent populations who also abuse psychoactive substances. All these compounds lead to complex behavioral syndromes and the effects of their interactions remain unclear. The main aim of the present study was to determine the influence of testosterone on the locomotor activity promoting effect of cocaine on male mice in an open field. In the three experiments, animals received two injections: firstly, testosterone or peanut oil, and secondly, cocaine or saline solution. In Experiments 1 and 2, testosterone (or oil) and cocaine (or saline) were injected 45 and 10 min, respectively, prior to activity recording. In the first experiment, we studied the effects of testosterone (2 mg/kg) on locomotor activity induced by different doses of cocaine (2, 4, 8, 10 or 12 mg/kg). In Experiment 2, we explored the effects of supraphysiological doses of testosterone (2, 6, 10 or 14 mg/kg) on animals treated with 10 mg/kg cocaine. Finally, in the third experiment, 14 mg/kg testosterone or vehicle was administered 15, 30, 45 or 75 min before activity data collection to animals that received 10 mg/kg cocaine or saline. Testosterone itself had no effects on spontaneous locomotor activity and, as was expected, cocaine increased locomotor activity dose-dependently. Given together, testosterone enhanced the cocaine-induced hyperactivity although not dose dependently, the highest effects being found 45 min after testosterone injection. The present study confirmed the existence of an interaction between testosterone and cocaine at the central nervous system. PMID- 12126998 TI - Age-associated difference in circadian sleep-wake and rest-activity rhythms. AB - Using actigraphic monitoring of wrist activity, we investigated the sleep and rest-activity patterns of 65 young, middle-aged, old and the oldest subjects in their natural environmental conditions. To assess the effects of age and gender on sleep and circadian rhythms in activity, multivariate analyses were performed. Age significantly affected circadian sleep and rest-activity rhythms. In the old and oldest groups, the actigraphic estimates of "actual sleep time" and "sleep efficiency" decreased significantly. The estimates of "sleep latency," the number of "nighttime awakening," sleep fragmentation and daytime naps significantly increased in the old and oldest groups. Concerning the circadian patterning of rest and activity, the interdaily stability (IS) was similar in the four age groups, while the old and oldest subjects showed significant increases in intradaily variability (IV) and nighttime activity and a decrease in amplitude (AMP). The present study demonstrated weakened and fragmented circadian sleep and rest-activity rhythms during aging. However, no gender-related difference was found. PMID- 12127000 TI - Behavioral effects of 7-nitroindazole on hyperbaric oxygen toxicity. AB - The aim of this study was to evaluate the role of nitric oxide (NO) upon hyperbaric oxygen (HBO) toxicity in male Sprague-Dawley rats during exposure to 0.5 MPa >99% O(2). In the first experiment, the selective neuronal NO synthase inhibitor 7-nitroindazole (7-NI) was injected intraperitoneally (ip) in 15 rats. Another 15 rats received vehicle injections of peanut oil intraperitoneally. Latency to observable tonic-clonic convulsions and motor activity during the HBO exposure were scored and compared between the control group and the 7-NI group. The results showed that injection of 7-NI (30 mg/kg) significantly prolonged the latency to observable tonic-clonic convulsions. The 7-NI group also showed a significant decrease in motor activity compared with the control group. A second experiment was performed to measure the effect of 7-NI injections upon open-field activity during normobaric conditions. Twenty-four male Sprague-Dawley rats were randomly divided into three groups, each consisting of eight rats receiving 30 mg/kg 7-NI injections, 10 mg/kg 7-NI injections or vehicle injections of peanut oil intraperitoneally, respectively. The results showed that injection of 7-NI led to a significant dose-dependent reduction in horizontal and vertical activities. This study shows that 7-NI prolongs the latency to hyperoxia-induced seizures. However, it also demonstrates that 7-NI in doses ranging from 30 to 10 mg/kg has a secondary effect upon motor behavior in general. It can therefore not be ruled out that the protective effect of 7-NI upon HBO intoxication is partly due to reduced motor activity. PMID- 12127001 TI - Exploration and motor activity in juvenile and adult rats exposed to hypergravity at 1.8 G during development: a preliminary report. AB - Pups from gestating rats exposed to hypergravity (1.8 G) or to normal gravity at the perinatal period were evaluated for motor activity, exploration and social interactions during juvenile and adult stages. By comparison to controls, the hypergravity group had shorter latencies before choosing a maze arm in a T-maze and a lower number of exploratory pokes in a hole board. During dyadic encounters, the hypergravity group had a lower number of self-grooming episodes and shorter latencies before crossing under the opposing rat. In contrast, no intergroup differences were observed during exploration of an elevated plus-maze and a light-dark box. These results indicate that exposure to 1.8 G during development appears to decrease exploratory tendencies in the hole board and fear related responses in T-maze and social interaction tests. PMID- 12127002 TI - Effect of the volume of liquid food infused intragastrically on satiety in women. AB - This experiment examined whether food volume and energy content affected satiety in lean and obese women, when visual and oral cues were bypassed by infusing food intragastrically. The effects of volume and energy content were examined separately by using liquid foods that differed in energy density (kcal/g). On 5 separate days, 25 lean and 29 obese women consumed all of their meals in the laboratory. A nasogastric tube was inserted 30 min before lunch on 4 days; on three of these occasions, a liquid preload was infused for 15 min and, on the fourth occasion (control), the preload was diverted covertly. On the remaining day, no tube was inserted. The three preloads varied two-fold in volume and energy content: 200 ml/200 kcal, 400 ml/200 kcal and 400 ml/400 kcal. The results showed that increasing the volume of infused food, but not the energy content, affected satiety in both lean and obese women. There was a mean decrease in energy intake at lunch of 77 kcal (13%) after the 400-ml preload compared with the iso-energetic 200-ml preload (P=.013). Increasing the energy content of infused food, but not the volume, did not affect satiety. Thus, when sensory cues were bypassed, the volume of liquid food infused intragastrically affected subsequent energy intake in both lean and obese women. These results suggest that gastric and postgastric mechanisms are involved in the effects of high-volume, low-energy-dense foods on satiety. PMID- 12127003 TI - Flavor preferences conditioned by sucrose depend upon training and testing methods: two-bottle tests revisited. AB - In confirmation of prior work, rats given one-bottle training with flavored 5% and 30% sucrose solutions (CS5 and CS30) strongly preferred the CS5 when both flavors were presented in intermediate 17.5% sucrose solutions. The CS5 preference has been attributed to a conditioned satiety response to the CS30 flavor, but equal intakes of CS5 and CS30 in one-bottle tests did not support this view. To determine if sweetness differences between training and test solutions contributed to the CS5 preference, new rats were trained and tested with flavored 10% sucrose solutions. One flavor (CS5) was paired with matched intragastric (ig) water infusions (=net 5% solution) and another flavor (CS30) was paired with matched infusions of 50% sucrose (=net 30% solution) during one bottle training. In two-bottle tests with both flavors paired with an intermediate infusion (25%=net 17.5%), the rats initially showed no overall preference for the CS5 or CS30. Following additional training, the rats significantly preferred the CS30 to the CS5. The intragastric data suggested that a change in sweet taste context between training and testing might have accounted for the strong CS5 preference obtained in the first experiment. This was confirmed in a third experiment in which rats were trained with flavored 5% and 30% sucrose solutions and then given two-bottle tests with both flavors presented either in 5% sucrose or 30% sucrose. Rats tested with 30% sucrose strongly preferred the CS5 flavor, whereas rats tested with 5% sucrose significantly preferred the CS30 flavor. Thus, the outcome of two-bottle flavor preference tests and presumably other tests of conditioned flavor reward may be greatly influenced by the solutions used in the tests. The impact of this variable may be greatest when the training solutions do not substantially differ in their net postingestive reinforcing actions. This appears to be the case with 5% and 30% sucrose solutions because the satiating effect of the concentrated solution tends to counteract its nutrient reinforcing action. PMID- 12127005 TI - Anterograde and retrograde influence of vestibular stimulation on spatial working memory. AB - Rats were trained in an eight-arm radial maze to explore the apparatus in search of a food reward. After completion of the training phase, some animals were submitted to a hemicerebellectomy (HCbed group), while others were used as a control group. To study the effects of vestibular stimulation on the recall of ongoing working memory information, both groups were exposed to radial maze sessions: in the first session (no-rotation), animals were confined for 30 s to the fourth arm visited without being further manipulated; in the second session (rotation), the animals were again confined for 30 s to the fourth arm visited, while the apparatus was rotated five times around its vertical axis. The effects of these manipulations on successive visits to complete the task were assessed, as well as the solving time and kinds of errors made. Errors were significantly more frequent in the control animals during the rotation session; HCbed animals were unaffected by confinement alone or by vestibular stimulation, but showed a decreased search speed. It was concluded that vestibular input is required for an adequate functioning of the working memory system devoted to the formation and consolidation of spatial mnesic traces and that the amnesic effect due to vestibular stimulation is both anterograde and retrograde in nature. PMID- 12127004 TI - Effects of perinatal octylphenol on ultrasound vocalization, behavior and reproductive physiology in rats. AB - A rodent diet containing paraffin wax was designed to administer the environmental estrogen octylphenol (OP) to nonpregnant, pregnant and lactating rats. The estrogenic activity of OP via this diet was first confirmed in ovariectomized adult animals: 20 mg OP/kg/day increased the mitoses in the vaginal epithelium, and 60 mg OP/kg/day stimulated mitoses in the uterine luminal epithelium. The effects on a variety of reproductive and nonreproductive parameters were then investigated in the offspring of dams fed OP (100-250 mg/kg/day during gestation and lactation). A number of modest reproductive and morphological effects observed in the offspring including decreased body weights in adults of both sexes, disrupted vaginal cyclicity and decreases in seminiferous tubule diameter and testis, kidney, spleen and ovary weights. Behavioral effects included increased sexual arousal in males, increased sexual motivation in females towards a female teaser and increased motor activity by females. Ultrasonic vocalizations by pups at Postnatal Day (PND) 7 were reduced in number and duration in both sexes. There were no effects of perinatal OP on ano-genital distance, prepuce separation, aggressive behavior or adult ultrasound vocalization. These observations confirm that the dietary intake of estrogenic amounts of OP during pregnancy and lactation can have a wide variety of effects in the offspring. PMID- 12127006 TI - Differential responses to odorant analogs after recovery from nerve transection. AB - We previously found that exposure-induced increase in odor sensitivity involves, at least in part, the olfactory epithelium. We did this by exposing mice to 5 alpha-androst-16-en-3-one (androstenone) and measuring changes in the epithelium. Past research showed that sensitivity to androstenone also could be induced by exposing individuals to 4-(4',4'-dimethylcyclohexyl)-2-methylcyclohexanone (DMCMC), a structural and functional analog of androstenone. What remained unknown is whether structural and/or functional odorant analogs share peripheral components. In the current work, we used a well-established model to disconnect the olfactory epithelium from the olfactory bulbs (BNX) to disrupt mechanisms underlying olfactory coding (when the afferents reinnervate the bulb, they do not synapse in their original glomeruli), and to examine the effects of disruption and restoration on exposure-induced odor sensitivity. In this study, we determined whether analogs of androstenone, viz., 5 alpha-androstan-3-one (androstanone) and DMCMC, could induce sensitivity to androstenone after BNX. Results demonstrate that exposure to either androstanone or DMCMC can induce sensitivity to androstenone in control mice. Different results were observed in mice that had recovered from bilateral BNX. Exposure to androstanone for 10 days immediately after surgery increased sensitivity to androstenone; however, exposure to DMCMC did not. These results suggest that androstanone and DMCMC, although apparent perceptual analogs of androstenone, may be using different pathways of olfaction within the central nervous system (CNS). PMID- 12127007 TI - Effects of incisor-cutting on muricidal behavior induced by olfactory bulbectomy in rats. AB - Muricidal behavior in rats is composed of two main components, attacking and killing performance. Since a large number of mice could be killed by rats during behavioral experiments, research has been limited in the past decade possibly because of ethical considerations. In preliminary studies, we found that the rat incisors play a key role in muricidal behavior in rats, so, in the present study, we cut off the incisors and assessed the following parameters of muricidal behavior: attack latency, first attack site, lethal attack site, attack frequency, total attack duration and mean attack duration. If after incisor cutting (IC) rats still tried to demonstrate muricidal activity, but failed to kill the mouse, this would be an ideal model for studying the mechanisms of muricidal behavior. Since muricide can be induced in rats by olfactory bulbectomy (OBX), young adult male Wistar rats with OBX displaying muricidal behavior were tested for muricidal activity 4 h after IC, then every 24 h for 3 days. At 4 and 28 h after IC, only 9% and 36% of rats killed mice, but these values rose to 73% and 82% 52 and 76 h after IC, respectively. At 4, 28 and 52 h after IC, there was no significant difference in attack latency, first attack site, lethal attack site or mean attack duration between IC-treated rats (both killers and nonkillers) and sham-operated controls, while the attack frequency was obviously increased in IC nonkiller rats, and a significantly longer total attack duration was seen in both IC killer and nonkiller rats compared to controls. Since IC treatment increases attack frequency and prolongs the total attack duration without affecting other basic components of muricidal behavior in rats, these results suggest that the killer rats treated with IC may provide a suitable model for research on muricidal behavior. PMID- 12127008 TI - Isolation rearing-induced facilitation of Pavlovian learning: abolition by postsession intra-amygdala nafadotride. AB - It has been shown previously in this laboratory that rats reared in social isolation acquire a Pavlovian-conditioned approach task much more rapidly than their respective controls. This study assessed the involvement specifically of the mesoamygdaloid dopamine pathway in this facilitated learning of isolates. Thus, animals were required to associate arbitrary stimuli with a pulsed light stimulus (unconditioned stimulus, US). The US, while without biological significance, was nevertheless capable of eliciting an intrinsic and sustained alerting response. Procedures ensured that the arbitrary stimuli (tone or clicker) did not elicit a response in the first instance, and were presented either paired (CS+) or unpaired (CS-) with the US. Isolates and socially reared controls received intra-amygdala infusions of the D3 dopamine receptor antagonist, L-nafadotride, or vehicle immediately following the end of each training session. The conditioned response increased over sessions in both groups of vehicle-infused rats during presentations of the CS+ stimulus, but not CS-, and isolates acquired this association more rapidly than controls. However, acquisition of this association was abolished by postsession intra-amygdala L nafadotride. Responding to the US was largely unaffected by drug or rearing conditions. Hence, these data provide strong evidence for the specific involvement of the mesoamygdaloid dopamine projection in the facilitation of associative learning by isolation rearing. PMID- 12127009 TI - Duration of sexual arousal predicts semen parameters for masturbatory ejaculates. AB - A man's semen parameters may vary considerably from one specimen to the next, partly due to variability in the conditions under which the specimens are produced. In the present study, the relationship between the duration of preejaculatory sexual arousal and the quality of semen produced by masturbation was investigated. Twenty-five regular semen donors aged 22-44 provided a total of 292 semen specimens (median 11 per donor) over a period of 4 months. Each specimen was produced after a minimum of 3 days of ejaculatory abstinence and measures included the time taken to produce the specimen, ejaculate volume, sperm concentration, and percent motility. Linear regression revealed that, controlling for donor identity, there was a significant (t=2.13, P<.05) positive relationship between the time taken to produce a specimen and sperm concentration. We conclude that the duration of preejaculatory sexual arousal is an important predictor of ejaculate quality for specimens produced by masturbation and that variation in the duration of preejaculatory arousal may contribute to within-male fluctuations in semen parameters over time. PMID- 12127010 TI - Maternal alcohol consumption during pregnancy and fetal startle behaviour. AB - The startle behaviour of the fetus (both spontaneous and elicited) was examined in fetuses of mothers who drank alcohol and mothers who did not. Fetuses exposed to alcohol showed a higher frequency of spontaneous startles and were less likely to exhibit a normal startle in response to a vibroacoustic stimulus. These differences illustrate a teratogenic effect of alcohol on CNS functioning in utero, possibly associated with brainstem damage. PMID- 12127011 TI - Oxytocin induces long-term depression on the rat dentate gyrus: possible ATPase and ectoprotein kinase mediation. AB - We studied the effects of the neuropeptide oxytocin (OT) on the long-term potentiation (LTP) paradigm in the dentate gyrus (DG) of urethane anesthetized rats. Intracerebroventricular injection of 1 microg of the hormone in 1 microl of physiological solution 2min before tetanization produced a significant decrease in both components of the perforant path evoked potentials (EP) in the DG. The effects appeared right after the tetanization stimuli and were more pronounced in the excitatory postsynaptic components of the EPs. The decrements lasted for the 2h of recording time. We concluded that OT induced and maintained long-term depression on the DG. In contrast, injection of OT in the absence of tetanic stimulation did not significantly affect perforant path EP in the DG. The results are discussed taking particular consideration of the inhibitory effects the OT has on (Ca(2+)+Mg(2+)) ATPase at membrane levels and the potential interference that this action may have with phosphorylation processes via an ectoprotein kinase isolated from membranes of hippocampal pyramidal neurons. Blocking of this ectoprotein kinase in vitro significantly impairs establishment and maintenance of LTP. PMID- 12127013 TI - Neuropathic pain is associated with alterations of nitric oxide synthase immunoreactivity and catalytic activity in dorsal root ganglia and spinal dorsal horn. AB - Previous experiments have suggested that nitric oxide may play an important role in nociceptive transmission in the spinal cord. To assess the possible roles of neuronal nitric oxide synthase (nNOS) in spinal sensitization after nerve injury, we examined the distribution of nNOS immunoreactivity in dorsal root ganglia (DRGs) and dorsal horn of the corresponding spinal segments. NOS catalytic activity was also determined by monitoring the conversion of [3H]arginine to [3H]citrulline in the lumbar (L4-L6) spinal cord segments and DRGs in rats 21 days after unilateral loose ligation of the sciatic nerve. Behavioral signs of tactile and cold allodynia developed in the nerve-ligated rats within 1 week after surgery and lasted up to 21 days. Immunocytochemical staining revealed a significant increase (approximately 6.7-fold) of nNOS-immunoreactive neurons and fibers in the DRGs L4-L6. No significant changes were detected in the number of nNOS-positive neurons in laminae I-II of the spinal segments L4-L6 ipsilateral to nerve ligation. However, an increased number of large stellate or elongated somata in deep laminae III-V of the L5 segment expressed high nNOS immunoreactivity. The alterations of NOS catalytic activity in the spinal segments L4-L6 and corresponding DRGs closely correlated with nNOS distribution detected by immunocytochemistry. No such changes were detected in the contralateral DRGs or spinal cord of sham-operated rats. The results indicate that marked alterations of nNOS in the DRG cells and in the spinal cord may contribute to spinal sensory processing as well as to the development of neuronal plasticity phenomena in the dorsal horn. PMID- 12127012 TI - Withdrawal from dependence upon butorphanol uniquely increases kappa(1)-opioid receptor binding in the rat brain. AB - Changes in kappa(1)-opioid receptor binding have been implicated in the development of dependence upon and withdrawal from butorphanol. Autoradiographic characterization of binding for brain kappa(1)-([3H]CI-977), mu-([3H]DAMGO), and delta-([3H]DPDPE) opioid receptors was performed in rats undergoing naloxone precipitated withdrawal from dependence upon butorphanol or morphine. Dependence was induced by a 72h i.c.v. infusion with either butorphanol or morphine (26nmol/microl/h). Withdrawal was subsequently precipitated by i.c.v. challenge with naloxone (48 nmol/5 microl/rat), administered 2h following cessation of butorphanol or morphine infusion. During withdrawal from butorphanol, but not morphine, kappa(1)-opioid receptor binding was increased significantly in the frontal cortex, posterior basolateral amygdaloid nucleus, dorsomedial hypothalamus, hippocampus, posterior paraventricular thalamic nucleus, ventral tegmental area and locus coeruleus. In contrast, mu-opioid receptor binding decreased in these brain regions in naloxone-precipitated withdrawal from morphine, but not butorphanol, while binding for delta-opioid receptors was altered in both withdrawal groups. The brain kappa(1)-opioid receptor appears to be more directly involved in the development of physical dependence upon, and the expression of withdrawal from, butorphanol, as opposed to the prototypical opioid analgesic, morphine. PMID- 12127014 TI - The regional distribution of nitric oxide synthase activity in the spinal cord of the dog. AB - The aim of this study was to examine the distribution of calcium-dependent nitric oxide synthase activity (cNOS) in the white and gray matter in cervical, thoracic, lumbar and sacral segments of the spinal cord and cauda equina of the dog. The enzyme's activity, measured by the conversion of [3H]arginine to [3H]citrulline revealed considerable region-dependent differences along the rostrocaudal axis of the spinal cord in general and in cervical (C1, C2, C4, C6 and C8) and lumbar (L1-L3, L4-L7) segments in particular. In the non compartmentalized spinal cord, the cNOS activity was lowest in the thoracic and highest in the sacral segments. No significant differences were noted in the gray matter regions (dorsal horn, intermediate zone and ventral horn) and the white matter columns (dorsal, lateral and ventral) in the upper cervical segments (C1 C4), except for a significant increase in the ventral horn of C4 segment. In C6 segment, the enzyme's activity displayed significant differences in the intermediate zone, ventral and lateral columns. Surprisingly, extremely high cNOS activity was noted in the dorsal horn and dorsal column of the lowest cervical segment. Comparing the enzyme's activity in upper and lower lumbar segments of the spinal cord, cNOS activity prevailed in L4-L7 segments in the dorsal horn and in all the above mentioned white matter columns. PMID- 12127015 TI - Effects of 192 IgG-saporin on acetylcholinesterase histochemistry in male and female rats. AB - Sex hormones may exert neuroprotective effects in various models of brain lesions. Male and female Long-Evans rats were subjected to intracerebroventricular injections of 2 microg 192 IgG-saporin or vehicle. Starting 2 days before surgery, half the male rats were treated with estradiol for 7 days. Three weeks after surgery, they were sacrificed for histochemical staining of acetylcholinesterase (AChE) and densitometric evaluations. The lesion induced a substantial to dramatic decrease of the AChE-positive fiber density in the cingulate, somatosensory, piriform, retrosplenial and perirhinal cortices, and in the hippocampus. Weak effects were found in the striatum. There was no significant decrease in the dorsal thalamus. Sex had no significant effect on AChE-positive staining in any brain area. In males, estradiol treatment did not alter the effects of 192 IgG-saporin. These results show that sex or estradiol treatment in male rats does not interfere with the immunotoxic effects of intracerebroventricular injections of 192 IgG-saporin on cholinergic projections from the basal forebrain. PMID- 12127016 TI - Acute effects of amperozide and paroxetine on social cohesion in male conspecifics. AB - The effects of acute administration of the selective 5-hydroxytrypamine (5-HT; serotonin) uptake inhibitor, paroxetine, and the potent 5-HT(2A) receptor antagonist, N-ethyl-4-[4',4'-bis(rho-flourophenyl)butyl]-1-piperazinecarboxamide (amperozide) on social cohesion was determined by using a tether paradigm, in which the movement of one of a pair of rats was restricted to one-half of an observation chamber. Administration of 0.1mg/kg paroxetine, 1.0, 3.0, and 5.0mg/kg amperozide but neither 1.0 nor 10.0mg/kg paroxetine increased time spent in contact with the untreated, tethered rat. Whereas only the lowest dose of paroxetine (0.1mg/kg) promoted social cohesion, all doses of amperozide (1.0, 3.0, and 5.0mg/kg) promoted social cohesion. The amperozide-induced increases in time spent in contact were greater than the paroxetine-induced increases in seconds spent in contact, regardless of dose. Acute administration of amperozide is more effective than acute administration of paroxetine in promoting social cohesion between pairs of male conspecifics. Amperozide may be an effective alternative treatment for patients with social anxiety disorder suffering from adverse side effects of paroxetine. Amperozide may prove to be a more effective treatment for social anxiety disorder than paroxetine, which is supported by our results. PMID- 12127017 TI - A new easy accessible and low-cost method for screening olfactory sensitivity in mice: behavioural and nociceptive response in male and female CD-1 mice upon exposure to millipede aversive odour. AB - In a previous study, mice were found to be repelled by the odour emitted by the millipede (Ommatoiulus sabulosus) as a defensive strategy against predators [Physiol. Behav. 74 (2001) 305-311]. To develop a standardised test for screening olfactory capabilities in rodents, we have characterised the behavioural response displayed by adult male and female CD-1 mice when exposed to a Stimulus Object (SO) consisting of a millipede-shaped sponge previously soaked either in a Toluquinone (TQ) solution (5g/100ml; Fluka), a chemical component of the exudate secreted by the millipede, or in distilled water. In Experiment 1, behaviours performed when exposed to the SO were scored (15min for 5 consecutive days). TQ exposure suppressed nearly completely Catching and Eating the SO, and increased general activity in a sex-dependent fashion. In Experiment 2, performances in a hot-plate test (50+/-0.5 degrees C, cut-off 60s) were assessed immediately after a 15-min exposure to the SO. Toluquinone-exposed mice showed a subtle yet significant decrease of pain threshold. TQ exposure assay is a new, easily testable, and low-cost method for measuring rodents olfactory sensitivity relevant for the analysis of the pharmacological agents, lesions and transgenesis. PMID- 12127018 TI - Beta-amyloid-induced increase in the resting intracellular calcium concentration gives support to tell Alzheimer lymphocytes from control ones. AB - Senile plaques containing beta-amyloid peptide (betaAP) comprise the major neuropathological lesions in Alzheimer's disease (AD). In line with ongoing studies investigating alterations of various biochemical processes of cells of peripheral tissues, the authors demonstrate differences in resting intracellular free calcium levels of lymphocytes harvested from sporadic Alzheimer patients and from age-matched controls. Resting intracellular calcium concentration was measured in Fura-2AM-loaded human lymphocytes by dual wavelength spectrofluorimetry. Resting calcium level appeared to be higher in Alzheimer cells when compared to control lymphocytes. After incubating cells in 10(-7)M of beta-amyloid, the resting calcium concentration of the control cells elevated, while that of Alzheimer lymphocytes did not differ considerably. PMID- 12127019 TI - Effects of androstenedione on long term potentiation in the rat dentate gyrus. Relevance for affective and degenerative diseases. AB - We studied the effects of the androgenic hormone androstenedione, a 17 ketosteroid, on long term potentiation (LTP) in the dentate gyrus (DG) of intact, urethane anesthetized rats. Intravenous injection of 10mg of the hormone dissolved in Nutralipid produced a significant increase of the population spike (PS), but not of the excitatory post-synaptic potentials (EPSPs). The results are discussed in terms of the potential enhancement that androstenedione may have on some aspects of memory processes as reported for other androgenic steroids. Also noted are the plausible beneficial effects of the hormone on depression as well as in recovery following both central and peripheral neural injury. PMID- 12127020 TI - Comparisons between C57BL/6J and A/J mice in motor activity and coordination, hole-poking, and spatial learning. AB - The C57BL/6J (B6) inbred mouse strain was compared to the A/J inbred strain for motor activity in an open-field, exploration of a hole-board, motor coordination in the coat-hanger test, and spatial learning in the Morris water maze. B6 mice displayed a higher number of segment crossings in the open-field and of hole poking responses than A/J mice. The performance of B6 mice was superior to that of A/J mice not only in the submerged but also in the visible platform version of the Morris water maze. By contrast to their hypoactivity, the A/J strain had shorter movement times in the coat-hanger test, indicating faster motor speed, although the groups did not differ in latencies before falling. These results indicate that recombinant inbred or congenic strains derived from B6 and A/J mice offer considerable potential for discerning the genetic basis of several behavioral phenotypes. PMID- 12127021 TI - Midbrain projecting dorsal column nucleus neurons in a reptile. AB - The origin of midbrain projecting cells in the dorsal column nucleus (DCN) was investigated in reptiles, Caiman crocodilus, using a retrograde tracer. Labeled neurons were confined to a caudal-central portion of the DCN. Midbrain projecting DCN neurons had round, oval, or triangular soma and were small. While neurons that project to the spinal cord in Caiman are also located in the caudal half of the DCN, midbrain projecting cells are located more dorsally and are smaller than those whose axons terminate in the spinal cord. Taken together, these observations suggest that the DCN in Caiman is subdivided, at least in part, according to target location. In view of similar findings in certain birds and mammals, subdivisions of the DCN into sectors is likely a phylogenetically ancient feature of amniote sensory systems transmitting somatosensory information from the body surface. PMID- 12127022 TI - Effect of long-term aluminum feeding on kinetics attributes of tissue cholinesterases. AB - Aluminum (Al) is considered a potential etiological factor in Alzheimer's disease (AD). Neurotoxicity from excess brain exposure to Al is documented from both clinical observations and animal experiments. A key role of the acetylcholine system in memory disturbances that characterize AD has been reported. On this basis, we studied the effect of long-term Al feeding on kinetic properties of cholinesterases employing the rat as experimental model. Animals were given prolonged treatment with soluble salts of Al (100mg AlCl(3)/kg body weight mixed with food for 100-115 days), and the kinetic properties of cholinesterases (acetylcholinesterase, AChE, and butyrylcholinesterase, BChE) were determined in different tissues. Prolonged treatment with Al had no effect on the K(m) values of the soluble and membrane-bound forms of AChE in the brain, but V(max) was instead decreased in all the components of soluble and membrane-bound forms of AChE in the brain. In addition, the Al treatment resulted in complete loss of the component II of erythrocyte membrane AChE. Surprisingly, after prolonged treatment with Al, higher V(max) was observed in all the components of soluble and membrane-bound forms of BChE in the heart and liver. Variable effects of Al exposure were observed on temperature kinetic properties of cholinesterases. Altogether these findings indicate that long-term Al feeding results in inhibition of AChE, while an opposite effect is observed on BChE. Decreased V(max) of the brain AChE could represent the mode of action through which Al may contribute to pathological processes in Al-induced neurotoxicity. PMID- 12127023 TI - Disruption of astroglial interlaminar processes in Alzheimer's disease. AB - A palisade of long, interlaminar astroglial processes in supragranular layers of the cerebral cortex is characteristic of adult individuals of anthropoid species. In the present study, this distinctive cytoarchitectonic feature was analyzed in tissue deriving from the neocortex of cases affected by Alzheimer's disease (n=14) and age-matched control cases (n=10). Samples of different cortical areas, and in particular prefrontal, temporal and striate fields, were analyzed. Astroglia was labeled by glial fibrillary acidic protein immunoreactivity, that allowed a clear distinction between the classical, stellate intralaminar astroglia and the interlaminar glial processes. The occurrence and relative density of neuritic plaques were ascertained in the same specimens with Bielchowsky staining. In most cortical regions of cases diagnosed as severe Alzheimer's disease by the donor institutions, interlaminar astroglia was found to be markedly altered or absent, and replaced by hypertrophic intralaminar astrocytes. Cases diagnosed as milder or uncertain Alzheimer's disease showed a less consistent involvement of the interlaminar glial palisade. Alterations of the interlaminar palisade in the cortex affected by Alzheimer's disease did not strictly correlate with the density of neuritic plaques in the examined specimens. The findings indicate that loss/severe disruption of the interlaminar palisade of astroglial processes is part of the array of neuropathological changes occurring in the cerebral cortex during Alzheimer's disease. In addition, our data indicate that different types of neocortical astrocytes (namely intralaminar and interlaminar astrocytes) respond differently to the pathobiology of Alzheimer's disease in the neocortex, inasmuch as interlaminar processes tend to disappear while intralaminar processes become reactive. PMID- 12127024 TI - Pain elicited by blunt pressure: neurobiological basis and clinical relevance. PMID- 12127025 TI - Unidimensional pain rating scales: a multidimensional affect and pain survey (MAPS) analysis of what they really measure. AB - Pain is now regarded as 'the fifth vital sign' and patients are frequently asked to score the intensity of their pain on a numerical pain rating scale (NPRS). However, the use of a unidimensional scale is questionable in view of the belief, overwhelmingly supported by clinical experience as well as by empirical evidence from multidimensional scaling and other sources, that pain has at least two dimensions: somatosensory qualities and affect. We used a Chinese translation of the 101 descriptor multidimensional affect and pain survey (MAPS) questionnaire to determine the relative contributions of various dimensions of postoperative pain to a patient's score on a unidimensional NPRS. MAPS and NPRS were administered postoperatively to 69 patients with descending colon carcinoma who were recovering from left hemi-colectomy. Multiple linear regression revealed that the emotional pain qualities supercluster (P=0.0005) and four of its eight subclusters, anxiety, depressed mood, fear and anger, significantly (P=0.001 0.007) predicted a patient's score on the unidimensional NPRS. Notably, none of the 17 subclusters in the somatosensory pain qualities supercluster predicted NPRS scores. It may be concluded that patient scores on unidimensional pain intensity scales reflect the emotional qualities of pain much more than its sensory intensity or other qualities. Accordingly such scales are poor indicators of analgesic requirement. The results also suggest that patients' postoperative anxiety and depression are inadequately treated. Based on our findings we present six unidimensional scales that should yield a more accurate assessment of the sources of a patient's pain. PMID- 12127026 TI - Nerve injury induces plasticity that results in spinal inhibitory effects of galanin. AB - Galanin is a 29-amino-acid neuropeptide that has been implicated in the processes of nociception. This study examines the effect of exogenous galanin on dorsal horn neurone activity in vivo in the spinal nerve ligation (SNL) model of neuropathic pain. SNL rats but not naive or sham-operated rats exhibited behaviour indicative of allodynia. In anaesthetized rats, extracellular recordings were made from individual convergent dorsal horn neurones following stimulation of peripheral receptive fields electrically or with natural (innocuous mechanical, noxious mechanical and noxious thermal) stimuli. Spinal administration of galanin (0.5-50 microg) caused a slight facilitation of the neuronal responses to natural and electrical stimuli in naive rats and up to a 65% inhibition of neuronal responses in sham-operated rats following 50 microg galanin. In contrast, there was a marked inhibition of up to 80% of responses to both natural and electrical stimuli in SNL rats following spinal galanin administration. These results suggest that following peripheral nerve injury, there is plasticity in the levels of galanin and/or its receptors at spinal cord level so that the effect of exogenous galanin favours inhibitory function. PMID- 12127027 TI - Increased synaptosomal [3H] GABA uptake in the rat brainstem after facial carrageenan injections. AB - The aim of the present study was to quantify synaptosomal [(3)H] gamma aminobutyric acid (GABA) uptake in the rat brainstem after facial carrageenan injections. Synaptosomal preparations from the brainstem of rats that had received one or four facial carrageenan injections showed greater GABA binding on the side of the brainstem ipsilateral to the carrageenan injection than on the contralateral side when compared to saline injected controls. In contrast, no difference in GABA binding between the injected and contralateral sides was observed in the same synaptosomal preparations that had been treated with GABA uptake inhibitors NNC-711, beta-alanine, or nipecotic acid. The difference between GABA binding in the absence of the GABA uptake inhibitor and GABA binding in a portion from the same synaptosomal preparation which had been incubated with the GABA uptake inhibitor was obtained to represent [(3)H] GABA binding to GABA transporters/transporter mediated [(3)H] GABA uptake. A significantly greater GABA uptake was observed on the side of the brainstem ipsilateral to the carrageenan injection(s) than on the contralateral side. A consequence of the observed increase in GABA uptake is that it could reduce the amount of GABA in the synaptic cleft. This could influence the transmission of nociceptive input from primary afferents to secondary neurons in the spinal trigeminal nucleus and could be a contributing factor in the development of hyperalgesia after carrageenan injections or other chronic inflammatory conditions. PMID- 12127028 TI - No pain, no gain: clinical excellence and scientific rigour--lessons learned from IA morphine. AB - The effectiveness of intra-articular (IA) morphine in arthroscopic procedures of the knee joint was analysed in all randomised and controlled trials that included injections of morphine and placebo into the knee joint, and where the data were analysable. Sensitivity of the studies and effectiveness were analysed for three different periods: immediate (0-2h), early (2-6h) and late (6-30h). Sensitivity for each period was assumed if pain intensity was at least 30% of the maximum of 100 on the visual analogue scale in the placebo group. Six different doses (1 10mg) of IA morphine were compared with placebo. The injections were made at the end of surgery, before the arthroscope was removed from the joint. In the immediate period 7/15 sensitive trials were positive, in the early period 8/12 sensitive trials were positive and in the late period 10/13 sensitive trials were positive. Most positive studies had used higher doses (3-5mg) compared with negative studies that had mainly used 1mg. Two studies using patient controlled analgesia consumption of analgesics as an outcome were also positive. The only sensitive study of four dose-response comparisons indicated that 5mg of IA morphine was more effective than 1mg. The only sensitive study of three cross route comparisons showed no difference between 5mg of IA and 5mg of intra muscular morphine. All insensitive trials, including placebo (except two individual comparisons), cross-route and dose-response comparisons, were negative. The analysis of sensitive studies indicates that 5mg of IA morphine injected into the knee joint provides postoperative pain relief for up to 24h. A minimum of 30% of the maximum possible pain intensity is needed for an analgesic effect to be detected in a study. PMID- 12127029 TI - Sensitization of trigeminal caudalis neuronal responses to intraoral acid and salt stimuli and desensitization by nicotine. AB - In human studies, repeated intraoral application of strong acidic or salt stimuli induces irritation that progressively increases across trials (sensitization), whereas irritation elicited by nicotine progressively decreases (desensitization). We investigated whether nociceptive neurons in trigeminal subnucleus caudalis (Vc) exhibit increasing or decreasing patterns of firing to the intraoral application of these irritants. In rats anesthetized with halothane and thiopental, single-unit recordings were made from nociceptive neurons in superficial layers of dorsomedial Vc that responded to mechanical and noxious thermal and chemical stimulation of the tongue. NaCl (5M), citric acid (300 mM), pentanoic acid (300 mM) or nicotine (600 mM) were separately delivered to the tongue by constant flow (0.32 ml/min) for 15 or 25 min. NaCl, citric acid and pentanoic acid each elicited a progressive, significant increase in Vc neuronal firing over the initial 10 min to a plateau level that was maintained for the stimulus duration. Nicotine induced a significant increase in firing rate of Vc neurons within 6 min, followed by a decline back to the baseline level over the ensuing 10 min. Following a rest period, reapplication of nicotine no longer activated Vc neurons, indicative of self-desensitization. We additionally tested for nicotine cross-desensitization to acid. After recording the responses of Vc neurons to pentanoic acid and noxious heat, nicotine was then applied for 15 min. Post-nicotine responses to pentanoic acid were markedly reduced (to 13% of control), indicative of cross-desensitization; responses to noxious heat were also reduced to a lesser degree (to 71% of control). The progressive increase in Vc neuronal firing elicited by NaCl and acid, and the decline in firing after initial nicotinic excitation, resemble psychophysical patterns of sensitization and desensitization, respectively, and support the involvement of Vc neurons in the signaling of oral irritant sensations. PMID- 12127030 TI - Expression of 5-HT1A receptor mRNA in rat lumbar spinal dorsal horn neurons after peripheral inflammation. AB - The present study observed the expression of the 5-hydroxytryptamine (5-HT) (1A) receptor mRNA in the lumbar spinal dorsal horn neurons following carrageenan inflammation using in situ hybridization (ISH). We also studied the co localization of 5-HT(1A) receptor mRNA and gamma-amino butyric acid (GABA) or enkephalin (ENK) immunoreactivities using a combined fluorescent ISH and immunofluorescent histochemical double-staining technique. The finding of this study demonstrated that 5-HT(1A) receptor mRNA was widely distributed in the spinal dorsal horn with the highest density in laminae III-VI. Following carrageenan-induced inflammation, the 5-HT(1A) receptor mRNA expression in all layers of ipsilateral dorsal horn was significantly enhanced, and the peak occurred after 8h. Furthermore, the number of 5-HT(1A) receptor mRNA and GABA or ENK immunoreactive double-labeled cells was also markedly increased 8h after carrageenan injection. These findings suggested that following peripheral inflammation, the synthesis of 5-HT(1A) receptor was increased in the lumbar spinal dorsal horn neurons, especially in spinal GABA and ENK neurons. PMID- 12127031 TI - Local cooling does not prevent hyperalgesia following burn injury in humans. AB - One of the oldest methods of pain relief following a burn injury is local application of ice or cold water. Experimental data indicate that cooling may also reduce the severity of tissue injury and promote wound healing, but there are no controlled studies in humans evaluating the anti-inflammatory or anti hyperalgesic potential of early cooling after thermal injury. Twenty-four healthy volunteers participated in this randomized, single-blinded study. Following baseline measurements, which included inflammatory variables (skin temperature, erythema index) and sensory variables (thermal and mechanical detection thresholds, thermal and mechanical pain responses, area of secondary hyperalgesia), first degree burn injuries were induced on both calves by contact thermodes (12.5 cm(2), 47 degrees C for 7 min). Eight minutes after the burn injury, contact thermodes (12.5 cm(2)) were again applied on the burns. One of the thermodes cooled the burn (8 degrees C for 30 min) whereas the other thermode was a non-active dummy on the control burn. Inflammatory and sensory variables were followed for 160 min after end of the cooling procedure. The burn injury induced significant increases in skin temperature (P<0.0005), erythema index (P<0.0001), thermal pain responses (P<0.0005), mechanical pain responses (P<0.005) and secondary hyperalgesia, and significant decreases in heat pain threshold (P<0.0005) and mechanical pain threshold (P<0.0005). There were no post cooling effects on skin temperature (P>0.5), erythema (P>0.9), heat pain threshold (P>0.5), thermal or mechanical pain responses (P>0.5) or the development of secondary hyperalgesia (P>0.4) compared with the control burn. However, a significant, albeit transient, increase in cold detection threshold was observed on the cooled burn side (P<0.0001). In conclusion, cooling with 8 degrees C for 30 min following a first degree burn injury in humans does not attenuate inflammatory or hyperalgesic responses compared with a placebo-treated control burn. PMID- 12127033 TI - Altered central sensorimotor processing in patients with complex regional pain syndrome. AB - Alterations in tactile sensitivity are common in patients with chronic pain. Recent brain imaging studies have indicated that brain areas activated by acute experimental pain partly overlap with areas processing innocuous tactile stimuli. However, the possible effect of chronic pain on central tactile processing has remained unclear. We have examined, both clinically and with whole-head magnetoencephalography, six patients suffering from complex regional pain syndrome (CRPS) of the upper limb. The cortical somatosensory responses were elicited by tactile stimuli applied to the fingertips and the reactivity of spontaneous brain oscillations was monitored as well. Tactile stimulation of the index finger elicited an initial activation at 65 ms in the contralateral SI cortex, followed by activation of the ipsi- and contralateral SII cortices at about 130 ms. The SI responses were 25-55% stronger to stimulation of the painful than the healthy side. The distance between SI representations of thumb and little finger was significantly shorter in the hemisphere contralateral than ipsilateral to the painful upper limb. In addition, reactivity of the 20-Hz motor cortex rhythm to tactile stimuli was altered in the CRPS patients, suggesting modified inhibition of the motor cortex. These results imply that chronic pain may alter central tactile and motor processing. PMID- 12127032 TI - Efficacy of continuous versus intermittent morphine administration after major surgery in 0-3-year-old infants; a double-blind randomized controlled trial. AB - A randomized double-blind clinical trial compared the efficacy of 10 microg/kg/h morphine continuous intravenous infusion (CM) with that of 30 microg/kg morphine (IM) every 3h after major abdominal or thoracic surgery, in 181 infants aged 0-3 years. Efficacy was assessed by the caregiving nurses with the COMFORT 'behavior' and a visual analogue scale (VAS) for pain, every 3h in the first 24h after surgery. Random regression modeling was used to simultaneously estimate the effect of randomized group assignment, actual morphine dose (protocol dosage plus extra morphine when required), age category, surgical stress, and the time varying covariate mechanical ventilation on COMFORT 'behavior' and the observational VAS rated pain, respectively. Overall, no statistical differences were found between CM and IM morphine administration in reducing postoperative pain. A significant interaction effect of condition with age category showed that the CM assignment was favorable for the oldest age category (1-3 years old). The greatest differences in pain response and actual morphine dose were between neonates and infants aged 1-6 months, with lower pain response in neonates who were on average satisfied with the protocol dosage of 10 microg/kg/h. Surgical stress and mechanical ventilation were not related to postoperative pain or morphine doses, leaving the inter-individual differences in pain response and morphine requirement largely unexplained. PMID- 12127034 TI - Disappearance of central thalamic pain syndrome after contralateral parietal lobe lesion: implications for therapeutic brain stimulation. AB - At present there is hardly any appropriate therapy for central pain syndromes available. We report on a unique case of a central thalamic pain syndrome that did not respond to any therapy but disappeared after an additional contralateral parietal lobe lesion. This example indicates that lesions affecting the bilateral balance of thalamo-parietal circuits may lead to pain relief in patients with central pain syndrome, which probably constitutes a bilateral disorder of functional plasticity. This should be taken into account in chronic brain stimulation for persistent pain states. PMID- 12127035 TI - Dissociation between cutaneous and deep sensibility in central post-stroke pain (CPSP). AB - We present three cases of central post-stroke pain after right hemorrhagic or ischemic stroke associated with severe impairment of cutaneous sensibility but preservation of stimulus-evoked pain from periosteum. This is the first such report of dissociation of cutaneous- from deep-tissue sensibility loss. PMID- 12127036 TI - Continuous intraventricular clonidine infusion in controlled morphine withdrawal- case report. AB - A patient with atypical bilateral facial pain reported the loss of analgesic effect of intracerebroventricular morphine delivered continuously via an implanted pump, accompanied by intolerable adverse side effects associated with the administered high dose of morphine. Clonidine was substituted for morphine over a period of 3 weeks to achieve a drug holiday. The patient did not have significant withdrawal symptoms or major discomfort from pain, leading to a reduced quality of life during this period. Six months after the treatment, the patient continues to require a significantly lower daily dose of morphine. Morphine withdrawal with clonidine substitution produced a significant improvement in the analgesic efficacy of morphine and in the quality of life in the absence of undesirable side effects. PMID- 12127037 TI - Life and science of Patrick Wall. Letter to the editor. PMID- 12127038 TI - Genetics of osteoporosis: role of steroid hormone receptor gene polymorphisms. AB - Osteoporosis is a common skeletal disease characterized by low bone mass and microarchitectural deterioration of bone tissue with a consequent increase in bone fragility and susceptibility to fracture. In the past years, twin and family study have shown that this disease recognizes a strong genetic component and that genetic factors play an important role in regulating bone mineral density (BMD). While in few isolate conditions osteoporosis can be inherited in a simple Mendelian pattern, due to single gene mutations, in the majority of cases has to be considered a multifactorial polygenic disease in which genetic determinants are modulated by hormonal, environmental and nutritional factors. Given the important role that steroid hormones play in bone cell development and in the maintenance of normal bone architecture, polymorphisms at receptor of the steroid/thyroid hormone receptor superfamily, such as estrogen receptor alpha (ERalpha) and Vitamin D receptor (VDR) have been thoroughly investigated in the last years and appeared to represent important candidate genes. The individual contribution of these genetic polymorphisms to the pathogenesis of osteoporosis remains to be universally confirmed and an important aim in future work will be to define their functional molecular consequences and how these polymorphisms interact with each other and with the environment to cause the osteoporotic phenotype. A further promising application of genetic studies in osteoporosis comes from their pharmacogenomic implications, with the possibility to give a better guidance for therapeutic agents commonly used to treat this invalidating disorder or to identify target molecules for new therapeutic agents. PMID- 12127039 TI - Structural specificity of steroids in stimulating DNA synthesis and protooncogene expression in primary rat hepatocyte cultures. AB - Among the chemical compounds of varied structure which possess liver tumour promoting are steroids, such as estrogens, pregnenolone derivatives and anabolic steroids. Although the mechanism(s) of tumour promotion in liver by these xenobiotics is not well understood, it is clear that growth stimulation is one important element in their action. As a basis for better defining whether steroids stimulate growth by a common mechanism or fall into sub-groups with differing actions, the effects of 46 steroids on DNA synthesis and the expression of protooncogenes c-fos and c-myc were examined in primary cultures of normal rat hepatocytes. Tentative groupings of steroids have been identified based on apparent structural requirements for stimulation of DNA synthesis, and effects of auxiliary factors in modulating this growth stimulus. For a "progestin" group, insulin appeared to be permissive for stimulation of DNA synthesis, and presence of an ester or hydroxyl group at 17alpha-position in combination with a non-polar group at C(6) appeared to be required for stimulation. For the pregnenes, dexamethasone was stimulatory. Structural requirements include a non-polar substitution at 16alpha-position and presence of a 6alpha-methyl group. Androgens were weak or ineffective stimulators of DNA synthesis. Anabolic steroids were weak to strong stimulators and alteration to A ring structure in combination with non-polar substitution at 17alpha-position appeared to be required for the activity. With the exception of the anabolic steroid, dianabol, there do not appear to be strong correlation between ability to stimulate DNA synthesis and ability to induce protooncogene expression among the steroids. This study provides a starting point for future more detailed examination of growth stimulatory mechanism(s) of action of steroids in the liver. PMID- 12127040 TI - Benzo(a)pyrene exposure induces CYP1A1 activity and expression in human endometrial cells. AB - Estrogen is a major risk factor for endometrial cancer and it has been well established that smokers have a significantly reduced risk of endometrial cancer. Localized levels of estrogen within the uterus may determine the estrogenic response. The objective of this research was to investigate effects of cigarette smoke related hydrocarbons (benzo(a)pyrene, BP) on uterine CYP1A1/2 and 1B1, enzymes involved in estrogen metabolism. Human endometrium epithelial cells (RL95 2) were incubated with various concentrations (0.05, 0.1, 0.5, 1, and 10mM) of BP for 48h. CYP1 catalytic activity, protein and mRNA levels were determined. Selective chemical and immuno-inhibitors were used to determine the contribution of individual CYP1 isoenzymes. Cells expressing CYP1A1, CYP1A2 and CYP1B1 were used for comparisons. CYP1A1/2 protein and mRNA levels were significantly elevated by BP. Low level of constitutive CYP1 activity was observed in RL95-2 cells, which was significantly induced by BP exposure (12-fold at 1mM). CYP1 activity in BP-induced cells was significantly inhibited by specific anti-CYP1A1 and high concentration of alpha-naphthoflavone (ANF, 100nM), but not by selective CYP1A2 (furafylline) and CYP1B1 (homoeriodictoyl) inhibitors and low concentration of ANF (5nM). These studies suggest that CYP1A2 and CYP1B1 are not induced by BP in the endometrial cells. It also appears that CYP1A1 is one of the major CYP450 enzymes induced by BP. PMID- 12127041 TI - Estrogen receptor binding assay of chemicals with a surface plasmon resonance biosensor. AB - We have developed a simple assay method for the evaluation of estrogen receptor (ER) binding capacity of chemicals without the use of radio- or fluorescence labeled compounds. We used the solution competition assay by the BIACORE biosensor, a surface plasmon resonance biosensor, with estradiol as a ligand, human recombinant ER(alpha) (hrER(alpha)) as a high molecular weight (hmw) interactant and test chemicals as analytes. For the ligand, aminated estradiol with a spacer molecule (E2-17PeNH) was synthesized and immobilized on a carboxymethyl dextran-coated sensor chip by the amine coupling method. The injection of the hmw interactant hrER(alpha) to the biosensor raised the sensorgram, indicating its binding to the ligand E2-17PeNH. The binding of test chemicals to hrERalpha was determined as a reduction in the hrER(alpha) binding to E2-17PeNH. The dissociation constant for the binding to hrER(alpha) was calculated for estrone (4.29 x 10(-9)M), estradiol (4.04 x 10(-10)M), estriol (8.35 x 10(-10)M), tamoxifen (2.16 x 10(-8)M), diethylstilbestrol (1.46 x 10( 10)M), bisphenol A (1.35 x 10(-6)M) and 4-nonylphenol (7.49 x 10(-6)M), by plotting the data according to an equation based on mass action law. This method can also be used as a high throughput screening method. PMID- 12127042 TI - Local uptake and synthesis of oestrone in normal and malignant postmenopausal breast tissues. AB - Uptake and local formation of oestrone (E1) were studied in vivo by a double isotopic technique in normal and malignant breast tissues from 24 postmenopausal women with breast cancer. Active uptake of radio-labelled E1 beyond plasma was found both in normal and malignant tissue, the effect being significantly greater in non-malignant compared with cancer tissue. The presence of local E1 formation was also demonstrated in most samples. Both uptake and synthesis positively correlated with total amount of radioactive E1 found in the tissues. Uptake appeared to make a greater contribution to E1 levels within the breast than in situ synthesis, although there were marked variations between specimens from different patients and the relative proportion of synthesis to uptake was higher in tumour compared with non-malignant tissue. These results demonstrate quantitative differences in the different compartments by which postmenopausal breasts obtain oestrogen and highlight variations between individual breasts. This may be important in optimising oestrogen deprivation therapy for postmenopausal patients with hormone-dependent cancers. PMID- 12127043 TI - Steroid sulfatase activity and expression in mammary myoepithelial cells. AB - PURPOSE: This investigation examined mRNA expression and enzymatic activity of steroid sulfatase (STS) in human mammary myoepithelial cells (MMECs) and MCF-7 cells and assessed the effects of 17-beta estradiol on the activity of STS. METHODS: The mRNA level of STS in MMECs was determined by RT-PCR analysis using specific primers for STS. STS enzymatic activity prior to and after treatment with 17-beta estradiol was determined by measuring 3H-metabolites formed after exposure to [3H]estrone 3-sulfate (E1S) and [3H]dehydroepiandrosterone-sulfate (DHEA-S). RESULTS: Our data demonstrate the presence of STS in the MMECs. Based on RT-PCR analysis, MMECs had slightly lower levels of STS compared to MCF-7 cells. However, sulfatase activity was about 120 times greater in the MMECs than the MCF-7 cells (E1S V(max)=2640nmol/(mg DNAh) compared to 20.9nmol/(mg DNAh)). Exposure to 17-beta estradiol was associated with 70% reduction in E1S sulfatase activity in the MCF-7 cells and 9% increase in the MMECs after 6 days. DISCUSSION: Our studies indicate for the first time the presence of STS in MMECs. This is suggestive of a previously undetermined role for MMECs in converting precursor hormones into active steroid hormones within mammary tissue. In addition, differential response of the MMECs and the MCF-7 cells to estrogen demonstrates differences in hormone metabolism between these two cell types, perhaps related to the absence of estrogen receptors in the MMECs and their presence in the MCF-7 cells. The MMECs may have an important role in hormonal regulation within mammary tissue. PMID- 12127044 TI - Activation of nucleolar DNA expression in hepatocytes by glucocorticoids and high density lipoproteins. AB - A novel mechanism of protein biosynthesis regulation in liver under the action of reduced forms of steroid hormones (tetrahydrocortisol) and apolipoprotein A-I (apoA-I) is presented. Kupffer cells play an important role in uptake of the cortisol and high density lipoproteins (HDL) as well as in formation of the active complex, tetrahydrocortisol+apolipoprotein A-I (THC-apoA-I). If macrophages are stimulated by lipopolysaccharides (LPS), these processes enhance dramatically, thus causing parallel activation of nucleolar DNA expression and ribosome formation in hepatocytes. THC-apoA-I complex accelerates protein biosynthesis in primary cultures of hepatocytes, but not in macrophages and endotheliocytes. PMID- 12127046 TI - Specific radioimmunoassay of estrone sulfate. Application to measurement in male plasma. AB - We described a new, specific and easy to use radioimmunoassay (RIA) of estrone sulfate (E1S) in males. After synthesis of an E1S-6-(O-carboxymethyl) oxime hapten then coupling to BSA, we obtained a specific anti-E1S antiserum. Although the cross-reactivity of DHEAS with our anti-E1S antiserum was low (CR=0.002%), we confirmed the absolute necessity of separating plasma DHEAS from plasma E1S, before E1S RIA, because in plasma, DHEAS is present at levels 3-6000-fold higher than E1S, which generally is ignored. Thus, we elicited an easy separation of DHEAS from E1S, by a fast chromatography on in-house minicolumns. This new RIA, was applied to the determination of E1S plasma normal values in males. In 27 young men (<35 years), mean+/-S.D. were 1.97nmol/l+/-1.07nmol/l and in 63 untreated healthy aged men (>55 years), 1.80nmol/l+/-1.21nmol/l. No significant difference was seen between young and older subjects. The ranges of E1S plasma levels in these subjects were rather large and the ratios between the highest and the lowest E1S plasma levels were seven in the young group and 23.4 in the older group. No decrease of E1S plasma levels was observed with ages. Contrary to large interindividual E1S plasma level variations, the intraindividual variations have been found to be no significant. Correlations between E1S and unconjugated estrogens, E2 and E1 were 0.22 (P=0.016) and 0.51 (0.001), respectively. PMID- 12127045 TI - Nuclear exclusion of the androgen receptor by melatonin. AB - Androgen receptors (AR) play a crucial role in androgen-mediated processes and prostate cancer progression. The pineal hormone melatonin attenuates the androgen dependent growth of benign and cancer prostate epithelial cells in vitro and may reverse clinical resistance to androgen ablation therapy in patients progressing on gonadotropin releasing hormone (GnRH) analogue. Where along the AR cascade does melatonin act remains to be determined. The effects of melatonin on AR localization, level and activity were assessed using androgen-insensitive prostate carcinoma PC3 cells stably transfected with a wild-type AR-expressing vector (PC3-AR).AR was localized to the PC3-AR cell nucleus in the absence of dihydrotestosterone (DHT). Melatonin caused a robust exclusion of the AR from the cell nucleus to the cytoplasm. The nuclear export inhibitor, leptomycin B prevented this process. The exclusion was selective since melatonin had no such effect on the nuclear localization of estrogen receptors alpha (ERalpha) in these cells. Melatonin also caused nuclear exclusion of the AR in the presence of DHT. In addition, it attenuated androgen induced reporter gene activity in PC3 cells co-transfected with the human AR and AR reporter plasmids. Elevated androgen concentrations counteracted melatonin's effects. Melatonin did not decrease AR level or androgen binding in the cells. The nuclear localization of the AR is a hallmark of its cellular activity. These data point to AR nuclear exclusion as a possible mechanism to attenuate androgen responses in target tissues. PMID- 12127047 TI - The IGF-system in healthy pre- and postmenopausal women: relations to demographic variables and sex-steroids. AB - Plasma insulin-like growth factor (IGF)-I, free IGF-I and -II, IGF-binding protein (IGFBP)-1, -2, and -3 together with IGFBP-3 protease activity were measured in 114 postmenopausal and 39 premenopausal healthy women. For each parameter, the mathematical distribution was characterised, and the normal range for pre- and postmenopausal women described, together with correlations to demographic variables and sex-steroids (postmenopausal women). Postmenopausal women had lower levels of plasma IGF-I (P<0.001) and free IGF-I (P<0.001) compared to premenopausal women, while plasma IGFBP-2 (P<0.05) and immunoreactive IGFBP-3 (P<0.001) were higher in postmenopausal women. Free IGF-I (but none of the other parameters) was significantly lower in postmenopausal smokers compared to non-smokers (P<0.05).IGF-I and -II both correlated positively to height (r=0.203, P<0.05 and r=0.198, P<0.05, respectively), while IGF-II correlated positively to weight (r=0.250, P<0.01). Plasma IGF-I correlated positively to androstenedione (r=0.292, P<0.01) and dehydroepiandrosterone sulphate (DHEAS, r=0.202, P<0.05), while a significant positive correlation was observed between IGF-II on the one side and oestradiol (E(2), r=0.227), oestrone sulphate (E(1)S, r=0.238) and androstenedione (r=0.213) on the other side (P<0.05 for all). Our results support a relation between sex-steroids and IGF-I and -II in healthy postmenopausal women. The lower levels of total and free IGF-I in postmenopausal compared to premenopausal women indicate lower bioavailability of this growth factor in elderly females. PMID- 12127049 TI - Differences between CEM and human peripheral blood T lymphocytes in cAMP dependent HIV viral fusion and CXCR4 expression. AB - CXCR4, a G-protein-coupled chemokine receptor and HIV coreceptor, has been shown to play a central role in both chemotaxis and HIV-1-entry into T lymphocytes. Recent efforts have focused on identifying the signaling pathways that modulate CXCR4 expression in order to modulate HIV infectivity. Toward this effort, we previously demonstrated cAMP-dependent up-regulation of CXCR4 mRNA and protein in human peripheral blood T cells (PBL), resulting in increased HIV infectivity. Regulation of CXCR4 mRNA was mediated, in part, by a CRE element within the CXCR4 promoter. In order to develop a model system to examine cAMP regulation, the responses of the T lymphoblastoid cell line CEM were compared to those of human PBL. In sharp contrast to that of human PBL, HIV-1 entry into CEM cells was dramatically reduced in response to dibutyryl cAMP (DcAMP). Furthermore, while total cellular and cell surface CXCR4 protein levels were up-regulated in human PBL and in Jurkat T cells in response to DcAMP or forskolin stimulation, CXCR4 levels were unchanged by stimulation in CEM cells. Surprisingly, the CXCR4 promoter (nucleotides -1098 to +59) fused to luciferase was found to be activated similarly in CEM and Jurkat cells in response to DcAMP in a concentration dependent manner. RT-PCR analyses confirmed that CXCR4 mRNA levels were increased by cAMP agonists. Taken together, our findings suggest that total and cell surface CXCR4 protein expression is regulated differently in human PBL than in CEM cells, a finding that correlates with the differential HIV-1 fusion in response to cAMP signaling. Moreover, our results suggest that, for CXCR4 expression and HIV viral infectivity, CEM cells may not be a faithful model of primary human lymphocytes. PMID- 12127048 TI - Eotaxin/CCL11 is a negative regulator of neutrophil recruitment in a murine model of endotoxemia. AB - Eotaxin/CCL11 is a chemokine that has been primarily characterized with respect to its eosinophil chemoattractant activity. However, the broad tissue expression of eotaxin/CCL11 suggests that it may have other unknown activities. We have used a murine model of endotoxemia to study the role of eotaxin/CCL11 in neutrophil recruitment. We demonstrate that eotaxin/CCL11 is acutely upregulated in the serum, peritoneal wash, and lungs of mice given an intraperitoneal lipopolysaccharide (LPS) challenge. Furthermore, immunoneutralization of eotaxin/CCL11 in this model results in a significant increase in the number of neutrophils within the lung after LPS challenge. When eotaxin/CCL11 knockout mice were challenged with LPS, these mice had increased peritoneal neutrophils, but not lung neutrophils, compared to the wild-type controls. Administration of eotaxin/CCL11 to eotaxin(-/-) mice suppressed endotoxemia-associated peritoneal neutrophils. The presence or absence of eotaxin/CCL11 did not affect the number of peritoneal macrophages in these mice. These data indicate that eotaxin/CCL11 plays a novel regulatory role during the acute inflammatory response and suggest that constitutive expression of this chemokine within tissues such as the gut, lung, heart, and placenta might be important in downregulating acute inflammatory processes within these tissues. PMID- 12127050 TI - Antibodies to squalene in recipients of anthrax vaccine. AB - We previously reported that antibodies to squalene, an experimental vaccine adjuvant, are present in persons with symptoms consistent with Gulf War Syndrome (GWS) (P. B. Asa et al., Exp. Mol. Pathol 68, 196-197, 2000). The United States Department of Defense initiated the Anthrax Vaccine Immunization Program (AVIP) in 1997 to immunize 2.4 million military personnel. Because adverse reactions in vaccinated personnel were similar to symptoms of GWS, we tested AVIP participants for anti-squalene antibodies (ASA). In a pilot study, 6 of 6 vaccine recipients with GWS-like symptoms were positive for ASA. In a larger blinded study, only 32% (8/25) of AVIP personnel compared to 15.7% (3/19) of controls were positive (P > 0.05). Further analysis revealed that ASA were associated with specific lots of vaccine. The incidence of ASA in personnel in the blinded study receiving these lots was 47% (8/17) compared to an incidence of 0% (0/8; P < 0.025) of the AVIP participants receiving other lots of vaccine. Analysis of additional personnel revealed that in all but one case (19/20; 95%), ASA were restricted to personnel immunized with lots of vaccine known to contain squalene. Except for one symptomatic individual, positive clinical findings in 17 ASA-negative personnel were restricted to 4 individuals receiving vaccine from lots containing squalene. ASA were not present prior to vaccination in preimmunization sera available from 4 AVIP personnel. Three of these individuals became ASA positive after vaccination. These results suggest that the production of ASA in GWS patients is linked to the presence of squalene in certain lots of anthrax vaccine. PMID- 12127051 TI - APO(a) variants and lipoprotein(a) in men with or without myocardial infarction. AB - The lipoprotein Lp(a) with high plasma concentration is an independent genetic determinant for cardiovascular diseases. It was investigated as a quantitative factor of risk for myocardial infarction. A total of 345 Italian subjects, 127 Cases and 218 Controls, were studied. Lipids and lipoproteins were compared. Cases had atherogenic traits, such as lower HDL cholesterol and higher triglycerides than Controls. In particular, they had Lp(a) concentrations over the risk threshold, (median, 27 mg/dl in Cases vs 17 mg/dl in Controls; P = 0.0075, Mann-Whitney test) which confirmed the association of this parameter with the disease. Two main functional variants of the apo(a) gene, KringleIV and penta nucleotide repeat, (PNR) were analyzed. Allele and genotype frequency distributions differed between Cases and Controls. Lp(a) concentrations differed according to PNR genotypes in Controls: subjects having alleles >8 showed lower Lp(a). This was not found in Cases. They had a higher prevalence of the smaller KringleIV alleles, the high Lp(a)-expressing ones. In Cases, genotypes consisting of two small KringleIV alleles were prevalently associated to PNR 8/9 and 8/10, thus preventing Lp(a) lowering. The putative apo(a) enhancer within LINE1 in the apo(a)-plasminogen intergenic region was investigated for functional polymorphisms. No variants that could be associated to the Lp(a) variability were found. PMID- 12127052 TI - Parenchymal microglia of naive adult C57BL/6J mice express high levels of B7.1, B7.2, and MHC class II. AB - In this study, we addressed B7.1, B7.2, and MHC class II expression on microglia of normal adult C57BL/6J mice, which are susceptible to MOG35-55 peptide-induced experimental autoimmune encephalomyelitis (EAE). We showed that there are two distinct major populations of CD11b(+) cells in the central nervous system (CNS) of naive mice: CD45 low (CD45(lo); parenchymal microglia) and CD45 intermediate (CD45(int); CNS-associated macrophages). These two populations compose CNS microglia. There is a rare CD45 high (CD45(hi)) population. By contrast, splenic CD11b(+) cells (macrophages) are CD45(int) and CD45(hi), but rarely CD45(lo). CD45(lo)CD11b(+) cells constitutively express much higher levels of B7.1, B7.2, and MHC class II compared to CD45(int) CD11b(+) cells. A shift of CD11b(+) cells from CD45(lo) to CD45(int) was observed in the CNS of EAE mice. Our study provides evidence that (1) CD45(lo) and CD45(hi), but not CD45(int), could be unique markers to differentiate parenchymal microglia from infiltrating macrophages in EAE; (2) the level of CD45 expression on parenchymal microglia (CD45(lo)) was upregulated in EAE; and (3) parenchymal microglia in normal CNS could be potent APCs by expressing high levels of B7.1, B7.2, and MHC class II molecules and could therefore play an important role in inflammation and autoimmunity in the CNS. PMID- 12127053 TI - Effect of pro-inflammatory stimuli on tumor cell-mediated induction of endothelial cell adhesion molecules in vitro. AB - The object of our study was the question about the relevance of the tumor surrounding inflammatory cells with respect to the metastatic potential of the tumor cells. To imitate the role of inflammatory cells, three colon carcinoma (HT 29, HRT-18, and SW-620), one breast carcinoma (MCF-7), and one melanoma (ST-ML 12) cell lines were treated with pro-inflammatory stimuli, LPS, TNF-alpha, or IL 1beta. HUVEC monolayers were then stimulated by the collected supernatants (SN) of the tumor cells, following washing out of the applied stimuli. Analysis of CAM expression on HUVEC was performed using cell enzyme immunoassay. E-selectin, VCAM 1, and, in part, ICAM-1 were significantly up-regulated on HUVEC by exposure to SN of all LPS-stimulated tumor cells. This was especially the case for the colon carcinoma cell lines. A minimal increase of expression of VCAM-1 was observed after exposure to SN from TNF-alpha-stimulated HT-29 and MCF-7 cells. IL-1beta stimulation had no effect on endothelial CAM expression. These observations indicate that LPS could play a crucial role in tumor metastasis by inducing the release of soluble factors from different tumor cell lines capable of up regulating CAM expression. This might be of special significance in colon carcinomas, where a large source of bacterial LPS is available in the intestinal lumen. PMID- 12127054 TI - Neoplastic reversal of human ovarian carcinoma cells transfected with connexin43. AB - Gap junctional intercellular communication and expression of gap junction proteins (connexins) are decreased frequently in neoplastic cells including human ovarian carcinoma cells. In order to test the hypothesis that these changes contribute to the neoplastic phenotype of ovarian carcinoma cells, we transfected human ovarian carcinoma SKOV-3 cells with connexin43. Stable, connexin43 expressing transfectants were characterized for cell proliferation in vitro in normal, low-serum, and serum-free culture medium, for tumorigenicity in nude mice, and for sensitivity to adriamycin in vitro. Transfected clones expressed higher levels of connexin43 and gap junctional intercellular communication, reduced proliferation and greater dependence upon serum for growth in vitro, decreased tumor formation, increased sensitivity to adriamycin, and reduced expression of p-glycoprotein. These data suggest that gap junctional intercellular communication and/or connexin43 expression suppresses the neoplastic phenotype of ovarian carcinoma cells and their downregulation is involved in neoplastic transformation of ovarian epithelial cells. The increased sensitivity to adriamycin and elevated expression of p-glycoprotein by the transfected cells also suggest that gap junctional intercellular communication and connexin43 expression are involved in drug sensitivity and might be manipulated to enhance the clinical response. PMID- 12127055 TI - The use of FISH on breast core needle samples for the presurgical assessment of HER-2 oncogene status. AB - HER-2 status has been used in breast carcinoma as a prognostic marker to predict drug response and to select patients for trastuzumab treatment. Since immunohistochemistry (IHC) is thought to be less reliable, HER-2 testing with FISH is preferred. The analysis of HER-2 is usually performed on formalin-fixed paraffin tissue sections obtained from surgery. The use of paraffin sections is very time consuming and labor intensive. The objectives of this study were to (1) develop a simple and quick FISH protocol using touch imprints of breast core needle biopsies, eliminating the deparaffinization and pretreatment; and (2) make the HER-2 status available at the presurgical multidisciplinary treatment planning conference. A total of 50 core samples of breast carcinoma were obtained from image-guided core needle biopsy. Both FISH and IHC data were available for 46 cases. Forty-four of 46 cases (95.7%) were consistent. Two IHC 2+ cases were nonamplified (ratios of 0.99 and 1.09). It is expected that, in the near future, additional molecular markers will be used before surgery when the overall treatment plan is being developed. We conclude that HER-2 gene analysis by FISH on breast touch imprints is easily done and is a useful and reliable technique. PMID- 12127056 TI - Ultrasonographic evidence of phenotypic instability during hepatocarcinogenesis in N-nitrosomorpholine-treated rats. AB - Carcinogen-induced hepatoma in immunocompetent animal models has shown a progress similar to the clinical course of human hepatoma. Ultrasonography (US) was used for consecutive evaluation of the phenotypic changes in Sprague-Dawley rats exposed for 8 weeks to N-nitrosomorpholine (NNM, 200 mg/L). Three distinctive US findings were ascites, coarseness (defined as small and heterogeneously widespread increased echogenecity), and nodularity (defined as a >0.6-cm-sized echogenic region and clearly showing a tumor-like mass). Abdominal ascites was observed in 5 of 26 rats at week 8 NNM posttreatment and the number of rats showing ascites gradually increased. Coarseness (22 of 26 rats) and nodularity (1 of 18) appeared at weeks 8 and 17 NNM posttreatment, respectively. The gross and histological findings indicated that coarseness and nodularity shown in US reflected fibrosis and hepatocellular carcinoma or cholangiofibroma, respectively. The computer-aided quantification of coarseness and nodularity showed that the regression-linked phenotypic instability was present in coarseness but not in nodularity. We conclude that the heterogeneity of preneoplasia in NNM-treated rats might be induced by phenotypic instability rather than random initiating events of preneoplastic lesion. PMID- 12127057 TI - AT1 angiotensin II receptor mediates intracellular calcium mobilization in normal and cancerous breast cells in primary culture. AB - Angiotensin II (Ang II) increases intracellular calcium concentration ([Ca2+]i) in both normal and cancerous human breast cells in primary culture. Maximal [Ca2+]i increase is obtained using 100nM Ang II in both cell types; in cancerous breast cells, [Ca2+]i increase (delta[Ca2+]i) is 135+/-10nM, while in normal breast cells it reaches 65+/-5 nM (P<0.0001). In both cell types, Ang II evokes a Ca2+ transient peak mediated by thapsigargin (TG) sensitive stores; neither Ca2+ entry through L-type membrane channels or capacitative Ca2+ entry are involved. Type I Ang II receptor subtype (AT1) mediates Ang II-dependent [Ca2+]i increase, since losartan, an AT1 inhibitor, blunted [Ca2+]i increase induced by Ang II in a dose-dependent manner, while CGP 4221A, an AT2 inhibitor, does not. Phospholipase C (PLC) is involved in this signaling mechanism, as U73122, a PLC inhibitor, decreases Ang II-dependent [Ca2+]i transient peak in a dose-dependent mode.Thus, the present study provides new information about Ca2+ signaling pathways mediated through AT1 in breast cells in which no data were yet available. PMID- 12127058 TI - Src kinase induces calcium release in Xenopus egg extracts via PLCgamma and IP3 dependent mechanism. AB - Mobilization of intracellular calcium is an indispensable step of fertilization induced egg activation. Recently, this process has been shown to require the sequential activation of Src family tyrosine kinases, phospholipase Cgamma (PLCgamma), and inositol-1,4,5-trisphosphate (IP3)-dependent receptor of endoplasmic reticulum. In the present study, we made an attempt to recapitulate the early events of egg activation by stimulating Src kinase activity in the cell free extracts of Xenopus eggs. We found that enhanced Src kinase activity can initiate calcium response of low magnitude in cytostatic factor (CSF)-arrested mitotic extracts without releasing them into interphase. The addition of catalytically active recombinant Src kinase, as well as the activation of endogenous Xenopus Src family kinase by hydrogen peroxide (H2O2), increased total tyrosine phosphorylation, tyrosine phosphorylation of PLCgamma, and IP3 production in the extracts. The treatment with the Src family kinase-specific inhibitor, PP1, or PLC inhibitor, U73122, or IP3 receptor antagonist, heparin, prevented calcium release in the extracts. We conclude, therefore, that possible mechanism of Src/H2O2 action in the extracts might involve tyrosine phosphorylation and activation of PLCgamma, accompanied by the increase in IP3 content and subsequent calcium release from IP3-regulated calcium stores. These results also suggest that monitoring calcium signals induced in the Xenopus egg extracts by various components of signaling pathways may provide a particularly useful approach to investigating their role in the signal transduction. PMID- 12127059 TI - Modulation of gene expression in transgenic mouse hearts overexpressing calsequestrin. AB - Calsequestrin (CSQ) is the major Ca2+ binding protein of the cardiac sarcoplasmic reticulum (SR). Transgenic mice overexpressing CSQ at the age of 7 weeks exhibit concentric cardiac hypertrophy, and by 13 weeks the condition progresses to dilated cardiomyopathy. The present study used a differential display analysis to identify genes whose expressions are modulated in the CSQ-overexpressing mouse hearts to provide information on the mechanism of transition from concentric cardiac hypertrophy to failure. Cardiac ankyrin repeat protein (CARP), glutathione peroxidase (Gpx1), and genes which participate in the formation of extracellular matrix including decorin, TSC-36, Magp2, Osf2, and SPARC are upregulated in CSQ mouse hearts at 7 and 13 weeks of age compared to those of non transgenic littermates. In addition, two novel genes without sequence similarities to any known genes are upregulated in CSQ-overexpressing mouse hearts. Several genes are downregulated at 13 weeks, including SR Ca2+-ATPase (SERCA2) and adenine nucleotide translocase 1 (Ant1) genes. Further, a functionally yet unknown gene (NM_026586) previously identified in the mouse wolffian duct is dramatically downregulated in CSQ mice with dilated hearts. Thus, CARP, Gpx1, and genes encoding extracellular matrix proteins may participate in the development of cardiac hypertrophy and fibrosis, and changes in SERCA2, Ant1, and NM_026586 mRNA expression may be involved in transition from concentric to dilated cardiac hypertrophy. PMID- 12127060 TI - LY294002, but not wortmannin, increases intracellular calcium and inhibits calcium transients in bovine and human airway smooth muscle cells. AB - To characterize the effect that a phosphatidylinositol 3-kinase (PI3-kinase) inhibitor, LY294002, has on cytosolic calcium concentrations ([Ca2+]i), bovine airway smooth muscle cells (BASMC) and cultured human bronchial smooth muscle cells (HBSMC) were loaded with fura 2-AM, imaged as single cells and [Ca2+]i measured ratiometrically. LY294002 (50 microM) increased [Ca2+]i by 294+/-76 nM (P<0.01, n=13) and 230+/-31 nM (P<0.001, n=10) in BASMC and HBSMC, respectively, and increases occurred in the absence of extracellular calcium. In contrast, after pre-treatment with thapsigargin, LY294002 no longer increased [Ca2+]i. This calcium mobilization by LY294002 was associated with a significant functional effect since LY294002 also inhibited calcium transients to carbachol (45+/-23 nM), caffeine (45+/-32 nM), and histamine (20+/-22 nM), with controls of 969+/ 190, 946+/-156, and 490+/-28 nM, respectively. Wortmannin, a different PI3-kinase inhibitor, neither increased [Ca2+]i nor inhibited transients. Also, LY294002 increased [Ca2+]i in the presence of wortmannin, U-73122, and xestospongin C. We concluded that LY294002 increased [Ca2+]i, at least in part, by mobilizing intracellular calcium stores and inhibited calcium transients. The effects of LY294002 on [Ca2+]i were not dependent on wortmannin-sensitive PI3-kinases, phospholipase C, or inositol trisphosphate receptors (IP3R). For BASMC and HBSMC, LY294002 has effects on calcium regulation that could be important to recognize when studying PI3-kinases. PMID- 12127061 TI - Xestospongin C is a potent inhibitor of SERCA at a vertebrate synapse. AB - The specificity of action of Xestospongin C (XeC) towards the inositol 1,4,5 trisphosphate (IP3) receptor has been studied using the frog neuromuscular junction. In perisynaptic Schwann cells (PSCs), glial cells at this synapse, Ca2+ stores are dependent upon IP3 activation. Bath application of XeC (700 nM) caused a transient calcium elevation and blocked Ca2+ responses evoked in PSCs by synaptic activity or various agonists (ATP, muscarine, adenosine) only when Ca2+ stores had previously been challenged with local application of agonists. Moreover, XeC occluded the effects of thapsigargin (tg; 2 microM), a blocker of the Ca2+ ATPase pump of internal stores, which failed to evoke Ca2+ transients following 20 min of exposure to XeC. In nerve terminals, where the Ca2+ stores are ryanodine-sensitive, application of XeC (700 nM) prolonged the recovery phase of Ca2+ transients evoked by single action potentials, due to a prolonged Ca2+ clearance in the nerve terminal. No effects of tg (2 microM) were observed on Ca2+ response evoked by nerve stimulation when applied on the preparation after XeC (700 nM). Conversely, XeC (700 nM) had no effect on the shape and duration of Ca2+ entry in nerve terminals when tg was applied before XeC. These results indicate that XeC acts as an inhibitor of the sarcoplasmic/endoplasmic reticulum Ca2+ ATPase (SERCA) pump of internal stores. PMID- 12127062 TI - Xestospongin C empties the ER calcium store but does not inhibit InsP3-induced Ca2+ release in cultured dorsal root ganglia neurones. AB - The action of Xestospongin C (XeC) on calcium concentration in the cytosol ([Ca2+]i) and within the lumen of endoplasmic reticulum (ER) ([Ca2+]L) was studied using cultured dorsal root ganglia (DRG) neurones. Application of 2.5 microM of XeC triggered a slow [Ca2+]i transient as measured by Fura-2 video imaging. The kinetics and amplitude of XeC-induced [Ca2+]i response was similar to that triggered by 1 microM thapsigargin (TG). The [Ca2+]L was monitored in cells loaded with low-affinity Ca2+ indicator Mag-Fura-2. The cytosolic portion of Mag-Fura-2 was removed by permeabilisation of the plasmalemma with saponin. Application of XeC to these permeabilised neurones resulted in a slow depletion of the ER Ca2+ store. XeC, however, failed to inhibit inositol 1,4,5 trisphosphate (InsP3)-induced [Ca2+]L responses. We conclude that XeC is a potent inhibitor of sarco(endo)plasmic reticulum calcium ATPase, and it cannot be regarded as a specific inhibitor of InsP3 receptors in cultured DRG neurones. PMID- 12127063 TI - Untying the regulation of the Raf-1 kinase. AB - The Raf-1 kinase is the entry point to the mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK-1/2) signaling pathway, which controls fundamental cellular functions including proliferation, differentiation, and survival. As such, Raf-1 is regulated by complex mechanisms that are incompletely understood. Recent results have shown that release from repression is an important event that facilitates the interaction of Raf-1 with the Ras activator and its substrate, MAPK/ERK-1/2 kinase. A number of distinct activation steps contribute in a combinatorial fashion to regulate and adjust Raf-1 activity. The efficiency of downstream signal transmission is modulated by protein:protein interactions, and new data consolidate an important role for kinase suppressor of ras (KSR) as a scaffolding protein. KSR is a dynamic scaffold whose function and localization is regulated by phosphorylation. PMID- 12127064 TI - In vitro low-density lipoprotein oxidation by copper or *OH/O*(2)(-): new features on carbonylation and fragmentation of apolipoprotein B during the lag phase. AB - The aim of our study was to evaluate the carbonylation and the carbonylated fragmentation of apolipoprotein B upon low-density lipoprotein (LDL) oxidation induced in vitro by copper and *OH/O*(2)(-) free radicals generated by gamma radiolysis. Therefore, we developed a very sensitive Western blot immunoassay using 2,4-dinitrophenylhydrazine which allows the revelation of the apolipoprotein B carbonylation and its carbonylated fragmentation. The main results of this study show that (i) apolipoprotein B carbonylation is present during the lag phase of LDL oxidation in the two oxidative processes and (ii) apolipoprotein B carbonylated fragmentation was not detected during the lag phase of copper-oxidized LDL but was detected during the propagation phase. By contrast, apolipoprotein B carbonylated fragmentation was detected in the lag phase of *OH/O*(2)(-) oxidized LDL. PMID- 12127065 TI - Protein synthesis is required for optimal induction of 25-hydroxyvitamin D(3)-24 hydroxylase, osteocalcin, and osteopontin mRNA by 1,25-dihydroxyvitamin D(3). AB - The regulation of the 25-hydroxyvitamin D(3)-24-hydroxylase gene by 1,25 dihydroxyvitamin D(3) (1,25(OH)(2)D(3)) has been extensively studied. It is well established that two vitamin D response elements in the promoter are responsible for the 1,25(OH)(2)D(3) induction of transcription. Surprisingly, this induction is blocked by the protein synthesis inhibitor, cycloheximide (CHX). In AOK-B50 cells, 1,25(OH)(2)D(3) caused a large induction of 24-hydroxylase mRNA by 7h; however, the addition of CHX simultaneously or 2h after 1,25(OH)(2)D(3) addition caused 76.4+/-13.0 and 37.1+/-18.8% reductions in the mRNA, respectively. Addition of CHX 4h after 1,25(OH)(2)D(3) had the opposite effect, and 21.7+/ 17.2% more mRNA was observed after 7h. Similar patterns of mRNA expression were observed in other cell lines. CHX also decreased the induction by 1,25(OH)(2)D(3) of osteocalcin and osteopontin mRNA in ROS17/2.8 cells when added together with 1,25(OH)(2)D(3). The effect of CHX on the expression of a stably transfected luciferase construct under the control of 1400bp of 24-hydroxylase promoter indicates that a 1,25(OH)(2)D(3)-inducible transcription factor(s) that acts in the promoter region may at least in part be responsible for the effect of CHX on mRNA production of target genes. PMID- 12127066 TI - Role of cysteine 306 in the catalytic mechanism of Ascaris suum phosphoenolpyruvate carboxykinase. AB - Biochemical and metabolic data lead to the conclusion that the enzyme phosphoenolpyruvate carboxykinase (PEPCK) contributes to a critical point of divergence in energy conservation pathways between mammals and nematodes. The Ascaris suum PEPCK shares considerable homology with PEPCK from avian liver and is a good candidate for mutagenesis studies. The Cys306 substitution by Ser and Ala produced active enzymes and the two mutants are kinetically indistinguishable from each other. This substitution affects the catalytic affinity for the formation of the specific enzyme-nucleotide complex (k(cat)/K(m)) in the forward and reverse reactions. Studies with the substrate analogs 2(')dGDP and 2(')dGTP indicate that Cys306 in A. suum PEPCK is one of the residues important in nucleotide binding and may interact with the 2(')OH group in the ribose ring. Alternatively, mutation of this residue could cause protein changes that interfere with the proper conformation of the nucleotides for optimal catalysis to take place. PMID- 12127067 TI - Assessment of acrosomal status in rat spermatozoa: studies on carbohydrate and non-carbohydrate agonists. AB - In the mouse and several other species, including man, capacitated acrosome intact spermatozoa interact with natural [soluble zona pellucida (ZP) and progesterone (P4)] and synthetic [neoglycoproteins (ngps) and calcium (Ca(2+)) ionophore] agonists, prior to the initiation of a Ca(2+)-dependent signal transduction cascade. The net result is the fusion of the sperm plasma membrane overlying the outer acrosomal membrane at multiple sites and exocytosis of acrosomal contents [i.e., induction of the acrosome reaction (AR)]. This step is believed to be a prerequisite that enables the acrosome-reacted spermatozoon to penetrate the ZP and fertilize the egg. Although the rat is one of the most commonly used laboratory animals, very little is known about the chemical nature of agonists that induce the AR in this species. The lack of this information is primarily due to the fact that the rat sperm acrosome is a relatively thin structure. Thus, it is difficult to assess the status of the sperm acrosome in this species. In this report, we describe the use of a Coomassie brilliant blue dye staining procedure to assess the status of the rat sperm acrosome by light microscopy. The procedure is highly reproducible and has allowed us to determine the effects of carbohydrate (ngps and mouse ZP) and noncarbohydrate (P4 and Ca(2+) ionophore) agonists on capacitated spermatozoa. In addition, we have used a pharmacological approach to examine the functional significance of calmodulin (CaM), a Ca(2+)-binding protein, in induction of the AR in spermatozoa. Data presented in this report demonstrate that several ngps, solubilized mZP, P4, and Ca(2+) ionophores induce the AR in rat spermatozoa. Furthermore, we demonstrate that, whereas CaM antagonists blocked P4-induced AR, most of the inhibitors used had no significant effect on the Ca(2+) ionophore-induced (nonphysiological) AR. PMID- 12127068 TI - Kinetic and thermodynamic characterization of adenylyl cyclase from Euglena gracilis. AB - Some kinetic and thermodynamic properties of the plasma membrane adenylyl cyclase (AC) from the protist Euglena gracilis were examined. The AC kinetics for Mg-ATP was hyperbolic with a K(m) value of 0.33-0.43 mM, whereas the inhibition exerted by 2('),5(')-dideoxyadenosine was of the mixed type with a K(i) of 80-147 microM. The V(m) value (0.9 or 1.8 nmol(mg protein)(-1)min(-1)) changed, depending upon the carbon source in the growth medium (lactic acid or glutamate plus malate). Lactic acid membrane AC was slightly more thermolabile (from 28 to 40 degrees C) and showed higher activation energy (range 15-25 degrees C). With lactate, the total and saturated fatty acid percentage content in the plasma membrane was significantly greater than with glutamate plus malate, whereas the percentage content of polyunsaturated (n-3) fatty acids was lower. The data suggest that the fatty acid composition, as changed by the carbon source in the growth medium, may modulate the AC activity in Euglena. PMID- 12127069 TI - Oct2 transcription factor of a teleost fish: activation domains and function from an enhancer. AB - Oct2 transcription factors of the catfish (Ictalurus punctatus) are expressed as alternatively spliced alpha and beta isoforms. Functional analysis revealed an N terminal glutamine (Q)-rich transactivation domain common to both isoforms of catfish Oct2. A C-terminal proline, serine, threonine (PST)-rich activation domain was identified exclusively in the beta isoform. Activation domains of fish and mammalian Oct2 showed cell type- and species-specific activity correlated with their biochemical composition (Q-rich vs PST-rich). In contrast the activation domains of the aryl hydrocarbon receptor and aryl hydrocarbon receptor nuclear translocator of fish and mammals showed no correlation of activity with biochemical composition or species of origin. Although isolated catfish Oct2 activation domains were unable to drive transcription from a site 1.9kb distal to the promoter, Oct2beta activated transcription from both an IgH enhancer and an array of octamer motifs at this distal position. The properties of catfish Oct2 activation domains differ depending on whether they are studied in isolation or as components of the intact transcription factor. PMID- 12127070 TI - Modification phenomena of solid-state lignin caused by electron-abstracting oxidative systems. AB - Oxidative treatments of wood pulp lignin by one-electron-abstracting enzymatic or chemical systems result in modification phenomena which are not fully described in terms of those known from lignin model compound studies. The generation of, e.g., long-lived radicals necessitates nondestructive spectroscopic analysis of the lignin polymer for a proper characterization of these. The present work exposes a complexity of spectroscopic modification phenomena, which has not previously been realized. This is achieved by a laccase-mediator system, where the mediator is an aromatic low-molecular-weight compound, which mediates the electron abstraction between the lignin and the enzyme laccase. It is demonstrated that the modification generated exhibits qualitatively different temporal phases. The mechanisms are partly explained in terms of Marcus electron transfer theory, and it is suggested that these may play a role in the in vivo synthesis and degradation of lignin. PMID- 12127071 TI - Glc8 is a glucose-repressible activator of Glc7 protein phosphatase-1. AB - Regulation of Glc7 type 1 protein phosphatase stability and activity was studied in budding yeast. We found that the Glc7 protein has a half-life of over 180min, which is sufficient for several generations. Glc7 protein stability was constant during the cell cycle and in batch culture growth. Furthermore, deletion of regulatory subunit Gac1, Reg1, Reg2, Sds22, or Glc8 had no influence on Glc7 protein half-life. The activity of Glc7 assayed as okadaic acid-resistant phosphorylase phosphatase activity was constant during the cell cycle. Deletion of the aforementioned regulatory subunits revealed that only Glc8 deletion had a significant effect in reducing Glc7 activity. Glc7 activity was induced during stationary phase in a Glc8-dependent manner. In addition, extracellular glucose repressed the induction of Glc7 activity. These results are consistent with glucose repression of Glc8 expression and favor the role of Glc8 as a major Glc7 activator. PMID- 12127072 TI - Determinants within the C-terminus of the human norepinephrine transporter dictate transporter trafficking, stability, and activity. AB - The function of the human norepinephrine transporter (hNET) depends on its presence at the cell surface. A role for the hNET C-terminus in trafficking the transporter to the surface has been suggested by the report of a bovine NET C terminal splice variant that accumulates within heterologous host cells, and a human variant homolog has also been reported. We examined the relevance of the C terminus of hNET to trafficking and function using transfected LLC-PK1 cells. The intracellular and surface expression of NET proteins was evaluated by Western blots, and their functional capacities were assessed using transport assays. We found that the C-terminal residues encoded by hNET 1a enable the efficient maturation and surface expression of hNET and therefore critically impact transporter activity. Alternative splicing causes the retention of immature hNETs within the cell, whereas introduced C-terminal deletions result in significant degradation. The loss of the terminal isoleucine alone (Delta617-hNET) is sufficient to cause the degradation of hNET, an effect that can be mimicked by nonconservative point mutations at the terminal position. The phenotype of Delta617-hNET is recapitulated in neuronal SK-N-MC cells, but is significantly less severe in HEK-293 cells, suggesting a role for host cell factors in enabling the biosynthetic progression of wild-type hNET. Additional proximal residues may act at other steps to affect the expression of the fully mature protein on the cell surface (Q608A) and to more directly affect transporter activity (F609A). Together our studies document a critical contribution of the hNET C-terminus to transporter trafficking, stability, and function. PMID- 12127073 TI - Cell density inversely regulates D- and L-aspartate levels in rat pheochromocytoma MPT1 cells. AB - In a previous report (FEBS Lett. 434 (1998) 231), we demonstrated for the first time that D-aspartate (D-Asp) is synthesized in rat pheochromocytoma 12 (PC12) cells. This unique amino acid is believed to act as a novel messenger in mammalian cell regulation. However, the dynamics of D-Asp homeostasis in mammalian cells is yet to be elucidated. In this communication, we demonstrate that D-Asp is also synthesized in MPT1 cells (a subclone of PC12 cells) and that the D- and L-Asp levels in cells are regulated by cell density of the culture. Our data show that D-Asp levels increase, while in contrast, L-Asp levels decrease as a function of increased cell density. Conversely, in PC12 cells, which do not express the glutamate transporter involved in the incorporation of D and L-Asp into cells, L-Asp levels decrease upon cell density increase while D Asp concentrations remain almost unchanged. The results indicate that the biochemical behaviors of D- and L-Asp in mammalian cells are distinct and that the cellular levels of these stereoisomers appear to be under different control mechanisms. PMID- 12127074 TI - Alpha-neurotoxin gene expression in Naja sputatrix: identification of a silencer element in the promoter region. AB - Alpha-neurotoxin (alpha-NTX) from the venom of cobra, Naja sputatrix, is a highly lethal post-synaptic toxin that is responsible for the lethality caused by the venom. However, this toxin is found at low levels (3%) in the crude venom. The expression of its gene is determined by a promoter which is 90% similar to the promoter of another three-fingered toxin, cardiotoxin (CTX), which is produced in large amounts (60%) in the same venom. Functional analysis of the NTX-2 gene promoter demonstrated the presence of a silencer element of 24 nucleotides (nt 678 to -655) at its 5(') flanking region. This element has been found to play a major role in the down-regulation of NTX-2 gene expression. A point mutation on this silencer appears to attenuate its repressive property in CTX-2 gene. PMID- 12127075 TI - 4,4(')-Dianilino-1,1(')-binaphthyl-5,5(')-disulfonate: report on non-beta-sheet conformers of Alzheimer's peptide beta(1-40). AB - The venerable fluorescent probe of protein hydrophobic regions, 4,4(')-dianilino 1,1(')-binaphthyl-5,5(')-disulfonate (bis-ANS), unexpectedly increases in fluorescence with soluble beta(1-40) in acidic buffer solutions but reacts weakly with amyloid fibrils while other hydrophobic probes react with the fibrils. CD analysis correlates reaction with the probe with random coil/mixed conformations and alpha-helical forms of beta(1-40) in buffer solutions but less so with soluble beta-sheet forms or amyloid fibrils. The kinetics of the fluoroalcohol induced interconversion of conformers can be followed by changes in bis-ANS fluorescence. Formation of the beta-sheet form in aqueous buffer is limited by a slow component (minutes) while fluoroalcohol-promoted changes between beta-sheet and alpha-helix occur over seconds. Variants of beta(1-40) such as beta(1-42) or the Dutch E22Q mutation of beta(1-40) and fragments beta(1-28), beta(12-28), beta(10-20 amide), and beta(10-35 amide) react with bis-ANS under conditions that do not support fibril formation. Primary amino acid sequence is important as beta(1-11) does not cause bis-ANS fluorescence while beta(1-16) does, but hydrophobicity is not as beta(25-35) and beta(15-20 amide) are unreactive. bis ANS is a useful biophysical tool for characterizing particular, but not all, soluble Abeta conformations distinct from the fibrillar form of amyloid peptides detected by Thioflavin T. PMID- 12127076 TI - In vivo and in vitro phosphorylation of Candida albicans 20S proteasome. AB - In this paper we demonstrate that the Candida albicans 20S proteasome is in vivo phosphorylated and is a good in vitro substrate (S(0.5) 14nM) of homologous protein kinase CK2 (CK2). We identify alpha6/C2, alpha3/C9, and alpha5/Pup2 proteasome subunits as the main in vivo phosphorylated and in vitro CK2 phosphorylatable proteasome components. In vitro phosphorylation by homologous CK2 holoenzyme occurs only in the presence of polylysine, a characteristic that distinguishes the yeast proteasomes from mammalian proteasomes which are phosphorylated by CK2 in the absence of polycations. The major in vivo phosphate acceptor is the alpha3/C9 subunit, being phosphorylated in serine, both in vivo and in vitro. The phosphopeptides generated by endoproteinase Glu-C digestion from in vivo labeled alpha3/C9 subunit, from in vitro phosphorylation by homologous CK2 holoenzyme, and from the recombinant alpha3/C9 subunit phosphorylated by recombinant human CK2-alpha subunit are identical, suggesting that CK2 is likely responsible for in vivo phosphorylation of this subunit. Direct mutational analysis shows that serine 248 is the residue of the alpha3/C9 subunit phosphorylated by CK2. The in vitro stoichiometry of phosphorylation of the alpha6/C2 and alpha3/C9 proteasome subunits by CK2 can be estimated as 0.7 0.8 and 0.4-0.5 mol of phosphate per mole of subunit, respectively. These results are consistent with the relative abundance of the unphosphorylated and phosphorylated isoforms of these subunits present in the purified 20S proteasome preparation. Our demonstration of phosphorylation of C. albicans proteasome suggests that phosphorylation might be a general mechanism of regulation of proteasome activity. PMID- 12127077 TI - Cysteine 116 participates in intermolecular bonding of the human VEGF(121) homodimer. AB - VEGF(121), the 121-amino acid form of vascular endothelial growth factor is a homodimer with nine cysteine residues per monomer. While three intramolecular and two intermolecular disulfide bonds have been mapped, the state of the ninth cysteine, Cys116, is not known. In this study, we determined that human VEGF(121) contains a third interchain disulfide bond between Cys116 of each monomer. We also isolated a VEGF(121) variant with two extra cysteines bound to each Cys116. No evidence was found for the exsistence of Cys116 in the reduced state. In fact, selective reduction of the Cys116 interchain disulfide bond yielded an unstable VEGF(121) molecule, which reoxidized quickly. Biological activities of VEGF(121) Cys116 variants were assessed. The oxidative state of Cys116 has no effect on binding or proliferation activities but may be important for overall stability of the molecule. PMID- 12127078 TI - Activation of substrate/product couples by medium-chain acyl-CoA dehydrogenase. AB - Natural substrate/product binding activates medium-chain acyl-CoA dehydrogenase (MCAD) to accept electrons from its substrate by inducing a positive flavin midpoint potential shift. The energy source for this activation has never been fully elucidated. If ground-state alterations of the ligand, such as polarization, are entirely responsible for enzyme activation, the ligand potential should shift equally to that of the flavin but in the opposite direction. Ligand polarization is likely responsible for only a small portion of this activation. Here, thiophenepropionoyl- and furylpropionoyl-CoA analogs were used to directly measure the redox modulations of several ligand couples upon binding to MCAD. These measurements identified the thermodynamic contribution of ligand polarization to enzyme activation. Because the ligand potential alterations are significantly smaller than modulations in the flavin potential due to binding, other phenomena such as pK(a) changes, desolvation, and charge alterations are likely responsible for the thermodynamic modulations required for MCAD's activity. PMID- 12127079 TI - The plastid translocon component TOC36 exhibits an affinity for the bacterial protein translocation process. AB - The 44-kDa envelope polypeptides are active components of the plastid translocon, but their role in plastid protein import remains elusive. One form from Brassica napus (bnToc36B) was previously observed to exert a significant overall effect on bacterial protein translocation, but the nature of the influence requires further characterization. The experimental strategies employed in this study thus focus specifically on the nature of the bnToc36B-bacterial Sec translocon relationship to gain an understanding of Toc36's function. BnToc36B's presence in bacteria created a number of effects related to the protein transport process that together point to functional interactions with the bacterial Sec translocon. These effects are (1) reduced sensitivity to azide impairment as measured by a higher recovery rate from azide treatment, (2) reduced sensitivity to suboptimal temperatures manifesting as sustained levels of protein synthesis and translocation, (3) sustained levels of growth and beta-lactamase transport in high ampicillin concentrations, and (4) evidence for a physical affinity for the bacterial translocon. A reduction in overall SecA levels and a more stable SecA profile, when subjected to azide treatment, was observed in bnToc36B-containing bacteria. The implications of the bacterial data are discussed. PMID- 12127080 TI - Redox properties of an engineered purple Cu(A) azurin. AB - Purple Cu(A) centers are a class of binuclear, mixed-valence copper complexes found in cytochrome c oxidase and nitrous oxide reductase. An engineered Cu(A) protein was formed by replacing a portion of the amino acid sequence that contains three of the ligands to the native type I copper center of Pseudomonas aeruginosa azurin with the corresponding portion of sequence from the Cu(A) center of cytochrome c oxidase from Paracoccus denitrificans [Proc. Natl. Acad. Sci. USA 93 (1996) 461]. Oxidation-reduction midpoint potential (E(m)) values of the Cu(A) azurin of +399+/-10 and +380+/-2mV, respectively, were determined by cyclic voltammetry and spectrochemical titration. An n value of one was obtained, indicating that the redox reaction is cycling between the mixed valence and the fully reduced states. Whereas the E(m) value of native azurin is pH dependent, the E(m) value of Cu(A) azurin is not, as expected for the Cu(A) center. Similarities and differences in the redox properties are discussed in terms of the known crystal structures of Cu(A) centers in cytochrome c oxidase and Cu(A) azurin. PMID- 12127081 TI - Effects of fluoxetine treatment on carbohydrate metabolism in human blood platelets: a 1H-NMR study. PMID- 12127082 TI - UCP1 muscle gene transfer and mitochondrial proton leak mediated thermogenesis. AB - Mitochondrial uncoupling protein 1 (UCP1) mediates the thermogenic transport of protons through the inner mitochondrial membrane. This proton leak uncouples respiration from ATP synthesis. The current study assessed the possible contribution of UCP1 muscle gene transfer to impair mitochondrial respiration in a tissue lacking UCP1 gene expression. Rats received an intramuscular injection of plasmid pXC1 containing UCP1 cDNA in the right tibialis muscles, while left tibialis muscles were injected with empty plasmid as control. Ten days after DNA injection, mitochondria from tibialis anterior muscles were isolated and analyzed. UCP1 gene transfer resulted in protein expression as analyzed by inmunoblotting. Mitochondria isolated from UCP1-injected muscles showed a significant increase in state 2 and state 4 oxygen consumption rates and a decreased respiration control ratio in comparison to mitochondria from control muscles. Furthermore, UCP1-containing mitochondria had a lower membrane potential in those states (2 and 4) when compared with control mitochondria. Our results revealed that UCP1 muscle gene transfer is associated with an induced mitochondrial proton leak, which could contribute to increase energy expenditure. PMID- 12127083 TI - Regionally selective neurotoxicity of NMDA and colchicine is independent of hippocampal neural circuitry. AB - The mechanisms by which cerebral ischemia and several neurotoxins cause regionally selective damages to the hippocampal formation are largely unknown. The CA1-selective toxicity of N-methyl--aspartate (NMDA), the CA3-selective toxicity of kainate, and the dentate gyrus (DG)-selective toxicity of colchicine were observed in organotypic entorhino-hippocampal cultures. The selective neurotoxicity of NMDA and colchicine but not kainate was present in isolated tissue cultures of each hippocampal subregion, suggesting that the regional vulnerability is irrespective of the hippocampal trisynaptic pathway. Dispersed cultures of neurons prepared from Ammon's horn and the DG still exhibited a preference for susceptibility to NMDA and colchicine, respectively. Thus, the neurons per se appear to be inherently susceptible to specific toxins independently of their original loci, intrinsic neural circuits, vascular system, or other systemic factors. PMID- 12127084 TI - Effect of muscle electrostimulation on afferent activities from tibialis anterior muscle after nerve repair by self-anastomosis. AB - Numerous previous studies were devoted to the regeneration of motoneurons toward a denervated muscle after nerve repair by self-anastomosis but, to date, few investigations have evaluated the regeneration of sensory muscle endings. In a previous electrophysiological study (Decherchi et al., 2001) we showed that the functional characteristics of tibialis anterior muscle afferents are affected after self-anastomosis of the peroneal nerve even when the neuromuscular preparation was not chronically stimulated. The present study examines the regeneration of groups I-II (mechanosensitive) and groups III-IV (metabosensitive) muscle afferents by evaluating the recovery of their response to different test agents after self-anastomosis combined or not with chronic muscle stimulation for a 10-weeks period. We compared five groups of rats: C, control; L, nerve lesion without suture; LS, nerve lesion with suture; LSE(m): nerve lesion plus chronic muscle stimulation with a monophasic rectangular current; and LSE(b): nerve lesion plus chronic stimulation with a biphasic current with modulations of pulse duration and frequency, eliciting a pattern of activity resembling that delivered by the nerve to the muscle. Compared to the control group, (1) muscle kept only its original weight in the LSE(b) group, (2) in the LS group the response curve to tendon vibration was shifted toward the highest mechanical frequencies and the response of groups III-IV afferents after fatiguing muscle stimulation lowered, (3) in the LSE(m) group, the pattern of activation of mechanoreceptors by tendon vibrations was altered as in the LS group, and the response of metabosensitive afferents to KCl injections was markedly reduced, (4) in the LSE(b) group, the response to tendon vibration was not modified and the activation of metabosensitive units by increased extracellular potassium chloride concentration was conserved. Both LSE(b) and LSE(m) conditions were ineffective to maintain the post muscle stimulation activation of metabosensitive units as well as their activation by injected lactic acid solutions. Our data indicate that chronic muscle electrostimulation partially favors the recovery of mechano- and metabosensitivity in a denervated muscle and that biphasic modulated currents seem to provide better results. PMID- 12127085 TI - Comparison of matrix metalloproteinase expression during Wallerian degeneration in the central and peripheral nervous systems. AB - The matrix metalloproteinases (MMPs) are a large family of zinc-dependent enzymes which are able to degrade the protein components of the extracellular matrix. They can be placed into subgroups based on structural similarities and substrate specificity. Aberrant expression of these destructive enzymes has been implicated in the pathogenesis of immune-mediated neuroinflammatory disorders. In this study we investigate the involvement of MMPs, from each subgroup, in Wallerian degeneration in both the central and peripheral nervous systems. Wallerian degeneration describes the process initiated by transection of a nerve fibre and entails the degradation and removal of the axon and myelin from the distal stump. A similar degenerative process occurs as the final shared pathway contributing to most common neuropathies. MMP expression and localisation in the peripheral nervous system are compared with events in the CNS during Wallerian degeneration. Within 3 days after axotomy in the peripheral nervous system, MMP-9, MMP-7 and MMP-12 are elevated. These MMPs are produced by Schwann cells, endothelial cells and macrophages. The temporospatial expression of activated MMP-9 correlates with breakdown of the blood-nerve barrier. In the CNS, 1 week after optic nerve crush, four MMPs are induced and primarily localised to astrocytes, not microglia or oligodendrocytes. In the degenerating optic nerve, examined at later time points (4, 8, 12 and 18 weeks), MMP expression was down-regulated. The absence of MMPs in oligodendrocytes and mononuclear phagocytes during Wallerian degeneration may contribute to the slower removal of myelin debris observed in the CNS. The low level of the inactive pro-form of MMP-9 in the degenerating optic nerve may explain why the blood-brain barrier remains intact, while the blood-nerve barrier is rapidly broken down. We conclude that the difference in the level of expression, activation state and cellular distribution of MMPs may contribute to the different sequence of events observed during Wallerian degeneration in the peripheral compared to the CNS. PMID- 12127086 TI - Behavioral and neurochemical characterization of alpha(2A)-adrenergic receptor knockout mice. AB - Genetic manipulation of mice now provides new tools to evaluate the biological functions of the alpha(2)-adrenergic receptor (alpha(2)-AR) subtypes (alpha(2A), alpha(2B), and alpha(2C)). To investigate the role of the alpha(2A)-AR in the modulation of mouse primary behavioral characteristics and brain neurochemistry, mice with targeted inactivation of the gene for the alpha(2A)-AR were compared with wild-type C57BL/6 control animals. First, a comprehensive behavioral screen was employed to provide a detailed characterization of basic neurologic functions. Thereafter, the mice were analyzed in three models of anxiety, i.e. the elevated-plus maze test, the marble burying test and the open field test. The diurnal activity pattern of the mice was assessed in a 24-h locomotor activity test. Furthermore, receptor autoradiography of the brain was performed using the subtype-non-selective alpha(2)-AR antagonist radioligand [(3)H]RS-79948-197. Lack of the alpha(2A)-AR was associated with alterations in autonomic functions, including increased heart rate and piloerection. The mutant mice also exhibited impaired motor coordination skills, increased anxiety-like behavior and an abnormal diurnal activity pattern. In addition, neurochemical analysis of monoamine neurotransmitters revealed a considerable increase in brain norepinephrine turnover in mice lacking alpha(2A)-AR. Our results provide further support for the crucial role of the alpha(2A)-AR in modulating brain noradrenergic neurotransmission and many aspects of mouse behavior and physiology. PMID- 12127087 TI - Seladin-1 transcription is linked to neuronal degeneration in Alzheimer's disease. AB - Seladin-1 is a gene recently shown to be down-regulated in brain regions selectively degenerated in Alzheimer's disease. The sequence of seladin-1 shares similarities with flavin-adenine-dinucleotide-dependent oxidoreductases and it has been found to protect cells from apoptotic cell death. In this work, we show that the transcription of seladin-1 is selectively down-regulated in the brain areas affected in Alzheimer's disease. The down-regulation in seladin-1 transcription was associated with hyperphosphorylated tau seen as linkage to immunohistochemically detected paired helical filament tau, neuritic plaques and neurofibrillary tangles. In contrast, no association was found between seladin-1 transcription and beta-amyloid deposition when analyzing human samples or tissue from transgenic animals. Furthermore, the relative transcription of seladin-1 was found to fluctuate during aging in the transgenic mouse model of Alzheimer's disease. The fluctuation was enhanced by Alzheimer's disease causing mutations in presenilin-1 and amyloid precursor protein genes. Finally, seladin-1 transcription was found to be up-regulated in mouse N2a cells induced to undergo apoptosis with okadaic acid. The results presented here indicate that seladin-1 transcription is selectively down-regulated in brain regions vulnerable to Alzheimer's disease and this down-regulation is associated with the hyperphosphorylation of tau protein. PMID- 12127088 TI - Response of GABAergic cells in the deep mesencephalic nucleus to dopaminergic cell degeneration: an electrophysiological and in situ hybridization study. AB - The deep mesencephalic nucleus (DMN) is a large midbrain reticular region located between the substantia nigra compacta and the superior colliculus. It contains GABAergic cells that share striatal afferents, thalamic and collicular efferents, as well as neurochemical and electrophysiological similarities, with those of the substantia nigra reticulata. In the present paper we used electrophysiological (firing rate and firing pattern) and morphological (densitometric analysis of in situ hybridization histochemical labeling for glutamic acid decarboxylase (GAD)65 and GAD67 mRNA) techniques, to study the response of DMN GABAergic cells to the degeneration of nigral dopaminergic cells. Our results showed that unilateral dopaminergic cell loss (after injection of 6-hydroxydopamine in the medial forebrain bundle) induces a bilateral and symmetrical increase in both firing rate and GAD67 mRNA levels and a decrease in GAD65 mRNA levels. These findings support the involvement of DMN GABAergic cells in the basal ganglia modifications that follow dopaminergic cell loss, also suggesting its participation in the pathophysiology of Parkinson's disease. The symmetry of effects, together with its recently reported bilateral projections to the thalamus and superior colliculus, suggest that unlike substantia nigra reticulata, DMN is involved in the interhemispheric regulation of basal ganglia, probably keeping their functional symmetry even after asymmetric lesions. PMID- 12127089 TI - Effects of sleep deprivation on extracellular serotonin in hippocampus and frontal cortex of the rat. AB - Sleep deprivation improves the mood of depressed patients, but the exact mechanism behind this effect is unclear. An enhancement of serotonergic neurotransmission has been suggested. In this study, we used in vivo microdialysis to monitor extracellular serotonin in the hippocampus and the frontal cortex of rats during an 8 h sleep deprivation period. These brain regions were selected since both have been implicated in depression. The behavioral state of the animal was continuously monitored by polygraphic recordings during the experiment. Sleep deprivation produced a gradual decline in extracellular serotonin levels, both in the hippocampus and in the frontal cortex. In order to investigate whether the reduction in serotonin was due to other factors than sleep deprivation, i.e. time of day effect, another experiment was performed. Here animals were allowed to sleep during most of the recording period. This experiment showed the expected changes in extracellular serotonin levels: consistently higher levels in the awake, non-sleep deprived animals compared to during sleep, but no time of day effect. The reduction in extracellular serotonin during sleep deprivation may suggest that serotonin does not play a major role in the mood-elevating effect of sleep deprivation. However, since 5-HT levels are strongly behavioral state dependent, by eliminating sleep, there may be a net increase in serotonergic neurotransmission during the sleep deprivation period. PMID- 12127090 TI - Muscarinic acetylcholine receptors induce neurite outgrowth and activate the synapsin I gene promoter in neuroblastoma clones. AB - The possible role of acetylcholine as a modulator of neuronal differentiation has been tested using a neuroblastoma cell line (N18TG2), which does not synthesize any neurotransmitter. Acetylcholine synthesis has been activated in this line by transfection with a construct containing a choline acetyltransferase (ChAT) cDNA; ChAT-positive clones share a higher ability to grow fibers and an activation of synapsin I expression compared to the parental cells. Atropine, a muscarinic antagonist, abolishes the higher ability to grow fibers of ChAT-positive transfected clones, and the cholinergic agonist carbachol induces higher neurite outgrowth in the parental line. In transient transfections of ChAT-positive clones, the expression of a reporter gene under the control of synapsin I promoter is considerably reduced by atropine, while it is not modified by carbachol; in contrast, in the parental cells, which do not synthesize acetylcholine, the reporter gene expression is induced by carbachol and this effect is abolished by atropine. The data presented provide evidence for the existence of a direct modulation of fiber outgrowth and synapsin I expression by muscarinic receptor activation, which may be related to early growth response gene-1 (EGR-1) levels. PMID- 12127091 TI - The role of mu-opioid receptors in inflammatory hyperalgesia and alpha 2 adrenoceptor-mediated antihyperalgesia. AB - The purpose of the present study was to investigate the role of mu-opioid receptor in inflammatory hyperalgesia in intact and in spinalized animals and the interaction between mu-opioid and alpha2-adrenergic receptor in acute pain and inflammatory hyperalgesia. Behavioral responses to mechanical and heat stimuli were studied in mu-opioid receptor knockout mice and wildtype control mice. Thermal nociception was evaluated by measuring paw withdrawal latencies to radiant heat applied to the hindpaws. Mechanical nociception was measured by von Frey monofilament applications to the hindpaws. Intraplantar carrageenan-induced (1 mg/40 microl) mechanical and heat hyperalgesia were compared in micro-opioid knockout and wildtype mice. The effect of systemically administered alpha2 adrenergic receptor agonist dexmedetomidine (1-10 microg/kg) was evaluated on mechanical and thermal withdrawal responses under normal and inflammatory state in knockout and wildtype mice. The role of micro-opioid receptor in descending modulation of nociception was studied by assessing mechanical and heat withdrawal responses before and after mid-thoracic spinalization. Withdrawal responses to radiant heat and von Frey monofilaments were similar in mu-opioid knockout and wildtype mice before and after the carrageenan induced hindpaw inflammation. Also, antinociceptive effects of dexmedetomidine in thermal and mechanical nociceptive tests were similar before carrageenan induced hindpaw inflammation. However, the potency of dexmedetomidine was significantly reduced in carrageenan induced mechanical hyperalgesia in mu-opioid knockout mice compared to the wildtype control mice. Thermal and mechanical withdrawal responses were similar between mu-opioid knockout and wildtype mice before and after mid-thoracic spinalization. Our observations indicate that the mu-opioid receptors do not play an important role in alpha2-adrenergic receptor agonist-mediated acute antinociception. In addition, micro-opioid receptors are not tonically involved in the modulation of inflammation-induced mechanical and thermal hyperalgesia, and the supraspinal control of spinal reflexes. However, in the presence of inflammation, mu-opioid receptors play an important role in the antihyperalgesic actions of an alpha2-adrenergic receptor agonist. PMID- 12127093 TI - Peripheral stimuli excite coronal beams of Golgi cells in rat cerebellar cortex. AB - Cerebellar granule cells constitute the largest neurone population of the brain. Their axons run as parallel fibres along the coronal axis, and the one dimensional spread of excitation that is expected to result from this arrangement is a key assumption of theories of cerebellar function. In many studies using various techniques, however, it was not possible to evoke such a beam-like propagation of excitation with natural stimuli. We recorded, in Crus I and II of anaesthetised rats, pairs of Golgi cells aligned along the parallel fibre axis and synchronising spontaneously. Each pair was subjected to two stimulation protocols: punctate and semi-continuous. Local punctate facial stimulation evoked distinct fast and late responses of variable strength and latency (fast: 4.0-10.2 ms; late: 13.6-22.7 ms). Semi-continuous stimulation with a brush increased the firing rate, and modified the precision and phase of synchronisation. Differences between a pair in response strength and phase to brush stimulation correlated strongly with the difference in latency to punctate stimulation. These observations were reproduced in a model of the granular layer. The stimulus activated a central patch of mossy fibres, and Golgi cells received short- and long-range excitation from mossy and parallel fibres, respectively. The strength and latency of the punctate response of a model Golgi cell were found to vary with its position, reflecting a systematic change in the contribution of mossy and parallel fibres to its excitation with distance from the activated patch. During brush stimulation, model Golgi cells inside the patch fired more precisely synchronised, whereas the other Golgi cells responded with a lag proportional to their distance from the patch, thereby reproducing the experimentally observed changes in synchronisation. Taken together with the previously reported large receptive fields of Golgi cells and with their spontaneous synchronisation, the variable, position-dependent latency of evoked Golgi cell responses indicates a beam-like spread of excitation along the parallel fibres in rat cerebellar cortex. PMID- 12127092 TI - Effects of two selective phosphodiesterase type 5 inhibitors, sildenafil and vardenafil, on object recognition memory and hippocampal cyclic GMP levels in the rat. AB - The present study investigated the effects of two cyclic GMP-specific phosphodiesterase enzyme type 5 inhibitors, sildenafil and vardenafil, on the memory performance in the object recognition task. Both compounds were given per orally (1, 3 and 10 mg/kg sildenafil; 0.1, 0.3, 1 and 3 mg/kg vardenafil) immediately after the exposure to two identical objects. The memory for the objects was tested 24 h later. Vehicle-treated rats spent equal times exploring a new and the familiar object demonstrating that they did not remember the familiar one. However, sildenafil improved the object discrimination performance of the rats with a high discrimination performance at a dose of 3 mg/kg. Rats treated with vardenafil also showed an improved object discrimination performance. Compared with sildenafil, vardenafil appeared to be even more potent in this respect since it already produced a high discrimination performance at a dose of 0.3 mg/kg. The effects of both compounds on cyclic GMP and cyclic AMP accumulation were studied in rat hippocampal slices incubated in vitro. Cyclic GMP levels were increased after incubation with the highest concentration of 100 microM vardenafil (together with 0.1 mM sodium nitroprusside), although no changes in cyclic GMP levels were detected after incubation with different concentrations of sildenafil. Both compounds had no effect on cyclic AMP levels. Additional cyclic GMP immunocytochemistry showed that incubation with vardenafil (in the presence of sodium nitroprusside) resulted in a concentration-dependent staining of cyclic GMP. Staining was predominantly found in neuronal fibres in the hippocampal CA2/CA3 region. It was already detected at a concentration of 0.1 microM vardenafil. Also positive fibres were detected after incubation with sildenafil but at a higher concentration of 10 microM. Taken together, these results suggest that inhibition of phosphodiesterase enzyme type 5 improves object recognition memory. This effect might be explained by increased levels of central cyclic GMP. PMID- 12127094 TI - Cerebral activation by the signals ascending through unmyelinated C-fibers in humans: a magnetoencephalographic study. AB - Cerebral processing of first pain, associated with A delta-fibers, has been studied intensively, but the cerebral processing associated with unmyelinated C fibers, relating to second pain, remains to be investigated. This is the first study to clarify the primary cortical processing of second pain by magnetoencephalography, through the selective activation of C-fibers, by the stimulation of a tiny area of skin with a CO2 laser. In the hemisphere contralateral to the side stimulated, a one-source generator in the upper bank of the Sylvian fissure (secondary somatosensory cortex, SII) or two-source generators in SII and the hand area of the primary somatosensory cortex (SI) were the optimal configurations for the first component 1M. The onset and peak latency of the two sources in SI and SII were not significantly different. In the hemisphere ipsilateral to the stimulation, only one source was estimated in SII, and its peak latency was significantly (approximately 18 ms on average) longer than that of the SII source in the contralateral hemisphere. From our findings we suggest that parallel activation of SI and SII contralateral to the stimulation represents the first step in the cortical processing of C-fiber-related activities, probably related to second pain. PMID- 12127095 TI - Inflammation induces serine protease inhibitor 3 expression in the rat pineal gland. AB - In the rat pineal gland, prominent expression of serine protease inhibitor 3 (SPI 3) mRNA is seen after systemic injection of lipopolysaccharide. The up-regulation of SPI-3 mRNA expression is also confirmed by northern blotting. Most SPI-3 mRNA positive cells simultaneously express synaptophysin, a marker for pinealocytes, but not glial fibrillary acidic protein, a marker for astrocytes. This indicates that SPI-3 mRNA-positive cells are pinealocytes. Almost all SPI-3 mRNA-positive cells also showed translocation of the signal transducers and activators of transcription 3 (STAT3) into nuclei after lipopolysaccharide injection. These data support previous in vitro results that SPI-3 expression is induced in a STAT3-mediated manner. In addition, the expression of ciliary neurotrophic factor receptor (CNTFR) and leukemia inhibitory factor receptor (LIFR) mRNAs, but not of interleukin 6 receptor mRNA, was up-regulated after systemic lipopolysaccharide treatment. Because these receptors are upstream of STAT3, the present results suggest that cytokines such as LIF and/or CNTF induce SPI-3 expression via STAT3 in the pineal gland in response to inflammatory stimulus. We conclude that although the functional consequences of SPI-3 in the pineal gland during systemic inflammation are unknown, SPI-3 may have a crucial role in preventing some degenerative proteolysis induced by inflammatory stimuli. PMID- 12127096 TI - Stress during adolescence alters behavioral sensitization to amphetamine. AB - In humans, chronic intermittent and uncontrollable stress during adolescence is viewed as a key factor for vulnerability to drug abuse and development of psychopathologies later in life. Less is known about the long-term effects of chronic stress in animals during the juvenile period. Although there is evidence of cross sensitization during prenatal period and adulthood between chronic stress and amphetamine-induced behavioral sensitization in the rat, no studies have been conducted on cross sensitization between chronic variable stress in adolescence and behavioral sensitization to amphetamine. To address this question, at the onset of adolescence (28 days) male rats were subjected to 28 days of intermittent non-habituating social stress (isolation, novel environment, crowding, litter-shifting, subordination), or physical stress (restraint, swim, cold, ether, noise), or were handled as controls. Twenty-four hours after the last stressor or handling, all groups were exposed to a novel environment for 1 h, after which they underwent a regimen of behavioral sensitization to amphetamine. Our results showed that socially stressed rats have low locomotor activity in the novel environment, when compared to the control and physical groups who were identical in the same test. Even though socially stressed rats had lower locomotor activity in response to amphetamine injections, there were no significant differences during the training phase between the three groups at this dose of amphetamine. However, when tested for behavioral sensitization to amphetamine control and physically stressed rats showed a robust sensitization, socially stressed rats were significantly inhibited. We conclude that our chronic variable social stress protocol during adolescence inhibits behavioral sensitization to amphetamine during adulthood. PMID- 12127097 TI - Effects of chronic restraint stress and estradiol on open field activity, spatial memory, and monoaminergic neurotransmitters in ovariectomized rats. AB - Twenty-one days of chronic restraint stress impairs male rat performance on the radial arm maze [Luine et al. (1994) Brain Res. 639, 167-170], but enhances female rat performance [Bowman et al. (2001) Brain Res. 904, 279-289]. To assess possible ovarian hormone mechanisms underlying this sexually dimorphic response to stress, we examined chronic stress effects in ovariectomized rats. Ovariectomized rats received Silastic capsule implants containing cholesterol or estradiol and were assigned to a daily restraint stress (21 days, 6 h/day) or non stress group. Following the stress period, subjects were tested for open field activity and radial arm maze performance. Stress and estradiol treatment affected open field activity. All stressed animals, with or without estradiol treatment, made fewer total outer sector crossings. In contrast, estradiol-treated animals, with or without stress, made more inner sector visits, an indication that estradiol decreased anxious behavior on the open field across time. As measured by the total number of visits required to complete the task, stress did not affect radial arm maze performance in ovariectomized rats, but estradiol-treated animals, with or without stress, performed better than non-treated animals on the radial arm maze. Stressed subjects receiving estradiol showed the best radial arm maze performance. Following killing, tissue samples were obtained from various brain regions known to contribute to learning and memory, and monoamine and metabolite levels were measured. Several changes were observed in response to both stress and estradiol. Most noteworthy, stress treatment decreased homovanillic acid levels in the prefrontal cortex, an effect not previously observed in stressed intact females. Estradiol treatment increased norepinephrine levels in CA3 region of the hippocampus, mitigating stress-dependent changes. Both stress and estradiol decreased dentate gyrus levels of 5-hydroxyindole acetic acid. In summary, the current study provides novel information showing that estradiol alters behavioral and neurochemical responses to stress in ovariectomized rats. Estradiol treatment decreased anxious behavior on the open field and stressed animals receiving estradiol had enhanced radial arm maze performance. In relation to interactions between stress and estradiol on cognition and anxiety, changes in the prefrontal cortex dopaminergic system, dentate gyrus serotonergic system, and norepinephrine levels in the CA3 region appear important. Results show that estradiol may moderate stress effects on cognition and anxiety through both organizational and activation effects. PMID- 12127098 TI - The influence of developmental period of aluminum exposure on synaptic plasticity in the adult rat dentate gyrus in vivo. AB - Previous studies from our group have demonstrated that chronic aluminum exposure from parturition throughout life impairs both long-term potentiation (LTP) and long-term depression (LTD) of the excitatory postsynaptic potential (EPSP) slope and reduces the population spike (PS) amplitude in the rat dentate gyrus in vivo. The present study sought to extend these findings by evaluating the developmental periods critical for aluminum-induced impairment of synaptic plasticity. Rats were exposed to aluminum (gestational, lactational and postlactational) through drinking 0.3% aluminum chloride in water over different developmental intervals: (1) prenatal exposure; (2) beginning from birth and terminating at weaning; (3) beginning at weaning throughout life; (4) beginning at birth and continuing throughout life. As adults (postnatal day 80-100), field potentials were measured in the dentate gyrus of hippocampus in response to stimulation applied to the lateral perforant path. The results showed: (1) Prenatal aluminum exposure had no effect on the magnitude of LTP as measured by the EPSP slope and LTD as measured for the PS amplitude, while it had a small effect on the magnitude of LTP as measured for the PS amplitude and LTD as measured by the EPSP slope. (2) Lactational, postlactational and throughout life exposure to aluminum impaired both LTP and LTD of the EPSP slope and PS amplitude, except that LTD of PS amplitude was not significantly changed in animals postlactationally exposed. (3) Aluminum exposure from parturition throughout life caused the greatest impairment of the range of synaptic plasticity, while prenatal aluminum exposure caused the least. From these results we conclude that the lactational period was the most susceptible to aluminum-induced impairment of synaptic plasticity and that chronic aluminum exposure from parturition throughout life is extremely disruptive to synaptic plasticity and should be avoided. PMID- 12127099 TI - Strain differences in urocortin expression in the Edinger-Westphal nucleus and its relation to alcohol-induced hypothermia. AB - The Edinger-Westphal nucleus is the primary source of urocortin in rodent brain. Mapping of inducible transcription factors has shown that the Edinger-Westphal nucleus is preferentially sensitive to ethanol self-administration. In the present study we have immunohistochemically compared expression of urocortin and c-Fos in naive and ethanol-treated C57BL/6J and DBA/2J mouse inbred strains. We found that C57BL/6J mice possess significantly higher numbers of urocortin expressing cells in the Edinger-Westphal compared to DBA/2J mice. Subsequent histological analysis confirmed a lower number of large neurons in the DBA/2J Edinger-Westphal nucleus. Surprisingly, despite the differences in structure, no strain differences were observed in the number of c-Fos-containing cells after acute (0.6-4.8 g/kg, i.p.) and repeated (2.4 g/kg, 14 days, one injection/day) administration of ethanol. Double-label immunohistochemistry showed that ethanol induced c-Fos expression is present in different sets of Edinger-Westphal cells between the strains. Specifically, expression of c-Fos in C57BL/6J mice is preferentially induced in urocortin cells, while c-Fos in DBA/2J mice occurs in a mixed population of cells. Behavioral analysis of the B6D2 F2 intercross, a heterogeneous mouse strain, showed that the number of urocortin cells is positively correlated with basal temperatures and ethanol-induced hypothermia. Involvement of the Edinger-Westphal in alcohol-induced hypothermia is further confirmed by analysis of urocortin cells in the HOT/COLD selected lines. These results provide evidence that C57BL/6J and DBA/2J mice have structural differences in the Edinger-Westphal that can result in activation of different populations of neurons upon alcohol intoxication contributing to differential thermoregulation between these inbred strains. PMID- 12127100 TI - Central pathways in the pons and midbrain involved in cardiac sympathoexcitatory reflexes in cats. AB - Activation of cardiac sympathetic afferents elicits pain and excitatory cardiovascular reflexes including acute hypertension and tachyarrhythmias. Our previous studies have shown that specific regions in the medulla, such as the nucleus of solitary tract and ventrolateral medulla, are involved in central regulation of cardiac sympathoexcitatory reflexes. However, the contributions of supramedullary nuclei to these reflexes have not been characterized. In the present study, we located activated neurons in the pons and midbrain induced by inputs from cardiac sympathetic afferents by detecting their c-Fos immunoreactivity. In anesthetized cats with bilateral carotid denervation and cervical vagotomy, epicardial application of bradykinin (1-10 microg, in 0.1 ml; n=7) was performed on the anterior surface of the left ventricle six times, every 20 min. Repetitive application of bradykinin caused consistent excitatory cardiovascular reflexes characterized by increases in blood pressure and heart rate. No responses were evoked by the vehicle for bradykinin (0.9% saline, n=7). Compared to control cats, c-Fos immunoreactive cells were significantly increased (P<0.05) in the rostral pons, caudal and intermediate midbrain in the bradykinin treated cats. The specific areas activated include the parabrachial nucleus, Kolliker-Fuse nucleus, locus coeruleus, dorsal nucleus of raphe, and dorsal, lateral and ventrolateral periaqueductal gray. From these results we suggest that cardiovascular-related regions in the pons and midbrain form part of a long loop in central integration of cardiac sympathoexcitatory reflexes. PMID- 12127101 TI - Immunocytochemical localization of neuronal calcium sensor-1 in the hippocampus and cerebellum of the mouse, with special reference to presynaptic terminals. AB - Neuronal calcium sensor-1 (NCS-1) is a member of the EF-hand calcium-binding protein superfamily, which is considered to modulate synaptic transmission and plasticity. In this work, we first examined the distribution patterns of NCS-1 in the hippocampus and cerebellum. The intense NCS-1-immunoreactive (IR) elements in the hippocampus were restricted to dendritic layers, while those in the cerebellum occurred in both dendritic and cellular layers. Then, we examined the exact localization of NCS-1 using immunofluorescent double labeling for NCS-1 and synaptophysin, a marker of presynaptic terminals. In the hippocampus, the mossy fiber systems (terminals and bundles) exhibited intense NCS-1 immunoreactivity. On the other hand, the presumed principal cell dendrites were also NCS-1-IR in the stratum lacunosum-moleculare of Ammon's horn and molecular layer of the dentate gyrus, where NCS-1-IR elements and synaptophysin-IR presynaptic terminals showed characteristic complementary distribution patterns. In the cerebellum, some of the basket cell axon terminals surrounding the somata of Purkinje cells exhibited NCS-1 immunoreactivity, while the pinceau showed consistent labeling for NCS-1. Higher magnification observations revealed that the NCS-1-IR presumed granule cell dendrites and synaptophysin-IR mossy fiber terminals in the glomeruli of the cerebellum showed characteristic complementary distribution patterns. Furthermore, we estimated quantitatively the relative amount of NCS-1 in the presynaptic terminals in individual layers, and confirmed that the mossy fiber terminals in the hippocampus contained comparatively high amounts of NCS-1. These results showed the diverse localization of NCS-1 in pre- and/or postsynaptic elements of the hippocampus and cerebellum, and suggest potential roles in specific synaptic transmission. PMID- 12127102 TI - Differential localization of alpha-, beta- and gamma-synucleins in the rat CNS. AB - Alpha-synuclein is a presynaptic protein that normally participates in the homeostasis of synaptic vesicles. Missense mutations in its gene cause the protein to participate actively in the development of heritable forms of Parkinson's disease. Moreover, its metabolism is perturbed in all cases of Parkinson's disease where alpha-synuclein accumulates in a filamentous form in the Lewy body nerve cell lesion. Lewy bodies also develop in other common neurodegenerative disorders, like dementia with Lewy bodies and Lewy body variant of Alzheimer's disease. In the present study, we have studied the detailed distribution of alpha-, beta- and gamma-synuclein in the rat CNS. Alpha-synuclein was not observed in perikarya, but was distributed with high intensity in nerve terminals in the caudate and putamen and ventral pallidum, where beta-synuclein was much weaker and less densely distributed in the caudate and putamen. Gamma synuclein was not found in the caudate and putamen. Alpha-synuclein was robustly distributed in the substantia nigra pars reticulata, but was very weak or virtually absent from the perikarya of the neurons in the pars compacta. In contrast, beta-synuclein was very weak or absent from the substantia nigra. gamma Synuclein was absent from the terminals of substantia nigra pars reticulata, but sparsely distributed gamma-synuclein-containing neurons were detected in the substantia nigra pars compacta. In the brainstem, alpha-synuclein as well as gamma-synuclein were present in the locus coeruleus with high intensity, while beta-synuclein was very weak. In addition, alpha-synuclein was intense in the vagus nucleus, but weak in the oculomotor, facial, hypoglossal, accessory and ambiguous nuclei, where beta-synuclein was very intensely present. Furthermore, gamma-synuclein was localized in the terminals and in cell bodies of the Edinger Westphal nucleus, the red nucleus, locus coeruleus, and most cranial nerve related nuclei. In the spinal cord, alpha- and gamma-synucleins were intensely present in laminae I and II and in the preganglionic sympathetic nuclei, whereas beta-synuclein was very weak. These results indicate that alpha-synuclein is abundant in central catecholaminergic regions. Beta-synuclein is more localized in the somatic cholinergic components, while it is particularly weak or absent from catecholaminergic neurons. Gamma-synuclein appears to be present in both cholinergic and catecholaminergic regions, but very weak in the forebrain. PMID- 12127103 TI - Ciliary muscle in avian is derived from mesenchymal and epithelial cells. AB - It has long been maintained that the ciliary muscle derives from mesenchymal cells. The embryonic development of the avian ciliary muscle was studied in chick embryos from stage 25 HH to the time of hatching. Serial sections of the eye were stained routinely or immunocytochemically using the monoclonal antibody 13F4, which recognizes a cytoplasmic antigen specific for all types of muscle cells. We found that the mesenchymal immunoreactive cells, at stage 37 HH, are arranged in two distinct orientations forming the anterior and posterior portions of the ciliary muscle. At stages 38 and 39 HH the pigmented epithelium contained 13F4 positive cells, which detach from the epithelium and apparently migrate into stroma. These epithelial cells may differentiate into muscle cells. Within this same time period a progressive accumulation of myoblasts was detected between the pigmented epithelium and the ciliary muscle. Some myoblasts containing melanin were also observed. At stage 40 HH the internal portion of the ciliary muscle was visible. These findings indicate that the immunopositive epithelial cells participate in the formation of the internal portion of the muscle. We conclude that the ciliary muscle derives not only from the mesenchymal cells but also from the pigmented epithelium. PMID- 12127104 TI - Afterimages in fly motion vision. AB - Afterimage-like effects modulate the responses of fly wide-field motion-sensitive cells following adaptation to stationary or slowly moving patterns. The origin of these afterimages is unclear. They have been interpreted as either the result of adaptation in the early visual system or as a direct consequence of the correlation scheme of motion detection. Using a combination of intracellular recording and computer modelling, we find that afterimage-like effects cannot be satisfactorily explained by a simple version of the correlation model previously proposed by Egelhaaf and Borst (J. Opt. Soc. Am. A 6 (1) (1989) 116). We propose a modified variant of the correlation model featuring a short delay filter and temporal high-pass filtering prior to motion correlation. Our model gives superior predictions of afterimage-like effects induced by a range of stimuli. Our model also predicts changes in cells' image step responses following exposure to motion, suggesting that previous experimental evidence for the "shortening delay" theory of motion adaptation (Biol. Cybern. 54 (1986) 223; Visual Neurosci. 14 (4) (1997) 741) should be re-interpreted in terms of afterimage effects. PMID- 12127105 TI - Electroretinographic oscillatory potentials are reduced in adenylyl cyclase type I deficient mice. AB - Electroretinography (ERG) of adult Adcy1(brl) mutant mice, which are deficient in adenylyl cyclase type 1 (AC1) activity, revealed decreased amplitude of the oscillatory potentials (OP) and of the primary rising phase of the b-wave intensity-response function in scotopic conditions. These abnormalities were less discernable in 3-6 week old mutants. No abnormalities were detected in the ERG signal obtained in photopic conditions or in the dark adaptation dynamics. The mutants displayed no histologic evidence of retinal degeneration. Retinal output, as measured by visual evoked potentials, was not different from heterozygous control mice. AC1-dependent pathways contribute to the generation of the retinal response to light. They may be necessary for the maintenance of the neural generators of the ERG OP. PMID- 12127106 TI - The spatial properties of opponent-motion normalization. AB - The final stage of the Adelson-Bergen model [J. Opt. Soc. Am. A 2 (1985) 284] computes net motion as the difference between directionally opposite energies E(L) and E(R). However, Georgeson and Scott-Samuel [Vis. Res. 39 (1999) 4393] found that human direction discrimination is better described by motion contrast (C(m))--a metric where opponent energy (E(L)-E(R)) is divided by flicker energy (E(L)+E(R)). In the present paper, we used a lateral masking paradigm to investigate the spatial properties of flicker energy involved in the normalization of opponent energy. Observers discriminated between left and right motion while viewing a checkerboard in which half of the checks contained a drifting sinusoid and the other half contained flicker (i.e. a counterphasing sinusoid). The relative luminance contrasts of flicker and motion checks determined the checkerboard's overall motion contrast C(m). We obtained selectivity functions for opponent-motion normalization by measuring C(m) thresholds whilst varying the orientation, spatial frequency, or size of flicker checks. In all conditions, performance (percent correct) decayed lawfully as we decreased motion contrast, validating the C(m) metric for our stimuli. Thresholds decreased with check size and also improved as we increased either the orientation or spatial-frequency difference between motion and flicker checks. Our data are inconsistent with Heeger-type normalization models [Vis. Neurosci. 9 (1992) 181] in which excitatory inputs are normalized by a non-selective pooling of inhibitory inputs, but data are consistent with the implicit assumption in Georgeson and Scott-Samuel's model that flicker normalization is localized in orientation, scale, and space. However, our lateral masking paradigm leaves open the possibility that the spatial properties of flicker normalization would be different if opponent and flicker energies spatially overlapped. Further characterization of motion contrast will require models of the spatial, temporal, and joint space-time properties of mechanisms mediating opponent-motion and flicker normalization. PMID- 12127107 TI - Failure of direction identification for briefly presented second-order motion stimuli: evidence for weak direction selectivity of the mechanisms encoding motion. AB - We sought to investigate why the direction of second-order motion, unlike first order motion, cannot be identified when the stimulus exposure duration is brief (<200 ms). In a series of experiments observers identified both the orientation (vertical or horizontal) and the direction (left, right, down or up) of a drifting sinusoidal modulation (0.93 c/ degrees ) in either the luminance (first order) or the contrast (second order) of a two-dimensional noise carrier. All motion stimuli were equated for visibility, and the duration was varied using the method of constant stimuli. Performance was measured for second-order motion over a range of drift temporal frequencies (0.63-5.04 Hz) and for first-order motion stimuli composed of two, opposite drifting modulations in luminance of unequal modulation depth. Orientation-identification performance was nearly 100% correct for both first-order and second-order motion stimuli, even at the briefest stimulus duration tested (26.49 ms). Direction identification for first-order motion was also typically good with brief presentations, but was poor for second order motion when the exposure duration was < approximately 200 ms. Importantly increasing either the drift temporal frequency of second-order motion or the bidirectional nature of the first-order motion patterns produced comparable levels of performance for the two varieties of motion (i.e. the minimum duration required for reliable direction identification could be equated). As orientation identification performance for the first-order and second-order motion stimuli was comparably good and minimally affected by duration, the marked differences on the direction-identification task must be specific to mechanisms that encode drift direction, rather than spatial structure. We propose that second-order motion detectors are much less selective for stimulus direction than first-order motion sensors, and thus are more susceptible to the deleterious effects of limiting stimulus duration (which introduces spurious motion in the opposite direction, particularly at low drift rates). Alternative explanations based on the delayed propagation of second-order motion signals or the temporal characteristics of the underlying motion mechanisms are not supported by our findings. PMID- 12127108 TI - Task-dependent transfer of perceptual to memory representations during delayed spatial frequency discrimination. AB - Discrimination thresholds were obtained using a delayed spatial frequency discrimination task. In Experiment 1, we found that presentation of a mask 3 s before onset of a reference Gabor patch caused a selective, spatial frequency dependent interference in a subsequent discrimination task. However, a 10 s interval abolished this masking effect. In Experiment 2, the mask was associated with a second spatial frequency discrimination task so that a representation of the mask had to be coded into short-term perceptual memory. This experiment was performed to assess whether absence of masking in the 10 s condition of Experiment 1 might be due to decay of the mask information in the perceptual or the memory representational domain. The presence of this second discrimination task now caused similar interference effects on the primary discrimination task at both the 3 s and 10 s interstimulus intervals (ISI) conditions. Finally, to test the robustness of the masking effect, the nature of the secondary masking task was changed from a spatial frequency discrimination task to an orientation discrimination task in Experiment 3. The masking effect was now abolished in both the 3 and 10 s ISI conditions. Together, the results from these experiments are consistent with the idea of a two-level perceptual memory mechanism. The results also suggest that stimulus representations during a perceptual discrimination task are shared between the perceptual and memory representation domains in a task-dependent manner. PMID- 12127109 TI - Separating the effects of response nonlinearity and internal noise psychophysically. AB - A psychophysical method is proposed to separate the contrast dependence of internal response and its noise. The resulting contrast relationships represent a signature of the visual processing stage that limits the human observer's performance. The method was applied to contrast discrimination for sustained and transient Gabor patches with a 3 cycle/deg spatial carrier. For both stimulus types the predominant noise was found to be multiplicative with a power exponent of 0.76-0.85 and the source of this noise preceded by an accelerating signal transducer with a power of 2-2.7. These exponents combine to account for the classic compressive power of about 0.4 for the signal-to-noise ratio in contrast discrimination. The estimated transducer acceleration suggests that there is a direct computation of contrast energy in the visual cortex. PMID- 12127110 TI - Transsaccadic perception of translating objects: effects of landmark objects and visual field position. AB - Previously Gysen, De Graef, and Verfaillie [Vision Research 42 (2002) 379] showed that, with stimulus displays presenting one stationary and one translating object, sensitivity for intrasaccadic displacements was higher for translating than for stationary objects. In the present paper the importance of the relative encoding of the path of the translating object towards the stationary object is investigated. In three experiments we compared detection of intrasaccadic displacements of translating objects in relative motion (moving towards the landmark object) and translating objects moving in isolation. No 'facilitatory' effect of relative motion was found. However a visual field effect was present. Performance was always better for the translating object presented in the lower part in comparison to the upper part of the visual field. A fourth experiment investigated the sensitivity for intrasaccadic displacements of stationary and translating objects presented in the upper as well as in the lower visual field. A lower visual field advantage was observed. The superior performance for translating objects, as was found previously, was confirmed in the lower and upper visual field. PMID- 12127111 TI - Cause of kinematic differences during centrifugal and centripetal saccades. AB - Measurements of eye movements have shown that centrifugal movements (i.e. away from the primary position) have a lower maximum velocity and a longer duration than centripetal movements (i.e. toward the primary position) of the same size. In 1988 Pelisson proposed that these kinematic differences might be caused by differences in the neural command signals, oculomotor mechanics or a combination of the two. By using the result of muscle force measurements that were made in recent years (Orbit 1.8 Gaze mechanics simulation, Eidactics, San Francisco, 1999) we simulated the muscle forces during centrifugal and centripetal saccades. Based on these simulations we show that the cause of the kinematic differences between the centrifugal and centripetal saccades is the non-linear force-velocity relationship (i.e. muscle viscosity) of the muscles. PMID- 12127112 TI - Refractive error and monochromatic aberrations in Singaporean children. AB - Higher order optical aberrations were measured in 273 cyclopleged Singaporean school children using a Bausch and Lomb Zywave aberrometer, with 268 of these subjects also undergoing corneal topography measurements (Tomey TMS 2 system). Subjects with low myopia (> -3.00 to -0.50 D) showed slightly, but significantly, less positive levels of spherical aberration than other refractive error groups. Chinese subjects also showed significantly higher amounts of aberrations than Malay subjects, particularly for vertical coma, but also for horizontal coma and spherical aberration. Anterior corneal spherical aberration (calculated from topography) was significantly correlated with whole eye spherical aberration, but did not vary significantly with refractive error or racial background. Residual spherical aberration (i.e. of posterior cornea and crystalline lens) did vary significantly with refractive error and race. Our results do not provide any evidence for aberration-driven form-deprivation as a major mechanism of myopia development. PMID- 12127114 TI - Cancer of the esophagogastric junction: epidemiology and pathogenesis. PMID- 12127115 TI - Surgical treatment of esophageal adenocarcinoma: concepts in evolution. PMID- 12127116 TI - Minimally invasive surgical approaches to esophageal cancer. PMID- 12127117 TI - Multimodality therapy for gastroesophageal cancers. PMID- 12127118 TI - Clinical and histologic follow-up after antireflux surgery for Barrett's esophagus. AB - There are few prospective studies that document the histologic follow-up after antireflux surgery in patients with Barrett's esophagus, as defined by the recently standardized criteria. We report the clinical, endoscopic, and histologic results of patients with Barrett's esophagus followed postoperatively for at least 2 years. Diagnosis of Barrett's esophagus required preoperative endoscopic evidence of columnar-lined epithelium in the esophagus and a biopsy demonstrating specialized intestinal metaplasia, which stains positively with Alcian blue stain. Between April 1993 and November 1998, a total of 104 patients meeting these criteria underwent fundoplication (laparoscopic [n = 84] or open [n = 6] nissen, laparoscopic Toupet [n = 11], laparoscopic Collis-Nissen [n = 1], Collins-Toupet [n = 1] or open Dor [n = 1]). Short-segment Barrett's esophagus (length of intestinal metaplasia <3 cm) was found preoperatively in 34% and low grade dysplasia in 4% of patients. All patients were contacted yearly by mail, phone, or clinic visit. At a mean follow-up of 4.6 years (range 2 to 7.5 years), 81% of patients had stopped taking antisecretory medications and 97% were satisfied with the results of their operations. Eight patients have undergone reoperation for recurrence of symptoms. Two patients have died and two were excluded from endoscopic biopsy because of portal hypertension. Sixty-six patients complied with the surveillance protocol, and their histologic results were returned to our center. Symptomatic follow-up of the 34 patients who refused surveillance esophagogastro and duodenoscopy revealed two patients who were taking medication for reflux symptoms. None of the patients have developed high grade dysplasia or esophageal carcinoma during surveillance endoscopy (337 total patient-years of follow-up). The incidence of regression of intestinal metaplasia to cardiac-fundic-type metaplasia after successful antireflux surgery is greater than previously reported. We suspect that this is a result of longer follow-up and the inclusion of patients with short-segment Barrett's esophagus. A substantial number of patients with Barrett's esophagus who are asymptomatic after antireflux surgery refuse surveillance endoscopy. PMID- 12127119 TI - Long-term efficacy of total (Nissen-Rossetti) and posterior partial (Toupet) fundoplication: results of a randomized clinical trial. AB - The efficacy of fundoplication operations in the long-term management of gastroesophageal reflux disease (GERD) has been documented. However, only a few prospective controlled series support the long-term (>10 years) efficacy of these procedures, and further data are required to also determine whether the type of fundoplication affects the frequency of postfundoplication complaints. The aim of this study was to conduct a randomized, controlled clinical trial to assess the long-term symptomatic outcome of a partial posterior fundoplication as compared to a total fundic wrap. During the years 1983 to 1991, a total of 137 patients with chronic gastroesophageal reflux disease were enrolled in the study; 72 were randomized to semifundoplication (Toupet) and 65 to total fundoplication (Nissen Rossetti). A standardized symptom questionnaire was used for follow-up of these patients. A total of 110 patients completed a median follow-up of 11.5 years; 54 had a total wrap and 56 underwent a partial posterior fundoplication. During this period, seven patients required reoperation (Nissen-Rossetti in 5 and Toupet in 2), 11 patients died, and nine patients were lost to follow-up or did not comply with the follow-up program. Control of heartburn (no symptoms or mild, intermittent symptoms) was achieved in 88% and 92% in the total and partial fundoplication groups, respectively, and the corresponding figures for control of acid regurgitation were 90% and 94%. We observed no difference in dysphagia scoring between the two groups, although odynophagia was somewhat more frequently reported in those undergoing a total fundoplication. On the other hand, a significant difference was observed in the prevalence of rectal flatus and postprandial fullness, which were recorded significantly more often in those undergoing a total fundoplication (P < 0.001 and P < 0.03, respectively). Posterior partial fundoplication seems to maintain the same high level of reflux control as total fundoplication. Earlier observations demonstrating the advantages of a partial fundoplication, which included fewer complaints associated with gas-bloat, continue to be valid after more than 10 years of follow-up. PMID- 12127121 TI - An integrated approach to intestinal failure: results of a new program with total parenteral nutrition, bowel rehabilitation, and transplantation. AB - Intestinal failure can be treated with bowel rehabilitation, total parenteral nutrition, or intestinal transplantation. Little has been done to integrate these therapies for patients with intestinal insufficiency or failure and to develop an algorithm for appropriate use and timing. We established a multidisciplinary program using bowel rehabilitation, total parenteral nutrition, or intestinal transplantation as appropriate in a large population. Evaluation included clinical, pathologic, and psychosocial assessments and assignment to therapy based on the results of this evaluation. Of 59 patients evaluated for life threatening complications of intestinal failure, 68% were considered appropriate candidates for transplantation, 10% were managed with rehabilitation, and 17% were maintained on optimized long-term parenteral nutrition. Nineteen transplants were performed, with 78% patient survival and 66% graft survival. Patient survival among isolated intestine recipients was 90%. All patients managed with rehabilitation were weaned from parenteral nutrition within 6 months. Long-term management with parenteral nutrition resulted in a significant number of deaths both among patients waiting for a transplant and those who were poor candidates for transplant. Intestinal rehabilitation, when successful, is optimal. For patients with irreversible intestinal failure, isolated intestinal transplantation holds particular promise. Parenteral nutrition is plagued by high failure rates among this population of debilitated patients compared with the general parenteral nutrition population. Integration of these therapies, with individualization of care based on a multidisciplinary approach and perhaps with earlier isolated intestinal transplantation for patients with irreversible intestinal failure, should optimize survival. PMID- 12127120 TI - Ligand activation of alternatively spliced fibroblast growth factor receptor-1 modulates pancreatic adenocarcinoma cell malignancy. AB - Pancreatic adenocarcinoma continues to be a devastating tumor (28,000 new cases per year in the United States; 10% 2-year survival). Pancreatic adenocarcinoma frequently (90% of the time) overexpresses fibroblast growth factor ligands (FGF 1 and FGF-2) and alternatively spliced high-affinity receptors (FGFR-1beta) (FGFR 1alpha was previously found in normal pancreatic tissue). To study the significance of this observation in vitro, PANC-1 cells were stably transfected via the pMEXneo vector containing FGFR-1alpha (PANC-1alpha) or FGFR-1beta (PANC 1beta) isoforms. Cells were treated with 1 mg/ml of 5-fluorouracil. Cells were evaluated for growth inhibition, apoptosis (propidium iodide staining and flow cytometry, caspase 3 activation) and for Bcl-x(L)/BAX expression (by Western blot analysis). In vivo, 7 x 10(6) cells of each isoform were injected into nude Balb/c mice for xenograft formation (N = 10). Compared to PANC-1beta (9%) in vitro, 5-fluorouracil-induced death was significantly (P < 0.05) increased in PANC-1alpha (20%) at 24 hours. Increased cell death in PANC-1alpha was mediated by activated caspase 3 and was correlated with decreased expression of Bcl x(L)/BAX. In vivo, PANC-1beta readily demonstrated formation of tumor xenograft at 2 weeks, whereas PANC-1alpha did not form tumors. Alternative splicing of FGFR 1 to the beta isoform appears to correlate with pancreatic adenocarcinoma cell growth in vivo and resistance to chemotherapy. Inhibition of FGFR-1 splicing or overexpression of FGFR-1alpha inhibits pancreatic adenocarcinoma cell growth in vivo and restores cytotoxic responses to chemotherapy, thereby suggesting the basis of rational interventional strategies for this devastating tumor. PMID- 12127122 TI - Reduction of intestinal neoplasia with adenomatous polyposis coli gene replacement and COX-2 inhibition is additive. AB - Mutations of the adenomatous polyposis coli (APC) gene are implicated early in colorectal tumorigenesis. Restoration of normal APC expression through gene therapy may prevent or reduce intestinal neoplasia. Furthermore, the relationship between colorectal tumors and increased cyclooxygenase-2 (COX-2) activity provides a rationale for the use of selective COX-2 inhibitors such as rofecoxib (Vioxx) to prevent the formation of polyps. This study was performed to determine the effects of liposome-mediated APC gene therapy and a selective COX-2 inhibitor on intestinal neoplasia in vivo. Five-week-old Min mice weaned on a 30% high-fat diet were randomized to receive no treatment (control), APC only, Vioxx only, and APC/Vioxx. APC-treated mice received a plasmid containing the human APC cDNA (pCMV-APC) mixed with a liposome preparation that was administered biweekly. Vioxx was administered at 200 ppm in the high-fat rodent chow. The control mice were treated similarly with a plasmid construct lacking the APC gene. Confirmation of exogenous APC gene expression was determined by Western blot analysis. After 2 months, there was a 54% and 70% reduction in the total number of intestinal polyps after APC and Vioxx treatment, respectively. Combined APC/Vioxx therapy reduced polyp formation by 87%. The reduction of intestinal neoplasia by APC gene replacement and COX-2 inhibition suggests their separate roles in intestinal tumorigenesis. Each modality, both individually and together, may prove therapeutic and therefore contribute to new strategies in the prevention and treatment of colorectal cancer. PMID- 12127123 TI - Use of a bipolar vessel-sealing device for parenchymal transection during liver surgery. AB - Most blood loss during liver resection occurs during parenchymal transection, and multiple approaches have been developed to limit blood loss. The purpose of this study was to evaluate a new bipolar vessel-sealing device in hepatic surgery, particularly whether the device would permit safe transection without routine inflow occlusion. Twenty-seven hepatic procedures were performed using the device after preliminary studies to adapt its use to the liver. Inflow occlusion was used when necessary to control blood loss but not as a routine. The device worked well for transection through normal liver during common liver operations such as right hepatectomy. It worked less well for enucleations. Inflow occlusion was used in approximately 10% of resections, exclusive of enucleations, and about 25% of these patients were transfused during surgery or in the postoperative period. We conclude that the device is a useful tool in standard liver resections. PMID- 12127124 TI - Effect of intraoperative cholangiography during cholecystectomy on outcome after gallstone pancreatitis. AB - Acute gallstone pancreatitis has traditionally been managed by early cholecystectomy with intraoperative cholangiography (IOC). To evaluate the effect of IOC on patient outcome, we analyzed all patients operated on for acute gallstone pancreatitis at our institution over a 3-year period. A total of 200 patients (37 open, 163 laparoscopic) were evaluated. Nineteen of 34 patients who underwent preoperative endoscopic retrograde cholangiopancreatography (ERCP) were found to have common bile duct (CBD) stones. The 59 patients who underwent cholecystectomy with IOC had significantly longer operative times compared to the 141 patients who underwent cholecystectomy alone (167 vs. 105 minutes for open [P = 0.008] and 89 vs. 68 minutes for laparoscopic [P < 0.0001] operations). Of the 59 patients who underwent IOC, only nine (15%) had abnormal cholangiograms, and CBD exploration in seven revealed stones in four patients, edematous ampullae in two, and no abnormality in one. Six of eight patients (5 IOC, 3 no IOC) who required immediate postoperative ERCP were noted to have CBD stones. Patients who underwent IOC had significantly longer postoperative hospital stays (3.8 vs. 2.0 days [P = 0.007]). The incidence of retained CBD stones following surgery was similar (5.1% IOC, 2.8% no IOC). Although 7 of 122 patients who underwent laparoscopic cholecystectomy without IOC were readmitted, only one was found on ERCP to have a retained CBD stone. Age, sex, preoperative days, procedure type, and biliary-pancreatic complications after discharge did not differ significantly between patients with and without IOC. We conclude that IOC in patients operated on for acute gallstone pancreatitis results in a longer operative time and a prolonged postoperative course, but has no effect on the incidence of retained CBD stones. PMID- 12127125 TI - Systemic response in patients undergoing laparoscopic cholecystectomy using gasless or carbon dioxide pneumoperitoneum: a randomized study. AB - In general, laparoscopic cholecystectomy produces a surgical stress response very similar to which occurs after open cholecystectomy. The question is whether the pneumoperitoneum constitutes a significant pathophysiologic trauma, which might be followed by profound changes in the stress response. We conducted a prospective, randomized trial involving 50 consecutive patients scheduled for laparoscopic cholecystectomy, who had a body mass index equal to or less than 30 kg/m(2) with no acute cholecystitis, pancreatitis, or liver or renal disease. These patients were randomized to undergo either the gasless (GLC, n = 24) or the carbon dioxide pneumoperitoneum (CLC, n = 26) procedure. Perioperative assessment of cortisol, insulin, glucose, and C-reactive protein levels was the main determinant of outcome. During the operative procedure, significantly higher levels of serum cortisol and insulin were found in the CLC group than in the GLC group (P < 0.05). No difference in glucose levels was observed between the two groups. The inflammatory response was moderate in both groups. However, on postoperative day 1 the median C-reactive protein level was significantly higher in the GLC group than that in the CLC group (P < 0.05). Carbon dioxide and the positive intra-abdominal pressure during conventional laparoscopy may contribute to the activation of the surgical stress response. PMID- 12127126 TI - Detecting blunt pancreatic injuries. AB - Pancreatic injury after blunt abdominal trauma is exceedingly uncommon, occurring in less than 5% of major abdominal injuries. When blunt pancreatic injury does occur, however, it is notoriously difficult to identify. The use of serum amylase has been advocated in the diagnosis of such injury, yet it is neither sensitive nor specific. Computed tomography has become widely accepted in the evaluation of hemodynamically stable patients after blunt abdominal trauma, although it is clearly not a sensitive modality for the detection of pancreatic injury. In fact, numerous examples of normal CT scans with missed pancreatic injury have been documented. However, careful attention to CT technique and awareness of the CT manifestations of pancreatic injury may facilitate the diagnosis of pancreatic injury. Additionally, important information about the pancreatic duct can be obtained with the use of MRI-pancreatography or endoscopic retrograde pancreatography. Accurate, timely identification of major pancreatic ductal injury is imperative because delay in diagnosis and associated vascular injuries are largely responsible for the high morbidity and mortality associated with blunt pancreatic trauma. Blunt pancreatic trauma can be managed successfully by means of both operative and selective approaches. PMID- 12127128 TI - Factors affecting surgical risk in elderly patients with inflammatory bowel disease. AB - The operative treatment of elderly patients with inflammatory bowel disease (IBD) has often been avoided in favor of medical management because of a perceived increase in surgical risk. This study sought to define the following in the elderly IBD patient population: (1) the risk of surgical management and (2) those factors affecting risk. Thirty patients with IBD, aged 60 years or more, who were surgically managed by a single surgeon over a 10-year period, were retrospectively matched to 75 patients with IBD who were less than 60 years of age; patients were matched according to sex, date of surgery, and type of surgery performed. Regression analysis using generalized estimating equation methodology to account for the matched clusters of patients was performed to evaluate the effect of age group on the complication rate, operating room time, and length of hospital stay. Presence of comorbid conditions, surgical indications, prior surgery for IBD, and the use of immunosuppressive medications were studied in multivariate models, adjusting for age group. By means of univariate analysis, the odds of complications in elderly IBD patients were shown to be statistically higher than the odds seen in younger patients (47% vs. 20%, P = 0.01). Also observed in the elderly group were a longer length of hospital stay (11.5 days vs. 7.1 days, P = 0.001) and longer operating room time (249 minutes vs. 212 minutes, P = 0.02). Multivariate analysis revealed that the effect of age remained statistically significant, even when adjusted for potential confounding variables such as comorbidity, medications, date of diagnosis of IBD, and indications for surgery. The complication outcome was significantly associated with the surgical indication, with obstruction, fistula, and bleeding having increased odds of complications as compared with other indications (odds ratio = 1.7 vs. 4.2 vs. 7.2, respectively, P = 0.02). The length of hospital stay similarly was significantly associated with the surgical indication (fistula, 10.5 days vs. bleeding, 9.8 days vs. obstruction, 7.4 days vs. other, 9.3 days; P = 0.04) and a history of prior surgery. A significant interaction for length of hospital stay was present between age group and prior surgery status (with prior surgery: old, 18 days vs. young, 6.4 days, P = 0.0001; without prior surgery: old, 9.5 days vs. young 7.3 days, P = 0.10). Elderly patients with IBD have an increased rate of postoperative complications along with an increased length of hospital stay and increased operating room time. This effect of age persists when adjusted for comorbidity and immunosuppressive therapy. Complications are most dependent on surgical indications, with obstruction being the least and bleeding the worst predictive factors. The longest hospital stay is associated with patients who require surgery for fistulous disease and patients who have undergone previous surgery. The fact that the higher complication rate seen in older patients with IBD is associated with disease-defined surgical indications suggests that IBD in elderly patients may be more aggressive than what is observed in younger individuals. PMID- 12127127 TI - Inducible nitric oxide synthase knockout mice are resistant to diet-induced loss of gut barrier function and intestinal injury. AB - Loss of gut barrier function has been documented to occur in animals receiving total parenteral nutrition (TPN) and certain liquid diets. However, the mechanisms responsible for diet-induced gut barrier dysfunction remain to be fully determined. Thus we tested the hypothesis that increased intestinal nitric oxide production contributes to this phenomenon. To test this hypothesis, iNOS deficient (iNOS -/-) mice and their wild-type littermates (iNOS +/+) were fed either chow or TPN solution for 14 days. Subsequently they were killed and gut barrier function was assessed by measuring bacterial translocation to the mesenteric lymph node (MLN) complex. Additionally, intestinal bacterial population levels, gut morphology, plasma and intestinal nitric oxide levels, as well as intestinal levels of the nitric oxide synthase (NOS) enzymes cNOS and iNOS, were measured. Bacterial translocation occurred in the iNOS +/+ but not the iNOS -/- mice receiving oral TPN solution. Oral TPN-induced bacterial translocation was associated with increased intestinal bacterial population levels as well as morphologic evidence of intestinal injury. Plasma and intestinal levels of the nitric oxide products, nitrite/nitrate, were increased in the iNOS +/+ mice fed the TPN solution but not in the chow-fed groups or the iNOS -/- mice receiving TPN solution. Last, intestinal iNOS, but not cNOS, activity was increased in the iNOS +/+ oral TPN-fed mice. These results implicate a role for increased intestinal nitric oxide production, through iNOS, in the pathogenesis of oral TPN-induced gut barrier dysfunction and injury. PMID- 12127129 TI - Breast cancer masquerading as a primary gastric carcinoma. AB - The differentiation between primary gastric cancer and a metastatic breast tumor to the stomach is important for planning of treatment and to spare the patient unnecessary surgery. We report a rare case of breast cancer with metastasis to the stomach. The diagnosis was established by histologic and immunohistochemical analyses of biopsies of the stomach lesion using gross cystic disease fluid protein-15, cytokeratin, carcinoembryonic antigen, and epithelial membrane antigen. Positivity for gross cystic disease fluid protein-15 with negative staining for carcinoembryonic antigen supported the diagnosis of a breast cancer metastasis. The patient was treated with systemic chemotherapy without surgery and is still alive 2 years after initial referral for a gastric neoplasm. PMID- 12127131 TI - Ruminations of an ordinary hepatic surgeon: a journey through the pitfalls of major liver resections. AB - Historically, major hepatic resections have been fraught with voluminous blood losses and, at times, high mortality rates. Improvements in patient selection and operative technique over the past 20 years have resulted in marked reduction in death and complications and have given the impression that liver surgery can be relatively effortless. Contrary to this belief, the present review illustrates some of the pitfalls and dangers of major hepatectomy and may serve to alert ordinary surgeons to approach this operation with a degree of trepidation and careful planning. Over a 22-year period, 147 liver resections were performed by one surgeon for solid liver tumors (range 0 to 21/yr). Of these, 101 were major hepatectomies comprising at least three anatomic segments (63 right, 24 left, 11 extended right, and 3 extended left) and form the basis for this report. The major resections were performed for benign disease in 16 patients and malignant tumors in 85 (24 primary and 61 metastatic lesions). All but one patient were noncirrhotic. Seventeen patients were more than 70 years and 84 were less than 70 years of age. There were five postoperative deaths among these 101 patients: two intraoperative (coagulopathy after venovenous bypass in 1 and air embolus in 1), two from postoperative liver failure, and one resulting from a myocardial infarction. Three deaths were in patients older than 70 (18%), and two were in patients younger than 70 (2%) (P = 0.03). Complications developed in 20 of 96 survivors, three patients required reoperation for postoperative bleeding, and nine patients had some duration of bile leakage. In contrast, among those undergoing "minor" hepatectomies (n = 46), there were no deaths and six (13%) patients had complications. In patients undergoing major hepatectomies, estimated blood loss was 3836 +/- 3346 ml. Estimated blood loss was unaffected by experience (first 50 patients vs. second 51 patients) or use of the ultrasonic surgical aspirator, but has been reduced by the use of the Harmonic scalpel (2650 +/- 2706.1 ml vs. 3997 +/- 3405.8 ml, P = 0.026). The use of rapid-infusion systems aided in preventing intraoperative hypotension and hypothermia. Estimated blood loss was significantly greater than with minor anterior or lateral segmentectomies (n = 24) (3836 +/- 3346 ml vs. 975 +/- 518.8 ml, P < 0.0001). Hospital length of stay has been shortened, primarily by the use of closed suction drainage compared to open drainage (7.5 2 +/-.4 days vs. 18.8 +/- 8.4 days, P < 0.0001). Major hepatectomies continue to be formidable operations with the potential for copious blood loss and intraoperative instability. Proper patient selection, anesthesia support and availability of rapid-infusion technology, and familiarity with liver anatomy are important in keeping operative mortality and postoperative morbidity at an acceptable level. PMID- 12127130 TI - Surgical treatment of hilar bile duct carcinoma: experience with 25 consecutive hepatectomies. AB - To evaluate our recent surgical policy regarding hilar bile duct carcinoma, we evaluated 62 cases treated between 1976 and 1993, and 25 cases treated between 1994 and 2000. In the late period we used percutaneous transhepatic portal vein embolization (PTPE) before extended right hepatectomy; S4a + S5 + S1 hepatectomy for elderly patients and those with poor liver function; and routine total caudate lobectomy including the paracaval portion and resection of the inferior portion of the medial segment (S4a). Sixty-five (74.7%) of the 87 patients underwent hepatectomy: 40 in the early period and 25 in the late period. Bile duct resection alone was performed in 22 patients, all in the early period. Resection was curative in 54.8% in the early period and 88.0% in the late period. The 3- and 5-year survival rates in the early period were 27.1% and 20.2%, respectively, as compared to 59.9% and 49.9% in the late period. Analysis of the 25 hepatectomies in the late period revealed improved survival times compared to patients treated by PTPE with extended right hepatectomy. No complications occurred after extended left hepatectomy or S4a + S5 + S1 hepatectomy, but four patients (16%) who underwent extended right hepatectomy plus PTPE died postoperatively. Our policy has resulted in improved outcome in patients with hilar bile duct carcinoma. PMID- 12127132 TI - An unusual case of squamous cell carcinoma arising at the stomal site: case report and review of the literature. AB - An unusual case of squamous cell carcinoma arising at the ileocutaneous stomal site is reported. The presenting symptoms were peristomal ulceration and bleeding. The patient was treated with wide local excision of the stoma and the peristomal skin, and relocation of the ileostomy. A search of the literature for other similar cases subsequently identified two additional cases that were reported in the literature in 1987 and 2000. PMID- 12127133 TI - Influence of functional bowel disease on outcome of surgical antireflux procedures. AB - Patients with gastroesophageal reflux disease (GERD) have a coexisting diagnosis of functional bowel disease (FBD) in approximately 30% of cases. Symptom improvement after surgical therapy for GERD may be less in patients with FBD when compared to patients without this coexisting problem. A retrospective review of patients undergoing Nissen fundoplication between 1996 and 2000 evaluated patients with documented FBD or FBD symptoms to determine operative outcome. Poor postoperative outcome included recurrent heartburn, gas bloat syndrome, dysphagia requiring reoperation or dilation, or delay in resumption of normal diet. Bivariate comparison and multivariate logistic regression evaluated the independent impact of a documented diagnosis of FBD or preoperative symptoms of FBD on outcome. This study examined 155 patients: 32% reported having symptoms of FBD and 10% had a confirmed diagnosis of FBD. Poor postoperative outcomes occurred in 27%. Patients with a documented diagnosis of FBD were significantly more likely to have a poor outcome when compared to patients without symptoms of FBD (53% vs. 23%, P = 0.01). Patients with preoperative symptoms of FBD (but without a documented diagnosis of FBD) also had a higher incidence of poor outcome (5% vs. 23%, P = 0.09). Patients with FBD are at increased risk of poor results after antireflux surgery. Patients with these conditions should be counseled preoperatively regarding the potential for recurrent postoperative symptoms. PMID- 12127134 TI - Effective treatment of rumination with Nissen fundoplication. AB - Rumination is a syndrome characterized by the effortless regurgitation of recently ingested food. It has been linked to severe medical and psychosocial conditions including malnutrition, aspiration pneumonia, and complete social withdrawal. Psychotherapy, the current treatment modality for rumination, may improve symptoms but requires significant motivation and is rarely curative. We hypothesized that a complete fundoplication would eliminate, or at least impair, the ability to regurgitate gastric contents through the esophagogastric junction. We performed a Nissen fundoplication in five patients with a classic history of rumination. In all cases, symptoms had been resistant to medical and psychiatric intervention prior to fundoplication. Formal preoperative testing included esophageal manometry, 24-hour pH monitoring, endoscopy, and upper gastrointestinal barium swallow studies. All patients reported their primary symptom to be effortless recurrent postprandial regurgitation for 1 to 2 hours after meals consistent with rumination. Four (80%) of the five patients had low resting lower esophageal sphincter pressures with evidence of gastroesophageal reflux disease on 24-hour pH monitoring. All patients reported complete cessation of ruminating behavior after Nissen fundoplication. We report, for the first time, complete elimination of rumination symptoms after a Nissen fundoplication. Although further trials are needed to confirm our results, we recommend considering a Nissen fundoplication for treatment of rumination refractory to behavioral and medical interventions. PMID- 12127137 TI - Introduction: membrane trafficking, secretion and development. PMID- 12127135 TI - Effect of duodenal diversion on low-grade dysplasia in patients with Barrett's esophagus: analysis of 37 patients. AB - It is well known that in patients with Barrett's esophagus (BE), even after antireflux surgery, intestinal metaplasia can progress to dysplasia or even adenocarcinoma. However, the opposite-that is regression of dysplastic changes to intestinal metaplasia after antireflux surgery-has been documented in only a few reports. The objective of this study was to determine the effect of a duodenal diversion operation on low-grade dysplasia in patients with BE. Thirty-seven patients with either short-segment (n = 12) or long-segment (n = 25) BE underwent antireflux surgery plus either a duodenal switch procedure (13 patients) or a partial distal gastrectomy with Roux-en-Y gastrojejunal anastomosis (24 patients). All of them were subjected to complete clinical, endoscopic, histologic, manometric, and 24-hour pH testing, and 24-hour monitoring of the bile exposure in distal esophagus. There were no deaths in this series, and morbidity occurred in only one patient (2.7%). Manometric assessment after surgery showed a significant increase in sphincter pressure, abdominal length, and total length (P < 0.001). Acid reflux showed a significant decrease after surgery, and duodenal reflux was completely abolished in all except one patient. Follow-up in all patients was longer than 24 months (mean 60 months). Three to four endoscopic procedures were performed after surgery in each patient, and several biopsy specimens were taken distal to the squamo-columnar junction during each endoscopic procedure. Eleven patients (91%) with short-segment BE demonstrated histologic regression to either cardiac mucosa or nondysplastic intestinal metaplasia. Among the 25 patients with long-segment BE, there was a 62.5% rate of histologic regression to nondysplastic epithelium when the length of BE measured between 31 and 99 mm and 33% histologic regression when the length of BE was 101 mm or more. There were no cases of progression to high-grade dysplasia or adenocarcinoma. The endoscopic length of the columnar-lined esophagus did not change late after surgery. In 65% of patients with BE, antireflux surgery, gastric acid reduction, and duodenal diversion produced histologic regression of low-grade dysplasia to nondysplastic mucosa. This effect was even more pronounced when the length of BE was shorter. It seems to be permanent, and no progression to high-grade dysplasia or adenocarcinoma has occurred. PMID- 12127138 TI - Exocytosis, endocytosis, and development. AB - During development, metazoans are faced with the daunting task of generating many different cell types in a temporally and spatially precise manner. This orderly process of cell generation relies on creating localized signals that activate or inhibit specific cellular pathways. Recent work has shown that some of these localized signals require the targeted secretion of proteins, or their uptake by endocytosis. The importance of these protein trafficking pathways in localized signal generation is further substantiated by endo- and exocytosis mutants which can phenocopy many developmental mutants. Genetic and molecular techniques that increase our ability to inhibit exocytosis and endocytosis in a temporal and cell type specific manner are likely to further elucidate the complexities of development. PMID- 12127139 TI - Membrane trafficking in cytokinesis. AB - Until recently, two distinct types of cytokinesis were thought to be responsible for the division of plant and animal cells. Plant cells divide through the formation of a membrane plate between the daughter cells, while animal cells divide by the constriction of a cortical actin-based ring around the cell. However, accumulating evidence suggests that the two mechanisms may have more in common than previously thought. In this review we will focus on recent developments that raise the possibility of unexpected similarities between the final steps in cytokinesis in animal and plant cells. PMID- 12127140 TI - Regulating morphogen gradients in the Drosophila wing. AB - During development, diffusible ligands, known as morphogens, are thought to move across fields of cells, regulating gene expression in a concentration dependent manner. The case for morphogens has been convincingly made for the Decapentapleigic (Dpp), Wingless (Wg) and Hedgehog (Hh) proteins in the Drosophila wing. In each case, the concentration of the morphogen's receptor plays an important role in shaping the morphogen gradient, through influencing ligand transport and/or stability. However, the relationships between each ligand/receptor pair are different. The role of heparan sulfated proteoglycans, endocytosis and novel exovesicles called argosomes in regulating morphogen distribution will also be discussed. PMID- 12127141 TI - Membrane trafficking in Drosophila wing and eye development. AB - It is clear that membrane transport is essential to the proper sorting and delivery of membrane bound receptors and ligands, and secreted signaling molecules. Molecular genetic studies in Drosophila are particularly well suited to studies of membrane transport in development. The conservation of cell signaling pathways and membrane transport molecules between Drosophila and other species makes the results obtained in these studies of general interest. In addition, the ability to generate gain- and loss-of-function genetic mutations of various strengths, and the ability to generate transgenic flies that direct protein expression to tissues during development are of particular advantage. Several recent papers suggest that interesting and novel roles for membrane transport processes will be uncovered by studying classically defined membrane transport proteins in developmental contexts. Together these studies suggest that regulation of membrane transport may represent an additional mechanism to regulate the strength of cell-cell signaling during development. PMID- 12127142 TI - Expression of SNAP-25 during mammalian retinal development: thinking outside the synapse. AB - The SNARE complex is the core machinery required for vesicle fusion events. Numerous structural, functional, and genetic studies have led to a better understanding of mechanisms that regulate vesicle fusion events during neural development. Studies using the mammalian retina as a model system have increased our understanding of the dynamic patterns of expression of SNARE proteins. In particular, the SNARE complex protein SNAP-25 is expressed in a dynamic fashion during the development of cholinergic amacrine cells in a number of mammalian species. SNAP-25 is also likely to play a crucial role during the development of vertebrate photoreceptors. The integration of comparative studies examining SNARE proteins, such as SNAP-25, provides a powerful approach for the study of CNS development. PMID- 12127144 TI - Say when: reversible control of gene expression in the mouse by lac. AB - In bacteria, coordinate expression of genes involved in lactose metabolism is regulated by the lac repressor and its DNA binding sequence, the lac operator. The lac operator-repressor complex can also be used to regulate gene expression in the laboratory mouse. In this review, I discuss the current state of murine trans-operons, and suggest ways this lac-based system might be used to build more advanced models of human diseases in the mouse. PMID- 12127145 TI - Tetracycline-controlled transcriptional regulation systems: advances and application in transgenic animal modeling. AB - Since the first tetracycline-controlled transcriptional activation system was designed nearly a decade ago, new variants, modifications, and improvements have been steadily added to this powerful set of tools for temporal control of transgene expression in mammalian systems. Tetracycline-based externally regulatable (Tet-based) systems have been successfully used to control the expression of numerous transgenes in cultured cells and in whole organisms, especially in mice. The application of these systems has provided invaluable insights into the function and regulation of a variety of genes under physiological and pathological conditions. Because of the favorable characteristics of the inducing agent doxycycline and the efficiency and effectiveness of the operating mechanism, the Tet-based systems have attracted substantial attention from the transgenic research community and are rapidly gaining popularity. The original tetracycline-controlled transcriptional activator (tTA) is a regulator with tight control of target gene expression and a broad range of inducibility. The reverse tetracycline-controlled transcriptional activator (rtTA) activates the responsive elements only in the presence of doxycycline, giving a convenient control over the target transgene. The recently developed tetracycline-controlled transcriptional silencer (tTS) has been successfully used in cultured cells and in transgenic mice. In combination with rtTA, tTS actively suppresses background expression or "leakiness" without impeding the inducibility of the target gene, providing a true "On/Off" transgenic switch. New variants of Tet-based regulators with improved features are still emerging and the utilities of these systems are constantly being tested. PMID- 12127147 TI - Chemical chaperones reduce aggregate formation and cell death caused by the truncated Machado-Joseph disease gene product with an expanded polyglutamine stretch. AB - Machado-Joseph disease/spinocerebellar ataxia-3 (MJD/SCA-3) is an inherited neurodegenerative disorder caused by expansion of the polyglutamine stretch in the MJD gene-encoded protein ataxin-3. The truncated form of mutated ataxin-3 causes aggregation and cell death in vitro and in vivo. Abnormal conformation and misfolding of the pathological protein are assumed critical to pathogenesis. To test this hypothesis, we transfected BHK-21 and Neuro2a cells transiently with N terminal truncated ataxin-3 with an expanded polyglutamine stretch. We then studied the effects of organic solvent dimethyl sulfoxide (DMSO), cellular osmolytes glycerol, and trimethylamine N-oxide (TMAO) on aggregate formation and cell death. These reagents stabilize proteins in their native conformation and are called chemical chaperones based on their influence on protein folding. Aggregate formation and cytotoxicity induced by truncated expanded ataxin-3 were reduced by exposing cells to these chemical chaperones. Our results indicate the potentially useful therapeutic strategy of the chemical chaperones in preventing cell death in MJD. PMID- 12127146 TI - Expression and differential processing of caspases 6 and 7 in relation to specific epileptiform EEG patterns following limbic seizures. AB - The caspase family of cell death proteases has been implicated in the mechanism of neuronal death following seizures. We investigated the expression and processing of caspases 6 and 7, putative executioner caspases. Brief limbic seizures were evoked by intraamygdala kainic acid to elicit unilateral death of target hippocampal CA3 neurons in the rat. Seizures rapidly induced cleavage of constitutively expressed caspase-6, followed by elevated VEIDase activity and the proteolysis of lamin A. Neuronal caspase-6 immunoreactivity was markedly upregulated within cortex and hippocampus in relation to bursts of polyspike paroxysmal discharges. In contrast, while caspase-7 expression also increased within cortical and hippocampal neuronal populations in response to the same seizure patterns, caspase-7 was not proteolytically activated. These data highlight differences in expression and activation of caspases 6 and 7 in response to identifiable seizure patterns, focusing potential therapeutic targets for neuroprotection in epilepsy. PMID- 12127148 TI - Mitochondrial sequestration and Ca(2+)-dependent release of cytosolic Zn(2+) loads in cortical neurons. AB - The endogenous divalent cations, Ca(2+) and Zn(2+), are both highly toxic upon excessive glutamate triggered intracellular accumulation. Given apparent parallels in their neurotoxic mechanisms, the present study aimed to explore interactions between these cations, by examining effects of moderate intracellular Zn(2+) loading on responses to subsequent Ca(2+) influx. Cortical cultures were briefly exposed to high-K(+) buffer in the presence or absence of Zn(2+) (50-100 microM), to activate and permit a modestly toxic amount of Zn(2+) to enter through VSCC. After 1 h, the cultures were loaded with fluorescent probes, and 2 h after the Zn(2+) exposure, imaged before and after induction of Ca(2+) entry or addition of other drugs. In Zn(2+) preexposed cultures loaded with the Zn(2+) probe, Newport Green, induction of Ca(2+) entry through either VSCC or NMDA channels induced cytoplasmic release of sequestered Zn(2+). The source of this Ca(2+) dependent intracellular Zn(2+) release appears largely to be mitochondria, as indicated by the ability of the mitochondrial protonophore, FCCP, the mitochondrial uncoupler, dinitrophenol with the K(+) ionophore, valinomycin, or the inducer of mitochondrial permeability transition (mPT), phenylarsine oxide (PAO), to substitute for NMDA in triggering Zn(2+) release. Suggesting functional consequences of mitochondrial Zn(2+) uptake, Zn(2+) preexposures resulted in long-lasting mitochondrial depolarization (assessed with rhodamine 123), and reduced mitochondrial reactive oxygen species generation (assessed with hydroethidine) in response to subsequent NMDA triggered Ca(2+) influx. PMID- 12127149 TI - Aggresome formation in neuropathy models based on peripheral myelin protein 22 mutations. AB - Alterations in peripheral myelin protein 22 (PMP22) gene expression are associated with demyelinating peripheral neuropathies. Overexpression of wild type (wt) PMP22 or inhibition of proteasomal degradation lead to the formation of aggresomes, intracellular ubiquitinated PMP22 aggregates. Aggresome formation has now been observed with two mutant PMP22s, the Tr- and TrJ-PMP22 when the proteasome is inhibited. The formation of these aggresomes required intact microtubules and involved the recruitment of chaperones, including Hsp40, Hsp70, and alphaB-crystallin. Spontaneously formed ubiquitinated PMP22 aggregates were also observed in Schwann cells of homozygous TrJ mice. Significant upregulation of both the ubiquitin-proteasomal and lysosomal pathways occurred in affected nerves suggesting that two pathways of PMP22 degradation are present. Thus, the presence of aggresomes appears to be a common finding in neuropathy models of PMP22 overexpression and of some point mutations known to cause neuropathy in mice and humans. PMID- 12127151 TI - Fibrillar inclusions and motor neuron degeneration in transgenic mice expressing superoxide dismutase 1 with a disrupted copper-binding site. AB - Mutations in Cu/Zn superoxide dismutase 1 (SOD1) have been linked to dominantly inherited forms of amyotrophic lateral sclerosis (FALS). To test the hypothesis that the toxicity of mutant SOD1 originates in Cu(2+)-mediated formation of toxic radicals, we generated transgenic mice that express human SOD1 that encodes disease-linked mutations at two of the four histidine residues that are crucial for the coordinated binding of copper (H46R/H48Q). We demonstrate that mice expressing this mutant, which possesses little or no superoxide scavenging activity, develop motor neuron disease. Hence, mutations in SOD1 that disrupt the copper-binding site do not eliminate toxicity. We note that the pathology of the H46R/H48Q mice is dominated by fibrillar (Thioflavin-S-positive) inclusions and that similar inclusions were evident in mouse models that express the G37R, G85R, and G93A variants of human SOD1. Overall, our data are consistent with the hypothesis that the aberrant folding/aggregation of mutant SOD1 is a prominent feature in the pathogenesis of motor neuron disease. PMID- 12127150 TI - Environmental risk factors and Parkinson's disease: selective degeneration of nigral dopaminergic neurons caused by the herbicide paraquat. AB - Environmental toxicants and, in particular, pesticides have been implicated as risk factors in Parkinson's disease (PD). The purpose of this study was to determine if selective nigrostriatal degeneration could be reproduced by systemic exposure of mice to the widely used herbicide paraquat. Repeated intraperitoneal paraquat injections killed dopaminergic neurons in the substantia nigra (SN) pars compacta, as assessed by stereological counting of tyrosine hydroxylase (TH) immunoreactive and Nissl-stained neurons. This cell loss was dose- and age dependent. Several lines of evidence indicated selective vulnerability of dopaminergic neurons to paraquat. The number of GABAergic cells was not decreased in the SN pars reticulata, and counting of Nissl-stained neurons in the hippocampus did not reveal any change in paraquat-treated mice. Degenerating cell bodies were observed by silver staining, but only in the SN pars compacta, and glial response was present in the ventral mesencephalon but not in the frontal cortex and cerebellum. No significant depletion of striatal dopamine followed paraquat administration. On the other hand, enhanced dopamine synthesis was suggested by an increase in TH activity. These findings unequivocally show that selective dopaminergic degeneration, one of the pathological hallmarks of PD, is also a characteristic of paraquat neurotoxicity. The apparent discrepancy between pathological (i.e., neurodegeneration) and neurochemical (i.e., lack of significant dopamine loss) effects represents another important feature of this paraquat model and is probably a reflection of compensatory mechanisms by which neurons that survive damage are capable of restoring neurotransmitter tissue levels. PMID- 12127152 TI - Beta-amyloid peptides decrease soluble guanylyl cyclase expression in astroglial cells. AB - In astroglial cells beta-amyloid peptides (betaA) induce a reactive phenotype and increase expression of NO synthase. Here we show that treatment of rat brain astrocytes with betaA decreases their capacity to accumulate cyclic GMP (cGMP) in response to NO as a result of a decreased expression of soluble guanylyl cyclase (sGC) at the protein and mRNA levels. Potentiation of betaA-induced NO formation by interferon-gamma did not result in a larger decrease in cGMP formation and inhibition of NO synthase failed to reverse down-regulation of sGC, indicating that NO is not involved. The betaA effect was prevented by the protein synthesis inhibitor cycloheximide. Intracerebral betaA injection also decreased sGC beta1 subunit mRNA levels in adult rat hippocampus and cerebellum. A loss of sGC in reactive astrocytes surrounding beta-amyloid plaques could be a mechanism to prevent excess signalling via cGMP at sites of high NO production. PMID- 12127153 TI - Increased lipid peroxidation in cerebrospinal fluid and plasma from patients with Creutzfeldt-Jakob disease. AB - Oxidative pathomechanisms play an important role in neurodegenerative diseases like Alzheimer's disease (AD). It has been shown that lipid peroxidation in cerebrospinal fluid (CSF) and plasma is increased in AD. To assess the role of oxidative stress in Creutzfeldt-Jakob disease (CJD), we investigated the oxidizability of lipids, the lipid composition and the levels of the antioxidants ascorbate and alpha-tocopherol in CSF and plasma of 15 CJD patients and 12 neurologically healthy controls. CSF and plasma lipid peroxidation was increased in CJD patients and polyunsaturated fatty acids were reduced in CSF of these patients. Ascorbate levels were lower in CSF and plasma of CJD patients, while alpha-tocopherol was found to be decreased in CSF but not in plasma. These results support the hypothesis that oxidative mechanisms are involved in the pathogenesis of CJD and provide a rationale for the use of antioxidants in the therapy of this disease. PMID- 12127154 TI - Substitutions in the conserved C2C domain of otoferlin cause DFNB9, a form of nonsyndromic autosomal recessive deafness. AB - DFNB, the nonsyndromic hearing loss with an autosomal recessive mode of inheritance constitutes the majority of severe to profound prelingual forms of hearing impairment, usually leading to inability of speech acquisition. We analyzed a consanguineous family with autosomal recessive deafness which has been shown to segregate within chromosomal region 2p23.1 (DFNB9; MIM 601071). By SSCP analysis and DNA sequencing of the 48 exons of the DFNB9 gene, coding for otoferlin, previously reported mutations in OTOF were excluded. Next to a frequent T > C single nucleotide polymorphism in exon 8, two novel mutations linked in exon 15 of the OTOF long splice form were identified comprising substitutions at positions 490 (Pro > Gln) and 515 (Ile > Thr), both located in the conserved Ca(2+) binding C2C domain of this peptide. Comparisons of homology using human and mice otoferlins and closely related peptides and computer simulation analyses suggest that changes in the mutated segment's secondary structure affect the Ca(2+) binding capacity of the C2C domain in otoferlin. PMID- 12127155 TI - L-DOPA-induced dyskinesia in the intrastriatal 6-hydroxydopamine model of parkinson's disease: relation to motor and cellular parameters of nigrostriatal function. AB - In order to assess the role of striatal dopamine (DA) afferents in L-DOPA-induced dyskinesia, we have studied a large series of rats sustaining 2, 3, or 4 unilateral injections of 6-hydroxydopamine (6-OHDA) in the lateral striatum. This type of lesion produced a dose-dependent depletion of DA fibers in the caudate putamen, which was most pronounced in the lateral aspects of this structure. An additional group of rats was injected with 6-OHDA in the medial forebrain bundle to obtain complete DA denervation on one side of the brain. During a course of chronic L-DOPA treatment, rats with intrastriatal 6-OHDA lesions developed abnormal involuntary movements (AIMs), which mapped onto striatal domains exhibiting at least approximately 90% denervation, as judged by DA transporter autoradiography. The denervated areas showed local upregulation of preproenkephalin and prodynorphin mRNA, and FosB-like immunoreactivity, which were highly correlated with the rats' AIM scores. When compared to completely DA denervated animals, the rats with intrastriatal 6-OHDA lesions showed an overall lower incidence, lower severity and different topographic distribution of AIMs. The involvement of proximal limb and axial muscles in the abnormal movements was proportional to the spreading of the lesion from lateral towards medial aspects of the caudate-putamen. Locomotive AIMs were only seen in rats with complete lesions, but not in any of the animals with intrastriatal 6-OHDA (which showed > 5% DA fiber sparing in the medial striatum). Intrastriatally 6-OHDA-lesioned rats had a larger therapeutic window for L-DOPA than did rats with complete bundle lesions, since they exhibited an overall lower predisposition to dyskinesia but a similar degree of drug-induced motor improvement in a test of forelimb stepping. Our results are the first to demonstrate that selective and partial DA denervation in the sensorimotor part of the striatum can confer cellular and behavioral supersensitivity to L-DOPA, and that the phenomenology of L-DOPA induced rat AIMs can be accounted for by the topography of DA denervation within the caudate-putamen. PMID- 12127156 TI - Triacylglycerol-rich lipoproteins trigger the phosphorylation of extracellular signal regulated kinases in vascular cells. AB - Postprandial triacylglycerol-rich lipoproteins (TRL) have been implicated in the pathophysiology of atherosclerosis, but the intracellular processes by which TRL could affect vascular function are still unknown. Incubation of TRL obtained at 2 h postprandial period with vascular smooth muscle cells (VSMC) produced a tyrosine phosphorylation of the extracellular signal-regulated kinases 1 and 2 (ERK1 and ERK2) that belong to the mitogen-activated protein kinase (MAPK) family. The activation of ERK1 and ERK2 had a maximum at 15 min, returned to baseline by 60 min, and was partially depleted after incubation of cells with a MAPKK inhibitor (PD 098059). In addition, postprandial TRL did competent VSMC for DNA replication through a MAPK pathway. These effects were dependent of the lipid composition of TRL. Our observations suggest that postprandial TRL can trigger activation of the MAPK pathway and induce a mitogenic response in VSMC in a lipid dependent fashion. PMID- 12127157 TI - In vivo biodistribution and pharmacokinetics of (18)F-labeled human C-peptide: evaluation in monkeys using positron emission tomography. AB - The recently observed beneficial effects exerted by C-peptide in insulin dependent diabetes patients (IDDM) have instigated research into the mechanisms of C-peptide action as well as the location for it. Here we report in vivo biodistribution studies performed in monkeys using positron emission tomography (PET) and C-peptide labeled in the N-terminal with fluorine-18. Following iv injection of the radiotracer, dynamic decay data were collected over the chest and/or abdomens of the monkeys. The radioactivity distributed mainly to the kidneys, less to the heart and to some extent to the liver. Excretion of radioactivity into the urinary bladder was observed. Brain uptake was not detected in a static emission scan of the head performed at late times. Accumulation of radioactivity in the skeleton as a result of in vivo defluorination was not observed. Pharmacokinetic modeling of the regional concentrations of radioactivity over time resulted, for most organs, in two compartment models. The organs with the highest radioactivity concentrations have been identified, enabling dose estimations for studies in humans with low or no C peptide. PMID- 12127159 TI - Aphrodisiac properties of Tribulus Terrestris extract (Protodioscin) in normal and castrated rats. AB - Tribulus terrestris (TT) has long been used in the traditional Chinese and Indian systems of medicine for the treatment of various ailments and is popularly claimed to improve sexual functions in man. Sexual behaviour and intracavernous pressure (ICP) were studied in both normal and castrated rats to further understand the role of TT containing protodioscin (PTN) as an aphrodisiac. Adult Sprague-Dawley rats were divided into five groups of 8 each that included distilled water treated (normal and castrated), testosterone treated (normal and castrated, 10 mg/kg body weight, subcutaneously, bi-weekly) and TT treated (castrated, 5 mg/kg body weight, orally once daily). Decreases in body weight, prostate weight and ICP were observed among the castrated groups of rats compared to the intact group. There was an overall reduction in the sexual behaviour parameters in the castrated groups of rats as reflected by decrease in mount and intromission frequencies (MF and IF) and increase in mount, intromission, ejaculation latencies (ML, IL, EL) as well as post-ejaculatory interval (PEI). Compared to the castrated control, treatment of castrated rats (with either testosterone or TT extract) showed increase in prostate weight and ICP that were statistically significant. There was also a mild to moderate improvement of the sexual behaviour parameters as evidenced by increase in MF and IF; decrease in ML, IL and PEI. These results were statistically significant. It is concluded that TT extract appears to possess aphrodisiac activity probably due to androgen increasing property of TT (observed in our earlier study on primates). PMID- 12127158 TI - In vitro and in vivo pharmacological profile of 5-[2-[4-(6-fluoro-1H-indole-3 yl)piperidin-1-yl]ethyl]-4-(4-fluorophenyl)thiazole-2-carboxylic acid amide (NRA0562), a novel and putative atypical antipsychotic. AB - In vitro and in vivo pharmacological properties of 5-[2-[4-(6-fluoro-1H-indole-3 yl)piperidin-1-yl]ethyl]-4-(4-fluorophenyl)thiazole-2-carboxylic acid amide (NRA0562), a novel atypical antipsychotic, were investigated. NRA0562 showed high affinities for human cloned dopamine D(1), D(2), D(3) and D(4) receptors with Ki values of 7.09, 2.49, 3.48 and 1.79 nM. In addition, NRA0562 had high affinities for the 5-HT(2A) receptor and the alpha(1) adrenoceptor with Ki values of 1.5 and 0.56 nM, and moderate affinity for the histamine H(1) receptor. Using in vivo and ex vivo receptor binding studies in rats, we showed NRA0562 occupied frontal cortical 5-HT(2A) receptors and alpha(1) adrenoceptor potently, while occupancy of striatal dopamine D(2) receptor was moderate as were other atypical antipsychotics. NRA0562 dose-dependently inhibited methamphetamine (MAP)-induced locomotor hyperactivity in rats. At higher dosage, NRA0562 dose-dependently antagonized MAP-induced stereotyped behavior and induced catalepsy dose dependently and significantly in rats. But, the ED(50) value in inhibiting MAP induced locomotion hyperactivity was 10 times lower than that in inhibiting MAP induced stereotyped behavior, and 30 times lower than that in inducing catalepsy. In addition, the potency of NRA0562 in antagonizing MAP-induced hyperactivity in rats was higher than that of other antipsychotics, clozapine, risperidone and olanzapine. NRA0562 had favorable properties in view of prediction of extrapyramidal side effects. As this antipsychotic has a unique profile with affinity and occupancy for receptors, we propose that NRA0652 may have unique atypical antipsychotic activities, and a moderate liability of extrapyramidal motor side effects seen in the treatment with classical antipsychotics. PMID- 12127160 TI - E-cadherin promoter methylation can regulate its expression in invasive ductal breast cancer tissue in Chinese woman. AB - Promoter methylation is an important mechanism of regulating E-cadherin expression. Methylation-specific PCR (MSP) assay was done to evaluate the promoter methylation status of E-cadherin gene in primary tumor samples from 23 cases of Chinese women with invasive ductal breast cancers. Western blotting assay was employed for E-cadherin and beta-actin expressions. Positive MSP results occurred in 26.1% (6/23) of primary tumor samples and none of four normal skin samples. These molecular events tended to occur in breast cancers associated with poor prognosis. Whereas the mean ratio of CDH1/beta-actin for six MSP positive cases was 0.0290 +/- 0.0355, the mean ratio for 17 MSP-negative cases was 0.4726 +/- 0.5049 (P = 0.046). In conclusion, aberrant E-cadherin methylation preferentially occurs in invasive ductal breast cancer associated with poor prognosis and is one of the mechanisms of E-cadherin expression silence in breast cancers from Chinese women. PMID- 12127161 TI - Control of postprandial hyperglycaemia by galactosyl maltobionolactone and its novel anti-amylase effect in mice. AB - The ability to control carbohydrate digestion is useful in the treatment of diabetes mellitus and obesity. In the present study, we examined whether recently developed 4(2)-O-beta-D-galactosyl maltobionolactone (LG2O) having anti-amylase activity is able to control postprandial blood glucose concentration in mice. In addition, we tried to determine how LG2O regulates carbohydrate delivery in the gut lumen by conducting in vivo and in vitro studies. Male non-diabetic ddY mice and KK-A(y) mice, a spontaneously diabetic strain, had free access to a carbohydrate rich diet supplemented with LG2O (3 or 10 g/kg) for 0.5 hr, and blood glucose concentration was measured. LG2O suppressed any steep increase in postprandial blood glucose concentration in both ddY and KK-A(y) mice. Corresponding to the blood glucose response, LG2O also markedly suppressed any increase in postprandial plasma insulin concentration. After ingestion of the diet, LG2O produced a 1.5-3.5 fold increase in the gut contents and reducible sugar content in the small intestine but not in the stomach. Although alpha amylase activity in the stomach was much lower compared with the activity in the small intestine, LG2O still strongly inhibited alpha-amylase activity in the stomach. In contrast, LG2O had little or no influence on alpha-amylase activity in the proximal intestine. From the in vitro carbohydrate digestion stimulation, LG2O at 7.5 mM decreased glucose production by 75% for dextrin, 25% for alpha starch and 60% for raw starch. In conclusion, administration of LG2O inhibits carbohydrate digestion in the gut, and produces significant improvements in both blood glucose and insulin response following ingestion as part of the diet, and this evidence provides support for its therapeutic potential in treating diabetes mellitus and obesity. PMID- 12127162 TI - Progesterone receptor in the liver and oviduct of the lizard Podarcis sicula. AB - In this paper we report the presence of a (3)H-Progesterone ((3)H-P) binding moiety, which has the characteristics of a true receptor, in the liver of the female of the lizard Podarcis sicula. (3)H-P binding studies show the presence of one type of binding site with an average Kd value of 6.2 +/- 2.0 nM in the cytoplasm and 6.3 +/- 1.1 nM in the nucleus. Competition experiments showed that progesterone (P) was the best competitor, while testosterone, deoxycorticosterone (DOC), corticosterone, 17 alpha-hydroxyprogesterone; R5020; RU486 and RU26988-5 were poor competitors. We have also investigated the immunological characteristics of progesterone receptor (PR) in both the liver and the oviduct of Podarcis sicula, by Western blotting using the monoclonal antibody PR22 raised against the PR isoforms A and B of chicken. One imunoreactive band of about 70 kDa was detected in cytoplasmic and nuclear extracts of both the liver and the oviduct. PR immunoreactivity was present in the liver during the quiescent phase. In the oviduct PR immunoreactivity increased from the recovery to the full grown phase. P treatment of estrogen-primed females did not affect the presence of PR in the liver, while brought about a PR increase in the oviduct. This study suggests that PR is expressed differently in the liver and the oviduct of Podarcis sicula throughout the reproductive cycle. PR might fulfill different requirements in relation to the different physiological functions of the tissue during the reproductive cycle. PMID- 12127163 TI - The role of the hypothalamic nitric oxide in the pressor responses elicited by acute environmental stress in awake rats. AB - We quantitatively investigated the change in nitric oxide (NO) in the hypothalamic paraventricular nucleus (PVN) and its effect on cardiovascular regulation during shaker stress (SS) using brain microdialysis in awake rats. Male Wistar rats were fed either N(G)-nitro-L-arginine methyl ester (L-NAME, 0.7 g/L) or tap water for 2 weeks. Two days after implantation of an arterial catheter and guide shaft, a microdialysis probe was placed to perfuse the PVN with degassed Ringer solution at 2 microl/min in awake normotensive Wistar (CONTROL) and chronic L-NAME-treated hypertensive rats. After the rat was placed in a plastic cage set on a shaker, the blood pressure and heart rate was monitored and 10-min SS was loaded at a frequency of 200 cycles/min. Dialysate samples were analyzed by NO analyzer (based on the Griess reaction) every 10 min, and NOx (NO(2)(-) + NO(3)(-)) was measured. Plasma NOx was also measured before and after SS. Pressor responses elicited by SS were significantly greater in L NAME-treated rats than in the CONTROL. Although NOx in the PVN dialysate were increased by SS in the CONTROL, these responses were attenuated in chronic L-NAME treated rats. Resting plasma NOx were higher in the CONTROL than in L-NAME treated rats. SS elicited no difference between two groups in plasma NOx. These results indicated that NO within the PVN, but not in systemic circulation, may play a role on the attenuation of the pressor responses elicited by SS. The dysfunction of NO release within the PVN may, in part, play a role in the exaggerated pressor responses in acute environmental stress. PMID- 12127164 TI - Augmentation of NO-mediated vasodilation in metabolic acidosis. AB - Reduction of perivascular pH in acidemia produces hyporesponsiveness of vascular bed to vasoconstrictors. In the present study, we examined the effects of modest acidification on dilatory responses of isolated rat thoracic aorta. Acetylcholine produced endothelium-dependent relaxation in phenylephrine-precontracted aorta, which was markedly enhanced by acidification of Krebs-Henseleit solution from pH 7.4 to 7.0. A similar augmentation was observed in the relaxing responses to NO donors (SNP, SIN-1, SNAP), 8-Br-cGMP and NS-1619 (a putative K(Ca) channel opener and/or Ca channel inhibitor) in endothelium-denuded, phenylephrine-contracted aorta. However, papaverine-induced relaxation was not affected by the change in pH. At pH 7.4, the relaxing responses to acetylcholine and SNP were partially inhibited by charybdotoxin (K(Ca) channel inhibitor) but not glibenclamide (K(ATP) channel inhibitor), while at pH 7.0 the relaxation induced by either drug was not affected by K(+) channel inhibitors. Relaxation induced by 8-Br-cGMP or NS-1619 was not inhibited by charybdotoxin or glibenclamide. Acidification to pH 7.0 increased the cGMP production in response to acetylcholine in endothelium intact aorta and to SNP in endothelium-denuded aorta. These results show that modest acidification augments NO-mediated relaxation in rat aorta, probably due to an enhancement of cGMP-dependent but K(+) channel-unrelated relaxation mechanisms. PMID- 12127165 TI - Anti-Helicobacter pylori flavonoids from licorice extract. AB - Licorice is the most used crude drug in Kampo medicines (traditional Chinese medicines modified in Japan). The extract of the medicinal plant is also used as the basis of anti-ulcer medicines for treatment of peptic ulcer. Among the chemical constituents of the plant, glabridin and glabrene (components of Glycyrrhiza glabra), licochalcone A (G. inflata), licoricidin and licoisoflavone B (G. uralensis) exhibited inhibitory activity against the growth of Helicobacter pylori in vitro. These flavonoids also showed anti-H. pylori activity against a clarithromycin (CLAR) and amoxicillin (AMOX)-resistant strain. We also investigated the methanol extract of G. uralensis. From the extract, three new isoflavonoids (3-arylcoumarin, pterocarpan, and isoflavan) with a pyran ring, gancaonols A[bond]C, were isolated together with 15 known flavonoids. Among these compounds, vestitol, licoricone, 1-methoxyphaseollidin and gancaonol C exhibited anti-H. pylori activity against the CLAR and AMOX-resistant strain as well as four CLAR (AMOX)-sensitive strains. Glycyrin, formononetin, isolicoflavonol, glyasperin D, 6,8-diprenylorobol, gancaonin I, dihydrolicoisoflavone A, and gancaonol B possessed weaker anti-H. pylori activity. These compounds may be useful chemopreventive agents for peptic ulcer or gastric cancer in H. pylori infected individuals. PMID- 12127166 TI - Contributions of Kv1.2, Kv1.5 and Kv2.1 subunits to the native delayed rectifier K(+) current in rat mesenteric artery smooth muscle cells. AB - A large array of voltage-gated K(+) channel (Kv) genes has been identified in vascular smooth muscle tissues. This molecular diversity underlies the vast repertoire of native Kv channels that regulate the excitability of vascular smooth muscle tissues. The contributions of different Kv subunit gene products to the native Kv currents are poorly understood in vascular smooth muscle cells (SMCs). In the present study, Kv subunit-specific antibodies were applied intracellularly to selectively block various Kv channel subunits and the whole cell outward Kv currents were recorded using the patch-clamp technique in rat mesenteric artery SMCs. Anti-Kv1.2 antibody (8 microg/ml) inhibited the Kv currents by 29.2 +/- 5.9% (n = 6, P < 0.05), and anti-Kv1.5 antibody (6 microg/ml) by 24.5 +/- 2.6% (n = 7, P < 0.05). Anti-Kv2.1 antibody inhibited the Kv currents in a concentration-dependent fashion (4-20 microg/ml). Co-application of antibodies against Kv1.2 and Kv2.1 (8 microg/ml each) induced an additive inhibition of Kv currents by 42.3 +/- 3.1% (n = 7, P < 0.05). In contrast, anti Kv1.3 antibody (6 microg/ml) did not have any effect on the native Kv current (n = 6, P > 0.05). A control antibody (anti-GIRK1) also had no effect on the native Kv currents. This study demonstrates that Kv1.2, Kv1.5, and Kv2.1 subunit genes all contribute to the formation of the native Kv channels in rat mesenteric artery SMCs. PMID- 12127167 TI - Tenses of insomnia epidemiology. PMID- 12127168 TI - Sleep and sleep habits from childhood to young adulthood over a 10-year period. AB - The aim of this epidemiological study was to utilise a cross-sectional as well as a longitudinal approach to examine sleep habits and how they develop in young people in Iceland. The 668 subjects (1-20 years) who responded to a postal survey in 1985 were followed up 5 and 10 years later. The majority of the variance in bedtime and sleep duration was explained by age, but also to a considerable degree by other factors such as residence, season, and year of survey or interaction of these factors. Natural phenomena, such as the diminution of total sleep duration in the first years of life and the tendency for longer sleep on weekends compared to weekdays were confirmed. The lengthening of sleep on weekends was first significant at the age of 9 and was greater among adolescents than young adults. The incidence of daytime sleepiness increased in adolescence, as did napping, at which time their nocturnal sleep time significantly decreased. Over a period of 10 years, a significant shift to earlier wake-up times occurred in children up to 15 years of age, which resulted in a shortened total sleep time. The idea that individual sleep duration is an inherent parameter is supported by the high positive correlation of total sleep time across a 10-year period (r=.73). The present data confirm that Icelandic adolescents (aged 11, 13, and 15) have delayed bedtimes and shorter nocturnal sleep compared to European peers. PMID- 12127169 TI - Sleep apnea, APOE4 and Alzheimer's disease 20 years and counting? AB - Alzheimer's disease (AD) is acknowledged to be at least partially genetic, and one of the key genotypic markers for this condition is the APOE4 allele. Links between sleep apnea and AD have long been suspected because of mental impairment seen in some sleep apnea patients and possible evidence suggesting higher rates of sleep apnea in some dementia patients. The recent demonstration of an association between the APOE4 genotype and sleep apnea has rekindled further interest in this topic, particularly because sleep apnea is characterized by multiple genetic vulnerabilities. We review here evidence related to associations between sleep apnea and dementia, the role of APOE4 as a likely marker for cerebrovascular disease, and discuss treatment considerations relevant to sleep apnea as a potentially reversible cause of dementia. PMID- 12127170 TI - Prevalence of restless legs syndrome and periodic limb movement disorder in the general population. AB - BACKGROUND: Periodic limb movement disorder (PLMD) and restless legs syndrome (RLS) are two sleep disorders characterized by abnormal leg movements and are responsible for deterioration in sleep quality. However, the prevalence of these disorders is not well known in the general population. This study aims to document the prevalence of RLS and PLMD in the general population and to identify factors associated with these conditions. METHODS: Cross-sectional studies were performed in the UK, Germany, Italy, Portugal and Spain. Overall, 18,980 subjects aged 15 to 100 years old representative of the general population of these five European countries underwent telephone interviews with the Sleep-EVAL system. A section of the questionnaire assessed leg symptoms during sleep. The diagnoses of PLMD and RLS were based on the minimal criteria provided by the International Classification of Sleep Disorders. RESULTS: The prevalence of PLMD was 3.9% and RLS was 5.5%. RLS and PLMD were higher in women than in men. The prevalence of RLS significantly increased with age. In multivariate models, being a woman, the presence of musculoskeletal disease, heart disease, obstructive sleep apnea syndrome, cataplexy, doing physical activities close to bedtime and the presence of a mental disorder were significantly associated with both disorders. Factors specific to PLMD were: being a shift or night worker, snoring, daily coffee intake, use of hypnotics and stress. Factors solely associated with RLS were: advanced age, obesity, hypertension, loud snoring, drinking at least three alcoholic beverages per day, smoking more than 20 cigarettes per day and use of SSRI. CONCLUSIONS: PLMD and RLS are prevalent in the general population. Both conditions are associated with several physical and mental disorders and may negatively impact sleep. Greater recognition of these sleep disorders is needed. PMID- 12127171 TI - Sleep complaints cosegregate with illness in older adults: clinical research informed by and informing epidemiological studies of sleep. AB - OBJECTIVE: The more recent epidemiological studies of sleep complaints have shown that when factors such as physical and psychiatric illness, medication use and drug and alcohol use are accounted for, the age-related increase in prevalence of sleep complaints is strikingly less than in those earlier studies without such controls. In an effort to support this finding, we examined the relationships between clinical health screenings and sleep complaints and disorders in two large groups of potential research volunteers (total N=2954). METHODS: As part of a study of older adults at risk for Alzheimer's disease, two large groups of potential research volunteers (n=1619 and 1335, respectively) replied to advertisements that made no mention of sleep and underwent three increasingly rigorous levels of health screening. At each level of screening, medical and psychiatric health and history were assessed and, where appropriate, subjects were excluded from further study participation before any mention of sleep quality or sleep complaint was made. The remaining subjects then underwent polysomnography. RESULTS: Of 1619 elderly adult volunteers screened in this manner, only 51 (3.14%) were found to have significant sleep complaints or disorders. The second group of 1335 screened in this manner revealed a similar pattern with only 18 (1.35%) having significant sleep complaints or disorders. CONCLUSIONS: These findings indicate that careful health assessments will screen out most sleep complaints and disorders in the older population and lend further support to the epidemiological evidence demonstrating that the bulk of geriatric sleep complaints and disorders is not the result of age per se, but rather cosegregates with medical and psychiatric disorders and related health burdens. PMID- 12127172 TI - Impact of insomnia on future functioning of adolescents. AB - OBJECTIVE: To examine the impact of insomnia among adolescents on somatic, interpersonal, and psychological functioning using data from a two-wave, prospective study. METHODS: Subjects were adolescents 11-17 years of age sampled from managed care enrollment rosters in the United States. The baseline sample was 4175 and the follow-up sample a year later was 3136. Data were collected using computer-assisted personal interviews and self-administered questionnaires. Insomnia was measured using DSM-IV symptom criteria, scored as a summated scale and as separate symptoms. Indicators of somatic functioning were perceived health, limitations due to health problems, and impact of illness on family activities. Indicators of interpersonal functioning were social support, relations with parents, relations with peers and relations at school. Indicators of psychological functioning were self-esteem, perceived mental health, life satisfaction, and depression. RESULTS: Symptoms of insomnia were common, with 17% reporting nonrestorative sleep, 6% difficulty initiating sleep, 7% daytime fatigue, and 5% daytime sleepiness almost every day. Without adjustment for insomnia at follow-up, the odds of dysfunction at follow-up for those with insomnia averaged 2.5 across 11 indicators of functioning. For 9 of 11 indicators, there was a clear dose-response relation such that moderate levels of insomnia increased risk and this risk increased with greater insomnia. Adjusting for insomnia at follow-up eliminated the association with somatic functioning and attenuated associations with interpersonal and psychological functioning. DISCUSSION: These prospective data provide additional evidence that insomnia can have adverse consequences for the functioning of adolescents. Given the growing evidence for a relation between disturbed sleep and impaired adolescent functioning, more attention needs to be directed to identifying causal pathways and possible strategies for intervention. PMID- 12127173 TI - Substance use for insomnia in Metropolitan Detroit. AB - OBJECTIVE: People with insomnia are not typically treated medically for their insomnia. Studies have reported approximately 30% of insomniacs self-medicate with alcohol or over-the-counter (OTC) medications. This study was done to identify determinants and risks of different insomnia therapeutics. METHODS: A random-digit-dial, computer-assisted survey of a representative sample of adults in Metropolitan Detroit, aged 18-65 years, is being conducted. A sample of all respondents over an 18-month period was collected (n=1324) with a 68% response rate. Exclusive past-year use of alcohol for sleep was reported by 10% (n=132), prescription medications by 8% (n=108), and OTC medications by 10% (n=135). Five percent used both alcohol and sleep medications. The three exclusive-use groups formed the comparison groups of the study. RESULTS: The prescription drug group used medications for more consecutive nights and for more total nights than the alcohol and OTC users. Alcohol users were predominantly male, while OTC and prescription drug users were predominantly female. Alcohol users were more likely to be single than the others, and prescription drug users were older than the others. Prescription drug users had more severe insomnia and had greater disability, neuroticism, and daytime fatigue than the others. In contrast, the alcohol users had greater daytime sleepiness than the others. CONCLUSIONS: In Metropolitan Detroit, insomniacs receiving medical treatment have more severe insomnia and greater disability than those who self-treat. However, while the insomnia of those self-treating is less severe, it is still associated with some risks. PMID- 12127174 TI - Prevalence and consequences of sleep disorders in a shift worker population. AB - INTRODUCTION: Irregular work schedules often results in a disruption of the normal circadian rhythm that can causes sleepiness when wakefulness is required and insomnia during the main sleep episode. METHOD: Two physicians using the Sleep-EVAL system interviewed 817 staff members of a psychiatric hospital. The interviews were done during the working hours. In addition to a series of questions to evaluate sleep and mental disorders, the evaluation included a standard questionnaire assessing work conditions, work schedule and their consequences. Three work schedules were assessed: (1) fixed daytime schedule (n=442), (2) rotating daytime shifts (n=323) and (3) shift or nighttime work (n=52). RESULTS: Subjects working on rotating daytime shifts were younger than the two other groups and had a higher proportion of women. Participants working on rotating daytime shifts reported more frequently than the fixed daytime schedule workers to have difficulty initiating sleep (20.1% vs. 12.0%). The sleep duration of shift or nighttime workers was shorter than that of the two other groups. Furthermore, subjects working rotating daytime schedule reported to have shorter sleep duration of about 20 min when they are assigned to the morning shift. Work-related accidents were two times more frequent among the rotating daytime workers (19.5%) compared with the fixed daytime schedule workers (8.8%) and the group of nighttime or shift workers (9.6%). Sick leaves in the previous 12 months were also more frequently reported in the rotating daytime schedule group (62.8%) as compared with the daytime group (38.5%, P<.001); 51.9% of nighttime or shift workers took sick leave. CONCLUSIONS: Working on a rotating daytime shifts causes significant sleep disturbances. As consequences, these workers are more likely to feel sleepy at work and are more likely to have work related accidents and sick leaves. PMID- 12127175 TI - Work load and work hours in relation to disturbed sleep and fatigue in a large representative sample. AB - OBJECTIVE: To study the relation between work and background factors on the one hand and disturbed sleep and fatigue on the other. METHOD: A representative national sample of 58,115 individuals was selected at regular intervals over a period of 20 years and interviewed on issues related to work and health. The data were subjected to a multiple logistic regression analysis. RESULTS: The number of cases was 18,828 (32.8%) for fatigue and 7347 (12.8%) for disturbed sleep. For disturbed sleep, the significant predictors became: female gender, age above 49 years, present illness, hectic work, physically strenuous work, and shift work. For fatigue, the significant predictors became female gender, age below 49 years, high socioeconomic status, present illness, hectic work, overtime work, and physically strenuous work. CONCLUSION: Work stress, shift work, and physical workload interfere with sleep and are related to fatigue. PMID- 12127176 TI - Insomnia in central Pennsylvania. AB - OBJECTIVE: Establish the association between insomnia and various physical and mental health symptoms as well as objective measures of sleep disturbance while controlling for age, gender and BMI in a large random sample of the general public. METHODS: A subsample (N=1741) was selected for a single-night sleep laboratory evaluation from a larger random sample (N=16,583) of the general public (20-100 years old). RESULTS: The prevalence of insomnia was 7.5% and difficulty sleeping an additional 22.4%. The complaints were more frequent in women and in non-Caucasian minorities. A multivariate logistic regression analysis indicated that depression was the single strongest factor followed by female gender associated with either insomnia or difficulty sleeping. Minority status and a history of colitis, hypertension and anemia were also associated, but to a lesser degree. The final model did not include age, BMI as well as any of the sleep laboratory findings. CONCLUSION: These findings support the conclusion that mental health variables have the primary independent association with a complaint of insomnia. Other factors including minorities and hypertension are also independently associated, though to a lesser degree. Other primary sleep disorders, e.g., sleep apnea, do not seem to play a major role in insomnia. These findings underscore the fact that insomnia is a symptom associated with a wide variety of mental and physical health problems requiring a proper psychiatric and medical management. PMID- 12127177 TI - Prevalence of insomnia and associated factors in South Korea. AB - INTRODUCTION: In Western countries, insomnia is associated with daytime impaired functioning, as well as physical and psychiatric illnesses. However, little information exists on insomnia in Asian countries. This study investigates the prevalence and correlates of insomnia in the general population of South Korea. METHODS: A representative sample of the South Korean general population composed of 3719 noninstitutionalized individuals aged 15 years or older were interviewed by telephone using the Sleep-EVAL system. The participation rate was 91.4%. The interviews covered sleep habits, sleep symptomatology, physical and psychiatric illnesses. DSM-IV sleep and psychiatric disorder diagnoses were also assessed. RESULTS: Insomnia symptoms occurring at least three nights per week were reported by 17.0% of the sample; difficulty initiating sleep (DIS) was mentioned by 4.0% of the sample, difficulty maintaining sleep (DMS) by 11.5%, early morning awakenings (EMA) by 1.8%, and nonrestorative sleep (NRS) by 4.7% of the sample. DSM-IV insomnia disorder diagnoses were found in 5% of the sample. Over 50% of subjects with insomnia symptoms reported important daytime consequences and another 20% reported mild or moderate consequences. However, the proportion of insomnia subjects seeking medical help for their sleep problems was very low (6.8%). CONCLUSIONS: As in Western countries, insomnia is widespread in South Korea, affecting nearly one in five individuals. Many of them would benefit from medical help; however, few insomnia subjects are consulting for this problem. An educational effort is needed for both the general population and the physicians. PMID- 12127178 TI - Gender differences in insomnia--a study in the Hong Kong Chinese population. AB - OBJECTIVE: To study the epidemiology of insomnia in the adult Chinese population in Hong Kong and to examine the potential gender-related demographic and lifestyle factors in insomnia. METHODS: A population study via random telephone survey with a structured questionnaire was carried out for noninstitutionalized Chinese adults aged 18-65 by trained lay interviewers. The questionnaire included demographic data, sleep habits and problems, insomnia symptoms and lifestyle questions. RESULTS: A total of 9851 subjects (46.4% male; 53.6% female) were included in the final analysis. The overall prevalence of Hong Kong Chinese as suffering from insomnia during the preceding month (with a frequency of sleep disturbance of at least three times per week) was 11.9% (95% CI 11.2-12.6), including difficulty in initiating sleep (DIS) (4.5%; 95% CI 4.1-5.0), difficulty in maintaining sleep (DMS) (6.9%; 95% CI 6.4-7.5) and early morning awakening (EMA) (4.0%; 95% CI 3.6-4.4). Females were about 1.6 times at higher risk for insomnia than males. The prevalence of insomnia was also shown to increase with age. Multivariate analysis showed that unemployment, lower economic status, alcohol consumption, regular medication and psychiatric disturbance were all associated with higher risks of insomnia in both sexes. Furthermore, lower education level and being retired was associated with a higher risk of insomnia in males, but being a housewife, divorced/widowed, and complaining of a nocturnal noisy environment were associated with a higher risk of insomnia in females. Among all these factors, psychiatric disturbance was the most influential risk factor for insomnia in both sexes. The reasons for gender differences of insomnia may include their differences in the prevalence of psychiatric morbidities, symptom endorsement, gonadal steroids, sociocultural factors and coping strategies. CONCLUSIONS: Overall, 11.9% of the Hong Kong Chinese adult population complained of frequent insomnia in the preceding month. There was a higher prevalence of insomnia in females. Although there were common risk factors for insomnia in both sexes, there existed gender-specific risk factors. PMID- 12127179 TI - Chronic insomnia, postmenopausal women, and sleep disordered breathing: part 1. Frequency of sleep disordered breathing in a cohort. AB - OBJECTIVES: A cohort of postmenopausal women complaining of chronic insomnia for over 6 months and free of hypnotic intake was recruited mostly from the community. Three hundred and ninety-four women were included. The following questions were addressed: How many presents sleep disordered breathing (SDB)? Which type of SDB (upper airway resistance syndrome [UARS] or obstructive sleep apnea syndrome [OSAS]) is the most frequent? Is there a specific upper airway anatomical abnormality in SDB patients predisposing to the syndrome? POPULATION: Subjects were recruited in the community or referred by the Sleep Clinic and all had complaint of chronic poor sleep. METHODOLOGY: First step. Questionnaires, visual analog scales, clinical interview, clinical evaluation with work-up, actigraphy, and ambulatory monitoring were used. Second step. Otolaryngologic evaluation, ambulatory sleep monitoring, and reading of results were used. Subjects negative for SDB at ambulatory monitoring had polysomnography (PSG) with pressure transducer/nasal cannula system and esophageal manometry measurements. RESULTS: Population. Three hundred and ninety-four individuals responded to all entry criteria. Ambulatory monitoring identified 194 subjects with OSAS. Two hundred individuals were not recognized with SDB and were submitted to PSG. This further testing showed that 68 subjects had normal breathing, 62 had UARS, and 100 mild OSAS. Based on otolaryngological evaluation, subjects were classified based on the presence or absence of narrow upper airway, and the location of narrowing was assessed. CONCLUSION: A total of 326 postmenopausal women complaining of chronic insomnia had a SDB, usually with low apnea-hypopnea index (AHI). This total represents about 83% of the studied women. Questions of the role of SDB in the complaint of chronic insomnia are raised. PMID- 12127181 TI - Attention and the control of posture and gait: a review of an emerging area of research. AB - Research on the relationship between attention and the control of posture and gait is a new and expanding area with studies on young adults revealing the role of cognitive factors in the control of balance during standing and walking. The use of dual task paradigms to examine the effect of age related changes in attentional requirements of balance control and age-related reductions in stability when performing a secondary task has shown that these are important contributors to instability in both healthy and balance-impaired older adults. The attentional demands of balance control vary depending on the complexity of the task and the type of secondary task being performed. New clinical assessment methods incorporating dual-task paradigms are helpful in revealing the effect of disease (e.g. Parkinson's disease) on the ability to allocate attention to postural tasks and appear to be sensitive measures in both predicting fall risk and in documenting recovery of stability. PMID- 12127180 TI - Chronic insomnia, premenopausal women and sleep disordered breathing: part 2. Comparison of nondrug treatment trials in normal breathing and UARS post menopausal women complaining of chronic insomnia. AB - OBJECTIVE: The question addressed here is: Can a discrete sleep disordered breathing (SDB) play a role in the insomnia complaint of postmenopausal chronic insomniacs? To respond to the query, two groups of individuals derived from a cohort of postmenopausal chronic insomniacs recruited mostly from the community were enlisted in a treatment protocol. These subjects were all individuals identified with normal breathing (n=68) and all those recognized with Upper Airway Resistance Syndrome (UARS) (n=62) pooled from a cohort of 349 postmenopausal insomniacs. TREATMENT PROTOCOL: The 62 UARS were allocated to either treatment of chronic insomnia by behavioral approaches or treatment of SDB. Based on ENT evaluation, health professionals in charge of patients selected either treatment with nasal CPAP or treatment of nasal turbinates. A stratification correction was performed to obtain a near equal number of both treatment modalities in each of the two subgroups. The 68 individuals with normal breathing were randomly allocated to immediate behavioral treatment of insomnia or delay treatment of insomnia. The delay treatment received a list of 10 sleep hygiene recommendations by mail. METHODOLOGY: Questionnaires, visual analog scales (VAS), Epworth Sleepiness Scale (ESS), clinical interviews, clinical evaluation with oto-laryngologic clinical assessment of a presence/absence of narrow upper airway and location of narrowing. Actigraphy and polysomnography (PSG) with pressure transducer/and nasal cannula system and esophageal manometry. DATA ANALYSES: All recording data were scored blind to patient's condition. RESULTS: Two subjects in the SBD-CPAP treated group (Group B) and two subjects in the delayed treatment group (Group D) dropped out. Total sleep time was improved compared to baseline in all groups, including the delayed treatment group. One group was significantly better (ANOVA, P=.05) with a more important delta score compared to baseline, and this was the behaviorally treated non-SDB. Sleep latency was significantly decreased in the behaviorally treated group (with or without SBD), P=.05, compared to SBD-treated and delayed treatment groups. Sleep latency was, however, improved in all groups. VAS for "quality of sleep" was higher at 6 months in all the groups when compared to "baseline" values. VAS for "daytime fatigue" showed significant differences among the four groups (ANOVA, P=.01); the overall score at the end of treatment was significantly better in the SDB-treated group than the other three groups. SBD was treated either by radio frequency on nasal turbinate or by nasal CPAP. CPAP-treated patients had a lower VAS score than nasal turbinate treatment, but the difference was only a trend. The delta improvement (6-month baseline condition) in "daytime fatigue" of each subgroup was calculated and compared within and between groups. Despite the small number of subjects, the turbinate-treated subgroup was significantly different from Groups B, C and D (ANOVA, P=.05). When a similar comparison was made with the nasal CPAP group, there was only a nonsignificant trend when compared to Groups B, C and D. CONCLUSION: Abnormal breathing during sleep significantly enhanced complaints of daytime fatigue in postmenopausal chronic insomniacs and this complaint improved with SDB treatment. This improvement is significantly better compared to SDB insomniacs treated with a behavioral regimen. Behavioral treatment, however, gave the best response in the non-SDB chronic insomnia group and improved better long sleep latency even in the SDB group. These results suggest the need to find an appropriate treatment for SBD even if mild and to recognize the role of SDB in relation to symptoms seen with chronic insomnia. PMID- 12127182 TI - The influence of multiple obstacles in the travel path on avoidance strategy. AB - Obstacle avoidance strategy was analysed as participants were subjected to varied quantities and positions of obstacles in their travel path. Kinematic data were collected with three optoelectric cameras sampling at 100 Hz. Calculated from this data were toe-elevation and take-off distance for both lead and trail limbs. Each subject completed a total of 120 randomly assigned trials (15 repetitions in each of eight test conditions). Test conditions were as follows: no obstacle (C1), a single obstacle at 0 (C2), 1 (C3), 1.5 (C4), and 2 m (C5) positions and double obstacles at 0 and 1 (C6), 0 and 1.5 (C7) and 0 and 2 m (C8) positions. Avoidance strategy was defined in terms of take-off distance and toe-elevation at obstacle crossing. Single obstacle trials demonstrated that take-off distance and toe-elevation are gait parameters, which are controlled in successful obstacle clearance. Double obstacle trials revealed that presence and position of a second obstacle in the travel path influences trail limb take-off distance for both first and second obstacles. PMID- 12127183 TI - Contributions of altered sensation and feedback responses to changes in coordination of postural control due to aging. AB - We used multivariate kinematics and joint torque measurements during dynamic posturography to determine the relative contributions of changes in overall control gain, relative weighting of sensors, and noise-like effects on posture control in the elderly. Our results show that sway coordination and amplitude both change with age, but that changes in overall feedback gains do not explain these differences. We propose that increased sway of elderly subjects in platform sway-referenced conditions is due to sensory noise or decreased ability to detect small motions of the platform, while increased sway during visual sway referencing is due to re-weighting of the various sensors. PMID- 12127184 TI - Knee and hip kinetics during normal stair climbing. AB - Understanding joint kinetics during activities of daily living furthers our understanding of the factors involved in joint pathology and the effects of treatment. In this study, we examined hip and knee joint kinetics during stair climbing in 35 young healthy subjects using a subject-specific knee model to estimate bone-on-bone tibiofemoral and patello-femoral joint contact forces. The net knee forces were below one body weight while the peak posterior-anterior contact force was close to one body weight. The peak distal-proximal contact force was on average 3 times body weight and could be as high as 6 times body weight. These contact forces occurred at a high degree of knee flexion where there is a smaller joint contact area resulting in high contact stresses. The peak knee adduction moment was 0.42 (0.15) Nm/kg while the flexion moment was 1.16 (0.24) Nm/kg. Similar peak moment values, but different curve profiles, were found for the hip. The hip and knee posterior-anterior shear forces and the knee flexion moment were higher during stair climbing than during level walking. The most striking difference between stair ascent and level walking was that the peak patello-femoral contact force was 8 times higher during stair ascent. These data can be used as baseline measures in pathology studies, as input to theoretical joint models, and as input to mechanical joint simulators. PMID- 12127185 TI - Development of a reliable method to assess footwear comfort during running. AB - The purposes of this study were: (a) to determine whether subjects are able to distinguish between differences in footwear with respect to footwear comfort; and (b) to determine how reliably footwear comfort can be assessed using a visual analogue scale (VAS) and a protocol including a control condition during running. Intraclass correlation coefficients (ICCs) between comfort ratings for repeated conditions were high (ICC = 0.799). Differences in comfort ratings between the insert conditions were significant. A paired t-test revealed a significant difference in overall comfort ratings for the control insert when tested after the soft insert compared to when tested after the hard insert (P = 0.008). The results of this study showed that VASs provide a reliable measure to assess footwear comfort during running under the conditions that: (a) a control condition is included; and (b) the average comfort rating of sessions 4-6 is used. PMID- 12127186 TI - Knee joint functional range of movement prior to and following total knee arthroplasty measured using flexible electrogoniometry. AB - The functional ranges of movement of the knee were investigated in a group of patients with knee osteoarthritis (n = 42, mean age 70 years) before, 4 months and at 18-24 months after total knee arthroplasty and then compared with age matched normal subjects (n = 20, mean age 67 years). Flexible electrogoniometry was used to record the maximum flexion-extension angle, the minimum flexion extension angle and flexion-extension excursions of both knees during eleven functional activities along with the active and passive knee joint range of motion measured using a manual goniometer. Over the eleven functional activities the patients pre-operatively exhibited 28% less knee joint excursion than normal age matched subjects. By 18-24 months following total knee arthroplasty only 2% of this deficit was recovered. Statistically this recovery was only significant in level walking, slope ascent and slope descent. A greater range of movement was measured in a non-weight bearing position than was used in weight bearing functional activity. It is concluded that total knee arthroplasty gives rise to little improvement in knee motion during functional activities and that functional range of movement of the knee remains limited when compared to normal knee function for a minimum of 18 months following operation. PMID- 12127187 TI - Standing balance evaluation using a triaxial accelerometer. AB - This paper presents a new inherently triaxial accelerometer-based system for determining the ability to maintain balance while standing. In this study, the accelerometer was placed at the back of the subject at the approximate height of the centre of mass. The data were processed to obtain five performance parameters. Paired t-tests indicated that the accelerometer measurements were able to distinguish between the different test conditions as well as or better than simultaneous AMTI force platform measurements (P < or = 0.05). The accelerometer system is fully portable, independent of inclination in space, low cost and allows long term measurements of standing balance. PMID- 12127188 TI - Trunk accelerometry as a measure of balance control during quiet standing. AB - The goal was to investigate whether body sway measured by trunk accelerometry during quiet standing could differentiate between young and elderly healthy subjects, and between conditions with eyes open and closed on firm and compliant surfaces. Raw data demonstrated poor discrimination, but horizontal transformation to eliminate a constant gravity component disclosed consistent mean differences across groups (P < or = 0.0025), and also between conditions (P < or = 0.0005), except for the most marginal difference. When drift associated with very low frequency body sway was adjusted for, a significant difference appeared also here (P = 0.001). This study indicates that trunk accelerometry can discriminate between populations and conditions during quiet standing. PMID- 12127189 TI - Acquirement of stability and mobility in infant gait. AB - We examined gait development in a longitudinal and cross-sectional study in 35 infants, age range 7-70 months. We estimated walking stability from mediolateral motion of body segments and mobility from the angular displacement of joints. Motion at the shoulder, hip, knee and ankle decreased significantly over several months after the onset of walking and thereafter changed gradually. The remarkable decrease began distally. The trunk-thigh and thigh-shank angles changed significantly until 9 months after the onset of walking. These results indicate that lateral stability, which develops earlier than mobility, is the most important factor in gait development in infants. PMID- 12127190 TI - Speed dependence of averaged EMG profiles in walking. AB - Electromyogram (EMG) profiles strongly depend on walking speed and, in pathological gait, patients do not usually walk at normal speeds. EMG data was collected from 14 muscles in two groups of healthy young subjects who walked at five different speeds ranging from 0.75 to 1.75 ms(-1). We found that average EMG profiles varied in a predictable way with speed. The average EMG profile for each muscle at any speed could be estimated in a simple way from two functions, one constant and one proportionally increasing with walking speed. By taking into account the similarity among profiles within functional groups, the number of basic functions could be reduced further. Any average EMG profile among the 14 leg muscles studied at all speeds in the measured range could be predicted from six constant and ten speed-dependent basic patterns. These results can be interpreted in terms of a central pattern generator for human walking. PMID- 12127191 TI - Changes in weight-bearing following injury or surgical reconstruction of the ACL: relationship to quadriceps strength and function. AB - PURPOSE: Changes in weight-bearing an average of 1.5 months after ACL injury (ACLD) and up to 3 months after surgical reconstruction (ACLR) were investigated using a force platform. Correlations between force platform test variables, quadriceps strength and functional self-report scores were also examined. RESULTS: Each force platform test revealed impaired weight-bearing by either ACLD and/or ACLR subjects. Many variables were correlated to quadriceps strength; therefore, improved strength in ACLR subjects by 6 weeks post-operative negated any weight-bearing impairment. No single force platform variable correlated with the subjects' perception of function. CONCLUSION: Only the step up/over test provided insight into disability in these patients. Modification of force platform analysis programs may allow for other clinical questions to be answered. PMID- 12127193 TI - History of abuse and drinking outcomes following inpatient alcohol treatment: a prospective study. AB - Little is known about the impact of sexual or physical abuse history on response to alcohol treatment. This prospective study investigated the relationships between sexual and physical abuse histories, participants' characteristics, and response to inpatient alcohol treatment. Forty-one women and 59 men were assessed monthly for 1 year following hospitalization for alcohol dependence. Survival analyses showed that sexual abuse history was associated with shorter times to first drink and relapse. Physical abuse history was not associated with poorer drinking outcomes. Although women were more likely than men to have a history of sexual abuse, no gender differences were found in drinking outcomes. Poorer drinking outcomes were found among participants who at baseline were not married, had less than a college education, were not employed full time, or carried a diagnosis of depression or other psychiatric disorder. When adjusted for these characteristics, the associations between sexual abuse history and times to first drink and relapse were no longer statistically significant. While sexual abuse history is a clinically meaningful predictor of return to drinking we note the importance of considering patients' background and clinical characteristics in examining the impact of sexual abuse history on drinking outcomes following treatment. PMID- 12127194 TI - Mood and anxiety symptoms among 140 children from alcoholic and control families. AB - OBJECTIVE: Children of alcoholics have been reported to have elevated levels of internalizing symptoms, including anxiety and depression. However, many studies have not adequately controlled for the influence of independent (i.e. not substance-induced) parental mood or anxiety disorders and other factors. The present evaluations assess the relationships of the family histories of alcohol use disorders and independent mood and anxiety disorders to internalizing symptoms in children of alcoholic and nonalcoholic subjects. METHOD: A behavioral checklist and a structured interview were administered to the parents of 140 children aged 7-18 years. The fathers of these offspring had been recruited 15 years previously from a university population to participate in a prospective study of 453 men from alcoholic and nonalcoholic families. RESULTS: While a higher score for one of four measures of internalizing symptoms in the children was found to relate to a higher density of alcoholic relatives, this pattern was more robust in children of parents with mood or anxiety disorders. In a hierarchical regression, the family history of alcohol use disorders did not add significantly to the prediction of any of the four internalizing scores in the children after considering the impact of a family history of independent mood and anxiety disorders. CONCLUSIONS: The results indicate that internalizing symptoms in children of alcoholics were more strongly influenced by a positive family history of mood and anxiety disorders than the family history of alcohol use disorders. PMID- 12127195 TI - Marijuana use and cessation of tobacco smoking in adults from a community sample. AB - Tobacco smokers are more likely to use marijuana than those who do not smoke tobacco. Little is known about how marijuana use affects the probability of tobacco smoking cessation. This analysis was based on 431 adults less than 45 years of age who reported recent tobacco smoking in the 1981 baseline interview in the household-based Baltimore Epidemiologic Catchment Area study and were re interviewed 13 years later. At baseline, 41% of the tobacco smokers reported ever use of marijuana, 27% reported use of marijuana in the previous 30 days, and 9% reported daily use of marijuana for 2 weeks or more in the last 30 days. Marijuana users in the past 30 days at baseline were more likely than nonusers to still be using tobacco at follow-up after adjusting for race, educational level and marital status (OR = 1.94, 95% CI = 1.03, 3.63). Daily use of marijuana at baseline was even more strongly related to continued tobacco smoking 13 years later. Difficulty in tobacco cessation might be considered one of the most important adverse effects of marijuana use. Clinicians working with patients who are trying to stop tobacco smoking may be aided by routinely assessing marijuana use history, particularly with the recent increase in co-smoking of marijuana and tobacco. PMID- 12127196 TI - Abstinence at intake for marijuana dependence treatment predicts response. AB - Abstinence prior to entering treatment is common among individuals seeking substance abuse treatment. The current study examined the relationship between abstinence at a pretreatment intake assessment and treatment response during outpatient treatment for marijuana dependence. At the intake assessment, 142 marijuana-dependent individuals completed past 30 day calendars of daily drug use. Forty-four (31%) participants were pretreatment abstainers, as defined by reports of one or more consecutive days of marijuana abstinence prior to the day of the intake assessment. Non-abstainers (69%) reported marijuana use the day prior or the day of the assessment. Pretreatment abstainers were more likely to enter treatment (P < 0.05) and showed better treatment response than non abstainers. Abstainers provided 50% more marijuana-negative urine screens during treatment (P < 0.05), and more than three times as many abstainers reported no marijuana use (P < 0.01). The groups did not differ on treatment completion. Marijuana abstinence at the time of initial clinic contact appears to be a strong predictor of success during treatment. Pretreatment abstinence may prove useful as a pretreatment matching strategy that could improve outcomes and cost effectiveness. Clinical trials might consider including pretreatment abstinence status as a stratification variable during participant assignment or as a covariate in outcome analyses. PMID- 12127198 TI - The validity of drug use self-reports among hard core drug users in a household survey in Puerto Rico: comparison of survey responses of cocaine and heroin use with hair tests. AB - The extent to which underreporting of drug use in household surveys affects the validity of epidemiological studies of drug use disorders is largely unknown. We developed a list of known hard core drug users as part of a larger household study in Puerto Rico. The known drug users were recruited and interviewed with the same procedures used for the respondents selected through area-probability sampling. Upon completion of the interview, subjects were asked to provide a sample of scalp hair. A total of 78 hair specimens were collected from the known drug users. Hair specimens were screened for cocaine and heroin using radio immunoassay, and confirmed using gas chromatography/mass spectrometry. Using the cutoff of 0.2 ng/mg of hair, 93.2% of the hair specimens were classified positive for cocaine and 75.7% for heroin. With the hair test results as the gold standard, we calculated specificity and sensitivity statistics as measures of the validity of self-reports. Self-reports of drug use in the past 3 months had a specificity of 78% or higher for both drugs. The sensitivity of self-reports was 69.6% for reports of recent cocaine use and 78.6% for reports of recent heroin use. Sensitivity increased with reports of use in more remote time periods, among subjects reporting DSM-IV drug disorder symptoms, and among those reporting use of both drugs. The results suggest that while drug reports of hard core drug users interviewed in household surveys might be more valid than those of the general population, there still remains considerable under-reporting. PMID- 12127197 TI - Physiochemical and pharmacological characterization of a Delta(9)-THC aerosol generated by a metered dose inhaler. AB - The goal of the present study was to formulate a Delta(9)-tetrahydrocannabinol (Delta(9)-THC) metered-dose inhaler (MDI) that can be used to provide a systemic dose of Delta(9)-THC via inhalation. Following physiochemical characterization and accelerated stability testing of the aerosol, mice were exposed to the aerosol and evaluated for pharmacological effects indicative of cannabinoid activity, including hypomotility, antinociception, catalepsy, and hypothermia. The fine particle dose of Delta(9)-THC was 0.22 +/- 0.03 mg (mean +/- S.D.) or 25% of the emitted dose and was not affected by accelerated stability testing. A 10-min exposure to aerosolized Delta(9)-THC elicited hypomotility, antinociception, catalepsy, and hypothermia. Additionally, Delta(9)-THC concentrations in blood and brain at the antinociceptive ED(50) dose were similar for both inhalation and intravenous routes of administration. Finally, pretreatment with the CB(1) receptor antagonist SR 141716A (10 mg/kg, i.p.) significantly antagonized all of the Delta(9)-THC-induced effects. These results indicate that an MDI is a viable method to deliver a systemic dose of Delta(9) THC that elicits a full spectrum of cannabinoid pharmacological effects in mice that is mediated via a CB(1) receptor mechanism of action. Further development of a Delta(9)-THC MDI could provide an appropriate delivery device for the therapeutic use of cannabinoids, thereby reducing the need for medicinal marijuana. PMID- 12127199 TI - Influence of estrous cycle and estradiol on behavioral sensitization to cocaine in female rats. AB - The hypotheses that the estrous cycle and estradiol modulate behavioral sensitization to cocaine in female rats were assessed. In an analysis of sensitization across the estrous cycle, female rats were administered saline or cocaine (15 mg/kg) twice daily for 5 days. Sensitization developed in the intact female rats as measured by the significant increase in stimulant behaviors seen between day 1 and day 5 of treatment. Rats were challenged with cocaine (5 mg/kg) at 3 days following discontinuation of drug treatment. The expression of sensitization as measured between cocaine and saline-treated rats was evident only in female rats in diestus at the time of the challenge test with cocaine. To explore the role of estradiol in sensitization, female rats were ovariectomized or ovariectomized and implanted with estradiol for two weeks prior to treatment with cocaine (15 mg/kg) twice daily for 5 days. Sensitization developed in both ovariectomized and ovariectomized+estradiol rats treated with cocaine as measured by the significant increase in stimulant-like behaviors seen between day 1 and day 5 of treatment. Rats were challenged with 5 mg/kg of cocaine at 3, 13 and 34 days following discontinuation of drug treatment. While neither hormone treatment group exposed to the cocaine regimen expressed sensitization at 3 days of withdrawal, both groups exhibited sensitization at 13 and 34 days following discontinuation of cocaine treatment. The estradiol-treated groups exhibited higher levels of activity relative to their untreated cohorts in both saline or cocaine treatment groups. These results suggest that detection of sensitization in female rats is not only influenced by injection regimen and length of abstinence but also by the presence of estrogens which effectively enhance the response to an acute cocaine challenge in the presence or absence of prior cocaine exposure. PMID- 12127200 TI - Patterns of cocaine self-administration in rats produced by various access conditions under a discrete trials procedure. AB - Frequency of drug access greatly affects the pattern and stability of cocaine self-administration. Previous research has shown that restricted drug availability produces remarkably consistent levels of daily cocaine intake, whereas increased or unlimited access produces more variable patterns of self administration that may change over time. In the present study we used a discrete trials (DT) procedure to document how levels of access affect the pattern of cocaine intake. Rats that had been implanted with a chronically indwelling jugular cannula and trained to self-administer cocaine on an FR1 schedule were given access to cocaine during 10-min DT that were initiated throughout the day/night cycle for 21 days. Frequency of access (2, 3, 4 or 5 trials/h; 1.5 mg/kg/inj) and dose (1.0, 1.5, 2.0 and 2.5 mg/kg/inj, 3 trials/h) were investigated in separate groups of rats. When rats were presented with two or three trials an hour (1.5 mg/kg/inj), a highly regular and circadian pattern of intake was observed across weeks. Increasing the number of trials or increasing the unit dose resulted in a significant increase in average daily cocaine intake. Access to higher dose or higher frequency conditions produced a sustained drug taking binge during the first few days on the schedule. Rats given access to five trials per hour typically responded at every opportunity for more than 48 h, then stabilized within a range of 80-100 mg/kg/day for the remainder of the experiment. To assess whether such high levels of cocaine intake had altered the motivation to respond, cocaine reinforced break-points were assessed on a progressive ratio schedule (0.32, 1.5 and 3.0 mg/kg/inj) in separate groups of animals before and 24 h after 5 days access on the DT procedure (5 trial/h). Sustained exposure to high levels of cocaine produced a shift in the dose response curve to the right indicating tolerance to the reinforcing effects. This DT procedure provides a method to examine the behavioral and neurochemical effects of high cocaine intake over extended periods without toxicity. PMID- 12127201 TI - Effects of oral THC maintenance on smoked marijuana self-administration. AB - Studies have shown that the Delta(9)-tetrahydrocannabinol (Delta(9)-THC) concentration in marijuana cigarettes is an important factor for the maintenance of marijuana self-administration. Yet, the impact of oral Delta(9)-THC treatment on marijuana self-administration is unknown. Because other agonist therapies have been demonstrated to be effective for the treatment of substance use disorders, the objective of this study was to evaluate the influence of oral Delta(9)-THC maintenance on choice to self-administer smoked marijuana. During this 18-day residential study, 12 healthy research volunteers received one of three doses of oral Delta(9)-THC capsules (0, 10, 20 mg q.i.d.) for 3 consecutive days, followed by 3 consecutive days of matching placebo. The order of active Delta(9)-THC administration was counterbalanced. Each morning, except on days 6, 12, and 18, participants smoked the 'sample' marijuana cigarette (1.8% Delta(9)-THC w/w) and received a $2 voucher (redeemable for cash at study's end). Following the sample, volunteers participated in a four-trial choice procedure during which they had the opportunity to self-administer either the dose of marijuana they sampled that morning or to receive the $2 voucher. Relative to placebo Delta(9)-THC maintenance, participants' choice to self-administer marijuana was not significantly altered by either of the two active Delta(9)-THC maintenance conditions. Some 'positive' subjective drug-effect ratings following the sample marijuana cigarette were reduced: by day 3 of active oral Delta(9)-THC maintenance, participants' rating of 'Good Drug Effect' and 'High' were significantly decreased. Smoked marijuana-related total daily caloric intake was not significantly altered under any maintenance conditions. Finally, the effects of smoked marijuana on psychomotor task performance were only minimally affected by oral Delta(9)-THC maintenance. These data indicate that participants' choice to self-administer marijuana was unaltered by the oral Delta(9)-THC dosing regimen used in the present investigation. PMID- 12127202 TI - Discriminative-stimulus effects of modafinil in cocaine-trained humans. AB - Modafinil is a novel stimulant that is effective in the treatment of narcolepsy and excessive daytime sleepiness. In vitro and in vivo neuropharmacological data suggest that the mechanism of action of modafinil is distinct from that of prototypical abused stimulants like cocaine and d-amphetamine. In the present experiment, six human volunteers with recent histories of cocaine use learned to discriminate 150 mg oral cocaine HCL. After acquiring the discrimination (i.e. > or = 80% correct responding on 4 consecutive days), a range of doses of oral cocaine (50, 100, and 150 mg), modafinil (200, 400, and 600 mg), and placebo were tested to determine if they shared discriminative-stimulus and self-reported effects with 150 mg cocaine. Methylphenidate (60 mg) and triazolam (0.5 mg) were included as positive and negative controls, respectively. Cocaine and methylphenidate, but neither modafinil nor triazolam, produced cocaine-like discriminative-stimulus, subject-rated, and cardiovascular effects. The results of the present experiment suggest that cocaine discrimination in humans is pharmacologically specific within and across drug classes. PMID- 12127203 TI - The use of divalproex in alcohol relapse prevention: a pilot study. AB - Anticonvulsant agents show promise in the treatment of the acute symptoms of alcohol withdrawal and may also treat some symptoms associated with the protracted abstinence syndrome. Impulsivity, hostility and irritability are common characteristics of alcohol-dependent individuals, and there is some evidence that anticonvulsant agents decrease these traits in individuals with a number of different psychiatric disorders. This pilot study is a 12-week, double blind, placebo-controlled trial of an anticonvulsant agent, divalproex (DVPX), in alcohol-dependent individuals. Alcohol use (Timeline Follow Back), impulsivity (Barratt Impulsivity Scale), irritability and aggression (Buss-Durkee Hostility Index; and Anger, Irritability, Aggression Scale) were measured at baseline and throughout the 12-week treatment period. Drinking decreased significantly in both the placebo and the DVPX-treated groups. In the DVPX group, a significantly smaller percentage of individuals relapsed to heavy drinking, but there were no significant differences in other alcohol-related outcomes. There were significantly greater decreases in irritability in the DVPX-treated group and a trend towards greater decreases on measures of lability and verbal assault. There were no significant between-group differences on measures of impulsivity. While DVPX did not have a robust effect on alcohol-related outcomes, it did have modest impact on a measure of irritability. This is consistent with the findings of other investigators exploring the use of DVPX in schizophrenia, personality disorder and a number of other psychiatric disorders. PMID- 12127204 TI - Substance abuse and the need for money management assistance among psychiatric inpatients. AB - Patients who mismanage their funds may benefit from financial advice, case management or the involuntary assignment of a payee who restricts direct access to funds. Data from a survey of psychiatric inpatients at four VA hospitals (N = 236) was used to evaluate the relationship between substance abuse and clinician rated need for money management assistance. Multivariate analytic techniques were used to control for sociodemographic factors and psychopathology. Alcohol and drug use severity both were modestly associated with need for assistance. The effect of substance use severity was greater in patients who were also diagnosed with a major mental illness. Clinicians indicated that 27 patients (11% of the sample) required an involuntary payee and 21 of the 27 (78%) had a Substance Abuse diagnosis. Only drug use severity was significantly associated with need for a payee. These data describe a substantial unmet need for money management assistance in psychiatric inpatients, particularly among those with substance abuse disorders. There is a need to examine the process by which the Social Security and Veterans Benefits Administrations assign payees to determine whether patients with co-morbid substance abuse are not being assigned a payee in spite of their discernible need for one. PMID- 12127205 TI - Paediatric uroradiology-where are we and where shall we go? PMID- 12127206 TI - Future expectations--what paediatric nephrologists and urologists await from paediatric uroradiology. AB - Cooperation between paediatric nephrology/urology and paediatric radiology is essential for timely and correct diagnosis and therapy of kidney and urinary tract disorders. We need the direct contact between doctors before or after investigations, interdisciplinary discussions, and rapid access to the images. This should lead to optimal settings for investigations, a reduction of radiation burden and the number of investigations, and further improvement in the management of patients. Modern sonography including colour Doppler sonography, amplitude-coded Doppler sonography, and eventually 3D-ultrasound is and will be the method of choice as the basic, non-invasive investigation. These investigations should become the routine in all institutions. Refinement and standardisation of already established investigations are needed. Recently introduced investigations, like MRI, will have to show their impact on future diagnostic imaging. Future introduction of new non-invasive methods is welcome, e.g. to correctly diagnose vesicoureteral reflux without catheterisation/puncture of the bladder. PMID- 12127207 TI - Conventional imaging in paediatric uroradiology. AB - OBJECTIVE: To briefly describe basic conventional imaging in paediatric uroradiology. METHOD: The state of the art performance of standard imaging techniques (intravenous urography (IVU), voiding cystourethrography (VCU), and ultrasound (US)) is described, with emphasis on technical aspects, indications, and patient preparation such as adequate hydration. Only basic applications as used in routine clinical work are included. RESULT AND CONCLUSION: Conventional imaging methods are irreplaceable. They cover the majority of daily clinical routine queries, with consecutive indication of more sophisticated modalities in those patients who need additional imaging for establishing the final diagnosis or outlining therapeutic options. PMID- 12127208 TI - Potential of modern sonographic techniques in paediatric uroradiology. AB - OBJECTIVE: To describe the potential of modern sonographic techniques in paediatric uroradiology. METHOD: Ultrasound (US)-now being the primary imaging tool-has revolutionised imaging diagnostic in the urinary tract. Constant developments and technical refinements have secured the role of US in uroradiology. Colour Doppler Sonography (CDS) and innovative applications such as the transperineal approach or application of m-mode US to the urinary tract have helped to develop US from just a basic tool to a sophisticated and respected method. The ongoing introduction of new and even more sophisticated methods further enhance the sonographic potential, which shall be demonstrated by a more detailed discussion of these methods. RESULTS: Harmonic imaging, extended field of view US, amplitude coded CDS, echo-enhanced US, and three-dimensional US as the most recent new sonographic techniques are successfully applicable to paediatric urinary tract disease. They improve sonographic diagnosis in many conditions, such as detection of vesico-ureteral reflux, renal parenchymal volume assessment, comprehensive visualisation of hydronephrosis and complex pathology, evaluation of renal perfusional disturbances or defects, superior documentation with improved comparability for follow-up, or simply by offering clearer tissue delineation and differentiation. CONCLUSION: Modern US techniques are successfully applicable to neonates, infants, and children, further boosting the value of US in the paediatric urinary tract. However, as handling became more sophisticated, and artefacts have to be considered, modern urosonography became not only a more powerful, but also a more demanding method, with the need for expert knowledge and dedicated training. PMID- 12127209 TI - Vesicoureteral reflux grading in contrast-enhanced voiding urosonography. AB - INTRODUCTION AND OBJECTIVE: The sonographic diagnosis of vesicoureteral reflux (VUR) with contrast-enhanced voiding urosonography (VUS) is gradually increasing. With the introduction of VUS as part of the routine diagnostic imaging modalities for reflux significant reduction in the number of voiding cystourethrographies (VCUG) was possible. Like in VCUG grading of reflux in VUS is becoming more and more relevant. The aim of this study was to find out if there are any sonomorphologic and sonomorphometric parameters that would correlate with reflux grading in VCUG. Furthermore, a reflux grading system for VUS is proposed and the correlation of this grading system tested with the one of VCUG. PATIENTS AND METHODS: In one examination session a total of 186 children underwent both VUS and VCUG of whom 89 had VUR in at least one and the same kidney-ureter-unit (KUU) in both diagnostic imagings. The VUS was conducted with intravesical administration of ultrasound (US) contrast medium (Levovist). Ureteral and pelvicalyceal dilatations before administration of US contrast medium and during reflux were documented. Renal pelvic diameter was measured. The density of microbubbles in the renal pelves was scored on a scale of 1-3 (low to high). A grading system for reflux in VUS was set up similar to the international reflux grading system for VCUG with the addition of one more differentiation parameter, namely whether the reflux was primarily in a dilated or non-dilated urinary tract. Reflux grades in VUS were compared with those in VCUG. RESULTS: None of the sonomorphologic and sonomorphometric parameters demonstrated any clear cut finding that would simplify reflux grading in VUS. In 59/95 (62%) KUUs the reflux grades were the same in both examinations. In 10/95 (11%) and 26/95 (27%) KUUs, the reflux was graded lower or higher, respectively, in VUS than in VCUG. Fifty seven percent were in a primarily dilated system and the remaining 43% in a non dilated one. Seventy percent of KUUs diagnosed as having grade I reflux in VCUG, showed as grade 2 on VUS. CONCLUSIONS: A reflux grading system similar to the one used in VCUG can be applied in VUS. Adding the parameter reflux into a primarily dilated or non-dilated ureter and/or pelvicalyceal system may bring in a further dimension to the reflux grading in VUS. Most of the refluxes labelled as grade I in VCUG are actually grade II or higher. PMID- 12127210 TI - The role of computed tomography in modern paediatric uroradiology. AB - Computed tomography (CT) has developed a well-recognised role within paediatric uroradiology, especially in imaging of trauma, malignancy (in particular Wilms' tumour), atypical infection, and congenital urogenital abnormalities. CT can also be used for problem solving in nephrolithiasis and renal transplant assessment. These applications are illustrated and discussed, with an emphasis on particular information that can be gained from the CT study. PMID- 12127211 TI - Single- and multi-slice spiral computed tomography of the paediatric kidney. AB - Single- and multi-slice computed tomography (CT) is regarded as the primary imaging tool in traumatology, both in adults and children. For complicated infectious disease and renal tumours, these techniques are recommended only as secondary diagnostic tools. Specifically, multi-slice CT (MSCT) provides excellent spatial resolution, which is a particular advantage for the evaluation of small structures as they are typical in children. However, MSCT offers more information than is required for diagnosis. Therefore, low-dose protocols are necessary for paediatric examinations. The CT dose-index (CTDI(vol)) should not exceed 2 mGy for newborns, 4 mGy for toddlers, 5 mGy for elementary school children, and 8 mGy for adolescents. PMID- 12127212 TI - Radionuclide studies in paediatric nephro-urology. AB - The main tool of radionuclide techniques applied to paediatric uro-nephrology is the quantitation of function, which is an information not easily obtained by other diagnostic modalities. The radiation burden is low. Drug sedation is only rarely needed, whatever the age of the patient. Accurate determination of glomerular filtration rate can be obtained by means of an intravenous injection of Cr-51 EDTA and one or two blood samples. Tc-99m DMSA scintigraphy is an accurate method for evaluation of regional cortical impairment during acute pyelonephritis and later on, for detection of permanent scarring. Tc-99m MAG3 renography is nowadays a well-standardized method for accurate estimation of the split renal function and of renal drainage with or without furosemide challenge. This technique is particularly indicated in uni- or bilateral uropathies with or without renal and/or ureteral dilatation. Direct and indirect radionuclide cystography are two alternative modalities for X-ray MCUG. Their relative place in the strategy of management of vesicoureteral reflux is discussed. PMID- 12127213 TI - MR urography in children. AB - Thanks to the development of rapid sequences with better resolution, applications of uro MR have rapidly increased in children. Difficulties that remain are related to the variable ages of the patients. It is therefore mandatory to standardize as much as possible the techniques that are used in order to obtain reproducible results. In this review, the examination protocols will be explained. In a second part the current applications in children will be illustrated and discussed, especially in comparison with the other imaging techniques. PMID- 12127214 TI - Imaging guided interventional procedures in paediatric uroradiology--a case based overview. AB - OBJECTIVE: To describe the potential and application of interventional image guided procedures in the paediatric urinary tract. PATIENTS AND METHODS: The different techniques are illustrated using case reports. The examples comprise established indications such as percutaneous nephrostomy for compromised kidneys in obstructive uropathy and infection, sonographic guided renal biopsy including monitoring or treatment of complications after biopsy, and evaluation and balloon dilatation of childhood renal artery stenosis. There are new applications such as treatment of stenosis in cutaneous ureterostomy or sonographically guided catheterism for deployment of therapeutic agents. RESULTS: Generally, the procedures are safe and successful. However, complications may occur, and peri /post-interventional monitoring is mandatory to insure early detection and adequate management. Sometimes additional treatment such as percutaneous embolisation of a symptomatic post biopsy arterio-venous fistula, or a second biopsy for recurrent disease may become necessary. CONCLUSION: Imaging guided interventional procedures are performed successfully in a variety of diseases of the paediatric urinary tract. They can be considered a valuable additional modality throughout infancy and childhood. PMID- 12127215 TI - Quiz case. Congenital infantile myofibromatosis. Diaphragmatic and skeletal involvement but no visceral changes. PMID- 12127216 TI - OK-432 therapy for lymphatic malformation in 32 patients (28 children). AB - BACKGROUND/PURPOSE: Operating lymphatic malformation (LM) may lead to nerve damage with permanent cosmetic disturbance. Even sclerosants as ethanol and Sotradecol may sometimes harm more than cure. The purpose with this study was to evaluate the effect of a relatively new drug for intralesional injections, OK 432. METHODS: The diagnosis of LM was made clinically by means of ultrasound and MRT and/or CT. Thirty-two patients (28 children) with LM were consecutively enrolled in the study. Twenty-nine (27 children) had not been treated previously: 17 (15 children) had macrocysts (MAC), four microcysts (MIC) and eight had combined cysts (CC). Three patients (one child) had got previous treatment without any curative effect. All patients got intralesional injections with OK 432 at intervals according to a previously published protocol (Lakartidningen, 95 (1998) 2074). RESULTS: No serious adverse effects were seen. The results obtained were excellent in all with macrocysts but in one, who was pretreated with ethanol, where no LM-regression was seen. None of four with MIC-LM required further therapy; for two of them the results were excellent. Of 10 with CC, seven showed excellent results. Only one required surgery. CONCLUSION: OK-432 is effective and is proposed to be the first choice of treatment for LM. PMID- 12127217 TI - Nasal resistances are useful in identifying children with severe obstructive sleep apnea before polysomnography. AB - OBJECTIVE: In this study, we would like to show that anterior rhinometry measurement of nasal resistance would be a simple and useful test to identify severe obstructive sleep apnea (OSA) in a population of children affected by adenotonsillar hypertrophy. METHODS: Seventy-three consecutive children (44 males; mean age 5.4+/-1.2 years) with adenotonsillar hypertrophy, who complained sleep-disordered breathing, were studied. All the parents completed a questionnaire concerning the children's sleeping habits and sleep complaints before consultation; each child underwent a general paediatric examination and an evaluation of craniofacial features and upper airway patency. In all 73 children polysomnography was performed and anterior rhinometry nasal patency was measured. RESULTS: The diagnosis of OSA was confirmed in 44/73 patients (60%). Total nasal resistance showed a significant direct correlation with apnea hypopnea index, arousal index, snoring time, percentage of sleep time spent at SaO(2)<90% and a significant inverse correlation with total sleep time, sleep efficiency and the mean of SaO(2)% during sleep. Total nasal resistance was significantly related to snoring, mouth breathing and daytime sleepiness. The receiver operator characteristics (ROC) curve indicates that in the range of age of our sample a nasal resistance value of 0.59 Pa/cm(3)/s has a sensitivity of 91% and specificity of 96% for identifying the children with adenotonsillar hypertrophy affected by OSA. CONCLUSIONS: Our study shows that in children with adenotonsillar hypertrophy nasal resistance seems to be risk factor for OSA. The anterior rhinometry appears as a useful tool in routine evaluation of sleep disordered breathing in these patients. PMID- 12127218 TI - Language development in hearing-impaired children. Establishment of a reference material for a 'Language test for hearing-impaired children', LATHIC. AB - OBJECTIVE: In Sweden, there has previously been no normalised test material for the evaluation of language development in individual hearing-impaired children, and for the assessment of various methods of auditory habilitation. The purpose of the present study was to compose, apply and evaluate a test for language development in hearing-impaired children, and to establish the first set of reference values related to age, sex, type and degree of hearing impairment. METHODS: A test consisting of nine subtests was assembled and developed for, and subsequently applied to, hearing-impaired children in the age range 4-6 years. The inclusion criteria were a pure tone average of 80 dBHL or less and oral language (Swedish) as the first language. Two hundred and eleven hearing-impaired children and 87 normal hearing control children were tested. RESULTS: The results show that: (1) children with hearing impairment-also unilateral-have a delayed language development; (2) the delay is greater in children with larger losses and tends to decrease with increasing age; (3) 6-year-olds with hearing loss greater than 60 dB have not reached the level of the control group; (4) no difference between right- or left sided deafness with respect to language development was observed; (5) a reference material, applicable during clinical assessment, was established for the most common types of hearing impairment. CONCLUSIONS: The test designed gave graded measures of important aspects of language development in hearing-impaired children. The results merit further application of the test material. PMID- 12127219 TI - Cine magnetic resonance imaging for evaluation of focal tracheomalacia: innominate artery compression syndrome. AB - BACKGROUND: The contribution of an 'aberrant innominate artery' to respiratory distress syndromes has been a matter of debate nearly since the introduction of this concept. Recent advances in dynamic imaging are proving to be of value in assessing tracheal function in patients with respiratory distress. We therefore evaluated patients with innominate artery compression syndrome using the cine magnetic resonance imaging (CMRI) modality. OBJECTIVES: To apply the CMRI modality to evaluate patients with respiratory distress who exhibited tracheal compression at the level of the innominate artery. METHODS: A cohort of three patients in respiratory distress underwent bronchoscopy, followed by CMRI using a Siemens 1.5T Vision system. RESULTS: These three patients exhibited tracheal compression at the level of the innominate artery in agreement with their findings during bronchoscopy. All three exhibited dynamic tracheal compression that varied with the respiratory cycle. The degree of tracheal compromise was readily appreciated using the dynamic, real-time CMRI modality. Due to the severity of symptoms, the two children underwent innominate arteriopexy with complete resolution of their symptoms. CONCLUSIONS: CMRI provides extremely rapid acquisition of images, as well as integrated information regarding relationships of mediastinal structures. By providing functional imaging of tracheal patency during the respiratory cycle, CMRI may provide additional insight into innominate artery compression syndrome as more patients are evaluated. PMID- 12127220 TI - Spontaneous resolution of recurrent tonsillitis in pediatric patients on the surgical waiting list. AB - OBJECTIVE: To assess the impact of the waiting list on the spontaneous resolution of recurrent acute tonsillitis in children. METHODS: We have evaluated 623 cases placed on the waiting list for elective tonsillectomy (with or without adenoidectomy) between February 1994 and May 1999 at our institution. In each child, age, time on the waiting list, type of procedure and outcome were registered. There were two possible outcomes after the preoperative evaluation: tonsillectomy was still indicated or tonsillectomy was not necessary. RESULTS: Mean length of time on the waiting list was 10.8 months (range: 3.0-35.6 months; median: 8.2 months). In 507 of the 623 children (81.4%), the operation was still indicated. However, 116 patients (18.6%) did not need surgery because of spontaneous resolution of the clinical picture. No relation was found between outcome and age, time on the waiting list or type of procedure (P>0.05). CONCLUSION: There was no clinical evidence for claiming that resolution of recurrent acute tonsillitis in children is spontaneous with time. PMID- 12127221 TI - Infrared tympanic thermometer can accurately measure the body temperature in children in an emergency room setting. AB - OBJECTIVE: The objective in this study was to compare the accuracy of the tympanic membrane infrared thermometer with the other conventional temperature measurement options. METHODS: One hundred and ten randomly selected pediatric patients who admitted to our emergency room were included in the study. Each child underwent simultaneous temperature measurement via rectum, axilla, and external auditory canal. The rectal and axillary measurements were performed using conventional mercury in glass thermometers. The aural measurement was performed using the non-contact infrared thermometer (Braun ThermoScan IRT 1020, Germany). RESULTS: On aural measurement, the results of both ears as well as the first, second and third measurements were similar (P<0.01). The mean results of the axillary, rectal and tympanic temperature measurements were 37.46+/-1, 38.18+/-1, and 38.01+/-1.1, respectively. The mean axillary temperature was 0.72 degrees C lower than the mean rectal temperature, and 0.55 degrees C lower than the tympanic temperature. The difference between the mean tympanic and rectal temperatures was 0.17 degrees C. The results of measurements via rectum, axilla and ear were similar (P<0.01). CONCLUSION: In conclusion, it is apparent that each of the temperature measurement options has some advantages and disadvantages. An optimal thermometer should have the following features; accurate temperature measurement; ease of application in a short while; safety and absence of potential risks; and tolerability by the patient. Since the aural infrared thermometer meets these criteria, its use in the routine clinical practice appears to be advantageous rather than or complementary to the conventional methods. PMID- 12127223 TI - Bilateral congenital choanal atresia in a 13-year-old patient. AB - Congenital choanal atresia (CCA) is the developmental failure of the nasal cavity to communicate with the nasopharynx. Surgical repair is recommended in the first weeks of life in bilateral cases, because in newborns this is a life-threatening situation. This is a case report of a 13-year-old patient complaining of long term bilateral nasal obstruction and rhinorrea, in whom bilateral choanal atresia was diagnosed by endoscopic exploration and CT scan, and who was treated by an endonasal endoscopic surgical technique. Bilateral choanal atresia is a life threatening disease in newborns; however, it can be diagnosed in adults with bilateral nasal obstruction and rhinorrea. PMID- 12127222 TI - Robin sequence: review of treatment modalities for airway obstruction in 110 cases. AB - OBJECTIVE: The objective of this paper is to assess the treatment protocols used in our institution for the management of airway obstruction among patients with Robin sequence and to suggest a rationale for management based on the findings. METHOD: A retrospective study of 110 children with Robin sequence seen from 1988 to 1997 at the Chang Gung Memorial Hospital was done. Results of the management in airway obstruction and feeding difficulty were studied. RESULTS: Prone posturing was effective in the treatment of mild airway obstruction in 82 patients who had noisy breathing sounds. Twenty-eight infants required endotracheal intubation due to severe respiratory distress. Seven received a tongue to lip adhesion (TLA) operation. Three of these patients showed a relief of airway obstruction, while four of them needed a tracheotomy to maintain a patent airway because of wound dehiscence. Two other patients underwent tracheotomy without TLA. All the six patients who received tracheotomy were eventually decannulated. A nasopharyngeal tube was inserted in two other patients, and one of them improved only temporarily. With regards to feeding difficulties, 46 patients needed nasogastric tube feeding, while the rest were aided through the use of a cleft palate bottle and nipple. A gastrostomy was not needed in any of the patients in this series. CONCLUSION: The morbidity and mortality among Robin sequence patients had been widely decreased through the teamwork of the pediatrician, anesthesiologist, otolaryngologist, dentist, and plastic surgeon. Based on our experience, conservative management was sufficient for those patients with mild airway obstruction, while endotracheal intubation was required for patients with cyanosis, respiratory failure and sleep apnea. Tracheotomy was a better option than TLA for airway management among patients with failed extubation. PMID- 12127224 TI - Tuberculous mastoiditis: when is surgery indicated? AB - Mycobacterium tuberculosis is a rare cause of mastoiditis, but diagnosis is often delayed, with potentially serious results. We present the case of a 7-year-old child who failed to improve even once the diagnosis was made and appropriate medical treatment initiated. At mastoidectomy, a bony sequestrum was found which had not been evident on CT scanning. We review the diagnosis and management of this condition and suggest that failure to respond to drug therapy even in the absence of demonstrable complications be added to the list of indications for surgical intervention. PMID- 12127225 TI - Tourette syndrome manifest as chronic cough. AB - Gilles de la Tourette syndrome (TS) is a neurobehavioral disorder characterized by the presence of fluctuating involuntary motor and vocal tics. We report the case of a child in whom TS manifest as an involuntary recurrent complex tic presenting as a chronic cough. The clinical and pathological features and management of TS are reviewed. PMID- 12127226 TI - McCune-Albright syndrome: a case report and review of the literature. AB - Nasal obstruction is one of the most common symptoms pediatric patients present to their physicians with. Usually this symptom is caused by allergic rhinitis or an upper respiratory infection, however, many conditions present with nasal obstruction. We present a 14-year-old patient with a rare cause of nasal obstruction, turbinate enlargement due to fibrous dysplasia, as diagnosed by CT scan and histopathologic analysis. Additionally, clinical history revealed precocious puberty, and a dermatologic exam revealed a cafe au lait macule, consistent with the McCune-Albright syndrome. This case illustrates the need to formulate a broad differential diagnosis and also consider the overall patient, not just the nasal cavity. The clinical presentation, diagnostic evaluation, long term management plan and relevant literature are discussed. PMID- 12127227 TI - A mild case of frontonasal dysplasia: the rhinologic perspective. AB - Frontonasal dysplasia (FND) is a congenital malformation characterized by hypertelorism, broad nasion with a midline cleft in the bony dorsum, midline defect of the frontal bone, absence of the nasal tip, and deformities in the nasal alar region. The clinician should be aware of the mild forms of FND. We presented absence of crista galli in a mild case of FND. Computed tomography scanning should assess the facial bones, nose, and paranasal structures. If a surgical correction is planned, this complete work-up prevents unexpected complications and complements the evaluation of paranasal deformities. PMID- 12127229 TI - Nootropic activity of Albizzia lebbeck in mice. AB - The effect of saponin containing n-butanolic fraction (BF) extracted from dried leaves of Albizzia lebbeck on learning and memory was studied in albino mice using passive shock avoidance paradigm and the elevated plus maze. Significant improvement was observed in the retention ability of the normal and amnesic mice as compared to their respective controls. We have also studied the effects of BF on the behavior influenced by serotonin (5-HT), noradrenaline and dopamine. The brain levels of serotonin, gamma-aminobutyric acid (GABA) and dopamine were also estimated to correlate the behavior with neurotransmitter levels. The brain concentrations of GABA and dopamine were decreased, whereas the 5-HT level was increased. The data indicate the involvement of monoamine neurotransmitters in the nootropic action of BF of A. lebbeck. PMID- 12127230 TI - Medicinal plants of the Popoluca, Mexico: organoleptic properties as indigenous selection criteria. AB - The taste and smell of the environment are important to humans in everyday life and are of particular relevance for the selection of medicinal versus non medicinal plant species. In a 16-months study with the Popoluca of southern Veracruz (Mexico), we focused on the indigenous selection criteria for medicinal plants. We provide evidence for a highly significant association between organoleptic properties of plants and the use of these species as medicine. Additionally, the doctrine of signature is an essential mnemonic aid, which facilitates remembering the use assigned to the plant. From the Popoluca point of view, it is essential to find substitutes or alternative treatments when a certain species is not at hand. We show that organoleptic properties and the doctrine of signature are excellent guides for selecting or memorising such medicinals. PMID- 12127231 TI - Glucose lowering effect of aqueous extract of Enicostemma littorale Blume in diabetes: a possible mechanism of action. AB - Diabetes mellitus is a chronic metabolic disorder characterized by hyperglycemia. Enicostemma littorale Blume is a small herb and recently we have reported its blood glucose lowering potential in alloxan induced diabetic rats. A single dose of aqueous extract of E. littorale (15 g dry plant equivalent extract per kg) had shown significant increase in the serum insulin levels in alloxan-induced diabetic rats at 8 h. The insulinotropic action of aqueous extract of E. littorale was further investigated using rat pancreatic islets. Extract has the potential to enhance glucose-induced insulin release at 11.1 mM glucose from isolated rat pancreatic islets and was partially able to reverse the effect of diazoxide (0.25 mM). Incubation with Ca(2+) chelator (EGTA) and Ca(2+) channel blocker (nimodipine) did not affect the glucose-induced insulin release augmented by the extract. Above results suggest the glucose lowering effect of aqueous extract of E. littorale to be associated with potentiation of glucose-induced insulin release through K(+)-ATP channel dependent pathway but did not require Ca(2+) influx. PMID- 12127232 TI - Tranquilizing, antihistaminic and purgative activity of Nyctanthes arbor tristis leaf extract. AB - The decoction of the leaves of Nyctanthes arbor tristis Linn. (Harsingar) is widely used in Ayurvedic system of medicine for the treatment of sciatica, arthritis, fevers, various painful conditions and as laxative. In the present investigation, the water soluble portion of the alcoholic extract of the leaves was screened for some CNS activities (viz. hypnotic, tranquilizing, local anaesthetic, hypothermic, anticonvulsant), antihistaminic and purgative activities. The extract produced general depression of spontaneous motor activity, significantly increased pentobarbitone sleeping time though it had no effect on righting reflex. Furthermore higher doses of the extract abolished CAR without affecting motor coordination. Moreover the extract exhibited hypothermic effect and protected guinea pigs from histamine aerosol. These activities are common to major tranquilizers and support the usage of the plant by Ayurvedic physicians in aforementioned conditions. In addition significant purgative activity was also exhibited by the extract. PMID- 12127233 TI - Inhibition of cancer cell growth by crude extract and the phenolics of Terminalia chebula retz. fruit. AB - A 70% methanol extract of Terminalia chebula fruit, was studied for its effects on growth in several malignant cell lines including a human (MCF-7) and mouse (S115) breast cancer cell line, a human osteosarcoma cell line (HOS-1), a human prostate cancer cell line (PC-3) and a non-tumorigenic, immortalized human prostate cell line (PNT1A) using assays for proliferation ([(3)H]-thymidine incorporation and coulter counting), cell viability (ATP determination) and cell death (flow cytometry and Hoechst DNA staining). In all cell lines studied, the extract decreased cell viability, inhibited cell proliferation, and induced cell death in a dose dependent manner. Flow cytometry and other analyses showed that some apoptosis was induced by the extract at lower concentrations, but at higher concentrations, necrosis was the major mechanism of cell death. ATP assay guided chromatographic fractionation of the extract yielded ellagic acid, 2,4-chebulyl beta-D-glucopyranose (a new natural product), and chebulinic acid which were tested by ATP assay on HOS-1 cell line in comparison to three known antigrowth phenolics of Terminalia, gallic acid, ethyl gallate, luteolin, and tannic acid. Chebulinic acid (IC(50) = 53.2 microM +/- 0.16) > tannic acid (IC(50) = 59.0 microg/ml +/- 0.19) > and ellagic acid (IC(50) = 78.5 microM +/- 0.24), were the most growth inhibitory phenolics of T. chebula fruit in our study. PMID- 12127234 TI - Anticancer and antithrombin activity of Russian plants. AB - A chromogenic bioassay was utilized to determine the antithrombin activity of the methylene chloride and methanol extracts prepared from forty-five plants of Russia. Mouse leukemia cells (L1210) were utilized to screen these extracts for activity against cancer. The results indicated that eight plant extracts demonstrated 90% or higher activity in the inhibition of thrombin. Also, nine methanol extracts demonstrated activity of 90% or higher in the inhibition of mouse leukemia L1210 cells. The methanol extracts of Quercus robur and Sanguisorba officinalis demonstrated high activity against both thrombin and cancer. PMID- 12127235 TI - Antidiabetic and hypolipidemic activity of Helicteres isora in animal models. AB - Helicteres isora (Sterculiaceae) root juice has been used in the treatment of diabetes by several ethnic groups in different parts of India. A program was initiated to elucidate the scientific basis for the antidiabetic activity of H. isora. Ethanolic extract of H. isora root caused significant reduction in plasma glucose, triglyceride and insulin levels at 300 mg/kg dose after 9 days of administration to insulin resistant and diabetic C57BL/KsJdb/db mice. In normoglycemic and mildly hypertriglyceridemic Swiss albino mice, the extract also showed significant reduction in plasma triglyceride and insulin levels, without affecting plasma glucose level. An ethanolic extract showed activity distinctly different from glybenclamide and acarbose but similar to troglitazone in these models. In high fat fed hamster model, the extract showed significant reduction in plasma lipid levels. In order to identify the active pharmacophore, the ethanolic extract was further subjected to sequential partitioning with low, medium and high polarity solvents, which yielded a semipurified fraction having both euglycemic and lipid-lowering activity. Our study suggests that the extract of H. isora has insulin-sensitizing and hypolipidemic activity and has the potential for use in the treatment of type-2 diabetes. PMID- 12127236 TI - Hypoglycaemic effect of Rubus fructicosis L. and Globularia alypum L. in normal and streptozotocin-induced diabetic rats. AB - The present study investigates the effect of oral administration of the aqueous extract of Rubus fructicosis L. (RF) and Globularia alypum L. (GA) leaves on blood glucose levels in normal and streptozotocin (STZ) diabetic rats. In normal rats, single and repeated oral administration of RF lowered significantly the blood glucose levels, while, GA treatment did not change blood glucose levels. In STZ rats, single and repeated oral administration of both RF and GA produced significant decrease of blood glucose levels. RF and GA treatments did not affect insulin secretion both in normal and STZ rats, indicating that mechanism(s) by which these plants decrease blood glucose levels is extra-pancreatic at least, for the doses used. In addition, the acute toxicity study revealed that the aqueous extracts may be considered relatively safe since that the LD(50) value was over 8.1 and 14.5 g/kg for RF and GA respectively. These findings indicate that RF and GA represent an effective blood glucose lowering and a potential source for discovery of new orally active component(s) for future therapy. PMID- 12127237 TI - Cancer prevention and therapy with kiwifruit in Chinese folklore medicine: a study of kiwifruit extracts. AB - Kiwi gold fruits were extracted successively with hexane, acetone, methanol and 70% methanol, and further fractionated by silica gel and ODS column chromatographies for the assays of various biological activities. Five fractions H1, H2 (hexane extract), Al, A2 (acetone extract) and M2 (methanol extract) showed selective cytotoxic activity against human oral tumor cell lines, which was more sensitive than human gingival fibroblasts. More hydrophilic fractions [70M3, 70M4, 70M5] of 70% methanol extract displayed higher anti-HIV activity, radical generation and O2- scavenging activity. The antibacterial activity of 70% methanol extracts [70M0, 70M1, 70M2, 70M3, 70M4] was generally lower than that of more lipophilic fractions (hexane, acetone, methanol extracts), although each fraction did not show any specific antimicrobial action. All fractions were inactive against Helicobacter pylori. These results demonstrate that gold kiwifruit extracts contain valuable, various bioactive materials, which can be separated with each other. PMID- 12127238 TI - Augmentation of immune cell activity against tumor cells by Rauwolfia radix. AB - In this study, we investigated the effect of Rauwolfia radix on heat shock protein (HSP) 70 expression and cytotoxicity against tumor cells in activated human T cells. When activated T cells were cultured with Rauwolfia radix for 18 h, HSP70 expression after heat shock was remarkably increased, and cytotoxicity against T98G tumor cells was augmented. Moreover, Rauwolfia radix also enhanced the cytotoxicity of heat shocked activated T cells against Molt-4 and T98G tumor cells. Secretions of interferon-gamma (IFN-gamma) and tumor necrosis alpha (TNF alpha), due to Concanavalin A (Con A) stimulation, were increased by Rauwolfia radix in activated T cells. To investigate the antitumor effect in vivo, EL-4 tumor-bearing mice were administered with Rauwolfia radix in drinking water. The survival period of the Rauwolfia radix treatment group was significantly prolonged compared with that of the control group. Reserpine, the major active ingredient of Rauwolfia radix, also enhanced the cytotoxicity of activated T cells against Molt-4 and T98G tumor cells, and prolonged the survival period of EL-4 tumor-bearing mice. Taken together, our results suggest that Rauwolfia radix can enhance the activity of immune cells against tumor cells. PMID- 12127239 TI - Anti-angiogenic activities of Cnidium officinale Makino and Tabanus bovinus. AB - This study investigated the anti-angiogenic activities of Cnidium officinale Makino and Tabanus bovinus by using cultured glomerular capillary endothelial cells (GECs), chorioallantoic membrane (CAM) and rat cornea. Treatment of GECs with several concentrations (5-50 microg/ml) of C. officinale Makino and T. bovinus extracts for 24 h inhibited angiotensin II (10(-8) M)-induced increases of [3H]thymidine uptake and cell numbers in a concentration-dependent manner. The extent of inhibitory rate of [3H]thymidine incorporation by C. officinale Makino and T. bovinus at 50 microg/ml was a similar to that by 10(-5) M of retinoic acid. Herbal extracts also conspicuously inhibited the neovascularization. In contrast to the normal branching of vascular vessels, blood vessel patterns in CAMs treated with extracts (50 microg per egg) of C. officinale Makino and T. bovinus were ran parallel to each other without much branching. Moreover, oral administration of herbal extracts (20 mg/kg per day) for 4 weeks significantly inhibited the rat corneal neovascularization induced by suture, and the length of blood vessels in herbal medicine-treated rat cornea was conspicuously lower than that in control animals. A similar inhibitory effect to these was also observed in the rat cornea treated with thalidomide (200 mg/kg per day). These findings indicate that the anti-angiogenic properties of C. officinale Makino and T. bovinus may be one of the pharmacological mechanisms underlying the anti-tumor and anti-metastatic activities of herbal extracts tested in this study. PMID- 12127240 TI - Antimutagenic potential of homoisoflavonoids from Muscari racemosum. AB - The potential antimutagenic effect of the plant extract of Muscari racemosum bulbs, rich on 3-benzylidene-4-chromanones, was evaluated on three genetic model organisms. The mixture of three homoisoflavonoids was applied together with diagnostic mutagens in the Ames assay on four bacterial strains Salmonella typhimurium TA97, TA98, TA100, TA102, in the toxicity and mutagenicity/antimutagenicity assay on the yeast strain Saccharomyces cerevisiae D7, and in the simultaneous phytotoxicity and clastogenicity/anticlastogenicity assay on Vicia sativa (L.). The extract exerted antimutagenic and anticlastogenic effects due to the presence of homoisoflavonoids, which may be included in the group of natural antimutagens. This genotoxicological study suggests that homoisoflavonoids from M. racemosum (L.) owing to antimutagenic and anticlastogenic properties are of great pharmacological importance, and might be beneficial for prevention of cancer. PMID- 12127241 TI - Antioxidant and antihepatotoxic activities of Phellinus rimosus (Berk) Pilat. AB - The antioxidant and antihepatotoxic activities of a wood inhabiting macrofungus, Phellinus rimosus were studied. The superoxide anion scavenging, Fe(2+)-ascorbate induced lipid peroxidation inhibiting, hydroxyl radical scavenging and nitric oxide scavenging activities of the ethyl acetate extract were determined. The results indicated that ethyl acetate extract of P. rimosus exhibited significant in vitro antioxidant activity. The ethyl acetate extract of P. rimosus also showed potent antihepatotoxic activity against carbontetrachloride-induced acute toxicity in rat liver. The amelioration of liver toxicity by the ethyl acetate extract was evident from its significant effect on the levels of serum glutamate pyruvate transaminase (SGPT), serum glutamate oxaloacetate transaminase (SGOT) and serum alkaline phosphatase (ALP). The results suggest that hepatoprotective effect of P. rimosus is possibly related to the free radical scavenging activity. PMID- 12127242 TI - Gastroprotective effect of fenugreek seeds (Trigonella foenum graecum) on experimental gastric ulcer in rats. AB - The effect of fenugreek seeds (Trigonella foenum graecum) compared to omeprazole was studied on ethanol-induced gastric ulcer. The aqueous extract and a gel fraction isolated from the seeds showed significant ulcer protective effects. The cytoprotective effect of the seeds seemed to be not only due to the anti secretory action but also to the effects on mucosal glycoproteins. The fenugreek seeds also prevented the rise in lipid peroxidation induced by ethanol presumably by enhancing antioxidant potential of the gastric mucosa thereby lowering mucosal injury. Histological studies revealed that the soluble gel fraction derived from the seeds was more effective than omeprazole in preventing lesion formation. These observations show that fenugreek seeds possess antiulcer potential. PMID- 12127244 TI - Concerning kif, a Cannabis sativa L. preparation smoked in the Rif mountains of northern Morocco. AB - The aim of the present paper is to present information about a kif preparation smoked by the Moroccan population. Results are considered as an advance of our actual investigations undertaken in the Rif zone to observe an improvement in night vision after smoking kif [Ethan et al., 2002. International Cannabinoid Research Society (ICRS) meeting, in preparation]. PMID- 12127243 TI - In vitro antiplasmodial activity of extracts of Alchornea cordifolia and identification of an active constituent: ellagic acid. AB - Extracts of leaves of Alchornea cordifolia were studied for their antiplasmodial activities. Chloroformic and ether extracts were found to be inactive while the ethanolic extract exhibited mild in vitro activity against Plasmodium falciparum. Fractionation of this extract led us to isolate ellagic acid as the active constituent of the extract with IC(50) in the range of 0.2-0.5 microM. Cytotoxicity of ethanolic fraction and ellagic acid was also estimated on human fibroblasts cells (IC(50) on Hela cells = 7.3 microM at 24 h for ellagic acid). PMID- 12127245 TI - The use of commercial saponin from Quillaja saponaria bark as a natural larvicidal agent against Aedes aegypti and Culex pipiens. AB - The larvicidal activity of commercial bark saponin extract (Sigma) from Quillaja saponaria was studied on 3rd-4th instar larvae of Aedes aegypti and Culex pipiens (vectors for dengue fever and Western Nile virus, respectively). The larvae were exposed to serial concentrations (1000, 800, 500, 300, 100, 10, 1, 0.1 and 0.01 mg/l) of the extract for 1, 3, 5, 7 and 11 days. The results indicate that commercial bark saponin is toxic, causing 100% larval mortality in A. aegypti and C. pipiens after 1 and 5 days at a dosage of 800 and 1000 mg/l, respectively. Interestingly, while bark saponins had a toxic effect on larvae there was no effect on egg hatchability in either species. The results obtained suggest that, in addition to their known activities, saponins can also serve as natural larvicidal compounds. PMID- 12127250 TI - An analysis of the performance characteristics of serological tests for the diagnosis of Neospora caninum infection in cattle. AB - AIMS: To analyse the performance characteristics (sensitivity/specificity) of two enzyme-linked immunosorbent assays (ELISA) against the indirect fluorescent antibody test (IFAT). METHODS: A total of 1199 sera were tested in two ELISAs and the IFAT and results analysed utilising software that performed a receiver operating characteristic (ROC) analysis. RESULTS: Sensitivity and specificity for the two ELISAs were calculated for a range of different cut-offs. Minimal misclassification was achieved at cut-offs that, in the case of the Central Animal Health Laboratory (CAHL)-ELISA were in line with previously published cut off values. In the case of the IDEXX-ELISA lower cut-off values than suggested by the manufacturer were calculated. CONCLUSIONS: ROC-analysis resulted in optimised, as compared to the IFAT, cut-off thresholds for the CAHL and IDEXX ELISA which can be further adjusted depending on the purpose of the investigation. PMID- 12127249 TI - Heterologous antibodies to evaluate the kinetics of the humoral immune response in dogs experimentally infected with Toxoplasma gondii RH strain. AB - An IgM capture ELISA using heterologous antibodies was developed to evaluate the kinetics of the humoral immune response in dogs experimentally infected with Toxoplasma gondii RH strain. Detection of parasite in tissues from inoculated dogs was evaluated by mouse bioassay and immunohistochemical techniques. Serum samples were obtained at regular intervals up to 62 days post-inoculation (p.i.), when the animals were necropsied and their tissues examined. Antibody levels were measured by IgM capture ELISA (McELISA), indirect hemagglutination (IHA), indirect fluorescent antibody test (IgG-IFAT) and indirect immunoenzymatic assay (IgG-ELISA). All dogs seroconverted but only one exhibited severe clinical signs of infection. IgM antibodies were detected by McELISA from the seventh day on, with decreasing IgM levels around the 27th day. Similar results were obtained from IHA, although McELISA showed earlier and longer detection of IgM antibodies. IgG antibodies were detected from the seventh day on, and throughout the period of observation. Immunohistochemical findings and mouse bioassay revealed the presence of free tachyzoites in tissues of the clinically affected dog only. These results suggest that T. gondii acute infection in dogs shows a remarkably transient IgM synthesis, and this feature may constitute an important marker of active infection. Furthermore, McELISA was shown to be a potential tool to diagnose canine toxoplasmosis. PMID- 12127251 TI - A longitudinal study of gastrointestinal parasites in Canadian dairy farms. The value of an indirect Ostertagia ostertagi ELISA as a monitoring tool. AB - The general objective of this study was to evaluate a crude Ostertagia ostertagi antigen enzyme-linked immunosorbent assay (ELISA) for monitoring gastrointestinal parasites in lactating dairy cattle. A longitudinal study of gastrointestinal parasites in lactating dairy cows was carried out in 38 herds in four provinces of Canada (Prince Edward Island, Quebec, Ontario and Saskatchewan) from September 1999 to October 2000. Bulk tank milk, cow milk, serum and fecal samples were collected monthly or quarterly from all these farms. Information on herd management factors was collected by a standard questionnaire and individual cow production data were obtained from an electronic database. The overall mean optical density ratio (ODR) was 0.30 and ranged from -0.05 to 1.55. Although a clear seasonal pattern was not observed, the ODR values tended to decrease during the housing period and start increasing in the spring before the cows went out to pasture. The second and third or greater lactation cows had significantly higher ODR values compared with first lactation animals. The individual cow ODR had a very low correlation with individual squared root fecal egg counts but showed a reasonably high correlation when herd averages values were computed (r=0.73). A moderate correlation (r approximately 0.50) between the bulk tank and herd average ODR was observed. Milk yield was negatively associated with individual cow milk ODR and a quadratic effect on ODR was observed for days in milk. Twenty eight of the herds participated in a clinical trial of eprinomectin (Ivomec Eprinex) treatment at calving. The cow level ODR values determined late in the previous lactation had a marginally significant effect (P=0.07) on treatment response, suggesting that high OD cows responded better to the anthelmintic treatment. However, because of the small sample size available in this model, more research is needed to better understand this relationship. In conclusion, the indirect ELISA using an O. ostertagi crude antigen appears useful as a technique for monitoring gastrointestinal parasite burdens in adult dairy cows and holds promise as a potential predictor of response to anthelmintic treatment. PMID- 12127252 TI - Effect of gastrointestinal nematode and liver fluke infections on weight gain and reproductive performance of beef heifers. AB - Spring born, crossbred beef heifers (n=372) were utilized over four years to measure reductions in body weights, reproductive performance and calf weights caused by gastrointestinal nematodes (primarily Ostertagia ostertagi) and the bovine liver fluke (Fasciola hepatica) and to differentiate losses attributable to each type of parasitism. Each year, weaned heifers were allotted to one of the four treatment regimens: Group 1, untreated controls; Group 2, treated for nematodes; Group 3, treated for liver fluke; and Group 4, treated for both nematodes and liver fluke. Nematodes were controlled with subcutaneous injections of either ivermectin (Ivomec, Merial) or doramectin (Dectomax, Pfizer), both at the recommended dose of 200 ug/kg bodyweight. Clorsulon (Curatrem, Merial) drench was given at the recommended rate of 7 mg/kg bodyweight to control flukes. Treatments and fecal collections were initiated at allotment each year and were repeated at 28-84-day intervals until palpation for pregnancy diagnosis. Open heifers were removed from the study at this time. Treatment dates were based on expected length of treatment efficacy, the stage of growth of the heifers and the seasonal risk of infection by the parasites. Pregnant females were pooled and received their assigned treatments prior to their calving and breeding seasons and remained together until their calves were weaned. Heifers treated for nematode infections were heavier and had higher condition scores (P<0.01) than untreated control heifers at initiation of breeding, and maintained that difference through pregnancy diagnosis. Liver fluke infection did not affect heifer gains or condition scores prior to palpation (P<0.01). At palpation, heifers treated for both forms of parasitism had the highest condition scores and weight gains (P<0.01), and also higher pregnancy rates than control heifers and heifers treated for nematodes only (P<0.01). Pregnancy rates for heifers treated for flukes only were not significantly different from those treated for both nematodes and flukes. Heifers treated for nematodes weaned heavier calves than those not treated for nematodes (P<0.05). PMID- 12127253 TI - Canine spirocercosis: clinical, diagnostic, pathologic, and epidemiologic characteristics. AB - The nematode Spirocerca lupi is a parasite of dogs with beetles of several species serving as intermediate hosts. The medical records of 50 dogs diagnosed with spirocercosis at the Hebrew University Veterinary Teaching Hospital (HUVTH) in Israel during 1991-1999 were retrospectively reviewed and compared to a control group (n=100). There was a seven-fold increase in the annual number of dogs diagnosed with spirocercosis during these years while the hospital caseload increased by 80%, indicating an emerging outbreak of this infection. Dogs from the greater Tel Aviv area were at the highest risk of being diagnosed with spirocercosis with 74% of the cases originating from this region compared to only 17% of the controls. The disease appeared to have a primarily urban pattern of distribution with a significantly higher percentage (P=0.025) of dogs from cities versus rural areas, as compared to the control group. Sixty-two percent of the cases were diagnosed during the colder months of December through April. The median age of infected dogs was 5 years, with dogs 1 year old or younger at the lowest risk of being diagnosed with spirocercosis. Large breeds were at a higher risk of infection in comparison to small breeds and the Labrador Retriever was significantly over represented (P=0.027) in the study group compared to the control population. The most common signs were vomiting or regurgitation (60%), pyrexia (24%), weakness (22%), respiratory abnormalities (20%), anorexia (18%), melena (18%) and paraparesis (14%). A caudal esophageal mass was identified by radiography in 53% of the dogs and spondylitis of the thoracic vertebrae in 33%. Fecal flotation was positive for S. lupi eggs in 80% of the dogs, and endoscopy was found to be the most sensitive diagnostic procedure and allowed diagnosis in 100% of the examined dogs. Fifty-three percent of the dogs were anemic and creatine kinase (CK) activities were elevated in 54%. Necropsy of 14 dogs revealed esophageal or gastric granulomas in 13 dogs, and an esophageal osteosarcoma in a single animal. Aortic aneurysms were found in six (43%) dogs. Out of 24, 15 dogs (63%) for which follow-up information was available died or were euthanized within 1 month of admission. The case-fatality rate decreased toward the end of the study period when improved therapy with avermectins became available. PMID- 12127254 TI - Mast cell and eosinophil mucosal responses in the large intestine of horses naturally infected with cyathostomes. AB - From December 1998 to March 2000, caecum and ascendant colon of 42 horses naturally infected with cyathostomes were collected during routine necropsy or from a local slaughterhouse. Changes in the numbers of mucosal and submucosal mast cells (MMC and SMMC), intraepithelial, mucosal and submucosal eosinophils (IE, ME and SME) in the large intestine were investigated by histochemical techniques in relation to the worm burdens. The effect of age was examined in three subgroups: 6-24-month-old horses (group 1), 2-10-year-old horses (group 2) and horses more than 10 years of age (group 3). No globule leucocytes were detected in any sections. No significant variations with breed or sex were observed in cell counts. The main variations were higher eosinophil counts in groups 2 and 3 and a marked increase of the MMC counts in the oldest horses (group 3). For each cell type, the infiltration was homogeneous and generalised along the large intestine. In the whole horse sample, the IE numbers were the only parameters that correlated with the MMC and SMMC counts. Very few significant relationships were found between mast cells and eosinophils in groups 1 and 3, whereas numerous positive correlations were recorded in group 2. In the whole horse sample, several correlations were found between different cell counts and cyathostome burdens. The numbers of larvae, adult worms, and the total worm burdens were related to some of the tissular eosinophil counts while the percentage of early third stage larvae (EL3) was linked to mast cell densities. These relations between cells and worm populations showed variations with age. In group 1, most of the significant associations were found between eosinophil counts (IE and SME) and the total numbers of larvae and worms; in group 2, they were noticed between the three eosinophil types and the total cyathostome burdens. In group 3, a MMC hyperplasia was observed and correlations were mostly recorded between these MMC and the total numbers of adult worms or the percentage of EL3. Several associations were also detected between eosinophils (mainly ME and/or IE) and different cyathostome burdens. These variations in the relationship between inflammatory cells and cyathostomes seemed to be consistent with the cellular changes observed among the three age groups. These results suggest that eosinophil and mast cell infiltrations quantified in the large intestine wall might be associated with cyathostome infection. PMID- 12127255 TI - Serologic responses of cats against experimental Sarcocystis neurona infections. AB - Sarcocystis neurona is the most important cause of a neurologic disease of horses, equine protozoal myeloencephalitis (EPM). Cats and other carnivores can act as its intermediate hosts and horses are aberrant hosts. Little is known of the sero-epidemiology of S. neurona infections in cats. In the present study, antibodies to S. neurona were evaluated by the S. neurona agglutination test (SAT). Cats fed sporocysts from the feces of naturally infected opossums or inoculated intramuscularly with S. neurona merozoites developed high levels (> or =1:4000) of SAT antibodies. Antibodies to S. neurona were not found in a cat inoculated with merozoites of the closely related parasite, Sarcocystis falcatula. These results should be useful in studying sero-epidemiology of S. neurona infections in cats. PMID- 12127257 TI - Elevated basal expression of liver peroxisomal beta-oxidation enzymes and CYP4A microsomal fatty acid omega-hydroxylase in STAT5b(-/-) mice: cross-talk in vivo between peroxisome proliferator-activated receptor and signal transducer and activator of transcription signaling pathways. AB - Long-term treatment of rodents with peroxisome proliferator chemicals, a group of structurally diverse nongenotoxic carcinogens, leads to liver cancer in a process dependent on the nuclear receptor peroxisome proliferator-activated receptor alpha (PPARalpha). Previous in vitro studies have shown that growth hormone (GH) can inhibit PPARalpha-dependent gene expression by down-regulation of PPARalpha expression and by a novel inhibitory cross-talk involving the GH-activated transcription factor STAT5b. Presently, we evaluate the role of STAT5b in mediating these inhibitory actions of GH on PPAR function using a STATb-deficient mouse model. Protein levels of three PPARalpha-responsive peroxisomal beta oxidation pathway enzymes (fatty acyl-CoA oxidase, 3-ketoacyl-CoA thiolase, and L bifunctional enzyme) were increased up to two- to threefold in STAT5b(-/-) relative to wild-type control mouse liver, as was the basal expression of two PPARalpha-regulated cytochrome P450 4A proteins. In contrast, protein levels of two PPARalpha-unresponsive peroxisomal enzymes, catalase and urate oxidase, were not affected by the loss of STAT5b. A corresponding increase in expression of fatty acyl-CoA oxidase and L-bifunctional enzyme mRNA, as well as PPARalpha mRNA, was observed in the STAT5b-deficient mice, suggesting a transcriptional mechanism for the observed increases. Although basal liver expression of PPARalpha and its target genes was thus elevated in STAT5b(-/-) mice, the clofibrate-induced level of enzyme expression was unaffected, suggesting that the inhibitory effects of STAT5b are overcome at high concentrations of PPARalpha activators. These findings support the hypothesis that GH and potentially other endogenous activators of STAT5b help to maintain liver PPARalpha function at a low basal level and may thereby moderate PPARalpha-dependent hepatocarcinogenesis and other responses stimulated by exposure to low levels of environmental chemicals of the peroxisome proliferator class. PMID- 12127258 TI - Dietary butylated hydroxytoluene protects against aflatoxicosis in Turkeys. AB - Turkeys are among the most sensitive species to the toxic effects of the mycotoxin aflatoxin B(1) (AFB(1)). In mammals, dietary antioxidants, such as butylated hydroxytoluene (BHT), have been shown to lessen the toxic effects of AFB(1) by various mechanisms. To test whether BHT protects against aflatoxicosis in turkeys, we supplemented the feed of 10-day-old male white turkeys with low (1000 ppm) and high (4000 ppm) BHT for 20 days. AFB(1) (1 ppm) was then added to the diets and continued for another 10 days. Birds in the AFB(1)-only group had a lower weight gain, a condition that had returned to near control in groups fed diets containing AFB(1) + BHT. Significant elevations in serum aspartate transaminase, alanine aminotransferase, and lactate dehydrogenase, which were evident in the AFB(1) group, were reversed in the AFB(1) + BHT groups. Histopathology revealed hepatic submassive necrotic lesions and biliary hyperplasia, the severity of which was lessened in the AFB(1) + BHT-treated birds. Hepatocellular hydropic degeneration was observed in the BHT-only group, but not in the AFB(1) + BHT groups. This condition associated with BHT treatment was found in a separate study to be reversible and without any long-term adverse effects. These results indicate that BHT counteracts many of the deleterious effects caused by AFB(1) and that this antioxidant may prove to be a viable feed additive for the reduction of aflatoxicosis in turkeys. PMID- 12127259 TI - Improved efficiency of hepatic mitochondrial function in rats with cholestasis induced by an acute dose of alpha-naphthylisothiocyanate. AB - Cholestasis is a feature of many chronic human liver diseases. Previous studies pointed out an impairment of mitochondrial function as a cause of hepatocyte dysfunction leading to cholestatic liver injury. This study was aimed to evaluate liver mitochondrial bioenergetics of alpha-naphthylisothiocyanate-treated Wistar rats, an experimental model of cholestasis. Serum markers of liver injury and endogenous adenine nucleotides were measured. Changes in membrane potential, mitochondrial respiration, and alterations in mitochondrial permeability transition pore induction were monitored. In rats injected with alpha naphthylisothiocyanate, liver injury with cholestasis developed within 48 h, as indicated by both serum enzyme activities and total bilirubin concentration. Liver mitochondria isolated from alpha-naphthylisothiocyanate-treated rats had a higher state 3 respiration, respiratory control ratio, ADP/O, and endogenous ATP/ADP ratio compared to controls. No change in state 4 respiration was observed. Associated with these parameters, cholestatic mitochondria exhibited an increased resistance to disruption of mitochondrial calcium homeostasis due to permeability transition pore induction. Hepatic mitochondria isolated from alpha naphthylisothiocyanate-treated rats exhibited an improved efficiency. These data indicate that an adaptive response to resist cell death occurs during alpha naphthylisothiocyanate-induced acute cholestasis. PMID- 12127260 TI - Salvia miltiorrhiza inhibits biliary obstruction-induced hepatocyte apoptosis by cytoplasmic sequestration of p53. AB - Cholestatic liver injury is caused by hepatocellular apoptosis induced by toxic bile salts. We have studied the effects of a traditional Chinese herbal medicine, Salvia miltiorrhiza, on apoptotic cell death in bile duct-ligated (BDL) rats. We also attempted to clarify the molecular mechanisms of the hepatoprotective effects of S. miltiorrhiza in this animal model. A water extract of S. miltiorrhiza (Sm-X; 200 mg/kg; po) was administered to BDL rats for 10 days. Rats were euthanized and apoptosis was detected in liver tissue by TUNEL staining. Western blot analysis and immunostaining for alpha-smooth muscle actin (alpha SMA), Bax, Bcl-2, and p53 were performed. Results show that the treatment of BDL rats with Sm-X significantly improved the liver function parameters, although the expression of alpha-SMA, a marker of hepatic stellate cell activation, was not affected. Treatment with Sm-X significantly reduced the number of apoptotic cells. A time-dependent decrease in Bax protein level and an increase in Bcl-2 protein level were observed in BDL rats treated with Sm-X. Immunohistochemical analysis demonstrated that p53 was strongly positive in hepatocyte nuclei of BDL rats but that treatment with Sm-X induced a cytoplasmic sequestration of p53. Taken together, hepatoprotective effects of Sm-X partially ascribe to the antiapoptotic property in hepatocytes. Treatment of Sm-X-induced cytoplasmic sequestration of p53, downregulation of Bax, and upregulation of Bcl-2 protein. This study identifies and delineates signaling factors involved in the antiapoptotic properties of Sm-X and suggests a potential application of S. miltiorrhiza in the clinical management of hepatic disease induced by toxic bile salts following biliary obstruction. PMID- 12127261 TI - Humic acid induces apoptosis in human endothelial cells. AB - Humic acid (HA) has been implicated as an etiologic factor in the vasculopathy of Blackfoot disease. In this study, the ability of HA to induce apoptosis was studied in cultured human umbilical vein endothelial cells. Treatment of endothelial cells with a variety of concentrations of HA (50-200 microg/ml) resulted in dose- and time-dependent sequences of events marked by apoptosis as shown by loss of cell viability, chromatin condensation, and internucleosomal DNA fragmentation. Antioxidants (superoxide dismutase, vitamin C, and vitamin E) and Ca(2+) chelator (BAPTA) effectively suppressed HA-induced DNA fragmentation (apoptosis). Further studies have shown that HA induced dramatic Ca(2+)-dependent caspase activation (2, 3, 6, 8, and 9). In contrast, negligible caspase-1 activation was observed. The increase in HA-induced apoptosis correlated with a reduction in Bcl-2, a potent cell death inhibitor, and an increase in Bax protein levels, which heterodimerizes with and thereby inhibits Bcl-2. Both of the antioxidants vitamin C and vitamin E prevented the dysregulation of Bcl-2 and Bax in HA-treated endothelial cells. Furthermore, the increase in p53 protein levels correlated with an increase in HA-induced apoptosis. We concluded that both Ca(2+) and oxidative stress were mediators of apoptosis caused by HA and the induction of apoptotic cell death on endothelial cells may be important to the etiology of HA-induced vascular disorder of Blackfoot disease. PMID- 12127262 TI - Induction and inhibition of aromatase (CYP19) activity by various classes of pesticides in H295R human adrenocortical carcinoma cells. AB - Various pesticides known or suspected to interfere with steroid hormone function were screened in H295R cells for effects on catalytic activity and mRNA expression of aromatase. Dibutyl-, tributyl-, and triphenyltin chloride decreased aromatase and ethoxyresorufin O-deethylase activities concentration dependently (1-300 nM; 24-h exposure). However, these decreases occurred only at cytotoxic concentrations, indicated by decreases in mitochondrial MTT reduction and intracellular neutral red uptake. The organotins did not cause direct inhibition during the catalytic assay (1-1000 nM; 1.5-h exposure). The same was true for p,p'-DDT, and o,p-DDT, and o,p-DDE, which decreased aromatase activity only at cytotoxic concentrations (> or =10 microM; 24-h exposure). p,p'-DDE had no effect on aromatase activity or cell viability at 1 and 10 microM. Various imidazole like fungicides were aromatase inhibitors. Imazalil and prochloraz were potent mixed inhibitors (K(i)/K(i)(') = 0.04/0.3 and 0.02/0.3 microM, respectively), whereas propiconazole, difenoconazole, and penconazole were less potent competitive inhibitors (K(i) = 1.9, 4.5, and 4.7 microM, respectively). Fenarimol, tebuconazole, and hexaconazole decreased aromatase activity close to cytotoxic concentrations. Vinclozolin, as was shown previously for atrazine, induced aromatase activity and CYP19 mRNA levels about 2.5- and 1.5-fold, respectively. To investigate the mechanism of aromatase induction in H295R cells, the ability of the pesticides to increase intracellular cAMP levels was examined. Vinclozolin (100 microM) and atrazine (30 microM) increased cAMP levels about 1.5 fold above control. Forskolin and isobutyl methylxanthine (IBMX) increased cAMP levels 3 and 1.8-fold, respectively. Time-response curves for cAMP induction and concentration-response curves for aromatase induction by vinclozolin, atrazine, and IBMX were similar, suggesting that the mechanism of aromatase induction by these pesticides is mediated through inhibition of phosphodiesterase activity. PMID- 12127263 TI - Contribution of dichloroacetate and trichloroacetate to liver tumor induction in mice by trichloroethylene. AB - Determining the key events in the induction of liver cancer in mice by trichloroethylene (TRI) is important in the determination of how risks from this chemical should be treated at low doses. At least two metabolites can contribute to liver cancer in mice, dichloroacetate (DCA) and trichloroacetate (TCA). TCA is produced from metabolism of TRI at systemic concentrations that can clearly contribute to this response. As a peroxisome proliferator and a species-specific carcinogen, TCA may not be important in the induction of liver cancer in humans at the low doses of TRI encountered in the environment. Because DCA is metabolized much more rapidly than TCA, it has not been possible to directly determine whether it is produced at carcinogenic levels. Unlike TCA, DCA is active as a carcinogen in both mice and rats. Its low-dose effects are not associated with peroxisome proliferation. The present study examines whether biomarkers for DCA and TCA can be used to determine if the liver tumor response to TRI seen in mice is completely attributable to TCA or if other metabolites, such as DCA, are involved. Previous work had shown that DCA produces tumors in mice that display a diffuse immunoreactivity to a c-Jun antibody (Santa Cruz Biotechnology, SC-45), whereas TCA-induced tumors do not stain with this antibody. In the present study, we compared the c-Jun phenotype of tumors induced by DCA or TCA alone to those induced when they are given together in various combinations and to those induced by TRI given in an aqueous vehicle. When given in various combinations, DCA and TCA produced a few tumors that were c-Jun+, many that were c-Jun-, but a number with a mixed phenotype that increased with the relative dose of DCA. Sixteen TRI-induced tumors were c-Jun+, 13 were c-Jun-, and 9 had a mixed phenotype. Mutations of the H-ras protooncogene were also examined in DCA-, TCA-, and TRI-induced tumors. The mutation frequency detected in tumors induced by TCA was significantly different from that observed in TRI-induced tumors (0.44 vs 0.21, p < 0.05), whereas that observed in DCA-induced tumors (0.33) was intermediate between values obtained with TCA and TRI, but not significantly different from TRI. No significant differences were found in the mutation spectra of tumors produced by the three compounds. The presence of mutations in H-ras codon 61 appeared to be a late event, but ras-dependent signaling pathways were activated in all tumors. These data are not consistent with the hypothesis that all liver tumors induced by TRI were produced by TCA. PMID- 12127264 TI - Mechanisms involved in responses to the poroxisome proliferator WY-14,643 on gap junctional intercellular communication in V79 hamster fibroblasts. AB - WY-14,643, a potent hepatic peroxisome proliferator, decreased gap junctional intercellular communication when used at low and intermediate concentrations (1 nM to 10 microM) in the V79 Chinese hamster fibroblast cell line. It did not decrease intercellular communication in early passage Syrian hamster embryo fibroblasts. The mechanism of WY-14,643-induced suppression of intercellular communication was studied by preexposure of V79 cells to inhibitors of protein kinase C, mitogen-activated protein kinases, phosphatidylinositol 3-kinase, or mammalian target-of-rapamycin before addition of WY-14,643. Only protein kinase C, particularly the delta isoform, appeared involved in the inhibition of communication by WY-14,643. Also clofibrate-induced suppression of GJIC in V79 cells appeared to involve protein kinase Cdelta. Furthermore, WY-14,643 did not cause any activation of the mitogen-activated protein kinases p44/42, p38, or Jun N-terminal kinase. When WY-14,643 was used at a higher concentration (100 microM), intercellular communication was increased both in V79 and Syrian hamster embryo cells. This effect was inhibited by preexposure of V79 cells to brefeldin A. Thus, there may be a pool of connexins in the Golgi complex that could be recruited to the plasma membrane upon exposure to high concentrations of WY 14,643. PMID- 12127265 TI - Effect of linoleic acid metabolites on Na(+)/K(+) pump current in N20.1 oligodendrocytes: role of membrane fluidity. AB - Metabolic derivatives of linoleic acid, both monoepoxides and diols, have been reported to be toxic in humans and multiple animal tissue preparations. A previous electrophysiological study has shown these compounds produce multiple effects on the electrical activity of rat ventricular myocytes. The hydrophobic nature of these compounds suggests the possibility that these effects may be due to nonspecific lipid interactions, i.e., changes in membrane fluidity. This study investigates membrane fluidity as a possible mechanism by which linoleic acid metabolites inhibit Na(+)/K(+) pump current (I(p)). This study showed that positional isomers 9,10- and 12,13-epoxy-octadecenoic acid (EOA) and 9,10- and 12,13-dihydroxy-OA (DHOA) inhibit I(p) in a dose-dependent manner in N20.1 mouse oligodendrocytes, with greater inhibition produced by EOAs. These compounds, at 10 microM, inhibited I(p) by 4.7 +/- 1.6, 18.2 +/- 0.5, 11.7 +/- 0.5, and 25.1 +/ 0.9% for 12,13-DHOA, 9,10-DHOA, 12,13-EOA, and 9,10-EOA, respectively, in oligodendrocytes. Fluorescence recovery after photobleaching measurements showed that both DHOA isomers produced a 7-8% increase in diffusion coefficient of the probe at 10 microM, whereas the diffusion coefficient was decreased by 5 and 13% by 9,10-EOA and 12,13-EOA, respectively. There was no apparent correlation between membrane fluidity and inhibition of I(p) by these four linoleic acid metabolites. These results indicate that membrane fluidity alone cannot explain the effects of these compounds on I(p) and suggest that they have a specific interaction with the Na(+)/K(+) pump. PMID- 12127266 TI - Antioxidant and prooxidant activities of alpha-lipoic acid and dihydrolipoic acid. AB - Reactive oxygen (ROS) and nitrogen oxide (RNOS) species are produced as by products of oxidative metabolism. A major function for ROS and RNOS is immunological host defense. Recent evidence indicate that ROS and RNOS may also function as signaling molecules. However, high levels of ROS and RNOS have been considered to potentially damage cellular macromolecules and have been implicated in the pathogenesis and progression of various chronic diseases. alpha-Lipoic acid and dihydrolipoic acid exhibit direct free radical scavenging properties and as a redox couple, with a low redox potential of -0.32 V, is a strong reductant. Several studies provided evidence that alpha-lipoic acid supplementation decreases oxidative stress and restores reduced levels of other antioxidants in vivo. However, there is also evidence indicating that alpha-lipoic acid and dihydrolipoic acid may exert prooxidant properties in vitro. alpha-Lipoic acid and dihydrolipoic acid were shown to promote the mitochondrial permeability transition in permeabilized hepatocytes and isolated rat liver mitochondria. Dihydrolipoic acid also stimulated superoxide anion production in rat liver mitochondria and submitochondrial particles. alpha-Lipoic acid was recently shown to stimulate glucose uptake into 3T3-L1 adipocytes by increasing intracellular oxidant levels and/or facilitating insulin receptor autophosphorylation presumably by oxidation of critical thiol groups present in the insulin receptor beta-subunit. Whether alpha-lipoic acid and/or dihydrolipoic acid-induced oxidative protein modifications contribute to their versatile effects observed in vivo warrants further investigation. PMID- 12127267 TI - Getting around lethality with inducible Cre-mediated excision. PMID- 12127268 TI - Discovering new genes with advanced homology detection. AB - Most genome annotation protocols combine ab initio predictions with transcription and homology analyses to produce reliable gene predictions but they often fail to detect many actual genes. Alternative approaches involving more sensitive homology recognition methods are playing an increasingly important role in the next stage of gene discovery. The hunt for new genes is far from over. PMID- 12127269 TI - The grand challenge: facing up to proteomics. PMID- 12127270 TI - Current trends in bioinformatics. PMID- 12127271 TI - Using proteomics to identify targets and validate leads. PMID- 12127272 TI - Crystallization comes of age. PMID- 12127276 TI - PCR as an emerging option in the microbial quality control of drinking water. PMID- 12127277 TI - ImClone's failed culture of expertise. PMID- 12127278 TI - Needed: models of biotechnology intellectual property. AB - Although never uncontroversial, intellectual property rights in biotechnological innovation are once more the focus of intense debate. The debate has yet to reach any result, largely because of several important errors in the way that various disciplines approach it. These errors include making assumptions without empirical basis and conflating various intellectual property regimes. What is needed is a transdisciplinary integrated method to correct these errors. Such a method can be implemented through the construction of alternative models of intellectual property protection designed to balance the various social, ethical and economic constraints that affect biotechnology. PMID- 12127279 TI - Toxicogenomics and toxic torts. AB - One of the first practical applications of toxicogenomics will probably be in the context of toxic tort personal injury litigation. Gene expression changes that'fingerprint' exposure to particular classes of toxic substances can potentially be used to demonstrate exposure, prove causation and support novel damage claims in lawsuits brought by citizens injured by toxic exposures. Although the potential use of toxicogenomic data in toxic tort litigation is immense, there is a danger of premature use of such data before they have been adequately validated and characterized. PMID- 12127280 TI - Antibody separation by hydrophobic charge induction chromatography. AB - Hydrophobic charge induction chromatography using 4-mercapto-ethyl-pyridine as the ligand is an effective method for the separation of antibodies from a variety of feedstocks. Antibodies are adsorbed in physiological conditions without preliminary concentration. Desorption occurs when the pH is lowered, thus inducing an ionic charge of the same sign to the ligand and the antibody. Antibody capture conditions are compatible with crude samples in terms of pH, conductivity, binding capacity and expression level. The final purity of the antibody is feedstock dependent, but can reach levels of purity as high as 98%. Examples of antibody separation are given and ligand structure information discussed. PMID- 12127281 TI - Biological substitutes for pesticides. AB - In the 20th century an increasing number of pesticides, based on biocidal molecules, were the means for a substantial increase in food and fibre production and quality. Because of health and environmental concerns continued extensive use of such molecules is intensively debated and substitutes are often urgently required. Beside crop plant resistance, various biological control methods based on natural pest suppressing organisms are regarded as main alternatives. Several approaches and concepts also have been tested and commercial organism-based preparations are steadily increasing. However, further biotechnological efforts are required to give them status of being practical substitutes to pesticides. At present they are not comparable to pesticides in meeting efficacy, market and other expectations, but they still have a promising future, especially where genetically modified organisms can be used. PMID- 12127282 TI - Combing the genome for genomic instability. AB - Genomic instability is one of the major features of cancer cells. The clinical phenotypes associated with several human diseases have been linked to recurrent DNA rearrangements and dysfunction of DNA replication processes that involve unstable genomic regions. Analysis of these rearrangements, which are frequently submicroscopic and can lead to loss or gain of dosage-sensitive genes or gene disruption, requires the development of sensitive, high-resolution techniques. This will lead to a better understanding of the mechanisms underlying genome instability and a greater awareness of the role of chromosomal rearrangements in disease. A new technology that involves molecular combing, a method that permits straightening and aligning molecules of genomic DNA, should make possible a detailed analysis of genomic events at the level of single DNA molecules. Such a single molecule approach could help to elucidate important properties that are masked in bulk studies. PMID- 12127283 TI - Tissue engineering: advances in in vitro cartilage generation. AB - Damaged or diseased articular cartilage frequently leads to progressive debilitation resulting in a marked decrease in the quality of life. Tissue engineering, a budding field in modern biomedical sciences, promises creation of viable substitutes for failing organs or tissues. It represents the amalgamation of rapid developments in cellular and molecular biology on the one hand and material, chemical and mechanical engineering on the other. Current tissue engineering approaches are mainly focused on the restoration of pathologically altered tissue structure based on the transplantation of cells in combination with supportive matrices and biomolecules. The ability to manipulate and reconstitute tissue structure and function in vitro has tremendous clinical implications and is likely to have a key role in cell and gene therapies in coming years. PMID- 12127284 TI - The history of the peer-review process. AB - The peer-review process is a turf battle with the ultimate prize of the knowledge, science or doctrine being published. On the one side, we have the writers and originators of ideas, on the other, we have the editors and critics. But it was not always so. PMID- 12127300 TI - Kyolic and Pycnogenol increase human growth hormone secretion in genetically engineered keratinocytes. AB - The amount of human growth hormone (HGH) decreases significantly after the age of 30. This decrease has been implicated as one of the major causes in the signs of aging, such as thinning of the skin and bones, a decrease in lean muscle mass and an increase in adipose tissue. Supplementing the body's dwindling supply with recombinant human growth hormone (rHGH) has been shown to reverse the signs and symptoms of aging. However, drawbacks in rHGH replacement therapy include prohibitively high cost, the need for repeated injection and side effects such as carpel tunnel syndrome, gynecomastia and insulin resistance. The purpose of this study was to establish an in vitro model using genetically-engineered keratinocytes to screen natural compounds for the ability to stimulate HGH secretion. We now report that a combination of equal amounts of L-arginine and L lysine, aged garlic extract (Kyolic), S-allyl cysteine and Pycnogenol significantly increased secretion of HGH in this in vitro model. The data indicate that this in vitro model may be used to screen for other secretagogues. PMID- 12127299 TI - Growth hormone replacement therapy for adults: into the new millennium. PMID- 12127301 TI - Differential induction of transcription factors and expression of milk protein genes by prolactin and growth hormone in the mammary gland of rabbits. AB - Previously we demonstrated that administration of lactogenic hormones - prolactin (PRL) and growth hormone (GH) - to pregnant rabbits differentially induces expression of casein and whey proteins in the mammary gland. Now we extend these observations to transcription factors (TFs) that are responsive for differential induction of milk protein genes. Analysis of correlation between the number of putative TF binding sites in 5'-upstream sequences and the levels of induction of milk protein genes allowed preselection of the TFs involved. An electrophoretic mobility shift assay with nuclear proteins derived from rabbit mammary glands showed changes in the patterns of Stat5, MAF, NF1 and Oct1 DNA-protein binding during progression of pregnancy and transition to lactation. Administration of lactogenic hormones - PRL or GH - to early-pregnant rabbits induced DNA-protein complexes similar to those formed by nuclear proteins from the mammary glands of lactating (Stat5, MAF, NF1) or late-pregnant (Oct1) animals. Induction of milk protein genes by PRL was several-fold greater than that by GH. However, PRL and GH similarly induced MAF DNA-protein complexes, thus suggesting that the amount of MAF factor in the mammary gland can be limiting for expression of these genes. Our study for the first time provided the evidence that in the mammary gland both PRL and GH can induce DNA-binding activity of transcription factors other than Stats. PMID- 12127302 TI - The relationship between serum levels of insulin-like growth factor-I and its binding proteins and microvascular function in acromegalic patients. AB - The system of insulin-like growth factor-I (IGF-I) and its binding proteins is thought to be involved in the pathogenesis of vascular damage under different pathological circumstances. The results of various studies are rather controversial. This study considers the relationship between the activity of this system and the function of microcirculation in acromegalic patients. Thirteen patients with hormonally active acromegaly and 15 healthy controls were included in the study. The growth hormone, free IGF-I, IGF-I, IGF binding protein (IGFBP) 1, -2, -3 and -6 serum levels and parameters of lipid metabolism were determined. The function of microcirculation was determined by laser Doppler fluxmetry and the intima media thickness of the common carotid artery was measured by ultrasound. We noted significant reduction in postocclusive reactive hyperaemia (PORH(max)) (P < 0.01), in thermal hyperaemia (TH(max)) (P < 0.05) and in the velocity of reaction in both tests in the group of acromegalic patients. A significant negative correlation between free IGF-I serum levels and maximal perfusion during thermal hyperaemia TH(max) (P < 0.02) was found in the control group. Statistically significant positive correlation between free IGF-I serum levels and the time to maximal perfusion in postocclusive reactive hyperaemia PORH(max) (P < 0.05) was found in the group with hormonally active acromegaly. Moreover, a positive relationship between IGFBP-1 serum levels and serum levels of total (P < 0.01) and low density lipoprotein (LDL) (P < 0.05) cholesterol was found in the group of patients with acromegaly. We conclude that the function of microcirculation is impaired in patients with acromegaly and that free IGF-I serum levels may affect the microvascular function as measured by laser Doppler fluxmetry. In addition, we found a significant relationship between the serum levels of IGFBP-1 and those of total and LDL cholesterol in the group of patients with hormonally active acromegaly. PMID- 12127303 TI - Glucagon and GLP-1 stimulate IGFBP-1 secretion in Hep G2 cells without effect on IGFBP-1 mRNA. AB - Glucagon has previously been reported to increase serum levels of insulin-like growth factor binding protein-1 (IGFBP-1) in humans. The in vitro effect of glucagon and glucagon-like peptide-1 (7-36) amide (GLP-1) was investigated in Hep G2 human hepatoma cells. The expression of IGFBP-1 mRNA was determined by solution hybridization assay and IGFBP-1 secretion was measured by radioimmunoassay. In contrast to forskolin the peptides glucagon and GLP-1 had no effect on IGFBP-1 mRNA at 3, 6 and 24 h incubation or any detectable effect on the apparent half-life of IGFBP-1 mRNA. However, the exposure to glucagon (10 microg/mL, 2.87 microM) and GLP-1 (1 microM) caused a two-fold stimulation in protein levels of IGFBP-1 after 6 h incubation, declining to control levels after 24 h. This transient effect was dose dependent, remained when transcription was inhibited and required protein synthesis. The regulation of IGFBP-1 secretion by glucagon and GLP-1 appeared to be cAMP independent. In conclusion, glucagon and GPL-1 were shown to have a post-transcriptional stimulatory effect on IGFBP-1 release. PMID- 12127304 TI - Diethylnitrosamine induces long-lasting re-expression of insulin-like growth factor II during early stages of liver carcinogenesis in mice. AB - The expression of the insulin-like growth factor II (IGF-II) gene (Igf2) in rodents is completely abrogated in almost all adult tissues. A prominent exception are neoplasms in which IGF-II frequently serves as an autocrine growth factor. We have investigated the potential role of Igf2 expression during liver carcinogenesis. After application of diethylnitrosamine (DEN) preneoplastic foci and adenomas emerged in liver tissue of wild-type and phosphoenolpyruvate carboxykinase (PEPCK)-IGF-II transgenic mice. Surprisingly, number and size of preneoplastic foci were not significantly increased in PEPCK-IGF-II mice as compared with wild-type animals. In situ preparation showed that early adenomas expressed Igf2 transcripts. Reverse transcriptase polymerase chain reaction (RT PCR) and restriction enzyme analysis confirmed that DEN treatment had indeed reactivated the hepatic expression of murine Igf2 in control mice in a dose dependent manner. This re-expression of Igf2 persisted for at least 18 months. Species-specific RT-PCR analyses also revealed the presence of murine Igf2 mRNAs in some PEPCK-IGF-II mice. A similar reactivation of Igf2 was detected in bovine growth hormone transgenic mice which develop hepatocellular neoplasms with high frequency. Our results suggest that reactivation of Igf2 is an early event during hepatocarcinogenesis in mice. Its appearance in two independent animal models suggests that Igf2 may be important at pivotal checkpoints of hepatocarcinogenesis. PMID- 12127305 TI - Integration of the regulation of reproductive function and energy balance: lactation as a model. AB - Lactation is a physiological model for studying how the hypothalamus integrates peripheral signals, such as sensory signals (suckling stimulus) and those denoting energy balance (leptin), to alter hypothalamic function regulating food intake/energy balance and reproduction. The characteristics of food intake/energy balance during lactation are extreme hyperphagia, coupled with negative energy balance. The arcuate nucleus Neuropeptide Y (ARH-NPY) system is activated by: (1) brainstem projections specifically activated by the suckling stimulus, and (2) the decrease in leptin in response to the metabolic drain of milk production. NPY neurons from the ARH make direct contact with GnRH neurons and with CRH neurons in the PVH. NPY neurons also make contact with orexin and MCH neurons in the LHA, which, in turn, make contacts with GnRH neurons. Thus, the ARH-NPY system provides a neuroanatomical framework by which to integrate changes in food intake/energy with the regulation of cyclic reproductive function. PMID- 12127307 TI - Development of gonadotropin-releasing hormone-1 neurons. AB - Gonadotropin releasing hormone-1 (GnRH-1) neurons, critical for reproduction, are derived from the nasal placode and migrate into the brain during prenatal development. Once within the brain, GnRH-1 cells become integral components of the CNS-pituitary-gonadal axis, essential for reproductive maturation and maintenance of reproductive function in adults. This review focuses on the lineage and development of the GnRH-1 neuroendocrine system. Although the migration of these cells from nose to brain has been well documented in a variety of species, many questions remain concerning the melecules and cues directing GnRH-1 cell differentiation, migration, axon targeting, and establishment and control of GnRH-1 secretion. These process most likely involve multiple and redundant cues because if these mechanisms fail, reproduction dysfunction will ensue and guarantee that this defect does not remain in the gene pool. PMID- 12127306 TI - Development of the neuroendocrine hypothalamus. AB - The development of the neuroendocrine hypothalamus has been studied using a variety of neuroanatomical and molecular techniques. Here, the major findings that mold our understanding of hypothalamic development are reviewed. The rat hypothalamus is generated predominantly from the third ventricular neuroepithelium in a "lateral early to medial late" pattern dictated perhaps by the medially receding third ventricle. Neuroendocrine neurons seem to exhibit a delayed migrational strategy, showing relatively early birthdates, although they are located in the latest-generated, periventricular nuclei. Several homeobox genes seem to play a role in hypothalamic development, and gene knockout experiments implicate a number of genes of importance in the generation of the neuroendocrine cell type. PMID- 12127308 TI - Detection of pyrogens in intravenous IgG preparations. AB - Five different intravenous IgG (i.v. IgG) preparations were assessed for their capacity to modify the pyrogenic response to bacterial lipopolysaccharide (LPS) of rabbits under the conditions of a pharmacopoeal test. Four of the five preparations were found to mitigate the reaction rendering the result "non pyrogenic" with an LPS dose proved pyrogenic when administered in saline or in albumin. Bacterial LPS was found readily detectable by a simple Limulus amoebocyte lysate (LAL) gelation test. Four of six brands of i.v. IgG were found reactive in the test under conditions adjusted to detect the FDA limit. The reaction obtained upon addition of standard LPS to the negative preparations supported the validity of the assay. The LAL reactivity of two of the reactive preparations was inhibited by laminarin, a compound known to inhibit Limulus lysate gelation by beta-D-glucan, but not by Polymyxin B. Specific detection of bacterial endotoxins in i.v. IgG solutions requires inhibition of the beta-D glucan pathway of the Limulus lysate coagulation. Using an appropriate inhibitor, the LAL gelation test is suitable to detect a potential endotoxin contamination in i.v. IgG which might have not been unravelled by the in vivo test for pyrogens. PMID- 12127309 TI - Development of a highly sensitive in vitro assay method for biological activity of endotoxin contamination in biological products. AB - The pyrogen test in rabbits has been replaced by the bacterial endotoxin test. The endotoxin test, however, showed a considerable discrepancy with pyrogenicity and was, therefore, assumed to have an efficacy limitation in directly predicting harmful biological effects of endotoxin. We developed a sensitive in vitro assay method by making use of tumour necrosis factor alpha (TNF-alpha) induction in RAW264.7 cells, which showed a fine correlation with pyrogenicity in rabbits. RAW264.7 cells maintained by serial subculture under an endotoxin-free condition have gained the similar level of sensitivity as the endotoxin test to allow extensive dilutions of a drug for eliminating adverse effects on the cells. The in vitro TNF-alpha induction assay was shown to be capable to detect quantitatively a synergistic effect of a drug and endotoxin. The synergy is assumed necessary to be taken into consideration to define the limit value for the endotoxin test for guaranteeing the similar level of safety as by the pyrogen test. PMID- 12127310 TI - Production of a Mycobacterium avium ssp. paratuberculosis purified protein derivative (PPD) and evaluation of potency in guinea pigs. AB - A Mycobacterium avium ssp. paratuberculosis purified protein derivative (PPD) was produced and the biologic activity evaluated in sensitized guinea pigs. The PPD when adjusted to a protein concentration of 1mg/ml induced a delayed-type hypersensitivity response comparable to USDA Johnin OT 133-8707. PMID- 12127311 TI - Quality control of polyvalent pneumococcal polysaccharide-protein conjugate vaccine by nephelometry. AB - A nephelometric method was used for quantitative analysis of individual polysaccharides (PSs) in a polyvalent pneumococcal conjugate vaccine using CRM(197) as carrier protein. Using this method, the individual types 4, 6B, 9V, 14, 18C, 19F and 23F PSs were found to range between 82.3 to 119% of the manufacturer's indicated values. During conjugation using reductive amination, pneumococcal PS was first oxidized to introduce aldehyde groups. Higher or lower levels of antigen-antibody reaction were observed in periodate activated and then reduced PS of some serotypes compared to non-treated PS. Use of oxidized and reduced PS may provide an early indication of change in conjugation process. Furthermore, since the final monovalent and polyvalent conjugate vaccines gradually change during the storage period, the nephelometry provides an useful analytical method for stability study of these vaccines. PMID- 12127312 TI - Potency control of diphtheria component in adsorbed vaccines by in vitro neutralization tests. AB - Samples from 20 lots of dT vaccine and from 20 lots of DTP vaccine were used to standardize and validate the Vero cell and the toxin binding inhibition (ToBI) tests for the potency control of diphtheria component. For the Vero cell method, violet crystal solution was used to stain the cells and estimate the endpoint of diluted diphtheria antitoxin. Diphtheria anatoxin was used for performing the ToBI test instead of toxin. The results obtained by both in vitro tests were similar to those obtained by in vivo toxin neutralization test in guinea pigs. The various analysis and the chi(2) test applied to evaluate the reproducibility and homogeneity, respectively, among in vitro tests and in vivo toxin neutralization test did not detect statistical significant difference for both analysed vaccines. An excellent correlation among in vitro tests and in vivo neutralization test was observed by Spearman's correlation coefficient. PMID- 12127313 TI - Usefulness of sugar mapping by liquid chromatography/mass spectrometry in comparability assessments of glycoprotein products. AB - We have previously reported that sugar-mapping by liquid chromatography/mass spectrometry (LC/MS) equipped with a graphitized carbon column (GCC) can be useful for structural analysis of carbohydrates in a glycoprotein. In this paper, we evaluated sugar-mapping with regard to its use in comparability assessment of glycoprotein products. Erythropoietins (EPO) produced from three different sources were chosen as models of the closely related glycoprotein products. The two-dimensional displays of sugar maps drawn by LC/MS with GCC clearly showed the differences in carbohydrate heterogeneity with regard to sialylation, acetylation, and sulphation patterns among three EPOs. Exoglycosidase digestion followed by sugar-mapping provided information regarding the structure of characteristic carbohydrates in each EPO. These results demonstrate that LC/MS with GCC can reveal the details of carbohydrate heterogeneity in order to distinguish between closely related glycoprotein products. Our method can thus be useful in comparability assessments of therapeutic glycoproteins. PMID- 12127314 TI - Effect of gamma irradiation on human cortical bone transplants contaminated with enveloped and non-enveloped viruses. AB - In the production of bone grafts intended for transplantation, basic safety measures to avoid the transmission of pathogens are selection and serological screening of donors for markers of virus infections. As an additional safety tool we investigated the effect of gamma irradiation on the sterility of human bone diaphysis transplants and evaluated its impact on the virus safety of transplants. Model viruses were included in the study to determine the dose necessary to achieve a reduction factor for the infectivity titres of at least 4 log(10) at a temperature of -30+/-5 degrees C. The following viruses were used: human immunodeficiency virus type 2 (HIV-2), hepatitis A virus (HAV), and poliovirus (PV-1), and the following model viruses: pseudorabies virus (PRV) as a model for human herpesviruses, bovine viral diarrhoea virus (BVDV) for HCV, and bovine parvovirus (BPV) for parvovirus B19. A first approach was to determine the D(10) values (kGy) for the different viruses (virus inactivation kinetics: BPV 7.3; PV-1 7.1; HIV-2 7.1; HAV 5.3; PRV 5.3; BVDV <3.0 kGy). Based on these results, inactivation of these viruses was studied in experimentally contaminated human bone transplants (femoral diaphyses). For BPV, the most resistant one of the viruses studied, a dose of approximately 34 kGy was necessary to achieve a reduction of infectivity titres of 4 log(10). We therefore recommend a dose of 34 kGy for the sterilisation of frozen bone transplants. PMID- 12127315 TI - Inactivation of lipid enveloped viruses by octanoic Acid treatment of immunoglobulin solution. AB - Inactivation of lipid enveloped viruses by treatment with octanoic acid has been investigated for three intravenous immunoglobulin preparations, using Human Immunodeficiency Virus, Bovine Viral Diarrhoea Virus, Sindbis Virus and Pseudorabies Virus as test viruses. At a concentration of 7.45 g octanoic acid per kg solution complete inactivation of lipid enveloped viruses to below detectable level (>5.36, >4.68, >6.25 and >5.55 log(10), respectively) was achieved within the first minutes of treatment. Octanoic acid treatment as described here, has been demonstrated as an effective and rapid virus inactivation procedure, which shows high robustness at the tested ranges of temperature, pH and protein content of the test material. However, pH must be considered as a critical parameter of treatment, as octanoic acid fails to inactivate lipid coated viruses at basic pH. At suitable conditions, e.g. pH<6.0 and a concentration of >3.7 g/kg, octanoic acid treatment gives reliable and highly effective inactivation of lipid enveloped viruses. PMID- 12127316 TI - Effect of preservatives on IgG aggregation, complement-activating effect and hypotensive activity of horse polyvalent antivenom used in snakebite envenomation. AB - Intravenous administration of antivenoms is associated with early adverse reactions in a number of cases, but the causes of this phenomenon are still unclear. The effect of preservatives (phenol and thimerosal) on IgG aggregate and dimer formation, in vitro complement-activating effect and hypotensive activity of a whole IgG horse liquid polyvalent antivenom, produced by caprylic acid fractionation, was assessed. These parameters were studied since they have been associated with the development of early adverse reactions to the administration of antivenoms and human immunoglobulins. After a three-year storage period at 4 degrees C, antivenoms with preservatives had an increased content of IgG aggregates and dimers when compared with antivenom devoid of phenol and thimerosal. These observations correlate with a slight increment in the turbidity of preservative-containing antivenoms. The three antivenoms studied (formulation: no preservatives; with phenol and thimerosal; with thimerosal alone) activated human complement in vitro, with only minor quantitative differences among them. When antivenoms were administered as a bolus intravenous injection in rats, a rapid and prominent hypotension of short duration was observed after injection of phenol-containing antivenom, whereas such an effect was absent in antivenom free of preservative and in the one containing only thimerosal. Bolus injection of saline solution with phenol resulted in a similar hypotension, indicating that the effect is due to phenol. However, when phenol-containing antivenom was diluted 1:5 with saline solution before infusion, as occurs in the clinical use of this product, no hypotension was observed. Our results stress the need to evaluate the effects of preservatives on the physicochemical and pharmacological characteristics of antivenoms. PMID- 12127317 TI - Enveloped virus inactivation by caprylate: a robust alternative to solvent detergent treatment in plasma derived intermediates. AB - Solvent-detergent treatment, although used routinely in plasma product processing to inactivate enveloped viruses, substantially reduces product yield from the human plasma resource. To improve yields in plasma product manufacturing, a new viral reduction process has been developed using the fatty acid caprylate. As licensure of plasma products warrants thorough evaluation of pathogen reduction capabilities, the present study examined susceptibility of enveloped viruses to inactivation by caprylate in protein solutions with varied pH and temperature. In the immunoglobin-rich solutions from Cohn Fraction II+III, human immunodeficiency virus, Type-1, bovine viral diarrhea virus (BVDV), and pseudorabies virus were inactivated by caprylate concentrations of >/=9 mM, >/=12 mM, and >/=9 mM, respectively. Compared to solvent-detergent treatment, BVDV inactivation in Fraction II+III solution was significantly faster (20-60 fold) using 16 mM caprylate. Caprylate-mediated inactivation of BVDV was not noticeably affected by temperature within the range chosen manufacturing the immunoglobulin product. In Fraction II+III solutions, IgG solubility was unaffected by -17 dB, the cut-off score of calibrated IBI discriminating non responders to beta-blocker therapy in our previous report, was related to the poor outcome (chi(2)=4.43, P=0.035). The stepwise multivariate analysis revealed that both calibrated IBI in the septum>-17 dB (chi(2)=4.43, P=0.035) and LVFS<15% (chi(2)=3.89, P=0.049) were useful to predict the poor clinical outcome. The event free rate assessed by the Kaplan-Meier method was also significantly reduced in patients with calibrated IBI in the septum >-17 dB (chi(2)=6.594, P=0.01) and calibrated IBI in the posterior wall>-17 dB (chi(2)=4.215, P=0.04). However, LVFS<15% (chi(2)=3.576, not significant) did not contribute to discriminating the event free rate in the clinical course. CONCLUSIONS: The present study demonstrated that myocardial IB intensity was higher in DCM patients who followed a poor clinical course rather than in those with a good outcome. Therefore, it is clarified that myocardial ultrasonic tissue characterization in DCM patients is useful for assessing their clinical outcome after receiving not only the standard treatment but also beta-blocker therapy. PMID- 12127367 TI - QT dispersion in patients with different etiologies of left ventricular hypertrophy: the significance of QT dispersion in endurance athletes. AB - Left ventricular hypertrophy (LVH) increases the risk of ventricular arrhythmias and sudden death and has a significant effect on total cardiovascular mortality. QT dispersion (QTd) is a measure of inhomogeneous repolarization and is used as an indicator of arrhythmogenicity. In this study we detected QTd in patients with different etiologies of left ventricular hypertrophy and the effect of LVH in QTd on endurance athletes. The study group consisted of 147 white male subjects with 3 different etiologies of LVH and 30 healthy male individuals. The underlying etiologies of LVH were essential hypertension, valvular aortic stenosis and long term training (athletic heart). QTd was measured by surface electrocardiogram and Bazett's formula was used to correct QTd for heart rate (QTcd). Left ventricular mass was determined by transthoracic echocardiography and left ventricular mass index was calculated in relation to body surface area. The QTcd was significantly higher in patients with pathological LVH (due to hypertension and aortic stenosis) than in the athletes' group (physiological LVH) and healthy subjects (P<0.05). The magnitude of QTcd was similar between athletes and the control group (P=0.6). The difference of QTcd between the groups with pathological LVH was not statistically significant (P=0.1). In conclusion; the increasing of QT dispersion is associated with only pathological conditions of LVH. The left ventricular hypertrophy has not a negative effect in QT dispersion on endurance athletes. The measurement of QT dispersion may be a non-invasive useful method for screening additional pathological conditions in endurance athletes. PMID- 12127366 TI - Heart rate variability before the onset of ventricular tachycardia: differences between slow and fast arrhythmias. AB - BACKGROUND: We tested whether or not heart rate variability (HRV) changes can serve as early signs of ventricular tachycardia (VT) and predict slow and fast VT in patients with an implantable cardioverter defibrillator (ICD). METHODS AND RESULTS: We studied the ICD stored 1000 beat-to-beat intervals before the onset of VT (131 episodes) and during a control time without VT (74 series) in 63 chronic heart failure ICD patients. Standard HRV parameters as well as two nonlinear parameters, namely 'Polvar10' from symbolic dynamics and the finite time growth rates 'Fitgra9' were calculated. Comparing the control and the VT series, no linear HRV parameter showed a significant difference. The nonlinear parameters detected a significant increase in short phases with low variability before the onset of VT (for time series with less than 10% ectopy, P<0.05). Subdividing VT into fast (cycle length 270 ms) events, we found that the onset of slow VT was characterized by a significant increase in heart rate, whereas fast VT was triggered during decreased heart rates, compared to the control series. CONCLUSIONS: Our data may permit the development of automatic ICD algorithms based on nonlinear dynamic HRV parameters to predict VT before it starts. Furthermore, they may facilitate improved prevention strategies. PMID- 12127368 TI - Evaluation of the phase-plane ECG as a technique for detecting acute coronary occlusion. AB - OBJECTIVE: To evaluate the phase plane (PP) ECG as a method for detecting acute coronary occlusion (ACO). BACKGROUND: Balloon inflation in a coronary artery during PTCA produces acute myocardial ischemia. The sensitivity of the standard ECG for detecting ACO is approximately 50%, depending on the number of leads recorded. METHODS: The standard ECG signals of 18 patients (91 leads), undergoing PTCA were sampled and converted to digital data, prior to, and during acute coronary occlusion. PPs were constructed by projecting the ECG signals and their first derivatives onto a two-dimensional plane. Standard ECG signals and PPs, prior to ACO, were compared to their respective recordings and PPs during ACO. RESULTS: Using the standard ECG analysis, the acute occlusion was detected in 39 of 91 leads (43%), and in 15 of 18 patients (83%), whereas using the PP analysis it was detected in 82 of 91 leads (90%), and in all 18 patients (100%) (P<0.001, for leads). The median number of leads per patient demonstrating standard ECG changes was 2.0, whereas for the PP analysis it was 5.5 (P<0.001). The specificity of the PP method was 83.5%. CONCLUSIONS: The sensitivity of the PP method for detecting ACO during PTCA was superior to that of the standard ECG analysis. A smaller lead system is required to detect changes of ACO, during PTCA, when the PP method is used. The PP method is simple, low-priced, and may serve to detect acute myocardial ischemia in a number of clinical settings. PMID- 12127369 TI - C/T polymorphism of the intercellular adhesion molecule-1 gene (exon 6, codon 469). A risk factor for coronary heart disease and myocardial infarction. AB - BACKGROUND: The intercellular adhesion molecule-1 (ICAM-1) mediates the interaction of activated endothelial cells with leukocytes and plays a fundamental role in the pathogenesis of coronary atherosclerosis. ICAM-1 single base C/T polymorphism, which determines an amino acid substitution in the ICAM-1 protein in exon 6 codon 469, has been described. Our purpose was to determine whether this C/T polymorphism influences the risk of coronary heart disease (CHD) and myocardial infarction (MI) in humans. METHODS AND RESULTS: We enrolled 349 patients with angiographically documented CHD, including a sub-group of 179 patients with acute or chronic MI. The control group consisted of 213 patients with normal left ventricular function and no documented evidence of CHD. All patients and controls were Germans genotyped by polymerase chain reaction and allele-specific oligonucleotide techniques for the ICAM-1 polymorphism. In the patients with CHD and MI the frequencies of the T genotype (TT+TC) were significantly higher than the CC genotype compared to the control subjects (P<0.001). With the additional use of multivariable logistic regression analysis for CHD (TT+TC versus CC; P=0.011, odds ratio 2.21, 95% CI 1.20-4.07), we found a significant association between CHD and MI and the TT and TC genotype of the ICAM 1 gene polymorphism. CONCLUSIONS: These results suggest that the TT and TC genotype of the ICAM-1 gene polymorphism in codon 469 is a genetic factor that may determine an individual's susceptibility for CHD and MI. PMID- 12127370 TI - Contribution of stress echocardiography to clinical decision making in unselected ambulatory patients with known or suspected coronary artery disease. AB - BACKGROUND: Due to its higher diagnostic accuracy stress echocardiography (SE) has been advocated as a substitute for stress ECG to detect coronary heart disease (CAD). However, its contribution to clinical decision-making in unselected patients presenting to the ambulatory care centre for known or suspected coronary artery disease is unclear. METHODS: To evaluate the clinical value of SE in unselected patients, we prospectively obtained SE and stress ECG in 221 consecutive patients (142 males; mean age 58+/-12 years) presenting to the ambulatory care centre with known or suspected CAD. Patients with acute coronary syndrome were not included. RESULTS: Results of stress ECG and SE were concordant in 181 (82%) and discordant in 40 patients (18%). The clinical decision to recommend or to currently withhold coronary angiography was possible solely on the basis of clinical criteria and stress ECG findings in 191 (86.4%) patients but was guided by the results of SE in 30 patients (13.6%). Left heart catheterization and coronary angiography were conducted in 61 patients. In this population SE was more accurate (82.6%) than stress ECG (65.6%) in indicating significant coronary artery stenosis. CONCLUSION: Despite its higher accuracy, SE adds little to the information derived from dynamic stress ECG and symptom evaluation in unselected outpatients with known or suspected CAD. Thus, SE should not in general replace stress ECG as a screening method for detecting significant coronary artery disease, for both clinical and economic reasons. PMID- 12127371 TI - Do clinical and angiographic parameters predict failure of medical therapy in patients suitable for coronary angioplasty? AB - AIMS: Recent studies have suggested that patients with coronary disease suitable for angioplasty have an equally good outcome with medical therapy if clinically stable. Complex lesion morphology may predict acute events without intervention and stenosis severity influences the degree of collateralisation. This study was designed to assess the influence of these factors on clinical outcome. METHODS AND RESULTS: A retrospective review of patients suitable for angioplasty who were randomised to initial medical therapy as part of a multicentre study. Angiograms were reviewed for lesion characteristics, TIMI flow grade, and degree of collateralisation. Angiograms were available on 79 patients (13 female, 66 male). Mean age was 54.8 years (range 43-68) in the group crossing-over to revascularisation, and 58.4 (range 37-78) in the group who did not (P=ns). Seventeen patients crossed-over (two to CABG, 15 to PTCA) at 5.4 months (range 0 10) after initial angiography. Disease progression had occurred in 10/17 patients (58.8%), three of whom developed a new occlusion. Collateralisation was more likely in smokers, independent of lesion severity (P<0.05). Time to cross-over was not influenced by progression of disease. Crossing-over was not affected by age, diabetic status, cholesterol level, vessel involved, lesion severity, TIMI flow, lesion morphology, collateralisation, or the number of vessels diseased, but was more likely in females (P<0.05). CONCLUSION: This group of patients generally does well with medical therapy. Whilst the numbers are relatively small, there does not appear to be any reliable prospective marker, including the presence of spontaneous collateral channels on diagnostic angiography, to indicate which patients will fail medical therapy and require revascularisation. PMID- 12127373 TI - Blood pressure elevation after phenylephrine infusion may adversely affect myocardial perfusion in patients with coronary artery disease. AB - BACKGROUND: Although blood pressure is a major determinant of myocardial oxygen demand, little information is currently available regarding the changes in blood pressure (BP) during myocardial ischemia. Since BP elevation may cause left ventricular (LV) wall stress and an increase in oxygen demand, infusion of an alpha-adrenergic agonist, such as phenylephrine (PH), may provoke changes in myocardial perfusion in coronary artery disease (CAD) patients. As the effects of BP changes alone on myocardial perfusion have never been assessed by thallium-201 (Tl) scintigraphy, we investigated the effects of BP elevation after PH infusion, in order to study the hypothesis that pressure loading alone without increases in heart rate, may provoke transient impairment of regional myocardial perfusion, in CAD patients. PATIENTS AND METHODS: Forty-one (41) patients with angiographically documented CAD, aged 54+/-8 years, were included in our study. Each patient was given, without any complications, a PH (0.1 mg/ml) dose infused at a rate of 0.8 ml/mm, determined by a standardisation procedure and producing a mean blood pressure elevation of approximately 30% above baseline levels and a heart rate response to levels of no less than 50 bpm. One minute after the desired blood pressure and heart rate responses were reached, 2 mCi of Tl were injected and the PH infusion continued until the termination of the test. Tl scintigraphy was performed both 2 min after Tl injection and 4 h later, while the results were correlated to coronary angiography findings. RESULTS: PH scintigraphy produced 152 total defects. The mean perfusion defect size (%) was 14+/-12 and was directly related to the number of diseased vessels, i.e., 2% for one-vessel disease, 15% for two-vessel disease and 25% for three-vessel disease (P<0.05). The lowest percentage Tl activity values were 56+/-14 and were inversely related to the number of diseased vessels (P<0.01). The mean Tl lung counts/pixel values were 25+/-8 while it increased as the number of diseased vessels increased (P<0.01). The mean lung/heart ratio values were 0.31+/-0.08 while it increased as the number of diseased vessels increased (P<0.01). CONCLUSION: BP elevation after PH loading, produces a significant impairment of myocardial perfusion, that correlates well with the extend of angiographic findings. PMID- 12127372 TI - Basic fibroblast growth factor changes in response to coronary angioplasty in patients with stable angina. AB - Basic fibroblast growth factor (bFGF) has been implicated in the pathogenesis of coronary atherosclerosis and in angiogenesis. We assessed the changes in serum bFGF before, immediately after, and 6 months after coronary angioplasty (PTCA). Using the ELISA methods we measured plasma bFGF in 28 patients who underwent PTCA, in 20 patients with coronary artery disease who underwent elective coronary angiography and in 28 healthy subjects. Before PTCA and coronary angiography, bFGF plasma levels were similar in both patient groups (4.4+/-1.0 vs. 3.3+/-0.5 pg/ml), but were significantly higher compared with those of the control group (0.8+/-0.1 pg/ml, P<0.05). By 24 h, 3 months and 6 months after PTCA, bFGF levels had decreased significantly in the PTCA group (3.2+/-0.6, 1.7+/-0.3 and 2.7+/-0.6 pg/ml, respectively, P<0.05). In conclusion, these findings show that bFGF levels are elevated in patients with coronary artery disease. Following PTCA, bFGF levels decreased significantly and remained stable for 6 months after the procedure. Thus, bFGF level may change in response to PTCA in patients with coronary artery disease and stable angina. PMID- 12127374 TI - Spread of heart diseases seen in an open-access paediatric echocardiography clinic. AB - All patients <13 years of age referred to the open-access paediatric echocardiography clinic at the Sultan Qaboos University Hospital, Muscat, Oman, during the five years from 1994 to 1998 were analysed. Among the 2633 patients studied, 1543 (58.6%) were normal, 845 (32.1%) had congenital heart disease, and 245 (9.3%) had acquired heart disease. The major congenital heart diseases identified were secundum atrial septal defect (22.5%), ventricular septal defect (22.5%), patent ductus arteriosus (15.7%), mitral valve prolapse (10.7%), pulmonary stenosis (9.7%) and atrioventricular septal defect (4.5%). Fifty-eight percent of the congenital heart diseases were identified in the first year of life. Among the acquired heart diseases, rheumatic heart disease (30.2%) and cardiac involvement secondary to haemoglobinopathies (16.7%), dilated cardiomyopathy (16.3%) and hypertrophic cardiomyopathy (12.7%) were significant. Although the presence of specific cardiac symptoms was associated with a high yield of abnormalities, such disorders were also discovered in a significant number of children with isolated cardiac murmur. The referral source did not influence significantly the frequency of heart diseases diagnosed in this study. Open-access echocardiography is important in early detection of heart disease in the paediatric population. PMID- 12127375 TI - Prognostic value of 2D echocardiography in patients presenting with acute chest pain and non-diagnostic ECG for ST-elevation myocardial infarction. AB - The purpose of the present study was to test the hypothesis that early detection of regional wall motion abnormalities (WMA) by 2D echocardiography (ECHO) accurately predicts further cardiac events in patients presenting with acute chest pain. A prospective analysis was performed in subjects admitted with the first presentation of acute chest pain and a non-diagnostic ECG for acute ST elevation myocardial infarction. Patients with known coronary artery disease were excluded. All subjects were contacted by phone for a 30days follow-up regarding cardiac events defined as PCI/CABG, AMI, and death. In 132 consecutive patients (89 male, 43 female) complete data sets consisting of case history (H; abnormal: typical angina), ECG (abnormal: ST-depression, T-inversion, atypical ST elevation, LBBB), serum markers (TnI; abnormal: elevation of troponin I=0.5 ng/ml), ECHO (abnormal: WMA) and follow-up were available. In 45 patients, 60 cardiac events occurred (three deaths, 24 AMI, 33 PCI/CABG). Positive (PPV; %) and negative predictive values (NPV; %) of ECHO were superior to all other diagnostic tests (P<0.05 each) for adverse cardiac events, evolving AMI or death, and superior to history and ECG for later need of revascularisation (PCI/ACVB). Multivariate analysis revealed that WMA in ECHO predict cardiac events independently of age, gender, and the common combination of investigations (H/ECG/TnI). A significant independent impact of ECHO was also determined for the prediction of AMI/death or PCI/CABG. The study shows that early 2D echocardiography provides superior prognostic information concerning the risk of subsequent complications in patients with acute chest pain and a non-diagnostic ECG for ST-elevation-AMI. PMID- 12127376 TI - Effects of short-term atorvastatin treatment on global fibrinolytic capacity, and sL-selectin and sFas levels in hyperlipidemic patients with coronary artery disease. AB - BACKGROUND: The beneficial effects of HMG-CoA reductase inhibitors (statins) in patients with coronary artery disease (CAD) appear to be attributable not only to their lipid-lowering properties, but also to their therapeutic effects on the coagulation system, and anti-inflammatory effect. Furthermore, statins mitigate the apoptosis of vascular smooth muscle cells (VSMC) and macrophages in atherosclerotic plaques. HYPOTHESIS: The purpose of this study was to evaluate the effects of short-term atorvastatin treatment on the fibrinolytic system and systemic inflammatory status, and on apoptosis in hyperlipidemic patients with CAD. METHODS: The study population consisted of 36 hyperlipidemic patients (14 women and 22 men, mean age 53+/-9 years) with stable CAD, untreated with lipid lowering medications. Serum lipoproteins, fibrinogen levels, sFas and sL selectin, and global fibrinolytic capacity (GFC) were measured at baseline and after 12 weeks of treatment with atorvastatin, 10 mg/day. RESULTS: Atorvastatin treatment decreased serum low-density lipoprotein (-39%, P=0.0001), total cholesterol (-32%, P=0.0001), and triglycerides (-22%, P=0.0001), and increased high-density lipoprotein (+13%, P=0.0001) at 12 weeks compared to baseline. These effects were associated with a decrease in plasma fibrinogen from 331+/-73 to 298+/-58 mg/dl (P=0.0001), and sL-selectin levels from 666+/-201 to 584+/-162 ng/ml (P=0.0001). sFas levels and GFC increased from 3754+/-1264 to 4873+/-1835 pg/ml and from 3.5+/-2.4 to 5.6+/-2.9 microg/ml, respectively (both P=0.0001). CONCLUSIONS: These results suggest that lipid lowering with atorvastatin therapy significantly increases GFC, decreases fibrinogen levels, and causes leukocyte deactivation. Our findings also suggest that atorvastatin treatment mitigates apoptosis of VSMC in the atherosclerotic plaque. These effects of atorvastatin may, in part, explain the early decrease in cardiovascular events observed in clinical trials of statins. PMID- 12127378 TI - The effect of the localization of Q wave myocardial infarction on ventricular electromechanics. AB - BACKGROUND: The exact location of a Q wave myocardial infarction has an important effect on overall left ventricular function. OBJECTIVES: To assess the effect of localization of Q wave infarction on left ventricular minor and long axis function, with particular reference to electromechanical disturbances. METHODS: We studied 72 patients with Q wave myocardial infarction; 35 anterior, age 61+/ 15 years and 37 inferior, age 62+/-12 years. ECG intervals were automatically measured by Hewlett-Packard Pagewriter and LV dimension and filling velocities studied by transthoracic echocardiography and simultaneous phonocardiogram. Findings were compared with 21 controls of similar age. RESULTS: Heart rate and all ECG intervals were similar in the two patient groups and controls. QRS axis was more to the left in patients with inferior MI. Normal septal q wave was absent in lead V5 and V6 in 33/35 (94%) patients with anterior MI and in only 3/37 (8%) with inferior MI, p<0.001. LV minor axis dimensions were enlarged vs. normal (p<0.001) in the two patient groups and to a greater extent in anterior MI compared with inferior MI, p<0.05. Isovolumic relaxation time was prolonged only in-patients with an inferior MI, p<0.01. Long axis amplitude was globally reduced (p<0.001) in the two patient groups as were shortening and lengthening velocities (p<0.001). The onset of septal long axis shortening with respect to the q wave was delayed by 30 and 40 ms in inferior MI and anterior MI and that of lengthening with respect to A2 by 20 and 30 ms, respectively, compared to normal (p<0.001 for both). Post ejection shortening was localized to the septal long axis in 32/35 patients with anterior MI but was generalized involving all three LV long axes in inferior MI, p<0.001. Transmitral Doppler flow velocities and the frequency of mild mitral regurgitation were similar in the two groups. CONCLUSION: These results confirm a close association between anterior Q wave infarction, septal incoordination and absent septal q waves. The global incoordinate long axis behaviour in inferior Q wave MI may be due to significant papillary muscle dysfunction, and results in significant shape change in early diastole. This disturbance in electromechanical behaviour might play an important role in the differing outcomes between the two different sites of myocardial infarction. PMID- 12127377 TI - Exercise training following myocardial infarction improves myocardial perfusion assessed by thallium-201 scintigraphy. AB - AIM: We assessed the effects of a 6-week exercise programme on the thallium-201 myocardial perfusion characteristics of patients following myocardial infarction. METHODS: Twenty-five patients presenting with a first acute myocardial infarction were randomised into two groups: (i) those undergoing a supervised exercise training programme over 6 weeks (n=15) and (ii) a control group who did not attend the exercise programme (n=10). All underwent three sequential stress thallium myocardial perfusion scans at 10 days, 6 weeks and 3 months after infarction. The stress conditions were identical on each occasion. The images were analysed using a polar plot with a computer assisted algorithm comparing stress and redistribution data. Values for extent, severity and percentage redistribution of the thallium images were generated. RESULTS: A total of 29 perfusion defects were identified, 18 in the exercise group and 11 in the control group. Over 3 months in the exercise group the mean extent of the stress image defect fell from 109+/-64 to 95+/-51 pixels (P<0.05) while in the control group there was an increase from 133+/-57 to 144+/-57 pixels (P=ns). Stress defect severity fell in the exercise group from 581+/-417 to 494+/-346 S.D. (P<0.05) but increased in the control group from 765+/-494 to 877+/-543 S.D. (P=ns). On redistribution imaging in the exercise group a significant decrease was observed in both extent (94+/-56 to 76+/-43 pixels (P<0.05)) and severity (541+/-387 to 438+/-291 S.D. (P<0.05)) of the defects. However in the control group no significant change was observed for extent (125+/-54 to 125+/-52 pixels) or severity (745+/-485 to 820+/-503 S.D.) of the redistribution defects (P=ns). Reversibility of the defects increased slightly in both the exercise group (from 14.6+/-17 to 17.5+/-20%) and the control group (5.2+/-5 to 9.6+10%) (P=ns). CONCLUSION: Following myocardial infarction a 6-week exercise programme improves myocardial perfusion characteristics. An exercise programme should be integrated into cardiac rehabilitation protocols for patients after infarction. PMID- 12127379 TI - Transcardiac gradient of soluble adhesion molecules predicts progression of coronary artery disease. AB - BACKGROUND: During the development of atherosclerotic lesion, several types of cellular adhesion molecules (CAMs) are overexpressed on the surface of vascular endothelium. Some parts of these membrane proteins are proteolysed and are detected in blood as soluble forms. AIMS: The purpose of this study is to investigate the relationship between the transcardiac gradient of soluble cellular adhesion molecules (sCAMs) and the clinical characteristics of coronary artery disease (CAD). METHODS: We studied 46 patients with clinically stable CAD. Serum sCAM levels in both aortic sinus and coronary sinus were measured using enzyme-linked immunosorbent assay, and the transcardiac gradient of sCAMs was calculated. We also evaluated the angiographic severity of CAD, response of coronary artery to acetylcholine (Ach), as well as progression of coronary atherosclerosis over a 6-month period. RESULTS: The transcardiac gradient of sCAMs did not correlate to the angiographic severity of coronary atherosclerosis. The transcardiac gradient of sVCAM-1 was significantly higher in patients with vasoconstrictive response to Ach than patients without vasoconstrictive response to Ach (191.5+/-98.2 vs. -9.2+/-14.1 ng/ml, P<0.05). Furthermore, patients who exhibited progression of coronary atherosclerosis had a higher transcardiac gradient of sVCAM-1 at the initial study than patients without progression (47.8+/-24.5 vs. -6.4+/-12.3 ng/ml, P<0.05). CONCLUSIONS: An elevated transcardiac gradient of sVCAM-1 may represent the persistent activation of coronary artery that is accompanied by endothelial dysfunction, and may be a predictive index of progression of coronary atherosclerosis. Measurement of coronary circulating sVCAM-1 could provide important functional and predictive information about atherosclerosis. PMID- 12127380 TI - Internists in adolescent medicine. PMID- 12127381 TI - Maintenance of lightweight correlates with decreased cardiovascular risk factors in early adolescence. AB - PURPOSE: To describe predictors of lightweight in 11-year-olds and how weight changes between ages 9-11 years affected selected cardiovascular risk factors. METHODS: Cohort study among an ethnically and geographically diverse group of 5098 nine-year-olds who participated in the Child and Adolescent Trial for Cardiovascular Health (CATCH) and were followed from 1991 to 1994 until age 11 years. Lightweight (body mass index [BMI] <15th percentile) was defined from gender- and age-adjusted data on the total population in the National Health and Nutrition Examination Survey I of 1971 to 1973. Weight (BMI) category at follow up (age 11 years) was the dependent variable in the logistic regression. Gender, race/ethnicity, school site, weight category at baseline, and participation in the CATCH (intervention or control group) were examined for possible predictors. RESULTS: The prevalence of lightweight was much lower than the 15% expected at both ages 9 and 11 years; 4.7% for males (M) and 7.4% for females (F) and at age 11 years; 5.9% for M and 8.3% for F. Lightweight at age 9 years was the best predictor of lightweight at age 11 years (OR = 56.8; CI 41.3, 78.2). Normal weight children who entered the overweight category exhibited striking adverse percent changes in serum cholesterol and in both serum high-density cholesterol and apolipoprotein B cholesterol levels. Among children in each weight group at age 9 years, the heavier children at age 11 years had higher percent changes in systolic blood pressure, and lower times on the 9-min run. Maintenance of lightweight was associated with the most favorable pattern of cardiovascular risk factors. Differences between groups were not found for percent change in total calorie intake or percentage of calories from fat. Normal-weight children who became overweight exhibited striking percentage increases in levels of serum cholesterol and in serum apolipoprotein B cholesterols. CONCLUSIONS: Population based anticipatory guidance about weight should focus on all children in mid childhood, not only those at the upper extremes of weight. These who are genetically lean and healthy should be encouraged to remain so. Maintenance or achievement of lightweight was associated with favorable changes in systolic blood pressure in CATCH children during early adolescence. Those below the 5th percentile should be assessed and treated if eating disorders, undernutrition, or chronic illness are present. Optimal weight and optimal health should not be confused. PMID- 12127382 TI - A school-based intervention can reduce body fat and blood pressure in young adolescents. AB - PURPOSE: To determine the effect of increasing the aerobic component of the school's physical activity program and improving the knowledge about weight control and blood pressure on the blood pressure and body fat of early adolescents. METHODS: The subjects were 1140 youth aged 11 to 14 years (630 females, 510 males; 64% white, 24.4% African-American, and 11.6% "other"), who were randomly assigned by school into four treatment groups: exercise only, education only, exercise and education combined, and control group. Heights, weights, and skinfold thicknesses were measured, and body mass index (BMI) was computed kg/m(2). Blood pressure was obtained in duplicate using a random-zero mercury sphygmomanometer. Maximal oxygen uptake was predicted from a submaximal cycle ergometer test. Data were analyzed using analysis of covariance statistics, adjusting for gender, ethnicity, age, socioeconomic status, and initial baseline characteristics. RESULTS: Systolic and diastolic blood pressures increased more in the control group than in the intervention groups (p =.001). The BMI did not change significantly (p =.709), but the sum of skinfolds increased less in subjects in the exercise intervention groups than the education only or control groups (p =.0001). The small increase in (p)VO(2)max of the combined exercise and education group was significantly greater than the education only group (p =.0001). CONCLUSION: An exercise program for youth can have a positive effect on blood pressure independent of body weight loss. PMID- 12127383 TI - Weight management goals and practices among U.S. high school students: associations with physical activity, diet, and smoking. AB - PURPOSE: To examine associations of physical activity, fruit and vegetable consumption, and cigarette smoking with weight management goals and practices of U.S. high school students. METHODS: Data were from the 1999 national Youth Risk Behavior Survey, a representative sample of U.S. high school students (n = 15,349). Adjusted odds ratios (OR) were calculated to describe associations, controlling for demographic characteristics. RESULTS: Based on self-reported height and weight, 25% of students were either overweight (11%) or at risk for becoming overweight (14%). However, 43% of students were trying to lose weight and 19% of students were trying to maintain their current weight. Female students were less likely than male students to be overweight, but more likely to be trying to lose weight. Trying to lose weight was associated with vigorous physical activity (OR = 1.5), strengthening exercises (OR = 2.2), and cigarette smoking (OR = 1.4) among female students; and vigorous physical activity (OR = 1.6), strengthening exercises (OR = 1.8), and eating > or =5 servings/day of fruits and vegetables (OR = 1.5) among male students. Among students trying to lose weight or stay the same weight, only 62% of females and 41% of males combined exercise with a reduced fat and calorie diet, while 32% of females and 17% of males used unhealthy weight control methods (fasting, diet pills, vomiting, or laxatives). CONCLUSIONS: Efforts to promote healthy weight management among adolescents are needed and should place greater emphasis on combining physical activity with a reduced fat and calorie diet, increasing fruit and vegetable consumption, and discouraging smoking and other unhealthy weight control practices. PMID- 12127384 TI - Unhealthy behaviors and psychosocial difficulties among overweight adolescents: the potential impact of familial factors. AB - PURPOSE: To compare overweight and non-overweight youth on a selection of self reported eating, physical activity, dieting, educational, and emotional variables and identify familial factors that serve as protective forces against unhealthy behaviors and psychosocial difficulties among overweight adolescents. METHODS: Data were taken from a 1996 cross-sectional school-based survey of 9957 adolescents in grades 7, 9, and 11. Based on self-reported heights and weights, respondents were categorized as "overweight" (body mass index) > or =85th percentile or "non-overweight." Student's t-tests were used to compare the non overweight and overweight sample on the self-reported health-related behaviors and psychosocial variables. Logistic and linear regressions were used to identify familial factors associated with a reduced risk of engaging in unhealthy behaviors and experiencing psychosocial distress. RESULTS: Overweight adolescents reported engaging in significantly more unhealthy behaviors and experiencing more psychosocial distress than their non-overweight peers. Among the overweight youth, higher levels of reported family connectedness and parental expectations and moderate levels of parental monitoring were associated with the lowest levels of unhealthy behaviors and psychosocial distress. CONCLUSIONS: Satisfying and developmentally appropriate parent-adolescent relationships are associated with reduced behavioral and psychosocial risk factors associated with overweight during adolescence. PMID- 12127385 TI - Supplementation with omega-3 polyunsaturated fatty acids in the management of recurrent migraines in adolescents. AB - PURPOSE: To examine whether dietary supplementation with fish oil rich in very long-chain n-3 polyunsaturated fatty acids might reduce frequency and severity of migraines in adolescents. METHODS: Twenty-seven adolescents suffering from frequent migraines for at least 1 year (mean 4 +/- 1 years since migraine onset) participated in a randomized, double-blind, cross-over study consisting of 2 months of fish oil, 1-month washout period, and 2 months of placebo (olive oil). Participants self-assessed severity and duration of headache episodes (7-point faces and 10-point visual analog pain scales, 5-point frequency and severity rating scale) throughout the study. At the end of every 2-month treatment period, participants rated the effectiveness of treatment on a 7-point Likert scale (1,"not effective, not worthwhile"; 4,"moderately effective, moderately worthwhile"; 7, "totally effective, totally worthwhile"). A score of > or = 4 on the Likert scale was considered as improvement. RESULTS: Twenty-three adolescents (16 girls, 7 boys, 18 Whites, 3 Hispanics, 1 African-American, 1 Cape Verdean, mean age 15 +/- 1 years) completed the study. Compared with frequency of headaches before the study (31 +/- 4 episodes/2 months), there was a significant (p <.0001) reduction in headache frequency during fish oil treatment (4 +/- 1 episodes/2 months) and during placebo (olive oil) treatment (4 +/- 1 episodes/2 months) but no significant (NS) difference between treatments. Likewise, self assessment on a 7-point faces pain scale revealed a significant reduction in headache severity during fish oil treatment (2.9 +/- 0.5, p =.01) and during placebo (olive oil) treatment (3.5 +/- 0.4,

or =10mg/kg) caused a sustained increase in canalicular bile salt-independent bile flow, combined with significant increases in the concentration and output of glutathione and of bicarbonate in bile. In rats receiving bosentan (> or =10mg/kg), both under basal conditions and under intravenous taurocholate perfusion (2micromol/min/kg), phospholipid and cholesterol secretions were profoundly inhibited and uncoupled from bile salt secretion. In TR(-) rats, the choleretic effect of bosentan was reduced to non-significant levels. The stimulation of bilirubin secretion and the uncoupling of phospholipid from bile salt secretion were absent, whereas that of cholesterol was maintained. CONCLUSIONS: Bosentan alters canalicular bile formation in major part via mrp2 mediated mechanisms. Intermittent uncoupling of lipid from bile salt secretion may contribute to bosentan hepatic adverse reaction. PMID- 12127423 TI - Cystic fibrosis transmembrane conductance regulator (CFTR) gene defects in patients with primary sclerosing cholangitis. AB - BACKGROUND/AIMS: Because biliary tract lesions that resemble those of primary sclerosing cholangitis (PSC) may occur in cystic fibrosis (CF), we examined the prevalence and influence of CF transmembrane conductance regulator (CFTR) gene mutations in PSC patients. METHODS: Genomic DNA was analyzed in 29 consecutive PSC patients and in 115 healthy control individuals. A scanning method followed by direct DNA sequencing was used to scan the CFTR coding regions. RESULTS: Four patients (13.8%) were heterozygous for a CFTR mutation, including a new putative severe CF-causing mutation (N782K), and three mild defects (L997F, D1270N, and S1235R). The comparison of PSC patients with healthy controls showed no significant difference in the frequency of CFTR mutations (P=0.415). In addition, two patients (6.9%) were heterozygous for the IVS8-5T allele, which is not significantly different from the 5-6%-prevalence in the general population. Unusual clinical features including a severe outcome in childhood, with a lethal outcome at age 22, and biliary aspergillosis were recorded in patients with a CFTR mutation. CONCLUSIONS: The proportion of CF carriers is not significantly higher in PSC patients than in the general population. The possibility that CFTR mutations may contribute to a severe clinical course in PSC patients is worth further examining. PMID- 12127425 TI - Leptin is essential for the hepatic fibrogenic response to chronic liver injury. AB - BACKGROUND/AIMS: Obesity is associated with hyperleptinemia and is also a risk factor for fibrosis and severity of fibrosis in several chronic liver diseases. The correlation between increased leptin, obesity and hepatic fibrosis prompted us to hypothesise that leptin has profibrogenic effects on the liver. METHODS: We analysed the role of leptin in liver fibrosis in leptin-deficient mice fed a diet which generates steatohepatitis, and in chronic carbon tetrachloride-induced hepatic injury. RESULTS: Leptin-deficient mice failed to develop fibrosis during steatohepatitis or in response to chronic toxic liver injury, and failed to up regulate collagen-I while developing similar hepatic injury as their genetic controls. Restitution of physiological levels of circulating leptin by injection of exogenous leptin, but not correction of the obese phenotype by dietary manipulation, restored liver fibrosis in leptin-deficient mice during chronic liver injury. These results confirmed the absolute requirement of leptin for hepatic fibrosis. We showed that leptin deficiency did not alter hepatic TNF regulation but that leptin is necessary for induction of bioactive transforming growth factor beta 1 (TGFbeta1) protein in the context of chronic liver injury. CONCLUSIONS: These data establish that leptin is an essential mediator of hepatic fibrosis in response to chronic liver injury, whether metabolic or toxic in aetiology. PMID- 12127424 TI - Hepatobiliary organic anion transporters are differentially regulated in acute toxic liver injury induced by carbon tetrachloride. AB - BACKGROUND/AIMS: Hepatobiliary transporters are down-regulated in cholestasis, but their expression in acute, non-cholestatic, cytokine-mediated liver injury is unknown. Thus we studied the molecular mechanisms, by which sodium taurocholate cotransporting polypeptide (Ntcp), organic anion transporting polypeptide 1 (Oatp1), Oatp2, Oatp4, multidrug-resistance protein 2 (Mrp2) and bile salt export pump (Bsep) are regulated in liver injury induced by carbon tetrachloride (CCl(4)). METHODS: mRNA and protein levels were determined in rats 24 and 72h after CCl(4) injection. Transporter gene transcription and binding activities of Ntcp and Mrp2 transactivators were assessed by nuclear runoff and electrophoretic mobility shift assays. RESULTS: mRNA levels significantly declined to 41+/-44% for Ntcp, 65+/-41% for Oatp1 and 64+/-28% for Oatp2, but remained unchanged for Oatp4, canalicular Mrp2 and Bsep. Protein levels declined only for Oatp4 (-50+/ 17%) and Ntcp (-23+/-13%) at 24h. Reduced mRNA levels (Ntcp, Oatp1, Oatp2) were associated with decreased transcriptional activities. Binding activity of Ntcp transactivators (hepatocyte nuclear factor 1 alpha (HNF1alpha) and CAAT enhancer binding protein alpha (C/EBPalpha) were reduced by 24h, whereas retinoid X receptor alpha (RXRalpha):retinoid acid receptor alpha (RARalpha) as transactivator of both Ntcp and Mrp2 remained unaltered. Recovery of acute hepatitis and changes in gene expression occurred after 72h. CONCLUSIONS: Acute liver injury results in down-regulation of basolateral organic anion transporters similar to liver regeneration after partial hepatectomy, but in contrast to endotoxin-induced cholestasis. Maintained binding activity of RXRalpha:RARalpha may explain differences in Mrp2 expression. PMID- 12127426 TI - Reversal of activation of human myofibroblast-like cells by culture on a basement membrane-like substrate. AB - BACKGROUND: Liver injury transforms hepatic stellate cells into myofibroblast (MFB)-like cells. With recovery from injury, MFBs undergo apoptosis, but it is unknown whether they can also revert to quiescence. AIM: To determine whether human (h)MFBs become quiescent if cultured on a basement membrane-like substrate (Matrigel). METHODS: hMFBs obtained from cirrhotic liver were re-cultured on plastic or Matrigel. Expression of genes of collagen metabolism was assayed before and after transforming growth factor beta (TGFbeta) and Oncostatin M (OSM) stimulation. RESULTS: hMFBs had typical MFB-like morphology, with abundant alpha smooth muscle actin (SMA) but no cytoplasmic lipid droplets. hMFBs re-cultured on Matrigel reverted to alphaSMA-negative, lipid droplet-positive quiescent morphology. alphaSMA, collagen alpha1(1) (COL1A1) and collagen alpha2(1) (COL1A2) messages were upregulated in hMFBs cultured on plastic, but suppressed by Matrigel. The opposite was true for metalloproteinase-1 mRNA. OSM but not TGFbeta reduced alphaSMA mRNA by 30% while TGFbeta but not OSM upregulated COL1A1 mRNA by 48%, in hMFBs on plastic. TGFbeta and OSM stimulated COL1A1 gene expression in Matrigel by 50 and 60%, respectively. CONCLUSIONS: Matrigel culture de-activates hMFBs yet collagen gene expression still responds to fibrogenic cytokines. The responses of hMFB gene expression to TGFbeta and OSM, are regulated differently by the extracellular matrix. PMID- 12127427 TI - Inhibition of rat hepatocellular carcinoma tumor growth after multiple infusions of recombinant Ad.AFPtk followed by ganciclovir treatment. AB - BACKGROUND/AIMS: The antitumor efficiency of thymidine kinase (tk) in Herpes Simplex virus-tk-based gene therapy of rat hepatocellular carcinoma (HCC) was examined by specific transcriptional targeting of tk to tumor cells by the alpha fetoprotein (AFP) gene promoter and by multiple infusions of recombinant adenovirus Ad.AFPtk. METHODS: We developed a surgical procedure that allows efficient, non-invasive delivery (during 2 months) of recombinant Ad via the intra-hepatic artery (IHA) route. RESULTS: Treatment of tumor-bearing rats with either three or five doses of 5x10(9)pfu Ad.AFPtk, administered every 3 days, and followed by intra-peritoneal treatment with ganciclovir (GCV), resulted in tumor growth inhibition and apoptosis, when compared to untreated tumor-bearing rats or animals treated with Ad.AFPlacZ or buffered saline. No treatment-related toxicity was noted. Antitumor efficacy, based on tumor size and number of tumors, was demonstrated in more than 50% of Ad.AFPtk+GCV-treated rats, as compared to control rats (P<0.0005). CONCLUSIONS: Our results demonstrate the safety and potential of multiple Ad.AFPtk administrations by the IHA route to inhibit HCC tumor growth, and support further clinical investigation of Ad.AFPtk gene therapy for treatment of multifocal tumor lesions in most primary liver cancers. PMID- 12127428 TI - Mutation of p53 gene in regenerative nodules in cirrhotic liver. AB - BACKGROUND/AIMS: Mutations of p53 gene have been detected in precancerous stages of several cancers, and the possible role in multistep carcinogenesis is suggested. The aim of this study was to examine the mutation profile of p53 gene in regenerative nodules in cirrhotic livers. METHODS: Ninety eight tissue specimens of regenerative nodules obtained from 15 cases of cirrhosis were used for analysis. Twenty cases of chronic hepatitis and two cases of fatty liver were used as controls. DNA was extracted from each of manually demarcated regenerative nodules, and nucleotide sequence analysis was performed on p53 gene exon 5. RESULTS: Direct sequencing detected p53 mutations in seven of 98 DNA samples (7.1%) from regenerative nodules in six cases of cirrhosis. Subcloning analysis revealed that mutation sites differed in each subclone and the incidences of the mutation varied from 7.7 to 58.8% depending on individual nodules. The mutation was not detected in any of chronic hepatitis and fatty liver. There were inconsistent p53 sequence with regenerative nodules and accompanied hepatocellular carcinomas in six cases. CONCLUSIONS: Mutations of p53 gene were frequently found in cirrhotic livers compared with livers of patients with chronic hepatitis (P<0.01), suggesting that p53 mutations at the stage of cirrhosis may be a causative factor that may potentially lead to hepatocellular carcinoma. PMID- 12127429 TI - Mechanical stress-dependent secretion of interleukin 6 by endothelial cells after portal vein embolization: clinical and experimental studies. AB - BACKGROUND/AIMS: Interleukin-6 (IL-6) is an essential early signal in liver regeneration, however, little is known about what triggers IL-6 release. Changes in portal hemodynamics after portal vein embolization (PVE) may contribute to IL 6 release, leading to regeneration of non-embolized lobe. METHODS: In 22 patients who underwent right PVE, the diameters of the left portal branches, liver volumes, and serum concentrations of IL-6, tumor necrosis factor-alpha (TNF alpha), and hepatocyte growth factor (HGF) were measured. We then studied endothelial cells cultured on an elastic silicone membrane and subjected to continuous uni-axial stretch. Supernatant cytokine concentrations were measured. RESULTS: The diameters of the portal branches increased by 150% after PVE. Serum IL-6 concentrations increased within 3h after PVE. The concentrations of TNF alpha and HGF remained unchanged. The left lobe volume increased 2 weeks after PVE. The IL-6 concentrations in the supernatant of endothelial cells with stretch stress were higher than that in the non-stretched control group. CONCLUSIONS: These findings indicate that PVE dilates the portal branches in the non-embolized lobe, exposing hepatic vasculature to stretch stress. This hemodynamic change may act as a trigger for IL-6 release from endothelial cells and contribute to the activation of regenerative cascade in the non-embolized lobes. PMID- 12127430 TI - Treatment of recurrent hepatitis C in liver transplants: efficacy of a six versus a twelve month course of interferon alfa 2b with ribavirin. AB - BACKGROUND/AIMS: Interferon (IFN) with ribavirin combination therapy (CT) was proposed for the treatment of hepatitis C recurring in liver transplants. We assessed the efficacy of two protocols of CT in transplanted patients with recurrent severe hepatitis C virus (HCV) hepatitis. METHODS: Fifty-seven patients (68% genotype 1b) were treated with IFN alfa-2b 3 million units three times weekly and oral ribavirin 800mg/die for 6 or 12 months. Study end-points were the end of treatment (ETVR) and the 12-month post-therapy sustained virologic response (SVR; negative HCV-RNA). RESULTS: ETVR was induced in 9/27 (33%) and in 7/30 patients (23%) treated, respectively, for 6 and 12 months (P=0.4); a SVR was induced in six (22%) of the former and five (17%) of the latter (P=0.4). HCV genotype non-1 patients responded better than genotype 1 (SVR: 43% in genotype non-1 versus 12% in genotype 1, P: 0.02). In ETV responders the hepatitis activity index improved by >2 points in biopsies taken after therapy compared to pre-therapy biopsies. Anemia and leukopenia required reduction of therapy in 51% of the patients. CONCLUSIONS: CT is efficacious in controlling HCV disease in about 20% of transplants with recurrent hepatitis C. Six months of therapy are as efficacious as 12 months. PMID- 12127431 TI - The influence of baseline characteristics on viral dynamic parameters in chronic hepatitis B patients treated with lamivudine. AB - BACKGROUND/AIMS: Viral decline during lamivudine therapy in chronic hepatitis B patients is bi-phasic. We studied the influence of lamivudine dose and baseline characteristics on parameters obtained from a mathematical model. METHODS: Chronic hepatitis B patients were randomized to receive 150 mg (group 1; n=11) or 600 mg (group 2; n=10) lamivudine daily for 4 weeks. Hepatitis B virus DNA was measured frequently with the Digene Hybrid Capture II test and the Roche PCR assay. RESULTS: The description of viral decline in our closely monitored patients by means of the mixed-effects approach with both the bi-phasic model and a piecewise linear regression model resulted in a good fit. Baseline alanine aminotransferase (ALT) was significantly related to the slope of the second phase of viral decline. Previous lamivudine-treated patients showed a significant slower first phase than patients naive to lamivudine treatment. CONCLUSIONS: The initial observed difference in viral decline between 150 and 600 mg of lamivudine disappeared when baseline ALT was taken into account. This strengthens the hypothesis that the level of intrinsic activity is related to the turnover of infected hepatocytes. Moreover, reintroduction of lamivudine in previously lamivudine-treated patients should be considered carefully. PMID- 12127432 TI - Detection of YMDD mutant using a novel sensitive method in chronic liver disease type B patients before and during lamivudine treatment. AB - BACKGROUND/AIMS: The emergence of lamivudine-resistant hepatitis B virus (HBV) was reported in patients with prolonged lamivudine administration. There was no report of the existence of tyrosine-methionine-aspartate-aspartate (YMDD) mutant in non-lamivudine treated chronic hepatitis B patients. In the present study, we developed a sensitive assay and applied it to the detection of YMDD mutant. METHODS: We developed peptide nucleic acid (PNA) mediated polymerase chain reaction clamping for detecting mutations in a YMDD motif of the hepatitis B virus DNA polymerase gene. We studied YMDD mutants in a patient with HBV DNA breakthrough longitudinally and in non-lamivudine treated patients (36 patients). RESULTS: We could detect as little as 0.01-0.001% of mutant viruses coexisting in 10(5)-10(9) copies of wild-type viruses using this assay. YMDD mutant was detected 7 months before clinical breakthrough, which was 6 months earlier than using the conventional restriction fragment length polymorphism assay. YMDD mutants were also detected in four of 18 anti-HBe antibody positive untreated chronic hepatitis type B: YMDD+tyrosine-valine-aspartate-aspartate (YVDD) in two patients and YMDD+tyrosine-isoleucine-aspartate-aspartate (YIDD) in two patients, however, none in HBe antigen positive patients. CONCLUSIONS: We developed a highly sensitive assay for detecting YMDD mutants. This is an effective procedure for monitoring patients during or before lamivudine treatment and can provide more insights into the therapeutic strategies for chronic hepatitis B patients. PMID- 12127433 TI - Famciclovir treatment of chronic delta hepatitis. AB - BACKGROUND/AIMS: Interferon is the only established therapy for chronic delta hepatitis and alternative treatment options are an urgent need. Since successful treatment of a case of post-transplant delta hepatitis with the nucleoside analogue famciclovir had been reported, a pilot study was undertaken to evaluate the use of famciclovir in the treatment of chronic delta hepatitis. METHODS: A total of 15 adult patients, 13 men, two women, ages 20-52 years, with chronic delta hepatitis were treated with famciclovir, 500 mg, three times a day for 6 months and were then followed-up for 6 months posttreatment. All patients had compensated chronic liver disease, elevated liver enzymes and were hepatitis delta virus (HDV) RNA positive by polymerase chain reaction at baseline. Patients were monitored and tested for HBsAg, hepatitis B virus (HBV) DNA and HDV RNA levels. Liver biopsies were obtained before starting famciclovir and within 1 month of completion of treatment. RESULTS: HBV DNA levels decreased in nine of the 15 patients and levels rose again after treatment (P<0.05). Famciclovir had no effect on alanine aminotransferase (ALT) and HBsAg levels or on serum HDV RNA and overall, there was no improvement in liver histology. CONCLUSIONS: Treatment of chronic delta hepatitis with famciclovir has no effect on disease activity and HDV RNA levels. PMID- 12127434 TI - Canalicular bile formation: beyond single transporter functions. PMID- 12127435 TI - Ribavirin and interferon combination for recurrent post-transplant hepatitis C: which benefit beyond 6 months? PMID- 12127436 TI - Hepatitis B virus viral dynamics: effects of drug dose and baseline alanine aminotransferase. PMID- 12127437 TI - Haemostatic abnormalities in patients with liver disease. PMID- 12127438 TI - Cholangiography in a patient with hilar peribiliary cysts. PMID- 12127439 TI - Nadolol-induced painful gingival bleeding. PMID- 12127440 TI - Fatal lactic acidosis in a HIV-positive patient treated with interferon and ribavirin for chronic hepatitis C. PMID- 12127441 TI - Hepatotoxicity of polyunsaturated fatty acids in alcohol abuser. PMID- 12127445 TI - RNA-protein interactions. AB - Recent discoveries have revealed that there is a myriad of RNAs and associated RNA-binding proteins that spatially and temporally appear in the cells of all organisms. The structures of these RNA-protein complexes are providing valuable insights into the binding modes and functional implications of these interactions. Even the common RNA-binding domains (RBDs) and the double stranded RNA binding motifs (dsRBMs) have been shown to exhibit a plethora of binding modes. PMID- 12127446 TI - The role of metal ions in RNA catalysis. AB - Understanding the catalytic mechanisms of RNA enzymes remains an important and intriguing challenge - one that has grown in importance since the recent demonstration that the ribosome is a ribozyme. At first, it seemed that all RNA enzymes compensate for the limited chemical versatility of ribonucleotide functional groups by recruiting obligatory metal ion cofactors to carry out catalytic chemistry. Mechanistic studies of the large self-splicing and pre-tRNA processing ribozymes continue to support this idea, yielding increasingly detailed views of RNA active sites as scaffolds for positioning catalytic metal ions. Re-evaluation of the methodologies used to distinguish catalytic and structural roles for metal ions, however, has challenged this notion in the case of the small self-cleaving RNAs. Recent studies of the small ribozymes blur the distinction between catalytic and structural roles for metal ions, and suggest that RNA nucleobases have a previously unrecognized capacity for mediating catalytic chemistry. PMID- 12127447 TI - RNA folding in vivo. AB - RNA folding in vivo is influenced by the cellular environment, the vectorial nature of transcription and translation, trans-acting factors and ion homeostasis. Specific RNA-binding proteins promote RNA folding by stabilizing the native structure or by guiding folding. In contrast, RNA chaperones, which are believed to interact nonspecifically with RNA, were proposed to resolve misfolded RNA structures and to promote intermolecular RNA-RNA annealing. Small trans acting noncoding RNAs are thought to modulate mRNA structures, thereby regulating gene expression. So far, there is some evidence that in vitro and invivo RNA folding pathways share basic features. However, it is unclear whether the rules deduced from in vitro folding experiments generally apply to invivo conditions. PMID- 12127448 TI - The accuracy of ribosomal RNA comparative structure models. AB - The determination of the 16S and 23S rRNA secondary structure models was initiated shortly after the first complete 16S and 23S rRNA sequences were determined in the late 1970s. The structures that are common to all 16S rRNAs and all 23S rRNAs were determined using comparative methods from the analysis of thousands of rRNA sequences. Twenty-plus years later, the 16S and 23S rRNA comparative structure models have been evaluated against the recently determined high-resolution crystal structures of the 30S and 50S ribosomal subunits. Nearly all of the predicted covariation-based base pairs, including the regular base pairs and helices, and the irregular base pairs and tertiary interactions, were present in the 30S and 50S crystal structures. PMID- 12127450 TI - Charge transport in DNA. AB - The base pair stack within double helical DNA provides an effective medium for charge transport. The DNA pi-stack mediates oxidative DNA damage over long molecular distances in a reaction that is exquisitely sensitive to the sequence dependent conformation and dynamics of DNA. A mixture of tunneling and hopping mechanisms have been proposed to account for this long-range chemistry, which is gated by dynamical variations within the stack. Electrochemical sensors have also been developed, based upon the sensitivity of DNA charge transport to base pair stacking, and these sensors provide a completely new approach to diagnosing single base mismatches in DNA and monitoring protein-DNA interactions electrically. DNA charge transport, furthermore, may play a role within the cell and, indeed, oxidative damage to DNA from a distance has been demonstrated in the cell nucleus. As a result, the biological consequences of and opportunities for DNA-mediated charge transport now require consideration. PMID- 12127449 TI - Protein surface salt bridges and paths for DNA wrapping. AB - The organization of large regions of DNA on the surface of proteins is critical to many DNA 'transactions', including replication, transcription, recombination and repair, as well as the packaging of chromosomal DNA. Recent thermodynamic and structural studies of DNA binding by integration host factor indicate that the disruption of protein surface salt bridges (dehydrated ion pairs) dominates the observed thermodynamics of integration host factor binding and, more generally, allows the wrapping of DNA on protein surfaces. The proposed thermodynamic signature of wrapping with coupled salt bridge disruption includes large negative salt-concentration-dependent enthalpy, entropy and heat capacity changes and smaller than expected magnitudes of the observed binding constant and its power dependence on salt concentration. Examination of the free structures of proteins recently shown to wrap DNA leads us to hypothesize that a pattern of surface salt bridges interspersed with cationic sidechains provides a structural signature for wrapping and that the number and organization of salt bridges and cationic groups dictate the thermodynamics and topology of DNA wrapping, which in turn are critical to function. PMID- 12127451 TI - Force spectroscopy of single DNA and RNA molecules. AB - Experiments in which single molecules of RNA and DNA are stretched, and the resulting force as a function of extension is measured have yielded new information about the physical, chemical and biological properties of these important molecules. The behavior of both single-stranded and double-stranded nucleic acids under changing solution conditions, such as ionic strength, pH and temperature, has been studied in detail. There has also been progress in using these techniques to study both the kinetics and equilibrium thermodynamics of DNA protein interactions. These studies generate unique insights into the functions of these proteins in the cell. PMID- 12127453 TI - Genomic insights into evolution. PMID- 12127452 TI - Residual dipolar couplings in nucleic acid structure determination. AB - Solution NMR spectroscopy of nucleic acids has been limited by the short-range nature of the nuclear Overhauser effect and scalar coupling restraints normally used in structure determination. The addition of residual dipolar couplings, obtained from slightly oriented mixtures, provides bond vector angles relative to a universal alignment tensor. The accurate determination of helix curvature, domain orientation and the stoichiometry of homomultimeric nucleic acid complexes is now possible. PMID- 12127454 TI - Protein structure prediction in 2002. AB - Central issues concerning protein structure prediction have been highlighted by the recently published summary of the fourth community-wide protein structure prediction experiment (CASP4). Although sequence/structure alignment remains the bottleneck in comparative modeling, there has been substantial progress in fully automated remote homolog detection and in de novo structure prediction. Significant further progress will probably require improvements in high resolution modeling. PMID- 12127455 TI - Functional transcriptomes: comparative analysis of biological pathways and processes in eukaryotes to infer genetic networks among transcripts. AB - Microarray technology enables us to monitor large changes in transcripts at any given time. The compilation of these data makes possible the comparison of such gene expression data on a genome-wide scale. As comparisons of genome sequence data yield new biological insights, comparative analyses of transcriptome data also promise new discoveries regarding metabolic pathways and cellular processes. The coordinated expression of genes shows that these genes physically interact with each other or are part of the same cascade. We have produced one of the largest expression profiles of adult mice and developmental tissues. These data, as well as the data on yeast from previous reports, were used to see whether coordinated expression (with high correlation coefficient) is closely coupled to the actual cascade on the pathway map. PMID- 12127456 TI - Automated detection of remote homology. AB - The classification of a newly identified protein as a member of a superfamily is important for focusing experiments on its most likely functions. Such classification, often performed by hand, has now been fully automated. This sophisticated new approach takes into account not only alignment scores but also a number of other computable attributes, such as functional sites deduced from sequence conservation patterns. PMID- 12127457 TI - Computational methods for the prediction of protein interactions. AB - Establishing protein interaction networks is crucial for understanding cellular operations. Detailed knowledge of the 'interactome', the full network of protein protein interactions, in model cellular systems should provide new insights into the structure and properties of these systems. Parallel to the first massive application of experimental techniques to the determination of protein interaction networks and protein complexes, the first computational methods, based on sequence and genomic information, have emerged. PMID- 12127458 TI - Pathway evolution, structurally speaking. AB - Small-molecule metabolism forms the core of the metabolic processes of all living organisms. As early as 1945, possible mechanisms for the evolution of such a complex metabolic system were considered. The problem is to explain the appearance and development of a highly regulated complex network of interacting proteins and substrates from a limited structural and functional repertoire. By permitting the co-analysis of phylogeny and metabolism, the combined exploitation of pathway and structural databases, as well as the use of multiple-sequence alignment search algorithms, sheds light on this problem. Much of the current research suggests a chemistry-driven 'patchwork' model of pathway evolution, but other mechanisms may play a role. In the future, as metabolic structure and sequence space are further explored, it should become easier to trace the finer details of pathway development and understand how complexity has evolved. PMID- 12127459 TI - Structural genomics of proteins from conserved biochemical pathways and processes. AB - During the past year, X-ray crystallographers and solution NMR spectroscopists have made significant progress towards the complete structural characterization of conserved biochemical pathways and processes. Some of these advances were made in the context of nascent structural genomics programs, which promise to accelerate structural studies of biologically and medically important proteins. The results of high-throughput protein production, crystallization, structure determination, homology modeling and functional annotation published by two such programs have provided insight into the evolution and function of enzymes in the isoprenoid biosynthesis and ribulose monophosphate pathways. PMID- 12127460 TI - Trends in protein evolution inferred from sequence and structure analysis. AB - Complementary developments in comparative genomics, protein structure determination and in-depth comparison of protein sequences and structures have provided a better understanding of the prevailing trends in the emergence and diversification of protein domains. The investigation of deep relationships among different classes of proteins involved in key cellular functions, such as nucleic acid polymerases and other nucleotide-dependent enzymes, indicates that a substantial set of diverse protein domains evolved within the primordial, ribozyme-dominated RNA world. PMID- 12127461 TI - Evolution of protein structures and functions. AB - Within the ever-expanding repertoire of known protein sequences and structures, many examples of evolving three-dimensional structures are emerging that illustrate the plasticity and robustness of protein folds. The mechanisms by which protein folds change often include the fusion of duplicated domains, followed by divergence through mutation. Such changes reflect both the stability of protein folds and the requirements of protein function. PMID- 12127462 TI - Did evolution leap to create the protein universe? AB - The genomes of over 60 organisms from all three kingdoms of life are now entirely sequenced. In many respects, the inventory of proteins used in different kingdoms appears surprisingly similar. However, eukaryotes differ from other kingdoms in that they use many long proteins, and have more proteins with coiled-coil helices and with regions abundant in regular secondary structure. Particular structural domains are used in many pathways. Nevertheless, one domain tends to occur only once in one particular pathway. Many proteins do not have close homologues in different species (orphans) and there could even be folds that are specific to one species. This view implies that protein fold space is discrete. An alternative model suggests that structure space is continuous and that modern proteins evolved by aggregating fragments of ancient proteins. Either way, after having harvested proteomes by applying standard tools, the challenge now seems to be to develop better methods for comparative proteomics. PMID- 12127463 TI - Brief interventions for problem drinking and women. AB - Early identification and intervention among problem drinking women may avert the more severe, adverse consequences of alcohol abuse and dependence. Screening and brief interventions, while generally effective, have not been adequately examined among subgroups, such as women. The purpose of this review article is to examine the efficacy of brief interventions for the population of women in need of some alcohol treatment. Representative studies with random assignment to treatment conditions and either substantial numbers of women, or a special focus on women, were included. Findings suggest brief interventions are not consistently helpful to women drinkers. PMID- 12127464 TI - A trial of "standard" outpatient alcoholism treatment vs. a minimal treatment control. AB - This study sought to examine the effectiveness of a "standard" outpatient alcoholism treatment (ST) program. An outpatient alcoholism treatment as it is commonly practiced in the US (with group and individual therapy, and an emphasis on Alcoholics Anonymous [AA]), was compared with a minimal treatment (MT) approach (weekly alcohol education movies). At 6 months, ST patients surpassed those in MT in terms of complete abstinence, reduction in amount of alcohol consumed, length of sobriety at follow-up, improvement in employment status, number of AA meetings attended, and lower initial drop-out. It is concluded that a ST approach is more helpful than MT in treating severely alcohol-dependent individuals who have not been able to cut down drinking on their own. Those already drinking less appeared to be helped by MT. PMID- 12127465 TI - Preliminary outcomes from the assertive continuing care experiment for adolescents discharged from residential treatment. AB - In many treatment systems, adolescents referred to residential treatment have the most serious alcohol or other substance use disorders and are at high risk of relapse. Upon discharge, these adolescents are typically referred to continuing care services, however, linkage to these services is often problematic. In this study, 114 adolescents (76% male) who stayed at least 7 days in residential treatment were randomly assigned to receive either usual continuing care (UCC) or UCC plus an assertive continuing care protocol (ACC) involving case management and the adolescent community reinforcement approach. ACC participants were significantly more likely to initiate and receive more continuing care services, to be abstinent from marijuana at 3 months postdischarge, and to reduce their 3 month postdischarge days of alcohol use. Preliminary findings demonstrate an ACC approach designed for adolescents can increase linkage and retention in continuing care and improve short-term substance use outcomes. PMID- 12127467 TI - Women in addictions treatment: comparing VA and community samples. AB - Despite increasing awareness of gender issues in substance use treatment, women with substance use disorders (SUD) and gender-specific treatment remain understudied. This study examines differences, including identification of comorbid issues and patients' perceived treatment needs, between women in different SUD treatment settings: an intensive VA outpatient program (VA; N = 76) and a private residential/outpatient program (Residence XII; N = 308). In both settings the Addiction Severity Index (ASI) was administered at intake; ASI data were collected from retrospective chart review. Results support previous findings that women entering SUD treatment endorse high rates of psychiatric and medical comorbidity, and past abuse. Women in VA SUD treatment experienced more impairment on indices of medical, psychiatric, and employment issues whereas the private agency sample had higher alcohol and family/social composite scores. The differences between and similarities among the two treatment groups have implications for design of women-specific SUD treatment programs. PMID- 12127468 TI - Magdalena Pilot Project: motivational outreach to substance abusing women street sex workers. AB - The Magdalena Pilot Project provided outreach to Albuquerque women sex workers who were also using illicit drugs, primarily cocaine and heroin. This initial uncontrolled trial evaluated the feasibility and potential impact of motivational interviewing (MI) on change in drug use and HIV risk behaviors. Twenty-seven women were enrolled and interviewed about their substance use, health risk behaviors, and plans for change, using the client-centered, directive method of MI. Four months later, 25 women (93%) were interviewed again to assess their drug use and health risk behaviors. Large reductions were reported in frequency (days) of drug use and sex work, with a corresponding increase in days of lawful employment. In identifying problems that most needed to be addressed in order to help them live healthier lives, the women prioritized (1) basic needs including decent housing, (2) mental health care, and (3) treatment for substance use disorders. PMID- 12127466 TI - Predicting treatment-outcome in cocaine dependence from admission urine drug screen and peripheral serotonergic measures. AB - We investigated whether urine drug screens (UDS) at admission and platelet paroxetine binding, a measure of serotonin transporter sites, were related to outcome measures for cocaine patients in treatment. Tritiated paroxetine binding sites on platelets were assayed and UDS were obtained for 105 African American cocaine-dependent outpatients. Outcome measures included number of negative urines, days in treatment, dropouts, and number of treatment sessions attended. A significant association was found between cocaine-positive UDS at admission and negative urines, treatment retention, dropouts, and treatment sessions; while Bmax values of paroxetine binding (density of serotonin transporter sites) were significantly associated with treatment retention and negative urines. Moreover, UDS and paroxetine binding combined to enhance prediction of retention and abstinence. Although both admission UDS and paroxetine binding seem to contribute individually in predicting outcome of cocaine patients, a combination of the two variables seems to have a stronger effect in terms of predicting treatment outcome. PMID- 12127469 TI - Staff beliefs about drug abuse clinical trials. AB - Staff from 10 community-based addiction treatment organizations in the National Drug Abuse Clinical Trials Network participated in an educational session about addiction research practices and human subject protections. This 1.5-hour presentation addressed "informed consent," "confidentiality of research information," "inclusion and exclusion criteria," "random assignment," "patient protections," and "patient payments." Pre- and postsession surveys were administered to 115 staff members measuring their beliefs about clinical trials. At baseline, 52% of staff believed patients could transfer out of a study even if they were doing poorly, and 55% believed staff had this right; 44% agreed that patients could participate in a clinical trial without understanding what would take place in the study. After the educational session, staff beliefs about patient protections were significantly increased in five of the seven items. A fourth of staff continued to believe patient payments were harmful, and 37% did not believe participation in a clinical trial would increase a patient's chances at recovery. PMID- 12127470 TI - Narcotics Anonymous participation and changes in substance use and social support. AB - In Victoria (a southern Australian state) in 1995, Narcotics Anonymous had a small but growing membership providing an opportunity to study the early experience of new self-help members. Ninety-one new members were interviewed and 62 (68%) were reinterviewed after 12 months. Three measures of self-help participation were examined: service role involvement, step work, and stable meeting attendance. Lower prior involvement in treatment services and greater participation in self-help predicted subsequent self-help participation. Higher levels of secondary school education predicted service role involvement and longer periods in stable meeting attendance. Higher self-help participation through the 12 months prior to follow-up was associated with lower levels of hazardous alcohol use and higher emotional support at reinterview. Multivariate regression analysis suggested stable self-help meeting attendance and step work continued to predict reductions in hazardous alcohol use and improvements in social support, after controlling for a range of alternative predictors. PMID- 12127471 TI - Research on psychiatric outcomes and interventions subsequent to disasters: a review of the literature. AB - Tragic events such as those of September 11, 2001, underscore the increasingly prominent role that psychiatrists play in aiding survivors, emergency workers, and broader communities to cope with disaster. The present review was undertaken to identify whether there exists a scientific basis for the practice of psychiatry in the aftermath of disasters. Most of the extensive literature over the past 30 years suggests that disasters have psychopathological consequences as well as medical and social ones. Pre-existing mood and anxiety disorders, although surprisingly not psychotic illness, appear to be risk factors for further psychopathology after a disaster. Thus, both acute psychopharmacological and psychotherapeutic interventions at disaster sites may prevent long-term sequelae, although their efficacy remains uncertain. Future controlled treatment trials are needed to determine the optimal treatment strategy. PMID- 12127472 TI - A pilot study of noradrenergic and HPA axis functioning in PTSD vs. panic disorder. AB - The biological literature in the anxiety disorders has focused on comparisons between patient groups and normal volunteers, with relatively little comparative study of the anxiety disorders. We therefore conducted this pilot study to compare a group of patients with post-traumatic stress disorder (PTSD) (n = 7) to a contiguously studied panic disorder group (n = 17) and healthy control subjects (n = 16) on baseline levels of cortisol and 3-methoxy-4-hydroxyphenylglycol (MHPG), and response to clonidine challenge. Despite the small sample size, highly significant differences were found on the following measures: PTSD patients had lower cortisol, lower MHPG, reduced MHPG volatility to clonidine challenge, and marginally reduced cortisol volatility compared to patients with panic disorder. These biological findings support existing clinical, epidemiologic, family study, and clinical trial findings that distinguish these two disorders as distinct syndromes. PMID- 12127473 TI - PTSD symptoms and cognitive performance in recent trauma survivors. AB - Chronic post-traumatic stress disorder (PTSD) has been associated with cognitive impairments involving memory and attention. The association between cognitive impairment and early PTSD symptoms is unknown, yet such association may lead to poorer processing of traumatic memories and thereby contribute to subsequent PTSD. This study evaluated the relationship between PTSD symptoms and cognitive functioning within 10 days of traumatic events. Forty-eight survivors were assessed for symptoms of PTSD, anxiety, depression and dissociation and for immediate and delayed verbal and figural memory, attention, learning and IQ. Survivors with high levels of PTSD symptoms showed impaired attention and immediate recall for figural information and lower IQ. They did not show, however, an impairment of verbal recall and learning. The observed difference was not explained by anxiety or dissociation. It disappeared, however, when the effect of depressive symptoms was controlled for. Lower IQ and impaired attention are associated with early PTSD and depressive symptoms. Poorer attention may have a role in shaping traumatic memories. PMID- 12127474 TI - Testosterone, sexuality and antisocial personality in rapists and child molesters: a pilot study. AB - Morning and afternoon levels of saliva testosterone levels in Finnish imprisoned rapists (n = 10) and child molesters (n = 10) were compared to those in randomly selected control subjects (n = 31). The associations of saliva testosterone with sexual behavior and antisocial personality traits were explored in all groups. The sexual offenders and control subjects did not differ in the between-subjects main effect estimated for the averaged morning and afternoon testosterone levels. Seven rapists and three child molesters met the criteria for antisocial personality disorder (ASP). In the sexual offenders, a summed ASP index was positively correlated with mean saliva testosterone. Sexual activity as estimated from self-reports of sexual intercourse and masturbation was significantly related to testosterone in both rapists and child molesters but not in the control males. The implications of these results are discussed. PMID- 12127475 TI - Decreased levels of brain-derived neurotrophic factor in serum of chronic schizophrenic patients. AB - Neurotrophic factors regulate neuronal development as well as synaptic plasticity, and their impairment is often implicated as a cause of schizophrenia. Among various neurotrophic molecules, brain-derived neurotrophic factor (BDNF) levels have been found to be increased in the corticolimbic regions of patients' brains. In the present study, we assessed peripheral BDNF levels in whole blood as well as in the serum of two independent groups of schizophrenic patients (n = 34 in each group) and healthy volunteers (n = 35 and n = 27, respectively). BDNF protein levels in fresh serum and blood of the patients and volunteers were measured using a two-site enzyme immunoassay and correlated with the number and decay of platelets. In addition to the studies of patients and volunteers, neuroleptic effects on BDNF levels were assessed by administering haloperidol to adult rats for 2 weeks or 5 months. The major findings were as follows: BDNF levels were significantly reduced in the serum of schizophrenic patients (P < 0.005, Mann-Whitney U-test) but not in their whole blood. Antipsychotic dose did not correlate with serum BDNF levels. Moreover, chronic administration of haloperidol failed to decrease serum BDNF levels in adult rats. Abnormal levels of BDNF are evident not only in the brain of schizophrenic patients, but also in their peripheral blood. The BDNF reduction in serum but not in whole blood suggests a potential deficit in neurotrophic factor release in patients with schizophrenia. PMID- 12127476 TI - Spatial working memory span, delayed response and executive function in schizophrenia. AB - This study investigated the spatial working memory span (SWMS) as well as the immediate memory span of schizophrenic patients, and examined the contribution of each span to the patients' executive function deficit in the visuospatial domain. SWMS measured the visuospatial working memory capacity that simultaneously processes and stores visuospatial information. Immediate memory span was measured with the spatial span (SS), a variant of Corsi's block-tapping test. A total of 16 patients diagnosed with schizophrenia and 16 normal control subjects participated in the study. SWMS, as well as the forward and backward SS, was significantly reduced in schizophrenia. The SWMS deficit observed in this study and previous findings of deficit in verbal working memory spans suggest that impairment in working memory capacity in schizophrenia is general, and not limited to the verbal domain. Executive function as assessed with the self ordered pointing task (SOPT) was also impaired in the patients, which is consistent with clinical observations of self-monitoring impairment in schizophrenia. SWMS was able to account for the performance on the SOPT, but its contribution in the patients' impairment did not reach statistical significance. Backward span deficit explained this executive function impairment. SWMS was effective in explaining schizophrenic patients' impaired performance on the spatial delayed response, a prefrontal function task. Implications of the relations observed between the spans and the prefrontal function tasks are discussed. PMID- 12127477 TI - Family attitude scale: measurement of criticism in the relatives of patients with schizophrenia in Japan. AB - Expressed emotion (EE) is traditionally measured with the Camberwell Family Interview (CFI), but the CFI requires considerable time for both execution and evaluation. As an alternative, we investigated the validity of the Family Attitude Scale (FAS), a questionnaire developed for the measurement of EE. The CFI, the FAS, the General Health Questionnaire (GHQ), and the Five-Minute Speech Sample (FMSS) were administered in 57 members of the families of 41 patients with acute episodes of schizophrenia. The relative sensitivity and specificity of EE assessment with the FAS compared with the criticism component of the CFI were 100% and 88.5%, respectively. EE assessment based on criticism as assessed with the FMSS compared with the CFI had a sensitivity of 40.0% and a specificity of 90.4%. The GHQ score tended to be higher in the high-scoring FAS group than in the low-scoring FAS group. The FAS showed excellent validity for the measurement of critical aspects of family attitudes, and the FAS score reflected the state of psychological health of the families. PMID- 12127478 TI - Race/ethnicity and depressive symptoms in community-dwelling young adults: a differential item functioning analysis. AB - To examine variations in the manifestation of depressive symptomatology across racial/ethnic groups, analyses of differential item functioning (DIF) on the Center for Epidemiologic Studies Depression Scale (CES-D) were separately conducted for representative samples of young adults in the following groups: African-Americans (n = 434), Hispanics born in the US (n = 493), and Hispanics born outside the US (n = 395). Non-Hispanic whites (n = 463) were employed as the reference group in all analyses. The effects of gender and age were controlled. DIF analyses indicated that: (1) about half of the CES-D items functioned differently among non-Hispanic whites compared to each of the other racial/ethnic groups; (2) the manifestation of symptoms seemed to be similar for both Hispanic groups, except for low positive affect; (3) African-Americans tended to favor somatic symptoms over affective (depressive) symptoms; (4) Immigrant Hispanics appeared to inhibit the expression of positive affect, and thus more high scorers on the total CES-D were observed within this subgroup. In contrast, no differences were observed when only negative items were considered. The use of positive affect items might artifactually induce spurious differences among people who were born outside the United States or North America. PMID- 12127479 TI - Cross-cultural validation of the Beck Depression Inventory-II in Japan. AB - The Beck Depression Inventory has undergone substantial revision recently as the BDI-II to correspond to DSM-IV criteria. We developed the Japanese version of the BDI-II and examined its psychometric properties. The linguistic equivalence was verified by a back-translation method. The final translation was administered to the visitors at a public health care center, and the responses of 766 adults (age = 24-82 years, women = 40%) were analyzed. Half of the participants completed the Center for Epidemiologic Studies Depression Scale (CES-D) as well. A high level of internal consistency reliability (Cronbach's alpha = 0.87) and item homogeneity was confirmed. Exploratory factor analysis showed a two-factor structure (cognitive and somatic-affective), which was almost identical to the original model demonstrated by Beck et al. (1996, Manual for the Beck Depression Inventor Psychological Corporation, San Antonio, TX, USA). The following confirmatory factor analysis also supported the two-factor structure. Adequate correlation (r = 0.69, P < 0.001) between the total score of the BDI-II and that of the CES-D was observed. A higher score for women compared to men, without significant age differences, was consistent with the results of previous reports. We conclude that the Japanese version of the BDI-II is psychometrically robust and can be used to assess depressive symptoms in Japanese people. PMID- 12127480 TI - Solubilization of zinc salts by a bacterium isolated from the air environment of a tannery. AB - Airborne bacteria isolated from a tannery air environment were screened for the property of solubilization of insoluble zinc oxide and zinc phosphate. Out of 10 strains tested, a strain of Pseudomonas aeruginosa (CMG 823) showed the best solubilization and solubilized both zinc oxide and zinc phosphate. Colonies of the bacterium produced clear haloes on solid medium which contained these insoluble metal compounds, but only when glucose was provided as a carbon source. Solubilization of zinc oxide and phosphate was accompanied by an increase in the H+ concentration of the medium, probably a consequence of the production of 2 ketogluconic acid. PMID- 12127481 TI - Plasmid profiling and antibiotic resistance of Vibrio strains isolated from cultured penaeid shrimp. AB - Resistance to different antibiotics was found in 26 of the 30 strains analyzed, more than 70% of the strains analyzed were resistant to carbenicillin and ampicillin and a significant correlation was found between the resistance to both antibiotics. Plasmids were found in 80% of the strains analyzed, and 11 different plasmid profiles were observed. The most common profile obtained had only a 21.2 kbp plasmid, a significant correlation was found between the presence of this plasmid and resistance to carbenicillin, although some exceptions could be detected. Plasmids were cured from a cephalothin resistant strain and reintroduced into the plasmid-free cell and into Escherichia coli DH5alpha, both strains gained resistance to this antibiotic. PMID- 12127482 TI - Lysobacter strain with high lysyl endopeptidase production. AB - A new lysyl endopeptidase producing strain, Lysobacter sp. IB-9374, was isolated from soil. This strain secreted the endopeptidase to culture medium at 6-12-fold higher levels relative to Achromobacter lyticus and Lysobacter enzymogenes. The mature Lysobacter sp. enzyme was enzymatically identical to Achromobacter lysyl endopeptidase bearing lysyl bond specificity, a high peptidase activity, a wide pH optimum, and stability against denaturants. Nucleotide sequence analysis of the Lysobacter sp. lysyl endopeptidase gene revealed that the enzyme is synthesized as a precursor protein consisting of signal peptide (20 amino acids (aa)), pro-peptide (185 aa), mature enzyme (268 aa), and C-terminal extension peptide (198 aa). The deduced amino acid sequence of the mature enzyme was totally identical to that of the Achromobacter enzyme. The Lysobacter sp. precursor protein has an 18-aa longer peptide chain following nine consecutive amino acid residues distinct from the Achromobacter counterpart at the C terminus. Total precursor protein is 671 aa of which only 268 aa are in the finally processed exoenzyme. PMID- 12127483 TI - Characterization of an inducible citrate uptake system in Penicillium simplicissimum. AB - When citrate was used as a sole source of carbon, citrate uptake by Penicillium simplicissimum increased 267-fold (if glucose-grown mycelium was adapted to citrate) or 1400-fold (if the fungus was grown on citrate) compared to glucose grown mycelium. Inhibition of macromolecular synthesis prevented this stimulation of citrate uptake. Citrate uptake by glucose-grown mycelium was low (0.0015 nmol min(-1) (mg DW)(-1)) and most probably due to diffusion of undissociated citric acid. Citrate-adapted mycelium had a K(M) of 65 micromol l(-1) and a V(max) of 0.34 nmol min(-1) (mg DW)(-1). In citrate-grown mycelium K(M) was 318 micromol l( 1) and V(max) was 8.5 nmol min(-1) (mg DW)(-1). Citrate uptake was inhibited by sodium azide and uncouplers (TCS, 3,3',4',5-tetrachlorosalicylanilide; FCCP, carbonyl cyanide p-trifluoromethoxyphenyl-hydrazone). Because of this we postulate that the induced citrate uptake must be an active transport process. The pH optimum of citrate uptake was between pH 6 and 7. EDTA and Mg2+, Mn2+, Cu2+, Zn2+, Fe2+, Ca2+ only weakly influenced the induced citrate uptake. The properties of citrate uptake by Aspergillus niger and P. simplicissimum are compared. PMID- 12127484 TI - Transcriptional and mutational analysis of the Helicobacter pylori urease promoter. AB - Urease is an essential virulence factor of the human gastric pathogen Helicobacter pylori, and is expressed to very high levels. The promoter of the urease operon contains sequences resembling the canonical -10 and extended -10 motifs, but no discernible -35 motif. To establish the role of different motifs and regions in the urease promoter, we fused the urease promoter to a genomic lacZ reporter gene in H. pylori, made substitutions in the aforementioned promoter motifs, and also made deletions in the upstream sequences removing regulatory sequences. Substitutions in the -10, extended -10 and predicted -35 motifs all significantly altered expression of the lacZ reporter gene, demonstrating their importance in transcription of the H. pylori urease operon. In contrast, sequential deletions upstream of the -35 region did not affect expression of the lacZ reporter gene. This demonstrates the modular structure of the H. pylori urease promoter, where basal levels of transcription are initiated from a typical sigma(70) promoter, which requires -10 and extended -10 motifs, and also its -35 motif for efficient transcription. Upstream sequences are not involved in basal levels of urease transcription, but play an important role in responses to environmental stimuli like nickel. PMID- 12127485 TI - Engineering a genetic transformation system for Colletotrichum acutatum, the causal fungus of lime anthracnose and postbloom fruit drop of citrus. AB - Postbloom fruit drop (PFD) of citrus is caused by Colletotrichum acutatum. PFD isolates infect flower petals, induce abscission of small fruit and can cause severe yield loss on most citrus cultivars. Isolates from Key lime anthracnose (KLA) cause that disease on the Mexican lime, but also cause PFD on sweet orange. Both PFD and KLA isolates exhibited resistance to the common selection agents including hygromycin, bialaphos, benomyl and geneticin/G418. A genetic transformation system was developed for C. acutatum to confer resistance to sulfonylurea (chlorimuron ethyl) by expressing an acetolactate synthase gene (sur) cassette from Magnaporthe grisea. The protocol was tested on 11 different KLA and PFD isolates. The transformation frequencies were highly variable among isolates and among experiments (0-17.9 per microg circular DNA using 10(7) protoplasts). Southern blot analysis of transformants indicated that the plasmid vector was randomly integrated in multiple copies into the genome of C. acutatum. Addition of restriction enzymes or use of a vector with homologous sequences did not change the transformation frequencies, but tended to reduce the number integrated. Over 97% of the transformants retained the sulfonylurea resistance phenotype under non-selective conditions. Of 300 transformants tested, three were unable to cause necrotic lesions on detached Key lime leaves. The transformation method opens up opportunities for the genetic manipulation of C. acutatum. PMID- 12127486 TI - A protein-farnesyl transferase inhibitor interferes with the serum-induced conversion of Candida albicans from a cellular yeast form to a filamentous form. AB - A commercially available, cell permeable, protein-farnesyl transferase inhibitor interfered with the serum-induced morphological change in Candida albicans from a cellular yeast form to a filamentous form. The inhibitor has a negligible effect on the growth of C. albicans cells in the cellular yeast form, at the levels used to interfere with the morphological change. Conversion of C. albicans from the yeast form to filamentous form is associated with pathogenicity and hence protein farnesyl transferase inhibitors are potentially of therapeutic value against C. albicans infection. PMID- 12127487 TI - Phage HK022-based integrative vectors for the insertion of genes in the chromosome of multiply marked Escherichia coli strains. AB - We constructed a series of plasmids that allow the insertion of cloned DNA in the Escherichia coli chromosome by site-specific integration into the bacteriophage HK022 bacterial attachment site. These plasmids make use of a ColE1 origin of replication, the phage HK022 attachment site attP, antibiotic resistance genes for selection and unique restriction sites. Circularisation of non-replicative fragments containing the HK022 attachment site attP is performed in vitro and site-specific integration of attP containing molecules is ensured by transfer into cells transiently expressing the HK022 integrase gene carried by a thermosensitive replicon. Insertion is very efficient and the inserted fragments are stably maintained without selection pressure. Since integrative fragments carry rarely used antibiotic markers conferring resistance to antibiotics hygromycin or apramycin, they can be used in most E. coli strains in conjunction with many replicative or integrative vectors. PMID- 12127488 TI - Functional characterization of 4'-phosphopantetheinyl transferase genes of bacterial and fungal origin by complementation of Saccharomyces cerevisiae lys5. AB - Lysine biosynthesis in yeast requires the posttranslational conversion of the alpha-aminoadipate semialdehyde reductase Lys2 by the 4'-phosphopantetheinyl transferase (PPTase) Lys5 from the inactive apo-form into the catalytically active holo-form. In this reaction, the peptidyl carrier domain of Lys2 is modified at a conserved serine residue side chain with the 4'-phosphopantetheine moiety derived from coenzyme A. We have deleted the lys5 gene in Saccharomyces cerevisiae to investigate the substrate specificity of various heterologous PPTase genes of bacterial and fungal origin by testing their ability to complement lys5 in trans. Genes encoding PPTases Sfp and Gsp from Bacillus spp., which are involved in non-ribosomal peptide antibiotic synthesis, complemented the lys5 deletion, whereas ydcB of Bacillus subtilis, which encodes the acyl carrier protein synthase involved in fatty acid synthesis, could not. Two yet uncharacterized fungal genes, q10474 of Schizosaccharomyces pombe, meanwhile annotated as the putative lys7 gene, and npgA of Aspergillus nidulans, also complemented the lys5 deletion and have thus been functionally characterized as PPTases. The complementation system described also provides the basis for a simple method of functional characterization of PPTase candidate genes and their cloning from chromosomal DNA or cDNA libraries of diverse origin. PMID- 12127489 TI - Enhanced production of D-(-)-3-hydroxybutyric acid by recombinant Escherichia coli. AB - Wild-type bacteria including Escherichia coli normally do not produce extracellular D-(-)-3-hydroxybutyric acid (3HB). To produce extracellular chiral 3HB, a new pathway for synthesis of 3HB was constructed by simultaneous expression of genes of beta-ketothiolase (phbA), acetoacetyl-CoA reductase (phbB), phosphor-transbutyrylase (ptb) and butyrate kinase (buk) in E. coli strain DH5alpha. E. coli DH5alpha containing any one of the four plasmids pBHR69, pUCAB, p68CM or pKKAB that harbor the phbA and phbB genes produced small amounts of 3HB, ranging from 75 to 400 mg l(-1), while E. coli DH5alpha harboring p68CMPTK containing genes of phbA, phbB, ptb and buk increased the 3HB concentration to 1.4 g l(-1) in shake flasks supplemented with LB broth and 20 g l(-1) glucose. 3HB production was further improved to over 2 g l(-1) in shake flasks when E. coli DH5alpha hosted two plasmids simultaneously that separately contained phbA and phbB in one plasmid while ptb and buk in the other. A batch fermentation run in a 5-l fermenter produced approximately 5 g l(-1) 3HB after 24 h. A fed-batch process increased 3HB production to 12 g l(-1) after 48 h of fermentation. PMID- 12127490 TI - Acid stress response in Helicobacter pylori. AB - To determine the existence of an acid stress response in Helicobacter pylori the global changes in the proteins synthesized by the bacterium when subjected to an acid stress were studied. H. pylori ATCC43504 previously adapted to pH 7 did not show an acid stress response as detected by the two-dimensional electrophoretic pattern of 35S-labeled proteins when incubated at pH 3. This was probably due to the neutralization of the external medium by the action of urease. However, H. pylori DW504UreI-negative, a mutant strain unable to transport urea into the cell, showed a large number of proteins changed, as is typical in an acid stress response. Some of these proteins were identified by N-terminal sequencing. PMID- 12127491 TI - Pseudomonas aeruginosa internalization by corneal epithelial cells involves MEK and ERK signal transduction proteins. AB - Invasion of epithelial cells represents a potential pathogenic mechanism for Pseudomonas aeruginosa. We explored the role of mitogen-activated protein kinase kinases (MEK 1/2) and the extracellular signal-regulated kinases (ERK 1/2) in P. aeruginosa invasion. Treatment of corneal epithelial cells with MEK inhibitors, PD98059 (20 microM) or UO126 (100 microM), reduced P. aeruginosa invasion by approximately 60% without affecting bacterial association with the cells (P=0.0001). UO124, a negative control for UO126, had no effect on bacterial internalization. Infection of cells with an internalization-defective flhA mutant of P. aeruginosa was associated with less ERK 1/2 tyrosine phosphorylation than infection with wild-type invasive P. aeruginosa. An ERK-2 inhibitor, 5 iodotubercidin (20 microM), reduced P. aeruginosa invasion by approximately 40% (P=0.035). Together, these data suggest that P. aeruginosa internalization by epithelial cells involves a pathway(s) that includes MEK and ERK signaling proteins. PMID- 12127492 TI - Salmonella typhimurium displays cyclical patterns of sensitivity to UV-C killing during prolonged incubation in the stationary phase of growth. AB - Stationary phase cells of Salmonella typhimurium were more resistant to killing by UV-C irradiation than those from the exponential phase. Analysis of the tolerance of cells taken at different stages of prolonged incubation as batch cultures to 60 or 100 J m(2) doses of UV-C revealed cycles of resistance and tolerance. The possible involvement of rpoS-controlled functions in mediating these cycles could be discounted because they were also detected in an rpoS minus mutant of S. typhimurium. The results are discussed in the context of heterogeneity in cells of stationary phase cultures of S. typhimurium. PMID- 12127493 TI - Characterization of the Streptomyces violaceoruber SANK95570 plasmids pSV1 and pSV2. AB - We have analyzed the structure of two extrachromosomal elements of the methylenomycin producing actinomycete Streptomyces violaceoruber SANK95570. The presence of the circular plasmid pSV1 which was supposed to contain the genes for methylenomycin biosynthesis could be verified. Physical mapping of pSV1 revealed a size of 175.35 kb for this plasmid. In addition we generated a restriction map for the 100-kb linear plasmid pSV2. Cloning and sequencing of the terminal ends of pSV2 indicated the presence of 426-bp terminal inverted repeats. Both pSV2 termini show significant homology to the chromosome ends of Streptomyces coelicolor A3(2) which is a closely related strain to S. violaceoruber SANK95570. PMID- 12127495 TI - Membrane-bound progesterone receptors coupled to G proteins in the fungus Rhizopus nigricans. AB - Steroid binding sites with high affinity for progesterone (Kd=40+/-14 nM determined by binding, and Kd=71+/-22 nM determined by displacement studies) and lower affinity for 21-hydroxyprogesterone and for testosterone, but no affinity for estradiol-17beta, onapristone and alpha-naphthoflavone were detected in the enriched plasma membrane fraction of the fungus Rhizopus nigricans. The amount of steroid binding sites is in accordance with the value of B(max)=744+/-151 fmol (mg protein)(-1). In the membrane fraction, progesterone induced about 30% activation of G proteins over basal level, as determined by GTPase activity (EC50=32+/-8 nM) and by the guanosine 5'-O-(3-thiotriphosphate) (GTPgammaS) binding rate (EC50=61+/-21 nM). The affinity of receptors for progesterone was substantially decreased in the presence of GTPgammaS and of cholera toxin. Our results suggest the existence of progesterone receptors in the membrane of Rhizopus nigricans and their coupling to G proteins. PMID- 12127494 TI - Extremely high frequency of common flagellar antigens between Bacillus thuringiensis and Bacillus cereus. AB - Bacillus cereus isolates, recovered from natural environments of Japan, were examined for their flagellar (H) antigenicities with the reference H antisera against Bacillus thuringiensis serotypes H1-H55. Of 236 B. cereus isolates tested, 165 (70%) were agglutinated with the reference antisera available. The frequencies of seropositive isolates were: 77% in soils, 68% on phylloplanes, and 60% in animal fecal populations. Among the 45 H serogroups detected, the serovar shandongiensis (H22) was the predominant, followed by the serovars entomocidus (H6), indiana (H16), pakistani (H13), and neoleonensis (H24ab). These five H serovars were commonly distributed in the three populations from different sources. PMID- 12127496 TI - BUT-1: a new member in the chromosomal inducible class C beta-lactamases family from a clinical isolate of Buttiauxella sp. AB - An atypical Enterobacteriaceae strain with a beta-lactam susceptibility pattern of inducible cephalosporinase was isolated in Tenon Hospital (Paris, France) from a patient's skull wound infection. Identifications by the API-50CHE biochemical system and 16S rRNA gene sequencing concluded that it was a member of the Buttiauxella genus. The bla gene was cloned and sequenced. The deduced translated product was a 383-amino acid protein (BUT-1) with 75-78% identity with the chromosomal AmpC beta-lactamases of Citrobacter freundii, Enterobacter aerogenes, Enterobacter cloacae and Escherichia coli. The isoelectric point of 9.0 and the kinetic constants of BUT-1 were comparable with results described for other Ambler class C enzymes. bla(BUT-1) and the associated ampR transcriptional regulator gene were divergently transcribed from a common intercistronic region, a genetic organization already described for other inducible class C beta lactamases. The deduced amino acid sequence of AmpR shared 85% and 81% identity with AmpR from E. cloacae and C. freundii respectively. PMID- 12127497 TI - Unexpected changes in photosystem I function in a cytochrome c6-deficient mutant of the cyanobacterium Synechocystis PCC 6803. AB - Cytochrome c6, the product of the petJ gene, is a photosynthetic electron carrier in cyanobacteria, which transfers electrons to photosystem I and which is synthesised under conditions of copper deficiency to functionally replace plastocyanin. The photosystem I photochemical activity (energy storage, photoinduced P700 redox changes) was examined in a petJ-null mutant of Synechocystis PCC 6803. Surprisingly, photosystem I activity in the petJ-null mutant grown in the absence of copper was not much affected. However, in a medium with a low inorganic carbon concentration and with NH4+ ion as nitrogen source, the mutant displayed growth inhibition. Analysis showed that, especially in the latter, the isiAB operon, encoding flavodoxin and CP43', an additional chlorophyll a antenna, was strongly expressed in the mutant. These proteins are involved in photosystem I function and organisation and are proposed to assist in prevention of overoxidation of photosystem I at its lumenal side and overreduction at its stromal side. PMID- 12127499 TI - Degradation of 2,7-dichlorodibenzo-p-dioxin by wood-rotting fungi, screened by dioxin degrading ability. AB - One hundred thirty-six strains of wood-rot fungi (74 strains, 66 species of 19 genera, and 62 unidentified strains) were screened for the dibenzo-p-dioxin (DD) decreasing activity. It was observed that 20% of additional DD (1 micromol) disappeared in the cultures of eight strains belonging to four genera (Aleurodiscus (one strain), Ceriporia (one strain), Phanerochaete (one strain), Phlebia (five strains)) and four unidentified strains. These 12 fungal strains were used for the examination of the degradation of [U-14C]2,7-dichlorodibenzo-p dioxin (2,7-diCDD). The fungi unidentified strain MZ-227, Phlebia sp. MG-60 and Phlebia lindtneri showed higher cumulative 14CO2 evolution rates than the other nine fungi. MZ-227, Phlebia sp. MG-60, and P. lindtneri converted 250 nmol of 2,7 diCDD to 196, 155 and 149 nmol of 14CO2, respectively, during a 30-day incubation period PMID- 12127498 TI - Nitric oxide releases intracellular zinc from prokaryotic metallothionein in Escherichia coli. AB - Nitric oxide (NO) has a broad spectrum of signalling and regulatory functions and multiple molecular targets. Recently, the intrabacterial toxicity of NO and mechanisms for NO resistance have been intensively investigated. Here we report for the first time that NO elicits release of zinc from a bacterial protein. Using the zinc-responsive expression of zntA (encoding a Zn-exporting P-type ATPase) fused to lacZ, i.e. Phi(zntA-lacZ), to monitor intracellular zinc, and SmtA (the Synechococcus metallothionein) as zinc store, we have shown that the NO donors NOC-5 and NOC-7 elicit zinc ejection. No increase in Phi(zntA-lacZ) activity was observed in a zntR mutant, indicating the specificity of the zntA promoter response to zinc ions. PMID- 12127500 TI - Phospholipid etherlipid and phospholipid fatty acid fingerprints in selected euryarchaeotal monocultures for taxonomic profiling. AB - Phospholipid etherlipid (PLEL) derived isoprenoids and phospholipid fatty acids (PLFA) were determined in eight Euryarchaeotal monocultures for taxonomic profiling. For the first time significant amounts of fatty acids in the PLFA of Euryarchaeota were determined. The PLFA proportion varied between 11.3 and 35.5% of the total phospholipid side chains except in Methanothermus fervidus where PLFA accounted for 89.0% of the total phospholipid side chains. Fractionation of fatty acids prior to gas chromatography mass spectrometry analysis revealed that non-ester-linked fatty acids dominated which accounted for 85.5-95.2% of total PLFA in all investigated archaeal strains. PLEL concentration and composition was estimated in accordance with previous studies with two exceptions. In the polar (phospho)lipid fraction of Methanopyrus kandleri side chains possibly derived from hydroxyarchaeol as well as acyclic and cyclic caldarchaeol were identified. In phospholipid extracts of Methanothermus fervidus the 'H-formed' caldarchaeol could not be detected. Overall, PLEL derived isoprenoids as well as PLFA enabled taxonomic differentiation of the selected microorganisms into phylogenetically related groups. PMID- 12127501 TI - Event templates in the lexical representations of verbs. AB - Four experiments support the hypothesis that syntactically relevant information about verbs is encoded in the lexicon in semantic event templates. A verb's event template represents the participants in an event described by the verb and the relations among the participants. The experiments show that lexical decision times are longer for verbs with more complex templates than verbs with less complex templates and that, for both transitive and intransitive sentences, sentences containing verbs with more complex templates take longer to process. In contrast, sentence processing times did not depend on the probabilities with which the verbs appear in transitive versus intransitive constructions in a large corpus of naturally produced sentences. PMID- 12127502 TI - The role of meaning in inflection: why the past tense does not require a rule. AB - How do we produce the past tenses of verbs? For the last 20 years this question has been the focal domain for conflicting theories of language, knowledge representation, and cognitive processing. On one side of the debate have been similarity-based or single-route approaches that propose that all past tenses are formed simply through phonological analogies to existing past tenses stored in memory. On the other side of the debate are rule-based or dual-route approaches which agree that phonological analogy is important for producing irregular past tenses (e.g., think-->thought), but argue that regular past tenses (e.g., walk- >walked) are generated via a +ed rule and that a principled account of regular inflection can only be given by recourse to explicit rules. This debate has become a crucial battleground for arguments concerning the necessity and importance of abstract mental rules, embracing not only language processing, but also the of nature cognition itself. However, in centering on the roles of phonological similarity and rules, the past tense debate has largely ignored the possible role of semantics in determining inflection. This paper presents five studies that demonstrate a striking and decisive role of semantic similarity in inflection. In fact, semantic factors appear to be more important in inflection than the grammatical considerations put forward by the dual-route account. Further, these new findings provide a new way of discriminating between the claims of single-route (similarity-based) and dual-route (rule-based) approaches. It appears that inflection is carried out through analogical reminding based on semantic and phonological similarity and that a rule-based route is not necessary to account for past tense inflection. PMID- 12127503 TI - Forgetting curves: implications for connectionist models. AB - Forgetting in long-term memory, as measured in a recall or a recognition test, is faster for items encoded more recently than for items encoded earlier. Data on forgetting curves fit a power function well. In contrast, many connectionist models predict either exponential decay or completely flat forgetting curves. This paper suggests a connectionist model to account for power-function forgetting curves by using bounded weights and by generating the learning rates from a monotonically decreasing function. The bounded weights introduce exponential forgetting in each weight and a power-function forgetting results when weights with different learning rates are averaged. It is argued that these assumptions are biologically reasonable. Therefore power-function forgetting curves are a property that may be expected from biological networks. The model has an analytic solution, which is a good approximation of a power function displaced one lag in time. This function fits better than any of the 105 suggested two-parameter forgetting-curve functions when tested on the most precise recognition memory data set collected by. Unlike the power-function normally used, the suggested function is defined at lag zero. Several functions for generating learning rates with a finite integral yield power-function forgetting curves; however, the type of function influences the rate of forgetting. It is shown that power-function forgetting curves cannot be accounted for by variability in performance between subjects because it requires a distribution of performance that is not found in empirical data. An extension of the model accounts for intersecting forgetting curves found in massed and spaced repetitions. The model can also be extended to account for a faster forgetting rate in item recognition (IR) compared to associative recognition in short but not long retention intervals. PMID- 12127505 TI - Biphenyl derivatives as novel dual NK(1)/NK(2)-receptor antagonists. AB - In a continuation of our efforts to simplify the structure of our neurokinin antagonists, a series of substituted biphenyl derivatives has been prepared. Several compounds exhibit potent affinities for both the NK(1) receptor (<10nM) and for the NK(2) receptor (<50 nM). Details on the design, synthesis, biological activities, SAR and conformational analysis of this new class of dual NK(1)/NK(2) receptor antagonists are presented. PMID- 12127506 TI - Apoptosis-inducing activity of synthetic intermediates of halichlorine. AB - Synthetic intermediates of alkaloid halichlorine with the azaspiro core structure have been found to induce apoptosis of cultured human cells including an acute monocytic leukemia cell line (THP-1) at micromolar concentrations. The novel biological activity of the intermediates was suggested to depend on the skeletal structure and silyloxymethyl functionality on the five-membered ring. PMID- 12127507 TI - A new class of potent nonpeptide luteinizing hormone-releasing hormone (LHRH) antagonists: design and synthesis of 2-phenylimidazo[1,2-a]pyrimidin-5-ones. AB - The design and synthesis of a new class of nonpeptide luteinizing hormone releasing hormone (LHRH) receptor antagonists, the 2-phenylimidazo[1,2 a]pyrimidin-5-ones, is reported. Among compounds described in this study, we identified the potent antagonist 15b with nanomolar in vitro functional antagonism. The result might suggest that the heterocyclic 5-6-ring system possessing a pendant phenyl group attached to the five-membered ring is the important structural feature for a scaffold of small molecule LHRH antagonists. PMID- 12127508 TI - Developing site-specific immobilization strategies of peptides in a microarray. AB - In peptide-based microarrays, most existing methods do not allow for site specific immobilization of peptides on the glass surface. We have developed two new approaches for site-specific immobilization of kinase substrates onto glass slides: (1) slides were functionalized with avidin for attachment of biotinylated peptides; and (2) slides were functionalized with thioester for attachment of N terminally cysteine-containing peptides via a native chemical ligation reaction. PMID- 12127509 TI - Antibody-based fluorescence detection of kinase activity on a peptide array. AB - Peptide-based microarrays allow for high-throughput identification of protein kinase substrates. However, current methods of detecting kinase activity require the use of radioisotopes. We have developed a novel fluorescence-based approach for quantitative detection of peptide phosphorylation on chip using fluorescently labeled anti-phosphoserine and anti-phosphotyrosine antibodies. This method is sensitive, specific and extremely fast, presenting obvious advantages and may find wider uses in high-throughput kinase screenings. PMID- 12127510 TI - Essential structural factors of acetogenins, potent inhibitors of mitochondrial complex I. AB - To elucidate the role of the hydrophobic alkyl tail of acetogenins in the inhibitory action, we synthesized an acetogenin derivative possessing the shortest tail (i.e., methyl group) and examined its inhibitory activity against bovine heart mitochondrial complex I. Our results indicated that the alkyl tail, which is one of the common structural features of natural acetogenins, is not an essential structural factor required for the potent inhibition. PMID- 12127511 TI - 4,4'-Benzophenone-O,O'-disulfamate: a potent inhibitor of steroid sulfatase. AB - We investigated whether the benzophenone moiety can be used as core element of steroid sulfatase (STS) inhibitors. While 4- and 3-benzophenone-O-sulfamates inhibit STS with IC(50) values between 5 and 7 microM irrespective of additional hydroxy and methoxy substituents at the second phenyl ring, benzophenone-O,O' disulfamates show increased activity. With an IC(50) value of 190 nM the 4,4' derivative is the first small monocyclic STS inhibitor coming close to the potency of the steroidal standard estrone sulfamate. PMID- 12127512 TI - Synthesis and biological evaluation of N-(7-indolyl)-3-pyridinesulfonamide derivatives as potent antitumor agents. AB - We herein report the synthesis and antitumor activity of E7070 analogues containing a 3-pyridinesulfonamide moiety. E7070 was selected from our sulfonamide-based compound collections, currently undergoing Phase II clinical trials because of its tolerable toxicity profile and some antitumor responses in the Phase I setting. Of the analogues examined, ER-35745, a 6-amino-3 pyridinesulfonamide derivative, demonstrated significant oral efficacy against the HCT116 human colon carcinoma xenograft in nude mice. PMID- 12127513 TI - Efficient asymmetric synthesis of (S)-2-ethylphenylpropanoic acid derivative, a selective agonist for human peroxisome proliferator-activated receptor alpha. AB - An optically active phenylpropanoic acid derivative, a selective agonist for human peroxisome proliferator-activated receptor alpha, was efficiently prepared in high optical purity by using Evans chiral oxazolidinone technique as a key step. PMID- 12127514 TI - Synthesis and hypoglycemic evaluation of substituted pyrazole-4-carboxylic acids. AB - The synthesis and in vivo activities of a series of substituted pyrazole-4 carboxylic acids as hypoglycemic agents are described. Modelization of some potent compounds, comparatively to the metformine, presents certain analogies permitting to predict the design of some novel antidiabetic drugs. PMID- 12127515 TI - SAR of 2,6-diamino-3,5-difluoropyridinyl substituted heterocycles as novel p38MAP kinase inhibitors. AB - 2,6-Diamino-3,5-difluoropyridinyl substituted pyridinylimidazoles, -pyrroles, oxazoles, -thiazoles and -triazoles have been identified as novel p38alpha inhibitors. Pyridinylimidazole 11 potently inhibited LPS-induced TNFalpha in mice, showed good efficacy in the established rat adjuvant (ED(50): 10 mg/kg po b.i.d.) and collagen induced arthritis (ED(50): 5 mg/kg po b.i.d.) with disease modifying properties based on histological analysis of the joints. PMID- 12127516 TI - Pyrrolylquinoxalinediones carrying a piperazine residue represent highly potent and selective ligands to the homomeric kainate receptor GluR5. AB - Pyrrolylquinoxalinediones carrying aminoalkyl residues were evaluated for affinity to the recombinant, homomeric kainate receptors GluR5, GluR6 and GluR7. Most derivatives preferred binding to GluR5. In particular, the piperazine 6e represents a highly potent and selective antagonist to GluR5. PMID- 12127517 TI - Identification of the ability of highly charged nanomolar inhibitors of protein kinases to cross plasma membranes and carry a protein into cells. AB - A fluorescently labeled adenosine-oligoarginine conjugate (ARC), nanomolar bisubstrate analogue-type inhibitor of basophilic protein kinases PKA and PKC, readily enters cells of different origin and localizes into cytoplasm and nucleus. Moreover, the biotinylated derivative of ARC is able to deliver avidin, a non-covalently attached protein cargo, into cells. PMID- 12127518 TI - The synthesis and biological evaluation of a novel series of antimicrobials of the oxazolidinone class. AB - A novel series of antimicrobials of the oxazolidinone class is disclosed. These compounds are characterized relative to previously described analogues by a 'halostilbene-derived' pharmacophore and demonstrate enhanced antimicrobial activity against key Gram-positive pathogens when compared to Linezolid. PMID- 12127520 TI - Rapid solid-phase synthesis of DNA-binding pyrrole-imidazole polyamides. AB - Pyrrole-imidazole polyamides can be synthesized to target predetermined sequences of DNA with nanomolar affinity and high specificity, and have been shown to modulate gene transcription both in vitro and in vivo. To make polyamides more readily available to biological laboratories, we have developed a rapid solid phase synthesis based on azabenzotriazole (OAt) activation that decreases synthesis time 60% compared to standard benzotriazole (OBt) techniques, without loss of yield or purity. PMID- 12127519 TI - Identification of a novel 1'-[5-((3,5-dichlorobenzoyl)methylamino)-3-(3,4 dichlorophenyl)-4-(methoxyimino)pentyl]-2-oxo-(1,4'-bipiperidine) as a dual NK(1)/NK(2) antagonist. AB - A novel series of dual NK(1)/NK(2) receptor antagonists, based on the 2-oxo-(1,4' bipiperidine) template, has been prepared. Compound 10R is a potent dual NK(1)/NK(2) antagonist and demonstrates excellent in vivo activity and good oral plasma levels in the dog. PMID- 12127521 TI - 3-Aryl pyrazolo[4,3-d]pyrimidine derivatives: Nonpeptide CRF-1 antagonists. AB - The synthesis of a series of 3-aryl pyrazolo[4,3-d]pyrimidines as potential corticotropin-releasing factor (CRF-1) antagonists is described. The effects of substitution on the aromatic ring, the amino group and the pyrazolo ring on CRF-1 receptor binding were investigated. PMID- 12127522 TI - Rapid synthesis of triazine inhibitors of inosine monophosphate dehydrogenase. AB - A series of novel triazine-based small molecule inhibitors (IV) of inosine monophosphate dehydrogenase was prepared. The synthesis and the structure activity relationships (SAR) derived from in vitro studies are described. PMID- 12127523 TI - N-(arylacetyl)-biphenylalanines as potent VLA-4 antagonists. AB - A series of potent N-(aralkyl-, arylcycloalkyl-, and heteroaryl-acyl)-4 biphenylalanine VLA-4 antagonists was prepared by rapid analogue methods using solid-phase chemistry. Further optimization led to several highly potent compounds (IC(50) <1 nM). Evaluation of rat pharmacokinetic revealed generally high clearance. PMID- 12127524 TI - Novel thiophene derivatives for the treatment of benign prostatic hyperplasia. AB - The syntheses and biological activities of a novel series of 2,4- and 2,5 disubstituted thiophenes are reported. These analogues have shown excellent affinity and selectivity against alpha(1)-adrenoreceptor subtypes. PMID- 12127525 TI - Improving metabolic stability of phosphodiesterase-4 inhibitors containing a substituted catechol: prevention of reactive intermediate formation and covalent binding. AB - A detailed study directed towards metabolic stability optimization of the alkoxy substituents on the catechol moiety of CDP-840 is reported. Replacement of the methoxy and cyclopentyloxy substituents by cyclobutyloxy and/or difluromethoxy groups resulted in the discovery of potent and selective PDE4 inhibitors where the formation of reactive metabolites that could covalently bind to microsomal protein was significantly reduced or eliminated. PMID- 12127526 TI - Rational design of 4,5-disubstituted-5,7-dihydro-pyrrolo[2,3-d]pyrimidin-6-ones as a novel class of inhibitors of epidermal growth factor receptor (EGF-R) and Her2(p185(erbB)) tyrosine kinases. AB - A novel class of 4,5-disubstituted-5,7-dihydro-pyrrolo[2,3-d]pyrimidin-6-ones has been discovered as potent and selective inhibitors of the EGF-R tyrosine kinase family. These compounds selectively inhibit EGF-R kinase activity at low nanomolar concentration and tyrosine autophosphorylation in cells expressing EGF R or Her2 (p185(erbB)). Structure-activity relationships (SARs) for this class of compounds are presented. PMID- 12127527 TI - Antifungal activity of bifunctional sphingolipids. Intramolecular synergism within long-chain alpha,omega-bis-aminoalcohols. AB - The in vitro antifungal activity of a series of alpha,omega-bifunctionalized aminoalcohols against Candida glabrata was measured. The dimeric bi functionalized lipids exhibited activity about approximately 10-fold higher higher than D-sphingosine, which is a larger factor than expected from the simple additive effects of vicinal aminoalcohols groups. PMID- 12127528 TI - Synthesis and biological evaluation of 9-substituted tetracycline derivatives. AB - The synthesis of 9-substituted tetracycline derivatives has been accomplished by the reaction of C9 diazonium tetrafluoroborate tetracycline salts with organotin reagents under modified Stille coupling conditions. Several of these unreported derivatives show promising in vitro biological activity against tetracycline resistant and antibiotic resistant bacteria. PMID- 12127529 TI - Antimalarial compounds from Parinari capensis. AB - The antimalarial activity of the raw petroleum ether and dichloromethane extracts of the stems of Parinari capensis (Chrysobalanceae) was determined. Phytochemical investigation of these extracts led to the isolation of three diterpene lactones that possess antimalarial activity with IC(50) values of 0.54, 0.67, and 1.57 microg/mL. Although their antimalarial activity is promising, the toxicity profiles of these diterpene lactones prevent further biological evaluation. They could however be used effectively as lead compounds in the synthesis of novel antimalarial agents. PMID- 12127530 TI - Design, synthesis, and biological evaluation of novel, centrally-acting thyrotropin-releasing hormone analogues. AB - Novel, metabolically stable and centrally acting TRH analogues with substituted pyridinium moieties replacing the [His(2)] residue of the endogenous peptide were prepared by solid-phase Zincke reaction. The 1,4-dihydropyridine prodrugs of these analogues obtained after reducing the pyridinium moiety were able to reach the brain and maintain a sustained concentration of the charged, degradation resistant analogues formed after enzymatic oxidation of the prodrug, as manifested by the analeptic action measured in mice. Among the four analogues reported, compound 2a showed the highest potency and longest duration of action in reducing the pentobarbital-induced sleeping time compared to the parent TRH. No binding to the endocrine TRH-receptor was measured for 2a; thus, this compound emerged as a potent, centrally acting TRH analogue. PMID- 12127531 TI - Diboronic acids as fluorescent probes for cells expressing sialyl Lewis X. AB - A series of fluorescent diboronic acids was synthesized in nine steps as potential sensors for sialyl Lewis X (sLex). The fluorescent binding studies of these sensors with sLex were carried out in a mixed aqueous solution. Compound 7e was found to show the strongest fluorescence enhancement upon binding with sLex. Using cell cultures, 7e was shown to label sLex-expressing HEPG2 cells at 1 microM, while non-sLex-expressing cells were not labeled. PMID- 12127532 TI - Synthesis and initial structure-activity relationships of a novel series of imidazolo[1,2-a]pyrimid-5-ones as potent GnRH receptor antagonists. AB - SAR studies of 2-arylimidazolo[1,2-a]pyrimid-5-ones 10a-m, which were derived from initial lead 3a, resulted in the discovery of a series of potent nonpeptide human GnRH receptor antagonists. Compounds with good potency (e.g., 10e, K(i)=7.5 nM) were prepared by introduction of a 2-(2-pyridyl)ethyl at the basic nitrogen and a 3-pentyl ester at the 6-position of the bicyclic core. PMID- 12127533 TI - Design, synthesis and structure-activity relationships of novel imidazolo[1,2 a]pyrimid-5-ones as potent GnRH receptor antagonists. AB - SAR studies of lead GnRH receptor antagonists 2a and 2b reported earlier resulted in the discovery of compound 10b which showed much higher potency (K(i)=4.6 nM, compared with 2b, K(i)=230 nM) in which the 7-position of the imidazolo[1,2 a]pyrimidone core was substituted with a methyl group, and the ester at the 6 position was replaced by the 3-methoxyphenyl group. PMID- 12127534 TI - Phosphonate and phosphinate analogues of N-acylated gamma-glutamylglutamate. potent inhibitors of glutamate carboxypeptidase II. AB - Phosphonate and phosphinate analogues of N-acylated gamma-glutamylglutamate were tested for the ability to inhibit glutamate carboxypeptidase II (GCP II). All of the compounds inhibit GCP II with IC(50) values in the low nanomolar range. The comparison of the results to previously reported inhibitory studies of the same compounds toward folylpoly-gamma-glutamyl synthetase (FPGS) and gamma-glutamyl hydrolase (gamma-GH) provides insight into structural and mechanistic features of each enzyme. Potential utility of these compounds as diagnostic agents and probes to understand folate or antifolate poly-gamma-glutamates metabolism is also described. PMID- 12127535 TI - Lipid A structures containing novel lipid moieties: synthesis and adjuvant properties. AB - Structurally well-defined immune stimulatory molecules are important components of new generation molecular vaccines. In this paper, the design and synthesis of two lipid A analogues containing an unnatural tri-lipid acyl group are described. In a totally synthetic liposomal vaccine system, these re-designed lipid A analogues demonstrate potent immune stimulatory properties including antigen specific T-cell activation. PMID- 12127536 TI - Discovery of potent, selective human granzyme B inhibitors that inhibit CTL mediated apoptosis. AB - A novel class of small molecule human granzyme B inhibitors is reported. Compound 20 has a K(i) of 7 nM against human granzyme B and blocks CTL mediated apoptosis with an IC(50) of 3 micromolar. PMID- 12127537 TI - Design and synthesis of novel inhibitors of gelatinase B. AB - A new method was developed to identify nonpeptidic metalloproteinase inhibitors with novel zinc binding groups. Application of this method to matrix metalloproteinase-9 resulted in the identification of aminomethyl benzimidazole analogue 7a with an IC(50)=13 microM. PMID- 12127538 TI - N-aryl-prolyl-dipeptides as potent antagonists of VLA-4. AB - The design, synthesis, and biological evaluation of N-arylprolyl-dipeptide derivatives as small molecule VLA-4 antagonists is described. Potency against VLA 4 and alpha(4)beta(7) and rat pharmacokinetic evaluation revealed some advantages over the related N-(arylsulfonyl)-prolyl-dipeptide analogues. PMID- 12127539 TI - Synthesis and biological evaluation of 3'-carboranyl thymidine analogues. AB - Boron neutron capture therapy (BNCT) is a chemoradio-therapeutic method for the treatment of cancer. It depends on the selective targeting of tumor cells by boron-containing compounds. One category of BNCT agents with potential to selectively target tumor cells may be thymidine derivatives substituted at the 3' position with appropriate boron moieties. Thus, several thymidine analogues were synthesized with a carborane cluster bound to the 3'-position either through an ether or a carbon linkage. The latter are the first reported carborane-containing nucleosides in which the carboranyl entity is directly linked to the carbohydrate portion of the nucleoside by a carbon-carbon bond. Low but significant phosphorylation rates in the range of 0.18% that of thymidine were observed for the carbon-linked 3'-carboranyl thymidine analogues in phosphoryl transfer assays using recombinant preparations of thymidine kinases 1 (TK1) and thymidine kinases 2 (TK2). Some of the ether-linked 3'-carboranyl thymidine analogues appeared to be slightly unstable under acidic as well as phosphoryl transfer assay conditions and were, if at all, poor substrates for TK1. PMID- 12127540 TI - Syntheses of dendritic linkers containing chlorambucil residues for the preparation of antibody-multidrug immunoconjugates. AB - A novel dendritic molecule with nine chlorambucil (CBL) residues on the surface and a maleimide moiety at the core terminus was synthesized using a convergent synthetic methodology. This molecule is ready for attachment to single-chain Fv antibodies (scFvs) to form antibody-multidrug immunoconjugates in an effort to study the relevance of drug/antibody molar ratio and the potency of these drug antibody immunoconjugates. A monomer and a trimer with a similar structural motif were also prepared for comparative purposes. PMID- 12127541 TI - Isothiazole dioxides: synthesis and inhibition of Trypanosoma brucei protein farnesyltransferase. AB - A series of isothiazole dioxides was synthesized and evaluated as inhibitors of protein farnesyltransferase from the parasite that causes African sleeping sickness (Trypanosoma brucei). The most potent compound in the series inhibited the parasite enzyme with an IC(50) of 2 microM and blocked the growth of the bloodstream parasite in vitro with an ED(50) of 10 microM. The same compound inhibited rat protein farnesyltransferase and protein geranylgeranyltransferase type I only at much higher concentration. PMID- 12127542 TI - Synthesis and antiparasitic activity of 1H-benzimidazole derivatives. AB - Compounds 1-18 have been synthesized and tested in vitro against the protozoa Giardia lamblia, Entamoeba histolytica and the helminth Trichinella spiralis. Inhibition of rat brain tubulin polymerization was also measured and compared for each compound. Results indicate that most of the compounds tested were more active as antiprotozoal agents than Metronidazole and Albendazole. None of the compounds was as active as Albendazole against T. spiralis. Although only compounds 3, 9 and 15 (2-methoxycarbonylamino derivatives) inhibited tubulin polymerization, these were not the most potent antiparasitic compounds. PMID- 12127543 TI - Identification of a novel partial inhibitor of dopamine transporter among 4 substituted 2-phenylquinazolines. AB - In an attempt to identify novel ligands for the dopamine transporter, a series of 4-substituted-2-phenylquinazolines were synthesized and evaluated. Among the compounds studied, 4-[(diphenylmethyl)amino]-2-phenylquinazoline (4 g) was identified as a novel partial inhibitor of [(125)I]RTI-55 binding to the dopamine transporter and a partial inhibitor of [(3)H]dopamine uptake. PMID- 12127544 TI - N-thiolated beta-lactams: novel antibacterial agents for methicillin-resistant Staphylococcus aureus. AB - In this report we describe a new family of N-thiolated beta-lactams that have antibacterial activity against methicillin-resistant Staphylococcus aureus (MRSA). The compounds show unprecedented structure-activity features and an unusual mode of action for a beta-lactam antibiotic. PMID- 12127545 TI - Binding modes of two novel dinucleotide inhibitors of HIV-1 integrase. AB - Insights into the binding modes on HIV-1 integrase of our novel dinucleotide inhibitors (pisodApdC and pdCpisodU) have been obtained using molecular docking experiments. In contrast to their base-stacked unbound state, these dinucleotides in their integrase-bound state prefer unstacked conformations for a more extensive interaction with the active site. The calculated free energies of binding are in concert with the experimentally acquired anti-HIV-1 integrase data. PMID- 12127546 TI - Convenient synthesis of human calcitonin and its methionine sulfoxide derivative. AB - The human calcitonin peptide chain was assembled using Fmoc solid-phase peptide synthesis chemistry. The combinations of cleavage Reagent H with trans [Pt(en)(2)Cl(2)](2+) and Reagents B, K, and R with trans-[Pt(CN)(4)Cl(2)](2-) provide convenient methods for the synthesis of human calcitonin and its methionine sulfoxide derivative; the formation of intramolecular disulfide bonds by the above Pt(IV) oxidants is essentially quantitative. PMID- 12127547 TI - Decoding region bubble size and aminoglycoside antibiotic binding. AB - Aminoglycoside antibiotics promiscuously bind to structurally diverse RNA molecules containing internal bubbles and bulges with affinities in the microM range. An interesting exception is found in the human 12S mitochondrial decoding region where aminoglycoside binding, unlike in the case of its bacterial and human cytoplasmic counterparts, is absent. Mutations that reduce the size of the bubble in the 12S decoding region immediately restore aminoglycoside binding, giving the system chemical switch like behavior. PMID- 12127548 TI - Sequence selective recognition of DNA by hairpin conjugates of a racemic seco cyclopropaneindoline-2-benzofurancarboxamide and polyamides. AB - Conjugates of racemic seco-cyclopropaneindoline-2-benzofurancarboxamide (CI-Bf) and four diamides (ImIm 1, ImPy 2, PyIm 3, and PyPy 4, where Py is pyrrole, and Im is imidazole), linked by a gamma-aminobutyrate group were synthesized. In addition to alkylating at adenine-N3 positions within an A(5) sequence, the imidazole-containing compounds 1 and 2 were found to also alkylate purine-N3 positions within a sequence 3'-GGGGGGA(888)CTGCTC(894)-5'. A model for the binding of hairpin conjugates 1 and 2 with the 3'-GACT-5' sequence is proposed. PMID- 12127549 TI - What are we learning from familial chronic lymphocytic leukemia? PMID- 12127550 TI - Malignant hematopoietic cell lines: in vitro models for the study of mantle cell lymphoma. AB - Mantle cell lymphoma (MCL) is a distinct type of B cell malignancy and accounts for approximately 5-10% of non-Hodgkin's lymphomas (NHL). The characteristic cytogenetic aberration in MCL is the translocation (11;14)(q13;q32) present in virtually all cases. This rearrangement at the BCL1 locus at 11q13 dysregulates the gene CCND1 following juxtaposition with immunoglobulin heavy chain (IGH) transcriptional enhancers at 14q32 and leading to overexpression of its protein product, cyclin D1, which plays a key role in the control of the cell cycle. Eight continuous cell lines (plus several sister cell lines) have been hitherto established from lymph nodes or peripheral blood of patients with MCL (n=5) or with a lymphoma which would nowadays be classified as MCL (n=3). Six of these cell lines carry the specific t(11;14) translocation and a seventh cell line while being negative for t(11;14) shows a rearranged BCL1 locus and cyclin D1 overexpression. Each of these MCL cell lines is unique with regard to its immunophenotypical, additional cytogenetic and functional features. In light of the relatively low frequency of this lymphoma and the poor results of current treatment strategies, the availability of various types of MCL-derived cell lines for immunologic, cytogenetic, molecular and functional studies is expected to illuminate the biology of this disease, which in turn will be hopefully translated into new and better therapies. PMID- 12127551 TI - Idiopathic thrombocytopenic purpura and myelodysplastic syndrome: distinct entities or overlapping syndromes? PMID- 12127552 TI - Telomere length and heavy-chain mutation status in familial chronic lymphocytic leukemia. AB - We examined whether telomere lengths of peripheral blood mononuclear cells are associated with immunoglobulin gene usage in 21 familial chronic lymphocytic leukemia (CLL) patients. Subjects with unmutated V genes tended to have shorter telomeres than those with somatic mutations, especially after adjusting for age. Unlike V(H) mutation status, telomere length was not predictive for survival. Our results suggest that telomere length is associated with V(H) gene mutation status and provides further evidence that the biological basis of familial B-CLL is similar to that of sporadic patients. PMID- 12127553 TI - Regulation of CD23 isoforms on B-chronic lymphocytic leukemia. AB - CD23 is constitutively and atypically expressed on malignant B-cells in patients with chronic lymphocytic leukemia. It exists in two isoforms that differ only in a short amino acid sequence at the N-terminus. The CD23a isoform exhibits an endocytosis signal, that renders it more efficient in antigen uptake than CD23b. Therefore, we analyzed the regulation of CD23 isoforms and tested the ability to stimulate T-cell clones by targeting antigen to CD23 on CLL B-cells. Investigation of several stimulators to promote CD23a expression on CLL versus normal B-cells confirmed a different CD23 regulation in B-CLL. We did not find any evidence for a differential regulation of the two CD23 isoforms in B-CLL. However, CD23a is always predominantly expressed with a constant ratio of CD23a:CD23b. We show that antigen targeted to CD23 on CLL B-cells is very efficiently presented. Therefore, CD23 is likely to provide a suitable target for receptor-mediated antigen presentation in B-CLL which can be used to activate a T cell response. PMID- 12127554 TI - Blastic CD4 NK cell leukemia/lymphoma: a distinct clinical entity. AB - We report the findings of three new cases of a distinct clinicopathologic natural killer (NK) cell malignancy characterized by cutaneous, nodal and bone marrow infiltration by CD3-CD4+CD56+ NK blastic cells. Tumor cells were detected in bone marrow and in peripheral blood smears and showed finely distributed nuclear chromatin with nucleoli and a moderate amount of cytoplasm. Epstein-Barr virus (EBV) DNA was negative in the two tested cases. The immunophenotypes determined by flow cytometry were identical concerning mCD3-cytCD3-CD4+weakCD56+ HLA-DR+. The TCR was in germline configuration in the two cases tested. NK cell activity was demonstrated only in one out of the two cases tested. The negative reactions with alpha-naphthyl-acetate-esterase (ANAE), CD11b and CD14 strongly suggested that the tumor cells were not of the monocytic lineage. PMID- 12127555 TI - Quantitative analysis of Cyclin D1 and CD23 expression in mantle cell lymphoma and B-chronic lymphocytic leukemia. AB - We studied Cyclin D1 (CyD1) and CD23 mRNA expression with real-time quantitative reverse transcription polymerase chain reaction (RQ-PCR) method. CyD1 expression in peripheral blood of seven mantle cell lymphoma (MCL) patients was found to be 1305.4 times higher than in 24 B-chronic lymphocytic leukemia (CLL) patients. CD23 expression in CLL was found to be 54.8 times higher than in MCL. These differences were statistically significant, and no overlap was found in CyD1 expression intensities between MCL and CLL. RQ-PCR allows rapid, simple and accurate quantification of CyD1 and CD23 expression, even from small samples, and is thus useful for the diagnosis of MCL and CLL. PMID- 12127556 TI - Loss of heterozygosity of p16 correlates with minimal residual disease at the end of the induction therapy in non-high risk childhood B-cell precursor acute lymphoblastic leukemia. AB - We evaluated the incidence of MTS1/p16 deletions by loss of heterozygosity (LOH) analysis in 36 non-high risk B-cell precursor childhood acute lymphoblastic leukemia (BCP-ALL) and correlated these results with clinical features and with the presence of minimal residual disease (MRD) at the end of induction therapy. LOH was analyzed using three microsatellite markers flanking the p16 gene. MRD was studied by the polymerase chain reaction (PCR) for IgH and TCRdelta genes. All patients were classified and treated according to the BFM-86 protocol. A slower response to the induction treatment (MRD) was associated with LOH of p16 and worse clinical outcome. Thus, LOH of p16 may be a marker of chemotherapy resistance among the children classified as non-high risk BCP-ALL. PMID- 12127557 TI - Two groups of chronic myelomonocytic leukaemia: myelodysplastic and myeloproliferative. Prognostic implications in a series of a single center. AB - The clinical records of 70 patients seen at our hospital between 1976 and 1998 and diagnosed as suffering from chronic myelomonocytic leukaemia (CMML) were reviewed in order to confirm the validity of the classification into two forms of disease that the French-American-British Co-operative Leukaemia Group (FAB) proposed in 1994: myelodysplastic (MD) and myeloproliferative (MP), depending on the peripheral white blood cell count (WBC) (less or more than 13 x 10(9)/l, respectively). After the rejection of incomplete records and lost to follow up patients, our study population consisted of 49 records. Our results confirm that, even though this classification is useful in order to separate two classes of patients, it is not enough to predict the prognosis in an accurate manner. A lot of studies have tried to find some prognostic factors, but the results have been discordant. The multivariate analysis of our group of patients showed three prognostic factors: serum lactate dehydrogenase (LDH) >1.5 times normal level, blasts in bone marrow >5%, and peripheral blood leukocytes >10 x 10(9)/l. A second multivariate analysis led us to distinguish two groups: high risk (2-3 risk factors) and low risk (0-1 risk factors) (median survival 7 and 44 months, respectively) with a very high statistic significance (P<0.0001). This score should be applied to other series of CMML patients in order to confirm its validity. PMID- 12127558 TI - Monoclonal constitution of neutrophils detected by PCR-based human androgen receptor gene assay in a subset of idiopathic thrombocytopenic purpura patients. AB - It is well known that some patients with monopathic thrombocytopenia in myelodysplastic syndrome (MDS) show clinico-hematologic features resembling chronic idiopathic thrombocytopenic purpura (ITP). This study examined the monoclonal nature of ITP to obtain a further insight into patients with borderline ITP and monopathic thrombocytopenia in MDS, using polymorphic trinucleotide CAG repeats in the X-linked human androgen receptor (HUMARA) gene. In this study, we separated peripheral neutrophils and mononuclear cells (MNCs) from 18 patients with chronic ITP, and analyzed them in comparison with those from normal or MDS female subjects by PCR-based HUMARA assay. All normal controls showed a polyclonal pattern of the HUMARA gene, whereas some MDS patients had monoclonality in MNC and/or neutrophils. Among ITP patients, two had a nonrandom inactivation pattern of the HUMARA gene in neutrophils, which was considered to be derived from hematopoietic cells of clonal origin, whereas no ITP patient had MNC of clonal nature. Two ITP patients with a monoclonal pattern in the neutrophil fraction were refractory to ordinary treatment. This approach may provide further information in patients with borderline hematologic disorders between chronic ITP and refractory thrombocytopenia of MDS. PMID- 12127559 TI - Expression of insulin-like growth factors IGF-I and IGF-II, and their receptors during the growth and megakaryocytic differentiation of K562 cells. AB - Insulin-like growth factors (IGFs) I and II are critical regulators of cell proliferation and differentiation and most of the growth promoting properties of both ligands are mediated by IGF-I receptor (IGF-IR). In the present study we have investigated the role of IGFs in K562 cell line during normal growth and 12 O-tetradecanoyl-phorbol-13-acetate (TPA)-induced megakaryocytic differentiation. Abundant expression of IGF-I, IGF-II and IGF-IR was demonstrated in resting cells and exogenous IGF-I and IGF-II increased 3H-thymidine incorporation in a dose dependent manner. In contrast, we found that basal growth of the cells was inhibited by using anti-IGF-IR mAb. Furthermore, also IGF-I and IGF-II induced DNA synthesis was significantly suppressed by anti-IGF-IR mAb. During megakaryocytic differentiation, expression of IGF-IR increased during first 12h, but after that the expression started to decrease together with IGF-I. Taken together, our data suggest that autocrine production of IGF-I and IGF-II may via IGF-IR play a significant role in the growth and megakaryocytic differentiation of K562 cells. PMID- 12127560 TI - Bone and bone marrow interactions: hematological activity of osteoblastic growth peptide (OGP)-derived carboxy-terminal pentapeptide. II. Action on human hematopoietic stem cells. AB - Osteogenic growth peptide (OGP) is a peptide exerting regulatory effects on the bone and on bone marrow. The carboxy-terminal pentapeptide (OGP10-14) is the biologically active portion of OGP. We evaluated OGP10-14 hematopoietic activity performing colony-forming tests on human stem cells derived by bone marrow, peripheral blood and cord blood. Granulocyte-macrophage colony-forming unit (CFU) were significantly increased in OGP10-14-treated samples, while granulocyte erythrocyte-monocyte-megakaryocyte CFU and burst-forming unit (BFU) erythroid were increased only in the cord blood test.Moreover, OGP10-14 preserves stem cells self renewal potential in long-term culture (LTC) initiating cells and acts directly on CD34+ enriched cells or by increasing activity of stem cell factor (SCF) and granulocyte-megakaryocyte colony-stimulating factor. PMID- 12127561 TI - Establishment and characterization of a new mantle cell lymphoma cell line, Mino. AB - Mantle cell lymphoma (MCL) is a distinct type of B-cell non-Hodgkin's lymphoma characterized by cyclin D1 overexpression and the cytogenetic abnormality, the t(11;14)(q13;q32). MCL cell lines have been difficult to establish and in vitro studies of these neoplasms are scarce. We describe the establishment and characteristics of a new MCL cell line, Mino. The cells are large, growing singly and in small clumps in vitro. By flow cytometry, the immunophenotype was compatible with MCL (i.e. CD5+CD20+CD23-FMC7+). Conventional cytogenetics showed hyperdiploidy with multiple complex karyotypic abnormalities, but no evidence of the t(11;14), proven to be present only by fluorescence in situ hybridization and polymerase chain reaction (PCR) methods. Western blots showed expression of cyclin D1 but no detectable cyclin D2 and cyclin D3; the retinoblastoma protein was predominantly phosphorylated. There was expression of tumor suppressor gene products including p53, p16(INK4a), and p21(WAF1). Sequencing of the TP53 gene revealed a mutation (codon 147(valine-->glycine)) in exon 5. Epstein Barr virus was absent. In summary, Mino is a new MCL cell line that may be useful to study the pathogenesis of MCL. PMID- 12127562 TI - NAIP-deltaEx10-11: a novel splice variant of the apoptosis inhibitor NAIP differently expressed in drug-sensitive and multidrug-resistant HL60 leukemia cells. AB - Alterations of neuronal apoptosis inhibitory protein (NAIP), a member of the inhibitory of apoptosis protein (IAP) family of inhibitors of apoptosis, have been previously associated with different neurodegenerative disorders. This study indicated the existence of a novel NAIP splice variant. This isoform, NAIP deltaEx10-11, was found in tumor cell lines of different origin and in normal adult brain. Analysis of the putative protein predicted that the NAIP variant lacks part of the third BIR domain as well as the COOH-terminal tail of regular NAIP. This might suggest that it is endowed with a reduced antiapoptotic activity. This view is supported by the fact that NAIP-deltaEx10-11 mRNA and protein were much less expressed in the multidrug- and apoptosis-resistant HL60R leukemia than in its parental counterpart HL60. The opposite occurred for regular NAIP. Overall, the NAIP transcripts might be involved in tumor resistance to chemotherapeutic agents. PMID- 12127563 TI - Anticancer-drug-induced apoptotic cell death in leukemia cells is associated with proteolysis of beta-catenin. AB - beta-Catenin is a known regulator of cell-cell adhesion and transcriptional regulation. However, the role of beta-catenin and its regulation in non-adherent cells has not been examined. Therefore, we examined the role and fate of beta catenin during hematopoietic cell apoptosis using Jurkat T-acute lymphoblastic and U937 acute myeloblastic leukemia cells. The results presented here demonstrate that the treatment of Jurkat cells with the apoptosis inducers anti Fas, TRAIL, staurosporine, and etoposide induces proteolytic fragments of beta catenin, as did TRAIL and staurosporine in U937 cells. In Jurkat cells, beta catenin was cleaved at both the N- and C-terminal after anti-Fas addition. Cleavage of intact beta-catenin was completely inhibited by caspase selective protease inhibitors. There was a clear accumulation of the large proteolytic fragment in Jurkat cells treated with lactacystin or N-acetyl-leucyl leucyl methioninal (ALLM). These results suggest that both the proteasome and calpain may recognize the large beta-catenin fragment as a substrate for further degradation. Densitometric analysis demonstrated that the loss of intact beta catenin was more rapid in the cell nucleus (beta-catenin T1/2 of approximately 1.5h in cytoplasm and 0.5h in nucleus). Down-regulation of beta-catenin associated transcription was an early event in response to anti-Fas. These results suggest that beta-catenin plays a role in promoting Jurkat survival. PMID- 12127564 TI - Establishment of the T-cell large granular lymphocyte leukemia cell line MOTN-1 carrying natural killer-cell antigens. AB - A novel interleukin-2 (IL-2) dependent leukemia cell line MOTN-1 was established from the peripheral blood of a 63-year-old woman with T-cell large granular lymphocyte (LGL) leukemia in chronic phase. Primary peripheral blood leukemia cells were CD3+, CD5+, CD7+, CD56+, CD94+, CD161+, TcRalphabeta+, and HLA-DR+. The immunoprofile of the established cell line MOTN-1, however, showed CD3-, CD5 , CD7+, CD56+, CD94+, CD159+, CD161+, TcRalphabeta- and HLA-DR+; the MOTN-1 cells were cytoplasmatically positive for CD3varepsilon and the products of the T-cell receptor (TcR) genes beta and gamma. While the TcRbeta and TcRgamma genes were rearranged, the TcRdelta gene was found to be deleted. DNA fingerprinting and chromosome analysis identifying the t(2;6)(q?23;q?21) and t(12;18)(q13;q?22) alterations demonstrated the authenticity and the malignant nature of the cell line. The scientific significance of MOTN-1 lies in (1) the rarity of this type of leukemia cell lines, (2) the co-expression of various T- and natural killer (NK)-cell-associated markers, and (3) its unique chromosomal aberrations. PMID- 12127565 TI - Response of idiopathic hypereosinophilic syndrome to treatment with imatinib mesylate. AB - Idiopathic hypereosinophilic syndrome (HES) is a rare hematologic disorder characterized by persistent eosinophilia with organ involvement. Patients with HES have a poor prognosis, but the disease course can be heterogeneous. Treatment of HES has included corticosteroids, chemotherapeutic agents such as cyclophosphamide, vincristine, hydroxyrea, and most recently interferon-alpha (IFN-alpha) which has shown long-term beneficial effects. We herein report on a patient with HES who had disease resistant to steroids, and chemotherapy with 2 chlorodeoxyadenosine and cytarabine, but who had a significant response after only 8 days of treatment with imatinib mesylate 100mg daily. The possible mechanism of response is discussed. This observation may lead to a better understanding of the pathophysiology of HES, and may provide a new form of effective therapy for the disease. PMID- 12127567 TI - Endophilin-1: a multifunctional protein. AB - Endophilin-1, a cytoplasmic Src homology 3 (SH3) domain-containing protein, localises in brain presynaptic nerve termini. Endophilin dimerises through its N terminus, and participates at multiple stages in clathrin-coated endocytosis, from early membrane invagination to synaptic vesicle uncoating. Both its C terminal SH3 domain and N-terminus are required for endocytosis. Through its SH3 domain, endophilin bound to proline-rich domains (PRDs) in other endocytic proteins, including synaptojanin and dynamin. The N-terminal region possesses unique functions affecting lipid membrane curvature, through lysophosphatidic acid acyl transferase (LPAAT) activity and direct binding and tubulating activity. In addition to synaptic vesicle formation, endophilin-1 complexes with signalling molecules, including cell surface receptors, metalloprotease disintegrins and germinal centre kinase-like kinase (GLK). Therefore, endophilin 1 may serve to couple vesicle biogenesis with intracellular signalling cascades. PMID- 12127568 TI - Eps8 in the midst of GTPases. AB - Eps8, originally identified as a substrate for the kinase activity of the epidermal growth factor receptor (EGFR), displays a domain organization typical of a signaling molecule that includes a putative N-terminal PTB domain, a central SH3 domain, and a C-terminal "effector region". This latter region directs Eps8 localization within the cell and is sufficient to activate the GTPase, Rac, leading to actin cytoskeletal remodeling. Eps8 binds, through its SH3 domain, to either Abi1 (also called E3b1) or RN-tre. Abi1 scaffolds together Eps8 and Sos1, a dual specificity guanine nucleotide exchange factor for Ras and Rac proteins, thus facilitating the formation of a trimeric complex, in turn required for activation of Rac. On the other hand, RN-tre, a Rab5 GTPase activating protein, by entering in a complex with Eps8, inhibits EGFR internalization. Furthermore, RN-tre competes with Abi1 for binding to Eps8, diverting the latter from its Rac activating function. Thus, depending on its engagement in different complexes, Eps8 participates to EGFR signaling through Rac and endocytosis through Rab5. PMID- 12127569 TI - The Tec family of tyrosine kinases in T cells, amplifiers of T cell receptor signals. AB - ITK and Rlk/Txk are the predominant Tec family of tyrosine kinases expressed in T cells, and are involved in T cell antigen receptor mediated activation of T cells. These kinases require prior activation of Lck, Zap-70 and PI3-kinase for efficient activation. They share major substrates with both Lck and Zap-70, however the pathways they regulate are unclear. Recent evidence suggests that these kinases may not activate unique pathways, but instead serve as amplifiers for the upstream kinases Lck and Zap-70. This review will discuss the evidence for this view. PMID- 12127570 TI - Possible physiological roles of mitochondrial uncoupling proteins--UCPn. AB - Five mitochondrial uncoupling proteins exist in the human gemone: UCP2, expressed ubiquitously; UCP1, exclusively in brown adipose tissue (BAT); UCP3, predominantly in muscle; UCP4 and BMCP (UCP5), in brain. UCP4 is the ancestral prototype from which the other UCPn diverged. Findings on the level of organism and reconstituted recombinant proteins demonstrated that UCPn exhibit a protonophoric function, documented by overexpression in mice, L6 myotubes, INS1 cells, muscle, and yeast. In a few cases (yeast), this protonophoric function was correlated with elevated fatty acid (FA) levels. Reconstituted UCPn exhibited nucleotide-sensitive FA induced H(+) uniport. Two mechanisms, local buffering or FA cycling were suggested as an explanation. A basic UCPn role with mild uncoupling is to accelerate metabolism and reduce reactive oxygen species. UCP2 (UCP3) roles were inferred from transcriptional up-regulation mediated by FAs via peroxisome proliferator-activated receptors, cytokines, leptin signalling via hypothalamic pathway, and by thyroide and beta2 adrenergic stimulation. The latter indicated a role in catecholamine-induced thermogenesis in skeletal muscle. UCP2 (UCP3) may contribute to body weight regulation, although obesity was not induced in knockout (KO) mice. An obesity reduction in middle-aged humans was associated with the less common allele of -866 G/A polymorphism in the ucp2 gene promoter enhancing the exon 8 insertion: deletion transcript ratio. Up regulated UCP2 transcription by pyrogenic cytokines (tumour necrosis factor alpha (TNFalpha)) suggested a role in fever. UCP2 could induce type 2 diabetes as developed from obesity due to up-regulated UCP2 transcription by FAs in pancreatic beta-cells. UCPn might be pro-apoptotic as well as anti-apoptotic, depending on transcriptional and biochemical regulation. PMID- 12127571 TI - Mode of action of two inhibitory peptides from heptad repeat domains of the fusion protein of Newcastle disease virus. AB - Peptides derived from heptad repeat (HR) sequences of viral fusion proteins from several enveloped viruses have been shown to inhibit virus-mediated membrane fusion but the mechanism remains unknown. To further investigate this, the inhibition mechanism of two HR-derived peptides from the fusion protein of the paramyxovirus Newcastle disease virus (NDV) was investigated. Peptide N24 (residues 145-168) derived from HR1 was found to be 145-fold more inhibitory in a syncytium assay than peptide C24 (residues 474-496), derived from HR2. Both peptides failed to block lipid-mixing between R18-labeled virus and cells. None of the peptides interfered with the binding of hemagglutinin-neuraminidase (HN) protein to the target cells, as demonstrated by hemagglutining assays. When both peptides were mixed at equimolar concentrations, their inhibitory effect was abolished. In addition, both peptides induced the aggregation of negatively charged and zwitterionic phospholipid membranes. The ability of the peptides to interact with each other in solution suggests that these peptides may bind to the opposite HR region on the protein whereas their ability to interact with membranes as well as their failure to block lipid transfer suggest a second binding site. Taken together these results, suggest a mode of action for C24 and N24 in which both peptides have two different targets on the F protein: the opposite HR sequence and their corresponding domains. PMID- 12127572 TI - Two-domain arginine kinases from the clams Solen strictus and Corbicula japonica: exceptional amino acid replacement of the functionally important D(62) by G. AB - Arginine kinases (AKs) isolated from the adductor muscle of the clams Solen strictus and Corbicula japonica have relative molecular masses of 80 kDa as estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) in contrast to the 40 kDa AKs found in Mollusca and Arthropoda. The cDNAs encoding Solen and Corbicula AKs have open reading frames of 2175 nucleotides (724 amino acid protein) and 2172 nucleotides (723 amino acid protein), respectively. The amino acid sequence clearly indicates that Solen and Corbicula AKs have a two-domain structure: the first-domain includes residues 1-363 and the second-domain includes residue 364 to the end. There is approximately 60% inter domain amino acid identity. It is clear that gene-duplication and subsequent fusion occurred in the immediate ancestor of the clams Solen, Corbicula, and Pseudocardium. During substrate binding, it is proposed that AK undergoes a substrate-induced conformational change and that the hydrogen bond between D(62) and R(193) stabilizes the substrate-bound structure. However, in Solen and Corbicula two-domain AKs, D(62) is replaced by a G, and R(193) by A, S, or D. Consequently, the two-domain AKs can not form the stabilizing hydrogen bond. Nevertheless, the enzyme activity of Corbicula AK is comparable to those of other molluscan 40 kDa AKs. We assumed that the substrate-bound structure of the two domain AK is stabilized not by the hydrogen bond between D(62) and R(193) but by the bond between H(60) and D(197), characteristic of the unusual two-domain AKs. This explains why D(62) and R(193), which remain highly conserved in other AKs, have undergone amino acid replacements in Solen and Corbicula AKs. PMID- 12127573 TI - Rat kidney porphobilinogen deaminase kinetics. Detection of enzyme-substrate complexes. AB - BACKGROUND AND AIMS: Acute intermittent porphyria (AIP) is an inherited disease resulting from a reduced activity of the enzyme porphobilinogen deaminase (PBG D). The kidney is an important target for numerous porphyrinogenic drugs and it may contribute to the clinical manifestations of porphyric attacks. An evaluation of kidney PBG-D role in the AIP pathophysiology requires detailed information on kidney PBG-D properties, under normal conditions. METHODS: Rat kidney PBG-D was purified to homogeneity and initial reaction velocities were calculated by measuring uroporphyrinogen I formation at pH 8.2 for different incubation times (0-20 min) and over a wide range of substrate concentrations (0.8-66 microM). RESULTS: Purified rat kidney PBG-D is a monomeric enzyme showing only a single protein band after SDS-PAGE, Western blot and isoelectric focusing (pI 4.9). Its molecular mass is 40 +/- 2.3 kDa, determined by SDS-PAGE and 39.8 +/- 2 kDa by gel filtration chromatography. Rat kidney PBG-D has an unusual kinetic behaviour, exhibiting a deviation from the Michaelis-Menten hyperbola. PBG-D kinetic data required a fitting to an equation of higher degree, leading to the following apparent kinetic constants: K(1) = 2.08 +/- 0.01 microM and K(2) = 0.102 +/- 0.003 microM. CONCLUSION: The values of these constants fulfil the restriction 4K(2) < or = K(1)(2), necessary for the occurrence of isoenzymes, interpreted in this work as enzyme-substrate intermediates. The initial reaction velocity expression here defined, correlates with an enzyme carrying only one active site but allowing, through conformational changes, the detection of at least two enzyme-substrate intermediates formed during PBG-D reaction. PMID- 12127574 TI - Refolding and reactivation of calf intestinal alkaline phosphatase with excess magnesium ions. AB - It is well known that Mg(2+) is an essential component in many biological processes. This research investigated the courses of both the reactivation and the refolding in the absence and presence of Mg(2+) ions. Calf intestinal alkaline phosphatase (CIP) was extensively denatured in 3 M guanidine hydrochloride (GdnHCl) solution for 2 h. Under suitable renaturation conditions, about 60-70% of the activity was recovered in the absence and presence of different magnesium ion concentrations. The refolding processes followed two phase courses, whereas the reactivation processes were monophasic after dilution in proper solutions with or without Mg(2+). The magnesium ions affected both the reactivation and the refolding courses of unfolded CIP. A comparison of rate constants for the refolding of unfolded CIP with those for recovery of enzyme activity at different Mg(2+) concentrations showed that they were not synchronized. The activity recovery was speeded up due to the presence of Mg(2+) ions; while the refolding course of unfolded CIP was somewhat inhibited by the excess Mg(2+). PMID- 12127575 TI - Physical state changes of membrane lipids in human lung adenocarcinoma A(549) cells and their resistance to cisplatin. AB - The properties of membrane lipids in sensitive A(549) and resistant A(549)/DDP cells to cis-dichlorodiammine platinum[II] (cisplatin) were examined by combining different approaches. The results showed that fluorescence intensity (deltaF) of Merocyanine 540 (MC540) was 93.5 +/- 21.8 for the sensitive A(549) cells and 49.5 +/- 11.2 for the resistive A(549)/DDP cells, monitored by flow cytometry, which may indicate that membrane lipid packing of the sensitive A(549) cells were looser than that of the resistant A(549)/DDP cells. Diffusion rate of N-(7-nitro 2,1,3-benzoxadiazol-4-yl)-1,2-hexadecanoyl-Sn-glycero-3-phosphatidyl-ethanolamine (NBD-PE) was slower in A(549)/DDP cells than in A(549) cells as detected by fluorescence recovery after photobleaching (FRAP) technique. Fatty acid analysis of the membrane lipids showed 21.6, 27.0 and 31.8% increase in the amount of C(18:1), C(18:2) and C(18:3) fatty acid, respectively, in A(549) cells as compared to A(549)/DDP cells. The total amount of unsaturated fatty acids in the plasma membrane lipid is 69.13% +/- 2.2% for A(549), and 55.08% +/- 1.8% for A(549)/DDP cells, respectively. The resistance to cisplatin in A(549)/DDP cells was confirmed by the measurements of the transmembrane influx of Rhodamine-123, cisplatin or Bodipy-cisplatin by fluorescence assay and inductively coupled plasma mass spectrometry (ICP-MS). From the results described previously, it is concluded that changes in the membrane lipids "composition" cause a change in the physical state of the plasma membrane lipids and that this may be associated with the resistance of A(549)/DDP cells to cisplatin. PMID- 12127576 TI - Correlation between airway responsiveness and proteoglycan production by bronchial fibroblasts from normal and asthmatic subjects. AB - Asthma is characterized by an airway remodeling process involving altered extracellular matrix deposition such as collagen, fibronectin and proteoglycans. Proteoglycans determine tissue mechanical properties and are involved in many important biological aspects. Not surprisingly, it has been suggested that proteoglycan deposition may alter airway properties in asthma including airway hyperresponsiveness. In chronically inflamed airway tissues, fibroblasts likely represent an activated fibrotic phenotype that contributes to the excessive deposition of different extracellular matrix components. To investigate whether this was the case for proteoglycans, the production of hyaluronan, perlecan, versican, small heparan sulphate proteoglycans (HSPGs), decorin and biglycan was quantified in the culture medium of primary bronchial fibroblast cultures, established from four normal and six asthmatic subjects. Values were further correlated to the airway responsiveness (PC(20) methacholine) of donor subjects. Fibroblasts from subjects with the most hyperresponsive airways produced up to four times more total proteoglycans than cells from subjects with less hyperresponsive or normoresponsive airways. We observed a significant negative correlation between the PC(20) and perlecan, small HSPGs and biglycan, while such correlation was absent for decorin and close to significant for hyaluronan and versican. Altered proteoglycan metabolism by bronchial fibroblasts may contribute to the increased proteoglycan deposition in the bronchial mucosa and to airway hyperresponsiveness characterizing asthma. PMID- 12127577 TI - Structural characterization and phylogenetic relationships of myotoxin II from Atropoides (Bothrops) nummifer snake venom, a Lys49 phospholipase A(2) homologue. AB - In order to analyze its structure-function relationships, the complete amino acid sequence of myotoxin II from Atropoides (Bothrops) nummifer from Costa Rica was determined. This toxin is a Lys49-type phospholipase A(2) (PLA(2)) homologue, devoid of catalytic activity, structurally belonging to class IIA. In addition to the Asp49 --> Lys change in the (inactive) catalytic center, substitutions in the calcium-binding loop suggest that its lack of enzymatic activity is due to the loss of ability to bind Ca(2+). The toxin occurs as a homodimer of basic subunits of 121 residues. Its sequence has highest similarity to Lys49 PLA(2)s from Cerrophidion, Trimeresurus, Bothrops and Agkistrodon species, which form a subfamily of proteins that diverged early from Asp49 PLA(2)s present in the same species, as shown by phylogenetic analysis. The tertiary structure of the toxin was modeled, based on the coordinates of Cerrophidion godmani myotoxin II. Its exposed C-terminal region 115-129 shows several differences in comparison to the homologous sequences of other Lys49 PLA(2)s, i.e. from Agkistrodon p. piscivorus and Bothrops asper. Region 115-129 of the latter two proteins has been implicated in myotoxic activity, on the basis of the direct membrane-damaging of their corresponding synthetic peptides. However, peptide 115-129 of A. nummifer myotoxin II did not exert toxicity upon cultured skeletal muscle cells or mature muscle in vivo. Differences in several amino acid residues, either critical for toxicity, or influencing the conformation of free peptide 115-129 from A. nummifer myotoxin II, may account for its lack of direct membrane-damaging properties. PMID- 12127578 TI - Oxidative stress in mice is dependent on the free glucose content of the diet. AB - In animals, chronic intake of diets with high proportions of rapidly absorbable glucose promotes the development of insulin resistance. High levels of glucose can produce permanent chemical alterations in proteins and lipid peroxidation. delta-Aminolevulinate dehydratase (delta-ALA-D) is a sulfhydryl-containing enzyme essential for all aerobic organisms and is highly sensitive to the presence of pro-oxidants elements. The heme synthetic pathway is impaired in porphyria and a frequent coexistence of diabetes mellitus and porphyria disease has been reported in humans and experimental animal models, which can be casually linked to the delta-ALA-D inhibition found in diabetics. The present study was designed to evaluate the effect of two different diets, a high glucose (HG) diet and a high starch (HS) diet, on lipid peroxidation levels in different tissues (brain, liver, and kidney) and on delta-ALA-D activity (from liver and kidney) in mice. Plasma glucose and triglyceride levels were significantly higher in mice fed HG than in mice fed HS (P < 0.02 and P < 0.03, respectively). Thiobarbituric acid reactive species (TBA-RS) content was significantly increased in kidney and liver from HG diet-fed mice when compared with animals fed HS diets (P < 0.001). Hepatic delta-ALA-D activity of HG diet-fed animals was significantly lower than that of HS diet-fed animals (P < 0.01). The results of this study support the hypothesis that consumption of a diet with high free glucose can promote the development of oxidative stress that we tentatively attribute to hyperglycemia. PMID- 12127579 TI - Amino acid sequence of the major form of toad liver glutathione transferase. AB - To investigate structural relationship between amphibian and mammalian GSTs the complete amino acid sequence of the major form of glutathione transferase present in toad liver (Bufo bufo) was determined. The enzyme subunit is composed of 210 amino acid residues corresponding to a molecular mass of 24,178 Da. In comparison with the primary structure of amphibian bbGSTP1-1, toad liver GST showed 54% sequence identity. On the other hand, toad liver GST showed about 45-55% sequence identity when compared with other pi class GST and less then 25% identity with GST of other classes. Amino acid residues involved in the H site and in the key and lock structure of the toad enzyme are significantly different from those of bbGSTP1-1 and other mammalian pi class GST. On the basis of its structural and immunological properties the toad liver GST, indicated as bbGSTP2-2, could represent the prototype of a subset of the pi family. PMID- 12127580 TI - Activity and properties of alpha-L-fucosidase are dependent on the state of enterocytic differentiation of HT-29 colon cancer cells. AB - Previously we have demonstrated an impairment in the activity of alpha-L fucosidase in colon tumours. In order to establish an in vitro model to study this enzyme in colon cancer, we have determined the activity and properties of the enzyme during the differentiation of HT-29 colon cancer cells. Cultures were committed to differentiate into enterocyte-like cells by placing them in a culture medium without glucose for 18-21 days. The state of differentiation was evaluated by assaying the activity of enterocytic marker enzymes, and the acquisition of enterocyte morphology was assessed by electron microscopy. The alpha-L-fucosidase activity was determined using a fluorometric method. Intracellular levels of alpha-L-fucosidase activity are lower in non differentiated cells (3.0 +/- 1.01 U/mg) than in differentiated ones (9.2 +/- 4.09 U/mg) (P < 0.001). This variation is not due to a greater secretion of the enzyme to the culture medium, and properties such as pH optimum or the affinity towards substrate are not dependent on differentiation. The enzyme however, is more stable at acidic pH and at high temperatures, and V(max) is higher in differentiated cells. Moreover, in undifferentiated cells the enzyme is mainly in a monomeric form whereas multimeric forms of the enzyme appear only upon differentiation. Most of these changes are very similar to those previously observed between normal colon tissue and colon tumours. Thus, we suggest that differentiation of HT-29 colon cancer cells could be used as a model to study the alterations of the enzyme alpha-L-fucosidase during the progression of the tumoural process. PMID- 12127581 TI - Ikaros: a key regulator of haematopoiesis. AB - Ikaros is an essential transcription factor for normal lymphocyte development. Because of its interaction with a number of closely related factors, Ikaros is required for correct regulation of differentiation and cell proliferation in T- and B-cell lineages. Interestingly, Ikaros appears to function both as a transcriptional repressor and as an activator through its ability to bind a large number of nuclear factors, including components of both histone deacetylase and ATP-dependent chromatin remodelling complexes. In addition, nuclear localisation is important for Ikaros function--unlike most transcription factors, Ikaros is localised to discrete nuclear foci in lymphoid cells, suggesting it employs novel mechanisms to regulate transcription. PMID- 12127582 TI - Identification and characterization of a cathepsin B-like protease in Physarum sclerotium. AB - In response to dry stress the plasmodium of a true slime mold, Physarum polycephalum, undergoes formation of sclerotium, which is a dormant body resistant to desiccation. The sclerotium can germinate within several hours after addition of water, followed by generation of the plasmodium. In the early phase of the germination many enzymes and other proteins of the sclerotium are required for formation of the plasmodium. As dehydration of proteins often leads to destruction of their structure or reduction in their activity, it is important to elucidate whether the dehydrated enzymes are present as the intact in the sclerotium. In this study three peaks of protease activity were detected with anion exchange column chromatography of the extract from the sclerotia. From among them, an acid protease was purified to homogeneity by gel filtration column chromatography, hydroxyapatite column chromatography, acid treatment, and cation exchange column chromatography. Treatment of the protease fractions with pH 4.0 resulted in approximately 20-fold activation of the activity. The purified protease was a monomer with a molecular mass of 35 kDa. The optimum pH and temperature were 6.3 and 40 degrees C, respectively. Beta-casein, histone H1, and H2B were degraded by the 35 kDa protease, but human hemoglobin and human serum albumin were very poor substrates. In addition, the enzyme was sensitive to the cysteine protease inhibitors chymostatin, E-64, and leupeptin. These results indicate that, in the sclerotium, a premature form of a cathepsin B-like protease remains non-denatured under dehydrated conditions. PMID- 12127583 TI - The role of peroxisomes in the integration of metabolism and evolutionary diversity of photosynthetic organisms. AB - The peroxisome is a metabolic compartment serving for the rapid oxidation of substrates, a process that is not coupled to energy conservation. In plants and algae, peroxisomes connect biosynthetic and oxidative metabolic routes and compartmentalize potentially lethal steps of metabolism such as the formation of reactive oxygen species and glyoxylate, thus preventing poisoning of the cell and futile recycling. Peroxisomes exhibit properties resembling inside-out vesicles and possess special systems for the import of specific proteins, which form multi enzyme complexes (metabolons) linking numerous reactions to flavin-dependent oxidation, coupled to the decomposition of hydrogen peroxide by catalase. Hydrogen peroxide and superoxide originating in peroxisomes are important mediators in signal transduction pathways, particularly those involving salicylic acid. By contributing to the synthesis of oxalate, formate and other organic acids, peroxisomes regulate major fluxes of primary and secondary metabolism. The evolutionary diversity of algae has led to the presence of a wide range of enzymes in the peroxisomes that are only similar to higher plants in their direct predecessors, the Charophyceae. The appearance of seed plants was connected to the acquirement by storage tissues, of a peroxisomal fatty acid oxidation function linked to the glyoxylate cycle, which is induced during seed germination and maturation. Rearrangement of the peroxisomal photorespiratory function between different tissues of higher plants led to the appearance of different types of photosynthetic metabolism. The peroxisome may therefore have played a key role in the evolutionary formation of metabolic networks, via establishing interconnections between different metabolic compartments. PMID- 12127584 TI - Molecular species of glycinin in some soybean cultivars. AB - A(4) polypeptide-containing (Shirotsurunoko and York) and A(4) polypeptide lacking (Raiden and Suzuyutaka) soybean cultivars were used to investigate the heterogeneity of glycinin molecular species. Purification of glycinin by DEAE Toyopearl column chromatography afforded molecular species eluting before the glycinin fraction. Analysis of this fraction by SDS-polyacrylamide gel electrophoresis (SDS-PAGE) and sucrose density gradient centrifugation indicated that this protein consisted of A(1) and A(2) polypeptides. The A(4)-containing soybean cultivars contained less of this protein than the A(4)-lacking soybean cultivars, as exhibited by the size of the early peak appearing during column chromatography. Alkaline PAGE and N-terminal amino acid sequence analysis confirmed that the A(1)- and A(2)-rich molecular species in the A(4) polypeptide lacking cultivars consisted of the A(1a) and A(2) polypeptides. Estimation of the molecular mass by gel permeation chromatography and multi-angle laser light scattering (GPC-MALLS) indicated that the A(1a)- and A(2)-rich molecular species were similar to a monomer of glycinin. PMID- 12127585 TI - Accumulation of tyrosol glucoside in transgenic potato plants expressing a parsley tyrosine decarboxylase. AB - As part of the response to pathogen infection, potato plants accumulate soluble and cell wall-bound phenolics such as hydroxycinnamic acid tyramine amides. Since incorporation of these compounds into the cell wall leads to a fortified barrier against pathogens, raising the amounts of hydroxycinnamic acid tyramine amides might positively affect the resistance response. To this end, we set out to increase the amount of tyramine, one of the substrates of the hydroxycinnamoyl CoA:tyramine N-(hydroxycinnamoyl)-transferase reaction, by placing a cDNA encoding a pathogen-induced tyrosine decarboxylase from parsley under the control of the 35S promoter and introducing the construct into potato plants via Agrobacterium tumefaciens-mediated transformation. While no alterations were observed in the pattern and quantity of cell wall-bound phenolic compounds in transgenic plants, the soluble fraction contained several new compounds. The major one was isolated and identified as tyrosol glucoside by liquid chromatography-electrospray ionization-high resolution mass spectrometry and NMR analyses. Our results indicate that expression of a tyrosine decarboxylase in potato does not channel tyramine into the hydroxycinnamoyl-CoA:tyramine N (hydroxycinnamoyl)-transferase reaction but rather unexpectedly, into a different pathway leading to the formation of a potential storage compound. PMID- 12127586 TI - (+)-(10R)-Germacrene A synthase from goldenrod, Solidago canadensis; cDNA isolation, bacterial expression and functional analysis. AB - Profiling of sesquiterpene hydrocarbons in extracts of goldenrod, Solidago canadensis, by GC-MS revealed the presence of both enantiomers of germacrene D and lesser amounts of germacrene A, alpha-humulene, and beta-caryophyllene. A similarity-based cloning strategy using degenerate oligonucleotide primers, based on conserved amino acid sequences in known plant sesquiterpene synthases and RT PCR, resulted in the isolation of a full length sesquiterpene synthase cDNA. Functional expression of the cDNA in E. coli, as an N-terminal thioredoxin fusion protein using the pET32b vector yielded an enzyme that was readily purified by nickel-chelate affinity chromatography. Chiral GC-MS analysis of products from of (3)H- and (2)H-labelled farnesyl diphosphate identified the enzyme as (+)-(10R) germacrene A synthase. Sequence analysis and molecular modelling was used to compare this enzyme with the mechanistically related epi-aristolochene synthase from tobacco. PMID- 12127587 TI - Fungal melanin inhibitor and related compounds from Penicillium decumbens. AB - Two polyketides, decumbenones A and B, and versiol were isolated from the culture filtrate of the fungus, Penicillium decumbens. Their respective structures were 1 (2,8-dihydroxy-1,2,6-trimethyl-1,2,6,7,8,8a-hexahydronaphthalen-1-yl)-3-hydroxy-1 propanone and 1-(2,8-dihydroxy-1,2,6-trimethyl-1,2,4a,5,6,7,8,8a octahydronaphthalen-1-yl)-3-hydroxy-1-propanone based on NMR spectroscopic data, chemical conversion, and X-ray analysis. Decumbenone A inhibited melanization in Magnaporthe grisea, the rice blast pathogen, whereas decumbenone B like versiol did not. PMID- 12127588 TI - Galiellalactone and its biogenetic precursors as chemotaxonomic markers of the Sarcosomataceae (Ascomycota). AB - (-)-Galiellalactone is a hexaketide metabolite with interesting pharmacological activities which was detected in four strains of Galiella rufa (Sarcosomataceae, Ascomycota) and in two unidentified fungi shown by their 18S rDNA sequences also to belong to the Sarcosomataceae. These were a wood-inhabiting apothecial species from Chile and an endophytic isolate from Cistus salviifolius (Sardinia). Other members of the family (Urnula helvelloides, one Strumella coryneoidea isolate) produced no galiellalactone but merely hexaketides structurally related to galiellalactone precursors, whereas a third group of species (Sarcosoma latahensis, Strumella griseola, one S. coryneoidea isolate) lacked hexaketide production altogether. Despite thorough screening programmes, galiellalactone and its precursors have not yet been found in any fungus outside the Sarcosomataceae and may thus be a chemotaxonomic marker of the family, supporting its current phylogenetic definition. Two pentaketide derivatives of the 6-pentyl-alpha-pyrone type were found in all G. rufa strains as well as in A111-95 and the hexaketide producing S. coryneoidea isolate. PMID- 12127589 TI - Diversity of cuticular wax among Salix species and Populus species hybrids. AB - The leaf cuticular waxes of three Salix species and two Populus species hybrids, selected for their ability to produce high amounts of biomass, were characterized. Samples were extracted in CH(2)Cl(2) three times over the growing season. Low kV SEM was utilized to observe differences in the ultrastructure of leaf surfaces from each clone. Homologous series of wax components were classified into organic groups, and the variation in wax components due to clone, sample time, and their interaction was identified. All Salix species and Populus species hybrids showed differences in total wax load at each sampling period, whereas the pattern of wax deposition over time differed only between the Salix species. A strong positive relationship was identified between the entire homologous series of alcohols and total wax load in all clones. Similarly strong relationships were observed between fatty acids and total wax load as well as fatty acids and alcohols in two Salix species and one Populus species hybrid. One Salix species, S. dasyclados, also displayed a strong positive relationship between alcohols and alkanes. These data indicate that species grown under the same environmental conditions produce measurably different cuticular waxes and that regulation of wax production appears to be different in each species. The important roles cuticular waxes play in drought tolerance, pest, and pathogen resistance, as well as the ease of wax extraction and analysis, strongly suggest that the characteristics of the cuticular wax may prove to be useful selectable traits in a breeding program. PMID- 12127590 TI - Scutellarein 4'-methyl ether glycosides as taxonomic markers in Teucridium and Tripora (Lamiaceae, Ajugoideae). AB - The flavonoid profiles of two monotypic genera, Teucridium and Tripora, have been studied by analytical methods. These genera were formerly placed in the Verbenaceae, but are now classified in the Lamiaceae, subfamily Ajugoideae. The major flavonoids of both genera were identified as glycosides of scutellarein 4' methyl ether (5,6,7-trihydroxy-4'methoxyflavone) and acacetin (5,7-dihydroxy-4' methoxyflavone). The new flavone glycoside, scutellarein 4'-methyl ether 7-O rutinoside, was isolated from Teucridium parvifolium and the rare scutellarein 4' methyl ether 7-O-glucuronide from Tripora divaricata. The latter compound has only been reported previously in the related genus Clerodendron. The potential of these flavonoids as taxonomic markers for the tribe Ajugoideae is discussed. PMID- 12127591 TI - Guaianolides from Viguiera gardneri inhibit the transcription factor NF-kappaB. AB - Five guaianolides and a germacrolide were isolated from the leaf rinse extract of Viguiera gardneri (Asteraceae), together with known compounds. All compounds were detected in glandular trichomes collected from the leaves and were analyzed by HPLC. Structure elucidation was based on the analysis of spectroscopic data. Low energy conformations were obtained by quantum mechanical calculations. Three closely related guaianolides which were isolated as the main compounds were studied for their anti-inflammatory activity using the transcription factor NF kappaB as molecular target. NF-kappaB DNA binding was inhibited at sesquiterpene lactones concentrations of 10 or 50 microM. PMID- 12127592 TI - Isofuranonaphthoquinone derivatives from cultures of the lichen Arthonia cinnabarina (DC.) Wallr. AB - Two isofuranonaphthoquinone derivatives, named arthoniafurones A (1-acetyl-8 hydroxynaphtho[2,3-c]furan-4,9-dione) and B [1-acetyl-4,8-dihydroxynaphtho[2,3 c]furan-9(4H)-one], were isolated from a spore-derived culture of the mycobiont of the lichen Arthonia cinnabarina, that is new to Japan. Bostrycoidin and 8-O methylbostrycoidin were also identified in the A. cinnabarina culture. PMID- 12127593 TI - Apotirucallane triterpenoids from Luvunga sarmentosa (Rutaceae). AB - The leaves of Luvunga sarmentosa (Bl.) Kurz. yielded eight apotirucallane triterpenoids named luvungins A-G, and 1alpha-acetoxyluvungin A. Characteristic of the structure are the seven-membered lactone-ring A, the alpha-hydroxyl or alpha-acetoxyl group at C-7 and an oxygen bridge in the side chain giving five-, six- or seven-membered rings, respectively. Because of a hemiacetal function at C 21, luvungin C occurred as a mixture of 21-epimers. The structures have been elucidated on the basis of MS and NMR spectral data. In addition, two known coumarins ostruthin (6-geranyl-7-hydroxycoumarin) and 8-geranyl-7-hydroxycoumarin as well as five known triterpenes friedelin, flindissone, melianone, niloticin and limonin were isolated. PMID- 12127594 TI - Immunological mechanisms and the spectrum of psychiatric syndromes in Alzheimer's disease. AB - Pathological, genetic and epidemiological studies support the opinion that inflammatory mechanisms are involved in the pathogenesis of Alzheimer's disease (AD). Recent pathological and neuroradiological (PET) data show that activation of microglia is an early pathogenic event that precedes the process of severe neuropil destruction in AD brains. In this paper we review the evidence that inflammatory mediators can play a pathogenic role in some behavioural disorders frequently encountered during the clinical course in AD patients. Motivational disturbances are the most striking of the depressive symptoms in AD and can be present in a preclinical stage of the disease. Experimental animal studies and clinical trials in humans have shown that cytokines can induce similar symptoms which were described as 'sickness behaviour' or 'depressive-like' state. Delirious states are frequently observed in more advanced stages of dementia. Delirium is generally considered the result of an imbalance in neurotransmitter systems with severe deficits of the cholinergic systems. Animal studies show that pro-inflammatory cytokines, such as interleukin-1, induce a reduced activity of the cholinergic system. In AD, the release of cytokines would exacerbate any already existing disturbances in the cholinergic neurotransmission. This could explain the susceptibility of demented patients to delirium provoked by a wide variety of trivial incidents that are accompanied by an acute phase response. The data reviewed in this paper suggest that it could be worthwhile employing a neuroimmunological approach to study at molecular level the pathogenesis of a broad spectrum of behavioural disturbances common in the clinical course of AD patients. PMID- 12127595 TI - Modeling the prevalence and incidence of Alzheimer's disease and mild cognitive impairment. AB - A number of systems have been proposed for classifying older adults who suffer from cognitive impairment or decline but do not yet meet criteria for Alzheimer's disease (AD). The classification, Mild Cognitive Impairment (MCI), has attracted much attention. It uses relatively specific diagnostic criteria and individuals who meet these criteria appear to be at substantial risk for the development of AD. However, little data is available to define the prevalence of MCI in any age group. We propose a simple mathematical model for the progression of patients from Non-Affected (NA) to MCI to AD. This first-order Markov model defines the likely prevalence of MCI at specific ages. Primary assumptions of the model include an AD prevalence of 1% at age 60 increasing to 25% at age 85 and a conversion rate from MCI to AD of 10% constant across all ages considered. We used the best available information for our model and found (1) that the MCI prevalence increased from 1% at age 60 to 42% at age 85 and (2) that the conversion rate from NA to MCI increased from 1% per year at age 60 to 11% at age 85. In conclusion, this model allows estimation of prevalence of MCI and conversion from NA to MCI based upon known prevalences of AD, conversion rates of MCI to AD, and death rates. Due to its substantial prevalence, MCI may be an important target for screening and possible intervention. PMID- 12127596 TI - The combined DEX-CRH test in treatment course and long-term outcome of major depression. AB - Neuroendocrine studies strongly suggest that the hypothalamic-pituitary adrenocortical (HPA) system plays a crucial role in the development and course of depression. The interaction between the disease process and HPA system function in long-term course, however, is unclear. Since improvement of HPA system deterioration has been demonstrated to be associated with treatment response, the question has arisen whether the course of therapy response as reflected by, for example, early improvement or response (after 1 or 2 weeks of therapy) is also based on HPA system dysfunction and whether the course of HPA regulation during treatment is only a state marker or has additional predictive implications for long-term outcome. In order to elucidate these questions a long-term study was carried out to investigate whether HPA system disturbance is associated (1) with the course of treatment response, predominantly early treatment response, during acute depression and (2) with the long-term course of depression, i.e. number of episodes. Twenty patients with affective disorders who participated in earlier controlled antidepressant treatment studies over 6 weeks were enrolled in an exploratory follow-up study. Using the combined DEX/CRH test it was demonstrated that (1) early improvement, early treatment response and beneficial treatment outcome after 6 weeks were associated with a lower HPA system activity and that (2) in long-term course of depression the HPA system deterioration increases in parallel with the number of previous episodes. These findings suggest that HPA system alterations are closely related to treatment response and long-term outcome of depression. PMID- 12127597 TI - Effects of REM sleep awakenings and related wakening paradigms on the ultradian sleep cycle and the symptoms in depression. AB - In 1975 Vogel and coworkers published their classical study where they compared selective rapid eye movement (REM) sleep deprivation by brief awakenings to a control intervention paradigm in depressed patients. The superior antidepressive impact of the first procedure was attributed to the REM pressure accumulating during the treatment period. The laborious procedure and the considerable effort necessary to evaluate the sleep profiles in real time have prevented similar experiments so far. Based on artificial neural networks we developed a software for the real time detection of REM sleep. In combination with an alarm system the algorithm allowed us to wake up subjects automatically and to reduce REM sleep by about 50%. The procedure was then compared to a modified nonREM intervention paradigm for a treatment period of ten consecutive nights in depressed patients (n(1)=14, n(2)=13). These simultaneously received moderate dosages of Trimipramine. We found a strong and robust but not significantly different reduction of the average Hamilton rating scores (33 and 41% of baseline levels). While the REM sleep awakenings shortened the sleep cycle duration considerably, our nonREM intervention paradigm lengthened the ultradian alternations. Both effects might be interpreted as a challenge imposed on the nonREM-REM alternating mechanism possibly responsible for the antidepressive impact. A different timing of the control interventions might have caused the discrepancy between our findings and those of Vogel and coworkers. PMID- 12127598 TI - The possible pathophysiological role of plasma nitric oxide and adrenomedullin in schizophrenia. AB - Evidence is accumulating for a possible role of nitric oxide (NO) in schizophrenia. Adrenomedullin (AM) induces vasorelaxation by activating adenylate cyclase and also by stimulating the release of NO. AM immune reactivity is present in the brain consistent with a role as neurotransmitter. We aimed to examine plasma levels of nitrite (a metabolite of NO) and AM in schizophrenic patients. Eighty-two patients with schizophrenia and 21 healthy control subjects were included in this study. DSM-IV diagnosis of chronic schizophrenia was established on the basis of independent structured clinical interviews and review of records by two qualified psychiatrists which included the Brief Psychiatric Rating Scale (BPRS), The Scale for the Assessment of Negative Symptoms (SANS) and The Scale for the Assessment of Positive Symptoms (SAPS). Total nitrite and AM have been studied in plasma. The mean values of plasma nitrite and AM levels in schizophrenic group were significantly higher than control values, respectively (P=0.03, P<0.0001). AM levels of schizophrenic patients were three fold higher than controls. In correlation analyses, there were statistically significant positive correlations between AM level and SAPS-delusion subscale (r=0.27, P=0.04); SAPS-bizarre behavior subscale (r=0.28, P=0.03) and SAPS-total (r=0.36, P=0.005). There is no correlation between total nitrite and AM levels (r=0.11, P=0.31). Both NO and AM may have a pathophysiological role in schizophrenia, and clinically symptomatology and prognosis of schizophrenia. This subject needs further study including treatment response and subtypes of schizophrenia. PMID- 12127599 TI - The increased activity of plasma manganese superoxide dismutase in tardive dyskinesia is unrelated to the Ala-9Val polymorphism. AB - That tardive dyskinesia (TD) may have its origins in free-radical toxicity has stimulated investigations into one enzyme important in the control of oxidative free radicals: superoxide dismutase (SOD). The manganese-containing form of this enzyme (MnSOD) is the major superoxide scavenger in mitochondria; a weak association between a functional genetic polymorphism (Ala-9Val) in the mitochondrial targeting sequence (MTS) of this enzyme and TD has been reported in a Japanese population. We have undertaken to determine both the plasma activity of MnSOD and the association of the Ala-9Val polymorphism in a well-matched series of male Chinese schizophrenic patients with (n=42) and without (n=59) TD, and normal male controls (n=50). MnSOD activity was elevated in the TD subjects over those without TD (P<0.05) and normal controls (P<0.05), an effect that was independent of age, age at first antipsychotic treatment, drug dosage and duration of illness. A significant positive correlation between total AIMS score and MnSOD activity was also observed (P<0.0001). No significant reduction in the frequency of the Ala allele was observed in the TD group (0.14) below non-TD (0.18) or control subjects (0.17); nor was there any relationship between MnSOD activity and the polymorphism. There was no difference between the mean AIMS scores for the two genotypes (V/V and A/V) in the TD group. We conclude that while we have further evidence of a disturbance in the mechanisms regulating oxidative free radicals in TD, this effect is not under the control of the genetic polymorphism investigated here. PMID- 12127600 TI - Familiality of clinical characteristics in schizophrenia. AB - Few studies have assessed the familiality of clinical characteristics in schizophrenia. Therefore, we set out to investigate the familiality of the following characteristics; age of onset, course of disorder, employment status at onset, impairment during disorder, marital status at onset, mode of onset and premorbid functioning. Clinical characteristics were recorded using the Operational Criteria Checklist for Psychotic Illness for 155 subjects with an RDC diagnosis of schizophrenia, schizoaffective disorder, or psychosis of unknown origin, from 61 families multiply affected by schizophrenia. Age of onset, course of disorder, impairment during disorder, mode of onset, and premorbid functioning were shown to be familial. The familiality of these clinical characteristics supports their use in the delineation of homogeneous subsets for future genetic studies. PMID- 12127601 TI - Decreased production of interleukin-2 (IL-2), IL-2 secreting cells and CD4+ cells in medication-free patients with schizophrenia. AB - There are a number of indications that schizophrenia is associated with changes in the immune system. Although functional studies have mostly demonstrated decreased in vitro production of IL-2 by peripheral blood mononuclear cells (PBMCs) stimulated with mitogen, the reason is unclear. The aim of the study was to explore the relationship between IL-2 production and CD4+ cells which mainly secret IL-2 in non-Caucasian patients with schizophrenia. Blood CD4+ cells and mitogen-stimulated IL-2 secreting cells identified by an immunohistochemical study with the alkaline phosphatase/anti-alkaline phosphatase (APAAP) technique, and in vitro IL-2 production with radioimmunometric assay (RIA) were measured in 30 schizophrenic patients and 30 normal control subjects matched for sex, age and race. The results showed that blood CD4+ cells and mitogen-induced IL-2 secreting cells and IL-2 production were significantly lower in schizophrenic subjects than in the normal controls. There was significantly positive correlation between CD4+ cells and IL-2 production in normal controls but not in patients. These findings suggest that immune disturbance may be present in schizophrenic patients. The lower in vitro IL-2 production is probably related to the decreased number of T cells that secret IL-2, as well as to the intrinsic disorder of the patients' T cells. PMID- 12127602 TI - Characterization of children of bipolar parents by parent report CBCL. AB - In past research the Child Behavior Checklist (CBCL) has differentiated among various diagnostic categories for children and adolescents. However, research has not been conducted on whether the CBCL differentiates among diagnostic categories for children at high risk for development of psychopathology. This study compares four diagnostic groups [bipolar disorder (BD), attention/deficit-hyperactivity disorder (ADHD), Depressed/Anxious and No Diagnosis] within a cohort of 58 children of bipolar parents to determine whether their CBCL scores will replicate the scores of children not at high risk for bipolar disorder. The cohort of children of bipolar parents received elevated scores on the CBCL scales in comparison with non-clinical populations. In addition, the CBCL distinguished between children of bipolar parents with and without clinical disorders. Finally the BD group differed from the ADHD group only on the Aggressive Behaviors, Withdrawn and Anxious/Depressed subscales of the CBCL. Therefore the CBCL did not discriminate between the BD and ADHD groups as it had in previous studies of children with BD and unspecified family history. It is possible that this discrepancy is due to a group of children of bipolar parents with ADHD who are currently prodromal for bipolar disorder and therefore received higher scores on the CBCL based on prodromal symptomatology. A longitudinal follow-up of this cohort is necessary to ascertain whether this is the case. PMID- 12127603 TI - A comparison of clinical ratings with vocal acoustic measures of flat affect and alogia. AB - In this report we compare clinical ratings of flat affect and alogia with objective measures of the patient's speech prosody and productivity. Thirty schizophrenic patients were evaluated with the Scale for the Assessment of Negative Symptoms (SANS) and the St. Hans Rating Scale for extra pyramidal side effects. Their speech was recorded and analyzed acoustically for measures of prosody and productivity. Correlations between pairs of SANS items and acoustic measures (e.g. Vocal Inflection and Fundamental Frequency Variance) were weak. The SANS item and global ratings were strongly related. Ratings of bradykinesia overlapped with the SANS ratings but not with the acoustic measures. The SANS ratings appear to be derived from global impressions, with diffuse confounding of flat affect with alogia, and with bradykinesia. Acoustic analysis has the potential to provide objective measures that may help develop operational definitions of these constructs and enhance clinical assessment. PMID- 12127604 TI - Usefulness of standard electrocardiographic parameters for predicting cardiac events after acute myocardial infarction during modern treatment era. AB - Comprehensive information about the independent value of different electrocardiographic (ECG) variables in predicting cardiac events after acute myocardial infarction (AMI) in the era of modern therapy is limited. Patients (n = 1,034) underwent standard electrocardiography from 5 to 7 days after an AMI. Several time intervals and PQRST abnormalities were analyzed from the electrocardiogram. During a mean +/- SD follow-up of 752 +/- 301 days on average, 42 patients (4%) experienced cardiac death, and 259 patients (25%) a cardiac death, nonfatal AMI, or unstable angina. Several ECG variables had a significant association with cardiac events in univariate comparisons. After adjustment for all risk variables in the Cox hazards model, lateral ST-segment depression (hazard ratio [HR] 4.76, 95% confidence interval [CI] 2.40 to 9.44, p <0.0001) and atrial abnormality with a terminal deflection of the P wave > or =0.1 mV deep and > or =40 ms in duration in lead V(1) (HR 2.46, 95% CI 1.25 to 4.82, p = 0.009) were the only ECG variables that independently predicted cardiac death. Lateral ST-segment depression also predicted the combined end point of cardiac death/nonfatal AMI/unstable angina in this model (HR 1.49, 95% CI 1.14 to 1.94, p = 0.003). In conclusion, lateral ST depression and atrial abnormality on the electrocardiogram are independent predictors of cardiac death after AMI. Lateral ST depression is also associated with ischemic cardiac events. PMID- 12127605 TI - Comparison between visual assessment and quantitative angiography versus fractional flow reserve for native coronary narrowings of moderate severity. AB - We tested the hypothesis that experienced interventional cardiologists can identify patients with fractional flow reserve (FFR) <0.75 either by visual assessment of the angiogram or by quantitative coronary angiography (QCA). Estimation of the significance of moderate lesions is difficult. FFR can determine the physiologic significance of a stenosis. Data comparing visual assessment and QCA of moderate lesions with FFR are limited. FFR was measured in 83 moderate lesions defined as having a 40% to 70% stenosis by visual inspection. An FFR <0.75 was considered "significant." Lesions were visually assessed by 3 experienced interventional cardiologists and their significance estimated. QCA was performed. Both analyses were compared with FFR. FFR averaged 0.82 +/- 0.11 and was <0.75 in 15 of 83 lesions (18%). The reviewers' classification was concordant with the FFR in about half the lesions. Concordance between reviewers was poor (Spearman's rho = 0.36). Visual assessment resulted in good sensitivity (80%) and negative predictive value (91%), but poor specificity (47%) and positive predictive value (25%) compared with FFR. By QCA, no patient with stenosis <60% or minimal luminal diameter >1.4 mm had FFR <0.75. QCA did not discriminate the significance of lesions outside of these parameters. Thus, neither visual assessment of an angiogram by experienced interventional cardiologists nor QCA can accurately predict the significance of most moderate narrowings. PMID- 12127606 TI - Relation of percutaneous coronary intervention of complex lesions to clinical outcomes (from the NHLBI Dynamic Registry). AB - Advances in percutaneous coronary intervention (PCI) have reduced complications but expanded indications. We used the National Heart, Lung, and Blood Insitute Dynamic Registry to determine clinical outcomes up to 1 year after PCI in 2,839 patients with at least 1 treated complex lesion (defined as a lesion showing evidence of thrombus, calcification, bifurcation or ostial location, or chronic occlusion) and 1,790 patients with only simple lesions treated. Complex lesion interventions were associated (p <0.05) with more sustained major dissections, distal embolization, side branch occlusion, and persistent flow reduction. Patients with treated complex lesions had a lower procedural success rate (93.8% vs 97.3%, p <0.001) and increased in-hospital rates (p <0.001) of death (2.0% vs 0.6%), death/myocardial infarction [MI] (5.2% vs 2.4%), or death/MI/coronary artery bypass graft [CABG] surgery (6.5% vs 2.9%). After adjustment for potential confounders, patients treated for multiple complex lesions were more likely to experience the in-hospital combined end points of death/MI (odds ratio 3.22, 95% confidence interval 2.10 to 4.92), or death/MI/CABG (odds ratio 2.55, 95% confidence interval 1.71 to 3.80). At 1 year, patients with treated complex lesions were more likely (p <0.001) to die (6.2% vs 3.7%), suffer death/MI (11.7% vs 7.5%), or death/MI/CABG/repeat PCI (27.2% vs 23.4%). Patients treated for multiple complex lesions were approximately 50% more likely to die or to have major adverse events than with patients only treated for simple lesions. An increased in-hospital adverse clinical event rate was independently noted for thrombotic, bifurcation, and calcified lesions, and bifurcation lesions had worse long-term event rates. PMID- 12127607 TI - Six-month angiographic and 12-month clinical follow-up of MultiLink long (25 to 35 mm) stents for long coronary narrowings in patients with angina pectoris. AB - There are limited prospective angiographic data on stent deployment for long coronary lesions. This multicenter prospective study enrolled 120 patients with a single de novo stenosis >20 mm in length, in a native vessel > or =3 mm diameter, suitable for a MultiLink stent 25 to 35 mm in length with additional stent deployment if required. Quantitative angiography before and immediately after stenting and at 6-month follow-up assessed restenosis for the complete lesion and for 5-mm segments of the stented and adjacent nonstented vessel. By 1 year, myocardial infarction had occurred in 3% and target vessel repeat revascularization in 12% of patients. The mean stented length (35.8 +/- 14.6 mm) closely matched mean lesion length (30.1 +/- 13.5 mm). Restenosis to > or =50% diameter loss occurred in 32% of patients, but to > or =70% in only 8%. Of the 147 segments (5 mm in length) with baseline stenosis <25%, only 3 patients (2%) developed restenosis of > or =50%, and only in 1 of these was it > or =70%. Stenting of long narrowings is associated with good clinical outcome and a low rate of severe restenosis. Mildly diseased segments of long lesions covered by a stent rarely became severely narrowed and had negligible influence on the overall restenosis rate. These data support a strategy of full lesion coverage by stent deployment. PMID- 12127608 TI - Reliability of resolution of ST-segment elevation after coronary reperfusion in predicting myocardial salvage in anterior wall acute myocardial infarction. AB - Resolution of ST-segment elevation (ST resolution) after reperfusion therapy has been shown to correlate with improved left ventricular (LV) function in patients with acute myocardial infarction (AMI). However, not all patients with ST resolution have preserved LV function. We evaluated the clinical significance of ST resolution in 129 patients with anterior wall AMI who underwent successful coronary recanalization within 6 hours after symptom onset by studying the relation to myocardial blush grade, another angiographic marker of myocardial reperfusion. A reduction of > or =50% in ST-segment elevation after recanalization was defined as ST resolution. Ninety-eight patients had ST resolution and 31 patients did not. Patients with ST resolution were subdivided into 2 groups according to myocardial blush grade after recanalization: 67 patients with blush grade 2 or 3, and 31 with blush grade 0 or 1. The QRS score after recanalization was higher (5.9 +/- 1.9 vs 3.4 +/- 2.0, p <0.01) and predischarge LV ejection fraction was lower (39 +/- 8% vs 57 +/- 9%, p <0.01) in patients with blush grade 0 or 1 than in those with blush grade 2 or 3. However, the QRS score after recanalization and the predischarge LV ejection fraction were similar in patients who had ST resolution with blush grade 0 or 1 and in those without ST resolution. Our findings suggest that ST resolution after recanalization does not consistently predict myocardial salvage in patients with anterior AMI. PMID- 12127609 TI - Thresholds for the electrocardiographic change range of biochemical markers of acute myocardial infarction (GUSTO-IIa data). AB - The definition of acute myocardial infarction (AMI) is increasingly dependent on levels of biochemical markers, including troponin. We aimed to determine the levels of biochemical markers associated with definite evolutionary electrocardiographic (ECG) changes in patients with ST-segment elevation myocardial infarction. By examining the database of 855 patients from the troponin substudy of GUSTO-IIa, we selected patients with ST-segment elevation at baseline, evidence of evolution of the QRS, T, and ST-segment waveforms on the predischarge electrocardiogram, and 3 measurements of > or =1 of the following: creatine kinase (CK)-MB, troponin T, or troponin I. We identified 222 patients with evolutionary ECG changes. The median QRS score for this population was 5 points; the fifth percentile was 1. For patients with 3 CK-MB measurements, the fifth percentile as a multiple of the upper limit of normal was 2.1 (upper limit of normal 7.0 ng/ml). For patients with troponin T measurements, the fifth percentile as a multiple of the upper limit of normal was 11.0 (upper limit of normal 0.1 ng/ml). For patients with troponin I measurements, the fifth percentile as a multiple of the upper limit of normal was 3.8 (upper limit of normal 1.5 ng/ml). This study revealed that 95% of the patients with definite ECG evidence of AMI had a more than twofold increase in CK-MB and more than a 3- to 11-fold increase in troponin. PMID- 12127610 TI - Usefulness of clinical risk markers and ischemic threshold to stratify risk in patients undergoing major noncardiac surgery. AB - The risk of cardiac events in patients undergoing major noncardiac surgery is dependent on their clinical characteristics and the results of stress testing. The purpose of this study was to develop a composite approach to defining levels of risk and to examine whether different approaches to prophylaxis influenced this prediction of outcome. One hundred forty-five consecutive patients (aged 68 +/- 9 years, 79 men) with >1 clinical risk variable were studied with standard dobutamine-atropine stress echo before major noncardiac surgery. Risk levels were stratified according to the presence of ischemia (new or worsening wall motion abnormality), ischemic threshold (heart rate at development of ischemia), and number of clinical risk variables. Patients were followed for perioperative events (during hospital admission) and death or infarction over the subsequent 16 +/- 10 months. Ten perioperative events occurred in 105 patients who proceeded to surgery (10%, 95% confidence interval [CI] 5% to 17%), 40 being cancelled because of cardiac or other risk. No ischemia was identified in 56 patients, 1 of whom (1.8%) had a perioperative infarction. Of the 49 patients with ischemia, 22 (45%) had 1 or 2 clinical risk factors; 2 (9%, 95% CI 1% to 29%) had events. Another 15 patients had a high ischemic threshold and 3 or 4 risk factors; 3 (20%, 95% CI 4% to 48%) had events. Twelve patients had a low ischemic threshold and 3 or 4 risk factors; 4 (33%, 95% CI 10% to 65%) had events. Preoperative myocardial revascularization was performed in only 3 patients, none of whom had events. Perioperative and long-term events occurred despite the use of beta blockers; 7 of 41 beta blocker-treated patients had a perioperative event (17%, 95% CI 7% to 32%); these treated patients were at higher anticipated risk than untreated patients (20 +/- 24% vs 10 +/- 19%, p = 0.02). The total event rate over late follow-up was 13%, and was predicted by dobutamine-atropine stress echo results and heart rate response. PMID- 12127611 TI - Effect of catheter-based iridium-192 gamma brachytherapy on the added risk of restenosis from diabetes mellitus after intervention for in-stent restenosis (subanalysis of the GAMMA I Randomized Trial). AB - Catheter-based intracoronary radiation therapy using iridium-192 (Ir-192) has been shown to be effective in reducing recurrent coronary restenosis after initial percutaneous treatment of in-stent restenosis. Patients with diabetes mellitus (DM) have a higher risk of recurrent restenosis than nondiabetics for nonstented and in-stent restenosis coronary lesions. The use of Ir-192 for preventing recurrent restenosis in such patients remains undefined. The GAMMA I trial was a prospective, randomized, double-blind, multicenter trial of 252 patients with in-stent restenosis who underwent percutaneous coronary intervention and were assigned to receive either Ir-192 (131 patients) or catheter-based placebo (121 patients). DM was present in 79 patients (31%) (41 patients received Ir-192 and 38 patients received placebo) and was absent in 173 patients (90 patients received Ir-192 and 83 patients received placebo). At 6 month follow-up in the GAMMA I trial, the angiographic in-lesion binary restenosis rate was lower in the Ir-192 arm than in the placebo arm (32.4 vs 55.3, p = 0.01). When patients were stratified by the presence of DM, the antirestenosis effect of Ir-192 was larger for diabetic patients than for nondiabetic patients (absolute in-lesion restenosis rate was reduced by 40% for diabetics and 16% for nondiabetics). Thus, adjunctive Ir-192 intracoronary radiation therapy reduces recurrent restenosis after intervention for in-stent restenosis in patients with and without DM. The relative impact of this treatment is more pronounced in diabetic patients because it appears to neutralize the added risk of recurrent restenosis seen in proliferative diabetic lesions. PMID- 12127612 TI - Atypical presentations among Medicare beneficiaries with unstable angina pectoris. AB - Chest pain is a hallmark symptom in patients with unstable angina pectoris (UAP). However, little is known regarding the prevalence of an atypical presentation among these patients and its relation to subsequent care. We examined the medical records of 4,167 randomly sampled Medicare patients hospitalized with unstable angina at 22 Alabama hospitals between 1993 and 1999. We defined typical presentation as (1) chest pain located substernally in the left or right chest, or (2) chest pain characterized as squeezing, tightness, aching, crushing, arm discomfort, dullness, fullness, heaviness, pressure, or pain aggravated by exercise or relieved with rest or nitroglycerin. Atypical presentation was defined as confirmed UAP without typical presentation. Among patients with confirmed UAP, 51.7% had atypical presentations. The most frequent symptoms associated with atypical presentation were dyspnea (69.4%), nausea (37.7%), diaphoresis (25.2%), syncope (10.6%), or pain in the arms (11.5%), epigastrium (8.1%), shoulder (7.4%), or neck (5.9%). Independent predictors of atypical presentation for patients with UAP were older age (odds ratio 1.09, 95% confidence interval 1.01 to 1.17/decade), history of dementia (odds ratio 1.49, 95% confidence interval 1.10 to 2.03), and absence of prior myocardial infarction, hypercholesterolemia, or family history of heart disease. Patients with atypical presentation received aspirin, heparin, and beta-blocker therapy less aggressively, but there was no difference in mortality. Thus, over half of Medicare patients with confirmed UAP had "atypical" presentations. National educational initiatives may need to redefine the classic presentation of UAP to include atypical presentations to ensure appropriate quality of care. PMID- 12127613 TI - Impact of age and sex on plasma natriuretic peptide levels in healthy adults. AB - Assays for natriuretic peptides have received considerable attention as potential screening tests for congestive heart failure and left ventricular dysfunction. However, information regarding the impact of age, sex, and other physiologic characteristics on natriuretic peptide levels is limited. We examined a healthy reference sample of 911 subjects (mean age 55 years, 62% women) from the Framingham Heart Study who were free of hypertension, valvular disease, diabetes, atrial fibrillation, obesity, coronary heart disease, congestive heart failure, and renal failure, and who had normal left ventricular systolic function. Plasma brain natriuretic peptide and N-terminal atrial natriuretic peptide levels were measured, and multivariable regression used to assess correlates of natriuretic peptide levels. The strongest predictors of higher natriuretic peptide levels were older age and female sex. Other multivariable predictors included lower diastolic blood pressure (higher pulse pressure), lower body mass index, and higher left atrial size. Reference limits were then formulated based on the empirical distribution of natriuretic peptide levels by gender both across all ages and partitioned by age. Age-pooled reference limits compared with age specific limits classified a higher proportion of healthy elderly subjects (17% vs 2.5%), but a lower proportion of healthy young subjects (1% vs 2.5%) as "abnormal." We conclude that interpretation of natriuretic peptide levels should take into consideration gender and possibly age. The reference limits derived from this large, healthy community-based sample will aid in the identification of elevated natriuretic peptide levels in clinical practice. PMID- 12127614 TI - "Interventional lipidology": tomographic plaque imaging and aggressive treatment of metabolic disorders. PMID- 12127615 TI - Dean Ornish, MD: a conversation with the editor. Interview by William Clifford Roberts, MD. PMID- 12127616 TI - Effects of statins on six-month survival and clinical restenosis frequency after coronary stent deployment. PMID- 12127617 TI - Validation of the thrombolysis in myocardial infarction (TIMI) risk score for unstable angina pectoris and non-ST-elevation myocardial infarction in the TIMI III registry. PMID- 12127618 TI - Mechanism of mitral regurgitation in inferior wall acute myocardial infarction. PMID- 12127619 TI - Relation of platelet activation and myocardial ischemia biomarkers dependent on type of chest pain (abrupt onset versus intermittent) in patients with angina pectoris or non-Q-wave acute myocardial infarction. PMID- 12127620 TI - Effect of loading with clopidogrel at the time of coronary stenting on platelet aggregation and glycoprotein IIb/IIIa expression and platelet-leukocyte aggregate formation. PMID- 12127621 TI - Frequency of incomplete reperfusion in patients with acute myocardial infarction undergoing primary angioplasty. PMID- 12127622 TI - Selecting who qualifies for optimal balloon angioplasty versus elective stenting in coronary artery disease (a subanalysis of the DESTINI trial). PMID- 12127623 TI - Usefulness of low-dose dobutamine echocardiography for selection of patients with severely depressed left ventricular function for coronary bypass grafting. PMID- 12127624 TI - Effect of cocaine use on coronary calcium among black adults in Baltimore, Maryland. PMID- 12127625 TI - Internal cardioversion of atrial fibrillation under transesophageal echocardiography guidance without fluoroscopy using single-lead catheter technique. PMID- 12127626 TI - Successful cardioversion of atrial fibrillation using 360-Joules biphasic shock. PMID- 12127627 TI - Relation between sinus rates preceding and following ectopic beats occurring in isolation and as episodes of bigeminy in young healthy subjects. PMID- 12127628 TI - QRS duration variability in patients with heart failure. PMID- 12127629 TI - Prevalence of and risk factors for atrial fibrillation and intra-atrial reentrant tachycardia among patients with congenital heart disease. PMID- 12127630 TI - A novel nonfluoroscopic catheter visualization system (LocaLisa) to reduce radiation exposure during catheter ablation of supraventricular tachycardias. PMID- 12127631 TI - Effect of repeated sauna therapy on survival in TO-2 cardiomyopathic hamsters with heart failure. PMID- 12127632 TI - Inaccurate normal values of heart rate variability spectral analysis in newborn infants. PMID- 12127633 TI - Blockpnea and silent myocardial ischemia. PMID- 12127635 TI - Perpetuating negative attitudes about the intrauterine device: textbooks lag behind the evidence. AB - This study examined current textbooks and manuals to discern whether their presentations of the Copper T380A intrauterine device are accurate, current, and objective. Thirteen medical student obstetrics and gynecology texts used in the US and five from the UK were evaluated for information presented about the Copper T-380A intrauterine device. A scoring system was developed to record the presence or absence of characteristics of intrauterine device usage. The two authors independently reviewed the texts and completed code sheets. In general, advantages of the device were under-reported while disadvantages were exaggerated. Review of US texts identified inaccuracies regarding the device's mechanism of action; 5 of 13 did not include its prefertilization action. Despite evidence to the contrary, 9 of 13 texts reported an increased risk of pelvic inflammatory disease associated with intrauterine device use; 6 of 13 reported an increased risk of ectopic pregnancy; and 4 of 13 reported an increased risk of infertility. Review of UK texts yielded similar results regarding advantages and disadvantages, as well as mechanism of action. The UK texts presented fewer inaccuracies regarding intrauterine device risks. Most texts from both countries implied that the intrauterine device is a method of last resort. Texts commonly used by medical students on women's health rotations may not be evidence-based in the information presented about the intrauterine device. PMID- 12127634 TI - Current opinion: consensus statement on intrauterine contraception. AB - Forty-five experts from around the world attended a 1-day seminar in September 2001 in Chapel Hill, North Carolina, USA, to identify ways that they might collaborate to overcome unnecessary barriers to the use of intrauterine devices (IUDs). Seminar participants formed working groups that produced at least three specific recommendations relating to: training/performance improvement; service delivery improvement; general public information; and clinical and programmatic research. Key recommendations included: integrating reproductive health knowledge and skills into curricula for all healthcare professionals; reviewing and reinforcing with providers evidence-based guidelines for IUD use; encouraging evidence-based review of the IUD label and package insert; and conducting further research about IUD client eligibility, potential health benefits, acceptability among clients and providers, and use by HIV-infected women. At the meeting's conclusion, a number of participants, representing the fields of research, policy, communications, donors, women's advocacy, and medicine, expressed an interest in refining and acting upon the recommendations. Hosted by Family Health International, the meeting was supported by the Mellon Foundation. PMID- 12127636 TI - Using a low-dose contraceptive in women 35 years of age and over: 20 microg estradiol/100 microg levonorgestrel. AB - The efficacy, safety, and cycle control of a low-dose oral contraceptive (OC) containing 20 microg ethinyl estradiol (EE) and 100 microg levonorgestrel (LNG) has been demonstrated in a large trial with 1708 women (>or=15 years old with regular menstrual cycles). The objective of this study was to analyze the same parameters in 218 of the 1708 women who were 35 years of age and older. Women were administered the 28-day, combination OC (20 microg EE/100 microg LNG; 21 days active medication/7 days placebo) for up to 3 years. During 3859 cycles evaluated for efficacy, one pregnancy occurred (Pearl index 0.34). The most common adverse events cited as reasons for discontinuation were hypertension (3% of subjects), headache (2%), and metrorrhagia (2%). One subject withdrew as a result of a serious adverse event. Breakthrough bleeding, spotting, and bleeding and spotting occurred in 2.9%, 11.0%, and 6.8%, respectively, of the 3739 cycles evaluated for cycle control. This low-dose, monophasic regimen of 20 microg EE/100 microg LNG is an effective, safe, and tolerable contraceptive and provides good cycle control for women 35 years of age and older. PMID- 12127637 TI - Attitudes and experiences with levonorgestrel 100 microg/ethinyl estradiol 20 microg among women during a 3-month trial. AB - To describe attitudes and experiences with a low-dose oral contraceptive pill (Alesse) over 3 months, women aged 18 years and older (n = 218) were enrolled from 16 locations to evaluate their experiences with Alesse. The questionnaire assessed demographic and personal characteristics, attitudes and experiences, and satisfaction. The participants had a mean age of 26.7 years and most were single, Caucasian, had completed high school, had a regular sexual partner, and had previously used OCs. Sixty percent of participants could discuss pill use easily with their mothers, 92% with friends, and 96% with partners; 45% of the women were unsure about their mother's previous OC use. Of the 11 side effects assessed, the most frequently anticipated side effect was weight gain. There was a significant relationship between anticipated and reported side effects for weight and mood changes; however, there remained a number of women for whom these differed. Most (90%) were satisfied with Alesse. Even when beginning on 20 microg pills, some women may still anticipate side effects such as weight gain typically associated with higher doses of estrogen. Healthcare providers should assess women's attitudes and anticipated experiences with OCs and counsel accordingly. PMID- 12127638 TI - Bone mineral density at various anatomic bone sites in women receiving combined oral contraceptives and depot-medroxyprogesterone acetate for contraception. AB - The association between users of combined oral contraceptives and depot medroxyprogesterone acetate (DMPA) for contraception and bone mineral density (BMD) has been controversial because of variations among studies. Like other studies, this cross-sectional study compares BMD in users of combined oral contraceptives and DMPA with that in nonusers. Unlike previous studies, we defined long-term use as >2 years, and we measured more bone sites than previous studies including lumbar spines, femurs, and forearms. The study group consisted of 59 women aged 30 years to 34 years who had been using combined oral contraceptives for 57.36 +/- 27.02 months with a minimum period of 24 months, 34 women of the same age who had been using DMPA as contraceptive for 55.76 +/- 35.31 months, and 62 women of the same age who had not used any steroid hormonal contraceptives for more than 6 months. BMD was measured by dual energy photon absorptiometer at lumbar spine 2-4, neck of femur, Ward's triangle of femur, greater trochanter of femur, ultradistal radius, and distal ulnar, respectively. Age, body mass index, and lifestyles of both groups were matched with nonusers. Mean BMD at lumbar spine (L2-4) in the DMPA users was significantly lower than in the controls (1.031 +/- 0.090 vs. 1.096 +/- 0.116, p = 0.007). There were no significant differences in BMD values at bone sites other than lumbar spine between DMPA users and the controls. There were no significant differences in BMD values at all bone sites between combined oral contraceptives users and the controls. We conclude that combined oral contraceptives are not associated with changes in values of BMD, while DMPA is associated with decreased BMD only at lumbar spine. We comment that steroid hormonal contraceptives are safe to use for long-term contraception regarding bone mass effects. PMID- 12127639 TI - Second trimester abortion by laminaria followed by vaginal misoprostol or intrauterine prostaglandin F2alpha: a randomized trial. AB - Intra-amniotic injection, as well as intravaginal application of prostaglandins, have been used to terminate second trimester pregnancies. There is as yet no consensus as to the most efficient protocol of such late abortions. Our goal was to compare the efficacy of intra-amniotic injection of prostaglandin F2 alpha (PGF2alpha) and intravaginal application of misoprostol in terminating second trimester pregnancies after pretreatment with intracervical laminaria. Women with live fetuses and requesting second trimester abortions were randomized into two groups. Eighteen hours following the insertion of intracervical laminaria, women were treated with either intra-amniotic injection of 40 mg PGF2alpha, or 12 hourly doses (to a maximum of 4 doses) of 200 mcg misoprostol. Fifty women were randomly assigned to each group. Failure to abort within 24 h of initiation of treatment occurred in 6 patients (12%) in the misoprostol group and 14 (28%) of the PGF2alpha group (p = 0.04). Mean time of induction of pharmacologic treatment to abortion was 13.6 h in the misoprostol group and 10.7 h in the PGF2alpha group (p = 0.03). The mean number of analgesic injections given were 0.8 in the misoprostol group and 1.6 in the PGF2alpha group (p = 0.0001). Only the method of abortion was predictive of abortion success and not other variables such as patient age, gestational age, gravidity, or parity. Following intracervical laminaria, vaginal misoprostol has been found to be more effective and less painful, compared with intra-amniotic PGF2alpha, for the termination of second trimester pregnancies with live fetuses. PMID- 12127640 TI - Buccal misoprostol as cervical preparation for second trimester pregnancy termination. AB - The objective of the study was to assess the efficacy of cervical preparation with misoprostol or laminaria suction abortion up to 18 weeks gestation. The study was conducted among a consecutive case series of patients presenting for dilatation and evacuation at a single center. Cervical preparation was effected by either 600 mg buccal misoprostol (n = 32) or laminaria (n = 78). Dilatation and evacuation was then performed using vacuum aspiration, with fetal parts or placenta removed as necessary with appropriate forceps. The principle outcome of interest was procedure time. Complete abortion was effected in 100% of the cases. In all cases, the cervix was adequately prepared to allow either the introduction of a size 14-mm suction cannula or was easily dilated to this diameter. Group size was sufficient for detection of a 20% difference in mean procedure time with 80% power (calculated with two separate standard deviations for procedure time). Procedure times were not significantly different when misoprostol was used compared to laminaria for cervical preparation. It is concluded that the use of misoprostol to provide cervical preparation in second trimester abortion procedures appeared effective and offers an alternative to laminaria. Buccal misoprostol should be more widely investigated for this purpose. PMID- 12127641 TI - Users' perspectives on medical abortion in Finland. AB - The objective of this study was to determine the reasons for choosing the medical instead of surgical method for the termination of early pregnancy, and to evaluate the experience and the level of satisfaction with the medical method among women in Helsinki who participated in a multinational efficacy study of medical abortion. The study respondents consisted of 123 women with less than 9 weeks gestation, undergoing medical abortion with mifepristone 200 mg orally followed 36 h to 48 h later by misoprostol 0.8 mg orally or vaginally. By using structured questionnaires, the reasons for choosing the medical method were assessed before the abortion, whereas the experience of medical abortion was evaluated at follow-up visits 2 weeks and 6 weeks later. The three most important reasons for choosing medical abortion instead of surgical abortion were 'avoidance of surgery,' 'avoidance of general anesthesia,' and 'the method being more natural.' At the 6-week evaluation, most women (92%) were satisfied or highly satisfied with the method, and 103 (88%) women said that the method met their expectations. The majority of all women, 105 (89%), would opt for the same method in the future should the need arise, and 102 (88%) patients would recommend the medical method to a friend. The length of pregnancy had an effect on acceptability; the shorter the gestational age, the more acceptable the medical abortion. Medical abortion was perceived as an acceptable method for termination of early pregnancy among the Finnish women studied. PMID- 12127642 TI - Oral contraceptives and risk of gestational trophoblastic disease. AB - Clinical reports suggested that the use of oral contraceptives (OC) after a molar pregnancy may increase the risk of persistent throphoblastic disease. However, few epidemiologic studies have analyzed the effect of OC use on the risk of developing gestational trophoblastic disease (GTD). To give further information, we have analyzed data from a case-control study on risk factors for GTD. Cases were 268 women with a histologically confirmed diagnosis of complete or partial mole referred to the participating Trophoblastic Disease Centers. A total of 268 subjects were interviewed; 79 cases were classified as partial and 159 as complete mole. Controls were randomly selected women who gave birth to healthy infants at term on randomly selected days in the same network of hospitals in which cases had been identified. A total of 104 cases and 130 controls reported ever OC use, and the corresponding odds ratio (OR) was 1.5 (95% CI, 1.1-2.1). The risk of GTD increases with duration of OC use: the OR was 1.7 (95% CI 1.2-2.6) for ever-users reporting >or=12 months of use. No consistent pattern of risk was observed with time since last OC use. We have analyzed separately the association between OC use and risk of complete and partial moles: no statistically significant difference emerged, but the OR for partial moles was higher (OR 3.0, 95% CI 1.6-8.4) than for complete mole (OR 1.0, 95% CI 1.8). In conclusion, we observed a weak association between OC use and GTD; such a weak association could be explained by factors other than causality. PMID- 12127643 TI - Inconsistent use of oral contraceptives in rural Bangladesh. AB - The purpose of this study is to explore predictors of inconsistent use of oral contraceptives (OCs) in rural Bangladesh. A total of 801 rural OC users were included in the study, about half of them (49%) missed one or more active pill(s) during the 6 months before the survey.Multivariate analysis revealed that Muslim women were 60% more likely to be inconsistent OC users compared to their non Muslim counterparts. Women who lacked knowledge about contraindications were 60% more likely to take the pill inconsistently than were women who had the knowledge. Women who were not visited by family planning workers or did not have access to mass media were 40% more likely to be inconsistent OC users.OC users need increased information about correct OC use, which could be provided via improved access to mass media with specific messages on how to use OCs properly. Better access to the community clinics could improve the pill-taking behaviors of rural Bangladeshi women. PMID- 12127644 TI - Effects of chelating agents on the rheological property of cervical mucus. AB - As an ongoing effort to elucidate the mechanisms involved in the calcium dependent fertility regulation process, the viscoelastic properties of the mucus obtained from lamb cervix and human semen, as well as their water and total protein contents after exposure to EDTA, a chelating agent, or Nonoxynol-9 (N-9), a spermicidal agent, were examined. The viscosity was measured using a Cone Plate Digital Viscometer, while the water and total protein contents were determined by the lyophilization process and the Lowry method, respectively. The significant changes in the rheological properties of mucus, such as its viscosity and the water content, upon exposure to EDTA were demonstrated. The viscosity of cervical mucus and human semen were significantly increased by EDTA treatment (as compared to the controls): lamb cervical mucus (2.9 +/- 0.3 vs. 2.2 +/- 0.3 cps) and human semen (5.0 +/- 0.3 vs. 4.3 +/- 0.3 cps), respectively. The hydration rate was decreased by EDTA treatment as compared with the control (93.6 +/- 0.7 vs. 96.8 +/- 0.8%). Among tested samples, the reduction in the percentage of sperm penetration through the cervical mucus was the highest in the mucus containing EDTA, which had the lowest water content (93.6 +/- 0.7%), indicating that there is a positive relationship between the hydration rate of the cervical mucus and its ability to permit the penetration of spermatozoa. This result indicates that spermicidal activity exerted by high concentrations of EDTA is in part due to its effect on the rheological properties of cervical mucus or semen. PMID- 12127645 TI - Inhibition of Ca(2+) channels in mouse spermatogenic cells by male antifertility compounds from Tripterygium wilfordii Hook. f. AB - The male antifertility effect of a water-chloroform extract (GTW) from the root xylem of Tripterygium wilfordii has attracted worldwide interest. In the present study, by using whole-cell recording, the effects of GTW and two isolated monomers from GTW, demethylzeylasteral and L-epicatechin, on the T-type Ca(2+) channels in mouse spermatogenic cells were investigated. The results showed that each of them concentration-dependently and partially reversibly inhibited T-type Ca(2+) current in the cells. The IC(50) of GTW and demethylzeylasteral were approximate, while L-epicatechin inhibited the channels at a much higher concentration. The voltage dependence of the inhibitory effect and the changes in activation and inactivation time constants after application of these compounds were also examined. These data suggest that the inhibition of T-type Ca(2+) currents could be responsible for the antifertility activity of these compounds. PMID- 12127646 TI - Side effects and oral contraceptive pill (OCP) discontinuation rate in rural Bangladesh. PMID- 12127648 TI - Multiple sclerosis 2001. PMID- 12127649 TI - Mechanisms of demyelination and tissue destruction in multiple sclerosis. PMID- 12127650 TI - Multiple sclerosis: from basic immunopathology to immune intervention. PMID- 12127651 TI - Compartmentalization of the immune response in the central nervous system and natural history of multiple sclerosis. Implications for therapy. PMID- 12127652 TI - The worldwide prevalence of multiple sclerosis. PMID- 12127653 TI - Epidemiology of multiple sclerosis in Croatia. PMID- 12127654 TI - The genetics of multiple sclerosis. PMID- 12127655 TI - The natural history of early onset multiple sclerosis: comparison of data from Moscow and Vancouver. PMID- 12127656 TI - The natural history of untreated multiple sclerosis in Iceland. A total population-based 50 year prospective study. PMID- 12127657 TI - The differential diagnosis of multiple sclerosis. PMID- 12127658 TI - Changes of attention and memory in a group of patients with multiple sclerosis. PMID- 12127659 TI - Using cognitive performance to investigate neurodegenerative disease. PMID- 12127660 TI - The symptomatic treatment of multiple sclerosis. PMID- 12127661 TI - MRI monitoring of MS in clinical trials. PMID- 12127662 TI - Long-term benefits of early and high doses of interferon beta-1a treatment in relapsing-remitting multiple sclerosis. PMID- 12127663 TI - Immunomodulatory treatment of MS in Slovenia. PMID- 12127664 TI - Interferon beta treatment in relapsing-remitting multiple sclerosis. A review. PMID- 12127665 TI - Multiple sclerosis and interferon beta-1b, past, present and future. PMID- 12127666 TI - Newer long-term treatments for multiple sclerosis. PMID- 12127667 TI - Quality of life and cost of multiple sclerosis. PMID- 12127669 TI - The nature of the autonomic dysfunction in multiple system atrophy. AB - The concept that multiple system atrophy (MSA, Shy-Drager syndrome) is a disorder of the autonomic nervous system is several decades old. While there has been renewed interest in the movement disorder associated with MSA, two recent consensus statements confirm the centrality of the autonomic disorder to the diagnosis. Here, we reexamine the autonomic pathophysiology in MSA. Whereas MSA is often thought of as "autonomic failure", new evidence indicates substantial persistence of functioning sympathetic and parasympathetic nerves even in clinically advanced disease. These findings help explain some of the previously poorly understood features of MSA. Recognition that MSA entails persistent, constitutive autonomic tone requires a significant revision of our concepts of its diagnosis and therapy. We will review recent evidence bearing on autonomic tone in MSA and discuss their therapeutic implications, particularly in terms of the possible development of a bionic baroreflex for better control of blood pressure. PMID- 12127670 TI - N-methyl-D-aspartate receptor subunit NR2A and NR2B messenger RNA levels are altered in the hippocampus and entorhinal cortex in Alzheimer's disease. AB - The N-methyl-D-aspartate (NMDA) receptor is a subtype of ionotropic glutamate receptor that is involved in synaptic mechanisms of learning and memory, and mediates excitotoxic neuronal injury. In this study, we tested the hypothesis that NMDA receptor subunit gene expression is altered in Alzheimer's disease (AD), especially in brain regions known to be important in memory. Quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) was used to determine the messenger RNA (mRNA) levels of the NMDA receptor subunits NR1, NR2A, and NR2B in the hippocampus and entorhinal cortex of postmortem brain samples from nine clinically well-characterized AD patients and nine aged controls. Cerebellum, a site minimally affected by AD, was also chosen for comparison assessment. Results showed decreased levels of the NR2 mRNAs in AD brains compared to controls. Reductions of NR2A (46.2%, p<0.01) and NR2B (43.2%, p<0.0001) mRNA levels were identified in the entorhinal cortex. Reductions of NR2A (41.4%, p<0.05) and NR2B (40.6%, p=0.058) mRNA levels were found in the hippocampus. NR1 mRNA levels were similar in all three brain regions in both AD and controls. No significant changes of subunit NR2A and NR2B mRNA levels were identified in the cerebellum. Postmortem delay (PMD), tissue storage time, brain weight, or age of the subjects did not affect these changes. These data suggest that alterations in NMDA receptor subunits, especially the NR2A and NR2B, may be important in AD, particularly in neuronal populations that underlie impaired learning and memory. PMID- 12127671 TI - Intranuclear inclusions, neuronal loss and CAG mosaicism in two patients with Machado-Joseph disease. AB - The presence of neuronal intranuclear inclusions (NIIs) and neuronal mosaicism has been described in some autosomal dominant spinocerebellar ataxias (SCA), but their implication in neurodegenerative mechanisms still remains unclear. OBJECTIVE: To investigate the correlation between neuronal loss and NIIs, and the size of CAG triplet expansion in selected areas of the CNS in two SCA3 patients. MATERIAL AND METHODS: Postmortem neuropathological study was carried out, and the regional distribution of neuronal loss was compared with NIIs. CAG expansion was analysed by PCR amplification in the same regions. RESULTS: Marked neuronal loss was seen in the anterior horn of the spinal cord, pontine nuclei and motor nuclei of the brain stem. Moderate neurone loss was found in the locus ceruleus, colliculus and substantia nigra. Loss of granule and Purkinje cells was found in the cerebellum, mainly in the vermis. NIIs were present in neurones of the involved nuclei of the anterior horn of the spinal cord, medulla oblongata and pons, but not in the locus ceruleus, substantia nigra and cerebellum. A few NIIs were found in the striatum. The number of CAG repeats was 27/70 in the first patient and 21/74 in the second patient. The variation of the expanded allele size among different cerebral areas was +/-1-3 CAG repeats. CONCLUSION: The partial correlation between neuronal loss and NIIs suggests that other factors distinct from NII formation may be involved in the neuronal death. Moreover, the low degree of mosaicism between regions without neuronal loss and regions with marked neuronal loss points to the existence of selective cellular vulnerability to the genetic defect. PMID- 12127672 TI - Differences in cerebral metabolic impairment between early and late onset types of Alzheimer's disease. AB - The purpose of this study was to delineate the specific patterns of cerebral glucose metabolism with regard to the time of onset of Alzheimer's disease (AD). METHODS: Two groups of 20 AD patients with different ages of onset were examined. The early onset (EO) and late onset (LO) groups had mean ages of onset of 53.9 and 72.7 years. Groups of age-matched normal subjects were used as controls. A regional relative cerebral glucose metabolic image of each subject was obtained by 2-[18F] fluoro-2-deoxy-D-glucose (FDG) positron emission tomography (PET). NEUROSTAT program was used for spatial normalization and voxel-based statistical parametric mapping (SPM) 99 was used for statistical analyses. RESULTS: Both AD groups had significant hypometabolic regions in the bilateral parieto-temporal regions compared with the age-matched groups. The EO group had more severe hypometabolism in the bilateral parietal and posterior cingulate cortices and precuneus region than the LO group. However, LO group showed no significant hypometabolic regions compared to the EO group. CONCLUSION: The effects of time of AD onset were delineated as a double dissociation, that is, EO AD patients have a more severe reduction of glucose metabolism. Our finding suggests the existence of biological subtypes of AD. PMID- 12127673 TI - Treatment of spinal cord impact injury in the rat with transforming growth factor beta. AB - To investigate the contribution of cytokines, proinflammatory TNF-alpha and inhibitory TGF-beta, to spinal cord injury (SCI) in a rat model, two studies were performed using adult Sprague-Dawley rats which were injured at T9/T10. In the first study, rats were sacrificed at 1, 6, 24, 96 and 168 h after SCI for immunocytochemistry of coronal sections for the presence of mononuclear phagocytes, astrocytes, TNF-alpha and TGF-beta, among other markers. From intervening frozen sections, RNA was extracted for semiquantitative polymerase chain reaction (RT-PCR) analysis of TNF-alpha and TGF-beta. In the second experiment, rats were treated with intravenous TGF-beta 30 min after injury and sacrificed at 6 and 48 h after injury. Spinal cord sections were immunocytochemically stained and RNA extracted for semiquantitative PCR as mentioned above, as well as quantitation of lesion volume. There were increases in mononuclear phagocytes and astrocytes, as early as 1 h after SCI, with steady progression over 168 h after injury. TNF-alpha and TGF-beta was produced locally by mononuclear phagocytes and astrocytes. There was an 18-h delay in peak mRNA production of TGF-beta compared to TNF-alpha. The treatment of SCI rats with TGF beta reduced lesion volume by 50% at 48 h and this was associated with decreased accumulation of mononuclear phagocytes in and around the injury site. This reduction of mononuclear phagocyte numbers around the site of trauma would reduce their contribution to secondary injury. PMID- 12127674 TI - Gelatinase B, PECAM-1 and MCP-3 gene polymorphisms in Belgian multiple sclerosis. AB - Polymorphic microsatellite markers in the genes for gelatinase B, PECAM-1 and MCP 3 have previously been analysed in Swedish and Sardinian individuals to test for association with multiple sclerosis (MS). Confirmation and comparison of genetic associations in various ethnic populations is mandatory and, therefore, we studied these three gene polymorphisms in 216 clinically definite MS patients and 193 normal controls, and in 148 simplex MS families, all of Belgian origin. No allelic associations were found between MS and the CA microsatellite marker in the promoter region of the gelatinase B gene, and the polymorphic CA repeat in the sixth intron of PECAM1. However, the two most abundant alleles of the CA/GA microsatellite polymorphism in the promoter-enhancer region of the MCP-3 gene, A2 (109 bp) and A3 (111 bp), were found to be significantly associated with disease in the case-control study [OR (95% CI)=0.68 (0.51-0.92), p (1 df)=0.015 and OR (95% CI)=1.62 (1.22-2.14), p (1 df)=0.0010, respectively], but not in the family study. These results are in agreement with previous findings in the Swedish and Sardinian populations and reinforce the possibility of a role for chemokines in MS pathogenesis. PMID- 12127675 TI - Home management of febrile convulsion in an African population: a comparison of urban and rural mothers' knowledge attitude and practice. AB - To determine the knowledge, attitude and practice (KAP) of home management of febrile convulsion (FC), by mothers in the community, focus group discussions (FGD) were conducted in two communities, Uselu (urban) and Evbuomodu village (rural), both in Edo State, Southern Nigeria. The study was conducted between December 2000 and February 2001. Our findings show that 71% of urban mothers compared to 25% of rural mothers attributed the cause of FC to fever (chi(2)=24.17: p<0.001). Seventy-five percent of mothers from rural community and 28.6% of urban mothers attributed the cause to witchcraft and/or evil spirits. Twenty-five percent of rural mothers also attributed abnormality of the spleen as a cause of FC. All the mothers, both urban and rural, were not directly involved in the management of the convulsive episode due to panic and confusion. Ninety two percent of urban and all the rural mothers permitted the use of traditional medicine while 7.1% of urban mothers employed prayers during convulsion. Twenty percent of urban and twenty-two percent of rural mothers use urine (human and or cow's) for treating FC at home. Other home remedies include kerosene, fuel and crude oil. Mass enlightenment campaign for the community, especially the rural, against use of harmful traditional remedies to treat FC at home is strongly advised. PMID- 12127676 TI - Serial analysis of cytokine mRNA profiles in whole blood samples from patients with early multiple sclerosis. AB - In this pilot study, we serially determined the cytokine messenger RNA (mRNA) expression pattern in whole blood samples from 12 patients with clinical isolated syndrome suggestive of early multiple sclerosis (MS) using a new sensitive quantitative polymerase chain reaction (PCR) method. Significantly higher levels of tumor necrosis factor-alpha (TNF-alpha; x 5.1), interferon-gamma (IFN-gamma; x 4.8) and interleukin-10 (IL-10; x 5.6) mRNA were detected in MS patients at the time of a relapse compared to healthy controls. Treatment with i.v. methylprednisolone (MP) led to an increase of IL-4 mRNA and a significant decrease of IFN-gamma and TNF-alpha mRNA expression. In this cohort of clinically stable patients, proinflammatory cytokines remained low during the 1-year follow up period. As several indications point to a cytokine dysregulation in MS, quantitative analysis of cytokine mRNA profiles in whole blood samples by real time PCR may be a useful immunological marker to monitor disease activity in future therapeutic trials in MS. PMID- 12127677 TI - Chorea associated with non-ketotic hyperglycemia and hyperintensity basal ganglia lesion on T1-weighted brain MRI study: a meta-analysis of 53 cases including four present cases. AB - BACKGROUND: Chorea associated with non-ketotic hyperglycemia and high signal intensity lesions on T1-weighted brain magnetic resonance images (C-H-BG) is recognized as a unique syndrome that affects elderly women exclusively. However, its overall clinical features are unclear. MATERIAL AND METHODS: The literature describing patients with C-H-BG from 1985 to 2001 was reviewed using MEDLINE. Their clinical features and those of four patients with C-H-BG at this hospital were analyzed. RESULTS: This study included 49 patients from the literature and four patients at this hospital. Their mean age at the onset was 71.1 years (range=22-92 years). Women were affected more frequently than men (men/women=17:30). The mean serum glucose level measured after the onset of chorea was 481.5 mg/dl (ranging from 169 to 1264), HbA1c level was 14.4% (ranging from 9.9 to 19.2), and the serum osmolarity was 305.9 mmol/kg (ranging from 291 to 335). Forty-seven patients developed hemichorea. Six patients developed bilateral chorea, and magnetic resonance imaging (MRI) showed bilateral basal ganglia lesions. MRI showed that putamen was involved in all cases (isolated putamen=31 patients, additional basal ganglia lesions=22 patients). None had lesions confined to the caudate nucleus or the globus pallidus. In all, except one, the anterior limb of the internal capsule was spared. Follow-up MRI studies were performed in 22 patients. In most, hemichorea improved along with the disappearance of the lesions. In 39 patients, chorea had ameliorated completely. The remaining 14 cases showed some improvement during the follow-up period. The chorea recurred in seven patients. CONCLUSION: C-H-BG is a benign disorder affecting the elderly. It affects men much more frequently than has been reported. The high signal intensity basal ganglia lesion on the T1-weighted brain MRI study was reversible, and correlated with the clinical improvement in chorea. PMID- 12127678 TI - Neurological features of West Nile virus infection during the 2000 outbreak in a regional hospital in Israel. AB - During the summer of 2000, 35 patients with West Nile Virus Fever were admitted to our hospital. Of these, the 26 (21 adults, mean age 56 (19-86) and 5 children (aged 9-15)) presented have neurological involvement, 33% with meningitis, 52% with meningoencephalitis, 10% with encephalitis and 5% with acute polyneuropathy. Presenting clinical features were fever in 95% of cases, headache in 90%, nausea/vomiting in 52%, confusion in 48%, somnolence in 38%, neck stiffness in 33%, a skin rash in 19%, diarrhea in 14%, cervical pain in 14%, seizure in 9%, photophobia in 9% and limb weakness in 4%. Leucopenia was not found. Two patients diagnosed with meningoencephalitis died. Three patients had signs of an acute polyneuropathy, this being the only complaint of one patient. The EEG was abnormal in all cases of meningitis or meningoencephalitis, except in three cases. Outbreaks of West Nile Virus Fever are emerging as a worldwide disease with high rates of neurological involvement and death. It should be considered in cases presenting with aseptic meningoencephalitis, meningitis and acute polyneuropathy, especially during the summer months and in areas along bird migration pathways. PMID- 12127679 TI - Alteration of midkine expression in the ischemic brain of humans. AB - Midkine (MK) is a heparin-binding growth factor that occurs as a product of the retinoic acid-inducible gene. Alteration of MK expression in ischemic brain lesions was examined in humans immunohistochemically in nine patients and in two control subjects without neurological disorders. Some neurons were MK immunopositive, but no evident MK-immunoreactivity was observed in astrocytes in brains of control subjects. In the ischemic lesions, significant elevation of MK immunoreactivity in the astrocytes and depletion of the reactivity in neurons were seen, especially in the early period, where edema and eosinophilic neurons were prominent. On the other hand, MK-immunoreactivity was not observed in hypertrophic and fibrillary astrocytes in the later period. These findings suggest that the MK in astrocytes play some role in the repair process in the early period of the ischemic brain lesions in humans. PMID- 12127680 TI - Neurologic services in sub-Saharan Africa: a case study among Zambian primary healthcare workers. AB - INTRODUCTION: In many parts of the developing world, access to physician consultation and neurologic expertise is limited or nonexistent. We conducted a survey among non-physician, primary healthcare workers (PHCWs) to determine the neurological needs and services in rural Zambia. METHODS: Semi-structured written questionnaire utilizing fill-in-the-blank, multiple-choice likert-scaled questions, and open-ended questions. RESULTS: Seizures were reported as the most common neurologic disorder by 66% of the PHCWs. Only 1/3 of PHCWs reported feeling adequately trained to care for seizures and seizure disorders. PHCWs reported even less expertise for other neurologic conditions. Over 40% of PHCWs surveyed work in primary care clinics without a physician available for consultation. Their patients must travel a median of 50 km to access a physician and geographic barriers are a frequent problem. In addition to difficulty physically accessing care, PHCWs reported that financial barriers to physician referral are substantial. Expenses cited include additional user fees for physician-level care, transportation costs, and the cost of maintaining the patient and/or family at a site distant from the home village. Traditional beliefs, social stigma, and discriminatory healthcare policies associated with neurologic conditions were also noted to deter and defer care and care seeking. CONCLUSIONS: PHCWs lack sufficient training and experience to care for the neurologic disorders in their patient populations, although such disorders are relatively common. Geographic, financial and cultural barriers substantially limit physician referrals. To assure at least a minimal quality of care for people with nervous system disorders in Zambia, PHCWs' neurologic education must be increased and barriers to physician referral decreased. PMID- 12127681 TI - Cerebral metabolic changes in early multiple system atrophy: a PET study. AB - Previous positron emission tomography (PET) studies have shown widespread hypometabolism in the brain of advanced MSA but the time course of these metabolic abnormalities is largely unknown. In order to clarify the principal disease processes in multiple system atrophy (MSA) in the early stage, we investigated regional cerebral glucose metabolism (rCMGglc) and nigral dopaminergic function in nine patients with early stage of MSA using [(18)F]fluorodeoxyglucose (FDG) and 6-L-[(18)F]fluorodopa ((18)F-Dopa) positron emission tomography (PET) (two men and seven women; age, 59.3+/-5.4 years; disease duration, 29.7+/-14.6 months). The rCMRglc in the early MSA patients significantly decreased in the cerebellum, brainstem, and striatum compared with that in nine normal subjects. A significant correlation was found between the severity of autonomic dysfunction and rCMRglc within the brainstem. The severity of extrapyramidal signs also correlated with the decline of F-Dopa uptake but not that of rCMRglc within the striatum. The degree of atrophy on MRI has correlated with neither the clinical symptoms nor rCMRglc at the cerebellum and the brainstem. Our PET studies demonstrated widespread metabolic abnormalities except for the cerebral cortex in the brain of MSA even in the early stage. The hypometabolism in the brainstem was tightly linked to the autonomic dysfunction. Not the striatal dysfunction but the nigral damage may be responsible for the extrapyramidal symptoms in early MSA. PMID- 12127682 TI - P301L tauopathy: confocal immunofluorescence study of perinuclear aggregation of the mutated protein. AB - The clinical and neuropathological features in the P301L tauopathy have been described in several kindreds. In this study, we present findings in two previously unreported patients, evaluated both genetically, neuropathologically, and with multiparametric confocal immunofluorescence. The patients were female, with age 65 and 75 years old, respectively. Both exhibited clinical symptoms of frontotemporal dementia (FTD). Marked atrophy of the frontal and temporal lobes with moderate atrophy of the remaining cerebral and brain stem structures was present. The substantia nigra was pale. The atrophic neocortical regions exhibited neuronal loss, marked gliosis, status spongiosus, and occasional ballooned neurons. By light microscopy, the most striking findings were argyrophilic perinuclear rings, frequently with an attached small inclusion (mini Pick-like body), especially prominent in dentate granule cells, entorhinal and temporal cortices, and to a lesser extent in CA1. These structures were immunopositive for tau protein (Tau-2, AT-8, PHF-1, MC-1). Numerous astrocytic plaques, tuft-shaped astrocytes, coiled bodies, and dystrophic neurites were also present. Confocal immunofluorescence with a P301L-specific antibody directly demonstrated the presence of the mutated protein in the PHF-1 positive aggregates. The mutated tau protein (4-repeat tau) was detected in the mini Pick like bodies, indicating an important biochemical difference between these inclusions and classical Pick bodies (3-repeat tau). Additionally, since 4-repeat tau protein is not normally present in dentate granule cells, this result also suggests an abnormality in the mRNA splicing mechanisms. The structural features of the involvement of proteolytic systems in this tauopathy were assessed by immunohistochemistry for the active form of calpain II (C-27) and ubiquitin. Colocalization of PHF-1 positive aggregates with C-27 points to the possible involvement of calpain in tau protein hyperphosphorylation. Absence of immunostaining for ubiquitin indicates possible dysfunction of the ubiquitin proteasome system in this tauopathy. PMID- 12127685 TI - Genetic parameters of neuroblastomas. AB - Neuroblastoma is a malignant childhood tumor of migrating neuroectodermal cells derived from the neural crest and destined for the adrenal medulla and the sympathetic nervous system. The biological behavior of neuroblastomas is extremely variable and in some respects unique. Neuroblastomas tend to regress spontaneously in a portion of infants or to differentiate into a benign ganglioneuroma in some older patients. Unfortunately, in the majority of patients neuroblastoma is metastatic at the time of diagnosis, and it usually undergoes rapid progression with a fatal outcome. The mechanisms leading to this diverse clinical behavior of neuroblastomas are largely unclear. From the analysis of tumors at the cytogenetic and molecular level non-random genetic changes have been identified, including ploidy changes, amplification of the oncogene MYCN, deletions of chromosome 1p, gains of chromosome arm 17q, and deletions of 11q as well as of other genomic regions that allow tumors to be classified into subsets with distinct biological features and clinical behavior. MYCN status is widely accepted for therapy stratification. Additional genetic parameters are currently under investigation to refine risk assessment, but so far the molecular monitoring tools for prediction of therapy response and disease outcome are still incomplete. This should lead to more risk-adapted therapies according to the clinical-genetic parameters by which individual tumors are characterized. This review aims at discussing the role of genomic changes in neuroblastomas of diverse biological and clinical types. PMID- 12127686 TI - Thyroid hormone receptor alpha 1 (c-erb A alpha 1) suppressed transforming phenotype of nasopharyngeal carcinoma cell line. AB - Only three thyroid hormone receptor (TR) isoforms, alpha 1, beta 1, and beta 2, bind thyroid hormone (TH) and are considered to be true TRs. TR alpha 2, unable to bind TH, binds to TH response element on DNA and has been shown to exert dominant negative action on TR alpha1. TR alphas regulate many important processes such as proliferation, differentiation and apoptosis. To find out if TR alphas played roles in growth control of nasopharyngeal carcinoma cells, transfectant with inducible expression of TR alpha 1 was generated from NPC-TW 04 cell lines. Induced expression of TR alpha 1 in nasopharyngeal carcinoma cell reduced proliferation and colony-formation ability in agar. Tumor formation ability in nude mice was reduced in NPC cells with TR alpha 1 expression than those without expression or vector-transfected cells. Our results supported the hypothesis that TR alpha 1 functions as a tumor suppressor gene in nasopharyngeal carcinoma tumorigenesis. PMID- 12127687 TI - N-ethyl-N-hydroxyethylnitrosamine (EHEN)-induced renal and hepatocarcinogenesis in the tumor suppressor Tsc2 transgenic rat. AB - Hereditary renal carcinomas (RCs) develop in Tsc2 gene mutant (Eker) rats around the age of 1 year. We previously reported that Tsc2 mutations were detected in chemically (N-ethyl-N-hydroxyethylnitrosamine (EHEN) and diethylnitrosamine) induced non-Eker rat RCs, suggesting an involvement of Tsc2 alteration in rat RC development. In this study, we evaluated the effect of extra copies of the Tsc2 gene on renal and hepatocarcinogenesis that was induced by EHEN in vivo. The incidence of RCs in non-transgenic rats (2/17) is slightly higher than in transgenic rats (0/32), although it is statistically not significant. These results suggest the presence of other target RC gene(s) in chemically (EHEN) induced renal carcinogenesis. We observed no difference in the numbers and areas of the hepatic glutathione S-transferase placental type positive foci. PMID- 12127688 TI - Intravenous anesthetic, propofol inhibits invasion of cancer cells. AB - Intravenous anesthetic, propofol (2,6-diisopropylphenol), is extensively used for general anesthesia without knowing the effects on cancer. We found here that clinically relevant concentrations of propofol (1-5 microg/ml) decreased the invasion ability of human cancer cells (HeLa, HT1080, HOS and RPMI-7951). In the HeLa cells treated with propofol, formation of actin stress fibers as well as focal adhesion were inhibited, and propofol had little effect on the invasion ability of the HeLa cells with active Rho A (Val(14)-Rho A). In addition, continuous infusion of propofol inhibited pulmonary metastasis of murine osteosarcoma (LM 8) cells in mice. These results suggest that propofol inhibits the invasion ability of cancer cells by modulating Rho A and this agent might be an ideal anesthetic for cancer surgery. PMID- 12127689 TI - Effect of combined photodynamic and chemotherapeutic treatment on lymphoma cells in vitro. AB - We investigated the cytotoxic and apoptotic effects of a combination of photodynamic therapy (PDT) and cisplatin (CDDP) on L5178Y (LY) cells. Treatment with PDT by photofrin((R)) (5 microg/ml) alone or with CDDP (20 microg/ml) alone killed 41.5+/-8.5% or 42.9+/-6.5% of LY cells, respectively, while a combination of these two treatments killed 99.7+/-0.6%. Apoptotic cell death after combination treatment was also revealed to be 49.6+/-7.8% compared to 12.4+/-3.4% after PDT alone, and 18.8+/-2.6% after CDDP. This study demonstrated that combined treatment of PDT and CDDP results in enhanced apoptotic cell death as well as a cytotoxic effect on LY cells. PMID- 12127690 TI - A combined gene delivery by co-transduction of adenoviral and retroviral vectors for cancer gene therapy. AB - A novel second generation retroviral producer cell strategy (an adenoviral/retroviral combined delivery system) has been developed by this laboratory. In the present studies, this delivery system was used to examine its delivery efficiency in vitro and in vivo by using a marker gene, LacZ, and a therapeutic gene, herpes simplex virus thymidine kinase (HSV-tk), both of which were transduced into a tumor cell line KBALB. In the in vitro experiments for delivery efficacy of the LacZ gene, the delivery efficiency of KBALB+KBALBLNPOZAdN/H (1:1) was 27.8% higher than that of KBALB+KBALBLNPOZ (1:1) (P<0.01). For the antitumor effect of HSV-tk/ganciclovir (GCV), the death ratio of KBALB+KBALBLNCTKAdN/H (1:1) was higher than that of KBALB+KBALBLNCTK (1:1), on 4, 6, and 8 days at a concentration of 0.1, 1, and 10 microg/ml, respectively (P<0.01 or P<0.05). In the in vivo experiments for LacZ gene expression, the delivery efficiency in KBALB+KBALBLNPOZAdN/H (1:1) was 21.5% more efficient than that in KBALB+KBALBLNPOZ (1:1) (P<0.01). For HSV-tk/GCV antitumor effect, the suppression of tumors by KBALB+KBALBLNCTKAdN/H (1:1) was more effective than that by KBALB+KBALBLNCTK (1:1) (P<0.05). Results suggest that this new delivery system is more efficient than the traditional in vitro and in vivo retroviral vector delivery system. PMID- 12127691 TI - Deletion of tumor suppressor genes in Chinese non-small cell lung cancer. AB - In the present study, we used 22 microsatellite markers flanking to or within 13 known or candidate tumor suppressor genes (TSGs) to detect loss of heterozygosity (LOH) in these chromosomal regions among 41 cases of non-small cell lung cancer, including 28 squamous cell carcinoma (SCC) and 13 adenocarcinoma (ADC). The studied TSGs comprised FHIT, VHL, APC, PRLTS, p16, IFNA, PTEN, p57, ATM, p53, BRCA1, DPC4 and DCC. Our data demonstrated frequent allelic losses of FHIT, p53, IFNA, VHL and p16 in both SCC and ADC. PTEN and ATM showed the least frequency of LOH, while no deletion of BRCA1 was detected in all tumor samples. LOH analysis of PRLTS was extended to 26 cases of ADC, which demonstrated significantly higher frequency of LOH than SCC. Our data indicated a possible correlation between specific TSG(s) and either histological type of lung cancer, and more attention should be paid to the PRLTS gene, which might play an important role in the development of ADC. PMID- 12127692 TI - Identification of genes differentially expressed between gastric cancers and normal gastric mucosa with cDNA microarrays. AB - To identify genes whose alterations lead to gastric cancer, gene expression profiles have been obtained from 22 gastric cancer tissues and their surrounding gastric mucosa tissues. A total of 16 genes were differentially expressed in more than 50% of gastric cancer tissues compared with surrounding gastric mucosa tissues. Genes such as HMG-Y, fibroblast collagenase inhibitor, and osteopontin are among those that are overexpressed in over 50% of the gastric cancer tissues. Dihydrodiol dehydrogenase, ribonuclease A, and glutathione peroxidase are among those genes that are underexpressed in over 50% of the gastric cancer tissues. We identified genes that are associated with clinical phenotypes of patients with gastric cancers. Alpha-II spectrin, Na/K-ATPase and KIAA0111 are those that are enhanced in intestinal type of gastric cancer. Gene such as platelet-endothelial tetraspan antigen 3 was enhanced in highly metastatic gastric cancer tissues. PMID- 12127693 TI - Fractionation of normal and leukemic T-cells by lectin-affinity column chromatography. AB - A method for rapid fractionation of normal and leukemic T-cells (Jurkat, RPMI 8402, MOLT-4), using lectin-affinity column chromatography, is described. CNBr activated Sepharose 6MB was used as a non-mobile phase. The gel was covalently conjugated with Dolichos biflorus agglutinin (DBA) over 24 h. The normal cells were eluted by phosphate buffered saline (Ca(2+) and Mg(2+) free), while the leukemic T-cells, interacting with DBA, were removed by N-acetyl-D-galactosamine or by low-concentrated acetic acid as a mobile phase. The cell fractions were detected spectrophotometrically at 600 nm. The rate of cell elution decreased in the order: normal>leukemic T-cells. The viability and the type of separated T cell fractions were characterized by flow cytometry, using adequate fluorescent antibodies. The interactions between leukemic T-cells and DBA-saturated Sepharose beads were examined by fluorescent microscopy, using fluorescent isothiocyanate DBA as a fluorescent marker. PMID- 12127694 TI - Clinical significance of serum and urinary c-erbB-2 levels in colorectal cancer. AB - In this study we measured serum and urinary c-erbB-2 levels in 63 patients with colorectal cancer and 29 healthy controls, assessing their role in cancer specific survival and the effects of resectional surgery. Serum and urinary c erbB-2 levels were measured by an enzyme-linked immunosorbent assay, preoperatively and 7 days following tumor resection. Preoperative serum c-erbB-2 concentrations were significantly higher in the cancer patients and correlated with disease stage and the presence of liver metastases. Urinary c-erbB-2 was detected more often in cancer patients, although levels did not differ from controls and there was no association with any clinicopathological variable. Serum c-erbB-2 levels decreased significantly in those patients resected for cure and were an independent prognostic factor for cancer-specific survival with higher preoperative concentrations correlating with worse overall survival. These findings suggest that serum assessment of c-erbB-2 concentration may be valuable in defining colorectal cancer prognosis. PMID- 12127695 TI - The expression of p53, c-erbB-1 and c-erbB-2 molecules and their correlation with prognostic markers in patients with head and neck tumors. AB - Molecular prognostic and predictive factors have extensively been studied in different cancers during the past decades, some of which were found to be useful in diagnosis, follow up or even treatment of some malignant tumors. To assess the significance of c-erbB-1, c-erbB-2 and p53 expression in head and neck tumors among Iranian patients and their correlation with known prognostic factors, samples from 53 patients with squamous cell carcinomas of larynx and tongue were studied immunohistochemically. Strong immunoreactivity of c-erbB-1, c-erbB-2 and p53 was observed in 37 (70%), 40 (76%) and 37 (70%) of cases, respectively. The coexpression of these molecules was detected in 27 (50.9%) samples. Neither histological grading nor nodal involvement revealed correlation with c-erbB-1 and/or c-erbB-2 expression. No correlation was found between p53 expression and histological grade. However, a significant positive association was observed between p53 expression and nodal involvement. This data, which is the first report on head and neck squamous cell carcinomas (HNSCC) in Iran, indicates the significance of p53 protein expression which may result from p53 tumor suppressor gene inactivation in lymph node metastasis of HNSCC among Iranian patients. PMID- 12127696 TI - Categorical perception of familiar objects. AB - We report three experiments where the categorical perception of familiar, three dimensional objects was investigated. A continuum of shape change between 15 pairs of objects was created and the images along the continuum were used as stimuli. In Experiment 1 participants were first required to discriminate pairs of images of objects that lay along the shape continuum. Then participants were asked to classify each morph-image into one of two pre-specified classes. We found evidence for categorical perception in some but not all of our object pairs. In Experiment 2 we varied the viewpoint of the objects in the discrimination task and found that effects of categorical perception generalized across changes in view. In Experiment 3 similarity ratings for each object pair were collected. These similarity scores correlated with the degree of perceptual categorization found for the object pairs. Our findings suggest that some familiar objects are perceived categorically and that categorical perception is closely tied to inter-object perceptual similarity. PMID- 12127697 TI - Thought before language: how deaf and hearing children express motion events across cultures. AB - Do children come to the language-learning situation with a predetermined set of ideas about motion events that they want to communicate? If so, is the expression of these ideas modified by exposure to a language model within a particular cultural context? We explored these questions by comparing the gestures produced by Chinese and American deaf children who had not been exposed to a usable conventional language model with the speech of hearing children learning Mandarin or English. We found that, even in the absence of any conventional language model, deaf children conveyed the central elements of a motion event in their communications. More surprisingly, deaf children growing up in an American culture used their gestures to express motion events in precisely the same ways as deaf children growing up in a Chinese culture. In contrast, hearing children in the two cultures expressed motion events differently, in accordance with the languages they were learning. The American children obeyed the patterns of English and rarely omitted words for figures or agents. The Chinese children had more flexibility as Mandarin permits (but does not demand) deletion. Interestingly, the Chinese hearing children's descriptions of motion events resembled the deaf children's descriptions more closely than did the American hearing children's. The thoughts that deaf children convey in their gestures thus may serve as the starting point and perhaps a default for all children as they begin the process of grammaticization--thoughts that have not yet been filtered through a language model. PMID- 12127698 TI - Sense of body and sense of action both contribute to self-recognition. AB - Recognizing oneself, easy as it appears to be, seems at least to require awareness of one's body and one's actions. To investigate the contribution of these factors to self-recognition, we presented normal subjects with an image of both their own and the experimenter's hand. The hands could make the same, a different or no movement and could be displayed in various orientations. Subjects had to tell whether the indicated hand was theirs or not. The results showed that a congruence between visual signals and signals indicating the position of the body is one component on which self-recognition is based. Recognition of one's actions is another component. Subjects had most difficulty in recognizing their hand when movements were absent. When the two hands made different movements, subjects relied exclusively on the movement cue and recognition was almost perfect. Our findings are in line with pathological alterations in the sense of body and the sense of action. PMID- 12127699 TI - A defense of the subordinate-level expertise account for the N170 component. AB - A recent paper in this journal reports two event-related potential (ERP) experiments interpreted as supporting the domain specificity of the visual mechanisms implicated in processing faces (Cognition 83 (2002) 1). The authors argue that because a large neurophysiological response to faces (N170) is less influenced by the task than the response to objects, and because the response for human faces extends to ape faces (for which we are not expert), we should reject the hypothesis that the face-sensitivity reflected by the N170 can be accounted for by the subordinate-level expertise model of object recognition (Nature Neuroscience 3 (2000) 764). In this commentary, we question this conclusion based on some of our own ERP work on expert object recognition as well as the work of others. PMID- 12127700 TI - Accounts for the N170 face-effect: a reply to Rossion, Curran, & Gauthier. AB - In their commentary, Rossion, Curran, and Gauthier (Rossion, B., Curran, T., Gauthier, I. (2002). A defense of the subordinate-level expertise account for the N170 component. Cognition, 85, 197-202) (RC&G) raise a series of arguments against the domain-specificity account for the N170 face-effect (Carmel, D., & Bentin, S. (2002). Domain specificity versus expertise: factors influencing distinct processing of faces. Cognition, 83, 1-29). This effect consists of a large difference (always significant) observed in the amplitude of a negative component peaking at the lower posterior-temporal sites in response to human faces relative to many other stimulus categories. As an alternative to domain specificity, RC&G advocate a "subordinate-level expertise" account, by which the N170 effect can be obtained for any type of stimulus for the individual identification of which the perceiver is an expert. While considering some of their arguments well taken and interesting, we believe that, overall, RC&G's interpretation of our current data (as well as some of theirs) and of our position ignores several important aspects and, therefore, their critique is not persuasive. PMID- 12127701 TI - Grammar overrides frequency: evidence from the online processing of flexible word order. AB - We show that online processing difficulties induced by word order variations in German cannot be attributed to the relative infrequency of the constructions in question, but rather appear to reflect the application of grammatical principles during parsing. Event-related brain potentials revealed that dative-marked objects in the initial position of an embedded sentence do not elicit a neurophysiologically distinct response from subjects, whereas accusative-marked objects do. These differences are predictable on the basis of grammatical distinctions (i.e. underlying linguistic properties), but not on the basis of frequency information (i.e. a superficial linguistic property). We therefore conclude that the former, but not the latter, guides syntactic integration during online parsing. PMID- 12127702 TI - The minimal unit of phonological encoding: prosodic or lexical word. AB - Wheeldon and Lahiri (Journal of Memory and Language 37 (1997) 356) used a prepared speech production task (Sternberg, S., Monsell, S., Knoll, R. L., & Wright, C. E. (1978). The latency and duration of rapid movement sequences: comparisons of speech and typewriting. In G. E. Stelmach (Ed.), Information processing in motor control and learning (pp. 117-152). New York: Academic Press; Sternberg, S., Wright, C. E., Knoll, R. L., & Monsell, S. (1980). Motor programs in rapid speech: additional evidence. In R. A. Cole (Ed.), The perception and production of fluent speech (pp. 507-534). Hillsdale, NJ: Erlbaum) to demonstrate that the latency to articulate a sentence is a function of the number of phonological words it comprises. Latencies for the sentence [Ik zoek het] [water] 'I seek the water' were shorter than latencies for sentences like [Ik zoek] [vers] [water] 'I seek fresh water'. We extend this research by examining the prepared production of utterances containing phonological words that are less than a lexical word in length. Dutch compounds (e.g. ooglid 'eyelid') form a single morphosyntactic word and a phonological word, which in turn includes two phonological words. We compare their prepared production latencies to those syntactic phrases consisting of an adjective and a noun (e.g. oud lid 'old member') which comprise two morphosyntactic and two phonological words, and to morphologically simple words (e.g. orgel 'organ') which comprise one morphosyntactic and one phonological word. Our findings demonstrate that the effect is limited to phrasal level phonological words, suggesting that production models need to make a distinction between lexical and phrasal phonology. PMID- 12127703 TI - Learning and development in neural networks--the importance of prior experience. AB - Infants can discriminate between familiar and unfamiliar grammatical patterns expressed in a vocabulary that is distinct from that used earlier during familiarization (Cognition 70(2) (1999) 109; Science 283 (1999) 77). Various models have captured the data, although each required that discrimination be distinct, in terms of the computational process, from familiarization. This article describes a simple recurrent network (SRN), equipped only with the assumption that it should predict what comes next, which models the data without distinguishing between familiarization and discrimination. To accomplish this, the SRN requires pre-training on a range of sequences instantiating different structures and different vocabulary items to those used subsequently during familiarization and test. Pre-training enables the network to avoid replacing structure acquired during familiarization with structure experienced at test. An equivalent enabling condition may underpin infants' resistance to catastrophic interference between the different structures and vocabulary items to which they are exposed. PMID- 12127704 TI - Modularity and spatial reorientation in a simple mind: encoding of geometric and nongeometric properties of a spatial environment by fish. AB - When disoriented in environments with distinctive geometry, such as a closed rectangular arena, human infants and adult rats reorient in accord with the large scale shape of the environment, but not in accord with nongeometric properties such as the colour of a wall. Human adults, however, conjoined geometric and nongeometric information to reorient themselves, which has led to the suggestion that spatial processing tends to become more flexible over development and evolution. We here show that fish tested in the same tasks perform like human adults and surpass rats and human infants. These findings suggest that the ability to make use of geometry for spatial reorientation is an ancient evolutionary tract and that flexibility and accessibility to multiple sources of information to reorient in space is more a matter of ecological adaptations than phylogenetic distance from humans. PMID- 12127705 TI - Procalcitonin as a marker of sepsis. AB - Prompt diagnosis and treatment with appropriate antimicrobial chemotherapy is of paramount importance to reduce morbidity and mortality associated with sepsis. Inflammatory markers currently in use, such as C-reactive protein (CRP) do not reliably differentiate between the systemic inflammatory response and sepsis. Procalcitonin (PCT), a precursor of calcitonin, is a 116 amino acid protein that has been proposed as a marker of disease severity in conditions such as septicaemia, meningitis, pneumonia, urinary tract infection (UTI) and fungal and parasitic infection. In particular, serial measurements are useful in order to monitor response to therapy. Together with good clinical judgement and judicious use of antimicrobial agents, PCT should serve as a valuable adjunct in the diagnosis and management of sepsis. PMID- 12127706 TI - Pathogen occurrence and antimicrobial resistance trends among urinary tract infection isolates in the Asia-Western Pacific Region: report from the SENTRY Antimicrobial Surveillance Program, 1998-1999. AB - Worldwide surveillance of antimicrobial resistance among urinary tract pathogens is useful to determine important trends and geographical variation for common Gram-positive and -negative species. The most common causative uropathogens often have intrinsic or acquired resistance mechanisms which include ESBL production among enteric bacilli, multi-drug resistant staphylococci and non-fermentative Gram-negative bacilli such as Pseudomonas aeruginosa and Acinetobacter spp. and vancomycin-resistant Enterococcus spp. This study evaluates pathogen frequency and the resistance rates among urinary tract infection (UTI) pathogens in 14 medical centres in the Asia-Pacific region between 1998 and 1999. The isolates were referred to a central monitor for reference NCCLS broth microdilution testing, identification confirmation and patient demographic analysis. Over 50% of the 958 pathogens were Escherichia coli and Klebsiella spp. followed by P. aeruginosa, Enterococcus spp. and Enterobacter spp. Susceptibility for the three enteric bacilli was high for carbapenems (100%), 'fourth-generation' cephalosporins (cefepime 94.9-98.6%) and amikacin (> or = 93.0%). Beta-lactamase inhibitor compounds were more active against E. coli (piperacillin/tazobactam; > 90% susceptible) than the other two enteric species and all other tested agents had a narrower spectra of activity. The rank order of anti-pseudomonal agents was amikacin (91.5% susceptible)> imipenem > piperacillin/tazobactam > tobramycin > ceftazidime and cefepime (77.4 and 76.4% susceptible, respectively). Susceptibility to quinolones for the P. aeruginosa isolates was only 63.2-67.0%. Only one vancomycin-intermediate Enterococcus spp. (van C phenotype) was detected among the 103 strains tested. Newer fluoroquinolones (gatifloxacin; MIC(50), mg/l) were more potent against enterococci than ciprofloxacin (MIC(50), 2 mg/l) and high-level resistance to aminoglycosides was common (41.7%). The data presented are compared to studies of similar design from other areas which are part of the SENTRY surveillance network. PMID- 12127707 TI - Lomefloxacin versus ciprofloxacin in the treatment of chronic bacterial prostatitis. AB - A total of 182 patients with chronic bacterial prostatitis (CBP) were recruited into this multicentre prospective, randomized clinical study. Of these, 93 were treated orally with lomefloxacin (LOMX) 400 mg once daily and 89 with ciprofloxacin (CIPX) 500 mg twice daily for 4 weeks. At 5-9 days 90/90 vs 86/86, at 4-6 weeks 82/83 vs 82/82, at 3 months 80/79 vs 78/75, and at 6 months 78/75 vs 75/72 patients aging from 18 to 70 years were evaluable bacteriologically/clinically according to a modified intention to treat evaluation. The most frequent pathogens were Escherichia coli, followed by staphylococci, enterococci and Proteus mirabilis. At 5-9 days, 4-6 weeks, 3 and 6 months after therapy the rates of eradication without superinfection per evaluable patients (100%) were 80, 72, 74, and 63% in the LOMX group and 84, 81, 82, and 72% in the CIPX group and (cure and improvement) rates were 98 vs 97%, 84 vs 90%, 86 vs 89%, and 81 vs 89%. There were no statistically significant differences (P < 0.05) between the results of the two treatment groups. Nine (5 vs 4) patients were withdrawn because of adverse events. From the bacteriological and clinical results including adverse events, the oral treatment of CBP over 4 weeks with LOMX 400 mg once daily was comparably effective and tolerable with that of CIPX 500 mg twice daily. PMID- 12127708 TI - Comparative efficacy and safety of 5-day cefaclor and 10-day amoxycillin treatment of group A streptococcal pharyngitis in children. AB - A total of 384 children with group A beta-haemolytic streptococcal (GABHS) pharyngitis were randomized to receive either 40 mg/kg/day of cefaclor in two doses for 5 days (192 patients), or 40 mg/kg/day of amoxycillin in three doses for 10 days (192 patients). The signs and symptoms of pharyngitis were recorded and throat cultures were obtained at presentation and on days 6-7, 11-15, 16-20 and 28-35. Patient compliance was significantly higher in the children treated with cefaclor (100 vs. 95.1%; P = 0.003). At the end of follow-up, the percentage of clinical success was 91.4% for cefaclor and 91.9% for amoxycillin (P = 0.974); bacteriological success was obtained in 85.7 and 89.6% children (P = 0.348), respectively. Both treatments were well-tolerated with adverse event rates of 8.3% in the cefaclor group and 9.4% in the amoxcillin group (P = 0.857). Our study shows that five days' treatment with cefaclor is as effective and safe as the conventional 10-day course of amoxycillin in the treatment of GABHS pharyngitis, but compliance seems to be significantly greater. PMID- 12127709 TI - Intracellular activity of clinical concentrations of phenothiazines including thioridiazine against phagocytosed Staphylococcus aureus. AB - The effect of thioridazine (TZ) was studied on the killing activity of human peripheral blood monocyte derived macrophages (HPBMDM) and of human macrophage cell line THP-1 at extracellular concentrations below those achievable clinically. These macrophages have nominal killing activity against bacteria and therefore, would not influence any activity that the compounds may have against intracellular localised Staphylococcus aureus. The results indicated that whereas TZ has an in vitro minimum inhibitory concentration (MIC) against the strains of S. aureus of 18, 0.1 mg/l of TZ in the medium completely inhibits the growth of S. aureus that has been phagocytosed by macrophages. The latter concentration was non-toxic to macrophages, did not cause cellular expression of activation marker CD69 nor induction of CD3+ T cell production of IFN-gamma, but blocked cellular proliferation and down-regulated the production of T cell-derived cytokines (IFN gamma, IL-5). These results suggest that TZ induces intracellular bactericidal activities independent of the capacity to generate Type 1 responses against S. aureus. PMID- 12127710 TI - Bacterial strain-independent AUC/MIC and strain-specific dose-response relationships reflecting comparative fluoroquinolone anti-pseudomonal pharmacodynamics in an in vitro dynamic model. AB - To compare the antimicrobial effects (AMEs) of two quinolones in terms of the AUC/MIC- and dose (D)-response relationships, five differentially susceptible clinical isolates of Pseudomonas aeruginosa were exposed to decreasing concentrations of ciprofloxacin (two 12-h doses with T(1/2) = 4 h) and trovafloxacin (a single dose with T(1/2) = 9.2 h). The simulated AUC/MICs of ciprofloxacin ranged from 58 to 932 and those of trovafloxacin, from 54 to 466 h. The intensity of the AME (I(E)) correlated well with log AUC/MIC for both ciprofloxacin and trovafloxacin (r(2) = 0.99 and 0.97, respectively) in a strain independent fashion. At a given AUC/MIC ratio, AMEs of trovafloxacin were greater than ciprofloxacin. However, based on the respective I(E)-logD curves, 200 mg trovafloxacin produced a slightly greater AME than 2 x 500 mg ciprofloxacin only with the most susceptible P. aeruginosa. With the less susceptible P. aeruginosa ciprofloxacin was more efficient than trovafloxacin. This study suggests that both bacterial strain-independent AUC/MIC- and the respective strain-specific dose-response relationships of the AME are important for comprehensive pharmacodynamic evaluation of antimicrobial agents. PMID- 12127711 TI - In vitro microbicidal activity of W/O/W multiple emulsion for vaginal administration. AB - The microbicidal activity of a W/O/W multiple emulsion destined for vaginal application, containing lactic acid in the internal aqueous phase, octadecylamine (ODA) in the oily phase and benzalkonium chloride (CBZ) in the external aqueous phase was evaluated against three microbial strains: Escherichia coli, Staphylococcus aureus and Candida albicans. The results were different depending on the procedure used. Interpretable results were obtained if only a gentle agitation was used just after the introduction of the microbial suspension to the product. This suggested that vigorous agitation lead to a variable fraction of CBZ or ODA entrapped in the micelles of ethylene and propylene oxide copolymer (COE). PMID- 12127712 TI - Synergistic activity of the novel des-fluoro(6) quinolone, garenoxacin (BMS 284756), in combination with other antimicrobial agents against Pseudomonas aeruginosa and related species. AB - Non-fermentative Gram-negative bacteria (Pseudomonas aeruginosa, Burkholderia cepacia, Stenotrophomonas maltophilia and Acinetobacter spp.) are intrinsically less susceptible to many antimicrobial agents. Two-drug combinations have been used to treat infections caused by less susceptible pathogens. In this study, the antibacterial activity of garenoxacin (GARX) with non-quinolones was examined. The non-quinolones evaluated were cefepime (CEPI), imipenem (IMIP), aztreonam (AZTR), piperacillin-tazobactam (PIPC/TZ), amikacin (AMK), ceftazidime (CTAZ), trimethoprim-sulphamethoxazole (TMP/SMX) and ticarcillin-clavulanate (TICC/CA). Synergism was determined by time-kill analysis using GARX (at 2 x its MIC, not to exceed 4 mg/l) and the second drug (at 1 x MIC, not to exceed its susceptible MIC breakpoint), and is defined as > or = 2 log(10) enhanced killing at 24 h with the combination. Partial synergy is defined as > or = 1.5 log(10) but < 2 log(10) enhanced killing with the drug combination. Synergy/partial synergy was observed most often with GARX plus: CEPI, AZTR, PIPC/TZ, IMIP (five strains each) or AMK (four strains) vs. eight P. aeruginosa; CTAZ, AZTR (five strains each) vs. six B. cepacia; TICC/CA (six strains), CEPI, CTAZ or AMK (five strains each) vs. eight S. maltophilia; and CEPI, AMK (three strains each) or CTAZ, TICC/CA (two strains each) vs. four Acinetobacter spp. In conclusion, synergistic killing was observed frequently with GARX plus a non-quinolone bactericidal agents against non fermentative Gram-negative bacteria, including strains intermediately susceptible/resistant to one or both agents. PMID- 12127713 TI - In vitro activity of linezolid, synercid and telithromycin against genetically defined high level fluoroquinolone-resistant methicillin-resistant Staphylococcus aureus. AB - The in vitro activity of levofloxacin, moxifloxacin, gatifloxacin, erythromycin, telithromycin, linezolid, synercid and vancomycin was measured against 36 genetically defined, gyrA/grlA double mutant MRSA clinical strains with an MIC to ciprofloxacin > or = 8 mg/l. The three newer fluoroquinolones tested were more active than ciprofloxacin. Resistance rates for levofloxacin and gatifloxacin were high (44.5 and 36.1%, respectively). All the strains were moxifloxacin susceptible, though most of them had MICs close to the break point. All the strains were intermediate or resistant to erythromycin and most were also resistant to telithromycin. No strains were resistant to linezolid, synercid or vancomycin (MIC(90): 2, 1 and 2 mg/l, respectively). PMID- 12127714 TI - Comparative antimicrobial activity of lysosubtilin and its acid-resistant derivative, Fermosorb. AB - Comparative data are given on the in vitro activity of lysosubtilin against micro organisms and its acid-resistant derivative, Fermosorb. The lytic activity Fermosorb against all micro-organisms tested was higher (in the range of 0.9-7.7 and 4.1-13.5% for bacteria and filamentous fungi/yeasts, respectively) than that achieved by the action of lytic enzymes present in lysosubtilin solution. All six lysosubtilin-resistant strains tested were Fermosorb-susceptible. Considering the increase in the incidence of antibiotic resistance antimicrobial enzymotherapy or enzymoprophylaxis by means of Fermosorb might be considered for the treatment of intestinal infections in animals. PMID- 12127715 TI - Antimicrobial anionic peptide binds in vivo to Mannheimia (Pasteurella) haemolytica attached to ovine alveolar epithelium. AB - Endogenous antimicrobial peptide activity in vivo has rarely been demonstrated. To assess this, Mannheimia haemolytica (log(10) 10.20 cfu) was deposited into the lungs of adult sheep, which were killed at 0, 5, 10 and 20 min for necropsy. At 0 min, M. haemolytica appeared normal and monoclonal antibody to antimicrobial anionic peptide (AP) and Protein A-colloidal gold identified AP already bound to the bacterial surface. At 5-20 min, many organisms were distorted with flocculated intracellular constituents characteristic of AP cellular damage indicating that AP can bind to and presumably help inactivate organisms in vivo. PMID- 12127717 TI - Encapsulation of pancreatic islets for transplantation in diabetes: the untouchable islets. AB - The aim of encapsulation of pancreatic islets is to transplant in the absence of immunosuppression. It is based on the principle that transplanted tissue is protected from the host immune system by an artificial membrane. Encapsulation allows for application of insulin-secreting cells of animal or other surrogate sources, to overcome human islet shortage. The advantages and pitfalls of the approaches developed so far are discussed and compared, together with some recent progress, in view of applicability in clinical islet transplantation. PMID- 12127716 TI - Smad7: a new key player in TGF-beta-associated disease. AB - Smad7 is a major inhibitory regulator of transforming growth factor (TGF)-beta signaling. Smad7 expression is induced by TGF-beta itself and other signaling pathways, indicating a key role for Smad7 in feedback or cross-talk control of TGF-beta signaling. Recent reports have implicated Smad7 as a crucial regulator of TGF-beta activity in human disease; aberrant expression of Smad7 is involved in inflammatory bowel disease and scleroderma. Thus, modulation of Smad7 expression could provide a novel therapeutic basis for TGF-beta-associated disorders. PMID- 12127722 TI - Atherosclerosis: another protein misfolding disease? AB - The secondary structure and conformation of apo-B 100 in low-density lipoproteins (LDL) are imposed by lipid-protein interactions and dynamics, and affected by the introduction or removal of lipids during the course of lipoprotein metabolism. Following an alteration of the water-lipid interface as a result of, for example, oxidation of lipids, the supramolecular structure becomes destabilized and apoB can misfold. These events have been observed in LDL(-), a fraction of oxidatively modified LDL isolated in vivo. This modified lipoprotein possesses several atherogenic properties and represents an in vivo counterpart of in vitro modified LDL that is implicated in atherosclerosis. The misfolding of apoB, its aggregation, resistance to proteolysis, and cytotoxicity are common motifs shared by LDL(-) and amyloidogenic proteins. Based on these analogies, we propose that atherogenesis could be considered as a disease produced by the accumulation of cytotoxic and pro-inflammatory misfolded lipoproteins. PMID- 12127723 TI - Pancreatic beta-cell growth and survival--a role in obesity-linked type 2 diabetes? AB - Obesity-linked type 2 diabetes is a disease of insulin resistance combined with pancreatic beta-cell dysfunction. Although a role for beta-cell mass in the pathogenesis of obesity-linked type 2 diabetes has recently gained prominence, the idea is still being developed. It is proposed that in early obesity an increase in beta-cell mass and function might compensate for peripheral insulin resistance. However, as time and/or the severity of the obesity continue, there is decay in such adaptation and the beta-cell mass becomes inadequate. This, together with beta-cell dysfunction, leads to the onset of type 2 diabetes. It is becoming evident that elements in insulin and insulin growth factor (IGF)-1 signal-transduction pathways are key to regulating beta-cell growth. Current evidence indicates that interference of insulin signaling in obesity contributes to peripheral insulin resistance. This article examines whether a similar interference of IGF-1 signaling in the beta-cell could hinder upregulation of beta-cell mass and/or function, resulting in a failure to compensate for insulin resistance. PMID- 12127724 TI - NF-kappaB as a therapeutic target in cancer. AB - The transcription factor nuclear factor (NF)-kappaB is activated in certain cancers and in response to chemotherapy and radiation. The transcriptional activation of genes associated with cell proliferation, angiogenesis, metastasis and suppression of apoptosis appears to lie at the heart of the ability of NF kappaB to promote oncogenesis and cancer therapy resistance. Supporting these findings are recent experiments, performed in vitro and using xenograft models of cancer, which implicate NF-kappaB inhibition as an important new approach for the treatment of certain hematological malignancies and as an adjuvant approach in combination with chemotherapy or radiation for a variety of cancers. Clinical trials with drugs that block NF-kappaB are currently in progress with promising results. However, as there is currently no drug that blocks specific NF-kappaB activation, conclusions drawn with small-molecule inhibitors must be interpreted carefully. PMID- 12127725 TI - Prostanoids and pain: unraveling mechanisms and revealing therapeutic targets. AB - Advances in our understanding of the synthesis, regulation and function of prostanoids have led to a new appreciation of their actions in health and disease. Prostanoid synthesis is essential for the generation of inflammatory pain and this depends not only on prostanoid production at the site of inflammation, but also on the actions of prostanoids synthesized within the central nervous system (CNS). Moreover, central prostanoid synthesis is controlled both by neural and humoral signals, the latter being a novel form of input to the CNS. Diverse compounds that act along the pathway of prostanoid synthesis and action, both in the periphery and in the CNS, might provide increased benefit for treating inflammatory pain hypersensitivity and its associated sickness syndrome, with a reduced risk of adverse effects. PMID- 12127726 TI - Reciprocal products of chromosomal translocations in human cancer pathogenesis: key players or innocent bystanders? AB - Chromosomal translocations are frequently involved in the pathogenesis of leukemias, lymphomas and sarcomas. They can lead to aberrant expression of oncogenes or the generation of chimeric proteins. Classically, one of the products is thought to be oncogenic. For example, in acute promyelocytic leukaemia (APL), reciprocal chromosomal translocations involving the retinoic acid receptor alpha (RARalpha) gene lead to the formation of two fusion genes: X RARalpha and RARalpha-X (where X is the alternative RARalpha fusion partner: PML, PLZF, NPM, NuMA and STAT 5b). The X-RARalpha fusion protein is indeed oncogenic. However, recent data indicate that the RARalpha-X product is also critical in determining the biological features of this leukemia. Here, we review the current knowledge on the role of reciprocal products in cancer pathogenesis, and highlight how their expression might impact on the biology of their respective tumour types. PMID- 12127732 TI - Dietary accumulation and biochemical responses of juvenile rainbow trout (Oncorhynchus mykiss) to 3,3',4,4',5-pentachlorobiphenyl (PCB 126). AB - Juvenile rainbow trout (Oncorhynchus mykiss) (initial weights 2-5 g) were exposed to three dietary concentrations (0, 12.4 and 126 ng g(-1), wet weight) of a 14C labelled 3,3',4,4',5-pentachlorobiphenyl (PCB 126) for 30 days followed by 160 days of clean food. We assessed bioaccumulation, histology (liver and thyroid) and biochemical responses (liver ethoxyresorufin-O-deethylase (EROD), liver vitamins (retinoids and tocopherol) and muscle thyroid hormone levels) along with growth and survival. The half-life of PCB 126 in the rainbow trout ranged from 82 to 180 days while biomagnification factors (BMF) ranged from 2.5 to 4.1 providing further evidence that PCB 126 is among the most bioaccumulative PCB congeners. Toluene extractable 14C declined with time in the trout suggesting the possibility of some biotransformation and/or covalent bonding with biological macromolecules. The threshold for liver EROD induction by PCB 126 was approximately 0.1 ng g(-1) (wet weight). EROD activities in the low- and high treatments were 9 and 44 times greater than control, respectively, and remained elevated throughout the experiment. EROD activity was correlated with whole body concentrations of PCB 126 although there was evidence of EROD activity suppression in the highly exposed fish. Liver didehydroretinoids and tocopherol concentrations were depressed by the high PCB 126 dose after 30 days exposure. Initially, muscle concentrations of thyroxine (T4) and triiodo-L-thyronine (T3) declined as the fish grew during the experiment, and exposure to PCB 126 accelerated the growth related decline. More information is needed to assess the functional significance of the reduced muscular stores of thyroid hormones. Despite the changes in liver EROD, liver vitamins and muscle thyroid hormones, liver and thyroid histology in trout examined after 30 days exposure and growth parameters were unaffected by PCB 126. This indicates that the functional competences of the physiological factors associated with growth were maintained under the experimental conditions. PMID- 12127733 TI - Photoperiod effects on the UV-induced toxicity of fluoranthene to freshwater mussel glochidia: absence of repair during dark periods. AB - The effect of photoperiod on the ultraviolet radiation (UV)-induced toxicity of fluoranthene to glochidia of the freshwater mussel, Utterbackia imbecillis, was evaluated using a series of static renewal toxicity tests conducted using one of four different photoperiods (24 h light, 16 h light:8 h dark, 12 h light:12 h dark, 8 h light:16 h dark). Rates of acute mortality were dependent both upon fluoranthene dose and the photoperiod. Median lethal time (LT50) values calculated on the basis of accumulated UV exposure time (UV-LT50) were compared with LT50 values calculated from real time of exposure (R-LT50) to determine relative rates of photoactivated fluoranthene damage versus physiologic repair during periods of darkness. UV-LT50 values were only dependent on fluoranthene dose and not on photoperiod. The fact that UV-LT50 values did not increase with decreasing light cycle length indicates that physiologic repair during dark periods was not an important process in these experiments. These findings suggest that (1) species-specific and/or life history-specific factors, in part, determine the ability of an organism to repair photoactivated polycyclic aromatic hydrocarbon (PAH) damage during dark periods; and (2) predictions of the UV induced toxic response of PAH in glochidia need only be based upon total UV dose (dose rate and duration) and PAH dose. PMID- 12127734 TI - Numerical chromosomal aberrations in the early life-history stages of a marine tubeworm, Pomatoceros lamarckii (Polychaeta: Serpulidae). AB - The marine environment provides a sink for a host of toxic chemicals, directly or inadvertently, released as a result of human activity. Some of these chemicals have the potential to act as aneugens, substances that cause numerical chromosomal aberrations (NCAs). NCAs are one of the most important classes of genetic abnormality and are implicated in a variety of deleterious effects, including premature ageing, birth defects and cancer. Clearly, any increase in the incidence of these agents in the marine environment poses a risk to the indigenous biota and its predators, including man. In this paper, we describe our recent success with applying the fluorescence in situ hybridisation technique (FISH) to detect NCAs in the interphase cell nuclei of Pomatoceros lamarckii, a common rocky shore invertebrate. Given the lack of requirement for any detailed cytogenetic knowledge, the method holds considerable promise for laboratory and field studies in general, and should lend itself to automated screening protocols, where large numbers of cells can be screened rapidly, for example, using a flow cytometer. When exposed either under acute or chronic (viz. adult) exposure conditions, colchicine and di-butylphthalate (DBP) (a widely-used plasticiser), two recognised aneugens, induced significant increases in the levels of NCAs, in the dose range 1 x 10(-6)-5 x 10(-6) M, in both four to eight cell embryo stages and 24 h-old larvae. In keeping with the severely debilitating effects of this class of agent, an inverse correlation was observed between the induced levels of NCAs and larval fitness based on the results of a standard 48-h larval bioassay. PMID- 12127735 TI - How estrogenic is nonylphenol? A transgenerational study using rainbow trout (Oncorhynchus mykiss) as a test organism. AB - The aim of the present study was to evaluate both estrogenic effects in directly NP-exposed sexually mature rainbow trout and possible transgenerational effects in the offspring of exposed fish. Four months prior to spawning, adult rainbow trout of both sexes were exposed intermittently to NP concentrations of 1 and 10 microg/l. At the end of the exposure period, which coincided with the beginning of spawning time, vitellogenin levels in the plasma of adult male rainbow showed a significant increase compared to the control group. After exposure to 10 microg NP/l reproduction was impaired as indicated by significantly reduced hatching rates. Histological examination of the testicular tissue of NP-exposed individuals revealed no morphological differences from the controls. In the offspring, vitellogenin levels of male individuals were not affected, whereas in females they were significantly higher than in the control progeny. The histological examination revealed no alteration in sex ratios. In single cases, intersex occurred in both male and female offspring of exposed fish. The analysis of sex steroid levels revealed a two-fold increase of estradiol in the plasma of male offspring and a 13-fold elevation of testosterone in the plasma of female progeny. The present findings indicate that NP, in an environmentally relevant concentration range, acts as a weak estrogen in directly exposed adult male rainbow trout as indicated by elevated plasma vitellogenin levels. Reproduction success was reduced as indicated by decreased hatching rates. Hormonal imbalances detected in the offspring of exposed fish indicate a transgenerational effect mediated by the endocrine system. PMID- 12127736 TI - Biodegradation and enzymatic responses in the marine diatom Skeletonema costatum upon exposure to 2,4-dichlorophenol. AB - The biodegradation and responses of selected detoxification and antioxidant enzymes in the marine diatom, Skeletonema costatum, upon exposure to sublethal concentrations of 2,4-dichlorophenol (2,4-DCP) were investigated. Results show that 2,4-DCP was readily metabolised, but bioaccumulation and adsorption were negligible. Glutathione S-transferase, ascorbate peroxidase and superoxide dismutase activities were increased markedly after exposure to 2,4-DCP for 96 h, while no appreciable change in peroxidase activity was observed. The addition of exogeneous glutathione to diatom culture enhanced the degradation of 2,4-DCP, and promoted diatom growth. The inhibition of glutathione synthesis enhanced the toxicity of 2,4-DCP. These results suggest that glutathione conjugation was one of the principal mechanisms involved in the degradation of 2,4-DCP in this diatom. PMID- 12127737 TI - Accumulation of microcystins by a tropical zooplankton community. AB - In the current study, the hepatotoxic peptide microcystins, were measured in the zooplankton community of Jacarepagua Lagoon during a 6-month period. Concurrent phytoplankton and seston samples were obtained. Microcystins were measured in seston and phytoplankton by High Performance Liquid Chromatography (HPLC), and in zooplankton by an Enzyme-Linked Immunosorbant Assay (ELISA). Zooplankton community was comprised mainly by the rotifers Brachionus angularis and B. plicatilis, the cladocerans Moina micrura and Ceriodaphnia cornuta and the copepod Metacyclops mendocinus. Phytoplankton was dominated by Microcystis aeruginosa during all the studied period. Microcystins in zooplankton ranged from 0.3 to 16.4 microg g(-1) DW, while in the sestonic samples they ranged from undetectable values to 5.8 ng g(-1) DW. Microcystins in net phytoplankton ranged from 0.3 to 3.9 mg g(-1) DW. We conclude that zooplankton from Jacarepagua Lagoon were efficient accumulators of microcystins from seston and that these animals can be potential vectors in the transferring of such toxins to higher trophic levels in the aquatic food chain. PMID- 12127738 TI - Algal esterase activity as a biomeasure of environmental degradation in a freshwater creek. AB - This study investigated the potential for using algal esterase activity of Microcystis aeruginosa and Selenastrum capricornutum as a rapid measure of the biological effects of acid mine drainage (AMD) in a South Australian stream (Australia) also affected by sewage pollution and dry-land salinity. Algal bioassays were based on the non-fluorescent substrate, fluorescein diacetate (FDA) which is metabolised by esterases to the fluorescent product, fluorescein. Esterase activity was interpreted as the mean rate of conversion of FDA to fluorescein and expressed as a percentage of the rate achieved by control algae (%FDAC). Flow cytometry was used to measure the fluorescence of individual algal cells, enabling differentiation of three esterase activity states (low=S(1), normal and stimulated) and calculation of the percentage of algal cells in each activity state relative to that found for control algae (e.g. %S(1)). Algal esterase activity responded rapidly to AMD-affected water but also to increased conductivity (associated with dry-land salinity) and nutrient concentrations (associated with sewage). Exposure to AMD-affected water for 1 h reduced %FDAC by 30-70%, and increased %S(1) by 60-90%, a depression of esterase activity that was maintained over 24 h. A similar depression of esterase activity occurred in both algae exposed to comparatively high-conductivity water (ca. 20 mS cm(-1)) for 1 h but the algae recovered from this 'shock' within 24 h. The %FDAC of S. capricornutum increased from 66 to 158% of control values after a 24 h exposure to nutrient-enriched water sampled downstream from a sewage treatment plant, despite the fact that the alga was grown in nutrient-sufficient culture. The combination of cyanobacterial (M. aeruginosa) and green (S. capricornutum) algal cultures with exposure times of 1 and 24 h was successful in distinguishing between the three types of pollution. Correlation of esterase activity measures with water quality parameters indicated that the clearest and least equivocal biological measure of AMD for the study area was the %S(1) for M. aeruginosa after a 24 h exposure. The use of the flow cytometer to define a low esterase activity state was therefore successful in clarifying the response to AMD affected water. The study demonstrates the successful application of algal esterase activity bioassays, in combination with flow cytometry, to rapidly assess the toxicity of AMD-affected waters and to differentiate this response from the effects of other pollutants (increased nutrients and conductivity). PMID- 12127739 TI - Physiological response of freshwater microalga (Chlorella vulgaris) to triazine and phenylurea herbicides. AB - The effects of two herbicides used wide-spread, isoproturon (phenylurea) and terbutryn (triazine), on growth, dry weight, elemental composition, photosynthetic pigments and protein content, and cell volume assayed by flow cytometry in the freshwater microalgae Chlorella vulgaris were studied. Different parameters for algal activity show widely different sensitivities to these aquatic pollutants. After 96 h of herbicide exposure, terbutryn was the strongest inhibitor of growth, giving an EC50 value for growth twice lower than that for isoproturon cultures (EC50 terbutryn=0.097 microM; EC50 isoproturon=0.199 microM). However, lower concentrations of the triazine herbicide provoked an increase in the cellular density and growth rate of this microalga, not observed in the phenylurea-treated cultures. Cellular volume and dry weight of C. vulgaris cells were increased strongly in the presence of isoproturon and terbutryn. Other cellular parameters, such as pigment and protein content, were stimulated with both herbicides at higher concentrations. PMID- 12127740 TI - Expression of P-glycoprotein in killifish (Fundulus heteroclitus) exposed to environmental xenobiotics. AB - P-glycoproteins (P-gp) are transmembrane efflux flippases that prevent the cellular accumulation of moderately hydrophobic compounds and are responsible for certain multidrug resistance phenotypes in tumor cell lines and human patients. We investigated whether P-gps could be involved in a contaminant resistant phenotype observed in a population of fish exposed over generations to high levels of planar halogenated aromatic hydrocarbons (PHAHs). Hepatic and intestinal epithelial P-gp expression was examined by immunoblot and immunohistochemistry in killifish (Fundulus heteroclitus) from New Bedford Harbor, MA (NBH), a Superfund site highly contaminated with PHAHs, and from Scorton Creek on Cape Cod, MA (SC), a relatively unpolluted site. The NBH population has developed resistance to the toxicity of PHAHs. Hepatic P-gp levels were more than 40% greater in fish freshly collected from SC than in fish freshly collected from NBH. When killifish from either site were maintained in clean water for up to 78 days to permit depuration of bioaccumulated contaminants, hepatic P-gp levels decreased approximately 50% by day 8. P-glycoprotein expression was detected in the intestinal epithelium in 55% of freshly collected NBH fish. However, depurated NBH fish and freshly caught and depurated SC fish rarely expressed P-gp in the intestine. In an effort to determine whether environmental chemicals at the two sites might contribute to altered P-gp expression, depurated fish were exposed either to sediment collected from SC or 2,3,7,8-tetrachlorodibenzofuran, a contaminant found at the NBH site and a model aryl hydrocarbon receptor agonist. Neither exposure affected hepatic P-gp levels in killifish. Elevated intestinal P-gp in NBH fish might counter the absorption of P-gp substrates/inducers and thus limit the amount of these compounds reaching the liver, which might account for the lower hepatic P-gp levels in NBH fish compared to SC fish. The differences in hepatic P-gp levels (SC>NBH) and intestinal P-gp (NBH>SC) in freshly collected fish also might reflect environmental exposure to different anthropogenic contaminants or microbial, algal, plant or other natural products via the water column, sediment, or diet at each site. PMID- 12127741 TI - Toxicity of selenium and other elements in food organisms to razorback sucker larvae. AB - Elevated selenium concentrations documented in water, sediment, and biota in irrigation drain water studies by U.S. Department of the Interior agencies and academia have raised concerns that selenium may be adversely affecting endangered fish in the upper Colorado River basin. The objective of the study was to determine the effects on endangered razorback sucker (Xyrauchen texanus) larvae from exposure to selenium and other trace elements in water and zooplankton collected from sites adjacent to the Colorado River near Grand Junction, CO. A 30 day study was initiated with 5-day-old larvae exposed in a 4 x 4 factor experiment with four food and four water treatments, and the biological endpoints measured were survival, growth, development, and whole-body residues of selenium. Mean selenium concentration in reference water (24-Road) was <0.7 microg/l, in reference food (brine shrimp) was 3.2 microg/g, at Horsethief was 1.6 microg/l in water and 6.0 microg/g in zooplankton, at Adobe Creek was 3.4 microg/l in water and 32 microg/g in zooplankton, and at Walter Walker was 13 microg/l in water and 52 microg/g in zooplankton. Although there were differences in concentrations of inorganic elements in water and biota among the three sites, selenium was apparently the only element elevated to concentrations of concern. Effects on survival were more prominent from dietary exposure compared to waterborne exposure. Selenium concentrations of >or=4.6 microg/g in food organisms adversely affected the survival of razorback sucker larvae. The onset of mortality in larvae exposed to food and water from Walter Walker seemed delayed compared to mortality in larvae exposed to food and water from Horsethief, which has been observed in two other studies. Elevated arsenic in one food source seemed to interact with selenium to reduce the toxic effects of selenium. PMID- 12127742 TI - Early oxidative damage in primary cultured trout hepatocytes: a time course study. AB - The aim of this study was to evaluate the influence of the two-step hepatocyte isolation procedure on primary cultured trout (Oncorhynchus mykiss) hepatocytes over time. We characterised the possible changes of a variety of some cellular parameters within the first 24-48 h after seeding. We followed the time dependent changes of these parameters during subsequent culture times in order to see if the cells maintained a differentiated status. Scanning electron microscopy revealed bleb formation and 20% cell damage in freshly isolated hepatocytes. During subsequent culture times the bleb dimension appear to be reduced. Heat shock proteins 70 and 50 (HSP70, HSP50) were induced by hepatocyte isolation. During the first 4 h of culture, the hepatocytes showed a variation in mitochondrial activity, an increase in free radical species (ROS), and a decrease in both glutathione (GSH) content and catalase (CAT) activity; the generation of free radicals led to an increase in the formation of 8-hydroxydeoxyguanosine (8 OHdG) in the DNA. The cells showed detectable ethoxyresorufin-O-deethylase activity after 4 h of culture, which had rapidly increased by the 24th hour. After 24 h, mitochondrial and CAT activity, free radical production, and the content of GSH and 8-OHdG returned to their original levels. P450 activity was retained for at least 48 h after seeding. Our data show that trout hepatocytes suffer significant cell injury as a result of the isolation procedure, but primary cultured cells metabolically recover from this stress after a few hours: they are capable of repairing their damaged surfaces, recovering their antioxidant defences and retaining their ability to repair DNA. Our results also confirm that trout hepatocytes in a primary culture maintain their in vivo-like metabolic activities for 3-8 days. PMID- 12127743 TI - Retrograde signaling by the neurotrophins follows a well-worn trk. AB - The mechanism that allows a neuron to send cues received at its terminal to its cell body and nucleus has proved elusive. However, a recent study by Howe and colleagues indicates that neurotrophin signaling via the trkA receptor requires formation of a signaling endosome containing NGF and trkA. Thus, endocytosis of the neurotrophin-receptor complex is a crucial step in the generation of intracellular signaling platforms required for activation and compartmentalization of signaling events. PMID- 12127744 TI - Sealed with a twist: complexin and the synaptic SNARE complex. AB - Knockout experiments show that complexins are crucial for Ca(2+)-dependent neurotransmitter release. In the recently solved crystal structure of complexin I bound to the synaptic SNARE complex, complexin binds along the groove between the syntaxin and synaptobrevin coils. This structure, together with NMR data, suggests that the role of complexin is to stabilize the SNARE complex as it forms from SNAREs in vesicle and target membranes. PMID- 12127745 TI - Meaningless minis? Mechanisms of neurotransmitter-receptor clustering. AB - Initiation and maintenance of the postsynaptic neurotransmitter-receptor field are important steps during synapse formation and maturation, as they play a determinative role in regulating synaptic strength. However, the mechanisms directing neurotransmitter-receptor clustering and maintenance are poorly understood. Recently, two models explaining glutamate-receptor clustering at the Drosophila neuromuscular junction have been proposed. One model postulates that release of an agent via single vesicle fusion events (minis) is required for the initiation of postsynaptic glutamate-receptor clustering, and that glutamate is not responsible for initiation or maintenance of the postsynaptic receptor field. The other model rules out a role for minis in initiation of clustering, and suggests a role for non-vesicular release of glutamate in receptor-field maintenance. Here, we compare and discuss the data underlying both models. PMID- 12127746 TI - Response: meaningless minis? PMID- 12127747 TI - Response: meaningless minis? PMID- 12127748 TI - Event-related brain dynamics. AB - Event-related potentials (ERPs) provide evidence of a direct link between cognitive events and brain electrical activity in a wide range of cognitive paradigms. It has generally been held that an ERP is the result of a set of discrete stimulus-evoked brain events. A recent study, however, provides new evidence to suggest that some ERP components might be generated by stimulus induced changes in ongoing brain dynamics. This is consistent with views emerging from several neuroscientific fields, suggesting that phase synchronization of ongoing rhythms across different spatio-temporal scales mediates the functional integration necessary to perform higher cognitive tasks. PMID- 12127749 TI - Response: event-related brain dynamics -- unifying brain electrophysiology. PMID- 12127750 TI - Cyclic amplification of protein misfolding: application to prion-related disorders and beyond. AB - Diverse human disorders, including the majority of neurodegenerative diseases, are thought to arise from the misfolding and aggregation of protein. We have recently described a novel technology to amplify cyclically misfolded proteins in vitro. This procedure, named protein misfolding cyclic amplification (PMCA), is conceptually analogous to DNA amplification by PCR and has tremendous implications for research and diagnosis. The PMCA concept has been proved on the amplification of prions implicated in the pathogenesis of transmissible spongiform encephalopathies. In this article we describe the rational behind PMCA and some of the many potential applications of this novel technology. PMID- 12127756 TI - Axonal and presynaptic protein synthesis: new insights into the biology of the neuron. AB - The presence of a local mRNA translation system in axons and terminals was proposed almost 40 years ago. Over the ensuing period, an impressive body of evidence has grown to support this proposal -- yet the nerve cell body is still considered to be the only source of axonal and presynaptic proteins. To dispel this lingering neglect, we now present the wealth of recent observations bearing on this central idea, and consider their impact on our understanding of the biology of the neuron. We demonstrate that extrasomatic translation sites, which are now well recognized in dendrites, are also present in axonal and presynaptic compartments. PMID- 12127757 TI - Cooperation between mglu receptors: a depressing mechanism? AB - Recent findings from the perirhinal cortex have shed new light on the ways in which metabotropic glutamate receptors could be involved in synaptic plasticity, and in particular in long-term depression (LTD) of synaptic transmission. Importantly, these findings have also led to a greater understanding of mechanisms that could regulate mglu-receptor signalling and the ways in which mglu receptors interact with one another. PMID- 12127758 TI - Lipid rafts in neuronal signaling and function. AB - Lipid rafts are plasma membrane microdomains rich in cholesterol and sphingolipids, which provide a particularly ordered lipid environment. Rafts are enriched in glycosylphosphatidylinositol (GPI)-anchored proteins, as well as proteins involved in signal transduction and intracellular trafficking. In neurons, lipid rafts act as platforms for the signal transduction initiated by several classes of neurotrophic factors, including neurotrophins and glial derived neurotrophic factor (GDNF)-family ligands. Emerging evidence also indicates that such rafts are important for neuronal cell adhesion, axon guidance and synaptic transmission. Thus, lipid rafts are structurally unique components of plasma membranes, crucial for neural development and function. PMID- 12127759 TI - An 'oligarchy' rules neural development. AB - Recent reports show that Olig genes, which encode the basic helix-loop-helix Olig transcription factors, are essential for development of oligodendrocytes. Surprisingly, Olig function is also required for formation of somatic motor neurons. These findings alter our views of how the oligodendrocyte lineage is generated and raise further questions about the underlying developmental relationships between neurons and glia. PMID- 12127761 TI - Genetic bases of mental illness -- a cure for stigma? AB - An increased emphasis on biological causes of mental illness has been viewed as having the potential to significantly reduce stigma. From this perspective, the current genetics revolution can be seen as a source of hope. However, some have argued that biological attributions could increase stigma, for example by making the ill person seem 'defective' or 'physically distinct' -- 'almost a different species'. In this paper, I use a multicomponent conceptualization of stigma as a guide in forming hypotheses about the likely impact of genetic attributions on the stigma of mental illness. PMID- 12127760 TI - Moving around in a worm: netrin UNC-6 and circumferential axon guidance in C. elegans. AB - How does an extracellular guidance molecule direct multiple growth cones to different positions? The answer is important for understanding the development of complex neural connections. UNC-6 is a member of the netrin family of guidance proteins. It has phylogenetically conserved domains that mediate its different guidance and branching activities. In the Caenorhabditis elegans embryo, UNC-6 is secreted ventrally and a pattern of circumferential axon tracts develops as pioneer growth cones bearing UNC-5 and UNC-40 receptors are directed towards, or away from, the ventral sources. Following the first migrations, UNC-6 from additional sources allows more complex migration patterns to emerge. In addition, at specific dorsoventral positions, locally restricted extracellular molecules alter growth cone responses to UNC-6, causing circumferentially migrating growth cones to turn and longitudinal nerves to develop. These observations show that extracellular guidance molecules can direct complex arrangements of migrating growth cones in vivo by eliciting different types of responses, by spatially and temporally regulating their expression and by working in concert with other extracellular molecules. PMID- 12127763 TI - The genus Pan: population genetics of an endangered outgroup. PMID- 12127764 TI - Small RNAs in Rickettsia: are they functional? PMID- 12127765 TI - Distinguishing the ORFs from the ELFs: short bacterial genes and the annotation of genomes. AB - A substantial fraction of hypothetical open reading frames (ORFs) in completely sequenced bacterial genomes are short, suggesting that many are not genes but random stretches of DNA. Although it is not feasible to authenticate the coding capacity of all such regions experimentally, comparisons of ORFs in related genomes can expose those that encode functional proteins. PMID- 12127766 TI - Human SNP variability and mutation rate are higher in regions of high recombination. AB - Understanding the co-variation of nucleotide diversity and local recombination rates is important both for the mapping of disease-associated loci and in understanding the causes of sequence evolution. It is known that single nucleotide polymorphisms (SNPs) around protein coding genes show higher diversity in regions of high recombination. Here, we find that this correlation holds for SNPs across the entire human genome, the great majority of which are not near exons or control elements. Contrasting with results from coding regions, we provide evidence that the higher nucleotide diversity in regions of high recombination is most likely due, at least in part, to a higher mutation rate. One possible explanation for this is that recombination is mutagenic. PMID- 12127767 TI - Wnt signaling--20 years and counting. PMID- 12127768 TI - Fission yeast blooms in Kyoto. PMID- 12127774 TI - Metastable epialleles in mammals. AB - There are some mammalian alleles that display the unusual characteristic of variable expressivity in the absence of genetic heterogeneity. It has recently become evident that this is because the activity of these alleles is dependent on their epigenetic state. Interestingly, the epigenetic state is somewhat labile, resulting in phenotypic mosaicism between cells (variegation) and also between individuals (variable expressivity). The establishment of the epigenetic state occurs during early embryogenesis and is a probabilistic event that is influenced by whether the allele is carried on the paternal or maternal alleles. In addition, the epigenetic state determines whether these alleles are dominant. We propose that mammalian alleles with such characteristics should be termed metastable epialleles to distinguish them from traditional alleles. At this stage, it is unclear how common these alleles are, but an appreciation of their existence will aid in their identification. PMID- 12127775 TI - X-chromosome inactivation: closing in on proteins that bind Xist RNA. AB - X inactivation is the developmentally regulated silencing of a single X chromosome in XX female mammals. In recent years, the Xist gene has been revealed as the master regulatory switch controlling this process. Parental imprinting and/or counting mechanisms ensure that Xist is expressed only on the inactive X chromosome. Chromosome silencing then results from the accumulation of the Xist RNA silencing signal, in cis, over the entire length of the X chromosome. A key issue has been to identify the factors that interact with Xist RNA to initiate heritable gene silencing. This review discusses recent progress that has put this goal in sight. PMID- 12127777 TI - Using mouse models to dissect the genetics of obesity. AB - Mice have proved to be powerful models for understanding obesity in humans and farm animals. Single-gene mutants and genetically modified mice have been used successfully to discover genes and pathways that can regulate body weight. For polygenic obesity, the most common pattern of inheritance, many quantitative trait loci (QTLs) have been mapped in crosses between selected and inbred mouse lines. Most QTL effects are additive, and diet, age and gender modify the genetic effects. Congenic, recombinant inbred, advanced intercross, and chromosome substitution strains are needed to map QTLs finely, to identify the genes underlying the traits, and to examine interactions between them. PMID- 12127776 TI - Imprinted genes: identification by chromosome rearrangements and post-genomic strategies. AB - Imprinted genes are differentially expressed from the maternally and paternally inherited alleles. Accordingly, inheritance of both copies of an imprinted chromosome or region from a single parent leads to the mis-expression of the imprinted genes present in the selected region. Strains of mice with reciprocal and Robertsonian chromosomal translocations or mice with engineered chromosomal rearrangements can be used to produce progeny where both copies of a chromosomal region are inherited from one parent. In combination with systematic differential expression and methylation-based approaches, these mice can be used to identify novel imprinted genes. Advances in genome sequencing and computer-based technologies have facilitated this approach to finding imprinted genes. PMID- 12127780 TI - Quasispecies, error catastrophe, and the antiviral activity of ribavirin. AB - Ribavirin is the first synthetic, broad-spectrum antiviral nucleoside. Despite its more than 30 year history, the mechanism of action of this compound remains unclear and somewhat controversial. Recent data suggest the possibility that the activity of ribavirin against RNA viruses is a reflection of incorporation of ribavirin into the viral genome. Because ribavirin incorporation is not specific, this event leads to lethal mutagenesis of the virus population. The data supporting this new proposal for the mechanism of action of ribavirin are reviewed herein. In addition, we discuss briefly the challenges that remain for development of lethal mutagenesis as an effective antiviral strategy. PMID- 12127781 TI - Identification of the bZIP and Rta homologues in the genome of rhesus monkey rhadinovirus. AB - Rhesus monkey rhadinovirus (RRV) is a gamma2-herpesvirus identified from the peripheral blood mononuclear cells of rhesus macaques (Macaca mulatta). Serologic studies suggested that the infections of RRV are prevalent among rhesus monkeys. RRV can be efficiently propagated and grows to a high titer in cultured rhesus monkey fibroblasts, thus providing a valuable model to study the rhadinovirus replication. By comparative genomic studies, here we describe the identification of two potential transcriptional factors encoded by RRV, designated as R-Rta and R-bZIP. Initial functional characterization of these products suggested that R Rta is a potent transcriptional activator and R-bZIP forms homodimers in vivo. Viral homologues of R-Rta and R-bZIP, previously identified from other rhadinoviruses, have been implicated in serving as molecular switches in lytic replication. PMID- 12127782 TI - Species tropism of chimeric SHIV clones containing HIV-1 subtype-A and subtype-E envelope genes. AB - To analyze HIV-1 genes in a nonhuman primate model for lentivirus infection and AIDS, recombinant SIV/HIV-1 (SHIV) clones were constructed from two HIV-1 subtype A isolates (HIV-1(SF170) and HIV-1(Q23-17) from individuals in Africa) and two HIV-1 subtype-E isolates (HIV-1(9466) and HIV-1(CAR402) from AIDS patients in Thailand and Africa), respectively. These four SHIV clones, designated SHIV-A 170, SHIV-A-Q23, SHIV-9466.33, and SHIV-E-CAR, contain envelope (env) genes from the subtype-A or -E viruses. Interestingly, SHIV-A-170, SHIV-A-Q23, and SHIV 9466.33 were restricted for replication in cultures of macaque lymphoid cells, whereas SHIV-E-CAR replicated efficiently in these cells. Additional studies to define the block to replication in macaque cells were focused on the subtype-E clone SHIV-9466.33. A SHIV intragenic env clone, containing sequence-encompassing V1/V2 regions of HIV-1(CAR402) and V3/V4/V5 regions of SHIV-9466.33, infected and replicated in macaque lymphoid cells. These results indicated that the sequence encompassing V1/V2 region of HIV-1(9466) was responsible for the block of the SHIV-9466.33 replication in macaque cells. Analysis of viral DNA in acutely infected macaque cells revealed that SHIV-9466.33 was blocked at a step at/or before viral DNA synthesis, presumably during the process of virion entry into cells. In a fluorescence-based cell-cell fusion assay, fusion pore formation readily took place in cocultures of cells expressing the SHIV-9466.33 env glycoprotein with macaque T-lymphoid cells. Taken together, these results demonstrated that the block of SHIV-9466.33 replication in macaque cells is at an early step after fusion pore formation but before reverse transcription. PMID- 12127783 TI - Subdomain specific functions of the RNA polymerase region of poliovirus 3CD polypeptide. AB - The 3D polymerase domain of the poliovirus 3CD polypeptide plays a role in modulating its RNA binding and protein processing activities, even though the proteinase catalytic site and RNA binding determinants appear to reside within the 3C(pro) portion of the molecule. In this study, we have generated recombinant 3CD polypeptides that contain chimeric 3D polymerase domains representing suballelic sequence exchanges between poliovirus type 1 (PV1) and coxsackievirus B3 (CVB3) to determine which portions of the 3D domain are responsible for influencing these activities. By utilizing these recombinant protein chimeras in protein processing and RNA binding studies in vitro, we have generated data suggesting the presence of separate subdomains within the polymerase domain of 3CD that may independently modulate its RNA binding and protein processing activities. In predicting where our sequence exchanges map by utilizing the previously published three-dimensional structure of the PV1 3D polymerase, we present evidence that sequences contained within the RNA recognition motif of the polymerase are critical for 3CD function in recognizing the 5' RNA cloverleaf. Furthermore, our protein processing data indicate that at least some of the substrate recognition and processing determinants within the 3D domain of 3CD are separate and distinct from the RNA binding determinants in this domain. PMID- 12127784 TI - Mapping of B-cell epitopes of hemagglutinin protein of rinderpest virus. AB - Monoclonal antibodies (mAbs) against secreted hemagglutinin (H) protein of rinderpest virus (RPV) expressed by a recombinant baculovirus were generated to characterize the antigenic sites on H protein and regions of functional significance. Three of the mAbs displayed hemagglutination inhibition activity and these mAbs were unable to neutralize virus infectivity. Western immunoblot analysis of overlapping deletion mutants indicated that three mAbs recognize antigenic regions at the extreme carboxy terminus (between amino acids 569 and 609) and the fourth mAb between amino acids 512 and 568. Using synthetic peptides, aa 569-577 and 575-583 were identified as the epitopes for E2G4 and D2F4, respectively. The epitopic domains of A12A9 and E2B6 mAbs were mapped to regions encompassing aa 527-554 and 588-609. Two epitopes spanning the extreme carboxy terminal region of aa 573 to 587 and 588 to 609 were shown to be immunodominant employing a competitive ELISA with polyclonal sera form vaccinated cattle. The D2F4 mAb which recognizes a unique epitope on RPV-H is not present on the closely related peste des petits ruminant virus HN protein and this mAb could serve as a tool in the seromonitoring program after rinderpest vaccination. PMID- 12127785 TI - The structure of P4 procapsids produced by coexpression of capsid and external scaffolding proteins. AB - The double-stranded DNA bacteriophage P4 has a T = 4 icosahedral arrangement of the gpN capsid protein derived from the P2 helper phage. The precursor procapsids in addition contain an external scaffold made up of the P4-encoded Sid protein. High yields of pure P4 procapsids have been obtained by coexpressing the gpN and Sid proteins from a chimeric plasmid. Biochemical measurements show that the ratio of gpN to Sid in the procapsids is 2:1, corresponding to 120 copies of Sid per procapsid particle. A reconstruction of the P4 procapsid, made from 213 particle images to an effective resolution of about 21 A, greatly improves on the previously determined P4 procapsid structures. The structure shows a T = 4 capsid shell and a unique tandem arrangement of 120 copies of chilli-shaped Sid monomers, which form trimers and dimers on the procapsid surface. PMID- 12127786 TI - Functional replacement of the tobacco rattle virus cysteine-rich protein by pathogenicity proteins from unrelated plant viruses. AB - Mutation of the 16K gene encoded by RNA1 of Tobacco rattle virus (TRV) greatly reduced the levels of viral RNA that accumulated in both infected protoplasts and plants, showing that the 16K cysteine-rich protein (CRP) is required for efficient multiplication of TRV. Overexpression of the 16K protein, either from an additional copy of the gene carried on TRV RNA2 or from a PVX vector, led to an increase in the severity of disease symptoms, suggesting that the protein has a role in the pathogenicity of the virus. Mutation of the 16K gene could be overcome by expression from RNA2 of the Cucumber mosaic virus 2b gene, the Soil borne wheat mosaic virus 19K gene, or the Barley stripe mosaic virus gammab gene, indicating that the proteins encoded by these diverse genes may have similar functions. One known function of the CMV 2b gene is as a suppressor of posttranscriptional gene silencing, suggesting that the TRV 16K protein may also possess this activity. PMID- 12127787 TI - Human papillomavirus type 11 alters the transcription and expression of loricrin, the major cell envelope protein. AB - Human papillomavirus (HPV) does not induce lysis of infected keratinocytes, and the exact mechanisms of viral escape are not known. As keratinocytes differentiate, the cornified cell envelope (CCE) develops, providing a protective barrier to the host. We previously showed that the normally durable CCE in HPV 11 infected epithelium is fragile compared to CCEs in healthy epithelium. In this study, we examined uninfected and HPV 11-infected human genital epithelium for expression of loricrin, the major CCE protein in healthy epidermis. In HPV 11 infected human genital epithelium, detection of loricrin was reduced in immunoelectron microscopic and immunoblot assays, suggesting that loricrin incorporation into the CCE was reduced or that loricrin synthesis was reduced. Loricrin detection was reduced in immunohistochemical assays in areas of high viral replication. Mathematical modeling by least squares was performed using the amino acid composition of highly purified CCE fragments, confirming that loricrin was markedly reduced and that the small proline-rich proteins and cytokeratins were increased in those derived from HPV 11-infected epithelium compared to healthy genital epithelium. In RNase protection and RT-PCR assays, loricrin transcripts were markedly reduced in HPV 11-infected epithelium compared to uninfected epithelium. Loricrin transcripts were detectable by RNA in situ analysis in isolated cells of HPV 11-infected epithelium, but were absent in large regions of epithelium. We conclude that HPV 11 infection reduces transcription of the loricrin gene and that this leads to a reduction in loricrin incorporation into the CCE. Further studies will examine the effects of specific HPV gene products on transcription of loricrin and other CCE components, as it is likely that viral egress from the infected keratinocyte depends on these effects. PMID- 12127788 TI - The product of the UL12.5 gene of herpes simplex virus type 1 is not essential for lytic viral growth and is not specifically associated with capsids. AB - The herpes simplex virus type 1 UL12 gene encodes a pH-dependent deoxyribonuclease termed alkaline nuclease. An N-terminally truncated version of the UL12 gene, called UL12.5, was shown to be translated independently from a subgenic mRNA which shares its 3' terminus with the full-length UL12 mRNA. We showed previously that the UL12.5 gene product cannot compensate for the absence of the full-length UL12 gene product (R. Martinez, L. Shao, J. C. Bronstein, P. C. Weber, and S. K. Weller, 1996, Virology 215, 152-164); however, it was not known whether UL12.5 itself performs an essential function during lytic viral growth. In this article the initiation codon for the UL12.5 gene product was mapped and altered to create a gene no longer capable of producing UL12.5. This mutation was introduced into the viral genome to create a virus which was capable of producing full-length UL12 but not UL12.5. The growth properties of this virus indicate that UL12.5 is not essential for viral growth in culture. UL12.5 was previously reported to represent a capsid-associated form of alkaline nuclease (J. C. Bronstein, S. K. Weller, and P. C. Weber, 1997, J. Virol. 71, 3039-3047). Sucrose sedimentation analysis of capsids from cells infected with wild-type or mutant viruses indicates that both UL12 and UL12.5 are found in fractions from across the sucrose gradient which do not always correlate with the presence of viral capsids. Furthermore, UL12.5 is found in fractions across the gradient even in cells infected under conditions in which no capsids are formed. These results indicate that UL12.5 does not specifically associate with viral capsids. Taken together, these data indicate that UL12.5 is not likely to play an important role in lytic viral infection. PMID- 12127789 TI - Functional properties of the predicted helicase of porcine reproductive and respiratory syndrome virus. AB - Porcine reproductive and respiratory syndrome virus (PRRSV) is a member of the positive-strand RNA virus family Arteriviridae. Although considerable research has focused on this important pathogen, little is known about the function of most PRRSV proteins. To examine characteristics of putative nonstructural proteins (nsp) encoded in ORF1b, which have been identified by nucleotide similarity to domains of equine arteritis virus, defined genomic regions were cloned and expressed in the pRSET expression system. One region, nsp10, encoded a protein with a putative helicase domain and was further examined for functional helicase-like activities. PRRSV nsp10 was found to possess a thermolabile and pH sensitive NTPase activity that was modulated by polynucleotides and to unwind dsRNA in a 5' to 3' polarity. These results provide the first evidence of the functional properties of PRRSV helicase and further support the finding that nidovirus helicases possess properties that distinguish them from other viral helicases. PMID- 12127790 TI - Differential production of cytokines and activation of NF-kappaB in HPV transformed keratinocytes. AB - We have proposed that chronic infection of keratinocytes by HPV modifies the expression of potentially important cytokines by interfering with the NF-kappaB signal pathway. We evaluated the constitutive and IL-1beta-induced expression of GM-CSF and TNF-alpha and the expression/activity of NF-kappaB in HPV+ and HPV- cell lines. Despite the enhanced expression of the functional components of the NF-kappaB signaling pathway in HPV+ cell lines by a mechanism implicating the HPV oncoprotein E6, the constitutive activity of NF-kappaB and the expression of GM CSF/TNF-alpha were significantly reduced relative to the HPV- cell line and normal keratinocytes. In contrast, we observed a superactivation of NF-kappaB activity after IL-1beta stimulation, a strong and transient induction of GM CSF/TNF-alpha mRNA, but undetectable levels of secreted proteins in HPV+ cell lines. Our data demonstrate that E6 modulates the NF-kappaB signaling pathway and suggest that other HPV proteins also interfere with GM-CSF/TNF-alpha expression by transcriptional and/or posttranscriptional mechanisms. PMID- 12127791 TI - Lyssavirus P gene characterisation provides insights into the phylogeny of the genus and identifies structural similarities and diversity within the encoded phosphoprotein. AB - A comprehensive phylogenetic analysis of the Lyssavirus genus, employing P gene sequences from 128 isolates recovered globally, is presented. While confirming prior suggestions of the genetic distinctness of the Australian bat lyssaviruses, these data also revealed a clear division within the rabies virus clade (Genotype 1) between globally distributed viruses circulating predominantly in canid species (subgroup 1-1), and American strains harbored by both chiropteran and terrestrial hosts (subgroup 1-2). Nucleotide substitution patterns within the P locus suggested differential selection operating along the length of the open reading frame (ORF) between rabies viruses of these two subgroups. Comparison of the deduced primary sequences of the encoded phosphoproteins of all isolates provided insights into the product's structural organization. Two conserved (CD1,2) and two variable (VD1,2) domains were evident; high variation in the VD2 region was reflected by differences in hydropathic profiles. Only two of five serine residues reported to function as phosphate acceptors in the P protein of the rabies challenge virus standard (CVS) strain were absolutely conserved; similarly, out of four internal methionines reported to direct internal translation initiation of the CVS strain to produce N-terminally truncated P proteins, only Met(20) was universally retained. In contrast, two sequence motifs, one believed to confer binding to the cytoplasmic dynein light chain LC8, and a lysine-rich sequence probably contributing to N protein binding, were conserved throughout the genus. Most rabies viruses of the carnivora (1-1) contain a potential C ORF in alternate frame to that of P, a feature limited or absent in most other isolates of the genus, an observation interpreted with consideration to the predicted course of lyssavirus evolution. PMID- 12127792 TI - Protection by intranasal immunization of a nef-deleted, nonpathogenic SHIV against intravaginal challenge with a heterologous pathogenic SHIV. AB - An effective vaccine against sexual transmission of human immunodeficiency virus (HIV) should elicit both systemic and mucosal immune responses. In this study, to examine the possibility of using an attenuated virus for mucosal immunization, four female macaques were intranasally or intravenously administered with a chimeric simian-human immunodeficiency virus with a deleted nef gene (SHIV-dn). Although all the monkeys had anti-HIV-1 antibodies with neutralizing activity in the plasma, the intranasally immunized monkeys had much higher levels of HIV-1 Env-specific IgG and IgA antibodies in mucosal secretions compared with the intravenously immunized monkeys. Moreover, three of four intranasally immunized monkeys were completely protected from intravaginal challenge with a pathogenic virus, SHIV-89.6P, whereas only one intravenously immunized monkey was protected. Thus, intranasal immunization of an attenuated virus can induce the protective efficacy against intravaginal infection. PMID- 12127794 TI - Cloning of a gene encoding a unique haemolysin from Klebsiella pneumoniae and its potential use as a species-specific gene probe. AB - A gene, designated khe, that encodes a haemolysin of Klebsiella pneumoniae CMC-1 has been cloned and sequenced. When expressed in Escherichia coli, a unique peptide of approximately 20kDa was identified. Nucleotide sequence analysis predicted a single open reading frame (ORF) of 486bp encoding a 162 amino acid polypeptide with an estimated pI of 6.77. No extensive sequence homology could be identified between khe and any reported sequence at either the nucleotide or amino acid level. Furthermore, DNA hybridizations under high stringency conditions failed to show any cross hybridizations to several bacteria including K. oxytoca, K. planticola, K. terrigena and K. ornithinolytica. These data indicate that we have cloned a unique gene, which is highly conserved among tested K. pneumoniae isolates. PMID- 12127793 TI - Effect of proteolytic processing at two distinct sites on shape and aggregation of an anchorless fusion protein of human respiratory syncytial virus and fate of the intervening segment. AB - We have examined the consequences of cleaving the fusion glycoprotein (F) of human respiratory syncytial virus (HRSV) at two distinct furin-recognition sites. Purified anchorless F is a mixture of unaggregated cone-shaped molecules and rosettes of lollipop-shaped spikes. The unaggregated molecules contain a proportion of uncleaved F0 and an intermediate, F(delta1-109), cleaved only at site I, residues 106-109. Inhibition of cleavage at site I, by two amino acid changes (R108N/R109N), reduces the proportion of aggregated molecules with a concomitant increase in the amount of unprocessed F0. Inhibition of cleavage at site II, residues 131-136, by deletion of four amino acids (delta131-134), abrogates aggregation of anchorless F and all molecules are seen as individual cone-shaped rods. In vitro cleavage of anchorless F, or mutant delta131-134, with trypsin at 4, 20, or 37 degrees C, under conditions in which cleavage at site II is complete in all molecules, leads to their aggregation in rosettes of lollipop shaped spikes. Thus, cleavage at site II is required for the structural changes in anchorless F that lead to changes in shape and to aggregation. The segment between sites I and II, residues 110-136, is not associated with anchorless F in the supernatant of infected cell cultures, indicating that it is released from the processed protein when cleavage at sites I and II is completed. PMID- 12127795 TI - Previous infection with the nematode Nippostrongylus brasiliensis alters the immune specific response against Chlamydophila abortus infection. AB - An experimental mouse model to analyze the interaction between the immune responses elicited following infection with Nippostrongylus brasiliensis and Chlamydophila abortus has been established. Mice infected with C. abortus 7 days after N. brasiliensis showed an increased bacterial multiplication in spleen and liver compared to bacteria-alone infected mice. However the morbidity of these mice, expressed as weight loss, was significantly lower. Analysis of the immune responses elicited showed that spleen from co-infected mice had reduced IFN-gamma production in response to C. abortus antigen. The bias towards a type 2 response in co-infected mice was confirmed by an increase in the production of IL-4 and in the lower ratio IgG2a/IgG1. In pregnant mice co-infection caused a delay in the time of abortion and an increased systemic susceptibility to C. abortus infection. PMID- 12127796 TI - Expression of green fluorescent protein in Rickettsia conorii. AB - Rickettsiae are obligate intracellular class III pathogens for which genetic manipulation has only recently been shown to be feasible. Such experiments were restricted to the typhus group rickettsiae, namely R. typhi and R. prowazekii. Here we report the first genetic manipulation of Rickettsia conorii, the bacterial agent responsible for the Mediterranean spotted fever. A gene encoding a variant of the green fluorescent protein under the control of the sterically repressed promoter (srp) from E. coli was integrated into the genome of this bacteria and detected by FACS analysis. PMID- 12127797 TI - Isolation of a putative laminin binding protein from Streptococcus anginosus. AB - Viridans streptococci, including Streptococcus anginosus, are a common cause of infective endocarditis in humans. Adherence mechanisms involved in colonization of non-diseased native valves (present in 40% of native valve endocarditis) are unknown. We have previously shown that an endocarditis isolate of S. anginosus adheres to exposed basement membrane of human and porcine valve tissue in a laminin dependent manner. We now describe the partial purification of an 80 kDa putative laminin binding protein (PLBP) by biochemical methods. Amino acid sequence of PLBP peptides is similar to substrate binding proteins of ABC transporters in other Gram-positive cocci. PMID- 12127798 TI - Characterization and molecular cloning of a glutamyl endopeptidase from Staphylococcus epidermidis. AB - A novel extracellular endopeptidase, designated GluSE, was purified from Staphylococcus epidermidis ATCC 14990 cultured by the dialysis membrane technique, and the structural gene (gseA) was cloned. GluSE was a 27kDa, glutamic acid-specific protease, and the optimal pH was 8.0. The proteolytic activity was specifically inhibited with diisopropyl fluorophosphate, indicating that it is a serine protease. The gseA encoded a single polypeptide of 282 amino acids with a deduced molecular weight of 30,809, in which the first 19 N-terminal amino acids completely matched the deduced sequence starting at Val-67, suggesting that GluSE is synthesized with a propeptide. The amino acid sequence of GluSE exhibited 50.5% identity to Staphylococcus aureus V8-protease (GluV8). Although GluSE lacks a C-terminal 12 repeats of the PBN/PBZ tripeptide of GluV8, a catalytic triad of His-117, Asp-159 and Ser-235 was conserved in GluSE. Southern hybridization analysis revealed that gseA exists as a single copy on the chromosomal DNA. The finding that production of GluSE was obviously observed in the adherent culture conditions of the dialysis membrane technique, but not in the planktonic culture conditions, strongly suggests that GluSE could be involved in an important etiologic process in S.epidermidis infection leading to multiple tissue damages. PMID- 12127799 TI - Roles of gamma interferon and tumor necrosis factor-alpha in shiga toxin lethality. AB - Shiga toxins (Stxs) have been specifically implicated as a causal factor of hemolytic uremic syndrome and acute encephalopathy. The first step of Stx-induced brain damage is considered to injure endothelial cells cooperating with tumor necrosis factor-alpha (TNF-alpha). Gamma interferon (IFN-gamma) is one of the proinflammatory cytokines as well as TNF-alpha is critical in activation of endothelial cells. Therefore we focused on the possibility of IFN-gamma-mediated lethality of Stx1 or Stx2 in mice. All of mice died within 3-4 days after injection with 400 ng of Stx1 and 37.5% of mice, which had been injected with 133 ng, survived. In contrast, a lethal dose of Stx2 was 40 times lower than that of Stx1. When mice were given 400 ng of Stx1 or 10 ng of Stx2, IFN-gamma mRNA was detected in the spleens 24h after injection. Moreover, when mice were injected with 133 ng of Stx1 or 3.3 ng of Stx2, survival rates of IFN-gamma-deficient mice and TNF-alpha-deficient mice were significantly higher than that of wild-type mice. The present study using cytokine-gene knockout mice directly demonstrated that not only TNF-alpha but also IFN-gamma is involved in lethality of Stx1 and Stx2. PMID- 12127800 TI - Induction of antinociception and increased met-enkephalin plasma levels by cyclosporine and morphine in rats: implications of the combined use of cyclosporine and morphine and acute posttransplant neuropsychosis. AB - BACKGROUND: Cyclosporine A (CsA) and morphine have neurotoxic and psychiatric side effects, respectively. Endogenous opiatelike peptides can elicit a number of behavioral responses that mimic the symptoms of psychiatric illness. The purpose of this study was to quantitiate the changes of Met-enkephalin (ME) and beta endorphin (BE) after administration of CsA and morphine in surgery and to assess the antinociceptive effect. PATIENTS AND MATERIALS: Pain sensitivity, an antinociceptive indicator in rats, was determined with the hotplate test. Plasma ME and BE levels were measured with radioimmunoassays. RESULTS: In normal unoperated rats, CsA induced a profound analgesic effect concomitant with an increased plasma ME level on day 1. Morphine produced an analgesic effect on days 1 and 2, with decreased ME levels on days 2 and 3. Coadministration of CsA and morphine prolonged the analgesia from days 1 to 4 and increased the plasma ME level on day 1. No change in plasma BE level was found. In surgically operated rats, CsA induced an analgesic effect and higher ME levels than those in unoperated rats. Interestingly, the combined use of CsA and morphine prolonged the analgesia and increased plasma ME levels from days 1 to 4, with no significant change in plasma BE levels. CONCLUSIONS: Our results showed that CsA can induce antinociception and increase plasma ME levels. This induction can be potentiated by the addition of morphine. Acute neuropsychiatric manifestations in the early posttransplant period might, therefore, be due to induction of ME after coadministration of CsA and morphine. PMID- 12127801 TI - NF-kappaB inhibition enhances peroxynitrite-induced enterocyte apoptosis. AB - BACKGROUND: Sustained overproduction of nitric oxide and peroxynitrite (ONOO(-)) in conditions such as necrotizing enterocolitis and inflammatory bowel disease may promote gut barrier failure by inducing enterocyte apoptosis. NF-kappaB is upregulated in the gut during inflammation and, in addition to its proinflammatory effects, may upregulate protective or antiapoptotic factors such as inhibitor of apoptosis proteins (IAPs). We have previously demonstrated that NF-kappaB inhibition increases cytokine-induced enterocyte apoptosis; however, the effect of NF-kappaB on ONOO(-)-induced enterocyte apoptosis is unknown. MATERIALS AND METHODS: Rat intestinal epithelial cells (IEC-6) were transfected with the adenoviral vector AdIkappaB or AdlacZ. AdIkappaB contains a mutated form of IkappaB which functions as a superrepressor of NF-kappaB. Cells were then treated with 50 microM ONOO(-) or decomposed ONOO(-). Apoptosis was then determined by flow cytometry with annexin V-FITC and propidium iodide staining. Caspase activation and IAP, Bcl-2, Bad, and Bax expression were examined using Western blot analysis, and NF-kappaB activation was determined via electrophoretic mobility shift assay (EMSA). RESULTS: Inhibition of NF-kappaB with AdIkappaB significantly enhanced ONOO(-)-induced apoptosis in IEC-6 cells. ONOO(-) treatment did not activate NF-kappaB in IEC-6 cells as determined by EMSA. There was no difference in IAP, Bcl-2, Bad, and Bax expression between nontransfected, AdlacZ-transfected, and AdIkappaB-transfected cells. Baseline procaspase 3 activation was increased in AdIkappaB-transfected cells. CONCLUSIONS: NF-kappaB inhibition enhances ONOO(-)-induced enterocyte apoptosis, suggesting that NF-kappaB upregulates a protective factor. This protective factor does not appear to be an IAP or Bcl-2 family member and may be expressed constitutively, since ONOO(-) did not activate NF-kappaB over baseline levels of activation. PMID- 12127802 TI - Covered stent exclusion of dialysis access pseudoaneurysms. AB - BACKGROUND: The traditional repair of hemodialysis graft pseudoaneurysms has been to surgically replace that segment of involved PTFE graft material or autogenous vein. We report a novel approach to these lesions, employing a covered stent (Wallgraft) for exclusion of arteriovenous graft (AVG) and arteriovenous fistula (AVF) pseudoaneurysms. METHODS: Ten patients with AVG or AVF pseudoaneurysms were treated endoluminally by covered stent exclusion. Wallgraft implantations were performed in the operating room with interventional capabilities under local anesthesia through a percutaneous access. Follow-up included physical examination at 2 weeks and duplex ultrasound of AVG/ AVF at 6 months after surgery. RESULTS: Ten patients with pseudoaneurysmal degeneration of their AVG/AVF were identified. The mean diameter of the pseudoaneurysmal segment was 3 cm (range, 1.5-5 cm). Immediately following covered stent implantation all the patients had palpable pulses in the pseudoaneurysms despite adequate coverage by angiography. At the 2 week follow-up visit all had lost the palpable pseudoaneurysm pulsation while the AVGs remained functional in nine patients. One patient had early thrombosis of the AVG. The follow-up duplex scans at 6 months showed complete exclusion of the pseudoaneurysms in seven patients. Two patients had thrombosis of their dialysis access, at 3 weeks (n = 1) and 3 months (n = 1) post-implantation. CONCLUSION: Endovascular covered stent exclusion of AV dialysis access pseudoaneurysms is safe and technically feasible in eliminating flow through dialysis access pseudoaneurysms and represents a novel and simple approach to this common problem, prolonging the functional life of the access site. PMID- 12127803 TI - Risk factors for conversion of laparoscopic to open cholecystectomy. AB - BACKGROUND: Laparoscopic cholecystectomy (LC) has become the treatment of choice for symptomatic gallstones; however conversion to open cholecystectomy (OC) remains a possibility. Unfortunately, preoperative factors indicating risk of conversion are unclear. Therefore, we aimed to identify risk factors associated with conversion of LC to OC. PATIENTS AND MATERIALS: Records of 564 patients undergoing LC in 1995 and 1996 were reviewed. Patients were assigned to one of two groups: (1) acute cholecystitis defined by the presence of gallstones, fever, leukocyte count >10(4), and inflammation on ultrasound or histology; (2) chronic cholecystitis that included all other symptomatic patients. Demographics, history, and physical, laboratory, and radiology data, operative note, and the pathology report were reviewed. RESULTS: 161 of 564 patients, had acute and 403 patients had chronic cholecystitis; 16 acute cholecystitis patients (10%) were converted from LC to OC and 17 chronic cholecystitis patients (4%) had LC converted to OC. Patients having open conversion were significantly older, had greater prevalence of cardiovascular disease, and were more likely to be males. LC conversion to OC in acute cholecystitis patients was associated with a greater leukocyte count; in gangrenous cholecystitis patients, 29% had open conversion. CONCLUSIONS: Importantly, these risk factors-older men, presence of cardiovascular disease, male gender, acute cholecystitis, and severe inflammation are determined preoperatively, permitting the surgeon to better inform patients about the conversion risk from LC to OC. While acute cholecystitis was associated with more than a twofold increased conversion rate, only 10% of these patients could not be completed laparoscopically. Therefore, acute cholecystitis alone should not preclude an attempt at laparoscopic cholecystectomy. PMID- 12127804 TI - Immune-enhancing enteral diet selectively augments ileal blood flow in the rat. AB - BACKGROUND: Clinical studies show that immune-enhancing enteral diets (IED; with L-arginine, fish oil, and RNA fragments) decrease the rate of sepsis and shorten the length of hospital stay after the start of enteral feeding. These beneficial effects are dependent on the route of administration (enteral vs parenteral) and on the nutrient composition (IED vs standard diets). Gut exposure to an IED seems to preserve and/or augment intestinal mucosal immunity. However, nutrient absorption stimulates gut blood flow in a nutrient-specific manner (i.e., postprandial hyperemia). We hypothesized that an IED would initiate a different pattern of whole organ blood flow compared to a standard diet. This suggests that a mechanism for the protective effect of IED might be the preferential augmentation of gut blood flow to gut-associated lymphoid tissue (GALT) or mucosa associated lymphoid tissue (MALT). METHODS: Male Sprague-Dawley rats (200-225 g) were anesthetized and cannulated for colorimetric microsphere determination of blood flow distribution (with the phantom organ technique). Animals received gastric gavage (2 ml) of an IED (Impact; Novartis) or an isocaloric, isonitrogenous control diet (Boost; Mead-Johnson). Blood flow to the antrum, duodenum, jejunum, ileum, colon, liver, kidneys, and spleen was determined at baseline and 30, 60, 90, and 120 min after gavage. RESULTS: Baseline blood flows to the left and right kidneys were within 10%, indicating the technical integrity of the microsphere technique and assay. Control diet augmented blood flow compared to IED in the antrum, duodenum, jejunum, and spleen. Conversely, IED gavage stimulated a delayed and sustained hyperemic response in the ileum. IED also increased hepatic blood flow early (30 min). IED increased blood glucose levels compared to control diet at 30, 60, and 90 min, suggesting enhanced nutrient absorption. CONCLUSIONS: These data show that blood flow distribution depends on nutrient composition and that IED preferentially augments blood flow to the ileum. Since the terminal jejunum and ileum contain much of the GALT, our data suggest that a mechanism for enterally stimulated mucosal immunity involves selective perfusion of the terminal ileum during IED nutrient absorption. PMID- 12127806 TI - A 24-h pneumoperitoneum leads to multiple organ impairment in a porcine model. AB - BACKGROUND: An intra-abdominal pressure (IAP) of 15 mm Hg reduces intestinal organ perfusion in humans and animals, but it is unknown whether this results in organ damage. The purpose of this study was to evaluate if an IAP of 15 mm Hg lasting for 24 h in a porcine model will lead to morphologic impairment of intestinal and adjacent organs. METHODS: We examined 12 intubated and anesthetized domestic pigs (51.8 +/- 4.4 kg). Using CO2 pneumoperitoneum, the IAP was raised to 15 mm Hg (study group, n = 6) for an investigation period of 24 h. In the control group, the IAP remained unchanged. Investigated parameters were cardiac output (CO), peak inspiratory pressure (PIP), and urine output (UO), as well as serum creatinine, alanine aminotransferase (ALT), lactate, lipase, and alkaline phosphatase (AP). Additionally, histopathological examinations were performed.Results. In comparison to the control, CO did not change, while UO decreased significantly by 59% and PIP increased significantly to more than 30 mbar. Serum ALT and AP increased significantly while there was no change in creatinine, lactate, and lipase. Histopathologically, low-grade liver necrosis (12% of liver lobuli), low-grade proximal tubular epithelial necrosis, and low bowel mucosal damage were observed.Conclusion. In this porcine model, an IAP of 15 mm Hg lasting for 24 h was found to result in functional and morphologic impairment of lungs, liver, kidneys, and bowel. These results imply that a prolonged IAP of 15 mm Hg predisposes to multiorgan dysfunction and that a safe duration of increased IAP still has to be determined. PMID- 12127805 TI - Increased nitric oxide production in hepatocytes is involved in liver dysfunction following obstructive jaundice. AB - BACKGROUND: Obstructive jaundice damages critical functions in the liver. However, the mechanisms involved in hepatic dysfunction are obscure. Nitric oxide is implicated in liver injury under various pathological conditions. We previously reported that proinflammatory cytokine interleukin-1beta (IL-1beta) stimulated the production of nitric oxide in hepatocytes, which was associated with mitochondrial dysfunction. Studies were performed to examine whether obstructive jaundice influences the production of nitric oxide in hepatocytes and alters hepatic energy metabolism. MATERIAL AND METHODS: Hepatocytes were isolated and cultured from a rat model of obstructive jaundice or sham control. Nitric oxide production, ATP content, and ketone body ratio (acetoacetate/beta hydroxybutyrate; KBR) were compared between the two groups in the presence of IL 1beta. RESULTS: Hepatocytes obtained from obstructive jaundice rats markedly increased the levels of nitric oxide production stimulated by IL-1beta compared with those from sham control. Western blot analysis revealed that the enhancement of nitric oxide production was a posttranslational event, since protein levels of inducible nitric oxide synthase (NOS) were unchanged between the two groups. IL 1beta decreased cellular ATP content in obstructive jaundice but not in sham control. Further, the KBR, which is a marker of mitochondrial redox state, was lower in obstructive jaundice than in sham control. Addition of N(G)-monomethyl-L arginine, an inhibitor of NOS, abolished the decreases in ATP content and KBR as well as the nitric oxide production. CONCLUSIONS: These results indicate that a priming of nitric oxide production following obstructive jaundice is associated with the alteration of hepatic energy metabolism in part through mitochondrial dysfunction. Regulation of nitric oxide production may be a useful therapy for preventing liver dysfunction in obstructive jaundice. PMID- 12127807 TI - Effect of nitric oxide in ischemia/reperfusion of the pancreas. AB - BACKGROUND: Ischemia/reperfusion injury, and thus graft pancreatitis, remains a major problem in pancreas transplantation. Contradictory results about the role of nitric oxide (NO) in pancreatic ischemia/reperfusion have been reported; however, in none of the reports has a detailed comparison between inhibition of NO synthase and NO supplementation been carried out. METHODS: Vascular isolation of the pancreatic tail was performed in landrace pigs. After splenectomy catheters placed in the distal part of the splenic vessels allowed collection of the venous effluent and perfusion of the pancreatic tail. Three hours of complete warm ischemia was followed by 6 h of reperfusion. The effect of the NO donor sodium nitroprusside (SNP) and L-arginine was compared to a control group and NO synthase inhibition with L-NAME. RESULTS: Lipase in the venous effluent of the pancreas was significantly decreased in the SNP and the L-arginine groups. Vascular resistance was markedly elevated in the L-NAME group and reduced in the NO donor groups. Tissue pO2 after reperfusion was only significantly elevated in the SNP group. Granulocyte infiltration and also overall histological tissue injury were most severe in the control group followed by the L-NAME group, the SNP group, and the L-ARG group. CONCLUSION: The data show that supplementation of nitric oxide is clearly protective in pancreatic ischemia/reperfusion. However, inhibition of NO synthesis does not lead to an equally clear aggravation of tissue injury. PMID- 12127808 TI - Influence of epidermal growth factor on bovine pancreatic duct cell bicarbonate. AB - BACKGROUND AND AIMS: Epidermal growth factor (EGF) is secreted in pancreatic juice and its receptor is expressed on pancreatic duct cells (PDCs), suggesting a physiological role which has yet to be defined. Here we examined the effects of EGF on bicarbonate production and carbonic anhydrase (CA) activity in a PDC explant model. METHODS: Bovine main PDCs were prepared and maintained in culture as explants. Levels of CA expression, phosphorylation, and enzymatic activity were measured in resting cells and compared to that of cells exposed to 10 nM secretin, 10 nM EGF, or both. Bicarbonate production was measured using the autoburette pH titration technique. RESULTS: CA protein levels were unchanged with any treatment, but enzyme activity increased by 180% with secretin treatment and was reduced by 54% with EGF. The combination treatment led to a synergistic increase 240% above basal. EGF alone did not affect bicarbonate secretion, but the normal increase observed with secretin stimulation (1.3 +/- 0.4 to 2.9 +/- 0.6 micromol/h/cm(2)) was abolished by acute EGF pretreatment. On the other hand, EGF pretreatment for 24 h significantly increased basal and stimulated secretion (2.2 +/- 0.5 and 3.8 +/- 0.5, respectively) compared to controls. CONCLUSIONS: EGF exerts a regulatory role on bicarbonate secretion by the pancreatic duct epithelium, independent of its effect on CA activity. Its inhibition of stimulated bicarbonate secretion could play a protective role in the setting of pancreatic inflammation, where increased levels of EGF are associated with reduced pancreatic juice production. PMID- 12127809 TI - Significance of altered bilirubin subfractions in bile following hepatectomy. AB - BACKGROUND: Hyperbilirubinemia occurs as a sign of hepatic failure after hepatectomy. The pathogenesis of this event has not been elucidated. In cases complicated with postoperative infection, hyperbilirubinemia is prolonged and the composition of bilirubin subfractions in bile changes markedly. A reduction in the proportion of bilirubin diglucuronide (BDG) is especially notable. This study was aimed at clarifying the relationship between infection and biliary bilirubin subfractions, with a view to shedding light on the mechanisms of change. MATERIALS AND METHODS: Rats underwent either laparotomy or partial hepatectomy (Hx). Daily intraperitoneal injections of lipopolysaccharide (LPS) or natural saline were administered for 3 days following surgery. Total serum bilirubin levels and proportions of BDG and bilirubin in bile were measured until Day 5 after the operation. Hepatic levels of UDP-glucuronic acid (UDP-GA), UDP-glucose, NAD(+), and total adenine nucleotides (TAN) and activities of UDP glucuronyltransferase (UDP-GT) and UDP-glucose dehydrogenase were measured on Day 4. RESULTS: In hepatectomized rats treated with LPS (Hx-LPS), total serum bilirubin levels were elevated, biliary bilirubin levels were decreased, and the proportion of biliary BDG was decreased on Day 4. Hepatic levels of UDP-GA, NAD(+), and TAN and activities of UDP-GT in Hx-LPS were reduced. In all groups tested, a significant linear correlation between BDG and UDP-GA and between UDP GA and NAD(+) was found. CONCLUSIONS: The reduction of UDP-GA might be effected by reduced hepatic levels of NAD(+) in endotoxemia following hepatectomy. It is therefore suggested that alterations in biliary bilirubin subfractions might accurately reflect the energy state of the remnant liver following hepatectomy. PMID- 12127811 TI - Left ventricular end-diastolic volume from ejection fraction and stroke volume in pigs during IVC occlusion. AB - BACKGROUND: Real-time measurement of left ventricular end-diastolic volume (LVEDV), combined with left ventricular end-diastolic pressure (LVEDP), would allow continuous measurement of intraoperative diastolic function. In pursuit of this goal, we examined stroke volume divided by ejection fraction for calculation of LVEDV(sv/ef). METHODS: Five anesthetized pigs underwent median sternotomy and pericardiotomy. A transit-time ultrasonic flow probe on the ascending aorta provided cardiac output. A micromanometer provided LV end-diastolic pressure. End diastolic and end-systolic areas were measured from LV short-axis cross sections to obtain ejection fraction. LVEDV(sv/ef) was calculated during IVC occlusion. Steady-state LVEDV(echo) was determined using a three-plane echocardiography model. LVEDV(echo) was used to validate steady-state LVEDA in each experiment. RESULTS: Correlation coefficients for linear and pressure-volume relation analyses ranged from 0.46 to 0.99. The two methods for measuring LVEDV generated compliance curves with an overall reliability coefficient of 0.84. CONCLUSIONS: The LVEDV(sv/ef) method may facilitate real-time determination of LV compliance. PMID- 12127810 TI - Regional variation in myocardial water content in the edematous pig heart. AB - BACKGROUND: The purpose of this study was to examine regional variation in myocardial water content (%MWC) throughout the iatrogenically edematous pig heart. METHODS: Edema was induced by hemodilution in domestic swine (n = 26). Hearts were arrested with potassium chloride. The left ventricular free wall (LVFW), interventricular septum (IVS), and right ventricular free wall (RVFW) were biopsied at the apex, base, and equator. Full-thickness biopsy (Full, 0.5-1 g) and subendocardial (Endo, 0.1-0.2 g) biopsies were removed from each region. %MWC was determined for each biopsy. RESULTS: The %MWC for all hearts were as follows: Endo, 81.1 +/- 0.1, and Full, 80.6 +/- 0.1* (*P < 0.02); apex, 81.1 +/- 0.2*, equator, 80.7 +/- 0.2, and base, 80.4 +/- 0.2 (*P < 0.02); RVFW, 81.4 +/- 0.3*, IVS, 80.8 +/- 0.2*, and LVFW, 80.3 +/- 0.1* (*P < 0.02). For 18 hearts with LV samples with average %MWC >or= 80%, the percentages were apex, 81.4 +/- 0.2*, equator, 81.0 +/- 0.2*, and base, 80.6 +/- 0.2* (*P < 0.02) (repeated measures, ANOVA). CONCLUSION: In the iatrogenically edematous porcine heart, a significant water gradient exists, with Endo > Full, apex > equator > base, and RVFW > IVS > LVFW. These results indicate that when examining edema, consistent biopsy results depend on a reproducible sampling site. Water content tends to be highest in thin walled portions of the heart, suggesting that contractile force may be important in the distribution of edema. PMID- 12127812 TI - Effect of nitric oxide on the contractile function of rat reperfused skeletal muscle. AB - BACKGROUND: The involvement of nitric oxide (NO) in ischemia-reperfusion injury remains controversial and has been reported to be both beneficial and deleterious. The purpose of this study was to examine the contribution of NO and superoxide to skeletal muscle function using an ischemic revascularized hind limb model in rats. PATIENTS AND MATERIALS: Warm ischemia produced by vascular pedicle clamping was sustained for 3 h. The animals were divided into four groups according to the solution administrated: (1) saline, (2) N-methyl-L-arginine acetate (L-NMMA), (3) L-NMMA + N-(N-L-g-glutamyl-S-nitroso-l-cysteinyl)glycine (S nitrosoglutathione), or (4) superoxide dismutase (SOD). Saline, L-NMMA, or L-NMMA + S-nitrosoglutathione was infused for the first 2 h of reperfusion. The SOD was administered as an intravenous bolus 5 min before the onset of reperfusion. Postischemic blood flow was measured by a Doppler flow meter. Muscle contractile function was determined after 24 h of reperfusion. RESULTS: Postischemic blood flow was significantly decreased by the L-NMMA infusion compared with that in the saline-treated group. No significant difference in postischemic blood flow was noted in the saline-, L-NMMA + S-nitrosoglutathione-, and SOD-treated groups. Contractile function of the gastrocnemius muscle in the L-NMMA-and SOD-treated groups, but not in the L-NMMA + S-nitrosoglutathione group, was significantly better than that in the saline-treated group. CONCLUSION: Limiting postischemic blood flow and SOD infusion are both beneficial in decreasing the ischemia reperfusion injury of skeletal muscle. S-Nitrosoglutathione infusion following suppression of endogenous NO production does not reduce ischemia-reperfusion injury. PMID- 12127814 TI - Comparative study of the efficacy of the common topical hemostatic agents with fibrin sealant in a rabbit aortic anastomosis model. AB - OBJECTIVE: The purpose of this study was to compare the hemostatic efficacy of the common surgical hemostatic agents with fibrin sealant (FS) and to assess their functional strength to secure hemostasis in lieu of placing additional sutures. METHODS: End-to-end anastomosis of transected abdominal aorta was performed in moderately anticoagulated rabbits using 4 or 6 interrupted sutures. The suture line was covered either with gauze alone ("untreated") or with gauze plus Gelfoam, Avitene, Surgicel, FloSeal, or FS, following which blood flow was restored. Blood loss was absorbed by gauze and measured. The surviving rabbits were recovered and the repaired vessel was examined histologically 4 weeks after operation. The investigators were blinded to the treatment groups. Aortic anastomoses using 8 or 12 sutures (untreated) were also performed. RESULTS: Untreated 4-suture anastomosis of aorta resulted in a profuse hemorrhage with an average 108.0 +/- 19.2 (mean +/- SD) ml blood loss and 100% mortality (n = 4). FS application sealed the anastomoses, prevented blood loss (P < 0.01 vs untreated) and exsanguination of the rabbits (n = 4). Other hemostatic agents reduced the bleeding to varying degrees compared to the untreated animals (Gelfoam 66.4 +/- 17.6, Avitene 80.6 +/- 34, Surgicel 66.7 +/- 16.7, FloSeal 44.2 +/- 8.5 ml blood loss, n = 4/group), but the changes were not statistically significant. One to three rabbits in each group survived the operation. Six-suture aortic anastomoses, untreated, resulted in 67.7 +/- 21.8 ml blood loss and 100% survival (n = 6). Application of FS produced immediate and sustained hemostasis in all the animals (P < 0.01 vs untreated). Other hemostatic agents also reduced the bleeding (Gelfoam 42.5 +/- 10, Avitene 50.9 +/- 12.4, Surgicel 32.1 +/- 14, FloSeal 33.9 +/- 5.4 ml blood loss, n = 6/group), but the changes were not statistically significant. The 8- and 12-suture aorta repairs resulted in a moderate blood loss (43.9 +/- 19 and 21.3 +/- 14.9 ml, respectively), followed by a stable hemostasis that precluded the need to use any hemostatic agent. The aortic cross-clamping time of the 12-suture and time to hemostasis for both the 8 and the 12-suture techniques were significantly longer than those of the 4 suture plus FS application (P < 0.01, P < 0.01 and P < 0.05, respectively). CONCLUSION: In a moderate coagulopathy, FS was proven to be the most efficacious hemostatic agent, producing immediate and sustained hemostasis at the arterial anastomotic site. This high efficacy is in part attributed to the strong tissue adhesive property of this agent. FS application may potentially ease the anastomosis and shorten the duration of timely critical vascular procedures. PMID- 12127813 TI - Anti-ETA IgG neutralizes the effects of Pseudomonas aeruginosa exotoxin A. AB - BACKGROUND: The opportunistic pathogen Pseudomonas aeruginosa causes severe infections in immunocompromised hosts. Among P. aeruginosa-infected burn patients, mortality may reach as high as 50%. Due to their immunocompromised status, burn patients may benefit from passive immunotherapy against infection. As a potential multivalent immunoglobulin therapy, specific polyclonal antibodies against four P. aeruginosa virulence factors, including exotoxin A (ETA), were prepared. MATERIALS AND METHODS AND RESULTS: In this study, we analyzed the ability of ETA antibody (ETA-Ab) to neutralize the in vivo effects of ETA. Adult mice injected with purified ETA suffered 100% mortality. The cytosolic DNA of their hepatocytes was fragmented, indicating ETA induction of apoptosis. In addition, multiprobe RNase protection assays showed that ETA upregulates the expression of the genes for proinflammatory cytokines as well as apoptosis genes in the livers of ETA-injected mice. Treatment with ETA-Ab prior to ETA injection prevented mortality, ETA-induced hepatocyte DNA fragmentation, and upregulation of the cytokine and apoptosis-related genes. The role of ETA during P. aeruginosa infection of the burn wound was examined by determining the in vivo virulence of P. aeruginosa PA103 and its isogenic, ETA-deficient mutant PA103Omega::toxA using the thermally injured mouse model. The lethality, local spread, and systemic spread of PA103Omega::toxA were significantly reduced compared to PA103. CONCLUSION: These results suggest that (1) ETA induces apoptosis in hepatocytes, (2) specific cytokines are produced in response to ETA, (3) ETA-Ab neutralizes these effects, and (4) ETA contributes to the spread of P. aeruginosa during burn wound infection. PMID- 12127815 TI - Impairment of activation of hepatocyte growth factor precursor into its mature form in rats with liver cirrhosis. AB - BACKGROUND: Hepatocyte growth factor (HGF) has a crucial role in liver regeneration following injury. The conversion of an inactive precursor form of HGF (proHGF) into a biologically active form (mature HGF) is essential, as HGF is involved in the recovery of liver damage. Liver regeneration is markedly poor in patients with liver cirrhosis after resection. We hypothesized that impairment of liver regeneration in cirrhosis is in part because of the absence of activation of proHGF to mature HGF. Studies were performed to clarify the molecular form of HGF in the liver of rats with fibrosis/cirrhosis before and after liver resection. METHODS: Rat models of liver fibrosis/cirrhosis were induced by intraperitoneal administration of dimethylnitrosamine, followed by 45% partial hepatectomy or sham operation. HGF was purified from the liver and plasma on a SP Sepharose column and was analyzed by Western blotting. RESULTS: Production of proHGF in the liver increased in the following order: rats with normal liver, rats with fibrosis, and rats with cirrhosis. However, the levels of proHGF were similar after liver resection in the liver of these groups. A small but significant level of mature HGF was detected before resection in the fibrosis group, but not in the normal and cirrhosis groups. Liver resection increased the levels of mature HGF in the normal and fibrosis groups, but marginally in the cirrhosis group. CONCLUSIONS: These results demonstrate that the conversion of proHGF into mature HGF is impaired after liver resection in liver cirrhosis, while proHGF production is similar in the livers of normal, fibrosis, and cirrhosis groups. Acceleration of the processing of the HGF molecule may contribute to the improvement of liver dysfunction in cirrhosis. PMID- 12127816 TI - Long-term culture of primary human hepatocytes with preservation of proliferative capacity and differentiated functions. AB - BACKGROUND: The aim of this study was to develop a suitable method for the prolonged culture and maintenance of human hepatocytes with preservation of both proliferative capacity and differentiated functions. MATERIALS AND METHODS: Primary human hepatocytes were isolated from small pieces of liver tissue obtained from 15 patients who underwent hepatic resection. Hepatocytes were cultured in keratinocyte-stimulating factor medium supplemented with 10% human serum, 10 mM nicotinamide, 10 ng/ml epidermal growth factor, 0.5 microg/ml insulin, 10(-7) M dexamethasone, and antibiotics. Hepatic differentiation and function were analyzed by immunocytochemistry, Western blot, ELISA, lidocaine metabolism, and urea synthesis. Ultrastructural analysis of cultured hepatocytes was performed by electron microscopy. RESULTS: Many primary hepatocytes were maintained for more than 56 days. Hepatocytes proliferated during the initial 14 days, and bromodeoxyuridine labeling indices were 15.2, 12.2, and 6.2% at days 5, 10, and 15, respectively. Electron micrographs of the hepatocytes at day 28 demonstrated numerous mitochondria, rough endoplasmic reticulum, large peroxisomes, and glycogen granules. Albumin secretion increased for the first 14 days and then gradually decreased thereafter but was maintained at levels greater than 2 microg/ml/h until day 56. alpha(1)-Antitrypsin, alpha(1)-antichymotrypsin, and ceruloplasmin production was also observed at day 56, while lidocaine metabolism and urea synthesis were maintained for a long time. CONCLUSION: This hepatocyte culture method facilitates the prolonged culture of primary human hepatocytes with preservation of hepatocyte differentiation, function, and proliferative capacity. PMID- 12127817 TI - Genetic detection of liver micrometastases that are undetectable histologically. AB - BACKGROUND: Predicting liver metastasis from colon cancer is essential for improving its prognosis. We studied to what extent genetic detection of cancer cells in the resected liver tissue can predict the incidence of macroscopic liver metastasis with a similar mouse model to clinical colorectal cancer that causes a several decade percentage of metachronous hepatic metastases after resection of the primary lesions. MATERIALS AND METHODS: A LS174T human colorectal cancer cell suspension was injected into the spleens of nude mice. One to 10 days after splenic injection, 3 x 3 mm of liver tissue was removed, and a splenectomy was performed. Liver tissue was used for genetic detection and histological examination. Five weeks after splenic injection, the number of macroscopic metastases on the surface of the liver was counted. RESULTS: Eight of the 45 cases were positive for tumor cells in liver tissue genetically, while only 1 was positive for tumor cells histologically. Macroscopic liver metastases were seen 5 weeks after splenic injection in 11 of 37 (29.7%) cases negative for tumor cells genetically and in 8 of 8 (100%) cases positive for tumor cells genetically. Five or more metastases were seen in 3 of 37 (8.1%) cases negative for tumor cells genetically and in 7 of 8 (87.5%) cases positive for tumor cells genetically. CONCLUSIONS: The cases which were positive for tumor cells in liver tissue genetically at the time of splenectomy had more significantly macroscopic liver metastases some weeks later than the cases negative for tumor cells. This study suggests that if micrometastasis was detected genetically, the development of metachronous macroscopic liver metastasis could be predicted. PMID- 12127818 TI - Impact of abdominal CT imaging on the management of appendicitis: an update. AB - BACKGROUND: Abdominal computed tomographic scanning (ACTS) has recently been shown to be an accurate diagnostic tool for appendicitis and may improve the negative exploration rate in our patient population. MATERIALS AND METHODS: We reviewed 224 patients evaluated for appendicitis during 1998. Forty-two patients underwent appendectomy on clinical grounds alone (Group I), 182 patients underwent ACTS (Group II), and 79 patients in Group II were explored for appendicitis. Diagnostic errors, alternative diagnoses, and perforation rates were noted. RESULTS: There were five negative explorations in Group I (11.9%) and five in Group II (6.3%), resulting in a combined negative rate of 8.3%. The negative exploration rate in women was 23.5% in Group I and 5.3% in Group II (P = 0.07), producing a combined negative rate of 10.9%. Fifty-eight alternative diagnoses were made by ACTS. The ACTS made a critical difference in the management of 67% of patients over 50 years of age and in 79% of Group II patients. CONCLUSIONS: The negative exploration rate for appendicitis at our institution fell from 13.6 to 8.3% with selective use of ACTS. The most striking benefit occurred in women and in patients over 50 years of age. PMID- 12127819 TI - Protein kinase C activation of intestinal glutamine transport is mediated by mitogen-activated protein kinases. AB - BACKGROUND: Glutamine is essential for the preservation of intestinal structure and function and its uptake by the bowel is augmented during catabolic states. However, the signal transduction pathways implicated in brush border glutamine transport have not been examined. The aim of this study was to investigate the intracellular signaling pathways involved in the regulation of accelerated intestinal glutamine transport. Our hypothesis was that the activation of intestinal glutamine transport involves protein kinase C (PKC) and is mediated by mitogen-activated protein kinases (MAPKs). METHODS: [3H]L-Glutamine (50 microM) transport activity and mRNA levels for the intestinal glutamine transporter ATB(0) were measured in intestinal epithelial Caco-2 cells. Confluent cells were treated with phorbol ester (PMA, 0-10 microM), the MAPK MEK inhibitor PD 98059 (0 100 microM), actinomycin (0-0.1 microM), MAPK p38 inhibitor SB 203580 (0-10 microM), protein kinase C inhibitor chelerythrine chloride (0-6.6 microM), or cycloheximide (0-10 microM) for 24 h. Data were analyzed by ANOVA with significance set at P < 0.05. RESULTS: Phorbol ester treatment increased intestinal System B glutamine transport activity by 75%, an increase that was blocked individually by PD 98059, chelerythrine chloride, actinomycin, and cycloheximide, but not SB 203580, an effect first noted at 6 h. The resulting activity increase was consistent with de novo synthesis of transporter units and enhanced expression of transporter gene ATB(0) as indicated by a threefold increase of ATB(0) mRNA levels in PMA-treated cells. CONCLUSIONS: Activation of glutamine transport in Caco-2 cells by phorbol ester occurs via signaling pathways that lead to transcription of the glutamine transporter gene. PKC and mitogen-activate protein kinase MEK are key intracellular mediators involved in this signal transduction cascade. PMID- 12127820 TI - Surgical manipulation of the small intestine and its effect on the lung. AB - BACKGROUND: Surgical manipulation of the intestine results in generation of oxygen free radicals leading to mucosal damage as evidenced by ultrastructural and biochemical changes. It is likely that the gut-derived mediators can bring about damage to distant organs such as the lung. METHODS: Surgical manipulation of the gut was performed by opening the abdominal wall and handling the intestine. Lung damage was assessed by histology, markers of oxidative stress, and protein content in bronchoalveolar lavage fluid. Protection offered by pretreatment with various compounds such as allopurinol, L-arginine, quinacrine, and indomethacin was also studied. RESULTS: Gut manipulation resulted in neutrophil infiltration, oxidative stress, and permeability changes in the lung and these changes were maximum 30 and 60 min following surgical manipulation, which recovered with time and reversed to normal by 24 h. Prior treatment with inhibitors of xanthine oxidase, phospholipase A(2), or cyclooxygenase showed a protective effect against lung damage. CONCLUSION: This study has shown that laparotomy and intestinal handling result in distant organ (lung) damage which is probably brought about by neutrophil infiltration and oxidative stress on the lung. This is likely mediated by compounds generated in the intestine and transported into the systemic circulation since inhibition of generation of chemical mediators in the intestine offers protection against lung damage. PMID- 12127821 TI - Regulation of hypoxia-inducible factor 1 in enterocytic cells. AB - BACKGROUND: Mucosal hypoxia due to intestinal hypoperfusion is characteristic of a number of clinical disorders. An early event in the adaptive response to cellular hypoxia is the binding of hypoxia-inducible factor 1 (HIF-1) to cis acting regulatory sites in target genes. METHODS: We characterized the expression of HIF-1 in transformed (Caco-2(BBe) and T84) and nontransformed human (FHs 74 Int) and rat (IEC-6) intestinal epithelial cell lines. RESULTS: The electrophoretic mobility shift assay detected increased HIF-1 DNA-binding activity in each cell line within 2 h of hypoxia (1% O2). HIF-1 binding was maximal within 4 h and remained stable for 24 h. HIF-1 DNA-binding activity was maximal in the established IEC-6 cell line below 2% oxygen, but HIF-1 DNA-binding activity was not detectable above 0.5% O2 in the primary human FHs 74 Int cell line. The nonspecific protein kinase inhibitor genistein (200 microM) inhibited HIF-1 binding at 4 h. Transfection of Caco-2 cells with a wild-type, but not a mutant, HIF-1-dependent luciferase expression vector resulted in a fourfold induction of reporter gene expression during hypoxia. CONCLUSIONS: In conclusion, HIF-1 regulates gene expression in enterocytes and an undefined phosphorylation event is important for O2 sensing. PMID- 12127822 TI - Role of PKC in the late phase of microvascular protection induced by preconditioning. AB - INTRODUCTION: We hypothesized that the late phase of microvascular protection induced by ischemic preconditioning or by adenosine is protein kinase C (PKC) dependent. MATERIALS AND METHODS: The cremaster muscle of male Sprague-Dawley rats underwent 45 min of ischemic preconditioning and, 24 h later, 4 h of warm ischemia followed by 60 min of reperfusion. To mimic the effects of IPC, adenosine (ADO; an adenosine receptor agonist) or 4-phorbol 12-myristate 13 acetate (PMA; a PKC activator) was delivered to the vascular network of the cremaster 24 h before the prolonged ischemia via local intra-arterial infusion. To block the microvascular protection induced by ADO or IPC, chelerythrine (CHE; a PKC blocker) was given by local intra-arterial infusion prior to the administration of ADO or the initiation of IPC. Microvascular responses in the cremaster muscle to ischemic preconditioning or pharmacological preconditioning were determined by measuring terminal arteriole diameter and capillary perfusion using intravital microscopy and by the evaluation of the endothelium-dependent nitric oxide system in terminal arterioles. RESULTS: Blockade of PKC using CHE on day 1 eliminated both ADO- and IPC-induced microvascular protections seen on day 2. However, the microvascular protection induced by the administration of PMA (without IPC) that was given 24 h before the 4 h of warm ischemia/reperfusion was significantly better than the control group response (sham IPC), but was not as good as the protection induced by IPC or ADO alone. CONCLUSION: The overall results from these studies suggest that ischemic or ADO preconditioning induces late-phase microvascular protection in skeletal muscle by a PKC-dependent mechanism. PMID- 12127823 TI - E-cadherin expression is inversely proportional to tumor size in experimental liver metastases. AB - OBJECTIVE: The role of E-cadherin in the metastatic process remains controversial. In this study, we examined what role E-cadherin plays in relation to tumor size and microvessel density (MVD) of the liver metastatic lesion. MATERIALS AND METHODS: In F344 male rats, metastatic lesions were made by injecting RCN-9 cells into the spleen. We performed an immunohistochemical evaluation of E-cadherin and MVD in the metastatic lesions. According to a previous report, we focused on tumor sizes ranging from 2 to 3 mm, and the metastatic nodules were divided into two groups considering the value of the MVD. RESULTS: The expression of E-cadherin was inversely proportional to tumor size (r = -0.882, P < 0.0001). MVD was directly proportional to metastatic tumor size (r = 0.726, P = 0.001). MVD in group A, in which the tumor size was 2.3 mm or more, was significantly higher than that in group B, in which the tumor size was less than 2.3 mm (3.13 +/- 1.65% vs 1.75 +/- 0.27%; P = 0.0069). The expression ratio of E-cadherin in group A was 28.21 +/- 16.36%, while it was 82.33 +/- 16.35% in group B (P = 0.0003). CONCLUSION: In liver metastasis, E-cadherin's function is preserved when the tumor is small and E-cadherin's expression is reduced in large tumors in which neovascularization is increased. PMID- 12127825 TI - Hepatic endosomal trafficking of lipoprotein-bound endotoxin in rats. AB - Triglyceride-rich lipoproteins (chylomicrons (CM), VLDL) can bind and protect against endotoxin (LPS)-induced shock and mortality in rodents. The protective effect of lipoproteins is in part due to the increased plasma clearance and biliary excretion of LPS. Specifically, CM-LPS complexes are principally removed from the circulation by the liver with a rapid plasma half-life approximating that for CM alone. Thus, we hypothesized that hepatocytes clear CM-bound LPS via known lipoprotein receptors and traffic the toxic macromolecule through the same endosomal pathway employed for the catabolism of triglyceride-rich lipoproteins. To examine the endosomal uptake and biliary excretion of LPS, we isolated early and late hepatic endosomal fractions and hepatic bile from rats following the injection of radiolabeled CM-bound LPS. The uptake of (125)I-LPS was compared in animals that overexpressed either the LDL receptor or the LDL receptor-related protein (LRP) versus untreated control with normal lipoprotein levels. Herein we present data indicating that both the LDL receptor and the LRP participate in the rapid internalization of CM-bound LPS by hepatocytes. Upregulation of the LDL receptor increased the accumulation of (125)I-LPS in late endosomes (P < 0.03). In contrast, increased levels of the LRP were associated with negligible movement of LPS into late endosomes but a trend toward the increased biliary excretion of the radiolabeled macromolecule. Taken together these data further elucidate the role of the liver in the host innate immune response to infection and potentially implicate distinct roles for the LDL receptor and LRP in the catabolism of CM bound LPS. PMID- 12127824 TI - Functional heterogeneity of the kupffer cell population is involved in the mechanism of gadolinium chloride in rats administered endotoxin. AB - BACKGROUND: The purpose of this study was to determine if evidence of functional heterogeneity between subtypes of the Kupffer cell (KC) may be involved in the mechanism of the protective effect of gadolinium chloride (GdCl3) in endotoxemia. METHODS: Rats pretreated with or without GdCl3 were administered lipopolysaccharide (LPS) or vehicle. Serum and liver tissues were collected after LPS administration for cytokine measurements and pathological and immunohistochemical evaluation. RESULTS: After LPS administration, increases in expression of TNF-alpha and IL-6 mRNA in the liver were blunted significantly by GdCl3. In control liver tissue, ED2-positive cells were a predominant fraction, with a few ED1-positive cells, and GdCl3 eliminated only ED2-positive cells. Further, ED2-positive cells were larger in size than ED1-positive ones. Importantly, the number of ED1-positive cells in the liver was increased about threefold in the control group but not in the GdCl3 group after LPS injection. Intermediate or large KCs isolated by counterflow centrifugal elutriation showed greater capacity for phagocytosis and production of superoxide and TNF-alpha than small ones. In contrast, IL-6 production was increased to a greater extent in small than in intermediate or large cells. GdCl3 eliminated the intermediate or large KC subpopulation predominantly. CONCLUSION: Collectively, functional heterogeneity of the KC population was involved in the mechanism of the protective effects of GdCl3 in endotoxemia. TNF-alpha derived from activated intermediate or large KCs may activate small KCs and the latter may be recruited to other organs, such as lungs and kidneys, and produce a large amount of IL-6, leading to multiple organ failure. PMID- 12127827 TI - Microinjection of DNA into the nuclei of human vascular smooth muscle cells. AB - BACKGROUND: It is challenging to successfully transfect human vascular cells by conventional techniques. We evaluated the efficiency of transfection of human smooth muscle cells (SMC) using a method of direct nuclear microinjection of DNA constructs. MATERIALS AND METHODS: The nuclei of explanted human saphenous vein SMC were microinjected with the plasmid pCMVbeta, containing the lacZ gene for beta-galactosidase (beta-gal). Efficiency of injection and expression were assessed by histochemical staining for beta-gal. Injected SMC were subjected to standard assays of viability and migration. RESULTS: Parameters affecting the conditions of injection were systematically analyzed to achieve optimal transfection efficiency. A vertical injection resulted in a twofold increase in expression of beta-gal compared to a horizontal approach. A DNA concentration of 100 ng/microl (390 copies/injection) provided a maximal rate of expression. No further increase in expression was evident at higher concentrations. Maximal expression was achieved with a time of injection of 200-500 ms, an injection pressure of 5-10 psi, and a pipette tip size of 0.6 microm, resulting in an injection volume of 0.03 pl. Cytoplasmic injection did not result in gene expression. The ability of SMC to migrate under videomicroscopy was not altered by the injection process. Optimizing all injection parameters resulted in cell viability >95% and efficiency of injection of 59%. CONCLUSION: DNA encoding a variety of intracellular proteins can be efficiently microinjected into human vascular SMC. Coupled with the use of videomicroscopy, this technique can allow for the evaluation of genes that might modulate important cellular processes such as proliferation and migration. PMID- 12127826 TI - Fertility assessment of hydatid cyst by proton MR spectroscopy. AB - BACKGROUND: Hydatid cysts, the larvae of the parasite Echinococcus granulosus, may lodge in any organ of intermediate hosts, namely, man and sheep. Complete cyst removal is the treatment of choice; however, spillage of fertile cysts during surgery leads to disease recurrence that may be prevented by preoperative detection of the fertility status of the cyst. With this perspective, ex vivo proton (1H) MR spectroscopy of hydatid fluid of human and sheep origin was performed to differentiate fertile from sterile cysts on the basis of their metabolite pattern. PATIENTS AND MATERIALS: Cysts of sheep and human origin were used as source of hydatid fluid. A fraction of this fluid was tested for cyst fertility and the rest was used for ex vivo1H spectroscopy. Histopathology of the cyst wall was done as a gold standard for this study. RESULTS: Of 10 sheep samples, 7 were fertile and 3 were sterile, while among 6 human samples, 5 were fertile and 1 was sterile. Spectroscopic and histopathological results corroborated each other. The fluid from microbiologically proven fertile cysts contained malate and fumarate along with other resonances and the histopathology of the fertile cyst wall demonstrated germinal lining and protoscoleces. CONCLUSIONS: The ex vivo spectroscopic differentiation of fertile and sterile cysts may be a stepping-stone for their in vivo separation in future and thus help in framing strategies for percutaneous/surgical management. PMID- 12127828 TI - Pulmonary hypoplasia and congenital diaphragmatic hernia: advances in the pathogenetics and regulation of lung development. PMID- 12127829 TI - Problems in measuring or interpreting change in patient outcomes. PMID- 12127830 TI - A prospective, randomized, pragmatic, health outcomes trial evaluating the incorporation of hylan G-F 20 into the treatment paradigm for patients with knee osteoarthritis (Part 1 of 2): clinical results. AB - OBJECTIVE: First, to assess the clinical effectiveness of hylan G-F 20 in an appropriate care treatment regimen (as defined by the American College of Rheumatology (ACR) 1995 guidelines) as measured by validated disease-specific outcomes and health-related quality of life endpoints for patients with osteoarthritis (OA) of the knee. Second, to utilize the measures of effectiveness and costs in an economic evaluation (see accompanying manuscript). DESIGN: A total of 255 patients with OA of the knee were enrolled by rheumatologists or orthopedic surgeons into a prospective, randomized, open-label, 1-year, multi centred trial, conducted in Canada. Patients were randomized to 'Appropriate care with hylan G-F 20' (AC+H) or 'Appropriate care without hylan G-F 20' (AC). Data were collected at clinic visits (baseline, 12 months) and by telephone (1, 2, 4, 6, 8, 10, and 12 months). RESULTS: The AC+H group was superior to the AC group for all primary (% reduction in mean Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC) pain scale: 38% vs 13%,P =0.0001) and secondary effectiveness outcome measures. These differences were all statistically significant and exceeded the 20% difference between groups set a priori by the investigators as the minimum clinically important difference. Health-related quality of life improvements in the AC+H group were statistically superior for the WOMAC pain, stiffness and physical function (all P< 0.0001), the SF-36 aggregate physical component (P< 0.0001) and the Health Utilities Index Mark 3 (HUI3) overall health utility score (P< 0.0001). Safety (adverse events and patient global assessments of side effects) differences favoured the AC+H group. CONCLUSION: The data presented here indicate that the provision to patients with knee OA of viscosupplementation with hylan G-F 20 within an appropriate care treatment regimen provides benefits in the knee, overall health and health related quality of life at reduced levels of co-therapy and systemic adverse reactions. PMID- 12127831 TI - A prospective, randomized, pragmatic, health outcomes trial evaluating the incorporation of hylan G-F 20 into the treatment paradigm for patients with knee osteoarthritis (Part 2 of 2): economic results. AB - OBJECTIVE: Viscosupplementation with hylan G-F 20 has recently become registered for treatment of patients with osteoarthritis (OA) of the knee in most parts of the world. The cost effectiveness and cost utility of this new therapeutic modality were determined as part of a Canadian prospective, randomized, 1-year, open-label, multicentered trial. DESIGN: A total of 255 patients were randomized to 'Appropriate care with hylan G-F 20' (AC+H) or 'Appropriate care without hylan G-F 20' (AC). Costs (1999 Canadian dollars) were collected from the societal viewpoint and included all costs related to OA of the knee and OA in all joints. Patients completed a number of outcomes questionnaires including the Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC) and the Health Utilities Index Mark 3 (HUI3). Data were collected at clinic visits (baseline, 12 months) and by telephone (1, 2, 4, 6, 8, 10, and 12 months). RESULTS: The AC+H group over the year had higher costs ($2125-$1415=$710, P< 0.05), more patients improved (69%-40%=29%,P =0.0001), greater increases in HUI3 (0.13-0.03=0.10, P< 0.0001) and increased quality-adjusted life years (QALYs) (0.071, P< 0.05). The incremental cost-effectiveness ratio was $2505/patient improved. The incremental cost-utility ratio was $10000/QALY gained. Sensitivity analyses and a second cost perspective gave similar results. CONCLUSION: The cost-utility ratio is below the suggested Canadian adoption threshold. The results provide strong evidence for adoption of treatment with hylan G-F 20 in the patients and settings studied in the trial. PMID- 12127832 TI - Novel cartilage-specific splice variants of fibronectin. AB - OBJECTIVE: To determine the nature of alternatively spliced isoforms of fibronectin expressed in healthy bovine articular cartilage and cartilage derived from human osteoporotic and osteoarthritic joints. DESIGN: Our study focused on a single alternatively spliced region of the fibronectin gene, the variable region. Bovine cartilage samples were obtained within 12h of slaughter and human cartilage samples were obtained within 8h of the time of joint replacement surgery. RNA was extracted and alternatively spliced isoforms of fibronectin were amplified using RT-PCR. RESULTS: Two novel alternatively spliced forms of fibronectin designated (V+I-10)(-) and (V+III-15)(-) were identified in bovine articular cartilage. Fibronectin is composed of multiple repeats of three types of homologous units and these two novel isoforms specifically splice out single individual repeating units. Expression of all these isoforms was dependent upon the presence of an extracellular matrix. In the human samples the expression profiles obtained with osteoporotic hip and osteoarthritic knee cartilage was not uniform. The (V+C)(-) isoform was present in all samples and the (V+I-10)(-) isoform was distributed between both osteoporotic and osteoarthritic cartilage. However, the (V+III-15)(-) isoform was shown to be associated with osteoarthritic cartilage with statistical significance (P< 0.015 ). In addition a third novel splice variant was identified and designated as III-15X. Translation of the III 15X isoform results in a truncated form of fibronectin lacking a significant portion of the C-terminus. Further RT-PCR analysis of several other tissue types suggests that these variants are cartilage specific. CONCLUSION: Our results demonstrate the existence of three new cartilage specific isoforms of fibronectin and indicate that expression of one or more may be associated with osteoarthritis. Published by Elsevier Science Ltd. All rights reserved. PMID- 12127833 TI - Effect of articular cartilage proteoglycan depletion on high frequency ultrasound backscatter. AB - OBJECTIVE: To study the effect of variations of articular cartilage proteoglycans (PG) on high-frequency ultrasound backscatter. DESIGN: The study was performed on patellar cartilages of immature and mature rats (N=36). The variation of PG content was induced by enzyme digestion. Control and treated cartilages were explored in vitro using a 55MHz scanning acoustic microscopy, then assessed by histology for the fibrillar collagen organization analysis. The variations of proteoglycan and collagen content were evaluated. Thickness measurements performed on both B-scan images and histologic sections were compared. Ultrasonic radio-frequency signals reflected by the cartilage surface and backscattered from its internal matrix were processed to estimate the integrated reflection coefficient (IRC) and apparent integrated backscatter (AIB). RESULTS: Although hyaluronidase treatment of immature and mature cartilages removed approximately 50% of the proteoglycans, the echogenicity level of ultrasound images of degraded cartilages was similar to that of controls. IRC and AIB parameters did not significantly vary. Histologic sections of degraded cartilage displayed no change in collagen fiber organization. The thickness mean values measured by ultrasound in PG-depleted groups were significantly higher than in controls, whereas no significant difference in thickness was detected by histological measurement. The increase in cartilage thickness may potentially be explained by a decrease of speed of sound in PG-depleted cartilages that is more likely subsequent to an increase of water content. CONCLUSION: Current results indicate that PG depletion has no significant effect on high frequency ultrasound backscattered from rat patellar cartilage. Ultrasound may provide information about variations of PG content via speed of sound measurement. PMID- 12127834 TI - Quantitative assessment of joint space width with an electronic caliper. AB - The progression of joint space narrowing (JSN) is considered to be the best available marker of osteoarthritis (OA) progression. Several techniques have been proposed for the measurement of joint space at its narrowest point in OA of the hips and knees. OBJECTIVE: To evaluate the properties of the technique using an electronic caliper for the measurement of JSN in OA patients. DESIGN: We used an electronic caliper to measure joint space width (JSW) for hips on 100 plain radiographs. JSW was measured in the vertical position at the center of the femoral head. Femoral head diameter was also determined to correct for variations due to differences in magnification of digitized X-rays. All films were read twice by each of two rheumatologists (one junior, one senior) and two radiologists (one junior, one senior). Intraclass correlation coefficients and their 95% confidence intervals were calculated. RESULTS: Detailed results are given for right hips (38 with OA, 18 inflammatory, 44 normal); very similar results were obtained for left hips. For JSW, the intraclass correlation coefficient was between 0.96 and 0.99 for intraobserver reliability. The level of reliability was similar for analysis of the diameter of the femoral head (R:0.84 to 0.98) and for the ratio of these two measurements (0.96 to 0.99). The most reliable measurements were those made by the senior radiologist, followed by those made by the two rheumatologists. In assessments of interobserver reliability for the measurement of JSW, R varied from 0.91 to 0.96 for the first reading and from 0.88 to 0.96 for the second reading. For the measurement of femoral head diameter, R varied from 0.86 to 0.96 for the first reading and from 0.74 to 0.96 for the second reading. CONCLUSION: The electronic caliper technique is an accurate method for measuring JSW in the hip. This technique seems to be reproducible, is simple, and could be used for routine evaluation. Further validation is required, with the measurement of serial X-rays from the same patients. PMID- 12127835 TI - Regulation by reactive oxygen species of interleukin-1beta, nitric oxide and prostaglandin E(2) production by human chondrocytes. AB - OBJECTIVES: To determine the effects of two drugs, N-monomethyl-L-arginine (L NMMA) and N-acetylcysteine (NAC), on interleukin-1beta (IL-1beta), nitric oxide (NO) and prostaglandin E(2) (PGE(2)) production by human chondrocytes. The effect of aceclofenac (ACECLO), a non-steroidal antiinflammatory drug (NSAID), was also examined. METHODS: Human chondrocytes were enzymatically isolated from osteoarthritic knee cartilage and then maintained in culture in suspension for 48h in the absence or in the presence of lipopolysaccharide (LPS) (10 microg/ml), L-NMMA (0.5mM), NAC (1mM) or ACECLO (6.10(-6)M). IL-1beta and PGE(2) productions were quantified by specific immunoassays. Nitrite was measured in the culture supernatants by a spectrophotometric method based upon the Griess reaction. Cyclooxygenase-2 (COX-2), inducible NO synthase (iNOS) and IL-1beta gene expressions were quantified by transcription of mRNA followed by real time and quantitative polymerase chain reaction. COX-2 protein expression was analysed by Western blot. RESULTS: LPS markedly increased the expression of IL-1beta, iNOS and COX-2 genes. In parallel, NO(2) and PGE(2) amounts found in the culture supernatants were significantly enhanced whereas IL-1beta was immunologically undetectable. The addition of L-NMMA (0.5mM) fully blocked LPS-induced NO production but greatly increased PGE(2) production, suggesting a negative effect of NO on PGE(2) synthesis. Inversely, NO production was stimulated by NAC while PGE(2) production was not affected. Interestingly, NAC increased the IL-1beta and iNOS mRNA levels but did not significantly modify COX-2 mRNA expression. L-NMMA did not significantly affect the expression of IL-1beta, iNOS and COX-2. The amount of COX-2 protein did not change in the presence of the antioxidants. Finally, ACECLO fully blocked the production of PGE(2) by chondrocytes without affecting the levels of COX-2 mRNA. CONCLUSIONS: The stimulation of IL-1beta, NO and PGE(2) production by LPS is differentially controlled by reactive oxygen species (ROS). In fact, L-NMMA and NAC have different mechanisms of action on the regulation of NO and PGE(2) productions. L-NMMA fully inhibits NO but increases PGE(2) production whereas NAC up-regulates NO but does not modify PGE(2) synthesis. The stimulating effect of L-NMMA on PGE(2) production is not controlled at the transcriptional level. These findings suggest that antioxidant therapy could have different effects according to the oxygen radical species targeted. PMID- 12127836 TI - Co-localization of insulin-like growth factor binding protein 3 and fibronectin in human articular cartilage. AB - OBJECTIVE: The anabolic cytokine insulin-like growth factor I (IGF-I) stimulates chondrocyte synthesis of matrix macromolecules and several lines of evidence suggest that it has a major role in maintaining articular cartilage and possibly in cartilage repair. Despite the apparent importance of IGF-I in articular cartilage metabolism and its potential importance in joint diseases, little is known about the regulation of IGF-I activity within the tissue. Insulin-like growth factor binding proteins (IGFBPs) bind IGF-I and can modify its activity. At least three IGFBPs are expressed by chondrocytes: IGFBP-3, -4 and -5. Localization of IGFPBs in the articular cartilage extracellular matrix (ECM) could create reservoirs of IGF-I within the articular cartilage ECM and thereby regulate local IGF-I levels. We hypothesized that ECM molecules bind and concentrate IGFPBs in the pericellular/territorial matrix. DESIGN: Semi quantitative immunohistological measures of co-localization were used to compare the spatial distribution of IGFBP-3, -4, and -5 with the distributions of three peri-cellularly-enriched matrix molecules fibronectin, tenascin-C, and type VI collagen in osteoarthritic and non-osteoarthritic human articular cartilage. Purified proteins were used in an agarose diffusion assay to compare IGFBP-3 binding to the same three matrix proteins. RESULTS: IGFBP-3 associated with fibronectin in the pericellular/territorial matrix (approximately 40% co localization) but not with tenascin-C, or type VI collagen (approximately 6% and approximately 15% co-localization respectively, P< 0.05). Neither IGFBP-4, nor IGFBP-5 were associated with any of the three ECM proteins (P< 0.05). In agarose diffusion assays IGFBP-3 interacted with fibronectin and heparan sulfate proteoglycan but not with type VI collagen or tenascin-C. CONCLUSIONS: Direct binding between purified IGFBP-3 and fibronectin and the strong co-localization the two proteins in the cartilage matrix support the hypothesis that IGFPB-3 and fibronectin help regulate local IGF-I levels. PMID- 12127837 TI - Quantitative structural organization of normal adult human articular cartilage. AB - OBJECTIVE: Data pertaining to the quantitative structural features and organization of normal articular cartilage are of great importance in understanding its biomechanical properties and in attempting to establish this tissue's counterpart by engineering in vitro. A comprehensive set of such baseline data is, however, not available for humans. It was the purpose of the present study to furnish the necessary information. DESIGN: The articular cartilage layer covering the medial femoral condyle of deceased persons aged between 23 and 49 years was chosen for the morphometric analysis of cell parameters using confocal microscopy in conjunction with unbiased stereological methods. The height of the hyaline articular cartilage layer, as well as that of the calcified cartilage layer and the subchondral bone plate, were also measured. RESULTS: The mean height of the hyaline articular cartilage layer was found to be 2.4mm, the volume density of chondrocytes therein being 1.65%, the number of cells per mm(3) of tissue 9626 and the mean cell diameter 13 microm. Other estimators (including matrix mass per cell and cell profile density) were also determined. CONCLUSIONS: A comparison of these normal human quantitative data with those published for experimental animals commonly used in orthopaedic research reveals substantial differences, consideration of which in tissue engineering strategies destined for human application are of paramount importance for successful repair. PMID- 12127838 TI - Increased knee joint loads during walking are present in subjects with knee osteoarthritis. AB - OBJECTIVE: This study tests the hypothesis that the peak external knee adduction moment during gait is increased in a group of ambulatory subjects with knee osteoarthritis (OA) of varying radiographic severity who are being managed with medical therapy. Tibiofemoral knee OA more commonly affects the medial compartment. The external knee adduction moment can be used to assess the load distribution between the medial and lateral compartments of the knee joint. Additionally, this study tests if changes in the knee angles, such as a reduced midstance knee flexion angle, or reduced sagittal plane moments previously identified by others as load reducing mechanisms are present in this OA group. DESIGN: Thirty-one subjects with radiographic evidence of knee OA and medial compartment cartilage damage were gait tested after a 2-week drug washout period. Thirty-one normal subjects (asymptomatic control subjects) with a comparable age, weight and height distribution were also tested. Significant differences in the sagittal plane knee motion and peak external moments between the normal and knee OA groups were identified using t tests. RESULTS: Subjects with knee OA walked with a greater than normal peak external knee adduction moment (P=0.003). The midstance knee flexion angle was not significantly different between the two groups (P=0.625) nor were the peak flexion and extension moments (P> 0.037). CONCLUSIONS: Load reducing mechanisms, such as a decreased midstance knee flexion angle, identified by others in subjects with endstage knee OA or reduced external flexion or extension moments were not present in this group of subjects with knee OA who were being managed by conservative treatment. The finding of a significantly greater than normal external knee adduction moment in the knee OA group lends support to the hypothesis that an increased knee adduction moment during gait is associated with knee OA. PMID- 12127839 TI - Biochemical quantification of DNA in human articular and septal cartilage using PicoGreen and Hoechst 33258. AB - OBJECTIVE: To compare two fluorometric assays, utilizing (1) the bisbenzimidazole Hoechst 33258 and (2) PicoGreen, for determining DNA content in human cartilage. METHODS: Human articular and nasal septal cartilage explants were digested using proteinase K. Portions of sample digest were analysed for intrinsic and dye enhanced fluorescence with either Hoechst 33258 or PicoGreen. RESULTS: Intrinsic tissue fluorescence in both articular and septal cartilage increased with age and was prominent at wavelengths used for Hoechst 33258 but relatively low at wavelengths used for PicoGreen. The relative contribution of intrinsic fluorescence to total dye-enhanced fluorescence of human cartilage was markedly greater for Hoechst 33258 (19-57%) than for PicoGreen (2-7%). Thus, in many situations, DNA in human cartilage can be assayed using PicoGreen without the need to correct for intrinsic cartilage fluorescence. The enhancement of fluorescence by each dye was found to be specific for DNA, as shown by fluorescence spectra, >90% sensitivity to DNase, and resistance to RNase. In addition, little or no interference was caused by non-DNA tissue components, since DNA caused an equal enhancement in the absence or presence of proteinase K digested human cartilage, once intrinsic cartilage fluorescence was subtracted. PicoGreen was more sensitive for assaying DNA (0.9ng DNA/ml) than Hoechst 33258 (6ng DNA/ml) and can also be used in a microplate reader. CONCLUSION: PicoGreen can be used in a rapid and sensitive assay to quantify DNA in small samples of human cartilage. PMID- 12127840 TI - Impact loading of articular cartilage. PMID- 12127841 TI - Who should treat patients with peripheral arterial disease - the vascular specialist. PMID- 12127843 TI - External supports and the prevention of neointima formation in vein grafts. AB - AIMS AND METHODOLOGY: the aim of this review is to provide an overview of the aetiology of neointima formation in vein grafts and to highlight the use of an external support to modulate this phenomenon. A systematic literature review was performed via computerised search on MEDLINE, OVID and the Cochrane Library. The search terms initially employed were broad-based; "vein graft", "neointima" and "external stent". Subsequently, more specific search terms were utilised; "perivenous mesh", "external prosthesis" and "varicose vein". Articles from indexed journals relevant to the objective, external venous supports, from the earliest reports in the 1960's to the latest in 2001 were included to obtain an exhaustive list. Reviews, abstracts and proceedings of scientific meetings, case reports and the results of both animal model investigations and human clinical trials in all languages were included. Articles describing an external support employed in both peripheral and aortocoronary bypass investigations were included. PMID- 12127842 TI - What constitutes best medical therapy for peripheral arterial disease? AB - Peripheral arterial disease (PAD) is associated with a high morbidity and mortality, largely from coronary and cerebrovascular disease, which often overshadows the PAD itself. Best Medical Therapy (BMT), comprising smoking cessation, antiplatelet agent use, cholesterol reduction, exercise therapy, and the diagnosis and treatment of hypertension and diabetes mellitus; is evidenced based and can result in significant reductions in cardiovascular risk, as well as some improvement in PAD. Previous data have largely been restricted to patients with coronary artery disease, and their relevance to PAD has been extrapolated. However, data are now starting to become available, such as the Heart Protection Study, with data specific to PAD patients. This article reviews the data regarding the use of BMT in patients with PAD, and based on this, makes recommendations for the use of BMT in this group of patients. PMID- 12127844 TI - The use of ischaemic vessels as prostheses or tissue engineering scaffolds after cryopreservation. AB - OBJECTIVE: to evaluate the condition of organ donor arteries subjected to prolonged cold-ischaemia followed by cryopreservation, for their possible use as vascular grafts. MATERIALS AND METHODS: fresh specimens of human iliac artery from organ donors were used as controls. These arteries were divided into two portions, one of which was cryopreserved in an automated freezer. A further group of arteries was immersed in Wisconsin solution and kept for 4 days at 4 degrees C (cold-ischaemia). After this period, the arteries were also cut into two, and one of these portions was cryopreserved. All the cryopreserved arterial segments were stored for a month and then subjected to automated gradual thawing. The thawed specimens were evaluated by light microscopy, scanning and transmission electron microscopy, immunohistochemical analysis (MMPs, elastin, CD31, von Willebrand factor) and the in situ detection of fragmented DNA (TUNEL method). RESULTS: the most marked changes induced by cryopreservation were partial vessel deendothelialisation and morphological changes in cells of the intima that were in the process of detachment. No significant changes were observed in the medial layer, other than discrete elastic fibre fragmentation. Following cold-ischaemia, the endothelium was the most affected layer, with large denuded areas and exposure of the fibroelastic layer. Increased MMP-2 expression was also noted after cold-ischaemia. When subjected to both cold-ischaemia and cryopreservation, a large proportion of endothelial cells showed positivity for the TUNEL technique, however, no significant difference was observed between the ischaemic and the ischaemic/cryopreserved specimens. CONCLUSIONS: prolonged cold-ischaemia causes some additional damage to the arterial wall compared to cryopreservation alone. However, the structural component of the ischaemic vessel remains in a condition that is suitable for subsequent cryopreservation and use as a vessel substitute or a scaffold for tissue engineering. PMID- 12127845 TI - Duplex scanning or arteriography for preoperative planning of lower limb revascularisation. AB - OBJECTIVE: to compare the accuracy of duplex and angiography for the planning of lower limb revascularisation. PATIENTS AND METHODS: Sixty limbs (82% with critical limb ischaemia) were assessed by means of duplex by one surgeon and by angiography by another in terms of the optimum inflow and outflow sites for arterial bypass. These data were then compared with the final operation performed which was used as the gold standard. Surgeons were blinded to the determinations of the other. RESULTS: surgical plans based on duplex scan and angiography were correct in 77% (40/52) and 79% (41/52), respectively and plans based on the one imaging modality was modified by the other in only 1 and 2 instances. The diagnostic agreement between duplex scanning and arteriography was excellent (Kappa value=0.94, 95% C.I. 0.89-0.98). CONCLUSIONS: the reliability of duplex scanning is comparable to digital angiography in the preoperative planning of lower extremity arterial reconstruction. However neither exam can be considered as the gold standard because intraoperative arteriography needs to be available in a significant number of infrapopliteal procedures. PMID- 12127846 TI - Doppler ultrasound monitoring for detection of microembolic signals in peripheral arteries. AB - OBJECTIVE: to use Doppler ultrasound to detect peripheral microemboluation. METHODS: standard Transcranial Doppler equipment was used to peripheral detect peripheral embolic high intensity transient signals (HITSs) in a pig model following injection of microparticles and atheroma, and in 23 patients who underwent open repair of an abdominal aortic aneurysm (AAA), six patients with blue toe syndrome and 10 age matched healthy subjects. RESULTS: the pig study showed increasing signal intensity with particle size. Particles of 100 (n=24), 200 (n=17), and 400 microm (n=31) elicited 14, 25, 33 dB signals, respectively (p<0.05). During AAA surgery, the intensity (median) of HITSs before clamping (n=226) and after declamping (n=1216) were 14, and 20dB, respectively (p<0.001). Quite a few HITSs were detected after surgery. In patients with blue toe syndrome, a total of 63 HITSs could be detected, and the frequency of HITSs (median: 5.72/30min) was significantly higher than that in patients with AAA before surgery (0.065/30min) (p<0.001). CONCLUSIONS: Doppler ultrasound technique may be a clinically useful test to guide the treatment of patients at risk of distal atheroembolic events. PMID- 12127847 TI - Critical review of non- or minimally invasive methods (duplex ultrasonography, MR and CT-angiography) for evaluating stenosis of the proximal internal carotid artery. AB - OBJECTIVE: to assess the performance of non- or minimally invasive methods (duplex ultrasonography, MR- and CT-angiography) in measuring stenosis of the proximal internal carotid prior to endarterectomy without preoperative intra arterial digital subtraction angiography (DSA). METHODS: systematic review of the literature (five databases, 1990 to February 2001). The value of each imaging technique was studied through its reproducibility and its sensitivity/specificity compared to DSA. RESULTS: sensitivity exceeded 80% and specificity 90% in over two-thirds of the methodologically sound studies, regardless of technique, although direct comparisons between results had to be avoided since the findings originated from different populations. The main drawback of duplex ultrasonography is its levels of reproducibility. In contrast, only a few studies have addressed the reproducibility of MR- and CT-angiography. When the results of duplex and MR-angiography agree, the combination use of these two techniques provides a better diagnosis than either technique taken alone. CONCLUSIONS: all three techniques appear suitable for measuring stenosis of the proximal internal carotid when compared to DSA. PMID- 12127848 TI - The haemodynamic effect of carotid endarterectomy. AB - OBJECTIVES: to assess the haemodynamic effect of carotid artery surgery, and to relate postoperative changes to the state of cerebral circulation before revascularisation. MATERIALS AND METHODS: using transcranial Doppler we studied bilateral middle cerebral artery (MCA) flow velocities before and on 1st day, 2nd or 3rd day and 4th or 5th day and 3 months after carotid surgery in 61 patients. In addition, ipsilateral MCA flow velocity was monitored continuously during surgery. Data were related to the internal carotid artery (ICA) perfusion pressure (cerebral perfusion pressure index, CPPI), measured directly before ICA clamping. RESULTS: postoperatively, MCA flow velocities increased significantly overall (p<0.01), mainly due to pronounced and longer lasting flow velocities in the group of 18 patients with CPPI<0.7 (p<0.05). Flow velocities peaked - absolute as well as relative - on the first postoperative day and then gradually levelled off to reach preoperative values after 4-5 days in patients with high CPPI, whereas MCA flow velocities remained increased in the group of patients with low CPPI. At 3 months flow velocities in both groups were normalised. New neurological symptoms occurred in four patients, who all had low CPPI preoperatively (22% (4/18) vs 0%; Fisher's exact test: p=0.006). CONCLUSION: some degree of hyperperfusion was seen in most patients, but the changes were significantly more pronounced in patients with preoperative hypoperfusion, who also suffered significantly more neurological complications. PMID- 12127849 TI - Cerebral haemodynamic aspects of severe carotid stenosis: asymptomatic vs symptomatic. AB - OBJECTIVES: to compare cerebral haemodynamics in patients with asymptomatic and symptomatic severe (> or =70%) internal carotid artery (ICA) stenosis. METHODS: we assessed 195 consecutive patients, 116 with asymptomatic carotid stenosis (ACS) and 79 with symptomatic carotid stenosis (SCS). Using transcranial Doppler we assessed cerebral vasoreactivity (CVR) following acetazolamide test, the middle cerebral artery flow velocity ratio after/before carotid clamping (mv-MCA ratio), and the carotid back pressure (CBP) during crossclamping. RESULTS: no significant differences between the two groups were demonstrated regarding CVR (47 vs 39%), mv-MCA ratio (50 vs 52%), or CBP (36 vs 44 mmHg). However, in patients with contralateral ICA occlusion all three variables were significantly lower as compared to patients with patent contralateral ICA. Also patients who needed a shunt during surgery had significantly lower values of mv-MCA ratio and CBP. Patients who suffered peri-operative neurologic deficits (n=6; 3%) did not differ from patients who had an uneventful course. CONCLUSIONS: clinical state of ICA stenosis is independent of cerebral haemodynamics. Occluded contralateral ICA is more important for predicting cerebral ischaemia caused by carotid clamping. Finally, none of the haemodynamic parameters showed predictive value for peri operative neurologic morbidity. PMID- 12127850 TI - Carotid endarterectomy with contralateral carotid artery occlusion: is this a higher risk subgroup? AB - OBJECTIVE: to evaluate early and mid-term term results of carotid endarterectomy (CEA) in patient with and without contralateral carotid occlusion. METHODS: between 1996 and 1999, 1324 CEAs were performed. In 82 patients contralateral carotid artery occlusion was present (group I); 1242 patients had patent contralateral carotid (group II). All patients were operated under general anaesthesia, and selective shunting was based on somatosensory evoked potentials (SEPs). Ultrasonographic follow-up was performed at 1, 6 and 12 months and then once a year. Early results and follow-up data were analysed retrospectively. RESULTS: in group I there was a significantly higher incidence of SEPs reduction and shunt insertion; however, there were no differences in terms of perioperative complications. The cumulative stroke and death rate at 30 days in group 1 and group 2 were 2.4% vs 1.4% (p=n.s.), respectively. At a mean follow-up of 15 months there were no differences between the two groups in terms of cumulative symptom-free survival. CONCLUSIONS: the presence of contralateral carotid occlusion caused an increased use of shunt, but not in early complications rates. PMID- 12127851 TI - Does the angiotensin-converting enzyme (ACE) gene polymorphism affect rate of abdominal aortic aneurysm expansion? AB - OBJECTIVES: the tissue renin-angiotensin system (RAS), which plays an important role in vascular structure and function, is regulated in part by an insertion deletion polymorphism of the angiotensin converting enzyme (ACE) gene. We hypothesised that ACE genotype might affect rate of AAA expansion via modulating long-term structural changes associated with RAS activation. METHODS: fifty-eight patients (50 M, mean age 70 years, mean initial aneurysm size 4.3 cm) with current or previous AAA and serial (>3) annual ultrasound measurements of antero posterior AAA size provided a sample of leucocyte DNA for ACE genotyping. AAA expansion rate (cm per year) for individual subjects was calculated by linear regression. RESULTS: median AAA expansion rate was 0.28 cm/year (range 0-1.8 cm/year), and the genotype distribution included DD (n=14), DI (n=29) and II (n=15). Corresponding median AAA expansion rates for each of the three genetic subgroups were 0.22, 0.32 and 0.30 cm/year, respectively (p=0.6, nonparametric). CONCLUSIONS: the wide inter-individual variability in AAA expansion rate is likely to reflect complex genetic and environmental interactions, but the lack of any relationship with ACE genotype suggests that differences in vascular ACE activity in aortic tissue are not major determinants of the variability in rate of AAA dilatation. PMID- 12127852 TI - Enhanced invasive properties exhibited by smooth muscle cells are associated with elevated production of MMP-2 in patients with aortic aneurysms. AB - BACKGROUND: abdominal aortic aneurysms (AAA) are associated with excessive vascular matrix remodelling. Recent findings suggest a systemic overproduction of matrix metalloproteinases-2 (MMP-2) by vascular smooth muscle cells (SMC) may be pivotal aetiologically. SMC migration is facilitated by MMP mediated proteolysis of the basement membrane and extracellular matrix. Our aim was to see if enhanced MMP-2 production by these SMC exhibit increased invasion, in an in vitro model of migration. METHOD: SMC were derived from inferior mesenteric vein (IMV) harvested from patients undergoing aneurysm repair (n=6) or colectomy for diverticulosis (n=6, control). Using a modified Boyden chamber chemotaxis was measured towards platelet derived growth factor (PDGF) and foetal calf serum (FCS) and invasion through a Matrigel layer. MMP-2 production was quantified by ELISA and gelatin zymography. RESULTS: chemoattractant studies demonstrated no difference in the effect of PDGF or FCS between the two populations of SMC. However, invasive studies demonstrated a significant increase in the number of migrating SMC isolated from IMV of AAA patients. Analysis of culture media extracts revealed that this difference was associated with a significant increase in production of MMP-2. CONCLUSION: SMC derived from patients with AAA demonstrate increased invasive properties when compared to a control group. Increased migration appears to be due to overproduction of MMP-2. The enhanced migratory potential of these SMC may lead to extracellular matrix remodelling and subsequent medial disruption demonstrated in the aneurysmal aorta. These data further support evidence of the proteolytic role of MMP-2 in cell migration. PMID- 12127853 TI - Immunoblotting analysis of abdominal aortic aneurysms using antibodies against Chlamydia pneumoniae recombinant MOMP. AB - OBJECTIVES: antibodies against Chlamydia pneumoniae have been associated with atherosclerosis and with expansion of abdominal aortic aneurysms (AAA). C. pneumoniae has been demonstrated in coronary arteries, AAA and the carotid arteries by use of polymerase chain reactions (PCR), immunohistochemical procedures and electron microscopy. However, the correlation between demonstrating C. pneumoniae DNA or antigen in tissue from plaque material or aneurysms and the antibody titres in serum is controversial. The specificity of immunohistochemical procedures is unknown. The aim of this study was to assess the possibility of potential non-specific findings for methods based on immunostaining. MATERIALS AND METHODS: twenty patients undergoing infrarenal AAA repair were studied. Full AAA thickness tissue was collected from the anterior wall of the aneurysm. Analysis was performed using polyacrylamide gelelectrophoresis, immunoblotting and mass spectrometric protein identification. RESULTS: C. pneumoniae antigen was not demonstrated in any of the AAA samples, whereas a major cross-reacting protein was present in all AAA samples. The protein was identified as the human haemoglobin beta chain. CONCLUSION: we were not able to find C. pneumoniae antigens reacting with an anti C. pneumoniae major outer membrane protein (MOMP). Direct detection of C. pneumoniae by immunohistostaining procedures should be interpreted with caution due to potential crossreaction with non chlamydial proteins. PMID- 12127854 TI - Accuracy of duplex scan of internal carotid arteries. PMID- 12127855 TI - Retrograde endovascular management of a mycotic internal carotid artery false aneurysm. PMID- 12127856 TI - Asymptomatic profunda femoris artery aneurysm: diagnosis and rationale for management. PMID- 12127859 TI - Recent progress in the diagnosis of fungal infections in the immunocompromised host. AB - The management of invasive fungal infections has been hampered by the inability to diagnose the infection at an early stage of disease. Although proving the presence of infection by histology and culture remains the cornerstone of the diagnosis, non-culture based methods are becoming available that enable early detection. Molecular diagnosis by PCR appears very promising since fungal DNA can be detected in the blood of infected patients before conventional methods. Furthermore, a broad range of yeasts and molds can be identified to species level. Automation of sample preparation and use of real-time PCR systems will help standardize the procedure and reduce false positive results due to contamination. Promising assays for the detection of fungal antigens in serum have been commercialized, including detection systems for mannan (Candida) and galactomannan (Aspergillus). Circulating antigens can be detected at an early stage of infection, often before the onset of clinical symptoms. Antigen detection is limited to detecting only one genus and not enabling speciation. Furthermore, both PCR and antigen detection can be used to monitor the response of patients to treatment with anti-fungal agents. Although prospective screening of high-risk patients for the presence of circulating markers of fungal infection appears to be an appropriate strategy, studies are needed to help to establish the optimal approach to managing invasive fungal infections that incorporates the benefits of non-culture based methods. PMID- 12127860 TI - EGF receptor targeting in therapy-resistant human tumors. AB - The development of resistance against cytotoxic or endocrine therapy limits the number of chemotherapeutic compounds used in the clinic. The receptor for EGF (EGFR) is not only involved in survival signaling, cell migration, metastasis formation and angiogenesis, but also confers reduced responses of tumor cells towards cytotoxic compounds or radiation. Clinical trials designed to combine EGFR inhibitors with standard chemo- or radiation therapy have been successful. Elucidation of some of the molecular mechanisms of EGFR-mediated chemoresistance may lead to novel treatment approaches against molecules linked to EGFR signal transduction. In human breast carcinomas, the presence of EGFR correlates with lack of response towards anti-estrogen therapy suggesting the concomitant inhibition of both the receptors for estrogen and EGF to improve breast cancer therapy. PMID- 12127861 TI - Determinants of resistance to 2',2'-difluorodeoxycytidine (gemcitabine). AB - The inherent or induced resistance of tumors to cytostatic agents is a major clinical problem. In this review, we summarize the pre-clinical mechanisms of acquired and inherent resistance to the fluorinated deoxycytidine analog gemcitabine (2',2'-difluorodeoxycytidine, dFdC, Gemzar((R))), which has proven activity in non-small cell lung carcinoma, pancreatic and bladder cancer. Extensive research has been performed to elucidate the complex mechanism of action of this relatively new drug. Gemcitabine requires phosphorylation to mono , di- and triphosphates to be active. Similar to the structurally and functionally related deoxycytidine analog ara-C, the first, crucial step in phosphorylation is catalyzed by deoxycytidine kinase (dCK). However, in contrast to ara-C, gemcitabine has multiple intracellular targets; up- or down-regulation of these targets may confer resistance to this drug. Resistance is associated with altered activities of enzymes involved in the metabolism of the drug, of target enzymes, and of enzymes involved in programmed cell death. However, the only strong correlations with gemcitabine sensitivity are dCK activity and dFdCTP pools, with a potential important role for ribonucleotide reductase. PMID- 12127862 TI - Emerging new therapies for chemotherapy-resistant cancer using adenoviral vectors. AB - The treatment of cancer by genetic manipulation of either the tumor itself or the patient as a whole offers new avenues for the treatment of otherwise refractory cancers. Gene therapy seeks to correct underlying genetic defects in malignant tissue or to augment the host defense response or to promote selectivity of other therapies. Many innovative and exciting genetic targets have been recently identified. However, the field as a whole is still constrained by limitations of gene delivery. The most common vector for gene delivery is modified adenovirus. In this review, we survey a sampling of current therapeutic approaches that depend upon adenoviral delivery vehicles and outline the advantages and disadvantages of this vector system. PMID- 12127863 TI - Cryptosporidiosis and the challenges of chemotherapy. AB - Cryptosporidium parvum is a protozoan parasite that infects the epithelial cells of the small intestine causing diarrheal illness in humans. Cryptosporidium has a worldwide distribution and is considered an emerging zoonosis. Despite intensive efforts to develop workable experimental models, and the evaluation of over 200 chemotherapeutic agents, adequate therapies to clear the host of these parasites are still lacking. The reasons for the lack of drug efficacy are probably manifold and may include the unusual location of the parasite in the host cell, distinct structural and biochemical composition, or its ability to either block import or rapidly efflux drug molecules. Understanding some of the basic mechanisms by which drugs are transported to the parasite and identifying unique targets is a first step in developing effective therapeutic agents. PMID- 12127866 TI - Changes and challenges--the world post-Gleevec (Glivec). PMID- 12127867 TI - 'Omic' and hypothesis-driven research in the molecular pharmacology of cancer. AB - Given the layered, evolutionary complexity of biological systems, it will not be possible to understand them comprehensively on the basis of hypothesis-driven research alone. Likewise, it will not be possible to do so solely through 'omic' studies of genes, proteins and other molecules in aggregate. The two modes of research are complementary and synergistic. The twin complexities of cancer biology and drug discovery demand such synergy as a basis for the next generation of oncopharmacomic research. PMID- 12127868 TI - Using cancer genetics to guide the selection of anticancer drug targets. AB - The genetic alterations that cause cancer are coming into view. Genes that are recurrently mutated in a particular form of human cancer flag the proteins (or molecular pathways) that are critical for the evolution of that malignancy. The first generation of anticancer agents prospectively guided by these principles, for which imatinib mesylate is a prototype, inhibit the biochemical activities that result from gain-of-function oncogenic mutations. Advances in somatic cell genetics and chemical biology should facilitate the development of a second generation of agents that will inhibit proteins that are selectively required for survival in the context of specific cancer-causing mutations, whether loss-of function or gain-of-function. PMID- 12127870 TI - Epidermal growth factor receptor tyrosine kinase inhibitors. AB - Clinical trials of selective small-molecule inhibitors of epidermal growth factor receptor tyrosine kinase activity have shown that these targeted inhibitors of proliferative signal transduction provide well-tolerated antitumour activity in patients. Preclinical pharmacology studies illustrate the potential use of these new cancer therapeutics in combination with chemotherapy, radiotherapy and hormone therapy, and in chemoprevention, in a spectrum of solid human tumours. PMID- 12127869 TI - Protein tyrosine kinase inhibitors: new treatment modalities? AB - Protein tyrosine kinases (PTKs) have been recognized as attractive cell-signaling targets for drug discovery in the treatment of cancer and other diseases. Most of the PTK inhibitors are small molecules, designed to compete for, or nearby, the ATP-binding site, and are currently in phase I-III clinical trials, mainly for oncological indications. Recent efforts focused on the synthesis of selective PTK inhibitors have generated several promising clinical candidates, which recently culminated in the approval of Gleevec, the first kinase inhibitor registered for the treatment of chronic myeloid leukemia and gastrointestinal stromal tumors. PMID- 12127871 TI - Farnesyltransferase inhibitors: promises and realities. AB - Farnesyltransferase inhibitors have recently shown clinical efficacy against leukemias, gliomas and even non-small-cell lung cancers, especially when administered in combination with taxanes. It is possible that the critical target downstream of farnesyltransferase responsible for these effects is not either Ras or RhoB, the two most cited possibilities - but the hunt is on. PMID- 12127872 TI - Inhibitors of integrins. AB - The inhibition of integrins--cell surface receptors with a crucial role in angiogenesis, tumour cell survival, invasion and metastases--has centred on the alpha(v)beta3 integrin. Work has culminated in two antagonists that are in clinical trials as cancer therapeutics. Other integrins appear to be candidate targets in the light of gene knockout studies. Surprisingly, genetic alpha(v)beta3 ablation did not confirm the pertinence of the use of alpha(v)beta3 antagonists. However, these apparent discrepancies could be explained by the new finding that this integrin has a role as a cell survival sensor, limiting rather than promoting angiogenesis. Accumulating data on the role of integrins and the mechanism of action of pharmacological antagonists will help to develop and apply an efficient anti-integrin therapy in cancer. PMID- 12127873 TI - Drug inhibition of angiogenesis. AB - Cancer is one of the leading causes of death in the Western world. Though advances in cancer therapy and diagnosis have considerably improved life expectancy, the overall survival rate of patients still remains poor - disseminated cancer at presentation and acquisition of tumour resistance are two reasons for this. Angiogenesis is one of the crucial steps in the pathogenesis of tumours. Drug inhibition of angiogenesis is an area of intense research and at least 10000 cancer patients worldwide have received some form of experimental antiangiogenic therapy. More than 300 angiogenesis inhibitors have been discovered to date; 80 antiangiogenic drugs are currently in clinical trials, 12 of which target the key angiogenic factor vascular endothelial growth factor. A convincing regression of tumours has been reported for drugs against this target. Antiangiogenic therapy has raised the hopes both of cancer sufferers and of the physicians looking after them. A concerted international effort by cancer researchers and the pharmaceutical industry will help to further develop this novel treatment strategy for cancer. PMID- 12127874 TI - Telomeres and telomerases as drug targets. AB - Recent advances in telomerase inhibition have been achieved by using antisense oligonucleotides and ribozymes to target the telomerase mRNA or the telomerase RNA template. Also, small molecules are potent catalytic inhibitors of telomerase. However, therapeutic regimes incorporating these agents will be challenging to implement in the clinic because of their delayed effectiveness. Drugs that directly bind to the telomeres and stabilize secondary DNA structures such as G-quadruplexes are also potent inhibitors of telomerase and disrupt telomere structure. These G-quadruplex-interactive drugs could feasibly be used in synergy with more conventional cytotoxic agents to bring about more immediate responses in cancer cells that are less dependent upon telomere length. Recently, an emerging possible novel use of G-quadruplex-interactive drugs employs their ability to target G-quadruplexes in promoter regions of genes (such as c-MYC), which then serves to repress the production of the human telomerase reverse transcriptase protein. PMID- 12127875 TI - Immunomodulation. PMID- 12127876 TI - Immunomodulatory effect of antidepressants. AB - Basic and clinical studies have provided evidence that the biophase level of monoamines, determined by the balance of their release and uptake, is involved in the pathophysiology and treatment of depression, whereas other arguments cite the role of inflammatory mediators in the etiology of psychiatric disorders. A bidirectional interaction between the monoamine system and the inflammatory system might explain the concurrent therapeutical efficacy and immunomodulatory features of antidepressants. PMID- 12127877 TI - Therapeutic use of interleukin-2 in HIV-infected patients. AB - Highly active antiretroviral therapy has improved the morbidity and survival of patients with the human immunodeficiency virus (HIV) infection. Currently available antiretroviral agents can suppress viral replication and partially reverse cellular immunity defects in HIV patients. Immune-based therapy with interleukin-2 when used as adjunctive therapy to antiretroviral therapy may further improve immune responses, as demonstrated by an increase in CD4(+) T lymphocyte counts in recent clinical trials. PMID- 12127879 TI - CD40-CD40L interaction in Alzheimer's disease. AB - Increasing evidence supports a role of the CD40 receptor-CD40 ligand (CD40-CD40L) interaction in the pathogenesis of Alzheimer's disease (AD). It has previously been shown that this dyad acts synergistically with the Alzheimer amyloid-beta peptide to promote microglial activation. Reactive microglia produce potentially neurotoxic substances such as tumor necrosis factor alpha and the reactive oxygen species nitric oxide, which can induce bystander neuronal injury at high levels. When a transgenic mouse model of AD is crossed with an animal deficient in CD40L, the resulting phenotype is deficient in the gliosis observed in a mouse model of AD in which CD40L is present. Additionally, these crossed animals have complete absence of AD-like neuronal Tau hyperphosphorylation, a marker of the preneuronal tangle pathology in AD patients. This suggests that the CD40-CD40L system is a critical enhancer of microglial activation in an AD transgenic mouse model and that such activation is associated with an increase in a key indicator of neuronal stress. Conversely, the finding that reduced CD40-CD40L interaction is associated with reduced chronic microgliosis and Tau hyperphosphorylation supports the view that, in general, mechanisms that reduce microgliosis will be beneficial in AD. PMID- 12127878 TI - Adenosine: a potential mediator of immunosuppression in multiple organ failure. AB - Multiple organ failure following a variety of insults, including, trauma, shock and pancreatitis, is the cause of 50-80% of all deaths in surgical intensive care units. In most patients, infections secondary to a general immunosuppressive state serve to trigger the development of multiple organ failure. This immunosuppressive state may be a consequence of excessive release of adenosine into the extracellular space, as adenosine has multiple immunosuppressive effects. Activation of adenosine receptors on immune cells inhibits the production of proinflammatory cytokines such as tumor necrosis factor alpha and interleukin (IL)-12, and increases the production of the anti-inflammatory cytokine IL-10. In addition, adenosine receptor activation appears to suppress cellular immunity by decreasing T helper cell (Th)1 and increasing Th2 responses. A deeper understanding of the role of adenosine in multiple organ failure may facilitate the development of adenosine-based therapeutic strategies. PMID- 12127880 TI - Immunomodulation in stem-cell transplantation. AB - Acute graft-versus-host disease is a complication that affects 30-60% of patients undergoing allogeneic stem-cell transplantation. The standard for prophylaxis for graft-versus-host disease has historically been the combination of cyclosporine and methotrexate. Recently, tacrolimus has been used more frequently and current studies are exploring the potential of mycophenolate mofetil. There is little published experience with the use of sirolimus in prophylaxis or treatment but studies are ongoing. There have been significant advances recently in the treatment of steroid-refractory acute graft-versus-host disease. Historically, antithymocyte globulin was used when patients did not respond to the steroid treatment. New monoclonal antibodies such as daclizumab, and tumor necrosis factor alpha inhibitors such as infliximab are producing more promising results. Chronic graft-versus-host disease continues to be a major complication of stem cell transplantation, affecting 35-50% of patients. Finding effective treatments for chronic graft-versus-host disease other than steroids continues to be a challenge. PMID- 12127881 TI - Serine/threonine protein phosphatases in apoptosis. AB - The importance of phosphorylation and dephosphorylation in intracellular signaling pathways has long been recognized, although attention has focused mainly on kinases. Recent studies have highlighted the importance of serine/threonine protein phosphatases in many processes including apoptosis. The phosphorylation state of antiapoptotic (Bcl-2, Bcl-X(L)) and proapoptotic (BAD, Bid, Bik) Bcl-2 proteins regulates their cellular activity and, therefore, cell survival and cell death. For example, dephosphorylation of BAD by the protein phosphatases PP1, PP2A and PP2B allows BAD to interact with Bcl-X(L) and initiate cell death. Caspases are also important in cell death and phosphorylation/dephosphorylation of caspases themselves, their targets and their regulators modulates apoptotic pathways. The activity of serine/threonine protein phosphatases needs further study, but it is clear that these enzymes are potential targets for novel therapeutics with applications in many diseases, including cancer, inflammatory diseases and neurodegeneration. PMID- 12127882 TI - Using genetic variation to study immunomodulation. AB - The generation of a draft sequence of a human genome has led to the identification of millions of common variants, known as single nucleotide polymorphisms, which constitute a resource for studying complex diseases. Currently, high-density maps of variants in candidate genes, chromosomal regions or the entire genome should encourage investigation of determinants of human immune response, using quantitative analysis. Ultimately, this approach should identify novel targets for therapeutic intervention. PMID- 12127883 TI - Ethical challenges in conducting pediatric environmental health research. Introduction. PMID- 12127884 TI - Appropriate risk exposure in environmental health research. The Kennedy-Krieger lead abatement study. PMID- 12127885 TI - What are we doing when we are doing research on humans? PMID- 12127886 TI - The risk/benefit ratio in pediatric environmental research. PMID- 12127887 TI - What is not found in the spreadsheets. PMID- 12127888 TI - Studies of pervasive toxic contaminants in children: staying the ethical course. PMID- 12127889 TI - Research ethics in pediatric environmental health: lessons from lead. PMID- 12127890 TI - Confronting the ethical challenges of environmental health research. PMID- 12127891 TI - Ethical considerations of research involving minorities, the poorly educated and/or low-income populations. PMID- 12127892 TI - Commentary on the Kennedy Krieger Institute Lead Paint Repair and Maintenance Study. PMID- 12127893 TI - Growth from birth onwards of children prenatally exposed to drugs: a literature review. AB - Reductions in birth weight and length have been independently attributed to prenatal exposure to alcohol, cigarettes and cocaine. While pregnant women often use multiple substances, studies have not consistently controlled for exposure to other agents or other important differences in maternal lifestyle associated with the use of these substances. Despite these difficulties, the preponderance of evidence suggests that prenatal alcohol and cocaine independently reduce birth measurements. This review synthesizes the scientific literature focusing on prenatal exposures and the relationship to child growth. First examined are studies that investigated the link between prenatal exposures and birth weight and length, followed by the effects of these substances on childhood growth. Studies vary in the number of subjects, cohort characteristics, measurement of exposure and control for potential confounders. Differences in sample characteristics and size, as well as degree of statistical control for potential confounders and the examination of moderating characteristics, have led to differing conclusions regarding the long-term effect of prenatal substance exposure on growth. Large-scale, well-designed studies are needed to clearly examine the unique contribution of both varying prenatal exposures and the magnitude and timing of these exposures on childhood growth deficits. PMID- 12127894 TI - Birth to age 7 growth of children prenatally exposed to drugs: a prospective cohort study. AB - Prenatal exposure to cocaine, alcohol, and cigarettes has been linked to decreased birth weight and length. Unclear, however, is whether growth deficits persist into childhood. Women who were pregnant, African-American, not HIV positive, and who delivered singleton infants were extensively screened throughout pregnancy for cocaine, alcohol, cigarette, and other illicit drug use. Of the approximately 1100 eligible subjects, 665 families were located at a 7 year postbirth follow-up and 540 participated. After appropriate control for potential confounders and prenatal exposures, prenatal exposure to cocaine, alcohol, and cigarettes each independently predicted birth weight and length. At age 7, prenatal cocaine exposure was significantly related to height deficits after accounting for other prenatal exposures and significant confounders. Children at age 7 exposed to cocaine in utero were up to 1 in. shorter and twice as likely to fall below the 10th percentile in height as the control children after accounting for other significant confounders including other prenatal exposures. Maternal age moderated the relation between prenatal exposures and child growth. Children born to women over 30 and exposed to cocaine were up to 2 in. shorter and four times more likely to have clinically significant height deficits at age 7. Children of older women and exposed to moderate-to-high levels of alcohol prenatally were up to 14 lb lighter and five times more likely to fall below the 10th percentile in weight. Similar growth restriction was not associated with prenatal exposures for children born to younger mothers. These outcomes add to the growing body of literature detailing long-term effects of prenatal drug exposure, suggesting differential effects for cocaine and alcohol, and indicating that maternal age may moderate these effects. Mechanisms for growth restriction and failure of catch-up under conditions of prenatal exposures are presented, suggesting further study of these developmental outcomes. PMID- 12127895 TI - Effect of prenatal haloperidol exposure on behavioral alterations in rats. AB - Pregnant Charles-Foster rats were exposed to haloperidol (HAL), a neuroleptic drug that binds to and blocks dopamine (DA) receptor subtypes at a dose of 2.5 mg/kg body weight (intraperitoneally) from Gestation Day (GD) 12 to 20. The animals from both treated as well as vehicle control groups were allowed to deliver on GD 21. The offspring culled at birth on the basis of sex and weight were subjected to behavioral tests at the age of 8 weeks. The HAL-treated rat offspring showed a significant increase in anxiogenic behavior on the open field, elevated plus-maze and elevated zero-maze tests when compared with the vehicle treated (control) rat offspring of the same age group. These findings suggest that prenatal exposure to HAL during a critical period of brain development leaves a lasting imprint on the brain, resulting in abnormal anxiety states, possibly through dopaminergic neurotransmission mechanisms. PMID- 12127896 TI - Differential susceptibility of rats and guinea pigs to the ototoxic effects of ethyl benzene. AB - The present study was designed to compare the ototoxic effects of volatile ethyl benzene in guinea pigs and rats. Rats showed deteriorated auditory thresholds in the mid-frequency range, based on electrocochleography, after 550-ppm ethyl benzene (8 h/day, 5 days). Outer hair cell (OHC) loss was found in the corresponding cochlear regions. In contrast, guinea pigs showed no threshold shifts and no OHC loss after exposure to much higher ethyl benzene levels (2500 ppm, 6 h/day, 5 days). Subsequently, a limited study (four rats and four guinea pigs) was performed in an attempt to understand these differences in susceptibility. Ethyl benzene concentration in blood was determined in both species after exposure to 500-ppm ethyl benzene (8 h/day, 3 days). At the end of the first day, blood of the rats contained 23.2+/-0.8-microg/ml ethyl benzene, whereas the concentration in guinea pig blood was 2.8+/-0.1 microg/ml. After 3 days, the concentration in both species decreased with respect to the first day, but the ethyl benzene concentration in rat blood was still 4.3 times higher than that in guinea pig blood. Thus, the difference in susceptibility between the species may be related to the ethyl benzene concentration in blood. PMID- 12127897 TI - Schedule-controlled behavior in rats exposed perinatally to the PCB mixture Aroclor 1254. AB - Exposure to polychlorinated biphenyls (PCBs) has been shown to detrimentally affect learning and memory in children as well as schedule-controlled behavior in experimental animals. The objective of the present series of experiments was to extend research into the effects of PCBs on behavior maintained under both short (30 s) and long (5 min) fixed-interval (FI) schedules as well as an FI 3-min with reinforcement omission. Long-Evans rats were exposed to 0 or 6 mg/kg/day Aroclor 1254 (A1254) via oral gavage from Gestation Day 6 (GD 6) through Postnatal Day 21 (PND 21). At approximately PND 90, acquisition and steady-state performance were assessed under a series of FI reinforcement schedules consisting of FI 30-s, FI 5 min, and FI 3-min with 33% of the scheduled reinforcers omitted. Performance measures included index of curvature (IOC), response rate, and postreinforcement pause (PRP). There were no effects of A1254 on the acquisition of behavior under the FI 30-s schedule. Subsequently, there was an initial decrease in response rate and IOC and an increase in PRP following the transition from FI 30-s to the FI 5-min; there were, however, no treatment-related effects on any measure. During the reinforcement-omission procedure, there was an increase in the rate of responding and a decrease in IOC and PRP following omission intervals irrespective of treatment. These data are inconsistent with previous findings and suggest that perinatal A1254 exposure in the rat does not disrupt temporally organized behavior. PMID- 12127899 TI - Long-term effects of developmental exposure to zidovudine on exploratory behavior and novelty discrimination in CD-1 mice. AB - Long-term changes in exploratory, social and agonistic behavior have been reported in rodents following developmental exposure to zidovudine (AZT), an agent commonly administered to pregnant seropositive women and their neonates to prevent HIV-1 transmission. The present study evaluates the effects of either prenatal or prolonged AZT treatment on spatial and nonspatial novelty discrimination in mice, using an open-field test with four objects, in which responses to both spatial rearrangement of familiar objects and object novelty are assessed. AZT (160 mg/kg) or Saline was given orally twice daily to pregnant mice from gestational days (GD) 10 to 19 (Experiment 1) or from GD 10 to lactation day 10 (Experiment 2). Offspring of both sexes were tested on postnatal day (PND) 28, 45 or 70. Depending on treatment schedule, AZT altered different behavioral responses, males being more affected than females. The prenatal treatment (Experiment 1) reduced exploration of the objects at all ages considered and increased wall and top rearing at ages 45 and 70. Following prolonged treatment (Experiment 2), AZT offspring were markedly more active than controls and displayed more wall rearing at age 70 while showing lower grooming frequency at all ages. Both AZT and control mice failed to respond to object rearrangement at adulthood, a discrepancy from previous data, which is discussed in relation to perinatal stress effects. PMID- 12127898 TI - Methanol-induced neurotoxicity in pups exposed during lactation through mother: role of folic acid. AB - Role of folic acid on methanol-induced neurotoxicity was studied in pups at Postnatal Day (PND) 45 exposed to methanol (1%, 2% and 4%, v/v) during lactation through mothers maintained on folic acid-deficient (FD) and folic acid-sufficient (FS) diet. A gradual loss in the body weight gain was observed in the pups exposed to 2% and 4% methanol in the FD group, while FS group exhibited this alteration only at 4% exposure. The assessment of spontaneous locomotor activity (SLA) showing a significant increase in the distance travelled was observed in the 2% and 4% methanol-exposed groups in both the FS and FD animals when compared with their respective controls, but the effect was more marked in the FD group. A significant decrease in the conditioned avoidance response (CAR) was observed in pups exposed to 2% and 4% methanol in the FD group at PND 45. The results also suggest that disturbances in dopaminergic and cholinergic receptors were more pronounced in the FD group as compared to the FS group. A significant decrease in striatal dopamine levels was also observed in the FD group at 2% and 4% methanol exposure, while in the FS group, a significant decrease was exhibited only at 4% methanol exposure. An aberrant increase in the expression of Growth-Associated Protein (GAP-43), a neuron-specific growth-associated protein was observed in pups in the FD group exposed to 2% and 4% methanol, while an increase in the expression of GAP-43 in the FS group was found only at 4% methanol exposure in the hippocampal region as compared to their respective controls. Results suggests that methanol exposure during growth spurt period adversely affects the developing brain, the effect being more pronounced in FD rats as compared to FS rats, suggesting a possible role of folic acid in methanol-induced neurotoxicity. PMID- 12127900 TI - Lead and conditioned fear to contextual and discrete cues. AB - Male Fischer 344 rats received either tap water or water containing 250 ppm lead for 90 days prior to training in either Pavlovian fear conditioning or consummatory contrast, an aversive reward reduction paradigm. In Experiment 1, lead-exposed and -unexposed rats were trained in operant chambers over a 6-min session. After 3 min elapsed, three tone-shock pairings were presented over the remainder of the session. Rats then received 7 days of extinction training in an identical procedure with only tones presented, no shocks. Lead-exposed rats exhibited greater behavioral suppression to both the contextual and auditory cues that predicted shock. In Experiment 2, rats were placed in operant chambers daily and allowed to consume either a 15% or a 5% fructose solution for 7 days. On Day 8, the rats consuming the 15% fructose solution were shifted to the 5% solution for 3 days. Lead-exposed rats did not differ from their controls in either initial consumption of either solution or in the suppression of their consumption after shifting to the 5% solution. Taken together, these findings suggest that lead impairs the extinction of fear conditioning and this finding is not due to a nonspecific increase in aversive emotionality. PMID- 12127901 TI - Lead and spatial vs. cued open field performance. AB - Male Fischer 344 rats received chronic exposure to either water containing 250 ppm lead or tap water. On the first day of the study, rats were allowed to habituate to a 1-m(2) open field arena with a rectilinear grid pattern of food wells on the floor for a 2-min session. On the following 7 days, half the rats were trained (four trials per day, 2-4-min intertrial interval) to find a food location based on extra-maze spatial cues and the other half were trained to find a food location based on a discrete intra-maze cue placed over the baited food well. While lead did not appear to significantly affect motor activity during the habituation phase, lead-exposed spatially trained rats exhibited superior acquisition and performance of the food-rewarded task compared to their controls and their cue-trained lead-exposed and counterparts. Furthermore, by the last day of training, Day 7, lead significantly reduced the relative amount of time spent on the periphery of the maze in spatially and cued-trained rats. PMID- 12127902 TI - The effect of administering ethanol as single vs. divided doses on blood alcohol levels in the rat. AB - The magnitude of peak blood alcohol levels (BALs) and duration of exposure are critical determinants of alcohol's effects. This technical report provides BAL data for different doses (2, 3, 4, 5, or 6 g/kg) administered as single (at 12:00 h) or dual doses (at 07:00 and 12:00 h) of alcohol when administered by intubation at several time points (0.5, 1, 2, 4, 6, and 24 h after the 12:00 h intubation) in male rats. Administration of the highest dose in a single intubation resulted in the highest peak BALs, a later peak in BAL, and a longer latency to return to 0 mg% ethanol in the blood. Other combinations resulted in different profiles. The differences are explained in terms of "first-pass" effects relating to alcohol's elimination via the liver. These findings should be of practical use to researchers using intubation as their method of alcohol administration, especially when the timing and magnitude of peak BAL are critical. PMID- 12127903 TI - Kinetics and organotropy of some polyfluorinated dibenzo-p-dioxins and dibenzofurans (PFDD/PFDF) in rats. AB - While polychlorinated dibenzo-p-dioxins and dibenzofurans (PCDD/PCDF) and the corresponding polybrominated congeners must be considered as animal teratogens and carcinogens, little information is available on corresponding polyfluorinated compounds (PFDD/PFDF). Kinetic studies on a few fluorinated dibenzo-p-dioxins and dibenzofurans revealed a rapid elimination, suggesting a much lower toxicity than the corresponding polychlorinated and polybrominated congeners. In order to obtain further clues on the possible toxicity, the kinetics and organ distribution (in liver, thymus and adipose tissue) of a PFDD/PFDF-mixture were studied in Wistar rats after intravenous application. The congeners investigated included four of the 2,3,7,8-substituted, and four of the not-2,3,7,8-substituted dibenzo-p-dioxins, as well as two dibenzofurans. The main result of our studies is the finding that the concentration in the thymus of several of the 2,3,7,8 substituted PFDD/PFDF greatly exceeded that in hepatic tissue. An organotropy quite different from that of the other polyhalogenated congeners must be expected, immunosuppressive effects presumably being the predominant ones. Overall, the elimination half-life of all the PFDD/PFDF studied is considerably shorter than that of the corresponding polychlorinated or polybrominated congeners, in the rat, suggesting a much lower toxicity in this species. No information is available for other species, e.g. nonhuman primates or humans. PMID- 12127904 TI - Metabolic profile of NO-flurbiprofen (HCT1026) in rat brain and plasma: a LC-MS study. AB - Male rats were given the nitrooxybutyl ester of flurbiprofen HCT1026 (15 mg/Kg) by oral administration and plasma and brain levels of the parent drug and its potential metabolites (HCT1027 and flurbiprofen) were determined at different times post-administration by a validated HPLC method (UV-DAD detection; LOQ: 0.13 nmoles/ml and 0.3 nmoles/g respectively in plasma and brain tissue). Structural confirmation of the analytes was achieved by MS monitoring of their de-protonated (negative ion mode) or cationized/protonated (positive ion mode) molecular ions and of the relative fragment ions obtained by collision-induced dissociation (CID) experiments. The results indicate that flurbiprofen is the only metabolite found at measurable levels in both plasma and brain, while HCT1026 or its de nitrated metabolite HCT1027 were always below the limit of detection at all the observation times. The same was observed after administration of the higher dose of HCT1026 (100 mg/Kg, i.p.). In orally-treated animals the time-course of flurbiprofen formation strictly parallels that of NOx (nitrite/nitrate) in plasma but not in brain, where the levels were always in the range of the controls. These data indicate that the NO molecule is not released from the parent drug within the brain. PMID- 12127905 TI - Neuroregulative mechanism of hypothalamic paraventricular nucleus on gastric ischemia-reperfusion injury in rats. AB - A rat model of gastric ischemia-reperfusion injury (GI-RI) was established by clamping the celiac artery for 30 min and allowing reperfusion for 1 h, on which the regulatory effect of the paraventricular nucleus (PVN) and its neural mechanisms were investigated. The results were: 1. Electrical stimulation of the PVN obviously attenuated the GI-RI. Microinjection of L-glutamic acid into PVN produced an effect similar to that of PVN stimulation. 2. Electrolytic ablation of the PVN aggravated the GI-RI. 3. Nucleus tractus solitarius (NTS) ablation could eliminate the protective effect of electrical stimulation of PVN on GI-RI. 4. Hypophysectomy did not alter the effect of electrical stimulation of PVN. 5. Vagotomy or sympathectomy both could increase the effect of PVN stimulation on GI RI. These results indicate that the PVN participates in the development of GI-RI as a specific area in the CNS, exerting protective effects on the GI-RI. The NTS and vagus and sympathetic nerve may be involved in the regulative mechanism of PVN on GI-RI, but the PVN mechanism here is independent of the PVN-hypophyseal pathway. PMID- 12127906 TI - Influence of different neural systems on the secretion of sex hormones in potassium deficient mice. AB - The influence of different neural systems that modulate GnRH secretion by hypothalamic neurons was investigated in mice exposed to hypokalemic conditions, in which the pulsatile release of GnRH has been shown to be altered and associated with a significant decrease of plasma sex steroids. Our results demonstrate that the potentiation of the inhibitory pathways mediated by opiates and GABA may be implicated in the decrease of sex hormones secretion produced by hypokalemia since treatment with higher doses of naloxone or flumazenil are required to restore progesterone or testosterone levels in potassium deficient mice. The combination treatment of prazoxin and naloxone suggests that the inhibitory action of opiates take place through its action on noradrenergic neurons. It is also possible that the inhibition of GnRH release could be due to a decrease in the tonic stimulatory action of noradrenergic pathway implicated in the control of GnRH release. Our results also reveal that it is unlikely that the glutamatergic system may play any relevant direct role in the decrease of sex steroid secretion observed in potassium deficient mice. Finally, these results together with the normal pattern of estradiol levels found along the estrus cycle in potassium deficient mice indicate that factors different from estradiol and acting on neural systems implicated in the regulation of GnRH-secreting neurons participate in the generation of the preovulatory surge of GnRH. PMID- 12127907 TI - Divergent effect of mometasone on human eosinophil and neutrophil apoptosis. AB - Mometasone is a potent synthetic glucocorticoid, which is under development as an inhaled preparation for the treatment of asthma. Previous studies have suggested that glucocorticoids have direct effects on human eosinophil and neutrophil apoptosis. The present study was designed to characterize the effects of mometasone on constitutive apoptosis and cytokine-afforded survival in isolated human eosinophils and neutrophils. The isolated eosinophils or neutrophils were cultured in vitro, and apoptosis was assessed by flow cytometric analysis of relative DNA content, by annexin-V binding and morphological analysis. Mometasone enhanced constitutive human eosinophil apoptosis in a concentration-dependent manner. The maximal enhancement of eosinophil apoptosis was 2.1-fold with an EC(50) value of 5.63 +/- 2.33 nM. This enhancing effect was reversed by the glucocorticoid receptor antagonist, mifepristone. In the presence of added cytokines, mometasone reversed tumor necrosis factor -alpha-induced eosinophil survival but not that afforded by interleukin -5. In contrast, mometasone inhibited human neutrophil apoptosis in a concentration-dependent manner. The maximal inhibition of neutrophil apoptosis was 50% with an EC(50) value of 0.17 +/- 0.03 nM. The inhibitory effect was partly reversed by mifepristone. In the presence of added cytokines, mometasone further enhanced neutrophil survival induced by the granulocyte-macrophage colony-stimulating factor and leukotriene B(4). The present data suggests that mometasone has opposite effects on apoptosis of human eosinophils and neutrophils at clinically relevant drug concentrations via an effect on glucocorticoid receptor. PMID- 12127908 TI - Effects of exogenous inositol hexakisphosphate (InsP(6)) on the levels of InsP(6) and of inositol trisphosphate (InsP(3)) in malignant cells, tissues and biological fluids. AB - InsP(6) is abundant in cereals and legumes. InsP(6) and lower inositol phosphates, in particular InsP(3), participate in important intracellular processes. In addition, InsP(6) possess significant health benefits, such as anti cancer effect, kidney stones prevention, lowering serum cholesterol. Because of the insensitivity of existing methods for determination of non-radiolabeled inositol phosphates, little is known about the natural occurrence, much less on the concentrations of InsP(6) and InsP(3) in biological samples. Using gas chromatography-mass detection analysis of HPLC chromatographic fractions, we report a measurement of unlabeled total InsP(3) and InsP(6) (a) as they occur within cells culture, tissues, and plasma, and (b) their changes depending on the presence of exogenous InsP(6). When rats were fed on a purified diet in which InsP(6) was undetectable (AIN-76A) the levels of InsP(6) in brain were 3.35 +/- 0.57 (SE) micromol.kg(-1) and in plasma 0.023 +/- 0.008 (SE) micromol.l(-1). The presence of InsP(6) in diet dramatically influenced its levels in brain and in plasma. When rats were given an InsP(6)-sufficient diet (AIN-76A + 1% InsP(6)), the levels of InsP(6) were about 100-fold higher in brain tissues (36.8 +/- 1.8 (SE)) than in plasma (0.29 +/- 0.02 (SE)); InsP(6) concentrations were 8.5-fold higher than total InsP(3) concentrations in either plasma (0.033 +/- 0.012 (SE)) and brain (4.21 +/- 0.55 (SE)). When animals were given an InsP(6)-poor diet (AIN 76A only), there was a 90% decrease in InsP(6) content in both brain tissue and plasma (p < 0.001); however, there was no change in the level of total InsP(3). In non-stimulated malignant cells (MDA-MB 231 and K562) the InsP(6) contents were 16.2 +/- 9.1 (SE) micromol.kg(-1) for MDA-MB 231 cells and 15.6 +/- 2.7 (SE) for K 562 cells. These values were around 3-fold higher than those of InsP(3) (4.8 +/ 0.5 micromol.kg(-1) and 6.9 +/- 0.1 (SE) for MDA-MB 231 and K562 cells respectively). Treatment of malignant cells with InsP(6) resulted in a 2-fold increase in the intracellular concentrations of total InsP(3) (9.5 +/- 1.3 (SE) and 10.8 +/- 1.0 (SE) micromol.kg(-1) for MDA-MB 231 and K562 cells respectively, p < 0.05), without changes in InsP(6) levels. These results indicate that exogenous InsP(6) directly affects its physiological levels in plasma and brain of normal rats without changes on the total InsP(3) levels. Although a similar fluctuation of InsP(6) concentration was not seen in human malignant cell lines following InsP(6) treatment, an increased intracellular levels of total InsP(3) was clearly observed. PMID- 12127909 TI - Expression of active human beta-glucuronidase in Sf9 cells infected with recombinant baculovirus. AB - Antibody directed enzyme prodrug therapy (ADEPT) using glucuronide prodrugs is an experimental approach to reduce systemic toxicity of anti-cancer agents. Bioactivation of such prodrugs is achieved by fusion proteins consisting of targeting moieties (e.g. ligands of tumor specific antigens) and human beta glucuronidase. In order to test a large panel of possible beta-glucuronidase fusion proteins for their applicability in ADEPT, an easy, rapid and high-yield expression system like the baculovirus/insect cell expression system would be needed. A prerequisite for using such fusion proteins is functional and biochemical characterization of human beta-glucuronidase expressed in baculovirus infected insect cells. Therefore, recombinant human beta-glucuronidase was expressed in Sf9 insect cells and characterized at the protein and functional level. As shown by Western blot analysis the recombinant enzyme consists of dimers with their monomers being linked via disulfide bonds. Posttranslational modifications of the monomers seem to be different as compared with mammalian cells or tissues. The enzyme is functionally active in cleaving the substrates 5 bromo-4-chloro-3-indolyl-beta-D-glucuronic acid, 4-methylumbelliferyl-beta-D glucuronide and the glucuronide prodrug HMR 1826, respectively, with similar enzyme kinetic parameters as those found in human tissues. Our data demonstrate that beta-glucuronidase expressed in Sf9 cells displays the same enzymatic features as the protein expressed in mammalian cells. Therefore, we suggest that beta-glucuronidase fusion proteins produced in this cell line will be valuable tools for testing a large panel of various targeting moieties in human tumor xenograft models or may be used for ADEPT in man. PMID- 12127910 TI - Alterations in 5-HT(1A) receptors and adenylyl cyclase response by trazodone in regions of rat brain. AB - The in vivo effect of trazodone on the density of [(3)H]5-HT binding sites and 5 HT(1A) receptors and adenylyl cyclase (AC) response was studied in regions of rat brain. The chronic administration of trazodone (10 mg/Kg body wt, 40 days) resulted in a significant downregulation of [(3)H]5-HT binding sites and 5-HT(1A) receptors in cortex and hippocampus. Trazodone significantly (p < 0.0001) decreased the density of [(3)H]5-HT binding sites in cortex (42.6 +/- 3.6 fmol/mg protein, 65%) and hippocampus (12.6 +/- 1.6 fmol/mg protein, 87%) when compared to control values of 121.9 +/- 5.4 and 99.3 +/- 7.5 fmol/mg protein in these regions, respectively. Similarly there was a significant (p < 0.0001) decrease in the density of 5-HT(1A) receptors in both cortex (7.2 +/- 0.5 fmol/mg protein, 70%) and hippocampus (6.3 +/- 1.2 fmol/mg protein, 79%) when compared to control values of 24.2 +/- 2.1 and 30.6 +/- 3.7 fmol/mg protein, in these regions respectively. However, the affinity of [(3)H]5-HT to 5-HT binding sites (1.83 +/- 0.26 nM, p < 0.0001) and [(3)H]8-OH-DPAT to 5-HT(1A) receptors (0.60 +/- 0.06 nM, p < 0.05) was significantly decreased only in cortex when compared to the control K(d) values of 0.88 +/- 0.04 nM and 0.47 +/- 0.02 nM in these regions, respectively. The basal AC activity did not alter in treated rats, where as, the inhibition of forskolin-stimulated AC activity by 5-HT (10 microM) was significantly (p < 0.0001) decreased both in cortex (43%) and hippocampus (40%) when compared to control levels. In conclusion, chronic treatment with trazodone results in downregulation of 5-HT(1A) receptors in cortex and hippocampus along with concomitant increased AC response, suggesting the involvement of 5-HT(1A) receptor-mediated AC response in the mechanism of action of trazodone. PMID- 12127911 TI - The retinal pigment epithelium of Cr-deficient rats. AB - The purpose of this study is to investigate the effect of Cr deficiency on the rat retina. Three-week-old Wistar Kyoto rats were divided into 2 groups. Cr deficient rats were fed AIN-93G diet without Cr and deionized distilled water. Control rats were fed AIN-93G diet and deionized distilled water. The Cr and sugar concentrations in the whole blood and cholesterol concentration in the serum were measured. We observed the retina with an electron microscope, and counted phagocytized lamellar structures in the retinal pigment epithelium (RPE) before and after the start of light exposure on negative electron microscopic films. The whole blood Cr level of Cr-deficient rats was less than 0.2 microg/l. The blood sugar level of Cr-deficient rats was significantly higher than that of normal rats (p < 0.05). There were significantly more phagocytized lamellar structures in the RPE of Cr-deficient rats 1, 2, 7, 11 and 12 h after the start of light exposure than in that of normal rats (p < 0.05). However, no morphological abnormalities were found in the photoreceptor cells of Cr-deficient rats. Phagocytosis in the photoreceptor outer segment discs in the RPE was accelerated, but the pattern of the retinal circadian rhythm with maximum phagocytosis 2 h after exposure to light was unchanged. The Cr-deficient state may cause the membrane to degenerate, and phagocytosis of the photoreceptor outer segment discs in the RPE may be accelerated. This study provided an evidence of the nutritional importance of Cr in rat retina. PMID- 12127912 TI - Effects of herbal components on cDNA-expressed cytochrome P450 enzyme catalytic activity. AB - We evaluated the effects of 25 purified components of commonly used herbal products on the catalytic activity of cDNA-expressed cytochrome P450 isoforms in in vitro experiments. Increasing concentrations of the compounds were incubated with a panel of recombinant human CYP isoforms (CYP1A2, CYP2C9, CYP2C19, CYP2D6 and CYP3A4) and their effects on the conversion of specific surrogate substrates measured fluorometrically in a 96-well plate format. For each test substance, the IC50 (the concentration required to inhibit metabolism of surrogate substrates by 50%) was estimated and compared with IC50's for the positive control inhibitory drugs furafylline, sulfaphenazole, tranylcypromine, quinidine, and ketoconazole. Constituents of Ginkgo biloba (ginkgolic acids I and II), kava (desmethoxyyangonin, dihydromethysticin, and methysticin), garlic (allicin), evening primrose oil (cis-linoleic acid), and St. John's wort (hyperforin and quercetin) significantly inhibited one or more of the cDNA human P450 isoforms at concentrations of less than 10 uM. Some of the test compounds (components of Ginkgo biloba, kava, and St. John's wort) were more potent inhibitors of the isoforms 1A2, 2C19, and 2C19 than the positive controls used in each assay (furafylline, sulfaphenazole, and tranylcypromine, respectively), which are known to produce clinically significant drug interactions. The enzyme most sensitive to the inhibitory of effects of these compounds was CYP2C19, while the isoform least effected was CYP2D6. These data suggest that herbal products containing evening primrose oil, Ginkgo biloba, kava, and St. John's Wort could potentially inhibit the metabolism of co-administered medications whose primary route of elimination is via cytochrome P450. PMID- 12127913 TI - Characterization of radioiodinated (-)-ortho-iodovesamicol binding in rat brain preparations. AB - We investigated the binding characteristics of optical isomers of three iodovesamicol analogs to vesicular acetylcholine transporters (VAChT) and to sigma receptors (sigma-1, sigma-2) in rat brains. In competitive inhibition studies, (-)-enantiomers [(-)-ortho-iodovesamicol ((-)-oIV), (-)-meta iodovesamicol ((-)-mIV), (-)-vesamicol] displayed a higher affinity for VAChT than (+)-enantiomer [(+)-oIV, (+)-mIV, (+)-vesamicol]. (-)-oIV and (-)-mIV showed the same high affinity for VAChT as (-)-vesamicol. For sigma receptors(sigma-1, sigma-2), (-)-oIV (Ki = 62.2 nM (to sigma-1) and 554 nM(to sigma-2)) showed a lower affinity than (-)-mIV (Ki = 4.5 nM (to sigma-1) and 42.9 nM (to sigma-2)). Furthermore, in a saturation binding study, (-)-[125I]-oIV exhibited a Kd of 17.4 +/- 5.1 nM with a maximum number of binding sites, Bmax, of 559 +/- 51 fmol/ mg of protein. These results showed that (-)-oIV binds to vesicular acetylcholine transporters (VAChT) more selectively than (-)-mIV, previously reported as a VAChT mapping agent, and may be suitable for VAChT imaging studies. PMID- 12127914 TI - Acute coronary syndromes observed from an international perspective. PMID- 12127915 TI - The quality of care in acute coronary syndromes. PMID- 12127916 TI - Thrombin inhibitors in acute coronary artery disease. PMID- 12127917 TI - Low molecular weight heparin: a bridge over troubled water. PMID- 12127918 TI - Late is perhaps not ... too late for primary PCI in acute myocardial infarction. PMID- 12127919 TI - The Cardiology Information System: the need for data standards for integration of systems for patient care, registries and guidelines for clinical practice. PMID- 12127920 TI - Management of acute coronary syndromes. Variations in practice and outcome; findings from the Global Registry of Acute Coronary Events (GRACE). AB - AIMS: Despite advances in the treatment of acute coronary syndromes based on randomized trial data and published guidelines, the extent to which such treatments are applied in practice remains uncertain. Data from clinical trials derive from selected geographical areas and in highly selected populations of patients, and hence may not reflect the overall population. The aim of the study was to investigate variations in hospital management and outcome using unselected data collected in the prospective Global Registry of Acute Coronary Events (GRACE). METHODS AND RESULTS: The 95 hospitals in GRACE were organized into 18 population-based clusters in 14 countries. Information was recorded about patient management and outcome during hospitalization and after discharge. Data on treatments administered were analysed by baseline condition, hospital type, by the presence or absence of a catheterization laboratory, and by geographical region. Of 11543 patients, 44% had an admission diagnosis of unstable angina, 36% presented with myocardial infarction, 9% were admitted to rule out a myocardial infarction, 7% had chest pain and 4% were hospitalized for 'other cardiac' and 'non-cardiac' diagnoses. Of the total GRACE population 38% had a final diagnosis of unstable angina, 30% ST-segment elevation myocardial infarction, 25% non-ST segment elevation myocardial infarction, and 7% of 'other cardiac' and 'non cardiac' final diagnoses. The event rates for hospital death or reinfarction were six and 2% for non-ST-segment elevation myocardial infarction, seven and 3% for ST-segment elevation myocardial infarction, and 3% hospital death for unstable angina. The use of aspirin was similar across all hospital types and geographical regions. In contrast, the use of percutaneous coronary intervention and glycoprotein IIb/IIIa inhibitors was higher (P<0.0001) in teaching hospitals and hospitals with catheterization laboratories and was also higher in the United States. At discharge a higher percentage (P<0.0001) of patients received angiotensin-converting enzyme inhibitors in hospitals without catheterization laboratories. The use of statins was lower in non-teaching hospitals and in centres without a catheterization laboratory. CONCLUSIONS: The GRACE study reveals substantial differences in the management of patients based on hospital type and geographical location. Further analyses will determine whether such variations translate into differences in longer term outcomes. GRACE provides a multinational reference for the implementation of therapies of proven efficacy. PMID- 12127921 TI - A prospective survey of the characteristics, treatments and outcomes of patients with acute coronary syndromes in Europe and the Mediterranean basin; the Euro Heart Survey of Acute Coronary Syndromes (Euro Heart Survey ACS). AB - AIMS: To better delineate the characteristics, treatments, and outcomes of patients with acute coronary syndromes (ACS) in representative countries across Europe and the Mediterranean basin, and to examine adherence to current guidelines. METHODS AND RESULTS: We performed a prospective survey (103 hospitals, 25 countries) of 10484 patients with a discharge diagnosis of acute coronary syndromes. The initial diagnosis was ST elevation ACS in 42.3%, non-ST elevation ACS in 51.2%, and undetermined electrocardiogram ACS in 6.5%. The discharge diagnosis was Q wave myocardial infarction in 32.8%, non-Q wave myocardial infarction in 25.3%, and unstable angina in 41.9%. The use of aspirin, beta-blockers, angiotensin converting enzyme inhibitors, and heparins for patients with ST elevation ACS were 93.0%, 77.8%, 62.1%, and 86.8%, respectively, with corresponding rates of 88.5%, 76.6%, 55.8%, and 83.9% for non-ST elevation ACS patients. Coronary angiography, percutaneous coronary interventions, and coronary bypass surgery were performed in 56.3%, 40.4%, and 3.4% of ST elevation ACS patients, respectively, with corresponding rates of 52.0%, 25.4%, and 5.4% for non-ST elevation ACS patients. Among patients with ST elevation ACS, 55.8% received reperfusion treatment; 35.1% fibrinolytic therapy and 20.7% primary percutaneous coronary interventions. The in-hospital mortality of patients with ST elevation ACS was 7.0%, for non-ST elevation ACS 2.4%, and for undetermined electrocardiogram ACS 11.8%. At 30 days, mortality was 8.4%, 3.5%, and 13.3%, respectively. CONCLUSIONS: This survey demonstrates the discordance between existing guidelines for ACS and current practice across a broad region in Europe and the Mediterranean basin and more extensively reflects the outcomes of ACS in real practice in this region. PMID- 12127922 TI - Effect of hirudin vs heparin on haemostatic activity in patients with acute coronary syndromes; the GUSTO-IIb haemostasis substudy. AB - AIMS: We compared the effects of hirudin and heparin on thrombin generation and activity among 350 patients with acute coronary syndromes enrolled in the GUSTO IIb trial. METHODS AND RESULTS: We obtained blood at baseline; at 4, 8, and 24h into infusion; at drug termination; and 6 and 24h after termination. We assayed for thrombin activity (fibrinopeptide A, activated protein C, thrombin antithrombin complex), thrombin generation (prothrombin fragment 1.2), and platelet activation (platelet factor 4). Median baseline fibrinopeptide A and platelet factor 4 levels were elevated. Thrombin formation tended to increase with hirudin and decrease with heparin; by 8h into infusion, thrombin formation was significantly less for heparin (P<0.01). Most patients showed reduced thrombin activity and platelet activation during infusion of either agent. Hirudin-assigned patients had significantly lower fibrinopeptide A levels during infusion. Six h post-termination, both groups had increased thrombin activity. Thrombin formation was increased in heparin patients (P<0.0001), significantly more than with hirudin (P=0.005). Higher values of haemostasis markers tended to be associated with poorer 30-day outcomes. CONCLUSION: Although hirudin did not prevent generation of new thrombin, it appeared to inhibit thrombin activity more than did heparin and produced slower increases in thrombin formation after discontinuation. The reelevation of thrombotic markers after stopping intravenous antithrombin therapy and the tendency toward increased thrombotic events with post-treatment increases in marker levels suggest an ongoing, clinically significant prothrombotic state. These results raise the possibility of improving on current antithrombotics by preventing thrombin generation and thrombin activity and by sustained suppression of the prothrombotic state. PMID- 12127923 TI - Benefits of extended treatment with dalteparin in patients with unstable coronary artery disease eligible for revascularization. AB - AIMS: The FRISC II trial demonstrated that, for patients with unstable coronary artery disease, an early invasive strategy following acute treatment with dalteparin and aspirin, was superior to a more conservative approach. We evaluated whether it is beneficial to extend treatment with dalteparin to patients eligible for revascularization but for whom these procedures are performed after the initial hospital stay. METHODS AND RESULTS: As a subanalysis of FRISC II, the efficacy and clinical safety of extended dalteparin treatment (5000 or 7500 IU.12h(-1) to day 90) compared with placebo was assessed in 1601 patients randomized to a non-invasive group who underwent revascularization only when necessary because of recurring symptoms, (re)infarction, or severe ischaemia. By day 90, 440 patients had undergone revascularization: 267 of these procedures occurred during the double-blind period. All patients initially received acute treatment (5-7 days from day 1) with dalteparin (120 IU/kg(-1) 12h(-1)). The incidence of death and/or myocardial infarction was monitored until revascularization or day 45 and until revascularization or day 90. There was a significant difference in the estimated probability of death and/or myocardial infarction until revascularization or day 90 in favour of dalteparin (log-rank test, P=0.0415) and there was a significant reduction in death and/or myocardial infarction in favour of extended dalteparin treatment at day 45, with a 57% relative risk reduction (P=0.0004). At day 90 the relative risk reduction was 29%. The safety profile of extended dalteparin treatment was similar to that of acute usage. CONCLUSION: Extended dalteparin treatment for up to 45 days is effective and safe as a bridging therapy for patients with unstable coronary artery disease awaiting revascularization. PMID- 12127924 TI - Impact of perioperative myocardial injury on early and long-term outcome after coronary artery bypass grafting. AB - AIMS: To establish the influence of perioperative myocardial injury on short- and long-term survival after coronary artery bypass grafting (CABG). METHODS AND RESULTS: The correlation of postoperative serum aspartate aminotransferase and creatine kinase MB to early cardiac-related death and to late survival was evaluated in 4911 patients who underwent CABG consecutively during a 6-year period. There were 93 early deaths (1.9%), 73 of them cardiac-related (1.5% of 4911). After a mean follow-up of 5 years, 409 additional deaths (8.5% of 4818) had occurred. Elevated enzyme levels on day 1 postoperatively highly increased the risk of early cardiac death (serum aspartate aminotransferase >or=2.35 microkat.l(-1): odds ratio 9.2; serum creatine kinase MB >or=61 microg.l(-1): odds ratio 6.0), and increased the risk of late death by approximately 50% (serum aspartate aminotransferase >or=2.35 microkat.l(-1): relative hazard 1.5; serum creatine kinase MB >or=61 microg.l(-1): relative hazard 1.4). This increased risk of death was independent of other risk factors and remained constant over time. CONCLUSIONS: Enzyme elevation after CABG implied an increased risk of both early and late death. The long-term effect persisted many years after surgery. PMID- 12127925 TI - Nutritional management: when and how should we become involved? PMID- 12127926 TI - How to manage central venous catheter-related sepsis. PMID- 12127927 TI - Epidural anaesthesia and analgesia - effects on surgical stress responses and implications for postoperative nutrition. AB - BACKGROUND: Surgical injury leads to an endocrine-metabolic and inflammatory response with protein catabolism, increased cardiovascular demands, impaired pulmonary function and paralytic ileus, the most important release mechanisms being afferent neural stimuli and inflammatory mediators. RESULTS: Epidural local anaesthetic blockade of afferent stimuli reduces endocrine metabolic responses, and improve postoperative catabolism. Furthermore, dynamic pain relief is achieved with improved pulmonary function and a pronounced reduction of postoperative ileus, thereby providing optimal conditions for improved mobilization and oral nutrition, and preservation of body composition and muscle function. Studies integrating continuous epidural local anaesthetics with enforced early nutrition and mobilization uniformly suggest an improved recovery, decreased hospital stay and convalescence. CONCLUSIONS: Epidural local anaesthetics should be included in a multi-modal rehabilitation programme after major surgical procedures in order to facilitate oral nutrition, improve recovery and reduce morbidity. PMID- 12127928 TI - Prevention and treatment of implanted central venous catheter (CVC) - related sepsis: a report after six years of home parenteral nutrition (HPN). AB - Catheter-related sepsis is a serious and common complication in patients receiving home parenteral nutrition (HPN). Prevention measures, prevalence of infections, types of agents and implanted central venous catheters (CVC), effectiveness of antibiotic therapy have been evaluated in 221 patients consecutively followed in our unit from January 1995 to December 2000. The clinical diagnosis of catheter-related infection was made using well-defined criteria. Patients were divided into two groups: A and B, receiving instructions with different modalities: standard (A) and detailed (B), respectively. Sixty CVC related sepsis occurred in 32 (14%) patients. A multivariate analysis showed that the duration of HPN (P<0.001; OR=0.9), type of catheter (P=0.009; OR=0.12) and type of disease (P=0.033; OR=4.92) significantly influence catheter infection. The type of implanted CVC (159 port-a-cath in 153 patients and 71 tunnelled in 68) seems to affect the infection rate, this being lower in tunnelled (P=0.03). Infection rate was lower in B vs A group (P<0.001) with all types of catheters, suggesting the preventive role of very careful training. In particular, the incidence of CVC-related sepsis was 6/1000 days of HPN (i.e. 6/1000 days of catheterization) in Group A and 3/1000 in Group B. Systemic and antibiotic lock therapy was performed with an 83% successful rate. Gram-positive bacteria were the most frequent CVC infection agents, which are usually eradicated by antibiotic therapy lasting 7 days. PMID- 12127929 TI - The effect of glutamine-supplemented total parenteral nutrition on nitrogen economy depends on severity of diseases in surgical patients. AB - BACKGROUND: Gln is an important substrate for enterocyte and rapid proliferation cells. Studies have shown that parenteral supplementation of Gln maintains the intracellular Gln pool, improves nitrogen balance and shortens hospital stay. However, some studies showed Gln-supplemented TPN had no effect on restoring the Gln pool in critically ill patients. OBJECTIVE: To evaluate the effect of glutamine (Gln) dipeptide supplementation of total parenteral nutrition (TPN) on postoperative nitrogen balance and immune response of patients undergoing surgery. METHODS: This study is a prospective, randomized double-blind clinical trial. APACHE II score and TISS were used to evaluate the patients after admission. Forty-eight patients with major abdominal surgery were allocated to two groups to receive isonitrogenous (0.228 g nitrogen/kg/day) and isoenergetic (30 kcal/kg/day) TPN for 6 days. Two groups (Conv and Ala-Gln) were further divided to high (APACHE>or=6) and low (APACHE <6) groups. Control group (Conv) received 1.5 g amino acids/kg/day, whereas the Ala-Gln group received 0.972 g amino acids/kg/day and 0.417 g of L-alanyl-L-glutamine (Ala-Gln)/kg/day. Blood samples were collected on day 1 and day 6 after surgery for plasma amino acid and CD4, CD8 cell and T lymphocyte analysis. Cumulative nitrogen balance were also measured on day 2, 3, 4, 5 postoperatively. RESULTS: Although there was a tendency to have better cumulative nitrogen balance on the postoperative days in the Ala-Gln group, no significant difference was observed between two groups. However, a better significant cumulative nitrogen balance was observed on the 2nd, 3rd and 5th postoperative day in the Ala-Gln group than in the Conv group in patients with APACHE II <6, whereas no significant difference was noted in patients with APACHE II >or= 6. No difference in urine 3-methylhistidine excretion were observed between the 2 groups. Patients in the Ala-Gln group had significant higher T lymphocyte and CD4 cells than did those in the Conv group. CONCLUSION: TPN supplemented with Gln dipeptide had beneficial effect on enhancing the immune response. However, the effect of Ala-Gln administration on improving nitrogen economy was only observed in patients with low APACHE II scores. These results may indicate that Gln required for reversing the catabolic condition may depend on the characteristics and severity of the diseases. PMID- 12127930 TI - Plasma total and free fatty acids composition in human non-alcoholic steatohepatitis. AB - BACKGROUND AND AIM: Non-alcoholic steatohepatitis (NASH), the association of steatosis with an inflammatory response, is a novel liver disease of unknown pathogenesis and prognosis. Triacylglycerols and their precursors, the fatty acids, are the likely candidates to accumulate in the hepatocyte. Disturbed fatty acid metabolism can be involved in the pathogenesis of NASH but there is no information concerning its plasma fatty acid profile. The aim of this study was to evaluate plasma total (esterified plus free) and free fatty acids concentrations to assess the association of NASH with plasma fatty acid accumulation. MATERIALS AND METHODS: Overnight fasting blood samples from 22 biopsy-proven NASH patients and of 6 matched age healthy controls were studied. RESULTS: NASH patients had significantly higher concentration of total and free fatty acids than controls (P<0.05), higher total saturated and monounsaturated levels in both studied lipid fractions (P<0.05), mainly due to the increase of hexadecanoic, hexadecenoic and octadecenoic acids. Absolute polyunsaturated fatty acids (PUFA) concentrations were similar in both groups. The C20:4/C18:2 and the C18:1/C18:0 ratios as well as the peroxidability index were not significantly different. CONCLUSION: In overweight/obese patients NASH is associated with deranged fatty acid metabolism which may be involved in its pathogenesis and/or progression. PMID- 12127931 TI - Fatty acid composition of plasma lipids in patients with pancreatic, lung and oesophageal cancer in comparison with healthy subjects. AB - BACKGROUND & AIMS: Despite the general notion of impaired nutritional status in cancer patients, studies on fatty acid status in cancer patients are limited. The aim of the present study was to investigate whether plasma n-3 fatty acids concentrations are reduced in patients with different tumour types. METHODS: We measured fatty acid composition in plasma phospholipids (PLs) and cholesteryl esters (CEs) in 71 newly diagnosed, untreated cancer patients of three tumour types: oesophageal or cardia cancer (n = 35), non-small cell lung cancer (n = 22) and pancreatic cancer (n = 15) and in 45 healthy subjects. RESULTS: In pancreatic cancer, plasma n-3 fatty acids showed a substantial reduction in both plasma PLs and CES. Although n-3 fatty acids in lung cancer also tended to be reduced, this difference failed to reach statistical significance. n-3 Fatty acid levels were especially reduced in pancreatic cancer patients without diabetes mellitus, and in lung cancer patients with weight loss. In oesophageal cancer, n-3 fatty acid concentrations were comparable to those in healthy subjects. CONCLUSION: We conclude that plasma n-3 fatty acid levels were reduced in pancreatic cancer, tended to be reduced in lung cancer, but were not altered in oesophageal cancer. Further studies are needed to assess the mechanisms underlying the observed changes in n-3 fatty acid concentrations. PMID- 12127933 TI - Does undernutrition contribute to diaphragm weakness in patients with severe COPD? AB - BACKGROUND AND AIMS: The assumption that undernourishment contributes to diaphragm weakness in chronic obstructive pulmonary disease (COPD) remains unproven. METHODS: We, therefore, studied diaphragm strength, measured as transdiaphragmatic pressure during a maximal voluntary sniff (Sn P(di)) and cervical magnetic stimulation (Tw P(di)), in two groups of 10 patients with severe COPD. The groups had equally severe COPD as judged by FEV(1) and thoracic gas volume (V(tg)). The malnourished group had a mean body mass index (BMI) of 17.3 kg/m(2) compared with 27 kg/m(2) for the normally nourished group (mean difference -9.7 kg/m(2); 95% confidence intervals -6.8 to -12.6 kg/m(2),P <0.0001). RESULTS: There was no significant difference between Tw P(di) (mean difference 2.1 cm H(2)O; 95% CI-3 to + 7.4 cm H(2)O, P=0.39) or Sn P(di) (mean difference -2.4 cm H(2)O; 95% CI-21 cm H(2)O to +16 cm H(2)O,P =0.8). CONCLUSION: We conclude that undernourishment of the severity studied does not contribute to diaphragm weakness in severe COPD. PMID- 12127932 TI - Nutrition beyond nutrition: plausibility of immunotrophic nutrition for space travel. AB - BACKGROUND AND AIMS: Microgravity has adverse effects on the immune system. We examined the effects of supplemental dietary nucleotides on immune function in ground-based in vivo anti-orthostatic tail-suspended (AOS) mice and in vitro (bioreactor-BIO) analogs of microgravity. METHODS: BALB/c mice were divided into the following three groups: group housed, single isolation, and AOS. Mice were fed either control chow or chow supplemented with RNA or uracil. Immune function was assessed by in vivo popliteal lymph node proliferation (PLN), in vitro PHA stimulated proliferation of splenocytes and cytokine production. BIO splenocytes were cultured in vitro with/without PHA, a nucleoside-nucleotide mixture (NS/NT) or uridine. The cell proliferation and scanning electron microscopic examination for cells were carried out. RESULTS: PLN response was significantly suppressed in AOS mice (P<0.05) and was restored by RNA and uracil diets. Splenocytes from AOS mice had decreased phytohemagglutinin (PHA)-stimulated proliferation, decreased IL-2 and IFN-gamma cytokine levels (P<0.05). These responses were restored by RNA and uracil diets. In BIO cultures, PHA response was suppressed significantly, and uridine and NS/NT restored the proliferative responses. Scanning electron microscopic analysis of cells cultured in BIO revealed cells with pinched, distorted and eroded membranes. Nucleotide supplementation especially uridine restored normal activated cell surface appearance and ruffling. CONCLUSION: In the microgravity analog environment of AOS and BIO, supplemental nucleotides and especially uracil/uridine have up-regulating and immunoprotective effects with potential as a countermeasure to the observed immune dysfunction in true microgravity. PMID- 12127934 TI - Folate supplementation after heart transplantation: effects on homocysteine plasma levels and allograft vascular disease. AB - BACKGROUND AND AIMS: After heart transplantation, the effects of folate supplementation on total homocysteine plasma levels (THcy) and heart allograft vascular disease (AVD) remain unclear. METHODS: Accordingly, we prospectively analyzed 48 heart transplant receipients referred for routine follow-up from July to September 1998 (age 54+/-11 years, 75% male, 35+/-27 months from transplant). Among these patients, 17 were treated with folate supplementation for 12 months (Group F), while 31 cross-matched for age, gender, serum creatinine and time from transplant (P>0.3 vs Group F for all) did not assume folate supplementation (Group NF). Routine coronary angiography for AVD detection was routinely obtained in every patient. RESULTS: THcy overall increased during the study period (from 16.6+/-6.5 to 19.4+/-7.6 micromol/l, P<0.001), and a strong trend toward higher THcy was observed in patients presenting AVD (22.4+/-8.7 vs 17.6+/-6.8 micromol/l, P=0.051). After 12 months THcy was lower in Group F as compared to Group NF (16.2+/-5.6 vs 21.1+/-8.1 micromol/l, respectively, P=0.033). CONCLUSIONS: Our results demonstrate that THcy increases over time in heart transplant recipients, and a strong trend toward higher THcy is observed in the presence of AVD. Since folate supplementation appears to positively influence THcy, a favorable effect of folate on AVD can be hypothesized. PMID- 12127935 TI - Nutritional status of elective gastrointestinal surgery patients pre- and post operatively. AB - BACKGROUND AND AIMS: Studies have indicated that undernutrition is common on admission to hospital but there is limited data on change in nutritional parameters during the hospital stay. We assessed the nutritional status of elective gastrointestinal surgery patients on admission and documented change in nutritional indices during hospitalisation. METHODS: Two hundred patients aged 18 80 years undergoing elective open gastrointestinal surgery were nutritionally assessed on admission and 150 were reassessed on commencement of oral diet post surgery. Data were collected on height, weight, upper arm anthropometry and hand grip dynamometry. RESULTS: On admission BMI <20, 20-24.9 and >25, respectively, were found in 9%, 34% and 57% of patients. Post-surgery, 34% of patients experienced a clinically significant weight loss. Males lost significantly more weight (3.7% vs 1.6%, P<0.001) and tended to lose muscle mass while females preferentially lost subcutaneous fat. CONCLUSIONS: The incidence of undernutrition on admission appears to be lower than previously reported. However, clinically significant weight loss was common and this study highlighted gender differences in the changes in nutritional parameters experienced by gastrointestinal surgery patients. This differential influence of gender warrants further investigation and may have implications for the nutritional management of such patients. PMID- 12127936 TI - Resting energy expenditure in children and adolescents: agreement between calorimetry and prediction equations. AB - BACKGROUND AND AIMS: To assess the degree of agreement between indirect calorimetry and five equations commonly used to predict resting energy expenditure (REE) in obese and non-obese children and adolescents. METHODS: In 116 children and adolescents (57 obese and 59 non-obese) aged between 7.8 and 16.6 years, REE was measured (MREE) by open-circuit indirect calorimetry under standardized conditions. REE was predicted (PREE) in all subjects with equations from the Food and Agriculture/World Health Organization/United Nations University (FAO/WHO/UNU), Maffeis et al., Harris and Benedict, and two from Schofield: one using weight (W) and one using height and weight (H-W). Agreement between indirect calorimetry and equations was assessed following the Bland-Altman method. RESULTS: In the entire cohort group, only data from FAO/WHO/UNU, Schofield-W and Schofield-HW equations showed non-statistic differences against calorimetry results. When agreement between equations and calorimetry was tested, Schofield-HW equation showed the lowest mean MREE-PREE difference: 3.7 kcal/d (limits of agreement -293 and 300 kcal/d; 95% confidence interval for the bias 24.0 to 31.5 kcal/d) and the best agreement. Group by group, equations which obtained the best agreement were: FAO/WHO/UNU in girls, Schofield-HW in boys, Schofield-HW in obese, and Schofield-W in non-obese. CONCLUSIONS: Until more accurate prediction equations are developed, we recommend Schofield-HW equations for REE studies with a mixed population of obese and non-obese children and adolescents; however, FAO/WHO/UNU equation may also be useful in girls and Schofield-W equation in non-obese children. PMID- 12127937 TI - Organization of higher-level chromatin structures (chromomere, chromonema and chromatin block) examined using visible light-induced chromatin photo stabilization. AB - The method of chromatin photo-stabilization by the action of visible light in the presence of ethidium bromide was used for investigation of higher-level chromatin structures in isolated nuclei. As a model we used rat hepatocyte nuclei isolated in buffers which stabilized or destabilized nuclear matrix. Several higher-level chromatin structures were visualized: 100nm globules-chromomeres, chains of chromomeres-chromonemata, aggregates of chromomeres-blocks of condensed chromatin. All these structures were completely destroyed by 2M NaCl extraction independent of the matrix state, and DNA was extruded from the residual nuclei (nuclear matrices) into a halo. These results show that nuclear matrix proteins do not play the main role in the maintenance of higher-level chromatin structures. Preliminary irradiation led to the reduction of the halo width in the dose-dependent manner. In regions of condensed chromatin of irradiated nucleoids there were discrete complexes consisting of DNA fibers radiating from an electron dense core and resembling the decondensed chromomeres or the rosette-like structures. As shown by the analysis of proteins bound to irradiated nuclei upon high-salt extraction, irradiation presumably stabilized the non-histone proteins. These results suggest that in interphase nuclei loop domains are folded into discrete higher-level chromatin complexes (chromomeres). These complexes are possibly maintained by putative non-histone proteins, which are extracted with high-salt buffers from non-irradiated nuclei. PMID- 12127938 TI - The expression patterns of integrin subunits on human breast tissues obtained during pregnancy. AB - Integrins have been shown to exert regulatory influences on mammary differentiation and morphogenesis in rodent experimental systems. We have, therefore, examined the expression patterns of integrin subunits on human breast tissues obtained at the 12th, 15th and 18th weeks of pregnancy. Myoepithelial cells were positive for all the integrin subunits examined. alpha2, alpha6 and beta4 showed increased and more defined labelling during pregnancy, indicating that myoepithelial cells and extracellular matrix strengthen their support for the expanding alveoli during pregnancy. Sub-populations of stromal cells were positive for alpha1, alpha3, alpha6, beta1 and beta4. On luminal epithelial cells, alpha1, alpha2, alpha3, alpha6 and beta1 were detectable. alpha2 showed a focal decrease, but the expression patterns of other integrins in luminal cells did not change during pregnancy. Therefore, the expression patterns of most integrins existing prior to pregnancy seem sufficient in this cell type to support the morphological and functional development during early pregnancy. PMID- 12127939 TI - Extremely low frequency (ELF) pulsed-gradient magnetic fields inhibit malignant tumour growth at different biological levels. AB - Extremely low frequency (ELF) pulsed-gradient magnetic field (with the maximum intensity of 0.6-2.0 T, gradient of 10-100 T.M(-1), pulse width of 20-200 ms and frequency of 0.16-1.34 Hz treatment of mice can inhibit murine malignant tumour growth, as seen from analyses at different hierarchical levels, from organism, organ, to tissue, and down to cell and macromolecules. Such magnetic fields induce apoptosis of cancer cells, and arrest neoangiogenesis, preventing a supply developing to the tumour. The growth of sarcomas might be amenable to such new method of treatment. PMID- 12127940 TI - Uptake of cadmium ions in white-rot fungus Trametes versicolor: effect of cd (II) ions on the activity of laccase. AB - The effects of cadmium Cd (II) ions on the physiology and biological activity of Trametes versicolor, a strain belonging to white-rotting Basidiomycetes, were examined. Cd (II) ions were added to 10-day-old cultures grown on a liquid medium, or at the time of inoculation. Our experiments showed that T. versicolor is a good cadmium biosorbent from aqueous solution, this strain removing almost all the Cd (ll) ions over the first 2h of incubation by what appears to be a rapid, energy-independent surface binding phenomenon, at the rate of approximately 2mg Cd per g mycelial dry weight. An additional slower and energy dependent transport mechanism was also present, taking in approximately 0.3mg Cd (II) perg dry weight. It is also shown that these Cd (II) ions significantly stimulate the activity of extracellular laccase when added to 10-day-old cultures. PMID- 12127941 TI - Expression and post-translational modification of human 4-hydroxy-phenylpyruvate dioxygenase. AB - 4-hydroxyphenylpyruvate dioxygenase (HPD) (EC 1.13.11.27) is a key enzyme involved in tyrosine catabolism. Congenital HPD deficiency is a rare, relatively benign condition known as hereditary type III tyrosinemia. The severe type I tyrosinemia, caused by a deficiency of fumarylacetoacetate hydrolase which functions downstream of HPD in the tyrosine degradation pathway, is often associated with decreased expression of HPD, and interestingly, inhibition of HPD activity seems to ameliorate the clinical symptoms of type I tyrosinemia. The HPD gene was previously mapped to the chromosomal region 12q24-->qter. In the present study high-resolution chromosome mapping localized the HPD gene to 12q24.31. DNase I footprinting, revealed that four regions of the HPD promoter were protected by rat liver nuclear proteins. Computer-assisted analyses suggested that these elements might bind Sp1/AP2, HNF4, HNF3/CREB, and C/EBP, respectively. In transient transfection experiments, the proximal 271bp of the promoter conferred basal transcriptional activation in human Chang cells. Sequences in intron 1 were able to enhance the activity of this basal promoter. Finally, vaccinia virus-based expression provided evidence that HPD is subject to phosphorylation, and furthermore, allowed mapping of the HPD protein in the human keratinocyte 2D database. PMID- 12127942 TI - Modification of glycosylation reduces microvilli on rat liver epithelial cells. AB - Effect of glycosylation modification on the shape of microvillus was investigated on rat liver epithelial cell line WB-F344. Since rat ER alpha-mannosidase has 89% identity to human 6A8 alpha-mannosidase, we used an antisense 6A8 cDNA fragment to inhibit expression in WB-F344 cells. Cells were transfected with antisense 6A8, the relevant sense fragment or the mock plasmid. Genomic PCR for neo(R) demonstrated integration of the transfected gene into host DNA. Enzymatic activity assay on p -nitro-phenyl-alpha-D-mannopyranoside showed suppression of ER alpha-mannosidase expression in the cells transfected with antisense 6A8. Concanavalin A binding to these cells was enhanced, indicating a modification in glycosylation. Number reduction and blunting of microvillus on these cells was observed. Transduction with the sense fragment or the mock had no effect. Cells with suppressed ER alpha-mannosidase expression grew slower in culture. Our results indicate an obvious effect of glycosylation modification on microvilli, which might be related with malfunctioning of cells. PMID- 12127943 TI - Spatial distribution of mDLG6 mRNA in embryonic and adult mouse brain. AB - We have isolated mouse DLG6 (mDLG6) cDNA clones by RT-PCR and then by using the RT-PCR products to screen a mouse brain cDNA library. The deduced amino acid sequence of mDLG6 shows 79.2% and 82.7% overall identity to human (hDLG6) and rat DLG6 (rDLG6), respectively. In situ hybridization revealed that mDLG6 mRNA is predominantly expressed in embryonic and adult brain. PMID- 12127944 TI - Unusual cytological patterns of microsporogenesis in Brachiaria decumbens: abnormalities in spindle and defective cytokinesis causing precocious cellularization. AB - Cytogenetic studies carried out in the tetraploid accession BRA001068 of Brachiaria decumbens, also known as cv. Basilisk, revealed an unusual pattern of microsporogenesis. The spindle in metaphase I and anaphase I became heavily stained with propionic carmine. In telophase I, the interzonal microtubules continued to be intensely stained, and during the phragmoplast formation the fibers were pushed to the cell wall, persisting until prophase II, even after cytokinesis. Due to its tetraploid condition, the accession presented many cells with precocious chromosome migration to the poles in metaphase I and laggards in anaphase I that gave rise to micronuclei in telophase I. While in other polyploid accessions of Brachiaria micronuclei remained in this condition until the second cytokinesis, the micronuclei in this accession organized their own spindle in the second division. In several microsporocytes, the micronuclei with their minispindle were divided further into microcytes by additional cytokinesis. Some curious planes of cytokinesis were found in some cells, with partitioning of cytoplasm into cells of irregular shape. The result consisted of a high frequency of abnormal products of meiosis. Quadrivalents were observed in diakinesis at low frequency, which suggests a segmental allotetraploid and the inability of both genomes to co-ordinate their activities, leading to multiple spindle and precocious cellularization. In spite of abnormal meiotic products reducing pollen fertility, seed production was normal. Enough normal pollen was available to fertilize the central-cell nucleus of the embryo sac and produce normal endosperm in this pseudogamous aposporous apomictic accession. PMID- 12127946 TI - Evidence that the novobiocin-sensitive ATP-binding site of the heat shock protein 90 (hsp90) is necessary for its autophosphorylation. AB - The 90kDa heat shock protein (Hsp90) is one of the most abundant protein and essential for all eukaryotic cells. Many proteins require the interaction with Hsp90 for proper function. Upon heat stress the expression level of Hsp90 is even enhanced. It is assumed, that under these conditions Hsp90 is required to protect other proteins from aggregation. One property of Hsp90 is its ability to undergo autophosphorylation. The N-terminal domain of Hsp90 has been shown to contain an unusual ATP-binding site. A well-known inhibitor of Hsp90 function is geldanamycin binding to the N-terminal ATP-binding site with high affinity. Recently it was shown that Hsp90 possesses a second ATP-binding site in the C terminal region, which can be competed with novobiocin. Autophosphorylation of Hsp90 was analysed by incubation with gamma(32)P-ATP. Addition of geldanamycin did not interfere with the capability for autophosphorylation, while novobiocin indeed did. These results suggest that the C-terminal ATP-binding site is required for autophosphorylation of Hsp90. PMID- 12127945 TI - Simple separation of chicken gonadal primordial germ cells with and without foreign genes. AB - Simplicity is the key element of an inexpensive technique described that is superior in performance to previous methods. It can make it the rapid method of choice to obtain reasonable yields of purified primordial germ cells (PGCs) for immediate production of germline chimeric chickens with integrated foreign genes. After Ficoll centrifugation, the purity of PGCs from gonads was 80.9+/-0.08% (mechanical) compared with 86.1+/-0.19% (enzymatic). GFP gene and lacZ-transduced chicken gonadal primordial germ cells (gPGCs) examined 72h after transduction had a transfection efficiency of approximately 61% and approximately 64%, respectively. After 10 days of G418 selection, approximately 90 and 92% of pure gPGCs did not contain other cells following this Ficoll gradient centrifugation method of preparation. PMID- 12127951 TI - The cure of thalassemia by bone marrow transplantation. PMID- 12127952 TI - Pathogenesis and treatment of anaemia of chronic disease. AB - Anaemia of chronic disease (ACD), the most frequent anaemia among hospitalized patients, develops under chronic inflammatory disorders such as chronic infections, cancer or autoimmune diseases. A number of different pathways contribute to ACD, such as diversion of iron traffic, a diminished erythropoiesis, a blunted response to erythropoietin, erythrophagocytosis and bone marrow invasion by tumour cells and pathogens. Nevertheless, ACD is a reflection of an activated immune system and possibly results from an innovative defence strategy of the body in order to withdraw the essential growth factor iron from invading pathogens and to increase the efficacy of cell-mediated immunity. Diagnosis of ACD can be assessed by examination of chances in serum iron parameters with low to normal serum iron, transferrin saturation and transferrin concentrations on the one hand and normal to increased ferritin, zinc protoporphyrin IX and cytokine levels on the other side. Therapy of ACD includes the cure of the underlying the disease. Apart from this transfusions for rapid correction of haemoglobin levels, and human recombinant erythropoietin for prolonged therapy are used. However, response rates to recombinant erythropoietin are sometimes low. Iron alone should be strictly avoided due to its growth promoting effect towards micro-organisms and tumour cells and because of it capacity to inhibit T-cell-mediated immune effector pathways. We urgently need prospective clinical trials to gain knowledge about the effects of anaemia correction and/or the use of erythropoietin towards the course of the underlying disease, to find out if a combination therapy with erythropoietin and iron may be beneficial in ACD and to define therapeutic end-points. PMID- 12127953 TI - Factor X deficiency. AB - Factor X is one of the vitamin K-dependent serine proteases. It plays a crucial role in the coagulation cascade, as the first enzyme in the common pathway of thrombus formation. The gene for factor X maps to the long arm of chromosome 13, approximately 2.8 kb downstream of the factor VII gene. The gene consists of eight exons, each of which encodes a specific functional domain within the protein. Both the gene structure and the amino acid sequence show homology to other vitamin K-dependent clotting factors, suggesting their origin in a common ancestral protein. Factor X deficiency is one of the rarest of the inherited coagulation disorders. Inheritance is in an autosomal recessive manner. The clinical phenotype is of a variable bleeding tendency. Homozygous factor X deficiency has an incidence of 1:1,000,000 in the general population. Heterozygotes are often clinically asymptomatic. Acquired factor X deficiency is rare, but when it occurs it is usually in association with amyloidosis. Treatment of factor X deficiency involves replacement of the protein with either fresh frozen plasma or prothrombin complex concentrates, although the latter should be used with caution as infusion may be associated with an increased risk of thrombosis. PMID- 12127954 TI - Hodgkin's disease. AB - Significant progress has been made over the past decade in the classification, diagnosis, staging, prognosis, and treatment of Hodgkin's disease (HD). A new classification system has recognized differences in the natural history of certain subtypes. The introduction of positron emission tomography has improved the accuracy of non-invasive staging. New prognostic indices have led to the development of risk-adapted treatment strategies. The serious long-term side effects of extended-field radiotherapy have prompted the increasing use of chemotherapy in conjunction with limited radiotherapy for early-stage patients. Combination chemotherapy remains the treatment of choice for advanced HD, and new dose-intense regimens appear to have improved activity. Patients who relapse now have a more favorable prognosis with the availability of active salvage regimens, autologous stem cell transplantation, and novel biologic agents. PMID- 12127955 TI - Hepatitis C and B-cell lymphoma: the hemato-hepatologist linkage. AB - Hepatitis C virus (HCV)-lymphotropism may be responsible for the development of mixed cryoglobulinemia (MC) and other lymphoproliferative disorders associated with HCV infection. An association between HCV infection and B-cell lymphoma has been largely demonstrated in several geographical areas with prevalence ranging between 7.4 and 37%. However, the intimate pathogenetic mechanism involved in HCV associated lymphomas remains considerably unknown. HCV may exerts its oncogenic potential via an indirect mechanism or utilizes other pathways directly. It is reasonable to assume that several different pathogenetic mechanisms operate in the wide spectrum of HCV-related lymphomas which includes the 'idiopathic', non cryoglobulinemic, intermediate to high-grade lymphoma, and the more common indolent, low-grade lymphoma, preceded by long standing symptomatic MC type II. In most cases, HCV has no significant impact on response to chemotherapy or survival of lymphoma patients. Treatment with chemotherapy is relatively safe, and interruption of treatment regimens is usually not required. Whether to treat low-grade HCV-related lymphomas with anti-viral therapy is still debatable, but encouraging data emerge from recent studies. PMID- 12127957 TI - Second malignancies after allogeneic hematopoietic stem cell transplantation: new insight and current problems. AB - With increased number of patients surviving on the long term, late effect after allogeneic hematopoietic stem cell transplantation have become of major clinical importance. Among these late effect, second malignancies have increasingly been recognized in the recent years. It has been usual to divide the problem of secondary malignancies following hematopoietic stem cell transplantation into three groups, i.e. leukemia, lymphoma and solid tumors. Recent clinical and biological data on these three types of malignancies, occurring after allogeneic stem cell transplantation, are summarized in this review. We will focus here only on second malignancies after allogeneic stem cell transplantation with particular emphasis on recent development on the pathogenesis, and early diagnosis, and treatment of these transplant-related complications. PMID- 12127956 TI - Orally active iron chelators. AB - In patients with transfusion-dependent anemias, iron accumulation is fatal in the absence of chelating therapy. Extended survival, free of most complications of iron overload is observed in patients treated with early, adequate parenteral deferoxamine. Despite its success in prevention and treatment of iron toxicity, the expense and inconvenience of this therapy have stimulated a continued quest for an effective chelating agent that is orally active. Unfortunately, studies emerging over the last five years have confirmed that the most widely administered orally active agent, deferiprone (L1; 1,2-dimethyl-3-hydropyrid-4 one) may be harmfully ineffective in many patients: 18-65% of patients in six studies which obtained hepatic irons after long term deferiprone treatment had body iron exceeding the threshold for cardiac disease and premature death. The impact of deferiprone on cardiac and liver disease must be evaluated further, while the association between deferiprone and accelerated hepatic fibrosis still awaits refutation in large prospective trials. In view of the striking therapeutic successes of deferoxamine over the past 20 years, administration of deferiprone outside the setting of prospective clinical trials may need to be reconsidered. Meanwhile, an orally active iron chelator of demonstrated safety and effectiveness remains an objective for development for transfused patients. PMID- 12127958 TI - Genome-wide multilocus analysis for immune-mediated complex diseases. AB - In postgenomic era, searching and identification of disease genes associated with complex diseases are still one of the great challenge for dissecting human complex diseases. To improve the disease gene localization for complex diseases, a group of closely immune-mediated disease loci were overlapped on each chromosome based on previously reported genome-wide scanning data. Interestingly, five overlapping chromosomal regions (1q21, 2q33, 5q31.1-q33.1, 6p21, and 11q13) were identified by co-localizing disease loci for the following diseases: diabetes, asthma, atopic dermatitis, osteoporosis, and inflammatory bowel disease. The development of specific disease was associated with different combinations of disease loci among five overlapped chromosomal regions. Therefore, the analysis of multiple genetic loci should be considered to determine the effects of multiple genes responsible for complex diseases resulting from the influence of multiple genes. PMID- 12127959 TI - Ubiquitination of Ro52 autoantigen. AB - Anti-Ro/SSA antibodies are antinuclear antibodies most commonly found in patients with Sjogren's syndrome, a chronic autoimmune disease characterized by dryness of the eyes and mouth. The autoantibodies recognize a RING-finger protein, Ro52/SSA (52 kDa), whose function is still unknown. In this study, the ubiquitination of Ro52 was investigated. We found that Ro52 was strongly conjugated by a single molecule of ubiquitin in cells. Although the biological relevance of this mono ubiquitination was not defined, the function of Ro52 might be modified by the mono-ubiquitination. We also found that Ro52 was conjugated with poly-ubiquitin chain in cells (poly-ubiquitination), suggesting that Ro52 may be downregulated by the ubiquitin-proteasome pathway in vivo. Interestingly, sera from patients with Sjogren's syndrome showed heterogeneity in their reactivity to poly ubiquitinated Ro52, probably because of their differing antigenic determinants. This heterogeneity of the reactivity might be associated with the varying clinical features found in patients with Sjogren's syndrome. PMID- 12127960 TI - Silkworm hemolymph as a potent inhibitor of apoptosis in Sf9 cells. AB - We have previously shown that silkworm hemolymph exhibits anti-apoptotic activity against baculovirus-induced Sf9 cell apoptosis. In this study, using various chemical inducers, such as actinomycin D, camptothecin, and staurosporine, we found that silkworm hemolymph inhibits insect cell apoptosis induced not only by baculovirus but also by chemical inducers. This indicates that silkworm hemolymph contains anti-apoptotic components that work directly in insect cell apoptosis without any booster expression of baculoviral genes. With the analysis of Sf caspase-1 activity, it was found that the inhibitory effect of silkworm hemolymph works in a further upstream step than the Sf-caspase-1 activation step. PMID- 12127961 TI - Downregulation of Wilms' tumor 1 protein inhibits breast cancer proliferation. AB - High levels of Wilms' Tumor 1 (WT1) mRNA have been correlated with poor prognosis in breast cancer patients. However, the function of WT1 protein in breast cancer is not known. We observed that the levels of WT1 protein correlated with the proliferation of breast cancer cells. When the proliferation of breast cancer cells was stimulated by 17beta-estradiol, WT1 protein expression increased. But when the proliferation of breast cancer cells was inhibited by tamoxifen or all trans retinoic acid (ATRA), WT1 protein expression decreased. We hypothesize that WT1 protein plays a role in regulating breast cancer cell proliferation. Using liposome-incorporated WT1 antisense oligodeoxynucleotides, we found that downregulation of WT1 protein expression led to breast cancer growth inhibition and reduced cyclin D1 protein levels. These results indicate that WT1 protein contributes to breast cancer progression by promoting breast cancer cell proliferation. PMID- 12127962 TI - Conotoxin MI inhibits the alpha-delta acetylcholine binding site of the Torpedo marmorata receptor. AB - The muscle-type nicotinic receptor has two pharmacologically distinguishable acetylcholine binding sites at the alpha-gamma and alpha-delta subunit interfaces; alpha-conotoxins can bind them selectively. As reported, alpha conotoxin MI has greater affinity for the site near the alpha-delta interface of the BC(3)H1 cell receptor but, in the case of the Torpedo californica receptor, displays greater affinity for that near the alpha-gamma interface. To further investigate ligand selectivity, we study the conotoxin MI-Torpedo marmorata receptor interaction. In this work, we show the binding of alpha-conotoxin MI to the T. marmorata receptor and the influence of the antagonist alpha-Bungarotoxin and the agonist carbamylcholine on such binding; in addition, and contrasting with the results for the Torpedo californica receptor, we identify the alpha delta subunit interface as the high affinity binding site. This is the first work describing different characteristics of the interaction between alpha-conotoxin MI and receptors from different species of the same genus. PMID- 12127963 TI - An anionic antimicrobial peptide from toad Bombina maxima. AB - Amphibian skin is a rich resource of antimicrobial peptides like maximins and maximins H from toad Bombina maxima. A novel cDNA clone encoding a precursor protein that comprises maximin 3 and a novel peptide, named maximin H5, was isolated from a skin cDNA library of B. maxima. The predicted primary structure of maximin H5 is ILGPVLGLVSDTLDDVLGIL-NH2. Containing three aspartate residues and no basic amino acid residues, maximin H5 is characterized by an anionic property. Different from cationic maximin H peptides, only Gram-positive strain Staphylococcus aureus was sensitive to maximin H5, while the other bacterial and fungal strains tested were resistant to it. The presence of metal ions, like Zn2+ and Mg2+, did not increase its antimicrobial potency. Maximin H5 represents the first example of potential anionic antimicrobial peptides from amphibians. The results provide the first evidence that, together with cationic antimicrobial peptides, anionic antimicrobial peptides may also exist naturally as part of the innate defense system. PMID- 12127965 TI - Effect of luminal and extravesicular Ca2+ on NAADP binding and release properties. AB - Nicotinic acid adenine dinucleotide phosphate (NAADP) has been shown to be a powerful Ca2+ release agent in numerous systems, including echinoderms, plants, and mammalian cells. NAADP has been shown to release Ca2+ via a separate mechanism to IP3 and ryanodine receptors, and specific binding sites have recently been characterised. However, functional studies have shown that there is a functional interplay between the NAADP-sensitive mechanism and the other two. In particular, it appears that activation of the NAADP receptor might act as a trigger to facilitate responses from IP3 and ryanodine receptors. To further characterise this interplay, we have investigated the effects of luminal and cytosolic Ca2+ on the NAADP receptor in sea urchin egg homogenates. We report that neither cytosolic nor luminal Ca2+ appears to influence NAADP binding. Conversely, emptying of stores significantly amplifies NAADP-induced fractional Ca2+-release, providing a mechanism of self-adjustment independent of store loading. PMID- 12127964 TI - Tyrosine phosphorylation of maspin in normal mammary epithelia and breast cancer cells. AB - Maspin is a 42kDa tumor suppressor protein that belongs to the serine protease inhibitor (serpin) family. It inhibits cell motility and invasion in vitro, and tumor growth and metastasis in nude mice; however, maspin's molecular mechanism of action has remained elusive. Maspin contains several tyrosine residues and we hypothesized that phosphorylation of maspin could play a role in its biological function. Our study reveals that maspin is phosphorylated on tyrosine moiety(ies) in normal mammary epithelial cells endogenously expressing maspin. In addition, transfection of the maspin gene, using either a stable or inducible system into maspin-deficient breast cancer cell lines, yields a protein product that is phosphorylated on tyrosine residue(s). Furthermore, recombinant maspin protein can be tyrosine-phosphorylated by the kinase domain from the epidermal growth factor receptor in vitro. These novel observations suggest that maspin, which deviates from the classical serpin, may be an important signal transduction molecule in its phosphorylated form. PMID- 12127966 TI - Role of cis prolines 112 and 126 in the functional activity of ribonucleolytic toxin restrictocin. AB - Restrictocin is a 149 amino acid ribonucleolytic toxin produced by the fungus Aspergillus, which specifically cleaves a single phosphodiester bond within 28S rRNA resulting in a potent inhibition of protein synthesis in eukaryotic cells. Restrictocin has 12 prolines out of which three at positions 48, 112, and 126 are cis. Prolines at position 112, 118, and 126 were individually mutated to alanine to investigate their role in the catalytic and membrane interaction activity of restrictocin. All mutants were expressed in Escherichia coli, and recombinant proteins purified to homogeneity. Mutation of P112 resulted in a remarkable 50- and 100-fold reduction, respectively, in the ribonucleolytic and cytotoxic activities of restrictocin, whereas the interaction of P112A with phospholipid membranes increased. Mutants P118A and P126A exhibited 3-5-fold decreased ribonucleolytic and cytotoxic activities, however, their membrane interaction activity was marginally reduced compared to restrictocin. The study demonstrates that P112 is absolutely essential to maintain the functionally active conformation of restrictocin. Also, prolines 112, 118, and 126 do not appear to be directly involved in the membrane interaction activity of restrictocin. PMID- 12127967 TI - A novel amylolytic enzyme from Thermotoga maritima, resembling cyclodextrinase and alpha-glucosidase, that liberates glucose from the reducing end of the substrates. AB - The gene previously designated as putative cyclodextrinase from Thermotoga maritima (TMG) was cloned and overexpressed in Escherichia coli. The recombinant TMG was partially purified and its enzymatic characteristics on various substrates were examined. The enzyme hydrolyzes various maltodextrins including maltotriose to maltoheptaose and cyclomaltodextrins (CDs) to mainly glucose and maltose. Although TMG could not degrade pullulan, it rapidly hydrolyzes acarbose, a strong amylase and glucosidase inhibitor, to acarviosine and glucose. Also, TMG initially hydrolyzes p-nitrophenyl-alpha-pentaoside to give maltopentaose and p nitrophenol, implying that the enzyme specifically cleaves a glucose unit from the reducing end of maltooligosaccharides unlike to other glucosidases. Since its enzymatic activity is negligible if alpha-methylglucoside is present in the reducing end, the type of the residue at the reducing end of the substrate is important for the TMG activity. These results support the fact that TMG is a novel exo-acting glucosidase possessing the characteristics of both CD-/pullulan hydrolyzing enzyme and alpha-glucosidase. PMID- 12127968 TI - The complete amino acid sequence of the pediocin-like antimicrobial peptide leucocin C. AB - The pediocin-like antimicrobial peptide leucocin C produced by a strain of Leuconostoc mesenteroides has been purified using a recently developed rapid two step procedure. The complete and corrected amino acid sequence of the peptide has been determined by Edman degradation of the intact peptide and a C-terminal fragment generated by cleavage with Asp-N endoprotease. Leucocin C contained 43 residues with the following sequence: KNYGNGVHCTKKGCSVDWGYAWTNIANNSVMNGLTGGNAGWHN. The molecular weight of leucocin C as determined by mass spectrometry was 4595, which is consistent with the theoretical molecular weight of 4596 calculated from the sequence. Moreover, the molecular weights of the two fragments generated by cleavage with Asp-N were also consistent with the determined sequence. PMID- 12127969 TI - Cross-linked crystals of chloroperoxidase. AB - Chloroperoxidase from Caldariomyces fumago was crystallized. The crystals were modified with several cross-linkers, but only glurataldehyde was able to produce catalytically active and insoluble crystals. Unlike other immobilized chloroperoxidase preparations, these catalytic crystals are more thermostable than the unmodified soluble enzyme. The enhanced stability is probably due to the structure conservation in the crystalline matrix. In addition, non-cross-linked chloroperoxidase crystals retained more activity than the soluble enzyme after incubation in an organic solvent with low water content. Although the cross linked crystals were catalytically active, they showed lower specific activity than the soluble enzyme. This low activity may be due to non-specific reactions between the cross-linker and essential residues for catalysis. Alternative cross linking strategies are discussed. PMID- 12127970 TI - Reversal of P-glycoprotein-mediated multidrug resistance by 5,6,7,3',4' pentamethoxyflavone (Sinensetin). AB - Multidrug resistance (MDR) cells can be sensitized to anticancer drugs when treated concomitantly with chemosensitizers. In this study, chemosensitizing effects of 5,6,7,3',4'-pentamethoxyflavone (sinensetin) and its analogs were investigated with respect to in vitro efficacy and structure-activity relationship. Sinensetin reversed the resistance of P-glycoprotein (Pgp) overexpressing AML-2/D100 to vincristine in a concentration-dependent manner. Chemosensitizing effect of sinensetin was 10- and 18-fold higher than those of 5,7,3',4'-tetramethoxyflavone and 3,7-dihydroxy-3',4'-dimethoxyflavone, respectively. Sinensetin cytotoxicity in AML-2/D100 was not changed by the complete inhibition of Pgp, suggesting that it is not a substrate for Pgp. Flow cytometry showed that sinensetin increased drug accumulation in the AML-2/D100 in a concentration-dependent manner. Unlike verapamil and cyclosporin A, the maximum non-cytotoxic concentrations of sinensetin were found to decrease the Pgp levels. Azidopine-binding assay showed that cyclosporin A or verapamil inhibited azidopine binding on Pgp partially but sinensetin did not. Taken together, these results suggest that sinensetin has a chemosensitizing effect in reversing Pgp mediated MDR by increasing the intracellular accumulation of drugs without competition in a binding site of azidopine. Thus, sinensetin is anticipated as a novel and highly potent second-generation flavonoid chemosensitizer, since sinensetin has significant advantages of having a high therapeutic index, of being a non-transportable inhibitor, and of effecting no induction of Pgp. PMID- 12127971 TI - SVK14 cells express an MCH binding site different from the MCH1 or MCH2 receptor. AB - Melanin-concentrating hormone (MCH) is a cyclic peptide, mainly involved in the regulation of skin pigmentation in teleosts and feeding behavior in mammals. The human keratinocyte SVK14 cell line has been previously shown to express binding sites for the MCH analog [125I]-[Phe13,3-iodo-Tyr19]MCH. We report here that: (1) this binding site similarly recognized [125I]-[3-iodo-Tyr13]MCH; (2) its pharmacological profile clearly differed from those observed at the two human MCH receptor subtypes, MCH1-R and MCH2-R; (3) MCH did not induce any effect on second messenger systems (including cAMP, calcium, and MAP kinase signaling pathways), and (4) no mRNAs corresponding to the MCH receptors were found. In conclusion, the binding site characterized in the SVK14 cell line is distinct from the MCH1 and MCH2 receptors and deserves therefore further investigation. PMID- 12127972 TI - Cloning and characterization of two novel zebrafish P2X receptor subunits. AB - In this report we describe the cloning and characterization of two P2X receptor subunits cloned from the zebrafish (Danio rerio). Primary sequence analysis suggests that one cDNA encodes an ortholog of the mammalian P2X(4) subunit and the second cDNA encodes the ortholog of the mammalian P2X(5) subunit. The zP2X(4) subunit forms a homo-oligomeric receptor that displays a low affinity for ATP (EC(50)=274+/-48 microM) and very low affinity (EC(50)>500 microM) for other purinergic ligands such as alphabetameATP, suramin, and PPADS. As seen with the mammalian orthologs, the zP2X(5) subunit forms a homo-oligomeric receptor that yields very small whole-cell currents (<20pA), making determination of an EC(50) problematic. Both subunit genes were physically mapped onto the zebrafish genome using radiation hybrid analysis of the T51 panel, with the zp2x4 localized to LG21 and zp2x5 to LG5. PMID- 12127973 TI - Distinct roles of alphaA- and alphaB-crystallins under thermal and UV stresses. AB - alpha-Crystallin, a major protein of all vertebrate lenses, consists of two subunits, alphaA and alphaB, which form polymeric aggregates with an average molecular mass of about 800kDa. In this study, we have employed various biophysical methods to study aggregate sizes and conformational properties of purified alphaA, alphaB subunits, and cloned recombinant alphaB subunit. From far and near-UV CD spectra, native alpha-, alphaA-, alphaB-, and recombinant alphaB crystallins from porcine lenses all show similar beta-sheet conformation to that from bovine and human lenses as reported previously. By means of gel-filtration chromatography and dynamic light scattering, we have found that the molecular sizes of all four crystallin aggregates are polydispersedly distributed in the following order of aggregate sizes, i.e., native alpha>alphaA>alphaB approximately recombinant alphaB. To investigate the structural and functional relationships, we have also compared the chaperone activities of all four alpha crystallin aggregates at different temperatures. From the results of chaperone activity assays, ANS (8-anilinonaphthalene-1-sulfonic acid) binding and thermal stability studies, there appeared to be at least two factors playing major roles in the chaperone-like activity of these lens proteins: one is the hydrophobicity of the exposed protein surface and the other is the structural stability associated with each protein. We showed that alphaA-crystallin is a better chaperone to protect gamma-crystallin against UV irradiation than alphaB crystallin, in contrast to the observation that alphaB is generally a better chaperoning protein than alphaA for enzyme protective assays at physiological temperatures. PMID- 12127974 TI - Identification and characterization of two novel zinc finger genes, ZNF359 and ZFP28, in human development. AB - Transcription factors play an essential role in controlling gene expression during cardiac and vascular pathogeneses. Identification of regulatory genes in the cardiovascular system is a necessary step toward an understanding of the pathogenesis of congenital heart disease and acquired cardiovascular diseases. The Cys2/His2 type zinc finger genes are the single largest class of transcription factors in the human genome and many numbers of these krupple-like zinc finger genes have been found to be involved in cardiac development or cardiovascular diseases. In this study, we have identified two novel human krupple-like zinc finger genes named ZNF359 and ZFP28 from the human heart cDNA library. The complete human ZNF359 cDNA sequence is 3270bp and contains a 1932-bp open reading frame (ORF) that encodes a 643 amino acid protein with an N-terminal KRAB domain and 16 C-terminus zinc finger C2H2 motifs. The ZFP28 cDNA sequence is 4104bp and contains a 2076-bp ORF that encodes an 868 amino acid protein with an N-terminal signal peptide, two KRAB domains, and 14 C-terminal C2H2 zinc finger motifs. Northern blot analyses showed a strong expression of ZNF359 and ZFP28 in various tissues of adult human. A further analysis using human embryonic tissues (18-23 weeks) showed a development-specific expression pattern in heart, skeletal muscle, liver, lung, kidney, and brain, suggesting a role for these genes in embryonic development. PMID- 12127975 TI - Model mice for Presbyterian hemoglobinopathy (Asn(beta108)-->Lys) confer hemolytic anemia with altered oxygen affinity and instability of Hb. AB - Hb Presbyterian is a variant hemoglobin that carries Lys at Asn-108 of beta globin. This variant Lys(beta108) residue enhances the stability of Hb in the deoxy-state, conferring the low affinity for oxygen-binding in vitro. In the present study, we generated mutant mice carrying the Presbyterian mutation (Asn(beta108)-->Lys) at the beta-globin locus by a targeted knock-in strategy. Heterozygous mice showed the expression of Hb Presbyterian in 27.7% of total peripheral blood without any hematological abnormalities, which well mimicked human cases. On the other hand, homozygous mice exclusively expressed Hb Presbyterian in 100% of peripheral blood associated with hemolytic anemia, Heinz body formation, and splenomegaly. Hb Presbyterian showed instability in an in vitro precipitation assay. Erythrocytes from homozygous mice showed a shortened life span when transfused into wild-type mice, confirming that the knocked-in mutation of Lys(beta108) caused hemolysis in homozygous mice. This is the first report on the hemolytic anemia of unstable hemoglobin in an animal model. These results confirm the notion that the higher ratio of an unstable variant beta globin chain in erythrocytes triggers the pathological precipitation and induces hemolysis in abnormal hemoglobinopathies. PMID- 12127976 TI - Lys 43 and Asp 46 in alpha-helix 3 of uteroglobin are essential for its phospholipase A2 inhibitory activity. AB - Uteroglobin (UG) is an anti-inflammatory, secreted protein with soluble phospholipase A2 (sPLA2)-inhibitory activity. However, the mechanism by which UG inhibits sPLA2 activity is unknown. UG is a homodimer in which each of the 70 amino acid subunits forms four alpha-helices. We previously reported that sPLA2 inhibitory activity of UG may reside in a segment of alpha-helix 3 that is exposed to the solvent. In addition, it has been suggested that UG may inhibit sPLA2 activity by binding and sequestering Ca++, essential for sPLA2 activation. By site-specific mutation, we demonstrate here that Lys 43 Glu, Asp 46 Lys or a combination of the two mutations in the full-length, recombinant human UG (rhUG) abrogates its sPLA2-inhibitory activity. We demonstrate further that recombinant UG does not bind Ca++ although when it is expressed with histidine-tag (H-tag) it is capable of binding Ca++. Taken together our results show that: (i) Lys 43 and Asp 46 in rhUG are critical residues for the sPLA2-inhibitory activity of UG and (ii) Ca++-sequestration by rhUG is not likely to be one of the mechanisms responsible for its sPLA2-inhibitory activity. PMID- 12127977 TI - Enzymatic properties of a new peptide deformylase from pathogenic bacterium Leptospira interrogans. AB - Peptide deformylase (LiPDF), a target protein for antibacterial agents from pathogenic bacteria Leptospira interrogans was identified and purified. Enzymatic studies including kinetics and inhibition revealed new inspiring highlights. The purified active enzyme was a dimer and showed a hyperbolic progress plot when the substrate was low but an excess substrate inhibition effect in higher substrate concentration. Variants on the metal-binding ligand-Cys102 were constructed to verify the indispensable attribute. Also the variant, LiPDF with the insertion residues (R(70)Y(71)P(72)G(73)T(74) P(75)D(76)V(77)) between the conserved motif 1 and motif 2 excised, was constructed and displayed no marked changes on enzymatic features. The results of atom absorbance proved that it contains a tightly bound Zn2+ rather than Fe2+ in E. coliPDF that is an essential cofactor for its high catalytic activity. PMID- 12127978 TI - Expression studies of two paralogous ppa genes encoding distinct Family I pyrophosphatases in marine unicellular cyanobacteria reveal inactivation of the typical cyanobacterial gene. AB - Genome sequence analyses revealed the occurrence of two paralogous ppa genes potentially encoding distinct Family I inorganic pyrophosphatases (sPPases, EC3.6.1.1) in the marine unicellular cyanobacteria Prochlorococcus marinus strains MED4 and MIT9313 and Synechococcus sp. WH8102. Protein sequence alignment and phylogenetic analysis indicated that the ppa gene proper of cyanobacteria (ppa1) encodes a presumably inactive mutant enzyme whereas the second gene (ppa2) might encode an active sPPase closely related to those of some proteobacteria. Heterologous expression of the two cloned P. marinus MED4 ppa genes in Escherichia coli confirmed this proposal, only the inactive ppa1 product being immunodetected by anti-cyanobacterial sPPase antibodies. A possible scenario of ppa gene inactivation and replacement in the context of the postulated rapid diversification of marine unicellular cyanobacteria, the most abundant photosynthetic prokaryotes in the oceans, is discussed. PMID- 12127979 TI - Synergistic activity of fibronectin and fibroblast growth factor receptors on neuronal adhesion and neurite extension through extracellular signal-regulated kinase pathway. AB - Fibroblast growth factor (FGF) is an important modulator of cell growth and differentiation of various cells including neuron. Cells need to adhere specifically to cellular and extracellular components of their environment to carry out diverse physiological functions. Here, we examined whether fibronectin (FN) and FGF can cooperate for neuronal adhesion and neurite outgrowth. Using recombinant FN peptide (FNIII9-10), we found that FNIII9-10-mediated adhesion promotes the effect of FGF1 on neurite outgrowth of PC12 cells, while FGF1 enhances the FNIII9-10-mediated neuronal adhesion of PC12 cells. This collaboration of FNIII9-10 and FGF1 was the result of the sustained activation of extracellular signal-regulated kinase (ERK)-type MAP kinase. Finally, the synergistic activity of FGF1 and FN was inhibited by PD98059, an MEK inhibitor. Taken together, these findings indicate that FN-mediated signaling can collaborate with FGFRs signaling for neurite outgrowth through selective activation of ERK-type MAP kinase in PC12 cells, and suggest that FN and FGF act in concert to regulate cell differentiation in the nervous system. PMID- 12127980 TI - Enhanced detection and characterization of protocatechuate 3,4-dioxygenase in Acinetobacter lwoffii K24 by proteomics using a column separation. AB - Acinetobacter lwoffii K24 known as an aniline degrading bacterium has also been found to utilize p-hydroxybenzoate as a sole carbon source. In this study, 2-DE using Q-Sepharose column separation was attempted for fast screening of protocatechuate 3,4-dioxygenase for catabolism of p-hydroxybenzoate in A. lwoffii K24. Two protocatechuate 3,4-dioxygenase subunits, pcaG and pcaH were detected and identified with N-terminal and internal sequencing, suggesting proteomics using a column separation may be helpful for the identification of specific protein spots and maximizing the detectable protein spots on the 2-DE gel. The PCR process using degenerate primers for protocatechuate 3,4-dioxygenase and sequence analyses of the PCR products revealed the existence of pcaH and pcaG in A. lwoffii K24. These two subunits were found to be closely located and share extensive homology with pcaH and pcaG of Pseudomonas marginata or Pseudomonas cepacia, providing the evidence that A. lwoffi K24 has the protocatechuate branches as well as catechol branches of beta-ketoadipate pathway. PMID- 12127981 TI - Identification of putative mammalian D-lactate dehydrogenase enzymes. AB - Mammalian L-isomer dehydrogenases represent an expansive and well characterized class of metabolic enzymes. Surprisingly, little is known regarding their evolutionarily distinct counterparts, D-isomer dehydrogenases, since few mammalian D-isomer 2-hydroxy acid enzymes have been isolated. Here we present the identification and initial characterization of putative human and murine D lactate dehydrogenases (DLD) that can interact with the muscle-specific cysteine rich protein CRP3/MLP. Sequence analysis reveals that the human and mouse transcripts encode novel proteins that display strong similarities to the yeast D lactate dehydrogenase proteins DLD1, AIP2, and YEL071W. Expression analysis of the mammalian proteins indicates widespread distribution with transcripts present in striated muscle tissues and a variety of other tissue types. Immunofluorescence subcellular localization of the mouse DLD protein indicates that it resides within mitochondria, a feature shared by many dehydrogenases. The identification of the human and mouse DLD clones provides new insight regarding the activity of D-isomer-specific enzymes in mammalian cells. PMID- 12127982 TI - Role of dietary phosphorus in the progression of renal failure. AB - Dietary phosphorus is thought to be a factor that impairs the residual renal function in patients with chronic renal failure. To determine the effect of dietary phosphorus on the prognosis of chronic renal failure, low-phosphorus milk was prepared from normal cow's milk using boehmite, a synthetic phosphate-ion absorbent. Regular diet, normal cow's milk, and low-phosphorus milk were then given to 5/6-nephrectomized rats and the serum levels of inorganic phosphorus, calcium, creatinine, and blood urine nitrogen in the rats in each group were compared. The serum levels of inorganic phosphorus and calcium were not different among the groups, despite a significant difference in phosphorus intakes. On the other hand, serum levels of creatinine (Cr) and blood urine nitrogen (BUN) in the rats fed low-phosphorus milk were significantly lower (Cr, 0.54+/-0.054mg/dl; BUN, 29.2+/-3.90mg/dl) than those in the rats fed a regular diet (Cr, 0.64+/ 0.057mg/dl; BUN, 37.4+/-3.55mg/dl) or normal milk (Cr, 0.61+/-0.040mg/dl; BUN, 34.5+/-3.59mg/dl). No beneficial effect of protein restriction was observed when residual renal functions in rats fed a regular diet and those fed normal milk were compared. The results suggest that dietary phosphorus plays a major role in the progression of renal failure. PMID- 12127983 TI - Involvement of in vivo induced icmF gene of Vibrio cholerae in motility, adherence to epithelial cells, and conjugation frequency. AB - Previously, using global transcription profile approach icmF gene of Vibrio cholerae was identified as an in vivo induced gene. In the present study, the icmF gene of V. cholerae O395 was cloned, sequenced, and used to construct an icmF insertion mutant. This IcmF is homologous to Legionella pneumophila IcmF, belonging to the icm cassette responsible for macrophage killing and intracellular survival of the organism. The icmF insertion mutant exhibited reduced motility and increased adherence to human intestinal epithelial cells. The presence of ATP-GTP-binding site suggests further a possible role of IcmF as a signaling molecule. Triparental-mating assay, with the mutant as a recipient, showed higher conjugation frequency than wild type. We propose that the increased adherence to epithelial cell line and increased conjugation frequency of the mutant result from some sort of cell surface reorganization. PMID- 12127984 TI - Effect of hypoxia on excitatory transmission in the rat substantia gelatinosa neurons. AB - We have investigated the effect of hypoxia on the excitatory synaptic transmission in the substantia gelatinosa neurons using perforated-patch-clamp configuration. Brief periods of hypoxia induced a depression in the evoked excitatory postsynaptic current (eEPSC) amplitude. The hypoxia-induced depression of eEPSC was not observed in the presence of theophylline, a nonselective adenosine receptor antagonist, and DPCPX, a selective adenosine receptor A1 antagonist. Application of adenosine (100 microM) also depressed eEPSC in a similar way as with hypoxia. This adenosine-induced depression of eEPSC was inhibited by DPCPX. Hypoxia and exogenous adenosine decreased the frequency of the spontaneous excitatory postsynaptic current (sEPSC) but not the amplitude of sEPSC and increased the paired-pulse ratio. From these results, it is suggested that acute hypoxia depresses the excitatory synaptic transmission by activating the presynaptic adenosine A1 receptor. PMID- 12127986 TI - Human mitochondrial transcription factor A binds preferentially to oxidatively damaged DNA. AB - Mitochondrial transcription factor A (mtTFA) is necessary for both transcription and maintenance of mtDNA, and is also one of the high mobility group (HMG) proteins that preferentially binds to cisplatin-damaged DNA. In this study we confirmed the preferential binding of mtTFA to cisplatin-damaged DNA, and also discovered that mtTFA binds to oxidatively damaged DNA. The affinity for oxidatively damaged DNA of mtTFA is higher for A/8-oxo-dG and C/8-oxo-dG than for G/8-oxo-dG and T/8-oxo-dG. Our findings suggest that mtTFA plays an important role in the recognition of oxidative DNA damage. PMID- 12127985 TI - TNF-alpha-mediated apoptosis in chondrocytes sensitized by MG132 or actinomycin D. AB - The mechanism of TNF-alpha-mediated chondrocyte apoptosis in human articular cartilage was investigated. First passage OA chondrocytes were treated with actinomycin D or MG132 in combination with TNF-alpha to facilitate cell death. The patterns of apoptosis-related proteins, NF-kappaB activation, and IkappaB degradation were analyzed. Cell death was increased by 0.2 microg/ml of actinomycin D or 20 microM MG132 in combination with TNF-alpha. Apoptosis potentiated by MG132 was more effectively inhibited by caspase inhibitors than that by actinomycin D. MG132 or actinomycin D both led to a significant increase in p53, but the expressions of the p53 response proteins increased only in MG132 treated chondrocytes. TNF-alpha induced chondrocyte IkappaB phosphorylation was unaffected by either MG132 or actinomycin D. MG132, but not actinomycin D, inhibited the chondrocyte IkappaB degradation induced by TNF-alpha and NF-kappaB activation. Our results suggest that MG132 and actinomycin D exert different influences upon TNF-alpha-mediated chondrocyte apoptotic signaling. PMID- 12127988 TI - Highly specific inactivation of triosephosphate isomerase from Trypanosoma cruzi. AB - We searched for molecules that selectively inactivate homodimeric triosephosphate isomerase from Trypanosoma cruzi (TcTIM), the parasite that causes Chagas' disease. We found that some benzothiazoles inactivate the enzyme. The most potent were 3-(2-benzothiazolylthio)-propanesulfonic acid, 2-(p-aminophenyl)-6 methylbenzothiazole-7-sulfonic acid, and 2-(2-4(4-aminophenyl)benzothiazole-6 methylbenzothiazole-7-sulfonic acid. Half-maximal inactivation by these compounds was attained with 33, 56, and 8 microM, respectively; in human TIM, half-maximal inactivation required 422 microM, 3.3 mM, and 1.6 mM. In TcTIM, the effect of the benzothiazoles decreased as the concentration of the enzyme was increased. TcTIM has a cysteine (Cys 15) at the dimer interface, whereas human TIM has methionine in that position. In M15C human TIM, the benzothiazole concentrations that caused half-maximal inactivation were much lower than in the wild type. The overall findings suggest that the benzothiazoles perturb the interactions between the two subunits of TcTIM through a process in which the interface cysteine is central in their deleterious action. PMID- 12127987 TI - Heterologous expression and purification of SpaB involved in subtilin biosynthesis. AB - Lantibiotic peptides contain thioether bridges termed lanthionines that are putatively generated by dehydration of Ser and Thr residues followed by Michael addition of cysteine residues within the peptide. The LanB and LanC proteins have been proposed to catalyze the dehydration and formation of the thioether rings, respectively. We report here the first heterologous overexpression in Escherichia coli of SpaB, the putative dehydratase for subtilin. Sequence analysis of spaB revealed several nucleotide differences with current gene database entries. The solubility of SpaB was increased dramatically when co-expressed with GroEL/ES, and soluble His(6)-tagged SpaB was purified. The protein is at least a dimer, and interaction between SpaB and SpaC was observed. SpaS the putative substrate for SpaB was overexpressed in E. coli as an intein fusion protein, and after cleavage, the peptide was obtained in good yield. PMID- 12127990 TI - Hrp3, a chromodomain helicase/ATPase DNA binding protein, is required for heterochromatin silencing in fission yeast. AB - Hrp3, a paralog of Hrp1, is a novel member of the CHD1 (chromo-helicase/ATPase DNA binding 1) protein family of Schizosaccharomyces pombe. Although it has been considered that CHD1 proteins are required for chromatin modifications in transcriptional regulations, little is known about their roles in vivo. In this study, we examined the effects of Hrp3 on heterochromatin silencing using several S. pombe reporter strains. The phenotypic analysis revealed that hrp3(+) is not an essential gene for cell viability. However, Hrp3 is required for transcriptional repression at silence loci of mat3. A chromatin immunoprecipitation assay showed that Hrp3 directly associates with mat3 chromatin. Thus, our results strongly suggest that Hrp3 is involved in heterochromatin silencing and plays a direct role as a chromatin remodeling factor at mat3 in vivo. PMID- 12127989 TI - c-Fos is a surface pressure-dependent diverter of phospholipase activity. AB - c-Fos, a transcription factor, associates to endoplasmic reticulum and modulates phospholipid biosynthesis. Its surface thermodynamic properties allow it to differentially interact with phospholipid monolayers with a selective dependence on the lipid polar head group and the lateral surface pressure. We explored the c Fos ability to modulate phospholipid degradation by phospholipases (ppPLA2, Bacillus cereus PLC, and sphingomyelinase) using the monolayer technique. Experiments conducted under constant packing conditions show that c-Fos modulates phospholipase activity in a finely tuned way, depending on the membrane intermolecular packing. Surface lateral pressures above 12-16 mN/m induce c-Fos to activate phospholipase A2 and sphingomyelinase, and abolish phospholipase C activity. The effects of c-Fos on other steps of the catalytic process, lag-time and extent, are synergic with those on activity. We show for the first time that c-Fos participates in modulating phospholipid degradation and that it can affect the formation of lipid second messenger products by PLA2, PLC, and sphingomyelinase. PMID- 12127991 TI - Affinity of carbon monoxide to hemoglobin increases at low oxygen fractions. AB - Following systemic inflammation, the lung induces an isoenzyme of heme oxygenase (HO-1), catalyzing carbon monoxide (CO) production through breakdown of heme molecules. However, it is still debated why the paradoxical arterio-venous carboxyhemoglobin (COHb) difference occurs only during critical illness but not in healthy volunteers. To elucidate whether oxygen fractions at (sub-)physiologic ranges alter the affinity of CO to hemoglobin (Hb), we performed an in vitro laboratory experiment, in which we exposed venous blood samples to fixed CO-doses at incrementing oxygen fractions (FiO2). ANOVA demonstrated that the affinity of CO (200 and 400 ppm) to Hb progressively increased with an FiO2 from 0% to 15%, whereas at higher oxygen tensions this effect vanished. This might explain why the arterio-venous COHb difference found in critically ill patients is not reproducible in healthy adults, since the latter ones are characterized by higher venous oxygen saturations. PMID- 12127992 TI - Uptake of lead and iron by divalent metal transporter 1 in yeast and mammalian cells. AB - Although the divalent metal transporter (DMT1) was suggested to transport a wide range of metals in Xenopus oocytes, recent studies in other models have provided contrasting results. Here, we provide direct evidence demonstrating that DMT1 expressed in yeast mutants defective for high affinity iron transport facilitates the transport of iron with an 'apparent K(m)' of approximately 1.2 microM, and transport of lead with an 'apparent K(m)' of approximately 1.8 microM. DMT1 dependent lead transport was H(+)-dependent and was inhibited by iron. Human embryonic kidney fibroblasts (HEK293 cells) overexpressing DMT1 also showed a higher uptake of lead than HEK293 cells without overexpressing DMT1. These results show that DMT1 transports lead and iron with similar affinity in a yeast model suggesting that DMT1 is a transporter for lead. PMID- 12127993 TI - Monocyte-derived soluble protein confers 5-lipoxygenase activity Ca2+-dependent. AB - 5-Lipoxygenase (5-LO) is a Ca2+-stimulated enzyme that initializes the formation of proinflammatory leukotrienes from arachidonic acid (AA). In this report, we demonstrate that a soluble protein of the monocytic cell line Mono Mac 6 confers 5-LO activity Ca2+-dependent in vitro. Thus, in broken cell preparations of human polymorphonuclear leukocytes (PMNL) and rat basophilic leukemia (RBL)-1 cells, 5 LO converted AA (>20 microM) in the absence of Ca2+, whereas Ca2+ was absolutely required for 5-LO activity in broken cell preparations of MM6 cells. 5-LO partially purified from MM6 cells was substantially active in the absence of Ca2+. Recombination experiments revealed that the cytosolic fraction of MM6 cells contains a factor that suppresses the activity of partially purified 5-LO from PMNL, RBL-1, and MM6 cells in the absence but not in the presence of Ca2+. Further characterization showed that this factor is a 80-100 kDa heat-sensitive protein. PMID- 12127994 TI - C-terminal modification of 6-phosphofructo-1-kinase from Saccharomyces cerevisiae and its influence on enzyme structure and activity. AB - Studies on limited proteolysis of 6-phosphofructo-1-kinase (Pfk-1) from Saccharomyces cerevisiae led to the suggestion that the C-terminal part of the alpha-subunit must contribute to the stabilisation of the octameric enzyme structure. To analyse the role of the C-terminus in vivo, the respective terminus of one of both types of subunits of Pfk-1 was sequentially truncated or extended. These modifications resulted in a decrease of the protein level of the mutated subunit and of the specific enzyme activity in the cell-free extract as well as in changes of the kinetic properties. Size exclusion HPLC demonstrated that the modified subunit is still able to assemble with the native counterpart generating an enzymatically active hetero-octamer. On the basis of our results we assume that the C-termini are important for the three-dimensional structure of the subunits determining their susceptibility to proteolysis and the ability to assembly to an active, oligomeric Pfk-1. PMID- 12127995 TI - Expression and binding properties of the two mannose-6-phosphate receptors differ during perinatal development in rat liver. AB - Mammalian tissues express both cation-dependent (CD-MPR) and cation-independent (CI-MPR) mannose-6-phosphate receptors, which mediate the targeting of acid hydrolases to lysosomes. The coexistence of the two receptors in all cell types and tissues is still poorly understood. To determine whether these receptors might play a role in maturation, we studied their expression and binding properties in rat liver during perinatal development. CI-MPR expression decreases progressively from 18-day fetuses to adults, whereas the CD-MPR showed a transient decrease in newborn and at the 5th day after birth. Immunostaining of the tissues showed that both receptors localize to hepatocytes at all the ages and, additionally, the CD-MPR was reactive in megakaryocytes at early stages. Binding assays showed differences in the B(max) and K(D) values between the ages studied. These results demonstrate that both receptors change differentially during perinatal development, suggesting that they play distinct roles during organ maturation. PMID- 12127996 TI - Galactose-binding lectin from the seeds of champedak (Artocarpus integer): sequences of its subunits and interactions with human serum O-glycosylated glycoproteins. AB - Our group has previously reported the isolation, partial characterisation, and application of a Galbeta1-3GalNAc- and IgA1-reactive lectin from the seeds of champedak (Artocarpus integer). In the present study, we have subjected the purified lectin to reverse-phase high performance liquid chromatography and sequenced its subunits. Determination of the N-terminal sequence of the first 47 residues of the large subunit demonstrated at least 95% homology to the N terminal sequence of the alpha chains of a few other galactose-binding Artocarpus lectins. The two smaller subunits of the lectin, each comprised of 21 amino acid residues, demonstrated minor sequence variability. Their sequences were generally comparable to the beta chains of the other galactose-binding Artocarpus lectins. When used to probe human serum glycopeptides that were separated by two dimensional gel electrophoresis, the lectin demonstrated strong apparent interactions with glycopeptides of IgA1, hemopexin, alpha2-HS glycoprotein, alpha1-antichymotrypsin, and a few unknown glycoproteins. Immobilisation of the lectin to Sepharose generated an affinity column that may be used to isolate the O-glycosylated serum glycoproteins. PMID- 12127997 TI - Upregulation of monocyte chemoattractant protein 1 and effects of transforming growth factor-beta 1 in Peyronie's disease. AB - Peyronie's disease (PD) is characterized by fibrosis in the tunica albuginea (TA) of the penis, which becomes bent as a result. We have previously shown that transforming growth factor-beta 1 (TGF-beta1) is upregulated in the TA of patients with PD and can initiate PD-like lesions in rat models. In this study we isolated three types of fibroblasts: P cells from the lesions of PD patients, C cells from the normal-appearing areas of the TA of the same patients, and N cells from the TA of patients without PD. We examined these cells for the expression of two fibrogenic cytokines, connective tissue growth factor (CTGF), and Monocyte Chemoattractant Protein 1 (MCP-1). We found that CTGF was expressed at similar levels in P, C, and N cells, whereas MCP-1 was significantly more expressed in P cells than in C cells and more in C cells than in N cells. Higher MCP-1 expression was also found in the lesions than in normal TA. Treatment with TGF beta1-induced higher expression of MCP-1 but not CTGF in all three types of cells, with C cells being the most responsive. Based on these observations, we propose that MCP-1 could be a novel therapeutic target in PD. PMID- 12127998 TI - An unusual orientation for Tyr75 in the active site of the aspartic proteinase from Saccharomyces cerevisiae. AB - The structures of the native Saccharomyces cerevisiae proteinase A have been solved by molecular replacement in the monoclinic and trigonal crystal forms and refined at 2.6-2.7A resolution. These structures agree overall with those of other uninhibited aspartic proteinases. However, an unusual orientation for the side chain of Tyr75, a conserved residue on the flexible "flap" that covers the active site and is important for the activity of these enzymes, was found in the trigonal crystals. A similar conformation of Tyr75 occupying the S1 substrate binding pocket was previously reported only for chymosin (where it was interpreted as representing a "self-inhibited" state of the enzyme), but for no other aspartic proteinases. Since this orientation of Tyr75 has now been seen in the structures of two members of the family of aspartic proteinases, it might indicate that the placement of that residue in the S1 substrate-binding pocket might have some functional significance, analogous to what was seen for self inhibited structures of serine proteinases. PMID- 12127999 TI - Immunomagnetic capture of lens membrane fractions containing steroid binding protein. AB - This study describes the use of magnetic Dynabeads to purify microsomes from a crude microsomal fraction. A 28 kDa membrane-associated protein is proposed to mediate the binding of progesterone and other steroid hormones to ocular lens membranes and the rapid-nongenomic actions of these steroids. The subcellular location of this membrane steroid binding protein (MSBP) was probed by capture of organelles containing MSBP by magnetic beads displaying an antibody to a cytoplasmic domain of the protein. The beads were exposed to a crude microsomal fraction from lens epithelia. Western blotting was used to identify captured organelles and confirm the presence of MSBP. Microsomes and trace fiber cell plasma membrane were captured. Microsomes contained the 28 kDa MSBP. Lens fiber cell membrane contained a 55 kDa immunoreactive protein. The role of this serendipitously recognized protein in binding of steroids is unknown. PMID- 12128000 TI - Functions of cannabinoid receptors in the hippocampus. AB - Marijuana smoking is recognised to impair human cognition and learning, but the mechanisms by which this occurs are not well characterised. This article focuses exclusively on the hippocampus to review the effects of cannabinoids on hippocampal function and evaluate the evidence that hippocampal cannabinoid receptors play a role in learning and formation of memory. Activation of cannabinoid receptors inhibits release of a variety of neurotransmitters, and modulates a number of intrinsic membrane conductances. Suppression of inhibitory GABAergic synaptic transmission has been repeatedly described, but whether there is also control of excitatory glutamatergic transmission is more controversial. The recognition that the commonly used WIN55,212-2 also acts via non-cannabinoid receptors may help resolve this issue. The involvement of endocannabinoids in depolarisation induced suppression of inhibition (DSI) and the demonstration that activation of metabotropic glutamate receptors can stimulate endocannabinoid release have provided the first insights into the physiological roles of the cannabinoids. Cannabinoids have consistently been reported to inhibit high frequency stimulation induced synaptic long-term potentiation but the experimental design of most behavioural experiments have meant it is not possible to categorically demonstrate a role for hippocampal cannabinoid receptors in learning and memory. PMID- 12128001 TI - Imipramine treatment up-regulates the expression and function of mGlu2/3 metabotropic glutamate receptors in the rat hippocampus. AB - We examined the effect of a chronic imipramine treatment (10 mg/kg, i.p., once daily for 21 days) on the expression and function of metabotropic glutamate (mGlu) receptors in discrete regions of the rat brain. Chronic imipiramine treatment up-regulated the expression of mGlu2/3 receptor proteins in the hippocampus, nucleus accumbens, cerebral cortex and corpus striatum. Expression of mGlu1a receptor protein was increased exclusively in the hippocampus, whereas no changes in the expression of mGlu4 and mGlu5 receptors or Homer-1a protein were detected. Using hippocampal slices, we examined the stimulation of polyphosphoinositide (PI) hydrolysis induced by mGlu receptor agonists in control and imipramine-treated rats. Imipramine treatment amplified the PI response to the non subtype-selective mGlu receptor agonist, 1S,3R-aminocyclopentane-1,3 dicarboxylated (1S,3R-ACPD) in both hippocampal and cortical slices, but failed to affect the response to the selective mGlu1/5 receptor agonist, S-3,5 dihydroxyphenylglycine (DHPG). Amplification was restored when DHPG was combined with the selective mGlu2/3 receptor agonist, LY379268. In addition, 1S,3R-ACPD stimulated PI hydrolysis was no longer enhanced in imipramine-treated rats when the mGlu2/3 component of the PI response was abrogated by the antagonist, LY341495. In contrast, the ability of LY379268 to inhibit forskolin-stimulated cAMP formation was reduced in hippocampal slices of rats chronically treated with imipramine. Taken together, these results suggest that neuroadaptive changes in the expression and function of mGlu2/3 receptors occur in response to chronic antidepressants. PMID- 12128002 TI - Effect of chronic imipramine or electroconvulsive shock on the expression of mGluR1a and mGluR5a immunoreactivity in rat brain hippocampus. AB - Previous studies showed that chronic electroconvulsive shock (ECS) or imipramine treatment induced a subsensitivity of group I metabotropic glutamate receptors (mGluR) in hippocampus. In the present study effects of antidepressant treatment on the expression of mGluR1a and mGluR5a, belonging to the group I mGluR, were investigated in rat brain hippocampus using immunohistochemical and Western blot methods, respectively. Male Wistar rats were treated singly or chronically for 21 days with imipramine, 10 mg/kg, twice daily; with ECS (90 mA, 50 Hz, 0.5 s) every second day; or with haloperidol, 1.2 mg/kg, once daily. Appropriate controls were injected with saline. Rats were sacrificed 24 h after the last treatment and their hippocampi were taken out for analysis. It was found that the mGluR1a immunoreactivity expression increased significantly in Ammon's horn (CA) regions after chronic ECS. The most pronounced effect was observed in the CA3. No significant effects were found after single treatment or after haloperidol. The expression of mGluR5a increased significantly after chronic imipramine in the CA1 and after chronic ECS in the CA3 region. The results obtained indicate an influence of antidepressant treatment on group I mGluR. This increase in the receptor protein level may be a compensatory mechanism developing after chronic treatment. PMID- 12128003 TI - Synergistic effect of uncompetitive NMDA receptor antagonists and antidepressant drugs in the forced swimming test in rats. AB - In spite of intensive research, the problem of treating antidepressant-resistant depressive patients has not yet been solved. The authors previously reported that combined administration of imipramine and the uncompetitive NMDA receptor antagonist amantadine reduced immobility time in the forced swimming test in rats to a much greater extent than either treatment alone. The present paper investigates the possibility of synergistic interactions between three antidepressants (imipramine, venlafaxine, fluoxetine) with three uncompetitive NMDA receptor antagonists (amantadine, memantine and neramexane). Most combinations resulted in synergistic (hyperadditive) antidepressive-like effects in the forced swim test. Most interesting was the observation that fluoxetine, which was inactive when given alone, showed a positive effect when combined with amantadine (10 and 20 mg/kg), memantine (2.5 and 5 mg/kg) or neramexane (2.5 and 5 mg/kg). The specificity of these observations is supported by control open field studies, which demonstrated no significant increase, or even a decrease in general locomotion after coadministration of the compounds. The present results suggest that the combination of traditional antidepressant drugs and NMDA receptor antagonists may produce enhanced antidepressive effects, and this is of particular relevance for antidepressant-resistant patients. PMID- 12128004 TI - Repeated treatment with antidepressants differentially alters 5-HT1A agonist stimulated [35S]GTP gamma S binding in rat brain regions. AB - Electrophysiological studies have led to the proposal that the neurobiological mechanism(s) underlying drug therapy of anxiety and depression involve(s) regionally specific adaptations in 5-HT(1A) receptor sensitivity. Depending on the drug utilized, a decrease in sensitivity of inhibitory somatodendritic autoreceptors, an increase in sensitivity of postsynaptic receptors, or both alterations, occur after several weeks of treatment. This hypothesis was tested using N,N-dipropyl-5-carboxamidotryptamine-stimulated guanosine-5'-O-(3 thio)triphosphate ([(35)S]GTPgammaS) binding assessed by autoradiography. Rats were treated for 21 days with one of four different anxiolytic/antidepressant drugs (in mg/kg): fluoxetine (10), imipramine (10), clorgyline (1), ipsapirone (2 x 20) or saline. Three brain regions rich in 5-HT(1A) receptors were examined: the dorsal raphe (somatodendritic), the dorsal hippocampus (postsynaptic) and the lateral septum (postsynaptic). Only imipramine (+17%) and fluoxetine (+54%) significantly increased agonist-stimulated binding in the dorsal hippocampus; all drugs except imipramine significantly decreased binding in the dorsal raphe (-19 to -41%). These results generally support the concept of a net enhancement of hippocampal 5-HT neurotransmission via one or more 5-HT receptor subtypes. The most consistent effect, however, was a significant decrease in stimulated [(35)S]GTPgammaS binding in the lateral septum after all four treatments (-14 to 23%), suggesting that this may be a heretofore unrecognized common outcome of antidepressant treatment deserving further study. PMID- 12128005 TI - Enhancement of long-term potentiation in the rat hippocampus following cocaine exposure. AB - The neural adaptations involved in initiating and maintaining the long-term consequences of utilizing drugs of abuse are the subject of intense investigation. It is commonly suggested that the neural plasticity mechanisms underlying physiological phenomena such as learning and memory may also be engaged when drug addiction occurs. The effect of cocaine on one prominent cellular mechanism for learning/memory, long-term potentiation (LTP), was assessed in the CA1 region of the rat hippocampus. Hippocampal slices obtained from animals treated in vivo for five days with cocaine (15 mg/kg i.p., daily) exhibited enhanced LTP vs saline treated controls. We suggest that this example of cocaine-induced enhancement of LTP provides an example of how synaptic plasticity mechanisms may be altered in a manner that contributes to the behavioral outcomes expressed, following exposure to psychostimulants. PMID- 12128006 TI - Involvement of sigma receptors in the behavioral effects of cocaine: evidence from novel ligands and antisense oligodeoxynucleotides. AB - Pharmacological and molecular biological tools were used to validate the involvement of sigma receptors in the actions of cocaine. Radioligand binding studies demonstrated significant levels of sigma receptors in the brain and heart, where cocaine interacts preferentially with the sigma(1) subtype. In behavioral pharmacological studies using mice, nine novel sigma receptor antagonists significantly attenuated cocaine-induced convulsions, while structural analogs with weak interactions with sigma receptors were ineffective. In contrast to the protection provided by the antagonists, a classical sigma receptor agonist exacerbated the convulsive effects of cocaine. The antagonists also attenuated cocaine-induced lethality, with the best compound protecting against death even when administered as a post-treatment. At doses where the antagonists had no effect on baseline locomotor activity, they significantly attenuated the locomotor stimulatory effects of cocaine, suggesting their ability to block the psychomotor as well as the toxic effects of cocaine. To further validate that the anti-cocaine effects were achieved by interfering with cocaine's access to sigma receptors, antisense oligodeoxynucleotides against sigma(1) receptors were shown to attenuate the convulsive and locomotor stimulatory effects of cocaine. Together, the studies support the involvement of sigma receptors, particularly the sigma(1) subtype, in the behavioral effects of cocaine. PMID- 12128007 TI - Estradiol protects dopaminergic neurons in a MPP+Parkinson's disease model. AB - The prevalence of Parkinson's disease is higher in males than in females. Although the reason for this gender difference is not clear, the level of female steroid hormones or their receptors may be involved in the pathogenesis. The estrogen receptor subtype expressed in the midbrain is limited to the novel beta subtype, whose role in the central nervous system has not been resolved. We demonstrated that ligand-activated estrogen receptor beta suppressed dopaminergic neuronal death in an in vitro Parkinson's disease model which uses 1-methyl-4 phenylpyridinium ions (MPP(+)). MPP(+) treatment caused the upregulation of c-Jun amino-terminal kinase (JNK) and dopaminergic neuronal death, the latter being blocked by curcumin, an inhibitor of the c-Jun/AP-1 cascade. 17alpha- and 17beta estradiol both protected dopaminergic neurons from MPP(+)-induced neuronal death and this was blocked by a pure antagonist of the estrogen receptor, ICI 182,780, but not by an inhibitor of estrogen receptor dimerization, YP537. These data indicated that the neuroprotection provided by 17alpha-estradiol was via inhibitory transcriptional regulation at the activator protein-1 (AP-1) site mediated by estrogen receptor beta. Thus, 17alpha-estradiol is a suitable candidate for neuroprotective therapy of Parkinson's disease because it is associated with few undesirable feminizing effects. PMID- 12128008 TI - Presynaptic mu-opioid receptors regulate a late step of the secretory process in rat ventral tegmental area GABAergic neurons. AB - Gamma-aminobutyric acid (GABA)-containing interneurons of the ventral tegmental area (VTA) regulate the activity of dopaminergic neurons. These GABAergic interneurons are known to be innervated by synaptic terminals containing enkephalin, an endogenous ligand of mu-opioid receptors. Bath application of mu opioid receptor agonists inhibits the activity of VTA GABAergic neurons but the mechanism whereby mu-opioid receptors regulate synaptic GABA release from these neurons has not been directly identified. Using cultured VTA neurons we have confirmed that mu-opioid receptor agonists inhibit synaptic GABA release. DAMGO, a selective mu-opioid receptor agonist, had four distinct effects on GABAergic IPSCs: (1) it inhibited the frequency and amplitude of spontaneous IPSCs (sIPSCs), (2) it reduced the amplitude of IPSCs evoked by single action potentials, (3) it inhibited the frequency, but not the amplitude of miniature IPSCs (mIPSCs), and (4) DAMGO inhibited mIPSCs evoked by ionomycin, a Ca(2+) ionophore. The inhibition of action potential-evoked IPSCs and of spontaneous and ionomycin-evoked mIPSCs by DAMGO was prevented by the K(+) channel blocker, 4 aminopyridine (4-AP). In conclusion, our work shows that one of the mechanisms through which mu-opioid receptors inhibit GABA release by VTA neurons is through inhibition of the secretory process at the nerve terminal level. In addition, considering that ionomycin stimulates exocytosis through a mechanism that should be insensitive to membrane polarization, our experiments with 4-AP suggest that K(+) channels are implicated in the inhibition of the efficacy of the secretory process by mu-opioid receptors. PMID- 12128009 TI - Enhanced antinociception by intrathecally-administered morphine in histamine H1 receptor gene knockout mice. AB - We previously reported that histamine H(1) receptor gene knockout mice (H1KO) showed lower spontaneous nociceptive threshold to pain stimuli when compared to wild-type mice. The objective of the present study was to examine the antinociceptive effect of intrathecally-administered morphine in H1KO mice. The antinociceptive effects of morphine were examined using assays for thermal (tail flick, hot-plate, paw-withdrawal), mechanical (tail-pressure) and chemical nociception (formalin and capsaicin tests) using H1KO and wild-type mice. In these nociceptive assays, intrathecally-administered morphine produced significant antinociceptive effects in wild-type mice. The antinociceptive effect produced by intrathecally administered morphine was enhanced in the knockout mice. We also examined the effect of an histamine H(1) receptor antagonist, an active (d-) isomer of chlorpheniramine, on morphine-induced antinociception in ICR mice. The intrathecal co-administration of d-chlorpheniramine enhanced the effect of morphine in all nociceptive assays examined. The pharmacological experiments using d-chlorpheniramine further substantiate the evidence for the histamine H(1) receptor-mediated suppression of morphine-induced antinociception. These results suggest that existing H(1) receptors play an inhibitory role in morphine-induced antinociception at the spinal cord level. PMID- 12128011 TI - Inhibitory effects of cilnidipine on peripheral and brain N-type Ca2+ channels expressed in BHK cells. AB - We investigated differences in electrophysiological characteristics between peripheral and central N-type Ca(2+) channels, containing alpha(1B-a) and a(1B c), respectively. In addition, we examined the inhibitory effects of cilnidipine, a dihydropyridine (DHP) derivative, on both channels. Both alpha(1B) subunits were transiently expressed in BHK cells, and then analyzed using the whole-cell patch-clamp technique. The current-voltage relationship showed that alpha(1B-c) currents were activated at more negative potentials than alpha(1B-a) currents. The voltage-dependent steady-state inactivation and activation showed that the V(1/2) values for inactivation and activation of alpha(1B-c) (-88.5+/-1.3 and 33.2+/-1.3 mV) were both significantly more negative than those for alpha(1B-a) ( 83.3+/-1.3 and -27.9+/-2.3 mV). Despite the different electrophysiological characteristics of these two N-type channels, cilnidipine blocked both with similar potency within the range 0.1 to 10 microM. Furthermore, cilnidipine had no effect on the I-V relationships or the steady-state inactivation curves. Our data indicate that the spliced positions of alpha(1B-a) and a(1B-c) may affect not only their voltage-sensing abilities but also the kinetics of channel activation and inactivation. The data also suggest that cilnidipine binds to sites independent of those controlling voltage-sensing and channel kinetics in these alpha(1B) subunits. PMID- 12128010 TI - Receptor-mediated internalization of [3H]-neurotensin in synaptosomal preparations from rat neostriatum. AB - Following its binding to somatodendritic receptors, the neuropeptide neurotensin (NT) internalizes via a clathrin-mediated process. In the present study, we investigated whether NT also internalizes presynaptically using synaptosomes from rat neostriatum, a region in which NT1 receptors are virtually all presynaptic. Binding of [(3)H]-NT to striatal synaptosomes in the presence of levocabastine to block NT2 receptors is specific, saturable, and has NT1 binding properties. A significant fraction of the bound radioactivity is resistant to hypertonic acid wash indicating that it is internalized. Internalization of [(3)H]-NT, like that of [(125)I]-transferrin, is blocked by sucrose and low temperature, consistent with endocytosis occurring via a clathrin-dependent pathway. However, contrary to what was reported at the somatodendritic level, neither [(3)H]-NT nor [(125)I] transferrin internalization in synaptosomes is sensitive to the endocytosis inhibitor phenylarsine oxide. Moreover, treatment of synaptosomes with monensin, which prevents internalized receptors from recycling to the plasma membrane, reduces [(3)H]-NT binding and internalization, suggesting that presynaptic NT1 receptors, in contrast to somatodendritic ones, are recycled back to the plasma membrane. Taken together, these results suggest that NT internalizes in nerve terminals via an endocytic pathway that is related to, but is mechanistically distinct from that responsible for NT internalization in nerve cell bodies. PMID- 12128012 TI - Cytoskeleton disruption causes apoptotic degeneration of dentate granule cells in hippocampal slice cultures. AB - Colchicine, a potent microtubule-depolymerizing agent, is well known to selectively kill dentate granule cells in the hippocampal formation in vivo. Using organotypic cultures of rat entorhino-hippocampal slices, we confirmed that in vitro exposure to 1 microM and 10 microM of colchicine reproduced a specific degeneration of the granule cells after 24 h. Similar results were obtained with other types of microtubule-disrupting agents, i.e., nocodazole, vinblastine, and Taxol. Interestingly, the actin-depolymerizing agents cytochalasin D and latrunculin A also elicited selective neurotoxicity in the dentate gyrus without affecting survival of hippocampal pyramidal cells. The selective pattern of degeneration was observable 24 h after a brief treatment with the toxins as short as 5 min, but this delayed neuronal death was unlikely to be a result of excitotoxicity because it was virtually unaffected by glutamate receptor antagonists, tetrodotoxin, or extracellular Ca(2+)-free conditions. The damaged tissues contained a large number of TUNEL-positive neurons and exhibited an increased level in caspase-3-like activity, suggesting that cytoskeleton disruption triggers an apoptosis-like process in dentate granule cells. Thus, this study may provide a basis for understanding the distinctive mechanism that supports granule cell survival. PMID- 12128013 TI - Surround suppression in the human visual cortex: an analysis using magnetoencephalography. AB - The responses of neurons in the primate and cat primary visual cortices (V1s) to the stimuli within their classical receptive fields (CRFs) are markedly suppressed by the surrounding stimuli outside CRFs. In the present study, we show that a similar suppressive effect occurs for visually evoked magnetic responses in the human visual cortex. The initial peak amplitude of the magnetic response (at a latency of around 90 ms) to a test grating accompanied by high-contrast surround gratings was smaller than that for the test without the surround. Current source localization with a single dipole model indicated that the initial response originated from cortical activity near the occipital pole in the contralateral hemisphere to the visual stimulation. The peak amplitude for the test decreased with increasing surround contrast, and increased with increasing test contrast. The contrast dependence and the early development of the surround suppression were in agreement with the results of the V1 single-cell studies of monkeys and cats. We suggest that the surround suppression of the initial peak amplitude of the magnetic response may be ascribed to the inhibition of the neural activity at the early processing stage(s), presumably at V1, in the human visual cortex. PMID- 12128014 TI - Spatial frequency discrimination in cyclopean vision. AB - It is well known that inspecting a cyclopean grating causes threshold for detecting a subsequently presented test cyclopean grating to be elevated, and that the threshold elevation is greatest at the adapting frequency. We report here that spatial frequency discrimination threshold is also elevated, but the elevation is greatest at frequencies offset from the adapting frequency, and the elevation at the adapting frequency is near-zero. We conclude that discrimination threshold is determined by the relative activity of cyclopean frequency-tuned channels, and suggest that relative activity is computed at an opponent-frequency stage. Discrimination threshold for cyclopean gratings was 2.5-4% for two observers, and remained approximately constant over the range 0.16-2.0 cycles/ degrees. Discrimination threshold for luminance-defined gratings was only slightly lower. Discrimination threshold was approximately independent of the grating's peak-to-peak disparity over a range of 45:1 for one observer and 17:1 for another. This finding as well as the low value of discrimination threshold are consistent with an opponent-process model. The dot density of every cyclopean grating used was chosen bearing in mind our finding that three or more spatial samples per grating cycle are required before sampling effects can be ignored. PMID- 12128015 TI - Measurement of generalization fields for the recognition of biological motion. AB - The human visual system processes complex biological motion stimuli with high sensitivity and selectivity. The characterization of spatio-temporal generalization in the perception of biological motion is still a largely unresolved problem. We present an experiment that investigates how the visual system responds to motion stimuli that interpolate spatio-temporally between natural biological motion patterns. Inspired by analogous studies in stationary object recognition, we generated stimuli that interpolate between natural perceptual categories by morphing. Spatio-temporal morphs between natural movement patterns were obtained with a technique that allows to calculate linear combinations of spatio-temporal patterns. The weights of such linear combinations define a linear metric space over the set of generated movement patterns, so that the spatio-temporal similarity of the motion patterns can be quantified. In our experiments, we found smooth and continuous variation of the categorization probabilities with the weights of the prototypes in the morphs. For bipedal locomotion patterns we could accurately predict the perceived properties of the morphs by linear combinations of the perceived properties of the prototypes. Such predictions were not possible for morphs between locomotion and very dissimilar movements. We conclude that the visual system shows generalization within classes of motion patterns with similar basic structure, such as bipedal locomotion. PMID- 12128016 TI - Content and context of monocular regions determine perceived depth in random dot, unpaired background and phantom stereograms. AB - Perceived depth was measured for three-types of stereograms with the colour/texture of half-occluded (monocular) regions either similar to or dissimilar to that of binocular regions or background. In a two-panel random dot stereogram the monocular region was filled with texture either similar or different to the far panel or left blank. In unpaired background stereograms the monocular region either matched the background or was different in colour or texture and in phantom stereograms the monocular region matched the partially occluded object or was a different colour or texture. In all three cases depth was considerably impaired when the monocular texture did not match either the background or the more distant surface. The content and context of monocular regions as well as their position are important in determining their role as occlusion cues and thus in three-dimensional layout. We compare coincidence and accidental view accounts of these effects. PMID- 12128017 TI - Orientation-selective summing mechanisms revealed in visual search. AB - We investigated properties of the neural mechanisms that mediate detection of complex grating targets in an orientation-based visual search task. Targets and distractors were composed of small patches of compound sinusoidal gratings. Components were chosen to differ enough in spatial frequency to stimulate separate and independent mechanisms at the primary cortical layer of processing. The orientations of the components were both vertical in distractor patches. In the uncrossed condition, both components of the target tilted either 3 degrees left or right. In the crossed condition, one component of the target tilted left and the other tilted right. Search was faster and more accurate in the uncrossed condition, ruling out mediation either by V1-like tuned mechanisms or by a higher level mechanism that signals differences in orientation. Results were consistent with two classes of mid-level summing mechanisms. We argue that mid-level mechanisms such as these may be the neural substrate for conceptual orientation feature maps. PMID- 12128018 TI - A psychophysical correlate of contrast dependent changes in receptive field properties. AB - Recent physiological investigations have demonstrated that a neuron's area of spatial summation can vary depending on stimulus contrast. Specifically, when the same stimulus is presented to a neuron at a low contrast, the area of summation (or neuron's receptive field) can increase by at least a factor of two, compared to that estimated with a high contrast stimulus. We sought to examine this phenomenon psychophysically by using an orientation discrimination task carried out in the presence of contextual stimuli. We have found previously that orientation discrimination thresholds for a sine-wave grating are elevated by the presence of a surround pattern of similar orientation (with an offset) and spatial frequency. However, when these patterns were separated by a gap of mean luminance exceeding roughly 1 deg, thresholds dropped to the level measured using the center pattern alone. Here, we examined the surround pattern's effect on orientation thresholds as a function of the contrast of the center and surround. We find that when both are presented at a low contrast, the detrimental influence of the surround on orientation thresholds is maintained over larger gap separations. We also find that the spatial frequency and orientation selectivity of the surround's masking effect on orientation thresholds is broader at low contrast than at high contrast. Although the results support the idea of a spatial reorganization of the mechanisms involved in the task at low contrast, these changes are insufficient, in and of themselves, to account for the data. We suggest that additional influences possibly reflecting image segmentation also affect performance. PMID- 12128019 TI - M-cone opsin gene number does not correlate with variation in L/M-cone sensitivity. AB - Molecular genetic studies demonstrate that the human cone opsin gene array on the q-arm of the X-chromosome typically consists of one long-wave-sensitive (L) cone opsin gene and from one to several middle-wave-sensitive (M) cone opsin genes. Although the presence of the single L-cone opsin gene and at least one M-cone opsin gene is essential for normal red-green colour discrimination, the function of the additional M-cone opsin genes is still unclear. To investigate whether any variations in phenotype correlate with differences in the number of M-cone opsin genes, we selected 13 normal trichromat males, for whom four independent molecular techniques have exactly determined their number of M-cone opsin genes, ranging from one to four. Their phenotype was characterized by estimating their foveal L- to M-cone ratio from heterochromatic flicker photometric (HFP) thresholds, by measuring the wavelength corresponding to their 'unique yellow', and by determining their L- and M-cone modulation thresholds (CMTs). No correlation was found between these psychophysical measures and the number of M cone opsin genes. Although, we found a reasonably good correlation between the L/M-cone ratios based on HFP and on CMT, we did not find any correlation between the estimated L/M-cone ratios and the settings of 'unique yellow'. Our results accord with previous molecular genetic studies that suggest that only the first two genes in the X-linked opsin gene array are expressed. PMID- 12128021 TI - Texture segmentation performance related to cortical geometry. AB - There are two prevailing explanations for the foveal deficit in texture segmentation reported in previous works. One is based on the spatial and temporal properties of the stimuli, which means in terms of physiology a strong contribution of the Magno-channel. The other one is purely spatial and assigns filters of different bandwidths to each eccentricity in the visual field. We have challenged the first explanation experimentally by using isoluminant stimuli. The central performance drop persisted although the Magno-channel is known to respond weakly to stimuli with low luminance contrast. Therefore, we agreed with the spatial explanation. But instead of the abstract filter theories from previous works we propose a computational neural model assuming local lateral interactions in a cortical map model. The psychophysical performance measures could be directly related to geometric properties of the primary visual cortex concerning its mapping geometry and its intrinsic interaction width. Our model accounts quantitatively for our own psychophysical data as well as for others from literature. In general, we claim that the high foveal retino-cortical magnification maps texture elements too far away from each other for being compared by local processes. PMID- 12128020 TI - Voluntary saccadic eye movements in humans studied with a double-cue paradigm. AB - In the classic double-step paradigm, subjects are required to make a saccade to a visual target that is briefly presented at one location and then shifted to a new location before the subject has responded. The saccades in this situation are "reflexive" in that they are made in response to the appearance of the target itself. In the present experiments we adapted the double-step paradigm to study "voluntary" saccades. For this, several identical targets were always visible and subjects were given a cue to indicate that they should make a saccade to one of them. This cue was then changed to indicate another of the targets before the subject had responded: double-cue (DC) paradigm. The saccadic eye movements in our DC paradigm had many features in common with those in the double-step paradigm and we show that apparent differences can be attributed to the spatio temporal arrangements of the cues/targets rather than to any intrinsic differences in the programming of these two kinds of eye movements. For example, a feature of our DC paradigm that is not seen in the usual double-step paradigm is that the second cue could cause transient delays of the initial saccade, and these delays still occurred when the second cue was reflexive--provided that it was at the fovea (as in our DC paradigm) and not in the periphery (as in the usual double-step paradigm). Thus, the critical factor for the delay was the retinal (foveal) location of the second cue/target--not whether it was cognitive or reflexive--and we argue that the second cue/target is here acting as a distractor. We conclude that the DC paradigm can be used to study the programming of voluntary saccades in the same way that the double-step paradigm can be used to study reflexive saccades. PMID- 12128022 TI - New, sensitive window on abnormal spatial vision: rarebit probing. AB - Clinical tests have a poor sensitivity to low to moderate degrees of neuro-visual damage, possibly because their test targets involve numerous receptive fields. A new test used briefly exposed microdots of high contrast. Multiple visual field areas were probed repeatedly, with ever-new microdot positions. Normal subjects responded to a median 96.0% of probes. Patients with different visual field defects missed larger numbers of probes within defects and the deeper the defects, the larger the number of misses. Patients with minor chiasmal lesions averaged 1.8 times larger defects in microdot perimetry than in high-pass resolution perimetry, indicating superior sensitivity to minor damage. PMID- 12128023 TI - Codon adaptation and synonymous substitution rate in diatom plastid genes. AB - Diatom plastid genes are examined with respect to codon adaptation and rates of silent substitution (Ks). It is shown that diatom genes follow the same pattern of codon usage as other plastid genes studied previously. Highly expressed diatom genes display codon adaptation, or a bias toward specific major codons, and these major codons are the same as those in red algae, green algae, and land plants. It is also found that there is a strong correlation between Ks and variation in codon adaptation across diatom genes, providing the first evidence for such a relationship in the algae. It is argued that this finding supports the notion that the correlation arises from selective constraints, not from variation in mutation rate among genes. Finally, the diatom genes are examined with respect to variation in Ks among different synonymous groups. Diatom genes with strong codon adaptation do not show the same variation in synonymous substitution rate among codon groups as the flowering plant psbA gene which, previous studies have shown, has strong codon adaptation but unusually high rates of silent change in certain synonymous groups. The lack of a similar finding in diatoms supports the suggestion that the feature is unique to the flowering plant psbA due to recent relaxations in selective pressure in that lineage. PMID- 12128024 TI - Genetic subdivision and biogeography of the Danubian rheophilic barb Barbus petenyi inferred from phylogenetic analysis of mitochondrial DNA variation. AB - The barb Barbus petenyi is a cyprinid widely distributed throughout the mountain regions in the Danube River basin. Phylogenetic analysis of the DNA sequence variation at the mitochondrial cytochrome b gene over much of this range yielded three deep-branching (5.9-9.4% average divergence), well-supported haplotype clades with mutually exclusive geographic distributions and divergence times estimated to be in the Tertiary. The clades did not form an altogether monophyletic group as the most divergent one coalesced more recently with haplotypes of phylogenetically close species than with the other B. petenyi haplotypes. This pattern was supported by bootstrap and log-likelihood Shimodaira Hasegawa tests. The other two were sister clades, but their distinctiveness was supported by previous allozyme data. Hence, from a taxonomic point of view, the current recognition of B. petenyi is erroneous, as it does not represent a single evolutionary lineage, and we suggest that three species be recognized instead. Substantial phylogeographic differences were evident among the three putative species, the two more southerly ones displaying significant structure, which suggested that they each survived in several glacial refugia throughout the Pleistocene. The phylogeographic pattern of multiple populations of rheophilic barbs with a history of long-term persistence and separation within the Danube River basin is novel within fishes and provides a hypothesis against which phylogeographic patterns among other similarly distributed rheophilic species may be compared. PMID- 12128025 TI - Paraphyly of the Blue Tit (Parus caeruleus) suggested from cytochrome b sequences. AB - The phylogenetic relationships of the Blue Tit-Azure Tit assemblage (genus Parus; Aves: Passeriformes) were studied using mitochondrial DNA sequences of 24 specimens representing seven subspecies from Eurasia and North Africa. Previous work based on comparative morphological and acoustic data suggested a division of the Blue Tit (Parus caeruleus) into two species. Our analyses clearly indicate that the Blue Tit represents a paraphyletic assemblage, including a European/Middle Asian clade that is the sister group to the Azure Tit (Parus cyanus) and a North African clade. The North African clade (teneriffae subspecies group) is a sister group to the European Blue Tit/Azure Tit clade. We suggest a division of the Blue Tit into two separate species, Eurasian Blue Tit (Parus caeruleus s. str.) and African Blue Tit (Parus teneriffae). However, our data give no support for assigning species rank to Parus cyanus flavipectus, a subspecies of the Azure Tit, as suggested by several authors on morphological grounds. PMID- 12128026 TI - Phylogeography of the Eurasian Willow Tit (Parus montanus) based on DNA sequences of the mitochondrial cytochrome b gene. AB - The phylogeographic relationships of the trans-Palearctic Willow Tit assemblage were studied by obtaining sequence data from the mitochondrial cytochrome b gene from 34 specimens representing nine subspecies from across the species range. Four distinct genetic groups were identified: Parus montanus weigoldicus, P. m. affinis, P. m. songarus, and a clade containing six Eurasian subspecies (ssp. baicalensis, borealis, montanus, restrictus, rhenanus, and sachalinensis). P. m. weigoldicus, P. m. affinis, and P. m. songarus were reciprocally monophyletic and separated from each other and other subspecies by uncorrected genetic distances between 1.9 and 5.8%. The remaining six subspecies were closely related and shared mitochondrial haplotypes, despite marked morphological and acoustical differences, suggesting a rapid evolution of distinct vocalization patterns. The current classification splitting the species into the songarus and montanus subspecies groups is not concordant with our phylogeny. Also, the four regiolect groups, Lowland, Alpine, Siberian, and Sino-Japanese, are not fully mirrored in the phylogeny. Our data suggest that the mono-frequency song type may be ancestral and was retained over a long evolutionary time in certain populations, but was altered or camouflaged by learning processes in others. PMID- 12128027 TI - Relationships within Cornales and circumscription of Cornaceae-matK and rbcL sequence data and effects of outgroups and long branches. AB - Phylogenetic relationships in Cornales were assessed using sequences rbcL and matK. Various combinations of outgroups were assessed for their suitability and the effects of long branches and outgroups on tree topology were examined using RASA 2.4 prior to conducting phylogenetic analyses. RASA identified several potentially problematic taxa having long branches in individual data sets that may have obscured phylogenetic signal, but when data sets were combined RASA no longer detected long branch problems. t(RASA) provides a more conservative measurement for phylogenetic signal than the PTP and skewness tests. The separate matK and rbcL sequence data sets were measured as not containing phylogenetic signal by RASA, but PTP and skewness tests suggested the reverse [corrected]. Nonetheless, the matK and rbcL sequence data sets suggested relationships within Cornales largely congruent with those suggested by the combined matK-rbcL sequence data set that contains significant phylogenetic signal as measured by t(RASA), PTP, and skewness tests. Our analyses also showed that a taxon having a long branch on the tree may not be identified as a "long-branched" taxon by RASA. The long branches identified by RASA had little effect on the arrangement of other taxa in the tree, but the placements of the long-branched taxa themselves were often problematic. Removing the long-branched taxa from analyses generally increased bootstrap support, often substantially. Use of non-optimal outgroups (as identified by RASA) decreased phylogenetic resolution in parsimony analyses and suggested different relationships in maximum likelihood analyses, although usually weakly supported clades (less than 50% support) were impacted. Our results do not recommend using t(RASA) as a sole criterion to discard data or taxa in phylogenetic analyses, but t(RASA) and the taxon variance ratio obtained from RASA may be useful as a guide for improved phylogenetic analyses. Results of parsimony and ML analyses of the sequence data using optimal outgroups suggested by RASA revealed four major clades within Cornales: (1) Curtisia-Grubbia, (2) Cornus-Alangium, (3) Nyssa-Camptotheca-Davidia-Mastixia-Diplopanax, and (4) Hydrangeaceae-Loasaceae, with clades (2) and (3) forming a monophyletic group sister to clade (4) and clade (1) sister to the remainder of Cornales. However, there was not strong bootstrap support for relationships among the major clades. The placement of Hydrostachys could not be reliably determined, although most analyses place the genus within Hydrangeaceae; ML analyses, for example, placed the genus as the sister of Hydrangeeae. Our results supported a Cornales including the systematically problematic Hydrostachys, a Cornaceae consisting of Cornus and Alangium, a Nyssaceae consisting of Nyssa and Camptotheca, a monogeneric Davidiaceae, a Mastixiaceae consisting of Mastixia and Diplopanax, and an expanded Grubbiaceae consisting of Grubbia and Curtisia, and two larger families, Hydrangeaceae and Loasaceae. PMID- 12128028 TI - Phylogenetic dating with confidence intervals using mean path lengths. AB - The mean path length (MPL) method, a simple method for dating nodes in a phylogenetic tree, is presented. For small trees the age estimates and corresponding confidence intervals, calibrated with fossil data, can be calculated by hand, and for larger trees a computer program gives the results instantaneously (a Pascal program is available upon request). Necessary input data are a rooted phylogenetic tree with edge lengths (internode lengths) approximately corresponding to the number of substitutions between the nodes. Given this, the MPL method produces relative age estimates with confidence intervals for all nodes of the tree. With the age of one or several nodes of the tree being known from reference fossils, the relative age estimates induce absolute age estimates and confidence intervals of the nodes of the tree. The MPL method relies on the assumptions that substitutions occur randomly and independently in different sites in the DNA sequence and that the substitution rates are approximately constant in time, i.e., assuming a molecular clock. A method is presented for identification of the nodes in the tree at which significant deviations from the clock assumption occur, such that dating may be done using different rates in different parts of the tree. The MPL method is illustrated with the Liliales, a group of monocot flowering plants. PMID- 12128029 TI - Molecular phylogenies support multiple morphological reductions in the liverwort subclass Marchantiidae (Bryophyta). AB - The phylogeny of one of the putative basal-most group of land plants, the Marchantiidae, is estimated with morphological characters and with sequences of the nuclear (LSU) rDNA gene (first four domains of the 5' end of the 26S rRNA and four subsequent regions) from 34 species and 27 genera. Molecular and morphological data display high degrees of incongruence. The molecular tree topology predominates in the combined analysis. A trend from complex towards simpler morphological traits is apparent from the molecular and combined trees, whereas a trend from simple towards complex traits prevails in the morphological tree. Previously published molecular data corroborate the molecular results. It is suggested that the incongruence stems from the presence of coherent sets of reduction-related morphological traits varying in concert in the morphological data. Marchantiidae is traditionally subdivided into Marchantiales, Sphaerocarpales and Monocleales, with the majority of taxa referred to the first group. The molecular and the combined data both indicate unequivocally that Sphaerocarpales and Monocleales are nested within Marchantiales, and this result is not explicitly refuted by the morphological data. PMID- 12128030 TI - Amino acid vs. nucleotide characters: challenging preconceived notions. AB - The 567-terminal analysis of atpB, rbcL, and 18S rDNA was used as an empirical example to test the use of amino acid vs. nucleotide characters for protein coding genes at deeper taxonomic levels. Nucleotides for atpB and rbcL had 6.5 times the amount of possible synapomorphy as amino acids. Based on parsimony analyses with unordered character states, nucleotides outperformed amino acids for all three measures of phylogenetic signal used (resolution, branch support, and congruence with independent evidence). The nucleotide tree was much more resolved than the amino acid tree, for both large and small clades. Nearly twice the percentage of well-supported clades resolved in the 18S rDNA tree were resolved using nucleotides (91.8%) relative to amino acids (49.2%). The well supported clades resolved by both character types were much better supported by nucleotides (98.7% vs. 83.8% average jackknife support). The faster evolving nucleotides with a smaller average character-state space outperformed the slower evolving amino acids with a larger average character-state space. Nucleotides outperformed amino acids even with 90% of the terminals deleted. The lack of resolution on the amino acid trees appears to be caused by a lack of congruence among the amino acids, not a lack of replacement substitutions. PMID- 12128031 TI - Molecular support for Afrotheria and the polyphyly of Lipotyphla based on analyses of the growth hormone receptor gene. AB - The order Lipotyphla has generally been viewed as a difficult group to classify. For example, recent morphologically based analyses only weakly support the lipotyphla while molecular evidence renders it polyphyletic, placing the golden moles and tenrecs in the superorder known as Afrotheria. Afrotheria is an hypothesized order that contains elephants, sirenians, hyraxes, aardvarks, elephant shrews, tenrecs, and golden moles. Within this group, it has been suggested that the African lipotyphlans (tenrecs and golden moles) form a monophyletic order sometimes referred to as "Afroscoricida," but more appropriately termed Tenrecoidea. The paper presents a molecular analysis of 36 taxa including representatives of five of the six families in Lipotyphla (Solenodontidae is absent) and all orders within Afrotheria. Parsimony analyses were completed using data from the nucleotide sequence of the tenth exon of the growth hormone receptor gene (GHR). These analyses support both the polyphyly of Lipotyphla and the monophyly of Afrotheria with high bootstrap and jackknife support. In addition, the remaining lipotyphlans (known as Eulipotyphla) appear polyphyletic, as does Tenrecoidea. PMID- 12128032 TI - Detection of recombination among Salmonella enterica strains using the incongruence length difference test. AB - Particular serovars of Salmonella enterica have emerged as significant foodborne pathogens in humans. At the chromosomal level, discrete regions in the Salmonella genome have been identified that are known to play important roles in the maintenance, survival, and virulence of S. enterica within the host. Interestingly, several of these loci appear to have been acquired by horizontal transfer of DNA among and between bacterial species. The profound importance of recombination in pathogen emergence is just now being realized, perhaps explaining the sudden interest in developing novel and facile ways for detecting putative horizontal transfer events in bacteria. The incongruence length difference (ILD) test offers one such means. ILD uses phylogeny to trace sequences that may have been acquired promiscuously by exchange of DNA during chromosome evolution. We show here that the ILD test readily detects recombinations that have taken place in several housekeeping genes in Salmonella as well as genes composing the type 1 pilin complex (14 min) and the inv-spa invasion gene complex (63 min). Moreover, the ILD test indicated that the mutS gene (64 min), whose product helps protect the bacterial genome from invasion by foreign DNA, appears to have undergone intragenic recombination within S. enterica subspecies I. ILD findings were supported using additional tests known to be independent of the ILD approach (e.g., split decomposition analysis and compatibility of sites). Taken together, these data affirm the application of the ILD test as one approach for identifying recombined sequences in the Salmonella chromosome. Furthermore, horizontally acquired sequences within mutS support a model whereby evolutionarily important recombinants of S. enterica are rescued from strains carrying defective mutS alleles via horizontal transfer. PMID- 12128033 TI - Molecular systematics, historical ecology, and phylogeography of Halimeda (Bryopsidales). AB - Halimeda (Bryopsidales), a genus of calcified, segmented green seaweeds, abounds in reefs and lagoons throughout the tropics. To investigate phylogenetic, phylogeographic, and historic ecological relationships of the genus, the nuclear rDNA including the SSU and both ITS regions were sequenced. A maximum likelihood tree revealed the following: (1) there were anatomical and morphological synapomorphies for five well-supported lineages; (2) the last common ancestor of one lineage invaded sandy substrata; those of two other lineages established in wave-affected habitats, whereas the cenancestor of the remaining two lineages occupied sheltered rocky slopes. Yet, several species adapted to new habitats subsequently, resulting in several cases of convergence; (3) all lineages separated into Atlantic and Indo-Pacific daughters, likely resulting from the rise of the Panamanian Isthmus. Each daughter pair gave rise to additional convergent species in similar habitats in different oceans; (4) Halimeda opuntia, the only monophyletic pantropical species detected so far, dispersed from the Indo-Pacific into the Atlantic well after the closure event; (5) minor SSU sequence differences across species and phylogeographic patterns of vicariance indicated a relatively recent diversification of the extant diversity. Cretaceous and Early Tertiary fossil look-alikes of modern species must then have resulted from iterative convergence. PMID- 12128034 TI - Phylogeny of Antarctic dragonfishes (Bathydraconidae, Notothenioidei, Teleostei) and related families based on their anatomy and two mitochondrial genes. AB - Although Antarctic teleosts of the suborder Notothenioidei are well studied, the status of some families remains unclear because of limited taxonomic sampling and sometimes poor statistical support from molecular phylogenies. It is true for the Bathydraconidae, the sister-family of the famous haemoglobin-less icefishes, the Channichthyidae. The present study is aimed at clarifying bathydraconid phylogeny and the interrelationships of higher notothenioid families, taking nototheniids as the outgroup. For this purpose, about 300 positions in the mitochondrial control region, 750 positions in the cytochrome b, and a matrix of morphological characters were employed for separate and simultaneous phylogenetic analyses. We conclude that (1) molecular data strongly support the split of bathydraconids into three clades, here called the Bathydraconinae (Bathydraco, Prionodraco, Racovitzia), the Gymnodraconinae (Gymnodraco, Psilodraco, Acanthodraco), and the Cygnodraconinae (Cygnodraco, Gerlachea, Parachaenichthys). Interrelationships between these three and the Channichthyidae remain unclear. Molecular data support neither paraphyly nor monophyly of the bathydraconids, while morphology leads to the monophyly of the family based on the synapomorphic loss of the spinous dorsal fin; (2) The Channichthyidae, the Harpagiferidae, and the Artedidraconidae are monophyletic families; (3) the phylogeny of the haemoglobin less channichthyids is completely resolved and congruent with the conclusions of based on anatomical characters; (4) The present molecular results as well as other molecular studies favour the hypothesis that harpagiferids are the sister group of artedidraconids, though our morphological matrix puts harpagiferids as the sister-group of all other families on the basis of a single character. With regard to harpagiferid relationships, it is interesting to notice that, when analysed simultaneously, morphological characters are not automatically "swamped" within molecular ones: in the tree based on the simultaneous analysis of all available data, morphological characters impose their topology on molecules. PMID- 12128035 TI - Historical relationships among Neotropical lowland forest areas of endemism as determined by mitochondrial DNA sequence variation within the Wedge-billed Woodcreeper (Aves: Dendrocolaptidae: Glyphorynchus spirurus). AB - Studies of the distribution of South American taxa have identified several areas of endemism that may have contributed to the historical diversification of the region. We constructed a phylogeny of Glyphorynchus spirurus (Aves: Dendrocolaptidae) populations using mtDNA sequence data from portions of cytochrome b, NADH dehydrogenase subunit II (ND2), and complete NADH dehydrogenase subunit III (ND3). Using this phylogeny we evaluate five previous hypotheses of area-relationships, two based on phylogenetic studies of morphological characters in birds and three based on parsimony analysis of endemism in birds and primates. Maximum likelihood and maximum parsimony analyses recovered two phylogenetic hypotheses that differed in the placement of one of the areas. Within each of the areas of endemism, the two analyses support the same clades. Neither of the phylogenetic hypotheses for Glyphorynchus exactly matches any of the five previous hypotheses of area-relationships, although ambiguous support exists for one of them. Five areas-Central America, Inambari, Napo, Para, and Rondonia-are supported as composites with component taxa having phylogenetic affinities with more than one area. Data reported here also indicate high levels of sequence divergence within Glyphorynchus. Genetic breaks within Glyphorynchus are only partially congruent with subspecific taxonomy. The regional sampling design used makes this study the largest scale genetic assay of a widespread Neotropical avian taxon published to date. PMID- 12128037 TI - Quality of life in young people: ratings and factor structure of the Quality of Life Profile-Adolescent Version. AB - The purpose of the study was to provide British data relating to the 54-item Quality of Life Profile Adolescent Version (QOLPAV; Raphael et al., 1996, Journal of Adolescent Health, 19, 366-375) and to explore the underlying factor structure of the questionnaire. The relationship between demographic variables (such as social economic status, gender and ethnicity) and quality of life (QOL) were investigated. Eight hundred and ninety-nine young people aged 12-16 were recruited from secondary schools in the south of England to participate in the study. Factor analysis highlighted eight dimensions embedded within the scale, which were largely consistent with the QOL model proposed by Raphael et al. (1996, Journal of Adolescent Health, 19, 366-375). A notable exception concerned items from the subdomain of "spiritual being", which did not cluster together but dispersed across multiple factors. Exclusion of complex items and those with low loadings on subdomains resulted in a briefer scale consisting of 32 items. Of the demographic variables examined, only age was significantly associated with quality of life scores. PMID- 12128038 TI - Adolescent quality of life, part I: conceptual and measurement model. AB - Few quality of life instruments exist that focus on the positive aspects of adolescence, incorporate adolescents' perspectives and language, and apply to both general and vulnerable populations. With these goals in mind, a conceptual and measurement model was developed using inductive qualitative methods to guide construction of the Youth Quality of Life Instrument-Research Version (YQOL-R). A conceptual model with four domains-Sense of Self, Social Relationships, Environment, and General Quality of Life-is reported. PMID- 12128039 TI - Adolescent quality of life, part II: initial validation of a new instrument. AB - The psychometric properties of the Youth Quality of Life Instrument-Research Version (YQOL-R) perceptual module are presented. Self-reports were obtained from 236 youth aged 12-18 years with mobility disability, attention-deficit hyperactivity disorder, or without self-reported chronic conditions. Item and factor analyses confirmed the hypothesized conceptual model derived from previous qualitative research. The scales of the YQOL-R showed acceptable internal consistency (Cronbach's alpha=0.77-0.96), reproducibility (ICCs=0.74-0.85), expected associations with other constructs, and ability to distinguish between known groups. The YQOL-R showed sufficient validity to encourage further use. PMID- 12128040 TI - Perceived social inadequacy and depressed mood in adolescents. AB - The goal of the study was to examine stability in, and the relationship between, perceived social inadequacy and depressed mood in a sample of community adolescents. The Checklist of Adolescent Problem Situations (CAPS) and Children's Depression Inventory (CDI) were administered to 224 high-school students on two occasions 4 months apart. CDI and CAPS scores were positively correlated and test retest reliability was high in both instruments. Cross-lagged panel correlations, controlled for within-time associations, did not significantly differ. Results indicated that perceived social inadequacy in adolescents is stable over time independent of its association with depressed mood. PMID- 12128041 TI - Locus of control, television viewing, and eating disorder symptomatology in young females. AB - The purpose of this study was to assess the effects of locus of control and television watching (number of hours of television watched per week) on eating disorder symptomatology in girls between the ages 10-17 years. A 2 x 2 factorial design was employed in which girls were identified either as (a) being higher or lower viewers of television, and (b) having either an external or internal locus of control. Girls with an external locus of control had significantly greater eating disorder symptomatology than those with an internal locus of control. For girls watching higher amounts of television, those having an external locus of control had significantly greater eating disorder symptomatology than those having an internal locus of control. PMID- 12128042 TI - A possible selves intervention to enhance school involvement. AB - We developed a 9-week after-school, small group, activities-based intervention focused on enhancing youth's abilities to imagine themselves as successful adults and connecting these future imagines to current school involvement. We describe and evaluate this programme comparing three cohorts of urban African American middle school students (n=62 experimental, n=146 control), controlling for sex and previous school involvement. By the end of the school year, intervention youth reported more bonding to school, concern about doing well in school, "balanced" possible selves, plausible strategies to attain these possible selves, better school attendance, and for boys, less trouble at school. PMID- 12128043 TI - Personality, peer relations, and self-confidence as predictors of happiness and loneliness. AB - This study is set out to examine to what extent peer relations, self-confidence, and school performance correlated with self-rated happiness (OHI) and loneliness (UCLA LS) in adolescents. Personality traits (EPQ), self-confidence (PEI), friendship and school grades were all significantly oppositely correlated with happiness and loneliness. Regressional analysis revealed that extraversion and neuroticism were direct predictors of happiness and self-confidence, while psychoticism and extraversion were direct predictors of loneliness. The effect of sex on happiness and loneliness was moderated by friendship and neuroticism, and by neuroticism and psychoticism, respectively. Extraversion was also a significant predictor of general confidence and social interactions which directly influenced loneliness whilst psychoticism was a direct predictor of loneliness. Self-rated school performance was the only direct predictor of happiness whereas general confidence and social interactions were related to adolescents' self-reported loneliness. PMID- 12128044 TI - High-schoolers' peer orientation priorities: a snapshot. AB - This short paper reports a set of qualitative research findings which reveal something of the peer orientation priorities of 462 British high-schoolers (14 year olds)-who they hang out with and why. Offering a snapshot of young people's current social and relational priorities, these data point to their over-riding concern for mutuality and "hanging" itself (e.g. "having a laugh"); boys' single minded preference for sport (and computers); girls' consistent attunement to pleasant, caring interactions; and the relative unimportance of ethnically based affiliation. PMID- 12128047 TI - Taenia crassiceps: androgen reconstitution of the host leads to protection during cysticercosis. AB - The effects of testosterone, dihydrotestosterone, and 17beta-estradiol in castrated mice of both sexes infected with Taenia crassiceps cysticerci were studied. The results showed that castration and treatment with either testosterone or dihydrotestosterone before infection decreased parasite loads by 50 and 70%, respectively, while the treatment with 17beta-estradiol increased it by three times in both genders, as compared with control mice. The specific splenocyte cell proliferation and IL-2 and IFN-gamma production were depressed in infected-castrated mice of both genders, while treatment with testosterone or dihydrotestosterone produced a significant proliferation recovery and enhanced production of IL-2 and IFN-gamma. On the other hand, the humoral response was unaffected with testosterone or dihydrotestosterone restitution, while the treatment with estradiol in both genders augmented the levels of anti-cysticerci IgG, as well as IL-6 and IL-10 production. These results suggest a protective role for androgens, possibly through the stimulation of the specific cellular immunity. PMID- 12128049 TI - Identification and distribution of carbohydrate moieties on the salivary glands of Rhodnius prolixus and their possible involvement in attachment/invasion by Trypanosoma rangeli. AB - In the present study, FITC-labelled lectins (WGA, Con A, PNA, HPA, and TPA) were utilized to investigate carbohydrate residues on the surface of Rhodnius prolixus salivary glands. The results revealed that the salivary glands are rich in carbohydrate moieties and the diversity in binding pattern of particular lectins showed the presence of specific carbohydrate residues in the basal lamina, muscle, and cell layers of the glands. Subsequently, the sugars detected on the salivary gland surface were employed to investigate the interaction between Trypanosoma rangeli and the R. prolixus salivary glands. In vitro adhesion inhibition assays using long epimastigote forms (the invasion/adhesion forms) showed that some sugars tested were able to block the receptors on both the surfaces of the salivary glands and on T. rangeli. Among the sugars tested, GlcNAc, GalNAc, and galactose showed the highest overall inhibitory effect, following pre-incubation of either the salivary glands or parasites. These results are discussed in relation to previous work on the role of carbohydrates and lectins in insect vector/parasite interactions. PMID- 12128048 TI - Trypanosoma cruzi: identification of a galactose-binding protein that binds to cell surface of human erythrocytes and is involved in cell invasion by the parasite. AB - Trypanosoma cruzi must invade mammalian host cells to replicate and complete its life cycle. Almost all nucleated mammalian cells can be invaded by the parasite following a receptor-ligand recognition as an early prerequisite. In this work, we describe a 67-kDa lectin-like glycoprotein that binds to desialylated human erythrocyte membranes in a galactose-dependent way. This protein is present on the parasite surface in both infective and non-infective stages of T. cruzi. More interestingly, we demonstrate by lectin-immuno-histochemistry assays that the 67kDa protein is involved in the recognition of host-cell receptors in mouse cardiac tissue and human cardiac aortic endothelium and mammary artery tissue. Moreover, antibodies against the 67kDa glycoprotein inhibit in vitro host-cell invasion by 63%. These data suggest that the 67kDa glycoprotein in vivo is needed for host-cell invasion by T. cruzi. PMID- 12128050 TI - Brugian infections in the peritoneal cavities of laboratory mice: kinetics of infection and cellular responses. AB - Standard, immunocompetent, inbred strains of mice are non-permissive for infection with the human filarial nematode, Brugia malayi or the closely related Brugia pahangi. This non-permissiveness allows one to address the mechanism(s) that might be used by mammalian hosts to eliminate large, multicellular, metazoan, extracellular invertebrate pathogens. We describe here the time course of intraperitoneal Brugian infections in naive and primed +/+ mice from two commonly used, inbred laboratory strains (C57BL/6J and BALB/cByJ). We believe that this documentation of the course of infection in normal mice will serve as a reference for future studies using mice with gene-targeted immunological deficits or which have been pharmacologically or immunologically manipulated to manifest such deficits. Our data show that even though both strains of mice eliminate the parasite before the onset of patency, there are significant differences in the time course of infection and in the fractions of input larvae that can be recovered at any time after infection. In a secondary infection, the time course of elimination is accelerated. We examined the cells in the peritoneal cavity, the site of infection, by flow microfluorimetry using forward and side scatter properties and cell surface antigen expression using fluorescent antibodies. These studies reveal a complex cellular pattern, predominated by B lymphocytes, macrophages, and eosinophils. The most notable gross morphological findings at necropsy during the phase of elimination of the parasite are nodules of tissue containing larvae, which appear viable in some cases and undergoing various stages of disintegration in others. These nodules, which are histologically granulomas, are primarily composed of macrophages and eosinophils, with few if any lymphocytes. Transmission electron micrographs reveal that eosinophils can penetrate under the cuticles of the larvae and be seen in close approximation with internal structures. These granulomas may represent an important mechanism by which worms are eliminated. PMID- 12128051 TI - Trypanosoma cruzi: methoprene is a potent agent to sterilize blood infected with trypomastigotes. AB - The effects of methoprene, a juvenile hormone analogue (JHA), on Trypanosoma cruzi bloodstream trypomastigotes (Tulahuen strain, Tul 2 stock) were studied. It was observed that 150microM of methoprene in in vitro experiments cause cellular death of T. cruzi. In contrast, methoprene was not able to clear bloodstream trypomastigotes in in vivo experiments, but it was observed a decrease of parasitemia levels of infected mice treated with 200microg of methoprene/mouse/day during 5 days. According to these results and the low toxicity of methoprene, we suggest that this compound will serve as an effective agent to sterilize blood for transfusions. PMID- 12128052 TI - A non-radiolabelled ferriprotoporphyrin IX biomineralisation inhibition test for the high throughput screening of antimalarial compounds. AB - Intraerythrocytic malaria parasites produce large amounts of toxic ferriprotoporphyrin IX (FP) during their digestion of host cell haemoglobin. The inhibition of biomineralisation of FP to haemozoin (or beta-haematin) by antimalarial drugs underlies their mode of action. We have developed an in vitro microassay for testing the inhibition of biomineralisation by drugs. It is based on the detection by optical density measurement of solubilised beta-haematin remaining after contact with drugs. The assay uses a 192-microM haemin chloride solution in dimethyl sulfoxide, 96-well filtration microplates as well as normal microplates; it lasts 18-24h and requires a spectrophotometer. We determined by this assay the IC(50) of chloroquine phosphate (28microM) and quinine base (324microM) and showed that unlike previous methods it is insensitive to inorganic anions. We also determined the activity of synthetic dyes and plant extract to determinate the interference of coloured compounds on the accuracy of the test. We found that methylene blue, thionine (IC(50) 38 and 87microM, respectively), and an extract of plants that contains quinoline derivatives, inhibited the biomineralisation of FP regardless of their intrinsic colour. PMID- 12128053 TI - Trypanosoma simiae and Trypanosoma congolense: surface glycoconjugates of procyclic forms-the same coats on different hangers? AB - Organic solvent extraction, reverse-phase high performance liquid chromatography and enzyme-linked immunosorbent assay with surface binding monoclonal antibodies were used to isolate membrane molecules of procyclic culture forms of Trypanosoma simiae and Trypanosoma congolense. Gel electrophoresis of the purified molecules revealed two predominant molecular species from each parasite that were broadly similar yet showed different apparent molecular masses and staining characteristics. The molecules were shown to be glycosylphosphatidylinositol lipid anchored glycoconjugates, rich in carbohydrates. Each moiety displayed surface-disposed carbohydrate epitopes that were recognized on the surface of both species of trypanosomes by monoclonal antibodies specific for procyclic parasites of the subgenus Nannomonas. The epitopes were previously shown to be displayed on the glutamic acid-alanine rich protein of T. congolense yet neither this protein, nor its encoding gene is present in T. simiae. The results indicate that although T. congolense and T. simiae share common carbohydrate surface epitopes, these are displayed on biochemically different molecules. We speculate that the surface disposed carbohydrate structures are involved in parasite-tsetse interactions since these species have the same developmental cycles in the insect vector. PMID- 12128054 TI - Trypanosoma cruzi: role of host genetic background in the differential tissue distribution of parasite clonal populations. AB - Chagas' disease, caused by the protozoan parasite Trypanosoma cruzi, has quite a variable clinical presentation, ranging from asymptomatic to severe chronic cardiac and/or gastrointestinal disease. The reason for that is not completely understood, but both parasite and host genetic traits are certainly involved. Recently, we have demonstrated clinically and experimentally that the genetic variability of T. cruzi is one of the determinants of the pattern of tissue involvement in Chagas' disease. We then decided to turn our attention to the role of host genetic background. To study this, we compared the infection of four lineages of mice [three inbred (BALB/c, DBA-2, and c57Black/6) and one outbred (Swiss)] with two T. cruzi clonal populations, the Col1.7G2 clone and the JG monoclonal strain. The tissue distribution of T. cruzi strains was identical for BALB/c and DBA-2 mice, but very different in C57BL/6 (H-2(b)) and outbred Swiss mice. This result clearly demonstrates the importance of host genetic aspects in the process. Since BALB/c and DBA-2 have the same H-2 haplotype (H-2(d)) and C57BL/6 does not (H-2(b)), it is possible that MHC variability may be involved in influencing the tissue distribution of involvement in experimental Chagas' disease of the mouse. PMID- 12128055 TI - Expression and evolution studies of ets genes in a primitive coelomate, the polychaete annelid, Hediste (Nereis) diversicolor. AB - The Ets family includes numerous proteins with a highly conserved DNA-binding domain of 85 amino acids named the ETS domain. Phylogenetic analyses from ETS domains revealed that this family could be divided into 13 groups, among them are ETS and ERG. The ets genes are present in the Metazoan kingdom and we have previously characterized the Nd ets and Nd erg genes in the polychaete annelid Hediste diversicolor. Here, we isolated a fragment encoding the ETS domain from Nd Ets, by genomic library screening. By Northern blot analysis, we showed that this gene was transcribed as one major mRNA of 2.6 kb and one minor mRNA of 3.2 kb. By in situ hybridization, we observed that Nd ets was expressed in the intestine and oocytes and that Nd erg was expressed in cellular clumps present in the coelomic cavity, in an area of proliferating cells situated between the last metamere and the pygidium. Finally, we showed that Nd erg shared the expression pattern of Nd ets in oocytes. Molecular modeling studies have revealed that the spatial structure of ETS domain of Nd Ets and Nd Erg was conserved, in comparison to the murine Ets-1 and human Fli-1 proteins, respectively. PMID- 12128056 TI - Beta-cyclodextrin facilitates cholesterol efflux from larval Manduca sexta fat body and midgut in vitro. AB - The ability of 2-hydroxypropyl-beta-cyclodextrin (HPbetaCD) and methyl-beta cyclodextrin (MbetaCD) to promote cholesterol efflux from [3H]cholesterol-labeled larval Manduca sexta fat body and midgut was tested. In fat body, both beta cyclodextrins induced a two-phase efflux of cholesterol. The first rapid phase depended on cyclodextrin concentration and was more rapid for MbetaCD than for HPbetaCD. The second, slower, phase was independent of cyclodextrin concentration and type. In midgut, only the concentration-dependent phase was observed; the rate constants are approximately 85% slower than for fat body. In both cases, a low activation energy for transfer was observed, consistent with a collision mechanism where cyclodextrin interacts directly with cholesterol in plasma membrane to affect transfer. In fat body, the second slower phase is suggestive of a second pool of exchangeable cholesterol and most likely represents transfer of cholesterol from internal membranes or different lateral domains of the plasma membrane. The lack of this second phase in midgut suggests that midgut has only a single pool of exchangeable cholesterol. Although the rates are somewhat different, the overall kinetic pattern for cyclodextrin-mediated cholesterol transfer in insect fat body closely resembles that for vertebrate cells, while the single pool behavior of the midgut is not found in vertebrate cells. PMID- 12128057 TI - Hemolymph of reproductives of Solenopsis invicta (Hymenoptera: Formicidae)-amino acids, proteins and sugars. AB - Free amino acid composition and carbohydrate and protein concentration of the hemolymph of the pupal and adult stages of reproductives of Solenopsis invicta are presented. The physico-chemical properties of the hemolymph differ between both sexes of fire ants during development. Male alates (459 mmol/kg) have a higher osmolality than pupae (388 mmol/kg). Osmolality during the pupal stage was 428 mmol/kg, decreasing to 354 mmol/kg soon after de-alation and increasing again to 463 mmol/kg in active queens. The hemolymph of newly eclosed adults was more basic than the pupa and older adults. The pH of the hemolymph ranged from 6.40 to 6.71 for males and 6.47 to 7.01 for females. An increase in the carbohydrate and protein content was found for both sexes after emergence. Newly de-alated females had a decrease in their carbohydrate titers, although no changes were found in the protein concentration. An increase in the total free amino acid concentration was recorded for males after emergence, while changes in females were noted only after de-alation. However, females had a 2.7-fold increase in the free amino acid pool after they de-alated, with proline, glutamine and taurine showing the highest increase. Changes in the organic composition and in the physico-chemical properties of the hemolymph of S. invicta are discussed in regard to the physiological processes involved in the maturation of the reproductives of this insect. PMID- 12128058 TI - Carotenol fatty acid esters: easy substrates for digestive enzymes? AB - To study the specificity of gastric lipases on carotenoid mono- and diesters, an enzymatic assay was applied. Digestions were carried out in phosphate buffer at pH 7.4 and 37 degrees C. As substrates we employed oleoresins from marigold (Tagetes erecta L.; lutein diesters), red paprika (Capsicum annuum L., mainly capsanthin diesters), papaya (Carica papaya L.; beta-cryptoxanthin esters), and loquat (Eriobotrya japonica Lindl.; beta-cryptoxanthin esters) as well as retinyl palmitate. These were reacted with porcine pancreatic lipase, porcine pancreatin, porcine cholesterol esterase, and human pancreatic lipase. As reference enzyme a yeast lipase from Candida rugosa was applied. A high turnover could be observed with porcine pancreatic lipase and porcine cholesterol esterase, indicating cholesterol esterase to be a plausible candidate for generation of free carotenoids in the gut. Human pancreatic lipase accepted only retinyl palmitate as substrate, carotenoid mono- and diesters were not hydrolyzed. The assay permits an approach for calculation of enzymatic activities towards carotenoid esters as substrates for the first time, which is based on the amount of enzyme formulation, present in the assay (U/mg solid). Furthermore, these studies provide deeper insight into carotenoid ester bioaccessibility. PMID- 12128059 TI - Lipid radical generation in polar (Laternula elliptica) and temperate (Mya arenaria) bivalves. AB - Lipid peroxidation in Laternula elliptica was assessed by detecting lipid radicals by electronic paramagnetic resonance. The values were compared with data from the temperate mud clam Mya arenaria. Lipid radical content was higher in the Antarctic bivalve than in the temperate mud clam, even within the range of its habitat temperature. The rate of generation of lipid radicals was affected by the iron content in the samples. The iron content in individual samples of digestive glands in L. elliptica ranged from 3 to 6 nmol g(-1) fresh weight (fwt) and in M. arenaria from 0.6 to 2.7 nmol g(-1) fwt. Arrhenius plots, developed from the rates obtained in the presence of 25 microM iron, showed no significant differences between the activation energy calculated for digestive glands of L. elliptica and M. arenaria. The Fe3+ reduction rate in L. elliptica was higher than in M. arenaria (4.7 +/- 0.9 vs. 1.8 +/- 0.4 nmol mg(-1) protein min(-1), respectively). L. elliptica had a higher content of alpha-tocopherol and beta carotene than M. arenaria. Our data suggest that increased lipid radical content in the membranes of cold-adapted organisms could be related to iron content. PMID- 12128060 TI - Proline biosynthesis genes and their regulation under salinity stress in the euryhaline copepod Tigriopus californicus. AB - Diverse organisms regulate concentrations of intracellular organic osmolytes in response to changes in environmental salinity or desiccation. In marine crustaceans, accumulation of high concentrations of proline is a dominant component of response to hyperosmotic stress. In the euryhaline copepod Tigriopus californicus, synthesis of proline from its metabolic precursor glutamate is tightly regulated by changes in environmental salinity. Here, for the first time in a marine invertebrate, the genes responsible for this pathway have been cloned and characterized. The two proteins display the sequence features of homologous enzymes identified from other eukaryotes. One of the cloned genes, delta1 pyrroline-5-carboxylase reductase (P5CR), is demonstrated to have the reductase enzyme activity when expressed in proline-auxotroph bacteria, while the second, delta1-pyrroline-5-carboxylase synthase (P5CS), does not rescue proline-auxotroph bacteria. In contrast to results from higher plants, neither levels of P5CS nor P5CR mRNAs increase in response to salinity stress in T. californicus. Hence, regulation of proline synthesis during osmotic stress in T. californicus is likely mediated by some form of post-transcriptional regulation of either P5CS or P5CR. Understanding the regulation this pathway may elucidate the mechanisms limiting the salinity ranges of marine taxa. PMID- 12128061 TI - Specific cleavage of beta-amyloid peptides by a metallopeptidase from Xenopus laevis skin secretions. AB - Dactylysin (EC 3.5.24.60) is a metalloendopeptidase first isolated from the skin granular gland secretions of Xenopus laevis. This peptidase hydrolyzes bonds on the amino-terminus of singlets and between doublets of hydrophobic amino acids and was considered to play a role in the in vivo inactivation of biologically active regulatory peptides. Here, we show that dactylysin has also the ability to cleave human beta[1-40]-amyloid peptide and related peptides. Cleavage of the wild type beta[1-40]-amyloid peptide form, and to a lesser extent Flemish and Dutch mutants, occurred predominantly at the His14-Glu15 bond. We demonstrate that frog skin exudate contains a full-length amyloid protein precursor detected by immunochemical cross-reactivity with monoclonal antibody against C-terminal human amyloid protein precursor. The possibility that dactylysin, might be involved in normal catabolism of beta amyloid peptide of Xenopus laevis is discussed. PMID- 12128062 TI - Identification and molecular cloning of Xenopus laevis SP22, a protein associated with fertilization in mammals. AB - SP22 is a novel sperm protein that has been shown to be highly correlated with fertility in rats. SP22 homologues have been studied in mouse and man but a definitive role for the protein has not yet been established. Using a polyclonal IgG to recombinant rat SP22, we detected the presence of this protein in Xenopus laevis tissues. Moreover, a Xenopus EST was found that shares a high degree of similarity with rat SP22 and the derived sequence codes for an 189 aa protein that is very similar to rat SP22. Finally, using Western blotting and RT-PCR analyses, we investigated the expression of Xenopus SP22 both in adult tissues and during embyronic development. SP22 protein expression predominated in the adult testis, and both mRNA and protein levels diminished subsequent to the initial day following fertilization. PMID- 12128063 TI - Expression of a hepatocyte growth-factor activator protein in turkey (Meleagris gallopavo) deferent duct epithelial cells. AB - In previous research, we discovered that turkey deferent duct epithelial cells express a serine protease. Our experimental objective was to identify the gene that encodes this protein. A lambda phage cDNA library from duct cell mRNA was constructed. The library was screened using monoclonal antibodies previously produced against the turkey deferent-duct serine protease. Phage containing the protease cDNA was excised and re-circularized into plasmids. E. coli were transformed with plasmids containing protease cDNA, which was then isolated for sequencing. NCBI BLAST searches within the GenBank database returned 63.5 and 61.7% identity with murine and human hepatocyte growth-factor activator (HGFA) precursor, respectively. The turkey protease cDNA was then cloned into the pQE-32 expression vector and transformed into M15 cells for HIS-tagged expression of the recombinant protein, which was then purified using nickel-chelated Sepharose spin columns. Afterwards, Western blot analysis of the purified recombinant turkey protein revealed recognition by a monoclonal antibody specific to the proteolytic subunit of the turkey deferent duct protease. Therefore, these findings indicate that the recombinant HGFA precursor isolated from the deferent duct is the turkey seminal plasma protease that is secreted from the deferent duct. HGFA, a member of the Kringle-serine proteinase superfamily, can initiate diverse mitogenic, morphogenic and motogenic effects through its substrate hepatocyte growth factor. Although the presence of hepatocyte growth factor and its c-MET receptor have been reported in male mammalian reproductive tracts, our novel findings on the secretion of HGFA precursor from turkeys may help to elucidate the regulation of activated hepatocyte growth factor. PMID- 12128064 TI - Changes of major carotenoids in gonads of sea urchins (Strongylocentrotus intermedius and S. nudus) at maturation. AB - Changes in the carotenoid content in gonads of two sea urchins species were investigated during maturation. The content of echinenones and carotenes, the two major carotenoid fractions in gonads, is highest for Strongylocentrotus intermedius at the spawning gametogenic stage of gonad maturation for both sexes. For S. nudus, the content of these pigments is highest at stages of active gametogenesis and spawning for males and at the growth stage for females. A comparison of the carotenoid content dynamics during maturation of gonads for males, females and animals at the resting (sexual inactivity) stage was also carried out. PMID- 12128065 TI - Chloride conductance and mitochondria-rich cell density in isolated skin of Rana catesbeiana acclimated to various environments. AB - The Cl- conductance in isolated skin of frogs (Rana catesbeiana) acclimated to 30 mM solutions of NaCl, Na2SO4, MgCl2 and distilled water (DW) was studied. Transepithelial potential difference (PDtrans), short-circuit current (ISC) and total conductance (Gt) were measured under conditions such that there was Cl- flux in the presence and absence of Na+ transport. The Cl- content of the mucosal solution was acutely replaced with SO42- or gluconate to evaluate the effect of removal of Cl- conductance on electrophysiological parameters. Mitochondria-rich cell density (DMRC) was also measured. Skins from frogs acclimated to NaCl and Na2SO4 showed the lowest and the highest D(MRC), respectively, but no difference could be found between the skins from frogs acclimated to DW and MgCl2 indicating that DMRC is not unconditionally dependent on environmental Cl- in this species. Frogs acclimated to NaCl showed marked differences when compared to the other groups: the highest Gt, probably represented by a higher paracellular conductance; the lowest transepithelial electrical potential difference which remained invariant after replacement of mucosal Cl- with SO42- or replacement of mucosal Cl- with gluconate and an inwardly oriented positive current in the absence of bilateral Na+. PMID- 12128066 TI - Methacholine-induced cGMP production is regulated by nitric oxide generation in rabbit submandibular gland cells. AB - Guanosine 3',5'-monophosphate (cGMP) is an intracellular messenger in various kinds of cell. We investigated the regulation of cGMP production by nitric oxide (NO) in rabbit submandibular gland cells. Methacholine, a muscarinic cholinergic agonist, stimulated cGMP production in a dose- and time-dependent manner, but the alpha-agonist phenylephrine, substance P and the beta-agonist isoproterenol failed to evoke cGMP production. In fura-2-loaded cells, methacholine induced an increase in intracellular Ca2+ ([Ca2+]i) in a concentration-dependent manner, which was similar to that for cGMP production. When the external Ca2+ was chelated with EGTA, methacholine failed to induce cGMP production. Ca2+ ionophore A23187 and thapsigargin, which induce the increase in [Ca2+]i without activation of Ca2+-mobilizing receptors, mimicked the effect of methacholine. cGMP production induced by methacholine, A23187 and thapsigargin was clearly inhibited by NG-nitro-L-arginine methylester (L-NAME), a specific inhibitor of nitric oxide synthase (NOS). S-Nitroso-N-acetyl-DL-penicillamine (SNAP), a NO donor, induced cGMP formation. In the lysate of rabbit submandibular gland cells, Ca2+-regulated nitric oxide synthase activity was detected. These findings suggest that cGMP production induced by the activation of muscarinic cholinergic receptors is regulated by NO generation via the increase in [Ca2+]i. PMID- 12128067 TI - Anhydrolutein in the zebra finch: a new, metabolically derived carotenoid in birds. AB - Many birds acquire carotenoid pigments from the diet that they deposit into feathers and bare parts to develop extravagant sexual coloration. Although biologists have shown interest in both the mechanisms and function of these colorful displays, the carotenoids ingested and processed by these birds are poorly described. Here we document the carotenoid-pigment profile in the diet, blood and tissue of captive male and female zebra finches (Taeniopygia guttata). Dietary carotenoids including: lutein; zeaxanthin; and beta-cryptoxanthin were also present in the plasma, liver, adipose tissue and egg-yolk. These were accompanied in the blood and tissues by a fourth pigment, 2',3'-anhydrolutein, that was absent from the diet. To our knowledge, this is the first reported documentation of anhydrolutein in any avian species; among animals, it has been previously described only in human skin and serum and in fish liver. We also identified anhydrolutein in the plasma of two closely related estrildid finch species (Estrilda astrild and Sporaeginthus subflavus). Anhydrolutein was the major carotenoid found in zebra finch serum and liver, but did not exceed the concentration of lutein and zeaxanthin in adipose tissue or egg yolk. Whereas the percent composition of zeaxanthin and beta-cryptoxanthin were similar between diet and plasma, lutein was comparatively less abundant in plasma than in the diet. Lutein also was proportionally deficient in plasma from birds that circulated a higher percentage of anhydrolutein. These results suggest that zebra finches metabolically derive anhydrolutein from dietary sources of lutein. The production site and physiological function of anhydrolutein have yet to be determined. PMID- 12128070 TI - Proteoglycan production in disomic and trisomy 7-carrying human synovial cells. AB - To gain further insight into the synthesis and structure of the synovial matrix of joints, we have established cell cultures from synovial specimens and elaborated their production of hyaluronan and proteoglycans. The cultures secreted mainly the small proteoglycan decorin, but also considerable amounts of the related biglycan and the large proteoglycan versican. Only minor amounts of heparan sulfate proteoglycans were found. All cultures also had a high production of hyaluronan, which highlights the important role for normal joint function of these cells. In joint diseases, a common feature is the presence of an extra chromosome 7 (trisomy 7) in the synovial cells. To study the possible consequences of trisomy 7 on the synovial cell function, we extended our study to cultures that had been sub-cloned to contain high amounts of trisomy 7-carrying cells. These cell cultures had approximately four times more versican than their disomic counterparts in the cell culture medium, indicating that versican may be a mediator in the processes of joint destructive disorders. To find an explanation for this increase in versican, we investigated the expression/secretion of PDGF-AA and IL-6, cytokines with their genes located to chromosome 7. Indeed, both these cytokines were increased in the cultures with high frequencies of trisomy 7. We then added the two cytokines to cell cultures of disomic synovial cells, but only cells treated with IL-6 displayed an increased amount of versican. Thus, we suggest that the increased amount of versican in cultures of trisomy 7-carrying cells relates to an autocrine loop involving an increased IL-6 production. PMID- 12128069 TI - A recombinant NH(2)-terminal heparin-binding domain of the adhesive glycoprotein, thrombospondin-1, promotes endothelial tube formation and cell survival: a possible role for syndecan-4 proteoglycan. AB - Thrombospondin-1 (TSP-1) is a multifunctional protein known to modulate angiogenesis, endothelial cell adhesion and apoptosis. In this study, we have demonstrated that TSP18, a recombinant 18 kDa protein encompassing the N-terminal residues 1-174 of human TSP-1, accelerated the process of tube-like structures formation by human umbilical vein endothelial cells (HUVECs) when included in fibrin matrices at 0.55-2.2 microM concentrations, for times ranging from 24 to 72 h. This effect was specifically inhibited by V58A4, a Mab raised against TSP18. Whole TSP-1 showed a dual effect, weakly enhancing tube formation at 22 nM (10 microg/ml), but causing inhibition at 45 and 90 nM (20 and 40 microg/ml, respectively). In order to investigate the possible effects of TSP18 on cell adhesion and viability, we performed adhesion assays on different protein supports. HUVECs adhered more weakly on TSP-1-coated surfaces, remaining round shaped, as compared to the well-spread phenotype displayed on fibronectin and gelatin. Cells adhering on TSP18-coated surfaces displayed a well spread phenotype, with this adhesion strongly inhibited by heparin. The binding of TSP18 to endothelial membrane extracts was blocked by a monoclonal IgG directed against the cell surface proteoglycan syndecan-4. The DNA fragmentation patterns and the nuclear morphology were comparable for HUVECs adhering on all proteins, including TSP18, showing minimal cell apoptosis. Our results indicate that the N-terminal region of TSP-1 constitutes a suitable adhesive support for HUVECs, protecting them from apoptosis, possibly mediated by syndecan-4 proteoglycan. PMID- 12128072 TI - Retention of the native chondrocyte pericellular matrix results in significantly improved matrix production. AB - The interaction of the cell with its surrounding extracellular matrix (ECM) has a major effect on cell metabolism. We have previously shown that chondrons, chondrocytes with their in vivo-formed pericellular matrix, can be enzymatically isolated from articular cartilage. To study the effect of the native chondrocyte pericellular matrix on ECM production and assembly, chondrons were compared with chondrocytes isolated without any pericellular matrix. Immediately after isolation from human cartilage, chondrons and chondrocytes were centrifuged into pellets and cultured. Chondron pellets had a greater increase in weight over 8 weeks, were more hyaline appearing, and had more type II collagen deposition and assembly than chondrocyte pellets. Minimal type I procollagen immunofluorescence was detected for both chondron and chondrocyte pellets. Chondron pellets had a 10 fold increase in proteoglycan content compared with a six-fold increase for chondrocyte pellets over 8 weeks (P<0.0001). There was no significant cell division for either chondron or chondrocyte pellets. The majority of cells within both chondron and chondrocyte pellets maintained their polygonal or rounded shape except for a thin, superficial edging of flattened cells. This edging was similar to a perichondrium with abundant type I collagen and fibronectin, and decreased type II collagen and proteoglycan content compared with the remainder of the pellet. This study demonstrates that the native pericellular matrix promotes matrix production and assembly in vitro. Further, the continued matrix production and assembly throughout the 8-week culture period make chondron pellet cultures valuable as a hyaline-like cartilage model in vitro. PMID- 12128071 TI - Regulation of the expression of lysyl oxidase mRNA in cultured rabbit retinal pigment epithelium cells. AB - Lysyl oxidase, an extracellular amine oxidase, controls the maturation of collagen and elastin. We examined the regulation of lysyl oxidase mRNA in cultured rabbit retinal pigment epithelium (RPE) cells in relation to the changes in subretinal fluid transport and phenotype of RPE cells. The level of the mRNA in cells grown on microporous membranes was markedly increased by application of hyperosmotic mannitol solution on the apical side (191% of control), implying that RPE cells express more lysyl oxidase in the condition which may cause the accumulation of subretinal fluid. Platelet-derived growth factor increased the mRNA level in subconfluent cells in culture (137% of control) and basic fibroblast growth factor decreased it (79% of control). In addition, exposure of cells to retinoic acid alone or in combination with dibutyryl cAMP for 22 days markedly decreased the level of lysyl oxidase mRNA (52 or 35% of control) while increasing the level of mRNA of N-acetylglucosaminidase (NAG), a marker enzyme for lysosomes (162 or 142% of control). Moreover, the level of lysyl oxidase mRNA in cells grown on microporous membranes was lower than that in cells grown on plastic dishes, while the level of NAG mRNA in the former cells was higher than that in the latter. Taken together, the expression of lysyl oxidase seemed to increase during proliferation of RPE cells and decrease toward differentiation. beta-Aminopropionitrile, an inhibitor of lysyl oxidase, significantly inhibited the contraction of collagen gels by fetal calf serum, suggesting that lysyl oxidase may be involved in pathogenesis caused by RPE cells. PMID- 12128073 TI - Lumican is a major proteoglycan component of the bone matrix. AB - MC3T3-E1 mouse calvaria cells are a clonal population of committed osteoprogenitors that in the presence of appropriate supplements form a mineralized bone matrix. The development of the MC3T3-E1 cells can be divided into three major stages, namely, proliferation, differentiation, and mineralization. Recently, using the cDNA microarray technology we found lumican to be abundantly expressed during the mineralization and differentiation stages of the MC3T3-E1 development and not during the proliferation stage. Lumican has been shown to play essential roles in regulating collagen fibril formation in different extracellular matrices but its expression in the developing bone matrix remains elusive. By examining the expression profile of this gene during the different stages of MC3T3-E1 development, utilizing the 'real-time' PCR technology, we observed that the expression of lumican increases as the osteoblast culture differentiates and matures, suggesting that lumican may be involved in regulating collagen fibrillogenesis in bone matrices. Using immunostaining, we observed that during the early embryonic development of mouse (E11 to E13), lumican is mainly expressed in the cartilaginous matrices. However, in the older embryos (E14 to E16), the expression of lumican is more prominent in the developing bone matrices. Our data suggest that lumican is a significant proteoglycan component of bone matrix, which is secreted by differentiating and mature osteoblasts only and therefore it can be used as a marker to distinguish proliferating pre-osteoblasts from the differentiating osteoblasts. PMID- 12128074 TI - Collagen fibril size and crimp morphology in ruptured and intact Achilles tendons. AB - The present study examined the hypothesis that collagen fibril diameter and crimp angle in ruptured human Achilles tendons differed from that of intact ones. Tissue samples were obtained from the central core (distal core) and the posterior periphery (distal superficial) at the rupture site, and the proximally intact (proximal superficial) part of the tendon in 10 subjects (38+/-8 years) with a complete tendon rupture. For comparisons corresponding tissue samples were procured from age (38+/-7 years) and gender matched intact Achilles tendons during routine forensic autopsy. The cross-sectional area density and diameter distribution of fibrils were analyzed using stereological techniques of digitized electron microscopy biopsy cross-sections, while crimp angle was measured by the changing banding pattern of collagen fibers when rotated between crossed polars. Nine of 10 persons with tendon ruptures reported that the injury did not occur during exceedingly large forces, and none experienced any symptoms in the days or months prior to the injury. Fibril diameter distribution showed no region specific differences in either the ruptured or intact tendons for either group. However, in the distal core there were fewer fibrils in the ruptured compared to the intact tendons in 60-150 nm range, P<0.01. Similarly, in the distal superficial portion there were fewer fibrils in the ruptured compared to the intact tendons in the 90-120 nm range, 2P<0.05, while there were no differences in the proximal superficial tendons. Crimp angle did not display any region specific differences, or any difference between the rupture and intact tendons. In conclusion, these data suggest that although crimp morphology is unchanged there appears to be a site-specific loss of larger fibrils in the core and periphery of the Achilles tendon rupture site. Moreover, the lack of symptoms prior to the rupture suggests that clinical tendinopathy is not an etiological factor in complete tendon ruptures. PMID- 12128077 TI - Correlation between beta-amyloid peptide production and human APP-induced neuronal death. AB - The production of amyloid peptide (Abeta) from its precursor (APP) plays a key role in Alzheimer's disease (AD). However, the link between Abeta production and neuronal death remains elusive. We studied the biological effects associated with human APP expression and metabolism in rat cortical neurons. Human APP expressed in neurons is processed to produce Abeta and soluble APP. Moreover, human APP expression triggers neuronal death. Pepstatin A, an inhibitor of aspartyl proteases that reduces Abeta production, protects neurons from APP-induced neurotoxicity. This suggests that Abeta production is likely to be the critical event in the neurodegenerative process of AD. PMID- 12128078 TI - Beta-amyloid (Abeta) protein in cerebrospinal fluid as a biomarker for Alzheimer's disease. AB - With the arrival of symptomatic treatment (acetylcholine esterase inhibitors) and the promise of drugs that may delay disease progression, development of diagnostic biomarkers for Alzheimer's disease (AD) are important. Beta-Amyloid (Abeta) protein is the main component of senile plaques. A marked reduction in cerebrospinal fluid (CSF)-Abeta42 in AD has been found in numerous studies. Importantly, reduced CSF-Abeta42 is also found very early in the disease process, before the onset of clinical symptoms. Recent studies suggest that CSF-Abeta42 have a satisfactory performance when used as a diagnostic marker for AD in clinical routine. This paper reviews CSF-Abeta42 as a biomarker for AD. PMID- 12128079 TI - Electrophysiologic properties of channels induced by Abeta25-35 in planar lipid bilayers. AB - Abeta25-35, a fragment of the neurotoxic amyloid beta protein Abeta1-42 found in the brain of Alzheimer patients, possesses amyloidogenic, neurotoxins and channel forming abilities similar to that of Abeta1-42. We have previously reported that Abeta25-35 formed voltage-dependent, relatively nonselective, ion-permeable channels in planar lipid bilayers. Here, we show that Abeta25-35 formed channels in both solvent-containing and solvent-free bilayers. We also report that for Abeta25-35, channel forming activity was dependent on ionic strength, membrane lipid composition, and peptide concentration, but not on pH. Lower ionic strength and negatively charged lipids increased channel formation activity, while cholesterol decreased activity. The nonlinear function relating [Abeta25-35] and membrane activity suggests that aggregation of at least three monomers is required for channel formation. PMID- 12128080 TI - Laminin affects polymerization, depolymerization and neurotoxicity of Abeta peptide. AB - Amyloid deposition in Alzheimer fibrils forms neurotoxic senile plaques in a process that may be modulated by associated proteins. In this work we demonstrate the ability of laminin-1 and laminin-2 to inhibit fibril formation and toxicity on cultured rat hippocampal neurons. We confirm that the laminin-1-derived peptide YFQRYLI inhibits efficiently both fibril formation and neurotoxicity and show that the IKVAV peptide inhibits amyloid neurotoxicity despite its slight inhibition of fibril formation. On other hand, laminin-1 induces disaggregation of preformed fibrils in vitro, characterized as a progressive disassembly of fibrils into protofibrils and further clearance of these latter species, leading to a continual inhibition of amyloid neurotoxicity. PMID- 12128081 TI - Exogenous induction of cerebral beta-amyloidosis in betaAPP-transgenic mice. AB - A key commonality of most age-related neurodegenerative diseases is the accumulation of aggregation-prone proteins in the brain. Except for the prionoses, the initiation and propagation of these proteopathies in vivo remains poorly understood. In a previous study, we found that the deposition of the amyloidogenic peptide Abeta can be induced by injection of dilute extracts of Alzheimeric neocortex into the brains of Tg2576 transgenic mice overexpressing the human beta-amyloid precursor protein. The present study was undertaken to assess the pathology after long-term (12 months) incubation, and to clarify the distinctive anatomical distribution of seeded Abeta-immunoreactivity. All mice were injected at 3 months of age; 5 months later, as expected, Abeta deposits were concentrated mostly in the injected hemisphere. After 12 months, abundant, transgene-derived Abeta deposits were present bilaterally in the forebrain, but plaque load was still clearly greater in the extract-injected hemisphere. There was also evidence of tau hyperphosphorylation in axons of the corpus callosum that had been injured by the injection, most prominently in transgenic mice, but also, to a lesser degree, in non-transgenic mice. Five months following injection of AD-extract, an isolated cluster of Abeta-immunoreactive microglia was sometimes evident in the ipsilateral entorhinal cortex; the strong innervation of the hippocampus by entorhinal cortical neurons suggests the possible spread of seeded pathology from the injection site via neuronal transport mechanisms. Finally, using India Ink to map the local dispersion of injectate, we found that Abeta induction is especially potent in places where the injectate is sequestered. The AD-seeding model can illuminate the emergence and spread of cerebral beta-amyloidosis and tau hyperphosphorylation, and thus could enhance our understanding of AD and its pathogenic commonalties with other cerebral proteopathies. PMID- 12128082 TI - Annexin 5 and apolipoprotein E2 protect against Alzheimer's amyloid-beta-peptide cytotoxicity by competitive inhibition at a common phosphatidylserine interaction site. AB - Amyloid-beta-protein (betaA/4, AbetaP) accumulates in the brains of patients with Alzheimer's disease (AD), regardless of genetic etiology, and is thought to be the toxic principle responsible for neuronal cell death. The varepsilon4 allele of apoE has been linked closely to earlier onset of AD and increased deposition of the amyloid peptide, regardless of the clinical status of AD, while the apoE varepsilon2 allele is generally protective. We have previously hypothesized that the cell target for amyloid peptide might be the apoptotic signal molecule phosphatidylserine (PS). We report here that annexin 5, a specific ligand for PS, not only blocks amyloid peptide AbetaP[1-40] cytotoxicity, but competitively inhibits AbetaP[1-40]-dependent aggregation of PS liposomes. In addition, we find that apoE2, but not apoE4, can not only perform the same protective effect on cells exposed to AbetaP[1-40], but can also competitively inhibit PS liposome aggregation and fusion by the amyloid peptide. Altogether, the in vivo and in vitro results reported here provide fundamental insight to the process by which amyloid targets specific neurons for destruction, and suggest that PS may be a surface "receptor" site for AbetaP binding. These results also provide a biochemical mechanism by which the apoE varepsilon2 allele, but not apoE varepsilon4, can be protective towards the incidence and progression of Alzheimer's disease. PMID- 12128083 TI - Imaging real-time aggregation of amyloid beta protein (1-42) by atomic force microscopy. AB - Amyloid beta protein (AbetaP) is the major fibrillar constituent of senile plaques. However, no causative role for AbetaP-fibers in Alzheimer's disease (AD) pathology is established. Globular AbetaPs are continuously released during normal cellular metabolism at pico- to nano-molar concentration. We used atomic force microscopy (AFM) to examine aggregation of freshly prepared AbetaP(1-42) and to examine the role of AbetaP concentration, imaging medium (air, water, or PBS) and agonists/antagonists on AbetaP-fibrillogenesis. At even very high and non-physiological AbetaP concentrations, 24-48 h of real-time AFM imaging (a) in water show only multiple layers of globular aggregates and no fibrils and (b) in PBS show mainly the globular structures and some short fibrils. On-line addition of Zn, an agonist for AbetaP-fibrillogenesis, induced a slow but non-fibrillar aggregation of globular AbetaPs. EDTA, a chelator of Zn and calcium (a modulator of AbetaP-mediated toxicity) induced a reversible change in the Zn-mediated aggregation. These results strongly suggest that no AbetaP-fibers are formed for the physiologically relevant concentration and thus the plaque-associated fibers may not account for the AD pathophysiology. PMID- 12128084 TI - Amyloid beta-peptide induces cholinergic dysfunction and cognitive deficits: a minireview. AB - Amyloid beta-peptide (Abeta) plays a critical role in the development of Alzheimer's disease (AD). Much progress has been made in understanding this age related neurodegenerative disorder, thus an insight into the cellular actions of Abeta and resulting functional consequences may contribute to preventive and therapeutic approaches for AD. In this review, recent evidence of Abeta-induced brain dysfunction, particularly of cholinergic impairment and memory deficits is summarized. Moreover, proposed mechanisms for Abeta-induced neurotoxicity such as oxidative stress, ion-channel formation, and Abeta-receptor interaction are discussed. PMID- 12128085 TI - Biogenesis and metabolism of Alzheimer's disease Abeta amyloid peptides. AB - Biochemical and genetic evidence indicates the balance of biogenesis/clearance of Abeta amyloid peptides is altered in Alzheimer's disease. Abeta is derived, by two sequential cleavages, from the receptor-like amyloid precursor protein (APP). The proteases involved are beta-secretase, identified as the novel aspartyl protease BACE, and gamma-secretase, a multimeric complex containing the presenilins (PS). Gamma-secretase can release either Abeta40 or the more aggregating and cytotoxic Abeta42. Secreted Abeta peptides become either degraded by the metalloproteases insulin-degrading enzyme (IDE) and neprilysin or metabolized through receptor uptake mediated by apolipoprotein E. Therapeutic approaches based on secretase inhibition or amyloid clearance are currently under development. PMID- 12128086 TI - Methionine residue 35 is critical for the oxidative stress and neurotoxic properties of Alzheimer's amyloid beta-peptide 1-42. AB - Amyloid beta-peptide 1-42 [Abeta(1-42)] is central to the pathogenesis of Alzheimer's disease (AD), and the AD brain is under intense oxidative stress. Our laboratory combined these two aspects of AD into the Abeta-associated free radical oxidative stress model for neurodegeneration in AD brain. Abeta(1-42) caused protein oxidation, lipid peroxidation, reactive oxygen species formation, and cell death in neuronal and synaptosomal systems, all of which could be inhibited by free radical antioxidants. Recent studies have been directed at discerning molecular mechanisms by which Abeta(1-42)-associated free radical oxidative stress and neurotoxicity arise. The single methionine located in residue 35 of Abeta(1-42) is critical for these properties. This review presents the evidence supporting the role of methionine in Abeta(1-42)-associated free radical oxidative stress and neurotoxicity. This work is of obvious relevance to AD and provides a coupling between the centrality of Abeta(1-42) in the pathogenesis of AD and the oxidative stress under which the AD brain exists. PMID- 12128087 TI - The channel hypothesis of Alzheimer's disease: current status. AB - The channel hypothesis of Alzheimer's disease (AD) proposes that the beta-amyloid (Abeta) peptides which accumulate in plaques in the brain actually damage and/or kill neurons by forming ion channels. Evidence from a number of laboratories has demonstrated that Abeta peptides can form ion channels in lipid bilayers, liposomes, neurons, oocyctes, and endothelial cells. These channels possess distinct physiologic characteristics that would be consistent with their toxic properties. Abeta channels are heterogeneous in size, selectivity, blockade, and gating. They are generally large, voltage-independent, and relatively poorly selective amongst physiologic ions, admitting calcium ion (Ca(2+)), Na(+), K(+), Cs(+), Li(+), and possibly Cl(-). They are reversibly blocked by zinc ion (Zn(2+)), and tromethamine (tris), and irreversibly by aluminum ion (Al(3+)). Congo red inhibits channel formation, but does not block inserted channels. Although much evidence implicates Abeta peptides in the neurotoxicity of AD, no other toxic mechanism has been demonstrated to be the underlying etiology of AD. Channel formation by several other amyloid peptides lends credence to the notion that this is a critical mechanism of cytotoxicity. PMID- 12128088 TI - Is gamma-secretase a multienzyme complex for membrane protein degradation? Models and speculations. AB - The beta- and gamma-secretases cleave the amyloid protein precursor (APP) to release the amyloid protein (Abeta). While the beta-secretase has now been identified, the gamma-secretase remains an enigma. A number of mutations in the presenilins (PS) and APP have been shown to alter the cleavage specificity of gamma-secretase. However, the relationship between PS and gamma-secretase remains unclear. This article presents some models of gamma-secretase and suggests that the simplest interpretation of current data is that gamma-secretase is a complex of several proteases located in the lumen of secretory vesicles. PMID- 12128089 TI - Glycosaminoglycans and beta-amyloid, prion and tau peptides in neurodegenerative diseases. AB - Protein aggregation into dense filamentous inclusions is a characteristic feature of many etiologically diverse neurodegenerative disorders including Alzheimer's disease (AD), spongiform encephalopathies, and tauopathies. Thus, beta-amyloid peptide (Abeta) accumulates within senile amyloid plaques in AD, protease resistant prion protein constitutes the amyloid deposits in spongiform encephalopathies and tau protein gives rise to neurofibrillary tangles (NFT) both in AD and in tauopathies. Curiously, these abnormal protein inclusions contain, in addition to their major peptide components, some associated sulfated glycosaminoglycans (sGAG). Here we discuss the proposal that the binding of sGAG to aggregate-forming peptides may modify the pathogenic process depending on their subcellular localization. PMID- 12128090 TI - The state versus amyloid-beta: the trial of the most wanted criminal in Alzheimer disease. AB - Investigators studying the primary culprit responsible for Alzheimer disease have, for the past two decades, primarily focused on amyloid-beta (Abeta). Here, we put Abeta on trial and review evidence amassed by the prosecution that implicate Abeta and also consider arguments and evidence gathered by the defense team who are convinced of the innocence of their client. As in all trials, the arguments provided by the prosecution and defense revolve around the same evidence, with opposing interpretations. Below, we present a brief synopsis of the trial for you, the jury, to decide the verdict. Amyloid-beta: guilty or not guilty? PMID- 12128091 TI - Senile plaque composition and posttranslational modification of amyloid-beta peptide and associated proteins. AB - Amyloid deposits are primarily composed of the amyloid-beta protein, although other proteins (and metal ions) also have been colocalized to these lesions. The pattern of oxidative modifications in amyloid plaques is very different to that associated with neurofibrillary tangles and neuronal cell bodies, likely reflecting the different composition of these structures, accessibility of oxidants, the generation of oxidants in and around these structures and the intrinsic antioxidant defense systems to protect these structures. Future studies directed at understanding Abeta interactions with other amyloid components, the role of oxidative modifications in stabilizing amyloid deposits and the determination of protease cleavage sites on Abeta may provide mechanistic insights regarding both amyloid formation and removal. PMID- 12128092 TI - Transmissible spongiform encephalopathies: the story of a pathogenic protein. AB - An overview is provided from the first description of the transmissible spongiform encephalopathies (TSE) to recent major discoveries in this research field. The TSE are a group of diseases in animal and in man caused by a unique pathogen: the prion protein. The exact nature of the etiological agent or the prion protein is thought to be a misfolded protein. Although current research has provided a wealth of data indicating that a structural isoform of the prion protein is the responsible pathogen, this hypothesis is not yet experimentally proven. PMID- 12128093 TI - The relation between daily activities and scrotal temperature. AB - Normal sperm production depends on a testicular temperature below body temperature, but the thermogenic effects of daily life activities are not well known. We described the association between scrotal temperature and daily activities in 101 males using a non-invasive method for measuring scrotal temperature. A thermistor was attached to the underwear and the temperature of the scrotal skin was logged by a portable device every 5 min for 24h. Participants reported sedentary position and activities at work and during spare time in a questionnaire. Scrotal temperature was strongly correlated with sedentary work position with a dose-effect association (<1 h sedentary: 33.1 degrees C, >6 h sedentary: 34.7 degrees C, median values). The variation in sedentary work accounted for 31.5% of the variation in median temperature during the entire 24h. Sedentary position during spare time did not correlate with scrotal temperature. Median temperature at night was 1.2 degrees C higher than during the daytime. No effect was found for size or reported tightness of the underwear. In a model experiment, the deviance between testicular and scrotal temperature was estimated as maximally 0.1-0.6 degrees C, depending on the type of activity. Measuring scrotal temperature provides a valid estimate of testicular temperature and is feasible in large cohorts. We conclude that work position is an important determinant of testicular temperature. PMID- 12128094 TI - Impact of diurnal scrotal temperature on semen quality. AB - A high scrotal temperature is a common finding in infertile patients and experimental studies indicate that specific types of heat exposure reduce semen quality. More and more men have a sedentary work position, which increases scrotal temperature. Semen and blood samples from 99 healthy men were analysed in relation to scrotal skin temperature obtained by a 24-h continuous monitoring protocol. Information on sedentary position at work and during spare time was collected by questionnaires. A negative correlation was found between high scrotal temperature and sperm output. Sperm concentration decreased 40% per 1 degrees C increment of median daytime scrotal temperature (95% CI: 8-71%). Similar results were found for total sperm count, FSH, and inhibin B. Motility, morphology, pH, and testosterone were not significantly associated with temperature. Only weak and inconsistent associations were found between sedentary position and semen quality. We conclude that scrotal temperature and semen quality are closely associated. Sedentary work position encountered in ordinary jobs, although a strong determinant of scrotal temperature, does not seem to have any effect on semen quality. PMID- 12128095 TI - Microarray analysis of altered gene expression in the TM4 Sertoli-like cell line exposed to chromium(III) chloride. AB - Chromium(III) chloride is a common human exposure metal that is a preconceptional carcinogen in mice, although it enters cells poorly, and is non-toxic and non carcinogenic in most biologic systems. An indirect effect on sperm is postulated, and this effect might be mediated through the testicular Sertoli cells that influence spermatogenesis. To test this possibility, we exposed mouse TM4 Sertoli like cultured cells to 1mM CrCl(3) x 6H(2)O, a non-toxic dose, for 7 days and then extracted mRNA for microarray analysis. The chromium(III) chloride had modest effects on the expression of many genes, in the range of 1.5-2.3-fold. These effects provided an opportunity for development of statistical approaches for sifting microarray data in a situation where differences were small. Data were winnowed by screening for those ratios that fell outside the 99% confidence limits and/or represented a > or = 50% change in expression in the three comparison pairs. Fifty-two genes/clones were significant after the Bonferroni adjustment for multiple comparisons. The largest average increase was observed for the transcription factor Bach2, and this increase was confirmed by RT-PCR. The results show that Cr(III) has significant effects on gene expression in a Sertoli-like cell line. PMID- 12128096 TI - Periods of vertebral column sensitivity to boric acid treatment in CD-1 mice in utero. AB - Boric acid (BA) has many uses as an industrial compound and is widely distributed in the environment. BA has been shown to produce rib agenesis, a rare effect in laboratory animals. This study was conducted to determine if there is a period of sensitivity to this unusual effect. BA (500 or 750 mg/kg) was administered p.o. to pregnant CD-1 mice once daily on gestational days (GDs) 6-10. A reduction of 13th rib length occurred at both dose levels. BA 400mg/kg was also administered twice daily on GD 6, 7, 8, 9, or 10 or on GDs 6-8. A significant decrease in average fetal weight was observed in all treatment groups. Significant increases in the incidence of cervical ribs/ossifications resulted from treatments on GD 7 and GDs 6-8. Rib agenesis occurred with treatment on GD 8 and GDs 6-8. Reduced rib length, a decreased incidence of supernumerary ribs (SNR), and an increased incidence of fused and/or branched ribs occurred when dams were treated GDs 6-8. Doses of 750 mg/kg given twice on day 8 produced significant increases in several thoracic skeletal anomalies. Further studies of pathogenesis are necessary to determine the earliest perturbations and the processes that are affected. The sensitivity of embryos to treatment on GD 8 to rib agenesis suggests that BA is affecting early processes such as gastrulation and presomitic mesoderm formation and patterning in this area. PMID- 12128097 TI - Expression of c-Myc and other apoptosis-related genes in Swiss Webster mouse fetuses after maternal exposure to all trans-retinoic acid. AB - The Myc family of genes regulates proliferation, differentiation, and apoptosis. Temporal expression of Myc family genes and several pro-apoptotic genes were investigated during Swiss Webster mice organogenesis after maternal treatment with an oral dose of 100 mg/kg trans-retinoic acid (RA) or vehicle on day 10 post coitum. Reverse transcriptase-polymerase chain reaction and ribonuclease protection assay revealed decreased c-myc expression at 48 h followed by an increase at 72 h in fetuses from RA-treated dams. Increased c-Myc protein was detected at 72 h in the RA-treated group. In utero RA-treatment resulted in decreased expression of max, mad, caspases, bax, and bad genes at 48 h. Terminal uridinetriphosphate nick end-labeling (TUNEL) analysis revealed increased apoptosis at 24-48 h, followed by decreased apoptosis 72 h after in utero RA exposure, which correlated with the decreased expression of pro-apoptotic genes noted at 48 h. Further investigations are needed to understand the role of Myc family genes during RA-mediated teratogenesis. PMID- 12128098 TI - Expression of genes for alcohol and aldehyde metabolizing enzymes in mouse oocytes and preimplantation embryos. AB - Alcohols and aldehydes are metabolized primarily by alcohol (ADH) and aldehyde (ALDH) dehydrogenase isozymes. Although significant progress has been made towards understanding the involvement of these isozymes in the oxidation of alcohol and aldehydes in the body, it is not known how these compounds are handled during fertilization and preimplantation embryogenesis. In this study, reverse transcription and the polymerase chain reaction (RT-PCR) was used to determine which ADH and ALDH isozymes are expressed at the oocyte, zygote, morula, and blastocyst stages of preimplantation development in the mouse. Transcripts of beta-actin and vimentin, assayed as controls, were detected at all stages, as well as Class III ADH (Adh-2) and Class 3 ALDH (Ahd-4), involved in the detoxification of formaldehyde and aromatic aldehydes, respectively. In contrast, transcripts for the major ethanol oxidizing isozyme, Class I ADH (Adh 1) was not detected during preimplantation development. Cytosolic retinol dehydrogenase (Adh-3) transcripts were marginally detected in oocytes and zygotes. The mRNA for cytosolic retinal dehydrogenase (Ahd-2), microsomal short chain retinol dehydrogenases (RoDH Type I), and the mitochondrial low-Km acetaldehyde dehydrogenase (Ahd-5) only appeared as maternal transcripts. Microsomal ALDH (Ahd-3), which is induced by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), was not expressed until the blastocyst stage. ADH and ALDH enzyme systems may guard mouse preimplantation embryos against the toxic effects of industrial pollutants, such as formaldehyde and TCDD, as well as peroxidatic aldehydes generated during lipid peroxidation. The absence of enzymes to convert ethanol to acetaldehyde, coupled with oocyte expression of the acetaldehyde-degrading enzyme, Ahd-5, may be protective for the early embryo. PMID- 12128099 TI - Comparative estrogenic effects of p-nonylphenol by 3-day uterotrophic assay and female pubertal onset assay. AB - Nonylphenol (NP) is widely used as a component of detergents, paints, pesticides, and many other formulated products. Several studies have demonstrated that NP is estrogenic in fish, avian, and mammalian cells. NP also competitively inhibits the binding of 17 beta-estradiol (E2) to the estrogen receptor (ER). However, there are relatively few in vivo data related to this issue in mammals. The aim of this study was to investigate the estrogenic activity of NP in animal models. We performed a 3-day uterotrophic assay using immature female rats for comparison with other endpoints of Tier I screening including vaginal opening (VO) in prepubertal intact female rats. For the uterotrophic assay, diethylstilbestrol (DES) (0.2 and 1.0 microg/kg) and p-NP (10, 25, 50, 100, and 200 mg/kg) were administered subcutaneously to immature Sprague-Dawley female rats for 3 consecutive days (postnatal days (PND) 20, 21, and 22). For the female pubertal onset assay, DES (0.2, 1.0, and 5.0 microg/kg) and p-NP (10, 50, and 100 mg/kg) were administered daily by oral gavage from 21 days of age for 20 days. In the uterotrophic assay, statistically significant increases in uterine wet weight were observed at doses of 100 and 200 mg/kg p-NP. DES (0.2 and 1.0 microg/kg) also significantly increased uterine weight compared to the vehicle control. In the female pubertal onset assay, the age of VO was advanced following oral exposure to DES (1.0 and 5.0 microg/kg) and p-NP (50 and 100 mg/kg). Estrous cyclicity was monitored in prepubertal rats from the day of VO to the day of necropsy. Irregular estrous cycles were observed in the groups treated with DES (5.0 microg/kg) and p-NP (50 and 100 mg/kg). High-dose DES (5.0 microg/kg) produced a persistent estrus state, whereas p-NP (50 and 100 mg/kg) increased the number of days in diestrus. Serum thyroxine (T(4)) concentrations were decreased in a dose-dependent manner by DES and p-NP treatment. A significant decrease in serum T(4) level was observed at high-dose DES (5.0 microg/kg) and p-NP (100 mg/kg). Serum TSH level was significantly increased by DES (5.0 microg/kg) treatment. Statistically significant decreases in ovarian weight were observed in female rats treated with DES (5.0 microg/kg) and p-NP (100 mg/kg). Our data demonstrate that p-NP can accelerate the onset of puberty and alter estrous cyclicity in prepubertal female rats at oral doses lower than the subcutaneous doses typically used in the uterotrophic assay. We therefore suggest that the female pubertal onset assay may be used as a sensitive testing method to detect environmental agents with weak estrogenic activity, but requires further research. PMID- 12128100 TI - Effect of acute ozone exposure on pregnant rat uterus contractile responses. AB - Pulmonary effects of ozone (O(3)) inhalation have been comprehensively studied, but little is known about its extrapulmonary consequences, particularly in the reproductive tract. Thus, the effects of an acute O(3) exposure on the contractile response of the pregnant rat uterus were evaluated. Nonpregnant and pregnant (5, 10, and 18 days of gestation) rats were exposed to air or O(3) (3 ppm) for1 h, and uterine strips isolated from these animals were studied 16-18 h later. Contractile responses to acetylcholine (ACh) and oxytocin (OT) were evaluated with respect to three parameters (area under the curve, amplitude, and frequency). O(3) did not modify the sensitivity (-logEC(50)) to either agonist at any pregnancy stage, but induced a statistically significant increase in all maximum responses to OT at gestational day 5, and increased the maximum response (area under the curve) to ACh at pregnancy days 5 and 10. Our results suggest that O(3) inhalation can produce abnormal contractility in the pregnant uterus, and identify the need for further investigation of this issue. PMID- 12128101 TI - Atrazine inhibition of testosterone production in rat males following peripubertal exposure. AB - Atrazine is currently one of the most widely used agricultural pesticides in the US and is the most frequently detected pesticide in ground and surface water. Earlier work by others has raised the possibility that atrazine can act as an endocrine disrupter in rat males. The current study examined testosterone levels following in vivo and in vitro exposure to atrazine. For in vivo exposures, juvenile rat males were administered atrazine 50mg/kg body weight per day by gavage acutely (from postnatal day (pnd) 46 to 48) and chronically (from pnd 22 to 48). In both acutely- and chronically-treated animals, serum and intratesticular levels of testosterone were significantly reduced by approximately 50%. For in vitro exposures, Leydig cells isolated from rats on pnd 49 were co-cultured with 232 microM atrazine in the presence of a maximally stimulating concentration of luteinizing hormone. Compared with cells cultured with vehicle and luteinizing hormone alone, testosterone production by Leydig cells treated with atrazine was reduced by 35%. A similar decrease was observed when Leydig cells were stimulated with dibuterol cAMP (db-cAMP) in lieu of luteinizing hormone, but not when cells were stimulated with hydroxycholesterol, indicating that the effects of atrazine on testosterone production can be bypassed. Taken together, these results demonstrate that atrazine acts as an endocrine disrupter in rat males by directly inhibiting Leydig cell testosterone production. PMID- 12128102 TI - Testicular changes induced by chronic exposure to the herbicide formulation, Tordon 75D (2,4-dichlorophenoxyacetic acid and picloram) in rats. AB - The second most used herbicide in the Vietnam war was Agent White, which contained the active components 2,4-dichlorophenoxyacetic acid (2,4-D) and 4 amino-3,5,6-trichloropicolinic acid (picloram). The herbicide formulation Tordon 75D is similar in terms of its active components to Agent White and is currently used by the agricultural industry in Australia. As part of an investigation into the possible adverse effects of this herbicide on male reproductive performance, groups of five male rats were gavaged 5 days a week for 9 weeks with either 0.125 ml/kg (low dose), 0.25 ml/kg (middle dose), or 0.5 ml/kg (high dose) Tordon 75D or water (controls). The high dose corresponded to 150 mg/kg body weight 2,4-D and 37.5 mg/kg picloram acid equivalents. At the end of the treatment period, the testes were collected, weighed, and examined histologically and blood samples were taken to determine serum testosterone. Groups of high dose animals were also examined after 1, 2, and 4 weeks treatment. The 9 weeks treatment with Tordon 75D caused severe reduction in testicular weight in some high dose animals. Histologically, the small testes showed shrunken tubules with germ cell depletion. This damage was still evident in some rats following a 21 weeks recovery period suggesting that the testicular damage was permanent. Testicular damage was not due to endocrine disruption as there were no significant differences in the serum concentration of testosterone in control animals compared to Tordon 75D-treated animals. Blood levels associated with the high dose were determined in a separate study and were much higher than those likely to be obtained by occupational exposure to this herbicide. PMID- 12128103 TI - Ginseng intestinal metabolite-I (GIM-I) reduces doxorubicin toxicity in the mouse testis. AB - The protective effect of ginseng intestinal metabolite-I (GIM-I) against doxorubicin-induced testicular toxicity was investigated in 5-week-old ICR male mice. GIM-I was administered orally to mice at a dose of 50 mg/kg daily for 4 weeks. From the second week, doxorubicin was coadministered intraperitoneally to the animals at a dose of 3 mg/kg once a week for 3 weeks (a total of 9 mg/kg). The body weight, spermatogenic activities (Sertoli cell, repopulation, and epididymal indices), and serum levels of lactate dehydrogenase (LDH) and creatine phosphokinase (CPK) were significantly decreased by doxorubicin treatment (P<0.01), while the combined treatment of GIM-I with doxorubicin resulted in parameters similar to the control. In the testes of doxorubicin-treated animals, almost all of the germ cells disappeared and were replaced by fibrinoid debris in the seminiferous tubules. Germ cell injury was significantly attenuated by GIM-I coadministration. The mRNA for phospholipid hydroperoxide glutathione peroxidase (PHGPx), a testis-specific antioxidant, was greatly decreased by doxorubicin treatment, and less decreased with GIM-I coadministration. These findings indicate that GIM-I may be partially protective against doxorubicin-induced testicular toxicity. PMID- 12128105 TI - Suppression of uterine decidualization correlated with reduction in serum progesterone levels as a cause of preimplantation embryonic loss induced by diphenyltin in rats. AB - In our previous study, diphenyltin dichloride (DPTCl) at 16.5 mg/kg and higher on days 0-3 of pregnancy was found to induce preimplantation embryonic loss in rats. In the present study, the effects of DPTCl on uterine decidualization in pseudopregnant rats, effects of ovarian hormones on uterine decidualization in ovariectomized rats, and effects of progesterone on the DPTCl-induced preimplantation embryonic loss in pregnant rats were determined. Female rats were given DPTCl by gastric intubation at 4.1, 8.3, 16.5, or 24.8 mg/kg on days 0-3 of pseudopregnancy and the decidual cell response was induced on day 4 of pseudopregnancy. The uterine weight on day 9 of pseudopregnancy served as an index of uterine decidualization. A significant decrease in uterine weight, which indicates suppression of uterine decidualization, was detected at 16.5 and 24.8 mg/kg. Ovarian weight and number of corpora lutea in the DPTCl-treated groups were comparable to the controls. A significant decrease in the serum progesterone levels was found at 16.5 and 24.8 mg/kg in pseudopregnant rats. The hormonal regimen consisting of progesterone and estrone-supported decidual development in ovariectomized rats given DPTCl. Pregnancy rate and number of implantations were significantly lower in the intact mated groups given DPTCl at 16.5 and 24.8 mg/kg on days 0-3 of pregnancy than in the control group and significantly higher in the groups given DPTCl plus progesterone than in the groups given DPTCl alone. These results show that reduction in serum progesterone levels is correlated with suppression of uterine decidualization and progesterone protects against preimplantation embryonic loss induced by DPTCl. PMID- 12128104 TI - Alteration in ovarian gene expression in response to 2,3,7,8-tetrachlorodibenzo-p dioxin: reduction of cyclooxygenase-2 in the blockage of ovulation. AB - 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is a reproductive toxicant and endocrine disrupter that is known to block ovulation. This study was designed to investigate alterations in relevant ovarian genes that may be involved in the blockage of ovulation by TCDD in immature intact rats primed with equine chorionic gonadotropin (eCG). In this ovulation model, rats were given either 32 microg/kg TCDD or corn oil by gavage on 25 days of age. The next day, eCG (5 IU) was injected subcutaneously (s.c.) to stimulate follicular development. Ovulation occurs 72 h after administration of eCG in controls of this model. TCDD blocked ovulation at the expected time and also reduced both ovarian and body weights. At 72 h after eCG (the morning after expected ovulation), TCDD did not alter significantly serum concentrations of progesterone (P4) and androstenedione (A4). However, estradiol (E2) was significantly higher at 72 h after eCG in TCDD treated rats when compared with controls. Western blots revealed that ovarian CYP1A1 was induced by TCDD. In addition, the aryl hydrocarbon receptor (AhR) and AhR nuclear translocator (ARNT) were down- and up-regulated by TCDD, respectively, indicating that AhR-mediated signal transduction was altered in the ovary. Ovarian estrogen receptor (ER)alpha, ER beta and progesterone receptor (PR) were not altered significantly by TCDD, but ovarian glucocorticoid receptor (GR) was increased at 24h after TCDD and decreased at 72 h after eCG when compared with controls. TCDD induced the early appearance of ovarian plasminogen activator inhibitor type-1 (PAI-1), plasminogen activator inhibitor type-2 (PAI 2), urokinase plasminogen activator (uPA), and tissue plasminogen activator (tPA) at 24h after dosing when compared with controls. On the morning after ovulation (72 h after eCG), no significant differences between control and TCDD-treated rats were observed except that TCDD had still increased tPA and decreased PAI-2 when compared with controls. Interestingly, ovarian COX-2 was induced on the morning after ovulation (72 h after eCG) in controls, but was greatly inhibited in TCDD-treated rats at that time. On the other hand, COX-1 was constitutively expressed throughout the ovulatory period and remained unaffected by TCDD. Immunolocalization of COX-2 in the ovary revealed that TCDD inhibited COX-2 expression in the granulosa cell layer when assessed in the morning of expected ovulation. In conclusion, AhR signaling is activated in the ovary by TCDD and inhibition of COX-2 appeared to be a critical step in the TCDD blockage of ovulation because blockage or reduction of COX-2 expression is well known to be associated with failure of ovulation. PMID- 12128106 TI - Valproate irreversibly alters steroid secretion patterns from porcine follicular cells in vitro. AB - The aim of the present study was to investigate whether exposure of porcine ovarian follicular cells to clinically relevant concentrations of valproate affected steroid production in vitro and to which extent these effects were reversed by removal of the drug from the culture medium. Small and medium follicles were obtained from ovaries collected, respectively, at days 8-10 and 14 16 of the estrous cycle. Theca and granulosa cells were collected from follicles and co-cultured in one well. To show whether the effect of valproate was reversible, cells were cultured for 24, 48, or 72 h with valproate 100 or 250 microg/ml. The medium was then changed to fresh medium without drugs for an additional 24, 48, or 72 h. Valproate added to the culture medium caused a significant reduction of estradiol secretion with concomitant increase in both testosterone and progesterone secretion by small follicles. In medium-sized follicles, 100 microg/ml valproate was without effect on estradiol secretion while 250 microg/ml caused a small, but statistically significant decrease. The effects of valproate on steroid secretion patterns were irreversibly independent of concentration, exposure time, and time of restoration after drug exposure up to 72 h in both small and medium follicles. In conclusion, the present study demonstrated that even short-term valproate treatment disrupted follicular steroidogenesis in isolated ovarian follicular cells resulting in increased testosterone and progesterone and decreased estradiol secretion. It was not possible to reverse the steroidogenic effects of valproate by removing the drug from the cell cultures. PMID- 12128108 TI - Lack of radiation dose response for patients with low-risk clinically localized prostate cancer: a retrospective analysis. AB - PURPOSE: The need for dose escalation for patients with low-risk clinically localized prostate cancer remains controversial. In this study, we report our pooled institutional experience of low-risk patients treated with a range of "standard" radiation doses to assess outcome in regard to biochemical failure and to determine whether a dose-response relationship exists within this conventional dose range. METHODS AND MATERIALS: Patients with low-risk clinically localized prostate cancer (T1 or T2a, Gleason grade 6660 cGy). To ensure that any differences in biochemical failure between patients at the lower and higher ends of the dose range used were not masked by analysis of the entire cohort, patients receiving or=6800 cGy were subsequently analyzed separately. Biochemical failure was defined per the ASTRO consensus definition. The log-rank test was used to assess for differences in follow-up and time to biochemical failure. A Kaplan-Meier plot of time to biochemical failure for the initial 3 subgroups was generated. RESULTS: A total of 264 patients were identified. Sixteen patients whose dose was not recorded in the database were excluded from analysis. The median follow-up was 35 months. No significant differences were found in baseline prostate-specific antigen, Gleason grade, or clinical stage among the groups. The overall actuarial rate of 5-year freedom from biochemical failure was 80.2%. By dose level, the 5-year biochemical failure free rate was 79.2%, 78.4%, and 84.5% for <6660 cGy, 6660 cGy, and >6660 cGy, respectively. These differences were not significant. Subsequently, 45 patients receiving or=6800 cGy. The difference between these groups was not significant. The 5-year freedom from biochemical failure rate for the patients receiving 10 ng/mL; the FFF rate was 62% for the 78 Gy arm vs. 43% for those who received 70 Gy (p = 0.01). For patients with a pretreatment PSA 10 ng/mL who were treated to 78 Gy (98% vs. 88% at 6 years, p = 0.056). Rectal side effects were also significantly greater in the 78 Gy group. Grade 2 or higher toxicity rates at 6 years were 12% and 26% for the 70 Gy and 78 Gy arms, respectively (p = 0.001). Grade 2 or higher bladder complications were similar at 10%. For patients in the 78 Gy arm, Grade 2 or higher rectal toxicity correlated highly with the proportion of the rectum treated to >70 Gy. CONCLUSION: An increase of 8 Gy resulted in a highly significant improvement in FFF for patients at intermediate-to-high risk, although the rectal reactions were also increased. Dose escalation techniques that limit the rectal volume that receives >or=70 Gy to <25% should be used. PMID- 12128109 TI - High-dose intensity modulated radiation therapy for prostate cancer: early toxicity and biochemical outcome in 772 patients. AB - PURPOSE: To report the acute and late toxicity and preliminary biochemical outcomes in 772 patients with clinically localized prostate cancer treated with high-dose intensity-modulated radiotherapy (IMRT). METHODS AND MATERIALS: Between April 1996 and January 2001, 772 patients with clinically localized prostate cancer were treated with IMRT. Treatment was planned using an inverse-planning approach, and the desired beam intensity profiles were delivered by dynamic multileaf collimation. A total of 698 patients (90%) were treated to 81.0 Gy, and 74 patients (10%) were treated to 86.4 Gy. Acute and late toxicities were scored by the Radiation Therapy Oncology Group morbidity grading scales. PSA relapse was defined according to The American Society of Therapeutic Radiation Oncology Consensus Statement. The median follow-up time was 24 months (range: 6-60 months). RESULTS: Thirty-five patients (4.5%) developed acute Grade 2 rectal toxicity, and no patient experienced acute Grade 3 or higher rectal symptoms. Two hundred seventeen patients (28%) developed acute Grade 2 urinary symptoms, and one experienced urinary retention (Grade 3). Eleven patients (1.5%) developed late Grade 2 rectal bleeding. Four patients (0.1%) experienced Grade 3 rectal toxicity requiring either one or more transfusions or a laser cauterization procedure. No Grade 4 rectal complications have been observed. The 3-year actuarial likelihood of >/= late Grade 2 rectal toxicity was 4%. Seventy-two patients (9%) experienced late Grade 2 urinary toxicity, and five (0.5%) developed Grade 3 urinary toxicity (urethral stricture). The 3-year actuarial likelihood of >/= late Grade 2 urinary toxicity was 15%. The 3-year actuarial PSA relapse-free survival rates for favorable, intermediate, and unfavorable risk group patients were 92%, 86%, and 81%, respectively. CONCLUSIONS: These data demonstrate the feasibility of high-dose IMRT in a large number of patients. Acute and late rectal toxicities seem to be significantly reduced compared with what has been observed with conventional three-dimensional conformal radiotherapy techniques. Short-term PSA control rates seem to be at least comparable to those achieved with three-dimensional conformal radiotherapy at similar dose levels. Based on this favorable risk:benefit ratio, IMRT has become the standard mode of conformal treatment delivery for localized prostate cancer at our institution. PMID- 12128111 TI - Implementation and utility of a daily ultrasound-based localization system with intensity-modulated radiotherapy for prostate cancer. AB - PURPOSE: To evaluate the clinical feasibility of daily computer-assisted transabdominal ultrasonography for target position verification in the setting of intensity-modulated radiotherapy (IMRT) for prostate cancer. METHODS AND MATERIALS: Twenty-three patients with clinically localized prostate cancer were treated using a sequential tomotherapy IMRT technique (Peacock) and daily computer-assisted transabdominal ultrasonography (BAT) for target localization. Patients were instructed to maintain a full bladder and were placed in the supine position using triangulation tattoos and a leg immobilizer to minimize pelvic rotation. The BAT ultrasound system is docked to the treatment collimator and electronically imports the CT simulation target contours and isocenter. The system is able to use the machine isocenter as a reference point to overlay the corresponding CT contours onto the ultrasound images captured in the transverse and sagittal planes. A touch screen menu is used to maneuver the CT contours in three dimensions such that they match the ultrasound images. The system then displays the three-dimensional couch shifts required to produce field alignment. Data were prospectively collected to measure the frequency by which useful ultrasound images were obtained, the amount of time required for localization/setup, and the direction/magnitude of the positional adjustments. RESULTS: Of the 23 patients, the BAT ultrasound system produced images of sufficient quality to perform the overlay of the CT contours in 19 patients such that positional verification could be reliably performed. Poor image quality was associated with patient inability to maintain a full bladder, large body habitus, or other anatomic constraints. Of the 19 assessable patients, a total of 185 treatment alignments were performed (mean 8.8/patient). For all cases, the average time required for the daily ultrasound imaging and positional adjustments was 11.9 min. After the initial 5 cases, the user experience skills improved such that the time required for image verification/positional adjustments decreased to a mean of 5.6 min. The average right-left, AP, and cranial-caudal adjustment was 2.6 +/- 2.1 mm, 4.7 +/- 2.7 mm, and 4.2 +/- 2.8 mm, respectively. Positional adjustments >10 mm were infrequent and related primarily to misidentification of the target structures on the ultrasound image, patient movement, or improper registration of the triangulation tattoos. CONCLUSION: Daily computer-assisted BAT ultrasound positional verification of the prostate can be successfully performed through the acquisition of high-quality images in most patients with only a modest increase in treatment setup time. Positional data obtained with this system resulted in clinically meaningful adjustments in daily setup for sequential IMRT that would not be otherwise apparent from other verification modalities. PMID- 12128110 TI - Three-dimensional intrafractional movement of prostate measured during real-time tumor-tracking radiotherapy in supine and prone treatment positions. AB - PURPOSE: To quantify three-dimensional (3D) movement of the prostate gland with the patient in the supine and prone positions and to analyze the movement frequency for each treatment position. METHODS AND MATERIALS: The real-time tumor tracking radiotherapy (RTRT) system was developed to identify the 3D position of a 2-mm gold marker implanted in the prostate 30 times/s using two sets of fluoroscopic images. The linear accelerator was triggered to irradiate the tumor only when the gold marker was located within the region of the planned coordinates relative to the isocenter. Ten patients with prostate cancer treated with RTRT were the subjects of this study. The coordinates of the gold marker were recorded every 0.033 s during RTRT in the supine treatment position for 2 min. The patient was then moved to the prone position, and the marker was tracked for 2 min to acquire data regarding movement in this position. Measurements were taken 5 times for each patient (once a week); a total of 50 sets for the 10 patients was analyzed. The raw data from the RTRT system were filtered to reduce system noise, and the amplitude of movement was then calculated. The discrete Fourier transform of the unfiltered data was performed for the frequency analysis of prostate movement. RESULTS: No apparent difference in movement was found among individuals. The amplitude of 3D movement was 0.1-2.7 mm in the supine and 0.4-24 mm in the prone positions. The amplitude in the supine position was statistically smaller in all directions than that in the prone position (p < 0.0001). The amplitude in the craniocaudal and AP directions was larger than in the left-right direction in the prone position (p < 0.0001). No characteristic movement frequency was detected in the supine position. The respiratory frequency was detected for all patients regarding movement in the craniocaudal and AP directions in the prone position. The results of the frequency analysis suggest that prostate movement is affected by the respiratory cycle and is influenced by bowel movement in the prone position. CONCLUSION: The results of this study have confirmed that internal organ motion is less frequent in the supine position than in the prone position in the treatment of prostate cancer. RTRT would be useful in reducing uncertainty due to the effects of the respiratory cycle, especially in the prone position. PMID- 12128112 TI - Initial experience with ultrasound localization for positioning prostate cancer patients for external beam radiotherapy. AB - PURPOSE: Transabdominal ultrasound localization of the prostate gland and its immediate surrounding anatomy has been used to guide the positioning of patients for the treatment of prostate cancer. This process was evaluated in terms of (1) the reproducibility of the ultrasound measurement; (2) a comparison of patient position between ultrasound localization and skin marks determined from a CT treatment planning scan; (3) the predictive indicators of patient anatomy not well suited for ultrasound localization; (4) the measurement of prostate organ displacement resulting from ultrasound probe pressure; and (5) quality assurance measures. METHODS AND MATERIALS: The reproducibility of the ultrasound positioning process was evaluated for same-day repeat positioning by the same ultrasound operator (22 patients) and for measurements made by 2 different operators (38 patients). Differences between conventional patient positioning (CT localization with skin markings) and ultrasound-based positioning were determined for 38 patients. The pelvic anatomy was evaluated for 34 patients with pretreatment CT scans to identify predictors of poor ultrasound image quality. The displacement of the prostate resulting from pressure of the ultrasound probe was measured for 16 patients with duplicate CT scans with and without a simulated probe. Finally, daily, monthly, and semiannual quality assurance tests were evaluated. RESULTS: Self-verification tests of ultrasound positioning indicated a shift of <3 mm in approximately 95% of cases. Interoperator tests indicated shifts of <3 mm in approximately 80-90% of cases. The mean difference in patient positioning between conventional and ultrasound localization for lateral shifts was 0.3 mm (SD 2.5): vertical, 1.3 mm (SD 4.7 mm) and longitudinal, 1.0 mm (SD 5.1). However, on a single day, the differences were >10 mm in 1.5% of lateral shifts, 7% of longitudinal shifts, and 7% of vertical shifts. The depth to the isocenter, thickness of tissue overlying the bladder, and position of the prostate relative to the pubic symphysis, but not the bladder volume, were significant predictive indicators of poor ultrasound imaging. The pressure of the ultrasound probe displaced the prostate in 7 of the 16 patients by an average distance of 3.1 mm; 9 patients (56%) showed no displacement. Finally, the quality assurance tests detected ultrasound equipment defects. CONCLUSION: The ultrasound positioning system is reproducible and may indicate the need for significant positioning moves. Factors that predict poor image quality are the depth to the isocenter, thickness of tissue overlying the bladder, and position of the prostate relative to the pubic symphysis. The prostate gland may be displaced a small amount by the pressure of the ultrasound probe. A quality assurance program is necessary to detect ultrasound equipment defects that could result in patient alignment errors. PMID- 12128113 TI - Influence of 3D-CRT pelvic irradiation on outcome in prostate cancer treated with external beam radiotherapy. AB - PURPOSE: The role of pelvic irradiation (PRT) in the treatment of prostate cancer remains unclear. We reviewed our institution's experience with three-dimensional conformal external beam radiotherapy (3D-CRT) during the prostate-specific antigen era to determine the influence of PRT on the risk of biochemical recurrence in patients who have a predicted risk of lymph node involvement. METHODS AND MATERIALS: Between March 1985 and January 2001, 1832 patients with clinically localized prostate cancer were treated with definitive 3D-CRT. All treatments involved CT planning to ensure coverage of the intended targets. Treatment consisted of prostate-only treatment, prostate and seminal vesicle treatment, or PRT of lymph nodes at risk followed by a boost. To create relatively homogenous analysis groups, each patient's percentage of risk of lymph node (%rLN) involvement was assigned by matching the patient's T stage, Gleason score, and initial prostate-specific antigen level to the appropriate value as described in the updated Partin tables. Three categories of %rLN involvement were defined: low, 0-5%; intermediate, >5-15%; and high, >15%. Biochemical recurrence was defined as the first occurrence of either the American Society for Therapeutic Radiology and Oncology consensus definition of prostate-specific antigen failure or the initiation of salvage hormonal therapy for any reason. RESULTS: The risk status (%rLN) could be determined for 709 low-risk, 263 intermediate-risk, and 309 high-risk patients. The actuarial freedom from biochemical recurrence (bNED) and the log-rank test for the similarity of the control and treatment survival functions are reported for each risk group. Multivariate analysis demonstrated a statistically significant benefit for the entire population treated with PRT, with a relative risk reduction of 0.72 (95% confidence interval 0.54-0.97). Although the multivariate analysis could not determine the patient population that would most benefit from PRT, the beneficial effect appeared to be most pronounced within the intermediate-risk group. Univariate analysis revealed that the intermediate-risk patients treated with PRT had an improved 2-year bNED rate, 90.1% vs. 80.6% (p = 0.02), and both low-risk and high-risk patients treated with PRT had statistically similar 2-year bNED rates compared with those who did not receive it. CONCLUSION: Pelvic 3D-CRT appears to improve bNED in prostate cancer patients. Additional studies are needed to elucidate the %rLN population for which this treatment should be recommended. PMID- 12128114 TI - Long-term use of combined radiation therapy and hormonal therapy in the management of stage D1 prostate cancer. AB - PURPOSE: To determine tumor response, patterns of relapse, and prognostic indicators in patients followed long-term after combined hormonal radiation therapy of adenocarcinoma of the prostate in men with tumor metastatic to pelvic lymph nodes. METHODS AND MATERIALS: Seventy-nine patients with adenocarcinoma of the prostate with pathologically confirmed pelvic lymph node metastases were treated with combined radiation therapy and hormonal therapy. Of these, 55 patients (70%) had T3 disease, with the remainder having earlier-stage disease; 45 (57%) patients had N2 disease (Whitmore-Jewett staging). No distant metastases were detected at initial staging, and no patient had radiographic or pathologic involvement of the para-aortic nodes. Pelvic lymph nodes were irradiated to a dose 45-54 Gy, and the prostate was irradiated to a dose 65-71.8 Gy. Hormonal therapy began up to 2 months before radiation and continued indefinitely. Patients were allowed to select their hormonal therapy and could choose diethylstilbestrol (DES) (2 patients), orchiectomy (21 patients), luteinizing hormone-releasing hormone agonist (12 patients), or combined androgen blockade (44 patients). Prognostic factors examined included microscopic vs. measurable lymph node involvement, one-sided vs. two-sided disease, T stage, pretreatment PSA, method of androgen blockade, and Gleason score. Log-rank analysis was used to determine statistical significance with respect to overall survival, disease free survival, clinical freedom from progression, and biochemical freedom from progression; Cox multivariate analysis was employed to determine potential confounders. RESULTS: Median follow-up was 6.7 years. There were 25 recurrences among the 79 patients, including 7 biochemical recurrences without clinical evidence of disease, three local recurrences in the prostate, and distant metastases in 14 patients; 2 patients were deceased, with cause of death listed as prostate cancer, though the location of recurrence was unknown. Patients with biochemical failure before 5 years were more likely to fail distantly, 16% vs. 4% (p < 0.001). Overall actuarial survival at 5, 8, and 12 years was 86%, 72%, and 53%, respectively, whereas actuarial disease-free survival was 90%, 87%, and 81%. Ten patients died of intercurrent disease; these included 4 patients who died of a separate (nonpelvic) malignancy of nonadenocarcinomatous histology with no elevation in PSA. When the potential prognostic variables were examined, a trend toward increased biochemical recurrence in patients with Gleason score >or=8 was observed; this became statistically significant when the 4 patients with known residual lymph node disease after biopsy were excluded (p < 0.03). Gleason score remained the only significant indicator on multivariate analysis. A single long term toxic event, recto-ureteral fistula, was observed. CONCLUSION: Combined hormonal and radiation therapy continues to represent an effective treatment option for patients with adenocarcinoma of the prostate with metastasis confined to pelvic lymph nodes. All patient groups seem to have a better disease-free survival than that reported previously in single-modality hormone or radiation treatment series. There is a suggestion that patients with lower Gleason score have a lower risk for recurrence. Combined modality therapy may also extend disease-free survival and allow patients to maintain independent function. PMID- 12128115 TI - Freedom from castration: an alternative end point for men with localized prostate cancer treated by external beam radiation therapy. AB - PURPOSE: Androgen deprivation therapy (ADT) is frequently given to men with localized prostate cancer. This study was designed to determine what proportion of men require subsequent ADT if their localized prostate cancer is first treated by radiation. METHODS AND MATERIALS: A retrospective review of the outcome of 768 men with T1-4NxM0 prostate cancer treated with external beam radiation at a single institution from 1988 to 1995. The median follow-up for the entire group was 5.8 years. The end points analyzed were biochemical failure (3 successive rises in prostate-specific antigen) and ADT (either medical or surgical castration). RESULTS: A total of 322 biochemical failure events occurred among the 768 treated patients, and 187 began ADT during the period of observation. Fifty-four percent of men starting ADT did so without rebiopsy or radiographic evidence of persistent disease. The overall freedom from biochemical failure rate at 10 years was 43.3% and the 10-year freedom from ADT rate was 56.3%. The corresponding values for the most favorable subgroup (T1-2a, Gleason sum 6, prostate-specific antigen or=70, and no pelvic lymphadenopathy or distant metastases. The total dose to the prostate was 70.2 Gy in 20 patients and 73.8 Gy in 9 patients. Therapy was delivered using a 4-field technique with three dimensional conformal planning. Amifostine was administered intrarectally as an aqueous solution 30 min before irradiation on the first 15 days of therapy. Amifostine was escalated in cohorts from 500 to 2500 mg. Proctoscopy was performed before therapy and at 9 months after completion. Most patients underwent repeat proctoscopy at 18 months. On Days 1 and 10 of radiotherapy, serum samples were collected for pharmacokinetic studies. The clinical symptoms (Radiation Therapy Oncology Group scale) and a proctoscopy score were assessed during follow-up. RESULTS: All patients completed therapy with no amifostine related toxicity at any dose level. The application was feasible and well tolerated. No substantial systemic absorption occurred. With a median follow-up of 26 months, 9 patients (33%) developed rectal bleeding (8 Grade 1, 1 Grade 2). At 9 months, 16 and 3 patients developed Grade 1 and Grade 2 telangiectasia, respectively. This was mostly confined to the anterior rectal wall. No visible mucosal edema, ulcerations, or strictures were noted. No significant differences were found between the proctoscopy findings at 9 and 18 months. Four patients (14%) developed symptoms suggestive of radiation damage that, on sigmoidoscopy, proved to be secondary to unrelated processes. These included preexisting nonspecific proctitis (n = 1), diverticular disease of the sigmoid colon (n = 1), rectal polyp (n = 1), and ulcerative colitis (n = 1). Symptoms developed significantly more often in patients receiving 500-1000 mg than in patients receiving 1500-2500 mg amifostine (7 [50%] of 14 vs. 2 [15%] of 13, p = 0.0325, one-sided chi-square test). CONCLUSION: Intrarectal application of amifostine is feasible and well tolerated. Systemic absorption of amifostine and its metabolites is negligible, and close monitoring of patients is not required with rectal administration. Proctoscopy is superior to symptom score as a method of assessing radiation damage of the rectal wall. The preliminary efficacy data are encouraging, and further clinical studies are warranted. PMID- 12128117 TI - Treatment outcome, body image, and sexual functioning after orchiectomy and radiotherapy for Stage I-II testicular seminoma. AB - PURPOSE: Orchiectomy followed by infradiaphragmatic irradiation is the standard treatment for Stage I-II testicular seminoma in The Netherlands. Because body image and sexual functioning can be affected by treatment, a retrospective study was carried out to assess treatment outcome, body image, and changes in sexuality after orchiectomy and radiotherapy. METHODS AND MATERIALS: The medical charts of 166 patients with Stage I-II testicular seminoma were reviewed. A questionnaire on body image and current sexual functioning regarding the frequency and quality of erections, sexual activity, significance of sex, and changes in sexuality was sent to 157 patients (at a mean of 51 months after treatment). RESULTS: Seventy eight percent (n = 123, mean age 42 years) completed the questionnaire. During irradiation, almost half of patients experienced nausea and 19% nausea and vomiting. Only 3 patients had disease relapse. After treatment, about 20% reported less interest and pleasure in sex and less sexual activity. Interest in sex, erectile difficulties, and satisfaction with sexual life did not differ from age-matched healthy controls. At the time of the survey, 17% of patients had erectile difficulties, a figure that was significantly higher than before treatment, but which correlated also with age. Twenty percent expressed concerns about fertility, and 52% found their body had changed after treatment. Cancer treatment had negatively influenced sexual life in 32% of the patients. CONCLUSIONS: Orchiectomy with radiotherapy is an effective and well-tolerated treatment for Stage I-II testicular seminoma. Treatment-induced changes in body image and concerns about fertility were detected, but the sexual problems encountered did not seem to differ from those of healthy controls, although baseline data are lacking. PMID- 12128118 TI - Determination and delineation of nodal target volumes for head-and-neck cancer based on patterns of failure in patients receiving definitive and postoperative IMRT. AB - PURPOSE: We present the guidelines for target volume determination and delineation of head-and-neck lymph nodes based on the analysis of the patterns of nodal failure in patients treated with intensity-modulated radiotherapy (IMRT). METHODS AND MATERIALS: Data pertaining to the natural course of nodal metastasis for each head-and-neck cancer subsite were reviewed. A system was established to provide guidance for nodal target volume determination and delineation. Following these guidelines, 126 patients (52 definitive, 74 postoperative) were treated between February 1997 and December 2000 with IMRT for head-and-neck cancer. The median follow-up was 26 months (range 12-55), and the patterns of nodal failure were analyzed. RESULTS: These guidelines define the nodal target volume based on the location of the primary tumor and the probability of microscopic metastasis to the ipsilateral and contralateral (Level I-V) nodal regions. Following these guidelines, persistent or recurrent nodal disease was found in 6 (12%) of 52 patients receiving definitive IMRT, and 7 (9%) of 74 patients receiving postoperative IMRT had failure in the nodal region. CONCLUSION: On the basis of our clinical experience in implementing inverse-planning IMRT for head-and-neck cancer, we present guidelines using a simplified, but clinically relevant, method for nodal target volume determination and delineation. The intention was to provide a foundation that enables different institutions to exchange clinical experiences in head-and-neck IMRT. These guidelines will be subject to future refinement when the clinical experience in head-and-neck IMRT advances. PMID- 12128119 TI - Effects of breathing a hyperoxic hypercapnic gas mixture on blood oxygenation and vascularity of head-and-neck tumors as measured by magnetic resonance imaging. AB - PURPOSE: For head-and-neck tumors, breathing a hyperoxic hypercapnic gas mixture and administration of nicotinamide has been shown to result in a significantly improved tumor response to accelerated radiotherapy (ARCON, Accelerated Radiotherapy with CarbOgen and Nicotinamide). This may be caused by improved tumor oxygenation, possibly mediated by vascular effects. In this study, both blood oxygenation and vascular effects of breathing a hyperoxic hypercapnic gas mixture (98% O2 + 2% CO2) were assessed by magnetic resonance imaging (MRI) in patients with head-and-neck tumors. METHODS AND MATERIALS: Tumor vascularity and oxygenation were investigated by dynamic gadolinium contrast-enhanced MRI and blood oxygen level dependent (BOLD) MRI, respectively. Eleven patients with primary head-and-neck tumors were each measured twice; with and without breathing the hyperoxic hypercapnic gas mixture. RESULTS: BOLD MR imaging revealed a significant increase of the MRI time constant of transverse magnetization decay (T2*) in the tumor during hypercapnic hyperoxygenation, which correlates to a decrease of the deoxyhemoglobin concentration. No changes in overall tumor vascularity were observed, as measured by the gadolinium contrast uptake rate in the tumor. CONCLUSION: Breathing a hyperoxic hypercapnic gas mixture improves tumor blood oxygenation in patients with head-and-neck tumors, which may contribute to the success of the ARCON therapy. PMID- 12128120 TI - Hypoxia-inducible factor (HIF1A and HIF2A), angiogenesis, and chemoradiotherapy outcome of squamous cell head-and-neck cancer. AB - PURPOSE: Hypoxia-inducible factors HIF1alpha and HIF2alpha (HIFalphas) regulate the expression of a variety of genes encoding proteins related to angiogenesis and to anaerobic metabolism of cells exposed to hypoxic stress. Their putative role as markers of clinically relevant hypoxia and, therefore, as predictors of response to chemoradiotherapy is herein examined. PATIENTS AND METHODS: Using immunohistochemistry, we assessed the expression of HIFalphas in normal head-neck mucosa and in 75 cancer specimens from patients with locally advanced squamous cell head-and-neck cancer (SCHNC), treated with concurrent carboplatin chemoradiotherapy. RESULTS: Head-and-neck mucosa from normal individuals did not show any HIF1alpha or HIF2alpha reactivity. SCHNC showed a varying expression of HIFalphas ranging through negative reactivity, to weak or focally strong cytoplasmic reactivity, or to strong diffuse cytoplasmic/nuclear reactivity. Fifty-two percent and 33% of cancer samples showed the latter expression pattern for HIF1alpha and HIF2alpha, respectively, and were considered to bear "high" HIF reactivity. Bone/cartilage involvement was more frequent in tumors with high HIF1alpha expression (p = 0.05). HIF1alpha and HIF2alpha overexpression were significantly associated with high microvessel density (p = 0.002 and 0.02, respectively) and with VEGF expression (p = 0.01 and 0.005, respectively). HIF1alpha was related to high thymidine phosphorylase expression (p = 0.03), whereas VEGF/KDR-activated tumor vasculature was significantly more frequent in HIF2alpha-overexpressing tumors (p = 0.02). High HIF1alpha and HIF2alpha were associated with incomplete response to chemoradiation (p = 0.007 and p = 0.02, respectively). In univariate analysis, high HIF1alpha and HIF2alpha expression were significantly associated with poor local relapse-free survival (p = 0.003 and 0.003, respectively) and with poor overall survival (p = 0.05 and 0.001, respectively). In multivariate models, HIF2alpha expression was an independent prognostic factor. In biopsies performed after the delivery of 20 Gy of radiotherapy, upregulation of HIFalphas was noted in some cases. CONCLUSIONS: It is concluded that the overexpression of HIFalphas in SCHNC is related to locally aggressive behavior, to intensification of angiogenesis, and to an important resistance to carboplatin chemoradiotherapy. PMID- 12128121 TI - Arterial baroreflex and peripheral chemoreflex function after radiotherapy for laryngeal or pharyngeal cancer. AB - PURPOSE: Denervation of the carotid sinus causes baroreflex and chemoreflex failure, resulting in labile hypertension and loss of hypoxic responsiveness. We investigated whether radiation therapy for laryngeal or pharyngeal cancer affects baroreflex and chemoreflex function. METHODS AND MATERIALS: Twelve patients were studied after radiation therapy for locally advanced laryngeal or pharyngeal cancer (11 male, 1 female, age: 56.0 +/- 7.9 years), 3.3 years (median; range 1.0 4.7) after radiotherapy and 15 healthy controls (11 male, 4 female, 53.4 +/- 9.2 years). We measured baroreflex sensitivity (phenylephrine), blood pressure level and variability (24-h Spacelabs and 5-h Portapres recordings), responses to cardiovascular reflex tests, and the ventilatory responses to normocapnic and hypercapnic hypoxia. RESULTS: Baroreflex sensitivity was lower in patients (9.7 +/- 7.8 ms/mm Hg) than in controls (17.5 +/- 10.3 ms/mm Hg, p = 0.011). Mean office blood pressure was significantly higher in patients (141.5 +/- 27.8/89.2 +/- 10.6 mm Hg, 63.3 +/- 12.3 bpm) than in controls (117.3 +/- 10.1/75.1 +/- 6.8 mm Hg, 61.8 +/- 10.8 bpm). Blood pressure variability was not different between groups, nor were the responses to reflex tests. The normo/hypercapnic ventilatory response to hypoxia was similar in patients (0.21 +/- 0.10/1.37 +/- 0.60 L/min/%) and controls (0.22 +/- 0.16/1.19 +/- 0.78 L/min/%). CONCLUSIONS: Radiation therapy for laryngeal or pharyngeal carcinoma does not affect chemoreflex function, but results in an attenuated baroreflex sensitivity. Clinically relevant blood pressure lability is absent however. PMID- 12128122 TI - What margins are necessary for incorporating mediastinal nodal mobility into involved-field radiotherapy for lung cancer? AB - PURPOSE: The mobility of mediastinal nodes was studied on multiple CT scans of the thorax from patients with non-small-cell lung cancer. PATIENTS AND METHODS: A total of 10 enlarged mediastinal nodes/masses were identified in 8 patients with non-small-cell lung cancer. Nodal locations were classified using the Naruke/ATS LCSG system, and between 3 and 6 scans were available for each site. The CT data sets were coregistered, and the contoured nodes were automatically projected onto the initial planning CT scan. An encompassing nodal volume (ENV) of all contours of a particular node was manually contoured on all scans. Individual nodal volumes were expanded in three dimensions to establish additional margins required to encompass the ENV. RESULTS: The mean volume of nodes studied ranged from 0.8 to 23.2 cc. The addition to individual nodes of a margin of 5 mm was found to result in a mean ENV coverage of >or=95% at all sites. For individual nodes at locations N4R, N5, and N6, however, the coverage ranged from 87.8% to 92.6%. CONCLUSION: The addition of a margin of 5 mm to individual mediastinal nodes seems to be adequate to account for variations in both contouring and mobility. PMID- 12128123 TI - Association of Ki-67, p53, and bcl-2 expression of the primary non-small-cell lung cancer lesion with brain metastatic lesion. AB - PURPOSE: The study was conducted to determine whether immunohistochemical analysis of Ki-67, p53, and bcl-2 in patients with non-small-cell lung cancer is associated with a higher rate of brain metastases and whether the intrapatient expression of these biomarkers (in the primary tumors vs. brain lesions) is similar. METHODS AND MATERIALS: At the M. D. Anderson Cancer Center, tumors from 29 case patients with primary lung tumor and brain metastasis and 29 control patients with primary lung tumor but no brain metastasis were resected and examined for immunohistochemical expression. Ki-67, p53, and bcl-2 were analyzed in resected primary lung, lymph node, and metastatic brain tumors. Each control patient was matched by age, gender, and histology to a patient with brain metastasis. RESULTS: No significant differences in patient survival characteristics were detected between the case group and control group. Also, difference in patient outcome between the two groups was not generally predicted by biomarker analysis. However, when the groups were combined, the biomarker analysis was predictive for certain patient outcome end points. Using median values as cutoff points between low and high expression of biomarkers, it was observed that high expression of Ki-67 (>40%) in lung primaries was associated with poorer disease-free survival (p = 0.04), whereas low expression of p53 in lung primaries was associated with poorer overall survival (p = 0.04), and these patients had a higher rate of nonbrain distant metastases (p = 0.02). The patients with brain metastases were particularly prone to developing nonbrain distant metastases if the percentage of p53-positive cells in brain metastases was low (p = 0.01). There was a positive correlation in the expression of Ki-67 (p = 0.02)(r(2) = 0.1608), as well as p53 (p < 0.001) (r(2) = 0.7380), between lung primaries and brain metastases. Compared to Ki-67 and p53, bcl-2 was the least predictive. CONCLUSION: Differences in biomarker expression between the case and control groups did not serve as significant predictors of brain metastasis or patient survival. There was a strong correlation between lung primary biomarker expression and brain metastasis expression for Ki-67 and p53. Univariate analysis showed that low p53 and high Ki-67 expression predicted poor prognosis. This study shows that there may be a strong correlation between biomarker expression in non-small-cell lung cancer primary tumors and their brain metastases. PMID- 12128124 TI - Locoregional treatment outcomes for inoperable anthracycline-resistant breast cancer. AB - PURPOSE: To assess the therapeutic outcomes and treatment-related morbidity of patients treated with radiation for inoperable breast cancer resistant to anthracycline-containing primary chemotherapy. METHODS AND MATERIALS: We analyzed the medical records of breast cancer patients treated on five consecutive institutional trials who had been designated as having inoperable locoregional disease after completion of primary chemotherapy, without evidence of distant metastases at diagnosis. The cohort for this analysis was 38 (4.4%) of 867 patients enrolled in these trials. Kaplan-Meier statistics were used for survival analysis, and prognostic factors were compared using log-rank tests. The median follow-up of surviving patients was 6.1 years. RESULTS: Thirty-two (84%) of the 38 patients were able to undergo mastectomy after radiotherapy. For the whole group, the overall survival rate at 5 years was 46%, with a distant disease-free survival rate of 32%. The 5-year survival rate for patients who were inoperable because of primary disease extent was 64% compared with 30% for those who were inoperable because of nodal disease extent (p = 0.0266). The 5-year rate of locoregional control was 73% for the surgically treated patients and 64% for the overall group. Of the 32 who underwent mastectomy, the 5-year rate of significant postoperative complications was 53%, with 4 (13%) requiring subsequent hospitalization and additional surgical revision. Preoperative radiation doses of >or=54 Gy were significantly associated with the development of complications requiring surgical treatment (70% vs. 9% for doses <54 Gy, p = 0.0257). CONCLUSION: Despite the poorer prognosis of patients with inoperable disease after primary chemotherapy, almost one-half remained alive at 5 years and one third were free of distant disease after multidisciplinary locoregional management. These patients have high rates of locoregional recurrence after preoperative radiotherapy and mastectomy, and the morbidity associated with this approach may limit dose-escalation strategies. Alternative therapeutic strategies such as novel systemic agents, use of planned myocutaneous repair for closure, or radiation combined with radiosensitizing agents, should be considered in this class of patients. PMID- 12128125 TI - Radiotherapy after high-dose chemotherapy and peripheral blood stem cell support in high-risk breast cancer. AB - PURPOSE: To assess the toxicity and efficacy of radiotherapy with respect to locoregional control after adjuvant high-dose chemotherapy for patients with breast cancer. At first, radiotherapy was withheld because of toxicity concerns, but it was introduced in 1995 because of reported high locoregional relapse rates. METHODS AND MATERIALS: Between 1992 and 1998, 40 patients with Stage II III high-risk breast cancer received adjuvant high-dose chemotherapy consisting of thiotepa, mitoxantrone, and cyclophosphamide and peripheral blood stem cell support after four cycles of induction chemotherapy. The chest wall or breast, as well as the supraclavicular nodes, were irradiated with electrons and photons to a median dose of 50.4 Gy in 20 patients. Six additional patients received only supraclavicular irradiation to a median dose of 50.4 Gy. Acute toxicity was scored clinically. Pulmonary function tests were performed in 14 irradiated patients before high-dose chemotherapy and 1.1-4.4 years (median 1.6) after irradiation. The median follow-up time of living patients was 33 vs. 67 months in irradiated (n = 26) and nonirradiated (n = 14) patients, respectively. RESULTS: G2 and G3 hematologic toxicity occurred in 1 patient each. No clinical pneumonitis or clinical impairment of lung function was observed. After 1-2 years, the lung function tests showed only minor changes in 4 patients. The 3 year locoregional control rate was 92% in the irradiated patients vs. 58% in the nonirradiated patients (p = 0.049, actuarial analysis). CONCLUSION: In this series, adjuvant radiotherapy after adjuvant chemotherapy for breast cancer appeared well tolerated, with improved local regional control and without significant side effects. Longer follow-up and more patient accrual, as well as Phase III trials, are necessary for confirmation. PMID- 12128126 TI - An evidence-based estimate of appropriate radiotherapy utilization rate for breast cancer. AB - PURPOSE: Current estimates of the proportion of cancer patients who will require radiotherapy (RT) are based almost entirely on expert opinion. We sought to use an evidence-based approach to estimate the proportion of incident cases of breast cancer that will require RT at any point in the evolution of the illness. METHODS AND MATERIALS: We undertook a systematic review of the literature to identify indications for RT for breast cancer and to ascertain the level of evidence that supported each indication. An epidemiologic approach was then used to estimate the incidence of each indication for RT in a typical North American population of breast cancer patients. The effect of sampling error on the estimated appropriate rate of RT was calculated mathematically, and the effect of systematic error was estimated by sensitivity analysis. RESULTS: It was estimated that 66.4% +/- 4.8% of breast cancer patients develop one or more indications for RT at some point in the course of the illness. The plausible range for this rate was 56.3%-72.4% on sensitivity analysis. Of all breast cancer patients, 57.3% +/- 4.7% require RT in their initial treatment and 9.1% +/- 1.0% do so later for recurrence or progression. The proportion of patients who ever require RT is stage dependent: 39.8% +/- 1.1% in ductal carcinoma in situ; 68.6% +/- 4.1% in Stage I invasive carcinoma; 81.5% +/- 2.3% in Stage II; 95.3% +/- 0.3% in Stage III; and 63.7% +/- 0.3% in Stage IV. CONCLUSION: This method provides a rational starting point for the long-term planning of RT services and for the audit of access to RT at the population level. By completing such evaluations in the major cancer sites, it will be possible to estimate the appropriate RT treatment rate for the cancer population as a whole. PMID- 12128127 TI - Metabolic imaging of low-grade gliomas with three-dimensional magnetic resonance spectroscopy. AB - PURPOSE: The role of radiotherapy (RT) seems established for patients with low grade gliomas with poor prognostic factors. Three-dimensional (3D) magnetic resonance spectroscopy imaging (MRSI) has been reported to be of value in defining the extent of glioma infiltration. We performed a study examining the impact MRSI would have on the routine addition of 2-3-cm margins around MRI T2 weighted hyperintensity to generate the treatment planning clinical target volume (CTV) for low-grade gliomas. METHODS AND MATERIALS: Twenty patients with supratentorial gliomas WHO Grade II (7 astrocytomas, 6 oligoastrocytomas, 7 oligodendrogliomas) underwent MRI and MRSI before surgery. The MRI was contoured manually; the regions of interest included T2 hyperintensity and, if present, regions of contrast enhancement on T1-weighted images. The 3D-MRSI peak parameters for choline and N-acetyl-aspartate, acquired voxel-by-voxel, were categorized using a choline/N-acetyl-aspartate index (CNI), a tool for quantitative assessment of tissue metabolite levels, with CNI 2 being the lowest value corresponding to tumor. CNI data were aligned to MRI and displayed as 3D contours. The relationship between the anatomic and metabolic information on tumor extent was assessed by comparing the CNI contours and other MRSI-derived metabolites to the MRI T2 volume. RESULTS: The limitations in the size of the region "excited" meant that MRSI could be used to evaluate only a median 68% of the T2 volume (range 38-100%), leaving the volume T2c. The CNI 2 volume (median 29 cm(3), range 10-73) was contained totally within the T2c in 55% of patients. In the remaining patients, the volume of CNI 2 extending beyond the T2c was quite small (median 2.3 cm(3), range 1.4-5.2), but was not distributed uniformly about the T2c, extending up to 22 mm beyond it. Two patients demonstrated small regions of contrast enhancement corresponding to the regions of highest CNI. Other metabolites, such as creatine and lactate, seem useful for determining less and more radioresistant areas, respectively. CONCLUSION: Metabolically active tumor, as detected by MRSI, is restricted mainly to the T2 hyperintensity in low-grade gliomas, but can extend outside it in a limited and nonuniform fashion up to 2 cm. Therefore, a CTV including T2 and areas of CNI extension beyond the T2 hyperintensity would result in a reduction in the size and a change in the shape of the standard clinical target volumes generated by adding uniform margins of 2 3 cm to the T2 hyperintensity. PMID- 12128128 TI - Intensity-modulated radiation therapy (IMRT) for meningioma. AB - PURPOSE: To assess the safety and efficacy of intensity-modulated radiation therapy (IMRT) in the treatment of intracranial meningioma. METHODS AND MATERIALS: Forty patients with intracranial meningioma (excluding optic nerve sheath meningiomas) were treated using IMRT with the NOMOS Peacock system between 1994 and 1999. Twenty-five patients received IMRT after surgery either as adjuvant therapy for incomplete resection or for recurrence, and 15 patients received definitive IMRT after presumptive diagnosis based on imaging. Thirty-two patients had skull base lesions, and 8 had nonskull base lesions. The prescribed dose ranged from 40 to 56 Gy (median 50.4 Gy) at 1.71 to 2 Gy per fraction, and the volume of the primary target ranged from 1.55 to 324.57 cc (median 20.22 cc). The mean dose to the target ranged from 44 to 60 Gy (median 53 Gy). Follow-up ranged from 6 to 71 months (median 30 months). Acute and chronic toxicity were assessed using Radiation Therapy Oncology Group (RTOG) morbidity criteria and tumor response was assessed by patient report, examination, and imaging. Overall survival, progression-free survival, and local control were calculated using the Kaplan-Meier method. RESULTS: Cumulative 5-year local control, progression-free survival, and overall survival were 93%, 88%, and 89%, respectively. Two patients progressed after IMRT, one locally and one distantly. Each was treated with IMRT after multiple recurrences of benign meningioma over many years. Both were found to have malignant meningioma at the time of relapse after IMRT, and it is likely their tumors had already undergone malignant change by the time IMRT was given. Defined normal structures generally received a significantly lower dose than the target. The most common acute central nervous system (CNS) toxicity was mild headache, usually relieved with steroids. One patient experienced RTOG Grade 3 acute CNS toxicity, and 2 experienced Grade 3 or higher late CNS toxicity, with one possible treatment-related death. No toxicity was observed with mean doses to the optic nerve/chiasm up to 47 Gy and maximum doses up to 55 Gy. CONCLUSION: IMRT is a promising new technology that is safe and efficacious in the primary and adjuvant treatment of intracranial meningiomas. A history of local aggression may indicate malignant degeneration and predict a poorer outcome. Toxicity data are encouraging, but the potential for serious side effects exists, as demonstrated by one possible treatment-related death. Larger cohort and longer follow-up are needed to better define efficacy and late toxicity of IMRT. PMID- 12128129 TI - Acute effects of irradiation on cognition: changes in attention on a computerized continuous performance test during radiotherapy in pediatric patients with localized primary brain tumors. AB - PURPOSE: To assess sustained attention, impulsivity, and reaction time during radiotherapy (RT) for pediatric patients with localized primary brain tumors. METHODS AND MATERIALS: Thirty-nine patients (median age 12.3 years, range 5.9 22.9) with primary brain tumors were evaluated prospectively using the computerized Conners' continuous performance test (CPT) before and during conformal RT (CRT). The data were modeled to assess the longitudinal changes in the CPT scores and the effects of clinical variables on these changes during the first 50 days after the initiation of CRT. RESULTS: The CPT scores exhibited an increasing trend for errors of omission (inattentiveness), decreasing trend for errors of commission (impulsivity), and slower reaction times. However, none of the changes were statistically significant. The overall index, which is an algorithm-based weighted sum of the CPT scores, remained within the range of normal throughout treatment. Older patients (age >12 years) were more attentive (p < 0.0005), less impulsive (p < 0.07), and had faster reaction times (p < 0.001) at baseline than the younger patients. The reaction time was significantly reduced during treatment for the older patients and lengthened significantly for the younger patients (p < 0.04). Patients with a shunted hydrocephalus (p < 0.02), seizure history (p < 0.0006), and residual tumor (p < 0.02) were significantly more impulsive. Nonshunted patients (p < 0.0001), those with more extensive resection (p < 0.0001), and patients with ependymoma (p < 0.006) had slower initial reaction times. CONCLUSION: Children with brain tumors have problems with sustained attention and reaction time resulting from the tumor and therapeutic interventions before RT. The reaction time slowed during treatment for patients <12 years old. RT, as administered in the trial from which these data were derived, has limited acute effects on changes in the CPT scores measuring attention, impulsiveness, and reaction time. PMID- 12128130 TI - Gamma Knife radiosurgery for numerous brain metastases: is this a safe treatment? AB - PURPOSE: Gamma Knife (GK) radiosurgery has recently been employed in patients with numerous brain metastases (METs), even those with 10 or more lesions. However, cumulative irradiation doses to the whole brain (WB), with such treatment, have not been determined. METHODS AND MATERIALS: Since the GammaPlan ver. 5.10 (ver. 5.31 is presently available, Leksell GammaPlan) became available in November 1998, 92 GK procedures have been performed for 80 patients with 10 or more brain METs at our facility. The median lesion number was 17 (range: 10-43) and the median cumulative volume of all tumors was 8.02 cc (range: 0.46-81.41 cc). The median selected dose at the lesion periphery was 20 Gy (range: 12-25 Gy). Based on these treatment protocols, the cumulative irradiation dose was computed. RESULTS: The median cumulative irradiation dose to the WB was 4.71 (range: 2.16-8.51) Gy. The median brain volumes receiving >2 Gy, >5 Gy, >10 Gy, >15 Gy, and >20 Gy were 1105 (range 410-1501) cc, 309 (46-1247) cc, 64 (13-282) cc, 24 (2-77) cc, and 8 (0-40) cc, respectively. CONCLUSION: The cumulative WB irradiation doses for patients with numerous radiosurgical targets were not considered to exceed the threshold level of normal brain necrosis. PMID- 12128131 TI - Postoperative low-pelvic irradiation for stage I-IIA cervical cancer patients with risk factors other than pelvic lymph node metastasis. AB - PURPOSE: To retrospectively investigate whether postoperative low-pelvic radiotherapy (RT) is an appropriate treatment for node-negative, high-risk Stage I-IIA cervical cancer patients. METHODS AND MATERIALS: A total of 228 Stage I-IIA cervical cancer patients treated by radical surgery and postoperative RT were included in this study. All patients had histopathologically negative pelvic node metastasis, but at least one of the following risk factors: parametrial involvement, positive or close resection margins, invasion depth two-thirds or greater cervical stromal thickness. Seventy-nine patients (35%) received 30-50 Gy (median 44) to whole pelvis and a boost dose to the low pelvis (whole-pelvic RT group); the other 149 patients (65%) received low-pelvic RT only (low-pelvic RT group). For both groups, the total external RT dose to the low pelvis ranged from 40 to 60 Gy (median 50). The potential factors associated with survival, small bowel (gastrointestinal) complications, and leg lymphedema were analyzed, and patients who had a relapse in the upper pelvis were identified. RESULTS: The 5 year overall and disease-specific survival rate was 84% and 86%, respectively. After multivariate analysis, only bulky tumor (>or=4 cm) and non-squamous cell carcinoma were significantly associated with survival. Parametrial involvement, lymph-vascular invasion, 50.4 Gy to the low pelvis, but not old age and treatment technique (AP-PA vs. box), were significantly associated with gastrointestinal complications. Three patients (2%) in the low-pelvic RT group and 6 patients (8%) in the whole-pelvic RT group were found to have Grade III or higher small bowel complications (p = 0.023). Thirty-one percent of patients developed lymphedema of the leg. A dose to the low pelvis >50.4 Gy and an AP-PA field, but not whole-pelvic RT, old age, or the number of sampled lymph nodes, were associated with lymphedema of the leg. Five patients (3.6%) of the low-pelvic RT group and none of the whole-pelvic RT group developed upper pelvis relapse. Three of these 5 patients had upper pelvic relapse alone. CONCLUSION: Compared with whole-pelvic RT plus low-pelvic boost, low-pelvic RT alone significantly reduces the small bowel complications in node negative, high-risk, Stage I-IIA cervical cancer patients. Although low-pelvic RT alone increases the incidence of upper pelvic relapse, its effect on survival is not substantial. Low-pelvic RT alone appears to be an appropriate treatment method for this group of patients. PMID- 12128132 TI - A predictive model for survival in metastatic cancer patients attending an outpatient palliative radiotherapy clinic. AB - PURPOSE: To develop a predictive model for survival from the time of presentation in an outpatient palliative radiotherapy clinic. METHODS AND MATERIALS: Sixteen factors were analyzed prospectively in 395 patients seen in a dedicated palliative radiotherapy clinic in a large tertiary cancer center using Cox's proportional hazards regression model. RESULTS: Six prognostic factors had a statistically significant impact on survival, as follows: primary cancer site, site of metastases, Karnofsky performance score (KPS), and fatigue, appetite, and shortness of breath scores from the modified Edmonton Symptom Assessment Scale. Risk group stratification was performed (1) by assigning weights to the prognostic factors based on their levels of significance, and (2) by the number of risk factors present. The weighting method provided a Survival Prediction Score (SPS), ranging from 0 to 32. The survival probability at 3, 6, and 12 months was 83%, 70%, and 51%, respectively, for patients with SPS or=20 (n = 133) (p < 0.0001). Corresponding survival probabilities based on number of risk factors were as follows: 85%, 72%, and 52% (or=5 factors)(n = 180)(p < 0.0001). CONCLUSION: Clinical prognostic factors can be used to predict prognosis among patients attending a palliative radiotherapy clinic. If validated in an independent series of patients, the model can be used to guide clinical decisions, plan supportive services, and allocate resource use. PMID- 12128133 TI - How iliopelvic lymphoscintigraphy can affect the definition of planning target volume in radiation therapy of pelvic and testicular tumors. AB - PURPOSE: External beam radiation therapy (EBRT) of most intrapelvic and testicular tumors has been generally performed with large fields encompassing both the primary disease and lymphatic drainage. This study was carried out to map the pelvic and periaortic lymphatics by means of iliopelvic lymphoscintigraphy (IPL) in preparation for radiotherapy planning. METHODS AND MATERIALS: Between January 2000 and October 2001, 70 patients scheduled for EBRT (61 operated on, 52 females, 18 males, mean age 61, range, 24-80), affected with uterine (43), rectal (11), testicular (8), anal (4), penile (2), and vulvar (2) cancers were enrolled in the study. IPL was performed by injection of 99mtechnetium-nanocolloids in the bipedal (70 cases) or bipedal plus perianal (20 cases) sites. The sensitivity of IPL in mapping the lymphatic anatomy was evaluated first. Then three radiation oncologists scored the modifications induced by IPL on the planning target volume (PTV) which had been previously delineated only on the basis of bony landmarks. The original fields were classified "inadequate" if they failed to match the new PTV by more than 1 cm. RESULTS: IPL sensitivity in showing the inguinal, external iliac, common, and periaortic lymphatics was 100%, 90%, 80%, and 70% in anterior-posterior (A-P) projections, and 100%, 80%, 70%, and 60% in lateral projections respectively. For the presacral and hypogastric ones the sensitivity was 40%. When compared with bony landmarks, IPL changed the delineation of PTV in 24 of 70 A-P P-A fields (34%) and 22 of 58 (38%) lateral fields. Furthermore, 8/12 (67%) lymphadenectomies resulted in being incomplete. No IPL-related toxicity was observed. CONCLUSION: IPL is a safe, inexpensive (cost: 100 Euros), and effective method to map the lymphatic chains. In the A-P scintigrams these structures were detected in 85% (70-100%) of the patients referred for total pelvis irradiation, and this figure could be higher in subjects not operated on. IPL can also give a reliable evaluation of the lymphadenectomies in order to schedule the proper treatments after surgery. Finally, IPL may change the conventional PTV for pelvic irradiation in about 36% (34-38%) of the cases; therefore, the fields should be tailored more around the lymphatic landmarks than the bony landmarks. PMID- 12128134 TI - Pronounced radiosensitization of cultured human cancer cells by COX inhibitor under acidic microenvironment. AB - PURPOSE: To demonstrate the influence of pH on the cytotoxicity and radiosensitization by COX (cyclooxygenase) -1 and -2 inhibitors using established human cancer cells in culture. METHODS AND MATERIALS: Nonselective COX inhibitor, ibuprofen (IB), and selective COX-2 inhibitor, SC-236, were used to determine the cytotoxicity and radiosensitization at varying pH of culture media. Human colon carcinoma cell line (HT-29) was exposed to the drug alone and in combination with radiation at different pH of the cell culture media. The end point was clonogenic ability of the single-plated cells after the treatment. RESULTS: Cytotoxicity and radiosensitization of IB increased with higher drug concentration and longer exposure time. The most significant radiosensitization was seen with IB (1.5 mM) for 2-h treatment at pH 6.7 before irradiation. The dose-modifying factor as defined by the ratio of radiation doses required to achieve the same effect on cell survival was 1.8 at 10% survival level. In contrast, SC-236 (50 microM for 2 8 h) showed no pH-dependent cytotoxicity. There was modest increase in the cell killing at lower doses of radiation. CONCLUSION: An acidic pH was an important factor affecting the increased cytotoxicity and radiosensitization by ibuprofen. Radiation response was enhanced at shoulder portion of the cell survival curve by selective COX-2 inhibitor. PMID- 12128135 TI - Retinoic acid modulates the radiosensitivity of head-and-neck squamous carcinoma cells grown in collagen gel. AB - PURPOSE: Collagen gels are increasingly regarded as reliable scaffolds for studying cells in vitro, displaying the same three-dimensional network of collagen fibers as encountered in vivo. As a contribution to therapeutic control of head-and-neck cancer, we grew HSCO86 cells in collagen gel and assessed their behavior in the presence of retinoic acid (RA) and radiation. METHODS AND MATERIALS: The malignant epithelial cell line HSCO86 was isolated from a postirradiation human oropharyngeal squamous carcinoma; it was EGFR-negative by immunocytochemical criteria. The cells were embedded in hydrated collagen I at a density of 10(6) cells/mL, and on Days 8, 10, and 12 of culture, they were treated with 10(-5) M retinoic acid. Radiation was administered using two different schedules: simultaneously with RA in three daily doses totaling 10 Gy, or with a single dose of 8 Gy on Day 29 of culture, after the effects of RA had taken place. Cell proliferation was evaluated by the MTT assay, whereas morphometric characteristics were detected in the cultured gels directly or in the gels after they were fixed and stained with hematoxylin. RESULTS: Contrary to growth in monolayer, where HSCO86 cells displayed a high proliferation rate, in collagen gel only a tiny fraction of the cells, usually less than 0.02%, survived the environmental stress; these cells spontaneously organized themselves into clonal multicellular spheroids growing up to 0.8 mm in diameter. After exposure to 10(-5) M retinoic acid, cell proliferation first declined and then, about 15 days after treatment, it started to increase to a level far above that in the control group. This surge in proliferation was ascribed to the appearance of numerous fibroblast-like cells at the edge of the spheroids. These cells, called HSCO-F, were the result of epithelial-to-mesenchymal conversion. When the gels were disaggregated by collagenase, and the cells were seeded in monolayer, HSCO-F cells reversed their morphology into parental HSCO86 cells. Treatment of collagen gels with 10 Gy, fractionated in three daily doses, did not substantially affect the growth of HSCO86 spheroids. However, when radiation was given simultaneously with RA, cell growth was significantly inhibited, both in terms of cell proliferation and size of spheroids (p < 0.0001 vs. untreated controls). This synergism applied mainly to parental HSCO86 cells, because no significant damage was induced by radiation on the HSCO-F cells previously generated by treatment with RA. CONCLUSION: Differences in the radiosensitivity of HSCO86 and HSCO-F cells are surprising in view of their common origin; this suggests a scenario in which, to overcome a microenvironmental stress, head-and-neck carcinoma cells can temporarily shift from an epithelial to a mesenchymal phenotype. In particular, morphologic and functional data suggested that HSCO-F cells were transformed into vascular endothelial cells whose characteristics included the following: (1) distinctive expression of Factor VIII and beta(1)-integrin, not detected in parental HSCO86 cells; (2) active migration in the collagen network by extruded pseudopodia, frequently appearing as colonies of filamentous cells aligned along the radial axis of the spheroids; and (3) efficient contraction of floating collagen gels. The implication of our study is that head-and-neck carcinomas may respond to RA treatment by selecting cell populations both resistant to radiation and capable of migrating inside the connective tissue, mimicking the behavior of vascular capillaries. PMID- 12128136 TI - Plan space: representation of treatment plans in multidimensional space. AB - PURPOSE: Treatment planning balances the need to provide adequate radiation coverage of the target with the need to reduce against the risk of overdosing normal tissues. An acceptable plan fulfills minimum dose-volume criteria for irradiation of tumor and normal tissues. However, multiple plans can satisfy these minimum criteria, and some plans provide for better protection of normal tissue than others. Here, we present a method to help the planner compare plans and decide whether a particular plan is the "best" plan on the basis of a set of certain dose-volume conditions. METHODS AND MATERIALS: Treatment plans are represented as points in multidimensional space. One dimension is assigned to the target and one to normal tissues of each anatomic structure under consideration. Minimum target dose is used as the target axis coordinate and the percentage volume of each normal structure receiving more than a specified dose as each normal-tissue coordinate. Images of plan space are developed for model phantom anatomy as well as for two clinical cases in the thorax and abdomen. RESULTS: When a sufficient number of plans have been plotted, a feasibility boundary becomes evident. This hypersurface in plan space represents the limit of the given treatment technique. By using a plan on this boundary, the benefits of a given treatment modality are maximized, providing assurance that the selected plan is the "best" plan. The beam angles and relative weights of a plan can be changed to alter its position in plan space, allowing improvements in an existing plan. Frequently, plans with normal-tissue dose distributions superior to the minimum acceptable criteria can be selected. The benefits of using plan-space images have been demonstrated at sites in the thorax and abdomen. CONCLUSION: Instead of defining dose-volume criteria at the outset, it is possible to select the best achievable plans by first evaluating the space of possible plans for a particular patient's unique anatomy and then choosing the plan with the optimum dose-volume characteristics. No attempt is made to arrive at a single plan score so that explicit judgments about the relative worth of individual structures are avoided. Visualization of plan-space images allows physicians to make choices based on their assessment of the relative significance of irradiation of each normal structure. PMID- 12128137 TI - Flat-panel cone-beam computed tomography for image-guided radiation therapy. AB - PURPOSE: Geometric uncertainties in the process of radiation planning and delivery constrain dose escalation and induce normal tissue complications. An imaging system has been developed to generate high-resolution, soft-tissue images of the patient at the time of treatment for the purpose of guiding therapy and reducing such uncertainties. The performance of the imaging system is evaluated and the application to image-guided radiation therapy is discussed. METHODS AND MATERIALS: A kilovoltage imaging system capable of radiography, fluoroscopy, and cone-beam computed tomography (CT) has been integrated with a medical linear accelerator. Kilovoltage X-rays are generated by a conventional X-ray tube mounted on a retractable arm at 90 degrees to the treatment source. A 41 x 41 cm(2) flat-panel X-ray detector is mounted opposite the kV tube. The entire imaging system operates under computer control, with a single application providing calibration, image acquisition, processing, and cone-beam CT reconstruction. Cone-beam CT imaging involves acquiring multiple kV radiographs as the gantry rotates through 360 degrees of rotation. A filtered back-projection algorithm is employed to reconstruct the volumetric images. Geometric nonidealities in the rotation of the gantry system are measured and corrected during reconstruction. Qualitative evaluation of imaging performance is performed using an anthropomorphic head phantom and a coronal contrast phantom. The influence of geometric nonidealities is examined. RESULTS: Images of the head phantom were acquired and illustrate the submillimeter spatial resolution that is achieved with the cone-beam approach. High-resolution sagittal and coronal views demonstrate nearly isotropic spatial resolution. Flex corrections on the order of 0.2 cm were required to compensate gravity-induced flex in the support arms of the source and detector, as well as slight axial movements of the entire gantry structure. Images reconstructed without flex correction suffered from loss of detail, misregistration, and streak artifacts. Reconstructions of the contrast phantom demonstrate the soft-tissue imaging capability of the system. A contrast of 47 Hounsfield units was easily detected in a 0.1-cm-thick reconstruction for an imaging exposure of 1.2 R (in-air, in absence of phantom). The comparison with a conventional CT scan of the phantom further demonstrates the spatial resolution advantages of the cone-beam CT approach. CONCLUSIONS: A kV cone-beam CT imaging system based on a large-area, flat-panel detector has been successfully adapted to a medical linear accelerator. The system is capable of producing images of soft tissue with excellent spatial resolution at acceptable imaging doses. Integration of this technology with the medical accelerator will result in an ideal platform for high-precision, image-guided radiation therapy. PMID- 12128138 TI - Open low-field magnetic resonance imaging in radiation therapy treatment planning. AB - PURPOSE: To evaluate the possibilities of an open low-field magnetic resonance imaging (MRI) scanner in external beam radiotherapy treatment (RT) planning. METHODS AND MATERIALS: A custom-made flat tabletop was constructed for the open MR, which was compatible with standard therapy positioning devices. To assess and correct image distortion in low-field MRI, a custom-made phantom was constructed and a software algorithm was developed. A total of 243 patients (43 patients with non-small-cell lung cancer, 155 patients with prostate cancer, and 45 patients with brain tumors) received low-field MR imaging in addition to computed tomographic (CT) planning imaging between January 1998 and September 2001 before the start of the irradiation. RESULTS: Open low-field MRI provided adequate images for RT planning in nearly 95% of the examined patients. The mean and the maximal distortions 15 cm around the isocenter were reduced from 2.5 mm to 0.9 mm and from 6.1 mm to 2.1 mm respectively. The MRI-assisted planning led to better discrimination of tumor extent in two-thirds of the patients and to an optimization in lung cancer RT planning in one-third of the patients. In prostate cancer planning, low-field MRI resulted in significant reduction (40%) of organ volume and clinical target volume (CTV) compared with CT and to a reduction of the mean percentage of rectal dose of 15%. In brain tumors, low-field MR image quality was superior compared with CT in 39/45 patients for planning purposes. CONCLUSIONS: The data presented here show that low-field MRI is feasible in RT treatment planning when image correction regarding system-induced distortions is performed and by selecting MR imaging protocol parameters with the emphasis on adequate images for RT planning. PMID- 12128139 TI - Treatment planning and dosimetry for the Harvard-MIT Phase I clinical trial of cranial neutron capture therapy. AB - PURPOSE: A Phase I trial of cranial neutron capture therapy (NCT) was conducted at Harvard-MIT. The trial was designed to determine maximum tolerated NCT radiation dose to normal brain. METHODS AND MATERIALS: Twenty-two patients with brain tumors were treated by infusion of boronophenylalanine-fructose (BPA-f) followed by exposure to epithermal neutrons. The study began with a prescribed biologically weighted dose of 8.8 RBE (relative biologic effectiveness) Gy, escalated in compounding 10% increments, and ended at 14.2 RBE Gy. BPA-f was infused at a dose 250-350 mg/kg body weight. Treatments were planned using MacNCTPlan and MCNP 4B. Irradiations were delivered as one, two, or three fields in one or two fractions. RESULTS: Peak biologically weighted normal tissue dose ranged from 8.7 to 16.4 RBE Gy. The average dose to brain ranged from 2.7 to 7.4 RBE Gy. Average tumor dose was estimated to range from 14.5 to 43.9 RBE Gy, with a mean of 25.7 RBE Gy. CONCLUSIONS: We have demonstrated that BPA-f-mediated NCT can be precisely planned and delivered in a carefully controlled manner. Subsequent clinical trials of boron neutron capture therapy at Harvard and MIT will be initiated with a new high-intensity, high-quality epithermal neutron beam. PMID- 12128140 TI - Dose distribution of narrow beam irradiation for small lung tumor. AB - PURPOSE: To aid in the selection of incident X-ray energy for stereotactic irradiation (STI) of lung tumor, dose distribution was investigated in a model of a thorax embedded with a tumor. METHODS AND MATERIALS: The dose distribution in a thorax model was calculated using the EGS4 Monte Carlo simulation; it was also measured with dosimetric film of a tentative thorax phantom. Uniformity of dose distribution in a tumor region was compared among the results of irradiation for several X-ray energies, and optimal X-ray energy for STI of a lung tumor was discussed. RESULTS: Dose distributions in the thorax were obtained. An increase in X-ray energy led not only to an increased dose delivered to the tumor, but also to an increased dose to surrounding normal lung tissue. CONCLUSIONS: The flat range in dose distribution along the beam axis and in the beam profiles of the tumor increases with decreasing X-ray energy. Consequently, lower energy, rather than higher energy, is recommended for STI of a lung tumor in terms of higher uniformity in the target volume. PMID- 12128141 TI - Usefulness of positron-emission tomographic images after proton therapy. AB - PURPOSE: To examine the positron emission tomography (PET) image obtained after proton irradiation and investigate the usefulness of the image for confirmation of the irradiated volume in proton radiotherapy (RT). METHODS AND MATERIALS: A homogenous phantom was irradiated separately by carbon-ion and proton beams and the images obtained were compared. The PET images of cancer patients just after proton RT were then taken after informed consent. RESULTS: In the PET image produced by carbon-ion beams, the high pixel counts in the image corresponded to the Bragg peak; however, in that produced by proton beams, they were visible throughout the entire track of the proton beams and were not related to the Bragg peak. The PET image of patients treated with proton RT was similar to that of the phantom experiment. CONCLUSION: The PET image after proton RT was different from that of carbon-ion RT. It was found that the PET image was very useful in proton RT to verify treatment planning. PMID- 12128142 TI - In regard to Brenner et al. Direct evidence that prostate tumors show high sensitivity to fractionation (low alpha/beta ratio) similar to late-responding normal tissue. PMID- 12128144 TI - In regard to Regine et al. Risk of symptomatic brain tumor recurrence and neurologic deficit after radiosurgery alone in patients with newly diagnosed brain metastases: results and implications. PMID- 12128146 TI - Prostate cancer: low alpha/beta the only consideration? In regard to Fowler, Chappell, and Ritter: the prospects for new treatments for prostate cancer. PMID- 12128148 TI - In regard to Antonadou et al. Randomized trial investigating the effects of daily amifostine in addition to radiation therapy in 146 patients with advanced lung cancers. PMID- 12128150 TI - Separate, parallel sensory and hedonic pathways in the mammalian somatosensory system. AB - We propose that separate sensory and hedonic representations exist in each of the primary structures of the somatosensory system, including brainstem, thalamic and cortical components. In the dorsal horn of the spinal cord, the hedonic representation, which consists primarily of nociceptive-specific, wide dynamic range, and thermoreceptive neurons, is located in laminae I and II, while the sensory representation, composed primarily by low-threshold and wide dynamic range neurons, is found in laminae III through V. A similar arrangement is found in the caudal spinal trigeminal nucleus. Based on the available anatomical and electrophysiological data, we then determine the corresponding hedonic and sensory representations in the area of the dorsal column nuclei, ventrobasal and posterior thalamic complex, and cortex. In rodent primary somatosensory cortex, a hedonic representation can be found in laminae Vb and VI. In carnivore and primate primary and secondary somatosensory cortical areas no hedonic representation exists, and the activities of neurons in both areas represent the sensory aspect exclusively. However, there is a hedonic representation in the posterior part of insular cortex, bordering on retroinsular cortex, that receives projections from two thalamic areas in which hedonics are represented. The functions of the segregated components of the system are discussed, especially in relation to the subjective awareness of pain. PMID- 12128151 TI - Place-learning, but not cue-learning training, modifies the hippocampal theta rhythm in rats. AB - Hippocampal theta activity accompanies behaviours such as running, swimming, head movements and spatially orientated responses in the rat. However, whether a relationship between this activity and information processing exists remains unclear. As place-learning depends on hippocampal integrity, whereas cue-learning does not, the hippocampal theta activity underlying each test was evaluated. Local CA1 hippocampal electroencephalograms (EEGs) were recorded over 6 days in a Morris maze for one place-learning test group of rats (n=10) and one cue-learning test group (n=8). The EEG of the corresponding test was taken during waking immobile, searching, and on-platform stages. The relative power (RP) values of EEGs were divided into 4-6.5Hz, 6.5-9.5Hz, and 9.5-12Hz frequency sub-bands. The place-learning training produced a separation of 4-6.5Hz and 6.5-9.5Hz sub-bands in searching and platform stages and a higher activity on the 6.5-9.5Hz sub-band during searching, compared with the basal record, while the cue-learning group did not show differences related either to the training or behavioural stage. As the motor activity and swimming velocity was similar in both groups, the results strongly suggest that the changes observed in theta activity reflect information processing by the hippocampus. PMID- 12128152 TI - Sphingomyelinase and ceramide analogs induce vasoconstriction and leukocyte endothelial interactions in cerebral venules in the intact rat brain: Insight into mechanisms and possible relation to brain injury and stroke. AB - This study was designed to test the hypothesis that the sphingomyelin-ceramide signaling pathway may be important in proinflammatory-like responses in the intact brain. Effects of neutral sphingomyelinase (N-SMase), ceramide analogs, phosphorylcholine and ceramide metabolites were studied on rat brain cerebral (cortical) venule lumen sizes, leukocyte rolling, velocity and endothelial cell wall adhesion, microvessel permeability, microvessel rupture and focal hemorrhages using in vivo high resolution TV microscopy. Perivascular and close intra-arterial administration of N-SMase, C(2)-, C(8)-, and C(16)-ceramide, but not either phosphorylcholine, C(6)-ceramide, nervonic (C(24):1) ceramide, lignoceric (C(24):0) ceramide, C(8)-ceramide-1-phosphate, glucosylceramide or 1-0 acylceramide, resulted in potent, concentration-dependent constriction (and spasm) of cortical venules, followed by increased leukocyte rolling, decreased leukocyte velocities, increased leukocyte-endothelial wall adhesion, increased venular wall permeability, postcapillary venule rupture and, often, micro hemorrhaging at high concentrations; angiotensin II, serotonin and PGF(2alpha) didn't demonstrate these characteristics. Pretreatment with either one of three different antioxidants, including inhibitors of NF-kappaB activation, or two different Ca(2+) channel blockers either prevented or attenuated the adverse venular effects of N-SMase and the ceramides. Likewise, pretreatment with either a PKCalpha-beta antagonist or a MAP kinase antagonist also attenuated the adverse venular effects. These results suggest that N-SMase and several ceramides can result in potent venular cerebrovasospasm, leukocyte-endothelial chemoattraction, and microvessel wall permeability changes in the intact rat brain. These proinflammatory-like actions suggest that N-SMase and ceramides could produce brain-vascular damage by reperfusion injury triggering lipid peroxidation, release of reactive oxygen species and activation of diverse signaling pathways: PKCalpha-beta isozymes, MAP kinase and NF-kappaB. PMID- 12128153 TI - Behavioral responses to social stress in noradrenaline transporter knockout mice: effects on social behavior and depression. AB - Noradrenaline has been implicated in the pathogenesis of depression and the noradrenaline transporter (NET) is a target for some antidepressants. Therefore, mice with disrupted NET gene expression (NET-KO) appear especially suitable for studying this behavioral disorder. We have examined the interaction between social stress (an etiological factor of depression) and the resulting depressive behaviors in NET-KO mice. Social stress was induced by daily defeats from larger resident mice while depression was assessed by the behavioral despair model. Animals subjected to repeated social stress showed reduced weight gain and a gradual shift from offensive to defensive behaviors. The latter may be considered a situation-specific depressive-like behavior. NET gene disruption did not prevent these changes that developed in a homotypic situation (i.e., during the repeated application of the same stressor). In contrast, stressed NET-KO mice showed more struggling in the behavioral despair model than stressed wild type (WT) animals. Thus, NET gene disruption inhibited depression-like behavior in chronically stressed animals tested in a situation heterotypic to the original cause of chronic stress. We suggest that the behavioral effects of NET gene disruption were overruled by experience and learning in the homotypic situation but manifested fully in the heterotypic situation. Tentatively, our data suggest that enhanced noradrenergic function does not prevent situation-specific social learning but impedes the generalization of depression to heterotypic circumstances. PMID- 12128154 TI - Cocaine modulates mu-opioid receptor mRNA but not c-fos mRNA levels in primary cortical astrocytes. AB - Cocaine is known to modulate the opioid system in several brain regions, including the cortex. Glial cells that are derived from the neonatal cortex have been shown to express opioid peptides and opioid receptors. In this study we investigated the effects of cocaine on c-fos and mu-opioid receptor mRNA levels in primary cortical astrocyte cultures, using RT-PCR and quantitative solution hybridization assays. Astrocyte cultures from 1-day-old Fischer rats were untreated or treated with cocaine for 30min, 2h, or 5h. While c-fos mRNA levels did not change at any time, mu-opioid receptor mRNA levels decreased by 75% after 2 and 5h of cocaine treatments. Our data suggest that cocaine differentially modulates c-fos and opioid signaling in astrocyte cell culture. PMID- 12128155 TI - Rate-independent inhibition of 5-HT release by 5-HT in the somadendritic regions of raphe neurons. AB - The somadendritic regions of raphe neurons respond to exogenous 5 hydroxytryptamine (5-HT) with an inhibition of spontaneous rate and a consequent reduction in local transmitter release, providing evidence for the operation of negative feedback regulation of spontaneous rate. Experiments were done to determine if a release process for 5-HT might also operate in the somadendritic regions that is independent of negative feedback and rate regulation. Slices of rabbit brain containing medullary or midbrain raphe nuclei, were stimulated in vitro at predetermined frequencies and the efflux of 3H-transmitter determined. The stimulation-induced pattern of transmitter release was independent of frequency, pointing to the absence of feedback. Further, exogenous 5-HT (1 x 10( 6)M) depressed the release of 3H-transmitter, but the inhibition, monitored over a range of frequencies, did not reflect competition with endogenous 5-HT for receptor sites. The antagonist methiothepin (3 x 10(-6)M) attenuated the inhibitions by 5-HT but did not by itself potentiate transmitter release, as expected if feedback inhibition were operative. Labeled transmitter release was antagonized by pretreatment with fluoxetine prior to 3H-HT incubation, and was severely curtailed in a calcium deficient medium, confirming that a neuronally relevant pool of transmitter was involved. It is concluded that serotonergic somadendritic sites contain inhibitory receptors for 5-HT release that operate independently of rate regulation and feedback. These findings could explain how other transmitters, and 5-HT itself (through dendritic release of transmitter), could exert synaptic effects on serotonergic and other neurons without being promptly countermanded by a somadendritic feedback-induced rate correction. PMID- 12128156 TI - Effects of cocaine on c-fos and preprodynorphin mRNA levels in intact and ovariectomized Fischer rats. AB - Psychostimulants such as cocaine have been shown to regulate c-fos and opioid gene expression in male rats. However, little information is available on cocaine effects in female rats or how the ovarian hormones, estrogen and progesterone, modulate these effects. In this study we used quantitative solution hybridization assays to measure c-fos and preprodynorphin (PDYN) mRNA levels after cocaine administration in the caudate/putamen of intact male and female rats or ovariectomized (OVX) female rats that were pretreated with vehicle, estrogen and/or progesterone. The c-fos mRNA levels were increased in intact male and female rats after 30min or 3h of one single cocaine injection and after 14 days of single daily cocaine injections. The c-fos mRNA levels were also increased after 30min of a single cocaine injection in OVX female rats that were treated with vehicle, estrogen and/or progesterone. The PDYN mRNA levels did not change after 30min, 3h or 14 days in intact male or female rats. However, PDYN mRNA levels were increased in the caudate/putamen of OVX female rats pretreated with vehicle or a combination of estrogen and progesterone but not in OVX female rats that were pretreated with either estrogen or progesterone alone. Our data suggest hormonal regulation of cocaine effects on PDYN mRNA levels which may modulate cocaine-induced behaviors in female rats. PMID- 12128157 TI - Contribution of N-methyl-d-aspartate receptors in the anteroventral third ventricular region to vasopressin secretion, but not to cardiovascular responses provoked by hyperosmolality and prostaglandin E2 in conscious rats. AB - The aim of this study is to pursue roles of N-methyl-d-aspartate (NMDA) receptors in the anteroventral third ventricular region (AV3V; a pivotal area for autonomic functions) in controlling vasopressin (AVP) release and cardiovascular system. In conscious rats, we examined effects of AV3V infusion of MK-801 (a selective antagonist for NMDA receptor) on plasma AVP, osmolality, electrolytes, arterial pressure and heart rate, in the absence or presence of NMDA, hyperosmotic or prostaglandin (PG) E2 stimulus. The AV3V infusion of NMDA caused significant increases in plasma AVP, osmolality and sodium, hematocrit, arterial pressure and heart rate after 5 or 15min. When NMDA was administered into the cerebral ventricle, relatively smaller elevations were observed only in plasma AVP and arterial pressure. The effects of AV3V infusion of NMDA were nearly completely prevented by MK-801 applied to the same region before 15min. The application of MK-801 was also potent to block rises of plasma AVP elicited by AV3V injection of PGE2 or i.v. infusion of hypertonic saline. However, it inhibited neither increases of arterial pressure and heart rate due to the PGE2 treatment nor those of arterial pressure, plasma osmolality and sodium in response to the osmotic load. Histological analysis on the AV3V infusion sites of NMDA, MK-801 and PGE2 indicated that they had been located in the structures such as the median and medial preoptic nuclei, periventricular nucleus and medial preoptic area. These results suggest that stimulation of AV3V NMDA receptors in the basal state may facilitate AVP secretion and cause pressor and tachycardiac actions, and that these receptors may be involved in both the hyperosmolality- and PGE2-induced hormone release, but not in the cardiovascular responses to these stimuli. PMID- 12128158 TI - Elevated plasma nociceptin level in patients with Wilson disease. AB - Plasma level of nociceptin, the endogenous agonist of orphanin FQ/ORL1 receptor was found to be significantly elevated in Wilson disease patients (13.98+/ 2.44pg/ml, p<0.001, n=20) compared to age-matched healthy controls (9.18+/ 1.63pg/ml, n=25). Wilson disease is an autosomal recessive disorder of copper metabolism caused by mutation of the gene ATP7B leading to toxic copper accumulation in the liver and other organs such as brain, kidney and cornea. Measurements were performed by 125I-radioimmunoassay. Neither sex differences nor correlation between plasma nociceptin levels and liver function test results were found. It is suggested that elevated plasma nociceptin level found in Wilson disease patients is due to inhibition of nociceptin-inactivating Zn metallopeptidases (aminopeptidase N (APN) and endopeptidase 24.15) by the toxic copper deposits in liver and/or brain. PMID- 12128159 TI - Environmental influences on functional outcome after a cortical infarct in the rat. AB - The effect of postoperative housing conditions on functional outcome and brain derived neurotrophic factor (BDNF) gene expression was evaluated 1 month after a distal ligation of the right middle cerebral artery (MCA) in spontaneously hypertensive rats. Two days postoperatively the rats were randomized into four groups; individually housed with no equipment (deprived group), individually housed with free access to a connected running wheel (running group), housed together in a large cage with no equipment (social group) or in the same size of cage furnished with bars, chains and various things to manipulate (enriched group). The enriched rats had significantly higher scores when crossing a rotating horizontal rod than deprived and running rats. The social group performed significantly better than the deprived group. The BDNF gene expression in the ipsi- and contralateral cortex, thalamus, hippocampus and cerebellum did not significantly differ between the groups. The weight of the adrenal glands was significantly increased in running rats suggesting that postischemic running may be stressful. We conclude that the beneficial effect of postischemic environmental enrichment is likely to be a combination of social and various physical activities, and that BDNF gene expression 1 month after a cortical infarct did not correlate with functional outcome. PMID- 12128160 TI - Septal cholinergic mediation of hippocampal theta in the cat. AB - The effects of intraseptally microinjected muscarinic (atropine sulfate, pirenzepine and gallamine) and nicotinic (hexamethonium) antagonists on spontaneous, sensory and electrically-induced hippocampal (HPC) theta EEG activity were investigated in the freely behaving cat. Administration of hexamethonium failed to elicit a detectable effect on HPC theta. Injections of atropine and pirenzepine abolished, whereas the injection of gallamine only reduced hippocampal theta. Moreover, a gradual recovery of theta amplitude and power was observed, while frequency recovered rapidly. Our data provide further evidence that the septal M1 and M2 muscarinic receptor subtypes mediate the HPC theta in this species. Intraseptal microinjection of cholinergic agonist (carbachol) produced almost a continuous HPC theta with increased amplitude and power. The contribution of the medial septal cholinergic projections to HPC theta frequency and amplitude was also discussed. PMID- 12128161 TI - Electrophysiological properties of the rat area postrema neurons displaying both the transient outward current and the hyperpolarization-activated inward current. AB - We found coexistence of the transient outward potassium current (I(TO)) and the hyperpolarization-activated inward current (I(H)) in 26 of 82 area postrema neurons tested using the whole-cell patch-clamp technique in rat brain slices. Cells displaying both the I(TO) and the I(H) typically showed "voltage sag" and "rebound potentials" in response to hyperpolarizing current injection and repetitive firing with strong adaptation was seen with depolarizing current injection. When cells were held at membrane potentials more negative than the resting level (e.g., -85mV), the afterhyperpolarization was enhanced. Voltage clamp recordings were performed to examine the characteristics of I(TO) and I(H) in and the contributions of these currents to the electroresponsiveness of area postrema cells. We show, in this study, the voltage-dependent properties of I(H) and I(TO), and how these currents modulate the intrinsic membrane properties of area postrema cells. We discuss the functional significance of the specific subset of area postrema neurons whose cells have both I(H) and I(TO) channels. PMID- 12128162 TI - Effects of droplet size and type of binder on the agglomerate growth mechanisms by melt agglomeration in a fluidised bed. AB - This study was performed in order to evaluate the effects of binder droplet size and type of binder on the agglomerate growth mechanisms by melt agglomeration in a fluidised bed granulator. Lactose monohydrate was agglomerated with melted polyethylene glycol (PEG) 3000 or Gelucire 50/13 (esters of polyethylene glycol and glycerol), which was atomised at different nozzle air flow rates giving rise to median droplet sizes of 40, 60, and 80 microm. Different product temperatures were investigated, below the melting range, in the middle of the melting range, and above the melting range for each binder. The agglomerates were found to be formed by initial nucleation of lactose particles immersed in the melted binder droplets. Agglomerate growth occurred by coalescence between nuclei followed by coalescence between agglomerates. Complex effects of binder droplet size and type of binder were seen at low product temperatures. Low product temperatures resulted in smaller agglomerate sizes, because the agglomerate growth was counteracted by very high binder viscosity or solidification of the binder. At higher product temperatures, neither the binder droplet size nor the type of binder had a clear effect on the final agglomerate size. PMID- 12128163 TI - Physico-chemical determinants of dermal drug delivery: effects of the number and substitution pattern of polar groups. AB - The aim of this study was to investigate the effect of number and substitution pattern of -OH groups of a set of phenols on the in vitro permeation of heat separated human epidermis. The diffusion was calculated from Log(D/x)=logk(p) 0.59logK(oct)+0.024 (D, diffusion coefficient; x, pathlength; k(p), permeability coefficient (cm/h); and K(oct), octanol-water partition coefficient). The main factors reducing D were the dipolar and hydrogen bonding capabilities of the permeants quantified as their Hansen partial solubility parameters delta(p) and delta(h). These parameters are significantly reduced by the degree of symmetry of the molecule, so that phloroglucinol (1,3,5-benzenetriol), with three -OH groups, diffuses more rapidly that phenol. When symmetry is absent, as in 1,2,4 benzenetriol, the number of -OH groups results in very slow diffusion. D/x (cm/h) was related to the combined solubility parameter delta(a) defined as radical(delta(p)(2)+delta(h)(2)) by: (D/x)=0.0024-0.000065delta(a) (n=7, R(2)=0.70, P=0.012). PMID- 12128164 TI - Determination of transport in the Caco-2 cell assay of compounds varying in lipophilicity using LC-MS: enhanced transport of Leu-enkephalin analogues. AB - PURPOSE: To synthesize a number of analogues of Leu-enkephalin with different lipophilicities and to develop an LC-MS method for determining the Caco-2 cell permeability values of these compounds. METHODS: A number of sugar and sugar plus lipoamino acid analogues of Leu-enkephalin were synthesized by solid-phase and solution methods. An LC-MS method was developed for analyzing the Caco-2 cell assay samples and validated against the traditional method using radiolabelled compounds. RESULTS: A sensitive and specific LC-MS assay was developed. Standard curves were linear in the range of 0.025-5 microM. Apparent permeability values determined by LC-MS and liquid scintillation counter were identical, for both a hydrophilic drug, cephalexin and a lipophilic Leu-enkaphalin analogue. Caco-2 permeability values for the analogues of Leu-enkephalin were determined and it was found that attachment of sugar or sugar and lipoamino acid to the Leu enkephalin peptide resulted in an increase in the apparent permeability values compared to the native peptide, which was not transported across the Caco-2 cell monolayers. CONCLUSIONS: A rapid, generic LC-MS method for analyzing a range of compounds was developed. Attachment of a sugar or sugar and lipoamino acid to Leu enkephalin improves the apparent permeability across Caco-2 cell monolayers. PMID- 12128165 TI - Altered drug disposition of the platelet activating factor antagonist apafant in mdr1a knockout mice. AB - The aim of the present study was to determine a potential impact of P glycoprotein (P-gp) on the tissue distribution and disposition of apafant (WEB 2086, CAS 105219-56-5), a selective platelet-activating factor antagonist, and on digoxin in mdr1a(-/-) and wildtype mice. Transport experiments in Caco-2 monolayers at low concentrations (<10 microM) showed that the secretory flux of [(14)C]apafant and [(3)H]digoxin exceeded the absorptive flux nine times. This efflux was concentration dependent and subject to inhibition by the P-gp substrates verapamil and cyclosporin A. This indicates that active drug transporter P-gp was involved in apafant and digoxin absorption. Mdr1a(-/-) mice showed a more than 70-fold higher concentration of digoxin-related radioactivity (P<0.001) in the brain than wildtype mice after intravenous doses of 0.05 mg/kg [(3)H]digoxin. Differences were less pronounced in other tissues. Both liquid scintillation counting and whole body autoradiography yielded comparable results and they also matched recently published data. Apafant-related radioactivity was about ten-fold higher in the brain of mdr1a(-/-) mice compared to wildtype mice following intravenous doses of 2 mg/kg radiolabelled apafant. Only slight or negligible differences were observed in other tissues. In wildtype mice, intestinal excretion of [(14)C]apafant (54.9%) exceeded biliary excretion (26.5%). However, in mdr1a(-/-) mice biliary excretion (50.7%) exceeded intestinal excretion (6.8%). These differences were mirrored in the urinary and faecal excretion. Pharmacokinetic parameters of apafant and radioactivity did not differ between wildtype and mdr1a(-/-) mice. The conclusions were: (1) apafant and digoxin are P-gp substrates, and (2) absence of mdr1a encoded P-gp significantly alters tissue distribution (especially in brain) and excretion routes (biliary and intestinal) of apafant. PMID- 12128166 TI - Anticonvulsant activity of hydrazones, Schiff and Mannich bases of isatin derivatives. AB - In the present study, anticonvulsant activity of hydrazones, Schiff and Mannich bases of isatin were evaluated by maximal electroshock method (MES) and metrazol induced convulsions (MET) at 30, 100 and 300 mg/kg dose levels. Neurotoxicity of the compounds was also assessed at the same dose levels. Eight compounds of the series exhibited significant anticonvulsant activity at 30 mg/kg dose level. 3-(4 chloro-phenylimino)-5-methyl-1,3-dihydro-indol-2-one (compound 10) was found to be the most potent compound of the series with 87% protection at 100 mg/kg and an ED(50) of 53.61 mg/kg (MET). All the compounds exhibited lesser neurotoxicity compared to phenytoin. All the active compounds showed greater protection than sodium valproate. The essential structural features responsible for interaction with receptor site are established within a suggested pharmacophore. PMID- 12128167 TI - Influence of the infusion rate on disposition of netilmicin in the isolated rat perfused kidney. AB - A study of the disposition of netilmicin in the isolated rat kidney was carried out in order to establish the influence of the infusion rate on the drug profile in this tissue. A dose of 800 microg administered as a bolus injection or at infusion times of 5, 7.5 and 10 min, respectively, was injected through the afferent cannula into the isolated kidney. Analysis of outflow curves was carried out using different kinetic approaches. Comparison of statistical moments and derived parameters pointed to changes in the distribution process with the infusion rate. In contrast, elimination remained constant, since the extraction coefficient and relative area under the curve values did not change with the infusion rate, although the MTT (mean transit time) and distribution volume decreased for the longest infusion times. The UDF (unit disposition function) profiles were not superimposed for the different infusion rates and combined with the results of the kinetic analysis revealed that the behaviour of netilmicin in the isolated kidney depends on infusion rate. The apparent partition coefficients in renal cortex and medulla showed higher values for the slower perfusion rates. Yet, a progressive decrease in the absolute amount of netilmicin was predicted in the tubular epithelium compartment whereas the residence time tended to increase. The latter phenomenon could account for the higher aminoglycoside nephrotoxicity reported when these drugs are administered over longer infusion times. PMID- 12128168 TI - Quantum chemical descriptors in the formulation of pectin pellets produced by extrusion/spheronisation. AB - The objective of this study was to employ quantitative structure-activity relationships (QSAR) to relate calculated molecular descriptors of granulation liquid additives to improvements in the size of pectin pellets produced by extrusion/spheronisation. Quantum chemical descriptors were calculated for a large number of candidate additives. Based on a principal component analysis (PCA) of descriptors of the candidates, a few substances were selected. The most suitable concentration for each additive was found, and pellets were prepared by an extrusion/spheronisation process. Three pectin grades of different methoxy and amide substitution were tested and the quality of the pellets was evaluated based on size. PLS models were constructed to identify the molecular properties that were most important in producing short, nearly spherical pellets. The results show that quantum chemical descriptors can be a useful tool in the formulation of pectin pellets. Acceptable models relating additive properties and pellet size were achieved. Independent of the pectin grade, the two most important factors favouring formation of small spherical pellets were a small molecular size and a strong hydrogen bond forming ability of the additive molecules. PMID- 12128169 TI - Nicotine permeability across the buccal TR146 cell culture model and porcine buccal mucosa in vitro: effect of pH and concentration. AB - The present study was conducted to investigate and compare the effect of pH and drug concentration on nicotine permeability across the TR146 cell culture model and porcine buccal mucosa in vitro. As a further characterization of the TR146 cell culture model, it was explored whether the results were comparable for bi directional and uni-directional transport in the presence of a transmembrane pH gradient. Nicotine concentrations between 10(-5) and 10(-2) M were applied to the apical side of the TR146 cell culture model or the mucosal side of porcine buccal mucosa. Buffers with pH values of 5.5, 7.4 and 8.1 were used to obtain different fractions of non- and mono-ionized nicotine. The apparent permeability (P(app)) of nicotine across both models increased significantly with increasing pH, and the P(app) values obtained with the two models could be correlated in a linear manner. With increasing concentrations of nicotine, the P(app) values decreased, which can partly be explained by an effect on the paracellular pathway. Similar results were also obtained when using the models for bi-directional as well as for uni-directional studies. The TR146 cell culture model may be used as model for buccal epithelium in studies with ionized drugs and a transmembrane pH gradient. PMID- 12128170 TI - Effects of 12 Ca2+ antagonists on multidrug resistance, MDR1-mediated transport and MDR1 mRNA expression. AB - The effects of 12 Ca(2+) antagonists on MDR1 were examined by two independent models: the inhibitory effect on MDR1-mediated transport of [(3)H]digoxin using MDR1-overexpressing LLC-GA5-COL150 cell monolayers and the reversal effect on cytotoxicity of vinblastine or paclitaxel using MDR1-overexpressing Hvr100-6 cells. The inhibitory effects on [(3)H]digoxin transport were assessed as the 50% inhibitory concentration during 4 h exposure, and the values were the lowest for nicardipine (4.54 microM), manidipine (4.65 microM) and benidipine (4.96 microM), followed by bepridil (10.6 microM), barnidipine (12.6 microM), efonidipine (13.0 microM), verapamil (13.2 microM) and nilvadipine (18.0 microM). The reversal effect on cytotoxicity was assessed by the 50% growth inhibitory concentration after 3 days exposure, and the resistance to vinblastine or paclitaxel in Hvr100 6 cells was reversed by manidipine, verapamil, benidipine, barnidipine, and nicardipine, in that order. Bepridil, barnidipine, efonidipine, verapamil and nilvadipine showed similar inhibitory effects on [(3)H]digoxin transport, but barnidipine and verapamil showed a stronger effect in reversal of cytotoxicity. Real-time quantitative RT-PCR assay indicated a decrease in MDR1 mRNA expression by barnidipine and verapamil. It is concluded that Ca(2+) antagonists cannot only be direct inhibitors of MDR1 but that some may at the same time act as inhibitors of expression of MDR1 via down-regulation of MDR1 mRNA. PMID- 12128171 TI - Synthesis and in vitro chemical and enzymatic stability of glycosyl 3'-azido-3' deoxythymidine derivatives as potential anti-HIV agents. AB - New glycosyl derivatives of 3'-azido-3'-deoxythymidine (AZT) (1 and 2) were synthesized in order to improve AZT retention in the blood and to guarantee its sustained release, overcoming the necessity of multiple drug administrations. The esters synthesized (1 and 2) link AZT, by a succinyl linker, to the C-3 position of glucose and to C-6 of galactose. Furthermore, the chemical and enzymatic stabilities of esters 1 and 2 were evaluated in order to determine both their stability in aqueous medium and their feasibility to undergo enzymatic cleavage by esterase to regenerate the original drug. The pharmacokinetic profiles of esters 1 and 2, obtained after systemic administration, showed an interesting controlled release, in particular for ester 2, compared to the pharmacokinetic profile of AZT. PMID- 12128172 TI - Characterization of azithromycin hydrates. AB - Azithromycin (AZI) is a macrolide antibiotic with an expanded spectrum of activity that is commercially available as a dihydrate. This study was carried out to characterize hydrates of azithromycin. A commercial dihydrate sample was used to prepare monohydrate from water/ethanol (1:1) mixture. Hydrates were characterized using DSC, TGA, KFT, XRD, HSM, SEM and FT-IR. TGA showed that the commercial samples are dihydrate and the sample prepared from water/ethanol (1:1) was a monohydrate. Solubility studies revealed that monohydrate converted to dihydrate during solubility studies and as a result there was no significant difference in the equilibrium solubility of MH and DH. Thermal analysis under various conditions revealed that dehydration and melting took place simultaneously. Anhydrous AZI was found to be hygroscopic and converted to DH on storing at room temperature. Molecular modeling studies revealed the probable sites of attachment of water molecules to AZI. PMID- 12128173 TI - In vitro drug release studies on guar gum-based colon targeted oral drug delivery systems of 5-fluorouracil. AB - Intravenous administration of 5-fluorouracil for colon cancer therapy produces severe systemic side-effects due to its cytotoxic effect on normal cells. The broad objective of the present study was to develop novel tablet formulations for site-specific delivery of 5-fluorouracil to the colon without the drug being released in the stomach or small intestine using guar gum as a carrier. Fast disintegrating 5-fluorouracil core tablets were compression coated with 60% (FHV 60), 70% (FHV-70) and 80% (FHV-80) of guar gum, and were subjected to in vitro drug release studies. The amount of 5-fluorouracil released from the compression coated tablets in the dissolution medium at different time intervals was estimated by a HPLC method. Guar gum compression-coated tablets released only 2.5 4% of the 5-fluorouracil in simulated GI fluids. When the dissolution study was continued in simulated colonic fluids (4% w/v rat caecal content medium) the compression-coated FHV-60, FHV-70 and FHV-80 tablets released another 70, 55 and 41% of the 5-fluorouracil respectively. The results of the study show that compression-coated tablets containing 80% (FHV-80) of guar gum are most likely to provide targeting of 5-fluorouracil for local action in the colon, since they released only 2.38% of the drug in the physiological environment of the stomach and small intestine. The FHV-80 formulation showed no change either in physical appearance, drug content or dissolution pattern after storage at 40 degrees C/RH 75% for 6 months. The differential scanning calorimetric study showed that 5 fluorouracil did not interact with the formulation excipients used in the study. PMID- 12128174 TI - Comparative evaluation of rate of hydration and matrix erosion of HEC and HPC and study of drug release from their matrices. AB - Hydrophilic polymers, in contact with the dissolution medium, may swell and make a continuous gel layer, erode or undergo combination of the two. The swelling action of these polymers is controlled by the rate of their hydration in the dissolution medium. The extent of polymer swelling, relative mobilities of dissolution medium and drug, and matrix erosion dictate the kinetics as well as mechanism of drug release from the polymeric matrices. The objective of the present investigations was to study the rate of hydration and the rate of matrix erosion of two hydrophilic, non-ionic cellulose ethers, i.e., hydroxyethylcellulose (HEC) and hydroxypropylcellulose (HPC), and to compare the kinetics and mechanism of drug release from their matrices. Chlorpheniramine maleate was used as the model drug. Matrix tablets containing chlorpheniramine maleate, HEC or HPC and dicalcium phosphate were compressed at 156 MPa pressure. The rate of hydration of the polymer, rate of erosion of the matrices and in vitro drug release studies were carried out in phosphate buffer (pH 7.4). The hydration studies of the two polymers demonstrated that due to relatively larger water uptake, the degree of swelling of HEC matrices was considerably higher as compared to the HPC matrices. Also, HEC matrices exhibited relatively higher erosion as compared to HPC matrices. The drug release from HEC matrices occurred by non-Fickian transport, i.e., combination of drug diffusion and polymer swelling, while drug release from HPC matrices was controlled primarily by diffusion through pores and channels in the structure. The t(50%), time to reach 50% drug release, for HEC matrices was 4.8 h and that for HPC matrices was 6.5 h which indicates that a higher polymer level was needed in the case of HEC matrices to sustain the drug release for up to 12 h of dissolution as compared to HPC matrices due to relatively higher hydrophilicity of HEC. PMID- 12128175 TI - Differential interaction of Sophora isoflavonoids with lipid bilayers. AB - The mechanisms of some biological effects exerted by flavonoids (e.g. activity against lipid oxidation, multidrug resistance modulation) may involve their interactions with lipid bilayers. Due to variety of substituents attached to the flavonoid nucleus individual isoflavones significantly differ in their properties; in particular they may differently interact with membranes. For this reason we have investigated the interactions of different isoflavones with lipid bilayers. The influence of four plant isoflavones on the phase transitions of dipalmitoylphosphatidylcholine (DPPC) and on liposome aggregation was studied, using microcalorimetry and absorption measurements, respectively. We found that isoflavones substituted with one or two prenyl groups less effectively induce liposome aggregation than more polar ones, possessing no prenyl groups. For aggregation-promoting compounds, rather small differences in the influence on phosphatidylcholine, phosphatidylserine and phosphatidylinositol liposomes were recorded. On the other hand, the alteration of DPPC phase transitions by prenyl substituted isoflavones was more pronounced than changes induced by non-prenyl ones. On the basis of observed effects we conclude that prenyl-substituted isoflavones penetrate deeper into the lipid bilayer while more polar ones act closer to the membrane surface. Comparing our results with biological tests it seems that interactions with the hydrophobic core of membranes are responsible for the activity of the studied isoflavones. PMID- 12128176 TI - Determination of the mechanical properties of pellets and film coated pellets using dynamic mechanical analysis (DMA). AB - The elastic and viscoelastic properties of pellets and film coatings applied to pellets were studied using Dynamic Mechanical Analysis (DMA). Four batches of spherical pellets produced with different methods were compared without film coating and after application of a plasticised ethylcellulose film of average thickness of 25 microm. The modulus of elasticity of the uncoated pellets was found to be related to the porosity of the pellets, as was the slope of the linear portion of the storage modulus-dynamic force curves. None of the other parameters obtained from DMA of the uncoated pellets appeared to be related to the macroscopic properties of these pellets. After application of the film coating the dynamic force at the onset of the linear portion of the storage modulus-dynamic force curve and the maximum storage modulus obtained at a dynamic force of 600 mN were found to be significantly different from the values of the uncoated pellets, but similar for all four coated pellet batches. Hence these parameters represented the viscoelastic behaviour of the film coatings. PMID- 12128177 TI - On the role of the macromolecular phase transitions in biology in response to change in solution volume or macromolecular composition: action as an entropy buffer. AB - We have used numerical simulation to demonstrate the potential for macromolecular precipitate solution phase transitions existing within the cell, to play a role in the minimization of changes in location or quaternary state of other macromolecular components, predicted to accompany changes in cell volume. For our modeling we have employed thermodynamic relations that take into account the large effects upon the thermodynamic activity coefficient produced by a solution environment that is highly volume occupied due to the presence of high concentrations of soluble macromolecule. The theoretical approach adopted, along with the simulated results, provide a framework for the interpretation of certain proteins' behavior (e.g. cytoskeletal elements such as tubulin and actin and possibly some prion structures) in response to cell volume change. PMID- 12128179 TI - Interaction of sodium dodecyl sulfate with human native and cross-linked hemoglobins: a transient kinetic study. AB - The interaction of sodium dodecyl sulfate (SDS) at a concentration range (0-515 microM) below the critical micelle concentration (CMC approximately 0.83 mM) with human native and cross-linked oxyhemoglobin (oxyHb) and methemoglobin (metHb) has been investigated by optical spectroscopy and stopped-flow transient kinetic measurements. It is observed that the interaction of SDS with human native and cross-linked oxyHb shows the disappearance of the bands of oxyHb at 541 and 576 nm and the appearance at 537 nm. The resultant spectra are characteristic of low spin (Fe(3+)) hemichrome. Similarly SDS has been found to convert human native and cross-linked high spin (Fe(3+)) metHb to low spin (Fe(3+)) hemichrome. The interaction of SDS with oxyHb suggests a conformational change of the protein in the heme pocket, which may induce the binding of distal histidine to iron leading to the formation of superoxide radical. The formation of hemichrome from metHb is found to be concentration-dependent with SDS. The stopped flow transient kinetic measurements of the interaction of SDS with metHb show that at least four molecules of SDS interact with one molecule of metHb. The interaction of SDS with human cross-linked oxy and met hemoglobin shows results similar to those for human native oxy and met hemoglobin indicating that the covalent modification does not alter the interaction of SDS with cross-linked hemoglobin. PMID- 12128178 TI - Hydrophobic forces between protein molecules in aqueous solutions of concentrated electrolyte. AB - Protein-protein interactions have been measured for a mutant (D101F) lysozyme and for native lysozyme in concentrated solutions of ammonium sulfate at pH 7 and sodium chloride at pH 4.5. In the mutant lysozyme, a surface aspartate residue has been replaced with a hydrophobic phenylalanine residue. The protein-protein interactions of D101F lysozyme are more attractive than those of native lysozyme for all conditions studied. The salt-induced attraction is correlated with a solvation potential of mean force given by the work required to desolvate the part of the protein surfaces that is buried by the protein-protein interaction. This work is proportional to the aqueous surface-tension increment of the salt and the fractional non-polar surface coverage of the protein. Experimental measurements of osmotic second virial coefficients validate a proposed potential of mean force that ascribes the salt-induced attraction between protein molecules to an enhancement of the hydrophobic attraction. This model provides a first approximation for predicting the protein-protein potential of mean force in concentrated aqueous electrolyte solutions; this potential is useful for determining solution conditions favorable for protein crystallization. PMID- 12128180 TI - The alterations of lipid bilayer fluidity induced by newly synthesized phenothiazine derivative. AB - Using fluorescence spectroscopy, calorimetry and ESR the interactions of the phenothiazine derivative 2-trifluoromethyl-10-(4-[methylsulfonylamid]buthyl) phenothiazine (FPhMS) with lipids were studied. Calorimetry showed biphasic effect of FPhMS on main phase transition of DPPC. At molar ratios up to 0.06 drug induced decrease of transition temperature and enthalpy, while at higher concentrations it caused subsequent increase of these parameters. For all concentrations studied we observed gradual broadening of transition peaks. Fluorescence polarization revealed that in FPhMS/lipid mixtures, order in bilayers is decreased in the gel state and increased in the liquid crystalline state. ESR experiment showed that at molar ratio of 0.06, FPhMS reduces the mobility of spin probes located in both polar and hydrophobic regions. Comparing observed effects with those reported for cholesterol/lipid mixtures, we conclude that at higher concentrations FPhMS presumably induces a new mode of bilayer packing. This structure is less co-operative than an unperturbed bilayer, but locally the mobility of lipid molecules is decreased. PMID- 12128181 TI - Molecular modeling of wild-type and antifolate resistant mutant Plasmodium falciparum DHFR. AB - The development of drug resistance is reducing the efficiency of antifolates as antimalarials. This phenomenon has been linked to the occurrence of mutations in the parasite's dihydrofolate reductase (DHFR). In this way, the resistance to pyrimethamine and cycloguanil, two potent inhibitors of P. falciparum DHFR, is mainly related to mutations (single and crossed) at residues 16, 51, 59, 108 and 164 of the enzyme. In this work, we have refined a recently proposed homology model of P. falciparum DHFR, and the resulting structure was used to obtain models for 14 mutant enzymes, employing molecular modeling. Ternary complexes of the mutant enzymes with these inhibitors have been superimposed to equivalent ternary complexes of the wild-type enzyme, allowing the proposition of hypotheses for the role of each mutation in drug resistance. Based on these results, possible reasons for antifolate resistance have been proposed. PMID- 12128182 TI - The conformation of dehydroalanine in short homopeptides: molecular dynamics simulations of a 6-residue chain. AB - A molecular dynamics study about the conformational preferences in a chloroform solution of a homo-oligomer constituted by six residues of dehydroalanine is presented. For this purpose, two sets of force-field parameters and explicit solvent molecules have been used. Furthermore, ab initio calculations have been performed in order to estimate 1[H]-NMR chemical shifts. Results have been compared with experimental data. PMID- 12128183 TI - Studies on surfactant-biopolymer interaction. I. Microcalorimetric investigation on the interaction of cetyltrimethylammonium bromide (CTAB) and sodium dodecylsulfate (SDS) with gelatin (Gn), lysozyme (Lz) and deoxyribonucleic acid (DNA). AB - The interaction of the surfactants cetyltrimethyl ammonium bromide (CTAB) and sodium dodecyl sulfate (SDS) with the biopolymers gelatin (Gn), lysozyme (Lz) and deoxyribonucleic acid (DNA) was studied by isothermal titration microcalorimetry at varied biopolymer concentration, pH and temperature. The nature of interaction of the surfactants with the biopolymers was assessed from the observed enthalpy [surfactant] profiles. The biopolymer-induced aggregation of the surfactants was observed. The enthalpies of aggregation of amphiphiles, binding of aggregates with macromolecules, organisational change of bound aggregates, and threshold concentrations for micelle formation of surfactants in the presence of biopolymers were estimated. The results collected on the three biopolymers were analysed and compared. PMID- 12128184 TI - Special issue in honour of Maurizio Brunori on the occasion of his 65th birthday. PMID- 12128185 TI - Focal contributions to molecular biophysics and structural biology: a personal view. Part III. PMID- 12128186 TI - Recombinant hemoglobins with low oxygen affinity and high cooperativity. AB - By introducing an additional H-bond in the alpha(1)beta(2) subunit interface or altering the charge properties of the amino acid residues in the alpha(1)beta(1) subunit interface of the hemoglobin molecule, we have designed and expressed recombinant hemoglobins (rHbs) with low oxygen affinity and high cooperativity. Oxygen-binding measurements of these rHbs under various experimental conditions show interesting properties in response to pH (Bohr effect) and allosteric effectors. Proton nuclear magnetic resonance studies show that these rHbs can switch from the oxy (or CO) quaternary structure (R) to the deoxy quaternary structure (T) without changing their ligation states upon addition of an allosteric effector, inositol hexaphosphate, and/or reduction of the ambient temperature. These results indicate that if we can provide extra stability to the T state of the hemoglobin molecule without perturbing its R state, we can produce hemoglobins with low oxygen affinity and high cooperativity. Some of these rHbs are also quite stable against autoxidation compared to many of the known abnormal hemoglobins with altered oxygen affinity and cooperativity. These results have provided new insights into the structure-function relationship in hemoglobin. PMID- 12128187 TI - Intramolecular electron transfer in cytochrome cd(1) nitrite reductase from Pseudomonas stutzeri; kinetics and thermodynamics. AB - Cytochrome cd(1) nitrite reductase from Pseudomonas stutzeri catalyzes the one electron reduction of nitrite to nitric oxide. It is a homodimer, each monomer containing one heme-c and one heme-d(1), the former being the electron uptake site while the latter is the nitrite reduction site. Hence, internal electron transfer between these sites is an inherent element in the catalytic cycle of this enzyme. We have investigated the internal electron transfer reaction employing pulse radiolytically produced N-methyl nicotinamide radicals as reductant which reacts solely with the heme-c in an essentially diffusion controlled process. Following this initial step, the reduction equivalent is equilibrating between the c and d(1) heme sites in a unimolecular process (k=23 s(-1), 298 K, pH 7.0) and an equilibrium constant of 1.0. The temperature dependence of this internal electron transfer process has been determined over a 277-313 K temperature range and yielded both equilibrium standard enthalpy and entropy changes as well as activation parameters of the specific rate constants. The significance of these parameters obtained at low degree of reduction of the enzyme is discussed and compared with earlier studies on cd(1) nitrite reductases from other sources. PMID- 12128188 TI - Hydration, slaving and protein function. AB - Protein dynamics is crucial for protein function. Proteins in living systems are not isolated, but operate in networks and in a carefully regulated environment. Understanding the external control of protein dynamics is consequently important. Hydration and solvent viscosity are among the salient properties of the environment. Dehydrated proteins and proteins in a rigid environment do not function properly. It is consequently important to understand the effect of hydration and solvent viscosity in detail. We discuss experiments that separate the two effects. These experiments have predominantly been performed with wild type horse and sperm whale myoglobin, using the binding of carbon monoxide over a broad range of temperatures as a tool. The experiments demonstrate that data taken only in the physiological temperature range are not sufficient to understand the effect of hydration and solvent on protein relaxation and function. While the actual data come from myoglobin, it is expected that the results apply to most or all globular proteins. PMID- 12128189 TI - Heterotropic effectors control the hemoglobin function by interacting with its T and R states--a new view on the principle of allostery. AB - Careful analyses of precise oxygenation curves of hemoglobin (Hb) clearly indicate that, contrary to the common belief, allosteric effectors exert a dramatic control of the oxygenation characteristics of the protein by binding not only to the T (unligated), but also to the R (ligated) state, in a process that is proton-driven and involves proton uptake. The most striking functional changes were obtained when the allosteric effectors were bound to the fully ligated Hb: the oxygen affinity decreased dramatically, Bohr effect was enhanced, and cooperativity of oxygen ligation was almost absent, emulating a Root effect-like behavior. However, structural analysis, such as Cys beta 93 sulfhydryl reactivity and ultraviolet circular dichroism, confirmed that the ligated Hb was in fact in the R state, despite its extremely low affinity state features. These findings provide a new global view for allosteric interactions and invoke for a modern interpretation of the role of allosteric effectors and a reformulation of the Monod-Wyman-Changeaux model for control of allosteric systems, and other complementary models as well. PMID- 12128190 TI - Production and characterisation of Met80X mutants of yeast iso-1-cytochrome c: spectral, photochemical and binding studies on the ferrous derivatives. AB - The iron ligand, Met80, of yeast iso-1-cytochrome c has been mutated to residues that are unable to bind to the iron. The resultant proteins, Met80Ala, Ser, Asp, Glu, have been expressed and purified. All mutant proteins exhibit well defined pH dependent spectral transitions that report the binding, at high pH, of an intrinsic ligand (probably the nitrogen of an epsilon-NH(2) of a lysine) that drives the heme low-spin. The pK values are mutant dependent. All the mutant proteins bind extrinsic ligands, such as CO, in their ferrous states and we report the apparent quantum yield (phi) for CO photo-dissociation. The values of phi range from 0.004 for Met80Ala to 0.04 for Met80Asp. We also report values for the rate constant for binding the intrinsic lysine residue. The values for this constant, for phi and for the pK values are discussed in terms of the rigidity of the cytochrome structure. We also show that the mutant proteins bind with high affinity to cytochrome c oxidase, both in the ferric and ferrous states. The potential of these proteins to act as light activated electron donors for the study of electron transfer is discussed. PMID- 12128191 TI - Description of hemoglobin oxygenation under universal solution conditions by a global allostery model with a single adjustable parameter. AB - The Monod-Wyman-Changeux allosteric model parameters evaluated from accurate oxygen equilibrium curves (OECs) of hemoglobin that were measured in an extremely wide range of structural constraints, imposed by allosteric effectors, yielded a closed circle when log K(T) and log K(R) were plotted against log L(0) and log L(4), respectively, showing novel phenomena that L(0) and L(4) have a maximal value and a minimal value, respectively, and K(T) and K(R) vary by more than three orders of magnitude. These phenomena were successfully described by a global allostery model, which mathematically keeps the frame work of the MWC model, but allows that K(T) under a set of solution conditions becomes larger than K(R) under another set of solution conditions and postulates that a representative allosteric effector binds to both the T and R states with a lower affinity but with a larger stoichiometry for the R state than for the T state. Thus, this global model can describe any given OEC measured under universal solution conditions with the single adjustable parameter, the concentration of the representative effector. PMID- 12128192 TI - Effects of inert volume-excluding macromolecules on protein fiber formation. I. Equilibrium models. AB - The equilibrium Oosawa-Asakura model for nucleated assembly of rod-like protein fibers is recast in terms of dimensionless (scaled) quantities. The model is then generalized to treat arbitrarily large deviations from thermodynamic ideality arising from high fractional volume occupancy by an inert protein or polymer. Each state of association of the self-associating protein is modeled as an equivalent rigid convex particle (sphere or spherocylinder) and the crowding species is modeled either as an equivalent sphere or cylindrical rod. The resulting conservation of mass relation is readily solved to yield the fractional abundance of monomer, from which the entire equilibrium distribution of oligomeric species can be calculated, either directly or through the use of an additional scaling relationship. Results indicating the potential effect of volume occupancy on the equilibrium solubility of the self-associating protein and upon the equilibrium distribution of polymer size are presented. It is found that the fractional (logarithmic) change in both solubility and in the breadth of the polymer size distribution scale almost linearly with the fractional (logarithmic) change in the thermodynamic activity of monomer. PMID- 12128193 TI - Relocation of an internal proton donor in cytochrome c oxidase results in an altered pK(a) and a non-integer pumping stoichiometry. AB - Cytochrome c oxidase from Rhodobacter sphaeroides has two proton-input pathways leading from the protein surface towards the catalytic site, located within the membrane-spanning part of the enzyme. One of these pathways, the D-pathway, contains a highly conserved Glu residue [E(I-286)], which plays an important role in proton transfer through the pathway. In a recent study, we showed that a mutant enzyme in which E(I-286) was re-located to the opposite side of the D pathway [EA(I-286)/IE(I-112) double mutant enzyme] was able to pump protons, although with a stoichiometry that was lower than that of the wild-type enzyme (approximately 0.6 H(+)/e(-)) (Aagaard et al. (2000) Biochemistry 39, 15847 15850). These results showed that the residue must not necessarily be located at a specific place in the amino-acid sequence, but rather at a specific location in space. In this study, we have investigated the effect of moving E(I-286) on the kinetics of specific reaction steps of the catalytic cycle in the pH range 6-11. Our results show that during the reaction of the four-electron reduced enzyme with O(2), the rates of the two first transitions (up to formation of the 'peroxy' intermediate, P(r)) are the same for the double mutant as for the wild type enzyme, but formation of the oxo-ferryl (F) and fully oxidized (O) states, associated with proton uptake from the bulk solution, are slowed by factors of approximately 30 and approximately 400, respectively. Thus, in spite of the dramatically reduced transition rates, the proton-pumping stoichiometry is reduced only by approximately 40%. The apparent pK(a) values in the pH dependencies of the rates of the P(R)-->F and F-->O transitions were >3 and approximately 2 units lower than those of the corresponding transitions in the wild-type enzyme, respectively. The relation between the modified pK(a)s, the transition rates between oxygen intermediates and the pumping stoichiometry is discussed. PMID- 12128194 TI - Introduction of a new regulatory mechanism into human hemoglobin. AB - Previous studies on bovine hemoglobin (HbBv) have suggested amino acid substitutions, which might introduce into human hemoglobin (HbA) functional characteristics of HbBv, namely a low intrinsic oxygen affinity regulated by Cl( ). Accordingly, we have constructed and characterized a multiple mutant, PB5, [beta(V1M + H2 Delta + T4I + P5A + A76K)] replacing four amino acid residues of HbA with those present at structurally analogous positions in HbBv, plus an additional substitution, beta T4I, which does not occur in either HbBv or HbA. This 'pseudobovine' hemoglobin has oxygen binding properties very similar to those of HbBv: the P(50) of HbA, PB5 and HbBv in the absence of Cl(-) are 1.6, 4.6 and 4.8 torr, respectively, and in 100 mM Cl(-) are 3.7, 10.5 and 12 torr, respectively. Moreover, PB5 has 3-fold slower autoxidation rate compared to HbA and HbBv. These are desirable characteristics for a human hemoglobin to be considered for use as a clinical artificial oxygen carrier. Although the functional properties of PB5 and HbBv are similar, van't Hoff plots indicate that the two hemoglobins interact differently with water, suggesting that factors regulating the R to T equilibrium are not the same in the two proteins. A further indication that PB5 is not a functional mimic of HbBv derives from PB5(control), a human hemoglobin with the same substitutions as PB5, except the beta T4I replacement. PB5(control) has a high oxygen affinity (P(50)=2.3 torr) in the absence of Cl(-), but retains the Cl(-) effect of PB5. The Cl(-) regulation of oxygen affinity in PB5 involves lysine residues at beta 8 and beta 76. PB4, which has the same substitutions as PB5 except beta A76K, and PB6, which has all the substitutions of PB5 plus beta K8Q, both have a low intrinsic oxygen affinity, like HbBv and PB5, but exhibit a decreased sensitivity to Cl(-). Since HbBv has lysine residues at both beta 8 and beta 76, these results imply that Cl(-) regulation in HbBv likewise involves these two residues. The mechanism responsible for the low intrinsic oxygen affinity of HbBv remains unclear. It is suggested that residues peculiar to HbBv at the alpha(1)beta(1) interface may play a role. PMID- 12128195 TI - Myoglobin as a model system for designing heme protein based blood substitutes. AB - The ligand binding properties and resistances to denaturation of >300 different site-directed mutants of sperm whale, pig, and human myoglobin have been examined over the past 15 years. This library of recombinant proteins has been used to derive chemical mechanisms for ligand binding and to examine the factors governing holo- and apoglobin stability. We have also examined the effects of mutagenesis on the dioxygenation of NO by MbO(2) to form NO(3)(-) and metMb. This reaction rapidly detoxifies NO and is a key physiological function of both myoglobins and hemoglobins. The mechanisms derived for O(2) binding and NO dioxygenation have been used to design safer, more efficient, and more stable heme protein-prototypes for use as O(2) delivery pharmaceuticals in transfusion therapy (i.e. blood substitutes). An interactive database is being developed (http://olsonnt1.bioc.rice.edu/web/myoglobinhome.asp) to allow rapid access to the ligand binding parameters, stability properties, and crystal structures of the entire set of recombinant myoglobins. The long-range goal is to use this library for developing general protein engineering principles and for designing individual heme proteins for specific pharmacological and industrial uses. PMID- 12128196 TI - A tertiary two-state allosteric model for hemoglobin. AB - The two-state allosteric model of Monod, Wyman, and Changeux (MWC) provides an excellent description of homotropic effects in a vast array of equilibrium and kinetic measurements on cooperative ligand binding by hemoglobin. However, in contrast to experimental observations, the model does not allow for alteration of the ligand affinity of the T quaternary structure by allosteric effectors. This failure to explain heterotropic effects has been appreciated for over 30 years, and it has been generally assumed to result from tertiary conformational changes in the absence of a quaternary change. Here we explore a model that preserves the essential MWC idea that binding without a quaternary conformational change is non cooperative, but where tertiary conformations of individual subunits play the primary role instead of the quaternary conformations. In this model, which we call the 'tertiary two-state (TTS) model', the two affinity states correspond to two tertiary conformations of individual subunits rather than the two quaternary conformations of the MWC two-state allosteric model. Ligation and the R quaternary structure bias the subunit population toward the high affinity tertiary conformation, while deligation and the T quaternary structure favor the low affinity tertiary conformation. We show that the model is successful in quantitatively explaining a demanding set of kinetic data from nanosecond carbon monoxide photodissociation experiments at times longer than approximately 1 micros. Better agreement between the model and the submicrosecond kinetic data may result from detailed considerations of the distribution and dynamics of conformational substates of the two tertiary conformations. The model is consistent with the results of solution, gel, and single crystal oxygen binding studies, but underestimates the population of doubly-liganded molecules determined in low-temperature electrophoresis experiments. PMID- 12128197 TI - Responses of normal and sickle cell hemoglobin to S-nitroscysteine: implications for therapeutic applications of NO in treatment of sickle cell disease. AB - Factors which govern transnitrosation reactions between hemoglobin (Hb) and low molecular weight thiols may define the extent to which S-nitrosated Hb (SNO-Hb) plays a role in NO in the control of blood pressure and other NO-dependent reactions. We show that exposure to S-nitrosylated cysteine (CysNO) produces equivalent levels of SNO-Hb for Hb A(0) and sickle cell Hb (Hb S), although these proteins differ significantly in the electron affinity of their heme groups as measured by their anaerobic redox potentials. Dolphin Hb, a cooperative Hb with a redox potential like that of Hb S, produces less SNO-Hb, indicating that steric considerations outweigh effects of altered electron affinity at the active-site heme groups in control of SNO-Hb formation. Examination of oxygen binding at 5-20 mM heme concentrations revealed increases due to S-nitrosation in the apparent oxygen affinity of both Hb A(0) and Hb S, similar to increases seen at lower heme concentrations. As observed at lower heme levels, deoxygenation is not sufficient to trigger release of NO from SNO-Hb. A sharp increase in apparent oxygen affinity occurs for unmodified Hb S at concentrations above 12.5 mM, its minimum gelling concentration. This affinity increase still occurs in 30 and 60% S nitrosated samples, but at higher heme concentration. This oxygen binding behavior is accompanied by decreased gel formation of the deoxygenated protein. S nitrosation is thus shown to have an effect similar to that reported for other SH group modifications of Hb S, in which R-state stabilization opposes Hb S aggregation. PMID- 12128198 TI - NO rebinding to myoglobin: a reactive molecular dynamics study. AB - The rebinding of NO to myoglobin after photolysis is studied using the 'reactive molecular dynamics' method. In this approach the energy of the system is evaluated on two potential energy surfaces that include the heme-ligand interactions which change between liganded and unliganded myoglobin. This makes it possible to take into account in a simple way, the high dimensionality of the transition seam connecting the reactant and product states. The dynamics of the dissociated NO molecules are examined, and the geometrical and energetic properties of the transition seam are studied. Analysis of the frequency of recrossing shows that the height of the effective rebinding barrier is dependent on the time after photodissociation. This effect is due mainly to protein relaxation and may contribute to the experimentally observed non-exponential rebinding rate of NO, as has been suggested previously. PMID- 12128199 TI - The interplay between heme iron and protein sulfhydryls in the reaction of dimeric Scapharca inaequivalvis hemoglobin with nitric oxide. AB - The homodimeric hemoglobin from the mollusk Scapharca inaequivalvis possesses a single reactive cysteine residue per monomer, Cys92, which is located in the subunit interface in the vicinity of the heme group. The interplay between the heme iron and Cys92 towards the reaction with NO has been investigated by the combined use of electrospray mass spectrometry, FTIR and UV-Visible spectroscopy. When the ferrous liganded or unliganded protein reacts with free NO in solution Cys92 is not modified, but undergoes nitrosation when the hemoglobin is exposed to the nitric oxide releaser S-nitrosocysteine. When the ferric protein reacts with free NO under anaerobic conditions the heme iron is reduced and Cys92 is nitrosated. At variance with other hemeproteins investigated to date, in Scapharca HbI the heme-iron NO driven reduction is not accompanied by the formation of a ferric iron nitrosyl intermediate in detectable amounts. The results are consistent with the hypothesis that the nitrosating agent is the NO(+) species, which is generated during the NO driven reduction of the ferric heme iron. The possible reaction mechanism is discussed in comparison with recent findings on human hemoglobin and myoglobin. PMID- 12128200 TI - Changes in the abnormal alpha-subunit upon CO-binding to the normal beta-subunit of Hb M Boston: resonance Raman, EPR and CD study. AB - Heme-heme interaction in Hb M Boston (His alpha 58-->Tyr) was investigated with visible and UV resonance Raman (RR), EPR, and CD spectroscopies. Although Hb M Boston has been believed to be frozen in the T quaternary state, oxygen binding exhibited appreciable co-operativity (n=1.4) and the near-UV CD spectrum indicated weakening of the T marker at pH 9.0. Binding of CO to the normal beta subunit gave no change in the EPR and visible Raman spectra of the abnormal alpha subunit at pH 7.5, but it caused an increase of EPR rhombicity and significant changes in the Raman coordination markers as well as the Fe(III)-tyrosine related bands of the alpha-subunit at pH 9.0. The UVRR spectra indicated appreciable changes of Trp but not of Tyr upon CO binding to the alpha-subunit at pH 9.0. Therefore, we conclude that the ligand binding to the beta heme induces quaternary structure change at pH 9.0 and is communicated to the alpha heme, presumably through His beta 92-->Trp beta 37-->His alpha 87. PMID- 12128201 TI - Requirements of high levels of Hedgehog signaling activity for medial-region cell fate determination in Drosophila legs: identification of pxb, a putative Hedgehog signaling attenuator gene repressed along the anterior-posterior compartment boundary. AB - We show that high levels of Hedgehog signaling activity are essential for medial region patterning in Drosophila legs. In mid-to-late third instar leg discs, high levels of Hedgehog signals repress the transcription of pxb, a newly identified gene encoding a transmembrane protein expressed specifically in the anterior compartment. Misexpression experiments indicate that Pxb may serve as a Hedgehog signaling attenuator capable of acting prior to Hedgehog-Patched interactions, suggesting that Hedgehog signaling in leg discs includes a pxb-repression mediated positive feedback loop. RNA interference and clonal analysis show that neither Wingless nor Decapentaplegic signaling is required for pxb repression but high levels of Wingless signaling activity are essential for patterning in the leg ventral medial region. PMID- 12128202 TI - Initiation of dorso-ventral axis during chick limb development. AB - We analysed spatio-temporal expression of dorso-ventral genes - Wnt-7a, En-1, Lmx 1 and Fgf-8 - during both normal and ectopic limb formation following fibroblast growth factor (FGF) application to the flank. We confirm that Wnt-7a is the first of these genes to be expressed in dorsal ectoderm in limb-forming regions. We also noticed patterns and kinetics of gene expression specific to chick that could account for differences observed in ridge formation between chick and mouse. We find that Wnt-7a expression, in dorsal ectoderm, is rapidly and locally induced by FGF application. In contrast, ectopic induction of Lmx-1 expression, in dorsal mesoderm, is much slower, occurs first at a distance from the FGF-2 bead and seems initially independent of direct Wnt-7a signalling during FGF-2 limb induction. Finally, we show that there is no contribution to extra-limb mesoderm from normal limb mesoderm and confirm that flank cells give rise to the extra limb. Furthermore, we suggest that an inhibitor present in the flank normally prevents Lmx-1 expression in this region and restricts its expression to limb-forming regions. PMID- 12128203 TI - Depletion of FGF acts as a lateral inhibitory factor in lung branching morphogenesis in vitro. AB - Previous studies have shown that the interaction of positive and inhibitory signals plays a crucial role during lung branching morphogenesis. We found that in mesenchyme-free conditions, the lung epithelium exerted a lateral inhibitory effect on the neighbouring epithelium via depletion of fibroblast growth factor 1 (FGF1). Contrary to previous suggestions, bone morphogenetic protein 4 could not substitute for the inhibitory effect. Based on of this observation, we used a reaction-diffusion model of the substrate-depletion type to represent the initial phase of in vitro branching morphogenesis of lung epithelium, with depletion of FGF playing the role of lateral inhibitor. The model was able to account for the effects of the FGF1 concentration, extracellular matrix degradation and different subtypes of FGF on morphogenesis of the lung bud epithelia. These results suggest that the depletion of FGF may be a key regulatory component in initial phase of branching morphogenesis of the lung bud epithelium in vitro. PMID- 12128205 TI - Na,K-ATPase alpha and beta subunit genes exhibit unique expression patterns during zebrafish embryogenesis. AB - We have used in situ hybridization to analyze Na,K-ATPase alpha and beta subunit gene expression during zebrafish embryogenesis. The most striking finding is that each of the 14 Na,K-ATPase genes exhibits a distinct expression profile. All alpha and beta subunit genes are expressed in the nervous system, although the pattern of expression in different regions varies dramatically. In peripheral tissues, three of the five alpha1-like genes are expressed in pronephros and mucous cells, one is expressed in heart, and one is predominant in skeletal muscle. The alpha2 gene is expressed in brain and heart but is most prominent in skeletal muscle, while the two alpha3 genes are restricted in their expression to the nervous system. Of the six beta subunit genes, beta1a is expressed at highest abundance in lens, pronephros, and heart, while beta1b transcripts are abundant in mucous cells. The two beta2-like genes are differentially expressed in the nervous system. One beta3 gene is expressed exclusively in brain while the other is abundantly expressed in skeletal muscle. Based on these expression patterns, we predict that at least 14 alpha/beta subunit pairs are likely to be formed in different tissues. PMID- 12128204 TI - FGF10 is a mesenchymally derived stimulator for epidermal development in the chick embryonic skin. AB - The development of avian cutaneous appendages, feathers and scales, is known to arise from the epithelial-mesenchymal interaction. Here we show that FGF10 is associated with this developmental process as an early signal from mesenchymal cells underlying nascent cutaneous placodes. Expression of Fgf10 was detected in the mesenchymal cells underneath the developing placodes. Forced expression of Fgf10 in the femoral skin suppressed expression of Shh and a zinc finger gene snail-related (cSnR), while induced expression of Bmp2 in the interbud region, resulting in thickening of the epidermal layer. Furthermore, forced expression of Fgf10 in the foot skin caused marked ingrowings of the epidermis. The cells in the epidermal ingrowings expressed beta-catenin, proliferating cell nuclear antigen, and an epidermal stem cell marker p63. These results support the idea that FGF10 is a mesenchymally derived stimulator of epidermal development through crosstalk with bone morphogenetic protein (BMP), beta-catenin, and other signaling pathways. PMID- 12128207 TI - Tramtrack69 is required for the early repression of tailless expression. AB - During embryogenesis, the activated Torso receptor tyrosine kinase (TOR RTK) pathway activates tailless (tll) expression by a relief-of-repression mechanism. Various lines of evidence have suggested that multiple factors are required for this repression. We show that Tramtrack69 (TTK69) binds to two sites within tll cis-regulatory DNA, TC2 and TC5, and that TTK69 is phosphorylated by mitogen activated protein kinase. In embryos lacking maternal ttk69 activity, the expression of both endogenous tll and lacZ driven by the tll minimal regulatory region (tll-MRR) are expanded. Further, in wild-type embryos, the tll-MRR mutated in TC5 drives uniform lacZ expression before late stage 4. We conclude that TTK69 is required for early (before the end of stage 4) repression of tll transcription. PMID- 12128206 TI - L-Maf, a downstream target of Pax6, is essential for chick lens development. AB - During lens development in vertebrates, the orchestration of multiple transcriptional regulators is essential for fate determination and terminal differentiation. In early development, Pax6, Sox2 and Six3 are expressed in the head ectoderm, while L-maf, Prox1 and crystallin genes are expressed at a later stage in the lens placode in a more restricted fashion. To uncover the genetic interactions among these factors during lens development, we examined the effects of dominant-negative molecules of Pax6 and L-Maf, which play decisive roles in lens formation. The two dominant-negative isoforms of Pax6 repress L-maf, Prox1 and delta-crystallin expression, resulting in failure of lens formation. These effects of dominant-negative Pax6 are fully rescued by co-expression with wild type L-Maf. In addition, dominant-negative L-Maf inhibits the expression of Prox1 and delta-crystallin, while misexpression of L-Maf causes ectopic induction of these genes in a Sox-2-dependent fashion. Our results demonstrate that L-Maf is a downstream target of Pax6 and mediates Pax6 activity in developing lens cells. PMID- 12128208 TI - Chick homeobox gene cbx and its role in retinal development. AB - Homeobox genes play important roles in animal development. We isolated a chick homeobox gene, cbx, and studied its function during embryonic development. The deduced Cbx protein contained 376 amino acid residues. Its homeodomain was related (with 65-71% sequence identity) to that of human Crx, human Cart-1, and chick Alx-4. On searching the human genome sequence, a human homologue was found, which had 78% overall sequence identity and a 100% identical homeodomain. In the developing chick retina, cbx was expressed in a small fraction of post-mitotic cells residing at anatomical locations typical of bipolar cells. These cells were Goalpha(+) and protein kinase C(-), suggesting that they were probably cone bipolar cells. cbx mRNA was also detected outside the retina, particularly in the tectum and Rathke's pouch. Replication-competent retrovirus was used to drive misexpression of cbx and of an Engrailed repression construct. Engrailed-mediated repression of Cbx was embryonic lethal, while misexpression of cbx itself was tolerated. In the retina, misexpression of cbx resulted in fewer PKC(+) bipolar cells. Our data suggest that cbx is essential for embryonic survival and may participate in the development of bipolar, probably cone bipolar, cells in the retina. PMID- 12128209 TI - Some distinctive features of zebrafish myogenesis based on unexpected distributions of the muscle cytoskeletal proteins actin, myosin, desmin, alpha actinin, troponin and titin. AB - The current myofibrillogenesis model is based mostly on in vitro cell cultures and on avian and mammalian embryos in situ. We followed the expression of actin, myosin, desmin, alpha-actinin, titin, and troponin using immunofluorescence microscopy of zebrafish (Danio rerio) embryos. We could see young mononucleated myoblasts with sharp striations. The striations were positive for all the sarcomeric proteins. Desmin distribution during muscle maturation changes from dispersed aggregates to a perinuclear concentration to striated afterwards. We could not observe desmin-positive, myofibrillar-proteins-negative cells, and we could not find any non-striated distribution of sarcomeric proteins, such as stress fiber-like structures. Some steps, like fusion before striation, seem to be different in the zebrafish when compared with the previously described myogenesis sequences. PMID- 12128210 TI - Nk6, a novel Drosophila homeobox gene regulated by vnd. AB - Nk(x)-type homeobox genes are an evolutionarily conserved family that regulate diverse developmental processes. Here we describe a novel Drosophila gene, Nk6, which encodes an Nk-type transcription factor most homologous to vertebrate Nkx6.1 and Nkx6.2. The homeodomains and NK decapeptide domains of all three proteins are highly conserved. Nk6 is expressed in the embryonic brain, ventral nerve cord, hindgut, and internal head structures. Nerve cord expression is in midline precursors, several ventral and intermediate column neuroblasts, and later in neurons but not glia, similar to the known expression of Nkx6 genes in the neural tube. We show genetically that Nk6 is positively regulated, directly or indirectly, by vnd in brain precursors. In vnd mutants, head neuroectoderm Nk6 expression is abolished where it is normally co-expressed with vnd. Conversely, vnd-overexpression leads to ectopic Nk6 expression in the brain. These findings further highlight the importance of interactions between Nk(x)-type genes in regulating their expression. PMID- 12128211 TI - Loss of the beta-catenin homologue aardvark causes ectopic stalk formation in Dictyostelium. AB - Aardvark (Aar) is a Dictyostelium beta-catenin homologue with both cytoskeletal and signal transduction roles during development. Here, we show that loss of aar causes a novel phenotype where multiple stalks appear during late development. Ectopic stalks are preceded by misexpression of the stalk marker ST-lacZ in the surrounding tissue. This process does not involve the kinase GSK-3. Mixing experiments show that ectopic ST-lacZ expression and stalk formation are cell non autonomous. The protein-cellulose matrix surrounding the stalk of aar mutant fruiting bodies is defective, and damage to the stalk of wild-type fruiting bodies leads to ectopic ST-lacZ expression. We postulate that poor synthesis of the stalk tube matrix allows diffusion of a stalk cell-inducing factor into the surrounding tissue. PMID- 12128212 TI - SH3 domain-mediated binding of the Drk protein to Dos is an important step in signaling of Drosophila receptor tyrosine kinases. AB - Activation of the Sevenless (Sev) receptor tyrosine kinase (RTK) in the developing Drosophila eye is required for the specification of the R7 photoreceptor cell fate. Daughter of Sevenless (Dos), a putative multi-site adaptor protein, is a substrate of the Sev kinase and is known to associate with the tyrosine phosphatase Corkscrew (Csw). Binding of Csw to Dos depends on the Csw Src homology 2 (SH2) domains and is an essential step for signaling by the Sev RTK. Dos, however, lacks a recognizable phosphotyrosine interaction domain and it was previously unclear how it is recruited to the Sev receptor. Here it is shown that the SH2/SH3 domain adaptor protein Drk can provide this link. Drk binds with its SH2 domain to the autophosphorylated Sev receptor while the C terminal SH3 domain is able to associate with Dos. The Drk SH3 domain binding motifs on Dos were mapped to two sites which do not conform the known Drk SH3 domain binding motif (PxxPxR) but instead have the consensus PxxxRxxKP. Mutational analysis in vitro and in vivo provided evidence that both Drk binding sites fulfil an important function in the context of Sev and Drosophila epidermal growth factor receptor mediated signaling processes. PMID- 12128213 TI - Expression of a vas::EGFP transgene in primordial germ cells of the zebrafish. AB - In zebrafish, maternally produced vasa (vas) transcripts become targeted to the cleavage planes of early embryos and subsequently incorporated into the primordial germ cells (PGCs). Zygotic vas transcription occurs from the onset of gastrulation. Here, we report on the characterisation of the zebrafish vas locus. The gene consists of 27 exons, spans about 25kb, and contains two CpG-rich regions. We have used vas regulatory regions to establish transgenic zebrafish lines expressing enhanced green fluorescent protein (EGFP) in their PGCs. Maternally encoded vas::EGFP transcripts and VAS::EGFP protein segregate with the PGCs during embryogenesis. We find that the maternally deposited vas::EGFP transcripts are stable during embryogensis at least up to 50h of development. Vas::EGFP transcripts could not be detected in embryos that inherit the transgene from males, most likely due to the lack of one or more regulatory elements required for early zygotic expression. We show that vas::EGFP transcripts become enriched to the cleavage planes in early embryos, a finding that supported an RNA localisation signal localised within the vas region of these transcripts. PMID- 12128214 TI - CRP2 transcript expression pattern in embryonic chick limb. AB - Members of the cysteine-rich protein (CRP) family are evolutionary conserved proteins that have been implicated in the processes of cell proliferation and differentiation via the cytoskeletal proteins. In this paper, we present the dynamic expression pattern of CPR2 transcripts during chick limb bud development. CRP2 transcripts are located in various tissues, including muscle, arteries, cartilage, ligaments and digit tendons and also in the apical ectodermal ridge and feather buds. PMID- 12128215 TI - Developmental expression of the T-box transcription factor T-bet/Tbx21 during mouse embryogenesis. AB - A novel type of DNA-binding domain, the 'T-box' domain, characterizes an increasingly large family of transcription factors (Trends Genet. 15 (1999) 154). We have identified and characterized the expression pattern of a new member of the Tbr1 subfamily of T-box genes; this gene has been recently named T-bet/Tbx21 (Genomics 70 (2000) 41; Cell 17 (2000) 655; Science 292 (2001) 1907; Science 295 (2002) 338). The sequence and expression of Tbr1 and eomesodermin/Tbr2 are closely related to T-bet/Tbx21. The expression of Tbr1 (Neuron 15 (1995) 63) and Tbr2 (Mech Dev 84 (1999) 133) have virtually identical onset, at around E10.5, and expression domains in the mouse telencephalon. While Tbr1 is expressed in postmitotic neurons, Tbr2 (which is also expressed during gastrulation is also expressed in neural progenitors. We have used in situ hybridization to determine the temporal and spatial distribution of T-bet/Tbx21 expression during mouse development. T-bet/Tbx21 expression is exclusively restricted to the olfactory bulb and the thymus. To assess the distribution of T-BET/TBX21 expression in the haematopoietic compartment we used reverse transcriptase-polymerase chain reaction and found its expression in several human blood cell lineages, including progenitors/stem cells, immature B cells and peripheral T cells. PMID- 12128216 TI - Expression of Notch genes and their ligands during gastrulation in the chicken embryo. AB - Notch signalling is an important evolutionary conserved mechanism known to control cell fate choices through local interactions. Here, the patterns of expression of Notch-1 and -2 genes and their ligands Delta-1, Serrate-1 and -2, were established in the early blastodisc of the chicken embryo from the pre streak to the first somite stages. Delta-1 was detected as early as the pre streak stage in the posterior part of the embryo shortly followed in the same region by Notch-1 at the initial streak stage. Thereafter both were strongly expressed in the posterior part of the primitive streak until HH4. Notch-2 was also found at the level of the streak although at low levels. Notch-1 was homogeneously expressed by the epiblast and by mesodermal cells ingressing at the level of the streak whereas Delta-1 expression formed a 'salt and pepper' pattern. The difference between the two was clearly detected by double in situ hybridisation. From the mid-streak to the first somite stages, Notch-1 and Delta 1 expressions appeared in the anterior part of the embryo. Serrate-1 and -2 were not detected at these stages. Taken together, these results constitute a framework for analysing the role(s) for Notch signalling during gastrulation. PMID- 12128217 TI - Ets2 is expressed during morphogenesis of the somite and limb in the mouse embryo. AB - Ets2 is a member of the ETS family of transcription factors. In order to address the developmental function of Ets2, we have examined its expression pattern in E8.5 to E13.5 embryos using RNA whole-mount in situ hybridization. In the paraxial mesoderm, Ets2 is expressed uniformly in the presomitic mesoderm and then restricted to the cells in the rostral portion of the segmenting and the next two recently formed somites. In the developing limb, Ets2 expression in the mesenchyme reflects the progressive formation of the hand or foot plate and the digital skeleton. In addition, Ets2 is expressed in the otic vesicle and its derivatives, the dorsal (posterior) root ganglia, the neuroepithelium in the dorsal part of the caudal neural tube and the inter-segmental vasculature. PMID- 12128218 TI - Isthmin is a novel secreted protein expressed as part of the Fgf-8 synexpression group in the Xenopus midbrain-hindbrain organizer. AB - Patterning of the central nervous system is regulated by a signaling center located at the midbrain-hindbrain boundary (MHB), or isthmus organizer. Fibroblast growth factors secreted from the MHB are required and sufficient to direct the ordered growth and regionalization of the midbrain and anterior hindbrain. In an unbiased secretion cloning screen of Xenopus gastrula embryos we identified a novel gene, which we designated as Isthmin (xIsm) due to its prominent expression at the MHB. xIsm encodes a secreted protein of 449 amino acids containing one copy of the thrombospondin type 1 repeat (TSR). We also found orthologous Isthmin genes in human (hIsm) and mouse (mIsm), as well as a gene encoding an Isthmin-like human unknown protein (hIsm-l). The conservation of a unique carboxy-terminal region between hIsm and hIsm-l suggests that Isthmin is the founding member of a new family of secreted proteins. xIsm was strongly expressed maternally in the Xenopus egg and showed zygotic expression in the ventral blastopore lip, notochord, and MHB. Additional expression domains were detected in neural crest, ear vesicle, and developing blood islands. Interestingly, xIsm was co-expressed with Fibroblast growth factor-8 (xFgf-8) at multiple sites including the MHB, indicating that these two genes are part of a synexpression group which also includes sprouty and sef homologs. PMID- 12128219 TI - The expression patterns of Wnts and their antagonists during avian eye development. AB - To determine the possible involvement of Wnt signaling in eye development, we analyzed the expression patterns of Wnts and Wnt inhibitors in the chicken eye at stage 25, when the first wave of neural crest migration into the cornea begins, and stage 27, just prior to the second wave of neural crest migration. Wnt expression is developmentally regulated in the chicken eye, and antagonists of Wnt signaling are generally expressed in patterns that are temporally distinct from the Wnts. PMID- 12128220 TI - Cloning and developmental expression of Baf57 in Xenopus laevis. AB - Mammalian and Drosophila homologues of Baf57 have been previously isolated as being a subunit of SWI/SNF-like chromatin remodeling complexes. Here, we report the cloning and developmental expression of Xenopus Baf57. We isolated XBaf57 by using an expression cloning approach to identify novel modulators of Xenopus Smad7. XBaf57 co-operates with XSmad7 by increasing the expression of neural markers in ectodermal explants. XBaf57 is expressed in the ectoderm and pre involuting mesoderm during gastrula stages and in the central nervous system during neurula and tailbud stages. These results raise the possibility that XBaf57 (or XBaf57-containing chromatin remodelling complexes) may be involved in the process of neural induction during Xenopus embryonic development. PMID- 12128221 TI - Cloning and expression of Xenopus Prickle, an orthologue of a Drosophila planar cell polarity gene. AB - We have cloned Xenopus orthologues of the Drosophila planar cell polarity (PCP) gene Prickle. Xenopus Prickle (XPk) is expressed in tissues at the dorsal midline during gastrulation and early neurulation. XPk is later expressed in a segmental pattern in the presomitic mesoderm and then in recently formed somites. XPk is also expressed in the tailbud, pronephric duct, retina, and the otic vesicle. The complex expression pattern of XPk suggests that PCP signaling is used in a diverse array of developmental processes in vertebrate embryos. PMID- 12128222 TI - The heart LIM protein gene (Hlp), expressed in the developing and adult heart, defines a new tissue-specific LIM-only protein family. AB - In a subtraction designed to identify transcripts accompanying mesodermal lineage specification in mouse ES differentiation cultures, we identified a gene encoding a two LIM-domain protein which we named heart LIM protein (Hlp). Hlp is most closely related to thymus LIM protein, and these two genes comprise a new gene family related to the cysteine-rich protein (CRP) gene family. In the embryo, Hlp expression is primarily restricted to the developing heart. In situ hybridization showed expression at E7.75 in the paired heart-forming primordia prior to linear heart-tube formation. At E8.5, strong expression is detected in the heart, with equal expression in both heart chambers. Hlp expression is detected in both myocardium and endocardium, and in vascular endothelium. Later in fetal development low levels of Hlp expression are detected outside the heart, including dorsal root ganglia and the spinal cord. In the adult, Hlp is expressed at highest levels in the heart, and at lower levels in the brain, skeletal muscle and aorta. Hlp expression is unchanged in hypertrophic hearts induced by aortic constriction. These data suggest a role for the two LIM-domain gene Hlp in the very earliest stages of heart differentiation and development. PMID- 12128223 TI - Characterization of Dir: a putative potassium inward rectifying channel in Drosophila. AB - Potassium channels vary in their function and regulation, yet they maintain a number of important features - they are involved in the control of potassium flow, cell volume, cell membrane resting potential, cell excitability and hormone release. The potassium (K(+)) inward rectifier (Kir) superfamily of channels are potassium selective channels, that are sensitive to the concentration of K(+) ions. They are termed inward rectifiers since they allow a much greater K(+) influx than efflux. There are at least seven subfamilies of Kir channels, grouped according to sequence and functional similarities (Curr. Opin. Neurobiol. 5 (1995) 268; Annu. Rev. Physiol. 59 (1997) 171). While numerous Kir channels have been discovered in a variety of organisms, Drosophila inward rectifier (Dir) is the first putative inward rectifier to be studied in Drosophila. In fact, there are only three genes (including Dir) encoding putative inward rectifiers in the Drosophila genome. Though there are other known potassium channels in Drosophila such as ether-a-go-go and shaker, most are voltage-gated channels. As an important first step in characterizing Kir channels in Drosophila, we initiated studies on Dir. PMID- 12128225 TI - FGFR1-IIIb is a putative marker of pancreatic progenitor cells. AB - The pancreas develops from buds that derive from the endodermal epithelium of the digestive tract. The progenitor cells that will give rise to the mature pancreatic cells reside within this epithelium. However, their exact identity remains unknown. In the present study, we searched for genes expressed by pancreatic progenitor cells. We focused our search on receptor tyrosine kinases. We found that fibroblast growth factor-IIIb (FGFR1-IIIb) expression is high in pancreatic epithelium enriched in progenitor cells. We next investigated FGFR1 IIIb expression throughout pancreatic development. At early stages of pancreas development, FGFR1-IIIb is expressed by pancreatic epithelial cells that resemble undifferentiated cells, while at later stages of development, FGFR1-IIIb expression decreases, concomitant with the expected decrease in the number of progenitor cells. PMID- 12128224 TI - The expression of neural-specific genes reveals the structural and molecular complexity of the planarian central nervous system. AB - Planarians are attractive animals in which various questions related to the central nervous system (CNS) can be addressed, such as its origin and evolution, its degree of functional conservation among different organisms, and the plasticity and regenerative capabilities of neural cells and networks. However, it is first necessary to characterize at the gene expression level how this CNS is organized in intact animals. Previous studies have shown that the planarian brain can be divided into at least three distinct domains based on the expression of otd/Otx-related genes. In order to further characterize the planarian brain, we have recently isolated a large number of planarian neural-specific genes through DNA microarrays and ESTs projects. Here, we describe new molecular domains within the brain of intact planarians by the expression of 16 planarian neural-specific genes, including the putative homologues of protein tyrosine phosphatase receptor, synaptotagmin VII, slit, G protein and glutamate and acetylcholine receptors, by in situ hybridization in both whole-mount and transverse sections. Our results indicate that planarian otd/Otx-positive domains can be further subdivided into distinct molecular regions according to the expression of different neural genes. We found differences at the gene expression level between the dorsal and ventral sides of the brain, along its antero posterior axis and also between the proximal and distal parts of the brain lateral branches. This high level of regionalization in the planarian brain contrasts with its apparent simplicity at the morphological level. PMID- 12128226 TI - The oculocutaneous albinism type IV gene Matp is a new marker of pigment cell precursors during mouse embryonic development. AB - Expression profile analysis demonstrated that the expression of membrane associated transporter protein (MATP) varied similarly to the melanogenic enzymes dopachrome tautomerase (DCT) and tyrosinase related protein 1 (TYRP1) (Proc. Natl Acad. Sci. USA (2002) in press). Mutations in MATP result in pigmentation alterations in mice (underwhite, uw), in medaka (b-locus), and in man (Oculocutaneous Albinism Type 4, OCA4) (Nat. Genet. 28 (2001) 381; Am. J. Hum. Genet. 69 (2001) 981). Consistent with MATP acting in a pigment cell autonomous manner, in situ hybridization analysis demonstrated expression of murine Matp in the presumptive retinal pigmented epithelium starting at E9.5, and in neural crest-derived melanoblasts starting at E10.5. Matp expression is reduced in embryos mutated for microphthalmia-associated transcription factor (Mitf) (Cell 74 (1993) 395; J. Biol. Chem. 268 (1993) 20687), suggesting Mitf regulates Matp expression. PMID- 12128227 TI - Differential expression of the Drosophila mitofusin genes fuzzy onions (fzo) and dmfn. AB - Mitofusins comprise a family of evolutionarily conserved, nuclear encoded mitochondrial guanosine triphoshatases that control mitochondrial fusion and morphology. The fuzzy onions (fzo) and Drosophila mitofusin (dmfn) genes, which encode the only Mitofusin homologs in Drosophila are differentially expressed during development. Dmfn-mRNA was widely expressed during embryogenesis accumulating in the mesoderm and endoderm during gut development, during oogenesis with transcripts maternally deposited into the early embryo and in the male germ line, where dmfn-mRNA was expressed in spermatogonia, spermatocytes and early spermatids. In contrast, expression of the fzo was tightly restricted to the male germ line, with mRNA accumulation in spermatocytes and early spermatids. In addition, expression of dmfn and fzo in the same cell type, primary spermatocytes, was under control of different regulatory mechanisms. PMID- 12128228 TI - Testis-specific expression of a novel mouse defensin-like gene, Tdl. AB - In order to isolate genes regulating sex differentiation of embryonic gonads, we have attempted to obtain genes specifically expressed in male embryonic gonads by means of subtractive hybridization screening, and we have cloned a novel mouse gene, Tdl. It potentially encodes a protein showing sequence homology to anti microbial protein, beta-defensins. Tdl reveals structural features such as the six cysteine residues with invariant spacing, which are found in beta-defensins, but the overall amino acid similarity of Tdl to other members of the beta defensin family is low. In addition, the Tdl gene shows genomic organization similar to that of other beta-defensin genes. We have found that Tdl is specifically expressed in Sertoli cell-lineage in seminiferous cords in embryonic testes, but not in embryonic ovaries after 12.5dpc when the sexual differentiation of gonads is initiated. Tdl is specifically expressed in gonads among adult tissues, and its expression persisted in Sertoli cells. PMID- 12128229 TI - Novel SOX9 expression during human pancreas development correlates to abnormalities in Campomelic dysplasia. AB - Haploinsufficiency of SOX9, which encodes a homeodomain transcription factor, results in Campomelic dysplasia. Classical features of this disorder (e.g. skeletal dysplasia and 46,XY sex reversal) are in concordance with SOX9 expression profiles during human embryonic development. We report the robust expression of SOX9 throughout the pancreas during human embryogenesis, at levels of detection equivalent to the developing skeleton and testis. In the early foetal period, SOX9 expression declines and, in particular, is not apparent within the pancreatic islets. In keeping with this profile, examination of three cases with Campomelic dysplasia revealed abnormal pancreatic morphology. Epithelial cells were less densely packed within the mesenchymal stroma and islets less clearly formed with variable expression of hormone and beta cell markers. Taken together, these data indicate a novel potential role for SOX9 in pancreas development during human embryogenesis and early foetal life. PMID- 12128230 TI - Platelet-derived growth factor receptor alpha (pdgfr-alpha) gene in zebrafish embryonic development. AB - Here we present the cloning of a full-length zebrafish pdgfr-alpha cDNA as well as the expression of this gene during zebrafish embryogenesis. We show that zebrafish pdgfr-alpha mRNA is present at high levels in the fertilized egg as well as in all embryonic cells up to the end of gastrulation. Spatially restricted expression of the gene started after the onset of segmentation and is mainly localized in premigratory neural crest cells, the placodes, the anterior paraxial cells of somites and the adaxial cells of the tailbud. Transient expression of this gene was also detected in the early Kupffer's vesicle, a teleost-specific structure. Expression of the zebrafish pdgfr-alpha is both conserved as well as diverged comparing to that of other vertebrate species. PMID- 12128231 TI - Cloning and developmental expression of AmphiBrn1/2/4, a POU III gene in amphioxus. AB - The large family encoding POU transcription factors has been described in several species. In particular, class III POU genes regulate critical steps of vertebrate and invertebrate neurogenesis. A novel amphioxus class III POU gene, AmphiBrn1/2/4, has been isolated and its spatio-temporal expression has been reported. AmphiBrn1/2/4 is first expressed in the dorsal epiblast, then throughout the neural plate except for a gap at level of the anterior region of the cerebral vesicle. Transcripts are also detected in the primordium of gill slits, pharynx and left Hatschek's diverticulum. PMID- 12128232 TI - Expression of hedgehog genes in Ciona intestinalis embryos. AB - The configuration of the ascidian tadpole larva represents the most simplified and primitive chordate body plan. The present study revealed that Ciona intestinalis contains two hedgehog genes (Ci-hh1 and Ci-hh2), which are likely to be independent duplicate genes in this animal and ancestral to the three types of hedgehog gene of vertebrates. Ci-hh1 was expressed maternally and its maternal transcript was distributed evenly in fertilized eggs and early embryos. Ci-hh2 was expressed zygotically in the tailbud embryo and its transcript was evident only in cells of the ventral nerve cord. The notochord cells did not express the hedgehog genes in Ciona embryos. PMID- 12128234 TI - Cloning and expression of the mouse dual-specificity mitogen-activated protein (MAP) kinase phosphatase Mkp3 during mouse embryogenesis. AB - Mitogen-activated protein (MAP) kinase phosphatases (MKPs) constitute a growing family of dual specificity phosphatases, which dephosphorylate both serine/threonine and tyrosine residues of MAP kinases. MAP kinase signaling cascades are involved in the control of cell proliferation, differentiation and apoptosis. In mammals, ten members of the dual-specificity MKP family have so far been identified. In this report, we describe the cloning and expression analysis of the mouse Mkp3 gene. During early development, expression of Mkp3 is most prominent in the primitive streak, presomitic mesoderm and somites, frontonasal prominence, midbrain/hindbrain boundary, branchial arches and limb buds. At later stages, expression is also detected in the tooth primordia, vibrissae, hair follicles, pinna, submandibular gland, mammary gland primordia, lung and kidney. Strong expression was detected in the adult brain. PMID- 12128233 TI - Expression of the chick vascular endothelial growth factor D gene during limb development. AB - Vascular endothelial growth factor D (VEGF-D) is a member of the VEGF/PDGF superfamily that has been implicated in angiogenesis and lymphangiogenesis. We have isolated a chick cDNA that shows homology with VEGF-D (also known as FIGF, c fos-induced growth factor) of other species. Here, we describe the expression pattern of cVegf-D in chick embryos. In the limb buds, cVegf-D shows a dynamic expression pattern that is restricted to the mesenchyme of the posterior region. cVegf-D expression is also detected in the ectoderm and mesenchyme of the head region, somites, notochord and pharyngeal arches. We also report on the capability of Sonic hedgehog and retinoic acid to regulate cVegf-D expression. PMID- 12128235 TI - To what extent do oral contraceptives influence mood and affect? AB - BACKGROUND: Studies examining the effects of oral contraceptives (OCs) on mood, affect, and affect variability are reviewed. METHODS: MEDLINE and PsycLIT data bases were examined to identify studies that compared OC users with nonusers using daily ratings of mood, affect, or affect variability. RESULTS: Compared to non-users, OC users experience less variability in affect across the entire menstrual cycle, and less negative affect during menstruation (i.e. withdrawal bleeding). In women with OC-related negative mood and affect change, potential mediators of the relation between OCs and mood or affect were identified: a history of depression, psychiatric symptoms, dysmenorrhea, and premenstrual mood symptoms prior to OC use; a history of pregnancy-related mood symptoms; a family history of OC-related mood complaints; being in the postpartum period; and age. Furthermore, a lower ratio of progesterone to estrogen is associated with more negative mood change in women with a history of premenstrual emotional symptoms, higher progesterone to estrogen ratios are associated with increased negative mood effects in women without such a history, and monophasic OCs have a greater stabilizing effect on mood than triphasic OCs. LIMITATIONS: The 'survivor effect', psychological factors, and indirect pharmacological effects (e.g. weight gain) have not yet been systematically investigated. Furthermore, most studies have examined only negative mood or affect, as opposed to both positive and negative affect and affect variability; and few affect studies have assessed potential mediators of OC-related affect change. CONCLUSIONS: While the only consistent OC-related mood effects experienced by most women are beneficial, a subgroup of women do experience negative mood change. Future research must focus on expounding the individual difference and OC-related risk factors for negative mood change. PMID- 12128236 TI - A general anxiety-prone cognitive style in anxiety disorders. AB - OBJECTIVE: This study compared scores on the Anxious Thoughts & Tendencies (AT&T) questionnaire, a putative measure of a general anxiety-prone cognitive style, among patients with panic disorder without agoraphobia (PD, n=62), panic disorder with agoraphobia (PDA, n=29), generalized anxiety disorder (GAD, n=43), limited social phobia (LSP, n=34), generalized social phobia (GSP, n=33), and community residents (n=319). METHOD: Candidates for treatment studies completed a diagnostic interview and the AT&T. AT&T scores were compared among anxious groups using analysis of variance. Then depressed and non-depressed patients were compared. The final analysis compared anxious groups without comorbid depressive or anxiety disorders. RESULTS: AT&T scores were highest in PDA patients, followed by patients with GAD or GSP, then patients with PD or LSP, with community residents lowest. Depressed patients were higher than non-depressed patients. Patients with current or past comorbid depressive disorders did not differ. The ranking of anxious groups on AT&T scores remained unchanged after exclusion of patients with comorbid disorders. Patients with PD or LSP without comorbidity had the same AT&T levels as the community sample. CONCLUSIONS: The AT&T discriminates PDA and GAD from PD per our hypothesis. The low AT&T levels among patients with PD and LSP suggest no association with a general anxiety-prone cognitive style. LSP and GSP may be distinct disorders. The cognitive style assessed by the AT&T is also associated with depression and may be a marker for vulnerability to depression. Definitive conclusions about a pathogenic role for cognitions require their measurement before the onset of the disorder. PMID- 12128237 TI - Is extended clonazepam cotherapy of fluoxetine effective for outpatients with major depression? AB - BACKGROUND: Clonazepam cotherapy of fluoxetine was previously demonstrated to accelerate efficacy over the first 3 weeks of treatment. A new 18-week double blind study attempted to replicate these findings to determine whether superiority would extend to 3 months and assess risks of extension. METHOD: Fifty outpatient volunteers aged 18-65 from Seattle and Portland with moderate-marked depression received fluoxetine (20 mg) doubled at 6 weeks if needed; half took clonazepam (0.5 mg) and half took an identical placebo, 1 or 2 tablets adjusted during the first 2 weeks, until a 3-week taper at 3 months. RESULTS: No serious adverse events and no special problems with sedation or discontinuation were noted. Cotherapy was superior to fluoxetine monotherapy at Day 7 for HAM-D (t=2.03, df=48, P<0.05) and CGI-I (32 vs. 4% responders, P<0.03, Fisher Exact Test) but not otherwise. Cotherapy was effective in reducing insomnia but not anxiety or core symptoms (low mood, suicidality, reduced interest). The only significant benefit of extending treatment was a more rapid response to increased fluoxetine at 6 weeks manifested in a mean HAM-D of 9.0 and CGI-I responder rate of 76% after 8 weeks compared to 16 weeks for monotherapy. LIMITATIONS: Small sample size (N=50) limited power and rendered conclusions tentative. CONCLUSIONS: Extended clonazepam cotherapy of fluoxetine appeared safe and effective for depressed outpatients: it was superior to fluoxetine alone early in treatment and again following fluoxetine dose increase. Cotherapy might be considered at the start of fluoxetine treatment, especially for those with insomnia, and when a dose increase of fluoxetine is anticipated. PMID- 12128238 TI - Depression in paranoid and nonparanoid schizophrenic patients compared with major depressive disorder. AB - BACKGROUND: Depression occurring in schizophrenia is a common problem; however, investigators have typically not studied it with the paranoid/nonparanoid dichotomy in mind. This study examines the quality as well as the severity of depression in three psychiatric groups: paranoid schizophrenia patients, nonparanoid schizophrenia patients, and nonpsychotic major depression patients. METHOD: Clinical and sociodemographic data were collected on 27 paranoid and 27 nonparanoid schizophrenia patients during their postpsychotic phase while they were at least mildly depressed, and a comparison group of 27 nonpsychotic patients diagnosed with major depressive disorder. The three groups were then assessed on various psychometric scales for severity of depression, profile of symptoms, suicidal risk, and anhedonia. RESULTS: The paranoid schizophrenia patients were more depressed and more at risk for suicide than the nonparanoid schizophrenia patients, yet their depressive profiles and levels of anhedonia were similar. CONCLUSIONS: Depressed mood and anhedonia constitute serious problems for schizophrenia patients, but particularly for paranoid schizophrenia patients during the postpsychotic phase of their illnesses. CLINICAL IMPLICATIONS: Schizophrenia patients, especially those with the paranoid features, should be routinely evaluated and monitored for depression. Apart from treatment with drugs, cognitive therapy may be considered a viable option, particularly for paranoid schizophrenia patients. LIMITATIONS: Gender was not matched for the two schizophrenia groups and extrapyramidal side effects were not measured. PMID- 12128239 TI - Heterogeneity in EEG sleep findings in adolescent depression: unipolar versus bipolar clinical course. AB - BACKGROUND: EEG sleep measures in child and adolescent subjects with depression have shown considerable variability regarding group differences between depressed and control subjects. This investigation was designed to assess whether some of the observed variability is related to undifferentiated unipolar and bipolar disorders in a sample that was reported previously. METHODS: Twenty-eight adolescents who met criteria for unipolar major depression and 35 controls with no lifetime psychiatric disorder participated in a cross-sectional sleep polysomnography study. Approximately 7 years later, follow-up clinical evaluations were conducted in 94% of the original cohort. Clinical course during the interval period was assessed without knowledge of subjects' initial diagnostic and psychobiological status. Re-analysis of the original sleep data were performed with the added information of longitudinal clinical course. RESULTS: Depressed subjects who had a unipolar course showed reduced REM latency, higher REM density, and more REM sleep (specifically in the early part of the night) compared with depressed adolescents who converted to bipolar disorder and controls who remained free from psychopathology at follow-up. In contrast to the unipolar group, depressed subjects who would later switch to bipolar disorder had demonstrated more stage 1 sleep and diminished stage 4 sleep. CONCLUSIONS: These preliminary results indicate that some of the observed variability in EEG sleep measures in adolescent depression appear to be confounded by latent bipolar illness. The findings also suggest that sleep regulatory changes associated with unipolar versus bipolar mood disorders may be different. PMID- 12128240 TI - The effect of a history of alcohol dependence in adult major depression. AB - We examined 180 patients with a principal diagnosis of Major Depression, with or without a lifetime diagnosis of alcohol dependence. In the ever-dependent group, the GAF was lower; cannabis dependence higher; and Borderline, Schizotypal and Paranoid personality disorders more common. They reported more paranoia and interpersonal sensitivity on the Hopkins Symptom Checklist and more friction and interpersonal behaviour on the Social Adjustment Scale. On the Temperament and Character Inventory, novelty seeking was higher and persistence and cooperativeness lower. PBI scores and family histories did not differ significantly. Treatment outcome did not vary, except that subjects with lifetime alcohol dependence and current heavy drinking did less well. PMID- 12128241 TI - Postpartum depression and mother-infant relationship at 3 months old. AB - BACKGROUND: This paper is part of a prospective, epidemiologic study concerning postpartum depression (PPD). The women were first examined during pregnancy; after delivery they were seen with their infants at 3 and 18 months. The present study focuses on the 3-months-postpartum results. METHODS: A sample of 570 women and their infants were examined 3 months after delivery. Using the EPDS (Edinburgh Postnatal Depression Scale; Cox et al., 1987. Br. J. Psychiatry 150:782-786), 10.2% of these new mothers presented PPD. The focus of the study concerned the effects of this neurotic disorder on the mother, the infant and on the mother-infant relationship. RESULTS: The deleterious effects concerning the infant were functional disorders such as eating or sleeping difficulties. The 'depressed' dyads presented less vocal and visual communications, less corporal interactions and less smiling. Conditions surrounding delivery and tiredness at 3 months are linked to difficulties in mother-infant relationship for the non depressed mothers. Logistic models showed that primiparous PPD mothers have difficulties bathing their infants, whereas multiparous PPD mothers are more tired. LIMITATION: This study did not take into account either protective factors or the effects of the infant himself. CLINICAL RELEVANCE: Knowledge of the mothers' and infants' difficulties may help caregivers to detect these at-risk dyads and initiate therapeutic measures. PMID- 12128242 TI - The interpersonal sensitivity measure in depression: associations with temperament and character. AB - OBJECTIVE: The first objective of this brief report is to examine the relationships between levels of interpersonal sensitivity and dimensions of personality, depression severity and early relationship with parents. An additional objective is to examine the differences between levels of interpersonal sensitivity in depressive subtypes. METHOD: One hundred and fifty four patients completed the Interpersonal Sensitivity Measure [IPSM], the Temperament and Character Inventory [TCI], and the Parental Bonding Instrument [PBI]. Other measures including Hamilton Depression Rating Scale [HDRS], DSM-IV atypical symptoms and DSM-IV melancholic symptoms were obtained using clinician rating scales. RESULTS: There were strong Pearson correlations between both the total and subscale scores of the IPSM and both temperament and character scores of the TCI. A joint principal components analysis isolated two main underlying constructs, both consisting of IPSM and TCI subscales. Patients with rejection sensitivity, an aspect of atypical depression, scored higher on the IPSM and three of its subscales, but there were no other differences in score by subtype. CONCLUSIONS: Both the IPSM and dimensions of the TCI measure similar constructs. These two constructs may help us to understand differences in symptom profile and response to therapy in depressed patients. PMID- 12128243 TI - A comparison of paroxetine and amisulpride in the treatment of dysthymic disorder. AB - BACKGROUND: The purpose of this study was to provide preliminary data on the effects of paroxetine and amisulpride on depressive dimensions, analyzed by factor analysis, in dysthymic patients. METHODS: One hundred and eighteen patients with DSM IV criteria for DD without concurrent major depression were enrolled in this 8-week, open study, and 100 completed it. Symptom dimensions were identified by principal components analysis with the SAS Factor procedure. RESULTS: Results of the symptom rating scales indicated that both drugs were equally effective. Response rate was 65% both in the paroxetine and the amisulpride group and the proportions of patients achieving a final HRSD score < or =7 were 46.7 and 55%, respectively. MADRS factor analysis identified two factors at baseline: the first corresponding to the global severity of depression and the second to somatic symptoms. After 8 weeks of treatment only one factor could be substantiated. At week 4 both paroxetine and amisulpride produced significant improvements on factor 1 while at week 8 mean changes of factor 1 were greater in the amisulpride-treated patients. LIMITATIONS: The main limitation was the open-label design. CONCLUSIONS: Both paroxetine and amisulpride appear to be effective in the short-term management of DD, improving its most characteristic symptoms. PMID- 12128244 TI - What are older peoples' experiences of taking antidepressants? AB - BACKGROUND: Antidepressants are prescribed widely to older people but little is known about older peoples' own reported experiences of taking them in routine practice. METHODS: A doctor interviewed 92 people, aged over 65, with a hospital diagnosis of depression, who had been prescribed an antidepressant in the past year. RESULTS: Most of the subjects were prescribed a selective serotonin reuptake inhibitor (SSRI), at a standard dose, for at least 8 weeks. Most people felt the antidepressants were helpful, although a third felt they made no difference. Two thirds of older people reported having adverse effects of which most were moderate or severe. The most common adverse effects were headache and dry mouth. Only a third said that they always took their tablets. LIMITATIONS: The sample was restricted to older people seen by hospital services. Interviews were carried out by a doctor and patients may not have been entirely truthful. CONCLUSIONS: Older people have important views about their treatment that they are prepared to tell a doctor. A lot of older people do not think their antidepressants are helpful, and the majority experience adverse effects. Many do not always take their medication. Doctors should routinely ask older people about their experiences of taking antidepressants. PMID- 12128245 TI - Severity of bipolarity in hospitalized manic adolescents with history of stimulant or antidepressant treatment. AB - BACKGROUND: Childhood bipolarity (BP) and ADHD frequently co-occur, these children often receive stimulants. METHOD: We retrospectively evaluated 80 adolescents hospitalized with BP, manic or mixed, assessed severity of hospital course, and compared groups according to current/past stimulant or antidepressant treatment. RESULTS: Lifetime ADHD rate was 49%; 35% of patients had exposure to stimulants and 44% to antidepressants. Stimulant-exposed patients were younger than non-exposed (mean+/-S.D.=13.7+/-2 vs. 15.1+/-2 years, Z=-3.1, P=0.002). Only stimulant exposure was associated with worse hospitalization course (MANCOVA, Wilks' Lambda=0.87, F=3.4; df=70; P=0.02). CONCLUSION: Stimulant-exposed BP adolescents may have more severe illness course not fully explained by ADHD comorbidity. LIMITATIONS: Retrospective methodology and lack of structured interviewing make it difficult to quantify exposure to stimulants and antidepressants. PMID- 12128246 TI - Effects of tricyclic antidepressants on protein kinase C activity in rabbit and human platelets in vivo. AB - BACKGROUND: The purpose of this study was to examine the effects of tricyclic antidepressants (TCA) on protein kinase C (PKC) in vivo. METHODS: PKC activity in rabbit and human platelets in vivo was measured after administration of TCA and in controls. RESULTS: Administration of TCA increased PKC activity in rabbit and human platelets in vivo. CONCLUSIONS: It has been reported that activation of PKC mediates inhibition of neurotransmitter uptake and down-regulation of beta adrenergic receptor. We suppose that TCA-induced activation of PKC may be associated, at least in part, with the mechanism of TCA. LIMITATIONS: Other signal transduction systems, such as those of protein kinase A, protein kinase G, and cyclic-AMP, also affect neurotransmitter uptake and/or down-regulation. In this study, the relationship between the TCA-PKC system and other signal transduction systems was not investigated. PMID- 12128247 TI - Erotomania preceding an aneurysmal subarachnid hemorrhage: is there an association? AB - Clear symptoms of erotomania were observed in a woman with past history of high blood pressure who, later, presented a slight subarachnoid hemorrhage from a ruptured anterior cerebral communicating artery aneurysm. The possibility that erotomania, in this case, may be related to the patient's organic brain damage is discussed. PMID- 12128248 TI - Seasonal affective disorder and social deprivation in Aberdeen. AB - BACKGROUND: Unlike non-seasonal depression, there is some evidence that seasonal affective disorder (SAD) is more common among more affluent socioeconomic groups. METHODS: In primary care settings in Aberdeen, 4557 subjects had previously completed a Seasonal Pattern Assessment Questionnaire (SPAQ). From the subjects' postcodes they were allocated a Carstairs score which placed them in one of seven categories of socioeconomic deprivation. These categories were compared with regard to seasonal pathology from the SPAQ ratings. RESULTS: Complete postcodes and Carstairs scores were established for 3772 (83%) of the 4557 subjects. No statistically significant relationship between socioeconomic deprivation and SPAQ ratings was detected. LIMITATIONS: The study population was an affluent one relative to Scotland as a whole which may have reduced the likelihood of a positive finding. The study was conducted 7 years after the census on which postcode deprivation scores were calculated, and changes therein may have occurred. CONCLUSIONS: SAD either has no relationship to social deprivation or is associated with affluence and this distinguishes it from non-seasonal depression. PMID- 12128249 TI - Immediate and delayed hippocampal neuronal loss induced by kainic acid during early postnatal development in the rat. AB - The degree to which the neonatal hippocampus is resistant to the effects of excitotoxins, such as kainic acid (KA) remains uncertain. Previously, we showed delayed loss of hippocampal neurons during pubescence in neonatal rats subjected to intracerebroventricular (i.c.v.) KA administration (10 nmol) at postnatal day 7 (P7). To further characterize the time course as well as the underlying mechanisms of this neuronal loss, we administered i.c.v. KA (10 or 50 nmol) to P7 preweanling rats. Brain sections were then examined at several neurodevelopmental time points (i.e., P8, P14, P25, P40, P60 and P75) using thionin staining and three-dimensional, non-biased cell counting to assess neuronal loss, and immunohistochemistry and electron microscopy to search for evidence of necrosis and apoptosis. Dose-dependent acute neuronal loss was observed at P8-P14 in hippocampal subfields CA3a and CA3c. Transient heat shock protein (HSP-70) immunostaining accompanied this acute neuronal loss. Progressive neuronal loss then continued in CA3 until P75, but without concomitant HSP-70 immunostaining. Progressive neuronal cell loss was also observed in the CA1 subfield of the hippocampus beginning at pubescence (i.e., P40) and continuing until P75. The appearance of TUNEL-positive hippocampal neurons accompanied the delayed neuronal loss in both CA3 and CA1 and electron micrographs confirmed that neurons in these subfields were undergoing apoptosis. KA administration (i.c.v.) to preweanling rats caused both immediate and delayed damage to hippocampal neurons. The effect of KA was dose-dependent, and the delayed neuronal damage occurred through an apoptosis-mediated mechanism. These findings may be relevant to the pathogenesis of some neuropsychiatric disorders, where early CNS injury is not apparent until the onset of clinical symptoms in young adulthood. PMID- 12128250 TI - Activation of protein kinase C isozymes in primary mouse myotubes by carbachol. AB - The activation of muscle PKC isozymes following treatment with carbachol, an acetylcholine receptor agonist, has been investigated. Primary mouse myotubes were treated with carbachol, and protein extracts from the cytosol and membrane fractions of the myotubes were subjected to Western blot analyses. Carbachol treatment resulted in a rapid translocation of PKC-theta; to the membrane. This effect was dependent on both carbachol concentration and incubation time. The treatment also resulted in a drastic increase of PKC-alpha in the cytosol followed by an increase of PKC-alpha in the membrane. The regulation of PKC-alpha in response to carbachol was quite distinct from that produced by the PKC activator, PMA, which rapidly translocated PKC-alpha from the cytosol to the membrane without any increases in PKC-alpha in the cytosol. Confocal microscopy demonstrated an enhanced membrane localization of PKC-theta; and overall increased intensity of PKC-alpha staining in the cytosol accompanied by a characteristic membrane staining of PKC-alpha in the myotubes treated with carbachol. Taken together, the results suggested that the activation of PKC isozymes in response to the receptor agonist is quite distinct, which indicates their diverse role in the muscle upon the release of neurotransmitter at the neuromuscular junction. PMID- 12128251 TI - Effects of dopamine and estrogen upon cortical neurons that express parvalbumin in vitro. AB - The purpose of these experiments was to study the effects of dopamine (DA) and 17 beta-estradiol (EST) upon parvalbumin expression in rodent frontal cortex during development. Organotypic slice cultures of the frontal cortex were prepared from neonatal rats (postnatal day 2/3) and maintained for 14 days in vitro in serum enriched medium and medium treated with either DA, EST or DA+EST. Cultured slices were then fixed and immunostained for parvalbumin immunoreactivity. Under control conditions, parvalbumin immunoreactive somata and fibers were primarily found in the deep laminae. In comparison, slices in all treatment groups exhibited a pattern of parvalbumin expression that was significantly different than controls. Specifically, DA treatment increased the percentage of parvalbumin immunoreactive somata, dendritic length and density in the deep cortical layers, but not in the superficial cortical layers. Both EST and DA+EST treatments induced similar changes in both the deep and the superficial cortical layers. These treatment induced changes represent more mature patterns of parvalbumin expression when compared to controls, indicating that both DA and EST enhance cortical expression of the protein. PMID- 12128252 TI - Acute neuronal injury after hypoxia is influenced by the reoxygenation mode in juvenile hippocampal slice cultures. AB - In neonates asphyxia is usually followed by hyperoxia due to resuscitation procedures. In order to study whether hyperoxic reoxygenation might cause additional cell injury we subjected organotypic hippocampal slice cultures of juvenile rats to normoxic or hyperoxic reoxygenation (19 or 85% oxygen, respectively) following hypoxia (3% oxygen) for 30, 60, and 120 min. Cell injury was quantified by lactate dehydrogenase (LDH) release and propidium iodide (PI) fluorescence 1 h after end of the reoxygenation period. In both experimental groups, LDH activity was significantly enhanced by hypoxia as compared to normoxic controls. However, hyperoxic reoxygenation caused a larger increase in LDH activity than normoxic reoxygenation (e.g., by a factor of 1.60 vs. 1.29 following 120 min hypoxia). PI fluorescence increased after hypoxia in all principal cell layers of the hippocampus but again showed a larger enhancement after hyperoxic reoxygenation as compared to normoxic reoxygenation (e.g., by a factor of 3.9 vs. 1.7 for CA1 and 120 min of hypoxia). After normoxic reoxygenation, PI fluorescence intensity was lower in the dentate gyrus as compared to CA1 and CA3, while it reached similar values like CA1 after high oxygen supply. In conclusion, juvenile hippocampal slice cultures subjected to hyperoxic reoxygenation display a greater amount of acute neuronal injury than slice cultures undergoing normoxic reoxygenation. PMID- 12128254 TI - Thyroid hormone-dependent development of mouse cerebellar Purkinje cells in vitro. AB - Using a well-defined medium with insulin, transferrin and selenium but without serum and albumin, we quantitatively determined the effect of thyroid hormones on the development of Purkinje cells in mouse cerebellar monolayer cultures. Addition of a thyroid hormone, T3 or T4, to the serum-free medium resulted in a highly elaborate dendritic development of Purkinje cells. The cultured Purkinje cells in the presence of T4 even showed similarities in shape and in synapse formation to normal Purkinje cells in vivo. Such effect of T4 on the dendritic arborization of Purkinje cells was dose dependent and significantly sensitive to a low dose of T4 even at 50 pM. The effect of T4 was confirmed by an inhibition experiment using amiodarone, which was reported to induce thyroid dysfunction. Furthermore, T4 affected not only Purkinje cell development but also the shape of other neural cells such as small interneurons (mainly granule cells) and astrocytes in cerebellar cultures. T4 induced development of both interneurons and astrocytes having long processes. These results indicate that thyroid hormones play a pivotal role in the development of mouse Purkinje cell dendrites acting on Purkinje cells directly and/or indirectly via the close interaction with interneurons and astrocytes. PMID- 12128253 TI - Rescue of the acetylcholinesterase knockout mouse by feeding a liquid diet; phenotype of the adult acetylcholinesterase deficient mouse. AB - Acetylcholinesterase (AChE, EC3.1.1.7) functions in nerve impulse transmission, and possibly as a cell adhesion factor during neurite outgrowth. These functions predicted that a mouse with zero AChE activity would be unable to live. It was a surprise to find that AChE -/- mice were born alive and survived an average of 14 days. The emaciated appearance of AChE -/- mice suggested an inability to obtain sufficient nutrition and experiments were undertaken to increase caloric intake. Pregnant and lactating dams (+/-) were fed 11% high fat chow supplemented with liquid Ensure. AChE -/- pups were weaned early, on day 15, and fed liquid Ensure. Although nullizygous animals showed slow but steady weight gain with survival over 1 year (average 100 days), they remained small at all ages compared to littermates. They demonstrated delays in temperature regulation (day 22 vs. 15), eye opening (day 13 vs. 12), righting reflex (day 18 vs. 12), descent of testes (week 7-8 vs. 4), and estrous (week 15-16 vs. 6-7). Significant physical findings in adult AChE -/- mice included body tremors, abnormal gait and posture, absent grip strength, inability to eat solid food, pinpoint pupils, decreased pain response, vocalization, and early death caused by seizures or gastrointestinal tract ileus. Behavioral deficits included urination and defecation in the nest, lack of aggression, reduced pain perception, and sexual dysfunction. These findings support the classical role for AChE in nerve impulse conduction and further suggest that AChE is essential for timely physical development and higher brain function. PMID- 12128255 TI - Fetal exposure to (+/-)-methylenedioxymethamphetamine in utero enhances the development and metabolism of serotonergic neurons in three-dimensional reaggregate tissue culture. AB - Methylenedioxymethamphetamine (MDMA, Ecstasy) is a potent psychomotor stimulant with neurotoxic potential which is widely abused by females of childbearing age raising serious public health concerns in terms of exposure of the fetus to the drug. The current study was conducted using the three-dimensional reaggregate tissue culture system as an approach to the assessment of risk to fetal brain cells following exposure to MDMA during early to mid-gestation. In this culture system, the serotonergic and dopaminergic mesencephalic-striatal projections are reconstructed and develop with a time course similar to that observed in vivo. Pregnant C57Bl/6J mice were injected twice daily with 40 mg/kg MDMA or saline from gestational day 6 to 13. On gestational day 14, mesencephalic and striatal cells from MDMA- and saline-exposed embryos were used to prepare reaggregate cultures. Levels of neurotransmitters and their metabolites in the reaggregates and culture medium were assessed at 22 and 36 days of culture. There was a long term enhancement of serotonergic development and metabolism by fetal exposure to MDMA as evidenced by increased reaggregate serotonin levels as well as the elevated production and release of 5-hydroxyindoleacetic acid in cultures prepared from MDMA-exposed embryos which persisted for up to 36 days of culture. Dopaminergic neurons in such cultures also exhibited increased metabolism as indicated by elevated levels of dihydroxyphenylacetic acid in reaggregate tissue and culture medium. The data obtained suggest that exposure to MDMA in utero during early to mid-gestation may result in more active serotonergic and dopaminergic neurons. PMID- 12128256 TI - Local proliferation of microglia cells in response to neocortical injury in vitro. AB - Studies examined whether increased numbers of microglia following neural damage result from induced mitotic activity of resident microglia in situ. Organotypic slice cultures of neocortex were maintained for 1 week prior to placement of lesions. Increased numbers of OX-6 or tomato lectin labeled microglial cells were detected 1-8 days following lesions. Administration of BrdU to the cultures demonstrated lectin and BrdU double labeled microglial cells, conclusively demonstrating that a portion of the microglial cells were generated by local mitotic activity in situ. PMID- 12128257 TI - Ontogeny of descending serotonergic innervation and evidence for intraspinal 5-HT neurons in the mouse spinal cord. AB - Neuronal networks in the mouse spinal cord express serotonin (5-HT)-induced rhythmic motor activity at early developmental stages (embryonic day (E) 12.5). Later in development, by post-natal day (P) 10, the 5-HT-evoked rhythmic motor activity matures and acquires an adult locomotor-like pattern. With the view to establishing a relationship between the ontogeny of locomotor networks and the maturation of spinal 5-HT systems, we have traced 5-HT immunoreactivity in the mouse spinal cord from E12.5 to PN10. By E12.5, descending 5-HT immunoreactive (5 HT-ir) fibers that likely originate from raphe nuclei were detected in the ventral and lateral funiculi, at anterior cervical spinal levels, but not at more caudal levels. Descending 5-HT-ir axons reached thoracic levels at E14.5 and lumbar levels at E16.5. Some 5-HT-ir fibers could be detected in the ventral and intermediate gray matter by E16.5, whereas the dorsal gray matter was not invaded before PN0. At PN10, a dense serotonergic innervation was restricted to the gray matter with a high concentration of 5-HT-ir fibers in three areas: dorsal horn, ventral horn (where motoneurons are located) and intermediate area. Surprisingly, from E16.5 to PN10, 5-HT-ir intraspinal neurons were found, exclusively at sacral levels. Their somata lay in the gray matter around the central canal and preferentially in the ventro-median part of the ventral horn. The functional significance of these sacral 5-HT-ir neurons is discussed. PMID- 12128258 TI - Development of the catecholaminergic system in the early zebrafish brain: an immunohistochemical study. AB - Tyrosine hydroxylase-containing cells (TH cells) were investigated immunohistochemically in early and late postembryonic zebrafish brain sections (at 2 and 5 days postfertilization [dpf]) yielding an improved neuroanatomical resolution of spatiotemporal developmental dynamics of the catecholaminergic system. Additionally, double-immunolabel preparations for visualizing TH cells and cells containing the proliferating cell nuclear antigen (PCNA cells) were carried out allowing for a prosomeric interpretation of early forebrain TH cell clusters. Many TH cell populations recently described in the adult zebrafish brain could be identified in the present study by location and cell type already in the 5 dpf (e.g. eight of 12 adult diencephalic TH cell populations) and 2 dpf (e.g. five of 12 adult TH cell populations) zebrafish brain. Early and adult diencephalic TH cells are restricted to the pretectum (P1) and ventral thalamus (P3) in the alar plate, and to various TH groups in the basal plate posterior tuberculum (P3), as well as to various populations in the hypothalamus (secondary prosencephalon). The alar plate ventral thalamic and most anterodorsal posterior tubercular TH cell populations range among the earliest detectable ones. There was no indication of migration of TH cells from the midbrain-hindbrain boundary or anterior neural ridge into the diencephalon. PMID- 12128259 TI - Delivery of the second twin: comparison of two approaches. AB - OBJECTIVE: To compare two obstetrical approaches toward delivery of the second twin: one of expectant management, and the other, active; to compare the neonatal and maternal results and thereby identify, if possible, the optimal approach. STUDY DESIGN: This retrospective study looked at twin births in two maternity units in the Paris, France metropolitan region: Antoine Beclere (AB) in Clamart, and Port-Royal (PR) in Paris and concerned 113 deliveries of pairs of twins at AB and 78 at PR. RESULTS: The mean duration of the interbirth interval was 9 min at AB and 5 min at PR (P < 0.001). The characteristics of the pregnancies and the deliveries of twin A were comparable. Spontaneous birth accounted for 51% of twin A births at AB and 27% at PR (P < 0.001). Intrauterine manipulation of twin B occurred in 2% of the births at AB and 43% at PR (P < 0.001). At AB, there were five cesareans to deliver the second twin, but none at PR. The Apgar scores at AB and PR were identical, at 1 and 5 min, and for births before 32 weeks' gestation as well as for those afterwards. At AB, 19% (n = 21) of second twins were transferred to the neonatal intensive care unit, and at PR, 18% (n = 14). CONCLUSION: The neonatal results were similar in both groups, even though both the rate of obstetric maneuvers and the interbirth interval differed significantly. The two methods therefore appear to be equivalent when judged by the second twin's neonatal indicators. Our data suggest that an active approach diminishes the likelihood of cesarean delivery for the second twin, without increasing the neonatal risk. PMID- 12128260 TI - Ultrastructural features and ICSI treatment of severe teratozoospermia: report of two human cases of globozoospermia. AB - Abnormal sperm morphology is associated with male infertility. We describe two human cases of globozoospermia (round-headed spermatozoa) together with fine diagnosis and proposed treatment. Scanning and transmission electron microscopy (SEM and TEM) were performed to identify the ultrastructural features. Female partners underwent ovarian hyperstimulation and intracytoplasmic sperm injection (ICSI). Fertilized oocytes were transferred 36 h later. One couple had a healthy live-birth. Ultrastructural analysis may help to diagnose sperm morphology and identify those which will respond to treatment. PMID- 12128262 TI - Non-Hodgkin lymphoma of the female genital tract. A five case series. AB - Non-Hodgkin lymphoma (NHL) of the female genital tract is extremely rare. We report five cases of NHL, which were collected during an 8-year period from the files of the Department of Pathology at the Onze Lieve Vrouwe Gasthuis, Amsterdam. In these five cases, the NHL was clinically not considered and the genital location was the primary site of presentation. PMID- 12128261 TI - Knowledge about contraception in women undergoing repeat voluntary abortions, and means of prevention. AB - AIMS: Despite reliable and effective means of contraception, cases of repeat abortion are on the increase in all developed countries. The aim of this work was to determine whether women undergoing repeat abortions are exposed to risk factors which might be amenable to preventative measures, and the methods employed by carers in these cases. METHODS: We set out to evaluate practices in the Family Planning Centre of l'Hopital Jean Verdier (Bondy, France) by sending a questionnaire to 147 women who had undergone two abortions up to 1997, and by conducting interviews with the care team. Thirty patients responded to the questionnaire. RESULTS: Twenty-two women (73%) underwent one or more further abortions between 1999 and 2000. Twenty-seven out of 30 women were unaware of the existence of emergency contraception. The 'morning after' pill, indicated for cases of unprotected sex, was unknown to one woman in two (15), nine out of 30 did not know what 'back-up' measures they should take after missing a dose of the contraceptive pill. Psychological problems were found in nine cases. These were followed up with a psychological consultation in three cases. The information given to the patients by the carers was the same irrespective of the number of abortions. Poverty and psychological problems were noted by the carers. CONCLUSION: Patients who have undergone two abortions might benefit, in addition to their routine visits, from a consultation with a psychologist and a consultation providing information about contraception. Providing the contraceptive pill free of charge to low-income patients is essential. PMID- 12128263 TI - Ultrasound guided aspiration in pathological adnexal processes. AB - OBJECTIVE: To establish the efficiency of ultrasonographically guided transvaginal adnexal cyst aspiration as a treatment and diagnostic method. STUDY DESIGN: In 72 patients with an adnexal cystic mass, transvaginal ultrasound guided cyst aspiration was performed. Before the procedure, presence of primary malignant disease was excluded by gynecologic and ultrasound examination. Cyst content was sent for cytological analysis. Cytological findings were staged according to Papanicolaou. Patients were re-examined 3 and 6 months after the ultrasound intervention. Cysts measuring 3 cm or more in diameter were considered to be recurrence of the disease. RESULTS: Recurrence of the disease appeared in 32 cases (44%) and was more common with larger cysts. Malignant cells were found in one case (1.5%), a recurrent ovarian cancer, previously treated by surgery and chemotherapy. CONCLUSION: In our study, ultrasound guided aspiration of adnexal cysts was not shown to be an efficient method of treatment because of the high recurrence rate. It may be used in selected patients at high anaesthesiologic risk for surgery as a therapeutic or a diagnostic procedure. PMID- 12128264 TI - Expression and clinical significance of pepsinogen C in epithelial ovarian carcinomas. AB - BACKGROUND: Pepsinogen C (pep C) is a gastric aspartic protease of which is associated with a favorable prognosis in breast cancer. Recently, it has been demonstrated in other tumors of extradigestive origin. STUDY DESIGN: We have analyzed pep C expression in 72 epithelial ovarian carcinomas by immunohistochemistry. RESULTS: Nineteen (26%) tumors stained positively for pep C. Overall this expression was not associated with the clinicopathologic characteristics or with outcome. However, in patients with serum levels of CA 125 less than 35 U/ml, pep C expression identified a group with a more favorable prognosis. CONCLUSION: Pepsinogen C is expressed in a quarter of ovarian carcinomas and might identify a subset of patients with different prognosis. PMID- 12128265 TI - The effectiveness of spectral and color Doppler in predicting ovarian torsion. A prospective study. AB - We evaluated the effectiveness of color and spectral Doppler examination of the ovarian vasculature flow, using transvaginal sonography (TVS) in 65 women prior to laparoscopy due to suspected ovarian torsion. There were 15 cases of ovarian torsion. In all of them, a pathology was detected by the color and spectral Doppler examination. Of the 50 patients without torsion at laparoscopy, one had abnormal Doppler studies. Color and spectral Doppler can demonstrate the presence or absence of arterial and venous flow in cases of suspected torsion of the ovary. PMID- 12128266 TI - Management and evolution of cervical intraepithelial neoplasia during pregnancy and postpartum. AB - OBJECTIVE: To investigate the evolution of cervical intraepithelial neoplasia (CIN), and to evaluate the safety of cytological and colposcopical surveillance of women with CIN during pregnancy. STUDY DESIGN: Ninety-eight women with antenatal cytological and/or colposcopical impression of CIN were followed up during pregnancy with cytology and colposcopy every 2 months. A cytological and colposcopical reevaluation 2 months postpartum was done, and large loop excision of the transformation zone (LLETZ) was performed if appropriate. Punch or loop biopsies were only taken if there was suspicion of microinvasion. RESULTS: In 14 of 39 (35.9%) and in 25 of 52 (48.1%) women with antenatal impression of CIN I and CIN II-III, respectively, there was postnatal impression of regression. Seven women with findings suspicious of microinvasion underwent small loop biopsies during pregnancy, but early stromal invasion (< 1 mm) was seen in just one case. There was one more case of microinvasion (1.5 mm) diagnosed postnatally in which the antenatal impression was of CIN III. 84.6% of the women with regression compared to 67.3% of the women with stable disease or progression had a vaginal delivery (P = 0.057). CONCLUSION: There is a considerable regression rate of CIN after pregnancy possibly attributable to the loss of the dysplastic cervical epithelium during cervical ripening and vaginal delivery. Frequent cytological and colposcopical evaluation seems to be safe. Small loop biopsies are recommended in cases of possible microinvasion. PMID- 12128267 TI - Sigmoid colon cancer during pregnancy. AB - Colorectal carcinoma during pregnancy is a rare event. We report a 23-year-old primigravida with advanced stage adenocarcinoma of the sigmoid colon diagnosed at 34 weeks of gestation. A healthy female infant was delivered by cesarean section. The treatment of choice was chemotherapy. The patient died 3 months after delivery. PMID- 12128268 TI - Fertilisation and implantation failure in an oral contraceptive user. AB - We report the case of a young woman taking a low-dose oral contraceptive (gestodene 0.075mg and ethinylestradiol 0.02mg) in whom we documented by both hormonal assays and sonographic evaluations the occurrence of ovulation, oocyte fertilization and embryo implantation. However, the implantation process did not complete and only a biochemical pregnancy was registered. On the basis of known actions of estroprogestin on endometrium that are not conducive to implantation, it is possible that the pregnancy loss was originated by oral contraceptive's hormonal influence at endometrial level. PMID- 12128269 TI - Incidental carcinoid of appendix in cesarean section. AB - A case with carcinoid tumor of the appendix was encountered incidentally during an elective cesarean section. The tumor was discovered as a result of the routine exploration of the whole abdomen. The investigation for metastasis proved no evidence of spread and the patient was treated solely by surgery. PMID- 12128270 TI - An infertile patient with breast cancer who delivered a healthy child under adjuvant tamoxifen therapy. PMID- 12128271 TI - Near-miss maternal mortality in developing countries. PMID- 12128273 TI - Specialist life-Richard Neale. PMID- 12128275 TI - The use of low-molecular-weight heparin for the management of venous thromboembolism in pregnancy. AB - Thromboembolic disease is a rare, but important, complication of pregnancy that remains a leading non-obstetric cause of maternal death. The prevention and management of venous thromboembolism (VTE) in pregnant women is a complex area of medicine: a balance must be found between protecting the health of the mother and minimizing the risk to the unborn fetus. Until now, unfractionated heparin has been regarded as the drug of choice for the prevention and treatment of VTE during pregnancy. However, because of its significant side effects (osteoporosis and heparin-induced thrombocytopenia), the inconvenient mode of administration and need for monitoring, unfractionated heparin is now being replaced by low molecular-weight heparin (LMWH). There is a convincing body of clinical evidence from well-designed studies and prospective case series that supports the efficacy and safety of LMWH in pregnant women. There are also encouraging observations on the efficacy of LMWH in the prevention of severe obstetric complications, which are frequently associated with inherited or acquired thrombophilias. The recently published guidelines of The American College of Chest Physicians (ACCP), summarized in this review, allows the development of higher clinical standards. However, there is concern over the greater cost of LMWH compared with unfractionated heparin and oral anticoagulants, and cost-effectiveness studies are needed. PMID- 12128276 TI - Mothers' knowledge of screening for trisomy 21 in 1999: a survey in Paris maternity units. AB - OBJECTIVE: To assess mothers' knowledge of screening tests for trisomy 21. STUDY DESIGN: Interview of all women who had recently delivered a healthy child and were present in 15 Paris maternity units during one of the two non-consecutive days in June 1999 (N = 734). RESULTS: Two-third said that they had access to a nuchal translucency measurement (NTM) and to maternal serum screening (MSS), and 16% to amniocentesis. Thirty-eight percent of the women who had NTMs and 69% of those who had serum screening said that they had been informed of the need for amniocentesis if the results were abnormal. Among the women who had amniocentesis, 20% did not know the risk of miscarriage and 41% had not been informed about the possibility of terminating the pregnancy if trisomy 21 was diagnosed. CONCLUSIONS: Mothers' knowledge about the screening tests for trisomy 21 remains fragmentary. Providing comprehensive information about all these tests should be considered in early pregnancy so that women can make informed choices. PMID- 12128277 TI - Myasthenia gravis in pregnancy: report on 69 cases. AB - OBJECTIVE: To review our experience with pregnancies in women with myasthenia gravis (MG). STUDY DESIGN: Sixty nine pregnancies among 65 women with MG patients managed by our department over 28 years were included. The course of the disease in pregnancy, mode of delivery and postpartal period were evaluated. RESULTS: One pregnancy miscarried. In 15% of patients the MG deteriorated in pregnancy a further 16% in the puerperium. 17% of pregnancies were delivered by cesarean section, one due to myasthenia exacerbation. All women with puerperal infections developed exacerbations. One neonatal death, not attributable to myasthenia, was recorded. Transitory neonatal myasthenia gravis (TNMG) was diagnosed in 30% infants. Its incidence was inversely associated with maternal disease duration (P < 0.05). Newborns of thymectomized mothers showed lower rate of neonatal myasthenia compared to those of non-thymectomized women (P < 0.05). CONCLUSIONS: MG patients can have normal pregnancy and delivery but the course is unpredictable. Shorter disease history and infection predispose to puerperal exacerbation. Maternal thymectomy lessens the likelihood of neonatal myasthenia. An interdisciplinary approach is required for managing the pregnant women with MG. PMID- 12128278 TI - Estrogens and neuroprotection. AB - In recent years, we have become increasingly aware that estrogen is a gonadal hormone that exerts diverse non-reproductive actions on multiple organs and in multiple physiological systems. Amongst these, estrogen has profound effects on plasticity and cell survival of the adult brain. Over the past 100 years, the lifespan of women has increased to >80 years, but the age of the menopause has remained fixed. Women are therefore living an ever-increasing proportion of their lives in a hypoestrogenic, postmenopausal state, which could contribute to an increased risk of cognitive dysfunction and a variety of neurodegenerative diseases. Recent experiments emphasize the importance of apoptosis as a mechanism of cell death after brain injury induced by global ischemia, and indicate that estrogen treatment has a neuroprotective effect by attenuating expression of selective markers of apoptosis. PMID- 12128279 TI - Early encounters, lifetime effects: hormones in the intrauterine environment. PMID- 12128283 TI - Dissecting human adrenal androgen production. AB - The human adrenal cortex produces aldosterone, cortisol and the so-called adrenal androgens, dehydroepiandrosterone (DHEA) and DHEA sulfate (DHEAS). Within the adult adrenal, the zona glomerulosa produces aldosterone, the zona fasciculata cortisol and the zona reticularis both DHEA and DHEAS. The processes regulating aldosterone and cortisol synthesis are well defined; however, the mechanisms regulating the production of DHEA(S) remain elusive. The emphasis of this review is based on increasing evidence that cytochrome b(5), DHEA sulfotransferase and 3 beta-hydroxysteroid dehydrogenase play crucial roles in regulating production of DHEA(S). Insight into the mechanisms that regulate the synthesis of these key components of DHEA(S) synthesis should provide important clues to the regulation of adrenal androgen biosynthesis. PMID- 12128284 TI - Regulation of insulin action by CEACAM1. AB - Activation of the tyrosine kinase of the insulin receptor by insulin binding initiates a cascade of signaling pathways that mediates the metabolic and growth promoting effects of insulin. Insulin action is regulated by the amount of circulating insulin, which is, in turn, partially regulated by insulin clearance in liver. Receptor-mediated insulin endocytosis followed by degradation mediates insulin clearance. Earlier studies in transfected cells suggested that the carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1), a substrate of the insulin receptor in liver, upregulates receptor-mediated insulin endocytosis and degradation in a phosphorylation-dependent manner. To test this hypothesis, a transgenic mouse, L-SACC1, overexpressing a dominant-negative phosphorylation-defective S503A CEACAM1 mutant in liver was established. The transgenic mouse demonstrated that CEACAM1 increases insulin clearance to maintain insulin sensitivity. Because insulin resistance is the hallmark of type 2 diabetes, understanding the mechanism of CEACAM1 regulation of insulin clearance and action might lead to novel therapeutic strategies against this disease. PMID- 12128285 TI - Prolactin as an autocrine/paracrine growth factor in human cancer. AB - Prolactin (PRL) has a dual function -- as a circulating hormone and as a cytokine. This understanding is based on PRL production and distinct regulation in extrapituitary sites, its binding to membrane receptors of the cytokine receptor superfamily, and activation of signaling pathways that promote cell growth and survival. There is increasing evidence that PRL plays a role in several types of cancer in reproductive and non-reproductive tissues via local production or accumulation. The expression of both PRL and its receptor in human cancer cell lines of diverse origin lends further support to its action as an autocrine/paracrine growth factor. Establishment of PRL as an active participant in tumorigenesis should inspire the development of novel therapies aimed at reducing tumor growth by suppressing PRL production or by blocking its receptors. PMID- 12128286 TI - The role of lifestyle modification in polycystic ovary syndrome. AB - Polycystic ovary syndrome (PCOS) is a common endocrine condition with reproductive and metabolic consequences, including anovulation, infertility and an increased prevalence of diabetes mellitus. Obesity, central obesity and insulin resistance are strongly implicated in its etiology and reduction of these risk factors should be a central treatment focus. Short-term weight loss has been consistently successful in reducing insulin resistance and restoring ovulation and fertility. However, problems arise with maintaining weight loss and precisely quantifying the associated long-term benefits of risk factor change. Although recent research indicates modest long-term lifestyle changes might reduce the extent of impaired glucose tolerance and delay the conversion to diabetes mellitus in the general population, this has not yet been examined in women with PCOS. Current conservative treatment should emphasize sustainable weight loss through dietary modification and exercise. Modifying additional lifestyle factors, including alcohol consumption, psychosocial stressors and smoking, are also crucial in long-term treatment of PCOS. PMID- 12128287 TI - The nature of FSH induction by GnRH. AB - Follicle-stimulating hormone (FSH), a major regulator of mammalian gonadal function, is induced by gonadotropin-releasing hormone (GnRH), but it is unclear how much induction is direct or indirect and what relevance each has in vivo. Two advances now make it possible to address these issues, which are central to understanding FSH regulation. The first is the use of transformed L beta T2 gonadotropes to define key promoter sequences of FSHB (the gene encoding the FSH beta subunit) that are needed for induction by GnRH and/or other factors; and the second is the ability to express FSHB promoter-reporter constructs in transgenic mouse gonadotropes to test the physiological relevance of promoter elements identified by using L beta T2 cells. Here, we summarize past studies on GnRH induction of FSH, and propose questions and approaches for the future. PMID- 12128288 TI - Calcitonin gene-related peptide in pregnancy and its emerging receptor heterogeneity. AB - Calcitonin gene-related peptide (CGRP) is the most potent vasodilator, and there is a growing body of evidence that this peptide might have multiple other functions. During pregnancy, circulating CGRP levels in rats increase up to the time of delivery, followed by a sharp decline at term and postpartum. In addition, the sensitivity of various vascular beds to CGRP in rats appears to increase with advancing pregnancy. This increased sensitivity might be involved in regulating uteroplacental blood flow, in addition to other vascular adaptations that occur during normal pregnancy. Furthermore, the uterine relaxation response to CGRP is elevated during pregnancy and decreased at term. Sex steroid hormones, estrogens and progesterone, regulate CGRP synthesis and its effects on both myometrial and uterine vascular tissues. These changes in smooth muscle relaxation sensitivity to CGRP appear to be a consequence of changes in CGRP-receptor levels in these tissues. There appear to be two receptors for CGRP: the CGRP-A receptor, a well-characterized receptor consisting of calcitonin receptor-like receptor and receptor activity modifying protein 1, and the CGRP-B receptor. The CGRP system might play a role in the maintenance of normal pregnancy, and a defect in this system might lead to complications. PMID- 12128295 TI - Surgical management of colloid cyst of the third ventricle--a study of 105 cases. AB - BACKGROUND: Colloid cyst of the third ventricle is a relatively rare intracranial tumor. It generates tremendous interest for the neurosurgeon because of its benign nature, deep location, and an excellent prognosis when diagnosed early and excised. METHOD: A retrospective analysis of 105 cases of third ventricle colloid cyst treated between 1967 to 1998 was conducted. The clinical presentation, radiological findings, different surgical approaches, and outcome were analyzed. The transcallosal and transcortical-transventricular approaches were predominantly used. Memory and psychological assessment were carried out both pre and postoperatively. A computerized tomography (CT) scan was performed during follow-up. RESULTS: The male to female ratio was 1.5:1. The age of the patients ranged from 10 to 68 years. Headache was the most common symptom. Papilledema and short-term memory disturbances were the most common signs. In 5 patients the colloid cyst was detected incidentally. Surgery for colloid cyst was performed in 93 patients. Transcallosal and transcortical-transventricular approaches were performed in 62 and 30 patients, respectively. In 1 patient the cyst was excised through the subfrontal lamina terminalis approach. Total excision was achieved in 90 patients, while partial cyst excision was done in three patients. Moderate to severe lateral ventricular enlargement was found in 76 patients at presentation. A ventriculoperitoneal shunt was the only surgical procedure performed in 7 patients. In 16 patients colloid cyst excision was conducted after cerebrospinal fluid (CSF) diversion via a shunt. No surgical treatment of any kind was performed in 5 patients. Five patients died. Eighty-six patients came for follow up, with a range from 1 month to 25 years (average 3 years and 8 months). Postoperatively, transient recent memory deficits occurred in 14 patients, while a permanent recent memory loss was noted in 2 patients. There was no incidence of postoperative disconnection syndrome or behavioral disturbance. A CT scan was performed in 44 patients during follow-up. Recurrence was detected in 1 patient in whom the cyst had been partially excised. CONCLUSIONS: Colloid cyst, although a benign tumor, is surgically challenging because of its deep midline location. Early detection and total excision of the colloid cyst carries an excellent prognosis. PMID- 12128299 TI - Colloid cysts of the third ventricle: some comments. PMID- 12128300 TI - Results of chronic subthalamic nucleus stimulation for Parkinson's disease: a 1 year follow-up study. AB - BACKGROUND: Deep brain stimulation (DBS) has been established as an alternative approach for the treatment of advanced Parkinson's disease (PD). Recently, the subthalamic nucleus (STN) has been identified as the optimal target for DBS. METHODS: Thirty-eight patients have undergone surgery for advanced PD since 1996. They include 12 females and 26 males with a mean age of 55.6 years. The mean stage on the Hoehn and Yahr Scale was 3.5 (off condition). Electrodes (Medtronic DBS 31389) were stereotactically implanted into the STN bilaterally. Targeting was performed using computerized tomography (CT) scans and ventriculography (VG). After 4 days of external stimulation, permanent neurostimulators were implanted. Patients were evaluated preoperatively and 1, 6, and 12 months postoperatively. Evaluations were performed in defined on and off states using the Unified Parkinson's Disease Rating Scale (UPDRS) as well as the Hoehn and Yahr Scale, the dyskinesia scale, and the Activities of Daily Living (ADL) Scale. RESULTS: Significant improvement of all motor symptoms was found in all patients (UPDRS motor score 32/48 preoperatively versus 15/30 at 12-month follow-up, p < 0.001). Daily off-times were reduced by 35%. Dyskinesias also improved markedly (UPDRS IV: 3.2/3.1 [on/off] vs. 0.9/1.3 at 12 months follow-up). Postoperative L-dopa medication was adjusted (mean reduction: 53%). Complications occurred in two patients (5%) who developed infections, leading to system removal. Systems were replaced after 6 months. Two patients (5%) had a permanent worsening of a previously known depressive state and developed progressive dementia. CONCLUSIONS: TN stimulation is a relatively safe procedure for treating advanced PD. The possibility of readjusting the stimulation parameters postoperatively improves the therapeutic outcome and reduces side effects in comparison to ablative methods. PMID- 12128306 TI - Carotid endarterectomy in patients with contralateral internal carotid artery occlusion without intraoperative shunting. AB - BACKGROUND: Controversy about the optimal method of performing a carotid endarterectomy (CEA) exists despite its widespread application and support from various randomized clinical trials. Many surgeons selectively or routinely use electroencephalography (EEG) monitoring as well as shunting when performing this operation. METHODS: We conducted this retrospective study to assess the maximum carotid clamp time without shunting or EEG monitoring during a CEA without the development of neurological deficits in an already compromised cerebral circulation. RESULTS: Fifteen consecutive patients who underwent CEAs between 1988 and 1999 met our criteria of angiographically documented ipsilateral internal carotid artery (ICA) stenosis with contralateral ICA occlusion. The patient presentations included asymptomatic (14%), transient ischemic attack (TIA) (50%), and stroke (36%). All patients were operated under general anesthesia without shunting and only 4 patients underwent EEG monitoring. On angiography, all 15 patients had ipsilateral ICA stenosis (70-99%) and contralateral occlusion. In 54% of patients, the vertebral arteries (VAs) were both patent, while in 46% of patients only 1 VA was patent. Eighty-five percent of patients had at least 1 patent anterior communicating (Pcomm) artery, while 15% had nonvisualized Pcomm arteries bilaterally. Of the 15 patients, 14 had a patent anterior communicating artery. The mean clamp time of the CCA was 18.5 minutes (range 14-30 minutes). None of the 15 patients had new neurological changes immediately postoperatively or during the 6 weeks of follow-up. CONCLUSION: We propose that shunting may not be necessary during CEA for high grade stenosis with contralateral ICA occlusion, presumably because of adequate distal small vessel collaterals. PMID- 12128303 TI - Bilateral supraorbital keyhole approach for multiple aneurysms via superciliary skin incisions. AB - BACKGROUND: Considering that multiple aneurysms carry a high risk for fatal rupture, there is a need for complete treatment of all lesions in one surgical session using either unilateral-contralateral or bilateral approaches. Contralateral approaches have been used mainly for small anteriorly projecting middle cerebral and medially expanding ophthalmic types of aneurysms. They are limited by the narrow space for surgical manipulation, forced elevation of frontal lobes, and stretching of the olfactory nerves. These problems might result in damage to structures along the unusually long intracranial way of the approach. The complications associated with the unnecessarily large conventional fronto-temporal and bifrontal craniotomies, and the developments in visualization, neuroanaesthesia, microneurosurgery, cerebrospinal fluid (CSF) drainage, and brain protection have led to less invasive methods in cerebral base surgery. These achievements have supplied the background for the supraorbital keyhole approach to aneurysms of the anterior circulation or basilar tip. Because the supraorbital keyhole approach offers several advantages over the classic fronto-temporal craniotomies to the anterior skull base, it was extended for both sides in one surgical session to treat bilateral multiple aneurysms as well. METHODS: Out of a series of 150 patients harboring 188 saccular aneurysms operated on via a supraorbital keyhole approach with a superciliar skin incision, 36 had multiple aneurysms. Thirty patients with multiple aneurysms underwent surgery for their ruptured aneurysms (17 cases in the acute phase and 13 patients during the chronic stage); in 6 cases silent aneurysms were operated on. The multiple aneurysms were managed from one side in 18 cases. A bilateral supraorbital keyhole approach was performed during one surgical session in 11 patients, and in 7 cases the unilateral supraorbital keyhole approach was combined with contralateral fronto-temporal (3 cases), suboccipital (2 cases), or frontal-parasagittal (2 cases) exploration. The operations were carried out through an approximately 2.5 x 3 cm supraorbital keyhole craniotomy following a skin incision just above the eyebrow. The roughly 4 cm superciliar skin incision begins medial to the supraorbital nerve and ends 3 to 10 mm beyond the lateral edge of the eyebrow. The technical details of the method are presented, and the benefits, limitations, and complications are discussed. RESULTS: In the 36 patients operated on via the supraorbital keyhole approach 74 aneurysms were clipped successfully. In 2 cases premature intraoperative rupture of the aneurysms occurred, but these events were managed successfully. Despite the small size of the craniotomy the approach allows enough room for intracranial manipulation with maximal protection of the brain and other intracranial structures. One patient died because of pulmonary embolism. There were no craniotomy-related complications in the present series. CONCLUSION: The supraorbital keyhole approach together with the advent of the modern neuroanaesthesia, CSF drainage, and microsurgical techniques is a safe approach in the hands of experienced neurosurgeons for the treatment of supratentorial or basilar tip aneurysms. Because the approach is simple and swift, the bilateral single-session craniotomy does not have any disadvantages compared to two-stage procedures. However, the one-sitting surgery reduces the high risk of fatal rupture in the perioperative period associated with multiple aneurysms. PMID- 12128309 TI - Assessment of cerebrovascular reserve capacity in asymptomatic and symptomatic hemodynamically significant carotid stenoses and occlusions. AB - BACKGROUND: Cerebrovascular reactivity measurements are believed to be a helpful tool for selecting patients who are at higher risk for hemodynamic strokes. The aim of this study was to compare cerebral vasoreactivity among patients suffering from internal carotid artery stenosis of different severity (asymptomatic stenosis, asymptomatic occlusion, symptomatic stenosis, symptomatic occlusion). METHODS: Sixty-two patients with asymptomatic and symptomatic internal carotid artery stenoses and occlusions underwent transcranial Doppler-acetazolamide tests. Absolute velocities of the middle cerebral arteries (MCAV), percent increases of the MCAV at different time points of the test (cerebrovascular reactivity, CVR) and maximal percent increase after administration of acetazolamide (cerebrovascular reserve, CRC) were compared on the affected and non-affected sides. Asymmetry indices (CRC (affected side)/CRC (non-affected side)) were compared between the groups of different severity of obstructive lesion. RESULTS: Resting MCAV was similar on both sides in all groups. A significant side-difference of the MCAV values after acetazolamide was observed only in the symptomatic groups. Difference of cerebrovascular reserve capacity between the affected and non-affected side was statistically significant only in the symptomatic groups (CRC symptomatic stenosis 36.6 +/- 20.9% vs. 71.1 +/- 27.9%, CRC symptomatic occlusion: 31.2 +/- 24.6% vs. 64.5 +/- 29.7%). Asymmetry index of the CRC was near to 1 in the asymptomatic stenosis group only, while in all the other groups this index referred to a significant hemispheric asymmetry of the vasoreactivity. CONCLUSIONS: Although in general cerebrovascular reserve capacity is compromised in cases of hemodynamically significant carotid lesions, there is a large individual variability within the subgroups. Further randomized studies are needed to clarify whether the clinical efficiency of carotid endarterectomy and extra-intracranial bypass may be improved by selecting the patients using hemodynamic criteria. PMID- 12128311 TI - Multiple cavernomas of brain presenting with simultaneous hemorrhage in two lesions: a case report. AB - BACKGROUND: Cavernomas are rare vascular lesions of the brain that can bleed. However, the risk of bleeding is lower than that of aneurysms or arteriovenous malformations. In selected cases, bleeding cavernomas require surgical management. Presented here is a case of multiple cavernomas of the brain with simultaneous bleeding in two different lesions, along with its management. Although multiple cavernomas have been described in the literature, simultaneous bleeding in two different lesions is rare. CASE DESCRIPTION: A 52-year-old woman presented with difficulty maintaining balance, double vision, and slurred speech. She had had multiple surgeries for cavernous angioma of the brain in the past. Examination and investigations revealed two cavernomas, one in the dorsal midbrain region and one in the left occipital region. Her clinical condition deteriorated suddenly, and further evaluation revealed bleeding in both the cavernomas. The lesion in the midbrain was removed surgically. Since the lesion was in the posterior midbrain, a posterior interhemispheric approach was used with the help of frameless stereotactic navigation. Total excision was achieved. The lesion in the occipital lobe was not operated on. The patient had an uneventful recovery. It was planned to observe the progress of the occipital cavernoma by serial magnetic resonance imaging scans. CONCLUSION: Although there may be simultaneous bleeding in two or even more lesions, surgical treatment should be undertaken for the lesions jeopardizing critical structures or exerting mass effects. Additionally, it was discovered that frameless stereotactic navigation was of immense help in delineating the lesion and for safe excision of the lesion in critical areas. PMID- 12128313 TI - Middle cerebral artery dissecting aneurysm causing intracerebral hemorrhage 4 years after the non-hemorrhagic onset: a case report. AB - BACKGROUND: We report a case of dissecting middle cerebral artery (MCA) aneurysm causing intracerebral hemorrhage 4 years after the non-hemorrhagic onset. CASE DESCRIPTION: A 30-year-old male was diagnosed with an unruptured dissecting MCA aneurysm after a severe pulsating headache in August 1993. Angiography revealed dilatation of the distal portion of the temporo-occipital artery. During 2 years of follow-up, there were no significant changes on magnetic resonance imaging. In August 1997, he became comatose because of massive intracerebral hemorrhage caused by a dilated fusiform dissecting aneurysm. Emergency surgery and postoperative mild hypothermia resulted in full recovery. CONCLUSION: To date only 30 cases of dissecting MCA aneurysm have been reported and for unruptured aneurysms, surgical intervention was not chosen. However, the present case strongly suggests that long-term follow-up is necessary in patients with unruptured dissecting aneurysms of the anterior circulation, especially those with ectatic components. PMID- 12128316 TI - Management of hydrocephalus associated with occipital encephalocoele using endoscopic third ventriculostomy: report of two cases. AB - BACKGROUND: Occipital encephalocoele is the most common cranial dysraphism in the western hemisphere and is often complicated by hydrocephalus. Management of hydrocephalus and reducing the CSF pressure is crucial in preventing dehiscence at the site of the encephalocoele repair. METHODS: Two female patients had presented with occipital encephalocoeles. The first patient (aged 42 days) had undergone repair of the occipital encephalocoele and then developed hydrocephalus with recurrence of the encephalocoele. The second patient (aged 12 months) had hydrocephalus associated with an occipital encephalocoele at initial presentation.Both the patients underwent endoscopic third ventriculostomy (ETV) through a right frontal burr hole. In the first patient, ETV was performed after shunt dysfunction at the age of 9 months. Because she presented with recurrence of the encephalocoele 15 months later, a repeat endoscopic third ventriculostomy was performed. She required a ventriculoperitoneal shunt during the same admission because of the early failure of the ventriculostomy. In the second patient, it was performed before the encephalocoele repair, both ETV and the repair being conducted under the same anesthesia. ETV was performed using a rigid scope and the perforation in the third ventricular floor was enlarged using a No. 4 Fogarty catheter. RESULTS: The first patient had no recurrence of encephalocoele at follow-up of 10 months but she presented with recurrence of the occipital encephalocoele after 15 months. The second patient had no evidence of recurrence at follow-up after 16 months. The lateral and third ventricular volumes had decreased in both the patients at initial follow-up. CONCLUSION: ETV can be an effective treatment option for encephalocoele-associated hydrocephalus, even in children under the age of 1 year. It may obviate the need for placement of CSF shunts that have a risk of infection and dysfunction. However, delayed failure of ETV may occur as seen in our first patient, indicating the need for careful and long-term follow-up. PMID- 12128320 TI - The neurosurgery-biomedical industrial complex. PMID- 12128318 TI - Microsurgical treatment for hypothalamic hamartoma in children with precocious puberty. AB - BACKGROUND: We review the surgical treatment of hypothalamic hamartoma causing precocious puberty. METHODS: Six children (three girls and three boys) with precocious puberty secondary to hypothalamic hamartoma were recruited for our study. The mean age of the patients was 30 months old (range 13 months to 5 years), and the mean age of the onset of puberty was 7.3 months. All patients were treated by microsurgery. RESULTS: All patients had higher then normal stature, body weight, bone growth, and serum levels of sexual hormones. The boys presented with mature external genitalia, pubic hair, frequent erection, and acne, while the girls presented with growth of breasts and menarche. Magnetic resonance image (MRI) revealed an isointense mass below the tuber cinereum extending into the supersellar and interpeduncular cistern, ranging from 4 to 12 mm in diameter, consistent with pedunculate hamartoma. The hamartoma was removed completely via a right pterional approach. The symptoms and signs of precocious puberty resolved completely, and sexual hormone levels decreased to the pre pubertal range in all six patients without any postoperative complications. CONCLUSION: We report a series of six children with hypothalamic hamartoma induced precocious puberty who underwent microsurgical treatment. All of them recovered completely to their age-appropriate state. Microsurgery is a good choice of treatment for pedunculate hypothalamic hamartoma. PMID- 12130397 TI - Effect of pitch type, pitch count, and pitching mechanics on risk of elbow and shoulder pain in youth baseball pitchers. AB - BACKGROUND: Joint pain is thought to be an early sign of injury to a pitcher. OBJECTIVE: To evaluate the association between pitch counts, pitch types, and pitching mechanics and shoulder and elbow pain in young pitchers. STUDY DESIGN: Prospective cohort study. METHODS: Four hundred and seventy-six young (ages 9 to 14 years) baseball pitchers were followed for one season. Data were collected from pre- and postseason questionnaires, injury and performance interviews after each game, pitch count logs, and video analysis of pitching mechanics. Generalized estimating equations and logistic regression analysis were used. RESULTS: Half of the subjects experienced elbow or shoulder pain during the season. The curveball was associated with a 52% increased risk of shoulder pain and the slider was associated with an 86% increased risk of elbow pain. There was a significant association between the number of pitches thrown in a game and during the season and the rate of elbow pain and shoulder pain. CONCLUSIONS: Pitchers in this age group should be cautioned about throwing breaking pitches (curveballs and sliders) because of the increased risk of elbow and shoulder pain. Limitations on pitches thrown in a game and in a season can also reduce the risk of pain. Further evaluation of pain and pitching mechanics is necessary. PMID- 12130398 TI - Biomechanical comparison of patellar tendon repairs in a cadaver model: an evaluation of gap formation at the repair site with cyclic loading. AB - BACKGROUND: Ruptures of the patellar tendon are rare injuries. Surgical treatment for this injury is mandatory. HYPOTHESIS: Gap formation does not differ between the three patellar tendon repair techniques. STUDY DESIGN: Controlled laboratory study. METHODS: Twelve fresh-frozen cadaveric knees were used to compare three techniques of patellar tendon repairs. The standard suture repair used two Krackow sutures placed in the avulsed patellar tendon, passed through transpatellar drill holes, and secured with the knee in 30 degrees of flexion. In the second group, suture repair was augmented with a No. 5 Ethibond suture. In the third group, suture repair was augmented with a 2.0 Dall-Miles cable. Testing was performed with the specimens mounted to a custom knee jig with the tibia free, simulating the knee moment of a 70-kg person. Each knee was then cycled 250 times at 0.25 Hz. RESULTS: Gap formation across the standard suture repair averaged 7.3 mm; across the suture augmentation and cable augmentation groups it averaged 4.9 mm and 3.5 mm, respectively. CONCLUSIONS: Augmentation of patellar tendon avulsions can decrease gap formation at the repair site, allowing early mobilization. CLINICAL RELEVANCE: Gap formation seen in repair without augmentation could lead to clinical failure with resultant patella alta and extensor mechanism lag. PMID- 12130399 TI - Arthroscopic surgery for isolated capitellar osteochondritis dissecans in adolescent baseball players: minimum three-year follow-up. AB - BACKGROUND: Osteochondritis dissecans of the capitellum of the humerus usually occurs in adolescence and is caused by the valgus forces associated with excessive throwing. HYPOTHESIS: Arthroscopic surgery is an appropriate procedure for this condition. STUDY DESIGN: Retrospective cohort study. METHODS: Arthroscopic surgery was performed on 10 baseball players (average age, 13.8 years) with osteochondritis dissecans whose symptoms had been apparent for an average of 9 months before the operation. Follow-up at an average of 3.9 years included use of a standard rating scale, radiographs, and a questionnaire regarding return to sport. RESULTS: There were two grade I, one grade II, two grade IV, and five grade V lesions. Symptoms and objective findings correlated poorly with the grade of the lesion. The postoperative score averaged 195, reflecting excellent results. Radiographically, the primary lesion was still apparent in one patient, secondary degenerative changes were evident in one patient, and, in one patient, the lesion was still evident and degenerative changes had occurred. Only four athletes returned to organized baseball. CONCLUSIONS: Arthroscopic surgery for symptomatic osteochondritis dissecans of the capitellum in adolescent baseball players can provide excellent rating scores with intermediate follow-up but does not assure return to baseball. PMID- 12130400 TI - Reliability of heel-height measurement for documenting knee extension deficits. AB - BACKGROUND: Heel-height difference has been used to detect subtle knee flexion contractures, but the effects of thigh circumference differences and patient positioning during testing have not been evaluated. HYPOTHESIS: Differences in thigh circumference measurements and whether the patient's patellae are on or off the examination table during heel-height difference measurement will not affect the accuracy of detecting knee flexion contracture. STUDY DESIGN: Prospective cohort study. METHODS: Bilateral knee range of motion, prone heel-height difference with the patellae on and off the table, and thigh circumference at 5 and 15 cm proximal to the proximal pole of the patella were measured by one investigator on 50 consecutive patients who had undergone unilateral anterior cruciate ligament reconstruction. RESULTS: A high degree of correlation was demonstrated between the heel-height difference and the standard range of motion measurement. Differences in thigh girth and patellar position did not statistically affect the accuracy of the heel-height difference as an indicator of knee flexion contracture. CONCLUSION: Heel-height difference is a valid method of documenting knee flexion contractures. Compared with traditional goniometer assessment, this test is a more meaningful and easier way for detecting subtle knee flexion contractures of less than 10 degrees. PMID- 12130401 TI - Motor control of the vastus medialis oblique and vastus lateralis muscles is disrupted during eccentric contractions in subjects with patellofemoral pain. AB - BACKGROUND: Inappropriate control of the vastus medialis oblique and vastus lateralis muscles by the central nervous system can contribute to maltracking of the patella. HYPOTHESIS: The activation timing and amplitude of the vastus medialis oblique and vastus lateralis muscles will be different between normal subjects and patients with patellofemoral pain. STUDY DESIGN: Controlled laboratory study. METHODS: Subjects with patellofemoral pain and asymptomatic control subjects performed maximum voluntary knee extension contractions initiated from a flexed and an extended position. The activation timing and amplitude of the vastus lateralis and vastus medialis oblique muscles were quantified from the recorded electromyographic signals. RESULTS: There were no between-group differences in activation timing. The activation amplitude of the vastus medialis oblique and vastus lateralis muscles of the patellofemoral pain subjects was altered to the greatest extent during eccentric contractions and differed significantly from that of control subjects. CONCLUSIONS: The activation amplitudes of the vastus medialis oblique and vastus lateralis muscles of subjects with patellofemoral pain are consistent with a laterally tracking patella during eccentric contractions. CLINICAL RELEVANCE: The findings suggest the clinical importance of determining whether altered activation patterns are sensitive to rehabilitation, and, if so, if subjective reports of knee joint pain and function parallel changes in the activation patterns as a result of rehabilitation. PMID- 12130402 TI - Significance of ultrasonographically detected asymptomatic tendinosis in the patellar and achilles tendons of elite soccer players: a longitudinal study. AB - BACKGROUND: Chronic tendinosis of the Achilles or patellar tendons, or both, is one of the most frequent and severe conditions that affects athletes in sports such as soccer. It can often end an athlete's sports activity. HYPOTHESIS: Ultrasonography of asymptomatic tendons can be used to predict which athletes will develop tendon symptoms. STUDY DESIGN: Longitudinal study. METHODS: Using ultrasonography of the ankle and knee, we examined 54 elite soccer players in the top Danish soccer league before and after a single season (in January and December). RESULTS: At the start of the season, ultrasonography had revealed abnormalities in 29% of those examined. Eighteen percent (18 of 98 tendons) were observed to have abnormal sonographic findings in the patellar tendon at the initial examination in January. These athletes were found to have a 17% risk of developing symptomatic jumper's knee during the 12-month season. Eleven percent (11 of 96 tendons) were observed to have abnormal sonographic findings in the Achilles tendon at the initial examination; it was calculated that they had a 45% risk of developing symptoms of Achilles tendinosis. Only one of the players with normal tendons in January actually developed symptoms by the end of the season. CONCLUSIONS: For the first time it is now possible to identify risk factors for the development of serious tendon disorders in asymptomatic athletes. Future studies may be directed at developing preventive treatment to reduce the risk of chronic, therapy-resistant symptoms of tendinosis and ruptures. PMID- 12130403 TI - A biomechanical analysis of meniscal repair techniques. AB - BACKGROUND: Various methods are available for repair of meniscal tears: a biodegradable meniscal implant without sutures (Biofix meniscus arrow), a suture anchor device (T-fix), and horizontal and vertical mattress sutures. HYPOTHESIS: There is no difference in repair strength or mode of failure among the techniques compared. STUDY DESIGN: Controlled laboratory study. METHODS: Reproducible tears were created in bovine menisci and repaired with each of the techniques. Residual displacement of the tear immediately after repair and the resistance to displacement under load applied perpendicular to the tear were measured and compared. RESULTS: The residual displacement after repair was highest in the Biofix arrow group (0.70 mm) and lowest in the vertical mattress suture group (0.21 mm). The ultimate strength of repair was strongest for the vertical sutures (202 +/- 7 N) and lowest for the arrow and T-fix (95.9 +/- 8 N and 99.4 +/- 8 N, respectively). The force required to generate 2 mm of tear displacement was greatest for the vertical sutures (143 N) and least for the arrow (43.6 N). CONCLUSIONS: Suture techniques were stronger at all levels of testing. CLINICAL RELEVANCE: Knowledge of biomechanical characteristics aids the surgeon in choosing the appropriate technique for each situation. PMID- 12130404 TI - The effect of graft-tunnel diameter disparity on intraosseous healing of the flexor tendon graft in anterior cruciate ligament reconstruction. AB - BACKGROUND: Graft-to-tunnel healing is a significant factor in anterior cruciate ligament reconstruction, but there have been few studies on the effect of graft tunnel diameter disparity on intraosseous healing of the flexor tendon graft. HYPOTHESIS: Graft-tunnel diameter disparity of 2 mm has no effect on the pull-out strength of the graft from the bone tunnel. STUDY DESIGN: Controlled laboratory study. METHODS: Forty-two beagle dogs were divided into three groups. In each animal, reconstruction was performed in the left knee by using a 4-mm diameter autogenous flexor tendon for groups 1 and 2 and by using a 4-mm wide bone patellar tendon-bone graft in group 3. A 4-mm diameter tunnel was drilled in the tibia of groups 1 and 3 and a 6-mm diameter tunnel, in group 2. In each group, seven animals were sacrificed at 3 and 6 weeks. RESULTS: The perpendicular fibers connecting the graft to the bone were generated in groups 1 and 2, and the number appeared to be higher in group 2, where the space was greater. There was no significant difference in the ultimate failure load between groups 1 and 2 at each period. CONCLUSION: Graft-tunnel diameter disparity of up to 2 mm may not adversely affect intraosseous healing of the flexor tendon graft. CLINICAL RELEVANCE: Surgeons need not be overly concerned about minor graft-tunnel diameter disparities. PMID- 12130405 TI - How four weeks of implantation affect the strength and stiffness of a tendon graft in a bone tunnel: a study of two fixation devices in an extraarticular model in ovine. AB - BACKGROUND: For a tendon graft to function as an anterior cruciate ligament, the tendon must heal to the bone tunnel. We studied the effect of 4 weeks of implantation on the strength and stiffness of a tendon in a bone tunnel using two different fixation devices in an ovine model. HYPOTHESIS: The type of fixation device in anterior cruciate ligament reconstruction may affect early healing, which can be measured as the strength and stiffness of a tendon in a bone tunnel. STUDY DESIGN: Controlled laboratory study. METHODS: An extraarticular tendon graft reconstruction was performed in ovine tibias. The graft was fixed with either a bioresorbable interference screw or a WasherLoc. After 4 weeks of implantation the strength and stiffness of the complex and the tendon graft-bone tunnel interface were determined by incrementally loading specimens to failure. RESULTS: For the interference screw, the strength deteriorated 63% and the stiffness deteriorated 40%. For the WasherLoc, the strength was similar and the stiffness improved 136%. CONCLUSIONS: The type of fixation device determines whether the strength and stiffness of a tendon in a bone tunnel increases or decreases after implantation. CLINICAL RELEVANCE: The pace of rehabilitation may need to be adjusted based on the type of fixation device used to secure a soft tissue graft. PMID- 12130406 TI - Relevance of arm position and muscle activity on three-dimensional glenohumeral translation in patients with traumatic and atraumatic shoulder instability. AB - BACKGROUND: No quantitative data on glenohumeral translation exist allowing one to distinguish insufficiency of the active or passive stabilizers in different forms of shoulder instability. HYPOTHESIS: To determine whether 1) in traumatic or atraumatic shoulder instability an increase of glenohumeral translation can be observed in specific relevant arm positions, 2) muscle activity leads to recentering of the humeral head, and 3) there exist differences between traumatic and atraumatic instability. STUDY DESIGN: Prospective clinical trial. METHODS: In 12 patients with traumatic and 10 patients with atraumatic instability, both shoulders were examined in different arm positions-with and without muscle activity-by using open magnetic resonance imaging and a three-dimensional postprocessing technique. RESULTS: At 90 degrees of abduction and external rotation, translation (anterior-inferior) was significantly higher in patients with traumatic unstable shoulders compared with their contralateral side (3.6 +/- 1.5 versus 0.7 +/- 1.6 mm). In patients with atraumatic instability, significantly increased translation (4.7 +/- 2.0 mm) was observed, with the direction being nonuniform. Muscle activity led to significant recentering in traumatic but not in atraumatic instability. CONCLUSIONS: In traumatic instability, increased translation was observed only in functionally important arm positions, whereas intact active stabilizers demonstrate sufficient recentering. In atraumatic instability, a decentralized head position was recorded also during muscle activity, suggesting alterations of the active stabilizers. CLINICAL RELEVANCE: These data are relevant for optimizing diagnostics and therapeutic strategies. PMID- 12130407 TI - A five-year comparison of patellar tendon versus four-strand hamstring tendon autograft for arthroscopic reconstruction of the anterior cruciate ligament. AB - BACKGROUND: The choice of graft material for anterior cruciate ligament reconstruction is believed to play a major role in outcome, but most comparisons of graft choice have not been well controlled. HYPOTHESIS: The choice of graft material (patellar tendon or hamstring tendon) does affect clinical outcome after anterior cruciate ligament reconstruction. STUDY DESIGN: Prospective, nonrandomized clinical trial. METHODS: Two groups of 90 patients each were followed for a minimum of 5 years. RESULTS: International Knee Documentation Committee assessment revealed that more than 85% of each group had an overall score of A or B at all follow-up intervals. The median Lysholm knee score was greater than 90 for both groups at 2 and 5 years. Instrumented testing revealed no significant difference between the two groups beyond 3 years. Thirty-one percent of the patellar tendon group (25) had a fixed flexion deformity and 19% of the hamstring tendon group (14) had fixed flexion deformity at 5 years. Radiologic assessment revealed early osteoarthritic changes in 4% of the hamstring tendon group (2) and in 18% of the patellar tendon group (11) at 5 years. CONCLUSIONS: Arthroscopic reconstruction with either graft results in a similar surgical outcome, reliably restoring knee stability over a 5-year period; however, patients with patellar tendon grafts are at greater risk of developing early signs of osteoarthritis. PMID- 12130408 TI - Injury to the anterior cruciate ligament during alpine skiing: a biomechanical analysis of tibial torque and knee flexion angle. AB - BACKGROUND: The anterior cruciate ligament has been shown to be particularly susceptible to injury during alpine skiing. Tibial torque is an important injury mechanism, especially when applied to a fully extended or fully flexed knee. PURPOSE: We wanted to record the forces generated in the anterior cruciate ligament with application of tibial torque to cadaveric knees in different positions. STUDY DESIGN: Controlled laboratory study. METHODS: Thirty-seven fresh frozen cadaveric knees were instrumented with a tibial load cell that measured resultant force in the anterior cruciate ligament while internal and external tibial torques were applied to the tibia at full extension, 90 degrees of flexion, full flexion, and forced hyperflexion. RESULTS: At each knee flexion position, mean force generated by 10 N.m of internal tibial torque was significantly higher than the mean generated by 10 N.m of external tibial torque. Mean forces generated by tibial torque at 90 degrees of flexion were relatively low. During flexion-extension without tibial torque applied mean forces were highest (193 N) when the knee was hyperflexed. CONCLUSIONS: Application of internal tibial torque to a fully extended or fully flexed knee represents the most dangerous loading condition for injury from twisting falls during skiing. CLINICAL RELEVANCE: Understanding of the mechanisms of falls can be used to design better equipment and to better prevent or treat injury. PMID- 12130409 TI - Medial collateral ligament reconstruction of the elbow using the docking technique. AB - BACKGROUND: Medial collateral ligament insufficiency of the elbow with resultant valgus instability in throwing athletes is typically treated with free tendon graft reconstruction as described by Jobe. HYPOTHESIS: Improved results could be obtained with the use of the docking technique. STUDY DESIGN: Uncontrolled retrospective review. METHODS: The study group consisted of 36 athletes who had symptomatic insufficiency of the medial collateral ligament confirmed by magnetic resonance imaging and by surgical findings. Average follow-up was 3.3 years. Key elements of the docking technique included a muscle-splitting approach without routine transposition of the ulnar nerve, routine arthroscopic assessment, treatment of associated lesions, and docking the two ends of the tendon graft into a single humeral tunnel. RESULTS: Thirty-three of 36 patients (92%) returned to or exceeded their previous level of competition for at least 1 year, meeting the Conway-Jobe classification criteria of "excellent." All 22 professional or collegiate athletes returned to or exceeded their previous competition level. CONCLUSIONS: The docking technique allowed simplified graft tensioning and improved graft fixation. PMID- 12130410 TI - Bone-patellar tendon-bone grafts for anterior cruciate ligament reconstruction: an in vitro comparison of mechanical behavior under failure tensile loading and cyclic submaximal tensile loading. AB - BACKGROUND: Secure fixation is an important factor in the success of anterior cruciate ligament reconstruction. HYPOTHESIS: There is no difference in the mechanical behavior of reconstructions from method of fixation or method of testing. STUDY DESIGN: Controlled laboratory study. METHODS: Anterior cruciate ligament reconstructions were performed with bone-patellar tendon-bone grafts in 48 human cadaveric knees. Three different fixation methods (Endobutton, interference screw, suture-post fixation) were compared under failure tensile loading and cyclic submaximal tensile loading. RESULTS: No difference was observed in ultimate load among the three techniques. Stiffness of the grafts was significantly lower for the suture technique than for the interference screw technique. Cyclic loading revealed significantly different failure rates: 0% of the Endobutton, 38% of the interference screw, and 100% of the suture-post groups. The relative movement of the femoral bone plug and the migration of the bone plug out of the femoral canal were lowest in the interference screw group. CONCLUSIONS: The suture-post fixation is not recommended. The interference screw technique showed the best results, but results were age-dependent, suggesting its best use is in younger patients. The Endobutton technique is recommended for older patients. CLINICAL RELEVANCE: Results of testing are useful to the surgeon in making a choice of reconstruction technique. PMID- 12130411 TI - Rotator cuff tears in middle-aged tennis players: results of surgical treatment. AB - BACKGROUND: Tennis players, like participants in other overhead sports, are vulnerable to rotator cuff tears. In players who continue to play into their middle-age years, the incidence of such injury increases. HYPOTHESIS: Surgical treatment of rotator cuff tears in middle-aged tennis players is largely successful in allowing return to tennis. STUDY DESIGN: Retrospective review. METHODS: We evaluated the results of surgical treatment of 51 middle-aged tennis players (average age, 51 years) with a rotator cuff tear in their dominant shoulder. Tennis participation among the group had averaged 3.5 hours per week for an average of 25 years. Forty-two patients underwent open repair of the tear with or without biceps tenodesis, whereas 9 patients underwent arthroscopic debridement of the tear with or without a biceps tenotomy. Patients were reviewed at an average of 57 months after surgery with an activities score, a subjective questionnaire, and a questionnaire regarding their postoperative participation in tennis. RESULTS: The activities score averaged 26.6 of 30 possible points. Forty seven patients were satisfied with their result, and 40 patients were able to return to tennis at an average of 9.8 months after surgery. No difference was found in the ability to return to tennis between the open repair group and the arthroscopic debridement group. CONCLUSIONS: The results of this study indicate that it is possible for nearly 80% of middle-aged tennis players to return to participation after operative treatment of rotator cuff tears. PMID- 12130412 TI - Quantitative review of operative and nonoperative management of achilles tendon ruptures. AB - BACKGROUND: There is no consensus on the best method for management of acute Achilles tendon ruptures. Individual preferences, drawn from experience and study, determine whether treatment is operative or nonoperative. PURPOSE: Our goal was to review the literature to try to determine what management method was the most popular and effective. We wanted to ascertain the best results in terms of complication rates and patient outcomes. STUDY DESIGN: Retrospective review of retrospectively and prospectively collected data. METHODS: We analyzed 125 articles in peer-reviewed journals for year of publication, patient numbers, sex, management method, follow-up complications, and patient satisfaction. Each article was graded using a validated methods score. Methods, patient satisfaction, and complication rates were correlated with the year each article was published. RESULTS: Skin-healing complications were lowest in conservatively managed patients (3 of 578, 0.5%) and highest in open repair and immobilized patients (543 of 3718, 14.6%). General complication rates were lowest in open repair and early-mobilization groups (16 of 238, 6.7%) and highest in percutaneous and early-mobilization groups (19 of 122, 15.6%). Rerupture rates were highest in immobilized conservative management groups (62 of 578, 10.7%) and lowest in groups with external fixation (0%). CONCLUSIONS: In general, the number of publications reporting Achilles tendon ruptures is increasing, the quality of articles is increasing, and the trend for the number of reported complications is decreasing. The published articles had a low methods score (mean, 50.9; range, 25 to 77) and showed a trend toward earlier mobilization. Open repair and early mobilization give the best functional recovery and an acceptable complication rate. PMID- 12130413 TI - A prospective, randomized evaluation of arthroscopic stabilization versus nonoperative treatment in patients with acute, traumatic, first-time shoulder dislocations. AB - BACKGROUND: Nonoperative treatment of traumatic shoulder dislocations leads to a high rate of recurrent dislocations. HYPOTHESIS: Early arthroscopic treatment for shoulder dislocation will result in a lower recurrence rate than nonoperative treatment. STUDY DESIGN: Prospective, randomized clinical trial. METHODS: Two groups of patients were studied to compare nonoperative treatment with arthroscopic Bankart repair for acute, traumatic shoulder dislocations in young athletes. Fourteen nonoperatively treated patients underwent 4 weeks of immobilization followed by a supervised rehabilitation program. Ten operatively treated patients underwent arthroscopic Bankart repair with a bioabsorbable tack followed by the same rehabilitation protocol as the nonoperatively treated patients. The average follow-up was 36 months. RESULTS: Three patients were lost to follow-up. Twelve nonoperatively treated patients remained for follow-up. Nine of these (75%) developed recurrent instability. Six of the nine have required subsequent open Bankart repair for recurrent instability. Of the nine operatively treated patients available for follow-up, only one (11.1%) developed recurrent instability. CONCLUSIONS: Arthroscopic stabilization of traumatic, first-time anterior shoulder dislocations is an effective and safe treatment that significantly reduces the recurrence rate of shoulder dislocations in young athletes when compared with conventional, nonoperative treatment. PMID- 12130414 TI - Long-term outcome of fasciotomy with partial fasciectomy for chronic exertional compartment syndrome of the lower leg. AB - BACKGROUND: Fasciotomy with partial fasciectomy for compartment syndrome has had good short-term results, but no long-term studies have been performed. HYPOTHESIS: Combining a partial fasciectomy with fasciotomy for compartment syndrome relieves pain and eliminates symptoms in the long term. STUDY DESIGN: Retrospective cohort study. METHODS: A self-administered questionnaire was given to 62 patients at a mean follow-up of 51 months after surgery. RESULTS: Of the 50 patients who underwent a single operation, 60% (30) reported an excellent or good outcome. Average pain and pain-on-running were significantly reduced, although some subjects still reported considerable levels of pain. Fifty-eight percent (36 of 62) were exercising at a lower level than before injury and, of these, 36% (13) cited the return of their compartment syndrome or the development of a different lower leg compartment syndrome as the reason for a reduction in exercise levels. Some subjects indicated early initial improvement followed by subsequent deterioration. CONCLUSION: This surgical technique reduces pain and allows the majority of patients to return to sports; however, patients should be counseled that they may not be able to return to their preinjury level of exercise or remain pain-free. PMID- 12130415 TI - Arthroscopic repair of meniscal tears extending into the avascular zone in patients younger than twenty years of age. AB - BACKGROUND: Limited data are available regarding repair results of meniscal tears extending into the central avascular region. HYPOTHESIS: Meniscal tears extending into the avascular region can be successfully repaired in patients less than 20 years old. STUDY DESIGN: Prospective cohort study. METHODS: We examined the results of 71 meniscal repairs (64 knees) for tears extending into the central avascular region in patients 19 years of age or younger; 67 were examined clinically (mean, 51 months after surgery) and 36, by follow-up arthroscopy (mean, 18 months). RESULTS: In 53 of 71 (75%) meniscal repairs patients had no tibiofemoral compartment symptoms and there were no clinical failures. In 18 (25%) meniscal repairs, patients showed tibiofemoral symptoms or a failed repair was detected on follow-up arthroscopy. In the subgroup of 45 knees with meniscal repair and anterior cruciate ligament reconstruction evaluated clinically, 39 (87%) patients rated their knee as normal or very good, 2 (4%) as good, 3 (7%) as fair, and 1 (2%) as poor. CONCLUSIONS: A stable repair of complex meniscal tears that extend into the avascular region can be obtained using a meticulous inside out vertical divergent suture technique. We recommend repair, particularly in young active patients in whom removal of complex tears would result in major loss of meniscal function and the risk of future arthrosis. PMID- 12130416 TI - Prevention of axillary nerve injury in anterior shoulder reconstructions: use of a subscapularis muscle-splitting technique and a review of the literature. AB - BACKGROUND: Previous authors have suggested that the axillary nerve should be explored or palpated during all anterior shoulder stabilization procedures. OBJECTIVE: The goal of this study was to document the axillary nerve injury rate in a cohort of patients who had undergone anterior shoulder stabilization without axillary nerve dissection. HYPOTHESIS: Use of a subscapularis muscle-splitting approach by using a retractor along the scapular neck does not result in significant risk of injury to the axillary nerve, and exploration of the axillary nerve is not necessary using this approach. STUDY DESIGN: Prospective cohort study. METHODS: One hundred and twenty-eight anterior stabilizations were performed with a subscapularis muscle-splitting approach that has been previously described. In all cases a retractor was placed along the inferior scapular neck to protect the axillary nerve. The axillary nerve was not exposed or palpated in any case. All patients were evaluated on the 1st postoperative day and again within 10 days for symptoms of axillary nerve palsy, including sensory loss and return of muscle function. One patient (0.8%) had paresthesia in an axillary nerve distribution; recovery occurred without the need for electromyography or other interventions. There were no clinically detected cases of axillary nerve motor dysfunction. CONCLUSIONS: Routine exposure of the axillary nerve is not necessary during anterior stabilization procedures using a subscapularis muscle splitting approach if proper precautions are taken to protect the nerve. Other techniques of anterior stabilization may require exposure of the axillary nerve. PMID- 12130417 TI - The surgical treatment of internal snapping hip. AB - BACKGROUND: Internal snapping hip is an underdiagnosed cause of hip pain that sidelines many recreational and competitive athletes. It originates from a taut iliopsoas tendon that snaps across bony prominences when the hip is extended from a flexed position. When nonoperative treatment methods fail, fractional tendon lengthening procedures may be used. HYPOTHESIS: Surgical tendon lengthening through a true ilioinguinal approach, which has not been previously reported, will achieve good results in patients with internal snapping hip. STUDY DESIGN: Retrospective cohort study. METHODS: In 30 patients with symptoms in their anterior hip, internal snapping hip was diagnosed by history and physical examination. All patients were initially treated nonoperatively; 19 (63%) improved and did not require further intervention. Eleven patients (12 hips) whose symptoms were recalcitrant to physical therapy were offered the surgical option of iliopsoas tendon lengthening. The procedure was performed via an ilioinguinal intrapelvic approach. Patients were followed up for an average of 3 years. RESULTS: All 11 surgically treated patients (100%) had complete postoperative mitigation of their snapping hip. Nine (82%) reported excellent pain relief. Moreover, nine patients thought that they had greatly benefited from the tendon lengthening and would repeat the surgery. CONCLUSION: Although nonoperative measures are usually successful in the treatment of internal snapping hip, surgical tendon lengthening is a viable approach in cases refractory to nonoperative therapy. PMID- 12130418 TI - Bilateral radial nerve compression syndrome in an elite swimmer: a case report. PMID- 12130419 TI - Neurovascular complications of knee arthroscopy. AB - During the last 3 decades, arthroscopy has revolutionized the way knee surgery is performed. The indications and the applications of arthroscopic procedures in the knee joint have enormously increased with the improvement in surgical technique and advent of new arthroscopic equipment. The use of arthroscopic techniques has led to a significant decrease in morbidity for the patient with intraarticular abnormalities, in terms of both diagnosis and surgical correction. Even though knee arthroscopy is a minimally invasive procedure with relatively low morbidity, it is not without risk of complications, of which neurovascular complications are among the most serious and devastating. The reported incidence of neurovascular complication is low, but it may be underestimated. Many neurovascular complications that occur are preventable with a thorough understanding of neurovascular anatomy, good preoperative and intraoperative planning, and attention to the details of basic techniques and the equipment used for the procedure. It is imperative that the surgeon who is performing arthroscopy be aware of these neurovascular complications, recognize them as early as possible, and initiate further evaluation and treatment as expeditiously as possible. In this article, the causes, management, prevention, and medicolegal implications of neurovascular complications of knee arthroscopy are reviewed. PMID- 12130420 TI - No evidence of impaired neurocognitive performance in collegiate soccer players. PMID- 12130421 TI - Creep behavior of a rabbit model of ligament laxity after electrothermal shrinkage in vivo. PMID- 12130422 TI - Dose reduction in CT: how low can we go? PMID- 12130424 TI - Multiplanar and three-dimensional imaging of the central airways with multidetector CT. PMID- 12130425 TI - Central venous access: a primer for the diagnostic radiologist. PMID- 12130426 TI - CT screening: why I do it. PMID- 12130427 TI - Role models in the education of radiologists. PMID- 12130428 TI - When does malpractice become manslaughter? PMID- 12130429 TI - Christ healing the sick. PMID- 12130430 TI - Placement of endovascular stent-grafts for emergency treatment of acute disease of the descending thoracic aorta. AB - OBJECTIVE: The aim of this study was to evaluate the feasibility, safety, and effectiveness of endovascular stent-graft placement for the emergency treatment of acute descending thoracic aortic disease. MATERIALS AND METHODS: From January 1996 through November 2001, 18 patients underwent emergency endovascular stent graft placement for various types of acute descending thoracic aortic disease. Five patients had Stanford type B aortic dissection, six had traumatic ruptures of the thoracic aorta, five had ruptured aortic aneurysms, and two had penetrating atherosclerotic aortic ulcers. All patients presented with life threatening symptoms requiring treatment with stent-grafts from the emergency kit. All were at high surgical risk due to serious comorbidities. The efficacy of the procedure was assessed at follow-up studies before discharge and at 3, 6, and 12 months after intervention and yearly thereafter. RESULTS: The primary technical success rate was 78%. Four patients had primary perigraft leaks. The secondary technical success rate was 83%. One patient died 20 hr after intervention from stent-graft-related causes. Follow-up studies revealed stent graft migration in one patient. Progression of disease was observed in one patient treated for dissection and in both patients treated for penetrating ulcers. One patient died 7 months after intervention of unknown reasons; all other patients are alive. The mean follow-up time was 17.4 months (range, 0-38 months). CONCLUSION: Emergency repair of acute descending thoracic aortic disease with stent-graft placement can be successfully accomplished and may be a promising alternative to open-chest surgery, especially in patients at high risk. PMID- 12130431 TI - Pulmonary arteriovenous malformations: effect of embolization on right-to-left shunt, hypoxemia, and exercise tolerance in 66 patients. AB - OBJECTIVE: This study assessed the effect and safety of percutaneous transcatheter coil embolization of pulmonary arteriovenous malformations. MATERIALS AND METHODS: In 58 (88%) of 66 patients, all malformations with feeding vessels greater than or equal to 3 mm in diameter were embolized with steel coils. Arterial oxygen saturation at rest and exercise, intrapulmonary right-to left anatomic shunt fraction ((99m)Tc-macroaggregate injection), maximum exercise capacity (incremental work rate test), and pulmonary function were measured before and after embolization. Complications were analyzed. RESULTS: Three categories of patients were identified. Patients in group 1 (27%) had complete occlusion of all angiographically visible pulmonary arteriovenous malformations; patients in group 2 (61%) had complete occlusion of all malformations with feeding vessels greater than or equal to 3 mm in diameter, but with smaller lesions persisting; and patients in group 3 (12%) had incomplete embolization, with feeding vessels greater than or equal to 3 mm in diameter remaining. The mean right-to-left shunt after embolization was least in group 1 (7%), intermediate in group 2 (10%), and greatest in group 3 (19%). Arterial oxygen saturation and right-to-left shunt fraction returned to normal levels (>96% and <3.5%, respectively) in 33% of patients. A significant improvement occurred after embolization in carbon monoxide diffusing capacity per unit of alveolar volume and in exercise capacity in 16 and 10 patients, respectively. In 93 procedures, 12 complications (13%) occurred. CONCLUSION: Coil embolization of pulmonary arteriovenous malformations is effective in reducing right-to-left anatomic shunt fraction and in improving arterial oxygenation. Coil embolization of pulmonary arteriovenous malformations is well tolerated and has a low complication rate. PMID- 12130432 TI - Analysis of early failure of tunneled hemodialysis catheters. AB - OBJECTIVE: Tunneled hemodialysis catheters are often placed by the interventional radiology service using sonographic guidance and fluoroscopy for safe and optimal placement. The aim of this study was to determine the causes of early failure ( 1 x 10(9)/L) and platelet (> 20 x 10(9)/L) counts of 16 (range, 11-24) and 23 days (range, 12-43), respectively. Complete donor chimerism was detected after a median of 30 days (range, 19-38). Acute GVHD stage II occurred in 4 patients (25%) and grade III GVHD in 2 patients (13%). Chronic GVHD developed in 40% of the patients, but only 1 patient experienced extensive chronic GVHD requiring further immunosuppressive therapy. Two patients died of alveolar hemorrhage and pneumonia, resulting in a day 100 mortality rate of 11%. The rate of complete remission with negative immunofixation increased from 18% after autografting to 73% after allografting. After a median follow-up of 17 months after autologous and 13 months after allogeneic transplantation 13 patients are alive and 12 of them free of relapse or progression. The tandem auto-allotransplant protocol is highly active and provides rapid engraftment with complete donor chimerism and tolerable toxicity. PMID- 12130483 TI - Transplantation of mobilized peripheral blood cells to HLA-identical siblings with standard-risk leukemia. AB - Allogeneic mobilized peripheral blood progenitor cells instead of bone marrow are increasingly used to restore hematopoiesis after myeloablative therapy. Data supporting this important change of clinical practice are scarce. We therefore assigned patients with early leukemias to peripheral blood or bone marrow transplantation; the occurrence of acute and chronic graft versus host disease, survival, transplantation-related mortality, and relapse rates were compared. A total of 350 patients between 18 and 55 years of age with acute leukemias in remission or chronic myelogenous leukemia in first chronic phase were randomized to receive either filgrastim-mobilized peripheral blood progenitor cells or bone marrow cells from HLA-identical sibling donors after standard high-dose chemoradiotherapy. Neutrophil and platelet recovery occurred significantly faster after transplantation of peripheral blood progenitor cells than after bone marrow transplantation. Acute graft versus host disease of grades II-IV was significantly more frequent in recipients of peripheral blood progenitor cells than in recipients of marrow cells (52% vs 39%, odds ratio 1.74, 95% confidence interval 1.12-2.69, P =.013). The cumulative incidence of chronic graft versus host disease was 67% with peripheral blood progenitor cells and 54% with bone marrow cells (hazard ratio 1.67, 95% confidence interval 1.15-2.42, P =.0066). The estimated overall probability of survival at 2 years was 65% with either source of stem cells (hazard ratio 1.15, 95% confidence interval 0.79-1.67, P =.46). Disease-free survival, transplantation-related mortality at day 100, and relapse rates did not significantly differ between treatment arms. Peripheral blood is an equivalent source of hematopoietic stem cells compared with bone marrow if administered to patients with standard-risk leukemias. Long-term observation of patients with different diseases and stages of disease is necessary to ultimately define the role of both sources of stem cells. PMID- 12130484 TI - Phase II trial of subcutaneous anti-CD52 monoclonal antibody alemtuzumab (Campath 1H) as first-line treatment for patients with B-cell chronic lymphocytic leukemia (B-CLL). AB - This phase II study determined the efficacy and safety of alemtuzumab, a humanized anti-CD52 monoclonal antibody, delivered subcutaneously as first-line therapy, over a prolonged treatment period of 18 weeks in 41 patients with symptomatic B-cell chronic lymphocytic leukemia (B-CLL). Injections were administered subcutaneously 3 times per week, from week 2 to 3 onward. An overall response rate (OR) of 87% (95% CI, 76%-98%; complete remission [CR], 19%; partial remission [PR], 68%) was achieved in 38 evaluable patients (81% of intent-to treat population). CLL cells were cleared from blood in 95% patients in a median time of 21 days. CR or nodular PR in the bone marrow was achieved in 66% of the patients and most patients achieved this after 18 weeks of treatment. An 87% OR (29% CR) was achieved in the lymph nodes. The median time to treatment failure has not yet been reached (18+ months; range, 8-44+ months). Transient injection site skin reactions were seen in 90% of patients. Rigor, rash, nausea, dyspnea, and hypotension were rare or absent. Transient grade IV neutropenia developed in 21% of the patients. Infections were rare, but 10% patients developed cytomegalovirus (CMV) reactivation. These patients rapidly responded to intravenous ganciclovir. One patient, allergic to cotrimoxazole prophylaxis, developed Pneumocystis carinii pneumonia. Alemtuzumab is highly effective as first-line treatment in patients with B-CLL. Prolonged treatment is important for maximal bone marrow response. Subcutaneous administration induced very few "first dose" flulike symptoms and may reduce health care costs in comparison with the intravenous infusions. PMID- 12130485 TI - Allogeneic bone marrow transplantation: cure for familial Mediterranean fever. AB - We describe data on a 7-year-old girl with congenital dyserythropoietic anemia (CDA), who also had familial Mediterranean fever (FMF). Repeated transfusions required since the age of 6 months to treat her CDA led to iron overload and a persistently high ferritin level. Her relapsing FMF made effective iron chelation therapy very difficult. Consequently, at the age of 4 years, she underwent allogeneic, sibling bone marrow transplantation (BMT). During conditioning for her BMT, symptoms of FMF, including splenomegaly, arthritis, and recurrent abdominal pain, began to resolve and she was gradually weaned off colchicine. Now, 2 years after the transplantation, she remains free from FMF symptomatology and is off all immunosuppressants. This case demonstrates that symptoms of FMF can be alleviated by the therapy used during allogeneic BMT. In this patient it is likely that the missing factor in FMF is now being provided by granulocytes derived from the stem cells within transplanted bone marrow. PMID- 12130488 TI - Myelodysplastic syndrome is not merely "preleukemia". AB - Myelodysplastic syndrome (MDS) is a disease characterized by ineffective hematopoiesis. There are significant biologic and clinical differences between MDS and acute myeloid leukemia (AML). We studied a cohort of 802 patients, 279 (35%) with newly diagnosed MDS and 523 (65%) with newly diagnosed AML, and compared clinical and biologic characteristics of the 2 groups. Complete clinical and cytogenetic data were available on all patients, and a subgroup of patients was studied for apoptosis, angiogenesis, proliferation, and growth factors. Our results demonstrate that MDS is a discrete entity that is different from AML and is characterized primarily by increased apoptosis in early and mature hematopoietic cells. Using cell sorting and loss of heterozygosity, we demonstrate that the leukemic cells from MDS patients are capable of differentiation into mature myeloid cells and monocytes. We also demonstrate that there is a significant overlap between AML and MDS when MDS is defined on the basis of an arbitrary percentage of blasts of 20% or 30%. These data suggest that despite similarities between AML and MDS in their responses to treatment and outcomes, MDS is biologically and clinically different from AML and should not be considered an early phase of AML. The data indicate that MDS must be better defined on the basis of its biology rather than the percentage of blasts; further, the data suggest that there is a need to develop therapeutic approaches that specifically address the biologic abnormalities of MDS. PMID- 12130487 TI - Risk factors for evolution of acquired aplastic anemia into myelodysplastic syndrome and acute myeloid leukemia after immunosuppressive therapy in children. AB - Long-term survivors of acquired aplastic anemia (AA) have an increased risk of developing myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML) after immunosuppressive therapy (IST). It is uncertain whether the increased survival time simply discloses the natural history of AA as a premalignant disease or whether secondary disease is related to the therapy itself. Between November 1992 and September 1997, 113 AA children with normal cytogenetics at diagnosis were treated with IST using antithymocyte globulin, cyclosporin, and danazol with or without granulocyte colony-stimulating factor (G-CSF). We assessed risk factors for developing MDS/AML by Cox proportional hazards models. Twelve of 113 patients developed MDS between 9 and 81 months following the time of diagnosis, giving a cumulative incidence of 13.7 +/- 3.9%. The following cytogenetic abnormalities were observed at the time of diagnosis of MDS: monosomy 7 (6 patients), monosomy7/trisomy21 (1 patient), trisomy 11 (1 patient), del (11) (9?:14) (1 patient), add (9q) (1 patient), add 7 (q 32) (1 patient), and trisomy 9 (1 patient). The number of days of G-CSF therapy and nonresponse to therapy at 6 months were statistically significant risk factors by multivariate analysis. The present study suggests a close relationship between long-term use of G-CSF and secondary MDS in nonresponders to IST. PMID- 12130486 TI - von Willebrand factor cleaving protease (ADAMTS13) is deficient in recurrent and familial thrombotic thrombocytopenic purpura and hemolytic uremic syndrome. AB - Whether measurement of ADAMTS13 activity may enable physicians to distinguish thrombotic thrombocytopenic purpura (TTP) from hemolytic uremic syndrome (HUS) is still a controversial issue. Our aim was to clarify whether patients with normal or deficient ADAMTS13 activity could be distinguished in terms of disease manifestations and multimeric patterns of plasma von Willebrand factor (VWF). ADAMTS13 activity, VWF antigen, and multimeric pattern were evaluated in patients with recurrent and familial TTP (n = 20) and HUS (n = 29). Results of the collagen-binding assay of ADAMTS13 activity were confirmed in selected samples by testing the capacity of plasma to cleave recombinant VWF A1-A2-A3. Most patients with TTP had complete or partial deficiency of ADAMTS13 activity during the acute phase, and in some the defect persisted at remission. However, complete ADAMTS13 deficiency was also found in 5 of 9 patients with HUS during the acute phase and in 5 patients during remission. HUS patients with ADAMTS13 deficiency could not be distinguished clinically from those with normal ADAMTS13. In a subgroup of patients with TTP or HUS, the ADAMTS13 defect was inherited, as documented by half-normal levels of ADAMTS13 in their asymptomatic parents, consistent with the heterozygous carrier state. In patients with TTP and HUS there was indirect evidence of increased VWF fragmentation, and this occurred also in patients with ADAMTS13 deficiency. In conclusion, deficient ADAMTS13 activity does not distinguish TTP from HUS, at least in the recurrent and familial forms, and it is not the only determinant of VWF abnormalities in these conditions. PMID- 12130489 TI - Outcome of 154 patients with severe aplastic anemia who received transplants from unrelated donors: the Japan Marrow Donor Program. AB - We retrospectively analyzed results for 154 patients with acquired severe aplastic anemia who received bone marrow transplants between 1993 and 2000 from unrelated donors identified through the Japan Marrow Donor Program. Patients were aged between 1 and 46 years (median, 17 years). Seventy-nine donor-patient pairs matched at HLA-A, -B, and -DRB1 loci, as shown by DNA typing. Among the 75 mismatched pairs, DNA typing of 63 pairs showed that 51 were mismatched at 1 HLA locus (18 HLA-A, 11 HLA-B, 22 HLA-DRB1) and 12 were mismatched at 2 or more loci. Seventeen patients (11%) experienced either early or late graft rejection. The incidence of grade III/IV acute graft versus host disease and chronic graft versus host disease was 20% (range, 7%-33%) and 30% (range, 12%-48%), respectively. Currently, 99 patients are alive, having survived for 3 to 82 months (median, 29 months) after their transplantations. The probability of overall survival at 5 years was 56% (95% confidence interval, 34%-78%). Multivariate analysis revealed the following unfavorable factors: transplantation more than 3 years after diagnosis (relative risk [RR], 1.86; P =.02), patients older than 20 years (RR, 2.27; P =.03), preconditioning regimen without antithymocyte globulin (RR 2.28; P =.04), and HLA-A or -B locus mismatching as determined by DNA typing. Matching of HLA class I alleles and improvement of preparative regimens should result in improved outcomes in patients with severe aplastic anemia who receive transplants from unrelated donors. PMID- 12130490 TI - Complete allogeneic hematopoietic chimerism achieved by a combined strategy of in utero hematopoietic stem cell transplantation and postnatal donor lymphocyte infusion. AB - In utero hematopoietic stem cell transplantation (IUHSCTx) can achieve mixed hematopoietic chimerism and donor-specific tolerance without cytoreductive conditioning or immunosuppression. The primary limitation to the clinical application of IUHSCTx has been minimal donor cell engraftment, well below therapeutic levels for most target diseases. Donor lymphocyte infusion (DLI) has been used in postnatal circumstances of mixed chimerism as targeted immunotherapy to achieve a graft-versus-hematopoietic effect and to increase levels of donor cell engraftment. In this report we demonstrate in the murine model that a combined approach of IUHSCTx followed by postnatal DLI can convert low-level, mixed hematopoietic chimerism to complete donor chimerism across full major histocompatibility complex barriers with minimal risk for graft-versus-host disease (GVHD). Time-dated embryonic day 14 (E14) to E15 Balb/c (H-2K(d), CD45.2) fetuses underwent intraperitoneal injection of 5 x 10(6) T-cell-depleted B6 (H 2K(b), CD45.2) bone marrow cells. Chimeric recipients then received transplants at either 4 or 8 weeks of age with 1 of 3 doses (5, 15, or 30 x 10(6) cells) of donor congenic splenocytes (B6-Ly5.2/Cr, H-2K(b), CD45.1). The response to DLI was dose dependent, with conversion to complete donor peripheral blood chimerism in 100% of animals that received high-dose (30 x 10(6) cells) DLI. Only 1 of 56 animals receiving this dose succumbed to GVHD. This study directly supports the potential therapeutic strategy of prenatal tolerance induction to facilitate nontoxic postnatal cellular therapy and organ transplantation, and it has broad implications for the potential treatment of prenatally diagnosed genetic disorders. PMID- 12130491 TI - Lentiviral vectors containing the human immunodeficiency virus type-1 central polypurine tract can efficiently transduce nondividing hepatocytes and antigen presenting cells in vivo. AB - High-titer self-inactivating human immunodeficiency virus type-1 (HIV-1)-based vectors expressing the green fluorescent protein reporter gene that contained the central polypurine and termination tract and the woodchuck hepatitis virus posttranscriptional regulatory element were constructed. Transduction efficiency and biodistribution were determined, following systemic administration of these improved lentiviral vectors. In adult severe combined immunodeficiency (SCID) mice, efficient stable gene transfer was achieved in the liver (8.0% +/- 6.0%) and spleen (24% +/- 3%). Most transduced hepatocytes and nonhepatocytes were nondividing, thereby obviating the need to induce liver cell proliferation. In vivo gene transfer with this improved lentiviral vector was relatively safe since liver enzyme concentration in the plasma was only moderately and transiently elevated. In addition, nondividing major histocompatibility complex class II positive splenic antigen-presenting cells (APCs) were efficiently transduced in SCID and normal mice. Furthermore, B cells were efficiently transduced, whereas T cells were refractory to lentiviral transduction in vivo. However, in neonatal recipients, lentiviral transduction was more widespread and included not only hepatocytes and splenic APCs but also cardiomyocytes. The present study suggests potential uses of improved lentiviral vectors for gene therapy of genetic blood disorders resulting from serum protein deficiencies, such as hemophilia, and hepatic disease. However, the use of liver-specific promoters may be warranted to circumvent inadvertent transgene expression in APCs. In addition, these improved lentiviral vectors could potentially be useful for genetic vaccination and treatment of perinatal cardiac disorders. PMID- 12130492 TI - Lentiviral vectors pseudotyped with a modified RD114 envelope glycoprotein show increased stability in sera and augmented transduction of primary lymphocytes and CD34+ cells derived from human and nonhuman primates. AB - Generating lentiviral vectors pseudotyped with different viral glycoproteins (GPs) may modulate the physicochemical properties of the vectors, their interaction with the host immune system, and their host range. We have investigated the capacity of a panel of GPs of both retroviral (amphotropic murine leukemia virus [MLV-A]; gibbon ape leukemia virus [GALV]; RD114, feline endogenous virus) and nonretroviral (fowl plague virus [FPV]; Ebola virus [EboV]; vesicular stomatitis virus [VSV]; lymphocytic choriomeningitis virus [LCMV]) origins to pseudotype lentiviral vectors derived from simian immunodeficiency virus (SIVmac251). SIV vectors were efficiently pseudotyped with the FPV hemagglutinin, VSV-G, LCMV, and MLV-A GPs. In contrast, the GALV and RD114 GPs conferred much lower infectivity to the vectors. Capitalizing on the conservation of some structural features in the transmembrane domains and cytoplasmic tails of the incorporation-competent MLV-A GP and in RD114 and GALV GPs, we generated chimeric GPs encoding the extracellular and transmembrane domains of GALV or RD114 GPs fused to the cytoplasmic tail (designated TR) of MLV-A GP. Importantly, SIV-derived vectors pseudotyped with these GALV/TR and RD114/TR GP chimeras had significantly higher titers than vectors coated with the parental GPs. Additionally, RD114/TR-pseudotyped vectors were efficiently concentrated and were resistant to inactivation induced by the complement of both human and macaque sera, indicating that modified RD114 GP-pseudotyped lentiviral vectors may be of particular interest for in vivo gene transfer applications. Furthermore, as compared to vectors pseudotyped with other retroviral GPs or with VSV-G, RD114/TR pseudotyped vectors showed augmented transduction of human and macaque primary blood lymphocytes and CD34+ cells. PMID- 12130493 TI - Development of a murine hematopoietic progenitor complementary DNA microarray using a subtracted complementary DNA library. AB - With the goal of creating a resource for in-depth study of myelopoiesis, we have executed a 2-pronged strategy to obtain a complementary DNA (cDNA) clone set enriched in hematopoietic genes. One aspect is a library subtraction to enrich for underrepresented transcripts present at early stages of hematopoiesis. For this, a hematopoietic cDNA library from primary murine bone marrow cells enriched for primitive progenitors was used as tester. The subtraction used 10 000 known genes and expressed sequence tags (ESTs) as driver. The 2304 randomly picked clones from the subtracted cDNA libraries represent 1255 distinct genes, of which 622 (50%) are named genes, 386 (30%) match uncharacterized ESTs, and 247 (20%) are novel. The second aspect of our strategy was to complement this subtracted library with genes known to be involved in myeloid cell differentiation and function. The resulting cDNAs were arrayed on polylysine-coated glass slides. The microarrays were used to analyze gene expression in primary and cultured murine bone marrow-derived progenitors. We found expression of various types of genes, including regulatory cytokines and their receptors, signal transduction genes, and transcription factors. To assess gene expression during myeloid differentiation, we examined patterns of change during induced differentiation of EML cells. Several hundred of the genes underwent fluctuations in expression level during myeloid cell differentiation. The complete database, accessible on the World Wide Web at http://yale130132115135.med.yale.edu/, allows for retrieval of information regarding these genes. Our microarray allows for genomewide expression analysis of myeloid stem cells, which will help in defining the regulatory mechanisms of stem cell differentiation. PMID- 12130494 TI - Down-regulation of DNA repair in human CD34(+) progenitor cells corresponds to increased drug sensitivity and apoptotic response. AB - Although DNA repair processes have been shown to considerably modulate the cytotoxic effects of alkylating agents, little information is available on the role of these mechanisms in chemotherapy-induced myelosuppression. Therefore, we have analyzed in detail the DNA repair capacity of primary human hematopoietic cells from cord blood (CB) or bone marrow (BM) by 2 functional assays, the immunocytologic assay (ICA) and single-cell gel electrophoresis (comet assay). Besides substantial interindividual differences, we consistently observed significantly lower repair capacity of CD34(+) cells in comparison to CD34(-), CD19(+), or CD33(+) cells of the same donor. After exposure to the alkylating agent ethylnitrosourea (EtNU), the comet assay displayed on average twice as many DNA single-strand breaks (SSBs) in CD34(+) cells and a tripled half-life of these lesions in comparison to corresponding CD34(-) cells. Similarly, reduced SSB repair activity in CD34(+) cells was detected following melphalan or cisplatin application. When specific antibodies were used to monitor DNA reaction products of these drugs, adduct levels were significantly higher and lesions persisted longer in the CD34(+) fraction. To assess the contribution of individual pathways to overall DNA repair, modulators blocking defined steps in repair processes were coapplied with alkylating drugs. Similar "modulation pattern" in corresponding CD34(+) and CD34(-) cell fractions indicated a generalized reduction in DNA repair capacity of CD34(+) cells, rather than deficiencies in a specific pathway. Because CD34(+) cells also displayed higher frequencies of apoptosis in response to melphalan or cisplatin, these findings may help to explain the myelosuppression after exposure to alkylating agents. PMID- 12130495 TI - Evaluation of role of G-CSF in the production, survival, and release of neutrophils from bone marrow into circulation. AB - In steady-state hematopoiesis, G-CSF (granulocyte-colony stimulating factor) regulates the level of neutrophils in the bone marrow and blood. In this study, we have exploited the availability of G-CSF-deficient mice to evaluate the role of G-CSF in steady-state granulopoiesis and the release of granulocytes from marrow into circulation. The thymidine analogue bromodeoxyuridine (BrdU) was used to label dividing bone marrow cells, allowing us to follow the release of granulocytes into circulation. Interestingly, the labeling index and the amount of BrdU incorporated by blast cells in bone marrow was greater in G-CSF-deficient mice than in wild-type mice. In blood, 2 different populations of BrdU-positive granulocytes, BrdU(bright) and BrdU(dim), could be detected. The kinetics of release of the BrdU(bright) granulocytes from bone marrow into blood was similar in wild-type and G-CSF-deficient mice; however, BrdU(dim) granulocytes peaked earlier in G-CSF-deficient mice. Our findings suggest that the mean transit time of granulocytes through the postmitotic pool is similar in G-CSF-deficient and control mice, although the transit time through the mitotic pool is reduced in G CSF-deficient mice. Moreover, the reduced numbers of granulocytes that characterize G-CSF-deficient mice is primarily due to increased apoptosis in cells within the granulocytic lineage. Collectively, our data suggest that at steady state, G-CSF is critical for the survival of granulocytic cells; however, it is dispensable for trafficking of granulocytes from bone marrow into circulation. PMID- 12130496 TI - Deregulated expression of HOXB4 enhances the primitive growth activity of human hematopoietic cells. AB - Identification of the molecular mechanisms that can promote human hematopoietic stem cell amplification is a major goal in experimental and clinical hematology. Recent data indicate that a variety of regulatory molecules active in early development may also play a role in the maintenance of hematopoietic stem cells with repopulating activity. One important class of early developmental genes determining hematopoietic development are homeobox transcription factors. Here, we report that retrovirally mediated expression of the homeobox gene HOXB4 rapidly triggers an increase in the number of human hematopoietic cord blood cells with stem cell and progenitor cell properties detected both by in vitro and in vivo assays. This growth enhancement extended across primitive myeloid erythroid and B-lymphoid progenitors but did not lead to alterations in the balance of lymphomyeloid reconstitution in vivo, suggesting that HOXB4 does not affect control of end-cell output. These findings reveal HOXB4 as a novel, positive regulator of the primitive growth activity of human hematopoietic progenitor cells and underline the relevance of early developmental factors for stem cell fate decisions. PMID- 12130497 TI - Functional analysis of human hematopoietic repopulating cells mobilized with granulocyte colony-stimulating factor alone versus granulocyte colony-stimulating factor in combination with stem cell factor. AB - Using in vitro progenitor assays, serum-free in vitro cultures, and the nonobese diabetic/severe combined immune-deficient (NOD/SCID) ecotropic murine virus knockout xenotransplantation model to detect human SCID repopulating cells (SRCs) with multilineage reconstituting function, we have characterized and compared purified subpopulations harvested from the peripheral blood (PB) of patients receiving granulocyte colony-stimulating factor (G-CSF) alone or in combination with stem cell factor (SCF). Mobilized G-CSF plus SCF PB showed a 2-fold increase in total mononuclear cell content and a 5-fold increase in CD34-expressing cells depleted for lineage-marker expression (CD34(+)Lin(-)) as compared with patients treated with G-CSF alone. Functionally, G-CSF plus SCF-mobilized CD34(+)CD38( )Lin(-) cells contained a 2-fold enhancement in progenitor frequency as compared with G-CSF-mobilized subsets. Despite enhanced cellularity and progenitor capacity, G-CSF plus SCF mobilization did not increase the frequency of SRCs as determined by limiting dilution analysis by means of unfractionated PB cells. Purification of SRCs from these sources demonstrated that as few as 1000 CD34(+)CD38(-)Lin(-) cells from G-CSF-mobilized PB contained SRC capacity while G CSF plus SCF-mobilized CD34(+)CD38(-)Lin(-) cells failed to repopulate at doses up to 500 000 cells. In addition, primitive CD34(-)CD38(-)AC133(+)Lin(-) cells derived from G-CSF plus SCF-mobilized PB were capable of differentiation into CD34-expressing cells, while the identical subfractions from G-CSF PB were unable to produce CD34(+) cells in serum-free cultures. Our study defines qualitative and quantitative distinctions among subsets of primitive cells mobilized by means of G-CSF plus SCF versus G-CSF alone, and therefore has implications for the utility of purified repopulating cells from these sources. PMID- 12130498 TI - Pro-oxidant and cytotoxic effects of circulating heme. AB - Numerous pathologies may involve toxic side effects of free heme and heme-derived iron. Deficiency of the heme-catabolizing enzyme, heme oxygenase-1 (HO-1), in both a human patient and transgenic knockout mice leads to an abundance of circulating heme and damage to vascular endothelium. Although heme can be directly cytotoxic, the present investigations examine the possibility that hemoglobin-derived heme and iron might be indirectly toxic through the generation of oxidized forms of low-density lipoprotein (LDL). In support, hemoglobin in plasma, when oxidized to methemoglobin by oxidants such as leukocyte-derived reactive oxygen, causes oxidative modification of LDL. Heme, released from methemoglobin, catalyzes the oxidation of LDL, which in turn induces endothelial cytolysis primarily caused by lipid hydroperoxides. Exposure of endothelium to sublethal concentrations of this oxidized LDL leads to induction of both HO-1 and ferritin. Similar endothelial cytotoxicity was caused by LDL isolated from plasma of an HO-1-deficient child. Spectral analysis of the child's plasma revealed a substantial oxidation of plasma hemoglobin to methemoglobin. Iron accumulated in the HO-1-deficient child's LDL and several independent assays revealed oxidative modification of the LDL. We conclude that hemoglobin, when oxidized in plasma, can be indirectly cytotoxic through the generation of oxidized LDL by released heme and that, in response, the intracellular defense-HO-1 and ferritin-is induced. These results may be relevant to a variety of disorders-such as renal failure associated with intravascular hemolysis, hemorrhagic injury to the central nervous system, and, perhaps, atherogenesis-in which hemoglobin-derived heme may promote the formation of fatty acid hydroperoxides. PMID- 12130499 TI - Characterization of Kaposi sarcoma-associated herpesvirus/human herpesvirus-8 infection of human vascular endothelial cells: early events. AB - Kaposi sarcoma-associated herpesvirus (KSHV)/human herpesvirus-8 (HHV-8) is causally associated with Kaposi sarcoma (KS). The absence of a cell culture system that effectively reproduces the composite mechanisms governing initiation and maintenance of HHV-8 infection (lytic and latent) in KS endothelial cells, however, has left important questions unanswered. Here, we report a culture system in which the earliest events that accompany HHV-8 infection could be surveyed in primary endothelial cells. Binding of HHV-8 to microvascular dermal endothelial cells (MVDECs) was directly compared with other primary target cells implicated in HHV-8-associated diseases. Virus attachment, fusion, internalization and transport within MVDECs was monitored by electron microscopy. Studies of genome configuration revealed that rapid circularization of the viral DNA occurred on entry, though by 72 hours after infection linear DNAs accumulated and early as well as late lytic RNAs (T1.1, K8.1) could be detected. The latency transcripts (LT1/LT2) were first detected on day 8, demonstrating that both lytic and latent infection were initiated. Although most lytic transcripts accrued until passage, open-reading frame-74 RNAs fluctuated with a fixed periodicity, suggesting that early replication after infection of MVDECs was synchronous. PMID- 12130500 TI - Stomatin is a major lipid-raft component of platelet alpha granules. AB - Lipid rafts are detergent-resistant, cholesterol- and sphingolipid-rich membrane domains that are involved in important cellular processes such as signal transduction and intracellular trafficking. Stomatin, a major lipid-raft component of erythrocytes and epithelial cells, is also an abundant platelet protein. Microscopical methods and subcellular fractionation showed that stomatin is located mainly at the alpha-granular membrane. The lipid-raft marker proteins flotillin-1 and flotillin-2 were also present in platelets but excluded from alpha granules. Stomatin and the flotillins were associated with Triton X-100 insoluble lipid rafts. Whereas stomatin was partly soluble in Triton X-100, it was insoluble in the detergents Lubrol and 3-[(3-cholamidopropyl)dimethylamonio] 1-propyl sulfonate (CHAPS). Flotation experiments after CHAPS lysis of platelets revealed a distinct set of lipid-raft-associated proteins, which were identified by matrix-assisted laser desorption/ionization mass spectrometry as stomatin, flotillin-1, flotillin-2, CD36, CD9, integrin alpha(IIb)beta(3), and the glucose transporter GLUT-3. Stomatin, the flotillins, and CD36 were exclusively present in this lipid-raft fraction. Activation of platelets by calcium ionophore A23187 or thrombin led to translocation of stomatin to the plasma membrane, cleavage by calpain, and specific sorting into released microvesicles. In conclusion, this study demonstrated the existence of alpha-granular lipid rafts and suggests an important role for stomatin in the organization and function of alpha granules. PMID- 12130501 TI - A monoclonal antibody to vascular endothelial-cadherin inhibits tumor angiogenesis without side effects on endothelial permeability. AB - Vascular endothelial cadherin (VE-cadherin) is an endothelial-specific, trans membrane protein that promotes homophilic cell adhesion. Inhibition of VE cadherin by the blocking monoclonal antibody (mAb) BV13 inhibited angiogenesis and tumor growth in vivo. However, this effect was accompanied by a marked increase in lung and heart permeability. In the present paper, we characterize a different VE-cadherin mAb (BV14) that is able to inhibit angiogenesis without affecting vascular permeability. In vitro studies show that BV14, in contrast to BV13, did not increase paracellular permeability of endothelial monolayers and did not disrupt VE-cadherin clusters at junctions. However, both antibodies could inhibit formation of vascularlike structures in collagen gels and increase migration of endothelial cells into wounded areas. In vivo, BV14 and BV13 were equally active in inhibiting angiogenesis in the mouse cornea and in reducing the growth of hemangioma and C6 glioma. In contrast to BV13, BV14 did not change vascular permeability in all the organs tested and at any dose used. BV14 and BV13 bind to VE-cadherin extracellular repeats EC4 and EC1, respectively. We propose that, in resting vessels, where junctions are stable and well-structured, antibody binding to EC1 but not EC4 disrupts their organization and increases permeability. In contrast, in growing vessels, where endothelial cells are migrating and junctions are weaker, antibody binding to EC4 may be sufficient to disrupt cell-to-cell adhesion and inhibit assembly of new vascular structures. PMID- 12130503 TI - NAD(P)H oxidase-dependent platelet superoxide anion release increases platelet recruitment. AB - Platelets, although not phagocytotic, have been suggested to release O. Since O producing reduced nicotinamide adenine dinucleotide (phosphate) (NAD(P)H) oxidases can be specifically activated by certain agonists and are found in several nonphagocytotic tissues, we investigated whether such an enzyme is the source of platelet-derived O. We further studied which agonists cause platelet O release and whether platelet-derived O influences thrombus formation in vitro. Collagen, but not adenosine 5'-diphosphate (ADP) or thrombin, increased O formation in washed human platelets. This was a reduced nicotinamide adenine dinucleotide (NADH)-dependent process, as shown in platelet lysates. Consistent with a role of a platelet, NAD(P)H oxidase expression of its subunits p47(phox) and p67(phox) and inhibition of platelet O formation by diphenylene iodoniumchloride (DPI) and by the specific peptide-antagonist gp91ds-tat were observed. Whereas platelet-derived O did not influence initial aggregation, platelet recruitment to a preformed thrombus following collagen stimulation was significantly attenuated by superoxide dismutase (SOD) or DPI. It was also inhibited when ADP released during aggregation was cleaved by the ectonucleotidase apyrase. ADP in supernatants of collagen-activated platelets was decreased in the presence of SOD, resulting in lower ADP concentrations available for recruitment of further platelets. Exogenous O increased ADP- concentrations in supernatants of collagen-stimulated platelets and induced irreversible aggregation when platelets were stimulated with otherwise subthreshold concentrations of ADP. These results strongly suggest that collagen activation induces NAD(P)H oxidase-dependent O release in platelets, which in turn enhances availability of released ADP, resulting in increased platelet recruitment. PMID- 12130502 TI - c-myc proto-oncogene expression in hemophilic synovitis: in vitro studies of the effects of iron and ceramide. AB - Hemophilia is a rare congenital bleeding disorder that is due to the deficiency of blood coagulation factor VIII or IX. Recurrent musculoskeletal bleeding is common and bleeding into joints results in a chronic inflammatory condition termed hemophilic synovitis. This destructive process is characterized by hemosiderin deposition in the superficial and deeper layers of the synovial membrane as well as a proliferation of synovial fibroblasts and vascular cells. The hyperplastic synovium and neovascular changes are reminiscent of the histopathologic appearance observed in malignant tissues. Indeed, the benign hyperplastic synovium in patients with hemophilia displays similar invasive and destructive behaviors suggesting the possibility of analogous disturbances in growth control and locally invasive mechanisms. Iron plays a role in malignant cell growth, local invasion, and tumor progression, possibly due to changes in the expression of the proto-oncogene, c-myc. We hypothesized that iron plays a similar role in hemophilic synovitis. To explore this hypothesis, we investigated the in vitro effects of iron on the proliferation of a primary, human synovial fibroblast cell (HSFC) line and the involvement of c-myc in this process. We also examined the role of ceramide, a sphingolipid capable of inducing apoptosis in this model system. HSFC proliferation was increased in a dose-dependent fashion and c-myc expression was enhanced by ferric citrate compared to sodium citrate control. Ceramide prevented both the iron-induced increases in HSFC proliferation and c-myc expression. These results indicate that iron probably plays a role in the proliferative changes observed in hemophilic joint disease and that aberrant expression of c-myc may underlie the iron effects. Furthermore, these results suggest that there may be a therapeutic role for ceramide in reversing these changes. PMID- 12130504 TI - Nucleotides induce chemotaxis and actin polymerization in immature but not mature human dendritic cells via activation of pertussis toxin-sensitive P2y receptors. AB - Dendritic cells (DCs) are considered the principal initiators of immune response because of their ability to migrate into peripheral tissues and lymphoid organs, process antigens, and activate naive T cells. There is evidence that extracellular nucleotides regulate certain functions of DCs via G-protein-coupled P2Y receptors (P2YR) and ion-channel-gated P2X receptors (P2XR). Here we investigated the chemotactic activity and analyzed the migration-associated intracellular signaling events such as actin reorganization and Ca(++) transients induced by common P2R agonists such as adenosine 5'-triphosphate (ATP) and 2 methylthioadenosine triphosphate, the P2YR agonists UTP and adenosine 5' diphosphate (ADP), or the P2XR agonists alphabeta-methylenadenosine-5' triphosphate and 2',3'-(4-benzoyl)benzoyl-ATP. The common P2R agonists and the selective P2YR agonists turned out to be potent chemotactic stimuli for immature DCs, but not for mature DCs. In contrast, P2XR agonists had only marginal chemotactic activity in both DC types. Chemotaxis was paralleled by a rise in the intracellular Ca(++) concentration and by actin polymerization. Studies with pertussis toxin implicated that intracellular signaling events such as actin polymerization, mobilization of intracellular Ca(++), and migration induced by nucleotides was mediated via G(i/o) protein-coupled P2YR. Moreover, functional studies revealed selective down-regulation of this G(i/o) protein-coupled chemotactic P2YR responsiveness during maturation, although immature and mature DCs expressed similar amounts of mRNA for the P2R subtypes (P2Y(2)R, P2Y(4)R, P2Y(5)R, P2Y(7)R, P2Y(11)R and P2X(1)R, P2X(4)R, P2X(7)R), and no major differences in respect to the mRNA expression of these receptors could be observed by semiquantitative reverse transcription and polymerase chain reaction (RT-PCR). In summary, our data describe a differential chemotactic response of immature and mature DCs to nucleotides, and lend further support to the hypothesis that P2R are a novel class of immunomodulatory plasma membrane receptors suitable for pharmacological intervention. PMID- 12130505 TI - Epstein-Barr virus-specific CD8(+) T cells that re-express CD45RA are apoptosis resistant memory cells that retain replicative potential. AB - During acute infection, latent and lytic Epstein-Barr virus (EBV) epitope specific CD8(+) T cells have a CD45RO(+) CD45RA(-) phenotype. However, after resolution of the infection, a large proportion of these cells, particularly those specific for lytic viral epitopes, re-express the CD45RA molecule. The role of CD8(+) CD45RA(+) T cells in ongoing immunity to EBV and other viruses is unknown. We now demonstrate that, relative to their CD45RO(+) counterparts, the EBV-specific CD8(+) T cells that revert to CD45RA expression after acute infectious mononucleosis are not in cell cycle, have longer telomeres, and are more resistant to apoptosis partly because of increased Bcl-2 expression. However, the EBV-specific CD8(+) CD45RA(+) T cells have shorter telomeres than the total CD8(+) CD45RA(+) T-cell pool and predominantly express low levels of the CCR7 chemokine receptor, indicating that they are not naive cells. In addition, EBV-specific CD8(+) CD45RA(+) T cells can be induced to proliferate and exhibit potent cytotoxic activity against target cells loaded with specific peptide. Our results strongly suggest, therefore, that EBV-specific CD8(+) CD45RA(+) T cells represent a stabilized virus-specific memory pool and not terminally differentiated effector cells. The identification of mechanisms that enable stable virus-specific CD8(+) T cells to persist after acute infection may lead to the enhancement of antiviral immunity in immunocompromised and elderly persons. PMID- 12130506 TI - Immunologic mechanisms of extracorporeal photochemotherapy in chronic graft versus-host disease. AB - Extracorporeal photochemotherapy (ECP) has been shown to be an effective therapy for patients with acute and chronic graft-versus-host disease (GVHD) following allogeneic bone marrow transplantation, but its biologic mechanism is not understood. We reported that clinical response to ECP was associated not only with normalization of skewed CD4/CD8 ratios but also with an increase in CD3( )/CD56(+) natural killer cells and a decrease in the number of CD80(+) and CD123(+) circulating dendritic cells (DCs). To further elucidate the effects of ECP on activated lymphocyte subpopulations and the interaction between effector lymphocytes and antigen-presenting DCs, we isolated and characterized DC populations from patients with chronic GVHD undergoing ECP therapy. Antigen presenting activity of DCs was measured as proliferation of antigen-stimulated autologous and allogeneic T cells by mixed-lymphocyte reaction (MLR). In MLR assays the proliferation of T cells was decreased in all 10 patients by a mean of 84% (range, 75%-95%; P < or =.002) after a 2-day cycle of ECP and longitudinally over the 12-month course of therapy. Immunophenotypic analysis of DC populations revealed a preponderance of DC1 monocytic dendritic cells in all patients before the initiation of ECP. Nine of 10 patients demonstrated a shift from DC1 to DC2 and as a concordant shift from a predominantly Th1 (interleukin-2 [IL-2], interferon-gamma) to Th2 (IL-4, IL-10) cytokine profile after ECP, and 8 of 10 had a clinical response to ECP. Our results suggest that ECP alters alloreactivity by affecting allo-targeted effector T cells and antigen-presenting DCs. PMID- 12130507 TI - Analysis of V(lambda)-J(lambda) expression in plasma cells from primary (AL) amyloidosis and normal bone marrow identifies 3r (lambdaIII) as a new amyloid associated germline gene segment. AB - Primary (AL) amyloidosis is a plasma cell dyscrasia characterized by extracellular deposition of monoclonal light-chain variable region (V) fragments in the form of amyloid fibrils. Light-chain amyloid is rare, and it is not fully understood why it occurs in only a fraction of patients with a circulating monoclonal component and why it typically associates with lambda isotype and lambdaVI family light-chain proteins. To provide insights into these issues, we obtained complete nucleotide sequences of monoclonal V(lambda) regions from 55 consecutive unselected cases of primary amyloidosis and the results were compared with the light-chain expression profile of polyclonal marrow plasma cells from 3 healthy donors (a total of 264 sequences). We demonstrated that: (1) the lambdaIII family is the most frequently used both in amyloidosis (47%) and in polyclonality (43%); (2) both conditions are characterized by gene restriction; (3) a very skewed repertoire is a feature of amyloidosis, because just 2 germline genes belonging to the lambdaIII and lambdaVI families, namely 3r (22% of cases, lambdaIII) and 6a (20%, lambdaVI), contributed equally to encode 42% of amyloid V(lambda) regions; (4) these same 2 gene segments have a strong association with amyloidosis if their prevalences are compared with those in polyclonal conditions (3r, 8.3%, P =.024; 6a, 2.3%, P =.0008, chi2 test); (5) the J(lambda)2/3 segment, encoding the fourth framework region, appears to be slightly overrepresented in AL (83% versus 67%, P =.03), and this might be related to preferential J(lambda)2/3 rearrangement in amyloid (11 of 12 cases) versus polyclonal 3r light chains (13 of 22 cases). These findings demonstrate that V(lambda)-J(lambda) expression is more restricted in plasma cells from amyloidosis than from polyclonal bone marrow and identify 3r as a new disease-associated gene segment. Overusage of just 2 gene segments, 3r and 6a, can thus account for the lambda light-chain overrepresentation typical of this disorder. PMID- 12130508 TI - Hematopoietic progenitor kinase 1 supports apoptosis of T lymphocytes. AB - Hematopoietic progenitor kinase 1 (HPK1) is a member of germinal center kinases that is predominantly expressed in hematopoietic cells and transiently activated by T-cell receptor (TCR) triggering. We show here that HPK1 supports apoptosis of T cells. When HPK1 was overexpressed in murine CD4(+) T cells, a substantial increase was observed in spontaneous and TCR/CD3-mediated apoptosis as well as in Fas ligand (FasL) expression. In H2O2-treated EL-4 thymoma cells, which show an increase in reactive oxygen species (ROS) and apoptosis, overexpression of HPK1 enhanced ROS-mediated apoptosis, whereas expression of HPK1 antisense (AS) RNA impaired apoptosis. HPK1 expression also led to a sustained increase in c-Jun N terminal kinase (JNK) activity, suggesting that JNK activation contributes to the HPK1-mediated apoptosis in H2O2-treated EL-4 cells. Under the same conditions, a rapid cleavage of HPK1 was observed, and overexpression of N- and C-terminal cleavage products in CD4(+) T cells resulted in, similar to full-length HPK1, an increase in apoptosis. In agreement with published data, we show that the C terminal portion of HPK1 suppresses IkappaBalpha degradation, thereby inhibiting nuclear factor (NF)-kappaB activation. These findings suggest that by inhibiting the antiapoptotic action of NF-kappaB and inducing the proapoptotic activity of JNK, OHPK1 supports apoptosis in T cells. PMID- 12130509 TI - Sperm protein 17 (Sp17) is a suitable target for immunotherapy of multiple myeloma. AB - Sperm protein 17 (Sp17) is a protein recently identified as a novel cancer-testis (CT) antigen in multiple myeloma (MM). Because this tumor antigen demonstrates a very restricted normal tissue expression, Sp17 may be an excellent target for tumor vaccine of MM. In this study, we determined the ability to generate Sp17 specific HLA class I-restricted cytotoxic T lymphocytes (CTLs) from the peripheral blood of 4 patients with MM, 3 consecutive Sp17(+) patients, and 1 Sp17(-) patient. Dendritic cells were generated from monocytes of 4 patients with MM and used to present a recombinant Sp17 protein to autologous T cells. Following 4 rounds of antigen stimulation, the CTLs were tested for their ability to kill autologous targets in an Sp17-dependent and HLA-class I- restricted manner in standard cytotoxicity assays. Despite previous chemotherapy and the immunosuppression so often associated with MM, CTL generation was successful in all 4 patients, irrespective of the Sp17 status of their tumors. Most importantly, the CTLs were able to lyse autologous tumor cells that expressed Sp17. Tumor cell lysis in all cases appeared to be mainly mediated by perforin and could be blocked by concanamycin A. We conclude that Sp17 is a suitable target for immunotherapy of MM. Our findings provide the basis for a clinical study aimed at inducing a cellular immune response directed at Sp17(+) MM. PMID- 12130510 TI - Functional involvement of Akt signaling downstream of Jak1 in v-Abl-induced activation of hematopoietic cells. AB - Activation of intracellular signaling pathways is important for cellular transformation and tumorigenesis. The nonreceptor tyrosine kinases Jak1 and Jak3, which bind to the v-Abl oncoprotein, are constitutively activated in cells transformed with the Abelson murine leukemia virus. A mutant of p160 v-Abl lacking the Jak1-binding region (v-Abl Delta858-1080) has a significant defect in Jak/STAT (signal transducers and activators of transcription) activation, cytokine-independent cell growth/survival, and tumorigenesis. To identify the pathways downstream of Jak kinases in v-Abl-mediated signaling, we examined the activation of several signaling molecules by p160 v-Abl or the v-Abl Delta858 1080 mutant. We demonstrate that, in addition to the decreased Ras activation, signaling through phosphatidylinositol-3 kinase and Akt are impaired in cells expressing mutant v-Abl. The proliferative defect of v-Abl Delta858-1080 was rescued by activated v-Akt and was also moderately rescued by activated v-H-Ras. However, constitutive active phosphatidylinositol-3 kinase (p110CAAX) did not complement this effect. Cells expressing v-Abl Delta858-1080 demonstrated reduced tumor formation in nude mice. In contrast, cells coexpressing v-Akt with v-Abl Delta858-1080 demonstrated reduced latency and increased frequency of tumor formation in nude nice compared with cells expressing v-Abl Delta858-1080 alone, whereas v-H-Ras or p110CAAX had minimum effects on tumor formation. These results suggest that Jak1-dependent Akt activation is important in v-Abl-mediated transformation. PMID- 12130511 TI - MDR1 protein expression is an independent predictor of complete remission in newly diagnosed adult acute lymphoblastic leukemia. AB - Little is known about the prognostic role of multidrug resistance (MDR) in adults with newly diagnosed acute lymphoblastic leukemia (ALL). In the context of the GIMEMA ALL0496 protocol, we evaluated the impact of MDR1 (protein expression and function) on the achievement of complete remission (CR) and clinical outcome. Flow cytometric analysis of MDR1 expression (D) and function (rhodamine-123 efflux) was obtained in 203 and 158 patients, respectively. MDR1 expression was detected in 44 (21.7%) of 203 patients, and function was found in 23 (14.6%) of 158 (14.6%) patients. Expression of the multidrug resistance-associated protein 1 (MRP1) and lung-resistance protein (LRP) evaluated in 43 samples was found in 13 and 26 patients, respectively. Among the 200 patients evaluable for the clinical correlation study, 125 (79.6%) of 157 without MDR1 expression achieved CR compared with 23 (53.5%) of 43 with MDR1 expression (P =.001). At univariate analysis, MDR1 expression was significantly associated with CR when considered as a dichotomized (P =.001) or continuous (P =.01) variable. At multivariate analysis, dichotomized evaluation of MDR1 expression independently predicted CR (P =.004) with age (P =.03) and CD34 (P =.03); as a continuous variable, MDR1 expression (P =.03) was the only significant factor other than CD34 (P =.01). MDR1 function failed to predict achievement of CR or of MRP1 and LRP expression. MDR1 expression did not correlate with CR duration, nor did it predict for survival duration. These results demonstrate that MDR1 expression in de novo adult ALL is an independent predictor of CR achievement. PMID- 12130512 TI - Regulation of leukemic cell adhesion, proliferation, and survival by beta catenin. AB - In epithelial cells beta-catenin plays a critical role as a component of the cell cell adhesion apparatus and as a coactivator of the TCF/LEF (T-cell transcription factor/lymphoid enhancer binding factor) family of transcription factors. Deregulation of beta-catenin has been implicated in the malignant transformation of cells of epithelial origin. However, a function for beta-catenin in hematologic malignancies has not been reported. beta-Catenin is not detectable in normal peripheral blood T cells but is expressed in T-acute lymphoblastic leukemia cells and other tumor lines of hematopoietic origin and in primary lymphoid and myeloid leukemia cells. beta-Catenin function was examined in Jurkat T-acute lymphoblastic leukemia cells. Overexpression of dominant-negative beta catenin or dominant-negative TCF reduced beta-catenin nuclear signaling and inhibited Jurkat proliferation and clonogenicity. Similarly, these constructs inhibited proliferation of K562 and HUT-102 cells. Reduction of beta-catenin expression with beta-catenin antisense down-regulated adhesion of Jurkat cells in response to phytohemagglutinin. Incubation of Jurkat cells with anti-Fas induced caspase-dependent limited proteolysis of beta-catenin N- and C-terminal regions and rapid redistribution of beta-catenin to the detergent-insoluble cytoskeleton, concomitant with a marked decline in nuclear beta-catenin signaling. Fas-mediated apoptosis was potentiated by inhibition of beta-catenin nuclear signaling. The data suggest that beta-catenin can play a significant role in promoting leukemic cell proliferation, adhesion, and survival. PMID- 12130513 TI - HOX11L2 expression defines a clinical subtype of pediatric T-ALL associated with poor prognosis. AB - The most frequent oncogenic activation events characterized in childhood T acute lymphoblastic leukemia (T-ALL) result in the transcriptional activation of genes coding for transcription factors. The main genes are TAL1/SCL, a member of the basic region helix-loop-helix gene family, and HOX11L2, a member of the homeobox containing protein family. To gain insight into the pathogenesis of this type of hematologic malignancy, we analyzed 28 T-ALL samples. SIL-TAL1/SCL fusion was detected in 6 patients; expression of HOX11L2 was observed in 6 patients and of HOX11 in 3 patients. With one exception, these activations did not occur simultaneously in the same patients, and they allowed the subclassification of 50% of the patients. SIL-TAL1 fusion was detected in association with HOX11 expression in one patient and with a t(8;14) (q24;q11) in another. High expression of LYL1, LMO2, or TAL1 was observed mainly in samples negative for HOX11L2 expression. HOX11L1 and HOX11 expression were observed in one instance each, in the absence of detectable chromosomal abnormality of their respective loci, on chromosomes 2 and 10, respectively. HOX11L2 expression was associated with a chromosome 5q abnormality, the location of the HOX11L2 locus in each case tested. Finally, our data show that HOX11L2 expression was a suitable marker for minimal residual disease follow-up and was significantly associated with relapse (P =.02). PMID- 12130514 TI - Heterozygous PU.1 mutations are associated with acute myeloid leukemia. AB - The transcription factor PU.1 is required for normal blood cell development. PU.1 regulates the expression of a number of crucial myeloid genes, such as the macrophage colony-stimulating factor (M-CSF) receptor, the granulocyte colony stimulating factor (G-CSF) receptor, and the granulocyte-macrophage colony stimulating factor (GM-CSF) receptor. Myeloid cells derived from PU.1(-/-) mice are blocked at the earliest stage of myeloid differentiation, similar to the blast cells that are the hallmark of human acute myeloid leukemia (AML). These facts led us to hypothesize that molecular abnormalities involving the PU.1 gene could contribute to the development of AML. We identified 10 mutant alleles of the PU.1 gene in 9 of 126 AML patients. The PU.1 mutations comprised 5 deletions affecting the DNA-binding domain, and 5 point mutations in 1) the DNA-binding domain (2 patients), 2) the PEST domain (2 patients), and 3) the transactivation domain (one patient). DNA binding to and transactivation of the M-CSF receptor promoter, a direct PU.1 target gene, were deficient in the 7 PU.1 mutants that affected the DNA-binding domain. In addition, these mutations decreased the ability of PU.1 to synergize with PU.1-interacting proteins such as AML1 or c-Jun in the activation of PU.1 target genes. This is the first report of mutations in the PU.1 gene in human neoplasia and suggests that disruption of PU.1 function contributes to the block in differentiation found in AML patients. PMID- 12130515 TI - Targeted removal of PML-RARalpha protein is required prior to inhibition of histone deacetylase for overcoming all-trans retinoic acid differentiation resistance in acute promyelocytic leukemia. AB - All-trans retinoic acid (tRA)-induced differentiation in NB4 cells, a cell line derived from an acute promyelocytic leukemia patient with t(15;17) translocation, is markedly facilitated by sodium butyrate (NaB), a histone deacetylase inhibitor (HDACI), or by hexamethylene bisacetamide (HMBA), a non-HDACI tRA-differentiation inducer, as determined by nitroblue tetrazolium reduction. The tRA-induced expression of RIG-G, Bfl-1/A1, and p21(waf1) and, to a lesser extent, of CCAAT/enhancer binding protein-epsilon (C/EBPepsilon) are also enhanced by such combined treatments. Both responses are associated with a facilitated diminution of the leukemogenic PML-RARalpha protein and retained DeltaPML-RARalpha, a cleavage product. Treatment with tRA in tRA differentiation-resistant NB4 subclones R4 and MR-2 does not result in PML-RARalpha diminution and the tested gene expressions. Moreover, the addition of HMBA or NaB with tRA results in only minimal increase of differentiation in the tRA differentiation-resistant subclones. The increases in acetylated histone H3 (AcH3) and AcH4 in NaB-treated NB4, R4, and MR-2 cells are similar and do not correlate with the extent of differentiation induction when NaB and HMBA are given in combination with tRA. Arsenic trioxide (As2O3) treatment results in the total degradation of PML RARalpha without increasing AcH3 or AcH4 or inducing differentiation in R4 cells. As2O3 in combination with tRA induces gene (Bfl-1/A1 and C/EBPepsilon) expression and partial differentiation. Both NaB and HMBA addition to As2O3-plus-tRA-treated R4 cells further enhances differentiation. These results suggest that elimination of the dominant negative PML-RARalpha protein is required prior to inhibition of histone deacetylase to fully overcome tRA-differentiation resistance in APL cells. PMID- 12130516 TI - Several types of mutations of the Abl gene can be found in chronic myeloid leukemia patients resistant to STI571, and they can pre-exist to the onset of treatment. AB - Targeting the tyrosine kinase activity of BCR-ABL represents a very promising therapeutic strategy in chronic myeloid leukemia (CML). Despite strong efficacy of the tyrosine kinase inhibitor STI571, resistance has been observed in a significant proportion of patients in advanced CML stage or in Ph-positive acute lymphoid leukemia (ALL). We investigated in this study the mechanism of resistance to STI571 through point mutations in the tyrosine kinase domain and/or BCR-ABL gene amplification in 24 patients (16 in chronic phase and 8 in accelerated phase of the disease) who obtained no cytogenetic response to STI571 treatment. Screening for the already-described Thr315Ile point mutation in the ABL domain using a reverse transcription polymerase chain reaction restriction fragment length polymorphism (RT-PCR-RFLP) technique, 3 patients showed a proportion of mutated transcript at the time of resistance. The same technique failed to detect mutation at diagnosis, but a specific allele-specific oligonucleotide (ASO)-PCR on DNA for the Thr315Ile mutation and, after sequencing, for 2 newly described Phe311Leu and Met351Thr substitutions, showed the presence of rare mutated cells prior to STI571 therapy. Furthermore, the increased proportion of mutated cells during treatment detected by ASO-PCR strongly suggested clonal selection by the functional inhibiting effect of these mutations. Finally, no BCR-ABL gene amplification was detected by fluorescent in situ hybridization (FISH) in the 24 STI571-resistant patients. Our data support that in CML patients treated with STI571, ABL mutations are not restricted to the accelerated phase of the disease and that, at least in some cases, mutations seem to occur prior to STI571 therapy, probably as second mutational events during the course of CML. PMID- 12130517 TI - Engagement of ILT2/CD85j in Sezary syndrome cells inhibits their CD3/TCR signaling. AB - Extensive phenotype analysis of cutaneous T-cell lymphoma (CTCL) malignant cell lines revealed surface expression of receptors usually not detected on normal circulating CD4(+)CD45RO(+) lymphocytes. We previously found that CTCL malignant cells express the killer cell immunoglobulinlike receptor (KIR) KIR3DL2/CD158k, whereas they fail to express the other KIRs. In the present study, we report for the first time that the CD85j/immunoglobulin (Ig)-like transcript 2 (ILT2) receptor is found on Sezary cell lines and on circulating Sezary malignant CD4(+) cells, while it is hardly detectable on circulating CD4(+) lymphocytes from healthy individuals. We demonstrate that ILT2 is functional on CTCL cells, as its triggering leads to the recruitment of Src homology 2 domain-containing tyrosine phosphatase (SHP-1) and to the specific inhibition of CTCL malignant cell proliferation induced by CD3/T-cell receptor (TCR) stimulation. Interestingly, we found that separated CD4(+)ILT2(+) circulating malignant Sezary cells are less susceptible to anti-CD3 monoclonal antibody (mAb)-induced cell death than autologous CD4(+)ILT2(-) lymphocytes. Therefore, the resistance to apoptosis of Sezary cells may result from distinct mechanisms including cytokine-induced high levels of bcl-2 and specific expression of inhibitory receptors involved in lymphocyte survival. PMID- 12130518 TI - Deletion of leucine 61 in glucose-6-phosphate dehydrogenase leads to chronic nonspherocytic anemia, granulocyte dysfunction, and increased susceptibility to infections. AB - In this study the blood cells of 4 male patients from 2 unrelated families with chronic nonspherocytic anemia and recurrent bacterial infections were investigated. The activity of glucose-6- phosphate dehydrogenase (G6PD) in the red blood cells and in the granulocytes of these patients was below detection level. Moreover, their granulocytes displayed a decreased respiratory burst upon activation. Sequencing of genomic DNA revealed a novel 3-base pair (TCT) deletion in the G6PD gene, predicting the deletion of a leucine at position 61. The mutant G6PD protein was undetectable by Western blotting in the red blood cells and granulocytes of these patients. In phytohemagglutinin-stimulated lymphocytes the G6PD protein was present, but the amount of G6PD protein was strongly diminished in the patients' cells. Purified mutant protein from an Escherichia coli expression system showed decreased heat stability and decreased specific activity. Furthermore, we found that the messenger RNA of G6PD(180-182delTCT) is unstable, which may contribute to the severe G6PD deficiency observed in these patients. We propose the name "G6PD Amsterdam" for this new variant. PMID- 12130519 TI - Decreased susceptibility of leukemic cells with PIG-A mutation to natural killer cells in vitro. AB - The cloning of the PIG-A gene has facilitated the unraveling of the complex pathophysiology of paroxysmal nocturnal hemoglobinuria (PNH). Of current major concern is the mechanism by which a PNH clone expands. Many reports have suggested that an immune mechanism operates to cause bone marrow failure in some patients with PNH, aplastic anemia, and myelodysplastic syndromes. Because blood cells of PNH phenotype are often found in patients with these marrow diseases, one hypothesis is that the PNH clone escapes immune attack, producing a survival advantage by immunoselection. To test this hypothesis, we examined the sensitivity of blood cells, with or without PIG-A mutations, to killing by natural killer (NK) cells, using 51Cr-release assay in vitro. To both peripheral blood and cultured NK cells, PIG-A mutant cells prepared from myeloid and lymphoid leukemic cell lines were less susceptible than their control counterparts (reverted from the mutant cells by transfection with a PIG-A cDNA). NK activity was completely abolished with concanamycin A and by calcium chelation, indicating that killing was perforin-dependent. There were no differences in major histocompatibility (MHC) class I expression or sensitivity to either purified perforin or to interleukin-2-activated NK cells between PIG-A mutant and control cells. From these results, we infer that PIG-A mutant cells lack molecules needed for NK activation or to trigger perforin-mediated killing. Our experiments suggest that PIG-A mutations confer a relative survival advantage to a PNH clone, contributing to selective expansion of these cells in the setting of marrow injury by cytotoxic lymphocytes. PMID- 12130520 TI - Cell-surface expression of RhD blood group polypeptide is posttranscriptionally regulated by the RhAG glycoprotein. AB - In most cases, the lack of Rh in Rh(null) red cells is associated with RHAG gene mutations. We explored the role of RhAG in the surface expression of Rh. Nonerythroid HEK293 cells, which lack Rh and RhAG, or erythroid K562 cells, which endogenously express RhAG but not Rh, were transfected with RhD and/or RhAG cDNAs using cytomegalovirus (CMV) promoter-based expression vectors. In HEK293 cells, a low but significant expression of RhD was obtained only when RhAG was expressed at a high level. In K562 cells, as expected from the opposite effects of the phorbol ester 12-O-tetradecanoyl phorbol 13-acetate (TPA) on erythroid and CMV promoters, the levels of endogenous RhAG and recombinant RhD transcripts were substantially decreased and enhanced upon TPA treatment of RhD-transfected cells (K562/RhD), respectively. However, flow cytometry and fluorescence microscopy analysis revealed a decreased cell-surface expression of both RhAG and RhD proteins. Conversely, TPA treatment of RhAG-transfected cells increased both the transcript and surface expression levels of RhAG. When K562/RhD cells were cotransfected by the RhAG cDNA, the TPA-mediated induction of recombinant RhAG and RhD transcription was associated with an increased membrane expression of both RhAG and RhD proteins. These results demonstrate the role of RhAG as a strictly required posttranscriptional factor regulating Rh membrane expression. In addition, because the postulated 2:2 stoichiometry between Rh and RhAG observed in the native red cell membrane could not be obtained in cotransfected K562 cells, our study also suggests that as yet unidentified protein(s) might be involved for optimal membrane expression of Rh. PMID- 12130521 TI - Plasmodium falciparum cysteine protease falcipain-2 cleaves erythrocyte membrane skeletal proteins at late stages of parasite development. AB - Plasmodium falciparum-derived cysteine protease falcipain-2 cleaves host erythrocyte hemoglobin at acidic pH and specific components of the membrane skeleton at neutral pH. Analysis of stage-specific expression of these 2 proteolytic activities of falcipain-2 shows that hemoglobin-hydrolyzing activity is maximum in early trophozoites and declines rapidly at late stages, whereas the membrane skeletal protein hydrolyzing activity is markedly increased at the late trophozoite and schizont stages. Among the erythrocyte membrane skeletal proteins, ankyrin and protein 4.1 are cleaved by native and recombinant falcipain 2 near their C-termini. To identify the precise peptide sequence at the hydrolysis site of protein 4.1, we used a recombinant construct of protein 4.1 as substrate followed by MALDI-MS analysis of the cleaved product. We show that falcipain-2-mediated cleavage of protein 4.1 occurs immediately after lysine 437, which lies within a region of the spectrin-actin-binding domain critical for erythrocyte membrane stability. A 16-mer peptide containing the cleavage site completely inhibited the enzyme activity and blocked falcipain-2-induced fragmentation of erythrocyte ghosts. Based on these results, we propose that falcipain-2 cleaves hemoglobin in the acidic food vacuole at the early trophozoite stage, whereas it cleaves specific components of the red cell skeleton at the late trophozoite and schizont stages. It is the proteolysis of skeletal proteins that causes membrane instability, which, in turn, facilitates parasite release in vivo. PMID- 12130522 TI - Gamma-globulins prepared from sera of multiparous women bind anti-HLA antibodies and inhibit an established in vivo human alloimmune response. AB - It has previously been shown that sera from multiparous women have increased levels of anti-idiotypic antibodies specific for anti-HLA molecules. gamma Globulins prepared from these sera may be superior to commercial preparations of intravenous gamma-globulin (IVIg) for inhibiting HLA alloimmunization. To test this, F(ab')2 fragments prepared from either commercial IVIg or from the sera of men or multiparous women were coupled to CNBr-Sepharose and tested for their ability to bind F(ab')2 fragments derived from polyspecific anti-HLA sera. As determined by flow cytometry, compared with columns coated with F(ab')2 derived from commercial IVIg or sera from men, columns coated with F(ab')2 prepared from the sera of multiparous women bound significantly more anti-HLA. In addition, intact IgG molecules prepared from the sera of multiparous women significantly neutralized the reactivity of the anti-HLA F(ab')2 fragments. To determine whether the intact IgG molecules or their corresponding F(ab')2 fragments could affect in vivo alloimmunity, they were tested for their ability to inhibit an established IgG human alloimmune response in humanized severe combined immunodeficient (SCID) mice. Compared with commercial IVIg, when intact IgG or F(ab')2 fragments derived from multiparous women were administered to SCID mice making human anti-HLA antibodies, a significant reduction in anti-HLA reactivity was observed. The findings suggest that IgG molecules prepared from the sera of multiparous women have increased anti-idiotypic reactivity against anti-HLA antibodies, which can significantly inhibit an established human IgG alloimmune response in an Fc-independent manner. PMID- 12130523 TI - Temporary increase in the risk for recurrence during pregnancy in women with a history of venous thromboembolism. AB - There is strong evidence that the incidence of venous thromboembolism (VTE) is increased during pregnancy. However, it is unknown whether and to which extent pregnancy influences the risk for recurrent VTE in women with previous thrombosis. To investigate whether pregnancy temporarily increases the risk for recurrent thrombosis, we retrospectively evaluated the recurrence rate in 109 women who had at least one pregnancy after an episode of VTE by comparing the time period during pregnancy with the nonpregnant period. Forty-three women had a first recurrence during a total observation time of 1014 years. Eight events (73 observation years) occurred during pregnancy, and 35 events (941 observation years) occurred outside pregnancy. Recurrence rates per 100 patient-years were 10.9% during and 3.7% outside pregnancy. Relative risk during pregnancy was 3.5 (95% confidence interval, 1.6-7.8; P =.002). Our data suggest that pregnancy leads to a temporary increase in the risk for recurrent thrombotic events. PMID- 12130524 TI - Short deletion within the blood group Dombrock locus causing a Do(null) phenotype. AB - A new alteration of the blood group DO*A allele was identified in a female Do(null) donor from Reunion Island with allo- anti-DO3 in her serum; her parents are consanguineous. Because the amplification of the DO transcript failed, each exon and intron-exon junction from the DO gene were examined. After polymerase chain reaction (PCR) amplification and sequencing, the only deviation from the wild-type DO*A allele sequence was an 8-nucleotide deletion (nt 343-350) within exon 2. This short deletion generates a premature stop codon and encodes a truncated protein lacking the predicted functional motif of the adenosine diphosphate-ribosyltransferase enzyme and the glycosyl-phosphatidylinositol anchor motif essential for RBC membrane attachment. An allele-specific PCR to detect the DO(Delta8nt) deletion was developed. PMID- 12130525 TI - Complete remission through blast cell differentiation in PLZF/RARalpha-positive acute promyelocytic leukemia: in vitro and in vivo studies. AB - Acute leukemia with the t(11;17) expressing the PLZF-RARalpha gene fusion is a rare variant of acute promyelocytic leukemia (APL) that has been associated with poor clinical response to all-trans retinoic acid (ATRA) treatment. However, some recent reports have put into question the absolute refractoriness of this leukemia to ATRA. We describe here a patient with PLZF/RARalpha APL who was treated at relapse with ATRA and low-dose hydroxyurea. Complete hematologic remission was obtained through differentiation of leukemic blasts, as proven by morphologic, immunophenophenotypic, and genetic studies carried out in sequential bone marrow samples. Moreover, in vitro studies indicated that blast differentiation was potentiated by the addition of the histone deacetylase inhibitor tricostatin A, but not of hydroxyurea, to ATRA. Our findings indicate that the maturation block may be overcome and terminal differentiation obtained in this leukemia subset and support the view that sensitivity/refractoriness of this form to ATRA should be revisited. PMID- 12130526 TI - Overcoming STI571 resistance with the farnesyl transferase inhibitor SCH66336. AB - The development of chronic myeloid leukemia (CML) is dependent on the deregulated tyrosine kinase of the oncoprotein BCR-ABL. STI571 (imatinib mesylate), an abl tyrosine kinase inhibitor, has proven remarkably effective for the treatment of CML. However, resistance to STI571 because of enhanced expression or mutation of the BCR-ABL gene has been detected in patients. In the current study we show that the farnesyl transferase inhibitor (FTI) SCH66336 (lonafarnib) inhibits the proliferation of STI571-resistant BCR-ABL-positive cell lines and hematopoietic colony formation from peripheral blood samples of STI571-resistant patients with CML. Moreover, SCH66336 enhances STI571-induced apoptosis in STI571-sensitive cells and, in patients with STI571 resistance from gene amplification, cooperates with STI571 to induce apoptosis. Our data provide a rationale for combination clinical trials of STI571 and SCH66336 in CML patients and suggest that combination therapy may be effective in patients with STI571 resistance. PMID- 12130527 TI - A 2-kb c-mpl promoter fragment is sufficient to direct expression to the megakaryocytic lineage and sites of embryonic hematopoiesis in transgenic mice. AB - Thrombopoietin receptor c-mpl is expressed on hematopoietic progenitors and cells of the megakaryocytic lineage. The c-mpl promoter may, therefore, be useful for directing the expression of transgenes. We tested whether a 2-kb genomic DNA fragment comprising the putative c-mpl regulatory elements and most of the 5' untranslated region of mouse c-mpl is able to direct the expression of a reporter gene to hematopoietic cells in transgenic mice. As a reporter gene we used the human placental alkaline phosphatase (PLAP). In adult transgenic mice, PLAP expression was specifically detected in megakaryocytes and platelets. Embryos showed PLAP reporter gene expression already in the yolk sac at embryonic day 6.5 (E6.5) and in blood islands at E7.5. At E9.5, expression was found in blood vessels of the yolk sac and the embryo proper, followed by high levels of expression in the fetal liver at E11.5. Expression in E6.5 yolk sac is compatible with a function of c-mpl and its ligand, thrombopoietin, in the earliest stages of embryonic hematopoiesis. PMID- 12130528 TI - Transferrin receptor 2 (TfR2) and HFE mutational analysis in non-C282Y iron overload: identification of a novel TfR2 mutation. AB - Hereditary hemochromatosis (HH) is classically associated with a Cys282Tyr (C282Y) mutation of the HFE gene. Non-C282Y HH is a heterogeneous group accounting for 15% of HH in Northern Europe. Pathogenic mutations of the transferrin receptor 2 (TfR2) gene have been identified in 4 Italian pedigrees with the latter syndrome. The goal of this study was to perform a mutational analysis of the TfR2 and HFE genes in a cohort of non-C282Y iron overload patients of mixed ethnic backgrounds. Several sequence variants were identified within the TfR2 gene, including a homozygous missense change in exon 17, c2069 A- >C, which changes a glutamine to a proline residue at position 690. This putative mutation was found in a severely affected Portuguese man and 2 family members with the same genotype. In summary, pathologic TfR2 mutations are present outside of Italy, accounting for a small proportion of non-C282Y HH. PMID- 12130529 TI - Distinct functions for signal transducer and activator of transcription 1 and PU.1 in transcriptional activation of Fc gamma receptor I promoter. AB - The myeloid cell-specific expression and interferon-gamma (IFN-gamma) induction of Fc gamma receptor I (FcgammaRI) requires cooperation between PU.1 and signal transducer and activator of transcription 1 (Stat1) by means of mechanisms that are unknown. We found that PU.1 and Stat1 mediated distinct functions in the activation of FcgammaRI promoter. The basal activity of the natural FcgammaRI promoter was strictly dependent on PU.1, and IFN-gamma induction required both PU.1 and Stat1. Recruitment of TATA-binding protein (TBP) to the FcgammaRI promoter did not replace PU.1 in promoter activation, suggesting that TBP is not sufficient for FcgammaRI activation and that PU.1 mediates additional contacts with basal transcription machinery. In contrast, Stat1 did not interact with basal transcription machinery, but the Stat1-mediated activation of FcgammaRI promoter critically required CREB-binding protein (CBP)/p300. These results define functional cooperativity between PU.1 and Stat1 in FcgammaRI promoter activation, in which PU.1 appears to serve as a bridging factor with the basal transcription machinery and IFN-gamma-mediated induction of transcription occurs through recruitment of CBP/p300 by Stat1. PMID- 12130530 TI - Vesicle-associated membrane protein 3 (VAMP-3) and VAMP-8 are present in human platelets and are required for granule secretion. AB - Secretion of platelet granules is necessary for normal hemostasis. Platelet secretion requires soluble N-ethylmaleimide-sensitive factor attachment protein (SNAP) receptor (SNARE) complex formation between different members of the syntaxin, SNAP-25, and vesicle-associated membrane protein (VAMP) gene families. Using microcapillary reverse-phase high-performance liquid chromatography-nano electrospray tandem mass spectrometry, we identified VAMP-3 and VAMP-8 as VAMP isoforms coimmunoprecipitated from platelets with syntaxin 4. Immunoblotting experiments confirmed the presence of VAMP-3 and VAMP-8 but not VAMP-1 or VAMP-2 in platelets. To examine the effect of VAMP proteins on platelet secretion, soluble recombinant (r) VAMP-2, rVAMP-3, and rVAMP-8 were incubated with streptolysin O-permeabilized platelets. Secretion of alpha granules (monitored by flow cytometric measurement of P-selectin) was blocked, and dense-granule secretion (assessed by release of carbon 14-serotonin) was almost completely inhibited by rVAMP-3, whereas rVAMP-8 inhibited secretion of dense granules but not alpha granules. In contrast, rVAMP-2, which formed SNARE complexes in vitro, had no effect on platelet exocytosis. We conclude that VAMP-3 and VAMP-8 form SNARE complexes with platelet syntaxin 4 and are required for platelet granule secretion. PMID- 12130531 TI - Rapamycin inhibits macropinocytosis and mannose receptor-mediated endocytosis by bone marrow-derived dendritic cells. AB - Dendritic cells (DCs) are professional antigen-presenting cells (APCs) that use 2 major pathways for antigen uptake: constitutive macropinocytosis and mannose receptor-mediated endocytosis. Efficient endocytosis is critical for DCs to fulfill their sentinel function in immunity. We investigated the influence of the immunosuppressive macrolide rapamycin on macropinocytosis of fluorescein isothiocyanate (FITC)-albumin and mannose receptor-mediated endocytosis of FITC dextran by murine bone marrow-derived DCs by flow cytometry. The data show that (1) at a low, physiologically relevant concentration (1 ng/mL), rapamycin impairs macropinocytosis and mannose receptor-mediated endocytosis; (2) the effects are independent of DC maturation and can be demonstrated specifically in immature CD11c(+) major histocompatibility complex (MHC) class II(lo) DCs by 3-color flow cytometry; (3) inhibition of endocytosis is not related to apoptotic cell death; and (4) molar excess of the structurally related molecule FK506 inhibits the actions of rapamycin. The inhibitory effects of rapamycin on DC endocytosis were confirmed in vivo. To our knowledge, this is the first report that a clinically relevant immunosuppressant inhibits DC endocytosis. PMID- 12130532 TI - Activity of STI571 in chronic myelomonocytic leukemia with a platelet-derived growth factor beta receptor fusion oncogene. AB - Platelet-derived growth factor beta receptor (PDGFbetaR) fusion genes have been shown to be critical transforming oncogenes in a subset of patients with chronic myelomonocytic leukemia (CMML). The sensitivity of dysregulated tyrosine kinase oncogenes to the tyrosine kinase inhibitor STI571 (imatinib mesylate) makes it a potentially attractive treatment option in this subset of patients. We have recently cloned a novel member of the PDGFbetaR fusion oncogene family, rabaptin 5-PDGFbetaR. A patient with CMML carrying the rabaptin-5-PDGFbetaR fusion gene underwent allogeneic stem cell transplantation (SCT) and was monitored closely with a sensitive reverse transcriptase-polymerase chain assay to detect the novel fusion gene transcript. After achieving a molecular remission at 5 months after transplantation, 15 months after SCT the patient showed persistent and progressive evidence of molecular relapse. After demonstrating in vitro that cells transformed with this specific fusion oncogene are efficiently killed by STI571, the patient was started on STI571. The patient responded rapidly and entered molecular remission after 6 weeks of therapy, and he continues to be in remission 6 months later. These results suggest that STI571 may be an effective targeted therapy in patients with CMML related to PDGFbetaR fusion oncogenes. PMID- 12130533 TI - Histone modification and replacement in chromatin activation. PMID- 12130534 TI - NASty effects on fibrillin pre-mRNA splicing: another case of ESE does it, but proposals for translation-dependent splice site choice live on. PMID- 12130535 TI - A nonsense mutation in the fibrillin-1 gene of a Marfan syndrome patient induces NMD and disrupts an exonic splicing enhancer. AB - A nonsense mutation in the fibrillin-1 (FBN1) gene of a Marfan syndrome (MFS) patient induces in-frame exon skipping of FBN1 exon 51. We present evidence, based on both in vivo and in vitro experiments, that the skipping of this exon is due to the disruption of an SC35-dependent splicing enhancer within exon 51. In addition, this nonsense mutation induces nonsense-mediated decay (NMD), which degrades the normally spliced mRNA in the patient's cells. In contrast to NMD, skipping of FBN1 exon 51 does not require translation. PMID- 12130536 TI - Brn-1 and Brn-2 share crucial roles in the production and positioning of mouse neocortical neurons. AB - Formation of highly organized neocortical structure depends on the production and correct placement of the appropriate number and types of neurons. POU homeodomain proteins Brn-1 and Brn-2 are coexpressed in the developing neocortex, both in the late precursor cells and in the migrating neurons. Here we show that double disruption of both Brn-1 and Brn-2 genes in mice leads to abnormal formation of the neocortex with dramatically reduced production of layer IV-II neurons and defective migration of neurons unable to express mDab1. These data indicate that Brn-1 and Brn-2 share roles in the production and positioning of neocortical neuron development. PMID- 12130537 TI - Fission yeast CENP-B homologs nucleate centromeric heterochromatin by promoting heterochromatin-specific histone tail modifications. AB - Heterochromatin is a functionally important chromosomal component, especially at centromeres. In fission yeast, conserved heterochromatin-specific modifications of the histone H3 tail, involving deacetylation of Lys 9 and Lys 14 and subsequent methylation of Lys 9, promote the recruitment of a heterochromatin protein, Swi6, a homolog of the Drosophila heterochromatin protein 1. However, the primary determinants of the positioning of heterochromatin are still unclear. The fission yeast proteins Abp1, Cbh1, and Cbh2 are homologs of the human protein CENP-B that bind to centromeric alpha-satellite DNA and associate with centromeric heterochromatin. We show that the CENP-B homologs are functionally redundant at centromeres, and that Abp1 binds specifically to centromeric heterochromatin. In the absence of Abp1 or Cbh1, the centromeric association of Swi6 is diminished, resulting in a decrease in silencing of the region. CENP-B homolog double disruptants show a synergistic reduction of Swi6 at centromeric heterochromatin, indicating that the three proteins are functionally redundant in the recruitment of Swi6. Furthermore, using chromatin immunoprecipitation assays, we show that disruption of CENP-B homologs causes a decrease in heterochromatin specific modifications of histone H3. These results indicate that the CENP-B homologs act as site-specific nucleation factors for the formation of centromeric heterochromatin by heterochromatin-specific modifications of histone tails. PMID- 12130538 TI - G9a histone methyltransferase plays a dominant role in euchromatic histone H3 lysine 9 methylation and is essential for early embryogenesis. AB - Covalent modification of histone tails is crucial for transcriptional regulation, mitotic chromosomal condensation, and heterochromatin formation. Histone H3 lysine 9 (H3-K9) methylation catalyzed by the Suv39h family proteins is essential for establishing the architecture of pericentric heterochromatin. We recently identified a mammalian histone methyltransferase (HMTase), G9a, which has strong HMTase activity towards H3-K9 in vitro. To investigate the in vivo functions of G9a, we generated G9a-deficient mice and embryonic stem (ES) cells. We found that H3-K9 methylation was drastically decreased in G9a-deficient embryos, which displayed severe growth retardation and early lethality. G9a-deficient ES cells also exhibited reduced H3-K9 methylation compared to wild-type cells, indicating that G9a is a dominant H3-K9 HMTase in vivo. Importantly, the loss of G9a abolished methylated H3-K9 mostly in euchromatic regions. Finally, G9a exerted a transcriptionally suppressive function that depended on its HMTase activity. Our results indicate that euchromatic H3-K9 methylation regulated by G9a is essential for early embryogenesis and is involved in the transcriptional repression of developmental genes. PMID- 12130539 TI - Cyclin D-cdk4 activity modulates the subnuclear localization and interaction of MEF2 with SRC-family coactivators during skeletal muscle differentiation. AB - Prior work has indicated that D-type cyclin-cdk4 complexes, which are only active in proliferating cells, can suppress the skeletal muscle differentiation program in proliferating myoblasts. In this study, we show that cyclin D-cdk activity can block the activity of the MEF2 family of transcriptional regulators, which are crucial regulators of skeletal muscle gene expression. We have found that cyclin D-cdk activity blocks the association of MEF2C with the coactivator protein GRIP 1 and thereby inhibits the activity of MEF2. During skeletal muscle differentiation, GRIP-1 is localized to punctate nuclear structures and can apparently tether MEF2 to such structures. Cotransfection of GRIP-1 can both potentiate the transcriptional activity of a Gal4-MEF2C construct and induce MEF2C localization to punctate nuclear structures. Consistent with the absence of punctate nuclear GRIP-1 in proliferating myoblasts, we have found that ectopic cyclin D-cdk4 expression disrupts the localization of both GRIP-1 and MEF2C to these punctate subnuclear structures. Our findings indicate that cyclin D-cdk4 activity represses skeletal muscle differentiation in proliferating cells by blocking the association of MEF2 with the coactivator GRIP-1 and concomitantly disrupts the association of these factors with punctate nuclear subdomains within the cell. PMID- 12130540 TI - Inhibition of apoptosis by ATFx: a novel role for a member of the ATF/CREB family of mammalian bZIP transcription factors. AB - The mammalian ATF/CREB family of transcription factors comprises a large group of basic-region leucine zipper (bZIP) proteins whose members mediate diverse transcriptional regulatory functions. Here we report that expression of a specific mouse ATF gene, ATFx, is down-regulated in a variety of cells undergoing apoptosis following growth factor deprivation. When stably expressed in an interleukin 3 (IL-3)-dependent cell line, ATFx suppresses apoptosis resulting from cytokine deprivation. Conversely, a dominant-negative ATFx mutant induces apoptosis of cells cultured in the presence of growth factors. We also show that 24p3, a secreted lipocalin that induces apoptosis when added to hematopoietic cells, represses ATFx expression. However, constitutive expression of ATFx renders cells resistant to 24p3-mediated apoptosis. Collectively, our results indicate that ATFx is an anti-apoptotic factor, a novel role for an ATF protein. PMID- 12130541 TI - C. elegans EOR-1/PLZF and EOR-2 positively regulate Ras and Wnt signaling and function redundantly with LIN-25 and the SUR-2 Mediator component. AB - In Caenorhabditis elegans, Ras/ERK and Wnt/beta-catenin signaling pathways cooperate to induce P12 and vulval cell fates in a Hox-dependent manner. Here we describe eor-1 and eor-2, two new positively acting nuclear components of the Ras and Wnt pathways. eor-1 and eor-2 act downstream or in parallel to ERK and function redundantly with the Mediator complex gene sur-2 and the functionally related gene lin-25, such that removal of both eor-1/eor-2 and sur-2/lin-25 mimics the removal of a main Ras pathway component. Furthermore, the eor-1 and eor-2 mutant backgrounds reveal an essential role for the Elk1-related gene lin 1. eor-1 and eor-2 also act downstream or in parallel to pry-1 Axin and therefore act at the convergence of the Ras and Wnt pathways. eor-1 encodes the ortholog of human PLZF, a BTB/zinc-finger transcription factor that is fused to RARalpha in acute promyelocytic leukemia. eor-2 encodes a novel protein. EOR-1/PLZF and EOR-2 appear to function closely together and cooperate with Hox genes to promote the expression of Ras- and Wnt-responsive genes. Further studies of eor-1 and eor-2 may provide insight into the roles of PLZF in normal development and leukemogenesis. PMID- 12130542 TI - The transcriptional repressor Brinker antagonizes Wingless signaling. AB - In the embryonic midgut of Drosophila, Wingless (Wg) signaling elicits threshold specific transcriptional response, that is, low-signaling levels activate target genes, whereas high-signaling levels repress them. Wg-mediated repression of the HOX gene Ultrabithorax (Ubx) is conferred by a response sequence within the Ubx B midgut enhancer, called WRS-R. It further depends on the Teashirt (Tsh) repressor, which acts through the WRS-R without binding to it. Here, we show that Wg-mediated repression of Ubx B depends on Brinker, which binds to the WRS-R. Furthermore, Brinker blocks transcriptional activation by ubiquitous Wg signaling. Brinker binds to Tsh in vitro, recruits Tsh to the WRS-R, and we find mutual physical interactions between Brinker, Tsh, and the corepressor dCtBP. This suggests that the three proteins may form a ternary repressor complex at the WRS-R to quench the activity of the nearby-bound dTCF/Armadillo transcription complex. Finally, brinker and tsh produce similar mutant phenotypes in the ventral epidermis, and double mutants mimic overactive Wg signaling in this tissue. This suggests that Brinker may have a widespread function in antagonizing Wg signaling. PMID- 12130543 TI - The Wilms tumor suppressor WT1 regulates early gonad development by activation of Sf1. AB - In mammals, several genes including the Wilms tumor suppressor gene Wt1, the Lim homeobox gene Lhx9, and the gene encoding steroidogenic factor 1 (Sf1) have been implicated in the development of the indifferent gonad prior to sexual differentiation. Interactions among these genes have not yet been elucidated. Using biochemical and genetic experiments, we demonstrate here that WT1 and LHX9 function as direct activators of the Sf1 gene. Interestingly, only the -KTS form of WT1 is able to bind to and transactivate the Sf1 promoter. This observation is consistent with differential roles for the -KTS and +KTS variants of WT1 which have been postulated on the basis of human disorders such as the Frasier syndrome. Our data suggest a pathway in which the products of the Wt1 and Lhx9 genes activate expression of Sf1 and thus mediate early gonadogenesis. PMID- 12130545 TI - Minireview: genetic models for the study of gonadotropin actions. AB - Fertility in both sexes relies on complex physiological and molecular processes with many levels of regulation, and our ability to alter the mammalian genome using transgenic technology has greatly enhanced our understanding of these processes. There are numerous commonalities in human and mouse physiology, and the list of mouse models recapitulating recognized and idiopathic human reproductive defects is growing at an ever-increasing rate. In this review, we focus on genetic models of gonadotropin actions, summarizing features of transgenic mice that phenocopy defects in gonadotropin production and gonadotropin receptor responses seen in patients. In addition, we provide examples of mouse models with genetic alterations influencing pituitary FSH and LH production and their effects. These include: 1) transgenic mice with aberrations in steroid hormone, inhibin, and activin feedback pathways; 2) knockouts that demonstrate specific in vivo functions of pituitary transcription factors; and 3) models with alterations in other pituitary hormones, IGF-1, and leptin signaling pathways, which affect both the central and peripheral endocrine axis. What we have to learn from these and other models will continue to revise our conceptions of physiology, identify new targets for contraception, and improve our tools for understanding, diagnosing, and treating cases of human endocrinopathies and pathologies of the reproductive tissues. PMID- 12130544 TI - TFIID and human mediator coactivator complexes assemble cooperatively on promoter DNA. AB - Activator-mediated transcription complex assembly on templates lacking chromatin requires the interaction of activators with two major coactivator complexes: TFIID and mediator. Here we employed immobilized template assays to correlate transcriptional activation with mediator and TFIID recruitment. In reactions reconstituted with purified proteins, we found that activator, TFIID, and mediator engage in reciprocal cooperative interactions to form a complex on promoter DNA. Preassembly of the coactivator complex accelerates the rate of transcription in a cell-free system depleted of TFIID and mediator. Our data argue that this coactivator complex is an intermediate in the assembly of an active transcription complex. Furthermore, the reciprocity of the interactions demonstrates that the complex could in principle be nucleated with either TFIID or mediator, implying that alternative pathways could be utilized to generate diversity in the way activators function in vivo. PMID- 12130546 TI - Editorial: Pitx-2 mutants and somatolactotroph gene regulation--deciphering the combinatorial code. PMID- 12130547 TI - Regulation of prolactin, GH, and Pit-1 gene expression in anterior pituitary by Pitx2: An approach using Pitx2 mutants. AB - The transcription factor Pitx2 is required for the morphogenesis of anterior structures such as the eye, teeth, and anterior pituitary. We investigated the functional properties of Pitx2 missense mutants previously reported in Axenfeld Rieger syndrome, using reporter genes under the control of pituitary target gene [human (h)PRL, hGH, hPit-1] promoters transfected in nonpituitary and pituitary cell lines. The five mutants appeared to be transcriptionally defective despite conserved DNA-binding in CV1 cells. In addition, one mutation, R91P, almost completely blocked the wt-Pitx2-induced activation of the target promoters, prevented the Pitx2/Pit-1 synergistic activation of the hPRL promoter, and was able to counteract the Pitx1-driven transactivation effects. The dominant negative properties of this mutant were further established in cells endogenously expressing Pitx2 because transfection of R91P in GH4C1 somatolactotroph cells resulted in a dose-dependent inhibition of basal activities of the pituitary promoters. These results, which show that Pitx2 mutants are defective in activating pituitary target genes, confirm the critical role of this homeodomain factor in the differentiated functions of the pituitary somatolactotroph cells. Furthermore, these results might form the basis for future experiments because dominant negative forms of Pitx2 such as R91P might provide instructive tools to further delineate the detailed mechanisms mediating Pitx2 functions in cell proliferation and differentiation. PMID- 12130548 TI - Marked regulatory shifts in gonadal, adrenal, and metabolic system responses to repeated restraint stress occur within a 3-week period in pubertal male rats. AB - We compared testosterone (T), corticotropin (ACTH), corticosterone (B), and leptin responses to three daily 3-h bouts of restraint and blood sampling as well as energy balance of male rats in early (40 d of age) and late (60 d of age) puberty. Rats either remained intact or were adrenalectomized and replaced with B clamped at basal mean values (ADX+B). Hormones, weight gain, food intake, and fat depot weight were measured during or after the days of stress. The major effects of restraint on T, ACTH, and energetic responses were age dependent, but clamped B affected the effects of restraint seen in intact rats at each age. T secretion was inhibited in 40-d-old and was stimulated in 60-d-old rats after restraint. ACTH responses were high, but diminished with repetition of stress in intact, but not ADX+B, 40-d-old rats. ACTH responses were lower, but constant across days, in both intact and ADX+B 60-d-old rats. Younger rats gained weight during the period of stress, whereas older rats stopped gaining weight. We conclude that the central regulation of stress responses shifts markedly between early and late puberty, although stress-induced B responses are important at both ages. In early puberty, priority is placed on maintaining normal ponderal growth, whereas in late puberty, priority is placed on maintaining reproductive capability. PMID- 12130549 TI - Ligands for the peroxisomal proliferator-activated receptor gamma and the retinoid X receptor inhibit aromatase cytochrome P450 (CYP19) expression mediated by promoter II in human breast adipose. AB - Local estrogen biosynthesis in breast adipose tissue, catalyzed by P450 aromatase, contributes to the growth of breast carcinomas. Aromatase expression is regulated by a number of alternative promoters, and in normal adipose tissue it is primarily regulated via the distal promoter I.4. However, in breast adipose containing a tumor, aromatase expression is regulated by the proximal promoter II in response to tumor-derived factors. Previously we have shown that peroxisomal proliferator-activated receptor gamma (PPARgamma) ligands inhibit aromatase expression in normal breast adipose tissue mediated by promoter I.4. In the present study, we investigated the effects of the PPARgamma ligand troglitazone and the retinoid X receptor (RXR) ligand LG101305 on aromatase expression mediated by promoter II. In cultured human breast adipose stromal cells, troglitazone or LG101305 alone inhibited aromatase activity and expression stimulated by inducers of promoter II, in a concentration-dependent manner, and this inhibition was greater in the presence of both ligands. Reporter gene assays showed that troglitazone and LG101305 inhibit transcription from promoter II of the CYP19 gene. However, EMSAs showed that PPARgamma and RXRalpha do not bind to promoter II of the CYP19 gene, indicating that PPARgamma- and RXR-mediated inhibition of aromatase expression via promoter II occurs through an indirect mechanism of action. Because ligands for PPARgamma and RXR inhibit aromatase expression in healthy breast adipose (via promoter I.4), as well as expression induced by tumor-derived factors (via promoter II), such compounds could find utility in the treatment of estrogen-dependent breast cancers. PMID- 12130550 TI - Antigen presentation by vaginal cells: role of TGFbeta as a mediator of estradiol inhibition of antigen presentation. AB - Estradiol is known to inhibit antigen presentation in the vagina. We report here that TGFbeta mediates the action of estradiol on vaginal antigen presenting cells (APC). When vaginal APC from ovariectomized rats were incubated with increasing concentrations of TGFbeta1 and TGFbeta2 in the presence of ovalbumin-specific T cells and ovalbumin, both TGFbeta1 and TGFbeta2 inhibited vaginal cell antigen presentation, whereas IL-6, IL-10, and TNFalpha had no consistent effect. In other experiments, estradiol-induced inhibition of antigen presentation by vaginal cells was partially reversed when vaginal APC were incubated with anti TGFbeta antibody. In contrast, anti-TNFalpha, anti-IL-6, and anti-IL-10 had no effect on antigen presentation. The effect of anti-TGFbeta was seen with vaginal APC from ovariectomized rats treated with estradiol for 1 d as well as 3 d. Finally, analysis of TGFbeta in the culture media of vaginal cells from saline- and estradiol-treated rats indicated that the TGFbeta produced is biologically active. In response to estradiol, vaginal cell production of TGFbeta was significantly greater than that seen with control cells. These studies suggest that estradiol regulation of antigen presentation by vaginal cells is mediated through the local production of TGFbeta by vaginal cells. PMID- 12130551 TI - Mechanism of selective retinoid X receptor agonist-induced hypothyroidism in the rat. AB - The retinoid X receptor (RXR) agonist bexarotene can cause clinically significant hypothyroidism in cutaneous T cell lymphoma patients. The mechanism by which the RXR agonist produces this effect is unclear. We have studied the impact of a selective RXR agonist (rexinoid), LG100268, on rat thyroid axis hormones and show that the acute phase of hypothyroidism is associated with reduced pituitary TSH secretion. A single oral administration of LG100268 to naive Sprague Dawley rats causes a rapid and statistically significant decline in TSH levels (apparent in 0.5-1 h). Total T(4) and T(3) levels decline more gradually, reaching statistical significance 24 h after treatment. Increasing doses of LG100268 produce greater suppression of thyroid axis hormones. To investigate the mechanism(s) mediating this suppression, we determined pituitary TSHbeta mRNA, TSH protein levels, and TRH-stimulated TSH secretion. Two hours after treatment, neither TSHbeta mRNA nor TSH protein levels were altered by LG100268. However, LG100268 treatment reduced the area under the curve for TRH-stimulated TSH secretion by 54%. We have identified an unexpected mechanism by which rexinoids induce hypothyroidism by acutely reducing TSH secretion from the anterior pituitary. This mechanism is independent of the rexinoid's previously demonstrated inhibition of TSHbeta gene transcription. PMID- 12130552 TI - Correlation between basal signaling and internalization of thyrotropin-releasing hormone receptors: evidence for involvement of similar receptor conformations. AB - Previous studies have shown that rat thyrotropin-releasing hormone (TRH) receptor type 2 exhibits higher basal signaling activity and internalizes more rapidly upon agonist binding than rat TRH receptor type 1. The mouse TRH receptor type 2 (mR2) was recently cloned and, similar to its rat homolog, shows a higher basal signaling activity than mR1. Taking advantage of the high degree of sequence homology between mR1 and mR2, we used chimeras/mutants of these receptors to gain insight into the properties of the receptors that influence internalization and basal signaling. Chimeric receptors that have the mR1 extracellular and transmembrane domains with the carboxyl terminus and intracellular loops of mR2 (R1/R2-tail; R1/R2-I3,tail; R1/R2-I2,3,tail; R1/R2-I1,2,3,tail) exhibited internalization rates and basal activities that were similar to that of mR1. In contrast, a chimeric receptor with the extracellular and transmembrane domains of mR2 and the carboxyl terminus of mR1 exhibited the more rapid internalization rate and higher basal signaling activity characteristic of mR2. We showed previously that mutation of a highly conserved tryptophan to alanine caused mR1 to exhibit a high basal signaling activity and rapid internalization rate. In contrast, mutation of this tryptophan to alanine in mR2 decreased the rate of internalization and inhibited basal signaling activity. The rates of receptor internalization did not correlate with the binding affinities, coupling efficiencies, or potencies of the receptors. Thus, we observed that receptors with more rapid internalization rates showed relatively higher basal signaling activities, whereas receptors with lower basal signaling activities showed slower internalization rates. These data suggest that similar receptor conformations are required for productive coupling to signaling G proteins and to proteins involved in internalization. PMID- 12130553 TI - Deficiency of acyl coenzyme a:diacylglycerol acyltransferase 1 increases leptin sensitivity in murine obesity models. AB - Acyl coenzyme A:diacylglycerol acyltransferase 1 (DGAT1) is one of two known enzymes that catalyze the final step in mammalian triglyceride synthesis. We have reported that DGAT1-deficient mice have increased insulin and leptin sensitivity, likely accounting for their protection against diet-induced obesity and insulin resistance. Here we show that DGAT1 deficiency enhanced the response to peripheral leptin infusion in Agouti yellow and leptin-deficient (ob/ob) mice, two genetic models of obesity and insulin resistance. Interestingly, DGAT1 deficiency did not enhance the response to intracerebroventricular leptin infusion. Moreover, DGAT1 deficiency did not alter the expression of key hypothalamic genes involved in leptin signaling or in the regulation of food intake and energy expenditure. Thus, the leptin-sensitizing effect of DGAT1 deficiency is present in both leptin-resistant and leptin-deficient genetic models of obesity and may occur in part by enhancing the effects of leptin in peripheral tissues. PMID- 12130554 TI - Role of estrogen receptor-beta in regulation of vasopressin and oxytocin release in vitro. AB - In rats, the magnocellular neurons that produce vasopressin (VP) and oxytocin (OT) express estrogen receptor-beta (ER-beta). Physiological concentrations of estrogen (E2) inhibit N-methyl-D-aspartate (NMDA)-stimulated VP and OT release from explants of the hypothalamo-neurohypophyseal system (HNS). To determine whether ER-beta mediates inhibition by E2, HNS explants were perifused with and without NMDA (50 microM) in the presence of E2 (50 pg/ml), E2 coupled to BSA (E2:BSA), genistein (100 nM, a phytoestrogen with affinity for ER-beta), or tetrahydrochrysene-R,R,-enantiomer (R,R-THC, a ligand that acts as an agonist on ER-alpha but an antagonist on ER-beta). VP and OT released into the perifusate were measured by RIA. E2 and genistein inhibited NMDA-stimulated VP release, but E2:BSA and R,R,THC were not effective inhibitors. However, R,R,THC blocked E2 inhibition of NMDA-stimulated VP release. The inability of E2:BSA to mimic the effect of E2 indicates that E2 inhibition is not mediated by membrane receptors. The ability of genistein to mimic the effect of E2 suggests that the effect is mediated by ERbeta. This interpretation is supported by the ability of R,R,THC to block but not to mimic the effect of E2. Thus, E2 inhibition of NMDA-stimulated VP and OT release may be mediated by ER-beta. PMID- 12130555 TI - Water permeability of an ovarian antral follicle is predominantly transcellular and mediated by aquaporins. AB - Ovarian folliculogenesis is characterized, in part, by the formation and expansion of the fluid-filled antrum. Development of this cavity requires water influx, which may occur by transcellular or pericellular transport mechanisms. To assess the contribution of these mechanisms to the water permeability of an antral follicle, the rate of (3)H(2)O and (14)C-inulin (a complex sugar restricted to the extracellular compartment) uptake into isolated follicles was determined. The rate of H(2)O movement was 3.5-fold greater than that of inulin, suggesting that water enters a follicle primarily by transcellular pathways. Preincubation of the follicles with 50 microM HgCl(2) [a nonspecific aquaporin (Aqp) inhibitor] decreased H(2)O movement to levels seen with inulin, indicating that transcellular water movement is mediated through Aqp. To demonstrate the functional presence of Aqp in granulosa cells, we show that swelling in response to a hypotonic insult is attenuated by preincubation with 50 microM HgCl(2). Flow cytometry demonstrated the presence of Aqps-7, -8, and -9, thus identifying candidate Aqp potentially mediating water movement into antral follicles. These results suggest that water permeability of antral follicles occurs primarily through transcellular mechanisms, which may be mediated by Aqps -7, -8, and/or -9 in granulosa cells. PMID- 12130556 TI - The phenotype of the aromatase knockout mouse reveals dietary phytoestrogens impact significantly on testis function. AB - Estrogen is synthesized in the testis, both in Leydig cells and seminiferous epithelium, and its importance in spermatogenesis is highlighted by the phenotype of the aromatase knockout (ArKO) mouse. These mice are unable to synthesize endogenous estrogens. The males develop postmeiotic defects by 18 wk of age. We hypothesized that maintenance of spermatogenesis in younger animals may be mediated by exogenous estrogenic substances. Dietary soy meal, contained in almost all commercial rodent diets, provides a source of estrogenic isoflavones. We thus investigated spermatogenesis in wild-type and ArKO mice raised on a diet containing soy, compared with a soy-free diet, to elucidate the biological action of phytoestrogens on the testis. In ArKO mice, dietary phytoestrogens could partially prevent disruptions to spermatogenesis, in that they prevented the decline in germ cell numbers. They also seemed to maintain Sertoli cell function, and they blocked elevations in FSH. The impairment of spermatogenesis seen in soy free ArKOs occurred in the absence of a decreased gonadotropic stimulus, suggesting that the effects of dietary phytoestrogens are independent of changes to the pituitary-gonadal axis. Our study highlights the importance of estrogen in spermatogenesis and shows that relatively low levels of dietary phytoestrogens have a biological effect in the testis. PMID- 12130557 TI - Content and activity of cAMP response element-binding protein regulate platelet derived growth factor receptor-alpha content in vascular smooth muscles. AB - Experiments in vascular smooth muscle cells (SMCs) indicate that the transcription factor cAMP response element-binding protein (CREB), the cyclic nucleotide response element-binding protein, suppresses expression of the platelet-derived growth factor-alpha receptor gene (PDGFRalpha). Adenovirus mediated expression of constitutively active CREB mutants decreases PDGFRalpha mRNA, PDGFRalpha protein, and PDGFRalpha promoter-luciferase reporter activity in cultured SMCs. Expression of dominant negative CREB protein, A-CREB, increases PDGFRalpha protein content and the PDGFRalpha-promoter activity in SMCs. Active CREB prevents activation of PDGFRalpha promoter-luciferase reporter activity by CCAAT/enhancer-binding protein-delta (C/EBPdelta), shown to mediate IL-1beta stimulation of PDGFRalpha expression. Exposure of cultured SMCs to high glucose or reactive oxidant stress, which decrease CREB protein content and activity, increases PDGFRalpha protein content and promoter activity. Expression of active CREB blunts reactive oxidant stress-induced PDGFRalpha accumulation in SMCs. Loss of CREB protein in aortic walls of rats with streptozotocin-induced diabetes is accompanied by an increase in PDGFRalpha content. In Ob/Ob mice (which demonstrate reduced aortic wall CREB content vs. Ob/- controls), treatment with the peroxisomal proliferator-activated receptor gamma rosiglitazone increases CREB content and decreases PDGFRalpha content in the aortic wall. Thus, both in vitro and in vivo loss of CREB content and activity and subsequent accumulation of PDGFRalpha may contribute to SMC activation during diabetes. PMID- 12130558 TI - Dihydrotestosterone inhibits granulosa cell proliferation by decreasing the cyclin D2 mRNA expression and cell cycle arrest at G1 phase. AB - Hyperandrogenism is known to perturb ovarian physiology resulting in anovulatory conditions. In the ovary, androgens produced by theca-interstitial cells are converted to estrogens in granulosa cells under the influence of FSH and LH. In some of the target organs, including the ovary, androgens are also converted into their 5alpha reduced metabolites. In the present study, we examined the molecular mechanism by which dihydrotestosterone (DHT), a 5alpha reduced metabolite of testosterone, mediates the inhibition of granulosa cell proliferation, using a rat model. Immature female rats were primed with estradiol, followed by DHT administration for 2 d and granulosa cells were cultured in the presence or absence of forskolin. Granulosa cells from the DHT-treated rats showed reduced [(3)H]thymidine incorporation into DNA and reduced cell number in response to forskolin stimulation, compared with control. The decreased responsiveness of DHT treated granulosa cells to forskolin was not due to increased apoptosis because the expression of cleaved caspase 3 remained the same in both control and DHT exposed granulosa cells stimulated with forskolin. Forskolin treatment stimulated the expression of cyclin D2 mRNA in control granulosa cells, whereas DHT treatment abolished this response. In vitro DHT treatment of granulosa cells for 48 h resulted in a cell cycle arrest with 70% of cells at G1 phase and 26% at S phase, and control cells exhibited a distribution of 42% and 55% at G1 and S phase, respectively. In conclusion, the present study shows that DHT inhibits the granulosa cell proliferation through a decrease in cyclin D2 mRNA expression, which leads to cell cycle arrest at the G1 phase. PMID- 12130559 TI - Expression of microsomal prostaglandin E synthase in bovine endometrium: coexpression with cyclooxygenase type 2 and regulation by interferon-tau. AB - Prostaglandins (PGs) are important regulators of reproductive functions. In ruminants, interferon (IFN)-tau is the embryonic signal responsible for recognition of pregnancy. This is effected by a reduction of the production of PGF(2alpha) relative to PGE(2.) This may be accomplished by a decrease in PGF(2alpha) production, but a stimulation of PGE(2) via the PGE synthase might also be involved. The purpose of the present study was to confirm the presence of PGE synthase (PGES) in the bovine endometrium, identify the factors affecting its expression, and compare it with that of cyclooxygenase-2 (COX-2). This was done by Northern blot analysis using primary cultures of bovine epithelial and stromal cells of the endometrium and bovine endometrial cell line. PGES mRNA expression was increased in the presence of lipopolysaccharides, TNF-alpha, and IFN-tau in stromal cells and IFN-tau in epithelial cells. In stromal cells, IFN-tau induced a rapid increase of PGES and COX-2 mRNA expression. In bovine endometrial cells, phorbol 12-myristate 13-actetate increased PGES mRNA, COX-2 mRNA and PGE(2) production. These results suggest that in endometrial cells, the expression of PGE synthase is correlated with that of COX-2 and is an important enzyme for the production of PGE(2). Increasing this production will modulate the PGE(2)/PGF(2alpha) ratio and contribute to establishment of pregnancy. PMID- 12130560 TI - The somatostatin analog octreotide inhibits GH-stimulated, but not IGF-I stimulated, bone growth in hypophysectomized rats. AB - IGF-I mediates growth-promoting actions of GH. In the present study we investigated whether the somatostatin analog octreotide blunts the stimulatory effects of GH and/or IGF-I on bone growth in hypophysectomized rats infused for 6 d with vehicle, GH, or IGF-I. We found that octreotide significantly suppressed the GH-induced rise in liver IGF-I mRNA (-27%) and peptide (-32%) and the serum IGF-I level (-26%) and concomitantly inhibited GH-stimulated, but not IGF-I stimulated, body weight gain (-31%), tibial epiphyseal width (-14%), and bone growth rate (-24%). Furthermore, octreotide significantly reduced the GH-induced increase in the number of IGF-I immunoreactive chondrocytes in all layers (except in the upper hypertrophic zone) of the tibial growth plate cartilage (P < 0.0001 for stem cell and proliferative zone; P < 0.0005 for lower hypertrophic zone). These findings demonstrate that octreotide does not interfere with IGF-I action, but does interfere with local GH-stimulated IGF-I production in the growth plate. Thus, besides inhibiting pituitary GH secretion, octreotide exerts inhibitory peripheral effects on GH-stimulated longitudinal bone growth. PMID- 12130561 TI - Hepatocyte growth factor is required for progestin-induced epithelial cell proliferation and alveolar-like morphogenesis in serum-free culture of normal mammary epithelial cells. AB - The steroid hormones, estrogen and progesterone, are required for mammary epithelial cell proliferation and alveolar morphogenesis in vivo. We have developed a minimally supplemented, serum-free medium, collagen gel primary mammary culture system to determine the mechanism of progestin-induced proliferation and alveolar morphogenesis. In epithelial cells cultured alone, treatment with progestin (R5020) alone produced a lumen within the epithelial organoids, but did not stimulate epithelial cell proliferation. The formation of lumens was associated with increased apoptosis, targeted within the organoids. We have previously reported that in our culture system hepatocyte growth factor (HGF) increases epithelial cell proliferation and induces a tubulo-ductal morphological response. In the present report we show that treatment with HGF and progestin (R5020) further increases epithelial proliferation above that with HGF alone and also produces an alveolar-like morphology similar to that observed in vivo in response to progestin treatment. To the best of our knowledge this is the first in vitro demonstration of both progestin-induced proliferation and alveolar like morphogenesis of normal nonpregnant mouse mammary epithelial cells in vitro. These results suggest that HGF may play a crucial role in progestin-induced proliferation and morphogenesis in vivo. PMID- 12130562 TI - The glucocorticoid receptor represses the positive autoregulation of the trout estrogen receptor gene by preventing the enhancer effect of a C/EBPbeta-like protein. AB - Stress and cortisol are known to have negative effects on vitellogenesis in oviparous species. This provides a physiological context in which to explore in more detail the molecular mechanisms involved in transcriptional interferences between two steroids receptors, the estradiol receptor (ER) and the glucocorticoid receptor (GR). We have previously shown that the cortisol inhibitory effect on rainbow trout (rt) vitellogenesis is the result of a repression of the estradiol-induced ER-positive autoregulation by activated GR. In the present study, we demonstrate that the GR repression involves a proximal region of the rtER promoter that is unable to bind GR. This inhibition is counteracted in part by the orphan receptor COUP-TF1 that has been previously shown to cooperate with ERs on the same promoter. A detailed analysis allowed us to identify a C/EBPbeta-like protein that is implicated in both the maximal stimulatory effect of estradiol and the GR repression. Indeed, GR, through its DNA-binding domain, suppresses the binding of C/EBPbeta on the rtER promoter by protein-protein interactions and thereby prevents the enhancer effect of this transcription factor. PMID- 12130563 TI - High glucose stimulates angiotensinogen gene expression via reactive oxygen species generation in rat kidney proximal tubular cells. AB - The present studies investigated whether the effect of high glucose levels on angiotensinogen (ANG) gene expression in kidney proximal tubular cells is mediated via reactive oxygen species (ROS) generation and p38 MAPK activation. Rat immortalized renal proximal tubular cells (IRPTCs) were cultured in monolayer. Cellular ROS generation and p38 MAPK phosphorylation were assessed by lucigenin assay and Western blot analysis, respectively. The levels of immunoreactive rat ANG secreted into the media and cellular ANG mRNA were determined by a specific RIA and RT-PCR, respectively. High glucose (25 mM) evoked ROS generation and p38 MAPK phosphorylation as well as stimulated immunoreactive rat ANG secretion and ANG mRNA expression in IRPTCs. These effects of high glucose were blocked by antioxidants (taurine and tiron), inhibitors of mitochondrial electron transport chain complex I (rotenone) and II (thenoyltrifluoroacetone), an inhibitor of glycolysis-derived pyruvate transport into mitochondria (alpha-cyano-4-hydroxycinnamic acid), an uncoupler of oxidative phosphorylation (carbonyl cyanide m-chlorophenylhydrazone), a manganese superoxide dismutase mimetic, catalase, and a specific inhibitor of p38 MAPK (SB 203580), but were not affected by an inhibitor of the malate-aspartate shuttle (aminooxyacetate acid). Hydrogen peroxide (>/=10(-5) M) also stimulated p38 MAPK phosphorylation, ANG secretion, and ANG mRNA gene expression, but its stimulatory effect was blocked by catalase and SB 203580. These studies demonstrate that the stimulatory action of high glucose on ANG gene expression in IRPTCs is mediated at least in part via ROS generation and subsequent p38 MAPK activation. PMID- 12130565 TI - Eating elicited by orexin-a, but not melanin-concentrating hormone, is opioid mediated. AB - Melanin-concentrating hormone (MCH) and orexin-A are orexigenic peptidergic neurotransmitters produced primarily in the lateral hypothalamus. Because two other hypothalamic peptides, neuropeptide Y and agouti-related peptide, increase food intake by a mechanism that depends on activation of opioid receptors, we assessed whether MCH or orexin-A also elicits food intake via opioid receptor activation. A dose of naloxone (0.3 mg/kg, ip) that had no effect on its own reduced the acute orexigenic effect of third ventricular (i3vt) orexin-A (3 ng/rat). However, this same dose of naloxone had no effect on i3vt MCH (5 microg/rat)-induced hyperphagia. Because the opioid system has also been linked to food selection, we investigated whether MCH or orexin-A alters food choice when rats have simultaneous access to two diets differing in the relative amounts of fat and carbohydrate. Whereas i3vt MCH stimulated intake of both diets and did not alter food choice, i3vt orexin-A stimulated intake of only the high fat diet. These data indicate that despite several similarities between MCH and orexin-A, these two lateral hypothalamic area peptides stimulate food intake by recruiting different neural circuits and exert different effects on food choice. PMID- 12130564 TI - Acute signaling by the LH receptor is independent of protein kinase C activation. AB - LH receptor activation leads to the phosphorylation/activation of p42/44 MAPK in preovulatory granulosa cells. As the LH receptor can activate both adenylyl cyclase and phospholipase C, we hypothesized that the LH receptor could elicit phosphorylation of p42/44 MAPK through activation of protein kinase A (PKA) and/or protein kinase C (PKC). Preovulatory granulosa cells in serum-free primary cultures were treated with ovulatory concentrations of human chorionic gonadotropin (hCG), an LH receptor agonist, with or without various inhibitors. The PKA inhibitor H89 as well as the myristoylated PKA inhibitor peptide PKI strongly inhibited hCG-stimulated p42/44 MAPK phosphorylation, whereas the PKC inhibitor GF109203X had no effect on p42/44 MAPK phosphorylation. LH receptor stimulated phosphorylation of cAMP response element-binding protein (CREB), histone H3, and MAPK kinase (MEK) was also strongly inhibited by H89 and not by GF109203X. The extent of PKC activation was assessed in preovulatory granulosa cells using three criteria: translocation of PKC isoforms to the membrane fraction, phosphorylation of a known PKC substrate, and autophosphorylation of PKC delta on an activation-related site. By all three criteria PKCs were partially activated before hCG stimulation, and hCG treatment failed to elicit further PKC activation, in vitro or in vivo. Taken together, these results indicate that, under primary culture conditions where physiological levels of signaling proteins are present, hCG signals to activate MEK, p42/44 MAPK, CREB, and histone H3 in a predominantly PKA-dependent and PKC-independent manner. Unexpectedly, PKCs were partially activated in the absence of LH receptor activation, and LH receptor activation did not elicit further detectable PKC activation. PMID- 12130566 TI - Pituitary hypoplasia and lactotroph dysfunction in mice deficient for cyclin dependent kinase-4. AB - The lactotroph undergoes dynamic regulation of cell cycle progression during pregnancy, as well as throughout the development of the pituitary. We recently reported that female mice with targeted disruption of Cdk4, one of the G(1) regulatory cyclin-dependent kinases, are unable to support embryo implantation because of defective progesterone secretion from the corpus luteum. In this study, we demonstrate that this phenotype is not attributable to a primary defect in the corpus luteum but is a consequence of defective prolactin (PRL) production caused by inappropriate development of the pituitary lactotroph population. Specifically, the pituitary of Cdk4-deficient mice is extremely hypoplastic. Lactotrophs and somatotrophs of prepubertal Cdk4-deficient mice were 80% decreased in number, relative to those in wild-type mice, whereas gonadotrophs were unaffected. Lactotrophs of Cdk4-deficient mice did not proliferate in response to estrogen administration, whereas estrogen could induce the expression of galanin, an estrogen-responsive factor required for lactotroph proliferation. The reduction in lactotroph numbers was reflected by markedly diminished serum PRL levels in both prepubertal and postcoital Cdk4-deficient mice. Administration of PRL, after mating, significantly increased serum progesterone levels and restored implantation in Cdk4-deficient female mice. These observations demonstrate that Cdk4 is required for normal proliferation of the lactotroph population. PMID- 12130568 TI - Identification of intracellular signaling pathways that induce myotonic dystrophy protein kinase expression during myogenesis. AB - Myotonic dystrophy (DM) is the most common inherited adult neuromuscular disorder. DM is caused by a CTG expansion in the 3'-untranslated region of a protein kinase gene (DMPK). Decreased DMPK protein levels may contribute to the pathology of DM, as revealed by gene target studies. However, the postnatal regulation of DMPK expression and its pathophysiological role remain undefined. We studied the regulation of DMPK protein and mRNA expression during myogenesis in rat L6E9 myoblasts, mouse C2C12 myoblasts, and 10T1/2 fibroblasts stably expressing the myogenic transcription factor MyoD (10T1/2-MyoD). We detected DMPK as an 80-kDa protein mainly localized to the cytosolic fraction of skeletal muscle cells. DMPK expression and protein kinase activity were enhanced in IGF-II differentiated cells. In L6E9 and C2C12 cells, DMPK expression was regulated through the same signaling pathways (i.e. phosphatidylinositol 3-kinase, nuclear factor-kappaB, nitric oxide synthase, and p38 mitogen-activated protein kinase) that had been described as being crucial for the myogenesis induced by either low serum or IGF-II. However, in 10T1/2-MyoD cells, p38 MAPK inhibition blocked cell fusion and caveolin-3 expression without affecting DMPK up-regulation. These results suggest that although DMPK is induced during myogenesis, its expression cannot be totally associated with the development of a fully differentiated phenotype. PMID- 12130567 TI - Expression of the nuclear receptor coactivator, cAMP response element-binding protein, is sexually dimorphic and modulates sexual differentiation of neonatal rat brain. AB - Recent studies indicate that the transcriptional activity of steroid receptors is governed by proteins called nuclear receptor coactivators. Using immunocytochemistry, we found that on the day of birth (postnatal d 0) males express higher levels of the nuclear receptor coactivator, cAMP response element binding protein-binding protein (CBP), within the ventromedial hypothalamus, medial preoptic area, and arcuate nucleus. Using Western immunoblots, we confirmed that males have higher levels of CBP on postnatal d 0, 1, and 5; however, there was no sex difference on postnatal d 11. To examine the functional role of CBP, we infused oligodeoxynucleotides that were antisense to CBP mRNA or a scrambled sequence as a control into the hypothalamus of female rats on postnatal d 0, 1, and 2. On postnatal d 1, all rats were injected with 100 microg testosterone propionate to both masculinize (increase male) and defeminize (decrease female) sexual behavior. Rats were ovariectomized in adulthood and tested for adult sexual behavior. Neonatal CBP antisense oligodeoxynucleotides treatment interfered with the defeminizing, but not the masculinizing, actions of testosterone. These results indicate that CBP expression in developing rat brain is sexually dimorphic and an important modulator for steroid hormone action. PMID- 12130569 TI - Leptin-induced leptin resistance reveals separate roles for the anorexic and thermogenic responses in weight maintenance. AB - The purpose of this study was to determine whether leptin induces leptin resistance by examining the temporal attenuation of the anorexic and energy expenditure responses to leptin. We administered recombinant adeno-associated virus encoding rat leptin cDNA or control viral vector into mildly obese rats for 138 d and compared these results with those from pair-fed rats. We measured food consumption, body weight, oxygen consumption, leptin signal transduction, and brown adipose tissue uncoupling protein 1. The anorexic response attenuated by d 25, whereas the increase in energy expenditure persisted for 83 d before attenuating. Despite attenuation of physiological responses, phosphorylated signal transducer and activator of transcription-3 remained elevated for the duration of the study. The temporal differential attenuation of the anorexic and thermogenic responses allowed us to determine the relative contributions of each response to weight maintenance. The anorexic response predominantly mediated the initial loss of body weight, but only the energy expenditure response was necessary to maintain the reduced weight. This study provides evidence that leptin induces leptin resistance. The leptin resistance was associated with persistent elevation in hypothalamic phosphorylated signal transducer and activator of transcription-3 and was characterized by a rapid attenuation of the anorexic response and slower onset for the attenuation of the energy expenditure response. We propose that both elevated leptin and obesity may be necessary for the development of leptin resistance. PMID- 12130570 TI - Transcript expression of the tuberoinfundibular peptide (TIP)39/PTH2 receptor system and non-PTH1 receptor-mediated tonic effects of TIP39 and other PTH2 receptor ligands in renal vessels. AB - Although lower than in brain, the type 2 PTH receptor (PTH2-R) has been shown to be expressed throughout the cardiovascular system. Tuberoinfundibular peptide (TIP) purified from brain is thought to be the endogenous selective ligand of the PTH2-R. In the present studies, TIP and PTH2-R mRNA expressions were evidenced by RT-PCR in rat intrarenal arteries as well as in renovascular smooth muscle cells cultured from these arteries. In the isolated perfused rat kidney (IPK), peptides known to bind to both PTH1- and PTH2-Rs, such as rat PTH (1-34) and the hybrid PTH/PTHrP peptide, [Ile(5), Trp(23)]PTHrP (1-36), failed to exhibit improved vasodilatory effect, compared with human PTHrP (1-36), which binds only to the PTH1-R. Thus, a non-PTH1-R seemed not to be involved in the vasodilatory effects of these peptides. On the other hand, TIP exhibited complex vasoactivity, constricting the IPK at 10 nM and dilating the IPK at 1, 100, and 1000 nM. Moreover, [p-benzoyl-L-Phe(4),Ile(5),Trp(23)]PTHrP (1-36), initially described as a selective PTH2-R antagonist, also displayed a strong vasodilatory effect and therefore could not be used to check that TIP-induced vasoactivity was mediated by the PTH2-R. However, both [p-benzoyl-L-Phe(4),Ile(5),Trp(23)]PTHrP (1-36) and TIP displayed similar or even enhanced vasodilation in IPK in which PTH1-R induced vasodilation was fully desensitized by sustained exposure to human PTHrP (1-36). Importantly, in IPK desensitized to the vasodilatory action of PTHrP (1 36), the hybrid PTH/PTHrP peptide and rat PTH (1-34), whose vasodilatory responses appeared exclusively PTH1-R dependent in naive IPK, produced a new and strong vasodilation. In conclusion, TIP and PTH2-R mRNAs are expressed in renal vessels and TIP appears as a new vasoactive peptide. Whether TIP interacts with PTH2-R could not be shown. However, these studies reveal the ability of TIP, as well as of other peptides known to bind to the PTH2-R, to dilate renal vessels in a PTH1-R-independent manner. Moreover, results obtained in IPK desensitized to the vasodilatory action of PTHrP (1-36) strongly suggest that TIP, along with PTHrP, might be coordinately involved in the regulation of renal hemodynamics. PMID- 12130571 TI - Gestational and lactational exposure of male mice to diethylstilbestrol causes long-term effects on the testis, sperm fertilizing ability in vitro, and testicular gene expression. AB - The objective of the study was to determine the long-term effects of gestational and lactational exposure to diethylstilbestrol (DES; 0, 0.1, 1, and 10 microg/kg maternal body weight) on mouse testicular growth, epididymal sperm count, in vitro fertilizing ability, and testicular gene expression using cDNA microarrays and real-time PCR in mice on postnatal day (PND) 21, 105, and 315. In the high dose group there was a persistent decrease in the number of Sertoli cells, and sperm count was decreased on PND315 (P < 0.05). Sperm motion was unaffected; however, the in vitro fertilizing ability of epididymal sperm was decreased in the high dose group on both PND105 (P < 0.001) and PND315 (P < 0.05). Early and latent alterations in the expression of genes involved in estrogen signaling (estrogen receptor alpha), steroidogenesis (steroidogenic factor 1, 17alpha hydroxylase/C17,20-lyase, P450 side chain cleavage, steroidogenic acute regulatory protein, and scavenger receptor class B1), lysosomal function (LGP85 and prosaposin), and regulation of testicular development (testicular receptor 2, inhibin/activin beta C, and Hoxa10) were confirmed by real-time PCR. The results demonstrate that early exposure to DES causes long-term adverse effects on testicular development and sperm function, and these effects are associated with changes in testicular gene expression, even long after the cessation of DES exposure. PMID- 12130572 TI - Annexin 1-dependent actions of glucocorticoids in the anterior pituitary gland: roles of the N-terminal domain and protein kinase C. AB - Annexin 1 (ANXA1) is an important mediator of glucocorticoid action in the neuroendocrine system. As the activity of this protein in other systems is modulated by phosphorylation of its N-terminal domain, we have explored the significance of this domain and its phosphorylation status to ANXA1 actions within the pituitary gland, using an established in vitro preparation. Two N terminal peptides, ANXA1(Ac2-26) and ANXA1(Ac1-50), inhibited forskolin-evoked ACTH and prolactin release; however, they lacked the potency and full efficacy of the parent molecule (ANXA1(1-346)), whereas other shorter N-terminal sequences were without effect. A chimeric protein comprising ANXA1(1-44) and the C-terminal core of ANXA5 (ANXA5(20-320)) also produced a partial inhibition of peptide release. Protein kinase C (PKC) blockade (PKC(19-36)) abolished the inhibitory effects of dexamethasone on forskolin-evoked peptide release and attenuated the antisecretory actions of ANXA1(Ac2-26.) ANXA5, which sequesters PKC in other systems, produced similar effects. PKC(19-36) also blocked the dexamethasone- induced translocation of a serine phosphorylated species of ANXA1 from the cytoplasm to the outer cell surface. These results suggest that 1) the N-terminal domain plays a fundamental role in effecting the inhibitory actions of ANXA1 on pituitary peptide release; 2) PKC-dependent mechanisms are essential for both the cellular exportation and the biological activity of ANXA1; and 3) ANXA1 exported from the cells is serine phosphorylated. PMID- 12130573 TI - Identification and characterization of novel estrogen receptor-beta-sparing antiprogestins. AB - The steroid hormones estrogen and progesterone together regulate the development and maintenance of the female reproductive system. The actions of these two hormones are mediated by their respective nuclear receptors located within overlapping cell populations in target organs. The molecular mechanism of action of these two hormones has been defined to a large extent using estrogen receptor (ER) and progesterone receptor (PR) antagonists. In the case of ER, the available antagonists are highly receptor selective. With respect to PR, however, the available antiprogestins also interact with the receptors for glucocorticoids, mineralocorticoids, and androgens. Whereas these cross-reactivities can usually be managed in studies of female reproductive function, it is the recent demonstration that RU486 is an effective antagonist of the beta-isoform of ER that suggested the need for more selective antiprogestins. In this study, we used cell-based transcriptional assays combined with screens using coactivator peptide analogs to identify two novel classes of antiprogestins that distinguish themselves from the antiprogestin RU486 in the manner they interact with PR. One class exhibits the characteristics of a pure antiprogestin in that its members bind to the receptor and induce a conformational change that prevents the presentation of two potential coactivator binding surfaces on the protein. The second class of compounds distinguish themselves from RU486 in that they are ERbeta sparing. When tested in vivo the ER-sparing antiprogestins were as effective as RU486 in suppressing superovulation. It is anticipated that the availability of these new antiprogestins will advance the studies of PR pharmacology in a manner similar to how the availability of selective ER modulators has helped the study of ER action. PMID- 12130574 TI - Photoperiodic regulation of leptin resistance in the seasonally breeding Siberian hamster (Phodopus sungorus). AB - Seasonal Siberian hamsters lose fat reserves, decrease body weight and leptin concentrations, and suppress reproduction on short-day photoperiod (SD). Chronic leptin infusion at physiological doses caused body weight and fat loss in SD animals but was ineffective in long-day (LD) hamsters. Using ovariectomized estrogen-treated females, we tested the hypothesis that responsiveness to leptin is regulated by photoperiod. On SD, hypothalamic neuropeptide Y, agouti-related peptide, and cocaine- and amphetamine-regulated transcript gene expression in the arcuate nucleus did not exhibit significant changes, and despite SD-induced fat loss, the catabolic peptide proopiomelanocortin was down-regulated. Food restriction of LD-housed animals caused significant reduction of fat reserves and serum leptin concentrations to SD levels, suppressed serum gonadotropins, and induced increased anabolic (neuropeptide Y, agouti-related peptide) and decreased catabolic (proopiomelanocortin, cocaine- and amphetamine-regulated transcript) gene expression in the arcuate nucleus. Leptin infusion in food-restricted animals had no effect on fat reserves or gonadotropins and did not modulate neuropeptide gene expression. Also, leptin treatment did not blunt the refeeding responses or weight and fat gain in LD-housed food-restricted animals. In conclusion, our results strongly suggest that hypothalamic responses to leptin are regulated primarily by photoperiod, rather than seasonal changes in fat reserves, sex steroids, or leptin concentrations. PMID- 12130575 TI - Angiotensin II stimulates contraction and growth of testicular peritubular myoid cells in vitro. AB - Seminiferous tubule contraction, an important step in the regulation of spermatogenesis and testicular sperm output, is regulated by several agonists. In the present paper, we investigated whether angiotensin II (Ang II) may have a place among them. In binding experiments performed to assess the presence of specific receptors in rat peritubular myoid cells (TPMC), binding of (125)I-Ang II to TPMC was saturable in a time-dependent manner. Competition binding experiments performed with Losartan and PD 123319 showed that Losartan was able to inhibit the binding of (125)I-Ang II, whereas PD 123319 was ineffective. Ang II induced a dose-dependent rise in intracellular Ca(2+). Depletion of intracellular calcium stores by thapsygargin resulted in a lower rise of intracellular calcium, and the L-type voltage-operated calcium channel (VOCC-L) blocker verapamil abolished the Ca(2+) influx in rat TPMC. Altogether, these findings indicate that the Ang II-induced increase in [Ca(2+)](i) involves both extracellular influx and Ca(2+) release from intracellular stores. Ang II induced a dose-dependent TPMC contraction, and Losartan and not PD 123319 inhibited the response. Ang II-induced contraction was inhibited by adrenomedullin, previously shown to antagonize endothelin 1-provoked contraction in those cells. Ang II elicited (3)H-thymidine DNA incorporation and proliferation in a dose-dependent manner in TPMC. Losartan and both MAPK inhibitor PD 98059 and tyrosine kinase inhibitor AG18 were able to inhibit Ang II-induced (3)H-thymidine uptake and cell proliferation. In conclusion, the present study documents that angiotensin II, the active mediator of the tissue and circulating renin-angiotensin system present in the mammalian testis, induces contraction, growth and rise in intracellular calcium in rat peritubular myoid cells via angiotensin II type 1 receptors, and suggests that Ang II is involved in the paracrine regulation of the seminiferous tubule function. PMID- 12130576 TI - p38 MAPK-mediated signals are required for inducing osteoclast differentiation but not for osteoclast function. AB - Receptor activator of nuclear factor-kappaB ligand (RANKL)-induced signals play critical roles in osteoclast differentiation and function. SB203580, an inhibitor of p38 MAPK, blocked osteoclast formation induced by 1alpha,25-dihydroxyvitamin D(3) and prostaglandin E(2) in cocultures of mouse osteoblasts and bone marrow cells. Nevertheless, SB203580 showed no inhibitory effect on RANKL expression in osteoblasts treated with 1alpha,25-dihydroxyvitamin D(3) and prostaglandin E(2). RANKL-induced osteoclastogenesis in bone marrow cultures was inhibited by SB203580, suggesting a direct effect of SB203580 on osteoclast precursors, but not on osteoblasts, in osteoclast differentiation. However, SB203580 inhibited neither the survival nor dentine-resorption activity of osteoclasts induced by RANKL. Lipopolysaccharide (LPS), IL-1, and TNFalpha all stimulated the survival of osteoclasts, which was not inhibited by SB203580. Phosphorylation of p38 MAPK was induced by RANKL, IL-1, TNFalpha, and LPS in osteoclast precursors but not in osteoclasts. LPS stimulated phosphorylation of MAPK kinase 3/6 and ATF2, upstream and downstream signals of p38 MAPK, respectively, in osteoclast precursors but not in osteoclasts. Nevertheless, LPS induced degradation of IkappaB and phosphorylation of ERK in osteoclasts as well as in osteoclast precursors. These results suggest that osteoclast function is induced through a mechanism independent of p38 MAPK-mediated signaling. PMID- 12130577 TI - Human chorionic gonadotropin inhibits Kaposi's sarcoma associated angiogenesis, matrix metalloprotease activity, and tumor growth. AB - Kaposi's sarcoma is a highly angiogenic, AIDS-associated neoplasm that is more frequent in male than in female patients. Cases of spontaneous regression during pregnancy have been reported and the pregnancy hormone human chorionic gonadotropin (hCG) has shown anti-Kaposi's sarcoma activity in several, but not all, clinical trials. Antiproliferative and proapoptotic activities specific for Kaposi's sarcoma (KS) cells have been shown. We report here further analyses of the anti-KS activity of the hormone and show that urinary hCG, the hCG beta subunit, the hCG beta-core, and to a lesser extent a recombinant hCG, directly inhibit the activity of matrix metalloproteases of different origin. The hCG hormone also inhibited angiogenesis in vivo in the matrigel sponge assay as well as growth of KS cell xenografts in nude mice. The effect of the pure recombinant hormone dimer on xenograft growth was transient, indicating that the activity of intact hCG alone is not sufficient to overcome the growth potential of this tumor and suggesting that active hCG fragments or other anti-KS activities contribute to the activity of urinary hCG. PMID- 12130579 TI - Differential expression of GATA-4 and GATA-6 in fetal and adult mouse and human adrenal tissue. AB - Earlier work implicates transcription factors GATA-4 and GATA-6 in murine adrenal function. We have now studied their expression during mouse and human adrenal development in detail. GATA-4 and GATA-6 mRNAs and protein are readily detectable from embryonic d 14 and gestational wk 19 onwards in the mouse and human adrenal cortex, respectively. In the postnatal adrenal, GATA-4 expression is down regulated, whereas GATA-6 mRNA and protein continue to be expressed. To clarify the significance of GATA-4 for early adrenocortical development, Gata4-/- ES cells were injected into eight-cell-stage embryos derived from ROSA26 mice, a transgenic line expressing beta-galactosidase in all cell types, including the adrenocortical cells. The resultant chimeric embryos were stained with X-gal to discriminate ES cell- and host-derived tissue. Gata4-/- cells contributed to adrenocortical cells in these chimeras, and these cells also expressed GATA-6. Taken together, our findings suggest that GATA-6 expression is needed throughout adrenal development from fetal to adult age. GATA-4, on the other hand, may serve a role in the fetal adrenal gene regulation, although it is not essential for early adrenocortical differentiation. PMID- 12130578 TI - Steroidogenic factor-1 is essential for compensatory adrenal growth following unilateral adrenalectomy. AB - While the orphan nuclear receptor steroidogenic factor-1 (SF-1) has been shown to function as an induction factor to define adrenocortical cell lineage, it remains unclear whether SF-1 plays an additional role as a growth promoting agent in the adult adrenal cortex. The proliferative potential of the adrenal cortex in adult SF-1(+/-) mice was examined using the model of compensatory adrenal growth following unilateral adrenalectomy (uADX). While the right adrenal gland of wild type (wt) mice grew significantly after uADX, the adrenal of SF-1(+/-) mice exhibited a blunted, statistically nonsignificant weight increase. Accordingly, a profound increase in the proliferation marker proliferating cell nuclear antigen could be detected only in wt mice after uADX but not in the SF-1(+/-) mice. The proposed key regulator in adrenal compensatory growth, the recently cloned adrenal secretory serine protease was up-regulated in the remaining adrenal of wt mice, whereas this increase was blunted in SF-1(+/-) mice. While no differences in preadipocyte factor-1, the presumed marker of primitive adrenocortical cells, were detectable in the adrenals of wt and SF-1(+/-) mice, an increase in the ACTH receptor as well as agouti-related protein was observed only in wt animals but not in the SF-1(+/-) mice following uADX. Taken together, these results reflect a primary inability of adrenal cortical cells of SF-1(+/-) mice to undergo compensatory adrenal growth in response to uADX. PMID- 12130580 TI - Estrogen sulfotransferase: discrete and androgen-dependent expression in the male reproductive tract and demonstration of an in vivo function in the mouse epididymis. AB - Estrogen sulfotransferase (EST) catalyzes the sulfoconjugation and inactivation of the steroid hormone estrogen. It is known previously that EST is expressed abundantly in Leydig cells of the testis. We recently have shown that male mice with targeted EST gene disruption developed age related Leydig cell and seminiferous tubule abnormalities as a consequence of increased local estrogen stimulation. In the same study, we also found that epididymal sperm isolated from the mutant mice had significantly reduced motility, but whether this reflected impaired epididymal function or was secondary to the testicular lesions was not known. The purpose of the current study was to investigate if EST is normally present in the mouse epididymis and/or other parts of the male reproductive tract where, as in testis, it may play a role in regulating local estrogen homeostasis. We describe here that EST is expressed in the epithelium of corpus and cauda but not caput regions of the mouse epididymis. It is also expressed in the luminal epithelium and smooth muscle cells of the vas deferens but was present at very low levels, if at all, in the prostate or seminal vesicle/ coagulating gland. Hypophysectomy, castration, and epididymal ligation experiments, together with the use of an androgen receptor antagonist, established that EST expression in the epididymis and vas deferens is critically dependent on pituitary hormone(s) and androgen but not on other factors in the testicular fluid. Administration of exogenous estradiol to mice with surgically ligated epididymis resulted in a more pronounced reduction in sperm motility in EST mutant mice than in wild-type mice. We conclude that EST is discretely expressed and regulated in the male reproductive tract and plays a physiological role in maintaining the functional integrity of the epididymis by regulating luminal estrogen homeostasis. PMID- 12130581 TI - Insulinotropic hormone glucagon-like peptide-1 differentiation of human pancreatic islet-derived progenitor cells into insulin-producing cells. AB - Glucagon-like peptide-1 (GLP-1) is an intestinal incretin hormone, derived from the processing of proglucagon, that exerts insulinotropic actions on insulin producing pancreatic islet beta-cells. Recently GLP-1 was shown to stimulate the growth and differentiation (neogenesis) of beta-cells and appears to do so by inducing the expression of the homeodomain protein IDX-1 (islet duodenum homeobox 1; also known as PDX-1, pancreatic and duodenal homeobox gene; and as IPF-1, insulin promoter factor), which is required for pancreas development and the expression of beta-cell-specific genes. Earlier we identified multipotential progenitor cells in the islet and ducts of the pancreas, termed nestin-positive islet-derived progenitor cells (NIPs). Here we report the expression of functional GLP-1 receptors on NIPs and that GLP-1 stimulates the differentiation of NIPs into insulin-producing cells. Furthermore, confluent NIP cultures express the proglucagon gene and secrete GLP-1. These findings suggest a model of islet development in which pancreatic progenitor cells express both GLP-1 receptors and proglucagon with the formation of GLP-1. Locally produced GLP-1 may act as an autocrine/paracrine developmental morphogen on receptors on NIPs, resulting in the activation of IDX-1 and the expression of the proinsulin gene conferring a beta-cell phenotype. GLP-1 may be an important morphogen both for the embryonic development of the pancreas and for the neogenesis of beta-cells in the islets of the adult pancreas. PMID- 12130582 TI - Gastric parietal cells: potent endocrine role in secreting estrogen as a possible regulator of gastro-hepatic axis. AB - Estrogen, if it is produced in the gastrointestinal tract, may overflow into the systemic circulation in the case of increased portal-systemic shunting. This idea is in accord with a significant step-up in serum estradiol (E2) concentration in the portal vein of rats, compared with that in the artery. Gene expression of aromatase, estrogen synthetase, was demonstrated by RT-PCR in the gastric mucosa of male and female adult rats, equivalent to that in the ovary. Aromatase activity and production of E2 in the gastric mucosa were demonstrated by (3)H(2)O assay and gas chromatography-mass spectrometry, and they were inhibited by aromatase inhibitor, 4-hydroxyandrostenedione. Conversion of (14)C androstenedione to (14)C-E2 through (14)C-testosterone in cultured gastric mucosa was also demonstrated. Parietal cells exhibited strong signals for aromatase mRNA and immunoreactive protein by in situ hybridization histochemistry and immunohistochemistry. Estrogen receptor alpha mRNA and immunoreactive protein were demonstrated in hepatocytes by RT-PCR, in situ hybridization histochemistry, and immunohistochemistry. Total gastrectomy reduced portal venous E2 concentration, without changing systemic E2 concentration, together with down regulation of estrogen receptor alpha mRNA level in the liver. These findings indicate that gastric parietal cells play a potent endocrine role in secreting estrogen that may function as a regulator of the gastro-hepatic axis. PMID- 12130583 TI - Purified gonocytes from the neonatal rat form foci of proliferating germ cells in vitro. AB - Due to the lack of a specific marker for gonocytes from newborn rats, isolation of these cells has proven difficult and laborious. We have found a specific cell membrane marker, the Epithelial Cellular Adhesion Molecular (Ep-CAM) that can be used to isolate these cells using antibody directed cell sorting. 4 days post partum (dpp) rat testes were enzyme treated to attain a cell suspension, which was labelled with an antibody (GZ1) against Ep-CAM and tagged with a fluorescent probe. The labelled cell suspension was run through a FACS cell sorter, from which a gonocyte suspension of >85% purity was attained. The cells remained viable in culture and proliferated actively as determined by double labelling the cells with anti-HSP90alpha (a specific germ cell marker) and anti-BrdU antibodies (after BrdU incorporation). During culture, these cells formed chains of 2 to 4 cells and aggregates of proliferating germ cells were found after 8 days of culture. PMID- 12130584 TI - Estradiol stimulates GDNF expression in developing hypothalamic neurons. AB - Estrogens stimulate the differentiation of neurons and neural networks in the CNS. The concordance of the cellular responses of estrogens and growth factors suggests that both factors may interact on the cellular level to ensure their developmental role. We have put forward this hypothesis and analyzed the effect of estrogens on the expression of glial cell line-derived neutrotrophic factor (GDNF) in developing hypothalamic cells. Using Western blotting and competitive RTPCR, we have demonstrated that 17beta-estradiol (E2) increases the expression of GDNF in hypothalamic cell cultures. E2-induced GDNF expression was seen in neurons but not astrocytes. GDNF induction by E2 appeared to be transmitted through nonclassical estrogen action, since the application of the nuclear estrogen receptor antagonists ICI 182, 780 did not abolish this effect. Only inhibitors of intracellular Ca(2+) and cAMP/protein kinase A signaling were effective in preventing E2 effects. We conclude that E2 is capable of influencing GDNF expression in the developing hypothalamus. Thus, it is conceivable that developmental E2 effects in the hypothalamus are partially mediated through the regulation of other important developmental signals such as growth factors. PMID- 12130586 TI - Regulation of the forkhead transcription factor FKHR (FOXO1a) by glucose starvation and AICAR, an activator of AMP-activated protein kinase. AB - Expression of the catalytic subunit of glucose-6-phosphatase (G6Pase) has recently been shown to be transactivated by the transcription factor FKHR. Insulin and conditions of energy depletion are known repressors of the G6Pase gene. Whereas insulin is known to inhibit G6Pase expression by phosphorylation and nuclear exclusion of FKHR, the mechanism of repression of G6Pase by energy depletion is unknown. Here, we have studied the effect of glucose starvation and AICAR, an activator of AMP-activated protein kinase (AMPK) on G6Pase expression and the expressional level of FKHR-protein in hepatic cells. Using a H4-hepatoma cell line stably overexpressing FKHR, we found that both glucose starvation and treatment of cells with AICAR strongly repressed G6Pase expression and led to an almost complete disappearance of the FKHR protein, whereas the levels of control proteins and FKHR mRNA were not affected. Our data suggest that AICAR and glucose starvation inhibit G6Pase expression by a reduction of the cellular level of FKHR, presumably mediated by specific degradation of the protein. PMID- 12130585 TI - Mutant FGF-23 responsible for autosomal dominant hypophosphatemic rickets is resistant to proteolytic cleavage and causes hypophosphatemia in vivo. AB - FGF-23 is involved in the pathogenesis of two similar hypophosphatemic diseases, autosomal dominant hypophosphatemic rickets/osteomalacia (ADHR) and tumor-induced osteomalacia (TIO). We have shown that the overproduction of FGF-23 by tumors causes TIO. In contrast, ADHR derives from missense mutations in FGF-23 gene. However, it has been unclear how those mutations affect phosphate metabolism. Therefore, we produced mutant as well as wild-type FGF-23 proteins and examined their biological activity. Western blot analysis using site-specific antibodies showed that wild-type FGF-23 secreted into conditioned media was partially cleaved between Arg(179) and Ser(180). In addition, further processing of the cleaved N-terminal portion was observed. In constrast, mutant FGF-23 proteins found in ADHR were resistant to the cleavage. In order to clarify which molecule has the biological activity to induce hypophosphatemia, we separated full-length protein, the N-terminal and C-terminal fragments of wild-type FGF-23. When the activity of each fraction was examined in vivo, only the full-length FGF-23 decreased serum phosphate. Mutant FGF-23 protein that was resistant to the cleavage also retained the activity to induce hypophosphatemia. The extent of hypophosphatemia induced by the single administration of either wild-type or the mutant full-length FGF-23 protein was similar. In addition, implantation of CHO cells expressing the mutant FGF-23 protein caused hypophosphatemia and the decrease of bone mineral content. We conclude that ADHR is caused by hypophosphatemic action of mutant full-length FGF-23 proteins that are resistant to the cleavage between Arg(179) and Ser(180). PMID- 12130589 TI - Hormone replacement therapy. PMID- 12130590 TI - An ethically defensible market in organs. PMID- 12130591 TI - Magnetic resonance imaging of the knee. PMID- 12130592 TI - Chest pain units. PMID- 12130593 TI - Health risks in babies born after assisted reproduction. PMID- 12130594 TI - GPs vote "yes" for new contract framework. PMID- 12130595 TI - UK regulatory authority challenged over embryo screening. PMID- 12130596 TI - US issues tablets for potential terrorist attack. PMID- 12130598 TI - Experts launch action on acrylamide in staple foods. PMID- 12130600 TI - Global Fund director admits to $8bn shortfall. PMID- 12130597 TI - Planning for death but not serious future illness: qualitative study of housebound elderly patients. AB - OBJECTIVE: To understand how elderly patients think about and approach future illness and the end of life. DESIGN: Qualitative study conducted 1997-9. SETTING: Physician housecall programme affiliated to US university. PARTICIPANTS: 20 chronically ill housebound patients aged over 75 years who could participate in an interview. Participants identified through purposive and random sampling. MAIN OUTCOME MEASURES: In-depth semistructured interviews lasting one to two hours. RESULTS: Sixteen people said that they did not think about the future or did not in general plan for the future. Nineteen were particularly reluctant to think about, discuss, or plan for serious future illness. Instead they described a "one day at a time," "what is to be will be" approach to life, preferring to "cross that bridge" when they got to it. Participants considered end of life matters to be in the hands of God, though 13 participants had made wills and 19 had funeral plans. Although some had completed advance directives, these were not well understood and were intended for use only when death was near and certain. CONCLUSIONS: The elderly people interviewed for this study were resistant to planning in advance for the hypothetical future, particularly for serious illness when death is possible but not certain. PMID- 12130601 TI - South African government forced to give mothers antiretroviral drug. PMID- 12130603 TI - Candidate for US surgeon general comes under fire from senators. PMID- 12130604 TI - Surgeons must know their limitations, but so must governments. PMID- 12130607 TI - Are inequalities in height underestimated by adult social position? Effects of changing social structure and height selection in a cohort study. AB - OBJECTIVES: To investigate whether changing social structure and social mobility related to height generate (inflate) inequalities in height. DESIGN: Longitudinal 1958 British birth cohort study. SETTING: England, Scotland, and Wales. PARTICIPANTS: 10 176 people born 3-9 March 1958 for whom data were available at age 33 years. MAIN OUTCOME MEASURES: Adult height and social class at age 33 years; class of origin (father's occupation when participant was 7 years old). RESULTS: Adult height showed a social gradient with class at age 7 years and age 33 years. The difference in mean height between extreme groups was greater for class of origin than for adult class, reducing from 2.21 cm to 1.62 cm for men and from 2.18 cm to 1.74 cm for women. This narrowing inequality was due mainly to a decrease in mean height in classes I and II. This was because of the pattern of height related social mobility in which, for example, men moving into classes I and II were taller (mean 177.2 cm) than men remaining in class III manual (mean 176.1 cm) yet shorter than men with class I and II origins (mean 178.3 cm) and the relatively large number of individuals moving into classes I and II. Changes in the structure of society, seen here with the general trend of upward social mobility, have acted to diminish inequalities in adult height. CONCLUSIONS: The combination of changing social structure and height related mobility constrains, rather than inflates, inequalities in height and may lead to an underestimation of the role of childhood socioeconomic factors in the development of inequalities in adult disease. PMID- 12130606 TI - Randomised factorial trial of falls prevention among older people living in their own homes. AB - OBJECTIVE: To test the effectiveness of, and explore interactions between, three interventions to prevent falls among older people. DESIGN: A randomised controlled trial with a full factorial design. SETTING: Urban community in Melbourne, Australia. PARTICIPANTS: 1090 aged 70 years and over and living at home. Most were Australian born and rated their health as good to excellent; just over half lived alone. INTERVENTIONS: Three interventions (group based exercise, home hazard management, and vision improvement) delivered to eight groups defined by the presence or absence of each intervention. MAIN OUTCOME MEASURE: Time to first fall ascertained by an 18 month falls calendar and analysed with survival analysis techniques. Changes to targeted risk factors were assessed by using measures of quadriceps strength, balance, vision, and number of hazards in the home. RESULTS: The rate ratio for exercise was 0.82 (95% confidence interval 0.70 to 0.97, P=0.02), and a significant effect (P<0.05) was observed for the combinations of interventions that involved exercise. Balance measures improved significantly among the exercise group. Neither home hazard management nor treatment of poor vision showed a significant effect. The strongest effect was observed for all three interventions combined (rate ratio 0.67 (0.51 to 0.88, P=0.004)), producing an estimated 14.0% reduction in the annual fall rate. The number of people needed to be treated to prevent one fall a year ranged from 32 for home hazard management to 7 for all three interventions combined. CONCLUSIONS: Group based exercise was the most potent single intervention tested, and the reduction in falls among this group seems to have been associated with improved balance. Falls were further reduced by the addition of home hazard management or reduced vision management, or both of these. Cost effectiveness is yet to be examined. These findings are most applicable to Australian born adults aged 70-84 years living at home who rate their health as good. PMID- 12130608 TI - Impact of upward social mobility on population mortality: analysis with routine data. AB - OBJECTIVE: To examine the contribution of changes in the distribution of social class to the mortality of the whole population between 1970-2 and 1991-3. DESIGN: Examination of routine data at two time points: 1970-2 and 1991-3. DATA SOURCE: Data provided by the Office for National Statistics. MAIN OUTCOME MEASURES: Difference for the total population in the number of deaths between 1971 and 1991. Proportion of difference accounted for by change in population size, change in risk of death within each social class, or change in distribution of population across social classes. RESULTS: Reductions in mortality between 1970-2 and 1991-3 among men in England and Wales were partially (16% of all deaths) attributable to increases in the proportion of men in higher social classes, representing 3943 fewer deaths per year or one less death for every 3056 men in 1991-3 compared with 1970-2. CONCLUSION: Some of the observed reduction in mortality seen between 1970-2 and 1991-3 can be accounted for by improved overall socioeconomic status of the population. PMID- 12130609 TI - Haemoglobin and ferritin concentrations in men and women: cross sectional study. PMID- 12130610 TI - Bicyclist's vulva: observational study. PMID- 12130611 TI - Morale among general practitioners: qualitative study exploring relations between partnership arrangements, personal style, and workload. AB - OBJECTIVES: To explore general practitioners' experiences of wellbeing and distress at work, to identify their perceptions of the causes of and solutions to distress, and to draw out implications for improving morale in general practice. DESIGN: Three stage qualitative study consisting of one to one unstructured interviews, one to one guided interviews, and focus groups. SETTING: Fife, Lothian, and the Borders, South East Scotland. PARTICIPANTS: 63 general practitioner principals. RESULTS: Morale of general practitioners was explained by the complex interrelations between factors. Three key factors were identified: workload, personal style, and practice arrangements. Workload was commonly identified as a cause of low morale, but partnership arrangements were also a key mediating variable between increasing workload and external changes in general practice on the one hand and individual responses to these changes on the other. Integrated interventions at personal, partnership, and practice levels were seen to make considerable contributions to improving morale. Effective partnerships helped individuals to manage workload, but increasing workload was also seen to take away time and opportunities for practices to manage change and to build supportive and effective working environments. CONCLUSIONS: Solutions to the problem of low morale need integrated initiatives at individual, partnership, practice, and policy levels. Improving partnership arrangements is a key intervention, and rigorous action research is needed to evaluate different approaches. PMID- 12130612 TI - Blood transfusion medicine. PMID- 12130613 TI - Interpretation of rubella serology in pregnancy--pitfalls and problems. PMID- 12130614 TI - Depression in medical patients. PMID- 12130615 TI - Syphilis: old problem, new strategy. PMID- 12130617 TI - Are selective COX 2 inhibitors superior to traditional NSAIDs? Rofecoxib did not provide unequivocal benefit over traditional NSAIDs. PMID- 12130616 TI - Pregnancy complications and maternal cardiovascular risk: opportunities for intervention and screening? PMID- 12130618 TI - NHS Direct audited. NHS Direct is value for money and improving. PMID- 12130619 TI - Nurses as NHS gatekeepers. Nurses tend to use social rather than medical model of care. PMID- 12130620 TI - Can nurse practitioners provide equivalent care to GPs? Nurses and doctors working together can complement each other. PMID- 12130621 TI - Australian GPs are beginning battle faced by other countries. PMID- 12130622 TI - Oncologist who stood up to US insurance companies lost work. PMID- 12130624 TI - Improving working lives for doctors. PMID- 12130625 TI - Career pathways. PMID- 12130626 TI - Study leave funding in the devolved nations. PMID- 12130628 TI - The doctors' mess should be the hub of the hospital. PMID- 12130629 TI - Clinical record keeping. PMID- 12130630 TI - Mutant products of the NF2 tumor suppressor gene are degraded by the ubiquitin proteasome pathway. AB - Neurofibromatosis type 2 (NF2), a syndrome associated with multiple tumors of the nervous system, mostly schwannomas, is caused by mutations in the NF2 tumor suppressor gene that encodes schwannomin (Sch). Here we examined NF2 pathogenetic mutations that result in misfolding of the FERM domain. We found that these mutant forms of Sch were efficiently degraded by the ubiquitin-proteasome pathway. In transfected cells, Sch Delta F118 was 3-fold more efficiently degraded than the related molecule ezrin bearing the equivalent mutation. In heterozygous Nf2 knock-out mouse fibroblasts, endogenous mutant Sch Delta 81-121, but not wild type Sch, was also degraded by proteasomes. We further show that this degradation pathway is functional in primary Schwann cells. We analyzed Sch Delta 39-121 expressed in a transgenic mouse model of NF2 and found that Sch Delta 39-121, but not the endogenous wild type Sch, was unstable due to proteasome-mediated degradation. Altogether these results suggest that degradation of mutant Sch mediated by the ubiquitin-proteasome pathway is a physiopathological pathway contributing to the loss of Sch function in NF2 patients. PMID- 12130631 TI - The Drosophila melanogaster brainiac protein is a glycolipid-specific beta 1,3N acetylglucosaminyltransferase. AB - Mutations at the Drosophila melanogaster brainiac locus lead to defective formation of the follicular epithelium during oogenesis and to neural hyperplasia. The brainiac gene encodes a type II transmembrane protein structurally similar to mammalian beta1,3-glycosyltransferases. We have cloned the brainiac gene from D. melanogaster genomic DNA and expressed it as a FLAG tagged recombinant protein in Sf9 insect cells. Glycosyltransferase assays showed that brainiac is capable of transferring N-acetylglucosamine (GlcNAc) to beta linked mannose (Man), with a marked preference for the disaccharide Man(beta1,4)Glc, the core of arthro-series glycolipids. The activity of brainiac toward arthro-series glycolipids was confirmed by showing that the enzyme efficiently utilized glycolipids from insects as acceptors whereas it did not with glycolipids from mammalian cells. Methylation analysis of the brainiac reaction product revealed a beta1,3 linkage between GlcNAc and Man, proving that brainiac is a beta1,3GlcNAc-transferase. Human beta1,3GlcNAc-transferases structurally related to brainiac were unable to transfer GlcNAc to Man(beta1,4)Glc-based acceptor substrates and failed to rescue a homozygous lethal brainiac allele, indicating that these proteins are paralogous and not orthologous to brainiac. PMID- 12130632 TI - Protein kinase C-zeta regulates transcription of the matrix metalloproteinase-9 gene induced by IL-1 and TNF-alpha in glioma cells via NF-kappa B. AB - The regulation of matrix metalloproteinase-9 (MMP-9) expression in glioma cells is one of the key processes in tumor invasion through the brain extracellular matrix. Although some studies have demonstrated the implication of classic protein kinase C (PKC) isoforms in the regulation of MMP-9 production by phorbol esters or lipopolysaccharide, the involvement of specific PKC isoforms in the signaling pathways leading to MMP-9 expression by inflammatory cytokines remains unclear. Here we report that the atypical PKC-zeta isoform participates in the induction of MMP-9 expression by interleukin-1 (IL-1) and tumor necrosis factor alpha (TNF-alpha) in rat C6 glioma cells. Indeed, zymography and semi quantitative reverse transcriptase-PCR analysis showed that pretreatment of C6 cells with PKC-zeta pseudosubstrate abolished MMP-9 activity and gene expression induced by IL-1 or TNF-alpha. Accordingly, IL-1 and TNF-alpha were able to induce PKC-zeta activity, as demonstrated by in vitro kinase assay using immunoprecipitated PKC-zeta. Furthermore, stable C6 clones overexpressing PKC zeta, but not PKC-epsilon, displayed an up-regulation of MMP-9 constitutive expression as well as an increase of mmp-9 promoter activity. These processes were inhibited by an NF-kappaB-blocking peptide and completely prevented by NF kappaB-binding site mutation in the mmp-9 promoter. Taken together, these results indicate that PKC-zeta plays a key role in the regulation of MMP-9 expression in C6 glioma cells through NF-kappaB. PMID- 12130633 TI - Identification of biologically active sequences in the laminin alpha 4 chain G domain. AB - Laminins are a family of trimeric extracellular matrix proteins consisting of alpha, beta, and gamma chains. So far five different laminin alpha chains have been identified. The laminin alpha 4 chain, which is present in laminin-8/9, is expressed in cells of mesenchymal origin, such as endothelial cells and adipocytes. Previously, we identified heparin-binding sites in the C-terminal globular domain (G domain) of the laminin alpha 4 chain. Here we have focused on the biological functions of the laminin alpha 4 chain G domain and screened active sites using a recombinant protein and synthetic peptides. The rec-alpha 4G protein, comprising the entire G domain, promoted cell attachment activity. The cell attachment activity of rec-alpha 4G was completely blocked by heparin and partially inhibited by EDTA. We synthesized 116 overlapping peptides covering the entire G domain and tested their cell attachment activity. Twenty peptides showed cell attachment activity, and 16 bound to heparin. We further tested the effect of the 20 active peptides in competition assays for cell attachment and heparin binding to rec-alpha 4G protein. A4G6 (LAIKNDNLVYVY), A4G20 (DVISLYNFKHIY), A4G82 (TLFLAHGRLVFM), and A4G83 (LVFMFNVGHKKL), which promoted cell attachment and heparin binding, significantly inhibited both cell attachment and heparin binding to rec-alpha 4G. These results suggest that the four active sites are involved in the biological functions of the laminin alpha 4 chain G domain. Furthermore, rec alpha 4G, A4G6, and A4G20 were found to interact with syndecan-4. These active peptides may be useful for defining of the molecular mechanism laminin-receptor interactions and laminin-mediated cellular signaling pathways. PMID- 12130634 TI - Defining requirements for heterodimerization between the retinoid X receptor and the orphan nuclear receptor Nurr1. AB - Nurr1, an orphan nuclear receptor mainly expressed in the central nervous system, is essential for the development of the midbrain dopaminergic neurons. Nurr1 binds DNA as a monomer and exhibits constitutive transcriptional activity. Nurr1 can also regulate transcription as a heterodimer with the retinoid X receptor (RXR) and activate transcription in response to RXR ligands. However, the specific physiological roles of Nurr1 monomers and RXR-Nurr1 heterodimers remain to be elucidated. The aim of this study was to define structural requirements for RXR-Nurr1 heterodimerization. Several amino acid substitutions were introduced in both Nurr1 and RXR in the I-box, a region previously shown to be important for nuclear receptor dimerization. Single amino acid substitutions introduced in either Nurr1 or RXR abolished heterodimerization. Importantly, heterodimerization deficient Nurr1 mutants exhibited normal activities as monomers. Thus, by introducing specific amino acid substitutions in Nurr1, monomeric and heterodimeric properties of Nurr1 can be distinguished. Interestingly, substitutions in the RXR I-box differentially affected heterodimerization with Nurr1, retinoic acid receptor, thyroid hormone receptor, and constitutive androstane receptor demonstrating that the dimerization interfaces in these different heterodimers are functionally unique. Furthermore, heterodimerization between RXR and Nurr1 had a profound influence on the constitutive activity of Nurr1, which was diminished as a result of RXR interaction. In conclusion, our data show unique structural and functional properties of RXR-Nurr1 heterodimers and also demonstrate that specific mutations in Nurr1 can abolish heterodimerization without affecting other essential functions. PMID- 12130635 TI - Brain-derived neurotrophic factor regulates surface expression of alpha-amino-3 hydroxy-5-methyl-4-isoxazoleproprionic acid receptors by enhancing the N ethylmaleimide-sensitive factor/GluR2 interaction in developing neocortical neurons. AB - In hippocampal neurons, the exocytotic process of alpha-amino-3-hydroxy-5-methyl 4-isoxazoleproprionic acid (AMPA)-type glutamate receptors is known to depend on activation of N-methyl-d-aspartate channels and its resultant Ca(2+) influx from extracellular spaces. Here we found that brain-derived neurotrophic factor (BDNF) induced a rapid surface translocation of AMPA receptors in an activity independent manner in developing neocortical neurons. The receptor translocation became evident within hours as monitored by [(3)H]AMPA binding and was resistant against ionotropic glutamate receptor antagonists as evidenced with surface biotinylation assay. This process required intracellular Ca(2+) and was inhibited by the blockers of conventional exocytosis, brefeldin A, botulinum toxin B, and N ethylmaleimide. To explore the translocation mechanism of individual AMPA receptor subunits, we utilized the human embryonic kidney (HEK) 293 cells carrying the BDNF receptor TrkB. After the single transfection of GluR2 cDNA or GluR1 cDNA into HEK/TrkB cells, BDNF triggered the translocation of GluR2 but not that of GluR1. Subsequent mutation analysis of GluR2 carboxyl-terminal region indicated that the translocation of GluR2 subunit in HEK293 cells involved its N ethylmaleimide-sensitive factor-binding domain but not its PDZ-interacting site. Following co-transfection of GluR1 and GluR2 cDNAs, solid phase cell sorting revealed that GluR1 subunits were also able to translocate to the cell surface in response to BDNF. An immunoprecipitation assay confirmed that BDNF stimulation can enhance the interaction of GluR2 with N-ethylmaleimide-sensitive factor. These results reveal a novel role of BDNF in regulating the surface expression of AMPA receptors through a GluR2-NSF interaction. PMID- 12130636 TI - Changes in the glycosylation pattern of prion protein in murine scrapie. Implications for the mechanism of neurodegeneration in prion diseases. AB - In prion diseases, the normal prion protein (PrP(c)) undergoes a conformational change that results in the abnormal form, named scrapie prion protein (PrP(sc)). The visual system of rodents provides a relatively simple neuronal model in which the cell bodies of neurons are confined to the retina and the axons constitute the optic nerve. We investigated by Western blot the profile of PrP(c) in the optic nerve and retina of normal hamsters and mice. We found that in the optic nerve the amount of PrP(c) is significantly higher than in the retina. A less abundant non-glycosylated band was observed in retinas compared with the optic nerve and brain. Similar results were found in the gray and white matter from normal mice and hamsters. After stereotaxic injection of ME7 or 139A in the superior colliculus, a visual target area, the proportion and glycopattern of PrP changed in the retina and optic nerve throughout the course of the disease. Similar results were found in the gray and white matter at terminal stage of scrapie after injection of ME7 and 139A in the dorsal hippocampus. This is the first time that changes in the distribution and glycopattern of PrP have been described in an in vivo model of prion diseases. PMID- 12130637 TI - Characterization of SMOC-1, a novel modular calcium-binding protein in basement membranes. AB - We have isolated the novel gene SMOC-1 that encodes a secreted modular protein containing an EF-hand calcium-binding domain homologous to that in BM-40. It further consists of two thyroglobulin-like domains, a follistatin-like domain and a novel domain. Recombinant expression in human cells showed that SMOC-1 is a glycoprotein with a calcium-dependent conformation. Results from Northern blots, reverse transcriptase-PCR, and immunoblots revealed a widespread expression in many tissues. Immunofluorescence studies with an antiserum directed against recombinant human SMOC-1 demonstrated a basement membrane localization of the protein and additionally its presence in other extracellular matrices. Immunogold electron microscopy confirmed the localization of SMOC-1 within basement membranes in kidney and skeletal muscle as well as its expression in the zona pellucida surrounding the oocyte. PMID- 12130638 TI - Circadian Transcription. Thinking outside the E-Box. AB - The E-Box is a widely used DNA control element. Despite its brevity and broad distribution the E-Box is a remarkably versatile sequence that affects many different genetic programs, including proliferation, differentiation, tissue specific responses, and cell death. The circadian clock is one of the latest pathways shown to employ this element. In this context, E-Boxes are likely to play a key role in establishing the robust waves of gene expression characteristic of circadian transcription. The regulatory flexibility of the E Box hinges on the sequence ambiguity allowed at its core, the strong influence of the surrounding sequences, and the recruitment of spatially and temporally regulated E-Box-binding factors. Therefore, understanding how a particular E-Box can accomplish a specific task entails the identification and systematic analysis of these cis- and trans-acting E-Box modifiers. In the present study we compared the E-Box-containing minimal promoters of vasopressin and cyclin B1, two genes that can respond to the transcriptional oscillators driving the circadian clock and cell cycle, respectively. Results of this comparison will help elucidate the manner in which discreet DNA modules associate to either augment or restrain the activation of potential circadian E-Boxes in response to competing regulatory signals. PMID- 12130639 TI - Dissecting the hydrolytic activities of sarcoplasmic reticulum ATPase in the presence of acetyl phosphate. AB - Sarcoplasmic reticulum vesicles and purified Ca(2+)-ATPase hydrolyze acetyl phosphate both in the presence and absence of Ca(2+). The Ca(2+)-independent activity was fully sensitive to vanadate, insensitive to thapsigargin, and proceeded without accumulation of phosphorylated enzyme. Acetyl phosphate hydrolysis in the absence of Ca(2+) was activated by dimethyl sulfoxide. The Ca(2+)-dependent activity was partially sensitive to vanadate, fully sensitive to thapsigargin, and associated with steady phosphoenzyme accumulation. The Ca(2+)/P(i) coupling ratio at neutral pH sustained by 10 mm acetyl phosphate was 0.57. Addition of 30% dimethyl sulfoxide completely blocked Ca(2+) transport and partially inhibited the hydrolysis rate. Uncoupling induced by dimethyl sulfoxide included the accumulation of vanadate-insensitive phosphorylated enzyme. When acetyl phosphate was the substrate, the hydrolytic pathway was dependent on experimental conditions that might or might not allow net Ca(2+) transport. The interdependence of both Ca(2+)-dependent and Ca(2+)-independent hydrolytic activities was demonstrated. PMID- 12130640 TI - Global gene expression profiling in Escherichia coli K12. The effects of leucine responsive regulatory protein. AB - Leucine-responsive regulatory protein (Lrp) is a global regulatory protein that affects the expression of multiple genes and operons in bacteria. Although the physiological purpose of Lrp-mediated gene regulation remains unclear, it has been suggested that it functions to coordinate cellular metabolism with the nutritional state of the environment. The results of gene expression profiles between otherwise isogenic lrp(+) and lrp(-) strains of Escherichia coli support this suggestion. The newly discovered Lrp-regulated genes reported here are involved either in small molecule or macromolecule synthesis or degradation, or in small molecule transport and environmental stress responses. Although many of these regulatory effects are direct, others are indirect consequences of Lrp mediated changes in the expression levels of other global regulatory proteins. Because computational methods to analyze and interpret high dimensional DNA microarray data are still an early stage, much of the emphasis of this work is directed toward the development of methods to identify differentially expressed genes with a high level of confidence. In particular, we describe a Bayesian statistical framework for a posterior estimate of the standard deviation of gene measurements based on a limited number of replications. We also describe an algorithm to compute a posterior estimate of differential expression for each gene based on the experiment-wide global false positive and false negative level for a DNA microarray data set. This allows the experimenter to compute posterior probabilities of differential expression for each individual differential gene expression measurement. PMID- 12130641 TI - NMR structure of PW2 bound to SDS micelles. A tryptophan-rich anticoccidial peptide selected from phage display libraries. AB - PW2 (HPLKQYWWRPSI) was selected from phage display libraries through an alternative panning method using living sporozoites of Eimeria acervulina as target. Synthetic PW2 shows anticoccidial activity against E. acervulina and Eimeria tenella with very low hemolytic activity. It also displays antifungal activity but no activity against bacteria. We present the solution structure of the PW2 bound to SDS micelles. In the absence of an interface, PW2 is in random coil conformation. In micelles, structural calculation shows that Trp-7 forms the hydrophobic core that is important for the peptide folding. Lys-4, Tyr-6, Trp-8, and Arg-9 are in the same surface, possibly facing the micelle interface. This possibility was supported by the fact that chemical shift differences for these residues were more pronounced when compared with PW2 in water and in SDS. PW2 gains structure upon binding to SDS micelles. Lys-4, Tyr-6, Trp-8, and Arg-9 were found to bind to the micelle. Trp-7, Trp-8, and Arg-9 composed the WW+ consensus found in the sequence of the peptides selected with the phage display technique against E. acervulina sporozoites. This suggested that Trp-7, Trp-8, and Arg-9 are probably key residues not only for the peptide interaction with SDS micelles but also for the interaction with E. acervulina sporozoites surface. PMID- 12130642 TI - Ophioluxin, a convulxin-like C-type lectin from Ophiophagus hannah (King cobra) is a powerful platelet activator via glycoprotein VI. AB - Ophioluxin, a potent platelet agonist, was purified from the venom of Ophiophagus hannah (King cobra). Under nonreducing conditions it has a mass of 85 kDa, similar to convulxin, and on reduction gives two subunits with masses of 16 and 17 kDa, slightly larger than those of convulxin. The N-terminal sequences of both subunits are very similar to those of convulxin and other C-type lectins. Ophioluxin induces a pattern of tyrosine-phosphorylated proteins in platelets like that caused by convulxin, when using appropriate concentrations based on aggregation response, because it is about 2-4 times more powerful as agonist than the latter. Ophioluxin and convulxin induce [Ca(2+)](i) elevation both in platelets and in Dami megakaryocytic cells, and each of these C-type lectins desensitizes responses to the other. Convulxin agglutinates fixed platelets at 2 microg/ml, whereas ophioluxin does not, even at 80 microg/ml. Ophioluxin resembles convulxin more than echicetin or alboaggregin B because polyclonal anti ophioluxin antibodies recognize both ophioluxin and convulxin, but not echicetin, and platelets adhere to and spread on ophioluxin- or convulxin-precoated surfaces in the same way that is clearly different from their behavior on an alboaggregin B surface. Immobilized ophioluxin was used to isolate the glycoprotein VI-Fcgamma complex from resting platelets, which also contained Fyn, Lyn, Syk, LAT, and SLP76. Ophioluxin is the first multiheterodimeric, convulxin-like snake C-type lectin, as well as the first platelet agonist, to be described from the Elapidae snake family. PMID- 12130643 TI - Complex N-linked glycosylated nicastrin associates with active gamma-secretase and undergoes tight cellular regulation. AB - The intramembranous proteolysis of Notch and the amyloid precursor protein by gamma-secretase exemplifies an unusual and newly recognized mechanism of signal transduction in multicellular organisms. Here, we show that only a form of nicastrin (NCT) containing N-linked complex oligosaccharides is present in active gamma-secretase complexes. Overexpression of NCT does not generate more of this mature protein, a phenomenon analogous to the strictly regulated formation of mature presenilin heterodimers from immature holoprotein. The absence of presenilin severely limits the maturation of NCT, yet combined overexpression of both proteins does not increase respective mature types. Taken together, our findings describe unusual regulatory features of this key signaling protease: the association of NCT with gamma-secretase is tightly regulated via glycosylation; at least one other cofactor exists; the least abundant member of the complex becomes limiting; and the cofactor that serves this role may vary by cell type. PMID- 12130644 TI - Peptides containing membrane-transiting motifs inhibit virus entry. AB - Several exceptional peptides have been identified that can cross plasma membranes and deliver various covalently linked moieties into cells. We report the surprising observation that each of four structurally distinct transiting peptides tested displayed antiviral activity and inhibited herpes simplex virus entry into cells. All four peptides inhibited infection at concentrations in the low micromolar range. Some of the peptides selectively and reversibly blocked entry without inactivating virions in a persistent manner. For other peptides, the effects on virus entry were not readily distinguishable from virus inactivation. High concentrations of nearly all peptides lead to irreversible inactivation of virions. By various criteria, the peptides differed in their ability to inactivate virions and in the temperature dependence of inactivation. Testing of peptides with modifications known to disrupt transport revealed that, in some instances, transport activity did not correlate with antiviral activity. These results identify inhibition of viral entry as another common property of membrane-transiting peptides in addition to their ability to cross membranes and transport materials into cells. These or related peptides may be useful as agents to prevent infection and to study the process of viral entry. PMID- 12130645 TI - Structure and flexibility of Streptococcus agalactiae hyaluronate lyase complex with its substrate. Insights into the mechanism of processive degradation of hyaluronan. AB - Streptococcus agalactiae hyaluronate lyase degrades primarily hyaluronan, the main polysaccharide component of the host connective tissues, into unsaturated disaccharide units as the end product. Such function of the enzyme destroys the normal connective tissue structure of the host and exposes the tissue cells to various bacterial toxins. The crystal structure of hexasaccharide hyaluronan complex with the S. agalactiae hyaluronate lyase was determined at 2.2 A resolution; the mechanism of the catalytic process, including the identification of specific residues involved in the degradation of hyaluronan, was clearly identified. The enzyme is composed structurally and functionally from two distinct domains, an alpha-helical alpha-domain and a beta-sheet beta-domain. The flexibility of the protein was investigated by comparing the crystal structures of the S. agalactiae and the Streptococcus pneumoniae enzymes, and by using essential dynamics analyses of CONCOORD computer simulations. These revealed important modes of flexibility, which could be related to the protein function. First, a rotation/twist of the alpha-domain relative to the beta-domain is potentially related to the mechanism of processivity of the enzyme; this twist motion likely facilitates shifting of the ligand along the catalytic site cleft in order to reposition it to be ready for further cleavage. Second, a movement of the alpha- and beta-domains with respect to each other was found to contribute to a change in electrostatic characteristics of the enzyme and appears to facilitate binding of the negatively charged hyaluronan ligand. Third, an opening/closing of the substrate binding cleft brings a catalytic histidine closer to the cleavable substrate beta1,4-glycosidic bond. This opening/closing mode also reflects the main conformational difference between the crystal structures of the S. agalactiae and the S. pneumoniae hyaluronate lyases. PMID- 12130646 TI - Identification of neuropeptide W as the endogenous ligand for orphan G-protein coupled receptors GPR7 and GPR8. AB - The structurally related orphan G-protein-coupled receptors GPR7 and GPR8 are expressed in the central nervous system, and their ligands have not been identified. Here, we report the identification of the endogenous ligand for both of these receptors. We purified the peptide ligand from porcine hypothalamus using stable Chinese hamster ovary cell lines expressing human GPR8 and cloned the cDNA encoding its precursor protein. The cDNA encodes two forms of the peptide ligand with lengths of 23 and 30 amino acid residues as mature peptides. We designated the two ligands neuropeptide W-23 (NPW23) and neuropeptide W-30 (NPW30). The amino acid sequence of NPW23 is completely identical to that of the N-terminal 23 residues of NPW30. Synthetic NPW23 and NPW30 activated and bound to both GPR7 and GPR8 at similar effective doses. Intracerebroventricular administration of NPW23 in rats increased food intake and stimulated prolactin release. These findings indicate that neuropeptide W is the endogenous ligand for both GPR7 and GPR8 and acts as a mediator of the central control of feeding and the neuroendocrine system. PMID- 12130647 TI - Engineering of metallothionein-3 neuroinhibitory activity into the inactive isoform metallothionein-1. AB - The third isoform of mammalian metallothioneins (MT-3), mainly expressed in brain and down-regulated in Alzheimer's disease, exhibits neuroinhibitory activity in vitro and a highly flexible structure that distinguishes it from the widely expressed MT-1/-2 isoforms. Previously, we showed that two conserved prolyl residues of MT-3 are crucial for both the bioactivity and cluster dynamics of this isoform. We have now used genetic engineering to introduce these residues into mouse MT-1. The S6P,S8P MT-1 mutant is inactive in neuronal survival assays. However, the additional introduction of the unique Thr5 insert of MT-3 resulted in a bioactive MT-1 form. Temperature-dependent and saturation transfer (113)Cd NMR experiments performed on the (113)Cd-reconstituted wild-type and mutant Cd(7) MT-1 forms revealed that the gain of MT-3-like neuronal inhibitory activity is paralleled by an increase in conformational flexibility and intersite metal exchange in the N-terminal Cd(3)-thiolate cluster. The observed correlation suggests that structure/cluster dynamics are critical for the biological activity of MT-3. We propose that the interplay between the specific Pro-induced conformational requirements and those of the metal-thiolate bonds gives rise to an alternate and highly fluctuating cluster ensemble kinetically trapped by the presence of the (5)TCPCP(9) motif. The functional significance of such heterogeneous cluster ensemble is discussed. PMID- 12130648 TI - Polarization of myosin II heavy chain-protein kinase C in chemotaxing dictyostelium cells. AB - Eukaryotic cells need morphological polarity to carry out chemotaxis (Parent, C. A., Blacklock, B. J., Froehlich, W. M., Murphy, D. B., and Devreotes, P. N. (1998) Cell 95, 81-91; Jin, T., Zhang, N., Long, Y., Parent, C., and Devreotes, P. N. (2000) Science 287, 1034-1036; Servant, G., Weiner, O. D., Herzmark, P., Balla, T., Sedat, J. W., and Bourne, H. R. (2000) Science 287, 1037-1040), but sensing direction does not require polarization of chemoattractant receptors. When cells are exposed to a gradient of chemoattractant, activation occurs selectively at the stimulated edge. Such localized activation, transmitted by the recruitment of cytosolic proteins, may be a general mechanism for gradient sensing by G protein-linked chemotactic systems. Here we show that in Dictyostelium discoideum cells exposed to a cAMP gradient the myosin II heavy chain kinase (MHC-PKC) and myosin II translocate to opposite ends of the cell. We further show that MHC-PKC C1 domain is responsible for the localization of MHC PKC to the cell leading edge, but it is not sufficient to promote cell polarization. Our findings suggest a mechanism by which MHC-PKC regulates myosin II, allowing cell polarization and movement in the direction of the cAMP source. PMID- 12130649 TI - Activations of ERK1/2 and JNK by transforming growth factor beta negatively regulate Smad3-induced alkaline phosphatase activity and mineralization in mouse osteoblastic cells. AB - Transforming growth factor (TGF) beta inhibits alkaline phosphatase (ALP) activity and mineralization in mouse osteoblastic MC3T3-E1 cells, whereas local administration of TGF-beta stimulates bone formation in vivo. We recently demonstrated that Smad3, a TGF-beta signaling molecule, promotes ALP activity and mineralization in MC3T3-E1 cells. Moreover, the target disruption of Smad3 in mouse is reported to cause a decrease in bone mineral density. These findings indicate that Smad3 plays an important role in the regulation of bone formation. However, why the effects of TGF-beta and Smad3 on ALP activity and mineralization are different remains unknown. The purpose of the present study is to clarify the role of mitogen-activated protein kinase (MAPK) in TGF-beta and Smad3 pathways in osteoblast. TGF-beta activated extracellular signal-regulated kinases/p42/p44 (ERK1/2), p38 MAPK, and c-Jun N-terminal kinase (JNK) in mouse osteoblastic MC3T3 E1 cells. The expression of dominant negative type Smad3, Smad3DeltaC, affected neither TGF-beta-activated MAPKs nor TGF-beta-inhibited ALP activity. Specific inhibitors of ERK1/2 activation (PD98059 and U0126), as well as JNK inhibitors (curcumin and dicumarol) antagonized the inhibitory effects of TGF-beta on ALP activity and mineralization, whereas the specific inhibitor of p38 MAPK (SB203580) did not affect them. PD98059 and curcumin enhanced Smad3-induced ALP activity and mineralization, whereas SB203580 inhibited them. In the luciferase reporter assay using 3TP-lux with the specific Smad3-responsive element, PD98059, and curcumin enhanced TGF-beta- and Smad3-induced transcriptional activity in MC3T3-E1 cells. On the other hand, TGF-beta-induced production of type I collagen was antagonized by curcumin but not by PD98059. The present study indicated that TGF-beta-responsive ERK1/2 and JNK cascades negatively regulate Smad3-induced transcriptional activity as well as ALP activity and mineralization in osteoblasts. PMID- 12130650 TI - Role of amino acid residues in transmembrane segments IS6 and IIS6 of the Na+ channel alpha subunit in voltage-dependent gating and drug block. AB - Alanine-scanning mutagenesis of transmembrane segments IS6 and IIS6 of the rat brain Na(v)1.2 channel alpha subunit identified mutations N418A in IS6 and L975A in IIS6 as causing strong positive shifts in the voltage dependence of activation. In contrast, mutations V424A in IS6 and L983A in IIS6 caused strong negative shifts. Most IS6 mutations opposed inactivation from closed states, but most IIS6 mutations favored such inactivation. Mutations L421C and L983A near the intracellular ends of IS6 and IIS6, respectively, exhibited significant sustained Na(+) currents at the end of 30-ms depolarizations, indicating a role for these residues in Na(+) channel fast inactivation. These residues, in combination with residues at the intracellular end of IVS6, are well situated to form an inactivation gate receptor. Mutation I409A in IS6 reduced the affinity of the local anesthetic etidocaine for the inactivated state by 6-fold, and mutations I409A and N418A reduced use-dependent block by etidocaine. No IS6 or IIS6 mutations studied affected inactivated-state affinity or use-dependent block by the neuroprotective drug sipatrigine (compound 619C89). These results suggest that the local anesthetic receptor site is formed primarily by residues in segments IIIS6 and IVS6 with the contribution of a single amino acid in segment IS6. PMID- 12130651 TI - The Drosophila gene brainiac encodes a glycosyltransferase putatively involved in glycosphingolipid synthesis. AB - The Drosophila genes fringe and brainiac exhibit sequence similarities to glycosyltransferases. Drosophila and mammalian fringe homologs encode UDP-N acetylglucosamine:fucose-O-Ser beta1,3-N-acetylglucosaminyltransferases that modulate the function of Notch family receptors. The biological function of brainiac is less well understood. brainiac is a member of a large homologous mammalian beta3-glycosyltransferase family with diverse functions. Eleven distinct mammalian homologs have been demonstrated to encode functional enzymes forming beta1-3 glycosidic linkages with different UDP donor sugars and acceptor sugars. The putative mammalian homologs with highest sequence similarity to brainiac encode UDP-N-acetylglucosamine:beta1,3-N-acetylglucosaminyltransferases (beta3GlcNAc-transferases), and in the present study we show that brainiac also encodes a beta3GlcNAc-transferase that uses beta-linked mannose as well as beta linked galactose as acceptor sugars. The inner disaccharide core structures of glycosphingolipids in mammals (Galbeta1-4Glcbeta1-Cer) and insects (Manbeta1 4Glcbeta1-Cer) are different. Both disaccharide glycolipids served as substrates for brainiac, but glycolipids of insect cells have so far only been found to be based on the GlcNAcbeta1-3Manbeta1-4Glcbeta1-Cer core structure. Infection of High Five(TM) cells with baculovirus containing full coding brainiac cDNA markedly increased the ratio of GlcNAcbeta1-3Manbeta1-4Glcbeta1-Cer glycolipids compared with Galbeta1-4Manbeta1-4Glcbeta1-Cer found in wild type cells. We suggest that brainiac exerts its biological functions by regulating biosynthesis of glycosphingolipids. PMID- 12130652 TI - The NH2-terminal domain of Golgin-160 contains both Golgi and nuclear targeting information. AB - Golgin-160 is a member of the golgin family of Golgi-localized membrane proteins. The COOH-terminal two-thirds of golgin-160 is predicted to form a coiled-coil, with an NH(2)-terminal "head" domain. To identify the Golgi targeting information in golgin-160, full-length and deletion constructs tagged with green fluorescent protein were generated. The head domain alone was targeted to the Golgi complex in the absence of assembly with endogenous golgin-160. Further truncations from both ends of the head domain narrowed the Golgi targeting information to 85 amino acids between residues 172 and 257. Surprisingly, certain truncations of the head domain also specifically accumulated in the nucleus. Both a nuclear localization signal (masked in the full-length protein) and information for nuclear retention contributed to the nuclear localization of these truncations. Because the golgin 160 head is cleaved by caspases during apoptosis, we examined the localization of epitope-tagged proteins corresponding to all potential caspase cleavage fragments. Our data suggest that three of six fragments could be targeted to the nucleus, provided that they are released from Golgi membranes after cleavage. The finding that both Golgi and nuclear targeting information is present in the same region of golgin-160 suggests that this protein may have more than one function. PMID- 12130653 TI - Absence of small conductance K+ channel (SK) activity in apical membranes of thick ascending limb and cortical collecting duct in ROMK (Bartter's) knockout mice. AB - The ROMK (Kir1.1; Kcnj1) gene is believed to encode the apical small conductance K(+) channels (SK) of the thick ascending limb (TAL) and cortical collecting duct (CCD). Loss-of-function mutations in the human ROMK gene cause Bartter's syndrome with renal Na(+) wasting, consistent with the role of this channel in apical K(+) recycling in the TAL that is crucial for NaCl reabsorption. However, the mechanism of renal K(+) wasting and hypokalemia that develop in individuals with ROMK Bartter's syndrome is not apparent given the proposed loss of the collecting duct SK channel. Thus, we generated a colony of ROMK null mice with approximately 25% survival to adulthood that provides a good model for ROMK Bartter's syndrome. The remaining 75% of null mice die in less than 14 days after birth. The surviving ROMK null mice have normal gross renal morphology with no evidence of significant hydronephrosis, whereas non-surviving null mice exhibit marked hydronephrosis. ROMK protein expression was absent in TAL and CCD from null mice but exhibited normal abundance and localization in wild-type littermates. ROMK null mice were polyuric and natriuretic with an elevated hematocrit consistent with mild extracellular volume depletion. SK channel activity in TAL and CCD was assessed by patch clamp analysis in ROMK wild-type ROMK(+/+), heterozygous ROMK(+/-), and null ROMK(-/-) mice. In 313 patches with successful seals from the three ROMK genotypes, SK channel activity in ROMK (+/+ and +/-) exhibited normal single channel kinetics. The expression frequencies are as follows: 67 (TAL) and 58% (CCD) in ROMK(+/+); about half that of the wild-type in ROMK(+/-), being 38 (TAL) and 25% (CCD); absent in both TAL or CCD in ROMK(-/-) between 2 and 5 weeks in 15 mice (61 and 66 patches, respectively). The absence of SK channel activity in ROMK null mice demonstrates that ROMK is essential for functional expression of SK channels in both TAL and CCD. Despite loss of ROMK expression, the normokalemic null mice exhibited significantly increased kaliuresis, indicating alternative mechanisms for K(+) absorption/secretion in the nephron. PMID- 12130654 TI - In vivo and in vitro phosphorylation at Ser-493 in the glutamate (E)-segment of neurofilament-H subunit by glycogen synthase kinase 3beta. AB - Neurofilament (NF), a major neuronal intermediate filament, is composed of three subunits, NF-L, NF-M, and NF-H. All three subunits contain a well conserved glutamate (E)-rich region called "E-segment" in the N terminus of the tail region. Although the E-segments of NF-L and NF-M are phosphorylated by casein kinases, it has not been observed in NF-H. Using mass spectrometric analysis, we identified phosphorylation of the E-segment of NF-H, prepared from rat spinal cords, at Ser-493 and Ser-501 in the Ser-Pro sequences. The E-segment kinase was isolated from rat brain extract using column chromatography and identified as glycogen synthase kinase (GSK) 3beta. GSK3beta was shown to phosphorylate at Ser 493 in vitro by phosphopeptide mapping and site-directed mutagenesis, and in vivo in HEK293 cells using the phospho-Ser-493 antibody, but did not phosphorylate Ser 501. GSK3beta preferred Ser-493 to the KSP-repeated sequences for phosphorylation sites in the NF-H tail domain. Moreover, Ser-493 was a better phosphorylation site for GSK3beta than other proline-directed protein kinases, Cdk5/p35 and ERK. GSK3beta in the spinal cord extract was associated with NF cytoskeletons. Taken together, we concluded that Ser-493 in the E-segment of NF-H is phosphorylated by GSK3beta in rat spinal cords. PMID- 12130656 TI - Carbamoyl-phosphate synthetase. Creation of an escape route for ammonia. AB - Carbamoyl-phosphate synthetase catalyzes the production of carbamoyl phosphate through a reaction mechanism requiring one molecule of bicarbonate, two molecules of MgATP, and one molecule of glutamine. The enzyme from Escherichia coli is composed of two polypeptide chains. The smaller of these belongs to the Class I amidotransferase superfamily and contains all of the necessary amino acid side chains required for the hydrolysis of glutamine to glutamate and ammonia. Two homologous domains from the larger subunit adopt conformations that are characteristic for members of the ATP-grasp superfamily. Each of these ATP-grasp domains contains an active site responsible for binding one molecule of MgATP. High resolution x-ray crystallographic analyses have shown that, remarkably, the three active sites in the E. coli enzyme are connected by a molecular tunnel of approximately 100 A in total length. Here we describe the high resolution x-ray crystallographic structure of the G359F (small subunit) mutant protein of carbamoyl phosphate synthetase. This residue was initially targeted for study because it resides within the interior wall of the molecular tunnel leading from the active site of the small subunit to the first active site of the large subunit. It was anticipated that a mutation to the larger residue would "clog" the ammonia tunnel and impede the delivery of ammonia from its site of production to the site of utilization. In fact, the G359F substitution resulted in a complete change in the conformation of the loop delineated by Glu-355 to Ala-364, thereby providing an "escape" route for the ammonia intermediate directly to the bulk solvent. The substitution also effected the disposition of several key catalytic amino acid side chains in the small subunit active site. PMID- 12130655 TI - 8-(3-Chlorostyryl)caffeine may attenuate MPTP neurotoxicity through dual actions of monoamine oxidase inhibition and A2A receptor antagonism. AB - Caffeine and more specific antagonists of the adenosine A(2A) receptor recently have been found to be neuroprotective in the MPTP (1-methyl-4-phenyl-1,2,3,6 tetrahydropyridine) model of Parkinson's disease. Here we show that 8-(3 chlorostyryl)caffeine (CSC), a specific A(2A) antagonist closely related to caffeine, also attenuates MPTP-induced neurotoxicity. Because the neurotoxicity of MPTP relies on its oxidative metabolism to the mitochondrial toxin MPP(+), we investigated the actions of CSC on striatal MPTP metabolism in vivo. CSC elevated striatal levels of MPTP but lowered levels of the oxidative intermediate MPDP(+) and of MPP(+), suggesting that CSC blocks the conversion of MPTP to MPDP(+) in vivo. In assessing the direct effects of CSC and A(2A) receptors on monoamine oxidase (MAO) activity, we found that CSC potently and specifically inhibited mouse brain mitochondrial MAO-B activity in vitro with a K(i) value of 100 nm, whereas caffeine and another relatively specific A(2A) antagonist produced little or no inhibition. The A(2A) receptor independence of MAO-B inhibition by CSC was further supported by the similarity of brain MAO activities derived from A(2A) receptor knockout and wild-type mice and was confirmed by demonstrating potent inhibition of A(2A) receptor knockout-derived MAO-B by CSC. Together, these data indicate that CSC possesses dual actions of MAO-B inhibition and A(2A) receptor antagonism, a unique combination suggesting a new class of compounds with the potential for enhanced neuroprotective properties. PMID- 12130657 TI - Cysteine activation is an inherent in vitro property of prolyl-tRNA synthetases. AB - Aminoacyl-tRNA synthetases are well known for their remarkable precision in substrate selection during aminoacyl-tRNA formation. Some synthetases enhance the accuracy of this process by editing mechanisms that lead to hydrolysis of incorrectly activated and/or charged amino acids. Prolyl-tRNA synthetases (ProRSs) can be divided into two structurally divergent groups, archaeal-type and bacterial-type enzymes. A striking difference between these groups is the presence of an insertion domain (approximately 180 amino acids) in the bacterial type ProRS. Because the archaeal-type ProRS enzymes have been shown to recognize cysteine, we tested selected ProRSs from all three domains of life to determine whether cysteine activation is a general property of ProRS. Here we show that cysteine is activated by recombinant ProRS enzymes from the archaea Methanocaldococcus jannaschii and Methanothermobacter thermautotrophicus, from the eukaryote Saccharomyces cerevisiae, and from the bacteria Aquifex aeolicus, Borrelia burgdorferi, Clostridium sticklandii, Cytophaga hutchinsonii, Deinococcus radiodurans, Escherichia coli, Magnetospirillum magnetotacticum, Novosphingobium aromaticivorans, Rhodopseudomonas palustris, and Thermus thermophilus. This non-cognate amino acid was efficiently acylated in vitro onto tRNA(Pro), and the misacylated Cys-tRNA(Pro) was not edited by ProRS. Therefore, ProRS exhibits a natural level of mischarging that is to date unequalled among the aminoacyl-tRNA synthetases. PMID- 12130658 TI - Methanocaldococcus jannaschii prolyl-tRNA synthetase charges tRNA(Pro) with cysteine. AB - Methanocaldococcus jannaschii prolyl-tRNA synthetase (ProRS) was previously reported to also catalyze the synthesis of cysteinyl-tRNA(Cys) (Cys-tRNA(Cys)) to make up for the absence of the canonical cysteinyl-tRNA synthetase in this organism (Stathopoulos, C., Li, T., Longman, R., Vothknecht, U. C., Becker, H., Ibba, M., and Soll, D. (2000) Science 287, 479-482; Lipman, R. S., Sowers, K. R., and Hou, Y. M. (2000) Biochemistry 39, 7792-7798). Here we show by acid urea gel electrophoresis that pure heterologously expressed recombinant M. jannaschii ProRS misaminoacylates M. jannaschii tRNA(Pro) with cysteine. The enzyme is unable to aminoacylate purified mature M. jannaschii tRNA(Cys) with cysteine in contrast to facile aminoacylation of the same tRNA with cysteine by Methanococcus maripaludis cysteinyl-tRNA synthetase. Although M. jannaschii ProRS catalyzes the synthesis of Cys-tRNA(Pro) readily, the enzyme is unable to edit this misaminoacylated tRNA. We discuss the implications of these results on the in vivo activity of the M. jannaschii ProRS and on the nature of the enzyme involved in the synthesis of Cys-tRNA(Cys) in M. jannaschii. PMID- 12130659 TI - A tetrameric porin limits the cell wall permeability of Mycobacterium smegmatis. AB - Mycobacteria protect themselves with an outer lipid bilayer, which is the thickest biological membrane hitherto known and has an exceptionally low permeability rendering mycobacteria intrinsically resistant against many antibiotics. Pore proteins mediate the diffusion of hydrophilic nutrients across this membrane. Electron microscopy revealed that the outer membrane of Mycobacterium smegmatis contained about 1000 protein pores per microm(2), which are about 50-fold fewer pores per microm(2) than in Gram-negative bacteria. The projection structure of the major porin MspA of M. smegmatis was determined at 17 A resolution. MspA forms a cone-like tetrameric complex of 10 nm in length with a single central pore. Thus, MspA is drastically different from the trimeric porins of Gram-negative bacteria and represents a new class of channel proteins. The formation of MspA micelles indicated that the ends of MspA have different hydrophobicities. Oriented insertion of MspA into membranes was demonstrated in lipid bilayer experiments, which revealed a strongly asymmetrical voltage gating of MspA channels at -30 mV. The length of MspA is sufficient to span the outer membrane and contributes in combination with the tapering end of the pore and the low number of pores to the low permeability of the cell wall of M. smegmatis for hydrophilic compounds. PMID- 12130660 TI - Tissue-specific regulation of retinal and pituitary precursor cell proliferation. AB - Mammalian organogenesis requires the expansion of pluripotent precursor cells before the subsequent determination of specific cell types, but the tissue specific molecular mechanisms that regulate the initial expansion of primordial cells remain poorly defined. We have genetically established that Six6 homeodomain factor, acting as a strong tissue-specific repressor, regulates early progenitor cell proliferation during mammalian retinogenesis and pituitary development. Six6, in association with Dach corepressors, regulates proliferation by directly repressing cyclin-dependent kinase inhibitors, including the p27Kip1 promoter. These data reveal a molecular mechanism by which a tissue-specific transcriptional repressor-corepressor complex can provide an organ-specific strategy for physiological expansion of precursor populations. PMID- 12130662 TI - Probing oxygen motion in disordered anionic conductors with 17O and 51V MAS NMR spectroscopy. AB - Identification of the local environments of the ions in a solid-state electrolyte that contribute to the ionic conductivity or remain trapped in the lattice represents a challenge for many experimental probes of structure. We show that high-resolution 17O magic angle spinning nuclear magnetic resonance (MAS NMR) spectra may be obtained even from the highly disordered, layered materials alpha Bi4V2O11 and gamma-Bi4V1.7Ti0.3O10.85, in which the different oxide sites in the lattice may be distinguished. The sites responsible for anionic conduction were determined directly from the variable-temperature 17O NMR spectra, and correlation times for motion were estimated. Double-resonance 17O/51V NMR methods were used as confirmation of the assignments of the resonances and as a second experimental probe of motion that is sensitive to mobility involving oxide ion hops between the same crystallographic sites. PMID- 12130661 TI - Impaired B and T cell antigen receptor signaling in p110delta PI 3-kinase mutant mice. AB - Class IA phosphoinositide 3-kinases (PI3Ks) are a family of p85/p110 heterodimeric lipid kinases that generate second messenger signals downstream of tyrosine kinases, thereby controlling cell metabolism, growth, proliferation, differentiation, motility, and survival. Mammals express three class IA catalytic subunits: p110alpha, p110beta, and p110delta. It is unclear to what extent these p110 isoforms have overlapping or distinct biological roles. Mice expressing a catalytically inactive form of p110delta (p110delta(D910A)) were generated by gene targeting. Antigen receptor signaling in B and T cells was impaired and immune responses in vivo were attenuated in p110delta mutant mice. They also developed inflammatory bowel disease. These results reveal a selective role for p110delta in immunity. PMID- 12130664 TI - Urokinase plasminogen activator receptor: Prognostic biomarker for endometrial cancer. AB - Endometrial adenocarcinoma is the most common gynecologic malignancy in the United States. However, reliable diagnostic or prognostic tumor markers have not been identified for endometrial cancer. In this study, we examined whether urokinase plasminogen activator receptor (UPAR), a glycosyl-phosphatidylinositol linked membrane protein, is a candidate diagnostic or prognostic marker for patients with cancer of the endometrium. Sixty-five surgically excised, formalin fixed endometrial tissue specimens were accessioned through the Department of Pathology Registry at the University of California, Los Angeles, and analyzed for UPAR expression by using immunohistochemical techniques. A retrospective review was also performed to determine stage and histopathologic grade of disease, recurrence, and mortality. No expression of UPAR protein was present in seven patients with benign neoplasia of the endometrium. UPAR protein expression highly correlated with stage of disease (ungrouped Spearman correlation = 0.625, P < 0.0001): 40% of patients with stage I, 66% of patients with stage II, 100% of patients with stage III, and 85% with stage IV demonstrated the highest level of UPAR expression. Moreover, high UPAR expression positively correlated with grade of disease (ungrouped Spearman correlation = 0.71, P < 0.0001): 29% of grade 1 specimens, 57% of grade 2, and over 90% of specimens with grade 3, the majority representing uterine papillary serous carcinoma and mixed malignant mesodermal tumor. Finally, UPAR protein expression also positively correlated with rate of recurrence and mortality in patients with adenocarcinoma of the endometrium (ungrouped P = 0.034). Our data suggest that UPAR is a useful prognostic marker for biologically aggressive forms of endometrial cancer. PMID- 12130663 TI - Heterologous desensitization of opioid receptors by chemokines inhibits chemotaxis and enhances the perception of pain. AB - The chemokines use G protein-coupled receptors to regulate the migratory and proadhesive responses of leukocytes. Based on observations that G protein-coupled receptors undergo heterologous desensitization, we have examined the ability of chemokines to also influence the perception of pain by cross-desensitizing opioid G protein-coupled receptors function in vitro and in vivo. We find that the chemotactic activities of both mu- and delta-opioid receptors are desensitized following activation of the chemokine receptors CCR5, CCR2, CCR7, and CXCR4 but not of the CXCR1 or CXCR2 receptors. Furthermore, we also find that pretreatment with RANTES/CCL5, the ligand for CCR1, and CCR5 or SDF-1alpha/CXCL12, the ligand for CXCR4, followed by opioid administration into the periaqueductal gray matter of the brain results in an increased rat tail flick response to a painful stimulus. Because chemokine administration into the periaqueductal gray matter inhibits opioid-induced analgesia, we propose that the activation of proinflammatory chemokine receptors down-regulates the analgesic functions of opioid receptors, and this enhances the perception of pain at inflammatory sites. PMID- 12130665 TI - Erythropoietin administration protects retinal neurons from acute ischemia reperfusion injury. AB - Erythropoietin (EPO) plays an important role in the brain's response to neuronal injury. Systemic administration of recombinant human EPO (rhEPO) protects neurons from injury after middle cerebral artery occlusion, traumatic brain injury, neuroinflammation, and excitotoxicity. Protection is in part mediated by antiapoptotic mechanisms. We conducted parallel studies of rhEPO in a model of transient global retinal ischemia induced by raising intraocular pressure, which is a clinically relevant model for retinal diseases. We observed abundant expression of EPO receptor (EPO-R) throughout the ischemic retina. Neutralization of endogenous EPO with soluble EPO-R exacerbated ischemic injury, which supports a crucial role for an endogenous EPO/EPO-R system in the survival and recovery of neurons after an ischemic insult. Systemic administration of rhEPO before or immediately after retinal ischemia not only reduced histopathological damage but also promoted functional recovery as assessed by electroretinography. Exogenous EPO also significantly diminished terminal deoxynucleotidyltransferase-mediated dUTP end labeling labeling of neurons in the ischemic retina, implying an antiapoptotic mechanism of action. These results further establish EPO as a neuroprotective agent in acute neuronal ischemic injury. PMID- 12130667 TI - The structure of a replication initiator unites diverse aspects of nucleic acid metabolism. AB - Rolling circle replication is a mechanism for copying single-stranded genomes by means of double-stranded intermediates. A multifunctional replication initiator protein (Rep) is indispensable for the precise initiation and termination of this process. Despite the ubiquitous presence and fundamental importance of rolling circle replication elements, structural information on their respective replication initiators is still missing. Here we present the solution NMR structure of the catalytic domain of Rep, the initiator protein of tomato yellow leaf curl virus. It is composed of a central five-stranded anti-parallel beta sheet, flanked by a small two-stranded beta-sheet, a beta-hairpin and two alpha helices. Surprisingly, the structure reveals that the catalytic Rep domain is related to a large group of proteins that bind RNA or DNA. Identification of Rep as resembling the family of ribonucleoprotein/RNA-recognition motif fold proteins establishes a structure-based evolutionary link between RNA binding proteins, splicing factors, and replication initiators of prokaryotic and eukaryotic single stranded DNA elements and mammalian DNA tumor viruses. PMID- 12130668 TI - Contributions of protein kinase A anchoring proteins to compartmentation of cAMP signaling in the heart. AB - The cAMP-dependent protein kinase (PKA) transduces signals in the heart initiated by beta(1)-adrenergic, G-protein-coupled receptors after norepinephrine, sympathetic stimulation. Signaling through this pathway results in a characteristic set of cellular responses, including increases in ion fluxes and contractile strength, mobilization of energy stores, and changes in gene expression. Not all receptors that activate adenylate cyclase and increase cAMP levels, however, cause the cardiac myocyte to react in this manner. Research in the field of signal transduction over the last 25 years has addressed this issue of specificity in signaling by diffusable second messengers. PKA is in part targeted to discrete cellular locations by A-kinase anchoring proteins. Through anchoring and formation of multienzyme complexes, specific, localized signal transduction is possible. I discuss in this review recent advances in the understanding of PKA signaling complexes in the cardiac myocyte. PMID- 12130669 TI - The yin and the yang of 5-lipoxygenase pathway activation. PMID- 12130666 TI - Mitochondrial DNA and the origins of the domestic horse. AB - The place and date of the domestication of the horse has long been a matter for debate among archaeologists. To determine whether horses were domesticated from one or several ancestral horse populations, we sequenced the mitochondrial D-loop for 318 horses from 25 oriental and European breeds, including American mustangs. Adding these sequences to previously published data, the total comes to 652, the largest currently available database. From these sequences, a phylogenetic network was constructed that showed that most of the 93 different mitochondrial (mt)DNA types grouped into 17 distinct phylogenetic clusters. Several of the clusters correspond to breeds and/or geographic areas, notably cluster A2, which is specific to Przewalski's horses, cluster C1, which is distinctive for northern European ponies, and cluster D1, which is well represented in Iberian and northwest African breeds. A consideration of the horse mtDNA mutation rate together with the archaeological timeframe for domestication requires at least 77 successfully breeding mares recruited from the wild. The extensive genetic diversity of these 77 ancestral mares leads us to conclude that several distinct horse populations were involved in the domestication of the horse. PMID- 12130670 TI - Fusion proteins with anticoagulant and fibrinolytic properties: functional studies and structural considerations. AB - In an effort to combine the benefits of fibrinolytics, such as staphylokinase, with those of thrombin inhibitors for the prevention of vessel reocclusion after vascular injury, we have produced several chimeric proteins with plasminogen activating and thrombin-inhibiting properties. Fusion proteins were constructed consisting of the modules staphylokinase (Sak), the factor Xa cleavage site, and various dipetalin (Dip) domains (H(6)-Sak-Dip-I+II, H(6)-Sak-Dip-I, and H(6)-Sak Dip-II). Sak stimulates fibrinolysis via activation of plasminogen, whereas dipetalin is a two-domain, Kazal-type inhibitor of thrombin. NMR spectroscopy of the fusion proteins revealed that the molecular structures of the modules are retained in the fusion protein and that no significant interactions occur between the modules in terms of their functionally relevant regions. In enzymatic thrombin inhibition tests and blood coagulation assays (thrombin, prothrombin, and activated partial thromboplastin times), no significant differences in anticoagulant capacity were observed between the fusion protein H(6)-Sak-Dip-I+II and isolated Dip-I+II, even at nanomolar concentrations. Similar results (i.e., the inhibition of thrombin-induced platelet aggregation and the inhibition of thrombin-induced vascular relaxation) were obtained when the cellular thrombin effects were studied. The fusion protein containing Dip-I has less but still significant thrombin inhibitory effects compared with those of H(6)-Sak-Dip-I+II. In contrast, the H(6)-Sak-Dip-II protein failed to inhibit thrombin in each of the assays used. The plasminogen-activating and fibrinolytic activities of the fusion proteins are similar to those of wild-type Sak. The individual dipetalin domains do not activate plasminogen. In conclusion, the fusion protein H(6)-Sak Dip-I+II is a bifunctional molecule able to activate fibrinolysis via plasminogen activation and inhibit blood coagulation via direct inhibition of thrombin. PMID- 12130671 TI - The Ca(2+) channel antagonists mibefradil and pimozide inhibit cell growth via different cytotoxic mechanisms. AB - We show that mitogenic cells expressing T-type Ca(2+) channels (T-channels) are more sensitive to the antiproliferative effects of the drugs pimozide and mibefradil than cells without significant T-channel expression. The growth of Y79 and WERI-Rb1 retinoblastoma cells, as well as MCF7 breast cancer epithelial cells, all of which express T-channel current and mRNA for T-channel subunits, is inhibited by pimozide and mibefradil with IC(50) values between 0.6 and 1.5 microM. Proliferation of glioma C6 cells, which show little T-channel expression, is less sensitive to these drugs (IC(50) = 8 and 5 microM for pimozide and mibefradil, respectively). Neither drug seems to alter cell cycle or the expression of cyclins. Although this strong correlation between T-channel expression and growth inhibition exists, the following results suggest that the drugs inhibit cell growth via different cytotoxic pathways: 1) pimozide and mibefradil have additive effects on T-channel current inhibition, whereas the antiproliferative activity of the drugs together is synergistic; 2) an increase in the number of apoptotic Y79 and MCF7 cells and a decrease in the mRNA for the antiapoptotic gene Bcl-2 is detected only in pimozide-treated cells, whereas in mibefradil-treated cells, the toxicity is primarily necrotic; and 3) growth inhibition by mibefradil is reduced in Y79 cells transfected with T-channel antisense and in differentiated Y79 cells (which have decreased T-channel expression), but growth inhibition by pimozide is affected to a lesser extent. These results suggest that pimozide and mibefradil inhibit cell proliferation via different cytotoxic pathways and that in the case of pimozide, it is unlikely that this effect is mediated solely by T-channel inhibition. PMID- 12130672 TI - Re-engineering butyrylcholinesterase as a cocaine hydrolase. AB - To address the problem of acute cocaine overdose, we undertook molecular engineering of butyrylcholinesterase (BChE) as a cocaine hydrolase so that modest doses could be used to accelerate metabolic clearance of this drug. Molecular modeling of BChE complexed with cocaine suggested that the inefficient hydrolysis (k(cat) = 4 min(-1)) involves a rotation toward the catalytic triad, hindered by Tyr332. To eliminate rotational hindrance and retain substrate affinity, we introduced two amino acid substitutions (Ala328Trp/Tyr332Ala). The resulting mutant BChE reduced cocaine burden in tissues, accelerated plasma clearance by 20 fold, and prevented cocaine-induced hyperactivity in mice. The enzyme's kinetic properties (k(cat) = 154 min(-1), K(M) = 18 microM) satisfy criteria suggested previously for treating cocaine overdose (k(cat) >120 min(-1), K(M) < 30 microM). This success demonstrates that computationally guided mutagenesis can generate functionally novel enzymes with clinical potential. PMID- 12130673 TI - Insulin-like growth factor-1-induced phosphorylation of transcription factor FKHRL1 is mediated by phosphatidylinositol 3-kinase/Akt kinase and role of this pathway in insulin-like growth factor-1-induced survival of cultured hippocampal neurons. AB - Insulin-like growth factor-1 (IGF-1) is a trophic factor promoting cell survival by activating phosphatidylinositol 3-kinase (PI3K)/Akt kinase pathway. FKHRL1, a member of the Forkhead family of transcription factors possibly involved in cell cycle and apoptosis, is a downstream target of Akt in fibroblasts. However, very little information is available concerning neurons. We report herein that IGF-1 rapidly induced the phosphorylation of endogenous FKHRL1 in hippocampal neurons. The PI3K/Akt kinase pathway mediates this action, as evidenced by the use of different kinase inhibitors, the expression of constitutively active Akt, and in vitro kinase assay. IGF-1 blocked the nuclear translocation of FKHRL1 in hippocampal neurons and promoted survival in parallel to the phosphorylation of Akt and FKHRL1. Similarly, the expression of constitutively active Akt in PC-12 cells increased the phosphorylation of FKHRL1 and promoted survival, whereas the expression of kinase dead Akt attenuated IGF-1-mediated survival of PC-12 cells. Moreover, the overexpression of wild-type FKHRL1 and its nonphosphorylated mutant induced apoptosis in cultured hippocampal neurons. Interestingly, IGF-1 and PI3 kinase inhibitors have no significant effect on the cell cycle inhibitor p27kip1 in hippocampal neurons. This finding suggests that in contrast to fibroblasts, FKHRL1 is unlikely to be involved in cell cycle in neurons. Taken together, these data reveal that endogenous FKHRL1 is a downstream substrate of PI3K/Akt in IGF-1 receptor signaling in hippocampal neurons and suggest that the phosphorylation of this transcription factor may play an important role in the neuronal survival properties of IGF-1. PMID- 12130674 TI - The zebrafish (Danio rerio) aryl hydrocarbon receptor type 1 is a novel vertebrate receptor. AB - Fish are known to have two distinct classes of aryl hydrocarbon receptors, and their roles in mediating xenobiotic toxicity remain unclear. In this study, we have identified and characterized a cDNA tentatively named zebrafish AHR1 (zfAHR1). Analysis of the deduced amino acid sequence reveals that the protein is distinct from zfAHR2 and is more closely related to the mammalian aryl hydrocarbon receptor (AHR). zfAHR1 and zfAHR2 share 40% amino acid identity overall and 58% in the N-terminal half. The zfAHR1 gene maps to linkage group 16 in a region that shares conserved synteny with human chromosome 7 containing the human AHR, suggesting that the zfAHR1 is the ortholog of the human AHR. zfAHR2 maps to a separate linkage group (LG22). Both zfAHR mRNAs are expressed in early development, but they are differentially expressed in adult tissues. zfAHR2 can dimerize with zfARNT2b and binds with specificity to dioxin-responsive elements (DREs). Under identical conditions, zfAHR1/zfARNT2b/DRE complexes are formed; however, the interactions are considerably weaker. In COS-7 cells expressing zfARNT2b and zfAHR2, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) exposure leads to a significant induction of dioxin-responsive reporter genes. In identical experiments, TCDD exposure fails to induce the reporter gene in zfAHR1-expressing cells. Ligand-binding experiments suggested that the differential zfAHR activities are attributable to differences in TCDD binding because only zfAHR2 exhibits high-affinity binding to [(3)H]TCDD or beta-naphthoflavone. Finally, using chimeric zfAHR1/zfAHR2 constructs, the lack of TCDD-mediated transcriptional activity was localized to the ligand-binding and C-terminal domains of zfAHR1. PMID- 12130675 TI - Cyclic AMP-mediated inhibition of 5-lipoxygenase translocation and leukotriene biosynthesis in human neutrophils. AB - 5-Lipoxygenase (5-LO) catalyzes the transformation of arachidonic acid to leukotrienes (LT). In stimulated human PMN, activation of 5-LO involves calcium, p38 MAP kinase (p38) phosphorylation, and translocation of 5-LO from the cytosol to nuclear membranes containing the 5-LO activating protein (FLAP). In this study, cAMP-elevating agents such as isoproterenol, prostaglandin E(2), CGS-21680 (an adenosine A(2a) receptor agonist), the type IV phosphodiesterase inhibitor RO 20-1724, the adenylate cyclase activator forskolin, and the Gs-protein activator cholera toxin all inhibited LT biosynthesis and 5-LO translocation to the nucleus in cytokine-primed human PMN stimulated with platelet-activating factor and in human PMN stimulated with the endomembrane Ca(2+)-ATPase blocker thapsigargin. Furthermore, monophosphorothioate analogs of cAMP, which activate protein kinase A (PKA), also inhibited LT biosynthesis and 5-LO translocation in stimulated cells. Treatment of PMN with CGS-21680 also prevented the phosphorylation of p38 by thapsigargin. Treatment of PMN with the PKA inhibitors H-89 and KT-5720 prevented the inhibitory effect of cAMP-elevating agents on LT biosynthesis, 5-LO translocation, and p38 phosphorylation, whereas the p38 inhibitor SB 203,580 dose dependently inhibited arachidonic acid-induced LT biosynthesis. The 5-LO translocation was also inhibitable by the FLAP antagonist MK-0591 and correlated with LT biosynthesis in all experimental conditions tested. These results indicate that cAMP-mediated PKA activation in PMN results in the concomitant inhibition of 5-LO translocation and LT biosynthesis and support a role of p38 in the signaling pathway involved. This represents the first physiological down regulation mechanism of 5-LO translocation in human PMN. PMID- 12130676 TI - Apoptosis in murine hepatoma hepa 1c1c7 wild-type, C12, and C4 cells mediated by bilirubin. AB - Elevated serum and tissue bilirubin concentrations that occur in pathological conditions such as cholestasis, jaundice, and other liver diseases are known to stimulate cytotoxic responses. In preliminary studies, we noted that bilirubin seemed to cause apoptosis in murine hepatoma Hepa 1c1c7 wild-type (WT) cells. Consequently, we investigated apoptosis caused by bilirubin in WT, mutant C12 [aryl hydrocarbon receptor (AHR)-deficient], and C4 (AHR nuclear translocator deficient) Hepa 1c1c7 cells. Three independent measures of apoptosis were used to quantify the effects of exogenous bilirubin (0, 1, 10, 25, 50, or 100 microM). Caspase-3 activity and cytochrome c release from mitochondria increased at 3 h post-treatment, before increased caspase-8 activity at 6 h, and nuclear condensation by 24 h after treatment with bilirubin. No differences in whole-cell lipid peroxidation were observed between the cell types; however, intracellular reactive oxygen species (ROS) production was greater in WT cells than C12 or C4 cells 3 h after bilirubin exposure. Pretreatment of cells for 1 h with 1 or 10 microM alpha-naphthoflavone, an AHR antagonist, before bilirubin exposure resulted in decreased caspase-3 activity at 6 h and nuclear condensation at 24 h in WT cells. These results indicate that bilirubin, a potential AHR ligand, causes apoptosis in murine Hepa 1c1c7 WT cells by a mechanism(s) partially involving the AHR, disruption of membrane integrity, and increased intracellular ROS production. PMID- 12130677 TI - Molecular requisites for drug binding to muscle CLC-1 and renal CLC-K channel revealed by the use of phenoxy-alkyl derivatives of 2-(p-chlorophenoxy)propionic acid. AB - CLC channels are a gene family of Cl(-) channels that serve a variety of functions, several of which are involved in genetic diseases. Few specific ligands of CLC channels are known that could be useful as pharmacological tools or potential drugs. We synthesized various derivatives of 2-(p chlorophenoxy)propionic acid, the S(-)-enantiomer of which is a specific blocker of the muscle channel CLC-1. In particular, compounds with different alkyl or phenoxy-alkyl groups on the chiral center, isosteres of the oxygen in the aryloxy moiety, or bioisosteres of the carboxy function were prepared. We found that compounds containing a phenoxy and a phenoxy-alkyl group on the chiral center (bis-phenoxy derivatives) specifically inhibited renal CLC-K channels from the extracellular side with an affinity in the 150-microM range and with almost no effect on other CLC channels when applied from the outside. Surprisingly, the same substances inhibited CLC-1 from the intracellular side in a voltage dependent manner with an apparent K(D) of <5 microM at -140 mV, thus being the most potent blockers of a CLC channel known so far. Although the chlorine atom in para- position of the second phenoxy group was essential for inhibition of CLC-K channels from the outside, it could be substituted by a methoxy group without changing the potency of block for CLC-1 from the inside. These newly identified substances provide powerful tools for studying the structure-function relationship and the physiological role of CLC channels and may represent a starting point for the development of useful drugs targeting CLC-K channels. PMID- 12130678 TI - Ethanol induces gene expression via nuclear compartmentalization of receptor for activated C kinase 1. AB - Scaffolding proteins such as receptor for activated C kinase (RACK) 1 are involved in the targeting of signaling proteins and play an important role in the regulation of signal transduction cascades. Recently, we found that in cultured cells and in vivo, acute ethanol exposure induces the nuclear compartmentalization of RACK1. To elucidate a physiological role for nuclear RACK1, the Tat protein transduction system was used to transduce RACK1 and RACK1 derived fragments into C6 glioma cells. We found that nuclear RACK1 is mediating the induction of the immediate early gene c-fos expression induced by ethanol. First, transduction of full-length RACK1 (Tat-RACK1) resulted in the induction of c-fos expression and enhancement of ethanol activities. Second, we determined that the C terminus of RACK1 (Tat-RACK1DeltaN) is mediating transcription. Third, we identified a dominant negative fragment of RACK1 that inhibited the nuclear compartmentalization of endogenous RACK1 and inhibited ethanol-induction of c-fos mRNA and protein expression. Last, acute exposure to ethanol or transduction of full-length Tat-RACK1 resulted in an increase in mRNA levels of an activator protein 1 site-containing gene, PAC1 (pituitary adenylate cyclase-activating polypeptide receptor type I), suggesting that nuclear RACK1 is involved in the regulation of the expression of genes that are altered upon acute ethanol treatment. These results may therefore have important implications for the study of alcohol addiction. PMID- 12130679 TI - Negative and positive regulatory epitopes in the C-terminal domains of the human B1 and B2 bradykinin receptor subtypes determine receptor coupling efficacy to G(q/11)-mediated [correction of G(9/11)-mediated] phospholipase Cbeta activity. AB - The human B1 bradykinin (BK) receptor (B1R) is more efficacious than the human B2 BK receptor (B2R) in both ligand-independent and agonist-dependent coupling to G(q/11)-mediated phospholipase Cbeta activity. In fact, B1R is constitutively active, whereas B2R exhibits little if any constitutive activity. To evaluate the role of the C-terminal domain in receptor G(q/11) coupling, we constructed chimeric and C-terminally truncated receptors. The slopes of the increase in basal and agonist-dependent cellular phosphoinositide hydrolysis as a function of receptor density in transiently transfected human embryonic kidney 293 cells provided parameters of receptor coupling. Exchanging the C-terminal domains between the two receptors revealed that these domains are largely responsible for the difference in coupling. B1R truncation showed that this receptor does not directly depend on the C-terminal domain for efficient coupling, although coupling is dramatically augmented by residues in the membrane-distal portion of the domain downstream from Tyr(327). On the other hand, coupling of B2R is absolutely dependent on a membrane-proximal epitope in the C-terminal domain upstream from Lys(315). This epitope is adjacent to a basic residue, Arg(311), which exerts an inhibitory effect on coupling. Arg(311) is not conserved in B1R, and complementary mutations in B2R and B1R showed that this residue, together with previously identified serines and threonines, acts to attenuate the coupling efficacy of B2R. Therefore, the C-terminal domain participates intimately in the efficacy of B1R and B2R G(q/11) coupling by contributing both positive and negative regulatory epitopes. PMID- 12130680 TI - Inhibition of branched-chain alpha-keto acid dehydrogenase kinase and Sln1 yeast histidine kinase by the antifungal antibiotic radicicol. AB - The 90-kDa heat shock family (HSP90) of protein and two-component histidine kinases, although quite distinct at the primary amino acid sequence level, share a common structural ATP-binding domain known as the Bergerat fold. The Bergerat fold is important for the ATPase activity and associated chaperone function of HSP90. Two-component histidine kinases occur in bacteria, yeast, and plants but have yet to be identified in mammalian cells. The antifungal antibiotic radicicol (Monorden) has been shown to bind to the Bergerat fold of HSP90 and to inhibit its ATPase activity. The structural similarity between the Bergerat fold of HSP90 and bacterial two-component histidine kinases prompted our inquiry into whether radicicol could be a potential inhibitor of histidine kinase-like proteins. Structural homology searches suggest that the ATP-binding domains of the yeast histidine kinase Sln1 and the mammalian, branched-chain alpha-keto acid dehydrogenase kinase are very similar to that of other Bergerat fold family members. On the basis of structural homology, we tested radicicol as a potential inhibitor of Sln1 and branched-chain alpha-keto acid dehydrogenase kinase (BCKDHK) and propose a mechanism of inhibition of these kinases. Although BCKDHK has been shown to have serine autophosphorylation activity, we speculate, based on the results from this study and other supporting evidence, that BCKDHK may also have intrinsic histidine kinase activity. PMID- 12130682 TI - Influence of ceramide metabolism on P-glycoprotein function in immature acute myeloid leukemia KG1a cells. AB - Previous studies have emphasized the role of glucosylceramide (Glu-Cer) synthase in multidrug resistance (MDR) regulation. However, the mechanism by which the inhibition of this enzyme results in increased drug retention and cytotoxicity remains unclear. In this study, we investigated the respective role of ceramide (Cer) accumulation and Glu-Cer derivatives depletion in MDR reversal effect of 1 phenyl-2-decanoylamino-3-morpholino-1-propanolol (PDMP), a Glu-Cer synthase inhibitor. We show here that treatment with PDMP resulted in increased rhodamine 123 (Rh123) retention and potent chemosensitization of P-glycoprotein (P-gp) expressing cells, including KG1a cells, KG1a/200 cells, K562/138 cells, and K562/mdr-1 cells. Metabolic studies revealed that PDMP induced not only time dependent Cer accumulation but also reduction of all glycosylated forms of Cer, including Glu-Cer, lactosylceramide (Lac-Cer), monosialo ganglioside (GM3) and disialo ganglioside (GD3). The influence of these metabolites on P-gp function was investigated by measuring Rh123 retention in PDMP-treated cells. P-gp function was found to be stimulated only by the addition of gangliosides in all resistant cell lines, whereas Glu-Cer, Lac-Cer, and Cer had no effect. Moreover, in KG1a/200 cells, GD3 and, to a lesser extent, GM3 were found to phosphorylate P gp on serine residues. Altogether, these results suggest that, at least in leukemic cells, gangliosides depletion accounts for PDMP-mediated MDR reversal effect, and that gangliosides are important P-gp regulators perhaps through their capacity to modulate P-gp phosphorylation. PMID- 12130683 TI - 7,12-Dimethylbenz[a]anthracene induces apoptosis in murine pre-B cells through a caspase-8-dependent pathway. AB - Polycyclic aromatic hydrocarbons (PAHs) have been demonstrated to cause a variety of tumors and immunosuppressive effects. Our laboratory, and others, have demonstrated that coculture of progenitor B lymphocytes (pre-B cells) with bone marrow stromal cells and the model PAH 7,12-dimethylbenz[a]anthracene (DMBA) results in pre-B cell apoptosis. In this study we investigated the molecular events that precede apoptosis in DMBA-treated 70Z/3 cells, a pre-B cell line. Using caspase activity assays and immunoblotting techniques, we determined the temporal pattern of caspase expression in the pre-B cells. Using caspase inhibitors, we demonstrated that DMBA-mediated pre-B cell apoptosis is dependent on activation of caspase-8, whereas caspase-9 activation is essential for maximal apoptosis. We also demonstrated that DMBA activated PKR, an interferon-inducible protein kinase, in pre-B cells. PKR in turn can activate caspase-8 independently of death receptor ligation. As a result of these studies, we propose a novel PKR dependent pathway for activation of apoptosis in DMBA-treated pre-B cells. PMID- 12130681 TI - trans-Activation and repression properties of the novel nonsteroid glucocorticoid receptor ligand 2,5-dihydro-9-hydroxy-10-methoxy-2,2,4-trimethyl-5-(1 methylcyclohexen-3-y1)-1H-[1]benzopyrano[3,4-f]quinoline (A276575) and its four stereoisomers. AB - Glucocorticoids are potent anti-inflammatory and immunosuppressant agents. However, they also produce serious side effects that limit their usage. It has been proposed that anti-inflammatory properties of glucocorticoids are caused mostly by repression of activator protein 1- and nuclear factor kappabeta stimulated synthesis of inflammatory mediators, whereas most of their adverse effects are associated with trans-activation of genes involved with metabolic processes. Our laboratories have sought to discover novel glucocorticoid receptor (GR) ligands that have high repression but low trans-activation activities. We describe here cellular properties of 2,5-dihydro-9-hydroxy-10-methoxy-2,2,4 trimethyl-5-(1-methylcyclohexen-3-y1)-1H-[1]benzopyrano[3,4-f]quinoline (A276575) and its four enantiomers. Similar to dexamethasone, A276575 exhibited high affinity for GR and potently repressed interleukin (IL) 1beta-stimulated IL-6 production in human skin fibroblasts, prostaglandin (PG) E(2) production in A549 human lung epithelial cells, and concanavalin A-induced monocyte proliferation. In contrast to dexamethasone, A276575 caused smaller induction of aromatase activity in human skin fibroblasts and antagonized dexamethasone-induced activation of an mouse mammary tumor virus-glucocorticoid-response element (GRE) reporter gene construct. Among the four enantiomers of A276575, the two (-) enantiomers showed 10- to 30-fold higher affinities for GR than their respective (+)-enantiomers. Both (-)-Syn and (-)-Anti enantiomers of A276575 were potent inhibitors of IL-1beta-stimulated PGE2 production in A549 lung epithelial cells; unexpectedly, however, only the (-)-Anti enantiomer inhibited regulated on T-cell activation, normal T-cell expressed and secreted (RANTES) production in A549 cells. In summary, A276575 is a novel, nonsteroidal GR ligand that possesses high repression activities against inflammatory mediator production but has lower GRE trans-activation activities than traditional steroids. Differential repression of RANTES and PGE2 production in a cell by the two (-)-enantiomers of A276575 illustrates the complexity of repression by GR. PMID- 12130684 TI - Alternative processing of the human FMO6 gene renders transcripts incapable of encoding a functional flavin-containing monooxygenase. AB - The flavin-containing monooxygenases (FMOs) are a family of five microsomal enzymes important for the oxidative metabolism of environmental toxicants, natural products, and therapeutics. With the exception of FMO5, the FMO are encoded within a single gene cluster on human chromosome 1q23-25. As part of the human genome effort, an FMO-like gene, FMO6, was identified between FMO3 and FMO2 (GenBank accession no. AL021026). Sequence analysis of this putative FMO family member revealed nothing that would a priori argue against a functional gene and encoded protein. When FMO6 expression was examined by reverse transcriptase coupled polymerase chain reaction DNA amplification, transcripts were identified in 8 of 11 human liver samples, but 0 of 4 kidney biopsy samples. However, in all cases, the observed transcripts were shorter than predicted. Sequence analysis revealed skipping of exon 4, exons 3 and 4, and/or the use of alternative splice donor or acceptor sites in introns 3, 4, 6, and 8, resulting in nine unique transcripts. Based on an analysis of possible open reading frames, none of these transcripts would encode a functional FMO enzyme. Taking advantage of the high sequence identity between FMO3 and FMO6, it is posited that the loss of binding sites for the serine-arginine-rich splicing factor protein family within exons 3 and 4 contributes to the exon skipping events, although the most commonly observed alternative splice event results from a 21-bp insertion immediately 3' to the predicted FMO6 intron 8 splice acceptor site, diminishing the efficiency of this site. PMID- 12130685 TI - Transcriptional regulation of basal cyclooxygenase-2 expression in murine lung tumor-derived cell lines by CCAAT/enhancer-binding protein and activating transcription factor/cAMP response element-binding protein. AB - Cyclooxygenase-2 (COX-2) is frequently expressed in cancer cells, contributing to tumor development. Most studies of COX-2 expression have examined artificially induced expression in noncancer cells rather than basal expression in cancer cells. Therefore, basal COX-2 expression and its regulation were examined in cell lines derived from a murine model of lung adenocarcinoma. The presence of COX-2 protein in these cells was demonstrated by Western analysis. COX-2 promoter activity was repressed by U0126 [1,4-diamino-2,3-dicyano-1,4-bis(2 aminophenylthio)butadiene], a mitogen-activated protein kinase kinase inhibitor, as well as SB202190 [4-(4-fluorophenyl)-2-(4-hydroxyphenyl)-5-(4-pyridyl)-1H imidazole], an inhibitor of p38 mitogen-activated protein kinase, substantiating the involvement of these signal transduction pathways in the regulation of basal COX-2 expression. Retinoic acid also repressed promoter activity, yet increased activity significantly in one cell line after 18 and 30 h of treatment. Deletions of the murine COX-2 promoter revealed that the 5' transcription factor binding sites were not required for basal expression, including the only nuclear factor kappaB sites of the promoter. Site-directed mutagenesis of the 3' C/EBP (CCAAT/enhancer-binding protein) sites inhibited promoter activity by 20 to 55%, while mutation of the 3' ATF/CREB/AP-1 (activating transcription factor/cAMP response element-binding protein/activator protein-1) site inhibited activity by 70%. Mutation of the 3' upstream stimulatory factor site did not affect promoter activity. Electrophoretic mobility shift assays indicated that the AP-1 transcription factor does not bind to the 3' ATF/CREB/AP-1 site, leaving C/EBP and ATF/CREB as the major transcriptional regulators of basal expression of COX-2 in these lung tumor-derived cell lines and identifying new targets for the prevention/treatment of lung cancer through the modulation of COX-2 expression. PMID- 12130686 TI - A polymorphism in the mouse neuronal alpha4 nicotinic receptor subunit results in an alteration in receptor function. AB - Nicotine-stimulated (86)Rb(+) efflux and [(3)H]cytisine binding, both of which seem to measure the nicotinic acetylcholine receptor, composed of alpha4 and beta2 subunits, were assessed in eight brain regions obtained from 14 inbred mouse strains. The potential role of a single nucleotide polymorphism (SNP) in the nicotinic receptor alpha4 subunit gene (Chrna4) on nicotinic receptor binding and function in mice was also evaluated. This SNP leads to an alanine-to threonine variation at amino acid position 529 of the nascent alpha4 subunit polypeptide. Both nicotine-stimulated (86)Rb(+) efflux and [(3)H]cytisine binding were found to vary across brain regions and among mouse strains. Variability in nicotine-stimulated (86)Rb(+) efflux was positively correlated (r > 0.9) within each strain with the number of [(3)H]cytisine binding sites. However, the number of [(3)H]cytisine binding sites was not correlated with nicotine-stimulated (86)Rb(+) efflux across mouse strains. In contrast, the Chrna4 polymorphism was associated with receptor function across mouse strains: (86)Rb(+) efflux was greater in seven of the eight brain regions studied in those mouse strains that carry the Ala-529 variant of Chrna4. The Chrna4 SNP did not seem to influence the number of [(3)H]cytisine binding sites across mouse strains. These data indicate that inbred mouse strains exhibit differences in receptor function that cannot be attributed to variation in receptor expression but may be explained, at least in part, by the missense polymorphism in the alpha4 subunit. PMID- 12130687 TI - The intracellular loops of the GB2 subunit are crucial for G-protein coupling of the heteromeric gamma-aminobutyrate B receptor. AB - The gamma-aminobutyrate B (GABA(B)) receptor is the first discovered G-protein coupled receptor (GPCR) that needs two subunits, GB1 and GB2, to form a functional receptor. The GB1 extracellular domain (ECD) binds GABA, and GB2 contains enough molecular determinants for G-protein activation. The precise role of the two subunits in G-protein coupling is investigated. GB1 and GB2 are structurally related to the metabotropic glutamate, Ca(2+)-sensing and other family 3 GPCRs in which the second (i2) as well as the third (i3) intracellular loop play important roles in G-protein coupling. Here, the role of the i2 loops of GB1 and GB2 in the GABA(B) receptor ability to activate G(alpha)-proteins is investigated. To that aim, the i2 loops were swapped between GB1 and GB2 heptahelical domains (HDs), either in the wild-type subunits or in the chimeric subunits GB1/2 that contain the ECD of GB1 and the HD of GB2. The effect of an additional mutation within the i3 loop of GB2 that prevents coupling of the heteromeric receptor was also examined. Combinations of interest were found to be correctly addressed at the cell surface and to assemble into heteromers. Taken together our data revealed the following new information on the G-protein coupling of the heteromeric GABA(B) receptor: 1) the i2 loop of GB2 within the GB2 HD is required for the heteromeric GABA(B) receptor to couple to G-proteins, whereas the i2 loop of GB1 is not; 2) the presence of the i2 loop of GB2 within the GB1 HD is not sufficient to allow coupling of GB1; 3) the GB2 HD activates the Gqi9 protein whether it is associated with the GB2 or GB1 ECD; 4) in the combination with two GB2 HDs, each is able to couple to G-proteins; and finally, 5) the use of mutations in i2, i3, or both within the GB2 HD brings evidence for the absence of domain swapping enabling the exchange of region including i2 and i3 between the subunits. PMID- 12130688 TI - Pharmacophore model for novel inhibitors of ubiquitin isopeptidases that induce p53-independent cell death. AB - The tumor suppressor p53 is mutated in more than 50% of all cancers. Importantly, most clinically useful antineoplastic agents are less potent and efficacious in the context of mutant p53. This situation has prompted a search for agents that cause tumor cell death via molecular mechanisms independent of p53. Our recent investigations with electrophilic prostaglandins enabled us to devise a pharmacophore and mechanism of action hypothesis relevant to this problem: a cross-conjugated alpha,beta-unsaturated dienone with two sterically accessible electrophilic beta-carbons is a molecular determinant that confers activity among this class of ubiquitin isopeptidases inhibitors, and that inhibitors of ubiquitin isopeptidases cause cell death in vitro independently of p53. Here, we report the use of the National Cancer Institute's Developmental Therapeutics Database to identify compounds to test this hypothesis. Shikoccin (a diterpene), dibenzylideneacetone, and curcumin fit the pharmacophore hypothesis, inhibit cellular isopeptidases, and cause cell death independently of p53 in isogenic pairs of RKO and HCT 116 cells with differential p53 status. The sesquiterpene achillin and 2,6-diphenyl-4H-thiopyran-4-one, which have cross-conjugated dienones with sterically hindered electrophilic beta-carbons, do not inhibit isopeptidases or cause significant cell death. Furthermore, we show that a catalytic-site proteasome inhibitor causes cell death independently of p53. Combined, these data verify the p53-independence of cell death caused by inhibitors of the proteasome pathway and support the proposition that the ubiquitin-dependent proteasome pathway may contain molecular targets suitable for antineoplastic drug discovery. PMID- 12130689 TI - Transcriptional regulation of the human CYP3A4 gene by the constitutive androstane receptor. AB - Cytochrome P450 3A4 (CYP3A4), the predominant P450 expressed in adult human liver, is both constitutively expressed and transcriptionally activated by a variety of structurally diverse xenochemicals. In this study, we examined the role of the constitutive androstane receptor (CAR), a member of the steroid/retinoid/thyroid hormone receptor superfamily, in the transcriptional regulation of CYP3A4. Herein, we demonstrate that CAR is capable of trans activating expression of the CYP3A4 gene, both in vitro and in vivo. Induction of CYP3A4 is dependent on cooperativity between elements within the promoter proximal region of the gene and the distal xenobiotic-responsive enhancer module. CAR responsiveness was shown to be primarily mediated by two high-affinity binding motifs located within the CYP3A4 gene 5'-flanking region, approximately 7720 and 150 bases upstream of the transcription initiation site. Importantly, the human CAR response elements also mediate trans-activation of CYP3A4 by the human pregnane X receptor, suggesting that interplay between these receptors is likely to be an important determinant of CYP3A4 expression. PMID- 12130690 TI - Modulation of mouse and human phenobarbital-responsive enhancer module by nuclear receptors. AB - The constitutive androstane receptor (CAR) regulates mouse and human CYP2B genes through binding to the direct repeat-4 (DR4) motifs present in the phenobarbital responsive enhancer module (PBREM). The preference of PBREM elements for nuclear receptors and the extent of cross-talk between CAR and other nuclear receptors are currently unknown. Our transient transfection and DNA binding experiments indicate that binding to DR4 motifs does not correlate with the activation response and that mouse and human PBREM are efficiently 'insulated' from the effects of other nuclear receptors despite their substantial affinity for DR4 motifs. Certain nuclear receptors that do not bind to DR4 motifs, such as peroxisome proliferator-activated receptor-alpha and farnesoid X receptor, can suppress PBREM function via a coactivator-dependent process that may have relevance in vivo. In competition experiments, mouse PBREM is clearly more selective for CAR than human PBREM. Pregnane X, vitamin D, and thyroid hormone receptors can potentially compete with human CAR on human PBREM. In contrast to the selective nature of PBREM, CYP3A enhancers are highly and comparably responsive to CAR, pregnane X receptor, and vitamin D receptor. In addition, the ligand specificities of human and mouse CAR were defined by mammalian cotransfection and yeast two-hybrid techniques. Our results provide new mechanistic explanations to several previously unresolved aspects of CYP2B and CYP3A gene regulation. PMID- 12130691 TI - Oxidative stress decreases G protein-coupled receptor kinase 2 in lymphocytes via a calpain-dependent mechanism. AB - G protein-coupled receptor kinase (GRK) 2 plays a crucial role in regulating the extent of desensitization and resensitization of G protein-coupled receptors (GPCRs). We have shown that the expression level of GRK2 in lymphocytes decreases during inflammatory diseases such as arthritis. Reactive oxygen species play an important role in a variety of inflammatory conditions, including arthritis. We demonstrate herein that oxidative stress, induced by exposure of lymphocytes to H(2)O(2), results in a 50% reduction in GRK2 protein levels and GRK activity with no changes in mRNA expression. Treatment of lymphocytes with the tyrosine kinase inhibitor genistein partially reverses the effect of H(2)O(2) on GRK2 levels, although we did not detect direct tyrosine phosphorylation of GRK2. Inhibition of the nonproteasomal protease calpain by calpeptin can prevent the H(2)O(2)-induced GRK2 decrease. In vitro experiments confirm that GRK2 is partially digested by m calpain in a calcium-dependent way. Functionally, H(2)O(2)-induced decrease in GRK2 levels is associated with an ~70% decrease in agonist-induced beta(2) adrenergic receptor sequestration. We describe oxidative stress as a novel mechanism for regulation of the intracellular level of GRK2 during inflammatory processes. Moreover, our data demonstrate that oxidative stress may change the functioning of GPCRs via calpain-dependent regulation of GRK2 levels. PMID- 12130692 TI - Behavioral characterization of mice lacking histamine H(3) receptors. AB - Brain histamine H(3) receptors are predominantly presynaptic and serve an important autoregulatory function for the release of histamine and other neurotransmitters. They have been implicated in a variety of brain functions, including arousal, locomotor activity, thermoregulation, food intake, and memory. The recent cloning of the H(3) receptor in our laboratory has made it possible to create a transgenic line of mice devoid of H(3) receptors. This paper provides the first description of the H(3) receptor-deficient mouse (H(3)(-/-)), including molecular and pharmacologic verification of the receptor deletion as well as phenotypic screens. The H(3)(-/-) mice showed a decrease in overall locomotion, wheel-running behavior, and body temperature during the dark phase but maintained normal circadian rhythmicity. H(3)(-/-) mice were insensitive to the wake promoting effects of the H(3) receptor antagonist thioperamide. We also observed a slightly decreased stereotypic response to the dopamine releaser, methamphetamine, and an insensitivity to the amnesic effects of the cholinergic receptor antagonist, scopolamine. These data indicate that the H(3) receptor deficient mouse represents a valuable model for studying histaminergic regulation of a variety of behaviors and neurotransmitter systems, including dopamine and acetylcholine. PMID- 12130693 TI - Destabilization of the HIV-1 reverse transcriptase dimer upon interaction with N acyl hydrazone inhibitors. AB - N-(4-tert-butylbenzoyl)-2-hydroxy-1-naphthaldehyde hydrazone (BBNH) inhibits both the DNA polymerase and ribonuclease H (RNase H) activities of the human immunodeficiency virus type 1 reverse transcriptase. In this study, we show that BBNH binding impacts on the stability of the human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) heterodimer. The Gibbs free energy of dimer dissociation of HIV-1 RT is decreased in the presence of increasing concentrations of BBNH, resulting in a loss in stability of 3.8 kcal mol(-1). To evaluate whether this observed phenomenon was mediated by BBNH binding to one or more sites in RT, we synthesized a variety of BBNH analogs and identified (4-t butylbenzoyl)-2-hydroxy-1-salicylyl hydrazone (BBSH) and (4,N,N dimethylaminobenzoyl)-2-hydroxy-1-naphthyl hydrazone as specific inhibitors of RT DNA polymerase or RT RNase H activity, respectively. Interestingly, only BBSH provided significant destabilization of the HIV-1 RT dimer. The identification of these specific inhibitors, in combination with other biochemical data, suggests a model in which two molecules of BBNH bind per RT heterodimer. In this regard, only the binding of hydrazone molecules in the DNA polymerase domain activity elicits the observed destabilization of the HIV-1 RT heterodimer. PMID- 12130694 TI - Nicotinic receptor M3 transmembrane domain: position 8' contributes to channel gating. AB - The nicotinic acetylcholine receptor (nAChR) is a pentamer of homologous subunits with composition alpha(2)(beta)(epsilon)(delta) in adult muscle. Each subunit contains four transmembrane domains (M1-M4). Position 8' of the M3 domain is phenylalanine in all heteromeric alpha subunits, whereas it is a hydrophobic nonaromatic residue in non-alpha subunits. Given this peculiar conservation pattern, we studied its contribution to muscle nAChR activation by combining mutagenesis with single-channel kinetic analysis. Construction of nAChRs carrying different numbers of phenylalanine residues at 8' reveals that the mean open time decreases as a function of the number of phenylalanine residues. Thus, all subunits contribute through this position independently and additively to the channel closing rate. The impairment of channel opening increases when the number of phenylalanine residues at 8' increases from two (wild-type nAChR) to five. The gating equilibrium constant of the latter mutant nAChR is 13-fold lower than that of the wild-type nAChR. The replacement of (alpha)F8', (beta)L8', (delta)L8', and (epsilon)V8' by a series of hydrophobic amino acids reveals that the structural bases of the observed kinetic effects are nonequivalent among subunits. In the alpha subunit, hydrophobic amino acids at 8' lead to prolonged channel lifetimes, whereas they lead either to normal kinetics (delta and epsilon subunits) or impaired channel gating (beta subunit) in the non-alpha subunits. The overall results indicate that 8' positions of the M3 domains of all subunits contribute to channel gating. PMID- 12130695 TI - Prenylamine block of Nav1.5 channel is mediated via a receptor distinct from that of local anesthetics. AB - We have shown previously that prenylamine, a calcium channel blocker, has potent local anesthetic activity in vivo and in vitro. We now characterize the tonic and use-dependent block of prenylamine on wild-type human cardiac voltage-gated sodium channels (hNav1.5) transiently expressed in human embryonic kidney 293t cells under whole-cell voltage-clamp condition. We also determine whether prenylamine and local anesthetics interact with a common binding site on the Nav1.5 channel by analyzing prenylamine block on mutant hNav1.5 channels that have substitution mutations in amino acids at the putative local anesthetic binding sites. Prenylamine exhibits tonic block at both hyperpolarizing and depolarizing potentials on hNav1.5 channels with 50% inhibitory concentrations of 9.67 +/- 0.25 microM and 0.72 +/- 0.02 microM, respectively. Substitutions of the amino acids at the putative local anesthetic binding site (i.e., F1760, N1765, Y1767, and N406) with lysine had much lesser effects on prenylamine block of the mutant hNav1.5 channels compared with local anesthetic block. The affinity of prenylamine was reduced at most by 5.8-fold, whereas that of bupivacaine, a known local anesthetic, was reduced by as much as 68-fold compared with wild-type by the mutations at the local anesthetic receptor site. Furthermore, equilibrium results between prenylamine-bupivacaine mixtures suggest two independent receptors. Thus, the data demonstrate that prenylamine has both tonic and use dependent block of hNav1.5 channels similar to that of local anesthetics, but the location of the prenylamine binding site on hNav1.5 differs from that of the local anesthetic binding site. PMID- 12130696 TI - Inflammation modulates the interaction of heterogeneous nuclear ribonucleoprotein (hnRNP) I/polypyrimidine tract binding protein and hnRNP L with the 3'untranslated region of the murine inducible nitric-oxide synthase mRNA. AB - Interaction of two members of the heterogeneous nuclear ribonucleoprotein (hnRNP) family with the 3'untranslated region (UTR) of the murine inducible nitric-oxide synthase (iNOS) mRNA is demonstrated in this study. An iNOS RNA-protein complex is formed using protein extracts from untreated and septic shock treated mouse liver. UV cross-linking reveals that the complex consists of at least two proteins, with apparent molecular masses of 60 and 70 kDa, respectively. The 60 kDa protein binding site lies within a 112-nt pyrimidine-rich sequence, approximately 160 nt from the coding sequence, and the RNA-protein complex can be precipitated by a monoclonal antibody directed against hnRNP I [also named polypyrimidine tract binding protein (PTB)]. The 70-kDa protein binds a 43-nt sequence near the 3'end of the 3'UTR and is immunoprecipitated by a monoclonal antibody against hnRNP L. A computer-simulated conformation of the 3'UTR suggests that both binding sites reside in regions easily accessible for a protein. Supershifts of the native RNA-protein complex could only be achieved with anti hnRNP L, suggesting that within this multiprotein RNA complex, only hnRNP L is exposed to the antibodies, whereas the hnRNP I/PTB is mainly responsible for its interaction with the mRNA. Up-regulation of iNOS by septic shock reduces the RNA protein complex formation, thus showing that hnRNP I/PTB and hnRNP L binding to the iNOS mRNA is modulated by inflammation. This suggests a novel function for the two previously described proteins as regulators of the iNOS gene. PMID- 12130697 TI - Regulation of expression of the multidrug resistance-associated protein 2 (MRP2) and its role in drug disposition. AB - The multidrug resistance protein 2 (MRP2; ABCC2) is an ATP-binding cassette transporter accepting a diverse range of substrates, including glutathione, glucuronide, and sulfate conjugates of many endo- and xenobiotics. MRP2 generally performs excretory or protective roles, and it is expressed on the apical domain of hepatocytes, enterocytes of the proximal small intestine, and proximal renal tubular cells, as well as in the brain and the placenta. MRP2 is regulated at several levels, including membrane retrieval and reinsertion, translation, and transcription. In addition to transport of conjugates, MRP2 transports cancer chemotherapeutics, uricosurics, antibiotics, leukotrienes, glutathione, toxins, and heavy metals. Several mutagenesis studies have described critical residues for substrate binding and various naturally occurring mutations that eliminate MRP2 expression or function. MRP2 is important clinically as it modulates the pharmacokinetics of many drugs, and its expression and activity are also altered by certain drugs and disease states. PMID- 12130698 TI - Genetic polymorphisms of the human CYP2A13 gene: identification of single nucleotide polymorphisms and functional characterization of an Arg257Cys variant. AB - Human cytochrome P450 2A13 (CYP2A13), which is highly efficient in the metabolic activation of a major tobacco-specific carcinogen, 4-(methylnitrosamino)-1-(3 pyridyl)-1-butanone (NNK), may play important roles in xenobiotic toxicity and tobacco-related tumorigenesis in the respiratory tract. The aim of this study was to identify any genetic polymorphisms of the CYP2A13 gene, which may alter the metabolic capacities of the enzyme. Polymerase chain reaction (PCR) single-strand conformational polymorphism analysis was used to identify single-nucleotide polymorphisms (SNPs) in all of the exons and at the exon-intron boundaries, and PCR-restriction fragment length polymorphism analysis and DNA sequencing were used to determine the frequencies of the newly identified variant alleles in the four major ethnic groups. Blood spot DNA from more than 100 individuals was used for these analyses. Seven variant alleles were found, but only one SNP was detected in the coding region, in exon 5, leading to an Arg257Cys amino acid change. The frequencies of the Arg257Cys allele in white, black, Hispanic, and Asian individuals are 1.9%, 14.4%, 5.8%, and 7.7%, respectively. Functional analysis of the variant protein was performed following its heterologous expression. The Arg257Cys variant was 37 to 56% less active than the wild-type Arg-257 protein toward all substrates tested. With NNK, Cys-257 had higher K(m) and lower V(max) values than did Arg-257, with a >2-fold decrease in catalytic efficiency. The Arg257Cys mutation could provide some protection against xenobiotic toxicity in the respiratory tract to individuals who are homozygous for the Cys-257 allele. PMID- 12130699 TI - Effects of methylmercury on human neuronal L-type calcium channels transiently expressed in human embryonic kidney cells (HEK-293). AB - Methylmercury (MeHg) disrupts the function of native, high voltage-activated neuronal Ca(2+) channels in several types of cells. However, the effects of MeHg on isolated Ca(2+) channel phenotypes have not been examined. The aim of the present study was to examine the action of MeHg on recombinant, neuronal L-type voltage-sensitive Ca(2+) channels. Human embryonic kidney cells (HEK-293) were transfected with human neuronal cDNA clones of the alpha(1C-1) subunit in combination with alpha(2b) and beta(3a) Ca(2+) channel subunits and the reporter jellyfish green fluorescent protein for transient expression. Current from expressed channels (I(Ba)) and their response to MeHg applied acutely were measured using whole-cell voltage-clamp recording techniques and Ba(2+) (5 mM) as charge carrier. Amplitude of I(Ba) in these cells was reduced by the dihydropyridine (DHP), nimodipine, and enhanced by Bay K8644 [S-(-)-1,4-dihydro 2,6-dimethyl-5-nitro-4-(2-[trifluoromethyl]phenyl)-3 pyridine carboxylic acid methyl ester]. MeHg (0.125-5.0 microM) caused a time- and concentration-dependent reduction in amplitude of the peak and sustained current through these channels. However, even at the highest concentration of MeHg tested, reduction of current amplitude by MeHg was incomplete. Washing with MeHg-free solution could not reverse its effects. The steady-state inactivation curve was unaltered by MeHg. Increasing the stimulation frequency or the extracellular Ba(2+) concentration each attenuated slightly the reduction in amplitude of I(Ba) by MeHg. In the presence of MeHg (5.0 microM), Bay K8644 still increased the remaining current, and nimodipine (10 microM) reduced residual current that was resistant to MeHg. Thus, although MeHg reduces the amplitude of recombinant, heterologously expressed L-type channel current, a portion of current is resistant to reduction by MeHg. Furthermore, DHP agonists and antagonists retain their ability to affect L-type Ca(2+) channel current even in the presence of MeHg. PMID- 12130700 TI - Dopamine modulates the function of group II and group III metabotropic glutamate receptors in the substantia nigra pars reticulata. AB - Recent findings have shown that dendritically released dopamine (DA) plays an important modulatory role in the substantia nigra pars reticulata (SNr). It is therefore possible that the loss of DA observed in Parkinson's disease (PD) could hold important consequences for nigral function. Previously, we have shown that activation of presynaptically localized group II metabotropic glutamate receptors (mGluRs) inhibits excitatory transmission at the subthalamic nucleus (STN)-SNr synapse and that activation of presynaptically localized group III mGluRs decreases excitatory and inhibitory transmission in the SNr. To test the hypothesis that nigral DA can modulate mGluR function in the SNr, we performed whole-cell patch-clamp recordings from gamma-aminobutyric acidergic SNr neurons in slices obtained from rats that were acutely reserpinized. In slices obtained from reserpinized animals, the effect of group II mGluR activation by the selective agonist (+)-2-aminobicyclo[3.1.0]-hexane-2,6-dicarboxylate monohydrate (LY354740) (100 nM), but not group III mGluR activation [L-(+)-2-amino-4 phosphonobutyric acid, L-AP4, 500 microM], at STN-SNr synapses is significantly decreased. This effect could be mimicked in control slices by prior bath application of haloperidol (20 microM) and R-(+)-7-chloro-8-hydroxy-3-methyl-1 phenyl-2,3,4,5-tetrahydro-1H-3-benzazepine (SCH23390) (20 microM) but not sulpiride (50 microM). Furthermore, application of dopamine (100 microM) and (+/ )-6-chloro-7,8-dyhydroxy-3allyl-1-phenyl-2,3,4,5-tetra-hydro-1H-benzazepine (SKF82958) (1 microM) but not quinpirole (10 microM) could rescue the group II mGluR effect in reserpinized slices. The effect of group III mGluR activation (L AP4, 100 microM) on inhibitory synaptic transmission was also significantly reduced in slices from reserpine-treated animals. This effect was mimicked by haloperidol (20 microM), SCH23390 (20 microM), and sulpiride (50 microM) in control slices. Thus, in a Parkinsonian state, the loss of nigral DA may add to the overall pathophysiological changes in basal ganglia output. PMID- 12130701 TI - Effect of hyperinsulinemia and type 2 diabetes-like hyperglycemia on expression of hepatic cytochrome p450 and glutathione s-transferase isoforms in a New Zealand obese-derived mouse backcross population. AB - In subgroups of a New Zealand obese mouse-derived backcross population with defined aberrations of glucose homeostasis, a comprehensive study of the hepatic expression of cytochrome P450 and glutathione S-transferase was performed. Three patterns of alterations in response to insulin resistance (normoglycemia/hyperinsulinemia) or diabetes (hyperglycemia/hypoinsulinemia) were observed: mRNA levels of Cyp2b9, Cyp3a16, Cyp4a14, and Gstt2 as assessed by Northern- and dot-blot analysis were increased markedly in liver from diabetic mice with no or only a slight increase in insulin resistant mice. Western-blot analysis detected the corresponding changes of the CYP2B and CYP4A proteins. In contrast, expression of Cyp2c22, Cyp2c29, and Cyp2c40 was reduced in diabetic, but normal in insulin resistant mice. These alterations were correlated with changes in serum free fatty acid levels and, therefore, seem to be mediated by the peroxisome proliferator activated receptor-alpha. Furthermore, expression of Cyp1a2, Cyp7b1, Gstm3, and Gstm6 was reduced in both diabetic and insulin resistant mice. Because this third pattern was not correlated with the alterations of serum free fatty acid levels, it seems to reflect an early alteration in the course of the disease, and may be related to the progression of the syndrome from insulin resistance to the type 2-like diabetes. PMID- 12130703 TI - Proteinase-activated receptor (PAR)-1 and -2 agonists induce mediator release from mast cells by pathways distinct from PAR-1 and PAR-2. AB - Because thrombin-induced inflammation is partially mast cell-dependent and involves proteinase-activated receptors (PARs), we hypothesized that mast cells express PAR and can be stimulated with PAR-activating peptides (PAR-AP). We demonstrated that rat peritoneal mast cells expressed PAR-1 and PAR-2 mRNA, and that PAR-2AP (tc-LIGRLO-NH(2), 1 microm) induced 64.2 +/- 4.4% specific beta hexosaminidase release from peritoneal mast cells, whereas another PAR-2AP (SLIGRL-NH(2), 10 microM), trypsin (40 U/ml), and mast cell tryptase (1.5 microg/ml) did not. PAR-1AP (ApfFRChaCitY-NH(2), 10 microM) (Cit) induced 11.7 +/ 3.7% specific beta-hexosaminidase release, whereas another PAR-1AP (TFLLR-NH(2), 40 microM) and human thrombin (10 U/ml) did not. PAR-AP, tc-LIGRLO-NH(2), and Cit increased the free intracellular Ca(2+) concentration, whereas trypsin, tryptase, thrombin, and other PAR-APs did not. Desensitization of Ca(2+) flux with different agonists suggests that although tc-LIGRLO-NH(2), Cit, and compound 48/80 have similar mechanisms of action, tc-LIGRLO-NH(2) also activates mast cells by a mechanism distinct from that of 48/80. Using benzalkonium chloride, which antagonizes the actions of 48/80 by competing for the same G(i) protein, we determined that benzalkonium chloride suppressed tc-LIGRLO-NH(2)-mediated (0.1 microM) beta-hexosaminidase release by 62%. Moreover, removal of sialic acid from peritoneal mast cells, using neuraminidase (2 U/ml), inhibited Cit- (10 microM, 52%) and tc-LIGRLO-NH(2) (0.5 microM, 29%)-mediated beta-hexosaminidase release. Thus, tc-LIGRLO-NH(2) and Cit have at least partially similar mechanisms of action as 48/80. PAR-AP may therefore activate mast cells via multiple mechanisms that are distinct from those of classical PAR-1 and PAR-2. The responsiveness of mast cells to PAR-AP via a non-PAR-1/non-PAR-2 mechanism complicates the interpretation of in vivo studies using these peptides. PMID- 12130702 TI - Delta(9)-tetrahydrocannabinol-induced apoptosis in the thymus and spleen as a mechanism of immunosuppression in vitro and in vivo. AB - Delta(9)-tetrahydrocannabinol (THC), the main psychoactive component of marijuana has been shown to suppress the immune response. However, the exact mechanism of THC-induced immunosuppression remains unclear. In the current study, we tested the hypothesis that exposure to THC leads to the induction of apoptosis in lymphocyte populations. Splenocytes of C57BL/6 mice cultured in the presence of 10 microM or greater concentrations of THC showed significantly reduced proliferative response to mitogens, including anti-CD3 monoclonal antibodies (mAbs), concanavalin A (Con A), and lipopolysaccharide (LPS) in vitro. Thymocytes and naive and activated splenocytes exposed to 10 microM or 20 microM THC showed significantly increased levels of apoptosis. Treatment with CB2 antagonist inhibited THC-induced apoptosis in thymocytes and activated splenocytes. Administration of 10 mg/kg body weight of THC into C57BL/6 mice led to thymic and splenic atrophy as early as 6 h after treatment. This effect could be partially inhibited by treatment with a caspase inhibitor in vivo. THC exposure led to reductions in the numbers of all subpopulations of splenocytes and thymocytes examined. Functional studies revealed that splenocytes from THC-treated mice had significantly reduced proliferative response to anti-CD3 mAbs, Con A, and LPS in vitro. Finally, thymocytes and splenocytes exposed to THC in vivo exhibited apoptosis upon in vitro culture. Together, these results suggest that in vivo exposure to THC can lead to significant suppression of the immune response by induction of apoptosis. PMID- 12130704 TI - Expression and induction of CYP2C P450 enzymes in primary cultures of human hepatocytes. AB - Although CYP2C8, CYP2C9, and CYP2C19 play an important role in drug biotransformation, factors influencing the expression and activity of these CYP2C P450s in human liver remain largely undefined. We used primary cultures of human hepatocytes from 15 subjects to assess the inducibility of CYP2C enzyme expression by prototypical inducer agents, including rifampicin, dexamethasone, and phenobarbital. After culture for 72 h in serum-free medium on collagen, Western blotting revealed that CYP2C9 was the only CYP2C enzyme expressed at appreciable levels in untreated hepatocytes. Subsequent treatment with 25 microMrifampicin for 48 h elicited marked increases in CYP2C8 (700 +/- 761%), CYP2C19 (854%), and CYP2C9 (209 +/- 176%) protein content versus a 550 +/- 170% enhancement of CYP3A4 enzyme levels. Parallel increases in CYP2C mRNAs, measured by Northern blotting and/or RNase protection, were found in rifampicin-treated hepatocytes, with CYP2C8, CYP2C9, and CYP2C19 transcripts exhibiting increases of 688 +/- 635, 207 +/- 49, and 230 +/- 60%, respectively, versus an 8.8-fold enhancement of CYP3A4 mRNA levels. Dexamethasone (10 microM) treatment enhanced CYP2C8 mRNA (360 +/- 100%) and protein (274%) content, although this steroid had less effect on CYP2C9 and CYP2C19 transcripts (23 +/- 21% and 21 +/- 36%, respectively) and enzyme levels (55 and 143%, respectively). Phenobarbital (100 microM) was a powerful inducer of CYP2C9 (850%) and CYP2C19 (735%) mRNA content, and also increased CYP2C8 (610%) and CYP3A4 (205%) transcripts. Our results show that CYP2C enzyme expression in human hepatocytes is highly inducible by rifampicin, dexamethasone, and phenobarbital. Because these xenobiotics are ligands and/or activators of the pregnane X receptor and/or constitutive androstane receptor, such orphan nuclear receptors and their response elements may partake in regulating CYP2C gene expression in humans. PMID- 12130705 TI - Gender difference in the urinary excretion of organic anions in rats. AB - This study was aimed at clarifying the gender differences in the urinary excretion of organic anions and the gene expression of organic anion transporters in rats. The renal clearance with regard to the plasma concentration (CL(urine,p)) of taurocholate, dibromosulfophthalein (DBSP), and zenarestat, all substrates and/or inhibitors of organic anion transporting polypeptide 1 (Oatp1), was much higher in female than in male rats. The following results imply that the transport system(s) for the reabsorption of zenarestat across the luminal side exhibits a gender difference: 1) the renal uptake clearance assessed by an in vivo integration plot analysis of zenarestat from the blood side does not show any clear gender differences; 2) the renal clearance with regard to the kidney concentration (CL(urine,k)) of zenarestat in female rats was approximately 30 times higher than in male rats; and 3) both CL(urine,p) and CL(urine,k) were increased in male rats by the coinfusion of DBSP, which is an inhibitor of organic anion transporters. Northern and Western blot analyses confirmed a previous finding that the gene expression of Oatp1, which is localized at the apical plasma membrane of the kidney, was much higher in the kidneys of male rats. Overall, a gender difference in urinary excretion is commonly observed for several organic anions, including Oatp1 substrates and inhibitors, and Oatp1 and/or transporters that have a similar substrate specificity to Oatp1 could be involved in such a phenomenon involving its substrates. PMID- 12130706 TI - Prevention of latently expressed CYP2C11, CYP3A2, and growth hormone defects in neonatally monosodium glutamate-treated male rats by the N-methyl-D-aspartate receptor antagonist dizocilpine maleate. AB - Neonatal administration of monosodium glutamate (MSG) can produce latently expressed defects in drug metabolism and growth hormone secretion as well as stunted growth and obesity. Instead of secreting growth hormone in the masculine episodic profile, plasma hormone levels are generally undetectable in affected adult male rats. Moreover, male-specific isoforms of cytochrome P450 (P450; e.g., CYP2C11 and CYP3A2), whose combined levels comprise the bulk of the total hepatic P450 in adult male rats, are similarly undetectable in these animals. Since "signaling elements" in the masculine episodic growth hormone profile are solely responsible for the elevated characteristic male-like expression levels of CYP2C11 and CYP3A2, suppression of the isoforms in the MSG-treated rats appeared to be caused by the simple absence of the hormone from the circulation. However, the reported failures of restored physiologic masculine growth hormone profiles to correct the P450 defects suggested the occurrence of direct MSG-induced liver damage independent of the well known hypothalamic lesions produced by the amino acid. Concurrent administration of dizocilpine maleate (MK-801), a selective and highly potent noncompetitive N-methyl-D-aspartate receptor antagonist of glutamate, completely prevented the adverse effects of neonatal MSG treatment on P450 expression, growth hormone secretion, and growth parameters, indicating that the amino acid-induced defects are solely a result of neuronal (i.e., hypothalamic) damage produced at the time of MSG exposure. The irreversibility of the P450 damage is described as resulting from secondary defects initially induced by the neuronal lesions. PMID- 12130707 TI - A single methamphetamine administration rapidly decreases vesicular dopamine uptake. AB - Recent studies demonstrated that vesicular dopamine (DA) uptake can be rapidly altered in synaptic vesicles purified from the striata of stimulant-treated rats. Specifically, a single administration of the plasmalemmal DA transporter inhibitor, cocaine, or the DA D(2) agonist, quinpirole, increases vesicular DA uptake in vesicles purified from the striata of treated rats. These effects of cocaine are prevented by pretreatment with a D(2), but not D(1), DA receptor antagonist. The purpose of the present study was to characterize the effect of a mechanistically different psychostimulant, methamphetamine (METH), on vesicular DA uptake. Results demonstrated that a single administration of this DA-releasing agent rapidly and reversibly decreased vesicular DA uptake. The METH-related decrease in vesicular DA uptake was attenuated by pretreatment with the D(2) antagonist, eticlopride, but not the D(1) antagonist, SCH23390 (R-[+]-7-chloro-8 hydroxy-3-methyl-1-phenyl-2,3,4,5-tetrahydro-1H-3-benzazepine). Core body temperature did not contribute to the effects of METH on vesicular DA uptake. Neither quinpirole nor cocaine increased vesicular DA uptake when rats were concurrently treated with METH. These studies provide further evidence that psychostimulants rapidly and differentially modify vesicular DA uptake. In addition, these studies demonstrate a complex role for D(2) DA receptors in altering vesicular DA transport. PMID- 12130709 TI - Involvement of organic cation transporter 1 in hepatic and intestinal distribution of metformin. AB - Metformin, a biguanide, is widely used as an oral hypoglycemic agent for the treatment of type 2 diabetes mellitus. The purpose of the present study was to investigate the role of organic cation transporter 1 (Oct1) in the disposition of metformin. Transfection of rat Oct1 cDNA results in the time-dependent and saturable uptake of metformin by the Chinese hamster ovary cell line with K(m) and V(max) values of 377 microM and 1386 pmol/min/mg of protein, respectively. Buformin and phenformin, two other biguanides, were also transported by rOct1 with a higher affinity than metformin: their K(m) values were 49 and 16 microM, respectively. To investigate the role of Oct1 in the disposition of metformin, the tissue distribution of metformin was determined in Oct1 gene-knockout mice after i.v. administration. Distribution of metformin to the liver in Oct1(-/-) mice was more than 30 times lower than that in Oct1(+/+) mice, and can be accounted for by the extracellular space. Distribution to the small intestine was also decreased in Oct1(-/-) mice, whereas that to the kidney as well as the urinary excretion profile showed only minimal differences. In conclusion, the present findings suggest that Oct1 is responsible for the hepatic uptake as well as playing a role in the intestinal uptake of metformin, whereas the renal distribution and excretion are mainly governed by other transport mechanism(s). PMID- 12130708 TI - Orphanin FQ/nociceptin-mediated desensitization of opioid receptor-like 1 receptor and mu opioid receptors involves protein kinase C: a molecular mechanism for heterologous cross-talk. AB - Morphine tolerance in vivo is reduced following blockade of the orphanin FQ/nociceptin (OFQ/N)/opioid receptor-like 1 (ORL1) receptor system, suggesting that OFQ/N contributes to the development of morphine tolerance. We previously reported that a 60-min activation of ORL1 receptors natively expressed in BE(2)-C cells desensitized both mu and ORL1 receptor-mediated inhibition of cAMP. Investigating the mechanism(s) of OFQ/N-mediated mu and ORL1 receptor cross-talk, we found that pretreatment with the protein kinase C inhibitor, chelerythrine chloride (1 microM), blocked OFQ/N-mediated homologous desensitization of ORL1 and heterologous desensitization of mu opioid receptors. Furthermore, depletion of PKC by 12-O-tetradecanoylphorbol-13-acetate exposure (48 h, 1 microM) also prevented OFQ/N-mediated mu and ORL1 desensitization. OFQ/N pretreatment resulted in translocation of PKC-alpha, G protein-coupled receptor kinase 2 (GRK2) and GRK3 from the cytosol to the membrane, and this translocation was also blocked by chelerythrine. Reduction of GRK2 and GRK3 levels by antisense, but not sense DNA treatment blocks ORL1 and mu receptor desensitization. This suggests that PKC alpha is required for GRK2 and GRK3 translocation to the membrane, where GRK can inactivate ORL1 and mu opioid receptors upon rechallenge with the appropriate agonist. Our results demonstrate for the first time the involvement of conventional PKC isozymes in OFQ/N-induced mu-ORL1 cross-talk, and represent a possible mechanism for OFQ/N-induced anti-opioid actions. PMID- 12130712 TI - Transport mechanisms of nicotine across the human intestinal epithelial cell line Caco-2. AB - Ulcerative colitis is a disease more commonly seen in nonsmokers. Because nicotine was postulated to be a beneficial component of tobacco smoke for ulcerative colitis, various formulations of nicotine have been developed to improve the local bioavailability within the gastrointestinal tissue. In the present study, to characterize the disposition of nicotine in the intestines, we investigated intestinal nicotine transport using Caco-2 cells. Nicotine was predominantly transported across Caco-2 cell monolayers in a unidirectional mode, corresponding to intestinal secretion, by pH-dependent specific transport systems. The specific uptake systems appear to be distinct from organic cation transporters and the transport system for tertiary amines, in terms of its substrate specificity and the pattern of the interaction. These transport systems could play a role in the intestinal accumulation of nicotine from plasma and could also be responsible for the topical delivery of nicotine for ulcerative colitis therapy. These findings could provide useful information for the design of effective nicotine delivery. PMID- 12130710 TI - Alcohol-induced c-Fos expression in the Edinger-Westphal nucleus: pharmacological and signal transduction mechanisms. AB - Mapping inducible transcription factors has shown that the Edinger-Westphal nucleus is preferentially sensitive to alcohol intoxication. Herein, we characterize the pharmacological and signal transduction mechanisms related to alcohol-induced c-Fos expression in Edinger-Westphal neurons. Using immunohistochemistry, we show that pretreatment with gamma-aminobutyric acid (GABA)-ergic antagonists (4 mg/kg bicuculline and 45 mg/kg pentylenetetrazole) attenuates induction of c-Fos expression by alcohol (2.4 g/kg, intraperitoneal). In addition, 10 mg/kg 2-(2,3-dihydro-2-methoxy-1,4-benzodioxin-2-yl)4,5-dihydro 1H-imidazole (RX 821002), an alpha(2A/D)-adrenoceptor antagonist, and 20 mg/kg haloperidol, a dopamine antagonist, also block alcohol-induced c-Fos expression in Edinger-Westphal neurons. No effects were seen in alcohol-induced c-Fos after the pretreatment of 20 mg/kg propranolol (beta-adrenoceptor antagonist), 10 mg/kg 2-(2-(4-(2-methoxyphenyl)piperazin-1-yl) ethy)-4,4-dimethyl-1,3-(2H,4H) isoquinolindione dihydrochloride (ARC 239) (alpha(2B/C)-adrenoceptor antagonist), or 30 mg/kg naltrexone (opioid antagonist). Although positive modulators for the GABA(A) receptor (20 mg/kg 3alpha-hydroxy-5alpha-pregnan-20-one and 10-30 mg/kg chlordiazepoxide) and opioid receptor (10 mg/kg morphine) produced significant elevations, agonists for alpha(2)-adrenoceptors (clonidine) and dopamine receptors (apomorphine) had no effect on Edinger-Westphal c-Fos expression. These findings suggest that alcohol-induced c-Fos expression in Edinger-Westphal results from direct interactions with GABA(A) receptors, which are modified by alpha(2A/D)-adrenoceptors and dopamine receptors. Also using immunohistochemistry to identify potential intracellular mechanisms associated with alcohol-induced c Fos expression in Edinger-Westphal, we show time-dependent increases in serine 727 phospho-signal transducer and activator of transcription 3 (Stat3) but no changes in phospho-cAMP response element-binding protein and phospho-Elk1. Time dependent increases in phospho-extracellular signal-regulated kinase (ERK) 1/2 were found to occur simultaneously with increases in serine 727 phospho-Stat3. Finally, blockade of ERK 1/2 phosphorylation with the mitogen-activated protein kinase (MEK) 1/2 inhibitor SL327 blocked alcohol-induced c-Fos expression, suggesting that alcohol induces c-Fos in Edinger-Westphal neurons through activation of the MEK1/2-ERK1/2-Stat3 pathway. PMID- 12130711 TI - Gender differences in hypothalamic tyrosine hydroxylase and alpha(2)-adrenoceptor subtype gene expression in cafeteria diet-induced hypertension and consequences of neonatal androgenization. AB - This study investigated the incidence of cafeteria-diet induced hypertension on hypothalamic tyrosine hydroxylase (TH) and alpha(2)-adrenoceptor subtype gene expression in male, female, and neonatally testosterone-imprinted female rats. After 10 weeks of cafeteria diet, all these rats were hyperleptinemic. In contrast, males and testosterone-treated females developed hypertension, whereas intact females remained normotensive. In these rats, cafeteria diet up-regulated TH gene expression only in males and testosterone-treated females. On the other hand, cafeteria diet differentially affected hypothalamic gene expression of alpha(2)-adrenoceptor subtypes. In fact, this diet increased alpha(2A) adrenoceptor mRNA levels only in intact normotensive females. In contrast, gene expression of the alpha(2B)-adrenoceptor was up-regulated only in male and testosterone-treated female cafeteria-fed rats. Furthermore, an alpha(2C) adrenoceptor gene over-expression was also induced, but only in male cafeteria fed rats. If one assumes that the up-regulations in TH and alpha(2B)-adrenoceptor gene expression are indicative of increased sympathetic nervous activity, then, these altered gene expressions could be responsible for the maintenance of high blood pressure in male and testosterone-treated female cafeteria-fed rats. Conversely, in intact females, the absence of these over-expressions and the up regulation of the alpha(2A)-adrenoceptor gene expression could reflect an adaptive response to the diet and, consequently, could be protective against cafeteria diet-induced hypertension. Moreover, neonatal testosterone imprinting in females could have induced an irreversible android susceptibility to the cafeteria diet, leading to the onset of hypertension. PMID- 12130713 TI - Ischemia promotes renin activation and angiotensin formation in sympathetic nerve terminals isolated from the human heart: contribution to carrier-mediated norepinephrine release. AB - We recently reported that in the ischemic human heart, locally formed angiotensin II activates angiotensin II type 1 (AT(1)) receptors on sympathetic nerve terminals, promoting reversal of the norepinephrine transporter in an outward direction (i.e., carrier-mediated norepinephrine release). The purpose of this study was to assess whether cardiac sympathetic nerve endings contribute to local angiotensin II formation, in addition to being a target of angiotensin II. To this end, we isolated sympathetic nerve endings (cardiac synaptosomes) from surgical specimens of human right atrium and incubated them in ischemic conditions (95% N(2,) sodium dithionite, and no glucose for 70 min). These synaptosomes released large amounts of endogenous norepinephrine via a carrier mediated mechanism, as evidenced by the inhibitory effect of desipramine on this process. Norepinephrine release was further enhanced by preincubation of synaptosomes with angiotensinogen and was prevented by two renin inhibitors, pepstatin-A and BILA 2157BS, as well as by the angiotensin-converting enzyme inhibitor enalaprilat and the AT(1) receptor antagonist EXP 3174 [2-N-butyl-4 chloro-1-[2'-(1H-tetrazol-5-yl)biphenyl-4-yl] methyl]imidazole-5-carboxylic acid]. Western blot analysis revealed the presence of renin in cardiac sympathetic nerve terminals; renin abundance increased ~3-fold during ischemia. Thus, renin is rapidly activated during ischemia in cardiac sympathetic nerve terminals, and this process eventually culminates in angiotensin II formation, stimulation of AT(1) receptors, and carrier-mediated norepinephrine release. Our findings uncover a novel autocrine/paracrine mechanism whereby angiotensin II, formed at adrenergic nerve endings in myocardial ischemia, elicits carrier mediated norepinephrine release by activating adjacent AT(1) receptors. PMID- 12130715 TI - The effect of chronic ethanol consumption and withdrawal on mu-opioid and dopamine D(1) and D(2) receptor density in Fawn-Hooded rat brain. AB - Previous studies have implicated the dopamine and opioid systems in the induction and maintenance of ethanol consumption. This study investigated, in alcohol preferring Fawn-Hooded (FH) rats, whether chronic free-choice ethanol consumption and subsequent withdrawal cause alterations in central mu-opioid, dopamine D(1), and D(2) receptor density using autoradiography. FH rats were given a free choice between a 5% ethanol solution and tap water (n = 25) and displayed a mean ethanol consumption of 5.6 g/kg/day. A parallel group of FH rats (n = 5) only had access to tap water. Rats were then withdrawn from ethanol for 0, 1, 2, 5, or 10 days and killed by cervical dislocation and decapitation. Increases in mu-opioid receptor density were observed in the nucleus accumbens and ventral tegmental area upon withdrawal compared with the ethanol naive group. In the lateral amygdala, binding in all withdrawal groups was significantly different from the ethanol naive FH rats, and also from the chronic ethanol rats. An increase in dopamine D(1) receptor density was observed in the substantia nigra, pars reticulata in the 5- and 10-day withdrawal groups compared with ethanol naive. Accumbal dopamine D(2) receptor density (+25-30%) increased in the 10-day withdrawal group compared with both naive and chronic ethanol groups. These findings demonstrate that the opioid and dopamine systems are susceptible to modulation by chronic ethanol consumption and withdrawal in the FH rat. Furthermore, although acute ethanol withdrawal results in modulation of mu-opioid receptors, effects on dopamine receptors are delayed and only become evident 5 to 10 days after withdrawal. PMID- 12130716 TI - Modulation of neuronal nicotinic acetylcholine receptors by mercury. AB - Mercuric chloride exerted a biphasic modulatory effect on rat neuronal nicotinic acetylcholine receptors (nAChRs) expressed in Xenopus laevis oocytes as heteromers of the alpha3 or alpha4 and beta2 or beta4 subunits. The degree of modulation was subunit-dependent, with beta4-containing receptors displaying greater potentiation and alpha4-containing receptors displaying greater inhibition. Thus, alpha4beta4 receptors displayed both robust potentiation and robust inhibition. During prolonged coapplication of HgCl(2), first potentiation then inhibition of the acetylcholine (ACh) response was observed. Upon coapplication of 1 microM HgCl(2), a 2-fold increase in ACh-induced current was achieved in 55 +/- 1 s. With continued HgCl(2) application, the ACh response was slowly inhibited until, after 5 min, less than 10% of the initial response remained. By measuring potentiation at its peak and inhibition 5 min after the start of HgCl(2) coapplication, we obtained EC(50) and IC(50) values of 262 +/- 75 and 430 +/- 72 nM, respectively. HgCl(2) potentiation was voltage-dependent, increasing at more positive holding potentials. Upon washout of mercury chloride, potentiation reversed with a t(1/2) of 4.6 min. Inhibition reversed more slowly, with less than half the initial response recovered after 15 min of wash. Although free cysteine residues are common targets for mercury, elimination of all free cysteines located in the extracellular domains of the alpha4 and beta4 subunits did not alter the effects of mercuric chloride. Potentiation and inhibition of neuronal nAChRs may occur through action at a transmembrane or cytoplasmic location after passive diffusion of mercuric chloride across the plasma membrane. PMID- 12130714 TI - Modulation of the human Kv1.5 channel by protein kinase C activation: role of the Kvbeta1.2 subunit. AB - Kv1.5 is the principal molecular component of I(Kur), an atrial-specific K(+) current in human myocytes that is suppressed by activation of protein kinase C (PKC). We examined the effect of phorbol 12-myristate 13-acetate (PMA), a direct activator of PKC, on Kv1.5 current. Although PMA had minimal effect when Kv1.5 was expressed alone, K(+) currents derived from coexpression of Kvbeta1.2 (but not another closely related beta subunit, Kvbeta1.3) with Kv1.5 were markedly reduced by PMA, associated with a small depolarizing shift in the voltage dependence of channel activation. Additional experiments with an inactive stereoisomer, 4alpha-PMA, and the PKC inhibitor chelerythrine indicated that the effects of PMA were mediated by PKC activation. Assembly of Kv1.5 in vivo with both beta subunits was demonstrated, and all three K(+) channel proteins were substrates for phosphorylation by PKC. These results demonstrate that coexpression of Kvbeta1.2 enhances the response of Kv1.5 to PKC activation and that direct phosphorylation of K(+) channel subunits is a potential molecular basis for the effect. Furthermore, they suggest that Kvbeta1.2 may be a component of the I(Kur) complex in human atrium. PMID- 12130717 TI - Morphine-3beta-D-glucuronide suppresses inhibitory synaptic transmission in rat substantia gelatinosa. AB - High doses of intrathecally applied morphine or morphine-3beta-D-glucuronide (M3G) produce allodynia and hyperalgesia. Whole-cell patch-clamp recordings were made from substantia gelatinosa neurons in transverse slices of adult rat lumbar spinal cord to compare the actions of M3G with those of the mu-opioid agonist, DAMGO ([D-Ala(2),N-Met-Phe(4),Gly-ol(5)]-enkephalin), and the ORL(1) agonist, nociceptin/orphanin FQ (N/OFQ). M3G (1-100 microM) had little or no effect on evoked excitatory postsynaptic currents (EPSC) and no effect on postsynaptic membrane conductance. In contrast, 1 microM DAMGO and 1 microM N/OFQ reduced the amplitude of evoked EPSCs and activated an inwardly rectifying K(+) conductance. M3G did not attenuate the effect of DAMGO or N/OFQ on evoked EPSC amplitude. However, 1 to 100 microM M3G reduced the amplitude of evoked GABAergic and glycinergic inhibitory postsynaptic current (IPSC) by up to 48%. This effect was naloxone-insensitive. The evoked IPSC was also attenuated by DAMGO, but not by N/OFQ. Because M3G reduced the frequency of tetrodotoxin-insensitive miniature IPSCs and increased paired-pulse facilitation, it appeared to act presynaptically to disinhibit substantia gelatinosa neurons. This effect, which does not appear to involve mu-opioid or ORL(1) receptors, may contribute to the allodynia and hyperalgesia observed after intrathecal application of high doses of morphine. PMID- 12130718 TI - Digoxin uptake, receptor heterogeneity, and inotropic response in the isolated rat heart: a comprehensive kinetic model. AB - The cardiac pharmacokinetics of digitalis glycosides is not well understood. In the present study, a mechanism-based pharmacokinetic/pharmacodynamic model was developed to describe the uptake kinetics, receptor interaction, and positive inotropic effect of digoxin in the single-pass isolated perfused rat heart. Three doses of digoxin (0.1, 0.2, and 0.3 micromol) were administered to the heart (n = 12) as consecutive 1-min infusions followed by 15-min washout periods. Outflow concentration and left ventricular developed pressure were measured and analyzed by the model. The uptake of digoxin by the heart was limited by capillary permeability with a permeation clearance of 2.35 ml/min/g (about one-third of perfusate flow). Binding kinetics was determined by a mixture of two receptor subtypes, a low-affinity/high-capacity binding site (K(D,1) = 20.9 nmol, 89% of total receptors) and a high-affinity/low-capacity binding site (K(D,2) = 1.5 nmol, 11%). The time course of inotropic response was linked to receptor occupation, with higher efficiency of the high-affinity receptor population. The results suggest that, in the rat heart, consecutive inhibition of first the alpha(2)- and then the alpha(1)-isoform of Na(+)/K(+)-ATPase mediates the positive inotropic effect of digoxin with increasing dosage. PMID- 12130719 TI - Persistent suppression of hepatic CYP2A1 expression and serum triiodothyronine levels by tamoxifen in intact female rats: dose-response analysis and comparison with 4-hydroxytamoxifen, fulvestrant (ICI 182,780), and 17beta-estradiol-3 benzoate. AB - Tamoxifen, a nonsteroidal antiestrogen, is used widely in the treatment of breast cancer and is undergoing evaluation as a chemopreventive agent. In this study, we investigated several long-term effects of tamoxifen in intact adult female rats following acute treatment at various dosages. The effects of tamoxifen on somatic growth, growth hormone (GH) levels, thyroid hormone levels, and on hepatic cytochrome P450 (P450) expression were compared with those of fulvestrant (ICI 182,780), 17beta-estradiol-3-benzoate, and 4-hydroxytamoxifen under the same experimental conditions. Each compound was injected s.c. for two consecutive days, and rats were killed 37 days after treatment. Tamoxifen decreased body weight and serum triiodothyronine (T3) levels at dosages ranging from 0.5 to 200 mg/kg. Ovary weight, uterus weight, peak plasma GH concentration, and hepatic CYP2A1 content were decreased 37 days after treatment with tamoxifen at a dosage of 20 mg/kg, but expression of other P450 enzymes was not affected. However, tamoxifen and 4-hydroxytamoxifen could not be detected in plasma by high performance liquid chromatography analysis at this time, which suggests that the effects of tamoxifen were mediated indirectly. 4-Hydroxytamoxifen exhibited effects similar to those of tamoxifen, indicating that this metabolite contributes to the in vivo activity of tamoxifen. Estradiol benzoate decreased CYP2A1 and increased CYP3A hepatic levels, but had no effect on serum T3 concentration. In contrast, treatment with ICI 182,780 had little or no effect on the endpoints measured. In summary, 2-day tamoxifen treatment of intact adult female rats resulted in persistent suppression of somatic growth, serum T3 levels, and hepatic CYP2A1 expression. PMID- 12130720 TI - The concordance of early antipyrine and thiopental distribution kinetics. AB - Studies of factors affecting the initial disposition of drugs with a rapid onset of effect following i.v. administration have used antipyrine as a surrogate for lipophilic drugs because it lacks cardiovascular effects. The present study tested the assumption that antipyrine is a useful surrogate for the flow dependent tissue distribution of the lipophilic drug thiopental by comparing the recirculatory pharmacokinetic models of antipyrine and thiopental disposition after concomitant administration to five dogs anesthetized with 1.5% halothane. The pharmacokinetics of indocyanine green, a marker of the intravascular behavior of antipyrine and thiopental, and antipyrine in these dogs was nearly identical to that described previously in dogs anesthetized with 1.5% halothane but not given thiopental. The total volume of distribution of the highly lipophilic drug thiopental was more than 60% larger than that of antipyrine, 53 versus 33 liters, respectively. Nonetheless, the initial distribution kinetics of the two drugs, including the pulmonary tissue volume and the volume of the nondistributive pathway as well as the clearance to it, were nearly identical. As a result, the fraction of cardiac output involved in distribution of the two drugs to peripheral tissues was similarly identical, although the distribution of cardiac output between clearance to the rapidly equilibrating tissues and clearance to the slowly equilibrating tissues differed slightly. This study validates the assumption that antipyrine is a useful surrogate for lipophilic drugs in pharmacokinetic studies in which physiologic stability is desirable to meet the assumption of system stationarity. PMID- 12130721 TI - Statins modulate oxidized low-density lipoprotein-mediated adhesion molecule expression in human coronary artery endothelial cells: role of LOX-1. AB - LOX-1, a receptor for oxidized low-density lipoprotein (ox-LDL), plays a critical role in endothelial dysfunction and atherosclerosis. LOX-1 activation also plays an important role in monocyte adhesion to endothelial cells. A number of studies show that 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitors (statins) reduce total LDL cholesterol and exert a cardioprotective effect. We examined the modulation of LOX-1 expression and its function by two different statins, simvastatin and atorvastatin, in human coronary artery endothelial cells (HCAECs). We observed that ox-LDL (40 microg/ml) treatment up-regulated the expression of E- and P-selectins, VCAM-1 and ICAM-1 in HCAECs. Ox-LDL mediated these effects via LOX-1, since antisense to LOX-1 mRNA decreased LOX-1 expression and subsequent adhesion molecule expression. Pretreatment of HCAECs with simvastatin or atorvastatin (1 and 10 microM) reduced ox-LDL-induced expression of LOX-1 as well as adhesion molecules (all P < 0.05). A high concentration of statins (10 microM) was more potent than the low concentration (1 microM) (P < 0.05). Both statins reduced ox-LDL-mediated activation of the redox-sensitive nuclear factor-kappaB (NF-kappaB) but not AP-1. These observations indicate that LOX-1 activation plays an important role in ox-LDL-induced expression of adhesion molecules. Inhibition of expression of LOX-1 and adhesion molecules and activation of NF-kappaB may be another mechanism of beneficial effects of statins in vascular diseases. PMID- 12130722 TI - Effects of antidepressants in rats trained to discriminate centrally administered isoproterenol. AB - Previous work has shown that the discriminative stimulus effects of centrally administered isoproterenol are mediated primarily via beta1-adrenergic receptors. In the present study, this model was used to investigate the ability of antidepressant drugs displaying various pharmacological profiles to stimulate beta1-adrenergic receptors in vivo; this was assessed by determining whether they substituted for the discriminative stimulus effects of isoproterenol. Rats were trained to discriminate centrally administered isoproterenol (10 microg i.c.v.) from artificial cerebral spinal fluid using a water-reinforced, two-lever operant task (fixed ratio 10 schedule). After acquisition of the discrimination, drugs were tested for substitution (i.p.). The tricyclic antidepressants protriptyline and desipramine, the norepinephrine uptake inhibitor nisoxetine, the monoamine oxidase inhibitor phenelzine, and the atypical antidepressants bupropion, mirtazapine, and venlafaxine all produced greater than 90% isoproterenol appropriate responding. The serotonin uptake inhibitor fluoxetine, the atypical antidepressants buspirone and trazodone, and the novel, putative antidepressants N(G)-nitro-L-arginine and N-acetyl-L-tryptophan 3,5-bis benzyl ester failed to substitute for isoproterenol at the dose ranges tested. Antagonism studies carried out with betaxolol for those drugs that fully generalized to isoproterenol's cue verified mediation by beta1-adrenergic receptors. The present results indicate that drugs with noradrenergic activity generalize to isoproterenol's discriminative stimulus. Although this suggests a role for central beta1-adrenergic receptors in the mechanism of action of certain antidepressant drugs, it does not seem that stimulation of these receptors is an effect shared by antidepressants from all pharmacological classes. PMID- 12130723 TI - Molecular actions of (S)-desmethylzopiclone (SEP-174559), an anxiolytic metabolite of zopiclone. AB - This study set out to profile the activity of (S)-desmethylzopiclone (SEP-174559) at subtypes of the gamma-aminobutyric acid type-A (GABA(A)) receptor and other neurotransmitter receptor ion channels. Recombinant receptors were expressed in human embryonic kidney 293 cells and examined functionally by patch-clamp recording with fast perfusion of agonist and drug solutions. Micromolar concentrations of SEP-174559 potentiated GABA(A) receptor currents evoked by subsaturating concentrations of GABA. The potentiation was related to a leftward shift in the GABA dose-response curves, suggesting the drug acts to increase GABA binding affinity. The potentiation strictly required the presence of the gamma2 subunit; no enhancement was seen for receptors containing instead the gamma1 subunit or lacking a gamma subunit altogether. SEP-174559 and its parent compound, racemic zopiclone, were not selective between alpha1-, alpha2-, or alpha3-bearing GABA(A) receptors. Within the nicotinic receptor superfamily, SEP 174559 did not affect serotonin type-3 receptor function but was found to inhibit nicotinic acetylcholine (nACh) receptors. The inhibition of nACh receptors was noncompetitive and was mimicked by zopiclone, alprazolam, and diazepam. In the glutamate receptor superfamily, SEP-174559 inhibited N-methyl-D-aspartate (NMDA) receptor currents but did not affect non-NMDA receptors. These data confirm that SEP-174559 has benzodiazepine-like actions at gamma2-bearing subtypes of the GABA(A) receptor and suggest additional actions of benzodiazepine-site ligands at nACh and NMDA receptors. PMID- 12130724 TI - bis-Cholesteryl-conjugated phosphorothioate oligodeoxynucleotides are highly selectively taken up by the liver. AB - We previously modulated, by conjugating a single cholesterol, plasma protein binding and liver cell uptake of a phosphorothioate oligodeoxynucleotide (PS ODN). In this study, we investigated the biological fate of a PS-ODN, denoted ISIS-9389 (3',5'-bis-cholesteryl-conjugated ISIS 3082), provided with two cholesteryl moieties. After intravenous injection of into rats, [(3)H]ISIS-9389 was cleared from plasma with a half-life of 23.6 +/- 0.3 min. After 90 min (approximately 95% cleared), the liver contained 83.0 +/- 0.8% of the dose. Spleen and bone (marrow), which constitute with the liver the reticuloendothelial system, contained 3.1 +/- 0.3 and 4.3 +/- 0.2%, respectively. All other tissues accumulated together <5% of the dose. The hepatic uptake of [(3)H]ISIS-9389 occurred mainly by endothelial cells (51.9 +/- 6.4% of the liver uptake). Parenchymal and Kupffer cells were responsible for 24.9 +/- 7.7 and 23.3 +/- 2.5%, respectively. Preinjected polyinosinic acid and polyadenylic acid reduced hepatic uptake, albeit the latter was less effective. This finding suggests implication of (multiple) scavenger receptors in liver uptake of ISIS-9389. The interaction of ISIS-9389 with plasma proteins, analyzed by size exclusion chromatography, differs from that of unconjugated PS-ODN and PS-ODN with a single cholesterol. Plasma-incubated ISIS-9389 was mainly recovered as a high molecular weight complex. In conclusion, conjugation of PS-ODNs with two cholesteryl moieties results in almost quantitative uptake by the liver. The liver targeting exceeds the already impressive gain in liver uptake achieved by conjugation of a single cholesterol, and is expected to increase the therapeutic activity against liver-associated targets and reduce side effects in nonhepatic tissues. PMID- 12130725 TI - Roles of substance P receptors in human colon circular muscle: alterations in diverticular disease. AB - The characteristics of [(125)I]Bolton-Hunter[Sar(9),Met(O(2))(11)]substance P ([(125)I]BH-SarSP) binding were investigated in membranes of human ascending, transverse, distal, and sigmoid colon circular muscle. Binding of [(125)I]BH SarSP was of high affinity (K(D) = 68 nM) and low capacity (B(max) = 0.31 fmol/mg of wet weight tissue), and showed no regional differences. [(125)I]BH-SarSP binding was inhibited by SP approximately equal to [Pro(9)]SP > or = (2S,3S)-3-(2 methoxybenzylamino)-2-phenylpiperidine (CP99994) >> neurokinin (NK) A > or = neuropeptide gamma > [Lys(5),MeLeu(9),Nle(10)]-NKA(4-10) approximately (S)-N methyl-N[4-acetylamino-4-phenylpiperidino)-2-(3,4-dichlorophenyl) butyl]benzamide (SR48968) >> senktide, suggesting binding to NK-1 sites. Most agonists seemed to bind to two sites. In autoradiographic studies, dense binding for [(125)I]BH SarSP was associated with submucosal and longitudinal muscle blood vessels, and the submucosal margin of circular muscle (corresponding to interstitial cells of Cajal), with moderate binding over most of the circular muscle. In normal colon circular muscle strips, [Pro(9)]SP was almost ineffective, and SP caused contractions with pD(2) values of 5.3 to 5.7. No regional differences were observed in potency or efficacy. Responses to SP were inhibited by the NK-2 receptor antagonist SR48968, but not by NK-1 antagonist CP99994, indicating the involvement of NK-2 rather than NK-1 receptors. Atropine significantly inhibited contractions induced by SP, indicating a minor cholinergic component. Contractile responses to SP were considerably reduced in preparations from patients with diverticular disease, and marginally reduced in ulcerative colitis compared with control. This study clearly demonstrates an NK-1 binding site on human colon circular muscle, but its role in this tissue remains unclear and may not involve contractile mechanisms. The attenuated contractility in specimens with diverticular disease may reflect disease-related alterations of the tachykinin receptor system. PMID- 12130727 TI - Piperine, a major constituent of black pepper, inhibits human P-glycoprotein and CYP3A4. AB - Dietary constituents (e.g., in grapefruit juice; NaCl) and phytochemicals (e.g., St. John's wort) are important agents modifying drug metabolism and transport and thereby contribute to interindividual variability in drug disposition. Most of these drug-food interactions are due to induction or inhibition of P-glycoprotein and/or CYP3A4. Preliminary data indicate that piperine, a major component of black pepper, inhibits drug-metabolizing enzymes in rodents and increases plasma concentrations of several drugs, including P-glycoprotein substrates (phenytoin and rifampin) in humans. However, there are no direct data whether piperine is an inhibitor of human P-glycoprotein and/or CYP3A4. We therefore investigated the influence of piperine on P-glycoprotein-mediated, polarized transport of digoxin and cyclosporine in monolayers of Caco-2 cells. Moreover, by using human liver microsomes we determined the effect of piperine on CYP3A4-mediated formation of the verapamil metabolites D-617 and norverapamil. Piperine inhibited digoxin and cyclosporine A transport in Caco-2 cells with IC(50) values of 15.5 and 74.1 microM, respectively. CYP3A4-catalyzed formation of D-617 and norverapamil was inhibited in a mixed fashion, with K(i) values of 36 +/- 8 (liver 1)/49 +/- 6 (liver 2) and 44 +/- 10 (liver 1)/77 +/- 10 microM (liver 2), respectively. In summary, we showed that piperine inhibits both the drug transporter P glycoprotein and the major drug-metabolizing enzyme CYP3A4. Because both proteins are expressed in enterocytes and hepatocytes and contribute to a major extent to first-pass elimination of many drugs, our data indicate that dietary piperine could affect plasma concentrations of P-glycoprotein and CYP3A4 substrates in humans, in particular if these drugs are administered orally. PMID- 12130726 TI - Correlation between anticonvulsant activity and inhibitory action on glial gamma aminobutyric acid uptake of the highly selective mouse gamma-aminobutyric acid transporter 1 inhibitor 3-hydroxy-4-amino-4,5,6,7-tetrahydro-1,2-benzisoxazole and its N-alkylated analogs. AB - The inhibitory effect of 3-hydroxy-4-amino-4,5,6,7-tetrahydro-1,2-benzisoxazole (exo-THPO) and its N-methylated (N-methyl-exo-THPO) and N-ethylated (N-ethyl-exo THPO) analogs, derived from gamma-aminobutyric acid (GABA) and 4,5,6,7 tetrahydroisoxazolo[4,5-c]pyridin-3-ol (THPO) on GABA transport was investigated using cultured neocortical neurons (GABA-ergic) and astrocytes and cloned mouse GABA transporters GAT1-4 expressed in human embryonic kidney (HEK) 293 cells. Anticonvulsant activity was assessed after i.c.v. administration to Frings audiogenic seizure-susceptible mice. Anticonvulsant activity of the O pivaloyloxymethyl prodrug of N-methyl-exo-THPO was assessed after i.p. administration. Results from these studies were compared with those obtained from similar studies with the novel anticonvulsant drug tiagabine, which acts via inhibition of GABA transport. exo-THPO and its N-alkyl analogs inhibited neuronal, astrocytic, and GAT1-mediated GABA transport but not GABA uptake mediated by GAT2-4. N-Methyl-exo-THPO was 8-fold more potent as an inhibitor of astrocytic versus neuronal GABA uptake. The IC(50) value for inhibition of GABA uptake by GAT1 closely reflected its IC(50) value for inhibition of neuronal uptake. Tiagabine was approximately 1000-fold more potent than exo-THPO and its alkyl derivatives as an inhibitor of GABA uptake in cultured neural cells and GAT1-expressing HEK 293 cells. exo-THPO, its alkylated analogs, and tiagabine displayed a time- and dose-dependent inhibition of audiogenic seizures after i.c.v. administration. N-Methyl-exo-THPO was the most potent anticonvulsant among the exo-THPO compounds tested and only slightly less potent than tiagabine. The findings suggest a correlation between anticonvulsant efficacy and selective inhibition of astroglial GABA uptake. Furthermore, results obtained with the N methyl-exo-THPO prodrug demonstrate the feasibility of developing a glial selective GABA uptake inhibitor with systemic bioavailability. PMID- 12130728 TI - Inhibition of human and pig ureter motility in vitro and in vivo by the K(+) channel openers PKF 217-744b and nicorandil. AB - The relaxing property of the K(+) channel opener and nitric oxide donor nicorandil and the new K(+) channel opener PKF 217-744b was investigated on isolated human ureteral tissue in vitro and in intact ureters of anesthetized pigs in vivo. In addition, nicorandil and its antagonists, glibenclamide and methylene blue, were tested on isolated pig ureter tissue in vitro. Nicorandil decreased the frequency of spontaneous contractions in isolated pig ureter rings. This effect was antagonized by glibenclamide and methylene blue suggesting that the nicorandil induced relaxation of the ureter is mediated by activation of ATP sensitive K(+) channels and involvement of soluble guanylate cyclase. Moreover, nicorandil and PKF 217-744b reduced the amplitude of electrically induced contractions in isolated human ureter rings. Calculations of EC(50) values showed that PKF 217-744b [EC(50) = 4.83 x 10(-8) M] was more potent than nicorandil [EC(50) = 4.38 x 10(-5) M]. Both drugs reduced the contraction frequency of the pig ureter after intravenous and topical administration in vivo. Intravenous, but not topical, administration of nicorandil and PKF 217-744b significantly decreased arterial blood pressure but did not affect the heart rate. The in vitro findings suggest that K(+) channel opening and nitric oxide release mediate the effect of nicorandil. Our in vivo results indicate that PKF 217-744b and nicorandil are promising drugs for clinical application in patients with acute stone colic to relieve obstruction and facilitate stone passage or to relax the ureter before ureteroscopy. PMID- 12130729 TI - Antinerve growth factor treatment prevents intestinal dysmotility in Trichinella spiralis-infected rats. AB - Nerve growth factor (NGF) could be involved in the development of hyperalgesia as well as in nervous remodeling consequence of inflammation. Both dysmotility and increase of visceral sensitivity have been described in functional gastrointestinal disorders such as irritable bowel syndrome. Trichinella spiralis infected rats show an exacerbated spontaneous motility and a significant increase of the excitatory response to cholecystokinin (CCK), both associated with a reversible inflammatory process and the hypertrophy of the muscle layers. In this study we determined the intestinal expression of NGF mRNA by polymerase chain reaction and NGF by enzyme-linked immunosorbent assay. We implanted serosal strain gauge transducers on duodenum, jejunum, and ileum of anesthetized Sprague Dawley rats to record circular muscle contractions. The experimental protocol included the evaluation of intestinal spontaneous motor activity (SMA), the response to CCK-8, and the ascending contraction induced by electrical mucosal stimulation. This protocol was performed in healthy and infected nontreated rats, in healthy rats with an NGF antibody treatment (1.6 mg/rat i.p.), and in infected rats with the same treatment applied at 0 or 3 days postinfection. NGF and NGF mRNA levels in the bowel were increased during inflammation. Although anti-NGF treatments did not prevent or reverse inflammatory response, the treatment was effective in preventing the motor alterations induced by the T. spiralis infection, i.e., inhibited increased SMA, reversed altered response to CCK, and reversed in part exacerbated response to electrical stimulation. PMID- 12130730 TI - Characterization of methotrexate transport and its drug interactions with human organic anion transporters. AB - Life-threatening drug interactions are known to occur between methotrexate and nonsteroidal anti-inflammatory drugs (NSAIDs), probenecid, and penicillin G. The purpose of this study was to characterize methotrexate transport, as well as to determine the site and the mechanism of drug interactions in the proximal tubule. Mouse proximal tubule cells stably expressing basolateral human organic anion transporters (hOAT1 and hOAT3) and apical hOAT (hOAT4) were established. The K(m) values for hOAT1-, hOAT3-, and hOAT4-mediated methotrexate uptake were 553.8 microM, 21.1 microM, and 17.8 microM, respectively. NSAIDs (salicylate, ibuprofen, ketoprofen, phenylbutazone, piroxicam, and indomethacin), probenecid, and penicillin G dose dependently inhibited methotrexate uptake mediated by hOAT1, hOAT3, and hOAT4. Kinetic analysis of inhibitory effects of these drugs on hOAT3-mediated methotrexate uptake revealed that these inhibitions were competitive. The K(i) values for the effects of salicylate, phenylbutazone, indomethacin, and probenecid on hOAT3-mediated methotrexate uptake were comparable with therapeutically relevant plasma concentrations of unbound drugs. In addition, in the presence of human serum albumin, the K(i) values were comparable with therapeutically relevant total plasma concentrations of drugs. In conclusion, these results suggest that methotrexate is taken up via hOAT3 and hOAT1 at the basolateral side of the proximal tubule and effluxed or taken up at the apical side via hOAT4. In addition, hOAT1, hOAT3, and hOAT4 are the sites of drug interactions between methotrexate and NSAIDs, probenecid, and penicillin G. Furthermore, it was predicted that hOAT3 is the site of drug interactions between methotrexate and salicylate, phenylbutazone, indomethacin, and probenecid in vivo. PMID- 12130731 TI - The endothelin A receptor antagonists PD 156707 (CI-1020) and PD 180988 (CI-1034) reverse the hypoxic pulmonary vasoconstriction in the perinatal lamb. AB - Endothelin-1 (ET-1) is considered an intermediary in the constrictor response of the pulmonary vasculature to hypoxia and, by extension, is assigned a prime role in the pathogenesis of pulmonary hypertension. We report here the antihypertensive action in the conscious newborn lamb of two novel endothelin A receptor antagonists, sodium 2-benzo-[1,3]dioxol-5-yl-4- (4-methoxy-phenyl)-4-oxo 3-(3,4,5-trimethoxy-benzyl)-but-2- enoate (PD 156707) and 4-(7-ethyl benzo[1,3]dioxol-5-yl)-1, 1-dioxo-2-(2-trifluoromethyl-phenyl)-1,2-dihydro-1l6 benzo-[e][1,2]thiazine-3-carboxylic acid potassium (PD 180988), differing in chemical properties and half-life within the body. PD 156707 and PD 180988, given in the right atrium as a bolus followed by infusion, had little or no effect on pulmonary and systemic hemodynamics under normoxia. Conversely, they both reversed the pulmonary hypertension due to alveolar hypoxia while producing minor changes, or no change at all, in systemic vascular resistance. Furthermore, their pulmonary vascular effect outlasted administration. Pulmonary hypertension being elicited by infusion of the thromboxane A(2) analog, 9,11-epithio-11,12-methano thromboxane A(2) (ONO-11113) was instead not amenable to ET(A)R inhibition. Blood levels of ET-1, which rose with hypoxia but not ONO-11113 treatment, were not changed by either antagonist. Consistent with findings in vivo, when using isolated pulmonary resistance arteries from term fetal lamb, PD 156707 curtailed the hypoxia- but not the ONO-11113-induced constriction. We conclude that PD 156707 and PD 180988 are selective inhibitors of pulmonary vasoconstriction resulting from hypoxia. Our findings support the use of these or allied compounds in the management of pulmonary hypertension in the neonate. PMID- 12130732 TI - Effects of blood flow on the in vivo recovery of a small diffusible molecule by microdialysis in human skin. AB - The aim of this study was to investigate the impact of changes in local blood flow on the recovery of a small, diffusible molecule (sodium fluorescein) from the extravascular tissue space of the skin, by microdialysis in vivo. Loss and recovery of fluorescein by linear microdialysis probes (5-kDa molecular mass cutoff, 0.2 mm diameter) inserted 1 mm apart in pairs, at three sites in the skin of the volar surface of the forearm of healthy volunteers, was measured under conditions of basal, reduced (noradrenaline, 0.005 mg/ml), and increased (glyceryl trinitrate, patch) blood flow. Whereas loss of tracer from the delivery probe appeared unaffected by changes in local blood flow, retrieval of fluorescein by the second probe was directly related to blood flux, measured using scanning laser Doppler imaging. Steady-state recovery at vasoconstricted sites was 4.0 +/- 0.7 microg. ml(-1) compared with 1.8 +/- 0.7 microg. ml(-1) at control sites (p < 0.001). Local vasodilatation reduced the retrieval of fluorescein by approximately 50% to give a steady-state concentration of fluorescein in the dialysate at 40 to 50 min after the start of perfusion of 0.9 +/- 0.3 microg. ml(-1) (p = 0.05). These studies in the skin are consistent with microdialysis theory. They suggest that clearance of solute by the blood will have a significant impact on microdialysis probe recovery and that, in the skin, the magnitude of this clearance is directly related to blood flow. PMID- 12130733 TI - Reboxetine: functional inhibition of monoamine transporters and nicotinic acetylcholine receptors. AB - The present study determined whether repeated administration of the antidepressant and selective norepinephrine (NE) uptake inhibitor reboxetine resulted in an adaptive modification of the function of the NE transporters (NETs), serotonin (5-HT) transporters, or dopamine (DA) transporters. Because antidepressants may be effective tobacco smoking cessation agents and because antidepressants have recently been shown to interact with nicotinic acetylcholine receptors (nAChRs), the interaction of reboxetine with nAChRs was also evaluated. Repeated administration of reboxetine (10 mg/kg i.p., twice daily for 14 days) did not alter the potency or selectivity of reboxetine inhibition of [(3)H]NE, [(3)H]DA, or [(3)H]5-HT uptake into striatal or hippocampal synaptosomes (IC(50) values = 8.5 nM, 89 microM, and 6.9 microM, respectively). In a separate series of experiments, reboxetine did not inhibit (K(i) > 1 microM) [(3)H]methyllycaconitine, [(3)H]cytisine, or [(3)H]epibatidine binding to rat whole brain membranes. However, at concentrations that did not exhibit intrinsic activity, reboxetine potently inhibited (IC(50) value = 7.29 nM) nicotine-evoked [(3)H]NE overflow from superfused hippocampal slices via a noncompetitive mechanism. In the latter experiments, the involvement of NET was eliminated by inclusion of desipramine (10 microM) in the superfusion buffer. Reboxetine also inhibited (IC(50) value = 650 nM) nicotine-evoked (86)Rb(+) efflux at reboxetine concentrations that did not exhibit intrinsic activity in this assay. Thus, in addition to inhibition of NET function, reboxetine inhibits nAChR function, suggesting that it may have potential as a smoking cessation agent. PMID- 12130734 TI - Pharmacological profile of (2R-trans)-4-[1-[3,5-bis(trifluoromethyl)benzoyl]-2 (phenylmethyl)-4-piperidinyl]-N-(2,6-dimethylphenyl)-1-acetamide (S) Hydroxybutanedioate (R116301), an orally and centrally active neurokinin-1 receptor antagonist. AB - In comparison with a series of reference compounds, (2R-trans)-4-[1-[3,5 bis(trifluoromethyl)benzoyl]-2-(phenylmethyl)-4-piperidinyl]-N-(2,6 dimethylphenyl)-1-acetamide (S)-Hydroxybutanedioate (R116301) was characterized as a specific, orally, and centrally active neurokinin-1 (NK(1)) receptor antagonist with subnanomolar affinity for the human NK(1) receptor (K(i): 0.45 nM) and over 200-fold selectivity toward NK(2) and NK(3) receptors. R116301 inhibited substance P (SP)-induced peripheral effects (skin reactions and plasma extravasation in guinea pigs) and a central effect (thumping in gerbils) at low doses (0.08-0.16 mg/kg, s.c. or i.p.), reflecting its high potency as an NK(1) receptor antagonist and excellent brain disposition. Higher doses blocked various emetic stimuli in ferrets, cats, and dogs (ED(50) values: 3.2 mg/kg, s.c.; 0.72 2.5 mg/kg, p.o.). Even higher doses (11-25 mg/kg, s.c.) were required in mice (capsaicin-induced ear edema) and rats (SP-induced extravasation and salivation), consistent with lower affinity for the rodent NK(1) receptor and known species differences in NK(1) receptor interactions. R116301 inhibited the ocular discharge (0.034 mg/kg) but not the dyspnoea, lethality, or cough (>40 mg/kg, s.c.) induced by [betaALA(8)]-neurokinin A (NKA) (4-10) in guinea pigs, attesting to NK(1) over NK(2) selectivity. R116301 did not affect senktide-induced miosis (>5 mg/kg, s.c.) in rabbits, confirming the absence of an interaction with the NK(3) receptor. R116301 was inactive in guinea pigs against skin reactions induced by histamine, platelet-aggregating factor, bradykinin, or Ascaris allergens (>10 mg/kg, s.c.). In all species, R116301 showed excellent oral over parenteral activity (ratio, 0.22-2.7) and a relatively long duration (6.5-16 h, p.o.). The data attest to the specificity and sensitivity of the animal models and support a role of NK(1) receptors in various diseases. PMID- 12130735 TI - Cocaine metabolism accelerated by a re-engineered human butyrylcholinesterase. AB - Plasma butyrylcholinesterase (BChE) is important in the metabolism of cocaine, but natural human BChE has limited therapeutic potential for detoxication because of low catalytic efficiency with cocaine. Here we report pharmacokinetics of cocaine in rats treated with A328W/Y332A BChE, an excellent cocaine hydrolase designed with the aid of molecular modeling. Compared with wild-type BChE, this enzyme hydrolyzes cocaine with 40-fold improved k(cat) (154 min(-1) versus 4.1 min(-1)) and only slightly increased K(M) (18 microM versus 4.5 microM). In rats given this hydrolase (3 mg/kg i.v.) 10 min before cocaine challenge (6.8 mg/kg i.v.), cocaine half-life was reduced from 52 min to 18 min. Mirroring the reductions of plasma cocaine were large increases in benzoic acid, a product of BChE-mediated cocaine hydrolysis. All other pharmacokinetic parameters confirmed a large, dose-dependent acceleration of cocaine removal by the injected cocaine hydrolase. These results show that A328W/Y332A, an efficient cocaine hydrolase in vivo as well as in vitro, might promote cocaine detoxication in a clinical setting. PMID- 12130736 TI - Renal cytochrome p450 oxygenases and preglomerular vascular response to arachidonic acid and endothelin-1 following ischemia/reperfusion. AB - This study tested the hypothesis that cytochrome P450 (P450) metabolites of arachidonic acid (AA) contribute to the vascular changes in ischemia/reperfusion (I/R) injury in the rat. In this study, P450-dependent omega-hydroxylase-mediated vascular reactivity of the rat renal interlobular and arcuate vessels [preglomerular vessels (PGMV)] was measured in left kidneys subjected to I/R. Clipping the left renal artery and vein for 30 min followed by reperfusion (I/R) for 3, 6, and 24 h markedly reduced renal microsomal omega-hydroxylase-mediated conversion of [(14)C]AA to 20-hydroxyeicosatetraenoic acid (HETE) that amounted to 34, 37, and 58% of the control enzyme activity, respectively. CYP4A protein expression was also reduced. There was no significant change in epoxygenase activity. Despite these changes, constriction of the rat PGMV by AA or endothelin 1 (ET-1) was not different in vessels from the clipped and nonclipped (contralateral) kidney. Clofibrate (250 mg/kg i.p.), an inducer of CYP4A protein and omega-hydroxylase enzymes, did not increase 20-HETE production but selectively enhanced the vasoconstriction produced by AA and ET-1 in the clipped but not the contralateral kidney without affecting the constriction produced by 9,11-dideoxy-9alpha,11alpha-methanoepoxy prostaglandin F(2alpha). On the other hand, administration of 2% NaCl (w/v, orally for 7 days) to induce P450-dependent epoxygenase activity attenuated AA-induced vasoconstriction but enhanced ET-1 induced vasoconstriction only in the clipped kidney. These data indicate that the reduction in CYP4A protein expression and enzyme activity in I/R is an adaptive mechanism to preserve renal vasculature from excessive vasoconstriction. Moreover, the increase in epoxygenase activity following salt loading may account for the diminished vasoconstriction evoked by AA. However, the enhancing effect of salt on ET-1-induced vasoconstriction in I/R appears to result from an overwhelming effect of salt-induced sensitization of the renal vasculature to ET 1 over the enhanced production of dilator epoxygenase products. PMID- 12130737 TI - Suppression of adenosine A(3) receptor-mediated hypotension and mast cell degranulation in the rat by dexamethasone. AB - Dexamethasone increases the expression of adenosine A(3) receptors and augments degranulation in response to their activation in the rat basophilic leukemia cell line, RBL-2H3. We have studied the effects of dexamethasone on mast cell activation induced by A(3) receptor stimulation in vivo. Administration of the A(3) receptor agonist APNEA [N(6)-2-(4 aminophenyl)ethyladenosine; 10-30 microg kg(-1) i.v.] to anesthetized Sprague-Dawley rats induced falls in blood pressure. Pretreatment with dexamethasone (1 mg kg(-1), i.p., -24 h) blocked the hypotensive response to APNEA but not those induced by the A(1) receptor agonist N(6)-cyclopentyladenosine, the A(2A) receptor agonist 2-[p-(2 carboxyethyl)phenylamino]-5'-N-ethylcarboxamidoadenosine, or the mast cell degranulating agent compound 48/80 (100-300 microg kg(-1), i.v.). APNEA (10 and 30 microg kg(-1), i.v.) and compound 48/80 (100 and 300 microg kg(-1), i.v.) increased plasma histamine concentrations dose dependently. Pretreatment with dexamethasone significantly inhibited the increases induced by the lower doses of each compound. APNEA induced degranulation of mast cells in thymus but not in skin or skeletal muscle, whereas compound 48/80 induced degranulation in each tissue. Pretreatment with dexamethasone inhibited APNEA-induced degranulation of mast cells in the thymus and slightly, yet significantly, reduced degranulation induced by compound 48/80. Thus, in contrast to the findings in RBL-2H3 cells in vitro, in the whole animal, dexamethasone down-regulates the response of the mast cell to A(3) receptor activation. The qualitatively similar effects on compound 48/80 suggest that dexamethasone suppresses mast cell responsiveness by modulating site(s) downstream from the adenosine A(3) receptor, possibly at the level of the G(i) family of trimeric GTP-binding proteins. PMID- 12130738 TI - SL65.0155, a novel 5-hydroxytryptamine(4) receptor partial agonist with potent cognition-enhancing properties. AB - SL65.0155 [5-(8-amino-7-chloro-2,3-dihydro-1,4-benzodioxin-5-yl)-3-[1-(2-phenyl ethyl)-4-piperidinyl]-1,3,4-oxadiazol-2(3H)-one monohydrochloride] is a novel benzodioxanoxadiazolone compound with high affinity for human 5-hydroxytryptamine (5-HT)(4) receptors (K(i) of 0.6 nM) and good selectivity (greater than 100-fold for all other receptors tested). In cells expressing the 5-HT(4(b)) and 5 HT(4(e)) splice variants, SL65.0155 acted as a partial agonist, stimulating cAMP production with a maximal effect of 40 to 50% of serotonin. However, in the rat esophagus preparation, SL65.0155 acted as a 5-HT(4) antagonist with a pK(b) of 8.81. In addition, SL65.0155 potently improved performance in several tests of learning and memory. In the object recognition task, it improved retention at 24 h when administered i.p. or p.o. (0.001-0.1 mg/kg). This effect was antagonized by the 5-HT(4) antagonist SDZ 205,557, itself without effect, demonstrating that the promnesic effects of SL65.0155 are mediated by 5-HT(4) agonism. SL65.0155 also reversed the cognitive deficits of aged rats in the linear maze task and the scopolamine-induced deficit of mice in the water maze task. Furthermore, the combined administration of an inactive dose of SL65.0155 with the cholinesterase inhibitor rivastigmine resulted in a significant promnesic effect, suggesting a synergistic interaction. SL65.0155 was devoid of unwanted cardiovascular, gastrointestinal, or central nervous system effects with doses up to more than 100-fold higher than those active in the cognitive tests. These results characterize SL65.0155 as a novel promnesic agent acting via 5-HT(4) receptors, with an excellent preclinical profile. Its broad range of activity in cognitive tests and synergism with cholinesterase inhibitors suggest that SL65.0155 represents a promising new agent for the treatment of dementia. PMID- 12130739 TI - Effect of interleukin-2 on intestinal P-glycoprotein expression and functionality in mice. AB - P-glycoprotein (Pgp), an active drug transporter expressed in enterocytes, can reduce intestinal absorption of drugs. Until now, interleukin-2 (IL2) has been reported as a Pgp modulator only in vitro. The present study examines the effects in vivo of IL2 after chronic treatment on intestinal Pgp protein expression and activity. This work also describes the effects of IL2 on the oral bioavailability of a Pgp substrate (digoxin) and of a Pgp/CYP3A cosubstrate (saquinavir). Human recombinant interleukin-2 (rIL2), administered to mice at 9 million international units/kg by intraperitoneal route twice daily for 4 days, led to a decrease in intestinal Pgp protein expression evaluated by Western blot with C219 antibody. In an in vitro everted gut sac model, rIL2 pretreatment decreased the Pgp mediated transport of rhodamine 123 across mouse intestine by 37%. Moreover, rIL2 pretreatment markedly raised the area under the curve of orally administered digoxin from 3.5 +/- 0.5 to 9.7 +/- 1.5 mg min l(-1) as a consequence of the reduction in intestinal Pgp activity. rIL2 treatment increased saquinavir bioavailability from 2.5 to 4.5%, showing that first-pass metabolism is not affected and that Pgp by itself has only a moderate effect on saquinavir oral bioavailability. In conclusion, rIL2 pretreatment reduces intestinal Pgp protein expression and activity in mice. However, the effect of such a treatment on drug bioavailability depends on the extent of their metabolism by CYP3A. PMID- 12130740 TI - Wild-type and A328W mutant human butyrylcholinesterase tetramers expressed in Chinese hamster ovary cells have a 16-hour half-life in the circulation and protect mice from cocaine toxicity. AB - Human butyrylcholinesterase (BChE) hydrolyzes cocaine to inactive metabolites. A mutant of human BChE, A328W, hydrolyzed cocaine 15-fold faster compared with wild type BChE. Although the catalytic properties of human BChE secreted by Chinese hamster ovary (CHO) cells are identical to those of native BChE, a major difference became evident when the recombinant BChE was injected into rats and mice. Recombinant BChE disappeared from the circulation within minutes, whereas native BChE stayed in the blood for a week. Nondenaturing gel electrophoresis showed that the recombinant BChE consisted mainly of monomers and dimers. In contrast, native BChE is a tetramer. The problem of the short residence time was solved by finding a method to assemble the recombinant BChE into tetramers. Coexpression in CHO cells of BChE and 45 residues from the N terminus of the COLQ protein yielded 70% tetrameric BChE. The resulting purified recombinant BChE tetramers had a half-life of 16 h in the circulation of rats and mice. The 16-h half-life was achieved without modifying the carbohydrate content of recombinant BChE. The protective effect of recombinant wild-type and A328W mutant BChE against cocaine toxicity was tested by measuring locomotor activity in mice. Pretreatment with wild-type BChE or A328W tetramers at a dose of 2.8 units/g i.p. reduced cocaine-induced locomotor activity by 50 and 80%. These results indicate that recombinant human BChE could be useful for treating cocaine toxicity in humans. PMID- 12130741 TI - Characterization of epibatidine binding to medial habenula: potential role in analgesia. AB - The objective of the present study was to characterize a recently described binding site in the habenula, which has high affinity for [(3)H]epibatidine and low affinity for nicotine and acetylcholine. We report that the extension of this binding area in coronal and horizontal sections corresponds to the anatomical extension of the medial habenula. The affinity (K(D)) of the medial habenula receptors for [(3)H]epibatidine was estimated to be 0.5 nM using an autoradiographic saturation assay, whereas the affinity of the binding site for nicotine and acetylcholine was estimated to be 5 and 8 microM, respectively. The receptor density (B(max)) in the medial habenula was estimated to be about 1100 fmol/mg wet weight using [(3)H]epibatidine. The subunit composition of the "epibatidine receptor" was investigated by the ability of different compounds with affinity to various subtypes of nicotinic receptors to displace [(3)H]epibatidine bound to the receptor. The results suggest that the receptor contains alpha3 subunits but that it is unlikely to be an alpha3beta4 nicotinic receptor. Systemic administration of epibatidine has analgesic effects in rats. Here we report that 2 x 1 microl of 10 nM epibatidine, resulting in a 2 x 10-fmol dose, administered directly to the medial habenula by bilateral stereotactic injection had an analgesic effect measured in the hot-plate test. This dose of epibatidine increased hot-plate latency significantly, whereas 2 x 2 fmol of epibatidine or 2 x 10 fmol of nicotine were without effect. This leads us to suggest that the medial habenular epibatidine binding site might be a valuable target for the development of non-opiate analgesics. PMID- 12130742 TI - Appetite-boosting property of pro-melanin-concentrating hormone(131-165) (neuropeptide-glutamic acid-isoleucine) is associated with proteolytic resistance. AB - Melanin-concentrating hormone (MCH) is a cyclic neuropeptide, with a major role in stimulation of feeding behavior in mammals. MCH signals in the brain occur via two seven-transmembrane G protein-coupled receptors, namely MCH1 (SLC-1, MCH(1), MCH-R1, or MCH-1R) and MCH2 (SLT, MCH(2), MCH-R2, or MCH-2R). In this study, we demonstrate that the pro-MCH(131-165) peptide neuropeptide-glutamic acid isoleucine (NEI)-MCH is more potent than MCH in stimulating feeding in the rat. Using rat MCH1-expressed human embryonic kidney 293 cells, we show that NEI-MCH exhibits 5-fold less affinity in a binding assay and 2-fold less potency in a cAMP assay than MCH. A similar 7- to 8-fold shift in potency was observed in a Ca(2+)(i) assay using rat MCH1 or human MCH2-transfected Chinese hamster ovary cell models. This demonstrates that NEI-MCH is not a better agonist than MCH at either of the MCH receptors. Then, we compared the proteolysis resistance of MCH and NEI-MCH to rat brain membrane homogenates and purified proteases. Kinetics of peptide degradation using brain extracts indicated a t(1/2) of 34.8 min for MCH and 78.5 min for NEI-MCH with a specific pattern of cleavage of MCH but not NEI MCH by exo- and endo-proteases. Furthermore, MCH was found highly susceptible to degradation by aminopeptidase M and endopeptidase 24.11, whereas NEI-MCH was fully resistant to proteolysis by these enzymes. Therefore, our results strongly suggest that reduced susceptibility to proteases of NEI-MCH compared with MCH account for its enhanced activity in feeding behavior. NEI-MCH represents therefore the first MCH natural functional "superagonist" so far described. PMID- 12130743 TI - Chronic morphine-induced changes in mu-opioid receptors and G proteins of different subcellular loci in rat brain. AB - Prolonged exposure to opioid agonists can induce adaptive changes resulting in tolerance and dependence. Here, rats were rendered tolerant by subcutaneous injections of increasing doses of morphine from 10 to 60 mg/kg for 3, 5, or 10 consecutive days. Binding parameters of the mu-opioid receptor in subcellular fractions were measured with [(3)H]DAMGO ([D-Ala(2),N-Me-Phe(4),Gly(5)-ol] enkephalin). Although the density of surface mu-sites did not change after the 5 day morphine treatment, up-regulation of synaptic plasma membrane binding was detected after the 10-day drug administration. In contrast, the number of mu binding sites in a light vesicle or microsomal fraction (MI) was elevated by 68 and 30% after 5 and 10 days of morphine exposure, respectively. The up-regulated MI mu-sites displayed enhanced coupling to G proteins compared with those detected in saline-treated controls. Pertussis toxin catalyzed ADP ribosylation, and Western blotting with specific antisera was used to quantitate chronic morphine-induced changes in levels of various G protein alpha-subunits. Morphine treatment of 5 days and longer induced significant increases in levels of Galpha(o), Galpha(i1), and Galpha(i2) in MI fractions that are part of an adaptation process. Up-regulation of intracellular mu-sites may be the result of post-translational changes and in part de novo synthesis. The results provide the first evidence that distinct regulation of intracellular mu-opioid receptor G protein coupling and G protein levels may accompany the development of morphine tolerance. PMID- 12130745 TI - Vasoactivity of diadenosine polyphosphates in human small mesenteric resistance arteries. AB - Diadenosine polyphosphates (ApnA) (n = 3-6) induced vasoconstrictions in isolated human mesenteric resistance arteries (hMRAs) mounted in a microvessel myograph (rank order of potency: Ap5A > Ap6A > Ap4A > Ap3A). The contractile effects of ApnA in hMRA were similar to their effects in rat MRA investigated previously. ATP, ADP, AMP, and adenosine had less contractile potency than ApnA, suggesting that the observed effects were not induced by the degradation products of ApnA. Ap4A- and Ap5A-induced vasoconstriction was inhibited by pyridoxalphosphate-6 azophenyl-2',4'-disulfonic acid (PPADS) (P2X purinoceptor antagonist) but not by ADP3'5' (P2Y purinoceptor antagonist). Thus, this purinergic vasoconstriction of hMRA seems to be P2X but not P2Y purinoceptor-mediated. In precontracted hMRA all ApnA caused vasorelaxations but (in contrast to rat MRA) the potencies of the ApnA did not differ significantly from each other. The ApnA degradation products had less vasorelaxing potency than ApnA, demonstrating that the vasorelaxations can be ascribed to the ApnA themselves. Ap5A-induced vasorelaxation of hMRA could neither be inhibited with ADP3'5' nor with PPADS, which reveals a decisive difference to the rat MRA where the inhibitory profile demonstrated the importance of the P2Y purinoceptor for Ap5A-induced vasorelaxation. However, Ap4A induced vasorelaxation in hMRA could be inhibited by ADP3'5'. These findings show that Ap4A-induced vasorelaxation in hMRA is due to P2Y purinoceptor activation, that Ap5A evokes vasorelaxation in hMRA via another mechanism than Ap4A, and that data derived from the animal model cannot be simply transferred to human conditions. PMID- 12130744 TI - The activation of neuronal nitric-oxide synthase by various divalent cations. AB - Nitric-oxide synthase (NOS; EC 1.14.13.39) catalyzes the oxidation of L-arginine to nitric oxide (NO(.)) and L-citrulline via the intermediate N(omega)-hydroxy-L arginine. Of the three distinct isoforms of NOS that have been characterized, the constitutive neuronal NOS (NOS I) generates NO(.) associated with long-term potentiation (LTP) and early brain development. All of the NOS isoforms contain an N-terminal oxidase and a C-terminal reductase domain connected by a Ca(2+)/calmodulin binding region. To activate NOS I, Ca(2+) has to bind to calmodulin, allowing electron transport through both domains. Calcium ions are tightly regulated in cells. However, a number of other metal ions that bind and activate calmodulin may also activate NOS I. One such metal ion may be Pb(2+), which is associated with neurobehavioral and psychological alterations, including the inhibition of LTP. The effect of various divalent cations on NOS I activity was tested, and the results presented herein demonstrate that Pb(2+) and Sr(2+) can activate NOS I to a level similar to that found for Ca(2+). Finally, there is a synergy between Pb(2+) and Ca(2+) resulting in maximal activation of NOS I using minimal concentrations of both metal ions. PMID- 12130747 TI - Genetic polymorphisms of human organic anion transporters OATP-C (SLC21A6) and OATP-B (SLC21A9): allele frequencies in the Japanese population and functional analysis. AB - Genetic polymorphisms of human organic anion transporting polypeptides OATP-C (SLC21A6) and OATP-B (SLC21A9) in the Japanese population were analyzed. The allele frequencies of OATP-C*1a, OATP-C*1b (N130D), OATP-C*1c (R152K and D241N), and OATP-C*5 (V174A) were 35.2, 53.7, 0, and 0.7%, respectively, in 267 healthy Japanese subjects. In the OATP-C gene, we found a novel allele called OATP-C*15 possessing two single nucleotide polymorphisms (SNPs), N130D and V174A, simultaneously. The allele frequency of OATP-C*15 was 3.0%. The allele frequencies of OATP-B*1, OATP-B*2 (T392I), and OATP-B*3 (S486F) were 69.1, 0, and 30.9%, respectively. For functional analysis, each OATP-C and OATP-B allele was expressed in human embryonic kidney (HEK293) cells, and the kinetics of uptake of [(3)H]estrone-3-sulfate was determined. In the case of OATP-C alleles, no significant alteration in K(m) or V(max) values of [(3)H]estrone-3-sulfate uptake was observed, even when the V(max) values were corrected for the expression levels of OATP-C protein. In contrast, V(max), corrected with the expression level of OATP-B*3, was decreased to 42.5% of OATP-B*1, whereas the K(m) values were comparable. Since the frequency of the OATP-B*3 allele was high (30.9%) in our subjects, the SNP of S486F may affect the physiological function and/or pharmacological effects of OATP-B substrates in vivo. PMID- 12130746 TI - Antisense oligonucleotides selectively regulate aspartyl beta-hydroxylase and its truncated protein isoform in vitro but distribute poorly into A549 tumors in vivo. AB - Alternative splicing of the human beta-aspartyl (asparaginyl) hydroxylase (BAH) gene results in the expression of humbug, a truncated form of BAH that lacks the catalytic domain of the enzyme. Overexpression of BAH and humbug has been associated with a variety of human cancers, and although humbug lacks enzymatic activity, it is expressed at levels comparable with that of BAH in various cancer cell lines. Phosphorothioate antisense oligonucleotides (ONs) were designed to dissect out the function of these hydroxylase protein isoforms. In A549 cells, these ONs differentially down-regulated BAH and humbug at the mRNA and protein level. Phosphorothioate ON uptake and antisense studies were conducted in parallel in nude mice bearing A549 tumor xenografts. Microscopic examination of the tumor after administration of a fluorescein-labeled ON showed strong labeling of the outer layers of the tumor connective tissue but cells within the interior of the tumor were sparsely labeled. A modest but significant effect on tumor growth was observed in animals treated with an antisense ON directed against both BAH and humbug transcripts. However, Northern analysis of tumor RNA did not indicate a down-regulation of the targeted mRNA species. These results demonstrate the successful development of antisense ONs that selectively differentiate between the closely related beta-hydroxylase protein isoforms. However, determination of the biological function of these proteins in vivo was limited by the poor uptake properties of phosphorothioate ONs in A549 tumors. PMID- 12130748 TI - Goblet cell hyperplasia, airway function, and leukocyte infiltration after chronic lipopolysaccharide exposure in conscious Guinea pigs: effects of rolipram and dexamethasone. AB - The effects of chronic exposures (nine, 48 h apart) of conscious guinea pigs to lipopolysaccharide (LPS) (30 microg. ml(-1), 1 h) on airway function, airway histology (in particular, goblet cell numbers), and inflammatory cell infiltration of the lungs were examined as a model of chronic inflammatory lung disease, such as chronic obstructive pulmonary disease. The sensitivity of these parameters to treatment with the corticosteroid, dexamethasone, or the phosphodiesterase-4 (PDE4) inhibitor, rolipram, was determined. As the number of LPS exposures increased, there was a progressively persistent bronchoconstriction after each exposure. After nine LPS exposures, there was evidence on histological examination of airway infiltration of, predominantly, neutrophils in perivascular, peribronchial, and alveolar tissues. After chronic LPS exposure, the airway epithelium possessed a marked goblet cell hyperplasia and evidence of inflammatory edema, features contributory to reduced airway caliber. Treatment with dexamethasone (20 mg. kg(-1)) or rolipram (1 mg. kg(-1)), administered (i.p.) 24 and 0.5 h before exposure and 24 and 47 h after each subsequent exposure, attenuated the inflammatory cell infiltration into the airway, goblet cell hyperplasia, and inflammatory edema. Dexamethasone exacerbated, whereas rolipram reversed, the chronic LPS-induced bronchoconstrictions. This study demonstrates that chronic LPS causes persistent bronchoconstriction, neutrophilic airway inflammation, goblet cell hyperplasia, and edema. These rolipram-sensitive features suggest the potential of PDE4 inhibitors in chronic inflammatory lung diseases. PMID- 12130749 TI - Extent and direction of ghrelin transport across the blood-brain barrier is determined by its unique primary structure. AB - The novel hormone ghrelin is a potent orexigen that may counterbalance leptin. Ghrelin is the only secreted molecule requiring post-translational acylation with octanoic acid to ensure bioactivity. Ghrelin, predominantly derived from the stomach, may target neuroendocrine networks within the central nervous system (CNS) to regulate energy homeostasis. This would require ghrelin to cross the blood-brain barrier (BBB). In mice, we examined whether ghrelin crosses the BBB and whether its lipophilic side chain is involved in this process. We found that saturable systems transported human ghrelin from brain-to-blood and from blood-to brain. Mouse ghrelin, differing from human ghrelin by two amino acids, was a substrate for the brain-to-blood but not for the blood-to-brain transporter and so entered the brain to a far lesser degree. des-Octanoyl ghrelin entered the brain by nonsaturable transmembrane diffusion and was sequestered once within the CNS. In summary, we show that ghrelin transport across the BBB is a complex, highly regulated bidirectional process. The direction and extent of passage are determined by the primary structure of ghrelin, defining a new role for the unique post-translational octanoylation. PMID- 12130750 TI - The relationship of clinical QT prolongation to outcome in the conscious dog using a beat-to-beat QT-RR interval assessment. AB - QT interval prolongation of the electrocardiogram has been associated with the occurrence of life-threatening fatal ventricular arrhythmias. To understand the relationship between preclinical cardiac conduction assessment to clinical outcome, comparisons of free (unbound)-plasma drug concentrations and their associated effects in the conscious mongrel dog were made to the free plasma concentrations in humans reported to produce QT prolongation. E-4031 (an experimental class III antiarrhythmic), cisapride, terfenadine, terodiline, and verapamil all affect cardiac repolarization and can produce QT prolongation in humans. In the conscious dog, the QT interval was assessed on a beat-to-beat basis in relation to each preceding RR interval at concentrations approximating the same unbound human concentrations. E-4031, cisapride and terodiline statistically increased the QT(RR1000) interval [the QT interval at a 60 beats/min (bpm) heart rate] 23, 8, and 9 ms, respectively, at concentrations 0.3 to 15.8 times their relevant clinical level. Increases were not observed for terfenadine or verapamil (p > 0.05 at all doses). Inspection of individual dog QT versus RR interval relationships showed clear QT interval responses specific to each treatment but not readily apparent when data are averaged at a heart rate of 60 bpm. For specific rectifier K(+) current (IKr) blockers, robust effects on mean QT prolongation can be detected. However, for drugs that affect repolarization through multiple channels, the effect on the mean QT interval may be more difficult to detect. Inspection of the beat-to-beat QT-RR interval relationship in an individual animal can increase the sensitivity for more accurate clinical prediction. PMID- 12130751 TI - Are cells viable at gigapascal pressures? PMID- 12130752 TI - The intelligence blame game. PMID- 12130754 TI - Heavy-ion physics. Heavy-element fizzle laid to falsified data. PMID- 12130753 TI - HIV/AIDS meeting. Tough challenges ahead on political and scientific fronts. PMID- 12130755 TI - Middle East. Archaeologists keep joint project rolling. PMID- 12130757 TI - Solar system exploration. Panel plots clear path for planetary program. PMID- 12130756 TI - Space station research. Bigger is better for science, says report. PMID- 12130758 TI - Former Soviet Union. Armenia gears up for synchrotron. PMID- 12130759 TI - Neuroscience. Gene's effect seen in brain's fear response. PMID- 12130760 TI - Foot-and-mouth disease. Report urges U.K. to vaccinate herds. PMID- 12130761 TI - Research integrity. U.S. universities urged to do a better job. PMID- 12130762 TI - Human cloning. President's bioethics council delivers. PMID- 12130763 TI - Women's health. The vanishing promises of hormone replacement. PMID- 12130764 TI - Development. Missized mutants help identify organ tailors. PMID- 12130765 TI - Protein structure. Stretching the limits. PMID- 12130766 TI - Science and the law. Is science different for lawyers? PMID- 12130767 TI - Parasitology. Drugs to combat tropical protozoan parasites. PMID- 12130768 TI - Parasitology. Malaria--from infants to genomics to vaccines. PMID- 12130769 TI - Geophysics. Tides, earthquakes, and volcanoes. PMID- 12130770 TI - Applied physics. Testing the limits for resists. PMID- 12130771 TI - Sea level changes. How Alaska affects the world. PMID- 12130772 TI - Cell biology. When wee meets whi. PMID- 12130773 TI - The amyloid hypothesis of Alzheimer's disease: progress and problems on the road to therapeutics. AB - It has been more than 10 years since it was first proposed that the neurodegeneration in Alzheimer's disease (AD) may be caused by deposition of amyloid beta-peptide (Abeta) in plaques in brain tissue. According to the amyloid hypothesis, accumulation of Abeta in the brain is the primary influence driving AD pathogenesis. The rest of the disease process, including formation of neurofibrillary tangles containing tau protein, is proposed to result from an imbalance between Abeta production and Abeta clearance. PMID- 12130774 TI - Lichen guilds share related cyanobacterial symbionts. PMID- 12130777 TI - Nonresonant multiple spin echoes. AB - Nonresonant manipulation of nuclear spins can probe large volumes of sample situated in inhomogeneous fields outside a magnet, a geometry suitable for mobile sensors for the inspection of roads, buildings, and geological formations. However, the interference by Earth's magnetic field causes rapid decay of the signal within a few milliseconds for protons and is detrimental to this method. Here we describe a technique to suppress the effects of Earth's field by using adiabatic rotations and sudden switching of the applied fields. We observed hundreds of spin echo signals lasting for more than 600 milliseconds and accurately measured the relaxation times of a liquid sample. PMID- 12130775 TI - Oscillations and sparsening of odor representations in the mushroom body. AB - In the insect olfactory system, oscillatory synchronization is functionally relevant and reflects the coherent activation of dynamic neural assemblies. We examined the role of such oscillatory synchronization in information transfer between networks in this system. The antennal lobe is the obligatory relay for olfactory afferent signals and generates oscillatory output. The mushroom body is responsible for formation and retrieval of olfactory and other memories. The format of odor representations differs significantly across these structures. Whereas representations are dense, dynamic, and seemingly redundant in the antennal lobe, they are sparse and carried by more selective neurons in the mushroom body. This transformation relies on a combination of oscillatory dynamics and intrinsic and circuit properties that act together to selectively filter and synthesize the output from the antennal lobe. These results provide direct support for the functional relevance of correlation codes and shed some light on the role of oscillatory synchronization in sensory networks. PMID- 12130776 TI - Regulation of cerebral cortical size by control of cell cycle exit in neural precursors. AB - Transgenic mice expressing a stabilized beta-catenin in neural precursors develop enlarged brains with increased cerebral cortical surface area and folds resembling sulci and gyri of higher mammals. Brains from transgenic animals have enlarged lateral ventricles lined with neuroepithelial precursor cells, reflecting an expansion of the precursor population. Compared with wild-type precursors, a greater proportion of transgenic precursors reenter the cell cycle after mitosis. These results show that beta-catenin can function in the decision of precursors to proliferate or differentiate during mammalian neuronal development and suggest that beta-catenin can regulate cerebral cortical size by controlling the generation of neural precursor cells. PMID- 12130778 TI - Direct measurement of the reaction front in chemically amplified photoresists. AB - The continuing drive by the semiconductor industry to fabricate smaller structures using photolithography will soon require dimensional control at length scales comparable to the size of the polymeric molecules in the materials used to pattern them. The current technology, chemically amplified photoresists, uses a complex reaction-diffusion process to delineate patterned areas with high spatial resolution. However, nanometer-level control of this critical process is limited by the lack of direct measurements of the reaction front. We demonstrate the use of x-ray and neutron reflectometry as a general method to measure the spatial evolution of the reaction-diffusion process with nanometer resolution. Measuring compositional profiles, provided by deuterium-labeled reactant groups for neutron scattering contrast, we show that the reaction front within the material is broad rather than sharply defined and the compositional profile is altered during development. Measuring the density profile, we directly correlate the developed film structure with that of the reaction front. PMID- 12130779 TI - Generation of spatially coherent light at extreme ultraviolet wavelengths. AB - We present spatial coherence measurements of extreme ultraviolet (EUV) light generated through the process of high-harmonic up-conversion of a femtosecond laser. With a phase-matched hollow-fiber geometry, the generated beam was found to exhibit essentially full spatial coherence. The coherence of this laser-like EUV source was shown by recording Gabor holograms of small objects. This work demonstrates the capability to perform EUV holography with a tabletop experimental setup. Such an EUV source, with low divergence and high spatial coherence, can be used for experiments involving high-precision metrology, inspection of optical components for EUV lithography, and microscopy and holography with nanometer resolution. Furthermore, the short time duration of the EUV radiation (a few femtoseconds) will enable EUV microscopy and holography to be performed with ultrahigh time resolution. PMID- 12130781 TI - Rapid wastage of Alaska glaciers and their contribution to rising sea level. AB - We have used airborne laser altimetry to estimate volume changes of 67 glaciers in Alaska from the mid-1950s to the mid-1990s. The average rate of thickness change of these glaciers was -0.52 m/year. Extrapolation to all glaciers in Alaska yields an estimated total annual volume change of -52 +/- 15 km3/year (water equivalent), equivalent to a rise in sea level (SLE) of 0.14 +/- 0.04 mm/year. Repeat measurements of 28 glaciers from the mid-1990s to 2000-2001 suggest an increased average rate of thinning, -1.8 m/year. This leads to an extrapolated annual volume loss from Alaska glaciers equal to -96 +/- 35 km3/year, or 0.27 +/- 0.10 mm/year SLE, during the past decade. These recent losses are nearly double the estimated annual loss from the entire Greenland Ice Sheet during the same time period and are much higher than previously published loss estimates for Alaska glaciers. They form the largest glaciological contribution to rising sea level yet measured. PMID- 12130780 TI - Adhesion and friction mechanisms of polymer-on-polymer surfaces. AB - The adhesion and friction of smooth polymer surfaces were studied below the glass transition temperature by use of a surface forces apparatus. The friction force of a crosslinked polymer was orders of magnitude less than that of an uncrosslinked polymer. In contrast, after chain scission of the outermost layers, the adhesion hysteresis and friction forces increase substantially. These results show that polymer-polymer adhesion hysteresis and friction depend on the dynamic rearrangement of the outermost polymer segments at shearing interfaces, and that both increase as a transition is made from crosslinked surfaces to surfaces with long chains to surfaces with quasi-free ends. The results suggest new ways for manipulating the adhesion and friction of polymer surfaces by adjusting the state of the surface chains. PMID- 12130782 TI - Freshening of the Ross Sea during the late 20th century. AB - Ocean measurements in the Ross Sea over the past four decades, one of the longest records near Antarctica, reveal marked decreases in shelf water salinity and the surface salinity within the Ross Gyre. These changes have been accompanied by atmospheric warming on Ross Island, ocean warming at depths of approximately 300 meters north of the continental shelf, a more negative Southern Oscillation Index, and thinning of southeast Pacific ice shelves. The freshening appears to have resulted from a combination of factors, including increased precipitation, reduced sea ice production, and increased melting of the West Antarctic Ice Sheet. PMID- 12130783 TI - The function of the cranial crest and jaws of a unique pterosaur from the Early Cretaceous of Brazil. AB - The discovery of a previously undescribed pterosaur, Thalassodromeus sethi, yields information on the function of cranial crests and the feeding strategy developed by these extinct flying reptiles. The material consists of a large skull (length: 1420 millimeters, including the crest) with a huge bony crest that was well irrigated by blood vessels and may have been used for regulation of its body temperature. The rostrum consists of two bladelike laminae, the arrangement of which is analogous to the condition found in the bird Rynchops, which skims over the water to catch food, indicating that T. sethi also may have been a skimmer. PMID- 12130784 TI - Serotonin transporter genetic variation and the response of the human amygdala. AB - A functional polymorphism in the promoter region of the human serotonin transporter gene (SLC6A4) has been associated with several dimensions of neuroticism and psychopathology, especially anxiety traits, but the predictive value of this genotype against these complex behaviors has been inconsistent. Serotonin [5- hydroxytryptamine, (5-HT)] function influences normal fear as well as pathological anxiety, behaviors critically dependent on the amygdala in animal models and in clinical studies. We now report that individuals with one or two copies of the short allele of the serotonin transporter (5-HTT) promoter polymorphism, which has been associated with reduced 5-HTT expression and function and increased fear and anxiety-related behaviors, exhibit greater amygdala neuronal activity, as assessed by BOLD functional magnetic resonance imaging, in response to fearful stimuli compared with individuals homozygous for the long allele. These results demonstrate genetically driven variation in the response of brain regions underlying human emotional behavior and suggest that differential excitability of the amygdala to emotional stimuli may contribute to the increased fear and anxiety typically associated with the short SLC6A4 allele. PMID- 12130785 TI - Enhanced CpG mutability and tumorigenesis in MBD4-deficient mice. AB - The mammalian protein MBD4 contains a methyl-CpG binding domain and can enzymatically remove thymine (T) or uracil (U) from a mismatched CpG site in vitro. These properties suggest that MBD4 might function in vivo to minimize the mutability of 5-methylcytosine by removing its deamination product from DNA. We tested this hypothesis by analyzing Mbd4-/- mice and found that the frequency of of C --> T transitions at CpG sites was increased by a factor of three. On a cancer-susceptible Apc(Min/+) background, Mbd4-/- mice showed accelerated tumor formation with CpG --> TpG mutations in the Apc gene. Thus MBD4 suppresses CpG mutability and tumorigenesis in vivo. PMID- 12130786 TI - A role for peroxisomes in photomorphogenesis and development of Arabidopsis. AB - The nuclear protein DET1 is a central repressor of photomorphogenesis in plants. We have identified the molecular lesion in ted3, a mutation that dominantly suppresses the phenotypes of det1-1. TED3 encodes a peroxisomal protein (AtPex2p) essential for Arabidopsis growth. Developmental defects and the abnormal expression of many genes in det1 are rescued by ted3. ted3 also partially suppresses another pleiotropic de-etiolated mutant cop1. Thus, peroxisomes, whose functions are still largely unexplored, play a key role in a photomorphogenetic pathway negatively regulated by the DET1 and COP proteins. PMID- 12130787 TI - Integrin-mediated long-term B cell retention in the splenic marginal zone. AB - The mechanisms that control localization of marginal zone (MZ) B cells are poorly understood. Here we show that MZ B cells express elevated levels of the integrins LFA-1 (alphaLbeta2) and alpha4beta1 and that they bind to the ligands ICAM-1 and VCAM-1. These ligands are expressed within the MZ in a lymphotoxin-dependent manner. Combined inhibition of LFA-1 and alpha4beta1 causes a rapid and selective release of B cells from the MZ. Furthermore, lipopolysaccharide-triggered MZ B cell relocalization involves down-regulation of integrin-mediated adhesion. These studies identify key requirements for MZ B cell localization and establish a role for integrins in peripheral lymphoid tissue compartmentalization. PMID- 12130788 TI - Tech.Sight. Gene therapy. Hurdles and hopes for cancer treatment. AB - Remarkable progress has been achieved in the field of gene therapy over the past decade. The initial excitement in this young field has led to the development of more than 500 gene therapy protocols approved for evaluation in clinical trials. As these clinical trials progress, many obstacles have been identified that investigators in the field will need to overcome. Among the most important areas of investigation in the field of cancer gene therapy are 1) development of novel vectors to improve gene delivery, 2) development of tumor-selective binding to improve specific tumor targeting, and 3) improvement of the therapeutic window to reduce the toxicity of gene therapy administered alone or in combination with conventional agents. The challenge is not to replace current strategies for cancer therapy with gene therapy but to incorporate gene therapy as a more rational, molecularly based approach to increase our therapeutic armamentarium. PMID- 12130789 TI - Energetics of potassium ion transport in Aplysia gut. AB - Basolateral membranes of Aplysia californica foregut epithelia contain an ATP dependent Na(+)/K(+) transporter (Na(+)/K(+) pump or Na(+)/K (+) -ATPase). This Na(+)/K(+) pump accounts for both the intracellular Na(+) electrochemical potential (micro) being less than the extracelluar Na(+) micro and the intracellular K(+) micro being more than the extracellular K(+ ) micro. Also, K(+) channel activity resides in both luminal and basolateral membranes of the Aplysia foregut epithelial cells. Increased activity of the Na(+)/K(+) pump, coupled to luminal and basolateral membrane depolarization altered the K(+) transport energetics across the basolateral membrane to a greater extent than the alteration in K(+) transport energetics across the luminal membrane. These results suggest that K(+) transport, either into or out of the Aplysia foregut epithelial cells, is rate-limiting at the basolateral membrane. PMID- 12130790 TI - Light-dependent changes in the chick pineal temperature and the expression of cHsp90 alpha gene: a potential contribution of in vivo temperature change to the photic-entrainment of the chick pineal circadian clock. AB - The circadian clock is entrained to the diurnal alteration of environmental conditions such as light and temperature, but the molecular mechanism underlying the entrainment is not fully understood. In the present study, we employed a differential display-based screening for a set of genes that are induced by light in the chick pineal gland, a structure of the central clock entrainable to both light and temperature changes. We found that the level of the mRNA encoding chicken heat shock protein 90 alpha (cHSP90 alpha) was rapidly elevated in the pineal gland within a 5-min exposure of chicks to light. Furthermore, the pineal cHsp90 alpha mRNA was expressed rhythmically under both 12-hr light/12-hr dark (LD) cycles and constant dark (DD) conditions. The total amount of the pineal cHSP90 alpha protein was, however, kept at nearly constant levels under LD cycles, and immunohistochemical analyses of the pineal cHSP90 alpha showed invariable localization at the cytoplasm throughout the day. In vivo measurement of the chick pineal temperature demonstrated its light-dependent and time-of-day dependent change, and the profile was very similar to that of the pineal cHSP90 alpha mRNA level. These observations suggest that the in vivo temperature change regulates the expression of temperature-responsive genes including cHSP 90 alpha in the pineal gland. The temperature change may induce a phase-shift of the pineal clock, thereby facilitating its efficient entrainment to environmental LD cycles. PMID- 12130791 TI - A comparative study of osmoregulation in four fiddler crabs (Ocypodidae: Uca). AB - This study aims to give an integrative description of the correlation of physiological parameters of osmoregulation and the habitats of the four common Uca species in Taiwan. Uca arcuata inhabits areas close to fresh water in the upper beach. Uca formosensis is only found in the areas near the mean high water of spring tide where there is a clear dry-wet transition within a single semilunar cycle. Uca vocans is found in the lower intertidal zone. Uca lactea, the most widely distributed species, can easily be found on most muddy sand shores. The number of gills was observed and histological sectioning performed on each species. The range of salinity in which the fiddler crabs maintained their hemolymph osmolality without any significant change (i.e. osmoregulatory homeostasis) and the gill Na(+), K(+)-ATPase activity were determined by transferring individuals to different salinity tanks. The results suggest that U. formosensis and U. lactea can sustain a wider range of salinity change through both modification in gill morphology and Na(+), K(+)-ATPase activity. Uca arcuata can regulate in a hypo-osmotic condition and U. vocans tends to be a weak osmoregulator. PMID- 12130793 TI - Switching from asexual to sexual reproduction in the planarian Dugesia ryukyuensis: change of the fissiparous capacity along with the sexualizing process. AB - Asexual worms of fissiparous strain of the planarian Dugesia ryukyuensis switch from asexual to sexual reproduction, if they are fed with sexually mature worms of Bdellocephala brunnea. This suggests that the sexually mature worms have a sexualizing substance(s) that induces the sexuality in the asexual worms. Here, we found by analysis of the sexualization that the cessation of the fission, namely their asexual reproduction, occurs immediately after the acquisition of sexuality. This result suggests that the downstream mechanisms induced by the putative sexualizing substance in B. brunnea become responsible for the cessation of fission. We also found that the decapitation triggers fission in the worms even after the acquisition of sexuality if they are not sexually mature, while the fully sexualized worms never fission even though they are decapitated. This result suggests that the cessation of fission takes place via at least two steps: (1) the mechanisms associated with the cephalic system; (2) other mechanisms independent of cephalic control. PMID- 12130792 TI - Anti-bovine rhodopsin monoclonal antibody recognizing light-dependent structural change. AB - The antigenic structure of the bovine rhodopsin molecule was investigated by using a bovine rhodopsin-specific monoclonal antibody designated Rh 29. Competition assay with sealed intact disks and broken disks indicated that the antibody-binding region was localized in the intradiscal surface. An antigenic peptide obtained by a cyanogene bromide cleavage of rhodopsin was purified and determined as residues 2-39 in the amino acid sequence. Further analysis suggested that the antigenic determinant included at least residues 21-25. These results were consistent with the structural model for membrane topology of rhodopsin. The antigenicity of the rhodopsin was compared among several states. The antibody bound to both ammonyx LO-solubilized unbleached and bleached rhodopsin. In contrast, upon membrane-embedded rhodopsin, unbleached one was 100 times less antigenic than bleached one. The results suggested that the segment around the determinant of membrane-embedded rhodopsin should undergo a structural change upon absorption of light. Rh 29 detected a band corresponding to bovine, porcine and octopus opsins in immunoblotting. Protein blot of crayfish rhabdome did not show any reactive band. These bands except for crayfish reacted with concanavalin A as well. The N-terminal structure may, therefore, conserved between mammal and erthropoda and diverge between them and cepharopoda. PMID- 12130794 TI - Signification of the sexualizing substance produced by the sexualized planarians. AB - Asexual worms of an exclusively fissiparous strain (the OH strain) of the planarian Dugesia ryukyuensis keep developing hermaphroditic reproductive organs and eventually undergo sexual reproduction instead of asexual reproduction, namely fission, if they are fed with sexually mature worms of an exclusively oviparous planarian, Bdellocephala brunnea, suggesting that the sexually mature worms has a sexualizing substance(s). The fully sexualized worms no longer need the feeding on sexual worms to maintain the sexuality. Here, we demonstrate that the sexualized worms produce enough of their own sexualizing substance similar to that contained in B. brunnea. In case of surgical ablation of the sexualized worms, the fragments with sexual organs regenerate to become sexual, while those without sexual organs, namely head fragments, regenerate to return to the asexual state. The asexual regenerants from the sexualized worms are also fully sexualized by being fed with B. brunnea. Additionally, it was reported that head region in sexually mature worms lacks the putative sexualizing substance necessary for complete sexualization (Sakurai, 1981). These results suggest that the fragments without sexual organ lack enough of an amount of the putative sexualizing substance and the sexuality is maintained by the sexualizing substance contained in the sexualized worms. PMID- 12130796 TI - Annual changes of urinary progesterone and estradiol-17beta of the Dugong (Dugong dugon) in captivity. AB - Levels of urinary progesterone and estradiol-17 beta were measured twice a week in a female dugong, Dugong dugon, in captivity for two years from April 1996 to April 1998. The dugong showed 14 ovarian cycles during the period of study. Concentrations of progesterone ranged from 0.01 ng/mg creatinine (Cr) to 1.94 ng/mg Cr and the length of estrous cycle was 53.6+/-8.6 (mean+/-SEM) days based on intervals of urinary progesterone peak-to-peak measurements. Concentrations of urinary estradiol-17 beta ranged from 0.9pg/mgCr to 23.7pg/mgCr, and tended to peak just prior to elevations of progesterone during the first year of study. This is the first report demonstrates that the ovulatory cycle of the dugong is about 50 days. The present findings suggest that measurement of urinary progesterone is a useful method to detect ovarian cycle of the dugong in captivity. PMID- 12130795 TI - Localization of the cytochrome p450 side-chain cleavage enzyme in the inactive testis of the naked mole-rat. AB - Spermatogenesis was histologically examined in non-breeding male of the naked mole rat (Heterocephalus glaber) using a light microscopy. Spermatogonia, spermatocytes and spermatids were confirmed in the seminiferous tubules. However, the spermatogenesis was disordered, and many spermatocytes and spermatids were sloughing. Sperms could not be seen in the lumen of the tubules. The characteristic accumulation of interstitial cells was the most noteworthy. In the immunohistochemistry for cytochrome p450 side-chain cleavage enzyme, immunoreactions were not entirely distributed in each interstitial cell, although positive reactions were scattered in the interstitial cell-mass. The findings indicate that few interstitial cells act as a testosterone-synthesizing apparatus in the characteristic structure with accumulated cell-mass. From the immunohistochemical data we suggest the possibility that spermatogonia and Sertoli cells may secrete 17 beta-estradiol. We also suggest that 17 beta estradiol from spermatogonia and Sertoli cells may inhibit the interstitial cells from synthesizing and secreting testosterone and may suppress the later stages of the spermatogenesis to induce apoptosis of germ cells. The TUNEL methods demonstrated that cell death occurred in some spermatocytes in non-breeding males. PMID- 12130797 TI - Melatonin and the wintering strategy of the tundra vole, Microtus oeconomus. AB - Short photoperiod induces physiological changes connected to the wintering of the tundra vole, Microtus oeconomus. The aim of the present study was to investigate the effects of continuous melatonin treatment on selected hormones and enzyme activities associated with energy metabolism in the species. Liver, kidney, and muscle glycogen concentrations and glycogen phosphorylase activities, as well as liver and kidney glucose-6-phosphatase and lipase esterase activities were determined. Plasma leptin, ghrelin, thyroxine, testosterone, cortisol, and melatonin concentrations were also measured. Exogenous melatonin stimulated gluconeogenesis, increased glycogen stores, and reduced fat mobilization in kidneys. Melatonin treatment also increased the food intake of the voles. This may have been mediated via elevated ghrelin levels of the melatonin-treated animals, as ghrelin is known to increase appetite of rodents. Winter metabolism of the species does not seem to require accumulation of fat or extra stores of liver or muscle glycogen. On the contrary, successful wintering of the tundra vole presumably depends on continuous food availability. PMID- 12130798 TI - Immunohistochemical study on the fetal rat pituitary in hyperthermia-induced exencephaly. AB - Hyperthermia of fetal rats is known to cause malformations of various organs including brain. The present study was carried out to investigate the effect of the hyperthermia-induced brain damages on the development of the adenohypophysis. Mother rats of day 9.5 of pregnancy were anesthetized and immersed in hot water (43 degrees C) for 15 min. At day 21.5 of gestation, fetuses were removed by caesarian section and examined for exencephaly. Hyperthermal stress induced varying degrees of exencephaly in 36% of surviving fetal rats. In extreme cases a considerable part of head was lost. Even in those fetuses with severe brain deformities, the hypophysial stalk and neural lobe were present though they were markedly underdeveloped. In exencephalic fetuses, no immunoreactive vasopressin was detected in the neural lobe of the hypophysis. Immunohistochemical examination of the adenohypophysis showed that exencephaly caused a marked decrease in the number of growth hormone (GH)-producing cells. Other types of hormone-producing cells appeared to be unaffected by brain anomaly. The reason for a decreased population of GH cells in exencephalic fetuses is discussed in relation to their adrenocortical hypotrophy. PMID- 12130799 TI - A comparative study of body wall structures of a pogonophore and an annelid from a phylogenetic viewpoint. AB - Pogonophores are tube worms that live in reducing deep-sea waters where sunlight does not penetrate. They are highly adapted for their special habitat in lacking guts and possessing endosymbiotic chemosynthetic bacteria. Because of these peculiar characteristics, it is not yet clear whether they should be classified as annelids or not. Electron-microscopic observations of sections of a Japanese pogonophore (Oligobrachia mashikoi) show that the body wall has circular and longitudinal muscular systems. These muscular systems, however, differ from the annelid (Branchiura sowerbyi) in these ways: (1) The outer circular muscle of the pogonophore was constructed of smooth muscle cells. In contrast, that of the annelid was composed of obliquely-striated muscle cells, even though the cells were small and bore undeveloped characteristics. (2) The inner longitudinal muscle of the pogonophore was constructed of undeveloped obliquely-striated muscle cells, whereas that of the annelid was composed of well-developed ones. These observations suggest that this pogonophore can not be classified as an annelid, although many previous studies have placed pogonophores in that phylum. PMID- 12130800 TI - Variation in cannibalistic polyphenism between populations in the salamander Hynobius retardatus. AB - Organisms sometimes change their phenotype to maximize fitness according to local environments. If the frequency of the broad-headed "cannibal" morph in the larvae of the salamander Hynobius retardatus has been evolutionarily maintained at a certain level within a population as a result of local adaptation, variations in its frequency should be found among different populations with environmental variation. We investigated whether variations in the frequency of the broad headed morph were present in 2 different populations, Nopporo (a low-density population) and Erimo (a high-density population), by raising larvae from the respective populations under the same experimental conditions. The occurrence rate of the broad-headed "cannibal" morph was significantly different between the 2 populations when examined with different experimental larval densities. These results suggest that the reaction norm with respect to the frequency of the broad headed morph is different between the Nopporo and Erimo populations. Because the local populations are assumed to be selected for under different environments, the different reaction norm might have evolved in response to different selection pressures. PMID- 12130801 TI - The effect of feeding habitats on dietary shifts during the growth in a benthophagous suction-feeding fish. AB - Several mechanisms of fish ontogenetic dietary shifts, which have important ecological implications, have been proposed. We studied the mechanism of dietary shifts of a benthophagous fish Goniistius zonatus, focusing on the effect of foraging on calcareous algal mat, one of its main feeding substrates, through comparison between two local populations (Morode and Arakashi) with different feeding ecology. The studied fish (11-29 cm SL) fed on various kinds of small invertebrates inhabiting the substrates, using a suctorial feeding mode without visual discrimination towards individual prey. At Morode, gut contents of small fish consisted of more epifaunal and less infaunal organisms than those of large fish, that is, mainly small crustaceans in small fish, and many invertebrates other than crustaceans in large fish. By contrast, at Arakashi, gut contents consisted mostly of epifaunal crustaceans regardless of fish size, i.e., fish showing no dietary shifts. At Morode, G. zonatus took foods mostly from thick calcareous algal mat, whereas fish foraged mainly on thin algal mat and bare rocks at Arakashi, the difference being due to the local differences in the substrate component. The algal mat of Morode harbored much larger amount of infaunal animals than any substrates of Arakashi. At Morode, large fish more forcefully and deeply thrust the mouth into thick algal mat than small fish, and was likely to suck up more infaunal prey using great suctorial force. The comparison clearly indicates that the dietary shifts of G. zonatus at Morode resulted from size-related efficiency in straining foods from heterogeneous micro topography of thick algal mat. PMID- 12130802 TI - Sex pheromone gland of the female European corn borer moth, Ostrinia nubilalis (Lepidoptera, Pyralidae): ultrastructural and biochemical evidences. AB - The sex pheromone gland of the female European corn borer moth, Ostrinia nubilalis was studied using light and electron microscopy. The pheromone gland is formed by hypertrophied epidermal cells at the mid-dorsal region of the intersegmental membrane between abdominal segments 8 and 9/10. Active glandular cells contain extensive apical membrane foldings, a single nucleus, many free ribosomes, numerous mitochondria, microtubules and lipid droplets. Smooth endoplasmic reticulum is scanty. In young moths, the glandular cells are smaller in size, the microvilli at the apical membrane are poorly developed and the cytoplasm contains fewer mitochondria, microtubules, and no lipid droplets. The surrounding unmodified epidermal cells are small cuboidal or squamous cells. These cells have ill-defined apical membrane foldings and do not contain lipid droplets in the cytoplasm and the overlying cuticle. Fatty acids analyses revealed the presence of the sex pheromone components, (E)-11-tetradecenyl acetate, and their immediate precursors, methyl (E)-11- and methyl (Z)-11 tetradecenoate, only in the dorsal portion of the cylindrical intersegmental membrane. Results of the present study show that the sex pheromone gland of O. nubilalis is restricted to the dorsal aspect of the intersegmental membrane between segments 8-9/10 and is not a ring-gland. PMID- 12130803 TI - Male accessory gland secretory proteins in nasuta subgroup of Drosophila: synthetic activity of Acp. AB - The quantity of male accessory gland secretory proteins in relation to the number of cells in the gland, size of the gland and the duration of copulation has been studied in seven members of the nasuta subgroup of Drosophila. The study revealed that the difference in the quantity of secretions is independent of the number of secretory cells in the gland. However, a positive correlation exists between the quantity of secretions and size of the gland; while there is no correlation between the copulation duration and the quantity of secretions. Further, there is an increase in the values of all the parameters studied, with increasing distance of the species from the ancestor. PMID- 12130804 TI - The germ cell lineage identified by vas-mRNA during the embryogenesis in goldfish. AB - vas RNA has been identified in germ-line cells and its precursors in zebrafish, with the result that the germ-line lineage can be traced throughout embryogenesis. In the present study, we described vas localization and the migration of vas-positive cells in goldfish, using whole mount in situ hybridization. The signals of vas mRNA localization appeared at the marginal part of the first to third cleavage planes. The eight signals were detected during the period from the 8- cells to the 512-cell stage. At the late-blastula stage, additional numbers of vas-positive cells were observed, suggesting the proliferation of these cells. At the segmentation period, vas-positive cells showed a long extended distribution along the embryonic axis, but did not form any clusters. vas-positive cells were occasionally distributed at the head region, especially around the future otic vesicle. These signals were inherited to the primordial germ cells, suggesting that vas-positive cells were primordial germ cells (PGCs) in goldfish. PMID- 12130805 TI - Re-examination of sibling cross-sterility in the ascidian, Ciona intestinalis: genetic background of the self-sterility. AB - Self-sterility of solitary ascidians is a typical example of the allogeneic recognition, though its molecular mechanism remains an open question. In this paper we analyze the fertility between siblings from selfed and crossed eggs to understand the genetic basis of self-sterility in the ascidian, Ciona intestinalis. First, we show that the self-sterility is strict and stable, and the individuality expressed in gametes is highly diversified in the wild population that we used. Secondly, we show one-way cross-sterility and reciprocal cross-sterility within the siblings that are self-sterile but fertile with non siblings. Thirdly, we show self-sterility and cross-sterility share some natures and both are closely related to the sperm capacity not to bind to the vitelline coat of the autologous eggs or the eggs sterile to the sperm concerned. In all, this paper shows that the self-sterility is genetically governed by a multiple locus system, and that most probably individual-specific determinants are haploid expression in sperm and diploid expression in eggs, given they recognize self but not non-self. PMID- 12130806 TI - Analyses of mRNA expression patterns of cohesin subunits Rad21 and Rec8 in mice: germ cell-specific expression of rec8 mRNA in both male and female mice. AB - A multisubunit protein complex called cohesin is required for the cohesion between sister chromatids in both mitosis and meiosis in yeast. We investigate here the mRNA expression patterns of mouse homologues of the yeast mitotic cohesin rad21 and the meiotic cohesin rec8 in various organs, with special attention to their expression in gonads. Northern blot and reverse transcription polymerase chain reaction (RT-PCR) analyses revealed that, in contrast to the ubiquitous expression of rad21 mRNA in all of the organs examined, rec8 was expressed only in the gonads. We conducted in situ hybridization analysis to identify the cells that express rad21 and rec8 mRNAs in the gonads. In the testis, rad21 mRNA was expressed in somatic cells and spermatogonia but not in spermatocytes, and conversely, rec8 mRNA was expressed in spermatocytes but not in spermatogonia or somatic cells. Spermatids expressed rad21 and rec8 mRNAs simultaneously. In the ovary, rad21 mRNA was detected in all of the ovarian cells including germ cells and somatic cells, whereas rec8 mRNA was detected only in oocytes. Unlike the widespread expression of rad21 gene, therefore, the gene expression of rec8 is strictly confined to spermatocytes and spermatids in male mouse and oocytes in female mouse. The restricted expression pattern of rec8 mRNA implies its essential role in meiosis in both sexes of mammals, as has been reported in yeast. We also discuss the cooperative functions of Rad21 and Rec8 on the basis of the finding that their mRNAs are coexpressed in oocytes and spermatids. PMID- 12130807 TI - Fine structure of the storage micropocket of spermatozoa in the ovary of the guppy Poecilia reticulata. AB - To investigate the internal fertilization of the guppy Poecilia reticulata, the present electron microscopic observations were focused on the morphology of the sperm storage site in females. In the ovary of the mature female guppy, many spermatozoa were found in a synaptic knob-shaped micropocket (SSP) as the sperm storage (probably sperm entry) site on the follicle surface which was the expanded blind alley of a small tract extending from the ovarian cavity. Oocytes in the developmental stage of oil droplet formation already showed the attachment of the terminal end of the small tract opening into the ovarian cavity. The lateral wall of the tract attaching to the follicle surface consisted of epithelial cells fast jointed with tight junctions and desmosomes. The thick lateral wall of SSP was constructed with complex epithelial cell layers, and the terminal bottom was comprised of a single layer of epithelial cells on the surface of the follicular layer, which consisted of a very thin thecal cell layer, basement membrane, and granulosa cell layer. The vitellus was enclosed by the follicular layer and thin chorion, in which the micropyle was absent. In fully-grown oocytes, the germinal vesicle containing comparative short chromosomes did not always locate in the vicinity of the storage SSP of spermatozoa. PMID- 12130808 TI - Histological studies on early oogenesis in barfin flounder (Verasper moseri). AB - There is much information on oogenesis from the resumption of the first meiotic division to oocyte maturation in many vertebrates; however, there have been very few studies on early oogenesis from oogonial proliferation to the initiation of meiosis. In the present study, we investigated the histological changes during early oogenesis in barfin flounder (Verasper moseri). In fish with a total length (TL) of 50mm (TL 50mm fish), active oogonial proliferation was observed. In TL 60mm fish, oocytes with synaptonemal complexes were observed. Before the initiation of active oogonial proliferation, somatic cells which surrounded a few oogonial germ cells, started to proliferate to form the oogonial cysts that accompanied oogonial proliferation. In TL 70mm fish, however, the cyst structure of the oocyte was gradually broken by the invagination of somatic cells, and finally the oocyte became a single cell surrounded by follicle cells. Upon comparison of nuclear size, DNA-synthesizing germ cells could be divided into two types: small nuclear cells and large nuclear cells. Based on histological observation, we propose that the small nuclear cells were in the mitotic prophase of oogonia and the large nuclear cells were in the meiotic prophase of oocytes, and that the nuclear size increases upon the initiation of meiosis. PMID- 12130810 TI - Intensity of larval diapause in the bamboo borer, Omphisa fuscidentalis. AB - Larvae of the bamboo borer, Omphisa fuscidentalis, enter larval diapause in September and pupate in the following June (Singtripop et al., 1999). We examined the changes in the responses of larvae to exogenous 20-hydroxyecdysone (20E) in order to estimate the progress of diapause development. In this respect, we adopted two terms, responsiveness and sensitivity of larvae to 20E. Responsiveness was estimated by the percentage of larvae that pupated, and sensitivity was evaluated by the duration from the day of 20E injection to pupation. The responsiveness of larvae declined gradually from September to November when larvae were least responsive to 20E, and then increased markedly from January to February. This indicates that the intensity of diapause increases from September to November and terminated gradually thereafter. Thus the sequence of events as the larval responses to 20E is characterized by a V-shaped curve. Sensitivity of larvae to 20E was at the same level from September to December, and increased remarkably from December to January. The abrupt increase in the sensitivity occurred one month earlier than the bottom of the V-shaped curve of larval responsiveness, suggesting that the increases in the responsiveness and sensitivity in the latter half of diapause may be brought about by respective mechanisms. PMID- 12130809 TI - A cryptic clonal line of the loach Misgurnus anguillicaudatus (Teleostei: Cobitidae) evidenced by induced gynogenesis, interspecific hybridization, microsatellite genotyping and multilocus DNA fingerprinting. AB - In Memanbetsu town, Hokkaido island, Japan, a high frequency of natural triploid loaches Misgurnus anguillicaudatus (7.4% on average) was detected by flow cytometry for relative DNA content. Among sympatric diploid females (n=6) from a single population, we found two unique females that laid unreduced diploid eggs. They gave normal diploid progeny even after induction of gynogenesis with genetically inert UV-irradiated sperm. When fertilized with normal loach sperm, some unreduced eggs developed into triploids, but the rest into diploids. Hybridization using goldfish Carassius auratus sperm gave both normal diploid loaches and inviable allotriploid hybrids possessing the diploid loach genome and the haploid goldfish genome. Microsatellite genotyping and DNA fingerprinting demonstrated that the diploid progeny developing from the unreduced eggs were genetically identical to the mother, while the triploids had some of the paternal DNA. These results indicate that the diploid eggs reproduced unisexually as a diploid clone and in other cases developed into triploids after accidental incorporation of sperm nucleus. The presence of at least one clonal line in this area was shown by the identical DNA fingerprint detected in five out of 17 diploid loaches examined. PMID- 12130811 TI - Comparison of the morphology of the inner ear between newts and frogs in relation to their locomotory capability. AB - The ultrastructural differences between the inner ears of Japanese red-bellied newts (Cynops pyrrhogaster) and black-spotted pond frogs (Rana nigromaculata) were investigated. Scanning electron microscopic observations showed apparent morphological differences in the shape of the ampulla cristae and the localization of the striola in the saccular macula. There were differences in the length of the kinocilia of the sensory hairs in each sensory region. In addition, the diameters of the bundles of stereocilia differed between the two species: the bundles of stereocilia in the semicircular cristae were thicker in frogs than in newts, while those of the utricular and lagenal maculae were thicker in newts than in frogs. PMID- 12130812 TI - Bayesian phylogenetic analysis supports monophyly of ambulacraria and of cyclostomes. AB - Vertebrates are part of the phylum Chordata, itself part of a three-phylum group known as the deuterostomes. Despite extensive phylogenetic analysis of the deuterostome animals, several unresolved relationships remain. These include the relationship between the three deuterostome phyla (chordates, echinoderms and hemichordates), and the monophyletic or paraphyletic origin of the cyclostomes (hagfish and lampreys). Using robust Bayesian statistical analysis of 18S ribosomal DNA, mitochondrial genes and nuclear protein-coding DNA, we find strong support for a hemichordate-echinoderm clade, and for monophyly of the cyclostomes. PMID- 12130813 TI - Genetic differentiation of piscivorous chub (genus Opsariichthys) in Japan, Korea and Russia. AB - A comparison of allozyme variation, restriction fragment length polymorphisms in the mitochondrial DNA and partial sequences of the ND II gene (496 bp) was made among two lacustrine populations of the piscivorous chub (Opsariichthys uncirostris uncirostris) in Japan, four fluvial populations in Korea, one lacustrine population in Russia and one specimen from the Amur River (O. u. amurensis). All analyses indicate that the Japanese populations of piscivorous chub are separable from the Asian mainland populations of Korea and Russia. The latter populations were further divided into the Korean and Russian fish. Although opinions are divided on the phylogenetic position of the population in Lake Mikata, Japan, which shows unique morphological traits intermediate between those of the population in Lake Biwa and the mainland populations, the current analysis indicates a closer relationship to the population in Lake Biwa. PMID- 12130814 TI - A new species of Thetispelecaris (Crustacea: Peracarida) from submarine cave on Grand Cayman Island. AB - A new species of the peracaridan order Bochusacea, Thetispelecaris yurikago, is described from a submarine cave on Grand Cayman Island, the Caribbean Sea. The new species is the fourth species of the order and family, and the second of the genus. Recent studies have strongly suggested a close phylogenetic affinity between cave-dwelling and deep-sea taxa in the Bochusacea as recognized in other cavernicolous/deep-sea crustaceans such as amphipods and copepods. The morphology of the gut and female reproductive system is observed for the first time in the Bochusacea: the stomach is complex with structures such as ridges, processes, spinules, and hairs in the lumen; paired gonopores are located near the base of the fifth pereiopods on the sternite. PMID- 12130815 TI - Noradrenergic and cholinergic nerves in the uterus of the Japanese long-fingered bat, Miniopterus schreibersii fuliginosus, change with reproductive cycle. AB - The pattern of uterine innervation by noradrenergic (NA) and acetylcholinesterase positive (AChE) nerves in different reproductive stages of the adult Japanese long-fingered bats were investigated histochemically and immunohistochemically. In the non-pregnant bat, the uterine horn was supplied with abundant NA and AChE nerves. These two types of nerves were closely associated with the uterine arteries and myometrial smooth muscles. In the pregnant bat, NA and AChE nerves supplying the uterus did not degenerate much during hibernating period, but reduced markedly after arousal. In the postpartum bat, the density of nerves recovered progressively. The significant change in the innervation pattern of uterine NA and AChE nerves in the pregnant bats under and after hibernation, and in the postpartum bat must be considered in relation to the adrenergic and cholinergic controlling mechanisms on the uterine function that is matched for the unique reproductive cycle of this bat. PMID- 12130816 TI - Neural function of the mesencephalic dorsomedial nucleus (DM) on distance call production in Bengalese finches. AB - In sexually mature Bengalese finches, acoustic structures of distance calls show sexual difference. The dorsomedial nucleus (DM) of intercollicular complex is known as the midbrain vocal center of distance calls. Neural input from the robust nucleus of archistriatum (RA) was observed in the DM of sexually mature males, but not observed in that of sexually mature females. The purpose of this study is to clarify somo more details of physiological function of the neural system in the DM in distance call production. Electrical stimulation to the DM of both sexes induced a call acoustically similar to distance calls, whose duration depended on the number of the pulses/train of electrical stimulation; electrical stimulation in relatively large (or small) numbers of pulses/train induced calls with relatively long (or short) duration, respectively. Multi-unit spikes were recorded from neurons in the DM. The increment of the frequency of recorded spikos was large when the bird vocalized distance calls, and the number of the frequency decreased when the bird vocalized calls whose duration was shorter than that of distance calls. These results suggest that the neural system in the DM controls duration of distance calls in sexually mature males and females. Electrical stimulation to the DM under different pulse frequencies induced calls with different patterns of time-frequency characteristics. The relation between the pulse frequencies and time-frequency characteristics showed sexual difference. The relation between them in RA-lesioned males was similar to that in females. These results suggest that the neural circuit in the DM of sexually mature males is consisted of sexually common neural circuit controlled by the neural input from the RA, and that these sexually different neural system produce sexually different acoustic structures of distance calls. PMID- 12130817 TI - Rearing conditions required for behavioral compensation after unilateral cercal ablation in the cricket Gryllus bimaculatus. AB - The rearing condition necessary for behavioral compensation after sensory deprivation was investigated in the cricket Gryllus bimaculatus. The right-cercus ablated cricket was reared in a glass vial with a slightly larger diameter than the body length of the cricket. After two weeks of rearing in the vial, the air puff-evoked escape behavior of the cricket was investigated. The response rate (relative occurrence of the escape behavior after a standard air puff) obtained was identical with that of crickets reared in a large cage. On the other hand, unlike crickets reared in a large cage, the distorted escape directional property of the cricket reared in the vial was not compensated at all. Control experiments proved that the restraint in the vial did not affect the motor system, and the air motion from environments was not essential for the compensational recovery of the escape direction. Therefore, the ablated crickets required spontaneous walking in order to compensate the directionality of their escape. A self generated wind caused by spontaneous walking appears necessary for the crickets to realize the defect of their sensory system and to compensate the related escape behavior. A hypothesis for the compensation mechanism based on the efference copy signal is proposed. PMID- 12130818 TI - Induction and characterization of mutations at the b locus of the medaka, Oryzias latipes. AB - The b locus is one of the most familiar pigmentation loci in the medaka, but its biochemical function is still unknown. Here we report induction of new mutations at the b locus by radiation and ENU. We also characterized all these mutations and previously isolated spontaneous ones on the phenotypic basis. Unexpectively, all the 18 induced mutations reduced melanin contents in both eyes and skin correlatively, although degree of reduction was varied from mutations to mutations. Moreover, presumed null mutants (bs8, bg8, bc2, bd3, bd6, bg13, bg19, bg24) had slightly melanized (dark red) eyes. These results suggest that the b locus product plays an important but not a critical role in melanogenesis. The spontaneous mutants were divided into two types: one (bdl2, bdl3, and bp) had similarities with the induced mutants in that they had slightly colored eyes and skin, the other (bv, B', bd, bdl1, and b) exhibited normally black eyes but lightly colored skin. The present study supports our recent results (Fukamachi et al., 2001) that mutational changes were found in the coding region of the b gene in some of the mutants which reduced both eyes and skin melanogenesis, while the mutational change for the b allele could not be found there. We speculate that the bv, B', bd, bdl1, and b alleles might arise by the mutations in the regulatory region for skin melanogenesis. PMID- 12130819 TI - Comparative studies on the energy metabolism in spermatozoa of four closely related species of sea urchins (genus Echinometra) in Okinawa. AB - Sea urchins of the genus Echinometra are abundant on Okinawa reef flats in southern Japan. The Okinawan Echinometra is designated into four sympatric and closely related species: A, B, C, and D (Ea, Eb, Ec, and Ed). The sperm head size and shape gradually changes to become longer and more slender according to the following order: Ea, Eb, Ec, and Ed. To obtain information regarding speciation in Okinawan Echinometra, this study examined comparatively the energy production system of spermatozoa of Ea, Eb, Ec, and Ed. All spermatozoa contained cholesterol and several kinds of phospholipids. Glycogen, glucose, and triglyceride were present at extremely low levels. After incubation in sea-water, a decrease in the level of phosphatidylcholine (PC) was observed in all spermatozoa concomitantly with activation of motility and respiration. The hydrolysis of PC correlated with the activity of phospholipase A2. Interestingly, the amount of PC consumed, the respiratory rate, and the phospholipase A2 activity in spermatozoa of Ea and Eb were approximately two-fold higher than those of Ec and Ed. Ultrastructural studies showed that lipid bodies within mitochondria were present in the midpieces of all species of spermatozoa. They became small or disappeared after incubation in seawater. Thus, the results obtained strongly suggest that spermatozoa of Ea, Eb, Ec, and Ed all use PC located in the lipid bodies as a substrate for energy metabolism. Also, it seems likely that energy production activities in Ea and Eb spermatozoa are stronger than those in Ec and Ed. The properties of energy metabolism in different species of sea urchin may be related to their habitat. PMID- 12130821 TI - Characterization of the sperm receptor for acrosome reaction-inducing substance of the starfish, Asterias amurensis. AB - Acrosome reaction-inducing substance (ARIS) in the jelly coat of starfish eggs is a highly sulfated proteoglycan-like molecule of an apparent molecular size over 10(4) kDa and plays a pivotal role in the induction of acrosome reaction in homologous spermatozoa. It is known in Asterias amurensis that ARIS binds to a restricted area of the anterior portion of sperm head, and that a glycan fragment of ARIS, named Fragment 1, consisting of 10 repeats or so of a pentasaccharide unit retains the biological activity of ARIS to an appreciable extent. In this report, we have shown the binding of Fragment 1, a relatively small pure glycan fragment of ARIS, to the putative ARIS receptor on the sperm surface by three independent methods. First, the specific binding of P-ARIS to isolated sperm membranes was monitored in real-time by using a surface plasmon resonance detector, namely a Biacore sensor system. The specific and quantitative binding of Fragment 1 to the intact sperm and to isolated sperm membranes was similarly monitored. Secondly, the binding of 125I-labeled Fragment 1 to the intact sperm was stoichiometrically measured, for which we had developed a unique procedure for radioiodination of saccharide chains. It is found that Fragment 1 competes with P-ARIS for the binding to ARIS-receptor, suggesting that Fragment 1 is a useful ligand in the search for ARIS receptor protein(s). Thirdly, the putative receptor molecules were specifically labeled by using Fragment 1 as a ligand for photoaffinity crosslink technique. Taking these results into account, we conclude that starfish sperm have the ARIS receptor, which consists most probably of 50 to 60 kDa proteins, of reasonably high affinity (for Fragment 1, Kd = 15 microM, Bmax = 8.4 x 10(4) per cell). PMID- 12130820 TI - PCR-cloning of cadmium-inducible peptides in the barnacle, Megabalanus volcano. AB - A 340 bp DNA fragment was amplified from barnacle (Megabalanus volcano) cDNA by polymerase chain reaction using primers designed based on the amino acid sequences of barnacle cadmium-inducible peptides CdlP1 and CdlP2. The whole sequence was determined by rapid amplification of cDNA ends method. The cDNA contained an open reading frame encoding 71 amino acid residues and the sequences for CdlP1 and CdlP2 were found to be located in the center of this coding region. Although CdlP1 and CdlP2 had been detected only in the cadmium-exposed barnacles, their mRNA was present both in cadmium-exposed barnacles and in unexposed barnacles. These results suggest that posttranslational proteolytic processing may be induced in the presence of cadmium. PMID- 12130822 TI - Cortactin-binding protein 90 (CBP90) expression in the mouse mammary glands during prolactin-induced lobuloalveolar development. AB - We have previously performed suppression subtractive hybridization to identify genes that were induced during prolactin (PRL)-driven lobuloalveolar development of the mammary gland. This suggested that cortactin-binding protein 90 (CBP90), which is known to be a brain-specific protein that binds to cortactin, was expressed under the regulation of PRL in the mammary glands (preliminary observation). In this study, the expression of CBP90 was examined in the mammary glands of mice under manipulated hormonal circumstances. PRL treatment by 9 days of pituitary grafting induced CBP90 expression in the normal mammary glands but not in the cleared fat pads, while cortactin was expressed constitutively in both the normal mammary glands and the cleared fat pads. Unlike milk proteins, longer treatment with PRL (36 days of pituitary grafting) did not increase the expression level of CBP90 mRNA, while it slightly increased the cortactin mRNA level. Mammary CBP90 mRNA expression was induced by pituitary grafting but not by progesterone treatment in PRL-deficient mice, while pituitary grafting induced mammary CBP90 expression in ovariectomized PRL-deficient mice only when estrogen and progesterone were appropriately supplemented to permit the formation of alveolar buds. The CBP90 protein was detected by immunohistochemistry in the luminal epithelium of the alveolar buds and more faintly in the ductal epithelium. Thus, from the unique expression pattern, CBP90 may be useful as a molecular marker for the hormone-stimulated development of mammary alveolar buds. PMID- 12130824 TI - Taxonomic study of the kinorhyncha in Japan. III. Echinoderes sensibilis n. sp. (Kinorhyncha: Cyclorhagida) from Tanabe Bay. AB - A new species of echinoderid kinorhynch, Echinoderes sensibilis, is described and illustrated using light and electron microscopy. The specimens were collected from masses of the red algae Corallina pilulifera growing in intertidal pools in Tanabe Bay, Honshu Island, Japan. Diagnostic characters of E. sensibilis include the presence of middorsal spines on segments 6-10; lateral spines/tubules on segments 4, 7-12; remarkable trapezium-like subventral fields of minute cuticular hairs on segments 5-12. The positions of numerous sensory spots, large sieve areas, glandular tubes and the shape of terminal tergal and sternal extensions are also diagnostic. All taxonomic characters used for this description are illustrated by SEM. Echinoderes sensibilis constitutes the fifty-eight valid species of the genus Echinoderes and the fourteenth species described from the Pacific Ocean. This is the third representative of Pacific kinorhynchs found only in the intertidal zone and the first Pacific Echinoderes living on red macroalgae in intertidal pools. PMID- 12130825 TI - Population differentiation and gene flow revealed by microsatellite DNA markers in the house mouse (Mus musculus castaneus) in Taiwan. AB - We analyzed population subdivision and gene flow of the Southeast Asian house mouse (Mus musculus castaneus) in Taiwan by using six microsatellite DNA markers. Seven populations of the house mouse (187 individuals), including one from Fukien Province in southeastern China, which is separated from Taiwan by the Taiwan Strait, were analyzed in this study. The overall polymorphic level at the six loci was high (He = 0.76) although individual populations varied in their levels of heterozygosity (He = 0.35-0.83). For the populations within Taiwan, there was no evidence of isolation by distance and the level of gene flow was not (inversely) correlated to geographic distances. Gene flow was estimated to be higher across the Taiwan Strait than within the island of Taiwan. These observations of gene flow cannot be understood unless in the context of the historical human settlements and agricultural expansion, and the commensal habits of the species. We also discussed the causes of population subdivision and genetic variation among populations in terms of ecological characteristics of the house mouse in Taiwan. PMID- 12130826 TI - Molecular phylogeography of the red deer (Cervus elaphus) populations in Xinjiang of China: comparison with other Asian, European, and North American populations. AB - To illustrate phylogeography of red deer (Cervus elaphus) populations of Xinjiang, we determined their mitochondrial DNA (mtDNA) control region sequences, and then investigated geographic variations and phylogenetic relationships between Xinjiang populations and other populations from Asia, Europe, and North America. The C. elaphus mtDNA control region shared different copy numbers of tandem repeats of 38 to 43-bp motifs which clearly distinguished the Western lineage from the Eastern lineage of this species in Eurasia. The western lineage comprised the Tarim populations from southern Xinjiang and the European populations, all of which had four copies of the motifs. By contrast, the Eastern lineage consisted of populations from northern Xinjiang (Tianshan and Altai Mountains), other Asian areas (Alashan, Gansu, Tibet, Mongolia, and northeastern China), and North America, all of which shared six copies of the motifs. MtDNA phylogenetic trees showed that there are two major clusters of haplotypes which referred to the Western and Eastern lineages, and that subgroupings of haplotypes in each cluster were congruent with their geographic distributions. The present study revealed that a boundary separating the Western lineage from the Eastern lineage occurs between Tarim Basin and Tianshan Mountains in Xinjiang. Meanwhile, North American populations were genetically closer to those of northern Xinjiang, northeastern China, and Mongolia, supporting that C. elaphus immigrated from northeastern Eurasia to North America through the glacier-induced land-bridge (Beringia) which had formed between the two continents after Late Pleistocene. PMID- 12130823 TI - Effects of starvation and refeeding on gonadotropin and thyrotropin subunit mRNAs in male Japanese quail. AB - The contents of mRNAs encoding LH beta-, FSH beta-, TSH beta- and common alpha subunit precursor molecules were measured in food-deprived and subsequently re fed male Japanese quail. Pituitary LH beta, FSH beta and common alpha mRNA levels were decreased by starvation, and increased to the control levels by re-feeding. The rates of decreases of LH beta and common alpha mRNA levels were greater the corresponding rate for FSH beta levels. Pituitary TSH beta mRNA levels were not decreased by starvation, but increased transitorily by re-feeding. Plasma LH and triiodothyronine levels were decreased by starvation, and then increased to control levels by re-feeding, while plasma FSH and thyroxine levels did not show significant changes. Plasma LH and FSH levels showed positive correlations with pituitary common alpha and FSH beta mRNA levels, respectively, while plasma thyroxine levels showed a negative correlation with TSH beta mRNA levels. Hepatic weight was decreased slightly but significantly by starvation, and then showed a remarkable rebound after re-feeding was started. These results suggest that LH synthesis and secretion are more sensitive to starvation than FSH synthesis and secretion in Japanese quail, and that LH production recovered to initial levels within several days when birds were fully fed. Also, there is a possibility that the synthesis of TSH is accelerated transitorily by re-feeding. Furthermore, these results showed that there are different relationships between the plasma levels of LH, FSH, and TSH and the various hormone subunit mRNA levels. The remarkable change in hepatic weight leads us to assume that hepatic thyroid hormone metabolism is affected by starvation and re-feeding. PMID- 12130827 TI - Androgen- and estrogen-dependent sex differences in host resistance to Strongyloides venezuelensis infection in Wistar rats. AB - The effects of male and female sex hormones on the protective capacity of Wistar rats against infection with Strongyloides venezuelensis were investigated. Male rats were more susceptible than females in terms of worm recovery from the lungs. Orchidectomy of male animals significantly reduced the plasma testosterone concentration and increased host resistance to the migratory stages of S. venezuelensis larvae. In contrast, ovariectomy of female animals significantly decreased host resistance in association with a significant reduction of estrogen levels. To examine the direct effect of sex hormones, exogenous testosterone and estrogen were implanted into animals. Susceptibility significantly increased or decreased in ovariectomized females given testosterone or estrogen, respectively. These results suggest that male and female sex hormones are important in the down and up-regulation of host resistance against S. venezuelensis in Wistar rats. PMID- 12130828 TI - Supplementation of heterologous complement induces anti-Thy-1.1 nephritis in the Mongolian gerbil (Meriones unguiculatus). AB - Anti-Thy-1.1 nephritis in the rat is a popular experimental model for mesangial proliferative glomerulonephritis (GN). This model is characterized by direct binding of anti-Thy-1.1 antibody with Thy-1.1 antigen expressed on mesangial cells (MCs) of glomeruli in the rat. A single injection of anti-rat thymocyte serum (ARTS) results in GN with proteinuria and extensive mesangiolysis. Development of mesangiolysis and proteinuria are complement-dependent. We previously demonstrated Thy-1.1 antigen, similar to the rat, in thymocytes, brain cells and MCs of the kidney in the Mongolian gerbil (MG). In this study, we attempted to develop a MG nephritis model, but an injection of ARTS did not induce GN. An additional injection of guinea pig serum as a complement after ARTS injection resulted in anti-Thy-1.1 nephritis in MG. Degeneration of MCs and neutrophil infiltration were observed 1 hr after GP serum injection. Mesangiolysis and fibrin exudation occurred 12 hr after the injection and MC proliferation was apparent 7 days after the injection. In the complement dependent hemolytic test, MG serum could not hemolyze sheep erythrocytes. These results suggested low activity, or depletion of some factors, in complements of MG serum. PMID- 12130830 TI - The identification and properties of chitin-binding protein b01 (CBPb01). AB - Bovine serum contains N-acetyl-D-glucosamine (GlcNAc)-sensitive opsonin inhibitory factors. In the present study, a major component of chitin-binding protein (chitin-binding protein b01, CBPb01) was purified from bovine serum, and identified CBPb01 as bovine IgM by its subunit structure, antigenic properties, and partial sequences. The results of a lectin-binding assay showed that the heavy chain of CBPb01 had a GlcNAc structure, but the commercial IgM did not. It is possible that CBPb01 interconnects through its GlcNAc structure, subsequently forming complexes. We also demonstrated that CBPb01 had opsonin-inhibitory activity, and that this activity was dependent on the binding of CBPb01 to GlcNAc on the zymosan surface. These findings indicate the presence of a kind of IgM that recognizes GlcNAc structure in the regulation of opsonization. PMID- 12130829 TI - Cardiovascular and renal effects of carvedilol in dogs with heart failure. AB - To determine the acute effects of carvedilol (beta-blocker) on cardiovascular and renal function and its pharmacokinetics in dogs. Fifteen mature mongrel dogs (7 15 kg) of both sexes were used in these experiments. Eight dogs served as controls, and seven dogs served as iatrogenic mitral regurgitation (MR) experimental animals. Carvedilol (0.2, 0.4, and 0.8 mg/kg, P.O.) was administered, and the blood carvedilol concentration was analyzed by reverse phase high-performance liquid chromatography. The response to isoproterenol or phenylephrine was also evaluated. Isoproterenol (0.025 microg/kg/min) was infused via the saphenous vein for 5 min, and phenylephrine (5 microg/kg) was injected with carvedilol (0.2, 0.4 mg/kg) or placebo for 4 days. The heart rate and arterial blood pressure were measured, and LV fractional shortening was measured by echocardiography. Glomerular filtration rate (GFR) and renal plasma flow (RPF) were measured by intravenous infusion of sodium thiosulfate and sodium para aminohippurate. Carvedilol (0.2 mg/kg) decreased the heart rate, whereas renal function, arterial blood pressure, and left ventricular contractile function were not affected. Carvedilol (0.4 mg/kg) decreased heart rate, blood pressure, and renal function. The tachycardic response to isoproterenol was significantly diminished for 36 hr by 0.4 mg/kg carvedilol. Carvedilol 0.2 mg/kg inhibited this effect for 24 hr. Thus, it is necessary to titrate the dosage of carvedilol, it should be initiated at less than 0.2 mg/kg and titrated up to 0.4 mg/kg for heart failure dogs. PMID- 12130831 TI - Effects of chemical ischemia on purine nucleotides, free radical generation, lipids peroxidation and intracellular calcium levels in C2C12 myotube derived from mouse myocytes. AB - To elucidate the mechanisms of ischemia-mediated myopathy using in vitro model, changes of purine nucleotides, membrane lipid peroxidation(TBARS), intracellular calcium ([Ca2+]i)levels, generation of free radicals, and deoxyribonucleic acid (DNA) fragmentation were examined in mouse-derived C2C12 myotubes under the condition with an inhibition of glycolytic and oxidative metabolism as the ischemic condition. In purine nucleotides, intracellular adenosine triphosphate (ATP) and guanosine triphosphate (GTP) concentrations rapidly and significantly decreased after the treatment with ischemia. No remarkable differences were observed in other purine nucleotides, with the exception of inosine monophosphate (IMP) and extracellular hypoxanthine levels, both of which increased significantly during the ischemia. The lactate dehydrogenase activity in culture supernatant of C2C12 myotubes increased significantly from 2 to 4 hr after the ischemia. On the generation of free radicals, no spectrum was detected in supernatants throughout the observation period, whereas supernatant TBARS concentration increased rapidly and significantly after the ischemia. The relative intensity of [Ca2+]i significantly increased after the ischemia. On the fragmented deoxyribonucleic acid(DNA), no TUNEL positive cells was detected in C2C12 myotubes after 1 hr of the ischemia, however the positive cell percentage subsequently increased. From these results, it was suggested that the ischemic condition induced changes of membrane permeability and increase of [Ca2+]i, both of which lead to cell membrane damage, although a free radical generation was not detected. The ischemic condition also induced the release of substrate hypoxanthine for free radical generation and might initiate the apoptotic pathway in C2C12 myotubes. PMID- 12130833 TI - Uterine disorders diagnosed by ventrotomy in 47 rabbits. AB - The type, ages of occurrence, primary complaints, clinical signs and mortality in forty-seven cases of uterine disorders diagnosed by ventrotomy in rabbits were analyzed retrospectively. Endometrial hyperplasia (29.8%) was most frequently observed, followed by uterine adenocarcinoma (21.3%). Tumorous lesions were seen in 46.8% of the cases. The age of occurrence ranged from two years and two months to seven years and six months, with a peak at four to five years of age. The most common primary complaint was bleeding (62.2%), followed by mammary gland abnormality (12.8%) and increased abdominal circumference (10.6%). Physical examinations revealed mammary gland disorders such as mammary cysts in 31.9% of the cases. Uterine disorders were detected by palpation in 15 out of 32 cases with a primary complaint of bleeding. Ultrasonography showed uterine disorders in 21 out of 24 cases, suggesting that ultrasonography could be useful in the diagnosis of uterine disorders. The outcome seemed to be influenced by physical status rather than malignancy of lesions. The mortality was higher in cases with symptoms such as anorexia, emaciation, severe anemia, and dehydration. PMID- 12130834 TI - Effects of milrinone on hemodynamics and regional blood flow in the hypoxic dog. AB - Milrinone, a therapeutic agent for acute congestive heart failure, has both inotropic and vasodilatory effects, but investigations of these effects of milrinone were almost all conducted under normoxia, and few reports have investigated how milrinone affects the hemodynamics and redistribution of regional blood flow under severe hypoxia. By using colored microspheres, we investigated how milrinone affects hemodynamics and the redistribution of regional blood flow under severe hypoxia. Twelve healthy mongrel dogs were divided into 2 groups. The milrinone group was infused with milrinone cumulatively at 25, 75 and 250 microg/kg for 5 min each. The intact group was infused with saline instead of milrinone. We measured the hemodynamics and cerebrum, cerebellum and kidney blood flow in both groups. Both groups were inspired with 10% oxygen. Milrinone induced significant decrease in mean pulmonary artery and pulmonary vascular resistance, compared with the intact group. In both groups slight decreases in mean arterial pressure, systemic vascular resistance and double-product were seen. In regional blood flow, milrinone-induced increases in blood flow were seen in the cerebrum, cerebellum, and especially in the kidneys. Milrinone's vasodilatory effects were sufficient even under hypoxia. And milrinone increased regional blood flow slightly in the cerebrum and cerebellum, and significantly in the kidneys. These results suggested that milrinone protects against hypoxia-induced organ damage especially in the kidneys. In addition, milrinone is very potent in improving severe congested hemodynamics which complicates hypoxic pulmonary vasoconstriction. PMID- 12130832 TI - Tumor promoting effect of phenolphthalein on development of lung tumors induced by N-ethyl-N-nitrosourea in transgenic mice carrying human prototype c-Ha-ras gene. AB - In order to examine tumor modifying effects of phenolphthalein (PhP), female transgenic mice carrying human prototype c-Ha-ras gene (rasH2 mice) were given a single intraperitoneal injection of 60 mg/kg body weight of N-ethyl-N-nitrosourea (ENU), followed by the diet containing 12,000 ppm PhP for 26-week. Histopathologically, alveolar hyperplasias, adenomas and adenocarcinomas were observed in the ENU + PhP group, but only hyperplasias and adenomas were observed in the ENU alone group. The incidence and multiplicity of adenocarcinomas in the ENU + PhP group was significantly increased as compared to that in the ENU alone group. The combined multiplicity of adenomas and adenocarcinomas in this group was also significantly higher than that of the ENU alone group. In addition, the ratio of area of adenomas in the ENU + PhP group was significantly higher than that in the ENU alone group. The result of our study suggests that PhP has a clear tumor promoting effect in the lung of rasH2 mice. PMID- 12130836 TI - A morphological study of the thyroid gland in Risso's Dolphin, Grampus griseus. AB - To accumulate histological information of cetaceans and basic information about metabolic systems of marine mammals, the thyroid gland of Risso's dolphins was examined by gross anatomical and light and electron microscopic observations. Gross anatomically, right and left lobes of the thyroid were not clearly discriminated, and no isthmus was observed. By light microscopy, irregular or oval follicular lumens were seen, and surrounded by follicular epithelial cells. By electron microscopy, the rough endoplasmic reticulum (rER) was seen adjacently to mitochondria at the basal and lateral regions of the follicular epithelial cells. RERs at the basal side of the cells sometimes contained flocculent material with the same electron density as the follicular lumen component. Microvilli were poorly developed at the apical surface of the cells. In the apical regions of the cells, there were typical Golgi complexes, multivesicular bodies, and granules with various size and electron density. The parafollicular cells were recognized among the follicular epithelial cells and in the interstitial regions but never protruded into the follicular lumen. These cells were present singly and/or formed clusters among the follicular epithelial cells, and often located adjacent to capillaries. They were obviously discriminated from follicular epithelial cells by higher electron density of their granules. In their cytoplasm, well-developed rERs, primary lysosomes, secondary lysosomes, multivesicular bodies, and phagosomes were recognized. PMID- 12130838 TI - Age- and sex-related changes in susceptibility of Wistar rats to Strongyloides venezuelensis infection. AB - The effects of host age and sex on susceptibility to Strongyloides venezuelensis in Wistar rats were examined by counting larvae recovered from the lungs of animals 3 days after infection. The susceptibility of female rats to S. venezuelensis rapidly decreased with age and elevated estrogen. Resistance in female rats inoculated at 6 and 10 weeks of age was nine and twenty-fold higher, respectively than that in the youngest group (3 weeks). In contrast, the susceptibility of male animals was lowest in the youngest group, then increased with age and elevated testosterone. Sex differences in susceptibility were not evident in the youngest group, but became apparent with age. PMID- 12130835 TI - Sex identification of Japanese black bear, Ursus thibetanus japonicus, by PCR based on amelogenin gene. AB - A method for sex identification of the Japanese black bear was examined using a polymerase chain reaction (PCR) and sequencing of a part of the amelogenin gene. This gene is located on the X and Y chromosomes, and there are 54 nucleotide deletions on the Y chromosome-specific gene. Forty-seven (26 male and 21 female) DNA samples and 23 (13 male and 10 female) DNA samples, respectively extracted from white blood cells and hairs of Japanese black bears were analyzed. The primers SE47 and SE48 from this X-Y homologous region were used in sex identification by PCR amplification. These primers amplified X- and Y-specific bands, which could be used to discriminate between sexes by a length polymorphism in all samples. We suggest that PCR amplification using the primers SE47 and SE48 is useful for sex determination of the Japanese black bear and could be applied to DNA analysis of small samples such as hairs. PMID- 12130837 TI - Evaluation of cytokine messenger RNA expression in peripheral blood mononuclear cells from dogs with canine demodicosis. AB - Using RT-PCR and semi-quantitative PCR, mRNA expression for canine interferon (IFN)-gamma, interleukin (IL)-2, IL-4, IL-5, IL-10, tumor necrosis factor (TNF) alpha and transforming growth factor (TGF)-beta in peripheral blood mononuclear cells (PBMCs) was examined in dogs with or without demodicosis. mRNA expression for IFN-gamma as well as TNF-alpha in dogs with demodicosis (localized (LD) and generalized (GD)) was slightly lower than those in dogs without demodicosis (healthy controls). Expression of IL-5 mRNA in dogs with demodicosis was higher than that in control dogs, but there were no significant differences in IL-4 and IL-10 mRNA expression levels among the three groups. On the other hand, expression levels of TGF-beta mRNA in dogs with GD were higher than those in control dogs and dogs with LD. The expression levels of IL-5 and TGF-beta mRNA decreased in all three dogs with GD which showed resolution of the clinical signs. Taken together, these results suggest that the Th2-like response in PBMCs from dogs with demodicosis is up-regulated, and that subsequent increased expression of IL-5 and TGF-beta mRNA in dogs with GD is reversible after treatment. Therefore, these cytokines, particularly IL-5, might be a useful clinical index of the clinical course in demodicosis. Also, increased TGF-beta mRNA expression might be a key factor for revealing the difference in the mechanism of onset between LD and GD. PMID- 12130840 TI - Psittacine beak and feather disease in three captive sulphur-crested cockatoos (Cacatua galerita) in Thailand. AB - Three sulphur-crested cockatoos (Cacatua galerita) were diagnosed as psittacine beak and feather disease (PBFD). Histopathology of the feather pulp and follicles showed intracytoplasmic botryoid clusters or granular inclusion bodies in epithelial cells and macrophages. Electron microscopy revealed multiple cytoplasmic clusters of electron dense viral particles corresponding to the inclusions. PBFD virus (circovirus) DNA-specific product was detected from formalin-fixed paraffin-embedded feathers by nested polymerase chain reaction (PCR) method. PMID- 12130839 TI - Immunohistochemical and ultrastructural identification of Fusobacterium necrophorum subsp. necrophorum in bovine fatal necrotizing glossitis. AB - A 37-day-old male Japanese black calf showing marked salivation and leucocytosis died and was examined the tissues histologically. Histological lesions were characterized by severe focal necrotic glossitis on the ventral side of the root of the tongue. Immunohistochemically, Fusobacterium necrophorum subsp. necrophorum antigen was detected in the necrotic tissues and its distribution corresponded to that of the gram-negative, nonsporeforming, long filamentous organisms. Ultrastructural similarities between the organism and F. necrophorum subsp. necrophorum, but not subsp. funduliforme were observed. These findings clearly demonstrated that the fatal necrotic glossitis was caused by F. necrophorum subsp. necrophorum. This is the first report of bovine fatal necrotizing glossitis with leucocytosis caused by F. necrophorum subsp. necrophorum infection, and this organism may be an important fatal pathogen in calves with glossal lesions. PMID- 12130841 TI - Immunohistochemical detection of P-glycoprotein (PGP) and multidrug resistance associated protein (MRP) in canine cutaneous mast cell tumors. AB - Fifty-four canine cutaneous mast cell tumors were evaluated immunohistochemically for the expression of P-glycoprotein (PGP) and multidrug-resistance-associated protein (MRP). All tumors examined were graded according to the histological malignancy. ranging from grade I to III. The expression of PGP was confirmed in 15% (8/54) of whole, 33% (5/15) of grade I, 10% (3/31) of grade II, and 0% (0/8) of grade III tumors. The expression of MRP was found in 18% (10/54) of whole, 26% (4/15) of grade I, 19% (6/31) of grade II, and 0% (0/8) of grade III tumors. The cases positive to at least one of these 2 multidrug markers were 26%, 47%, 23% and 0% of whole and grade I to III tumors, respectively. These results indicate that at least 26% of canine cutaneous mast cell tumors express PGP and/or MRP and that these tumors may be resistant to several anti-cancer drugs. PMID- 12130842 TI - Lectin histochemistry of Peyer's patches in the porcine ileum. AB - Lectin staining pattern in Peyer's patches of porcine ileum was studied using twenty one biotinylated-labeled lectins as cell markers which were visualized with avidin-biotin-peroxidase complex method (ABC). WGA appears to be a selective marker for tingible body macrophages in the porcine germinal centers. ConA may be a positive marker for the lymphoid tissues, whereas 9 lectins (DBA, SBA, SJA, s WGA, PNA, ECL, UEA-I, PHA-E, and PHA-L) may be negative markers for the lymphoid tissues in all areas. PMID- 12130844 TI - Elucidation of solid-state complexation in ground mixtures of cholic acid and guest compounds. AB - The solid-state complexation between cholic acid (CA) and either methyl p hydroxybenzoate (MPB) or ibuprofen (IBP) was investigated. Powder X-ray diffractometry, IR spectroscopy and thermal analysis suggested the complex formation between CA and MPB as well as between CA and IBP by co-grinding method. The stoichiometry of CA-MPB was 1 : 1 while that of CA-IBP was 2 : 1, reflecting the effect of guest size on complex formation. The guest compounds were assumed to be included in the channel of complexes formed by hydrogen bonds among CA molecules. PMID- 12130843 TI - Langerhans cells within the follicular epithelium and the intradermal sweat duct in equine insect hypersensitivity "Kasen". AB - Histopathologic and electron microscopic observations were given on Langerhans cells (LCs) within the follicular epithelium (FE) and intradermal sweat duct (ISD) of equine "Kasen". By light microscopy, LCs were present in the greatest numbers within the FE and ISD than within the epidermal layer and the normal skin, with an occasional formation of several aggregated foci. By electron microscopy, LCs within the FE and ISD widely extended their dendritic processes between the keratinocytes and contained Birbeck granules (Bgs), mitochondria, rough endoplasmic reticula and ribosomes in the cytoplasm. Numerous Type 2 LCs, with a number of Bgs and endocytosis, and Type 3 LCs, with multivesicular bodies and endosomes of various sizes, were recognized within the FE and ISD, although inactive Type 1 LCs, with a narrow and lucid cytoplasm, were rarely seen. LCs observed within the FE and ISD in the "Kasen" skin lesions might express the particular stage corresponded to recognize, intake and process the antigens which permeate them. PMID- 12130846 TI - Evaluation of variation of acteoside and three major flavonoids in wild and cultivated Scutellaria baicalensis roots by micellar electrokinetic chromatography. AB - Micellar electrokinetic chromatography (MEKC) conditions were developed to analyze the constituents of Scutellariae Radix (SR) and Scutellaria baicalensis roots. Using the MEKC method, the major flavonoid constituents of baicalin, baicalein and wogonin of wild and cultivated S. baicalensis roots were compared. In a preliminary comparison of electropherogram, one special peak was found in a wild sample but not in a 2-year-cultivated one. The compound corresponding to the peak was isolated and identified as a phenylethanoid glycoside, acteoside, by comparing the 1H- and 13C-NMR spectral data with that of the authentic compound. This is the first time acteoside has been isolated from the Scutellaria genus. It could only be found in SR derived from wild S. baicalensis roots and 4-year cultivated plants, but not in plant materials cultivated for 3 years. Applying the MEKC method established in this study, rapid and simultaneous determinations of acteoside together with 3 flavonoids in samples were achieved. The method can thus be used for the quality control of SR in a shorter analysis period than HPLC. PMID- 12130845 TI - An in vitro and in vivo investigation into the suitability of bacterially triggered delivery system for colon targeting. AB - The colon specific drug delivery systems based on polysaccharides; locust bean gum and chitosan in the ratio of 2 : 3, 3 : 2 and 4 : 1 were evaluated using in vitro and in vivo methods. The in vitro studies in pH 6.8 phosphate buffer containing 2% w/v rat caecal contents showed that the cumulative percentage release of mesalazine after 26 h were 31.25+/-0.56, 46.25+/-0.96, 97.5+/-0.26 (mean+/-S.D.), respectively. The in vivo studies conducted in nine healthy male human volunteers for the various formulations revealed that, the drug release was initiated only after 5 h (i.e.) transit time of small intestine and the bioavailability (AUC(0-->t*)) of the drug was found to be 85.24+/-0.10, 196.08+/ 0.12, 498.62+/-0.10 microg x h/ml 26 (mean+/-S.D.), respectively. These studies on the polysaccharides demonstrated that the combination of locust bean gum and chitosan as a coating material proved capable of protecting the core tablet containing mesalazine during the condition mimicking mouth to colon transit. In particular, the formulation containing locust bean gum and chitosan in the ratio of 4 : 1 held a better dissolution profile, higher bioavailability and hence a potential carrier for drug targeting to colon. PMID- 12130847 TI - Activity-guided isolation of saponins from Kalopanax pictus with anti inflammatory activity. AB - By bioassay-guided separation, a known saponin, kalopanaxsaponin A (1) and a new saponin, pictoside A (2) were isolated from the stem bark of Kalopanax pictus as anti-inflammatory components when evaluated by vascular permeability test. Another novel saponin, pictoside B (3) was also isolated but was inactive in the test system used. The structures of pictosides A and B were elucidated as caulophyllogenin 3-O-alpha-L-rhamnopyranosyl(1-->2)-alpha-L-arabinopyranoside (2) and pictogenin (3beta,6beta,16alpha,23-tetrahydroxyolean-12-ene-28-oic acid) 3-O alpha-L-arabinopyranoside (3), respectively, by spectral analysis and by chemical degradation. Kalopanaxsaponin A and pictoside A showed significant anti inflammatory activity at the oral doses of 50 mg/kg. PMID- 12130848 TI - Synthesis of new azulene derivatives and study of their effect on lipid peroxidation and lipoxygenase activity. AB - The relationship between free radicals and acute or chronic inflammation has been well established. We have previously reported the significant antioxidant activity of the natural azulene derivatives chamazulene and guaiazulene. Furthermore, some synthetic azulene analogues have been found to possess anti inflammatory activity. In this investigation we report the synthesis of five 3 alkyl or 3-(hydroxy)alkylazulene-1-carboxylic acids and esters, from tropolone, via the corresponding furanone. The synthesised compounds were tested for their effect on the peroxidation of rat hepatic microsomal membrane lipids, applying the 2-thiobarbituric acid test. Their anti-inflammatory activity was evaluated in vitro by the offered inhibition of soybean lipoxygenase. All the tested molecules were found to inhibit lipid peroxidation by 100% at 1 mM. They were also found to considerably inhibit lipoxygenase activity. The above results are discussed in relation to the structure and physicochemical properties of the examined azulene derivatives. PMID- 12130849 TI - The effects of Lewis acid on the 1,3-dipolar cycloaddition reaction of C arylaldonitrones with alkenes. AB - The regio- and stereoselectivity of the 1,3-dipolar cycloaddition reactions of C aryl-N-alkylaldonitrones (1a-e) with some alkenes were found to be affected significantly by the addition of Lewis acid. The rate of the reaction was also affected by adding the Lewis acid. In the reactions using allyl alcohol as a dipolarophile an addition of Lewis acid caused a remarkable acceleration of the reaction and a great change in the stereoselectivity. In the reactions using ethyl acrylate as a dipolarophile the regioselectivity was reversed whether the reaction was performed in the presence or the absence of Lewis acid; i.e. isoxazolidine-5-carboxylates were obtained mainly in the absence of Lewis acid although isoxazolidine-4-carboxylates were obtained mainly in the presence of Lewis acid. When the reaction of C,N-diarylaldonitrones (1k, 1m, 1n) with ethyl acrylate was carried out in the presence of Lewis acid, the cleavage of the N-O bond of the cycloadducts giving gamma-aminoalcohols was also observed besides a reverse phenomenon of regioselectivity. PMID- 12130850 TI - Inhibitors of adhesion molecules expression; the synthesis and pharmacological properties of 10H-pyrazino[2,3-b][1,4]benzothiazine derivatives. AB - During a search for novel, orally-active inhibitors of upregulation of adhesion molecules such as intercellular adhesion molecule-1 (ICAM-1), we found a new series of 10H-pyrazino[2,3-b][1,4]benzothiazine derivatives to be potent ICAM-1 inhibitors. Of these compounds, N-[1-(10H-Pyrazino[2,3-b][1,4]benzothiazin-8 ylmethyl)piperidin-4-yl]-N',N'-dimethylsulfamide 7p showed the potent oral inhibitory activities against neutrophil migration in a murine interleukin-1 (IL 1) induced paw inflammation model. The synthesis and structure-activity relationships of these amide derivatives are described. PMID- 12130852 TI - Coordination of sodium cation to an oxygen function and olefinic double bond to form molecular adduct ion in fast atom bombardment mass spectrometry. AB - Steroidal allylic alcohols formed Na+ adduct ion peaks [M+Na]+ by the addition of NaCl in FAB mass spectrometry. A comparison of the intensities of the adduct ion peaks of allylic alcohols with those of the corresponding saturated alcohols and olefin suggested that the olefinic double bond and the proximal hydroxyl group had coordinated to Na+. The adduct ion was stable and did not undergo dehydroxylation. We suggest that the Na+ adduction will be useful for the molecular weight determination of allylic alcohols which are susceptible to dehydroxylation under FAB mass spectrometric conditions. Na+ adduct ions of alpha,beta-unsaturated carbonyl compounds were also investigated. PMID- 12130851 TI - First synthesis of (+)-alpha- and (+)-gamma-polypodatetraenes. AB - First synthesis of (+)-alpha-polypodatetraene (1) and (+)-gamma-polypodatetraene (2) was achieved from (+)-albicanol (6) and (-)-drimenol (8), respectively. The absolute structure of natural (+)-2 was established to be (5S,9S,10S)-polypoda 7,13(E),17(E),21-tetraene. PMID- 12130853 TI - Synthesis and structure-activity relationships of 4-amino-5-chloro-N-(1,4 dialkylhexahydro-1,4-diazepin-6-yl)-2-methoxybenzamide derivatives, novel and potent serotonin 5-HT3 and dopamine D2 receptors dual antagonist. AB - In search of a dopamine D2 and serotonin 5-HT3 receptors dual antagonist as a potential broad antiemetic agent, a number of benzamides were prepared from 4 amino-5-chloro-2-methoxybenzoic acid derivatives and 6-amino-1,4-dialkylhexahydro 1,4-diazepines and evaluated for their binding affinity for the dopamine D2 and the serotonin 5-HT3 receptors using rat brain synaptic and rat cortical membranes, respectively. From the results of both in vitro receptor binding and in vivo biological assays for the dopamine D2 receptor, 1-ethyl-4-methylhexahydro 1,4-diazepine ring was selected as an optimum amine moiety. Introduction of one methyl group on the nitrogen atom at the 4-position and/or modification of the substituent at the 5-position of the 4-amino-5-chloro-2-methoxybenzoyl moiety caused a marked increase in the dopamine D2 receptor binding affinity along with a potent 5-HT3 receptor binding affinity. Among the compounds, 5-chloro-N-(1 ethyl-4-methylhexahydro-1,4-diazepin-6-yl)-2-methoxy-4-methylaminobenzamide (82), 5-bromo (110), and 5-iodo (112) analogues exhibited a much higher affinity for the dopamine D2 receptor than that of metoclopramide (IC50=17.5-61.0 nM vs. 483 nM). In particular, 82 showed a potent antagonistic activity for both receptors in vivo tests. Optical resolution of the racemate 82 brought about a dramatic change in the pharmacological profile with the (R)-enantiomer exhibiting a strong affinity for both the dopamine D2 and the 5-HT3 receptors, while the corresponding (S)-enantiomer had a potent and selective serotonin 5-HT3 receptor binding affinity. PMID- 12130854 TI - Formation of an unusual dimeric compound by lead tetraacetate oxidation of a corynanthe-type indole alkaloid, mitragynine. AB - Lead tetraacetate oxidation of a Corynanthe-type indole alkaloid, mitragynine, produced mainly 7-acetoxyindolenine derivative (2) together with a dimeric compound (4) as a minor product. The novel structure having a bridge between the C-11' and C-7 positions in the respective indolenine parts and its formation mechanism were studied. PMID- 12130855 TI - The crystal structure of biatractylolide, an 8,8' (C-C) linked dimeric 12,8 eudesmanolide from the resin of Trattinickia rhoifolia WILLD. AB - A symmetrical dimeric sesquiterpenoid, biatractylolide (1), was isolated from the resin of Trattinickia rhoifolia WILLD. The structure of compound 1 was elucidated by one- and two-dimensional NMR techniques and electron impact-mass spectra (EI MS) data, and confirmed by X-ray crystallographic analysis. PMID- 12130856 TI - Conformational analysis of the NMDA receptor antagonist (1S,2R)-1-phenyl-2-[(S)-1 aminopropyl]-N,N-diethylcyclopropanecarboxamide (PPDC) designed by a novel conformational restriction method based on the structural feature of cyclopropane ring. AB - (1S,2R)-1-phenyl-2-[(S)-1-aminopropyl]-N,N-diethylcyclopropanecarboxamide (2b, PPDC), a new class of potent N-methyl-D-aspartic acid (NMDA) receptor antagonist, was designed based on a new method for restricting the conformation of compounds having a cyclopropane ring. The three-dimensional structures of PPDC obtained by the three different methods of X-ray crystallographic analysis, usual MM2 calculations in vacuum, and MM2 calculations based on the nuclear Overhauser effect (NOE) data in D2O are similar, which are in accord with that hypothesized. These results suggest that this conformational restriction method is particularly effective in designing novel biologically active molecules. PMID- 12130858 TI - Medicinal flowers. VI. Absolute stereostructures of two new flavanone glycosides and a phenylbutanoid glycoside from the flowers of Chrysanthemum indicum L.: their inhibitory activities for rat lens aldose reductase. AB - Two new flavanone glycosides, (2S)- and (2R)-eriodictyol 7-O-beta-D glucopyranosiduronic acids, and a new phenylbutanoid glycoside, (2S, 3S)-1-phenyl 2,3-butanediol 3-O-beta-D-glucopyranoside, were isolated from the flowers of Chrysanthemum indicum L. cultivated in China together with eight flavonoids. The absolute stereostructures of the new compounds were determined on the basis of chemical and physicochemical evidence. Both of the new flavanone glycosides were found to show inhibitory activity for rat lens aldose reductase. PMID- 12130857 TI - Solvent effect on photoisomerisation of 3-methyl-1-phenylbutane-1,2-dione 2 oxime. AB - Photoisomerisation of (2E)- and (2Z)-3-methyl-1-phenylbutane-1,2-dione 2-oxime (MPBDO) in several solvents was studied. With increasing dielectric constants of solvents, kinetic constants of forward reactions (E-form-->Z-form) did not change appreciably but those of reverse reactions (Z-form-->E-form) decreased. The positive correlation was found between equilibrium constants of photoisomerisation and dielectric constants of solvents. PMID- 12130859 TI - Biosynthetic study of amphidinolide W. AB - The biosynthetic origins of amphidinolide W (1) were investigated on the basis of (13)C-NMR data of 13C-enriched samples obtained by feeding experiments with [1 13C], [2-13C], and [1,2-13C2] sodium acetate in cultures of a strain Y-42 of the dinoflagellate Amphidinium sp. These incorporation patterns suggested that 1 was generated from a hexaketide chain, two acetate units, four isolated C1 units from C-2 of acetates, and four branched C1 units from C-2 of acetates. The acetate incorporation patterns for C-1-C-2-(C-21) and C-8-C-18-(C-23, C-24) of 1 corresponded well to those for C-1-C-2-(C-27) and C-5-C-15-(C-28, C-29) of amphidinolide H (2) isolated from this strain. PMID- 12130860 TI - New lignans from the heartwood of Chamaecyparis obtusa var. formosana. AB - Four new lignans, 3',4'-O,O-demethylenehinokinin (1), chamalignolide (2), 8'beta hydroxyhinokinin (3) and 7beta,8beta-epoxyzuonin A (4), as well as (-)-hinokinin (5), and (-)-zuonin A (6), were isolated from the heartwood of Chamaecyparis obtusa var. formosana. The structures of these lignans were unambiguously determined by spectroscopic methods. And the absolute configuration of 1 was elucidated with a circular dichroism (CD) spectrum. PMID- 12130861 TI - New flavonol tetraglycosides from Astragalus caprinus. AB - A new flavonol tetraglycoside, together with four acylated derivatives, were isolated from the leaves of Astragalus caprinus. Their structures were elucidated by spectroscopic methods, mainly 2D NMR, as kaempferol-3-O-[[beta-D xylopyranosyl(1-->3)-alpha-L-rhamnopyranosyl(1-->6)][alpha-L-rhamnopyranosyl(1- >2)]]-beta-D-galactopyranoside (1), its 3(Gal)-p-coumaric (2) and 3(Gal)-ferulic (3) esters, and its 4(Gal)-p-coumaric (4) and 4(Gal)-ferulic (5) esters. PMID- 12130863 TI - Chemical profiling and standardization of Lepidium meyenii (Maca) by reversed phase high performance liquid chromatography. AB - Lepidium meyenii (Maca) is one of the few plants that can be cultivated in the harsh climate of the Andes. Its nutritious hypocotyl is traditionally used as food and medicine, and Maca products are increasingly becoming popular in the western world as tonics. This paper describes the first analytical method allowing the determination of the main macamides and macaenes, the marker compounds of L. meyenii. A separation within 35 min was possible by using a C-12 stationary phase, an acidic mobile phase comprising of acetonitrile and water, and raising the column temperature to 40 degrees C. By monitoring the separation at 210 and 280 nm, the markers were detectable as low as 0.40 microg/ml. In order to validate the method, accuracy, precision, linearity, limit of detection and intra/inter day repeatability were determined. The analysis of several commercially available Maca products showed a similar qualitative pattern but significant differences in the quantitative composition. The percentage of total markers in the preparations varied from 0.15 to 0.84%, resulting in daily intakes for the consumer from 1.52 to 14.88 mg, respectively. PMID- 12130862 TI - Synthesis of water-soluble C60-porphyrin hybrid compounds. AB - Water-soluble C60-porphyrin hybrid molecules were first synthesized toward their pharmaceutical applications. PMID- 12130865 TI - Solvents inducing oxidation of hydroxylamines. AB - Hydroxylamines gradually undergo oxidation to their oximes on being dissolved in organic solvent (e.g. methanol). This phenomenon was followed by (1)H-NMR and liquid chromatography-mass spectrometry (LC-MS). The oxidation rate was estimated from the peak area observed on the mass chromatogram at the protonated molecule or fragment ion on LC-atmospheric pressure chemical ionization (APCI)-MS. The results showed that the oxidation rate of hydroxylamines depended on the solvent type. PMID- 12130864 TI - Four new 3,5-cyclosteroidal saponins from Dracaena surculosa. AB - Further search for steroidal compounds contained in Dracaena surculosa (Agavaceae) led to the isolation of two new 3,5-cyclospirostanol saponins (1, 2) and two new 3,5-cyclofurostanol saponins (3, 4). Their structural assignment was established by spectroscopic analysis and a few chemical transformations as (24S,25R)-1beta-[(beta-D-fucopyranosyl)oxy]-6beta-hydroxy-3alpha,5alpha cyclospirostan-24-yl beta-D-glucopyranoside (1), (24S,25R)-1beta-[(beta-D glucopyranosyl)oxy]-6beta-hydroxy-3alpha,5alpha-cyclospirostan-24-yl beta-D glucopyranoside (2), (25S)-1beta-[(beta-D-glucopyranosyl)oxy]-6beta-hydroxy 22alpha-methoxy-3alpha,5alpha-cyclofurostan-26-yl beta-D-glucopyranoside (3), and (25S)-1beta-[(beta-D-fucopyranosyl)oxy]-6beta-hydroxy-22alpha-methoxy 3alpha,5alpha-cyclofurostan-26-yl beta-D-glucopyranoside (4), respectively. PMID- 12130866 TI - Improved preparation of an amino acid type poly(ethylene glycol) derivative. AB - An amino acid type poly(ethylene glycol) is a useful tool for preparation of a bi or multivalent poly(ethylene glycol) hybrid of bioactive peptides, but synthesis is problematic. The amino acid type poly(ethylene glycol) was prepared from poly(oxyethylene)diglycolic acid followed by introduction of a fluorenylmethyloxycarbonyl group. The resulting product could be purified easily by LH-20 column chromatography and HPLC. PMID- 12130867 TI - Improvement in the properties of 3-phenyl-3-trifluoromethyldiazirine based photoreactive bis-glucose probes for GLUT4 following substitution on the phenyl ring. AB - We have developed two novel 3-phenyl-3-trifluoromethyldiazirinyl bis-glucose derivatives to investigate the properties of the adipocyte glucose transporter GLUT4. These compounds were substituted by electron-withdrawing (iodo and nitro) groups on the aromatic ring of 3-phenyl-3-trifluoromethyldiazirine photophore and were found to be more photosensitive than compounds without such substituents. The compounds were used as inhibitors of insulin-stimulated glucose transport activity in order to assess half-maximal inhibition or relative affinity values for GLUT4. The affinities were found to be 60-130 times higher than the parent compound D-glucose. Because of the increased photo-reactivity and high affinity these compounds will be useful in studies directed at further elucidation of GLUT4 function. PMID- 12130868 TI - Mutagenicity of steviol and its oxidative derivatives in Salmonella typhimurium TM677. AB - Stevioside is natural non-caloric sweetner isolated from Stevia rebaudiana BERTONI, which has been used as a non-caloric sugar substitute in Japan. Pezzuto et al. demonstrated that steviol shows a dose-dependent positive response in forward mutation assay using Salmonella typhimurium TM677 in the presence of metabolic activation system (Aroclor induced rat liver S9 fraction). Our studies were carried out to identify the genuine mutagenic active substance from among the eight steviol derivatives. Steviol indicate almost similar levels of mutagenicity under the presence of S9 mixture, as reported by Pezzuto et al. 15 Oxo-steviol was found to be mutagenic at the one tenth the level of steviol itself under the presence of S9 mixture. Interestingly, specific mutagenicity of the lactone derivative under the presence of S9 mixture was ten times lower than that of the lactone derivative without the addition of S9 mixture. PMID- 12130869 TI - Re-revision of the stereo structure of piperidine lactone, an intermediate in the synthesis of febrifugine. AB - The stereo structure of piperidine lactone (3), an intermediate of the antimalarial agent febrifugine ((+)-1) prepared by a synthetic method, was re revised to the cis-form from the trans-form. PMID- 12130870 TI - A national strategic change in treatment policy for rectal cancer--implementation of total mesorectal excision as routine treatment in Norway. A national audit. AB - INTRODUCTION: Rectal cancer surgery has been characterized by a high incidence of local recurrence, an occurrence which influences survival negatively. In Norway there was a growing recognition that local recurrence rates were related to surgeon performance and that surgeons applying a standardized surgical technique in the form of total mesorectal excision could achieve better results. This contrasts with the prevailing argument voiced by many opinion leaders that local recurrence rates and possibly survival rates can only be improved by adjuvant or neoadjuvant treatment strategies. The Norwegian Rectal Cancer Project-initiated in 1993-aimed at improving the outcome of patients with rectal cancer by implementing total mesorectal excision as the standard rectal resection technique. METHODS: This observational national cohort study covers all new patients (3,319) with rectal cancer from a population of 4.5 million treated between November 1993 and August 1997. The main outcome measures were local recurrence, survival, and postoperative mortality and morbidity rates. The technique of total mesorectal excision was compared with conventional surgery. RESULTS: The proportion of patients undergoing total mesorectal excision was 78 percent in 1994, increasing to 92 percent in 1997. The observed local recurrence rate for patients undergoing a curative resection was 6 percent in the group treated by total mesorectal excision and 12 percent in the conventional surgery group. Four-year survival rate was 73 percent after total mesorectal excision and 60 percent after conventional surgery. Postoperative mortality rate was 3 percent and the anastomotic dehiscence rate was 10 percent. Radiotherapy was given to 5 percent and chemotherapy to 3 percent of the patients in the curative resection group. CONCLUSION: A refinement of the surgical resection technique for rectal cancer can be achieved on a national level, the technique of total mesorectal excision can be widely distributed, and surgery alone can give good results. PMID- 12130873 TI - Salvage radical surgery after failed local excision for early rectal cancer. AB - OBJECTIVES: Local recurrence after transanal excision of rectal cancer is often amenable to salvage radical proctectomy, but the long-term results remain unknown. This study was designed to determine the outcome of salvage radical surgery after failed local excision in patients with early rectal cancer. METHODS: We retrospectively reviewed the charts of 29 patients who underwent salvage radical surgery for local recurrence after a full-thickness transanal excision for Stage I rectal cancer. End points included local and distant recurrences and disease-free survival after salvage radical surgery. Comparisons between groups were performed by chi-squared test. RESULTS: Recurrence involved the rectal wall in 26 patients (90 percent) and was purely extrarectal in only 3 (10 percent). Mean time between local excision and radical operation was 26 months. The resection was considered curative in 23 patients (79 percent). The stage of the recurrent tumor was more advanced than the primary tumor in 27 patients (93 percent). At a mean follow-up of 39 (range, 2-147) months after radical surgery, 17 patients (59 percent) remained free of disease. The disease free survival rate was 68 percent for patients with tumors with favorable histology vs. 29 percent for patients with tumors with unfavorable histology. CONCLUSION: Salvage surgery for recurrence after local excision of rectal cancers may not provide results equivalent to those of initial radical treatment. In the present study the poor results of salvage surgery emphasize the importance of appropriate selection of the initial treatment of Stage I rectal cancer. PMID- 12130871 TI - Laparoscopic colorectal surgery for cancer: intermediate to long-term outcomes. AB - PURPOSE: Since 1991, a laparoscopic-assisted resection has been used at the Royal Brisbane Hospital selectively for patients with colorectal cancer. This article audits the intermediate to long-term postoperative complications and cancer follow-up data. METHODS: All patients undergoing a laparoscopic resection for cancer were prospectively followed up with regard to long-term outcomes. RESULTS: One hundred eighty-one patients have been studied. One hundred fifty-four patients had potentially curative procedures performed in the study period. Median follow up was 71 (range, 7-108) months. The overall recurrence rate in this group was 6 percent (21 recurrences). There was one port site recurrence after a potentially curative procedure (0.6 percent) and one port site recurrence after a palliative resection. Perioperative mortality was 1 percent (2 patients). Only six patients suffered an adhesive small-bowel obstruction postoperatively. There was one incisional hernia. Unadjusted five-year median survival data for Australian Clinico-pathological Staging A was 91 percent (3.5 percent recurrence); for Australian Clinico-pathological Staging B, 83 percent (15 percent recurrence); and for Australian Clinico-pathological Staging C, 74 percent (26 percent recurrence). CONCLUSION: In selected patients a laparoscopic resection for colorectal cancer produces acceptable intermediate to long-term oncologic outcomes and a low long-term complication rate. PMID- 12130874 TI - Reconstructive surgery for failed ileal pouch-anal anastomosis: a viable surgical option with acceptable results. AB - PURPOSE: Salvage procedures for failed ileal pouch-anal anastomoses frequently require total reconstruction with a combined abdominal and perineal approach. The aim of this study was to determine the indications for surgery and the outcomes in this group of patients. METHODS: All patients who underwent combined abdominal and perineal ileal pouch-anal anastomosis reconstruction at the Mount Sinai Hospital between 1982 and 2000 were reviewed. Data were collected prospectively in the inflammatory bowel disease database. RESULTS: Sixty-three reconstructive procedures were performed in 57 patients, with a mean age of 33.9 (+/-10.4) years at the time of reconstruction. There were 14 males. The mean follow-up was 69.1 months. The initial indication for ileal pouch-anal anastomosis was ulcerative colitis in 98 percent. The primary indication for reconstruction was pouch vaginal fistula in 21 patients, long outlet in 14, pelvic sepsis in 14, ileoanal anastomotic stricture in 5, pouch-perineal fistula in 2, and chronic pouchitis in 1. The mean operative time was four hours (+/-1.1), the average blood loss was 500 mL (+/-400), and the average length of stay was 10.3 days (+/-4.6). All patients had a diverting ileostomy. Forty-two (73.6 percent) of the patients have a functioning pouch. Seven (12.3 percent) patients have had their pouch excised. The ileostomy has not yet been closed in 8 (14 percent) patients; 3 of these patients are awaiting closure, whereas the remaining 5 have a permanently defunctioning ileostomy. Eighty-nine percent have ten or fewer bowel movements per day. No patients are incontinent of stool during the day, whereas two patients are incontinent at night. Seventeen percent complain of frequent urgency. Despite this, more than 80 percent rate their physical and psychological health as good to excellent. CONCLUSION: Reconstructive pouch surgery has a high success rate in experienced hands. The functional results in those whose pouch is in use are good. PMID- 12130875 TI - Teenagers with familial adenomatous polyposis: what is their risk for colorectal cancer? AB - PURPOSE: Familial adenomatous polyposis is a general growth disorder caused by highly penetrant germline mutations in the tumor suppressor gene APC. The major manifestation of these mutations is colorectal adenomatous polyposis, which, if untreated, leads to early development of colorectal cancer. To prevent this from happening, endoscopic screening of at-risk family members begins early in the second decade of life. Patients with adenomas are offered surgery sometime in that decade. There is a concern about the risk of cancer in teenagers if such surgery is deferred. We conducted this study to investigate that risk. METHODS: A brief survey was sent by facsimile or mail to all familial adenomatous polyposis registries affiliated with the Leeds Castle Polyposis Group. This questionnaire asked for the number of teenage or younger patients in the registry diagnosed with invasive colorectal carcinoma. Other questions addressed the stages and treatment of the tumors and the outcome of their treatment. Patients with carcinoma-in-situ or intramucosal carcinoma were excluded. RESULTS: Replies were received from 26 of 52 registries, but not all questions were answered by all registries. There were 14 patients identified as having invasive colorectal cancer younger than 20 years, the youngest of whom was 9 and the oldest 19. Two patients had two cancers each. Three patients were diagnosed at surgery, and seven were diagnosed when they presented with symptoms. Of the 13 cancers that had staging information, 8 were T1N0M0; 1 was T2N0M0; 2 were TxN1M0; 1 was T3N0M0; and 1 was TxNxM1. Only one patient died of their colorectal cancer. CONCLUSION: Cancer occurs rarely in familial adenomatous polyposis patients younger than 20 years, and only 1 case was reported younger than 15 years. Surgery for colorectal polyposis usually can be deferred safely until at least the age of 15, unless suspicious lesions are found. PMID- 12130876 TI - Functional outcome, quality of life, and complications after ileal pouch-anal anastomosis in selected septuagenarians. AB - PURPOSE: Concerns about morbidity and functional outcome have lead some authors to suggest that ileal pouch-anal anastomosis should not be performed in older patients. This article evaluates the outcome of selected septuagenarians undergoing ileal pouch-anal anastomosis at this institution. METHODS: Seventeen of 1,911 patients undergoing ileal pouch-anal anastomosis for ulcerative colitis were older than the age of 70 at the time of surgery. Functional outcome, quality of life, and manometric data were assessed prospectively, whereas complications were assessed by chart review. RESULTS: There was one mortality related to sepsis after small-bowel obstruction and one reoperation at 18 months for pelvic abscess. Minor complications occurred in five patients. Median (interquartile range) quality of life and health and levels of energy and happiness (scored out of 10) were 9 (7-10), 9 (7-10), 8 (5-10), and 9.5 (7-10), respectively. Medical Outcomes Study Short Form 36 quality of life scores were not different from those for the healthy population older than 65 years. There was complete continence in 38 percent, rare incontinence in 12 percent, and some incontinence in 50 percent. Nobody was usually or always incontinent. Overall, 82 percent would undergo pouch surgery again, and 89 percent would recommend it to others. CONCLUSIONS: Ileal pouch-anal anastomosis is an acceptable surgical option for selected healthy, motivated septuagenarians with ulcerative colitis who are eager to preserve fecal continence. PMID- 12130878 TI - T-level downstaging and complete pathologic response after preoperative chemoradiation for advanced rectal cancer result in decreased recurrence and improved disease-free survival. AB - PURPOSE: Preoperative chemoradiation therapy is used widely in the treatment of rectal cancer. The predictive value of response to neoadjuvant remains uncertain. We retrospectively evaluated the impact of response to preoperative and, specifically, of T-level downstaging, nodal downstaging, and complete pathologic response after chemoradiation therapy on oncologic outcome of patients with locally advanced rectal cancer. METHODS: There were 88 patients with ultrasound Stage T3/T4 midrectal (n = 37) and low rectal (n = 51) cancers (63 males; mean age 62.6 years). All patients were treated by preoperative 5-fluorouracil-based chemotherapy and pelvic radiation followed by surgical resection in six weeks or longer (56 sphincter-preserving resections). RESULTS: T-level downstaging after neoadjuvant treatment was demonstrated in 36 (41 percent) of 88 patients, and complete pathologic response was observed in 16 (18 percent) of the 88. Of the 42 patients with ultrasound-positive nodes, 27 had no evidence of nodal involvement on pathologic evaluation (64 percent). The overall response rate (T-level downstaging or nodal downstaging) was 51 percent. At a median follow-up of 33 months, 86.4 percent of patients were alive. The overall recurrence rate was 10.2 percent (three patients had local and six had metastatic recurrences). Patients with T-level downstaging and complete pathologic response were characterized by significantly better disease-free survival (P = 0.03, P = 0.04) and better overall survival (P = 0.07, P = 0.08), according to Wilcoxon's test comparing Kaplan-Meier survival curves. None of the patients with complete pathologic response developed recurrence or died during the follow-up period. CONCLUSION: T level downstaging and complete pathologic response after preoperative chemoradiation therapy followed by definitive surgical resection for advanced rectal cancer resulted in decreased recurrence and improved disease-free survival. Advanced rectal cancers that undergo T-level downstaging and complete pathologic response after chemoradiation therapy may represent subgroups that are characterized by better biologic behavior. PMID- 12130879 TI - Surgeon specialty is associated with outcome in rectal cancer treatment. AB - PURPOSE: The aim of this study was to determine the effect of surgeon specialty on disease-free survival and local control in patients with adenocarcinoma of the rectum. Patients underwent curative treatment with neoadjuvant external beam radiotherapy and proctectomy by colorectal surgeons and noncolorectal surgeons. METHODS: The records of 384 consecutive patients treated by colorectal surgeons (n = 251) and noncolorectal surgeons (n = 133) from 1977 to 1995 were reviewed independently by physicians in the Division of Radiation Oncology. Local recurrence was defined as pelvic recurrence occurring in the presence or absence of distant metastatic disease. RESULTS: The study population comprised 213 males, mean age 64 (range, 19-97) years. Preoperative radiotherapy was delivered as 4,500 cGy in 25 fractions six to eight weeks before surgery (n = 293) or 2,000 cGy in 5 fractions immediately before surgery (n = 91). Concurrent preoperative chemotherapy was given to 14 patients, postoperative chemotherapy to 55. Overall actuarial disease-free survival and local control rates were 74 and 90 percent, respectively, at five years. Actuarial disease-free survival and local control rates at five years were 77 and 93 percent for colorectal surgeons vs. 68 and 84 percent for noncolorectal surgeons (P < or = 0.005 for both, Tarone-Ware). Multivariate analysis revealed that pathologic stage and background of the surgeon were the only independent predictors of disease-free survival (both P < or = 0.006, Cox proportional hazards) and that pathologic stage, background of the surgeon, and proximal location of the tumor were independent predictors of local control (all P < or = 0.02, Cox proportional hazards). Radiation dose and use of chemotherapy were not significant factors. Sphincter preservation was more common by colorectal surgeons (131/251, 52 percent) than noncolorectal surgeons (40/133, 30 percent; P = 0.00004, Fisher's exact test, two-tailed). CONCLUSION: Good outcome for patients with adenocarcinoma of the rectum who undergo neoadjuvant external beam radiotherapy and proctectomy is associated with subspecialty training in colon and rectal surgery. PMID- 12130880 TI - Radio-frequency energy delivery to the anal canal for the treatment of fecal incontinence. AB - PURPOSE: In this prospective study we investigated the feasibility, safety, and efficacy of radio-frequency energy delivery deep to the mucosa of the anal canal for the treatment of fecal incontinence. METHODS: We studied ten patients with fecal incontinence of varying causes. All patients underwent anoscopy, anorectal manometry, endorectal ultrasound, and pudendal nerve terminal motor latency testing at baseline and six months. The Cleveland Clinic Florida scale for fecal incontinence (Wexner, 0-20), fecal incontinence-related quality of life score, and Short Form 36 were administered at baseline, 1, 2, 3, 6, and 12 months. Using conscious sedation and local anesthesia, we delivered temperature-controlled radio-frequency energy via an anoscopic device with multiple needle electrodes to create thermal lesions deep to the mucosa of the anal canal. RESULTS: Ten females (age, 55.9 +/- 9.2 years; range, 44-74) were enrolled and treated. Median discomfort by visual analog scale (0-10) was 3.8 during and 0.9 two hours after the procedure. Bleeding occurred in four patients (14-21 days after procedure), spontaneous resolution (n = 3) and anoscopic suture ligation (n = 1). At 12 months, the median Wexner score improved from 13.5 to 5 (P < 0.001), with 80 percent of patients considered responders. All parameters in the fecal incontinence-related quality of life were improved (lifestyle (from 2.3 to 3.4), coping (from 1.4 to 2.7), depression (from 2.2 to 3.5), and embarrassment (from 1.3 to 2.8); P < 0.05 for all parameters). Protective pad use was eliminated in five of the seven baseline users. At six months, there was a significant reduction in both initial and maximum tolerable rectal distention volumes. Anoscopy was normal at six months. CONCLUSION: Radio-frequency energy delivery to the anal canal for treatment of fecal incontinence is a new modality that, in this study group, safely improved Wexner and fecal incontinence-related quality of life scores, eliminated protective pad use in most patients, and improved patient quality of life. PMID- 12130881 TI - Pudendal nerve "complete" motor latencies at four different levels in the anal sphincter system in young adults. AB - PURPOSE: Pudendal nerve complete motor latency, or the sum of the conduction time from the root of the sacral nerve to the end of the pudendal nerve and the synaptic delay, varied in length (from shortest to longest) in the puborectalis, the deep external anal sphincter, and the superficial/subcutaneous external anal sphincter, in that order, for middle-aged people. The aim of this study was to elucidate whether such a phenomenon was physiologic or pathologic. METHODS: In 20 young adults (21.9 +/- 1.37 years old, 10 females), pudendal nerve complete motor latencies were measured after magnetic stimulation to the sacral region. Electromyographic recordings were taken at depths of 5, 3.8, 2.6, and 1.5 cm from the perineal skin using a needle electrode and at 3 cm from the anal verge using surface electrodes within the anal canal. The data were compared with the data of the middle-aged cohort (65.4 +/- 7.70 years old) in our previous study. RESULTS: The pudendal nerve complete motor latencies were 3.85 +/- 1.24 ms at 5 cm, 3.97 +/- 1.25 ms at 3.8 cm, 5.41 +/- 2.42 ms at 2.6 cm, 9.98 +/- 4.01 ms at 1.5 cm, and 3.45 +/- 0.52 ms while using surface electrodes. The pudendal nerve complete motor latencies at 5, 3.8, and 2.6 cm were significantly shorter in the young adults than in the middle-aged subjects. The pudendal nerve complete motor latency using surface electrodes was significantly shorter than the pudendal nerve complete motor latency at 2.6 and 1.5 cm (mean +/- standard deviation). CONCLUSIONS: Because pudendal nerve complete motor latency was progressively longer at 5, 3.8, 2.6, and 1.5 cm, in that order, in young adults as well as in middle-aged people, this phenomenon was considered to be physiologic and may be mechanically reasonable and safe in shutting the anal canal and might be useful for milking the residual mucus out of the anal canal to prevent soiling. Aging disturbed the innervation of the upper three levels of the anal sphincter system. Pudendal nerve complete motor latency using intra-anal surface electrodes approximated that at the highest of the anal sphincters. PMID- 12130883 TI - Effects of preoperative radiotherapy for primary resectable rectal adenocarcinoma on male sexual and urinary function. AB - PURPOSE: The purpose of this study was to determine whether preoperative radiotherapy had an influence on the urinary and sexual function of patients having a sphincter-saving, nerve-preserving total mesorectal excision. METHODS: Urinary and sexual function of male patients undergoing sphincter-saving, nerve preserving total mesorectal excision for primary resectable rectal carcinoma between January 1998 and December 1999 were evaluated retrospectively. Assessment was by standardized questionnaires. RESULTS: Fifty male patients met the inclusion criteria. Three patients had died (hepatic metastases), and five were living outside the European community and could not be contacted. Sixteen patients underwent preoperative radiotherapy (Group 1), and 26 patients were not treated preoperatively (Group 2). There was no perioperative mortality. Age, tumor stage, and localization of the tumor were comparable. Median follow-up was 20 months. Urinary function was not significantly different. One patient in Group 1 and 2 patients in Group 2 were impotent before surgery. All remaining patients in Group 2 (n = 24) and 11 of 15 remaining patients in Group 1 were sexually active (P = 0.016). All sexually active patients (n = 24) in Group 2 and 9 of the 11 sexually active patients in Group 1 have normal ejaculation (P = 0.09). CONCLUSION: Preoperative radiotherapy for primary resectable rectal carcinoma treated by total mesorectal excision with autonomic nerve preservation may impair male sexual function. PMID- 12130882 TI - Rectoanal reflex parameters in incontinence and constipation. AB - PURPOSE: The transient relaxation of the internal anal sphincter in response to rectal distention is believed to play an important role in the continence mechanism. Most anorectal physiology laboratories merely report the rectoanal inhibitory reflex as being either present or absent. This study aimed to assess the parameters of the rectoanal inhibitory reflex in incontinent and constipated patients and healthy control subjects, in an attempt to analyze differences in internal anal sphincter function in these groups. We analyzed each response of the internal anal sphincter to rectal distention with progressively increasing volumes of air at a single site (proximal anal canal). METHODS: Fifty-five constipated and 99 incontinent patients and healthy control subjects underwent manometry. Various parameters of the rectoanal inhibitory reflex were analyzed, and percentage sphincter relaxation was calculated at each volume at which rectoanal inhibitory reflex occurred. RESULTS: There was no difference in the volume of rectal distention required to elicit sensation (P = 0.626) or the rectoanal inhibitory reflex (P = 0.371) in the three groups. There was a significant correlation between the volume required to elicit the rectoanal inhibitory reflex and that at which sensation was first felt only in the incontinent (P = 0.0001) group. Significantly greater sphincter relaxation was seen at each volume (P = 0.001) in the incontinent as compared with the constipated patients. With progressive rectoanal inhibitory reflex, consistently progressive increases in internal anal sphincter relaxation were found only in the incontinent group. This consistent relationship was not seen in the constipated patients or in healthy control subjects. CONCLUSIONS: Assessment of various parameters of the rectoanal inhibitory reflex yielded important information regarding the continence mechanism. Altered responses of the internal anal sphincter in anorectal disorders plays a role in the associated physiologic impairment. This may have significant clinical implications with regard to sphincter-saving resections. PMID- 12130885 TI - Adults in a high-risk area are unaware of the importance of colorectal cancer: a telephone and mail survey. AB - PURPOSE: There are few studies assessing public awareness of colorectal cancer even in developed countries. This questionnaire and telephone-based survey aimed to determine the degree of awareness of colorectal cancer among adults in a high risk population with a high level of literacy. METHODS: Two thousand randomly selected adults living in Singapore were invited to answer a questionnaire by telephone (T, n = 1,000) or by returned mail (M, n = 1,000). The questionnaire consisted of 20 questions relating to epidemiology, presentation, screening, and management of colorectal cancer. RESULTS: There was a compliance rate of 85.2 percent for telephoned subjects, but only 21.5 percent of mailed subjects returned a completed questionnaire. Only 2.7 percent of T and 1.4 percent of M named colorectal cancer as a fatal disease. Only 49.6 percent of T and 60.9 percent of M were aware that the colon and rectum are part of the intestines. A minority (T 11.7 percent, M 35.8 percent) were aware of screening as an important means against developing colorectal cancer. A large proportion (T 46.5 percent, M 34.9 percent) were unable to name even one symptom of colorectal cancer. There was low awareness of causative factors for colorectal cancer (T 39.6 percent, M 28.4 percent). A proportion (T 24.8 percent, M 28.4 percent) believed that colorectal cancer affects people less than 40 years of age. Newspapers and popular magazines exert far more influence (T 58.2 percent, M 52.1 percent) than television, the internet. or doctors in educating the public about colorectal cancer. Respondents with higher education level (junior college and above), as well as those with a positive family history, tend to score better in the knowledge section. CONCLUSIONS: Knowledge of colorectal cancer among the surveyed population was poor despite a relatively high incidence of the disease in Singapore. Public education regarding colorectal cancer is best done via newspapers and magazines. PMID- 12130884 TI - Defunctioning stoma in low anterior resection with colonic pouch for rectal cancer: a comparison between two hospitals with a different policy. AB - PURPOSE: The aim of this study was to compare surgical outcome, after low anterior resection for rectal cancer with colonic J-pouch, at two departments with a different policy regarding the use of a routine diverting stoma. METHODS: A total of 161 consecutive patients with invasive rectal carcinomas operated on between 1990 and 1997 with a total mesorectal excision and a colonic J-pouch were included in the study. Eighty patients were operated on in a surgical unit using routine defunctioning stomas (96 percent), whereas 81 were operated on in a department in which diversion was rarely used (5 percent). Recorded data with respect to surgical outcome were analyzed and compared. RESULTS: There was no difference between the two centers in postoperative mortality in connection with the primary resection and subsequent stoma reversal (3.7 vs. 3.8 percent). No significant difference could be found in the number of patients with pelvic sepsis (anastomotic leaks; 9 vs.12 percent). Surgical outcome in patients with pelvic sepsis was also similar. The frequency of reoperations associated with the anterior resection and subsequent stoma reversal was identical (14 percent). The total hospital stay (primary operation and stoma reversal) was significantly longer with than without a routine stoma (17 (range, 2-59) vs. 12 (range, 5-55) days, respectively; P < 0.001). CONCLUSION: This study suggests that the routine use of diversion does not protect the patient from anastomotic complications or pelvic sepsis and its use requires a second admission for closure. PMID- 12130887 TI - Long-term follow-up after first acute episode of sigmoid diverticulitis: is surgery mandatory?: a prospective study of 118 patients. AB - PURPOSE: This study was designed to evaluate the long-term natural history of sigmoid diverticulitis in patients treated nonoperatively after a first acute episode and to assess the role of elective colectomy. METHODS: Between 1986 and 1991, 144 patients were admitted for acute diverticulitis diagnosed by abdominal computed tomography and had a successful nonoperative treatment. Remote complications (persisting or recurring diverticulitis) were also diagnosed by computed tomography. Patients had a poor outcome if they had one of these complications. Diverticulitis was graded mild or severe on computed tomography according to Ambrosetti's criteria. We determined statistically whether young age (< or =50 years old) and severe diverticulitis were risk factors for a poor outcome. RESULTS: One hundred eighteen patients with a contributive computed tomographic scan at admission were followed up. Median age was 63 (range, 23-93) years, with a median follow-up of 9.5 (range, 0.2-13.8) years. Eighty patients had no complications, and 38 had remote complications. The incidence of remote complications was the highest (54 percent at 5 years) for young patients with severe diverticulitis on computed tomography and the lowest (19 percent at 5 years) for older patients with mild disease. Young age and severe diverticulitis taken separately were both statistically significant factors of poor outcome (P = 0.007 and P = 0.003, respectively), although age was no longer significant after stratification for disease severity on computed tomography (P = 0.07). Twenty four patients died. The cause of death was unrelated to diverticulitis in 21 cases and unknown in the remaining 3. CONCLUSIONS: We propose that after a first acute episode of diverticulitis treated nonoperatively, elective colectomy should be offered to young patients (< or =50 years old) with severe diverticulitis on computed tomography. PMID- 12130886 TI - Prevalence of perforated sigmoid diverticulitis is increasing. AB - INTRODUCTION: The population of Finland is aging fast, and dietary fiber consumption has decreased during the past few decades; the prevalence of sigmoid diverticular perforation can therefore be anticipated to increase. This study presents our experience concerning the outcome of 133 patients admitted to a university hospital for diverticular perforation during a 15-year period. METHODS: One hundred thirty-three patients admitted into our hospital for sigmoid diverticular perforation during the 15-year period from 1986 to 2000 were identified using a computer database. Clinical data were reviewed from the database and the patients' records. RESULTS: The annual prevalence of perforated sigmoid diverticulitis is increasing. It was 2.4 per 100,000 in the year 1986 and 3.8 per 100,000 in the year 2000. The resection rate was 90 percent; after resection, 45 primary anastomoses, 75 Hartmann's colostomies, and 1 covering colostomy were performed. The overall complication rate was 32 percent, without any significant difference between the procedures. Of the clinical variables, the Mannheim Peritonitis Index scores (P = 0.0088) and the number of previous hospital treatments (P = 0.035) correlated with postoperative complications. Overall mortality was 9 percent, without any significant difference between the procedures. Of the clinical variables, the Mannheim Peritonitis Index scores correlated with mortality. Of the 12 patients who died, 11 had Mannheim Peritonitis Index scores of 21 or more (P = 0.0001). Forty-five percent of the colostomies have been closed. CONCLUSIONS: The prevalence of perforated sigmoid diverticulitis is increasing in northern Finland. Mannheim Peritonitis Index score can be used in predicting the outcome of patients admitted for perforated sigmoid diverticulitis. PMID- 12130888 TI - Hyperbaric oxygen ameliorates bacterial translocation in rats with mechanical intestinal obstruction. AB - PURPOSE: The aim of this study was to demonstrate bacterial translocation after experimentally induced intestinal obstruction as well as investigate the preventive effects of hyperbaric oxygen on obstruction-induced bacterial translocation in rats. METHODS: Forty Wistar-albino male and female rats were used. Although no procedure was done in the control group (n = 8), hyperbaric oxygen treatment under 2.5 atm absolute for 90 minutes daily was applied for two days in the hyperbaric oxygen group (n = 8). In the sham group (n = 8), after laparotomy the small bowel was only handled gently, and tissue sampling was done 48 hours later. In the obstruction group (n = 8) the ileum was ligated by 5-0 polypropylene just 5 cm proximal to the ileocecal valve. In the obstruction and hyperbaric oxygen group (n = 8), after obstruction hyperbaric oxygen treatment was applied. Forty-eight hours after the procedures, tissue samples from small bowel, mesenteric lymph nodes, spleen, and liver were taken and 1 ml of blood from the portal vein was withdrawn. All samples were cultured for microbiologic examination. RESULTS: Hyperbaric oxygen treatment significantly reduced the endogenous bacterial overgrowth in the small intestine of normal rats. Endogenous bacteria in the small intestine were significantly increased in the obstruction group, and the presence of bacterial overgrowth was proven by bacterial presence on mesenteric lymph nodes, spleen, liver, and blood. Hyperbaric oxygen treatment significantly reduced the endogenous bacterial overgrowth in the small intestine and prevented the bacterial translocation almost completely in obstruction induced rats. CONCLUSIONS: Intestinal obstruction causes bacterial overgrowth and translocation. Hyperbaric oxygen treatment prevents the bacterial translocation effectively. PMID- 12130889 TI - Fecal incontinence induced by spontaneous internal anal sphincter relaxation: report of a case. AB - It has been previously suggested that an increase in the frequency of internal anal sphincter relaxations may cause fecal incontinence in patients in whom a structural lesion of the anal sphincter or its nerve supply is not ruled out. We here report a case of fecal incontinence in which the sphincter and its innervation was not damaged, and prolonged recordings of anal resting pressure detected frequent and prolonged internal anal sphincter relaxations. Moreover, a spontaneous improvement in fecal incontinence occurred at the same time as a reduction in the frequency and duration of internal anal sphincter relaxations. This case suggests that prolonged recordings of anal resting pressure are advisable in incontinent patients without detectable lesions of the anal sphincter or its nerve supply to detect any increase in the frequency of internal anal sphincter relaxations as a possible cause of fecal incontinence. PMID- 12130890 TI - Inflammatory bowel perforation during immune restoration after one year of antiretroviral and antituberculous therapy in an HIV-1-infected patient: report of a case. AB - PURPOSE: This article reports an unusual presentation of bowel perforation. METHODS: We report the case of a 30-year-old HIV-infected male who suffered from an advanced state of CD4 cell depletion (29 CD4 cells per 106/l). Abdominal pain and diarrhea led to further examinations. RESULTS: Colonoscopy revealed a severe tuberculous ileocecal inflammation. Tuberculosis and HIV infection were treated. The patient's response to antiretroviral therapy was excellent. After 11 months of potent antiretroviral treatment and 12 months of antituberculous therapy he suffered from acute abdominal pain with fever and ileus. Laparotomy revealed two intestinal perforations of the jejunum and inflammation of the whole ileocecal region. CONCLUSION: Immunopathologic reactions caused by immune restoration are novel presentations of highly active antiretroviral treatment as shown here. The presented patient is an unusual case with a very late onset of inflammatory response, which led to intestinal perforation. PMID- 12130892 TI - Tamponade of presacral venous hemorrhage. PMID- 12130891 TI - Ectopic prostatic tissue of the anal canal presenting with rectal bleeding: report of a case. AB - PURPOSE: Ectopic prostatic tissue at various sites within and outside the genitourinary system has been reported previously. A case of ectopic prostatic tissue located in the anal canal causing rectal bleeding is presented. METHOD: The patient was referred to our clinic with rectal bleeding. At rectal examination a bleeding sessile polypoid mass 2.5 cm in size was found in anal canal and removed surgically. RESULTS: Histopathologic and immunohistochemical staining of the specimen confirmed the prostatic nature of the tissue. CONCLUSION: Prostatic heterotopia is significant in several respects. Either it may be an important cause of hematuria or unusually, as in our case, it may cause rectal bleeding. In addition, ectopic tissue may be endoscopically confused with malignancy in either urinary or lower gastrointestinal system. This and other reports may disclose the genesis and significance of this peculiar tissue remnant. PMID- 12130893 TI - Tamponade of presacral venous hemorrhage. PMID- 12130897 TI - Evidence-based treatment of eating disorders. AB - Anorexia nervosa and bulimia nervosa are common problems facing adolescents and young adults. Treatment of these disorders poses a challenge to health care providers given the general paucity of clinical trials to guide management. There is evidence to support the use of CBT as well as psychopharmacotherapy to decrease binge and purge behaviors in bulimia nervosa. Significantly fewer trials have examined the efficacy of such therapies for anorexia nervosa. Short-term trials appear promising regarding potential treatments for bone loss in anorexia nervosa. The role of exercise in the management of anorexia nervosa remains controversial and begs future investigative efforts. PMID- 12130896 TI - Risk factors for thromboembolism in teens: when should I test? AB - The discoveries of the factor V Leiden mutation and the prothrombin gene variant 20210 in the last decade have markedly contributed to the understanding of the molecular pathophysiology of inherited risk factors for thrombophilia. Population studies in the adult literature have shown that although the overall prevalence of these defects is low, affected individuals are at increased risk of thrombosis particularly if acquired risk factors for thrombosis are also present. The use of combined hormonal oral contraceptive pills is a well-known acquired risk factor, and recent studies have shown significant increased risk of thrombosis for women who carry the factor V Leiden mutation and use oral contraceptive pills. Despite this significant increased risk, mass screening of asymptomatic women for factor V Leiden prior to prescribing oral contraceptive pills is not a cost-effective use of health care dollars and could result in unnecessarily preventing many women from the contraceptive and noncontraceptive benefits of this medication. Instead, clinicians can use thoughtful screening questions to identify potentially high-risk patients for thrombophilia and consider testing for inherited risk factors on a case-specific basis. PMID- 12130898 TI - Preparing adolescent patients for college. AB - Adolescents making the transition to college should have a thorough medical evaluation during the year prior to matriculation. In addition to required and recommended immunizations and tests, a comprehensive history and physical examination is important. Screening for substance abuse, sexual activity, depression, and suicidality is needed with appropriate anticipatory guidance, examinations, and treatment, if indicated. The teen should also be counseled on stress, sleep, and self-care, with information on when to seek medical care. The adolescent should be encouraged to continue communications with the primary care clinician during college. While respecting the adolescent's confidentiality, it is important that the physician communicate all significant medical and psychiatric health information to the college health center before the adolescent arrives on campus. PMID- 12130899 TI - Teaching adolescent medicine in the office setting. AB - Despite recent educational improvements in adolescent medicine, challenges remain. Residents in pediatrics are required to have a 1-month block rotation in adolescent medicine, and residents in family medicine and internal medicine are strongly encouraged to receive training in adolescent health issues. Meeting the educational challenges for all of these residents can be a daunting task but can be accomplished with proper curricular development, utilization of learner centered materials, and effective planning of the clinical education program. A significant amount of adolescent teaching occurs in the ambulatory setting. Clinical preceptors should be familiar with teaching approaches such as the 1 minute preceptor and should regularly reflect on their own teaching styles and skills. Proper learner feedback is crucial to any clinical teaching endeavor. PMID- 12130901 TI - Congenital melanocytic nevi: an update for the pediatrician. AB - While melanoma is uncommon in childhood, recent evidence suggests that its incidence is increasing in both adolescence and adulthood. Risk factors may be identifiable during childhood and include congenital nevi. Large congenital nevi carry a significantly increased risk for the development of melanoma, both cutaneous and extracutaneous. This risk appears to be greatest during early childhood. Large congenital nevi, particularly those overlying the head and neck, may also be associated with neurocutaneous melanosis. Symptomatic neurocutaneous melanosis, although rare, carries a poor prognosis. Conversely, asymptomatic neurocutaneous melanosis may be more common than previously recognized. For the most part, large congenital nevi are treated with primary excisions and closures, assisted by tissue-expansion techniques and skin grafting. Until the associated risks are better defined, therapy of small congenital nevi should be individualized, with consideration given to additional melanoma risk factors. PMID- 12130902 TI - Childhood psoriasis. AB - Psoriasis commences by the age of 15 years in one third of patients and is an important childhood disorder. Various human leukocyte antigen associations are important in predisposition to psoriasis, and streptococcal disease is important in its precipitation and exacerbation. The disorder takes many clinical forms, with guttate lesions and anogenital and facial involvement being particularly prominent in the childhood age group. Pustular psoriasis and psoriatic arthropathy are rare in this group of patients. It is important to take the age of the patient into account when making treatment choices. PMID- 12130903 TI - Atopic dermatitis: review 2000 to January 2001. AB - A review on atopic dermatitis is given, considering as basic information articles published over the period of February 1, 2000 to January 31, 2001. Atopic dermatitis is a chronic, inflammatory, primarily genetic-determined skin disease of which the cause is unknown. Its prevalence is rising in the industrialized countries, and no one knows why. The hygienic theory is most promising. Although most cases of atopic dermatitis are mild, the disease may be severe and widespread, with much impact on morbidity and social life. Mild cases usually clear and compromise 60% of the cases. Atopic dermatitis may always turn back. The clinical features are age related. Recently, a new subgroup was noted by several dermatologists in different parts of the world, consisting of people who suffer from atopic dermatitis for the first time at adult age. Food allergy, intolerance, and diet are still controversial and play a role especially in children until the age of 5 years. Diagnostic tests, such as the Atopy Patch Test, using food allergens, adds 10% or more positives and imitates the late phase clinical manifestations. The Atopy Patch Test is still experimental as a diagnostic tool and has shortcomings, like difficulty in reading. Financial costs of treating and caring for atopic dermatitis may be high, stressing the importance of health care. PMID- 12130904 TI - Topical immunomodulators for atopic dermatitis. PMID- 12130905 TI - Molecular advances in genetic skin diseases. AB - The genes for several genetic skin diseases have been identified in recent years. This development improves diagnostic capabilities and genetic counseling, and investigators can now turn to the molecular mechanisms involved in the pathogenesis of these diseases. The identification of the causative genes has led to the generation of mouse models for some genetic skin diseases. A study of the keratin 10 deficient mouse, a model for epidermolytic hyperkeratosis, and a mouse model for Bloom syndrome are reviewed in this article. Several studies also evaluate the relation between genotype and phenotype. In this article, the clinical findings and molecular advances in tuberous sclerosis complex, neurofibromatosis type 1, Bloom syndrome, epidermolytic hyperkeratosis, X-linked ichthyosis, Netherton syndrome, and Hermansky-Pudlak syndrome are reviewed. PMID- 12130906 TI - Pemphigus vulgaris in an adolescent. PMID- 12130908 TI - Metabolic effects of growth hormone in the child and adolescent. AB - During the year 2000, several original studies were published regarding the metabolic effects of growth hormone therapy in pediatric patients. Pharmacologic doses of growth hormone were rarely associated with abnormalities in glucose tolerance in children with intrauterine growth retardation and Turner syndrome; however, serum insulin levels were elevated. A report from the Pharmacia International Growth Study database suggested a possible increase in type 2 diabetes in growth hormone-treated patients, indicating the need for continued surveillance for this condition. Growth hormone therapy increased markers of bone turnover and bone mineral density in children with chronic renal failure and Prader-Willi syndrome. In Prader-Willi syndrome, 2 years of growth hormone therapy also induced a sustained decrease in body fat, improvement in strength and physical skills, and increased lean body mass. Serum leptin, a reflection of body fat, declined with growth hormone therapy in a dose-dependent manner in intrauterine growth retardation children; the magnitude of the decline correlated with linear growth response. Skin is a target organ for growth hormone in children; growth hormone increased dermal thickness and reduced skin stiffness in growth hormone-deficient children. Reassuring data were published regarding the risk of tumor recurrence and mortality in children with brain tumors treated with growth hormone. Growth hormone administered to short children prior to kidney transplantation did not have adverse effects on subsequent graft survival or number of rejection episodes. PMID- 12130910 TI - Maternal hypothyroidism and cognitive development of the offspring. PMID- 12130909 TI - Expanded spectrum of recombinant human growth hormone therapy. AB - The efficacy of recombinant human growth hormone in the treatment of growth hormone deficiency is well established. In recent years, the use of recombinant human growth hormone as a therapeutic modality has greatly increased and has expanded beyond the realm of replacement for growth hormone deficiency. Recombinant human growth hormone has been employed to ameliorate growth failure in multiple other disorders. For some, like Turner syndrome, recombinant human growth hormone has become the standard of care. For others, the ultimate benefit of recombinant human growth hormone remains to be determined. Although recent investigations provide encouraging short-term data, it is important to recognize that the impact of recombinant human growth hormone therapy on adult height has not been established in a number of conditions. PMID- 12130911 TI - Thyroxine supplementation in preterm infants: critical analysis. PMID- 12130912 TI - The ex-utero intrapartum treatment. AB - Advances in prenatal diagnosis, combined with a better understanding of the natural history of prenatally diagnosed anomalies, are providing increasing opportunities to consider fetal intervention in selected cases of life threatening malformations. Accurate prenatal diagnosis can now accurately identify fetal pathophysiology that poses an immediate threat to the life of the newborn infant on separation from the placental circulation. In this circumstance, the ex-utero intrapartum treatment (EXIT) procedure, which maintains intrapartum uteroplacental support, can be life saving. The most common indications for the EXIT procedure are fetal lesions causing extrinsic or intrinsic airway obstruction. However, fetuses with other anomalies that may compromise neonatal resuscitation can also benefit from this approach. The EXIT procedure differs significantly from a cesarean delivery, and caution must be taken to avoid maternal morbidity. As with all endeavors involving maternal-fetal intervention, a team approach is crucial to ensure accurate diagnosis and optimal perinatal management. PMID- 12130913 TI - Support of respiratory failure in the pediatric surgical patient. AB - Severe respiratory failure in newborn and pediatric patients is associated with significant morbidity and mortality. Basic science laboratory investigation has led to advances in the understanding of ventilator-induced lung injury and in optimizing the supportive use of conventional ventilation strategies. Over the past few years, progress has been made in alternative therapies for supporting children and adults with severe respiratory failure. This review will focus on recent laboratory and clinical data regarding the techniques of lung protective ventilator strategies, inhaled nitric oxide, liquid ventilation, and extracorporeal life support (ECLS, ECMO). Some of these modalities are commonplace, while others may have much to offer the pediatric clinician if their benefit is clearly demonstrated in future clinical trials. PMID- 12130914 TI - Nutritional support of the critically ill child. AB - The pediatric metabolic response to injury and operation is proportional to the degree of stress and causes an increase in the turnover of proteins, fats, and carbohydrates. Thereby, substrates are made readily available for the immune response and wound healing. Because this process requires energy, the resting energy expenditure of ill patients increases. Whole-body protein degradation rates are elevated out of proportion to synthetic rates, and negative protein balance also ensues. Neonates and children are particularly susceptible to the loss of lean body mass and its attendant increased morbidity and mortality caused by an intrinsic lack of endogenous stores and greater baseline requirements. An appropriately designed mixed fuel system of nutritional support replete in protein does not quell this metabolic response but can result in anabolism and continued growth in ill children. In addition, the use of adequate analgesia and anesthesia is a readily available and proven means of reducing the magnitude of the catabolism associated with operation and injury. Finally, as hormonal- and cytokine-mediated metabolic alterations are better understood, therapeutic interventions may become available to directly modulate the metabolic response to illness, thus potentially further improving clinical outcome in pediatric surgical patients. PMID- 12130915 TI - Multiple trauma: liver and spleen injury. AB - The management of hepatic and splenic injuries in childhood has evolved over the past 30 years from prompt operation upon recognition of injury to nonoperative management in the large majority of children. Many aspects of nonoperative management have become increasingly standardized and efforts continue to further refine this strategy. The appropriate intensive care unit and acute care unit length of stay, the number of laboratory draws, the length of activity restriction and the need for radiographic evidence of healing prior to release from activity restriction remain areas of study. Previously demonstrated variation in the management and outcome of injured children between adult and pediatric surgeons has led to debate over which type of facility should best care for injured children. The Pennsylvania Trauma Systems Foundation dataset was used to derive a series of children with severe liver injuries. Finally, the risk of post-splenectomy sepsis, a stimulus for the initial development of nonoperative management, has been further clarified by a literature review. While falls from a low height may infrequently lead to a significant injury, falls from greater heights are more likely to induce a solid organ injury. PMID- 12130916 TI - Congenital diaphragmatic hernia. AB - Congenital diaphragmatic hernia occurs in approximately 1 in every 2500 live births and is associated with a reported mortality of almost 35% in live-born patients and a higher mortality when in utero deaths are counted. Ventilator induced lung injury, pulmonary hypoplasia, and other associated anomalies account for the high death rate. Numerous adjunctive measures have been used to treat these patients. Inhaled vasodilators (nitric oxide), intravenous vasodilators, and fetal therapy have no proven benefit. While animal models of congenital diaphragmatic hernia are surfactant deficient, controversy remains over the use of surfactant in infants. There has been no clinical trial showing any clear benefit with the use of exogenous surfactant in these patients. Similarly, prenatal corticosteroids show some improvements in animal models, but again, there is a complete absence of supportive data to show benefit in humans. Mechanical ventilator strategies that limit ventilator-induced lung injury by avoiding hyperventilation and lung over inflation are the strategies currently in use that have been associated with improved survival. Long-term follow-up of these patients is quite important since gastroesophageal reflux, developmental delay, chronic lung disease, and chest wall deformity are all seen with increased frequency in these children. PMID- 12130917 TI - Abdominal wall defects. AB - Survival for newborns with congenital abdominal wall defects (primarily omphalocele and gastroschisis) has improved, but controversy remains regarding etiology, anatomy and embryology, the role of prenatal diagnosis and mode of delivery, and initial management. A number of recent studies have added to our knowledge and understanding of several of these topics, while several others have raised questions regarding traditional initial management of these infants. Continued improvement in the survival of these infants can be anticipated with further understanding of the in utero and antepartum diagnosis and management of infants with these common congenital abnormalities. PMID- 12130918 TI - Neonatal jaundice, animal-induced injuries, and immunizations. PMID- 12130920 TI - Best supportive care versus palliative chemotherapy in nonsmall-cell lung cancer. AB - The incidence of lung cancer continues to rise. The need and demand for more effective treatment to improve survival and palliate symptoms increases at a great rate. The most recent evidence for the use of chemotherapy in the palliative setting is summarized in this review of the literature from the last few years. It indicates that in advanced nonsmall-cell lung cancer survival, symptom control and physical functioning can be improved with the use of chemotherapy not only in the first-line but also in the second-line setting, in the elderly, and at disease relapse. PMID- 12130919 TI - Cost-effectiveness of chemotherapy for nonsmall-cell lung cancer. AB - After decades of research into its prevention and treatment, lung cancer remains the leading cause of cancer death in North America and Europe. Approximately 75% of all new lung cancer diagnoses are of the nonsmall-cell subtype, and less than 25% of these patients are potentially operable upon first detection. First generation cisplatin-based chemotherapy regimens for patients with metastatic disease achieved a median survival of 175 days, with 15 to 20% of patients alive at 1 year.In recent years, vinorelbine, gemcitabine, paclitaxel, and docetaxel have emerged as promising agents in the treatment of advanced nonsmall-cell lung cancer. Evidence from randomized trials demonstrates that when these agents are combined with cisplatin, the objective tumor response is 25 to 40%, with a median overall survival approaching 300 days. In addition, recent studies have shown that single-agent docetaxel improves survival and quality of life in patients with platinum-refractory nonsmall-cell lung cancer. Since these modest but important improvements in the management of nonsmall-cell lung cancer are achieved at a significant cost, cost has emerged as a major consideration in health policy decision-making. This article reviews the pharmacoeconomic literature to provide guidance on the cost-effective use of chemotherapy in the treatment of advanced nonsmall-cell lung cancer. PMID- 12130921 TI - Benefits of chemotherapy for quality of life in patients with advanced nonsmall cell lung cancer. AB - It has now become clear that chemotherapy for advanced nonsmall-cell lung cancer, compared with best supportive care only, improves survival, even if the amplitude of the benefit remains disappointing. However, some clinicians are still reluctant to prescribe chemotherapy in this patient population, arguing that the survival gain is too small to counterbalance the side effects. Therefore, randomized trials using quality of life as an endpoint and comparing best supportive care with or without chemotherapy were reviewed. Although there are difficulties in the methodology of quality of life assessments and in the analysis of quality of life data, most of the selected trials showed an improvement for quality of life in various components in the chemotherapy arm. Therefore, even if some of the reported results might be biased and should be interpreted with caution, the data analyses constitute a further argument to stop denying chemotherapy for patients with advanced nonsmall-cell lung cancer, at least for those who might be eligible for inclusion in a clinical trial. PMID- 12130922 TI - Dyspnea: the continuing challenge of palliative management. AB - The management of dyspnea is a challenge even for the most experienced palliative medicine teams. In the absence of effective treatment for the underlying disease, therapeutic options are limited to the supplementation of oxygen, the use of opioids, and multidisciplinary nonpharmacologic interventions. There is increased research into both the physiology of dyspnea and the correlates of the symptom in advanced disease. Hopefully, this research will lead to improved therapy in the future. This article reviews current literature on dyspnea with a focus on publications in 2001. PMID- 12130924 TI - Retroperitoneal sarcomas: combined modality treatment approaches. AB - Retroperitoneal soft tissue sarcomas are rare tumors estimated to account for 15%of all patients with soft tissue sarcoma seen in referral populations. The standard of care for patients with localized, resectable retroperitoneal sarcomas is surgical resection with gross and microscopically negative margins. However, owing to the large size and locally advanced nature of these tumors, this goal is difficult to achieve in most patients. As a result, the disease is characterized by a high propensity for local recurrence and a grade-specific risk for distant metastasis. Over the past decade, there has been considerable research into combined modality treatment of these tumors. The present report outlines current concepts relating to the diagnosis, staging, and management of retroperitoneal sarcomas. Emphasis is placed on evolving combined modality treatment approaches and current investigational strategies. PMID- 12130926 TI - Chromosomal translocations and sarcomas. AB - This review examines how the identification of tumor-specific translocations and fusion proteins has advanced the basic scientific and clinical understanding of sarcomas. Recent genetic advances, including the ASPL-TFE3 fusion of alveolar soft part sarcoma, the JAZF1-JJAZ1 fusion of endometrial stromal sarcoma, and HMGIC fusions in liposarcoma, are discussed. Next, the review addresses the ways in which molecular genetic data have influenced diagnostic and prognostic paradigms. For example, recent studies describe the detection of occult tumor cells and the identification of primary renal neoplasms that are genetically related to alveolar soft part sarcoma. In addition, the review discusses potential therapies based on the targeting of sarcoma-specific fusion proteins. These reports describe the potential use of Gleevec (STI571) for dermatofibrosarcoma protuberans and the use of tumor-specific fusion proteins as potential targets for immunotherapy. Finally, basic scientific findings are reviewed that elucidate, for example, the aberrant functions of SYT-SSX in chromatin remodeling and of EWS-FLI1 in transcription and mRNA splicing. These and other emerging models of tumorigenesis will help identify new therapeutic targets. PMID- 12130925 TI - cDNA microarray analysis of global gene expression in sarcomas. AB - Sequencing of the human genome and rapidly evolving microarray technology have combined to provide investigators with the ability to analyze individual tumors and groups of tumors for global patterns of gene expression. Few of these types of studies have been performed on rhabdomyosarcomas and osteogenic sarcomas, including cell lines and animal models. Groups of expressed genes that may characterize rhabdomyosarcomas and their subgroups and separate them from other types of tumors have been identified. More specifically, genes involved in myogenesis or the inhibition of myogenesis have been identified, as have genes that may play a role in metastatic activity in osteogenic sarcomas. Also, a study documenting the consistent and specific gene expression profile of gastrointestinal stromal tumors has been published. While the data regarding gene expression patterns in sarcomas is accruing, numerous investigators are working on developing and enhancing bioinformatic skills and tools such that the vast amount of data can be converted into knowledge regarding biology, therapeutic responsiveness or resistance, and prognosis. PMID- 12130927 TI - Nonsurgical methods for liver metastases including cryotherapy, radiofrequency ablation, and infusional treatment: what's new in 2001? AB - Surgical resection is now well accepted as the standard treatment in 10 to 20% of patients with liver metastases. Tumor ablative techniques have been developed in recent years. The basic idea is to use them in patients with a limited number of intrahepatic deposits that are not totally resectable. Several papers published in 2001 have addressed cryotherapy. Cryotherapy can be considered an effective method for local destruction of liver metastases up to 3 to 4 cm in diameter but is also associated with a significant rate of complications. In many centers, cryoablation has now been replaced by radiofrequency ablation, the most widely used method for ablation of unresectable liver metastases. It can be performed during laparotomy, at laparoscopy, or percutaneously. Tumors less than 3 cm in their greatest diameter can be destroyed with one placement of the needle electrode. Metastases larger than 3 cm require several placements. Both cryotherapy and radiofrequency ablation are effective methods to induce necrosis of liver metastases. It is likely that in the near future, most patients with liver metastases will receive a multimodality treatment: a local treatment such as surgical resection or tumor ablation, and a general treatment such as hepatic infusional or systemic chemotherapy. Trials published in 2001 have shown that oral prodrugs of fluorouracil were probably equivalent to fluorouracil bolus administration. Regimens containing oxaliplatin or irinotecan have also been evaluated for efficacy and tolerance and by the intravenous route alone or in combination with hepatic artery infusion. Effective systemic chemotherapy regimens have resulted in increased survival rates and improved quality of life and in some cases have allowed resection of initially unresectable liver metastases. PMID- 12130928 TI - Update on pancreatic cancer. AB - Pancreatic cancer remains the fourth leading cause of cancer deaths in males and females in the beginning of this new millennium. The 5-year survival for all stages remains less than 5%. The frequent diagnosis at late stages of the disease limits the role of surgery as a curative modality in pancreatic cancer. Despite recent advances, systemic treatment continues to have a limited role in the adjuvant setting, and chemotherapy is mostly palliative in advanced and metastatic pancreatic cancer patients. The differential diagnosis of pancreatic cancer and other gastrointestinal malignancies is, many times, challenging. Advances in the understanding of the disease biology may help in better diagnosis and treatment approaches. Clinical trials with molecular targeting agents are starting to emerge. PMID- 12130930 TI - Infectious risks and outcomes after stem cell transplantation: are nonmyeloablative transplants changing the picture? AB - PURPOSE OF REVIEW: Opportunistic infections contribute to morbidity and mortality after myeloablative allogeneic stem cell transplantation. The development of nonmyeloablative or toxicity-reduced conditioning regimens for allogeneic hematopoietic stem cell transplantation might change this picture significantly. These regimens are in general highly immunosuppressive, but effects on myelopoiesis and mucosal toxicities are usually reduced compared with myeloablative hematopoietic stem cell transplantation conditioning regimens. This review summarizes the infectious risks associated with each type of hematopoietic stem cell transplantation conditioning regimen, and presents the results of early clinical studies. RECENT FINDINGS: Although the data are preliminary, the results of recent studies suggest that nonmyeloablative conditioning regimens may decrease the risks of bacterial infections associated with mucosal damage and persistent neutropenia; however, risks for late viral and fungal infections persist during severe graft versus host disease. Results of several case reports and series emphasize that therapeutic outcomes of infections may be improved in patients who receive nonmyeloablative conditioning regimens. SUMMARY: Infectious risks and outcomes after hematopoietic stem cell transplantation appear to be in evolution given the introduction of alternative, nonmyeloablative conditioning regimens. Although infections remain a prominent cause of transplant-related mortality, the timing and types of infections may differ. Further studies are necessary to define appropriate preventative strategies, and to determine whether patients with ongoing infections might benefit from nonmyeloablative hematopoietic stem cell transplantation. PMID- 12130931 TI - Viral infections in immunocompromised patients: what's new with respiratory viruses? AB - PURPOSE OF REVIEW: The leading cause of death in solid organ and hematopoietic stem cell transplant recipients is infection. The respiratory viruses, particularly respiratory syncytial virus, influenza, parainfluenza, adenovirus, and picornaviruses, are increasingly recognized as significant pathogens in these populations. RECENT FINDINGS: Respiratory syncytial virus has again been found to be the most common of the respiratory viruses causing severe infections in transplant recipients. Advances in prevention, particularly with regard to infection control practices, and to lesser extent treatment have had a substantial impact on the frequency and outcomes of this infection. New studies have clarified the impact of influenza in the hematopoietic stem cell transplant recipients and have provided evidence to support the use of M2 and neuraminidase inhibitors for early treatment. The epidemiology of parainfluenza and adenovirus in transplant recipients has been clarified, although therapeutic modalities are still limited and understudied. New antiviral medications may bring improved outcomes of picornavirus infections in this population. Finally, a new virus, the human metapneumovirus, has recently been described and may be a significant respiratory pathogen in immunocompromised transplant recipients. SUMMARY: Studies published over the past year have documented a new respiratory pathogen. They have also resulted in improved understanding of the epidemiology of all of the respiratory virus pathogens, and have contributed to improve management of respiratory syncytial virus and influenza infection in hematopoietic stem cell transplant and solid organ transplant recipients. PMID- 12130929 TI - Biologic approaches to the treatment of gastrointestinal malignancy. AB - Though new agents have recently been approved for the treatment of gastrointestinal malignancies, cure rates remain low and survival times of patients short. Because of these discouraging numbers, there is a vigorous search for new agents and new strategies. This article reviews some of these strategies and the status of agents in clinical development. Though an exhaustive search is impossible and the field is changing so rapidly, the agents selected here for discussion may be considered representative of others. Discussed here are agents targeting the vascular endothelial growth factor as an example of an angiogenesis inhibitor, and agents targeting the epidermal growth factor receptor, the ras oncogene, and the immune system. PMID- 12130933 TI - Treatment of febrile neutropenia: what is new? AB - PURPOSE OF THE REVIEW: To identify the more recent challenges in the treatment of patients with febrile neutropenia following antineoplastic chemotherapy or bone marrow transplant published in the English language in the period late 2000-early 2002 regarding: changes in etiology of bacteremia in neutropenic patients; new options for initial empirical antibacterial therapy; factors associated with the risk of developing infection in cancer patients; prediction of prognosis in febrile neutropenia; oral therapy; need for a specific anti-Gram-positive coverage in persistently febrile and neutropenic patients. RECENT FINDINGS: Findings may be summarized according with the identified topics as follows: many centers are reporting an increase in the incidence of Gram-negative bacteremias; piperacillin-tazobactam could be safely administered as monotherapy of febrile neutropenia; congenital factors and intensity of chemotherapy and other medical interventions, such as antifungal prophylaxis, are recognized as of increasing importance in the determination of infectious risk; it is now possible to identify patients with a good prognosis (low risk) by means of validated scoring systems; oral therapy is feasible in low risk patients; the empirical addition of a glycopeptide in persistently febrile neutropenic patients is not indicated. SUMMARY: Many of the identified points may have a great impact in the daily management of febrile granulocytopenic patients. However, all recent epidemiological and therapeutical studies underline the absoloute need for the knowledge of the pattern of infecting organisms in each center. PMID- 12130932 TI - Prophylaxis, empirical therapy, or pre-emptive therapy of fungal infections in immunocompromised patients: which is better for whom? AB - Immunocompromised patients are at risk of developing fungal infections. Over time, the incidence of fungal infections and the spectrum of causative organisms have changed. In addition, treatment strategies in this high-risk population have also changed. Traditional approaches (using polyene-based therapy and older azoles), including empirical treatment strategies, have evolved to include prophylaxis in populations at the greatest risk. These strategies, although effective against Candida species, have not really impacted infections caused by Aspergillus spp. With the recent approval of antifungal agents with demonstrated activity against Aspergillus and other mould infections, there is hope for better outcomes in the treatment of established infections. Several agents, with activity against Aspergillus, have been shown to be effective in the empirical setting. The role of these new antifungal agents in the prophylactic setting remains unknown at present, but the potential for reducing Aspergillus infections is promising and requires ongoing study. The other area of significant research in fungal infections has been the search for accurate, non-invasive, rapid diagnostic tests. Over the past year, several publications have indicated that early diagnosis is possible in immunocompromised patients. These new diagnostics have paved the way for a new strategy, called pre-emptive therapy, enabling infected patients to be identified at an earlier stage of infection. This strategy will permit targeted antifungal therapy in those at greatest risk, and will avoid unnecessary, potentially toxic therapy in those not infected. Validations of the various techniques show promise and are reviewed in this paper. PMID- 12130934 TI - Molecular typing: a key tool for the surveillance and control of nosocomial infection. PMID- 12130937 TI - Infection in the intensive care unit: prevention strategies. AB - PURPOSE OF THE REVIEW: Nosocomial infections remain among the most common treatment complications, particularly in intensive care unit patients. In many countries antibiotic resistance is increasingly hampering treatment of these infections. Preventive strategies have therefore become more important and have been directed both against the development of specific infections and against the spread of antibiotic-resistant pathogens. The present review addresses recent data on the latter issue. In particular, we discuss the first approaches to use mathematical modelling as a tool to analyse and guide strategies to prevent infection, and the effects of antibiotic cycling. RECENT FINDINGS: Several mathematical models to address the dynamics of pathogen transmission in hospital settings have been developed. One of the models may allow quantification of the effects of different strategies to prevent infection in intensive care units, and another may be used to determine the relative importance of different colonization routes, without the need for expensive genotyping methods. The results of the first prospective studies on antibiotic cycling are inconclusive, and again mathematical modelling may help to provide testable hypotheses for such interventions. Finally, recent studies have shown that alcohol-based hand rubs are better than hand washing with soap and water for most hand disinfection purposes. SUMMARY: The first results of use of mathematical modelling to guide infection control strategies should be subjected to prospective, empirical testing in order to determine their clinical usefulness. More rigorously designed studies are needed to determine the benefits of antibiotic cycling strategies. Hands should be disinfected with alcohol-based hand rubs, which should be available at each bedside. PMID- 12130935 TI - Nosocomial pneumonia. AB - PURPOSE OF REVIEW: Despite abundant literature on the management of nosocomial pneumonia, a number of aspects, from diagnosis to the therapy of nosocomial pneumonia, are still controversial. This review focuses on recent advances that can aid in the day-to-day care of these critically ill patients. RECENT FINDINGS: The risk factors for nosocomial pneumonia in specific subsets of trauma, postoperative and burn injury patients have been identified, with emphasis on the type of pneumonia developing in these populations - early or late onset nosocomial pneumonia. Resolution of nosocomial pneumonia, in terms of improvement of clinical parameters such as oxygenation, fever, leukocytosis and bacterial eradication, has been reported, and these data can lead to a better understanding of the natural course of the disease. The importance of initial, accurate empiric therapy in improving mortality in nosocomial pneumonia has been reinforced by multiple studies. Newer techniques to study colonization and the routes of spread of pathogenic organisms in the intensive care unit are adding to our understanding of how pneumonia develops, the role of infection control measures and the types of strategies that are needed for prevention. Oral decontamination is showing promise as a technique to prevent ventilator-associated pneumonia, and noninvasive ventilation has been shown to be useful in various etiologies of respiratory failure, with the beneficial effect of reducing the incidence of ventilator-associated pneumonia and its associated mortality. The implementation of protocolized treatment guidelines and antibiotic rotation policies are emerging as useful tools for reducing the frequency of antibiotic resistance and the impact of nosocomial pneumonia. SUMMARY: There is a better understanding of nosocomial pneumonia risk factors, mechanisms of bacterial colonization, and resolution of illness, with exciting developments in prevention and treatment emerging, and these data can help us achieve more effective management of this complex illness. PMID- 12130938 TI - New trends in Staphylococcus aureus infections: glycopeptide resistance in hospital and methicillin resistance in the community. AB - PURPOSE OF REVIEW: Methicillin-resistant Staphylococcus aureus is prevalent in hospitals throughout the world, and we have got used to its presence in daily clinical practice. However, methicillin-resistant S. aureus has not remained static over the past four decades, but seems to be evolving in unfamiliar directions. This review focuses on recent findings on two directions of methicillin-resistant S. aureus evolution: the acquisition of multiple antibiotic resistance in the hospital and the trend towards methicillin-resistant S. aureus as a community pathogen. RECENT FINDINGS: We looked at reports on glycopeptide resistance in S. aureus and those on community-acquired methicillin-resistant S. aureus strains, with some references of historical value to explain the entire picture of this 'new field' of the methicillin-resistant S. aureus problem. SUMMARY: The references given here (excluding some of low credibility) attest the increasing awareness of the two conspicuous problems concerning methicillin resistant S. aureus infection. One is the increasing trend of glycopeptide resistance, making difficult the successful treatment of multi-drug-resistant methicillin-resistant S. aureus infection in the hospital. On the other hand, non multi-drug-resistant methicillin-resistant S. aureus strains are emerging as novel threats in the community, the genetic analysis of which indicates that they are independent clones from those found in hospitals. PMID- 12130936 TI - Antibiotic policies and control of resistance. AB - PURPOSE OF THE REVIEW: The current worldwide pandemic of antibiotic resistance shows no signs of abating. It is clear that it is driven mainly by heavy and often inappropriate antibiotic use. Although control measures are widely practised, it is important that we assess their efficacy critically in order to concentrate expensive control efforts where they will be most effective. The past year has seen much activity in this area, with evidence-based assessments of the literature according to strict guidelines, as well as progress in basic science studies of mechanisms of resistance, and their causes and relations to pathogenicity and adaptability. RECENT FINDINGS: The present review summarizes current developments in the causes of antibiotic resistance, the classification of antibiotic stewardship and control measures, the evidence base for their efficacy, current problems in hospital practice, the adaptability of bacteria, the content of antibiotic policies and anticipated activities. SUMMARY: The conclusions from the published literature are that much of it that pertains to changing prescribing practices does not stand up to modern evidence-based analysis concepts. Nevertheless, we can learn from experience in changing other areas of medical practice. We must be pragmatic and must not expect to change the world, but rather take it step by step, recognizing barriers and measuring outcomes and quality indicators. Studies into the molecular basis of resistance confirm the superb genetic adaptability of micro-organisms. They will always be several steps ahead of us. Nevertheless, we are learning how to modify our prescribing habits to minimize resistance, not only by using antibiotics less frequently but also by altering dosing schedules in various ways. PMID- 12130939 TI - Surveillance in infection control: are we making progress? AB - PURPOSE OF THE REVIEW: Surveillance of nosocomial infections is an indispensable tool in infection control, and is used for detecting problem areas, defining patients who are at risk and evaluating intervention strategies. Surveillance techniques are continuously being evaluated and improved. RECENT FINDINGS: Problems with definitions, risk stratification and case finding render development of (inter)national surveillance systems difficult with respect to the comparability of data between different hospitals. These problems also influence surveillance in specific areas of nosocomial infections, such as urinary tract infections, ventilator-associated pneumonia and surgical site infections. Examples of such problem areas are discussed in the present review. SUMMARY: Despite continuing efforts made to improve quality of (inter)national and local surveillance systems, issues of infection definition and risk stratification are still under debate and need further research. PMID- 12130940 TI - Managing the risk of nosocomial transmission of prion diseases. AB - PURPOSE OF REVIEW: Prion diseases such as Creutzfeldt-Jakob disease represent a unique infection control problem because prions exhibit an unusual resistance to conventional chemical and physical decontamination methods. This paper reviews the recent literature and provides recommendations for the prevention of nosocomial transmission of prion agents. RECENT FINDINGS: Recommendations to prevent the cross-transmission of infection from medical devices potentially contaminated with prions have been based primarily on prion inactivation studies. Newer recommendations consider inactivation data, but also use epidemiological studies of prion transmission, the infectivity of human tissues, and the efficacy of removing microbes by cleaning. Prion-specific disinfection/sterilization is required in only limited settings. Healthcare workers are not at risk of acquiring transmissible spongiform encephalopathies. Blood or blood products have not been demonstrated to be vehicles for transmission. SUMMARY: On the basis of scientific data, only critical (e.g. surgical instruments) and semicritical devices contaminated with high-risk tissue (i.e. brain, spinal cord, eye) from high-risk patients (e.g. with known or suspected Creutzfeldt-Jakob disease) require special treatment. PMID- 12130941 TI - Prevention of postsurgical infections: some like it hot. AB - PURPOSE OF THE REVIEW: The present review covers the literature on prevention of surgical site infections published during 2001. Only papers that offer new insights or question current recommendations are included. RECENT FINDINGS: The most interesting advances have been made in the preoperative preparation of the patient. In particular, the effects of preoperative warming of the patient, either generally or locally at the incision site, on the rate of surgical site infections are impressive and warrant further investigation. Also, the effects of strict control of perioperative blood glucose levels and of preoperative eradication of nasal carriage of Staphylococcus aureus should be studied in greater detail. SUMMARY: The protective effect of local and systemic warming before surgical procedures has been described in two well designed studies. Although a final judgement regarding effectiveness can only be made after more extensive investigations have been conducted, there appear to be few arguments against application of this cheap and safe measure. Thus, implementation of preoperative warming can be justified in settings associated with high rates of surgical site infections. PMID- 12130942 TI - Drug allergy. PMID- 12130943 TI - Immunoglobulin E binding determinants on beta-lactam drugs. AB - PURPOSE OF REVIEW: Allergies to penicillins and cephalosporins remain an important clinical problem, but structural and immunochemical knowledge of the allergenic structures involved has tended to lag behind the heavy usage, consequent adverse reactions and introduction of new therapeutic members of these two families of antibiotics. Evidence of the increasing incidence of reactions to cephalosporins and to "minor" determinants of the beta-lactams is accumulating. Also, although numerous reports detail unique allergic recognitions of individual members of the two families, particularly the cephalosporins, information remains predominantly clinical. The present review summarizes the most recent advances in studies of structural aspects of beta-lactams as allergens. RECENT FINDINGS: For the cephalosporins, a pyrazinone allergenic degradation product of cefaclor and cephalexin has been identified and characterized. The widely used cephalosporin cephalothin was shown to cross-react allergenically with benzylpenicillin and the common cross-reacting structure was identified. The fine structural features on the amoxicillin molecule recognized by antibodies that distinguish "major" and "minor" determinants were identified, and steric factors were used to explain antibody recognition of the amoxicillin determinants. A recent study elucidated the molecular basis of some cases of the multiple drug allergy syndrome and its relationship to beta-lactam allergy. SUMMARY: Findings of the type described in the present review provide fundamental insights into the nature and size of antigenic determinants on "small" molecules such as drugs and other chemicals. At the clinical level, such structure/activity findings have implications for our understanding of drug allergenic cross-reactions, for selection for therapy of an appropriate member from a family of structurally related drugs and, ultimately, for desensitization of drug-allergic patients. PMID- 12130944 TI - Pharmacological interaction of drugs with antigen-specific immune receptors: the p-i concept. AB - PURPOSE OF REVIEW: Drug allergies are examples of immune reactions to small molecular compounds. In many drug allergies drug specific CD4+ and CD8+ T-cells can be detected, which recognize small chemicals via their alphabeta-T-cell receptor in a major histocompatibility complex dependent way. In this review a new concept of drug presentation to T-cells is presented. RECENT FINDINGS: Drugs were stimulatory for T-cells if they bound covalently to peptides or proteins, but also if the drug had structural features allowing it to bind in a labile way (noncovalently) to the major histocompatibility peptide complex. This latter binding method has some similarities to superantigen stimulations and can explain allergies to drugs that are not metabolized. It has been described in patients with maculopapular, bullous and neutrophilic drug eruption, as well as in contact dermatitis. SUMMARY: Noncovalent drug presentation leads to the stimulation of immune cells, namely T-cells. The drug needs two surface molecules (one inert serving as a scaffold, major histocompatibility complex, and one reactive, T-cell receptor) to exert its function. Although two receptor structures are involved, the process is reminiscent of a pharmacological interaction between a drug and its receptors and, from the phrase pharmacological interaction with immune receptors, was thus termed the p-i concept. PMID- 12130946 TI - Pathophysiology of fixed drug eruption: the role of skin-resident T cells. AB - PURPOSE OF REVIEW: Fixed drug eruption is a distinct variant of drug-induced dermatoses characterized by their relapse in the same location after the administration of the causative drug. We have recently shown that intraepidermal CD8+ T cells phenotypically resembling effector memory T cells are greatly enriched in the resting lesions of FDE. Although effector memory T cells have been implicated as the mediators of protection in epithelial tissues, our observation raises an alternative possibility that improper, enhanced or uncontrolled activation of intraepidermal T cells could contribute to severe tissue injury. Until recently, however, their detrimental effects on epithelial tissues have rarely been examined. The focus of this review is on how intra epidermal T cells originally evolved to protect tissue integrity can exert an opposite action that is deleterious to the host. RECENT FINDINGS: Because those T cells residing in the lesions, upon activation, can rapidly produce large amounts of IFN-gammaepsilon followed by localized epidermal injury, their activation is probably essential for the initiation of deleterious inflammatory responses in the lesions. The activity of these potent effector T cells is therefore carefully controlled to prevent unwanted tissue injury under physiological conditions. A complex interplay of stop and go signals to the skin-resident T cells provides a delicate balance between cell death and survival, thereby determining the degree and outcome of inflammation generated in response to pathogens or antigens. SUMMARY: This consideration may provide important insights into the way in which skin-resident T cells maintain immunological homeostasis in the skin. PMID- 12130945 TI - Mechanistic perspectives on sulfonamide-induced cutaneous drug reactions. AB - PURPOSE OF REVIEW: Idiosyncratic drug reactions continue to limit the therapeutic utility of sulfonamide drugs because of their associated morbidity and mortality. Cutaneous reactions are the predominant reasons for withdrawal of such drugs from use in patients. As a consequence of the recognized metabolic and immunologic capability of the skin, an understanding of the pathogenic role of this tissue in the development of sulfonamide-induced cutaneous drug reactions may provide insight into the mechanisms and risk factors for these and other adverse drug events. RECENT FINDINGS: In the present review we discuss currently available mechanistic information, including issues related to drug bioactivation and adduct formation, immunoresponsiveness, and immune dysregulation, for the development of sulfonamide-induced (delayed-type) cutaneous drug reactions. The potential application of findings from several related areas of research are also discussed within the context of the pathogenesis of these cutaneous reactions. SUMMARY: Despite progress, numerous unresolved issues support the testing of novel hypotheses, the search for additional risk factors, and the need for a global approach, including links between laboratory and clinical paradigms. These issues must be addressed if we are to gain an understanding of the mechanistic bases for these cutaneous drug reactions. PMID- 12130947 TI - Acute generalized exanthematous pustulosis, a clue to neutrophil-mediated inflammatory processes orchestrated by T cells. AB - PURPOSE OF REVIEW: Circumstantial evidence exists that certain neutrophilic inflammatory processes are regulated by T cells, but how this occurs is not well understood. The present review presents data on how T cells may directly orchestrate a neutrophilic inflammation by specific release of the neutrophil attracting chemokine CXCL8 (formerly known as interleukin-8). RECENT FINDINGS: Acute generalized exanthematous pustulosis (AGEP) is an uncommon cutaneous eruption that is most often provoked by drugs, by acute infections with enteroviruses, or by mercury. It is characterized by acute, extensive formation of nonfollicular sterile pustules on an erythematous background, fever and elevated numbers of blood neutrophils. Involvement of T cells in drug-induced AGEP was suggested by positive patch tests and lymphocyte transformation tests. Moreover, drug-specific CD4+ and CD8+ T cells could be isolated and propagated in vitro from patch test sites and blood from AGEP patients. Their main characteristic is a high level of CXCL8 production. SUMMARY: T cells are involved even in some neutrophil-rich inflammatory responses, and they may orchestrate the immune reaction directly by high CXCL8 production or indirectly via interleukin 17 production, which induces CXCL8 production in various cell types. AGEP serves as a valuable model for characterizing T cells with a particular function--namely production of CXCL8--leading to neutrophilic inflammation. It is tempting to speculate that elucidation of this pathomechanism will help to improve our understanding of similar neutrophilic eruptions (e.g. pustular psoriasis) and may reveal new targets for pharmacotherapeutic interventions in such diseases. PMID- 12130949 TI - Epidemiology of Hymenoptera allergy. AB - PURPOSE OF REVIEW: Epidemiology and especially the natural history of Hymenoptera allergy form the background that is essential to improving the clinical management of insect venom allergic patients. This review focuses on the emergence of recent data which could help provide further enlightenment in this field. RECENT FINDINGS: The latest data on the extent of the disease, the risk factors for sensitization and for local, systemic and fatal reactions after a hymenoptera sting are reviewed. The emerging problems concerning asymptomatic sensitization, the meaning of constitutively elevated tryptase serum levels and the persisting widespread poor awareness of available therapies in Hymenoptera allergic patients are particularly emphasized. SUMMARY: The assessment of the risk for systemic reaction in skin-positive subjects with a negative case history, and the suggestion of the baseline serum tryptase level as a risk factor for reaction severity after a sting, are the most important clinical implications of the latest studies. The genetic and environmental factors involved in the persistence of venom specific immunoglobulin E after a sting and the factors which orient towards a systemic or a large local reaction after apparently the same sting remain open questions. PMID- 12130948 TI - Late-onset allergy-like reactions to X-ray contrast media. AB - PURPOSE OF REVIEW: The purpose of the present review is to describe recent insight into the pathomechanism of the late-onset allergy-like reactions which affect 2-3% of contrast medium-exposed patients, and to give advice regarding prophylactic measures to avoid recurrence of such reactions. RECENT FINDINGS: It is well recognized that the majority of contrast media-induced late-onset reactions are cutaneous reactions of the maculopapular, urticarial and angioedema types, with occasional occurrence of more serious, bullous eruptions. Recent findings strongly indicate that these reactions are T cell-mediated. The current evidences for this concept are the described clinical symptoms; the high incidence of reactions in interleukin-2-treated patients; the positive results from skin, provocation and lymphocyte transformation testing of affected patients; and the histopathology of skin eruptions and positive skin test sites. SUMMARY: On the basis of this new knowledge, several prophylactic measures are proposed to avoid repeat reactions in patients with previous late-onset contrast medium reactions. All persons receiving contrast media should be informed that transient skin reactions may develop up to 7 days after contrast medium exposure. Those who experience such reactions should be advised to see an allergist for diagnosis of the reaction. Skin testing with a panel of different contrast media appears to be useful for confirming the presence of an allergic reaction and for identifying alternative contrast media that can be safely used. For undiagnosed persons, a contrast medium that is structurally different from the product that precipitated the reaction should be chosen if re-exposure to contrast media is required. Pretreatment with corticosteroids may also be tried, although its protective effect has not been systematically studied. PMID- 12130950 TI - Diagnosis of Hymenoptera venom sensitivity. AB - PURPOSE OF REVIEW: The objective of this review is to highlight recent advances in the preparation, documentation and performance of reagents and methods used in the diagnosis of Hymenoptera-venom-induced immediate-type hypersensitivity. RECENT FINDINGS: The following potent allergens have been reported: (1) a low molecular-weight honey-bee allergen (Api m 6) has been described; (2) venom allergens in the North American species of bumble-bee (Bombus pennsylvanicus) have been more fully characterized, with the focus on phospholipase A2; (3) the vespid venom Ves v 5 allergen has been structurally mapped to identify immunoglobulin-E-binding epitopes; (4) the possible role of carbohydrate antigen epitopes as a cause of cross-reactivity among honey-bee and vespid venom proteins has been reported; and (5) the venom of Pachycondyla chinensis, an ant found commonly in the Far East, has been described. The most significant reports during this period have focused on the less-than-ideal performance of the intradermal venom skin-test reagents. The issue of the patient that is positive for venom allergy history but negative for an intradermal venom skin test is raised, and it is suggested that there is a need for caution and the use of serology as a supplementary diagnostic test. SUMMARY: The important issue this year is the reminder that intradermal skin tests may be negative in venom-allergic patients, possibly because of changes in the potency of the extracts. The clinical history should drive the diagnosis of insect-sting allergy. When negative confirmatory venom skin-test or serology results are considered to be inconsistent with a positive history, they should be repeated. PMID- 12130951 TI - Patients with negative skin tests. AB - PURPOSE OF REVIEW: The aim of this article is to present and discuss the clinical problem of systemic anaphylaxis to Hymenoptera venoms in patients without detectable immunoglobulin E, as it appears in recent literature. Reported at variable frequency in large series of patients undergoing evaluation, systemic anaphylaxis was previously considered to reflect lost sensitization or to involve non-immunoglobulin E mediated mechanisms. Sporadic case-reports drew attention to the fact that severe or even fatal reactions may occur in patients with negative skin tests. RECENT FINDINGS: A breakthrough article by Golden et al., who performed deliberate stings on skin test negative venom anaphylaxis patients, demonstrated that clinical sensitivity was still present in a subset of these subjects and pointed out to the limitations of present diagnostic methods or reagents. New immunobiochemical methods and highly specific recombinant allergens -when all clinically relevant Hymenoptera venom allergens have been identified, cloned, sequenced and expressed in the proper system--are anticipated to increase the diagnostic yield. Non-specific mechanisms causing anaphylactoid reactions will probably explain some enigmatic, skin test negative radioallergosorbent test negative cases in the future. Occult mastocytosis, predisposing patients to anaphylactoid reactions, has been reported with increasing frequency among skin test negative patients. Lastly, other causes mimicking venom anaphylaxis may on rare occasions contribute to the problem. SUMMARY: With the present understanding of venom allergy, the practising clinician is not infrequently faced with the dilemma of the skin test negative patient. Once other identifiable causes have been carefully ruled out, referral to a specialized center for deliberate sting challenges appears in selected cases to be a medically appropriate and ethically justified management approach. PMID- 12130952 TI - Maintenance venom immunotherapy. AB - PURPOSE OF REVIEW: Venom immunotherapy has proven to be a very effective method for the prevention of future re-sting reactions. However, initiation of the venom injection program is just the beginning. This review looks at recent papers which shed light on other issues that arise during maintenance venom immunotherapy. RECENT FINDINGS: Prophylactic antihistamines taken before venom injections reduce the frequency of reactions and one report suggests that they may improve efficacy. The schedule of venom injections usually does not have to be adjusted for patients who develop local reactions; a very large reaction may be the exception. Patients who react to stings should have their maintenance doses increased. Most patients are able to extend the interval between injections to 8 weeks in the third year of treatment. Two groups have proposed a maintenance interval of 12 weeks for routine use. SUMMARY: Our understanding of insect sting sensitivity continues to improve, leading to better outcomes for allergic patients. PMID- 12130953 TI - Increasing respiratory rate to improve CO2 clearance during mechanical ventilation is not a panacea in acute respiratory failure. AB - BACKGROUND: Increasing respiratory rate has recently been proposed to improve CO2 clearance in patients with acute respiratory failure who are receiving mechanical ventilation. However, the efficacy of this strategy may be limited by deadspace ventilation, and it might induce adverse hemodynamic effects related to dynamic hyperinflation. SETTING: An intensive care unit of a university hospital. PATIENTS: We studied 14 patients with acute respiratory failure during the adjustment of ventilator settings on the first day of mechanical ventilation in volume-controlled mode. MEASUREMENTS: After determining the positive end expiratory pressure that suppresses any intrinsic positive end-expiratory pressure at a respiratory rate of 15 breaths/min, we compared blood gas analysis, respiratory measurements, and Doppler evaluation of right ventricular systolic function by using two different respiratory strategies with the same airway pressure limitation (plateau pressure, < or =25 cm H2O), a low-rate conventional respiratory strategy with a respiratory rate of 15 breaths/min, and a high-rate strategy with a respiratory rate of 30 breaths/min. RESULTS: Compared with the low-rate strategy, the high-rate strategy neither significantly reduced PaCO2 (47 +/- 8 vs. 51 +/- 7 mm Hg with the low-rate strategy) nor significantly improved PaO2 (99 +/- 40 vs. 95 +/- 35 mm Hg with the low-rate strategy). It significantly increased alveolar deadspace to tidal volume ratio (21% +/- 8%, vs. 14% +/- 6% with the low-rate strategy) and produced dynamic hyperinflation, resulting in a substantial intrinsic positive end-expiratory pressure (6.4 +/- 2.7 cm H2O). Right ventricular outflow impedance was increased, resulting in a significant drop in the cardiac index (2.9 +/- 0.6 vs. 3.3 +/- 0.7 L/min/m with the low-rate strategy). CONCLUSION: We conclude that a high respiratory rate strategy during mechanical ventilation in patients with acute respiratory failure did not improve CO2 clearance, produced dynamic hyperinflation, and impaired right ventricular ejection. PMID- 12130955 TI - Cryptococcal encephalitis in thermally injured mice is accelerated by type 2 T cell responses. AB - OBJECTIVE: To explore the pathogenic role of burn-associated type 2 T-cell responses on the development of cryptococcal encephalitis in mice with severe thermal injuries. DESIGN: Experimental Cryptococcus neoformans infection in normal mice was compared with that in thermally injured mice (TI mice), normal mice treated with a mixture of interleukin (IL)-4 and IL-10, or normal mice inoculated with burn-associated type 2 T cells. SETTING: University research laboratory. SUBJECTS: Male BALB/c mice, 8 to 10 wks of age. INTERVENTIONS: We prepared four groups of mice as follows: a) normal mice, b) TI mice, c) normal mice treated with the IL-4/IL-10 mixture, and d) normal mice inoculated with burn associated type 2 T cells. These groups of mice were anesthetized and exposed to 1 x 10 cells/mouse of C. neoformans intratracheally. Cryptococcal growth in brains and lungs in normal mice were compared with those of the other three groups. Also, cytokine-producing profiles of T lymphocytes from brains of both normal mice and TI mice were determined. MEASUREMENTS AND MAIN RESULTS: Compared with normal mice, TI mice were susceptible to C. neoformans infection. At the maximum (15 days after infection), numbers of C. neoformans organisms in brains of TI mice were 10 times higher than those of the pathogen in brains of normal mice. After stimulation with anti-CD3 monoclonal antibody, IL-4 (but not interferon gamma) was produced in cultures of T lymphocytes from brains of TI mice 15 days after the infection, whereas the same cell preparation from normal mice produced interferon gamma (but not IL-4). TI mice and mice that were treated with a IL-4/IL-10 mixture or inoculated with burn-associated type 2 T cells were equally susceptible to the cryptococcal infection. CONCLUSIONS: Burn-associated type 2 T cells or their cytokine products play a key role in the severity of cryptococcal encephalitis that develops in TI mice. PMID- 12130954 TI - Family satisfaction with care in the intensive care unit: results of a multiple center study. AB - OBJECTIVE: To determine the level of satisfaction of family members with the care that they and their critically ill relative received. DESIGN: Prospective cohort study. SETTING: Six university-affiliated intensive care units across Canada. METHODS: We administered a validated questionnaire to family members who made at least one visit to intensive care unit patients who received mechanical ventilation for >48 hrs. We obtained self-rated levels of satisfaction with 25 key aspects of care related to the overall intensive care unit experience, communication, and decision making. For family members of survivors, the questionnaire was administered while the patient was still in the hospital. For family members of nonsurvivors, the questionnaire was mailed out to the family member 3-4 wks after the patient's death. MAIN RESULTS: A total of 891 family members received questionnaires; 624 were returned (70% response rate). The majority of respondents were satisfied with overall care and with overall decision making (mean +/- sd item score, 84.3 +/- 15.7 and 75.9 +/- 26.4, respectively). Families reported the greatest satisfaction with nursing skill and competence (92.4 +/- 14.0), the compassion and respect given to the patient (91.8 +/- 15.4), and pain management (89.1 +/- 16.7). They were least satisfied with the waiting room atmosphere (65.0 +/- 30.6) and frequency of physician communication (70.7 +/- 29.0). The variables significantly associated with overall satisfaction in a regression analysis were completeness of information received, respect and compassion shown to the patient and family member, and the amount of health care received. Satisfaction varied significantly across sites. CONCLUSIONS: Most family members were highly satisfied with the care provided to them and their critically ill relative in the intensive care unit. Efforts to improve the nature of interactions and communication with families are likely to lead to improvements in satisfaction. PMID- 12130956 TI - Electrocardiogram in Pneumocystis carinii pneumonia: can it be used as a prognostic variable? AB - OBJECTIVE: Many prognostic variables have been studied in patients with Pneumocystis carinii pneumonia and acquired immunodeficiency syndrome (AIDS). The role of the electrocardiogram in this setting has not been previously evaluated. We analyzed the admission electrocardiogram in patients with Pneumocystis carinii pneumonia and AIDS in an attempt to identify electrocardiogram findings that could be associated with adverse clinical outcomes and worse prognostic variables. DESIGN: A retrospective medical chart review. SETTING: All confirmed cases of Pneumocystis carinii pneumonia in patients positive for human immunodeficiency virus admitted to Albert Einstein Medical Center from 1994 to 2000. METHODS: Patients were assigned increasing severity ranks based on the findings on the admission electrocardiogram (normal sinus rhythm, sinus tachycardia, and right ventricular strain pattern). Data were extracted regarding study outcomes (admission to intensive care unit, mechanical ventilation, and hospital mortality) and prognostic variables. MAIN RESULTS: Of the 40 study patients, 14 (35%) had normal sinus rhythm, 15 (37.5%) had sinus tachycardia, and 11 (27.5%) presented with signs of right ventricular strain. The number of admissions to the intensive care unit, use of mechanical ventilation, and hospital mortality rate all increased with the severity of the electrocardiogram findings (p < or =.03). The serum lactate dehydrogenase concentrations and the alveolar-arterial oxygen gradient both increased with the severity of the electrocardiogram findings (p < or =.02). CONCLUSION: Electrocardiogram findings of sinus tachycardia and right heart strain are common in Pneumocystis carinii pneumonia. These findings are associated with adverse clinical outcomes as well as worsening of prognostic variables. The electrocardiogram may be useful in predicting outcome in patients with Pneumocystis carinii pneumonia. PMID- 12130957 TI - Gastrointestinal promotility drugs in the critical care setting: a systematic review of the evidence. AB - CONTEXT: Gastrointestinal promotility agents may improve tolerance to enteral nutrition, reduce gastroesophageal reflux and pulmonary aspiration, and therefore have the potential to improve outcomes of critically ill patients. OBJECTIVE: To systematically review and critically appraise studies of promotility agents in the critical care setting. DATA SOURCES: Computerized bibliographic search of published research (1980-2001), citation review of relevant articles, and contact with primary investigators. STUDY SELECTION: Randomized trials of critically ill adult patients that evaluated the effect of promotility agents on measures of gastrointestinal motility were included. DATA EXTRACTION: Relevant methods and outcome data were abstracted in duplicate by independent investigators. DATA SYNTHESIS: We reviewed 60 citations; 18 articles met the inclusion criteria (six studies of feeding tube placement, 11 studies evaluating gastrointestinal function, and one study of clinical outcomes). The heterogeneity of study methods and outcomes measured precluded a quantitative synthesis of the data. Although there are conflicting studies, the larger and more methodologically robust studies suggest that metoclopramide has no effect on feeding tube placement. Erythromycin has been shown to increase success rates with small-bowel tube placement in two studies. Eight of ten studies evaluating the effect of cisapride, metoclopramide, or erythromycin on measures of gastrointestinal transit demonstrated positive effects; the two studies that did not were relatively small (n = 27 and 10) and likely had inadequate power to detect a difference in treatment effect. No study demonstrated a positive effect on clinical outcomes. CONCLUSIONS: As a class of drugs, promotility agents appear to have a beneficial effect on gastrointestinal motility in critically ill patients. A one-time dose of erythromycin may facilitate small-bowel feeding tube insertion. Administration of metoclopramide appears to increase physiologic indexes of gastrointestinal transit and feeding tolerance. Concerns about safety and lack of effect on clinically important outcomes preclude strong treatment recommendations. PMID- 12130958 TI - Gastric versus small-bowel tube feeding in the intensive care unit: a prospective comparison of efficacy. AB - OBJECTIVE: To compare the outcomes of intensive care unit patients fed through a nasogastric vs. a nasal-small-bowel tube including the time from tube placement to feeding, time to reach goal rate, and adverse events. DESIGN: Sixty patients were prospectively randomized to receive gastric or small-bowel tube feedings. Nursing staff attempted to place a feeding tube in the desired position, and placement was confirmed radiographically after each bedside attempt. After two unsuccessful attempts, the feeding tube was placed under fluoroscopy. Feedings were started at 30 mL/hr and advanced to the patient's specific goal rate. SETTING: Twenty-bed medical intensive care unit. PATIENTS: Sixty medical patients admitted/transferred to the intensive care unit. INTERVENTIONS: Tube feeds were held for 2 hrs if any residual was >200 mL. MEASUREMENTS: Times were recorded at the initial tube insertion, onset of feeding, achievement of goal rate, and termination of feeding. Adverse outcomes included witnessed aspiration, vomiting, and clinical/radiographic evidence of aspiration. Patients were followed up for the duration of feeding, until leaving the intensive care unit, or for a maximum of 14 days. MAIN RESULTS: Patients fed in the stomach received nutrition sooner from initial placement attempt (11.2 hrs vs. 27.0 hrs) and with fewer attempts (one vs. two) than those fed in the small bowel. Patients achieve goal rate sooner (28.8 hrs vs. 43.0 hrs) with gastric feeding compared with small-bowel feeding. There was no difference in aspiration events. CONCLUSIONS: Gastric feeding demonstrates no increase in aspiration or other adverse outcomes compared with small-bowel feeding in the intensive care unit. Gastric feeding can be started and advanced to goal sooner with fewer placement attempts than small bowel feeding. PMID- 12130960 TI - Prone-position ventilation induces sustained improvement in oxygenation in patients with acute respiratory distress syndrome who have a large shunt. AB - OBJECTIVES: Prone-position ventilation (PPV) induces acute improvement in oxygenation in many patients with acute respiratory distress syndrome (ARDS), with some maintaining their oxygenation even after they were returned to the supine position, but it is unclear what clinical factors determine the sustained oxygenation benefit. We hypothesized that patients with ARDS who have a larger shunt would have a better acute and sustained oxygenation response to PPV. DESIGN: Prospective, nonrandomized interventional study. SETTING: Medical and surgical intensive care units, university tertiary care center. PATIENTS: Twenty two consecutive patients, with ARDS with an average PaO2/FiO2 of 94, were administered PPV for 12 hrs followed by supine-position ventilation for 2 hrs. MEASUREMENTS: Hemodynamic and gas exchange variables were monitored. The shunt was measured as venous admixture at an FiO2 of 1.0, and compliances of the respiratory system, lung, and chest wall were measured by the esophageal balloon technique before PPV, during PPV, and during subsequent supine-position ventilation. MAIN RESULTS: Fourteen patients (64%) responded to PPV, with PaO2/FiO2 increasing by > or =20. These changes were associated with a decrease in chest wall compliance. Responders had significantly shorter time from ARDS to PPV, a lower baseline PaO2/FiO2, and a higher venous admixture. All responders maintained the improvement in oxygenation and had a greater respiratory system compliance after returning to the supine position. Time from ARDS to PPV and baseline lung injury score were negatively associated, whereas chest wall compliance, heart rate, and PaCO2 were positively associated with sustained improvement in oxygenation. CONCLUSIONS: PPV induced acute and sustained improvement in oxygenation in many patients with ARDS. The sustained improvement is more significant if PPV is administered early to patients with a larger shunt and a more compliant chest wall. Measuring venous admixture and chest wall compliance before PPV may help select a subgroup of patients with ARDS who may benefit the most from PPV. PMID- 12130962 TI - Internal and external validation of the NOSEP prediction score for nosocomial sepsis in neonates. AB - OBJECTIVE: To evaluate the performance of a scoring system (NOSEP) to predict nosocomial sepsis in neonates at the hospital where the score was developed (internal validation) and in an independent data set from other centers (external validation). DESIGN: Multiple center prospective cohort study. SETTING: Six neonatal intensive care units from the Flanders in Belgium. PATIENTS: We analyzed two groups of patients: 62 episodes of presumed nosocomial sepsis in the internal validation cohort and 93 episodes of presumed nosocomial sepsis in a multiple center external validation cohort. INTERVENTIONS: Assessment of the predictive power of the NOSEP score 24 hrs preceding sepsis workup and the patients' basic demographic characteristics and co-morbidity was performed. Diagnosis of nosocomial sepsis and the microbiology results were registered. MAIN RESULTS: The NOSEP score's discriminative capability was very good in the internal validation (area under receiver operating characteristic curve = 0.73 +/- 0.08 [sem]). The NOSEP score performed satisfactory in the external validation (area under receiver operating characteristic curve = 0.66 +/- 0.06). The calibration capability in both validation sets as measured by goodness-of-fit tests (internal validation, p =.56; external validation, p =.48) was good. An improvement of the NOSEP score was obtained for the external centers by redefining the cut-off of the items of the NOSEP score (area under receiver operating characteristic curve for NOSEP-NEW-I = 0.71 +/- 0.05) or adding co-morbidity factors (area under receiver operating characteristic curve for NOSEP-NEW-II = 0.82 +/- 0.04), with good calibration performance (goodness-of-fit test, p >.50). Finally, the fit of the NOSEP score demonstrated no significant variation across subgroups of patients. CONCLUSIONS: The predictive power of the original NOSEP score is very good in neonates at the original neonatal intensive care unit. In other neonatal intensive care units, its discriminatory performance is satisfactory but could be improved after modification of the variables in the model or adding additional variables. To use such a NOSEP score in other neonatal intensive care units, its accuracy has to be validated and adjusted if necessary. PMID- 12130961 TI - Noninvasive ventilation in acute exacerbations of chronic obstructive pulmonary disease in patients with and without home noninvasive ventilation. AB - OBJECTIVE: The frequency of home ventilation has increased greatly. The objective of the study was, first, to compare the outcome of episodes of acute exacerbation of chronic obstructive pulmonary disease treated with mask intermittent positive pressure ventilation (MIPPV) in patients with home MIPPV and in patients without home ventilatory support and, second, for each category of patients, to compare patients successfully ventilated with MIPPV with those who failed with MIPPV. DESIGN: Prospective, controlled, nonrandomized clinical study. SETTING: Medical intensive care unit of a university hospital. PATIENTS: In the groups with and without home MIPPV, respectively, 31 and 78 episodes of acute exacerbations of chronic obstructive pulmonary disease were studied. INTERVENTIONS: MIPPV was performed in a sequential mode and delivered through a full-face mask with a bilevel positive airway pressure system. MEASUREMENTS AND MAIN RESULTS: The clinical and functional characteristics of the two groups, at admission, were similar. In groups with and without home ventilation, respectively, success rates were 68% and 72% (p =.68), length of intensive care unit stay was 8 +/- 6 and 10 +/- 4 days (p =.02), and intensive care unit deaths were 13% and 8% (p =.30). In survivors and in groups with and without home ventilation, respectively, the total time of ventilatory assistance in intensive care unit was 5 +/- 4 and 8 +/- 4 days (p =.004), and the length of intensive care unit stay was 7 +/- 5 and 10 +/- 4 days (p =.003). A greater correction of pH, after 45 mins of MIPPV with optimal settings, was recorded in the success patients than in the failure patients, respectively; in the group with home MIPPV, the pH after 45 mins was 7.34 +/- 0.04 vs. 7.31 +/- 0.04 (p =.06), and in the group without home MIPPV, pH was 7.34 +/- 0.04 vs. 7.30 +/- 0.04 (p =.001). CONCLUSION: MIPPV may also be favorable during episodes of acute exacerbations in patients with chronic obstructive pulmonary disease. Experience with MIPPV could benefit selected patients in the management of acute respiratory failure. PMID- 12130963 TI - Automatic tube compensation in patients after cardiac surgery: effects on oxygen consumption and breathing pattern. AB - OBJECTIVE: To evaluate patients without prior pulmonary disease after cardiac surgery and to determine whether resistive unloading by automatic tube compensation, pressure support ventilation, and continuous positive airway pressure has different effects on oxygen consumption, breathing pattern, gas exchange, and hemodynamics. DESIGN: Prospective, randomized, controlled study. SETTING: Tertiary care, postoperative intensive care unit. PATIENTS: Twenty-one patients scheduled for open heart coronary artery bypass graft surgery. INTERVENTIONS: Each patient was ventilated with all three modes in random order. MEASUREMENTS AND MAIN RESULTS: Patients were ventilated in three modes, each applied for 30 mins according to computer-generated randomization: pressure support ventilation with 5 cm H2O, continuous positive airway pressure, and automatic tube compensation. Oxygen consumption was calculated by means of indirect calorimetry. The hypnotic state of the patients was monitored by Bispectral Index. For hemodynamic measurements, a fiberoptic pulmonary artery catheter was inserted. The main finding of our study was that oxygen consumption and breathing pattern (tidal volume and respiratory rate) did not differ significantly during automatic tube compensation and pressure support ventilation compared with continuous positive airway pressure (oxygen consumption, 170 +/- 29 vs. 170 +/- 26 vs. 174 +/- 29 mL.min.m, respectively; tidal volume, 466 +/- 132 vs. 484 +/- 125 vs. 470 +/- 119 mL, respectively; respiratory rate, 16 +/- 4 vs. 15 +/- 4 vs. 16 +/- 4 breaths/min, respectively). Automatic tube compensation and pressure support ventilation had no clinical effects on gas exchange and hemodynamic variables compared with continuous positive airway pressure. None of the variables differed significantly during the three ventilatory settings. CONCLUSION: In postoperative tracheally intubated patients with normal ventilatory demand, automatic tube compensation and pressure support ventilation with 5 cm H2O lead to identical oxygen consumption, breathing patterns, gas exchange, and hemodynamics. We, therefore, suggest that this group of patients does not need any additional positive pressure support from the ventilator to overcome the additional work of breathing imposed by the endotracheal tube during the weaning phase from mechanical ventilation. PMID- 12130964 TI - Toward understanding evidence uptake: semirecumbency for pneumonia prevention. AB - OBJECTIVE: Randomized trials show that the semirecumbent position compared with the supine position is associated with less gastroesophageal aspiration and pneumonia in patients receiving mechanical ventilation. However, semirecumbency is inconsistently used in practice. The objective of this study was to understand the perspectives of intensive care unit clinicians regarding the determinants and consequences of semirecumbency. DESIGN: Qualitative study using semistructured interviews and focus groups. SETTING: Three university-affiliated intensive care units. PARTICIPANTS: A total of 93 intensive care unit clinicians, including bedside nurses, respiratory therapists, physiotherapists, nutritionists, residents, fellows, and intensivists. METHODS: We elicited perceptions about benefits and harms of semirecumbency, factors promoting and deterring use, and health systems changes to encourage semirecumbency. Interview and focus group notes were analyzed inductively to identify emerging themes. Validation methods involved triangulation by multidisciplinary analysis of several data sources collected through multiple methods and member checking. MEASUREMENTS AND MAIN RESULTS: Intensivists and nutritionists were familiar with semirecumbency as a potential pneumonia prevention strategy, whereas other clinicians were not. When made aware of the evidence, all participants endorsed semirecumbency. Nurses perceived that the main determinant of semirecumbency was physicians' orders, whereas intensivists perceived that the main determinant was nursing preference. Participants identified barriers to semirecumbency related to useful alternative positions (e.g., lateral position), contraindications (e.g., hemodynamic instability), risk of harm (e.g., decubitus ulcers), safety (e.g., sliding out of the bed), and resources (e.g., insufficient beds facilitating semirecumbency). Education, guidelines, reminders, audit and feedback, charting, and quality improvement initiatives were advocated to promote semirecumbency. CONCLUSIONS: Under-utilization of semirecumbency for pneumonia prevention is influenced by insufficient awareness of its benefit, real and perceived deterrents, poor agreement about implementation responsibility, and lack of enabling and reinforcing strategies. Cognitive, behavioral, and administrative approaches to enhancing evidence uptake may be needed in the complex, dynamic intensive care unit setting. PMID- 12130965 TI - Plasma and tissue pharmacokinetics of cefpirome in patients with sepsis. AB - OBJECTIVE: Broad initial antibiotic treatment is crucial for patients experiencing septic shock or severe sepsis. Fourth-generation beta-lactam antibiotics, such as cefpirome, are frequently favored in these conditions because of their low toxicity and wide antimicrobial coverage. From recent data, however, there is circumstantial evidence that one reason for the high mortality rate of patients with sepsis might be an impaired penetration of antimicrobial agents from the central compartment to the infectious focus. Thus, the present study aimed at describing penetration properties of cefpirome to the target site of many bacterial infections, which is the extracellular space fluid of soft tissues. DESIGN: Prospective comparative study of two groups. SETTING: An intensive care unit and research ward in a university hospital. SUBJECTS: The study population included 12 patients with septic shock or severe sepsis and a control group of six overall age-matched healthy volunteers. INTERVENTIONS: To measure cefpirome penetration into the interstitial space fluid of skeletal muscle, we employed microdialysis after single intravenous administration of 2.0 g of cefpirome to patients and healthy volunteers. MEASUREMENTS AND MAIN RESULTS: Maximum concentration and area under the concentration vs. time values in interstitium were significantly lower in patients compared with the control group (p <.004). Cefpirome area under the concentration time values for plasma were 16.0 +/- 1.1 mg.min/mL (mean +/- sem) and 18.8 +/- 1.1 mg.min/mL in patients and healthy volunteers, respectively (p =.075, not significant). In both study groups, mean cefpirome concentrations in interstitium and plasma exceeded 28 microg/mL throughout the observation period of 240 mins and covered completely minimal inhibitory concentration values for a range of clinically relevant pathogens. CONCLUSION: Cefpirome concentrations reached in tissue interstitium and plasma exceeded minimal inhibitory concentrations of most clinically relevant pathogens in patients with sepsis. Thus, cefpirome exhibits a tissue pharmacokinetic profile, which seems to be particularly valuable for the empirical therapy of patients with sepsis. PMID- 12130966 TI - How reliable is the Bispectral Index in critically ill patients? A prospective, comparative, single-blinded observer study. AB - OBJECTIVE: To establish a correlation between a reliable subjective measure, the Sedation-Agitation Scale (SAS), and an objective tool, the Bispectral Index (BIS), for monitoring critically ill patients with a decreased level of consciousness. DESIGN: Prospective, comparative, single-blinded observer study. SETTING: Surgical and medical intensive care units of the Tufts-New England Medical Center, a 349-bed tertiary care, academic medical center. PATIENTS: A convenience sample of 20 adult, critically ill patients with a decreased level of consciousness. The data from one patient were excluded because the patient did not meet inclusion criteria. MEASUREMENTS AND MAIN RESULTS: Patients were prospectively evaluated by a blinded observer using the SAS to subjectively determine their level of consciousness. Sedation levels varied from unarousable (SAS score of 1), to very sedated (SAS score of 2), to mildly sedated (SAS score of 3). Simultaneously, the patients were continuously monitored for 4-6 hrs with the BIS device. There was wide variability in BIS scores for any given level of consciousness as compared with the SAS. Unarousable patients had BIS scores ranging from 23 to 97, with a median score of 50 and an interquartile range of 24. Very sedated patients had BIS scores ranging from 35 to 98, with a median score of 68 and an interquartile range of 36. Mildly sedated patients had BIS scores ranging from 67 to 91, with a median score of 76 and an interquartile range of 8. Overall, there was a less than satisfactory correlation between BIS values and SAS scores (r =.36, p <.001). However, the correlation improved with subgroup analysis when BIS values associated with excessive muscle movement were excluded (r =.50, p <.001). CONCLUSIONS: The correlation between SAS and BIS scores was suboptimal and inconsistent in a heterogeneous group of critically ill patients. The generation of BIS hardware and software, studied herein, is neither reliable nor valid for routinely monitoring the level of consciousness in the critically ill patient. Excessive muscle movement by the patient is an important and spurious influence on BIS values and seriously undermines BIS reliability. PMID- 12130967 TI - Immunoparalysis in patients with severe trauma and the effect of inhaled interferon-gamma. AB - OBJECTIVE: To evaluate the local immune status in patients with severe trauma and the influence of interferon-gamma on patients with immunoparalysis. PATIENTS: Fifty-two mechanically ventilated patients with severe multiple trauma. SETTING: A 14-bed polyvalent intensive care unit. INTERVENTIONS: The local immune status was evaluated by examining bronchoalveolar lavage fluid. Subsequently, the patients were divided into two groups: immunoparalyzed (group 1) and nonimmunoparalyzed (group 2). Immunoparalysis was defined as a decreased level of human leukocyte antigen-DR expression of alveolar macrophages in <30%. Patients with immunoparalysis were treated with 100 microg of inhaled recombinant human interferon-gamma, three times daily (group 1a, 11 patients) or placebo (group 1b, ten patients). A second bronchoalveolar lavage fluid was obtained 3 days after the initiation of therapy. MEASUREMENTS: The alterations in human leukocyte antigen-DR expression, as well as in pro- and anti-inflammatory markers, such as platelet-aggregating factor, phospholipase A2, interleukin-1beta, platelet aggregating factor acetylhydrolase, and interleukin-10, were evaluated in the bronchoalveolar lavage fluids. RESULTS: In 21 of 52 (40%) patients, immunoparalysis was established. After interferon-gamma administration, the level of human leukocyte antigen-DR expression increased in group 1a from 17 +/- 5% to 46 +/- 9%. In parallel, platelet-aggregating factor and interleukin-1beta as well as the specific activities of phospholipase A2 and platelet-aggregating factor acetylhydrolase significantly increased. In contrast, interleukin-10 decreased after interferon-gamma therapy. In group 1b, no statistically significant changes appeared in the levels of human leukocyte antigen-DR expression or in the concentrations of inflammatory mediators. The incidence of ventilator-associated pneumonia was significantly lower in group 1a than in group 1b. The administration of interferon-gamma did not affect the outcome of the patients. CONCLUSIONS: A significant proportion of multiply injured patients developed immunoparalysis. The administration of interferon-gamma resulted in the recovery of levels of human leukocyte antigen-DR expression in alveolar macrophages, influenced the inflammatory reaction, and decreased the incidence ventilator associated pneumonia, without affecting the patients' outcome. PMID- 12130968 TI - Incidence, risk factors, and prognosis of a moderate increase in plasma creatinine early after cardiac surgery. AB - OBJECTIVE: To evaluate the incidence and prognosis of a moderate increase in serum creatinine early after cardiac surgery. DESIGN: Retrospective clinical study. SETTING: Surgical intensive care unit in a university hospital. PATIENTS: Five hundred and ninety-one consecutive adult patients operated on for cardiac surgery during 1 year. INTERVENTIONS: Plasma creatinine was measured systematically before and during the first 3 days after surgery. Comorbid events were assessed as organ dysfunction (cardiac, pulmonary, hematologic, and neurologic), allowing us to calculate for each patient a dysfunction score (0-5). MEASUREMENTS AND MAIN RESULTS: Postoperative plasma creatinine increased by > or =20% in 15.6% of patients; eight of these required dialysis. A 20% increase in plasma creatinine was associated with other organ dysfunction in 79.3% of patients. Overall mortality rate was 2.7% and increased with the dysfunction score (17.7% for a dysfunction score > or =3). Mortality rate was 12.0% for patients who had 20% increased plasma creatinine with other organ dysfunction but was 0% for patients without other organ dysfunction. A logistic regression analysis revealed that the most important prognostic factors of death were cardiac dysfunction (odds ratio, 8.5; 95% confidence interval, 2.2-32.5) and the association of renal dysfunction and hematologic dysfunction (odds ratio = 12.0; 95% confidence interval, 3.9-37.2). Mean intensive care unit stay of patients with increased plasma creatinine was significantly longer (8.1 +/- 5.6 vs. 4.3 +/ 1.4 days, p <.01) and increased significantly with the dysfunction score (p <.01). Patients with isolated increased plasma creatinine had a significantly longer stay in the intensive care unit than patients without any organ dysfunction (4.6 +/- 1.4 vs. 3.9 +/- 0.9, p <.01). CONCLUSION: Our results suggest that a postoperative 20% increase in plasma creatinine after cardiac surgery is not rare and has a significant impact on postoperative outcome, mainly when multiple organ dysfunction occurs. Any preoperative reduced renal reserve or perioperative renal ischemia increases the renal risk. PMID- 12130969 TI - Moderate hypothermia delays proinflammatory cytokine production of human peripheral blood mononuclear cells. AB - OBJECTIVE: To clarify the influence of moderate hypothermia on the production of proinflammatory cytokines. DESIGN: Controlled in vitro study. SETTING: Research laboratory. SUBJECTS: Peripheral blood mononuclear cells from healthy adult human subjects. INTERVENTIONS: Stimulation with 1 microg/mL lipopolysaccharide at 33 degrees C and 37 degrees C. MEASUREMENTS: Concentrations of released tumor necrosis factor-alpha, interleukin-1beta, and interleukin-6 were measured chronologically by enzyme immunoassay. The number of mRNA copies of these cytokines was determined by competitive reverse transcriptase-polymerase chain reaction analysis, and nuclear factor-kappaB activations were assessed by electrophoretic mobility shift assay. MAIN RESULTS: Significant reduction of the released-tumor necrosis factor-alpha concentration was observed 1 and 2 hrs after the stimulation with lipopolysaccharide at 33 degrees C compared with 37 degrees C. The peak release of interleukin-1beta at 33 degrees C was delayed 12 hrs later than that at 37 degrees C. A delayed peak in the release of interleukin-6 also was observed at 33 degrees C. The peaks of cytokines were confirmed at the mRNA expression level by competitive reverse transcriptase-polymerase chain reaction analysis at both temperatures. The peak of the tumor necrosis factor-alpha mRNA expression level was observed at 1 hr after the stimulation at 37 degrees C and 2 hrs after the stimulation at 33 degrees C. In the interleukin-1beta mRNA expression, at 37 degrees C the first peak appeared 1 hr and the second 6 hrs after the stimulation. In contrast, at 33 degrees C, the first peak appeared 2 hrs and the second 12 hrs after the stimulation. Whereas interleukin-6 mRNA expression at 37 degrees C peaked 6 hrs after the stimulation, no definite peak was observed at 33 degrees C and the expression level was approximately half of that at 37 degrees C. The maximum intensity of nuclear factor-kappaB activation at 33 degrees C was delayed by 1.5 hrs compared with that at 37 degrees C. CONCLUSIONS: Moderate hypothermia delays the induction of proinflammatory cytokines in human peripheral blood mononuclear cells. PMID- 12130970 TI - When small degrees of bias in randomized trials can mislead clinical decisions: an example of individualizing preventive treatment of upper gastrointestinal bleeding. AB - OBJECTIVE: Although randomized trials yield less biased estimates of treatment effects than other study designs, unconcealed randomization and lack of blinding can lead to overestimates of the treatment benefit on the order of 15% to 40% on average. In applying the results of clinical trials to patient care, clinicians need to be concerned about bias of this order when it would change patient management. The aim of this study is to assess under which circumstances clinicians need to be concerned about bias in clinical trials. DESIGN: Sensitivity analysis. SETTING: Recently published meta-analysis of RCTs on the benefit of H2-blockers on the prevention of gastrointestinal bleeding in critically ill patients. INTERVENTIONS: Assessment of the effect of different degrees of bias on clinical decision making in various clinical scenarios. RESULTS: Bias of even a modest degree (15% to 40% overestimation of effect) changed clinical decisions more frequently when treatment benefits were small and when the patients' risk to suffer an adverse outcome was low. When the benefit was large and the patients were at high risk, clinical decisions remained unchanged, despite bias in the estimation of effect. CONCLUSIONS: When treatments of moderate benefit are applied to patients of low to moderate risk, even small biases in the estimation of effects carry a high risk of erroneous decisions. When the treatment benefit is large and the patients are at high risk, clinicians need to be less concerned about bias in RCTs. PMID- 12130971 TI - Is dopamine administration possibly a risk factor for delirium? AB - OBJECTIVE: We explored the possibility that the administration of intravenous dopamine increases the risk for delirium as manifested by need for haloperidol. DESIGN: This study was based on a retrospective analysis. To examine the contribution of dopamine in the prediction of need for haloperidol, a multivariate logistic regression model was used. SETTING: University hospital. PATIENTS: All inpatient admissions to Stanford University Hospital over a 1-year period (n = 21,844). INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: Dopamine administration was associated with nearly a tripling of the odds of subsequent need of the antipsychotic drug (chi-square = 108, df = 1, p =.0001, odds ratio = 2.89), even after intensive care unit admission and diagnostic related group weight were considered as indicators of severity of illness. Even when analysis was limited to patients seen in the intensive care unit setting (n = 3,308), dopamine administration remained a very strong risk factor for haloperidol and hence possibly for delirium. The increased risk of need for haloperidol in patients administered dopamine is evident in every age group after age 20. CONCLUSIONS: The retrospective nature of this study, the inexact method to assess acuity, and, most of all, the use of haloperidol as an indicator of the presence of delirium preclude concluding that dopamine is directly a risk factor for delirium, much less a causal risk factor. However, the association is potent enough to suggest this possibility strongly and thus supports the need for prospective studies to examine the relationship between dopamine and delirium and to consider possible prophylactic treatment against delirium in those given dopamine. PMID- 12130972 TI - Tin chloride pretreatment prevents renal injury in rats with ischemic acute renal failure. AB - OBJECTIVE: To investigate whether tin chloride pretreatment ameliorates renal injury in rats with ischemic acute renal failure (IARF) by virtue of its kidney specific heme oxygenase-1 induction. DESIGN: Randomized, masked, controlled animal study. SETTING: University-based animal research facility. SUBJECTS: Sprague-Dawley male rats, weighing 200-230 g (n = 359). INTERVENTIONS: Rats were injected with tin chloride subcutaneously, because subcutaneous administration of tin chloride is known to specifically and potently induce renal heme oxygenase activity in the rat. Anesthetized rats were subjected to bilateral flank incisions, and the right kidney was removed. Renal ischemia for 40 mins was performed by left renal microvascular clamping, followed by reflow of the blood. MEASUREMENTS AND MAIN RESULTS: Tin chloride treatment specifically induced heme oxygenase-1 mRNA and protein in the proximal tubular epithelial cells of the kidney without apparent cell injury in the rat. Tin chloride treatment before renal ischemia augmented the induction of heme oxygenase-1 in IARF rats at both transcriptional and protein concentrations in the renal epithelial cells compared with IARF animals. Tin chloride pretreatment, which decreased microsomal heme concentration, ameliorated the ischemic renal injury as judged by the significant decrease in serum creatinine and blood urea nitrogen concentrations and the lesser tubular epithelial cell injuries. In contrast, inhibition of heme oxygenase activity by treatment with tin mesoporphyrin, which increased microsomal heme concentration, abolished the beneficial effect of tin chloride pretreatment. CONCLUSION: These findings indicate that tin chloride pretreatment significantly ameliorates renal injury in rats with IARF by virtue of its specific heme oxygenase-1 induction in renal epithelial cells. These findings also suggest that heme oxygenase-1 induction plays an important role in protecting renal cells from oxidative damage caused by heme. PMID- 12130973 TI - Specific compliance and gas exchange during high-frequency oscillatory ventilation. AB - OBJECTIVES: To evaluate the use of specific compliance (static compliance/functional residual capacity) to adjust mean airway pressure, resulting in optimal gas exchange during high-frequency oscillatory ventilation in a surfactant-deficient newborn piglet. DESIGN: Prospective controlled animal study. SETTING: Laboratory. SUBJECTS: Eight newborn piglets at 5 days of age. BACKGROUND: High-frequency oscillatory ventilation enables the use of relatively high mean airway pressures without the lung damage associated with conventional positive pressure ventilation. Mean airway pressures can be increased, resulting in static lung expansion that approaches total lung capacity with its negative impact on venous return. Therefore, knowledge of lung volume is important for safe patient management. A simple, noninvasive technique to enable the clinician to determine the optimal mean airway pressure likely would improve patient management. INTERVENTIONS: The lungs were lavaged after placement of central catheters and tracheostomy to lower respiratory system compliance and worsen ventilation perfusion matching. The animals were ventilated with high-frequency oscillatory ventilation at the same mean airway pressure as before lung lavage. Mean airway pressures then were increased in a step-wise fashion up to 30 cm H2O or until clinical deterioration occurred. All other ventilator variables, Fio2, frequency, and pressure amplitude were constant throughout the experiment. MEASUREMENTS AND MAIN RESULTS: Before lavage and at each level of mean airway pressure after lung lavage, respiratory system compliance and functional residual capacity were measured. Additionally, central arterial pressure, central venous pressure, heart rate, arterial blood gas, and pulse oximetric saturation were recorded. Lung lavage significantly lowered respiratory system compliance (static as well as specific compliance) and worsened ventilation perfusion matching as evidenced by an increase in Paco2 and a decreased arterial to alveolar oxygen ratio. With increasing mean airway pressures, static/specific compliance improved and then peaked before declining, functional residual capacity increased, and blood gas improved until reaching the flat portion of the pressure-volume relationship of the lung. Optimal gas exchange as reflected by the highest arterial to alveolar oxygen ratio and lowest Paco2 at constant ventilation was found at a mean airway pressure that maintained the functional residual capacity and static respiratory system compliance at the same level as the preinjury levels ("normalized" functional residual capacity and respiratory system compliance). CONCLUSIONS: These results suggest that specific compliance measurement that incorporates static respiratory system compliance and functional residual capacity during high-frequency oscillatory ventilation can be used to adjust mean airway pressure and achieve "normalized" functional residual capacity, static compliance, and gas exchange. These measurements may provide a simple method to optimize lung volume in a surfactant-deficient patient during high-frequency oscillatory ventilation. PMID- 12130974 TI - Fas/Fas ligand pathway is involved in the resolution of type II pneumocyte hyperplasia after acute lung injury: evidence from a rat model. AB - OBJECTIVE: We used a rat model of acute lung injury to evaluate the role of apoptosis of type II pneumocytes in alveolar remodeling during the resolution phase. DESIGN: Controlled animal study. SETTING: University research laboratory. SUBJECTS: Sprague-Dawley rats. INTERVENTIONS: Sprague-Dawley rats had Escherichia coli lipopolysaccharide instilled transtracheally to induce acute lung injury. Animals were killed on various days after lipopolysaccharide instillation. Lung specimens from all animals were examined for the presence of apoptosis in type II pneumocytes by an in situ apoptosis assay and for proliferative nuclear antigen, cytokeratin-18, Fas, and Fas ligand with an immunohistochemical stain. Fas and Fas ligand expression in both lung tissue and bronchoalveolar lavage fluid was examined by Western blot analysis. MEASUREMENTS AND MAIN RESULTS: Histologic examination revealed that the lungs of rats with acute lung injury showed infiltration of numerous inflammatory cells in the intra-alveolar and/or interstitial space and hyperplasia of type II pneumocytes. Type II pneumocyte proliferation, detected by proliferative nuclear antigen staining, developed maximally around day 3 after acute lung injury. In the in situ apoptosis assay, positive signals in type II pneumocytes were obvious and were distributed diffusely in the lung parenchyma from day 1 after acute lung injury, became maximal around day 7, then declined until day 21. DNA fragmentation analysis revealed that a DNA ladder pattern was detectable from day 3, persisted until day 10, and disappeared after day 14. The major cell types expressing Fas ligand are macrophages and neutrophils. Western blot analysis showed that Fas ligand, both membrane-bound form and soluble form, was present from day 1 to day 21 after acute lung injury, with highest level occurring during the first week of acute lung injury. Fas expression in type II pneumocytes reached its maximum on days 3 5 and then gradually declined until day 21. Fas and Fas ligand expression appeared to proceed type II pneumocyte apoptosis. After the acute stage, Fas and Fas ligand expression declined, and type II pneumocyte apoptosis also decreased. These findings correlate with histologic resolution of type II pneumocyte hyperplasia. CONCLUSIONS: Our results confirm that type II pneumocyte proliferation in response to acute lung injury is mainly a reparative phenomenon. During the resolution phase of acute lung injury, extensive apoptosis of type II pneumocytes is the main cellular mechanism that accounts for the disappearance of these cells, and Fas/Fas ligand is involved in the resolution of type II pneumocytes. Our model may provide a useful tool to assess the mechanisms of tissue remodeling after acute lung injury. PMID- 12130975 TI - Mild hypothermia alters the oxygen consumption/delivery relationship by decreasing the slope of the supply-dependent line. AB - OBJECTIVE: To evaluate the effect of mild hypothermia on the relationship between systemic oxygen consumption and oxygen delivery. DESIGN: Prospective animal study. SETTING: University research laboratory. SUBJECTS: Anesthetized and ventilated rabbits. INTERVENTIONS: Rabbits were subjected to stepwise cardiac tamponade to reduce oxygen delivery while body temperature was maintained at 34 degrees C (group H, n = 8) or 39 degrees C (group N, n = 8). MEASUREMENTS AND MAIN RESULTS: The oxygen consumption/oxygen delivery relationship was analyzed by the dual-line method. The slope of the supply-dependent line was significantly decreased in group H (y = 0.57x + 1.3) compared with that in group N (y = 0.72x + 1.7), indicating that the ability of tissues to extract oxygen was impaired during hypothermia. Consequently, the proportion of the supply-independent area over the entire range of oxygen delivery was decreased in response to hypothermia. CONCLUSION: The potential for tissue hypoxia is likely to be increased during hypothermia when the circulation becomes unstable and oxygen delivery decreases. PMID- 12130976 TI - Natural history of long-term lung injury in mouse experimental pancreatitis. AB - OBJECTIVE: In patients suffering from acute pancreatitis, the pathogenesis of pancreatitis-associated lung injury is not completely understood. Several rodent models of pancreatitis-associated lung injury suggested that activated neutrophils and the release of proinflammatory mediators after the activation of inflammatory cells within the pancreas might play an important role in translating the pancreatic inflammation to the lungs. In this study, we examined the natural history of pancreatitis-associated lung injury during an entire week. SUBJECTS: Mice were administered 12 hourly intraperitoneal injections of a supramaximal dose of cerulein. MEASUREMENTS AND MAIN RESULTS: The severity of pancreatitis was time-dependent, with a maximal injury by 12-24 hrs after the start of cerulein administration. Pancreatitis was associated with a significant lung injury characterized by a rise in lung microvascular permeability, sequestration of neutrophils within the lungs, and a marked thickening of alveolar membranes. Within the lungs, the peak of macrophage inflammatory peptide 2, which attracts inflammatory cells within the injured area, preceded the peaks of both tumor necrosis factor-alpha and intercellular adhesion molecule-1. Moreover, histologic injury peaked by 12 hrs, with a full recovery at day 7. Serum macrophage inflammatory peptide-2 concentrations were significantly correlated with the occurrence of pulmonary leakage. Lung macrophage inflammatory peptide-2 concentrations peaked 12 hrs before pancreatic concentrations. CONCLUSIONS: Mediators released by the pancreas into the blood during acute pancreatitis induce within the lungs the chronological expression of macrophage inflammatory peptide-2, tumor necrosis factor-alpha, and intercellular adhesion molecule-1. PMID- 12130977 TI - Intravenous L-lactate application in minipigs partially protects acetylcholinesteratic but not butyrylcholinesteratic activity in plasma from inhibition by paraoxon. AB - OBJECTIVE: Intoxications with organophosphorous compounds such as paraoxon, an inhibitor of serine hydrolases, mainly butyrylcholinesterase and acetylcholinesterase, are frequent. Oximes are the only enzyme reactivators clinically available. In vitro studies have shown that L(+)-lactate reduces the inhibition of acetylcholinesteratic (AChEA) and butyrylcholinesteratic activity of plasma (BChEA) by paraoxon. DESIGN: The purpose of this in vivo study was to determine whether intravenous L(+)-lactate application under normoxic/normocapnic/normohydrogenemic conditions is able to protect AChEA and BChEA from paraoxon inhibition. SETTING: University research institute. SUBJECTS: Eighteen female minipigs. INTERVENTIONS: Animals were anesthetized, intubated, and mechanically ventilated. Every animal received 1 mg of paraoxon per kilogram of body weight in 50 mL of saline over 50 mins. In addition to receiving paraoxon, six pigs of 18 received 2.5 g (0.125 g kg-1 of body weight) of intravenous L(+)-lactate in 50 mL of saline over 50 mins, and six other pigs received 10 g of L(+)-lactate (0.5 g kg-1 of body weight), whereas the six remaining served as controls. MEASUREMENTS AND MAIN RESULTS: In central venous blood, plasma acetylcholinesteratic and butyrylcholinesteratic activity were measured before paraoxon (baseline, 0 mins), immediately after paraoxon (50 mins after start), and 110, 170, 230, 290, 530, and 1010 mins after the start of infusion. Although 10 g of intravenous L(+)-lactate application had a statistically significant protective effect in vivo on AChEA, 2.5 g did not. No significant protective effect on BChEA was achieved with either 2.5 g or 10 g of L(+)-lactate. CONCLUSIONS: Ten grams of L(+)-lactate can increase AChEA when administered simultaneously with paraoxon. Further study of the in vivo effects of L(+)-lactate after paraoxon intoxication and a formal comparison with standard oxime therapy seem warranted. Also, methods for achieving a prolonged elevated lactate concentration in vivo should be investigated. PMID- 12130978 TI - Organ damage in zymosan-induced multiple organ dysfunction syndrome in mice is not mediated by inducible nitric oxide synthase. AB - OBJECTIVE: To examine the role of inducible nitric oxide synthase (iNOS) in the development of the multiple organ dysfunction syndrome (MODS) in a murine model by using either a selective iNOS inhibitor or iNOS knockout mice. DESIGN: Prospective randomized laboratory study. SETTING: Central animal laboratory and experimental laboratory. SUBJECTS: Fifty inbred C57BL/6 mice, 39 iNOS knockout ( /-) mice, and 30 wild-type (+/+) mice, 7-9 wks old, weighing 20-25 g. INTERVENTIONS: Mice received an aseptic intraperitoneal injection of 40 microg of lipopolysaccharide followed by zymosan at a dose of 1 mg/g of body weight 6 days later (day 0). In experiment 1, C57BL/6 mice additionally received intraperitoneal injections with 5 mg of aminoguanidine or saline every 12 hrs, from 4 days after the injection of zymosan onward. In experiment 2, both iNOS-/- mice and corresponding wild-type (iNOS+/+) mice were treated with lipopolysaccharide and zymosan. MEASUREMENTS AND MAIN RESULTS: In all animals, the injection of zymosan induced an acute peritonitis, followed by an apparent recovery. From approximately day 6 onward, animals entered the third-MODS-like phase, indicated by weight loss, a decrease in body temperature, and significant mortality rates. Quantitative reverse transcriptase polymerase chain reaction and immunochemistry revealed a strongly increased expression of iNOS messenger RNA and iNOS protein in livers of mice in the last phase. However, neither the in vivo administration of aminoguanidine to C57BL/6 mice nor the complete absence of iNOS enzyme (iNOS-/- mice) had a beneficial effect on survival rate, body temperature, or body weight. In addition, relative lung, liver, and spleen weights and lung scores were not different between experimental groups. CONCLUSIONS: The current results strongly argue against an essential and causative role of iNOS in the development of organ damage in our murine model of MODS. PMID- 12130979 TI - Protein synthesis inhibiting clindamycin improves outcome in a mouse model of Staphylococcus aureus sepsis compared with the cell wall active ceftriaxone. AB - OBJECTIVE: The release of proinflammatory components from bacteria depends on the mode of action of the antibacterial therapy used. We studied whether this influences mortality in experimental sepsis. DESIGN: In a lethal murine model of Staphylococcus aureus sepsis, animals were randomly assigned to receive the protein synthesis inhibitor clindamycin (CLI) or the beta-lactam ceftriaxone (CRO). SETTING: Therapy was introduced subcutaneously 5 hrs after intraperitoneal injection of 10 colony forming units of S. aureus American Type Culture Collection 29213 and was continued every 8 hrs for 3 days. MEASUREMENTS AND RESULTS: Survival was higher in mice receiving CLI (29/50 animals [58%]) than in mice receiving CRO (16/50 animals [32%]; p =.015). Mice treated with CRO died earlier than mice receiving CLI (p =.002). Eight hours after the first antibiotic dose, the motor performance of mice receiving CRO had deteriorated more than it did for mice receiving CLI (p =.009). Higher levels of tumor necrosis factor alpha were measured in serum (p =.027) and peritoneal fluid (p =.001) of CRO treated mice. In vitro, CLI released smaller amounts of staphylococcal enterotoxin A than CRO. CONCLUSIONS: Antibiotic treatment of Gram-positive sepsis with a protein synthesis inhibitor decreases morbidity and mortality compared with a bacteriolytic compound. This may be caused by a reduction of the concentrations of proinflammatory/toxic bacterial components and cytokines. PMID- 12130980 TI - Abnormal pulmonary vascular tone in canine oleic acid lung injury. AB - OBJECTIVE: To characterize the endothelium-dependent and endothelium-independent components of abnormal pulmonary vascular tone in canine oleic acid lung injury. DESIGN: Prospective, interventional study. SETTING: University laboratory. SUBJECTS: Twenty anesthetized mongrel dogs. INTERVENTIONS: Right heart catheterization was performed to measure pulmonary vascular resistance before and after induction of oleic acid lung injury in ten anesthetized and ventilated dogs. Pulmonary and internal mammary artery rings were sampled in these ten dogs with oleic acid injury and in ten anesthetized healthy control dogs. We also studied the responses to acetylcholine, to phenylephrine, and to hypoxia of the intact or endothelium-denuded rings mounted in organ baths. MEASUREMENTS AND MAIN RESULTS: Oleic acid lung injury was associated with an increase in pulmonary vascular resistance from 118 +/- 11 to 245 +/- 47 dyne.sec.cm-5.m-2 and a decrease in the Pao2/Fio2 ratio from 451 +/- 42 to 139 +/- 26 mm Hg (mean +/- se, p <.05 and p <.01, respectively). Acetylcholine-induced relaxation was decreased in the oleic acid pulmonary artery rings compared with the controls (85 +/- 3% vs. 99 +/- 6% of precontraction level, p <.05). Phenylephrine-induced contraction was decreased in denuded oleic acid pulmonary artery rings compared with the controls (81 +/- 8% vs. 102 +/- 10% of contraction to KCl 120 mM, p <.05). In vitro hypoxia induced a small endothelium-dependent contraction followed by an endothelium-independent relaxation. These responses were not different in oleic acid lung artery rings and in controls, except for a decrease in hypoxic contraction in the oleic acid pulmonary artery rings. In vitro hypoxic pulmonary vasoconstriction and relaxation were, respectively, directly (r =.48) and inversely (r = -.67) correlated with oleic acid-induced increase in pulmonary vascular resistance. There was no correlation between in vitro internal mammary artery reactivity and oleic acid-induced increase in pulmonary vascular resistance. CONCLUSIONS: Oleic acid-induced lung injury slightly impairs pulmonary arterial endothelium-dependent relaxation and endothelium-independent contraction. In vitro hypoxic pulmonary vasoreactivity is related to in vivo oleic acid-induced increase in pulmonary vascular resistance. PMID- 12130981 TI - Atrial natriuretic peptide improves pulmonary gas exchange by reducing extravascular lung water in canine model with oleic acid-induced pulmonary edema. AB - OBJECTIVE: The purpose of this study was to examine and compare the effects of atrial natriuretic peptide and furosemide on pulmonary gas exchange, hemodynamics, extravascular lung water, and renal function in a dog model of oleic acid-induced pulmonary edema. DESIGN: Prospective, comparable, experimental study. SETTING: Laboratory at a university hospital. SUBJECTS: Eighteen male beagle dogs were studied under mechanical ventilation with pentobarbital anesthesia. INTERVENTIONS: Oleic acid (0.08 mL/kg) was injected and allowed for 1 hr to achieve pulmonary edema with hypoxemia at Fio2 of 0.3. After lung injury, dogs were divided into three groups; control group (n = 6) receiving saline (2.5 mL/hr for 5 hrs), atrial natriuretic peptide group (n = 6) receiving atrial natriuretic peptide (1 microg x kg(-1) x min(-1) for 5 hrs), and furosemide group (n = 6) receiving furosemide (1 mg x kg(-1) x hr(-1) for 5 hrs). MEASUREMENTS AND MAIN RESULTS: Hemodynamics, arterial blood gases, extravascular lung water, and renal function were measured hourly for 7 hrs after injury. Oleic acid increased extravascular lung water and induced hypoxemia. In the atrial natriuretic peptide group, extravascular lung water was significantly (p <.05) lower and Pao2 was significantly (p <.05) higher than in the control and furosemide groups, respectively. Pulmonary hypertension induced by oleic acid was attenuated by atrial natriuretic peptide infusion but not by saline or furosemide. Increased natriuresis/diuresis did not significantly differ between the atrial natriuretic peptide and the furosemide group, whereas creatinine clearance in the atrial natriuretic peptide group was significantly higher than that in the furosemide group. CONCLUSIONS: These findings suggest that atrial natriuretic peptide improves pulmonary gas exchange by reducing extravascular lung water and pulmonary arterial pressure, possibly independently from natriuresis/diuresis in oleic acid-induced pulmonary edema. PMID- 12130983 TI - Volume resuscitation does not alleviate peripheral organ ischemia in dogs injected with scorpion venom. AB - OBJECTIVES: To examine the effect of fluid resuscitation on the hemodynamic changes in dogs injected with scorpion venom and to explore the effects of the venom on the determinants of venous return (i.e., circulatory compliance, time constant, and resistance to venous return). DESIGN: A prospective, controlled animal study. SETTING: University animal research laboratory. SUBJECTS: Mixed breed dogs. INTERVENTIONS: The effect of volume resuscitation (20 mL/kg of the synthetic colloid polygeline) 1 hr after venom injection (a time previously found to be related to severe decrease in cardiac output) was tested in two series of experiments. In the first series, 12 dogs were given venom and fluid, eight dogs were given venom alone, and four dogs served as the time-controlled group. In the second series, eight dogs were given venom and ten dogs served as controls. Scorpion venom (Leiurus quinquestriatus) at 0.1 mg/kg in the first series and 0.05 mg/kg in the second series was given intravenously. MEASUREMENTS AND MAIN RESULTS: In the first series of experiments, the venom decreased cardiac output from 5.0 +/- 1.1 to 2.9 +/- 0.7 L/min at 60 mins (p <.001). Arterial pH decreased from 7.39 +/- 0.05 to 7.16 +/- 0.1 (p <.001). Blood lactate increased from 0.9 +/ 0.8 to 3.2 +/- 1.9 mM (p <.05). Gastric pH decreased from 7.28 +/- 0.2 to 6.7 +/ 0.18 (p <.001). Arterial acidosis was secondary to gastrointestinal ischemia because the gradient between mucosal and arterial Pco2 increased from 17.5 +/- 7.7 to 98.6 +/- 75 (p <.01) 60 mins after venom injection. In the second series of experiments, circulatory compliance and time constant increased by 150% and 128%, respectively (p <.05), in dogs injected with venom compared with control dogs. Resistance to venous return increased after venom injection but did not change after fluid infusion. In both series of experiments, volume administration improved cardiac output but had no effect on oxygen delivery, arterial pH, HCO3-, lactate, and gastric mucosal pH. CONCLUSIONS: Metabolic acidosis and cardiovascular abnormalities seen after scorpion venom injection in dogs are closely related to gastrointestinal hypoperfusion. Fluid resuscitation increased cardiac output but had no effect on gastrointestinal perfusion and acidosis induced by the venom. PMID- 12130982 TI - Epidermal growth factor reduces ischemia-reperfusion injury in rat small intestine. AB - OBJECTIVE: To measure the effect of pre-ischemic administration of intraluminal epidermal growth factor on the changes in intestinal permeability induced by 30 mins of superior mesenteric artery occlusion, followed by 2 hrs of reperfusion. DESIGN: Prospective, randomized, placebo-controlled experimental study. SETTING: University basic science research laboratory. SUBJECTS: Healthy, young, adult, male Sprague-Dawley rats. INTERVENTIONS: A 10-cm segment of small intestine was isolated and studied in situ in rats that were anesthetized with fentanyl and mechanically ventilated. Intestinal ischemia-reperfusion injury was induced by temporary occlusion of the superior mesenteric artery for 30 mins, followed by 2 hrs of reperfusion. Three groups were studied: time controls with a sham operation, saline-treated ischemia-reperfusion, and epidermal growth factor treated ischemia-reperfusion. Epidermal growth factor, 100 ng/min, was infused intraluminally, beginning 30 mins before and continued until 40 mins after ischemia. MEASUREMENTS AND MAIN RESULTS: Intestinal permeability was measured for each 10-min time period by using chromium-labeled EDTA. Histopathologic injury was assessed by light microscopy. After superior mesenteric artery occlusion, intestinal permeability increased approximately ten-fold and was sustained for 2 hrs of reperfusion in saline-treated rats. Pretreatment with epidermal growth factor significantly reduced the permeability changes during reperfusion by >60% compared with saline-treated animals (p <.05). Histopathologic sections revealed apparently more extensive loss of epithelial cells and mucosal disruption in saline-treated intestine compared with epidermal growth factor-treated intestine. CONCLUSION: Pre-ischemic administration of intraluminal epidermal growth factor significantly protects against intestinal ischemia-reperfusion injury. PMID- 12130984 TI - Intratidal compliance-volume curve as an alternative basis to adjust positive end expiratory pressure: a study in isolated perfused rabbit lungs. AB - OBJECTIVE: Repeated collapse and reopening of alveoli have been shown to aggravate lung injury, which could be prevented by positive end-expiratory pressure (PEEP). Yet, how to adjust optimum PEEP is a matter of debate. We suggest a new strategy to adjust PEEP, which is based on the analysis of the intratidal compliance-volume curve. This approach was compared with a strategy based on the static pressure-volume curve. Furthermore, two other ventilator settings were investigated. One served as a negative control likely to provoke atelectasis, and the other was expected to induce overdistension. DESIGN: Prospective, randomized block design. SETTING: Laboratory. SUBJECTS: Isolated, perfused, and ventilated rabbit lungs. INTERVENTIONS: Tidal volumes of 8 mL/kg of body weight were used throughout. After stabilization, the lungs were randomized to one of four protocols (lasting 120 mins; n = 6 per group). Group 1 was ventilated at zero end-expiratory pressure. In group 2, PEEP was set above the lower inflection point of the static pressure-volume curve. In group 3, adjustment of PEEP was based on the intratidal compliance-volume curve, as determined by the slice method. In group 4, increasing PEEP levels ensured a plateau airway pressure of 20-25 cm H2O likely to provoke overdistension. MEASUREMENTS AND MAIN RESULTS: The ventilation/perfusion (VA/Q) distribution was analyzed by the multiple inert gas elimination technique. Alveolar derecruitment was indicated by shunt and low VA/Q areas as observed in group 1. In groups 2 and 3, VA/Q data initially indicated full recruitment. In contrast to group 3, shunt increased in group 2 near completion of the experiments. Group 4 showed complete recruitment, but the VA/Q distribution included high VA/Q areas. CONCLUSIONS: The intratidal compliance-volume curve represents a rational basis for adjusting PEEP in the isolated lung model. Because this strategy does not require invasive measures and facilitates continuous assessment of ventilator settings, it may be of clinical interest. PMID- 12130985 TI - Different strategies to keep the lung open: a study in isolated perfused rabbit lungs. AB - OBJECTIVE: Atelectatic alveoli can be recruited or kept open either by sustained inflation maneuvers or by positive end-expiratory pressure (PEEP). Little is known about potential interactions between both approaches. Especially, it is not known whether the recruiting effect of sustained inflation maneuvers is maintained in combination with a low PEEP, as suggested recently. In an attempt to answer this question, we combined sustained inflation maneuvers with either high or low PEEP. Both approaches were compared with a strategy likely to result in alveolar atelectasis and with another ensuring adequate alveolar recruitment by adjustment of PEEP alone. DESIGN: Randomized block design. SETTING: Laboratory. SUBJECTS: Isolated perfused rabbit lungs (n = 28). INTERVENTIONS: The lungs were ventilated with a tidal volume of 8 mL/kg. After stabilization, the lungs were randomized to one of four ventilatory strategies, which then were followed for 120 mins: a) PEEP 1 cm H2O (PEEP1, negative control); b) PEEP 1 cm H2O and 30 sec-sustained inflations (20 cm H2O) every 30 mins (SI-1); c) PEEP 3 cm H2O combined with sustained inflations (SI-3); and d) PEEP repeatedly adjusted following a previously established strategy ensuring full alveolar recruitment (DYN, positive control). MEASUREMENTS AND MAIN RESULTS: Distribution of ventilation and perfusion (Va/Q distribution) was analyzed by the multiple inert gas elimination technique. Volume-dependent compliance within the tidal volume was determined by using the slice method. Shunt and Va/Q mismatch significantly differed between SI-1 and SI-3, indicating full alveolar recruitment only in the latter. Data of SI-1 did not differ substantially from those of PEEP1, and data obtained in SI-3 were similar to those of DYN. CONCLUSIONS: First, enduring alveolar recruitment by sustained inflation maneuvers is only possible when the alveoli are stabilized thereafter by sufficient PEEP. Second, a ventilation strategy that uses repeated sustained inflations on a comparably high PEEP may not be superior to adequate adjustment of PEEP alone. PMID- 12130986 TI - Fructose-1,6-diphosphate attenuates acute lung injury induced by ischemia reperfusion in rats. AB - OBJECTIVE: To determine whether fructose-1,6-diphosphate (FDP) pretreatment can attenuate acute lung injury induced by ischemia-reperfusion in our isolated lung model in rats. DESIGN: Randomized, controlled study. SETTING: Animal care facility procedure room. SUBJECTS: Twenty-four adult male Sprague-Dawley rats each weighing 250-350 g. INTERVENTIONS: Typical acute lung injury in rats was induced successfully by 10 mins of hypoxia followed by 75 mins of ischemia and 50 mins of reperfusion. Ischemia-reperfusion significantly increased microvascular permeability as measured by the capillary filtration coefficient, lung weight gain, lung weight to body weight ratio, pulmonary arterial pressure, and protein concentration of bronchoalveolar lav-age fluid. MEASUREMENTS AND MAIN RESULTS: Pretreatment with FDP significantly attenuated the acute lung injury induced by ischemia-reperfusion as shown by a significant decrease in all of the assessed variables (p <.05 p <.001). The protective effect of FDP was nearly undetectable when promazine (an ecto-adenosine 5-triphosphatase inhibitor) was added before FDP pretreatment. CONCLUSIONS: Pretreatment with FDP significantly ameliorates acute lung injury induced by ischemia-reperfusion in rats. PMID- 12130987 TI - Prognostic factors, clinical course, and hospital outcome of patients with chronic obstructive pulmonary disease admitted to an intensive care unit for acute respiratory failure. AB - OBJECTIVE: To describe prognostic factors, clinical course, and hospital outcome of patients with chronic obstructive pulmonary disease admitted to an intensive care unit for acute respiratory failure. DESIGN: Analysis of prospectively collected data. SETTING: A multidisciplinary intensive care unit of an inner-city university hospital. PATIENTS: Patients with chronic obstructive pulmonary disease admitted to an intensive care unit for acute respiratory failure from August 1995 through July 1998. MEASUREMENTS AND MAIN RESULTS: Data were obtained concerning demographics, arterial blood gas, Acute Physiology and Chronic Health Evaluation (APACHE) II score, sepsis, mechanical ventilation, organ failure, complications, and hospital mortality rate. Fifty-nine percent of patients were male, 63% white, and 36% African-American; the mean age was 63.1 +/- 8.9 yrs. Noninvasive mechanical ventilation was tried in 40% of patients and was successful in 54% of them. Invasive mechanical ventilation was required in 61% of the 250 admissions. Sepsis developed in 31% of patients, nonpulmonary organ failure in 20%, pneumothorax in 3%, and acute respiratory distress syndrome in 2%. Multiple organ failure developed in 31% of patients with sepsis compared with 3% without sepsis (p <.0001). Predicted and observed hospital mortality rates were 30% and 15%, respectively. Differences in age and arterial carbon dioxide and oxygen tensions between survivors and nonsurvivors were not significant. Arterial pH was lower in nonsurvivors than in survivors (7.21 vs. 7.25, p =.0408). The APACHE II-predicted mortality rate (p =.0001; odds ratio, 1.046; 95% confidence interval, 1.022-1.070) and number of organ failures (p <.0001; odds ratio, 5.524; 95% confidence interval, 3.041-10.031) were independent predictors of hospital outcome; invasive mechanical ventilation was not an independent predictor. CONCLUSIONS: Physiologic abnormalities at admission to an intensive care unit and development of nonrespiratory organ failure are important predictors of hospital outcome for critically ill patients with chronic obstructive pulmonary disease who have acute respiratory failure. Improved outcome would require prevention and appropriate treatment of sepsis and multiple organ failure. PMID- 12130988 TI - Changes in intracranial pressure and cerebral autoregulation in patients with severe traumatic brain injury. AB - BACKGROUND: Impaired cerebral autoregulation is frequent after severe traumatic head injury. This could result in intracranial pressure fluctuating passively with the mean arterial pressure. OBJECTIVE: This study examines the influence of autoregulation on the amplitude and direction of changes in intracranial pressure in patients with severe head injuries during the management of cerebral perfusion pressure. DESIGN: Prospective study. SETTING: Neurosurgical intensive care unit PATIENTS: A total of 42 patients with severe head injuries. INTERVENTIONS: Continuous recording of cerebral blood flow velocity, intracranial pressure, and mean arterial pressure during the start or change of continuous norepinephrine infusion. MEASUREMENTS AND MAIN RESULTS: Cerebrovascular resistance was calculated from the cerebral perfusion pressure and middle cerebral artery blood flow velocity. The strength of autoregulation index was calculated as the ratio of the percentage of change in cerebrovascular resistance by the percentage of change in cerebral perfusion pressure before and after 121 changes in mean arterial pressure at constant ventilation between day 1 and day 18 after trauma. The strength of autoregulation index varied widely, indicating either preserved or severely perturbed autoregulation during hypotensive or hypertensive challenge in patients with or without intracranial hypertension at the basal state (strength of autoregulation index, 0.51 +/- 0.32 to 0.71 +/- 0.25). The change in intracranial pressure varied linearly with the strength of autoregulation index. There was a clinically significant change in intracranial pressure (> or =5 mm Hg) in the same direction as the change in mean arterial pressure in five tracings of three patients. This was caused by the mean arterial pressure dropping below the identified lower limit of autoregulation in three tracings for two patients. It seemed to be caused by a loss of cerebral autoregulation in the remaining two tracings for one patient. CONCLUSION: Cerebral perfusion pressure oriented therapy can be a safe way to reduce intracranial pressure, whatever the status of autoregulation, in almost all patients with severe head injuries. PMID- 12130989 TI - Antioxidant protection against iron in children with meningococcal sepsis. AB - OBJECTIVE: To assess antioxidant protection against iron-catalyzed reactive oxygen species in meningococcal sepsis and to establish whether severity of illness is related to deficiencies in these antioxidant systems. DESIGN: Prospective, controlled study. SETTING: Pediatric intensive care unit of a postgraduate teaching hospital. PATIENTS: Twenty children aged 6 months to 15 yrs (median, 5 yrs) with meningococcal septic shock were studied. Paired convalescent samples taken 8-10 wks after discharge were available in nine children. INTERVENTIONS: Routine management for meningococcal sepsis. MEASUREMENTS AND MAIN RESULTS: Patients were classified for disease severity using the Glasgow Meningococcal Septicaemia Prognostic Score. Paired acute and convalescent samples were compared. Transferrin level (1.77 +/- 0.08 g/L) and total iron-binding capacity (46.2 +/- 2.0 microM) were significantly decreased in acute patients compared with paired convalescent samples (2.85 +/- 0.10 g/L and 74.4 +/- 2.5 microM, respectively; p <.0001). The iron saturation of transferrin was significantly increased in acute disease (36.9% +/- 2.5%) compared with convalescence (18.8% +/- 1.5%; p =.0003). Iron-binding antioxidant protection was not significantly different in acute (81.4% +/- 1.7%) and paired convalescent samples (85.6% +/- 2.5%; p =.54). However, patients with more severe meningococcal septicemia (GMSPS, >10; n = 12) had significantly diminished protection (77.5% +/- 2.4%) compared with less severe disease (87.1% +/- 1.6%; p =.0028), and there was a significant correlation between disease severity and iron-binding antioxidant protection (R =.48; p =.00067) in acute disease. Paired ceruloplasmin levels were available in six patients and were decreased in acute disease (0.29 +/- 0.02 g/L) compared with convalescence (0.40 +/- 0.04 g/L), although not statistically significant (p =.076). However, there was a significant correlation between plasma ceruloplasmin and disease severity (Pearson product moment correlation, p =.038) in the acute patients. Iron oxidizing antioxidant assays were performed in four paired samples and were diminished in acute patients (53.3 +/- 4.4%) compared with convalescence (67.8 +/ 3.2%; p =.015). Acute samples demonstrated a significant relationship between iron-oxidizing antioxidant protection and both disease severity (r =.30; p =.012) and plasma ceruloplasmin levels (r =.48; p =.00067). CONCLUSIONS: Children with meningococcal septicemia exhibit abnormal plasma iron chemistry and decreased protection against iron-catalyzed oxidative damage. Such deficiencies correlate with disease severity. PMID- 12130990 TI - Survival and functional outcome in pediatric traumatic brain injury: a retrospective review and analysis of predictive factors. AB - OBJECTIVE: The aim of this study was to evaluate the relationship of patient care variables to survival and functional outcome in the pediatric population with traumatic brain injury. DESIGN: Retrospective chart review. SETTING: A 16-bed pediatric critical care unit in an academic community children's hospital. PATIENTS: A total of 320 consecutive pediatric patients with traumatic brain injuries admitted to our pediatric critical care unit between 1992 and 1996. INTERVENTIONS: Patients were managed using our standard traumatic brain injury protocol. MEASUREMENTS AND MAIN RESULTS: A total of 230 patient variables encompassing demographic data, prehospital, emergency department, and pediatric critical care unit care were recorded. A total of 79 patients were severely injured, with admitting Glasgow Coma Scale scores of < or =10. There were 18 deaths. Only two patients survived without cognition. Ninety-five of 302 survivors required inpatient rehabilitation. Of these, 73 were old enough to be compared using FIMTM scores. At the time of discharge from rehabilitation, 52 patients (71%) were functioning independently, 20 (27%) were moderately dependent, and one patient was completely dependent. Analysis of variables with respect to survival revealed that an inability to maintain a cerebral perfusion pressure of > or =50 mm Hg on the first pediatric critical care unit day (p =.0002) and the presence of bradycardia in the emergency department (p =.0139) were the strongest factors associated with mortality. By using the regression equation generated from this model, we could correctly identify survivors and nonsurvivors with a predictive value of 94%. CONCLUSIONS: The ability to maintain a cerebral perfusion pressure of > or =50 mm Hg was the single most important predictor of traumatic brain injury survival in this study. This suggests that monitoring and optimizing cerebral perfusion pressure is critical to the management of these patients. The relationship between cognitive outcome and therapeutic interventions used to optimize cerebral perfusion pressure is unclear and requires further evaluation in a large prospective study. PMID- 12130991 TI - Multicenter clinical research in adult critical care. AB - OBJECTIVE: To describe the development, organization, and operation of several collaborative groups conducting investigator-initiated multicenter clinical research in adult critical care. DESIGN: To review the process by which investigator-initiated critical care clinical research groups were created using examples from Europe, Australia, the United States, and Canada. Various models of group structure and function are discussed, highlighting complementary approaches to protocol development, multicenter study management, and project funding. DATA SOURCES: Published peer review research and unpublished terms of reference documents on the structure and function of these groups. DATA SYNTHESIS: The overall goal of clinical critical care research groups engaged in multicenter studies is to improve patient outcomes through conducting large, rigorous investigations. Research programs we reviewed included the following: a) multicenter epidemiologic studies and surveys; b) technology evaluations of mechanical ventilation; c) investigations focused on three priority fields (acute lung injury, infection, and acute brain injury); d) a series of randomized trials of treatments for one syndrome (acute respiratory distress syndrome); and e) diverse methodologies addressing several clinical problems. The structure and function of these research groups differ according to their historical development, research culture, and enabling resources. Specific protocols emerge from clinical questions generated by investigators or from collectively prioritized research agendas. Project funding includes government support, peer review grants, intensive care foundations, industry, local hospital funds, and hybrid models. Infrastructure for study management varies widely. CONCLUSIONS: Several national and international groups have engaged in investigator-initiated multicenter critical care research. The development, organization, and operational methods of these groups illustrate several collaborative models for clinical investigations in the intensive care unit. Common characteristics of these groups are a cohesive spirit, a sense of mission to achieve shared research goals, and acknowledgment that such an organization is much more than the sum of its parts. PMID- 12130992 TI - Lemierre's syndrome: an unusual cause of sepsis and abdominal pain. AB - OBJECTIVE: To describe a patient with Lemierre's syndrome who presented with acute abdominal findings and to describe the evaluation and treatment of this syndrome. DESIGN: Case report. SETTING: A 38-bed, pediatric intensive care unit at a tertiary care children's hospital. PATIENT: One patient presenting with signs of severe sepsis and acute abdominal pain. INTERVENTIONS: Intravenous hydration, inotropic support, thoracostomy tube drainage of a pleural effusion, and prolonged antimicrobial therapy. MEASUREMENT AND MAIN RESULTS: The patient presented with severe sepsis and abdominal pain. After Fusobacterium necrophorum grew in blood cultures, anaerobic antimicrobial therapy was initiated. Doppler duplex ultrasonography and magnetic resonance venography demonstrated thrombus formation in the left internal jugular vein. Computed tomography of the chest demonstrated bibasilar lung nodules consistent with septic emboli. The patient was treated with ampicillin-sulbactam and metronidazole intravenously for 3 wks, followed by a 3-wk course of oral amoxicillin/clavulanate. He had a good recovery, and his thrombus had resolved at the time of discharge. CONCLUSION: Lemierre's syndrome occurs in young, otherwise healthy patients, and it thus needs to remain high on the differential diagnosis for this group of patients presenting with severe sepsis. The diagnosis can be confounded by a lack of symptoms of pharyngitis at the time of presentation and end-organ dysfunction associated with severe sepsis, suggesting alternative sources of infection. PMID- 12130993 TI - All that's gold does not glitter: effects of an increase in respiratory rate on pulmonary mechanics and CO2 kinetics in acute respiratory failure. PMID- 12130994 TI - Family satisfaction surveys to improve the fit between the intensive care unit and its concept. PMID- 12130995 TI - Insights into the immune dysfunction associated with thermal injury. PMID- 12130996 TI - Promoting enteral feeding 101. PMID- 12130997 TI - Is there a benefit to postpyloric feeding? PMID- 12130998 TI - How should patients feel about prolonged mechanical ventilation? Can we predict their choices, and if so, should we? PMID- 12130999 TI - There and back again: does prone positioning have any value in respiratory failure? PMID- 12131000 TI - Nosocomial infection in neonatology: in search of the best score, in search of an oracle. PMID- 12131001 TI - Sedation in the intensive care unit: refining the models and defining the questions. PMID- 12131002 TI - Interferon-gamma: titration of inflammation. PMID- 12131003 TI - Increased serum creatinine: a marker for adverse outcome before and after cardiac surgery. PMID- 12131004 TI - Cytokines and therapeutic hypothermia. PMID- 12131005 TI - Fas-mediated alveolar epithelial cell apoptosis in acute lung injury: friend or foe? PMID- 12131006 TI - Scorpion sting--the inflammatory dimension. PMID- 12131007 TI - Force or finesse: maintaining functional residual capacity while practicing lung protective ventilation. PMID- 12131008 TI - Increasing blood pressure causes a decrease in intracranial pressure in patients with brain injury. PMID- 12131009 TI - Short-term high-volume hemofiltration in sepsis: perhaps the right way is to start with. PMID- 12131010 TI - Use of a capnograph during feeding tube insertion. PMID- 12131011 TI - Hyperkalemic cardiac arrest: the method chosen depends on the local circumstances. PMID- 12131012 TI - Questions of clinical importance and statistical significance. PMID- 12131013 TI - Predicting mortality or recognizing death in meningococcal disease? Pediatric Risk of Mortality (PRISM) and Pediatric Index of Mortality (PIM). PMID- 12131014 TI - Heart rate variability after abdominal aortic surgery. PMID- 12131016 TI - Prognostic significance of plasma concentrations of transforming growth factor beta in patients with coronary artery disease. AB - BACKGROUND: Cytokines play an important role in modulating inflammatory and proliferative responses, including atherosclerosis. Transforming growth factor beta (TGF-beta) and macrophage-colony stimulating factor (M-CSF) are one of the major antiinflammatory and proinflammatory cytokines, respectively. We have previously demonstrated that plasma concentrations of TGF-beta are decreased while those of M-CSF are increased in patients with coronary artery disease (CAD). In this study, we examined whether those alterations in plasma levels of cytokines have a prognostic significance in patients with CAD. METHODS AND RESULTS: Sixty-eight consecutive patients with proven CAD were studied. The plasma concentrations of TGF-beta and those of M-CSF were measured by enzyme linked immunosorbent assay (ELISA). They were divided into groups: high (> or =6 ng/ml, n = 19) and low (<6 ng/ml, n = 49) TGF-beta groups and high (>500 ng/ml, n = 52) and low (< or =500 ng/ml, n = 16) M-CSF groups. The long-term prognosis of these patients was prospectively followed up for a mean period of 979 +/- 27 days. The prognosis was analyzed by Kaplan-Meier analysis in terms of total survival, survival without myocardial infarction, survival without cardiovascular events and survival without coronary interventions. The analysis showed that the low TGF-beta group had a significantly poor prognosis in terms of survival without cardiovascular events and survival without coronary interventions as compared with the high TGF-beta group (both P < 0.05), while other prognoses were comparable between the two groups. By contrast, no significant prognostic influence was noted regarding M-CSF. CONCLUSIONS: These results suggest that plasma concentrations of TGF-beta may have a prognostic significance in patients with CAD. PMID- 12131017 TI - Comparison of low-dose dobutamine stress echocardiography and echocardiography during glucose-insulin-potassium infusion for detection of myocardial viability after anterior myocardial infarction. AB - BACKGROUND: Low-dose dobutamine stress echocardiography (LDDSE) is one of the methods most used to assess myocardial viability. Glucose-insulin-potassium (GIK) infusion has been shown to increase contraction of the ischemic zone. The aim of this study was to compare LDDSE and echocardiography during GIK infusion for detection of myocardial viability. METHODS: Thirty-two patients who had first anterior myocardial infarction (MI) without previous MI were included in the study. Echocardiographic evaluation was carried out on the 7th +/- 2 days after MI. During continuous electrocardiographic, blood pressure and echocardiographic monitoring, an intravenous infusion of dobutamine (3 microg/kg body weight/min) was started with an infusion pump, continued for 5 min and then increased to 5 microg/kg/min and 10 microg/kg/min for another 5 min. The GIK protocol consisted of a fixed dose of insulin (100 microU/kg/h intravenously) and a variable glucose/potassium infusion rate. GIK echocardiography was done at baseline and after 60 min of GIK. The detected viable myocardium was defined as one or two scores decreasing in at least two adjacent abnormal segments during LDDSE and GIK echocardiography. RESULTS: Under resting conditions 225 segments (44%) were normokinetic, 21 segments (4%) dyskinetic, 117 segments (23%) akinetic and 149 segments (29%) hypokinetic. Viability was detected in 20% (57 segments) of the asynergic segments at baseline with GIK echocardiography and in 22% (62 segments) of those segments with LDDSE (P < 0.05). Left ventricular wall motion score index at baseline was 1.87 and it decreased significantly indicating improvement in left ventricular systolic function during both LDDSE and GIK echocardiography (P < 0.001, versus 1.75 and 1.76 respectively). The agreement between LDDSE and GIK echocardiography for detection of myocardial viability was 96%. CONCLUSION: We have shown that GIK echocardiography is similar to LDDSE for detection of myocardial viability. With the support of further clinical studies GIK echocardiography could be used to detect myocardial viability after acute MI. PMID- 12131018 TI - Relationship of C-reactive protein to presence and severity of coronary atherosclerosis in patients with stable angina pectoris or a pathological exercise test. AB - BACKGROUND: C-reactive protein (CRP) level is a sensitive marker of inflammation and a probable predictor of cardiovascular risk. The aim of this study was to assess the relationship between the presence and the extent of coronary atherosclerosis and CRP level in patients referred for coronary angiography for stable angina pectoris or a pathological exercise test. PATIENTS AND METHODS: A group of 200 patients were prospectively analyzed for the relationship between the presence and extent of coronary atherosclerosis and high-sensitivity CRP. Patients with stable angina pectoris or a pathological exercise test were included. RESULTS: For the whole group the CRP geometric mean was 2.92 mg/l and the median 3.0 mg/l. There was no difference between groups of patients with different extents of coronary lesions (P = 0.320, one-way analysis of variance). In patients without significant coronary disease the CRP geometric mean was 3.1 (2.28-4.21) mg/l with a variation coefficient of 118.4%; in patients with coronary artery disease the geometric mean was 2.83 (2.34-3.43) mg/l with a variation coefficient of 104.0%. The difference in CRP between both groups was not significant (P = 0.601). There was also no significant difference in CRP levels between groups of patients with and without a history of myocardial infarction (2.65 (2.08-3.36) mg/l and 3.18 (2.54-3.98) mg/l, P = 0.266) respectively. There was no correlation between the classification of angina pectoris and the logarithm of CRP level (P = 0.331). This relationship was not confirmed even in the group of patients with significant coronary artery disease (P = 0.693). CONCLUSIONS: CRP level is not related to the extent or the presence of coronary atherosclerosis assessed by coronary angiography, history of myocardial infarction or class of stable angina pectoris in patients referred for coronary angiography for stable angina pectoris or a pathological exercise test. PMID- 12131019 TI - Coronary collateralization: determinants of adequate distal vessel filling after arterial occlusion. AB - BACKGROUND: The protective effect of collateral vessels in coronary artery disease (CAD) is well established. Little is known, however, about factors that influence collateral formation. METHODS: We studied the coronary angiograms of 200 consecutive patients with single-vessel coronary artery occlusion. Patients were excluded if obstructive stenoses were present in other vessels or if prior revascularization had been undertaken. Collateral circulation to the occluded artery was graded as 'poor' (no or incomplete filling) or 'rich' (complete filling). Patient characteristics, including mode of presentation, medications and CAD risk factors, were assessed. RESULTS: Positive univariate correlates of rich collaterals included increasing age [odds ratio (OR) 1.03, P = 0.016], 'statin' use (OR 2.50, P = 0.005), nitrate use (OR 1.96, P = 0.034), calcium channel blocker (CCB) use (OR 4.07, P < 0.001), presentation with stable angina (OR 2.34, P = 0.006), longer time since diagnosis of CAD (OR 1.12, P = 0.002) and history of hyperlipidemia (OR 3.55, P < 0.001). Significantly poorer collateralization was observed in the setting of acute myocardial infarction (MI) (OR 0.23, P < 0.001), diabetes mellitus (OR 0.33, P = 0.003), impaired left ventricular function (OR 0.64, P = 0.015) and occlusion of the left anterior descending coronary artery (LAD) (OR 0.28, P < 0.001). On multivariate analysis, rich collateralization was associated with hyperlipidemia (P = 0.003) and CCB use (P = 0.028). Independent predictors of poor collaterals were presence of diabetes (P < 0.001), LAD occlusion (P = 0.001) and presentation with acute MI (P = 0.017). CONCLUSION: Diabetes mellitus, occlusion of the LAD and presentation with acute MI are independently associated with poor distal vessel collateralization, whereas hyperlipidemia and use of CCBs are associated with rich collateralization. Factors determining coronary collateral formation may in turn influence outcomes after coronary artery occlusion. PMID- 12131020 TI - Circulating T-lymphocyte activation in patients with variant angina. AB - BACKGROUND: Both experimental and pathological studies suggest that immune response and inflammation may play an important role in the pathogenesis of coronary spasm. DESIGN: To elucidate the role of systemic immune and inflammatory responses in the pathogenesis of coronary spasm, we studied circulating T lymphocyte activation in variant angina patients (VAPs), stable effort angina patients (EAPs) and in control participants. METHODS: Twenty documented VAPs, 13 EAPs and 20 control participants were studied. To evaluate T-lymphocyte activation, T-lymphocyte surface antigen expression, including CD3, CD4, CD8 and HLA-DR, was measured by two-colour flow cytometric analysis. Serum-soluble interleukin-2 receptor (sIL-2R) and C-reactive protein (CRP) were also measured by enzyme-linked immunosorbent assay. We restudied 10 of the VAPs to investigate the relationship between the disease activity of variant angina and T-lymphocyte activation. RESULTS: The percentage of CD3+/DR+ T-lymphocytes in VAPs (14.8%) was significantly higher than in EAPs (10.7%, P < 0.05) and control participants (9.7%, P < 0.005); however, levels of sIL-2R were the same among the three groups. Levels of CRP were within normal range in all VAPs. The percentage of CD8+/DR+ T-lymphocytes was significantly higher in VAPs (9.5%, P < 0.005) than in EAPs (5.5%) and control participants (5.9%), whereas the percentage of CD4+/DR+ T lymphocytes was similar among the three groups. The percentage of activated T lymphocytes in VAPs was unchanged during the follow-up period (mean intervals, 10 months). CONCLUSIONS: These results indicate that the chronic activation of T lymphocytes, especially CD8+ T-lymphocytes, may be involved in the pathogenesis of coronary spasm. PMID- 12131021 TI - Continuous vectorcardiography is superior to standard electrocardiography in the prediction of long-term outcome after thrombolysis in patients with acute myocardial infarction. AB - BACKGROUND: Thrombolytic therapy results in reperfusion of the occluded coronary vessel in approximately 75% of treated patients with acute myocardial infarction (AMI). Unsuccessful thrombolysis results in impaired outcome. This study was undertaken to evaluate reperfusion assessments with 12-lead standard static electrocardiography (ECG) and continuous vectorcardiography (VCG) in AMI patients treated with thrombolytic therapy, with particular emphasis on the value of these assessments in relation to long-term outcome. METHODS: ST-recovery analysis 90 and 180 min after the start of thrombolytic therapy was performed by repeated ECG and by VCG in 63 AMI patients. Median follow-up was 255 days. RESULTS: No significant differences in long-term outcome were found between patients with or without obtained reperfusion, as assessed by ECG. For VCG, we found significant elevated relative risks for experiencing death (relative risk = 11.00, confidence interval = 2.70-44.90); P = 0.0008 for the group with ST-vector magnitude recovery of less than 50% at 90 min from start of thrombolytic therapy. CONCLUSION: We demonstrated that early reperfusion assessment with VCG enables the prediction of long-term outcome and is superior to reperfusion assessment with standard static ECG in this regard. We therefore recommend continuous ischemia monitoring of AMI patients treated with thrombolytic therapy as a routine procedure. PMID- 12131022 TI - Elevated body temperature during myocardial ischemia/reperfusion exacerbates necrosis and worsens no-reflow. AB - PURPOSE: The effects of an elevated body temperature on infarct size were tested in a rabbit model of ischemia/reperfusion. METHODS: Before coronary artery occlusion, body temperature was raised from baseline at 38.6 +/- 0.1 degrees C to 40.3 +/- 0.2 degrees C in nine treated rabbits. Temperature in eight normothermic rabbits was 38.4 +/- 0.2 degrees C. Both groups received 30 min coronary occlusion and 3 h reperfusion. RESULTS: In normothermic rabbits, 36 +/- 6% of the ischemic risk region became necrotic but in hyperthermic rabbits myocardial necrosis was significantly increased to 57 +/- 3% of the risk region (P < 0.005) despite similar risk regions and an equal degree of regional myocardial blood flow (RMBF) reduction during ischemia in both groups. Infarct size correlated positively with body temperature (r = 0.66, P < 0.004). RMBF was 43% lower during reperfusion in the previously ischemic areas of hyperthermic hearts compared with the control group (P < 0.04), suggesting worsened no-reflow. CONCLUSION: Elevation in body temperature by even a few degrees can aggravate necrosis during acute myocardial infarction and worsens no-reflow. PMID- 12131023 TI - Twenty-eight-day efficacy and phamacokinetics of the sirolimus-eluting stent. AB - BACKGROUND: In-stent restenosis is caused by neointimal hyperplasia. Sirolimus (rapamycin; Wyeth Research, Radnor, Pennsylvania, USA) inhibits vascular smooth muscle cell proliferation and we evaluated the efficacy of sirolimus in reducing neointimal formation in a rabbit iliac model and in-vivo pharmacokinetics in the porcine coronary model. DESIGN: Randomized, blinded, prospective animal study. METHODS: Bilateral rabbit iliac artery stent implantation was performed using crossflex stents (Cordis Corporation, Warren, New Jersey, USA) coated with sirolimus incorporated in a nonerodable polymer. Arteries were randomized to one of four stent groups: uncoated stents (n = 8); polymer control stents (n = 10); low-dose sirolimus-eluting stents (n = 9); and high-dose sirolimus-eluting stents (n = 10). Histomorphometry was performed at 28 days. Arterial tissue and stents were retrieved at 8, 14 and 28 days and blood samples were obtained daily during the first week. RESULTS: Treatment with low-dose sirolimus was associated with a 23% (P = NS) reduction in neointimal area and treatment with high-dose sirolimus with a 45% (P < 0.05) reduction. Sustained drug release from the stent and prolonged intramural arterial deposition were confirmed for up to 28 days. No detectable sirolimus was found in the blood after 2 days. CONCLUSION: Controlled release local delivery of a cell-cycle inhibitor from a nonerodable polymer coated stent reduced neointimal formation in rabbit iliac arteries in a dose dependent manner and represents a promising strategy for preventing restenosis. PMID- 12131024 TI - Beta-cyclodextrin tetradecasulfate, a novel cyclic oligosaccharide, inhibits thrombus and neointimal formation after coronary vascular injury. AB - BACKGROUND: Neointimal formation is a major cause of restenosis after interventional vascular procedures. Beta-cyclodextrin tetradecasulfate (beta-CDT) has been shown to inhibit fibroblast growth factor activity and we hypothesized that beta-CDT would reduce intimal formation. DESIGN: Three studies were performed: (1) pharmacokinetics of oral and intravenous beta-CDT and determination of optimal dose, (2) determination of efficacy of oral and intravenous beta-CDT in reducing neointimal formation after balloon-overstretch injury and (3) determination of the effect of beta-CDT on cellular proliferation, factor Xa activity, activated clotting time, activated partial thromboplastin time and thrombus formation. METHODS: Pharmacokinetics were determined in eight domestic swine following administration of oral beta-CDT and intravenous beta-CDT at three doses each. In the efficacy study, balloon-overstretch injury of 37 pigs (69 arteries) was performed and randomized into three groups (n = 23 arteries/group): control, oral administration of 300 mg beta-CDT/kg per day or intravenous infusion of 100 mg beta-CDT/kg per day. Animals were sacrificed 14 days later. Cellular proliferation and mural thrombus were determined in six arteries/group at 5 days and endothelial coverage was evaluated at 5 and 14 days. RESULTS: Oral and intravenous beta-CDT reduced the intimal hyperplasia area normalized to injury index by 24 and 48%, respectively: control, 3.03 +/- 0.75 mm2, oral, 2.31 +/- 0.83 mm2 (P = 0.004) and intravenous, 1.67 +/- 0.73 mm2 (P = 0.0000002). beta-CDT reduced cellular proliferation (control, 55 +/- 18%, oral, 35 +/- 7%, P = 0.03 and intravenous, 30 +/- 12%, P = 0.01) and mural thrombus formation (control, 0.84 +/- 0.4 mm2, oral, 0.44 +/- 0.14 mm2, P = 0.04, intravenous, 0.42 +/- 0.09 mm2, P = 0.03). Endothelial coverage was increased in the experimental groups (P = 0.008, oral versus control, P < 0.0001, intravenous versus control). Factor Xa activity was inhibited 9-10 fold following intravenous administration while oral administration demonstrated no effect. CONCLUSIONS: Both oral and intravenous formation of beta-CDT reduced intimal hyperplasia with the greatest reduction in the intravenous group. We postulate that beta-CDT was effective by the combination of increasing endothelial coverage, reducing mural thrombus formation, inhibiting factor Xa activity and reducing cellular proliferation. PMID- 12131025 TI - Quantitative assessment of diffuse coronary artery disease using a three dimensional reconstruction method compared with intravascular ultrasound images. PMID- 12131026 TI - Eye banking in Latin America. PMID- 12131027 TI - Efficacy and safety of recurrent pterygium surgery using human processed pericardium. AB - PURPOSE: To investigate the safety and efficacy of human processed pericardium used as an onlay after pterygium excision. METHODS: Twenty-five eyes of 25 patients (11 female, 14 male) with recurrent pterygium were included in this study. The median age was 50 years (range 24-89 years). Twenty-eight percent of the eyes previously had been operated on two or more times. The mean follow-up was 9.4 months (+/- 2.1 months, range 8-15 months). Following bare-sclera surgical removal of recurrent pterygium, a patch of processed human pericardium was sutured to cover the area of excision. The pericardium was not covered with conjunctiva, nor were adjuvant radiation or antimetabolite administered. All patients were treated with a combination of dexamethasone 0.1% / chloramphenicol drops three times per day for 1 month. Ketorolac tromethamine 0.5% three times per day was added to this regimen after complete corneal reepithelialization. RESULTS: Recurrence (any growth >1 mm onto the cornea) was detected in 12 patients (48%). The mean time of recurrence was 4.1 +/- 1.7 months. Three patients (12.0%) presented a regrowth of fibrovascular tissue not reaching 1 mm of the cornea. Pyogenic granulomas occurred in three patients, and all of them required surgical excision. Corneal thinning was present in one patient and was treated with a therapeutic contact lens and artificial tears with complete resolution and vascularization of the thinned area. No decrease in visual acuity was observed in any patient. CONCLUSIONS: The use of processed pericardium in pterygium surgery is a safe procedure but is associated with a relatively high rate of recurrence. It should only be considered as an option in managing recurrent pterygium when conjunctival autografting is not an available alternative. PMID- 12131028 TI - Delayed-onset mycobacterial keratitis after LASIK. AB - PURPOSE: To describe the time course, diagnosis, clinical features, and treatment of seven patients with Mycobacterium szulgai keratitis that developed from 7 to 24 weeks after laser in situ keratomileusis (LASIK). METHODS: Seven of 30 eyes of 18 patients were identified with keratitis after LASIK. The first two patients presented 12 to 14 weeks after LASIK; nontuberculous mycobacteria were identified 1 month after the flaps were cultured. Patient recall identified three additional cases by culture and two cases by clinical features alone. Pulsed-field gel electrophoresis (PFGE) was used to type the isolates, and treatment was modified based on susceptibilities. RESULTS: M. szulgai was identified in five patients for whom cultures were performed, but response to empiric therapy based on cultures proved unsatisfactory. The keratitis resolved in all patients with treatment including clarithromycin based on susceptibilities. Medical therapy was sufficient, although one patient required flap amputation. Six of seven patients recovered best-corrected visual acuity (BCVA), while one patient lost one line of BCVA. Two patients lost one line of postoperative uncorrected visual acuity (UCVA), two patients gained one line of UCVA, and three patients recovered postoperative UCVA. PFGE analysis revealed that the M. szulgai strains were identical, and the infection source was contaminated ice used to chill syringes for saline lavage. CONCLUSIONS: Nontuberculous mycobacterial keratitis after LASIK is a diagnostic and management challenge, but outcomes can be preserved with treatment based on susceptibilities. This cluster underscores the importance of adherence to sterile protocol during LASIK. PMID- 12131029 TI - The epidemiological features and laboratory results of fungal keratitis: a 10 year review at a referral eye care center in South India. AB - PURPOSE: To report the epidemiological features and laboratory results of 1,352 cases of fungal keratitis diagnosed at the L.V. Prasad Eye Institute (LVPEI) in south India. METHODS: The medical and microbiology records of 1,352 culture proven cases (1,354 eyes) of fungal keratitis diagnosed at the LVPEI between January 1991 to December 2000 was retrospectively reviewed for demographic features, risk factors, seasonal variation, and laboratory findings. RESULTS: Males (962) were affected significantly more (p< 0.0001) than females (390). Of 1,352 patients, 853 (64.4%) were in the younger age group (16-49 years). Ocular trauma predisposed to infection in 736 (54.4%) of 1,354 eyes. There was a higher incidence of fungal keratitis during the monsoon and winter than summer. A fungal cause was established by smears of corneal scrapings in 1,277 (95.4%) eyes. The potassium hydroxide preparation (KOH), Calcofluor white (CFW), Gram-, and Giemsa stained smears revealed fungus in 1,219 (91.0%), 1,224 (91.4%), 1,181 (88.2%), and 1,139 (85.1%) eyes, respectively. Fusarium(506, 37.2%) and Aspergillus species (417, 30.7%) predominated the hyaline fungal spectrum (1,133) and Curvularia species (39, 2.8%) were the highest among the dematiaceous isolates (218). CONCLUSIONS: To the best of our knowledge, this review presents the epidemiological features and laboratory results of the largest series of fungal keratitis ever reported in the literature. Keratomycosis is predominant in young adults with trauma as the major predisposing factor. With fungal keratitis being a major ophthalmologic problem in the tropical regions of the world, data available on the epidemiological features of a large series would greatly help medical practitioners at primary and secondary health care centers in the management of the disease. A simple KOH preparation of corneal scraping alone is highly beneficial in confirming the diagnosis. PMID- 12131030 TI - Sporadic diffuse lamellar keratitis (DLK) after LASIK. AB - PURPOSE: To examine the incidence of sporadic diffuse lamellar keratitis (DLK) in a large series of LASIK eyes and to suggest the hypothesis that the etiology of sporadic DLK differs from that of epidemic DLK. METHODS: The incidence and severity of DLK was noted in 1352 consecutive eyes that had primary LASIK for myopia or hyperopia and 217 consecutive eyes that had LASIK enhancement. RESULTS: Twelve of the eyes having primary LASIK had stage 1 DLK and 5 had stage 2 DLK (.9% total). No eyes had stage 3 or stage 4 DLK. Three of the 217 eyes (1.4%) that had LASIK enhancement had stage 1 DLK. The difference in the rate of DLK for primary LASIK compared with LASIK enhancement was not statistically significant (p = 0.69). All eyes responded to intensive corticosteroid therapy, with the addition of flap lifting and irrigation for the eyes with stage 2 DLK. Two of the eyes (one primary LASIK and one LASIK-enhancement) had implanted epithelial nests associated with the DLK. None of the cases of DLK occurred in eyes of patients who had surgery on the same operating day in this series. Two other eyes that had epithelial abrasions more than 3 months after LASIK or LASIK enhancement developed stage 1 DLK. CONCLUSIONS: Many cases of sporadic DLK, including cases associated with epithelial trauma after LASIK, are likely attributable to endogenous factors that trigger inflammation. One trigger is the release of epithelium-derived cytokines such as interleukin-1 that stimulate keratocytes to produce chemokines that are chemotactic to inflammatory cells. Cells likely accumulate at the interface because it is potential space, representing a path of least resistance for cell movement. Some sporadic cases may also be related to exogenous factors such as Betadine. Epidemic DLK is likely associated with exogenous factors that stimulate inflammation, such as endotoxin contaminating sterilizer reservoirs or detergents on instruments. PMID- 12131031 TI - Correction of myopia and astigmatism after penetrating keratoplasty with laser in situ keratomileusis. AB - PURPOSE: We evaluated whether laser in situ keratomileusis (LASIK) was a safe and effective treatment for myopia and astigmatism after penetrating keratoplasty (PK). METHODS: We performed a retrospective review of medical records of all the patients who underwent LASIK following PK at the University of Minnesota between January 1999 and March 2000. RESULTS: Seventeen patients (20 eyes) underwent LASIK following PK between January 1999 and March 2000. Mean age of the patients at the time of LASIK was 37 years (range, 20-62). Keratoconus was the indication for PK in the majority of the eyes (73.7%). Anisometropia and/or contact lens intolerance was the indication for LASIK following PK. No intraoperative complications occurred. Following LASIK, the best spectacle-corrected visual acuity remained within 1 line of preoperative visual acuity in 94.7% of the eyes. The mean sphere was reduced by 3.93 diopters (80.0%) and the mean cylinder was reduced by 2.83 diopters (69.9%) from the preoperative values at the last follow up visit. Uncorrected visual acuity became 20/40 or better in 73.7% of the eyes after LASIK. CONCLUSIONS: LASIK is a safe procedure in eyes in which PK has previously been performed. LASIK is effective in the treatment of myopia and astigmatism following PK. PMID- 12131032 TI - Recurrence of corneal dystrophy resulting from an R124H Big-h3 mutation after phototherapeutic keratectomy. AB - PURPOSE: The purpose of the study was to investigate the recurrence-free interval after phototherapeutic keratectomy (PTK) in patients with corneal dystrophies resulting from an Arg124His (R124H) mutation of the Big-h3 gene. METHODS: Patients with corneal dystrophy resulting from a genetically confirmed Big-h3 R124H mutation were examined with a slit lamp. The patients were divided into two groups on the basis of the mutation genotype, and the recurrence-free interval was analyzed. RESULTS: In the 4 eyes of 3 homozygous patients, the mean (+/- standard deviation [SD]) recurrence-free interval was 9.5 +/- 3.1 months, whereas in the 7 eyes of 4 heterozygous patients it was 38.4 +/- 6.2 months. The former interval was statistically shorter than the latter (Kaplan-Meier survival analysis with log-rank test, p = 0.004). CONCLUSIONS: These results strongly suggest that the mutation genotype of Big-h3 gene determined the recurrence-free interval as well as the clinical picture after PTK. Therefore, PTK should be considered for patients with Big-h3 R124H corneal dystrophy, on the basis of the expected recurrence-free interval deduced from molecular analysis of the zygosity of the Big-h3 R124H mutation. PMID- 12131033 TI - The relationship between first postoperative day epithelial status and eventual health of the ocular surface in penetrating keratoplasty. AB - PURPOSE: To determine a possible relationship between donor epithelial status on the first postoperative day after keratoplasty and the eventual health of the corneal surface. METHODS: We analyzed 91 patients who underwent penetrating corneal transplantation between January 1998 and January 2000, monitoring the epithelial status of the corneas with fluorescein staining using slit-lamp biomicroscopy. Recipient pre- and postoperative variables and donor characteristics were recorded. Macroepithelial defects were classified into three groups according to the extent of the epithelial defect. The results on the first postoperative day were compared with the first and third operative month. Donor and recipient variables were compared with the epithelial status on the first and third month as well. RESULTS: On the first postoperative day, 64.84% of the patients had epithelial defects, 10.99% had defects at the 1-month postoperative visit, and none had defects at the third month. Graft recipients with macroepithelial defects in the first postoperative month were older, had a higher prevalence of blepharitis, higher prevalence of inadequate eye hydration, and slightly increased corneal sensation compared with the group without epithelial defects; however, none of these trends were statistically significant. Patients with macroepithelial defects in the first postoperative month received older donor tissue, and the average preservation-to-surgery time was longer. These donor variables, however, were not significant statistically (p value >0.10) in determining outcome of the epithelial status at the first or third months. CONCLUSIONS: Our results suggest that the epithelial status on the first postoperative day is not predictive of surface integrity at 1-month postoperative (p value is 0.2676 for the likehood ratio test). The epithelial status on the first postoperative day is not predictive of the status of the third month after keratoplasty, because none of the 91 patients had epithelial defects after 3 months. PMID- 12131034 TI - Performance and repeatability of the NEI-VFQ-25 in patients with dry eye. AB - PURPOSE: The use of the National Eye Institute Visual Function Questionnaire (NEI VFQ) has increased as a method of assessing patients' impressions of their vision specific quality of life. The purpose of this study was to assess the performance and test-retest repeatability of the 25-question format of the NEI-VFQ in patients with dry eye. METHODS: The self-administered NEI-VFQ-25 was administered to 75 patients with dry eye on two occasions in a university-based optometry practice. Dry eye severity was assessed with use of the European criteria for dry eye. The weighted kappa statistic (kappa(w)) was used to evaluate test-retest repeatability of the NEI-VFQ-25 individual test questions, and the 95% limits of agreement and the intraclass correlation coefficients were calculated for the overall VFQ and subscale scores. RESULTS: With use of the European dry eye criteria, 21.9% of participants were classified with moderate to severe dry eye. For the sample, the ocular pain subscale score was lower (indicating more ocular pain) than published normative values. Repeatability of individual NEI-VFQ-25 questions ranged from moderate to substantial (kappa(w) values: 0.42 [pain and discomfort] to 0.90 [stay home because of vision]). The intervisit mean (+/- standard deviation) difference in the overall VFQ score was -0.66 +/- 4.26 (95% limits of agreement: -9.02, 7.69), and the intraclass correlation coefficient for the ocular pain subscale was 0.57. CONCLUSION: Patients with dry eye have lower ocular pain subscale scores. The repeatability of the overall NEI-VFQ score and the subscale scores was moderate to high, and it may be influenced by the number of questions in each subscale. PMID- 12131035 TI - Role of protein kinase C signaling in collagen degradation by rabbit corneal fibroblasts cultured in three-dimensional collagen gels. AB - PURPOSE: To understand the mechanism of corneal ulceration by characterizing the intracellular signaling pathways that regulate collagen degradation by corneal fibroblasts cultured in three-dimensional type I collagen gels. Specifically, the potential roles of protein kinase C (PKC) and protein kinase A (PKA) in collagen degradation were investigated. METHODS: Rabbit corneal fibroblasts were cultured in three-dimensional type I collagen gels for 24 hours in the presence of plasminogen and in the absence or presence of activators or inhibitors of PKC or PKA. Degradation of collagen fibrils was then evaluated by measurement of released hydroxyproline, and the production of matrix metalloproteinases (MMPs) was assessed by gelatin zymography and immunoblot analysis. RESULTS: The PKC activator phorbol 12-myristate 13-acetate (PMA) increased the extent of collagen degradation by corneal fibroblasts in a dose-dependent manner, with the maximal effect apparent at a concentration of 0.1 microM. The inactive analog 4alpha-PMA had no effect on collagen degradation. The PKC inhibitor H-7 reduced the extent of collagen degradation by corneal fibroblasts in the absence or presence of PMA. Phorbol 12-myristate 13-acetate also increased the production of proMMP-1, -3, and -9 by corneal fibroblasts, whereas H-7 inhibited this effect. Neither the PKA activators 8-bromo-cAMP, isobutylmethylxanthine, and forskolin nor the PKA inhibitor HA1004 affected collagen degradation by corneal fibroblasts. CONCLUSION: These results demonstrate that PKC plays an important role in collagen degradation by corneal fibroblasts in three-dimensional type I collagen gels, whereas PKA does not appear to participate in this process. PMID- 12131036 TI - A portable microkeratome-based anterior corneal surface harvesting device. AB - PURPOSE: To determine the reproducibility of anterior sclerokeratectomy using a portable nonelectric microkeratome-based device capable of harvesting the entire anterior corneal surface for lamellar transplantation. METHODS: A modified gas turbine-driven microkeratome (LSK One, Moria/Microtech, Doylestown, PA) with a redesigned head large enough to incorporate the whole human anterior corneal surface in a pass and was coupled to a manual vacuum pump. This instrument was tested on 25 fresh porcine globes divided into 2 groups (170-microm and 200 microm head). To assess cut reproducibility the physical dimensions (diameter and thickness) of the obtained lenticules were measured. RESULTS: The obtained lenticules were fairly circular (horizontal versus vertical diameters, p >0.2), with average diameters of 12.85 +/- 0.52 mm and 13.25 +/- 1.15 mm for the 170 and 200-microm heads, respectively. The average central lenticule thickness was 176.92 +/- 34.68 microm and 166.00 +/- 53.74 microm for the 170 and 200-microm heads, respectively. CONCLUSION: This new system presents an economical and portable alternative to electric-powered systems. In addition to being used by surgeons in the operating room, eye bank technicians in the field could theoretically use this system; including in developing countries where cost, availability of electricity, and portability are issues. PMID- 12131037 TI - Effect of locally administered anti-CD154 (CD40 ligand) monoclonal antibody on survival of allogeneic corneal transplants. AB - PURPOSE: To determine the effect of ocular administration of anti-CD154 monoclonal antibody on the survival of orthotopic murine corneal transplants. METHODS: BALB/c mice were used as recipients of multiple minor H- and MHC mismatched orthotopic corneal transplants. Recipient beds were either avascular (normal-risk) or neovascularized (high-risk) at the time of surgery. Mice were randomized to receive either anti-CD154 antibody or control immunoglobulin by subconjunctival injection. All grafts were evaluated for signs of rejection by slitlamp biomicroscopy until week 20-24 with the therapy tapered and discontinued after week 8 and week 12, respectively. RESULTS: In normal-risk transplantation, the 8-week survival rate improved from 30% in control mice to 90% in anti-CD154 treated mice (p= 0.0061). In high-risk transplantation, the survival rate of anti CD154-treated mice was enhanced to 55% compared with 0% in control mice at week 8 (p= 0.0184); however, tapering and termination of anti-CD154 led to some loss in graft survival, with a survival rate of 56% in normal-risk recipients, and 22% in high-risk recipients by week 20. Anti-CD40L treated animals displayed lower grades of postoperative corneal neovascularization (p<0.05), in particular in normal-risk recipients. CONCLUSIONS: Local ocular administration of anti-CD154 is effective in the prevention of corneal allograft rejection in normal-risk recipients, and in delaying the incidence of rejection in high-risk recipients. Long-term graft survival may not be fully achieved following termination of the CD40-CD154 pathway blockade. PMID- 12131038 TI - Interlacing of collagen lamellae in the midstroma of the human cornea. AB - PURPOSE: To investigate by means of scanning electron microscopy (SEM) the interlacing of collagen lamellae in the midstroma of the human cornea after opening the interlamellar spaces. MATERIAL AND METHODS: For SEM, cells and noncollagenous extracellular matrix were removed with 10% sodium hydroxide. Specimens were dehydrated in a series of graded tertiary butanols, frozen at -24 degrees C and dried in an exsiccator by sublimation of the frozen butanol. Dried corneas were cut vertically with a razor blade, and the interlamellar spaces were exposed by stretching the stroma along its anterior-posterior axis by pulling apart the inner (endothelial) and outer (epithelial) edges. Specimens were sputtered with gold and examined with a Cambridge Stereoscan 90 microscope. RESULTS: The opened interlamellar spaces gave the stroma at the cutting edge the appearance of a polymorphic honeycomb. The stromal openings differed in size, from approximately 10 microm to over 150 microm length and up to 80 microm in height. Adjacent lamellae remained connected at several interconnecting regions, either through an exchange of short merging sublamella or single fibrils. Interlacing lamellae crossed through fissures between the branches of splitting lamellae. Others crossed clefts of neighboring lamellae, and other lamellae tunneled crosswise through a horizontally split lamellae hanging in the inferior branch as in a hammock. Large interweaving zones in which a mixture of several types of interlacing was localized close together could also be found. CONCLUSION: The current study indicates that interlacing is a distinct and important feature of the human cornea, and it gives new insights into the stromal morphology by demonstrating various types of interlacing that occur between collagen lamellae. PMID- 12131039 TI - Burkholderia gladioli keratitis associated with consecutive recurrent endophthalmitis. AB - PURPOSE: To report a case of Burkholderia gladioli keratitis with consecutive endophthalmitis. METHODS: Case report and literature review. RESULTS: A 76-year old man with a history of diabetes mellitus developed bacterial keratitis and consecutive endophthalmitis in the corneal graft of the left eye. Corneal, aqueous, and vitreous cultures yielded Burkholderia gladioli. Emergent keratoplasty, pars plana vitrectomy, and injection of intravitreal antibiotics led to resolution of the infection and improved vision. Four months later, the patient developed recurrent Burkholderia keratitis and endophthalmitis, necessitating a total keratoplasty and repeat injection of intravitreal antibiotics. CONCLUSION: This is the first report, to our knowledge, of ocular Burkholderia gladioli infection, an uncommon aerobic, gram-negative rod, recently subclassified from the genus Pseudomonas based on DNA-rRNA homology studies. PMID- 12131040 TI - Late-onset traumatic flap dislocation and diffuse lamellar inflammation after laser in situ keratomileusis. AB - PURPOSE: To report a case of traumatic flap partial dislocation and subsequent diffuse lamellar inflammation 14 months after laser in situ keratomileusis (LASIK) retreatment. METHODS: Case report of a late flap dislocation that occurred during routine recreational activity (struck with a finger in the right eye while playing basketball). RESULTS: The partially dislocated LASIK flap was reflected nasally, and the stromal surfaces of the flap and bed were thoroughly scraped to remove debris and epithelial cells. The flap was repositioned, and a bandage contact lens was placed. Diffuse lamellar inflammation, which developed on post-trauma day number two, was successfully treated with frequent topical steroids. Three weeks after the injury, the patient had regained 20/20 uncorrected visual acuity. CONCLUSIONS: Patients should be appropriately warned of the possibility of late flap dislocation with traumatic forces encountered during routine recreational activities. Full visual recovery is possible if the dislocation is promptly diagnosed and appropriately managed. PMID- 12131041 TI - Keratopathy from congenital syphilis. AB - PURPOSE: Therapeutic decision-making in syphilitic corneal disease is discussed after seeing a patient with keratopathy from congenital syphilis. METHODS: Case report and review of the literature. RESULTS: Laboratory evaluation of our patient revealed a nonreactive serum rapid plasma reagin (RPR) test and a positive microhemagglutination assay-Treponema pallidum (MHA-TP) test. CONCLUSION: Any patient with residual interstitial keratopathy, especially if bilateral, should be suspected to have congenital syphilis if the patient or parents were ever treated for a sexually transmitted disease, if stigmata of congenital syphilis are present, or if childhood ocular inflammation occurred. PMID- 12131042 TI - Detection of measles virus genomic RNA in tear samples from a patient with measles keratitis. AB - PURPOSE: To report a case of measles keratitis, in which measles virus genomic RNA was detected in tear samples. METHODS: Samples of tears and pharyngeal mucus were obtained from a 19-year-old man with active systemic measles on day 7 after illness onset. These specimens were then subjected to reverse-transcriptase polymerase chain reaction (RT-PCR), using measles-specific primers. RESULTS: One week after systemic measles outbreak, the patient presented with a chief complaint of irritation in both eyes. Slit-lamp examination revealed bilateral superficial punctate keratopathy in the central cornea with small diffuse subepithelial infiltrations, and RT-PCR detected measles virus genomic RNA in tears and pharyngeal mucus. The patient was treated with topical 0.1% fluorometholone and 0.5% levofloxacin three times a day for 1 week, and the keratopathy resolved without sequelae. CONCLUSION: This report demonstrates for the first time the presence of measles virus genomic RNA in the tears of a patient with measles keratitis. It also highlights the utility of RT-PCR for rapid detection of measles RNA in tear or mucus samples. Furthermore, the presence of virus in bodily fluids and secretions signifies the potential contagiousness of this type of keratopathy. PMID- 12131043 TI - Danger of systemic cyclosporine for corneal graft. AB - PURPOSE: To report a case of posttransplant lymphoproliferative disorder (PTLD) in a patient receiving oral cyclosporine (CS) for immunosuppression in a high risk keratoplasty. METHODS: A systemic CS therapy was given to prevent graft rejection of a keratoplasty. Risk rejection was high in regard to a previous graft rejection and persistence of a corneal vascularization. One year after the keratoplasty, the patient developed a gastric Epstein-Barr virus (EBV)-induced B cell lymphoma. The outcome was favorable after chemotherapy. CONCLUSIONS: This unique case of lymphoma (PTLD) in the course of corneal graft management questions the indications and the follow-up of patients with CS therapy and raises the issue of topical CS treatment. PMID- 12131044 TI - Intralamellar autopatch with lamellar keratoplasty for paracentral corneal perforations. AB - PURPOSE: To report a new technique of tectonic intralamellar autopatch with lamellar keratoplasty (LK) for paracentral corneal perforations. METHODS: A partial thickness lamellar dissection of the host was undertaken to remove the superficial epithelium and the anterior stroma. A second lamellar dissection began at the periphery to approximately 1 mm away from the perforation to fashion a hinge. The hinged autolamellar flap was reflected over the perforation and sutured to the host with interrupted sutures. Finally, the autolamellar patch graft was supported by a lamellar graft, which was secured in place with 16 interrupted sutures. This technique was undertaken in 4 eyes with paracentral perforation after trauma (2 eyes), after pterygium (1 eye), and inadvertent perforation during host bed dissection of large LK (1 eye). RESULTS: All eyes achieved a stable ocular surface and a postoperative visual acuity of more than 6/60. CONCLUSIONS: Intralamellar autopatch with lamellar keratoplasty provides adequate tectonic support in cases of paracentral corneal perforation and thereby maintaining the integrity of the globe. It also provides ambulatory visual acuity. PMID- 12131045 TI - Confocal microscopy in multiple myeloma crystalline keratopathy. AB - PURPOSE: To report confocal microscopy findings of a patient with multiple myeloma crystalline keratopathy and the response to treatment. METHODS: Confocal microscopy images of the cornea were taken OU and the corneal crystals analyzed using the Cell Counter software. RESULTS: Numerous hyperreflective globules 6-11 nm in size were located within the corneal epithelium and anterior stroma. These crystals obscured normal architectural detail of the cornea. After 6 months of chemotherapy, confocal microscopy was repeated and demonstrated decrease in the size and number of hyperreflective globules. CONCLUSION: Confocal microscopy can enable the clinician to monitor the clinical response of multiple myeloma crystalline keratopathy to chemotherapeutic agents. PMID- 12131046 TI - Marginal keratitis associated with administration of filgrastim and sargramostim in a healthy peripheral blood progenitor cell donor. AB - PURPOSE: To describe a 61-year-old healthy peripheral blood progenitor cell (PBPC) donor who developed marginal keratitis and mild uveitis on the third day after receiving daily recombinant human granulocyte colony-stimulating factor (rhG-CSF; filgrastim) and recombinant human granulocyte-macrophage colony stimulating factor (rhGM-CSF; sargramostim) to mobilize PBPCs for allogeneic transplantation. METHODS: Interventional case report. RESULTS: The keratitis was treated with topical administration of 1% prednisolone acetate solution and resolved within 24 hours. The topical steroid dose was tapered and ultimately discontinued without recurrence of keratitis. CONCLUSION: Healthy PBPC donors receiving rhG-CSF or rhGM-CSF should be monitored for ocular complications, particularly marginal keratitis and uveitis. PMID- 12131047 TI - Conjunctival ulceration in recurrent Wegener granulomatosis. AB - PURPOSE: To describe a patient with Wegener granulomatosis (WG) previously in remission who developed conjunctival ulceration as the first sign of disease recurrence. METHODS: Case report and review of the literature. RESULTS: Twenty one years after WG was originally diagnosed and with the disease thought to be in remission, a 52-year-old man with complaints of ocular irritation for the previous year was found to have multiple palpebral conjunctival ulcerations of the left eye. Incisional biopsy revealed mixed inflammation consistent with WG. Within 3 months of recognition of his conjunctival ulcers, newly recurrent pulmonary inflammation developed and serologies for cytoplasmic-pattern antineutrophil cytoplasmic antibodies (C-ANCA) became positive. CONCLUSION: Conjunctival ulceration is a rare manifestation of WG but may presage more widespread disease. Mucosal ulceration in a patient with a previous diagnosis of WG should stimulate an aggressive search for renewed systemic disease activity. PMID- 12131049 TI - Co-infection of the human cornea with Stenotrophomonas maltophilia and Aspergillus fumigatus. AB - PURPOSE: To report a case of corneal co-infection of Stenotrophomonas maltophilia and Aspergillus fumigatus. METHODS: We describe a culture and biopsy proven infectious keratitis with a large, brown, round anterior chamber mass attached to the endothelium. RESULTS: Stenotrophomonas maltophilia was cultured from external scrapings of a corneal ulcer and septate hyphae were stained with Gomori's methenamine silver(GMS) stain along the wall of the excised intracameral mass. Aspergillus fumigatus was cultured from the mass and pus pockets developed along the corneoscleral incision for removal of the mass. CONCLUSION: Co-infection of cornea with Stenotrophomonas maltophilia and Aspergillus fumigatus with existence of a large, brown, smooth-surfaced mass in the anterior chamber makes this case unique and interesting. PMID- 12131048 TI - Positive polymerase chain reaction and histology with borderline serology in Parinaud's oculoglandular syndrome. AB - PURPOSE: To report a case of Parinaud's oculoglandular syndrome (POS) in which, despite a borderline serology, polymerase chain reaction (PCR) testing for a conjunctival biopsy was positive for Bartonella henselae, a source of cat-scratch disease. A Steiner silver stain demonstrated the organism. METHODS: Case Report. RESULTS: A 65-year-old man was seen for a foreign body in his left eye (OS) associated with chemosis and a preauricular node. CONCLUSION: B. henselae is a known cause of POS. This gram-negative pleomorphic rod has been more frequently discovered in connection with this syndrome due to improved diagnostic testing such as indirect immunofluorescence antibody and PCR testing. Frequently, serology is positive if the organism is present. This report describes a patient with clinical findings of POS who, despite borderline serology, had pleomorphic rods on Steiner silver stain and positive PCR testing compatible with Bartonella henselae. PMID- 12131050 TI - Amniotic membrane, tear film, corneal and aqueous levels of ofloxacin in rabbit eyes after amniotic membrane transplantation. PMID- 12131051 TI - Spontaneous expulsive suprachoroidal hemorrhage. PMID- 12131052 TI - Drugs, sport, and medical practice. PMID- 12131053 TI - Sports physicians and the doping crisis in elite sport. AB - The participation of sports physicians in the "doping" of athletes with banned drugs can be documented as far back as the 1890s. Concern about the ethics and safety of doping elite athletes appeared during the 1920s and 1930s as sport became an increasingly important form of popular culture. While organized medicine has opposed doping as a matter of policy at least since the 1950s, sports physicians have never adequately confronted the conflicts of interest that arise when they choose to work with elite athletes whose first priority is performance rather than with healing in the traditional sense. Confronted with the demands of their athlete-clients, sports physicians have divided into two factions regarding the wisdom and propriety of administering doping drugs to athletes. While most physicians are, in all likelihood, unwilling to violate laws, regulations, and medical standards by doping athletes, a significant minority of doctors has used one or more arguments to justify doping athletes: drugs are necessary to compete effectively; athletes should be free to medicate themselves as they please; drugs do not differ essentially from other performance enhancing techniques or equipment; and medically supervised doping is safer than self-medication by athletes. Physicians can also rationalize doping as an occupational requirement of some professional athletes. In summary, physicians have played a significant, and largely unacknowledged, role in the doping of many elite athletes over the past 50 years. PMID- 12131054 TI - Informed decision-making on sympathomimetic use in sport and health. AB - The International Olympic Committee, the World Anti-Doping Agency, and International Sport Federations have banned and restricted the use of many stimulants including prescription and over-the-counter medications and dietary supplements. In addition to elite athletes, people of all ages use stimulants in attempts to improve athletic performance, alter body composition, and increase levels of energy. Here we introduce a seven-stage model designed to facilitate informed decision-making by individuals taking or thinking of taking stimulants for sport, health, and/or appearance reasons. We review for amphetamines, over the counter sympathomimetics, and caffeine their performance-enhancing and performance-degrading effects, health benefits and mechanisms of action, medical side effects, and legal, ethical, safety, and financial implications. PMID- 12131055 TI - The asthmatic athlete, inhaled beta agonists, and performance. AB - INTRODUCTION: The large increase in the number of athletes who apply to use inhaled beta agonists (IBAs) at the Olympic Games is a concern to the medical community. This review will examine the use of IBAs in the asthmatic athlete, the variability that exists between countries and sport, and outline a plan to justify the use of these medications. DATA SOURCES: Much of this article is a result of an International Olympic Committee (IOC) Medical Commission-sponsored meeting that took place in May 2001. Records of the use of IBAs at previous Olympics were reviewed. MEDLINE Searches (PubMed interface) were performed using key words to locate published work relating to asthma, elite athletes, performance, treatment, and ergogenic aids. MAIN RESULTS: Since 1984 there have been significant increases in the use of IBAs at the Olympic Games as well as marked geographical differences in the percentage of athletes requesting the use of IBAs. There are large differences in the incidence of IBA use between sports with a trend towards increased use in endurance sports. There are no ergogenic effects of any IOC-approved IBA given in a therapeutic dose. CONCLUSIONS: In many cases, the prescription of IBAs to this population has been made on empirical grounds. Beginning with the 2002 Winter Games, athletes will be required to submit to the IOC Medical Commission clinical and laboratory evidence that justifies the use of this medication. The eucapnic voluntary hyperpnea test will be used to assess individuals who have not satisfied an independent medical panel of the need to use an IBA. PMID- 12131056 TI - Strategies for rhEPO detection in sport. AB - This article examines available strategies for the detection of recombinant erythropoietin (rhEPO) abuse in sport. RhEPO was quickly recognized as an effective but hazardous performance-enhancing agent. In the absence of a valid procedure to detect rhEPO doping, at-competition health checks were introduced, which excluded athletes from competition when their hemoglobin or hematocrit values exceeded an arbitrary limit. This limited the danger to athletes, but did nothing to eliminate the use of rhEPO. Through the last decade, both direct and indirect methods for detecting rhEPO were investigated. No single indirect marker was found that satisfactorily demonstrated rhEPO use. A combination of blood and urine tests together formed the procedure and strategy approved by the International Olympic Committee (IOC) for detecting rhEPO use at the Sydney Olympics. However strategies for testing for EPO are as important as the developed laboratory analytical procedures. The use of extensive out-of competition testing and analysis within the IOC accredited laboratory system is critical to any testing program. At-competition blood tests have merit as true health checks and will also be needed to detect acutely useful agents such as hemoglobin-based oxygen carriers. However the persistence of the "health check" rationale for on-site at-competition rhEPO testing has led to much wasted testing effort, as rhEPO use by athletes will rarely occur near to or at the time of the competition for fear of detection. Thus, direct testing methods (such as the rhEPO urine test) especially will fail due to the completed metabolism and elimination of administered rhEPO before the test, unless the international sporting federations use the information gathered to assist in targeted out-of competition testing. This article discusses the limitations of testing at competition and proposed strategies for dealing with various phases of EPO doping in detail, concluding that no one single currently used strategy will detect all users of rhEPO. The development of strategies to diagnose rhEPO abuse may serve as a model to detect other biological agents. PMID- 12131057 TI - DHEA and sport. AB - Dehydroepiandrosterone (DHEA), a 19-carbon steroid, is situated along the steroid metabolic pathway. It is the most abundant circulating hormone in the body and can be converted to either androgens or estrogens. It is readily conjugated to its sulphate ester DHEAS, and they are designated as DHEA(S) here when used together. Its secretion reaches a peak in early adulthood and thereafter decreases, until approximately age 70 years when it reaches a concentration of approximately 20%. Many hormonal changes may take place with aging but none is as marked as this. This "relative DHEA deficiency" resulted in DHEA being enthusiastically labelled by some as a fountain of youth or an antidote to aging that would prove to be the panacea they are seeking. Its use was also taken up enthusiastically by the athletic community and used as a prohormone in the belief or hope that it would be converted mainly to testosterone in the body. PMID- 12131058 TI - Beyond EPO. AB - The objective of this review is to examine newer compounds coming on the market capable of boosting red blood cell concentration and thus improving aerobic activities in particular. Erythropoietin (EPO) has been used extensively in the past by the athletic community in this role. Its main disadvantages are the side effects due to increased viscosity, and the recent development of a blood test for drug testing methods. Two new methods of increasing red blood cell concentration for use in medicine are hemoglobin oxygen carriers and perfluorocarbons, each having a different structure, but which allow oxygen to be delivered to the tissues. Hemoglobin oxygen carriers physically alter the hemoglobin molecule by several methods so that complications such as renal toxicity are obviated. Perfluorocarbons belong to a group of synthetic compounds containing hydrocarbons to which fluorine is added. Evidence is available to suggest that athletes are already adapting these newer molecules for their benefit. PMID- 12131059 TI - Nutritional supplements and doping. AB - CONTEXT: The problems of doping in sport and the increasing use of nutritional supplements by athletes are issues that intersect to the degree that a large number of supplements may contain substances that are banned in sport. Many supplements contain substances that are associated with significant health hazards. Athletes consuming such supplement products may jeopardize their sporting status, and their health. OBJECTIVES: To clarify and summarize the current status of dietary supplements in general, and to describe specific problems that can be associated with supplement use so that sport physicians might be better prepared to address these issues with their athlete-patients. DATA SOURCE: An analysis of recent and relevant literature accessed through MEDLINE, and interactions with clinicians, laboratory scientists, colleagues, and athletes. CONCLUSIONS: The dietary supplement industry is completely unregulated in the United States; as a consequence, an abundance of supplement products of dubious value, content, and quality are now available around the world. It is known that many supplement products contain substances that are prohibited in sport-typically stimulants or anabolic steroid precursors. Many supplements contain substances (e.g., ephedrine) that have been associated with significant morbidity and mortality. Sport practitioners have particular responsibilities in addressing this issue. Athletes need to be aware of the problems that can follow supplement use, and sport authorities need to ensure that nutritional education and guidance for athletes is of the highest standard. The need for the appropriate regulation of dietary supplements is emphasized. PMID- 12131060 TI - Does exogenous growth hormone improve athletic performance? AB - OBJECTIVE: To conduct a critical appraisal of the literature to address the question of whether human growth hormone (HGH) improves performance in trained athletes. DATA SOURCES: Used PubMed using the search terms of "growth hormone athletes" and the reference lists of previous reviews of the subject. STUDY SELECTION: Randomized double-blind placebo-controlled study of exogenous HGH on muscle power in trained athletes. Only one study matched the search criteria. CONCLUSION: There is no evidence of increased muscle strength with HGH in trained athletes. PMID- 12131061 TI - The war on drugs in sport: a perspective from the front-line. AB - CONTEXT: Recent international developments have served to solidify the international approach to doping in sport. The development of the World Anti Doping Agency (WADA) has resulted in new, coordinated efforts to address this important sport issue. An array of new efforts and initiatives has been initiated by the new agency. The Sydney and Salt Lake City Olympics were characterized by intensive efforts to minimize doping. The antidoping environment is evolving rapidly, and several profoundly important developments will take place in the immediate future. OBJECTIVES: To outline the challenges, opportunities, and changing circumstances of the current antidoping environment so that sport medicine practitioners might understand the context in which a variety of new initiatives and approaches will develop. At the same time, to ensure that practitioners understand the importance of appropriately developed and delivered antidoping policies, programs, and procedures, and the need for their harmonization. To ensure that sport medicine practitioners appreciate the need for a comprehensive approach to doping control, i.e., programs that include much more than drug testing. DATA SOURCE: A review of relevant policy documents derived from a variety of sport and antidoping organizations; selected references drawn from MEDLINE; and materials prepared by colleagues drawn from the international antidoping community. CONCLUSIONS: The increased global effort to address doping is welcome. It will require that several critical issues be addressed that will test the resolve of all involved. PMID- 12131062 TI - Complex regional pain syndrome type I: use of the International Association for the Study of Pain diagnostic criteria defined in 1994. AB - OBJECTIVES: The objective was to assess the reported use in recent publications of the diagnostic criteria for complex regional pain syndrome type I (CRPS I) proposed by the International Association for the Study of Pain (IASP) in 1994. METHODS: A literature search of MEDLINE (January 1996 to July 2000) was performed with use of the medical subject heading "reflex sympathetic dystrophy" and the free texts words "complex," "regional," "pain," and "syndrome." Publications in English, German, and Dutch were analyzed. From the search, 65 original publications were selected. Another 27 publications (referenced publications) that were referenced in the 65 original publications for the description of diagnostic criteria for CRPS I also were included. A standard form was used to assess a total of 92 publications. A sensitivity analysis was performed by means of analyzing three scenarios in which the diagnostic criteria were used as proposed and two combinations of less stringent criteria. RESULTS: Use of the diagnostic criterion pain was reported in 35 (38%) of the analyzed publications. None of the original publications satisfied the proposed IASP diagnostic criteria. Four (15%) of the referenced publications satisfied the proposed IASP diagnostic criteria. Ten (15%) of the original publications referred correctly to the referenced publications. With the less strict criteria used in scenarios 2 and 3, 2 (3%) and 3 (5%), respectively, of the original publications fulfilled these criteria. CONCLUSIONS: If the diagnostic criteria for CRPS I are not used uniformly, the populations in clinical studies may not be uniform either. Whether different authors are describing the same syndrome and whether their findings can be compared is open to question. On the basis of the results of this study, it is concluded that the IASP criteria for CRPS I are poorly used in clinical studies. PMID- 12131063 TI - Defining the therapeutic role of local anesthetic sympathetic blockade in complex regional pain syndrome: a narrative and systematic review. AB - OBJECTIVE: There is growing controversy on the value of blocking the sympathetic nervous system for the treatment of complex regional pain syndromes (CRPS). The authors sought to evaluate the efficacy of sympathetic blockade with local anesthetic in these syndromes. In addition, they performed a comprehensive review of the pathophysiology and other treatments for CRPS. DESIGN: Systematic review of the literature was performed. MEDLINE was searched from 1966 through 1999. The authors identified only three randomized controlled trials (RCTs) that evaluated sympathetic blockade with local anesthetic, but because of differences in study design they were unable to pool the study data. The authors therefore included nonrandomized studies and case series. INTERVENTIONS: Studies were included if local anesthetic sympathetic blockade was used in at least 10 patients. Studies were excluded if continuous infusion techniques, somatic nerve blocks, or combined sympatholytic therapies were evaluated. OUTCOME MEASURES: Pain relief was classified as full, partial, or absent. The lack of a comparison group in the studies allowed only the calculation of distribution of the response categories, and the sum of the pooled rates does not equal 100%. RESULTS: Twenty-nine studies were included that evaluated 1,144 patients. Nineteen studies were retrospective, 5 prospective case series, 3 RCTs, and 2 nonrandomized controlled studies. The quality of the publications was generally poor. Twenty-nine percent of patients had full response, 41% had partial response, and 32% had absent response. It was not possible to estimate the duration of pain relief. CONCLUSIONS: This review raises questions as to the efficacy of local anesthetic sympathetic blockade as treatment of CRPS. Its efficacy is based mainly on case series. Less than one third of patients obtained full pain relief. The absence of control groups in case series leads to an overestimation of the treatment response that can explain the findings. PMID- 12131064 TI - Comparison of multiple against single pain intensity measurements in complex regional pain syndrome type I: analysis of 54 patients. AB - OBJECTIVE: To describe the comparison of multiple and single pain ratings in patients with complex regional pain syndrome type I (CRPS I). DESIGN: Correlation, agreement, and reliability analyses were performed between the average pain intensity measured 3 times a day over a course of 4 days and one single pain rating (designated the "recalled average" pain, as assessed by the patient) before treatment and at 1-, 3-, and 6-month periods after treatment. PATIENTS: The patient population consisted of 54 patients with CRPS I in a randomized trial. RESULTS: The results show that both measurements correlate and have excellent agreement. Furthermore, both ratings measure significant pain reduction after treatment; "recalled average" pain, however, reflects greater change in pain intensity. CONCLUSIONS: In patients with CRPS I a single pain rating is an accurate predictor of the average pain measured by a multiple pain rating test. Moreover, both assessments are accurate enough to determine changes in pain over time with an effective treatment. PMID- 12131065 TI - Prediction of outcome in whiplash-associated disorders using West Haven-Yale Multidimensional Pain Inventory. AB - OBJECTIVE: To investigate the predictive capacity of the West Haven-Yale Multidimensional Pain Inventory (MPI) with regard to prolonged pain, using car occupants who had sustained a neck sprain in a traffic accident. DESIGN: A prospective cohort study including a one-year follow-up. PATIENTS: One hundred thirty adults were examined by a specialized neck-injury team after a first visit to an accident and emergency department. The subjects answered the MPI questionnaire within one month of the accident. OUTCOME MEASURES: One year later, the patients answered a questionnaire about residual neck pain. The main outcome was determined by the question, "Do you have residual pain which you relate to the accident?" RESULTS: One hundred twenty-three (95%) of the subjects completed the study. Ninety-seven reported pain of some degree that they related to the accident. All but one of the MPI variables differed significantly between the group with residual pain and the group without pain. The variable interference had the strongest correlation with the outcome. Its discriminative capacity was 81% for those with pain and 94% for those without pain one year later. CONCLUSIONS: The MPI may be used at an early stage to identify patients who may develop chronic neck-pain after a traffic accident, at least in those who want a follow-up session after an initial visit to an accident and emergency department. PMID- 12131066 TI - The neck pain and disability scale: test-retest reliability and construct validity. AB - OBJECTIVE: This research established test-retest reliability and construct validity for the Neck Pain and Disability Scale (NPAD). METHODS: Two groups of patients with neck pain completed the NPAD. The first group filled out the scale twice before treatment, whereas the second completed it with a number of other outcome measures once a month for 4 months, for evaluation of treatment with injections. RESULTS: The reliability coefficient (r2 = 0.93) calculated from the data for the first group of patients indicated high test-retest reliability. Construct validity was demonstrated with the second group when the NPAD was compared with a number of other pain measures and found to have a larger treatment effect. The Neck Pain and Disability Scale factor scores also indicated that treatment effects varied across the four factors. CONCLUSIONS: The NPAD is a stable and responsive measure for patients with neck pain. The Neck Pain and Disability Scale factor scores are useful in identifying treatment effects on the specific dimensions involved in the pain experience. PMID- 12131067 TI - The treatment of fear of movement/(re)injury in chronic low back pain: further evidence on the effectiveness of exposure in vivo. AB - BACKGROUND AND OBJECTIVE: Several cognitive-behavioral factors contribute to the persistence of pain disability in patients with chronic back pain. Fear-avoidance beliefs and fear of movement/(re)injury in particular have been shown to be strong predictors of physical performance and pain disability. Patients reporting substantial pain-related fear might benefit from exposure in vivo to a set of individually tailored, fear-eliciting, and hierarchically ordered physical movements rather than more general graded activity. PATIENTS AND INTERVENTIONS: Six consecutive patients with chronic low back pain who reported substantial fear of movement/(re)injury were included in the study. After a no-treatment baseline measurement period, the patients were randomly assigned to one of two interventions. In the first intervention, patients received exposure in vivo first, followed by graded activity. In the second intervention, the sequence of treatment modules was reversed. Before each treatment module, treatment credibility was assessed. Daily measures of pain-related fear, pain catastrophizing, and pain intensity were completed using visual analog scales. In addition, standardized measures of pain disability, pain-related fear, and pain vigilance were taken before and after each treatment module and at the 1-year follow-up. To obtain more objective data on actual activity levels, an ambulatory activity monitor was carried by the patients during 1 week before and after each treatment module. RESULTS: Time series analysis of the daily measures showed that improvements in pain-related fear and pain catastrophizing occurred only during the exposure in vivo and not during the graded activity, irrespective of the treatment order. Analysis of the pretreatment to post-treatment differences also revealed that decreases in pain-related fear also concurred with decreases in pain disability and pain vigilance and an increase in physical activity levels. All improvements remained at the 1-year follow-up. PMID- 12131068 TI - Pain in hospitalized pediatric patients: how are we doing? AB - OBJECTIVE: The purpose of this study was to provide a baseline description of the prevalence of pain and pain management strategies in a pediatric hospital and to compare the prevalence of pain in this hospital to that in published reports in the literature. METHODS: Two hundred thirty-seven children ranging in age from 10 days to 17 years and 223 parents participated in an 8-hour survey on 5 inpatient units. Information about pain intensity and pain affect was collected from the children older than 6 years of age and from parents of those who were younger at 4 2-hour intervals. Information about procedural pain was collected from children, parents, and health care professionals over this 8-hour period. The type and amount of analgesia were also noted. RESULTS: More than 20% of the children had clinically significant pain at each of the 2-hour intervals, and 7 had pain scores of 5/10 or greater for the majority of the study day. At least 50% of the children were found to be pain-free during the 4 intervals, and there was a high level of agreement between parents and children's pain-intensity ratings. One hundred fifty-seven children had medication ordered and 80 children had no analgesia ordered. There was no significant correlation between characteristics of the patients and amounts or types of medication given. No analgesia was administered via intramuscular or subcutaneous injection. DISCUSSION: Although these results are encouraging in that a significant portion of the children were pain-free during the study day, the number of children who had clinically significant pain was too high. The results of this study compare with others in that a significant number of children were inadequately treated for pain. Clinical implications are discussed. PMID- 12131069 TI - Peripheral blood mononuclear cell beta-endorphin concentration is decreased in chronic fatigue syndrome and fibromyalgia but not in depression: preliminary report. AB - OBJECTIVE: The aim of this study was to examine the possible role of the immune system in the pathophysiology of chronic fatigue syndrome and fibromyalgia syndrome and in the differential diagnosis of depression by investigating changes in peripheral blood mononuclear cell levels of beta-endorphin, an endogenous opioid known to be involved in regulation of the immune system function. DESIGN: Beta-endorphin concentrations were measured by radioimmunoassay in peripheral blood mononuclear cells from healthy controls (n = 8) and patients with chronic fatigue syndrome (n = 17), fibromyalgia syndrome (n = 5), or depression (n = 10). RESULTS: Beta-endorphin concentrations were significantly lower in patients with chronic fatigue syndrome or fibromyalgia syndrome than in normal subjects and depressed patients (p <0.001 and p <0.01, respectively). They were significantly higher in depressed patients than in controls (p <0.01). CONCLUSIONS: Evaluation of peripheral blood mononuclear cell beta-endorphin concentrations could represent a diagnostic tool for chronic fatigue syndrome and fibromyalgia and help with differential diagnosis of these syndromes versus depression. The results obtained are also consistent with the hypothesis that the immune system is activated in both chronic fatigue syndrome and fibromyalgia syndrome. PMID- 12131070 TI - Frequency of spontaneous fist clenching during routine blood pressure measurements and its effect on measurement accuracy. AB - BACKGROUND: Guidelines for office blood pressure reading techniques do not warn us about the possibility that patients may clench their fist during cuff inflation. It is unknown how often patients do this, and what effect it has on measured blood pressure readings. DESIGN AND METHODS: We registered double blood pressure readings in 150 outpatient clinic subjects, who were not given specific instructions as to how to hold their hand during the procedure. If they clenched their fist during the first reading, they were asked to relax their hand during the second reading, and vice versa. Double readings with a relaxed hand on both occasions were registered in 100 matched control patients as well. RESULTS: Twenty-two of 150 patients (15%) spontaneously made a fist during the first reading. No systematic effect (lower or higher blood pressure) from making a fist was observed, but individual effects were often marked, as evidenced by the median absolute differences (regardless of '+' or '-' sign) between the duplicate readings: 5 (range 0-31) versus 3 (0-18) mmHg for systolic blood pressure, and 4 (0-22) versus 2 (0-16) mmHg for diastolic blood pressure in the study group versus the control group, respectively (P < or = 0.05). A different hypertension classification occurred twice as often when a fist was made during one of the two blood pressure readings than with two fistless readings (23 versus 12%, P = 0.04). CONCLUSION: About one out of every seven patients makes a fist during blood pressure taking. This can seriously affect measurement accuracy, and doubles the risk of misclassification of hypertension. PMID- 12131071 TI - Twenty-four-hour ambulatory blood pressure monitoring in atrial fibrillation. AB - BACKGROUND: Due to large beat-to-beat blood pressure variation the use of 24-h ambulatory blood pressure monitoring in patients with atrial fibrillation has been questioned. METHODS: Repeatability and variability of 24-h ambulatory blood pressure (Accutraccer II or Diasys Integra), and daily blood pressure variation was examined in 42 patients aged 51-81 (median 73.5) years admitted for elective electrocardioversion of atrial fibrillation. RESULTS: Before cardioversion 24-h ambulatory systolic blood pressure was slightly lower and nocturnal blood pressure reduction was larger in the group of patients who achieved sinus rhythm than in the group who maintained atrial fibrillation (11.5/10.5 versus 4.1/4.7 mmHg; P < 0.05). No statistically significant change was observed in ambulatory blood pressure after cardioversion in any of the two groups. Blood pressure variability (SD/mean) was 10-14% both in patients with and without conversion to sinus rhythm. Coefficient of repeatability (2 SD of difference) was 13.6 mmHg (16.6%) for diastolic blood pressure and 30.2 mmHg (24.7%) for systolic blood pressure in patients with normalized heart rhythm and 17.0 and 29.0 mmHg (21.5 and 22.4%) in patients with maintained atrial fibrillation, respectively. CONCLUSION: Ambulatory blood pressure monitoring provides data with similar variability and repeatability in patients with atrial fibrillation as in subjects with normal cardiac rhythm. Twenty-four-hour ambulatory blood pressure measurement is applicable in atrial fibrillation in the same way as during sinus rhythm. PMID- 12131072 TI - Effect of recorded home blood pressure measurements on the staging of hypertensive patients. AB - OBJECTIVE: To compare clinic and home blood pressures for use in classifying patients in relation to a recent guideline for the diagnosis of hypertension. METHODS: Fifty patients were studied and classified on the basis of clinic pressures, using the Joint National Committee VI criteria, into the categories of normal, high-normal and stage 1, 2 or 3 hypertension. The patients were given instructions for using the Omron IC home-recording device to take their blood pressure daily for 1 week and then return the units for data recall and entry. Average home-recorded pressures were calculated and patients reclassified in terms of the Joint National Committee VI criteria if their home pressures were higher or lower than their clinic pressures. RESULTS: According to the clinic results, 18% of the participants had normal blood pressure, 16% had high-normal pressure, 48% were hypertensive stage 1, 16% were hypertensive stage 2 and 2% were hypertensive stage 3. Reclassification by recorded home pressures occurred in 54% of the participants: 40% downwards and 14% upwards. Only 46% remained in the same category for both clinic and recorded home pressures. CONCLUSION: Recorded home blood pressure measurement provides an accurate, reliable and unbiased assessment. Using the Joint National Committee VI classification system for both clinic and recorded home blood pressures, the data on the home pressures led, in this sample, to a downward classification three times more frequently than an upward one. We therefore conclude that recording home blood pressure is a highly useful method for assigning the appropriate blood pressure classification when using the Joint National Committee VI guidelines. PMID- 12131073 TI - Patient-specific differences between blood pressure estimated from 24 h ambulatory measurements and serial office self-recordings. AB - The long-term true blood pressure, estimated in terms of the repeatable 24 h ambulatory mean (24 hAmb) pressure, is probably best related to progressive cardiovascular deterioration and is vital for clinical decision-making. Serial self-measurements (SELF) under quiet conditions may reflect this level, but with an annoying uncertainty in individual patients. This uncertainty is characterized by analyzing differences between the SELF readings of seated/supine patients on their own and the reference 24 hAmb values based on 72 recordings. From at-random replicated sessions of 59 subjects, the sources of uncertainty can be separated into three components: a systematic (non-random) variable difference in level from person to person, a mean systematic difference in level between methods, and random variability determining repeatability between sessions. An unstable alerting reaction distorts about six initial self-recordings and increases the random variability. The following 7-12 or 13-24 'steady-state' values show a comparable random variability but reflect the 24 hAmb level only when compared as the average of all patients. The standard deviation for repeatability is about 50 100% higher for SELF than for 24 hAmb. The patient-specific difference between methods contributes more than +/-10 mmHg to the uncertainty interval of the SELF results. These patient-specific differences recur at repeat sessions and thus cannot be reduced by averaging the results of sequential SELF sessions. In contrast, two 24 hAmb results provide a 95% uncertainty interval of +/-5 mm Hg. Thus, the averaged results of multiple SELF sessions can be used to detect major changes in blood pressure but are, as a result of patient-specific differences, too unreliable when the value to be estimated (the true value) is the expected long-term value of 24 hAmb results. PMID- 12131074 TI - Terminal digit preference and single-number preference in the Syst-Eur trial: influence of quality control. AB - BACKGROUND: Terminal digit and single-number preference may produce inaccuracy and biased results when measuring blood pressure. We describe these preferences in the Syst-Eur randomized placebo-controlled trial of the treatment of isolated systolic hypertension and describe how we sought to eliminate these problems. METHODS: The Data Monitoring Committee of the trial conducted yearly quality control meetings in Belgium and visited the participating centres to check their adherence to the protocol. These meetings involved identifying terminal digit preference, improving blood pressure control and boosting recruitment. RESULTS: The prevalence of use of terminal digit zero when measuring sitting systolic blood pressure (first readings) reduced from an average of 42.4% in the year prior to the date when a centre first randomized a patient to 31.5, 25, 22.3, 26.3, 23.2 and 22% in the subsequent 6 years. This trend was independent of the calendar year during which a centre entered the trial and supports the hypothesis that data-quality monitoring, including the feedback of digit preference to centres, led to a reduction in terminal digit zero preference. In addition, a higher than expected prevalence of the systolic blood pressure value of 148 mmHg was found in the active treatment groups in the double-blind phase. Selection for 148 mmHg persisted over time and constituted a single-number preference bias. This arose from the instruction to investigators to reduce systolic blood pressure to below 150 mmHg. CONCLUSION: Monitoring and feedback of data quality should be undertaken to minimize digit and number preference. Automatic devices should ideally be employed to help to avoid these problems as long as the devices are fully validated and regularly serviced, and providing that readings are not rejected and repeated. PMID- 12131075 TI - Comparison of Portapres non-invasive blood pressure measurement in the finger with intra-aortic pressure measurement during incremental bicycle exercise. AB - BACKGROUND AND DESIGN: In order to evaluate the accuracy of continuous, non invasive blood pressure measurements in the finger during stress, blood pressure was measured in six patients with a Portapres Model 2 during increasing levels of bicycle exercise, using simultaneously registered intra-aortic (aortic arch) pressure as a reference. METHODS: Blood pressure was measured continuously and non-invasively in the left middle finger using the Portapres Model 2 device, at the same time invasively using a pigtail catheter situated in the aortic arch at rest and during and after standardized bicycle exercise. At the end of each stress level, in accordance with the protocol of the Association for the Advancement of Medical Instrumentation, systolic and diastolic blood pressure and pulse values were determined from the non-invasive (Portapres Model 2) and invasive measurements for 15 s each, and the mean values and standard deviations were calculated. RESULTS: Compared with simultaneous invasive blood pressure measurement in the aortic arch, the difference in measured blood pressure value (the value determined by the device minus the value determined invasively) at rest fell within the authorized normal range for German and European blood pressure measurement devices: systolic +1.7 +/- 1.8 mmHg and diastolic -3.7 +/- 6.2 mmHg. The difference in measured value increased with the stress level. CONCLUSION: Blood pressure and pulse can be determined reliably and accurately using the Portapres Model 2, non-invasively, both at rest and at low-to-moderate levels of stress. PMID- 12131076 TI - Accuracy of the Welch Allyn Vital Signs Monitor 52000 automatic blood pressure measuring device according to a modified British Hypertension Society protocol. AB - The accuracy of the Welch Allyn Vital Signs Monitor, a compact device for the oscillometric measurement of blood pressure, was determined according to the British Hypertension Society protocol. The monitor achieved a grade C for diastolic and a grade D for systolic blood pressure. The device is suitable for monitoring a patient, for example post-operatively, in the emergency department or during an intervention. The device cannot, however, be recommended for an exact determination of blood pressure when compared with the mercury sphygmomanometer. In an earlier validation report, the Welch Allyn Vital Signs Monitor achieved a grade A for both diastolic and systolic blood pressure. After adjusting for the difference in method of calculating the grades used in the two studies, there remained a considerable difference in grading results, for which no clear reason could be found. PMID- 12131077 TI - Evaluation of the Tensioday ambulatory blood pressure monitor according to the protocols of the British Hypertension Society and the Association for the Advancement of Medical Instrumentation. AB - BACKGROUND: The validation of ambulatory blood pressure monitoring devices is necessary to obtain information on their accuracy. The objective of the present study was to evaluate the accuracy of the Tensioday oscillometric ambulatory blood pressure monitor according to the protocols of the British Hypertension Society and the Association for the Advancement of Medical Instrumentation (AAMI). DESIGN: We followed the phases recommended by the British Hypertension Society protocol: before-use calibration, in-use assessment, after-use calibration, static device validation and report of the evaluation. However, we expanded on the protocol to accommodate features required by the AAMI. METHOD: The accuracy of calibration of three Tensioday devices was tested before and after the in-use phase when each of three devices was performing 10 24 h sessions of ambulatory monitoring. As all three devices passed these phases, the accuracy of blood pressure measurement was tested in one arbitrarily selected device on 85 subjects for systolic and 85 for diastolic blood pressure values. This was done by comparing three sequential same-arm blood pressure readings obtained by the device with three readings obtained by two observers using standard mercury sphygmomanometer. The comparisons were carried out while resting in the seated, supine and standing positions for all subjects. The results were used to grade the performance of the device according to the British Hypertension Society protocol and to calculate the mean +/- standard deviation of the difference between the device and the observers, as required by the AAMI. RESULTS: The Tensioday device achieved an overall grade of A for both the systolic and diastolic measurements, and had a mean difference compared with the observer measured blood pressure of 1.4 +/- 5.3/1.0 +/- 4.7 mmHg, which satisfies the AAMI criteria for accuracy. The British Hypertension Society grading did not change when patients with low, medium, and high blood pressure were analysed separately. The AAMI accuracy criteria were fulfilled in the standing and lying positions as well. CONCLUSION: On the basis of these results, the Tensioday ambulatory blood pressure monitoring device can be recommended for clinical use for ambulatory monitoring. The accuracy of the device needs, however, further testing in special situations, such as in pregnancy, in elderly patients and during exercise. PMID- 12131078 TI - Therapeutic value of gastrografin in adhesive small bowel obstruction after unsuccessful conservative treatment: a prospective randomized trial. AB - OBJECTIVE: To assess the therapeutic value of Gastrografin in the management of adhesive small bowel obstruction after unsuccessful conservative treatment. SUMMARY BACKGROUND DATA: Gastrografin is a hyperosmolar water-soluble contrast medium. Besides its predictive value for the need for surgery, there is probably a therapeutic role of this contrast medium in adhesive small bowel obstruction. METHODS: Patients with clinical evidence of adhesive small bowel obstruction were given trial conservative treatment unless there was suspicion of strangulation. Those who responded in the initial 48 hours had conservative treatment continued. Patients showing no clinical and radiologic improvement in the initial 48 hours were randomized to undergo either Gastrografin meal and follow-through study or surgery. Contrast that appeared in the large bowel within 24 hours was regarded as a partial obstruction, and conservative treatment was continued. Patients in whom contrast failed to reach the large bowel within 24 hours were considered to have complete obstruction, and laparotomy was performed. For patients who had conservative treatment for more than 48 hours with or without Gastrografin, surgery was performed when there was no continuing improvement. RESULTS: One hundred twenty-four patients with a total of 139 episodes of adhesive obstruction were included. Three patients underwent surgery soon after admission for suspected bowel strangulation. Strangulating obstruction was confirmed in two patients. One hundred one obstructive episodes showed improvement in the initial 48 hours and conservative treatment was continued. Only one patient required surgical treatment subsequently after conservative treatment for 6 days. Thirty five patients showed no improvement within 48 hours. Nineteen patients were randomized to undergo Gastrografin meal and follow-through study and 16 patients to surgery. Gastrografin study revealed partial obstruction in 14 patients. Obstruction resolved subsequently in all of them after a mean of 41 hours. The other five patients underwent laparotomy because the contrast study showed complete obstruction. The use of Gastrografin significantly reduced the need for surgery by 74%. There was no complication that could be attributed to the use of Gastrografin. No strangulation of bowel occurred in either group. CONCLUSIONS: The use of Gastrografin in adhesive small bowel obstruction is safe and reduces the need for surgery when conservative treatment fails. PMID- 12131079 TI - Contrast radiography and intestinal obstruction. PMID- 12131080 TI - Bacillus anthracis as an agent of bioterrorism: a review emphasizing surgical treatment. AB - OBJECTIVE: To familiarize surgeons with the specific complications of cutaneous, gastrointestinal, inhalation, and systemic infection with Bacillus Anthracis, which may require surgical treatment. SUMMARY BACKGROUND DATA: The recent cases of intentional exposure to Bacillus Anthracis in the United States make familiarity with the basic microbiology, clinical manifestations, diagnosis, treatment, and control of this disease essential if mortality and morbidity is to be minimized, particularly following mass exposure. Although the treatment of Bacillus Anthracis infection is primarily medical, there are specific surgical complications with which the surgeon should be familiar. METHODS: A review of the literature was undertaken, utilizing electronic databases on infection with Bacillus Anthracis, as well as consultation with experts in this field. Emphasis was placed on the diagnosis and treatment of complications of infection that might require surgical intervention. RESULTS: Cutaneous anthrax infection results in eschar formation and massive soft tissue edema. When involving the extremities, increased compartment pressure requiring fasciotomy may result. Primary infection of the gastrointestinal tract may result in oropharyngeal edema and respiratory compromise requiring a surgical airway. Direct involvement of the lower gastrointestinal tract can result in intestinal ulceration, necrosis, bleeding, and perforation, which would require surgical exploration and resection of affected segments. Systemic sepsis, most often associated with inhalation anthrax, can cause massive ascites, electrolyte derangements, and profound shock requiring aggressive fluid resuscitation and careful hemodynamic monitoring and respiratory support. Systemic anthrax infection can also lead to gastrointestinal involvement by hematogenous dissemination, resulting in complications and requiring surgical management similar to direct gastrointestinal infection. CONCLUSIONS: Cutaneous, gastrointestinal, inhalation and systemic infection with Bacillus Anthracis can result in complications which would require familiarity with the pathogenesis and manifestations of this disease in order to recognize and treat promptly and successfully by surgical intervention. PMID- 12131081 TI - A meta-analysis on the efficacy of preoperative biliary drainage for tumors causing obstructive jaundice. AB - OBJECTIVE: To review the effectiveness of preoperative biliary drainage (PBD) in patients with obstructive jaundice resulting from tumors. SUMMARY BACKGROUND DATA: This was a systematic review, including a meta-analysis, of randomized controlled trials and comparative cohort studies conducted worldwide and published between 1966 and September 2001, classified on methodologic strength and subdivided into level 1 (randomized controlled trials) and level 2 (comparative cohort studies). METHODS: Comparison was made of PBD versus no PBD in jaundiced patients undergoing resection of a tumor. Outcome measures were in hospital death rate, overall complications resulting from the treatment modality (drainage- and surgery-related complications), and hospital stay. Effect sizes were calculated and combined in meta-analyses. Relative differences (%) were calculated to compare effects on outcome measures. RESULTS: Five randomized controlled studies comprising 302 patients met the inclusion criteria for level 1 studies, and 18 cohort studies comprising 2,853 patients met the criteria for level 2 studies. Meta-analysis of level 1 studies showed no difference in the overall death rate between patients who had PBD and those who had surgery without PBD. The overall complication rate, however, was significantly adversely affected by PBD compared with surgery without PBD. At level 2, there was no difference in the death rate between the two treatment modalities. The overall complication rate, however, was significantly adversely affected by PBD compared with surgery without PBD. If PBD had been without complications, then complications would be in favor of drainage based on level 1 studies, and no difference based on level 2 studies. Further, PBD was not able to reduce the length of postoperative hospital stay compared with surgery without PBD; instead, it prolonged the stay. CONCLUSIONS: This meta-analysis shows that PBD with current standards for patients with obstructive jaundice resulting from tumors carries no benefit and should not be performed routinely. The potential benefit of PBD in terms of postoperative rates of death and complications does not outweigh the disadvantage of the drainage procedure. Only if PBD-related complications could be reduced by 27% and consequently diminish hospital stay could PBD be beneficial. Further randomized controlled trials with improved PBD techniques are necessary. PMID- 12131082 TI - Ileocecal valve as substitute for the missing pyloric sphincter after partial distal gastrectomy. AB - OBJECTIVES: Accelerated gastric emptying (including dumping syndrome) occurs frequently after gastric resections, largely resulting from rapid entry of meal contents into the small intestine. The authors hypothesized that an ileocecal segment used as an interpositional graft placed between the remaining part of the stomach and the small intestine would slow down food transit and thus replace pyloric function. METHODS: Thirty Gottingen minipigs were randomized into three groups. Group 1: partial gastrectomy and Roux-en-Y reconstruction; Group 2: partial gastrectomy and ileocecal interpositional graft; and Group 3: sham laparotomy. Gastric emptying in the nonsedated animals was quantified using radioscintigraphy at 3 and 6 months postoperatively. The animals ingested 300 grams of soft food containing 99mTc labeled resin- pellets using a technique previously described. Data were analyzed using ANOVA. RESULTS: Three months postoperatively, the ileocecal group had a significantly prolonged gastric emptying time compared with the Roux-en-Y group, but gastric emptying time was also significantly faster compared to the control group (sham laparotomy). After 6 months no significant difference was seen between the ileocecal group and the controls, while emptying rates were still significantly faster in the Roux-en-Y group. CONCLUSIONS: Reconstruction of the gastric reservoir with an ileocecal segment largely restores gastric emptying patterns of food in minipigs. Six months postoperatively, gastric emptying time is similar to that of controls, and significantly slower when compared with the group with Roux-en-Y reconstruction. These results suggest that the ileocecal interposition graft could offer specific advantages over current reconstruction procedures. PMID- 12131083 TI - Circulating VEGF levels in the serum of gastric cancer patients: correlation with pathological variables, patient survival, and tumor surgery. AB - OBJECTIVE: To evaluate the clinical usefulness of serum vascular endothelial growth factor (VEGF) levels in gastric cancer patients. SUMMARY BACKGROUND DATA: Vascular endothelial growth factor plays an important role in the formation of new blood vessels involved in the growth and metastatic spread of solid tumors, but there is limited information regarding the clinical significance of serum VEGF levels in cancer patients. METHODS: Serum VEGF concentrations were measured by an enzyme linked immunosorbent assay in 61 healthy controls and in 58 gastric cancer patients before surgery, and then again at 7 and 30 days after surgery. The association between preoperative serum VEGF levels, clinicopathological features and patient survival, and their changes following surgery were evaluated. RESULTS: Serum VEGF levels in gastric cancer patients were significantly higher than those in controls. There was a significant association between serum VEGF levels and disease stage, as well as invasion depth of the tumor and the presence of distant metastases. Serum VEGF levels decreased significantly after radical resection of the primary tumor and increased in patients with unresectable tumors. Multivariate regression analysis showed that serum VEGF level is an independent prognostic factor for survival. CONCLUSIONS: Serum VEGF levels in gastric patients are significantly higher compared with normal controls and correlate with local tumor extent, disease stage, and the presence of distant metastases. Preoperative serum VEGF concentration decreases significantly after radical resection of the primary tumor and is an independent prognostic factor for patient survival suggesting that determination of serum VEGF levels may be clinically useful. PMID- 12131084 TI - Equivalent function, quality of life and pouch survival rates after ileal pouch anal anastomosis for indeterminate and ulcerative colitis. AB - OBJECTIVE: To compare the function, complications, and quality of life after ileal pouch-anal anastomosis (IPAA) for patients with indeterminate colitis (IndC) and ulcerative colitis (UC). SUMMARY BACKGROUND DATA: Reports on the outcome of IPAA for IndC have been inconclusive because of the small numbers available for analysis. Concerns about functional outcome, infectious perineal complications, pouch loss and the development of Crohn's disease remain, while there is no data on the quality of life after IPAA for IndC. METHODS: One thousand nine hundred and eleven patients undergoing IPAA for Ind and UC from 1983 to 1999 were evaluated. IndC was confirmed by repeat pathologic evaluation in 115 patients. Functional outcome and quality of life were assessed prospectively for all office visits (IndC = 230; UC = 5388) using previously reported systems. Complications were evaluated retrospectively. RESULTS: Functional results and the incidence of anastomotic complications and major pouch fistulae were the same in UC and IndC patients. Although IndC patients were more likely to develop minor perineal fistulae, pelvic abscess, and Crohn's disease, the rate of pouch failure was 3.4%, identical to that of UC patients. There was no clinically significant difference in quality of life, or satisfaction with IPAA surgery. Patients were equally happy to recommend surgery to IndC or UC patients, but 3% fewer IndC would undergo the same surgery again for their disease. CONCLUSIONS: While functional outcome, quality of life, and pouch survival rates are equivalent after IPAA for IndC and UC, there is an increase in some complications and the late diagnosis of Crohn's disease. Over 93% of IndC patients would undergo the same procedure again, and 98% would recommend IPAA to others with IndC. Patients with IndC should not be precluded from having IPAA surgery. PMID- 12131085 TI - Comparison of J-pouch and coloplasty pouch for low rectal cancers: a randomized, controlled trial investigating functional results and comparative anastomotic leak rates. AB - OBJECTIVE: To assess the efficacy of a novel coloplasty colonic pouch design in optimizing bowel function after ultralow anterior resection. SUMMARY BACKGROUND DATA: A colonic J-pouch may reduce excessive stool frequency and incontinence after anterior resection, but at the risk of evacuation problems. Experimental surgery on pigs has suggested that a coloplasty pouch (CP) may be a useful alternative. Although CP has recently been shown to be feasible in patients, there is no randomized controlled trial comparing bowel function with the J pouch. METHODS: After anterior resection for cancer, patients were allocated to either J-pouch or CP-anal anastomoses. Continence scoring, anorectal manometry, and endoanal ultrasound assessments were made before surgery. All complications were recorded, and these preoperative assessments were repeated at 4 months. The assessments were repeated again at 1 year, and a quality of life questionnaire was added. RESULTS: Eighty-eight patients were recruited from October 1998 to April 2000. Both groups were well matched for age, gender, staging, adjuvant therapy, and mean follow-up. There were no differences in the intraoperative time and hospital stay. CP resulted in more anastomotic leaks. At 4 months, J-pouch patients had 10.3% less stool fragmentation but poorer stool deferment and more nocturnal leakage. However, there were no differences in the bowel function, continence score, and quality of life at 1 year. There were no differences in the anorectal manometry and endoanal ultrasound findings. CONCLUSIONS: Coloplasty pouches resulted in more anastomotic leaks and minimal differences in bowel function. At present, the J-pouch remains the benchmark for routine clinical practice, and due care (including defunctioning stoma) should be exercised in situations requiring CP. PMID- 12131086 TI - Colonic postoperative inflammatory ileus in the rat. AB - OBJECTIVE: To investigate local inflammatory events within the colonic muscularis as a causative factor of postoperative ileus. SUMMARY BACKGROUND DATA: Surgically induced intestinal muscularis inflammation has been hypothesized as a mechanism for postoperative ileus. The colon is a crucial component for recovery of gastrointestinal motor function after surgery but remains unaddressed. The authors hypothesize that colonic manipulation initiates inflammatory events that directly mediate postoperative smooth muscle dysfunction. METHODS: Rats underwent colonic manipulation. In vivo transit and colonic motility was estimated using geometric center analysis and intraluminal pressure monitoring. Leukocyte extravasation was investigated in muscularis whole mounts. Mediator mRNA expression was determined by real-time reverse transcriptase-polymerase chain reaction. In vitro circular muscle contractility was assessed in a standard organ bath. The relevance of iNOS and COX-2 inhibition was determined using DFU or L NIL perfusion. RESULTS: Colonic manipulation resulted in a massive leukocyte recruitment and an increase in inflammatory mRNA expression. This inflammatory response was associated with an impairment of in vivo motor function and an inhibition of in vitro smooth muscle contractility (56%). L-NIL but not DFU significantly ameliorated smooth muscle dysfunction. CONCLUSIONS: The results provide evidence for a surgically initiated local inflammatory cascade within the colonic muscularis that mediates smooth muscle dysfunction, which contributes to postoperative ileus. PMID- 12131087 TI - Effects of hypothermia and rewarming on the mucosal villus microcirculation and survival after rat intestinal ischemia-reperfusion injury. AB - OBJECTIVE: To determine the effects of hypothermia and rewarming on changes in the villus microcirculation induced by intestinal ischemia-reperfusion (I/R). SUMMARY BACKGROUND DATA: The small intestine is extremely sensitive to I/R injury, and although hypothermia can reduce cellular injury, its capacity to influence the villous microcirculation after intestinal I/R is unclear, especially after the return to normothermic conditions. METHODS: Core body temperature of PVG rats was maintained at either 36 degrees to 38 degrees C (n = 12) or 30 degrees to 32 degrees C (n = 24) and then subjected to 30 minutes of intestinal ischemia. A subgroup of hypothermic animals (n = 12) were returned to normothermic conditions 120 minutes after clamp removal. The mucosal surface was visualized in an exteriorized ileal segment and macromolecular leak (MML) and leukocyte adhesion were monitored using in vivo microscopy (n = 6 in each group). MML from individual villi and numbers of adherent leukocytes within villi were determined for 2 to 4 hours after clamp removal. Heart rate and mean blood pressure were monitored in all animals. Control animals underwent sham surgery (n = 12). RESULTS: Ten of 12 normothermic animals failed to survive the reperfusion period, whereas all hypothermic animals and 11 of 12 of the hypothermic animals that were returned to normothermic conditions survived. MML was significantly increased in all animals subjected to I/R, although leakage was more marked in animals subjected to continuous normothermia. Enhanced leukocyte adhesion and decreased blood flow were observed only in normothermic animals. CONCLUSIONS: Hypothermia might prove to be an effective strategy for preventing adverse side effects in clinical settings in which intestinal I/R can be predicted. PMID- 12131088 TI - Long-term prognostic significance of extent of rectal cancer response to preoperative radiation and chemotherapy. AB - OBJECTIVE: To determine whether selected clinicopathologic factors, including the extent of pathologic response to preoperative radiation and chemotherapy (RT +/- chemo), have an impact on long-term recurrence-free survival (RFS) in patients with locally advanced primary rectal cancer after optimal multimodality therapy. SUMMARY BACKGROUND DATA: Although complete pathologic response to preoperative RT +/- chemo has been detected in up to 30% of rectal cancers, its significance on long-term outcome has not been widely reported. Previous retrospective studies evaluating clinical outcome in patients with complete or near-complete pathologic response documented good prognosis in this population but were limited by median follow-up in the range of 2 to 3 years. METHODS: Sixty-nine patients with locally advanced (T(3-4) and/or N1) primary rectal cancer were prospectively identified. All were treated at one institution with preoperative RT to the pelvis (at least 4,500 cGy). Forty patients received concurrent preoperative 5-fluorouracil-based chemotherapy and 27 received both pre- and postoperative chemotherapy. Patients underwent resection 4 to 7 weeks after completion of RT. TNM stage, angiolymphatic or perineural invasion, and extent of response to preoperative RT +/- chemo were determined by pathologic evaluation. Adverse pathologic features were defined as the presence of angiolymphatic and/or perineural invasion. RFS at 5 years was determined by the Kaplan-Meier method. RESULTS: With a median follow up of 69 months, 5-year RFS was 79%. RFS was significantly worse for patients with aggressive pathologic features and positive nodal status identified in the postirradiated surgical specimen. Risk ratios for RFS were 3.68 for the presence of aggressive pathologic features and 4.64 for node-positive rectal cancers. In patients with greater than 95% rectal cancer response to preoperative RT +/- chemo, only one patient has died as a consequence of cancer, another has died of an unrelated cause, and the remainder were free of disease with a minimum follow up of 47 months. CONCLUSIONS: These data suggest that a marked response to preoperative RT +/- chemo may be associated with good long-term outcome but was not predictive of RFS. The presence of poor histopathologic features and positive nodal status are the most important prognostic indicators after neoadjuvant therapy. PMID- 12131089 TI - Extrahepatic portal hypertension in chronic pancreatitis: an old problem revisited. AB - OBJECTIVE: To evaluate the impact of concomitant nonhepatic portal hypertension in chronic pancreatitis on immediate and long-term outcome after major pancreatic surgery. METHODS: A total of 154 patients (96 male, 58 female) with a history of pancreatitis of at least 12 months, severe incapacitating pain, and radiologic evidence of pancreatic head enlargement was evaluated. One hundred thirty-five patients underwent duodenum-preserving resections of the pancreatic head according to Beger or Frey, and 19 patients underwent pancreatoduodenectomy without (classical Whipple) or with pyloric preservation (PPPD) in cases of suspected malignancy. Outcome parameters included operative time and blood loss, early and late complications and death, recurrent pancreatitis, professional rehabilitation, and alterations in portal venous flow. Median follow-up in this prospective study was 51 months. RESULTS: Patients with portal hypertension required significantly more blood transfusions and had longer operative times than their counterparts. The overall postoperative complication rate was significantly higher in this subgroup. Restoration of postoperative portal venous blood flow was complete after Beger, Whipple, and PPPD procedures but was little affected by Frey procedures. There was no evidence of variceal hemorrhage during the observation period in all operative groups. CONCLUSIONS: Concomitant extrahepatic portal hypertension entails a substantial risk in pancreatic surgery for chronic pancreatitis. When surgery is considered in a symptomatic patient, surgical strategy is determined more by pancreatic morphology than by the intent to restore portal blood flow. PMID- 12131091 TI - Effect of truncal vagotomy on sphincter of oddi cyclic motility in conscious dogs. AB - OBJECTIVE: To evaluate the effects of truncal vagotomy at the diaphragmatic level on the sphincter of Oddi (SO) motility. SUMMARY BACKGROUND DATA: Cholelithiasis is a well-known late complication after gastrectomy and/or vagotomy. The mechanism of gallstone formation is only partly understood, and few studies address the effects of vagotomy on SO cyclic motility in conscious subjects. METHODS: In conscious dogs, SO motility was recorded by retrograde infusion manometry through a duodenal cannula before and after bilateral truncal vagotomy at the diaphragmatic level. Effects of cholecystokinin-octapeptide and feeding were also evaluated before and after vagotomy. RESULTS: SO cyclic motility and the gastroduodenal migrating motor complex continued to occur during postvagotomy fasting. Intermittent inhibitions of the SO and duodenal contractions disappeared during phase 3 of the migrating motor complex. SO basal pressure significantly decreased, whereas the amplitude significantly increased. Cholecystokinin octapeptide inhibited SO contractions before and after vagotomy. The amplitude of SO contractions increased and their frequency decreased after feeding; however, these effects disappeared after vagotomy. CONCLUSIONS: SO cyclic motility and the effects of feeding change after truncal vagotomy at the diaphragmatic level. These facts may at least partly explain gallstone formation after gastric surgery and/or vagotomy. PMID- 12131090 TI - Laparoscopic liver resection for malignant liver tumors: preliminary results of a multicenter European study. AB - OBJECTIVE: To assess the feasibility, safety, and outcome of laparoscopic liver resection for malignant liver tumors. SUMMARY BACKGROUND DATA: The precise role of laparoscopy in resection of liver malignancies (hepatocellular carcinoma [HCC] and liver metastases) remains controversial despite an increasing number of publications reporting laparoscopic resection of benign liver tumors. METHODS: A retrospective study was performed in 11 surgical centers in Europe regarding their experience with laparoscopic resection of liver malignancies. Detailed questionnaires were sent to each surgeon focusing on patient characteristics, clinical data, type and characteristics of the tumor, technical details of the operation, and early and late clinical outcome. All patients had radiologic investigations at follow-up to exclude disease recurrence. RESULTS: From February 1994 to December 2000, 37 patients with malignant liver tumors were included in this study. Ten patients had HCC, including 9 with cirrhotic liver, and 27 patients had liver metastases. The mean tumor size was 3.3 cm, and 89% of the tumors were located in the left lobe or in the anterior segments of the right liver. Liver procedures included 12 wedge resections, 9 segmentectomies, 14 bisegmentectomies (including 13 left lateral segmentectomies), and 2 major hepatectomies. The transfusion rate, the use of pedicular clamping, the conversion rate (13.5% in the whole series), and the complication rate were significantly greater in patients with HCC. There were no deaths. Postoperative complications occurred in eight patients (22%). The surgical margin was less than 1 cm in 30% of the patients. During a mean follow-up of 14 months, the 2-year disease-free survival was 44% for patients with HCC and 53% for patients having hepatic metastases from colorectal cancer. No port-site metastases were observed during follow-up. CONCLUSIONS: In patients with small malignant tumors, located in the left lateral segments or in the anterior segments of the right liver, laparoscopic resection is feasible and safe. The complication rate is low, except in patients with HCC on cirrhotic liver. By using laparoscopic ultrasound, a 1-cm free surgical margin should be routinely obtained. The late outcome needs to be evaluated in expert centers. PMID- 12131092 TI - Role of cyclase activating parathyroid hormone (1-84 PTH) measurements during parathyroid surgery: potential improvement of intraoperative PTH assay. AB - SUMMARY BACKGROUND DATA: Quick intraoperative parathyroid hormone assays are widely used as a guide to the adequacy of resection during parathyroid surgery. However, some authors have reported a 15% error rate of these assays because of the presence of false-positive and false-negative results. Recently the authors have found that most commercial intact PTH (iPTH) assays cross-react with non-(1 84) PTH (likely 7-84 PTH) and that the proportional levels of non-(1-84) PTH in patients were variable in a much wider range, accounting mostly for 20% to 60% of the immunoreactivity in samples obtained from hyperparathyroid patients. A cyclase activating PTH (CAP) measured by a novel immunoradiometric assay was shown to measure specifically 1-84 PTH. Using a CAP assay, the authors studied the rate of decline of CAP after parathyroidectomy and compared it with iPTH as measured by the Nichols intact PTH immunoradiometric assay. METHODS: This study comprised 29 patients with primary hyperparathyroidism (pHPT) caused by a single adenoma and 7 patients with secondary hyperparathyroidism (secondary HPT) who underwent parathyroidectomy. Blood samples were drawn after anesthesia, before excision of one enlarged parathyroid gland in pHPT and of the last gland in secondary HPT, and at 5, 10, and 15 minutes after excision. The 7-84 PTH level was calculated by subtracting the CAP value from the iPTH value. RESULTS: The percentage of 7-84 PTH in iPTH in plasma samples was 27.5 +/- 14.4% in pHPT and 39.6 +/- 15.1% in secondary HPT. In pHPT patients the plasma CAP and iPTH value decreased to 23.4 +/- 10.8 and 32.0 +/- 11.3% of the preexcision level at 5 minutes, 10.6 +/- 7.7 and 21.1 +/- 8.8% at 10 minutes, and 8.5 +/- 4.9 and 16.1 +/- 6.8% at 15 minutes after removal of the enlarged gland, respectively. At 5 minutes, CAP levels of all 29 pHPT patients had decreased to less than 40% of the preparathyroidectomy level; however, 7 (24%) patients still had an iPTH level of more than 40%. In secondary HPT patients, CAP and iPTH values had dropped to 43.3 +/- 20.2 and 66.1 +/- 19.7% at 5 minutes, 28.6 +/- 16.6 and 53.6 +/- 18.1% at 10 minutes, and 14.2 +/- 9.0 and 41.0 +/- 12.9% at 15 minutes after removal of the last enlarged gland, respectively. At 10 minutes, CAP levels of all seven secondary HPT patients had decreased to less than 50% of the preexcision level; however, three (43%) patients still had an iPTH level of more than 50%. In pHPT and secondary HPT, the 7-84 PTH level had dropped to 57.4 +/- 85.9 and 62.1 +/- 84.9%, respectively, of the preexcision value 15 minutes after removal of the enlarged gland or glands. CONCLUSIONS: The percentage of 7-84 PTH in iPTH in plasma samples varies substantially between patients with HPT. In both pHPT and secondary HPT, the plasma CAP value decreased more rapidly than iPTH after parathyroidectomy, depending on the amount of 7-84 PTH in circulation. These results suggest that the CAP assay may be a more useful adjunct to parathyroidectomy than the currently used iPTH assay. PMID- 12131094 TI - Donor health assessment after living-donor liver transplantation. AB - OBJECTIVE: To elicit donor opinions on liver living donation through use of a survey that protected the anonymity of the respondent and to assay long-term (follow-up > 1 year) donor health by a widely recognized instrument for health assessment. SUMMARY BACKGROUND DATA: Living-donor liver transplantation is an accepted technique for children that has recently been extended to adults. Limited donor outcomes data suggest favorable results, but no outcomes data have been reported using an instrument that elicits an anonymous response from the donor or employs a widely recognized health survey. METHODS: Forty-one living donors between June 1992 and June 1999 were identified and included in this study, regardless of specific donor or recipient outcome. Each donor received a 68-question survey and a standard McMaster Health Index. RESULTS: Survey response was 80%. All donors were satisfied with the information provided to them before donation. Eighty-eight percent of donors initially learned of living donation only after their child had been diagnosed with liver disease: 44% through the transplant center, 40% by popular media, 12% by their pediatrician, and 4% by their primary care physician. Physical symptoms, including pain and the surgical wound, were recurrent items of concern. Perception of time to "complete" recovery were less than 3 months (74%), 3 to 6 months (16%), and more than 6 months (10%). Donors' return to physical activities was shown by above-mean McMaster physical scores; scores for social and emotional health were not different from population data. There were no reported changes in sexual function or menstruation after donation, and five of six donors procreated. CONCLUSIONS: Donors overwhelmingly endorsed living donation regardless of recipient outcome or the occurrence of a complication. Eighty-nine percent advocated "increased" application of living donation beyond "emergency situations," and no donor responded that living donation should be abandoned or that he or she felt "forced" to donate. PMID- 12131093 TI - Appearance of monocyte chemoattractant protein 1 (MCP-1) early after thermal injury: role in the subsequent development of burn-associated type 2 T-cell responses. AB - OBJECTIVE: To determine whether monocyte chemoattractant protein-1 (MCP-1), which initiates subsequent development of burn-associated type 2 T cells, is produced in mice early after thermal injury. SUMMARY BACKGROUND DATA: A predominance of type 2 T-cell responses is commonly observed in animals and patients with severe thermal injuries. Burn-associated type 2 T cells have been identified as the cells responsible for the increased susceptibility of thermally injured mice to infections with herpes simplex virus type 1 and Candida albicans. Recently, the necessity of MCP-1 for the generation of type 2 T cells was shown in MCP-1 knockout mice. MCP-1 may have an important role in the increased susceptibility of thermally injured mice to various intracellular opportunistic pathogens. METHODS: The production of MCP-1 in sera or in cultures of various cells prepared from thermally injured mice was measured. Dual-chamber transwell cultures were performed to determine the influence of MCP-1-producing cells on the generation of burn-associated type 2 T cells. RESULTS: Without any stimulation, splenic macrophages from mice (1/2D-M(phi)) produced MCP-1 into their culture fluids 12 hours after thermal injury. Interleukin-4 was detected in culture fluids of splenic T cells from normal mice cultured with 1/2D-M(phi) in a dual-chamber transwell system; however, this cytokine was not produced by normal T cells cultured with normal macrophages in the transwells. Also, normal T cells cultured with 1/2D-M(phi) did not produce interleukin-4 when transwell cultures were performed in the presence of anti-MCP-1 monoclonal antibody. Further, normal T cells directly stimulated with MCP-1 produced interleukin-4 into their culture fluids. Normal T cells, cultured with 1/2D-M(phi) for 24 hours in the transwells and recultured with fresh medium for an additional 7 days, produced interleukin 10 (but not interferon-gamma) and expressed ST2L mRNA (but not interleukin-12 receptor beta2 chain) when they were stimulated with anti-CD3 monoclonal antibody. CONCLUSIONS: Results indicate that MCP-1 is produced in mice within 1 day of thermal injury, and the subsequent development of burn-associated type 2 T cell responses may be triggered by MCP-1 produced early after thermal injury. PMID- 12131095 TI - Impact of intraoperative donor management on short-term renal function after laparoscopic donor nephrectomy. AB - OBJECTIVE: To determine whether intraoperative diuresis, postoperative recovery, and early graft function differ between laparoscopic open nephrectomy (LDN) and open donor nephrectomy (ODN). SUMMARY BACKGROUND DATA: Laparoscopic donor nephrectomy can reduce donor complications in terms of decreased pain and shorter convalescence. Although its technical feasibility has been established, concerns have been raised about the impaired renal function resulting from pneumoperitoneum and short- and long-term function of kidneys removed by LDN. METHODS: Between December 1997 and December 2000, 89 LDNs were performed at the authors' institution. These were compared with 83 conventional ODNs performed between January 1994 and December 1997. Graft function, intraoperative variables, and clinical outcome were compared. RESULTS: Laparoscopic donor nephrectomy was attempted in 89 patients and completed in 91% (81/89). Length of hospital stay was significantly shorter in the laparoscopic group. During kidney dissection, the amount of fluids administered and intraoperative diuresis were significantly lower for LDN. In recipients, mean serum creatinine was higher after LDN compared with ODN 1 day after surgery. From postoperative days 2 until 28, there were no differences in serum creatinine. Graft survival rates were similar for LDN and ODN. CONCLUSIONS: Donors can benefit from an improvement in postoperative recovery after LDN. Assessment of an adequate perioperative hydration protocol is mandatory to ensure optimal kidney quality during laparoscopic procurement. The initial graft survival and function rates justify continued development and adoption of LDN. PMID- 12131096 TI - Editorial view: anesthetic preconditioning: serendipity and science. PMID- 12131097 TI - Volatile anesthetics mimic cardiac preconditioning by priming the activation of mitochondrial K(ATP) channels via multiple signaling pathways. AB - BACKGROUND: Volatile anesthetics induce pharmacological preconditioning in cardiac tissue. The purpose of this study was to test whether volatile anesthetics mediate this effect by activation of the mitochondrial adenosine triphosphate-sensitive potassium (mitoK(ATP)) or sarcolemmal K(ATP) (sarcK(ATP)) channel in rat ventricular myocytes and to evaluate the signaling pathways involved. METHODS: A cellular model of ischemia with subsequent hypoosmolar trypan blue staining served to determine the effects of 5-hydroxydecanoate, a selective mitoK(ATP) channel blocker, HMR-1098, a selective sarcK(ATP) channel blocker, diazoxide, a preconditioning mimicking agent, and various modulators of putative signaling pathways on cardioprotection elicited by sevoflurane and isoflurane. Microscopy was used to visualize and measure autofluorescence of flavoproteins, a direct index of mitoK(ATP) channel activity. RESULTS: Volatile anesthetics significantly enhanced diazoxide-mediated activation of mitoK(ATP) channels as assessed by autofluorescence of myocytes. Conversely, volatile anesthetics alone did not alter mitoK(ATP) channel activity, implying a priming effect of volatile anesthetics on mitoK(ATP) channels. Administration of the protein kinase C inhibitor chelerythrine completely blocked this effect. Also, pretreatment with volatile anesthetics potentiated diazoxide-mediated protection against ischemia, as indicated by a reduction in trypan blue-positive myocytes. Importantly, cardioprotection afforded by volatile anesthetics was unaffected by the sarcK(ATP) channel blocker HMR-1098 but sensitive to modulations of nitric oxide and adenosine-G(i) signaling pathways. CONCLUSIONS: Using autofluorescence in live cell imaging microscopy and a simulated model of ischemia, the authors present evidence that volatile anesthetics mediate their protection in cardiomyocytes by selectively priming mitoK(ATP) channels through multiple triggering protein kinase C-coupled signaling pathways. These observations provide important new insight into the mechanisms of anesthetic-induced preconditioning. PMID- 12131099 TI - Differential effects of anesthetics on mitochondrial K(ATP) channel activity and cardiomyocyte protection. AB - BACKGROUND: Mitochondrial adenosine triphosphate-sensitive potassium (mitoK(ATP)) channels play a pivotal role in mediating cardiac preconditioning. The effects of intravenous anesthetics on this protective channel have not been investigated so far, but would be of importance with respect to experimental as well as clinical medicine. METHODS: Live cell microscopy was used to visualize and measure autofluorescence of flavoproteins, a direct reporter of mitoK(ATP) channel activity, in response to the direct and highly selective mitoK(ATP) channel opener diazoxide, or to diazoxide following exposure to various anesthetics commonly used in experimental and clinical medicine. A cellular model of ischemia with subsequent hypoosmolar trypan blue staining served to substantiate the effects of the anesthetics on mitoK(ATP) channels with respect to myocyte viability. RESULTS: Diazoxide-induced mitoK(ATP) channel opening was significantly inhibited by the anesthetics R-ketamine, and the barbiturates thiopental and pentobarbital. Conversely, urethane, 2,2,2-trichloroethanol (main metabolite of alpha-chloralose and chloral hydrate), and the opioid fentanyl potentiated the channel-opening effect of diazoxide, which was abrogated by coadministration of chelerythrine, a specific protein kinase C inhibitor. S ketamine, propofol, xylazine, midazolam, and etomidate did not affect mitoK(ATP) channel activity. The significance of these modulatory effects of the anesthetics on mitoK(ATP) channel activity was substantiated in a cellular model of simulated ischemia, where diazoxide-induced cell protection was mitigated by R-ketamine and the barbiturates, while urethane, 2,2,2-trichloroethanol, and fentanyl potentiated myocyte protection. CONCLUSIONS: These results suggest distinctive actions of individual anesthetics on mitoK(ATP) channels and provide evidence that the choice of background anesthesia may play a role in cardiac protection in both experimental and clinical medicine. PMID- 12131100 TI - Isoflurane pretreatment inhibits cytokine-induced cell death in cultured rat smooth muscle cells and human endothelial cells. AB - BACKGROUND: Anesthetics are protective during ischemic-reperfusion injury and associated inflammation; therefore, the authors hypothesized that anesthetic pretreatment may provide protection in culture from cytokine-induced cell death. METHODS: Rat vascular smooth muscle (VSM) cell and human umbilical vascular endothelial cell (HUVEC) cultures were used to determine whether pretreatment with 30 min of isoflurane decreases cell death from tumor necrosis factor alpha (TNF-alpha), interleukin 1 (IL-1 beta), and interferon (IFN-gamma) alone or in combination. Cell survival and viability were determined by trypan blue staining and cell proliferation assay, as well as by DNA fragmentation assays. The roles of protein kinase C (PKC) and adenosine triphosphate-sensitive potassium (K(ATP)) channels in mediating isoflurane (and halothane) protection were evaluated with the antagonists staurosporine or glibenclamide in cytokine- and also hydrogen peroxide (H(2)O(2))-induced cell death. RESULTS: Pretreatment with 1.5% isoflurane immediately prior to cytokine exposure increased cell survival and viability from cytokines by 10-60% for 24, 48, 72, and 96 h in VSMs and up to 72 h in HUVECs. DNA fragmentation (TUNEL) was also attenuated by isoflurane. Isoflurane was equally effective in VSMs at 0.75, 1.5, and 2.5%, whereas in HUVECs, 1.5 and 2.5% were more effective than 0.75%. In VSMs, isoflurane administered 1 h prior to or simultaneously with cytokines was also effective, whereas isoflurane 2 h prior to cytokines was less effective, and either 4 h prior to or 30 min after cytokines was not effective. In both cytokine- and H(2)O(2)-induced cell death, isoflurane and halothane pretreatment were equally protective, and staurosporine and glibenclamide attenuated the protective effect. CONCLUSIONS: Thirty minutes of isoflurane attenuates cytokine-induced cell death and increases cell viability in VSMs for 96 h and in HUVECs for 72 h. Isoflurane must be administered less than 2 h prior to or simultaneously with the cytokines to be protective. These initial inhibitor studies suggest involvement of PKC and K(ATP) channels in isoflurane and halothane protection against both cytokine- and H(2)O(2)-induced cell death of VSMs and HUVECs. PMID- 12131101 TI - Mechanisms of desflurane-induced preconditioning in isolated human right atria in vitro. AB - BACKGROUND: The authors examined the role of adenosine triphosphate-sensitive potassium (K(ATP)) channels, adenosine A1 receptor, and alpha and beta adrenoceptors in desflurane-induced preconditioning in human myocardium, in vitro. METHODS: The authors recorded isometric contraction of human right atrial trabeculae suspended in oxygenated Tyrode's solution (34 degrees C; stimulation frequency, 1 Hz). Before a 30-min anoxic period, 3, 6, and 9% desflurane was administered during 15 min. Desflurane, 6%, was also administered in the presence of 10 microm glibenclamide, a K(ATP) channels antagonist; 10 microm HMR 1098, a sarcolemmal K(ATP) channel antagonist; 800 microm 5-hydroxy-decanoate (5-HD), a mitochondrial K(ATP) channel antagonist; 1 microm phentolamine, an alpha adrenoceptor antagonist; 1 microm propranolol, a beta-adrenoceptor antagonist; and 100 nm 8-cyclopentyl-1,3-dipropylxanthine (DPX), the adenosine A1 receptor antagonist. Developed force at the end of a 60-min reoxygenation period was compared (mean +/- SD). RESULTS: Desflurane at 3% (95 +/- 13% of baseline), 6% (86 +/- 6% of baseline), and 9% (82 +/- 6% of baseline) enhanced the recovery of force after 60 min of reoxygenation as compared with the control group (50 +/- 11% of baseline). Glibenclamide (60 +/- 12% of baseline), 5-HD (57 +/- 21% of baseline), DPX (63 +/- 19% of baseline), phentolamine (56 +/- 20% of baseline), and propranolol (63 +/- 13% of baseline) abolished desflurane-induced preconditioning. In contrast, HMR 1098 (85 +/- 12% of baseline) did not modify desflurane-induced preconditioning. CONCLUSIONS: In vitro, desflurane preconditions human myocardium against simulated ischemia through activation of mitochondrial K(ATP) channels, adenosine A1 receptor, and alpha and beta adrenoceptors. PMID- 12131102 TI - Sevoflurane but not propofol preserves myocardial function in coronary surgery patients. AB - BACKGROUND: Sevoflurane has been shown to protect against myocardial ischemia and reperfusion injury in animals. The present study investigated whether these effects were clinically relevant and would protect left ventricular (LV) function during coronary surgery. METHODS: Twenty coronary surgery patients were randomly assigned to receive either target-controlled infusion of propofol or inhalational anesthesia with sevoflurane. Except for this, anesthetic and surgical management was the same in all patients. A high-fidelity pressure catheter was positioned in the left ventricle and the left atrium. LV response to increased cardiac load, obtained by leg elevation, was assessed before and after cardiopulmonary bypass (CPB). Effects on contraction were evaluated by analysis of changes in dP/dt(max). Effects on relaxation were assessed by analysis of the load dependence of myocardial relaxation (R = slope of the relation between time constant tau of isovolumic relaxation and end-systolic pressure). Postoperative concentrations of cardiac troponin I were followed during 36 h. RESULTS: Before CPB, leg elevation slightly increased dP/dt(max) in the sevoflurane group (5 +/- 3%), whereas it remained unchanged in the propofol group (1 +/- 6%). After CPB, leg elevation resulted in a decrease in dP/dt(max) in the propofol group (-5 +/- 4%), whereas the response in the sevoflurane group was comparable to the response before CPB (5 +/- 4%). Load dependence of LV pressure fall (R) was similar in both groups before CPB. After CPB, R was increased in the propofol group but not in the sevoflurane group. Troponin I concentrations were significantly lower in the sevoflurane than in the propofol group. CONCLUSIONS: Sevoflurane preserved LV function after CPB with less evidence of myocardial damage in the first 36 h postoperatively. These data suggest a cardioprotective effect of sevoflurane during coronary artery surgery. PMID- 12131103 TI - Differential modulation of the cardiac adenosine triphosphate-sensitive potassium channel by isoflurane and halothane. AB - BACKGROUND: The cardiac adenosine triphosphate-sensitive potassium (K(ATP)) channel is activated during pathophysiological episodes such as ischemia and hypoxia and may lead to beneficial effects on cardiac function. Studies of volatile anesthetic interactions with the cardiac K(ATP) channel have been limited. The goal of this study was to investigate the ability of volatile anesthetics halothane and isoflurane to modulate the cardiac sarcolemmal K(ATP) channel. METHODS: The K(ATP) channel current (I(KATP)) was monitored using the whole cell configuration of the patch clamp technique from single ventricular cardiac myocytes enzymatically isolated from guinea pig hearts. I(KATP) was elicited by extracellular application of the potassium channel openers 2,4 dinitrophenol or pinacidil. RESULTS: Volatile anesthetics modulated I(KATP) in an anesthetic-dependent manner. Isoflurane facilitated the opening of the K(ATP) channel. Following initial activation of I(KATP) by 2,4-dinitrophenol, isoflurane at 0.5 and 1.3 mm further increased current amplitude by 40.4 +/- 11.1% and 58.4 +/- 20.6%, respectively. Similar results of isoflurane were obtained when pinacidil was used to activate I(KATP). However, isoflurane alone was unable to elicit K(ATP) channel opening. In contrast, halothane inhibited I(KATP) elicited by 2,4-dinitrophenol by 50.6 +/- 5.8% and 72.1 +/- 11.6% at 0.4 and 1.0 mm, respectively. When I(KATP) was activated by pinacidil, halothane had no significant effect on the current. CONCLUSIONS: The cardiac sarcolemmal K(ATP) channel is differentially modulated by volatile anesthetics. Isoflurane can facilitate the further opening of the K(ATP) channel following initial channel activation by 2,4-dinitrophenol or pinacidil. The effect of halothane was dependent on the method of channel activation, inhibiting I(KATP) activated by 2,4-dinitrophenol but not by pinacidil. PMID- 12131104 TI - Isoflurane-induced facilitation of the cardiac sarcolemmal K(ATP) channel. AB - BACKGROUND: Volatile anesthetics have cardioprotective effects that mimic ischemic preconditioning, including the involvement of adenosine triphosphate sensitive potassium (K(ATP)) channels. However, evidence for a direct effect of volatile anesthetic on the K(ATP) channel is limited. In this study, the effects of isoflurane on the cardiac sarcolemmal K(ATP) channel were investigated. METHODS: Single ventricular myocytes were enzymatically isolated from guinea pig hearts. Whole cell and single-channel configurations, specifically the cell attached and inside-out patch mode, of the patch clamp technique were used to monitor sarcolemmal K(ATP) channel current. RESULTS: In the cell-attached patch configuration, 2,4-dinitrophenol (150 microm) opened the sarcolemmal K(ATP) channel. Isoflurane (0.5 mm) further increased channel open probability and the number of active channels in the patch. In contrast, in the inside-out patch experiments, isoflurane had no significant effect on the K(ATP) channel activated by low ATP (0.2-0.5 mm). In addition, isoflurane had no effect on the K(ATP) channel when activated by adenosine diphosphate, adenosine + guanosine triphosphate, bimakalim, and 2,4-dinitrophenol under inside-out patch configurations. When K(ATP) current was monitored in the whole cell mode, isoflurane alone was unable to elicit channel opening. However, during sustained protein kinase C activation by 12,13-dibutyrate, isoflurane activated the K(ATP) current that was sensitive to glibenclamide. In contrast, isoflurane had no effect on the K(ATP) channel activated by 12,13-dibutyrate in a cell-free environment. CONCLUSIONS: Isoflurane facilitated the opening of the sarcolemmal K(ATP) channel in the intact cell, but not in an excised, inside-out patch. The isoflurane effect was not due to a direct interaction with the K(ATP) channel protein, but required an intracellular component, likely including the translocation of specific protein kinase C isoforms. This suggests that the sarcolemmal K(ATP) channel may have a significant role in anesthetic-induced preconditioning. PMID- 12131105 TI - Eliminating intensive postoperative care in same-day surgery patients using short acting anesthetics. AB - BACKGROUND: A multidisciplinary effort was undertaken to determine whether patients could safely bypass the postanesthesia care unit (PACU) after same-day surgery by moving to an earlier time point evaluation of recovery criteria. METHODS: A prospective, outcomes research study with a baseline month, an intervention month, and a follow-up month was designed. Five surgical centers (three community-based hospitals and two freestanding ambulatory surgical centers) were utilized. Two thousand five hundred eight patients were involved in the baseline period, and 2,354 were involved in the follow-up period. Outcome measures included PACU bypass rates and adverse events. Intervention consisted of a multidisciplinary educational program and routine feedback reports. RESULTS: The overall PACU bypass rate (58%) was significantly different from baseline (15.9%, P < 0.001), for patients to whom a general anesthetic was administered (0.4-31.8%, P < 0.001), and for those given other anesthetic techniques (monitored anesthesia care, regional or local anesthetics; 29.1-84.2%, P < 0.001). During the follow-up period, the average (SD) recovery duration for patients who bypassed the PACU was significantly shorter compared to that for patients who did not bypass, 84.6 (61.5) versus 175.1 (98.8) min, P < 0.001, with no change in patient outcome. Patients receiving only short-acting anesthetics were 78% more likely (P < 0.002) to bypass the PACU after adjusting for various surgical procedures. CONCLUSIONS: This study represents a substantial change in clinical practice in the perioperative setting. Same-day surgical patients given short-acting anesthetic agents and who are awake, alert, and mobile requiring no parenteral pain medications and with no bleeding or nausea at the end of an operative procedure can safely bypass the PACU. PMID- 12131106 TI - Positioning in anesthesiology: toward a better understanding of stretch-induced perioperative neuropathies. AB - BACKGROUND: Stretch-induced neuropathy of the brachial plexus and median nerve in conventional perioperative care remains a relatively frequent and poorly understood complication. Guidelines for positioning have been formulated, although the protective effect of most recommendations remains unexamined. The similarity between the stipulated potentially dangerous positions and the components of the brachial plexus tension test (BPTT) justified the analysis of the BPTT to quantify the impact of various arm and neck positions on the peripheral nervous system. METHODS: Four variations of the BPTT in three different shoulder positions were performed in 25 asymptomatic male participants. The impact of arm and neck positions on the peripheral nervous system was evaluated by analyzing the maximal available range of motion, pain intensity, and type of elicited symptoms during the BPTT. RESULTS: Cervical contralateral lateral flexion, lateral rotation of the shoulder and fixation of the shoulder girdle in a neutral position in combination with shoulder abduction, and wrist extension all significantly reduced the available range of motion. Elbow extension also challenged the nervous system substantially. A cumulative impact could be observed when different components were simultaneously added, and a neutralizing effect was noted when an adjacent region allowed for unloading of the nervous system. CONCLUSIONS: The experimental findings support the experientially based guidelines for positioning. Especially when simultaneously applied, submaximal joint positions easily load the nervous system, which may substantially compromise vital physiologic processes in and around the nerve. Therefore, even when the positioning of all upper limb joints is carefully considered, complete prevention of perioperative neuropathy seems almost inconceivable. PMID- 12131107 TI - Patient State Index: titration of delivery and recovery from propofol, alfentanil, and nitrous oxide anesthesia. AB - BACKGROUND: The Patient State Index (PSI) uses derived quantitative electroencephalogram features in a multivariate algorithm that varies as a function of hypnotic state. Data are recorded from two anterior, one midline central, and one midline posterior scalp locations. PSI has been demonstrated to have a significant relation to level of hypnosis during intravenous propofol, inhalation, and nitrous oxide-narcotic anesthesia. This multisite study evaluated the utility of PSI monitoring as an adjunct to standard anesthetic practice for guiding the delivery of propofol and alfentanil to accelerate emergence from anesthesia. METHODS: Three hundred six patients were enrolled in this multicenter prospective randomized clinical study. Using continuous monitoring throughout the period of propofol-alfentanil-nitrous oxide anesthesia delivery, PSI guidance was compared with use of standard practice guidelines (both before [historic controls] and after exposure to the PSA 4000 monitor [Physiometrix, Inc., N. Billerica, MA; standard practice controls]). Anesthesia was always administered with the aim of providing hemodynamic stability, with rapid recovery. RESULTS: No significant differences were found for demographic variables or for site. The PSI group received significantly less propofol than the standard practice control group (11.9 microg x kg(-1) x min(-1); P < 0.01) and historic control group (18.2 microg x kg(-1) x min(-1); P < 0.001). Verbal response time, emergence time, extubation time, and eligibility for operating room discharge time were all significantly shorter for the PSI group compared with the historic control (3.3 3.8 min; P < 0.001) and standard practice control (1.4-1.5 min; P < 0.05 or P < 0.01) groups. No significant differences in the number of unwanted somatic events or hemodynamic instability and no incidences of reported awareness were found. CONCLUSIONS: Patient State Index-directed titration of propofol delivery resulted in faster emergence and recovery from propofol-alfentanil-nitrous oxide anesthesia, with modest decrease in the amount of propofol delivered, without increasing the number of unwanted events. PMID- 12131108 TI - The influence of mild hypothermia on the pharmacokinetics and time course of action of neostigmine in anesthetized volunteers. AB - BACKGROUND: The pharmacokinetics, maximum effect, and time course of action of neostigmine were studied in seven human volunteers. METHODS: Each volunteer was studied twice, during both normothermia and hypothermia. Anesthesia was induced with 30 microg/kg alfentanil and 3 mg/kg propofol, and was maintained with 60-70% nitrous oxide and 0.7-0.9% isoflurane. The mechanical response of the adductor pollicis to train-of-four stimulation of the ulnar nerve was recorded, and central body temperature maintained stable at either less than 34.5 degrees C or greater than 36.5 degrees C by surface cooling or warming. Before neostigmine administration, a stable 5% twitch height was obtained by an infusion of vecuronium, and the infusion rate remained unchanged thereafter. Neostigmine, 70 microg/kg, was then infused over 2 min, and blood samples for estimation of neostigmine concentrations were collected at intervals for 240 min. RESULTS: With hypothermia, the central volume of distribution of neostigmine decreased by 38%, and onset time of maximum effect increased (4.6 vs. 5.6 min). Hypothermia did not change the clearance (696 ml/min), maximum effect, or duration of action of neostigmine. CONCLUSIONS: The efficacy of neostigmine as an antagonist of vecuronium-induced neuromuscular block is not altered by mild hypothermia. PMID- 12131109 TI - Effects of subhypnotic doses of propofol on gastric emptying in volunteers. AB - BACKGROUND: Drugs which accelerate gastric emptying (GE) decrease nausea and vomiting. This could contribute to the antiemetic potential of subhypnotic doses of propofol. On the contrary, subhypnotic doses of propofol used for sedation could decrease GE and thus favor regurgitation and pulmonary inhalation. Therefore, the aim of this study was to assess the effect of low-dose propofol infusion on GE. METHODS: On three separate occasions, 10 volunteers received either a propofol infusion at a rate set to achieve a target plasma concentration of 0.5 microg/ml or equivalent volumes of 10% Intralipid(R) or 0.9% saline. GE for solids was measured by using the octanoic acid breath test. An acetaminophen absorption technique measured the GE rate for liquids. Blood samples were assayed for acetaminophen and propofol. Breath samples were analyzed for (13)CO(2) concentration by isotope-ratio mass spectrometry. Carbon dioxide production (VCO(2)) was measured instead of calculated by indirect calorimetry. Sedation was evaluated by the Bispectral Index of the electroencephalogram. RESULTS: Propofol blood concentrations were 0.32 +/- 0.20 and 0.45 +/- 0.18 microg/ml at 60 and 165 min, respectively. These concentrations were not sedative. Propofol or its solvent did not modify GE for solids or liquids. In all groups, differences in GE were obtained if measured VCO(2) was integrated in the formula instead of calculated VCO(2) (P < 0.002). CONCLUSIONS: Subhypnotic doses of propofol known to be antiemetic do not inhibit GE. These results suggest that the antiemetic properties of propofol are not peripheral and that propofol cannot be considered as a prokinetic agent. V(13)CO(2) must be measured instead of calculated to accurately determine GE. PMID- 12131110 TI - Gabapentin suppresses cutaneous hyperalgesia following heat-capsaicin sensitization. AB - BACKGROUND: The anticonvulsant gabapentin, proven effective for neuropathic pain in two large, placebo-controlled clinical trials, is widely used for treatment of chronic pain. Preclinical studies have demonstrated analgesic and antiallodynic effects in models involving neuronal sensitization and nerve injury, without affecting acute pain transmission. The aim of the present study was to link data from animal models and clinical trials for chronic pain by investigating the effect of gabapentin on acute nociception and experimentally induced cutaneous hyperalgesia in healthy volunteers. METHODS: The human experimental hyperalgesia model, the heat-capsaicin sensitization model, was induced in 25 healthy male volunteers. Subjects received oral gabapentin (1,200 mg) or placebo after heat capsaicin sensitization was established on the forearm. The primary outcome measures were the sizes of the areas of secondary hyperalgesia to von Frey hair and brush stimulation on the forearm. Secondary outcome measures were as follows: (1) size of secondary hyperalgesia area in response to brief thermal sensitization procedure on the thigh; (2) heat pain detection thresholds in normal and sensitized skin; and (3) painfulness of 1 min of 45 degrees C stimulation in normal skin. RESULTS: Oral gabapentin profoundly suppressed established cutaneous sensitization on the forearm and prevented development of cutaneous sensitization on the thigh. Thermal nociception in normal skin was unchanged. Side effects were modest. CONCLUSION: The results link preclinical findings with results from clinical trials of neuropathic pain. The results further suggest that gabapentin may prove effective in acute pain disorders involving neuronal sensitization, such as postoperative pain and acute herpetic pain, and could prove effective in prevention of chronic pain. PMID- 12131111 TI - Anesthetic-related cardiac arrest and its mortality: a report covering 72,959 anesthetics over 10 years from a US teaching hospital. AB - BACKGROUND: A prospective and retrospective case analysis study of all perioperative cardiac arrests occurring during a 10-yr period from 1989 to 1999 was done to determine the incidence, cause, and outcome of cardiac arrests attributable to anesthesia. METHODS: One hundred forty-four cases of cardiac arrest within 24 h of surgery were identified over a 10-yr period from an anesthesia database of 72,959 anesthetics. Case abstracts were reviewed by a Study Commission composed of external and internal members in order to judge which cardiac arrests were anesthesia-attributable and which were anesthesia contributory. The rates of anesthesia-attributable and anesthesia-contributory cardiac arrest were estimated. RESULTS: Fifteen cardiac arrests out of a total number of 144 were judged to be related to anesthesia. Five cardiac arrests were anesthesia-attributable, resulting in an anesthesia-attributable cardiac arrest rate of 0.69 per 10,000 anesthetics (95% confidence interval, 0.085-1.29). Ten cardiac arrests were found to be anesthesia-contributory, resulting in an anesthesia-contributory rate of 1.37 per 10,000 anesthetics (95% confidence interval, 0.52-2.22). Causes of the cardiac arrests included medication-related events (40%), complications associated with central venous access (20%), problems in airway management (20%), unknown or possible vagal reaction in (13%), and one perioperative myocardial infarction. The risk of death related to anesthesia attributable perioperative cardiac arrest was 0.55 per 10,000 anesthetics (95% confidence interval, 0.011-1.09). CONCLUSIONS: Most perioperative cardiac arrests were related to medication administration, airway management, and technical problems of central venous access. Improvements focused on these three areas may result in better outcomes. PMID- 12131113 TI - The unexpected difficult airway and lingual tonsil hyperplasia: a case series and a review of the literature. AB - BACKGROUND: An unexpected difficult intubation occurs because physical examination of the airway is imperfect in predicting it. Lingual tonsil hyperplasia (LTH) is one risk factor for an unanticipated failed intubation that is not detectable during a routine oropharyngeal examination. The authors attempted to determine the incidence of LTH in unanticipated failed intubation in patients subjected to general anesthesia. METHODS: Thirty-three patients with unanticipated failed intubation via direct laryngoscopy were subjected to airway examinations and fiberoptic pharyngoscopy to determine the cause(s) of failure. Mouth opening, mandibular subluxation, head extension, thyromental distance, and Mallampati airway class were recorded. Fiberoptic pharyngoscopy was then performed to evaluate the base of the tongue and valleculae. RESULTS: Of these 33 patients, none had an airway examination that suggested a difficult intubation. The lungs of 12 patients were difficult to ventilate by mask. In 15 patients, airway measurements were within normal limits with Mallampati class of I or II. Ten patients had a Mallampati class III airway, 6 associated with obesity and 5 with mildly limited head extension. Among the 5 morbidly obese patients, most of the weight was distributed on the lower trunk and body. The 3 remaining patients had a thyromental distance of 6 cm or less but otherwise had a normal airway examination. The only finding common to all 33 patients was LTH observed on fiberoptic pharyngoscopy. CONCLUSION: Lingual tonsil hyperplasia can interfere with rigid laryngoscopic intubation and face mask ventilation. Routine physical examination of the airway will not identify its presence. The prevalence of LTH in adults and the extent of its contribution to failed intubation is unknown. PMID- 12131112 TI - High-dose amrinone is required to accelerate rewarming from deliberate mild intraoperative hypothermia for neurosurgical procedures. AB - BACKGROUND: Since the time available to provide the cooling and rewarming is limited during deliberate mild hypothermia, the technique to accelerate the cooling and rewarming rate of core temperature has been studied. Amrinone has been reported to accelerate the cooling rate but not the rewarming rate of core temperature during deliberate mild hypothermia. The failure of amrinone effect on the rewarming rate might be due to an insufficient dose of amrinone during hypothermic conditions. The authors therefore tested whether higher doses of amrinone can accelerate the rewarming rate of core temperature during deliberate mild hypothermia for neurosurgery. METHODS: After institutional approval and informed consent, 30 patients were randomly assigned to one of three groups. Patients in the control group (n = 10) did not receive amrinone; patients in the AMR 15 group (n = 10) received 15 microg x kg(-1) x min(-1) amrinone with a 1.0 mg/kg loading dose of amrinone at the beginning of cooling; and patients in the ReAMR group (n = 10) received 5 microg x kg(-1) x min(-1) amrinone with 1.0-mg/kg loading and reloading doses of amrinone at the beginning of cooling and rewarming, respectively. Administration of amrinone was started just after the induction of cooling and continued until the end of anesthesia. Anesthesia was maintained with nitrous oxide in oxygen, propofol, and fentanyl. After induction of anesthesia, patients were cooled, and tympanic membrane temperature was maintained at 34.5 degrees C. After completion of the main surgical procedures, patients were actively rewarmed and extubated in the operating room. RESULTS: The cooling and rewarming rates of core temperature were both significantly faster in both amrinone groups than in the control group. During the cooling and rewarming periods, forearm minus fingertip temperature gradient was significantly smaller in both amrinone groups than in the control group. During the rewarming period, heart rate and mean arterial pressure in the AMR 15 group were significantly faster and lower, respectively, than in the control group. Systemic vascular resistance in the AMR 15 group was smaller than in the control group throughout the study; on the other hand, only the value after the start of rewarming in the ReAMR group was smaller than in the control group. CONCLUSIONS: Amrinone at an infusion rate of 15 or 5 microg x kg(-1) x min(-1) with a reloading at the beginning of rewarming accelerated the rewarming rate of core temperature during deliberate mild hypothermia. This suggests that high-dose amrinone is required to accelerate rewarming from deliberate mild intraoperative hypothermia for neurosurgical procedures. PMID- 12131114 TI - In vivo uptake and elimination of isoflurane by different membrane oxygenators during cardiopulmonary bypass. AB - BACKGROUND: Volatile anesthetics are frequently used during cardiopulmonary bypass (CPB) to maintain anesthesia. Uptake and elimination of the volatile agent are dependent on the composition of the oxygenator. This study was designed to evaluate whether the in vivo uptake and elimination of isoflurane differs between microporous membrane oxygenators containing a conventional polypropylene (PPL) membrane and oxygenators with a new poly-(4-methyl-1-pentene) (PMP) membrane measuring isoflurane concentrations in blood. METHODS: Twenty-four patients undergoing elective coronary bypass surgery with the aid of CPB were randomly allocated to one of four groups, using either one of two different PPL-membrane oxygenators for CPB or one of two different PMP-membrane oxygenators. During hypothermic CPB, 1% isoflurane in an oxygen-air mixture was added to the oxygenator gas inflow line (gas flow, 3 l/min) for 15 min. Isoflurane concentration was measured in blood and in exhaust gas at the outflow port of the oxygenator. Between-group comparisons were performed for the area under the curve (AUC) during uptake and elimination of the isoflurane blood concentrations, the maximum isoflurane blood concentration (C(max)), and the exhausted isoflurane concentration (F(E)). RESULTS: The uptake of isoflurane, expressed as AUC of isoflurane blood concentration and a function of F(E), was significantly reduced in PMP oxygenators compared to PPL oxygenators (P < 0.01). C(max) was between 8.5 and 13 times lower in the PMP-membrane oxygenator groups compared to the conventional PPL-membrane oxygenator groups (P < 0.01). CONCLUSIONS: The uptake of isoflurane into blood via PMP oxygenators during CPB is severely limited. This should be taken into consideration in cases using such devices. PMID- 12131115 TI - Reengineering intravenous drug and fluid administration processes in the operating room: step one: task analysis of existing processes. AB - BACKGROUND: A reengineering approach to intravenous drug and fluid administration processes could improve anesthesia care. In this initial study, current intravenous administration tasks were examined to identify opportunities for improved design. METHODS: After institutional review board approval was obtained, an observer sat in the operating room and categorized, in real time, anesthesia providers' activities during 35 cases ( approximately 90 h) into 66 task categories focused on drug/fluid tasks. Both initial room set-up at the beginning of a typical workday and cardiac and noncardiac general anesthesia cases were studied. User errors and inefficiencies were noted. The time required to prepare de novo a syringe containing a mock emergency drug was measured using a standard protocol. RESULTS: Drug/fluid tasks consumed almost 50 and 75%, respectively, of the set-up time for noncardiac and cardiac cases. In 8 cardiac anesthetics, drug/fluid tasks comprised 27 +/- 6% (mean +/- SD) of all prebypass clinical activities. During 20 noncardiac cases, drug/fluid tasks comprised 20 +/- 8% of induction and 15 +/- 7% of maintenance. Drug preparation far outweighed drug administration tasks. Inefficient or error prone tasks were observed during drug/fluid preparation (e.g., supply acquisition, waste disposal, syringe labeling), administration (infusion device failure, leaking stopcock), and organization (workspace organization and navigation, untangling of intravenous lines). Anesthesia providers (n = 21) required 35 +/- 5 s to prepare a mock emergency drug. CONCLUSIONS: Intravenous drug and fluid administration tasks account for a significant proportion of anesthesia care, especially in complex cases. Current processes are inefficient and may predispose to medical error. There appears to be substantial opportunity to improve quality and cost of care through the reengineering of anesthesia intravenous drug and fluid administration processes. General design requirements are proposed. PMID- 12131116 TI - Impact of unplanned extubation and reintubation after weaning on nosocomial pneumonia risk in the intensive care unit: a prospective multicenter study. AB - BACKGROUND: The authors prospectively evaluated the occurrence and outcomes of unplanned extubations (self-extubation and accidental extubation) and reintubation after weaning, and examined the hypothesis that these events may differ regarding their influence on the risk of nosocomial pneumonia. METHODS: Data were taken from a prospective, 2-yr database including 750 mechanically ventilated patients from six intensive care units. RESULTS: One hundred five patients (14%) experienced at least one episode of these 3 events; 51 self extubations occurred in 38 patients, 24 accidental extubations in 22 patients, and 56 reintubations after weaning in 45 patients. The incidence density of these 3 events was 16.4 per 1,000 mechanical ventilation days. Reintubation within 48 h was needed consistently after accidental extubation but was unnecessary in 37% of self-extubated patients. Unplanned extubation and reintubation after weaning were associated with longer total mechanical ventilation (17 vs. 6 days; P < 0.0001), intensive care unit stay (22 vs. 9 days; P < 0.0001), and hospital stay (34 vs. 18 days; P < 0.0001) than in control group, but did not influence intensive care unit or hospital mortality. The incidence of nosocomial pneumonia was significantly higher in patients with unplanned extubation or reintubation after weaning (27.6% vs. 13.8%; P = 0.002). In a Cox model adjusting on severity at admission, unplanned extubation and reintubation after weaning increased the risk of nosocomial pneumonia (relative risk, 1.80; 95% confidence interval, 1.15-2.80; P = 0.009). This risk increase was entirely ascribable to accidental extubation (relative risk, 5.3; 95% confidence interval, 2.8-9.9; P < 0.001). CONCLUSION: Accidental extubation but not self-extubation or reintubation after weaning increased the risk of nosocomial pneumonia. These 3 events may deserve evaluation as an indicator for quality-of-care studies. PMID- 12131117 TI - Differences in systemic opioid use do not explain increased fever incidence in parturients receiving epidural analgesia. AB - BACKGROUND: It has been hypothesized that an increased incidence of fever in patients receiving epidural analgesia might result not from epidural per se, but rather from the antipyretic effect of opioids preferentially administered to women in the no-epidural group. If this were the case, then one would expect the incidence of fever in parturients who did not receive systemic opioids to be independent of whether they received epidural analgesia. METHODS: Using a cohort study design, the authors evaluated the records of 1,233 nulliparous patients whose labor analgesia was managed with (1) no medication (N = 170); (2) 10 mg intravenous systemic nalbuphine plus 10 mg intramuscular every 3 to 4 h as required (N = 327); (3) epidural analgesia with continuous infusion of 0.125% bupivacaine with 2 microg/ml fentanyl (N = 278); or (4) patients who received both systemic nalbuphine and epidural analgesia (N = 458). Fever was diagnosed if the maximum temperature during labor exceeded 100.4 degrees F (38 degrees C). RESULTS: The incidence of fever did not differ according to nalbuphine administration for women not receiving epidural analgesia (1% no nalbuphine, 0.3% with nalbuphine, P = 0.27) or for women receiving epidural analgesia (17% no nalbuphine, 17% with nalbuphine, P = 1.0). However, the incidence of fever differed significantly between patients who received no analgesia as compared to those who received epidural analgesia alone (1% vs. 17%, P = 10(-6)). Controlling for confounding did not alter these associations. CONCLUSIONS: Our findings suggest that an antipyretic effect of nalbuphine in patients who do not receive an epidural does not explain the greater incidence of fever observed in women who receive epidural analgesia for labor. PMID- 12131118 TI - Esmolol improves left ventricular function via enhanced beta-adrenergic receptor signaling in a canine model of coronary revascularization. AB - BACKGROUND: Recent American Heart Association guidelines highlight the paucity of data on effectiveness and/or mechanisms underlying use of beta-adrenergic receptor (beta AR) antagonists after acute coronary syndromes in patients subsequently undergoing revascularization. It is important to assess whether beta AR antagonists might protect the heart and improve ventricular function in this scenario. The authors therefore used esmolol (an ultra-short-acting beta AR antagonist) to determine whether beta AR antagonist treatment improves left ventricular function in a canine model of acute reversible coronary ischemia followed by coronary reperfusion during cardiopulmonary bypass (CPB). The authors also tested whether the mechanism includes preserved beta AR signaling. METHODS: Dogs were randomized to either esmolol or saline infusions administered during CPB (n = 29). Pre-CPB and end-CPB transmyocardial left ventricular biopsies were obtained; plasma catecholamine concentrations, myocardial beta AR density, and adenylyl cyclase activity were measured. In addition, left ventricular systolic shortening and postsystolic shortening were determined immediately prior to each biopsy. RESULTS: While beta AR density remained unchanged in each group, isoproterenol-stimulated adenylyl cyclase activity decreased 26 +/- 6% in the control group but increased 38 +/- 10% in the esmolol group (pre-CPB to end-CPB, mean +/- SD, P = 0.0001). Left ventricular systolic shortening improved in both groups after release of coronary (LAD) ligature; however, the esmolol group increased to 72 +/- 23% of pre-CPB values compared to 48 +/- 12% for the control group (P = 0.0008). CONCLUSIONS: These data provide prospective evidence that esmolol administration results in improved myocardial function. Furthermore, the mechanism appears to involve enhanced myocardial beta AR signaling. PMID- 12131119 TI - Effects of a novel selective agonist for prostaglandin receptor subtype EP4 on hyperalgesia and inflammation in monoarthritic model. AB - BACKGROUND: Cytokines have crucial role in the development and maintenance of inflammation and pain in arthritis. Activation of prostaglandin receptor subtype EP(4) suppresses cytokine production in immune cells. The purpose of this study was to evaluate whether a novel EP(4) agonist would be able to suppress thermal and mechanical hyperalgesia and paw swelling in acute and chronic phases in rat monoarthritic model. METHODS: Monoarthritis was induced by an injection of complete Freund's adjuvant (CFA) intracapsularly into the tibiotarsal joint of the rats. Withdrawal latencies to thermal stimulation on the hind paw, withdrawal thresholds to mechanical stimulation, paw volume, and ankle diameter were measured 24 h and 4 weeks after the CFA injection. A novel selective EP(4) receptor agonist, ONO-AE1-329 (10, 25, or 50 microg) or saline was administered intracapsularly into the joint. RESULTS: Withdrawal latencies and withdrawal thresholds were significantly (P < 0.05) shortened and decreased, respectively, on the arthritic side but not on the contralateral side 24 h and 4 weeks after the CFA injection. In addition, significant (P < 0.05) increases in paw volume and ankle diameter on the arthritic side were observed. Intracapsularly administered ONO-AE1-329 showed significant (P < 0.05) inhibition of thermal and mechanical hyperalgesia and significant (P < 0.05) decrease in paw volume and ankle diameter in a dose-dependent manner at 24 h and 4 weeks after CFA. CONCLUSION: Intracapsular administration of EP(4) receptor agonist effectively inhibited mechanical and thermal hyperalgesia and inflammatory reactions in acute and chronic monoarthritis. An EP(4) agonist would be a potential strategy for inflammatory pain in arthritis. PMID- 12131120 TI - Effect of intrathecal non-NMDA EAA receptor antagonist LY293558 in rats: a new class of drugs for spinal anesthesia. AB - BACKGROUND: Excitatory amino acid receptors are important for both sensory and motor function in the spinal cord. We studied the effects of intrathecal LY293558, a competitive non-N-methyl-D-aspartate excitatory amino acid receptor antagonist, on motor and sensory function in rats to determine whether drugs blocking these receptors could potentially be used as alternative agents to local anesthetics for spinal anesthesia. METHODS: Rats were tested before and 15-240 min after intrathecal injection of 5 nmol (in 10 microl) LY293558. Sensory function was tested at the hind paw using withdrawal response to pin prick and withdrawal to pinch with sharp forceps. Motor performance (ambulation, placing reflex, and Rotorod time), blood pressure, and heart rate were also evaluated. Some tests were repeated the next day. Responses after LY293558 were compared to injection of 40 microl bupivacaine, 0.75%. Pin-prick responses at the forepaw, chest, abdomen, hind leg, and hind paw were also examined after intrathecal LY293558. RESULTS: Intrathecal LY293558 blocked both sensory and motor responses through 180 min; complete recovery was present the following day. No change in blood pressure or heart rate occurred. The effects of LY293558 were more pronounced and sustained than those of bupivacaine. Segmental blockade of the response to pin prick was present after LY293558. CONCLUSION: Drugs like LY293558 that block alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid (AMPA)/kainate receptors may be an alternative to local anesthetics for spinal anesthesia in humans. PMID- 12131121 TI - The role of transcranial motor evoked potentials in predicting neurologic and histopathologic outcome after experimental spinal cord ischemia. AB - BACKGROUND: Monitoring of myogenic motor evoked potentials to transcranial stimulation (tcMEPs) is clinically used to assess motor pathway function during aortic and spinal procedures that carry a risk of spinal cord ischemia (SCI). Although tcMEPs presumably detect SCI before irreversible neuronal deficit occurs, and prolonged reduction of tcMEP signals is thought to be associated with impending spinal cord damage, experimental evidence to support this concept has not been provided. In this study, histopathologic and neurologic outcome was examined in a porcine model of SCI after different durations of intraoperative loss of tcMEP signals. METHODS: In 15 ketamine-sufentanil-anesthetized pigs (weight, 35-45 kg) the spinal cord feeding lumbar arteries were exposed. tcMEP were recorded from the upper and lower limbs. Under normothermic conditions, animals were randomly allocated to undergo short-term tcMEP reduction (group A, < 10 min, n = 5) or prolonged tcMEP reduction (group B, 60 min, n = 10), resulting from temporary or permanent clamping of lumbar segmental arteries. Neurologic function was evaluated every 24 h, and infarction volume and the number of eosinophilic neurons and viable motoneurons in the lumbosacral spinal cord was evaluated 72 h after induction of SCI. RESULTS: In all animals except one, segmental artery clamping reduced tcMEP to below 25% of baseline. All but one animal in group A had reduced tcMEP for less than 10 min and had normal motor function and no infarction at 72 h after the initial tcMEP reduction. Seven animals in group B (70%) had reduced tcMEP signals for more than 60 min and were paraplegic with massive spinal cord infarction at 72 h. Two animals (one in both groups) had tcMEP loss for 40 min, with moderate infarction and normal function. In general, histopathologic damage and neurologic dysfunction did not occur when tcMEP amplitude recovered within 10 and 40 min after the initial decline, respectively. CONCLUSION: Prolonged reduction of intraoperative tcMEP amplitude is predictive for postoperative neurologic dysfunction, while recovery of the tcMEP signal within 10 min after the initial decline corresponds with normal histopathology and motor function in this experimental model. This finding confirms that intraoperative tcMEPs have a good prognostic value for neurologic outcome during procedures in which the spinal cord is at risk for ischemia. PMID- 12131123 TI - Influence of lung aeration on pulmonary concentrations of nebulized and intravenous amikacin in ventilated piglets with severe bronchopneumonia. AB - BACKGROUND: Pulmonary concentrations of aminoglycosides administered intravenously are usually low in the infected lung parenchyma. Nebulization represents an alternative to increase pulmonary concentrations, although the obstruction of bronchioles by purulent plugs may impair lung deposition by decreasing lung aeration. METHODS: An experimental bronchopneumonia was induced in anesthetized piglets by inoculating lower lobes with a suspension of 10(6) cfu/ml Escherichia coli. After 24 h of mechanical ventilation, 7 animals received two intravenous injections of 15 mg/kg amikacin, and 11 animals received two nebulizations of 40 mg/kg amikacin at 24-h intervals. One hour following the second administration, animals were killed, and multiple lung specimens were sampled for assessing amikacin pulmonary concentrations and quantifying lung aeration on histologic sections. RESULTS: Thirty-eight percent of the nebulized amikacin (15 mg/kg) reached the tracheobronchial tree. Amikacin pulmonary concentrations were always higher after nebulization than after intravenous administration, decreased with the extension of parenchymal infection, and were significantly influenced by lung aeration: 197 +/- 165 versus 6 +/- 5 microg/g in lung segments with focal bronchopneumonia (P = 0.03), 40 +/- 62 versus 5 +/- 3 microg/g in lung segments with confluent bronchopneumonia (P = 0.001), 18 +/- 7 versus 7 +/- 4 microg/g in lung segments with lung aeration of 30% or less, and 65 +/- 9 versus 2 +/- 3 microg/g in lung segments with lung aeration of 50% or more. CONCLUSIONS: In a porcine model of severe bronchopneumonia, the nebulization of amikacin provided 3-30 times higher pulmonary concentrations than the intravenous administration of an equivalent dose. The greater the lung aeration, the higher were the amikacin pulmonary concentrations found in the infected lung segments. PMID- 12131122 TI - Isoflurane hyperalgesia is modulated by nicotinic inhibition. AB - BACKGROUND: The inhaled anesthetic isoflurane inhibits neuronal nicotinic acetylcholine receptors (nAChRs) at concentrations lower than those used for anesthesia. Isoflurane produces biphasic nociceptive responses, with both hyperalgesia and analgesia within this concentration range. Because nicotinic agonists act as analgesics, the authors hypothesized that inhibition of nicotinic transmission by isoflurane causes hyperalgesia. METHODS: The authors studied female mice at 6-8 weeks of age. They measured hind paw withdrawal latency at isoflurane concentrations from 0 to 0.98 vol% after the animals had received a nicotinic agonist (nicotine), a nicotinic antagonist (mecamylamine or chlorisondamine), or saline intraperitoneally. In addition, the authors tested the interactions between mecamylamine and isoflurane and nicotine and isoflurane in heterologously expressed alpha(4)beta(2) nAChRs. RESULTS: Female mice had significant hyperalgesia from isoflurane. Nicotine administration prevented isoflurane-induced hyperalgesia without altering the antinociception produced by higher isoflurane concentrations. Mecamylamine treatment caused a biphasic nociceptive response similar to that caused by isoflurane. Mecamylamine and isoflurane had an additive effect, both at heterologously expressed alpha(4)beta(2) nAChRs and on the production of hyperalgesia in vivo. Mecamylamine thus potentiated hyperalgesia but did not affect analgesia. CONCLUSIONS: Since hyperalgesia occurs in vivo at isoflurane doses that antagonize nAChRs in vitro, is prevented by a nicotinic agonist, and is mimicked and potentiated by nicotinic antagonists, the authors conclude that isoflurane inhibition of nAChRs activation is involved in the pathway that causes hyperalgesia. At subanesthetic doses, isoflurane can either enhance pain responses (produce hyperalgesia) or be analgesic (antinociceptive). In rats, low volatile anesthetic concentrations (0.1-0.2 minimum alveolar concentration [MAC]) elicit hyperalgesia, while 0.4-0.6 MAC elicits antinociception. PMID- 12131124 TI - Ca(2+)-calmodulin-dependent protein kinase II plays a major role in halothane induced dose-dependent relaxation in the skinned pulmonary artery. AB - BACKGROUND: Previously, the authors have shown in Ca(2+)-clamped skinned arterial strips that protein kinase C (PKC) plays a role in 3% halothane- or isoflurane increased force. PKC in the pulmonary artery and Ca(2+)-calmodulin-dependent protein kinase II (CaMKII) in the femoral artery have been implicated in isoflurane-induced relaxation. For this study, the authors used clinical concentrations of halothane to examine the role of PKC and CaMKII in the halothane-induced biphasic effect on contraction in skinned pulmonary arterial strips. METHODS: Rabbit pulmonary arterial strips were mounted on force transducers and treated with saponin to make the sarcolemma permeable ("skinning"). Skinned strips were activated by low Ca(2+) (pCa 6.3) buffered with 7 mm EGTA, or the PKC activator phorbol-12,13-dibutyrate (PDBu, 1 microm) until force reached a steady state (control). Halothane (1, 2, and 3%) was administered, and the force was observed at peak and 15 min (test results). Ca(2+) ionophore (A23187, 10 microm) and inhibitors were preincubated in a relaxing solution and present in subsequent contracting solutions. Inhibitors were bisindolylmaleimide and Go6976 for PKC, and KN-93 and the inhibitor protein (CKIINtide) for CaMKII. RESULTS: Halothane (1-3%) dose-dependently caused an initial increase (18-35%) and a subsequent decrease (48-68%) in pCa 6.3-induced force. Bisindolylmaleimide, 3 and 10 microm, completely blocked the increase in force at 2% and 3% halothane, respectively. CKIINtide, 0.1 microm, reduced the force at 3% halothane. The decrease in force at 1% and 2% halothane was partially prevented by 0.01 microm bisindolylmaleimide, and at 1, 2, and 3% halothane by 0.01, 0.1, and 1 microm CKIINtide, respectively. At 3% halothane, the increased force was abolished by A23187. In PDBu-induced force, 3% halothane-induced relaxation was also partially prevented by lower concentrations of KN-93 and CKIINtide. CONCLUSIONS: In skinned pulmonary arterial strips, the dose-dependent increase in force by halothane is associated with PKC activation, and that of decrease is associated with CaMKII activation. PMID- 12131125 TI - The systemic inflammatory response to cardiac surgery: implications for the anesthesiologist. PMID- 12131126 TI - The invention and development of blood gas analysis apparatus. AB - In 1953, the doctor draft interrupted Dr. Severinghaus' anesthesia and physiology training and sent him to the National Institutes of Health as director of anesthesia research at the newly opened Clinical Center. He developed precise laboratory partial pressure of carbon dioxide (PCO(2)) and pH analysis to investigate lung blood gas exchange during hypothermia. Constants for carbon dioxide solubility and pK' were more accurately determined. In August 1954, he heard Richard Stow describe invention of a carbon dioxide electrode and immediately built one, improved its stability, and tested its response characteristics. In April 1956, he also heard Leland Clark reveal his invention of an oxygen electrode. Dr. Severinghaus obtained one and constructed a stirred cuvette in which blood partial pressure of oxygen (PO(2)) could be accurately measured. Technician Bradley and Dr. Severinghaus combined these, making the first blood gas analysis system in 1957 and 1958, and shortly thereafter, they added a pH electrode. Blood gas analyzers rapidly developed commercially. Dr. Severinghaus collaborated with Astrup and other Danes on the Haldane and Bohr effects and their concepts of base excess during two sabbaticals in Copenhagen. Work with both Astrup and Roughton on the oxygen dissociation curve led Dr. Severinghaus to devise a modified Hill equation that closely fit their new, better human oxygen dissociation curve and a blood gas slide rule that solved oxygen dissociation curve, PCO(2), pH, and acid-base questions. Blood gas analysis revolutionized both clinical medicine and cardiorespiratory and metabolic physiology. PMID- 12131127 TI - Nerve root inflammation demonstrated by magnetic resonance imaging in a patient with transient neurologic symptoms after intrathecal injection of lidocaine. PMID- 12131128 TI - Spinal anesthesia for a patient with familial hyperkalemic periodic paralysis. PMID- 12131129 TI - Prolonged desflurane administration for refractory status epilepticus. PMID- 12131130 TI - Propofol induced marked prolongation of QT interval in a patient with acute myocardial infarction. PMID- 12131131 TI - EXIT to ECMO. PMID- 12131132 TI - Anaphylactic shock due to suxamethonium complicated by a coronary thrombus. PMID- 12131133 TI - Halothane-induced calcium release in cultured human skeletal muscle cells from a family susceptible to malignant hyperthermia with an unidentified mutation in chromosome 19. PMID- 12131134 TI - Cultured rat trachea as a model for the study of ciliary abundance: the effect of oxygen. PMID- 12131135 TI - Are "international" medical graduate students second-class anesthesiologists? PMID- 12131136 TI - The distribution of the probability of survival and its impact on hypothesis testing in randomized controlled trials. PMID- 12131137 TI - Neurological complications after interscalene brachial plexus blockade: what to make of it? PMID- 12131138 TI - Interscalene block-, sedation-, lateral positioning-, and hydralazine-induced hypotension: is it really prudent? PMID- 12131139 TI - Anesthesia-induced alterations in plasma tracer concentrations may have relevance for brain imaging studies. PMID- 12131140 TI - Combined spinal-epidural versus epidural labor analgesia on progress and outcome of labor. PMID- 12131141 TI - Multiple estimates of EC(50). PMID- 12131142 TI - Pneumatic compression boots, lithotomy stirrups, and lower limb compartment syndrome. PMID- 12131143 TI - Upon what is such a claim founded? PMID- 12131144 TI - Arrhythmia risk of antiemetic agents. PMID- 12131145 TI - FDA "black box" warning regarding use of droperidol for postoperative nausea and vomiting: is it justified? PMID- 12131146 TI - Droperidol "box warning" warrants scrutiny. PMID- 12131147 TI - The FDA droperidol warning: is it justified? PMID- 12131149 TI - Medicine, technology, and human factors in trauma care: a civilian and military perspective. Baltimore, Maryland. November 15-16, 2001. PMID- 12131150 TI - Desmoplastic small round cell tumor: a clinicopathologic, immunohistochemical, and molecular study of 32 tumors. AB - Desmoplastic small round cell tumor is a rare, aggressive neoplasm that mainly affects young male patients and is characterized by a reciprocal translocation t(11;22)(p13;q12) associated with the EWS-WT1 gene fusion transcript. Clinical, histopathologic, immunohistochemical, and molecular genetics features were reviewed for 32 tumors. There were 29 male and three female patients, with ages from 6 to 54 years (mean, 25 years). The main clinical signs and symptoms included abdominal pain (eight patients), weight loss (five patients), and presence of umbilical hernia (four patients). Two tumors primarily involved the ethmoid sinus and the soft tissues of the scalp; the other tumors (mean size, 10 cm) involved the abdominal cavity (88%). One patient presented initially with an axillary lymph node metastasis. Generally, all tumors showed the typical histologic findings of variably sized clusters of small, round, or spindled cells lying in a desmoplastic stroma. The neoplastic cells in formalin-fixed, paraffin embedded tissue sections were positive for desmin (dot pattern) (81% of the cases), WT1 (91%), keratin (87%), neuron-specific enolase (84%), CD99 (23%), and actin (3%). The EWS-WT1 gene fusion transcript was detected in 29 of 30 tumors. One tumor with typical clinicopathologic and immunohistochemical features did not show the gene fusion. Follow-up for 27 patients showed that 19 patients (70%) died of uncontrolled, local, or widespread metastatic disease 3-46 months (mean, 20 months) after diagnosis, and eight patients were alive with known evidence of disease. Occasionally, desmoplastic small round cell tumor lacks the classic clinical, histologic, and immunohistochemical features. This study emphasizes the utility of analysis of the EWS-WT1 gene fusion transcript, which was performed on paraffin-embedded tissues, to confirm the diagnosis. PMID- 12131151 TI - Chondrosarcoma of the larynx: a clinicopathologic study of 111 cases with a review of the literature. AB - Chondrosarcomas of the larynx are rare tumors accounting for about 0.5% of all laryngeal primary tumors. A total of 111 laryngeal chondrosarcoma cases, diagnosed between 1970 and 1997, were retrieved from the Otorhinolaryngic-Head & Neck Tumor Registry of the Armed Forces Institute of Pathology. There was a 3.6:1 male/female ratio of patients 25-91 years of age (mean, 64.4 years). Patients presented most frequently with hoarseness (n = 72 patients) present for a mean of 28.2 months. The majority of tumors involved the cricoid cartilage (n = 77) with a mean size of 3.5 cm. All tumors were invasive and malignant by radiology and/or histology (into bone within the ossified laryngeal cartilages in 52 tumors). Most tumors were low-grade lesions: grade 1 (n = 51), grade 2 (n = 54); there were six grade 3 tumors. An associated benign chondroma with (n = 41 tumors) or without ischemia (n = 24 tumors) was noted. All patients had surgery and five had radiation therapy. Wide excision or voice-sparing surgery was used in 73 patients, whereas 37 patients had a laryngectomy. Recurrences occurred in 20 (18%) patients, 10 of whom underwent salvage laryngectomy. At the last follow-up, 102 patients had no evidence of disease (alive or dead, mean 11.2 years) and five patients had evidence of disease (alive, one patient, 6.5 years; dead, four patients, mean 6.4 years). The six patients with high-grade chondrosarcoma were all without disease at the last follow-up (mean, 15.1 years). There was no difference in clinical outcome based on grade (p = 0.210), location (p = 0.078), or treatment (p = 0.607) but was worse for patients with a myxoid-type chondrosarcoma (p = 0.044). Primary laryngeal chondrosarcomas are typically low- to moderate-grade lesions involving the cricoid cartilage, frequently associated with a chondroma. They usually portend an excellent overall long-term prognosis with initial conservative voice-sparing surgery. PMID- 12131152 TI - 'Agranular CD4+ CD56+ hematodermic neoplasm' (blastic NK-cell lymphoma) originates from a population of CD56+ precursor cells related to plasmacytoid monocytes. AB - In 1999, we reported seven cases of an unusual hematologic malignancy with primary cutaneous presentation that appeared as a distinct clinicopathologic entity characterized by medium-sized tumor cells with a peculiar CD3- CD4+ CD56+ CD43+ HLA-DR+ cell surface phenotype. Because the origin of tumor cells was not clear and they exhibited a nonlineage-specific phenotype, we hypothesized that such tumors likely originated from hematologic-myeloid precursor cells and were tentatively assigned the designation "agranular CD4+ CD56+ hematodermic neoplasms." In the present study we report 14 cases (seven already reported and seven additional cases) of these tumors, and simultaneously we present now a rare population of cells that we have identified in the peripheral blood of healthy volunteers treated with Flt3 ligand. These cells express all the characteristic markers of CD4+ CD56+ hematodermic neoplasms. This population appears to be related to plasmacytoid monocytes because they also expressed CD68 and bright levels of CD123. To confirm the relationship between these normal cells and CD4+ CD56+ hematodermic neoplasms, we conducted an extensive comparative phenotypic study. Results show that these two cell types are indeed related because they share many phenotypic features, including the presence of CD4, CD56, CD43, CD68, and HLA-DR and the absence of other T, B, NK, or myelomonocytic markers. More importantly, we found that the bright expression of CD123 by immunohistochemistry is a distinctive characteristic of CD4+ CD56+ hematodermic neoplasms because all (n = 14) cases expressed this marker, whereas only two specimens in a control panel comprising 30 samples of related tumors expressed comparable levels of CD123. We therefore propose that oncogenic transformation of NCAM-expressing plasmacytoid monocyte-like cells may lead to "agranular CD4+ CD56+ hematodermic neoplasm." PMID- 12131154 TI - Quality assurance in immunohistochemistry: results of an interlaboratory trial involving 172 pathologists. AB - The practicability of quality assurance in immunohistochemistry and its integration into the diagnostic process were both tested in this Germany-wide interlaboratory trial. One hundred seventy-two pathologists received one hematoxylin and eosin and five unstained slides from five cases; all cases were selected by a panel because immunohistochemistry was required for their final diagnosis. Participants rendered a morphologic diagnosis and then substantiated it immunohistochemically. Stained slides and evaluation sheets were reviewed by the panel, and the diagnostic process was analyzed in individual steps: morphologic diagnosis, selection of antibodies, staining quality, interpretation of stained slides, conclusions, and final diagnosis. Diagnosis-independent immunohistochemical performance was tested using a multisample tissue block (30 samples) that was stained and evaluated for six common antigens. For individual cases, corresponding to their difficulty, 21-89% of the final diagnoses (altogether 57% from 828 diagnoses) were correct. In a statistical analysis, the tentative diagnosis, the interpretation of stains and conclusions drawn from immunohistochemistry, were independent factors in reaching the diagnosis. Sensitivity to detect estrogen receptors on the multisample tissue block was only 48%. However, 24% of the stains were interpreted as falsely negative. The low staining sensitivity was not correlated to the number of correct diagnoses. The major problem of applying immunohistochemistry in surgical pathology appears to be its integration into the diagnostic process and not the staining quality. Both future quality control projects and training will have to regard these integrative requirements. Multisample tissue blocks provide a promising tool to standardize quantitative immunohistochemical parameters, such as receptor or proliferation scores. PMID- 12131153 TI - Increased endocrine cells in treated rectal adenocarcinomas: a possible reflection of endocrine differentiation in tumor cells induced by chemotherapy and radiotherapy. AB - The presence of focal endocrine cells in colorectal adenocarcinoma is a relatively common phenomenon. However, endocrine differentiation in treated adenocarcinomas of the gastrointestinal tract has received little attention. We noted striking numbers of cells with endocrine morphology and phenotype in the residual tumor of six randomly encountered cases of rectal adenocarcinoma that were subjected to neoadjuvant therapy. All six cases had a substantial treatment response (> or =50%). To validate our initial observation and to explore its clinicopathologic significance, further morphologic and immunohistochemical studies were performed on 53 cases of rectal adenocarcinomas treated with preoperative radiation with (33 cases) or without (20 cases) chemotherapy. Pretreatment biopsies from 20 of the 53 cases and 79 resection specimens of rectal adenocarcinoma that received no neoadjuvant therapy were used as controls. Chromogranin positivity was identified in the posttreatment resection specimens in 36 of the 53 study cases (67.9%). Twenty of the 36 showed positive staining in > or =20% of the residual tumor cells. The chromogranin-positive cells in these cases often formed cords or nests. On hematoxylin and eosin sections these cells had markedly eosinophilic cytoplasm and round and uniform or sometimes pleomorphic nuclei with an often dense chromatin pattern. The proportion of chromogranin-positive cells was significantly associated with the extent of treatment response (p = 0.0005). Tumors treated with both chemotherapy and radiotherapy were more likely to have abundant chromogranin-positive cells compared with tumors treated with radiotherapy alone (p = 0.0004). In contrast, only 30% of the pretreatment biopsies and 17.7% of the control resection specimens of untreated rectal carcinomas showed chromogranin-positive cells, predominantly arranged as scattered individual positive cells, constituting <10% of the tumor. No significant correlation was observed between pretreatment and posttreatment specimens with regard to chromogranin positivity (p = 1.0). Ten of 15 patients (66.7%) whose resection specimens showed positive chromogranin staining failed to demonstrate any chromogranin positivity in their pretreatment biopsy specimens. In addition, groups or nests of chromogranin-positive cells noted in posttreatment specimens showed a very low Ki67 labeling index (<5%) but showed a frequency of abnormal p53 protein expression comparable with that observed in tumor foci resembling conventional adenocarcinoma (66.7% vs 62.5%). Our findings demonstrate that there is an increased frequency and density of cells with an endocrine phenotype in rectal adenocarcinomas that were subjected to neoadjuvant therapy and that the extent of endocrine cells appears proportional to the degree of treatment response. The possible mechanism for the increased endocrine cells in treated rectal adenocarcinomas may be related to induction of endocrine differentiation in tumor cells by cytotoxic insult. PMID- 12131155 TI - Prognostic factors for malignant melanoma of the squamous mucosa of the head and neck. AB - Primary malignant melanomas of the squamous mucosa of the head and neck are rare. To learn more about the prognostic significance of various histologic parameters we examined the pathologic features of squamous mucosa from 40 patients seen at a single institution and correlated them with clinical outcome. Follow-up information was available on 37 patients. Thirty-five were treated with surgical resection and two were treated with radiotherapy. Twenty-six were dead at follow up. Twenty-one of them died of disease. The interval between diagnosis and death ranged from 1 month to 16.5 years (median survival, 2.4 years). Eleven patients were alive at 4 months to 19.5 years after the diagnosis: six of them with disease and five of them free of disease (mean follow-up, 3.5 years). Predictors of poor survival by univariate analysis were the presence of vascular invasion (overall survival, p = 0.007; disease-specific survival, p = 0.01), a polymorphous tumor cell population (overall survival, p = 0.007; disease-specific survival, p = 0.008), and necrosis (overall survival, p = 0.007; disease-specific survival, p = 0.056). However, because these three parameters were associated with each other, none of them remained of independent predictive value for outcome by multivariate analysis. No prognostic significance was found for tumor thickness, level of invasion, ulceration, mitotic index, or nerve/nerve sheath involvement. Thus, the histologic parameters relevant for the prognosis of squamous mucosa differ significantly from those of cutaneous melanomas. PMID- 12131157 TI - CD10: a valuable tool for the light microscopic diagnosis of microvillous inclusion disease (familial microvillous atrophy). AB - Microvillous inclusion disease (MID) is a specific disorder of the intestinal brush border that leads to intractable secretory diarrhea in infants. At present, electron microscopic analysis is required for its definitive diagnosis. However, this technique is not always available or feasible, and the diagnostic microvillous inclusions may not be evident in all specimens. Accordingly, the availability of a panel of histochemical and immunohistochemical stains displaying a specific staining pattern for MID will allow pathologists to reach a definitive diagnosis of this disorder without recourse to electron microscopy. CD10 is a membrane-associated neutral peptidase, shown to have a linear brush border staining pattern in normal small intestine. We studied the staining pattern of CD10 in small intestinal biopsies from six patients with MID and in 24 control cases (10 normal small intestine, 10 celiac disease, two autoimmune enteropathy, and two allergic enteropathy). All MID cases revealed prominent cytoplasmic CD10 immunoreactivity in surface enterocytes. In contrast, all control cases showed linear brush-border staining. Similar results were obtained with periodic acid-Schiff, polyclonal carcinoembryonic antigen, and alkaline phosphatase, three stains known to show cytoplasmic staining of surface enterocytes in MID. In conclusion, CD10 is a valuable tool for the diagnosis of MID. It may be used as part of a panel that includes other stains with a distinctive staining pattern in MID such as periodic acid-Schiff, polyclonal carcinoembryonic antigen, and alkaline phosphatase. We suggest that the definitive diagnosis of MID can be reached when small bowel biopsies from infants with intractable diarrhea display cytoplasmic staining of surface enterocytes with the above-mentioned stains. PMID- 12131156 TI - Prevalence and prognostic significance of acinar cell differentiation in pancreatic endocrine tumors. AB - We have noted that many histologically and immunohistochemically confirmed pancreatic endocrine tumors show immunophenotypic evidence of acinar cell differentiation, but the clinical relevance of this finding is unknown. We performed this study to evaluate the prevalence and prognostic significance of exocrine differentiation by immunohistochemistry in pancreatic endocrine tumors that do not show morphologic features of acinar cell differentiation. Routinely processed tissue sections from 87 pancreatic endocrine tumors were immunohistochemically stained with monoclonal antibodies against acinar (lipase, chymotrypsin, trypsin) and endocrine cell markers (chromogranin A, neuron specific enolase, synaptophysin, Leu-7) and for the proliferation-associated peptide Ki67. The degree of staining with each marker was graded on a three-tier scale for acinar markers (grade 0, <5%; grade 1, 5-10%; grade 2, 11-25%; and grade 3, >25%) and on a four-tier scale for endocrine markers (grade 0, <5%; grade 1, 5-25%; grade 2, 26-50%; grade 3, 51-75%; and grade 4, >75%), and the results were correlated with clinical outcome (mean follow-up 53 months). Greater than 75% of the tumor cells stained for chromogranin A, neuron-specific enolase, synaptophysin, and Leu-7 in 100%, 96%, 93%, and 27% of cases, respectively. Overall, 66% of tumors stained positively for at least one acinar cell marker, 31% stained for at least two acinar cell markers, and 13% stained for all three acinar cell markers. Forty-seven percent stained for lipase (23 grade 1, 11 grade 2, seven grade 3), 37% for trypsin (22 grade 1, three grade 2, seven grade 3), and 25% stained for chymotrypsin (13 grade 1, five grade 2, four grade 3). No correlation was noted between the presence or extent of expression of any single or combination of acinar cell markers and clinical outcome. However, higher tumor stage correlated with a poor clinical outcome (p = 0.002), and location in the tail of the pancreas was associated with a longer interval to tumor recurrence (p = 0.03). The presence of synaptophysin (p = 0.03) and Leu-7 expression (p = 0.03) correlated significantly with less aggressive clinical behavior. An association was observed between increased Ki67 labeling and poorer clinical outcome, but this was not statistically significant (p >0.05). In conclusion, immunophenotypic evidence of acinar cell differentiation is common in pancreatic endocrine tumors, but this feature does not have any relevance to clinical prognosis. However, in addition to tumor stage, location in the pancreatic tail and the immunohistochemical expression of synaptophysin and/or Leu-7 may be useful prognostic indicators in patients with these lesions. PMID- 12131158 TI - Interobserver interpretative reproducibility of GOLDFISH, a first generation gold facilitated autometallographic bright field in situ hybridization assay for HER 2/neu amplification in invasive mammary carcinoma. AB - Clinical laboratory testing for HER-2/neu gene amplification by fluorescence in situ hybridization is not widely used in diagnostic pathology laboratories. A bright field alternative permitting direct visualization of gene amplification using conventional microscopy may be more readily incorporated into routine diagnostic pathology practice. Interobserver reproducibility represents an important component of the validation of such an assay. We tested the hypothesis that a first-generation bright field alternative to fluorescence in situ hybridization, a Nanogold (Nanoprobes, Inc, Yaphank, NY, USA) (or gold label)/autometallographic assay for HER-2/neu gene amplification in breast carcinoma, can be reproducibly interpreted by pathologists. Reference standard was direct fluorescence in situ hybridization supplemented by RNA/RNA in situ hybridization. Reproducibility of selected conventional histologic parameters was captured based on a hematoxylin and eosin slide accompanying the GOLDFISH preparation (gold-facilitated autometallographic in situ hybridization) as an indication of comparative reproducibility. The average kappa among GOLDFISH observers was 0.84, which was at least or concordant of observers scoring nuclear grade (kappa = 0.50) and the presence of in situ carcinoma (kappa = 0.57) by conventional histopathology. The GOLDFISH assay was specifically designed for qualitative interpretation, thus obviating the need for oil immersion microscopy and signal enumeration, and its interpretation was highly reproducible among five pathologists. PMID- 12131159 TI - HLA-G immunoreactivity is specific for intermediate trophoblast in gestational trophoblastic disease and can serve as a useful marker in differential diagnosis. AB - HLA-G is a nonclassical MHC class I antigen that has been shown to be a specific marker for normal intermediate trophoblast (IT). In this study HLA-G immunoreactivity assessed with an HLA-G specific antibody (4H84) was detected in all 14 cases of choriocarcinoma, 14 placental site trophoblastic tumors, 13 epithelioid trophoblastic tumors, 16 placental site nodules, and nine exaggerated placental sites. In contrast, HLA-G immunoreactivity was not detected in 34 nontrophoblastic uterine neoplasms. HLA-G immunoreactivity was present in all the IT cells of exaggerated placental sites and placental site trophoblastic tumors and in 70-100% of IT cells in placental site nodules and epithelioid trophoblastic tumors. The pattern of distribution of HLA-G in different subpopulations of IT confirms the relationship of various trophoblastic lesions to different types of IT (exaggerated placental site and placental site trophoblastic tumor to implantation site IT and placental site nodule and epithelioid trophoblastic tumor to chorionic-type IT) and suggests that choriocarcinoma is related to villous-type IT because the majority of mononucleate cells in this neoplasm were HLA-G immunoreactive. In conclusion, HLA G immunoreactivity appears to be specific for IT in gestational trophoblastic disease and can serve as a useful marker in the differential diagnosis of these lesions. PMID- 12131160 TI - Expression of alpha-Methylacyl-CoA racemase (P504S) in atypical adenomatous hyperplasia of the prostate. AB - Atypical adenomatous hyperplasia (AAH) of the prostate, also known as adenosis, is characterized by a proliferation of prostatic glands with abnormal architectural patterns, but without significant cytologic atypia. In some cases it may be difficult to distinguish AAH from prostatic carcinoma. Additionally, it is not clear whether AAH is a precursor lesion of prostatic adenocarcinoma. P504S, a protein highly expressed in prostatic adenocarcinoma, has been recently shown to be a marker of prostate cancer. The goal of this study is to examine the expression of P504S in AAH by immunohistochemistry. A total of 80 prostate specimens, including 40 cases of AAH (prostatectomy N = 30, biopsy N = 6, transurethral resection N = 4), 20 cases of prostatic adenocarcinomas, and 20 cases of benign prostatic hyperplasia, were studied. Immunohistochemistry for a prostate cancer marker alpha-methylacyl-CoA racemase (P504S) and a basal cell specific marker 34betaE12 was performed in all the cases. The 34betaE12 stain confirmed the presence of patchy basal cells in all 40 cases of AAH. P504S was undetectable in the majority of AAHs (33 of 40, 82.5%), focally expressed in four of 40 (10.0%), or diffusely positive only in three of 40 (7.5%) cases of AAH. Interestingly, two of seven P504S-positive AAHs were found adjacent to adenocarcinoma. In contrast, all benign prostatic hyperplasias (20 of 20, 100%) were negative for P504S, and all 20 cases of prostatic carcinomas (100%) showed a diffuse P504S staining pattern. These findings suggest that AAH is a heterogenous entity. The biologic significance of P504S expression in a small subset of AAH remains to be determined. Because most cases of AAH are negative for P504S, immunostaining of P504S is also of diagnostic value in distinguishing the majority of AAHs from prostatic adenocarcinoma. PMID- 12131161 TI - Alpha-Methylacyl-CoA racemase: a novel tumor marker over-expressed in several human cancers and their precursor lesions. AB - alpha-Methylacyl-CoA racemase (AMACR) is a mitochondrial and peroxisomal enzyme involved in the metabolism of branched-chain fatty acid and bile acid intermediates. Recently, AMACR has been demonstrated to be over-expressed in localized and metastatic prostate cancer, suggesting that it may be an important tumor marker. This study examines AMACR expression in a variety of human cancers and their precursor lesions. A survey of online Expressed Sequence Tags (ESTs) and Serial Analysis of Gene Expression (SAGE) databases revealed that AMACR was over-expressed in multiple cancers. The findings were confirmed by AMACR immunohistochemistry performed on several tissue microarrays containing common human tumors, including prostate, colon, and breast. Based on prior work, AMACR protein expression was divided into two categories: negative (negative to weak staining intensity) and positive (moderate to strong staining intensity). AMACR protein over-expression was found in a number of cancers, including colorectal, prostate, ovarian, breast, bladder, lung, and renal cell carcinomas, lymphoma, and melanoma. Greatest over-expression was seen in colorectal and prostate cancer with positive staining in 92% and 83% cases, respectively. AMACR over-expression was present in 44% of breast cancer cases. AMACR was also over-expressed in precursor lesions. Sixty-four percent of high-grade prostatic intraepithelial neoplasia and 75% colonic adenomas demonstrated positive AMACR protein expression. Reverse transcriptase-polymerase chain reaction for AMACR using laser capture microdissected prostate tissue confirmed gene over-expression at the mRNA level. In conclusion, our study suggests that AMACR is potentially an important tumor marker for several cancers and their precursor lesions, especially those linked to high-fat diets. PMID- 12131162 TI - COL1A1-PDGFB gene fusion demonstrates a common histogenetic origin for dermatofibrosarcoma protuberans and its granular cell variant. AB - Granular cell variant of dermatofibrosarcoma protuberans is very rare with only one report of two cases. We report a new case in which we demonstrated the presence of the dermatofibrosarcoma protuberans-specific COL1A1-PDGFB fusion from paraffin-embedded tissue. This case analysis demonstrated the utility of molecular genetics as a powerful tool for the diagnosis of atypical forms of dermatofibrosarcoma protuberans. PMID- 12131163 TI - Primary cutaneous epidermotropic alveolar rhabdomyosarcoma with t(2;13) in an elderly woman: case report and review of the literature. AB - We report a case of a primary cutaneous alveolar rhabdomyosarcoma presenting on the lower limb of a 60-year old woman. The tumor was characterized by aggregates of round blue cells in an alveolar growth pattern in the dermis and subcutis, with the additional unique finding of epidermotropism. By immunohistochemistry tumor cells were positive for vimentin, muscle-specific actin, desmin, myogenin, and Myo-D1 with focal positivity for CD56, neuron-specific enolase, and S-100 protein. Staining for pan-keratin, HMB-45, melan-A, epithelial membrane antigen, chromogranin, CD99, leukocyte common antigen, and alpha-smooth muscle actin was negative. Interphase fluorescence in situ hybridization analysis from paraffin embedded tumor demonstrated the presence of the translocation (2;13)(q35;q14) confirming the diagnosis. Further investigations revealed no tumor in the underlying deep soft tissues, and there was no evidence of metastasis in other organs. A local recurrence associated with a metastasis to a regional lymph node on the right groin was treated with an above-knee amputation and local radiotherapy to the groin area. The patient subsequently developed cutaneous metastases in the amputation stump and died 2 years after initial presentation. This case indicates that rhabdomyosarcoma may rarely present in the skin in adults and should be included in the differential diagnosis of primary cutaneous small round blue cell tumors not only in children but also in this age group. PMID- 12131164 TI - Primary paraganglioma strictly confined to the liver and mimicking hepatocellular carcinoma: an immunohistochemical and in situ hybridization study. AB - We describe a case of primary nonfunctioning paraganglioma that, unlike any other previously reported case, was strictly confined to the liver and must therefore have arisen on liver parenchyma. An asymptomatic 46-year-old man was referred to us for laparotomy and a right hemihepatectomy after a preoperative diagnosis of fibrolamellar hepatocellular carcinoma, based on a fine-needle biopsy. An 8-cm resiliently firm, pale gray nodule with a large central area of fibrosis and a thin fibrous capsule was resected. The polygonal eosinophilic tumor cells containing round nuclei lacking nucleoli were arranged in small nests set in a vascularly rich stroma. At immunohistochemistry neoplastic cells were strongly positive for chromogranin A, neuron-specific enolase, synaptophysin, and IGF-II protein; they were negative for keratin, S-100 protein, CD10, vimentin, and smooth muscle actin. In situ hybridization confirmed that, as in other sites, liver paraganglioma can express IGF-II gene. Conversely (and unlike hepatocellular carcinomas), the neoplastic cells did not express albumin mRNA, which was detected only in surrounding hepatocytes. The clinical course was benign and the patient is well and free of neoplastic disease 9 years after surgery. Knowledge of the entity should avoid possible confusion with hepatocellular carcinoma, especially of the fibrolamellar variety. PMID- 12131165 TI - Epithelial-myoepithelial carcinomas of the bronchus. PMID- 12131166 TI - Colloid carcinomas of the pancreas and periamullary region. PMID- 12131171 TI - Prediction of follow-up living setting in patients with lower limb joint replacement. AB - OBJECTIVE: The living setting to which older adults are discharged from medical rehabilitation has important social and economic implications. This study was undertaken to develop statistical models to predict living setting after medical rehabilitation in persons with lower limb joint replacement. DESIGN: Information submitted from 1994 through 1998 to the Uniform Data System for Medical Rehabilitation was examined. Hip replacement was experienced by 42% of the patients, 51% experienced a knee replacement, and 7% received some combination or other lower limb procedure. RESULTS: Persons with hip replacements were slightly older than persons with knee replacements and had a longer length of stay. Logistic regression was used to develop a predictive model based on 60% of the cases. The model included five statistically significant predictor variables. CONCLUSION: Cognitive and basic motor function associated with activities of daily living, age, length of stay, and marital status were important variables in predicting if older adults would be living at home 80-180 days after rehabilitation for lower limb joint replacement. PMID- 12131172 TI - Proprioceptive function, clinical results, and quality of life after unicondylar sledge prostheses. AB - OBJECTIVE: Evaluation of proprioceptive performance and quality of life after implantation of unicondylar sledge knee prostheses in comparison with normal control subjects. DESIGN: A total of 17 patients were examined after implantation of unicondylar sledge prostheses. Clinical examination was performed, quality of life was assessed, and proprioceptive performance was examined. A total of 11 healthy subjects of comparable age served as a control group. RESULTS: Clinical results of both groups differed significantly in all categories. Quality-of-life assessment differed only in the following items: physical functioning, role limitation caused by physical problems, and bodily pain. Other than angle reproduction at 15 degrees, no significant differences in proprioceptive testing, neither sway measurement nor angle reproduction, were found. Statistical analysis revealed no correlation between clinical scores and proprioceptive results. CONCLUSIONS: In comparison with normal control subjects of comparable age, the implantation of unicondylar sledge prostheses does not result in proprioceptive deficits. Except for physical functioning, role limitations caused by physical problems and bodily pain, most aspects of quality of life do not differ between patients and control subjects, although the patients' clinical results were significantly lower than the controls. PMID- 12131173 TI - Adrenal gland volume after spinal cord injury. AB - OBJECTIVE: Spinal cord injury in adult men may result in hypothalamic-pituitary adrenal axis dysfunction. Atrophy of adrenal glands was speculated in these patients. This study was undertaken to clarify the functional-anatomic correlation between adrenal volume and body surface area in subjects with spinal cord injury with impaired adrenal reserve. DESIGN: Twenty male subjects with chronic spinal cord injury with impaired adrenal reserve were identified by adrenocorticotropic hormone stimulation test from a group of 42 subjects. All subjects with spinal cord injury and healthy volunteers underwent computed tomographic imaging with contiguous 3-mm section over adrenal glands for volumetric measurements. Ten pairs of subjects with spinal cord injury and controls with matched height and weight were included in the statistical analysis. RESULTS: Significantly increased relative adrenal volumes were noted among subjects with chronic spinal cord injury and impaired adrenal reserve as compared with the body weight-matched and height-matched control group. CONCLUSIONS: Increased relative adrenal volumes were found after chronic spinal cord injury. Hyperplasia of the zona glomerulosa may be the cause of increased relative adrenal volume after chronic spinal cord injury. PMID- 12131174 TI - Evaluation of rehabilitation outcomes in older patients with hip fractures. AB - OBJECTIVE: This study evaluated functional outcomes in patients with hip fracture after inpatient rehabilitation. DESIGN: The physical and cognitive functioning of 100 patients with hip fracture were determined by using the FIM instrument. The Montebello rehabilitation factor score was used to reflect rehabilitative outcome. Follow-up data were collected from 44 patients by using a telephone FIM interview. RESULTS: Discharge total FIM scores improved. The Montebello rehabilitation factor score for rehabilitation efficacy and efficiency scores both demonstrated improvement for patient function during inpatient rehabilitation. The mean motor FIM domain scores for transfer mobility and locomotion were lower at discharge compared with the domains of self-care and sphincter control. A subgroup of 44 patients showed no change in mean motor FIM domain scores. CONCLUSIONS: Inpatient rehabilitation improves overall functional independence as measured by the FIM instrument. Relative change, as measured by the Montebello rehabilitation factor score, indicated that rehabilitation outcome for locomotion was not maximized, despite exhibiting large absolute gains during inpatient rehabilitation. The improvements demonstrated at discharge were maintained at follow-up for a subgroup of 44 patients. Improved locomotion skills and maximizing ability to transfer independently are areas in which inpatient rehabilitation may be targeted to improve function in the future. PMID- 12131175 TI - Healthy swing: a golf rehabilitation model. AB - OBJECTIVE: To describe a rehabilitation model using a multidisciplinary team approach for the diagnosis and treatment of individuals with golf injuries or physically challenged persons desiring to play golf. DESIGN: A retrospective, descriptive study of a multidisciplinary golf rehabilitation program that included evaluation by a physiatrist, a physical therapist, and a Professional Golf Association golf professional. RESULTS: A total of 145 individuals were treated in this program between 1994 and 1997. The majority of subjects were amateur (95%), male golfers (80%), with a mean age of 55.7 yr (range, 14-80 yr). Golfing injuries of the lower back were the most common diagnosis and had a higher frequency in men than women (49% vs. 28%); women were more likely to have shoulder (28% vs. 10%) and elbow (13% vs. 9%) injuries than men. Interventions used included medical or surgical treatment (89%), physical rehabilitation, including exercises or diathermy (92%), and modification of golf swing technique (83%). Outcomes included a return to sports participation in 98% of subjects. All subjects with golf-induced injuries returned to sports participation, and one male and one female subject won state amateur golf championships. CONCLUSION: A comprehensive, multidisciplinary model for the evaluation and rehabilitation of golf injuries has been developed, using a team of healthcare professionals and a golf teaching professional. This approach may play a role in facilitating recovery and sports participation in injured golfers. PMID- 12131176 TI - Comparison of the revised 2000 American Spinal Injury Association classification standards with the 1996 guidelines. AB - OBJECTIVE: This study was undertaken to determine the level of agreement between the most recent change in the American Spinal Injury Association International Standards (2000) and the previous (1996) classification. DESIGN: In a spinal cord injury rehabilitation hospital, data were collected on 94 subjects who had an initial neurologic examination according to the International Standards within 1 wk of injury and again at 1 yr. Comparisons were examined of the level of agreement between the 1996 and 2000 revisions in classification of the motor incomplete levels and ability to prognosticate outcome at 1 yr on the basis of the initial examination. RESULTS: Near perfect agreement between the 1996 and 2000 revised guidelines in the classification of motor incomplete injuries was found, with no statistically significant difference for prognosticating neurologic recovery at 1 yr on the basis of the initial examination. CONCLUSION: The 2000 revisions do not offer a significant difference in American Spinal Injury Association impairment classification or in predicting neurologic recovery at 1 yr. PMID- 12131177 TI - Peak flow and peak cough flow in the evaluation of expiratory muscle weakness and bulbar impairment in patients with neuromuscular disease. AB - OBJECTIVE: To study the expiratory muscle force and the ability to cough estimated by the peak expiratory flow and peak cough flow in patients with Duchenne muscular dystrophy and amyotrophic lateral sclerosis. DESIGN: A total of 27 patients with amyotrophic lateral sclerosis and 52 patients with Duchenne muscular dystrophy were studied. From the group of 144 normal subjects of this laboratory, we selected 38 for comparison. RESULTS: The maximal inspiratory pressure in patients with Duchenne muscular dystrophy and amyotrophic lateral sclerosis was 64.5 +/- 24.7% and 37.8 +/- 21.8%, respectively, and maximal expiratory pressure was 64.2 +/- 32.5% and 37.7 +/- 21.6%, respectively. Patient groups showed a significant lower peak expiratory flow than normal subjects. Higher peak cough flow than peak expiratory flow was found in all groups. The peak cough flow-peak expiratory flow difference was 46 +/- 18% in normal subjects, 43 +/- 23% in patients with Duchenne muscular dystrophy, and 11 +/- 17% in patients with amyotrophic lateral sclerosis. The peak expiratory flow and peak cough flow were not different in bulbar onset amyotrophic lateral sclerosis. In patient groups, the dynamic and static behavior correlated positively. CONCLUSIONS: These results suggest that peak cough flow-peak expiratory flow is useful to monitor expiratory muscle weakness and bulbar involvement and to assess its evolution in these patients. PMID- 12131178 TI - Effect of botulinum toxin on endplate noise in myofascial trigger spots of rabbit skeletal muscle. AB - OBJECTIVE: To assess the effect of botulinum toxin type A (BTX-A) on the endplate noise prevalence in rabbit myofascial trigger spots to confirm the role of excessive acetylcholine release on the pathogenesis of myofascial trigger points and to develop an objective indicator of the effectiveness of BTX-A in the treatment of myofascial trigger points. DESIGN: Eighteen adult New Zealand rabbits were divided into three groups that received a single bolus of BTX-A over a myofascial trigger spot region on one side of the biceps femoris muscle. Another 10 rabbits received multiple-point injections in a myofascial trigger spot where endplate noises were found. A control study was performed on the other side of the biceps femoris muscle. The endplate noise prevalence in a myofascial trigger spot region was assessed. RESULTS: It was found that injection of BTX-A reduced the prevalence of endplate noise. No significant differences between a single bolus injection and multiple-point injections were noted, although there was some evidence that multiple-point injections might maintain the endplate noise decreasing effect much longer than a single injection. CONCLUSIONS: This study demonstrated the suppressive effect of BTX-A on endplate noise prevalence in a myofascial trigger spot region. The prevalence of endplate noise in the myofascial trigger point region may be a useful objective indicator for evaluating the therapeutic effectiveness of BTX-A injection to treat myofascial trigger points. PMID- 12131180 TI - Comparing the effects of different speech targets on cognitive event-related potentials: theoretical implications for evaluating brain injury. PMID- 12131181 TI - Abstracts of scientific papers and posters presented at the Annual Meeting of the Association of Academic Physiatrists. Las Vegas, Nevada, USA. February 28-March 2, 2002. PMID- 12131182 TI - AIDS in Mexico: lessons learned and implications for developing countries. PMID- 12131183 TI - T cell receptor V beta repertoire of the antigen specific CD8 T lymphocyte subset of HIV infected children. AB - OBJECTIVE: Analysis of the T cell receptor V beta repertoire during HIV infection reveals expansions in multiple V beta families of CD8 T cells, but their antigenic specificity is ill-defined. We sought to determine the TCR V beta repertoire of HIV specific CD8 T lymphocytes in infected children. DESIGN/METHODS: We performed flow cytometry to examine TCR V beta families as identified by specific monoclonal antibodies and binding of HIV peptide loaded tetrameric MHC complexes in peripheral blood samples from a group of HIV infected children. RESULTS: Simultaneous assessment of 12 selected expanded V beta families amongst nine HIV infected patients for tetramer binding revealed only one child in whom the expanded V beta population bound HIV Gag or Pol tetramers. In four HIV infected children, percentage tetramer binding cells was determined in 21 TCR V beta families. The tetramer binding cells of three children exhibited a widely distributed TCR V beta repertoire while in the fourth patient they were preferentially localized within two TCR V beta families. Repeat analysis revealed that the TCR V beta repertoire of tetramer binding cells was stable. CONCLUSIONS: These data provide evidence that the HIV-specific CD8 T cell response in children is usually distributed widely among many different TCR V beta families. The heterogeneity of the TCR V beta repertoire usage by the antigen specific CD8 T cells may reflect the dynamic interaction between host and pathogen during the course of HIV infection and may be influenced by the rate of viral mutation, CD4 T cell helper activity, or other factors. PMID- 12131184 TI - CCR5 mediates Fas- and caspase-8 dependent apoptosis of both uninfected and HIV infected primary human CD4 T cells. AB - DESIGN: HIV Env interaction with the corresponding chemokine receptor dictates the molecular mechanism of death of both HIV-infected and uninfected primary CD4 T cells. CXCR4/T tropic HIV virus (X4) triggers CD4 T cell death through a caspase independent mechanism, whereas CCR5/M tropic HIV virus (R5) HIV triggers a caspase dependent death. In the present study, we have investigated the pathway whereby R5 Env-CR5 interactions lead to a caspase dependent cell death. METHODS: CD4 T cells were infected with X4 or R5 HIV strains, or were mock infected. After infection, cells were treated with caspase inhibitors or decoys of death receptor signaling pathways and cell viability was analyzed. The role of R5 HIV Env in induction of cell death of uninfected T cells was analyzed by co-culturing uninfected CD4 T cells with R5 Env expressing cells in the absence or presence of various inhibitors of death receptor signaling. RESULTS: Infection of CD4 T cells with R5, but not with X4 HIV strains results in the activation of caspase-8 and cell death that is reversed by a decoy of the Fas receptor. Isolated activation of CCR5 by membrane-bound, or soluble R5 Env causes a Fas- and caspase-8 dependent death also of uninfected CD4 T cells. Additional studies demonstrate that isolated CCR5 activation by R5 Env leads to both de novo expression of FasL and induction of susceptibility to Fas-mediated apoptosis in resting primary CD4 T cells. CONCLUSIONS: These results ascribe to CCR5 a novel role in activating the Fas pathway and caspase-8 as well as triggering FasL production when activated by R5 Env, ultimately causing CD4 T cell death. PMID- 12131185 TI - Effects of interleukin-2 plus highly active antiretroviral therapy on HIV-1 replication and proviral DNA (COSMIC trial). AB - BACKGROUND: The effect of interleukin-2 (IL-2) in combination with antiretroviral therapy on HIV-1 replication and reservoirs was investigated. METHODS: In a prospective, open-label trial, 56 asymptomatic HIV-1-infected subjects (CD4 T cell count > 350 x 10(6) cells/l) were randomized to highly active antiretroviral therapy (HAART: stavudine, lamivudine, nelfinavir, saquinavir) with or without IL 2 (9 megaunits daily for 5 days in 6-weekly intervals for a total of eight cycles). Productive and latent infection were analysed in peripheral blood, and residual virus replication in the lymphoid tissue and in the cerebrospinal fluid. The influence of IL-2 on viral rebound after treatment discontinuation was studied. RESULTS: Virus replication was detected in 21 of 31 on-treatment lymph nodes despite undetectable plasma viraemia. Viral RNA was found in resting as well as in proliferating cells. RNA-negative patients tended towards more rapid proviral DNA elimination. Supplementary IL-2 led to a greater increase in CD4 T cell counts than HAART alone (P < 0.001), resulting in normalization in approximately 90% of IL-2-treated patients compared with approximately 50% HAART only subjects. IL-2 had no beneficial effect on virus replication and on proviral DNA in peripheral blood. CONCLUSIONS: Viral persistence during HAART is partly a result of continued low-level replication, calling for more active regimens. IL-2 accelerates the normalization of CD4 T cell counts but does not impact on virus production or latency. PMID- 12131187 TI - The fraction of perforin-expressing HIV-specific CD8 T cells is a marker for disease progression in HIV infection. AB - OBJECTIVE: Perforin is an important component of the death machinery of cytotoxic T cells (CTL). To evaluate functional differences between HIV- and cytomegalovirus (CMV)-specific CTL of coinfected patients, the frequencies of the respective perforin-expressing T cells were analysed in a rapid whole blood assay. METHODS: Whole blood of HIV- and CMV-infected individuals was specifically stimulated by HIV-1 Pr55(gag) or complete CMV antigen, and activation-induced intracellular cytokine and perforin expression in CD8 T cells was analysed by flow cytometry. RESULTS: Perforin-expressing HIV-1- and CMV-specific CD8 T cells can be quantified simultaneously. Within a patient, the frequency of such HIV specific CD8 T cells in peripheral blood was lower than the frequency of the respective CMV-specific cells. The number of the perforin-expressing HIV-specific CD8 T cells inversely correlated with the peripheral blood CD4 T cell count. CONCLUSIONS: The differential fractions of perforin-expressing virus-specific CD8 T cells in HIV and CMV double infection might be caused by differences in priming and trafficking to or from replication sites. However, without knowing the underlying mechanism, the fraction of perforin-expressing HIV-specific CD8 T cells provides another surrogate marker for disease progression. PMID- 12131186 TI - SIV(mac) pathogenesis in rhesus macaques of Chinese and Indian origin compared with primary HIV infections in humans. AB - OBJECTIVE: To develop a SIV-rhesus macaque (Rh) model of AIDS that more closely approximates HIV pathogenesis in humans. DESIGN: The pathogenesis of SIV was compared in two different types of Rh, the Chinese (Ch) and Indian (Ind) subspecies. METHODS: Ch Rh and Ind Rh origin were identified genetically and infected with the SIV(mac)239 molecular clone. Plasma viral loads, depletion of intestinal lymphocytes with memory phenotype, humoral immune responses and CD4/CD8 cell ratios were compared during acute and steady-state periods of infection. RESULTS: Plasma viral loads from 7 days after infection through 240 days were significantly lower in Rh of Ch origin compared with Ind Rh. Viral loads in Ch Rh were closer to viral loads observed in untreated humans infected with HIV-1. Depletion of intestinal effector cells was less evident in SIV infected Ch Rh compared with Ind Rh. An index of intestinal pathogenesis was devised that closely paralleled viral load and severity of infection. There were no rapid progressors to AIDS among 10 Ch Rh. In contrast, three of four Ind Rh progressed rapidly to AIDS. CONCLUSIONS: Compared with Ind Rh, SIV(mac) pathogenesis in Ch Rh was closer to HIV-1 infections in untreated adult humans. The differences were statistically significant. The Ch Rh subspecies is a suitable AIDS model and may have advantages over the rapid and highly pathogenic Ind Rh model. Moreover, Ind Rh supplies are limited and use of Ch Rh provides a new resource. PMID- 12131188 TI - Immune activation and induction of HIV-1 replication within CD14 macrophages during acute Plasmodium falciparum malaria coinfection. AB - OBJECTIVES: To determine the impact of Plasmodium falciparum malaria coinfection and its treatment on cellular reservoirs of viral replication in HIV-1-infected persons and to relate this to changes in systemic immune activation. METHODS: Plasma samples were obtained from HIV-1-infected individuals (n = 10) at diagnosis of acute malaria, 4 weeks after parasite clearance and from HIV infected aparasitemic controls (n = 10). Immunomagnetic HIV-1 capture analysis was used to determine the cellular origin of cell-free virus particles present in all 30 plasma samples and indices of immune activation were measured using enzyme linked immunosorbent assays. RESULTS: Compared with controls, the detectable proportion of HIV-1 particles derived from CD14 macrophages and CD26 lymphocytes was increased in persons with acute malaria coinfection and correlated with markedly increased plasma concentrations of both proinflammatory cytokines and soluble markers of macrophage and lymphocyte activation. Parasite clearance following treatment with antimalarial drugs resulted in decreased detection of HIV-1 particles derived from the CD14 macrophage cell subset and correlated with a marked diminution in systemic immune activation. CONCLUSIONS: Acute P. falciparum malaria coinfection impacts virus-host dynamics in HIV-1-infected persons at the cellular level, notably showing a reversible induction of HIV-1 replication in CD14 macrophages that is associated with changes in immune activation. PMID- 12131189 TI - Evolving patterns of HIV-1 resistance to antiretroviral agents in newly infected individuals. AB - OBJECTIVE: To assess temporal changes in prevalence of transmitted HIV-1 drug resistance in a homogeneous cohort of newly infected individuals. METHODS: Pretreatment genotypic and phenotypic drug resistance was tested in 154 subjects with primary HIV-1 infection identified between 1995 and 2001 (group A; n = 76) and 1999 and 2001 (group B; n = 78). Sequence analysis was assessed by population based sequencing. Virus susceptibility to antiretroviral agents was determined by the PhenoSense assay (ViroLogic). RESULTS: The frequency of resistance-associated mutations in protease (PR) and reverse transcriptase (RT) genes increased from 13.2% (1995-1998) to 19.7% (1999-2001). Although the overall prevalence of viruses with phenotypic resistance did not vary (1995-1998, 10.0%; 1999-2001, 10.8%), the distribution of drug classes changed [nucleoside RT inhibitor (NRTI): 8.3% to 2.7%; non-NRTI: 5.0% to 8.1%; protease inhibitors (PI): 1.7% to 5.4%]. The decrease of phenotypic resistance to NRTI in 1999-2001 was caused by the absence of transmitted lamivudine-resistant variants. Phenotypically susceptible variants with aspartic acid or serine residues at position 215 of RT (5.3%; P = 0.04) instead emerged. Hypersusceptibility to PI decreased from 18.3% to 5.4% (P = 0.02) while the amino acid substitutions in PR increased over time: M36I (6.6% to 19.7%) and A71V/T (3.9% to 15.8%). CONCLUSIONS: There was an increase in the number of HIV-1 variants with both genotypic and phenotypic resistance to non NRTI and PI over time. Furthermore, viruses with altered genotypes compatible with thymidine analogue or PI exposure but susceptible phenotypes were seen in 1999-2001. The latter findings suggest transmission of viruses from subjects who have either changed or discontinued therapy. PMID- 12131190 TI - The natural history and clinical significance of intermittent viraemia in patients with initial viral suppression to < 400 copies/ml. AB - OBJECTIVES: To determine the prevalence and prognostic significance of intermittent viraemia (IV) in patients who attained an undetectable viral load (VL) < 400 copies/ml within 6 months on highly active antiretroviral therapy (HAART). METHODS: Retrospective analysis of viral load rebound > or = 400 copies/ml and CD4 cell counts rise for 765 patients followed for > or = 12 months following initial VL undetectability, comparing the 226 (29.5%) who maintained an undetectable VL for > 1 year from initiation of HAART and 122 (15.9%) who had one or more episodes of IV. Genotypic resistance was evaluated at the time of the first episode of IV > or = 2000 copies/ml. RESULTS: Patients with IV had a threefold higher rate of sustained virological rebound [hazards ratio (HR), 3.15; 95% confidence interval (CI), 1.72-5.77; P < 0.001). For patients with and without IV, the Kaplan-Meier estimates at 24 and 36 months after initiation of HAART were 19.3% (95% CI, 8.9-21.5) versus 7.7% (95% CI, 4.5-13.0) and 31.6% (95% CI, 21.8-44.2) versus 12.9% (95% CI, 7.5-21.5), respectively (P < 0.001). The median CD4 cell count rise at 18 and 24 months was significantly lower in those with IV than in those without: 138 [interquartile range (IQR), 58-221] versus 224 x 10(6) cells/l (IQR, 119-357) (P = 0.0001) and 200 (IQR, 89-294) versus 260 x 10(6) cells/l (IQR, 125-384) (P = 0.003), respectively. In a subgroup of 16 patients, genotypic resistance mutations were found in the reverse transcriptase gene for five (31%) and in the protease gene in one. A probable contributing factor/event was identified for most patients with IV, such as poor adherence (42.6%), intercurrent infection (26.2%) or drug interaction (6.8%). CONCLUSIONS: Patients with IV > 400 copies/ml are three times more likely to experience sustained viral rebound and to have an impaired CD4 cell rise relative to those who maintain undetectable VL. This supports the adoption of a more pro-active approach to treatment intensification and the need for caution with structured treatment interruptions. PMID- 12131191 TI - Sexual risk behaviors and implications for secondary HIV transmission during and after HIV seroconversion. AB - OBJECTIVES: To determine the potential for secondary HIV transmission among newly HIV-infected men who have sex with men (MSM) during their HIV antibody seroconversion period, and for the 12 months after seroconversion. DESIGN: A cohort study. METHODS: Risk assessment questionnaires administered before receipt of the first positive HIV antibody result, plasma and seminal viral load measurements, and risk assessments one month and quarterly after receipt of the first HIV-positive test, and generalized estimating equation modelling techniques to analyse behavioral trends. RESULTS: Of 66 seroconverters, more than half reported unprotected anal intercourse (UAI) with HIV-negative or unknown serostatus partners during seroconversion, with 27% reporting insertive UAI with an HIV-negative partner. The initial median plasma viral load was 4.6 log/ml, the median seminal viral load was 2.7 log/ml, suggesting a high level of infectiousness. Compared with risk behavior during seroconversion, UAI with HIV negative or unknown-serostatus partners was reduced after the receipt of positive antibody results; however, a substantial proportion of participants reported high risk behaviors for transmission for 12 months of follow-up. After learning of their HIV infection, recent seroconverters did not reduce the risk of secondary transmission by engaging in proportionally more high-risk practices with HIV infected partners (compared with HIV-negative or unknown-serostatus partners), or engaging in proportionally more receptive compared with insertive UAI. CONCLUSION: Substantial potential exists for secondary HIV transmission during and for one year after HIV seroconversion. Receipt of an HIV-positive test is associated with a significant reduction in risk behavior, reinforcing the need to identify and counsel recently HIV-infected MSM. PMID- 12131192 TI - High-risk sexual behaviour increases among London gay men between 1998 and 2001: what is the role of HIV optimism? AB - OBJECTIVE: To examine whether HIV optimism (i.e. optimism in the light of new HIV drug therapies) can account for the recent increase in high-risk sexual behaviour among London gay men. METHODS: Gay men (n = 2938) using London gyms were surveyed annually between 1998 and 2001. Information was collected on HIV status, unprotected anal intercourse (UAI) in the previous 3 months, and agreement with two statements concerning the severity of and susceptibility to HIV infection. Those who agreed were classified as 'optimistic'. RESULTS: Between 1998 and 2001, the percentage of men reporting high-risk UAI (i.e. UAI with a casual partner of unknown or discordant HIV status) increased: HIV-positive men 15.3-38.8%; HIV negative men 6.8-12.1%; never-tested men 2.1-7.7%; (P < 0.01). Overall, less than a third were optimistic. In cross-sectional analysis, optimistic HIV-positive and -negative men were more likely to report high-risk UAI than other men (P < 0.05). However, the increase in high-risk UAI between 1998 and 2001 was seen in those who were optimistic and those who were not (P < 0.05). In multivariate analysis, the modelled increase in high-risk UAI over time remained significant after controlling for HIV optimism (P < 0.01), with no significant interaction between optimism and time. CONCLUSION: Among London gay men, no difference was detected between those who were optimistic and those who were not in the rate of increase in high-risk sexual behaviour between 1998 and 2001. Our findings suggest that HIV optimism is unlikely to explain the recent increase in high-risk sexual behaviour in these men. PMID- 12131193 TI - Long-term follow-up of HIV-positive and HIV-negative individuals in rural Malawi. AB - OBJECTIVE: To measure the effect of HIV on survival in rural Africa. DESIGN: A retrospective cohort study with more than 10 years follow-up. METHODS: Individuals with known HIV status in the 1980s were identified from previous population surveys in Karonga District, northern Malawi. Follow-up studies were conducted in 1998-2000 to trace 197 HIV-positive and 396 age-sex-matched HIV negative individuals and their spouses. RESULTS: Information was obtained on all but 11 index individuals. Half (302) were found and the others were reported to have died (161) or to be alive outside the district (119). Ten year survival was 36% in the HIV-positive cohort and 90% in the initially HIV-negative cohort. The death rate was 93.3 per 1000 person-years in the HIV-positive individuals, and 11.3 in the initially HIV-negative individuals. Survival time since the initial test in HIV-positive individuals decreased with age, but relative survival, compared with HIV-negative individuals, was similar across age groups. The effect of HIV on survival was similar in men and women. Spouses of HIV-positive individuals had four times the mortality rate, and among survivors, four times the HIV prevalence, of spouses of initially HIV-negative individuals. CONCLUSION: HIV-infected individuals had very high mortality rates, but one-third were still alive at 10 years. This is consistent with median survival from seroconversion being similar to that found in developed countries before antiretroviral therapy. Mortality rates in HIV-positive individuals increased with age, but relative mortality changed little with age. PMID- 12131195 TI - High effectiveness of efavirenz-based highly active antiretroviral therapy in HIV 1-infected patients with fewer than 100 CD4 cells/microl and opportunistic diseases: the EfaVIP Study (Efavirenz in Very Immunosuppressed Patients). AB - We evaluated the therapeutic outcomes of all antiretroviral-naive HIV-1-infected patients with fewer than 100 CD4 cells/microl, who received efavirenz-based highly active antiretroviral therapy (HAART). Sixty-one percent suffered AIDS defining diseases, and after a median follow-up of 45 weeks there were three deaths and five AIDS-related conditions (two relapses, three new). Efavirenz based HAART was found to be effective in profoundly immunosuppressed HIV-1 infected patients. PMID- 12131194 TI - Kinetics of CD4 cells after discontinuation of antiretroviral therapy in patients with virological failure and a CD4 cell count greater than 500 cells/microl. AB - After a median of 37 months on antiretroviral therapy, 16 patients were asked to discontinue treatment instead of changing it. After a median observation time of 10.5 months, most patients experienced a rapid and progressive decrease in their CD4 cell count, even without a high viral load rebound. This decline was unrelated to the CD4 cell count and HIV-RNA values at interruption, but was more profound in patients in whom the M184V mutation had disappeared after lamivudine discontinuation. PMID- 12131196 TI - Effects of co-infection with hepatitis C virus and GB virus C on CD4 cell count and HIV-RNA level among HIV-infected patients treated with highly active antiretroviral therapy. AB - The effects of co-infection with hepatitis C virus (HCV) and GB virus C (GBV-C) on CD4 cell counts and plasma HIV-RNA levels has been investigated in HIV infected patients treated with highly active antiretroviral therapy (HAART). Patients co-infected with HCV and GBV-C experienced a CD4 cell increase during 4 years of HAART, whereas the increase stopped after 2 years in the other groups. PMID- 12131197 TI - Angioedema and transient acquired C1 inhibitor functional deficiency in HIV infection: case report. PMID- 12131198 TI - Acute Guillain-Barre syndrome during the chronic phase of HIV infection and dramatic improvement under highly active antiretroviral therapy. PMID- 12131199 TI - Differential induction of P-glycoprotein and MRP by rifamycins in T lymphocytes from HIV-1/tuberculosis co-infected patients. PMID- 12131200 TI - Ritonavir-associated hyperparathyroidism, osteopenia and bone pain. PMID- 12131201 TI - Female sex but not ethnicity is a strong predictor of non-nucleoside reverse transcriptase inhibitor-induced rash. PMID- 12131202 TI - Sildenafil as a successful treatment of otherwise fatal HIV-related pulmonary hypertension. PMID- 12131203 TI - Treatment of tuberculosis in HIV-infected individuals. PMID- 12131205 TI - Does highly active antiretroviral therapy induce sickle cell crises? PMID- 12131207 TI - Prophylaxis of opportunistic infections in HIV-infected adults in sub-Saharan Africa: opportunities and obstacles. PMID- 12131206 TI - HIV incidence on the increase among homosexual men attending an Amsterdam sexually transmitted disease clinic: using a novel approach for detecting recent infections. AB - OBJECTIVE: Dramatic increases have occurred in sexually transmitted diseases (STD) and in sexual risk behaviour among homosexual men in Amsterdam and internationally. We investigated whether these trends indicate a resurgence of the HIV epidemic. METHODS: HIV incidence was determined among homosexual attendees of an STD clinic in Amsterdam, who had participated in semi-annual anonymous unlinked cross-sectional HIV prevalence studies from 1991 to 2001. Stored HIV-seropositive samples were tested with a less-sensitive HIV assay and, if non-reactive, were further tested for the presence of antiretroviral drugs, indicative of the use of highly active antiretroviral therapy. Seropositive men who tested non-reactive on the less-sensitive assay and had not used antiretroviral drugs were classified as recently infected (< 170 days). Annual HIV incidence and its changes were examined. RESULTS: Among 3090 homosexual participants (median age 34 years), 454 were HIV infected, of whom 37 were recently infectioned. From 1991 to 2001 the overall incidence was 3.0 infections/100 person-years. Incidence increased over time (P = 0.02) and, strikingly, the increase was evident in older (> or = 34 years) men (P < 0.01), but not in the young. Of men recently infected, 84% (n = 31) were unaware of their infection and 70.3% (n = 26) had a concurrent STD. These 26 men reportedly had sex with a total of 315 men in the preceding 6 months. CONCLUSION: HIV incidence is increasing among homosexual attendees of an STD clinic. It is imperative to trace recently infected individuals, because they are highly infectious, and can thus play a key role in the spread of HIV. PMID- 12131208 TI - Activation of HIV-1 specific CD4 and CD8 T cells by human dendritic cells: roles for cross-presentation and non-infectious HIV-1 virus. AB - BACKGROUND: The CD4 T cells in mucosal subepithelia are the first cells to become infected during sexual transmission of HIV-1. Dendritic cells (DC) are located in the same area and are known to play a central role in antiviral immune responses. However, extensive viral replication, syncytia formation and cell death follows the interaction between T cells and DC previously exposed to HIV-1. Despite this, anti-HIV responses are generated that control viremia following acute infection. OBJECTIVE: The anti-HIV-1 cellular immune responses observed may be activated by sources other than productively infected DC. HIV-1 induces apoptosis both in cells it infects and in bystander cells. Furthermore, retroviral replication typically generates a predominance of defective particles. We tested whether DC exposed to antigen from either of these sources could elicit anti-HIV specific immune responses. DESIGN AND METHODS: Apoptotic or necrotic monocytes infected with vaccinia virus vectors encoding HIV antigens, a cell line with integrated HIV-1 and apoptotic CD4 T cells pulsed with non-infectious or infectious HIV-1 virus were used as sources of antigens to assess cross presentation by DC. Furthermore, direct DC presentation of antigen from non-infectious and infectious HIV-1 was examined. RESULTS: We find that dead cells expressing HIV-1 antigens as well as non-infectious HIV-1 particles can be acquired and processed by DC, leading to the activation, differentiation and expansion of viral antigen specific CD4 and CD8 T cells from seropositive individuals. CONCLUSIONS: These sources of antigens may be critical for the generation and maintenance of anti HIV-1 immunity by DC. PMID- 12131209 TI - Virologic response to nelfinavir-based regimens: pharmacokinetics and drug resistance mutations (VIRAPHAR study). AB - OBJECTIVE: To assess the impact of HIV-1 protease and reverse transcriptase (RT) mutations, and pharmacokinetic parameters on virological responses to nelfinavir (NFV)-containing highly active antiretroviral therapy. DESIGN: Naive or antiretroviral-experienced HIV-1-infected subjects were included in a non randomized, observational cohort study and received two nucleoside RT inhibitors + NFV (750 mg three times per day or 1250 mg twice per day). Virologic success was defined as a virus load < 50 copies/ml for > 6 months. METHODS: RT and protease genes were sequenced at baseline and at the time of virological failure. Plasma NFV trough concentration (Cmin), maximum concentration (Cmax), and AUC0 tau at steady-state were subjected to population pharmacokinetic analysis. RESULTS: Patients (n = 154) enrolled between November 1998 and February 2000 started a twice per day (n = 84) or three times per day (n = 70) NFV-based regimen as first- (n = 48) or second-line therapy when protease inhibitor-naive (n = 64) or -experienced (n = 42). Median follow-up duration was 16 months. Virologic failure occurred in 88 patients. No significant differences were observed between twice per day and three times per day regimens. According to multivariate analysis, NFV Cmin and Cmax, CD4 cell count, number of baseline RT + protease gene mutations, D67N, M184V, T215F/Y in RT, and M36I in protease, were independent factors that were significantly predictive of failure. At failure, L10I, D30N, M36I, V77I, N88S/D or L90M protease mutations had emerged since baseline. Pharmacokinetic parameters were similar in patients with or without emergence of these neo-mutations. The more discriminating NFV Cmin efficacy threshold was estimated to be 1 mg/l. CONCLUSIONS: Our data confirm the association among individual pharmacokinetic parameters, genotype pattern and virological response to NFV-containing regimens. PMID- 12131210 TI - Hyperlactataemia and lactic acidosis during antiretroviral therapy: relevance, reproducibility and possible risk factors. AB - OBJECTIVE: To evaluate the prevalence, outcome and possible risk factors for hyperlactataemia and lactic acidosis in HIV-positive persons receiving antiretroviral therapy. METHODS: Cross-sectional and longitudinal data from a prospectively collected clinical database. Associations with antiretroviral regimen, clinical and laboratory parameters were assessed using univariate and multivariate Cox's proportional hazards model. RESULTS: Patients naive to therapy and patients on current therapy for a minimum of 4 months were assessed. Median lactate was 1.1 mol/l in 253 untreated individuals and 1.4 mmol/l in 1239 patients stable on therapy for at least 4 months. At least two on-therapy samples were available for 750 of the 1239 individuals, taken a median 92 days apart. Lactate measurement showed a low positive predictive value of 38.9% but a high negative predictive value (98%) for normal values. Lactate was elevated > or = 2.4 mmol/l in 102 individuals on at least one occasion. In the multivariate Cox's proportional hazards model, no demographic characteristics were associated with hyperlactataemia. Didanosine-containing regimens doubled the relative hazard of hyperlactataemia compared with those sparing didanosine. Abacavir-containing regimens reduced the hazard of hyperlactataemia. Choice of thymidine analogue did not influence risk. Hyperlactataemia was associated with acid-base disturbance. Use of didanosine and female sex were over-represented amongst nine patients with severe hyperlactataemia (> 5 mmol/l) or lactic acidosis. CONCLUSIONS: Screening of lactate is of limited use in asymptomatic individuals on antiretroviral therapy. Raised lactate represents part of a spectrum of lactate and acid-base disturbance that infrequently includes lactic acidosis. Didanosine appears associated with an increased risk of hyperlactataemia. PMID- 12131211 TI - The microbicide cyanovirin-N expressed on the surface of commensal bacterium Streptococcus gordonii captures HIV-1. AB - OBJECTIVE: To explore the feasibility of expressing the potent HIV-inactivating protein, cyanovirin-N (CV-N), in the human commensal bacterium Streptococcus gordonii, as a possible approach for local delivery of CV-N to prevent sexual transmission of HIV-1. DESIGN AND METHODS: To express CV-N in S. gordonii, we used the host-vector system we had previously developed. CV-N was expressed as a fusion protein both attached to the bacterial surface and secreted in soluble form in the supernatant of liquid cultures. The soluble form of recombinant CV-N was tested for gp120-binding activity in an enzyme-linked immunosorbent assay, whereas S. gordonii strain expressing CV-N on the surface was analyzed in an in vitro HIV capturing assay. RESULTS: Two recombinant S. gordonii strains secreting or displaying CV-N on the bacterial surface were constructed and the expression of CV-N was confirmed by immunoblot and flow-cytometric analysis. The secreted form of recombinant CV-N exhibited a concentration-dependent binding to the envelope glycoprotein gp120 of HIV-1, whereas CV-N displayed on the bacterial surface was able to capture HIV virions efficiently. CONCLUSION: The anti-HIV protein CV-N in S. gordonii was expressed in a biologically active form. This represents a first step in the development of a system to deliver and maintain an effective concentration of a microbicide in the vaginal mucosa. PMID- 12131212 TI - Is hepatitis C virus co-infection associated with survival in HIV-infected patients treated by combination antiretroviral therapy? AB - OBJECTIVE: To study whether hepatitis C virus (HCV) co-infection or the severe elevation of transaminases is associated with survival after the initiation of antiretroviral combination therapy. DESIGN: Prospective hospital-based cohort (Aquitaine Cohort). METHODS: HIV-infected adults started on an antiretroviral combination before 30 June 1999. HCV infection was defined as antibody detection or positive HCV RNA. Severe elevation of transaminases was defined as a value of aspartate or alanine aminotransferase (AST, ALT) above five times the upper limit of normal values. Survival was studied using a Cox model, including at least baseline HCV status and transaminases as a time-dependent covariate. RESULTS: Overall, 995 patients were analysed, including 576 HCV-positive individuals (58%). At baseline, HCV-positive patients were younger, more often injecting drug users and women, and had more frequently elevated transaminases. A shorter survival was associated with AIDS stage [hazard ratio (HR) versus non-AIDS 1.67; 95% confidence interval (CI) 1.03; 2.68], lower CD4 cell count (HR for 50 cells/mm3 lower 1.33; CI 1.17; 1.51), lower haemoglobin (HR for 1 g/dl lower 1.20; CI 1.07; 1.35), lower platelet count (HR for 10 000 cells/mm3 lower 1.04; CI 1.01; 1.07), and AST during follow-up (HR for > or = 200 IU/l 2.30; CI 1.32; 4.03). HCV co-infection (HR 1.20; CI 0.75; 1.92) was not statistically associated with survival. CONCLUSION: The occurrence of a severe elevation of transaminases was associated with poorer survival, although HCV was not. If liver toxicity may be treatment induced, plasma drug concentrations could guide dosage adjustments of antiretroviral treatments currently prescribed to optimize their use. PMID- 12131213 TI - The Senegalese government's highly active antiretroviral therapy initiative: an 18-month follow-up study. AB - OBJECTIVE: To study the feasibility, effectiveness, adherence, toxicity and viral resistance in an African government HAART initiative. METHODS: A prospective observational cohort study started in Dakar in August 1998. Initial treatment consisted of two nucleoside reverse transcriptase inhibitors and one protease inhibitor. The patients attended monthly medical examinations. Plasma HIV-1 RNA and CD4 cell counts were determined at baseline and every 6 months. Intention-to treat analyses were performed. RESULTS: Fifty-eight treatment-naive patients, mostly infected by HIV-1 strain CRF02-AG, were enrolled. Most were at an advanced stage of HIV disease (86.2% had AIDS). Adherence was good in 87.9% of patients and treatment was effective in most of them. Thus, HIV-1 RNA was undetectable in 79.6, 71.2, 51.4 and 59.3% of patients at months 1, 6, 12 and 18, respectively and the median viral load reduction was approximately 2.5 log10 copies/ml. The CD4 cell count rose by a median of 82, 147 and 180 x 106 cells/l at months 6, 12 and 18, respectively. At the same time points, the cumulative probability of remaining alive or free of new AIDS-defining events was 94.8, 85.0 and 82.3%. Most adverse effects (80.8%) were mild or moderate and only two cases of drug resistance occurred. CONCLUSION: This study shows that HAART is feasible and well tolerated in African patients. Clinical and biological results were comparable to those seen in western cohorts, despite differences in the HIV-1 subtype distribution and an advanced disease stage when the treatment was initiated. Contrary to other recent studies in Africa, viral resistance rarely emerged. PMID- 12131214 TI - Clinical efficacy of early initiation of HAART in patients with asymptomatic HIV infection and CD4 cell count > 350 x 10(6) /l. AB - OBJECTIVE: To evaluate the efficacy of early initiation of highly active antiretroviral therapy (HAART), we compared the clinical course of two nested, matched cohorts within the Swiss HIV Cohort Study. METHODS: We selected all asymptomatic patients who started HAART between 1 January 1996 and 31 December 1999 with a CD4 cell count > 350 x 10(6)/l. We then matched them with asymptomatic participants who were seen at around the same time and who remained untreated during the following 12 months. This control group was further matched for age, sex, CD4 cell count, viral load, and HIV risk category, generating 283 pairs of treated versus untreated patients. RESULTS: During observation of median 3.19 versus 2.66 years, CDC stage B/C occurred in 6.4% versus 21.2%, AIDS in 1.8% versus 5.3%, death in 2.1% versus 6.4%, and AIDS or death of 'natural' causes in 2.8% versus 6.7% of the treated versus untreated patients. In multivariable Cox regression analysis, treatment reduced the risk of clinical progression by a factor of four- to five fold. During follow-up, the treated group had significantly higher CD4 counts and lower HIV-1 RNA levels. Intolerance/adverse events led to change or stop of at least one drug in 35% of treated patients. The entire regimen was interrupted at least once by 41% of patients, and 24% had no treatment anymore at the end of follow-up. CONCLUSIONS: The initiation of HAART in asymptomatic patients with CD4 cell count > 350 x 10(6)/l significantly delayed clinical progression. However, the risk of severe clinical events with deferred therapy was low and must be counter balanced against the burden and toxicity of HAART. PMID- 12131215 TI - Reversal of atherogenic lipoprotein profile in HIV-1 infected patients with lipodystrophy after replacing protease inhibitors by nevirapine. AB - BACKGROUND: The widespread use of protease inhibitors (PI) has been associated with abnormalities in the lipid profile of HIV-1-infected patients. Treatment simplification approaches in which PI are replaced by nevirapine (NVP) have been shown to improve PI-related toxicity. OBJECTIVE: To assess the impact on plasma lipids of replacing the PI by NVP in HIV-1 infected patients with lipodystrophy. METHODS: We studied 34 patients with lipodystrophy who had been the first to be enrolled in a prospective, randomized trial of continuing current treatment, or replacing PI with NVP. Sixteen patients replaced their PI with NVP and 18 continued their current PI-containing treatment. Total, low density lipoprotein (LDL), very low density lipoprotein (VLDL), intermediate density lipoprotein and high density lipoprotein (HDL) cholesterol and triglyceride levels, the size and particle number of LDL were determined at baseline and after 24 weeks, by nucleic magnetic resonance spectroscopy. FINDINGS: After 24 weeks of replacing the PI with NVP, we observed a reduction of total cholesterol (P = 0.028), LDL cholesterol (P = 0.001), the number of circulating LDL particles (P = 0.003) and the VLDL-1 triglyceride level (P = 0.032). A concomitant significant increase was observed in both HDL-cholesterol level (P = 0.002) and HDL particle size (P < 0.001). No significant changes were observed in the group that continued taking the PI. CONCLUSIONS: The replacement of PI by NVP improved the lipid profile both by reducing the number and lipid content of atherogenic LDL particles, and increasing the protective HDL fraction. Although total triglyceride levels remained unchanged, a reduction in the VLDL-1 fraction contributes to the reduction of LDL particles. These changes are expected to reduce the risk of cardiovascular disease in HIV-1-infected patients on highly active antiretroviral therapy. PMID- 12131216 TI - Phase I/II trial of HIV-1 hyperimmune globulin for the prevention of HIV-1 vertical transmission in Uganda. AB - OBJECTIVES: To assess the safety, tolerance, pharmacokinetics, and virologic and immunologic changes associated with the use of Ugandan HIV hyperimmune globulin (HIVIGLOB) in HIV infected pregnant Ugandan women and their infants. DESIGN: A prospective, phase I/II, three-arm dose escalation trial of HIVIGLOB. METHODS: HIVIGLOB was prepared from discarded HIV infected units of blood collected from the National Blood Bank in Kampala. From June 1996 to April 1997, 31 HIV positive pregnant women were enrolled with HIVIGLOB infusions given at 37 weeks gestation and within 16 h of birth for infants. The first 10 mother-infant pairs were infused at a dose of 50 mg/kg, followed by 11 pairs at 200 mg/kg, and 10 pairs at 400 mg/kg. Study participants were followed for 30 months. RESULTS: Thirty-one women and 29 infants were infused with HIVIGLOB. The infusions were safe and well tolerated by the women and their infants at all doses. There were no significant changes in virologic or immunologic parameters after HIVIGLOB infusion. Pharmacokinetic properties of this product were similar to other immune globulin products with a median half-life of 28 days in women and 30 days in infants. CONCLUSION: An HIV immune globulin product derived from HIV infected Ugandan donors is safe, well tolerated, and has pharmacokinetic properties consistent with other immunoglobulin products. Data suggest that a 400 mg/kg dose of HIVIGLOB would be the most appropriate dose for a subsequent efficacy trial of HIVIGLOB for the prevention of mother to child HIV transmission. PMID- 12131217 TI - On-going generation of multiple forms of HIV-1 intersubtype recombinants in the Yunnan Province of China. AB - OBJECTIVES: To investigate the molecular epidemiology of HIV in China's Yunnan Province, where the initial HIV-1 outbreak among injecting drug users (IDU) occurred in 1989, and to analyse the genesis and interrelationship of the epidemic with that in surrounding areas. DESIGN: A molecular epidemiological investigation was conducted among IDU in three prefectures in Yunnan Province, including Wenshan (east), Honghe (southeast) and Dehong (west). METHODS: Thirty nine specimens were collected from consenting IDU in 2000-2001. The nucleotide sequences of 2.6 kb gag-RT and 340 base pair (bp) env (C2/V3) regions were determined. Phylogenetic tree and recombination breakpoint analyses were performed. RESULTS: The circulating recombinant form (CRF), CRF08_BC, predominated in east Yunnan near Guangxi Province (89% in Wenshan and 81% in Honghe), whereas it was not detected in Dehong (0/14) in the west. In contrast, 71% (10/14) of the Dehong isolates were unique recombinant forms (URF), mostly between subtypes B' (Thailand variant of subtype B) and C, with distinct profiles of recombination breakpoints. The subtype B' accounts for the remaining 29% (4/14) of Dehong isolates. Interestingly, two Honghe isolates (2/16) shared some of the precise B'/C recombination breakpoints with CRF07_BC. CONCLUSION: New recombinant strains are arising continually in west Yunnan near the Myanmar border. Some appeared to be secondary recombinants derived from CRF07_BC that had further recombined with other strains. The uneven distribution of subtypes, CRF and URF, suggests the presence of independent transmission networks and clusters among IDU in Yunnan. PMID- 12131219 TI - HIV/HCV co-infection: clinical and therapeutic challenges. PMID- 12131218 TI - Pneumococcal disease in HIV-infected Malawian adults: acute mortality and long term survival. AB - OBJECTIVE: HIV-infected patients in Africa are vulnerable to severe recurrent infection with Streptococcus pneumoniae, but no effective preventive strategy has been developed. We set out to determine which factors influence in-hospital mortality and long-term survival of Malawians with invasive pneumococcal disease. DESIGN, SETTING AND PATIENTS: Acute clinical features, inpatient mortality and long-term survival were described among consecutively admitted hospital patients with S. pneumoniae in the blood or cerebrospinal fluid. Factors associated with inpatient mortality were determined, and patients surviving to discharge were followed to determine their long-term outcome. RESULTS: A total of 217 patients with pneumococcal disease were studied over an 18-month period. Among these, 158 out of 167 consenting to testing (95%) were HIV positive. Inpatient mortality was 65% for pneumococcal meningitis (n = 64), 20% for pneumococcaemic pneumonia (n = 92), 26% for patients with pneumococcaemia without localizing signs (n = 43), and 76% in patients with probable meningitis (n = 17). Lowered consciousness level, hypotension, and age exceeding 55 years at presentation were associated with inpatient death, but not long-term outcome in survivors. Hospital survivors were followed for a median of 414 days; 39% died in the community during the study period. Outpatient death was associated with multilobar chest signs, oral candidiasis, and severe anaemia as an inpatient. CONCLUSION: Most patients with pneumococcal disease in Malawi have HIV co-infection. They have severe disease with a high mortality rate. At discharge, all HIV-infected adults have a poor prognosis but patients with multilobar chest signs or anaemia are at particular risk. PMID- 12131220 TI - Viral load response to a pneumococcal conjugate vaccine, polysaccharide vaccine or placebo among HIV-infected patients. AB - We determined the HIV viral load in 66 adults randomly assigned to receive pneumococcal immunization with one or two doses of protein conjugate vaccine, one dose of polysaccharide vaccine, one dose of each, or placebo. Second doses were given 8 weeks after the first. Mean baseline viral load and CD4 cell count were 3.41 copies/ml (log10) and 457 cells/microl, respectively. We found no change in viral load during 24 weeks of follow-up for any vaccine or combination of vaccines or placebo. PMID- 12131221 TI - Liver injury after beginning antiretroviral therapy in HIV/hepatitis C virus co infected patients is not related to immune reconstitution. AB - Transaminase elevations occur more frequently after beginning antiretroviral therapy in HIV-positive patients co-infected with hepatitis C virus (HCV). The mechanism of liver injury in these individuals is unknown, although immune reconstitution phenomena have been postulated. In 42 HIV/HCV co-infected individuals followed after beginning potent antiretroviral therapy, the development of liver injury was not associated with significant changes in serum HCV-RNA levels nor with greater CD4 cell increases. Underlying chronic hepatitis may thus increase the risk of liver toxicity by other mechanisms. PMID- 12131222 TI - Increased risk of HIV and sexually transmitted disease transmission among gay or bisexual men who use Viagra, San Francisco 2000-2001. AB - The potential role of sildenafil (Viagra) in the risk of HIV and sexually transmitted disease (STD) transmission was evaluated among gay or bisexual men seeking public STD services in San Francisco. Viagra users reported greater numbers of recent sex partners, higher levels of unprotected anal sex with an HIV positive partner, and higher rates of prevalent STD than non-users. Moreover, mixing Viagra with illicit drugs was commonly reported. Further studies are needed to determine whether a causal role exists. PMID- 12131224 TI - No evidence of benefical effect of GB virus type C infection on the course of HIV infection. PMID- 12131223 TI - Bacillary splenitis (Bartonella henselae) during immune restoration in an HIV infected patient. PMID- 12131225 TI - Long-term efficacy of dual nucleoside reverse transcriptase inhibitor antiretroviral therapy in HIV-1 infection. PMID- 12131226 TI - Pneumocystis carinii pneumonia after the discontinuation of secondary prophylaxis. PMID- 12131227 TI - Triglyceride increase can predict lipodystrophy in HIV patients under highly active antiretroviral therapy. PMID- 12131228 TI - Highly active antiretroviral therapy for patients with tuberculosis: the solution or the problem? PMID- 12131229 TI - Cytomegalovirus-induced hemorrhagic cystitis in AIDS patient treated successfully with valganciclovir. PMID- 12131230 TI - Efficacy of imiquimod on external anogenital warts in HIV-infected patients previously treated by highly active antiretroviral therapy. PMID- 12131232 TI - Scientific considerations for the regulation and clinical evaluation of HIV/AIDS preventive vaccines: report from a WHO-UNAIDS Consultation 13-15 March 2001, Geneva, Switzerland. AB - The consultation was jointly organized by the WHO-UNAIDS HIV Vaccine Initiative and the Quality Assurance and Safety of Biologicals Team of the World Health Organization (WHO). Thirty-four experts from 16 developed and developing countries attended the meeting, bringing together expertise from academic institutions, clinical trial centres, national and international regulatory authorities. Representatives of major pharmaceutical companies were also invited. The primary objective of the meeting was to identify gaps that need to be addressed from regulatory perspective to ensure appropriate progress of HIV vaccine development from basic research to human trials, licensing and future application, with a special focus on needs of developing countries. As a result of discussions, the following priority needs were identified and recommendations were made in order to establish an appropriate regulatory framework for the development and evaluation of preventive HIV/AIDS vaccines, which were divided in two main areas: (a) standardization and control of candidate HIV/AIDS vaccines, and (b) approaches to the conduct of clinical trials of candidate HIV/AIDS vaccines. PMID- 12131231 TI - Rhodococcus equi: pulmonary cavitation lesion in patient infected with HIV cured by levofloxacin and rifampicin. PMID- 12131233 TI - The Minnesota Living With Heart Failure Questionnaire: sensitivity to differences and responsiveness to intervention intensity in a clinical population. AB - BACKGROUND: The Minnesota Living With Heart Failure Questionnaire (LHFQ) is a commonly used measure of health-related quality of life in persons with heart failure. Researchers have questioned whether LHFQ is sensitive to subtle differences and sufficiently responsive to clinical interventions because the instrument has demonstrated variable performance in clinical trials. OBJECTIVES: A secondary analysis was conducted to assess the LHFQ for sensitivity to different clinical states and responsiveness to varying intensities of clinical intervention. METHODS: A convenience sample of nine experimental or quasi experimental studies from eight clinical sites in the United States yielded data from 1,136 patients with heart failure. Data in the studies had been collected at enrollment and one, three, and/or six months later. Data were analyzed using descriptive, univariate, and multivariate techniques. RESULTS: Total and subscale scores on LHFQ were poorer in those with worse New York Heart Association functional class, although there was no difference in LHFQ scores between classes III and IV. No difference in LHFQ scores was found when patients were classified by ejection fraction. Scores improved significantly following hospital discharge, even in those in the control group. Changes in LHFQ scores were greatest in those receiving high intensity interventions. CONCLUSIONS: The LHFQ is sensitive to major differences in symptom severity but may not be sensitive to subtle differences. It is responsive to high intensity interventions. Investigators are cautioned against using this instrument without first maximizing intervention power or without a control group for comparison. PMID- 12131234 TI - Effects of behavioral interventions on disruptive behavior and affect in demented nursing home residents. AB - BACKGROUND: Disruptive behaviors are prevalent in nursing home residents with dementia and often have negative consequences for the resident, caregiver, and others in the environment. Behavioral interventions might ameliorate them and have a positive effect on residents' mood (affect). OBJECTIVES: This study tested two interventions-an activities of daily living and a psychosocial activity intervention-and a combination of the two to determine their efficacy in reducing disruptive behaviors and improving affect in nursing home residents with dementia. METHODS: The study had three treatment groups (activities of daily living, psychosocial activity, and a combination) and two control groups (placebo and no intervention). Nursing assistants hired specifically for this study enacted the interventions under the direction of a master's prepared gerontological clinical nurse specialist. Nursing assistants employed at the nursing homes recorded the occurrence of disruptive behaviors. Raters analyzed videotapes filmed during the study to determine the interventions' influence on affect. RESULTS: Findings indicated significantly more positive affect but not reduced disruptive behaviors in treatment groups compared to control groups. CONCLUSIONS: The treatments did not specifically address the factors that may have been triggering disruptive behaviors. Interventions much more precisely designed than those employed in this study require development to quell disruptive behaviors. Nontargeted interventions might increase positive affect. Treatments that produce even a brief improvement in affect indicate improved quality of mental health as mandated by federal law. PMID- 12131235 TI - Anger in early adolescent boys and girls with health manifestations. AB - BACKGROUND: Some theorists suggest that boys and girls might experience and express anger in different ways, while others do not, making the study of sex differences in anger an important area for investigation. Further, much has been written theoretically about the health implications of anger, but there is a paucity of studies that have examined the relationship between anger and health variables in early adolescent boys and girls separately. OBJECTIVES: The objectives of this study were to examine sex differences in anger in early adolescents, and to examine the relationship between anger and several health variables, e.g., current health status, clinical health, eudaimonistic health for boys and girls separately. METHODS: This study compared differences in five anger variables between boys and girls, and examined relationships between the anger variables and health variables for boys and girls separately. The final sample consisted of 148 seventh and eighth graders, ages 12 to 14; 81 were girls and 67 were boys. They responded to the State-Trait Anger Expression Inventory and instruments measuring three health variables. RESULTS: Using multisample analysis via LISREL 7 and independent t tests, findings indicated that boys and girls did not differ in the experience and expression of anger. Pearson correlations were used to examine the relationships between the anger variables and the health variables for boys and girls separately. Of the 30 relationships examined, 12 were statistically significant; seven of these correlations were for girls, while five were for boys. CONCLUSIONS: Early adolescent boys and girls may not differ in any meaningful way in self-reported experiences and expressions of anger, but they may differ in health outcomes in relation to various types of anger. PMID- 12131236 TI - A randomized trial of a cognitive coping intervention for acutely ill HIV positive men. AB - BACKGROUND: People who are HIV-positive now live longer when they have contracted AIDS, and nursing interventions can help improve their quality of life. OBJECTIVES: To test the effects of an intervention based on developing cognitive coping skills as compared to one focused on facilitating the expression of emotions. Both interventions were intended to help regulate emotional response to an exacerbation of HIV-related symptoms. METHOD: In a randomized, controlled trial, 90 hospitalized HIV-positive men were randomly assigned to one of three groups: cognitive, expression, or control. The intervention was administered on three consecutive days in 20-30 minute sessions. Preintervention and postintervention data were gathered on mood, distress, and anxiety. RESULTS: Both interventions produced a beneficial effect on negative affect (cognitive group p =.002, expression group p =.011), and immediately following the first daily session (p =.001). No change in positive affect was produced by either intervention. Paired t tests indicated a decrease in distress (p =.039), specifically, of intrusive ideation (p =.03), for the cognitive group, which also experienced a decrease in anxiety from immediately before to immediately after each session. Conversely, the expression group experienced an increase in anxiety (p =.018). DISCUSSION: The cognitive coping skills nursing intervention was effective in helping to regulate HIV-positive persons' emotional responses to advanced disease. This nursing intervention is feasible for use by skilled practitioners providing daily care. PMID- 12131237 TI - Tobacco: an emerging topic in nursing research. AB - BACKGROUND: Tobacco use is the leading cause of preventable death in the United States. Since the first Surgeon General's Report in 1964 on the health risks of tobacco use, overwhelming evidence regarding increased tobacco-attributable morbidity and mortality has been reported. The purpose of this review was to explore nursing research contributions to this public health issue by evaluating the emergence of publications focused on tobacco in a leading nursing research journal. OBJECTIVES: The specific aims of this review were to determine, among data-based articles published in Nursing Research (1952-2000), how often tobacco use was included (a) in sample descriptions, (b) as a variable potentially associated with study outcomes, and (c) as a finding. Additionally, the frequency of publication of research instruments developed to study tobacco use was evaluated. METHODS: Data-based articles (n = 1,705) and research briefs (n = 197) were evaluated. Inter-rater reliability (100%) was established by the re-review of 20% of the issues in each decade. RESULTS: A total of 40 data-based articles (2% of those reviewed) either included tobacco use in the sample description only (n = 11), as an independent or mediating variable (n = 11), or as a finding (n = 18). The majority (53%) of the articles were published since 1990; and 71% of the outcome studies were published within the past 5 years. One study focused on tobacco use among youth, and 1 of 197 instrument articles reviewed focused on tobacco. None of the studies reviewed addressed prevention of tobacco use or strategies to decrease exposure to second-hand smoke. CONCLUSIONS: This review demonstrates that the cessation of tobacco use is emerging as a topic for nursing research, reflecting the increased public health attention on this topic. Increased research efforts are needed in the areas of tobacco cessation and prevention of tobacco use. Researchers should be encouraged to consider tobacco use as a variable potentially affecting outcomes in other research studies. PMID- 12131238 TI - The mediating effect of pain and fatigue on level of functioning in older adults. AB - BACKGROUND: Medical conditions and symptoms have been shown to predict level of functioning in older adults, but medical conditions and symptoms have rarely been investigated together in a comprehensive model that included both medical conditions and symptoms as predictors of functioning in older adults. OBJECTIVE: The purpose of this study was to determine whether the adverse effect of medical conditions on different aspects of functioning in older adults is mediated by the level of symptoms (pain and fatigue). If so, level of functioning may improve if pain or fatigue can be mitigated, even when underlying medical conditions cannot be cured. METHOD: Data from 225 adults aged 65-90 were used to test whether medical conditions, symptoms (pain and fatigue), and six covariates predicted lower body performance, self-reported physical functioning, and self-reported role and social functioning. The fit of a series of models to the data was analyzed using structural equation modeling. RESULTS: Medical conditions affected self-reported physical functioning and self-reported role and social functioning by increasing the level of symptoms, rather than by direct association. Further descriptive studies are needed to identify other symptoms and modifiable mechanisms by which medical conditions affect functioning. Researchers who investigate the causes of poor functioning in older adults are encouraged to include symptoms in models that hypothesize medical conditions as predictors of functioning outcomes. PMID- 12131239 TI - Issues in protection of human subjects in internet research. AB - BACKGROUND: Despite the increasing use of the Internet among nurses, the use of the Internet in nursing research has been rarely discussed and critiqued in terms of issues in protection of human subjects. APPROACH: In this article, issues in protection of human subjects in Internet research are explored by analyzing an Internet study to propose directions for human protection in Internet research. RESULTS: Issues raised through the study include those related to (a) anonymity and confidentiality, (b) security, (c) self-determination and authenticity, (d) full disclosure, and (e) fair treatment. DISCUSSION: Based on discussion of the five issues, development of standardized guidelines, investigator triangulation, and information sharing are proposed as directions for protection of human subjects in Internet research. PMID- 12131240 TI - Retrospective data collection using event history calendars. AB - BACKGROUND: Event history calendars are used to collect retrospective data about events and life transitions over short and long periods of time. Event history calendars are highly structured, but flexible, approaches to interviewing respondents about past events that use their own past experiences as cues to remembering. Event history calendars incorporate autobiographical memory retrieval mechanisms to assist respondents in reconstructing past events and experiences accurately and completely. APPROACH: A sample event history calendar and experiences from an ongoing study of adolescent risk behavior are described to illustrate event history calendar methodology application in nursing research. Event history calendar design, recording, interviewing, and interviewer training descriptions are included. DISCUSSION: Event history calendars have been used extensively for retrospective data collection of occurrence, timing, and sequencing of a variety of life events in population studies, psychology, and sociology research, but not in nursing research. Because event history calendars improve recollection of complex sequences of personal events, they would be ideal for retrospective data collection in quantitative and qualitative nursing studies. Nursing expertise in history-taking make this a natural method of choice for retrospective data collection and as a means of stimulating communication during interviewing. PMID- 12131241 TI - Ruptured Achilles tendons show increased lectin stainability. AB - PURPOSE: To ascertain whether lectins could be a useful tool for investigation of the extracellular matrix of degenerated and normal tendons. METHODS: Hematoxylin eosin-stained slides were assessed blindly using a semiquantitative grading scale for fiber structure, fiber arrangement, rounding of the nuclei, regional variations in cellularity, increased vascularity, decreased collagen stainability, hyalinization, and glycosaminoglycan, with a pathology score giving up to three marks per each of the above variables, with 0 being normal and 3 being maximally abnormal. For lectin staining with Aleuria aurantia, Canavalia ensiformis, Galanthus nivalis, Phaseolus vulgaris, Arachis hypogea, Sambucus nigra, and Triticum vulgaris, assessment of staining on a scale from 0 (no staining) to 5 (strong staining) was performed blindly. RESULTS: The mean pathology sum score of ruptured tendons (N = 14; average age 46.5 yr, range 29 61) was significantly greater than the mean pathology score of the control tendons of Achilles tendons from individuals with no known tendon pathology (N = 16; average age 62.5 yr, range 49-73) (pathology score: 18.5 +/- 3.2 vs 6.1 +/- 2.3). Four of the seven lectins used exhibited significantly positive results. CONCLUSIONS: Ruptured tendons were histologically significantly more degenerated than control tendons. Ruptured tendons showed different lectin staining properties than nonruptured ones. This difference may have resulted from posttranslational changes in the extracellular matrix producing alterations in the biochemistry of the tendon, which might interfere with the interaction with the lateral sugar residues of the collagen molecules or cause steric blockade. PMID- 12131242 TI - Body mass index and mortality: the influence of physical activity and smoking. AB - PURPOSE: To study the association between body mass index (BMI) and mortality, and to evaluate the effect of physical activity during leisure time and smoking on this association in a general male population. METHODS: During 1974-1978, all men aged 35-49 yr living in three Norwegian counties were invited to a cardiovascular screening, and 87.1% attended and had their weight and height measured. Men with recognized cardiovascular diseases, diabetes mellitus, or cancer at screening were excluded. The cohort (N = 22,304) was followed for an average of 16.3 yr with respect to total and cause-specific mortality. RESULTS: During follow-up, 1909 men died. We found a J-shaped association between BMI and total mortality, and the form of association was similar for death from cardiovascular diseases. Although not statistically significant, a J-shaped association was also suggested in never-smokers. Irrespective of BMI level, ex- and never-smokers had lower mortality than current smokers. Obese smoking men had a relative risk of dying of 2.01 (95% CI: 1.29-3.11) compared with obese never smokers, and a relative risk of 4.55 (95% CI: 3.34-6.20) compared with normal weight never-smokers (BMI 22-24.9 kg x m(-2)). Within each category of physical activity during leisure time, obese men had a similar increased relative risk of death compared with normal-weight individuals. However, the U- to J-shaped association between BMI and mortality seemed to disappear by increasing level of physical activity, but this finding was not significant. CONCLUSION: This study suggests a J-shaped association between BMI and total mortality, also when stratified on smoking habits and physical activity. The suggested linear trend in the most physical active men needs to be reassessed. PMID- 12131243 TI - Is there a threshold intensity for aerobic training in cardiac patients? AB - PURPOSE: Recent guidelines have recommended the use of a percentage of oxygen uptake reserve (VO2R) for prescribing aerobic exercise intensity for cardiac patients. Moreover, these guidelines suggest that a threshold intensity may exist, below which no improvement in peak oxygen uptake (VO2peak) occurs. The purpose, therefore, was to translate the intensity of aerobic exercise in previous training studies using cardiac patients into %VO2R units, and determine whether a threshold intensity exists. METHODS: Twenty-three studies, using 28 groups of aerobically trained cardiac patients, were identified in which VO2peak was measured before and after training by gas exchange. Intensity of exercise was variously described as a percentage of VO2peak, percentage of peak heart rate (HRpeak), percentage of heart rate reserve (HRR), or percentage of peak workload. These intensities were translated into equivalent units of %VO2R. RESULTS: Of the 28 groups of patients, three failed to show significant improvements in VO2peak. These groups exercised at intensities corresponding to 47-55% of VO2R. However, six other groups exercised at comparable intensities (i.e., 42% to 55% of VO2R) and experienced significant increases in VO2peak. Other confounding variables in these studies were similar, including the initial VO2peak of the subjects, suggesting that the failure of three groups to significantly improve aerobic capacity was due to their small sample size. CONCLUSION: No threshold intensity for aerobic training was identified in cardiac patients, with the lowest intensity studied being approximately 45% of VO2R. It is possible that intensities below this value may be an effective training stimulus, especially in extremely deconditioned subjects, but further research is needed to test that possibility and to determine whether a threshold exists. PMID- 12131244 TI - Low-back stiffness is altered with warm-up and bench rest: implications for athletes. AB - INTRODUCTION: General practice in many team sports is to have the athletes who do not start in a game sit on a bench while waiting to play. The purpose of this study was to examine the effect of a warm-up followed by bench rest on the stiffness of the lumbar spine in athletes. METHODS: Nine varsity-level volleyball players volunteered to have their lumbar-spine stiffness measured. The protocol consisted of an initial stiffness measurement followed by a 30-min warm-up, then another stiffness measurement, then 30 min of bench rest, and finally a third stiffness measurement. RESULTS: In general, lumbar spine stiffness increased as a result of bench rest after a warm-up. This effect was seen in both the spine extension and lateral bend axes but not in the flexion or axial twist axes. However, there was no decrease in stiffness associated with the active warm-up portion of the task. CONCLUSIONS: It was concluded that a warm-up followed by bench rest does lead to an increase in stiffness of the lumbar spine, suggesting this practice is not in the best interest of reducing the risk of back injury or optimal performance. PMID- 12131245 TI - EMG activity normalization for trunk muscles in subjects with and without back pain. AB - PURPOSE: The aims of the present study were to examine electromyographic (EMG) activity of six bilateral trunk muscles during maximal contraction in three cardinal planes and to determine the direction of contraction that gives maximal activation for each muscle, both for healthy subjects and back-pain patients. METHODS: Twenty-eight healthy subjects and 15 back-pain patients performed maximum voluntary contractions in three cardinal planes. Surface EMG signals were recorded from rectus abdominis, external oblique, internal oblique, latissimus dorsi, iliocostalis lumborum, and multifidus bilaterally. Root mean square values of the EMG data were calculated to quantify the amplitude of EMG signals. RESULTS: For both healthy subjects and back-pain patients, one single direction of contraction was found to give the maximum EMG signals for most muscles. Rectus abdominis demonstrated maximal activity in trunk flexion, external oblique in lateral flexion, internal oblique in axial rotation, and multifidus in extension. For the latissimus dorsi and iliocostalis lumborum, maximal activity was demonstrated in more than one cardinal plane. CONCLUSION: This study has implications for future research involving normalization of muscle activity to maximal levels required in many trunk EMG studies. As the latissimus dorsi and iliocostalis lumborum demonstrate individual differences in the plane that gives maximal activity, these muscles may require testing in more than one plane. PMID- 12131246 TI - Physical activity, sports participation, and suicidal behavior among college students. AB - PURPOSE: To evaluate the relationship between physical activity, sports participation, and suicidal behavior among college students (N = 4,728). METHODS: Data from the 1995 National College Health Risk Behavior Survey were analyzed. Students were classified as engaging in frequent vigorous activity 6-7 d.wk-1, vigorous activity 3-5 d.wk-1, moderate activity, low activity, or no activity. Sports participation was dichotomized into "yes" or "no" participation. Suicidal behavior was defined as thoughts about, plans for, or attempts at suicide during the 12 months before completing the survey. Data were stratified by sex and multivariable logistic regression modeling, calculated odds ratios (ORs) (adjusted for age, race/ethnicity, Body Mass Index/weight perception, cigarette smoking, episodic heavy alcohol use, drug use, and either activity level or sport participation) for suicidal behavior as associated with physical activity and sports participation. RESULTS: Adjusted ORs show that men in the "low activity" group were at almost half the odds (adjusted OR = 0.54; P < 0.015) of reporting suicidal behavior than men in the "not active" group. Women who engaged in moderate or frequent vigorous activity were at greater odds of reporting suicidal behavior compared with inactive women; OR = 1.76 (P < 0.035) and 1.99 (P < 0.034) respectively. Sports participation was protective against suicidal behavior. Adjusted ORs show that men who did not participate in sports were 2.5 times (P < 0.0003) more likely to report suicidal behavior than men who were sports participants. Women not participating in sports had 1.67 times the odds of reporting suicidal behavior than women sports participants (P < 0.05). CONCLUSIONS: Associations were found between sports participation/selected patterns of physical activity and suicidal behavior. Causal factors mediating the relationships need to be identified. PMID- 12131247 TI - Hemodynamic responses to stress among black women: fitness and parental hypertension. AB - PURPOSE: We compared hemodynamic aspects of the relationship between cardiorespiratory fitness and blood pressure changes during and after laboratory stress in young black women with or without parental history of hypertension. METHODS: Participants were 30 normotensive, black American women having low to moderate fitness levels (i.e., VO2peak) assessed by cycle ergometry who performed standard active and passive coping laboratory stressors. Blood pressure, heart rate, stroke volume, cardiac output, total peripheral resistance (TPR), calf blood flow (CBF), and calf vascular resistance (CVR) were assessed during exposure to forehead and hand cold pressors, and mental arithmetic, as well as during recovery after the tasks. RESULTS: Fitness was positively related to increases in either TPR or CVR during each stressor. In contrast, fitness was positively related to blunted blood pressure during or after passive stress (i.e., hand or forehead cold) and enhanced recovery of blood pressure and TPR after the active stressor (i.e., mental arithmetic); effects of fitness on the vascular responses during and after mental arithmetic were stronger among women having a negative history of parental hypertension. CONCLUSION: The findings confirm our previous report that fitness blunts systolic blood pressure response during the hand cold pressor in young women. They also suggest that future studies of fitness and blood pressure reactivity during stress should focus on the regulation of vascular responses and their recovery after stress. Weaker effects of VO2peak after mental arithmetic in the positive history group indicate that the level of fitness required to modify recovery from mental stress among black American women may differ according to parental history of hypertension. PMID- 12131249 TI - Training down-regulates fatty acid synthase and body fat in obese Zucker rats. AB - INTRODUCTION: The purpose of this study was to investigate whether chronic exercise training attenuates fatty acid synthase, the rate-limiting enzyme for hepatic lipogenesis, and the accumulation of body fat by using obese Zucker rats (OZR) as a model. METHODS: Female obese Zucker (fa/fa) rats (O, N = 16) and their lean litter mates (L, N = 16) were randomly divided into a trained (T) and untrained (U) group. T was performed on a treadmill for 2 h.d-1, 5 d.wk-1, 10 wk with running speed and grade adjusted to elicit similar workloads. All rats were meal-fed a high-cornstarch diet for 4 h.d-1 and killed 8 h after the initiation of the last meal and 27 h after the last T session, in the resting state. RESULTS: O rats exhibited twofold higher FAS activity and sixfold higher FAS mRNA abundance in the liver than L rats (P < 0.05), accompanied by a severe hyperinsulinemia (P < 0.05) but normal glucagon and glucose levels. FAS activity, but not mRNA level, was decreased by 18% with T in O rats (P < 0.05). T decreased percent body fat in both O and L rats (P < 0.05), and increased lean body mass in O rats (P < 0.05). Hepatic fatty acid profile showed higher 16:0, 16:1, and 18:1 concentrations in O rats, whereas 18:0, 18:2, and 20:4 were lower (P < 0.05). Training increased 20:4 in both O and L rats (P < 0.08). Nuclear protein binding to the insulin response sequence (IRS/A) and carbohydrate response element (ChoRE) on FAS gene promoter was decreased, whereas inverted CAATT box element (ICE) binding was increased in O versus L rats (P < 0.05). Training did not affect the binding of these gene sequences. CONCLUSION: De novo lipogenesis was greatly enhanced in OZR. Endurance training decreased body fat, which is partly explained by a decreased FAS activity. However, FAS down-regulation was not due to altered nuclear protein binding to FAS gene. PMID- 12131250 TI - Aging does not attenuate plantaris muscle hypertrophy in male Fischer 344 rats. AB - PURPOSE: The present study examined the effect of extreme old age on the plasticity of the rat plantaris muscle in response to an increase in mechanical load. METHODS: Male Fischer 344 rats, aged 7 months (adult) and 25 months (old) underwent bilateral surgical ablation of the gastrocnemius muscle to functionally overload (OV) the fast-twitch plantaris muscle for 8 wk RESULTS: At 27 months of age, plantaris wet weight and cross-sectional area (CSA) were unaffected by age, but aging decreased peak isometric tension (Po) 27% (P < 0.05). Plantaris muscle myosin heavy chain composition indicated a loss of faster myosin heavy chains (MHC) isoforms with concomitant increases in slower MHC in old rats (P < 0.05). In adult rats, OV increased muscle CSA and Po 72% and 83%, respectively (P < 0.05). Similarly, OV increased CSA and Po 69% and 73%, respectively, in old rats (P < 0.05). Average fiber CSA increased 57% and 68% in adult-OV and old-OV rats, respectively (P < 0.05). CONCLUSION: Collectively, our data indicate that plantaris muscle mass and plasticity in response to increased mechanical load are well conserved in very aged male Fischer 344 rats. PMID- 12131251 TI - Peripheral vascular responses to heat stress after hindlimb suspension. AB - PURPOSE: The purpose of this study was to determine whether hindlimb suspension (which simulates the effects of microgravity) results in impaired hemodynamic responses to heat stress or alterations in mesenteric small artery sympathetic nerve innervation. METHODS: Over 28 d, 16 male Sprague-Dawley rats were hindlimb suspended, and 13 control rats were housed in the same type of cage. After the treatment, mean arterial pressure (MAP), colonic temperature (Tcol), and superior mesenteric and iliac artery resistances (using Doppler flowmetry) were measured during heat stress [exposure to 42 degrees C until the endpoint of 80 mm Hg blood pressure was reached (75 +/- 9 min); endpoint Tcore = 43.6 +/- 0.2] while rats were anesthetized (sodium pentobarbital, 50 mg x kg(-1) BW). RESULTS: Hindlimb suspended and control rats exhibited similar increases in Tcol, MAP, and superior mesenteric artery resistance, and similar decreases in iliac resistance during heat stress (endpoint was a fall in MAP below 80 mm Hg). Tyrosine hydroxylase immunostaining indicated similar sympathetic nerve innervation in small mesenteric arteries from both groups. CONCLUSION: Hindlimb suspension does not alter the hemodynamic or thermoregulatory responses to heat stress in the anesthetized rat or mesenteric sympathetic nerve innervation, suggesting that this sympathetic pathway is intact. PMID- 12131253 TI - Physical activity and glucose tolerance in elderly men: the Zutphen Elderly study. AB - PURPOSE: To determine whether physical activity is associated with glucose tolerance in the elderly. METHODS: We examined current and 5-yr change in physical activity in relation to glucose tolerance in 424 randomly selected male inhabitants of the Dutch town Zutphen, aged 69-89 yr, without known diabetes mellitus. Physical activity was assessed by a validated questionnaire designed for retired men. Glucose intolerance was assessed by an oral glucose tolerance test and defined as impaired glucose tolerance or diabetes mellitus. RESULTS: Men with 30 min x d(-1) or more of physical activity of at least moderate intensity had a lower prevalence of glucose intolerance as compared to men without these activities (age-adjusted odds ratio 0.32; 95% CI, 0.18-0.57). Adjustment for family history of diabetes, smoking, alcohol intake, dietary factors, body mass index, and subscapular skin-fold thickness or exclusion of men with cardiovascular diseases or disabilities did not substantially change the results. With specific activities modeled simultaneously, bicycling (P for trend = 0.01) and gardening (P for trend = 0.02) were inversely associated with glucose intolerance. Men whose amount of physical activity had decreased during the past 5 yr had significantly higher age-adjusted 2-h glucose concentrations as compared with men who remained at least as active (difference 0.7 mmol x L(-1); 95% CI, 0.1-1.3). CONCLUSION: These findings suggest that common types of physical activity such as bicycling and gardening may contribute to the prevention of glucose intolerance in elderly men PMID- 12131252 TI - Oral amino-acid provision does not affect muscle strength or size gains in older men. AB - PURPOSE: The intent of this investigation was to examine the effects of a daily oral provision consisting of amino acids (L-lysine, L-leucine, L-valine, L phenylalanine, L-threonine, L-histidine, L-isoleucine, and L-methionine) in combination with carbohydrates (dextrose, sucrose, and fructose) on whole muscle strength and size characteristics during a 12-wk progressive knee extensor resistance training (PRT) program in older men (>65 yr). METHODS: Seventeen older men were randomly assigned to either the experimental (EX) or control (CN) groups. The EX (N = 8) and CN (N = 9) groups had the following characteristics EX: 70.8 +/- 1.5 yr, 91.0 +/- 4.9 kg, and 177.0 +/- 3.9 cm; CN: 72.1 +/- 1.9 yr, 75.4 +/- 4.7 kg, and 176.1 +/- 3.0. Pre and post PRT maximal unilateral isometric torque (N.m), isokinetic torque (1.05, 1.57, 2.09, 3.14, 4.19, and 5.24 rad.s-1), work capacity (30 consecutive reps at 3.14 rad.s-1) torque, one repetition maximum (1RM) bilateral isotonic strength, and whole muscle cross-sectional area (CSA) of the mid-thigh were performed by computed tomography on each subject. RESULTS: All variables showed an improvement with training (P < 0.05); however, there were no differences between the groups. Both groups increased in isometric strength by 21%, and isokinetic torque by 24% to 11% with the varying velocities (1.05-5.24 rad.s-1). Whole muscle 1RM strength and thigh CSA increased 50% and 6.5%, respectively. Additionally, voluntary torque/CSA increased 12% in both the EX and CN groups (P < 0.05). CONCLUSIONS: In conclusion, these data suggest that whole muscle strength and size are not enhanced with a postexercise daily provision of an oral amino-acid complex during 12 wk of PRT in older men. PMID- 12131254 TI - Familial aggregation of physical activity levels in the Quebec Family Study. AB - PURPOSE: Familial aggregation of physical activity phenotypes was investigated in 696 subjects from 200 families of the Quebec Family Study. The mean age of offspring and parents was 27 and 53 yr, respectively. METHODS: The levels of physical activity were estimated using a 3-d diary and a questionnaire dealing with physical activity during the past year. RESULTS: An ANOVA performed on the age and sex adjusted physical activity phenotypes revealed that there were 1.40 1.52 times more variation in physical activity levels between families than within families (0.001 < P < 0.0001), suggesting that physical activity levels aggregate in families. Maximal heritabilities (SEGPATH), adjusted for the degree of spouse resemblance, reached 25%, 16%, 19%, and 17% for the degree of inactivity, time spent in moderate to strenuous physical activities, total level of daily activity, and weekly time spent in the main activity during the previous year, respectively. CONCLUSION: These results suggest that physical activity level is characterized by a significant degree of familial resemblance, and that inactivity has a slightly higher heritability level than moderate to strenuous physical activity or total physical activity phenotypes. The pattern of familial correlations suggests that shared familial environmental factors along with genetic factors are also important in accounting for the familial resemblance in physical activity level. PMID- 12131255 TI - The assessment of intersegmental motion and pelvic tilt in elite oarsmen. AB - Low back pain (LBP) is a common problem in elite oarsmen. The relevance of spinal and pelvic flexibility to good rowing technique and the incidence of LBP is unclear. PURPOSE: The aim of this study was to investigate patterns of spinal and pelvic mobility in a group of elite oarsmen with and without a history of LBP. METHODS: Twenty elite oarsmen were recruited into this study, including nine with no history of spinal problems, four with a current spinal problem, and the remainder with a history of LBP. Subjects were scanned using an interventional magnetic resonance imaging (MRI) scanner. Four key stages of the rowing stroke were simulated within the scanner, and sagittal images of the lumbar spine and sacrum were obtained. From these images intersegmental motion was determined along with the angle of lordosis and position of the lumbar spine and sacrum. RESULTS: Different mobility trends were seen; oarsmen with no history of LBP demonstrated the greatest mobility in their lower lumbar regions (at the L5/S1 level in the catch position 7.5 degrees +/-1.3 in normals; 4.8 degrees +/-1.2 in previous LBP groups; and 2.8 degrees +/-5.5 in current LBP group) and the lowest rotation of their pelvis (level in the catch position 13.9 degrees +/-11.2 in normals; 16.1 degrees +/-6.8 in previous back pain groups; and 15.2 degrees +/ 11.2 in current back pain group). In contrast, those with either current or previous LBP presented with a hypomobility of their spine which appeared to be compensated for by increased pelvic rotation. CONCLUSIONS: Marked differences were observed in the motion characteristics of these 3 groups of oarsmen. At present it is not known if these changes are causative or effect. PMID- 12131256 TI - Frontal plane knee angle affects dynamic postural control strategy during unilateral stance. AB - PURPOSE: Center of plantar pressure (COPP) location moves toward the forefoot as ankle plantar flexor muscles attempt to maintain postural control during single leg stance. This study evaluated relationships between frontal plane tibiofemoral joint angulation during relaxed bilateral stance and mean COPP locations during vision-denied single leg stance at 20 degrees knee flexion. METHODS: Fifty-six nonimpaired athletes (29 female, 27 male) were evaluated for frontal plane tibiofemoral joint angulation and standing foot angle by using two-dimensional videography (30 Hz). Mean anterior-posterior and mediolateral COPP locations were assessed during single leg stance on a mat (25 Hz, 15 s). One-way ANOVA and Tukey HSD tests evaluated group differences (P < or = 0.05) based on frontal plane tibiofemoral joint angulation. RESULTS: Group 1 (genu varus or genu valgus < 5 degrees ) displayed a mean anterior-posterior COPP location of 54.2 +/- 6% from the (0,0) coordinate starting point at the anterolateral foot (10.3 +/- 2 cm from the posterior sensor edge). Group 2 (genu varus angulation > or = 5 degrees ) and group 3 subjects (genu valgus angulation > or = 5 degrees ) displayed mean anterior-posterior COPP locations of 60.6 +/- 8% and 60.7 +/- 7% (8.8 +/- 2 cm and 8.7 +/- 2 cm from the posterior sensor edges), respectively. Group 2 (12.5 +/ 3 N x kg(-1)) and group 3 (12.4 +/- 3.1 N x kg(-1)) subjects also displayed greater mean plantar force magnitude/body weight than group 1 (10.3 +/- 2 N x kg( 1)) subjects. Mean ankle plantar flexor moment magnitudes did not differ between groups. CONCLUSIONS: Rearfoot directed mean anterior-posterior COPP locations and greater plantar force magnitudes/body weight suggests that subjects with genu varus or genu valgus relied more on the subtalar and midtarsal joint control function of the ankle plantar flexor muscle group for lower extremity dynamic postural control. PMID- 12131257 TI - Emotional responsiveness after low- and moderate-intensity exercise and seated rest. AB - PURPOSE: Few experiments have been conducted regarding the effects of exercise on emotional responsiveness. The aim of this experiment was to determine whether anxiety-reducing conditions of low- and moderate-intensity cycling exercise lead to changes in emotional responsiveness to pictures designed to elicit pleasant, neutral, and unpleasant emotions. METHODS: 24 healthy college women completed counterbalanced conditions of 25 min of low- and moderate-intensity cycling exercise and seated rest. Indices of emotional responsiveness, including the acoustic startle eyeblink and corrugator supercilii responses, as well as baseline corrugator supercilii electromyographic (EMG) activity, were measured immediately before and 20 min after each condition while participants viewed pleasant, neutral, and unpleasant pictures from the International Affective Picture System. RESULTS: State anxiety was significantly reduced 20 min after each condition. Startle response magnitude was modulated by the affective content of the pictures and was reduced after each condition in response to each type of picture. Baseline corrugator EMG activity did not change after seated rest but decreased in an exercise intensity-dependent fashion after cycling. Corrugator EMG responses during the pictures were not different between conditions or from pre- to post-conditions. CONCLUSION: The findings suggest that cycling exercise results in decreased baseline activity of facial muscles involved in the expression of emotion but does not lead to changes in appetitive or defensive responses to emotional stimuli. Furthermore, anxiolytic conditions of low- and moderate-intensity cycling exercise and seated rest are related to decreased startle magnitude in healthy college women. PMID- 12131258 TI - Self-regulated cycling using the Children's OMNI Scale of Perceived Exertion. AB - PURPOSE: An estimation and production paradigm was used to determine whether clinically normal 8- to 12-yr-old female (N = 18) and male (N = 18) children could (a) self-regulate intermittent cycle ergometer exercise using a prescribed target rating of perceived exertion (RPE), (b) discriminate between target RPEs, and (c) produce intermittent target RPEs in both an ascending and descending sequence. METHODS: Overall body RPE was assessed with the Children's OMNI Scale (0-10). Subjects underwent (a) one orientation trial, (b) one estimation (E) trial, and (c) two production (P) trials. During E, RPE was estimated each minute of a progressive cycle ergometer test. During the 3-min intermittent P trials, subjects titrated cycle brake force to produce either an RPE sequence of 2 and 6 (ascending) or 6 and 2 (descending). The P trials simulated short, intermittent exercise typical of children's play. RESULTS: Oxygen uptake (VO2) did not differ between E and P at a target RPE of 2 (0.63 versus 0.66 L x min(-1)) and 6 (1.27 vs 1.21 L x min(-1)). Heart rate (HR) did not differ between E and P at a target RPE of 2 (104.1 vs 102.6 beats.min-1) and 6 (153.7 vs 154.5 beats x min(-1)). Both VO2 and HR were higher (P < 0.01) at a target RPE-6 than -2. Responses were not affected by gender or production sequence. CONCLUSION: Young female and male children were able to use the OMNI Scale to self-regulate short-duration intermittent cycle exercise intensity. PMID- 12131259 TI - Effects of creatine on isometric bench-press performance in resistance-trained humans. AB - PURPOSE: The purpose of this study was to investigate the effects of creatine (Cr) supplementation on force generation during an isometric bench-press in resistance-trained men. METHODS: 32 resistance-trained men were matched for peak isometric force and assigned in double-blind fashion to either a Cr or placebo group. Subjects performed an isometric bench-press test involving five maximal isometric contractions before and after 5 d of Cr (20 g.d-1 Cr + 180 g.d-1 dextrose) or placebo (200 g.d-1 dextrose). Body composition was measured before and after supplementation. Subjects completed 24-h urine collections throughout the study period; these were subsequently analyzed to provide total Cr and creatinine excretion. RESULTS: The amount of Cr retained over the supplementation period was 45 +/- 18 g (mean +/- SD), with an estimated intramuscular Cr storage of 43 (13-61) mmol x kg(-1) x dry weight muscle (median [range]). Four subjects in the Cr group were classified as "nonresponders" (< or =21 mmol x kg(-1) x dry weight muscle increase following Cr supplementation) and the remaining 17 subjects were classed as "responders" (> or =32 mmol x kg(-1) x dry weight muscle). For the Cr group, peak force and total force pre- or post supplementation were not different from placebo. However, when the analysis was confined to the responders, both the change in peak force [Repetition 2: 59(81) N vs -26(85) N; Repetition 3: 45(59) N vs -26(64) N) and the change in total force (Repetition 1: 1471(1274) N vs 209(1517) N; Repetition 2: 1575(1254) N vs 196(1413) N; Repetition 3: 1278(1245) N vs -3(1118) N; Repetition 4: 918(935) N vs -83(1095) N] post-supplementation were significantly greater compared with the placebo group (P < 0.01). For the Cr group, estimated Cr uptake was inversely correlated with training status (r = -0.68, N = 21, P = 0.001). Cr significantly increased body weight (84.1 +/- 8.6 kg pre- vs 85.3 +/- 8.3 kg post supplementation) and fat-free mass (71.8 +/- 6.0 kg pre- vs 72.6 +/- 6.0 kg post supplementation), with the magnitude of increase being significantly greater in the responder group than in the placebo group. CONCLUSION: Five days of Cr supplementation increased body weight and fat-free body mass in resistance trained men who were classified as responders. Peak force and total force during a repeated maximal isometric bench-press test were also significantly greater in the responders compared to the placebo group. PMID- 12131260 TI - Dose effects of oral bovine colostrum on physical work capacity in cyclists. AB - PURPOSE: There is interest in the potential long-term use of dietary supplementation with bovine colostrum to enhance exercise performance. The purpose of the present study was to determine the dose effects of bovine colostrum on cycling performance. METHODS: Forty-two competitive cyclists were randomly divided into three groups and required to consume either 20 g/d bovine colostrum + 40 g whey protein concentrate (wpc), 60 g of bovine colostrum, or 60 g of wpc (placebo). Two measures were used to assess performance before (pre-) and after (post-) an 8-wk supplementation period. The first measure required subjects to complete two VO2max tests separated by 20 min with the amount of work completed in the second test used to evaluate performance. The second performance measure was the time to complete a work-based time trial following a 2-h cycle at 65% VO2max. Subjects were required to maintain their regular training and keep a food and training diary over the study period. RESULTS: After supplementation, the performance enhancement in Measure One was not statistically significantly different in the colostrum groups compared to the placebo group (placebo = 3.4%, 20 g = 4.0%, 60 g = 3.9%; 95% confidence interval (CI) for differences, +/-1.8%, P > 0.05). In performance Measure Two subjects in the 20 g and 60 g groups completed the time trial significantly (P < 0.05) faster post supplement compared to pre supplement (improvements in performance times, placebo = 37 s, 20 g = 158 s, 60 g = 134 s; 95% CI for differences, 47 s). CONCLUSION: Oral bovine colostrum supplementation at 20 g or 60 g/d provided a small but significant improvement in time trial performance in cyclists after a 2-h ride at 65% VO2max. PMID- 12131261 TI - No effect of a noradrenergic reuptake inhibitor on performance in trained cyclists. AB - INTRODUCTION: According to the central fatigue hypothesis, serotonin (5-HT) is related to fatigue, whereas the noradrenergic system is primarily concerned with arousal and motivation, and therefore hypothesized to enhance performance. The purpose of the present study was to examine the effects of a selective noradrenergic reuptake inhibitor (reboxetine 2 x 4 mg REB-NARI) on exercise performance. METHODS: Seven healthy well-trained male cyclists (age: 23 +/- 1.7 yr, height: 182 +/- 5.8 cm, weight: 73.5 +/- 8.5 kg, VO2max: 73.5 +/- 6.4 mL x kg(-1) x min(-1), Watt(max): 376 +/- 11.7 W) participated to the study. Subjects completed two endurance tests (time trials) starting at 65% Wmax in a double blind randomized cross-over design. Blood samples were collected for adrenocorticotropin, prolactin, cortisol, growth hormone (GH), beta-endorphins, and catecholamines and were taken at 30-min time intervals until the end of exercise. Performance was analyzed with a paired t-test, whereas data for hormonal and metabolic differences during the trials were analyzed using an ANOVA repeated measures design and an LSD-planned comparisons test. Significance level was set at P < 0.05. RESULTS: Performance was not influenced by the NARI (REB: 97 min +/- 3 min, placebo (PLAC): 92 min +/- 1 min). All hormones increased during exercise except for GH in the REB trial, which was significantly lower than PLAC. The other hormones were significantly higher in the REB trial versus the PLAC trial at the end of exercise and during recovery. CONCLUSION: In conclusion, the results demonstrate that the drug had a central effect. In particular, the higher resting GH concentrations indicated a marked and selective noradrenergic effect of REB. However, performance was not influenced by a selective NARI in well trained endurance athletes. PMID- 12131262 TI - Inspiratory muscle training fails to improve endurance capacity in athletes. AB - PURPOSE: The purpose of this study was to examine the effects of specific inspiratory muscle training (IMT) on respiratory muscle strength and endurance and whole-body endurance exercise capacity in competitive endurance athletes. METHODS: Seven collegiate distance runners (5 male/2 female; VO2max = 59.9 +/- 11.7 mL.kg-1.min-1) were recruited to participate in this study. Initial testing included maximal oxygen consumption (VO2max), sustained maximal inspiratory mouth pressure (MIP), breathing endurance time (BET) at 60% MIP, and endurance run time (ERT) at 85% VO2max. Heart rate (HR), minute ventilation (VE), oxygen consumption (VO2), and ratings of perceived dyspnea (RPD) were recorded at 5-min intervals and during the last minute of the endurance run. Blood lactate concentration (BLC) was also obtained immediately before and at 2 min after the endurance run. All testing was repeated after 4 wk of IMT (50-65% MIP, approximately 25 min x d( 1), 4-5 sessions/week, 4 wk). RESULTS: After 4 wk of IMT, MIP and BET were significantly increased compared with pretraining values (P < 0.05). No significant differences between pre and post values were observed in VO2max or ERT at 85% VO2max after IMT. No significant differences between pre and post values were detected in HR, VE, VO2, or RPD during the endurance run as measured at steady state and end of the test after IMT. BLC was not significantly different before or at 2 min after the endurance run between pre and post IMT. CONCLUSION: These results suggest that IMT significantly improves respiratory muscle strength and endurance. However, these improvements in respiratory muscle function are not transferable to VO2max or endurance exercise capacity as assessed at 85% VO2max in competitive athletes. PMID- 12131263 TI - Muscle function in elite master weightlifters. AB - PURPOSE: To determine whether explosive power and isometric strength of the lower limb muscles in elite master Olympic weightlifters declines at a similar rate to nontrained healthy controls with increasing age. METHODS: 54 elite level masters weightlifters (aged 40-87), who were competitors at the World Masters Weightlifting Championships (1999), were compared with a similar number of aged matched, healthy untrained individuals. Isometric knee extensor strength and lower-limb explosive power were tested. Extent of antagonist co-contraction during isometric knee extension was determined by EMG and power loading characteristics by using a variable inertial system. Muscle volume was estimated using anthropometry. RESULTS: On average, the weightlifters were able to generate 32% more peak power (P < 0.05) in the lower limbs and 32% more isometric knee extensor force (P < 0.05) than the control subjects. No significant differences in lower-leg volume were observed between the two groups. Peak power declined at a similar rate with increasing age in the weightlifters and controls (1.2 and 1.3% of a 45-yr-old's value per year), as did strength, but at a lower rate (0.6 and 0.5% per year). The inertial load at which the weightlifters achieved their maximal peak power output was greater (P < 0.05) than the controls. The torque generated at this optimal inertia was also greater in the weightlifters (P < 0.05), whereas the time taken for the weightlifters to reach their maximal peak power was on average 13% shorter (P < 0.05). No differences in antagonist co contraction during isometric knee extension were observed between the two groups. CONCLUSIONS: Muscle power and isometric strength decline at a similar rate with increasing age in elite master weightlifters and healthy controls. In spite of inertial load optimization, muscle power declined in both groups at approximately twice the rate of isometric strength. Although similar rates of decline were observed, the absolute differences between the weightlifters and controls were such that an 85-yr-old weightlifter was as powerful as a 65-yr-old control subject. This would therefore represent an apparent age advantage of approximately 20 yr for the weightlifters. PMID- 12131264 TI - Cerebral hemodynamics and resistance exercise. AB - PURPOSE: Repetitive resistance exercise with large muscle mass causes rapid fluctuations in mean arterial blood pressure (MAP). We sought to determine the effect of these fluctuations on the cerebrovasculature response determined by mean flow velocity (Vmean) of the middle cerebral artery. METHODS: Nine subjects performed 10-repetition maximum leg press exercise. MAP was estimated by finger photoplethysmography, Vmean by Doppler ultrasound, and end-tidal CO2 (PETCO2) by mass spectrometry. RESULTS: Vmean fluctuated with MAP with each repetition however averaged over the 10 repetitions, Vmean was unchanged from resting baseline values (66.9 +/- 10.8 vs 67.7 +/- 12.3 cm.s-1, baseline vs exercise, P > 0.05) despite an increased MAP (89.5 +/- 8.4 vs 105.0 +/- 4.9 Torr, P < 0.05). PETCO2 also remained unchanged from rest to exercise (37.7 +/- 2.8 vs 36.6 +/- 2.7 Torr, P > 0.05). Vmean decreased below resting levels for the first 5 s of recovery (59.8 +/- 9.1 cm.s-1, P < 0.05) as MAP returned rapidly to slightly below baseline (83.3 +/- 6.1, P > 0.05). MAP/Vmean, an index of cerebrovascular resistance, was elevated during exercise and returned to baseline after exercise. An increase in Vmean at 30 s post exercise (78.4 +/- 10.6 cm.s-1, P < 0.05) corresponded with elevated PETCO2 (43.0 +/- 4.8 Torr, P > 0.05). CONCLUSION: The results suggest that fluctuations in MAP with individual muscle contractions during resistance exercise appear to be too rapid to be countered by cerebrovascular autoregulation. However, the progressive increase in MAP over a number of contractions was effectively countered to maintain Vmean near baseline values before a decrease in Vmean immediately after exercise. PMID- 12131265 TI - The team physician and return-to-play issues: a consensus statement. PMID- 12131266 TI - VO2max and lactate production are not normal in all patients with chronic fatigue. PMID- 12131267 TI - The Arizona Activity Frequency Questionnaire using doubly labeled water. PMID- 12131270 TI - Should ECG monitoring during the postanesthesia period be the standard of care for all women who have cesarean birth? PMID- 12131271 TI - Should ECG monitoring during the postanesthesia period be the standard of care for all women who have cesarean birth? PMID- 12131272 TI - Mothering multiples: a meta-synthesis of qualitative research. AB - Increasing numbers of qualitative studies in maternal-child nursing are being published. However, clinical application and knowledge development based on those studies will be hampered unless the rich understandings gleaned from these individual studies can be synthesized. Meta-synthesis is one technique to help accumulate knowledge from qualitative research. The first section of this article explains the technique of meta-synthesis and reviews meta-syntheses published in nursing. The focus then becomes an illustration of a meta-synthesis in maternal child nursing: Mothering multiples during the first year of life. Six qualitative studies comprised the sample for the meta-synthesis. The meta-synthesis revealed a shared set of five themes that help increase our understanding of mothering multiples: "bearing the burden," "riding an emotional roller coaster," "lifesaving support," "striving for maternal justice," and "acknowledging individuality." Implications for practice derived from this meta-synthesis are addressed. PMID- 12131273 TI - Touchpoints: changing the face of pediatric nurse practitioner education. AB - Touchpoints is an interdisciplinary relational model of healthcare primarily used with parents and young children. The underlying premise of the Touchpoints approach is to support the parent/child relationship during the health encounter by enhancing parents' efforts to optimize their child's physical and psychological development. Nurse practitioners who use this approach in practice find they are able to connect quickly to the parents' most urgent concerns for their child. Our experience has been that a pediatric nurse practitioner program that uses Touchpoints as the underlying framework can assist students in achieving a holistic view of families by focusing the curriculum more directly on development and relationships. Students learn that building a relationship with parents, and joining them in the care of their child, produces an atmosphere in the health encounter of mutual respect and trust. Parents leave the encounter feeling satisfied their concern for their child has been heard and questions have been seriously discussed; students leave feeling competent and valued by their patients. Touchpoints provides a model for teaching and demonstrating the development of interpersonal relationships by using the language of the child's behavior. PMID- 12131274 TI - Kangaroo care for adoptive parents and their critically ill preterm infant. AB - In this case study kangaroo care (KC) was facilitated for an adoptive mother and father who were planning to attend the birth of the infant they had arranged to adopt. Unexpectedly, the birth mother delivered at 27 weeks gestation. The infant was critically ill and required mechanical ventilation. However, in this neonatal intensive care unit where all adoptive parents and parents of mechanically ventilated infants are offered KC, these adoptive parents began KC on Day 3 while their infant daughter was still mechanically ventilated. She thrived thereafter and the entire experience was profoundly beneficial for this beginning family both at the hospital and after discharge home. PMID- 12131275 TI - Perceived health status in urban minority young adolescents. AB - PURPOSE: To describe perceptions of health status among a sample of urban minority adolescents and the contribution of demographics, intrinsic motivation, general self-efficacy, risk taking, and stressful life experiences on the adolescent's perception of health status. METHOD: Correlational design. A total of 71 adolescents were studied using the Adolescent Health Chart for perceived health status, the Health Self-Determinism Index for Children, the Self-Efficacy Scale, the Risk Taking Instrument, and the Life Events Checklist. RESULTS: There were no statistically significant effects of demographics on perceived health status. Scores of Perceived Health Status correlated with scores of self-efficacy (r = 0.56; p <.0001), risk-taking (r = 0.39; p <.001), negative events (r = 0.32; p <.01) and violent events (r = -0.41; p <.001). CLINICAL IMPLICATIONS: The study findings suggest that self-efficacy, risk taking, and life events are predictive of perceived health status in urban minority adolescents. The results contribute to the present body of knowledge about patterns of adolescent health as perceived by the adolescent. In addition to expanding the understanding of the minority adolescent experience in relation to health promotion attributes and health compromising behaviors, the results identify antecedents that are predictive of improved perceived health status for the urban adolescent. PMID- 12131276 TI - Management of fetal airway obstruction: an innovative strategy. AB - This article describes a planned ex utero intrapartum treatment (EXIT) procedure at the Children's Hospital of Philadelphia for a fetus with an airway obstruction resulting from a giant neck mass. The EXIT procedure is a technique that establishes a fetal airway while the utero-placental circulation is maintained for up to 1 hour. As a part of the planned EXIT procedure, a multidisciplinary, highly skilled team was developed to care for both mother and baby. This team consisted of obstetric and surgical personnel to care for the mother during the procedure, the birth, and the recovery, and a neonatal surgical team to care for the newborn. Nursing expertise necessary to conduct this procedure and safely care for the woman and fetus are discussed. PMID- 12131277 TI - "Waiting" as experienced by women hospitalized during the antepartum period. AB - PURPOSE: To better understand the lived experience of "waiting" among women who are hospitalized during the antepartum period. STUDY DESIGN AND METHODS: Phenomenology based on Parse's theory of human becoming. Audiotapes were made of dialogues between the participants and the researcher. Participants were asked to respond to this question: "Tell me what your experience of 'waiting' is like." The sample consisted of 14 hospitalized women. RESULTS: Unpredictable as well as paradoxical realities of waiting evolved that differed person to person, yet each participant's dialogue reflected waiting as an enduring vigil of burdening toil while engaging-disengaging with close others in cherishing the can-be. CLINICAL IMPLICATIONS: Listening to what antepartum women really feel about waiting can give direction to nursing care. Women wanted to know that nurses supported their efforts to carry their babies to full term. This study helps us to see that although most of western society does not value waiting, perhaps there is unseen value in waiting. The women in this study realized the agony of waiting, but understood its importance for the health of their babies. Research such as this which encourages active listening and reflecting on women's stories can help to improve nursing practice and healthcare for women. PMID- 12131278 TI - Electronic fetal monitoring and the legal medical record. PMID- 12131279 TI - The antimicrobial defense mechanism of the female urethra: a reassessment. AB - PURPOSE: This review of the literature and study of the cytology of the urethra were done to define the potential role of the female urethra as a staging site for urinary tract infection and examine the evidence for a urethral defense mechanism. MATERIALS AND METHODS: We re-analyzed data on the quantitative microbiology of the female urethra published 3 decades ago, reviewed the literature on the initiation of ascending urinary tract infections, the cytology and anatomy of the urethra, and performed studies of the morphology of urethral cells in boys and girls, men, menstruating and menopausal women, and women with acute cystitis. We also considered clues about the urethral microenvironment provided by gonococcal cervicitis and urethritis. RESULTS: We found strong statistical evidence that the female urethra has a powerful antimicrobial defense mechanism, which appears to differ in women with and without recurrent urinary tract infections. We corroborated the findings of previous investigators that the female urethra is lined by cells identical to those of the vagina that respond similarly to estrogens. We found immature basal and parabasal cells in children, and a modest inflammatory response to urinary tract infection. CONCLUSIONS: The female urethra may provide a favorable environment for colonization by uropathogens but it is protected by a powerful defense mechanism. This mechanism may be explained by the shedding of uropathogens bound to exfoliating urethral cells, trapping of bacteria by mucus secreted by the paraurethral glands, intermittent washout by urine, local production of Ig, cytokines and defensins and mobilization of leukocytes. PMID- 12131280 TI - Asymptomatic infections of the urinary tract. PMID- 12131281 TI - Wolff and Muller: fundamental eponyms of embryology, nephrology and urology. AB - PURPOSE: Kaspar Friedrich Wolff and Johannes Petrus Muller, German physicians in the 18th and 19th centuries, respectively, contributed significantly to the study of the biological sciences, specifically embryology and the development of the genitourinary tract. Their eponyms are a part of daily conversation in medical education, practice and research. We reviewed their lives and works. MATERIALS AND METHODS: Commentaries on the works of these scientists as well as available English translations of their works were analyzed. RESULTS: Wolff and Muller were pioneers in their fields. Wolff gained renown for his Theoria Generatonis, a great embryological treatise, produced in 1759. Muller performed significant studies in the fields of anatomy, pathology, physiology and embryology. In 1834 he became editor of the physiology journal that later came to be known as Muller's Archiv. CONCLUSIONS: Studies of and inferences about the development of the urogenital system a century apart by these 2 men contributed greatly to the current understanding of embryology. Their names remain a part of everyday medical dialogue. PMID- 12131282 TI - Bicalutamide as immediate therapy either alone or as adjuvant to standard care of patients with localized or locally advanced prostate cancer: first analysis of the early prostate cancer program. AB - PURPOSE: We determine the efficacy and tolerability of bicalutamide as immediate therapy, either alone or as adjuvant to treatment of curative intent, in patients with clinically localized or locally advanced prostate cancer. MATERIALS AND METHODS: This international program consists of 3 ongoing, randomized, double blind, placebo controlled clinical trials (trials 23, 24, and 25). Men with localized or locally advanced (T1-T4, Nx/N0, M0) prostate cancer were randomized to receive 150 mg. bicalutamide daily or placebo, in addition to standard care with radical prostatectomy, radiotherapy or watchful waiting. Primary end points are time to objective progression and overall survival. In this first analysis data from the trials were combined in a single overview analysis according to protocol. RESULTS: Data are available for 8,113 patients (4,052 randomized to bicalutamide, 4,061 to standard care alone) at a median followup of 3.0 years. Treatment with bicalutamide provided a highly significant reduction of 42% in the risk of objective progression compared with standard care alone (9.0% versus 13.8%, hazards ratio 0.58; 95% confidence interval 0.51, 0.66; p <<0.0001). The overall result was reflected in 2 of the 3 trials (trials 24 and 25) with trial 3 (trial 23) showing a nonsignificant difference at this time. Reductions in the risk of disease progression were seen across the entire patient population, irrespective of primary treatment or disease stage. Overall survival data are currently immature and longer followup will determine if there is also a survival benefit with bicalutamide. The most frequently reported side effects of bicalutamide were gynecomastia and breast pain. CONCLUSIONS: Immediate treatment with 150 mg. bicalutamide daily, either alone or as adjuvant to treatment of curative intent, significantly reduces the risk of disease progression in patients with localized or locally advanced prostate cancer. This benefit must be balanced with the morbidity associated with long-term hormonal therapy. Followup is ongoing to determine potential survival benefits of this treatment approach. PMID- 12131283 TI - Erbium: YAG laser lithotripsy mechanism. AB - PURPOSE: We tested the hypothesis that the mechanism of long pulse erbium:YAG laser lithotripsy is photothermal. MATERIALS AND METHODS: Human urinary calculi were placed in deionized water and irradiated with erbium:YAG laser energy delivered through a sapphire optical fiber. Erbium:YAG bubble dynamics were visualized with Schlieren flash photography and correlated to acoustic emissions measured by a polyvinylidene fluoride needle hydrophone. The sapphire fiber was placed either parallel or perpendicular to the calculus surface to assess the contribution of acoustic transients to fragmentation. Stones were irradiated using desiccated stone irradiated in air, hydrated stone irradiated in air and hydrated stone irradiated in water. Ablation crater sizes were compared. Uric acid stones were irradiated in water and the water was assayed for cyanide. RESULTS: During the early phase of vapor bubble expansion, acoustic transients had minimal effects on calculus fragmentation. Fragmentation occurred due to direct absorption of laser energy transmitted to the calculus through the vapor channel between the sapphire fiber tip and calculus. The forward axial expansion of the bubble occurred more rapidly than the radial expansion. A parallel oriented fiber on the calculus surface produced no fragmentation but generated larger amplitude acoustic transients compared to perpendicular orientation. In perpendicular orientation the erbium:YAG laser did not generate any collapse acoustic waves but fragmentation occurred. Crater width was greatest for desiccated stones irradiated in air (p <0.03). Cyanide production increased as erbium:YAG irradiation of uric acid calculi increased, (r2 = 0.98). CONCLUSIONS: The erbium:YAG laser fragments stones through a photothermal mechanism. PMID- 12131284 TI - Safety and efficacy of holmium: YAG laser lithotripsy in patients with bleeding diatheses. AB - PURPOSE: We assessed the safety and efficacy of ureteroscopy and holmium:YAG laser lithotripsy for treating upper urinary tract calculi in patients with known and uncorrected bleeding diathesis. MATERIALS AND METHODS: We retrospectively reviewed the charts at 2 tertiary stone centers to identify patients with known bleeding diathesis who were treated with holmium:YAG laser lithotripsy for upper urinary tract calculi. A total of 25 patients (29 upper urinary tract calculi) underwent ureteroscopic holmium laser lithotripsy. Bleeding diathesis involved warfarin administration for various conditions in 17 patients, liver dysfunction in 3, thrombocytopenia in 4 and von Willebrand's disease in 1. The mean international normalized ratio, platelet count and bleeding time were 2.3, 50 x 109/l. and greater than 16 minutes in patients on warfarin and in those with liver dysfunction, thrombocytopenia and von Willebrand's disease, respectively. RESULTS: Overall the stone-free rate was 96% (27 of 28 cases) and 29 of 30 procedures were completed successfully without significant complication. In a patient treated concomitantly with electrohydraulic lithotripsy significant retroperitoneal hemorrhage required blood transfusion. CONCLUSIONS: Upper tract urinary calculi in patients with uncorrected bleeding diathesis can be safely managed by contemporary small caliber ureteroscopes and the holmium laser as the only modality of lithotripsy. Ureteroscopic holmium laser lithotripsy without preoperatively correcting hemostatic parameters limits the risk of thromboembolic complications and costs associated with an extended hospital stay. Avoiding electrohydraulic lithotripsy is crucial for decreasing bleeding complications in this cohort of patients. PMID- 12131285 TI - Noninvasive anesthesia, analgesia and radiation-free extracorporeal shock wave lithotripsy for stones in the most distal ureter: experience with 165 patients. AB - PURPOSE: Spontaneous ureteral stone passage often causes severe renal colic, especially when the stone passes through the narrow ureteral orifice. In these situations noninvasive anesthesia-free, analgesia-free and radiation-free extracorporeal shock wave lithotripsy (ESWL) (Dornier Medical Systems, Marietta, Georgia) is a valuable tool. It can be performed at any time without needing any further patient preparation. MATERIALS AND METHODS: A total of 165 patients underwent ESWL using the Lithostar Ultra device (Siemens, Erlangen, Germany). Only ureteral calculi within 5 cm. of the ureterovesical junction were included in this study. Patients were treated while supine and stones were localized by ultrasound through the filled bladder without x-ray exposure. Treatment was started without anesthesia or analgesia and analgesics were administered only at patient request during treatment. RESULTS: Of the patients 93% were treated without anesthesia or analgesia and 7% required a single intravenous dose of 25 mg. pethidine. Postoperatively renal colic developed in 40 patients (24%). In 4 cases (2.4%) renal drainage was required for analgesia resistant pain or obstructive pyelonephritis. On day 1 after ESWL 90% of the patients were stone free or had fragments 2 mm. or less, while 10% had residual fragments 3 mm. or greater. Of all patients 7% were re-treated once. At 3 months postoperatively 129 of the 130 evaluable patients (99%) were stone-free. CONCLUSIONS: ESWL of stones located in the most distal ureter using the Lithostar Ultra device is effective, safe and radiation-free. It is done without anesthesia and in most cases without analgesics. This simple and noninvasive procedure is an excellent first line treatment modality for prevesical stones and it represents a valid alternative to conservative management or invasive endoscopy. PMID- 12131286 TI - Evaluation of disintegration in prevesical ureteral calculi by 3-dimensional endo ultrasound with surface rendering. AB - PURPOSE: Lower ureteral calculi are accessible by transrectal endo-ultrasound, including 3-dimensional image processing with surface rendering. We analyzed the efficiency of this innovative technology compared with that of standard radiographic examinations with respect to the outcome of extracorporeal shock wave lithotripsy (ESWL) (Dornier Medical Systems, Marietta, Georgia). MATERIALS AND METHODS: In patients with prevesical or intramural calculi we performed excretory urography and transrectal endo-ultrasound using a Combison 530 device (Kretz Technik, Zipf, Austria) with integrated optional 3-dimensional image processing and surface rendering. A total of 102 ESWL cases (108 radiopaque stones, 145 ESWL sessions) were included in the study. One day after ESWL an evaluation was independently performed by a radiologist (plain x-ray) and a urologist (transrectal endo-ultrasound). Results were compared to the outcome determined during further followup. RESULTS: All stones were evaluable by endo ultrasound with surface rendering, whereas in 27 cases the stone was hidden by extraureteral processes on followup radiography. Immediately after endo ultrasound 8 calculi were passed. In 7 of the remaining 110 cases (6.4%) radiographic examination did not confirm sonographic findings. In these cases evaluation by surface rendering proved to be correct on followup. CONCLUSIONS: Endo-ultrasound with surface rendering proved to be highly effective for evaluating ESWL success in cases of prevesical ureteral stones. This technique is independent from bowel gas or other factors that impede radiological imaging. It is safe, easy to learn, well tolerated by patients and does not expose them to radiation. PMID- 12131287 TI - Renal cell carcinoma in the solitary kidney: an analysis of complications and outcome after nephron sparing surgery. AB - PURPOSE: We evaluated surgical techniques, pathological features and extended outcomes in patients with renal cell carcinoma in a solitary kidney treated with surgical excision. MATERIALS AND METHODS: Between 1970 and 1998, 76 patients underwent nephron sparing surgery for sporadic renal cell carcinoma in a solitary kidney, including 63 with tissue specimens available for pathological review who comprised the cohort. Six (9.5%) patients had a congenitally absent kidney and 57 (90.5%) had previously undergone contralateral nephrectomy for renal cell carcinoma. The clinical and pathological features examined were patient age at nephron sparing surgery, sex, type of nephron sparing surgery (enucleation, partial nephrectomy or ex vivo resection), tumor size, nuclear grade, histological subtype and 1997 tumor stage. Overall cancer specific, local recurrence-free and metastasis-free survival as well as early (within 30 days of nephron sparing surgery) and late (30 days to 1 year after nephron sparing surgery) complications were assessed. Univariate and multivariate analyses were done to test for the associations of clinical and pathological features with outcome. RESULTS: Most patients were treated with enucleation (36.5%), standard partial nephrectomy (38.1%) or the 2 procedures (11.1%) and in 8 (12.7%) ex vivo tumor resection was done. The renal cell carcinoma histological subtypes were clear cell in 82.5% of cases, papillary in 15.9% and chromophobe in 1.6%. Grade was 1 to 3 in 10 (15.9%), 42 (66.7%) and 10 (15.9%) tumors, respectively. At 5 and 10 years the overall survival rate was 74.7% and 45.8%, the cancer specific survival rate was 80.7% and 63.7%, the local recurrence-free survival rate was 89.2% and 80.3%, and the metastasis-free survival rate was 69% and 50.4%, respectively. Tumor stage and nuclear grade were significantly associated with death from any cause, death from renal cell carcinoma and distant metastases on multivariate analysis. Notably no patient with papillary or chromophobe renal cell carcinoma died of renal cell carcinoma, or had recurrence or metastasis. The type of nephron sparing surgery was not significantly associated with outcome, although there were too few patients with recurrence to assess the association of the type of nephron sparing surgery with local recurrence. The most common early complication was acute renal failure in 12.7% of cases, while the most common late complications were proteinuria in 15.9% and renal insufficiency in 12.7%. CONCLUSIONS: The 1997 tumor stage and nuclear grade were significant predictors of death from any cause, death from renal cell carcinoma and distant metastases in patients treated with nephron sparing surgery for renal cell carcinoma involving a solitary kidney. Nephron sparing surgery in a solitary kidney can be performed safely and with minimal morbidity. PMID- 12131288 TI - Primary apoptosis as a prognostic index for the classification of metastatic renal cell carcinoma. AB - PURPOSE: We identified novel biological markers of prognosis in primary histopathological specimens from patients with metastatic renal cell carcinoma. MATERIALS AND METHODS: Apoptotic indexes (terminal deoxynucleotidyl transferase mediated deoxyuridine triphosphate nick end-labeling), proliferation rates (Ki-67 antigen), p21 (WAF1/cip1) expression and CD95 (APO-1/Fas) expression were determined in paraffin embedded nephrectomy specimens from 73 patients with histologically confirmed, progressive metastatic disease. Kaplan-Meier survival analysis, log rank statistics and 2-proportional Cox regression analysis were done to identify new risk factors in addition to conventional classification criteria, and demonstrate statistical independence. RESULTS: Multivariate analysis indicated that primary tumor apoptosis (p = 0.0116) and the interval from diagnosis to metastatic disease (p = 0.002) had a high predictive impact on overall survival after initial diagnosis. Patients were assigned to 2 risk groups, namely a poor prognosis group with a median survival of 20 months, defined by apoptosis less than 6% in the primary tumor nephrectomy specimen and a time from initial diagnosis to metastatic disease of less than 6 months, and a good prognosis group with a median survival of 56 months, defined as the absence of 1 or 2 risk factors. CONCLUSIONS: Our findings showed that primary tumor apoptosis is a novel independent predictor in patients with metastatic renal cell carcinoma at initial diagnosis. It leads to a new prognostic index in the pretreatment classification of metastatic renal cell carcinoma. PMID- 12131289 TI - Comparative evaluation of the nuclear matrix protein, fibronectin, urinary bladder cancer antigen and voided urine cytology in the detection of bladder tumors. AB - PURPOSE: We evaluate the diagnostic efficacy of nuclear matrix protein-22 (NMP22, Matritech, Newton, Massachusetts), fibronectin and urinary bladder cancer antigen (UBC, IDL Biotech, Borlange, Sweden) compared with voided urine cytology in the detection of bladder cancer. MATERIALS AND METHODS: A total of 168 patients provided a single voided urine sample for NMP22, fibronectin an ideal monoclonal for urinary bladder cancer and cytology before cystoscopy. Cystoscopy was done for all patients as the reference standard for identification of bladder cancer. Biopsy of any suspicious lesion was performed for histopathological examination. Of the 168 cases 100 were histologically diagnosed as bladder cancer, whereas the remaining 68 had benign urological disorders. A group of 47 healthy volunteers were also enrolled in this study. Voided urine was evaluated by NMP22, fibronectin and UBC, and their values were expressed relative to mg. creatinine. RESULTS: The optimal threshold values for NMP22, fibronectin and UBC were calculated by receiver operator characteristics curves as 27 units per mg. creatinine, 198 mg./mg. creatinine and 13 ng./mg. creatinine, respectively. The levels and positive rates of the 3 parameters were significantly higher in the malignant group compared to either the benign group or normal controls. Of the entire group NMP22, fibronectin and UBC were positive in 93.2%, 91% and 68.2%, respectively in bladder cancer cases with positive cytology. Moreover, these positive rates were significantly higher in bilharzial bladder cancer cases (58.8%, 67.5%, 58.8%, respectively) compared to nonbilharzial cases (35.6%, 36.3%, 31.1%). Overall sensitivity and specificity were 85% and 91.3% for NMP22, 83% and 82.6% for fibronectin, 67% and 80.8% for UBC and 44% and 100% for voided urine cytology. Combined sensitivity of voided urine cytology with the 3 biomarkers together was higher than either combined sensitivity of voided urine cytology with 1 of the biomarkers or than that of the biomarker alone. CONCLUSIONS: Our data indicate that NMP22 and fibronectin had superior sensitivities compared to UBC and voided urine cytology, while NMP22 and voided urine cytology had the highest specificities. The combined use of markers increased the sensitivity of cytology from 44% to 95.3%. The higher sensitivities of markers in bilharzial than nonbilharzial bladder cancer highlight their clinical use in screening patients with urinary bilharziasis. PMID- 12131290 TI - Clinical use of urinary markers for the detection and prognosis of bladder carcinoma: a comparison of immunocytology with monoclonal antibodies against Lewis X and 486p3/12 with the BTA STAT and NMP22 tests. AB - PURPOSE: The noninvasive detection of urothelial carcinoma remains challenging. We prospectively evaluated urine markers for bladder carcinoma. We compared the NMP22 (Matritech, Cambridge, Massachusetts) and BTA Stat (Bard Diagnostics, Redmond, Washington) tests with immunocytology using mAbs 486p3/12 and BG7 against Lewis X antigen. MATERIALS AND METHODS: The NMP22 and BTA Stat tests were performed in urine samples and immunocytology with mAbs 486p3/12 and BG7 staining were performed in bladder washing specimens in 146 samples of 115 patients undergoing transurethral resection for suspected bladder carcinoma (70) or 45 undergoing followup cystoscopy for a history of bladder carcinoma (76). Bladder carcinoma was detected in 54 patients, including stages pTa in 25, pT1 in 20, pT2 in 8 and carcinoma in situ in 1, while 61 had no evidence of bladder carcinoma. The cutoff was 10 units per ml. for the NMP22, 30% positive cells for 486p3/12 and 5% positive cells for the Lewis X tests. RESULTS: BTA Stat was positive in 65 samples (44.5%) and NMP22 was positive in 69 (47.3%). Immunocytology with mAbs 486p3/12 and BG7 against Lewis X was positive in 52 (35.6%) and 109 (74.7%) samples, respectively. Sensitivity was 70.3% for BTA Stat, 68.5% for NMP22, 68.5% for 486p3/12 and 94.4% for Lewis X. Specificity was 70.6% for BTA Stat, 65.2% for NMP22, 83.6% for 486p3/12 and 36.9% for Lewis X. Area under the receiver operating characteristics curve was 0.6804 for NMP22, 0.7226 for Lewis X and 0.8002 for 486p3/12. False-positive results on BTA Stat in 2 of 22 patients (9%), on NMP22 in 2 of 25 (8%), on 486p3/12 in 3 of 11 (27%) and on Lewis X in 4 of 43 (9.3%) were associated with tumor recurrence. Furthermore, negative results on BTA Stat in 2 of 39 patients (2%), on NMP22 in 2 of 36 (0.5%), on Lewis X in 0 of 18 (0%) and on 486p3/12 in 1 of 50 (2%) was associated with tumor recurrence during followup. CONCLUSIONS: Immunocytology with mAbs against Lewis X showed higher sensitivity than all commercially available tests evaluated. Because of its high sensitivity and high negative predictive value, it may be useful for screening in a high risk population. Patients with a false-positive 486p3/12 test results are at increased risk for tumor recurrence compared with those with negative results. PMID- 12131291 TI - Transurethral resection for bladder cancer using 5-aminolevulinic acid induced fluorescence endoscopy versus white light endoscopy. AB - PURPOSE: Endoscopy done under fluorescence induced by 5-aminolevulinic acid has proved to be a procedure with high sensitivity for detecting transitional cell carcinoma of the bladder. In this multicenter, parallel group, phase III study we compared 5-aminolevulinic acid fluorescence endoscopy guided transurethral bladder resection with transurethral bladder resection done using only white light endoscopy. The proportion of tumor-free resected cases in the 2 groups was evaluated. MATERIALS AND METHODS: After patient stratification according to participating centers and European Organization for the Research and Treatment of Cancer risk score 65 and 64 were randomized to the 5-aminolevulinic acid fluorescence and white light endoscopy groups, respectively. Residual tumor was evaluated in the 2 groups by repeat transurethral resection 10 to 14 days later. Analysis was performed according to the intent to treat principle with all patients randomized, followed by per protocol analysis. RESULTS: Intent to treat analysis revealed that in the white light endoscopy group 40.6% of cases were resected tumor-free at primary resection, whereas with 5-aminolevulinic acid fluorescence endoscopy guided transurethral resection 61.5% were resected tumor free (p <0014). On protocol analysis 46.9% patients in the white light and 67.3% in the 5-aminolevulinic acid fluorescence endoscopy groups were resected tumor free (p <0.031). No difference was noted in the 2 groups in regard to side effects or laboratory findings. CONCLUSIONS: The risk of residual tumor after transurethral resection of transitional cell carcinoma is significantly decreased by 5-aminolevulinic acid fluorescence endoscopy. PMID- 12131292 TI - Prostate cancer stage shift has eliminated the gap in disease-free survival in black and white American men after radical prostatectomy. AB - PURPOSE: The initiation of prostate specific antigen (PSA) testing has led to increased public awareness, early detection and a stage shift in prostate cancer. We have previously reported that black American men have worse disease-free survival independently of pathological or clinical factors. We tested the stage shift effects on disease-free survival in our cohort of patients treated with radical prostatectomy. MATERIALS AND METHODS: A total of 1,042 consecutive patients underwent radical prostatectomy performed by Wayne State University full time faculty. The cohort was divided by the year of surgery as before (585 men in group 1) or after (457 in group 2) 1996. Clinicopathological and disease-free survival data were obtained from the Karmanos Cancer Institute multidisciplinary prostate cancer database. RESULTS: Improvements in clinical stage, preoperative PSA and biopsy Gleason score were observed in group 2 (p = 0.0001). Pathological organ confined disease increased in group 2 versus 1 in the 2 races, including 89 of 153 (58%) from 66 of 178 (37%) in black men and 189 of 304 (62%) from 194 of 407 (48%) in white men (p = 0.003 and 0.001, respectively). Calculated cancer recurrence-free median probability in group 1 at 42 months was 81% and 68% in white and black men, respectively (log rank test p = 0.001). These differences became insignificant in group 2 patients at 42 months with a median probability of 90% and 88% in white and black men, respectively (log rank test p = 0.39), representing a net increase in disease-free survival of 20% in black men. Specimen Gleason score, PSA and pathological stage were independent predictors of survival in the 2 groups. In contrast, race was an independent predictor only in group 1. CONCLUSIONS: The increased rate of pathological organ confined disease is translating into improved disease-free survival rates. These early data suggest that the survival gap in black and white American men is narrowing and may become statistically insignificant. PMID- 12131293 TI - Targeted screening for prostate cancer in high risk families: early onset is a significant risk factor for disease in first degree relatives. AB - PURPOSE: Targeted screening for prostate cancer in high risk families is generally suggested by ages 40 to 45 years in first degree relatives. We support this concept by reporting higher risk and earlier onset of the disease in these families. MATERIALS AND METHODS: We proposed serum prostate specific antigen (PSA) testing in 40 to 70-year-old first degree relatives of 435 patients with prostate cancer treated between July 1994 and June 1997. A previous systematic genealogical analysis allowed us to define the familial prostate cancer status of each patient as sporadic or familial. RESULTS: Of the 747 potential candidates 442 (59%) accepted into the study have been screened, including 240 who were 40 to 49 years old (mean age 44.8) and 202 who were 50 to 70 years old (mean age 57.4). Two of the 240 subjects (0.8%) had PSA greater than 4 ng./ml. in the 40 to 49-year-old group. Prostate biopsies were negative in 1 relative but diagnostic for prostate cancer in the other. In the 50 to 70-year-old group 25 of 202 subjects (12.4%) had a PSA of greater than 4 ng./ml. Prostate cancer was diagnosed in 9 individuals (4.5%), 9 had negative biopsy results, 1 died before biopsy and 6 refused biopsy. The proportion of relatives with PSA greater than 4 ng./ml. and prostate cancer detection was not different according to familial status (sporadic or familial) but it was significantly higher in first degree relatives with early onset prostate cancer in the family at ages younger than 65 years (p = 0.037 and 0.012, respectively). CONCLUSIONS: Our results emphasize the usefulness of PSA screening in high risk families, including those without obvious hereditary features. Furthermore, early onset prostate cancer is a significant risk factor for prostate cancer in first degree relatives. PMID- 12131294 TI - Prostate cancer is not increased in men with vasectomy in denmark. AB - PURPOSE: The risk of prostate cancer after vasectomy was studied using population based register data from Denmark. MATERIALS AND METHODS: Men hospitalized for vasectomy in Denmark from 1977 to 1989 were followed for death, emigration and incident cancer by administrative registers. The observed number of patients with cancer in the cohort was divided by the number expected based on the cancer incidence rate for all Danish men, providing a standardized incidence ratio. RESULTS: A total of 46 cohort patients were diagnosed with prostate cancer, while 46.93 were expected (standardized incidence ratio 0.98, 95% confidence interval 0.7 to 1.3). Time since vasectomy or age at vasectomy showed no trend. CONCLUSIONS: This population based Danish cohort study indicated no increase in the risk of prostate cancer in men hospitalized for vasectomy. The study was free of reporting bias since all data were collected from administrative registers. The cohort was not comprehensive but it most likely represented an unbiased sample of Danish men who underwent vasectomy. The study strongly adds to the evidence that there is no excess prostate cancer risk after vasectomy is done for sterilization. PMID- 12131295 TI - Contemporary trends in imaging test utilization for prostate cancer staging: data from the cancer of the prostate strategic urologic research endeavor. AB - PURPOSE: Previous investigators have reported widespread overuse of imaging tests for staging clinically localized prostate cancer. In this study imaging test utilization rates were analyzed in a contemporary group of patients, and clinical and demographic predictors of testing were identified. MATERIALS AND METHODS: Data were abstracted from the Cancer of the Prostate Strategic Urologic Research Endeavor (CaPSURE), a longitudinal registry of men with various stages of prostate cancer. A total of 4,966 men met study inclusion criteria of available treatment and staging data. The rates of computerized tomography, magnetic resonance imaging and bone scans performed between the dates of diagnosis and primary treatment were analyzed in patients at 3 levels of clinical risk based on serum prostate specific antigen, Gleason sum and T stage. Time trends in test utilization were analyzed by linear regression. Contemporary rates were compared with those identified in a previous analysis of an earlier CaPSURE cohort. Demographic and clinical predictors of utilization were identified using generalized linear model analysis. RESULTS: Since June 1997, the overall use of staging tests has decreased 63%, 25.9% and 11.4% in patients at low, intermediate and high risk, respectively. The most precipitous decrease was noted for bone scan but the use of cross-sectional imaging also decreased in all groups. Utilization rates were lower in 2001 than in any other year studied in CaPSURE. CONCLUSIONS: The rates of testing decreased significantly in all risk groups. However, in the absence of established clinical practice guidelines many patients at low and intermediate risk continue to undergo unnecessary testing, while a growing number of those at high risk are proceeding to treatment without previous imaging. PMID- 12131297 TI - Results of the 5 region prostate biopsy method: the repeat biopsy population. AB - PURPOSE: The decision to repeat prostate biopsy in a patient in whom the first biopsy did not detect prostate cancer poses a challenge to urologists. Many published series show a low yield on repeat biopsy using standard techniques. We reviewed our data on the 5 region prostate biopsy method to evaluate its yield in the repeat biopsy population. MATERIALS AND METHODS: A total of 125 repeat transrectal ultrasound guided prostate needle biopsy sessions were done in 110 patients for standard indications using the 5 region method. Pathological findings were reviewed and the yield of the additional regions was analyzed. RESULTS: Patients were categorized with respect to the initial biopsy technique. In those who underwent 1 and more than 1 previous sextant biopsy the relative increase in yield of the 5 region technique over the standard sextant technique was 31% and 33%, respectively. In the cohort that underwent previous 5 region biopsy the relative increase in yield of the 5 region technique over the standard sextant technique was 38%. CONCLUSIONS: In the setting of repeat biopsy the 5 region method results in an increased yield over the sextant method. It is true in patients who have previously undergone biopsies with the sextant or 5 region technique. PMID- 12131296 TI - Individual prostate biopsy core embedding facilitates maximal tissue representation. AB - PURPOSE: The probability of detecting small foci of prostate cancer is related to the amount of tissue represented. When multiple prostate biopsy cores are embedded in a single block, less tissue is evaluated because it is difficult to embed all cores in a single plane for optimal tissue representation. A computer simulation of sectioning biopsy cores was devised to examine the total surface area available at various angles of embedding and predict the ability to detect small tumor foci. MATERIALS AND METHODS: The computer simulation of biopsy core was done using commercially available software. Biopsy cores were represented as 3-dimensional cylindrical objects and the cutting blade was represented by a 2 dimensional plane. The intersection of the plane and cylinder represented the cut surface, which varied depending on cylinder angle and position. The simulation program calculates available surface area. RESULTS: Maximal surface area was obtained when the plane was horizontal to the long axis of the core. Any divergence of the cylinder from horizontal decreased the represented area. A single section through a 1 x 15 mm. biopsy core at 0 degrees yielded a surface area of 15 mm(2). The surface area was decreased to 13.3, 9.01 and 4.52 mm(2) at 3, 5 and 10 degrees, respectively. At a small focus of 0.6 mm. there was 100% detection under optimal circumstance, which decreased to 56.2% and 27.9% as the angle increased to 3 and 10 degrees, respectively. CONCLUSIONS: Optimal sectioning, that is maximal surface area, of the core is obtained when a biopsy core is sectioned at a 0-degree angle, that is horizontal to its long axis. It is much more likely when each biopsy core is embedded individually. When multiple cores are embedded together, it is difficult to position all cores in the same plane since cores move to different planes and the cut surface of the cylinders substantially decreases. Thus, for optimal surface representation and cancer detection embedding individual cores is appropriate. PMID- 12131298 TI - Percent free prostate specific antigen in the total prostate specific antigen 2 to 4 ng./ml. range does not substantially increase the number of biopsies needed to detect clinically significant prostate cancer compared to the 4 to 10 ng./ml. range. AB - PURPOSE: Percent free prostate specific antigen (PSA) is useful to select patients for prostate biopsy with total PSA 4 to 10 ng./ml. However, 20% of men with PSA between 2.6 and 4 ng./ml. harbor significant prostate cancer and percent free PSA has been suggested to aid in the decision to biopsy in this total PSA range as well. Concerns exist that the number of biopsies needed to detect 1 cancer in this range may be inappropriately high. In a prospective referral population we evaluated sensitivity and specificity of various percent free PSA cutoffs and determined the biopsy-per-cancer ratio in the PSA 2 to 4 ng./ml. range in men with a benign digital rectal examination, and report on the biological nature of the detected cancers based on Gleason score. Results were compared to those obtained from a reference group of patients (PSA 4 to 10 ng./ml., benign digital rectal examination) from the same prospective referral cohort. MATERIALS AND METHODS: Total PSA and free PSA were measured and percent free PSA was calculated. Of the initial 1,602 men 756 had a benign digital rectal examination and PSA 4 to 10 ng./ml., and 219 had a benign digital rectal examination and PSA 2 to 4 ng./ml. Sensitivity, specificity, the number of true positive (evidence of cancer) and false-positive (no evidence of cancer) biopsies were determined. The ratio of true positive biopsies-to-all biopsies performed was used to determine the biopsy-per-cancer ratio. Gleason score of the detected cancers was evaluated. The procedure was repeated for the PSA 4 to 10 ng./ml. range. RESULTS: In the PSA 4 to 10 ng./ml. range a sensitivity of 63.7% to 92.5% with a specificity of 57.5% to 18.7% was found when percent free PSA was 18% to 25%. On average 3 biopsies were needed to detect 1 cancer. When PSA was 2 to 4 ng./ml. sensitivity was 46.3% to 75.6% and specificity was 73.6% to 37.6% when the same percent free PSA cutoff was examined. Calculation of the biopsy-per cancer ratio for various percent free PSA cutoffs revealed that 3 to 5 biopsies were needed to find 1 cancer. Of 41 cancers detected in the PSA 2 to 4 ng./ml. range 6 had a Gleason score 5. The majority (28 of 41) of cases had a Gleason score of 6. Gleason score was 7 in 5 patients and 8 in 1. CONCLUSIONS: In the PSA 4 to 10 ng./ml. range high sensitivity for prostate cancer detection is critical and 3 biopsies are needed to detect 1 cancer. In the PSA 2 to 4 ng./ml. range a percent free PSA cutoff of 18% to 20% detected about 50% of cancers while sparing up to 73% of unnecessary biopsies with a biopsy-to-cancer ratio of 3 to 4:1. Percent free PSA can be applied to the PSA 2 to 4 ng./ml. range to detect prostate cancer and only moderately increases the number of biopsies needed to detect 1 significant cancer compared to the greater than 4 to 10 ng./ml. range. PMID- 12131299 TI - Percent Gleason grade 4/5 as prognostic factor in prostate cancer diagnosed at transurethral resection. AB - PURPOSE: We investigated the value of percent Gleason grade 4/5 as a predictor of long-term outcome in men with prostate cancer diagnosed at transurethral resection who received deferred treatment. MATERIALS AND METHODS: A series of 305 men with prostate cancer diagnosed at transurethral resection from 1975 to 1990 who had subsequent expectancy was analyzed. Mean patient age at diagnosis was 74 years (range 52 to 95). Slides were reviewed, and the Gleason score, percent Gleason grade 4/5 and modified Gleason score (the sum of the dominant and worst grades) were assessed. RESULTS: At followup 271 men (89%) had died, including 110 (36%) of prostate cancer. Gleason score, percent Gleason grade 4/5 and modified Gleason score were significant predictors of disease specific survival (p <0.001). Of all men 34% had tumors without any grade 4/5 pattern, of whom only 8% died of prostate cancer compared with 52% with any grade 4/5 pattern (p <0.001). Gleason score 6 tumors with focal grade 4 (less than 5%) had a worse prognosis than pure Gleason score 3 + 3 = 6 tumors (p = 0.008). There was nonsignificantly shorter survival for Gleason score 4 + 3 = 7 than for Gleason score 3 + 4 = 7 disease (p = 0.19). In Cox models including all possible pairs of Gleason score, percent Gleason grade 4/5 and modified Gleason score the percent Gleason grade 4/5 and modified Gleason score were stronger than Gleason score, although all 3 were independently significant prognosticators. CONCLUSIONS: Percent Gleason grade 4/5, modified Gleason score and Gleason score are predictors of disease specific survival in patients with prostate cancer on deferred treatment. Our study indicates that any grade 4/5 pattern impairs the prognosis significantly. PMID- 12131300 TI - Is a limited lymph node dissection an adequate staging procedure for prostate cancer? AB - PURPOSE: Generally lymph node dissection is only considered a staging procedure for prostate cancer. Therefore, the need for meticulous lymph node dissection is often questioned and only sampling is suggested. We performed a prospective study to identify the pattern of lymph node metastasis in prostate cancer and determine how extensive lymph node dissection must be not to under stage cases. MATERIALS AND METHODS: All patients with clinically organ confined prostate cancer, no prior hormonal treatment, negative preoperative staging computerized tomography and bone scan, who underwent radical prostatectomy between 1989 and 1999, were evaluated prospectively as to the number and location of lymph node metastasis. A meticulous lymph node dissection was performed along the external iliac vein, obturator nerve and internal iliac (hypogastric) vessels. Nodes from each location and side were submitted separately for histological evaluation. RESULTS: In 365 patients with a median serum prostate specific antigen of 11.9 ng./ml. (range 0.4 to 172) the median number of nodes removed was 21 (range 6 to 50). Lymph nodes were positive in 88 (24%) patients and the median number of positive nodes was 2 (range 1 to 19). Internal iliac lymph nodes were positive in 51 (58%) of the 88 patients, including 34 with additional positive lymph nodes along the external iliac vein and/or obturator nerve. Internal iliac lymph nodes alone were positive in 17 (19%) of 88 patients. CONCLUSIONS: There were significant numbers of lymph node metastases at all 3 different areas of lymphadenectomy. Positive lymph nodes were found along the internal iliac artery in more than half (58%) of the patients and exclusively in 19%. Therefore, we consider lymph node dissection along the internal iliac (hypogastric) vessels essential for representative staging. Without this dissection a fifth of node positive cases would have been under staged and diseased nodes would have remained in more than half of the cases. PMID- 12131301 TI - Pathological parameters of radical prostatectomy for clinical stages T1c versus T2 prostate adenocarcinoma: decreased pathological stage and increased detection of transition zone tumors. AB - PURPOSE: Studies of radical prostatectomy specimens have suggested that the majority of prostate specific antigen detected (clinical stage T1c) tumors are clinically significant. We compared tumor location and pathological parameters in the radical prostatectomy specimens of stages T1c versus T2 cases in a 3-year period. The percent of stage T1c disease represented a stable majority of patients undergoing treatment for clinically localized prostate cancer. MATERIALS AND METHODS: From January 1, 1998 to December 31, 2000, 417 radical prostatectomies were performed at Vanderbilt University, including 246 for stage T1c and 108 for stage T2 disease. A total of 37 patients were excluded from study because of neoadjuvant antiandrogen treatment. Pathological parameters, including tumor location in the transition and/or peripheral zone, tumor Gleason grade, tumor stage, total tumor volume and surgical margins were compared in stages T1c and T2 cases, and in transition versus peripheral zone stage T1c tumors in completely embedded whole mount specimens. RESULTS: In contrast to stage T2 lesions, stage T1c tumors were of significantly lower Gleason score with a higher percent of Gleason score 5 and lower percent of Gleason score 6, 7 and 8 or greater. They also had a significantly smaller volume and lower pathological stage. Of stage T1c tumors 77% were organ confined versus 62% of stage T2 tumors. There was no statistically significant increase in clinically insignificant neoplasms in stages T1c versus T2 cases (13% versus 7%) when using a volume criterion of less than 0.2 cc but a statistically significant increase in clinically insignificant disease was observed using a volume criterion of less than 0.5 cc (22% versus 9%). Whereas none of the T2 tumors were located in the transition zone and 17% were located in the transition and peripheral zones, 14% of stage T1c lesions were exclusively in the transition zone, with another 17% in the transition and peripheral zones. Compared with peripheral zone tumors transition zone stage T1c tumors had a lower Gleason score with an increase in Gleason score 5 and lower percent of Gleason score 6, 7 and 8 or greater. Although transition zone stage T1c lesions were significantly larger than peripheral zone stage T1c lesions, they had a lower pathological stage with 94% versus 72% organ confined. CONCLUSIONS: Prostate specific antigen detected stage T1c tumors had a lower grade, stage and volume than stage T2 tumors during the same period. Lower tumor grade in stage T1c cases is due at least in part to the increased detection of Gleason pattern 2 containing transition zone tumors. Despite the larger size, T1c transition zone tumors appear to be more favorable with higher rates of organ confined and lower grade tumors. If such transition zone tumors prove to be biologically distinct, improved strategies to identify these lesions preoperatively may result in more conservative treatment recommendations. PMID- 12131302 TI - Competing risk analysis after radical prostatectomy for clinically nonmetastatic prostate adenocarcinoma according to clinical Gleason score and patient age. AB - PURPOSE: Factors important for determining appropriate therapy for localized prostate cancer are biopsy tumor grade, patient age and co-morbidity. We estimated the probability of dying from prostate cancer or other competing causes stratified by age at diagnosis and clinical histological grade in men diagnosed with clinically nonmetastatic prostate cancer who were treated with radical prostatectomy. MATERIALS AND METHODS: A total of 751 men comprised a retrospective cohort with clinically nonmetastatic prostate cancer diagnosed and treated with bilateral pelvic lymphadenectomy and radical prostatectomy at our institution between 1971 and 1984. All patients were between 55 and 74 years old (median age 65) at diagnosis and they were followed a median of 14.7 years. The cumulative incidence of prostate cancer death or death from any cause was estimated using methods of competing risk survival analysis. RESULTS: Overall 435 men died with 32% of the deaths attributable to prostate cancer. In 62%, 27% and 11% of patients the Charlson co-morbidity score was 0, 1 and 2+, respectively. The only significant predictor of death from prostate cancer was clinical Gleason score (p <0.001), while only age and Charlson co-morbidity score were significant independent predictors of death from other causes (p <0.001). The estimated cumulative incidence of prostate cancer death after considering competing risks increased with Gleason score regardless of patient age. In men with Gleason scores 2 to 4, 5, 6, 7 and 8 to 10 disease the cumulative incidence of prostate cancer death within 20 years was 6% to 7%, 10% to 13%, 15% to 19%, 29% to 35% and 36% to 43%, respectively, depending on age at diagnosis. Clinical stages T2 and T3 outcomes were indistinguishable. CONCLUSIONS: This study shows that for any given Gleason score the prostate cancer death rate is similar in older and younger patients with few to no co-morbidities. Men with a score of 7 to 10 were at 29% to 43% risk of death from prostate cancer even when cancer was diagnosed as late as age 74 years and treated surgically. PMID- 12131303 TI - Predictors of secondary cancer treatment in patients receiving local therapy for prostate cancer: data from cancer of the prostate strategic urologic research endeavor. AB - PURPOSE: Secondary cancer treatment is common after definitive local therapy for prostate cancer and it may be an indicator of the efficacy and cost of primary local treatment. We determined predictors of secondary cancer treatment in patients initially treated with radical prostatectomy or external beam radiation. MATERIALS AND METHODS: We examined 2,336 patients in Cancer of the Prostate Strategic Urologic Research Endeavor, a longitudinal registry of patients with prostate cancer, who underwent initial treatment with radical prostatectomy (1,744) or external beam radiation (592). Patients had at least 1 month of followup and all pretreatment information was available. The percent of patients receiving secondary cancer treatment, time to secondary treatment and type of secondary treatment delivered was determined. Multivariate analysis was done to determine independent predictors of secondary cancer treatment. In patients initially treated with prostatectomy a similar analysis was performed to identify predictors of receiving androgen deprivation versus radiation. RESULTS: A total of 590 patients (25%) received secondary cancer treatment, including prostatectomy in 391 (22%) and radiation in 199 (34%). Secondary cancer treatment was equally divided between radiation and androgen deprivation in 52% and 47%, respectively, of those initially treated with prostatectomy, while 92% initially treated with radiation received androgen deprivation. Predictors of any secondary treatment included patient age, biopsy Gleason score and prostate specific antigen at diagnosis. There was a trend toward increased secondary treatment more than 6 months after local therapy in patients initially treated with radiation. Increased age and lymph node metastases were independent predictors of receiving androgen deprivation after prostatectomy, while there was increased use of radiation in patients with positive surgical margins or extracapsular disease extension. CONCLUSIONS: Secondary treatment differs in patients initially treated with radical prostatectomy and radiation. Pretreatment factors can be used to counsel patients regarding the likelihood of secondary treatment, while age and prostatectomy results appear to determine the type of secondary treatment in those initially treated with prostatectomy. PMID- 12131304 TI - Biochemical disease-free survival in men younger than 60 years with prostate cancer treated with external beam radiation. AB - PURPOSE: We determined the biochemical failure rate in patients 60 years and younger, and older than 60 years old who were treated with external beam radiation for localized prostate cancer or locally advanced prostate cancer. We also evaluated prognostic factors in the 2 age groups. MATERIALS AND METHODS: We reviewed the medical records of 964 patients who received full dose radiotherapy as the only treatment for prostate cancer. Followup prostate specific antigen was measured 3 to 6 months after the completion of radiotherapy and every 3 to 6 months thereafter. Biochemical failure was defined using the criteria established by the American Society for Therapeutic Radiology and Oncology Consensus Panel. Median followup in the whole study group was 48 months. RESULTS: Of the 98 men 60 years or younger 46 (47%) had biochemical failure, whereas 261 (30%) of the 866 older than 60 years old had biochemical failure. The 5 and 7-year biochemical disease-free survival rates were 55% and 47% in the younger men, and 65% and 59% in the older men, respectively. These rates were significantly lower in the younger men (p = 0.017 and 0.027, respectively). Multivariate regression showed that in men 60 years or younger initial prostate specific antigen, Gleason score and lower radiation doses were predictive of biochemical failure. CONCLUSIONS: Men with prostate cancer who are 60 years or younger and treated with radiotherapy may be at significant risk for long-term biochemical failure. PMID- 12131305 TI - Low dose ketoconazole with replacement doses of hydrocortisone in patients with progressive androgen independent prostate cancer. AB - PURPOSE: High-dose (400 mg.) oral ketoconazole 3 times daily with replacement doses of hydrocortisone has become a standard treatment option for patients with advanced prostate cancer which progresses after androgen deprivation. However, toxicity can hinder the ability to deliver treatment and the cost of the regimen can be substantial. Therefore, a prospective phase II study was conducted to assess the efficacy and safety of a regimen of low dose (200 mg.) oral ketoconazole 3 times daily with replacement doses of hydrocortisone in men with androgen independent prostate cancer. MATERIALS AND METHODS: The study included 28 patients with progressive prostate cancer despite anorchid levels of testosterone and ongoing testicular androgen suppression. Treatment consisted of low dose ketoconazole and replacement doses of oral hydrocortisone (20 mg. every morning and 10 mg. at bedtime). At the time of disease progression patients were treated with high dose ketoconazole and continued on the same dose of hydrocortisone. Adrenal androgen levels were measured, and baseline and followup levels correlated with clinical outcome. RESULTS: Overall, 13 (46%) of 28 patients had a prostate specific antigen decrease of more than 50% (95% confidence interval 27.5% to 66.1%). Median duration of prostate specific antigen decrease for all responders was 30+ weeks and 5 patients continue to exhibit a response, ranging from 36+ to 53+ weeks. In general, therapy was well tolerated. There were no grade 4 toxicities. Grade 3 toxicities included hepatotoxicity in 1 patient and depression in 2. The most common toxicities were nausea (29% grades 1 and 2), dry skin (18% grade 1) and fatigue (14% grade 1). Four (14%) patients discontinued low dose ketoconazole due to toxicities. Of the 16 patients who received high dose ketoconazole after disease progression with low dose ketoconazole, 3 were removed from treatment due to toxicity and no patient responded to high dose ketoconazole. There was no difference in the distribution of pretreatment endocrine values between responders and nonresponders, and the magnitude of change in adrenal androgen levels was not associated with response to therapy, although a potential association could easily have been missed due to small sample size. CONCLUSIONS: The regimen of low dose ketoconazole with replacement doses of hydrocortisone is well tolerated and has moderate activity in patients with progressive androgen independent prostate cancer. PMID- 12131306 TI - Hormone therapy for locally advanced prostate cancer. AB - PURPOSE: We assessed cause specific and all cause survival in men with locally advanced prostate cancer after hormone therapy. MATERIALS AND METHODS: Between February 1991 and November 2000, 208 men with locally advanced prostate cancer were treated with gonadal androgen ablation or gonadal androgen ablation and an antiandrogen at a single medical center. Median PSA was 46 ng./ml. (range 2 to 748). Median potential followup was 78 months (range 4 to 122) and the median observation period was 46 months (range 3 to 122). RESULTS: Of the patients 14 (7%) died of causes related to cancer and 71 (34%) died of competing co-morbid disease. Actuarial cause specific survival at 5 and 8 years was 92% and 80%, respectively. The only demographic or tumor related variable that influenced cause specific survival was Gleason score less than 8 versus 8 or greater (p = 0.02). Actuarial all cause survival at 5 and 8 years was 59% and 41%, respectively. The only variable that influenced all cause survival was a Charlson weighted co-morbidity score of less than 2 versus 2 or greater (p <0.0001). Major morbidity from the primary tumor, including bothersome obstructive voiding symptoms requiring transurethral prostate resection, ureteral obstruction or persistent hematuria, developed in 13 patients (6%), while major treatment related morbidity, including flutamide hepatotoxicity and hip fracture, developed in 4. CONCLUSIONS: Hormone therapy for locally advanced prostate cancer is associated with minimal morbidity from the primary tumor and from treatment. All cause survival parallels that reported for integrated hormone and radiation therapy. PMID- 12131307 TI - Steroid therapy for idiopathic retroperitoneal fibrosis: dose and duration. AB - PURPOSE: Idiopathic retroperitoneal fibrosis is an uncommon disease of unknown etiology that may involve the ureters and other retroperitoneal structures. Surgical ureterolysis as well as medical treatment with steroids have been used to treat these patients. However, there is no agreement as to the dose and duration of steroid. We adopted a regimen of long-term steroid use in this prospective study. MATERIALS AND METHODS: We treated 12 patients with idiopathic retroperitoneal fibrosis with a regimen of steroids during a 10-year period. Tissue diagnosis was established by biopsy of all lesions and ureteral obstruction was managed with insertion of a nephrostomy tube or a ureteral stent. The initial dose of prednisolone was 60 mg. on alternate days for 2 months and was tapered during the following 2 months to a daily dose of 5 mg. The total duration of prednisolone use was 2 years. RESULTS: Of the 12 patients 11 who completed this treatment regimen have been followed for a duration of 26 to 132 months (median 63.1) after discontinuation of treatment. Good response in the form of relief of symptoms and regression of the mass occurred in 9 cases and there were 2 failures. In 1 case the retroperitoneal mass did not regress and surgical ureterolysis was required. In the 2nd case symptoms recurred after discontinuation of steroid and a further small dose of steroids was required. Function deteriorated in 1 of 19 functioning renal units. No steroid related serious side effects developed. CONCLUSIONS: This regimen of steroid may be used as the primary mode of treatment for the majority of patients with idiopathic retroperitoneal fibrosis with minimal complications. Patients with idiopathic retroperitoneal fibrosis should be followed periodically for the rest of their lives. PMID- 12131308 TI - Does the potassium stimulation test predict cystometric, cystoscopic outcome in interstitial cystitis? AB - PURPOSE: We establish the relationship among symptom duration, cystometric and cystoscopic findings and potassium stimulation test in patients with interstitial cystitis. MATERIALS AND METHODS: A retrospective chart review was performed of 189 patients treated at an ambulatory clinic between 1992 and 1998. Urodynamic parameters, potassium stimulation test results and subjective response to treatment were evaluated. Fisher's exact test was used for statistical analysis. RESULTS: Of the 189 patients diagnosed with interstitial cystitis 173 (92%) were female and 16 (8%) were male. The potassium stimulation test was positive in 105 (83%) patients, negative in 16 (13%) and equivocal in 6 (4%). A cystometrogram and potassium stimulation test were done in 118 patients. Bladder capacity averaged 259 ml. in patients with tests potassium positive and negative, while average bladder volume at first sensation to void was 85 ml. and 148 ml. in those with negative and positive tests, respectively. Among the 102 patients with a positive potassium stimulation test 52 had normal cystoscopic findings. CONCLUSIONS: The potassium stimulation test is not correlated with either bladder capacity or cystoscopic findings. PMID- 12131309 TI - Is periprostatic local anesthesia for transrectal ultrasound guided prostate biopsy associated with increased infectious or hemorrhagic complications? A prospective randomized trial. AB - PURPOSE: Periprostatic local anesthesia for prostate biopsy requires 2 or more extra needle punctures and injection of the local anesthetic through the highly colonized rectum. To our knowledge we report the first prospective randomized trial to assess the infectious or hemorrhagic complications associated with this method. MATERIALS AND METHODS: A total of 100 consecutive patients with sterile urine cultures underwent transrectal ultrasound guided prostate biopsy. They were randomized to receive a periprostatic nerve block or no anesthesia. Patients were evaluated for the amount of rectal and urethral bleeding, and symptoms and signs of infection after biopsy. RESULTS: The amount of urethral bleeding was slight and similar in the 2 groups. Rectal bleeding was significantly less in the patients who received anesthesia. High fever (greater than 37.8C) was more frequent in the nerve block group and 2 patients in this group required rehospitalization. Bacteriuria in post-biopsy urine cultures was significantly more common in the anesthesia group. CONCLUSIONS: Our results suggest that periprostatic local anesthesia for prostate biopsy does not increase the risk of urethral bleeding. It is associated with a decreased incidence of rectal bleeding, presumably due to decreased patient discomfort. The incidence of bacteriuria was significantly higher in the anesthesia group. High fever and hospitalization due to infectious complications were also more common in the local anesthesia group, although not statistically significant. Prospective randomized trials seem warranted to determine the optimum antibiotic prophylaxis regimen in patients undergoing biopsy with a periprostatic nerve block. PMID- 12131310 TI - Variability in catheter microwave sterilization techniques in a single clinic population. AB - PURPOSE: We designed a survey to assess the microwave sterilization technique practiced by patients at our clinic who perform clean intermittent catheterization. MATERIALS AND METHODS: A 23-question survey addressing urinary catheter use and home sterilization techniques was mailed to 129 patients. A followup survey was mailed to 47 respondents who reported using a microwave oven to sterilize the catheters to assess the microwave technique further. RESULTS: Of the 129 initial surveys 84 (64%) were returned, while 40 (85%) of the 47 followup questionnaires on microwave sterilization were returned. All patients surveyed have used clean intermittent catheterization for at least 1 year and 75% have used it more than 5 years. Of the respondents 80% perform clean intermittent catheterization 4 to 5 times daily, although sterilization frequency varies from daily to less than once weekly. Of the respondents 71% reported no difficulty with microwave sterilization, although 31 (63%) reported a history of catheter melting during microwaving. Of the respondents 35% reported using a rotation table, all used a heat sink containing 1/2 to 4 cups of water, 39% used 500 to 1,000 W., 37% used greater than 1,000 W., 73% set the microwave for 6 minutes, others set it for 3 to 30 minutes and 98% used a power setting described as high, full, 10 or 100%. CONCLUSIONS: Significant variation exists in the cleaning and sterilizing techniques used by our patients, although they were given uniform written and verbal instructions. It is unclear from the data in the literature how this variation affects sterilization. PMID- 12131311 TI - Relative bladder outlet obstruction. AB - PURPOSE: Currently bladder outlet obstruction in males is defined by the provisional International Continence Society nomogram which is partly based on expert opinion and partly on measurements before and after transurethral prostate resection. Recently there has been some interest in the development of a similar nomogram for females. MATERIALS AND METHODS: We studied the possibility of defining bladder outlet obstruction based on a sign that it causes, namely post void residual urine. RESULTS: The probability of relative post-void residual urine exceeding 20% of bladder capacity was modeled in males and females using 1 parameter, that is URA/w20 or the ratio of the obstruction parameter urethral resistance factor (URA)-to-the bladder contractility parameter Watts factor at 20% (w20). URA/w20 represents relative bladder outlet resistance or bladder outlet resistance normalized to bladder contractility. Above a threshold of URA/w20 = 6.8 in females and 8.2 in males a relative post-void residual exceeding 20% was noted in 90% of measurements. These thresholds may be used to define relative obstruction. The provisional International Continence Society nomogram for obstruction in males was transformed into an identical nomogram for females by equating the probabilities of post-void residual urine in each gender. The latter differed from that in men, in that the lines demarcating the zones were horizontal or flow rate independent but the intercepts were approximately the same at 20 and 40 cm. water. CONCLUSIONS: Instead of defining obstruction as an absolute level of bladder outlet resistance we suggest that it is better to define it relatively, that is as a level of bladder outlet resistance that depends on bladder contractility. PMID- 12131312 TI - alpha1-Adrenergic blockers in young men with primary bladder neck obstruction. AB - PURPOSE: We assessed the efficacy and safety of the mid term use of alpha1 adrenergic blockers for treating primary bladder neck obstruction in young and middle-aged men. MATERIALS AND METHODS: Between May 1998 and February 2001 primary bladder neck obstruction was diagnosed by videourodynamic study in 28 men younger than 55 years. The degree of obstruction was graded by the Schafer nomogram. Mean patient age and mean symptom duration were 39.3 years and 18.1 months, respectively. The presenting symptom was frequency in 22 cases (78.6%), urgency in 10 (35.7%), weak stream in 9 (32.1%), nocturia greater than 2 times in 7 (25%) and hesitancy in 7 (25%). A dose of 1 to 2 mg. doxazosin was administered at bedtime for at least 3 months. International Prostate Symptom Score (I-PSS), quality of life, uroflowmetry and post-void residual urine were assessed before and 3 months after medication. Improved urine flow was defined as at least a 2.5 ml. per second increase in the maximum flow rate. Improved symptoms was defined as more than a 50% decrease in I-PSS. Successful treatment was defined as improved urine flow and symptoms. RESULTS: Followup data were available in 24 patients. The medication period and followup were 7 and 12 months, respectively. Mean I-PSS plus or minus standard deviation decreased from 18.3 +/- 4.6 to 11.6 +/- 5.2 (p <0.01) and mean quality of life decreased from 4.1 +/- 1.1 to 2.6 +/- 1.0 (p <0.01). Mean maximum flow increased from 11.8 +/- 3.2 to 15.9 +/- 3.9 ml. per second (p <0.01). Mean post-void residual urine decreased from 80.2 +/- 17.1 to 48.5 +/- 10.3 ml. (p <0.01). Treatment was successful in 13 patients (54.2%) and 3 (12.5%) were free of medication for at least 6 months. More successful outcomes were noted in patients with high grades III to IV obstruction than in those with low grades 0 to II obstruction (80% versus 35.7%, p = 0.03). Before treatment higher detrusor pressure at maximum flow (70.1 versus 47.8 cm. water, p = 0.01) and lower maximum flow (10.6 versus 13.3 ml. per second, p = 0.02) were observed in patients with successful versus unsuccessful treatment, respectively. Differences in patient age, I-PSS, quality of life, prostate size and post-void residual urine were not statistically significant. No significant adverse effects were noted. CONCLUSIONS: alpha1-Blockers were effective and safe for treating young men with primary bladder neck obstruction. PMID- 12131313 TI - The effect of intravesical resiniferatoxin in patients with idiopathic detrusor instability suggests that involuntary detrusor contractions are triggered by C fiber input. AB - PURPOSE: We evaluated the role of bladder C-fiber input in involuntary detrusor activity in patients with idiopathic detrusor instability. MATERIALS AND METHODS: Filling cystometry and a voiding chart were done in 13 patients with idiopathic detrusor instability. The first detrusor contraction, maximal cystometric capacity, daily frequency and the number of episodes of urinary incontinence were determined. A 50 nM. solution of resiniferatoxin, a specific C-fiber neurotoxin, was then instilled in the bladder for 30 minutes. Patients were reevaluated 30 and 90 days later. RESULTS: Resiniferatoxin instillation delayed or suppressed involuntary detrusor contractions during filling cystometry. The mean first detrusor contraction plus or minus standard deviation increased from 170 +/- 109 ml. at baseline to 440 +/- 130 ml. (p = 0.0001) at 30 days and to 391 +/- 165 ml. (p = 0.008) at 90 days. Mean maximal cystometric capacity increased from 291 +/- 160 to 472 +/- 139 ml. (p = 0.01) at 30 days and to 413 +/- 153 ml. (p = 0.1) at 90 days. The mean number of episodes of urinary incontinence daily decreased from 4.3 +/- 2.7 to 0.9 +/- 2.7 (p = 0.001) at 30 days and to 0.7 +/- 0.9 (p = 0.009) at 90 days. Mean frequency daily also decreased from 12 +/- 3.2 to 9.7 +/- 3.2 (p = 0.003) and to 9.9 +/- 3.5 (p = 0.001) times at the same time points, respectively. CONCLUSIONS: C-fiber input seems to have an important role in the generation of involuntary detrusor contractions and lower urinary tract symptoms in patients with idiopathic detrusor instability. Substances that block C-fiber input may represent a new strategy for treating this bladder dysfunction. PMID- 12131314 TI - Efficacy and safety of transdermal oxybutynin in patients with urge and mixed urinary incontinence. AB - PURPOSE: We evaluated the efficacy and safety of an oxybutynin transdermal delivery system (TDS) in a general population of patients with overactive bladder and urge or mixed urinary incontinence. MATERIALS AND METHODS: Following symptom stabilization or treatment withdrawal 520 adult patients were randomized to 12 weeks of double-blind daily treatment with 1.3, 2.6 or 3.9 mg. oxybutynin TDS or placebo administered twice weekly, followed by a 12-week open-label, dose titration period to assess efficacy and safety further. Evaluations included patient urinary diaries, incontinence specific quality of life and safety. RESULTS: A dose of 3.9 mg. daily oxybutynin TDS significantly reduced the number of weekly incontinence episodes (median change -19.0 versus -14.5, p = 0.0165), reduced average daily urinary frequency (mean change -2.3 versus -1.7, p = 0.0457), increased average voided volume (median change 24 versus 6 ml., p = 0.0063) and significantly improved quality of life (Incontinence Impact Questionnaire total score, p = 0.0327) compared with placebo. Average voided volume increased in the daily 2.6 mg. group (19 ml., p = 0.0157) but there were no other significant differences between 1.3 and 2.6 mg. oxybutynin TDS and placebo. The most common adverse event was application site pruritus (oxybutynin TDS 10.8% to 16.8%, placebo 6.1%). Dry mouth incidence was similar in both groups (7.0% versus 8.3%, p not significant). In the open-label period a sustained reduction of nearly 3 incontinence episodes per day was reported for all groups. CONCLUSIONS: Doses of 2.6 and 3.9 mg. oxybutynin TDS daily improve overactive bladder symptoms and quality of life, and are well tolerated. Transdermal oxybutynin is an innovative new treatment for overactive bladder. PMID- 12131315 TI - Critical evaluation of the problem of chronic urinary retention after orthotopic bladder substitution in women. AB - PURPOSE: We studied the possible causes of chronic retention after radical cystectomy and orthotopic bladder substitution in women. MATERIALS AND METHODS: Between January 1995 and January 2001, 136 women with a mean age plus or minus standard deviation of 52 +/- 8 years underwent standard radical cystectomy and orthotopic substitution for organ confined bladder cancer. Videourodynamics, pelvic floor electromyography, pelvic floor magnetic resonance imaging and pan endoscopy were done. In the last 37 cases some technical modifications were adopted to circumvent the development of chronic urinary retention. RESULTS: One woman died postoperatively of massive pulmonary embolism. Of the 100 patients evaluable at a mean followup of 36 months 95 were continent in the daytime, 86 were continent at night, 2 were completely incontinent and 16 were in chronic retention. Videourodynamics showed that retention was mechanical in nature due to the pouch falling back in the wide pelvic cavity, resulting in acute angulation of the posterior pouch-urethral junction. In addition, herniation of the pouch wall through the prolapsed vaginal stump was observed in most cases. Pelvic floor electromyography demonstrated complete pelvic floor silence during voiding. No abnormality of the pelvic floor or rhabdosphincter was noted on magnetic resonance imaging. Pan-endoscopy showed a normal urethra with no urethroileal stricture. A 4 mg. dose of the alpha1-adrenergic blocker doxazosin daily was ineffective, excluding the possibility that sprouting from adjacent adrenergic neurons into the denervated proximal urethral muscles may have been the cause of this problem. After omental packing behind the pouch, suturing of the peritoneum on the rectal wall to the vaginal stump, suspension of the latter by the preserved round ligaments and suspension of the pouch near its dome to the back of the rectus muscle at cystectomy the incidence of chronic retention decreased from 18.7% (14 of 75 cases) before to 8% (2 of 25) after modifications. Furthermore, after vaginal wall descent was mechanically corrected by a pessary there was significant improvement in evacuation. CONCLUSIONS: Strong evidence was provided that chronic urinary retention after orthotopic substitution is due to anatomical rather than to functional or neurogenic reasons. Modifications to increase back support of the pouch with ventral suspension near its dome and support the vaginal stump are recommended to avoid this complication. PMID- 12131317 TI - Body size and serum levels of insulin and leptin in relation to the risk of benign prostatic hyperplasia. AB - PURPOSE: Obesity has been implicated in the etiology of benign and malignant prostatic growth due to its influence on metabolic and endocrine changes. Because obesity is an important determinant of serum levels of insulin and leptin (the product of the obesity gene Ob), we investigated the role of obesity and serum levels of insulin and leptin in benign prostatic hyperplasia (BPH) etiology. MATERIALS AND METHODS: Fasting serum levels of insulin and leptin as well as the body mass index, a measure of overall obesity, and waist-to-hip ratio, an indicator of abdominal obesity, were determined in 200 men newly diagnosed with BPH who were hospitalized for surgery and in 302 randomly selected healthy male subjects from the population in Shanghai, China. RESULTS: A higher waist-to-hip ratio and higher serum insulin were significantly associated with an increased risk of BPH. Relative to men in the lowest waist-to-hip ratio quartile (less than 0.856) those in the highest quartile (greater than 0.923) were at 2.4-fold risk (odds ratio 2.42, 95% confidence interval [CI] 1.34 to 4.37, test for trend p = 0.01). Similarly relative to men in the lowest quartile of insulin (less than 5.87 microU. per ml.) those in the highest quartile (greater than 9.76 microU. per ml.) were at significantly increased risk (odds ratio 2.47, 95% CI 1.35 to 4.54, test for trend p = 0.009). The effect of insulin on BPH risk was more pronounced in men in low and middle tertiles of the waist-to-hip ratio (odds ratios comparing high to low insulin tertiles 2.8 and 2.7, respectively), while among men in the highest waist-to-hip ratio tertile insulin was not significantly associated with BPH risk. In contrast, we found no significant odds ratio comparing the highest to lowest quartiles of leptin (odds ratio 0.62, 95% CI 0.33 to 1.17) or body mass index (odds ratio 1.64, 95% CI 0.96 to 2.81). CONCLUSIONS: Our results suggest that abdominal obesity and increasing serum insulin, and possibly overall obesity but not serum leptin are associated with a higher risk of BPH. Further prospective and laboratory studies are needed to confirm these results and elucidate the underlying mechanisms. PMID- 12131316 TI - Demographic and clinical characteristics of men with chronic prostatitis: the national institutes of health chronic prostatitis cohort study. AB - PURPOSE: We describe the study design of the National Institutes of Health Chronic Prostatitis Cohort (CPC) study characterizing men with chronic prostatitis/the chronic pelvic pain syndrome. MATERIALS AND METHODS: All 488 men screened into the CPC study before close of recruitment on August 22, 2001 were selected for analysis. The National Institutes of Health Chronic Prostatitis Symptom Index, including subscores, was used to measure symptoms. A comprehensive history, physical examination and demographic profile were obtained from each participant. Generalized Mantel-Haenszel procedures were used to investigate baseline associations between selected factors and symptoms. RESULTS: Chronic prostatitis/chronic pelvic pain syndrome is a chronic syndrome affecting men over a wide age range. The majority of CPC study participants are white, well educated and affluent. However, lower education, lower income and unemployment were associated with more severe symptoms. Patients most frequently reported pain in the perineum and tenderness in the prostate. The highest self-reported diseases were genitourinary (55%), allergies (53%), neurological (40%) and hematopoietic, lymphatic or infectious (40%). This disease has a significant negative impact on mental and physical domains of quality of life. Almost all patients (95%) reported antimicrobial drug use. Of these 488 participants 280 (57%) reported the previous or current use of 5 or more categories of prostatitis related treatments. CONCLUSIONS: Chronic prostatitis/chronic pelvic pain syndrome is a multifactorial problem affecting men of all ages and demographics. Patients with the chronic pelvic pain syndrome have dismal quality of life and many have benefited only minimally from empirical, goal directed therapy. Long-term followup of this cohort may answer important questions on the natural treated history of this syndrome. PMID- 12131318 TI - Correlations of urodynamic changes with changes in symptoms and well-being after transurethral resection of the prostate. AB - PURPOSE: To establish the predictive value of urodynamics on the outcome of transurethral prostate resection for benign prostatic enlargement we correlated urodynamic changes with symptomatic improvement, decreased bother, and increased general well-being and quality of life after transurethral prostate resection. MATERIALS AND METHODS: Men with lower urinary tract symptoms were selected if they met study criteria and underwent tests recommended by the International Scientific Committee on Benign Prostatic Hyperplasia, and if post-void residual urine volume and prostate size were estimated. Patients answered quality of life, symptom index, symptom problem index and benign prostatic hyperplasia impact index questions. Patients also underwent urodynamic evaluation. Men were included in analysis when transurethral prostate resection was selected as the treatment modality. Of the 132 patients included 93 were reevaluated 6 months after transurethral prostate resection. RESULTS: Improvements after transurethral prostate resection were significantly associated with decreased bladder outlet obstruction (p <0.01). However, 32 cases that were unobstructed or equivocal preoperatively also benefited moderately from resection. Effective capacity, that is cystometric capacity minus post-void residual urine volume, increased by an average of 45% postoperatively. The increase in effective capacity contributed to a significant decrease in symptoms and bother, and to improved well-being. Of the men with a urodynamically proved stable bladder 90% maintained a stable bladder after prostatectomy, while in 50% with a urodynamically proved unstable bladder it became stable postoperatively. CONCLUSIONS: Performing urodynamics preoperatively helps to predict the degree of symptom relief, decreased bother and increased well-being after transurethral prostate resection. PMID- 12131319 TI - The efficacy and safety of perioperative low molecular weight heparin substitution in patients on chronic oral anticoagulant therapy undergoing transurethral prostatectomy for bladder outlet obstruction. AB - PURPOSE: Candidates for prostatectomy who require chronic anticoagulant therapy present a major perioperative management problem due to the threat of significant hemorrhage associated with surgery and the risk of thromboembolism associated with discontinuation of the anticoagulants. We evaluated prospectively a perioperative routine using low molecular weight heparin substitution to allow safe discontinuation of prophylactic oral anticoagulants in patients undergoing transurethral resection of the prostate. MATERIALS AND METHODS: We treated 20 patients on chronic oral anticoagulant therapy who required prostatectomy for bladder outlet obstruction according to a prospective protocol based on exchange of the oral anticoagulants with perioperative injections of low molecular weight heparin and resumption of oral anticoagulants early postoperatively. The safety and efficacy of this regimen were assessed in comparison to a control group comprised of 20 randomly selected nonwarfarin treated patients who underwent prostatectomy during the same period. RESULTS: The need for blood transfusions and mean number of units transfused did not significantly differ between the 2 groups. Due to persistent hematuria routine removal of the catheter was possible only in 9 of 20 patients (45%) in the heparin group compared to 18 of 20 (90%) in the control group. Average catheterization and hospitalization period in the heparin group was 3.2 days and 4.2 days respectively, compared to 2.1 and 2.1 days in the control group, respectively (p <0.01). No long-term hemorrhagic or thromboembolic complications were noted at 3 months postoperatively. CONCLUSIONS: Despite longer hospitalization in the heparin treated group, our substitution protocol is safe and effective. PMID- 12131320 TI - Diagnostic steps in the evaluation of patients with erectile dysfunction. AB - PURPOSE: The necessity for a thorough diagnostic evaluation for erectile dysfunction has been questioned after the availability of effective oral therapies. We determined the impact of the different diagnostic steps on the management strategy for erectile dysfunction. MATERIALS AND METHODS: The study included all patients who presented at an andrology outpatient clinic during a 4 year period. Baseline evaluation included medical and sexual history, blood tests, physical examination and intracavernous injection test. Patients with normal initial screening were evaluated with specific diagnostic procedures. The results were analyzed to identify the diagnostic potential of each screening step separately. RESULTS: Overall 1,644 patients presented at the clinic during the study period, of whom 368 (22.4%) were excluded from study due to severe psychiatric (5.2%) or cardiovascular (2.7%) disease, or to a history of erectile dysfunction less than 3 months in duration (14.5%). In the remaining 1,276 patients with a mean age plus or minus standard deviation of 56 +/- 14 years, and a mean duration of erectile dysfunction of 4.9 +/- 3.4 years medical history revealed erectile dysfunction associated medical conditions in 57%, blood tests identified previously undiagnosed medical conditions in 6.2%, and physical examination and the intracavernous injection test were diagnostic in 13.9% and 2.6%, respectively. Initial screening was negative in 259 cases (20.3%), in which specific diagnostic procedures identified an underlying vascular pathology in 165 (12.9%) and unfavorable penile geometry in 16 (1.3%). The remaining 78 men (6.1%) had no evidence of organic disease. CONCLUSIONS: Baseline diagnostic evaluation for erectile dysfunction can identify the underlying pathological condition or erectile dysfunction associated risk factors in 80% of patients. Such screening may diagnose reversible causes of erectile dysfunction and also unmask medical conditions that manifest with erectile dysfunction as the first symptom. Specific diagnostic procedures may be limited in patients with primary erectile dysfunction or those without risk factors. Such clinical data support previously published guidelines for erectile dysfunction management. PMID- 12131321 TI - Experience with intraplaque injection of verapamil for Peyronie's disease. AB - PURPOSE: We examined the use of intraplaque injection of verapamil for the treatment of Peyronie's disease through its effects on pain, curvature, indentation, sexual function and erectile capacity. MATERIALS AND METHODS: A total of 156 men underwent treatment with intraplaque verapamil injection. Patients were assessed objectively, during dynamic penile duplex ultrasound, as well as subjectively using a questionnaire before and after initiation of the treatment protocol. Patients were also stratified by duration of disease before therapy and into 1 of 3 Kelami classification groups based on pretreatment plaque size and severity of curvature. Differences before and after treatment and among the Kelami classification groups were assessed. RESULTS: Of the 140 patients who completed treatment 73 (60%) had an objectively measured decrease in curvature while 79 (62%) reported a subjective decrease in curvature during the followup interview. After treatment 111 (83%) men reported an increase in girth, 107 (80%) an increase in rigidity distal to the plaque and 92 (71%) an improvement in sexual function. Among each Kelami class curvature was objectively measured to decrease in 41%, 68% and 62% of patients in classes I, II and III, respectively. There was no significant difference in response based on duration of disease (60% improvement versus 61% improvement for disease duration of less or greater than 1 year in duration, respectively). Mean followup was 30.4 months (range 10 to 81) and there was no reported recurrence of penile deformity in those men with an initial posttreatment positive response. CONCLUSIONS: Verapamil injection of Peyronie's plaques appears to be a clinically effective treatment option for pain and curvature and can contribute to subjective improvement in sexual function and erectile capacity. The low incidence of complications indicates that this therapy is also clinically safe. PMID- 12131322 TI - Drain-free simple retropubic prostatectomy with fibrin sealant. AB - PURPOSE: We present our preliminary experience with liquid fibrin sealant during simple retropubic prostatectomy. MATERIALS AND METHODS: We reviewed 18 consecutive simple retropubic prostatectomies performed for symptomatic advanced benign prostatic hyperplasia at our institution between 1997 and 2001. Adenoma enucleation was performed via transverse anterior prostatic capsulotomy. In the first 13 cases (group 1) a Jackson-Pratt suction drain was placed in the pelvis after prostatic capsular closure. In the remaining 5 cases (group 2) 2 ml. liquid fibrin sealant were administered over the closed prostatic capsule instead of a pelvic drain. RESULTS: The 2 groups were matched for age and prostate size. Average time to drain removal in control group was 3.92 days, while the fibrin sealant group had no clinically apparent adverse sequelae despite the lack of pelvic drainage. Average hospitalization in group 1 was 4.38 days, while all group 2 patients were discharged home after 2 days (p = 0.001). In addition, a trend toward earlier resumption of a full diet was noted in the sealant group (2.61 versus 1 day, p = 0.075). CONCLUSIONS: Liquid fibrin sealant appears to expedite recovery and shorten hospitalization when used as an adjunct during simple prostatectomy. PMID- 12131323 TI - Lithiasis in varicocele veins: "varicolithiasis". PMID- 12131325 TI - Primary synovial sarcoma of the penis. PMID- 12131324 TI - Prurigo nodularis as initial presentation of metastatic transitional cell carcinoma of the bladder. PMID- 12131326 TI - Symptomatic renal metastasis of a testicular seminoma mimicking pelvicaliceal transitional cell carcinoma. PMID- 12131327 TI - Leiomyo-adenomatoid tumor of the epididymis. PMID- 12131328 TI - Dermatomyositis: a rare initial presentation of adenocarcinoma of the prostate. PMID- 12131329 TI - Rapidly progressing adenocarcinoma of the prostate presenting as prostatitis. PMID- 12131330 TI - Primary squamous cell carcinoma of the prostate: a novel chemotherapy regimen. PMID- 12131331 TI - Metastasis of an ascending colon carcinoma in the prostate 10 years after hemicolectomy. PMID- 12131332 TI - Primary rhabdomyosarcoma of the seminal vesicle. PMID- 12131333 TI - Splenosis presenting as a right suprarenal retroperitoneal mass. PMID- 12131334 TI - Can oxybutynin cause peripheral neuropathy? PMID- 12131335 TI - Unique management of a congenital polyp of the prostatic urethra. PMID- 12131336 TI - 2001 American Urological Association Gallup Survey: changes in physician practice patterns, satisfaction with urology, and treatment of prostate cancer and erectile dysfunction. AB - PURPOSE: The Health Policy Survey and Research Committee of the American Urological Association and the Gallup organization have performed 9 surveys of American urologists since 1992 for the purpose of assessing demographics and practice patterns. The results of the 2001 survey are presented. MATERIALS AND METHODS: A random sample of 507 urologists was interviewed in February and March 2001. Major content areas were physician practice patterns, cryosurgery/brachytherapy, prostate specific antigen, erectile dysfunction, Medicare and the Internet. RESULTS: Membership in the American Urological Association continues to increase among American urologists. The number of patients seen weekly in the office also continues to increase. While age at retirement has not changed significantly, most urologists are satisfied with the specialty and increasing numbers are using the Internet. CONCLUSIONS: Minimally invasive procedures such as brachytherapy for prostate cancer continue to proliferate and there is evidence that the specialty of urology is continuing to become more office based. The demand for urological services appears to be continuing to increase. PMID- 12131337 TI - Highlights of the Society of Urologic Oncology meeting, June 2, 2001. PMID- 12131338 TI - Re: Percutaneous radio frequency ablation of small renal tumors: initial results. PMID- 12131339 TI - Re: Bilateral laparoscopic inguinal hernia repair can complicate subsequent radical retropubic prostatecomy. PMID- 12131340 TI - Re: Plasma selenium level before diagnosis and the risk of prostate cancer development. PMID- 12131341 TI - Re: Skeletal fracture associated with androgen suppression induced osteoporosis: the clinical incidence and risk factors for patients with prostate cancer. PMID- 12131342 TI - Re: The chronic prostatitis-chronic pelvic pain syndrome can be characterized by prostatic tissue pressure measurements. PMID- 12131343 TI - "Cup-patch" technique of ileocystoplasty for bladder enlargement or partial substitution. PMID- 12131345 TI - Subcapsular urinoma: an unusual form of page kidney in a high school wrestler. PMID- 12131346 TI - Laparoscopic management of page kidney. PMID- 12131347 TI - Development of renal function after neonatal urinary ascites due to obstructive uropathy. AB - PURPOSE: Neonatal urinary ascites is a rare complication of obstructive uropathy with possible lethal outcome if not treated adequately. We demonstrate that with adequate therapy the survival rate can be high and long-term survival, kidney function and lower urinary tract function of patients with urinary ascites can be good. MATERIALS AND METHODS: The study included 4 females and 8 males born with urinary ascites. Followup ranged from 3 to 14 years. Blood analysis for renal function, electrolytes and blood gas was performed at hospitalization and during followup. Ultrasound, cystourethrograms or cystoscopy showed the site of obstruction and leakage of urine. All patients were initially treated with drainage of the ascites and decompression of the obstructed urinary tract. All patients underwent surgery to remove the obstruction and reconstruct the urinary tract. Bladder and kidney function was evaluated at long-term followup. RESULTS: All patients had severe abdominal distention at presentation. Severe metabolic acidosis was present in cases that had not been detected prenatally or immediately after birth. Two patients died of causes related to pulmonary hypoplasia. Surprisingly urinary continence and renal function were good in 9 of 10 survivors. CONCLUSIONS: Long-term outcome of bladder and kidney function is surprisingly good in cases of severe obstructive uropathy with ascites. Intrauterine pressure relief of the bladder through urinary extravasation protects renal function and this decompression of the urinary tract prevents severe secondary changes to bladder function. Although not proven we believe that high intrauterine pressures in the abdominal cavity are prevented by peritoneal absorption of the extravasated urine and consequent dialysis through the placenta. PMID- 12131348 TI - Fate of the retained ureteral stump after upper pole heminephrectomy in duplex kidneys. AB - PURPOSE: We review the long-term outcome of retained ureteral stumps in children undergoing heminephrectomy for nonfunctioning upper pole moieties in duplex kidneys. MATERIALS AND METHODS: The medical records of 50 patients who underwent 50 upper pole heminephrectomies for a nonfunctioning upper pole moiety of a duplex kidney between January 1990 and December 2000 were reviewed retrospectively. RESULTS: Median patient age at heminephrectomy was 2.5 years (range 3 weeks to 16.5 years) and median followup was 6 years (range 1 to 11). Indications for heminephrectomy in the 50 renal units were obstructive ureterocele in 25 (50%) cases, ectopic ureter in 15 (30%), obstructive megaloureter in 5 (10%) and reflux nephropathy in 5 (10%). A total of 48 (96%) of the corresponding ureters were taken down as low as possible and transfixed through the heminephrectomy incision. Residual stump excision was required in 5 (10%) of the 50 units for recurrent urinary tract infection due to vesicoureteral reflux. CONCLUSIONS: Our long-term followup suggests that the majority of patients with residual ureteral stumps after upper pole heminephrectomy do not require stump resection. PMID- 12131349 TI - Expression and prognostic value Of CD44 isoforms in nephroblastoma (Wilms tumor). AB - PURPOSE: CD44 is a group of transmembranous glycoproteins formed by alternative splicing of a single messenger RNA. An abnormal pattern of CD44 expression has been demonstrated in several human malignancies. We evaluate the prognostic value of standard CD44 (CD44s) and some of its isoforms in treating clinical Wilms tumor. MATERIALS AND METHODS: The immunohistochemical expression of CD44 isoforms was studied in paraffin material of 61 clinical Wilms tumors. Patients were treated preoperatively with chemotherapy and mean followup was 5.7 years. RESULTS: Generally CD44s, CD44v5 and CD44v10 were expressed in normal kidney tissues and at variable levels in the 3 cell types of Wilms tumor (blastemal, epithelial and stromal). No CD44v6 expression was found neither in normal kidney or in tumor tissue. CD44s, CD44v5 and CD44v10 epithelial expression gradually decreased from stage T1 to T3. By contrast the percentage of CD44 positive cells in the blastemal component significantly increased from T1 to T3. CD44 immunoreactive blastema cells were found in 62%, 44% and 41% for CD44s, CD44v5 and CD44v10, respectively. Blastemal expression of CD44s and CD44v5 statistically significantly correlated with clinicopathological stage. Univariate and multivariate analyses showed that blastemal CD44v5 expression was indicative of poor prognosis. CONCLUSIONS: Increased expression of CD44v5 in the blastemal part of Wilms tumor correlated with tumor stage, clinical progression and tumor related death. Therefore, blastemal CD44v5 expression may be of value in identifying patients with a high propensity for distant metastases. These patients might benefit from adjuvant chemotherapy and/or radiotherapy. PMID- 12131350 TI - New perspectives on therapy for vaginal endodermal sinus tumors. AB - PURPOSE: Malignant germ cell tumors account for 3% of childhood cancers. Endodermal sinus tumor, the most common malignant germ cell tumor, requires treatment primarily with chemotherapy and surgery is reserved as a last resort. It is rare for the vagina to be the primary site of endodermal sinus tumor, and we report on our experiences with this phenomenon at a single institution. MATERIALS AND METHODS: We retrospectively reviewed the clinical features, treatment and outcomes of 3 children with vaginal endodermal sinus tumor. RESULTS: Initial treatment was combination chemotherapy, in 2 patients. alpha fetoprotein decreased rapidly and returned to normal in both. One patient is disease-free 7 years after chemotherapy and the other patient is currently disease-free with a normal alpha-fetoprotein 3 months after induction chemotherapy. In the remaining case progressive disease developed following initial chemotherapy and subsequent salvage surgery combined with radiation, chemotherapy and ultimately autologous bone marrow transplant was performed. The patient has remained disease-free for 6.5 years since completing this extensive therapy. CONCLUSIONS: Endodermal sinus tumor of the vagina is rare. All of our patients presented with painless bleeding of no obvious source. In such cases one must maintain a high index of suspicion for possible underlying pathological conditions even if ultrasound is negative. Evaluation must include endoscopic examination of the lower genitourinary tract. Bone marrow transplant should be considered as a last therapeutic resort in salvage cases of unresponsive vaginal endodermal sinus tumor. PMID- 12131351 TI - Extensive upper and mid ureteral loss in newborns: experience with reconstruction in 2 patients. AB - PURPOSE: We describe our experience with reconstruction of the ureter in 2 patients who sustained extensive upper and mid ureteral loss as newborns. MATERIALS AND METHODS: Two male patients, a 1-month-old and a neonate, sustained extensive ureteral loss due to candidal infection involving the retroperitoneum and ureter. The 1-month-old sustained a loss of the middle third of the ureter, and the neonate sustained a 3 cm. loss of the upper ureter. The first case was managed with a combination of renal mobilization and an extensive Boari flap, while the second was managed with renal mobilization and nephropexy with primary ureteropyelostomy. RESULTS: Both patients had a successful outcome with no evidence of anastomotic stenosis or obstruction. CONCLUSIONS: Extensive upper and middle third ureteral defects may be primarily bridged successfully in pediatric patients using the standard technique of renal mobilization combined with ureteropyelostomy and a Boari flap, respectively. PMID- 12131352 TI - Intravesical morphine analgesia is not effective after bladder surgery in children: results of a randomied double-blind study. AB - PURPOSE: Intravesical morphine was recently recommended to reduce postoperative pain after reimplantation surgery for vesicoureteral reflux in children. The efficacy of such treatment, so far solely evaluated by open study, needed to be confirmed. MATERIALS AND METHODS: After parental informed consent was obtained, 80 children requiring Cohen cross-trigonal reimplantation were considered for inclusion in a double-blind study. On the day of surgery patients were randomly assigned to receive either 0.04 mg./kg. morphine per hour or placebo (normal saline) at a constant intravesical infusion rate of 0.08 ml./kg. per hour. Postoperative pain was assessed every 3 hours using a pain score adapted to patient age. If the score was above a predefined limit, patients received intravenous acetaminophen and nalbuphine alternately every 3 hours. Bladder infusion was discontinued after 48 hours. RESULTS: Mean and maximum pain scores as well as the number of scores above the limit were not statistically different when comparing the morphine and placebo groups. There was no difference in the number of doses of analgesics administered. Urine output, voiding frequency and the number of painful voiding episodes were not significantly different between the 2 groups. Plasma morphine concentrations were 3.0 +/- 2.7 and 1.9 +/- 1.9 ng./ml. at 24 and 48 hours in the morphine group and undetectable in the placebo group. CONCLUSIONS: Intravesical administration of morphine is not effective for relieving postoperative pain during the first 48 hours after intravesical ureteral reimplantation. This study emphasizes the importance of controlled studies in evaluating the effectiveness of a new drug or procedure before recommending its use for all patients. PMID- 12131353 TI - Histological findings after colocystoplasty and gastrocystoplasty. AB - PURPOSE: We conducted a prospective, long-term assessment of the histological changes that can occur following bladder augmentation with colon or stomach. MATERIALS AND METHODS: Histological evaluations of biopsies from 44 consecutive patients undergoing augmentation (colocystoplasty in 26, gastrocystoplasty in 18) were performed. Patients underwent endoscopic assessment and tissue sampling at 2 or 4-year intervals following the initial augmentation procedure. Patients with less than 2 years of followup were excluded from the analysis. Specimens were taken from the native bladder, the augment segment (large bowel or stomach) and the anastomotic line. Sections (4 mu.) were examined using standard histological staining methods (hematoxylin and eosin and periodic acid-Schiff) and immunohistochemistry was performed for different markers of neoplasia, cellular proliferation and blood group antigens. Histological findings were correlated with the incidence of stone formation and urinary tract infection. RESULTS: Group 1 consisted of 20 patients undergoing colocystoplasty who met the criteria for study inclusion. Of the patients 10 (50%) had stones, 19 (95%) had a positive urine culture and 6 had no histological changes. While no cases of malignancy were identified, other forms of pathological change were noted in 14 of the 20 patients (70%). Group 2 included 15 patients undergoing gastrocystoplasty who met the criteria for study inclusion. No stones or malignancy were identified in this group. Positive urine cultures were recorded in 2 patients (13%), no histological changes were found in 6 and 9 (60%) had pathological changes. CONCLUSIONS: Periodic prospective biopsy evaluation of children who have undergone either colocystoplasty or gastrocystoplasty failed to reveal any histological evidence of malignancy after 10-year followup. However, histological evidence of a premalignant lesion 13 years after followup suggests that screening for premalignant lesions should be initiated no later than 6 to 10 years following enterocystoplasty. PMID- 12131354 TI - Foreign body reaction to bovine pericardium: a previously unreported complication of pediatric chordee repair. PMID- 12131355 TI - Long-term followup of 158 young adults surgically treated for vesicoureteral reflux in childhood: the ongoing risk of urinary tract infections. AB - PURPOSE: We recorded urinary tract infections in the long term after surgical reflux correction. MATERIALS AND METHODS: A total of 158 of 189 patients (160 females and 29 males) who were followed in 1985, an average of 10.8 years after reflux surgery were contacted again in 1995. At that time median patient age was 26 years (range 15.7 to 38.8) and the average period of observation was 20.3 years (range 13.4 to 26). RESULTS: In 82% of the patients febrile and in 18% afebrile symptomatic urinary tract infections had developed preoperatively. In the first 10-year period after operation 46% of patients continued to have symptomatic urinary tract infections compared with 52% in the second 10-year interval. In the 2 periods the incidence of febrile urinary tract infection was about 17%. In the whole postoperative observation period symptomatic urinary tract infections developed in 66% of all patients, including 74% of female patients. Symptomatic urinary tract infections were observed during 8 of 46 pregnancies (17%). CONCLUSIONS: After successful surgical reflux correction susceptibility to urinary tract infection continues for a number of years in many girls and women. However, postoperatively urinary tract infections are primarily afebrile. PMID- 12131356 TI - Risk of urinary tract infections in adults surgically treated for vesicoureteral reflux in childhood. PMID- 12131357 TI - Changing patterns of keratin expression could be associated with functional maturation of the developing human bladder. AB - PURPOSE: We investigate the keratin phenotype of human transitional epithelium at various gestational ages and whether keratin composition of transitional epithelium is related to bladder function and morphology. MATERIALS AND METHODS: Consecutive sections from formalin fixed paraffin embedded blocks of autopsy bladder tissue from 21 male and 5 female fetuses, gestational age 12 to 40 weeks and 7 infants 2 days to 19 months old were cut and stained with antibodies recognizing basal cell keratins 5, 14 and 17, intermediate squamous cell keratin 13 and columnar cell keratins 7, 8, 18 and 20. RESULTS: With gestational age there were distinct changes in expression of keratins recognizing columnar cells, consisting of focal loss of keratin 7 in transitional epithelium, restriction of keratin 20 expression to umbrella cells and expression of keratin 18 throughout the full thickness of transitional epithelium. Basal cell keratin 5 was found above the basal cell layer while keratins 14 and 17 were not found. Squamous cell keratin 13 was found throughout the full thickness of the urothelium. CONCLUSIONS: The changes with gestational age in expression of some keratins may be related to the development of the reservoir function of the bladder. The impermeability of transitional epithelium, particularly during early fetal development, is possibly a function of umbrella and intermediate transitional cells. PMID- 12131358 TI - Is bladder dysfunction and incontinence associated with ureteroceles congenital or acquired? AB - PURPOSE: Bladder dysfunction (a disorder often characterized by incontinence, urgency, patterns of dysfunctional voiding, incomplete emptying and so forth) in association with ureteroceles has been attributed to surgical intervention. A previous study suggested that patients with ectopic ureteroceles may have bladder dysfunction as part of this disorder regardless of the type of surgical intervention. We reviewed all types of ureteroceles (ectopic versus intravesical, simple versus duplex) to characterize the patterns of bladder dysfunction and its association with prior surgical treatments. MATERIALS AND METHODS: A retrospective review of medical records was performed as part of a multi institutional study. From 1986 to 2000, 616 patients were identified with ureteroceles. Bladder dysfunction was determined by detailed history (that is, voiding diary) plus urodynamic evaluation when deemed appropriate. RESULTS: Based on initial history, 39 of 616 (6.3%) patients had some form of bladder dysfunction and 34 of the 39 underwent urodynamics. All patients had ectopic ureteroceles of duplex systems. The most common symptoms of bladder dysfunction were urinary urgency and incontinence. Infrequent voiding, less than 4 voids daily, occurred in 13% (5 of 39) of the patients. Of the 33 incontinent patients 7% (2) had undergone endoscopic surgery, 12% (4) open lower tract surgery, 45% (15) a combination of upper and lower tract surgery and 36% (12) open upper tract surgery alone. Bilateral ureteroceles did not seem to increase the risk of bladder dysfunction. The majority (35 of 39) of patients with bladder dysfunction responded to behavioral modifications and medical therapy. CONCLUSIONS: Bladder dysfunction associated with ureteroceles occurs in approximately 6% of patients regardless of surgical therapy. The fact that patients treated with upper tract surgery alone have similar rates of incontinence suggests that bladder dysfunction is congenital as opposed to surgically acquired. PMID- 12131359 TI - Endocrine screening in 32 consecutive patients with hypospadias. AB - PURPOSE: Various endocrine studies performed in the hypospadias population show an unsatisfactory response to the human chorionic gonadotropin (HCG) test and abnormal androgen biosynthesis with possible enzyme defects. We evaluated the incidence of disorders in androgen production in boys with isolated hypospadias. MATERIALS AND METHODS: A total of 32 consecutive children (46,XY) with hypospadias were prospectively enrolled in the study. Severity of the defect was assessed with a new classification based on the location of the division of the corpus spongiosum. Endocrine evaluation consisted of measuring luteinizing hormone, follicle-stimulating hormone, anti-mullerian hormone (AMH), testosterone, dihydrotestosterone, progesterone, 17alpha-hydroxypregnenolone, 17alpha-hydroxyprogesterone, dehydroepiandrosterone sulfate and delta4 androstenedione. In all but 3 patients gonadal stimulation with 1,500 IU HCG every other day for 12 days was performed and steroid concentrations were reassessed after the test. An adrenocorticotropic hormone test was performed in 2 patients and molecular study of the androgen receptor was performed in 28. RESULTS: An increase to 37.37 nmol./l. progesterone (normal 0.1 to 0.5) and 17alpha-hydroxyprogesterone to 25.48 nmol./l. (normal 1.18 +/- 0.66) before HCG stimulation was noted in 1 patient. These abnormal results were not found after HCG stimulation but reappeared after the adrenocorticotropic hormone test. This result might be related to a partial mix of 17alpha-hydroxylase/17,20-lyase deficiency but no mutation was found after complete sequencing of gene CYP17. Of the 32 patients 4 had an insufficient response to HCG stimulation (testosterone less than 10 nmol./l.), including 1 with a low AMH level of 180 pmol./l. (normal 451 +/- 198) and an increased dehydroepiandrosterone sulfate level of 1,995 nmol./l. (normal 59 +/- 41) before HCG stimulation. Partial androgen insensitivity was suspected in 1 patient because he had a high testosterone response (29.96 nmol./l.) after HCG stimulation but no mutation of the gene of the androgen receptor was detected. Two patients with proximal hypospadias had isolated decreased AMH levels, which was evidence of Sertoli cell insufficiency. CONCLUSIONS: Although our series of 32 patients had several abnormal endocrine screenings, these results indicate no significant endocrine defects. PMID- 12131361 TI - The effect of varicocele repair on testicular volume in children and adolescents with varicocele. AB - PURPOSE: We investigated the effect of varicocele repair on testicular volume according to age in children and adolescents and review the long-term results of varicocele surgery. MATERIALS AND METHODS: The study included 39 boys 11 to 19 years old with clinical palpable varicocele who underwent varicocele surgery with at least 1 year of postoperative followup. Preoperative and postoperative testicular volumes were monitored and measured with an ellipsoid Prader orchidometer. Physical examination findings (testicular volumes and testicular consistency) in all boys, and serum hormone values and semen parameters in 16 adolescents were recorded and compared before and after surgery. RESULTS: Left unilateral varicocelectomy was done in 29 boys (74%) and bilateral varicocelectomy in 10 (26%). While no postoperative hematoma, infection or testicular atrophy was observed, 1 boy (2.5%) had varicocele recurrence and 2 boys (5.1%) had minimal hydroceles that required no intervention. Significant increases were observed in postoperative sperm concentration (p = 0.01), total motile sperm count (p = 0.009), testis volume (p = 0.000) and serum testosterone level (p = 0.014). All 15 boys with preoperative soft testis had normal testicular consistency postoperatively. Of the 19 boys with preoperative testicular atrophy 10 (53%) did regain normal testicular growth, while 9 (47%) retained testicular volume loss after surgery. When comparing preoperative to postoperative increase in testicular volume according to age in all boys, the mean was statistically significantly higher in boys younger than 14 years (left testis p = 0.037, right testis p = 0.000). CONCLUSIONS: Testicular consistency achieved normal firmness after varicocelectomy in all boys with preoperative soft testis. While there was catch-up growth in comparison to the contralateral testis, testicular consistency improved but testicular volumes may not increase significantly after varicocele repair at ages older than 14 years. However, in these adolescents postoperative semen parameters and serum hormone values may significantly improve regardless of testicular volume. Therefore, boys with varicocele and their families should be fully informed in light of these findings. PMID- 12131360 TI - Variables in successful repair of urethrocutaneous fistula after hypospadias surgery. AB - PURPOSE: We evaluate variables affecting the success of repairs of urethrocutaneous fistula after hypospadias surgery. MATERIALS AND METHODS: The records of 123 boys who underwent fistula repair at Primary Children's Medical Center were reviewed. Of these patients 100 underwent initial fistula repair at our center (surgery was performed at our center in 82 and elsewhere in 18) and 23 were referred from elsewhere after unsuccessful fistula repairs. Patient age was 6 months to 34 years (median 3.21 years) and interval between surgeries was 3.7 months to 12 years (median 12.6 months). Several variables potentially affecting the success of fistula closure were retrospectively assessed. RESULTS: Including those patients referred from outside hospitals, fistulas were successfully closed in 71%, 72%, 77%, 100% and 100% of these patients after fistula repairs 1 to 5, respectively. Variables studied yielded stent 67.7% (36 of 54 cases) versus no stent 76.1% (35 of 46) and operating microscope 70.4% (59 of 71) versus loupes 72.4% (21 of 29) in terms of success. Success based on patient age yielded 65.5% for younger than 2 years (n = 29 patients), 71.7% for 2 to 5 (46), 64.7% for 6 to 12 (17) and 87.5% for older than 12 (8). When considering the type of original hypospadias repair and its affect on fistula closure success, a significantly lower success was noted with Yoke and King procedures (p = 0.007 and 0.037, respectively). In patients who underwent hypospadias surgery and all subsequent fistula closure attempts at our center, fistulas were successfully repaired in 72%, 67% and 100% of patients after attempts 1 to 3, respectively. Initial fistula repair was successful in 72% (59 of 82) of patients who underwent original hypospadias surgery at our center and in 67% (12 of 18) of those referred after hypospadias surgery at an outside hospital. CONCLUSIONS: Regarding urethrocutaneous fistula closure, the data from this study suggest that there is no clear difference in stent versus no stent and microscope versus loupes, age at fistula closure does not affect success, type of original hypospadias procedure may influence success (King and Yoke procedures were least successful), success rate is not negatively impacted in recurrent fistula cases, given a diverse group of fistulas, success of fistula repair for attempts 1 to 5 was 71%, 72%, 77%, 100% and 100%, respectively, and success rate in a tertiary pediatric urology setting is not influenced by whether the original hypospadias procedure or initial fistula closure was performed in the pediatric urology setting versus outside hospital. PMID- 12131362 TI - Prostate specific antigen inhibits immune responses in vitro: a potential role in prostate cancer. AB - PURPOSE: Prostate specific antigen (PSA) is found in high concentration in prostate tissue and in semen, in which its physiological function appears to be liquefaction. In prostate cancer the peripheral PSA concentration is elevated, which may be used as a disease marker. Systemic and local immune defects have been demonstrated in prostate cancer and we postulated a role for PSA in this immunosuppression. We explored the effects of PSA on human T-lymphocyte proliferation in vitro. MATERIALS AND METHODS: PSA was purified from normal seminal plasma using a modified chromatographic technique. The effect of PSA or control protein on lymphocyte responses to mitogens, tetanus toxoid and alloantigens was tested. The inhibitory effect observed was further explored by varying the time of PSA addition, denaturing PSA and including interleukin-2 and anti-PSA antibodies. RESULTS: PSA suppressed in vitro phytohemagglutinin and alloantigen stimulated lymphocyte proliferation in a dose dependent manner. This effect was reversed by adding anti-PSA antibodies but not by interleukin-2. CONCLUSIONS: These in vitro PSA effects suggest another T-lymphocyte mediated immunosuppressive mechanism. In vivo high levels of PSA may compromise natural immune responses to cancer and current attempts at immunotherapy for prostate cancer. PMID- 12131363 TI - Resveratrol induced serine phosphorylation of p53 causes apoptosis in a mutant p53 prostate cancer cell line. AB - PURPOSE: Resveratrol (Calbiochem, La Jolla, California) is a naturally occurring stilbene reported to cause apoptosis in various cultured cancer cells. In the current study the effect of resveratrol was determined in the androgen insensitive DU 145 prostate cancer cell line. Induction of apoptosis and activation of apoptosis related signal transduction pathways were measured. MATERIALS AND METHODS: DU 145 cells were treated with resveratrol and apoptosis was measured by determining nucleosome content. Activation of mitogen activated protein kinase (MAPK) (extracellular signal-regulated kinase 1/2), p53 content and serine-15 phosphorylation of p53 were measured by immunoblot. Electrophoretic mobility shift assay of p53 binding to DNA, and measurement of p21 and glyceraldehyde-3-phosphate dehydrogenase messenger RNA were also done. RESULTS: Resveratrol induced apoptosis in DU 145 cells. The stilbene activated MAPK and caused increased abundance of p53 and serine-15 phosphorylated p53. Resveratrol induced serine-15 phosphorylation of p53 was blocked by PD 98059 (Calbiochem), a MAPK kinase inhibitor, implicating MAPK activation in the phosphorylation of p53. PD 98059 also inhibited resveratrol induced apoptosis. These results suggest that apoptosis induction by resveratrol in DU 145 cells requires serine-15 phosphorylation of p53 by MAPK. Inhibition of MAPK dependent serine-15 phosphorylation resulted in reduced p53 binding to a p53 specific oligonucleotide on electrophoretic mobility shift assay. Pifithrin-alpha (Calbiochem), a p53 inhibitor, blocked resveratrol induced serine-15 phosphorylation of p53 and p53 binding to DNA. Resveratrol caused a p53 stimulated increase in p21 messenger RNA. Transfection of additional wild-type p53 into DU 145 cells induced apoptosis, which was further enhanced by resveratrol treatment. CONCLUSIONS: Resveratrol causes apoptosis in DU 145 prostate cancer cells. This action depends on the activation of MAPK, increase in cellular p53 content, serine-15 phosphorylation of p53 and increased p53 binding to DNA. PMID- 12131364 TI - Calcitriol (1,25-dihydroxycholecalciferol) potentiates activity of mitoxantrone/dexamethasone in an androgen independent prostate cancer model. AB - PURPOSE: Mitoxantrone combined with glucocorticoids is widely used for androgen independent prostate cancer. It is well tolerated, reduces prostate specific antigen, diminishes pain and improves quality of life. Calcitriol (1,25 dihydroxycholecalciferol) inhibits proliferation, modulates cell cycle progression, induces apoptosis and potentiates the cytotoxic effects of a number of agents. Glucocorticoids potentiate the antitumor effects of calcitriol and blunt calcitriol induced hypercalcemia. Therefore, we investigated the effect of calcitriol on the antitumor efficacy of mitoxantrone and dexamethasone or mitoxantrone/dexamethasone in the PC-3 androgen independent prostate cancer model. MATERIALS AND METHODS: We treated PC-3 cells in vitro with various concentrations of mitoxantrone/dexamethasone with and without calcitriol, and assessed growth inhibition by crystal violet assays. We similarly treated mice bearing PC-3 xenografts and performed excision clonogenic assays and tumor outgrowth studies to assess antitumor activity. RESULTS: Calcitriol significantly increased mitoxantrone/dexamethasone mediated growth inhibition in PC-3 cells (p <0.05). Median dose effect analysis indicated that calcitriol is synergistic with mitoxantrone. Adding calcitriol to mitoxantrone/dexamethasone significantly reduced the surviving fraction per gm. tumor compared with mitoxantrone/dexamethasone or untreated controls (p <0.03). Calcitriol plus mitoxantrone/dexamethasone also caused significantly greater tumor regression in PC-3 xenografts compared with treatment with mitoxantrone/dexamethasone or untreated controls (p <0.02). CONCLUSIONS: These preclinical data demonstrate that calcitriol increases the antitumor activity of mitoxantrone/dexamethasone in the PC-3 model system. This combination may be efficacious for prostate cancer. PMID- 12131365 TI - Activation of signal transducer and activator of transcription 3 in renal cell carcinoma: a study of incidence and its association with pathological features and clinical outcome. AB - PURPOSE: Signal transducer and activator of transcription 3 (STAT3) is known to have an important role in cytokine and growth factor signaling pathways. In various types of human malignant tumors STAT3 has been shown to be constitutively activated due to aberrant production of cytokines and growth factors. We examined the presence of STAT3 activation and its association with pathological features and clinical outcome in renal cell carcinoma cases. MATERIALS AND METHODS: We examined 48 paraffin embedded renal cell carcinoma specimens and corresponding nonneoplastic kidney tissues for the activation status of STAT3 on immunohistochemistry using anti-phospho-specific (p)-STAT3 antibody, which recognizes only activated STAT3. Based on the percent of cells with positive nuclear staining the activation status of STAT3 was determined and categorized into 2 groups, including low-less than 10% and high-90% or more tumor cells positive. The associations of the activation status of STAT3 with pathological features and clinical outcome were analyzed. RESULTS: Of 48 tumors 24 (50%) demonstrated high levels of nuclear immunostaining for p-STAT3, while the other 24 (50%) showed low levels. Adjacent nonneoplastic kidney tissues showed only little immunostaining for p-STAT3. A significant association of high levels of p STAT3 with metastasis was observed (p = 0.0094). No significant associations of p STAT3 immunostaining with pathological stage or grade were observed. A high level of p-STAT3 was a significant indicator of a poor prognosis on univariate and multivariate analysis (p = 0.0117 and 0.0439, respectively) CONCLUSIONS: Our results indicate a high frequency of STAT3 activation in renal cell carcinoma, especially in metastatic disease. STAT3 activation was an independent prognostic variable in renal cell carcinoma cases. Our results strongly suggest that the activation of STAT3 contributes to the development and progression of renal cell carcinoma. PMID- 12131366 TI - A recombinant adenovirus expressing p7(Kip1) induces cell cycle arrest and apoptosis in human 786-0 renal carcinoma cells. AB - PURPOSE: We evaluated the effects of the over expression of p27Kip1, a cyclin dependent kinase inhibitor and tumor suppressor protein, on the 786-0 human renal carcinoma cell line. MATERIALS AND METHODS: The recombinant adenovirus Adp27Kip1 was evaluated for the induction of p27 protein expression in 786-0 renal carcinoma cells. Expression time and optimal vector concentration were determined. Growth curve studies, cell cycle analysis and terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end-labeling were done to determine the effects of p27Kip1 on the cell cycle. Cyclin dependent protein kinase (Cdk) inhibitor (CDKI) activity assays were done to determine the expression/activities of Cdks and Western blot analysis was performed to determine the presence of CDKIs and other cell cycle regulator proteins. Nude mouse xenografts were established to demonstrate the in vivo efficacy of Adp27Kip1. RESULTS: p27Kip1 protein expression was detected within 12 hours after Adp27Kip1 infection and it remained stable for at least 48 hours. Growth studies demonstrated that Adp27Kip1 infection resulted in the inhibition of proliferation by 3 days after infection and cell death was detected by day 5. Cell cycle analysis of DNA content indicated an accumulation of cells in the G1 phase of Adp27Kip1 infected cells and a corresponding decrease in S phase cells within 48 hours after infection. Cdk activity was determined, and Cdk2, Cdk4 and Cdc2 kinase activities were inhibited, consistent with p27Kip1 over expression. The levels of the CDKIs p16 and p18 were elevated 24 hours after Adp27Kip1 infection, while p21 levels remained unchanged. Terminal deoxynucleotidyl transferase mediated deoxyuridine triphosphate nick end-labeling revealed that Adp27Kip1 infection but not infection by control virus induced detectable apoptosis within 24 hours. Adp27Kip1 significantly caused the reduction in the size of tumors of the renal cell carcinoma xenografts. CONCLUSIONS: This study demonstrates the potential effectiveness of Adp27Kip1 as a vector for gene therapy studies of renal cell carcinoma. PMID- 12131367 TI - Characterization of a renal cell carcinoma cell line derived from a human bone metastasis and establishment of an experimental nude mouse model. AB - PURPOSE: We provide the necessary tools to study the biology of bone metastasis using renal cell carcinoma as a model. A relevant renal cell carcinoma cell line developed from a human bone metastasis is described and an experimental model in the nude mouse was established. MATERIALS AND METHODS: The novel cell line RBM1 was developed from bone metastasis from a patient with renal cell carcinoma. In vitro methods used to study the cell line included karyotype analysis, reverse transcriptase polymerase chain reaction, ribonuclease protection assay and Western blot analysis. An intratibial injection model in the nude mouse resulted in bone lesions similar to those in human patients. RESULTS: Chromosomal analysis of this cell line revealed characteristic cytogenetic abnormalities common in renal cell carcinoma. RBM1 cells expressed parathyroid hormone parathyroid hormone related peptide, interleukin-6 and macrophage colony-stimulating factor, which are cytokines involved in osteoclast activation and subsequent bone resorption. Western blot analysis revealed the presence of high levels of epidermal growth factor receptor and c-MET. Reproducible osteolytic lesions developed in the nude mouse after injecting 1 x 106 cells into the tibia. CONCLUSIONS: This cell line and animal model may allow further study of the biology and mechanism of renal cell carcinoma bone metastasis. PMID- 12131368 TI - N-acetylcysteine augments the cellular redox changes and cytotoxic activity of internalized mycobacterium bovis in human bladder cancer cells. AB - PURPOSE: We determined whether changes in cellular reactive oxygen species correlated with mycobacteria internalization and bladder cancer cell death. MATERIALS AND METHODS: Reactive oxygen species and thiols in RT112 and MGH bladder cancer cells were determined using the fluorescence probes 5-(and 6) carboxy-2', 7' dichlorodihydrofluorescein diacetate and monobromobimane. Superoxide and nitrite production were measured using bis-N-methylarcridinium nitrate and Griess reagents. Cytotoxicity was determined by the release of 14C thymidine from cells with 14C labeled DNA. RESULTS: MGH cells that internalize bacillus Calmette-Guerin (BCG) had decreased cellular reactive oxygen species and thiols, although superoxide and nitric oxide production increased. RT112 cells, which do not internalize BCG, did not show a decrease in reactive oxygen species after incubation with BCG. Blocking BCG uptake in MGH cells abrogated reactive oxygen species reduction, confirming that the changes in reactive oxygen species were internalization dependent events. Treating cells with BCG and the antioxidant N-acetylcysteine caused a greater reduction in reactive oxygen species, and induced earlier and greater cytotoxicity in MGH but not in RT112 cells. CONCLUSIONS: The induction of bladder cancer cell killing by BCG parallels the ability of cells to internalize BCG, which in turn indicates that the susceptibility of tumor cells to the cytotoxic effects of BCG may be related to changes in cellular levels of reactive oxygen species and thiols. Supplementation with an antioxidant could enhance the antitumor effect of BCG. PMID- 12131369 TI - Interleukin-6 production by human bladder tumor cell lines is up-regulated by bacillus Calmette-Guerin through nuclear factor-kappaB and Ap-1 via an immediate early pathway. AB - PURPOSE: The expression of interleukin-6 (IL-6) by normal and malignant urothelium in response to bacillus Calmette-Guerin (BCG) may have direct and indirect effects on the antitumor activity of BCG. We evaluated the molecular signaling pathway through which BCG induces IL-6 expression in human transitional cell carcinoma lines. MATERIALS AND METHODS: We evaluated IL-6 messenger RNA and protein expression by human transitional carcinoma cell lines in response to BCG. Pharmacological inhibition of protein synthesis was used to determine if BCG mediated IL-6 induction occurred via an immediate-early or delayed pathway. We used 5' deletion analysis and site directed mutagenesis to identify BCG responsive regions in the human IL-6 promoter. Electrophoretic mobility shift assays were done to assess nuclear translocation of the putative signaling proteins AP-1 and nuclear factor-kappaB (NF-kappaB) in response to BCG. RESULTS: BCG increased IL-6 messenger RNA and protein in a time and dose dependent manner. IL-6 induction by BCG occurred via an immediate-early response. Promoter analysis identified 2 areas in the -1,200 to 14, 5' region of the IL-6 gene, which when deleted were associated with significant losses of absolute or BCG responsive activity. Site specific mutation of putative AP-1 or NF-kappaB elements associated with each region demonstrated that these elements were necessary but not sufficient for BCG induced IL-6 transcription. Gel mobility shift assays showed that AP-1 and NF-kappaB were induced in response to BCG exposure. CONCLUSIONS: Our results show that BCG induced IL-6 expression by human transitional cell neoplasms occurs as an immediate-early gene pathway that requires NF-kappaB and AP-1. PMID- 12131370 TI - The bladder acellular matrix graft in a rat chemical cystitis model: functional and histologic evaluation. AB - PURPOSE: We investigated the feasibility of augmentation in a diseased bladder with a bladder acellular matrix graft. MATERIALS AND METHODS: In 50 female Sprague-Dawley rats chemical cystitis was induced by intravesical instillation of HCl repeated monthly to maintain chronic inflammation. Urodynamic studies were performed in all rats 1 week after the induction of chemical cystitis and repeated at sacrifice. The 29 rats in the experimental group underwent partial cystectomy (50% or greater), followed by bladder acellular matrix graft augmentation, while the 21 controls underwent monthly HCl instillation only. The rats were sacrificed at 2 weeks, 1, 2 and 3 months, respectively. The bladder was removed and examined for histological changes. RESULTS: Urodynamic studies showed that bladder capacity and compliance were significantly higher in the grafted than in the control group (p = 0.008 and 0.006, respectively, at 3 months). Histological studies revealed urothelial and smooth muscle regeneration within the bladder acellular matrix graft at 1 month and nerve regeneration at 3. The number of mast cells was significantly lower in the grafted region than in the host bladder of all grafted rats (p <0.001). CONCLUSIONS: In this rat chemical cystitis model bladder augmentation with a bladder acellular matrix graft led to functional and histological improvement over diseased host bladder. PMID- 12131371 TI - Extracellular matrix changes in urethral stricture disease. AB - PURPOSE: Glycosaminoglycans (GAGs) and collagen are major components of the extracellular matrix and they have key roles in fibrotic diseases. Little is known about the molecular environment in urethral stricture and the majority of the studies available focused on collagen analysis. However, to our knowledge there are no data on GAG composition in urethral stricture disease. MATERIALS AND METHODS: Bulbar urethral strictured segments were obtained from 10 patients 18 to 61 years old (mean age 41.8) who underwent end-to-end anastomotic urethroplasty. GAGs in dry tissue samples were extracted by papain digestion and cetylpyridinium chloride/ethanol precipitation. The concentration of total GAGs was assessed by hexuronic acid assay and expressed in microg. hexuronic acid per mg. dry tissue, while the proportion of sulfated GAGs was determined by agarose gel electrophoresis. The concentration of hyaluronic acid was determined by ion exchange chromatography and total tissue collagen was estimated as its hydroxyproline content. The control group consisted of 10 bulbar urethras obtained from fresh normal cadavers 22 to 53 years old (mean age 32.8). RESULTS: Mean total GAG concentration plus or minus standard deviation in the stricture group was 1.09 +/- 0.13, which was significantly lower than in controls (p <0.05). While the predominant GAG in normal bulbar urethras was hyaluronic acid, dermatan sulfate predominated in strictured urethras (mean 44.1% +/- 8.4 and 45.6% +/- 7.7%, respectively). Hyaluronic acid decreased 49.9% and dermatan sulfate increased 68.3%. There were no significant changes in the concentration of heparan sulfate or chondroitin sulfate in normal and strictured bulbar urethras. Mean total collagen significantly increased 32.3% (p <0.05). CONCLUSIONS: Composition changes in GAGs in strictured urethras could contribute to the noncompliant nature of urethral scar tissue and cause functional changes. These results may be useful for defining new targets for therapy for urethral stricture disease. PMID- 12131372 TI - Evaluation of a dissolvable ureteral drainage stent in a Swine model. AB - PURPOSE: Ureteral stents are commonly placed after routine ureteroscopic procedures to prevent acute ureteral obstruction. However, stents can cause significant symptoms, may require a secondary procedure for removal and may possibly be forgotten. Toward this end a temporary ureteral drainage stent capable of dissolving spontaneously was developed to obviate the problems associated with more commonly used stents. MATERIALS AND METHODS: We evaluated 3 stents of various compositions in a swine model. These stents were composed of dense dissolving material with and without radiopaque filling (dense filled and dense unfilled, respectively) and soft material with radiopaque filling (soft filled). Standard soft hydrogel coated plastic stents served as controls. Three female swine were placed into 4 groups each, including group 1-dense unfilled versus standard stents with harvest at day 7, group 2-dense unfilled versus standard stents with harvest at day 3, group 3-dense filled versus dense unfilled stents with harvest at day 2 or 3 and group 4-soft filled versus dense unfilled stents with harvest at day 2 or 3. Stents were placed bilaterally. The radiopaque quality of stent fragments on plain x-ray of the kidneys, ureters and bladder, and excretory urography, stent condition and histological changes of the urinary tract were compared among the groups. RESULTS: Successful cystoscopic placement of all stents was achieved. The standard stent was present at autopsy in all animals in group 1, although the dense unfilled stent was absent in group 1 at day 7. In group 2 the standard stent was in position but the dense unfilled stent was fragmented in 2 of the 3 animals and absent in 1. In groups 3 and 4 (dense filled versus dense unfilled and soft filled versus dense unfilled, respectively) all stents were fragmented in similar fashion on harvest day 2 or 3. Standard stents and filled stent fragments were evident on imaging, although dense unfilled stents were not discernible. Histological testing revealed inflammatory changes associated with all stents with no significant difference among stent types. CONCLUSIONS: These temporary drainage stents appeared to dissolve in benign fashion, as evidenced by the appearance of the collecting system on excretory urography and by histological changes. In addition, inflammatory changes throughout the urinary tract were similar for each stent evaluated. Further analysis is warranted in human subjects to assess the exact time of stent retention, potential adverse symptoms, stent fragment passage and safety. PMID- 12131373 TI - A system for the enhancement of adenovirus mediated gene transfer to uro epithelium. AB - PURPOSE: Recombinant adenovirus has been used widely as an in vivo gene transfer vector, although its transfection efficiency in bladder tissue is limited. Several studies have indicated that the bladder surface glycosaminoglycan (GAG) layer functions as a nonspecific anti-adherence factor and possibly as a first line anti-infection defense mechanism. We determined whether recombinant adenovirus mediated gene transfer could be enhanced in intact bladders by HCl pretreatment and by alterations in the GAG layer. MATERIALS AND METHODS: In vitro viral transfection efficiencies with and without the GAG analog pentosan polysulfate (Sigma Chemical Co., St. Louis, Missouri) were determined in bladder muscle and urothelial cells. Immunocytochemical studies and Western blot analysis were performed to determine whether urothelial cells possessed the Coxsackievirus and adenovirus receptor. Rat bladders were intravesically pretreated with HCl at various concentrations and for various periods. After 60 mM. HCl pretreatment for 10 minutes 2 x 109 pfu of recombinant adenovirus carrying the Escherichia coli LacZ gene were intravesically instilled into the bladders. RESULTS: Adenoviral infection of urothelial cells was significantly reduced in the presence of pentosan polysulfate in vitro. Coxsackievirus and adenovirus receptor expression was detected in urothelial cells in vivo and in vitro. Bladders pretreated with HCl resulted in an alteration of the bladder GAG layers. After intravesical gene instillation reporter gene analyses using X-5-bromo-4-chloro-3-inodolyl beta-D galactopyranoside (Sigma Chemical Co.) showed approximately 80% urothelial cell transfection efficiency in bladders pretreated with HCl. However, less than 10% of the urothelial cells expressed the transfected gene in control HCl untreated bladders. CONCLUSIONS: Primary urothelial cells and bladder carcinoma cells can be efficiently transfected using an adenoviral vector with similar infectivity. In vitro viral infection shows that the efficiency of adenoviral transfection is significantly reduced in the presence of pentosan polysulfate, a GAG analog. Adenoviral mediated gene transfer to bladder urothelium is enhanced by HCl pretreatment. PMID- 12131374 TI - Mast cell activation triggers a urothelial inflammatory response mediated by tumor necrosis factor-alpha. AB - PURPOSE: Mast cells have been implicated in bladder inflammation and pathogenesis. To determine if mast cell secretion products can modulate urothelial inflammatory responses we developed an in vitro model of mast cell urothelial cell interactions. MATERIALS AND METHODS: Cultures of the immortalized urothelial cell line TEU-2 were incubated in the conditioned medium of mast cell cultures. The urothelial inflammatory response to mast cell secretion products was then determined by quantifying nuclear factor kappaB activity, the expression of endogenous nuclear factor kappaB dependent genes and the protein expression of inflammation markers. RESULTS: Conditioned medium from RBL-2H3 mast cells induced a 4-fold increase in TEU-2 nuclear factor kappaB activity that was independent of the activation state of the mast cells. In contrast, ribonuclease protection assays revealed that the nuclear factor kappaB dependent transcripts tumor necrosis factor-alpha (TNF-alpha), interleukin (IL) 8 and 1beta, and intracellular adhesion molecule 1 (ICAM-1) were induced by mast cell conditioned medium in a manner that strictly depended on mast cell activation (antigen challenge of IgE sensitized RBL-2H3 cells). The dependence on mast cell activation was confirmed by the observation that IL-8 secretion and ICAM-1 protein expression in TEU-2 cultures were induced only by conditioned medium of stimulated RBL-2H3 cells The induction of TEU-2 IL-8 secretion and ICAM-1 expression by mast cell conditioned medium could be blocked by an anti-TNF-alpha antibody or the cysteine protease inhibitor N-acetyl-leucinyl-leucinyl norleucinal. CONCLUSIONS: Our data support the hypothesis that mast cells may participate in bladder inflammation. Furthermore, TNF-alpha acting via the nuclear factor kappaB signaling pathway may be a mediator of the urothelial response to mast cell secretion products. PMID- 12131375 TI - Optimal treatment of systemic bacillus Calmette-Guerin infection: investigations in an animal model. AB - PURPOSE: Hematogenous spread of bacillus Calmette-Guerin (BCG) after intravesical instillation for bladder cancer is rare but it may result in systemic infection and hypersensitivity reaction. We investigated fluoroquinolones and steroids in an animal model to improve the therapeutic options in local and systemic BCG infection. Furthermore, the antitumor effectiveness of intravesical BCG with simultaneous application of fluoroquinolones and/or steroids was tested. MATERIALS AND METHODS: Oral antimicrobial therapy with and without steroids was started immediately after intraperitoneal injection using fluoroquinolones or trimethoprim-sulfamethoxazole. To evaluate the therapeutic options against a hyperergic reaction after repeat systemic BCG infection re-challenge was performed with intraperitoneal BCG 7 days after primary infection and oral therapy was given with fluoroquinolones or trimethoprim-sulfamethoxazole with and without steroids. The influence of continuous oral fluoroquinolone therapy on the antitumor effect of BCG was also tested in the MB 49 orthotopic murine bladder tumor model. RESULTS: After primary systemic infection fluoroquinolone therapy alone led to significantly prolonged survival in mice (log rank test p = 0.041), whereas trimethoprim-sulfamethoxazole was ineffective. There was no additional effect of steroid administration. Steroids alone led to premature death (log rank test p = 0.022). After secondary BCG infection only steroid treated animals had prolonged survival (log rank test p = 0.032), whereas antimicrobials alone had no effect. The therapeutic efficacy of BCG in the orthotopic bladder tumor model was not affected by continuous oral fluoroquinolones in terms of survival (log rank test p = 0.001) or bladder weight (Wilcoxon test p = 0.001) compared with untreated controls. CONCLUSIONS: In a mouse model fluoroquinolones had a beneficial effect for primary systemic BCG infections, whereas the hyperergic reaction after repeat BCG infection was susceptible only to steroids. Administering fluoroquinolones during an intravesical treatment course does not affect the antitumor efficacy of BCG. PMID- 12131376 TI - Kit positive cells in the guinea pig bladder. AB - PURPOSE: We describe the presence of interstitial cells of Cajal (ICC) throughout the wall of the guinea pig bladder. MATERIALS AND METHODS: Bladders obtained from male guinea pigs were prepared for immunohistochemical investigations using various primary antibodies, including the specific ICC marker c-kit (Gibco BRL, Grand Island, New York). Enzymatically dispersed cells with a branched morphology were identified as ICC using anti-c-kit. They were loaded with fluo 4acetoxymethyl (Molecular Probes, Eugene, Oregon) and studied using confocal laser scanning microscopy. RESULTS: Anti-c-kit labeling demonstrated that ICC were oriented in parallel with the smooth muscle bundles that run diagonally throughout the bladder. Double labeling with anti-smooth muscle myosin (Sigma Chemical Co., St. Louis, Missouri) revealed that ICC were located on the boundary of smooth muscle bundles. When anti-c-kit was used in combination with the general neuronal antibody protein gene product 9.5 (Ultraclone Ltd., Isle of Wight, United Kingdom) or anti-neuronal nitric oxide synthase, it was noted that there was a close association between nerves and ICC. Enzymatic dissociation of cells from tissue pieces yielded a heterogeneous population of cells containing typical spindle-shaped smooth muscle cells and branched cells resembling ICC from other preparations. The latter could be identified immunohistochemically as ICC using anti-c-kit, whereas the majority of spindle-shaped cells were not Kit positive. Branched cells responded to the application of carbachol by firing Ca2+ waves and they were often spontaneously active. CONCLUSIONS: ICC are located on the boundary of smooth muscle bundles in the guinea pig bladder. They fire Ca2+ waves in response to cholinergic stimulation and can be spontaneously active, suggesting that they could act as pacemakers or intermediaries in the transmission of nerve signals to smooth muscle cells. PMID- 12131377 TI - Effects of ZD6169 and ZD0947, 2 potassium adenosine triphosphate channel openers, on bladder function of spinalized rats. AB - PURPOSE: The use of K+ channel openers is emerging as an attractive possibility for treating bladder overactivity. We tested the efficacy of the 2 adenosine triphosphate dependent K channel openers ZD6169 and ZD0947 on detrusor hyperreflexia after spinal cord injury in rats. MATERIALS AND METHODS: Included in this study were 72 adult Sprague-Dawley rats. Six animals served as normal controls, while 66 underwent spinal cord transection at the 10th thoracic vertebra. Two weeks after spinal cord injury 6 animals underwent filling cystometrography to confirm detrusor hyperreflexia, while another 12 served as control paraplegics. For each drug 24 animals were used and divided into 2 equal groups of 12. Group 1 received the drug in a dose of 3 mg./kg. daily, while group 2 received a dose of 0.3 mg./kg. daily. Each control paraplegic and treatment group was further subdivided into 2 subgroups of 6 rats. In subgroup 1 filling cystometrography was done 3 weeks after spinal cord injury, while in subgroup 2 it was done 4 weeks after spinal cord injury. RESULTS: Three weeks after spinal cord injury detrusor hyperreflexia developed in all control paraplegic animals with a mean bladder capacity plus or minus standard deviation of 0.7 +/- 0.2 ml. and a mean voiding pressure of 59 +/- 14.2 cm. water. Detrusor hyperreflexia resolved in 66% of the animals that received ZD6169 for 1 week at either dose. For example, mean bladder capacity was 2.5 +/- 1.8 versus 1.8 +/- 1.2 ml. and mean voiding pressure was 42.1 +/- 15.9 versus 43.2 +/- 21.4 cm. water in animals that received 3 versus 0.3 mg./kg. daily, respectively. All animals that received a dose of 3 mg./kg. ZD0947 daily for 1 week showed no detrusor hyperreflexia with a mean bladder capacity of 2.7 +/- 1.8 ml. and mean voiding pressure of 34 +/- 8.5 cm. water, while at 0.3 mg./kg. daily 83% showed no detrusor hyperreflexia with a mean bladder capacity of 2.5 +/- 2.0 ml. and a mean voiding pressure of 41.5 +/- 13.8 cm. water. Each drug produced better urodynamic results when given for 2 weeks. CONCLUSIONS: ZD6169 and ZD0947 are effective treatment for detrusor hyperreflexia after spinal cord injury and they may provide alternative treatment options for overactive bladder. Each drug has time and dose dependent response effects that reflect their wide range of efficacy. However, ZD0947 shows an efficacy profile that is relatively superior to that of ZD6169. PMID- 12131378 TI - A causative factor of copulatory disorder in rats following social stress. AB - PURPOSE: We investigated the causative role of testosterone in copulatory disorder and the expression of c-fos messenger (m)RNA in the medial preoptic area in rats after social stress. MATERIALS AND METHODS: To generate copulatory disorder rats in the experimental defeated group were attacked by residents for 10 minutes daily for 7 consecutive days (social stress). We then investigated the effect of repeat defeat on the frequency of mounting behavior and plasma testosterone levels. The effects of testosterone replacement and/or apomorphine (100 microg./kg. subcutaneously), a dopamine receptor agonist, on the frequency of mounting behavior were also studied. After experiencing social stress the brain area within the medial preoptic area was removed for analysis of c-fos and androgen receptor mRNA expression. Real-time reverse transcription-polymerase chain reaction was done to analyze gene expression. RESULTS: Rats in the defeated group showed a reduced frequency of mounting behavior and a decrease in plasma testosterone levels compared with values in control rats (p <0.01). After testosterone replacement the frequency of mounting behavior became significantly higher than that of socially stressed rat (p <0.05) but did not achieve control levels. The frequency of mounting behavior by socially stressed rats after apomorphine treatment was significantly higher than that of vehicle treated rats (p <0.05) but the frequency produced by the combination of testosterone replacement and apomorphine injection did not achieve control levels. After the social stress experience c-fos mRNA expression was significantly increased compared with that in control rats (p <0.05). The expression of androgen receptor mRNA was not affected by social stress. Testosterone replacement significantly reduced the expression of c-fos mRNA in the medial preoptic area (p <0.05). CONCLUSIONS: Our results indicate that a reduction in plasma testosterone may have a causative role in copulatory disorder induced by social stress. Changes in c-fos mRNA expression in the medial preoptic area correlated with copulatory disorder and, thus, they are suitable for monitoring that disorder. PMID- 12131380 TI - Spinal cord injury without radiographic abnormality: results of the National Emergency X-Radiography Utilization Study in blunt cervical trauma. AB - BACKGROUND: The purpose of this study was to better define the incidence and characteristics of patients with spinal cord injury without radiographic abnormality (SCIWORA), using the database of the National Emergency X-Radiography Utilization Study (NEXUS). METHODS: This was a prospective, observational study of blunt trauma patients in 21 U.S. medical centers undergoing plain cervical radiography. SCIWORA was defined as spinal cord injury demonstrated by magnetic resonance imaging, when a complete, technically adequate plain radiographic series revealed no injury. RESULTS: Of the 34,069 patients entered, there were 818 (2.4%) with cervical spine injury, including 27 (0.08%) patients with SCIWORA. Over 3,000 children were enrolled, including 30 with cervical spine injury, but none had SCIWORA. The most common magnetic resonance imaging findings among SCIWORA patients were central disc herniation, spinal stenosis, and cord edema or contusion. Central cord syndrome was described in 10 cases. CONCLUSION: In the large NEXUS cohort, SCIWORA was an uncommon disorder, and occurred only in adults. PMID- 12131381 TI - Spinal injuries in motorcycle crashes: patterns and outcomes. AB - BACKGROUND: The purpose of this study was to determine patterns of spinal injury and clinical outcomes resulting from motorcycle crashes. METHODS: We analyzed data collected on 1,121 motorcyclists involved in road traffic accidents (from 1993-2000) and identified those who had sustained a spinal injury. RESULTS: Spinal injury occurred in 126 (11.2%) riders (112 male riders [88.9%] and 14 female riders [11.1%]), with a mean age of 30.2 years (range, 16-61 years) and Injury Severity Score of 18.8 (range, 4-66). Isolated injuries to the spine occurred in 30 (23.8%) riders. The thoracic spine was injured in 69 (54.8%), the lumbar spine in 37 (29.4%), and the cervical spine in 34 (27.0%) cases. Multiple vertebral levels were affected in 54 (42.9%). Neurologic injury occurred in 25 riders (19.8%), with complete distal neurologic injury in 14 (4 cervical, 9 thoracic, and 1 lumbar). Eleven (8.7%) patients required spinal surgery. There were 13 (10.3%) deaths. CONCLUSION: The thoracic spine is the most commonly injured spinal region in motorcycle crashes. Multiple level injuries are common. Protocols concentrating on the radiographic clearance of the cervical region may miss a significant number of spinal injuries. Vigilance is required in assessing these patients, who often have multiple injuries. PMID- 12131382 TI - Evaluation of the lower spine after blunt trauma using abdominal computed tomographic scanning supplemented with lateral scanograms. AB - BACKGROUND: Patients at risk for thoracolumbar junction (TLJ) and lumbar spine (LS) injury after blunt trauma are classically evaluated using conventional radiographs. Frequently, these patients also undergo abdominal and pelvic computed tomographic (CT) scanning to exclude the presence of associated intra abdominal injuries. Standard abdominal and pelvic CT scan usually includes an anteroposterior (AP) scout film (scanogram) obtained before the cross-sectional imaging. The objective of this study was to determine whether a lateral CT scanogram and axial CT views would provide adequate imaging to allow for evaluation of the TLJ and LS and therefore eliminate the need for conventional screening computed lumbar spine radiographs (CLSRs). METHODS: Patients who sustained blunt injury and required both CLSRs as well as abdominal and pelvic CT scans were prospectively identified. The study protocol (CT + S) added lateral CT scanograms to all helical abdominal and pelvic CT scan studies. The AP and lateral CT scanograms were included with the axial images, and these views were reviewed together during final radiographic interpretation and diagnosis. The results of CT + S were compared with readings of the CLSRs (AP and lateral) in a blinded fashion by a trauma radiologist. RESULTS: Lateral scanograms were generated for 71 patients. All scanograms were technically adequate, with image quality equal or superior to computed plain radiographs. Ten patients were found to have 20 fractures, 19 acute and 1 chronic. All abnormalities identified by plain radiographs were seen using CT + S (sensitivity, 100%; specificity, 100%). Eight transverse process and two spinous process fractures not seen on CLSRs were identified using CT + S. CONCLUSION: Our CT + S protocol (axial CT images plus AP and lateral scanograms) outperformed screening CLSRs in the detection of fractures of the lower spine (TLJ + LS) after blunt trauma. In addition, scanogram imaging is less dependent on body habitus and adds no additional cost or time to abdominal and pelvic CT scanning. Further study is required to determine whether CT + S can routinely replace conventional radiographs of the lower spine after blunt trauma. PMID- 12131383 TI - Pelvic fracture in geriatric patients: a distinct clinical entity. AB - BACKGROUND: The purpose of this study was to describe differences in demographics, injury pattern, transfusion needs, and outcome of pelvic fractures in older versus younger patients. METHODS: This was a retrospective registry review of all patients with pelvic fractures admitted directly from the scene between January 1998 and December 1999. RESULTS: We cared for 234 patients with pelvic fractures during the study period. Mean age was 37.2 years, 51% were men, and mean Injury Severity Score (ISS) was 19. Overall mortality was 9%. Eighty three percent were under the age of 55 years and 17% were older than 55 years. Severe pelvic fractures (AP3, LC3) were more common in young patients (p < 0.05). Admitting systolic blood pressure was lower and heart rate higher, although ISS was not different between the two age groups. Older patients were 2.8 times as likely to undergo transfusion (p < 0.005), and those undergoing transfusion required more blood (median, 7.5 units vs. 5 units). Older patients underwent angiography more frequently and were significantly more likely to die in the hospital even after adjusting for ISS (p < 0.005). This was most marked with ISS 15 to 25. Lateral compression (LC) fractures occurred 4.6 times more frequently in older patients than anteroposterior (AP) compression, and 8.2 times more frequently in those older patients undergoing transfusion as compared with AP compression. Ninety-eight percent of LC fractures in older patients were minor (LC1,2). However, older patients with LC fractures were nearly four times as likely to require blood compared with younger patients. CONCLUSION: In older patients, pelvic fractures are more likely to produce hemorrhage and require angiography. Fracture patterns differ in older patients, with LC fractures occurring more frequently, and commonly causing significant blood loss. The outcome of older patients with pelvic fractures is significantly worse than younger patients, particularly with higher injury severity. Recognition of these differences should help clinicians to identify patients at high risk for bleeding and death early, and to refine diagnostic and resuscitation strategies. PMID- 12131384 TI - Turning point: rethinking violence--evaluation of program efficacy in reducing adolescent violent crime recidivism. AB - BACKGROUND: The link between medicine and violence prevention is self-evident, yet the literature reveals few studies that scientifically document effective interventions. The Turning Point: Rethinking Violence (TPRV) program is a unique multiagency program developed to expose, educate, and remediate first-time violent offenders and their parents regarding the real-world consequences of violence. Four key components are integrated into a 6-week, court-ordered program (14 total contact hours): the Trauma Experience (tour, video, discussions), the Victim Impact Panel (presented by parent survivors), Group Process, and Community Networking (individualized mental health referral). We hypothesize that TPRV delivers lower outcomes regarding violence recidivism (VR) when compared with standard court sentencing options (100 hours of community service) for first-time violent offenders. METHODS: The study group subjects (n = 38) met inclusion criteria and were blindly and randomly sentenced to attend the TPRV program. The control group (n = 38) were pulled from a subject pool of first-time offenders who received standard sentencing options, met the same inclusion criteria with regard to age and types of offenses, and were matched by race to the study group. Both groups were studied for VR within the year after the first violent conviction, and comparison was performed by a (2 analysis of recidivism rates. RESULTS: Results reveal a statistically significant difference between the study group and the control group for VR (p 3) from January 1, 2000, through December 31, 2000. Two groups were formed on the basis of the timing of UFH administration: within 72 hours of admission (Early group), or after the third day of hospitalization (Late group), if at all. Intracranial bleeding related to UFH administration was assessed by computed tomographic scan of the head and/or clinical examination. RESULTS: Sixty-four of 76 patients with intracranial blood on admission head computed tomographic scan fulfilled study criteria. Seventy-three percent (n = 47) were in the Early group and 27% (n = 17) were in the Late group. None of the Early group had an increase in intracranial bleeding or deterioration on neurologic examination as a result of UFH administration. However, there was no statistical difference in VTE events between the two groups. CONCLUSION: Early use of UFH in the severe head injured patient does not increase bleeding complications. PMID- 12131389 TI - Pediatric and adolescent tibial eminence fractures: arthroscopic cannulated screw fixation. AB - BACKGROUND: Fractures of the intercondylar spine of the tibia are enigmatic injuries. The mechanism of injury remains obscure, and appropriate treatment is unclear. METHODS: The authors analyzed a series of 26 cases of displaced fractures of the intercondylar eminence of the tibia treated with an arthroscopically placed, intrafocal screw with spiked washer. The patients were reviewed after a minimum follow-up of 24 months and a maximum of 8 years. RESULTS: Sixteen patients had a type II tibia eminence fracture according to Meyers and McKeever (mean age, 15 years; male/female ratio, 11:5). Ten patients had a type III tibia eminence fracture (mean age, 17 years; male/female ratio, 1:1). We encountered neither stiffness nor iatrogenic chondral abrasion. Only three patients with type II had no laxity. The 13 other patients in this fracture group had a minor laxity without correlation with the clinical result. In four patients with a type III lesion, a residual laxity without functional deficit was noticed. In two cases with a type III lesion, a reconstruction of the anterior cruciate ligament was necessary 3 years after trauma. In four patients with a type III fracture, the fragment remained elevated, with minor impairment of the mobility (extension lag). No mechanical failure or infection was seen in this series. CONCLUSION: The authors found the intrafocal screw fixation for displaced fracture of the intercondylar eminence to be a reliable and safe technique, although complete restoration of the anteroposterior knee stability was seldom seen. PMID- 12131388 TI - Fluoroscopic positioning of sacroiliac screws in 88 patients. AB - BACKGROUND: Fluoroscopic placement of guided sacroiliac screws is a well established method of fixation of the posterior pelvic ring, leading to biomechanical results similar to an intact pelvic ring. The main problem remains the risk of neurologic injury resulting from the penetration of the intervertebral root or the vertebral canal. METHODS: Eighty-eight patients in whom the posterior pelvic ring was stabilized for several indications were reviewed retrospectively. On perioperative and direct postoperative radiographs and postoperative computed tomographic (CT) scans, positioning was scored for 285 screws and compared with clinical results. RESULTS: Depending on the type of imaging (radiography or CT scan), only 2.1% to 6.8% of the screws showed malpositioning. In several cases, the malpositioned screws did not cause any complaints. Postoperative radiographs did not show any additional value above perioperative radiographs in predicting malpositioning. Seven of 88 patients had neurologic complaints and underwent reoperation. All complaints resolved completely, and no permanent neurologic damage occurred. Positioning both sacroiliac screws in the first vertebral body had a significantly lower rate of neurologic complaints compared with the lower screw in the second vertebral body. CT scanning was able to predict neurologic complaints most accurately. CONCLUSION: Percutaneous sacroiliac screws can be positioned safely, in experienced hands, using perioperative fluoroscopic techniques. A position in the first vertebral body had a significantly lower incidence of neurologic injury compared with a position in the second. In case of postoperative neurologic deficit, only CT scan can predict the clinical outcome. Further research toward improving the perioperative imaging technique must be undertaken. PMID- 12131390 TI - Results of ankle fractures with involvement of the posterior tibial margin. AB - BACKGROUND: Ankle fractures have a significantly worse functional outcome when they include a posterior tibial fragment. In 57 trimalleolar fractures, the effect of size, internal fixation, and anatomic reduction of the posterior fragment on the prognosis was evaluated. METHODS: A modified Weber protocol was used, providing a rating system for subjective, objective, and radiographic results. A visual analogue scale for subjective actual pain was also scored. RESULTS: The involvement of the articular surface ranged from 8% to 55%. Size or fixation of the fragment did not influence prognosis. Joint congruity in fragments >or= 10% of the articular surface was a significant factor influencing prognosis. Overall, the modified Weber protocol result was excellent in 10%, good in 15%, fair in 25%, and poor in 50% of patients. However, the low average visual analogue scale of 3.0 in the whole group does not appear representative of 50% poor results, indicating that the modified Weber protocol is fairly strict and overestimates the number of poor results. CONCLUSION: Joint congruity with or without fixation was a significant factor influencing prognosis. Congruity should be achieved for fragments >or= 10% of the tibial articular surface. PMID- 12131391 TI - The utility of both muscle and fascia flaps in severe upper extremity trauma. AB - BACKGROUND: Severe isolated upper extremity injuries are rarely lethal; however, they invariably are resource intensive, create significant disability, and promote resistance to a return to gainful employment. Appropriate soft tissue restoration is an essential component of any treatment protocol, and often requires a vascularized flap to protect the superficial neurovascular and musculotendinous structures. A basic schema to facilitate flap selection in the upper extremity is introduced. METHODS: The role of local muscle and fascia flaps or free tissue transfers for severe upper extremity injuries was retrospectively reviewed from a two-decade experience. Excluding digital injuries, primary treatment of soft tissue traumatic wounds requiring some form of vascularized flap occurred in 33 limbs in 31 patients. The choice of flap donor site, type, specific complications and benefits as related to the severity of injury, and the effect of timing of wound closure were compared. RESULTS: Initial coverage after significant upper extremity trauma in these 33 limbs required 16 local fascia flaps, 22 free flaps, 1 multistaged distant pedicled flap, and 1 local muscle flap. Flaps were selected in a nonrandom fashion on the basis of wound location, severity of injury, and flap availability. Complication rates were similar for local fascia and free flaps. The upper extremity could be divided into three regions that were differentiated according to the observed incidence of flap preference. Free flaps were more commonly used for hand and wrist wounds, or anywhere the defect was moderately large in size or extremely severe in overall injury. Local fascia flaps were a simpler option most applicable for the central upper limb. Local muscles as flaps were intentionally avoided to minimize any functional derangement. CONCLUSION: A schema to guide flap selection for upper extremity coverage is introduced that is predicated on using the best available option. The shoulder girdle and axilla are reached by many local trunk muscle or fascia flaps. The central upper limb about the elbow often is conducive to coverage with specific local fascia flaps. The distal upper extremity may be best served by a free flap, as would any large wound in all upper limb regions. PMID- 12131392 TI - Moderate hypothermia prevents brain stem oxidative stress injury after hemorrhagic shock. AB - BACKGROUND: The purpose of this study was to investigate the effects of temperature on oxidative stress in brain stem tissue induced by hemorrhagic shock. We researched the hemorrhagic oxidative stress at various core temperatures using reduced glutathione (GSH) levels and thiobarbituric acid reactive substances (TBARS) as markers of lipid peroxidation in brain stem homogenate. METHODS: Forty rats were divided into four groups, of which one constituted the nonbleeding normothermia control group. In all of the three study groups, 40% of estimated blood volume was removed while they were being held at normothermia, mild hypothermia (32 degrees C), or moderate hypothermia (28 degrees C). Parameters including mean arterial pressure, rectal temperature, and heart and breathing rates were monitored and recorded during the procedures. After an hour at shock state, tissue samples were removed by craniectomy. RESULTS: The tissue levels of TBARS increased significantly in normothermic and mild hypothermic hemorrhagic shock groups (10.74 nmol/g and 8.26 nmol/g) as compared with the control group (3.50 nmol/g) (p < 0.001). However, the tissue TBARS level in the moderate hypothermia group was only minimally increased (4.53 nmol/g). GSH showed a slight decrease in normothermic and mild hypothermic bleeding rats, and were unchanged in the moderate hypothermic rats. CONCLUSION: Moderate systemic hypothermia (28 degrees C) appears to protect brain stem tissue from oxidative stress during severe hemorrhagic shock in rats, as indicated by insignificant change in tissue TBARS and GSH concentrations. These results suggest antioxidant protective effects of moderate systemic hypothermia in metabolically active brain stem tissue during hemorrhagic shock. Similar effects in humans remain to be studied. PMID- 12131393 TI - Pterional orbita decompression in orbital hemorrhage and trauma. AB - BACKGROUND: This article presents a series of patients with traumatic retrobulbar hematoma and orbital trauma, treated with extended pterional orbital decompression. METHODS: Fifteen patients, showing symptomatic retrobulbar hematoma or symptoms of orbital injury after various trauma mechanisms, were treated with deep lateral orbital decompression and removal of orbital blood/bone fragments via this approach. Preoperative and postoperative course, neuroradiologic findings, additional brain or facial injuries, and outcome of eye function are analyzed in detail. RESULTS: Mean delay between trauma and decompression was 70 hours (3 days), with a range from 2 hours to 15 days. Proptosis decreased in all patients and visual acuity improved or remained normal in nine patients and stayed defective in four. Impaired extraocular movements and pupillary changes recovered in 10 patients. Apart from one case of permanent deficit of the frontal branch of the facial nerve, no severe complications were seen. CONCLUSION: The presented pterional orbital decompression represents an effective alternative approach for patients with sight-threatening retrobulbar hematoma or orbital trauma, especially in cases that require direct access to damaged structures and maximal decompression of the orbit. Immediate detection and treatment of orbital hematomas is mandatory for acceptable outcome of eye function. PMID- 12131394 TI - Isolated free fluid on computed tomographic scan in blunt abdominal trauma: a systematic review of incidence and management. AB - BACKGROUND: Abdominal computed tomographic (CT) scan is accepted as the primary diagnostic modality in stable patients with blunt abdominal trauma. A recent survey of 328 trauma surgeons demonstrated marked variation in the management of patients with head injuries and the finding of free intra-abdominal fluid without solid organ injury on CT scan. This study was undertaken to attempt to determine what to do when free fluid without solid organ injury is seen on abdominal CT scan in patients with blunt trauma. METHODS: Articles concerning the incidence and significance of free intra-abdominal fluid on CT scan of blunt trauma patients without solid organ injury were systematically reviewed. A MEDLINE search was performed using terms such as tomography-x-ray computed, wounds nonpenetrating, small intestine/injuries, time factors, and abdominal trauma and diagnostic tests. Bibliographies of pertinent articles were reviewed. Appropriate articles were evaluated for quality and data were combined to reach a conclusion. RESULTS: Meta-analysis could not be performed because no randomized, prospective, controlled trials could be found. Forty-one articles were excluded from the analysis because they looked at only patients with known injuries to intestine, diaphragm, or pancreas and the investigation of the CT scan findings did not include negative scans. Ten articles, which described CT scan results for all patients presenting with blunt abdominal trauma for a defined period of time, formed the basis of this study. Isolated free fluid was seen in 463 (2.8%) of over 16,000 blunt trauma patients scanned. A therapeutic laparotomy was performed in only 122 (27%) of these patients. CONCLUSION: The isolated finding of free intra-abdominal fluid on CT scan in patients with blunt trauma and no solid organ injury does not warrant laparotomy. Alert patients may be followed with physical examination. Patients with altered mental status should undergo diagnostic peritoneal lavage. PMID- 12131395 TI - Incidence, outcome, and long-term consequences of herpes simplex virus type 1 reactivation presenting as a facial rash in intubated adult burn patients treated with acyclovir. AB - BACKGROUND: Increased mortality, extensive visceral involvement, and necrotizing tracheobronchitis associated with herpes viruses have been reported after burns. It is unclear whether herpes presenting as a facial rash results in outcome changes after burns. METHODS: A retrospective study characterizing the incidence, presentation, and outcome of 14 patients with facial herpes rashes out of 95 severely burned intubated adults was performed. RESULTS: Facial rashes attributed to herpetic infections were found in at least 15% of patients. The problem was recognized during the second week after burn. There was no difference in mortality or length of stay noted between patients with or without the infection. CONCLUSION: The course of this infection was relatively benign in this group of acyclovir-treated patients. Even so, the lesions clearly contributed to patient discomfort and often produced fevers requiring costly investigations. Early recognition could help prevent diffuse spread of the lesions, decreasing patient discomfort and improving patient care. PMID- 12131396 TI - Improvements in prehospital trauma care in an African country with no formal emergency medical services. AB - BACKGROUND: A large proportion of trauma patients in developing countries do not have access to formal Emergency Medical Services. We sought to assess the efficacy of a program that builds on the existing, although informal, system of prehospital transport in Ghana. In that country, the majority of injured persons are transported to the hospital by some type of commercial vehicle, such as a taxi or bus. METHODS: A total of 335 commercial drivers were trained using a 6 hour basic first aid course. The efficacy of this course was assessed by comparing the process of prehospital trauma care provided before versus after the course, as determined by self-report from the drivers. RESULTS: Follow-up interviews were conducted on 71 of the drivers a mean of 10.6 months after the course. Sixty-one percent indicated that they had provided first aid since taking the course. There was considerable improvement in the provision of the components of first aid in comparison to what was reported before the course: crash scene management (7% before vs. 35% after), airway management (2% vs. 35%), external bleeding control (4% vs. 42%), and splinting of injured extremities (1 vs. 16%). CONCLUSION: Even in the absence of formal Emergency Medical Services, improvements in the process of prehospital trauma care are possible by building on existing, although informal, patterns of prehospital transport. PMID- 12131397 TI - Outcomes after injury: a comparison of workplace and nonworkplace injury. AB - BACKGROUND: Factors affecting recovery from injury are investigated comparing male emergency department patients involved in work-related and non-work-related accidents. METHODS: This was a prospective cohort study of 154 injured employed male emergency department patients recording demographic and accident details, return to work information, and involvement in litigation. Standardized questionnaires measured psychological, physical, and social responses. Evaluations were at admission, and at 6 weeks, 6 months, and 18 months after injury. RESULTS: Work-related injuries were less severe than non-work-related injuries (p = 0.006), and more patients became involved in litigation (p = 0.02) and suffered symptoms of posttraumatic stress disorder (p = 0.04). Psychosocial symptoms increased with nonreturn to work (p < 0.05). Factors predicting return to work include injury severity, blaming others, involvement in litigation, and subsequent physical and social functioning. CONCLUSION: Patients injured at work are more likely to commence litigation and develop symptoms consistent with posttraumatic stress disorder. Nonreturn to work is associated with higher psychosocial morbidity. Return to work is predicted from event and recovery period variables. PMID- 12131398 TI - Delayed cardiac tamponade complicating airbag deployment. PMID- 12131399 TI - Aortoduodenal fistula in the acute trauma setting: case report. PMID- 12131400 TI - Critical cerebral ischemia revealed by magnetic resonance imaging in a traumatic carotid-cavernous fistula without high-risk patterns on angiograms: a case report. PMID- 12131401 TI - Approach for drainage of descending necrotizing mediastinitis on the basis of the extending progression from deep neck infection to mediastinitis. PMID- 12131402 TI - Novel uses of a negative-pressure wound care system. PMID- 12131403 TI - Idiopathic compartment syndrome: a case report. PMID- 12131404 TI - Synchronous fractures of the trochlea and the radial neck without elbow dislocation. PMID- 12131405 TI - Irreducible lateral patellar dislocation with vertical axis rotation: case report and review of the literature. PMID- 12131406 TI - Successful reimplantation of retrieved large segment of open femoral fracture: case report. PMID- 12131407 TI - Blindness due to anterior ischemic optic neuropathy in a burn patient. PMID- 12131409 TI - Practice management guidelines for the prevention of venous thromboembolism in trauma patients: the EAST practice management guidelines work group. PMID- 12131410 TI - Missed cervical spinal cord injuries. PMID- 12131411 TI - Recurrent penetrating spinal cord injury. PMID- 12131412 TI - Placental abruption. PMID- 12131413 TI - Off-pump extraction of an embedded high posterior left ventricular bullet utilizing a new cardiac stabilization device. PMID- 12131414 TI - Thoracic handlebar hernia: a rare etiologic variant of traumatic intercostal hernia. PMID- 12131416 TI - Effects of lifestyle and work-related physical activity on the degree of lumbar lordosis and chronic low back pain in a Middle East population. PMID- 12131417 TI - The influence of lumbar lordosis on spinal fusion and functional outcome after posterolateral spinal fusion with and without pedicle screw instrumentation. AB - The aim of the current study was to examine the correlation between lumbar lordosis, spinal fusion, and functional outcome in patients suffering from severe low back pain, treated by posterolateral spinal fusion with or without pedicle screw instrumentation. One hundred thirty patients were randomly allocated to posterolateral lumbar fusion with or without Cotrel-Dubousset instrumentation. Functional outcome was assessed preoperatively, and 1 and 2 years postoperatively. Lordosis angles of the lumbar spine and fusion rates were assessed at the 1- and 2-year follow-up. No difference in lordosis angle was found between the two groups at any time. Lordosis was unchanged at 2 years compared with preoperative status in both groups. In the instrumented group, nonunion (23%) was followed by a decrease in lordosis at follow-up (p < 0.05). However, in the noninstrumented group, nonunion (14%) resulted in increased lordosis (p < 0.05). No correlation was found between functional outcome and lordosis angle. The current study showed no correlation between functional outcome and lordosis angle either before or after posterolateral spinal fusion. Use of instrumentation did not influence lumbar spinal alignment compared with noninstrumented fusions. The sagittal alignment was stable both 1 and 2 years after solid fusion. The failure mode of instrumented fusions was a reduced degree of lordosis in contrast to an increased degree of lordosis in patients with noninstrumented fusion. PMID- 12131418 TI - Diagnosis of symptomatic disc by magnetic resonance imaging: T2-weighted and gadolinium-DTPA-enhanced T1-weighted magnetic resonance imaging. AB - Although radial tear of the annulus fibrosus can be detected on T2-weighted and Gd-DTPA-enhanced magnetic resonance (MR) images, the association between the annular tear on MR images and the symptomatic discs is unclear. The purpose of this study was to investigate the relationship between T2-weighted, gadolinium DTPA-enhanced MR images and pain response through discography in patients with chronic low back pain. A total of 56 lumbar discs from 23 patients with chronic low back pain (13 to 47 years old) underwent MR imaging (T2-weighted, gadolinium DTPA-enhanced MR images) followed by provocative discography. The sensitivity, specificity, positive predictive value, and negative predictive value of T2 weighted and gadolinium-DTPA-enhanced MR images in detecting the symptomatic discs were calculated. The sensitivity, specificity, positive predictive value, and negative predictive value of T2-weighted images in detecting the symptomatic disc were 94%, 71%, 59%, and 97%, respectively. The sensitivity, specificity, positive predictive value, and negative predictive value of gadolinium-DTPA enhanced images were 71%, 75%, 56%, and 86%, respectively. The high sensitivity and the high negative predictive value of T2-weighted MR imaging in detecting the symptomatic disc indicated that MR imaging can be a useful screening tool in avoiding unnecessary discography in patients with chronic low back pain. PMID- 12131419 TI - Anatomic and radiographic considerations for placement of transiliac screws in lumbopelvic fixations. AB - Lumbopelvic fixation in spinal and pelvic surgery relies on rods or screws as an iliac anchor. Secure placement of screws with maximum diameter and length for the greatest pullout strength requires knowledge of the iliac structure and of intraoperative fluoroscopic landmarks for secure placement. Therefore, the authors evaluated the intrailiac length, inner width, and cortical thickness of three different transiliac screw anchor paths aimed toward the anterior inferior iliac spine and initiated at the iliac tubercle, posterior superior iliac spine, or posterior inferior iliac spine. Measurements were made using two- and three dimensional computed tomographic reformations in 40 consecutive trauma patients (27 measurements in 21 males, 16 to 75 years old; 28 measurements in 19 females, 16 to 78 years old). In addition, fresh and dry human cadaveric specimens were marked with metal wires at the previously determined optimal screw path to determine fluoroscopic landmarks for easiest and best controlled transiliac screw placement. The posterior superior iliac spine-anterior inferior iliac spine path had the largest bony canal lengths, with 141 mm in male and 129 mm in female patients. Two stereotypic iliac constrictions allowed placement of 8-mm implants in male and 6- to 7-mm implants in female patients. Cortical thickness at that optimal extraarticular path was 5.2 mm in the male and 4.7 mm in the female patients. Transiliac screws can be placed during operation under fluoroscopic control using standard lateral and obturator oblique-outlet views, the latter presenting a stereotypical teardrop figure above the acetabulum. PMID- 12131421 TI - Spinal cage retrieval and assessment of biologic response. AB - Implant retrieval programs have been effective in understanding implant failure and biomaterial compatibility in joint arthroplasty; however, its application has not been extended extensively to the assessment of spinal constructs and implants. The objective of this study is to determine the efficacy of implant retrieval analysis as a standard for the assessment of explanted spinal implants. The limitations of clinical radiographic assessment of fusion through metal interbody devices are also identified. The implant analysis protocol is shown through a case report of a titanium mesh spinal fusion cage retrieved from a 54 year-old woman who had a pseudoarthrosis at the T12 cage interface. The implant analysis techniques include backscattered electron imaging, high-resolution contact radiography, histology, and fluorochrome analysis. An implant retrieval analysis program similar to the one discussed in the presented case study will enable an accurate assessment of outcomes of these commonly used implants and will guide future development. PMID- 12131422 TI - Can the thoracic kyphosis be modeled with a simple geometric shape? The results of circular and elliptical modeling in 80 asymptomatic patients. AB - Many Cobb measurements have been reported at various levels for the thoracic kyphosis, but geometric models of the shape of kyphosis are rare. Thoracic vertebral bodies were digitized on 80 normal lateral full-spine radiographs to obtain the mean thoracic kyphosis. Global and segmental angles were determined. Computer iteration processes passed geometric shapes through the posterior body coordinates of the mean thoracic kyphosis to determine the best fit model in the least squares sense. The kyphosis was closely modeled with ellipses. The T1 and T12 areas tended to be flatter in curvature when compared with T2-T11, indicating these are inflection points. Mean global angles were Cobb(T1-T12) = 44.2 degrees, Cobb(T2-T11) = 39.9 degrees, and Cobb(T3-T10) = 33.3 degrees. The T2-T11 kyphotic region was closely modeled with approximately a 70-degree portion of an ellipse, with minor axis to major axis ratios of 0.6 to 0.72, and with major axis parallel to the posterior body margin of T11. PMID- 12131423 TI - Anterior dynamic plates in complex cervical reconstructive surgeries. AB - The aim of this work was to evaluate the efficacy of dynamic anterior cervical plates following one-level anterior corpectomy with fusion (ACF) (i.e., C5-7) and two- to four-level ACF with posterior wiring and fusion (PWF). Dynamic plates (ABC Aesculap, Tuttlingen, Germany), allowing for 10 mm of cephalad and 10 mm of caudad plate migration, were applied for one-level ACF (28 patients) and multilevel ACF/PWF (20 patients). Two (7%) dynamic plates failed after one-level ACF. One pseudarthrosis and one plate extrusion occurred, both in morbidly obese patients. No multilevel ACF/PWF failed. The 7% failure rate for dynamic plates applied for one-level ACF is somewhat high, whereas the 0% failure rate for multilevel ACF/PWF with halo immobilization proves more promising. Perhaps PWF with halo stabilization for morbidly obese individuals undergoing one-level ACF would avoid future plate-related problems. PMID- 12131425 TI - Simultaneous cervical and lumbar surgery for combined symptomatic cervical and lumbar spinal stenoses. AB - Spinal stenosis may rarely involve both cervical and lumbar spines. An alternative surgical strategy used for the treatment of combined cervical and lumbar spinal stenosis is presented. Two cases with symptomatic combined stenosis of the cervical and lumbar spinal canal are described. Simultaneous surgery was performed in both cases. The combined stenosis of the cervical and lumbar spinal canal dictates careful neurologic and neuroradiologic examinations. Simultaneous surgery is an alternative approach for patients with symptomatic multilevel spinal stenoses, whose general conditions necessitate a one-session and short lasting surgery. PMID- 12131426 TI - Effects of polyaxial pedicle screws on lumbar construct rigidity. AB - Pedicle screw constructs have been shown to increase fusion rates in the lumbar spine. Manufacturers have created pedicle screws with one or two degrees of freedom built into the screw head to allow for easier incorporation of the interlocking rod, but the effects of these screws on construct stiffness has not been tested. The purpose of this study is to compare and contrast the stiffness of lumbar pedicle screw constructs with and without the use of polyaxial pedicle screws. Nontapered, self-taping pedicle screws (6.0-mm diameter x 30-mm length, titanium) were used in the fixation of porcine spines from L3-L5. Group 1 (n = 5) contained six standard pedicle screws from one manufacturer. Group 2 (n = 5) contained six standard pedicle screws from a second manufacturer. Group 3 (n = 5) contained four standard pedicle screws placed at L3 and L5, as well as two polyaxial screws placed at L4. Group 4 (n = 5) contained six polyaxial pedicle screws. A rotational variable differential transformer was used to record angular displacement between vertebrae in the construct as it is loaded in flexion, extension, right bend, left bend, clockwise torque, and counterclockwise torque. Stiffness curves were linear throughout the range of applied force. The average r2 value for the generated stiffness graphs was 0.94 (SD = 0.06). No construct failure occurred during any of the testing. There were no significant differences (p < 0.05, two-way analysis of variance) in moment versus angle noted in any of the four groups tested. For torque tests, the all-polyaxial screw constructs showed significantly increased stiffness compared with the other groups. The current study has shown that the incorporation of polyaxial screws in pedicle screw constructs did not significantly decrease the construct stiffness. There is a suggestion that the use of all polyaxial screws may increase the resistance to torque by allowing better purchase of intervertebral rods. PMID- 12131427 TI - Hydroxyapatite granule graft combined with recombinant human bone morphogenic protein-2 for solid lumbar fusion. AB - The purpose of this study was to evaluate the availability of recombinant human bone morphogenetic protein-2 (rhBMP-2) combined with hydroxyapatite (HA) and autogenous bone. Posterolateral intertransverse fusion between the fifth and sixth lumbar vertebrae was performed in 27 adult Japanese white rabbits. These 27 rabbits were classified into three groups: the autogenous bone group, the HA group, and the bone morphogenic protein (BMP) group. In the HA group, HA (0.5 g) mixed with iliac bone was grafted. In the BMP group, HA (0.5 g) soaked with rhBMP 2 (100 mg) and iliac bone was grafted. At 6 weeks after the procedure, bone union was evaluated. In the BMP group, all cases showed solid bone union, and fusion masses were stiffer than the masses obtained in the other group. Biomechanically and histologically, grafts of HA soaked with rhBMP-2 and iliac bone was clearly effective in obtaining a solid intertransverse arthrodesis. PMID- 12131428 TI - Does provocative discography screening of discogenic back pain improve surgical outcome? AB - The value of preoperative provocative discography in the setting of discogenic low back pain was investigated by evaluating surgical outcomes. Seventy-three consecutive patients who underwent posterolateral interbody and posterior spinal arthrodesis for discogenic low back pain refractory to nonoperative management were reviewed. Chronologically, the first 41 patients (group A) were indicated without discography, whereas the remaining 32 (group B) had been indicated only if their pain had been reproduced during disc injection. The two groups were similar in demographic, psychometric, and radiologic parameters. Average follow up time in group A was 2.8 years and in group B it was 2.4 years, both with a 2 year minimum. Using modified Oswestry scoring, group A and group B patients had satisfactory outcomes of 75.6% and 81.2%, respectively. This difference was neither statistically significant nor suggestive. In this study, provocative discography screening did not improve surgical outcomes after circumferential fusion for lumbar discogenic back pain. PMID- 12131430 TI - Endodermal cyst of the cervical spine treated by an anterior approach for resection and shunting. AB - In most reported cases, a posterior approach to the cervical spine was used to remove an endodermal cyst or to place a shunt for cyst drainage. The authors performed partial excision, shunt insertion, and anterior spinal fusion in a patient with endodermal cyst of the cervical spine. Symptoms resolved after surgery and magnetic resonance images indicated morphologic restoration of the spinal cord. PMID- 12131429 TI - Vertebra plana caused by primary Ewing sarcoma: case report and review of the literature. AB - A 7-year-old boy presented with vertebra plana of T11. The presumptive diagnosis suggested by clinical presentation, conventional radiographs, and computed tomographic scans was eosinophilic granuloma. Progressive neurologic symptoms required surgical excision of the lesion and decompression. Histopathologic examination of the surgical specimen confirmed the diagnosis of Ewing sarcoma. PMID- 12131431 TI - Scoliosis in a patient with Alexander disease. AB - Alexander disease is a rare, degenerative disorder of the central nervous system. It is characterized clinically by spasticity, seizures, dementia, loss of developmental milestones, and macrocephaly. Here we describe a 13-year-old boy with Alexander disease and severe scoliosis. The patient initially presented at 9 months of age, with profound mental retardation and a history of seizures. When he was 7 years old, a pediatrician had diagnosed Alexander disease (hypotonia, macrocephaly, and progressive low-density white matter predominantly in the frontal region on computed tomography examination). From the age of 10, thoracolumbar scoliosis had gradually become severe. Because treatment using a corrective brace would have produced major problems because of the patient's mental retardation, the scoliosis was successfully treated surgically, by careful posterior spinal fusion with instrumentation, and an autologous iliac crest bone graft. A 64 degrees curve was corrected to 18 degrees (72% correction). Scoliosis with Alexander disease is considered to be very rare because patients with the disease seldom survive long enough to develop spinal deformities. PMID- 12131432 TI - Sacrococcygeal ganglioneuroma. AB - Ganglioneuromas are benign slow-growing masses that can be treated with complete surgical extirpation without any adjuvant therapy. Such lesions involving the sacrococcygeal region are exceedingly rare. The authors present the case of a 70 year-old woman with a sacrococcygeal ganglioneuroma treated by total en bloc resection. This patient also had a previous coccygeal fracture. To the authors' knowledge, there are no other reports of ganglioneuroma in association with a history of trauma. PMID- 12131433 TI - Treatment of rigid hypertrophic posttraumatic pseudarthrosis of the tibia in children using distraction osteogenesis. AB - Posttraumatic tibial pseudarthrosis is a relatively uncommon complication of tibia fractures in children. Although reported as a successful method of treating tibial nonunions in adults, distraction osteogenesis through a nonunion site via ring external fixation has not been described in children. The authors report three consecutive cases of distraction through an angulated, shortened, hypertrophic, posttraumatic nonunion to achieve successful union and concurrent correction of deformity. Distraction was carried out using a ring fixator with computer-guided correction. Union was achieved in each of the three patients with complete correction of deformity and length. Lengthening of 8 to 31 mm was achieved. The length of time in the external fixator ranged from 7 to 27 weeks. Pin tract infections developed in all patients; they were treated successfully using oral antibiotics. No deep infections or other significant complications developed. Each patient was followed up for at least 1 year. PMID- 12131434 TI - Mechanical properties of different combinations of flexible nails in a model of a pediatric femoral fracture. AB - Flexible nailing of pediatric femoral shaft fractures is based on the principle of using two C-shaped nails to create six points of fixation. However, clinical studies using various nail combinations demonstrate similar outcomes. This study aimed to compare the mechanical properties of different nail combinations by testing them in a model of a child's midshaft femoral fracture. The two C-shaped nails were compared with two straight nails and with paired S- and C-shaped nails. The constructs were tested in four-point bending and torsion. Graphs of the data were produced, from which the bending and torsional stiffness of the constructs was calculated. The results showed that there was no significant difference between the mechanical properties of the three different constructs. The conclusion is that any of the tested nail combinations can be used to treat a midshaft fracture of the femur in a child. PMID- 12131435 TI - Comparison of dynamic versus static external fixation for pediatric femur fractures. AB - External fixation of pediatric femoral shaft fractures has the advantages of minimal dissection and early weight bearing. However, it is associated with slow healing and potential for refracture. Some surgeons have proposed that axial dynamization may improve the speed and strength of callus formation. to test this hypothesis, we performed a randomized controlled trial using 53 femur fractures in 52 patients between 1995 and 1999. Patients were randomized to receive dynamic or static fixation. Average time until early callus formation was 23.2 days for dynamic fixation and 24.9 days for static fixation (P = 0.627). Average time until complete radiographic healing was 70.1 days for dynamic fixation and 63.1 days for static fixation (P = 0.370). Similarly, the differences in time to fixator removal and to full weight bearing did not reach statistical significance. The conclusion was that axial dynamization of external fixation for pediatric femur fractures has no significant effect on time to healing or frequency of complications. PMID- 12131436 TI - Factors affecting forearm compartment pressures in children with supracondylar fractures of the humerus. AB - This study evaluated forearm compartment pressures in 29 children with supracondylar humerus fractures. Pressures were measured before and after reduction in the dorsal, superficial volar, and deep volar compartments at the proximal 1/6th and proximal 1/3rd forearm. Pressures in the deep volar compartment were significantly elevated compared with pressures in other compartments. There were also significantly higher pressures closer to the elbow within each compartment. Fracture reduction did not have a consistent immediate effect on pressures. The effect of elbow flexion on post-reduction pressures was also evaluated; flexion beyond 90 degrees produced significant pressure elevation. We conclude that forearm pressures after supracondylar fracture are greatest in the deep volar compartment and closer to the fracture site. Pressures greater than 30 mm Hg may exist without clinical evidence of compartment syndrome. To avoid unnecessary elevation of pressures, elbows should not be immobilized in >90 degrees of flexion after these injuries. PMID- 12131437 TI - Displaced pediatric supracondylar humerus fractures: biomechanical analysis of percutaneous pinning techniques. AB - Supracondylar humerus fractures are a common childhood occurrence. Displaced fractures are typically treated with closed reduction and percutaneous pinning. Controversy continues over the appropriateness of various pinning techniques. The most common include crossed or lateral pins. A biomechanical comparison of crossed pins, "parallel" lateral pins, and "divergent" lateral pins was performed using a pediatric synthetic bone model. Mechanical testing of each pin configuration was performed in extension, varus, valgus, internal rotation, and external rotation. The divergent configuration provided statistically greater stability than parallel pins under varus and valgus loading. Divergent pins had similar stability compared with crossed pins in extension, varus, and valgus testing. In axial rotation testing, crossed pins were more stable. If the surgeon feels confident in the ability of lateral pins to provide satisfactory fracture stability, divergent lateral pins provide greater stability than parallel lateral pins while avoiding ulnar nerve injury (associated with crossed pins). PMID- 12131438 TI - Predicting ulnar nerve location in pinning of supracondylar humerus fractures. AB - Thirty-four consecutive patients with displaced supracondylar humerus fractures were treated with reduction and percutaneous pinning. The precise location of the ulnar nerve to the medial pin was determined by intraoperative nerve stimulation. In 22 of the 34 patients, the authors attempted to predict the location of the ulnar nerve by palpation and placing a mark on the skin. They also recorded the ability to feel the anatomic landmarks for pin fixation, including the medial epicondyle and ulnar nerve. The average distance from the medial pin to the predicted location was 9.3 mm, whereas the actual distance measured 7.6 mm, for a significant difference of 1.7 mm. Statistically, the authors could not accurately predict the location of the ulnar nerve prior to blind percutaneous crossed K wire fixation of supracondylar humerus fractures. However, clinically they were fairly close in their prediction and documented safe insertion and distance from the nerve. Intraoperative nerve stimulation may assist in localizing the nerve prior to placement of the medial pin. Stimulation of the pin itself following insertion is another technique to ensure safe pin placement and decrease the risk of injury. PMID- 12131439 TI - Carpal scaphoid fracture nonunion in children. AB - Four new cases of carpal scaphoid nonunion in children are presented. This rare injury is characterized by delay in treatment of the scaphoid fracture or incorrect immobilization. The Matti-Russe procedure was performed with autograft harvested from the proximal cubitus, achieving complete consolidation in every patient. No complications occurred and there was no significantly limited mobility. The author speculates about the posibility of partial or complete consolidation of the scaphoid fracture without treatment as a part of the natural history of this fracture in children. PMID- 12131440 TI - Management and complications of anterior cruciate ligament injuries in skeletally immature patients: survey of the Herodicus Society and The ACL Study Group. AB - Expert opinion regarding experience with the management and complications of pediatric anterior cruciate ligament (ACL) injuries was studied by surveying members of The Herodicus Society and The ACL Study Group. There was large practice variation in initial management and ACL reconstruction technique. There were 15 reported cases of growth disturbance: 8 cases of distal femoral valgus deformity with arrest of the lateral distal femoral physis, 3 cases of tibial recurvatum with arrest of the tibial tubercle apophysis, 2 cases of genu valgum without arrest, and 2 cases of leg length discrepancy. Associated factors included fixation hardware across the lateral distal femoral physis in 3 cases, bone plugs of a patellar tendon graft across the distal femoral physis in 3 cases, large (12 mm) tunnels in 2 cases, fixation hardware across the tibial tubercle apophysis in 3 cases, lateral extra-articular tenodesis in 2 cases, and over-the-top femoral position in 1 case. Based on this experience, we recommend a guarded approach to ACL reconstruction in the skeletally immature patient with careful attention to technique and follow-up. PMID- 12131442 TI - Interlocking triple pelvic osteotomy in severe Legg-Calve-Perthes disease. AB - The authors reviewed 21 patients who underwent triple pelvic osteotomy for severe Legg-Calve-Perthes disease to evaluate their clinical, functional, and radiologic results. The mean age at presentation was 7 years 7 months (range 5-11 years). Seventeen hips were Herring group C and 5 were group B. All of them had "at risk" radiologic signs. A new technique of interlocking iliac osteotomy was used to provide extra stability, allow early weight bearing, and prevent inadvertent retroversion. The average period of follow-up was 30 months (range 1-5 years). The average gain in Harris score was 34.3 (range 4-55). The average gain in acetabular head index was 18% and that in center-edge angle was 22 degrees, more than reported for any other single surgical procedure for containment of the subluxed femoral head. Average gains in abduction, internal rotation, and flexion were 17 degrees, 12 degrees, and 28 degrees respectively. Longer follow-up is required to see the results at maturity, but the early results are promising. PMID- 12131441 TI - Bilateral Legg-Calve-Perthes disease: presentation and outcome. AB - Most patients who develop Legg-Calve-Perthes disease have unilateral involvement. For those children who do develop bilateral involvement, the disease and its outcome have not been characterized. This study reviewed the records and radiographs of 83 patients (20 girls and 63 boys) with bilateral Legg-Calve Perthes disease. The patients were then divided into 3 groups based on the Waldenstrom stage at the time of the first radiograph. In Group I (26 patients), both hips were in the same stage. In Group II (45 patients), the hips were in a different stage. In Group III (12 patients), the first hip was well into the remodeling stage by the time the second hip became affected. Twenty of the 83 patients (24%) were girls. There were significantly more lateral pillar group A hips on the second side than the first side in Groups II and III, and only 10 of the 45 patients (22%) in these groups had more severe disease in the second hip. When compared with data from a group of hips with unilateral involvement, there were significantly more hips with a Catterall group I rating in the patients with bilateral involvement. In general, the Stulberg et al. class assigned appeared to be independent of bilaterality. It appears that the development of bilateral disease is an independent event. The data in the present study do not support that onset of disease in one hip leads to disease or causes a more severe disease in the second hip. PMID- 12131443 TI - Contrast-enhanced power Doppler imaging: comparison with scintigraphic phases of revascularization of the femoral head in Legg-Calve-Perthes disease. AB - The authors evaluated the usefulness of an ultrasound contrast agent (SHU 508A) to help identify different scintigraphic phases of revascularization of the femoral head in children with Legg-Calve-Perthes (LCP) disease. Eighteen unenhanced and contrast-enhanced power Doppler images and scintigrams of the pathologic hip in 18 children with LCP disease were compared. The scintigraphic stages of Conway's classification for LCP disease (stage A, recanalization; stage B, neovascularization) were compared with the degree of vascularity and mean peak enhancement ratios obtained from analysis of Doppler sonograms. Qualitatively, the patient's age (< or = or >7 years old) at the time of examination had a significant effect on the degree of vascularity visualized on postcontrast ultrasound images according to the scintigraphic stages. Quantitatively, there were no differences for overall mean peak enhancement ratios between stages A and B. Power Doppler ultrasound increased visualization of Doppler signals significantly but did not help in differentiating scintigraphic phases. PMID- 12131444 TI - Herring classification: how useful is the initial radiograph? AB - The Herring lateral pillar classification has been shown to be a useful predictor of subsequent femoral head deformity in Perthes disease. The initial Herring classification, however, can often be misleading with regard to outcome, as further collapse can occur. The purpose of this study was to assess the predictability of the initial Herring classification. The radiographs of 253 patients (275 hips) conservatively treated were reviewed. The Herring grade was recorded on both the first and subsequent radiograph, and any change in grade was noted. Ninety-two hips required upgrading. The average duration of symptoms to the initial radiograph was 108 days compared with 204 days in the group showing no change. The predictability of the initial Herring classification is related to time from onset of symptoms. The greater the duration of symptoms, the more accurate the initial grading; 7 months is the average time at which one could expect no further collapse. PMID- 12131446 TI - Three-dimensional evolution of scoliotic curve during instrumentation without fusion in young children. AB - This study respectively evaluated 12 young children with progressive curves who underwent subcutaneous rod instrumentation (SCRI) without fusion and were followed until definitive surgery. The mean curve Cobb angle was 58 degrees at the time of SCRI. The mean apical vertebral rotation at the time of presentation was 20 degrees. Curves were observed to have progressed to an average of 59 degrees before definitive surgery and were corrected to 34 degrees with instrumentation. The average apical vertebral rotation was found to be 34 degrees before the definitive treatment. This value was significantly larger than the pre SCRI value (P = 0.002). The average number of lengthening operations per patient was 4.6, and the number of all operations per patient was 7. Six patients developed complications. The average time spent in hospital was 101 days. The findings demonstrate that the curves were essentially unchanged in the frontal plane during the period of lengthening. The sagittal curves remained mostly within normal limits, but there was an alarming increase in rotation. PMID- 12131445 TI - Sagittal profiles of the spine in scoliosis associated with an Arnold-Chiari malformation with or without syringomyelia. AB - The sagittal spine of children with Arnold-Chiari I malformation with or without syringomyelia and associated scoliosis (ACS) has been poorly studied. A retrospective review of scoliosis secondary to ACS from three centers was undertaken. Sagittal and coronal plane variables were measured from standing radiographs. There were 30 ACS children (19 girls, 11 boys) with an average curve of 50 +/- 20 degrees and age of 11.2 +/- 3.2 years. Syringomyelia was present in 26 (87%). The scoliosis was thoracic in 25, thoracolumbar in 3 and lumbar in 2; 18 curves were right and 12 were left. A positive correlation was noted between cervical lordosis (CL) and thoracic kyphosis (TK). The 30 children with ACS scoliosis were compared with 26 children with adolescent idiopathic scoliosis (AID). The ACS group had more left curves (40% vs. 0%, P = 0.0002), more boys (37% vs. 8%, P = 0.01), and was younger (11.3 +/- 3.7 years vs. 14.2 +/- 1.8 years, P = 0.004). TK and CL were increased in ACS (TK: 40 +/- 13 degrees vs. 30 +/- 13 degrees, P = 0.005; CL: 16 +/- 21 degrees vs. -5 +/- 12 degrees, P < 0.0001). The ACS and AID groups were subdivided by CL >0 degrees and <0 degrees. In the ACS group, 19% (5 of 21) had CL <0 degrees, in the AID group 77% (20 of 26) had CL <0 degrees (P = 0.0001). When cervical lordosis is >0 degrees or thoracic kyphosis is >40 degrees (Cobb method), the clinician should strongly suspect the presence of an Arnold-Chiari I malformation with or without syringomyelia. PMID- 12131447 TI - Callotasis lengthening in patients with brachymetacarpia. AB - Callotasis lengthening is an established method, but few cases of metacarpal lengthening have been reported. The authors performed callotasis lengthening to treat brachymetacarpia in six digits in three patients. The patient age at the time of distraction ranged from 10 to 19 years. The period of application of an external fixator averaged 13.9 weeks (range 10-19 weeks). All the metacarpals achieved the target length, and all patients were satisfied with the esthetic improvement. The length of the metacarpal distraction averaged 15.2 mm (range 10 18 mm). Complete consolidation of the transport gap was achieved for five of six digits. One digit that had a history of previous lengthening required bone graft. The average healing index was 62.3 days/cm. There were no serious complications. In four digits, the range of flexion of the metacarpophalangeal joint was increased after lengthening, and this enabled the patient to use the digit easily in a palmar grasp. Brachymetacarpia is an appropriate indication for callotasis lengthening in congenital deformities of the upper extremities. PMID- 12131448 TI - Chronic recurrent multifocal osteomyelitis: review of orthopaedic complications at maturity. AB - Chronic recurrent multifocal osteomyelitis (CRMO) is a childhood, bone disorder causing bone pain, swelling, malaise, and fever. A study of 22 children with CRMO, treated at the Royal Children's Hospital, Melbourne, was reported previously. This present study reviews 8 of these, together with 4 additional patients. The patients were interviewed and examined by the first two authors, who had not been involved in their management. The patients were 9 females and 3 males, with a mean age of 22 years (16-31 years). Age at onset of symptoms was between 4 and 11 years and duration 2.5-20 years. Only 1 patient experienced difficulties in school and in maintaining a job. However, only 2 patients experienced complete resolution of symptoms. The number of affected sites was 2 9, most commonly ankle, knee, and clavicle. Seven patients have noticeable deformity. Five had leg-length inequality of at least 1.5 cm, one of whom, with shortening of 5.5 cm, has undergone a lengthening procedure. CRMO is not a benign condition and if not followed to maturity can have disabling sequelae. PMID- 12131449 TI - Outpatient parenteral antimicrobial therapy in osteoarticular infections in children. AB - There are few data on the use of outpatient parenteral antimicrobial therapy (OPAT) in the management of osteoarticular infections (OAIs) in childhood. The objective of this study was to determine if OPAT is safe and effective in the management of OAIs. Using their OPAT database, the authors evaluated the use of OPAT in children younger than 18 years old treated for OAIs between January 1, 1995, and December 31, 1999. One hundred eighty-four OAIs were treated in 179 patients over 5 years. OPAT involved central venous lines (CVLs) in 110 (59.8%), peripherally inserted central catheters (PICCs) in 71 (38.6%), and peripheral cannulas in 3 (1.6%). One hundred eighteen (64%) OPAT courses were completed without interruption. Rehospitalization occurred in 48 (26.1%) courses and occurred earlier with PICC. OPAT complications were catheter-related in 58 (30%) courses, not catheter-related in 60 (32%), and unknown in 10 (5.3%). The mechanical complication rate was 6.3 per 1,000 catheter-days (CVL 4.2, PICC 10.6), and the rate of infectious complications was 2.7 per 1,000 catheter-days (CVL 2.8, PICC 2.4). One hundred sixty-eight (98%) of 172 evaluable OAIs were cured. Four (2.2%) patients failed treatment: one had recurrence and three had persistent infection. The authors conclude that OPAT can be safely used to manage OAIs in children without compromising outcome. Mechanical complications are more common with PICCs. PMID- 12131450 TI - Osteochondral sequelae of meningococcemia: radiographic aspects. AB - The authors retrospectively analyzed the files of 11 patients with osteochondral sequelae of meningococcemia, which were referred to this service between 1988 and 1996. The purpose of this study was to radiographically evaluate bone and physeal injuries observed in these patients, deformities caused by them and correlate these findings with the current literature. During radiographic evaluation of patients with sequelae of meningococcemia, two distinct parameters should be observed: bone injury and physeal plate injury patterns. The most frequent bone injury pattern was lytic lesion and the most common physeal plate injury pattern was peripheral asymmetric physis destruction. The most frequent deformities were lower limb length discrepancy, angular deformities and digital amputations. Based upon the findings of the present research, a descriptive radiographic classification was proposed. PMID- 12131451 TI - Ponseti versus traditional methods of casting for idiopathic clubfoot. AB - Serial casting is successful in avoiding extensive posteromedial release (PMR) in only 11% to 58% of patients with idiopathic congenital clubfoot. Extensive open surgery is commonly associated with long-term stiffness and weakness. Ponseti claims to avoid PMR in 89% of cases by using his specific technique of manipulation, casting, and limited surgery. The authors report their first 27 patients undergoing the Ponseti technique (34 feet) with a group of 27 matched control patients (34 feet). All patients underwent serial casting, begun within the first 3 months of life. The parameter studied was the need to perform PMR within the first year of life. In the Ponseti group, only 1 (3%) of 34 feet required PMR. In 31 (91%) of 34 feet, percutaneous Achilles tenotomy was performed at age 2 to 3 months. The average duration of casting was 2 months. In the control group, 32 (94%) of 34 feet required PMR within the first year of life, despite a longer casting period. Based on the authors' initial success with the Ponseti method, they no longer believe that PMR is required for most cases of idiopathic clubfoot. Foot abduction splints are crucial to avoid recurrence. Longer follow-up will determine whether the authors can continue to match Ponseti's reported outcomes. PMID- 12131452 TI - Association of fibular hemimelia and clubfoot. AB - This study was designed to determine the incidence of clubfoot in our fibular hemimelia population. A retrospective review of fibular hemimelia patients treated at our institution over the past seventy years was conducted. We identified twenty-three cases of clubfoot in 121 patients with 147 involved limbs. Nineteen of twenty-three limbs retained the foot, four were converted to Syme's amputation because a plantigrade functional foot could not be obtained. Hindfoot coalitions were present in eighteen limbs and nineteen had one or more missing lateral rays. In summary, the association of clubfoot deformity with fibular hemimelia is more common than previously reported. It was not always evident to the surgeon treating these patients that fibular hemimelia syndrome was present in addition to clubfoot deformity. The presence of a coalition is a relatively constant finding in this condition and should be anticipated by the treating clubfoot surgeon. PMID- 12131453 TI - Clubfoot posteromedial release: advantages of tibialis anterior tendon lengthening. AB - The aim of this study is to evaluate the eventual advantages of tibialis anterior (TA) tendon lengthening during clubfoot posteromedial release. A continuous series of 60 idiopathic clubfeet has been retrospectively studied. Tibialis anterior lengthening (TAL) began to be performed in 1984. Two groups of 30 feet have been distinguished: without TAL (before 1984) and with TAL (after 1984). There was no significant difference between the 2 groups concerning mean age at surgery, preoperative clinical and radiologic data. Mean postoperative follow-up was 10 years and minimal follow-up required was 5 years. TAL decreased Triceps surae relative insufficiency and improved monopodal jump. TAL balanced TA and peroneus longus, decreased dynamic supination and balanced forefoot pronation and supination. The feet without TAL presented lack of anteromedial support (20% without TAL, 0% with TAL) and medial arch cavus with dorsal talo-navicular subluxation (20% without TAL, 3,3% with TAL). TAL decreased the rate of recurrence and surgical revision. PMID- 12131454 TI - Aneurysmal bone cyst in 29 children. AB - The purpose of this study was to review longitudinally the clinical features, method of treatment, and recurrence rates of 40 cases of aneurysmal bone cyst in children treated at one institution. Twenty-nine patients with histologic confirmation of the diagnosis and minimum follow-up of 2 years were included. Thirteen patients were less than 10 years of age, and 16 were in the second decade of life. The most frequent location of the lesion was the tibia (seven cases). Patients were treated with curettage, curettage and bone grafting, or resection. The overall recurrence rate was 27.5%. Five lesions recurred once, and three recurred twice. The average time before recurrence was 18.7 months. Complications occurred in six cases, three of them with physeal arrest. The lesion occurred more commonly in females in the second decade of life and was not associated with a pre-existing tumor. The recurrence rate dropped after the use of a high-speed bur. PMID- 12131455 TI - Minimal curettage, multiple drilling, and continuous decompression through a cannulated screw for treatment of calcaneal simple bone cysts in children. AB - This report concerns a series of 12 patients with simple bone cysts of the calcaneus, who were treated between 1988 and 2000 by a minimal surgical intervention of curettage, multiple drilling and continuous decompression through insertion of a cannulated screw. Inserted screws were made of titanium in 8 cases and ceramics in 4 cases. The mean operative time was 58.8 +/- 25.4 minutes, and complete healing was achieved in 11 cases after 9.2 +/- 6.4 months. One patient needed an additional surgery for replacement of a titanium screw. Final results were excellent after a mean follow up of 91 +/- 52.1 months. This series of patients represents one of the largest reported series of calcaneal simple bone cyst in conjunction with long-term follow-up. Our technique of minimal curettage, drilling and continuous decompression with a cannulated screw insertion is considered to be a good option for management of simple bone cysts of the calcaneus. PMID- 12131456 TI - Sternoclavicular joint ganglion cysts in young children. AB - Ganglion cysts originating from the sternoclavicular joint in children have not been previously reported. In this study, 5 children who presented with a small mass over the anterior aspect of the sternoclavicular joint were evaluated and treated. Only 1 patient was symptomatic. A ganglion cyst was suspected in each case and confirmed by magnetic resonance imaging in 3 patients and ultrasound in one patient. Excisional biopsy was performed in 3 patients and the diagnosis of a ganglion cyst confirmed histopathologically. No patient has had a recurrence. Observation of asymptomatic cystic lesions that arise in the sternoclavicular location is recommended. PMID- 12131457 TI - Changes in the management of monteggia fractures. PMID- 12131458 TI - Results of Charleston Bracing in skeletally immature patients with idiopathic scoliosis. PMID- 12131459 TI - Sonographic healing stages of achilles tendon after tenomuscular lengthening in children with cerebral palsy. PMID- 12131460 TI - Ender nail fixation of pediatric femur fractures: a biomechanical analysis. PMID- 12131462 TI - Managing carotid-cavernous fistulas in Ehlers-Danlos syndrome type IV. PMID- 12131463 TI - Spontaneous direct carotid-cavernous fistula in Ehlers-Danlos syndrome type IV: two case reports and a review of the literature. AB - Two unrelated adults with Ehlers-Danlos syndrome type IV developed acute unilateral ophthalmoplegia and ipsilateral headache as a consequence of spontaneous (nontraumatic) direct carotid-cavernous fistulas. Because the interventional radiologist suspected the diagnosis of Ehlers-Danlos syndrome type IV, the carotid-cavernous fistulas were closed via the venous rather than the more standard arterial route in an attempt to avoid arterial dissection or rupture. In any patient presenting with a spontaneous direct carotid-cavernous fistula, family history and clinical examination should be targeted toward a diagnosis of Ehlers-Danlos syndrome type IV because of risks attendant to angiography and repair of the fistula. For these patients, ancillary medical care must be approached cautiously to avoid hollow viscus rupture. Molecular tests can be used to confirm the diagnosis and provide family members with accurate genetic counseling and predictive genetic testing. PMID- 12131464 TI - Migraine-like visual hallucinations in occipital lesions of cysticercosis. AB - Four Indian patients with occipital lesions of cysticercosis presented with visual hallucinations. Neuro-ophthalmic and systemic examinations were normal in all cases except for one patient who had a partial homonymous hemianopia. Electroencephalography was normal in all cases. Neuroimaging revealed ring enhancing lesions in the occipital lobe typical of neurocysticercosis. In endemic regions like India, neurocysticercosis should be suspected in patients presenting with visual hallucinations, even when there are no other clinical findings. PMID- 12131465 TI - Oscillopsia without nystagmus caused by head titubation in a patient with multiple sclerosis. AB - Oscillopsia in patients who have brain stem disorders but not nystagmus is attributed to a failure of the vestibular-ocular reflex (VOR) to compensate for head movements. We report a patient who had marked head titubation and oscillopsia in aggressive multiple sclerosis but no nystagmus. Her severe head titubation precluded our ability to measure a VOR accurately. Because oscillopsia has also been described after rapid voluntary head oscillations in normal subjects, we queried whether the oscillopsia in our patient could be ascribed to the head movement alone. Six normal control subjects did not experience oscillopsia while shaking their heads at the same frequency as the patient's titubation. We conclude that the oscillopsia in our patient was probably the result of an impaired VOR or an alternative compensatory mechanism. PMID- 12131466 TI - Isolated unilateral adduction deficit and ptosis as the presenting features of chronic inflammatory demyelinating polyradiculoneuropathy. AB - A patient with chronic inflammatory demyelinating polyneuropathy (CIDP) presented with an isolated unilateral adduction deficit and ptosis. Investigations were negative until the onset of limb weakness and fatigue 2 years later. At that time, electroneuromyography, cerebrospinal fluid examination, and magnetic resonance imaging confirmed the diagnosis of CIDP. Thus, ophthalmic signs can precede extremity and bulbar signs with a long latency in CIDP. PMID- 12131467 TI - Fourth nerve palsy, homonymous hemianopia, and hemisensory deficit caused by a proximal posterior cerebral artery aneurysm. AB - A 21-year-old man developed an ipsilateral fourth nerve palsy, contralateral hemianopia, and contralateral hemisensory deficit as manifestations of a proximal right posterior cerebral artery aneurysm. This unusual constellation of signs reflects the involvement of the structures that run in the ambient cistern. The fourth nerve palsy and homonymous hemianopia are attributed to compression by the aneurysm. The hemisensory loss is ascribed to compromise of thalamoperforate arteries emanating from a thrombosed portion of the aneurysm. PMID- 12131468 TI - Ocular motor features of alternating hemiplegia of childhood. AB - A 14-month-old boy with alternating hemiplegia of childhood, an idiopathic disorder of early childhood causing episodic hemibody tonic spasms and hemiplegia, showed repetitive jerks of abduction of the ipsilateral eye during the spells. The mechanism of this ocular motor abnormality is unknown but may be unique to this disorder. PMID- 12131469 TI - Delayed visual loss following pergolide treatment of a prolactinoma. AB - A patient who had achieved marked improvement in vision and shrinkage of a prolactinoma following treatment with pergolide (0.1 mg/day) suffered a marked worsening of vision 7 months after continued treatment at the same dose. Brain magnetic resonance imaging (MRI) at the time of visual loss showed further shrinkage of the tumor and prolapse of the chiasm into the pituitary fossa. The dose of pergolide was cut in half (0.05 mg/day); 12 months later, vision had completely recovered. Brain MRI at the time of visual recovery showed no change in the position of the prolapsed chiasm. This is the 11th reported case of delayed visual loss following dopaminergic treatment of prolactinoma. Recovery of vision always occurs with reduction of the medication dosage. Many patients whose prolactinomas are treated in this fashion display chiasmal prolapse, and few suffer visual loss. Considering that visual recovery occurs without a visible change in the position of the chiasm, traction is an unlikely cause of delayed visual loss. Therefore, the term chiasmal traction syndrome, used to describe visual loss with prolapsed chiasm following surgical and radiation treatment of sellar tumors, should not be applied in this setting lest it prompt consideration of surgical chiasmapexy. The proper management is reduction of the dopaminergic agonist dosage. PMID- 12131470 TI - Periodic alternating nystagmus provoked by an attack of Meniere's disease. AB - Periodic alternating nystagmus is a rare central nervous system disorder in which the eyes undergo a horizontal jerk nystagmus that periodically reverses direction. A patient with a hypoplastic cerebellum and enlarged cisterna magna exhibited transient periodic alternating nystagmus following an attack of Meniere's disease. We hypothesize that in susceptible individuals with cerebellar disturbances, periodic alternating nystagmus may be transiently induced by vestibular stimuli. PMID- 12131471 TI - Prolonged premonitory optic disc signs in anterior ischemic optic neuropathy. AB - A patient displayed a pink mass on the right optic disc and normal visual function that was diagnosed as a capillary hemangioma. Seven months later, he developed typical features of nonarteritic anterior ischemic optic neuropathy (NAION) in that eye. Such a long latency between "preeruptive" and "eruptive" disc edema has not been well documented in NAION. PMID- 12131472 TI - Bilateral crocodile tears in a patient with Guillain-Barre Syndrome. AB - We describe the first reported case of the development of bilateral crocodile tears in Guillain-Barre Syndrome. This finding is an expression of axonal degeneration in the acute phase and misdirection-in-regeneration in the chronic phase. PMID- 12131473 TI - Superior segmental optic nerve hypoplasia. AB - A visually asymptomatic 27-year-old man was found to have inferior altitudinal visual field defects binocularly. Ophthalmoscopy revealed superior segmental optic pallor with superior nerve fiber layer atrophy, nicely highlighted in red free photographs. The patient's mother had insulin-dependent diabetes mellitus. Recognition of this entity is important for prognosis and for avoidance of unnecessary diagnostic studies. PMID- 12131474 TI - Diffusion-weighted magnetic resonance imaging. AB - Diffusion-weighted magnetic resonance imaging is a specialized technique that measures the degree of diffusion of water molecules within extracellular space and between intracellular and extracellular space. Diffusion-weighted imaging signal is high (bright) when diffusion is restricted, as occurs in cytotoxic damage from ischemia, inflammation, trauma, or tumor. This technique, now available on most magnetic resonance imaging units, is especially helpful in detecting early ischemic stroke and multiple sclerosis and in differentiating arachnoid cyst from epidermoid tumor and brain abscess from neoplasm. PMID- 12131475 TI - Best catch from NANOSNET. PMID- 12131476 TI - The 27th International Stroke Conference San Antonio, Texas, February 7-9, 2002. PMID- 12131499 TI - Downbeat nystagmus and dolichoectasia of the vertebrobasilar artery. PMID- 12131504 TI - Presidential address. PMID- 12131505 TI - Test your knowledge: preparing to take the CRNI Exam. AB - The CRNI Certification Examination consists of 200 multiple-choice questions covering the nine core content areas of infusion nursing: Technology and Clinical Applications, Fluid and Electrolyte Balance, Pharmacology, Infection Control, Pediatrics, Transfusion Therapy, Antineoplastic Therapy, Parenteral Nutrition, and Quality Assurance. The review questions provided below are modeled on the CRNI exam and are intended to help exam candidates test their knowledge of infusion therapy practice. This special section is a regular addition to the Journal of Infusion Nursing, with each edition focusing on a single content area. PMID- 12131507 TI - Evaluating new technology to improve patient outcomes: a quality improvement approach. AB - The nurses in the peripherally inserted central catheter (PICC) program at the University of Washington Medical Center perform ongoing data tracking to measure team and patient outcomes. Quality improvement initiatives have included the transition to microintroducer technology and ultrasound-guided placement. Used together, this technology has allowed the PICC team to increase their bedside insertion success rate to 91%. The group has also changed PICC securement methods, and use of the Statlock anchoring device has reduced the incidence of catheter migration from 6% to 1.5% of all catheter lines placed. Catheter durability also was assessed. The Pressure Activated Safety Valve PICC was compared to the Groshong PICC and the rate of catheter repair and exchange due to breakage has been reduced from 11% to 1%. PMID- 12131506 TI - A clinical study comparing the skin antisepsis and safety of ChloraPrep, 70% isopropyl alcohol, and 2% aqueous chlorhexidine. AB - ChloraPrep (2% chlorhexidine gluconate + 70% isopropyl alcohol [CHG + IPA] in a 3.0-mL applicator) is a recently approved antiseptic for preoperative skin preparation. This controlled open-label trial assessed the immediate and persistent antimicrobial efficacy and safety of CHG + IPA compared with 70% IPA or a 2% CHG aqueous solution alone. Each antiseptic significantly reduced abdominal and inguinal microbial counts from baseline at 10 minutes, 6 hours, and 24 hours (P =.0001). CHG + IPA provided significantly more persistent antimicrobial activity on abdominal sites than IPA (P =.003) or CHG (P =.028) at 24 hours. No skin irritations were reported for any of the three antiseptics. PMID- 12131508 TI - Comparison of two methods of peripheral intravenous cannula securement in the pediatric setting. AB - This prospective, nonrandomized study compared the effect of two peripheral catheter dressings (a combination transparent polyurethane film/soft cloth surgical tape dressing, and adhesive tape) on the incidence of complications in children and adolescents. A total of 407 catheter dressings were studied: 212 in the control group (adhesive tape) and 195 in the study group (transparent dressing). Catheter insertion site assessments at 24-hour intervals showed increased site visibility, better dressing adherence and less dressing reinforcement in the study group. There were few differences in the observed incidence of phlebitis or extravasation. The new combination dressing may be considered for use in children when prolonged catheterization is anticipated. PMID- 12131509 TI - Using unconjugated antibodies as an immunotherapeutic approach to treatment of B cell neoplasms. AB - B-cell neoplasms have the sixth highest mortality incidence and one of the largest rates of increase of all malignancies. Frequent relapse and the development of refractory disease in this patient group have stimulated research to find alternative treatments. The use of unconjugated antibodies offers a new approach to the treatment of non-Hodgkin's lymphoma and chronic lymphocytic leukemia. This article will explore the pharmacology, dosage, and administration guidelines as well as side effect management of the available drugs in this classification. Expectations for future approaches to the treatment of B-cell neoplasms will be discussed. PMID- 12131510 TI - Get a grip on patient safety: outcomes in the palm of your hand. AB - Documentation of what nurses do and the consequential impact on the care and safety of the patient is essential for the optimal use of intravascular devices. The University of Louisville Hospital's infection control department collaborated with the infusion therapy team on a project designed to provide an easier and more reliable way to quantify what the infusion therapy team did and the resultant patient outcomes. This project was based on software developed by the infection control department for use with the handheld personal digital assistant (PDA). This article will discuss how use of the PDA and software meet individual departmental needs and impact patient outcomes and patient safety by using evidence-based decision-making. PMID- 12131511 TI - Selective leptin resistance: a new concept in leptin physiology with cardiovascular implications. AB - Leptin, an adipocyte secreted hormone, acts in the hypothalamus to inhibit appetite and promote thermogenic metabolism, thereby reducing adiposity and body weight. Leptin has multiple autonomic and cardiovascular actions, including sympathetic activation, increases in endothelium derived nitric oxide (NO), and angiogenesis. The predominant cardiovascular effect of chronic hyperleptinemia is a pressor effect mediated by increased sympathetic activity. The sympathetic and cardiovascular actions of leptin are discussed and evidence derived from studies of obese mice for the novel concept of selective leptin resistance is reviewed. This concept holds that in some obese states, there is preservation of the sympathoexcitatory actions of leptin despite resistance to the satiety and weight reducing actions of the hormone. Selective leptin resistance might explain how hyperleptinemia could contribute to increases in sympathetic activity and arterial pressure in obese states where there is resistance to the metabolic (satiety and weight-reducing) actions of leptin. It is speculated here, that this concept may have potential implications for human obesity, which is often associated with elevated plasma leptin and partial resistance to the satiety effects of leptin. If selective leptin resistance occurs in obese humans, then leptin could contribute to the sympathetic overactivity and hypertension despite resistance to its metabolic actions. PMID- 12131512 TI - Blood pressure control and the implementation of guidelines in clinical practice: can we fill the gap? PMID- 12131513 TI - Enhancing risk stratification in hypertensive subjects: how far should we go in routine screening for target organ damage? PMID- 12131514 TI - Microarray analysis of gene expression in vascular smooth muscle cells from spontaneously hypertensive rats at an early age. PMID- 12131515 TI - Prostaglandin D2 : a Cinderella of vascular cell biology? PMID- 12131517 TI - Angiotensin II, the endothelium and superoxide anions. PMID- 12131516 TI - Angiotensin II type 1 receptors and oestrogen status: interaction or dissociation? PMID- 12131518 TI - Collagen and hypertension. PMID- 12131519 TI - Obesity-related hypertension: relevance of vascular responses to mental stress. PMID- 12131520 TI - Insulin resistance, oxidative stress and aspirin: therapeutic implications? PMID- 12131521 TI - Angiotensin II-dependent structural changes of preglomerular resistance may affect long-term control of blood pressure by resetting the renal 'baroreceptor'. PMID- 12131522 TI - Dipyridamole or dobutamine in arterial hypertension: are sensitivity and specificity the sole keywords? PMID- 12131523 TI - Left ventricular remodelling impacts on coronary flow reserve in hypertensive patients: is there a vascular mechanism? PMID- 12131524 TI - Left ventricular hypertrophy and remodelling of resistance arteries: the role of activation of the renin-angiotensin-aldosterone system in hypertension. PMID- 12131525 TI - Survey on treatment of hypertension and implementation of World Health Organization/International Society of Hypertension risk stratification in primary care in Belgium. AB - OBJECTIVE: To gain insight into the prevalence, treatment and control of hypertension and into the implementation of the 1999 World Health Organization/International Society of Hypertension guidelines for the management of hypertension in general practice in Belgium. DESIGN: A prospective cross sectional survey. SETTING: Primary care. METHODS: Participating physicians enrolled the first 15 men, at least 55 years old, who visited the surgery, measured their blood pressure with a validated automatic device and recorded data on age, medical history, drug utilization, cardiovascular risk factors and target organ damage. Patients were considered to have hypertension when systolic blood pressure was >or= 140 mmHg, diastolic blood pressure was >or= 90 mmHg or when they were under antihypertensive therapy. RESULTS: Among 3761 evaluable patients, 74% were considered to be hypertensive, 80% of whom were treated with antihypertensive drugs. Blood pressure was under control in 38% of the treated patients and in 31% of all hypertensives. Among the 1316 hypertensive patients in whom risk stratification was possible, 47, 56 and 86% of the patients in, respectively, the medium, high and very high risk groups were treated with antihypertensive drugs. Among the treated patients, 46, 37 and 31%, respectively, had reached goal pressure. Within each risk category, patients were treated more frequently when baseline blood pressure was higher. Logistic regression analysis revealed that hypertension grade and level of risk contributed independently to the odds of being treated. CONCLUSIONS: The results indicate that a large number of older hypertensive men are treated with antihypertensive drugs in primary care, but that the goal blood pressure is not reached in a substantial number of patients due to undertreatment. Furthermore, whereas patients at higher risk are treated more frequently than patients at lower risk, blood pressure itself remains an important factor for the initiation of antihypertensive drug therapy within each risk category. PMID- 12131526 TI - Migraine is more frequent in individuals with optimal and normal blood pressure: a population-based study. AB - BACKGROUND: The notion that hypertension causes headache is widely accepted despite the absence of confirmation by well-designed studies. OBJECTIVE: To investigate the association between headache, characterized as tension type and migraine like, with blood pressure and hypertension. METHODS: In a cross sectional study we evaluate this association in a sample of 1174 individuals older than 17 years, representative of inhabitants of Porto Alegre, RS, Brazil. Headache and its subtypes were defined according to International Headache Society criteria. Hypertension was defined as the mean of two blood pressure readings >or=140/90 mmHg or use of antihypertensive drugs. RESULTS: Headache in lifetime, in the last year, and defined as episodic and chronic tension-type headache was not associated with hypertension. Individuals with optimal or normal blood pressure (Sixth Joint National Committee criteria) complained of migraine more frequently than the participants with high-normal blood pressure or hypertension. This association persisted after adjustment for several potential confounding factors (risk ratio, 0.56; confidence interval, 0.41-0.77). CONCLUSION: Our findings confirm that high blood pressure is not associated with the complaint of headache in the population. Individuals with migraine-like episodes of headache may have lower blood pressure than individuals without headache. PMID- 12131527 TI - Role of echocardiography and carotid ultrasonography in stratifying risk in patients with essential hypertension: the Assessment of Prognostic Risk Observational Survey. AB - BACKGROUND: Echocardiography and carotid ultrasonography, by providing a more accurate assessment of cardiac and vascular damage related to hypertension, may lead to a more precise stratification of the global cardiovascular risk. However, current guidelines do not recommend systematic use of ultrasound examination of heart and large arteries in evaluating the cardiovascular risk in patients with hypertension. OBJECTIVE: To assess the impact of echocardiography and carotid ultrasonography on global risk stratification in hypertensive patients classified as being at low or medium risk according to routine clinical work-up as suggested by current hypertension guidelines. METHODS: Among 8502 consecutive patients screened at 44 outpatient hypertension hospital clinics in different parts of Italy, 1074 untreated individuals with low-to-medium risk essential hypertension were identified on the basis of the diagnostic routine procedures suggested by 1999 World Health Organization/International Society of Hypertension guidelines: medical history, physical examination and clinic blood pressure measurement; routine blood chemistry and urine analysis; electrocardiogram. The extent of risk for the 1074 individuals was reassessed by adding the results of ultrasound examinations of heart and carotid arteries: left ventricular hypertrophy (defined as left ventricular mass index > 120 g/m(2) in men and > 100 g/m(2) in women), carotid intima-media thickening (defined as diffuse thickening if >or= 0.8 mm), and presence of plaque (defined as focal thickening > 1.3 mm). RESULTS: According to routine classification, 18.7% (n = 201) of the 1074 patients were considered at low risk and 81.3% (n = 873) at medium risk. A marked change in risk stratification was obtained when ultrasound markers of target-organ damage were taken into consideration: the proportion of low-risk patients decreased to 11.1%, and that of medium risk patients to 35.7%, whereas more than 50% of the patients previously classified at low-medium risk were found to be at high absolute risk. According to a multivariate analysis, age, grade of hypertension, male sex, and serum cholesterol concentration were the variables with the greatest impact on risk class change. CONCLUSIONS: Ultrasound assessment of the heart and carotid wall helps to obtain a more valid assessment of global cardiovascular risk in hypertensive patients without evidence of target-organ damage after routine examination. PMID- 12131528 TI - Microalbuminuria identifies overall cardiovascular risk in essential hypertension: an artificial neural network-based approach. AB - BACKGROUND: Ultrasound (US) examination of heart and carotid arteries provides an accurate assessment of target organ damage (TOD) and may influence the stratification of the absolute cardiovascular risk profile. Microalbuminuria has recently proved to be a useful cost-effective marker of increased cardiovascular risk but is still too often neglected in clinical practice. OBJECTIVE: To evaluate how well artificial neural networks (ANNs) predict cardiovascular risk stratification by means of routine data and urinary albumin excretion, as compared to prediction by the clinical work-up suggested by the International Society of Hypertension (ISH), with and without ultrasound-determined TOD. METHODS: A group of 346 never previously treated essential hypertensives (212 men, 134 women, mean age 47 +/- 9 years) was studied. Risk was stratified according to the criteria suggested by the 1999 WHO/ISH guidelines; first, by routine procedures alone, and subsequently by reassessment, using data on cardiac and vascular structures obtained by US evaluation. The ANN was trained and tested to predict the overall cardiovascular risk on the basis of routine clinical data and urinary albumin excretion (UAE). The impact of these three approaches on the determination of cardiovascular risk profile was evaluated. RESULTS: According to the first classification, 5.5% (n = 19) of patients were considered at low risk, 47.3% (n = 164) at medium, 26.7% (n = 92) at high and 20.6% (n = 71) at very high risk. A marked change in risk stratification, namely an increase in the prevalence of high- and very-high-risk patients (2.3% low, 29.8% medium, 42.8% high and 25.2% very high risk; chi(2) 15.201, P < 0.0001), was obtained when US examination of TOD was taken into consideration. On the basis of routine clinical data and UAE, the artificial neural network successfully predicted overall cardiovascular risk and allocated patients in different classes as accurately as the US-based evaluation. CONCLUSIONS: The use of US techniques allows a more precise stratification of absolute cardiovascular risk in hypertensive patients as compared to routine clinical data. An ANN can accurately identify the patients' risk status by using low-cost routine data and UAE. These results further emphasize the value of UAE in the stratification of cardiovascular risk. PMID- 12131529 TI - Growth characteristics, angiotensin II generation, and microarray-determined gene expression in vascular smooth muscle cells from young spontaneously hypertensive rats. AB - BACKGROUND: We have demonstrated that vascular smooth muscle cells (VSMCs) derived from spontaneously hypertensive rats (SHR) show exaggerated growth and produce angiotensin (Ang) II and growth factors. These may reflect intrinsic abnormalities in SHR that are not caused by excessive blood pressure, and are associated with genetic abnormalities. OBJECTIVE: To evaluate whether these characteristics of VSMCs from SHR are associated with hypertension or genetic factors. DESIGN AND METHODS: VSMCs were obtained by an explant method from aortas of 4-week-old male SHR/Izumo and Wistar-Kyoto (WKY)/Izumo rats. We evaluated growth characteristics by [3H]thymidine incorporation and cell number increases, immunofluorescence of alpha-smooth muscle (alpha-SM) actin, mRNA expressions of phenotype markers, Ang II-generating system components, and growth factors by reverse transcription and polymerase chain reaction analysis, and Ang II levels by radioimmunoassay in VSMCs. Expression of 850 genes in VSMCs was evaluated by microarray. RESULTS: VSMCs from young SHR showed increased basal DNA synthesis and higher responses of DNA synthesis and cell numbers in response to calf serum. Ang II was significantly increased in conditioned medium and cell extracts from SHR-derived VSMCs than in those from WKY rat-derived VSMCs. mRNA expression of Ang II-generating proteinases, such as cathepsin D and angiotensin-converting enzyme, was greater in VSMCs from SHRs than in cells from WKY rats. Expression of transforming growth factor-beta1, platelet-derived growth factor A-chain and basic fibroblast growth factor mRNAs was greater in VSMCs from SHRs than in cells from WKY rats. Expression of mRNAs of phenotype markers, such as matrix gamma carboxyglutamic acid (Gla) and osteopontin, was also greater in VSMCs from SHR than in cells from WYK rats. Microarray study showed that VSMCs derived from young SHR increasingly express genes for many enzymes, adhesion molecules and cytokines. CONCLUSION: This study determined that VSMCs derived from young SHR show exaggerated growth, produce Ang II and increasingly express several enzymes, adhesion molecules and cytokines, which are independent of hypertension and possibly associated with genetic abnormalities. PMID- 12131530 TI - Early decline in the catecholamine release-inhibitory peptide catestatin in humans at genetic risk of hypertension. AB - BACKGROUND: Hypertension is a complex trait with an ill-defined genetic predisposition, in which adrenergic mechanisms seem to be involved even at the early stages. Chromogranin A is a pro-hormone stored and released with catecholamines by exocytosis; its fragment catestatin, formed in vivo, inhibits further catecholamine release as an antagonist at the physiologic trigger for secretion, the neuronal nicotinic cholinergic receptor. METHODS: We measured catestatin by radioimmunoassay in n = 277 subjects stratified by blood pressure (n = 61 hypertensive, n = 216 normotensive), and if normotensive by genetic risk of developing hypertension: family history positive (n = 176) versus negative (n = 40). Maximum likelihood analysis tested for bimodality. Involvement of catestatin in pathophysiology was probed by measurements of catecholamines and leptin, and the hemodynamic responses to environmental (cold) stress. RESULTS: The normotensive offspring of patients with hypertension already had diminished catestatin (P = 0.024), and family history was a better predictor of catestatin than age, ethnicity or gender (P = 0.014). Greater catestatin variance among family history-positive individuals (P = 0.021) suggested heterogeneity in this group, and a bimodal distribution (P < 0.001) identified 4.3% of individuals in a lower mode of catestatin values, all with positive family histories (P = 0.05). Catestatin correlated inversely with body mass index (r = -0.215, r(2) = 0.046, n = 276, P < 0.001) and plasma leptin (r = -0.203, r(2) = 0.041, n = 212, P = 0.003), while body mass index and leptin correlated directly (r = 0.59, r(2) = 0.350, n = 212, P < 0.001). Family history-positive individuals had greater epinephrine excretion (P = 0.037) in addition to diminished catestatin, suggesting an inhibitory effect of catestatin on chromaffin cells in vivo. Low plasma catestatin predicted enhanced pressor response to a sympathoadrenal stressor (cold stress; r = -0.184, r(2) = 0.034, n = 211, P = 0.007), suggesting an adrenergic mechanism whereby diminished catestatin might predispose to later development of hypertension. In white subjects, diminished catestatin also predicted greater systemic vascular resistance responses to cold stress (r = 0.307, r(2) = 0.094, n = 75, P = 0.007), a relationship not found in Blacks (r = 0.122, r(2) = 0.015, n = 94, P = 0.243). CONCLUSIONS: We conclude that catestatin is diminished early in the course of development of hypertension, even in the normotensive offspring of patients with the disease. Low catestatin predicts augmented adrenergic pressor responses, suggesting a mechanism whereby diminished catestatin might increase the risk for later development of hypertension. PMID- 12131531 TI - Endogenous prostaglandin D2 synthesis reduces an increase in plasminogen activator inhibitor-1 following interleukin stimulation in bovine endothelial cells. AB - OBJECTIVE: We examined the role of prostaglandin D2 (PGD2) in the formation of plasminogen activator inhibitor (PAI)-1 following interleukin-1beta (IL-1) stimulation in bovine endothelial cells (EC) transfected with lipocaline-type PGD2 synthase (L-PGDS) genes. DESIGN AND METHODS: EC were isolated from bovine thoracic aorta and incubated with 20 U/ml IL-1 and various concentrations of authentic PGD2. The isolated EC were also transfected with L-PGDS genes by electroporation. The L-PGDS-transfected EC were used to investigate the role of endogenous PGD2 in IL-1 stimulated PAI-1 biosynthesis. We also used an anti-PGD2 antibody to examine whether an intracrine mechanism was involved in PAI-1 production. PGD2 and PAI-1 levels were determined by radio- and enzyme immunoassay, respectively. PAI-1 mRNA was assessed by reverse transcription polymerase chain reaction. RESULT: IL-1 stimulated PAI-1 production by EC was dose-dependently inhibited by authentic PGD2 at concentrations greater than 10-6 mol/l. L-PGDS gene-transfected EC produced more PGD2 than EC transfected with the reporter gene alone. IL-1 induced increases in PAI-1 production in EC transfected with reporter genes alone. However, this effect was significantly attenuated in the case of IL-1 stimulation of EC transfected with L-PGDS genes, and accompanied by an apparent suppression of PAI-1 mRNA expression. The effects of PGD2 on PAI-I formation were reversed to the basal levels by the inhibition of synthesis of endogenous PGD2. Neutralization of extracellular PGD2 by anti-PGD2 antibody influenced neither PAI-1 mRNA expression nor PAI-1 biosynthesis. CONCLUSION: EC transfected with L-PGDS genes increased PGD2 synthesis. This was associated with attenuation of both PAI-1 formation and PAI-1 mRNA expression. It is suggested that endogenous PGD2 decreases PAI-1 synthesis and PAI-1 mRNA expression, probably through an intracrine mechanism. PMID- 12131532 TI - Cerebrovascular alterations in pressure and protein kinase C-mediated constriction in Dahl salt-sensitive rats. AB - BACKGROUND: Dahl salt-sensitive (DSS) rats fed an 8.7% sodium chloride diet from weaning spontaneously developed hypertension and a 50% mortality rate by 5 weeks. Before death the rats exhibited behavioural signs of stroke and disruption of the blood-brain barrier. OBJECTIVES: To test the hypothesis that rats exhibiting stroke had middle cerebral arteries (MCAs) that had lost the ability to constrict in response to pressure, and to assess whether this defect was associated with abnormalities in protein kinase C (PKC)-mediated constriction. METHODS: MCAs were sampled from DSS rats before and after stroke and from Dahl salt-resistant (DSR) rats fed 8.7% NaCl. Constrictions in response to a 100 mmHg pressure step and to PKC activation by phorbol dibutyrate (PDB) (0.1 micromol/l) in the presence of nifedipine (3 micromol/l) were measured. RESULTS: MCAs from DSS rats after stroke constricted in response to vasopressin but were unable to constrict in response to pressure or PDB in the presence of nifedipine, whereas those from DSS rats before stroke and from DSR rats constricted in response to all the stimuli. The PKC inhibitors, chelerythrine (12 micromol/l) and bisindolylmaleimide (5 micromol/l) inhibited constrictions in response to pressure and to PDB in the presence of nifedipine. CONCLUSIONS: Constriction of the MCA in response to pressure is dependent on functional PKC signalling. Development of stroke in DSS rats fed a high-salt diet is associated with an inability of the MCAs to constrict in response to pressure, possibly because of the presence of an incompetent PKC system. The inability to constrict in response to pressure may cause blood flow abnormalities that contribute to disruption of the blood-brain barrier in these rats. PMID- 12131533 TI - Angiotensin II type 1 receptor blocker irbesartan ameliorates vascular function in spontaneously hypertensive rats regardless of oestrogen status. AB - BACKGROUND: Angiotensin II type 1 (AT1) receptor overexpression may play a decisive role in endothelial dysfunction during oestrogen deficiency in spontaneously hypertensive rats (SHRs). Similarly, exaggerated production of angiotensin II and enhanced expression of AT1 receptor have been reported in vessels of SHRs compared with normotensive rats. OBJECTIVE: To test the hypothesis that antihypertensive treatment with the AT1 receptor antagonist, irbesartan, could not only decrease blood pressure but also ameliorate endothelial dysfunction associated with both hypertension and oestrogen deficiency. METHODS: Ovariectomized and sham-ovariectomized SHRs were treated with 50 mg/kg irbesartan per day, administered with chow for 30 weeks. Sham ovariectomized and ovariectomized rats receiving no treatment were used as control groups. At the end of the treatment period, the vascular reactivity of aortic rings was studied. RESULTS: In the irbesartan-treated groups, vasoconstriction induced by Nomega-nitro-l-arginine methyl ester (l-NAME) was increased and the response to phenylephrine exhibited greater potentiation in the presence of l-NAME, demonstrating a greater availability of basal nitric oxide in these groups. In addition, chronic treatment with irbesartan similarly enhanced the responsiveness of aortic rings from ovariectomized or sham-ovariectomized rats to acetylcholine and sodium nitroprusside. Incubation with indomethacin did not significantly alter acetylcholine- and sodium nitroprusside-induced relaxations in the irbesartan-treated rats. However, relaxations induced by acetylcholine and sodium nitroprusside in aortic rings from non-treated rats were significantly greater in the presence of indomethacin. CONCLUSION: Our data suggest that irbesartan enhances basal nitric oxide availability and ameliorates vascular relaxations in SHRs, by decreasing the production of cyclooxygenase dependent contracting factors in smooth muscle cells, regardless of oestrogen status. PMID- 12131534 TI - Increased contraction to noradrenaline by vasopressin in human renal arteries. AB - OBJECTIVE: Arginine vasopressin (AVP) not only acts directly on blood vessels through vasopressin V1 receptor stimulation but also may modulate adrenergic mediated responses in animal experiments. The aim of the present study was to assess whether subpressor concentrations of AVP could contribute to an abnormal adrenergic contractile response of human renal arteries. METHODS: Renal artery rings were obtained from 27 patients undergoing nephrectomy. The rings were suspended in organ bath chambers for isometric recording of tension. RESULTS: AVP (10(-10) mol/l) and the vasopressin V1 receptor agonist [Phe2, Orn8]-vasotocin (10(-10) mol/l) produced a leftward shift of the concentration-response curve to noradrenaline (half-maximal effective concentration decreased from 1.1 x 10(-6) mol/l to 3.1 x 10(-7) mol/l). The enhancement of noradrenaline-induced contractions was inhibited by the vasopressin V1 receptor antagonist d(CH2)5Tyr(Me)AVP (10-8 mol/l) and unaffected by endothelium removal or pretreatment with the inhibitor of nitric oxide (NO) synthase NG-monomethyl-l arginine (l-NMMA). The vasopressin V2 receptor agonist 1-desamino-8-D-arginine vasopressin (dDAVP) (10(-10)-10(-8) mol/l) did not modify contractile responses to noradrenaline. In the presence of the dihydropyridine calcium antagonist nifedipine (10(-6) mol/l), vasopressin failed to enhance the contractile response to noradrenaline. CONCLUSIONS: The results demonstrate that subpressor concentrations of vasopressin potentiate the contractile effects of noradrenaline without intervention of the NO system. The effects appear to be mediated by vasopressin V1 receptor stimulation, which brings about an increase in calcium entry through dihydropyridine-sensitive calcium channels. PMID- 12131535 TI - Tempol selectively attenuates angiotensin II evoked vasoconstrictor responses in spontaneously hypertensive rats. AB - OBJECTIVE: To assess whether superoxide anions mediate vasoconstrictor responses to agonists in blood vessels of spontaneously hypertensive rats (SHRs). METHODS: The effect of the superoxide dismutase mimetic, 4-hydroxy-2,2,6,6-tetramethyl piperidinoxyl (tempol), on responses to angiotensin II (Ang II), endothelin-1, phenylephrine and potassium chloride was determined in aortic rings and perfused mesenteric vascular beds (MVB) of adult male rats of the Sprague-Dawley, Wistar Kyoto (WKY) and spontaneously hypertensive (SHR) strains. The effect of tempol on Ang II-evoked superoxide production was assessed in aortic rings. RESULTS: There were no differences in the maximum tension (Emax) attained in response to agonists, but the negative logarithm of the concentration required to produce 50% of the maximal response (EC50) for Ang II was lower (P < 0.05) in aortic rings of SHRs. In the MVBs of SHRs, the Emax but not the EC50 values attained in response to Ang II, endothelin-1 and phenylephrine were greater. Tempol significantly and selectively reduced the Emax of Ang II in both aorta and MVB preparations with intact endothelium. The reduction in Emax attained in response to Ang II was more pronounced in SHRs (P < 0.01) than in WKY rats (P < 0.05) or Sprague-Dawley rats (P < 0.05). The inhibitory effect of tempol was absent when a nitric oxide synthase inhibitor was included or endothelium was denuded. A significant increase in lucigenin chemiluminescence evoked by Ang II in both intact and endothelium-denuded aortic rings of SHRs was abolished when tempol was included in the buffer. CONCLUSIONS: These data suggest that increased superoxide anions mediate vasoconstrictor responses to Ang II, but not to other agonists, in an endothelium-dependent manner, by quenching vasodilatory mediator, nitric oxide. This may account for the exaggerated vasoconstrictor responses to Ang II in SHRs. PMID- 12131536 TI - Collagen I and III mRNA gene expression and cell growth potential of skin fibroblasts in patients with essential hypertension. AB - OBJECTIVES: Despite the claimed disregulation of extracellular matrix synthesis and the increased proliferation rate of different cell types in experimental models of hypertension, very few data are available on collagen synthesis and the proliferation rate of fibroblasts in essential hypertensive patients. DESIGN: We measured collagen I, collagen III, histone H3 mRNA gene expression, collagen protein concentration and thymidine incorporation in fibroblasts from 17 essential hypertensive patients (EH) and 13 healthy normotensive control subjects (NC). METHODS: A Northern blot analysis was performed on fibroblasts in culture obtained from skin biopsies. Collagen protein concentration and DNA synthesis were measured by means of incorporation of tritiated proline and tritiated thymidine, respectively. RESULTS: In cultivated fibroblasts from hypertensives, the expression of collagen III mRNA after addition of fetal calf serum was significantly increased in comparison with that of normotensive-derived cells. After addition of fetal calf serum, collagen protein was statistically increased in cultures from EH patients as compared to NC. In hypertensives, the expression of histone H3 mRNA as well as tritiated thymidine incorporation were both increased as compared to normotensives. CONCLUSIONS: Our data suggest that cultivated fibroblasts from essential hypertensive patients are characterized by an increased expression of type III collagen mRNA and collagen protein synthesis in response to fetal serum, as compared to normotensive-derived cells. Cells from hypertensives are characterized by an increased rate of proliferation after addition of fetal serum, as ascertained by increased thymidine incorporation and increased histone H3 mRNA gene expression, as compared to normotensive-derived cells. This phenotype could be genetically determined and may have an important role in the pathogenesis of essential hypertension. PMID- 12131537 TI - Impaired skeletal muscle and skin microcirculatory function in human obesity. AB - BACKGROUND: Obesity is associated with exaggerated blood pressure and systemic vascular resistance responses to mental stress. OBJECTIVE: To test the hypothesis that skin and muscle microvascular dilatation in response to mental stress is blunted in obesity. DESIGN AND METHODS: Blood pressure, heart rate and forearm and skin blood flow responses to mental stress were compared in 23 obese and 23 age- and sex-matched lean normotensive individuals. RESULTS: Blood pressure was almost identical in both obese (mean 94 +/- 1 mmHg) and lean (93 +/- 2 mmHg) individuals. The increase in blood pressure during mental stress was similar in obese and lean individuals (2.0 +/- 0.9% compared with 3.1 +/- 4.0%; P = 0.8). Forearm vascular resistance decreased during mental stress in both groups, but this decrease was significantly blunted in obese individuals compared with controls (decreases of 14 +/- 4% and 26 +/- 3%; P < 0.01). Skin microcirculatory dilatation was also significantly blunted in obese individuals compared with controls (decreases of 5 +/- 2 and 17 +/- 4%; P = 0.02). CONCLUSIONS: Normotensive obese individuals exhibit markedly impaired muscle and skin microcirculatory responses to mental stress. The increased propensity of obese individuals to develop hypertension under conditions of chronic psychosocial stress may underlie obesity-related hypertension and cardiovascular disease. PMID- 12131538 TI - Prevention of hypertension, hyperglycemia and vascular oxidative stress by aspirin treatment in chronically glucose-fed rats. AB - OBJECTIVES: To examine whether an in vivo chronic treatment with aspirin could prevent insulin resistance, oxidative stress and blood pressure elevation associated with high glucose feeding in rats. METHODS: Sprague-Dawley rats (SD) were given a normal chow diet for 3 weeks combined or not with a 10% glucose drinking solution with or without aspirin added to their drinking water, and were compared to control SD rats which received normal chow and tap water to drink for 3 weeks. Oxidative stress was evaluated by measuring superoxide anion (O2-) production in the aorta using the lucigenin-enhanced chemiluminescence method. Antioxidant reserve was assessed by measuring the activities of glutathione peroxidase (GPx) and superoxide dismutase (SOD) in the blood. Fasting blood sugar and insulin levels were measured at the end of the study. RESULTS: The systolic blood pressure (SBP), the aortic basal superoxide production, plasma levels of insulin and glucose, as well as the insulin resistance index, were all significantly higher in rats fed glucose for 3 weeks, compared to control rats. The simultaneous treatment with aspirin prevented the increase in SBP, in plasma glucose levels and in aortic O2- production, and attenuated the rise in insulin levels as well as insulin resistance in the glucose-fed rats. Positive correlations between aortic O2- production and SBP, as well as between insulin resistance and SBP or between O2- production and insulin resistance, were found in control, glucose-fed and aspirin-treated, glucose-fed rats. The activities of GPx and SOD in the erythrocytes did not differ in the three groups. An increase in plasma SOD activity was observed in glucose-fed rats. CONCLUSIONS: Chronic in vivo treatment with aspirin prevented the development of hypertension and reduced insulin resistance significantly in chronically glucose-fed rats. Aspirin seems to produce these effects through its antioxidative properties, since it was found to prevent the increase in aortic O2- production observed in chronically glucose fed rats. PMID- 12131539 TI - Brief losartan treatment in young spontaneously hypertensive rats abates long term blood pressure elevation by effects on renal vascular structure. AB - OBJECTIVE: We studied the importance of regional vascular structural changes for the long-term antihypertensive effect of brief angiotensin II receptor blockade with losartan in young spontaneously hypertensive rats (SHRs). DESIGN/METHODS: SHRs were treated from 3 to 8 weeks of age with losartan (SHRLos, 30 mg/kg per day in drinking water) or vehicle (SHRCon). Mean arterial blood pressure (MAP) was measured using a telemetric technique from 12 to 25 weeks of age. Indices of vascular structure in the renal and hindquarter limb (HQ) were assessed using a haemodynamic perfusion technique at 13-15 weeks of age. RESULTS: MAP in SHRLos was reduced by 20-30 mmHg throughout the study (P < 0.001) and left ventricular weights were reduced (P < 0.05). The slope of the pressure/flow relationship was significantly changed (P < 0.001) in both kidneys and HQ vascular beds, suggesting greater average lumen dimensions in SHRLos. Pressure-glomerular filtration rate (GFR) curves of SHRLos kidneys were shifted to the left (P < 0.001), suggesting that the reduction in renal vascular resistance was predominantly preglomerular. The changes in structural indices of the heart and HQ closely followed the reduction in MAP. However, resistance at maximal dilatation in SHRLos kidneys was changed out of proportion to the lowering in MAP (P < 0.01). CONCLUSIONS: Brief losartan treatment in young SHRs reduces long-term MAP. The reduced MAP is associated with higher average renal and skeletal muscle vascular dimensions at maximal dilatation, predominantly in the pre-capillary vasculature. The reduction in vascular resistance of the kidney appears to be out of proportion to the reduction in MAP and it may be speculated that this is of primary importance in the long-term hypotensive effect of brief angiotensin II antagonism in SHRs. PMID- 12131540 TI - Safety and feasibility of dobutamine and dipyridamole stress echocardiography in hypertensive patients. AB - OBJECTIVES: To establish whether safety and feasibility of dobutamine and dipyridamole stress echocardiography are affected by history of hypertension. METHODS: Data on 2200 consecutive pharmacologic stress echocardiography (959 dobutamine and 1241 dipyridamole) performed between October 1990 and February 2001 at a single cardiology centre, were analysed. RESULTS: There were two complications (1/480 tests) during dobutamine (one sustained ventricular tachycardia and one severe asthmatic attack following antidote administration) and two (1/620 tests) during dipyridamole (one non-Q wave myocardial infarction and one sustained ventricular tachycardia) stress. Complications or limiting side effects were observed in 83/959 patients (48/430 hypertensives and 35/529 normotensives) with dobutamine and in 34/1241 patients (17/571 hypertensives and 17/670 normotensives) with dipyridamole stress. Therefore, the overall feasibility was 88.8% in hypertensives and 93.4% in normotensives (P = 0.013) for dobutamine, and 97% in hypertensives and 97.5% in normotensives (P = 0.64) for dipyridamole. Dipyridamole was significantly more feasible than dobutamine in both hypertensive (P < 0.0001) and normotensive (P = 0.0006) subjects. Logistic regression analysis failed to identify clinical or echocardiographic predictors of adverse reactions with dipyridamole, while history of hypertension [odds ratio (OR) = 1.8, 95% confidence interval (CI) 1.1-2.8, P = 0.0138] was the only independent predictor of cumulative adverse reactions with dobutamine stress. In addition, history of hypertension (OR = 3.2, 95% CI, 1.2-8.5, P = 0.0166), resting wall motion abnormalities (OR = 1.8, 95% CI, 1.1-3.1, P = 0.0282), and age over 70 years (OR = 4.8, 95% CI, 1.5-15.3, P = 0.0087) predicted hypertensive response, ventricular tachycardia, and atrial fibrillation, respectively. No covariate was associated with hypotensive response during dobutamine test. CONCLUSIONS: Dipyridamole has a slightly better safety profile and significantly higher feasibility than dobutamine stress both in hypertensives and in normotensives. In addition, the history of systemic hypertension is an independent predictor of cumulative adverse effects during dobutamine but not during dipyridamole stress. PMID- 12131541 TI - Left ventricular remodeling impairs coronary flow reserve in hypertensive patients. AB - BACKGROUND: In arterial hypertension, changes in both left ventricular mass and geometry may occur. Concentric left ventricular remodeling (i.e. an increased wall thickness relative to end diastolic diameter) has been implicated as an independent cardiovascular risk factor in hypertensive patients. The influence of concentric remodeling on the coronary microcirculation is not known. OBJECTIVE: To investigate the impact of left ventricular geometry on coronary flow reserve in patients with arterial hypertension and angiographically normal coronary arteries. METHODS: Following exclusion of coronary artery disease by cardiac catheterization, coronary flow reserve (dipyridamole, 0.5 mg/kg body weight intravenously; argon gas-chromatographic method) was measured in 49 patients with arterial hypertension and in six age-matched controls. Hypertensive patients were grouped by echocardiographic findings according to left ventricular mass and relative left ventricular wall thickness (i.e. left ventricular posterior wall plus septal thickness divided by end diastolic diameter): seven patients had normal left ventricular mass and geometry, 19 had eccentric hypertrophy (i.e. normal relative wall thickness but increased mass), concentric remodeling (i.e. normal mass but increased relative wall thickness) was present in nine patients, and 14 patients had concentric hypertrophy. RESULTS: There was a marked reduction in coronary flow reserve in all hypertensive groups as compared with control values (4.2 +/- 0.5). Within the hypertensive subgroups, the coronary flow reserve was differentially reduced in the following rank order: concentric remodeling (2.0 +/- 0.7) approximately concentric hypertrophy (2.3 +/- 0.8) < eccentric hypertrophy (2.9 +/- 0.6) mu normal geometry (2.7 +/- 0.4). Multi factorial regression analysis showed that the relative wall thickness but not left ventricular mass was independently linked to the coronary flow reserve. CONCLUSIONS: Concentric left ventricular remodeling is an independent predictor of the coronary flow reserve in hypertensive patients with chest pain and normal coronary angiogram. The impairment of the coronary microcirculation may contribute to the excess cardiovascular event rate associated with hypertensive concentric left ventricular remodeling. PMID- 12131542 TI - Structural changes in small resistance arteries and left ventricular geometry in patients with primary and secondary hypertension. AB - OBJECTIVE: To prospectively evaluate the interrelationships between left ventricular (LV) geometry and structural characteristics of the vessel wall in small resistance arteries in patients with consecutive primary and secondary hypertension. METHODS: In 14 patients with phaeochromocytoma, 12 with primary aldosteronism, 25 with renovascular, 25 with essential hypertension and 12 normotensive controls, an echocardiographic study for the measurement of LV mass index and relative wall thickness (RWT) was performed. Morphological characteristics of small resistance arteries (relaxed diameter < 300 microm) were directly evaluated by a micromyographic technique. RESULTS: A total of 25 patients had normal LV mass and geometry, 28 patients had normal RWT (< 0.45) and 23 patients had a RWT >or= 0.45; all normotensive subjects had normal LV mass and geometry. Media to lumen ratio (M/L) in subcutaneous small arteries was greater in hypertensive patients with concentric LV hypertrophy in respect to normotensives (ANOVA P = 0.01) and hypertensives with normal LV geometry (ANOVA P = 0.05). In the whole group of hypertensive patients the correlation coefficient between M/L and LV mass index was 0.33 (P < 0.05); the correlation coefficient between M/L and RWT was 0.46 (P < 0.01) and it was higher in primary aldosteronism (r = 0.67) and renovascular hypertension patients (r = 0.46). CONCLUSIONS: A close relation between morphology of subcutaneous small resistance arteries and LV geometric patterns may be observed in hypertensive patients; this relationship is more evident when the renin-angiotensin-aldosterone system is activated. PMID- 12131543 TI - Progressive hypertrophy regression with sustained pressure reduction in hypertension: the Losartan Intervention For Endpoint Reduction study. AB - OBJECTIVE: To examine the time course of left ventricular (LV) geometric response to blood pressure (BP) control during 2 years of systematic antihypertensive treatment. DESIGN: A total of 754 hypertensive patients with left ventricular hypertrophy (LVH) by Cornell voltage-duration product or Sokolow-Lyon voltage criteria on a screening electrocardiogram had their LV mass measured by echocardiogram at enrolment in the Losartan Intervention For Endpoint Reduction (LIFE) trial, and after 12 and 24 months of blinded therapy with losartan-based or atenolol-based regimens. SETTING: The LIFE trial, in which hypertensive patients with electrocardiographic LVH (Cornell voltage-duration product > 2440 mm x ms and/or Sokolow-Lyon voltage criteria SV1 + RV5-6 > 38 mm) were randomized to >or= 4 years double-blinded treatment with losartan or atenolol. PARTICIPANTS: A total of 754 LIFE participants with serial echocardiographic measurements of LV geometry. INTERVENTIONS: None. MAIN OUTCOME MEASURES: LV wall thicknesses, diameter and mass, and its indices. RESULTS: Mean systolic/diastolic BP fell from 173/95 to 150/84 mmHg after 1 year (P < 0.001) and to 148/83 mmHg at year 2 (P = not significant). Mean echocardiographic LV mass fell from 233 g at baseline to 206 g after 1 year (P < 0.001, adjusted for change in systolic BP) and to 195 g at year 2 (P < 0.001 versus year 1), with a parallel decrease in indexed LV mass [from 56.1 to 49.7 g/m2.7 (P < 0.001), to 47.1 g/m2.7 (P < 0.001 versus year 1)]. Relative wall thickness decreased from 0.41 at baseline to 0.37 at year 1 (P < 0.001), to 0.36 at year 2 (P < 0.001 versus year 1). As a result, there were serial decreases in prevalences of eccentric LVH [44 to 38%, and to 30% (P < 0.001 versus year 1)] and concentric LVH [24 to 7% (P < 0.001), to 2% (P < 0.05 versus year 1)], and increases in the proportion with normal LV geometry [22 to 50% (P < 0.001), and to 64% (P < 0.01 versus year 1)]. CONCLUSIONS: Sustained BP reduction in hypertensive patients with target organ damage causes continued decrease in echocardiographic LV mass and prevalence of anatomic LVH for at least 2 years despite only small BP decreases after the first year of blinded therapy. These data document cardiac benefit of sustained BP control and suggest that maximum LVH regression with effective antihypertensive treatment requires at least 2 years. PMID- 12131545 TI - Muscular side effects of statins. AB - Lipid lowering has been shown to be effective in preventing primary and recurrent cardiovascular events and to save life. Statins almost exclusively used for this purpose meanwhile became one of the most widely prescribed families of drugs world-wide. Myopathies--mainly not well characterized--are the major group of side effects. We here review different types of clinical appearances, localizations, symptoms and the biochemical background. The data indicate that severe muscular side effects are rare. Patients and their doctors, however, easily overlook mild ones. Myopathic symptoms without any known biochemical correlate are not rare. No general guideline exists about exact diagnosis and differential diagnosis. Strict adherence to the measures of life-style change and performance of regular exercise can even further enhance significantly these side effects. Much more research should be directed onto the pathophysiological (genetic?) background to finally evaluate possible therapeutic consequences rather than simply to withdraw or change the respective statin. PMID- 12131544 TI - Dual ACE and NEP inhibitor MDL-100,240 prevents and regresses severe angiotensin II-dependent hypertension partially through bradykinin type 2 receptor. AB - OBJECTIVE: To investigate the effects of the dual angiotensin-converting enzyme (ACE) + neutral endopeptidase (NEP) inhibitor, MDL-100,240 (MDL), on hypertension and cardiovascular damage in male heterozygous transgenic Ren2 rats. METHODS: Blood-pressure-matched 5-week-old transgenic rats were allocated to receive a placebo, MDL (40 mg/kg body weight) or ramipril (5 mg/kg body weight) for 8 weeks. During the last 4 weeks, the bradykinin B2 receptor antagonist, icatibant (0.5 mg/kg body weight), was also administered subcutaneously via osmotic minipumps to 50% of the transgenic rats receiving MDL or ramipril. We measured blood pressure, heart weight, structural changes in the aorta and small resistance mesenteric arteries, and the plasma concentrations of adrenomedullin, aldosterone, atrial natriuretic peptide and cGMP. To verify if MDL could regress long-standing hypertension and full-blown cardiovascular damage, 3-month-old transgenic rats received MDL subcutaneously (3 and 10 mg/kg body weight, osmotic minipumps) for 4 weeks. RESULTS: Compared with placebo, MDL decreased blood pressure (P < 0.001) and prevented left ventricular hypertrophy (P < 0.001), being as effective as ramipril. Hypertrophy and dilatation of the aorta and hypertrophy of the resistance arterioles were all prevented by MDL. Plasma aldosterone was decreased by MDL (P < 0.001), but not by ramipril. Icatibant blunted the decrease in blood pressure (P < 0.001), decreased cGMP concentrations and blunted the decrease in cross-sectional area of the resistance arteries in MDL-treated, but not in ramipril-treated, transgenic rats. In 3-month-old transgenic rats, MDL normalized blood pressure, regressed left ventricular hypertrophy and decreased adrenomedullin concentrations. CONCLUSIONS: The dual ACE+NEP inhibitor MDL prevented and regressed severe hypertension and cardiovascular damage, even in this model of severe angiotensin II-dependent hypertension with pronounced cardiovascular damage. Enhancement of the effects of bradykinin has a role in such favourable outcomes. PMID- 12131547 TI - Cyclosporin effect on rat aorta alpha(1)-adrenoceptors and their transduction mechanisms. AB - A possible explanation for cyclosporin-induced arterial hypertension may be its action on the adrenergic system. In spite of the controversial results reported in literature, it seems that cyclosporin changes the vascular response to noradrenaline. Therefore, after observation that two cyclosporin doses increase rat blood pressure and vascular reactivity in response to noradrenaline, the aim of this work was to study the cellular mechanisms beside the cyclosporin-induced changes in response to noradrenaline. Therefore, the cyclosporin influence on alpha(1)-adrenoceptors as well as on their transduction mechanism in smooth muscle cells was studied. Through Scatchard analysis of specific [(3)H]-prazosin binding, the alpha(1)-adrenoceptor number and related affinity were studied, before and after cyclosporin exposure. The cyclosporin influence on alpha1 adrenoceptor transduction mechanisms was also evaluated by the quantification of intracellular free calcium contents [Ca2+]i and inositol phosphate (InsP) turnover. All in vitro experiments were performed in rat aortic smooth muscle cells in culture. Results showed that both cyclosporin concentrations (10(-6) and 10(-7) M) changed alpha1-adrenoceptor number but only 10(-7) M cyclosporin increased its affinity for [(3)H]-prazosin. Compared with control cells, only 10( 7) M cyclosporin increased InsP levels. Stimulation by noradrenaline increased InsP in 10(-7) M cyclosporin-treated cells but decreased InsP in the presence of 10(-6) M cyclosporin. Both cyclosporin concentrations increased [Ca2+]i in basal conditions and after noradrenaline stimulation. The results suggest that after noradrenaline stimulation cyclosporin increases [Ca2+]i, probably through different mechanisms, depending on the cyclosporin concentration used. However, 10(-7) M cyclosporin increases alpha1-adrenoceptor affinity and their related transduction mechanisms. The higher cyclosporin concentration (10(-6) M) seems to induce downregulation of alpha1-adrenoceptors, probably by activation of protein kinase C. PMID- 12131546 TI - Chronic hydroxymethylglutaryl coenzyme a reductase inhibition and endothelial function of the normocholesterolemic rat: comparison with angiotensin-converting enzyme inhibition. AB - Hydroxymethylglutaryl coenzyme A (HMG-CoA) reductase inhibitors seem to have clinical benefits beyond those predicted by their lipid-lowering action. The objective was to evaluate the vascular effect of long-term treatment with statins on isolated rat aorta and their ability to prevent the acute toxicity of oxidized low-density lipoproteins (oxLDLs) compared with angiotensin-converting enzyme (ACE) inhibitors. Four groups of Wistar rats were treated for 5 weeks. Group 1 received pravastatin 20 mg/kg/d orally; group 2 received atorvastatin 10 mg/kg/d; group 3 received ciprofibrate 25 mg/kg/d; and group 4 served as control. Total cholesterol and triglyceride plasma levels were not altered, except in group 3, in which both parameters were decreased. The inhibitory effect of the endothelium on serotonin-induced contractions was significantly increased in group 1. A significant leftward shift of the concentration-response curves to acetylcholine (1 nM-0.1 mM) was observed in group 1 but the maximal relaxation to acetylcholine was similar in the four groups. In contrast, in the presence of human Cu2+-oxLDL (300 microg/ml, 30 min of preincubation), the maximal relaxation to acetylcholine was markedly decreased (p < 0.02) in groups 3 and 4 versus that of groups 1 and 2. No difference in superoxide accumulation was observed by the chemiluminescence technique. Cyclic guanosine monophosphate (cGMP), measured by enzyme immunoassay in aortic tissues, was increased in group 1 in the presence of superoxide dismutase. Endothelial nitric oxide synthase (eNOS) expression was not altered (Western blot and enzyme-linked immunosorbent assay). In aortas isolated from a fifth group of rats treated with an ACE inhibitor (ramipril 10 mg/kg/d for 6 weeks), similar results to those of group 1 were observed except that the eNOS abundance was significantly enhanced. Thus, long-term statin treatment upregulates the eNOS pathway and attenuates the acute toxicity of human oxLDL. In contrast to chronic ACE inhibition, the eNOS abundance is not increased. PMID- 12131548 TI - Systemic and regional hemodynamic and cardiac remodeling effects of candesartan in dilated cardiomyopathic hamsters with advanced congestive heart failure. AB - The effects of the selective angiotensin II type 1 receptor antagonist candesartan on cardiac, systemic, and regional hemodynamics and on cardiac, pulmonary, and hepatic histomorphometry were investigated in cardiomyopathic hamsters (CMHs), Bio TO-2 dilated strain, with advanced congestive heart failure (CHF). Two groups were treated orally with candesartan cilexetil at 22 or 50 mg/kg/d from 190 days of age and compared with a control group (38 animals/group). Investigations were performed at 225, 255, and 285 days of age. Left ventricle (LV) and systemic blood pressures and cardiac output and mesenteric and femoral blood flows were measured in anesthetized animals. LV cavity area, LV and right ventricle (RV) wall thickness and collagen density, and pulmonary and hepatic congestion were assessed. Compared with the control group, candesartan did not modify cardiac hemodynamics but significantly and dose dependently decreased systemic vascular resistances (on average: -23 and -32% after 22 and 50 mg/kg, respectively) and increased stroke volume (+32 and +42%) and cardiac output (+27 and +34%). Candesartan did not modify mesenteric vascular resistances and blood flow but significantly and dose-dependently decreased femoral vascular resistances (-19 and -33%) and increased femoral blood flow (+33 and +43%). Candesartan significantly decreased LV cavity area (-14 and -8%) and LV (-15 and -31%) and RV (-16 and -24%) collagen density but did not modify LV and RV wall thickness. Candesartan decreased pulmonary congestion at 255 and 285 days of age but did not modify hepatic congestion. In CMHs with advanced CHF, candesartan cilexetil improves systemic and femoral hemodynamics, partly reverses cardiac remodeling, and decreases pulmonary congestion. PMID- 12131550 TI - Nitric oxide in responses of regional kidney blood flow to vasoactive agents in anesthetized rabbits. AB - To determine whether differential release of nitric oxide underlies the diversity of regional kidney blood flow responses to vasoactive agents, this study examined how nitric oxide synthase blockade with IV N(G)-nitro-L-arginine (L-NNA), and also IV L-NNA plus co-infusion of glyceryl trinitrate, affected responses to renal arterial boluses and infusions of vasoactive agents. L-NNA, but not vehicle, or L-NNA plus glyceryl trinitrate, increased mean arterial pressure (35%) and reduced renal blood flow (20%), cortical perfusion (11%), and medullary perfusion (54%). L-NNA plus glyceryl trinitrate, but not L-NNA alone, blunted renal vasodilatation in response to boluses of bradykinin and acetylcholine, abolished increased medullary perfusion after bolus angiotensin II, and enhanced reductions in medullary perfusion, and to a lesser extent those in renal blood flow and cortical perfusion, during norepinephrine infusion. Neither L-NNA, nor L NNA plus glyceryl trinitrate, affected responses to infusions of angiotensin II, [Phe(2),Ile(3),Orn(8)]-vasopressin, or endothelin-1. The data indicate roles for nitric oxide in angiotensin II-induced increases in medullary perfusion and in protecting medullary perfusion from norepinephrine-induced vasoconstriction. However, differential engagement of nitric oxide synthase cannot completely account for the diversity of responses of regional kidney perfusion to vasoactive agents. Effects of nitric oxide synthase blockade on renal vascular responses to vasoactive agents were revealed only when glyceryl trinitrate was co-infused to restore resting nitrergic vasodilator tone. This may reflect interactions between nitric oxide and other vasodilator mediators, in modulating renal hemodynamic responses to vasoactive agents. PMID- 12131549 TI - Nitroglycerin tolerance: different mechanisms in vascular segments with or without intact endothelial function. AB - In vivo tolerance to nitroglycerin seems to be induced by an increase in vascular superoxide anion levels. In rabbits with normal endothelial function, in vivo induced tolerance is functionally reversed by ex vivo removal of the endothelium, probably due to a reduction in superoxide anion levels. However, the impact of in vivo endothelial dysfunction on tolerance development has not been examined. This study investigated how in vivo endothelium denudation affects the development of in vivo nitroglycerin tolerance. The effect of in vivo endothelium denudation was examined ex vivo (myograph experiments) after prolonged continuous nitroglycerin infusion in a conscious rat model. The vascular reactivity to nitroglycerin was studied in vivo in endothelium-denuded and corresponding endothelium-intact arteries. The results show that in vivo endothelium denudation does not affect the degree of tolerance development but significantly alters the effect of interventions targeted to inhibit tolerance development. In endothelium-intact vessels, superoxide dismutase and the angiotensin II receptor blocker losartan significantly inhibited tolerance-inducing properties of the prolonged nitroglycerin infusion (E[max, nitroglycerin] response in % of normal controls: nitroglycerin tolerant 70%, superoxide dismutase 93%, losartan 99%). This effect was absent in in vivo endothelium-denuded segments (nitroglycerin tolerant 57%, superoxide dismutase 72%, losartan 60%). These findings suggest that interventions against in vivo tolerance development, within the same animal, may elicit different results depending on the presence or absence of an in vivo dysfunctional endothelium. PMID- 12131551 TI - Altered effect of cyclopiazonic acid on endothelium-dependent relaxation in femoral arteries from hypertensive rats. AB - The function of endoplasmic reticulum in hypertensive vascular endothelium has not been intensively studied. The current study was designed to investigate a role of intracellular Ca2+ stores in endothelium-dependent relaxations to acetylcholine using femoral arteries obtained from Wistar-Kyoto (WKYs) and spontaneously hypertensive rats (SHRs). Rings were prepared from the femoral arteries and changes in isometric tension were recorded. Endothelium-dependent relaxations induced by acetylcholine in rings contracted with serotonin were identical in WKYs and SHRs. Cyclopiazonic acid (CPA) inhibited the relaxation in SHRs but not in WKYs. In WKYs, acetylcholine evoked smaller relaxations in rings contracted with KCl than in those contracted with serotonin, whereas in SHRs the relaxation was not affected by the contractile agonists used. The relaxation in rings contracted with KCl was abolished by Nw-nitro-l-arginine methyl ester (l NAME) and was reduced by CPA to a similar extent in both strains. In rings contracted with serotonin, l-NAME abolished the relaxation in SHRs, but the inhibitor only partially reduced the relaxation in WKYs. CPA did not alter the relaxation in the presence of l-NAME. Endothelium-independent relaxations to sodium nitroprusside were not affected by CPA. These results suggest that acetylcholine relaxes rat femoral arteries by releasing both nitric oxide and endothelium-derived hyperpolarizing factor (EDHF). In SHRs, the relaxation is preserved, but the release of EDHF is absent. CPA inhibits the relaxation mediated by nitric oxide, but not EDHF and, thus, inhibits the relaxation in SHRs but not in WKYs. Functional alteration of endoplasmic reticulum in the hypertensive endothelium cannot be detected. PMID- 12131552 TI - Time-dependent cardioprotection with calcium antagonism and experimental studies with clevidipine in ischemic-reperfused pig hearts: part I. AB - Clevidipine is a new ultrashort-acting dihydropyridine calcium antagonist developed for blood pressure regulation during cardiac surgery. When given locally to the ischemic and reperfused myocardium, clevidipine exerts a cardioprotective effect that varies depending on the timing of administration during ischemia. The current study explored the pharmacokinetics of clevidipine when administered locally into the coronary circulation. Pentobarbital anesthetized pigs were randomly divided into four groups, of which three were subjected to myocardial ischemia through ligation of the left anterior descending coronary artery (LAD) for 15, 35, or 45 minutes (n = 4 in each group). The fourth group (n = 4) was not subjected to LAD occlusion and acted as nonischemic controls. Clevidipine (0.3 nmol/kg per minute) was infused over a period of 5 minutes through a catheter distal to the LAD ligation in the ischemic pigs, or at the corresponding site of the nonligated LAD in the nonischemic pigs. Following release of the LAD ligation, or in the nonligated group following the drug infusion, the pigs were subjected to 60 minutes of reperfusion. Simultaneous blood samples were obtained for analysis of clevidipine from the femoral artery and the coronary vein during reperfusion and during drug infusion in the nonischemic pigs. Blood samples for estimating the in vitro hydrolysis rate both in whole blood and in plasma were also obtained. In nonischemic hearts, clevidipine reached a steady state level in the coronary venous blood of about 30 nM during the infusion. The concentration declined almost to the detection limit (1 nM) within 3 minutes of the end of infusion. The mean blood clearance of clevidipine was calculated to be 0.17 l/min per kilogram, and the estimated half life was 0.5 minute. In animals subjected to different periods of ischemia, very low levels of clevidipine were detected in the coronary venous blood only during the first 2 minutes of reperfusion. There were no detectable levels in the arterial blood at any time. Blood concentration profiles of clevidipine did not differ with the length of myocardial ischemia. The in vitro half-life in pig blood was 13 minutes, and the corresponding half-life in plasma was 111 minutes. At a dose known to exert cardioprotection when given as an intracoronary injection, the systemic concentration of clevidipine does not reach pharmacologically active levels. Clevidipine has an ultrashort blood half-life, and ischemic duration of up to 45 minutes does not seem to change the cardiac metabolism of this drug. Thus, it represents a pharmacological tool well suited for the study of time windows in cardioprotection. Moreover, considering the possibility of exerting myocardioprotection without any systemic effects, it could be an interesting compound to test in a clinical setting. PMID- 12131553 TI - Torsadegenic action of the antipsychotic drug sulpiride assessed using in vivo canine models. AB - A patient had QT prolongation and syncope after starting sulpiride therapy. The present experiments were performed to clarify the causal link among the sulpiride administration, QT prolongation, and the onset of torsade de pointes. Two in vivo models were used: halothane-anesthetized dogs and chronic atrioventricular (AV) block dogs. In the halothane-anesthetized animals (n = 6), sulpiride (2 and 20 mg/kg intravenously) decreased total peripheral resistance and increased heart rate, cardiac output, and ventricular contractility concomitantly. A transient attenuation of these effects occurred soon after the high-dose administration. No significant change was detected in left ventricle preload and depolarization, but repolarization and effective refractory period were significantly prolonged after high-dose administration. The extent of changes was greater in repolarization than in refractoriness, indicating prolongation of the final repolarization phase (electrically vulnerable period). In the chronic AV block animals (n = 4), onset of torsades de pointes with marked QT prolongation was demonstrated after the administration of 60 and 120 mg/kg orally of sulpiride. These results suggest that the QT prolongation and the change in the final repolarization phase with increased sympathetic tone may be the mechanisms responsible for the arrhythmogenic effect of sulpiride. Thus, caution should be paid with the use of sulpiride in patients at risk for elevated plasma concentrations and having preexisting susceptibility to QT prolongation. PMID- 12131554 TI - Isoproterenol and angiotensin I-converting enzyme in lung, left ventricle, and plasma during myocardial hypertrophy and fibrosis. AB - This study investigated whether long-term administration of isoproterenol (ISO) induces differential expression of angiotensin-converting enzyme (ACE) in lung, plasma, and left ventricle (LV) during development of left ventricular hypertrophy (LVH) and myocardial fibrosis. Male Sprague-Dawley rats (n = 7-9 per group) were treated with isoproterenol (ISO) 5 mg/kg per day for 10 days or saline and examined at 1, 15, and 33 days after the last injection. ISO stimulated the development of left ventricular hypertrophy (LVH); relative LV weight (mg LV 100/body weight), LV protein content, and LV beta-myosin heavy chain levels increased at day 1. LVH regressed at days 15 and 33. ISO also increased myocardial fibrosis (assessed by hydroxyproline content and morphometry) at days 15 and 33. There no were changes in arterial blood pressure. Long-term beta-adrenergic stimulation with ISO increased ACE expression in lung, LV, and plasma during development of LVH and myocardial fibrosis. However, time courses were markedly different. ISO stimulated a sustained increase in lung and plasma ACE activities, whereas ISO induced a high LV ACE. Plasma ACE activity paralleled lung ACE activity. LV ACE activity correlated with ACE mRNA levels and paralleled development of LVH. Our data suggest long-term beta-adrenergic stimulation induced a differential temporal expression of LV, lung, and plasma ACE in rat during development of LVH and myocardial fibrosis. PMID- 12131555 TI - Thromboxane A2 in vasomotor effects of phenylephrine, acetylcholine, and bradykinin in rat mesenteric bed. AB - In vessels, pharmacological agents displace the balance between relaxing and contracting factors. A cross-talk between those factors has been shown in some vascular beds. To examine whether NO may regulate the vascular tone by modulating prostanoid synthesis or release, we analyzed the response of resistance rat mesenteric arterial bed to vasoactive agents. Phenylephrine, bradykinin (BK), and acetylcholine (ACH) were administered in the absence or in the presence of either NO synthesis inhibition (N(G)-nitro-L-arginine methyl ester [L-NAME]), cyclo oxygenase inhibition (indomethacin), and/or a thromboxane A2 (TXA2) receptor antagonist (S18886), or a combination thereof. In the presence of L-NAME, the response to phenylephrine was markedly increased. Indomethacin limited these changes, which are attributed to TXA2 release since they were abolished by S18886 and a marked increase in TXB2 release (stable metabolite of TXA2) was found during phenylephrine infusion under NO blockade. Similarly, BK response under NO blockade was markedly attenuated with an improved response with indomethacin and a restoration of vasorelaxation with S18886. In contrast, indomethacin decreased further the response to ACH during L-NAME treatment, and TXA2 inhibition had no effect. Thus, in pathophysiological conditions where an endothelial dysfunction is present, TXA2 stimulation induced by NO release impairment may contribute to an altered response to phenylephrine or BK. PMID- 12131556 TI - Ca2+ entry channels involved in contractions of rat aorta induced by endothelin 1, noradrenaline, and vasopressin. AB - Endothelin-1 (ET-1) has been shown to activate three types of Ca2+ channel, namely two Ca2+-permeable nonselective cation channels (designated NSCC-1 and NSCC-2) and a store-operated Ca2+ channel (SOCC), and that these channels can be discriminated by Ca2+ channel blockers such as LOE 908 (a blocker of NSCC-1 and NSCC-2) and SK&F 96365 (a blocker of NSCC-2 and SOCC). This study pharmacologically compared Ca2+ entry channels involved in contractions of rat thoracic aorta without endothelium induced by ET-1, noradrenaline (NA), or arginine-vasopressin (AVP). These agonists-induced contractions of aortic rings without endothelium and increases in the intracellular free Ca2+ concentration ([Ca2+]i) of cultured aortic smooth muscle cells were abolished by removal of extracellular Ca2+. A blocker of L-type voltage-operated Ca2+ channel (VOCC), nifedipine had no effect on the responses to ET-1, but it suppressed the responses to NA and AVP to 70% and 65% of control responses, respectively. LOE 908 partially suppressed the nifedipine-resistant responses to ET-1 and AVP, but not those to NA. SK&F 96365 also partially suppressed the nifedipine-resistant responses to ET-1 and AVP, whereas it abolished the responses to NA. LOE 908 in combination with SK&F 96365 abolished the nifedipine-resistant responses to either of the agonists. These results show that the contraction of rat aorta involves different Ca2+ entry channel depending on agonists: (a) NSCC-1, NSCC-2, and SOCC for ET-1; (b) VOCC and SOCC for NA; and (c) VOCC, NSCC-1, NSCC-2, and SOCC for AVP. PMID- 12131557 TI - Mechanisms of inhibitory effects of cerivastatin on rat vascular smooth muscle cell growth. AB - The aim of this study was to clarify the mechanism(s) of an inhibitory effect of cerivastatin on cultured rat vascular smooth muscle cell (VSMC) growth. After being starved, cultured VSMCs were stimulated by 5% fetal bovine serum with either various concentrations of cerivastatin or 10-4 M of mevalonate. Cerivastatin dose-dependently decreased the values of [3H]-thymidine incorporation and cell numbers and the level of phosphorylated extracellular signal-regulated protein kinase 1/2. It also suppressed the level of proliferative cell nuclear antigen in a dose-dependent manner. These reductions were abolished by the addition of mevalonate. Similarly, the level of phosphorylated p38 was also decreased by cerivastatin. In contrast, cerivastatin dose-dependently activated the phosphorylation of both c-jun NH2-terminal protein kinase and activating transcription factor-2, and these activations were abolished by the addition of mevalonate. The levels of phosphorylated Akt and p70 S6 kinase as well as those of Bcl-2 were dose-dependently reduced by cerivastatin, and these reductions were abolished by the addition of mevalonate. Cerivastatin could dose-dependently elevate the levels of CPP32/caspase-3 activity and cytoplasmic histone-associated DNA fragments in VSMCs without causing cytotoxicity. These results indicate that cerivastatin suppresses cell survival and activates the apoptotic cellular signaling in VSMCs, suggesting that it could be effective for preventing the progression of restenosis after angioplasty. PMID- 12131558 TI - Heart rate in hypertensive patients treated with ACE inhibitors and long-acting dihydropyridine calcium antagonists. AB - Rapid heart rate (HR) has been reported to be a predictor of cardiovascular morbidity and mortality in hypertensive patients. The aim of this study was to evaluate the effect of ACE inhibitors and long-acting dihydropyridine calcium antagonists on clinic and ambulatory HR in patients with essential hypertension. We selected 292 hypertensive patients treated with ACE inhibitors and 198 hypertensive patients treated with dihydropyridine calcium antagonists. Groups were balanced for age, gender, body mass index, baseline blood pressure (BP) and HR, treatment duration and occupation. Patients had been submitted to clinic evaluation and noninvasive monitoring of BP and HR at baseline and during chronic therapy. Clinic and ambulatory BP were significantly and similarly reduced in patients treated with ACE inhibitors and calcium antagonists. Globally, clinic and 24-h HR were significantly reduced in patients treated with ACE inhibitors and remained unchanged in those treated with calcium antagonists. When patients were grouped according to baseline clinic HR (< 65 beats/min, 65-74 beats/min, 75 84 beats/min, and >85 beats/min), ACE inhibitors did not significantly change HR in subjects with baseline clinic HR <74 beats/min but significantly reduced clinic, 24-h, daytime and nighttime HR in those with baseline HR >75 beats/min and particularly in those with baseline HR >85 beats/min. Calcium antagonists did not significantly change clinic, 24-h, daytime and nighttime HR in various subgroups. Our study shows that ACE inhibitors reduce both clinic and ambulatory HR in hypertensive patients with faster HR, who seem to be at higher risk, and that long-acting dihydropyridine calcium antagonists do not induce significant changes in HR during chronic treatment (neither decrease nor increase). Whether this aspect is associated with a better prognostic impact of ACE inhibitors in essential hypertension should be determined in prospective studies. PMID- 12131559 TI - Anti-platelet effects of GPIIb/IIIa and P-selectin antagonism, platelet activation, and binding to neutrophils. AB - Platelet activation with GPIIb/IIIa binding to fibrinogen, aggregation and interaction with leukocytes constitute the principal mediator of thrombosis. Although the clinical benefits of GPIIb/IIIa antagonists have been documented, the relationship between their anti-platelet properties, platelet activation and binding to leukocytes is still debated. We investigated the effects of abciximab, tirofiban, roxifiban, and an anti-P-selectin blocking monoclonal antibody (Mab) on isolated human platelet aggregation using optical aggregometer, and on platelet P-selectin and GPIIb/IIIa expression, and platelet-neutrophil binding using flow cytometry. Thrombin at 0.025 U/ml induced maximal platelet aggregation (76.3 +/- 2.6%), P-selectin expression (88.5 +/- 4%), GPIIb/IIIa activation (PAC 1 binding, 86.2 +/- 8.9%) and platelet-neutrophil binding (58.0 +/- 6.4%). The GPIIb/IIIa antagonists inhibited in a concentration-dependent manner platelet aggregation (IC50 of 100 nM for abciximab and tirofiban and 50 nM for roxifiban) and PAC-1 binding, without any effect on P-selectin. None of these agents affected significantly platelet-neutrophil binding, whereas an anti-P-selectin Mab abolished this binding and amplified the effect of abciximab on platelet aggregation. These results indicate that the effects of these GPIIb/IIIa antagonists on platelet aggregation are not related to inhibition of platelet activation, as P-selectin levels and platelet-neutrophil binding remained unaffected, and highlight the participation of P-selectin with GPIIb/IIIa in platelet aggregation. PMID- 12131560 TI - Extracellular Ca2+ influx and endothelin-1-induced intracellular mitogenic cascades in rabbit internal carotid artery vascular smooth muscle cells. AB - Endothelin-1 (ET-1) has been proven to activate two types of Ca2+-permeable nonselective cation channels (designated NSCC-1 and NSCC-2) and a store-operated Ca2+ channel (SOCC) in rabbit internal carotid artery vascular smooth muscle cells (ICA VSMCs). Ca2+ influx through these channels plays an essential role for ET-1-induced mitogenesis in ICA VSMCs. The purpose of the current study was to investigate the effects of Ca2+ influx on intracellular pathways of ET-1-induced mitogenesis in ICA VSMCs using receptor-operated Ca2+ channel blockers, SK&F 96365 and LOE 908. We focused on extracellular-signal regulated kinase 1 and 2 (ERK1/2) in this context. PD 98059, an inhibitor of mitogen-activated protein kinase kinase, abolished the ET-1-induced increase in ERK1/2 activity, but only partially suppressed the mitogenesis. ERK1/2 activation by ET-1 was partially suppressed in the absence of extracellular Ca2+. Moreover, based on the sensitivity to SK&F 96365 and LOE 908, Ca2+ influx through NSCC-1, NSCC-2 and SOCC plays essential roles in the extracellular Ca2+-dependent component of ERK1/2 activity. In addition, Ca2+ influx through these channels was also involved in the PD 98059-resistant component of ET-1-induced mitogenesis. These results indicate that (1) the ET-1-induced mitogenesis involves both ERK1/2 dependent and -independent mechanisms in ICA VSMCs (2), ERK1/2 activation by ET-1 involves a Ca2+ influx-dependent cascade as well as a Ca2+ influx-independent cascade (3), Ca2+ influx through NSCC-1, NSCC-2 and SOCC has important roles in the Ca2+ influx-dependent component of ERK1/2-dependent mitogenesis, and (4) Ca2+ influx through these channels also plays important roles in mitogenic pathways downstream of ERK1/2. PMID- 12131561 TI - Antihypertensive effects of lacidipine during effort in mild to moderate hypertension. AB - The aim of this study was to evaluate the effects of chronic treatment with lacidipine on blood pressure, heart rate and double product during and immediately after physical effort in mild to moderate hypertensive patients. This was a single-center, randomized, double-blind, crossover, placebo-controlled clinical trial. Eighteen hypertensive patients (56% males, median age 53 years) were randomized to lacidipine 4 mg o.i.d. followed by placebo or to placebo followed by lacidipine 4 mg o.i.d. Lacidipine compared with placebo exerted a significant antihypertensive effect, lowering SBP and DBP both at baseline and either during or after exercise test. The average incremental changes of SBP and DBP between pre-exercise stage and maximal effort did not show any significant differences between treatments. HR during treatment with lacidipine was higher than during treatment with placebo both at rest and after exercise, but at maximal effort, HR was not different from placebo. The average values of DP at maximal effort, and during recovery, did not show any significant differences. Lacidipine 4 mg was effective in lowering blood pressure and in maintaining its antihypertensive effect throughout and after physical exercise, without enhancing double product value, which is an indirect index of myocardial oxygen consumption. PMID- 12131562 TI - The role of resistance characteristics of viral strains in the prediction of the response to antiretroviral therapy in HIV infection. AB - OBJECTIVE: To study the role of resistance characteristics of viral mutants in the prediction of virologic and immunologic response to antiretroviral therapy in HIV-infection. METHODS: This study is based on a mathematical model that generates viral and immunologic dynamics of HIV infection, taking into account drug-resistant mutants and therapy. We analyzed predictive factors of the increase in CD4 cell count and of the decrease in viral load from baseline after 6 months of HAART on a sample of 300 simulated individuals. The set of potential predictors was constituted by patients' state at initiation of therapy and by resistance characteristics of viral strains at that time. Predictive models, obtained by stepwise regression, were selected and compared using Mallows' Cp criterion. RESULTS: In addition to baseline viral load and CD4 cell count, known to influence response to therapy, baseline CD8 cell count and resistance characteristics of detectable strains are shown to improve the accuracy of the prediction. On the contrary, resistance parameters of low frequency viral mutants have no predictive value. CONCLUSIONS: Characteristics of preexisting detectable resistant mutants as determinants of virologic and immunologic response to antiretroviral therapy increase the capacity to predict the outcome of the treatment. Therefore, the use of phenotypic and genotypic testing could be crucial and should be considered for the choice of therapy. PMID- 12131563 TI - Exposure to dideoxynucleosides is reflected in lowered mitochondrial DNA in subcutaneous fat. AB - OBJECTIVES: Nucleoside reverse transcriptase inhibitors (NRTIs), particularly dideoxynucleoside analogs (ddNs), used in the treatment of HIV, inhibit mitochondrial DNA polymerase gamma in vitro. Mitochondrial DNA (mtDNA) depletion is proposed as the underlying mechanism of many of the in vivo side effects of these agents. A reliable and valid laboratory test to detect this is not yet available. The objective of this study was to correlate tissue mtDNA quantification in HIV-infected patients with exposure to nucleoside analogs. METHODS: 60 HIV-infected adults underwent detailed clinical assessment and blood and tissue sampling. Clinical and antiretroviral treatment details were correlated with results of plasma lactate assays, and real-time polymerase chain reaction quantification of mtDNA in peripheral blood mononuclear cells (PBMCs) and subcutaneous fat from the lower limb. RESULTS: Forty-nine (82%) subjects were on combination antiretroviral therapy. Of these, 33 (55%) were currently receiving one or more ddNs (stavudine, didanosine, or zalcitabine). mtDNA in subcutaneous fat was lower in subjects currently on ddNs than in those not taking ddNs (mean [log10] 2.47 vs. 2.74, p =.002). Plasma lactate was somewhat higher in subjects currently taking ddNs than those on no antiretroviral treatment (median 1.5 vs. 1.0, p =.03), but was not significantly different in either of these groups compared with subjects on other NRTIs. mtDNA in PBMCs did not vary with treatment status. CONCLUSIONS: mtDNA in subcutaneous fat was significantly reduced in patients currently taking ddNs. mtDNA in PBMCs was independent of patient exposure to NRTIs. PMID- 12131564 TI - Virologic rebound on HAART in the context of low treatment adherence is associated with a low prevalence of antiretroviral drug resistance. AB - The contributions of viral drug resistance, drug pharmacokinetics, and treatment adherence to failure of protease inhibitor (PI)-based highly active antiretroviral therapy (HAART) have been explored in isolation. We examined the interactions between these factors. Patients on their first PI-based HAART combination for >6 months with plasma HIV viral load (VL) >1000 copies/mL ("viremic" group) were compared with patients with a stable VL <50 copies/mL ("nonviremic" group). Data were collected on adherence (measured electronically), PI plasma levels (sampled randomly, at trough and twice after an observed dose), viral genotype, and phenotype. Mean adherence was significantly lower in the viremic group (84.3% versus 100.1%; p =.005). In the viremic group, substitutions in HIV protease were detected most frequently in the following positions in subjects on indinavir (IDV): L10I/V (35.7%), M46I/L (35.7%), A71T/V (35.7%), V82A (42.9%); and for subjects on nelfinavir (NFV): D30N (50.0%), V77I (56.3%), N88D (37.5%). Phenotypic resistance was detected in 20% of isolates from patients on IDV and 42% on NFV. Lower adherence was associated with detection of fewer resistance substitutions (r = 0.52; p =.005) and less phenotypic resistance (rho = 0.56, p =.005). There were no differences between the groups in pharmacokinetics. However, in viremic subjects, lower postdose NFV levels were associated with higher resistance (rho = -0.61, p =.02). PMID- 12131565 TI - Hyperlipidemia in HIV-infected children treated with protease inhibitors: relevance for cardiovascular diseases. AB - Cases of severely hypercholesterolemic HIV-infected children taking protease inhibitors (PIs) have been reported. Because high cholesterol levels (> or =15 mmol/L), as seen in homozygous familial hypercholesterolemia (FH), may lead to heart disease in childhood, the authors performed a systematic retrospective survey of all plasma lipid levels recorded for children who had received ritonavir or nelfinavir between 1995 and 2001 in Switzerland. Administration of PIs was associated with a significant increase in plasma cholesterol levels, which was more pronounced for those given ritonavir (from 3.3 +/- 0.7 mmol/L, n = 5 to 6.3 +/- 2.8 mmol/L, n = 19 [mean +/- SD]; p =.03) than for nelfinavir (from 3.0 +/- 0.7 mmol/L, n = 11 to 4.9 +/- 1.0 mmol/L, n = 30; p = <.001). Cholesterol levels exceeded 10.0 mmol/L in 3 of 49 (6%) PI-treated children and culminated at 13.8 mmol/L. Plasma cholesterol levels in PI-treated children were comparable with levels reported for heterozygous FH children but were all lower than in homozygous FH children. Because heterozygous FH patients usually develop heart disease in middle age, the authors conclude that the risk for heart disease in PI treated children is minimal. Long-term monitoring of these children, however, will be necessary. PMID- 12131566 TI - Clinical tolerance and immunologic effects after single or repeated administrations of the synthetic immunomodulator murabutide in HIV-1-infected patients. AB - Correction of the virus-induced deficits in innate immunity of HIV-infected subjects could well contribute to enhanced immune recovery and efficacious control of viral replication. The safe synthetic immunomodulator Murabutide (ISTAC Biotechnology, Lille, France) has been found to regulate the function of antigen-presenting cells and to selectively activate CD4 lymphocytes leading to dramatic suppression of HIV replication, in vitro. Therefore, as a first step toward the evaluation of the immunotherapeutic potential of Murabutide in HIV disease, we have conducted two phase 1/2 clinical trials to address the safety and the immunologic effects of Murabutide administration into HIV-infected subjects receiving antiretroviral therapy. The first study revealed that single administration of 5, 7, or 9 mg of Murabutide, to 6 patients per dose, was well tolerated. This was accompanied by a selective induction of cytokines and chemokines detectable in the serum, and the levels appeared to plateau at the 7 mg dose. The second study then evaluated the safety and biological effects of repeated administrations of 7 mg Murabutide, on 5 consecutive days, in 12 HIV-1 infected patients. A good clinical tolerance was noted throughout the study. Moreover, changes in several immune parameters, including downregulation of coreceptor expression on lymphocytes and improved lymphoproliferative responses, were detected during or/and up to 3 weeks after Murabutide administration. These encouraging results warrant further evaluation of longer periods or cycles of immunotherapy with Murabutide in HIV-infected subjects. PMID- 12131568 TI - Prenatal and postpartum zidovudine adherence among pregnant women with HIV: results of a MEMS substudy from the Perinatal Guidelines Evaluation Project. AB - Adherence to HIV treatment regimens during pregnancy may affect efforts to eliminate vertical transmission and influence the emergence of drug-resistant HIV strains that can affect maternal health and the risk of vertically-transmitted resistant strains. Study objectives were to document patterns of adherence to zidovudine (ZDV) during the perinatal period. Pregnant women with HIV who were seen at public clinics, taking ZDV, and willing to use Medication Event Monitoring Systems (MEMS) caps participated in this adherence substudy. Fifty three women were included in prenatal analyses; however, 19 women were excluded from postnatal analyses because medical records failed to confirm a postpartum maternal prescription for ZDV. Adherence to ZDV, defined as doses per day taken/prescribed during the last 3 weeks of pregnancy, was extremely low (mean = 50.0%), and declined significantly 3 weeks postpartum (mean = 34.1%) (p =.004). Clinical emphasis must be placed on enhancing adherence during and particularly after pregnancy when ZDV is continued for a mother's own care. PMID- 12131567 TI - Use of HIV protease inhibitors is associated with left ventricular morphologic changes and diastolic dysfunction. AB - HIV protease inhibitor (PI) therapy may be associated with cardiac and vascular complications. We assessed the effects of PIs on cardiac function and structure. M-mode, cross-sectional, and Doppler echocardiography were performed in 98 consecutive black adults aged 25 to 45 years with HIV infection who were receiving antiretroviral therapy. Forty-five (56.1%) took PIs (mean duration of PI use: 29.6 +/- 12.2 months). No significant differences between the PI and non PI groups were found in left ventricular (LV) systolic function and cardiac valve regurgitation. Those who took PIs had a significantly higher interventricular septum thickness (1.1 +/- 0.3 vs. 1.0 +/- 0.2 cm; p =.049), LV posterior wall thickness (1.1 +/- 0.2 vs. 1.0 +/- 0.2; p =.027), and lower ratio of early peak velocity (E wave) to late peak velocity (A wave) (E/A ratio) (1.36 +/- 0.30 vs. 1.53 +/- 0.31; p =.023) than did those who did not take PIs. Linear regression analyses indicated that posterior wall thickness, septum thickness, left atrial dimension, LV mass, and E/A ratios were significantly associated with the log transformed duration of PI therapy. Despite the proven benefits of PIs in patients with HIV infection, this report demonstrates an association between continued PI intake and LV hypertrophy that should be known and taken into consideration in the analysis of cardiac structure and function in patients with HIV infection. PMID- 12131569 TI - Correlates of plasma HIV-1 RNA viral load among HIV-1-seropositive women in Dar es Salaam, Tanzania. AB - This study was conducted to determine the predictors of plasma HIV-1 RNA viral load in HIV-1-positive pregnant women (N = 151) participating in a clinical trial in Tanzania. Viral load was measured at randomization, delivery, and approximately 7 months after delivery. The median viral load was 20,400 copies/mL at baseline, 20,216 copies/mL at delivery, and 19,100 copies/mL 7 months after delivery. The absolute CD4+ lymphocyte count had a strong negative correlation with HIV-1 RNA viral load at baseline (r = -.38), time of delivery (r = -.36), and 7 months after delivery (r = -.53). The association between CD4+ lymphocyte count and HIV-1 RNA viral load was modified by the per capita daily food expenditure in the household, although the difference in viral load became small as the food expenditure in the household increased and was marginally significant at the 75th percentile of the per capita food expenditure. The presence of malaria parasites at baseline was associated with an approximate 116% higher viral load at the three evaluation points (p =.007). Although the long-term effects of malaria on viral load are unknown, prevention of malaria among people living with HIV-1 should be given the highest priority. PMID- 12131571 TI - Females experiencing sexual and drug vulnerabilities are at elevated risk for HIV infection among youth who use injection drugs. AB - OBJECTIVES: To compare sociodemographic, drug-related, and sexual risk variables between young (13-24 years of age) and older (> or =25 years of age) injection drug users (IDUs); and to determine HIV prevalence and associated risk factors for HIV infection among young IDUs. METHODS: Data were collected through the Vancouver Injection Drug Users Study (VIDUS). To date, over 1400 Vancouver area IDUs have been enrolled and observed during follow-up. Sociodemographic, drug related, and sexual risk variables were compared between younger and older IDUs using nonparametric methods. Mantel-Haenszel and logistic regression methods were used to compare HIV-positive and HIV-negative female youth. RESULTS: Younger injectors (N = 232) were more likely to be female; work in the sex trade; report condom use; inject heroin daily; smoke crack cocaine daily; and need help injecting. HIV prevalence at baseline among the youth was 10%. HIV prevalence was associated with female gender; history of sexual abuse; engaging in survival sex; injecting heroin daily; injecting speedballs (a mixture of heroin and cocaine) daily; and having numerous lifetime sexual partners. CONCLUSION: Our data show that HIV positivity among young IDUs is concentrated among females engaged in dual sexual and drug-related risk exposure categories. Over half the HIV-positive youth were Aboriginal (a classification used by the federal government in Canada to include native peoples of all ethnic groups). Targeted interventions that take into account sexual and drug risk for young female and Aboriginal peoples are urgently needed. PMID- 12131570 TI - Assessment of adherence to HIV protease inhibitors: comparison and combination of various methods, including MEMS (electronic monitoring), patient and nurse report, and therapeutic drug monitoring. AB - BACKGROUND: Adherence to protease inhibitor-containing antiretroviral therapy is crucial, but difficult to measure. OBJECTIVE: To compare and combine various methods of measuring adherence to the strict protease inhibitor-containing regimens. METHODS: The following methods were used: medication event monitoring system (MEMS) caps (electronic monitoring), therapeutic drug monitoring, pill count, pharmacy refill data, questionnaires, diaries (for registration of food patterns and special events related to the use of MEMS), adherence assessment by the physician and clinical nurse specialist, and in-depth interviews. In addition, ultrasensitive viral load and resistance testing was performed. RESULTS: Twenty-eight patients were included; data could be evaluated in 26. According to MEMS data, 25% of the patients took fewer than 95% of all doses, and two thirds of the patients took fewer than 95% of the doses on time. Only 43% of the patients showed good adherence with food restrictions. Methods that showed significant correlations with MEMS results were patients' self-reported adherence; therapeutic drug monitoring, indicating plasma levels outside predefined ranges; and estimation of adherence by a clinical nurse specialist, especially by in-depth interview. CONCLUSION: Diary-corrected MEMS data gave a detailed insight into patients' adherence patterns. Patients' self-report and therapeutic drug monitoring were significantly correlated with the MEMS data, and the clinical nurse specialist may also play a role in identifying patients who are imperfectly adherent. PMID- 12131573 TI - Predicting death from HIV/AIDS: a case-control study from Florida public HIV/AIDS clinics. AB - BACKGROUND: After markedly decreasing for 3 years, HIV/AIDS mortality declined only slightly in 1999. METHODS: The authors conducted a case-control study in four Florida urban public health HIV clinics to evaluate modifiable factors associated with HIV/AIDS mortality in a non-research setting. Structured chart review was conducted for 120 case-patients who died in 1999 and for 240 randomly selected control-patients. Risk factors associated with death in univariate analysis were entered into three conceptually related, matched logistic regression models. RESULTS: In the final multivariate model, homelessness (adjusted odds ratio [AOR], 9.98; 95% confidence interval [CI], 2.34-42.5), Medicaid insurance (AOR, 3.10; 95% CI, 1.43-6.74), having a documented adherence problem (AOR, 3.50; 95% CI, 1.64-7.47), injection drug use (AOR, 2.46; 95% CI, 1.11-5.43), non-specific liver failure (AOR, 76.9; 95% CI, 6.79-870.9), interrupted highly active antiretroviral therapy (HAART) secondary to side effects (AOR, 4.00; 95% CI, 1.46-10.9), and not receiving HAART (AOR, 2.62; 95% CI, 1.03-6.68) were independent predictors of mortality. CONCLUSIONS: In addition to medical and clinical indicators, several sociobehavioral-demographic factors remained important throughout the multivariate analysis. Improvement in care should include a focus on social circumstances of infected people. Special attention to the homeless, those with adherence problems, and those with liver disease is clearly indicated. PMID- 12131572 TI - Increased prevalence of HTLV-1 among HIV-2-infected women but not HIV-2-infected men in rural Guinea-Bissau. AB - OBJECTIVE: To assess the prevalence of HTLV infection and its association with HIV and other potential risk factors. DESIGN AND SETTING: A cross-sectional survey and a case-control study in a rural community in Guinea-Bissau. METHODS: A total of 2770 people were included in an HIV and HTLV seroepidemiologic survey. Three hundred of these participants were selected for a case-control study on HIV 2. Sera from both studies were tested for HTLV. RESULTS: In all, 2501 and 298 subjects in the survey and case-control study, respectively, were tested for HTLV. Overall HTLV-1 prevalence was 5.2% and it was higher in women (odds ratio [OR], 1.36; confidence interval [CI], 0.92-2.02). Apart from an infected spouse, no significant risk factors could be identified for men. In women, HIV-2 infection (adjusted OR, 5.58; CI, 3.09-10.1), having an infected spouse, and area of residence were significantly associated with HTLV-1 infection. The association between HTLV-1 and HIV-2 was significantly different for men and women (test of interaction, p =.002). CONCLUSIONS: In women, the most important determinant of HTLV-1 seropositivity was HIV-2 infection. Because the pattern was significantly different for men and women, common sexual risk factors may not be sufficient to explain the co-occurrence of HIV-2 and HTLV-1 in women. These observations may have implications in geographic areas where both types of retroviruses are prevalent. PMID- 12131574 TI - Decline in prevalence of HIV-1 infection and syphilis among young women attending antenatal care clinics in Addis Ababa, Ethiopia: results from sentinel surveillance, 1995-2001. AB - From 1995 to 2001, five rounds of sentinel surveillance were carried out for young women attending antenatal care clinics at four health centers in Addis Ababa, the capital city of Ethiopia, to monitor trends in the prevalence of HIV infection and syphilis. Serum samples were tested for antibodies to HIV (enzyme linked immunosorbent assay and Western blotting) and antibodies to Treponema pallidum (T. pallidum hemagglutination assay and rapid plasma reagin test). Prevalence ratios for an increase in one calendar year were estimated using log binomial models. Between 1995 and 2001, the prevalence of HIV infection among young women (age range, 15-24 years) attending antenatal care clinics in inner city health centers declined from 24.2% to 15.1% (prevalence ratio for an increase in one calendar year, 0.91; 95% confidence interval, 0.87-0.95). No change was observed for older age groups or in outer city health centers. The decline in the prevalence of active syphilis (T. pallidum hemagglutination assay and rapid plasma reagin testing positive for antibodies to T. pallidum) was more pronounced among and also restricted to the young age groups (age range, 15-24 years) in the inner city (from 7.6% in 1995 to 1.3% in 2001; prevalence ratio, 0.69; 95% confidence interval, 0.59-0.80). The declining trends in the prevalence of HIV infection and syphilis among young women attending antenatal care clinics in the inner city are encouraging, but these findings require confirmation in future years and for other population groups. PMID- 12131575 TI - The lack of therapeutic benefit of adding hydroxyurea to highly active antiretroviral therapy (HAART). PMID- 12131576 TI - Highly active antiretroviral therapy (HAART) modifies the incidence and outcome of visceral leishmaniasis in HIV-infected patients. PMID- 12131577 TI - Clinical impact of HTLV-1 infection in Spain: implications for public health and mandatory screening. PMID- 12131578 TI - Neuroophthalmology and trauma. PMID- 12131580 TI - Management of traumatic cataracts. PMID- 12131579 TI - Corneal wound healing: cytokines and extracellular matrix proteins. PMID- 12131581 TI - Trauma after refractive surgery. PMID- 12131582 TI - Management of astigmatism after corneal trauma. PMID- 12131583 TI - Traumatic hyphema. PMID- 12131584 TI - Angle recession glaucoma. PMID- 12131585 TI - Closed-globe contusion injuries of the posterior segment. PMID- 12131586 TI - Management of subretinal hemorrhage. PMID- 12131587 TI - Traumatic macular holes. PMID- 12131588 TI - Anterior and posterior segment intraocular foreign bodies. PMID- 12131589 TI - Ultrasonography in the traumatized eye: intraocular foreign body versus artifact. PMID- 12131591 TI - Ocular injuries in shaken baby syndrome. PMID- 12131590 TI - Proliferative vitreoretinopathy after trauma. PMID- 12131592 TI - Traumatic ruptured globe injuries in children. PMID- 12131593 TI - Combat eye trauma: intraocular foreign body injuries during the Iran-Iraq war (1980-1988). PMID- 12131594 TI - Sympathetic ophthalmia. PMID- 12131595 TI - Management of complex eyelid lacerations. PMID- 12131596 TI - Adaptation to monocular vision. PMID- 12131598 TI - Options for pharmacological management of obesity in patients treated with atypical antipsychotics. AB - Obesity is associated with considerable morbidity and decreased life expectancy. Weight gain is a commonly encountered problem associated with antipsychotic treatment. We reviewed the literature regarding the mechanisms of weight gain in response to these agents and eight substances implicated as potential obesity prevention or treatment: orlistat, sibutramine, fluoxetine, topiramate, amantadine, nizatidine and cimetidine, and metformin. Weight gain in response to antipsychotic treatment may be mediated through serotonergic, dopaminergic, adrenergic, cholinergic, histaminergic and glutaminergic receptors. Sex hormone dysregulation and altered insulin sensitivity have also been implicated. Two compounds, orlistat and sibutramine, have been shown to help prevent weight gain following a hypocaloric diet, but orlistat requires compliance with a fat-reduced diet, and sibutramine is unsuitable for patients taking serotonergic agents. The weight reducing effect of fluoxetine, even in conjunction with a hypocaloric diet, is only transient. Topiramate, amantadine and metformin may have adverse side-effects potentially outweighing the weight reducing potential. The effectiveness of cimetidine and nizatedine remains unclear. The hazards of these agents in a psychiatric population are discussed. It is concluded that the current evidence does not support the general use of pharmacological interventions for overweight patients treated with antipsychotic medication, although individually selected patients may benefit. PMID- 12131599 TI - Moclobemide is effective and well tolerated in the long-term pharmacotherapy of social anxiety disorder with or without comorbid anxiety disorder. AB - Social phobia (social anxiety disorder) is a highly prevalent and chronic disorder that is associated with significant comorbidity and disability. Despite recent advances in the pharmacotherapy of the disorder, there is a paucity of randomized controlled trials on patients with comorbid disorders and on maintenance treatment. A randomized placebo-controlled, double-blind multi-site trial of moclobemide, a reversible inhibitor of monoamine oxidase A, was undertaken with 390 subjects. After an initial 12 weeks, there was the option of continuing for an additional 6 months of treatment. The primary efficacy parameter chosen was responder status as defined by the Clinical Global Impression scale change item. From week 4 onwards, there was a significantly higher response rate on moclobemide than on placebo. Superiority of medication over placebo was similar in patients with comorbid anxiety disorders (33% of subjects) and without, as well as in patients with different subtypes of social anxiety disorder; indeed, treatment with moclobemide rather than placebo was the strongest predictor of response. Adverse events were similar across treatment groups, and were typically mild and transient. In the extension phase, response rates remained higher in the moclobemide group, and ratings of tolerability were equally high in both groups. Thus, in a large sample of social anxiety disorder patients with and without comorbid anxiety disorders, moclobemide was both effective and well-tolerated in the short as well as long-term. These data confirm and extend previous findings on the value of moclobemide in the treatment of social anxiety disorder, and strengthen the range of therapeutic options for managing this important disorder. PMID- 12131600 TI - Social adjustment in depressed patients treated with venlafaxine and amitriptyline. AB - Social dysfunction is often associated with depressive disorders and its evaluation is an important measure of treatment outcome. The aim of this study was to assess the effect of two treatments, venlafaxine and amitriptyline, on the social functioning of depressed patients. Twenty-eight outpatients, meeting criteria for recurrent or single major depressive episodes, took part in a double blind, 8-week trial with amitriptyline or venlafaxine (maximum of 150 mg/day) and were assessed by the Self-Report Social Adjustment Scale (SAS-SR) before and at the end of treatment. The scale was also applied to a carefully selected non psychiatric sample. Both drugs showed the same efficacy on a clinical scale, but venlafaxine improved social functioning more than amitriptyline as only venlafaxine-treated patients reached SAS-SR values estimated for normal subjects. This effect might be linked to the higher rate of side-effects observed with amitriptyline. PMID- 12131601 TI - Effect of a single-dose of olanzapine on sleep in healthy females and males. AB - The effect of a single dose of 10 mg olanzapine on healthy volunteers of both sexes was examined using polysomnography and power spectral analysis. The structure and continuity of sleep were unaffected by olanzapine in both sexes. The increase in both actual sleep time and slow wave sleep in females correlated with the increase in theta power, while delta power was not significantly elevated, suggesting that theta power may be a sensitive indicator of changes in sleep. The changes in sleep had the same tendency in men, but they were not significant. The difference between the sexes could not be explained by differences in body mass index. Olanzapine affects sleep probably through 5 HT(2C) receptors. The receptor gene is located on the X-chromosome, inducing an allelic difference between the females and males. This difference may contribute to the different effects of olanzapine on sleep. Olanzapine seems to preserve the normal structure of sleep and increase the amount of slow-wave sleep, which might be of additional benefit in treatment of schizophrenia. The effective clinical dose may be lower for females than males. PMID- 12131602 TI - A placebo-controlled study of Kava kava in generalized anxiety disorder. AB - We assessed the efficacy and safety of a botanical anxiolytic, Kava kava (Piper methysticum), in treating generalized anxiety disorder (GAD). Thirty-seven adults with DSM-IV GAD were randomly assigned to 4 weeks of double-blind treatment with kava or a matching placebo. Weekly efficacy assessments [Hamilton Anxiety Scale, Hospital Anxiety and Depression Scale (HADS), Self Assessment of Resilience and Anxiety (SARA)] and safety evaluations were conducted. Improvement was observed with both treatments but no differences were found in the principal analysis. Post-hoc analyses revealed significant differences based on baseline anxiety severity, whereby kava was superior on the SARA in low anxiety and placebo was superior on the HADS and SARA in high anxiety. Both treatments were well tolerated. Although kava was not superior to placebo, it would be premature to rule it out as efficacious in GAD. PMID- 12131603 TI - Clozapine-treated subjects with treatment-resistant schizophrenia: a systematic review of experimental and observational studies. AB - Randomized clinical trials have limitations because they focus on small samples of highly selected patients. Observational studies, which follow large cohorts of typical patients receiving pharmacological treatments, should overcome some of these trial limitations and provide information that cannot be generated with clinical trials. The present study aimed to compare experimental and observational studies of clozapine-treated subjects with treatment-resistant schizophrenia. A systematic review of experimental and observational studies evaluating clozapine-treated subjects in treatment-resistant schizophrenia was carried out. We identified 50 studies that met the inclusion criteria. Less than one-third of clinical trials enrolled more than 50 patients compared to 44% of prospective and nearly 90% of retrospective studies. In addition, 78% of prospective and 89% of retrospective observational studies lasted more than 12 weeks, while the majority of trials lasted less than 8 weeks. Most clinical trials defined treatment-resistant schizophrenia according to Kane's criteria, while the majority of observational studies adopted implicit criteria. In comparison with clinical trials, observational studies provided a higher weighted mean rate of clozapine-responders and a lower weighted mean rate of clozapine dropouts. This literature survey suggests that the role of observational studies in the evaluation of medicines should be reconsidered. A new generation of observational studies should be developed to provide evidence on patient outcome in typical settings and under real-world circumstances, and on variables which may affect outcome. PMID- 12131604 TI - Do triglycerides modulate the effectiveness of clozapine? AB - We describe a case in which a patient's clinical response to clozapine appears to correlate positively with his serum triglyceride concentrations. We propose that the observed clinical response may partly be the result of the physical interaction of clozapine with the very low-density lipoproteins. We base this supposition on our previous in-vitro study showing that the plasma distribution of clozapine is significantly altered by increases in plasma triglyceride concentrations. Although this case only represents one patient, it highlights the possibility that serum lipids may be potential contributors to the clinical effectiveness of clozapine. PMID- 12131605 TI - Treatment of compulsive sexual behaviour with naltrexone and serotonin reuptake inhibitors: two case studies. AB - Although there is no universally accepted definition of compulsive sexual behaviour (CSB), the term is generally used to indicate excessive sexual behaviour or sexual cognitions that lead to subjective distress, social or occupational impairment, or legal and financial consequences. Similar to impulse control disorders, CSB is characterized by a failure to resist the impulse for sex. Opioid antagonists have been effective in treating urge-driven disorders, such as pathological gambling disorder, alcoholism, borderline personality disorder with self-injurious behaviour, cocaine abuse, mental retardation with self-injurious behaviour and eating disorders. Based upon the efficacy of opioid antagonists in treating disorders associated with urges, we hypothesized that naltrexone would reduce both the urges associated with CSB and therefore reduce the sexual behaviour. We present two case reports of individuals with CSB treated successfully with naltrexone, a novel treatment for CSB. In both cases, symptoms dramatically decreased and psychosocial functioning improved with the use of naltrexone. Although more research is needed to determine the mechanism that leads to the excessive sexual behaviour in individuals with CSB, the present case reports suggest that naltrexone may be effective in treating some cases of CSB. PMID- 12131606 TI - Infliximab improves quality of life in patients with Crohn's disease. AB - OBJECTIVE: The aim of this study was to assess the effect of infliximab on quality of life in patients with active Crohn's disease (CD) inadequately responsive to concomitant therapies. METHODS: We examined responses to the Inflammatory Bowel Disease Questionnaire (IBDQ) from patients enrolled in a previously reported, randomized, placebo-controlled study. Patients with active CD received a single intravenous infusion of either placebo or infliximab 5, 10, or 20 mg/kg. Most patients received stable doses of mesalamine, corticosteroids, azathioprine, or 6-mercaptopurine throughout the study. Changes from baseline in overall IBDQ score and individual dimensions at 4 weeks postinfusion were compared. RESULTS: Patients treated with infliximab had a significantly larger improvement in overall IBDQ score than those treated with placebo at 4 weeks (p < 0.001). Infliximab-treated patients also had larger improvements in all IBDQ dimensions: bowel (p = 0.007), social (p = 0.002), emotional (p < 0.001), and systemic (p < 0.001). A significantly larger proportion of infliximab-treated patients reported having normal or near-normal frequency of bowel movements in the past week (p < 0.001), full or a lot of energy (p = 0.019), and no or hardly any difficulty doing leisure or sports activities (p = 0.011), and being extremely or very satisfied with their personal life (p = 0.046). They also significantly differed in responses regarding fatigue, frustration, ability to work, general well-being, depression, anxiety, and anger resulting from bowel problems. CONCLUSIONS: These results indicate that infliximab significantly improved quality of life in patients with active CD, increasing their ability to work and participate in leisure activities, and decreasing feelings of fatigue, depression, and anger. PMID- 12131607 TI - Long-term evolution of disease behavior of Crohn's disease. AB - BACKGROUND: The Vienna classification of Crohn's disease (CD) distinguishes three patient subgroups according to disease behavior: stricturing, penetrating, and inflammatory. Our aim was to assess the long-term evolution of the disease behavior of CD and to determine the predictive factors and prognostic implications of this evolution. METHODS: Occurrence and predictive factors of a stricturing and/or a penetrating complication were searched for in 2,002 patients with CD studied retrospectively. In addition, the 1995-2000 disease course was assessed prospectively in a cohort of 646 patients with disease duration >5 years, classified according to their previous disease behavior. RESULTS: 1,199 patients (60%) developed a stricturing (n = 254) or a penetrating (n = 945) complication. Twenty-year actuarial rates of inflammatory, stricturing, and penetrating disease were 12, 18, and 70%, respectively. The initial location of lesions was the main determinant of the time and type of the complication. In the cohort study, year-by-year activity and therapeutic requirements did not show significant sustained differences between behavioral subgroups. CONCLUSION: Most patients with CD will eventually one day develop a stricturing or a perforating complication. Initial location determines the type of the complication. Classification of patients into a behavioral group from previous history has no impact upon activity during the following years. PMID- 12131608 TI - Doppler enhancement after intravenous levovist injection in Crohn's disease. AB - Although transabdominal bowel sonography (TABS) has been proposed as a reliable tool to assess increased bowel wall thickness (BWT), the most common sonographic pattern in patients with Crohn's disease (CD), its accuracy is limited in the diagnosis of CD. We therefore tried to assess whether color Doppler enhancement with Levovist, a galactose-based intravenous sonographic contrast agent able to enhance the arterial Doppler signal, increases TABS accuracy. Thirty-one patients with ileal CD, diagnosed by endoscopy and enteroclysis, and 20 healthy volunteers were examined with conventional TABS. Color Doppler of the intramural enteric vessels was then performed before and after intravenous injection of Levovist. Twenty-two CD patients had a BWT >4 mm, and 16 of them presented with active disease. Two of the remaining nine CD patients, all with BWT <4 mm, presented with active disease. By means of color Doppler we identified six patients with inactive disease, normal BWT, and normal basal Doppler signal intensity, who showed an enhanced Doppler signal in intramural vessels after contrast agent bolus. Four of these patients, identified only by color Doppler after Levovist injection, relapsed within 6 months. In our experience, sensitivity and specificity of TABS, integrated with additional stimulated acoustic emission mode, were 96.7% and 100%, respectively. The use of Levovist in color Doppler increases the accuracy of TABS in CD diagnosis and follow-up. PMID- 12131609 TI - Chronic colitis is associated with a reduction of mucosal alkaline sphingomyelinase activity. AB - BACKGROUND AND AIMS: The hydrolysis of sphingomyelin (SM) generates key molecules regulating cell growth. Animal cancer studies support an inhibitory role for this pathway in the malignant transformation of the colonic mucosa. The activity of a specific intestinal alkaline sphingomyelinase (SMase), which hydrolyzes SM, is reduced in colorectal tumors. In this study we measured alkaline SMase activity in patients with longstanding colitis and assessed if a reduction can be used as a marker in surveillance of high risk patients. METHODS: Alkaline SMase activity was measured in 139 colonic biopsies from 34 patients with longstanding, extensive colitis and from 11 controls. Fifteen patients had earlier diagnosis of dysplasia or DNA aneuploidy. Alkaline SMase activity was related to histologic dysplasia and DNA aneuploidy assessed by flow cytometry, patient age, and duration of disease. RESULTS: Alkaline SMase activity was significantly lower in the patient group with and without dysplasia compared with controls (p = 0.006). In biopsies, an association was not found between alkaline SMase activity, dysplasia, or DNA ploidy. However, alkaline SMase activity decreased with age both in patients and controls (p = 0.008). CONCLUSIONS: Reduction of alkaline SMase activity seen in colorectal cancer and adenomas is also present in patients with chronic colitis. It is not complementary to dysplasia or DNA-aneuploidy in the identification of high risk patients. The age-associated decrease of alkaline SMase activity seems to be a general phenomenon indicating premature senescence of the mucosa in longstanding colitis. PMID- 12131610 TI - Providing disease-related information worsens health-related quality of life in inflammatory bowel disease. AB - BACKGROUND: Patients with inflammatory bowel disease (IBD) have identified a need for more information about their disease. PURPOSE: To assess the effect of an educational intervention on health-related quality of life (HRQOL) in patients with IBD. METHODS: Consecutive ambulatory IBD patients were randomized to receive four IBD-specific educational booklets or usual care. Subjects completed two disease-specific HRQOL questionnaires-the Inflammatory Bowel Disease Questionnaire (IBDQ) (range 1-poor to 7-excellent) and the Quality Index in Crohn's and Colitis (QuICC) (range 1-excellent to 5-poor) at entry and after 2 weeks. The mean change in HRQOL scores at follow-up was compared between the education and control groups. RESULTS: 59 subjects participated, with a mean age of 40.0 +/- 11.9 years. 34 were given educational booklets and 25 received standard care. 6 patients (10%) did not complete the study. Mean IBDQ scores became significantly worse in the education group with a change of -0.17 +/- 0.49 compared with controls at +0.28 +/- 0.62 (p = 0.006). This could be explained by worsened disease activity in the education group. There was no significant change in the QuICC scores (p = 0.61). Education group patients who had not received prior educational material had improved mean IBDQ scores of +0.24 +/- 0.47 compared with education patients who had received educational material prior to this study, with a score change of -0.25 +/- 0.46 (p = 0.09). CONCLUSIONS: The addition of educational booklets to IBD patients in a tertiary center does not improve, and may worsen, short-term HRQOL. Education of newly diagnosed or less informed patients should be studied further. PMID- 12131611 TI - Are parents able to rate the symptoms and quality of life of their offspring with IBD? AB - The aim of this study was to investigate the degree of agreement between parents and their offspring with inflammatory bowel disease for the presence of symptoms and the assessment of health-related quality of life (HRQOL). Factors influencing parent-child agreement were studied. Eighty-three Children and 81 parents separately filled out a five-item symptom card and a validated generic HRQOL instrument, which assesses seven domains of HRQOL, using the child and parent form. The parent also filled out the GHQ-30, an instrument assessing nonpsychotic psychiatric disorder in the parent, and an item on marital status. Intraclass correlation coefficients and paired student t-test were used to assess the level of agreement between raters. On one domain, parents reported their children as having a worse QOL than did the children themselves (social functioning). The parents were adequate raters of objective components of their child's HRQOL (overall correlation coefficient: 0.88). However, on more subjective components, the coefficient dropped to 0.62. In 82% of the cases did parents correctly classify their child into the disease activity category the child classified him- or herself. In conclusion, agreement between parents and offspring is good for the child's symptoms, but for HRQOL assessment only when it concerns objective states. PMID- 12131612 TI - Role of appendicitis and appendectomy in the pathogenesis of ulcerative colitis: a critical review. AB - Besides a genetic predisposition, a causal role of various environmental factors has been considered in the etiology of ulcerative colitis (UC). The association between appendectomy and UC has recently been the subject of intense scrutiny in the hope that it may lead to the identification of important pathogenetic mechanisms. Published data from animal models of colitis demonstrated reduction in experimental colitis after appendectomy, especially if performed at an early age. Several epidemiological case control and cohort studies have shown a strong and consistent relationship. The metaanalysis of 17 case-controlled studies showed an overall odds ratio 0.312 (95% confidence intervals = 0.261-0.373) in favor of appendectomy (p < 0.0001). One of the two recent large cohort studies is in agreement with these results, but the other failed to confirm them. All these studies have suggested that alterations in mucosal immune responses leading to appendicitis or resulting from appendectomy may negatively affect the pathogenetic mechanisms of UC. Further investigation of the role of appendectomy in UC is expected to open new fields for basic scientific research and may lead to the improvement of our understanding for the disease pathogenesis. PMID- 12131613 TI - Adenocarcinoma of the cecum as the first manifestation of ulcerative colitis complicated by primary sclerosing cholangitis and endomyocardial fibrosis. AB - A 47-year-old male Caucasian patient, with no previous relevant medical history, presented in September 1996 with persistent right lower quadrant abdominal pain. A tumor in the cecum was identified and the patient was submitted to ileocecal resection with ileocolic anastomosis. Histological examination showed a moderately differentiated adenocarcinoma. One year later he developed bloody diarrhea, urgency, and loss of weight. Based on clinical presentation and histology of large bowel biopsies, a diagnosis of ulcerative colitis (UC) was established. The previously resected surgical specimen was reevaluated, and lesions resembling UC were identified in the nonneoplastic mucosa. High levels of alkaline phosphatase and gamma-glutamyl transferase were detected. These alterations could be traced back to 1991. Endoscopic retrograde cholangiopancreatography was performed, showing diagnostic features of primary sclerosing cholangitis (PSC), and the patient was put on ursodeoxycholic acid therapy. In March 1999, he started to have progressive dyspnea and signs of cardiac failure. Endomyocardial biopsy was performed showing extensive lesions of endomyocardial fibrosis. This case illustrates a rather silent course of UC in the presence of PSC, and supports the postulated increased risk in the development of proximally located colorectal carcinoma in these patients. Additionally, the development of endomyocardial fibrosis constituted an unexpected finding, not previously reported in this setting. PMID- 12131614 TI - Antiadhesion molecule therapy in inflammatory bowel disease. AB - Adhesion molecules regulate the influx of leukocytes in normal and inflamed gut. Some of these molecules such as MadCAM-1 are specific for the gastrointestinal endothelium, but in inflammatory bowel diseases most of the adhesion factors are up-regulated. Adhesion molecules also are involved in local lymphocyte stimulation and antigen presentation within the intestinal mucosa. Recently, therapeutic compounds directed against trafficking of lymphocytes toward the gut mucosa have been designed, and are being developed as a novel class of drugs in the treatment of Crohn's disease (CD) and ulcerative colitis. This review deals with the immunological aspects of leukocyte trafficking focused on gut homing of T cells. Secondly, the changes in adhesion molecules and T-cell trafficking during intestinal inflammation are discussed. Finally, we review the clinical data that have been gathered in trials of biological therapies directed against adhesion molecules. Both antiintercellular adhesion molecule-1 (ICAM-1) and anti alpha4 integrin strategies are being developed. Trials with the anti-ICAM-1 antisense oligonucleotide, ISIS-2302, in steroid-refractory CD have provided conflicting efficacy data. The anti-alpha4 integrin antibodies natalizumab (Antegren) and LDP-02 are in phase III and phase II trials, respectively. In the near future, these novel biological agents may prove valuable therapeutic tools in the management of refractory IBD. PMID- 12131615 TI - The future of inflammatory bowel disease therapy. PMID- 12131618 TI - Your HIPAA "to-do" list. PMID- 12131619 TI - Assessing medicare eligibility: suggestions for improving processes. PMID- 12131620 TI - Using special foods and supplements as alternative cancer treatments. AB - This column investigates the harm/benefit issues involved in coupling alternative methods of cancer treatment with traditional cancer therapy. PMID- 12131622 TI - Providing a home care clinical experience that benefits patients, students, and agencies. AB - This article describes how a university baccalaureate program partnered with an established home care organization to provide services to the frail elderly who were no longer eligible for reimbursed home care. The planning, implementation, and evaluation of the program, and the benefits to the nurse interns, the clients, and the agency are described. PMID- 12131621 TI - Mobile chest drainage: coming soon to a home near you. AB - Home care nurses are seeing more patients with chest drainage devices than ever before. This article reviews the conditions that may be treated at home with mobile chest drainage devices, specific devices nurses are likely to see in the home, focused nursing assessments, tips for monitoring the devices, and key aspects of patient and caregiver teaching. PMID- 12131623 TI - Understanding urinary catheter problems from the patient's point of view. AB - Nurses are provided with a unique viewpoint in this article by examining the lived experience of individuals with long-term urinary catheters. Fourteen adults (ages 35-95) who had used a catheter from 6 months to 18 years were interviewed. In their own words, study participants discussed how they respond to catheter related problems, urinary tract infection, leaking/ blocking, catheter discomfort, and autonomic dysreflexia. Using this insight will help clinicians implement mutually developed care-plan goals with patients as partners. PMID- 12131624 TI - Advanced telecare for wound care delivery. AB - Applying dressings remains the mainstay of wound care, and skilled nursing makes real differences in patient care. Nurses using telehealth tools can better observe and manage wounds meaning interventions are more scientific, cost effective, and result in better patient outcomes. One agency shares how telehealth works for it. PMID- 12131625 TI - Making the admission process more efficient. PMID- 12131626 TI - The end user of home care computer technology...the clinician. PMID- 12131627 TI - CMS update. PMID- 12131628 TI - Infectious disease. PMID- 12131629 TI - Filling the family fuel tank. PMID- 12131630 TI - Interpretation of accident and emergency radiographs. PMID- 12131631 TI - Cardiovascular depression resulting from atenolol intoxication. AB - A case of massive atenolol ingestion leading to hypotension in association with PR and QRS interval prolongation on the electrocardiogram is presented. These clinical findings are identical to those attributed to the membrane-stabilizing activity of propranolol and other lipophilic beta-blockers. It is commonly believed that hydrophilic agents such as atenolol lack this activity. A review of the literature reveals that hydrophilic beta-blockers may have membrane stabilizing activity, though much higher concentrations are required to produce this action in comparison with lipophilic agents. This case and a review of the literature provides a potential pathophysiological basis for atenolol-induced haemodynamic depression. PMID- 12131632 TI - Decision rule and utility of routine urine toxicology screening of trauma patients. AB - The objective of this study was to determine the impact of urine drug screening of major trauma victims on patient care and derive a decision rule for selective screening. Retrospective chart review of 170 trauma patients at a Level I Trauma Center, certified by the American College of Surgeons, was undertaken. The decision rule was developed by Classification and Regression Tree (CART) analysis to maximize sensitivity, with secondary attention to specificity. Eighty-nine percent of trauma patients were screened, while 26.0% had positive tests for illicit drugs. Serum ethanol was positive in 31.2%, over the legal limit of 0.08 g/dl. Both a legally intoxicated ethanol level and positive illicit drug screen were found in 11.0%. Additionally, 42.5% of patients with a positive illicit drug screen were also intoxicated (blood alcohol level above legal limit). Conversely, 35.4% of legally intoxicated patients also had positive illicit screens. Drug treatment referral occurred in 17.5% of positive drug screens. For urgent surgery, median time to drug screen result was 117 min, while median time to operation was 110 min. Of operative patients, 57% had the drug screen result recorded on the chart at any time, but only 14.3% of illicit screens were noted in the anaesthesia record. For all patients with and without operations, 71.1% had the result noted on the chart. We derived a 'low risk rule' to identify most patients with positive illicit drug screens (95% sensitivity, 55% specificity, 66% positive and 93% negative predictive values; accuracy 74%), while limiting the number of unnecessary tests. The rule avoids screening 48% of patients, missing only 5% of true positives. It is concluded that urine screening for illicit drugs in trauma patients can be performed selectively according to a decision rule based on demographics, mechanism of injury and time of presentation. This rule, which captures most positive screens while eliminating screening in low risk patients, could result in significant cost savings. Only prospective validation of these rules in patient populations of other trauma centres will offer confidence that the decision points are valid. Urine drug screening infrequently affected patient management or resulted in drug treatment referral in our sample. We call for increased vigilance in recording results and referring patients for treatment. PMID- 12131633 TI - Attitude towards rape among doctors working in the emergency department. AB - In this study, a 29-item questionnaire survey was conducted to explore the attitude and knowledge toward rape amongst doctors working in Hong Kong. The Likert scale was used for questions on attitude and knowledge. A composite score was computed for both attitude and knowledge. Comparisons of the attitude and knowledge scores were made between doctors of different genders and ranks. Existing practices in the handling of rape cases were also explored. The response rate was 58%; 175 questionnaires were completed by 147 (84%) male and 28 (16%) female doctors. There were 12 (6.3%) consultants, 44 (23%) senior medical officers and 119 (62%) medical officers. The average length of emergency department experience was 4.7 years. Most doctors held a favourable attitude towards rape victims, and there was no significant difference between senior and junior doctors (P=0.062) or between doctors of different genders (P=0.793). However, 36% of the doctors agreed that 'a women should be responsible for preventing her own rape', and more female doctors agreed that 'a woman can successfully resist rape if she tries hard enough' (P<0.042). Senior doctors had a higher knowledge score (P<0.0001). The correlation between knowledge and attitude scores was low (Spearman coefficient 0.258; P<0.05). In conclusion, doctors working in the emergency department generally held favourable attitudes towards rape victims. However, there is still room for improvements in attitude and knowledge. A one-stop service is suggested to avoid fragmentation and prevent 'revictimization'. PMID- 12131634 TI - Moon cycles and violent behaviours: myth or fact? AB - We formulated the hypothesis that lunar phases, identified by the fraction of the illuminated visible surface of the moon, have a relationship with the frequency of victims of aggression seen in an emergency department. If such a relationship exists, an increase in the frequency of incidents with the phases of full moon or new moon would be expected. In order to test this hypothesis, the daily frequency of victims of violent behaviour seen in the emergency department was used to create a temporal series of data. This was then correlated with a temporal series of lunar luminosity data from the same time period. Crossed correlations in the delay range -7 to +7 days showed coefficient values ranging between -0.102 and +0.034, demonstrating weak correlations without statistical significance. Despite the attractiveness of the popular belief that the moon influences human behaviour, the analysis of our data does not support an association between lunar phases and frequency of violent behaviour. That is, we cannot predict the frequency of cases from a knowledge of lunar luminosity, at least in the period over which our study was performed. PMID- 12131635 TI - Are accident and emergency consultants as accurate as consultant radiologists in interpreting plain skeletal radiographs taken at a minor injury unit? AB - The objective of this study was to compare the accuracy of an accident and emergency (A&E) consultant in interpreting plain skeletal radiographs with that of a consultant radiologist (CR). It took the form of a retrospective study of 2133 radiographs taken in a Minor Injury Unit (MIU). A&E consultant reports on these films were compared with those of a CR and also with a gold standard. The A&E consultant diagnoses achieved an accuracy of 98.5% (CR 97.8%), sensitivity of 97.8% (CR 98.1%), specificity of 98.8% (CR 97.7%), positive predictive value of 97.3% (CR 95.1%) and negative predictive value of 98.97% (CR 99.07%) (gold standard of 100%). In conclusion, the A&E consultant reports of plain skeletal radiographs generated from an MIU were as accurate as those of a consultant radiologist. This could have significant implications for the wet reporting of A&E departmental radiographs. PMID- 12131636 TI - Comparison of stone size and response to analgesic treatment in predicting outcome of patients with renal colic. AB - The aim of this study was to compare the prognostic value of stone size and response to analgesic treatment in patients with renal colic. We reviewed the charts of patients treated for renal colic in our Emergency Department. The eligibility criteria were a radiological examination demonstrating direct or indirect signs of ureteral obstruction and/or a stone. The primary endpoint was the requirement for surgical treatment. The parameters considered as prognostic factors were pain relief with ketorolac (K) or ketorolac plus opiate treatment (KO), and stone size (>or= or <6 mm). Ninety-five patients were considered for analysis. Of these, 49 (52%) had a stone demonstrated radiologically. Four out of 27 patients (15%) in the KO group and six out of 68 patients (8.8%) in the K group required a surgical procedure to relieve the obstruction (NS). Four out of five patients (80%) with a stone >or=6 mm required a surgical procedure, compared with one out of 44 (2.2%) who had a stone smaller than 6 mm (P<0.001). In conclusion, stone size is a better prognostic factor than the response to analgesic treatment in predicting the clinical outcome of patients with renal colic. A stone >or=6 mm in patients with renal colic should alert the emergency physician that urological complications requiring surgical intervention may occur and that urological management may be warranted. PMID- 12131637 TI - Validity of urinalysis and microscopy for detecting urinary tract infection in the emergency department. AB - This study assessed the validity of standard urinalysis, urinalysis for leucocyte esterase and nitrites, and urgent microscopy in the diagnosis of urinary tract infection (UTI) in 60 female patients with a triage diagnosis of UTI. There were 24 (40%) proven UTIs after culture. Simple urinalysis was sensitive for UTI (95.8%) but the positive predictive value was only 45.1%. The addition of leucocyte esterase and nitrite urinalysis testing did not improve the sensitivity, but if both of these were positive the positive predictive value improved to 100%. Urgent microscopy alone was sensitive (100%) but non-specific (38.9%). The specificity of the diagnosis improved to 94.4% for organism counts of >or=10/microl and to 88.9% for leucocyte counts of >or=50/microl. The negative predictive value of no detectable leucocytes on microscopy was 94.7%. Screening for UTI in the emergency department (ED) population is improved by the addition of leucocyte esterase and nitrite test. A positive urinalysis test for leucocytes and nitrites, or urinalysis positive at levels of >or=500 leucocytes or >or=5 g/l protein should confirm a clinical diagnosis of UTI. Urgent urine microscopy should be performed only if the above criteria are not met yet a minimum of one urinalysis result is positive. PMID- 12131638 TI - Severe accidents due to windsurfing in the Aegean Sea. AB - Windsurfing is a popular sport and has recently become an Olympic event. As an open-air water activity that requires the participant to be in perfect physical condition, windsurfers may be prone to accidents when certain basic rules or procedures are violated. The current study monitored severe injuries due to windsurfing over a period of 12 months in the Aegean Sea in Greece. Our study revealed 22 cases of severe accidents due to windsurfing, with a wide range of injuries including head injuries, spinal cord injuries, and severe fractures of the extremities. Prolonged hospitalization, severe disability and two deaths occurred as consequences of these accidents. The study examined the characteristics of these patients and the possible risk factors and conditions associated with the accidents. We also focused on the most common types of injuries and reviewed the mechanisms that may provoke them. Water sports and particularly windsurfing represent a major challenge for the emergency medical system, especially in the Aegean Sea. Hundreds of islands, kilometres of isolated coasts, millions of tourists, an extended summer period and rapidly changing weather create conditions that constantly test the efficacy of the emergency services. The development of an appropriate infrastructure and maximum control of the risk factors causing these accidents could reduce the morbidity and mortality that, unfortunately but rather predictably, accompany this popular summer activity. PMID- 12131639 TI - Comparison of tissue adhesive and suturing in the repair of lacerations in the emergency department. AB - The objective of this study was to compare the applications of Histoacryl Blue (HAB) and suturing regarding cosmetic outcome, cost and patient and physician satisfaction in the emergency department (ED). A total of 92 consecutive adult patients with lacerations equal to or shorter than 5 cm were enrolled in the study. Patients were randomized to either HAB or suturing. Ten-day and three month cosmetic outcomes were evaluated via visual analogue scale (VAS) by a blinded surgeon. Cosmetic outcome, cost and patient and physician satisfaction of both groups were compared. Only 52 patients completed the follow-up at three months. Twenty-eight had been repaired with sutures and 24 with HAB. The differences regarding ten-day and three-month cosmetic outcome scales between the patients repaired with HAB and sutures were not statistically significant. Application of HAB resulted in greater satisfaction of the patient and the physician (p=0.007 and p=0.0001, respectively). Costs of HAB were significantly lower than sutures (p=0.0001). It is concluded that HAB is a cheaper method of laceration repair and results in greater satisfaction of both patients and physicians, while cosmetic outcomes were comparable. These results suggest that HAB is a viable alternative to suturing for selected lacerations in the ED. PMID- 12131640 TI - Traumatic retropharyngeal haematoma treated by embolization of the thyrocervical trunk. AB - Trauma involving the retropharyngeal space is relatively infrequent. Upper airway obstruction due to a retropharyngeal haematoma can be life threatening and requires immediate intervention. We present a well-documented case that illustrates the unexpected clinical course of such a haematoma and its management. PMID- 12131642 TI - Adult acute epiglottitis and foreign body in the throat - chicken or egg? AB - A 53 year old man presented with the chief complaint of having a fish bone stuck in the throat for about 1 h. There was no dysphagia or respiratory symptoms. Plain lateral neck X-ray, direct laryngoscopy and oesophagogastroduodenoscopy showed a grossly swollen epiglottis with narrowing of the laryngeal lumen. No foreign body was found. His condition improved rapidly with intravenous antibiotic therapy. As acute epiglottitis may be a sudden life-threatening condition, a high index of suspicion should be maintained for patients who present with alleged foreign bodies in the throat. PMID- 12131641 TI - Portal vein air embolization after blunt abdominal trauma: a case report and review of the literature. AB - Gas in the portal vein is a rare and often fatal condition in surgical patients. However, the presence of gas in the mesenteric and portal veins in association with abdominal trauma is a transient incidental finding that resolves spontaneously. We describe a young patient with Crohn's disease who suffered air embolism of the portal veins secondary to blunt abdominal trauma. The condition was clinically benign and resolved spontaneously. The pathogenesis is discussed and a review of the literature is provided. PMID- 12131643 TI - Sipple's syndrome presenting acutely as severe heart failure. AB - We describe the case of a 56-year-old man who developed acute hypertension leading to cardiac insufficiency, arrhythmia, severe heart failure and death. The autopsy revealed Sipple's syndrome (multiple endocrine neoplasia syndrome type IIa) and catecholamine-induced cardiomyopathy. This man had received a false diagnosis of primary hypertension 1 year before. The prime objective of this report is to call attention to the necessity of an in depth diagnosis of labile and paroxysmal hypertension. The clinical diagnostic features of phaeochromocytoma as well as the main therapeutic approaches suggested in the literature are commented on. PMID- 12131644 TI - Spinal epidural haematoma as a result of warfarin/fluconazole drug interaction. AB - This is the first reported case in the emergency medicine literature of a drug interaction between warfarin and fluconazole. We present a case of spinal epidural haematoma and summarize four other case reports reported elsewhere from 1988 to 1996. We admonish emergency physicians to be aware of this dangerous drug combination. Warfarin and fluconazole are frequently encountered drugs in the emergency department and thus any interaction between these drugs is of considerable importance. PMID- 12131645 TI - Salbutamol intoxication: is salbutamol a drug-inducing fever? A case report and treatment strategy. AB - A four-year-old female with salbutamol intoxication was referred to our paediatric emergency medicine unit, due to agitation, tremulousness, sinus tachycardia, mild hypokalaemia and hyperglycaemia. On admission the child was agitated and had a noticeable tremor, an axillary temperature of 38 degrees C and a pulse rate of 185 beats/min. She had no identifiable focus of infection on physical examination to explain her fever. Gastric lavage, activated charcoal, intravenous hydration and electrocardiogram (ECG) monitoring were performed. Her plasma potassium level, blood sugar and QT interval were closely monitored during her hospital stay. Her fever, tachycardia and serum potassium and glucose levels returned to normal and she was discharged in good condition 24 h after admission. The difference of this case from prior cases of salbutamol intoxication was the observation of fever in the absence of evidence of infection. Since the cause of fever was not a reaction to the medication used in the treatment or related to environmental factors, it is assumed that salbutamol is a fever-inducing drug. PMID- 12131646 TI - TV medical dramas - British or American: which approach do you prefer? PMID- 12131647 TI - Complications from folk remedies at the emergency department. PMID- 12131649 TI - Advanced cardiac life support update: the new ILCOR cardiovascular resuscitation guidelines. International Liaison Committee on Resuscitation. PMID- 12131648 TI - Advances in disaster medicine. PMID- 12131650 TI - Problems with current methods of data analysis and reporting, and suggestions for moving beyond incorrect ritual. PMID- 12131651 TI - The changing spectrum of gastroesophageal cancer. PMID- 12131652 TI - Costs and effectiveness in the care of patients with oral and pharyngeal cancer: analysis of a paradox. AB - Oropharyngeal cancer is estimated to be the ninth most common cancer worldwide. Its prognosis is largely dependent upon tumour-stage at the time of diagnosis. Stage I and II oropharyngeal cancers are characterized by a 5-year survival rate ranging from 70% to 90%, and the management of these early carcinomas is usually of short duration, easy and very cost-effective. On the other hand, the diagnostic evaluation, treatment and management of complications and recurrences of advanced stage oral tumours (stage III and IV) are often very long, complex and costly. They also have very poor prognosis with survival figures dropping to about 20%. Nowadays, most oropharyngeal cancers are detected at a late stage with an overall 5-year survival rate of around 45-50%, and with a conspicuous increase in treatment costs and a worsening of prognosis. Even if formal and comprehensive cost-effectiveness and cost-benefit analyses are not currently available in the oropharyngeal cancer literature, it seems clear that, in the care of these patients, the enormous consumption of resources is not associated with acceptable outcomes. New initiatives should be evaluated, planned and developed for the care of patients with oral and pharyngeal cancer. These strategies should be directed at prevention and early diagnosis in order to increase patient survival and quality of life and decrease the consumption of health care resources. PMID- 12131653 TI - Participation in a colorectal cancer screening programme: influence of the method of contacting the target population. AB - We assessed the effect of two different methods of contacting the target population on the rate of participation in a colorectal cancer screening programme. All individuals aged between 50 and 74 years enlisted in one primary health care centre in Barcelona (Spain) were included in a prospective randomized controlled trial. An invitation letter signed by a doctor together with two containers for faecal sample collection were sent by post to subjects in the 'standard' group (n = 1060), while subjects in the 'study' group (direct contact, n = 965) were visited by a trained non-health professional who supplied them with the same documentation as the standard group. The screening test consisted of an immunological method for the detection of faecal blood which does not require any prior specific dietary measures. Specimens were collected on two successive days. A significantly higher participation was observed in the study group (557/965, 57.7%) compared with the standard group (388/1060, 36.5%, P < 0.005). Specimen collection correctness was also higher in the study group (419/557, 75.1%) compared with the standard group (262/388, 67.5%, P < 0.014). There were no differences in terms of either age group or sex for the participation, nor for degree of correctness of specimen collection. Participation and specimen collection can be raised in colorectal cancer screening programmes by means of an invitation made through direct contact by a suitably trained non-health professional. PMID- 12131654 TI - The changing spectrum of gastroesophageal reflux disease. AB - The incidences of Barrett's oesophagus and oesophageal adenocarcinoma are rising. However there is no evidence on whether the incidence of gastroesophageal reflux disease is rising. This was a retrospective study investigating the incidence of gastroesophageal reflux disease at endoscopy from 1980 to 1995. The study took place in Tayside, Scotland. Using the Tayside endoscopy database, patients with new diagnoses of Barrett's oesophagus and endoscopic oesophagitis were identified. Cases and rates (per 1000 endoscopies) for oesophagitis, Barrett's oesophagus and combined group of oesophagitis + Barrett's oesophagus were calculated for each year. There was a significant decline in the incidence of oesophagitis. There was a significant large increase in the incidence of Barrett's oesophagus and a small but significant rise in the incidence of endoscopically identified gastroesophageal reflux disease (oesophagitis + Barrett's oesophagus). There was a significant decrease in the ratio of new cases of oesophagitis to new cases of Barrett's oesophagus. At endoscopy there has been a small increase in incidence of gastroesophageal reflux disease. There has, however, been a dramatic change in the spectrum of gastroesophageal reflux disease, with a larger proportion having Barrett's oesophagus than previously. PMID- 12131655 TI - Trends in gastric cancer incidence in a well-defined French population by time period and birth cohort. AB - Epidemiological studies have shown a marked decline in gastric cancer incidence and opposite patterns between proximal and distal sites among different populations. Little is known about trends by histological type. The aim of this study was to analyse the change in gastric cancer incidence patterns by investigating the role of temporal components as determinants of such trends in the population of the Cote d'Or area (France) registered between 1976 and 1995. Gastric cancer incidence decreased over time. There was a decrease in incidence rates for distal cancers (-3.5% P < 0.001 per year in men and -4.6% P < 0.01 in women). In contrast, there was a non-significant increase of proximal cancer incidence in men and in women. Rates of adenocarcinomas decreased, whereas the incidence rates of undifferentiated carcinomas and of other histological types remained quite stable. There was a decrease in cumulative risk throughout the studied cohorts, whereas risk for proximal cancer remained stable and decreased slightly for distal localization. For adenocarcinomas, earlier birth cohorts showed a slight decrease in rates, whereas there was an increase for recent cohorts. Subsite and histological-specific analysis, in revealing different time trends in incidence, suggest, at least partly, different aetiologies for gastric cancer, and future aetiological studies must distinguish proximal and distal cancers. PMID- 12131656 TI - Radiation exposure from diagnostic and therapeutic treatments and risk of breast cancer. AB - An association between low-dose diagnostic X-ray exposure or therapeutic radiation treatment and breast cancer risk has not been established. To further investigate the issue, we analysed data from a case-control study of breast cancer in Connecticut in 1994-1997. A total of 1217 subjects (608 breast cancer cases and 609 controls), 30-80 years old, participated in the study. A standardized, structured questionnaire was used to collect information through in person interviews on diagnostic or therapeutic radiation and other breast cancer risk factors. An odds ratio (OR) of 1.7 (95% confidence interval (CI) 0.8-3.6) was observed for postmenopausal women with therapeutic radiation treatment for skin problems such as ringworm and acne, and an OR of 2.5 (95% CI 1.0-6.8) for those who reported having been treated six or more times. Radiation treatment received at younger ages seems to carry a higher risk. In earlier studies therapeutic radiation for skin problems has been associated with an increased risk of breast cancer. Therefore, it is possible that scattered radiation from these treatments could increase the risk of breast cancer. Radiation exposure from diagnostic X-rays was not associated with a significantly increased risk of breast cancer in this study. PMID- 12131657 TI - Cigarette and alcohol consumption and the risk of colorectal cancer in Shanghai, China. AB - The relation of cigarette smoking and alcohol drinking to colorectal cancer risk has been inconsistent in the epidemiological literature. In a population-based case-control study of colorectal cancer in Shanghai, China, where the incidence rates are rising sharply, we examined the association with tobacco and alcohol use. Cases were aged 30-74 years and newly diagnosed with cancers of the colon (N = 931) or rectum (N = 874) between 1990 and 1992. Controls (N = 1552) were randomly selected among Shanghai residents, frequency-matched to cases by gender and age. Information on lifetime consumption of tobacco and alcohol, as well as demographic and other risk factors, was obtained through in-person interviews. Associations with cigarette smoking and alcohol use were estimated by odds ratios (ORs) and 95% confidence intervals (CIs). Among women, the prevalence of smoking and alcohol drinking was low, and no significant association with colon or rectal cancer was observed. Although cigarette smoking among men was not related overall to colon or rectal cancer risk, there was a 50% excess risk of rectal cancer (OR 1.5, 95% CI 0.9-2.5) among those who smoked 55 or more pack-years. Among men, former alcohol drinkers had an increased risk of colon cancer (OR 2.3, 95% CI 1.4 3.7) but not rectal cancer, while current drinkers had a 30-50% excess risk of colon cancer only among those with long-term (30+ years) and heavy (>560 g ethanol/week) consumption. The excess risks were mainly associated with hard liquor consumption, with no material difference in risk between proximal and distal colon cancer. Although cigarette smoking and alcohol drinking in general were not risk factors for colorectal cancers in Shanghai, there were small excess risks for rectal cancer among heavy smokers and colon cancer among heavy drinkers. PMID- 12131658 TI - Prognostic importance of serum vascular endothelial growth factor in relation to platelet and leukocyte counts in human renal cell carcinoma. AB - It has been shown that both serum vascular endothelial growth factor (VEGF) and also platelet counts are associated with survival in renal cell carcinoma (RCC). It is not known, however, whether VEGF in serum relates to the angiogenic activity of the tumour or is derived from circulating blood components. Therefore, the interrelation between serum VEGF, platelet and leukocyte counts compared with health history, clinicopathological findings and outcome was evaluated in patients with RCC. Blood samples were collected before nephrectomy in 161 patients. Serum VEGF165 was assessed by a quantitative ELISA method. Platelet and leukocyte counts were analysed routinely and obtained from medical records. The variables were compared using univariate and multivariate analysis. There were significant correlations between VEGF levels, and platelet (P < 0.001) and leukocyte counts (P < 0.001). Serum VEGF levels, platelet counts, as well as leukocyte counts correlated significantly to stage and grade. Platelet counts were significantly lower in men with medication (P = 0.042), and decreased with age particularly in women (P = 0.001). Age or medication did not affect VEGF levels or leukocyte counts. Both VEGF and platelets gave significant prognostic information in univariate analysis. Using Cox multivariate analysis, VEGF was the last variable to be excluded. Only stage and grade remained as independent prognostic factors. Both VEGF levels and platelet counts gave prognostic information but VEGF was more reliable as predictor of survival in patients with RCC. PMID- 12131659 TI - Risk factors for male breast cancer in Canada, 1994-1998. AB - Relatively little attention has been paid to the aetiology of male breast cancer and the current understanding of female breast cancer, primarily related to reproductive events, cannot be readily transferred to understanding the cancer in males. However, since male breast cancer occurs in the absence of factors related to childbearing and menstruation, its aetiology may provide special insights into the causes of breast cancer in women. We examined lifestyle risk factors for male breast cancer as part of a Canadian, multi-site, population-based, case-control study. Eighty-one newly diagnosed, histologically confirmed cases and 1905 male controls aged 42-74 were analysed using unconditional logistic regression. Increased risks were found for men with a mother or sister with breast cancer (adjusted odds ratio (OR) 3.65, 95% confidence interval (95% CI) 1.62-8.19). Higher physical activity levels (moderate, and strenuous recreational plus occupational) were associated with a decreased risk of male breast cancer (highest quartile, adjusted OR 0.48, 95% CI 0.26-0.91). Similarly, higher risks were associated with higher weight 2 years before interview (2.19, 95% CI 1.08 4.43), maximum weight (OR 2.66) and higher body mass index (OR 1.60). Higher vegetable consumption and coffee consumption were associated with decreased risk, whereas higher beta-carotene, vitamin E and calcium supplementation were associated with statistically significant increased risk. The small number of cases and multiple comparisons preclude strong conclusions, but our study is consistent with studies suggesting obesity and family history increase risk, and physical activity decreases risk of breast cancer. PMID- 12131660 TI - Oral contraceptive use and mammographic patterns. AB - High-risk mammographic patterns represent an increased risk of contracting breast cancer and may be used as a surrogate endpoint for the disease. We examined the relationship between oral contraceptive (OC) use and mammographic patterns among 3218 Norwegian women, aged 40-56 years. Information on ever OC use, duration, and age of first OC use and other epidemiological data were obtained through questionnaires. The mammograms were categorized into five groups. Patterns I-III were combined into a low-risk group and patterns IV and V into a high-risk group. Odds ratios (OR) and 95% confidence intervals (CI) were estimated using logistic regression and adjusted for age, menopausal status, parity, age at first birth, and body mass index. Women who reported ever having used OCs were 20% more likely (OR 1.27, 95% CI 1.0-1.6) to have high-risk mammographic patterns compared with those reporting never having used OCs. There was no dose response between different measures of OC use and high-risk patterns. Among nulliparous women, ever OC users were four times more likely (OR 4.65, 95% CI 2.1-10.3) to have high risk patterns compared with never users. Our findings suggest that, especially among nulliparous women, ever OC use may exert its effect on breast cancer risk through changes in breast tissue, which can be observed on a mammogram. PMID- 12131661 TI - Butyrate and aspirin in combination have an enhanced effect on apoptosis in human colorectal cancer cells. AB - Laboratory and epidemiological studies suggest that butyrate, a metabolic product of microbial fermentation of dietary fibre, and aspirin, a non-steroidal antiphlogistic drug, both reduce the risk of developing colon cancer. Notably, few data exist on potential interactions of these two substances. In this study, the effects of a butyrate-aspirin combination on human colon cancer cells were compared with treatment with aspirin or butyrate alone. Both substances decreased proliferation and induced differentiation and apoptosis. Butyrate reduced mutant p53 expression, whereas aspirin did not affect p53 expression. Butyrate-induced apoptosis correlated with an increase in Bak expression and a decrease in the expression of Bcl-XL. Aspirin had no effect on the investigated apoptosis controlling factors. The antiproliferative and pro-apoptotic effects of the butyrate-aspirin combination were markedly enhanced. The combination resulted in a stronger decrease in the expression of PCNA and cdk2. Our data suggest that the anticarcinogenic effect of aspirin might effectively be augmented by combination with the short-chain fatty acid butyrate. PMID- 12131662 TI - Correlates of pregnancy oestrogen, progesterone and sex hormone-binding globulin in the USA and China. AB - The objective of this study is to examine perinatal correlates of oestradiol (E2), oestriol (E3), progesterone and sex hormone-binding globulin (SHBG) among pregnant women in the USA and China. Three hundred and four Caucasian women in Boston and 335 Chinese women in Shanghai were studied. Levels of E2, E3, progesterone and SHBG were measured in maternal blood at weeks 16 and 27 of gestation, and correlated with maternal, gestational and perinatal characteristics. Height, weight and body mass index (BMI) before pregnancy is inversely associated with E2 and SHBG, whereas E3 is inversely associated with height and progesterone is inversely associated with weight and BMI. A previous live birth is associated with lower E2 and SHBG in the index pregnancy. Total gestation duration is inversely associated with E2, E3 and progesterone, whereas weight gain during pregnancy is inversely associated with progesterone and SHBG. In the US, pregnancies with female fetuses are characterized by significantly reduced progesterone. Pregnancy hormones are associated with several maternal, gestational and neonatal characteristics. PMID- 12131664 TI - Can physical trauma cause breast cancer? AB - The objective of this study is to explore the effect of lifestyle on the risk of invasive breast carcinoma in women aged 50-65 years. A case-control study using a questionnaire and a semi-structured interview. Cases (n = 67) and controls (n = 134) were closely matched on known risk factors for breast cancer including age, family history, age at menarche, parity, age at first birth and menopausal status. Controls were chosen from a pool of 5600 women who attended for breast screening and filled in a questionnaire giving details to allow matching with cases. The study took place at the North Lancashire Breast Screening Service. Women were aged 50-65 years and presented with breast cancer or attended for breast screening. Women with breast carcinoma were more likely to report physical trauma to the breast in the previous 5 years than were the controls (odds ratio (OR) 3.3, 95% confidence interval (CI) 1.3-10.8, P < 0.0001). There were no significant differences in a wide range of other lifestyle indicators including factors relevant to social class, education, residence, smoking and alcohol consumption. In conclusion, recall bias is an unlikely explanation for these results in view of the nature and severity of physical trauma. Models of epithelial cell generation indicate that a causal link between physical trauma and cancer is plausible. A latent interval between cancer onset and presentation of under 5 years is also plausible. The most likely explanation of the findings is that physical trauma can cause breast cancer. PMID- 12131663 TI - Is use of hormone replacement therapy associated with increased detection of human papillomavirus and potential risk of HPV-related genital cancers? AB - Oral contraceptives (OC) are a risk factor for female genital cancers and in vivo studies have shown that progestins stimulate human papillomavirus (HPV) gene expression. A similar role for hormone replacement therapy (HRT) has received little evaluation. Cervical/vaginal specimens were obtained to detect HPV from postmenopausal women (n = 429) seeking annual gynaecologic care. HPV was detected in 14% of women and 4.4% had high-risk, oncogenic types. HPV prevalence was similar across current, past and never HRT users. After adjustment for HPV related risk factors, current and past user status showed no increased viral detection compared with never users. HRT duration also did not elevate risk among current users. However, longer duration (adj. OR 1.5/year, 95% CI 1.0-2.3) and longer latency (adj. OR 1.2/year, 95% CI 0.9-1.7) among past users of oestrogen/progestin regimens were associated with greater risk. Overall use of HRTs was not associated with HPV detection or disease. However, past users of combination HRTs had significantly greater risk of HPV detection with longer HRT duration and latency, similar to OC-HPV findings. The recommendation that postmenopausal women continue HRTs long term may lead to an increased development of HPV-related diseases, of particular concern among those who discontinue HRTs and subsequent gynaecologic care for early cancer detection. PMID- 12131677 TI - Anti-idiotypic antibodies from highly sensitized patients stimulate B cells to produce anti-HLA antibodies. AB - BACKGROUND: Sustained allosensitization increases waiting time for transplantation and increases the risk of rejection. The purpose of this study is to examine the effect of anti-idiotypic antibodies on B-cell responses and to define their role in alloantibody production. METHODS: The Immunoglobulin G (IgG) fraction, or the sera of 19 highly sensitized (HS) patients was absorbed to remove anticlass I antibody and was incubated with B cells. The culture supernatant was assayed for antihistocompatibility leukocyte antigen (HLA) antibody and tested for reactivity against a panel of normal lymphocytes. Similar studies were performed in 5 of the 19 patients who had a fall in alloantibody levels. RESULTS: The IgG (HS) fraction induced anti-HLA antibody from normal and autologous B cells in all 19 HS patients studied. The reactivity to HLA antigens in the culture supernatant was similar to the sera for each patient studied. The in vitro generated anti-HLA antibody bound to the IgG fraction used to stimulate the B cells. The in vitro production of anti-HLA antibodies was absent in the serum of all five patients who became nonsensitized. CONCLUSIONS: All patients who have high levels of alloantibody have anti-idiotypic antibodies in their sera that stimulate B cells to produce anti-HLA class I antibody similar in reactivity to that of their own sera. In the patients who have nondetectable alloantibodies in their sera, the stimulating anti-idiotypes are not measurable. Anti-idiotypic antibodies may act as a vaccine and cause sustained levels of alloantibody production. PMID- 12131679 TI - Improving the quality of kidneys from non-heart-beating donors, using streptokinase: an animal model. AB - BACKGROUND: Non-heart-beating donors (NHBD) offer a promising potential to increase the cadaveric organ donor pool, especially for kidneys. However, almost half of NHBD kidneys are discarded after viability assessment. This wastage is costly in both human and monetary terms. Intravascular thrombosis at the time of donor death is an event leading to failure in the viability assessment. We have developed an animal model to investigate the effects of the addition of streptokinase to the in situ flush medium before transplant in an attempt to redress the situation. METHODS: Two groups of eight healthy young Landrace Yorkshire white pigs were entered into the study. Both groups were subjected to approximately 70 min warm ischemia. Both groups received an intravascular flush with 4 L of Marshall's solution with heparin (1000 IU/L); one group of pigs also had streptokinase (1.5 MIU/L) added. After donor nephrectomy, all kidneys were machine perfused for 4 hr. Data on perfusion characteristics were taken and samples of kidney effluent were assayed for tissue damage biomarkers, glutathione S-transferase (GST) and alanine aminopeptidase (Ala-AP). Wedge sections of porcine kidneys were taken at the end of perfusion, for histological analysis. RESULTS: Data on machine perfusion parameters (temperature, mean pressure index, resistance) indicate better cooling, lower resistance, and lower mean pressure index in the streptokinase-treated group of pigs. GST and Ala-AP levels were increased in the nonstreptokinase group perfusates. Histopathological analysis of porcine kidneys indicated more ischemic injury and tissue damage in the nonstreptokinase group. CONCLUSION: The use of streptokinase in this porcine NHBD model conferred benefits to donor kidneys when assessed by machine perfusion. Markers of biochemical injury indicated that less renal tissue damage occurred with the incorporation of streptokinase in the in situ flush medium. PMID- 12131678 TI - Elimination of anti-Gal B cells by alpha-Gal ricin1. AB - BACKGROUND: A major barrier in pig to human organ transplantation is the binding of human anti-Gal to alpha-gal epitopes (Gal alpha 1-3Gal beta 1-4GlcNAc-R) on pig cells, resulting in hyperacute and acute vascular rejection of pig xenografts. Moreover, the immune system in xenograft recipients is activated by these epitopes to produce high affinity anti-Gal, which is also detrimental to xenografts. Production of anti-Gal can be prevented by specific elimination of anti-Gal B cells. This was achieved with the toxin ricin A, coupled to human alpha1-acid glycoprotein modified to carry alpha-gal epitopes. This complex, designated alpha-gal ricin, is targeted in vivo to anti-Gal B cells by interaction with the immunoglobulin molecules (i.e., B cell receptors) on these cells. METHODS: Carbohydrate chains on alpha 1-acid glycoprotein were converted to carry alpha-gal epitopes by enzymatic treatment with recombinant alpha 1,3 galactosyltransferase (alpha 1,3GT). This molecule and ricin A were biotinylated and coupled by avidin to generate alpha-gal ricin. The efficacy of alpha-gal ricin in eliminating anti-Gal B cells was studied in the experimental model of alpha 1,3GT knockout (KO) mice. These mice produce large amounts of anti-Gal immunoglobulin G when immunized with pig kidney membranes, as measured by ELISA with alpha-gal epitopes linked to bovine serum albumin (BSA). In the absence of anti-Gal B cells, these mice lack the ability to produce anti-Gal. RESULTS: Repeated administration of alpha-gal ricin into alpha1,3GT KO mice resulted in elimination of anti-Gal B cells, thereby preventing production of anti-Gal immunoglobulin G after immunization with pig kidney membranes. This prevention of anti-Gal production occurred with doses of alpha-gal ricin that were not toxic to the mice and did not affect production of antibodies with other specificities. CONCLUSIONS: Administration of alpha-gal ricin results in specific elimination of anti-Gal B cells in alpha 1,3GT KO mice. The elimination of these B cells may prove to be helpful in attempts to achieve immune tolerance to alpha-gal epitopes in primates. PMID- 12131680 TI - Role of anti-tumor necrosis factor-alpha in ischemia/reperfusion injury in isolated rat liver in a blood-free environment. AB - BACKGROUND: Warm ischemia/reperfusion injury during liver transplantation is the most important cause of primary nonfunction of liver allografts. Tumor-necrosis factor (TNF)-alpha apparently mediates tissue damage by inducing apoptosis and/or necrosis in liver transplants. The aim of the study was to determine, using an isolated rat liver model, if pretreatment with anti-TNF-alpha monoclonal antibodies can attenuate ischemia/reperfusion liver injury. Specifically, its effect on liver cell apoptosis through the modulation of caspase activity was examined in a blood-free environment. METHODS: Isolated rat livers were perfused with Krebs-Henseleit solution and randomly divided into three groups: (1) continuous perfusion for 165 min (control); (2) perfusion for 90 min, break for 60 min (ischemia), and reperfusion for 15 min; (3) as with group 2, but with administration of monoclonal mouse anti-rat TNF-alpha monoclonal antibodies before induction of ischemia. Caspase-3- and -9-like activity was measured by fluorometric assay, and apoptotic cells were identified by morphological criteria and application of the terminal deoxnucleotidyl transferase-mediated dUTP nick end-labeling (Tunel) assay. RESULTS: Portal pressure increased significantly in group 2 (14.8+/-2.3 mm Hg) compared to group 3, which showed no change (P<0.05). Significant amounts of TNF-alpha were detected in the effluent in group 2 at 1 min of reperfusion (147+/-8.9 pg/ml) compared to group 3 (30+/-6.7 pg/ml, P<0.05). Statistically significant reductions in liver enzyme levels were also noted in the animals pretreated with TNF-alpha antibodies (P<0.02). Caspase-3 and -9 activity was significantly decreased (270 and 160%, respectively) in group 3 compared to group 2 (P<0.005 and <0.05, respectively). A significant reduction in postischemic hepatic injury was noted on Tunel assay: many apoptotic hepatocyte cells were detected in group 2 but not in livers pretreated with monoclonal mouse anti-TNF-alpha antibodies (group 3). CONCLUSIONS: Neutralization with specific monoclonal antibodies against TNF before ischemia induction can attenuate postischemic hepatic injury. Apoptotic injury seems to be ameliorated through modulation of caspase-3- and -9-like activity. PMID- 12131681 TI - Recipient bone marrow engraftment in donor tissue after long-term tolerance to a composite tissue allograft. AB - BACKGROUND: An important component of a composite tissue limb allograft (CTA) is the vascularized bone marrow and bone marrow stroma, which when transplanted could create immediate marrow space and engraftment. We have previously demonstrated that tolerance to musculoskeletal allografts can be achieved with a 12-day course of cyclosporine without the presence of long-term peripheral donor cell chimerism. The objective of this study was to determine the fate of the donor bone marrow after transplantation of a limb allograft in a miniature swine model. METHODS: CTAs from donor swine were heterotopically transplanted into six MHC-matched, minor-antigen-mismatched recipients, and a 12-day course of cyclosporine was given. Previous animals transplanted without cyclosporine rejected their grafts in less than 42 days. A non-MHC-linked marker, pig allelic antigen (PAA), was used to distinguish host and donor cells. Three PAA- animals received PAA+ CTAs, and three PAA+ animals received PAA- CTAs. Bone marrow was harvested from the donor limb grafts and the recipient and analyzed by flow cytometry and histology. Thymus, spleen, and mesenteric lymph nodes were also harvested from the recipient swine and evaluated for the presence of donor cells by flow cytometry. RESULTS: All animals receiving cyclosporine demonstrated permanent tolerance to their allografts. Donor bone marrow cells were present in all grafts at the time of transplantation and during the immediate postoperative period. By 48 weeks, donor cells were no longer detectable within the marrow space of the allograft. In long-term animals host bone marrow cells replaced donor cells in the graft marrow space. No evidence of donor cell engraftment was found in recipient animals. CONCLUSION: This study demonstrates that in long-term tolerant recipients of musculoskeletal allografts there is no evidence of persistent donor bone marrow cells in the hematopoietic tissues of the graft or the host. Rather, the recipient's bone marrow cells and lymphocytes repopulate the donor marrow space of the graft. PMID- 12131682 TI - Investigation of promoter polymorphisms in the tumor necrosis factor-alpha and interleukin-10 genes in liver transplant patients. AB - BACKGROUND: Cytokines such as tumor necrosis factor (TNF)-alpha and interleukin (IL)-10 play significant roles in the inflammatory and immune responses that mediate allograft rejection. The presence of a G-->A polymorphism at position 308 in the promoter region of the TNF-alpha gene increased its transcription 6- to 7-fold. A similar polymorphism at position -1082 of the IL-10 promoter results in decreased production of IL-10 protein. In this study we have determined whether the single nucleotide polymorphisms in the promoter regions of the TNF alpha and IL-10 genes can predict the outcome of the allograft in liver recipients. METHODS: DNA was extracted from whole blood of liver recipients. The genotype of the patients was determined by polymerase chain reaction using sequence-specific primers. The level of TNF-alpha and IL-10 protein was measured by ELISA after stimulation of peripheral blood mononuclear cells with concanavalin A. RESULTS: There was significant correlation between acute cellular rejection and the presence of the -308A polymorphism (P<0.001), with 8 of 13 patients with the TNF-alpha polymorphism having evidence of acute rejection. Cell stimulation studies revealed that the level of TNF-alpha protein produced by patients with liver rejection was significantly higher than for patients without rejection (P=0.001). There were no strong associations between the presence of the IL-10 polymorphisms and rejection (P=0.71). CONCLUSIONS: This study adds to the understanding of the role of cytokine polymorphisms in liver transplants. The data suggest that cytokine promoter polymorphisms may be a risk factor associated with allograft rejection in the liver. PMID- 12131683 TI - Sensitized patients require sharing of highly matched kidneys. AB - BACKGROUND: Improved HLA matching can allow for renal transplantation in sensitized patients. United Network for Organ Sharing (UNOS) is currently considering removing the points awarded for HLA matching. METHOD: Twenty-six transplant centers shared kidneys, according to the algorithm: A, UNOS mandatory 0-mismatch (MM) share; B, UNOS mandatory 0-mm payback; C, 0 - B,Dr-MM; PRA> or = 40%; ROP tray negative; D, 0-B,Dr-MM; panel reactive antibodies (PRA) <40%; E, 0 B,Dr payback; F, local use. RESULTS: A total of 354 patients received transplants through the program and were compared with 6492 control patients. There was a significantly greater primary nonfunction rate in sensitized patients in spite of similar length cold ischemia time. Blacks were not significantly disadvantaged by the HLA matching requirements of the program. White recipients were favored by the matching program to the detriment of race group "other." Women were more frequently found in the sensitized groups but were not favored by sharing. Retransplant patients and patients with current PRA > or = 40% were favored by the sharing program. Sharing for HLA match does not improve graft survival at 1, 2, 3 years. CONCLUSIONS: (1). Elimination of suboptimal HLA matching by UNOS will probably not adversely affect 1-, 2-, and 3-year graft survival; (2). Removing the consideration for HLA matching will result in fewer transplant opportunities for highly sensitized patients such as retransplants and currently sensitized patients. PMID- 12131684 TI - Anatomical variations and surgical strategies in right lobe living donor liver transplantation: lessons from 120 cases. AB - BACKGROUND: Anatomical variations in right liver lobe are common. However, clinical implications and surgical management of these variations in living donor liver transplantation have not been analyzed systematically. METHODS: Surgical anatomy of vascular and biliary structures in 120 right lobe grafts were reevaluated by reviewing the results of preoperative (computerized tomography and Doppler ultrasonography) and intraoperative (cholangiography) imaging as well as surgical findings. The data were analyzed in relation to surgical management of anatomical variations. RESULTS: The incidence of variants leading to multiple portal vein anastomoses was 7.5%. The incidence of dual right hepatic veins was 0.8%; 30% of the grafts had significant accessory hepatic veins (>5 mm) and 13.9% of these were multiple. All of them were successfully reconstructed with technical modifications including venoplasty and venous grafts, except for two cases with multiple intraparenchymal portal vein branches to the anterior segment. The incidence of dual hepatic arteries was 1.7%, but only one of them was reconstructed without negative sequelae. The incidence of variants potentially leading to multiple bile duct anastomoses was 35.0%, and eventually 39.2% of the grafts had multiple orifices. With a variety of techniques including ductoplasty, hepaticohepaticostomy, and biliary stent, total incidence of leakage and stenosis was 10.8% and 9.2%, respectively. Although ductoplasty, internal stent or no stenting, seemed to be associated with increased risk of complications, anatomical variants, multiple bile ducts, and duct-to-duct reconstruction did not bear a significant risk. CONCLUSIONS: Anatomical variations of vascular and biliary structures in right lobe grafts are common. However, most can be managed safely with technical modifications. Only cases with intraparenchymal origin of the anterior portal vein(s) may form a relative contraindication, especially when combined with similar biliary variants. Otherwise, intraoperative assessment of biliary anatomy was enough for successful management. Detailed and precise assessment of vascular and biliary anatomy is vital for appropriate surgical management. PMID- 12131686 TI - Treatment of a patient with hemophilia A and hepatitis C virus-related cirrhosis by living-related liver transplantation from an obligate carrier donor. AB - BACKGROUND: Decompensated hepatitis C virus (HCV)-related cirrhosis is the main indication for liver transplantation. We report the first successful living related liver transplantation in a 49-year-old hemophilia A patient with end stage HCV-related cirrhosis using a graft obtained from his 20-year-old daughter, an obligate carrier. METHODS: The donor's autologous fresh-frozen plasma rich in factor VIII (FVIII) by treatment with 1-deamino-8-D-arginine vasopressin was prepared before the operation. At induction, 1-deamino-8-D-arginine vasopressin was given to the donor to increase plasma FVIII level. In addition, autologous fresh-frozen plasma containing high FVIII concentrate was infused intraoperatively. The right lobe was harvested from the donor and transplanted orthotopically. The recipient was treated postoperatively with recombinant FVIII and immunosuppressive agents. RESULTS: The donor did not receive recombinant FVIII or allogenic blood during perioperative periods. No bleeding was encountered in the donor perioperatively. The recipient showed a steady increase in FVIII activity postoperatively and was discharged 40 days after transplantation. Ribavirin and interferon-alpha were provided for 3 months postoperatively to prevent potential recurrence of HCV infection. Serum HCV-RNA by RT-PCR became negative after such treatment. CONCLUSIONS: End-stage liver disease in patients with hemophilia A can be an indication for living-related liver transplantation. Furthermore, a graft from a living-related donor with hemophilia A carrier seems to be suitable provided such individuals receive adequate support for coagulopathies. PMID- 12131685 TI - A randomized, prospective, double-blinded evaluation of selective bowel decontamination in liver transplantation. AB - BACKGROUND: Bacterial infection is a frequent, morbid, and mortal complication of liver transplantation. Selective bowel decontamination (SBD) has been reported to reduce the rate of bacterial infection after liver transplantation in uncontrolled trials, but benefits of this intervention have been less clear in controlled studies. METHODS: Eighty candidates for liver transplantation were randomly assigned in a double-blinded fashion to an SBD regimen consisting of gentamicin 80 mg+polymyxin E 100 mg+nystatin 2 million units (37 patients) or to nystatin alone (43 patients). Both treatments were administered orally in 10 ml (increasing to 20 ml, according to predefined criteria), four times daily, through day 21 after transplantation. Anal fecal swab cultures were performed on days 0, 4, 7, and 21. Rates of infection, death, and charges for medical care were assessed from day 0 through day 60. RESULTS: More than 85% of patients in both treatment groups began study treatment more than 3 days before transplantation. Rates of infection (32.4 vs. 27.9%), death (5.4 vs. 4.7%), or charges for medical care (median $194,000 vs. $163,000) were not reduced in patients assigned to SBD. On days 0, 4, 7, and 21, growth of aerobic gram negative flora in fecal cultures of patients assigned to SBD was significantly less than that of patients taking nystatin alone; growth of aerobic gram-positive flora, anaerobes, and yeast was not significantly different. CONCLUSION: Routine use of SBD in patients undergoing liver transplantation is not associated with significant benefit. PMID- 12131688 TI - Late nephrologist referral and access to renal transplantation. AB - BACKGROUND: Our aim was to explore a possible association between late nephrologist referral before onset of renal replacement therapy (RRT) and the likelihood of receiving a renal transplant. METHODS: For the period of 1991 to mid-1996 we defined an inception cohort of all patients with new-onset chronic RRT who were New Jersey Medicare and/or Medicaid beneficiaries in the year before RRT and who had been diagnosed with renal disease more than 1 year before RRT. To control for known risk factors and confounders of access to renal transplantation, we conducted a matched case-control study. Using number of days from onset of RRT to transplantation as the index date for cases, we successfully matched 32 transplant recipients (cases) with 197 controls who shared the cases' age (+/-2 years), gender, race (white/black/other), and year of onset of RRT (+/ 1 year) but had not received a transplant on index date. Using conditional logistic regression, we evaluated the effects on the likelihood of transplantation of late referral (< or = 90 days vs. >90 days before first RRT) and socioeconomic status (lower socioeconomic status indicated by enrollment in Medicaid or another state program for the poor), further controlling for comorbidity (Charlson score) in the year before index date. RESULTS: In the full multivariate conditional model, late referral was significantly associated with a much lower rate of renal transplantation (odds ratio [OR]=0.22; 95% confidence interval [CI]: 0.05, 0.97), as were socioeconomic status (OR=0.18; 95% CI: 0.04, 0.82) and comorbidity status (OR=0.69; 95% CI: 0.48, 1.00). CONCLUSIONS: Delayed referral of renal patients to a nephrologist before RRT is significantly associated with reduced access to renal transplantation, independent of age, gender, race, socioeconomic and comorbidity status. The validity of our result needs to be confirmed in larger populations. PMID- 12131687 TI - Accreditation of skin from a methanol-poisoned victim for banking and grafting. AB - BACKGROUND: Acute poisoning is a contraindication for organ and tissue donation. In this study the suitability of skin from a methanol-poisoned (MP) donor for future grafting and keratinocytes culturing was investigated. METHODS: A patient was admitted with a methanol blood level of 2.7 mg/mL, which became undetectable after 4 days of treatment with 4-methylpyrazole (fomepizole). Upon declared brain death and family consent, organs and skin were harvested. For approving MP skin for grafting, the following parameters were studied: viability and plating efficiency of MP keratinocytes, integrity of MP skin after cryopreservation, and its performance as xenografts on wounds in a pig model. Nonpoisoned (NP) controls included skin of matching age, cryopreservation period, and NP keratinocytes. RESULTS: No significant differences were observed for any parameter between NP and MP samples. Furthermore, in vitro exposure of NP keratinocytes and fibroblasts to <10 mg/mL methanol inhibited their growth by <20%, with an extrapolated LD50 of 100 mg/mL. A parallel exposure to formaldehyde, a spontaneous metabolite of methanol, yielded LD50 of 20 microg/mL and eradication of viability at 300 microg/mL. CONCLUSIONS: These results indicate that skin from a carefully monitored MP donor is suitable for banking toward massive burns and skin losses. This methodology may be applied to approve skin harvested from other types of poisoned donors for banking and future grafting. PMID- 12131689 TI - The changing causes of graft loss and death after kidney transplantation. AB - BACKGROUND: The results of kidney transplantation have improved markedly over the last three decades. Despite this, patients still lose grafts and die. We sought to determine whether the causes of graft loss and death have changed over the last 30 years. METHODS: We reviewed patients who underwent transplantation or who died between January 1, 1970 and December 31, 1999. We compared the causes of graft loss or death for three decades: 1970 to 1979, 1980 to 1989, and 1990 to 1999. RESULTS: From January 1, 1970 to December 31, 1999, we performed 2501 kidney transplantations in 2225 patients. For the three periods, 210, 588, and 383 patients lost their grafts, respectively. Graft survival increased substantially. Graft loss occurred later after transplantation, with 36.0% losing grafts in the first year during 1970 to 1970, 22.8% during 1980 to 1989, and 11.4% during 1990 to 1999. Death with a functioning graft increased from 23.8% for 1970 to 1979 to 37.5% for 1990 to 1999. Concomitantly, rejection as a cause of graft loss fell from 65.7% for 1970 to 1979 to 44.6% for 1990 to 1999. Approximately two thirds of the patients who died after transplantation died with a functioning graft and one third died after returning to dialysis. Cardiac disease as a cause of death increased from 9.6% for 1970 to 1979 to 30.3% for 1990 to 1999. Deaths from cancer and stroke also increased significantly over the three decades from 1.2% and 2.4%, respectively, for 1970 to 1979, to 13.2% and 8.0%, respectively, for 1990 to 1999. CONCLUSIONS: The causes of graft loss and death have changed over the last three decades. By better addressing the main causes of death, cardiac disease, and stroke with better prevention, graft loss due to death with a functioning graft will be reduced. PMID- 12131690 TI - Higher frequency of high-grade rejections in cardiac allograft patients after Quilty B lesions or grade 2/4 rejections. AB - BACKGROUND: To better understand how different histologic patterns of allograft inflammation found on biopsies of human cardiac allografts progress to high-grade rejection, we undertook a statistical analysis of our institutional database to detect statistical patterns among different types of myocardial allograft inflammations found on sequential biopsies. METHODS: Biopsies were analyzed for statistical associations between high-grade rejections (International Society of Heart and Lung Transplantation [ISHLT] grade > or = 3A/4) and the type of cardiac allograft inflammation found on prior biopsies. Case cross-over and case control designs were used to compare the antecedent patterns of inflammation on biopsies with high-grade rejection compared to biopsies with low-grade rejection, all within the same subject. Quilty lesions were correlated with cyclosporine levels. RESULTS: Patients with Quilty B lesions or ISHLT grade 2/4 rejections show an increased risk for high-grade rejection on their next biopsies (Odds ratio 5.9 to 11.2). The presence of two pathological findings, especially Quilty B and grade 2/4 rejection, creates additional risk in excess of that found independently (Odds ratio >14). Quilty lesions are found only in cardiac allografts, and do not correlate with trough cyclosporine levels. CONCLUSIONS: The morphological patterns of several types of human cardiac allograft inflammation found on sequential protocol biopsies are not randomly associated. Patients with grade 2/4 rejections and Quilty B lesions show an increased risk for high-grade rejections on their next biopsies. Quilty B lesions, similar to ISHLT grade 2/4 rejections, may represent subclinical rejection. Both are more likely to progress to a high grade rejection. PMID- 12131691 TI - Bladder carcinoma in a transplant recipient: evidence to implicate the BK human polyomavirus as a causal transforming agent. AB - The BK polyomavirus (BKV) infects most of the human population, but clinically relevant infections are mostly limited to individuals who are immunosuppressed. In transplant recipients, BKV has been associated with ureteral stenosis, interstitial nephritis, and hemorrhagic cystitis. The role of BKV in the development of human tumors is intriguing but uncertain. BKV has been identified in various tumor types including urothelial carcinoma, but the ubiquitous presence of BKV as a latent infection has confounded efforts to validate any causal role in cancer development. We report the case of a simultaneous pancreas and kidney transplant recipient who developed BKV interstitial nephritis and carcinoma of the bladder with widespread metastases. High level expression of BKV large T antigen in the primary and metastatic carcinoma, but not in the nonneoplastic urothelium, implicates BKV as an etiologic agent in the development of this tumor. PMID- 12131692 TI - Acute cellular rejection in liver transplant recipients under cyclosporine immunosuppression: predictive factors of response to antirejection therapy. AB - BACKGROUND: Predictive factors of response to antirejection therapy in acute cellular rejection (ACR) in liver transplantation are not well established. METHODS: To investigate the possible existence of these factors, we reviewed 111 consecutive episodes of ACR fulfilling the following criteria: histologically confirmed ACR; cyclosporine-based immunosuppression; initial antirejection treatment with high-dose steroid boluses; minimum follow-up of 2 weeks after treatment; and no other graft complication interfering with evaluation of therapeutic response. ACR episodes not responding to initial steroid therapy were given additional treatment (OKT3 and/or repeated steroid boluses). We analyzed the association of the response to the antirejection treatment with different clinical, laboratory, histological, and donor-recipient compatibility variables at two times: after the initial antirejection therapy, and after all the antirejection therapy administered. RESULTS: Eighty episodes of ACR (72%) resolved after the initial therapy with high-dose steroid boluses, and another 18 (16%), initially steroid-resistant, resolved with additional antirejection treatment. Thirteen episodes (12%) were refractory to all antirejection treatment administered. Variables with independent predictive value of nonresponse to initial therapy with steroid boluses were late-onset ACR (>2 months after transplantation), high serum bilirubin and alanine aminotransferase, low blood cyclosporine concentration in the week before antirejection treatment, and severe histological endothelialitis. Late-onset ACR and high serum bilirubin were also independent predictors of refractoriness to all the treatment administered. CONCLUSIONS: Response to antirejection treatment in ACR in liver transplantation can be predicted by several clinical and laboratory data. ACR episodes with factors predictive of therapeutic unresponsiveness could benefit from more aggressive antirejection treatment. PMID- 12131693 TI - Successful tolerance induction under CD40 ligation in a rodent small bowel transplant model: first report of a study with the novel antibody AH.F5. AB - BACKGROUND: Intestinal transplantation has been hampered by high rates of intestinal allograft rejection. One mechanism of altering rejection in other organ transplant models has been blockade of second set T-cell costimulatory signals. AH.F5, a novel hamster anti-rat monoclonal antibody to CD154, blocks CD40-dependent T-cell costimulation. We hypothesized that blockade of this pathway might abrogate rejection in a rodent orthotopic survival model of intestinal transplantation. METHODS: Eight groups were studied with different dosing schema, including syngeneic transplants (group 1), untreated allogeneic transplants (group 2), allogeneic transplants plus multiple doses of AH.F5 alone given IV or s.c. (groups 3 and 4), allogeneic transplants plus donor splenocyte preconditioning with and without single dose AH.F5 (groups 5 and 6), and donor splenocyte preconditioning followed by multiple doses of AH.F5 with and without thymectomy (groups 7 and 8). RESULTS: Control animals all died within 12 days of transplantation, whereas antibody-alone and splenocytes-alone resulted in modest prolongation of survival to 16 days. Only animals treated with splenocytes before transplantation and AH.F5 survived long-term (>60 days, group 8). These animals tolerated donor-specific skin grafts, rejected third-party grafts, and fed normally. However, their weight gain was subnormal and they demonstrated intestinal muscular thickening, which might represent chronic rejection. Thymectomy prevented the induction of tolerance. CONCLUSIONS: AH.F5 prevents acute intestinal allograft rejection in combination with donor-specific splenocyte preconditioning. We achieved long-term survival and the animals appeared tolerant. Central conditioning is essential for success with this antibody when used alone. Further studies with different dosing regimens or second agents seem warranted. PMID- 12131694 TI - A CD200FC immunoadhesin prolongs rat islet xenograft survival in mice. AB - BACKGROUND: A solubilized form of the CD200 molecule, CD200Fc, has been shown to suppress allograft rejection and development of collagen-induced arthritis in mice. We investigated whether the same molecule could prolong survival of rat islet xenografts. METHODS: Streptozocin-treated mice, receiving injections with anti-asialo-GM1 antibody, received rat islets ( approximately 400/mouse) under the kidney capsule or injected into the portal vein, along with rapamycin treatment. Thereafter mice received injections of CD200Fc (10 microg/mouse/injection) or control mouse IgG2. Blood glucose was monitored daily. Some mice received additional injections of anti-CD200/-CD200R monoclonal antibodies. RESULTS: Portal vein delivery of islets led to more extended resolution of diabetes than did transplantation under the kidney capsule. CD200Fc further prolonged survival in either case, an effect abolished by anti-CD200 or F(ab')2 anti-CD200R mAbs, but not by whole anti-CD200R (anti-CD200R Ig). Spleen cells taken from CD200Fc-treated mice showed polarization to type-2 cytokine production (interleukin-4, interleukin-10) on restimulation with rat splenocytes in culture, in comparison to cells from control mice (type-1 cytokines, interlulin-2, interferon-gamma). CONCLUSION: CD200:CD200R interactions are important in regulating rat islet xenograft survival. PMID- 12131696 TI - OKT3 (muromonab-CD3) associated hepatitis in a kidney transplant recipient. AB - The introduction of OKT3 (muromonab-CD3) revolutionized the management of steroid resistant rejection in transplant patients. Indeed, after the success of OKT3 for treatment of rejection, some centers used OKT3 for perioperative prophylaxis against rejection after transplantation. Despite the success of this agent for prevention and treatment of rejection, its use is associated with side effects. These side effects, including the cytokine release syndrome and flash pulmonary edema, are well recognized in the transplant community. However, there have been no published reports of acute severe hepatitis associated with the use of OKT3 in non-liver transplant patients. We describe here a case of acute severe hepatitis encountered during treatment of acute renal allograft rejection with OKT3 in a 31 year-old black man. PMID- 12131695 TI - Peritransplant streptavidin recipient treatment prolongs rat cardiac allograft survival. AB - BACKGROUND: Because streptavidin shows high localization in inflamed tissues, it might also interfere with the proliferation of cells involved in allograft rejection. METHODS AND RESULTS: Treatment of naive ACI recipients with 20 mg/kg streptavidin i.p. alone significantly prolonged Lewis cardiac allografts from a mean survival time of 9.8+/-0.7 days in controls to 19.8+/-6.5 days, with one recipient accepting the graft permanently (>250 days). Peritransplant streptavidin treatment combined with 0.5 ml of antilymphocyte serum (ALS) transient immunosuppression led to permanent graft survival (>250 days) in 6 of 10 recipients. Second-set skin grafts performed 60 days after the primary cardiac allograft were prolonged to 45 days, whereas the third party Wistar-Furth (WF) skin grafts were rejected in 15 days without the rejection of the primary Lewis cardiac allografts. Pathology of transplanted cardiac allografts at 100 days showed no mononuclear cell infiltration or chronic allograft vasculopathy. Streptavidin given for 5 days at 20 mg/kg caused a moderate initial weight loss but had no effect on hematologic, biochemical, and histologic parameters in the treated recipients. CONCLUSION: This study demonstrates that peritransplant recipient treatment with streptavidin combined with peritransplant ALS induces prolonged cardiac and second-set skin allograft survival. We conclude that recipient peritransplant streptavidin treatment may provide a new strategy for the induction of transplant tolerance. PMID- 12131697 TI - Intraportal infusion therapy as a novel approach to adult ABO-incompatible liver transplantation. AB - BACKGROUND: ABO-incompatible liver transplantation is associated with an extremely complicated postoperative course, especially when the recipients are adults. METHODS: Two adult patients underwent living-donor liver transplantation from ABO-incompatible donors. The antirejection therapy included multiple perioperative plasmapheresis, splenectomy, systemic triple immunosuppressive regimen with tacrolimus, methylprednisolone, and cyclophophamide, or azathioprine. In addition to these conventional approaches, we performed intraportal infusion therapy after transplantation with methylprednisolone, prostaglandin E1, and gabexate mesilate. RESULTS: With our protocol, antidonor blood group antibody titers in both cases remained low without any evidence of rejection or vascular complications throughout the postoperative course. Biliary complications were transient and resolved completely. The patients have now survived 30 and 12 months posttransplantation and have regained normal life activity with good liver function. CONCLUSIONS: Our experience has shown the feasibility of controlling rejection and other complications in adult ABO incompatible liver transplantation under intraportal infusion therapy. PMID- 12131698 TI - Itraconazole-induced rhabdomyolysis and acute renal failure in a heart transplant recipient treated with simvastatin and cyclosporine. AB - BACKGROUND: Statins are widely used to decrease cholesterol and improve morbidity and mortality associated with coronary artery disease. Myopathy constitutes a rare but potentially life-threatening adverse reaction, which is related to plasma HMG-CoA reductase inhibitory activity. Therefore, the incidence of rhabdomyolysis increases dramatically when statins are co-administered with drugs that inhibit their hepatic transformation, such as cyclosporine or azoles. METHODS AND RESULTS: We present a case of severe rhabdomyolysis and acute renal failure induced by itraconazole in a heart transplant recipient chronically treated with cyclosporine and simvastatin. The literature with regard to the pathogenetic mechanisms and the clinical implications are reviewed. CONCLUSIONS: To avoid severe myopathy, cyclosporine levels should be monitored sooner than weekly intervals and statins should be discontinued or their dosage should be reduced, as long as azoles need to be prescribed in transplant recipients. Rhabdomyolysis and acute renal insufficiency should be promptly recognized and aggressively treated. PMID- 12131699 TI - Subclinical rejection in tacrolimus-treated renal transplant recipients. AB - BACKGROUND: Subclinical rejection, defined as histologic acute rejection in the absence of graft dysfunction, has been suggested as a cause of chronic allograft rejection. In cyclosporine-treated patients, the incidence of subclinical rejection 3 months after transplant is reported to be approximately 30%. The intent of our study was to determine the incidence of subclinical rejection in tacrolimus-treated renal allograft recipients. METHODS: We prospectively studied the incidence of subclinical rejection on surveillance biopsies performed 3 months after transplantation in 114 patients transplanted between September 1, 1998 and November 30, 2000. All patients received tacrolimus, mycophenolate mofetil, and prednisone, and 56% received antibody induction. RESULTS: Subclinical rejection was detected in 2.6% of patients (3/114, 95% confidence interval 0.5-7.5%). Borderline changes were detected in 11% (12/114). Subclinical rejections were treated with bolus methylprednisolone. CONCLUSIONS: The incidence of subclinical rejection early after kidney transplantation is extremely low in tacrolimus-treated patients in whom early rejections are aggressively treated, suggesting that surveillance biopsies may not be necessary with this regimen. PMID- 12131700 TI - Case report of combined pediatric heart-lung-liver transplantation. AB - We present the youngest successful patient with combined heart-lung-liver transplantation. The patient was a 2.5-year-old child with Alagille syndrome suffering from tetralogy of Fallot with pulmonary atresia and multiple aortopulmonary collaterals and familial cholestasis. PMID- 12131701 TI - Living-related right lobe liver transplantation for a patient with fulminant hepatic failure during the second trimester of pregnancy: report of a case. AB - A 28-year-old pregnant Japanese woman developed fulminant hepatic failure (FHF) with coma grade IV at 15 weeks' gestation and underwent emergency orthotopic living-related liver transplantation (LRLT) using the right hepatic lobe of her father. Blood type was identical. On postoperative day 2, she regained consciousness and was extubated. For fear of possible negative effects of exposure to various drugs and from x-ray examinations on the fetus as well as the maternal burden of a continuing pregnant state on the patient, artificial abortion was a treatment choice in this woman on posttransplant day 31. The patient was discharged and is currently doing well. Until the present, 11 pregnant women were reported to have liver transplantation during the second trimester of pregnancy, including 2 pregnant women with LRLT. This is the third case of LRLT, and the first successful case in which the right hepatic lobe was used for graft. PMID- 12131702 TI - Multidetector computed tomographic cholangiography in the evaluation of potential living liver donors. AB - BACKGROUND: Lacking awareness of biliary variations causes complications in adult living donor liver transplantation. The study was performed to determine the diagnostic value of preoperative multidetector computed tomographic cholangiography (MDCT-CA). METHODS: MDCT-CA after the intravenous administration of meglumine iodipamide was performed in 12 potential liver donors. MDCT angiography was added to depict the topographic relationship between biliary and vascular structures. MDCT findings were correlated with intraoperative findings (n=7). RESULTS: MDCT-CA was diagnostic in all 12 patients. Nine patients revealed variants in biliary anatomy: drainage from liver segment four into right hepatic duct (n=4), additional segmental ducts draining into common (n=4) or left hepatic duct (n=2), and trifurcation at the upper confluence (n=1). Biliary vascular topography was variable and well depicted. Intraoperative assessment confirmed the preoperative findings. CONCLUSIONS: Variations in biliary anatomy appear to be the rule rather than the exception. MDCT-CA represents a noninvasive means for accurately assessing biliary morphology. PMID- 12131703 TI - Successful conversion from cyclosporine to tacrolimus for gastric motor dysfunction in a lung transplant recipient. AB - BACKGROUND: Conversion after transplantation from cyclosporine to tacrolimus is often performed because of recurrent acute/chronic rejection or unacceptable side effects such as nephrotoxicity, arterial hypertension, and cosmetic disorders. Although gastrointestinal discomfort is often reported after transplantation, it is usually not considered a sufficient reason for conversion, although tacrolimus seems beneficial with regards to gastric motor activity in renal transplant patients. METHODS: A lung transplantation was performed in a 41-year-old woman with alpha-1 antitrypsin deficiency emphysema. Because the patient presented severe symptoms of nausea, vomiting, and dyspepsia, without obvious endoscopic explanation, that resulted in highly variable cyclosporine trough levels, she was converted from cyclosporine to tacrolimus. RESULTS: After conversion, dyspepsia, nausea, and vomiting resolved. Neurological complications caused by a transient high trough level of tacrolimus resolved completely upon dose reduction with tacrolimus trough levels remaining very stable afterwards. CONCLUSION: Tacrolimus may be the immunosuppressant of choice after solid organ transplantation in patients with problems related to gastric motor dysfunction. PMID- 12131704 TI - Preemptive therapy for cytomegalovirus with oral ganciclovir after liver transplantation. PMID- 12131705 TI - Bed rest or normal activity for patients with acute low back pain: a randomized controlled trial. AB - BACKGROUND: The management of common low back pain has two principal objectives: to relieve acute pain and to attempt prevention of transition to chronicity. Several studies have shown the ineffectiveness of prolonged periods of bed rest. OBJECTIVE: To compare 4 days of bed rest with continued normal daily activity in acute low back pain, taking into account the type of work (physical or sedentary labor). METHODS: This open, comparative multicenter study enrolled 281 ambulatory patients, ages 18 to 65 years, with low back pain (onset < 72 hours). The subjects did not have pain radiating below the buttocks and did not have work related injuries. They were randomized into two treatment groups: one instructed to continue normal activity (insofar as the pain allowed), and the other prescribed 4 days of bed rest. After inclusion, patients were seen at three visits: on day 6 or 7, after 1 month, and after 3 months. RESULTS: On day 6 or 7, pain intensity was similar for both groups, as was the overall judgment of the treatment by patients and physicians. At 1 and 3 months, the groups again had equivalent intensity of back pain, functional disability, and vertebral stiffness. A higher proportion of patients in the bed rest group than in the normal activity group had an initial sick leave (86% vs 52%; P < 0.0001). This difference was greater for the patients whose work was sedentary. CONCLUSIONS: For patients with acute low back pain, normal activity is at least equivalent to bed rest. The findings of this study indicate that prescriptions for bed rest, and thus for sick leaves, should be limited when the physical demands of the job are similar to those for daily life activities. PMID- 12131706 TI - Anatomic suitability of the C1-C2 complex for pedicle screw fixation. AB - STUDY DESIGN AND OBJECTIVES: A computed tomography (CT) study of 60 consecutive patients (120 sides) was performed to assess suitability for either transarticular or pedicle screw fixation. SUMMARY OF BACKGROUND DATA: A C1 lateral mass and C2 pedicle screw fixation with a rigid cantilever beam system has been described. The anatomic constraints relevant for this technique have not. METHODS: Fifty consecutive patients underwent standard CT of the cervical spine. Pedicle and transarticular screw trajectories were plotted, and the maximum safe diameter for screw placement was determined for each trajectory. Also, trajectories were plotted in 10 additional patients with known craniocervical junction abnormalities using three-dimensional (3-D) imaging and computer-aided navigation tools. Screw placement was considered feasible if a 4 mm diameter trajectory could be plotted without impingement on neural or vascular structures. RESULTS: Four-millimeter diameter pedicle screws could be placed in 91 of 100 C2 pedicles in the CT studies and in 20 of 20 pedicles in the 3-D studies. Four-millimeter diameter C1-C2 transarticular screws could be placed in 94 of 100 sides in the CT study and in 19 of 20 sides in the 3-D study. Four sides could tolerate a C2 pedicle screw and not a transarticular screw; the opposite situation existed in five sides. Placement of screws into C1 was not an issue in any patient. The mean maximum diameter of potential transarticular screws was 6.5 mm, and the mean maximum diameter of the pedicle screws was 5.3 mm (P < 0.01). CONCLUSIONS: C1-C2 pedicle screw fixation is a technique that appears to be widely applicable and may represent an alternative fixation technique in selected patients. PMID- 12131707 TI - Does the morphology of the iliolumbar ligament affect lumbosacral disc degeneration? AB - STUDY DESIGN: An anatomic study of the iliolumbar ligament was performed in association with lumbosacral disc degeneration. OBJECTIVES: To determine whether the morphology of the iliolumbar ligament contributes to lumbosacral disc degeneration. SUMMARY OF BACKGROUND DATA: There have been few reports concerning the clinical significance of the iliolumbar ligament. METHODS: We dissected 25 male and 27 female cadavers and measured the length and cross-sectional area of the anterior and posterior bands of the iliolumbar ligament. The specimens were classified into three groups based on the grade of L4-L5 and L5-S1 disc degeneration: the L4-L5 disc was more degenerated than the L5-S1 disc (group L), the L4-L5 disc was less degenerated than the L5-S1 disc (group S), and both discs were equally degenerated (group E). The results were statistically compared among the three groups. RESULTS: The length of the posterior bands and the summation of the length of the anterior and posterior bands were significantly shorter in group L than in group S, and the cross-sectional area of the posterior bands and the summation of the cross-sectional area of the anterior and posterior bands were significantly larger in group L than in groups S or E in the male cadaver specimens. CONCLUSION: If the iliolumbar ligaments (especially the posterior band of the ligament) of a male patient are short and have a large cross-sectional area, the lumbosacral junction can be stabilized by the ligaments, with the L5-S1 disc being protected from degeneration. The L4-L5 disc may be prone to degeneration. PMID- 12131708 TI - Animal models used in spinal cord regeneration research. AB - STUDY DESIGN: A literature review was conducted. OBJECTIVES: To review animal models and injury paradigms used in the neurobiologic study of spinal cord regeneration, and to assist the spinal clinician in interpreting the many encouraging reports of potential therapies emerging from basic science laboratories. SUMMARY OF BACKGROUND DATA: An enormous amount of interest in spinal cord regeneration research has been generated within the past 20 years with the hope that experimental therapies will become available for individuals with spinal cord injuries. The use of various animal models in the laboratory setting has been critical to the development of such experimental therapies. METHODS: A literature review was conducted. RESULTS: Experimental interventions in animal models of spinal cord injury were evaluated both anatomically and functionally. Anatomic assessments use various histologic techniques and frequently include the use of anterograde and retrograde axonal tracers. Functional assessments can be performed neurophysiologically or by the observation of motor and sensory performance on a number of different tests. Sharp spinal cord injury paradigms in which the cord is completely or partially transected are useful for assessing axonal regeneration anatomically. In contrast, blunt injury models in which the cord is compressed or contused more accurately mimic the typical human injury and provide a good setting for the study of secondary pathophysiologic processes immediately after injury. CONCLUSIONS: Animal models will continue to play a critical role in the development of experimental therapies for spinal cord injuries. Both sharp and blunt spinal cord injury paradigms have unique characteristics that make them useful in addressing slightly different neurobiologic problems. PMID- 12131709 TI - Chemokine profile of herniated intervertebral discs infiltrated with monocytes and macrophages. AB - STUDY DESIGN: Herniated lumbar disc specimens were analyzed using reverse transcriptase-polymerase chain reaction to determine the profile of chemokine expression. OBJECTIVE: To investigate the mechanism underlying the recruitment of inflammatory cells into herniated discs during the process of spontaneous regression. SUMMARY OF BACKGROUND DATA: Spontaneous regression of herniated intervertebral discs has been increasingly reported. Although macrophages are suggested to play a central role in this process, it remains unclear how these macrophages accumulate in the herniated discs. METHODS: RNA was extracted from 36 surgical specimens of the herniated lumbar disc, a disc specimen of idiopathic scoliosis and pyogenic spondylitis, and activated peripheral blood mononuclear cells of a normal donor. The RNA was reverse transcribed, and the resultant cDNA was amplified by PCR using primer pairs specific to the CXC chemokines (IL-8, MGSA-alpha, IP-10, MIG), the CC chemokines (MCP-1, MCP-2, MCP-3, MCP-4, MIP 1alpha, MIP-3alpha, RANTES, STCP-1), the C chemokine (lymphotactin), and the glyceraldehyde phosphate housekeeping gene. Thin cryostat sections also were made from the disc specimens and stained with hematoxylin and eosin. RESULTS: All the chemokines examined except MCP-4 were expressed by activated peripheral blood mononuclear cells. Glyceraldehyde phosphate was detected in 8 of 36 herniated discs and in 1 disc specimen each of idiopathic scoliosis and pyogenic spondylitis. Chemokine expression was examined for these 10 disc specimens. From among the 13 chemokines examined, MCP-3, MCP-4, RANTES, and IP-10 were detected in the disc from the idiopathic scoliosis, and MCP-3, MCP-4, RANTES, IP-10, MIG, and MGSA-alpha were detected in the infected or herniated discs. Histologic analysis showed infiltration of inflammatory cells in the infected disc and all 8 herniated discs. CONCLUSIONS: The findings suggest that chemoattractive properties exist in a selected population of human intervertebral discs, and that unique sets of chemokinesplay a role in spontaneous regression of these herniated disc tissues. PMID- 12131712 TI - Fas ligand exists on intervertebral disc cells: a potential molecular mechanism for immune privilege of the disc. AB - STUDY DESIGN: Rat and human intervertebral disc specimens were examined immunohistochemically. Reverse transcription polymerase chain reaction (RT-PCR) analysis was also performed on rat disc tissue to demonstrate the existence of Fas ligand. OBJECTIVE: To clarify the existence of Fas ligand on intact intervertebral disc cells. SUMMARY OF BACKGROUND DATA: The nucleus pulposus has been reported to be an immune-privileged site. The immune-privileged characteristic in other tissues such as the retina and testis has been attributed to the local expression of Fas ligand, which acts by inducing apoptosis of invading Fas-positive T-cells. The existence of Fas ligand in normal disc cells has not yet been addressed. METHODS: Skeletally mature SD male rats were killed, and the coccygeal discs were harvested. Human disc specimens were obtained from idiopathic scoliosis patients during surgical procedures. Immunohistochemical staining for Fas ligand was performed for cross-sections of the discs by standard procedures. Reverse transcription polymerase chain reaction analysis was also carried out to demonstrate Fas ligand mRNA expression on rat intervertebral discs. Testes of the rats were used for positive controls, and muscles were used for negative controls. The sections were observed by light microscopy. RESULTS: The nucleus pulposus cells exhibited intense positive immune staining for Fas ligand. The outer anulus fibrosus cells and notochordal cells exhibited little immunopositivity. The positive controls exhibited positive immune staining, and the negative control showed no immunopositivity. The result of RT-PCR confirmed the existence of Fas ligand in disc cells. The human nucleus pulposus cells showed a similar predilection to rat disc cells. CONCLUSIONS: We demonstrated the existence of Fas ligand on disc cells, which should play a key role in the potential molecular mechanism to maintain immune privilege of the disc. Immune privilege and Fas ligand expression of the intervertebral disc may provide a new insight for basic science research as well as clinical treatments for disc degenerative diseases, including disc herniation with radicular pain. PMID- 12131710 TI - Changes in expression of voltage-dependent ion channel subunits in dorsal root ganglia of rats with radicular injury and pain. AB - STUDY DESIGN: Changes in expression of voltage-dependent ion channel subunits were examined in the radicular pain state. Furthermore, antinociceptive effects of gabapentin on radicular pain were compared with effects on peripheral neuropathic pain. OBJECTIVES: To clarify molecular substrates involved in the development of radicular pain, and to investigate the responsiveness of radicular pain to gabapentin. SUMMARY OF BACKGROUND DATA: Peripheral nerve injuries are known to induce dynamic changes of voltage-dependent Na+ and Ca2+ channel subunits expression in dorsal root ganglion neurons. However, the expression profiles of Na+ and Ca2+ channel subunits in the radicular pain state have not been examined. METHODS: Two radicular pain models and one peripheral neuropathic pain model were prepared. By using semiquantitative reverse transcriptase polymerase chain reaction, the expression levels of several Na+ and Ca2+ channel subunits in the dorsal root ganglions of these pain model rats were investigated. The antinociceptive effects of gabapentin were examined in a behavioral study using the aforementioned pain models. RESULTS: All three neuropathic pain operations induced comparable mechanical allodynia and thermal hyperalgesia. The upregulation of the Na(v)1.3 Na+ channel and Ca(v)alpha2delta Ca2+ channel subunits was observed only in the peripheral nerve injury model. A downregulation of the Na(v)1.9 channel was observed in all three pain model rats. A lower dose of gabapentin was significantly more effective in alleviating the mechanical allodynia of rats with radicular pain. CONCLUSIONS: The reduction of Na(v)1.9 found in all three models may link to the neuropathic pain state, including radicular pain. The lower sensitivity to gabapentin in rats with peripheral neuropathic pain might be partly explained by the marked upregulation of Ca(v)alpha2delta in the dorsal root ganglions, suggesting that gabapentin may be more effective in radicular pain treatment. PMID- 12131713 TI - The effect of ultrasound on the healing of muscle-pediculated bone graft in spinal fusion. AB - STUDY DESIGN: A rabbit model of posterolateral intertransverse process spine arthrodesis with and without the application of low-intensity ultrasound was used. OBJECTIVE: To determine the effects of low-intensity ultrasound on the healing of muscle-pediculated bone graft. SUMMARY OF BACKGROUND DATA: Earlier animal and clinical studies demonstrated the efficacy of low-intensity ultrasound stimulation in the acceleration of osteogenesis and fracture healing. This is the first study in which the beneficial effects of ultrasound on the healing of muscle-pediculated bone graft in spinal fusion have been assessed. METHODS: In this study, 20 New Zealand rabbits were randomly assigned to two groups to undergo either spinal fusion using muscle-pedicle bone graft with ultrasound (ultrasound group) or muscle-pedicle bone graft without ultrasound (control group). Muscle-pediculated bone grafts were prepared from the posterosuperior iliac crest, erector spine muscle, and internal and external oblique muscle. This graft was placed bilaterally between the L5 and L6 transverse processes. Ultrasound was performed 20 minutes per day over the rabbits' lumbar spine. The rabbits were killed 6 weeks after surgery. The lumbar spines were evaluated radiologically, macroscopically, and histologically. RESULTS: By macroscopic and radiologic findings, fusion was detected in 11 control group rabbits (55%) of the control group, and in 17 ultrasound group rabbits (85%). The fusion success rate for the rabbits treated with low-intensity ultrasound were statistically higher (P <0,05) than that for the control group. Histologic specimens showed increased bone formation in the fusions exposed to ultrasound. Mature fusions were present in rabbits that received ultrasound. CONCLUSION: Low-intensity ultrasound in white rabbits increased the rate and quality of spinal fusion using muscle pediculated bone grafts. PMID- 12131714 TI - Outcome of posterior lumbar interbody fusion versus posterolateral fusion for spondylolytic spondylolisthesis. AB - STUDY DESIGN: This retrospective study analyzed the outcome of 44 patients who had decompression, pedicle screw-rod fixation, and fusion for Grades 1 and 2 spondylolytic spondylolisthesis. OBJECTIVE: To evaluate the outcome of two methods for stabilization and fusion: posterolateral fusion and circumferential fusion involving posterior lumbar interbody fusion and posterolateral fusion for low grades of isthmic spondylolisthesis. SUMMARY OF BACKGROUND DATA: It has been suggested that stabilization with instrumented fusion is somewhat unpredictable due to lack of anterior support. Does circumferential fusion using posterior lumbar interbody fusion circumvent all the problems, and is it better than posterolateral fusion clinically? METHODS: A single surgeon treated 21 patients with instrumented posterolateral fusion and 23 patients with instrumented circumferential fusion, (i.e., posterior lumbar interbody fusion, and posterolateral fusion. These two groups were compared for clinical outcome, fusion rate, and correction of slippage. RESULTS: The minimum follow-up period for the patients was 2.1 years. The clinical satisfactory outcome on the Oswestry index was 81% for posterolateral fusion and 69% for posterior lumbar interbody fusion. The subjective outcome was 86% and 65%, respectively, for the two groups (P > 0.05). However, a consideration of subjective scores showed that three patients (14.3%) in posterolateral fusion group and eight patients (34.8%) in posterior lumbar interbody fusion group had an unsatisfactory outcome (P = 0.0135), which was very significant. There were two nonunions in the posterolateral fusion group and none in the posterior lumbar interbody fusion group (P > 0.05). The correction of slippage and the loss of correction at the last follow-up assessment was better in the posterior lumbar interbody fusion group, although this was not statistically significant (P > 0.05). Radicular symptoms and neurologic improvement were statistically similar between the two groups. CONCLUSIONS: Posterolateral fusion has a better clinical outcome in low grades of isthmic spondylolisthesis, although posterior lumbar interbody fusion is more predictable in maintaining correction and achieving union. Careful patient selection is needed for each operation, and adjacent level disc degeneration may influence the procedure offered to the patient. PMID- 12131715 TI - Patient and parental perception of adolescent idiopathic scoliosis before and after surgery in comparison with surface and radiographic measurements. AB - STUDY DESIGN: This prospective 2-year follow-up study evaluated patients treated surgically for adolescent idiopathic scoliosis (AIS). OBJECTIVE: To report parents' perception, patients' perception, and pain and disability before and after surgery and to examine their relationship to anthropometric, back surface, and radiographic measurements. SUMMARY OF BACKGROUND DATA: No longitudinal studies have examined these factors and their interrelationships. METHODS: Between 1995 and 1999, 39 AIS patients treated by anterior or posterior USS (Universal Spine System, Stratec, Oberdorf, Switzerland) instrumentation had complete prospective questionnaire, back surface, and radiographic appraisal. RESULTS: The preoperative Visual Analogue Score (VAS) for pain-predominantly mild backache-was 24 mm (range, 0-78 mm), and the Oswestry Disability Index (ODI) score was 9.2% (0-44.4%). Patients and parents wanted surgery to correct spinal curvature, stop curve progression, and correct the rib-hump (thoracic) and hip and waist asymmetry (thoracolumbar curves). The maximum angle of trunk inclination correlated with VAS and with short-form McGill Pain Questionnaire scores for thoracic curves (P = 0.005, Spearman rank correlation coefficient). Apical vertebral translation correlated with short-form McGill scores and ODI for thoracolumbar curves (P < 0.006, Spearman rank correlation coefficient). Parents rated scoliosis problems more severely than did their children (P < 0.0001, repeated measures of multivariate analysis of variants). There was no change in body image, VAS, ODI, or short-form McGill scores by 2 years' follow-up. Parents and patients perceived scoliosis problems to be less by 2 years' follow-up (P < 0.0005, Wilcoxon matched-pairs signed ranks test). The preoperative surface asymmetry score correlated with the patients' grading of their rib-hump (P = 0.007). CONCLUSIONS: Back pain incidence was higher than reported for healthy adolescents. Oswestry Disability Index was within normal adult limits. Pain varied by curve type, related to the maximum angle of trunk inclination and the maximum apical vertebral translation. After surgery, back pain and ODI were unchanged, but concerns regarding scoliosis were reduced. PMID- 12131717 TI - The use of short and rigid anterior instrumentation in the treatment of idiopathic thoracolumbar scoliosis: a retrospective review of 24 cases. AB - STUDY DESIGN: The results of short anterior rigid rod instrumentation used to treat thoracolumbar scoliosis in 24 patients were reviewed retrospectively. OBJECTIVE: To assess the safety and efficiency of anterior correction of thoracolumbar scoliosis using a single rigid rod with segmental fixation. SUMMARY OF BACKGROUND DATA: This method of treatment results from the historical difficulties noted in obtaining adequate correction in both planes for this particular type of curve while saving as many segments as possible. This approach seems to combine the advantages and avoid the limitations of posterior segmental and previous anterior segmental techniques. METHODS: For this study, 24 patients with thoracolumbar scoliosis underwent anterior spinal correction and fusion using a single rigid rod and segmental fixation. Additionally, in five patients, this construct was supplemented by interbody titanium cages. The patients were assessed for the amount of correction obtained in the main curve, and for the spontaneous correction of the compensatory curves above and below. Trunk balance was measured on standing radiographs. The average age of the patients was 17.3 years (range, 10-43 years), and the average follow-up period was 3.6 years (range, 2-4.5 years). RESULTS: Correction of the major curve at follow-up assessment was 54%, with a 73% correction of the instrumented area. The upper compensatory curve showed a 21% correction, and the lower curve a 59% correction. Over the duration of the follow-up period, the instrumented area showed kyphosis progression of 6.5 degrees. The five patients who had insertion of Harms cages tended to have a lower rate of correction. The average number of intervertebral mobile segments fused was 2.9 (range, 2-5 segments). No intraoperative complication occurred in relation to the anterior approach. A temporary sympathectomy effect on the side of the approach was noted in most patients. Two radiologic pseudarthrosis and one loss of fixation at the top of the construct were observed. CONCLUSIONS: Anterior correction and fusion using solid rod instrumentation constitute effective and safe treatment of thoracolumbar scoliosis. As compared with posterior systems, it provides correction and rebalance of the trunk through a shorter fusion segment. PMID- 12131718 TI - Accuracy of thoracic pedicle screws in patients with and without coronal plane spinal deformities. AB - STUDY DESIGN: This retrospective observational study evaluated 399 transpedicular thoracic screws using postoperative computed tomography (CT). OBJECTIVES: To examine the in vivo accuracy of transpedicular thoracic screws in patients with and without coronal plane spinal deformities. SUMMARY OF BACKGROUND DATA: There are no comparative studies regarding the safety and accuracy of thoracic pedicle screws in patients with and without coronal plane spinal deformities. METHODS: Curve magnitude and segmental vertebral rotation were determined from preoperative radiographs. Postoperative CT was used to assess the placement accuracy of titanium thoracic pedicle screws. RESULTS: Forty-seven patients underwent instrumented posterior spinal fusion using 399 titanium thoracic pedicle screws. Fully contained screw accuracy in patients with coronal plane spinal deformities was less than in patients without coronal plane spinal deformities at T9-T12 (59% vs. 73%, P = 0.04) and overall (42% vs. 62%, P = 0.001). There was no difference between the overall percentages of acceptably positioned screws (< or = 2 mm of medial or < or = 6 mm of lateral pedicle perforation) in patients with coronal plane spinal deformities (98%) versus patients without coronal plane spinal deformities (99%) (P = 0.69). Penetration of the anterior vertebral cortex was more frequent in patients with coronal plane spinal deformities than in those without coronal plane spinal deformities (8.0% vs. 1.0%, P = 0.008). There was no correlation between the accuracy of screw placement and the degree of segmental rotation, screw proximity to the curve apex, or screw position relative to the curve concavity or convexity (P > 0.12). There were no neurologic or vascular complications. CONCLUSIONS: The overall percentage of acceptably positioned screws was 98% in patients with coronal plane spinal deformities and 99% in patients without coronal plane spinal deformities. In patients with coronal plane spinal deformities, penetration of the pedicle wall and the anterior vertebral cortex was increased at T9-T12 and overall. PMID- 12131719 TI - A quantitative analysis of sensory function in lumbar radiculopathy using current perception threshold testing. AB - STUDY DESIGN: Peripheral sensory functions in patients with radiculopathy resulting from lumbar disc herniation and in control individuals were analyzed using current perception threshold testing. OBJECTIVE: To evaluate the severity of sensory disturbance quantitatively in patients with lumbar radiculopathy. SUMMARY OF BACKGROUND DATA: Subjective evaluation of the severity of sensory disturbance associated with spinal disorders using conventional methods often is difficult. Current perception threshold evaluation is a recently proposed method for studying peripheral nerve dysfunction. This is a quantitative sensory test for analyzing functions of A-beta, A-delta, and C fibers. METHODS: In this study, 48 patients with lumbar radiculopathy resulting from lumbar disc herniation were examined. The mean age of the patients was 37.9 years. All the patients had pain distribution from the compression of one lumbar nerve root (L5 or S1), and unequivocal unilateral disc herniation of the corresponding level was shown by magnetic resonance imaging. Eleven healthy volunteers were used as control subjects. Their mean age was 38.2 years. Current perception threshold evaluation using a Neurometer device was performed at three frequencies: 2000, 250, and 5 Hz. The stimulus sites were located on the dorsal side of the first metatarsus (the L5 dermatome) and the dorsal side of the fifth metatarsus (the S1 dermatome). These sites were investigated on both legs in all the patients and control subjects. The intensity of pain was scored using a visual analog scale. RESULTS: In the control group, there were no significant differences in current perception threshold values at any frequency between the left and right legs. In the patient group, the current perception threshold values in the affected legs were significantly higher than those in the contralateral legs at all frequencies. The current perception threshold values in the affected legs in the patient group were significantly higher than those in the control subjects at 2000 and 250 Hz, whereas there were no significant differences at 5 Hz. The current perception threshold values in the affected legs were significantly higher in patients with hypesthesia than in those without hypesthesia at 2000 and 250 Hz, and in patients with severe pain than in those with less pain at 5 Hz. CONCLUSIONS: Current perception threshold testing showed that the functions of A beta, A-delta, and C fibers deteriorated in patients with lumbar radiculopathy. This technique may be useful for quantifying sensory nerve dysfunction in patients with radiculopathy. PMID- 12131720 TI - Combined spinal cord monitoring using neurogenic mixed evoked potentials and collision techniques. AB - STUDY DESIGN: Neurogenic mixed evoked potentials are used routinely to monitor the spinal cord during spine surgery. This study investigates the differential sensory-motor contribution by using collision techniques. OBJECTIVE: To demonstrate that neurogenic mixed evoked potentials do contain a motor component. SUMMARY OF BACKGROUND DATA: Spinal cord monitoring is now routinely used during spine deformity surgery. Neurogenic mixed evoked potentials (i.e., potentials recorded from lower limb nerves after spinal cord stimulation) represent a reliable and sensitive technique. However, their specificity (sensory and motor spinal pathways) remains debated. METHODS: Neurogenic mixed evoked potentials and collisions were performed in 24 consecutive patients during scoliosis surgery. Neurogenic mixed evoked potentials were elicited by a high thoracic spinal test stimulation and recorded from the tibial nerve at the ankle. A peripheral conditioning stimulation was delivered at the popliteal fossa 15 ms before spinal stimulation, inducing an ascending volley. The antidromic ascending motor component stops at the anterior horn cell level, whereas the orthodromic sensory component reaches the dorsal columns. The 15-ms interstimulus interval between peripheral conditioning and spinal test stimulation makes the collision with descending volleys occur in the spinal cord. The descending sensory volley is blocked, whereas the descending motor volley is unaffected. RESULTS: Reproducible evoked potentials were recorded from the tibial nerve in all the patients studied when the conditioning stimulation was performed. These conditioned neurogenic mixed evoked potentials consist of a small and polyphasic wave whose amplitude represents approximately 26% that of the wave of unconditioned neurogenic mixed evoked potentials. It is likely that they correspond to motor spinal pathway activation. CONCLUSION: Both standard and conditioned neurogenic mixed evoked potentials are proposed to provide combined sensory and motor spinal pathway monitoring. PMID- 12131721 TI - Pedicle screws with high electrical resistance: a potential source of error with stimulus-evoked EMG. AB - STUDY DESIGN: Clinically relevant aspects of pedicle screws were subjected to electrical resistance testing. OBJECTIVES: To catalog commonly used pedicle screws in terms of electrical resistance, and to determine whether polyaxial-type pedicle screws have the potential to create a high-resistance circuit during stimulus-evoked electromyographic testing. SUMMARY OF BACKGROUND DATA: Although stimulus-evoked electromyography is commonly used to confirm the accuracy of pedicle screw placement, no studies have documented the electrical resistance of commonly used pedicle screws. METHODS: Resistance measurements were obtained from eight pedicle screw varieties (5 screws of each type) across the screw shank and between the shank and regions of the screw that would be clinically accessible to stimulus-evoked electromyographic testing with a screw implanted in a pedicle. To determine measurement variability, resistance was measured three times at each site and with the crown of the polyaxial-type screw in three random positions. RESULTS: Resistance across the screw shank ranged from 0 to 36.4 ohms, whereas resistance across the length of the monoaxial-type screws ranged from 0.1 to 31.8 ohms. Resistance between the hexagonal port and shank of polyaxial-type screws ranged from 0 to 25 ohms. In contrast, resistance between the mobile crown and shank of polyaxial-type screws varied widely, ranging from 0.1 ohms to an open circuit (no electrical conduction). Polyaxial-type screws demonstrated an open circuit in 28 of 75 measurements (37%) and a high-resistance circuit (exceeding 1000 ohms) in 5 of 75 measurements (7%). CONCLUSIONS: Polyaxial-type pedicle screws have the potential for high electrical resistance between the mobile crown and shank, and therefore may fail to demonstrate an electromyographic response during stimulus-evoked electromyographic testing in the setting of a pedicle breech. To avoid false-negative stimulus-evoked electromyographic testing, the cathode stimulator probe should be applied to the hexagonal port or directly to the screw shank, and not to the mobile crown. PMID- 12131722 TI - The use of interventional open MRI to assess the kinematics of the lumbar spine in patients with spondylolisthesis. AB - STUDY DESIGN: Open interventional MRI techniques were used to investigate the intervertebral mobility of the lumbar spine in subjects with isthmic and degenerative spondylolisthesis. The findings were compared with those in a published database of subjects with no history of low back pain. OBJECTIVE: To investigate patterns of intervertebral mobility in subjects with spondylolisthesis to determine the level of spinal instability in this population. SUMMARY OF BACKGROUND DATA: Subjects with spondylolisthesis have been considered to present with a special form of spinal instability. Consequently, this condition is frequently managed by spinal fusion. However, confusion exists regarding whether there is excessive motion at the level of the defect. METHODS: For this study, 29 subjects presenting to spinal clinics with spondylolisthesis (15 isthmic and 14 degenerative) were recruited and compared with an existing database of control subjects. The motion characteristics of these subjects in flexed and extended positions were investigated using interventional open MRI of known precision. In all the subjects, the level of resting pain, the grade of slip, and the level of the defect were noted. RESULTS: No mobility differences, in terms of both angular and translational motion, were found between the subjects with spondylolisthesis and those with no history of low back pain, suggesting that subjects with spondylolisthesis do not present with either instability or hypermobility. CONCLUSION: A spondylolytic defect does not lead to detectable instability or hypermobility in the lumbar spine. PMID- 12131723 TI - Re: Combined laminectomy and thoracoscopic resection of dumbbell-type thoracic cord tumor (Spine 2002; 26:E130-4.). PMID- 12131724 TI - Re: Karppinen J. et al. Periradicular infiltration for sciatica. A randomized controlled trial. Spine 26, 1059-1067:2001. PMID- 12131725 TI - Re: Harms J, Melcher P. Posterior C1-C2 fusion with polyaxial screw and rod fixation. (Spine 2001;26: 2467-71). PMID- 12131726 TI - Re: Koes B, van Tulder M, Ostelo R, et al. Clinical guidelines for the management of low back pain in primary care: an international comparison. (Spine 2001;26:2504-14). PMID- 12131727 TI - Re: Safety, efficacy, and cost-effectiveness of evidence-based guidelines for the management of acute low back pain in primary care (Spine 2001;26:2615-22). PMID- 12131728 TI - Re: Koch et al. Adolescents undergoing surgery for idiopathic scoliosis: how physical and psychological characteristics relate to patient satisfaction with cosmetic result. (Spine 2001;26:2119-24). PMID- 12131729 TI - Thoracic vertebral osteophyte-causing myelopathy: early diagnosis and treatment. AB - STUDY DESIGN: A case report is described. OBJECTIVE: To highlight an unusual cause of thoracic myelopathy. METHODS: Clinical evaluation of 63-year-old male revealed myelopathy. Thoracic cord compression from a solitary projection of a facetal joint osteophyte at the T9-T10 level was documented on magnetic resonance imaging scans and computed tomography scans. RESULTS: The osteophyte was successfully excised. CONCLUSIONS: Thoracic cord compression can be caused by various space-occupying lesions, and a high index of suspicion will lead to diagnosis before neurologic deficit is clinically expressed. Magnetic resonance imaging scans and computed tomography scans both demonstrate the osteophyte, and expedient surgery avoids the progression of the neurologic deficit. PMID- 12131730 TI - The destroyed lung syndrome: report of a case after Harrington rod instrumentation and fusion for idiopathic scoliosis. AB - STUDY DESIGN: A case report is described. OBJECTIVE: To describe the very rare complication of destroyed lung syndrome after scoliosis correction. SUMMARY OF BACKGROUND DATA: The destroyed lung syndrome has, to our knowledge, never been associated with scoliosis in the literature. Bronchial kinking and compression by the vertebral column have been described in severe scoliosis cases. METHODS: The patient, a 40-year-old woman was operated on in 1976 for a thoracic scoliosis and hypokyphosis using Harrington rod instrumentation and fusion with autologous bone graft. With a follow-up of 26 years, she has developed a very severe functional defect of the right lung, the so-called destroyed lung syndrome. RESULTS: After the index procedure, the patient developed various episodes of pneumonia and abscess formation in the right lung because of kinking and obstruction of the bronchial tree of the right lung. This seemed to be caused by a severe hypokyphosis and by residual scoliosis of the thoracic spine with direct compression of the right bronchus by the vertebral column. Eventually two stents were placed, but this prevented further deterioration only temporarily. CONCLUSIONS: After Harrington instrumentation and fusion for thoracic hypokyphotic idiopathic scoliosis, kinking and obstruction of a main bronchus are possible. In this patient, this complication gave rise to recurrent infections of the right lung, eventually progressing to destroyed lung syndrome. PMID- 12131731 TI - Irreducible thoracic spondyloptosis in a child with neurofibromatosis: a rationale for treatment. AB - STUDY DESIGN: A case report and review of literature are presented. OBJECTIVES: To describe the surgical management of a midthoracic spondyloptosis associated with kyphosis in a child with neurofibromatosis, an extremely rare but a potentially high-morbidity complication. SUMMARY OF BACKGROUND DATA: Dystrophic kyphoscoliotic spinal deformity is the most common orthopedic sequela of neurofibromatosis. Spondyloptosis is a rare complication but with the potential for high morbidity if the diagnosis is missed or undertreated. Reported cases are rare. METHODS: A severe thoracic spondyloptosis occurred in a 7-year-old girl with peripheral neurofibromatosis who presented with transient paraparesis after a fall. The kyphosis was reduced by cantilever correction forces, achieving side to-side (bayonet) apposition rather than anatomic reduction of the spondyloptosis. This was followed by anterior spinal arthrodesis and structural grafting. RESULTS: Two and a half years after the surgery there is no loss of correction, and the patient has remained neurologically recovered. CONCLUSIONS: Posterior correction of the gibbus in a bayonet apposition and stabilization with a two-rod construct followed by anterior spinal arthrodesis and structural grafting seem to offer efficient surgical treatment. PMID- 12131732 TI - Combined magnetic fields accelerate and increase spine fusion: a double-blind, randomized, placebo controlled study. AB - STUDY DESIGN: The clinical study conducted was a prospective, randomized, double blind, placebo-controlled trial. OBJECTIVES: The purpose of this study was to evaluate the effect of combined magnetic fields on the healing of primary noninstrumented posterolateral lumbar spine fusion. SUMMARY OF BACKGROUND DATA: Combined magnetic fields, a new type of biophysical stimulus, have been shown to act by stimulating endogenous production of growth factors that regulate the healing process. This is the first placebo-controlled study to assess the effect of an electromagnetic stimulus on primary noninstrumented posterolateral lumbar spine fusion surgery as well as the first evaluation of combined magnetic fields as an adjunctive stimulus to lumbar spine fusion. METHODS: This multicenter investigational study was conducted at 10 clinical sites under an Investigational Device Exemption from the United States Food and Drug Administration. Eligible patients had one-level or two-level fusions (between L3 and S1) without instrumentation, either with autograft alone or in combination with allograft. The combined magnetic field device used a single posterior coil, centered over the fusion site, with one 30-minute treatment per day for 9 months. Randomization was stratified by site and number of levels fused. Evaluation was performed 3, 6, and 9 months after surgery and 3 months after the end of treatment. The primary endpoint was assessment of fusion at 9 months, based on radiographic evaluation by a blinded panel consisting of the treating physician, a musculoskeletal radiologist, and a spine surgeon. RESULTS: Of 243 enrolled patients, 201 were available for evaluation. Among all patients with active devices, 64% healed at 9 months compared with 43% of patients with placebo devices: a significant difference (P = 0.003 by Fisher's exact test). Stratification by gender showed fusion in 67% of women with active devices, compared with 35% of those with placebo devices (P = 0.001 by Fisher's exact test). By contrast, there was not a statistically significant effect of the active device in this male study population. In the overall population of 201 patients, repeated measures analyses of fusion outcomes (by generalized estimating equations) showed a main effect of treatment, favoring the active treatment (P = 0.030). In a model with main effect and a time by treatment interaction, the latter was significant (P = 0.024), indicating acceleration of healing. Performed in the full sample of 243 patients, results of the intent-to-treat analysis were qualitatively the same as in the evaluable sample of 201 patients. DISCUSSION: This investigational study demonstrates that combined magnetic field treatment of 30 min/d increases the probability of successful spine fusion, and statistical analysis using the generalized estimating equations model suggests an acceleration of the healing process. This is the first randomized clinical trial of noninstrumented primary posterolateral lumbar spine fusion, with evaluation by a blinded, unbiased panel. This is the first double-blind study performed to date assessing noninstrumented fusion outcome with extremely critical radiographic criteria. The lower overall fusion rates in this study are attributed to the high-risk patient group with an average age of 57 years, the use of noninstrumented technique with posterolateral fusion only, and the reliance on extremely critical radiographic and clinical criteria and blinded panel for fusion assessment without surgical confirmation. CONCLUSIONS: In conclusion, the adjunctive use of the combined magnetic field device was statistically beneficial in the overall patient population, as has been shown in previous studies of adjunctive bone growth stimulation for spine fusion. For the first time, stratification of fusion success data by gender demonstrated that the female study population responded positively to the adjunctive combined magnetic field treatment, with no statistically significant effect observed in the male study population. Adjunctive use of the combined magnetic field device significantly increased the 9-month success of radiographic spinal fusion and showed an acceleration of the healing process. PMID- 12131734 TI - Laparoscopic anterior lumbar interbody fusion at L4-L5: an anatomic evaluation and approach classification. AB - STUDY DESIGN: An anatomic classification system was devised on the basis of operative reports and preoperative magnetic resonance imaging or computerized tomography from 139 patients who underwent laparoscopic anterior lumbar interbody fusion involving L4-L5. OBJECTIVE: To devise a classification system for laparoscopic exposure of the L4-L5 disc space that would allow prediction of the safest approach for any given vascular configuration. SUMMARY OF BACKGROUND DATA: The laparoscopic technique has gained acceptance at L5-S1 but has been less successful at L4-L5. The vascular bifurcation and the variability of the anatomy have led to difficulties with exposure. METHODS: Data were collected on 139 patients undergoing laparoscopic anterior lumbar interbody fusion involving the L4-L5 disc space. Operative notes and preoperative magnetic resonance imaging and computed tomography scans were reviewed, and a classification system was devised based on the aortic bifurcation and confluence of the left iliac vein with the vena cava. Three variations were identified. Complications, particularly ejaculatory dysfunction, were described. RESULTS: Three classification categories were described. Twenty-five patients (18%) were classified as category A (above the bifurcation of both vessels), 52 patients (37%) were classified as category B (below the bifurcation of both vessels), and 51 patients (37%) were classified as category C (between the left iliac artery and vein). There were 8 (5.8%) intraoperative and 17 (12.2%) postoperative complications. Ejaculatory dysfunction constituted the majority of the postoperative complications, representing 16% of the male population. The incidence of ejaculatory dysfunction correlated with exposure from the left side of the aorta or the left iliac artery. For two-level fusions from L4 to S1, the incidence of ejaculatory dysfunction was 63% for category A but 0% for categories B and C. An alternative approach was suggested for category A: exposing the disc space between the aorta and vena cava. CONCLUSION: The laparoscopic approach to L4-L5 is complicated by the variability of the vascular anatomy encountered during the exposure. Routine magnetic resonance imaging or computed tomography can be used to classify the vascular anatomy and plan the optimal approach. Avoiding the left side of the aorta or the left iliac artery may minimize the risk of ejaculatory dysfunction. PMID- 12131735 TI - Mechanism of intervertebral disc degeneration caused by nicotine in rabbits to explicate intervertebral disc disorders caused by smoking. AB - STUDY DESIGN: The effects of nicotine on intervertebral discs in rabbits were studied experimentally. OBJECTIVES: To investigate the effects of nicotine on the vascular buds in rabbits for elucidating the mechanism of nicotine-induced vertebral disc degeneration. BACKGROUND DATA: Several groups have suggested that cigarette smoking is associated with low back pain, but the exact mechanism is not yet fully understood. METHODS: The pump was filled with a diluted nicotine solution, then implanted under the skin of rabbits for 8 weeks. This model was designed to maintain blood nicotine concentration at approximately 110 ng/mL. Rabbits receiving physiologic saline were used as control animals. RESULTS: Nicotine treatment resulted in necrosis and hyalinization of the nucleus pulposus in all rabbits. The anulus fibrosus showed a disturbance of the pattern of overlapping laminae with and without clefts and separation. These resulted in changes indicative of stenosis of vascular buds and perivascular calcification. Nicotine treatment resulted in hypertrophy of vascular walls, necrotic changes in endothelial cells, and narrowing of the vascular lumen. Nicotine treatment resulted in delineation of vascular buds in the vicinity of the vertebral endplate and a reduction of their numbers. However, the control animals showed a dense vascular network. The number of vascular buds decreased in nicotine treatment. CONCLUSION: The authors believe that both reduction in the density of vascular buds and narrowing of the vascular lumen result in decreased oxygen tension, leading to decreased synthesis of proteoglycan and collagen, thus facilitating degeneration of the disc. PMID- 12131736 TI - Spontaneous production of monocyte chemoattractant protein-1 and interleukin-8 by the human lumbar intervertebral disc. AB - STUDY DESIGN: Scoliotic and herniated human intervertebral disc tissue obtained intraoperatively was cultured, and the medium was analyzed for the production of monocyte chemoattractant protein-1 and interleukin-8. OBJECTIVES: This study was conducted to determine whether the human intervertebral disc is capable of spontaneous production of the chemokines monocyte chemoattractant protein-1 and interleukin-8. SUMMARY OF BACKGROUND DATA: Lumbar disc herniations undergo spontaneous regression with time. This is believed to occur via macrophage mediated phagocytosis of herniated disc material. Monocyte chemoattractant protein-1, a chemotactic agent for macrophages, has recently been identified in rat intervertebral disc tissue. METHODS: Disc material obtained from patients undergoing surgery for scoliosis and sciatica was cultured using a serumless technique, and the medium was subsequently analyzed for levels of monocyte chemoattractant protein-1 and interleukin-8. RESULTS: Monocyte chemoattractant protein-1 and IL-8 were detected in both control and herniated disc specimens. Noncontained herniations produced higher levels of chemokines than those with an intact anulus. CONCLUSIONS: Human intervertebral disc tissue is capable of spontaneously producing the proinflammatory chemokines monocyte chemoattractant protein-1 and interleukin-8. These are chemotactic for macrophages and capillaries and may explain the ingrowth of granulation tissue seen in spontaneous disc herniation resorption. PMID- 12131737 TI - Eosinophilic granuloma of the cervical spine. AB - STUDY DESIGN: A meta-analysis was performed based on 53 cases of cervical eosinophilic granuloma reported in the literature and 1 in an adult treated by the authors. OBJECTIVE: To stress the clinical and radiologic differences between cervical and thoracolumbar spinal eosinophilic granuloma and to point out differences between adults and children with cervical eosinophilic granuloma to avoid false diagnosis. SUMMARY OF BACKGROUND DATA: Until now, cervical eosinophilic granuloma has been reported in 43 children and 9 adults. In 1 case the age is unknown. In previous studies, differences between adults and children with cervical eosinophilic granuloma have not been analyzed, nor has cervical eosinophilic granuloma been compared with thoracolumbar eosinophilic granuloma. METHODS: All reported cases of cervical eosinophilic granuloma were analyzed concerning age and sex distribution, clinical and radiologic presentation, therapy, and outcome. The authors' case in a 46-year-old patient is discussed. RESULTS: The presenting symptoms of cervical eosinophilic granuloma are usually pain and restricted range of motion. In contrast to eosinophilic granuloma of the thoracic spine and lumbar spine, the neurologic symptoms are less frequent, and the first radiographic sign is an osteolytic lesion. Vertebra plana is a rare sign in cervical eosinophilic granuloma. In children, the middle cervical spine is most often affected, whereas in adults it is the second vertebra. The outcome of the patients has been good in most cases, independently of treatment. CONCLUSION: In most cases of cervical eosinophilic granuloma, immobilization is an adequate therapy. If the process continuous to progress, radiotherapy is recommended. Surgical treatment should be reserved for cases with instability or neurologic defects. PMID- 12131738 TI - Does reconstruction of posterior ligamentous complex with extensor musculature decrease axial symptoms after cervical laminoplasty? AB - STUDY DESIGN: The authors retrospectively determined the prevalence of neck and shoulder symptoms (axial symptoms) after expansive laminoplasty with reattachment of spinous process and extensor musculature in patients with cervical myelopathy. OBJECTIVES: To determine the prevalence of both preoperative and postoperative axial symptoms of expansive laminoplasty when they occur after expansive laminoplasty. SUMMARY OF BACKGROUND DATA: Several clinical reports have noted that laminoplasty for cervical myelopathy produces positive clinical outcomes. However, recent reports have pointed out that complications from laminoplasty, such as axial symptoms, may be severe enough to interfere with daily activities. METHODS: The authors used a modified spinous process-splitting laminoplasty, which involved reattaching the spinous process with extensor musculature after enlarging the spinal canal by use of the French window method. Postoperative axial symptoms were investigated in 173 of 214 patients (80.1%) who underwent expansive laminoplasty between January 1989 and December 1998. The patients included 121 men and 52 women, and their average age was 61.5 years. The presence or absence and grade of axial symptoms before and after laminoplasty were investigated. The severity and duration of complications were also recorded, along with differences between age, sex, spinal alignment, and cervical diseases. RESULTS: Neck and/or shoulder stiffness worsened in 15% of the patients and declined in 21%. Neck pain worsened in 10% of the patients and improved in 11%. Neck and/or shoulder stiffness worse than moderate was recognized in 14.4% of the patients. Neck pain worse than moderate was recognized in 5% of the patients. In the 137 patients who had no axial pain before surgery, only 13 patients experienced such symptoms after surgery, and in most cases these symptoms were minimal. In only 1 case, significant postoperative neck pain arose de novo as a result of this surgery. In 88 patients who had no neck and/or shoulder stiffness before surgery, only 16 patients experienced such symptoms after surgery, and in most cases these were minimal. A similar pattern held true for each of the other grades of preoperative axial symptoms. The recovery rate score (Japanese Orthopedic Association) was 47.5 +/- 32.3 in the patients whose axial symptoms were worse than moderate and 60 +/- 28.9 in patients whose axial symptoms were less than mild. This difference was significant (P < 0.05). CONCLUSION: Laminoplasty is an appropriate operation for cervical spondylotic myelopathy and did not, in this study, seem to have any significant influence on the development or resolution of axial symptoms. PMID- 12131739 TI - Cervical kyphosis: predictive factors for progression of kyphosis and myelopathy. AB - STUDY DESIGN: A retrospective study of 13 patients with cervical kyphosis. The authors propose new methods of measuring spinal cord compression and predicting the progression of kyphosis. OBJECTIVES: To ascertain predictive factors for progression of cervical kyphosis and myelopathy. SUMMARY OF BACKGROUND DATA: Cervical kyphosis may be congenital, result from decompression surgery, or occur as a posttraumatic deformity. Although there is the potential for progressive deformity and the development of myelopathy in all these situations, there are few previous reports of predictive factors for progression of cervical kyphosis and myelopathy in patients with cervical kyphosis. METHODS: The authors studied radiographs and magnetic resonance imaging scans of 13 patients with cervical kyphosis, including 9 who had been operated on and had postsurgical secondary kyphosis, and 4 with idiopathic kyphosis without any of the above causes. Compression of the spinal cord at the apex of the cervical kyphosis was evaluated by magnetic resonance imaging of the ratio between the anteroposterior diameter of the medulla-pons junction and the spinal cord at the apex. RESULTS: The mean ratio between the anteroposterior diameter of the medulla-pons junction and the spinal cord at the apex in five patients in whom myelopathy did not develop was 0.37, and was 0.21 in the patients in whom myelopathy developed. Progression of cervical kyphosis was associated with osteophyte formation at the anterior aspect of the vertebral body. CONCLUSION: A ratio below 0.3 between the anteroposterior diameter of the medulla-pons junction and the spinal cord at the apex was a risk factor for cervical myelopathy. One of the most predictable risk factors of progression of the cervical kyphosis was osteophyte formation at the anterior aspect of the vertebral body. PMID- 12131740 TI - A pilot study on the recovery from paresis after lumbar disc herniation. AB - BACKGROUND: Although the existence of a motor defect in discogenic sciatica is a sign of severity, the literature does not provide evidence for an immediate requirement for surgery. OBJECTIVE: To assess the course of sciatica with discogenic paresis and to determine possible prognostic factors for recovery or improvement. STUDY DESIGN: This open prospective multicenter study included patients with discogenic sciatica with paresis that had been developing for less than 1 month and was rated < or =3 on a 5-grade scale. Pain, the strength of 11 muscles, return to work, and analgesic intake were assessed at 1, 3, and 6 months. Recovery and improvement were defined by pain not exceeding 20 mm or < or =50% of the initial pain score and a score of either 5 (recovery) or 4 (improvement) for the weakest muscle at inclusion. RESULTS: Sixty-seven patients were enrolled; 39 (58%) patients were treated surgically and 28 (42%) medically. Surgically treated patients differed from medically treated patients by a higher rate of extruded herniation, a higher number of paretic muscles (6.3 vs. 5; P = 0.051), and a longer course of sciatica (31.4 vs. 17.3 days; P = 0.034). At 6 months, 7 (10.4%) patients were lost to follow-up; 32 (53.3%) had improved, including 18 (30%) recovered, 33 (85%) back to work and having a professional activity, and 22 (39%) still taking analgesics. The only significant difference between recovered and not recovered patients was mean age at inclusion (43 vs. 51 years, P = 0.034). There were no significant differences between improved and not improved patients. Moreover, the outcome was not different in the two treatment groups: there were 17 (53%) improvements in surgically treated patients, including 8 (25%) recoveries, and 14 (56%) improvements in medically treated patients, including 8 (40%) recoveries. CONCLUSION: This pilot study showed no difference between surgical or medical management for recovery or improvement in patients with discogenic paresis. These results need confirmation by a randomized study. PMID- 12131742 TI - Gadolinium diethylenetriaminepentaacetic acid enhancement in magnetic resonance imaging in relation to symptoms and signs among sciatic patients: a cross sectional study. AB - STUDY DESIGN: A cross-sectional descriptive study. OBJECTIVES: Gadolinium enhancement in lumbar magnetic resonance imaging is not used routinely. The current study explored the possible intercorrelations of enhancement patterns with clinical symptoms and signs. SUMMARY OF BACKGROUND DATA: Rim enhancement has been reported to occur in the periphery of disc herniations, and it is thought to represent neovascularization. To the authors' knowledge, the significance of the enhancement in relation to clinical symptoms has not been studied. METHODS: Magnetic resonance imaging of the lumbar spine with intravenous gadolinium diethylenetriaminepentaacetic acid was performed in each patient. Various contrast enhancement parameters and volume of herniation were evaluated, and their correlations with clinical signs and symptoms (straight leg raising, motor defect, Achilles reflex, leg and back pain, disability) were analyzed. RESULTS: The extent of rim enhancement correlated highly significantly with the degree of disc displacement, being most pronounced in the case of sequesters. The duration of sciatic symptoms correlated negatively with enhancement. The clinical symptoms did not correlate significantly with the different enhancement parameters or disc herniation volume. Straight leg raising correlated only slightly with the extent of rim enhancement (P = 0.04) when bulges were excluded. Achilles reflex abnormality correlated significantly with all enhancement parameters for lesions at L5-S1. In the final stepwise logistic regression model, contrast enhancement extent correlated most significantly with abnormal Achilles reflex (P = 0.0002). CONCLUSIONS: Although rim enhancement of disc herniation is thought to represent a beneficial phagocytotic phenomenon, it may also have a harmful effect on the adjacent nerve root. PMID- 12131743 TI - Segmental recording of cortical motor evoked potentials from thoracic paravertebral myotomes in complete spinal cord injury. AB - STUDY DESIGN: A study of thoracic paravertebral muscle motor-evoked potentials using transcranial magnetic stimulation in spinal cord injury patients and control participants. OBJECTIVES: To develop a method to study the level and density of corticospinal lesions in thoracic spinal cord injury. SUMMARY OF BACKGROUND DATA: Cervical and lumbar spinal cord injury, unlike thoracic spinal cord injury, can be quantified by recording muscle motor-evoked potentials from limb muscles. For thoracic spinal cord injury, the use of paravertebral muscles is limited by complex innervation patterns and the greater difficulty in obtaining muscle motor-evoked potentials. METHODS: In 10 patients with complete midthoracic spinal cord injury (T4-T7) and 10 age-matched control participants, muscle motor-evoked potentials were recorded from all thoracic paravertebral muscles using transcranial magnetic stimulation with a double-cone stimulating coil over the vertex. RESULTS: In control participants, muscle motor-evoked potential responses evoked in all myotomes had progressively increasing latency in a rostrocaudal direction. Threshold was comparable in all segments. The duration of muscle motor-evoked potentials was unrelated to the spinal level. In spinal cord injury, responses were elicited in all segments above a lesion and in a varying range of segments below the lesion. In comparison with control participants, threshold was lower above and higher below the lesion (P < 0.001) in patients with spinal cord injury. Latency was longer than normal both above and below the lesion (P < 0.001). Duration was not significantly different from that in control participants at any level. CONCLUSIONS: Paravertebral muscle motor-evoked potentials can be elicited below the level of a complete spinal cord injury. Possible reasons for this include the multisegmental innervation of these muscles and the long muscle fiber conduction. Stretch reflex activation elicited by contraction of muscles above the lesion is thought to be an unlikely mechanism because of the latency of the response. Although the presence or absence of muscle motor-evoked potentials does not appear to be a sensitive indicator of the level of thoracic spinal cord injury lesion, analysis of muscle motor-evoked potentials reveals abnormal patterns that may assist in defining lesions. Finally, lower threshold above the lesion suggests corticospinal hyperexcitability of this pathway as a result of central plasticity after spinal cord injury. PMID- 12131744 TI - Neurophysiologic monitoring of spinal nerve root function during instrumented posterior lumbar spine surgery. AB - STUDY DESIGN: Retrospective review of 61 consecutive patients. OBJECTIVES: To determine the effectiveness of combining intraoperative monitoring of both spontaneous electromyographic activity and compound muscle action potential response to stimulation for detecting a perforation of the pedicle cortex irritation of nerve root during lumbar spine fusion surgery. SUMMARY OF BACKGROUND DATA: The complication rate from instrumentation used with lumbar spine fusion varies from 1 to 33%. To prevent neurologic complications, several monitoring techniques have been used to alert surgeons to possible neurologic damage being introduced during nerve decompression or placement of instrumentation with spine procedures. Because of different sensitivities, one monitoring technique may not be as effective for preventing complications as a combination of techniques. METHODS: Sixty-one consecutive patients who underwent instrumented posterior lumbar fusions received continuous electromyographic monitoring and stimulus-evoked electromyographic monitoring. A significant neurophysiologic event was signaled by sustained neurotonic electromyographic activity, prompting an alert and a pause in the surgical manipulations that precipitated the activity. After insertion of the transpedicular screws, the integrity of the pedicle cortex was tested by stimulating each screw head and recording compound muscle action potentials. In the presence of a pedicle breach, stimulus intensities below 7 mA were sufficient to evoke compound muscle action potentials from the muscle group innervated by the adjacent spinal nerve root, prompting a surgical alert and subsequent repositioning of the screw. RESULTS: Fourteen significant neurophysiologic events occurred in 13 of 61 patients (21%). Sustained neurotonic electromyographic discharges occurred in 5 of 40 patients during placement of interbody fusion cages, in 2 patients during placement of transpedicular screws, and in 1 patient during tightening of rods. On pedicle screw stimulation, breaches of the pedicle cortex were detected in 6 patients. After surgery, no new neurologic deficits were found in 60 of the 61 patients. One patient who experienced temporary paraparesis had sustained neurotonic electromyographic discharges during retraction of the thecal sac and distraction of the disc space before placement of the cage. CONCLUSION: These results suggest that intraoperative electromyographic monitoring provides a real-time measure of impending spinal nerve root injury during instrumented posterior lumbar fusion, allowing for timely intervention and minimization of negative postoperative sequela. PMID- 12131745 TI - Chiari I malformation associated with syringomyelia and scoliosis: a twenty-year review of surgical and nonsurgical treatment in a pediatric population. AB - STUDY DESIGN: Retrospective review of patients with Chiari I malformation with or without associated scoliosis. OBJECTIVES: Determine the effect of decompression of Chiari I malformation with syringomyelia on stabilization or improvement of associated scoliosis. SUMMARY OF BACKGROUND DATA: Chiari malformations are often associated with spinal deformities, including scoliosis. Studies have suggested a causal relation between syringomyelia and scoliosis. METHODS: Patients with Chiari I malformation and syringomyelia with or without scoliosis treated over the last 20 years were reviewed. Patients with any other anomalies were excluded. Scoliotic curves were classified by magnitude and curve type. All patients were treated with surgical decompression of the Chiari malformation with or without drainage of the syringomyelia. RESULTS: Twenty-five patients were identified, ranging in age from 19 months to 16.5 years. Nineteen patients (76%) had associated scoliosis. The majority of the patients with scoliosis (13 of 19) sought treatment for spinal deformity, and only 6 had for pain or neurologic symptoms. Eleven of 19 patients with scoliosis (58%) underwent fusion. Eight of 19 (42%) patients have not undergone fusion: 3 have experienced progress, 1 remains in a stable condition, and 4 have experienced improvement of curvature since undergoing decompression. The mean age of patients who experienced progress after decompression was 14.5 years, compared to 6 years for patients who experienced improvement. CONCLUSION: Early decompression of Chiari I malformation with syringomyelia and scoliosis resulted in improvement or stabilization of the spinal deformity in 5 cases. Each of these patients underwent decompression before 8 years of age and before the curve was severe. However, this series represents a few patients demonstrating this trend, and further follow-up and investigation are warranted. PMID- 12131746 TI - Quality of life and back pain: outcome 16.7 years after Harrington instrumentation. AB - STUDY DESIGN: A study on the quality of life of 82 patients with idiopathic scoliosis treated with Harrington instrumentation. OBJECTIVE: To analyze long term health-related quality of life and low back pain an average of 16.7 years after surgery. SUMMARY AND BACKGROUND DATA: Quality of life evaluated by self assessment questionnaires is an accepted outcome measure of surgical procedures. The purpose of this study was to evaluate the health status with the German version of internationally accepted and psychometrically tested questionnaires. METHODS: Quality of life was measured with the Short Form-36 health profile. Low back pain was assessed using the Roland-Morris Questionnaire. Demographic data (age, sex, follow-up time), radiographic analysis (Kings classification, Cobb angle, extension and site of fusion), and rib cage deformity were correlated with these data. Radiologic parameters were analyzed longitudinally. RESULTS: In comparison with the age-matched healthy population, there was no significant difference in the physical Short Form-36 scale (P = 0.98). Surgically treated patients showed significantly lower scores than at baseline in the psychologic Short Form-36 scale (P = 0.005); vitality (P < 0.001), general mental health (P = 0.003), and role activities because of emotional problems (P < 0.001) were significantly different from those of the age-matched population. Sixty-five (79.3%) of the 82 patients reported no or occasional back pain in the Roland Morris Questionnaire. Five patients (6.1%) reported chronic back pain. Neither patient age at the time of surgery (P = 0.74) nor time of follow-up (P = 0.44), type of scoliosis (P = 0.56), or extent of fusion (P = 0.12) was associated with health-related quality of life or pain. The size of the preoperative (P = 0.06) and postoperative (P = 0.12) curves and preoperative (P = 0.28) and postoperative (P = 0.7) rib cage deformities did not correlate with the data of the Short Form 36 scale and the Roland-Morris Questionnaire. CONCLUSIONS: In comparison with the age-matched population, the long-term effect of surgery does not affect the physical quality of life. The psychologic health status is, however, significantly impaired. Neither the type of curve, the size of scoliosis, nor the rib cage deformity influences the data. PMID- 12131748 TI - The evaluation of the surgical management of nerve root compression in patients with low back pain: Part 1: the assessment of outcome. AB - STUDY DESIGN: This was a prospective study investigating the outcome of decompression surgery using validated measures of outcome. OBJECTIVES: To investigate the outcome of lumbar decompressive surgery in the initial postoperative year period in terms of function, disability, general health, and psychological well-being. SUMMARY OF BACKGROUND DATA: The majority of studies investigating the outcome of lumbar decompression surgery have been retrospective and have not used validated measures of outcome. This limits their interpretation and usefulness. METHODS: Eighty-four patients undergoing lumbar spinal stenosis surgery were recruited into this study. Patients were assessed by use of validated measures of outcome including the Oswestry Disability Index and the Short Form SF-36 General Health Questionnaire before surgery and 6 weeks, 6 months, and 1 year after surgery. RESULTS: A significant reduction in pain (P < 0.001) was observed at the 6-week postoperative stage; this did not change at the subsequent assessment stages. Only some of the SF-36 categories were sensitive to change. The subcategories that were sensitive to change were physical function (P < 0.05), bodily pain (P < 0.001), and social function (P < 0.05). Improvements were observed in these categories at the 6-week and 6-month reviews. A gradual reduction in the Oswestry Disability Index was observed with time, with changes principally being observed between the 6-week and 6-month review and the 6-week and 1-year review stages (P < 0.05). Minimal changes were observed in the psychological assessments with time. The outcome of surgery could not be predicted reliably from psychological, functional, or pain measures. CONCLUSIONS: The visual analogue pain scales, the Oswestry Disability Index, and certain categories of the SF-36 Questionnaire, namely bodily pain and physical and social function, appeared to be the most sensitive outcome measures, with significant improvements occurring at the 6-week and 6-month reviews. PMID- 12131749 TI - The evaluation of the surgical management of nerve root compression in patients with low back pain: Part 2: patient expectations and satisfaction. AB - STUDY DESIGN: This was a prospective study investigating patient expectations of and satisfaction with the outcome of decompression surgery. OBJECTIVES: To investigate patient expectations of surgery and short- and long-term satisfaction with the outcome of decompression surgery in terms of pain, function, disability, and general health. SUMMARY OF BACKGROUND DATA: Information is scarce regarding patient-rated expectations of surgery and measures of satisfaction with surgery in terms of specific outcome measures such as pain. METHODS: Eighty-four patients undergoing spinal stenosis surgery were recruited into this study. Before surgery, patients were also asked to rate their expectations in terms of improvement in pain, general health, function, and other such characteristics. In addition, at each postoperative review stage, patients were asked to rate their satisfaction with their improvement in these key outcome measures. RESULTS: The results demonstrated that patients had very high expectations of recovery, particularly in terms of pain and function, and that patients were confident of achieving this recovery (76.8%). Levels of satisfaction, however, varied considerably: 41% of patients were 50% satisfied with the outcome, and 30% were dissatisfied. Most patients felt that they had made the right decision to have surgery, although the surgery had achieved only 43.4% +/- 37.8 of the outcome they had expected. CONCLUSIONS: Examination of patients' expectations of and satisfaction with surgery revealed that patients frequently had unrealistic expectations of their surgery and as a consequence tended to have lower levels of satisfaction. PMID- 12131750 TI - Controversies in spine: Subspecialty certification should not be a requirement for spine surgery. PMID- 12131751 TI - Controversies in spine: Should there be subspecialty certification in spine surgery? PMID- 12131752 TI - Inverse laminoplasty for the treatment of lumbar spinal stenosis. AB - STUDY DESIGN: Fifteen patients with lumbar spinal stenosis were treated by a new technique, inverse laminoplasty, and the results were evaluated clinically and radiologically. OBJECTIVE: To present the advantages of inverse laminoplasty over laminectomy for the treatment of lumbar spinal stenosis. SUMMARY AND BACKGROUND DATA: Laminectomy has been used widely in the treatment of lumbar spinal stenosis. Destruction of the spinal bony structure, instability, and peridural scar formation are the main problems with this procedure. To overcome these disadvantages, a practical technique is presented here. MATERIAL AND METHODS: In a prospective study, 15 patients who underwent surgery with the inverse laminoplasty technique were evaluated clinically and radiologically. The Oswestry Disability Index was used for clinical assessment. L4-L5 spinal stenosis was detected in all patients. As the operative technique, the L4 lamina was elevated en bloc using a high-speed drill and rongeur. After removal of the ligamentum flavum, the roof of the foramina, and/or disc, the lamina was rotated 180 degrees, rested on facets, and reattached by use of a titanium miniplate. RESULTS: All patients improved clinically and neurologically after this procedure. The mean Oswestry Disability Score was 38.33 preoperatively and 7.0 postoperatively. The mean follow-up time was 17.3 months. Spinal canal diameters were calculated by preoperative and postoperative computed tomography, and the mean enlargement was 77.8%. No complications were observed. CONCLUSION: With this technique, the important integrity of the spinal osseous structures is preserved, and a significant enlargement of the spinal canal area is achieved. This technique prevents peridural scar formation after laminectomy caused by a mechanical barrier effect. Long-term follow-up is needed to evaluate spinal stability in these patients. PMID- 12131753 TI - Traumatic enucleation of the body of the sixth cervical vertebra without neurologic sequelae: a case report. AB - STUDY DESIGN: Case report of an adult man with traumatic enucleation of the body of the sixth cervical vertebra with initial minimal neurologic damage and without vascular injury, which was successfully treated with surgery. OBJECTIVES: To describe the traumatic mechanism, diagnostic procedures, operative findings, and adopted management. SUMMARY AND BACKGROUND DATA: Literature review suggests that the observed case is exceptional. Similar vertebral injuries are usually associated with severe neurologic and vascular damages. METHODS: A 38-year-old farmer with enucleation of the body of the sixth cervical vertebra was admitted to the Orthopedic Clinic of Padua University. The patient initially was treated with halo traction. Two days later, he underwent anterior open reduction and internal fixation with plate. He was discharged 2 months later with a halo vest. RESULTS: Two years after surgery, the pedicles' fracture had healed solidly with normal intervertebral alignment and a slight loss of the height of the C6-C7 intervertebral space (fibrous union). He is asymptomatic and has totally resumed his habitual work. CONCLUSION: Traumatic enucleation of the body of a lower cervical vertebra is a rare injury. The case presented here demonstrates that such a lesion can be less severe than a complete dislocation or a burst fracture. The good outcome of surgery suggests that the treatment adopted was effective, although an interbody fusion would have been more adherent to conventional surgical principles. PMID- 12131754 TI - Vertebral osteonecrosis associated with the use of intradiscal electrothermal therapy: a case report. AB - STUDY DESIGN: This report describes a case of vertebral body osteonecrosis associated with the use of intradiscal electrothermal therapy. OBJECTIVES: To alert clinicians to a previously unreported complication, vertebral body osteonecrosis, after an intradiscal electrothermal therapy procedure. SUMMARY OF BACKGROUND DATA: The intradiscal electrothermal therapy procedure is a new treatment that has been advocated in the management of chronic low back pain of discogenic origin. The intradiscal electrothermal therapy procedure uses a fluoroscopically guided thermal catheter to heat the intervertebral disc. A review of the literature regarding this procedure has not revealed osteonecrosis as a complication. METHODS: The patient's work-up and postoperative course are documented, and all medical records were reviewed retrospectively. RESULTS: The patient's pain had improved only minimally at initial follow-up after L5-S1 combined anterior and posterior spinal fusion, undertaken after intradiscal electrothermal therapy failure. CONCLUSIONS: Intradiscal electrothermal therapy has gained increasing popularity in the treatment of discogenic low back pain, in large part because of its minimally invasive nature and perceived low risk for major complications. Previous reports in the literature have not noted any major complications associated with the proper use of this device. Clinicians should be advised that intradiscal electrothermal therapy can be associated with complications, which in the current case, led to osteonecrosis of the vertebral body. PMID- 12131755 TI - Healing of autologous bone in a titanium mesh cage used in anterior column reconstruction after total spondylectomy. AB - STUDY DESIGN: Autologous bone inside a titanium mesh cage, used as an anterior strut in a reconstruction after total spondylectomy, was histologically examined in a postmortem specimen. OBJECTIVES: To determine whether the autologous bone inside the titanium mesh cage attained fusion and remodeling in a combined reconstruction, consisting of an anterior titanium mesh cage with posterior multilevel instrumentation, after total spondylectomy. SUMMARY OF BACKGROUND DATA: There are few previous reports on the histologic analysis of the bone inside a titanium mesh cage when it is used clinically as an anterior column support in a spinal fusion. Attaining biologic bony fusion is desirable for long term stability after total spondylectomy. METHODS: A postmortem specimen from a 16-year-old boy with Ewing's sarcoma at T6, who died of lung metastasis 16 months after total spondylectomy and combined reconstruction, was analyzed. RESULTS: Histologic examination revealed many viable cells and normal lamella of trabecular bone formation in the grafted bone inside the mesh. Consecutive trabecular cancellous bony fusion between the grafted bone and the adjacent vertebral bodies was achieved. CONCLUSION: Remodeling and fusion of the grafted bone inside the titanium mesh cage was observed. Combined reconstruction using an anterior titanium mesh cage with posterior multilevel instrumentation after total spondylectomy makes it possible to achieve biologic fusion of the bone inside the mesh cage with the adjacent vertebral bodies. PMID- 12131756 TI - The Tower of Babel? An assessment of the development and state of genetic and SNP technologies. PMID- 12131757 TI - Treasure hunting in a new era: genotyping of single nucleotide polymorphisms and the search for complex disease genes by association scans. AB - Genotyping single nucleotide polymorphisms by technologies like those described in this issue are propelling us towards a new era in the study of human genetics. This new era promises improved prospects for mapping genes responsible for complex human diseases and other complex traits. In this article, I sketch empirical results and statistical theory that together form the scientific rationale for anticipating enhanced success in gene mapping in the new era that is now approaching. PMID- 12131758 TI - Psychiatric genetics in silico: databases and tools for psychiatric geneticists. AB - Bioinformatics can significantly impact the laboratory genetics process from the study design phase to conclusive identification of a disease gene. The present review will highlight key databases to enhance psychiatric genetic study design, based on full use of genomics data and the golden path sequence. It will address methods to ensure comprehensive genetic data mining, using the best available genomic and genetic databases such as the University of California Santa Cruz human genome browser, Ensembl, Mapview, dbSNP and GDB, and locus-specific databases such as Online Mendelian Inheritance In Man. Using the golden path sequence as a template, with the necessary quality checks, it is possible to design detailed genetic studies from sequence information alone. Drawing together this diverse information, it is possible to characterize a locus or gene in silico to a very detailed level. This in turn can have real cost and efficiency benefits by assisting in the identification of markers that are most likely to be informative, or by highlighting the best candidate genes for study. PMID- 12131759 TI - A nanobiotechnology roadmap for high-throughput single nucleotide polymorphism analysis. AB - Genetic analysis based on single nucleotide polymorphisms (SNPs) has the potential to enable identification of genes associated with disease susceptibility, to facilitate improved understanding and diagnosis of those diseases, and should ultimately contribute to the provision of new therapies. To achieve this end, new technology platforms are required that can increase genotyping throughput, while simultaneously reducing costs by as much as two orders of magnitude. Development of a variety of genotyping platforms with the potential to resolve this dilemma is already well advanced through research in the field of nanobiotechnology. Novel approaches to DNA extraction and amplification have reduced the times required for these processes to seconds. Microfluidic devices enable polymorphism detection through very rapid fragment separation using capillary electrophoresis and high-performance liquid chromatography, together with mixing and transport of reagents and biomolecules in integrated systems. The potential for application of established microelectronic fabrication processes to genetic analyses systems has been demonstrated (e.g. photolithography-based in situ synthesis of oligonucleotides on microarrays). Innovative application of state-of-the-art photonics and integrated circuitry are leading to improved detection capabilities. The diversity of genotyping applications envisaged in the future, ranging from the very high-throughput requirements for drug discovery through to rapid and cheap near-patient genotype analysis, suggests that several SNP genotyping platforms will be necessary to optimally address the different niches. PMID- 12131760 TI - Novel and alternate SNP and genetic technologies. AB - There are many different genotyping technologies and chemistries. Other articles in this special issue focus on nanotechnology, bioinformatics, DNA chips and genotyping methods. This article focuses on four method categories not featured elsewhere in this, or the following special issue: (1) melting curve-based technologies such as dynamic allele-specific hybridization (DASH), melting curve single nucleotide polymorphism (McSNP), fluorescent resonance energy transfer (FRET), hybridization-based melting curves, and homogeneous assay formats based on melting curves; (2) non-PCR-dependent assays such as the oligonucleotide ligation assay and Invader, and isothermal amplification techniques such as rolling circle amplification; (3) rapid whole genome sequencing with methods such as the use of single molecule arrays and molecular resonance sequencing; (4) and other promising novel technologies. PMID- 12131761 TI - Discovery and utilization of haplotypes for pharmacogenetic studies of psychotropic drug response. AB - The treatment of seriously mentally ill patients is complicated by variability in individual response to psychotropic drugs. Some patients remain treatment refractory even after two to three therapeutic modalities. Other patients experience adverse events that range from mild discomfort, to poor compliance, to life threatening. Genaissance Pharmaceuticals is actively engaged in a candidate gene-based haplotype (HAP Marker) approach to the pharmacogenetics of drug response and adverse events. In the present article, we review reasons why HAP Markers are more useful than single nucleotide polymorphisms (SNPs) for discovering genetic correlations to clinical response. In addition, we review our approach to HAP Marker discovery, which involves discovering SNPs in the functional regions of genes by sequencing, organizing these SNPs into HAP Markers for an index population of ethnically diverse individuals and calculating population frequencies for these HAP Markers. For clinical correlations, HAP Markers are defined and correlated to clinical data using the in-house DecoGen Informatics System. This approach has clear implications for the discovery of psychiatric disease-associated genes as well as for the development of safer, more efficacious psychiatric drugs. PMID- 12131762 TI - Single nucleotide polymorphisms and their characterization with oligonucleotide microarrays. AB - Small variations in human DNA sequences influence our predisposition to disease and our response to medications. Many psychiatric diseases appear to be polygenic. Because our molecular understanding of the genetic etiology of neuropsychiatric disorders is very limited, the discovery and characterization of these variations is crucial in identifying disease genes. Furthermore, this knowledge may enable the customized selection of drug treatments based on an individual's genotype so as to maximize efficacy yet avoid adverse side effects. Examples of findings using conventional methods for determining genotypes, as well as new and future technologies for the characterization of single nucleotide polymorphisms, will be discussed, with a focus on psychiatric diseases and medications. PMID- 12131763 TI - High-throughput genetic screening using matrix-assisted laser desorption/ionization mass spectrometry. AB - Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) has become a powerful and widespread analytical tool in all fields of life science. Compared with other techniques, its high accuracy and sensitivity makes it a superior method, especially for the analysis of nucleic acids. Recent problems in the analysis of nucleic acids by MALDI-TOF MS can be solved using an automated MALDI-compatible sample-preparation system. Together with the reliable minisequencing assay, high-throughput genotyping of single nucleotide polymorphisms by MALDI-TOF MS is able to become a routine method in research, clinical genetics and diagnostics. PMID- 12131765 TI - Relation between morphologic changes in the main pancreatic duct and exocrine pancreatic function after a secretin test. AB - INTRODUCTION: Because pancreatic exocrine function testing methods are problematic, both imaging and functional tests are important in the diagnosis of chronic pancreatitis. AIM: To evaluate the usefulness of ultrasonographic monitoring of the main pancreatic duct after a secretin test. METHODOLOGY: A total of 70 subjects (30 control subjects, 26 patients with probable chronic pancreatitis, and 14 patients with definite chronic pancreatitis) were selected. The main pancreatic duct diameters were measured serially after an injection of secretin (100 IU/body). The relation between the magnitude of the duct dilation and exocrine pancreatic function on the secretin test was evaluated. RESULTS: The main pancreatic duct dilated immediately after a bolus injection of secretin, showed a peak after 2-5 minutes, and recovered gradually. The response curve of the definite group had a flatter pattern than that of the other groups. For the maximal to basal duct diameter ratio, statistically significant differences were found between the control and definite groups and between the control and probable groups. In addition, the ratio correlated significantly with the maximal bicarbonate concentration and secretory volume on the secretin test. CONCLUSIONS: The results of the current study indicate that exocrine pancreatic function and the morphologic changes of the main pancreatic duct are significantly related. Dynamic ultrasonographic findings may reflect pancreatic function; consequently, this test may be a useful tool in the diagnosis of chronic pancreatitis. PMID- 12131766 TI - Clinical application of (11)C-methionine positron emission tomography for evaluation of pancreatic function. AB - INTRODUCTION: In recent years, it has become increasingly necessary to evaluate pancreatic function after pancreatectomy, but few precise methods are available. AIMS: To evaluate different surgical techniques for pancreatectomy in terms of the preservation of pancreatic function by (11)C-methionine positron emission tomography (PET), which determines amino acid metabolism in the pancreas. METHODOLOGY: The study included 33 pancreatectomy cases: 5 of distal pancreatectomy, 5 of pancreaticoduodenectomy, 10 of pylorus-preserving pancreaticoduodenectomy, 7 of duodenum-preserving pancreatic head resection, and 6 of inferior pancreatic head resection. The method was as follows. Approximately 370 MBq (11)C-methionine was intravenously injected. Cross-sectional imaging of the pancreas was performed by PET after 30 minutes. The images obtained were used to determine the radioactivity concentration in the pancreas. By adjustment of the radioactivity concentration for body weight and dosage, the differential absorption ratio could be determined to indicate the level of accumulation in the pancreas. Each surgical method used was evaluated on the basis of the differential absorption ratio. Postoperative total pancreatic accumulation was divided by preoperative level to calculate the total preserved pancreatic function rate (TPPFR), and postoperative local pancreatic accumulation was divided by preoperative level to calculate the local preserved pancreatic function rate (LPPFR). These rates were then compared for the individual techniques used. RESULTS: The results indicated that TPPFR and LPPFR were 61.2 +/ 20.0% and 114.6 +/- 29.4% for distal pancreatectomy (n = 5), 31.8 +/- 20.0% and 58.7 +/- 30.0% for pancreaticoduodenectomy (n = 5), 21.6 +/- 14.7% and 58.4 +/- 29.8% for pylorus-preserving pancreaticoduodenectomy (n = 10), 47.9 +/- 35.5% and 67.7 +/- 30.6% for duodenum-preserving pancreatic head resection (n = 7), and 48.1 +/- 29.5% and 83.9 +/- 30.5% for inferior pancreatic head resection (n = 6). TPPFR was highest in distal pancreatectomy cases. Among the pancreatic head resections, TPPFR was quite high for both inferior pancreatic head resection and duodenum-preserving pancreatic head resection. In contrast, TPPFR for pancreaticoduodenectomy and pylorus-preserving pancreaticoduodenectomy was quite low. LPPFR was highest for distal pancreatectomy and only slightly lower for inferior pancreatic head resection. In contrast, LPPFR was markedly lower for pancreaticoduodenectomy and pylorus-preserving pancreaticoduodenectomy. CONCLUSION: In conclusion, this method using (11)C-methionine PET is clearly useful for the evaluation of pancreatic function after pancreatectomy. PMID- 12131767 TI - Cyclooxygenase-2 is overexpressed in chronic pancreatitis. AB - INTRODUCTION: Cyclooxygenase enzymes catalyze a critical step in the conversion of arachidonic acid to prostaglandins, which are important mediators of acute and chronic inflammation. The constitutively expressed cyclooxygenase-1 (COX-1) appears to regulate many normal physiologic functions in several cell types, whereas the inducible cyclooxygenase-2 (COX-2) enzyme mediates the inflammatory response. AIMS AND METHODOLOGY: We investigated the expression of COX-2 in tissues of 35 patients with chronic pancreatitis, 6 patients with pancreatic cancer, and 5 control patients by immunohistochemical analysis and correlations to clinicopathologic features. RESULTS: We found an overexpression of COX-2 in the atrophic acinar cells (80% of patients), hyperplastic ductal cells (86% of patients), and islets cells (97% of patients) but not in normal pancreatic tissues. The COX-2 overexpression in the tissue of patients with chronic pancreatitis was significantly correlated with the frequency of acute attacks of pancreatitis. Tissue from patients who had more than five acute attacks of pancreatitis (n = 10) exhibited COX-2 immunoreactivity of a significantly higher score in atrophic acinar cells (p = 0.004). No correlation could be found with other examined clinical features such as duration of the disease, diabetes, alcohol consumption, smoking, or pain. CONCLUSION: Our results support the hypothesis that COX-2 may be involved in inflammatory responses in chronic pancreatitis and in the progression of this chronic inflammatory disease. PMID- 12131768 TI - Characterization of the neurotrophic response to acute pancreatitis. AB - INTRODUCTION: Interesting preliminary data on changes in the neurotrophin system in various digestive diseases have recently begun to emerge. AIMS: To measure changes in messenger RNA (mRNA) levels of neurotrophins and to identify cell types expressing neurotrophins in the pancreas of rats with L-arginine-induced pancreatitis. METHODOLOGY: Rats were killed at time points from 2 hours to 4 weeks after the induction of pancreatitis, and responses were measured by assay. RESULTS: By RNase protection assay, ciliary neurotrophic factor (CNTF) mRNA expression showed a rapid response (sixfold increase over control) in the inflamed pancreas at 2 hours. The levels of mRNA expression of brain-derived neurotrophic factor (BDNF), glial cell line-derived neurotrophic factor (GDNF), neurotrophin-3 (NT-3), and neurotrophin-4 (NT-4) in the inflamed pancreas reached a peak at 1 week (2.5-fold, twofold, fourfold, and fivefold increase, respectively). By immunohistochemistry, immunoreactivity for all neurotrophins examined was observed in the islets of Langerhans in the control pancreas at all time points, but it was markedly reduced in the islets in the inflamed pancreas at 2 and 6 hours. Acinar and ductal cells, inflammatory cells, and neural elements were immunoreactive for those neurotrophins in the inflamed pancreas from 2 hours to 2 weeks. CONCLUSION: The temporal and spatial expression of neurotrophins in the course of experimental pancreatitis suggests that their upregulation is a critical component of the response of the pancreas to injury in this model. PMID- 12131769 TI - Human aspartyl (asparaginyl) beta-hydroxylase monoclonal antibodies: potential biomarkers for pancreatic carcinoma. AB - INTRODUCTION: Pancreatic adenocarcinoma is among the top 10 leading causes of death due to cancer in the United States. The lack of reliable and sensitive biomarkers for this disease makes it difficult to render an early diagnosis. AIMS: To evaluate carcinoma-associated monoclonal antibodies (MoAbs), including AF-20, SF-25, and FB-50, for their binding specificity to pancreatic adenocarcinoma relative to normal pancreatic tissue. In addition, binding of the Th9 MoAb to human Reg 1 protein was studied because of its potential role in cell growth. METHODOLOGY: Adjacent histologic sections were immunostained with each of the MoAbs and graded on a scale of 0 to 4+, corresponding to the relative distribution and intensity of immunoreactivity within the tumor and normal adjacent tissue. RESULTS: Intense levels (grade 3 or 4) of FB50 immunoreactivity were detected in 19 of 19 tumors but not in normal adjacent pancreatic tissue. In addition, increased levels of FB50 immunoreactivity were detected in at least 75% of the tumor cells in 18 of the 19 cases. SF-25 immunoreactivity similarly distinguished pancreatic adenocarcinoma from normal pancreas in 14 of 19 cases. In contrast, AF20 immunoreactivity was detected in 6 of 19 pancreatic adenocarcinomas, and for the most part, the labeling was focal and of a low level. TH9 immunoreactivity was detected in 5 of 19 tumors but also in normal as well as inflamed adjacent pancreatic tissue. CONCLUSION: These results suggest that the FB50 and SF25 MoAbs represent excellent potential biomarkers of pancreatic adenocarcinoma and could be configured in an immunoassay for detecting pancreatic adenocarcinoma cells in biologic fluids. PMID- 12131770 TI - Suppression of human pancreatic carcinoma cell growth and invasion by epigallocatechin-3-gallate. AB - INTRODUCTION: The consumption of green tea is associated with a lower risk of several types of human carcinomas. A number of studies have focused on the possible mechanisms of cancer prevention by tea extracts, especially polyphenols such as epigallocatechin-3-gallate (EGCG). AIMS AND METHODOLOGY: Green tea derived EGCG was tested in human pancreatic carcinoma cells. The cells (PANC-1, MIA PaCa-2, and BxPC-3) were treated with different doses of EGCG (0, 25, 50, 100, and 200 micromol/L) for 48 hours in culture medium. Proliferation of pancreatic carcinoma cells was measured by means of the WST-1 colorimetric assay. For the study of cell invasion, the cells were incubated with 100 micromol/L EGCG for 2 hours. Then, the cells were added into the cell insert, coated with Matrigel basement membrane matrix. After incubation at 37 degrees C for 24 hours, the cells that had invaded through the Matrigel were counted visually under the microscope. RESULTS: The growth of all three pancreatic carcinoma cells was significantly suppressed by EGCG treatment in a dose-dependent manner. EGCG treatment caused significant suppression of the invasive ability of pancreatic carcinoma cells PANC-1, MIA PaCa-2, and BxPC-3 but did not affect the cell cycle protein cyclin D1. CONCLUSION: EGCG may be a potent biologic inhibitor of human pancreatic carcinomas, reducing their proliferative and invasive activities. PMID- 12131771 TI - The expression of retinoic acid receptors and the effects in vitro by retinoids in human pancreatic cancer cell lines. AB - INTRODUCTION: Analogues of vitamin A have been shown to influence growth of malignant tissue, such as pancreatic cancer. AIMS: To study the expression of retinoic acid receptors (RAR) in pancreatic cancer cells and the effect of three different retinoids on the cell number in vitro were studied. METHODOLOGY: Cell lines were established from 13 patients who underwent surgery for pancreatic adenocarcinoma. The expression of the retinoic acid receptors (RAR) and retinoic X receptor (RXR) subtypes (alpha, beta, and gamma) was studied with western blotting and specific antibodies. The effect of incubation with all-trans retinoic acid (atRA; tretinoin), 9-cis-retinoic acid (9-cis-RA), and 13-cis retinoic acid (13-cis-RA; isotretinoin) on the cell number was examined with use of a Roche XTT cell proliferation kit. RESULTS: The RXR alpha receptor was expressed in all cell lines. RAR alpha,beta and RXR beta were expressed in most of them. RXR gamma was expressed in about half of the cell lines and RAR gamma in only one. Incubation of the cells with retinoids showed a decreased cell number at concentrations of 10(4) M, except for 9-cis-RA, to which only about half of the cell lines responded. CONCLUSION: Two or more of the RAR subtypes were expressed in each pancreatic cell line. There was no uniform pattern of receptor expression; however, the cell lines responded with decreased cell number to high concentrations of atRA and 13-cis-RA but not to 9-cis-RA. PMID- 12131772 TI - Dibutyltin dichloride modifies amylase release from isolated rat pancreatic acini. AB - INTRODUCTION: Dibutyltin dichloride (DBTC) is widely used as a stabilizer for polyvinylchloride plastics and is of particular toxicologic interest. AIM: To examine the effects of DBTC on pancreatic exocrine function in isolated rat pancreatic acini. METHODOLOGY: Isolated rat pancreatic acini were incubated with various secretagogues in the presence or absence of DBTC. We investigated the effects of DBTC on amylase release, receptor binding, and protein kinase C (PKC) enzyme activity. RESULTS: DBTC reduced cholecystokinin octapeptide (CCK-8) stimulated and carbamylcholine-stimulated amylase release and the binding of [(125)I]CCK-8 to isolated rat pancreatic acini. Conversely, DBTC potentiated secretin-stimulated amylase release, although it slightly inhibited [(125)I]secretin binding to its receptors. In addition, DBTC potentiated amylase release stimulated by vasoactive intestinal peptide, 8-bromoadenosine 3', 5' monophosphate (8Br-cAMP) or calcium ionophore A23187, whereas it had no influence on amylase release stimulated by 12-O-tetradecanoylphorbol 13-acetate. The protein kinase C (PKC) inhibitor calphostin C abolished the DBTC-induced potentiation of amylase release stimulated by 8Br-cAMP or A23187. Moreover, DBTC caused a significant translocation of PKC enzyme activity from cytosol to membrane fraction. CONCLUSIONS: These results indicate that DBTC reduces CCK-8- and carbamylcholine-stimulated amylase release by inhibiting their receptor bindings to pancreatic acini, whereas it potentiates cAMP-mediated amylase release by activating PKC in isolated rat pancreatic acini. PMID- 12131773 TI - Visualization of the heterogeneous internal structure of so-called "pancreatic necrosis" by magnetic resonance imaging in acute necrotizing pancreatitis. AB - INTRODUCTION: Contrast-enhanced computed tomography (CT) is the gold standard for assessing the severity of acute pancreatitis, especially for evaluating the presence of pancreatic necrosis (poorly perfused area). However, the contrast medium used for CT is potentially toxic to the pancreas and kidney. Therefore, medical institutions without facilities for hemodialysis hesitate to acquire contrast-enhanced CT images. Diagnostic values of magnetic resonance imaging (MRI) in pancreatic diseases have been shown. AIM: To evaluate the usefulness of MRI in the assessment of the severity of acute pancreatitis. RESULTS: All necrotic regions in the pancreas were visualized by gadolinium-enhanced MRI. Furthermore, MRI can discriminate the poorly perfused pancreatic area, namely so called "pancreatic necrosis" judged on CT, into three parts: 1) necrotic area of the pancreatic parenchyma, 2) perinecrotic fluid collection, and 3) hemorrhagic foci. Inflammatory changes that were required for severity grading were also evaluated sufficiently by MRI. CONCLUSION: These results suggest that MRI is useful for the assessment of severity of acute pancreatitis. PMID- 12131774 TI - Primary culture of porcine pancreatic acinar cells. AB - INTRODUCTION: The methodology of acinar cell culture has become of primary importance in the research of pancreatic physiology and pharmacology. AIM: To develop a method for primary culture of porcine pancreatic acinar cells. METHODOLOGY: Dispersed pancreatic acinar cells were made by RPMI-1640 medium containing collagenase III. After purification, the isolated acinar cells were cultured in RPMI-1640 medium with 2.5% fetal bovine serum. The morphologic characteristics of acinar cells were described. (3)H-thymidine incorporation of acinar cells and activity of amylase or lipase were determined during the culture. RESULTS: There were no remarkable morphologic changes in the pancreatic acinar cells during the 20-day culture. The acini showed the tendency of gathering but did not attach to the walls of the culture disks. Incorporation of (3)H-thymidine in acinar cells in the primary culture was well kept. The secretion of amylase or lipase from acini decreased with the time of culture. CONCLUSIONS: In the primary culture of acinar cells from porcine pancreas developed in this study, the acinar cells retained normal morphology and ability of growth but not secretion of amylase or lipase. The method would be beneficial for further experiments on acini of porcine pancreas. PMID- 12131775 TI - Nonadhesive organ culture of human exocrine pancreatic cells with their stroma. AB - INTRODUCTION: Studies using explanted tissue have shown that it is possible to keep adult human cells in organ culture with a preserved morphology for up to 1 month as spheres in a nonadhesive organ culture. AIMS: The current study was to determine whether human exocrine pancreatic cells also can be grown in this manner. METHODOLOGY: Small tissue samples from organ donors and tumor-free resection rim from patients with pancreatic carcinoma were obtained (n = 16 adults). From each patient, fragments of approximately 300 microm in diameter were cultured and investigated with light microscopy and scanning and transmission electron microscopy at the time of explantation and after 5, 10, 20, 30, and 40 days of culture. RESULTS: Incubation of cultured fragments with vital dyes revealed a viable epithelium. At the time of explantation all the tissue fragments had a rough appearance with an uneven, torn periphery. During the first week of culture the fragments became rounder, with a smooth surface covering the whole circumference. This spheroid morphology persisted for the rest of the 6 week culture period. The fragments were within 1 week covered by a highly differentiated, polarized epithelium with secretory apparatus, apical secretion granules, and microvilli, as well as specialized cell junctions, with the same appearance as acinoductal pancreatic cells of the original tissue. The core of the fragments consisted of connective tissue with vascular elements, fibroblasts, leukocytes, and a few ductal and acinar elements. Transmission electron microscopy of the spheroids revealed a continuous basal lamina underneath the epithelium. Immunostaining for cytokeratin 5, 6, 7, 8, 17, and 18 was strongly positive in the epithelium. CONCLUSION: These results show that normal exocrine pancreatic cells can be grown in vitro in a nonadhesive organ culture with their stroma. PMID- 12131776 TI - Activated rat pancreatic stellate cells express intercellular adhesion molecule-1 (ICAM-1) in vitro. AB - INTRODUCTION: Activated pancreatic stellate cells (PSCs) have recently been implicated in the pathogenesis of pancreatic fibrosis. AIMS: To examine the role of PSCs in pancreatic inflammation by determining whether activated PSCs express intercellular adhesion molecule-1 (ICAM-1). METHODOLOGY: Culture-activated rat PSCs were treated with interleukin-1 beta (IL-1 beta), tumor necrosis factor alpha (TNF-alpha), ethanol, or acetaldehyde. ICAM-1 expression was analyzed by flow cytometry. The induction of mRNA was assessed by Northern blot analysis. The binding activity of transcription factors was examined by electrophoretic mobility shift assay. The activation of mitogen-activated protein kinases was assessed by Western blotting with use of antiphosphospecific antibodies. The adhesion of MOLT-4 cells to activated PSCs was also assessed. RESULTS: Culture activated PSCs expressed ICAM-1. The expression was increased in response to IL-1 beta and TNF-alpha but not to alcohol, with comparable mRNA induction. IL-1 beta and TNF-alpha increased the nuclear factor-kappa B (NF-kappa B)-specific and activator protein-1-specific DNA binding activity, whereas the NF-IL6 activity was not altered. Alcohol did not increase NF-kappa B binding activity. IL-1 beta and TNF-alpha activated extracellular signal-regulated kinase 1/2, c-Jun N terminal kinase/stress-activated protein kinase, and p38 mitogen-activated protein kinase. Inducible ICAM-1 expression was blocked by pyrrolidine dithiocarbamate, a specific inhibitor of NF-kappa B activation, but not by inhibitors of mitogen-activated protein kinase pathways. IL-1 beta and TNF-alpha increased the ICAM-1-mediated binding of MOLT-4 cells to activated PSCs, indicating a role of ICAM-1 in the adhesion of leukocytes to PSCs. CONCLUSION: Our results suggest that activated PSCs express ICAM-1 mainly through the activation of NF-kappa B, thus playing a role in the pathogenesis of pancreatic inflammation. PMID- 12131777 TI - Effect of stimulators such as GLP-1, PACAP, and nicotinamide on glucose stimulated insulin secretion from porcine pancreatic endocrine cells in long-term culture. AB - INTRODUCTION: Transplantation of glucose-responsive insulin-secreting cells has the potential to result in a cure for diabetes. AIM: To report the development of a model of adult porcine pancreatic endocrine cells (PE cells) exhibiting glucose stimulated insulin secretion during a long-term culture period, in vitro. METHODOLOGY: The PE cells were prepared by non-enzymatic digestion and purified by modifying a technique developed in our laboratory. The cells were first cultured for 7 days in RPMI-1640 medium containing 10% fetal bovine serum and 10 mM nicotinamide. On adhesion to the culture flasks, cells were collected by trypsinization, and then cultured in tissue culture dishes in medium with or without stimulators such as glucagon-like peptide-1 (GLP-1), pituitary adenylate cyclase-activating polypeptide (PACAP), and nicotinamide. The ability of the cells to respond to glucose-stimulated insulin secretion was also observed with and without stimulators. The immunocytochemical studies demonstrated pancreatic islets with well-preserved insulin and glucagon-containing cells. The morphologic integrity of cultured porcine cells was observed for up to 5-6 weeks after the purification. RESULTS: At a concentration of 3.3 mM glucose, PACAP and nicotinamide did not affect glucose-dependent insulin secretion, whereas 10 nM GLP-1 stimulated insulin secretion significantly. However, when glucose concentration was increased to 20 mM, 10 nM GLP-1 had no effect on insulin secretion. We also demonstrated that GLP-1 and PACAP could maintain insulin secretion better than control in the culture up to 5 weeks. Also GLP-1 and PACAP increased the number of insulin-secreting cells in culture. CONCLUSION: These results demonstrate that GLP-1 and PACAP increased the number of pancreatic beta cells in culture. PMID- 12131778 TI - Xenotransplantation of pig islets in diabetic dogs with use of a microcapsule composed of agarose and polystyrene sulfonic acid mixed gel. AB - INTRODUCTION: The authors have designed a microcapsule composed of agarose and polystyrene sulfonic acid (PSSa) mixed gel that provides a protective barrier against complement attack. Xenografts of islets, encapsulated in an agarose-PSSa microcapsule, have been shown to normalize blood glucose in rodents with chemically induced diabetes for extended periods of time without immunosuppression. AIM: To investigate the efficacy of agarose-PSSa microencapsulated pig islets in reversing diabetes in a large animal model. METHODOLOGY: Diabetes was induced in beagle recipients by total pancreatectomy. Each recipient received three to five intraperitoneal injections of either encapsulated (n = 5) or nonencapsulated pig islets (n = 2). RESULTS: In all dogs receiving microencapsulated islets, the graft function was achieved for 7.4 +/- 3.1 weeks (mean +/- standard error), as determined by elimination or reduction of exogenous insulin requirement. In three recipients, the fasting blood glucose levels were maintained at < or = 200 mg/dL without any exogenous insulin for a period of 6, 50, and 119 days. Circulating porcine C-peptide was detected in the sera of all dogs after transplantation of encapsulated islets. Immunohistologic examination revealed the presence of insulin-positive cells in the microcapsules. In contrast, in two dogs receiving nonencapsulated islets there was no graft function. CONCLUSIONS: This preliminary study demonstrates that agarose-PSSa microencapsulated pig islets can survive and function for weeks or months in totally pancreatectomized diabetic dogs without immunosuppression. PMID- 12131779 TI - Pancreatic cholesterol esterase, ES-10, and fatty acid ethyl ester synthase III gene expression are increased in the pancreas and liver but not in the brain or heart with long-term ethanol feeding in rats. AB - INTRODUCTION: Chronic alcohol consumption predisposes susceptible individuals to both acute and chronic pancreatitis. AIMS: Our hypothesis was that alcohol increases the risk of pancreatitis by disrupting defense mechanisms and/or enhancing injury-associated pathways through altered gene expression. Hence, we studied the expression of pancreatic genes in rats chronically exposed to ethanol. METHODOLOGY: Male Wistar rats were pair-fed liquid diets without and with ethanol for 4 weeks. Total RNA was extracted from rat pancreas and other organs. The mRNA expression patterns among pancreatic samples from ethanol-fed rats and controls were compared with use of mRNA differential display. The differentially expressed cDNA tags were isolated, cloned, and sequenced. RESULTS: One cDNA tag that was overexpressed in the pancreas showed 99% sequence homology to a rat pancreatic cholesterol esterase mRNA (CEL; Enzyme Commission number [EC] 3.1.1.13). The differential expression was confirmed by realtime PCR. Gene expression was also increased in the liver but not in the heart or brain of the alcohol-fed rats. Because CEL has fatty acid ethyl ester (FAEE)-generating activity and FAEEs play a major role in acute alcoholic pancreatitis, we determined the expression of other genes encoding for FAEE-generating enzymes and showed similar organ-specific expression patterns. CONCLUSION: Our results demonstrate that chronic ethanol consumption induced expression of FAEE-related genes in the pancreas and liver. This upregulation may be a central mechanism leading to acinar cell injury. PMID- 12131780 TI - Thrombotic microangiopathy in acute pancreatitis. PMID- 12131781 TI - Clinical evaluation of oral administration of a cholecystokinin-A receptor antagonist (loxiglumide) to patients with acute, painful attacks of chronic pancreatitis: a multicenter dose-response study in Japan. AB - INTRODUCTION: Cholecystokinin (CCK)-receptor antagonists have been found to markedly reduce the severity of pancreatitis and improve survival in experimental animal models of acute pancreatitis. CCK appears to play an important role in the development and progression of acute pancreatitis, and the recent development of CCK antagonists has provided a new approach to the treatment of acute pancreatitis in humans. AIMS: The therapeutic efficacy of a CCK-A receptor antagonist, loxiglumide, in patients with painful acute attacks of chronic pancreatitis was evaluated. METHODOLOGY: A multicenter dose-response controlled trial was conducted at 110 institutions in Japan from June 1993 to December 1994. Chronic pancreatitis was diagnosed for all patients on the basis of the Japanese criteria for chronic pancreatitis. Two-hundred seven patients were randomized to oral treatment with loxiglumide (300, 600, and 1,200 mg/d) or placebo for 4 weeks. The efficacy of treatment was evaluated on the basis of clinical symptoms, physical signs, and serum pancreatic enzyme levels. The groups were comparable with respect to age, sex, etiology, complications, and previous treatment. RESULTS: The improvement rate of the abdominal and/or back pain was 46% in the loxiglumide 300-mg group, 59% in the 600-mg group, and 52% in the 1,200-mg group, and it was 36% in the placebo group (600 mg versus placebo: p < 0.05). The physical signs evaluated--abdominal tenderness and resistance--improved in all three loxiglumide groups, and the serum pancreatic amylase and trypsin levels decreased significantly in the 600-mg group (p < 0.05). The overall clinical improvement rate was 46% in the 300-mg loxiglumide group, 58% in the 600-mg group, and 52% in the 1,200-mg group, and it was 34% in the placebo group. CONCLUSION: These results indicate that oral administration of loxiglumide may be useful in the treatment of patients with acute, painful attacks of chronic pancreatitis, and 600 mg/d is recommended as a beneficial dosage. PMID- 12131782 TI - Fecal elastase 1 measurement compared with endoscopic retrograde cholangiopancreatography for the diagnosis of chronic pancreatitis. AB - INTRODUCTION: Indirect tests of exocrine pancreatic function are thought to be of little sensitivity and specificity in mild to moderate insufficiency as compared with direct function tests. Direct tests, which are claimed to be the "gold standard" of functional diagnosis, are too complicated to be performed on great numbers of patients and are not standardized. AIMS: To characterize the use of an indirect function test (fecal elastase 1 measurements determined independently from a direct test), in this study we compared it with the gold standard of morphologic diagnosis, endoscopic retrograde cholangiopancreatography (ERCP). METHODOLOGY: Data for 213 patients who underwent ERCP (104 males and 109 females; mean age, 54 years [8-89]) were collected prospectively, including fecal elastase 1 measurements and clinical and ERCP data. RESULTS: Elastase 1 findings correlated with pancreatic duct changes (p < 0.05). At a cutoff point of 200 microg/g, the positive predictive value of elastase 1 measurement for moderate/severe duct changes was 90.4%, and for any duct changes it was 96.8%. The sensitivity was only 45.3% for any duct changes but 76.5% for severe changes. Specificity for moderate/severe changes was 86%. CONCLUSION: Fecal elastase 1 measurements appear to be valuable for characterizing patients at high risk for chronic pancreatitis, even if their sensitivity is lower than that of direct tests. PMID- 12131784 TI - Hunger After the Millennium: Perspectives and Demands. AB - Problems of over- and under-nutrition are changing with continued world population growth, rapid urbanization, and improved economic access to food. Still about 800 million lack reliable access to good quality and safe food. This article suggests action necessary to develop systems of sustainable food production in developing countries to help improve national development and food security, and give people better access to food. PMID- 12131783 TI - Palliative MEFLEP therapy in advanced pancreatic cancer: excellent response in a patient with Her-2/neu amplification. AB - INTRODUCTION: Patients with pancreatic cancer often present initially in advanced disease with many compromising factors, and yet they may still be responsive to chemotherapy. AIMS: The response of 23 patients with advanced pancreatic cancer to continuous infusion therapy was investigated. METHODOLOGY: From September 1995 to February 1998, 23 patients with advanced pancreatic cancer, many with compromising factors, were treated with a MEFLEP regimen: biweekly 24-hour infusions of etoposide, 5-fluorouracil, leucovorin, epirubicin, and cisplatin, all given through an infusion pump, plus megestrol acetate, 160 mg/d, taken daily. A total of 145 courses were given. Overall response rate was 21% (4/19) for assessable chemo-naive patients; median survival for all 23 patients was 6 months; 22% of patients were alive at 1 year; and a clinical response benefit was attained in 35%. RESULTS: Toxicity was manageable; grade 3 or 4 leukopenia occurred in 1 patient each, 1 patient had fever and grade 3 infection, and grade 3 and 4 hyperammonemic encephalopathy developed in 3 and 1 patients, respectively. All four of the latter patients recovered uneventfully within 2 days of initiation of therapy. Nine patients were evaluated by fluorescence in situ hybridization for the Her-2/neu oncogene, but for only one patient did amplification of the gene occur. She attained complete remission with treatment and lived for 26.7 months after diagnosis. CONCLUSION: Biweekly MEFLEP is an active and manageable regimen for patients with advanced pancreatic cancer with compromised clinical status. PMID- 12131785 TI - Chocolate: A Heart-healthy Food? Show Me the Science! AB - Cocoa and chocolate foods produced by appropriate methods can contribute significant amounts of heart-healthy flavanols to the diet. These flavanols may enhance cardiovascular health by delaying blood clotting, improving vascular endothelial function, and helping to moderate inflammation. The benefits of chocolate can be enjoyed without guilt as part of a healthful balanced diet. PMID- 12131786 TI - Food in American History Part 5: Pork: A Nation Divided: The American Civil War Era (1861-1865). AB - This fifth installment of the Food in American History series considers the period from 1861 through 1865, the era of the American Civil War, with pork as the central food theme. PMID- 12131787 TI - Dietary Supplement Use in Children: Who, What, Why, and Where Do We Go From Here? Executive Summary. AB - This executive summary is an overview of the goals, presentations, and recommendations from a conference on dietary supplement use in children conducted by the National Institute of Child Health and Human Development in partnership with the National Institutes of Health Office of Dietary Supplements, other members of the federal research community, and the private sector. PMID- 12131789 TI - Access to Dietary Supplements for Valid Medical Uses in Children. AB - Dietary supplements are marketed primarily to prevent disease. In certain rare genetic metabolic disorders, nutritional supplements can be therapeutic, but the absence of government regulation of the supplement industry makes these products unreliable. PMID- 12131788 TI - What Is a Nutrient? "If It Walks Like a Duck. " AB - The identification of an increasing number of substances in foods that have health-related biologic activity has opened a serious debate concerning the definition of a nutrient. Although there may be advantages to differentiating these food-borne substances as nutrients and non-nutrients, it is important to recognize that these substances have both positive and adverse effects. Moreover, the same substances may have multiple effects other than classic deficiency syndromes. In any case, it is important to differentiate the definitional needs for purposes of marketing from those required by science and health. PMID- 12131790 TI - Herbal Supplements: Indications, Clinical Concerns, and Safety. AB - Most herbal home remedies used in children, including teas made from chamomile, fennel, ginger, or mints, are benign. Most topical preparations are benign; however, garlic poultices can cause burns. Internal use of herbs containing saturated pyrrolizidine alkaloids should be avoided. PMID- 12131791 TI - Current Patterns of Supplement Use in Adolescents. AB - Eighth-grade students use vitamin and mineral supplements to a much greater extent than previously. Implications for dietary assessment and counseling are provided. PMID- 12131792 TI - National Nutrition Data: Contributions and Challenges. AB - National nutrition surveys in the United States indicate that about 1 in 2 infants or preschoolers and 1 in 3 school-aged children use at least 1 vitamin and mineral supplement, typically a multivitamin. Among users, adolescents are most likely to use 2 or more supplements, often a vitamin/mineral or multiple vitamin supplement plus single vitamins. PMID- 12131793 TI - Dietary Supplements in American Children: Scientific vs Marketing Justifications. AB - The American public receives conflicting messages from dietitians, nutritionists, physicians, and manufacturers regarding food supplements. Consumers commonly distrust scientists and justify supplement use based upon word of mouth and friendship patterns. Scientific-based education regarding supplement use is vital in the present atmosphere where consumer misinformation is rampant. PMID- 12131794 TI - Nutrition Monitoring Application in the Food Industry. AB - Nutrition scientists in the food industry use nutrition monitoring data in a variety of ways that include developing nutrition communications for consumers and health professionals, guiding product development and reformulation, and applying research applications. Continuous nutrition monitoring is essential to influence positively the nutrient content of the food supply and meet the changing nutrition needs of the population. This article reviews food industry application of nutrient intake information and provides specific examples of use. PMID- 12131795 TI - We're Fat and Getting Fatter! What Is the Food Industry's Role? AB - Is the food business to blame for the fattening of America? Can nutrition professionals play the blame game and get themselves off the hook? Some perspectives on the strategy and tactics of the war against obesity. PMID- 12131796 TI - Psychiatrists keep NP prescribing debate alive. PMID- 12131797 TI - Treating hypertension in patients with diabetes. AB - The coexistence of hypertension and diabetes synergistically increases the risk of cardiovascular and renal diseases. Although aggressive blood pressure reduction can decrease the rate of cardiovascular events in general, blockade of the renin-aldosterone-angiotensin system is a particularly effective target for antihypertensive treatment that may provide additional benefits for patients with diabetes. This article details evidence for blood pressure targets and antihypertensive drug choice in hypertensive patients with diabetes. PMID- 12131798 TI - Managing vulvar vestibulitis. AB - Vulvar vestibulitis, a type of vulvodynia, affects many American women. Patients typically present with a history of intermittent or continuous, localized, vulvar pain and frequently can't tolerate sexual intercourse. Here, review the etiology, history and physical examination, and comprehensive treatment of vulvar vestibulitis, including nonpharmacologic, pharmacologic, psychosocial, and surgical measures. PMID- 12131799 TI - Staying on the ethical high ground with complementary and alternative medicine. AB - The lack of substantive research regarding complementary and alternative medicine (CAM) and its rising use stimulate interest in the ethical obligations that primary care providers face when advising patients about these therapies. Here, we explore how ethical principles relate to decision-making regarding CAM. We also provide a framework for making decisions that involve CAM interventions. PMID- 12131802 TI - Who's supporting your CE program? PMID- 12131803 TI - Tracking the cause of recurrent thrombosis. PMID- 12131804 TI - Document everything. PMID- 12131805 TI - Evaluation of outcomes in nursing students using clinical concept map care plans. PMID- 12131806 TI - A transcultural nursing educational environment: an imperative for multicultural students. PMID- 12131807 TI - "Puzzle Patients" and critical thinking. PMID- 12131808 TI - Planning for an accreditation visit. PMID- 12131809 TI - Competent and safe practice: a profile of disciplined registered nurses. PMID- 12131810 TI - Transformational leadership in nursing education. PMID- 12131811 TI - Experiential learning: a strategy to teach conflict management. PMID- 12131813 TI - Embedding a pedagogical model in the design of an online course. AB - The importance of course design and its relationship to learning is overlooked when the electronic version of a course is static. The author discusses the process used to redesign a traditional course into a dynamic and interactive Web based course by using a pedagogical model developed specifically for hypermedia. Congruence between the instructional methods and the media is a must for a successful design that meets the needs of a wide variety of learners. The hyperlearning model, Dimensions, provides the infrastructure to facilitate the congruence of methods and media while supporting multiple learning styles. PMID- 12131812 TI - Nursing student creativity: teaching healthcare concepts to elementary students. PMID- 12131814 TI - The senior practicum. AB - Senior practicum offers students valuable time to apply knowledge from the entire nursing program within a supportive setting while having the benefit of direct supervision from an experienced clinician. Benefits to agencies include a well prepared practitioner who is thoroughly oriented to agency policies and procedures. The authors describe the practicum experience of a baccalaureate nursing program and discuss the benefits from multiple points of view. PMID- 12131815 TI - A clinical experience with foster families. AB - A structured, preceptored community clinical experience was established in foster care to help nursing students better understand the health problems of foster children and the nurse's role in interdisciplinary teamwork. The public health nurse preceptor identified specific opportunities for student involvement including intake, health record review, home visiting, and case management. The student was able to blend physical assessment skills, knowledge of child development and parenting, and psychosocial evaluations to have a rewarding service learning experience. PMID- 12131816 TI - Discovering co-learning with students in distance education sites. AB - Distance education is promoted as an effective way to reach more students, but how effective are interactive learning strategies in distance nursing courses? The authors discuss how nursing students adjusted to distance technologies and became actively involved in creating an environment that enabled development of supportive co-learning relationships. Learner perspectives on teaching strategies, distance learning, and becoming nurses are described. PMID- 12131817 TI - Cultural competence immersion experiences: public health among the Navajo. AB - Holistic nursing practice requires cultural competence. The author describes a cultural immersion experience involving public health among the Navajo as having four components: inspiration, preparation, immersion, and reentry. Such experiences require faculty with requisite attitudes and abilities, as well as a supportive academic environment to nurture this student learning opportunity. Important faculty development issues discussed include faculty cultural competence and interpersonal skills, fiscal support, curricular coordination, and administrative commitment. PMID- 12131819 TI - Education and high quality community care. PMID- 12131818 TI - Service learning: preparing a healthcare workforce for the next century. AB - Service learning coupled with community-based education is an effective way for educators to prepare nurses for their roles in healthcare for the 21st century. Structuring the curriculum to provide learning experiences that prepare nurses to meet the healthcare needs of aggregates and populations is the responsibility of nursing faculty today. Students need opportunities to work both with and within communities as partners, as well as experiences with interdisciplinary collaboration, to achieve this goal. The authors describe how two disciplines within the university and a community in rural eastern North Carolina partnered to accomplish this task. PMID- 12131820 TI - Families' experiences of the last office of deceased family members in the hospice setting. AB - This study aimed to describe the experience of family members who had been involved in the last office of a deceased relative and explored what the participation meant to each individual. Undertaken in Hong Kong, data were collected through semi-structured interviews of six bereaved family members who had participated in the procedure. Five major themes emerged with regard to the meanings of the participation: a sense of completeness; a continuation of relationship; fulfillment of responsibility; conveyed messages of gratitude and respect; and an enhancement of the acknowledgement of the reality of death. The most significant part of each individual's experience is the humanistic approach of care throughout the procedure including the attitude of the nurses. PMID- 12131821 TI - Massage and aromatherapy massage: nursing art and science. AB - This article begins by reviewing the place of complementary therapies in palliative care from the perspective of UK professional organizations, namely the Nursing and Midwifery Council (NMC) (formerly the United Kingdom Central Council for Nursing, Midwifery and Health Visiting (UKCC)) and the British Medical Association (BMA). It then reviews recent research on the use of massage and aromatherapy massage in palliative care and comments on their credibility and implications, thereby addressing nursing science. The art of nursing is explored through four case histories where massage was used intuitively when words were difficult or seemed inadequate. To conclude, by focusing on the therapies provided by a UK hospice, there is consideration of the practicalities of implementing massage and aromatherapy massage in a specialist palliative care unit. PMID- 12131823 TI - Defining and determining quality in end-of-life care. AB - Health care providers are directing increased attention toward improving end-of life care for patients who are dying and their families (Support Principle Investigators, 1995; Ferrell et al, 2000). Integral to the goal is determining what constitutes quality care for the dying. One of the major problems, however, in making such a determination results from the lack of consensus regarding a conceptual definition of quality. The purpose of this article is to stimulate debate and discussion on the meaning of quality in the context of providing end of-life care. To aid in this discussion, the various perspectives of what constitutes quality in health-care practice will be discussed with special attention being paid to the delivery of quality end-of-life care. Conceptual decisions drive approaches to research method and measurement (Kristjanson, 1992), therefore an examination of the issues surrounding the measurement of quality health care in the context of care for the dying will also be explored. PMID- 12131822 TI - The risk assessment of bereavement in a palliative care setting. AB - An effective bereavement risk assessment document used in a palliative care setting was identified which could ensure bereavement support for those in need, be it a family or carer group. The nursing team were used as assessors with a system to indicate carers' and relatives' immediate need following the death of a patient. The aim of this bereavement risk assessment system was to lessen the possible long-term effects of unresolved grief for family and carers of patients who died within a hospice setting. The assessment document is a useful education tool for nurses, allied support professionals and bereavement support volunteers in their work with grief and loss. PMID- 12131824 TI - Multiprofessional palliative care in a general hospital: education and training needs. AB - Palliative care for those who need it should be an integral part of National Health Service (NHS) practice in all settings in the UK. This study used a focus group and individual structured questionnaires, completed during a semi structured interview, to gain an insight into how different professionals in the hospital view palliative care. An assessment was made of their understanding of it, previous training and current knowledge. The respondents also discussed training needs and personal experiences. The findings show that previous training was variable but, in general, relatively young and inexperienced staff were working in the hospital under enormous pressures of workload, staff shortages and without experienced role models from whom to learn the complexities of palliative care. Communication difficulties focused around the lack of support and the inherent problems of multidisciplinary team working resulting in frustration and less than optimum care being given. Rapid staff turnover compounded the problems. Although staff were keen to learn, clinical workload always took priority over training. The following recommendations arose from the study: undergraduate training should be standardized, and postgraduate training and support should be ongoing and flexible to accommodate the diversities of knowledge and difficulties encountered in attending set sessions. PMID- 12131825 TI - The role of oral transmucosal fetanyl citrate in the management of breakthrough cancer pain. AB - Breakthrough pain affects at least two-thirds of cancer patients at some point during their illness (Portenoy and Hagen, 1990; di Palma et al, 2000). The ideal preparation for managing breakthrough pain is one that is able to mirror its characteristics, i.e. rapid in onset and relatively short in duration. Oral transmucosal fentanyl citrate (OTFC, Actiq[R]) is an opioid formulation with a unique delivery system that fulfils these criteria. It has a role to play in the treatment of breakthrough pain for some patients who are receiving opioid treatment for cancer pain. OTFC was preferred over their routine medication by most patients who recently participated in a UK clinical trial (Hanks et al, 2002, unpublished data). This article aims to give an overview of this new and novel preparation developed for the control of breakthrough cancer pain. PMID- 12131826 TI - Adverse incidents must be reported and learnt from. PMID- 12131827 TI - New evidence reaffirms the safety of the MMR vaccine. PMID- 12131828 TI - Nurse who made a video of herself mocking patients. PMID- 12131829 TI - Cardiorespiratory physical assessment for the acutely ill: 2. AB - The second of this two-part article aims to develop advanced cardiovascular and respiratory knowledge to enhance practice for the nurse caring for an acutely ill patient on a general ward. The first part dealt with the core aspects of care in respiratory and cardiovascular physical assessment, including respiratory function and failure, pulse oximetry, oxygen therapy, fluid therapy, pulse measurement, blood pressure and electrocardiogram monitoring (Vol 11(11): 750-8). As the use of critical care beds within trusts becomes more difficult to manage, with ever-increasing patient dependency and occupancy figures remaining high, critically ill patients are likely to remain in wards longer or be discharged from a critical care environment earlier. These patients will require more frequent, direct, non-invasive and invasive monitoring to ensure that they do not deteriorate, and that any deteriorations are detected early and managed effectively. The skills and information discussed here will help nurses advance their practice and improve the quality of their care. PMID- 12131830 TI - National Patient Safety Agency and the reporting of errors. PMID- 12131831 TI - Evaluating Snoezelen for relaxation within chronic pain management. AB - This experimental study investigated the use of Snoezelen - a sensory environment purported to produce relaxation - against traditional relaxation within the pain clinic setting. The variables measured included pain, anxiety, depression, coping, self-efficacy and disability. Assessments were carried out at three time intervals on a range of symptoms designed to reflect the multidimensional nature of the chronic pain experience, including pain intensity and quality, anxiety, depression, coping, confidence and quality of life. The experimental group experienced significant reductions in pain (sensory score P=0.002), and an improvement in self-efficacy (P=0.02) and sickness impact for the following scales: physical (P=0.009), psychosocial (P=0.009), recreation (P=0.001), sleep (P=0.001) and sickness impact total (P=0.001). The control group experienced significant improvements in sickness impact scales of psychosocial (P=0.05), sleep (P=0.01) and sickness impact total (P=0.004). The findings suggest that Snoezelen environments are as effective as, if not slightly better than, teaching relaxation within the traditional pain clinic environment for this group of patients. PMID- 12131832 TI - Foetal programming and coronary heart disease in later life. AB - The maternal environment has traditionally been seen as having the utmost importance for the developing foetus. However, a debate has been raging within medicine for the last decade, which points to the key role of the mother's state of health for the subsequent health of the baby. In particular, her nutritional state has been argued to be of critical importance to the health of her offspring not only in infancy but also as it approaches middle age. Evidence has been accumulating that the baby's state of intrauterine nutrition will determine the likely health problems that the mature individual will exhibit. This proposition has enormous implications for anybody involved in health promotion and for public policy. This article aims to summarize the main points in this debate and to draw tentative conclusions for the practising nurse, midwife, and health visitor. PMID- 12131833 TI - Views of family carers and older people of information technology. AB - This article is the second in a series of four describing recent developments in Sweden aimed at promoting partnerships between older people, their families and formal service providers. The last article (Vol 11(11): 759-63) described the development of an information and communication technology (ICT) project ACTION - Assisting Carers using Telematics Interventions to meet Older persons' Needs -- and focused on the use of CT to help family carers to be more prepared for their caregiving role. This article focuses on the concept of usability within the ACTION project and the importance of working closely with participants in order to create an information and communication service that is both acceptable and of direct benefit to family members in their everyday caring situations. Nielsen's (1993) Model of Usability is described within the overall context of the project and is used as a framework for the cycle of development and testing that underpin ACTION. A variety of research methods are highlighted, with a central theme being that of user involvement, with particular reference to the USERfit approach (Poulson et al, 1996). The education and training of older people and their family carers to use the ACTION technology is outlined and examples are given of the empowering effects of the use of the service and its user-driven focus. Recommendations for the further technical development of ACTION are firmly based on the comments and suggestions provided by older people and the family carers themselves. PMID- 12131834 TI - The concept of hope in nursing 1: its origins, background and nature. AB - Hope has been described in theoretical terms for many years but the recognition of the importance of hope within the practice of nursing is a more recent phenomenon. Despite the recent growth of references to hope within contemporary nursing literature, it is reasonable to suggest that there remain gaps in the substantive knowledge base and that there appears to be room for both additional research and further discursive literature. Accordingly, this series of six articles will explore the nature of hope, review the existing theoretical and empirical work in several discrete areas of nursing, and provide case studies to illustrate the role that hope plays in clinical situations. This article focuses on the origins, background and definitions of hope. The next article will examine hope within mental health nursing, and further articles will focus on hope within palliative care nursing, hope in gerontological nursing and hope in critical and acute care nursing. PMID- 12131835 TI - RCN and NMC have both reached crucial stages. PMID- 12131836 TI - The number of complaints made about nurses is rising. PMID- 12131837 TI - A new Mental Health Act: where is the evidence for it? PMID- 12131838 TI - Nurse who swore and shouted at patients and staff. PMID- 12131839 TI - Treatment of an infected venous leg ulcer with honey dressings. AB - An infectious diseases unit is potentially an ideal environment in which to carry out research into honey-based dressings. This article looks at the barriers to carrying out case study-based research, and describes the treatment of an elderly gentleman with venous leg ulcers. The patient's wounds improved with the honey based dressing, but it failed to free the wound of microbes. One possible explanation is that the honey, instead of killing the microbes, actually provided them with a food source. PMID- 12131840 TI - Case study: benefits of IT for older people and their carers. AB - The first article in this series (Vol 11(11): 759-63) described the development of the ACTION project, an information technology-based initiative designed to enhance the quality of life for family carers and older people, and the second article (Vol 11(12): 827-31) considered how the usability of the ACTION system as a whole was evaluated. This, the third article, presents a case study which describes in more detail the impact of the ACTION project on the lives of both a family carer (Rolf) and his wife (Kerstin), and on the technical worker (Thomas) who helped Rolf and Kerstin to learn to use the ACTION system. The impact of the intervention is assessed using two broad sets of criteria: (1) those relating to the PREP (Preparedness, Enrichment, Predictability) model of nursing interventions, and (2) those relating to the wider implications of participation in a research and development project of this nature. PMID- 12131841 TI - Research-based care on an acute inpatient psychiatric unit. AB - Many studies of research-based practice in nursing highlight factors that impede the development of practice. With the aim of adding to this body of knowledge, a modified grounded theory approach was used in order to understand more about these barriers and how individual nurses utilize research in their practice. A selective sample of five staff nurses from one acute inpatient psychiatric unit took part in semi-structured interviews. Three main themes were identified, each with two sub-themes. These were (a) activities to utilize research with (i) a 'systematic' model and (ii) a 'latent' model of research utilization (b) enhancing research utilization with (i) organizational culture and (ii) individual attitude and knowledge and (c) impeding research utilization with (i) resources (ii) resistance to change. It is suggested that for these nurses research utilization occurs through their individual knowledge, skill and motivation coupled with organizational commitment. Recommendation is made that further investigation of the 'systematic' and 'latent' models should be carried out. Additionally, it is suggested that these research findings might be used to inform future training, further research-based initiatives and to raise managerial awareness of the impeding factors of research utilization. PMID- 12131842 TI - The concept of hope in nursing 2: hope and mental health nursing. AB - This article is the second in a series of six that explores the nature of hope, reviews the existing theoretical and empirical work in several discrete areas of nursing, and provides case studies to illustrate the role that hope plays in clinical situations. In this article we focus on hope within the formal area of psychiatric/mental health nursing. The article points out that there is a limited empirical literature covering several aspects and issues of hope, hopelessness and hope inspiration within the domain of psychiatry. However, despite these studies many questions still remain unanswered. The bulk of this empirical literature focuses on hopelessness, and in the main hopelessness associated with suicide/depression. As a result of these investigations, a range of interventions has been identified for inspiring hope in different client groups with mental health problems, and a summary of these is given. Importantly, however, the basic social process of hope inspiration for each of these client groups is fundamentally the same, in that the process remains subtle, unobtrusive and associated with the therapy/relationship. We conclude by indicating key areas/questions for future research, and raise key questions regarding future policy/education issues. PMID- 12131843 TI - Reporting of injuries, diseases and dangerous occurrences. AB - Case Scenario: Clematis, a staff nurse in an orthopaedic ward, injured her back when lifting a patient with another nurse from the bed to a wheelchair. She was admitted to hospital and was off sick for many weeks. When she returned to work she noticed that the accident from which she had completed was still on the ward and copies did not appear to have been circulated. What is the law and what is her responsibility? PMID- 12131844 TI - Survey of continuing professional education within nursing homes. AB - This article reports a survey of qualified nurses employed in nursing homes in a large city in the north of England. The aim of the study was to describe nurses' experiences and perceptions of continuing professional education (CPE), and their views on the development of their knowledge and skills through formal and informal education. Findings revealed that nurses in nursing homes perceived the value of educational programmes but had limited opportunities to attend formal education programmes. They faced a number of barriers and challenges in accessing formal education. Various informal ways of learning were used, including reading professional journals, watching videotapes and television programmes and accessing the Internet. However, respondents appeared not to use fully opportunities for sharing knowledge with colleagues. Although this study is based on a small sample, the findings support those of other studies in this field. Collectively, these studies suggest an urgent need to develop a range of approaches to CPE within care homes, both formal and informal, if the standards outlined within recent policy initiatives are to be achieved. PMID- 12131846 TI - How we use language and its value in nursing. PMID- 12131845 TI - Wound healing/product selection for a critically ill obese patient. AB - The selection of wound management products by nurses can be an area of confusion. Healthcare professionals involved with tissue viability issues have to make choices from a wide range of commercially available products to suit wound conditions. This article recounts the challenges facing a specialist nurse and his selection of a range of Smith & Nephew Healthcare products, when he was asked for expert advice to promote granulation in a massive wound following debridement for extensive abscess formation in the lower abdominal, perineum and ischial areas. PMID- 12131847 TI - Time to make headway on Agenda for Change. PMID- 12131848 TI - Hydrocolloids in wound management: pros and cons. AB - Hydrocolloids are interactive dressings which are well established in wound management. Because of their sophisticated composition, they can be used to manage a variety of wound types, from clean and granulating to sloughy and necrotic. While they may be more expensive per item than many other modern dressings, if used correctly they can be cost-effective, as they have a longer wear time. Hydrocolloid formulations appear to be changing and improving with ongoing research by manufacturers, but a significant limitation appears to be sensitivity reactions. Dressings that would benefit from improvement are those containing gelatin, and those containing colophony within the adhesive matrix. These known sensitizers are largely unpublicised, but can produce serious side effects in some individuals. PMID- 12131849 TI - Diet and coronary heart disease: advice on a cardioprotective diet. AB - Coronary heart disease is a major disease leading to early morbidity and mortality in the UK. The causes are multifactorial, a combination of modifiable and non-modifiable risk factors. Although recent statistics show a reduction in the coronary heart disease mortality rate, the disease continues to be a major focus in health care, with strategies being developed to improve prevention and treatment. Dietary modification has a role to play in both primary and secondary prevention. PMID- 12131850 TI - Fall prevention among older adults: is London ready for the NSF? AB - Fall prevention is a key standard of the National Service Framework for Older People (Department of Health, 2001). This article describes a study exploring the extent and nature of fall prevention initiatives among London primary care NHS services. It outlines fall prevention approaches used, considers the shortfalls of current practice and recommends ways to meet the demands of the national service framework. PMID- 12131851 TI - Promoting nursing participation within primary care groups. AB - This article describes the evaluation of a scheme to improve communication between primary care nurses and newly established primary care groups. Findings suggest that the successes of the scheme were in part attributable to the appointment of experienced practitioners to a facilitator/link nurse role and the sustained support of the scheme by the NHS community trust. The article concludes by suggesting that, in a time of constant change within primary care, innovations such as these contribute to the development of practitioner participation in the organization and delivery of primary care services. PMID- 12131852 TI - Subcutaneous infusion: non-metal cannulae vs metal butterfly needles. AB - This review aimed to evaluate the effectiveness of non-metal cannulae compared to metal butterfly needles in maintaining subcutaneous infusion sites in patients receiving palliative care. The Cochrane Library, Medline, Pre-Medline, Embase, CINAHL, Amed and Cancerlit were searched for relevant studies. Controlled trials comparing non-metal cannulae with metal butterfly needles for giving subcutaneous infusion to palliative care patients were included. The outcome considered was site duration in terms of hours of patency or until change was required. Four trials met the inclusion criteria although overall quality was poor due to low follow-up. Studies examined either Teflon or Vialon-coated catheters. All studies showed non-metal cannulae to be superior to metal. In individual studies estimates in mean increase in duration of the site range from 21 to 159 hours. It seems that non-metal cannulae are more effective in maintaining the duration of subcutaneous infusion sites than butterfly needles. Both types of non-metal catheter showed clear benefits. This review has not examined other outcomes but in general adverse effects lead to the removal of the catheter and so would be reflected in the outcome of considered. Although historically non-metal cannulae have been considerably more expensive there is now little difference between metal and Teflon-coated catheters. This review recommends the use of non-metal cannulae in preference to butterfly needles. PMID- 12131853 TI - Verification of expected death by district nurses. AB - Pressures on a local out-of-hours GP deputizing service led to lengthy delays before a doctor could visit to verify the (expected) death of patients under the care of the district nursing service. This resulted in distress to the relatives and delayed transfer of the body to a funeral home. This article explores an initiative to enable community nursing staff to verify those expected deaths and to provide more effective support to relatives at a difficult time. The article highlights the benefits of multidisciplinary collaboration in the development of an appropriate protocol and provides an overview of its implementation in one area. The initiative contributes to the Government's agenda to improve access to primary care and to create more patient-friendly services. PMID- 12131854 TI - The scope of guidelines to prevent healthcare-associated infections. AB - As care is increasingly delivered in the community rather than acute settings, there has been concern that this might be accompanied by a rise in healthcare associated infection. Consequently, the National Institute for Clinical Excellence (NICE) has commissioned a set of infection prevention guidelines for healthcare workers in community and primary care. The guideline developers were anxious to concentrate this guidance on the areas of most concern to practitioners, particularly in relation to devices. This article describes how a survey and focus groups were employed to identify the areas for guideline development, namely standard principles, long-term indwelling urinary catheters, enteral feeds and central intravascular devices. PMID- 12131855 TI - Nurses can be good substitutes. PMID- 12131856 TI - Tissue viability nurses need a forum for debate. PMID- 12131857 TI - Are wound care assessments real or imaginary and what can be done? PMID- 12131858 TI - Injecting drug use: developing a drop-in wound care clinic. AB - The first part of this two-part series examined the difficulties facing nurses working with injecting drug users with skin problems (Finnie and Nicolson, 2002). This article describes the needs of homeless people with skin complaints and the innovative development of a specific drop-in wound care clinic within The Big Issue Scotland premises in Glasgow. It illustrates some practical and political difficulties of working with a unique and challenging client group, and of developing a wound care service outwith the NHS. Case studies illustrate individual people and their own challenges. PMID- 12131859 TI - The effect of containment on the properties of sterile maggots. AB - A laboratory-based study undertaken to examine the effect of confinement in net bags upon the feeding mechanisms and growth rate of maggots of Lucilia sericata showed that free-range maggots survived better and grew significantly faster than maggots in bags (P<0.005). In a separate study it was also demonstrated that maggots in bags could survive on wound fluid that passed through the net without their having access to any form of solid food. This finding was consistent with clinical experience that suggests that although there may be some aesthetic advantages to the use of maggots in bags, their ability to combat infection or remove necrotic tissue from wounds is greatly reduced. PMID- 12131860 TI - Clinical practice benchmarking: implications for tissue viability. AB - Healthcare provision has been described as a 'lottery', reflecting a deficiency in equity of care (Ellis, 2000). Recent Department of Health (DoH, 1998, 1999) attempt to overcome this disparity and assure quality care by quality assessment and continuous quality improvement (Ellis and Morris, 1997). These documents advocate clinical benchmarking as a means to supporting this practice. This article provides an overview of the benchmarking process, with particular focus being applied to the pressure ulcer element. It reflects on the coalescence that appears to exist between implementing clinical benchmarking and the characteristics of specialist practice. It also analyses the effects of establishing the process on an existing tissue viability service. PMID- 12131861 TI - Best practice statement of the prevention of pressure ulcers. AB - The Nursing and Midwifery Development Unit, Scotland, has developed a range of best practice statements, one of which relates to the prevention of pressure ulcers. The development of the statement involved achieving a consensus as to what constituted best practice in this field among the 30 nurse specialists in tissue viability working within Scotland. The statement aims to provide practical, relevant and factual information to guide pressure ulcer prevention practice. PMID- 12131862 TI - Pressure ulcer classification: the systems and the pitfalls. AB - The classification of pressure ulcers constitutes an essential part of patient assessment and forms a baseline on which to monitor patients' progress and measure the outcomes of care. Four classification systems - Torrance, National Pressure Ulcer Advisory Panel, European Pressure Ulcer Advisory Panel, and Stirling - are discussed. Many classification systems have been adopted by trusts despite insufficient research in their use in clinical practice. Regardless of which classification system is implemented, nurses need to be familiar with the one they are using and how to stage accurately a pressure ulcer. This can only be attained through regular education. PMID- 12131863 TI - Review of an independent audit into the clinical efficacy of VACUTEX. AB - An independent report was completed and analysed by Pharmaceutical Research Associates International in August 2001 (Pro-Tex, 2001). This company provides a service to carry out independent project work and research studies within the healthcare sector. The project involved 73 tissue viability nurses and 93 patients, whereby the performance of VACUTEX capillary action wound dressing was assessed on both acute and chronic wounds. The audit takes into account the varying cultures across England and Wales and demonstrates this rapid method of comparing previous dressings used on a multitude of wound types, with analysis focusing on the versatility and efficacy of the VACUTEX capillary action wound dressing. This article reviews the findings of the audit. PMID- 12131864 TI - Kawasaki disease: the mystery continues. AB - Kawasaki disease (KD) is an acute vasculitis of infancy and early childhood that continues to baffle researchers and clinicians alike. Although the acute illness resolves spontaneously, permanent damage to the coronary arteries occurs in 20 25% of untreated children. The cause of KD remains unknown and there is no specific laboratory test to identify affected children. Nonetheless, high dose intravenous g-globulin plus aspirin administered within the first 10 days of fever significantly reduces the risk of coronary artery damage by unknown mechanisms. KD thus presents a unique dilemma: the disease may be difficult to recognize, there is no diagnostic laboratory test, there is an extremely effective therapy, and there is a 25% chance of serious cardiovascular damage or death if the therapy is not administered. This review will highlight some of the many unanswered questions about KD. PMID- 12131865 TI - The pneumococcal conjugate vaccine. AB - Streptococcus pneumoniae is the most frequent cause of otitis media, sinusitis, and pneumonia in children. It is also one of the most common causes of invasive bacterial infections in children including bacteremia and meningitis. One of the current issues regarding S. pneumoniae is the emergence of pneumococcal strains resistant to penicillin and other antibiotics. Children less than two years of age suffer an increased incidence of invasive pneumococcal disease but fail to respond to the 23-valent polysaccharide vaccine because of the immaturity of the T-cell independent immune function. Covalently conjugating the polysaccharide antigen to a carrier protein improves the immune response by permitting the host to utilize a T-cell dependent immune response that is adequately mature in children less than two years of age. Immunogenicity studies of the currently licensed heptavalent conjugated polysaccharide vaccine, (Prevnar, marketed by Wyeth Lederle Vaccines) demonstrated that infants vaccinated with three doses 2 months apart at 2, 4, and 6 months of age successfully developed antibodies to all 7 serotypes; booster doses at 12-15 months demonstrated an amnestic response for each serotype. Immunogenicity studies have similarly demonstrated successful responses in children with sickle cell disease and human immunodeficiency virus infection. An efficacy trial involving nearly 38,000 subjects demonstrated the vaccine's effectiveness in healthy children against invasive pneumococcal disease as well as against pneumonia and otitis media. Currently the US Advisory Committee on Immunization Practices (ACIP) recommends that all infants and children under 24 months of age receive the vaccine. The ACIP recommends that infants receive the vaccine routinely at 2, 4 and 6 months with a fourth dose at 12 to 15 months of age. Infants may receive the first dose as early as 6 weeks of age. The vaccine is also indicated for children 24 to 59 months of age who are at high risk for pneumococcal infection. Adverse events include local reactions in the first two days following vaccination such as approximately 10% reporting erythema, 10% induration, and 20% tenderness. Fever of 38 degrees C or higher occurred in 15% to 25% of children in the first two days following vaccination. Follow-up studies should address important questions regarding the use of pneumococcal conjugate vaccine and other age groups. PMID- 12131866 TI - Long-term safety and efficacy of low-fat diets in children and adolescents. AB - The safety and efficacy of low-fat diets in children and adolescents were evaluated through a systematic review of the current literature. Eight major studies were reviewed. The safety of the diets was judged by measures of growth and development and by meeting nutritional requirements. The efficacy of the diets was evaluated by their effect on the plasma levels of total cholesterol and low-density lipoprotein (LDL) cholesterol. All studies except 1 showed that children and adolescents exhibited normal growth and development while on a low fat diet. In 3 of the studies, nutritional requirements for calcium, zinc, phosphorous and vitamin E were below the recommended daily intake. In each of the 5 studies in which efficacy was determined, a significant decrease in the levels of total or LDL cholesterol was observed. Low-fat diets are generally safe and efficacious when performed under medical supervision. PMID- 12131867 TI - Clinical evaluation and changes of the respiratory epithelium function after administration of Pidotimod in Greek children with recurrent respiratory tract infections. AB - BACKGROUND: Several studies have been conducted on young children with recurrent respiratory infections using several compounds (synthetic derivates or lyophilized bacterial extracts) causing improvement in the clinical process. METHODS: We conducted a prospective, randomized study comparing the clinical results and the changes of the respiratory epithelium function after the administration of immunostimulating drug (Pidotimod) to children with respiratory infections over a 9 month period. A total of 32 children (group A) were randomly assigned to receive Pidotimod therapy while a second group of 18 children (group B) weren't. All the children in group A received Pidotimod (400 mg x 2 daily) for fifteen days and 400 mg daily for the next twenty days. The proper function of the ciliary respiratory epithelium in all children was checked, using the Edicol Orange and CaH PO4 2H2O, coloring method before the therapeutic intervention and after the first and the sixth month. RESULTS: 87.5% of group A, responded exceptionally well to treatment presenting two or less infections in the nine month period, whereas only 33.3% of group B showed improvement (p<0.001). In group A, the clearance of the respiratory epithelium, from a primary 37 minutes decreased to 32 minutes in the first month and 19'5" six months after the therapy. In group B, the corresponding time was decreased from a primary 36'4" to 34'2" and 31' respectively (p=0.01). CONCLUSIONS: Our results suggest that Pidotimod therapy is a reliable, simple and safe approach to treat children with recurrent respiratory infections and it can reduce the frequency of such infections as a result of improvement of the ciliary respiratory epithelium. PMID- 12131868 TI - Ventriculoperitoneal shunting complicated with cerebrospinal fluid pseudocyst and acute appendicitis. AB - Ventriculoperitoneal shunting (VPS) is the standard treatment of hydrocephalus in children but can be followed by various intraabdominal complications. Formation of a cerebrospinal fluid (CSF) pseudocyst is a rare VPS complication. A case of a non-infected CSF pseudocyst complicated with acute appendicitis is presented. PMID- 12131869 TI - Neonatal hyperinsulinemic hypoglycemia. Two case reports. AB - Neonatal hyperinsulinemic hypoglycemia must be suddenly and appropriately diagnosed and treated to prevent any further neurological dysfunction and damage. Therefore, we report two cases of our observation. Case 1: birth asphyxia, episodes of hypoglycemia after delivery, hyperinsulinism and reduced IGFBP1 blood concentration. Clinical and laboratory pictures resolved progressively after 8 days of life, perfusions were stopped and the neonate began to suck breast milk. Case 2: negative familial and perinatal history. On the 3rd day of life he developed cyanosis, hypotonia, tremors and hypoglycemia. He was discharged with a diagnosis of cerebral injury and neonatal hypoglycemia. At 1 year of life the child showed progressive and heavy neurological damage. The RMN of the brain showed: enlarged ventricles and liquor spaces around the brain, particularly in the frontal region. Hyperinsulinism was diagnosed in our Clinic. He began pharmacological treatment with Diazoxide that permitted euglycemia. The ammonium was normal and excluded glutamate dehydrogenase deficiency (mutation of GLUD1 gene); Diazoxide responsivity excluded mutations of SUR1 and KIR6.3 genes. At 9 years of life he showed motor and language retardation. Newborns with perinatal history of asphyxia may develop transient hyperinsulinism with absent neurological consequences. Persistent hypoglycemic or epileptic-like episodes, in particular on waking up, after meals or during banal infections, must be studied to reveal hyperinsulinism. PMID- 12131870 TI - Unresolved questions regarding the mediaeval child. PMID- 12131871 TI - [Transient neonatal hypocalcemia. Onset Manifestation of the 22q11.2 deletion syndrome]. PMID- 12131872 TI - [Neonatal thrombosis of the middle cerebral artery]. PMID- 12131873 TI - Evaluation of the spermiogram in the adolescents. PMID- 12131874 TI - Pubertal gynecomastia. PMID- 12131875 TI - The microarray: potential applications for ophthalmic research. AB - The microarray is a revolutionary technology combining molecular biology and computer technology in the high throughput, simultaneous analysis of global gene expression. It is emerging as a powerful and valuable research tool that holds great promise in elucidating the molecular mechanisms involved in complex diseases. The information gained may provide direction toward identifying appropriate targets for therapeutic intervention. Despite the enormous potential of this technology, however, a number of issues exist that complicate gene expression analysis and require further resolution. This paper reviews these issues as well as the conceptual, practical and statistical aspects of microarray technology, including its current use in research and clinical applications. Furthermore, the advantages and potential benefits of this technology in ophthalmic research are discussed, with particular attention to retinal diseases, and its possible application in the identification of genes involved in ocular disease progression that may serve as clinical markers or potential therapeutic targets. PMID- 12131876 TI - Identification of novel bovine RPE and retinal genes by subtractive hybridization. AB - PURPOSE: Understanding of the specialized function of the retinal pigment epithelium (RPE) can be aided by the identification and characterization of genes that are preferentially expressed in the RPE. With this aim, we undertook a systematic effort to identify and begin characterization of such genes. METHODS: A subtracted bovine RPE cDNA library was generated through subtractive hybridization using a single-stranded circular bovine RPE cDNA library as target and biotinylated mRNA from bovine heart and liver as alternate drivers. Approximately one thousand of the resulting subtracted cDNA clones were partially sequenced and analyzed, and a non-redundant set of one hundred of these cDNAs were examined for tissue expression pattern using a mini-Northern blot procedure and for identity by sequence analysis. RESULTS: The subtraction method successfully allowed the enrichment of cDNAs that are preferentially expressed in the RPE. Out of the analyzed clones, expression of forty-five clones was verifiable by Northern blotting. Of these, a significant proportion of cDNAs were preferentially expressed in the RPE. We observed that the expression of some subtracted cDNAs was restricted to the retina and no expression was detected in the RPE. These retinal clones were obtained in addition to RPE clones presumably because the initial RPE RNA population was contaminated with a small proportion of retinal RNA. Two thirds of the identified RPE and retinal cDNAs are likely to represent novel genes because they do not have homology to known genes in the databases. CONCLUSIONS: Genes that are specifically or predominantly expressed in the RPE/retina are likely to be important for retinal function. We have identified novel cDNAs from bovine RPE and retina by subtractive hybridization. These cDNAs can be used as starting material for the identification of corresponding human genes expressed in the RPE and retina. The human genes thus identified are likely to contain good candidate genes for retinal disease. PMID- 12131877 TI - Expression of bone morphogenetic proteins (BMP), BMP receptors, and BMP associated proteins in human trabecular meshwork and optic nerve head cells and tissues. AB - PURPOSE: Bone morphogenetic proteins (BMPs) are multi-functional cytokines that have wide ranging effects on a variety of cells and tissues. In the present study, we profile the expression of BMPs, BMP receptors, and BMP associated proteins in the human trabecular meshwork (TM) and optic nerve head (ONH), two tissues involved in glaucoma pathogenesis. METHODS: Total RNA was isolated and subjected to reverse transcriptase-polymerase chain reaction (RT-PCR) to examine the expression of BMP, BMP receptor, and BMP associated proteins in tissues and cultured cells from the human TM and ONH (ONH astrocytes and lamina cribrosa cells). Western immunoblotting was used to study the expression of BMP and BMP receptor proteins in cultured human TM and ONH cells. RESULTS: Both TM and ONH cells and tissues expressed mRNAs for BMP2, BMP4, BMP5, BMP7, BMP-RIA, BMP-RIB, BMP-RII, DRM (gremlin), follistatin, chordin and NMA (BAMBI). The proteins for BMP2, BMP4, BMP5, BMP7, and all three BMP receptors were expressed in cultured human TM and ONH cells. CONCLUSIONS: Members of the BMP family of genes, including BMPs, BMP receptors, and inhibitory BMP associated proteins are expressed in the human TM and ONH. These molecules may be involved in the normal formation and function of these tissues. Altered expression of members of this gene family may lead to functional changes in the TM and ONH. These genes and their respective signaling pathways merit further research to examine their possible role in glaucoma. PMID- 12131879 TI - Prosthodontic management of the curve of Spee: use of the Broadrick flag. AB - Proper management of the occlusal plane is an essential consideration when multiple long-span posterior restorations are designed. When restorations are added to an existing tooth arrangement characterized by rotated, tipped, or extruded teeth, excursive interferences may be incorporated, resulting in detrimental sequelae. The curve of Spee, which exists in the ideal natural dentition, allows harmony to exist between the anterior tooth and condylar guidance. An instrument called the Broadrick flag has been used to assist in the reproduction of tooth morphology that is commensurate with the curve of Spee when posterior restorations are designed; its use prevents the introduction of protrusive interferences. Consideration also must be given to lateral excursive movements when the occlusal plane is designed. In this article, the importance of the curve of Spee in prosthodontic and restorative dentistry is discussed, and a patient treatment demonstrating use of the Broadrick flag is described. PMID- 12131878 TI - Maxillary and mandibular overlay removable partial dentures for the treatment of posterior open-occlusal relationship: a clinical report. AB - This clinical report describes the use of maxillary and mandibular overlay removable partial dentures to treat a patient with class III skeletal malocclusion and a posterior open-occlusal relationship. Overlay removable partial denture therapy was used as an alternative to other options such as orthodontics and combined orthodontic/oral surgery procedures; it satisfied the esthetic and functional requirements of the patient and provided a stable occlusion. Overlay removable partial dentures are a reversible and relatively inexpensive treatment for patients with congenital or acquired anomalies, but the potential disadvantages of these prostheses include compromised esthetics when the dentures are removed; caries and periodontal disease as a result of poor oral hygiene; and veneer material fracture, debonding, discoloration, and wear. PMID- 12131880 TI - Esthetic provisional replacement of a single anterior tooth during the implant healing phase: a clinical report. AB - Replacement of a missing anterior tooth with an implant-assisted restoration rather than a fixed partial denture is considered common practice. Implant placement with a 2-stage surgical approach in the anterior area of the mouth may be imperative and require an esthetic provisional replacement for the missing tooth during the healing phase. This provisional restoration should help optimize the health of hard and soft tissues around the implant without exerting pressure on the residual ridge and maintain the position of the adjacent and opposing teeth. This clinical report presents a technique for fabricating an anterior fixed provisional restoration with composite reinforced with a leno-woven polyethylene ribbon. PMID- 12131881 TI - Application of a 3-dimensional measurement system to complete denture impressions. AB - STATEMENT OF PROBLEM: Little is known about tissue displacement at the tissue/denture base interface during impression making. PURPOSE: This study used a new 3-dimensional measurement system to analyze and compare 2 complete denture reline impression techniques (1 occluding and 1 digital) to determine which resulted in less displacement of the tissue/denture base interface during impression making. MATERIAL AND METHODS: The experiment included 10 completely edentulous subjects. For each subject, 3 mandibular casts were obtained: (1) a reference cast of the existing denture base, (2) a cast made with an occluding reline impression technique, and (3) a cast made with a digital reline impression technique. With the use of an optical 3-dimensional measurement system, the corresponding casts in a common coordinate system were analyzed geometrically. For each cast, the coordinates of the barycentric point and the high point were determined. For the same subject, the differences between the coordinates of the barycentric points of the 3 casts were calculated 2-by-2. The same calculation was performed for the high points. To determine whether these differences, which represent the displacements of the barycentric points and the high points, were statistically significant, the Wilcoxon signed-rank test for paired group comparisons was applied (P<.05). Cartographic differences among the casts from the same subject also were compared. RESULTS: The uncertainty of measurement for the location of the barycentric and highest point was +/-57 microm. The uncertainty in the differences among the cartographic displays was +/-0.1 mm. Vertically, the mean distance that separated the barycentric points obtained with the 2 impression methods was 0.04 mm. Cartography data showed that the greatest dimensional differences between the 2 impression techniques (minimum 0.25 mm; maximum 1.5 mm) were located near the retromolar pad and along the lingual flange. CONCLUSION: Within the limitations of this 3-dimensional in vivo study, displacement of the tissue/denture base interface was essentially equivalent with the use of an occluding and a digital mandibular impression technique. PMID- 12131882 TI - Evaluation of porosity in microwave-processed acrylic resin using a photographic method. AB - STATEMENT OF PROBLEM: The effect of microwave polymerization on the porosity of denture base resin has not been fully determined. PURPOSE: This study investigated the effect of microwave energy on the porosity of 2 heat-activated denture base resins. MATERIAL AND METHODS: Two heat-activated denture base resins, one conventional (Paladon 65) and one designed for microwave polymerization (Acron MC), were used to prepare 50 test specimens. Five groups of 10 specimens each were established: Group A (Paladon 65, water bath); Group B(1) (Paladon 65, short microwave cycle); Group B(2) (Paladon 65, long microwave cycle); Group C(1) (Acron MC, short microwave cycle); and Group C(2) (Acron MC, long microwave cycle). Half of the specimens in each group were 3 mm thick, the other half 6 mm thick. After being polymerized, specimens were cut so that 3 cross-sectional areas were formed (S(1), S(2), and S(3)). These surfaces were polished and photographed under a microscope at x100 magnification. On the developed photographs, the area of each pore was measured with a digital planimeter, and the total area of pores per surface was calculated in percentage form. The total number of pores on each surface and the topographical distribution of the pores also were recorded. A 3-factor repeated-measures analysis of variance was used to compare porosity data, and a 1-way analysis of variance was performed to determine possible interactions between groups based on material and specimen thickness (P<.05). The effect of surface area on porosity data was analyzed with the use of contrasts. RESULTS: Group A specimens exhibited no pores. In the thicker specimens of Groups B(1) and B(2), giant pores (area as great as 3.69 mm(2)) and small, gaseous pores of almost uniform shape and size were found. In Groups C(1) and C(2), only the smaller pores were found; these were not clinically significant. Of the observed surfaces, 75.3% were free of pores and 24.7% contained at least one pore. In a selected group of pore-bearing surfaces, the majority (81%) had pores located near the center. The thicker specimens in Group B exhibited the greatest amount of porosity (P<.0001); Group C specimens exhibited the least porosity. Repeated-measures analysis of variance showed that polymerization cycle had no effect on porosity (P=.19). The 3 other factors (material, specimen thickness, and surface) and all possible interactions among them were significant (P<.05). Among the surfaces, S(1) and S(2) exhibited the highest total mean value of porosity (0.71% and 0.74%, respectively) and S(3) the lowest (0.025%). S(3) showed a different pattern of porosity than S(1) and S(2). CONCLUSION: Within the limitations of this in vitro study, minor porosity was identified in thin and more severe porosity in thicker areas of conventional resin specimens that underwent microwave polymerization. The resin designed specifically for microwave polymerization exhibited no clinically significant porosity. PMID- 12131884 TI - The accuracy of dual-arch impressions: a pilot study. AB - STATEMENT OF PROBLEM: Dual-arch impression trays often are used for addition silicone final impressions of fixed prosthodontic preparations, but concerns about distortion of the impression are common because such trays lack rigidity. PURPOSE: This in vitro pilot study was designed to determine the accuracy of addition silicone impressions made with custom trays or made with either passive or stressed dual-arch trays. MATERIAL AND METHODS: Complete crown preparations of a mandibular molar, premolar, and incisor were made on a dentoform. These tooth preparations received flat, parallel indexes on the facial and lingual axial walls for accurate and reproducible positioning of a digital caliper. Gypsum dies were produced with an addition silicone impression material in either custom trays or dual-arch trays seated passively or with induced flexure (3 dies per tray group). The facio-lingual dimensions of the dies were measured with a digital caliper accurate to +/-5 microm and compared to the dimensions of the original preparations. Flexure in the latter group was induced by contact of the tray with a simulated torus, made of resin, in the lingual vestibule of the dentoform. Data were analyzed with 2-way analysis of variance and Tukey's multiple comparison test (alpha=.05). RESULTS: Dies fabricated with either the custom or passive dual-arch tray reproduced the facio-lingual dimensions of the preparations within a -27 to +13 microm range. Dies fabricated with the flexed dual-arch tray exhibited greater discrepancy, in the range of -47 to -67 microm relative to the preparations. Tray type was a significant factor (P=.002): the flexed tray group was significantly different than the other 2 groups, which did not differ from each other. CONCLUSION: Within the limitations of this pilot study, dual-arch impressions were comparable in accuracy to impressions made with custom trays. Accuracy was reduced, however, when the trays were flexed during closure of the arches. PMID- 12131883 TI - Investigation of microleakage at the interface between a Co-Cr based alloy and four polymeric veneering materials. AB - STATEMENT OF PROBLEM: . Marginal adaptation and resistance to microleakage are important factors for clinical success in fixed prosthodontics. Alloy corrosion that sometimes occurs under a veneer in the cervical area may result in cervical staining, a metallic taste, or even failure of the interface. PURPOSE: This study investigated cervical microleakage between a Co-Cr alloy and 4 indirect polymeric veneering materials used with different conditioning systems. MATERIAL AND METHODS: Sixteen metallic frameworks (copings) were obtained by fabricating 0.6 mm calibrated wax patterns on a master cast abutment. The wax patterns were provided with 0.4-mm beaded retention on the veneering surfaces and cast in a Co Cr based alloy (Biosil F) used for fixed partial dentures. The Co-Cr copings were divided equally into 4 groups and veneered with 4 polymeric materials (Signum, Solidex, Superpont C+B, and Targis). Three chemical conditioning systems (Siloc, Targis-Link, and Silicoater MD) were used with the respective veneering system recommended by the manufacturer; Conolar opaque was used for Superpont C+B. No control group was created. After 2 weeks of storage in distilled water at 37 degrees C, 2000 cycles at 5-55 degrees C, and 24 hours of storage in 0.5% basic fuchsine, specimens were embedded in clear liquid casting resin and sectioned along a perpendicular cervical-incisal plane through the middle of the cervical collar. The surfaces of the resulting sections were examined in the cervical area with a metallurgical microscope to detect dye penetration. Leakage was quantitatively evaluated with the use of a scoring system (established by the authors) that indicated the presence/absence of dye in 3 regions of the cervical interface from the collar to the incisal margin. Scores were compared and analyzed with the use of 1-way analysis of variance followed by post-hoc Tukey's honestly significant difference test (P=.05). RESULTS: Superpont C+B was associated with the highest microleakage scores (3.75 +/- 0.5). The least microleakage at the interface was produced by Targis (1 +/- 0.816), followed by Solidex (2.5 +/- 1.0) and Signum (2.25 +/- 0.975). Only the difference between Targis and Superpont C+B was significant (P<.05). CONCLUSION: Within the limitations of this in vitro study, the extent of cervical microleakage between the coping and veneer depended on the particular polymeric material used for veneering. The extremes of the study were Targis/Targis-Link (lowest leakage scores) and Superpont C+B (highest leakage scores). Differences among the chemical retention systems tested were not significant. PMID- 12131885 TI - Effect of irrigation solutions on dentin bonding agents and restorative shear bond strength. AB - STATEMENT OF PROBLEM: The presence of irrigation solution prior to the application of dentin bonding agents may be one source of contamination and may adversely affect bond strength. PURPOSE: The purpose of this in vitro investigation was to evaluate the effect of irrigation solutions of different purity levels on the shear bond strength of a hybrid composite to dentin. MATERIAL AND METHODS: Forty extracted, intact human molars were hand-scaled. All soft tissue was removed, and the teeth were stored in room-temperature tap water for 1 week. Subsequently, each tooth was embedded in autopolymerizing acrylic resin with the coronal portion exposed. After complete polymerization of the resin, the dentin surfaces were exposed, and the crown was cut longitudinally on all 4 sides to produce flat dentin surfaces for bonding. The prepared specimens were assigned to 4 groups of irrigation solutions: tap water (control), sterile water, 0.9% NaCl irrigation solution, and filtered water. Within each irrigation group were 4 dentin bonding agent subgroups: Prime & Bond NT, One-Step, Single Bond Adhesive, and OptiBond Solo. The dentin surfaces (n = 10 per irrigation/bonding agent pair) were etched with 37% phosphoric acid, rinsed with the assigned irrigation solution for 15 seconds, and then dried. Plastic cylinders (3 mm long x 3 mm inner diameter) were filled with a hybrid composite (Herculite XRV) and bonded to the dentin. Specimens were loaded in a universal testing machine, and shear force was applied to the base of the composite cylinders, parallel to the dentin surfaces, at a crosshead speed of 0.05 in/min until fracture. Failure loads were recorded. Bond strength data were compared with analysis of variance (P<.05). Post-hoc comparisons of means were performed with t tests and P values adjusted for multiple comparisons (Bonferroni method). RESULTS: Tap water irrigation (control) produced no significant difference in bond strengths for all dentin bonding agents tested. When other irrigation solutions were used, One-Step consistently produced significantly lower bond strengths than other dentin bonding agents (P<.05). No preferential choice of irrigation solution could be made for Prime & Bond or Single Bond Adhesive. However, when One-Step or Solo was used, irrigation with normal saline significantly reduced bond strengths (P<.05). CONCLUSION: Within the limitations of this study, the bond strength to dentin of a hybrid composite irrigated with different solutions was dependent on the dentin bonding agent used. PMID- 12131886 TI - Interfacial micromorphological differences in hybrid layer formation between water- and solvent-based dentin bonding systems. AB - STATEMENT OF PROBLEM: Many dentin bonding systems of different compositions, and in particular containing different solvents, have been introduced to the market. Their effect on the quality of the interface requires clarification by means of comparative trials. PURPOSE: This study investigated micromorphological differences in hybrid layer formation with a variety of commercially available water- or solvent-based dentin bonding products and their recommended compomers. MATERIAL AND METHODS: Five bonding systems were used on groups of 10 teeth each as follows: group I, acetone-based system used with 36% phosphoric acid; group II, a different acetone-based system containing nano-sized particles for filler loading and used with a non-rinsing conditioner containing maleic acid; group III, the acetone-based system of group II used with 36% phosphoric acid (the only difference in the treatment for groups II and III was the acid etching system); group IV, a mixed-solvent-based system (water/ethanol) used with 37% phosphoric acid; and group V, a water-based system used with 37% phosphoric acid. Each bonding system was covered with the recommended compomer. Class I occlusal preparations were made in extracted teeth and restored with one of the above systems. Five specimens of each group were studied with optical microscopy after staining. Scanning electron microscopy was used to examine the interface of the bonding system/dentin of the other 5 teeth in each group. The optical microscopy measurements were made with a 10 x 10 reticle. A micron mark with scale was used for the scanning electron microscope. All measurements were made in microm. The following criteria were used to define a good interface: absence of voids between the different parts of the interface, uniformity of the hybrid layer, good opening of the tubuli orifices, and tag adherence to the tubuli walls. RESULTS: Morphological differences were found at the interface depending on dentin treatment and adhesive composition. The acetone-containing systems were associated with a continuous, gap-free hybrid layer that was linked intimately with the dentin. The tags adhered well to the tubuli walls and were often joined by side branches. In the water-based solvent systems, a lack of contact was visible between the resin tags and the tubuli walls, with some incompletely filled tubuli and some gaps in the hybrid layer. The 2 observational methods used, optical and scanning electron microscopy, proved to be complementary. CONCLUSION: Within the limitations of this study, use of the acetone-based systems after phosphoric acid etching resulted in a continuous, thick hybrid layer with reverse-cone-shaped tags in close contact with the tubuli walls. Use of the water-based systems resulted in a thinner hybrid layer with some incompletely sealed dentinal tubules. PMID- 12131887 TI - The effect of filler loading and morphology on the mechanical properties of contemporary composites. AB - STATEMENT OF PROBLEM: Little information exists regarding the filler morphology and loading of composites with respect to their effects on selected mechanical properties and fracture toughness. PURPOSE: The objectives of this study were to: (1) classify commercial composites according to filler morphology, (2) evaluate the influence of filler morphology on filler loading, and (3) evaluate the effect of filler morphology and loading on the hardness, flexural strength, flexural modulus, and fracture toughness of contemporary composites. MATERIAL AND METHODS: Field emission scanning electron microscopy/energy dispersive spectroscopy was used to classify 3 specimens from each of 14 commercial composites into 4 groups according to filler morphology. The specimens (each 5 x 2.5 x 15 mm) were derived from the fractured remnants after the fracture toughness test. Filler weight content was determined by the standard ash method, and the volume content was calculated using the weight percentage and density of the filler and matrix components. Microhardness was measured with a Vickers hardness tester, and flexural strength and modulus were measured with a universal testing machine. A 3 point bending test (ASTM E-399) was used to determine the fracture toughness of each composite. Data were compared with analysis of variance followed by Duncan's multiple range test, both at the P<.05 level of significance. RESULTS: The composites were classified into 4 categories according to filler morphology: prepolymerized, irregular-shaped, both prepolymerized and irregular-shaped, and round particles. Filler loading was influenced by filler morphology. Composites containing prepolymerized filler particles had the lowest filler content (25% to 51% of filler volume), whereas composites containing round particles had the highest filler content (59% to 60% of filler volume). The mechanical properties of the composites were related to their filler content. Composites with the highest filler by volume exhibited the highest flexural strength (120 to 129 MPa), flexural modulus (12 to 15 GPa), and hardness (101 to 117 VHN). Fracture toughness was also affected by filler volume, but maximum toughness was found at a threshold level of approximately 55% filler volume. CONCLUSION: Within the limitations of this study, the commercial composites tested could be classified by their filler morphology. This property influenced filler loading. Both filler morphology and filler loading influenced flexural strength, flexural modulus, hardness, and fracture toughness. PMID- 12131888 TI - In vitro wear rates of materials under different loads and varying pH. AB - STATEMENT OF PROBLEM: Despite the need for information about the wear characteristics of restorative materials, there have been few systemic studies of the factors that influence the rate of material wear. PURPOSE: This study compared the wear rates of enamel and 3 tooth-colored restorative materials under different loads (0, 3.2, 6.7, and 9.95 kg) and pH levels (1.2, 3.3, and 7.0). MATERIAL AND METHODS: An electromechanical tooth wear machine was used so that standard restorations representing 3 materials could be worn by opposing enamel under controlled conditions. The wear rates of enamel, composite (Z100), a conventional glass ionomer cement (Fuji IX), and a resin-modified glass ionomer cement (Fuji II LC) were compared at a range of loads (0 to 9.95 kg) and pH levels (1.2 to 7.0) and also at different sites across each restoration. Ten specimens were randomly assigned to each experimental group. Wear assessment was performed with a modified light microscope to quantify the height changes at defined points across wear facets. Four-way analysis of variance was used to compare wear rates among materials, pH levels, loads, and sites. Post-hoc t tests identified significant differences between specific pairs of experimental conditions (P<.05). RESULTS: The wear rates of enamel and the other test materials varied significantly with pH (P<.0001), load (P<.0001), and type of material (P<.0001). Enamel wear was influenced most by varied pH, whereas the composite was least affected by acid. The conventional glass ionomer cement was more susceptible than the composite to the effects of varied pH; the acid susceptibility of the resin-modified glass ionomer cement was generally between that of the composite and conventional glass ionomer cement. Enamel and the conventional glass ionomer cement were affected similarly by load. The composite was more resistant than the conventional glass ionomer cement to wear at higher loads; the resin-modified glass ionomer cement exhibited intermediate load resistance. CONCLUSION: Within the limitations of this study, the 3 test materials were more resistant than enamel to acid, with the composite demonstrating the lowest susceptibility to acid. The acid- and load-resistance of the resin-modified glass ionomer cement was consistently less than that of the composite and greater than that of the conventional glass ionomer cement. PMID- 12131889 TI - Variation in color between intended matched shade and fabricated shade of dental porcelain. AB - STATEMENT OF PROBLEM: The total quantifiable color difference between shade matching and shade duplication has not been investigated formally. PURPOSE: The purpose of this in vitro study was to evaluate and compare the color difference of the total color replication process and the direction of the individual color parameters for 3 dental porcelain shade-matching systems. MATERIAL AND METHODS: The shade of 11 porcelain master disks was determined visually and instrumentally using 3 porcelain shade-matching systems: (1) Vita Lumin/Vita VMK 68, (2) Vitapan 3D-Master/Vita Omega 900, and (3) Shofu ShadeEye-EX/Vintage Halo. Corresponding porcelain disks made of 4.5 mm opaque and 1 mm dentin porcelain were fabricated with each of the porcelain systems. The colors of the master disks and fabricated disks (CIE L* a* b* coordinates) were measured with a spectroradiometer with a 45 degrees /0 degrees configuration. Repeated-measures analysis of variance was used to evaluate within-group differences among the porcelain systems for the total color difference (Delta E) and direction of the color parameters (Delta L, Delta a, and Delta b). The Ryan-Einot-Gabriel-Welsch multiple range test was used for post-hoc analysis (alpha=.05). RESULTS: The largest mean Delta E was recorded for the Vitapan 3D-Master system, which was significantly different from the other systems (P=.0024). A significant difference was found between the interaction of the different systems and the direction of color (P=.0024). The amount of change within each color parameter was dependent on the porcelain system, as well as the amount of change among the color parameters. CONCLUSION: Within the limitations of this study, the results suggest that reliable delivery of a properly matched restoration to existing porcelain restorations cannot be ensured regardless of the shade assessment method used (visual or computer-generated). PMID- 12131890 TI - Influence of impaired mastication on nutrition. AB - It has been suggested that people who suffer from impaired masticatory function may adapt food consistency to their oral status (which may lead to deficient nutrient intake) or rely on the digestive system to compensate for the lack of oral preparation of food (which may increase the likelihood of digestive diseases and decrease gut absorption). Masticatory deficiency thus may be detrimental to health. This article reviews evidence of the effects of masticatory deficiency on nutrition. The selection of relevant literature was based on Medline queries using the following key words: mastication, nutrition, digestion, diet, and disease risk. Earlier work not listed in Medline but related to the subject also was reviewed. Only publications available in English were selected for inclusion. It is difficult to draw conclusions from many of the reviewed studies due to issues related to study design, confounding variables, and the subjective nature of the measurements. In particular, data supporting a link between masticatory function and deficient dietary intake often are based on relatively weak correlations and cannot confer a causal relationship. PMID- 12131891 TI - Endodontic failure caused by inadequate restorative procedures: review and treatment recommendations. AB - A review of the literature was performed to determine whether prompt placement of coronal restorations, including sealing and placement of posts and cores, can positively influence the long-term prognosis of teeth after root canal therapy. Both hand and MEDLINE searches were employed to identify peer-reviewed articles on radicular apical integrity after coronal restorations, especially where root canal space was used for post and core fabrication. A total of 41 articles published between 1969 and 1999 (the majority from the 1990s) were reviewed. The literature suggests that the prognosis of root canal-treated teeth can be improved by sealing the canal and minimizing the leakage of oral fluids and bacteria into the periradicular areas as soon as possible after the completion of root canal therapy. PMID- 12131892 TI - Fabrication of an educational model for patients considering prostheses retained by osseointegrated implants. AB - Patients considering the use of implant-retained craniofacial prostheses are faced with a large body of new information. This article describes the fabrication of a 3-dimensional teaching aid that helps educate such patients about the role of osseointegration in prosthetic treatment. The model illustrates the stages of auricular prosthesis fabrication and can be used to demonstrate proper prosthesis hygiene. PMID- 12131893 TI - A simple method of making an implant-level impression when presented with limited space, unfavorable implant positions, or problematic implant angulations. AB - Making an implant-level impression for the purpose of abutment selection when implants are placed in limited space, unfavorable positions, or compromising angulations can be a time-consuming procedure. An impression procedure is presented that makes use of either prefabricated screw-retained titanium implant index copings or plastic snap-on implant index copings to help resolve problematic implant placement. Both the titanium and plastic implant index copings are easy to modify and therefore make impression procedures more predictable. PMID- 12131894 TI - Bonded provisional restorations for esthetic soft tissue support in single implant treatment. AB - During the early phases of single-implant treatment, clinician needs and patient expectations can be challenging. This article describes a technique for making a simple bonded provisional restoration. In addition to minimizing patient discomfort without increasing laboratory costs, the technique enables soft tissue preservation through all treatment phases, from tooth loss to implant loading. PMID- 12131895 TI - A transfer method for multiple cement-retained implant restorations. AB - This article describes a time-saving method for impression making and abutment preparation performed as a laboratory procedure on multiple cement-retained ITI implants. PMID- 12131896 TI - Block-out prior to pick-up impression of overdenture with ERA attachments. PMID- 12131897 TI - Pinlay to prevent and restore excessive wear in anterior teeth. PMID- 12131898 TI - A customized guide for transferring angled abutments. PMID- 12131899 TI - Customizing the size of toothbrush handles for patients with restricted hand and finger movement. PMID- 12131900 TI - Leading from the heart. PMID- 12131901 TI - Supreme Court decides US Airways v. Burnett. PMID- 12131902 TI - To change or not to change? You have the option. PMID- 12131903 TI - Another FDA warning: Kava supplements. PMID- 12131904 TI - Mission possible: selecting, buying, and implementing case management software. PMID- 12131905 TI - Moral distress, part 2: I still can't take it! PMID- 12131907 TI - HMOs and the future of telemedicine and telehealth: Part 2. PMID- 12131906 TI - Outcomes and depression. PMID- 12131908 TI - The Case Report: a newsletter from the Case Management Society of America: the presidents' messages. PMID- 12131909 TI - Using your head while case managing from your heart. PMID- 12131910 TI - Pressure ulcers: assessment, prevention, and compliance. PMID- 12131911 TI - Direct outcomes of case management: involvement/participation, empowerment, and knowledge: The Council for Case Management Accountability's second state of the science paper. PMID- 12131912 TI - Case management for pediatric respiratory syncytial virus outcomes. PMID- 12131913 TI - The case for hand therapy. PMID- 12131914 TI - Membrane proteins: functional and structural studies using reconstituted proteoliposomes and 2-D crystals. AB - Reconstitution of membrane proteins into lipid bilayers is a powerful tool to analyze functional as well as structural areas of membrane protein research. First, the proper incorporation of a purified membrane protein into closed lipid vesicles, to produce proteoliposomes, allows the investigation of transport and/or catalytic properties of any membrane protein without interference by other membrane components. Second, the incorporation of a large amount of membrane proteins into lipid bilayers to grow crystals confined to two dimensions has recently opened a new way to solve their structure at high resolution using electron crystallography. However, reconstitution of membrane proteins into functional proteoliposomes or 2-D crystallization has been an empirical domain, which has been viewed for a long time more like "black magic" than science. Nevertheless, in the last ten years, important progress has been made in acquiring knowledge of lipid-protein-detergent interactions and has permitted to build upon a set of basic principles that has limited the empirical approach of reconstitution experiments. Reconstitution strategies have been improved and new strategies have been developed, facilitating the success rate of proteoliposome formation and 2-D crystallization. This review deals with the various strategies available to obtain proteoliposomes and 2-D crystals from detergent-solubilized proteins. It gives an overview of the methods that have been applied, which may be of help for reconstituting more proteins into lipid bilayers in a form suitable for functional studies at the molecular level and for high-resolution structural analysis. PMID- 12131915 TI - Presence of the RHD pseudogene and the hybrid RHD-CE-D(s) gene in Brazilians with the D-negative phenotype. AB - The molecular basis for RHD pseudogene or RHD Psi is a 37-bp insertion in exon 4 of RHD. This insertion, found in two-thirds of D-negative Africans, appears to introduce a stop codon at position 210. The hybrid RHD-CE-Ds, where the 3' end of exon 3 and exons 4 to 8 are derived from RHCE, is associated with the VS+V- phenotype, and leads to a D-negative phenotype in people of African origin. We determined whether Brazilian blood donors of heterogeneous ethnic origin had RHD Psi and RHD-CE-Ds. DNA from 206 blood donors were tested for RHD Psi by a multiplex PCR that detects RHD, RHD Psi and the C and c alleles of RHCE. The RHD genotype was determined by comparison of size of amplified products associated with the RHD gene in both intron 4 and exon 10/3'-UTR. VS was determined by amplification of exon 5 of RHCE, and sequencing of PCR products was used to analyze C733G (Leu245Val). Twenty-two (11%) of the 206 D-negative Brazilians studied had the RHD Psi, 5 (2%) had the RHD-CE-Ds hybrid gene associated with the VS+V- phenotype, and 179 (87%) entirely lacked RHD. As expected, RHD was deleted in all the 50 individuals of Caucasian descent. Among the 156 individuals of African descent, 22 (14%) had inactive RHD and 3% had the RHD-CE-Ds hybrid gene. These data confirm that the inclusion of two different multiplex PCR for RHD is essential to test the D-negative Brazilian population in order to avoid false positive typing of polytransfused patients and fetuses. PMID- 12131917 TI - Respiratory panic disorder subtype and sensitivity to the carbon dioxide challenge test. AB - The aim of the present study was to verify the sensitivity to the carbon dioxide (CO2) challenge test of panic disorder (PD) patients with respiratory and nonrespiratory subtypes of the disorder. Our hypothesis is that the respiratory subtype is more sensitive to 35% CO2. Twenty-seven PD subjects with or without agoraphobia were classified into respiratory and nonrespiratory subtypes on the basis of the presence of respiratory symptoms during their panic attacks. The tests were carried out in a double-blind manner using two mixtures: 1) 35% CO2 and 65% O2, and 2) 100% atmospheric compressed air, 20 min apart. The tests were repeated after 2 weeks during which the participants in the study did not receive any psychotropic drugs. At least 15 of 16 (93.7%) respiratory PD subtype patients and 5 of 11 (43.4%) nonrespiratory PD patients had a panic attack during one of two CO2 challenges (P = 0.009, Fisher exact test). Respiratory PD subtype patients were more sensitive to the CO2 challenge test. There was agreement between the severity of PD measured by the Clinical Global Impression (CGI) Scale and the subtype of PD. Higher CGI scores in the respiratory PD subtype could reflect a greater sensitivity to the CO2 challenge due to a greater severity of PD. Carbon dioxide challenges in PD may define PD subtypes and their underlying mechanisms. PMID- 12131916 TI - Six years of treatment with the HELP system of a patient with familial hypercholesterolemia. AB - The purpose of the present report is to demonstrate the long-term efficacy and safety of heparin-induced extracorporeal lipoprotein precipitation (HELP) of LDL c and fibrinogen in the management of familial hypercholesterolemia. From June 1992 to June 1998 a 22-year-old young male patient with familial hypercholesterolemia (double heterozygote for C660X and S305C) resistant to medication and diet and with symptomatic coronary artery disease (angina) was treated weekly with 90-min sessions of the HELP system. The patient had also been previously submitted to right coronary artery angioplasty. The efficacy of the method was evaluated by comparing the reduction of total cholesterol, LDL-c and fibrinogen before and after the sessions and before and after initiation of the study (data are reported as averages for each year). During the study, angina episodes disappeared and there were no detectable adverse effects of the treatment. Total cholesterol (TC), fibrinogen, and LDL-c decreased significantly after each session by 59.6, 66.1 and 64%, respectively. HDL-c showed a nonsignificant reduction of 20.4%. Comparative mean values pre- and post treatment values in the study showed significant differences: TC (488 vs 188 mg/dl), LDL-c (416.4 vs 145 mg/dl), and fibrinogen (144.2 vs 57.4 mg/dl). There was no significant change in HDL-c level: 29.4 vs 23 mg/dl. These data show that the HELP system, even for a long period of time, is a safe and efficient mode of treatment of familial hypercholesterolemia and is associated with disappearance of angina symptoms. PMID- 12131918 TI - Fungal infections in marrow transplant recipients under antifungal prophylaxis with fluconazole. AB - Fungal infection is one of the most important causes of morbidity and mortality in bone marrow transplant (BMT) recipients. The growing incidence of these infections is related to several factors including prolonged granulocytopenia, use of broad-spectrum antibiotics, conditioning regimens, and use of immunosuppression to avoid graft-versus-host disease (GvHD). In the present series, we report five cases of invasive mold infections documented among 64 BMT recipients undergoing fluconazole antifungal prophylaxis: 1) A strain of Scedosporium prolificans was isolated from a skin lesion that developed on day +72 after BMT in a chronic myeloid leukemic patient. 2) Invasive pulmonary aspergillosis (Aspergillus fumigatus) was diagnosed on day +29 in a patient with a long period of hospitalization before being transplanted for severe aplastic anemia. 3) A tumoral lung lesion due to Rhizopus arrhizus (zygomycosis) was observed in a transplanted patient who presented severe chronic GvHD. 4) A tumoral lesion due to Aspergillus spp involving the 7th, 8th and 9th right ribs and local soft tissue was diagnosed in a BMT patient on day +110. 5) A patient with a history of Ph1-positive acute lymphocytic leukemia exhibited a cerebral lesion on day +477 after receiving a BMT during an episode of severe chronic GvHD. At that time, blood and spinal fluid cultures yielded Fusarium sp. Opportunistic infections due to fungi other than Candida spp are becoming a major problem among BMT patients receiving systemic antifungal prophylaxis with fluconazole. PMID- 12131919 TI - Relationship between microalbuminuria and cardiac structural changes in mild hypertensive patients. AB - The aim of this study was to determine the relationship between urinary albumin excretion (UAE), cardiac structural changes upon echocardiography and 24-h ambulatory blood pressure (ABPM) levels. Twenty mild hypertensive patients (mean age 56.8 +/- 9.6 years) were evaluated. After 2 weeks of a washout period of all antihypertensive drugs, all patients underwent an echocardiographic evaluation, a 24-h ABPM and an overnight urine collection. Systolic and diastolic blood pressure during 24-h ABPM was 145 +/- 14/91 +/- 10 mmHg (daytime) and 130 +/- 14/76 +/- 8 mmHg (nighttime), respectively. Seven (35%) patients presented UAE > or = 15 microg/min, and for the whole group, the geometric mean value for UAE was 10.2 x// 3.86 microg/min. Cardiac measurements showed mean values of interventricular septum thickness (IVS) of 11 +/- 2.3 mm, left ventricular posterior wall thickness (PWT) of 10 +/- 2.0 mm, left ventricular mass (LVM) of 165 +/- 52 g, and left ventricular mass index (LVMI) of 99 +/- 31 g/m2. A forward stepwise regression model indicated that blood pressure levels did not influence UAE. Significant correlations were observed between UAE and cardiac structural parameters such as IVS (r = 0.71, P<0.001), PWT (r = 0.64, P<0.005), LVM (r = 0.65, P<0.005) and LVMI (r = 0.57, P<0.01). Compared with normoalbuminuric patients, those who had microalbuminuria presented higher values of all cardiac parameters measured. The predictive positive and negative values of UAE > or = 15 microg/min for the presence of geometric cardiac abnormalities were 75 and 91.6%. These data indicate that microalbuminuria in essential hypertension represents an early marker of cardiac structural damage. PMID- 12131920 TI - Use of single photon emission computed tomography and magnetic resonance to evaluate central nervous system involvement in patients with juvenile systemic lupus erythematosus. AB - The objective of the present study was to identify the single photon emission computed tomography (SPECT) and magnetic resonance (MR) findings in juvenile systemic lupus erythematosus (JSLE) patients with CNS involvement and to try to correlate them with neurological clinical history data and neurological clinical examination. Nineteen patients with JSLE (16 girls and 3 boys, mean age at onset 9.2 years) were submitted to neurological examination, electroencephalography, cerebrospinal fluid analysis, SPECT and MR. All the evaluations were made separately within a period of 15 days. SPECT and MR findings were analyzed independently by two radiologists. Electroencephalography and cerebrospinal fluid analysis revealed no relevant alterations. Ten of 19 patients (53%) presented neurological abnormalities including present or past neurological clinical history (8/19, 42%), abnormal neurological clinical examination (5/19, 26%), and abnormal SPECT or MR (8/19, 42% and 3/19, 16%, respectively). The most common changes in SPECT were cerebral hypoperfusion and heterogeneous distribution of blood flow. The most common abnormalities in MR were leukomalacia and diffuse alterations of white matter. There was a correlation between SPECT and MR (P<0.05). We conclude that SPECT and MR are complementary and useful exams in the evaluation of neurological involvement of lupus. PMID- 12131921 TI - In vitro antimicrobial activity of a new series of 1,4-naphthoquinones. AB - The antibacterial activity of a series of 1,4-naphthoquinones was demonstrated. Disk diffusion tests were carried out against several Gram-positive and Gram negative bacteria. The compound 5-amino-8-hydroxy-1,4-naphthoquinone was the most effective, presenting inhibition zones measuring 20 mm against staphylococci, streptococci and bacilli at 50 microg/ml. Methicillin-resistant Staphylococcus aureus and several clinical isolates of this bacterium were also inhibited. Naphthazarin, 5-acetamido-8-hydroxy-1,4-naphthoquinone, and 2,3-diamino-1,4 naphthoquinone were the next most active compounds. The minimal inhibitory concentration of the active compounds was determined against S. aureus, ranging from 30 to 125 microg/ml. All compounds presented a minimal bactericidal concentration higher than 500 microg/ml, indicating that their effect was bacteriostatic. The EC50, defined as the drug concentration that produces 50% of maximal effect, was 8 microg/ml for 5-amino-8-hydroxy-1,4-naphthoquinone against S. aureus, S. intermedius, and S. epidermidis. These results indicate an effective in vitro activity of 5-amino-8-hydroxy-1,4-naphthoquinone and encourage further studies for its application in antibiotic therapy. PMID- 12131922 TI - DNA topoisomerase inhibitors: biflavonoids from Ouratea species. AB - Topoisomerase inhibitors are agents with anticancer activity. 7"-O-Methyl agathisflavone (I) and amentoflavone (II) are biflavonoids and were isolated from the Brazilian plants Ouratea hexasperma and O. semiserrata, respectively. These biflavonoids and the acetyl derivative of II (IIa) are inhibitors of human DNA topoisomerases I at 200 microM, as demonstrated by the relaxation assay of supercoiled DNA, and only agathisflavone (I) at 200 microM also inhibited DNA topoisomerases II-alpha, as observed by decatenation and relaxation assays. The biflavonoids showed concentration-dependent growth inhibitory activities on Ehrlich carcinoma cells in 45-h culture, assayed by a tetrazolium method, with IC50 = 24 +/- 1.4 microM for I, 26 +/- 1.1 microM for II and 10 +/- 0.7 microM for IIa. These biflavonoids were assayed against human K562 leukemia cells in 45 h culture, but only I showed 42% growth inhibitory activity at 90 microM. Our results suggest that biflavonoids are targets for DNA topoisomerases and their cytotoxicity is dependent on tumor cell type. PMID- 12131923 TI - Effect of a Brazilian regional basic diet on the prevalence of caries in rats. AB - The aim of the present study was to determine the effect of a regional basic diet (RBD) on the prevalence of caries in the molar teeth of rats of both sexes aged 23 days. The animals were divided into six groups of 10 rats each receiving the following diets for 30 and 60 days after weaning: RBD, a cariogenic diet, and a commercial diet. The prevalence and penetration of caries in the molar teeth of the rats was then analyzed. The RBD produced caries in 37.5% of the teeth of animals fed 30 days, and in 83.4% of animals fed 60 days, while the cariogenic diet produced caries in 72.5% and 77.5% of the teeth of animals fed 30 and 60 days, respectively. Rats fed the RBD for 30 days had caries in the enamel in 38% of their teeth, 48% had superficial dentin caries, and 7.5% moderate dentin caries. The effect of the RBD did not differ significantly from that of the cariogenic diet in terms of the presence of caries in rats fed 60 days. The penetration depth of the caries produced by the RBD was the same as that produced by the cariogenic diet. Our results show that the RBD has the same cariogenic potential as the cariogenic diet. Since the RBD is the only option for the low income population, there should be a study of how to compensate for the cariogenicity of this diet. PMID- 12131924 TI - Immune response induced in mice oral immunization with cowpea severe mosaic virus. AB - There is increasing interest in the immune response induced by plant viruses since these could be used as antigen-expressing systems in vaccination procedures. Cowpea severe mosaic virus (CPSMV), as a purified preparation (300 g of leaves, 2 weeks post-inoculation), or crude extract from cowpea (Vigna unguiculata) leaves infected with CPSMV both administered by gavage to Swiss mice induced a humoral immune response. Groups of 10 Swiss mice (2-month-old females) were immunized orally with 10 daily doses of either 50 microg viral capsid protein (boosters of 50 microg at days 21 and 35 after immunization) or 0.6 mg protein of the crude extract (boosters of 0.6 mg at days 21 and 35 after immunization). Anti-CPSMV antibodies were quantified by ELISA in pooled sera diluted at least 1:400 at days 7, 14, 21, 28, 35 and 42 after the 10th dose. IgG and IgA against CPSMV were produced systemically, but IgE was not detected. No synthesis of specific antibodies against the proteins of leaf extracts from V. unguiculata, infected or not with CPSMV, was detected. The use of CPSMV, a plant infecting virus that apparently does not induce a pathogenic response in animals, induced a humoral and persistent (at least 6 months) immune response through the administration of low antigen doses by gavage. These results raise the possibility of using CPSMV either as a vector for the production of vaccines against animal pathogens or in quick and easy methods to produce specific antisera for viral diagnosis. PMID- 12131925 TI - Evaluation of behavioral states among morning and evening active healthy individuals. AB - The Horne-Ostberg questionnaire partly covers some factors that may be important determinants of peak time and characterize patterns of behavior. We conducted a study for the evaluation of self-reported behavioral states (hunger sensation, availability for study, physical exercise, solving daily problems, and time preferences) as expressions of underlying cyclic activity. Three hundred and eighteen community subjects without history of medical, psychiatric, or sleep disorders were evaluated in a cross-sectional design. A self-report about daily highest level of activity was used to categorize individuals into morning, evening, and indifferently active. Time-related behavioral states were evaluated with 23 visual analog questions. The responses to most analogic questions were significantly different between morning and evening active subjects. Logistic regression analysis identified a group of behaviors more strongly associated with the self-reported activity pattern (common wake up time, highest subjective fatigue, as well as wake up, bedtime, exercise and study preferences). These findings suggested that the patterns of activity presented by normal adults were related to specific common behavioral characteristics that may contribute to peak time. PMID- 12131926 TI - Reflex control of arterial pressure and heart rate in short-term streptozotocin diabetic rats. AB - Impaired baroreflex sensitivity in diabetes is well described and has been attributed to autonomic diabetic neuropathy. In the present study conducted on acute (10-20 days) streptozotocin (STZ)-induced diabetic rats we examined: 1) cardiac baroreflex sensitivity, assessed by the slope of the linear regression between phenylephrine- or sodium nitroprusside-induced changes in arterial pressure and reflex changes in heart rate (HR) in conscious rats; 2) aortic baroreceptor function by means of the relationship between systolic arterial pressure and aortic depressor nerve (ADN) activity, in anesthetized rats, and 3) bradycardia produced by electrical stimulation of the vagus nerve or by the iv injection of methacholine in anesthetized animals. Reflex bradycardia (-1.4 +/- 0.1 vs -1.7 +/- 0.1 bpm/mmHg) and tachycardia (-2.1 +/- 0.3 vs -3.0 +/- 0.2 bpm/mmHg) were reduced in the diabetic group. The gain of the ADN activity relationship was similar in control (1.7 +/- 0.1% max/mmHg) and diabetic (1.5 +/- 0.1% max/mmHg) animals. The HR response to vagal nerve stimulation with 16, 32 and 64 Hz was 13, 16 and 14% higher, respectively, than the response of STZ treated rats. The HR response to increasing doses of methacholine was also higher in the diabetic group compared to control animals. Our results confirm the baroreflex dysfunction detected in previous studies on short-term diabetic rats. Moreover, the normal baroreceptor function and the altered HR responses to vagal stimulation or methacholine injection suggest that the efferent limb of the baroreflex is mainly responsible for baroreflex dysfunction in this model of diabetes. PMID- 12131927 TI - The effect of feeding on the respiratory activity of the sloth. AB - The aim of the present study was to confirm whether feeding influences the resting breathing rate and to observe possible alterations in blood gas and pH levels produced by feeding in unanesthetized sloths (Bradypus variegatus). Five adult male sloths (4.1 +/- 0.6 kg) were placed daily in an experimental chair for a period of at least 4 h for sitting adaptation. Five measurements were made for each sloth. However, the sloths one, two and five were studied once and the sloths three and four were studied twice. Breathing rate was determined with an impedance meter and the output signal was digitized. Arterial blood samples were collected for blood gas analysis with a BGE electrolytes analyzer and adjusted for the animal's body temperature and hemoglobin content. The data are reported as mean +/- SD and were collected during the resting period (8:00-10:00 h) and during the feeding period (16:00-18:00 h). The mean breathing rate increased during mastication of ymbahuba leaves (rest: 5.0 +/- 1, feeding: 10 +/- 1 bpm). No significant alterations were observed in arterial pH (rest: 7.42 +/- 0.05, feeding: 7.45 +/- 0.03), PCO2 (rest: 35.2 +/- 5.3, feeding: 33.3 +/- 4.4 mmHg) or PO2 (rest: 77.5 +/- 8.2, feeding: 78.4 +/- 5.2 mmHg) levels. These results indicate that in unanesthetized sloths 1) feeding evokes an increase in breathing rate without a significant change in arterial pH, PCO2 or PO2 levels, and 2) the increase in breathing rate produced by feeding probably is due to the act of mastication. PMID- 12131928 TI - Stimulatory effects of adenosine on prolactin secretion in the pituitary gland of the rat. AB - We investigated the effects of adenosine on prolactin (PRL) secretion from rat anterior pituitaries incubated in vitro. The administration of 5-N methylcarboxamidoadenosine (MECA), an analog agonist that preferentially activates A2 receptors, induced a dose-dependent (1 nM to 1 microM) increase in the levels of PRL released, an effect abolished by 1,3-dipropyl-7-methylxanthine, an antagonist of A2 adenosine receptors. In addition, the basal levels of PRL secretion were decreased by the blockade of cyclooxygenase or lipoxygenase pathways, with indomethacin and nordihydroguaiaretic acid (NDGA), respectively. The stimulatory effects of MECA on PRL secretion persisted even after the addition of indomethacin, but not of NDGA, to the medium. MECA was unable to stimulate PRL secretion in the presence of dopamine, the strongest inhibitor of PRL release that works by inducing a decrease in adenylyl cyclase activity. Furthermore, the addition of adenosine (10 nM) mimicked the effects of MECA on PRL secretion, an effect that persisted regardless of the presence of LiCl (5 mM). The basal secretion of PRL was significatively reduced by LiCl, and restored by the concomitant addition of both LiCl and myo-inositol. These results indicate that PRL secretion is under a multifactorial regulatory mechanism, with the participation of different enzymes, including adenylyl cyclase, inositol-1 phosphatase, cyclooxygenase, and lipoxygenase. However, the increase in PRL secretion observed in the lactotroph in response to A2 adenosine receptor activation probably was mediated by mechanisms involving regulation of adenylyl cyclase, independent of membrane phosphoinositide synthesis or cyclooxygenase activity and partially dependent on lipoxygenase arachidonic acid-derived substances. PMID- 12131929 TI - Dorsal ulnar cutaneous nerve conduction: reference values. AB - We investigated the reference values of the dorsal ulnar cutaneous nerve (DUC) sensory nerve conduction (SNC) in 66 healthy individuals. Measurements were processed using stimulating electrodes positioned between the ulnar bone and the flexor carpi ulnaris muscle, 11-13 cm proximal to the active electrode recording. Superficial recording electrodes were placed on the fourth intermetacarpal space. The mean sensory conduction velocity (SCV) in males was 63.7 - 0.16 x age +/- 3.36 m/s and in females was 57.7 +/- 3.37 m/s. The mean sensory nerve action potential (SNAP) amplitude in males was 19.5 +/- 10.7 microV and in females was 24.6 +/- 5.8 microV. The mean SNAP duration was 0.96 +/- 0.13 ms. No significant differences regarding the DUC-SCV, distal latency, and SNAP duration or amplitude were found between both sides of the same subject. The amplitude of the SNAP was higher in females than males. The effects of age on DUC-SCV were distinct for each gender. PMID- 12131930 TI - Correlation of sleep macrostructure parameters and idiopathic epilepsies. AB - Sleep and epilepsy share some common mechanisms. The objective of the present investigation was to study the macrostructure of sleep in patients with idiopathic epilepsies, focal and generalized, comparing these two groups to each other and to a control group of 12 individuals without epilepsy. A total of 35 polysomnographies were performed, 12 of them in the control group, 10 in patients with idiopathic generalized epilepsies, and 13 in patients with idiopathic focal epilepsies. Antiepileptic medications were maintained for ethical reasons. The group with idiopathic focal epilepsy showed an increase in the total recording time (p = 0.04) and the group with idiopathic generalized epilepsy had a reduction of phase 4 NREM sleep. The efficiency of total sleep period and of total sleep time was also lower in the group with idiopathic generalized epilepsy (p = 0.03 in both cases). We concluded that the group with idiopathic generalized epilepsy presents sleep of poorer quality, whereas the group with idiopathic focal epilepsy presents a tendency toward an excessive somnolence. PMID- 12131931 TI - Carbon dioxide test as an additional clinical measure of treatment response in panic disorder. AB - OBJECTIVE: We aim to determine if a treatment with a dose of clonazepam--2 mg/day, for 6 weeks, blocks spontaneous panic attacks and the ones induced by the inhalation of 35% carbon dioxide (CO2) in panic disorder (PD) patients. The CO2 challenge-test may be a useful addition tool for measuring the pharmacological response during the initial phase (6 weeks) in the treatment of PD. METHOD: Eighteen PD patients drug free for a week participated in a carbon dioxide challenge test. Fourteen had a panic attack and were openly treated for a 6-week period with clonazepam. At the end of the 6-week period they were submitted again to the CO2 challenge test. RESULTS: After 6 weeks of treatment with clonazepam, 12 of 14 PD patients (85.7%) did not have a panic attack after the CO2 challenge test. Just 2 of 14 patients (14.3%) had a panic attack after the CO2 challenge test. Ten of 14 (71.4%) PD patients had panic free status after clonazepam treatment. The 2 patients who had a panic attack in the sixth week, after the CO2 test, did not have panic free status after the treatment with clonazepam. CONCLUSION: The CO2-test may be a valid tool for testing and predicting the drug response. PMID- 12131932 TI - Stereotactic fibrinolysis of spontaneous intracerebral hematoma using infusion of recombinant tissue plasminogen activator. AB - PURPOSE: The authors present a prospective study on 10 patients with stereotactic infusion of tissue plasminogen activator (rtPA) intraparenchimal hemorrhage. METHODS: Between 1999 and 2000, 10 patients with deep seated hematomas in the basal ganglia were selected for stereotactic infusion of rtPA and spontaneous clot drainage. RESULTS: All cases had about 80% reduction of the hematoma volume in the CT scan at the third day. The intracranial pressure was normalized by the third day too. There were no local or systemic complications with the use of this thrombolytic. The results were shown by the Glasgow Outcome Scale with six patients in V, three in IV and one in III after 3 months. CONCLUSION: Early treatment and drainage with minimally invasive neurosurgery, can make these patients with deep-seated hematomas recover the consciousness and they can be rehabilitated earlier avoiding secondary complications. PMID- 12131933 TI - Guillain Barre syndrome in a population less than 15 years old in Brazil. AB - To know the impact of the Guillain Barre syndrome (GBS) in the population less than 15 years old, after the eradication of poliomyelitis. Data bank from the program of epidemiological surveillance of acute flaccid palsies (AFP) from the Fundacao Nacional de Saude were analyzed between 1990-1996. From 3619 notifications of AFP there were 1678 GBS. GBS yearly incidence rates is 0.39-0.63 cases/100,000. No consistent seasonal variation existed or relationship to vaccines. Weakness at inclusion were, moderate 52.1%, severe in 47.9%, sixty days after 57.1% normal, 7.4% mild, 15.7% moderate, 10.4% with severe deficits, death in 5.4%. 67 (4.0%) cases unknown. Death rates varies from 2.8% in southeast to 7.9% in the northeast. GBS was the most frequent cause of AFP. In spite of the severity of this disease being similar in the different regions, the outcome varies according to origin of the cases, possibly reflecting the economical conditions in those places. PMID- 12131934 TI - Comparison of the efficacy of carbamazepine, haloperidol and valproic acid in the treatment of children with Sydenham's chorea: clinical follow-up of 18 patients. AB - In order to compare and contrast the efficacy of haloperidol, carbamazepine, and valproic acid in the treatment of Sydenham's chorea a prospective study including 18 cases of this disorder was undertaken. Age of patients ranged from 7 to 15 years. Ten children were female and 8 were male. All but one had generalized, either symmetric or asymmetric chorea. The patients were divided in three equal groups, and were given a standardized dose of each of the drugs built-up over a week. Following therapy, the six children receiving valproic acid showed remarkable improvement, without side effects. Five patients receiving carbamazepine showed improvement without side effects. Only three of the patients that received haloperidol improved. In the 4 cases that did not show clinical improvement after one week of treatment, therapy with valproic acid led to disappearance of the symptoms in a lapse that ranged from 4 to 7 days. Recurrence related to discontinuation of treatment was observed in two patients. In view of the present results we recommend valproic acid as the first choice drug to treat Sydenham chorea. PMID- 12131936 TI - Clinical, neuroimaging and cytogenetic findings in 20 patients with corpus callosum dysgenesis. AB - Twenty children with corpus callosum agenesis or hypoplasia were evaluated under a standardized investigation protocol. Psychomotor retardation, seizures, and craniofacial anomalies were the most prominent findings. There were three cases of chromosomal anomalies, all of them representing trisomy of chromosome 8. PMID- 12131935 TI - Rasmussen's encephalitis: the relevance of neuropsychological assessment in patient's treatment and follow up. AB - Rasmussen's encephalitis is characterized by refractory epilepsy, neurological deterioration and progressive atrophy of one cerebral hemisphere. The objective of this study is to describe the importance of neuropsychological evaluation in the treatment decision and follow-up of patients with Rasmussens encephalitis. Neuropsychological assessment was performed in two steps. Firstly, the clinical history was obtained and the Vineland adaptative behavior scale (VABS) was applied. After this first step, the patients with social maturity level equal or higher than the inferior limit underwent a battery of neuropsychological assessment. We evaluated three patients before any specific treatment was started, and six months after the intervention (surgery or plasmapheresis). Patient 1 underwent left hemispherectomy and had global improvement on second neuropsychological assessment. This suggests that the decision of performing surgery was adequate. Patients 2 and 3 underwent plasmapheresis. They did not present cognitive decline between both evaluations which suggest that our decision of postponing surgery was adequate as well. We conclude that neuropsychological assessment is important when evaluating patients with Rasmussens encephalitis. That is especially true for patients in whom disease progression is slow, and surgery timing has to be carefully planned. PMID- 12131937 TI - Neuropsychological evaluation of children after ischemic cerebrovascular disease. AB - The purpose of this study is to associate neuropsychological evaluation with neuroimaging results in children with cerebral tomography indicating ischemic cerebrovascular disease (ICVD). Neuroimaging, neurological exams and neuropsychological instruments were used to evaluate five children. The study revealed that the cognitive and perceptive skills in two children were normal and motor sequele in four cases. The rhythm, visual and speech receptive skills remained unchanged. In four cases the SPECT exam showed regions with hypoperfusion and in four cases the EEG was normal. Neuropsychological, neurological and image indication some degree of sequele demonstrating the importance of follow up of children who had suffered cerebrovascular disease. PMID- 12131938 TI - Benign focal epilepsy of childhood with centrotemporal spikes (BECTS): clinical characteristics of seizures according to age at first seizure. AB - BECTS is characterized by the presence of simple partial motor seizures in the face and/or oropharynx, with or without sensory symptoms and often with secondary generalization. These seizures tend to occur more often during sleep or drowsiness. According to some authors, generalized seizures prevail over other types particularly among children aged five or less. The purpose of this study is to determine the characteristics of the first epileptic episode among children with BECTS, grouped by age as of their first epileptic seizure, as well as to analyze how such seizures change over the course of clinical evolution. A total of 61 children were examined, 16 of whom below the age of 5 and 45 above. With regard to the first and recurrent epileptic episodes, our final assessment showed that partial seizures occurred more frequently than generalized tonic-clonic seizures in both groups. Although no conclusive relation could be established between the type of partial seizure (i.e. simple versus complex) and the children's age as of their first epileptic episode, it was possible to correlate the type of epileptic seizure with their clinical evolution, in which case simple partial seizures proved to be more frequent than complex partial seizures. It should be noted that the number of children under the age of five was relatively small, which evidences the need for further studies. It should also be borne in mind that the reported frequency of generalized seizures in these children's first epileptic episode may be due to their parents' lack of attention and familiarity with this pathology and their attendant difficulty in characterizing its clinical symptoms. PMID- 12131940 TI - Enzyme linked immunosorbent assay (ELISA) for the detection of IgG, IgM, IgE and IgA against Cysticercus cellulosae in cerebrospinal fluid of patients with neurocysticercosis. AB - The objective of this study was to analyze different immunoglobulins classes (IgG, IgM, IgE and IgA) against Cysticercus cellulosae in the cerebrospinal fluid (CSF), through enzyme linked immunosorbent assay (ELISA), correlating them to clinical and tomographic profiles in patients with neurocysticercosis (NCC). Eighty-five specimens of CSF were obtained from 43 cases with NCC (26 with the active form and 17 with the inactive form) and from 42 patients with other neurological diseases. The inactive form of NCC presented a profile in CSF similar to the group without NCC. The active form of NCC presented elevation of specific immunoglobulins (IgG, IgM, IgE, and IgA) in decreasing order, with the highest values being detected among the cases with intraventricular cysts, or with inflammation signs in CSF or in those with multiple clinical manifestations. The highest sensitivity and specificity were obtained with ELISA-IgG (88.5% and 93.2%, respectively). This study confirmed the importance of ELISA in the immunologic diagnosis of NCC. PMID- 12131939 TI - IgG intrathecal synthesis and specific antibody index in patients with neurocysticercosis. AB - We analyzed cerebrospinal fluid (CSF) and blood serum from 55 patients with neurocysticercosis (NC) at different clinical stages. According to inflammatory activity in the CSF, three stages were identified: (1) reactive, when there was at least an increase in the number of cells; (2) weakly reactive, when significant alterations were found in the CSF, including an increase in gamma globulins, albeit without hypercytosis; (3) non-reactive, when there was neither hypercytosis nor increase in gamma globulins. Nineteen patients had the reactive form; 18 had the weakly reactive form; 18 displayed the non-reactive form. Local immunoproduction was intense in the reactive group, moderate in the weakly reactive group, and absent in the non-reactive group. The specific antibody index was raised in approximately 2/3 of patients with the reactive form, 2/3 in those with the weakly reactive form, and 1/3 in those with the non-reactive form. IN CONCLUSION: (1) the classical CSF syndrome in NC can present both in complete and partial modes; (2) local immunoproduction can occur in weakly reactive forms; (3) a raised specific antibody index can occur in the absence of an inflammatory reaction in the CSF. PMID- 12131941 TI - [Efficacy of three drugs in the treatment of migrainous aura: a randomized placebo-controlled study]. AB - In spite of the enormous amount of research regarding classification, epidemiology, diagnosis, pathophysiology and treatment of migraines, aura, one of its hallmarks, stays much less studied. The objective of this study is, therefore, to evaluate the evolution of aura in patients submitted to placebo and three different drugs available in Brazilian public health units. We studied 86 patients during an acute migrainous attack, with aura at the moment of the evaluation. Patients were randomized to receive one of the following substances, in a parenteral route: placebo, dipyrone, chlorpromazine and magnesium sulphate. Magnesium sulphate was superior to placebo (p <0,05) 30 and 60 minutes after its administration. Dipyrone and chlorpromazine reduced the duration of the aura, when compared to placebo, 60 minutes following their administration. Our findings make possible some speculations about the pathophysiology of migraine, as well as about the therapeutic approach of patients presenting migrainous aura. PMID- 12131942 TI - [Intramuscular diclofenac in the acute treatment of migraine: a double-blind placebo controlled study]. AB - The aim of this study is to assess, in a double blind fashion, the effect of diclofenac on the pain and associated symptoms in patients with acute migraine. 60 patients with migraine with aura and 60 patients with migraine without aura were assigned at random to receiving intramuscular diclofenac, 75 mg associated to intravenous physiological saline, 10 ml, or physiological saline alone. We used 3 parameters of analgesic evaluation and an analogical scale to asses associated symptoms. We also observed the recurrence and rescue medication percentiles. Patients receiving diclofenac showed a statistically significant improvement of pain 60 minutes after the administration in two of the three parameters(migraine without aura) and in all parameters (migraine with aura). They did not present difference, compared to placebo, on the intensity of associated symptoms. We observed a reduction on the recurrence and rescue medication utilization. Despite being an option to treat migraines attacks in the emergency room settings, diclofenac has a slow onset action and no effects on the associated symptoms. PMID- 12131943 TI - [Tomographic findings in 1000 consecutive patients with antecedents of epileptic seizures]. AB - We analyzed the cranial computerized tomography scans (CT) of 1000 consecutive patients with the history of seizures. The seizures were classified in generalized tonic-clonic seizures (GS) and partial seizures (PS) on the basis of the clinical semiology, as it was described by the patients and family members. Seizure types were correlated with age group, sex and CT findings. GS were observed in 70.7% of the patients and PS in 29.3 %. An increased incidence of GS was noted among the age groups 0 to 10 yrs (31%) and 11 to 20 yrs (21.8%). For PS the 0 to 10 yrs (24.5 %), 21 to 30 yrs (16.7%) and 31 tp 40 yrs (18.5%) age groups were the most prevalent groups. The CT scan findings for the GS group were the following: normal studies 48.8 % of the patients, calcifications/cysticercosis (14.0%), neurocysticercosis/cysts (9.6 %), hydrocephalus (4.3%), ischemia (4.2 %), non-definite lesions (4.1 %), tumor (2.5%), and others (12.5%). In the PS group we found 37.4% of normal studies, calcifications/cysticercosis (11.2%), neurocysticercosis/cysts (12.2%), tumor (10.5%), ischemia (5.4%), hydrocephalus (3.7%), non-definite lesions (8.1%) and others (11.6%). We emphasize the importance of CT scan imaging in patients presenting with seizures, particularly in the diagnosis of cysticercosis. PMID- 12131944 TI - [Reliability of volumetric measures of mesial temporal structures]. AB - RATIONALE: The development of reliable techniques for volumetric measurement of mesial temporal structures (amygdala, hippocampus and parahippocampal gyrus) on magnetic resonance imaging (MRI) can provide data for the study of neuropsychiatric disorders, mainly temporal lobe epilepsy, Alzheimer's disease and schizophrenia. METHOD: We investigated these techniques performing intraobserver and interobserver reliability study concerning normal controls, epilepsy and Alzheimer's disease patients using the intra-class correlation coefficient. RESULTS: Intra-observer reliability of evaluated structures ranged from 0.93 to 0.99 (p<0.001). Inter-observer reliability ranged from 0.70 to 0.95 (p < or = 0.001). CONCLUSION: The results suggest that the technique of MRI morphometry of mesial temporal regions can be considered a reliable tool which may help in the investigation of neuropsychiatric disorders, since used by adequately trained clinicians and researchers. PMID- 12131945 TI - [Deep brain stimulation of thalamus for tremor control]. AB - PURPOSE: We present our results in 4 patients with tremor, in whom electrodes (uni and bilateral) for Deep Brain Stimulation (DBS) were implanted in the ventral intermediate nucleus (VIM) of the thalamus. METHOD: Four patients with disabling tremor, with drug-resistant spite of optimum therapeutic trials with poor response were referred to do surgery. Two patients had bilateral essential tremor. These patients were implanted with electrodes for DBS 3387 (Medtronic). Two patients had unilateral parkinsonian tremor and they received unilateral implantation of model 3387 DBS. RESULTS: All four patients showed relieve of the tremor symptoms with significant tremor control seen at the scores. There were no definite adverse events after the electrodes implants for DBS; adverse events were transient and promptly reversed after the adjustment of the parameters. CONCLUSIONS: The results the authors found in this study indicate that VIM stimulation is effective for tremor control either parkinsonian or essential tremor. The results correlate with the data in the literature. PMID- 12131946 TI - [Daily living activities evaluation in Parkinson's disease patients underwent to stereotactic surgery]. AB - OBJECTIVE: To evaluate the impact of stereotactic surgery on daily activities of Parkinson's disease (PD) patients. METHOD: Thirty patients with idiopathic PD were evaluated before surgery and one, three, six and twelve months after surgery. Patients were evaluated with the UPDRS - part II (Unified Parkinson's Disease Rating Scale) and the Schwab & England scales. Nine of the patients had also been evaluated after twenty four months. RESULTS: We performed nine posteroventral pallidotomies (PVP), two on the left hemisphere and seven on the right; 17 ventrolateral thalamotomies (VLT), 12 on the left and five on the right; two VLT on the left associated with PVP on the right at the same surgical procedure. The mean "off" phase scores of Schwab & England scale were: 60.6 before surgery, 74 after the first, 76.6 after the third, 75.6 after the sixth, 72.3 after the twelfth and 71.1 after the twenty fourth months after surgery. The mean "off" phase scores of UPDRS - part II scale were: 21 before surgery, 12.3 after the first, 14.7 after the third, 15.27 after the sixth, 17.1 after the twelfth and 17.5 after the twenty fourth months after surgery. CONCLUSION: VTL and PVP are useful procedures to improve daily living activities of the PD patients and the best results are seen by six months after surgery. PMID- 12131948 TI - [Comparison of functional activity performance in normally developing children and children with cerebral palsy]. AB - OBJECTIVE: To compare the pattern of self-care performance in normal children and children with cerebral palsy (CP). METHOD: 142 normal children and 33 children with CP were evaluated by 22 items from the self-care scale of the PEDI functional test. Rasch methodology transformed scores into interval measures of difficulty from 0 to 100 (logit). Spearman rank correlation coefficient compared the order of logits in the two groups. RESULTS: Eleven items showed significant differences in the logit values received. Out of these, 7 items showed relative difficulty values greater in the group of children with CP and 4 items showed relative difficulty values greater among normal children. A significant correlation was observed in the order of the 22 items displayed in the two interval scales. CONCLUSION: The results suggest that the development of self care functional activities may be influenced by the presence of CP. These results may support assessment and intervention strategies for children with neuromotor disorders. PMID- 12131947 TI - [Impact of epilepsy at work: evaluation of quality of life]. AB - This study searched for the knowledge of the most affected aspects in the quality of life (QoL) of patients with epilepsy. The sample consisted of 134 individuals with epilepsy, aging 18 to 59 years (M=35.38; DP=9.86) chosen at random from the Ambulatory of Epilepsy of HC/UNICAMP, interviewed by the QoL-65. Most of the subjects have not completed elementary school (58.2%), have not had any paid work (69.5%) and were single (48.5%). The most affected factor in epilepsy was the work area (31.29%), what reinforces studies showing the high level of unemployment or subemployment in epileptic population and aware to the importance of the insertion in the work-market as an economic and social integration factor and as a way of improving self-esteem. PMID- 12131949 TI - Long-term evolution of papilledema in idiopathic intracranial hypertension: observations concerning two cases. AB - Chronic headaches, associated with papilledema and pulsatile tinnitus without any neuroradiologic, cytobiochemical or cerebrospinal fluid abnormalities are suggestive of idiopathic intracranial hypertension (IIH). However the absence of the papilledema does not rule out this diagnosis. The reason why some patients do not develop papilledema in IIH is ignored, however there are some hypotheses concerning the structure of the optical nerve. In this study we described two female patients that presented diagnosis of IIH with papilledema, with subsequent resolution of papilledema without the due resolution of intracranial hypertension. The long-term behavior of the optic nerve (ON) facing an increased intracranial pressure was evaluated through repeated measurements of the intracranial pressure. We concluded that the ON submitted to high intracranial pressure for a certain length of time can adapt itself with subsequent disappearance of the papilledema. The presence or not of papilledema in IIH can be related to the period in which the diagnosis is accomplished. PMID- 12131950 TI - Iatrogenic Creutzfeldt-Jakob disease following human growth hormone therapy: case report. AB - We report the case of a 41-year-old man with iatrogenic Creutzfeldt-Jakob disease (CJD) acquired after the use of growth hormone (GH) obtained from a number of pituitary glands sourced from autopsy material. The incubation period of the disease (from the midpoint of treatment to the onset of clinical symptoms) was rather long (28 years). Besides the remarkable cerebellar and mental signs, the patient exhibited sleep disturbance (excessive somnolence) from the onset of the symptoms, with striking alteration of the sleep architecture documented by polysomnography. 14-3-3 protein was detected in the CSF, and MRI revealed increased signal intensity bilaterally in the striatum, being most evident in diffusion-weighted (DW-MRI) sequences. This is the second case of iatrogenic CJD associated with the use of GH reported in Brazil. PMID- 12131951 TI - Frontoparietal cortical atrophy with gliosis in the gray matter of cerebral cortex: case report. AB - The case of a patient who suffered from progressive amnesia, depressive humor, language and visuospatial disturbances, and hallucination episodies with interference at the daily living activities is reported. She had moderate neuropsychological diffuse deficits at the first examination, especially at the executive and visuo-constructive functions. Her cerebrospinal fluid test presented high total protein. Magnetic resonance image showed slight white matter increase in periventricular, semi-oval center bilateral and left external capsule regions, besides light frontal and parietal lobe atrophy, bilaterally. Brain single photon emission computerized tomography revealed both a bilateral moderate frontal and a severe parietal lobe hypoperfusion, especially on the left side. Macroscopic examination showed cortical atrophy, severe on the frontal, moderate on the parietal and mild on the posterior third temporal lobes, bilaterally. There was a slight atrophy on the neostriatum in the basal ganglia. The histopathological findings of the autopsy showed severe neuronal loss with intensive gemioscytic gliosis and variable degrees of status spongiosus in cortical layer. Hematoxylin-eosin and Bielschowsky staining did not show neuronal swelling (balooned cell), argyrophilic inclusion (Pick's bodies), neurofibrillary tangles nor senile plaques. Immunohistochemical staining for anti-ubiquitin, anti tau, anti-beta-amyloide, and anti-prion protein were tested negative. PMID- 12131952 TI - Glioma and multiple sclerosis: case report. AB - We report a case of a 44-years-old woman with relapsing-remitting and secondarily progressive form of multiple sclerosis (MS) since aged 24 years, who developed an anaplastic astrocytoma. The neurological manifestations of the tumor were misinterpreted as resulting from MS. Sequential MRI examination and seizures raised the possibility of another nature of her symptoms, besides MS. Her initial good response to high doses corticosteroids led to the initial assumption her symptoms were only exclusively due to the demyelinating process. She underwent craniotomy with radical excision of the lesion. Pathological examination disclosed anaplastic astrocytoma. Other cases of coincidental MS and primary CNS tumors are reviewed, as well as their possible relation. PMID- 12131953 TI - Spinal cord tumor in a patient with multiple sclerosis: case report. AB - The association between multiple (MS) sclerosis and cerebral gliomas has been sporadically reported in the literature, causing a long lasting discussion if these lesions occur coincidentally or if MS plaques may actually lead to the genesis of gliomas. We report a 36 year old man who developed a rapid onset of right side weakness and loss of vision, having established a diagnosis of MS which was confirmed by CSF analysis and MRI. Nine years later he developed progressive tetraparesis, leading initially to suspicion of illness relapse and a demyelinating plaque in the spinal cord. However, after MRI investigation, a spinal cord tumor was diagnosed. The patient underwent cervical spine laminotomy for microsurgical removal of the spinal cord tumor diagnosed as ependymoma. The neurological deficits improved significantly. PMID- 12131954 TI - Progression of an arterial infundibulum to aneurysm: case report. AB - In this case an aneurysm of the right posterior communicating artery developed 11 months after an infundibular dilation of this artery had been angiographycally and surgically demonstrated. In the best of the authors' knowledge, there are only eleven such cases reported in the literature. This report brings about diagnostic and therapeutic questions regarding arterial infundibula and the need of a better understanding of those lesions. PMID- 12131955 TI - [Giant cavernous angioma: report of two cases]. AB - Cavernous angiomas or haemangiomas or yet cavernomas are malformations of the central nervous system classified as occult vascular brain lesions. These rare lesions are clinically silent. They are defined by the presence of abnormally large vascular cavities or sinusoids channels of variable size, with sharp walls, located inside but not invading the brain parenchyma. They can occur at any age, including the neonatal period. Most of the small lesions are located inside the brain parenchyma. No abnormal circulation can be demonstrated in angiography and CT scan can be helpful for diagnosis only in rare occasions. Magnetic resonance is the best exam to demonstrate the lesion. Despite the benign character some lesions may cause neurologic dysfunction when their removal may be difficult. Complete extirpation is the best treatment if the lesion is favorable located and is causing neurological dysfunction. Two cases of giant cavernomas with good outcome after total removal are present. Only three cases of giant cavernomas were reported in the literature. PMID- 12131956 TI - [Central pain due to parietal cortex compression by cerebral tumor: report of 2 cases]. AB - Central pain derived from cerebral tumors is considered rare by most authors. The few cases described in literature refer to central pain caused by expansive lesions in the contralateral parietal cortex. Usually, not even thalamic tumors cause central pain. We describe two cases of central pain related to expansive lesions in the parietal cortical region - a low grade astrocytoma and a meningioma. Both patients reported pain relief after lesions were removed by surgery. PMID- 12131957 TI - [Hypokalemic periodic paralysis as first manifestation of hyperthyroidism: case report]. AB - We report a case of thyrotoxicosis in a 37 year old male, in whom hypokalemic periodic paralysis was the first manifestation of the disease. We comment about the differential diagnosis with other types of periodic paralysis, and the importance of a correct diagnosis to avoid permanent muscle damage. PMID- 12131958 TI - [Focal epileptic seizures ipsilateral to the tumor: case report]. AB - Focal somatosensory epileptic seizures ipsilateral to a brain tumor is reported and the literature reviewed. It is an exceptional occurrence, having been described only six cases, with several mechanisms being proposed. The proximity of the lesions with the low cerebral convexity (perisylvian) suggests the compromising of the secondary somatosensorial area, seeming to prove the experimental observation of somatosensorial crises originating in this area. PMID- 12131959 TI - [Pharmacologically-induced hypertension in a patient with vertebro-basilar territory ischemia associated with bilateral vertebral stenosis]. AB - Hypertension is one of the main risk factors for stroke. However, treating hypertension in the acute phase may cause further neurological deterioration. We present a case of an 81-year-old woman, admitted after multiple infarcts in the posterior circulation. While fully anticoagulated, her neurological deficits worsened, coinciding with normalization of her blood pressure levels. Magnetic resonance angiography documented bilateral vertebral artery stenoses. Induced hypertension was followed by rapid clinical improvement. In this first report of induced hypertension in the Brazilian literature, we illustrate the potential benefit of this therapeutic strategy in patients with documented hemodynamic mechanism of clinical deterioration. PMID- 12131960 TI - [Endovascular treatment of spontaneous vertebral arteriovenous fistula in children: case report]. AB - Spontaneous cervical artery fistulas are rare arteriovenous malformations between the artery and veins of the neighborhood. We report a case of non traumatic vertebral arteriovenous fistula in a girl, aged 9 years, treated by endovascular approach. Under general anesthesia and with fluoroscopic guidance, using a endovascular technique, latex detachable balloons were used to successfully occlude the fistula. Complete clinical and angiographic recovery was achieved and no complications related to the embolization procedure occurred. The result obtained in this case corroborates that endovascular techniques, especially balloon occlusion embolization, has become the standard treatment for this vascular disorder. PMID- 12131961 TI - The uncinated crisis of George Gershwin. AB - George Gershwin, renowned composer and pianist, well known for his popular works, died on the 11th July 1937 due to a brain tumor. His neurological symptoms first appeared on that same year, in February, with a simple olfactory partial seizure, characterized by an unpleasant smell of burnt rubber (uncinated seizure). He later had a quick clinical descend, with severe headache that occurred in bouts, dizziness, coordination compromise and olfactory seizures, eventually lapsing into a coma on the 9th July 1937. It was then that a gliomatosus cyst was diagnosed, which on microscopic examination proved to be a "glioblastoma multiforme". Despite the surgical intervention, Gershwin died soon after the procedure without recovering his consciousness. We make a brief review of Gershwin's neurologic disease, with emphasis on the initial symptoms, namely the uncinated seizures. PMID- 12131962 TI - [Evolutionary epidemiological thought on infections]. AB - The objective of the study is to analyse the main aspects of current epidemiological knowledge on the evolutionary status of infections. Living organisms in the biosphere are part of dynamic systems of variable intensities. Some of these systems are on the surface while others take place inside the genome core. Parasitism is a phenomenon commonly seen in nature. Infective parasites relate to each other through several mechanisms, such as genetic DNA exchange, and because of the connections established communities of infectious agents are not isolated. The internalization process allows the parasites to get into their hosts' cells, which is accomplished through the phagocytosis of infectious agents or other more sophisticated mechanisms such as pill production. To leave the intracellular medium, some organisms make use of apoptosis, a highly specialized genetic mechanism that makes possible to destroy macrophages. It is currently accepted that molecular DNA can flow into the blood stream as the so called infectrons. Thus it is hypothesized the existence of infection networks that allows the coadaptability of parasites and their hosts, and creates coevolutionary forces between hosts and their parasites facilitating the emergence of new pathogens. PMID- 12131963 TI - [Characteristics of anticontraception methods used in Sao Paulo State, Brazil (correction)]. AB - OBJECTIVE: To analyze data on contraceptive use in the State of Sao Paulo, Brazil, collected by the Demographic and Health Survey (DHS) conducted in 1996. METHODS: The study data were compared to 1986 DHS and 1996 data on the Brazilian population. Contraceptive use among married or cohabiting women was evaluated focusing on age, number of children, schooling, and age and timing of female sterilization. Statistical analysis was performed using Student t-test and Kendall's non-parametric test. RESULTS: Unlike data on the Brazilian population, female sterilization rates were steady in the State of Sao Paulo during the studied period. The same contraceptive pattern is seen in both Brazil and Sao Paulo: women aged up to 30 years use largely pills; female sterilization predominates in women over 30, increasing with the number of children and decreasing with years of schooling. Male methods have also increased in recent years, being greater in Sao Paulo than in Brazil. Sao Paulo also shows a greater variety of reversible contraceptive methods. CONCLUSIONS: Although there were some differences, the prevalence of only two contraceptive methods in both Brazil and Sao Paulo suggests a tendency regarding the contraceptive methods offered and in reproductive health in the view of new regulations on family planning. PMID- 12131964 TI - [Male perspective on induced abortion]. AB - OBJECTIVE: To analyze the perspective on induced abortion of men of a university community living in legal or consensual wedlock. METHODS: A descriptive cross sectional study was carried out and 361 members of different categories of an university were interviewed. Chi-square test was used to assess the association between dependent and independent variables. RESULTS: Fifth-three percent of the participants acknowledged that women have the right to end pregnancy. Men were more favorable to abortion when there is a risk to woman's life (85%); rape related pregnancy (80%); and fetal anomalies (75%). Higher schooling of both men and their partners and the interviewees' position (teacher/student) were associated to a positive attitude towards abortion. CONCLUSIONS: Men tended to be more prone to abortion in legally and/or socially accepted instances. Better education of both men and their partners was relevant to determine their attitude towards abortion. PMID- 12131965 TI - Survival of adult AIDS patients in a reference hospital of a metropolitan area in Brazil. AB - OBJECTIVE: To evaluate the influence of sociodemographic, clinical, and epidemiological factors in AIDS patients survival in a reference hospital. METHODS: A sample of 502 adult AIDS patients out of 1,494 AIDS cases registered in a hospital in Fortaleza, Brazil, was investigated between 1986 and 1998. Sixteen cases were excluded due to death at the moment of the AIDS diagnosis and 486 were analyzed in the study. Socioeconomic and clinical epidemiological were the variables studied. Statistical analysis was conducted using the Kaplan-Meier survival analysis and the Cox proportional hazards model. RESULTS: Three hundred and sixty two out of the 486 patients studied took at least one antiretroviral drug and their survival was ten times longer than those who did not take any drug (746 and 79 days, respectively, p <0.001). Patients who took two nucleoside reverse transcriptase inhibitors (NRTI) plus protease inhibitor were found to have higher survival rates (p <0.001). The risk of dying in the first year was significantly lower for patients who took NRTI and a protease inhibitor compared to those who took only NRTI. In addition, this risk was much lower from the second year on (0.10; 95%CI: 0.42-0.23). The risk of dying in the first year was significantly higher for less educated patients (15.58; 95%CI: 6.64-36.58) and those who had two or more systemic diseases (3.03; 95%CI: 1.74-5.25). After the first year post-diagnosis, there was no risk difference for these factors. CONCLUSIONS: Higher education revealed to exert a significant influence in the first-year survival. Antiretroviral drugs had a greater impact in the survival from the second year on. A more aggressive antiretroviral therapy started earlier could benefit those patients. PMID- 12131966 TI - [Factors associated to hospitalization of children under five years of age, Sao Paulo, Brazil]. AB - OBJECTIVE: In developing countries acute respiratory infection (ARI) is the leading cause of hospitalization among children under five years of age. Their underprivileged life conditions and restricted access to health care services are important determining factors. The objective of the study was to assess hospital morbidity and to identify factors associated to hospitalization of children under five years of age. METHODS: A data set derived from a cross-sectional study on health conditions of children under five years of age in the city of Embu, a county located in the metropolitan region of Sao Paulo, Brazil, was used. The inclusion criteria were one child per family (random selection). The exclusion criteria were missing data on any study variable. The sample size was 893 children. Data was collected using household interviews with mother or caretaker. Statistical analysis was performed using logistic regression models to identify factors associated with hospitalization. RESULTS/CONCLUSIONS: Sixty-five (7.7%) children were hospitalized. Of them, 41.5% were admitted with a respiratory tract disease, mainly due to an ARI (27.7 %). Factors associated to hospitalization included: low birth weight; perinatal problems; chronic illness; death of a sibling under the age of five; grandmother as day caretaker; living in overcrowded places, and mother's higher educational level. PMID- 12131967 TI - [A strategy for the management of hospitalized children with acute lower respiratory infections]. AB - OBJECTIVES: To improve the quality of care provided to hospitalized children having acute lower respiratory infections (ALRI), to increase the knowledge on this health condition, and to broaden the utilization of health care resources through a program called "Winter Plan". METHODS: The program comprised the use of guidelines for diagnosis and treatment, disease-oriented hospitalizations to provide an increased level of care, management of health care resources and implementation of computerized medical records. Systematic investigation of viral etiology was performed in order to rationalize the use of medications and reduce nosocomial infections. RESULTS: During program implementation (19/V-19/IX/99), 3,317 children were admitted; 1,347 (40.61%) had ALRI, of which 1,096 (81%) were included in the study. Of them, 71% aged less than 1 year. Most ALRI were viral (68%). Admission criteria were: oxygen saturation <95%, tachypnea, retractions or pleural effusion (92.4% of the children). The demand magnitude prevented compliance with isolation guidelines in all cases. Treatment guidelines were followed in a high percentage of cases: 73% of children having bronchiolitis and 72% of those with viral pneumonia received no antibiotics and 96% of children with bacterial pneumonia were put on antibiotics as recommended; use of bronchodilators and corticosteroids was reduced. Medication costs were reduced especially in the corticosteroid group, which meant a greater impact on hospitalization costs. CONCLUSIONS: To decrease ALRI morbidity and mortality there is a need to continue improving the quality of health care during hospitalization and to reinforce health promotion actions and preventive programs at the primary level. PMID- 12131968 TI - [Validity of death certificate variables in cases of death from external causes, Brazil]. AB - OBJECTIVE: To validate variables other than the basic cause of death on death certificates from external causes of residents younger than 20 years old living in Recife, Brazil, in 1995. METHODS: A survey of death records of the Institute of Forensic in Recife was carried out in order to validate death certificate variables included in official mortality registries. Agreement analysis was performed using Kappa indexes and sensitivity. As a result of the methodology used, the study data was considered to be more consistent and therefore regarded as standard. RESULTS: Those variables forensic specialists indicated as their direct responsibility showed an excellent (gender, age, and category of violence) and good (type of accident) agreement. However, there were significant discrepancies in those variables seen as either of indirect or no forensic responsibility, such as locality and area where death took place, medical care provided and accident location. CONCLUSIONS: Data reveal a dissociation between the reasons for including some variables in death certificates and the obtaining strategy and their social purpose. PMID- 12131969 TI - [Validity of the Nordic Musculoskeletal Questionnaire as morbidity measurement tool]. AB - OBJECTIVES: To validate the Portuguese version of the Nordic Musculoskeletal Questionnaire (NMQ), and to evaluate the relationship between musculoskeletal morbidity and demographic, occupational and behavior variables. METHODS: Ninety government bank employees were interviewed using the Portuguese version of NMQ in Brasilia, Brazil, in 1999. The respondents' answers were compared to clinical data found in their medical records. Descriptive and associative variable analyses were carried out. Comparative statistical analysis was performed using Student t-test and Pearson's test was applied for correlational analysis. RESULTS: The results revealed an 86% agreement rate between symptoms reported in the NMQ and the respondent's clinical history. There were found differences in symptoms prevalence for the variables gender, occupation and physical activity. Women showed a higher severity average of symptoms for almost all body anatomical regions; managers reported greater severity of symptoms in the lumbar area than clerks; physical activity was associated to the lowest severity of symptoms in upper extremities. CONCLUSIONS: The results show that the Portuguese NMQ version includes a good validity indicator and is satisfactory as a musculoskeletal morbidity measurement tool. However, there is a need of (1) more items to measure the severity of symptoms; and (2) changes in scale layout in order to make it more easily understandable and less prone to avoidable missing data. PMID- 12131970 TI - [Frequency and determinants of breastfeeding in the State of Sao Paulo, Brazil]. AB - OBJECTIVE: To meet the information need of Brazilian municipalities concerning breastfeeding practices as part of health care planning, a study was carried out to describe breastfeeding and to identify weaning-related factors. METHODS: Of all municipalities in the State of Sao Paulo, Brazil, openly called to participate in the study, 84 joined in. Personnel underwent training to collect data during the national mass immunization day in 1998. A sample strategy, proportional to the infant population, was developed for each participant. Standardized questionnaires were applied to assess infant feeding practices in the previous 24 hours. Descriptive statistic analysis on breastfeeding prevalence and logistic regression analysis of risk factors for discontinuing exclusive breastfeeding of infants aged less than 4 months and weaning of infants aged less than a year were performed. RESULTS: Exclusive breastfeeding rates in the first four months of life were under 30%. The risk factors were: lower maternal education status; lack of access to the so-called "Baby Friendly Hospital Initiative"; primiparity; and early age pregnancy. Around 50% of children under a year old were breastfed. CONCLUSIONS: Lacks of access to the "Baby Friendly Hospital Initiative", primiparity, and mother's unemployment or engagement in an informal occupation activity were risk factors for weaning. Local breastfeeding rates are extremely variable in the State of Sao Paulo, reinforcing the importance of local, swift, and easily feasible health care actions. PMID- 12131971 TI - [Aflatoxins and ochratoxin A in food and risks to human health]. AB - OBJECTIVES: The presence of mycotoxins in food has been associated with several human diseases, and health authorities have taken actions to decrease the ingestion of these compounds in the diet. A study was carried out to assess aflatoxins and ochratoxin A concentrations found in food, and to evaluate the potential risk to human health resulting from mycotoxin exposure. METHODS: Between July 1998 to December 2001, 366 food samples were analyzed, including peanuts and its products, nuts, maize, oat and/or wheat products, rice and beans. Samples were processed and the extracted mycotoxins were detected and separated using thin layer chromatography, and then quantified with fluorescence. RESULTS: Aflatoxins were detected in 19.6% of the samples: raw peanuts and its products, pop corn, maize and Brazilian nuts (>2mg/kg). Peanuts and its products showed the highest levels of aflatoxin contamination (34.7%) with up to 1280 mg/kg of AFB1+AFG1 and 1706 mg/kg of total aflatoxins. Of the positive samples, AFB1 was detected in 98.5%, AFB2 in 93%, AFG1 in 66.7%, and AFG2 in 65.4%. Ochratoxin A was not detected (<25 mg/kg) in any sample analyzed. CONCLUSION: It was found that contamination levels mainly seen in peanuts and its products exceed Brazilian regulated standards, and they can be a potential risk to regular consumers of these products. Food producers' awareness allied to monitoring programs is essential to reduce human exposure to these compounds and prevent ensuing chronic diseases. PMID- 12131972 TI - [Health sector decentralization and divergences with the medical society in Mexico]. AB - OBJECTIVE: To evaluate the medical society's perception and actions in the context of health sector decentralization in the states of Guanajuato and Sonora, Mexico. METHODS: Qualitative research techniques were applied. Thirty-five semi structured interviews were conducted with medical college members of both public and private institutions, and collegiate and union representatives of both states studied. RESULTS: Members of medical society in both states acknowledged that decentralization implied in insecurity due to the lack of clarity of health sector regulations. As a result of actions of the medical college in both states there was a growing politicization of medical college members, elaboration of proposals to increase their control over the medical labor market and their participation in the regional political power structure. CONCLUSIONS: This research supports the existing re-articulation of the medical society at a regional level preserving its status as a respected group. In contrast to studies conducted in the United States and Mexico medical authorities have pressed on the regulations in order to preserve their privileged status within the existing contention. PMID- 12131973 TI - [Chromosome abnormalities caused by computer video display monitors' radiation]. AB - OBJECTIVE: Concerns were raised about the potential damaging effects of electromagnetic field (EMF) radiation emissions of ELF (extremely low frequency) and VLF (very low frequency) computer video display monitors (VDM), it was assessed the frequency of structural chromosome abnormalities and investigated the cell cycle kinetics in individuals occupationally exposed to VDM radiation. METHODS: Chromosome aberrations were investigated in 2,000 first cell cycle metaphases obtained after 48-hour cultures of peripheral blood lymphocytes drawn from 10 individuals occupationally exposed to VDM radiation (group E) and 10 controls (group C). Cell cycle kinetics was studied using the mitotic index (MI) and cellular proliferation index (CPI). RESULTS: Statistical analysis showed significantly higher frequencies of anomalous metaphases (E=5.9%; C=3.7%) and anomaly/cell (E=0.066+/-0.026; C= 0.040+/-0.026) in individuals exposed to VDM radiation. The most common cytogenetic alterations seen were chromatid breaks at frequencies of 0.034+/-0.016 in group E and 0.016+/-0.015 in group C. There was no significant difference between MI and CPI frequencies in both groups. CONCLUSIONS: The study findings suggest genotoxic effects of EMF emissions revealed by the higher frequency of chromatid breaks in individuals exposed to VDM radiation. However, there is a need of further studies on EMF genetic effects using other investigation methods. PMID- 12131974 TI - [Morbidity leading to grants due to temporary work disability]. AB - INTRODUCTION: To identify health conditions leading to benefits due to temporary work disability in a population of insured workers. METHODS: International Classification of Diseases (ICD) codes for conditions resulting in temporary work disability (E-31) were retrieved from the National Institute of Social Security (INSS) data bank in Porto Alegre, Brazil, in 1998. The ICD codes related to the worker's disability assigned in the early medical expert examination (aX1) were used to describe the main disability causes and groups of conditions. RESULTS: A total of 6,898 disability benefits were allowed to insured workers: 1,486 (22%) were attributed to "external causes"; 1,181 (17%) to "surgery recovery" (subdivided as follows: 34% gastrointestinal; 26% genitourinary; 11% musculoskeletal; and 10% external causes ); and 4,119 (61%) to "medical conditions" (subdivided as follows: 24.8% musculoskeletal diseases; 18.9% mental diseases; and 16.2% cardiovascular diseases). When compared to a similar Brazilian study conducted in 1986, external causes moved up from fourth to the first position as a determinant of temporary work disability. CONCLUSIONS: The main causes of disability identified, accidents and violence, musculoskeletal diseases and mental diseases, are potentially associated to the worsening of the quality of life and work conditions during the study and should be a (preventive and therapeutic) priority in the nation's unified health care (SUS) agenda. The study shows the viability of referring to the INSS data bank in morbidity studies. PMID- 12131975 TI - Popular beliefs regarding the treatment of senile cataract. AB - OBJECTIVE: To identify popular beliefs regarding the treatment of senile cataract in patients enrolled in the community health programs on eye rehabilitation. METHODS: A cross-sectional survey was carried out using an interview questionnaire that was applied to 776 subjects drawn from a non-probabilistic sample in five cities of the state of Sao Paulo. The sample was made up of 47.2% males and 52.8% females, aged 50 to 96 years (average age 71.6 years). RESULTS: Of the total of subjects studied, 41.9% had never attended school, and 78.5% were no longer in the employment market. Most (85.1%) credited the sight restoration to cataract surgery. Among those unconvinced, 47.4% asserted that sight restoration depended only on God's will. A greater proportion of women than men (p 0.0000) believed in the association of cataract and menopause, maternity, and menstrual periods and they admitted using herbal and rose teas for treating cataract. CONCLUSIONS: Misbeliefs related to the causes and treatment of senile cataract were identified, most probably of sociocultural basis, indicating the need of education on the subject. PMID- 12131976 TI - [Hygienic conditions of hot dogs sold on the streets, Brazil]. AB - The study objective was to identify the circumstances where there was risk of contamination of food (hot dogs) prepared and sold on the streets and to establish a preventive action through the creation of key surveillance sites. Data were collected from 20 vending sites using interviews, questionnaires, observations of food handling and storage, and temperature and pH measurements of both meat and sauce, respectively. In 30% of the studied sites, hygienic conditions were rated as regular to extremely poor. Mashed potatoes, chicken and beef preparations were of high risk. These findings showed inappropriate hygienic practices of food preparation and lack of basic knowledge regarding food handling - a public health problem. Given the scarcity of literature and official data on food sold in the streets of Brazil, further studies are recommended. PMID- 12131977 TI - [Amount and quality of food advertisement on Brazilian television]. AB - The main objective of the study was to analyze the amount and quality of food advertisement on Brazilian television in three different times of the day. The results showed that food products, when compared to other products, were the most frequently advertised, regardless of the time of the day in a given week analyzed. Television promotes food predominantly high in fat and/or sugar and salt. The large number of high fat and/or sugar and salt products advertised can contribute to changing food habits of children and teenagers, and increasing the incidence of obesity in the population. PMID- 12131978 TI - [Epidemiological aspects of Helicobacter pylori infection in childhood and adolescence]. AB - The scope of the review is to study the epidemiological aspects of Helicobacter pylori infection and its importance during childhood and adolescence, focusing on incidence, prevalence, transmission and risk factors. The study's references included the following databases: LILACS (PAHO/ Bireme), MEDLINE, the US's National Library of Medicine and the thesis developed at University of Sao Paulo for the period 1983 to 1999. It was noted that Helicobacter pylori infection is mainly acquired during childhood, age-related prevalence, main risk factors are associated to low socioeconomic status, and its transmission mechanism remains unclear. PMID- 12131979 TI - [Water resources deterioration and its impact on human health]. AB - The objective of the study is to analyse the actual availability of water resources and its impact on human health deterioration. The following aspects were studied: (a) human activities and environmental deterioration; (b) statistics on the availability and demand of water resources; (c) urban and industry wastes as sources of water resources contamination; and (d) deleterious effect of contaminated water on human health. Statistical data on the impact of contaminated water on human health and the increasing demand of water resources are alarming. It is paramount that modern generations develop an environmental awareness to avoid overstressing water systems, as predicted to come about in the very near future. PMID- 12131980 TI - [A new medical school for a new health system: Ministries of Health and Education are launching plan to change the medical curriculum]. PMID- 12131981 TI - The influence of mouthrinses with antimicrobial solutions on the inhibition of dental plaque and on the levels of mutans streptococci in children. AB - The effect of daily mouthrinses on dental plaque accumulation and on salivary mutans streptococci was investigated in 200 children. The utilized solutions were: a placebo solution composed of mentholated deionized water (group I); 0.12% chlorhexidine gluconate associated to 0.05% sodium fluoride (group II); 0.2% chlorhexidine digluconate (group III), and 0.5% stevioside mixed with 0.05% sodium fluoride, with pH 3.4 (group IV). In order to verify the effect on plaque formation, the accumulation of plaque was assessed by means of the Loe12 index, at the beginning and at the end of the experiment, whereas the quantification of cariogenic streptococci was accomplished on three saliva samples collected at 3 different moments: before the first mouthrinse, 24 hours after the first mouthrinse and 1 week after the last mouthrinse. The mouthrinsing routine was carried out on a daily basis during 4 weeks. Five milliliters of solution were rinsed during 1 minute. The results revealed 4.10, 26.75, 41.20, and 5.91% of reduction in plaque accumulation for groups I, II, III, and IV, respectively. Comparisons between the groups as to plaque reduction revealed that groups II and III were significantly different from groups I (control) and IV (p < 0.05), but did not differ from each other. The solution utilized by group III was the least accepted by children and, as the solution utilized by group II, caused mild dental pigmentation. There were no statistically significant differences as to the levels of mutans streptococci, probably due to the low initial levels observed in each one of the four groups. PMID- 12131982 TI - [Evaluation of surface disinfectants utilized in dentistry]. AB - Surface disinfection is a procedure carried out on the external parts of the dental equipment as well as on other items of the dental office. The aim of this study was to analyze the efficacy of 4 surface disinfectants utilized in dentistry: 77 degrees GL alcohol, phenolic compound (Duplofen), iodophor (PVP-I) and 77 degrees GL alcohol with 5% of chlorhexidine. Four surfaces of the equipment were analyzed in the study (the carter, the washbasin for hand-washing, the headrest of the chair and the external surface of the reflector), and the spray-wipe-spray procedure was carried out. From each surface, samples were collected by means of surface plates containing Mitis Salivarius bacitracin sucrose agar, Sabouraud Dextrose agar with chloramphenicol, MacConkey agar and blood agar, for counting mutans streptococci, Candida yeasts, gram-negative bacteria and total microorganisms, respectively (ufc/plate). The results were statistically analyzed by means of the Student's t test in order to compare the mean ufc/plate values. The most effective disinfectant was 77 degrees GL alcohol with 5% of chlorhexidine, mainly against gram-positive bacteria. Iodophor and phenolic compound were also effective in microbial reduction. 77 degrees GL alcohol was the least effective product - however, although it is not considered as a surface disinfectant, it produced, in this study, statistically significant microbial reduction after the disinfecting procedure. PMID- 12131983 TI - Morphology and thickness of the diffusion of resin through demineralized or unconditioned dentinal matrix. AB - The formation of a hybrid layer is the main bonding mechanism of current dentin bonding systems. This study evaluated the morphology and thickness of the resin infiltrated dentinal layer after the application of adhesive systems. The dentin bonding agents were evaluated on flat dentinal preparations confected on the occlusal surfaces of human teeth. The test specimens were prepared and inspected under scanning electron microscopy at a magnification of X 2,000. The adhesive systems were responsible for different hybrid layer thicknesses (p < 0.05), and the mean values were: for Scotchbond MP Plus (SM), 7.41 +/- 1.24 micrometer for Single Bond (SB), 5.55 +/- 0.82 micrometer for Etch & Prime 3.0 (EP), 3.86 +/- 1.17 micrometer and for Clearfil SE Bond (CB), 1.22 +/- 0.45 micrometer. The results suggest that the conventional three-step adhesive system (SM) was responsible for the thickest hybrid layer, followed by the one-bottle adhesive (SB). The self-etching adhesives, EP and CB, produced the formation of the thinnest hybrid layers. PMID- 12131984 TI - [Comparative study of the fracture resistance of sound upper premolars and upper premolars restored with bonded amalgam]. AB - The purpose of this in vitro study was to determine the fracture resistance of upper premolars which had received class II preparations (conservative and extensive) and were restored with bonded amalgam, with two different adhesive systems. Seventy teeth were divided in four groups: group 1 (control), with ten sound teeth; group 2, with twenty prepared teeth (10 teeth received conservative cavities and 10, extensive cavities) restored with amalgam without any kind of liner; groups 3 and 4, similar to group 2, though with linings of glass ionomer cement (Vitrebond - 3M) (group 3) and dental adhesive (Scotchbond Multi-Purpose Plus - 3M) (group 4). The teeth were previously fixed in PVC cylinders with acrylic resin. After being restored and thermocycled, the test specimens were submitted to fracture by means of compression in an EMIC-MEM 2000 universal testing machine. After the application of the analysis of variance and complementary Tukey's test, we concluded that the utilized adhesive systems produced an increase of the fracture resistance of teeth presenting with conventional cavities; the teeth presenting with conservative cavities were more resistant in all experimental situations. PMID- 12131985 TI - Influence of the spatulation of two zinc oxide-eugenol-based sealers on the obturation of lateral canals. AB - The objective of this research was to evaluate, in vitro, the importance of the correct manipulation of endodontic sealers, correlating it with flow rate and with the consequent obturation of root canals. Twenty-four human canines were prepared, 1 mm from the apex, with K-files up to size 50, by means of the step back technique. Six lateral canals were then drilled in each tooth, with size 10 file fixed to a low-speed handpiece. The teeth were randomly divided into 4 groups, and root canals were obturated either with the Endomethasoneregister mark or target sealer or Grossman sealer, prepared at ideal or incorrect clinical consistency. After obturation by means of the lateral condensation technique, the teeth were radiographed and evaluated as to the number of sealed lateral canals. Statistical analysis revealed significant differences (p < 0.001) between the tested sealers, and indicated the higher capacity of the well-manipulated Grossman sealer to fill lateral canals. It can be concluded that the flow rate of a sealer and its correct manipulation are very important for the satisfactory obturation of lateral canals. PMID- 12131986 TI - [Clinical evaluation of external radicular resorption in non-vital teeth submitted to bleaching]. AB - The purpose of this study was to evaluate the presence of external resorption in non-vital teeth submitted to bleaching. The evaluated patients had at least one non-vital tooth, which had been bleached between 1986 and 1996. All teeth were submitted to bleaching with hydrogen peroxide and sodium perborate, as described by Busato et al.5,6. From 193 patients recalled for clinical and radiographic evaluation of bleached teeth, only 43 attended (54 teeth). The average time elapsed after bleaching was 3.5 years. The results revealed that none of the examined teeth had any degree of external cervical resorption. PMID- 12131987 TI - Microstructural analysis of demineralized primary enamel after in vitro toothbrushing. AB - The aim of this study was to investigate, in vitro, the morphological characteristics of demineralized primary enamel subjected to brushing with a dentifrice with or without fluoride. In order to do so, 32 enamel blocks were divided in 4 different groups containing 8 blocks each. They were separately immersed in artificial saliva for 15 days. The experimental groups were: C - control; E - submitted to etching with 37% phosphoric acid gel (30 s); EB - submitted to etching and brushing 3 times a day with a non-fluoridated dentifrice; EBF = submitted to etching and brushing 3 times a day with a fluoridated dentifrice. The toothbrushing force was standardized at 0.2 kgf and 15 double strokes were performed on each block. After the experimental period, the samples were prepared and examined under SEM. The control group (C) showed a smooth surface, presenting scratches caused by habitual toothbrushing. The etched samples (E) exhibited different degrees of surface disintegration, but the pattern of acid etching was predominantly the type II dissolution. The brushed surfaces were smooth, with elevations which corresponded to the exposure of Tomes' process pits and depressions which corresponded to interrod enamel. Particles resembling calcium carbonate were found in the most protected parts of the grooves. No morphological differences were observed between brushing with fluoridated (EBF) and non-fluoridated (EB) dentifrice. The results suggest that the mechanical abrasion caused by brushing demineralized enamel with dentifrice smoothes the rough etched surface, and the presence of fluoride does not cause morphological modifications in this pattern. PMID- 12131988 TI - [Relation between biofilm, caries activity and gingivitis in HIV + children]. AB - The utilization of medicines to treat HIV-infected children has been promoting a decrease in the prevalence of soft-tissue oral lesions, as years pass by. In contrast, it has been observed that the experience of caries and gingivitis is constant in this population, mostly because of the chronic influence of some factors involved in the HIV-infection process, such as the chronic utilization of sweetened liquid medicines and carbohydrate-enriched diet, as well as frequent episodes of hospitalization. So, the purpose of this study was to evaluate if the quality and quantity of biofilm are important factors in the activity of dental caries and gingivitis, also in this special group. After examination of the biofilm (biofilm index - Ribeiro23, 2000), the activity of caries and gingivitis was assessed in 56 children, aging from 0 to 14 years, who were patients with definitive diagnosis of HIV infection. It was observed that only 7 subjects (12.5%) did not present with clinically visible biofilm, and 33 (58.9%) presented with gingivitis, with the average of 4.44 bleeding sites. As to dental caries, 73.2% of the patients presented with active carious lesions. A strong correlation was verified between Biofilm Index, gingival status and active carious lesions (Spearman's correlation test, r s = +0.57 and r s = +0.49, respectively). It was concluded that, also in HIV-infected children, the quality and quantity of biofilm over the dental surfaces are important etiologic factors related to the activity of caries and gingivitis. Biofilm should, thus, be controlled in order to reestablish the oral health of HIV-infected children. PMID- 12131989 TI - [Dissection of carious lesions: a new technique for three-dimensional histopathological study]. AB - Two-dimensional analyses of the histopathological aspect of carious lesions present bias that may compromise the scientific value of studies. In this paper, the authors present a new three-dimensional technique for the histopathological analysis of carious lesions. It consists in the dissection of the affected areas under the stereomicroscope, utilizing refrigerated dental burs, without any loss of carious tissue. The dissection of three occlusal lesions located on first and second permanent molars is presented. It was possible to observe the internal reactions of enamel and dentin, and correlate the results to the macroscopic features of the occlusal surface. Some comparative aspects between the dissecting and two-dimensional techniques are discussed. PMID- 12131990 TI - [Immunohistochemical study of components of the basement membrane in odontogenic cysts]. AB - The pattern of distribution and expression of laminin and type IV collagen was studied in ten radicular cysts, ten dentigerous cysts and ten odontogenic keratocysts, by means of the streptavidin-biotin method. The purpose of this study was to investigate changes in the distribution of components of the basement membrane in an attempt to contribute to the understanding of the differences, as to evolution and clinical behavior, between these cysts. The results revealed a weak and discontinuous linear staining, in odontogenic keratocysts, for both laminin and type IV collagen, while, in radicular cysts, staining was more intense and continuous. In dentigerous cysts, an intermediary pattern was observed, which was more similar to that observed in keratocysts. Our results suggest that, in view of the weak expression of proteins of the basement membrane observed in odontogenic keratocysts, modifications must take place in the interaction between the epithelium and the adjacent connective tissue, which could, in part, contribute to the pattern of more rapid growth exhibited by these cysts. PMID- 12131991 TI - [Prevalence of aggressive periodontitis in adolescents and young adults from Vale do Paraiba]. AB - The aim of the present study was to determine the prevalence of localized and generalized aggressive periodontitis, as well as of incidental attachment loss, in a population of adolescents and young adults aging between 15 and 25 years (19.4 +/- 3.44) from Vale do Paraiba - SP, who searched for general dental care at the Department of Dentistry, University of Taubate, Sao Paulo. Six hundred patients, 244 male and 356 female subjects, were included in the studied sample. The periodontal status of this population was evaluated by measuring the depth of periodontal pockets, as well as attachment loss. The data were confirmed by means of radiographic examination. Measurements were performed in six sites per tooth. Ten subjects (1.66%) were diagnosed as having localized aggressive periodontitis, 2 males (aging 18.5 +/- 2.12 years) and 8 females (aging 19.2 +/- 3.91 years); 22 (3.66%) presented with generalized aggressive periodontitis, 6 males (aging 19.1 +/- 3.06 years) and 16 females (aging 20.1 +/- 2.71 years); and 86 individuals (14.3%) presented with incipient periodontitis, 29 males (aging 20.2 +/- 2.87 years) and 57 females (aging 21.1 +/- 2.79 years). There was a positive correlation between the female gender and the occurrence of periodontal disease. PMID- 12131992 TI - [Evaluation of the reduction of the visible plaque index and of the gum bleeding index in a program of oral health promotion for children]. AB - The purpose of this research was to evaluate the reduction of the VPI (visible plaque index) and of the GBI (gum bleeding index) in children seen at a unit of SUS, in the state of Rio de Janeiro. A comparative and statistical procedure (t test, by means of the GMC program) was employed in order to assess the variation between initial and final mean VPI and GBI values. Initial values were registered in the beginning of the assistance program, and final values were registered after the children had participated in health promotion activities, which included weekly supervised brushing sessions, individual dietary guidance, collective instructive activities in which parents were present, adjusting of the oral environment, as well as restorative and surgical dental care, during an average period of six months. The studied sample comprised fourty-two children, with an average age of nine years, who presented an average of four teeth affected by caries. The technique of indirect documentation, by means of documental research, was employed. The following results were obtained: initial VPI = 29%; final VPI = 11%; initial GBI = 13% and final GBI = 5%. The observed variations were statistically significant at the level of 1%, which was revealed by the applied statistical test. The program proved efficient as to plaque control, reducing VPI to a more acceptable level. Although GBI presented considerable reduction, the presence of bleeding at the end of the program calls for a better approach in motivating patients as to regular oral hygiene. PMID- 12131993 TI - [Total resection of the zygomatic arch in the growth period: experimental study in rats]. AB - The results obtained after total unilateral resection of the zygomatic arch in Wistar rats were evaluated by means of cephalometric measurements. The resection of the right zygomatic arch was carried out in one-month-old rats, and the animals were sacrificed two months later. The skull and hemimandibles were submitted to axial and lateral radiographic incidences. Based on the obtained radiographs, measurements were carried out by means of a computer system, which compared both sides of the specimens. The obtained values were submitted to statistical analysis. There was significant difference as to the extent of the temporal fossa, but there was no significant difference as to other measures of the maxilla. There was significant difference between both sides regarding the height of the body and the length of the base of the mandible. PMID- 12131995 TI - Tuberculosis control: how the world has changed since 1990. PMID- 12131996 TI - Response to a major disease of poverty: the Global Partnership to Stop TB. PMID- 12131997 TI - Joining forces to develop weapons against TB: together we must. PMID- 12131998 TI - The DOTS strategy in China: results and lessons after 10 years. AB - OBJECTIVE: To analyse the five-point tuberculosis (TB) strategy, DOTS, 10 years after its implementation in one-half of China's population, and to suggest lessons for future implementation of the DOTS strategy. METHODS: We analysed trends in case-finding and treatment outcome over time following implementation of the DOTS strategy in each county, using routine reporting data from the Infectious and Endemic Disease Control (IEDC) project (1991 - 2000). We also determined the proportion of counties with different levels of case-finding for the fifth and sixth years of DOTS implementation. FINDINGS: From 1991 to 1995, DOTS expanded rapidly to cover more than 90% of target population and counties. By 2000, 8 million TB suspects had received free diagnostic evaluation: 1.8 million TB cases were diagnosed, free treatment was provided to 1.3 million smear positive cases, and more than 90% were cured. During DOTS implementation, the percentage of previously treated cases decreased among all smear-positive cases and treatment outcomes improved. Despite these achievements, the detection rate for new smear-positive cases in the project was estimated to be only 54% in 1998, and 41.2% of the counties had a below average or low level of case-finding (with substantial variation between provinces). CONCLUSIONS: The IEDC project demonstrated that it is feasible to rapidly expand DOTS on a large scale. The global target of an 85% cure rate was quickly achieved, and the level of drug resistance was probably reduced by this project. However, case-detection did not reach the 70% global target, and more research is needed on how to enhance this. PMID- 12131999 TI - Low access to a highly effective therapy: a challenge for international tuberculosis control. AB - OBJECTIVE: To determine the scale of the tuberculosis (TB) problem facing the international Stop TB Partnership by measuring the gap between present rates of case detection and treatment success, and the global targets (70% and 85%, respectively) to be reached by 2005 under the WHO DOTS strategy. METHODS: We analysed case notifications submitted annually to WHO from up to 202 (of 210) countries and territories between 1980 and 2000, and the results of treatment for patients registered between 1994 and 1999. FINDINGS: Many of the 148 national DOTS programmes in existence by the end of 2000 have shown that they can achieve high treatment success rates, close to or exceeding the target of 85%. However, we estimate that only 27% of all the new smear-positive cases that arose in 2000 were notified under DOTS, and only 19% were successfully treated. The increment in case-finding has been steady at about 133 000 additional smear-positive cases in each year since 1994. In the interval 1999- 2000, more than half of the extra cases notified under DOTS were in Ethiopia, India, Myanmar, the Philippines, and South Africa. CONCLUSION: With the current rate of progress in DOTS expansion, the target of 70% case detection will not be reached until 2013. To reach this target by 2005, DOTS programmes must find an additional 333 000 cases each year. The challenge now is to show that DOTS expansion in the major endemic countries can significantly accelerate case finding while maintaining high cure rates. PMID- 12132000 TI - Cost-effectiveness of community health workers in tuberculosis control in Bangladesh. AB - OBJECTIVE: To compare the cost-effectiveness of the tuberculosis (TB) programme run by the Bangladesh Rural Advancement Committee (BRAC), which uses community health workers (CHWs), with that of the government TB programme which does not use CHWs. METHODS: TB control statistics and cost data for July 1996 - June 1997 were collected from both government and BRAC thanas (subdistricts) in rural Bangladesh. To measure the cost per patient cured, total costs were divided by the total number of patients cured. FINDINGS: In the BRAC and government areas, respectively, a total of 186 and 185 TB patients were identified over one year, with cure rates among sputum-positive patients of 84% and 82%. However, the cost per patient cured was US$ 64 in the BRAC area compared to US$ 96 in the government area. CONCLUSION: The government programme was 50% more expensive for similar outcomes. Although both the BRAC and government TB control programmes appeared to achieve satisfactory cure rates using DOTS (a five-point strategy), the involvement of CHWs was found to be more cost-effective in rural Bangladesh. With the same budget, the BRAC programme could cure three TB patients for every two in the government programme. PMID- 12132001 TI - DOTS-based tuberculosis treatment and control during civil conflict and an HIV epidemic, Churachandpur District, India. AB - OBJECTIVE: To pilot the WHO guidelines on DOTS for tuberculosis (TB) among displaced people affected by conflict in Churachandpur District, Manipur State, north-east India, which has endured an HIV epidemic, injecting drug use, civil unrest, high levels of TB, and poor TB treatment and prevention services for many years. METHODS: Prerequisites for TB control programmes were established. WHO guidelines and protocols were adapted for local use. Outreach workers were appointed from each ethnic group involved in the conflict, and training was conducted. Quality control and evaluation processes were introduced. FINDINGS: TB was diagnosed in 178 people between June and December 1998. Of the 170 with pulmonary disease, 85 were smear-positive. Successful outcomes were recorded in 91% of all patients and in 86% of smear-positive cases of pulmonary TB. The default rate and the mortality rate were low at 3% each. HIV positive serostatus was the only factor associated with a poor treatment outcome. CONCLUSION: TB treatment and control were possible in a conflict setting and WHO targets for cure were attainable. The factors associated with the success of the programme were strong local community support, the selection of outreach workers from each ethnic group to allow access to all areas and patients, the use of directly observed therapy three times a week instead of daily in the interest of increased safety, and the limiting of distances travelled by both outreach workers and patients. PMID- 12132002 TI - Rapid DOTS expansion in India. AB - Since late 1998 the coverage of the DOTS strategy in India has been expanded rapidly. In both 2000 and 2001 the country probably accounted for more than half the global increase in the number of patients treated under DOTS and by early 2002 more than a million patients were being treated in this way in India. As a result, nearly 200 000 lives were saved. The lessons learnt relate to the importance of the following elements of the programme: (1) getting the science right and ensuring technical excellence; (2) building commitment and ensuring the provision of funds and flexibility in their utilization; (3) maintaining focus and priorities; (4) systematically appraising each area before starting service delivery; (5) ensuring an uninterrupted drug supply; (6) strengthening the established infrastructure and providing support for staff; (7) supporting the infrastructure required in urban areas; (8) ensuring full-time independent technical support and supervision, particularly during the initial phases of implementation; (9) monitoring intensively and giving timely feedback; and (10) continuous supervision. Tuberculosis (TB) control still faces major challenges in India. To reach its potential, the control programme needs to: continue to expand so as to cover the remaining half of the country, much of which has a weaker health infrastructure than the areas already covered; increase its reach in the areas already covered so that a greater proportion of patients is treated; ensure sustainability; improve the patient-friendliness of services; confront TB associated with human immunodeficiency virus (HIV) infection. It is expected that HIV will increase the number of TB cases by at least 10% and by a considerably higher percentage if HIV becomes much more widespread. India's experience shows that DOTS can achieve high case-detection and cure rates even with imperfect technology and often with an inadequate public health infrastructure. However, this can only happen if the delivery programme is appropriately designed and effectively managed. PMID- 12132003 TI - Highly active antiretroviral therapy and tuberculosis control in Africa: synergies and potential. AB - HIV/AIDS (human immunodeficiency virus/acquired immunodeficiency syndrome) and TB (tuberculosis) are two of the world's major pandemics, the brunt of which falls on sub-Saharan Africa. Efforts aimed at controlling HIV/AIDS have largely focused on prevention, little attention having been paid to care. Work on TB control has concentrated on case detection and treatment. HIV infection has complicated the control of tuberculosis. There is unlikely to be a decline in the number of cases of TB unless additional strategies are developed to control both this disease and HIV simultaneously. Such strategies would include active case-finding in situations where TB transmission is high, the provision of a package of care for HIV-related illness, and the application of highly active antiretroviral therapy. The latter is likely to have the greatest impact, but for this therapy to become more accessible in Africa the drugs would have to be made available through international support and a programme structure would have to be developed for its administration. It could be delivered by means of a structure based on the five-point strategy called DOTS, which has been adopted for TB control. However, it may be unrealistic to give TB control programmes the responsibility for running such a programme. A better approach might be to deliver highly active antiretroviral therapy within a comprehensive HIV/AIDS management strategy complementing the preventive work already being undertaken by AIDS control programmes. TB programmes could contribute towards the development and implementation of this strategy. PMID- 12132004 TI - TB and HIV: joint problems, joint solutions? PMID- 12132005 TI - The research agenda for improving health policy, systems performance, and service delivery for tuberculosis control: a WHO perspective. AB - The development of WHO's DOTS strategy for the control of tuberculosis (TB) in 1995 led to the expansion, adaptation and improvement of operational research in this area. From being a patchwork of small-scale studies concerned with aspects of service delivery, TB operational research shifted to larger-scale, often multicountry projects that were also concerned with health policy and the needs of health systems. The results are now being put into practice by national TB control programmes. In 1998 an ad hoc committee identified the chief factors inhibiting the expansion of DOTS: lack of political will and commitment, poor financial support for TB control, poor organization and management of health services, inadequate human resources, irregular drug supplies, the HIV epidemic, and the rise of multidrug resistance. An analysis of current operational research on TB is presented on the basis of these constraints, and examples of successful projects are outlined in the article. We discuss the prerequisites for success, the shortcomings of this WHO- supported programme, and future challenges and needs. PMID- 12132006 TI - Molecular epidemiology of tuberculosis: achievements and challenges to current knowledge. AB - Over the past 10 years, molecular methods have become available with which to strain-type Mycobacterium tuberculosis. They have allowed researchers to study certain important but previously unresolved issues in the epidemiology of tuberculosis (TB). For example, some unsuspected microepidemics have been revealed and it has been shown that the relative contribution of recently acquired disease to the TB burden in many settings is far greater than had been thought. These findings have led to the strengthening of TB control. Other research has demonstrated the existence and described the frequency of exogenous reinfection in areas of high incidence. Much recent work has focused on the phenotypic variation among strains and has evaluated the relative transmissibility, virulence, and immunogenicity of different lineages of the organism. We summarize the recent achievements in TB epidemiology associated with the introduction of DNA fingerprinting techniques, and consider the implications of this technology for the design and analysis of epidemiological studies. PMID- 12132007 TI - What's new in tuberculosis vaccines? AB - Over the past 10 years, tuberculosis (TB) vaccine development has resurged as an active area of investigation. The renewed interest has been stimulated by the recognition that, although BCG is delivered to approximately 90% of all neonates globally through the Expanded Programme on Immunization, Mycobacterium tuberculosis continues to cause over 8 million new cases of TB and over 2 million deaths annually. Over one hundred TB vaccine candidates have been developed, using different approaches to inducing protective immunity. Candidate vaccines are typically screened in small animal models of primary TB disease for their ability to protect against a virulent strain of M. tuberculosis. The most promising are now beginning to enter human safety trials, marking real progress in this field for the first time in 80 years. PMID- 12132008 TI - Controlling multidrug-resistant tuberculosis and access to expensive drugs: a rational framework. AB - The emergence and spread of multidrug-resistant tuberculosis (MDR-TB), i.e. involving resistance to at least isoniazid and rifampicin, could threaten the control of TB globally. Controversy has emerged about the best way of confronting MDR-TB in settings with very limited resources. In 1999, the World Health Organization (WHO) created a working group on DOTS-Plus, an initiative exploring the programmatic feasibility and cost-effectiveness of treating MDR-TB in low income and middle-income countries, in order to consider the management of MDR-TB under programme conditions. The challenges of implementation have proved more daunting than those of access to second-line drugs, the prices of which are dropping. Using data from the WHO/International Union Against Tuberculosis and Lung Disease surveillance project, we have grouped countries according to the proportion of TB patients completing treatment successfully and the level of MDR TB among previously untreated patients. The resulting matrix provides a reasonable framework for deciding whether to use second-line drugs in a national programme. Countries in which the treatment success rate, i.e. the proportion of new patients who complete the scheduled treatment, irrespective of whether bacteriological cure is documented, is below 70% should give the highest priority to introducing or improving DOTS, the five-point TB control strategy recommended by WHO and the International Union Against Tuberculosis and Lung Disease. A poorly functioning programme can create MDR-TB much faster than it can be treated, even if unlimited resources are available. There is no single prescription for controlling MDR-TB but the various tools available should be applied wisely. Firstly, good DOTS and infection control; then appropriate use of second-line drug treatment. The interval between the two depends on the local context and resources. As funds are allocated to treat MDR-TB, human and financial resources should be increased to expand DOTS worldwide. PMID- 12132011 TI - Annual risk of tuberculous infection. 1988. PMID- 12132012 TI - Diagnostic testing in the control of tuberculosis. PMID- 12132014 TI - Global Alliance at full steam for new TB drugs. PMID- 12132015 TI - A new TB drug by 2010--or sooner? PMID- 12132016 TI - Estonia races to halt multidrug-resistant TB before HIV takes hold. PMID- 12132017 TI - Raju has TB and AIDS and lives on the street. PMID- 12132018 TI - Reading the rice genome starts a new chapter in food science. PMID- 12132019 TI - South Africa breaks through to realism on antiretrovirals for AIDS. PMID- 12132020 TI - New report explains how genome research is transforming the world's prospects for health. PMID- 12132024 TI - Increased medial TUNEL-positive staining associated with apoptotic bodies is linked to smooth muscle cell diminution during evolution of abdominal aortic aneurysms. AB - Apoptosis has recently been identified as an important process in large vessel structural integrity. We examined whether the size of the abdominal aortic aneurysm might be associated with programmed cell death. We performed in situ labeling of the 3' ends of DNA fragments by apoptosis-associated endonucleases in 20 aneurysms, 10 controls with aortoiliac occlusive disease, and 4 controls with healthy aortas. Antibodies against a-smooth muscle actin were used to quantify smooth muscle cell alterations in the medial layer. Inflammatory cell characterization was made by using four monoclonal mouse antibodies (UCHL1, L26, PG-M1, and KP1). The results confirm the assumption that an apoptotic process may be of consequence for the loss of medial smooth muscle cells in the early evolution of an abdominal aortic aneurysm process. PMID- 12132026 TI - Differences in neointima formation between impervious and porous polytetrafluoroethylene vascular patch material. AB - The aim of this study was to compare the neointima formation and blood loss of an impervious ePTFE with those of the porous ePTFE patch. Ten mongrel dogs were selected for the study. Both the impervious and porous ePTFE patches were implanted into the common iliac arteries in each dog. The blood loss of each patch was recorded and the patches were explanted 30-60 days later for microscopic analysis. The arteries with either the impervious or the porous ePTFE patch were all patent at the end of the study. The impervious ePTFE patch had a significant reduction in blood loss when compared with the porous ePTFE patch (p = 0.04). The neointima covering both patches showed no statistically significant difference in its thickness or in its cellular composition. It has been speculated that wall porosity is essential for tissue ingrowth, endothelial cell proliferation, and neointima formation. In this study, the impervious ePTFE patch did not inhibit neointima formation. Our study shows that graft porosity does not improve neointima formation or patency. Neointimization of the impervious ePTFE patch is the result of pannus ingrowth and deposition of circulating blood elements. There is a statistically significant reduction in blood loss (p = 0.04) with impervious ePTFE patch, compared with that of the porous ePTFE patch. PMID- 12132027 TI - Percutaneous balloon pericardial window for patients with symptomatic pericardial effusion. AB - PURPOSE: To describe the technique and our experience in percutaneous creation of a pericardial window in patients with recurrent pericardial effusion. METHODS: Eleven consecutive patients (9 men, 2 women; mean age 61 years, range 37-79 years) with recurrent pericardial effusion were treated from December 1994 to October 2000. Malignant effusion was the cause of cardiac tamponade in nine patients. Puncture of the pericardial space was performed under ultrasound and fluoroscopic guidance. Once a guidewire was safely positioned in the pericardial cavity and the wall of the pericardium was identified, the pericardial keyhole was dilated with an 18 mm balloon catheter. A temporary (1-3 day) 8 Fr pigtail catheter was inserted in order to stabilize the tract and aspirate the fluid. RESULTS: The aim of creating a pericardial window was achieved in all 11 patients. There were three recurrences (27%) of pericardial effusion and three early deaths (27%) due to progression of the underlying disease, which occurred within 30 days following the procedure. Follow-up ranged from 6 days to 382 days, mean 147 days. CONCLUSIONS: Percutaneous creation of a pericardial window can be a safe therapeutic alternative for patients with symptomatic pericardial effusion, particularly if this has a malignant etiology. PMID- 12132028 TI - Carbon-dioxide-guided vascular interventions: technique and pitfalls. AB - PURPOSE: To evaluate the usefulness of carbon dioxide (CO2) angiography to guide vascular interventions. METHODS: A prospective study was carried out of 50 procedures (angioplasty, stenting, stent-grafting and embolization) using CO2 angiography. Indications for using CO2 were renal impairment, cardiac failure, previous reaction to conventional iodinated contrast, or likelihood of needing high doses of conventional contrast. CO2 was intended to be the sole contrast agent. The use of additional conventional contrast or gadolinium was recorded, as were procedural complications. Radiation dose was compared with similar procedures using conventional contrast. RESULTS: Angiographic quality was satisfactory in 44 (88%) procedures and CO2 guidance was all that was required; in 6 (12%) cases adjunctive use of conventional contrast or gadolinium was necessary. Contrast doses were significantly reduced and there was a trend toward decreased radiation doses with CO2. There were two significant complications but only one related to the use of CO2. CONCLUSION: CO2 angiography is well tolerated and can be successfully used to guide even complex vascular interventions. High risk patients can be spared the risks of conventional contrast agents. PMID- 12132029 TI - TIPS creation in a patient with situs inversus totalis. AB - We describe the successful creation of a transjugular intrahepatic portosystemic shunt (TIPS) in a patient with complete situs inversus using a simple modification of the standard TIPS technique. PMID- 12132030 TI - A prospective randomized trial comparing the safety and efficacy of three commercially available closure devices (Angioseal, Vasoseal and Duett). AB - PURPOSE: We compared the safety and efficacy of three closure devices (Angioseal, Vasoseal and Duett) used to close arterial puncture sites in patients who underwent coronary percutaneous procedures. METHODS: A prospective randomized, single-center trial was carried out of consecutive patients who underwent coronary angiography [705 patients: Angioseal (243),Vasoseal (228) and Duett (234)] or angioplasty [146 patients:Angioseal (47), Vasoseal (52) and Duett (47)]. RESULTS: In the angiography patients the device deployment rates were similar, with the Angioseal been significantly slower in achieving hemostasis (p = 0.0001) but resulting in earlier ambulation (p = 0.0001). In the coronary angioplasty patients the deployment rates were similar to those for angiography: time to hemostasis was longer for the Angioseal (p = 0.003), while ambulation times were not different, although prolonged compared with angiography (p = 0.0001). The three devices had similar major complication rates. The Vasoseal had a higher major complication rate after angioplasty than after angiography (p = 0.004). The incidence rate of peripheral embolization was lower when the Angioseal was utilized. Severe complications were mainly seen in patients who received abciximab. CONCLUSIONS: The three closure devices had high rates of successful deployment and were relatively safe. The Angioseal resulted in earlier ambulation after angiography. Utilization of closure devices after abciximab administration possibly increased the complications. PMID- 12132038 TI - Mechanism of retinoic acid induced attenuation of PTH action in UMR 106-01 cells. AB - Studies from our laboratory in osteoblast-like cells have shown that the increase in EGF receptor expression in response to PTH was cyclic AMP mediated and was blocked by treatment with retinoic acid (RA). The present studies investigate the mechanism for this effect of RA on PTH actions. UMR 106-01 cells were exposed to RA and were tested for cAMP response to PTH as well as for (125)I PTH binding. cAMP production in response to PTH was markedly decreased by RA (25.1 +/- 1.6% of control) whereas there was only a slight decrease in PTH binding in response to RA. For the study of adenylate cyclase activity, membranes were isolated from intact cells that had been exposed to RA. Treatment with RA decreased PTH stimulated adenylate cyclase activity; however, forskolin-stimulated enzyme activity was unchanged. Treatment of intact cells with pertussis toxin, to inactivate Gi, did not alter the inhibitory effect of RA on PTH-stimulated adenylate cyclase activity. Addition of GppNHp, a non-hydrolyzable analogue of GTP, completely restored the response to PTH in the membranes. Therefore, we examined the activity of IMP dehydrogenase, the rate-limiting enzyme for GTP biosynthesis, and GMP reductase which counteracts the effect of the synthetic enzyme. Treatment with RA for 48 hours increased GMP reductase activity by 240.9 +/- 24.2% and decreased IMP dehydrogenase activity to 67.5 +/- 8.8% of control values. These data indicate that RA impairs the response to PTH in intact cells. This blunted response was preserved in membrane preparations but was corrected by GTP. The RA-induced alterations of enzymes involved in the GTP biosynthetic pathway in a direction that favors a decrease in GTP biosynthesis provide an explanation for the inhibitory effect of RA on PTH actions. PMID- 12132043 TI - Low transplant-related mortality after second allogeneic peripheral blood stem cell transplant with reduced-intensity conditioning in adult patients who have failed a prior autologous transplant. AB - Standard allogeneic stem cell transplantation (SCT) has been associated with a high transplant-related mortality (TRM) in patients who have failed a prior autologous SCT (ASCT). Reduced-intensity conditioning (RIC) regimens may reduce the toxicities and TRM of traditional myeloablative transplants. We report 46 adults who received a RIC peripheral blood SCT from an HLA-identical sibling in two multicenter prospective studies. The median interval between ASCT and allograft was 16 months, and the patients were allografted due to disease progression (n = 43) and/or secondary myelodysplasia (n = 4). Conditioning regimens consisted of fludarabine plus melphalan (n = 41) or busulphan (n = 5). The 100-day incidence of grade II-IV acute graft-versus-host disease (GVHD) was 42% (24% grade III-IV), and 10/30 evaluable patients developed chronic extensive GVHD. Early complete donor chimerism in bone marrow and peripheral blood was observed in 35/42 (83%) patients, and 16 evaluable patients had complete chimerism 1 year post transplant. With a median follow-up of 358 days (450 in 29 survivors), the 1-year incidence of TRM was 24%, and the 1-year overall (OS) and progression-free survival were 63% and 57%, respectively. Patients who had chemorefractory/ progressive disease, a low performance status or received GVHD prophylaxis with cyclosporine A alone (n = 32) had a 1-year TRM of 35% and an OS of 46%, while patients who had none of these characteristics (n = 32) had a 1 year TRM of 35% and an OS of 46% while patients who had none of these characteristics (n = 14) had a TRM of 0% and an OS of 100%. Our results suggest that adult patients who fail a prior ASCT can be salvaged with a RIC allogeneic PBSCT with a low risk of TRM, although patient selection has a profound influence on early outcome. PMID- 12132044 TI - A large-scale method for T cell depletion: towards graft engineering of mobilized peripheral blood stem cells. AB - We have investigated the feasibility and efficacy of large-scale T cell depletion from granulocyte colony-stimulating factor (G-CSF) mobilized peripheral blood stem cells (PBSC). The method is based on the use of a CD3 antibody conjugated to magnetic microbeads and magnetic activated cell sorting (Clinimacs). A total of eight large-scale experiments were performed. In four experiments, CD3(+) T cells were depleted from PBSC obtained from volunteers mobilized with G-CSF whereas, in four experiments, T cells were depleted from PBSC from stem cell donors, in which the CD34(+) stem cells had been removed for allogeneic transplantation by positive selection prior to T cell depletion. The mean number of processed mononuclear cells (MNCs) was 3.3 x 10(10) (range 1.5 x 10(10)-5.1 x 10(10)) with a mean T cell proportion of 35.8% (range 16.7-64.0%). After T cell depletion, the percentage of contaminating T cells was 0.15% (range 0.01-1.01%) with a mean log(10) depletion of 3.4 (range 2.8-4.1). The mean recovery of CD3-negative MNCs after depletion was 76% (range 52-100%). The mean recovery of CD34(+) stem cells in the four evaluable experiments was 82% (range 75-92%). In vitro colony assays and in vivo NOD/SCID repopulation assays showed that this large-scale T cell depletion method has no negative impact on the function of the hematopoietic precursor cells. Therefore, we conclude that this T cell depletion method is a valuable tool for further graft engineering strategies involving mobilized PBSCs. PMID- 12132045 TI - Circulating hematopoietic progenitor cells in a fetus with alpha thalassemia: comparison with the cells circulating in normal and non-thalassemic anemia fetuses and implications for in utero transplantations. AB - Our aim was to evaluate the number of progenitor cells circulating in an alpha thalassemic fetus during its infusion in utero with paternal CD34(+) and adult red cells and to compare those values with those circulating in normal and non thalassemic anemic fetuses of matched gestational age. The treatment of the alpha thalassemic fetus has been described elsewhere. Fetal blood was obtained from normal and anemic fetuses by fetal blood sampling for diagnostic or therapeutic purposes according to a protocol approved by the human subject committee. The number of progenitor cells in fetal blood was estimated on the basis of the number of colonies they gave rise to in semisolid cultures. The alpha-thalassemic fetus, as did the other fetuses analyzed, contained high numbers (10(6)-10(7) depending on the age) of progenitor cells, values which were higher than the number (10(4)-10(5)) of paternal progenitor cells being transplanted. Progenitor cells with adult characteristics (adult kinetics of differentiation) were detected rapidly (10 min) after the CD34(+) cell infusion, but were not detectable 2-3 weeks after the transplant. These results indicate that adult progenitor cells do not have a numerical advantage when transplanted into alpha thalassemic fetuses. PMID- 12132046 TI - GM-CSF-based mobilization effect in normal healthy donors for allogeneic peripheral blood stem cell transplantation. AB - It is important to optimize methods to mobilize hematopoietic stem cells into peripheral blood (PB) for successful allogeneic peripheral blood stem cell (PBSC) transplantation. Our primary intent was to investigate the role of GM-CSF for mobilization in normal healthy donors and to compare its efficacy in mobilizing stem cells alone, in concurrent combination and in sequential combination with G CSF in this study. We analyzed the results of the PBSC harvest through large volume leukapheresis from 48 normal healthy donors mobilized by three different regimens including GM-CSF. Donors were assigned sequentially to one of the following regimens for mobilization: GM-CSF 10 microg/kg/day alone (group 1, n = 9); concurrent combination (group 2, n = 20) of G-CSF 5 microg/kg/day and GM-CSF 5 microg/kg/day; sequential combination (group 3, n = 19) of GM-CSF alone 10 microg/kg/day for 3 days followed by G-CSF alone 10 microg/kg/day for 2-3 days. The harvested CD34(+) cell count (P < 0.05) was statistically higher in group 3 than in group 1 or 2. Pre-collection WBC count in donors (P < 0.05), harvested MNC (P < 0.05) and CD3(+) cell count (P < 0.05) of group 2 or 3 were significantly higher than those of group 1. Recipients who received stem cells mobilized with combination regimens showed an earlier recovery of WBC and platelets count than those with GM-CSF alone. The incidence of acute graft-versus host disease was not statistically different among three recipient groups. GM-CSF based mobilization was well tolerated in normal healthy donors. The sequential combination regimen appears to be an excellent mobilization strategy and might be preferred as the optimal method in some clinical situations that need a higher number of stem cells. PMID- 12132047 TI - Consequences of ABO incompatibility in allogeneic hematopoietic stem cell transplantation. AB - Aside from causing hemolytic reactions the ABO blood group system does not have an impact on outcome after allogeneic bone marrow or peripheral blood stem cell transplantation (SCT). However, only a few studies have addressed the effect of ABO incompatibility on the incidence of GVHD, time to engraftment, relapse and survival. Therefore, we performed a retrospective two-center analysis of 562 consecutive patients receiving allogeneic SCT, including 361 ABO-identical, 98 minor, 86 major and 17 bidirectional ABO-incompatible SCT. In multivariate analysis adjusted for potential confounders survival was significantly associated with ABO incompatibility (P = 0.006). Compared to ABO-identical SCT, bidirectional ABO incompatibility increased the risk significantly (RR, 2.8; 95% CI, 1.5-5.1; P = 0.0009), whereas survival of patients with minor (RR, 1.2; 95% CI, 0.9-1.7; P = 0.27), or major ABO-incompatible SCT (RR, 1.3; 95% CI, 0.9-1.8; P= 0.18) was not significantly different. RBC engraftment was delayed in major ABO-incompatible SCT (RR, 0.66; 95% CI, 0.51-0.85; P = 0.001). The incidence of acute GVHD (grade I-IV) was higher in minor ABO-incompatible SCT as compared to ABO identity (RR, 2.8; 95% CI, 1.3-5.9, P = 0.009). This difference was limited to mild GVHD; in moderate-to-severe GVHD (grade II-IV) no significant difference was found among the groups (P = 0.53). The relapse rate was not influenced by ABO incompatibility (P = 0.78). In conclusion, these results suggest that ABO incompatibility represents a risk factor not only for post-transplant hemolysis, but also for survival and the rate of mild GVHD after allogeneic SCT. PMID- 12132048 TI - Allogeneic hematopoietic stem cell transplantation in ovarian carcinoma: results of five patients. AB - Allogeneic hematopoietic stem cell transplantation is often used to treat hematologic malignancies. The efficacy of this procedure is due to both myeloablative conditioning and graft-versus-leukemia (GVL). However, the disadvantages of allogeneic transplantation include graft-versus-host disease (GVHD), relapse from the original tumor, and patient susceptibility to opportunistic infections. Lately, allogeneic transplantation has been developed to treat solid tumors, with the expectation that graft-versus-tumor (GVT), like GVL, will have a significant anti-tumor effect. This effect has been demonstrated in renal carcinomas, and with less evidence in breast cancers. Five patients with malignant ovarian tumors resistant to chemotherapy underwent allogeneic transplantation, four from bone marrow, and one from peripheral blood stem cells. All donors were HLA-identical siblings. One patient received a myeloablative conditioning regimen, while the other four received a non-myeloablative regimen. Two patients received donor lymphocyte infusions (DLI). Four of the patients presented with acute or chronic GVHD associated with tumor regression of at least 50%. These tumor regressions were measured by CA-125 levels and CT scans. The fifth patient died of rapid progression just after transplantation. Of the four transplantation survivors, three received a non-myeloablative regimen which did not seem to reduce treatment effectiveness. While it did reduce toxicity, one of these patients died of GVHD after 127 days. DLI was administered to two patients. These infusions seemed to promote GVHD which was able to control disease progression for one patient and had no apparent effect on the other. Allograft of hematopoietic stem cells might be of interest in ovarian cancer. The results in one patient also suggest that DLI may be an effective immunotherapy, although doses and timing need to be determined. The number of cases presented is small, however, and clinical experience on a larger scale will be required to determine the real clinical efficacy of graft versus cancerous ovarian cells. PMID- 12132050 TI - Selective growth advantage of wild-type lymphocytes in X-linked SCID recipients. AB - The cytokine receptor common gamma chain (gamma c) plays a pivotal role in multiple interleukin signaling, and gamma c gene mutations cause an X-linked form of SCID (X-SCID). Recently, gamma c gene transfer into the autologous X-SCID BM achieved appreciable lymphocyte reconstitution, contrasting with the limited success in previous gene therapy trials targeting hematopoietic stem cells. To understand the mechanisms underlying this success, we examined the repopulating potential of the wild-type (WT) BM cells using an X-SCID mouse model. Limited numbers of WT cells were infused into non-ablated WT and X-SCID hosts. Whereas no appreciable engraftment was observed in WT recipients, donor-derived lymphocytes repopulated well in X-SCID, reaching 37% (10(6)cells given) and 53% (10(7) cells given) of the normal control value 5 months post BMT. A lineage analysis showed a predominance of the donor-derived lymphocytes (CD4(+) T, CD8(+) T, B and NK cells) in X-SCID while the donor-derived granulocytes and monocytes engrafted poorly. These results showed a selective advantage of WT cells in X-SCID, and that the advantage was restricted to lymphocytes. In human gene therapy for X SCID, an analogous growth advantage would greatly enhance the repopulation of lymphocytes derived from a very small number of gamma c gene-supplemented precursors. PMID- 12132049 TI - Pre-transplant positron emission tomography (PET) using fluorine-18-fluoro deoxyglucose (FDG) predicts outcome in patients treated with high-dose chemotherapy and autologous stem cell transplantation for non-Hodgkin's lymphoma. AB - We investigated the predictive value of sequential FDG PET before and after high dose chemotherapy (HDT) and autologous stem cell transplantation (ASCT) in 24 patients suffering from non-Hodgkin's lymphoma (NHL). FDG PET was performed at baseline, after three cycles of induction therapy, before and after HDT with ASCT. Response assessment from sequential PET scans using standardized uptake values (SUV) was available in 22 patients at the time of transplantation. Partial metabolic response (PMR) was defined as a >25% decrease of SUV between successive PET scans [corrected]. Six of seven patients who did not achieve a PMR after complete induction therapy developed lymphoma progression, while 10 of 15 patients with complete metabolic response (CMR) or PMR remained in continuous remission. Four of seven patients with less than PMR after induction therapy died vs two of 15 patients with CMR/PMR. Median progression-free and overall survival of patients with less than PMR after HDT and ASCT was 9 and 29 months, respectively. In contrast, neither conventional re-staging nor the International Prognostic Index were predictive. These data suggest that sequential quantitative PET imaging does enlarge the concept of chemosensitivity used to select patients with high-risk NHL for HDT and ASCT or to route them to alternative treatments. PMID- 12132051 TI - Outcomes of antiemetic prophylaxis in children undergoing bone marrow transplantation. AB - A prospective survey of the control of acute and delayed antineoplastic and radiation-induced nausea and vomiting was undertaken in children undergoing bone marrow transplantation (BMT) at The Hospital for Sick Children. Prior administration of antineoplastic agents or irradiation, presence of anticipatory nausea or vomiting prior to starting the conditioning regimen, antiemetic use within 24 h of conditioning, the prescribed antineoplastic and/or radiation ablative regimen, and prescribed antiemetic regimens were recorded. Emetic episodes, dietary intake, administration of conditioning agents and antiemetics, and adverse effects were monitored on each day of the conditioning regimen and for 96 h thereafter. Children older than 3 years of age assessed their nausea on each study day. Twenty-five children were followed for 258 patient days. Children did not vomit or retch on 73% and 43% of patient days, in the acute and delayed phases, respectively. Nausea data were evaluable for 21 children on 200 patient days. Nausea was absent on 55% and 26% of patient days in the acute and delayed phases, respectively. Five children never had an emetic episode during the entire study period. One child was completely free from nausea and vomiting throughout the study period. Antineoplastic and radiation-induced nausea and vomiting can be successfully prevented in the majority of children undergoing BMT. However, effective treatment strategies must be developed in the event of antiemetic failure and for effective prophylaxis in children who cannot tolerate dexamethasone. PMID- 12132052 TI - Energy expenditure in children undergoing hematopoietic stem cell transplantation. AB - Prior studies suggest that patients undergoing hematopoietic stem cell transplantation (HSCT) for malignancy have nutritional needs that are greater than their estimated needs. To determine whether energy estimation equations accurately predict energy expenditure of pediatric patients undergoing HSCT, we prospectively compared the estimated energy expenditure (EEE) and measured energy expenditure (MEE) of 40 patients at four time-points. We also investigated whether energy requirements changed during the transplant period. MEE was determined by indirect calorimetry. Data from 34 patients (autologous HSCT = 10, allogeneic HSCT = 24) were sufficient for analysis. The World Health Organization equation adequately approximated MEE only on day 14 after HSCT. At all other time points, measured energy expenditure was significantly less than estimated energy expenditure obtained by using the WHO equation (applicable to all patients), the Seashore equation (for patients <15 years of age; n = 19), or the Harris-Benedict equation (for patients > or =15 years of age; n = 15). The median measured energy expenditure varied significantly over the study period and was greatest on day 14 after HSCT. Until accurate equations have been identified for estimating these patients' needs, the use of indirect calorimetry may be medically warranted. PMID- 12132053 TI - Septic shock and multiple organ failure after hematopoietic stem cell transplantation: treatment with recombinant human activated protein C. AB - Severe sepsis with multiple organ failure after hematopoietic stem cell transplantation (HSCT) results in extremely high morbidity and mortality. Recent studies have highlighted the importance of sepsis-induced activation of the coagulation system in the pathophysiology of severe sepsis. Activated protein C is an important modulator of coagulation and inflammatory derangements during severe sepsis. Low levels of protein C occur in severe sepsis and are predictive of poor outcome. Recombinant human activated protein C (drotrecogin alfa (activated)) was recently approved by the Food and Drug Administration (FDA) for severe sepsis. The phase III trial that resulted in the approval of this agent, however, enrolled a general sepsis population and excluded patients undergoing HSCT. We report a case of fulminant septic shock and multiple organ failure after HSCT that was treated with drotrecogin alfa (activated) in addition to standard therapy, and recovered. The high mortality rates of patients who develop severe sepsis after HSCT demand that new avenues of treatment be considered for this very high-risk patient population. This case illustrates the potential application of a novel therapeutic approach. Clinical trials are warranted to further investigate the safety and efficacy of drotrecogin alfa (activated) in patients with severe sepsis after HSCT. PMID- 12132056 TI - The future of myeloscopy. PMID- 12132057 TI - Targeted methylprednisolone acetate/hyaluronidase/clonidine injection after diagnostic epiduroscopy for chronic sciatica: a prospective, 1-year follow-up study. AB - BACKGROUND AND OBJECTIVES: It has been claimed that epiduroscopy offers an ideal combination of diagnostic and therapeutic interventions in one session. We prospectively evaluated whether abnormalities at the lumbar level as diagnosed by magnetic resonance imaging (MRI) are confirmed by epiduroscopy, and assessed if targeted epidural injection of medication alleviates sciatic pain. METHODS: A flexible, 0.9-mm fiberoptic endoscope was introduced through a disposable steering shaft into the caudal epidural space and advanced until the targeted spinal nerve was identified. Adhesions were mechanically mobilized under direct vision, and a mixture of 120 mg methylprednisolone acetate, 600 IU hyaluronidase, and 150 microg clonidine was applied locally. Pain scores were measured by visual analog scale (VAS) and global subjective efficacy rating. RESULTS: Nineteen of 20 patients studied showed adhesions via epiduroscopy. In 8 patients, 6 of whom had never undergone surgery, these were not detected with earlier magnetic resonance imaging. Six patients showed concomitant signs of active root inflammation. Of 20 patients treated with a targeted epidural injection, 11 patients (55%) experienced significant pain relief at 3 months. This was maintained at 6, 9, and 12 months for 8 (40%), 7 (35%), and 7 (35%) patients, respectively. Mean VAS at 3 months was significantly reduced (n = 20; DeltaVAS = 3.55; P <.0001), and this persisted at 12 months (DeltaVAS = 1.99, P =.0073). CONCLUSIONS: Epiduroscopy is of value in the diagnosis of spinal root pathology. In sciatica, adhesions unreported by MRI can be identified. Targeted epidural medication, administered near the compromised spinal nerve, results in substantial and prolonged pain relief. PMID- 12132058 TI - The epidural fibrous sheath: a guide for the replacement of a spinal cord stimulation electrode. AB - BACKGROUND AND OBJECTIVE: During spinal cord stimulation there is sometimes a need to replace defective leads. Percutaneous lead replacement by recannulating the epidural space and "steering" the new lead to the prior location is sometimes very difficult, resulting in diminished analgesia. Since fibrous deposits are known to form around epidural catheters and epidural obstructions have been noted with other techniques, we have inserted the new lead through the well-dissected opening in the interspinal ligament. We will report the results of our case series. METHODS: In 11 patients with lead malfunction we reinserted a new electrode into the epidural space by first withdrawing the lead with one hand and inserting the new one through the interspinal ligament with the other. In using this method, we found we could position the new electrode almost identically to the first. In only 3 patients did we experience difficulty in identifying the opening for the insertion. In the successfully cannulated patients identical stimulation parameters and paresthesia areas were obtained. By experimentally injecting contrast dye through an epidural catheter inserted into the interspinal opening and epidural pathway, we could visualize a thin dense line representing the fibrous sheath. CONCLUSION: Foreign bodies in the epidural space lead to fibrous deposits. Spinal cord stimulation, when those deposits form a sheath, the sheath is useful for lead revision. The procedure, if meticulously performed, has a high success rate. PMID- 12132059 TI - Expression of neuron-associated tumor necrosis factor alpha in the brain is increased during persistent pain. AB - BACKGROUND AND OBJECTIVES: Evidence implicates the pleiotropic cytokine tumor necrosis factor alpha (TNFalpha) in the pathogenesis of persistent pain. The present study employs a chronic constriction injury (CCI) model of neuropathic pain to examine TNFalpha production in the central nervous system (CNS) and in the periphery in this pain model. METHODS: CCI-induced hyperalgesia is assessed by measuring the nociceptive threshold using the hot-plate test. The development of hyperalgesia is correlated to levels of TNFalpha by assessing: bioactive TNFalpha in homogenates of sciatic nerves, cervical spinal cord, thoracolumbar spinal cord, as well as in plasma using the WEHI-13 variant cytotoxicity bioassay; and mRNA for TNFalpha in sections of locus coeruleus by in situ hybridization. RESULTS: We have previously demonstrated that TNFalpha bioactivity in the region of the brainstem containing the locus coeruleus is increased concurrent with the development of hyperalgesia, returning to baseline values by day 14, when hyperalgesia has ceased. Constitutive levels of TNFalpha are demonstrated in the plasma, sciatic nerves, and cervical and thoracolumbar spinal cord of control rats, sham-operated rats, and rats undergoing CCI. Levels of TNFalpha are significantly elevated in the injured sciatic nerve by day 8 postligature placement, concurrent with maximal hyperalgesia, and remain elevated when hyperalgesia has abated at day 14 postligature placement. Additionally, TNFalpha activity is increased in the thoracolumbar region of the spinal cord by day 4 postligature placement and remains elevated during hyperalgesia (day 8), as well as after hyperalgesia has dissipated (day 14). The increase in TNFalpha expression is specific to discrete regions of the CNS, rather than being the result of a systemic inflammatory response, since TNFalpha bioactivity in plasma is, in fact, decreased in rats undergoing CCI. Additionally, accumulation of mRNA specific for TNFalpha is significantly increased in neurons within a region of the brain containing the locus coeruleus at days 2, 8, and 14 postligature placement, contemporaneous with the development of hyperalgesia. CONCLUSIONS: The increases in TNFalpha within regions of the brain and spinal cord that are associated with adrenergic neuron function, as well as with modulation of pain perception, and the time course and distribution of the increases in TNFalpha accumulation support a neuromodulatory role for TNFalpha within the CNS in the development and maintenance of neuropathic pain. PMID- 12132060 TI - Patient-controlled thoracic epidural infusion with ropivacaine 0.375% provides comparable pain relief as bupivacaine 0.125% plus sufentanil after major abdominal gynecologic tumor surgery. AB - BACKGROUND AND OBJECTIVES: We tested the hypothesis that an opioid-free local anesthetic alone is able to provide comparable analgesia to the opioid supplemented epidural application of local anesthetics using thoracic epidural catheters after major abdominal surgery. METHODS: In a prospective, randomized, and double-blind study, we have compared the analgesic efficacy and side effects of ropivacaine 0.375% (group R) versus bupivacaine 0.125% in combination with sufentanil 0.5 microg/mL(-1) (group B/S) via a thoracic epidural catheter for a duration of 96 hours after major abdominal surgery in 30 gynecologic tumor patients. Piritramide was given for breakthrough pain. Assessments were performed every 12 hours after start of the epidural infusion using continuous (first 24 hours) and patient-controlled epidural analgesia (PCEA) (24 to 96 hours). RESULTS: No differences were seen in demographic and perioperative data. Dynamic pain scores (visual analog scale [VAS] values) were comparable between groups during mobilization (group R v group B/S: 24 hours: 40 +/- 30 v 36 +/- 14, P =.9; 48 hours: 46 +/- 33 v 42 +/- 25, P =.93; 72 hours: 42 +/- 24 v 48 +/- 26, P =.78; 96 hours: 42 +/- 25 v 29 +/- 28, P =.49) and on coughing during the whole study period. Hemodynamics, intensity of motor block (Bromage scale), and side effects like nausea, vomiting, pruritus, and bladder disfunction also did not differ between groups. CONCLUSION: The present study shows that thoracic epidural infusion of ropivacaine 0.375% provides comparable pain relief and incidence of side effects after major abdominal gynecologic surgery as bupivacaine 0.125% in combination with 0.5 microg/mL(-1) sufentanil and may therefore represent an alternative in epidural pain management. PMID- 12132061 TI - Epinephrine is not a useful addition to intrathecal fentanyl or fentanyl bupivacaine for labor analgesia. AB - BACKGROUND AND OBJECTIVES: Intrathecal fentanyl provides effective labor analgesia for a limited time with frequent side effects. We evaluated the effects of adding epinephrine to intrathecal fentanyl with and without bupivacaine. METHODS: Eighty healthy, term, nulliparous parturients with cervical dilation of 5 cm or less received combined spinal-epidural (CSE) analgesia. Subjects were randomized in a double-blind fashion to 1 of 4 intrathecal solutions containing fentanyl 35 microg with either saline (F); bupivacaine 2.5 mg + saline (FB); bupivacaine 2.5 mg + epinephrine 100 microg (FBE); or epinephrine 100 microg + saline (FE). Patients were evaluated for visual analog pain score, duration of spinal analgesia (time until patient request for additional analgesia), nausea/vomiting, pruritus, sensory and motor block, maternal blood pressure, and fetal heart rate (FHR). RESULTS: Intrathecal bupivacaine significantly prolonged fentanyl analgesia with or without epinephrine (P =.018), but epinephrine did not significantly prolong the duration of fentanyl alone or with bupivacaine (F, 92 +/- 39 minutes; FB, 125 +/- 31 minutes; FBE, 134 +/- 42 minutes; and FE, 117 +/- 48 minutes). Intrathecal epinephrine was associated with a higher incidence of severe nausea (P =.001), and the FBE group had more lower extremity weakness (P =.047). There was no difference in the incidence of severe pruritus, FHR deceleration, or delivery outcome between the groups. CONCLUSIONS: These results suggest that intrathecal epinephrine does not prolong the duration of fentanyl or fentanyl with bupivacaine for labor analgesia in nulliparous parturients. Additionally, intrathecal epinephrine did not decrease the incidence of side effects and therefore cannot be recommended. PMID- 12132062 TI - Nefopam and tramadol for the prevention of shivering during neuraxial anesthesia. AB - BACKGROUND AND OBJECTIVES: In patients undergoing neuraxial anesthesia, heat loss and core-to-peripheral redistribution of body heat causes the core temperature to decrease. The shivering threshold is therefore reached soon, and more shivering is required to prevent further hypothermia. Because shivering has deleterious metabolic and cardiovascular effects, it should ideally be prevented by pharmacologic or other means. We evaluated the usefulness of intravenous (IV) nefopam and tramadol in preventing and reducing the severity of shivering in patients undergoing neuraxial anesthesia for orthopedic surgery. METHODS: Ninety patients, scheduled for neuraxial anesthesia (epidural or subarachnoid) for lower limb orthopedic surgery, were prospectively enrolled. Patients were randomly assigned to 1 of 3 groups. Immediately before neuraxial anesthesia, 30 patients received 0.15 mg/kg(-1) IV nefopam in 10 mL saline, 30 patients received 0.5 mg/kg(-1) IV tramadol in 10 mL saline, and a control group of 30 patients received 10 mL IV saline. Neuraxial anesthesia was induced at the L3-L4 or L4-L5 interspaces with 1 mg/kg(-1) mepivacaine for epidural anesthesia and 0.2 mg/kg( 1) for subarachnoid anesthesia. An investigator blinded to the antishivering drug injected recorded the frequency and degree of shivering. RESULTS: The overall frequency and the intensity of shivering was significantly lower in patients treated with nefopam than in those treated with tramadol or placebo (P <.05 and P <.01) and in patients treated with tramadol than in those treated with placebo (P <.05). CONCLUSIONS: As a pharmacologic means of preventing shivering in patients undergoing neuraxial anesthesia, nefopam may hold the greatest promise. PMID- 12132063 TI - Animal pain models. PMID- 12132064 TI - Brachial plexus anesthesia: essentials of our current understanding. PMID- 12132065 TI - Mechanical effects of leg position on vertebral structures examined by magnetic resonance imaging. AB - BACKGROUND AND OBJECTIVES: Leg manipulation has been postulated to affect spinal curvature and position of the cauda equina within the dural sac. However, no evidence of such mechanical effects has been shown in living subjects. We used magnetic resonance imaging to evaluate the mechanical effects of leg position on these 2 parameters. METHODS: Sagittal and axial magnetic resonance images of the lumbosacral vertebral canal were obtained in 5 healthy, female volunteers with the subject in the supine position with knees straight, knees slightly flexed, and knees fully flexed. RESULTS: In the straight leg position, physiologic lumbar lordosis was evident in all subjects on midline sagittal slices, whereas lumbar lordosis disappeared in the fully flexed leg position. On the axial slices the cauda equina moved ventrally within the dural sac in all subjects in the fully flexed leg position. In 1 of the 5 subjects the cauda equina moved ventrally and also separated completely into right and left parts. CONCLUSIONS: Our findings indicate that 2 potential factors, flattening of the lumbar lordosis and some added tension on the lumbosacral nerve roots, may contribute to postoperative back and leg aching after spinal anesthesia in the lithotomy position. PMID- 12132066 TI - Seizures after epidural blood patch with undiagnosed subdural hematoma. AB - OBJECTIVE: We present a case of new-onset seizures in a parturient who received an epidural blood patch (EBP) in the presence of an undiagnosed cerebral subdural hematoma. We review the relevant literature and examine the implications for management. CASE REPORT: A 33-year-old parturient developed symptoms of postdural puncture headache 16 hours after initiation of epidural analgesia for labor. On the third postpartum day, she likely had an unwitnessed and unrecognized seizure. Presenting to hospital with headache and fatigue, she received an EBP. Forty-five minutes after the EBP, she had a generalized seizure followed by another 2 hours later. Imaging revealed a small cranial subdural hematoma. She had no further seizures, her headache improved, and she was discharged home on postpartum day 5. CONCLUSIONS: We believe that the subdural hematoma and not the EBP was the cause of the seizures. Earlier recognition of confusion and fatigue as a postictal state may have led to earlier diagnosis of the hematoma. Our experience suggests that EBP be avoided in the setting of subdural hematoma. PMID- 12132067 TI - In defense of radiofrequency neurotomy. PMID- 12132070 TI - Axillary block by double-, triple-, or quadruple-nerve stimulation. PMID- 12132072 TI - Use of an ultrasound doppler flowmeter for occipital nerve block. PMID- 12132073 TI - Horner's syndrome following epidural anesthesia with ropivacaine for cesarean delivery. PMID- 12132074 TI - Continuous brachial plexus block at the cervical level using a posterior approach in the management of neuropathic cancer pain. PMID- 12132076 TI - Vibration sensation testing over the medial malleolus. PMID- 12132078 TI - Advances in composite tissue allograft transplantation as related to the hand and upper extremity. AB - The clinical transplantation of composite tissue allografts (CTA) such as human hand or larynx is stimulating discussions among surgeons at national and international forums on the indications, ethical aspects, toxic effects of immunosuppression, and functional results of the first reported cases of unilateral and bilateral hand transplantation. This Clinical Perspective article presents the latest advances in clinical and experimental research related to the field of CTAs. The article presents the historic aspects of CTA, a broad view of the current state of composite tissue transplantation, the mechanism of allograft rejection, current experimental and clinical protocols, and, finally, the future prospects of the standard use of CTAs. It is clear that there is a substantial demand for routine use of CTAs but the treatment protocols need to be optimized and the functional outcomes need to be improved. PMID- 12132079 TI - Staged flexor tendon reconstruction fingertip to palm. AB - Thirty-five fingertip to palm staged flexor tendon reconstructions were performed between 1971 and 1998. Tendon injuries involved 5 avulsions and 30 lacerations, 9 of which had failed primary tenorrhaphies. Follow-up evaluation averaged 30 months. Thirteen patients had total active motion (TAM) of > or =220 degrees (excellent), 11 patients had TAM of 200 degrees to 219 degrees (good), 7 patients had TAM of 180 degrees to 199 degrees (fair), and 4 patients had TAM of <180 degrees (poor). Less favorable results occurred in those with a >1-year interval between injury and stage I, in those with a >6-month interval between stage I and stage II, and in those with a higher injury severity classification. Fingertip to palm staged flexor tendon reconstruction produced 69% good to excellent results. This technique allows the use of the injured digit profundus as the motor, preserves lumbrical function, and requires less tendon graft length (palmaris longus usually suffices). PMID- 12132080 TI - Management of flexor tendon sheath ganglions: a cost analysis. AB - The purpose of this study was to determine success rates of 1 or more aspirations on flexor tendon sheath ganglions compared with surgical excision and to determine what treatment method is most cost-effective. Data were collected from documented history and physical examinations, operative reports, billing records, and telephone interviews. Of the 259 patients coded as having flexor tendon sheath ganglions, 175 met the inclusion criteria. In addition, 2001 Medicare assigned relative value units and fees were used to calculate the most cost effective treatment. Of the 141 patients treated with aspiration, 66% exhibited no recurrence after 2 consecutive treatments. Thirteen of 14 patients (93%) who had ganglions excised without prior aspiration showed no recurrence. All 29 patients who had excision after 1 (n = 24) or 2 (n = 5) failed aspirations were cured. Because few patients require excision after 2 aspirations, the most cost effective treatment for recurrent flexor tendon sheath ganglions is 2 aspirations before excision. PMID- 12132081 TI - Flexor tendon rupture caused by gout: a case report. AB - We present a case of primary gouty infiltration of flexor tendons in the hand, causing rupture of both flexor digitorum superficialis and profundus tendons in a single digit. The patient was managed by a single-stage reconstruction of the less involved flexor digitorum superficialis tendon using a segment of the proximal stump of flexor digitorum profundus tendon as a bridge graft. This uncommon etiology of tendon rupture should be considered in all patients with a history of gout presenting with tendon insufficiency. PMID- 12132082 TI - Repair of flexor digitorum profundus tendon avulsions from bone: an ex vivo biomechanical analysis. AB - Avulsions or distal transections of the flexor digitorum profundus tendon are typically repaired by direct suture of the tendon stump to the distal phalanx. The optimal repair technique to withstand in vivo rehabilitation forces is unknown. Our objective was to determine the time-zero tensile mechanical properties of 4-strand tendon-bone repair site constructs performed with 3-0 and 4-0 sutures and with modified Kessler and modified Becker grasping techniques. We hypothesized that the 3-0 modified Becker grasping suture technique not described previously for the reattachment of tendon to bone would show improved biomechanical properties compared with the 4-0 or modified Kessler techniques. All modified Kessler repairs failed by suture pullout from the tendon, whereas all modified Becker repairs failed by rupture of the suture at the tendon-bone junction. Although the 3-0 modified Becker repair group showed greater ultimate force then the other groups (p <.01), tendon-bone gap observed did not differ markedly between Becker or Kessler groups. Neither suture caliber nor repair technique had a notable effect on strain at 20-N force, suggesting that early gap formation at the tendon-bone repair site may occur regardless of technique. PMID- 12132083 TI - The strength of distal fixation of flexor digitorum profundus tendon grafts in human cadavers. AB - Early active motion limits adhesion formation and thus improves functional performance after tendon grafting. The early strength of distal fixation is critical to successful tendon grafting. We describe a new Y-tunnel technique of distal fixation and compare it with 2 established methods, the Pulvertaft transverse tunnel and the Bunnell button over the fingernail techniques, in a human cadaver model to determine which is the strongest method. Hands with a grafted tendon were rigidly mounted on an anatomic tensiometer testing apparatus and loaded to failure. Mean load to failure (newtons +/- 95% confidence intervals) of the Y-tunnel technique (155.2 +/- 29.4) was greater than those for the Pulvertaft (100.2 +/- 13.2) and Bunnell (57.1 +/- 4.7) techniques. Two-way analysis of variance showed significant differences, and the Bonferroni multiple pairwise comparison test showed that all 3 intergroup comparisons were statistically significant. These results indicate marked improvement in immediate strength with the Y-tunnel technique and lay the groundwork for further studies using a healing tendon model. PMID- 12132084 TI - Characterization of the tensile properties and histologic/biochemical changes in normal chicken tendon at the site of suture insertion. AB - The hypothesis of this study is that inserting a grasping suture into tendon tissue causes adverse changes at the suture-to-tendon interface and thus compromises the outcome of tendon repair. This study characterized the histologic, biomechanical, and biochemical effects that 2 types of grasping sutures, Kessler and Savage, produced on normal chicken flexor tendon in the digital flexor sheath. Variables caused by a healing response were avoided by not cutting the tendon. Findings included a reduction in ultimate tensile strength beginning soon after the suture was inserted, cell proliferation, lysis of types I and III collagen by collagenase, and presence of gelatinase. No histologic or biochemical differences were detected between the 2 suture patterns. These findings are presumed to be adverse and suggest an additional reason why the ultimate tensile strength of repaired tendon may be too low to allow patients to move tendons actively without a high risk for causing gap formation or rupture in the early weeks of healing. PMID- 12132085 TI - Flexor tendon healing in vitro: effects of TGF-beta on tendon cell collagen production. AB - Flexor tendon healing is complicated by adhesions to the surrounding sheath. Transforming growth factor beta (TGF-beta) is a cytokine with numerous activities related to wound healing. We examined the effects of TGF-beta-1, -2 and -3 on tendon cell proliferation and collagen production. Three separate cell lines- sheath fibroblasts, epitenon and endotenon tenocytes--were isolated from rabbit flexor tendons and cultured separately. Cell culture media was supplemented with 1 or 5 ng/mL of TGF-beta-1, -2, or -3. Cell number and collagen I and III production were measured and compared with unsupplemented control cultures. The addition of TGF-beta to cell culture media resulted in a decrease in cell number in all 3 lines that did not reach statistical significance. There was a significant increase (p <.05) in collagen I and III production with the addition of all 3 TGF-beta isoforms. Modulation of TGF-beta production may provide a mechanism to modulate adhesion formation clinically. PMID- 12132086 TI - Biomechanical studies of 3 different 6-strand flexor tendon repair techniques. AB - We investigated the gap formation and ultimate strength, energy to failure, and patterns of failure of tendon repairs with the modified Savage, Lim, and Tang methods. Fifty-four fresh-frozen flexor digitorum profundus tendons were assigned to 3 groups and repaired with one of the previously mentioned methods. Nine tendons from each group were pulled to failure in a tensile testing machine when they were subjected to a linear load. The 2-mm gap formation force of the tendons was 37.4 N for the modified Savage, 44.5 N for the Tang, and 40.2 N for the Lim method. Ultimate strength was 57.8 N for the modified Savage, 60.2 N for the Tang, and 51.3 N for the Lim method. Statistically, the gap formation force was significantly higher in the Tang method than in the modified Savage and the Lim methods. Ultimate strength of the modified Savage and Tang methods was similar and significantly higher than that of the Lim method. The rest of the tendons were subjected to angular tension by placement of the tendons against a pulley. The tests show that resistance of the modified Savage and Tang methods to angular tension was significantly greater than that of the Lim method. The results indicate that the resistance to failure and failure modes of the repairs vary according to number of locking junctions with the tendon, location, and orientation of the sutures regardless of an equal number of strands across the repair site. We conclude that the modified Savage and Tang methods have greater tensile strength than the Lim method and may more effectively resist linear and angular tension generated by postoperative tendon motion. PMID- 12132087 TI - The pulley system of the thumb: anatomic and biomechanical study. AB - To examine the precise conformation of the annular and oblique pulleys of the thumb flexor sheath, anatomic dissections were undertaken on 14 hands. In all specimens a distinct pulley was found between the A1 and oblique pulleys. This is named the variable annular pulley or Av pulley. There appear to be 3 discrete forms of this pulley designated type I to III. The biomechanical study was done on 8 limbs by using linear strain transduction techniques. The analysis showed that the strain in the oblique pulley was greater in extension than in flexion of the thumb. This statement remains true even after division of either the A1 or Av pulley and after section of both pulleys. The oblique pulley does not prevent bowstringing of the flexor pollicis longus when A1 and Av pulleys have been sectioned. These studies challenge current concepts of both the anatomy and mechanics of the thumb pulley system with implications for clinical procedures such as trigger thumb release and pulley reconstruction. PMID- 12132088 TI - Posttraumatic volar tendon subluxation out of the first extensor compartment: a case report. AB - Symptomatic volar subluxation of the abductor pollicis longus and the extensor pollicis brevis tendons developed in a 29-year-old man after a sprain that occurred with the wrist in flexion and ulnar deviation. The extensor retinaculum, which forms the extensor compartment, was partially avulsed from its insertion on the radius. Palmar abduction and extension of the thumb with the wrist flexed produced subluxation of the tendons over the volar side of the radius ridge where the retinaculum forming the first extensor compartment attached. Nonoperative treatment including steroid injection and splinting was ineffective. Surgery was performed to reconstruct a new tendon restraint with part of the extensor retinaculum. PMID- 12132090 TI - Predictors of carpal tunnel syndrome: an 11-year study of industrial workers. AB - In 1984 we initiated a study of factors associated with carpal tunnel syndrome (CTS) in industrial workers by using a case definition based on both symptoms and electrophysiologic findings. Medical history, lifestyle factors, and symptoms were assessed by interview, and electrodiagnostic studies were used to measure median nerve function. Job tasks were classified by both interview and direct observation of work activities. Follow-up evaluations were conducted in 1989 and 1994-1995. The analytic sample consisted of 111 women and 145 men free of CTS in 1984 who were examined at both subsequent contact points. In logistic regression analyses, greater age, female gender, relative overweight, cigarette smoking, and vibrations associated with job tasks were found to significantly increase risk for dominant-hand CTS, whereas presence of an endocrine disorder was marginally related to reduced risk for CTS. These findings were generally similar when analyzed separately for men and women. Similar to other chronic noninfectious diseases, personal factors may play an important role in determining risk for CTS. PMID- 12132089 TI - Interosseous-lumbrical adhesions of the hand: contribution of magnetic resonance imaging to diagnosis and treatment planning. AB - Previous investigators have not found magnetic resonance imaging (MRI) to be helpful in the diagnosis of interosseous-lumbrical tendon adhesions of the hand. We present 2 cases in which preoperative magnetic resonance images correlated with the clinical diagnosis and intraoperative finding of adhesions between the interosseous-lumbrical tendons at the level of the metacarpal head. Because there are no specific signs for the diagnosis, no palpable lesion, and no specific provocative test, the MRI acted to confirm the diagnosis in the presence of vague, nonspecific symptoms and provide objective evidence of the source of the patient's discomfort. This finding preserved surgery as a definite therapeutic rather than exploratory procedure, raising the confidence level of the operating surgeon as well as the patient. We believe that an MRI is beneficial in narrowing the differential diagnosis of interosseous-lumbrical adhesions, especially in difficult cases in which other underlying pathology may exist. PMID- 12132091 TI - Outcomes of multiple microvascular toe transfers for reconstruction in 2 patients with digitless hands: 2- and 4-year follow-up case reports. AB - Two farmers who sustained amputations of all digits in the dominant hand had hand reconstructions with 3 toes from both feet. Both patients were monitored with hand function tests and validated outcomes questionnaires (Michigan Hand Outcomes Questionnaire and Short Form-36). Long-term follow-up data showed that both patients were able to use the reconstructed hands for activities of daily living and heavy manual farm labor. Until the immunologic problems associated with hand transplantation are solved, multiple toe transfer for total hand reconstruction is a reliable surgical treatment for these patients. PMID- 12132092 TI - Physiolysis for correction of clinodactyly in children. AB - Congenital clinodactyly is a lateral deviation of a finger frequently caused by an abnormal middle phalanx (trapezoidal or triangular delta phalanx). The physis extends longitudinally on the short side of the middle phalanx. Resection of the abnormal longitudinal physis and fat graft interposition (physiolysis) has been reported to correct the lateral finger deviation in growing children. We reviewed 35 fingers that had a physiolysis procedure. The age at surgery varied from 2.9 to 10.9 years (mean, 6.6 y), the preoperative angulation was 20 degrees to 29 degrees in 9 fingers, 30 degrees to 39 degrees in 16 fingers, and 40 degrees or more in 10 fingers. Thirty-one fingers presented a trapezoidal phalanx and 4 fingers a triangular phalanx. Ten fingers had a second surgery using the same procedure. Follow-up time ranged from 1.2 to 5.3 years (mean, 3.2 y). After 1 procedure the degree of correction varied from 0 degrees to 30 degrees (mean, 11.1 degrees). The residual angulation was <15 degrees in 8 fingers, 15 degrees to 19 degrees in 4 fingers, 20 degrees to 29 degrees in 15 fingers, 30 degrees to 39 degrees in 6 fingers, and > or =40 degrees in 2 fingers. Correction obtained in the trapezoidal phalanges was better (mean 12.5 degrees) than in the triangular phalanges (mean, 2.8 degrees). The fingers presenting a more severe preoperative deformity (angulation > or =40 degrees) had a better correction (mean, 20 degrees) compared with fingers with a lesser deformity (mean, 7.5 degrees). The correction was also better in children who had surgery before 6 years of age (mean, 17.9 degrees) compared with older children (mean, 6.5 degrees). A second physiolysis procedure was not beneficial in 8 of 10 fingers reoperated and 2 premature fusions of the proximal transverse physis were found among these 10 fingers. There were no other complications. The physiolysis procedure is simple and effective, particularly in children presenting with a trapezoidal phalanx who have surgery before 6 years of age. PMID- 12132093 TI - Bilateral biceps-to-triceps transfer to salvage failed bilateral deltoid-to triceps transfer: a case report. AB - The treatment of failed deltoid-to-triceps tendon transfers for elbow extension in persons with tetraplegia is a difficult problem. This situation was managed in a 19-year-old man by bilateral biceps-to-triceps tendon transfers. Successful restoration of elbow extension resulted in an increase in the available workspace and enhanced ability to perform activities of daily living. PMID- 12132094 TI - Malignant epithelioid hemangioendothelioma presenting as an aneurysm of the superficial palmar arch: a case report. AB - A patient with a malignant epithelioid hemangioendothelioma that presented as an aneurysm of the superficial palmar arch and third common digital artery had complete excision of the aneurysm and vascular reanastomosis. After pathologic diagnosis and surgical staging she received local irradiation to the hand. This case suggests that an aneurysm in the hand may develop in the setting of an underlying malignancy of blood vessel. When aneurysms in the hand require surgical intervention, we recommend that resection be complete, pathologic examination undertaken, and marginal tissue well-labeled to allow precise pathologic examination. PMID- 12132095 TI - Long-term results after repair and augmentation ligamentoplasty of rotatory subluxation of the scaphoid. AB - Twenty-two patients had scapholunate ligament repairs combined with a new augmentation ligamentoplasty for chronic scapholunate dissociation. All were evaluated by physical and radiologic examination after a mean postoperative follow-up period of 63 months (range, 12-134 mo). According to Green and O'Brien and Johnson and Carrera scores 5/8 had excellent, 13/12 good, and 4/2 fair results. Thirteen were free of pain; 6 had mild pain and 3 had moderate pain. Nineteen returned to their original occupation. There was an average loss of 10 degrees of flexion, 9 degrees of extension, and 11% of grip force compared with the opposite wrist. Radiologic examination showed an average decrease of 12 degrees of the scapholunate and 10 degrees of the radiolunate angles compared with the levels before surgery. No signs of degenerative osteoarthritis were found in 16 (73%) cases. Five wrists showed a distinct pattern of midcarpal degeneration correlating with notable dorsal intercalated segment instability after surgery, and 2 cases had signs of radioscaphoid degeneration. PMID- 12132096 TI - Treatment of scaphoid nonunions: quantitative meta-analysis of the literature. AB - We conducted a systematic quantitative meta-review of the literature to provide evidence-based suggestions for the treatment of scaphoid nonunion. This search identified 1,121 articles of which 36 met eligibility requirements. In unstable nonunions, screw fixation with grafting at 94% union was superior to K-wires and wedge grafting (77% union). Immediate mobilization versus 6 weeks or more of casting showed the same union rate of 74%. For avascular necrosis of the proximal fragment, union was achieved in 88% of those patients with a vascularized graft versus 47% with screw and wedge fixation. These results suggest that established unstable nonunions should be treated with screw fixation and wedge grafting. There is not evidence supporting the need for postoperative immobilization in patients with solid screw fixation. A vascularized graft may be preferable for patients with avascular necrosis of the proximal fragment or with a previously failed surgery. PMID- 12132097 TI - Scaphocapitolunate arthrodesis for idiopathic avascular necrosis of the capitate: a case report. AB - Idiopathic avascular necrosis of the capitate is a disorder of unknown etiology that causes wrist pain and limitation of function. Many different treatments have been presented in the literature with varying clinical results and levels of patient satisfaction. We present a case of a 23-year-old woman successfully treated with scaphocapitolunate arthrodesis. PMID- 12132098 TI - Surgical treatment of intra-articular fractures of the trapezium. AB - Eleven patients with intra-articular fractures of the trapezium were evaluated after surgical treatment with a mean follow-up time of 47 months (range, 25-80 mo). There were 6 vertical split and 5 comminuted fractures. All fractures involved high-energy trauma in men. Five resulted from motorcycle accidents. An associated Bennett's fracture occurred in 4 patients. Three fractures went undiagnosed at the time of initial evaluation. At late follow-up evaluation, 8 of 11 patients experienced some pain, mainly at the trapeziometacarpal articulation. There was no statistical difference in thumb motion, wrist motion, or grip and pinch strength between the affected and unaffected extremity. Radiographs revealed degenerative changes at the trapeziometacarpal articulation in 5 of 11 fractures. No patient was disabled and no patient changed occupation as a result of the hand injury. Based on the good results obtained with surgical intervention we advocate open reduction and internal fixation for fractures with either articular displacement >2 mm or carpometacarpal subluxation. PMID- 12132099 TI - Avascular necrosis of trapezium bone: a case report. AB - A 26-year-old woman presented with radiologic signs of avascular necrosis of the trapezium bone. Treatment by bone excision and suspension arthroplasty gave an excellent clinical result. Pathology studies confirmed the diagnosis of avascular necrosis of the trapezium. PMID- 12132100 TI - Antegrade intramedullary K-wire fixation for distal radial fractures. AB - Manual reduction and antegrade intramedullary K-wire fixation were performed for 29 selected Colles-type distal radial fractures in 29 patients. Severely comminuted intra-articular fractures and Barton's fracture were not included in this study. The tips of the K-wires supported an articular surface of the distal radius. Except for 1 case of skin irritation in the forearm, there were no complications related to tendon or nerve injuries or reflex dystrophy. There was no pin loosening, but protrusion of sharp tips of K-wires into the wrist joint occurred in 3 patients. Average volar tilt, radial shortening, and step-off at follow-up evaluation were 5 degrees, 2.6 mm, and 0.2 mm, respectively. These radiologic results were consistent with values reported in other pinning studies; however, radial shortening was not controlled effectively. Fourteen (48%) of 29 patients had radial shortening >3 mm. The advantages of this surgical procedure are the low occurrence rate of soft-tissue complications and prevention of dorsal angulation of fractures. PMID- 12132101 TI - Surgical treatment of redisplaced fractures of the distal radius in patients older than 60 years. AB - Twenty patients aged 60 years or older (average age, 68 y) presented to our institution with a distal radius fracture made complex by virtue of displacement after a closed reduction and cast or external fixation immobilization. Ten of the fractures were volarly angulated and 10 were dorsally angulated. Fifteen patients' hands had associated soft-tissue swelling. Surgical exposure and stable internal fixation were undertaken to both realign the fracture as well as enable functional rehabilitation. The final functional result and radiographic results were physician rated and patient rated by using the Patient Rated Wrist Evaluation (PRWE). The patients' overall activity level was quantified and compared with population norms by using the Physical Activity Scale for the Elderly (PASE). At an average follow-up visit of 38 months, all the fractures healed and none of the implants loosened or broke. The functional results were excellent in 7, good in 11, and fair in 2. The average PRWE score was 14, which compares favorably with prior studies of adult patients of all ages treated for a distal radius fracture. The PASE score averaged 177, representing a return to preoperative activity levels in 17 of 20 patients. Complications included loss of alignment, tendon rupture, transient radial sensory neuritis, and nonfatal pulmonary embolism in 1 patient each. Six plates were removed because of soft tissue irritation. PMID- 12132102 TI - Failure of a retinacular flap to prevent dorsal wrist pain after titanium Pi plate fixation of distal radius fractures. AB - Patients with distal radius fractures that had been treated with a dorsal Pi plate and retinacular flap covering the transverse limb of the Pi plate were evaluated clinically and radiographically. Nine of 20 patients (45%) required plate removal for dorsal wrist pain. Three of the remaining 11 who retained the plate had dorsal tenderness over the wrist extensors. There were no differences evident between the 2 groups in plate size, position, or number of screws used. In addition there were no significant differences between the groups in either radial height or inclination. The palmar tilt did show a trend toward statistical significance: those patients who required plate removal had an average of 4.1 degrees of dorsal tilt, patients whose plate was not removed averaged 2.8 degrees of palmar tilt. Our results show that the retinacular flap covering the distal transverse limb of the Pi plate did not prevent the occurrence of dorsal wrist pain. Dorsal wrist pain remained a problem with dorsal plating of distal radius fractures. PMID- 12132103 TI - Sigmoid notch reconstruction using osteoarticular graft in a severely comminuted distal radius fracture: a case report. AB - A case of a young patient with a severely comminuted intra-articular distal radius fracture dislocation and severe injury of the distal radioulnar joint is presented. Early reconstruction of the sigmoid notch and radioulnar ligaments was performed using the remaining scaphoid facet of the distal radius articular surface, an autogenous tendon graft for ligament reconstruction, and radioscapholunate arthrodesis. The patient was able to return to his manual work without limitations. We present additional information on the comparative anatomy of the sigmoid notch and scaphoid facet that may guide surgeons in treating this severe injury. PMID- 12132104 TI - Coincidental rupture of the scapholunate and lunotriquetral ligaments with volar intercalated segment instability complicating a closed distal forearm fracture in an adolescent. AB - A 15-year-old boy is described with concomitant scapholunate and lunotriquetral ligament injuries of the wrist, complicating a closed forearm fracture. The rotatory dislocation could be reduced and stabilized with a soft-tissue procedure. PMID- 12132105 TI - End-to-side nerve repair. PMID- 12132107 TI - Chronic urticaria and angioedema. PMID- 12132108 TI - Chronic urticaria and angioedema. PMID- 12132109 TI - Hypercarotenemia. PMID- 12132111 TI - Serious and enduring mental illness--schizophrenia:. PMID- 12132112 TI - International teaching material for mental health studying in English. PMID- 12132110 TI - The impact of depression on stroke recovery in the U.S. AB - Globally, stroke is an important cause of physical, cognitive and emotional disability. The success of rehabilitation efforts for stroke patients is often measured in terms of physical functioning. However, because the adaptation process also involves psychological and social factors, and nursing goal is to holistically meet patient's health care needs, it is important to address emotional and social variables that impact the healing process. Therefore, the purpose of this prospective longitudinal study was to determine the impact of depression on functional ability in adults (n = 50) who have had a stroke in the U.S. The study was conducted at two large urban medical centers and one community hospital in the Midwest of the U.S. Depression screening and functional status evaluations were conducted within 10 days (Time 1) and three months (Time 2) post stroke. In the total sample 46% (n = 23) described fewer depressive symptoms at Time 2; while 44% (n = 22) indicated more depressive symptoms, and 10% (n = 5) showed no difference between Time 1 and 2. In the total sample, 27 improved in functional status between Time 1 and Time 2, while 5 lost function at Time 2. The impact of stroke on depression in the U.S. as well as nursing implications globally are discussed. PMID- 12132113 TI - Efficiency of visual selective attention is related to the type of target. AB - Lavie and Cox (1997, Psychological Science, 8, 395) suggested that stimuli outside the focus of attention can have more distracting effects when the task requires less attention than when it requires more. This idea provides an explanation for the proposed dissociation found between two forms of attentional control (Folk & Remington, 1998, Journal of Experimental Psychology: Human Perception and Performance, 24, 847). This proposal was based on a red (green) distractor that captured attention when a red (green) target was used, but not when a green (red) target was used, and the further result that, if there were no distractor, reaction times were faster than with either type of distractor. We tested whether a target that requires serial search would eliminate the dissociation. With the same targets used by Folk and Remington (ones that can be found with parallel search), we also obtained a dissociation. However, with the serial-search targets, dissociation was not found. Our interpretation is that the dissociation represents two forms of the same attentional process. PMID- 12132114 TI - Facilitation of local information processing in the attentional blink as indexed by the shooting line illusion. AB - Perception of the second of two rapidly sequential targets is impaired if the temporal lag between them is relatively short. This attentional blink (AB) is said to occur because the first target preoccupies attentional resources, leading to a shortage of attention for, and subsequent failure of judgement of, the second target. In the present study, we examined whether the attention which is preoccupied by the first target carries the common characteristic of spatial attention, that is local facilitation of information processing. To index the facilitation, we presented an attention-sensitive probe known as the shooting line illusion in which observers see a progressively drawn line from the attended end to the other. We found that illusory motion was perceived to emanate from the end where the first target was presented. This result demonstrates the inherently spatial nature of attention, even when it is defined by temporal and symbolic variables. PMID- 12132115 TI - Effect of the similarity between target and global and local levels in hierarchical stimuli processing. AB - The hypothesis of global precedence was investigated by manipulating the similarity between target and relevant level, and between target and global and local levels of hierarchical stimuli, both in divided and selective attention conditions. The similarity was measured on the basis of the amount of shared contour between two visual forms. The stimuli were circles, right-facing semicircles, i.e., 1/2 of the circumference missing, and right-facing partial circles with a 1/4 of the circumference missing. The circle was used as a target and, consequently, the similarity between target and semicircle and between it and the partial circle was 50% and 75%, respectively. In the divided attention condition, the results showed a local advantage in the two similarity conditions between target and distractor level (50% and 75%) and a bidirectional and symmetrical interference at both levels of similarity. However, in the 'selective attention condition', a local reaction time advantage was only found when the similarity between target and irrelevant level was of 50%. When attention was directed towards the global level, local interference increased depending on the decrease in the degree of the local level similarity. When the attention was directed towards the local level, there was global interference but it was independent of the degree of the global level similarity. These results show the importance of the physical shape of the local elements used in experiments with hierarchical stimuli, since they may determine the relative speed of global processing, facilitating or impairing it. PMID- 12132116 TI - Effects of type of cognitive demand on bilateral advantage in interhemispheric processing. AB - The relationship between cognitive task demand and effect of bilateral advantage (BLA) was examined. In Experiment 1, the task demand based upon visual stimulus complexity was manipulated. One-digit and two-digit numbers were presented in the left, right or both visual fields tachistoscopically and subjects were requested to identify the numbers. The results showed no BLA, although a right visual field advantage was demonstrated. In Experiment 2, cognitive demand in terms of memory was manipulated. Subjects were asked to recall one-digit numbers that were presented successively once, three and five times in the left, right or the both visual fields. The results did not show any sign of BLA in the recall of numbers. In Experiment 3, a greater memory load task was given than that in Experiment 2, where two-digit numbers were presented successively (once, three and five times). Subjects were asked to recall the numbers. The results showed a significant BLA in the recall of numbers, i.e., the correct numbers in the correct temporal positions. These results strongly suggest that a benefit of redundant bilateral visual fields presentation is shown only in a highly cognitively demanding task, especially when it involves phonological memory loads. PMID- 12132117 TI - Learning to discriminate simple sinusoidal gratings is task specific. AB - Learning transfer effects of a modified spatial frequency discrimination task with simple sinusoidal gratings on various untrained test tasks, testing performance in relative position, local width and global size discrimination, have been investigated. Six subjects were exposed to grating stimuli of varying spatial frequency and size but constant relative position while they had to respond only to spatial frequency. In the course of 7 consecutive days all subjects showed significant reduction of spatial frequency discrimination thresholds. Comparison of discrimination thresholds for the untrained test tasks, taken before and after the learning epoch, reveals complete learning transfer to spatial frequency discrimination with gratings of constant size and variable relative position, but complete lack of transfer when grating spatial frequency is shifted about one octave lower. Further, there is improvement of relative position discrimination and width discrimination of single luminance bars, which is not very specific for the learnt spatial frequency. Although size variation was part of the learning procedure, discrimination of global stimulus size did not improve. Generally, the observed scheme of learning transfer reveals that there is learning only for stimulus attributes that are behaviorally relevant in the learning task. The differential scheme of improvement and code usage in the test tasks strongly indicates involvement of higher stages capable of independent access to different coding domains, as well as attentionally guided attribute selection and suppression. Supported by other recent findings, it is suggested that discrimination learning can be understood as a higher level process of gradual refinement of code selection out of a rich code base provided by lower level stages. PMID- 12132119 TI - Effects of a secondary task on "implicit" sequence learning: learning or performance? AB - Traditionally, implicit learning has been defined in terms of a lack of awareness of the process and products of learning. In the face of a number of conceptual and empirical difficulties with this definition, it has recently been suggested instead that the critical feature of implicit learning is that it proceeds without making any demands on attentional resources. As disconfirmatory evidence for this, we describe the results of two experiments which each used a sequential reaction time task. With a tone-counting secondary task, measures of sequence learning were significantly affected by whether training occurred under single- or dual-task conditions, regardless of whether testing took place under single- or dual-task conditions. PMID- 12132118 TI - Free recall and organization as a function of varying relational encoding in action memory. AB - A new approach was taken to study the question whether the free recall advantage of enacting actions (SPT), over only listening to their verbal descriptions (VT), is due to better relational encoding of SPTs than of VTs. The approach consisted of studying related lists and manipulating the degree of relational encoding by repeated list presentation in Experiment 1 and by presenting the list items at random or blocked in Experiment 2. In both experiments, free recall and adjusted ratio of clustering (ARC) scores increased as to be expected with repeated list presentation and blocking. However, these effects proved to be independent of the type of encoding. There was a clear SPT effect in free recall, but in contrast a slight though nonsignificant advantage of VTs over SPTs in ARC scores. Altogether, the experiments show that the SPT effect in free recall is not due to better relational encoding in SPTs than in VTs, in line with the assumption that this effect is due to good item-specific encoding. PMID- 12132120 TI - [Process management and controlling in diagnostic radiology in the hospital]. AB - Systematic process management and efficient quality control is rapidly gaining importance in our healthcare system. What does this mean for diagnostic radiology departments? To improve efficiency, quality and productivity the workflow within the department of diagnostic and interventional radiology at the University Hospital of Essen were restructured over the last two years. Furthermore, a controlling system was established. One of the pursued aims was to create a quality management system as a basis for the subsequent certification according to the ISO EN 9001:2000 norm. Central to the success of the workflow reorganisation was the training of selected members of the department's staff in process and quality management theory. Thereafter, a dedicated working group was created to prepare the reorganisation and the subsequent ISO certification with the support of a consulting partner. To assure a smooth implementation of the restructured workflow and create acceptance for the required ISO-9001 documentation, the entire staff was familiarized with the basic ideas of process- and quality-management in several training sessions. This manuscript summarizes the basic concepts of process and quality management as they were taught to our staff. A direct relationship towards diagnostic radiology is maintained throughout the text. PMID- 12132121 TI - [Change in process management by implementing RIS, PACS and flat-panel detectors]. AB - Implementation of radiological information systems (RIS) and picture archiving and communicating systems (PACS) results in significant changes of workflow in a radiological department. Additional connection with flat-panel detectors leads to a shortening of the work process. RIS and PACS implementation alone reduces the complete workflow by 21-80%. With flatpanel technology the image production process is further shortened by 25-30%. The workflow-steps are changed from original 17-12 with the implementation of RIS and PACS and to 5 with the integrated use of flatpanels. This clearly recognizable advantages in the workflow need an according financial investment. Several studies could show that the capitalisation-factor calculated over eight years is positive, with a gain range between 5-25%. Whether the additional implementation of flatpanel detectors results also in a positive capitalisation over the years, cannot be estimated exactly, at the moment, because the experiences are too short. Particularly critical are the interfaces, which needs a constant quality control. Our flatpanel detector-system is fixed, special images--as we have them in about 3-5% of all cases--need still conventional filmscreen or phosphorplate-systems. Full spine and long-leg examinations cannot be performed with sufficient exactness. Without any questions implementation of integrated RIS, PACS and flatpanel detector-system needs excellent training of the employees, because of the changes in workflow etc. The main profits of such an integrated implementation are an increase in quality in image and report datas, easier handling--there are almost no more cassettes necessary--and excessive shortening of workflow. PMID- 12132122 TI - [Improving productivity by implementing RIS and PACS throughout the clinic: a case study]. AB - PROBLEM: How are improvements in productivity in connection with RIS/PACS to be defined? What do they cost? To limit the problem to the relevant topics, we first describe the objectives of a radiology department and the identified bottlenecks in the workflow. How to define and assess the improvements is discussed. METHODS: The case in question for this study is the RIS/PACS project at the "Klinikum der Universitat Munchen, Campus Grosshadern". The goals of the project and its present status are reviewed. The project is not yet completed, so this is a "midterm" report. RESULTS AND DISCUSSION: We describe the status of the achieved and not yet achieved goals and of the eliminated bottlenecks. On the plus side, for example, nearly 100% of all digitally generated images (except mammogramms) are digitally archived. They are accessible to the same percentage in radiology via PACS and in the hospital via the webbased intranet image distribution system when needed. In some radiology areas, such as multislice CT, already the reporting can no longer be performed without softcopy image interpretation. However, the full elimination of hardcopy images is still not reality, since the distribution to DICOM viewers for selected requesters with demands for almost reporting quality, high cost image displays is still in the testphase. To reduce film costs, images are being printed on a high resolution paper printer in addition to the intranet distribution during this transition period. On the negative side, due to a lack of job positions in the transcription rooms, about 40% of the reports are still being handwritten by radiologists. Furthermore, the dictated and transcribed reports are usually still not available early enough in the RIS and thereby in the intranet report distribution of the hospital. Here only a speech recognition system can remedy the situation. As soon as this problem is solved and the image distribution to the DICOM viewers works routinely, the reports and the images will be accessible within minutes to maximally within some hours after the examination. CONCLUSION: The goals reached so far suffered delays due to unforeseen problems and pitfalls. Altogether, a quieter operation and workflow in radiology has already been achieved, due to less inquiries from the requestors for unfinished examinations, images and/or image copies. PMID- 12132123 TI - [Quality management in a radiology department]. AB - PURPOSE: To increase the quality of internal and external interactions (patients, clinical colleagues, technicians, radiologists) in a department of radiology. METHOD: Accompanied by a well-experienced adviser workshops have been performed dealing with different topics like "contact to patients," "performance of the radiological report and interaction with the referring colleague" or "research and teaching." A catalogue of different actions was defined to reduce hindrances within the internal and external work-flow. RESULTS: A total number of 53 actions was defined and related to different persons who were responsible for the realisation of the measures within a time interval. Six months after starting the quality management 46 (86%) of the defined actions were realised successfully, and another 4 (8%) measures were still running. There was a moderate increase of satisfaction of the patients and clinical colleagues considering the waiting time. CONCLUSIONS: A quality management in a radiological department allows an optimisation of the internal and external interactions. However, the guidance of a well-experienced adviser is as essential as the continuous control of successful finished measures. PMID- 12132124 TI - ["Activity based costing" in radiology]. AB - BACKGROUND: The introduction of diagnosis related groups for reimbursement of hospital services in Germany (g-drg) demands for a reconsideration of utilization of radiological products and costs related to them. METHODS: Traditional cost accounting as approach to internal, department related budgets are compared with the accounting method of activity based costing (ABC). The steps, which are necessary to implement ABC in radiology are developed. CONCLUSIONS: The introduction of a process-oriented cost analysis is feasible for radiology departments. ABC plays a central role in the set-up of decentralized controlling functions within this institutions. The implementation seems to be a strategic challenge for department managers to get more appropriate data for adequate enterprise decisions. The necessary steps of process analysis can be used for other purposes (Certification, digital migration) as well. PMID- 12132125 TI - [Telemedicine: chances and risks]. AB - PROBLEM: Due to the rapid progress in the fields of information technology and data networks, telemedicine applications are growing in number. Besides curative telemedicine, the electronic exchange of medical data and the integration of health information systems between health care providers is gaining importance. Through the improved accessibility of electronic patient record information, considerable risks arise. METHODS: A project for the interconnection of medical picture archiving and communication systems (PACS) between several hospitals is presented and the possibilities for further developing such networking systems utilizing new software technologies for transparent data access between different locations (GRID) and for decision support (software agents) are considered. RESULTS: The availability of the electronic patient record via the data network and the perspective of semi intelligent software systems automatically preparing the data bears great potential for a boost in treatment quality and efficiency. Systems for unique electronic patient identification and for secure digital signature are a prerequisite, but per se not enough to ensure the protection of data against illegitimate access. CONCLUSION: Despite quality and efficiency benefits, challenges in the protection of sensible data and in the change of the physicians role result. PMID- 12132126 TI - [Certification according to ISO 9001--waste of time or necessity?]. AB - Quality management (QM) systems are widely established management tools in the industry and business world today. In the health care sector, the need for quality, the implementation of quality management systems and economic orientation of the hospital management are strongly encouraged. Centers of excellence are created to offer special medical services of the highest possible quality. Furthermore, there is a worldwide tendency to establish standards in clinical medical settings, in teaching profession and medical science. These trends make the implementation of a quality management system in health care system highly desirable. The present manuscript reviews the fundamental principles and concepts and the aims of internationally accepted QM systems. It focuses on the ISO-9000 certificates and the European Foundation of Quality Management (EFQM) model. The advantages and disadvantages are discussed. The resources necessary for installation of a QM system and the different phases of implementation are reported. According to the experience of several groups, QM systems can be reliably used in a radiology department and lead to continuous improvement of the quality of services. PMID- 12132127 TI - [The value of magnetic resonance tomography (MRI) with reference to economic aspects of radiotherapy planning of vertebral metastases. A cost benefit analysis]. AB - Is magnetic resonance imaging (MRI) based target volume definition for treatment planning of vertebral metastasis effective under economic considerations. From 1994 to 1999, a total of 137 patients with bone metastases affecting the vertebral column underwent MRI of the cervical, thoracic, or lumbar spine for the treatment planning of palliative radiation therapy. The following radiation treatment consisted in a irradiation of the affected vertebral region up to a total dose of 30-40 Gy. The cost calculation for radiotherapy and magnetic resonance tomography was done using the common tariff model (EBM) of the German Health Insurances. In 73% of patients (101 patients), magnetic resonance imaging resulted in marked corrections of the irradiation fields which would have resulted in the necessity of treatment for recurrence in the case of treatment planning without MRI. Consequently, the higher cost of MRI of 345.00 DEM (176,40 EUR) lead to a saving of 497.00 DEM (254,11 EUR) compared to a recurrence treatment of 10 fractions and of 1,428.00 DEM (730,12 EUR) compared to 20 fractions. The transport expenses for the second treatment could be saved as well. Even under economic considerations MRI is effective. PMID- 12132128 TI - [Space-occupying lesion in the area of the 3rd ventricle. Cerebral pseudocystic metastasis of a neuroendocrine carcinoma of unknown primary tumor]. PMID- 12132129 TI - [Drugs in interventional radiology. 1: Prevention of allergy, antibiotic prophylaxis, inhibiting gastrointestinal motility, antiemetics, conscious sedation, vasodilation]. PMID- 12132130 TI - Dracunculiasis: Ethiopia and Uganda. PMID- 12132131 TI - Progress towards poliomyelitis eradication. PMID- 12132132 TI - New therapy for visceral leishmaniasis. PMID- 12132133 TI - [Request for labia correction: sometimes more than a simple question]. AB - Three women aged 19, 50 and 33 years, requested surgical correction of their labia minora because of subjective complaints attributed to the size of their labia minora. However, during consultation, for one of the patients it transpired that she did not know anything about the normal physiological changes of the external genitals during puberty and the enormous variety and diversity of the length of labia minora between women. It turned out that the other woman felt uncertain about her genitals following a recent divorce. For the third woman a vulvar pain syndrome and a sexual abuse history became clear. Two of the patients decided not to undergo surgery and the third sought a cure elsewhere. The request for a surgical correction of the labia minora seems quite simple and the operation does not seem to be complicated either. However, the question remains as to whether an operation is the solution for the psychological and behavioural consequences the woman experiences. A conservative approach is recommended, with attention for other possible problems that can be hidden behind the request for labia minora correction. PMID- 12132134 TI - [Combatting river blindness by means of chemotherapy directed at the symbiotic Wolbachia bacteria in the causative filariae]. AB - In a mouse model of river blindness it was demonstrated that Wolbachia bacteria, endosymbionts of filarial nematodes, play an important role in the inflammatory process leading to the disease and that this process depends on Toll-like receptor 4. Wolbachia is found in many arthropods and in all filariae pathogenic for man. Treatment with doxycycline depletes female filariae of Wolbachia and renders them infertile for at least 18 months. Chemotherapy of filarial nematodes should be studied for the reduction and possibly prevention of pathology due to filariae, and for a contribution to control and eradication programmes. PMID- 12132135 TI - [Spreading of hereditary diseases through donor sperm: no reason for reducing the number of offspring per donor in the Netherlands]. AB - In the Netherlands, a limit of 25 offspring per sperm donor has been applied for many years. This figure was based on the assumption that children from sperm donors are not at greater risk of having consanguineous relationships than a random individual of the Dutch population. After the report in 2002 that a donor, whose sperm was used to father 18 children by artificial insemination, had developed a serious hereditary brain disease, the question of whether the limit of 25 should be reduced was raised. From a population genetics point of view, reducing the number of children per donor will only result in greater genetic diversity being transmitted to the offspring of sperm donors. Moreover, the number of children from sperm donors is a negligibly small proportion of the general population (0.5% of the total number of newly-borns). Since no knowledge of other offspring from one particular donor is available to individuals making use of donor insemination, only optimally guaranteed safety (both genetic and microbial) is important to these people and their offspring, given that the risk of inbreeding is acceptably low. Finally, the interests of the donor can be properly safeguarded through individual agreements. Therefore, to conclude, there are no reasons to reduce the limit of 25 offspring per sperm donor. PMID- 12132136 TI - [Levetiracetam: an anti-epileptic drug with interesting pharmacokinetic properties]. AB - Levetiracetam is a new anticonvulsant for adjunctive treatment of partial epilepsy. It is well tolerated, with no significant risks, at a dose of 1000-3000 mg/day in adults. The efficacy (> 50% reduction in attacks) in refractory partial epilepsy is 22-40%, depending on the dose. Efficacy was also seen with levetiracetam monotherapy in more than half of the positive responders. Levetiracetam does not cause induction or inhibition of the P450 enzyme system or other enzyme systems, there is no active metabolite and it exhibits almost no protein binding. These factors mean that this drug undergoes no significant interactions with other medication and appears suitable for elderly patients and for conditions requiring complex pharmacotherapy. Compared with other recently registered anti-epilepsy drugs, levetiracetam appears promising in terms of efficacy, tolerability and pharmacokinetics. The simple dosing schedule is an additional benefit. PMID- 12132137 TI - [A flow diagram for the diagnosis of acute, non-traumatic joint complaints during childhood, retrospectively applied to 115 children]. AB - Acute, non-traumatic joint complaints during childhood can be caused by conditions which require a quick and adequate recognition and treatment as well as by conditions in which an expectant policy can be pursued. On the basis of certain data from the anamnesis, supplemented with findings from the physical examination it is often possible to arrive at a (probable) diagnosis. An algorithm was designed, the differential steps of which were: fever, C-reactive protein titre, involvement of the hip joint, the presence of extra-articular manifestations and the results of a full blood count, erythrocyte sedimentation rate and imaging techniques. When this algorithm was retrospectively applied to the disease data of 115 children with acute, non-traumatic joint complaints, for whom the diagnosis in the status was taken as the gold standard, the correct diagnosis was established for every single child: for 98 (85.2%) by the shortest route and for 17 (14.8%) indirectly. In the case of 4 children, use of this algorithm would have led to unnecessary laboratory investigations and/or treatment. None of the diseases requiring immediate treatment were missed. PMID- 12132138 TI - [Diagnostic image (94). A man with a bleeding cutaneous nodule. Lobular or capillary hemangioma]. AB - A 24-year-old man presented with an easily compressible cutaneous tumour of the chest. Histological examination confirmed the diagnosis 'granuloma teleangiectaticum' (lobular or capillary hemangioma). PMID- 12132139 TI - [Idiopathic scoliosis in adolescents; an inventory into the possibilities of studying the efficacy of screening and treatment]. AB - OBJECTIVE: To collect information for the purpose of establishing starting points and possibilities for a cost-effectiveness analysis of screening for adolescent idiopathic scoliosis (AIS). DESIGN: Interviews, literature review, questionnaires, an estimation of costs and discussions with experts and involved parties. METHOD: Following an initial interview with 16 orthopaedic surgeons and school doctors a literature study into the efficacy of treatment was carried out regarding the years 1989-1999. The variation in current practice was delineated by means of a questionnaire sent to all 51 municipal health services in the Netherlands. The costs of screening and treatment were estimated on the basis of health insurance premiums and a municipal health service cost model. All of the results were presented to five methodological experts and finally the study results and the recommendations of the five methodological experts were evaluated during a meeting of persons especially invited for this purpose. RESULTS: Screening for AIS was established to realise early diagnosis and treatment with a brace, so as to reduce unsatisfactory cosmetic outcomes and the need for surgery. Screening was performed using the bending test and was performed in 40/48 (83%) of the participating municipal health services. The overall costs of screening and treatment amount to 6 million euros per year. There was no convincing evidence that the screening programme was sufficiently sensitive and bracing sufficiently effective. Neither was there proof of the opposite. The following was recommended: obtain reliable data by carrying out a randomised controlled trial on the effectiveness of treating AIS with bracing in an early stage; carry out a case-control study combined with a retrospective patient follow-up study to evaluate the current screening practice; draw up a national standard for the screening of postural disorders in youth healthcare to ensure effective practice. PMID- 12132140 TI - [Dyspnoea and dysphagia in an adult caused by a tracheo-esophageal fistula]. AB - In a 64-year-old man who was suffering from chronic obstructive pulmonary disease, recurrent airway infections, dysphagia, and weight loss, achalasia was diagnosed on the basis of endoscopic and radiological examinations. Afterwards he underwent flexible bronchoscopy, which revealed a benign looking fistula between trachea and oesophagus. This appeared to be a congenital tracheo-oesophageal fistula. The fistula was closed surgically. Three months later breathlessness and a sputum-producing cough were the only remaining symptoms. This rare anomaly is mostly diagnosed during childhood, but can also manifest itself in adulthood. If a tracheo-oesophageal fistula is suspected, the diagnostic procedures of choice are a barium oesophagogram in a forward-sitting or supine position or endoscopy of the trachea. Treatment consists of division and closure of the fistula. The prognosis is good. PMID- 12132141 TI - [Delirium during withdrawal of venlafaxine]. AB - A 35-year-old man with anxiety and depression who was treated with venlafaxine, 300 mg a day, developed severe withdrawal symptoms in the form of a delirium during gradual tapering of the dosage. The symptoms resolved when the dosage was kept constant and did not recur when the dosage was reduced more gradually. Withdrawal symptoms are common during discontinuation of antidepressants, particularly after prolonged use of agents with a short half-life. The symptoms are usually mild and transient, especially in the case of selective serotonin reuptake inhibitors and venlafaxine. The occurrence of delirium as a result of the withdrawal of venlafaxine has not been reported previously. Even when antidepressants are being withdrawn with care, one should remain alert to the possible development of severe withdrawal symptoms. PMID- 12132142 TI - [One hundred years of the Association of Surgeons in the Netherlands. VI. The training]. AB - Over the last 25 years the organisation and content of the residency training program for general surgeons have been adapted to meet the needs of changing surgical practice. Recently more profound changes have been dictated by the Dutch Working Hours Act, which has strictly limited the working hours of resident physicians. With this the emphasis will be on improving theoretical and practical training methods. Because of the limiting working hours resident physicians will have a smaller role in patient care. These changes will require a huge effort from both the teaching surgeons and the resident physicians, as well as substantial financial investments from the government and healthcare providers. PMID- 12132143 TI - [A European discussion about stem cells for therapeutic use]. AB - Stem cells as a source material for growing cellular transplants to repair dysfunctional organs appear to be a new challenge for medical science. Though stem cells are also present in foetal and adult organs, embryonic stem cells from the pre-implantation embryo in particular have the potency to proliferate easily in vitro and the capacity to differentiate into all the body's organ-specific cells. Therefore, these are the ideal cells for developing new cell transplantation therapies for diseases such as Parkinson's disease, diabetes mellitus and heart failure. The use of spare in vitro fertilization (IVF) embryos or pre-implantation embryos specially created to harvest human embryonic stem cells is, however, controversial and an ethical problem. In a European discussion platform organised by the European Commission Research Directorate-General, the status quo of the progress was presented and subsequently commented upon and discussed in terms of medical-ethical, social, industrial and patient interests. The expectations of this new medical technology were high, but clinical trials seem only acceptable once the in vitro differentiation of stem cells can be adequately controlled and once it is known how in vitro prepared stem cells behave after implantation. The ethical justification of the use of in vitro pre implantation embryos remains controversial. The prevailing view is that the interests of severely ill patients for whom no adequate therapy exists, surmounts the interest of protection of a human in vitro pre-implantation embryo, regardless of whether it was the result of IVF or of transplantation of a somatic cell nucleus of the patient in an enucleated donor egg cell (therapeutic cloning). PMID- 12132144 TI - [Postprandial glucose peaks in the pathogenesis of cardiovascular disease in diabetes mellitus; implications for metabolic control]. PMID- 12132145 TI - [HIV prevalence among injecting drug users in South Limburg, 1994-1998/1999: increasing trend in Heerlen, not in Maastricht]. PMID- 12132146 TI - [Disinfection of the skin prior to injections does not influence the incidence of infections; a literature study]. PMID- 12132147 TI - [Cardiovascular effects of hyperthyroidism and treatment thereof]. PMID- 12132148 TI - [Analysis of natural regeneration barriers of Pinus sylvestris var. mongolica plantation on sandy land]. AB - By employing comparison analysis, and field experiment of watering and soil covering before overwintering for seedlings, the barriers of natural regeneration for Mongolian pine plantations on sandy soil were identified. The experimental area was divided into 3 parts according to the state of natural regeneration. Crown closure, litter, understory coverage, and site condition were the factors which affect natural regeneration. Water deficit, but not low temperature during overwintering, is the key factor that limits survival for 1-2 years old seedlings in plantation area. The water deficit is due to higher air temperature, less accumulated snow, higher soil evaporation, higher plant transpiration in plantation area than in the area in which Mongolian pine is naturally distributed. Based on the above research, it is necessary to take effective artificial measures to promote its natural regeneration so as to guarantee its successful development. The following measures are recommended: 1) covering the 1 2 years old seedlings with soil before wintering, 2) irrigating seedlings before wintering, 3) suitable forest harvest, e.g., belt clear-cutting, or patch cutting, 4) planting the pine with broadleaf species, 5) protecting forest stands from livestock grazing, seed collecting and litter gathering. PMID- 12132149 TI - [Analysis on the sustainability of Amomum villosum cultivation under the tropical rainforest in Xishuangbanna]. AB - Amomum villosum has been cultivated under tropic rainforest in Xishuangbanna area since 1970s. The cultivation area of Amomum villosum had expanded to 5811 hm2 by 1998, and the dry fruit yield had reached to 542 t. In this study, the ecological and economical benefits of this cultivation model were analyzed. The results showed that plant diversity, litter, soil humidity, and soil fertility of rainforest decreased at different levels after Amomum villosum cultivated under rainforest, and the distribution of nutrition had been changed in the soil. The economical benefits had an obvious downtrend after 7-8 years cultivating due to irrational management. So, the present cultivation model of Amomum villosum under the tropical rainforest was not sustainable. For the objectives of harmonizing the ecological, economic, and social benefits and sustainable utilization of the rainforest, and improved model was also discussed in this study. PMID- 12132150 TI - [A comparative study on life-form spectra of evergreen broad-leaved forest in different successions in Jinyun Mountain]. AB - The life-form of plant communities in different succession stages in Jinyun Mountain was studied and the life-form spectrum was established. The results showed that according to the characters of life-form of plant communities, the vegetation in Jinyun Mountain belonged to typical subtropical evergreen broad leaved forest vegetation. Evergreen phaenerophytes were absolutely dominant in the life-form spectrum and its quantity accounted for more than 80%. Among phaenerophyte, quantity of microphanerophyte was the most, and quantities of mesophanerophtye and nanophanerophyte were less. Different successions had different life-form spectra. With the development of succession, the percentage of phaenerophytes increased, and other life-forms decreased correspondingly. According to the development of life-form and comparative analysis of environmental factors in different succession stages, coniferous species would be replaced by evergreen phaenerophytes. PMID- 12132151 TI - [Neighbor diversity and interspecific association of Quercus mongolica]. AB - Neighbor diversity and interspecific association of Quercus mongolica were studied through point sample method. With the increase of plots number, species richness and Shannon-Wiener (H') diversity index increased. At last they came close to fixed value. LS value (average number of neighboring species) was first introduced in this article. Different sits had different PC value (value of interspecific association). According to PC value and LS value, accompany tree and shrub species of Quercus mongolica were determined. The similar environmental requirement and mutualism were the main reason why they live together. The result of interspecific association between Quercus mongolica and other species in 4 locations was analyzed. The reason why the differences existed was that Quercus mongolica studied was in different succession stages. PMID- 12132152 TI - [Effect of drought stress on apparent resource utilization efficiency of Quercus mongolica]. AB - To study the effect of drought stress on the physiological characteristics and resource utilization efficiency of Quercus mongolica, a dominant species in the Korean pine broad-leaved forest in Changbai Mountain, a comparative experiment was carried out under three soil moisture gradients as 85%-100% (high water, CK), 65%-85% (Medium water, MW), and 45%-65% (low water, LW) of field water-bolding capacity. The results showed that net photosynthetic rate, stomatal conduction, transpiration rate, water utilization efficiency, apparent CO2 utilization efficiency and apparent light utilization efficiency of the seedlings of Quercus mongolica were all significantly influenced by simulated drought stress. Stomatal conduction, water utilization efficiency and net photosynthetic rate of large tress of Quercus mongolica were affected by the soil drought stress, while transpiration rate, apparent CO2 utilization efficiency, and apparent light utilization efficiency were not influenced. Moderate soil water condition could greatly increase gas exchange and water utilization efficiency. These results indicated that the responses of gas exchange and resource utilization efficiency to various water stresses were different. Quercus mongolica was a species with variable resistance to drought stress, and the resistance would be improved by drought stress, especially moderate water stress. PMID- 12132153 TI - [Clutch size, nesting success and breeding success rate in Emberiza jankowskii in the grassland at Baicheng in Jilin Province]. AB - In this article, clutch size, nesting success and breeding success rate of Emberiza jankowskii living in grassland habitats in Baicheng Region of Jilin Province were studied from May to July in 1999. The results showed average clutch size was 5.09 +/- 0.58 eggs per nest. There was negative correlation between clutch size and initial laying date, brood quantity and initial laying date, clutch size and fresh egg weight. There was exceedingly negative correlation between initial laying date and incubated rate. There was exceedingly positive correlation between nest size and clutch size. There was positive correlation between other attributes of nest and clutch size. Average incubation period was 12 +/- 0.94 days, incubated rate was 36.3%, and breeding success rate was 11.11%. The population size of brood size above seven days was 2.56 +/- 1.53. Nestling survival rate was 27.69%. PMID- 12132154 TI - [Quantitative characteristics of degenerative succession in Festuca sinensis sowing grassland in the alpine pastoral area]. AB - Quantitative characteristics of degenerative succession in Festuca sinensis sowing grassland in the alpine pastoral area were studied. The results showed that with the succession developing, number of species, richness index, diversity index, and evenness index were all increased, but ecological dominance was decreased. Analysis on important values, niche width, and niche overlaps proved that some herb species such as Ajania tenuifolia and Polygomum sibiricum were pioneer plants invading sowing grassland in the succession process. The pioneer plants had wider niche widths than other species, and their important values increased with the succession developing. Niche overlaps between Festuca sinensis and other species were smaller than that between two latifoliate herbs species in most case. The invasion of latifoliate herbs was one of the most important factors, which made the sowing grassland more degenerative. PMID- 12132155 TI - [Selection of optimum periods for rice estimation using remote sensing data based on GIS]. AB - The optimum temporal for rice estimation using remote sensing data included the optimum temporal for rice planting area estimation and the optimum temporal for rice yield estimation. Based on GIS, the rice potential planting area was extracted. Therefore, the study area became smaller and the plant species considered decreased. So the optimum temporal for rice planting area estimation were determined through comparing the crop phonological calendars. In order to select the optimum temporal for rice yield estimation using remote sensing data, the relationships between the rice yield and agronomic parameters, the agronomic parameters and spectral variables, the rice yield and spectral variables for different development stages using field experiment data in 1989 and 1990 were studied. The result showed that the key period of rice production was from boot stage to heading stage. At last, the optimum temporal for rice yield estimation using remote sensing data was selected using the rice phenological calendars of 1998. PMID- 12132156 TI - [Morphology of wheat roots under low-phosphorus stress]. AB - The morphology of root systems of different wheat (Triticum aestivum L.) genotypes under low-phosphorus stress were studied to determine the effects of external factors on components of root system and the early morphological indicators related to phosphorus efficiency. The number of root axes and the length of lateral root of P-deficient plant were significantly lower than those of P-sufficient plant. The length of root axis and root system, and the number of lateral roots were sharply increased under low-P stress. The number and length of root axis were significantly different under different levels of phosphorus supply and among different wheat genotypes under same phosphorus supply. This implied that the two traits (number and length of root axis) were controlled by genotype and external factors. The difference in the characteristics of lateral root of the given wheat genotypes was significant only between different levels of P supply. It showed that the traits of lateral root mainly depended on external factors. The length and number of root axis, root length, and root angle were significantly different among 6 wheat genotypes. There exited significant linear relationships between relative grain yield and the interaction of the morphological traits, and it implied that the traits could be used as early indicators of selecting high P-efficiency wheat varieties. PMID- 12132157 TI - [Microbial biomass affected by cadmium with the occurrence of organic acids in red soil]. AB - Soil microbial biomass carbon and nitrogen were determined after adding fixed concentration of organic acids and changeable concentration of cadmium in pre incubated red soil and 14-day incubation at 25 degrees C. The results show that soil microbial biomass carbon and nitrogen in the soil added with organic acids decreased with increasing concentration of cadmium. When the concentration of cadmium was higher than 25 mg.kg-1, the microbial biomass carbon and nitrogen in the soil added with low molecular weight of organic acids (oxalic acid, acetic acid, citric acid, tartaric acid) were lower than those without the addition of organic acids, indicating that low molecular weight of organic acids could increase the toxicity of cadmium. On the contrary, the microbial biomass carbon and nitrogen in the soil added with low molecular weight of organic acids were higher than those without the addition of organic acids when the concentration of cadmium was lower than 25 mg.kg-1, indicating that low molecular weight organic acids could decrease the toxicity of cadmium. The microbial biomass carbon and mitrogen in the soil containing humic acid were higher than those without the addition of organic acids, indicating that humic acid could decrease the toxicity of cadmium. However, the C:N ratio of soil microbial biomass was increased with increasing concentration of cadmium. Organic acids could increase microbial biomass carbon and nitrogen when soil was not added with cadmium. PMID- 12132158 TI - [Effects of maize stubble remaining in field on dynamics of soil microbial biomass C and soil enzyme activities]. AB - This study dealt with the effects of maize stubble remaining in field on dynamics of soil microbial biomass C and soil enzyme activities. The results showed that maize stubble remaining in field could raise soil microbial biomass C and the activities of urease, phosphatase, cellulase, and invertase in soil remarkably. According to the dynamics of soil microbial biomass C and activities of the four enzymes, the five characters reached their maximum values about 60 days after sowing, and the crops were in bloom of growth. The results also showed that maize stubble remaining in field had positive effects on improving fertility and keeping high and steady yield of maize. PMID- 12132159 TI - [Exudation and accumulation of citric acid in common bean in response to Al toxicity stress]. AB - Significant differences in the exudation and accumulation of citric acid in common bean genotypes were observed in response to Al toxicity stress by hydroponic cultural experiments. Secreted citric acid increased with increasing external concentrations of Al3+ which ranged from 0 to 50 mumol.L-1, while ranged from 50 to 80 mumol.L-1, secreted citric acid decreased with increasing external concentrations of Al3+. Among different genotypic common beans, citric acid secreted in G19842 was the largest, while Al uptake per unit dry weight in G19842 was the least. No difference in the accumulation of citric acid in leaves was found among different genotypic common beans, while the size of the content of citric acid in roots was G19842 > AFR > ZPV > G5273. The amount of citric acid exuded was smaller induced by phosphorus deficiency than that induced by Al toxicity stress. Exposure to 50 mumol.L-1 LaCl3 could not induce the exudation of citric acid, and it implied that the exudation and accumulation of citric acid in common bean was an important physiological response of resistance to Al toxicity stress. PMID- 12132160 TI - [Effects of magnesium deficiency on senescence of Dimocarpus longana leaves]. AB - 180 days old Dimocarpus longana seedlings were grown in nutrition solution with three magnesium concentrations as 4 mmol.L-1 Mg2+ (sufficient supply), 0.4 mmol.L 1 Mg2+ (deficient supply), and Mg-free (without supply). Leaves were sampled 120 and 150 days later after treatment. With an increase in severity of Mg-deficient, the chlorophyll content, PS II activities, photosynthetic rate, protein, nucleic acid (DNA, RNA), and ZRs contents declined progressively. Whereas the content of H2O2, the rate of O2-. generation as well as the content of malondialdehyde (MDA) increased. The results suggested that magnesium deficiency had significant effect on the senescence of Dimocarpus longana leaves. PMID- 12132161 TI - [Contribution to vegetable mercury from atmosphere and soil]. AB - In this research, the characteristics of Hg accumulation in vegetable and the contribution of atmosphere and soil to vegetable Hg were studied by field observation and pot experiments. 11 kinds of vegetable were experimented in field. The results indicated that the Hg contents in roots and leaves of vegetables were higher than that in stems and fruits. In the natural condition, more than 60% of the assimilated Hg was distributed in the up-ground edible parts. The results of pot experiments showed that the Hg content of the edible parts of carrot and lettuce exceeded the Food Health Standard Value (FASV), when the atmospheric Hg content reached 57.6 +/- 14.7 ng.m-3. This result indicted that the contribution of atmosphere and soil to vegetable Hg were 70.4-90.7% and 9.3-29.6% respectively. So, the vegetable Hg was affected mainly by atmosphere Hg. PMID- 12132163 TI - [Ecological economic analysis of a rice-crab model]. AB - Based on field investigation and experimental data analysis, ecological economic principles were applied to study the structure and function, the characteristics of material and energy flows, and the economic and ecological benefits of a rice crab model. The results show that rice-crab model had a higher integral benefit than rice model, and its net income, cost-benefit ratio, and labor productivity were increased by 382%, 67.7% and 295%, respectively. In view of the ecological benefits produced, the total nitrogen, alkaline-hydrolyzal nitrogen, total phosphorus, rapidly available phosphorus, rapidly available potassium, and organic matter in the soil after 1-year carb breeding were increased by 10.6%, 3.3%, 5.8%, 11.7%, 3.5% and 10.5%, respectively. Moreover, 6.375 kg N.hm-2 of non point source pollution load could be cut down annually. PMID- 12132162 TI - [Changes of soil physical, chemical and ecological factors under mechanized cultivation]. AB - Three-years agricultural mechanization extension project in Huang-Huai-Hai regions showed that the application of comprehensive agricultural technologies which included the return of straw and stalk to field by mechanization, deep application of fertilizer, deep plough and soil no-tillage with mulch, had an obvious biological effects. In comparing with traditional cultivation, the comprehensive mechanized cultivation could decrease soil bulk density by 0.08 g.cm-3, increase soil organic mater by 12%, improve moisture utilization by 10.1 13.6%, and increase the grain yields of wheat and corn by 1218 kg.hm-2. PMID- 12132164 TI - [Bioremediation of PAHs contaminated soil using bio-slurry reactor process]. AB - Studies on bioremediation of PAHs contaminated soil were carried out using bio slurry reactor process. Phenanthrene (PHE) and pyrene (PY) were chosen as the test pollutants. The results showed that the physical and chemical properties of the pollutants were key factor affecting the bio-remedying possibility of PAHs in soil. PAHs with less benzene rings and lower molecule weight were easier to be biodegraded. So, phenanthrene had higher bio-remedying possibility than pyrene. Temperature and airflow were important control factors in bio-slurry reactor process. In this research, the optimized parameters were operating temperature as 20-30 degrees C, ratio between water and soil as 2:1, airflow as 8 L.h-1.L-1, and the inoculating amount as 50 g.kg-1. PMID- 12132165 TI - [Toxicity assessment of soil contaminated by heavy metals using algae growth inhibition test]. AB - Scenedesmus obliquus growth inhibition test was carried out to assess the toxicity of soil contaminated by heavy metals. The results showed that the growth rate of Scenedesmus obliquus was positively related to the concentration of heavy metals added to soil, and it was decreased with the increasing concentration of heavy metals. Two parameters were chosen and the sensitivities were compared. It was found that the cell growth rate was more sensitive than OD growth rate. The orders of general detection limit by EC50 of different metals were Pb > Cu > Zn > Cd by using cell growth rate as the test index and Pb > Cu > Cd > Zn by using OD as the test index. This study also indicated that under the complex pollution condition, the toxicity of heavy metals in soil was stronger than that in the case of soil contaminated by single metal. PMID- 12132166 TI - [Agricultural application of sludge dredged from landscape water bodies]. AB - The feasibility and ecological effects of agricultural application of sludge dredged from the Grand Canal (Hangzhou Section) were studied. Applied too much dredged sludge to red soil and paddy soil affected the germination of alpine fescue [Fescuta ovina var. brachyphylla (Schult.) Piper] and colver (Trifolium repens. L) seeds, while there wasn't significantly affects in pot experiment. While the application rate was lower than 270 t.hm-2, the growth of pakchoi (Brassica chinensis L.) increased as application rate increasing. While the application rate was higher than 270 t.hm-2, the growth of pakchoi decreased. The flowers and grasses in garden were more suitable for the dredged sludge application, and there was significant increase of growth while the application rate was lower than 1080 t.hm-2. Contents of copper and zinc exceed hygiene standard, while the application rate was above 1350 t.hm-2. While the application rate was lower than 450 t.hm-2, the pollution of the groundwater had not been observed. The results showed that land application was an economical and feasible way for the disposal of sludge dredged from landscape water bodies, and horticultural application was more safe and economical than agricultural application. PMID- 12132167 TI - [Influences of body weight and temperature on standard metabolic rate of Paralichthys olivaceus]. AB - The standard metabolic rate of Paralichthys olivaceus was increased with its increasing body weight, whose relationship could be described as a power function. The rate change with temperature could be expressed by an exponential function, Rs = 0.2340 W0.6695 e0.0372T. There was no significant interaction between body weight and temperature in the process of the standard metabolism for the living organism. PMID- 12132168 TI - [Stimulation effect of anthracene on marine microalgae growth]. AB - Two species of marine microalgae, Isochrysis galbana 8701 and Skeletonema costatum, were exposed to low concentration of anthracene, and their cell density, contents of chlorophyll a, carotinoid and protein, and superoxide dismutase (SOD) activity were examined. It was showed that low concentration of anthracene (1.5-6.0 micrograms.L) could obviously stimulate the growth of these microalgae, and their protein, chlorophyll a, and carotinoid contents increased with increasing cell density. SOD of treated groups remained high activity, compared with the controlled group during the whole experiment. PMID- 12132169 TI - [On the tasks and developmental problems of forest ecology]. PMID- 12132170 TI - [Frontier fields of plant chemical ecology in the 21st century]. AB - It has focused on chemical interactions between plant and other organisms, which mediated by secondary plant metabolites in recent years. Induced chemical defense in plant, plant chemical communication, relationships between secondary plant metabolites and evolution, chemical relationships between plant and human and chemical ecology of marine plant are the frontier fields needed attention of plant chemical ecology in the 21st century. Progress in these frontier fields of plant chemical ecology will play an important role in sustainable development, particular in increasing agricultural production and effective control of pest, disease and weed under ecological security in the 21st century. PMID- 12132171 TI - [On the basic concepts and contents of ecological security]. AB - Security is the inverse function of risk, generally regarded as safeguard degree expectation state of assessment object or reliability of prevent imperfect and uncertainty event to happen. Ecological security can defined as mankind's ensure degree un-effected by ecological destroy and environmental pollution in yield, living and health, including basic element of water and food security, air quality and green environment. The mostly content of ecological security consists of ecological health diagnosis, regional ecological risk analysis, landscape security pattern, ecological security monitoring and prediction, and ecological security management and guarantee etc. Study on regional ecological security has characteristics of macro-scope and pertinence, assessment criterion of relativity and expansibility. Ecological security prediction and design should embody the capability of human activity. At last, authors discuss the measures of the ecological security ensure of inland watershed and ecological security analysis of oasis landscape. PMID- 12132172 TI - [Present status and prospects in research on effect of enhanced UV-B radiation on plants]. AB - Current research status on the effect of enhanced UV-B radiation on plants was reviewed in this paper. The main research fields involved plant growth and morphological structure, plant physiological and biochemical metabolism, plant genetic material, UV-B-absorbing compounds and some gene expression, population, and ecosystem. In addition, some issues worthy to be studied in this field in future were proposed. PMID- 12132173 TI - [Ecological effects of cover crops]. AB - This paper reviewed the effects of cover crops in reducing soil loss, surface runoff, NO3- leaching and water pollution, and elucidated roles of cover crops in controlling pest insects, weeds and diseases, and increasing soil nutrients. The potential roles and appropriate application of cover crops in sustainable development of agriculture were also discussed. PMID- 12132174 TI - [Microbial reduction of iron, manganese as well as other metals and their individual significance in environmental bioremediation]. AB - In this paper, microbial reduction of iron, manganese and other metals in soils and waters were discussed. Microorganisms reducing to different metals such as iron, manganese and uranium as well as selenium and their enzymatic and non enzymatic mechanisms for reducing to different metals were summarized. Moreover, the significance of reduction of different microorganisms of metals in the bioremediation of metals-contaminated environment was also evaluated. PMID- 12132175 TI - [Impact of root exudates from transgenic plants on soil micro-ecosystems]. AB - With the commercialization of transgenic plants, ecological risk assessment of transgenic plants has been scheduled. Many problems such as gene transfer from transgenic plants to related wild species, production of super weeds and super virus, tolerance to insect-resistant transgenic plants, and distruption of biodiversity have been taken place in some transgenic plants. The influences of root exudates from transgenic plants on soil micro-ecosystems were reviewed. PMID- 12132176 TI - [Genetics study on photo- and thermo-sensitive genic male sterility of indica rice (O. satival L.)]. AB - The sterility segregation and its genetic patterns of indica P(T)GMS rice were analyzed mainly based on the investigation of bagged seed-set of individuals in reciprocal F2 population from the crosses between P(T)GMS lines of different origin and typical indica conventional varieties of different ecotype under long day-length and high temperature condition by using the software of the maximum likelihood method. The results showed that the sterility of Annong S-1, Hengnong S-1, and W6154S, of which the sterility alteration was mainly induced by temperature fluctuation, was controlled by one major recessive gene. But the sterility of W7415S was controlled by more than two major recessive genes. Apart from the major recessive gene, there was a group of polygenes in controlling of the sterility segregation in F2 population. Moreover, the polygenic variance was different from line to line. It implied that the polygenes of W6154S or W7415S was far more complicated than that of Annong S-1 and Hengnong S-1 according to the maximum likelihood method. All the conventional indica varieties of different ecotypes possessed the major dominant fertility genes. Meanwhile, the genetic background in different ecotype conventional varieties, the essentiality of which lies in the existence of polygenes, was different and influenced the major gene expression deeply. It was also indicated that genetic background of middle-season variety Nanjing 11 was more complicated than that of early-season variety Erjiuqing and late-season variety GER-1 as well. Finally, the differences of drift in critical temperature inducing male sterility of P(T)GMS lines (for short term DCT) and the strategy of breeding practically usable P(T)GMS line with slighter DCT were discussed in the paper. PMID- 12132177 TI - [Effect of phosphorus deficiency on activity of acid phosphatase exuded by wheat roots]. AB - The activity of acid phosphatase exuded by roots, the tissue location of the enzyme, and the relationship between the enzyme activity and phosphorus efficiency of wheat were studied. The results showed that the activity of acid phosphatase exuded by wheat 81(85)5-3-3-3 and NC37 under P-sufficiency treat were lower than those under P-deficiency, and the enzyme activity of the former variety was significantly higher than that of the latter. There was a significant difference in the enzyme activity among 12 wheat genotypes grown under P deficiency treat. Acid phosphatase was exuded by epidermis cell of root, especially by epidermal cell of root apex. Thus, there was a linear relationship between the enzyme activity and the surface area of root or the number of root apexes. It implied that the enzyme activity was markedly related to the size of root system. The linear relationship between relative grain yield and acid phosphatase activity was significant. It indicates that the enzyme activity could be used as an early indicator to select P-efficient wheat genotypes. PMID- 12132178 TI - [Effects of enhanced ammonium nutrition on content and accumulation of nitrogen and mineral nutrients in wheat plant]. AB - The hydroponics experiment was conducted to study the nutrition basis for growth promotion of wheat by Enhanced Ammonium Nutrition(EAN). Compared with fertilizing NO3- alone, EAN significantly increased N concentration and accumulation in leaves, sheaths, and whole plant, but there were not significant effects on roots. EAN had no significant effects on concentration of P and K, but decreased that of Ca and Mg in whole plant. EAN increased accumulation of P, K, and Ca in whole plant, and decreased that of Mg, whereas single NH4+ decreased accumulation of all mineral nutrients. EAN had different effects on accumulation of mineral nutrients in different genotypes. Compared with fertilizing NO3- alone, EAN increased accumulation of P, K, and Ca of YM158 and LZ953. For the accumulation of JDM, there was little responsive to EAN. These results indicated that EAN promoted N absorption and increased accumulation of N, P, K, and Ca in whole plant. PMID- 12132179 TI - Benchmarking: the backbone of successful performance improvement. AB - Although there are many valid applications for comparative data--sometimes simply "knowing where you stand" is valuable--it is time for home care agencies to use benchmarking that moves beyond that point. Benchmarking is integral to organizational and management strategies whose ultimate goal is to improve, and not simply measure, performance. Peer benchmarks can be a critical resource in addressing the full spectrum of management challenges. PMID- 12132180 TI - Responding to PPS claims denials and downcoding. AB - Home care agencies must become aware of the claims targeting process within medical review, undertake steps to prevent adverse claim determinations, and adjust strategies when responding to these decisions through the administrative appeals process. PMID- 12132181 TI - Strategic planning in home health care: the capital plan. AB - Home care organizations today have realized the importance of market-driven strategic planning. The strategies developed are unique to the specific organization doing the planning after careful consideration of the characteristics of its marketplace. In addition to a competitive analysis that analyzes the capabilities of competing organizations, the agency must consider the external dynamics occurring within the home care community. PMID- 12132182 TI - Good cash flow = come in fast, go out slow! AB - The formula for successful cash management in home care is a simple one: The agency must bring cash in as quickly as possible, while keeping expenditures at as low and slow a pace as possible. However, while the formula may be simple, success may be elusive unless agency administrators have a well-thought-out plan to handle cash management. PMID- 12132183 TI - Financial management of a hospice program. AB - Agencies interested in starting hospice programs or maximizing the benefits of existing programs need to implement and maintain accurate and effective internal cost accounting systems. Once established, a cost accounting system provides the administrators of the hospice program with information to prepare budget projections, perform break-even analysis, and develop other reports to assist in making sound business decisions to ensure success. PMID- 12132184 TI - One key to reasonably achieved outcomes: incentives. AB - Incentives have changed--in many cases, for the better. Innovation, discipline, and leadership are critical as organizations strive for profitability and excellence under a new set of rules. PMID- 12132185 TI - Strategic planning in five easy steps. AB - Every organization benefits from developing a strategic plan, but each organization needs to shape the plan and the development process to their unique needs and culture. PMID- 12132186 TI - Medicaid prospective pay--the next wave of reimbursement? AB - Given nationwide challenges with Medicaid reimbursement and the increasing pressures on states to curb escalating Medicaid expenditures, experimentation with pilot programs such as the one that VNAB implemented deserves serious consideration. PMID- 12132187 TI - [Ankle joint prosthesis implantation]. PMID- 12132188 TI - [Prosthetic replacement of the upper ankle joint]. AB - Results after total ankle arthroplasty in the 1970s and 1980s were poor. The outcomes of these surgeries deteriorated rather dramatically with time. Causes of failure were multifactorial, but the two main reasons for failure were constrained designs and cement fixation. Today, the design of total ankle arthroplasty is unconstrained and the fixation is uncemented. Total ankle arthroplasties are considered technically demanding procedures, with relatively high early postoperative complication rates. As yet, the ideal total ankle patient remains to be defined. Good alignment and ligamentous stability are essential. Osteonecrosis and profound osteoporosis are associated with poor results due to problems with bony fixation. Patients should be advised that the implant may fail and that this may require further surgery, including the potential need for an ankle fusion. The results of ankle fusions, although usually initially good, seem to deteriorate with time. Not uncommonly, patients frequently develop peritarsal degenerative joint disease several years after an ankle arthrodesis. Because of the associated pain and functional limitations that can follow ankle fusion, efforts to develop a workable total ankle replacement continue. At present, the long-term results of new designs are unknown. Today, total ankle arthroplasty should be limited to centers where patient volume and infrastructure allows critical review and prospective clinical trials to determine the factors leading to successful and unsuccessful outcomes. PMID- 12132189 TI - [Technique of intramedullary osteosynthesis of the clavicle with elastic titanium nails]. AB - This prospective controlled clinical trial was performed to assess fracture healing and clinical outcome after intramedullary nailing of midclavicular fractures. Within 3.5 years elastic-stable intramedullary nailing was performed in 62 patients with 65 midclavicular fractures. Surgery was performed in supine position. The ventral cortex of the proximal clavicle was opened using a 2.5 mm drill. The nail was advanced laterally under fluoroscopic control. If closed reduction failed, an additional incision was made to enable direct manipulation of the fragments. There were no infections, no implant displacements or refractures. Postoperatively, the mean subjective pain was significantly lower, and the range of motion improved. We observed one nonunion. The mean Constant score 6 months after hardware removal was 96.9 +/- 3.3 points. Intramedullary fixation of midclavicular fractures with an elastic titanium nail is a safe minimally invasive surgical technique, producing excellent functional and cosmetic results. PMID- 12132190 TI - [Fixed and functional decentering of the head of the humerus in patients with omarthrosis]. AB - AIM: To determine whether in patients with specific types of osteoarthritis of the shoulder not only a fixed but also a functional decentering of the humeral head exist. METHOD: The shoulder joints of 10 healthy volunteers and of 16 patients with osteoarthritis of the shoulder were examined in various arm positions, using an open MR scanner. After segmentation, 3D reconstruction of the scapula and humerus were performed and the position of the midpoint of the humeral head calculated relative to the center of mass of the glenoid cavity. RESULTS: At 30 degrees of abduction, 4 of 16 patients demonstrated a fixed posterior (12.9 +/- 2.8 mm) position and 8 (all patients with cuff-arthropathy) a fixed superior (6.6 +/- 2.6 mm) position of the humeral head. At 90 degrees of abduction the patients showed a significant (p < 0.001) combined decentering in the superior and posterior direction as compared to the healthy shoulders (functional decentering). CONCLUSIONS: This study demonstrates, that in most of the patients with osteoarthritis of the shoulder, a significant functional decentering occurred during abduction and external rotation, even if they showed no fixed decentering of the humeral head at 30 degrees of abduction. PMID- 12132191 TI - [Clinical and radiological results of surgical removal of periarticular ossifications after hip prosthesis implantation]. AB - The occurrence of heterotopic ossification (HO) is a well-recognized problem after total hip replacement. In a retrospective study, we investigated 32 patients who had undergone surgical excision of symptomatic HO followed by radiation with 7 Gy and nonsteroidal anti-inflammatory drug therapy between 1994 and 1999. The mean follow-up was 20 months (range: 12-60). Clinical and radiographic follow-up examinations included Harris hip score and classification according to Brooker. The preoperative Brooker class was III in 16 cases and IV in 16 patients. Comparison of the Brooker classification at follow-up revealed a statistically significant improvement (p < 0.0001; class 0:3, class I: 14, class II: 8, class III: 7 patients). In one case with symptomatic Brooker class III ossification, surgical reexcision of HO was necessary. A statistically significant increase (p < 0.05) in mean range of motion (ROM) was observed in flexion [preoperative: 57 degrees (+/- 26), follow-up: 83 degrees (+/- 21)], in abduction [preoperative: 17 degrees (+/- 12), follow-up: 24 degrees (+/- 9)], and in rotation (preoperative: 16 degrees (+/- 17), follow-up: 31 degrees (+/- 18)]. Comparison of preoperative Harris hip score (60 +/- 11) and Harris hip score at the time of follow-up examination (73 +/- 17) revealed a statistically significant increase (p < 0.0001) after treatment. At the time of follow-up examination, 18 patients (56%) assessed their pain symptoms as low but 6 patients (19%) reported strong pain symptoms. Nevertheless, the score at the time of examination (35 +/- 10) was statistically improved (p < 0.02) when compared to the preoperative score (30 +/- 8). Surgical excision of Brooker class III or IV heterotopic ossification with limited ROM followed by irradiation and anti inflammatory prophylaxis results in significant improvement in flexion, abduction, and rotation arc and significant reduction of HO in radiographic examination at follow-up, but pain relief was only satisfactory. PMID- 12132192 TI - [Modification of human osteoblasts by various analgesics]. AB - Analgesia plays a major role in the therapy of fractures. This raises the question whether frequently used analgetics as Tramadol and Diclofenac have negative effects on the healing of fractures. Human osteoblasts were isolated from human spongiosa and incubated with Diclofenac, Tramadol and without analgetic substance in an in vitro experiment. After 9 days the absolute number of cells as a marker for proliferation and their mitochondrial activity were quantified. The mitochondrial activity was measured using the metabolisation of XTT (sodium-3'-(1-[phenylamino-carbonyl]-3,4-tetrazolium)-bis(4- methoxy-6-nitro) benzene-sulfonic acid hydrate). Both drugs led to a concentration-dependent decrease of cell proliferation. Tramadol showed a significant effect at a concentration of 20 micrograms/ml, which is much higher than the therapeutical concentration of 0.25 microgram/ml in serum. Diclofenac decreased cell proliferation at a concentration of 6 micrograms/ml, having a therapeutical concentration of 1.5 micrograms/ml in serum. Vitality of cells had constant correlation to absolute number of cells (R = 0.95). Our results don't suggest any negative effects of Tramadol on the osteoblast activities in vitro. Diclofenac significantly decreased the proliferation of human osteoblasts at concentrations probably reachable in vivo. A prolonged healing of fractures under treatment with Diclofenac may be possible in critical situations (pseudarthrosis revision, callus distraction). PMID- 12132193 TI - [Differential therapy of radial head fracture: a critical analysis based on outcome of 53 patients]. AB - We investigated 53 patients with 57 radial head fractures (4 patients with bilateral fractures) treated between 1993 and 1998. We focused on patients with radial head fractures asking about (1) the relation between fracture type and therapy and (2) the correlation between chosen treatment and result. We saw the following fractures: Mason I: 3 cases, Mason II: 26 cases, Mason III: 11 cases, and Mason IV: 15 cases. Good results were achieved by 30 patients with 31 fractures, fair results by 8 patients with 9 fractures, and poor results by 13 patients with 14 fractures. Patients with a Mason I fracture achieved good results with functional therapy. Of the 26 Mason II fractures, 14 were treated with screws, 14% of whom had poor results subjectively. Six patients were treated with a K wire, titanium nail, or prevot nail, none of whom had poor results. Of 11 patients with a Mason III fracture, 10 were treated by resection of the radial head, and in 1 patient we implanted a prosthesis due to an intraoperatively detected elbow instability after resection and achieved good postoperative results. Only one patient (9%) had poor long-term results subjectively. Of 15 patients with a Mason IV fracture, 11 were treated by resection of the radial head: 5 patients (33%) had poor long-term results, only 3 of whom (20%) subjectively considered the results poor. PMID- 12132194 TI - [Fracture of the semilunar bone]. PMID- 12132195 TI - [Injuries of the alar ligaments in children and adolescents]. AB - Cervical spine trauma most commonly involves the lower parts in adults. In children lesions of the cervical spine can predominantly be found in the region of C1/C2 including ligament injuries at this level. However such injuries are difficult to detect and only few data are available concerning therapy and prognosis of atlantoxial ligament lesions. We report on two children suffering from isolated rupture of the alar ligaments. Both injuries were proven by magnetic resonance imaging which is recommended as the resource of choice for the evaluation of the cervical spine soft tissues in children. Although the biomechanic properties of the alar ligaments remain unclear non-operative treatment for the rupture of these ligaments seems to be adequate. In order to avoid neurologic symptoms or long term complications an immediate diagnosis is indispensable. PMID- 12132196 TI - [Isolated traumatic rupture of the subscapular muscle tendon as an adolescent injury]. AB - INTRODUCTION: Until now no case of a traumatic tear of the subascapularis muscle in children was described in the German speaking literature. Using the example of 2 cases of a 12 and 14 year boys youth history, clinic, diagnostics and therapy will be presented. METHODS: The accident happened in extension and external rotation of the arm without dislocation. Beside the complete tear of the SCP tendon in one case an accompanying expanded humeral flake fracture at the minor tuberosity was found. Under protection of the epiphysis line the refixation was performed using suture anchors. RESULTS: The post-operative control after 12 months showed a complete tendon healing, no arthritis or delayed bony ingrowth with return to full activity. CONCLUSION: Isolated traumatic SCP-tears can be occur also in young patients. As major consequence, it is necessary to perform a thorough clinical examination with additional apparative diagnostics (Sonography, MRI). This way, this rare but important lesion can be detected early and lead to adequate surgery without any delay. PMID- 12132197 TI - [Grisel syndrome--a trauma surgery rarity]. AB - Atlanto-axial fixed subluxation (Grisel's syndrome) is an uncommon complication of upper neck inflammatory processes and head and neck surgery. We present the case of a 6 year old patient who developed a Grisel's syndrome after an upper respiratory infection. X-rays and CT scans demonstrated a fixed C1 and C2 subluxation which was treated with reposition and application of a Halofixateur. We give a review of the pathogenesis, diagnosis and treatment of this rare syndrome. PMID- 12132199 TI - [Teutschlander disease. A rare benign differential diagnosis in proliferating space-occupying lesions of soft tissues]. AB - Tumoral calcinosis is a very rare benign soft tissue calcification. It occurs in all age-groups and prefers the shoulder, hip and elbow region as localisation. In most cases, local pain is the leading symptom. The aetiology remains unclear, so far, however, a certain connection to an imbalance of the calcium-/phosphate homeostasis is proposed. The adequate therapy is the complete surgical removal. Our presented case describes an extended occurrence at an unusual localisation and discusses characteristic signs in contrast of the differential diagnoses. PMID- 12132198 TI - [Delayed radial paralysis after Monteggia fracture--a case report]. AB - Concomittant lesions of neural structures represent a rare type of complications in Monteggia's fractures. In acute fractures spontaneous neurological remission usually occurs after reduction of the dislocated radial head. In the presented case a 33-year old man experienced a trady palsy of the posterior interosseus nerve 27 years after a Monteggia's fracture with the radial head left dislocated. Following a minimal trauma in badminton a neurological deficiency probably caused by distraction occurred and resulted in impairment of wrist extension and extension of the fingers. Initiated conservative treatment including intensive physiotherapy and electrotherapy for 4 months was unsuccessful. Consecutively the radial nerve was surgically exposed and released from an entrapping and thickened arcade of Frohse. The radial head was left dislocated. Full neurological recovery was obtained 9 months after surgery. PMID- 12132200 TI - [Injury caused by patient positioning in the surgical theater. Responsibility, documentation, legal liability]. PMID- 12132201 TI - [Determining responsibility for surgical positioning of the patient and malpractice for damage caused by patient position]. AB - The cooperation of surgeon and anaesthetist in positioning of the patient is subject to the principles of horizontal division of labour recognized in the interdisciplinary agreement and confirmed by the legislature: anaesthetist and surgeon carry out their respective tasks independently of each other, each bearing full responsibility for their own work (principle of strict separation of functions), they tailor their procedures to fit in with each other (duty of coordination), and each is entitled to expect and rely on due care in the other (principle of trust). In the case of conflict--when the best position for the specific intervention leads to a higher anaesthesiological risk--the principle of predominance of the actual requirements applies. If no agreement is reached it is incumbent on the surgeon to make the decision; this means that the surgeon bears the medical and legal responsibility for appropriate deliberation. Faults in organization are regarded under the law as faulty treatment. Anaesthetist and surgeon are each responsible for their own errors. According to the interdisciplinary agreements, positioning and checks on position are the task of the surgeon, while the anaesthetist is responsible for the "infusion arm". This does not exclude the possibility that anaesthetist and surgeon may agree on a different division of labour in the operating room. The patient bears the burden of proof that errors were committed in a case for damages. The doctor does, however, have to prove that the patient was correctly positioned. The demands of jurisdiction in terms of documentation of the positioning and of presentation of evidence are practically oriented and can basically be met. The same is true of the information supplied to the patient on the risk that positioning can cause harm. The doctor is obliged to supply evidence of the patient's substantive consent and the provision of information that this implies. PMID- 12132202 TI - [Systematic development of a scale for determination of health-related quality of life in multiple trauma patients. The Polytrauma Outcome (POLO) Chart]. AB - Even years after having sustained multiple injuries patients often suffer from its sequelae. These comprise restrictions in physical function, but also pain, social and psychological impairments. Although the Meran Consensus Conference in 1990 defined the contents of "quality of life" (QoL) measures in surgery, still no instrument is available for the valid assessment of all relevant QoL domains in multiple injured patients. This paper describes the systematic development of a modular instrument for the assessment of health related QoL. Within three phases (phase I: generation of items, phase II: item reduction, phase III: pre testing in 70 multiple injured and control patients) a questionnaire of 57 items was developed, which measures all relevant trauma-related aspects of QoL after acute hospital care. In combination with the Glascow Outcome Scale (GOS), the EUROQOL and the SF-36, the newly developed instrument builds the Polytrauma Outcome Chart (POLO-Chart) which will also be used as "Part E" for outcome assessment within the "Trauma registry" of the German Society for Trauma Surgery. In phase IV, the POLO-Chart will finally be validated in five trauma centres (Celle, Essen, Hanover, Cologne und Munich). PMID- 12132203 TI - [Outcome of long-term intensive therapy of surgery patients]. AB - OBJECTIVE: The treatment of severely ill patients remains a medical and human challenge. The aim of the study was to determine the survival rate of patients with prolonged intensive care unit (ICU) treatment. Additionally, the somatic, psychological, and social sequelae of the survivors should be determined. METHODS: Data of all patients who stayed for at least 30 consecutive days on a surgical ICU were evaluated with respect to age, sex, diagnosis on admission, APACHE II-Score, ISS, pre-existing diseases, therapeutic procedures, complications, organ dysfunctions, and mortality. The survivors passed a follow up examination after 35 +/- 14 months. This included somatic, psychological, and social parameters. RESULTS: Data of 101 patients were analysed (m/f: 78/23, mean age: 49.9 +/- 18.2 years, mean stay on ICU: 57 +/- 37 days, trauma patients: 46%). 31 subjects died on the ICU. Until the follow-up, another 24 patients deceased. Thus, the total mortality rose to 55%. Age, diagnosis on admission and severity of organ failure influenced the ICU mortality. Concerning the mortality after discharge, age, pre-existing morbidity and diagnosis on admission affected the outcome. 41 of the remaining 46 patients (89%) underwent the follow-up. Nearly half of them showed no or minor signs of impairment in any of the investigated areas. One third had severe handicaps. Trauma patients had the lowest mortality rates but showed worse results in rehabilitation. CONCLUSIONS: The mortality after prolonged ICU-treatment is substantially higher compared to average ICU patients. However, having survived the acute phase of the illness, successful rehabilitation in somatic, psychic as well as social terms could be performed to a considerable extent. This outcome is comparable to the one of other ICU populations. The results encourage to a consequent use of all medical options. PMID- 12132204 TI - [Retrospective analysis of healing problems after reamed and unreamed nailing of femoral shaft fractures]. AB - 168 fractures of the femoral shaft treated by intramedullary nailing were analyzed retrospectively. From 1986-1992 116 fractures had been treated with the reamed AO universal nail (RFN) and from 1993-1996 52 fractures with the AO unreamed femoral nail (UFN). In 24% of the RFN-group and in 2% of the UFN-group (p < 0.0001) open reduction of the fracture had been necessary. The time to radiological consolidation was similar in both groups (18.1 weeks +/- 6.1 vs. 18.3 weeks +/- 5.7, [mean +/- SD]). Delayed unions were less frequent in the RFN group than in the UFN-group (3% vs. 13%, p = 0.01). Non-unions occurred in the RFN-group in 4%, in the UFN-group in 8%, the difference is not statistically significant (p = 0.46). Fractures with impaired consolidation (delayed-unions and non-unions) in the RFN group were distributed randomly along the femoral diaphysis, whereas all 11 fractures with retarded healing in the UFN group were short transverse or oblique fractures localized immediately distal to the femoral isthmus. We believe that there is mainly a mechanical reason for this phenomen, in addition to fracture type and fracture localization the (insufficient) length of the unreamed nails might have impaired stability further. The different factors should be investigated in larger series. As a consequence we now treat transverse and short oblique fractures of diaphyseal femoral fractures distal to the femoral isthmus with a RFN whereas in other types and localizations of diaphyseal femoral fractures we continue to use the UFN with special attention to maximal nail diameter and length. PMID- 12132205 TI - [Effect of cyclical stretch on matrix synthesis of human patellar tendon cells]. AB - The significance of mobilization and loading for healing ligaments and tendons is generally accepted today. Local deformation of cells thereby represents the key stimulus for the cellular response. Less is known, however, about the effects of cyclic strain on the cellular and molecular level. The aim of the in vitro investigation was to determine the effect of cyclic mechanical strain on collagen type I and III and on fibronectin formation in human patellar tendon derived fibroblasts. Human patellar tendon derived fibroblasts from 5 donors (mean age 29.2 years) were cultured under standard conditions. Monolayers of subconfluently grown 3rd passage cells were stretched in rectangular silicone dishes with cyclic movement along their longitudinal axes. Cyclic strain (5%, 1 Hz) was applied for 30 min and 60 min, respectively. Carboxyterminal procollagen type I propeptide (P I-CP) and aminoterminal procollagen type III propeptide (P-III-NP) release was measured by a radio-immunoassay 6 h and 12 h after stretching. The release of fibronectin was measured by nephelometry following immunoreaction with a specific antiserum. Cells from each donor without any cyclic stretching served as controls. Compared with the controls, only cyclic stretching for 60 min resulted in a significantly increased release of P-I-CP and fibronectin after 6 h. The release of P-III-NP was significantly increased 12 h after 30 min of cyclic stretching as well as 6 h and 12 h after 60 min of cyclic stretching, respectively. We conclude that cyclic stretching causes a time-dependent, differential regulation of formation of fibronectin and collagen type I and III. This may effect the quality and thus the mechanical properties of healing tendon and ligament tissues. In order to improve current treatment protocols and to enlarge our knowledge of tissue healing, it is necessary to understand the cellular response to cyclic strain. PMID- 12132206 TI - [Mega-OATS. Technique and outcome]. AB - Big osteochondral defects in the weight-bearing zone of the medial respectively the lateral femoral condyle are still an unsolved problem especially in younger patients. The transfer of the posterior aspect of the femoral condyle was described as a salvage procedure. Mega-OATS is a technical improvement of the transfer of the posterior condyle-procedure. Essential advantages of the conventional OATS-technique are integrated in the Mega-OATS procedure, so that iatrogenic lesions of the transferred cartilage by press-fit-fixation and secondary hardware removal can be avoided. However, the Mega-OATS procedure itself remains a salvage procedure and should only be reserved for younger patients. The results of the first series of 17 patients (average follow-up 12 (5 19) months) showed an improvement of quality of life and a significant (p = 0.003) increase in the Lysholm-score. PMID- 12132207 TI - [First aid in sports injuries]. PMID- 12132208 TI - [Diagnosis and surgical therapy of diffuse pigmented villonodular synovitis of the hip joint]. AB - Pigmented villonodular synovitis (PVNS) of the hip is a monoarticular proliferative process involving the synovial membrane. A chronic inflammation as well as a neoplastic process have been proposed in the literature. PVNS is usually found in adults aged 20-50 years, without sex predilection. The knee is by far the commonest location, followed by the hip. We present a detailed case report of a 25-year-old man with PVNS of the hip. The physical examination was completely normal. Radiographs of the hip show erosions in the head and neck of the femur and the acetabulum. Magnetic resonance imaging showed a suspected malignant soft tissue mass involving the hip joint. The diagnosis of PVNS was confirmed by arthroscopy and biopsy, and the treatment of choice was an open synovectomy. One year after the operation the clinical examination was normal. PMID- 12132209 TI - [The BEHAC nail--a new intramedullary implant for managing 2-stage fractures of the humerus]. AB - The BEHAC-nail is a new implant for the treatment of complex humeral fractures, particularly for segmental fractures. It is an elastic intramedullary implant that is inserted retrograde into the distal humerus. The special feature of this nail is its proximal "loop design" which reduces the implant penetration at the proximal fixation site in the subchondral area of the humeral head in comparison to implants with tips such as Rush pins or Hackethal's nails. Thus, the risk of nail migration and perforation of the humeral head is less with the BEHAC-nail than with other nails. Another advantage of the "loop design" is that the humeral head fragment can be fixed adequately. Even short proximal fragments can be stabilized with this design. In contrast a short humeral head fragment cannot be held with implants like the UHN, HVN or Seidel's nail. The BEHAC-nail is a useful implant for segmental fractures of the humerus. PMID- 12132210 TI - [Anterograde intramedullary tibio-talo calcaneus arthrodesis (aIMTCA) with spongiosaplasty in pseudarthrosis]. AB - Pseudarthrosis occur in 65% of all ankle joint arthrodesis. From the therapeutical point of view we make a distinction between vital (hypertrophic) and avital (hypotrophic) respectively stable and instable pseudarthrosis. The hypotrophic forms demand an additional cancellous or bone grafting. Especially instable pseudarthrosis have to be treated with a biological osteosynthesis. In the hindfoot the so called compression arthrodesis made one's way. But there is still a discussion about the best method, intern or extern fixation. We report about a case of hypotrophic pseudarthrosis with a mal-position occurring after an ankle joint arthrodesis with a Charnley-Fixateur. A fusion of the ankle joint could be carried out with a proximal respectively anterograde intramedullary nail and allogene cancellous graft. PMID- 12132211 TI - [Corrective interventions after elbow para-articular fractures in childhood]. AB - Besides supracondylar fracture of the humerus there are several injuries of the elbow joint, which may lead to major disability. In this study 5 cases of corrective procedures are described after elbow fractures. Initially the lesions were overlooked. These were a fracture of the radial condyle, producing a pseudarthrosis, three cases of Monteggia fractures with persisting dislocation of the radial head in young children and a periarticular calcification issuing from an avulsion of the radial epicondyle and the radial capsule in a 13-year-old. All children had marked functional limitation of the elbow joint. The primarily overlooked fractures were corrected early. Various osteosynthesis procedures including movement and distraction extend fixator were employed. Overall, in all patients an almost complete movement of the joint at existing stability could be achieved. Transcondylar and Monteggia fractures should not be overlooked at the initial diagnosis as secondary operations for correction always have a less favorable outcome than the primary one. For the management of ankylosis of the elbow a movement extend fixator after distraction is a useful additional management. PMID- 12132212 TI - [Prevention of epidural fibrosis. Initial experiences with non-resorbable membrane]. AB - Scars around the neural structures after opening the spinal canal are common and severe problems in spine surgery. This paper presents the use of a special membrane to avoid epidural scarring in two cases. PMID- 12132213 TI - [Ruptures of the subscapular tendon. A diagnostic problem?]. AB - Isolated ruptures of the subscapularis tendon are rare injuries which are often missed initially. After recent treatment of 3 patients treated in our clinic typical courses and pitfalls are demonstrated. Due to the traumatic genesis of subscapularis tendon ruptures, lesions of the rotator cuff must be excluded generally by an adequate diagnostic concept. Conventional x-rays of the shoulder in 2 views to exclude a bony lesion, clinical and ultrasound examination are seen as a standard because these methods will allow the correct diagnosis in most cases. Typical clinical signs for a subscapularis lesion are the "lift-off-test" and the "napoleon sign." The ultrasound examination should be done statically and dynamically by comparing the injured with the uninjured shoulder. The typical view in case of a subscapularis tendon rupture is a thinned tendon overlying the humeral head. In our opinion nuclear magnetic resonance imaging should not be the first diagnostic procedure. It should be reserved for such cases with unclear clinical and ultrasound results. PMID- 12132214 TI - [Arterial vascular injury of the diaphragm caused by serial rib fracture]. AB - About 10% of all trauma patients sustain rib fractures. The average age is 58. Men are more often affected than women. Hemothorax, pneumothorax, and lesions of the lung are not uncommon. Very rare are injuries of the pericardium, aorta, and subclavia caused by fractured ribs. We present a very unusual case where a broken rib caused a severe diaphragmatic hemorrhage with a hemothorax and acute hypovolemia. The primary chest X-ray was thought to be without pathology. Arterial bleeding without exact localization could be found with computed tomography. Thoracotomy revealed the correct diagnosis and the cause of bleeding. PMID- 12132215 TI - [Staged medical rehabilitation of patients with chronic obstructive bronchitis]. AB - The author believes that adequate rehabilitation of patients with chronic obstructive bronchitis (COB) can be achieved only under staged approach, i.e. cooperation of outpatient, inpatient and sanatorium treatments. Each of the stages is characterized. This three-stage rehabilitation improves the patient's mobility due to higher physical performance and tolerance to dyspnea. PMID- 12132216 TI - [Features of predicting and control of clinico-functional states of patients with respiratory diseases during physical therapy]. AB - Changes in bronchial permeability in response to ultrasound and electric stimulation of the reflexogenic nasal zones by flow-volume forced expiration parameters were studied in patients with vasomotor rhinitis and bronchial asthma. The procedure employed step-by-step rise in intensity of the physical impact, change of polarity of the intranasal electrode. The rhinobronchial reflex proved heterogenic, it can lead to either deteriorated or improved bronchial permeability. In view of possible bronchoconstrictive reactions in the course of endonasal procedures it is necessary to furnish physiotherapeutic departments with devices of urgent control over clinico-functional condition of the patient. PMID- 12132217 TI - [Superhigh frequency electromagnetic fields and iodobromine baths in the rehabilitation of patients with cardiospasms]. PMID- 12132218 TI - [Method for micropolarization in treating various central nervous system diseases]. PMID- 12132219 TI - [Use of laser- and extremely high frequency magnetic therapy in the preoperative period before diskectomy]. AB - Electromagnetic therapy and tractions contributed to reduction of neurovascular structures compression evident not only from regression of clinical symptoms but also from improvement of regional hemodynamics, functional condition of the affected nerves and muscles of the limbs. This prediscectomy preparation appeared an effective conservative treatment for 69% patients. The rest patients benefited from such preoperative preparation which provided stabilization of the patients' condition before and after dyscectomy. PMID- 12132220 TI - [Rehabilitation of children with diffuse muscular hypotonia and neurophysiologic criteria of its effectiveness]. AB - Neuromapping, neuromyography, cerebrovascular mapping, cardiointervalography were conducted in children with the diagnosis "natal trauma of the cervical spine and vertebral arteries with ischemia of the reticular formation of the cerebral trunk in the form of myatonic syndrome". Adaptation reserves in the children were also studied. In addition to conventional methods, the treatment included kinesitherapy. The efficacy of the kinesitherapy was assessed. PMID- 12132221 TI - [Complex treatment of vertebrogenic pain syndrome in women during pregnancy]. AB - The proposed system of non-pharmacological treatment has been tried in 325 pregnant women with pain syndrome associated with spinal lesions (osteochondrosis, scoliosis, malformations, sequela of compression vertebral fractures, spondylolisthesis). The treatment combined orthopedic aids, relaxing massage, muscular relaxation, mobilization of functional blocks of intervertebral joints and pelvic junctions, therapeutic exercises. The above non-pharmacological system relieved vertebral pain syndrome partially or completely in 82% of the treated pregnant women. PMID- 12132222 TI - [Combined effect of musically-modulated electrical current and mineral drinking water from Khadyzhensky spring in experimental atherosclerosis]. AB - Male rats with experimental atherosclerosis drank mineral water (Khadyzhensky spring) and were exposed to music-modulated electric current. This combined treatment showed synergism of physical (current) and balneological (mineral water) factors providing lipolytic, antioxidant, stress-limiting and antiinflammatory intravascular effects and recovery of microcirculatory processes. PMID- 12132223 TI - [Adaptive reactions of rat testicular endocrine cells exposed to mineral drinking waters after radiation exposure]. AB - Early after irradiation (2 Gy) white male rats were given sulfate mineral water, sulfate mineral water with silicon, iodine-containing mineral water (iodine concentration 30 mg/l). It was found that sulfate mineral water alone or with silicon reduces the number of endocrine Leidig's cells, inhibits their functional activity, lowers blood level of testosterone. These effects were not observed after intake of iodine-containing mineral water. PMID- 12132224 TI - [Effect of boron-containing mineral waters on the lipid peroxidation status and antioxidant defense factors in experimental gastroduodenitis]. AB - We made experiments to study effects of effervescent hydrocarbonate mangesium calcium-sodium boronic mineral water on lipid peroxidation and factors of antioxidant defense in gastric tissue in gastroduodenitis. A course treatment with mineral water improves histomorphology of the gastric mucosa, stabilizes enzymes of the antioxidant defense system and down-regulates intensity of lipid peroxidation. PMID- 12132226 TI - [Experience in using acupuncture in children with opisthorchiasis]. PMID- 12132225 TI - [Differential approach to the use of drug ultraphonophoresis for scars]. AB - A differential approach to administration of drug ultraphonophoresis (gel contractubex, collitin, elastolitin) in scarry deformations is proposed. The analysis of 82 treatment outcomes has shown that early after trauma (on day 6-12) it is more beneficial to use ultraphonophoresis (UPP) of heparin-containing compounds improving blood rheology, e.g. lidase. This prevents development of pathological scars. Later, when the scar tissues has already formed, more effective is UPP of enzyme medicines with fibrinolytic properties (collitin, elastolitin). Differentiated choice of UPP in scar therapy reduces the time of rehabilitation considerably. PMID- 12132227 TI - [Use of glycyrrhiza extract in restoring local nonspecific defense of the lungs in patients with chronic obstructive bronchitis]. PMID- 12132228 TI - [Traction of the cervical spine region in osteochondrosis and phenomenon of cerebral circulatory insufficiency]. PMID- 12132229 TI - [Growth of bone and muscle tissue in children with delayed physical development under various physical loads]. PMID- 12132230 TI - [Ultraphonophoresis of omega-3 polyunsaturated fatty acids in the complex therapy of rheumatoid arthritis]. PMID- 12132231 TI - [Physical factors in the treatment of migraine]. PMID- 12132232 TI - [Mechanisms of the therapeutic and prophylactic effect of radon procedures in light of modern studies in the area of molecular biology]. PMID- 12132233 TI - [Conservative therapy of chronic cystitis patients]. PMID- 12132234 TI - [Long-term results of hydrogen sulfide balneotherapy in ischemic heart disease]. PMID- 12132235 TI - [Effectiveness of intermittent normobaric hypoxia in patients with bronchial asthma in various modes of chronotherapy]. PMID- 12132236 TI - [Pain--man's faithful companion]. PMID- 12132237 TI - [Multiple cerebral aneurysms]. AB - BACKGROUND: To define risk factors for the multiplicity of cerebral aneurysms, as well as clinical and therapeutical characteristics of patients with single aneurysms (SA) and multiple aneurysms (MA). METHODS: Retrospective study on 95 patients with SA and 22 patients with MA. For patients with SA and MA the following parameters were compared: gender, age, clinical state, aneurysmal localization and size, incidence of rebleeding and vasospasm, manner and outcome of treatment, preoperative interval, intraoperative rupture and postoperative complications. RESULTS: Aneurysms on anterior communicating artery existed in 37.4% of SA and in 17.8% of all MA (p < 0.05). As much as 44.2% of all aneurysms on middle cerebral artery and only 19% of all aneurysms on anterior communicating artery were associated with some other aneurysm (p < 0.02). The average size of SA was 15.4 +/- 11.8 mm, and 9.8 +/- 9 mm for MA (p < 0.05). Surgery was performed in 77.3% of patients with MA and 78.9% of patients with SA (p > 0.05), but complete surgical clipping was performed in 89.3% of patients with SA and in 47.1% of patients with MA (p < 0.01). Among operated patients with MA and SA, intraoperative rupture occurred in 36% and 17.6% of cases, respectively (p < 0.05) and ischemic postoperative complications were found in 29.4% and 17.3% of the cases (p > 0.05). Among 72.7% of all patients with MA and in 69.5% of all patients with SA the outcome was good, while among surgically treated patients it was good in 76.5% and 70.7% of cases, respectively. CONCLUSION: The treatment outcome was similar for patients with MA and SA, but complete operative treatment is significantly more frequent for SA. Multiple aneurysms were considerably smaller and with different anatomical distribution in relation to solitary aneurysms. PMID- 12132238 TI - [Endovascular procedures in the treatment of obstructive lesions of the brachiocephalic arteries]. AB - BACKGROUND: To assess the early effects, possible risks, and long term results of percutaneous transluminal angioplasty (PTA) of brachiocephalic trunk (BT) and subclavian arteries (SA). METHODS: During the period of 11 years, in 92 patients (57 males--62%, mean age 53.5 +/- 7.8 years) 93 PTA of SA/BT were performed; 70 (75%) lesions were stenosis, while 23 (25%) lesions were occlusions with mean diameter stenosis percent of 83.1 +/- 6.2%. Clinical indications were: vertebrobasilar insufficiency (n = 57), upper limb ischemia (n = 40), coronary steal syndrome (n = 4) and scheduled aorto-coronary bypass, using internal thoracic artery (ITA) (n = 4 asymptomatic patients). Mean lesion length was 22 +/ 8 mm. RESULTS: Eighty one (87%) out of 93 lesions were successfully dilated; all of 12 (13%) failures were due to unsuccessful recanalisation of occluded arteries. In 10 patients 10 stents were implanted (2 in BT and 8 in left SA). There were 6 (6.5%) procedural complications: 1 dissection, 1 thrombosis of the left SA, transient ischemic attack in 2 patients, and 2 cases of dislocation of atheromatous plaque from the right SA into the right common carotid artery. During the follow-up of 48 +/- 3 months, 16(20%) restenoses were treated by PTA (n = 7) or operatively (n = 9). Primary and secondary patency for all lesions treated during 11 years was 87% and 80%, respectively (stenosis: 97% and 89%; occlusions: 58% and 58%). CONCLUSION: PTA with or without stenting was relatively simple, efficient and safe procedure. It required short hospitalization with low treatment costs. If any of suboptimal results or chronic occlusions were present, the implantation of endovascular stents should have been considered. PMID- 12132239 TI - [Effect of primary and secondary below-knee amputation of war injuries on the length of hospitalization and rehabilitation]. AB - The results of below-knee amputations in 36 war wounded (mean age 35.42) were reviewed. The majority of the patients was wounded by land mines (94.4%). Most of them were between 25 and 35 years old. Bilateral amputation was done in 2.8% of cases. The amputation was performed on the day of wounding (primary below-knee amputation) in 30 (83.3%) amputees. Secondary amputation after the attempt to save the severely injured lower-limb was performed in 6 patients (16.7%) average 4.61 +/- 11.67 days after wounding. Reamputation was necessary in 6 cases (16.7%). Time period from the beginning of rehabilitation to the fitting of prosthesis, was 36.25 +/- 14.97 days for primary amputations, 32 +/- 17.8 days for secondary amputations and 68.66 +/- 33.52 days for reamputations. There was no significant correlation between the duration of rehabilitation to prosthetic management and the period between wounding and amputation (r = -0.102). The attempt to save the limb after severe below-knee injuries and the secondary amputation afterwards, did not significantly influence the ensuing rehabilitation and prosthetic works. PMID- 12132240 TI - Intrathecal synthesis of complement components C3c and C4 in the central nervous system infections with signs of the acute serous meningitis syndrome. AB - Two hundred and ten patients with meningismus and the infections of the central nervous system (CNS) with the clinical symptoms and signs of the acute serous meningitis syndrome, were divided in to groups according to etiology (enterovirus meningitis-ENTERO, serous meningitis various etiology-SM and tuberculous meningitis-TBC). Intrathecal synthesis (ITS) of C3c and C4 complement components and IgG were determined by the method of cerebrospinal indexes (I), to examine their role in differential diagnosis of this syndrome. Correlative study between the CSF/serum ratio (Q) for albumin (Alb) and QC3c and QC4 in patients with no proven ITS of this two complement proteins, and the comparative study of the increased value of C3cI and C4I (and IgGI) between the examined groups of the patients was done. Highly significant correlations were found between QAlb and QC3c (r = 0.89, p < 0.001) and QC4 (r = 0.85, p < 0.001). In 22.4% of the examined patients ITS of C3c and C4 were found. There was no difference in frequency of ITS of the two complement proteins between the examined groups, nor inside any particular group. TBC group had significantly lower (p < 0.05) intensity of ITS of C3c and C4 than MNG and ENTERO, and significantly higher intensity of ITS of IgG (p < 0.05) than the other tested groups. CSF index was confirmed as a valid method to detect intrathecal C3c and C4 production. Determination of ITS C3c and C4 could not be of great help in differential diagnosis in the acute serous meningitis syndrome. The intensity of ITS of C3c and C4, related to the intensity of ITS of IgG, could be of help in the determination of the duration of the disease. PMID- 12132241 TI - [Reliability in the detection of Escherichia coli based on their serogroup as a cause of sporadic and epidemic occurrence of enterocolitis]. AB - The purpose of this study was to determine the presence of virulence factors (heat-labile, heat-stable enterotoxin, verotoxin, invasiveness, localized, aggregative and diffuse adherence) among E. coli strains isolated from sporadic cases and outbreaks of enterocolitis, which belonged to serogroups characteristic for enteropathogenic E. coli. Serogroup was determined in 57.2% of 622 strains isolated from sporadic cases, and among them virulence factors were detected in 23.6%. Serogroup was also determined in 73.3% of 90 outbreaks isolates tested and virulence factors were detected in 97% of them. The detection rate of virulence factors rarely exceeded 50% among strains belonging to any of serogroup that was determined. The obtained data suggested that the identification of E. coli as a causative agent of enterocolitis by serogroup determination was a reliable method in outbreaks, but not in sporadic cases of this disease. PMID- 12132243 TI - [Reperfusion therapy in acute myocardial infarct]. PMID- 12132242 TI - [Determination of peracetic acid and hydrogen peroxide in a preparation]. AB - Iodometric and permanganometric titrations were used for determination of peracetic acid and hydrogen peroxide (H2O2) in the mixture. Two procedures were described and compared. Titrations could be done in only one vessel, in the same reaction mixture, when iodometric titration of peracetic acid was continued after the permanganometric titration of H2O2, (procedure A). Peracetic acid and H2O2, as oxidizing agents, reacted with potassium iodide in an acid medium, evolving iodine. This reaction was used for the quantitative iodometric determination of total peroxide in procedure B. H2O2 reacted with potassium permanganate in acid medium, but peracetic acid did not react under the same conditions. That made possible the selective permanganometric determination of H2O2 in the presence of peracetic acid. The procedure B was performed in two titration vessels (KV = 3.4% for peracetic acid, 0.6% for H2O2). The procedure A for iodometric determination of peracetic acid in one titration vessel after permanganometric titration of H2O2 was recommended (KV = 2.5% for peracetic acid, 0.45% for H2O2). PMID- 12132244 TI - [Effect of metabolic control on the onset and development of renal lesions in diabetes mellitus]. PMID- 12132246 TI - [The hepatitis G virus--a new hepatotropic virus]. PMID- 12132245 TI - [Febrile states and the encephalopathy syndrome]. PMID- 12132247 TI - [Immune defense mechanisms in infection with the tuberculosis bacillus]. PMID- 12132248 TI - [Multiorgan tuberculosis]. AB - Tuberculosis is an unusual infectious disease because of the latent period between the infection and the appearance of the disease may be prolonged for many weeks, months, or years as it is in case of the secondary tuberculosis. Tuberculosis in organs other than the lung has been observed for many years but has not always been recognized as tuberculosis, and it has been given many names. Extrapulmonary tuberculosis gained new importance, because it represented a progressively greater proportion of new cases. Multiple extrapulmonary sites were reported rarely except for one anatomical site, which was reported frequently. Extrapulmonary rates increase with age, so there are marked differences in age in specific rate patterns among the sites. Extrapulmonary tuberculosis occurred in respiratory organs other than lung, such as lymphatic, urogenital, and central nervous system, abdominal, osteoarticular, as well as tuberculosis of other organs such as skin, pericardium and endocrine glands. This case was reported to analyse clinical, morphologic and laboratory characteristics, method of diagnosis and the outcome in patients with multiorgan tuberculosis in order to explore the factors which might contribute to the decision making, concerning these forms of tuberculosis. Recent knowledge of pathogenesis was summarized as well as clinical presentation and the effects of cytokines produced by T lymphocytes and cellular population on antimycobacterial immune defences, and also susceptibility to tuberculosis. Mortality remains high and the treatment should start as soon as tuberculosis is suspected. PMID- 12132249 TI - [Successful resuscitation of a patient with prolonged tocolytic therapy and an emergency cesarean section]. AB - Ritodrine is the only medicament approved by FDA in the USA as well as in our country for prevention of the threatening preterm labor. Its adverse effects upon the respiratory and cardiovascular systems, including pulmonary oedema and myocardial ischemia, occur more frequently during the intravenous therapy than during the oral maintenance therapy. The aim of this report was to present a patient with cardiovascular adverse effects of ritodrine, who had her pregnancy terminated by an urgent cesarean section under general anesthesia. In the course of operation, the patient had two cardiac arrest (total of 70 min). Resuscitation was performed by direct and indirect heart massage. The patient's condition was stabilized during the next six hours. The patient was transferred to the coronary unit, where the treatment was continued for 30-days period, after which the patient was released home as completely recovered. PMID- 12132250 TI - [From the history of the Academy of Military Medicine: The 1st Military Hospital in Belgrade from 1836 to 1938]. PMID- 12132251 TI - Dental schools' relations with organized dentistry and accreditation: the Gies Report reconsidered. AB - The American Dental Education Association was formed with Gies' help in 1923 from four existing groups to better represent the interests of dental schools. An independent organization of examiners, practitioners, and educators, the Dental Education Council of America, started in 1909 to rate and later accredit schools. Later this group came under the influence of the ADA as the Council on Dental Education. Modern accreditation of dental education began in 1941, under the ADA Council on Dental Education, but has gradually achieved more independence. Gies favored the work of guiding and developing dental education under the hand of a small number of representative experts, and he cautioned against the sway of politics--a warning as valid today as it was three-quarters of a century ago. PMID- 12132253 TI - Identity theft. PMID- 12132254 TI - The Gies Report and Research. AB - In the eyes of the intellectually curious William Gies, dentistry and dental education in 1926 was mechanical, empirical, commercial, reparative, and isolated from other disciplines. The solution proposed in the Gies Report included making dental schools parts of universities and collaborative equals with medical schools, increasing full-time teachers, promoting graduate study, and especially, grounding dentistry in science. Early attempts by the American Dental Association to develop and support a research agenda floundered, and scholarship was left to the universities and the government, and more recently to commercial interests. To a disappointing extent, the separation of practice from science remains today as the technologies of the scientific spirit remain unfamiliar and vaguely threatening. PMID- 12132252 TI - The dental curriculum in Gies' time, now, and in the future. AB - Writing in the 1920s, William J. Gies was a champion for the importance of dentistry to general health, the scientific foundation for dentistry, and the need to develop and integrate an efficient curriculum. He argued that admissions standards should be made uniform and raised. He also proposed that the predoctoral dental curriculum should be reduced to three year's length by eliminating unnecessary material and improving efficiency. Gies called for available postgraduate experiences on an optional basis. Perhaps it is time to consider a three-plus-one model combining a three-year program focused on general dentistry with a mandatory fourth year along the lines of a postdoctoral general education experience. PMID- 12132255 TI - Eighty years of dental education in Canada. AB - The many similarities between dentistry and dental education in 1920 between Canada and the United States continue to exist today. These have lead to parallel development of dental education and practice and to extensive sharing between the two countries. However, the provincial rather than national approach to education and health care in Canada has not facilitated national outcomes. PMID- 12132256 TI - Dentistry and medicine, then and now. AB - Two factors have, at times, pushed dentistry and medicine together and pulled them apart. The factor acting to create a symbiosis is the common biomedical or scientific foundation for these fields. The factor causing independence deals with socio-cultural matters impacting on the professions and the public. These two factors will be examined at three points in time when the relationship between the two professions was significantly important for the welfare of the public: the 1920s and '30s, the 1960s and '70s, and our own time. Contemporary major discussion about the alignment of dental education, scientific advances, and societal needs point to a need for a new look at how dentistry and medicine relate to one another. PMID- 12132257 TI - What about dental care for people with mental retardation? A commentary. AB - People with mental retardation have untreated oral health needs that are comparable or even greater than those of individuals in the general population. But dental students have limited experiences in providing care for patients with special needs. In addition, there are numerous other barriers that current practitioners must overcome if we are to provide needed services for a population that increasingly resides in our communities. PMID- 12132258 TI - Is it ethical to involve patients in state board examinations? AB - It is argued that the state becomes an ethical agent when it requires that candidates for licensure perform dentistry on patients. As an ethical agent, the state is required to give full information, obtain true voluntary cooperation of patients, not expose patients to increased risk, and provide oversight while unlicensed dentists are practicing and follow-up care where untoward outcomes occur. The possibility of unsuccessful outcomes is known in advance, and there is no evidence showing that known exposure of individual patients to risk is compensated by decreased risk to patients generally. PMID- 12132259 TI - Ethical issues of performing invasive/irreversible dental treatment for purposes of licensure. AB - A case involving a patient sitting for an initial licensure examination is used to develop several ethical issues in detail. These include misalignment of those bearing the risks and dangers of such tests, lack of respect for patient autonomy, and compromises in the standard of care and avoidable and dysfunctional stress inherent in the examination system as it is currently conducted. PMID- 12132261 TI - Agents. AB - Although health care is inherently an economic activity, it is inadequately described as a market process. An alternative, grounded in organizational economic theory, is to view professionals and many others as agents, contracted to advance the best interests of their principals (patients). This view untangles some of the ethical conflicts in dentistry. It also helps identify major controllable costs in dentistry and suggests that dentists can act as a group to increase or decrease agency costs, primarily by controlling the bad actors who damage the value of all dentists. PMID- 12132260 TI - A response from the American Association of Dental Examiners. AB - The American Association of Dental Examiners supports a testing environments that include supervision of patient care. Extensive steps are taken in licensure examinations to ensure content validity through standardization by practice surveys, standards of competency, common core content, dental school curricula, and the limitations of practical constraints. Examining agencies report consistent, high collaboration among examiners. The examining community has developed comprehensive standards that compare favorably with standards in the testing community generally. When more reliable and valid alternatives to existing testing methods become available, they will be employed. PMID- 12132262 TI - Legal status of dentistry and licensure. AB - The lack of a national standard for dental education and lingering proprietary interests in the 1920s formed part of the context for development of licensure statues by individual states. In this report, Gies called for high standards on state boards and urged the National Association of Dental Examiners (now the AADE) to develop uniform statutes and examination practices. Significant progress has been made in the past seven decades (and especially recently) through regional examining agencies and in increasing the representation of membership on boards. The challenge posed by Gies to increase reciprocity has been refocused on credentialing, accreditation of specialists passed from state boards to the specialty groups, and uniform statues have proven elusive. PMID- 12132263 TI - Combating blindness. PMID- 12132264 TI - When is red eye not just conjunctivitis? AB - Red eyes are not 'just conjunctivitis' when there is significant pain or associated loss of sight. However, you are pretty safe to treat pain-free eyes and the normally seeing red eye with reasurance, lid hygiene advice and bland topical medication. But beware patients wearing soft lenses, and the red eyes of tiny babies or elderly patients, particularly those with apparent acute gastroenteritis. PMID- 12132265 TI - Casebook: flashes and floaters. PMID- 12132266 TI - Surgical treatment of cataracts. PMID- 12132267 TI - Managing dyslipidaemia: current concepts. PMID- 12132268 TI - Making the most of the accredited professional development programme. PMID- 12132269 TI - [Metastatic renal cell carcinoma]. PMID- 12132270 TI - [The value of tumor nephrectomy in metastatic renal cell carcinoma]. AB - In the case of an organ-confined RCC, tumor nephrectomy is the undisputed therapy of choice even though overall 5-year survival has not surpassed the 60% threshold. Further improvement will most likely have to await the development of more effective systemic treatment strategies. For an exclusively surgical therapy of metastatic RCC, tumor nephrectomy, sometimes in combination with metastasectomy, can be applied. However, more commonly used is a multimodality approach consisting of a cytoreductive operation followed by immunotherapy. Alternatively, one may select immunotherapy first followed by adjuvant nephrectomy in the case of a response, or one may proceed directly to immunotherapy only. Long-term survival does not exceed 5-10%, and patient selection appears to have a higher prognostic impact than any treatment strategy available. Concepts and progress in the field clearly are of increasing value for modern oncologic urologists. The current standard, a multimodality treatment of metastatic RCC, in which an operation becomes necessary at a certain point in time, easily justifies a central role for the urologic surgeon. PMID- 12132271 TI - [Value of metastases surgery in metastatic renal cell carcinoma]. AB - Metastasectomy in patients with renal cell carcinoma has to be considered as a palliative approach for symptomatic metastases (e.g., pathologic fracture) or as a curative approach in patients with the option for radical resection of all metastases. By modern perioperative management, even extended resections can be performed with limited morbidity and mortality. The survival rate is significantly higher after resection of pulmonary metastases than after resection of extrapulmonary metastases. Solitary metastases show a better prognosis than multiple metastases. Metachronous metastases that develop after a tumor-free interval of at least 12 months after tumor nephrectomy have a better prognosis than earlier metastases. For metastases that are resected with a curative intent, the best long-term results can be achieved after complete or radical resection. PMID- 12132272 TI - [Systemic immunotherapy of metastatic renal cell carcinoma and long-term outcome]. AB - Within the last 10 years, immunotherapy has progressively become an established treatment for patients with metastatic renal cell carcinoma. The cytokines interleukin-2 (IL-2) and interferon-alpha (IFN-alpha) are the substances that have shown the greatest effects. Both have been approved for the treatment of patients with metastatic renal cell carcinoma in Germany. Subcutaneous application of these frequently combined cytokines is the schedule of immunotherapy used most often in Germany. Combined cytokine therapy (IL-2 and IFN alpha) achieves response rates comparable to more aggressive immunotherapies. The retrospective analysis of treatment results from 66 patients with a follow-up of at least 5 years after the start of combined s.c. IL-2 and s.c. IFN-alpha +/- 5 fluorouracil (response classification: CR: 7, PR: 11, SD: 20, PD: 28) shows that the classification of the treatment results according to WHO criteria is the strongest predictor for survival compared with basic factors such as TNM status, grading, or number of metastatic sites. The combination of cytokine treatment with other treatment modalities (for example, surgical intervention) leads to a differentiated treatment according to the tumor status of the patient with metastatic renal cell carcinoma. Specific immunotherapies are still experimental. No approval has been granted for any of these treatments. Only standardization of these protocols can lead to a supplemental form of immunotherapy. Although several aspects of cytokine-based immunotherapy need further scientific evaluation, it is the treatment of choice for patients with metastatic renal cell carcinoma. However, for further progress in this field, prospective evaluation of immunotherapy for metastatic renal cell carcinoma is still needed. The German society for immunotherapy serves as a platform for this research. PMID- 12132273 TI - [Regional immunotherapy of metastatic renal cell carcinoma]. AB - We summarized the current literature concerning regional immunotherapy of pulmonary metastases in metastatic renal cell carcinoma and other malignancies using inhaled interleukin-2 (IL-2). Inhaled IL-2 therapy is associated with minimal toxicity and is effective in preventing progression in metastatic renal cell carcinoma, melanoma, and possibly other diseases such as breast cancer. Local (physiologic) use and systemic (pharmacologic) use of IL-2 are not mutually exclusive; a combination may be very appropriate in metastatic cancer. Local physiologic therapy intensifies treatment without intensifying toxicity. PMID- 12132274 TI - [Specific cellular immunotherapy of renal cell carcinoma. Current status and prospects]. AB - Renal cell carcinoma (RCC) is susceptible to immunomodulating therapies. This is proven by clinical responses to unspecific immunotherapy with cytokines. Understanding the mechanisms of antigen presentation and recognition by T cells enables us to expand T-cell clones which are capable of recognizing specific tumor-associated antigens (TAA). The use of dendritic cells (DC) in specific cellular immunotherapy could be beneficial because of their outstanding properties in antigen presentation and T-cell costimulation. In order to circumvent the escape of some tumor cells under T-cell pressure, polyvalent vaccination strategies should be developed. This goal can be achieved by either pulsing respective transfecting DC with tumor cell lysates, RNA or DNA libraries, or a pool of peptide antigens. Careful monitoring of the elicited T-cell response and quality assurance (GMP and GCP) are mandatory to establish a rationale for specific immunotherapy against RCC and to bring it from the bench to the bedside. PMID- 12132275 TI - [Manually-assisted laparoscopic donor nephrectomy]. AB - The number of patients suffering from terminal renal failure awaiting kidney transplantation has continued to increase. Since the number of available cadaver donor kidneys has largely remained unchanged, a special publicity campaign achieved that almost one-half of the kidneys transplanted in Basle come from live donors. We performed a laparoscopic live donor nephrectomy in 14 patients between March 1998 and August 1999. Despite good experience regarding transplant function and patient satisfaction, the hand-assisted technique was introduced in 1999 because of the long average operating time of 230 min (200-270) and warm ischemia time of 7.5 min (4-9). In the 33 operations performed up to now, the average operating time and warm ischemia time could be significantly reduced to 185 min (135-240) and 2.4 min (1.5-4), respectively. Most recently, the technique of retroperitoneoscopic live donor nephrectomy was successfully employed in 4 patients and could possibly represent the method of choice for the future. PMID- 12132276 TI - [Long term outcome after percutaneous sclerotherapy of renal cysts with polidocanol]. AB - Methodology and long-term results of treatment of uncomplicated kidney cysts by percutaneous sclerotherapy with polidocanol are presented. Between 1991 and 1998, 132 patients with 151 kidney cysts were treated by percutaneous sclerotherapy with polidocanol as the sclerosing agent. The average volume of the cysts was 288 ml. Over a mean period of 25.8 months, 118 patients with 132 cysts were followed up. In 56% of the cysts treated the cystic cavity disappeared completely, and in 30% the remaining volume was less than 10% of the initial volume. The existing symptoms before intervention remained unchanged only in four (3.4%) patients. The morbidity of the method was 9% and surgical reintervention was not necessary. There was no mortality. The mean hospitalization was 1.06 days. Percutaneous sclerotherapy of uncomplicated kidney cysts with polidocanol offers a high rate of success without the costs and invasiveness associated with laparoscopic surgery and also without the need for repeated interventions associated with sclerotherapy performed with ethanol. PMID- 12132277 TI - [Self-expanding permanent endoluminal stents (SPES) in therapy of benign ureteral obstruction. 6 years personal experience and review of the literature]. AB - At the urologic department of the Public General Hospital in Wels between 1994 and 2000, a self-expanding permanent endoluminal stent (SPES) was implanted in 13 women and 1 man (mean age: 52.2 years) with ureteral strictures. All patients had previously been treated unsuccessfully. An open operative approach was not possible because of various contraindications or because the patient had refused surgery. During a total follow-up period of 730 months (median: 59.9 months), primary patency lasted 669.3 months (median: 47.8 months). In nine patients no further intervention was necessary, and in four patients patency could be regained by additional interventions. In one patient the kidney had to be removed because of progressive renal insufficiency. Worldwide 98 cases of a SPES implant have been published so far. The patency rate was 68 of 98 stents (69.3%). The implantation of a SPES in the ureter for benign and complicated strictures represents a serviceable alternative to other urinary diversions. Additional clinical and experimental studies and long-term results are needed to make a definitive assessment of this method. PMID- 12132278 TI - [Postoperative pain therapy in urology. A prospective study]. AB - A sufficient analgesic treatment in the early postoperative period is important for the patients comfort level. Moreover, physical therapy for prophylaxis of pneumonia and thrombosis is better tolerated. In a prospective study, we compared two postoperative pain management regimens to establish a sufficient pain management without the need of additional costs or manpower. Of 215 patients undergoing major urologic surgery, 111 patients received on demand medication exclusively (group 1), whereas 104 patients were treated with basic analgesics combined with on demand medication (group 2). Pain intensity, side effects and subjective well being were evaluated with a visual analogue scale and a standardised interview. Pain intensity and side effects were significantly lower in group 2. Thus, with combined analgesic treatment, postoperative pain relieve can be achieved safely and without additional costs. PMID- 12132279 TI - [Microsurgical testicular spermatic cord denervation as therapy of chronic testicular pain]. AB - Diagnosis and treatment of chronic testicular pain (CTP) has been a difficult and often unrewarding clinical situation. Success rates of conservative and surgical measures rarely exceed 55% to 73% and 10% to 40%, respectively. We report on our experience with microsurgical testicular denervation as therapeutic option in CTP. Following an extensive preoperative work-up and a positive response to spermatic cord block, 25 patients underwent microsurgical testicular denervation. After a mean follow-up of 31.5 months 24/25 patients are painfree; no intra- or postoperative complications were encountered. In none of the cases testicular atrophy or testicular hydrocele was observed during postoperative follow-up. Microsurgical testicular denervation produces reliable and reproducible excellent therapeutic success rates and should be integrated in the management of CTP at an early stage. High success rates, however, require adequate and meticulous diagnostic workup of the patients with the spermatic cord block using saline and different local anaesthetics being an initial diagnostic armentarium predicting postoperative outcome. PMID- 12132280 TI - [Immunotherapy of metastatic renal cell carcinoma in Germany. An assessment of the current status]. AB - This survey was established to evaluate everyday use of interferon-alpha (IFN alpha) and interleukin-2 (IL-2) in metastatic renal-cell carcinoma (mRCC). Of 186 centers (with 2200 patients per year) that responded, 182 support immunotherapy by using it themselves (147 centers) or by referring patients (35 centers). Effectiveness and tolerance are the main reasons for use. 133 centers use IL-2 subcutaneously, 64 by inhalation, 24 use it locally or intratumorally. Continuous intravenous IL-2 is used in 13 centers only. Most centers use subcutaneous combinations of IL-2 and IFN-alpha, either alone or with 5-fluorouracil and/or isotretinoin; IFN-alpha/Vinblastin combinations, IFN-alpha-monotherapy, and IL-2 s.c.-monotherapy are used with similar frequency. Average treatment duration is 3 6 months. Maintenance therapy is used in responding patients in 118 centers. Subcutaneous and local application of IL-2 is standard treatment for mRCC in Germany and subcutaneous IL-2 and IFN-alpha represents the most frequently used combination. PMID- 12132281 TI - [Diagnostic imaging before kidney tumor operations]. PMID- 12132282 TI - [Preventive pacemaker stimulation in atrial fibrillation]. AB - Preventive pacing algorithms and the use of alternative or multifocal pacing sides are new approaches for treatment of paroxysmal atrial tachyarrhythmias. However, present data are not sufficient to define a new indication for pacemaker implantation in patients with refractory atrial fibrillation. Yet, preventive pacing should be predominantly performed either in patients with an established pacemaker indication or during controlled study projects. In patients undergoing cardiac surgery, biatrial overdrive pacing using temporary epicardial wires can be recommended for prevention of postoperative atrial fibrillation. PMID- 12132283 TI - Is left atrial appendage occlusion useful for prevention of stroke or embolism in atrial fibrillation? AB - Since in atrial fibrillation more than 90% of the thrombi are located in the left atrial appendage, an "elimination" of the left atrial appendage, either by resection or occlusion, seems an attractive alternative to oral anticoagulation. Although frequently regarded as an useless appendage, data from animal and human investigations show that the left atrial appendage may play an important role in the maintenance and regulation of the cardiac function, especially in arterial hypertension, atrial fibrillation, coronary heart disease, valvular heart disease and heart failure. Elimination of the left atrial appendage may impede thirst in hypovolemia, deteriorate hemodynamic responses to volume or pressure overload, decrease cardiac output and promote heart failure. Instead of preventing stroke, the consequences of left atrial appendage elimination may create new risk factors for stroke and thus might induce more harm than benefit to patients with atrial fibrillation. As long as the physiologic and pathophysiologic role of the left atrial appendage is not fully understood, left atrial appendage elimination should not be an alternative to oral anticoagulation. PMID- 12132284 TI - [Clinical aspects and molecular genetics of the Jervell- and Lange-Nielsen Syndrome]. AB - In contrast to the Romano-Ward (R-W) syndrome, the Jervell and Lange-Nielsen (J LN) syndrome is an autosomal recessive inherited disease characterized by QT prolongation in the electrocardiogram (ECG) and recurrent syncopal attacks which are also typical for the R-W syndrome, but also by congenital deafness. Recently, defect alleles in the genes for KCNQ1 and KCNE1 have been identified in patients with the J-LN syndrome. These genes may be causative for the R-W syndrome as well but in J-LN patients, they are only present in the homozygote or compound heterozygote form. In the present paper, we review the clinical and genetic similarities and differences of the J-LN and the R-W syndrome as well as the diagnostic and therapeutic management of these patients and their family members. PMID- 12132285 TI - [Long-term outcome of AV node modulation in 387 consecutive patients with AV nodal reentrant tachycardia]. AB - Aim of this study was to assess the long-term results of AV-node modulation in patients with AV nodal reetrant tachycardia. METHODS: From December 1991 until September 1999, AV node modulation (ablation of the fast pathway or ablation/modification of the slow pathway) was performed in 387 consecutive patients with clinically apparent AV nodal reentrant tachycardia. Follow-up data was available in 95% of patients with a mean of 41 +/- 26 months after ablation. RESULTS: Acute success rate was 97%. During long-term follow-up recurrence rate was 7.4% without any difference between fast and slow pathway ablation. Recurrence occurred in 23% of patients with persistent dual AV node physiology after ablation (modification of the slow pathway) in contrast to 3% without dual AV node physiology (ablation of the slow pathway) (p = 0.002). The presence of a dual AV node physiology after slow pathway modulation was the only predictor of recurrence during long-term follow-up. The complication rate was 5.7%. The incidence of complete heart block was 1% without any difference between fast and slow pathway ablation. CONCLUSIONS: Catheter modulation of the AV node for the treatment of AV nodal reentrant tachycardia is effective and safe. During long term follow-up, the recurrence rate was low. Modulation of the slow pathway is associated with a significantly higher recurrence rate than ablation of the slow pathway. PMID- 12132286 TI - Efficacy of antitachycardia pacing confirmed by stored electrograms. A retrospective analysis of 613 stored electrograms in implantable defibrillators. AB - The implantable defibrillator (ICD) is an established therapy in the prevention of sudden cardiac death by defibrillation of ventricular fibrillation. Another specific feature of the ICDs is antitachycardia pacing (ATP) of ventricular tachycardia. Several studies report success rates of ATP in 83 to 98% of cases. In clinical practice the success of terminating ventricular tachycardia is estimated only by automatic device analysis. Therefore the objective of this study was to confirm the efficacy of ATP based on the evaluation of stored electrograms. From the German Ventritex MD-register stored electrograms of 613 monomorphic ventricular tachycardias in 44 patients were analyzed retrospectively. The cycle length of the ventricular tachycardias was between 265 and 560 ms. The success rate of ATP-induced termination of the episodes reached 89.3%; another 2.3% of the ventricular tachycardias were accelerated by antitachycardia pacing into ventricular fibrillation. Left ventricular function did not influence the success rate, but the success rate was lower for fast ventricular tachycardias > 200/min (63.9%). For ventricular tachycardias < 150 bpm there was no restriction of ATP effectiveness. Of the episodes 72.9% were terminated by the first ATP burst. In these cases the duration of tachycardia was very short (11.9 +/- 2.8 s). Fifty-eight ventricular tachycardias (9.5%) had to be terminated by means of a shock, and only one case required 2 shocks. In patients with more than 10 episodes an individual therapy success > 90% was recorded for 80% of them. The very high success rate of the first ATP attempt in ICD therapy can be achieved with uniform programming, and is confirmed for ventricular tachycardias analyzed on the basis of stored electrograms. PMID- 12132288 TI - [Catheter ablation of ventricular tachycardia in the left ventricular outflow tract beyond the aortic valve]. AB - This case report describes successful catheter ablation of an ectopic focus in the left ventricular outflow tract just beyond the aortic valve. Extended pace mapping revealed a focus about 1 cm beneath the origin of the left coronary artery--above the base of the left coronary cusp. During a follow-up of 1 year no further spontaneous episode of ventricular tachycardia was documented. This case report describes an unusual localization of an idiopathic ventricular tachycardia and it demonstrates that radiofrequency catheter ablation can be done even in a critical position. PMID- 12132287 TI - Relationship between clinical and echocardiography-derived parameters and atrial activation as measured by the P-wave signal-averaged electrocardiogram. AB - P-wave signal averaging is used in an attempt to detect patients at risk of atrial arrhythmias, particularly atrial fibrillation. Although many studies utilized this method, data on the relationship between clinical and echocardiographic variables and signal averaged P-wave parameters are sparse. Thus, we performed a prospective study to evaluate the influence of important clinical and echocardiographic variables on P-wave parameters. The study included 100 probands without demonstrable cardiac disease. P-wave signal averaging was performed after echocardiographic examination in all subjects. Overall P-wave duration averaged 122 +/- 16 ms and correlated significantly with age, left atrial and left ventricular end-diastolic diameter on univariate analysis (r = 0.287, 0.393 and 0.375; p = 0.004, < 0.0001 and < 0.0001, respectively). On multivariate analysis, however, age was the only independent factor affecting signal averaged P-wave duration (p = 0.02). There were statistically significant differences in left atrial size and P-wave duration if probands were grouped according to their age. CONCLUSION: There is a significant relationship between signal averaged P-wave duration and age. This may be due to increased atrial fibrosis in elderly subjects. These data should be considered when signal averaged P wave duration is used as a risk stratifier in patients prone to atrial fibrillation. PMID- 12132289 TI - [Brugada syndrome or ARVD (arrhythmogenic right ventricular dysplasia) or both? Significance and value of right precordial ECG changes]. AB - We report about a 20-year old patient suffering cardiopulmonary resuscitation due to ventricular fibrillation. We diagnosed Brugada syndrome after exclusion structural heart disease and a positive Ajmalin test and implanted an ICD. In that there is a 20-30% familiar disposition, it was necessary that all family members undergo a cardiac examination. It was found that one brother and one sister presented the beginning of a right ventricular dilatation and a fibrolipomatous area in the anterior wall segment of the right ventricle. This result is compatible with a "concealed" arrhythmogenic right ventricular dysplasia (ARVD). As a prognostic indication we decided to implant an ICD prophylactically. The case report demonstrates the value of familiar examination of patients with an unclear ventricular arrhythmogenic event. PMID- 12132291 TI - [The value of hormone replacement therapy for prevention of coronary heart disease in women]. PMID- 12132290 TI - [Increased thrombocyte activation in dilated cardiomyopathy: a risk factor for development of ventricular thrombosis despite anticoagulant therapy?]. AB - HISTORY AND CLINICAL FINDINGS: A 48-year-old patient with dilated cardiomyopathy complained of dyspnea at rest, severe sleeplessness and a slight pain in the stomach. The clinical examination was normal except for a murmur at the apex of the heart. There was no evidence of edema or congestion of the jugular veins. INVESTIGATION: The echocardiography demonstrated a dilated left ventricle with severely compromised function. No ventricular thrombi were present at this time. Coronary artery disease was excluded by coronary angiography. Endomyocardial biopsies were obtained from the right ventricular septum. The immunohistological analysis of the endomyocardial biopsy specimens revealed pathologically increased lymphocytic infiltrates and increased expression of interstitial and endothelial MHC I and II antigens. Flow cytometric analysis of platelets surface antigens (P selectin, GP53, thrombospondin) was performed as a measure for intravasal platelet activation. Our patient compared to a healthy control group (> 4 SD) and to other patients with dilated cardiomyopathy (> 2 SD). A high grade increase of platelet activation was found. TREATMENT AND COURSE: ACE inhibitor, diuretics, spironolactone and digitalis were used to treat the heart insufficiency. Due to the severe left ventricular dysfunction phenprocoumone and aspirin were also prescribed. A follow-up echocardiography was performed 6 months later. Comparable to the first examination left ventricular contractility was found to be severely reduced. In addition, a marginal thrombus was now present in the left ventricle despite antithrombotic therapy. DISCUSSION: An increased platelet activation was found in the peripheral circulation of our patient with dilated cardiomyopathy. After 6 months, ventricular thrombi were found in the dilated ventricle, although aspirin and phenprocoumone had been administred. We speculate that an additional thrombotic treatment with clopidogrel is necessary in patients with dilated cardiomyopathy and increased platelet activation. PMID- 12132292 TI - ["The process was but too enjoyable". What one can learn from myths]. PMID- 12132293 TI - [Angiogenesis--anti-angiogenesis. Significance for tumor growth and metastasis]. AB - Angiogenesis plays an important role in the local growth and metastasis of a variety of malignant tumors. During the past three decades so-called angiogenesis factors have been discovered and characterized in more detail. These are the major contributors to angiogenesis. The term angiogenesis factors includes other functionally heterogeneous molecules. The best characterized angiogenesis factors are endothelial growth factors, such as "vascular endothelial growth factor" (VEGF), "fibroblast growth factor" (FGF), "platelet-derived growth factor" (PDGF), Angiogenin and interleukin-8 (IL-8). However, members of the family of matrix metalloproteinases are also included. The list of factors involved in angiogenessis and their receptors are increasing steadily as does the list of molecules with antiangiogenic capacity. The latter comprises the endogenous factors, Angiostatin, Endostatin, Thrombospondin-1 and 2, and chemical compounds, such as the Fumagillin derivative, AGM-1470, or inhibitors of matrix metalloproteinases. These have attracted more interest during the past years, since investigations on these molecules foster hope for new therapeutic strategies. Some of these antiangiogenic factors have already been used in various therapeutic approaches to influence tumor growth. While impressive results have been obtained in well controlled experimental animal models, the results of clinical studies in humans fall short of these successes. This is at least in part due to the fact that angiogenesis of tumors is a complex process based on the interaction of a variety of factors. The present article summarizes the current knowledge about tumor angiogenesis with a particular emphasis on antiangiogenic molecules. The presented data open new interesting therapeutic perspectives for future dermatology. PMID- 12132294 TI - ["Skin rejuvenation" by non-ablative laser and light systems. Literature research and overview]. AB - Currently, ablative laser therapy (with CO2/Er:YAG lasers) and deep chemical peeling are effective and promising methods of skin rejuvenation. The induction of collagen synthesis was observed after peelings with trichloroacetic acid or phenol as well as after treatments with the CO2 laser. In past years, the undesirable side effects and risks of these methods have led to intensified research in the fields of non-ablative facial rejuvenation and subsurfacing by means of ablative laser systems and intense pulsed light systems. The objective is to achieve selective, heat-induced denaturalisation of dermal collagen that leads to subsequent reactive synthesis but does not damage the epidermis. Recently, the results of numerous clinical and histological studies have indicated that these new technologies are successful. After critical review and assessment of current literature, we can say that in terms of their efficacy, non ablative methods are not a comparable alternative to ablative skin resurfacing. PMID- 12132295 TI - [Experiences with therapy of stage IV metastatic malignant melanoma with "Legha Protocol" polychemoimmunotherapy]. AB - BACKGROUND AND OBJECTIVE: Metastatic malignant melanoma (stage IV) is one of the most aggressive tumors. At the moment there is no safe therapy. Therefore the report of Legha et al., who achieved a rate of almost 10% of long-lasting complete remissions with a polychemoimmunotherapy using interleukin-2 and interferon-alpha in combination with cisplatin, vinblastine and dacarbazine, is a promising one. Because of these promising trends, we decided to treat our own patients with this therapy to examine the results, the side effects and the practicability on normal dermatological wards. PATIENTS/METHODS: From 1997 to 2000 we treated 28 patients with metastatic malignant melanoma with the polychemoimmunotherapy according to Legha's protocol. RESULTS: We achieved three complete (11.1%) and seven partial (25.9%) remissions (altogether 37%). Two of these patients are living relapse-free at the moment (7.4%). Three patients (11.1%) showed a stabilization of their disease, five patients (18.5%) had a mixed response and nine patients (33.3%) suffered progressive disease. CONCLUSIONS: The rate of complete and partial remissions was lower than those reported by Legha et al., however the rate of long-lasting complete remissions was almost identical. The follow-up time is still ongoing, so we have to limit our results to this period. We want to emphasize the practicability of this kind of therapy on normal dermatological wards in spite of the relatively high toxicity. PMID- 12132296 TI - [Lipoma of the lip. A rare differential diagnosis]. AB - We report on a 49 year-old female patient with a cherry-sized tumor of the lower lip, which over several years had steadily increased in size. The patient was psychologically affected by the appearance of the tumor and had difficulties to eat. We excised the tumor in local anaesthesia. Histology confirmed the clinical suspicion of lipoma. A total of four lipomas at this location have been reported world-wide. PMID- 12132297 TI - [Multiple lentigines (LEOPARD) syndrome. Case reports and review of the literature]. AB - The rarely occurring multiple lentigines (LEOPARD) syndrome represents a complex of skin, heart, skeleton and other malformations and is described in a 36-year old male and his 9-year-old daughter. With the occurrence of multiple lentigines, the diagnostic search for further malformations should always be undertaken. Its differential diagnosis and its pathogenetic and clinical aspects are discussed in this paper. PMID- 12132298 TI - [Kossard frontal fibrosing alopecia in a man]. AB - In 1994 Steven Kossard described a new and peculiar type of hair loss that he named postmenopausal frontal fibrosing alopecia. In 6 elderly women he observed a symmetric regression of the frontal hair line. Often the eyebrows were also affected. Histology showed lichen planopilaris. There were no clinical signs of lichen planus on the rest of the body. Since the original description by Kossard, several cases of frontal fibrosing alopecia have been described--almost all of them in elderly women. We report a man with frontal fibrosing alopecia of the Kossard type. PMID- 12132299 TI - [Simultaneous onset of pemphigoid and factor VIII antibody hemophilia]. AB - A 47-year old patient who had been suffering from hypertension and chronic renal failure for many years developed progressive extensive haemorrhagic erosions of the mouth within 3 months and less severe erosions of the genital and nasal mucosa. Additionally, subcutaneous haematomas developed spontaneously. Laboratory investigations demonstrated circulating antibodies against factor VIII while direct and indirect immunofluorescent microscopy showed discrete tissue-bound and circulating IgG reactive with the epidermal basement membrane in a pemphigoid like fashion. Immunoblot analysis of the patient's serum revealed an "atypical" IgG reactivity against a central portion of the extracellular domain of the BP180 antigen. These findings were unexpected, since the clinical aspect showed striking resemblance to (paraneoplastic) pemphigus. The patient developed life threatening complications. Eventually, reduction of circulating autoantibodies by a combination of plasmapheresis and subsequent immunosuppressive therapy led to a stable remission of both autoimmune bullous skin disorder and acquired haemophilia. PMID- 12132301 TI - [Restoration of moulages]. AB - Inappropriate storage threatens to destroy old collections of moulage. An experienced maker of moulage describes the time-consuming restoration of these three-dimensional illustrative objects, which requires specialist knowledge. Their teaching value in dermatology is particularly emphasised. PMID- 12132300 TI - [Skin blackish hyperpigmentation in 3 patients]. PMID- 12132302 TI - [Antimicrobial peptides: effector molecules of the skin as immune organ]. PMID- 12132303 TI - [Comment on the contribution by RE Schopf et a.: "Neurotrophic ulcer as a late sequela of trigeminal exharesis"]. PMID- 12132304 TI - [A startling beginning! Revolutionary local therapy of bullous pemphigoid?]. PMID- 12132305 TI - What's new on the dental scene? Browsing through the dental literature. PMID- 12132306 TI - The soft-tissue facial profile of patients with unilateral clefts of the lip, alveolus, and palate compared with healthy adults. AB - PATIENTS AND METHOD: In this study the soft-tissue profile of 84 patients aged between 16 and 29 years after complete rehabilitation of a unilateral cleft of the lip, alveolus, and palate was compared on lateral cephalograms with that of 58 adult probands with no cleft formation. RESULTS AND CONCLUSION: In the patient group, the thickness at nasion, the subnasal thickness, the upper alveolabial sulcus thickness, the upper lip prominence, and the mental thickness were less pronounced than in the control group. In contrast, the patients showed significantly higher values in the prominence and thickness of the lower lip. These results indicate a less dominant development of the upper lip and a more voluminous lower lip in comparison to the control group. In contrast, the length of the upper lip did not differ significantly between patients and controls. In both groups the facial integument was thicker in males than in females. The soft tissue anb-angle, the soft-tissue Holdaway II angle, and the nasolabial angle were significantly smaller in the patients than in the controls. Furthermore, the patients showed a distinctly more concave soft-tissue profile than the controls in measurements of the soft-tissue convexity angle. Thus, it is important for all surgeons involved in the rehabilitation of these patients to pay attention not only to an adequate length of the cleft-sided upper lip, but also to attaining a physiological alignment and reconstruction of the muscles, since the orbicularis oris muscle represents the most important component in the form and function of the lip. In addition, discontinuous muscular slings in the facial area may lead to severe midface growth disturbances. PMID- 12132307 TI - Evidence of intrafamilial variability of CBFA1/RUNX2 expression in cleidocranial dysplasia--a family study. AB - AIM: To investigate the phenotypical expression of an identical mutation of the CBFA1/RUNX2 gene within a family with cleidocranial dysplasia. PATIENTS AND METHOD: A five-member family underwent clinical examination. Two members, father and son, showed dissimilar symptoms of cleidocranial dysplasia. The two affected patients were examined for syndrome-typical symptoms, and the genotype was determined by molecular-genetic analysis. RESULTS: In both patients an identical missense mutation (G146R) in exon 2 of the CBFA1/RUNX2 gene was identified. In father and son the dental disturbances were similarly clearly expressed. However, the craniofacial skeleton of the son exhibited fewer dysostotic ossification features than that of the father. In the three clinically healthy family-members no mutation of the CBFA1/RUNX2 gene was found. CONCLUSION: In two patients with cleidocranial dysplasia an identical missense mutation in the CBFA1/RUNX2 gene leading to a divergent craniofacial phenotype was determined. The results indicate marked variability in the phenotypical expression of CBFA1/RUNX2 mutations. PMID- 12132308 TI - The frictional behavior of coated guiding archwires. AB - BACKGROUND: The vast range of orthodontic wires made of different alloys makes it increasingly difficult for orthodontists to judge them. Coated orthodontic wires form a group of innovative guiding archwires. MATERIAL AND METHODS: In the present in vitro study the frictional behavior of eight coated wires of different dimensions was investigated in archwire-guided canine retraction in the upper jaw. For this purpose five superelastic nickel titanium alloy wires (Titanol Low Force River Finish Gold and Gold 2: Forestadent, Pforzheim Germany; Titanol Superelastic tooth colored: Forestadent, Pforzheim Germany; BioForce Sentalloy Ionguard: GAC, Central Islip, NY, USA; NITI Imagination: GAC, Central Islip, NY, USA), two beta-titanium wires (TMA Low Friction Ionguard: Ormco, Glendora, CA, USA; TMA Low Friction Ionguard Purple: Ormco, Glendora, CA, USA), and one steel wire (Stainless steel Imagination: GAC, Central Islip, NY, USA) were selected. The coatings were made of Teflon or polyethylene, and by ion implantation. Three uncoated archwires (Rematitan Lite Dimple: Dentaurum, Pforzheim, German; Titanol Low Force River Finish: Forestadent, Pforzheim, Germany; BioForce Sentalloy: GAC, Central Islip, NY, USA) were used for comparison purposes. The force losses due to friction were measured using the Orthodontic Measurement and Simulation System (OMSS). RESULTS: The results indicated that all coatings can reduce frictional losses compared with an uncoated reference wire by the same manufacturer. Measured frictional losses ranged from 48.3-6.1%, with the Teflon coatings reducing the frictional losses to less than 10% in some cases. CONCLUSION: An unequivocal correlation between the surface roughness and frictional forces of the wires could not be verified by scanning electron microscopy. PMID- 12132309 TI - Corrosion susceptibility of lingual wire extensions in removable appliances. An in vitro study. AB - MATERIAL AND METHODS: The corrosion resistance of ten different round orthodontic wires as plastic-encased extensions was evaluated in vitro. Following the production of defined test samples with free as well as with acrylic-embedded wire segments, ten samples of each product were subjected to gap provocation by deflecting the free-running wire ends with an electric drive, while 10 samples remained provocation-free. The tests were run under standardized conditions (7 days, 37 degrees C, pH 2.3, 0.1 mol NaCl/CH3CHOHCOOH), and the findings were analyzed by stereo light microscopy and scanning electron microscopy. RESULTS: The following alterations were observed on the wire surfaces: a) crevice corrosion with substantial linear surface erosion and opaque discolorations, b) localized pitting corrosion, and c) mild surface erosion with translucent discolorations. The reactions of the test samples were similar in both those that were subjected to addition mechanical loading in the electrolyte and those that were not. Cr-Ni steels were susceptible to corrosion, whereas Co-Cr alloys as well as low-nickel manganese steels had only slight visible alterations, if any, in the embedded wire sections. CONCLUSIONS: Clinically occurring discolorations in the acrylic can be attributed to corrosion processes on the embedded wire extensions. These processes can be simulated and observed in vitro. During the processing of acrylic materials now customary in the orthodontic laboratory, crevices are inevitably formed between the wire and the acrylic, furthering the corrosion process. Our results suggest that, being more resistant to crevice corrosion, Co-Cr alloys or low-nickel steels can be considered as alternatives. PMID- 12132310 TI - Lingual technique--patients' characteristics, motivation and acceptance. Interpretation of a retrospective survey. AB - AIM: The aim of the present study was to collect personal data of patients treated solely with fixed lingual appliances. In addition, motivation and appliance acceptance were to be assessed. PATIENTS AND METHODS: The collective comprised 98 patients from our orthodontic clinic (C) and from an orthodontic practice (P). In addition to personal data such as gender, age, profession and marital status, the questionnaire covered the following parameters: initial source of information, motivation, phonetic-functional impairment, and subjective assessment of the treatment outcome. RESULTS: Statistical analysis of the personal data revealed a characteristic type of patient with a preference for lingual treatment: females below 40 years of age, whose preference for the lingual technique was based primarily on esthetic considerations. The main impairment (65%) due to the lingual appliance was reported to be injury to or irritation of the tongue and the restricted functional space for the tongue. The phonetic-functional adaptation period was 1-3 weeks. 99% of the patients were content or very content with the treatment outcome. 87% would recommend the lingual technique without reserve to relatives and friends. The majority of the patients would have refused orthodontic treatment with the labial technique. CONCLUSION: The results indicate that treatment with a fixed lingual appliance appeals to a characteristic patient group. In general, the level of acceptance among patients was positive and impairments during the adaptation period were tolerated after 1-3 weeks. The lingual technique should be integrated into routine orthodontic practice. PMID- 12132311 TI - A new bracket system for lingual orthodontic treatment. Part 1: Theoretical background and development. AB - BACKGROUND AND METHOD: This paper presents a new lingual bracket system differing fundamentally both in design and in manufacturing methods from existing appliances. Using state-of-the-art CAD/CAM technology, the two normally separate processes of bracket production and bracket positioning are fused into one unit. In this process, the demand for maximum individuality with simultaneously minimized space requirements is put consistently into practice. RESULTS AND CONCLUSION: Improved patient comfort, simplified rebonding in the event of bracket loss, and enhanced finishing precision are the improvements resulting for the course of treatment. In addition, bracket manufacture by a Rapid Prototyping technique permits direct transfer to clinically purposeful further developments. PMID- 12132312 TI - Rolf Berg: orthodontic treatment--yes or no? A difficult decision in some cases. PMID- 12132314 TI - Bioterrorism: what is and what may never be. Part 2. PMID- 12132313 TI - Lithium toxicity: two case reports. AB - Lithium has many medical and psychiatric uses. These include cluster and migraine headaches, alcoholism, impulsive behavior, and bipolar disorder. Toxicity from lithium can occur by overdose (intentional or accidental) or, more commonly, from alteration in its clearance by the kidney. We present two cases of lithium toxicity. The first is a 57-year-old male who presented with confusion, ataxia, and lethargy. The second case involves a 52-year-old female with bizarre behavior who was unable to care for herself. Both patients received dialysis and recovered without sequelae. PMID- 12132315 TI - [Prevalence of iron and iodine deficiency, and parasitosis among children from Arandas, Jalisco, Mexico]. AB - OBJECTIVE: To estimate the prevalence of iron deficiency, iodine deficiency and parasitosis in children attending the Instituto Alteno para el Desarrollo de Jalisco (Highlands Institute for Development of Jalisco State, INADEJ), Arandas, Jalisco, Mexico. MATERIAL AND METHODS: A cross-sectional study was conducted between 1997 and 1999, among 432 children aged 12 to 120 months attending the INADEJ. Measurements included hematological values, urine iodine concentration, and presence of parasites. Student's t test chi square tests were used for parametric and nonparametric analysis. RESULTS: The prevalence figures of anemia (20 vs 7.4%, p = 0.007) and iron deficiency (60.9 vs 44.4%, p = 0.02) were higher in preschool than in school children. Iodine deficiency was found in 29% (10.5% moderate or severe) and parasitosis in 47.2% of children, mainly E. histolytica (30.2%) and G. lamblia (28.9%). Low income, male gender and lack of social security policy holding were associated to parasitosis. CONCLUSIONS: The high prevalence rates of iron deficiency, iodine deficiency, and parasitosis, should be addressed by state health services with effective interventions to restrain these preventable diseases. The English version of this paper is available at: http://www.insp.mx/salud/index.html. PMID- 12132316 TI - [Acute respiratory infections in children attending a child day care center]. AB - OBJECTIVE: To assess the incidence of acute respiratory infections and bacterial colonization in children attending a daycare center. MATERIAL AND METHODS: A cohort study was conducted from April to Octuber 1999, among 85 children aged under four years, who attended the daycare center at Hospital Infantil de Mexico (Mexico City's Children's Hospital) "Federico Gomez". Acute respiratory infection incidence rates and quarterly point prevalence figures of nasopharyngeal colonization were obtained. Data were analyzed using descriptive statistics. RESULTS: A total of 85 children were studied (40 girls and 45 boys) during 9,090 children-days of follow-up. Three children had a history of atopia (3.5%), six a history of asthma (7%), and 39 (46%) were exposed to passive smoking. There were 258 events of respiratory tract infection for an incidence rate of 10.3 infections per person-year (95% CI 8.7-12.0). The main clinical syndromes were pharyngitis (95%), acute otitis media (3.5%), and bronchiolitis (1%). The incidence rates of otitis and bronchiolitis were 0.36 and 0.12 per child-year of observation, respectively. The prevalence figures of nasopharyngeal colonization for the three main bacteria were: S. pneumoniae 20.4%; nontypable H. influenzae 13%; and Moraxella catarrhalis 8%. CONCLUSIONS: Study results show a high prevalence of colonization due to invasive strains, as well as a two-fold incidence rate of acute respiratory infection, higher than those reported in community surveys. These results add to the description of this poorly documented infectious disease in Mexico. The English version of this paper is available at: http://www.insp.mx/salud/index.html. PMID- 12132317 TI - AIDS mortality trends in Mexico, 1988-1997. AB - OBJECTIVE: To assess the geographic distribution and trends of AIDS deaths for the 1988-1997 period in Mexico. MATERIAL AND METHODS: Crude and adjusted mortality rates were estimated for the 1988-1997 period. A trend test was performed using the simple linear regression method. Standardized mortality ratios (SMR) and years of potential life lost (YPLL) were calculated for each Mexican state. RESULTS: During the study period (1988-1997), there were 26,999 AIDS deaths in Mexico; 86.5% (23,354) of them were among men. The mean age at the time of death was 38.4 years for men and 37.7 years for women (p > 0.05). The crude AIDS mortality rate for the period of study was 3.02 cases (95% CI: 2.94, 3.06) per 100,000 inhabitants. The adjusted rate was 3.13 (95% CI: 3.09, 3.17), with 5.22 (95% CI: 5.16-5.29) for men and 0.82 (95% CI: 0.79-0.84) for women. The states with the highest SMR were: Baja California (SMR: 248.69; 95% CI: 234.02 263.36), Mexico City (SMR: 220.74; 95% CI: 215.57-225.91), and Jalisco (SMR: 169.16; 95% CI: 162.88-175.44). Similarly, a Potential Lost Life Years Index (PLLYI) analysis by state showed a greater risk of premature AIDS mortality in the same states [Baja California (PLLYI index: 236.33; 95% CI: 233.97-238.68), Mexico City (PLLYI: 194.68; 95% CI: 193.88-195.48), and Jalisco (PLLYI: 170.69; 95% CI: 169.60-171.79)]. CONCLUSIONS: Mortality trends indicate that AIDS mortality in Mexico increased by an annual rate of 23% between 1988 and 1997. The adjusted AIDS mortality rate increased from 0.75 per 100,000 in 1988, to 4.20 per 100,000 in 1997, with the largest burden of mortality in men (male to female ratio of 6:1). We therefore expect that a decreasing effect on AIDS mortality trends will be observed in the next years. The English version of this paper is available too at: http://www.insp.mx/salud/index.html. PMID- 12132318 TI - [Risk factors associated with systemic lupus erythematosis in a Mexican population]. AB - OBJECTIVE: To assess risk factors associated with systemic lupus erythematosus (SLE) in the Mexican population. MATERIAL AND METHODS: A case-control study was conducted on June 1996, at the Reumathology Clinic of Hospital de Especialidades del Centro Medico Nacional Siglo XXI (HE CMN), Instituto Mexicano del Seguro Social, in Mexico City. Cases were one hundred thirty subjects with four or more SLE criteria and disease evolution of +/- 5 years. Controls were hospitalized patients with acute diseases but without autoimmune diseases. Cases and controls were matched 1:1 by age and gender; both groups were evaluated by direct interview through a structured questionnaire. The following risk factors were assessed: genetic family history of SLE and connective tissue disease; socioedemographic (ethnicity, geographic distribution, education, monthly income); hormonal (use of oral contraceptives, replacement therapy and gynecoobstetric background); environmental (use of hair products, living with dogs, bacterial/viral infections, and allergies). Statistical analysis consisted of odd ratios (OR) with 95% confidence intervals (CI) and multivariate analysis using logistic regression. RESULTS: The multivariate model showed association with family history of SLE (OR 4.2, CI 95% 1.17-15.2), family history of connective tissue disorder (OR 2.6, CI 95% 1.15-4.5), use of oral contraceptives for more than one year (OR 2.1, CI 95% 1.13-4.3), repetitive pharyngitis (OR 2.1, CI 95% 1.18-3.6), and use of medications (OR 5.0 IC 95% 1.62-21.6). No association was found with socieconomic status, hair dye products, asthma, or allergies. CONCLUSIONS: Genetic factors, such as family history of SLE and connective tissue disease in first-degree relatives, persist as important factors in the development of SLE. Other factors, such as use of some drugs, oral contraceptives, and repetitive pharyngitis, may also favor the onset of disease in genetically susceptible hosts. The English version of this paper is available at: http://www.insp.mx/salud/index.html. PMID- 12132319 TI - Hemoglobin measured by Hemocue and a reference method in venous and capillary blood: a validation study. AB - OBJECTIVE: To assess the comparability of hemoglobin concentration (Hb) in venous and capillary blood measured by Hemocue and an automated spectrophotometer (Celldyn) and to document the influence of type of blood (capillary or venous) and analysis method on anemia prevalence estimates. MATERIAL AND METHODS: Between February and May 2000, capillary and venous samples were collected from 72 adults and children at Hospital del Nino Morelense (Morelos State Children's Hospital) in Cuernavaca, Morelos, Mexico, and assessed for Hb using the Hemocue and Celldyn methods. Estimated Hb levels were compared using the concordance correlation coefficient and Student's t test for paired data. The sensitivity and specificity for anemia diagnosis were estimated and compared between type of blood and method of assessment. RESULTS: Capillary blood had higher Hb (+0.5 g/dl) than venous blood in adults and children, as did samples assessed by Celldyn compared to Hemocue (+0.3 g/dl). Specificity to detect anemia was adequate (> 0.90) but sensitivity was low for capillary blood assessed by Hemocue (< 0.80). CONCLUSIONS: The difference in Hb between venous and capillary blood is likely related to biological variability. Hemoglobin concentration in capillary blood assessed by Hemocue provides an adequate estimation of population anemia prevalence but may result in excess false negative diagnoses among individuals. The results of this study stress the importance of sample collection technique, particularly for children. Method of analysis and sampling site need to be taken into consideration in field studies. The English version of this paper is available too at: http://www.insp.mx/salud/index.html. PMID- 12132320 TI - Phylogenetic analysis of the envelope protein (domain III) of dengue 4 viruses. AB - OBJECTIVE: To evaluate the genetic variability of domain III of envelope (E) protein and to estimate phylogenetic relationships of dengue 4 (Den-4) viruses isolated in Mexico and from other endemic areas of the world. MATERIAL AND METHODS: A phylogenetic study of domain III of envelope (E) protein of Den-4 viruses was conducted in 1998 using virus strains from Mexico and other parts of the world, isolated in different years. Specific primers were used to amplify by RT-PCR the domain III and to obtain nucleotide sequence. Based on nucleotide and deduced aminoacid sequence, genetic variability was estimated and a phylogenetic tree was generated. To make an easy genetic analysis of domain III region, a Restriction Fragment Length Polymorphism (RFLP) assay was performed, using six restriction enzymes. RESULTS: Study results demonstrate that nucleotide and aminoacid sequence analysis of domain III are similar to those reported from the complete E protein gene. Based on the RFLP analysis of domain III using the restriction enzymes NIa III, Dde I and Cfo I, Den-4 viruses included in this study were clustered into genotypes 1 and 2 previously reported. CONCLUSIONS: Study results suggest that domain III may be used as a genetic marker for phylogenetic and molecular epidemiology studies of dengue viruses. The English version of this paper is available too at: http://www.insp.mx/salud/index.html. PMID- 12132321 TI - Gender-related family head schooling and Aedes aegypti larval breeding risk in southern Mexico. AB - OBJECTIVE: To investigate if family head genre-associated education is related to the risk of domiciliary Aedes aegypti larval breeding in a dengue-endemic village of Southern Mexico. MATERIAL AND METHODS: A family head was considered to have a low education level if he/she had not completed elementary school. To estimate larval breeding risk within each household, a three-category Maya index was constructed using a weighted estimation of controllable and disposable domestic water containers. A socio-economic index was constructed based on household construction characteristics. RESULTS: Low-level education of either family head was associated to higher larval breeding risk. Households with low-educated mothers had more larval breeding containers. These associations persisted after adjusting for household socio-economic level. CONCLUSIONS: These results indicate that households with female family heads with low education levels accumulate more containers that favor Ae. aegypti breeding, and that education campaigns for dengue control should be addressed to this part of the population. The English version of this paper is available too at: http://www.insp.mx/salud/index.html. PMID- 12132322 TI - [Association between cardiovascular disease and anti-Chlamydia pneumoniae antibodies]. AB - OBJECTIVE: To evaluate the association between cardiovascular disease (CVD) and antibodies against Chlamydia in Mexican population. MATERIAL AND METHODS: A cross sectional study was conducted from August 1988 to April 2000, at the Immunology and Infectology Research Unit of Hospital de Infectologia, Centro Medico Nacional La Raza (CMNR)--and at the Cardiovascular Surgery and Circulatory Care, Hospital General CMNR, Instituto Mexicano del Seguro Social (IMSS). Study subjects were 70 CVD hospitalized patients, older than 30 years, from both genders. Serum IgG and IgM antibodies against C. psitaccii, C. trachomatis and C. pneumoniae were determined by microimmunofluorescence in study subjects and compared with those from 140 healthy individuals, matched by age and sex. Simple random sampling was used, for an expected prevalence of 50% and a 99% confidence level; the sample size was 110 subjects. The chi-squared test and odds ratios were used to compare proportions. RESULTS: IgG antibodies against C. pneumoniae were found in 94.3% (66/70) patients, as compared to only 37% (52/140) of healthy individuals (p < 0.001). CONCLUSIONS: An association between IgG antibodies against C. pneumoniae and CVD was found. This finding warrants further studies to evaluate the role of C. pneumoniae as a predictor of CVD. The English version of this paper is available at: http://www.insp.mx/salud/index.html. PMID- 12132323 TI - [Significance of prostaglandins in hepatic amebiasis]. AB - Prostaglandins are important mediators of inflammation; they also play a role in the regulation of both lymphocyte and macrophage functions. Hamster's liver lesions resulting from intraportal or intrahepatic inoculation of living Entamoeba histolytica trophozoites are characterized by an acute inflammatory response, where trophozoites are successively surrounded by polymorphonuclear leukocytes, lymphocytes, and macrophages. Incapability of these cells to counteract amebic invasion has been demonstrated in some studies. Prostaglandin E2 (PGE2) has potent effects on immune cells; its participation in amebic liver abscess has been reported recently. This paper presents a review of recent discoveries on biochemical mediators produced during inflammation due to Entamoeba histolytica infection, and their possible role in establishing the host's immune response. The English version of this paper is available at: http://www.insp.mx/salud/index.html. PMID- 12132324 TI - [Ideas, beliefs, and perceptions about health. History summary]. AB - To get a closer approach to the concept of health, this paper reviews the ideas, beliefs, and perceptions about health that have been accepted by Eastern and Western cultures. Health conceptions by philosophers, physicians, and thinkers are presented, particularly as they relate to substances, forces, and elementary biological principles of the human body that determine human life and fate. Analysis of these conceptions makes it possible to recognize different definitions of health as a status of balance, harmony, and equity. On the contrary, disease has been defined with the antonyms of health words. The ecological theory of "resilience" may be useful to analyze the adaptive capacity of ecosystems and to suggest ideas for physiological research on processes leading the human body to a new status of balance with the external environment, and thus define the blurring boundaries between health and disease. PMID- 12132325 TI - [Mortality statistics in Mexico: deaths registered in the year 2000]. PMID- 12132326 TI - [International classification of daily living function, disability and health status]. AB - The World Health Organisation issued the international classification of impairments, disabilities and handicap in 1980. As no Hungarian translation appeared, it was mainly utilised in Hungary in education, with good results. Based on practical application in foreign countries and on positive and negative experiences, as a result of an almost ten years development process a new version was published, entitled the International classification of functioning, disability and health. The new classification, employing qualifiers, enables the description of both the nature and the magnitude of the problems, thus it may be applied in rehabilitation medicine practice. It may also support the establishment of still missing disability data bases. Applying together with quality of life assessments, it may serve obtaining better knowledge of the persons in need of rehabilitation. PMID- 12132327 TI - [Health risks for the Hungarian population due to the nuclear accident in Chernobyl]. AB - The review presents the health risks upon irradiation, refers to the radiation sources and doses, the biological effects of ionizing radiation, the dose response relationships, the risk assessment. Based on the dosimetry data as presented at the scientific sessions held at the Hungarian Academy of Sciences on the 15th anniversary it is concluded that no health effects occurred and it was not expected at detectable levels according to the risk coefficients of the International Commission on Radiological Protection. PMID- 12132328 TI - [Thrombolytic therapy in acute ischemic stroke with streptokinase]. AB - AIM AND METHODS: In an open, observational study, 40 consecutive ischemic stroke patients eligible for thrombolytic therapy using the combined ECASS/NINDS inclusion criteria have been treated intravenously with 1.5 M units of streptokinase. The therapeutic window was 3 hours or shorter. RESULTS: The safety analysis documented a low rate (5%) of intracerebral hemorrhages, and an additional 13% rate of hemorrhagic transformation of the initial infarction. Two patients died due to intracerebral bleeding. The efficacy of the SK thrombolysis was significant in 53% of the patients (the mean of the improvement on the NIH stroke scale was 15 points), while an other 42% of the patients achieved only the mean of 4 points improvement on their NIHSS score. DISCUSSION: These results are of the same magnitude, as those documented in the NINDS trial with rt-PA. Time window rather than the thrombolytic agent itself seems to be the decisive factor for successful thrombolysis. CONCLUSION: The good safety profile of SK in acute stroke using the ECASS/NINDS criteria, and the cost-effectiveness of the drug underline the necessity of a new SK trial with the recently accepted inclusion and exclusion criteria. PMID- 12132329 TI - [Correlation of Barrett's esophagus and colonic adenomas]. AB - INTRODUCTION: According to the case-control studies, colon adenomas are not more frequent as usual in patients with Barrett's oesophagus. AIM: Analysis of the relationship of Barrett's oesophagus with colon adenomas through a case presentation. METHOD: The 62-years old male patient has been treated with pantoprazole for reflux oesophagitis and histologically proven short-segment Barrett's oesophagus. Pantoprazole treatment was started and the patient became symptom-free. In 1997 colonoscopy was performed for rectal bleeding and 4 recto sigmoideal polyps were removed (tubular adenomas). After 4 years of PPI treatment, laparoscopic fundoplication was performed but after one year of symptom-free period, PPI was started again because of relapse. The patient was followed-up for 5 years. The histological examinations included also Ki67 and p53 antibody staining. RESULTS: On control colonoscopies, small rectal and sigmoid polyps were found. The retrospective histologic examination of the polyps revealed the increase of Ki-67-positive cells from 2-5% to 45-55% during the 5 years of follow-up, which is higher than the values found in controls and may express the increasing proliferative activity of the epithelium. CONCLUSION: Although according to the literature, Barrett's oesophagus does not increase the risk of colon adenomas, this may occur in individual cases, warranting close follow-up. PMID- 12132330 TI - [Dr.Karoly Csepai--life work and its current significance for giving expert opinion in insurance medicine]. PMID- 12132331 TI - [ATRA (all-trans retinoic acid) in chronic myeloid leukemia?]. PMID- 12132332 TI - [Physician-patient relations and patient advocates]. PMID- 12132333 TI - [Comment on the paper "Hungarian physicians in Vienna"]. PMID- 12132334 TI - Contexts and culture in psychological research. PMID- 12132335 TI - Parenting self-efficacy among Japanese mothers: qualitative and quantitative perspectives on its association with childhood memories of family relations. PMID- 12132336 TI - Learning models in different cultures. PMID- 12132337 TI - Achievement motivation across cultures: some puzzles and their implications for future research. PMID- 12132338 TI - The culture and contexts of achievement motivation. PMID- 12132339 TI - Implicit theories of intelligence across academic domains: a study of meaning making in adolescents of Mexican descent. PMID- 12132340 TI - [Mental health among indigenous population. An international view]. PMID- 12132341 TI - Ecological benefits of contaminated sediment remediation. AB - Contaminated sediment has been identified as a source of ecological impacts in marine and freshwater systems throughout the world, and the importance of the contaminated sediment management issue continues to increase in all industrialized countries. In many areas, dredging or removal of sediments contaminated with nutrients, metals, oxygen-demanding substances, and persistent toxic organic chemicals has been employed as a form of environmental remediation. In most situations, however, the documentation of the sediment problem has not been quantitatively coupled to ecological impairments. In addition, the lack of long-term, postactivity research and monitoring for most projects has impeded a better understanding of the ecological significance of sediment contamination. Establishing quantitatively the ecological significance of sediment-associated contamination in any area is a difficult time- and resource-consuming exercise. It is, however, absolutely essential that it be done. Such documentation will likely be used as the justification for remedial and rehabilitative action(s) and also as the rationale for proposing when intervention is necessary in one place but not another. Bounding the degree of ecological impact (at least semiquantitatively) provides for realistic expectations for improvement if sediment remediation is to be pursued. It should also provide essential information on linkages that could be used in rehabilitating other ecosystem components such as fish or wildlife habitat. The lack of information coupling contaminated sediment to specific ecological impairments has, in many instances, precluded a clear estimate of how much sediment requires action to be taken, why, and what improvements can be expected to existing impairment(s) over time. Also, it has likely resulted in either a delay in remedial action or abandonment of the option altogether. A clear understanding of ecological links not only provides adequate justification for a cleanup program but also represents a principal consideration in the adoption of nonintervention, alternative strategies. In developing this understanding, it is important to know not only the existing degree of ecological impairment associated with sediment contaminants but also the circumstances under which those relationships and impacts might change (i.e., contaminants become more available and more detrimental). Because contaminated sediment remediation often costs millions of dollars per area, adequate assessment, prediction, and monitoring of recovery would seem obvious. However, experience has shown that this is not always the case, particularly for prediction and monitoring of ecological recovery. This scenario would never happen in the business world and should not occur in the environmental management field. PMID- 12132342 TI - Environmental fate of triclopyr. AB - Triclopyr is nonpersistent in surface water. It has limited mobility and low to medium persistence in soil. Considering its adsorptive characteristics and that it dissipates via multiple pathways, such as photolysis, plant metabolism, and microbial degradation, its potential to leach to depth in soil and to contaminate groundwater is low. This conclusion is corroborated by field leaching and groundwater monitoring data, both derived from use areas in several states in the U.S. and sites directly near handling/mixing facilities. Even when detected in the groundwater, e.g., five reported detections in two states in the U.S., the highest concentration was well below the estimated HAL of the USEPA. PMID- 12132343 TI - Physical and chemical properties of pyrethroids. AB - The physical and chemical properties of the pyrethroids bifenthrin, cyfluthrin, cypermethrin (also zetacypermethrin), deltamethrin, esfenvalerate (also fenvalerate), fenpropathrin, lambda-cyhalothrin (also cyhalothrin), permethrin, and tralomethrin have been reviewed and summarized in this paper. Physical properties included molecular weight, octanol-water partition coefficient, vapor pressure, water solubility, Henry's law constant, fish biocencentration factor, and soil sorption, desorption, and Freundlich coefficients. Chemical properties included rates of degradation in water as a result of hydrolysis, photodecomposition, aerobic or anaerobic degradation by microorganisms in the absence of light, and also rates of degradation in soil incubated under aerobic or anaerobic conditions. Collectively, the pyrethroids display a highly nonpolar nature of low water solubility, low volatility, high octanol-water partition coefficients, and have high affinity for soil and sediment particulate matter. Pyrethroids have low mobility in soil and are sorbed strongly to the sediments of natural water systems. Although attracted to living organisms because of their nonpolar nature, their capability to bioconcentrate is mitigated by their metabolism and subsequent elimination by the organisms. In fish, bioconcentration factors (BCF) ranged from 360 and 6000. Pyrethroids in water solution tend to be stable at acid and neutral pH but [table: see text] become increasingly susceptible to hydrolysis at pH values beyond neutral. Exceptions at higher pH are bifenthrin (stable), esfenvalerate (stable), and permethrin (half-life, 240 d). Pyrethroids vary in susceptibility to sunlight. Cyfluthrin and tralomethrin in water had half-lives of 0.67 and 2.5 d; lambda-cyhalothrin, esfenvalerate, deltamethrin, permethrin, and cypermethrin were intermediate with a range of 17 110 d; and bifenthrin and fenpropathrin showed the least susceptibility with half lives of 400 and 600 d, respectively. Pyrethroids on soil can also undergo photolysis, often at rates similar to that in water. Half-lives ranged from 5 to 170 d. [table: see text] Pyrethroids are degradable in soils with half-lives ranging from 3 to 96 d aerobically, and 5 to 430 d anaerobically. For those pyrethroids studied in water (cypermethrin, deltamethrin, esfenvalerate, fenpropathrin, and lambda-cyhalothrin), aerobic and anaerobic degradation often continued at rates similar to that displayed in soil. PMID- 12132344 TI - [Interventional cardiology--present possibilities]. PMID- 12132345 TI - [Surgical myocardial revascularization--present status] ]. AB - The number of operations increased and except of classical revascularisation with cardiopulmonary bypass and ischaemic heart arrest several new methods were introduced. Less invasive approach, especially without cardiopulmonary bypass is used more frequently due to technical development in stabilizing systems. The use of arterial grafts became a standard. Due to the improvements in surgical technique and postoperative care the significance of traditional risk factors decreased. The early results improve in spite of older patients with more risk factors are operated on every year. PMID- 12132346 TI - [Contribution of an experimental animal diabetologist to the diagnosis and therapy of human diabetes mellitus]. PMID- 12132347 TI - [Hypoglycemia, an acute complication or a syndrome?] ]. PMID- 12132348 TI - [Prediction of type 1 diabetes mellitus in first degree Czech relatives of diabetic patients]. AB - Diagnosis of autoimmune beta cell destruction by genetic risk analysis, autoantibody evaluation and the test of stimulated insulin secretion performance in first-degree relatives of diabetic patients. 208 Czech children and adults (101 boys and 107 girls, 186 siblings, 22 offspring of diabetic parents, aged 1 22 years, mean age 11.5 +/- 5.4 years) were enrolled in the study. Complete DQB1, DQA1 typing and DRB1*04 subtyping were performed by the PCR in 202 subjects. Sera of all children were investigated for anti-GAD65, anti-IA2 and insulin antibodies using RIA methods. The cut-off normal levels were determined as the 99th percentile of 105 non-diabetic children. IVGTT was performed in children with significant titre of one or more autoantibodies. Total level of stimulated insulin secretion < 48 mU/l was assessed as defect of FPIR. Risk genotype DQA1*05 DQB1*0201/DQA1*03-DQB1*0302 (OR = 100, CI 95% 13-730) was found in 24 of 202 first-degree relatives (12%). 22 children (11%) carried strong protective allele DQB1*0602 (OR = 0.03, CI 95% 0.01-0.12). Autoantibody positivity was recognised in 9 of 208 children (2.9%) and IVGTT was performed. Positivity of anti-GAD65, anti-IA2 or IAA was identified in 5 of 24 children with the highest risk genotype (21%) and in 4 children of 113 with lower risk or neutral genotypes (3.5%). Borderline positivity of one autoantibody was found in 1 boy with the highest risk genotype and in 2 children with lower risk genotypes. Only temporary anti GAD65 positivity was found in girl with protective genotype. Type 1 diabetes mellitus was diagnosed in boy during IVGTT and disease manifested 6 months after IVGTT in girl with defect of FPIR. Standardised methods for prediction of Type 1 diabetes were introduced in first-degree relatives of diabetic patients. These methods are used for Czech registry of diabetic children. PMID- 12132350 TI - [Obesity, type 2 diabetes and their quantitative relation]. AB - Diabetes type 2 and obesity are diseases which have marked mutual relations. The pathogenesis of these relations is in many respects obscure. Also the molecular biological knowledge of the two diseases is inadequate so far. In the submitted brief paper the authors present only a review of some quantitative aspects of the relationship of obesity and diabetes supplemented by some of their own results. The authors summarize the epidemiological relations of diabetes and obesity, possible prediction of the development of diabetes in obese subjects, possible prevention of diabetes in obese subjects and possible treatment of obese diabetics. Slight weight reduction, e.g. by using modern anti-obesity drugs is of fundamental importance for reducing the incidence of diabetes and for its treatment. Based on their own experience the authors present their own results of cluster and factor analysis for evaluating the links between diabetes and obesity with the metabolic syndrome. The authors' work similarly as work quoted in the discussion indicate that diabetes as well as obesity are beyond the so-called nucleus of the metabolic syndrome. Even slight body weight reduction by 5-10%, within the reach of everybody, influences in a fundamental way the incidence and compensation of diabetes as well as other components of the metabolic syndrome. PMID- 12132349 TI - [C-peptide as the decisive factor for classification of type 1 diabetes mellitus and type 2 diabetes mellitus]. AB - OBJECTIVE: The main objective was to seek, based on defined groups of diabetics, C-peptide levels on fasting and after stimulation which would help to differentiate diabetes mellitus type 1 from diabetes mellitus type 2 in patients with manifestation of diabetes in adult age. GROUPS: Group A comprised 65 non obese diabetics type 2 with failure of PAD treatment. Group B included 304 newly manifested diabetics type 1 and 2 aged 31-65 years. Group C was formed by 424 patients with diabetes mellitus type 1 and type 2 with different duration of diabetes. RESULTS: Group A: mean C-peptide levels on fasting 0.32 and after stimulation with a standard breakfast 0.59 pmol/ml suggest absolute insulin deficiency in type 2 diabetics with failure of PAD treatment. Group B: 29.2% diabetics type 1 had already during manifestation of diabetes C-peptide levels on fasting < 0.43 pmol/ml and 47.9% C-peptide of < 0.6 after a meal. There were 1.9 and 4.9% subjects among type 2 diabetics with such low C-peptide levels. After a six-year follow up the mean C-peptide levels on fasting declined in type 1 diabetics from 0.49 to 0.16 pmol/ml and in patients originally with type 2 diabetes reclassified to type 1 the levels dropped from 0.56 to 0.26 pmol/ml. Group C served as the basic group for statistically (linear regression method) detected discrimination values of C-peptide differentiating diabetes mellitus type 1 and diabetes mellitus type 2--the liminal value being 0.59 pmol/ml on fasting and 1.0 pmol/ml after a meal. CONCLUSION: In clinical practice it is not possible to assess reliably slowly manifesting diabetes type 1 (LADA by age, BMI and compensation of diabetes. Positivity of antiGAD antibodies does not rule out diabetes mellitus type 1. In unequivocal cases the decisive factor is therefore the C-peptide level on fasting and after a meal. PMID- 12132351 TI - [Nutritional support of elderly diabetics with acute diseases from the viewpoint of the diabetologist]. AB - Acute diseases in seniors suffering from diabetes significantly influence their nutritional status which is one of the factors participating in the development of acute hyperglycaemic decompensation of diabetes or hypoglycaemia. Inadequately compensated diabetes in seniors is conversely associated with nutritional disorders and the patient is at greater risk of an acute intercurrent disease. Ensuring optimal nutrition in old diabetics is one of the basic provisions in the treatment of intercurrent acute disease. PMID- 12132352 TI - [Changes in the concept of the diabetic diet]. PMID- 12132353 TI - [New oral antidiabetic agents and current strategies of peroral antidiabetic therapy (PAD)]. AB - Optimal compensation in patients with type 2 diabetes is individualization of therapy according to the stage of the disease and ratio of the main pathophysiological disorders. Because of the hitherto inadequate effectiveness of known procedures attention is focused on seeking new more effective procedures either based on the use of known molecules of oral antidiabetics or introduction of new substances. PMID- 12132354 TI - [Current trends in insulin therapy]. AB - More than 80 years have elapsed since the discovery of insulin. The submitted paper analyzes the main contemporary trends of the development of insulin therapy which comprise the development of new insulin analogues and their use in modified insulin regimes, the development of preparations for inhalatory or oral administration, new techniques of administration and new ways of self-monitoring. The author mentions also non-traditional forms of insulin therapy such as transplantation of insulin producing tissue and in the conclusion she discusses trends of insulin administration in subjects without diabetes. PMID- 12132355 TI - [Atherosclerosis, alcohol and diabetes]. AB - Diabetes is a known risk factor of atherosclerosis, moderate consumption of alcohol was reported to reduce the cardiovascular morbidity and mortality. This review presents information on the possible mechanisms of the antiatherogenic effect of alcohol and data of the effect of moderate alcohol drinking on the prevention of cardiovascular diseases. Red wine contains polyphenols with an antioxidant effect, alcohol is metabolized to acetaldehyde which was proved to inhibit the formation of advanced glycation endproducts (AGE) and lipoprotein oxidation. Diabetic patients may benefit from moderate consumption of alcohol beverages only if the risk of hypoglycemia is safely excluded. PMID- 12132356 TI - [Autonomic neuropathy in diabetics, treatment possibilities]. AB - Diabetic neuropathy is a chronic complication of diabetes. It involves non inflammatory damage of the function and structure of peripheral nerves by metabolic vascular pathogenic processes. In case of affection of vegetative nerves (small non-myelinated C fibres) autonomic neuropathy develops. It is a relatively frequent form of neuropathy which remains for a long time without clinical symptoms and therefore is rarely diagnosed and treated. Manifestations of the affection are encountered in all organs which are supplied by vegetative nerves. The presence of this complication of diabetes is signalized by tachycardia at rest, deterioration of gastric evacuation, diabetic diarrhoea or constipation, erectile dysfunction, impaired function of the sweat glans or impaired pupillary reaction. The advanced form involves the danger of latent myocardial ischaemia, serious postural hypotension and sudden death. It increases significantly the mortality of the affected patients. Similarly as the treatment of other complication of diabetes, treatment of autonomic neuropathy is difficult. The objective of the present paper is to review contemporary therapeutic possibilities. An essential prerequisite remain efforts to achieve optimal compensation. The authors draw attention to the effect of alpha-lipoic acid which exerts a positive effect not only on subjective symptoms but also on the objective finding. The other mentioned drugs are used either only experimentally or for purely symptomatic treatment. PMID- 12132357 TI - [Basic questions in therapy of the diabetic foot]. AB - The diabetic foot syndrome including foot tissue impairment distally from the ankle associated with neuropathy and angiopathy in diabetic patients is the main cause of amputations. However, amputations are not inevitable and the provision of rapid and intensive treatment of foot complications in multidisciplinary foot clinics can reduce the number of amputations in diabetic patients. The fundamental principle in the therapy is comprehensive approach; the omitting of any of the principle of the therapy may contribute to its failure. The most frequent case in the praxis is the omitting of the of-loading of the ulcers e.g. by the therapeutic shoes (half-shoes) or special contact cast. The infection control by local debridement and systemic antibiotic therapy is crucial. The higher risk of infection may be related to abnormalities in immunity. The condition for wound healing is good vascular supply by percutaneous transluminal angioplasty and/or by peripheral vascular bypasses most frequent performed on infrapopliteal vessels. Especial attention is necessary to give to therapy of Charcot osteoarthropathy. Good metabolic control in some patients is achievable only by intensive insulin treatment, sometimes by insulin pumps. Early recognition of the at-risk patients and the prompt institution of preventive measures may stop new ulcerations and amputations. PMID- 12132358 TI - [Diagnosis of familial adenomatous polyposis]. AB - Familial adenomatous polyposis (FAP) is a hereditary disease with autosomal dominant transmission, it is a practically 100% precancerosis. For detection of further patients in the family careful examination of the patient and all subjects at risk, i.e. above all grade 1 relatives, is decisive. The presented paper summarizes the author's own experience with the follow up and examination of a group of 96 patients from 42 families. The group of patients has been assembled gradually since 1967. The basis of the examination is preparation of a pedigree, somatic examination, endoscopic examination of the large intestine and the oral portion of the digestive tract. Examination of the ocular fundus is valuable as it evaluates the presence of congenital hypertrophy of the retinal pigmented epithelium (CHRPE). A positive finding in relatives permits conclusions on the presence of FAP. Most evidence is provided by molecular genetic examination at the DNA level which makes presymptomatic diagnosis of FAP possible. PMID- 12132359 TI - [Screening programs for sporadic colorectal carcinoma]. AB - In developed countries colorectal cancer (CRC) is the second most frequent organ malignancy of both genders. According to the world statistics Czech Republic occupies the top position in incidence of this disease. Approximately 75% of all CRC are the sporadic CRC in subjects with negative family or personal history of the disease. The low (average) risk factor in these subjects is age over 50 years, from which the incidence of CRC nearly doubles in each decade. The following options of screening are available for these subjects: 1. faecal occult blood test (FOBT), 2. flexible sigmoidoscopy, 3. combination of both previous procedures, 4. colonoscopy, 5. virtual colonography. FOBT is the mostly used programme in asymptomatic subjects over age 50 at one-year or at least two-years intervals. In FOBT-positive persons colonoscopy is considered the optimal diagnostic and in the case of polyps also therapeutic method. Prospective randomized studies proving a decrease of CRC-mortality in the range of 15-33% are available only for this type of programme. Screening of sporadic CRC was introduced on national basis in Federal Republic Germany in 1977 and in Czech Republic since the second half of the year 2000. With so many people dying of the disease, we cannot afford not to do its screening. PMID- 12132360 TI - [Gastrointestinal surgery and its perspectives]. PMID- 12132361 TI - [Etiopathogenesis of diarrhea and basic principles of diagnosis and therapy]. AB - Acute diarrhoea is worldwide the second most frequent disease after acute inflammations of the airways. Chronic diarrhoea is a less frequent disease, nevertheless the GP, specialist in internal medicine or gastroenterologist encounters it very frequently. For correct understanding of basic diagnostic and therapeutic principles of diarrhoea knowledge of its etiopathogenesis is necessary. In the submitted review the author mentions the functions of the small and large intestine and their part in the development of diarrhoea. He gives also the definition and classification of diarrhoea. The author presents the basic characteristics of osmotic, secretory, inflammatory, motor diarrhoea and diarrhoea associated with increased intestinal filtration. The basic diagnostic and therapeutic principles are in the last part of the review which has an educational character. PMID- 12132362 TI - [Diagnosis of the Zollinger-Ellison syndrome. I: Significance of anamnestic, clinical and laboratory examinations]. AB - The authors describe the diagnostic algorithm of Zollinger-Ellison's syndrome which proved useful in the diagnosis of 73 patients with a confirmed diagnosis. They evaluate the diagnostic validity of anamnestic and clinical data, of different examination methods and compare them with experience assembled abroad. For the diagnosis of sporadic gastrinomas the onset of the disease after the age of 40 years is important, the development of serious peptic complications (haemorrhage F-70%, M-59%, perforation M-54%, F-47%) and the presence of watery diarrhoea (41%). As to laboratory parameters they rely on high BAO values (96% > 15 mmol H+/hour and 100% > 5 mmol H+ after gastric resection. Less important is the examination of basal serum gastrin (almost 30% patients have normal or liminal values of BSG--empirically set at 100-150 pg. ml-1). The authors draw attention to the fact that patients with ZES after gastric resections may have BSG values lower than 100 pg/ml (12%). A positive secretin test has a higher validity (rise of SG by 150-200 pg. ml-1 above basal values) positive in 82.2% patients, liminally positive in 11% and negative in 6.3% patients. An even higher diagnostic value was possessed by a BAO/MAO index higher than 0.6 which was positive in 93.4% patients. At present it is not used as pentagastrin is not available. Every year they diagnose 4-6 new cases of ZES which with regard to the number of inhabitants (5 million) places Slovakia along with Denmark and Sweden among the countries with the highest detection rate (0.8-1.2 ZES cases/1 million per year). PMID- 12132363 TI - [Problems in portal hypertension]. PMID- 12132364 TI - [A new indication for transcutaneous cholangioscopy]. AB - The main indication for transcutaneous cholangioscopy through a channel of percutaneous transhepatic drainage are choledocholithiasis, hepatolithiasis and non-malignant or malignant stenosis of the bile ducts. In our department we were able to treat thus repeatedly also complete severing of the biliary pathways. GROUP: Twelve patients with complete severing of the right hepatic ducts (4x) or common bile duct (8x) after cholecystectomy could not be operated on account of great surgical risk or adhesions after operation. On account of cholangitis they had percutaneous transhepatic drainage. After three weeks transcutaneous cholangioscopy was performed and recanalization by means of a needle-shaped knife. The operation was terminated by introduction of a metal prosthesis Palmaz. RESULTS: The method was successful in all patients, there were no complications associated with the procedure. CONCLUSION: Transcutaneous cholangioscopy and recanalization of complete severing of the biliary pathways is a new safe method. It is indicated in patients with increased surgical risk. For restoration of the continuity of the bile ducts a metal prosthesis Palmaz is inserted. Short-term results of treatment are satisfactors, long-term results call for a longer follow up. PMID- 12132365 TI - [Rifaximin in the treatment of hepatic encephalopathy]. AB - Hepatic encephalopathy is a frequent and serious complication of liver cirrhosis. Usually it is treated by non-absorbable disaccharides or antibiotics and its treatment is often difficult and associated with undesirable effects. The objective of our investigation was to evaluate the safety and effectiveness of a new antibiotic used in this indication--rifaximine. With rifaximine, 400 mg three times per day, a total of 25 patients were treated for a 10-day period. Significant improvement of the manifestations of encephalopathy occurred (evaluated by the grade of encephalopathy, test of combining numerals, the degree of flapping tremor and the arterial ammonia level). None of the patients developed undesirable effects. Rifaximine seems an effective, safe drug for hepatic encephalopathy. PMID- 12132366 TI - [Anti-cytokines in the treatment of idiopathic intestinal inflammations--theory and practice]. AB - Into treatment of idiopathic inflammations of the gut cytokines or their antagonists entered less than 5 years ago and they extended the range of classical medicamentous treatment with aminosalicylates, corticosteroids and immunosuppressives. The theoretical models of their therapeutic application pertained to the blocking of anti-inflammatory cytokines (IL-1, IL-6, IL-8, TNF alpha), the use of immunomodulating cytokines (IL-2, IL-6, IL-8, IL-9) similarly as the therapeutic administration of cytokines with a predominant growth and regulating activity (CSF, TGFalpha, TGFbeta, ODGF, IL-10, IL-11, IL-12). The range is supplemented by ICAM, VCAM antibody oligonucleotides and PAG antagonists. The stage of animal experiments was so far passed only by rhuIL-10, antiIL-2 and PAF antagonists. The only anticytokine which within the record time of 10 years found clinical indication in Crohn's disease, is antiTNF. PMID- 12132367 TI - [Barrett's esophagus]. AB - Barrett's oesophagus is a premalignant metaplastic change of the oesophageal mucosa. Due to its relationship with oesophageal reflux disease and the development of adenoma-carcinoma of the oesophagus the problem arouses increasing interest. In the wide pathogenesis of the disease most probably the composite effect of the refluxed HCl content and duodenal juices play a part. In the diagnosis in addition to fundamental methods--endoscopy and histology- increasingly chromoendoscopy and fluorescent endoscopy are involved. Dispensarization of patients is essential and depends on the degree of pathohistological epithelial changes. Treatment of Barrett's oesophagus can be divided into conservative, where the drug of choice are proton pump inhibitors, and surgical treatment. Promising is endoscopic ablation of the epithelium in combination with subsequent antisecretory therapy. PMID- 12132369 TI - [Methods for diagnosis of thrombophilia caused by disorders in the protein C system (comments on publications by A L Berkovskii et al.)]. PMID- 12132368 TI - [Use of domestic thromboplastins with attested international indices of sensitivity in the treatment of thrombophilia]. AB - Clinical evaluation of thromboplastins manufactured by RENAM (Moscow) with international index of sensitivity (IIS) 1.1-1.5 and comparison of the results with the data obtained in the same patients with foreign thromboplastins (Dade, USA; Behring, Germany; Diamed, Switzerland) and Mediolab thromboplastins (Russia) manufactured from different raw materials (rabbit brain and human placenta) showed that by sensitivity to indirect anticoagulants, Russian thromboplastins made by RENAM and Mediolab firms are not inferior to the best foreign analogs. PMID- 12132370 TI - [Morphofunctional characteristics of erythrocytes in chronic kidney failure and purulent intoxication]. AB - The morphology of blood erythrocytes was examined under a scanning electron microscope and erythrocyte surface charge, lipid peroxidation on erythrocyte membranes, and blood viscosity were studied in 213 urological patients with chronic renal insufficiency (CRI) and purulent intoxication (PI) of different severity. CRI and PI were characterized by morphofunctional changes in erythrocytes, which depended on the type and severity of intoxication. Changes in erythrocyte morphology in CRI and PI were paralleled by a decrease in their negative surface charge and activation of lipid peroxidation. These parameters can serve as diagnostic tests for evaluation of the severity of intoxication. PMID- 12132371 TI - [Lectin-induced aggregation of neutrophil granulocytes in patients with unstable angina]. AB - Lectin-induced aggregation of neutrophilic granulocytes was studied in patients with unstable angina pectoris. PHA-P, WGA, ConA, LCL, PHA-L, PHA-E, PSL, HPL, and LABA were used as aggregation inductors. The results indicate restructuring of carbohydrate determinants of glycoprotein receptors in neutrophilic granulocytes which stimulates their aggregation. PMID- 12132372 TI - [Mixed infections. Basic conceptions, trends in research development, clinical and biological significance]. PMID- 12132373 TI - [Increased serum level of the acute inflammation phase parameter CRP and the high level of low density lipoprotein cholesterol--factors of increased risk of development of atherosclerosis and its complications (a literature review)]. PMID- 12132374 TI - [Test-methods in diagnosis of antibiotic resistance in microorganisms (a literature review)]. PMID- 12132375 TI - [Use of specific antibodies labelled with colloid silver particles for detection of Brucella antigens with dot immune analysis]. AB - Fitness of dot immuno-analysis for detection of Brucella antigens labeled with colloid silver is evaluated. Soluble lipopolysaccharides and protein-saccharide antigen and corpuscular antigens of 22 Brucella strains (7 species) pathogenic for humans and animals in the S and R forms were used. The specificity of the method was tested on 10 heterologous microorganisms whose antigens were closely related. The suggested test system is simple, economic, highly sensitive (from 62 thousands to 8 million CFU/ml) and specific, requires no expensive equipment, and is an alternative to enzyme immuno-assay and dot immuno-analysis with gold immunosole. PMID- 12132376 TI - [Evaluation of the first domestic immunoenzyme recombinant test-system "Khlami IGA-DS-Tr" for detection of class A antibodies to Chlamydia trachomatis]. AB - A test system for detecting species-specific class A antibodies to Chlamydia trachomatis, making use of recombinant chlamydial antigen, is developed in Russia for the first time. Its efficiency was demonstrated in blind clinical trials. The sensitivity, specificity, activity, and use of Chlamy-IgA-DC-Tr test system are not inferior to foreign analogs. PMID- 12132377 TI - [Laboratory diagnosis of systemic chlamydiosis]. AB - Modern methods for diagnosis of chlamydial infection are compared. In order to detect the total systems' manifestations in patients with chlamydial infection, the authors suggest to extend the list of clinical materials to be collected in patients with suspected chlamydial infection and to use direct immunofluorescent test, polymerase chain reaction, and ligase chain reaction in diagnostic studies. PMID- 12132378 TI - [Detection of anti-Borrelia antibodies by immunoblotting in Lyme borreliosis]. AB - IgM and IgG were detected in the sera from 25 Russian patients with Lyme disease by immunoblotting test with Immunetics kit (USA). Early stage of the disease was diagnosed in 12 patients and late stage in 13. Specific protein lines were detected in virtually all patients but their number and combination in the sera were different. Eleven (43%) sera were regarded as positive according to the American criteria of seropositive test, 3 of them by IgM and 10 by IgG. In patients with short disease serum reaction was either negative or confined to 1-2 lines per strip. Patients with late stage showed a more manifest extensive reaction, which was the most pronounced in patients with a long lasting disease and articular involvement; the reaction was similar to that characteristic of chronic Lyme arthritis. Parallel testing of IgG antibodies in 25 sera from American patients with Lyme disease (confirmed by direct isolation of the agent) showed notable similarity of reactions of both American and Russian sera. More essential differences were observed only for low molecular weight proteins (p28, p23, p21). PMID- 12132379 TI - [Detection of nucleic acids in stomach and duodenal contents in patients with gastroduodenal pathology]. AB - The proposed method shows the velocity of gastroduodenal epitheliocyte exchange by evaluating the total content of nucleic acids in gastric and duodenal aspirates. The concentration of nucleic acids in the cavitary secretion phase reflects the state of the gastroduodenal mucosa and physicochemical properties of the mucus. Sixty patients with gastroduodenal diseases (chronic gastritis, chronic duodenitis from surface to atrophic and during exacerbation) were examined. Changes in the distribution of nucleic acids by phases of cavitary secretion were revealed. A decrease in the total content of nucleic acids in cavitary contents was paralleled by decreased activity of chronic gastritis and duodenitis and normalization of colloid and gel-forming properties of the mucus, which manifested by an essential increase of nucleic acids concentrations in the mucus and decrease in the gastric juice. PMID- 12132380 TI - [Bone marrow analysis in a centralized laboratory in Ekaterinburg]. PMID- 12132381 TI - [Analysis of serum and bile lipids in toxic hepatitis by gas chromatography]. PMID- 12132382 TI - [Membrane-stabilizing effect of phospholipid liposomes in toxic hepatitis]. PMID- 12132383 TI - [Significance of detection of hyperlipoproteinemia as a risk factor for development of ischemic heart disease in a polyclinic]. PMID- 12132384 TI - [Oxygen-dependent phagocyte metabolism and blood plasma antioxidant defense in acute coronary syndromes depending on the outcome during hospitalization]. PMID- 12132385 TI - Outbreaks of multi-drug resistant Escherichia coli in long-term care facilities in the Durham, York and Toronto regions of Ontario, 2000-2002. PMID- 12132386 TI - World Health Organization study of venous thrombosis and air travel. PMID- 12132387 TI - Implant surgery using customized surgical templates: the Compu-Guide Surgical Template System. Interview. PMID- 12132388 TI - Naturally occurring cyclic tetrapyrroles. PMID- 12132389 TI - The chemistry of taxol and related taxoids. PMID- 12132390 TI - Development of visceral smooth muscle. AB - The development of the smooth musculature of viscera has attracted the interest of only relatively few investigators, and thus the field appears somewhat underexplored. The major emphasis on histochemical evidence--at the expense of ultrastructural and functional studies--may have limited the progress in this area. Mature tissue is formed through the differentiation of precursors into muscle cells and through the organization of these cells into a complex tissue where distribution and orientation of muscle cells, deployment of abundant extracellular materials and addition of other cellular elements (interstitial cells, fibroblasts, nerves, blood vessels) are characteristic and specific features. The precursor cells are found at sites where a muscle develops, and they derive predominantly from the mesoderm, but also from the neuroectoderm and from the endoderm. The process starts at different times in different organs. The earliest stages of differentiation are characterized by the precursor cells aggregating and becoming elongated; their longitudinal axis lies in a position similar to the one they will have in the mature muscle. Both the cytological and the histochemical differentiation follow distinct patterns in various muscles, with characteristic temporal sequences in the appearance of key features. This process must impart distinct functional properties to a muscle cell at each stage of its development. However, the chronological correspondence between ultrastructural and histochemical development is poorly understood. Histochemical studies have detected gradients of maturation of the muscle cells, for example, across the thickness of the gizzard musculature and along the length of the small intestine; ultrastructural studies have not yet confirmed the existence of these gradients. Muscle growth is accounted for by muscle cell enlargement (without nucleus duplication) and an increase in muscle cell number by mitosis of pre existing differentiated muscle cells. De-differentiation and division of muscle cells, migration of muscle cells and late development of muscle cell precursors have all also been considered as possible mechanisms for muscle growth. Several authors have described the presence of precursor cells within developing smooth muscles, and they have described late differentiation of some muscle cells or waves of differentiation that would give rise to phenotypic heterogeneity of the mature muscle cell population. In contrast, other studies, mainly by electron microscopy, have suggested that, within large visceral muscles, the muscle cells differentiate synchronously. There are interesting data on the influence of adjacent tissues on the development of a smooth muscle, but the interplay of these and other factors has not been fully investigated. Smooth muscles contract from early in their development, hence mechanical factors are likely to influence development: on the one hand, passive stresses imposed on the muscle by other tissues, such as adjacent muscles or the contents of the viscera and, on the other hand, active forces generated by the muscle itself. The very attraction of visceral smooth muscles in the study of cellular morphogenesis--an attraction that has not yet been highlighted or exploited in scientific studies, either descriptively or experimentally--is that, onto a single type of cell, a large range of factors interact, such as the genetic expression, chemical influences (from other muscles, endocrine glands, nerves, other intramuscular cells) and mechanical factors. PMID- 12132391 TI - Transcription factors in skeletal myogenesis of vertebrates. PMID- 12132392 TI - Hypaxial muscle development. PMID- 12132393 TI - Inhibition of skeletal muscle development: less differentiation gives more muscle. AB - The fact that stem cells have to be protected from premature differentiation is true for many organs in the developing embryo and the adult organism. However, there are several arguments that this is particularly important for (skeletal) muscle. There are some evolutionary arguments that muscle is a "default" pathway for mesodermal cells, which has to be actively prevented in order to allow cells to differentiate into other tissues. Myogenic cells originate from very small areas of the embryo where only a minor portion of these cells is supposed to differentiate. Differentiated muscle fibres are unconditionally post-mitotic, leaving undifferentiated stem cells as the only source of regeneration. The mechanical usage of muscle and its superficial location in the vertebrate body makes regeneration a frequently used mechanism. Looking at the different inhibitory mechanisms that have been found within the past 10 or so years, it appears as if evolution has taken this issue very serious. At all possible levels we find regulatory mechanisms that help to fine tune the differentiation of myogenic cells. Secreted molecules specifying different populations of somitic cells, diffusing or membrane-bound signals among fellow myoblasts, modulating molecules within the extracellular matrix and last, but not least, a changing set of activating and repressing cofactors. We have come a long way from the simple model of MyoD just to be turned on at the right time in the right cell. PMID- 12132394 TI - Control of muscle size during embryonic, fetal, and adult life. PMID- 12132395 TI - Cadherins in skeletal muscle development. PMID- 12132396 TI - Slow myosins in muscle development. AB - Myogenesis has been a system central to investigations on mechanisms of diversification within groups of differentiating cells. Diversity among cell types has been well described in striated muscle tissue at the protein and enzymatic-function levels for decades, but it is only in recent years that some understanding of the molecular mechanisms responsible for this diversity has begun to emerge. Study of the expression of the slow isoforms of the myosin heavy chain has contributed to our understanding of how cell diversity arises within skeletal and cardiac muscle. Slow MyHc isoforms are developmentally responsive to a number of cues provided by the nervous systems, the endocrine system and, later in development, to functional demands on these developing tissues. Perhaps most informative have been studies on the mechanism for regulation of slow MyHc expression in mammals and birds where studies on the calcineurin-NF-AT pathways and nuclear hormone action have been shown to control MyHC gene expression in skeletal muscle and in the developing heart. The mechanisms involved in cell diversification in myogenesis are undoubtedly more varied and complex than those currently offered to explain cell diversification, but these recent studies have broadened our understanding of the interplay between the nervous system, the endocrine system and cell lineages in controlling cell diversification. Greater focus on the first fibers and cardiomyocytes to form in the embryo are likely to bring additional insights into the mechanism crucial for establishing the patterns of diversity required for successful formation of embryonic tissues. PMID- 12132397 TI - Molecular characterization of early cardiac development. PMID- 12132398 TI - Mammalian smooth muscle differentiation: origins, markers and transcriptional control. PMID- 12132399 TI - The genetics of murine skeletal muscle biogenesis. PMID- 12132400 TI - Somite patterning: a few more pieces of the puzzle. PMID- 12132401 TI - [Government and training demands]. AB - The availability of adequately trained professionals in sufficient numbers is an essential condition for guaranteeing good quality of health care. The Individual Health Care Professions Act fosters the quality by defining the educational requirements and the area of professional competence. The proposal of the working group 'Competition, Deregulation and Quality of Legislation in the health care' to make the dental hygienist free accessible for the patient is in accordance with the policy of the government to foster the efficiency by task delegation. The replacement of sectorial EU-directives by general directives for the free movement of professionals within the European Economic Area has more drawbacks than advantages. Most EU-memberstates are not a proponent for a vocational year after the basic dental training. Deficiencies in the basic dental training should have been met by lengthening the basic training. PMID- 12132402 TI - [Careless treatment?]. AB - Comments are given on a recent judgement of a medical disciplinary tribunal in The Netherlands. It is stated that the interexaminer variation in the interpretation of periapical radiolucencies is large. Therefore the problem arises if a dentist may be blamed for not detecting a periapical radiolucency on a dental radiograph. A demand for systematic radiographic examination of all teeth with large fillings is also questionable, especially during periodic examinations of the dentition in patients without pain or symptoms of inflammation. PMID- 12132403 TI - Top blood pressure number warrants your attention. PMID- 12132404 TI - My wife and I are both over 70 and suffer from frequent nighttime leg cramps that interfere with our sleep. What can we do to prevent or treat these cramps? PMID- 12132405 TI - I've heard that chronic heartburn can lead to esophageal cancer. How commonly does heartburn progress to cancer, and can it be prevented? PMID- 12132406 TI - Physical fitness prolongs lives. PMID- 12132408 TI - Folic acid lowers colon cancer risk. PMID- 12132407 TI - Can blood tests detect cancer early? PMID- 12132409 TI - Drug coating improves stent performance. PMID- 12132410 TI - Blood pressure drug helps prevent stroke. PMID- 12132411 TI - Are isoflavones safe? PMID- 12132412 TI - Targeted treatment for non-Hodgkin's lymphoma. PMID- 12132413 TI - Lyme disease rash: no usually a bull's-eye. PMID- 12132414 TI - Low cholesterol linked to lower dementia risk. PMID- 12132415 TI - PET scan detects recurrent breast cancer. PMID- 12132416 TI - Daily aspirin may thwart prostate cancer. PMID- 12132417 TI - Antibiotics for heart health? PMID- 12132418 TI - Obesity has become an epidemic among our children...and can school interventions solve the problem? PMID- 12132419 TI - Child safety. A perspective on bioterrorism. PMID- 12132420 TI - Child safety. The importance of the chickenpox vaccine. PMID- 12132421 TI - When parents decide to eliminate milk and eggs in their child's diet, are they right? PMID- 12132422 TI - Parent/child. "Time outs" and how to use them. PMID- 12132423 TI - Product recalls. Recall of Curious George toys. PMID- 12132424 TI - Hepatitis B vaccine encouraged for all newborn infants...and do multiple vaccines overwhelm a child's immune system? PMID- 12132425 TI - Risks for birth defects or low birth weight: air pollution...medications...and caffeine. PMID- 12132427 TI - Parental disapproval of smoking CAN make a difference. PMID- 12132426 TI - Avoiding allergic reactions to foods in restaurants. PMID- 12132429 TI - Product recalls. Kid Cool, Baby Cool girl's infant/toddler garments sold at Sears. PMID- 12132428 TI - Parent/child. More on "time outs". PMID- 12132430 TI - Product recalls. Pop Links toys. PMID- 12132431 TI - Agromedicine program development: a commentary and book review. PMID- 12132432 TI - Alpha 1-acid glycoprotein levels in human immunodeficiency virus-infected subjects on antiretroviral regimens. PMID- 12132434 TI - Staff education at the click of a mouse. PMID- 12132433 TI - Presence of functional peroxisomal targeting signal in viral protein. PMID- 12132436 TI - Anti-HIV agents. Lopinavir--results after one year. PMID- 12132435 TI - Influence of rapid changes in cytosolic pH on oxidative phosphorylation in skeletal muscle: theoretical studies. AB - Cytosolic pH in skeletal muscle may vary significantly because of proton production/consumption by creatine kinase and/or proton production by anaerobic glycolysis. A computer model of oxidative phosphorylation in intact skeletal muscle developed previously was used to study the kinetic effect of these variations on the oxidative phosphorylation system. Two kinds of influence were analysed: (i) via the change in pH across the inner mitochondrial membrane and (ii) via the shift in the equilibrium of the creatine kinase-catalysed reaction. Our simulations suggest that cytosolic pH has essentially no impact on the steady state fluxes and most metabolite concentrations. On the other hand, rapid acidification/alkalization of cytosol causes a transient decrease/increase in the respiration rate. Furthermore, changes in pH seem to affect significantly the kinetic properties of transition between resting state and active state. An increase in pH brought about by proton consumption by creatine kinase at the onset of exercise lengthens the transition time. At intensive exercise levels this pH increase could lead to loss of the stability of the system, if not compensated by glycolytic H+ production. Thus our theoretical results stress the importance of processes/mechanisms that buffer/compensate for changes in cytosolic proton concentration. In particular, we suggest that the second main role of anaerobic glycolysis, apart from additional ATP supply, may be maintaining the stability of the system at intensive exercise. PMID- 12132437 TI - Anti-HIV agents. The benefit of HAART in older people. PMID- 12132438 TI - Infection fighters. Large study finds hepatitis C virus infection linked to reduced benefit from anti-HIV drugs. PMID- 12132439 TI - Side effects. Safety and effectiveness of interferon and ribavirin in HIV+ people co-infected with hepatitis C virus. PMID- 12132440 TI - Nutrition. Vitamin A and abnormal cells on the cervix. PMID- 12132441 TI - Side effects. FDA issues warning about d4T and ddI during pregnancy. PMID- 12132442 TI - Side effects. Severe nevirapine rash found more likely in women than men. PMID- 12132443 TI - Infection fighters. Maintenance therapy for TB works. PMID- 12132444 TI - Infection fighters. Interferon and ribavirin for hepatitis B. PMID- 12132445 TI - Nutrition. How nutrient deficiencies occur. PMID- 12132447 TI - Side effects. Effect of combination therapy on pregnancy--results from Europe. PMID- 12132446 TI - Nutrition. Two German studies find NAC supplements helpful. PMID- 12132448 TI - Side effects. Protease inhibitors and pregnancy--reports from the U.S. PMID- 12132449 TI - Immune boosters. Bone marrow stimulant may boost immunity. PMID- 12132450 TI - Infection fighters. MAC prevention takes a holiday. PMID- 12132451 TI - Side effects. Treating parts of the lipodystrophy syndrome--blood sugar problems. PMID- 12132452 TI - Side effects. Metformin for blood sugar problems. PMID- 12132453 TI - Side effects. Metformin--caution and concerns. PMID- 12132454 TI - Side effects. Calcium supplements help metformin users absorb vitamin B12. PMID- 12132455 TI - Can antioxidants help reduce side effects from ribavirin? PMID- 12132456 TI - High levels of testosterone develop in some women with body shape changes. PMID- 12132457 TI - A combination of testosterone and exercise for men. PMID- 12132458 TI - Anti-HIV agents. Different effects of nukes in men and women. PMID- 12132459 TI - Anti-HIV agents. Long-term use of hydroxyurea. PMID- 12132460 TI - Anti-HIV agents. Indinavir--3 years and still going strong. PMID- 12132461 TI - Anti-cancer agents. Lymphoma in the age of HAART. PMID- 12132462 TI - Anti-viral therapy for lymphoma. PMID- 12132463 TI - Anti-cancer agents. Hydroxyurea as anti-viral therapy for brain lymphoma. PMID- 12132464 TI - Anti-cancer agents. Will cidofovir be useful against cervical lesions? PMID- 12132465 TI - Side effects. Bone problems may occur with hepatitis treatment. PMID- 12132466 TI - Side effects. Indinavir and insulin. PMID- 12132467 TI - Side effects. Metformin for insulin and heart problems? PMID- 12132468 TI - Side effects. Niacin for cholesterol and weight problems? PMID- 12132470 TI - Health tips. Sun-protective clothes. PMID- 12132471 TI - Some antidepressants may reduce heart attack risk. PMID- 12132469 TI - Asthma. Not just for kids. PMID- 12132472 TI - Concerns over kava have the FDA's attention. PMID- 12132473 TI - What are the chances? Putting cancer risk in perspective. PMID- 12132475 TI - Ganglion cyst. A lump of unknown origin. PMID- 12132474 TI - Vulvar cancer. A potentially deadly skin cancer. PMID- 12132476 TI - What is a hammer toe, and what causes it? PMID- 12132477 TI - My friend told me there are health benefits to eating fish at least twice a week. Is that true? PMID- 12132478 TI - Two physicians who helped to establish the Chinese Republic. PMID- 12132479 TI - Prevention of heterotopic bone formation and type-II errors. PMID- 12132480 TI - Prevention of heterotopic bone formation and type-II errors. PMID- 12132481 TI - [(C(wedge)N(wedge)N)Pt(C identical to C)nR] (HC(wedge)N(wedge)N = 6-aryl-2,2' bipyridine, n = 1-4, R = aryl, SiMe3) as a new class of light-emitting materials and their applications in electrophosphorescent devices. AB - Tridentate cyclometalated platinum(II) complexes bearing sigma-alkynyl ligands exhibit tunable photoluminescence and enhanced stability during vacuum deposition; OLEDs based on these materials display orange to red electrophosphorescence with low turn-on voltages (approximately 4 V), maximum luminance approaching 10,000 cd m-2 and efficiency up to 4.2 cd A-1. PMID- 12132482 TI - New microporous coordination polymer affording guest-coordination sites at channel walls. AB - Utilization of a metalloligand, ([Cu(2,4-pydca)2(H2O)].2Et3NH) (1) (2,4-pydca = pyridine-2,4-dicarboxylate), as a building unit provides a novel porous coordination polymer, ([ZnCu(2,4-pydca)2(H2O)3(DMF)].DMF)n (2), in which the Zn(II) ion at the node of the network acts as a linker and the Cu(II) ion in the channel wall is available for guest-coordination. PMID- 12132483 TI - A rod-like polymer containing (Ru(terpy)2) units prepared by electrochemical coupling of pendant thienyl moieties. AB - A new rod-like coordination polymer consisting of (Ru-(terpy)2) motifs bridged by bithiophene units has been prepared by electrochemical polymerisation in acidic organic medium. PMID- 12132484 TI - Synthesis and structural characterisation of stable pyridine- and phosphine functionalised N-heterocyclic carbenes. AB - Stable, uncoordinated (1-[2-(6-trimethylsilyl)pyridyl]-3-[(2,6- diisopropyl)phenyl]imidazol-2-ylidene), I, and (1-[beta-(diphenylphosphino)ethyl] 3-[(2,6-diisopropyl)phenyl]imidazol- 2-ylidene), II, have been synthesised; in the solid state they adopt a conformation with the lone pairs in a mutually anti arrangement. PMID- 12132485 TI - Transient FTIR spectroelectrochemical and stopped-flow detection of a mixed valence (Fe(I)-Fe(II)) bridging carbonyl intermediate with structural elements and spectroscopic characteristics of the di-iron sub-site of all-iron hydrogenase. AB - Iron(I) in biology?: one-electron oxidation of an (Fe(I)-Fe(I)) carbonyl cyanide precursor bearing a proximal thioether group leads to an (Fe(I)-Fe(II)) bridging carbonyl transient with spectral features similar to the di-iron sub-site of the CO inhibited paramagnetic centre of all-iron hydrogenase. PMID- 12132486 TI - 3(5)-tert-butylpyrazole is a ditopic receptor for zinc(II) halides. AB - The complexes [ZnX(HpztBu)3]X (X- = Cl-, Br-, I-) contain a non-coordinated X- anion hydrogen-bonded within a pocket formed by the HpztBu tert-butyl groups. PMID- 12132487 TI - A three-dimensional zeolite-like organic-inorganic hybrid material constructed from (CuMo2O8N)n double helical chains linked via [Cu(4,4'-bpy)]n fragments. AB - The three-dimensional microporous [Cu2Mo2O8(4,4'-bpy)]n.3nH2O 1 contains (CuMo2O8N) double helical chains, which are built up from (CuIIO4N) square pyramids linked by (MoVIO4) tetrahedra and further connected to each other by 4,4'-bpy coordinated (CuIIO5N); the study of the physical properties of 1 demonstrates it is a paramagnetic semiconductor and a zeolite material; the specific channeling cavities in 1 have potential adsorption activities, indicating that 1 might be an attractive functional microporous solid material. PMID- 12132488 TI - Novel solid-state polycondensation I. Oxidative-coupling polymerization of 2,6 dihydroxynaphthalene. AB - Grinding crystals of 2,6-dihydroxynaphthalene-benzylamine complex with FeCl3.6H2O powder in a mortar resulted in the 1,5-oxidative-coupling polymerization of 2,6 dihydroxynaphthalene at room temperature. PMID- 12132489 TI - Serendipitous syntheses and structures of [Cp2Nb(H)((SiMe2)2(mu-NR))]. AB - The compound [Cp2Nb(SiMe2I)2H] reacts with H2NBut and HNEt2 to afford the unexpected imido-bridged 1,3-disilaniobocyclobutanes [Cp2Nb(H)((mu-SiR2)2NR)] (R = But (4), Et (5)) exhibiting surprising structural and spectral properties. PMID- 12132490 TI - Mass casualty management of a large-scale bioterrorist event: an epidemiological approach that shapes triage decisions. AB - The threat of a BT event has catalyzed serious reflection on the troublesome issues that come with event management and triage. Such reflection has had the effect of multiplying the efforts to find solutions to what could become a catastrophic public health disaster. Management options are becoming more robust, as are reliable detection devices and rapid access to stockpiled antibiotics and vaccines. There is much to be done, however, especially in the organizing, warehousing, and granting/exercising authority for resource allocations. The introduction of these new options should encourage one to believe that, in time, evolving standards of care will make it possible to rethink the currently unthinkable consequences. Unfortunately the cost of such preparedness is high and out of reach of most governments. Most of the developing world has neither the will nor the means to plan for BT events and remains overwhelmed with basic public health concerns (i.e., water, food, sanitation, shelter) that must take priority. Therefore, developed countries will be expected to respond using international exogenous resources to mitigate the effects of such a disaster. As a result, the state capacity of the effected government will be severely compromised. If triage and management of casualties is further compromised, terrorists will have met their goals. One could argue that health sciences will continue for decades to play catch up with the advanced technology driving potential bioagent weaponry. If one lesson was learned from the review of the former Soviet Union's biological weapons program, it is that the unthinkable remains an option to terrorists who have comparable expertise. It is crucial to develop realistic strategies for a BT event. Triage planning (the process of establishing criteria for health care prioritization) permits society to see cases in the context of diverse moral perspectives, limited resources, and compelling health care demands. This includes a competent and compassionate management and triage system and an in-depth and accurate health information system that appropriately addresses every level of threat or consequence. In a PICE stage I to III BT event resources will be compromised. Triage and management will be one process requiring multiple levels of cooperation, coordination, and decision-making. An immediate challenge to existing emergency medical services systems (EMSS) is the recognition that locally there will be a shift of emphasis and decision-making from prehospital first responders to community public health authorities. The author suggests that a working relationship, in most areas, between EMSS and the public health system is lacking. As priorities shift in a BT event to hospitals and public health care systems, they need to: 1. Improve their capabilities and capacities in surveillance, discovery, and in the consequences of different triage and management decisions and interventions in a BT environment, starting at the local level. 2. Develop triage and management systems (with clear lines of authority) based on public health and epidemiologic requirements, capability, and capacity (triage teams, categories, tags, rapid response, established operational priorities, resource-driven responsible management process), and link local level surveillance systems with those at the national or regional level. 3. Use a triage and management system that reflects the population (cohort) at risk, such as the epidemiologic based SEIRV triage framework. 4. Develop an organizational capacity that uses lateral decision making skills, pre-hospital outpatient centers for triage-specific treatments, health information systems, and resource-driven hospital level pre-designated protocols appropriate for a surge of unprecedented proportions. Such standards of care, it is recommended, should be set at the local to federal levels and spelled out in existing incident-management system protocols. PMID- 12132491 TI - Synthesis, structure determination and properties of MIL-53as and MIL-53ht: the first CrIII hybrid inorganic-organic microporous solids: CrIII(OH).(O2C-C6H4 CO2).(HO2C-C6H4-CO2H)x. AB - The first three-dimensional chromium(III) dicarboxylate compounds have been isolated and their structures solved from powder X-ray diffraction data; the flexible framework of these materials delimits large pores. PMID- 12132492 TI - [Use of implantable cardioverter defibrillators in the secondary prevention of malignant ventricular arrhythmias: lessons from large studies]. AB - The efficacy of implantable cardioverter defibrillator (ICD) therapy and medical therapy in the treatment of patients with ventricular fibrillation and sustained ventricular tachycardia had been compared in large randomized studies. In this section, we aimed to summarize what we have learned from large studies on the effects of amiodarone and ICD therapy in prevention of cardiac arrest in patients with malignant ventricular arrhythmias. The analyzed studies are; The multicenter unsustained Tachycardia Trial (MUSTT), Antiarrhythmics vs. implantable defibrillator (AVID), Cardiac Arrest Study Hamburg (CASH) and the Canadian Implantable Defibrillator Study (CIDS). PMID- 12132493 TI - [Etiopathogenesis of venous thrombosis and pulmonary thromboembolism: pathophysiologic changes in the cardiovascular system]. AB - It is known that pulmonary embolism is accompanied by quite complex pathophysiological changes in cardiovascular system. From cardiovascular point of view, the diagnosis of pulmonary thromboembolism may be easily based on echocardiographic signs of right ventricular hypokinesia. Physiologic abnormalities caused by venous emboli are related to the cross-sectional area of occluded pulmonary arterial bed. Recent studies has demonstrated, that in patients with massive pulmonary thromboembolism and signs of pulmonary hypertension, increase of right ventricular afterload can lead to both right ventricular failure and reduction of left ventricular preload. Despite development of pulmonary hypertension in acute massive pulmonary thromboembolism, there are no signs of right ventricular hypertrophy. The main ECG changes include right ventricular overload. Previous normal ECG is of special importance. Documentation of serious increase of pulmonary arterial pressure by Doppler echocardiography will assist to link right ventricular pressure overload and dysfunction with embolisms. Transesophageal echocardiography had the similar diagnostic value as transthoracic one, but especially is helpful in bedside diagnosis in patients with signs of shock secondary to pulmonary thromboembolism. PMID- 12132494 TI - [Mitral valve prolapse]. PMID- 12132496 TI - How long before a scheduled surgery do I have to stop eating and drinking? PMID- 12132495 TI - Abdominal aortic aneurysms: when and how to treat. PMID- 12132497 TI - How much daily aspirin should I take to prevent a heart attack or stroke? PMID- 12132498 TI - Does sunscreen block the skin's ability to make vitamin D? If so, how can I get enough of this vitamin without raising my risk of skin cancer? PMID- 12132499 TI - Peanut allergy: a growing problem. PMID- 12132501 TI - Hydration staves off heart disease? PMID- 12132500 TI - Treating enlarged prostates. PMID- 12132503 TI - Routine depression screening recommended. PMID- 12132502 TI - Stick with tried-and-true breast cancer treatment. PMID- 12132504 TI - Low-fat, low-cal diet doesn't mean high cost. PMID- 12132505 TI - New gadgets make carotid stenting safer. PMID- 12132506 TI - Lower doses of HRT protect bones. PMID- 12132507 TI - Foodborne illness on the decline. PMID- 12132508 TI - Hypnotherapy effective for IBS. PMID- 12132509 TI - Device delivers local radiation after breast-lump removal. PMID- 12132510 TI - SAM-e on par with NSAIDs for osteoarthritis? PMID- 12132511 TI - Prostate screening frequency questioned. PMID- 12132512 TI - A new paradigm of chemotherapy for gastric cancer: speeding up, and more clinical trials to catch up. PMID- 12132513 TI - Grading of gastritis: an impossible dream? PMID- 12132514 TI - Different phenotypes in dysferlinopathy. PMID- 12132515 TI - Pure red cell aplasia as an autoimmune-mediated disorder. PMID- 12132516 TI - Preconditioning effect during coronary angioplasty in patients with stable angina pectoris. AB - OBJECTIVE: To determine whether collateral recruitment is involved in the preconditioning effect on the electrocardiogram, chest symptoms, and lactate metabolism during coronary angioplasty in patients with stable angina pectoris. METHODS AND PATIENTS: Sixteen patients with stable angina pectoris underwent three consecutive 2-min balloon inflations 5-min apart. The greatest ST elevation (deltaSTmax), the sum of ST elevations in all leads (sum(ST)), and QT dispersion (QTd) were measured at the end of each balloon inflation. Chest pain score was evaluated on a scale ranging from no pain (0) to the most severe pain (10). Lactate extraction ratio (LER) was determined by simultaneous blood sampling from the aorta and the coronary sinus. Collateral flow index (CFI) was derived from simultaneous measurements of mean aortic pressure and coronary wedge pressure obtained from a pressure guidewire during balloon inflation. RESULTS: Significant decreases were noted in deltaSTmax (3.3+/-2.1 vs. 3.0+/-1.9 vs. 2.6+/-1.8 mm, p<0.01), sum(ST) (9.7+/-7.2 vs. 8.5+/-6.1 vs. 6.9+/-5.3 mm, p<0.01), QTd (55.3+/ 13.8 vs. 46.9+/-9.0 vs. 42.5+/-10.0 ms, p<0.01), and chest pain score (4.3+/-3.1 vs. 2.8+/-2.6 vs. 1.4+/-1.5, p<0.01) during the three sequential balloon inflations. LER significantly increased (-55.5+/-47.8 vs. -36.7+/-34.3 vs. 19.6+/-26.2%, p<0.01), indicating decreased lactate production. No significant difference was observed in CFI (0.16+/-0.10 vs. 0.15+/-0.10 vs. 0.15+/-0.10). CONCLUSION: Repeated balloon inflations during coronary angioplasty elicited a preconditioning effect on ST-segment shift, QT dispersion, chest pain, and lactate production that does not involve collateral recruitment. PMID- 12132517 TI - Hypertriglyceridemia is an independent risk factor for development of impaired fasting glucose and diabetes mellitus: a 9-year longitudinal study in Japanese. AB - OBJECTIVE: Hypertriglyceridemia is often associated with impaired fasting glucose (IFG) and diabetes mellitus. But the contribution of hypertriglyceridemia to the development of IFG and diabetes mellitus remains unclear. We evaluated whether or not hypertriglyceridemia is a risk factor for the development of IFG and diabetes mellitus. METHODS: From 1990 through 1999, 7, 222 Japanese with normoglycemia at baseline were followed. Fasting plasma glucose levels were measured. IFG and diabetes mellitus were defined by ADA criteria. RESULT: The multivariate-adjusted relative risks for the development of IFG were 1.38 for hypertriglyceridemia (p=0.001), 1.30 for obesity (p=0.003), 1.29 for hypertension (p=0.007), 1.26 for family history of diabetes (p=0.027), and 1.02 for age (p=0.035). The multivariate-adjusted relative risks for the development of diabetes mellitus were 1.003 for triglyceride level (p=0.013), 1.30 for level of body mass index (p=0.003), and 2.38 for family history of diabetes (p=0.001). CONCLUSION: Hypertriglyceridemia is an independent risk factor for the development of IFG and diabetes mellitus in Japanese patients. PMID- 12132518 TI - Transient increase in endothelin-1 levels in the pulmonary circulation during exercise while breathing of oxygen in patients with chronic obstructive pulmonary disease. AB - OBJECTIVE: The effects of endothelin-1 (ET-1) on pulmonary vascular tone depend on the complex interplay of ET-1-induced vasoconstriction and vasodilation due to the secondary generation of endothelium-derived vasorelaxants. Therefore, it is likely that the response to ET-1 varies, depending on whether it is applied to the luminal or adventitial side of pulmonary vessels. Therefore, this study was designed to determine the change in luminal ET-1 levels during exercise in patients with chronic obstructive pulmonary disease (COPD). METHODS: All subjects performed a constant-load exercise test for 5 minutes on the ergometer with right heart catheterization while breathing room air or oxygen. ET-1 levels at rest, just after exercise, and 1 hour after exercise were measured in the pulmonary capillary wedge region. PATIENTS: Thirty-six patients with COPD. RESULTS: While breathing room air, ET-1 levels did not significantly differ between at rest, just after exercise and 1 hour after exercise [at rest; 4.15 (0.43) pg/ml, just after exercise; 4.15 (0.44) pg/ml, 1 hour after exercise; 4.13 (0.42) pg/ml]. In contrast, while breathing oxygen, ET-1 levels were significantly higher just after exercise [4.41 (0.43) pg/ml] than at rest [3.90 (0.37) pg/ml, p=0.0116] and 1 hour after exercise [3.93 (0.38) pg/ml, p=0.0246]. The change in ET-1 levels between before and just after exercise (delta ET-1) was negatively correlated with change in mPAP (delta mPAP) (r=-0.638, p=0.0001). However, delta ET-1 was not significantly correlated with any FEV1 (% predicted), DLCO, PaO2, or baseline pulmonary hemodynamics. CONCLUSION: The impairment of ET-1 release into the luminal side was observed in patients with COPD during exercise while breathing room air. However, oxygen supplementation reversed the capacity of ET-1 release, and delta ET-1 with exercise was negatively correlated with delta mPAP. PMID- 12132519 TI - Prognosis of the elderly with asymptomatic atherosclerotic plaques of the carotid arteries. AB - OBJECTIVE: To elucidate the prognosis of the elderly with neurologically asymptomatic atheromatous plaques of the carotid arteries. METHODS: A total of 228 subjects aged 60 years or older, examined by carotid ultrasonography and platelet aggregation test, were studied. They were divided into 3 groups based on plaque morphology: the no lesion group (n=110), the nodular plaque group (n=47), and the mural plaque group (n=71). Platelet aggregability was assessed as suppressed, normal, or accelerated. RESULTS: During the 4.0 years of mean follow up period, 31 subjects died, and 16 of the deaths were due to vascular events such as cerebral infarction or ischemic heart disease. The annual mortality rate due to vascular events was 0.5% in the no lesion group, 1.4% in the nodular plaque group, and 4.1% in the mural plaque group, and Kaplan-Meier survival curve showed poor prognosis of the mural plaque group (logrank statistics: 12.8, Df=2, p=0.0017). According to the Cox proportional hazard model, a high hazard ratio (HR) was seen in the mural plaque group (5.3) and also the accelerated platelet aggregability group (4.0). CONCLUSION: These findings suggested that subjects with mural plaques and accelerated platelet aggregability, even when asymptomatic, have a poor prognosis due to vascular events. Antiplatelet therapy and exercise stress test for detecting coronary artery disease should be considered in these subjects. PMID- 12132520 TI - Clinical heterogeneity in dysferlinopathy. AB - OBJECTIVE: To clarify the clinical heterogeneity and genotype-phenotype correlation in dysferlinopathy. METHODS: We evaluated clinical parameters of 74 dysferlinopathy patients with known dysferlin gene mutations who were previously reported in the literature. RESULTS: The age at onset varied from 12 to 59 years (mean 21.7 years). Based on the initial distribution of muscle involvement, clinical phenotypes were divided into four subtypes: limb-girdle type, Miyoshi's type, distal anterior compartment type, or scapuloperoneal type. These phenotypic differences were prominent at the early stages, but were difficult to recognize later in the progression of the disease. Patients with missense mutations had significantly more severe functional status at examination and higher creatine kinase levels than those with frameshift or nonsense mutations. CONCLUSION: Dysferlinopathy exhibited marked heterogeneity in the age at onset, initial distribution of muscle involvement, and rate of disease progression. As this heterogeneity was observed even within the same family, some additional factors distinct from dysferlin might be involved. PMID- 12132521 TI - Groove pancreatitis: report of a case and review of the clinical and radiologic features of groove pancreatitis reported in Japan. AB - We report a case of groove pancreatitis in which a hypoechoic mass between the duodenum and pancreas head was clearly imaged, and narrowing of the supra ampullary area of the duodenum and bile duct stenosis were also found. The diagnosis was confirmed by surgery. Microscopic examination showed extensive scarring between the duodenum and pancreas head. Protein plugs were found in Santorini's duct. We consider that the disturbance of the pancreatic juice outflow in Santorini's duct is one of the important pathogenic factors in the development of groove pancreatitis. Therefore, we emphasize the finding of Santorini's duct in the differential diagnosis of groove pancreatitis. PMID- 12132522 TI - Inferior mesenteric arteriovenous fistula eight years after sigmoidectomy. AB - We report a 69-year-old woman with liver cirrhosis due to chronic hepatitis C virus (HCV) infection in whom iatrogenic arteriovenous fistula (AVF) developed after sigmoidectomy. A soft mass with bruit led to the diagnosis of inferior mesenteric AVF. Most mesenteric AVF cases have portal hypertension, but this patients showed none of the usual symptoms of portal hypertension; however, she had a splenomegaly that became worse after sigmoidectomy. Clinicians should be aware of the possibility of AVF in patients with a history of abdominal surgery. PMID- 12132523 TI - Cushing's syndrome with a large pituitary adenoma producing both corticotropin releasing hormone (CRH) and adrenocorticotropin (ACTH). AB - A 57-year-old man showed high serum cortisol, plasma adrenocorticotropin (ACTH) and corticotropin-releasing hormone (CRH) levels with a large pituitary tumor and a prostatic cancer. High dose dexamethasone did not suppress cortisol secretion and CRH administration did not stimulate cortisol secretion. After surgical removal of the pituitary tumor, plasma CRH, ACTH and serum cortisol levels were normalized. Histological examinations showed pituitary adenoma and prostatic adenocarcinoma, and pituitary adenoma was stained with both anti-CRH and anti ACTH antibodies, but prostatic cancer was not stained. A CRH-producing pituitary adenoma is a new type of Cushing's syndrome. PMID- 12132524 TI - Recovery from marked altered consciousness in a patient with adult-onset type II citrullinemia diagnosed by DNA analysis and treated with a living related partial liver transplantation. AB - A 21-year-old woman was admitted with altered consciousness and hyperammonemia. She was diagnosed as having adult-onset type II citrullinemia (CTLN2) by DNA analysis. The patient had mutations of the SLC25A13 gene, which were compound heterozygotes of 851 del 4 and IVS11+1G>A. CTLN2 has a poor prognosis, in spite of various intensive medications, and we performed a living related partial liver transplantation (LRLT). Over a 2-year follow-up, the patient has been well. CTLN2 can be diagnosed by the DNA analysis and can be treated by LRLT. PMID- 12132525 TI - Isolated adrenocorticotropin deficiency presenting with impaired renin angiotensin-aldosterone system and suppressed parathyroid hormone-vitamin D axis. AB - We report here a 47-year-old woman with isolated adrenocorticotropin (ACTH) deficiency (IAD). She presented impaired renin-angiotensin-aldosterone (R-A-A) system and suppressed parathyroid hormone (PTH)-vitamin D system. She showed severe hyponatremia due to secondary adrenocortical insufficiency, which was deteriorated by hypoaldosteronism. She also showed hyperphosphatemia and relative hypercalcemia with suppressed PTH-vitamin D axis. Moreover, she showed hypothyroidism, which was thought to be important to maintain normal Ca levels under secondary hypoadrenalism via decrease in bone resorption by T3. Replacement with glucocorticoid completely normalized PTH-vitamin D axis and R-A-A system. Thus, the present case implicates that severe adrenocortical deficiency due to IAD might affect both R-A-A system and PTH-vitamin D axis. These findings suggest that the ACTH-cortisol axis has an important role in mineral metabolism in vivo. PMID- 12132526 TI - Subclinical Cushing's disease accompanied by malignant hypertension and diabetes mellitus. AB - A 53-year-old woman was admitted because of hypertension and diabetes mellitus. Elevated diastolic blood pressure, hypertensive retinopathy and renal dysfunction indicated malignant hypertension. Adrenocorticotropic hormone (ACTH) and cortisol levels were high although there were no Cushingoid features. One mg dexamethasone administration decreased neither ACTH nor cortisol levels. Brain magnetic resonance imaging revealed a left pituitary tumor (7 mm x 6 mm). Upon removal, the tumor showed positive ACTH staining by immnohistochemistry, and was diagnosed as pituitary ACTH-secreting adenoma (Cushing's disease). Her blood pressure, renal function, blood glucose and hormone levels subsequently improved. Malignant hypertension and deteriorated diabetes mellitus may have been due to subclinical Cushing's disease. PMID- 12132527 TI - Hypersensitivity pneumonitis induced by the spore of Pleurotus Eryngii (Eringi). AB - We reported the first case of hypersensitivity pneumonitis (HP) by an edible mushroom, Pleurotus Eryngii (Eringi). A 54-year-old woman had worked in a Bunashimeji mushroom factory for 42 months, and she moved to a new factory producing Eringi. Two months after, she was found to have HP by the spore of Eringi. Although no radiological finding was detected 6 months before the onset of HP, serum surfactant protein D (SP-D) had been elevated. We speculated that type II pneumocyte activation might prepare the ground for HP during the former exposure to Bunashimeji, and serum SP-D levels might reflect their conditions. PMID- 12132528 TI - Visceral larva migrans due to Ascaris suum which presented with eosinophilic pneumonia and multiple intra-hepatic lesions with severe eosinophil infiltration- outbreak in a Japanese area other than Kyushu. AB - A 32-year-old man presented with the chief complaint of severe cough. Examination of peripheral blood showed a marked increase in eosinophils. Chest CT demonstrated multiple ground glass opacities in both lungs. Bronchoalveolar lavage showed abundant eosinophils. Abdominal CT demonstrated multiple low attenuation areas in the liver. Liver biopsy with ultrasonography revealed severe eosinophil infiltrations around the portal veins. Serologically, a multi-dot enzyme linked immunosorbent assay (DOT-ELISA) and ELISA inhibition test using microtiter plates were positive for Ascaris suum. Thus, visceral larva migrans due to Ascaris suum was diagnosed. Outbreaks of this disease in Japan have previously been confined to the Kyushu area. The present case which occurred outside that area, illustrates the importance of constant attention to the epidemiology of this disease. PMID- 12132529 TI - Familial cases of psittacosis: possible person-to-person transmission. AB - A 29-year-old woman with primary myelofibrosis developed severe pneumonia, and 20 days later her 31-year-old sister also contracted pneumonia. The first patient had been in contact with parakeets but the second patient had not been in contact with any birds. Psittacosis was diagnosed in both cases by microplate immunofluorescence antibody technique. Person-to-person transmission between the sisters was suspected to have taken place. PMID- 12132530 TI - Rapidly progressive interstitial lung disease in a dermatomyositis patient with high levels of creatine phosphokinase, severe muscle symptoms and positive anti Jo-1 antibody. AB - It has been reported that there is a subgroup of dermatomyositis (DM) patients with rapidly progressive interstitial lung disease (ILD) who have mild muscle symptoms, slightly increased levels of muscle enzymes, and absence of anti-Jo-1 antibody. A 51-year-old woman with DM was intubated requiring mechanical ventilation because of a rapidly progressing ILD in spite of the absence of the typical poor prognostic factors. A high dose or pulse therapy of corticosteroids was not effective, but additional treatment of cyclosporine gradually improved her respiratory condition. It is not clear why a rapidly progressive ILD occurred in this case lacking poor prognostic factors. However, if corticosteroid treatment is not effective, additional administration of cyclosporine in the early period of rapidly progressive ILD may rescue deteriorating cases. PMID- 12132531 TI - A patient with acquired pure red cell aplasia showing a positive antiglobulin test and the presence of inhibitor against erythroid precursors. AB - A 66-year-old Japanese man developed severe anemia and erythroid hypoplasia in bone marrow without any significant underlying disease. The results of an antiglobulin test were strongly positive, and serum erythropoietin (Epo) was high. The patient was diagnosed as having acquired pure red cell aplasia (PRCA) and was treated with steroids. Anemia was subsided by reticulocyte production in parallel with a decrease in the titer of antiglobulin test and the level of Epo. We studied the immunological mechanism directed against erythroid cells in vitro by using the patient's serum. In vitro analysis indicated the presence of an inhibitor of erythroid precursors at onset, and its disappearance at remission, suggesting the presence of inhibitor against erythroid precursors. PMID- 12132532 TI - Cauda equina syndrome caused by idiopathic sacral epidural lipomatosis. AB - The patient, who was a non-obese woman with no predisposing conditions of lipomatosis, slowly developed cauda equina syndrome. Spinal magnetic resonance imaging (MRI) presented mass lesion of high intensity on T1-weighted image (WI) and an intermediate signal intensity in T2 WI in the epidural space of S1 to coccyges. It has been reported that most idiopathic epidural lipomatosis (IEDL) is observed in obese men, and all cases have involved the thoracic or lumbar region. This is the first report of a patient with cauda equina syndrome caused by idiopathic sacral epidural lipomatosis (EDL). PMID- 12132533 TI - Adenomatous goiter with recurrent thyrotoxicosis. PMID- 12132534 TI - What is the clinical importance of an evanescent rash in a patient with fever of unknown origin? PMID- 12132535 TI - Field dependence of mobilities for gas-phase-protonated monomers and proton-bound dimers of ketones by planar field asymmetric waveform ion mobility spectrometer (PFAIMS). AB - The dependence of the mobilities of gas-phase ions on electric fields from 0 to 90 Td at ambient pressure was determined for protonated monomers [(MH+(H2O)n] and proton bound dimers [M2H+(H2O)n] for a homologous series of normal ketones, from acetone to decanone (M=C3H6O to C10H20O). This dependence was measured as the normalized function of mobility alpha (E/N) using a planar field asymmetric waveform ion mobility spectrometer (PFAIMS) and the ions were mass-identified using a PFAIMS drift tube coupled to a tandem mass spectrometer. Methods are described to obtain alpha (E/N) from the measurements of compensation voltage versus amplitude of an asymmetric waveform of any shape. Slopes of alpha for MH+ versus E/N were monotonic from 0 to 90 Td for acetone, butanone, and pentanone. Plots for ketones from hexanone to octanone exhibited plateaus at high fields. Nonanone and decanone showed plots with an inversion of slope above 70 Td. Proton bound dimers for ketones with carbon numbers greater than five exhibited slopes for alpha versus E/N, which decreased continuously with increasing E/N. These findings are the first alpha values for ions from a homologous series under atmosphere pressure and are preliminary to explanations of alpha (E/N) with ion structure. PMID- 12132536 TI - Mortality among farmers and cotters in the Sundsvall area of Sweden during the nineteenth century. AB - During the latter half of the nineteenth century, the Sundsvall area of Sweden underwent a dramatic economic transformation powered by the exceptional growth of the sawmill industry. Industrialization certainly had a devastating consequences for the health of sawmill employees and their families but also contributed to a gradual reduction in mortality rates among many farmers and cotters. This was particularly true among those who could benefit from the increased market for their produce found in the town and near the sawmills. Much of this study examines the consequences of industrial growth on the population's health. Nonetheless, the marked increase in tuberculosis mortality among women toward the end of the century illuminates the need to consider the social and cultural implications of industrialization rather than simply its economic characteristics when examining mortality patterns. PMID- 12132537 TI - The Black Death: end of a paradigm. PMID- 12132538 TI - Worrying about emotions in history. PMID- 12132539 TI - Evaluating the work of clinical nurse specialists in palliative care. PMID- 12132540 TI - Depression in palliative care: a systematic review. Part 2. Treatment. AB - OBJECTIVE: To summarize available literature containing data on the treatment of depression in palliative care patients. METHODS: A systematic review was conducted using extensive electronic databases and hand searches. All randomized controlled trials (RCTs) of interventions for depression in patients with advanced disease were eligible. RESULTS: Three RCTs assessed pharmacological treatments. Of these, two were placebo controlled and assessed mianserin and thioridazine. The third compared two antidepressants. There were no RCTs that specifically assessed psychotherapy for patients with depression. CONCLUSION: There are too few adequate studies to draw clear conclusions about management of depression in this setting. The treatment of depression in patients with advanced disease must, for now, be informed by the larger body of evidence on effective treatments for depression in patients with either no physical illness or less severe medical conditions. PMID- 12132542 TI - Truth may hurt but deceit hurts more: communication in palliative care. AB - Healthcare professionals often censor their information giving to patients in an attempt to protect them from potentially hurtful, sad or bad news. There is a commonly expressed belief that what people do not know does not harm them. Analysis of doctor and nurse/patient interactions reveals that this well intentioned but misguided assumption about human behaviour is present at all stages of cancer care. Less than honest disclosure is seen from the moment that a patient reports symptoms, to the confirmation of diagnosis, during discussions about the therapeutic benefits of treatment, at relapse and terminal illness. This desire to shield patients from the reality of their situation usually creates even greater difficulties for patients, their relatives and friends and other members of the healthcare team. Although the motivation behind economy with the truth is often well meant, a conspiracy of silence usually results in a heightened state of fear, anxiety and confusion--not one of calm and equanimity. Ambiguous or deliberately misleading information may afford short-term benefits while things continue to go well, but denies individuals and their families opportunities to reorganize and adapt their lives towards the attainment of more achievable goals, realistic hopes and aspirations. In this paper, some examples and consequences of accidental, deliberate, if well-meaning, attempts to disguise the truth from patients, taken verbatim from interviews, are given, together with cases of unintentional deception or misunderstandings created by the use of ambiguous language. We also provide evidence from research studies showing that although truth hurts, deceit may well hurt more. 'I think the best physician is the one who has the providence to tell to the patients according to his knowledge the present situation, what has happened before, and what is going to happen in the future' (Hippocrates). PMID- 12132541 TI - Clinical nurse specialists in palliative care. Part 1. A description of the MacMillan Nurse caseload. AB - Macmillan Nurses play a significant role in specialist palliative care services in the UK, providing direct and indirect services to patients with complex palliative care needs and to their families. Existing literature shows a developing understanding of the role; however, little detailed data exist regarding the clinical work that they undertake. This paper provides evidence from a major evaluation study, commissioned by Macmillan Cancer Relief. It reports the methods of data collection for the study and then goes on to use data from the evaluation to describe the caseload of Macmillan Nurses. Between September 1998 and October 1999, a team of researchers worked alongside 12 Macmillan teams for a period of 8 weeks with each team. Prospective data were gathered on all new referrals to the services within the 8-week period. This included demographic details, timing of referral, the nature and purpose of contacts, and interventions, recorded from case notes and Macmillan Nurse records. Where possible, a date of death was obtained for all patients. A total of 814 new patients were referred during the study period (range 45-114 per site). The most common reasons for referral were emotional care for the patient (57%), pain control (27%), and other physical symptoms (33%). Thirteen per cent of the patients referred to the services died within 1 week of referral while 40% died within 6 weeks; thus, a significant proportion of patient work is focused on care at the end of life. It is also noteworthy that one-third of patients were still alive, indicating that some patients are being cared for earlier in the illness trajectory. On average, each new patient referral received two or more 'face-to-face' visits and two follow-up phone calls within the 8-week period. It would appear that Macmillan Nurse teams have been successful in getting access to relevant patients. As with any service that provides a complex set of interventions, the Macmillan teams have to adapt and develop the services in each setting. Whilst it is clearly important for the development of a Macmillan service to be tailored to the local conditions, the evidence on diversity suggests that in some cases, stronger guidance, in partnership with both Macmillan Cancer Relief and core providers, may be justified. PMID- 12132543 TI - How do patients and spouses deal with the serious facts of malignant glioma? AB - Malignant glioma is a severe disease with little chance for recovery. Due to its effect on cerebral function, it is also a disease with an obvious social impact on family life. Brain tumours, therefore, produce much of the anguish associated with cancer diseases. There is a lack of prospective studies concerning how patients and spouses together deal with the serious facts of cancer. In this study, a series of 25 consecutive patients with malignant gliomas and their spouses was followed during the whole course of the disease process by repeated thematic interviews. The spouses were also subjected to a summarizing interview after the patient's death. The interviews were analyzed qualitatively in order to detect the various ways the patient-spouse couples dealt with this severe situation and how they discussed it with each other. Four different social processes were detected: 1) the patient does not seem to be aware, the spouse is aware but pretends not to be; 2) both are aware, but the patient does not want to share; they drift apart; 3) both are aware, they do/do not talk openly about the gravity of the situation; nevertheless, there is a joint platform; and 4) neither patient nor spouse seems to be aware; they carry on living as before. The patients, compared to the spouses, seemed content with received information. A few of the couples openly discussed death and dying. More common, and apparently sufficient, was a mutual acknowledgement of the serious facts, without using the words 'death' and 'dying'. In a sense, the present findings challenge the awareness categories suggested by Glaser and Strauss. PMID- 12132544 TI - Information needs and awareness of diagnosis in patients with cancer receiving chemotherapy: a report from Greece. AB - This study assessed the information needs of Greek cancer patients and examined whether awareness of diagnosis had any impact on patients. One hundred patients were interviewed about overall and specific information needs, satisfaction, emotional distress, and quality of life. Patients exhibited a great desire for information overall. The need to have more information was high especially regarding the aftermath of chemotherapy, prognosis, how chemotherapy worked, and diagnosis. Patients were more satisfied with care but less satisfied with the information received. Only 37% knew they had cancer, especially the younger, the better educated, and those with breast cancer. Awareness was not related to satisfaction, emotional distress, or quality of life. Our findings suggest that Greek cancer patients need more factual information about their condition and management. Greek oncologists may feel freer to inform their patients about the diagnosis and other issues following their judgement, rather than employing the policy of concealing the truth. PMID- 12132545 TI - Cancer patients' interpretations of verbal expressions when given information about ending cancer treatment. AB - BACKGROUND: Cancer patients' interpretations of the meaning of words used when given bad news are not well studied in medical settings. The aim of this study was to ascertain what significance verbal expressions had for cancer patients when they were given information about ending active tumour treatment, and what message they felt they received. METHODS: Tape-recorded semi-structured interviews were performed and analysed using a qualitative phenomenographical approach. RESULTS: Thirty patients with incurable cancer admitted to hospital based home care unit in Sweden participated. Three main categories about the significance of words emerged: 1) words could indicate indirect warnings as being forewarnings, evasive or ambiguous; 2) words could also be perceived as emotionally trying, as threats or abandoning; 3) other words were fortifying and strengthened the patient in this situation. The overall message given during the information could be interpreted differently: either focused on treatment, on quality of life, or on threat and death. CONCLUSION: The understanding of the significance of words to tailor the information to patients helps the physician to use forewarnings and fortifying words and to identify and avoid the use of threatening words. PMID- 12132546 TI - Psychosocial-spiritual correlates of death distress in patients with life threatening medical conditions. AB - The purpose of this study was to identify demographic, disease, health care, and psychosocial-spiritual factors associated with death distress (death-related depression and anxiety). Cross-sectional baseline data from a randomized controlled trial were used. Outpatients (n=70) were recruited from an urban academic medical centre and proprietary hospital. All patients had life threatening medical conditions, including cancer; pulmonary, cardiac, liver, or kidney disease; HIV/AIDS; or geriatric frailty. Measures of death distress, physical symptom severity, depression and anxiety symptoms, spiritual well-being, social support, patient-perceived physician communication, and patient-perceived quality of health care experiences were administered. In a hierarchical multiple regression model, higher death distress was significantly associated with living alone, greater physical symptom severity, more severe depression symptoms, lower spiritual well-being, and less physician communication as perceived by the patient. Death distress as a unique experiential construct was discriminable among younger patients with specific, diagnosable life-threatening conditions, but less so among geriatric frailty patients. The findings suggest that the experience of death distress among patients with life-threatening medical conditions is associated with the psychosocial-spiritual dimensions of the patient's life. Attention to these dimensions may buffer the negative affects of death distress. PMID- 12132548 TI - Learner-centred education in end-of-life care improved well being in home care staff: a prospective controlled study. AB - The aim of this controlled study was to evaluate a 1-year learner-centred educational project in end-of-life care for home care staff in a rural district of Sweden. Another rural district in the same region served as a control area. A 20-item questionnaire measuring attitudes towards end-of-life care was designed, and the Hospital Anxiety and Depression (HAD) scale was used to measure mental well being. Increased agreement to 18 of 20 attitude statements was seen in the education group, while 2 of 20 items showed a decreased agreement in the control group. Test-retest reliability of the 20-item questionnaire was good (r=0.92). The total HAD score decreased from 8.3 pretest to 5.3 post-test in the education group (95% CI = 2.1 -3.7; P<0.001), and was 6.8 for both years in the control group. Our study shows that a comprehensive educational programme not only improved attitudes towards end-of-life care, but also the mental well being of the home care staff. PMID- 12132547 TI - Terminally ill cancer patients' wish to hasten death. AB - This exploratory study investigated factors associated with the wish to hasten death among a sample of terminally ill cancer patients. Semi-structured interviews conducted on a total of 72 hospice and home palliative care patients were subjected to qualitative analysis using QSR-NUDIST. The main themes to emerge suggested that patients with a high wish to hasten death had greater concerns with physical symptoms and psychological suffering, perceived themselves to be more of a burden to others, and experienced higher levels of demoralization, while also reporting less confidence in symptom control, fewer social supports, less satisfaction with life experiences and fewer religious beliefs when compared with patients who had a moderate or no wish to hasten death. The implications of these findings will be discussed. PMID- 12132549 TI - Gabapentin for coeliac plexus pain. PMID- 12132550 TI - Tumour-induced hypoglycaemia. PMID- 12132551 TI - Statistical aspects of measurement in palliative care. PMID- 12132553 TI - FDA seeks comments on compounding guide. PMID- 12132552 TI - Guidelines promote COX-2 inhibitors for managing chronic pain. PMID- 12132554 TI - NIH panel recommends combination HCV treatment. PMID- 12132555 TI - Alosetron to return to market. PMID- 12132557 TI - Preparing your hospital to respond to a terrorist attack. PMID- 12132558 TI - Training requirements and opportunities in planning responses to bioterrorism. PMID- 12132559 TI - Esomeprazole: a clinical review. AB - The pharmacology, pharmacodynamics, pharmacokinetics, clinical efficacy, and adverse effects of esomeprazole are reviewed. Esomeprazole, a proton-pump inhibitor (PPI), is the S-isomer of omeprazole. Esomeprazole has FDA-approved labeling for use in the treatment of symptomatic gastroesophageal reflux disease (GERD), including healing and maintenance of healing of erosive esophagitis and as part of a triple-drug regimen for Helicobocter pylori infection. Esomeprazole is structurally similar to other PPIs but is the first PPI to include only the active isomer, which may lead to improved pharmacokinetic and pharmacodynamic characteristics. Esomeprazole maintains intragastric pH at a higher level and above 4 for a longer period than other PPIs. Clinical studies have shown that esomeprazole is at least equivalent in safety and efficacy to other drugs in the class. Esomeprazole has demonstrated efficacy in the treatment of erosive esophagitis, the maintenance of healing of erosive esophagitis, and the treatment of signs and symptoms of GERD. Effective dosages are 20 or 40 mg orally every day or as needed. Esomeprazole magnesium 40 mg once daily in combination with amoxicillin and clarithromycin is effective in eradicating H. pylori infection. The potential for interacting with other drugs is limited and is similar to that of omeprazole. The most common adverse effects are headache, respiratory infection, and abdominal symptoms. Esomeprazole has pharmacokinetic properties that may make it more effective than omeprazole in some patients. PMID- 12132560 TI - Using personal digital assistants to access drug information. AB - The use of personal digital assistants (PDAs) to access drug information in a health system is described. Given the widespread use of PDAs at an 872-bed university health system, an opportunity existed to provide current drug information to physicians via these devices. As part of the health system's intranet, extensive online content had been made available through a browser; extension to PDAs was a natural next step. There were two primary requirements: the ability to synchronize information with the database server when a PDA was used and the development of content and applications by using existing staff. Mobile enterprise software was chosen that supports multiple PDA platforms, is easy to use, and does not require programming skills. The software works through customized "channels," or collections of information from a content provider. The customized channel service works over the Internet. Two channels of content were created, an ambulatory care channel and an inpatient care channel. The ambulatory care channel contains a list of preferred ambulatory care agents, poison control information, the locations of outpatient pharmacies, drug information, and safety tips for prescribing. The inpatient channel contains the inpatient formulary, current news and events, information on currrent drug shortages and recalls, pharmacy contact information, and medication safety tips. When a user synchronizes his or her PDA, the software contacts the department's intranet servers and processes the request. The data are compressed and downloaded to the user's PDA. A university health system successfully used PDAs to access drug and other information. PMID- 12132561 TI - Cost-effectiveness of propofol anesthesia using target-controlled infusion compared with a standard regimen using desflurane. AB - The cost-effectiveness of propofol anesthesia using target-controlled infusion (TCI) versus a standard regimen using desflurane for anesthesia maintenance was analyzed. This observational study consisted of 100 inpatients 18 to 75 years old with an American Society of Anesthesiologists physical status of I or II who were scheduled for otological surgery lasting less than four hours. Patients received one of two treatments. The desflurane-maintenance group received propofol 2-4 mg/kg and sufentanil 0.15-0.30 microg (as the citrate)/kg. A constant fresh gas flow of 1 L/min was used during maintenance of anesthesia. The propofol maintenance group received TCI propofol and an additional infusion of sufentanil. Anesthesia was induced with 0.15-0.30 microg/kg. One blinded evaluator assessed the postoperative recovery from anesthesia for all patients. The cost of drugs and medical devices used during the intraoperative and postoperative periods was calculated. Effectiveness was defined as the absence of postoperative nausea and vomiting (PONV), while the cost-effectiveness of each procedure was the cost per PONV-free episode. The efficiency of each procedure represented the production of effectiveness per dollar invested. Chi-square and t tests, sensitivity analysis, and logistic regression were also performed. The only intergroup difference detected was the frequency of PONV occurring in the early recovery phase (11 in the desflurane group versus 2 in the propofol group). Of those patients requiring antiemetic rescue, 9 were in the desflurane group and only 2 were in the propofol group (p < 0.05). The TCI propofol regimen was more expensive than the desflurane regimen ($45 versus $28 per patient, respectively) (p < 0.001). The differential cost-effectiveness ratio was $94.7 per PONV-free episode. PONV 24 hours after surgery and patient satisfaction were similar between groups. A standard regimen of desflurane was more cost-effective than TCI propofol for anesthesia maintenance in achieving PONV-free episodes. PMID- 12132562 TI - Stability of dacarbazine in amber glass vials and polyvinyl chloride bags. AB - The stability of dacarbazine in commercial glass vials and polyvinyl chloride (PVC) bags in various storage conditions and the emergence of 2-azahypoxanthine, a major degradation product possibly linked with some adverse effects, were studied. Triplicate samples of reconstituted (11 mg/mL) and diluted (1.40 mg/mL) dacarbazine admixtures were prepared and stored at 4 degrees C or at 25 degrees C in daylight, fluorescent light, or the dark. The effect of several light protective measures (amber glass vials, aluminum foil wrapping, and opaque tubing) on dacarbazine stability in a simulated i.v. infusion system was also evaluated. Dacarbazine quantification and main degradation product determination were performed by high-performance liquid chromatography. Stability was defined as conservation of 90-105% of initial dacarbazine concentration without major variations in clarity, color, or pH and without precipitate formation. Reconstituted dacarbazine solutions were stable for 24 hours at room temperature and during light exposure and stable for at least 96 hours at 2-6 degrees C when stored in the dark. After dilution in PVC bags, stability time increased from 2 hours in daylight to 24 hours in fluorescent light and to 72 hours when covered with aluminum foil. After two hours of simulated infusion, dacarbazine remained stable. Diluted dacarbazine solutions stored at 2-6 degrees C were stable for at least 168 hours. The only degradation product found was 2-azahypoxanthine, which was detected in every sample. The storage and handling of dacarbazine should take into account both the loss of the drug and the production of its potentially toxic degradation product. Dacarbazine must be carefully protected from light, administered using opaque infusion tubing, and, if necessary, refrigerated before administration to reduce 2-azahypoxanthine formation. PMID- 12132563 TI - Cost-effectiveness of gemifloxacin: results from the GLOBE study. AB - The cost-effectiveness of treatment with oral gemifloxacin versus oral clarithromycin for acute exacerbations of chronic bronchitis (AECB) was evaluated. Economic outcomes were assessed for the Gemifloxacin Long-term Outcomes in Bronchitis Exacerbations study. This prospective double-blind, controlled, health outcomes study compared health, economic, and clinical outcomes after randomized treatment with either oral gemifloxacin or oral clarithromycin for AECB. Base case analysis was performed from the third-party payer's perspective and considered the costs of respiratory tract infection related medical care. Analysis from the societal perspective also included costs of lost productivity. Treatment effectiveness was measured as the proportion of patients without recurrence requiring antimicrobial treatment following resolution of the initial AECB. Data sources included the outcomes study itself and standard U.S. cost sources. Compared with clarithromycin, gemifloxacin treatment resulted in significantly more patients without AECB recurrence requiring antimicrobial treatment after 26 weeks (73.8% versus 63.8%, p = 0.024). Fewer patients receiving gemifloxacin were hospitalized (5 of 214 patients versus 14 of 224 patients, p = 0.059), and they had less time off from usual activities (8.3 days versus 10.1 days). The mean direct cost per patient receiving gemifloxacin was $127 less than with clarithromycin ($247 versus $374, respectively); mean total costs (direct plus indirect) per patient were $329 less for patients receiving gemifloxacin ($1413 versus $1742). Gemifloxacin dominated clarithromycin in cost-effectiveness analysis. Bootstrap analysis indicated that the probability of gemifloxacin being both cost saving and more effective than clarithromycin is 88% from a payer's perspective and 84% from the societal perspective. Gemifloxacin was more cost-effective, improving AECB outcomes and producing substantial cost offsets compared with clarithromycin. PMID- 12132565 TI - Delivering medications via a pneumatic tube system. PMID- 12132564 TI - Economic evaluations of olanzapine and risperidone. PMID- 12132566 TI - Simple extraction: antiquated term or needed paradigm shift? PMID- 12132567 TI - Dentin bonding. Interview by Edward J. Swift Jr. PMID- 12132568 TI - Dynesthetic and dentogenic concept revisited. AB - The dynesthetic and dentogenic concept, when applied, provides a more natural, harmonious prosthesis, which not only is desired by patients, but also is a quality of care they deserve. Outstanding esthetics can be achieved by simple guidelines, using tooth molds specifically sculpted for males and females, arranging prosthetic teeth to correspond with personality and age and sculpting the matrix (visible denture base) with more natural contours. There is no reason for edentulous individuals to be provided with care of any less quality than that available with other procedures, such as crowns, bridges, veneers, or implant restorations. Providing this upscale product can be rewarding and satisfying to patient and operator alike. This concept produces superior results no matter what posterior occlusal scheme is employed but, in the opinion of the author, works best when used in conjunction with a noninterceptive linear occlusion approach (not to be confused with lingualized occlusion), which precludes anterior contact. CLINICAL SIGNIFICANCE: Dentogenics provides an approach to esthetics in prosthodontics that enables the dentist to create a restoration in harmony with the patient's objective personality. This concept considers gender, age, and personality to restore the patient's dignity and unique individuality that has been missing in far too many prostheses. PMID- 12132569 TI - A novel filling technique for packable composite resin in Class II restorations. AB - The use of composite resin restorations in posterior teeth has increased considerably in the past few years. Specific composite resins for posterior teeth, as well as new operative techniques, have been developed to overcome some clinical difficulties. The present article describes a new technique to reconstitute the interproximal contour and contact of Class II restorations using a packable composite resin. Two lower first molars from the same patient are reported in detail, illustrating the technique step by step. The technique indicates the use of a metallic matrix band and wood wedges to provide an interproximal contour and contact with the adjacent tooth as well as to provide an adequate cervical adaptation. The first increment of the packable composite resin is applied on the gingival wall of the proximal box, packed cervically near the axial wall and, automatically, the resin climbs up in contact with the inner surface of the matrix band. This increment is sculpted and light-cured and the metallic matrix band is removed. Thus, the Class II cavities are transformed into Class I, with free access for light-curing. Small incremental layers of composite fill the remaining cavities. This technique is faster than conventional techniques and permits appropriate embrasure, better contour, and contact points. CLINICAL SIGNIFICANCE: Packable composite resins present primarily new handling characteristics that permit the development of novel filling techniques. The technique proposed in this article allows easier Class II buildup, with proper proximal contact and proximal smooth surfaces, once there are no irregularities among the composite increments. Another advantage is that there is adequate light exposure for polymerization, because the metal matrix can be removed during the restorative procedure. PMID- 12132570 TI - Curing-dependent changes in color and translucency parameter of composite bleach shades. AB - PURPOSE: The goal of this study was to evaluate curing-dependent changes in color and translucency parameter (TP) values of composite bleach shades. MATERIALS AND METHODS: Thirty bleach shades of microhybrid and microfill composites were analyzed. Specimens (n = 5) were made as disks, 10 mm in diameter and 2 mm thick, using cylindrical molds. Specimens were polymerized for 60 seconds using a light curing unit. Data were collected before and after composite curing using a spectrophotometer and analyzed using the appropriate color-difference metric equations. RESULTS: L*a*b* ranges (maximum minus minimum values) for microhybrids were 17.7, 2.91, and 7.97, respectively. Corresponding ranges for microfills were 14.4, 1.26, and 4.27, respectively. Curing-dependent color differences varied from 3.7 to 12.0 deltaE* units, whereas TP values of cured resin composites varied from 2.0 to 7.1. Light-curing caused increase of microhybrid TP values (+0.7) and decrease of microfill TP values (-0.7). Color differences were found to be acceptable for five of six composite pairs of the same shade designation (each of them made by the same manufacturer) in post-curing measurements against white background. CLINICAL SIGNIFICANCE: Curing-dependent color and TP changes indicate that dentists should use cured composite for matching of shade and translucency. Tested materials became less saturated, with microhybrids becoming darker and microfills becoming lighter after polymerization. Light-curing caused an increase in translucency of microhybrids and a reduced translucency in microfills. PMID- 12132571 TI - Air-abrasion enamel microsurgery to treat enamel white spot lesions of traumatic origin. PMID- 12132572 TI - Sealants. PMID- 12132573 TI - Immortalization and transformation of human cells. AB - The disruption of homeostatic mechanisms that regulate normal cell growth and proliferation is a hallmark of cancer. Experimentally, many of the same genetic changes that lead to abnormal cell proliferation conspire to confer replicative immortality upon cells in culture. Correspondingly, several lines of evidence implicate cellular immortalization as a prerequisite for cell transformation. Recently much progress has been made in elucidating the cellular machinery that regulates cell lifespan. This review summarizes these recent advances in our understanding of these molecular mechanisms that contribute to human cell immortalization and transformation. PMID- 12132574 TI - Diversity and varietal classification of Hibiscus syriacus L. with the heterogeneity within retrotransposon-like elements. AB - Retrotransposons are present in multi-copy numbers that are integrated into plant genomes with considerable heterogeneous sequences within a single plant and between plant species, which allows the use of retrotransposons as additional sources of DNA polymorphism. A primer design for the sequence-tagged specific site and cleaved amplified polymorphic sequences (STS-CAPs) that are derived from retrotransposon-like sequences was developed for the molecular marker analysis in Hibiscus syriacus. This method was applied for the detection of sequence variations of intact retrotransposons that exist in plant genomes, which resulted in higher polymorphisms than in the amplified fragment length polymorphism (AFLP). Through STS-CAPs, specific fingerprinting data among H. syriacus varieties can be easily distinguished and generated with reproducible results. It could also be adapted to any species that possess multi-copy retrotransposons for varietal identification as well as the assessment of genetic relationships. PMID- 12132575 TI - Characterization of the conserved region of the mxaF gene that encodes the large subunit of methanol dehydrogenase from a marine methylotrophic bacterium. AB - The highly conserved region of the mxaF gene that encodes the large subunit of methanol dehydrogenase (MDH) was cloned and sequenced from Methylophaga sp. strain MP cells. The calculated G + C content of the conserved region was found to be 44.9%. The nucleotide sequence homology of the region to those from methylotrophs was approximately 43.5%, while the identity of the deduced amino acid sequence to other MxaF peptides was approximately 76.8%. Analysis of the codon usage revealed that UUC and CGU codons seem to be used only for phenylalanine and arginine, respectively. The aligned amino acid sequences show that several key amino acids that are required for the MDH enzyme activity are located in the deduced MxaF peptide, together with tryptophan-docking motifs, called W4 and W5. PMID- 12132576 TI - Partial purification and properties of a phosphatidylinositol 4,5-bisphosphate hydrolyzing phospholipase C from the soluble fraction of soybean sprouts. AB - Three soluble enzyme fractions (F-I, F-II, and F-III) that hydrolyze phophoinositides were separated from soybean sprouts by using Matrex green gel column chromatography. Among the three phosphatidylinositol (PI)-specific phopholipsase C (PLC) enzymes, only the third fraction (F-III) was able to hydrolyze phosphatidylinositol 4,5-bisphosphate (PIP2) as well as phosphatidylinositol (PI) and phosphatidylinositol phosphate (PIP) as substrates. The F-I and F-II fractions only showed enzymatic activities for PI and PIP. The PIP2-hydrolyzing PLC protein, F-III, was partially purified using the chromatographic steps of the Matrex green gel, phenyl Toyopearl, Matrex orange gel, Mono S cation exchange, and superose 6 gel filtration columns. The molecular weight of the F-III protein was estimated to be about 64 kDa on SDS-PAGE. The protein showed immunocross-reactivity with a polyclonal antibody that was prepared against the X and Y motifs of animal PLC enzymes, the conserved catalytic domains. Ca2+ ion critically affected the PIP2-hydrolyzing PLC activity of the F-III protein, representing maximal activity at 10 microM Ca2+ concentration. The PIP2-hydrolyzing PLC activity of the protein was also significantly increased by sodium deoxycholate (SDC) from 0.05 to 0.08%. However, the activity was greatly reduced above the concentration, and no activity was detected at 0.3% SDC. In addition, the protein exhibited maximal PIP2-hydrolyzing PLC activity at pH, in the range of 6.5-7.5. PMID- 12132577 TI - Molecular diagnosis of Duchenne/Becker muscular dystrophy by polymerase chain reaction and microsatellite analysis. AB - Duchenne muscular dystrophy (DMD) and Becker muscular dystrophy (BMD) are X linked recessive genetic disorders resulting from mutations in the dystrophin gene. About two-thirds of the affected patients have large deletions or duplications, which occur in the 5' and central region of the gene. The remaining DMD/BMD cases show no deletions, so they cannot be easily identified by current strategies. In these DMD/BMD families, a linkage analysis that involves DNA markers of the flanking and intragenic dystrophin gene are necessary for carrier and prenatal diagnosis. We analyzed eighteen deletion-prone exons of the gene by a polymerase chain reaction (PCR) in order to characterize the molecular defects of the dystrophin gene in Korean DMD/BMD families. We also performed a linkage analysis to assess the usefulness and application of six short tandem repeat markers for molecular diagnosis in the families. We observed a deletion that eliminated the exon 50. Also, a linkage analysis in the families with six short tandem repeat (STR) markers showed heterozygosity at most of the STR markers. The haplotype analysis was useful for detecting the carrier status. This study will be helpful for a molecular diagnosis of DMD/BMD families in the Korean population. PMID- 12132578 TI - Cloning and characterization of the kinesin-related protein, Krp1p, in Schizosaccharomyces pombe. AB - Kinesin have been cloned in many organisms. They played important roles in the transport of cell organelles, polarized growth, and secretion. We report here the identification of a kinesin-related protein in Schizosaccharomyces pombe, which was named kinesin-related protein (Krplp). The primer sequences were driven from the highly conserved area of the kinesin genes in other organisms. We cloned kinesin genes from S. pombe using the PCR technique. Sequence analysis revealed that krp1+ has a 1,665 bp open-reading frame (ORF) that encoded a protein that consisted of 554 amino acids with a molecular weight of 61,900. It is homologous to the proteins that belong to the kinesin heavy chain (KHC) superfamily [GenBank accession No. AF156966 (genomic DNA) and AF247188 (mRNA)]. To characterize Krplp, the gene was disrupted and overexpressed in S. pombe. Cells that contained a krp1+ null allele were viable. Overexpression of Krp1p resulted in the inhibition of mitotic growth; cells became elongated, branched, and formed aberrant septa. To identify proteins that interact with Krplp, the yeast two-hybrid system was used. As a result, the novel protein, designated kinesin associated protein (Kap1p), was identified and showed structural homology to the proteins of the myosin family (GenBank accession No. AF351206). The data from the overexpression and two-hybrid study of Krplp may provide information that Krplp can have roles in cytokinesis with myosin. PMID- 12132579 TI - Age-related decline of inducible nitric oxide synthase gene expression in primary cultured rat hepatocytes. AB - Hepatocytes exhibit a non-specific immune response by expressing the enzyme inducible nitric oxide (NO) synthase (iNOS, NOS2) through the stimulation of a mixture of cytokines, or a single cytokine such as interleukin-1beta. We examined the age-dependent inducibility of the iNOS gene expression and the capacity of NO production in response to lipopolysaccharide (LPS) or interleukin-1beta (IL 1beta) in primary cultured rat hepatocytes that were isolated from the livers of rats, 3 (young) and 24 (aging) months of age. NO production (NO2-), indicating iNOS activity, was much higher in the young rat hepatocytes following stimulation with LPS or IL-1beta. Likewise, in the young hepatocytes, Western blot analyses showed much higher protein levels in the iNOS expression; it was also a little higher in mRNA levels that were analyzed by RT-PCR. Furthermore, after stimulation with IL-1beta, the levels of transactivation of nuclear factor-KB (NF kappaB) that were involved in the induction of the iNOS gene were reduced without a significant difference in the aged cells. Therefore, the decrease of NO formation in the aged hepatocytes was due to the belated and incomplete inducibility of the iNOS protein expression, together with a minor contribution of the reduced-transactivation of NF-kappaB. These results suggest that the age related decline of the iNOS gene expression in primary rat hepatocytes may be associated with the increased incidence of many infective diseases with aging. PMID- 12132580 TI - Carnitine uptake by AGP2 in yeast Saccharomyces cerevisiae is dependent on Hog1 MAP kinase pathway. AB - The AGP2 gene encodes a plasma membrane carnitine transporter in S. cerevisiae. Here, we report the identification of AGP2 as an osmotic stress response gene. AGP2 was isolated from mTn3 tagged mutants that contained in-frame fusions with lacZ. The expression of AGP2 was down-regulated by osmotic stresses, including NaCl, sorbitol, and KCI. We also found that carnitine uptake was inhibited by NaCl. In the ssk1delta stelldelta double-mutant strain, the expression of AGP2 and the uptake of carnitine were greatly reduced compared to the wild-type strain. Furthermore, carnitine uptake was inhibited by the constitutive expression of PBS2, which encodes a MAPKK that activates Hog1. We concluded, therefore, that the HOG pathway plays an important role in the regulation of carnitine uptake in S. cerevisiae. PMID- 12132581 TI - Karyotype analysis of a Korean cucumber cultivar (Cucumis sativus L. cv. Winter Long) using C-banding and bicolor fluorescence in situ hybridization. AB - An intensive karyotype analysis of a Korean cucumber cultivar (Cucumis sativus L. cv. Winter Long) was carried out with three different methods. These included Feulgen staining, Giemsa C-banding, and fluorescence in situ hybridization (FISH). The mitotic chromosomes of the cucumber (2n = 2x = 14) were characterized, based on the length and arm ratio values. A C-banding analysis showed dark stains on the centromeric, telomeric, and intercalary regions of the chromosomes, except that chromosome 2 had a heavy staining in the long arm. Bicolor FISH, using 45S and 5S rDNA probes, provided additional information to identify cucumber chromosomes. The signals for 45S rDNA were detected on the pericentromeric regions of chromosomes 1, 2, and 4. The signals for 5S rDNA were on the short arm of chromosome 5. Similar band patterns (as the C-banding) were observed when the chromosomes were counter-stained with 4',6-diamidino-2 phenyoindole (DAPI). The data implied that the karyotype of the Korean cucumber cultivar is peculiar and different from previous reports. PMID- 12132582 TI - Differential susceptibility of photosynthesis to light-chilling stress in rice (Oryza sativa l.) depends on the capacity for photochemical dissipation of light. AB - The primary target for light-chilling stress in chilling-sensitive cucumber leaves is the chloroplast Cu,Zn-Superoxide dismutase, followed by subsequent inactivation of the photosystem (PS) I by reactive oxygen species (ROS). To test this hypothesis, two rice cultivars that were different in their ecological origins (a chilling-resistant Stejaree 45 and a chilling-sensitive Milyang 23) were evaluated with respect to photosynthetic properties, the ROS scavenging system, and expression of genes that are involved in sucrose synthesis and allocation upon the light-chilling stress. As expected, when the leaves were exposed to various low temperatures with illumination (150 micromol m(-2)s(-1)) for 6 h, the leaf photosynthesis of Milyang 23 decreased faster than that of Stejaree 45. The light-chilling induced differential photoinhibition of photosynthesis between the two cultivars was caused by the photoin-activation of PSII but not of PSI, since the potential quantum yield of PSII followed a similar trend to the changes in photosynthetic rates. The activities of the two chloroplastic antioxidant enzymes (superoxide dismutase and ascorbate peroxidase) that are known to be sensitive to oxidative stress were barely affected by the light-chilling treatments. Among various genes in sucrose metabolism (such as cytosolic FBPase, SPS, SUT, SuSy, and AGPase), the transcript levels of SuSy in Milyang 23 were significantly decreased by light-chilling stress compared to that of Stejaree 45. Based on these results, we propose that PSII, not PSI, is the sensitive site for light-chilling stress in chilling-sensitive rice. The extent of PSII photoinhibition depends on its capacity for the photochemical utilization of light. PMID- 12132583 TI - Osteopontin in kainic acid-induced microglial reactions in the rat brain. AB - The present study was performed to investigate the spatial and temporal expression of osteopontin (OPN) mRNA in the rat brain after kainic acid-induced seizures, and to determine whether this phenomenon is associated spatiotemporally with the microglial reaction. The expression of OPN mRNA was detected using an in situ hybridization technique and Northern blot analysis. Following intraperitoneal injection of kainic acid (10 mg/kg), OPN mRNA was expressed in selective vulnerable areas, including the hippocampus, thalamus, hypothalamus, amygdala, and entorhinal cortex. Comparison of the morphology and localization with the established microglial marker OX-42 in the adjacent sections positively identified the OPN-expressed cells as microglia. Furthermore, double labeling experiments revealed that OPN mRNA expression was confined to ameboid-like cells among microglia stained with GSI-B4, an another microglial marker. These findings from a rat model of seizure support the notion that OPN can be synthesized in a subpopulation of reactive microglial cells. It can therefore be assumed that in the response of the brain to excitotoxic injury, synthesis of OPN occurs generally in a subset of activated microglia. PMID- 12132584 TI - Cloning and expression of squalene synthase cDNA from hot pepper (Capsicum annuum L.). AB - We isolated and artificially expressed a cDNA clone of the Capsicum annuum squalene synthase (CASS) gene to elucidate the pattern of alternatively regulated two-branch point enzymes. The 1,674-bp CASS cDNA contained an open reading frame of 411 amino acids, yielding a predicted molecular mass of about 45 kDa. A deduced amino acid sequence comparison to other squalene syntheses showed identities with Nicotiana tabacum (91%), Nicotiana benthamiana (90%), Arabidopsis thaliana (79%), and rats (40%). The artificially expressed soluble form of the CASS enzyme was identified by the enzyme activity that converted FPP to squalene and by SDS-PAGE. A Southern blot analysis indicated that at least two copies of the squalene synthase gene exist in the hot pepper genome. In hot pepper, the regulation of the branch point enzymes, squalene synthase and sesquiterpene cyclase was investigated in the UV-challenged leaves of Capsicum annuum. The transcript level and enzyme activity of the CASS were slightly reduced by UV. However, those of the CASC were rapidly induced within 24 h and slowly decreased thereafter. PMID- 12132585 TI - Determination of S-genotypes of pear (Pyrus pyrifolia) cultivars by S-RNase sequencing and PCR-RFLP analyses. AB - The pear (Pyrus pyrifolia) has gametophytic self-incompatibility (GSI). To elucidate the S-genotypes of Korean-bred pear cultivars, whose parents are heterozygotes, the PCR amplification using S-RNase primers that are specific for each S-genotype was carried out in 15 Korean-bred pear cultivars and 5 Japanese bred pear cultivars. The difference of the fragment length was shown in the following order: S6 (355 bp) < S7 (360 bp) < S1 (375 bp) < S4 (376 bp) < S3 and S5 (384 bp) < S8 (442 bp) < S9 (1,323 bp) < S2 (1,355 bp). We analyzed the sequence of the S-RNase gene, which had introns of various sizes in the hypervariable (HV) region between the adjacent exons with a fairly high homology. The sizes of the introns were as follows: S1 = 167 bp, S2 = 1,153 bp, S3 = 179 bp, S4 = 168 bp, S5 = 179 bp, S6 = 147 bp, S7 = 152 bp, S8 = 234 bp, S9 = 1,115 bp. There were five conservative and five hypervariable regions in the introns of S1, S3, S4, S5, S6 and S-RNases. A pairwise comparison of these introns of S RNases revealed homologies as follows: 93.7% between S1- and S4-RNases, 93.3% between S3- and S5-RNases and 78.9% between S6- and S7-RNases. PCR-RFLP and S RNases sequencing determined the S-genotypes of the pear cultivars. The S genotypes were S4S9 for Shinkou, S3S9 for Niitaka, S3S5 for Housui, S1S5 for Kimizukawase, S1S8 for Ichiharawase, S3S5 for Mansoo, S3S4 for Shinil, S3S4 for Whangkeumbae, S3S5 for Sunhwang, S3S5 for Whasan, S3S5 for Mihwang, S5S? for Chengsilri, S3S5 for Gamro, S3S4 for Yeongsanbae, S3S4 for Wonhwang, S3S5 for Gamcheonbae, S3S5 for Danbae, S3S4 for Manpoong, S3S4 for Soowhangbae and S4S6 for Chuwhangbae. The information on the S-genotypes of pear cultivars will be used for the pollinizer selection and breeding program. PMID- 12132587 TI - Enzymatic characterization of glycosidase antibodies raised against a chair transition state analog and the retained catalytic activity from the expressed single chain antibody fragments. AB - Catalytic antibodies with a glycosidase activity have been generated against a chair-like transition state analogue. Two monoclonal antibodies with the highest activity were selected for cloning and sequencing. Sequence analysis of the two antibodies showed four amino acids differences in the framework region. Such a difference resulted in 8-fold difference in catalytic activity with p-nitrophenyl beta-D-glucopyranoside between the two antibodies. Several Asp and Glu residues were found in the complimentarity determining region and some of these residue(s) might form the catalytic core for the glycosidase. Cloned antibody genes were expressed as a single chain antibody fragment. The expressed proteins showed the retained glycosidase activities. PMID- 12132586 TI - Hepatitis C virus core inhibits the Fas-mediated p38 mitogen activated kinase signaling pathway in hepatocytes. AB - The p38 mitogen activated kinase (MAPK) signaling pathway plays an essential role in regulating many cellular processes, including inflammation, cell differentiation, and cell death. Here, we report that the hepatitis C virus (HCV) core inhibits the Fas-mediated p38 signaling pathway. The Fas-mediated p38 activation is suppressed in core-expressing HepG2 cell lines, as well as in the hepatocytes of transgenic mice. In addition, core protein blocked the Fas mediated activation of apoptosis signal-regulating kinase 1 (ASK1), a major upstream MAPKKK of p38. Treatment of a specific p38 inhibitor (SB203580) or overexpression of a kinase-defective mutant, ASK1 (K709R), promoted Fas-mediated cell death in HepG2 cells. This suggests that the p38 and ASK1 activation is required for cell survival against Fas-mediated cell death. In addition, we observed that the HCV core protein enhances Fas-mediated liver injury and lethality in transgenic mice. Collectively, our findings suggest that the HCV core inhibits the Fas-mediated p38 signaling pathway, which results in accelerated Fas-mediated cell death. PMID- 12132588 TI - Rac GTPase activity is essential for lipopolysaccharide signaling to extracellular signal-regulated kinase and p38 MAP kinase activation in rat-2 fibroblasts. AB - Lipopolysaccharide (LPS) has potent proinflammatory properties by acting on many cell types. Recently, mitogen-activated protein kinases (MAPKs) including extracellular signal-regulated kinase (ERK), p38 kinase, and c-jun N-terminal kinase (JNK) were shown to be involved in signal transduction in response to LPS. However, the detailed mechanism of LPS-induced signaling in the cell, especially the role of the Rho family GTPases remains largely unknown. In the present study, we investigated the role of Rac1, a member of the Rho family GTPases, in the LPS induced MAPKs activation in Rat-2 fibroblasts. Our results showed that LPS induced the activation of ERK and p38 MAP kinase in a Rac-dependent manner, suggesting a mediatory role of Rac1 in LPS signaling to MAPKs stimulation. We also observed that LPS caused a time-dependent activation of Rac1. In addition, our results have shown that pretreatment with herbimycin or wortmannin dramatically inhibited Rac1 activation induced by LPS. These suggest that tyrosine kinase(s) and phosphatidylinositol 3-kinase (PI 3-kinase) are possibly acting upstream of Rac1 in the LPS signaling to MAPKs. PMID- 12132589 TI - Oxidative DNA damage and alteration of glutamate transporter expressions in the hippocampal Ca1 area immediately after ischemic insult. AB - Although oxidative stress and excitotoxicity may be interdependent mechanisms that are involved in delayed neuronal death, the temporal participation of these events in the early stage after ischemia-reperfusion insult is unclear. Therefore, in the present study, using the gerbil global ischemic model we investigated whether oxidative stress could be correlated with the expression of the glutamate transporters in the hippocampus, and whether these events are related and cooperate in the events that link ischemia to neuronal death in vivo. Thirty minutes after ischemia, the intensities of glutamate transporter-1 (GLT 1), glutamate/aspar-tate transporter (GLAST), and 8-hydroxy2'-deoxy-guanosine (8 OHdG) immunoreactivities were markedly increased in the hippocampal CA1 area. In contrast, excitatory amino acid carrier-1 (EAAC-1) immunoreactivity was 30% lower in the CA1 area than in the sham level. At 3 h post-reperfusion, the EAAC-1 expression began to increase in the CA1 area. Twelve hours after reperfusion, the reduction of both GLT-1 and GLAST immunoreactivity was salient, while the EAAC-1 immunoreactivity level intensified significantly. The 8-OHdG immunoreactivity peaked at this time point. These findings suggest that oxidative stress and alterations in the glutamate transporter expression in the CA1 area may simultaneously trigger neuronal damages very early after ischemia. PMID- 12132590 TI - Quantification of tumor suppressor mRNA expression by poly-competitive RT-PCR using a TS-IS that contained multiple internal competitors. AB - Despite the recent introduction of real-time PCR methods and cDNA microarrays, competitive PCR techniques continue to play an important role in nucleic acid quantification because of the significantly lower cost of equipment and consumables. In this study, we developed a construct, termed tumor suppressor internal standard (TS-IS) that produced polycompetitive RNA templates as an internal standard to quantify cellular RNA concentration of tumor suppressor genes. This construct is composed of not only sets of primers for detecting the expression of several tumor suppressor genes (such as pRB, p16(INK4A) 15(INK4B), p14(ARF) p53, and p21(WAF1)), but also HPRT as an endogenous marker. Using an internal standard RNA that was synthesized from the TS-IS construct, we were able to establish optimized conditions for the quantification of tumor suppressor genes with minimal amounts (50 ng) of cellular RNA. In addition, the usefulness of this method was confirmed by analyzing the expression levels of tumor suppressor genes in fourteen hepatoma cell lines as a model. The TS-IS assay that we used was inexpensive and a widely applicable method that permitted the reliable and accurate quantification of tumor suppressor genes. PMID- 12132591 TI - Genomic organization and identification of a novel alternative splicing variant of mouse mitochondrial thioredoxin reductase (TrxR2) gene. AB - Eukaryotic mitochondria are equipped with a complete thioredoxin system, composed of thioredoxin and thioredoxin reductase, which has been implicated in the protection against the reactive oxygen intermdiates generated during the respiratory process in this organelle. Like its cytosolic counterpart, mammalian mitochondrial thioredoxin reductase is a homodimeric selenoprotein. We report here the genomic organization of the mouse mitochondrial thioredoxin gene (TrxR2) that spans 53 kb and consists of 18 exons ranging from 20 to 210 bp. All splicing sites conformed to the GT/AG rule with the exon-intron boundaries located exactly at the same position as the human TrxR2 gene, the only mammalian mitochondrial thioredoxin reductase gene whose genomic structure has been elucidated to date. In addition, we have identified a novel mRNA splicing variant lacking intron 14 resulting in a protein subunit with a shorter interface domain. This new splicing variant provides a framework for further analysis of this important enzyme as its predicted homodimeric conformation can now be expanded to a putative heterodimeric structure as well as a small subunit homodimer with the obvious implications at the regulatory level. PMID- 12132592 TI - Involvement of subcomplexes of 32 and 14 kDa subunits in RPA's DNA binding activity through redox change. AB - The eukaryotic replication protein A (RPA) is a heterotrimeric protein complex. It consists of 70, 32, and 14 kDa subunits that are involved in DNA replication, repair, and genetic recombination. RPA is a 4-cysteine type zinc-finger protein. RPA's zinc-finger domain is not essential for DNA binding activity, but it is involved in the regulation of RPA's DNA binding activity through reduction oxidation (redox). In this study, we show that yeast RPA's ssDNA binding activity is regulated by redox potential through its subcomplexes of 32 and 14 kDa subunits. In contrast, the subunits' complex, RPA70, formed a stable complex with ssDNA, even under non-reducing conditions. The addition of DTT and H202 had no effect on its DNA binding activity. In RPA70, since the addition of the subcomplexes of the 32 and 14 kDa subunits, it restored the modulating ssDNA binding activity to native RPA's DNA binding activity. These results suggest that the subcomplexes of the 32 and 14 kDa subunits may be involved in the modulating RPA's DNA binding activity through redox change. These studies, therefore, show the novel structure and function relationship of a multiprotein complex in that the role of a specific domain (or one subunit) is regulated by the other subunits. PMID- 12132593 TI - Carnosine and related dipeptides protect human ceruloplasmin against peroxyl radical-mediated modification. AB - Ceruloplasmin (CP) is the major plasma antioxidant and copper transport protein. In a previous study, we showed that the aggregation of human ceruloplasmin was induced by peroxyl radicals. We investigated the effects of antioxidant dipeptides carnosine, homocarnosine and anserine on peroxyl radical-mediated ceruloplasmin modification. Carnosine, homocarnosine and anserine significantly inhibited the aggregation of CP induced by peroxyl radicals. When CP was incubated with peroxyl radicals in the presence of three compounds, ferroxidase activity, as measured by the activity staining method, was protected. All three compounds also inhibited the formation of dityrosine in peroxyl radicals-treated CP. The results suggest that carnosine and related compounds act as peroxyl radical scavenger to protect the protein modification. It is proposed that carnosine and related peptides might be explored as potential therapeutic agents for pathologies that involve CP modification mediated by peroxyl radicals generated in the lipid peroxidation. PMID- 12132594 TI - Primary care, self-rated health, and reductions in social disparities in health. AB - OBJECTIVE: To examine the extent to which good primary-care experience attenuates the adverse association of income inequality with self-reported health. DATA SOURCES: Data for the study were drawn from the Robert Wood Johnson Foundation sponsored 1996-1997 Community Tracking Study (CTS) Household Survey and state indicators of income inequality and primary care. STUDY DESIGN: Cross-sectional, mixed-level analysis on individuals with a primary-care physician as their usual source of care. The analyses were weighted to represent the civilian noninstitutionalized population of the continental United States. DATA COLLECTION/EXTRACTION METHODS: Principal component factor analysis was used to explore the stricture of the primary-care indicators and examine their construct validity. Income inequality for the state in which the community is located was measured by the Gini coefficient, calculated using income distribution data from the 1996 current population survey. Stratified analyses compared proportion of individuals reporting had health and feeling depressed with those with good and bad primary-care experiences for each of the four income-inequality strata. A set of logistic regressions were performed to examine the relation between primary care experience, income inequality, and self-rated health. PRINCIPAL FINDINGS: Good primary-care experience, in particular enhanced accessibility and continuity, was associated with better self-reported health both generally and mentally. Good primary-care experience was able to reduce the adverse association of income inequality with general health although not with mental health, and was especially beneficial in areas with highest income inequality. Socioeconomic status attenuated, but did not eliminate, the effect of primary-care experience on health. In conclusion, good primary-care experience is associated not only with improved self-rated overall and mental health but also with reductions in disparities between more- and less-disadvantaged communities in ratings of overall health. PMID- 12132595 TI - Can high quality overcome consumer resistance to restricted provider access? Evidence from a health plan choice experiment. AB - OBJECTIVE: To investigate the impact of quality information on the willingness of consumers to enroll in health plans that restrict provider access. DATA SOURCES AND SETTING: A survey administered to respondents between the ages of 25 and 64 in the West Los Angeles area with private health insurance. STUDY DESIGN: An experimental approach is used to measure the effect of variation in provider network features and information about the quality of network physicians on hypothetical plan choices. Conditional logit models are used to analyze the experimental choice data. Next, choice model parameter estimates are used to simulate the impact of changes in plan features on the market shares of competing health plans and to calculate the quality level required to make consumers indifferent to changes in provider access. PRINCIPAL FINDINGS: The presence of quality information reduced the importance of provider network features in plan choices as hypothesized. However, there were not statistically meaningful differences by type of quality measure (i.e., consumer assessed versus expert assessed). The results imply that large quality differences are required to make consumers indifferent to changes in provider access. The impact of quality on plan choices depended more on the particular measure and less on the type of measure. Quality ratings based on the proportion of survey respondents "extremely satisfied with results of care" had the greatest impact on plan choice while the proportion of network doctors "affiliated with university medical centers" had the least. Other consumer and expert assessed measures had more comparable effects. CONCLUSIONS: Overall the results provide empirical evidence that consumers are willing to trade high quality for restrictions on provider access. This willingness to trade implies that relatively small plans that place restrictions on provider access can successfully compete against less restrictive plans when they can demonstrate high quality. However, the results of this study suggest that in many cases, the level of quality required for consumers to accept access restrictions may be so high as to be unattainable. The results provide empirical support for the current focus of decision support efforts on consumer assessed quality measures. At the same time, however, the results suggest that consumers would also value quality measures based on expert assessments. This finding is relevant given the lack of comparative quality information based on expert judgment and research suggesting that consumers have apprehensions about their ability to meaningfully interpret performance-based quality measures. PMID- 12132596 TI - Is managed care leading to consolidation in health-care markets? AB - OBJECTIVE: To determine the extent to which managed care has led to consolidation among hospitals and physicians. DATA SOURCES: We use data from the American Hospital Association, American Medical Association, and government censuses. STUDY DESIGN: Two stage least squares regression analysis examines how cross section variation in managed care penetration affects provider consolidation, while controlling for the endogeneity of managed-care penetration. Specifically, we examine inpatient hospital markets and physician practice size in large metropolitan areas. DATA COLLECTION METHODS: All data are from secondary sources, merged at the level of the Primary Metropolitan Statistical Area. PRINCIPAL FINDINGS: We find that higher levels of local managed-care penetration are associated with substantial increases in consolidation in hospital and physician markets. In the average market (managed-care penetration equaled 34 percent in 1994), managed care was associated with an increase in the Herfindahl of .054 between 1981 and 1994, moving from .096 in 1981 to .154. This is equivalent to moving from 10.4 equal-size hospitals to 6.5 equal-sized hospitals. In the physician market place, we estimate that at the mean, managed care resulted in a 14 percentage point decrease of physicians in solo practice between 1986 and 1995. This implies a decrease in the percentage of doctors in solo practice from 38 percent in 1986 to 24 percent by 1995. PMID- 12132598 TI - Concurrent prediction of hospital mortality and length of stay from risk factors on admission. AB - OBJECTIVE: To develop a method for predicting concurrently both hospital survival and length of stay (LOS) for seriously ill or injured patients, with particular attention to the competing risks of death or discharge alive as determinants of LOS. DATA SOURCES: Previously collected 1995-1996 registry data on 2,646 cases of injured patients from three trauma centers in Maine. STUDY DESIGN: Time intervals were determined for which the rates of discharge or death were relatively constant. Poisson regression was used to develop a model for each type of terminal event, with risk factors on admission contributing proportionately to the subsequent rates for each outcome in each interval. Mean LOS and cumulative survival were calculated from a combination of the resulting piecewise exponential models. PRINCIPAL FINDINGS: Age, Glasgow Coma Scale, Abbreviated Injury Scores, and specific mechanisms of injury were significant predictors of the rates of death and discharge, with effects that were variable in different time intervals. Predicted probability of survival and mean LOS from the model were similar to actual values for categorized patient groups. CONCLUSIONS: Piecewise exponential models may be useful in predicting LOS, especially if determinants of mortality are separated from determinants of discharge alive. PMID- 12132597 TI - Nurse staffing and postsurgical adverse events: an analysis of administrative data from a sample of U.S. hospitals, 1990-1996. AB - OBJECTIVE: To examine the impact of nurse staffing on selected adverse events hypothesized to be sensitive to nursing care between 1990 and 1996, after controlling for hospital characteristics. DATA SOURCES/STUDY SETTING: The yearly cross-sectional samples of hospital discharges for states participating in the National Inpatient Sample (NIS) from 1990-1996 were combined to form the analytic sample. Six states were included for 1990-1992, four states were added for the period 1993-1994, and three additional states were added in 1995-1996. STUDY DESIGN: The study design was cross-sectional descriptive. DATA COLLECTION/EXTRACTION METHODS: Data for patients aged 18 years and older who were discharged between 1990 and 1996 were used to create hospital-level adverse event indicators. Hospital-level adverse event data were defined by quality indicators developed by the Health Care Utilization Project (HCUP). These data were matched to American Hospital Association (AHA) data on community hospital characteristics, including registered nurse (RN) and licensed practical/vocational nurse (LPN) staffing hours, to examine the relationship between nurse staffing and four postsurgical adverse events: venous thrombosis/pulmonary embolism, pulmonary compromise after surgery, urinary tract infection, and pneumonia. Multivariate modeling using Poisson regression techniques was used. PRINCIPAL FINDINGS: An inverse relationship was found between RN hours per adjusted inpatient day and pneumonia (p < .05) for routine and emergency patient admissions. CONCLUSIONS: The inverse relationship between pneumonia and nurse staffing are consistent with previous findings in the literature. The results provide additional evidence for health policy makers to consider when making decisions about required staffing levels to minimize adverse events. PMID- 12132599 TI - Do hospital length of stay and staffing ratio affect elderly patients' risk of readmission? A nation-wide study of Norwegian hospitals. AB - OBJECTIVE: To test whether there is an association between hospital operating conditions such as average length of stays (LOS) and staffing ratio, and elderly patients' risk of readmission. DATA SOURCES: The main data source was a national patient database of admissions to all acute-care Norwegian hospitals during the year of 1996. STUDY DESIGN: It is a cross-sectional study, where Cox' regression analysis was used to test the factors acting on the probability of early unplanned readmission (within 30 days), and later occuring ones. The principal hospital variables included average hospital LOS and staffing ratio (discharges per man-years of personnel). Adjusting patient variables in the model included age, gender, and cost-weights of the Diagnosis Related Groups (DRGs). DATA EXTRACTION METHODS: The selected material included discharges from 59 hospitals, and 113,055 elderly patients (> or = 67 years). Multiple admissions to the same hospital were linked together chronologically, and additional hospital data were matched on. To maximize the association between the index stay and the defined outcome (unplanned readmission), no intervening planned admission was accepted. PRINCIPAL FINDINGS: Being admitted to a hospital with relatively short average LOS increased the patient's risk of early readmission significantly. In addition it was found that more intensive care (more staff) could have a compensatory effect. Furthermore, the predictive factors were shown to be time dependent, as hospital variables had much less impact on readmissions occurring late (within 90 180 days). CONCLUSIONS: The results give support to the assumption of a link between hospital operating conditions and patient outcome. PMID- 12132600 TI - Geographic variation in the use of post-acute care. AB - OBJECTIVE: To assess the extent and consistency of geographic differences in the use of post-acute care (PAC), and the stability of this pattern of variation. DATA SOURCES: The 5 percent Medicare data sample for 1996, 1997, and the first eight months of 1998 were used. STUDY DESIGN: Patterns of PAC use for various Diagnosis-related Groups (DRGs) cross states (33 with enough cases per year) and census divisions were examined. The consistency of relative rankings for overall PAC use and use within defined DRGs was compared. PRINCIPAL FINDINGS: PAC use varied substantially across regions. For example, the extent of any PAC use for stroke patients varied by 12 percentage points among census regions in 1998. The pattern of PAC use was quite consistent across years; 30 of the 36 possible Spearman rank order correlations were statistically significant with coefficients ranging from 0.35 to 0.95 among the DRGs studied. The correlations among DRGs were generally high. For skilled nursing facility use, all the correlations were above 0.5 and were statistically significant; in general the patterns were highest within medical DRGs (0.65-0.93). CONCLUSIONS: The variation in PAC use is not a statistical artifact. It is likely the result of several forces: practice styles, supply of services, and local regulatory practices. PMID- 12132601 TI - The relationship of post-acute home care use to Medicaid utilization and expenditures. AB - RESEARCH OBJECTIVES: To describe the use of post-acute home care (PAHC) and total Medicaid expenditures among hospitalized nonelderly adult Medicaid eligibles and to test whether health services utilization rates or total Medicaid expenditures were lower among Medicaid eligibles who used PAHC compared to those who did not. STUDY POPULATION: 5,299 Medicaid patients aged 18-64 discharged in 1992-1996 from 29 hospitals in the Cleveland Health Quality Choice (CHQC) project. DATA SOURCES: Linked Ohio Medicaid claims and CHQC medical record abstract data. DATA EXTRACTION: One stay per patient was randomly selected. DESIGN: Observational study. To control for treatment selection bias, we developed a model predicting the probability (propensity) a patient would be referred to PAHC, as a proxy for the patient's need for PAHC. We matched 430 patients who used Medicaid-covered PAHC ("USE") to patients who did not ("NO USE") by their propensity scores. Study outcomes were inpatient re-admission rates and days of stay (DOS), nursing home admission rates and DOS, and mean total Medicaid expenditures 90 and 180 days after discharge. PRINCIPAL FINDINGS: Of 3,788 medical patients, 12.1 percent were referred to PAHC; 64 percent of those referred used PAHC. Of 1,511 surgical patients, 10.9 percent were referred; 99 percent of those referred used PAHC. In 430 pairs of patients matched by propensity score, mean total Medicaid expenditures within 90 days after discharge were $7,649 in the USE group and $5,761 in the NO USE group. Total Medicaid expenditures were significantly higher in the USE group compared to the NO USE group for medical patients after 180 days (p < .05) and surgical patients after 90 and 180 days (p < .001). There were no significant differences for any other outcome. Sensitivity analysis indicates the results may be influenced by unmeasured variables, most likely functional status and/or care-giver support. CONCLUSIONS: Thirty-six percent of the medical patients referred to PAHC did not receive Medicaid-covered services. This suggests potential underuse among medical patients. The high post-discharge expenditures suggest opportunities for reducing costs through coordinating utilization or diverting it to lower-cost settings. Controlling for patients' need for services, PAHC utilization was not associated with lower utilization rates or lower total Medicaid expenditures. Medicaid programs are advised to proceed cautiously before expanding PAHC utilization and to monitor its use carefully. Further study, incorporating non-economic outcomes and additional factors influencing PAHC use, is warranted. PMID- 12132602 TI - A national study of efficiency for dialysis centers: an examination of market competition and facility characteristics for production of multiple dialysis outputs. AB - OBJECTIVE: To examine market competition and facility characteristics that can be related to technical efficiency in the production of multiple dialysis outputs from the perspective of the industrial organization model. STUDY SETTING: Freestanding dialysis facilities that operated in 1997 submitted cost report fonns to the Health Care Financing Administration (HCFA), and offered all three outputs--outpatient dialysis, dialysis training, and home program dialysis. DATA SOURCES: The Independent Renal Facility Cost Report Data file (IRFCRD) from HCFA was utilized to obtain information on output and input variables and market and facility features for 791 multiple-output facilities. Information regarding population characteristics was obtained from the Area Resources File. STUDY DESIGN: Cross-sectional data for the year 1997 were utilized to obtain facility specific technical efficiency scores estimated through Data Envelopment Analysis (DEA). A binary variable of efficiency status was then regressed against its market and facility characteristics and control factors in a multivariate logistic regression analysis. PRINCIPAL FINDINGS: The majority of the facilities in the sample are functioning technically inefficiently. Neither the intensity of market competition nor a policy of dialyzer reuse has a significant effect on the facilities' efficiency. Technical efficiency is significantly associated, however, with type of ownership, with the interaction between the market concentration of for-profits and ownership type, and with affiliations with chains of different sizes. Nonprofit and government-owned Facilities are more likely than their for-profit counterparts to become inefficient producers of renal dialysis outputs. On the other hand, that relationship between ownership form and efficiency is reversed as the market concentration of for-profits in a given market increases. Facilities that are members of large chains are more likely to be technically inefficient. CONCLUSIONS: Facilities do not appear to benefit from joint production of a variety of dialysis outputs, which may explain the ongoing tendency toward single-output production. Ownership form does make a positive difference in production efficiency, but only in local markets where competition exists between nonprofit and for-profit facilities. The increasing inefficiency associated with membership in large chains suggests that the growing consolidation in the dialysis industry may not, in fact, be the strategy for attaining more technical efficiency in the production of multiple dialysis outputs. PMID- 12132603 TI - Use of the SF-12 instrument for measuring the health of homeless persons. AB - OBJECTIVE: To evaluate the construct validity of the Short Form 12-item Survey (SF-12) among users of a homeless day shelter. Adding brief health assessments has potential to provide information regarding the effect that programs have upon the health status and functioning of homeless persons. STUDY SETTING: A convenience sample of 145 homeless persons at a day shelter in an urban setting. STUDY DESIGN: Participants were verbally administered the SF-12 that provides information on mental and physical health status and the Dartmouth Improve Your Medical Care Survey (IYMC) that provides information on functional health, clinical symptoms, medical conditions, and health risk. The IYMC survey system has been widely used in clinical settings to assess health status and the outcomes of care. DATA COLLECTION/EXTRACTION METHODS: Construct validity was assessed by the following approaches: (a) the method of extreme groups was used where multivariate analysis of variance determined if SF-12 summary scores varied for individuals who differed in self-reported clinical symptoms and medical conditions, and (b) convergent validity was assessed by correlating SF-12 summary scores with the subscales. PRINCIPAL FINDINGS: Four to 10 point differences in physical health (PCS12) and 5-11 point differences in mental health (MCS12) were found between those who reported acute symptoms and medical conditions and those who did not. A 13-point difference in PCS12 scores and a 7-16-point difference in MCS12 scores were found for those who reported none or few to several symptoms or conditions. The summary scores and subscales yielded satisfactory convergent validity coefficients that ranged from 0.62 to 0.88 with one exception. CONCLUSIONS: The SF-12 shows promise as a valid outcome indicator for assessing and monitoring health status among the homeless. Its strengths include brevity and availability of norms for specific medical conditions. PMID- 12132604 TI - Agreement between self-reported and routinely collected health-care utilization data among seniors. AB - OBJECTIVE: To examine the agreement between self-reported and routinely collected administrative health-care utilization data, and the factors associated with agreement between these two data sources. DATA SOURCES/STUDY SETTING: A representative sample of seniors living in an Ontario county within Canada was identified using the Ontario Ministry of Health's Registered Persons Data Base in 1992. Health professional billing information and hospitalization data were obtained from the Ontario Ministry of Health and Long-Term Care (OMH) and the Ontario Health Insurance Plan (OHIP). STUDY DESIGN: A cross-sectional survey was carried out to assess any contact and frequency of contacts with health professionals and hospital admissions. Similar information was obtained from routinely collected administrative data. The level of agreement was assessed using the proportion of absolute agreement, Cohen's kappa statistic (kappa), and the intraclass correlation coefficient (ICC). Logistic and linear regressions were used to identify factors that were associated with the magnitude and direction of disagreement respectively. DATA COLLECTION/EXTRACTION METHODS: Telephone interviews were conducted on 1,054 seniors, and complete data were available for 1,038 seniors. Each respondent's personal health number was used to electronically link survey data with health professional billing and hospitalization databases. PRINCIPAL FINDINGS: Substantial to almost perfect agreement was found for the contact utilization measures, while agreement on volume utilization measures varied from poor to almost perfect. In surveys, seniors overreported contact with general practitioners and physiotherapists or chiropractors, and underreported contact with other medical specialists. Seniors also underreported the number of contacts with general practitioners and other medical specialists. The odds of agreement decreased if respondents were male, aged 75 years and older, had incomes of less than $25,000, had poor/fair/good self-assessed health status, or had two or more chronic conditions. CONCLUSION: The findings of this study indicate that there are substantial discrepancies between self-reported and administrative data among older adults. Researchers seeking to examine health-care use among older adults need to consider these discrepancies in the interpretation of their results. Failure to recognize these discrepancies between survey and administrative data among older adults may lead to the establishment of inappropriate health-care policies. PMID- 12132605 TI - Clinical effectiveness research in managed-care systems: lessons from the Pediatric Asthma Care PORT. Patient Outcomes Research Team. AB - OBJECTIVE: To highlight the unique challenges of evaluative research on practice behavior change in the "real world" settings of contemporary managed-care organizations, using the experience of the Pediatric Asthma Care PORT (Patient Outcomes Research Team). STUDY SETTING: The Pediatric Asthma Care PORT is a five year initiative funded by the Agency for Healthcare Research and Quality to study strategies for asthma care improvement in three managed-care plans in Chicago, Seattle, and Boston. At its core is a randomized trial of two care improvement strategies compared with usual care: (1) a targeted physician education program using practice based Peer Leaders (PL) as change agents, (2) adding to the PL intervention a "Planned Asthma Care Intervention" incorporating joint "asthma check-tips" by nurse-physician teams. During the trial, each of the participating organizations viewed asthma care improvement as an immediate priority and had their own corporate improvement programs underway. DATA COLLECTION: Investigators at each health plan described the organizational and implementation challenges in conducting the PAC PORT randomized trial. These experiences were reviewed for common themes and "lessons" that might be useful to investigators planning interventional research in similar care-delivery settings. CONCLUSIONS: Randomized trials in "real world" settings represent the most robust design available to test care improvement strategies. In complex, rapidly changing managed-care organizations, blinding is not feasible, corporate initiatives may complicate implementation, and the assumption that a "usual care" arm will be static is highly likely to be mistaken. Investigators must be prepared to use innovative strategies to maintain the integrity of the study design, including: continuous improvement within the intervention arms, comanagement by researchers and health plan managers of condition-related quality improvement initiatives, procedures for avoiding respondent burden in health plan enrollees, and anticipation and minimization of risks from experimental arm contamination and major organizational change. With attention to these delivery system issues, as well as the usual design features of randomized trials, we believe managed-care organizations can serve as important laboratories to test care improvement strategies. PMID- 12132607 TI - Evaluation of greater petrosal nerve function in patients with acute peripheral facial paralysis: comparison of soft palate electrogustometry and Schirmer's tear test. AB - We tested sensory and secretomotor function of the greater petrosal nerve (GPN) by means of electrogustometry (EGM) of the soft palate and Schirmer's tear test in 115 patients (59 males, 56 females) with acute peripheral facial paralysis. Facial paralysis was caused by Bell's palsy in 78 cases, Ramsay Hunt syndrome in 27 cases and zoster sine herpetic lesions in 10. All patients had dysfunction of the stapedial nerve. An electrogustometer was used to test taste (GPN sensory function), and elevation of the threshold by > 6 dB on the affected side was considered abnormal. Schirmer's test was used to evaluate lacrimal (GPN secretomotor) function, which was considered abnormal when tear secretion on the affected side was < 50% of secretion on the non-affected side. Of the 78 patients with Bell's palsy, 28.2% had altered taste on the soft palate (sensory dysfunction) and 10.3% had lacrimal dysfunction, indicating that EGM of the soft palate is more sensitive than Schirmer's test for identifying dysfunction of the GPN in patients with facial paralysis due to Bell's palsy. Of the total of 115 patients, 32 (28%) had taste dysfunction and 9 (28.1%) of these 32 patients also had lacrimal dysfunction. This finding indicates that facial paralysis has different effects on the sensory and secretory nerve fibers of the GPN. The results of Schirmer's test were more closely related to the severity of, and prognosis for, facial paralysis than the results of EGM. PMID- 12132606 TI - Assessment of chronic illness care (ACIC): a practical tool to measure quality improvement. AB - OBJECTIVE: To describe initial testing of the Assessment of Chronic Illness Care (ACIC), a practical quality-improvement tool to help organizations evaluate the strengths and weaknesses of their delivery of care for chronic illness in six areas: community linkages, self-management support, decision support, delivery system design, information systems, and organization of care. DATA SOURCES: (1) Pre-post, self-report ACIC data from organizational teams enrolled in 13-month quality-improvement collaboratives focused on care for chronic illness; (2) independent faculty ratings of team progress at the end of collaborative. STUDY DESIGN: Teams completed the ACIC at the beginning and end of the collaborative using a consensus format that produced average ratings of their system's approach to delivering care for the targeted chronic condition. Average ACIC subscale scores (ranging from 0 to 11, with 11 representing optimal care) for teams across all four collaboratives were obtained to indicate how teams rated their care for chronic illness before beginning improvement work. Paired t-tests were used to evaluate the sensitivity. of the ACIC to detect system improvements for teams in two (of four) collaboratives focused on care for diabetes and congestive heart failure (CHF). Pearson correlations between the ACIC subscale scores and a faculty rating of team performance were also obtained. RESULTS: Average baseline scores across all teams enrolled at the beginning of the collaboratives ranged from 4.36 (information systems) to 6.42 (organization of care), indicating basic to good care for chronic illness. All six ACIC subscale scores were responsive to system improvements diabetes and CHF teams made over the course of the collaboratives. The most substantial improvements were seen in decision support, delivery system design, and information systems. CHF teams had particularly high scores in self-management support at the completion of the collaborative. Pearson correlations between the ACIC subscales and the faculty rating ranged from .28 to .52. CONCLUSION: These results and feedback from teams suggest that the ACIC is responsive to health care quality-improvement efforts and may be a useful tool to guide quality improvement in chronic illness care and to track progress over time. PMID- 12132608 TI - Drug-related taste disturbances. AB - Drug-induced change in taste is the second most common cause of taste disturbance among our patients, accounting for approximately 25% of cases. About 170 drugs have been associated with taste disturbances, either when used alone or in combinations. We studied the zinc-chelating capability of 20 drugs associated with taste disturbance, using the pH titration test, DC polarography, spectrophotometry and tests of intestinal absorption of zinc in the presence of these drugs. The results of these analyses and our review of the literature indicate that there are a number of possible mechanisms for drug-related taste disturbance and that zinc plays a key role. PMID- 12132609 TI - Electron microscopic observations of glossal circumvallate papillae in dysgeusic patients. AB - A number of reports have been published describing the ultrastructure of normal taste buds but few have discussed their pathology, particularly in patients with gustatory abnormalities. We therefore conducted an electron microscopic study of biopsy specimens of glossal circumvallate papillae from patients with dysgeusia. We found depletion of microvilli and dense substance in the taste pores, vacuolation and hyaline degeneration of the taste bud cells and condensation of nuclear material and cytoplasm in cells of indeterminate histological type. In one patient with idiopathic dysgeusia, disruption of subsurface cisternae contiguous with the nerve endings was evident in Type II cells. In another patient, who had undergone radiotherapy, the number of cored vesicles in Type III cells was reduced. PMID- 12132610 TI - Double-blind, placebo-controlled trial of zinc picolinate for taste disorders. AB - A total of 73 patients with an idiopathic zinc-deficiency taste disorder completed a double-blind, placebo-controlled trial of the efficacy of zinc picolinate. Patients in the zinc treatment group (n = 37) received 29 mg of zinc picolinate orally by capsule 3 times a day for 3 months. No significant difference was noted between the treatment and placebo groups in terms of improvement in subjective symptoms or whole-mouth taste sensation. However, assessment of taste using the filter paper disk method showed significantly better improvement for the group given zinc, and the serum zinc level was also significantly higher in this group than in the placebo control group. PMID- 12132611 TI - Secretory function of the salivary gland in patients with taste disorders or xerostomia: correlation with zinc deficiency. AB - A notable proportion of patients with taste disorders complain of xerostomia and when zinc is prescribed the xerostomia is often improved in conjunction with the taste disorder. To study the relationship between taste disorders, zinc deficiency and xerostomia, we measured salivary gland function and zinc levels in 93 patients with hypogeusia and/or xerostomia and 60 patients with unilateral acute peripheral facial palsy who served as controls. We then prescribed zinc for patients with low serum zinc levels and evaluated xerostomia and taste sensation after 6 months of this treatment. The salivary gland secretory ratio (SGSR), determined by dynamic salivary 99mTc scintigraphy, was found to be an objective measure of salivary gland function and was reduced in patients with xerostomia. Patients with salivary gland dysfunction also had abnormal morphology of the papillae of the tongue. No significant relation was found between the severity of taste disorders and SGSR values, but low SGSR values were found in patients with zinc deficiency. Patients with taste disorders and/or xerostomia who were treated with zinc had relief of symptoms at 6 months, indicating that both taste disorders and xerostomia are among the symptoms of zinc deficiency. PMID- 12132613 TI - Changes in gustatory sense during pregnancy. AB - Changes in gustatory sense were investigated in 97 pregnant women and in 30 healthy, non-pregnant women who served as controls. All 97 pregnant women completed a questionnaire regarding taste changes and 32 of them underwent serial tests of gustatory function, including electrogustometry and testing with filter paper disks placed over the areas of the chorda tympani nerve and glossopharyngeal nerve. All gustatory testing was performed by the same person. In addition, serum levels of trace elements were measured in 72 of the pregnant women. Of the 97 pregnant women, 90 (92.8%) reported in the questionnaire that they had experienced some type of change in taste during pregnancy, usually a change in sour taste (59 women; 65.6%). Pregnant women had higher gustatory thresholds than non-pregnant women, with an especially marked decrease in gustatory function being noticed in the first trimester. Although serum zinc levels decreased in pregnant women between the second and third trimesters, zinc levels were in the normal range in the early stage of pregnancy. Thus, it is difficult to explain dysgeusia in the early stage of pregnancy as being associated with a deficiency of zinc. The decrease in gustatory function during the first trimester is considered to be due to the notable changes in secretion of hormones that occur during this stage. PMID- 12132612 TI - Incidence and treatment of dysgeusia in patients with glossodynia. AB - In 96 patients who visited our hospital with glossodynia, we conducted gustatory tests, measured serum zinc and copper levels, examined lingual papillae using biomicroscopy, conducted psychological tests and investigated the effectiveness of treatments directed at the cause of dysgeusia. Gustatory test results showed that 43 (44.8%) of the patients had dysgeusia, which was mild in 62.8%, moderate in 30.2% and severe in 7.0%. By giving higher priority to treatment of dysgeusia than to glossodynia, pain disappeared or was relieved and gustatory sensation improved in 27 (62.8%) of these 43 patients. Overall, pain disappeared or was improved in 65 (67.7%) cases. When treating glossodynia, it is important to diagnose the cause of pain and to give higher priority to treating that cause. The clinical efficacy of treatment for glossodynia will be improved when the presence or absence of dysgeusia is diagnosed early in the course of treatment. PMID- 12132614 TI - A case of Cronkhite-Canada syndrome whose major complaint, taste disturbance, was improved by zinc therapy. AB - A patient whose major complaint was taste disturbance and who was diagnosed as having Cronkhite-Canada syndrome was prescribed zinc sulfate. Improvement in taste disturbance was noted after 3 weeks of treatment, followed by gradual improvement in skin and gastrointestinal symptoms. Cronkhite-Canada syndrome can be considered a zinc-deficiency disorder caused by gastrointestinal polyposis. PMID- 12132615 TI - Taste disorder involving Hunter's glossitis following total gastrectomy. AB - We treated five patients with Hunter's glossitis following total gastrectomy. The major complaints of the patients were taste disorder and abnormal glossal sensation. In all five cases, the patient's tongue was red and smooth, and laboratory testing showed the presence of macrocytic anemia and decreased serum concentration of vitamin B12 (cyanocobalamin). Gustometry was carried out in four cases and the results documented the presence of taste disorder. All five patients were treated by administration of vitamin B12, which led to improvements in the appearance of the tongue, the patients' subjective complaints and the results of taste testing. When patients present with a red, smooth tongue, Hunter's glossitis (which can easily be improved by administration of vitamin B12) should be considered in the differential diagnosis. PMID- 12132616 TI - Influence of aging on electrogustometry thresholds. AB - Taste thresholds were measured using electrogustometry by the same researcher at three pairs of sites in the mouth (soft palate, tongue tip and tongue base) in both smokers and non-smokers (n = 461; age range 15-94 years). Comparison of the results by decades showed that the threshold increased significantly with age, starting at age 60 for areas innervated by the chorda tympani and glossopharyngeal nerves and at age 70 for areas innervated by the greater petrosal nerve. For the older teenage subjects, females had significantly lower thresholds than males and thresholds for women tended to be lower than those for men at all sites examined. Thresholds for the two sides of the tongue were within approximately 6 dB of each other. Thresholds on the soft palate were consistently higher than those on the tongue tip and tongue base. Little difference was detected between thresholds on the tongue tip and those on the tongue base except in older teenage subjects, in whom thresholds on the tongue tip were significantly lower than those on the tongue base. Male smokers in their 30s and 40s had significantly lower thresholds on the soft palate than non-smokers in this age group. PMID- 12132617 TI - Taste disturbance after tonsillectomy. AB - Of the 3583 outpatients treated at our taste disorder clinic over a period of 15 years, 11 (0.31%) complained of taste disorder after tonsillectomy. The cause of taste disorder was identified in 8 of the 11 cases: in 3 cases it was caused by direct or indirect damage to the lingual branch of the glossopharyngeal nerve; in 2 cases it was attributable to medication taken by the patient after tonsillectomy; and in 3 cases taste disturbance was caused by a lack of dietary zinc, even though this was identified at the time of tonsillectomy. These findings indicate the importance of (i) informing patients when consent for tonsillectomy is obtained that there is a risk of postoperative taste disorder; (ii) measuring the patient's taste threshold and serum zinc level preoperatively; and (iii) obtaining a thorough drug history, including details of non prescription medications habitually taken by the patient. PMID- 12132618 TI - Dysgeusia due to an orthodontic wire: a case report. AB - A 14-year-old female developed gustatory disorder due to an orthodontic wire having pierced the right trigonal retromolar. The patient's complaints included traction pain on the right lower jaw, numbness on the right front half of the tongue and hypogeusia with the exception of sweet tastes. Possible causes of dysgeusia in this case were: (i) direct mechanical compression of a gustatory nerve by the orthodontic wire; and (ii) disturbance of blood supply to a gustatory nerve by edema that developed in nerve tissues around the wire. The patient's retention of a normal threshold for the recognition of sweet tastes is not fully understood. PMID- 12132619 TI - Clinical use of electrogustometry: strengths and limitations. AB - Electrogustometry (EGM) has a number of strengths and a few limitations in clinical use. The strengths of EGM are: (i) the range of measurements can be kept constant; (ii) quantitative control of the intensity of the stimulation is possible; (iii) only a short period of time is required for testing; (iv) it is possible to detect even slight taste disorders for which the patient has no subjective symptoms; (v) it is useful for topognosis of lesions of taste pathways and for determining prognosis; and (vi) it is the only quantitative method for diagnosing disorders of the glosssopharyngeal nerve. The limitations of EGM are: (i) it is not useful for determining or diagnosing some of the symptoms often complained of by patients with taste disorder, namely dissociated taste disorder, heterogeusia and spontaneous dysgeusia; and (ii) it is not useful for following the progress of taste disorder. The many strengths and few limitations of EGM make it the first choice among taste examinations. This paper describes the clinical use of EGM as well as discussing other taste examinations used in our taste clinic and, in particular, the advantages and disadvantages of filter paper disk testing with taste solutions. PMID- 12132620 TI - A new whole-mouth gustatory test procedure. 1. Thresholds and principal components analysis in healthy men and women. AB - Gustatory testing using the whole-mouth method was performed in 123 healthy young male and female subjects. The average thresholds for detection and recognition of the four basic tastes were not greatly different from the normal thresholds previously reported in Japan: a 0.0165 M solution of sucrose for sweet taste, a 0.0316 M solution of table salt for salty taste, a 0.000743 M solution of tartaric acid for sour taste and a 0.0000203 M solution of quinine hydrochloride for bitter taste. These results indicate that the whole-mouth gustatory test procedure employed in this study may be useful for evaluating gustatory function clinically. Principal components analysis confirmed that sweet, salty, sour and bitter are indeed the four basic tastes and revealed that the sensation of taste is detected before the specific taste is recognized, regardless of the specific taste tested. PMID- 12132621 TI - A new whole-mouth gustatory test procedure. II. Effects of aging, gender and smoking. AB - Taste detection and recognition thresholds for the 4 basic tastes were measured in 670 healthy subjects using a new whole-mouth method in which test solutions of the 4 basic tastes were diluted 1:1 with distilled water successively in 13 steps. The taste thresholds were measured by spraying a 1 ml aliquot of solution into the mouth, starting with the lowest concentration, and asking the subject to swallow and note if a taste was detected and, if so, which one. The mouth was rinsed with distilled water only between different taste test solutions. Multiple comparison analysis showed a clear age-related increase in thresholds for salty, sour and bitter tastes for subjects in their eighth decade of life. From the third decade onwards, female subjects had significantly lower thresholds compared to males for sour taste in about half of the age groups and for salty and bitter tastes in some age groups. In contrast, for 18- and 19-year-old subjects, the gustatory thresholds of male subjects were lower than those of female subjects. Regarding smoking, male smokers in their third decade had significantly higher thresholds for bitter taste compared to male non-smokers in this age group. PMID- 12132622 TI - A review of objective measures of gustatory function. AB - Several studies of diagnostic tests of gustatory function have been reported. Results show that none of the subjective tests are satisfactory because they cannot detect psychogenic disease or malingering. Of the objective modalities that have been investigated, gustatory-evoked potential testing has been the most widely studied because it does not require expensive equipment and can be used in multicenter studies. The techniques developed to date for measuring gustatory evoked potentials are not yet clinically useful for diagnosing taste disorders. Future directions for improving this method of evaluating gustatory function include improving stimulation methods and equipment, in order to obtain better stimulation-related potentials and better means of distinguishing between normal and abnormal gustatory-evoked potential patterns, and applying these new standards to the evaluation of data. PMID- 12132623 TI - Gustatory function of the soft palate. AB - Several characteristics of the gustatory function of the soft palate principally elucidated by our own recent investigations, as well of those of others, are described. The soft palate has a gustatory function which is independent of the tongue and is innervated by the greater petrosal nerve. Taste papillae of the soft palate are morphologically similar to the fungiforme papillae which exist on the anterior part of the tongue. In individuals aged approximately 20 years, gustatory functions of the soft palate, examined by means of the electro gustatory and filter paper disk test's are as good as those of the tongue innervated by the chorda tympani and glossopharyngeal nerve. In younger individuals, the gustatory function of the soft palate plays as important a role as that of the tongue. Gustatory thresholds of the soft palate, as well as those of other sites innervated by other taste nerves, tend to be elevated with aging, and the degree of elevation in the soft palate is much greater than that of other sites. The most sensitively perceived taste at the soft palate is a sweet taste in the majority of subjects. Taste papillae of the palate in rats become flattened and decrease in number with age. These changes are more prominent in zinc-deficient rats. PMID- 12132624 TI - Characteristics of 2278 patients visiting the Nihon University Hospital Taste Clinic over a 10-year period with special reference to age and sex distributions. AB - We investigated the incidence of taste disorders in 2278 patients (871 men, 1407 women) who visited our Taste and Smell Clinic during the 10-year period between 1981 and 1990. Although a higher proportion of our patients were women, there was a significant gender difference only in the proportion of women with mild symptoms; the proportions of men and women with moderate or severe taste disorders were not significantly different. We propose that the incidence of taste disorders is the same among men and women but that women with milder symptoms are more likely to seek treatment. The time from onset of symptoms to the first clinic visit lengthened with increasing patient age. The efficacy of treatment decreased with increasing patient age, regardless of the severity of taste disorder at the initial visit. PMID- 12132625 TI - Sensitivity of three loci on the tongue and soft palate to four basic tastes in smokers and non-smokers. AB - The filter paper disk method was used to determine the map of the 4 tastes (sweet, salty, sour and bitter) on the tongue and soft palate in 69 healthy individuals. The equipment used for testing included filter paper disks 8 mm in diameter and taste solutions, which were sequentially diluted (in 13 stages from 80% sucrose, 12 stages from 20% NaCl, 12 stages from 8% tartaric acid and 14 stages from 4% quinine hydrochloride). The threshold at which each taste was recognized in three locations (center of the tongue tip, foliate papillae on the left side and center of the soft palate) was determined by testing increasing concentrations of each solution. We found no difference by sex for sweet taste but women were more sensitive to sour taste on the tongue tip and to salty and bitter tastes on the soft palate. No difference in taste threshold was noted at the different locations for sweet, salty or bitter tastes, but both sexes were less sensitive to sour taste on the soft palate than on the tongue tip, and in men the soft palate was even less sensitive to sour taste than the tongue root. Smoking was associated with increased (worse) taste thresholds for all four tastes, for both men and women. PMID- 12132626 TI - Course of recovery from taste receptor disturbance. AB - The course of recovery from taste receptor disturbance was studied in 119 patients with moderate-to-severe taste receptor disturbance that was cured or improved with zinc therapy. Taste receptor disturbance was idiopathic in 45 patients, drug-induced in 38 and due to zinc deficiency in 36. Recovery of taste, evaluated by filter paper disk testing and electrogustometry, followed 1 of 3 patterns: (i) in 54 (45.4%) of the 119 cases, taste improved simultaneously in the anterior (innervated by the chorda tympani nerve) and posterior (innervated by the glossopharyngeal nerve) portions of the tongue; (ii) in 53 (44.5%) of the cases, taste improved in the posterior portion first; and (iii) in 12 (10.1%) of the cases, taste improved in the anterior portion first. Zinc therapy was more effective in patients with the "posterior" pattern of recovery, and these patients also recovered the ability to sense sweet and bitter tastes earlier than other tastes. These results indicate that recovery of taste begins on the posterior portion of the tongue, which has an abundance of taste buds. The results of electrogustometry were not helpful in assessing recovery from taste disturbance, but testing for taste using the filter paper disk method on the posterior portion of the tongue was useful for identifying the onset of recovery. PMID- 12132627 TI - Taste disturbance in two patients after dental anesthesia by inferior alveolar nerve block. AB - We report two cases of temporary taste disturbance after inferior alveolar nerve block. The first patient to present with this rare complication of anesthesia for dental surgery was a 41-year-old woman. She lost the sense of taste on the left side of her tongue after local anesthesia for treatment of a left mandibular molar and first visited our outpatient clinic complaining of taste disorder 3 months later. Electrogustometry (EGM) and filter paper disk (FPD) testing revealed a taste disturbance in the innervation area of the left chorda tympani nerve and atrophy of the fungiform papillae on the left side of the tongue was observed. Eleven months after the dental treatment, the fungiform papillae and the results of EGM were normal. The second patient, a 22-year-old woman, received local anesthetic for extraction of a right mandibular molar and subsequently developed loss of taste on the right side of the tongue. When she visited our outpatient clinic 3 months later, atrophy of the fungiform papillae on the right side was observed. Her gustatory sense began to improve 4 months after the dental surgery and was normal at 13 months. From these findings we conclude that taste disturbance on the same side as the inferior alveolar nerve block in each case was due to direct injury to the chorda tympani and lingual nerves during administration of the local anesthetic. The results of EGM and FPD testing were diagnostic: atrophy of the fungiform papillae on the same side and disappearance of taste on the same side in the intravenous taste test provided complementary diagnostic information. The outcome was satisfactory in both cases. PMID- 12132628 TI - Anatomy of the tonsillar bed: topographical relationship between the palatine tonsil and the lingual branch of the glossopharyngeal nerve. AB - Taste disturbance may result from injury to the lingual branch of the glossopharyngeal nerve (LBGN) during tonsillectomy. Because an understanding of the anatomy of this nerve is required in order to avoid injuring it, a gross, histologic anatomic study was undertaken of the topographical relationship between the LBGN and the muscle layer of the palatine tonsillar bed. Evaluation of 107 sides of 83 Japanese adult cadavers (aged 27-88 years) confirmed that the muscular composition and lamination of the tonsillar bed do not change with age or pathological conditions such as inflammation. In about a quarter (23.4%) of cases, the LBGN traveled inferior to the styloglossus muscle and lateral to the superior constrictor pharyngeal muscle over its whole course to the base of the tongue, so that the palatine tonsil was clearly separated from the LBGN. In 55.1% of cases, however, the muscle lining of the tonsillar bed was discontinuous and thin muscle bundles, derived from the stylopharyngeus, palatopharyngeus or superior constrictor pharyngeal muscle, partially covered the tonsillar capsule externally. Moreover, in 21.5% of cases the LBGN was firmly adherent to the tonsillar capsule, due to the complete absence of muscles lining the tonsillar bed. In these cases, and also probably in a similar percentage of patients undergoing tonsillectomy, taste disturbance may occur on removal of the hypertrophic tonsillar capsule. Therefore, minimal disturbance of the tonsillar bed is recommended in all cases of tonsillectomy. PMID- 12132629 TI - Bad company: internet sites with dangerous information. PMID- 12132630 TI - Schizophrenia and the motivation for smoking. AB - PROBLEM: People with mental illness are twice as likely to smoke than people without a mental illness. METHODS: Data were collected through interviews with individuals who smoke and have been diagnosed with schizophrenia (N = 100). The research design included a descriptive, correlational design that described and examined the relationships among psychiatric symptoms, medication side effects, and reasons for smoking; and a qualitative analysis of the subjective experience of smoking. FINDINGS: A positive relationship was found between the age of onset of smoking and the onset of schizophrenia. Subjects reported they smoked primarily for sedative effects and control of negative symptoms of schizophrenia. Subjects also reported smoking related to addiction. Most indicated they would like to quit smoking or at least cut down on the number of cigarettes. CONCLUSIONS: Among people with schizophrenia, the motivation to smoke is related to their schizophrenia. PMID- 12132631 TI - Selective serotonin reuptake inhibitors and treatment of premenstrual dysphoric disorder. AB - TOPIC: Premenstrual dysphoric disorder (PMDD) has reentered the spotlight following the FDA's recent approval of fluoxetine hydrochloride to treat its symptoms. Although the diagnosis and treatment of PMDD has long been a source of contention, the FDA move has heightened the debate over this diagnostic category and the most appropriate treatment. PURPOSE: To explore several diagnoses related to PMDD and review recent research findings pertaining to the effectiveness of SSRIs to treat PMDD. SOURCES OF INFORMATION: Published literature. CONCLUSIONS: Advanced practice nurses need to remain well informed about premenstrual conditions and emerging evidence-based treatment alternatives. In particular, they need to remember that the FDA has approved fluoxetine for the treatment of a very small subset of women with premenstrual complaints, among whom treatment efficacy is limited. PMID- 12132632 TI - The nature and sources of hope: perspectives of family caregivers of people with serious mental illness. AB - PURPOSE: To explore the importance and meaning of hope for family members of people with mental illness. METHODS: Focused in-depth interviews with 16 family members in Queensland and Tasmania, Australia. FINDINGS: The data confirm the argument that hopefulness appears to be central to a family's coping with the impact of mental illness. Their definitions of hope, descriptions of what they hoped for, and the sources of their hope reflect issues of future orientation, positive expectation, and realism. Families drew their hopefulness from both formal and informal supports, from within and without. CONCLUSIONS: Health professionals need to be respectful of family hopes and aware of the role of hope and time in the process of grief and acceptance. Nurses should be mindful of their capacity to sustain or diminish the hopes of family members. PMID- 12132633 TI - Messy purse girls: adult females and ADHD. PMID- 12132634 TI - Monitoring contraceptive continuation: links to fertility outcomes and quality of care. AB - This study examines the fertility consequences of contraceptive discontinuation, describes cross-national variation in continuation rates, and assesses the usefulness of the contraceptive discontinuation rate as a summary outcome indicator of quality of care. In the 15 countries included in this analysis, the total fertility rate would be between 28 and 64 percent lower if the births following discontinuations that were not the result of a desire to become pregnant had not occurred. The all-method discontinuation rate for quality related reasons emerges as the most likely candidate for a summary measure of quality of care. Within a year of starting use of a method, between 7 and 27 percent of women cease to practice contraception for reasons related to the quality of the service environment. The results imply that as fertility declines, family planning programs would profit from a shift in emphasis from providing methods to new clients toward providing services to reduce discontinuation rates. PMID- 12132635 TI - The impact of the Navrongo Project on contraceptive knowledge and use, reproductive preferences, and fertility. AB - The Navrongo Community Health and Family Planning Project is a quasi-experimental study designed to test the hypothesis that introducing health and family planning services in a traditional African societal setting will introduce reproductive change. This article presents the impact of the initial three years of project exposure on contraceptive knowledge, awareness of supply sources, reproductive preferences, contraceptive use, and fertility. Findings show that knowledge of methods and supply sources increased as a result of exposure to project activities and that deployment of nurses to communities was associated with the emergence of preferences to limit childbearing. Fertility impact is evident in all treatment cells, most prominently in areas where nurse-outreach activities are combined with strategies for involving traditional leaders and male volunteers in promoting the program. In this combined cell, the initial three years of project exposure reduced the total fertility rate by one birth, comprising a 15 percent fertility decline relative to fertility levels in comparison communities. PMID- 12132636 TI - Changing patterns of Mongolian fertility at a time of social and economic transition. AB - In 1989, after a long period of socialist rule, Mongolia initiated a democratization process of its political system together with a transition toward a market economy. This study examines how changes in socioeconomic conditions in Mongolia have affected fertility patterns in recent times. It also provides an outline of changes that have taken place in terms of pronatalist policies. Data are drawn from the Reproductive Health Survey of Mongolia conducted in 1998. Among the older cohort, the relationship between economic activity and fertility is inverse but weak, whereas among the younger cohort, the economic downturn has had a strongly depressing effect on fertility. Important effects of micro-level variables, including education and housing, are also noted. The findings suggest that the fertility decline observed for the older cohorts is part of the first demographic transition, in which the collapse of pronatalist policies was influential. PMID- 12132638 TI - Spousal communication and family planning behavior in Navrongo: a longitudinal assessment. AB - The poor performance of most family planning programs in the 1980s, especially in sub-Saharan Africa, generated concern among researchers and led to a quest for explanations. In most countries, the alienation of men from participation in these programs was subsequently identified as one of the major causes, a finding that led researchers to redirect their attention to couples instead of individuals as the focus of such programs. Lack of spousal communication about family planning was identified as one reason for the low level of contraceptive use among women. Subsequent research has persistently demonstrated a positive relationship between spousal communication and contraceptive use. Most prior studies on this topic have been based on cross-sectional data, so that whether the identified relationships are causal remains unclear. Does communication, in fact, predict contraceptive use, or does the use of contraceptives generate communication among couples? This study addresses the question of causality by using longitudinal data from the Navrongo Health Research Centre panel survey. Results from both cross-sectional and longitudinal analysis demonstrate that spousal communication does, indeed, predict contraceptive behavior, even when other factors are controlled. PMID- 12132637 TI - The impact of female genital cutting on first delivery in southwest Nigeria. AB - To date, data linking obstetric morbidity to female genital cutting in populations with less severe types of cutting have been limited to case reports and speculation. In this cross-sectional study, 1,107 women at three hospitals in Edo State, Nigeria, reported on their first-delivery experiences. Fifty-six percent of the sample had undergone genital cutting. Although univariate analyses suggest that genital cutting is associated with delivery complications and procedures, multivariate analyses controlling for sociodemographic factors and delivery setting show no difference between cut and noncut women's likelihood of reporting first-delivery complications or procedures. Whereas a clinical association between genital cutting and obstetric morbidity may occur in populations that have undergone more severe forms of cutting, in this setting, apparent associations between cutting and obstetric morbidity appear to reflect confounding by social class and by the conditions under which delivery takes place. PMID- 12132639 TI - Reducing unplanned pregnancy and abortion in Zimbabwe through postabortion contraception. AB - In many countries, women treated for complications from spontaneous or unsafely induced abortion lack access to contraceptive services. As a result, many of them soon have a subsequent unplanned pregnancy or a repeat abortion, placing their health at increased risk. This report presents the results of a prospective intervention study on postabortion family planning conducted in the two largest public hospitals in Zimbabwe. Women at Harare Central Hospital, in the capital, received a postabortion family planning intervention, and Mpilo Central Hospital, in Bulawayo, served as the control site. The study cohort was 982 women, 527 of whom were followed for a 12-month period. During the follow-up period, significantly more women used highly effective methods of contraception, significantly fewer unplanned pregnancies occurred, and fewer repeat abortions were performed at the intervention site than at the control site. These results offer compelling evidence that ward-based contraceptive services provided to women treated for incomplete abortion can significantly reduce subsequent unplanned pregnancies. The results also suggest that postabortion family planning services can reduce the incidence of repeat abortion. PMID- 12132640 TI - Guinee 1999: results from the demographic and health survey. PMID- 12132641 TI - Zimbabwe 1998-99: results from the Demographic and Health Survey. PMID- 12132642 TI - Electro-acupuncture stimulation to muscle afferents in anesthetized rats modulates the blood flow to the knee joint through autonomic reflexes and nitric oxide. AB - Recent reports have focused on the mechanisms of the action of electro acupuncture stimulation (EAS) in the regulation of blood flow to different tissues. In the knee joint, blood flow is known to be modulated mainly by sympathetic postganglionic fibers, but recently the release or induction of nitric oxide (NO) synthesis in response to electrical stimulation has also been suggested. Therefore, a direct observation of the microcirculation is needed to further understand the mechanism by which blood flow is regulated by somatic afferent stimulation. In the present study, the effects of EAS to the vastus medialis muscle on systemic hemodynamics and the knee joint microcirculation were observed in vivo using a real-time confocal laser-scanning microscope system (CLMS). Electrical stimulation (5 mA, 0.5 ms, 5 Hz) was applied for 30 min using a pair of acupuncture needles introduced into the vastus medialis muscle. To clarify a plausible involvement of NO in the responses to EAS, the stimulus was applied either in the presence or absence of N(omega)-nitro-L-arginine methyl ester (L-NAME). Stimulation to either the muscle or the skin of the thigh after blockade of neuromuscular transmission was performed to determine the involvement of muscle contraction during EAS treatment. The changes in mean arterial pressure (MAP) and diameter of the arterioles supplying the knee joint were monitored continuously until 60 min poststimulus. Significant and persistent increases in arteriolar diameter by 26 +/- 6% and MAP by 17 +/- 2%, respectively, were observed after EAS to the muscle. Electro-acupuncture to the vastus medialis in the presence of L-NAME produced a strong decrease in diameter of the knee joint arterioles by -38 +/- 14% under the baseline with a simultaneous increase of 35 +/- 5% in MAP. EAS to the skin did not produce changes in arteriolar diameter while a slight increase in MAP by 12 +/- 6% over the baseline occurred after the stimulus. EAS to the muscle after neuromuscular blockade did not produce significant changes in diameter, while an increase in MAP by 24 +/- 8% was still observed, which facts suggest that the muscle contraction is required to produce vasodilatation. These responses suggest that a dynamic balance between the autonomic nervous system and the release of NO is the primary mechanism mediating the EAS effects on knee joint microcirculation. PMID- 12132643 TI - Cardiovascular responses to chemoreflex activation with potassium cyanide or hypoxic hypoxia in awake rats. AB - Although intravenous (iv) injection of potassium cyanide (KCN) activates the arterial chemoreflex, it has been questioned whether cytotoxic hypoxia reproduces a physiological stimulus such as hypoxic hypoxia (low inspired O2 tension). Thus, the goal of the present study was to compare the cardiovascular responses elicited by intravenous injection of KCN to those caused by hypoxic hypoxia in awake rats before and after bilateral ligature of carotid body arteries. We tested the hypothesis that hypoxic hypoxia activates the cardiovascular chemoreflex just as KCN does, causing an increase in arterial pressure and bradycardia. Intact adult Wistar rats received an intravenous injection of KCN (160 microg/kg) and were exposed to hypoxic hypoxia (7-5% O2 breathing) for 10-15 s at random while mean arterial pressure (MAP) and heart rate (HR) were measured. After the experiments, the animals were submitted to bilateral ligature of carotid body arteries or sham operation and the protocol was repeated on the subsequent day. Before surgery, all rats showed an abrupt rise in arterial pressure accompanied by a marked bradycardia in response to KCN or hypoxic hypoxia, with a very similar pattern. After surgery, these responses persisted only in the sham-operated group and were totally abolished in the ligature group. In conclusion, our data show that KCN is an appropriate stimulus to activate arterial chemoreflex because its cardiovascular responses are comparable to those induced by hypoxic hypoxia. Thus, the use of KCN as a tool to evaluate different aspects of the complex pattern of cardiovascular, respiratory, and behavioural responses to chemoreflex activation seems to be physiologically acceptable. PMID- 12132644 TI - Predicting motion sickness during parabolic flight. AB - BACKGROUND: There are large individual differences in susceptibility to motion sickness. Attempts to predict who will become motion sick have had limited success. In the present study, we examined gender differences in resting levels of salivary amylase and total protein, cardiac interbeat intervals (R-R intervals), and a sympathovagal index and evaluated their potential to correctly classify individuals into two motion sickness severity groups. METHODS: Sixteen subjects (10 men and 6 women) flew four sets of 10 parabolas aboard NASA's KC-135 aircraft. Saliva samples for amylase and total protein were collected preflight on the day of the flight and motion sickness symptoms were recorded during each parabola. Cardiovascular parameters were collected in the supine position 1-5 days before the flight. RESULTS: There were no significant gender differences in sickness severity or any of the other variables mentioned above. Discriminant analysis using salivary amylase, R-R intervals and the sympathovagal index produced a significant Wilks' lambda coefficient of 0.36, p=0.006. The analysis correctly classified 87% of the subjects into the none-mild sickness or the moderate-severe sickness group. CONCLUSIONS: The linear combination of resting levels of salivary amylase, high-frequency R-R interval levels, and a sympathovagal index may be useful in predicting motion sickness severity. PMID- 12132645 TI - Effect of whole-body and local heating on cutaneous vasoconstrictor responses in humans. AB - Animal studies suggest that alpha-adrenergic-mediated vasoconstriction is compromised during whole-body heating. The purpose of this study was to identify whether whole-body heating and/or local surface heating reduce cutaneous alpha adrenergic vasoconstrictor responsiveness in human skin. Protocol I: Six subjects were exposed to neutral skin temperature (i.e., 34 degrees C), whole-body heating, and local heating of forearm skin to increase skin blood flow to the same relative magnitude as that observed during whole-body heating. Protocol II: In eight subjects forearm skin was locally heated to 34, 37, 40, and 42 degrees C. During both protocols, alpha-adrenergic vasoconstrictor responsiveness was assessed by local delivery of norepinephrine (NE) via intradermal microdialysis. Skin blood flow was continuously monitored over each microdialysis membrane via laser-Doppler flowmetry. In protocol I, whole-body and local heating caused similar increases in cutaneous vascular conductance (CVC). The EC50 (log NE dose) of the dose-response curves for both whole body (-4.2 +/- 0.1 M) and local heating (-4.7 +/- 0.4 M) were significantly greater (i.e., high dose required to cause 50% reduction in CVC) relative to neutral skin temperature (- 5.6 +/- 0.0 M; P<0.05 for both). In both local and whole-body heated conditions CVC did not return to pre-heating values even at the highest dose of NE. In protocol II, calculated EC50 for 34, 37, 40, and 42 degrees C local heating was - 5.5 +/- 0.4, -4.6 +/- 0.3, -4.5 +/- 0.3, - 4.2 +/- 0.4 M, respectively. Statistical analyses revealed that the EC50 for 37,40 and 42 degrees C were significantly greater than the EC50 for 34 degrees C. These results indicate that even during administration of high concentrations of NE, alpha-adrenergic vasoconstriction does not fully compensate for local heating and whole-body heating induced vasodilatation in young, healthy subjects. Moreover, these data suggest that elevated local temperatures, above 37 degrees C, and whole-body heating similarly attenuate cutaneous alpha-adrenergic vasoconstriction responsiveness. PMID- 12132646 TI - Pupillary and cardiovascular responses to a video movie in senior human subjects. AB - The effects of watching video movies on autonomic functions were estimated by measuring changes in pupillary and cardiovascular parameters in 10 senior subjects. The subjects looked at a series of video images (with accompanied sounds) taken during the execution of motor vehicles. The images were rear projected on a large screen for 15 min. Pupil diameter and parameters of the light reflex were measured by an infrared pupillometer before and after the video presentation. Their electrocardiograms (ECG) and blood pressure were measured continuously. Subjects were divided into two groups depending on their values of blood pressure and fasting plasma glucose level. Subjects in Group A had blood pressures of less than 140 mm Hg and a fasting plasma glucose level of less than 7 mmol/dl (normal group). Other subjects were included in Group B (mild hypertension or diabetes mellitus group). While changes in pupillary light reflex after video viewing were minimal in the members of Group A, amplitudes of the pupillary reflex in the members of Group B varied over a significantly wide range. By the spectral analysis of cardiovascular rhythm, %LF and %HF components of blood pressure rhythm were significantly different between the two groups before video viewing. However, the ratios of frequency components before and after video viewing were not significantly different between the two groups. Our findings suggest that pupillary light reflex was less precisely controlled in subjects with mild autonomic dysfunction after prolonged audiovisual stimulation. PMID- 12132647 TI - Origins and projections of nerve fibres in rat pyloric sphincter. AB - Nerve fibres play an important role in the regulation of gastric emptying. The aims of this study were to clarify the distribution, projections and origin of neuronal type nitric oxide synthase (NOS)-, tyrosine hydroxylase (TH)-, vesicular acetylcholine transporter (VAchT)- and peptide-containing nerve fibres of the rat pyloric sphincter. Extrinsic and local denervations of the sphincter were performed in order to reveal the origin and projections of the various nerve fibre populations. Pylorus from control and denervated animals were processed for the immunocytochemical demonstration of cholecystokinin (CCK), enkephalin, gastrin-releasing peptide (GRP), somatostatin, calcitonin gene-related peptide (CGRP), neuropeptide Y (NPY), pituitary adenylate cyclase-activating peptide (PACAP), substance P (SP), vasoactive intestinal peptide (VIP), galanin, NOS, VAchT and TH. VAchT, TH, nNOS, and all of the peptides investigated were found in nerve fibres innervating the pyloric sphincter, and coexistence of several putative neurotransmitters were revealed. Extrinsic denervation caused a total loss of NPY/TH-, SP/CGRP- and SP/CGRP/VIP/NOS/PACAP-containing nerve fibres. Local denervation immediately proximal to the sphincter markedly reduced the numbers of VIP/NOS/galanin- and VIP/NOS/galanin/PACAP +/- NPY-containing fibres within the sphincter suggesting an origin of these fibres in myenteric ganglia in the antral region; denervation at the level of the oxyntic-pyloric border had no effect. Local denervation immediately distal to the sphincter caused a marked decrease in VAchT-, SP/enkephalin-, enkephalin-, somatostatin-, CCK- and GRP containing fibres within the sphincter suggesting that these emanate from the duodenum. The latter procedure also reduced the number of SP/CGRP-containing fibres of extrinsic origin within the pyloric sphincter. PMID- 12132648 TI - Comparison of capsaicin-evoked calcium transients between rat nodose and jugular ganglion neurons. AB - The objectives of this study were to describe the size distribution of capsaicin sensitive neurons in nodose and jugular ganglia and to determine whether there is a difference in capsaicin sensitivity between these two types of ganglia. Functional identification was made by measurement of the capsaicin-evoked calcium (Ca2+) transients in cultured vagal sensory neurons of young adult Sprague-Dawley rats using the Fura-2-based ratiometric imaging technique. In the first study series, cells on the second day of culture were perfused with capsaicin solution (10(-7) M) for 15 s, and the Ca2+ transients were continuously recorded before, during, and after the capsaicin challenge. Out of 603 viable neurons, 57.5% were capsaicin-sensitive; the percentages of capsaicin-sensitive cells in the nodose and jugular ganglia were 59.8% and 55.4%, respectively. Capsaicin sensitivity predominated in the small- and medium-sized neurons; the capsaicin-sensitive cells generally had a diameter less than 35 microm in both types of ganglia. Although the results did not indicate any differences in the size distribution of capsaicin-sensitive neurons between the two ganglia, results of our second study series showed that a near-maximal concentration of capsaicin (3 x 10(-6) M) evoked a significantly greater peak Ca2+ transient in jugular neurons (382.5 +/- 85.5 nM) than in nodose neurons (134.3 +/- 17.5 nM). In summary, our results showed that an increase in cell diameter was accompanied by a decreasing trend in percentage of capsaicin-sensitive neurons in both vagal ganglia. Capsaicin at high concentration evoked a greater peak Ca2+ transient in jugular ganglion neurons, despite no difference in the responses to KCl between these two types of ganglion neurons. PMID- 12132649 TI - ANP potentiates nonarterial baroreflex bradycardia: evidence from sinoaortic denervation in rats. AB - Previous findings indicate that atrial natriuretic peptide (ANP) enhances the reflex bradycardia arising from stimulation of cardiac mechanoreceptors and chemoreceptors, but not that from arterial baroreceptors. The present study tests this proposal by examining the effect of ANP on these reflexes in six chronically sinoaortic-denervated (SAD), and eight sham-operated (sham), conscious rats. Arterial baroreceptor-heart rate (HR) reflex function was examined by constructing full-range steady-state blood pressure (BP)-HR curves using alternating doses of pressor (methoxamine, 2-100 microg/kg) and depressor (nitroprusside, 1-50 microg/kg) agents. Nonarterial baroreceptor reflex function was assessed by the 'ramp' bradycardic response to the rapid BP rise after i.v. methoxamine (100 microg/kg bolus dose). The cardiopulmonary chemoreflex was evoked by i.v. injections of serotonin (1-20 microg/kg). These three tests were performed on each rat during infusions, in random order, of rat ANP (150 ng/kg/min i.v.) and saline vehicle. The ability of ANP to significantly enhance ramp reflex bradycardia was not diminished in SAD compared with sham rats (+54 +/ 12% vs. +42 +/- 15%, respectively). ANP also significantly enhanced cardiopulmonary chemoreflex bradycardia in both groups (+60 +/- 15% in SAD, +40 +/- 8% in sham). Neither the normal steady-state BP-HR response in sham rats nor the small residual response in SAD rats was enhanced by ANP (-1 +/- 7% in sham, 11 +/- 8% in SAD). We conclude that ANP enhances reflex bradycardias of nonarterial, probably cardiac mechanoreceptor, origin. PMID- 12132650 TI - Effects of L-carnosine on renal sympathetic nerve activity and DOCA-salt hypertension in rats. AB - The effects of L-carnosine (beta-alanyl-L-histidine) on the neural activity of the renal sympathetic nerve and on DOCA-salt hypertension in rats were examined. Intravenous injection of 1 microg L-carnosine inhibited renal sympathetic nerve activity in urethane-anesthetized animals, and a diet containing 0.0001% or 0.001% L-carnosine decreased blood pressure elevation in DOCA-salt hypertensive rats. Since L-carnosine is mainly synthesized in the skeletal muscles of mammals, it is not unreasonable to postulate that L-carnosine is an endogenous factor controlling the blood pressure in a manner possibly antagonistic to the obesity associated hypertensive effect of leptin. PMID- 12132651 TI - Increases in plasma dihydroxyphenylacetic acid levels in decapitated mice after exposure to various stresses. AB - This study investigated changes in plasma levels of the dopamine metabolite dihydroxyphenylacetic acid (DOPAC) in decapitated mice in response to the variable stresses of restraint, restraint and water immersion, and foot shock. DOPAC levels, but not norepinephrine (NE) and epinephrine (EPI) levels, increased upon exposure to these stresses. Plasma DOPAC levels measured using the decapitation method in rats were then compared with those measured using the catheter method. The NE and EPI levels in plasma measured using the decapitation method were much higher than those using the catheter method under basal conditions. In contrast, differences in the levels of DOPAC in plasma were smaller than those of NE and EPI under basal conditions using in both methods; furthermore parallel changes in plasma DOPAC levels occurred during restraint and water immersion stresses. These results indicate that the plasma DOPAC levels measured in mice using the decapitation method were clearly increased by the different stresses. Furthermore, in rats there were correlations between the decapitation and catheter methods for plasma levels of DOPAC. Thus the change in plasma DOPAC levels measured using the decapitation method is a good indicator of stress responses involving sympathoneural activity. PMID- 12132652 TI - Direct reaction between shikonin and thiols induces apoptosis in HL60 cells. AB - Shikonin (beta-alkannin), a naphthoquinone compound, was found to induce apoptotic features such as chromatin condensation, DNA fragmentation, and activation of caspase 3 in HL60 cells. The mechanism was examined in terms of oxidative stress in the cells. Exposure of the cells to shikonin greatly reduced the total thiols, protein thiols, and glutathione levels, however, lipid peroxide levels were enhanced. The depletion of thiol levels in the cells was thus thought to induce lipid peroxidation and DNA fragmentation. An electron spin resonance study revealed that shikonin reacts directly with glutathione and other oxidative stress-relevant compounds in the lysate of HL60 cells. Pretreatment of such cells with N-acetylcysteine before shikonin treatment completely inhibited the DNA fragmentation. From these results, it was proposed that the chemical reaction between shikonin and cellular thiols such as glutathione and protein thiols induces apoptosis in HL60 cells. PMID- 12132653 TI - A novel protein with alkaline phosphatase and protease inhibitor activities in Streptomyces hiroshimensis. AB - A novel alkaline phosphatase (S-ALP) was found in the culture filtrate of Streptomyces hiroshimensis IFO 12785. Purification was achieved on Sephadex G-75 column, palmitoylated gauze column, and Superdex 75 HR column chromatographies. The molecular weight of S-ALP was estimated to be 14200 by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The isoelectric point is 9.2. S ALP had maximum enzyme activity at pH 9.5. S-ALP efficiently catalyzed both p nitrophenyl phosphate and p-nitrophenyl phosphorylcholine substrates, particularly the latter. The N-terminal amino acid sequence (25 residues) of S ALP was 60 to 72% identical to that of Streptomyces subtilisin inhibitor-like proteins. S-ALP exhibited trypsin inhibition in addition to a strong inhibition of subtilisin. PMID- 12132654 TI - The role of protein kinase C in the transient association of p57, a coronin family actin-binding protein, with phagosomes. AB - Phagocytosis of opsonized zymosan (OpZ) particles by differentiated cells of the human leukemic cell line HL-60 induced transient periphagosomal association of p57, a coronin family actin-binding protein, and F-actin with dissociation from the phagosomes after ingestion was completed. Coincident with OpZ ingestion, p57 phosphorylation increased transiently and peaked with its dissociation from phagosomes. Since p57 contains several putative sites for protein kinase C (PKC) phosphorylation, we examined the effect of PKC on p57 phosphorylation and association with the phagosome. Purified p57 was phosphorylated in vitro by PKC isoforms alpha and delta, and PMA, an activator of PKC, induced p57 phosphorylation in HL-60 cells. Furthermore, chelerythrine, a specific PKC inhibitor, blocked p57 phosphorylation and the dissociation of p57 and F-actin from phagosomes, whereas wortmannin, genistein, and H-89 did not. Chelerythrine also inhibited the translocation of LAMP-1, a marker protein of lysosomes, to the OpZ-containing phagosomes, indicating that PKC-mediated phosphorylation is required for phagosome-lysosome fusion. Taken together, these data suggest that PKC-mediated phosphorylation of p57 triggers its dissociation from phagosomes, an event that may be necessary for the fusion of phagosomes with lysosomes. PMID- 12132655 TI - Metabolism of 2-methyl analogs of 1alpha,25-dihydroxyvitamin D3 in rat osteosarcoma cells (UMR 106). AB - Several novel A-ring modified analogs of 1alpha,25-dihydroxyvitamin D3 [1alpha,25(OH)2D3] have been synthesized in order to investigate the structure function relationships of 1alpha,25(OH)2D3. We synthesized A-ring modified analogs which contain a methyl group on C-2 of the A-ring. There are eight 2 methyl diastereomers, which differ in the stereochemistry of the methyl group on C-2 and the hydroxyl groups on C-1 and C-3. Further our biological activity studies of the 2-methyl diastereomers indicated that the potency of each analog is highly dependent on the stereochemistry of the A-ring substituents [Konno et al., Biorg. Med. Chem. Letts. 8(2), 151-156 (1998); Nakagawa et al., Biochem. Pharmacol. 60(12), 1937-1947 (2000)]. For example, the VDR binding affinities exhibited by the 1alpha-isomers are significantly higher than those exhibited by the 1beta-isomers. Furthermore, out of all the 1alpha-isomers, the 2alpha-methyl isomers, when compared to the corresponding 2beta-methyl isomers, showed much higher potency in inducing cell differentiation of HL-60 cells, but failed to stimulate apoptosis. In contrast the 2beta-methyl isomers strongly stimulated apoptosis. At present it is unknown how the addition of the 2-methyl modification to the hormone, 1alpha,25(OH)2D3 alters its metabolism in target tissues. Previously, we reported that 1alpha,25(OH)2D3 is metabolized in rat osteosarcoma (UMR 106) cells via both the C-24 oxidation and the C-3 epimerization pathways. Therefore, we studied the metabolism of the four 1alpha,2-methyl diastereomers in UMR 106 cells. Our results indicated that in UMR 106 cells, all four diastereomers were metabolized into several polar metabolites via the C-24 oxidation pathway. Thus, the presence of the 2-methyl group on the A-ring did not inhibit the metabolism of the analogs via the C-24 oxidation pathway. However, it is significant to note that the 2-methyl group prevented the metabolism of the analogs via the C-3 epimerization pathway. In summary, we report that the 2 methyl group interferes with the action of the enzyme(s) involved in C-3 epimerization, but not with the enzyme 1alpha,25(OH)2D3-24-hydroxylase, which is responsible for C-24 oxidation pathway. PMID- 12132656 TI - DJ-1, a target protein for an endocrine disrupter, participates in the fertilization in mice. AB - DJ-1 was first identified as an activated ras-dependent oncogene product and was later also found to be an infertility-related protein affected by sperm toxicants such as ornidazole (OR) and epichlorohydrin. These findings suggest that DJ-1 has functions in both somatic cells and sperm. In this study, to determine the relationship between DJ-1 and an endocrine disrupter and to determine the functions of DJ-1 in sperm, in vitro fertilization experiments were carried out using eggs and sperm extracted from mice that had or had not been treated with OR. We found that the amount of DJ-1 in sperm and the efficiency of fertilization decreased with the increasing dose of OR to which the mice were exposed. The addition of an anti-mouse DJ-1 serum to sperm solution before the in vitro fertilization reaction with eggs resulted in a decrease in the efficiency of fertilization to about one-third of that when pre-immune serum was added to sperm solution, indicating that DJ-1 participates in the fertilization. PMID- 12132657 TI - Effects of angiotensin II type 1 receptor antagonist, YM358, on cardiac hypertrophy and dysfunction after myocardial infarction in rats. AB - This study was undertaken to investigate the effects of an angiotensin II type 1 receptor antagonist, YM358 (2,7-diethyl-5-[[2'(1H-tetrazol-5-yl)biphenyl-4 yl]methyl]-5H-pyrazolo [1,5-b] [1,2,4]triazole potassium salt monohydrate), on cardiac hypertrophy and dysfunction in rats with heart failure after myocardial infarction (MI). One day after the coronary ligation, rats were randomized, and administered YM358 or vehicle for 2, 4 or 8 weeks. In MI rats, mean blood pressure, left ventricular (LV) systolic pressure, and the first derivative of LV pressure significantly decreased, and LV end-diastolic pressure (LVEDP) markedly increased after 2 to 8 week treatment of YM358. From 2 weeks after the ligation, ratios of cardiac weight and lung weight to body weight (BW) significantly increased, which indicated the progression of cardiac hypertrophy and lung congestion in MI rats. Two weeks after the ligation, YM358 did not improve LV function, although it decreased the elevated LVEDP and cardiac weights/BW ratios 8 weeks after the ligation. These results indicated that long-term treatment with YM358 improves the reduced cardiac function and reduces cardiac hypertrophy after MI, and may be useful for the treatment of congestive heart failure. PMID- 12132658 TI - Prevention of growth and metastasis of murine melanoma through enhanced natural killer cytotoxicity by fatty acid-conjugate of protopanaxatriol. AB - Ginsenosides, the glycosides of Panax ginseng, are metabolized (deglycosylated) by intestinal bacteria after oral administration. 20(S)-Protopanaxatriol (M4) is the main bacterial metabolite of protopanaxatriol-type ginsenosides and mediates their antitumor effects. To clarify the mechanism of the M4-mediated antitumor effect, the antitumor activity and metabolism of M4 was examined, using the C57BL/6 mice implanted with B16-BL6 melanoma. The chronic oral administration of M4 inhibited the growth of B16-BL6 melanoma at the implanted site. Analyses using TLC, HPLC, MS and NMR suggest that orally administered M4 was absorbed from the small intestine into the mesenteric lymphatics followed by the rapid esterification of M4 with fatty acids and its accumulation in the tissues including the liver and lung. The administration of M4 prior to the intravenous injection of B16-BL6 cells abrogated the enhanced lung metastasis in the mice pretreated with 2-chloroadenosine more effectively than in those pretreated with anti-asialo GM1. The esterified M4 (EM4) did not directly affect tumor growth in vitro, whereas it stimulated splenic NK cells to become cytotoxic to tumor cells. These results indicate that the antitumor activity of M4 is based on the NK cell mediated tumor lysis enhanced by EM4. PMID- 12132659 TI - Exhalation behavior of four organic substrates and water absorbed by human skin. AB - The simultaneous measurement of several volatile organic compounds and water released from the human skin can be achieved successfully by using a modified gas chromatographic system. After the thumb of each subject was dipped in aqueous solution containing acetone, diethyl ether, ethanol, and toluene, it was dried in the air. Then the thumb attached to the sampling probe for measuring the released gases. It is found that 90% of all these chemical substrates were desorbed after 20 min. The initial exhalation rate factor for each chemical substrate was determined in every subject. Correlation factors of the linear relationships between the initial exhalation rate for hydrophilic substrates (acetone and ethanol) and the total amount of water (TAW) released from the skin were 0.94 and 0.92, respectively. However, the rate of hydrophobic toluene was not dependent on the TAW. Therefore, the exhalation rate of substrates is greatly influenced by both their hydrophilicity and TAW. Additionally, an interesting personal specific character among the 6 subjects was observed on plotting the exhalation rate of organic substrates and water during the elapsed time. With the released water mostly due to insensible perspiration, the exhalation rate of all simultaneous organic substrates decreased monotonically over the elapsed time. On the contrary, when subjects sweated emotionally, the exhalation rate of organic substrates showed some variation, namely a higher of exhalation rate compared to the case of mostly due to insensible perspiration. Therefore, emotionally-induced sweating can enhance the release of organic substrates. PMID- 12132660 TI - Pharmacological properties of traditional medicines (XXVII). Interaction between Ephedra Herb and Gypsum under hyperthermal conditions in rats. AB - There are many important considerations in the interactions among the herbal constituents in a prescription of traditional Chinese medicine (TCM). Ephedra Herb [Chinese characters: see text] (Eph) is described a warm and acrid agent in TCM. The combination of Eph and Gypsum [Chinese characters: see text] (Eph-Gyp) shows specific actions in patients with different body temperatures. Previous reports suggested that Gypsum prevents the thermogenesis effect induced by ephedrine at an ambient temperature of 22 degrees C. In this investigation, the properties of Eph-Gyp in hyperthermal rats were studied in detail. It was shown that Gypsum Extract (GyE) enhanced the thermogenesis of Eph in hyperthermal rats, although not in normal rats. The results support not only the opposite actions of Eph-GyE but also the clinical differences in the symptomatic patterns of body temperature for Makyo-Kanseki-To [Chinese characters: see text] and Dai-Seiryu-To [Chinese characters: see text]. PMID- 12132661 TI - Antiproliferative constituents in plants 10. Flavones from the leaves of Lantana montevidensis Briq. and consideration of structure-activity relationship. AB - The flavonoid fraction from the leaves of Lantana montevidensis Briq. (Verbenaceae) showed antiproliferative activity against human gastric adenocarcinoma (MK-1, GI50: 12 microg/ml), human uterus carcinoma (HeLa, 5 microg/ml), and murine melanoma (B16F10, 5 microg/ml) cells in vitro. Bioactivity guided chemical investigation of the fraction has resulted in the isolation of apigenin (10) and ten 5,6,7-oxygenated flavones: cirsilineol (1), eupatorin (2), 5,4'-dihydroxy-6,7,3',5'-tetramethoxyflavone (3), 5,6-dihydroxy-7,3',4' trimethoxyflavone (4), 5,6,4'-trihydroxy-7,3',5'-trimethoxyflavone (5), 5,6,3' trihydroxy-7,4'-dimethoxyflavone (6), 5,3',4'-trihydroxy-6,7,5'-trimethoxyflavone (7), cirsiliol (8), hispidulin (9), and eupafolin (11). Antiproliferative activity of the isolated flavones, some other related flavones (luteolin, baicalein, 6-hydroxyluteolin, pectolinarigenin, jaceosidin, desmethoxycentaureidin, eupatilin, and chrysin) from other plant materials, and synthetic 6- and 7-methoxyflavones was evaluated, and the structure-activity relationships were examined. PMID- 12132662 TI - Suppressive effect of Shichimotsu-koka-to (Kampo medicine) on pulmonary metastasis of B16 melanoma cells. AB - Shichimotsu-koka-to (SKT) is a Kampo (traditional Japanese herbal) medicine, which is used in Japan to treat hypertension and atherosclerosis. We investigated the inhibitory effect of SKT on experimental pulmonary metastasis of B16 melanoma cells. The intake of SKT at a dose of 430 mg/kg for 6 weeks from 2 weeks before tumor inoculation significantly reduced the number of metastatic surface nodules in the lung and extended the life span. When the duration of SKT intake was examined, survival time was not affected by preintake before B16 melanoma cell inoculation and was slightly extended by postintake after B16 melanoma cell inoculation, although the life span was prolonged by intake throughout the experiment. To address the mechanism underlying the antimetastatic effect of SKT, we studied whether SKT modulated macrophage function, which is involved in killing tumor cells. The intake of SKT for 6 weeks dose dependently increased nitric oxide (NO) production by macrophages following stimulation with lipopolysaccharide. The elevated NO was found to serve as a cytotoxic mediator against B16 melanoma cells in co-culture with macrophages. On the contrary, B16 melanoma-conditioned medium reduced NO production by macrophages. However, SKT treatment reversed the reduction in NO production by the conditioned medium significantly. These findings may suggest that macrophage function-modulating activity by SKT appears to underlie its antimetastatic activity, which leads to a decrease in the number of lung metastatic surface nodules and the extension of life span. PMID- 12132664 TI - Comparison of the efficiency and safety of non-viral vector-mediated gene transfer into a wide range of human cells. AB - Non-viral gene transfer into a wide range of human cells was examined in order to clarify the factors that affect the efficiency and safety of non-viral vectors and to optimize the conditions so that high efficiency and low toxicity could be achieved. Six non-viral vectors (Lipofectin, LipofectAMINE PLUS, SuperFect, Effectene, DMRIE-C and DOTAP) were used to transfect a mammalian expression plasmid pCMVbeta into 16 types of human primary cells and cultured cell lines. Transfection efficiency was quantified using a galactosidase assay. Cytotoxic effects were measured by lactate dehydrogenase (LDH) assay and WST-8 assay. In serum-free conditions, LipofectAMINE PLUS, Effectene and SuperFect, on average, transfected DNA more successfully than Lipofectin, DMRIE-C, and DOTAP, although the levels of gene expression with these vectors varied remarkably in different cells. The most effective vector also differed depending on the cell type. Serum was found to inhibit gene transfer and reduce the cytotoxicity of all of these vectors except Effectene. The efficiency and toxicity of the non-viral vectors used depended on the type of vector, the DNA/vector ratio, the type of cell, and the presence of serum. These results provided useful information for the optimization of transfer conditions of these non-viral vectors. PMID- 12132663 TI - Studies on intestinal absorption of sulpiride (1): carrier-mediated uptake of sulpiride in the human intestinal cell line Caco-2. AB - We investigated whether the uptake of a specific antipsychotic agent, sulpiride, in Caco-2 cells is mediated by a carrier-mediated system. Caco-2 cell monolayers were cultured in plastic culture dishes and uptake and efflux studies were conducted. The determination of sulpiride was performed by HPLC. At 37 degrees C, sulpiride uptake in pH 6.0 was twice as much as in pH 7.4. At 4 degrees C, however, no significant difference was observed between pH 6.0 and 7.4. The uptake at 4 degrees C was markedly lower than that obtained at 37 degrees C. The subtraction of the uptake at 4 degrees C from the uptake at 37 degrees C indicated a saturable process, and the result of the Eadie-Hofstee plot analysis indicated that the uptake consists of two or more saturable components. The uptake was significantly inhibited by uncoupler, protonophore, amino acid modifying agent and proteinase. Sulpiride efflux was temperature-dependent and was significantly inhibited by uncoupler and amino acid modifying agent. These findings indicate that sulpiride uptake and efflux in Caco-2 cells are carrier mediated. Furthermore, the uptake was significantly decreased by some substrates and inhibitors of peptide transporter, PEPT1, and organic cation transporters, OCTN1 and OCTN2, and was significantly increased by preloading with them. The uptake was also significantly increased by a typical substrate of P-glycoprotein. From these findings, we presumed that peptide transporter PEPT1 and organic cation transporters OCTN1 and OCTN2 are involved with this uptake. P-glycoprotein may also contribute to the efflux of sulpiride. PMID- 12132665 TI - In vitro antioxidant potentials of traditional Chinese medicine, Shengmai San and their relation to in vivo protective effect on cerebral oxidative damage in rats. AB - The preventive effects of Shengmai San (SMS), a traditional Chinese herbal medicine (TCM), was studied on cerebral ischemia-reperfusion injury in rats as a model of antioxidant-based composite therapy. Two biochemical indicators of oxidative damage, thiobarbituric acid reactive substance (TBARS) formation and glutathione peroxidase (GPX) loss were measured in the brain after forebrain ischemia-reperfusion treatment and both were inhibited in all rats administered SMS (15 g original herbs/kg) 2 h before the ischemia-reperfusion. Histochemical study of the brain slice using TTC staining revealed that the SMS effectively reduced infarct area caused by the cerebral ischemia-reperfusion. The antioxidant potentials of SMS preparations were determined in vitro by five different assay methods and were related to the in vivo effectiveness of SMS in protection against brain damage. Inhibitory effect on TBARS formation in vivo showed better correlation with superoxide radical scavenging and DPPH quenching activity in vitro rather than with the other in vitro antioxidant indicators. On the other hand, the in vivo prevention of GPX activity loss showed better correlation with in vitro crocin bleaching inhibition than with the other in vitro antioxidant indicators. It was also suggested that the in vitro TBARS inhibitory activty of SMS is not a good indication to predict the in vivo effectiveness of SMS on inhibition of either TBARS formation or GPX activity loss. PMID- 12132666 TI - Isolation of a novel peptide homing to tumor-derived neovasculature that suppresses tumor growth. AB - Novel peptides homing to angiogenic vessels were recently isolated from a phage displayed random pentade-capeptide library, and peptides having WRP sequence showed tumor growth suppression. In this study, we observed that another novel sequence, PVVLFPLH, suppressed tumor growth in vivo. Through the study of tumor growth suppression by the 5-mer peptides derived from this sequence, we determined the epitope sequence to be LFPLH. LFPLH, but not the shuffled peptide FHLLP, suppressed the migration of vascular endothelial growth factor-stimulated human umbilical vein endothelial cells. Interestingly, growth suppression of LFPLH against the cells as well as tumor cells was not observed in vitro. Therefore LFPLH may function to induce tumor dormancy through inhibition of angiogenesis. PMID- 12132667 TI - Suppression of macrophage function by substances with a molecular weight lower than 3000 Da in B16 melanoma-conditioned medium. AB - In the present study, B16 melanoma cells were found to produce inhibitory and cytotoxic substances with a molecular weight lower than 3000 Da against macrophages in a conditioned medium. The B16 melanoma-conditioned medium suppressed nitric oxide (NO) production only by mouse peritoneal macrophages and the mouse macrophage-like cell line, RAW264.7 cells, but not by rat peritoneal macrophages. In addition, it showed cytotoxicity against mouse peritoneal macrophages and mouse macrophage-like cell lines, RAW264.7 and J774A.1 cells, but not against rat cells (peritoneal macrophages, 3Y1, hepatocytes), human cells (HeLa, KB, MCF-7), or mouse 3T3-L1 cells. The inhibitory activity of NO production was not affected by trypsin treatment or arginine supplementation, but it was abolished by heat treatment at 95 degrees C for 3 min. On the other hand, the cytotoxicity was not influenced by these treatments. Inducible NO synthase induction following lipopolysaccharide stimulation was reduced by treatment of mouse peritoneal macrophages with B16 melanoma-conditioned medium. These results suggest that metastatic B16 melanoma cells produce two distinct substances: to suppress NO production by macrophages and to kill macrophages and macrophage-like cell lines. We propose that these activities may help metastatic B16 melanoma cells to escape a host immunosurveillance system and to metastasize to target organs. PMID- 12132668 TI - Mutational analysis of conserved hydrophobic amino acid residues in the N terminal region of DnaA protein. AB - DnaA is the initiator of chromosomal DNA replication in E. coli. We previously reported that conserved hydrophobic amino acid residues in the N-terminal region of DnaA (I26 and L40) are essential for DNA replication in vivo and in vitro using mutant DnaA proteins (DnaAI26S and DnaAL40S). In this study, we introduced further random mutations to find intragenic suppressors for dnaAI26S or dnaAL40S. By direct DNA sequence, a mutation which causes substitution of the Ser (Ile, in the wild-type DnaA) with Phe (DnaAI26F or DnaAL40F) was found in all of the suppressors. Site-directed mutational analysis showed that DnaAI26L, and DnaAL40I, but not DnaAI26S or DnaAL40S, were active for oriC DNA replication in cells. Furthermore, purified DnaAI26F but not DnaAI26S was active for oriC DNA replication in a crude extract. These results strongly suggest that hydrophobic amino acid residues in these positions of DnaA (I26 and L40) are important for the function of this protein as an initiator of DNA replication both in vivo and in vitro. PMID- 12132669 TI - Synthesis, DNA-binding activity and cytotoxicity of carbamate derivatives of Hoechst 33258 in breast cancer MCF-7 cells. AB - A series of carbamate derivatives of Hoechst 33258 was prepared as potential anticancer agents. These new compounds (1-4) were readily prepared in good yields by addition of chloroethyl, bromoethyl, chloropropyl or 4-(chloromethyl)phenyl isocyanates to Hoechst 33258. Their cytotoxic activity was evaluated on human breast cancer MCF-7. Compounds 1-4 were more cytotoxic than Hoechst 33258. In particular derivative 4, the most active of the series, is up to 3 times more potent than Hoechst 33258. The DNA-binding ability of these compounds was evaluated by an ultrafiltration method using calf thymus DNA. These data show that in broad terms the cytotoxic potency of 1-4 in cultured breast cancer MCF-7 cells increases, in accord with their increases in DNA affinity, as shown by the binding constant values. PMID- 12132670 TI - Antiproliferative constituents in plants 9. Aerial parts of Lippia dulcis and Lippia canescens. AB - The antiproliferative constituents in the MeOH extracts of the aerial parts of Lippia dulcis Trev. and Lippia canescens Kunth (Verbenaceae) were investigated. Activity-guided chemical investigation of the MeOH extracts resulted in the isolation of the three bisabolane-type sesquiterpenes [(+)-hernandulcin (1), (-) epihernandulcin (2), and (+)-anymol (3)] and four phenylethanoid glycosides [acteoside (4), isoacteoside (5), martynoside (6), and a new diacetylmartynoside (7)] from the former, and four phenylethanoid glycosides [acteoside (4), isoacteoside (5), arenarioside (8), and leucosceptoside A (9)] and three flavones [desmethoxycentaureidin (10), eupafolin (11), and 6-hydroxyluteolin (12)] from the latter. Antiproliferative activity of the isolated compounds against murine melanoma (B16F10), human gastric adenocarcinoma (MK-1), and human uterine carcinoma (HeLa) cells was estimated. (+)-Anymol (3), acteoside (4), isoacteoside (5), arenarioside (8), eupafolin (11), and 6-hydroxyluteolin (12) had GI50 values of 10-16 microM against B16F10 cell. Desmethoxycentaureidin (10) and eupafolin (11) showed high inhibitory activity against HeLa cell growth (GI50 9 microM, and 6 microM, respectively). PMID- 12132671 TI - Optimal dose of omeprazole for CYP2C19 extensive metabolizers in anti Helicobacter pylori therapy: pharmacokinetic considerations. AB - In anti-Helicobacter pylori therapy using omeprazole and antimicrobials, the efficacy can be related to the CYP2C19 genotype groups; the eradication rates were 83% in extensive metabolizers and 100% in poor metabolizers. The present study was undertaken to help predict the optimal dosage of omeprazole for extensive metabolizers in this therapy. Seven healthy Japanese subjects, classified based on the CYP2C19 genotype into extensive metabolizers (n=4) and poor metabolizers (n=3), participated in this study. Each subject received a single oral dose of omeprazole 20, 40, and 80 mg, with at least a 1-week washout period between each dose. Plasma concentrations of omeprazole and its two metabolites were monitored for 12 h after each dose of medication. After each dose was administered, the pharmacokinetic profiles of omeprazole and its two metabolites were significantly different between extensive metabolizers and poor metabolizers. The area under the plasma concentration-time curve (AUC) of omeprazole in extensive metabolizers was disproportionally increased 3.2- or 19.2 fold with dose escalation from 20 to 40 or 80 mg omeprazole, respectively. In contrast, the AUC of omeprazole was proportionally increased with the higher dose in poor metabolizers. The AUC of omeprazole after 20 mg administration to poor metabolizers was almost equal to the AUC in extensive metabolizers after 80 mg administration. In anti-H. pylori therapy, the recommended dose of omeprazole for extensive metabolizers is suggested to be a maximum of 80 mg x 2/d based on pharmacokinetic considerations. PMID- 12132672 TI - Kidney- and site-selective delivery of 5-fluorouracil utilizing the absorption on the kidney surface in rats. AB - The present study was undertaken to elucidate the kidney- and site-selective delivery of 5-fluorouracil (5-FU) utilizing the absorption on the kidney surface in rats. An experimental system utilizing a cylindrical diffusion cell attached to the right kidney surface was established. After intravenous administration of 5-FU, the concentration of 5-FU in the right and left kidneys was almost the same and was rapidly eliminated. After right kidney surface application of 5-FU, however, the concentration of 5-FU in the right kidney was significantly higher than in the left kidney and other tissues. The 5-FU concentration in four sites of the right kidney after intravenous administration was almost the same. In contrast, 5-FU was site selectively delivered in the kidney after kidney surface application. The blood concentration of 5-FU was low (<1.7 microg/ml) until 120 min after kidney surface application. The maximum blood concentration of 5-FU after kidney surface application was much lower than after intravenous administration. PMID- 12132674 TI - Lethality of yeasts with low pathogenicity in mice immunocompromised by cyclophosphamide treatment. AB - One strain each of Arxiozyma telluris, Saccharomyces cerevisiae, and S. kluyveri showed lethal activity in cyclophosphamide (CY)-treated mice. Accumulation of these yeast cells in the kidneys and elevation of the levels of cytokines, tumor necrosis factor-alpha, and interleukin-1alpha in the sera were recognized in the CY-treated infected-debilitated mice. PMID- 12132673 TI - Effect of SCG, 1,3-beta-D-glucan from Sparassis crispa on the hematopoietic response in cyclophosphamide induced leukopenic mice. AB - Sparassis crispa Fr. is an edible mushroom recently cultivable in Japan. It contains a remarkably high content of 6-branched 1,3-beta-D-glucan showing antitumor activity. Using ion-exchange chromatography, a purified beta-glucan preparation, SCG, was prepared. In this study, we examined the hematopoietic response by SCG in cyclophosphamide (CY)-induced leukopenic mice. SCG enhanced the hematopoietic response in CY induced leukopenic mice by intraperitoneal routes over a wide range of concentrations. SCG enhanced the hematopoietic response in CY-treated mice by prior or post administration. Analyzing the leukocyte population by flow cytometry, monocytes and granulocytes in the peritoneal cavity, liver, spleen and bone marrow (BM) recovered faster than in the control group. The ratio of natural killer cells and gammadelta T cells in the liver, spleen and peritoneal cavity was also increased. In contrast, CD4+ CD8+ cells in the thymus were temporarily significantly decreased by the administration of SCG. Interleukin-6 (IL-6) production of CY+SCG-treated peritoneal exdated cells (PECs), spleen cells and bone marrow cells (BMCs) were higher than that of the CY-treated group. By in vitro culture of CY-treated PEC and spleen cells, IL-6 production was enhanced by the addition of SCG. These facts suggested the possibility that IL-6 might be a key cytokine for the enhanced hematopoietic response by SCG. PMID- 12132675 TI - The effect of ascorbic acid and ethanol on the level of thiobarbituric acid reactive substances (TBARS) in selected tissues of albino BALB/c mice. AB - The aim of the paper was to study the effect of ascorbic acid (vitamin C) and ethanol on the level of thiobarbituric acid reactive substances (TBARS) in the brain, liver, lungs and blood serum of inbred albino BALB/c mice. Sixty male mice divided into 4 groups were used for the study; the mice in each group were injected intraperitoneally (i.p.): Group 1--control, saline; Group 2--H2O2 and FeSO4; Group 3--ascorbic acid (AA); Group 4--AA and ethanol. The level of TBARS in the investigated tissues of the mice in all of the groups was statistically significant higher than in the control group. The highest content of TBARS in blood serum, brain, liver and lung tissue was observed in mice which were given ascorbic acid and ethanol. PMID- 12132676 TI - Pinguisane-type sesquiterpenes from the South American liverwort Porella recurva (Taylor) Kuhnemann. AB - The chemical composition of a dichloromethane extract of the South American liverwort Porella recurva has been examined. Two new pinguisane-type norsesquiterpenoid natural products were isolated: 6,11-epoxy-15-nor-3,4-dioxo 5,10-pinguisadien-12-acetate and 6,11-epoxy-15-nor-4-oxo-5,10-pinguisadien-12 acetate. In addition two known pinguisane-type sesquiterpenes were also isolated: norpinguisone and norpinguisone methyl ester. All structures were elucidated by means of NMR spectroscopic techniques and mass spectrometry. PMID- 12132677 TI - New spirorotenoids from Tephrosia candida. AB - The ethyl acetate extract of roots of Tephrosia candida afforded three new spirorotenoids belonging to a new class of spirocompounds, named tephrospirolactone, tephrospiroketone I, and tephrospiroketone II. The structures of these compounds were determined mainly based on spectral analysis. The only known spirorotenoid described in the literature is amorphispironone, isolated from Amorpha fruticosa. PMID- 12132678 TI - A new bioactive steroidal saponin from Agave attenuata. AB - A new steroidal saponin was isolated from the leaves of Agave attenuata Salm Dyck. Its structure was established as (3beta,5beta,22alpha,25S)-26-(beta-D glucopyranosyloxy)-22-methoxyfurostan-3-yl O-beta-D-glucopyranosyl-(1-->2)-beta-D glucopyranosyl-(1-->2)-O-[beta-D-glucopyranosyl-(1-->3)]-beta-D-glucopyranosyl-(1 ->4)-beta-D-galactopyranoside. The structural identification was performed using detailed analyses of 1H and 13C NMR spectra including 2D NMR spectroscopic techniques (COSY, HETCOR and COLOC) and chemical conversions. The haemolytic potential of the steroidal saponin was evaluated and the anti-inflammatory activity was performed using the capillary permeability assay. PMID- 12132679 TI - Pyrrolizidine alkaloid profiles of some Senecio species from Egypt. AB - Alkaloid profiles of two Egyptian Senecio species (Senecio aegyptius var. discoideus and S. desfontainei) in addition to a cultivated species (S. cineraria) were studied using capillary GLC and GLC-mass spectrometry with respect to pyrrolizidine alkaloids (PAs). Four alkaloids were identified in S. aegyptius var. discoideus, 8 in S. desfontainei and 13 in S. cineraria. Some of these alkaloids have not been reported from these plants. The alkaloidal pattern of different plant organs (flowers, leaves, stem, root) were also investigated. Senecionine has been found to be a one of the major alkaloid in all studied species, it was isolated and its structure was elucidated by 1H- and 13C-NMR. PMID- 12132680 TI - Chemical composition and biological activity of the essential oils of Senecio aegyptius var. discoideus Boiss. AB - The essential oil of Senecio aegyptius var. discoideus flowers, leaves, stems and roots were isolated by hydrodistillation. Analysis of the oils by capillary GLC and GLC-mass spectrometry were performed and 34 out of 37 compounds were identified. The main component was isolated and characterized as 1,10 epoxyfuranoeremophilane using a combination of GLC, GLC-MS, and NMR analyses. The oils of flowers, leaves and stems were rich in monoterpene hydrocarbons while the root oil mainly contains furanoeremophilanes. Flower and leaf volatile oils showed significant level of antifungal activity against C. albicans, moderate effect against Gram positive bacteria, however, it has weak activity against Gram negative bacteria. The isolated sesquiterpene (1,10-epoxyfuranoeremophilane) exhibited substantial inhibitory activity against Gram negative bacteria. PMID- 12132681 TI - Flavonoids from Ficaria verna Huds. AB - A phytochemical investigation of the flowers and leaves of Ficaria verna Huds. (Ranunculaceae) yielded four additional known flavonoid compounds including: kaempferol 3-O-beta-D-(6"-a-L-rhamnopyranosyl)-glucopyranoside (nicotiflorin), apigenin 8-C-beta-D-glucopyranoside (vitexin), luteolin 8-C-beta-D glucopyranoside (orientin) and apigenin 8-C-beta-D-(2"-O-beta-D-glucopyranosyl) glucopyranoside (flavosativaside). The characterisation of these compounds was achieved by various chromatographic and spectroscopic methods (UV, 1H NMR, 13C NMR and MS). PMID- 12132682 TI - On the occurrence of exudate flavonoids in the borage family (Boraginaceae). AB - Externally accumulated flavonoid aglycones have been found for the first time in Nonea species. They exhibit only flavones, one of them being the rare tricetin-4' methyl ether. Within the subfamily Boraginoideae, exudate flavonoids appear to be a rare character. PMID- 12132683 TI - Gas chromatographic-mass spectrometry study of the essential oils of Pimenta racemosa var. terebinthina and P. racemosa var grisea. AB - The essential oil of the leaves of P. racemosa var. terebinthina and P. racemosa var. grisea were examined by GC and GC/MS. The major constituents were alpha terpineol acetate (27%), alpha-terpineol (20%) and 4-methoxy eugenol (12.6%) for Pracemosa var. terebinthina and 4-methoxy-isoeugenol (75.2%) and 4-methoxy eugenol (4.5%) for P. racemosa var. grisea. PMID- 12132684 TI - Extracellular polysaccharides of Penicillium vermiculatum. AB - Extracellular Polysaccharide, Isolation, Characterization Extracellular polysaccharide mixture that forms a viscous mucilate in the fermentation medium during industrial cultivation of Penicillium vermiculatum was isolated by ethanol precipitation and its structural characteristics were investigated by a combination of physicochemical methods. The mixture contained two structurally different polysaccharides similar to those previously described for some fungal species. This is the first report of the fully structurally characterized extracellular polysaccharides of Penicillium vermiculatum. PMID- 12132685 TI - Aspterric acid and 6-hydroxymellein, inhibitors of pollen development in Arabidopsis thaliana, produced by Aspergillus terreus. AB - Aspterric Acid, 6-Hydroxymellein, Arabidopsis thaliana, Aspergillus terreus Aspterric acid (1) and 6-hydroxymellein (2), inhibitors of pollen development in Arabidopsis thaliana, have been isolated from the fungus Aspergillus terreus. 1 and 2 inhibited the pollen development at concentrations of 38 and 52 microM, respectively. The microscopic examination of pollen development suggested that the inhibition by the treatment with 1 caused at meiosis and the inhibition by the treatment with 2 caused at microspore stage. 1 and 2 could be useful agents for the molecular investigation of anther and pollen development in higher plants. PMID- 12132686 TI - Trichosetin, a novel tetramic acid antibiotic produced in dual culture of Trichoderma harzianum and Catharanthus roseus Callus. AB - The dual culture of Trichoderma harzianum and Catharanthus roseus callus produced an antimicrobial compound with a remarkable activity against the Gram-positive bacteria Staphylococcus aureus and Bacillus subtilis. Structural elucidation revealed that this compound, which we have named trichosetin, is a novel tetramic acid (2,4-pyrrolidinedione) antibiotic and a homolog of the fungal metabolite equisetin. This compound however, was not produced in the individual culture of T. harzianum or C. roseus callus. PMID- 12132687 TI - Mycosporine-like amino acids in Antarctic sea ice algae, and their response to UVB radiation. AB - Mycosporine like amino acids (MAAs) were detected in low concentration in sea ice algae growing in situ at Cape Evans, Antarctica. Four areas of sea ice were covered with plastics of different UV absorption exposing the bottom- ice algal community to a range of UV doses for a period of 15 days. Algae were exposed to visible radiation only; visible + UV radiation; and visible + enhanced UV radiation. MAA content per cell at the start of the experiment was low in snow covered plots but higher in samples from ice with no snow cover. During the study period, the MAA content per cell reduced in all treatments, but the rate of this decline was less under both ambient UV and visible radiation than under snow covered plots. While low doses of UVB radiation may have stimulated some MAA production (or at least slowed its loss), relatively high doses of UVB radiation resulted in almost complete loss of MAAs from ice algal cells. Despite this reduction in MAA content per cell, the diatoms in all samples grew well, and there was no discernible effect on viability. This suggests that MAAs may play a minor role as photoprotectants in sea ice algae. The unique structure of the bottom ice algal community may provide a self-shading effect such that algal cells closest to the surface of the ice contain more MAAs than those below them and confer a degree of protection on the community as a whole. PMID- 12132688 TI - Antifungal activity of the essential oils from some species of the genus Pinus. AB - The chemical composition of the essential oils from the needles of Pinus ponderosa (north american pine), P. resinosa (red pine) and P. strobus (eastern white pine) has been determined by GC/MS (FID). The essential oils from P. resinosa and P. ponderosa in comparison to P. strobus have been characterized by the higher content of beta-pinene (42.4%, 45.7% and 7.9% respectively). On the other hand, a-pinene (17.7%) and germacrene D (12.2%) were dominant compounds of P strobus. Moreover the essential oil from P. resinosa was more rich in myrcene 15.9%. Estragole and delta-3-carene, each one in amount ca 8% were identified only in P. ponderosa. The content of essential oils in the needles slightly varied--0.65%--P. resinosa, 0.4%--P strobus, 0.3%--P. ponderosa. The antifungal activity has been investigated towards Fusarium culmorum, F solani and F. poae. The strongest activity was observed for the essential oil from P. ponderosa, which fully inhibited the growth of fungi at the following concentrations--F. culmorum, F. solani at 2% and F. poae at 5%. PMID- 12132689 TI - Two epimeric flavalignans from Trichilia catigua (Meliaceae) with antimicrobial activity. AB - A mixture of flavalignan cinchonains Ia and Ib was isolated from the bark of Trichilia catigua. The structures were established on the basis of spectroscopic data of the natural products and their methylated derivatives including 2D NMR experiments, and compared with data in the literature. These flavalignans exhibited antibacterial activity against Bacillus PMID- 12132690 TI - Microbial thansformation of a mixture of argentatin A and incanilin. AB - The biotransformation of a mixture of argentatin A (20%) 1 and incanilin (80%) 2 by Gibberella suabinetti ATCC 20193 and Septomyxa affinis ATCC 6737 demonstrated the conversion of incanilin to 16beta-hydroxylanosta-2, 8, 23-triene, while argentatin A did not react. The acetate of this triterpenoid mixture was biotransformed by Septomyxa affinis ATCC 6737 to give five metabolites. Argentatin A acetate was transformed to 3beta, 16beta,30-trihydroxycycloart-20, 24-diene, 20R, 24R-epoxy-16beta, 25-dihydroxy-3, 4-seco-cycloart-4(28)-en-3-oic acid acetate and 20R, 24R-epoxy-16beta, 25-dihydroxy-3, 4-seco-cycloart-4(28)-en 3-oic acid. Incanilin acetate was converted to 16beta-hydroxylanosta-2, 8, 23 triene and 20R, 24R-epoxy-16beta, 25-dihydroxy-3, 4-seco-lanost-1, 4(28), 8-trien 3-oic acid acetate. The structural elucidations of these metabolites were achieved by different spectroscopic methods. PMID- 12132691 TI - Inhibition of enzymatic reactions. A rapid method to determine the index pI50. AB - The activity of every substance I inhibiting an enzymatic reaction can be approximately evaluated by the index PI50. This paper describes a simple and fast method of estimate and/ or determination of this index. The method is based on the linearity of the dependence of the ratio of reaction rates of uninhibited and inhibited reaction vs. concentration of the inhibitor at constant initial substrate and enzyme concentrations for fully competitive, noncompetitive, uncompetitive and mixed type of inhibition by the one inhibitor. The validity of the method is demonstrated by four inhibitors of hydrolysis of acetylthiocholine by butyrylcholine esterase. PMID- 12132692 TI - Ecto-phosphatase activity on the cell surface of Crithidia deanei. AB - In the present work we have partially characterized an ecto-phosphatase activity in Crithidia deanei, using viable parasites. This enzyme hydrolyzed p nitrophenylphosphate at a rate of 3.55 +/- 0.47 nmol Pi/h x 10(8) cells. The dependence on p-NPP concentration shows a normal Michaelis-Menten kinetics for this phosphatase activity and the value of the apparent Km for p-NPP was 5.35 +/- 0.89 mM. This phosphatase activity was inhibited by the product of the reaction, the inorganic phosphate. Experiments using classical inhibitors of acid phosphatases, such as ZnCl2 and sodium fluoride, as well as inhibitors of phosphotyrosine phosphatase, such as sodium orthovanadate and ammonium molybdate, showed a decrease in this phosphatase activity, with different patterns of inhibition. PMID- 12132694 TI - Photo-induced decomposition of 2-chloroaniline in aqueous solution. AB - A study was performed on the oxidizing degradation of 2-chloroaniline (used as a model pollutant in water) by photolysis (lambda = 254 nm). The change of spectrum and substrate concentration of treated solutions was measured spectrophotometrically as well as by HPLC. The yields of the degradation products (chloride ions, ammonium ions, formaldehyde, etc.) were studied as a function of UV-dose. Their initial quantum yields (Qi) were determined by specific analysis. It was shown that the substrate photolysis in the presence of N2O is most efficient, followed by degradation in media saturated with pure oxygen and air. A probable reaction mechanism for the photo-induced degradation of 2-ClA is presented. PMID- 12132693 TI - Alanine reverses the inhibitory effect of phenylalanine on acetylcholinesterase activity. AB - The aim of this work was to evaluate, in vitro, the effect of L-alanine (Ala) on suckling rat brain acetylcholinesterase (AChE) and on eel Electrophorus electricus pure AChE inhibited by L-phenylalanine (Phe) as well as to investigate whether Phe or Ala is a competitive inhibitor or an effector of the enzyme. AChE activity was determined in brain homogenates and in the pure enzyme after 1 h preincubation with 1.2 mM of Phe or Ala as well as with Phe plus Ala. The activity of the pure AChE was also determined using as a substrate different amounts of acetylthiocholine. Ala reversed completely the inhibited AChE by Phe (18-20% in 500-600 microM substrate, p<0.01). Lineweaver-Burk plots showed that Vmax remained unchanged. However, Km was found increased with Phe (150%, p<0.001), decreased with Ala alone (50%, p<0.001) and unaltered with Phe plus Ala. It is suggested that: a) Phe presents a competitive inhibitory action with the substrate whereas Ala a competitive activation; b) Ala competition with Phe might unbind the latter from AChE molecule inducing the enzyme stimulation; c) Ala might reverse the inhibitory effect of Phe on brain AChE in phenylketonuric patients, if these results are extended into the in vivo reality. PMID- 12132695 TI - Discriminatory power of RAPD, PCR-RFLP and southern blot analyses of ureCD or ureA gene probes on Helicobacter pylori isolates. AB - The genetic diversity of 33 Nigerian Helicobacter pylori isolates were studied using RAPD, PCR-RFLP and Southern blot analysis of ureA or ureCD gene probes. RAPD was able to distinguish the following number of isolates using the primers 3880: 5'-AAGAGCCCGT-3' (28), 3881 :5'-AACGCGCAAC-3' (33) and OPH8 :5'-GAAACACCCC 3' (25). Southern blot analysis using the ureCD probe was also able to distinguish the 12 isolates tested into ten different patterns. The PCR-RFLP technique distinguished all 33 isolates into six types. In conclusion, considering typeability, discriminatory power, and convenience, RAPD with the 3881 primer was considered the most useful technique. PMID- 12132697 TI - Chemical composition of European propolis: expected and unexpected results. AB - Ten propolis samples from Bulgaria, Italy and Switzerland were analyzed by GC-MS. As expected, most samples displayed the typical chemical pattern of "poplar" propolis: they contained pinocembrin, pinobanksin and its 3-O-acetate, chrysin, galangin, prenyl esters of caffeic and ferulic acids. Two samples differed significantly: one from the Graubunden Alpine region, Switzerland, rich in phenolic glycerides, and one from Sicily which contained only a limited number of phenolics and was rich in diterpenic acids. PMID- 12132696 TI - Cyanophycin synthetase-like enzymes of non-cyanobacterial eubacteria: characterization of the polymer produced by a recombinant synthetase of Desulfitobacterium hafniense. AB - Some bacterial genomes were found to contain genes encoding putative proteins with considerable sequence homology to cyanophycin synthetase CphA of cyanobacteria. Such a gene from the Gram-positive, spore-forming anaerobe Desulfitobacterium hafniense was cloned. Expression in Escherichia coli resulted in the formation of a polydispers copolymer of aspartic acid and arginine, with a minor amount of lysine, of about 30 kDa molecular mass. In contrast to cyanophycin, this polymer was water-soluble. The structure of the polymer formed by the synthetase from Desulfitobacterium hafniense was studied by enzymatic degradation with the cyanophycin-specific hydrolase cyanophycinase, and by chemical and mass-spectroscopic analyses. Despite of the differences in solubility, indicating that both polymers cannot be completely identical, the chemical structure was found to be very similar to that of cyanophycin. The results suggest that the use of cyanophycin-like polymers as a nitrogen-rich reserve material is not restricted to cyanobacteria, and that such polymers may not necessarily be stored in granules. PMID- 12132698 TI - Composition of lipophylic extracts from two Tinicates, Styela sp. and Phallusia sp. from the Eastern Mediterranean. AB - Sterols, volatiles and lipids were isolated and identified from lipophylic extracts from two tunicates, Styela sp. and Phallusia sp., occurring in the Eastern Mediterranean. Seventeen sterols were identified. The sterol composition of the two organisms appeared to be similar except for the concentrations of 5alpha-stanols. Both tunicates were characterized by the presence of sterols with a (22Z)-double bond. In the volatiles significant amounts of chlorinated compounds were found (phenols in Styela sp. and hydrocarbons in Phallusia sp.). The fatty acid composition of triacylglycerols and phospholipids of the two tunicates showed significant differences. PMID- 12132699 TI - Aspartic proteinase in Dugesia tigrina (Girard) planaria. AB - A proteolytic activity was identified in Dugesia tigrina planaria using the chromogenic substrate Phe-Ala-Ala-Phe (4-NO2)-Phe-Val-Leu-O4MP. The activity of the enzyme increased four times during the regeneration and presented a maximum at 120 hr being higher in tail than head regenerating segments. The protease that displays this activity was purified from worms by a single step on pepstatin agarose followed by gel-filtration high performance liquid chromatography. The purification resulted in a 34-fold increase in specific activity and the final yield was 10%. The active D. tigrina hydrolase appears to be a dimeric protein composed of identical subunits with 34 kDa associated by disulphide bridges similar to vertebrate cathepsin D. By SDS-PAGE several bands were detected but upon gel filtration HPLC one proteolytically active component, termed Asp-68, was detected and isolated. The maximal activity was observed in a range between pH 3.5-5.0 and the enzyme became inactivated at a pH value above 7.2. The purified enzyme was not inhibited by inhibitors from serine (aprotinin, TPCK, PMSF and TLCK), metallo (EDTA) and cysteine proteinase (E-64) classes. In contrast, inhibitors such as pepstatin, EPNP, and 4-beta-PMA efficiently inhibited the activity of the 68-kDa protease. PMID- 12132701 TI - Stimulatory beetle volatiles for the Asian longhorned beetle, Anoplophora glabripennis (Motschulsky). AB - Two male-specific beetle volatiles were found that elicited strong gas chromatographic-electroantennographic responses from both sexes of Asian longhorned beetle adults, Anoplophora glabripennis. The secretion consisted of a approximately 1:1 (v/v) blend of functionalized dialkyl ethers, 4-(n heptyloxy)butanal and 4-(n-heptyloxy)butan-1-ol. These compounds are chemically unusual natural products that are previously unknown from insects. Laboratory olfactometer studies showed that a blend of 10 microg of each synthetic compound on a filter paper strip was significantly attractive to ALB adults. PMID- 12132700 TI - Antimicrobial activity of the marine alkaloids haminol and pulo'upone and related compounds. AB - The marine alkaloids haminol A, haminol B and pulo'upone as well as 17 related compounds (twelve 2-substituted pyridine derivatives, four 3-substituted ones and one analogue of the bicyclic terminus of pulo'upone) were tested for antimicrobial activity against a panel of six microbes (Bacillus cereus, Escherichia coli, Pseudomonas aeruginosa, Staphylococcus epidermidis, Candida albicans and Saccharomyces cerevisiae) using the paper disc agar diffusion method. Six compounds were tested also against the mold Aspergillus niger. Some of the compounds displayed noteworthy antimicrobial activity, only one congener being completely devoid of activity. Nearly all compounds had activity against B. cereus and S. epidermidis. The growth of E. coli, C albicans and S. cerevisiae was also distinctly inhibited by many compounds. In contrast, most compounds were inactive or had minimal activity against P. aeruginosa. Interestingly, most of the compounds tested against the opportunistic pathogen A. niger were active, one of them having noteworthy inhibitory potency. PMID- 12132702 TI - GC-MS investigation of tropane alkaloids in Datura stramonium. AB - Alkaloids, GS-MS, Datura stramonium The alkaloid spectrum in roots, leaves and seeds of Datura stramonium L. was investigated by GC-MS. Twenty-nine tropane alkaloids are detected. Twelve of them are new constituents for the species and the two tropane esters 3-(3'-acetoxytropoyloxy)tropane (21) and 3-(2' hydroxytropoyloxy)tropane (26) are described for the first time. PMID- 12132703 TI - A quick assay for Na+-K+-AtPase specific activity. AB - The method describes a simultaneous determination of inorganic phosphate (Pi) and protein content from a reaction mixture used for assay of adult rat cerebrocortical synaptosomal membrane Na+-K+-ATPase specific activity. The present method is more convenient, accurate and quicker compared to the existing methods for the determination of Na+-K+-ATPase activity. It also eliminates the possible errors in protein estimation by other classical methods in brain, which have a high lipid content. PMID- 12132704 TI - Safety and security of radiation sources in the aftermath of 11 September 2001. AB - The attack on the United States on 11 September 2001 resulted in an increased awareness of the need for safety and security measures to protect against terrorism. The potential use of radiation sources in terrorism, in particular radioactive sources, was recognized prior to 11 September 2001, but has taken on new significance since. The planning of security measures for radioactive sources must take greater account of the potential for deliberate acts to attack or use radioactive sources to expose people and cause contamination. The potential consequences of an act of terrorism using radioactive sources can be gauged from the consequences of serious accidents that have occurred involving radioactive sources. These include fatal and injurious radiation exposures, contamination of the environment, and serious economic and psychosocial costs the total effect of which is mass disruption. Steps are being taken to improve security for radioactive sources but strategic approaches that can minimize the threat of radiological terrorism should be considered. When justifying a practice that uses radioactive sources, the potential for diversion or use in terrorism should be considered to be a detriment. In this regard, the consideration and development of alternatives to radioactive sources, such as radiation producing machines, have been recommended by terrorism experts as measures to reduce the threat of radiological terrorism. If a practice using radioactive sources is determined to be justified, the need for special security measures to protect against terrorism should then become part of the safety assessment. PMID- 12132705 TI - Plutonium in Colorado residents: results of autopsy bone samples collected during 1975-1979. AB - Concentrations of (239,240)Pu and the 240Pu/239Pu atom ratios were measured in rib samples from 55 non-occupationally exposed Colorado residents. Samples were collected at autopsy during 1975-1979 under an earlier study intended to compare plutonium levels in liver and lung of people who lived at various proximities to the Rocky Flats Environmental Technology Site (RFETS) near Denver. Overall, median (239,240)Pu concentrations from rib samples were 100, 80, and 57 microBq g(-1) ash weight for area locations A, B, and C, respectively. Area A encompassed subjects who lived within 25 km of RFETS, area B was between 25 and 50 km from RFETS, and area C included all of Colorado outside 50 km from the site and east of the continental divide. The corresponding median plutonium skeletal burdens estimated for these area locations were 146, 93, and 71 mBq, respectively. A statistically significant difference was noted only between plutonium concentrations in male rib samples and their skeletal burdens from area A compared to area C. However, based on a regression analysis of all study subjects, distance from RFETS was not statistically correlated to plutonium rib concentrations or skeletal burdens in this sample. Overall, median 240Pu/239Pu atom ratios were 0.20, 0.18, and 0.17 for areas A, B, and C, respectively. Although higher (239,240)Pu concentrations and skeletal burdens were indicated in area A males than area C males, we cannot conclude that RFETS releases may have caused this difference. The decreasing trends in the 240Pu/239Pu ratios with distance from RFETS are contrary with such a conclusion and strongly indicate that the material was primarily global fallout rather than weapons-grade plutonium that was processed at RFETS. Furthermore, there are other plausible explanations for the differences observed between area A and C residents. These include a decreasing trend in global fallout from the Rocky Mountain foothills eastward, smoking history differences, sample selection criteria, and possibly other unidentified factors. Nevertheless, estimated levels of total plutonium in this sample of Colorado residents were low and within the range of reported literature values for general, world-wide populations. PMID- 12132706 TI - Micronucleus assay reveals no radiation effects among nuclear power plant workers. AB - The micronucleus assay was applied as biomarker for exposure effect and radiosensitivity in a group of 99 radiation workers of the Nuclear Power Plant Doel (Belgium). The difference in micronucleus frequency between the group of radiation workers with annual dose exceeding 2 mSv and a non-exposed control population was statistically not significant. With respect to the micronucleus frequency after an in vitro challenge dose of 2 Gy, which can be considered as biomarker for radiosensitivity, the data of present study can be represented by a normal distribution without a high frequency tail. This means that a subpopulation of workers with elevated radiosensitivity could not be identified in the population under study. PMID- 12132707 TI - Individual dose reconstruction among residents living in the vicinity of the Semipalatinsk nuclear test site using EPR spectroscopy of tooth enamel. AB - Individual accumulated doses were determined by EPR spectroscopy of tooth enamel for 26 adult persons residing in territories adjacent to the Semipalatinsk Nuclear Test Site (SNTS). The absorbed dose values due to radiation from nuclear tests were obtained after subtracting the contribution of natural background radiation from the total accumulated dose. The determined dose values ranged up to 250 mGy, except for one person from Semipalatinsk city with a measured dose of 2.8 +/- 0.4 Gy. Increased dose values were determined for the individuals whose teeth were formed before 1962, the end of the atmospheric nuclear tests. These values were found to be significantly larger than those obtained for a group of younger residents of heavily exposed territories and the residents of territories not exposed to radioactive fallout. These increased dose values are consistent with those based on officially registered data for the Northeastern part of Kazakstan adjacent to SNTS, which was exposed to high levels of radioactive fallout from nuclear tests in period 1949-1962. PMID- 12132708 TI - Variation of uranium isotopic composition in soil within the JCO grounds from the 30 September 1999 criticality accident at JCO, Tokai-mura, Japan. AB - Following the 30 September 1999 criticality accident at JCO, 29 surface and 3 core soil samples were collected inside and outside the JCO grounds to evaluate possible contamination by 235U-enriched uranium (18.8%) being handled at the time of the accident. Uranium (234U, 235U, and 238U) and thorium (228Th, 230Th, and 232Th) isotopes were determined by alpha-spectrometry and ICP-MS after radiochemical separation. Concentrations of 238U and 234U ranged from 11.3 to 63.5 and 11.6 to 360 mBq g(-1), respectively. Higher amounts of 238U and/or 234U were found in the vicinity of the uranium conversion building. The calculated 234U/235U activity ratios ranged from a 1.0 radioactive equilibrium value to an unusually high 5.7 value. Several of the soil samples showed considerably higher 235U/238U atomic ratios (1.06-4.37%) than 0.725% for natural uranium. Based on the assumption that measured U-series nuclides in soil samples taken from the JCO grounds were almost at radioactive equilibrium up to 230Th, excess uranium could be calculated for each sample. The results suggest that the excess uranium in the soils have lower 235U/238U atomic ratios (a few %) than the 18.8% enrichment of the precipitation tank uranium. PMID- 12132709 TI - A re-evaluation of the 131I atmospheric releases from the Hanford site. AB - The atmospheric release of 131I from the Hanford site for the 1950's and 1960's, focused on the period of releases after the year 1950, has been re-evaluated using processing plant stack monitoring data to address a series of questions and concerns that have arisen related to the source term. Historical stack monitoring data have been used to re-assess the releases by creating either a release factor to use with the calculated plant throughput or using the stack monitoring results as the basic estimate, and the results have been verified using historical atmospheric monitoring data from a location several kilometers distant. Uncertainties in all of the historical data have been addressed in the re assessment. Compared to the original estimate between 1950 and 1971 of 2.46 +/- 0.71 PBq, the stack monitoring results show a release of 131I to the atmosphere of 1.55 +/- 0.23 PBq. The concurrent atmospheric monitoring results imply a release of 1.75 +/- 0.11 PBq over the same period, but this result is inflated by inclusion of global fallout The total effective dose estimated to a full-time, nearby adult resident from 131I using the Heeb source term from 1950 through 1972 is 0.73 mSv; using the source term based on stack monitoring data in the Hanford Environmental Dose Reconstruction project models, it is 0.51 mSv. PMID- 12132710 TI - Measuring the external exposure dose in the contaminated area near the Chernobyl nuclear power station using the thermoluminescence of quartz in bricks. AB - We collected bricks from buildings in the heavily contaminated evacuated area of Belarus in a 30-km zone around the Chernobyl nuclear power station and the Gomel Bryansk area of 150-250 km from Chernobyl and estimated the cumulative radiation dose caused by the reactor accident by measuring the thermoluminescence (TL) of the bricks. The annual dose at each location was measured using glass dosimeters and thermoluminescence dosimeters (TLD). The dose rate was measured using an energy-compensated NaI scintillation survey meter. The soil contamination near the location of each brick was measured using a germanium semiconductor detector. The main purpose of the project was to extrapolate the relation between the cumulative external dose and the present dose rate or contamination level to the lower contaminated areas. The results of the glass dosimeter, TLD, and survey meter determinations were almost identical. For a determination of the annual dose higher than 10 mGy y(-1), the cumulative dose by TL (TL dose) was roughly proportional to the annual dose and about 1.5 times larger than the cumulative dose calculated from the annual dose and 137Cs half life. The difference is expected due to the contribution of short-lived nuclides immediately after the accident or localized heavy contamination of the ground surface with 137Cs that migrated afterwards. For annual dose smaller than 10 mGy y(-1), the proportionality was not observed and most of the locations facing indoors showed TL doses very much larger than that expected from the proportionality. The cumulative dose outdoors by TL was also roughly proportional to the regional 137Cs contamination level and the proportional constant is about 10(-1) mGy per GBq km(-2), and is about 250 times larger than the present annual internal dose derived from published results. The correlation between the present dose rate where the brick was sampled and the average 137Cs concentration in the ground soil near the point is not clear, possibly because of the large spatial fluctuation in the 137Cs concentration in the soil. PMID- 12132711 TI - Do long-term average radon concentrations in schools and kindergartens differ from the average during working hours? AB - Radon concentrations in air in schools and kindergartens were measured by alpha scintillation cells, electret and etched track detectors, and a continuous device. Values obtained by different devices under different conditions are compared and their correlation discussed. The radon concentration averaged over working hours may differ substantially from the whole-day average, although Pearson's correlation coefficients between the two averages of 0.83 and 0.82 were obtained for schools and kindergartens, respectively. PMID- 12132712 TI - Monte Carlo simulation of single and two-dosimeter approaches in a steam generator channel head. AB - In a steam generator channel head, it was not unusual to see radiation workers wearing as many as twelve dosimeters over the surface of the body to avoid a possible underestimation of effective dose equivalent (H(E)) or effective dose (E). This study shows that only one or two dosimeters can be used to estimate H(E) and E without a significant underestimation. MCNP and a point-kernel approach were used to model various exposure situations in a steam generator channel head. The single-dosimeter approach (on the chest) was found to underestimate H(E) and E significantly for a few exposure situations, i.e., when the major portion of radiation source is located in the backside of a radiation worker. In this case, the photons from the source pass through the body and are attenuated before reaching the dosimeter on the chest. To assure that a single dosimeter provides a good estimate of worker dose, these few exposure situations cannot dominate a worker's exposure. On the other hand, the two-dosimeter approach (on the chest and back) predicts H(E) and E very well, hardly ever underestimating these quantities by more than 4% considering all worker positions and contamination situations in a steam generator channel head. This study shows that two dosimeters are adequate for an accurate estimation of H(E) and E in a steam generator channel head. PMID- 12132713 TI - Measurement of TVLs in lead for 4, 6 and 10 MV bremsstrahlung x-ray beams at scattering angles between 30 degrees and 135 degrees. AB - Measurements have been made of the transmission factors through lead for scattered radiation produced by 4, 6, and 10 MV bremsstrahlung x-ray beams incident on a polystyrene phantom; these measurements cover the range of scattering angles between 30 degrees and 135 degrees. The results show that the tenth value layer (TVL) for scattered radiation decreases sharply with increasing scattering angle and increases slightly with increasing beam energy, although at large scattering angles there is little energy dependence. TVLs range from 3.5 cm to 0.3 cm for 4 MV at scattering angles between 30 degrees and 135 degrees, from 3.8 cm to 0.6 cm for 6 MV, and from 4.2 cm to 0.7 cm for 10 MV, respectively, at scattering angles between 30 degrees and 120 degrees. Monte Carlo calculations, performed at 4 MV to simulate the transmission of scattered radiation, are in good agreement with the experimental measurements. PMID- 12132714 TI - 210Po implanted in glass surfaces by long term exposure to indoor radon. AB - Recent epidemiologic investigations of the relationship between residential radon gas exposure and lung cancer relied on contemporary radon gas measurements to estimate past radon gas exposures. Significant uncertainties in these exposure estimates can arise from year-to-year variation of indoor radon concentrations and subject mobility. Surface implanted 210Po has shown potential for improving retrospective radon gas exposure estimates. However, in previous studies, the ability of implanted 210Po activity to reconstruct cumulative radon gas exposure was not tested because glass was not available from homes with known radon-gas concentration histories. In this study, we tested the validity of the retrospective radon gas reconstruction using implanted 210Po surface activity by measuring glass surfaces from homes whose annual-average radon gas concentrations had been measured almost every year during two decades. Regression analysis showed a higher correlation between measured surface activity and cumulative radon gas exposure in these homes (R2>0.8) than was observed in homes where only contemporary radon gas measurements were available. The regression slope (0.57 ky m(-1)) was consistent with our earlier retrospective results. Surface activity measurements were as reliable for retrospective radon gas exposure reconstruction as yearlong gas measurements. Both methods produced estimates that were within 25% of the long-term average radon gas concentrations in a home. Surface measurements can be used for home screening tests because they can provide rapid, reliable estimates of past radon gas concentrations. Implanted 210Po measurements are also useful in retrospective epidemiologic studies that include participants who may have been exposed to highly variable radon concentrations in previously occupied or structurally modified homes. PMID- 12132715 TI - A new graphical user interface for fast construction of computation phantoms and MCNP calculations: application to calibration of in vivo measurement systems. AB - The paper reports on a new utility for development of computational phantoms for Monte Carlo calculations and data analysis for in vivo measurements of radionuclides deposited in tissues. The individual properties of each worker can be acquired for a rather precise geometric representation of his (her) anatomy, which is particularly important for low energy gamma ray emitting sources such as thorium, uranium, plutonium and other actinides. The software discussed here enables automatic creation of an MCNP input data file based on scanning data. The utility includes segmentation of images obtained with either computed tomography or magnetic resonance imaging by distinguishing tissues according to their signal (brightness) and specification of the source and detector. In addition, a coupling of individual voxels within the tissue is used to reduce the memory demand and to increase the calculational speed. The utility was tested for low energy emitters in plastic and biological tissues as well as for computed tomography and magnetic resonance imaging scanning information. PMID- 12132716 TI - Depleted uranium in military conflicts and the impact on the environment. AB - Kosovo was bombarded by fired shells (bullets) with depleted uranium (DU) during April 1999. Around 30,000 depleted uranium rounds (projectiles) were fired, and about 10 tons of the DU debris were scattered across Kosovo. In reviewing the data on environmental measurements for depleted uranium collected by field missions in the Kosovo area during the period of 5-19 November 2000 (1.5 y following the 1999 conflict), evidence of depleted uranium was found only in soil samples at localized points of concentrated contamination. Concentrations varied from a few mg (2.34 mg) DU per kg soil (29 Bq DU/kg soil) at depths of 39.5-44.5 cm up to about 18 g DU per kg soil (225,760 Bq DU/kg soil) at depths of 0-5 cm surface soil. There were no signs of depleted uranium in waters. However, in most (80%) of the 145 soil (core) samples reported by UNEP, 238U was lower than 100 Bq per kg soil (the lowest was 8.8 Bq per kg soil) in 112 locations of widespread contamination. PMID- 12132717 TI - Estimating populations for collective dose calculations. AB - The collective dose provides an estimate of the effects of facility operations on the public based on an estimate of the population in the area. Geographic information system software, electronic population data resources, and a personal computer were used to develop estimates of population within 80-km radii of two sites. PMID- 12132718 TI - Temporal variation of radon progeny ratio in outdoor air. AB - This paper investigates the concentration ratio of radon progeny (218Po, 214Pb, and 214Bi) in outdoor air. From 1988 to 1998, alpha spectroscopy was used intermittently to collect progeny data outside the author's office located in the northeastern part of Japan. Let the quantity R(EEC) represent the ratio EEC(218Po)/[EEC(214Pb) + EEC(214Bi)], where, e.g., EEC(218Po) denotes the equilibrium equivalent radon concentration contributed by 218Po. The concentration ratio R(EEC) correlates with outdoor air stability. Statistical and time series analyses of R(EEC) indicate (1) outdoor air is more stable in the morning than in the afternoon, (2) outdoor air is more stable in summer/autumn than in winter/spring, and (3) in spite of no significant correlation between R(EEC) and wind speed, the power spectrum of R(EEC) appears similar to that of wind speed. This is probably due to the fact that near-surface mixing is not very sensitive to wind speed. PMID- 12132720 TI - Data sets for transport code testing. PMID- 12132719 TI - French approach for the distribution of iodine tablets in the vicinity of nuclear power plants. AB - In the event of an accident, isotopes of iodine including 131I can be released into the atmosphere. In 1997, as a safety measure, the French government decided to begin the distribution of stable iodine tablets in advance, directly to those living in the vicinity of the nuclear power plants, to avoid having to do so in an emergency. The tablets were previously stored by Electricite de France (EDF), which held them at the disposal of the government authorities. This year, as the existing tablets pass their use--by date, EDF has begun redistributing stable iodine within a ten-kilometer radius around its nineteen nuclear sites. We review the effectiveness of this countermeasure as well as the nature and incidence of possible side effects while measuring the duration of its action under the conditions in which it was administered. A bibliographic study of the kinetics of iodine in the human body has enabled the indications and the means of use to be determined. The effectiveness of the preventive effect and the onset of thyroid dysfunction depends on both external and individual factors: uptake of iodine from food, functional condition of the thyroid, and age. In an individual with a healthy thyroid, taking 100 mg of stable iodine immediately before exposure to radioactive iodine reduces the dose to the thyroid by at least 95%. In cases of prolonged exposure, the reduction is smaller. Therefore, if exposure lasts for a number of days, consideration needs to be given to taking stable iodine again, to maintain maximum protection. In addition to the bibliographic study, this presentation covers the impact of making iodine available and the action taken to educate the public; the attitudes of populations concerned; and the reaction of the health professionals. PMID- 12132721 TI - INEEL cleanup. Idaho National Engineering and Environmental Laboratory. PMID- 12132722 TI - Radioactive drug research program audit checklist. AB - Many institutions conduct human research studies that involve the administration of radiopharmaceuticals. The Radiation Safety Officer (RSO) may be the person responsible for ensuring that such studies are performed safely and in compliance with applicable regulations. Sometimes, RSOs assigned this responsibility lack the knowledge and experience necessary to oversee such research activities. In an effort to assist RSOs unfamiliar with radioactive drug research regulations to establish an effective compliance program in this field, the authors developed a comprehensive audit checklist. Periodic use of this checklist may be effective in ensuring compliance with applicable regulations. PMID- 12132723 TI - A whole body counting facility in a remote Enewetak Island setting. AB - The U.S. Department of Energy (DOE) has recently implemented a series of strategic initiatives to address long-term radiological surveillance needs at former U.S. test sites in the Marshall Islands. The plan is to engage local atoll communities in developing shared responsibilities for implementing radiation protection programs for resettled and resettling populations. As part of this new initiative, DOE agreed to design and construct a radiological laboratory on Enewetak Island, and help develop the necessary local resources to maintain and operate the facility. This cooperative effort was formalized in August 2000 between the DOE, the Republic of the Marshall Islands (RMI), and the Enewetak/Ujelang Local Atoll Government (EULGOV). The laboratory facility was completed in May 2001. The laboratory incorporates both a permanent whole body counting system to assess internal exposures to 137Cs, and clean living space for people providing 24-h void urine samples. DOE continues to provide on-going technical assistance, training, and data quality review while EULGOV provides manpower and infrastructure development to sustain facility operations on a full time basis. This paper will detail the special construction, transportation and installation issues in establishing a whole body counting facility in an isolated, harsh environmental setting. PMID- 12132724 TI - The investigation and identification of a civil defense canister. AB - A Civil Defense canister was found in a high school storage area with no radioactive labels or instructions. Initial screening with a 2" x 2" NaI detector indicated elevated count rates above background. Using field instruments with gamma peak analysis features and a laboratory counting system, the collective effort indicated that the contents included 238U and 235U. A Civil Defense Item Specification introduced later in the investigation show that the canister contained approximately 3.0 g of uranyl acetate. The specification also stated that the purpose of the canister was to determine the amount of radioactive material in water and solid, or liquid food samples. PMID- 12132725 TI - Does elemental mercury exist in the workplace? AB - Elemental mercury can exist at sites undergoing decontamination and decommissioning operations from instruments containing mercury, mercury vacuum pumps, chemical operations, electrical components, and pharmaceutical production. Elemental mercury has been known to be a severe health hazard. Recent studies, utilizing more sensitive methods, are able to detect subclinical effects from mercury exposure at lower concentrations than in past studies. Precautions must be taken if elemental mercury may be present in the workplace. PMID- 12132726 TI - A comprehensive laser safety program for the research university setting. AB - Laser safety often receives less attention and fewer resources than other institutional safety and health program elements. This diminished support occurs despite the fact that laser-related occupational injuries and deaths greatly exceed those caused by biomedical research use of radioactive material. Therefore, a small emphasis on laser safety can potentially produce a much greater return on investment than more conventional health physics programs. Developing and implementing a formal comprehensive laser safety program enhances safe laser use and offers a great professional development opportunity in this important area of health physics expertise. This paper outlines such a university laser safety program. PMID- 12132727 TI - Medical laser safety hazard evaluation. AB - Health care laser systems offer general laser hazards and additional specific concerns unique to the clinical environment. A formal laser hazard evaluation procedure provides an efficient mechanism for identifying potential laser safety hazards. This paper outlines such a medical laser hazard evaluation program and highlights the unique characteristics of medical lasers. PMID- 12132728 TI - RSO interview. Interview with John P. Hageman. PMID- 12132729 TI - Field comparison of the sampling efficacy of two smear media: cotton fiber and kraft paper. AB - Two materials were compared in field tests at the Defense Waste Processing Facility: kraft paper (a strong, brown paper made from wood pulp prepared with a sodium sulfate solution) and cotton fiber. Based on a sampling of 46 pairs of smears, the cotton fiber smears provide a greater sensitivity. The cotton fiber smears collected an average of 44% more beta activity than the kraft paper smears and 29% more alpha actvity. Results show a greater sensitivity with cotton fiber over kraft paper at the 95% confidence level. Regulatory requirements for smear materials are vague. The data demonstrate that the difference in sensitivity of smear materials could lead to a large difference in reported results that are subsequently used for meeting shipping regulations or evaluating workplace contamination levels. PMID- 12132730 TI - Equations for modeling of grab samples of radon decay products. AB - A mathematical model of the process of collecting and analyzing a grab sample of radon decay products in air can be useful for a number ofpurposes. Among them is predicting the observed counts of alpha particles and/or beta particles (1) under various conditions of radon decay product equilibrium; (2) for various sampling, delay and counting times; and (3) for various counting efficiencies for the particles emitted by the collected decay products. PMID- 12132731 TI - Interview with John J. Pickering. PMID- 12132732 TI - Interview with Brenda Pangborn [interivew]. PMID- 12132733 TI - A novel lymphatic mapping technique to improve localization and staging of early colon cancer during laparoscopic colectomy. AB - Encouraging results from our previous studies of sentinel lymph node (SLN) mapping in colorectal cancer (CRC) prompted investigation of its feasibility and accuracy during laparoscopic colectomy for early CRC. Between 1996 and 2000, 14 patients with clinically localized colorectal neoplasms underwent colonoscopic tattooing of the primary site and SLN mapping. In each case 0.5 to 1 cm3 of isosulfan blue dye was injected submucosally via the colonoscope. The blue stained lymphatics were visualized through the laparoscope and followed to the SLN, which was marked with a clip, and laparoscopic colectomy was completed in the routine fashion. All lymph nodes were examined by hematoxylin and eosin (H&E) staining; in addition each SLN was subjected to focused examination by multisectioning and immunohistochemical staining using cytokeratin antibody. In all 14 patients the primary neoplasm and an SLN were identified laparoscopically. An average of 13.5 total lymph nodes and 1.7 SLNs per patient were identified. The SLN correctly reflected the tumor status of the nodal basin in 93 per cent of the cases. In four cases with unexpected lymphatic drainage, the extent of mesenteric resection was altered. In two cases (14%), nodal involvement was micrometastatic, confined to an SLN, and identified only by immunohistochemical staining. Lymphatic mapping caused no complications and added only 10 to 15 minutes to the overall operative time. Comparison of results in this group with results for a matched group of 14 patients undergoing SLN mapping during open colon resection showed that the laparoscopic technique had similar rates of accuracy and success. These preliminary findings indicate that colonoscopic/laparoscopic SLN mapping during laparoscopic colon resection is a feasible and technically simple means of identifying the primary colorectal neoplasm and its SLN. Focused pathologic examination of this node can upstage CRC and thereby may improve selection of patients for adjuvant chemotherapy. PMID- 12132734 TI - Blood transfusions correlate with infections in trauma patients in a dose dependent manner. AB - Infections are a common and significant sequela of major traumatic injury. The objective of this study was to evaluate the relationship between infections in trauma patients and the transfusion of packed red blood cells (pRBCs) within the first 48 hours of admission. We hypothesized that transfusions of pRBCs were associated with an increased risk of infection in a dose-dependent manner. All adult patients admitted to the trauma service of a Level I trauma center from November 1996 to December 1999 were studied. Secondary analysis was performed on prospectively collected data. One thousand five hundred ninety-three consecutive patients were studied; of these 12.6 per cent developed at least one infection. The overall transfusion rate was 19.4 per cent. The infection rate in patients who received at least one transfusion was significantly higher (P < 0.0001) at 33.0 versus 7.6 per cent in patients receiving no pRBCs. Transfusions per patient ranged from 0 to 46 units. There was a clear exponential correlation in patients receiving between 0 and 15 transfusions (R2 = 0.757). Multivariate logistic regression, which was used to identify risk factors for the development of infection, demonstrated the odds ratio of receiving pRBCs to be 1.084, with a 95 per cent confidence interval of 1.028 to 1.142 (P = 0.0028). In summary there is a clear dose-dependent correlation between transfusions of pRBCs and the development of infection in trauma patients. Multivariate analysis further demonstrated that pRBCs were an independent risk factor for the development of infections. Although transfusions are frequently indicated, they should be administered appropriately and with no more pRBCs than absolutely necessary. PMID- 12132735 TI - When the bough breaks: a 10-year review of logging injuries treated at a rural trauma center in Pennsylvania. AB - The purpose of this study was to describe logging-related injuries at a Pennsylvania trauma center and evaluate the impact of helmets and profession. Charts of patients from 1990 through 1999 admitted after logging accidents were retrospectively reviewed. Most injuries were to the head (71 in 28 patients). Injury Severity Score was greater for helmeted loggers (21.0 vs 11.8; P = 0.02) but did not differ by profession. Mean hospital stay was not significantly different for helmeted loggers (9.6 vs 5.4 days, P = 0.499). Mean length of stay was 8.2 days and 3.5 days for professionals and nonprofessionals, respectively (P = 0.01). Professional loggers tended to remain hospitalized longer. Helmet use does not appear to affect injury severity; however, this could be a result of the catastrophic nature of many logging accidents. Most loggers in this study were not wearing helmets, which suggests that improved educational efforts are needed to improve safety in this dangerous occupation. PMID- 12132736 TI - Hemoperitoneum from spontaneous rupture of a liver cell adenoma in a male with hyperthyroidism. AB - Spontaneous liver rupture is uncommon, is difficult to diagnose, and carries a high mortality. Liver cell adenoma is a rare benign liver tumor with increasing incidence in women on oral contraceptive pills, and they have been reported to rupture spontaneously. In men such a phenomenon is an extreme rarity. In animal experiments thyroid hormone is proven to play a role in the growth of liver cell derived neoplasms as they do in normal hepatocyte proliferation. An association of liver cell adenoma and hyperthyroidism in humans has not been previously reported. We present the successful management of an unusual case of spontaneous hemoperitoneum from rupture of a liver cell adenoma in a young man with hyperthyroidism. PMID- 12132737 TI - Adenocarcinoma of the gastric cardia: treatment via a left thoracoabdominal approach. AB - We examined the outcome of adenocarcinomas of the gastric cardia treated by total or proximal gastrectomy, lower esophagectomy, and D2 lymphadenectomy via a left thoracoabdominal approach. We compare these results with those of other methods as well as review the literature. During a 10-year period (1991-2000) 180 patients with primary gastric cancer were admitted to our department. Thirty-six of the patients had adenocarcinoma of the cardia. Twenty-four patients underwent total gastrectomy, D2 lymphadenectomy, and esophagectomy, and four others underwent proximal gastrectomy and esophagectomy with esophagogastric anastomosis via a left thoracoabdominal approach. These latter 28 patients compose our study group. We had no operative mortality, the morbidity varied, and the quality of life and the loss of body weight ranged within satisfactory levels, but the survival rate was rather poor. The median survival time was 19 +/- 1.2 months. Survival was significantly longer in patients with less than 40 per cent positive resected lymph nodes (P = 0.035). From the resulting data and our experience we believe that the left thoracoabdominal approach gives excellent exposure for radical resection of cancer of the gastric cardia and should be the procedure of choice for curative resection of such tumors. This approach combined with total gastrectomy and D2 lymphadenectomy can be performed with an acceptably low mortality rate; it provides good palliation but not encouraging survival rates. Although it is less radical proximal gastrectomy gives the same results and a better quality of life but may be performed only in the early stages of the disease. PMID- 12132738 TI - Leg revascularization in patients with calciphylaxis. AB - Calciphylaxis is a rare complication in patients undergoing hemodialysis. The pathogenesis and risk factors for this disease are poorly understood, although an association with secondary hyperparathyroidism has been suggested. Only two cases of lower-extremity revascularization of patients with calciphylaxis appear in the literature; this report adds two more. PMID- 12132739 TI - A new perspective in appendicitis: calculation of half time (T(1/2)) for perforation. AB - Appendicitis is generally a more serious disease in the elderly than in the young. In the former, perforation is seen commonly leading to the belief that the appendix perforates more readily and rapidly in the elderly. A competing view is that the appendix perforates relatively more frequently in the elderly than in the young. To distinguish between these two views we analyzed 126 cases of acute appendicitis stratified by age group. The time between onset of symptoms and perforation was calculated with a novel method that utilized the biological concept of T(1/2) for perforation. Our findings suggest that the rate of perforation in the elderly is not significantly different from that in the young but the frequency of perforation is higher in the elderly. We concluded that appendicitis carries a graver prognosis in the elderly because the frequency of appendiceal perforation is higher in the elderly. PMID- 12132740 TI - Recurrent ischemia in a young man with the antiphospholipid syndrome. AB - Recurrent thromboses, cerebral disease, miscarriages, and antiphospholipid antibodies are characteristic of the antiphospholipid syndrome. A 31-year-old man presented with limb ischemic and isolated right ventricular failure. Antiphospholipid syndrome was suspected and limb salvage was accomplished by anticoagulation and tibial- to-plantar artery bypass surgery. However, recurrent ischemic episodes were successfully treated with thrombolytic therapy and anticoagulants. The surgeon should be aware that patients with antiphospholipid antibodies and lupus anticoagulant antibodies have a high propensity for recurrent arterial thromboses and should use multiple therapeutic approaches to effect successful long-term limb salvage. PMID- 12132741 TI - Transmural migration of a retained laparotomy sponge. AB - The most common surgically retained foreign body is the laparotomy sponge. The clinical presentation of a retained sponge can vary from an incidental finding on plain radiograph to an intense inflammatory response with obstruction or perforation. In the case described here a patient reported abdominal pain 11 months after her hysterectomy. Although two sponge counts appeared in the operative record one laparotomy sponge had been overlooked. Apparently an inflammatory response created an abscess pocket around the sponge between the abdominal wall and the ileum resulting in perforation of the ileum. Through this opening the sponge migrated into the lumen of the small bowel, from which it was surgically removed. The patient recovered without complications. The case highlights the importance of a thorough exploration of all quadrants of the abdomen at the termination of surgical cases. PMID- 12132742 TI - Major lower-extremity amputation: contemporary experience in a single Veterans Affairs institution. AB - Our objective is to describe our current experience with major lower-extremity amputation secondary to vascular disease. We conducted a retrospective review of sequential amputations over a 3-year period at one Veterans Affairs institution. One hundred thirteen amputations were performed in 99 men (age 70 +/- 11 years). Seventy-five per cent were diabetic and 23 per cent were on dialysis. Fifty-six per cent were primary amputations. The final AKA/BKA (above-knee to below-knee amputation) ratio was 3:2 and was not related to prior bypass, ethnicity, or dialysis status (P > 0.5). Forty-three per cent of amputations were BKAs in diabetics versus 26 per cent in nondiabetics (P = 0.08). The in-hospital and 30 day mortality rates were 2.6 and 8 per cent and were not related to amputation level (P = 0.76). Forty per cent experienced postoperative complications that were most frequently wound related (22%). Wound complications were more frequent with BKA than AKA (P = 0.04). At an average follow-up of 10 +/- 8 months only 65 per cent were alive. Although 51 per cent were discharged to rehabilitation units only 26 per cent regularly wore a prosthesis with 23 per cent ambulating. BKA patients were more likely to ambulate than AKA (34% vs 9%; P = 0.001), and dialysis patients were less likely to ambulate than nondialysis patients (5% vs 25%; P < 0.02). During follow-up 17 per cent of patients discharged with an intact contralateral limb required amputation of that limb and 7 per cent had bypass surgery on that limb. Complication rates were higher in African Americans and Hispanics than in whites (59%, 45%, and 23%, respectively; P < 0.001), although mortality and ambulation rates were similar. Despite an acceptable perioperative mortality complication rates remain high especially in nonwhites. One-year mortality is high. Low rehabilitation rates especially in dialysis patients mandate further efforts in this regard. Vigilant follow-up of the contralateral limb is essential. PMID- 12132743 TI - Non-greater saphenous vein grafting for infrageniculate bypass. AB - Infrainguinal bypass grafting with greater saphenous vein has proven to be a highly effective procedure with primary 5-year patency and limb salvage rates exceeding 80 per cent. However, because of prior usage or intrinsic venous disease the greater saphenous vein is often not available as a conduit. Numerous studies have shown that patency rates for prosthetic bypass grafting to the infrageniculate vessels are clearly inferior to that reported for greater saphenous vein bypass. In this report we summarize our experience with the use of alternate autogenous vein grafting to the infrageniculate vessels. The records of all patients undergoing autogenous bypass grafting to the infrageniculate vessels using a conduit other than the greater saphenous vein between 1992 and 1999 were reviewed. Graft survival curves were plotted using the Kaplan-Meier method and results are reported using the Society for Vascular Surgery/International Society for Cardiovascular Surgery guidelines. Forty-eight patients underwent a total of 51 infrageniculate bypass procedures using non-greater saphenous autogenous conduits. Thirty-nine patients had reconstructions performed with single segments of arm vein, two had their operations performed with lesser saphenous vein, and ten had grafts created with two segments of non-greater saphenous autogenous vein. Twenty-one grafts were performed to the infrageniculate popliteal artery and 30 were performed to the tibial vessels. Primary and primary assisted patency rates at 30 months were 49 and 75 per cent. Limb salvage was 87 per cent. Infrainguinal bypass grafting using non-greater saphenous autogenous conduits can yield quite satisfactory intermediate limb salvage and patency rates. However, close graft surveillance and prompt intervention are required to avoid graft failure. PMID- 12132744 TI - Closure of proximal colorectal fistulas using fibrin sealant. AB - Fibrin glue has been used in upper gastrointestinal and perianal fistula disease, but its success in proximal colorectal pathology has not been widely documented. This report describes the use of endoscopically injected fibrin glue as a successful adjunct to traditional methods in accelerating the closure of colorectal fistulas. A retrospective review was performed on cases of colon and rectal fistulas treated with fibrin glue using an endoscopic technique of injection. Fistulas were injected via a flexible fiberoptic endoscope with fluoroscopic guidance (three) or directly with a rigid proctoscope (one). Fibrin glue was mixed directly from cryoprecipitate, thrombin, and calcium (one) or using a Tisseel kit (three) (Baxter, Deerfield, IL). Four patients were identified and included: two J-pouch fistulas, a colocutaneous fistula, and a complex rectocutaneous fistula. The median duration of fistula was 33 days (range 4-365 days). Total parenteral nutrition and bowel rest were used in two patients and three required drainage of an abscess. All fistulas were obliterated and patients required a mean of one application of fibrin glue (range one to two). The mean time to resuming a regular diet postinjection was 2 days (range 1-5). No complications were identified. Fistula resolution was documented in all cases with a contrast enema and no patient has had a fistula recurrence at a median follow-up of 12 months (range 6-65). This preliminary series demonstrates that fibrin glue can be used to obliterate proximal rectal, colonic, and pouch fistulas. Endoscopy and fluoroscopy may aid in administering the fibrin glue. This adjunctive technique may shorten the time to fistula closure and may allow some patients to avoid further surgery. PMID- 12132745 TI - Incidental cholecystectomy during laparoscopic antireflux surgery. AB - Cholelithiasis and gastroesophageal reflux are both very common diseases that may occur simultaneously. Management of asymptomatic gallstones is still controversial. Because severe complications due to gallstones may occur incidental cholecystectomy during nonrelated abdominal surgery may be offered to patients with coexisting gallbladder disease. The aim of this study was to assess the clinical outcome of patients after laparoscopic fundoplication and incidental cholecystectomy for cholelithiasis compared with the outcome of patients after fundoplication alone. We conducted a retrospective chart review and prospective analysis using a questionnaire of the clinical outcome of patients who underwent laparoscopic fundoplication and incidental cholecystectomy from June 1991 to January 2000 in comparison with sex- and age-matched patients who had antireflux surgery alone. Sixty-seven (6.3%) of 1065 patients had a laparoscopic cholecystectomy at the time of laparoscopic antireflux surgery; 101 (75%) of 134 answered the questionnaire. The mean follow-up time was 4.6 years. Laparoscopic cholecystectomy did not influence surgical morbidity or mortality. Postoperative symptom score (1-10) did not show a statistically significant difference regarding bloating, diarrhea, abdominal pain, nausea, vomiting, biliary problems, jaundice, pancreatitis, dysphagia for liquids and solid, heartburn, regurgitation, and chest pain when the two groups were compared. We conclude that incidental cholecystectomy during laparoscopic antireflux surgery is safe and does not appear to influence the clinical outcome of the antireflux procedure. PMID- 12132746 TI - Pattern of injury from personal watercraft. AB - Injury from personal watercraft has continued to increase. Prior attempts to delineate patterns of injury and relative frequencies have yielded varied results. We retrospectively reviewed Trauma Registry data and charts of all patients who suffered personal watercraft injury treated at the R. Adams Cowley Shock Trauma Center between August 1996 and January 2001. Patient demographics included mechanism of injury, injuries sustained, and outcomes. Attempts were made to correlate events around the injury and injury pattern. During the study period 24 patients were treated. Mechanisms consisted of direct collision, fails from the watercraft, handlebar straddle injuries, axial loading, and hydrostatic jet injury. Traumatic brain injury was most common occurring in 54 per cent of patients. Spinal injury was also common occurring in 29 per cent of patients. Axial loading from falls while wave jumping seemed to correlate with skeletal injury. Thoracolumbar spine injury were often skeletally unstable requiring either brace or operative fixation. Inexperience and reckless behavior were found to be the greatest contributing factors. Substance abuse did not influence injury. PMID- 12132747 TI - Anal complications after restorative proctocolectomy (J-pouch). AB - A prospective assessment was performed to determine the incidence of anal complications after ileoanal J-pouch anastomosis procedures for ulcerative colitis (UC) and familial adenomatous polyposis (FAP). From 1989 to 2000, 75 patients (50 male and 25 female) underwent proctocolectomy and ileal pouch-anal anastomosis with temporary loop ileostomy for UC (N = 68) and FAP (N = 7). Overall 33 patients (44%) developed anal complications postoperatively. Nineteen patients (25%) had mild anal stenosis amenable to digital dilatation in the office. Ten patients (13%) had severe anal stenosis requiring operative dilatation. Ileostomy closure was delayed longer than 3 months in four patients because of anal stenosis. One patient never had his ileostomy closed secondary to severe anal stenosis. Anal fissures developed in one patient that resolved with conservative treatment. Three patients developed fistula-in-ano and one patient developed a pouch-vaginal fistula. Of these four patients two later manifested signs of Crohn's disease. Four patients developed perirectal abscesses (three without fistulas) that were treated with incision and drainage. Two patients had presacral (anastomotic) abscesses; one patient was treated with temporary anastomotic diversion and the other underwent a permanent ileostomy and pouch resection. Both of these patients were later diagnosed with Crohn's disease. Anal complications developed in 17 of 41 (41%) handsewn anastomoses, 16 of 34 (47%) stapled anastomoses, three of seven (43%) patients with FAP, and 30 of 68 (44%) patients with UC. Operative technique and disease type did not significantly correlate with the type of anal complication. However, hand-sewn anastomoses had a higher incidence of severe strictures and FAP patients did not develop anal abscesses, fistulas, or fissures. Forty-five per cent of our patients with abscesses/fistulas and all of our patients with presacral abscesses from anastomotic dehiscence were later diagnosed with Crohn's disease. Anal complications after ileoanal J-pouch anastomosis are relatively common. PMID- 12132748 TI - Solid-pseudopapillary tumor of the pancreas masquerading as a pancreatic pseudocyst. AB - Solid-pseudopapillary tumor of the pancreas is a very rare low-grade malignancy, predominantly occurring in adolescent girls and young women. Accurate diagnosis of this unusual tumor is important because the prognosis after surgical resection is excellent. We report a rare case of solid-pseudopapillary tumor of the pancreas in a 22-year-old woman that was misdiagnosed as a pancreatic pseudocyst on abdominal CT scan. This case emphasizes the importance of biopsying the pseudocyst wall at the time of drainage procedure if misdiagnosis is to be avoided. CT scan findings alone can not reliably rule out malignant cystic lesions of the pancreas. PMID- 12132749 TI - The diagnostic dilemma of diaphragm injury. AB - Prompt diagnosis of acute traumatic injury to the diaphragm remains a challenge when the admission chest X-ray is unrevealing and immediate laparotomy or thoracotomy is not indicated. Diagnostic delay may contribute to significant morbidity and mortality. A retrospective review of our 15-year experience with diaphragm injury (DI) revealed 13 patients (nine male/four female; mean age, 40 +/- 34 years) who sustained injuries to the left (77%) and right (23%) diaphragm respectively as a result of motor vehicle crashes (MVCs) (69%), penetrating trauma (30%), and pedestrian-versus-car accidents (1%). Nine (69%) patients with timely diagnosis of DI underwent laparotomy for suggestive chest X-rays or other indications for immediate exploration. Four (31%) patients sustaining blunt trauma had DI missed on initial evaluation; all patients had initial radiographic evaluations of the chest and abdomen which ascribed abnormalities to intrathoracic pathology. In the one-day delay the diagnosis (right sided) was made at exploratory laparotomy for persistent abdominal pain. This 74-year-old patient, who also had sustained a duodenal injury, succumbed to sepsis. In the 17 day delay the patient had two chest CT scans and multiple bronchoscopies yet failed to wean from the ventilator before exploratory laparotomy which revealed the diagnosis. The third patient sustained multiple injuries after a MVC and underwent multiple imaging studies and back stabilization before discharge. Ten years later, after multiple negative gastrointestinal workups for abdominal pain a contrast study finally diagnosed herniated transverse colon in the left chest. This patient underwent successful repair via laparotomy. The fourth delayed diagnosis was made in a 72-year-old women who had been involved in an MVC 8 years earlier and had sustained multiple back fractures. She is scheduled for exploration in the near future. DI particulary after blunt trauma and on the right side may be missed in the absence of other indications for immediate surgery because radiographic abnormalities of the diaphragm particularly on the right are often attributed to thoracic pathology or may be absent initially. A high index of suspicion for DI may help lead to an earlier diagnosis especially when the patient's clinical condition fails to improve. PMID- 12132750 TI - Internal hernia formation after laparoscopic Roux-en-Y gastric bypass for morbid obesity. AB - There is mounting concern that internal hernia formation after laparoscopic Roux en-Y gastric bypass (LRYGB) for morbid obesity remains unrecognized until complications develop. In this report we present our experience with this complication. Out of 100 patients who underwent LRYGB we identified five patients who were diagnosed with postoperative internal hernia formation. The medical records and operative details of these patients were reviewed. Of the five patients four were female and the average age was 36 years (range 30-43). All Roux limbs were placed in a retrocolic position. The average time interval to presentation was 104 days (range 4-305). All patients had abdominal pain and four patients experienced vomiting. One patient had obstipation. Only one patient had fever (38.1 degrees C) and the highest white cell count was 14,500. The average loss in body-mass index was 5.21 kg/m2 (range 2.5-14.8). Plain abdominal films revealed dilated bowel in the upper abdomen in three patients. Contrast bowel series was diagnostic in only one patient. One patient had a CT scan, which was diagnostic of small bowel obstruction. All patients underwent operative reduction of the internal hernia; two of these were completed laparoscopically. All hernias had occurred at the mesocolic window and were caused by sutures that had pulled through tissue at the dorsal and lateral aspect of the initial repair. One patient had a nonviable segment of small bowel. There were no deaths. Patients who undergo LRYGB are at a 5 per cent risk for developing small bowel obstruction secondary to internal hernia formation at the mesocolic window. Clinical evaluation and traditional study modalities may not be effective diagnostic tools. A high index of suspicion and low threshold to explore these patients may be the best way to avoid serious sequelae. Modification of operative techniques may reduce the occurrence of internal hernia formation. PMID- 12132751 TI - Elevated admission white blood cell count in pregnant trauma patients: an indicator of ongoing placental abruption. AB - Placental abruption (PA) is a common cause of fetal demise in pregnant trauma patients. Diagnosis of PA may be difficult, and multiple diagnostic clues are currently used including uterine/abdominal pain and tenderness, bleeding, maternal hemodynamic instability, and evidence of PA by ultrasonography or other fetal monitoring. Although diagnosis may be problematic fetal and maternal survival are dependent on early diagnosis and intervention. The purpose of this study is to determine predictive factors present at admission associated with PA in trauma victims. Records of all pregnant trauma patients admitted to Wake Forest University Baptist Medical Center over a 5-year period were reviewed for injury characteristics and outcome. Inclusion criteria included a confirmed pregnancy and fetal disposition. Specific admission parameters evaluated included temperature, heart rate, systolic blood pressure, partial pressure of CO2 in arterial blood, total white blood cell count (WBC) and differential, hematocrit, base deficit, and lactic acid. PA is defined as a spontaneous abortion in the first trimester or abruptio placenta in the second or third trimester. Between April 1, 1996 and October 30, 2000, 30 patients met study criteria. Six of 30 patients were found to have PA (20%). Of the studied parameters WBC was significantly elevated in PA patients (27 +/- 4.6 vs 17 +/- 7.8 WBC x 10(3)/mm3; P = 0.005) as was band count (10 +/- 6.6% vs 4 +/- 6.1%; P = 0.03). Hematocrit was lower in the PA group (27 +/- 4.3% vs 32 +/- 5.4%; P = 0.04). Within this group of variables which differed on univariate analysis, WBC was the best discriminator between patients with and without PA (sensitivity 100%, specificity 79%, negative predictive value 100%, and positive predictive value 54%). In pregnant trauma patients WBC >20,000/mm3 on admission should raise suspicion of the possibility of PA, and close monitoring is warranted. Conversely WBC <20,000/mm3 rules out PA in the pregnant trauma patient (negative predictive value of 100%). PMID- 12132752 TI - Use of a human patient simulator for the advanced trauma life support course. AB - A highly anticipated and rewarding component of the Advanced Trauma Life Support (ATLS) program is the surgical skill station. Logistic, societal, and economic issues have resulted in development of human patient simulators (HPSs) as an alternative to the animal model. We studied initial student reaction to a simulator designed for this skill station. Fourteen participants in an ATLS Provider course completed the standard surgical skill stations and an experimental station using the Simulab Trauma Man HPS. After completion of the stations the students were asked to complete a 13-point satisfaction survey using a modified Likert scale (1 = strongly negative/dissatisfied, 5 = strongly positive/satisfied). Overall response was favorable. Students found the HPS to be superior to the animal model in teaching surgical airways [mean 3.64; standard deviation (SD) 0.93] and for management of pneumothorax (mean 3.86; SD 0.77). The students felt the HPS would be useful in ATLS and should be included as an option in training (mean 4.07; SD 0.92). Preliminary experience with an interactive HPS to teach the ATLS surgical skill station is well received by students when compared with standard methods. This strong acceptance supports inclusion of simulators in teaching ATLS skills. PMID- 12132753 TI - Re: Effectiveness of the ultrasonic coagulating shears, LigaSure vessel sealer, and surgical clip application in biliary surgery: a comparative analysis. PMID- 12132754 TI - Object pose: perceiving 3-d shape as sticks and slabs. AB - Estimating the pose (three-dimensional orientation) of objects is an important aspect of 3-D shape perception. We studied the ability of observers to match the pose of the principal axes of an object with the pose of a cross consisting of three perpendicular axes. For objects, we used a long and a flat spheroid and eight symmetric objects with aspect ratios of dimensions of approximately 4:2:1. Stimulus cues were the contour and stereo for the spheroids, and contour, stereo, and shading for the symmetric objects. In addition, the spheroids were shown with or without surface texture and with or without active motion. Results show that observers can perform the task with standard deviations of a few degrees, though biases could be as large as 30 degrees. The results can be naturally decomposed in viewer-centered coordinates, and it turns out that the estimation of orientation in the frontoparallel plane (tilt) is more precise than estimation of orientation in depth (slant, roll). A comparison of long and flat spheroids shows that sticks lead to better performance than do slabs. This can even be the case within the same object; the pose of the stick-like aspect is seen with more precision than is the pose of the slab-like aspect. The largest biases occurred when the spheroids were displayed with the binocular contour as the only cue. We can explain these biases by assuming that subjects' settings are influenced by the orientation of the rim. PMID- 12132755 TI - Object recognition is mediated by extraretinal information. AB - Many previous studies of object recognition have found view-dependent recognition performance when view changes are produced by rotating objects relative to a stationary viewing position. However, the assumption that an object rotation is equivalent to an observer viewpoint change ignores the potential contribution of extraretinal information that accompanies observer movement. In four experiments, we investigated the role of extraretinal information on real-world object recognition. As in previous studies focusing on the recognition of spatial layouts across view changes, observers performed better in an old/new object recognition task when view changes were caused by viewer movement than when they were caused by object rotation. This difference between viewpoint and orientation changes was due not to the visual background, but to the extraretinal information available during real observer movements. Models of object recognition need to consider other information available to an observer in addition to the retinal projection in order to fully understand object recognition in the real world. PMID- 12132756 TI - Perception of two-body center of mass. AB - Participants estimated the perceptual center of mass between two horizontally oriented black dots varying in size and distance. Experiment 1 showed that estimates, measured as distance from the larger dot's center, decreased with an increase in size ratio between the dots and a decrease in the distance between them, as predicted by the physical center-of-mass equation. The results were replicated and extended in further experiments with different ratios and distances. In all experiments, the true center was consistently overestimated, either because the stimuli were perceived in a low dimensionality or because of a hesitancy to place estimates near the larger dot's edge. In Experiment 4, the center was deliberately placed near this edge for one- and two-dimensional solutions. Participants still overestimated, indicating an "edge effect" as responsible. PMID- 12132757 TI - Prioritization in visual search: visual marking is not dependent on a mnemonic search. AB - Visual marking (VM) refers to our ability to completely exclude old items from search when new stimuli are presented in our visual field. We examined whether this ability reflects an attentional scan of the old items, possibly allowing observers to apply inhibition of return or maintain a memory representation of already seen locations. In four experiments, we compared performance in two search conditions. In the double-search (DS) condition, we required participants to pay attention to a first set of items by having them search for a target within the set. Subsequently, they had to search a second set while the old items remained in the field. In the VM condition, the participants expected the target only to be in the second (new) set. Selection of new items in the DS condition was relatively poor and was always worse than would be expected if only the new stimuli had been searched. In contrast, selection of the new items in the VM condition was good and was equal to what would be expected if there had been an exclusive search of the new stimuli. These results were not altered when differences in Set 1 difficulty, task switching, and response generation were controlled for. We conclude that the mechanism of VM is distinct from mnemonic and/or serial inhibition-of-return processes as involved in search, although we also discuss possible links to more global and flexible inhibition-of-return processes not necessarily related to search. PMID- 12132758 TI - Response scale transfer for visual speed. AB - By reversing the presentation order and frequency of stimuli between two series of trials, we studied how the category scale for visual speed is transferred across stimulus contexts. Participants judged five stimulus speeds, using three categories (slow, moderate, and fast). In Experiment 1, mainly frequent speeds (either low or high) occurred on the initial trials. This manipulation produced divergent preshift ratings for identical stimuli. Although subsequent reversal of stimulus context resulted in a reversal of scales, the adjustment was incomplete: The postshift ratings did not match the comparable preshift ones. In Experiment 2, mainly infrequent speeds were presented initially. Now the preshift ratings coincided, but higher postshift ratings occurred with the frequent high-speed rather than with the frequent low-speed stimuli. We conclude that with transfer into a new context, the spontaneous adjustment of response scale is determined (1) by the preshift primacy and the postshift frequency effects and (2) by the preshift frequency effect extended to the postshift trials. PMID- 12132759 TI - What makes a categorization task difficult? AB - To understand why some categorization tasks are more difficult than others, we consider five factors that may affect human performance--namely, covariance complexity, optimal accuracy level with and without internal noise, orientation of the optimal categorization rule, and class separability. We argue that covariance complexity, an information-theoretic measure of complexity, is an excellent predictor of task difficulty. We present an experiment that consists of five conditions using a simulated medical decision-making task In the task human observers view hundreds of hypothetical patient profiles and classify each profile into Disease Category A or B. Each profile is a continuous-valued, three dimensional stimulus consisting of three vertical bars, where each bar height represents the result of a medical test. Across the five conditions, covariance complexity was systematically manipulated. Results indicate that variation in performance is largely a function of covariance complexity and partly a function of internal noise. The remaining three factors do not explain performance results. We present a challenge to categorization theorists to design models that account for human performance as predicted by covariance complexity. PMID- 12132760 TI - Generalizing a neuropsychological model of visual categorization to auditory categorization of vowels. AB - Twelve male listeners categorized 54 synthetic vowel stimuli that varied in second and third formant frequency on a Bark scale into the American English vowel categories [see text]. A neuropsychologically plausible model of categorization in the visual domain, the Striatal Pattern Classifier (SPC; Ashby & Waldron, 1999), is generalized to the auditory domain and applied separately to the data from each observer. Performance of the SPC is compared with that of the successful Normal A Posteriori Probability model (NAPP; Nearey, 1990; Nearey & Hogan, 1986) of auditory categorization. A version of the SPC that assumed piece wise linear response region partitions provided a better account of the data than the SPC that assumed linear partitions, and was indistinguishable from a version that assumed quadratic response region partitions. A version of the NAPP model that assumed nonlinear response regions was superior to the NAPP model with linear partitions. The best fitting SPC provided a good account of each observer's data but was outperformed by the best fitting NAPP model. Implications for bridging the gap between the domains of visual and auditory categorization are discussed. PMID- 12132761 TI - Top-down gain control in the auditory system: evidence from identification and discrimination experiments. AB - The influence of intensity range in auditory identification and intensity discrimination experiments is well documented and is usually attributed to nonsensory factors. Recent studies, however, have suggested that the stimulus range effect might be sensory in origin. To test this notion, in one set of experiments, we had listeners identify the individual tones in a set. One baseline condition consisted of identifying four 1-kHz, low-intensity tones; the other consisted of identifying four 1-kHz, high-intensity tones. In the experimental conditions, these baseline tone sets were augmented by adding a fifth tone at either 1 or 5 kHz. Added 5-kHz tones had little effect on identification accuracy for the four baseline tones. When an added 1-kHz tone differed substantially in intensity from the four baseline tones, it adversely affected performance, with the addition of a high-intensity tone to a set of low intensity tones having a more deleterious effect than the addition of a low intensity tone to a set of high-intensity tones. These and further results, obtained in an exploration of this asymmetrical range effect in a third identification experiment and in two intensity-discrimination experiments, were consistent with the notion of a nonlinear amplifier under top-down control whose functions include protection against sensory overload from loud sounds. The identification data were well described by a signal-detection model using equal variance Laplace distributions instead of the usual Gaussian distributions. PMID- 12132762 TI - Tactile "capture" of audition. AB - Previous research has demonstrated that the localization of auditory or tactile stimuli can be biased by the simultaneous presentation of a visual stimulus from a different spatial position. We investigated whether auditory localization judgments could also be affected by the presentation of spatially displaced tactile stimuli, using a procedure designed to reveal perceptual interactions across modalities. Participants made left-right discrimination responses regarding the perceived location of sounds, which were presented either in isolation or together with tactile stimulation to the fingertips. The results demonstrate that the apparent location of a sound can be biased toward tactile stimulation when it is synchronous, but not when it is asynchronous, with the auditory event. Directing attention to the tactile modality did not increase the bias of sound localization toward synchronous tactile stimulation. These results provide the first demonstration of the tactile capture of audition. PMID- 12132763 TI - Goodness ratings of melodic openings and closures. AB - The purpose of the present study was to map the suitability of melodic intervals for opening and closing a melody. Listeners (n = 13), belonging to two groups with different levels of expertise, rated the 25 within-octave intervals (unison plus the 12 nonunison intervals in ascending and in descending directions), for their suitability as openings; another, similarly subdivided group of listeners (n = 12) rated the intervals as candidates for closure. Both data sets were modeled by means of multiple regression. For openings, a three-factorial model (intervallic size, direction, and implicit harmony) provided a satisfactory description of the data; for closures, an extra factor (intervallic gravity) had to be added to the model. Differences involving level of expertise were established-notably, the importance of harmony for the experts, which played only a subordinate role in the responses of the nonexperts. The two sets of data were contrasted with those of closely related perceptual studies of openings and closures. In addition, the findings were discussed in relation to their contribution to insight into overlapping, a syntactical construction in which tones serve the double function of closing a musical phrase and opening the next one. PMID- 12132764 TI - Effect of harmonic relatedness on the detection of temporal asynchronies. AB - Speeded intonation judgments of a target chord are facilitated when the chord is preceded by a harmonically related prime chord. The present study extends harmonic priming to temporal asynchrony judgments. In both tasks, the normative target chords (consonant, synchronous) are processed more quickly and accurately after a harmonically related prime than after a harmonically unrelated prime. However, the influence of harmonic context on sensitivity (d') differs between the two tasks: d' was higher in the related context for intonation judgments but was higher in the unrelated context for asynchrony judgments. A neural net model of tonal knowledge activation provides an explanatory framework for both the facilitation in the related contexts and the sensitivity differences between the tasks. PMID- 12132765 TI - The impact of spatiotemporal sampling on time-to-contact judgments. AB - When motion in the frontoparallel plane is temporally sampled, it is often perceived to be slower than its continuous counterpart. This finding stands in contrast to humans' ability to extrapolate and anticipate constant-velocity motion. We investigated whether this sampling bias generalizes to motion in the sagittal plane (i.e., objects approaching the observer). We employed a paradigm in which observers judged the arrival time of an oncoming object. We found detrimental effects of time sampling on both perceived time to contact and time to passage. Observers systematically overestimated the time it would take a frontally approaching object to intersect their eye plane. To rule out artifacts inherent in computer simulation, we replicated the experiment, using real objects. The bias persisted and proved to be robust across a large range of temporal and spatial variations. Energy and pooling mechanisms are discussed in an attempt to understand the effect. PMID- 12132766 TI - Stimulus-based lexical distinctiveness as a general word-recognition mechanism. AB - Word recognition is generally assumed to be achieved via competition in the mental lexicon between phonetically similar word forms. However, this process has so far been examined only in the context of auditory phonetic similarity. In the present study, we investigated whether the influence of word-form similarity on word recognition holds in the visual modality and with the patterns of visual phonetic similarity. Deaf and hearing participants identified isolated spoken words presented visually on a video monitor. On the basis of computational modeling of the lexicon from visual confusion matrices of visual speech syllables, words were chosen to vary in visual phonetic distinctiveness, ranging from visually unambiguous (lexical equivalence class [LEC] size of 1) to highly confusable (LEC size greater than 10). Identification accuracy was found to be highly related to the word LEC size and frequency of occurrence in English. Deaf and hearing participants did not differ in their sensitivity to word LEC size and frequency. The results indicate that visual spoken word recognition shows strong similarities with its auditory counterpart in that the same dependencies on lexical similarity and word frequency are found to influence visual speech recognition accuracy. In particular, the results suggest that stimulus-based lexical distinctiveness is a valid construct to describe the underlying machinery of both visual and auditory spoken word recognition. PMID- 12132767 TI - The locus of "memory displacement" is at least partially perceptual: effects of velocity, expectation, friction, memory averaging, and weight. AB - When observers are asked to localize the final position of a moving target, the judged position is usually displaced from the actual position. It has been suggested that mental processes derived from a number of invariant and noninvariant principles produce the mislocalization in memory. In this study, the effects of velocity, expectation, friction, memory averaging, and weight were reconsidered, and evidence was accumulated that supports the alternative view that the distortions arise to a large degree at a perceptual level. Effects of velocity and expectation were present when observers still perceived a persisting image of the target. It is suggested that the active reorienting of the perceptual organs explains the distortions. Furthermore, distortions of the perceived center of a visible stimulus may explain effects that have previously been attributed to memory averaging and mental analogues of weight. Thus, the locus of memory displacement is at least partially perceptual. PMID- 12132768 TI - Burkholderia cepacia complex infection in patients with cystic fibrosis. AB - The word 'complex' has several meanings and synonyms such as composite, obsession, heterogeneous, mixed and network, can all be used in its place. Our obsession with bacteria from the Burkholderia cepacia complex started in the early 1990s. In less than 10 years, we have seen the status of this bacterium move from: (i) a lesser known pseudomonad opportunist pathogen, (ii) to devastating infections transmitted between patients with cystic fibrosis (CF), (iii) through divisions into several new species, and (iv) now on towards one of the largest gram-negative genome sequencing projects. For microbiologists, hospital infection control officers, caregivers, and most of all the CF community, the changes in our understanding of the taxonomy, epidemiology and pathogenesis of the bacterium 'B. cepacia' are complex. PMID- 12132769 TI - Characterisation of an acapsular mutant of Burkholderia pseudomallei identified by signature tagged mutagenesis. AB - A Burkholderia pseudomallei mutant which was attenuated in a mouse model of melioidosis was identified by a signature tagged mutagenesis approach. The transposon was shown to be inserted into a gene within the capsular biosynthetic operon. Compared with the wild-type bacteria this mutant demonstrated a 10(5) fold increase in the median lethal dose in a mouse model and it did not react with a monoclonal antibody against high mol. wt polysaccharide of B. pseudomallei. To determine the kinetics of infection, mice were dosed intraperitoneally (i.p.) and intravenously (i.v.) with mutant and wild-type bacteria. After i.p challenge, the number of mutant bacteria in the peritoneal cavity declined, whereas wild-type bacteria proliferated. When administered by the i.v. route, the mutant was able to cause disease but the time to death was increased compared with the wild type. Mice were dosed with the mutant and subsequently challenged with wild-type B. pseudomallei, but the mutant failed to induce a protective immune response. PMID- 12132770 TI - Virulence properties of Escherichia coli faecal strains isolated in Poland from healthy children and strains belonging to serogroups O18, O26, O44, O86, O126 and O127 isolated from children with diarrhoea. AB - Four hundred and twenty-seven Escherichia coli isolates from 427 cases of infantile diarrhoea in Poland, belonging to serogroups O18, O26, O44, O86, O126 and O127 and 150 E. coli isolates from 52 healthy children were examined for selected virulence properties. The presence of the plasmid pAA, a plasmid encoding enterohaemolysin, the genes encoding intimin (eae), bundle-forming pili (bfp), Shiga toxins I and II (stxI, stxII) and cytotoxic necrotising factor types 1 and 2 (cnfl, cnf2) was investigated by PCR. Adhesion to HEp-2 cell monolayers was also tested and selected strains were investigated for the presence of P fimbriae and haemolytic activity. Typical enteropathogenic E. coli isolates (typical EPEC, strains possessing eae and bfp, but not stx) were not found. The particular classes of E. coli among 427 isolates from ill children were: atypical EPEC (eae+ bfp, stx-), 21.3%; Shiga toxin-producing E. coli (STEC), 0.7%; diffusely adherent E. coli (DAEC), 4%; enteroaggregative E. coli (EAEC), 16.9%; necrotoxic E. coli type 1 (NTEC1), 0.2%; and cell-detaching E. coli (CDEC), 29%. With the exception of STEC, all the above classes of E. coli were found among the isolates from healthy children which comprised: atypical EPEC 8.0%, DAEC 6.7%, EAEC 17.3%, NTEC1 14.0% and CDEC 40.0%. Cell detachment (CD) was significantly associated with 3-h haemolytic activity. There was also strong correlation between haemolytic activity (Hly) and the presence of P-fimbriae. No correlation was found between the presence of the cnf1 gene and CD, Hly or P-fimbriae. PMID- 12132771 TI - Distribution of fim3 and flaA TTGE sequence types amongst isolates of Bordetella bronchiseptica from different host animals. AB - Isolates of Bordetella bronchiseptica associated with different animal hosts were analysed by fim3 and flaA genotyping by temporal temperature gradient gel electrophoresis (TTGE). All the isolates from cats (n = 138), dogs (n = 42) and pigs (n = 13) could be assigned to one of two fim3 and one of three flaA TTGE sequence types, respectively. Two avian isolates and a marmoset isolate exhibited novel fim3 sequence types. Dominant but different TTGE sequence types were apparent in isolates from dogs and pigs for both fim3 (87.5% and 100%, respectively, of isolates were the dominant type) and flaA (95% and 92%, respectively, of isolates were the dominant type). There was a wider distribution of TTGE sequence types amongst cat isolates. As both fimbriae and flagella have been implicated in adherence of bordetellae to host cells, sequence variations in fimbrial proteins and FlaA may have a role to play in host preferences. PMID- 12132772 TI - Transferable tetracycline resistance in Listeria monocytogenes from food in Italy. AB - Mechanisms of tetracycline resistance were investigated in two recent Listeria monocytogenes isolates from food, with L. innocua 52P tet(r) as a control. Tetracycline resistance was transferred conjugatively from all three strains to L. ivanovii and from one isolate and the control to Enterococcus faecalis. Molecular analysis demonstrated a chromosomal location for the tet determinant, which was identified as tetM in all cases. These studies are the first to show that L. monocytogenes from food could be a source of tetracycline resistance genes able to spread to other micro-organisms. PMID- 12132773 TI - Inhibition of listeriolysin O and phosphatidylcholine-specific production in Listeria monocytogenes by subinhibitory concentrations of plant essential oils. AB - Successful infection by Listeria monocytogenes is dependent upon a range of bacterial extracellular proteins including a cytolysin termed listeriolysin O and phosphatidylcholine-specific phospholipase C. Five plant essential oils--bay, clove, cinnamon, nutmeg and thyme--significantly reduced the production of listeriolysin O by L. monocytogenes. The greatest change was observed after culture with oil of thyme, which reduced haemolysis to 52.1 haemolytic units (HU)/ml compared with 99.8 HU/ml observed with the control. Oil of clove was the only oil that also significantly reduced phosphatidylcholine-specific phospholipase C activity. These changes were observed despite the oils causing no change to the final bacterial concentration or total extracellular protein concentration. PMID- 12132774 TI - Phenotypic and genotypic properties of the genus Hafnia. AB - The present study characterised 73 Hafnia alvei isolates and five Escherichia isolates (originally identified as H. alvei) isolated from cases of diarrhoeal disease by the International Centre for Diarrhoeal Disease Research Branch (ICDDRB) in Bangladesh. Based upon the hydrolysis of arbutin and aesculin and the fermentation of salicin and D-arabinose, four distinct biotypes could be recognised among the 73 H. alvei isolates tested; biotype 1 (D-(-)-arabinose positive only) accounted for 75% of all isolates analysed. Hydrolysis of aglycone compounds such as arbutin, salicin and aesculin appeared to be associated with expression of beta-glucosidase activity. ICDDRB isolates, when compared with type or reference strains of H. alvei, were shown not to belong to the genus Hafnia based upon resistance to Hafnia-specific bacteriophage 1672, possession of the phoE gene, expression of glutamate decarboxylase activity and significant 16S rDNA sequence divergence (approximately 8%) from the type strain, ATCC 13337T. True H. alvei strains, implicated in outbreaks of diarrhoeal disease in Canada, lacked the eaeA gene in contrast to ICDDRB isolates. Twenty-two H. alvei isolates were selected for further study. Based upon partial 16S rDNA sequencing, these 22 isolates fell into two genomic groups (genomospecies), identical to DNA groups previously established by DNA hybridisation studies. Markers such as motility, biotype, or enzymic or carbohydrate fermentation patterns did not correlate totally with DNA grouping, although malonate utilisation appeared to be the single best discriminatory phenotype. The results indicate that the genus Hafnia is heterogeneous and there do not appear to be any laboratory data available specifically linking these organisms to gastro-enteritis. PMID- 12132775 TI - The role of macrophages in the induction of murine immune response to Actinobacillus actinomycetemcomitans. AB - The aim of this study was to determine the role of macrophages in the Actinobacillus actinomycetemcomitans-induced murine immune response. BALB/c mice were given carrageenan solution by intraperitoneal injection before immunisation with heat-killed A. actinomycetemcomitans. Mice immunised with antigens and phosphate-buffered saline served as positive and negative controls, respectively. One week after the last immunisation, the delayed-type hypersensitivity (DTH) response was assessed by measurement of footpad swelling. Serum IgG and IgM anti A. actinomycetemcomitans antibody levels and culture supernate levels of interferon (IFN)-gamma were determined by ELISA. The diameter of abscess formation was determined every 5 days. Sham-immunised spleen cells were transferred to carrageenan-untreated recipients (groups A and B) and to carrageenan-treated recipients (group D). Antigen-immunised spleen cells were transferred to carrageenan-untreated (group C) and carrageenan-treated (group E) recipients. The carrageenan-treated recipients in groups F and G received macrophages from antigen- and sham-immunised mice respectively. All mice except those in group A were immunised with antigen 24 h after cell transfer. After 1 week, a partial suppression of DTH response, reduced levels of IFN-gamma, serum IgG and IgM anti-A. actinomycetemcomitans antibodies and delayed healing were seen in carrageenan-treated mice when compared with the positive control. The immune response to A. actinomycetemcomitans in groups A, B and D was lower than that in groups C and E. Healing of the lesion in the former groups was also delayed when compared with the latter groups. The immune response and the healing of the lesion could be partially restored in carrageenan-treated mice that received antigen-pulsed macrophages (group F) but not in those that received naive macrophages (group G). These results suggest that macrophages play a partial role in the induction of the murine immune response to A. actinomycetemcomitans. PMID- 12132776 TI - Antibodies to streptococcal inhibitor of complement function and M peptides in a post-streptococcal glomerulonephritis endemic region of Australia. AB - Post-streptococcal glomerulonephritis (PSGN) is an immune-mediated disease in which an immune complex containing a streptococcal antigen are deposited in affected glomeruli. Strains of only some M types are known to be associated with PSGN. A secretory protein called SIC inhibits complement function. Whereas all M1 and M57 strains express closely related SIC (CRS), all M12 and M55 strains express distantly related SIC (DRS) proteins. Strains belonging to these four M types are historically associated with PSGN. This study used ELISA to analyse 112 sera from individuals with a recorded history of PSGN and 86 sera from individuals who had no such recorded history, all from a PSGN endemic region in tropical Australia. Antibody reactions to CRS, DRS and peptides corresponding to the N-termini of M1, M5, M12, M49, M55 and M57 antigens were assessed. A large proportion of the population showed reactions to each of these antigens and there was no correlation between CRS seropositivity and antibodies to CRS-positive M types. Likewise there was no correlation between DRS seropositivity and antibodies to DRS-positive M types. Interestingly, in this community endemic for PSGN a significantly higher proportion of DRS seropositive subjects had a recorded history of PSGN than did DRS seronegative subjects. DRS may have a predictive value for PSGN diagnosis or a role in PSGN pathogenesis. PMID- 12132777 TI - Typing of human isolates of Streptococcus agalactiae (group B streptococcus, GBS) strains from Zimbabwe. AB - Serotyping and genotyping are important tools in epidemiological studies of group B streptococcal (GBS) infections, which are important diseases in man, particularly in newborns. In the present study, 241 GBS isolates from Zimbabwe, comprising 124 carrier isolates from pregnant women and 117 isolates from patients hospitalised for various diseases, were serotyped. Antibodies specific for the capsular polysaccharide antigens (CPAs) Ia, Ib and II-V and antibodies specific for the surface-localised proteins, c(alpha), c(beta), R1, R3 and R4 were used for serotyping. Strains of the CPA types Ia (17%), III (47.7%) and V (23.2%) predominated. Of the various protein antigens, c(alpha) and R4 were expressed with highest frequency, c(alpha) by 100% of the CPA type Ia strains and R4 by 92% of the CPA type III strains. The R3 protein occurred frequently (24%), especially in type V strains (84%). A total of 25 serovariants was detected in the strain collection with the variants Ia/c(alpha) (16%), III/R4 (43.5%) and V/c(alpha), R3 (14.1%) occurring with the highest frequency. Serotype and subtype distribution of the carrier isolates were essentially similar to those of the disease-associated isolates. Genomic heterogeneity was demonstrated by pulsed field gel electrophoresis of type III/R4 and type V/c(alpha), R3 isolates, but to a much lesser extent than recorded with Norwegian strains. These results demonstrate that many variants of GBS occur in the Zimbabwean population. The data obtained may assist in the formulation of a possible future GBS vaccine for Zimbabwe and perhaps for other African countries. PMID- 12132778 TI - A prospective study on fungal infection in children with cancer. AB - A prospective study was conducted in 1999 at the National Cancer Institute, Cairo University, to estimate the incidence, morbidity and mortality of fungal infections along with the evaluation of risk factors influencing outcome of infections among paediatric cancer patients. Of 1917 infectious episodes, the fungal infection rate as documented both clinically and microbiologically was 3.7% (70 cases). Fungal pathogens isolated were yeasts in 55 patients (78.6%) and moulds in 15 patients (21.11%). Among yeasts, Candida parapsilosis was the commonest, followed by C. tropicalis. Pneumonia was the most common fungal infection (n = 25, 35.7%), followed by fungaemia (n = 18, 25.7%). The overall mortality rate was 40% (n = 28), with an infection-related mortality of 28.5% (n = 20). Risk factors that accompanied mortality were relapsing or recurrent disease, profound neutropenia, ADE (Ara-C, daunorubocin and etoposide) protocol of chemotherapy, C. tropicalis isolated and fungaemia as a site of infection. Early use of empirical antifungal therapy (day 4) was not associated with a better outcome. In the light of the poor outcome of patients with fungaemia and fungal pneumonia, every effort should be made to prevent these infections in paediatric cancer patients. PMID- 12132779 TI - A simple 'paper smear' method for dry collection, transport and storage of cervical cytological specimens for rapid screening of HPV infection by PCR. AB - Human papillomaviruses (HPVs) are major pathogens associated with the development of cancer of the uterine cervix, the most common malignant tumour of women worldwide. Reliable diagnosis of HPV infection, particularly the 'high-risk' types (16/18), may facilitate early identification of 'high-risk' populations for developing cervical cancer and may augment the sensitivity and specificity of primary cervical cancer screening programmes by complementing the conventional Pap test. A simple paper smear method has been developed for dry collection, transport and storage of cervical smears/scrapes at room temperature for subsequent detection of HPV DNA by PCR assay. Imprint biopsies, blood and fine needle aspirates were also collected by this method. The cervical scrapes or other body fluids were smeared (within 0.5-1 cm diameter) and dried on to sterile small slides made of Whatman 3MM filter paper, and stored individually at room temperature or at 4 degrees C. A small piece (2-3 mm) of the paper smear was punched or cut out with a sterile surgical blade, boiled in an eppendorf tube containing 50 microl of distilled water for 5 min and used directly for PCR amplification. The quality and quantity of DNA derived from paper smears and the results of PCR amplifications for HPV type 16, BRCA1 and p53 genes were identical to those obtained from the same samples following collection in PBS, storage (-70 degrees C) and phenol-chloroform-based DNA extraction. DNA was stable in the paper smears for up to a year, whether stored at room temperature or at 4 degrees C. This method is simple, rapid and cost-effective, and can be effectively employed for large-scale population screening, especially for regions where the specimens are to be transported from distant places to the laboratory. PMID- 12132780 TI - Pneumocystis carinii carriage in immunocompromised patients with and without human immunodeficiency virus infection. AB - Eighty-one bronchoalveolar lavage (BAL) specimens obtained from 26 HIV-infected, 45 non-HIV immunosuppressed and 10 immunocompetent patients with primary pulmonary diseases were analysed for the presence of Pneumocystis carinii by staining and by P. carinii 5S rDNA determined by PCR. P. carinii was observed by staining of BAL specimens from HIV-infected patients significantly more frequently than those from immunocompromised hosts without HIV infection (57.7% versus 20.0%, respectively). P. carinii 5S rDNA was detected by PCR assay in seven (26.9%) HIV-infected individuals, which was significantly more frequent than for four (8.9%) immunosuppressed patients without HIV infection, for whom staining was negative. None of these patients developed P. carinii pneumonia (PCP) within the follow-up period. BAL specimens from 10 immunocompetent patients with pulmonary disorders were negative for PCP by both staining and PCR assay. PMID- 12132781 TI - A clinical index predicting mortality with Pseudomonas aeruginosa bacteraemia. AB - The aim of this study was to define risk factors associated with mortality in Pseudomonas aeruginosa bactaeremia and to combine them in a clinical index predicting the risk of death. The study investigated 125 consecutive episodes of P. aeruginosa bacteraemia at this hospital. Crude mortality was 34%, corresponding to 43 patients who died, with 67% of deaths, directly attributable to bacteraemia. A regression logistic model identified five variables that were independently and significantly associated with an increased risk of death: 1) hospitalisation in the intensive care unit; 2) coagulopathy; 3) septic shock; 4) age > or = 65 years; and 5) the clinical condition of the patient. These variables were as recorded at the time that the first positive blood culture was obtained. The sensitivity and specificity of a prediction of death based on the model were 84% and 85%, respectively. An index score, calculated from these variables, divided patients into three groups with increasing likelihood of mortality resulting from P. aeruginosa bacteraemia. PMID- 12132782 TI - Community-acquired Pseudomonas aeruginosa sacro-iliitis in a previously healthy patient. AB - Pyogenic sacro-iliitis is an uncommon osteo-articular infection that occurs usually in immunocompromised patients and is associated with gram-positive cocci. It is very rarely linked with a gram-negative aetiology. The first case of Pseudomonas aeruginosa sacro-iliitis is described, which occurred in a previously healthy young man, without history of prior traumatic events, hospitalisation or chronic underlying disease. PMID- 12132783 TI - Bile acid-mediated hepatocyte apoptosis and cholestatic liver disease. PMID- 12132785 TI - Colorectal cancer. A novel approach to adjuvant chemotherapy with fluoropyrimidines. PMID- 12132784 TI - Safer non-steroidal anti-inflammatory drugs: are cyclo-oxygenase inhibitors or nitric oxide non-steroidal anti-inflammatory drugs the grand finale? PMID- 12132786 TI - Detection of Helicobacter pylori specific DNA in human atheromatous coronary arteries and its association to prior myocardial infarction and unstable angina. AB - BACKGROUND: Chronic infections have been proposed to play a role in the aetiology or progression of atherosclerotic plaques. Increased risk of coronary artery disease has been suggested in patients seropositive for Helicobacter pylori. AIM: To analyse coronary specimens in patients with severe (coronary artery disease) for Helicobacter pylori specific DNA. PATIENTS AND METHODS: Atherosclerotic plaques were obtained in 46 consecutive patients (9 female, 37 male, mean age 62.7+/-9.17 years) during coronary bypass procedures. Serum was analysed for IgG /cagA-antibodies specific for Helicobacter pylori. Polymerase chain reaction and sequence analysis were used to identify bacterial DNA. Coronary artery biopsies from 19 autopsies without coronary artery disease were examined as a control group. RESULTS: Of the 46 coronary artery disease patients, 32 (69.6%) were Helicobacter pylori seropositive. Positive results for Helicobacter pylori DNA showed 18 seropositive and 4 seronegative (with anamnesis of eradication therapy). A total of 22 patients (47.8%) of the coronary artery disease group but none of controls revealed positive DNA. In the coronary artery disease group, a correlation between DNA presence and prior myocardial infarction (p=0.008) and unstable angina (p<0.001) was found. CONCLUSION: Identification of DNA in atherosclerotic plaques of patients with severe coronary artery disease supports the hypothesis that Helicobacter pylori infection may influence the development of atherosclerosis. Our results may indicate an direct involvement of Helicobacter pylori in the progression and instability of plaques in these patients. PMID- 12132787 TI - Gastroprotective effects of amtolmetin guacyl: a new non-steroidal anti inflammatory drug that activates inducible gastric nitric oxide synthase. AB - BACKGROUND: The novel non-steroidal anti-inflammatory drug amtolmetin guacyl has been shown to possess markedly reduced ulcerogenic effects and nitric oxide mediated gastroprotective activity against the damage induced by ethanol in the rat. AIMS: To investigate, in the rat, the role of nitric oxide and of inducible nitric oxide synthase isoform in the protective effect of amtolmetin guacyl against the gastric damage induced by ethanol. METHODS: The effects of amtolmetin guacyl on gastric transmucosal potential difference and on gastric mucosal blood flow were investigated in the anaesthetised rat; myeloperoxidase activity, inducible and endothelial nitric oxide synthase protein content were determined in rat gastric mucosal homogenates. The anti-inflammatory drug tolmetin and the bacterial lipopolysaccharide from Escherichia coli were studied for comparison. RESULTS: In the anaesthetised rat, amtolmetin guacyl, but not tolmetin, reduced by approximately 50% the fall in gastric potential difference and, to a lesser extent, the macroscopic damage induced by ethanol. The effect of amtolmetin guacyl on transmucosal potential difference was prevented by the selective inducible nitric oxide synthase inhibitor 1400W. In amtolmetin guacyl-treated rats, 1400W decreased gastric mucosal blood flow, whereas it was inactive in vehicle- and tolmetin-treated animals. In gastric mucosal homogenates, both amtolmetin guacyl and lipopolysaccharide, but not tolmetin, increased inducible, but not endothelial, nitric oxide synthase protein content, as revealed by Western immunoblotting. CONCLUSIONS: These data confirm that amtolmetin guacyl is a non-steroidal anti-inflammatory agent devoid of gastrolesive properties, that can actually reduce the damaging effects of ethanol through the increase in nitric oxide production, via the inducible isoform of nitric oxide synthase. PMID- 12132788 TI - Infliximab in treatment of Crohn's disease: the Milan experience. AB - BACKGROUND: Efficacy of infliximab in treatment of patients with moderate-to severe refractory and fistulizing Crohn's disease has been shown in controlled clinical trials. Moreover, audit data from North America and North Europe have confirmed efficacy in clinical practice comparable to that in clinical trials. AIM: To report clinical experience using infliximab in treatment of Crohn's disease in Italy, comparing efficacy and safety with those reported in clinical trials and other published series. PATIENTS AND METHODS: The study population comprised 63 patients (31 males and 32 females, median age 33 years) treated with infliximab for refractory/inflammatory (31 patients) and/or fistulizing Crohn's disease (32 patients). All patients received an infusion of infliximab at a dose of 5 mg/kg at weeks 0, 2 and 6. After the first infusion, clinical and laboratory assessments were repeated at weeks 2, 6 and 10. For refractory inflammatory Crohn's disease, clinical remission was defined as a Crohn's Disease Activity Index of < or = 150 at each scheduled visit, clinical response as a reduction in the Crohn's Disease Activity Index score of > or = 70 points in comparison to baseline. For fistulizing Crohn's disease, a complete response was defined as closure of any draining fistulae at week 10. A fistula was defined as closed when it no longer drained despite gentle finger pressure. A partial response was defined as reduction in number, size or drainage of fistulae, at the same visit. RESULTS: According to an intention-to-treat evaluation on the 31 patients with refractory/inflammatory Crohn's disease, at week 2, 42.5% (14 patients) had a clinical response and 31.3% of patients (10 patients) were in clinical remission. At week 10 (4 weeks after the end of third infusion), 80.6% (25 patients) had a clinical response and 71% (22 patients) were in clinical remission and 14/19 (74%) had discontinued steroid treatment. Of the 32 patients with fistulizing Crohn's Disease, 15 (46.9%) had a complete response, 8 (25%) a partial response, and 9 (28.1%) no response at week 10 check-up. The incidence of side-effects was low (16%) and not influenced by concurrent immunomodulatory therapy. CONCLUSION: The present experience with infliximab in clinical practice confirms its efficacy, in particular in inflammatory/refractory Crohn's disease and its safety, at least, in short-term follow-up. PMID- 12132789 TI - Long-term maintenance treatment in ulcerative colitis: a 10-year follow-up. AB - BACKGROUND: With the extensive use of mesalamine, the natural history of ulcerative colitis is probably changed. AIM: To evaluate the relapse rate and the duration of remission in patients with ulcerative colitis on maintenance treatment with mesalamine. PATIENTS AND METHODS: Enrolled in the study were 95 patients divided into 4 groups according to macroscopic location of the disease and treated with the same therapy starting from the date of enrolment. Patients in all 4 groups were followed-up until relapse occurred. The disease activity was evaluated by the Clinical Activity Index and Endoscopic Index. Patients suitable for recruitment showed a Clinical Activity Index and Endoscopic Index lower than 6 and 4, respectively. The patients with ulcerative pancolitis or left-sided colitis were treated with 1.6 g/day while the cases with proctosigmoiditis or proctitis were treated with 5-acetylsalicylic acid enemas 4 g/day Each patient was evaluated with clinical and endoscopic assessment at a 6-month interval. Relapse was defined as an increase in Clinical Activity Index and Endoscopic Index, of more than 6 and 4, respectively. RESULTS: Five patients dropped-out. All enrolled patients showed a clinical and/or endoscopic relapse within 10 years, the majority 2 or 3 years after diagnosis: pancolitis and left-sided colitis within 2-3 years and patients with distal colitis within 9-10 years. CONCLUSIONS: A relapse was observed in most cases within 3 years, and in all recruited patients within a space of ten years. The extent of the disease in the colon is an important prognostic factor, as patients with distal colitis showed a lesser tendency to relapse. PMID- 12132790 TI - Whole gut lavage fluid interleukin-1beta and interleukin-8 in smokers and non smokers with Crohn's disease in clinical remission. AB - INTRODUCTION: Smoking in patients with Crohn's disease is associated with more frequent relapse. The mechanism responsible is unknown but a direct pro inflammatory action on intestinal mucosa has been postulated. Mucosal inflammation in clinically inactive Crohn's disease predicts forthcoming relapse. Whole gut lavage fluid obtained after bowel cleansing with a polyethylene glycol electrolyte solution is an assessment of gut inflammation and immunity. AIM: To assess whether whole gut lavage fluid interleukin-1beta and interleukin-8 differed between smokers and non-smokers with clinically inactive Crohn's disease. METHODS: A total of 34 patients with inactive Crohn's disease (Crohn's disease activity index <150 and whole gut lavage fluid IgG concentration of <10 mg/ml) underwent whole gut lavage with interleukin-1beta and interleukin-8 analysed by enzyme-linked immunosorbent assay. Clinical details and blood markers of inflammation were collected. RESULTS: In this series, 14 patients smoked (10 females, mean age 44.3+/-14.3 years), 20 did not (12 females, mean age 40.7+/ 14.3). Surgical resection was more common in smokers (12/14 vs 8/20, p<0.008). Whole gut lavage fluid IgG was significantly lower in smokers (median 1.5 mg/ml (range 1.0-8.0 mg/ml) vs median 3.5 mg/ml (range 1.0-7.0 mg/ml), p<0.05). Whole gut lavage fluid interleukin-1beta was also lower in smokers [median 14.5 pg/ml (range 2-72 pg/ml) vs 26 pg/ml (range 7-1700 pg/ml)], p<0.03. CONCLUSION: Markers of mucosal inflammation in inactive Crohn's disease are lower in smokers than non smokers. This is against the hypothesis that nicotine exerts a direct pro inflammatory action via interleukin-1beta and interleukin-8. Further research is required to elucidate the exact mechanisms involved. PMID- 12132791 TI - Anti-inflammatory effects of enteral diet components on Crohn's disease-affected tissues in vitro. AB - BACKGROUND: The mechanism of action of elemental diet in Crohn's disease treatment, is unknown. Alteration of bacterial flora, low antigenicity, low fat content and improvement of nutritional status are postulated to play a role in the anti-inflammatory effect of elemental diet. AIM: To determine whether elemental diet or its modifications has a direct anti-inflammatory effect on colonic tissue biopsies in vitro. PATIENTS AND METHODS: Colonic or ileal biopsies from 39 patients with inflammatory bowel disease and control patients were incubated for 24 hours with enteral diets in which nitrogen sources were amino acids as in elemental diet, casein or whey. Tissues were incubated with elemental diet, casein or whey, at dilutions of 1:5, 1:10 or 1:20 in Waymouth's complete medium; a medium control was also included. Tissue viability was assessed by bromodeoxyuridine uptake. Interleukin-1beta, interleukin-1 receptor antagonist and interleukin-10 concentrations in supernatants were measured by immunoassay (enzyme-linked immunosorbent assay). RESULTS: Incubation of tissues from Crohn's disease with elemental diet resulted in an increase in the ratio of interleukin-1 receptor antagonist/interleukin-1beta vs control statistically significant at 1:10 (89.6+/-17 vs 45.7+/-9. 1, p<0.05). Incubation of Crohn's tissue with casein resulted in a significant increase of interleukin-1 receptor antagonist/interleukin-1beta ratio at dilutions 1:20, 1:10 and 1:5 (101.8+/-22.0, p=0.05, 142.8+/-24.6, p<0.05; 109.7+/-25.0, p=0.05). In ulcerative colitis tissue and non-inflamed non-inflammatory bowel disease control tissue, no significant increase in interleukin 1 receptor antagonist/interleukin-1beta ratio was seen after incubation with elemental diet, casein and whey. CONCLUSION: Elemental diet incubation increases anti-inflammatory:proinflammatory cytokine ratio in Crohn's disease and this anti-inflammatory effect is not specifically due to amino acid composition, as diets containing casein have similar anti-inflammatory effects. PMID- 12132792 TI - Floxuridine coupling with lactosaminated human albumin to increase drug efficacy on liver micrometastases. AB - BACKGROUND: Conjugates of nucleoside analogues with galactosyl terminating peptides selectively enter hepatocytes through the asialoglycoprotein receptor. After intracellular release from the carrier, the drugs partly exit from hepatic cells into hepatic blood. AIMS: To establish whether administration of a conjugate of floxuridine with lactosaminated human albumin selectively enhances drug concentrations in hepatic blood. Floxuridine is a fluoropyrimidine active on human colorectal cancer, a tumour which metastasises first to the liver. METHODS: In rats injected with free or conjugated floxuridine, plasma levels of the drug were determined in hepatic veins and in inferior vena cava, in order to measure drug concentrations in hepatic blood and in the systemic circulation, respectively. RESULTS: Ratios between floxuridine levels in hepatic veins and those in systemic circulation were found to be seven times higher in rats injected with the conjugate (p=0.000). CONCLUSIONS: The present results suggest that coupling to lactosaminated albumin might improve the effect of floxuridine in adjuvant chemotherapy of colorectal cancer by exposing the cells of liver micrometastases (nourished by hepatic sinusoids) to enhanced drug concentrations. PMID- 12132793 TI - Inappropriate pemoline therapy leading to acute liver failure and liver transplantation. AB - A 36-year-old female, presenting with jaundice, developed acute liver failure requiring orthotopic liver transplantation. On admission, none of the known causative factors for acute hepatitis, including use of drugs, were found to be present. Several days after hospitalization, the patient admitted taking therapy prescribed by a "non-traditional" physician, that she had been using for several years due to overweight and which had recently been modified with the introduction of pemoline. A considerable body of evidence exists in the medical literature showing that pemoline, which is a central nervous system stimulant, has variable hepatotoxic effects, ranging from a mild transient increase of serum transaminases to liver failure, including some lethal cases. PMID- 12132794 TI - Management of hepatitis C in human immunodeficiency virus-infected patients. AB - Hepatitis C virus-related liver disease and its associated complications are steadily emerging health concerns in persons co-infected with human immunodeficiency virus. The increasing number of liver-related deaths in human immunodeficiency virus-hepatitis C virus co-infected individuals supports the compelling argument for more aggressive treatment in these patients. The safety and efficacy of interferon/ribavirin in human immunodeficiency virus/hepatitis C virus co-infected patients is currently under evaluation. Despite well-documented concern over highly active antiretroviral therapy-associated hepatotoxicity human immunodeficiency virus/hepatitis C virus co-infected patients should be offered antiretroviral therapy. Since management of co-infected patients is complex a multidisciplinary approach is needed in order to facilitate care and help patients to achieve a positive outcome. PMID- 12132795 TI - Recertification: an analysis. PMID- 12132796 TI - Femoral shaft fracture after hip arthroplasty: a system for classification and treatment. AB - Twenty-one consecutive cases of femoral shaft fracture after hip arthroplasty treated at the University of Illinois affiliated hospitals were reviewed. Adequate follow-up and radiographs were available for 19 patients. The length of follow-up after fracture ranged from 2 to 13 years, with a mean of 3.1 years. The time from index procedure to fracture averaged 2.6 years, with a range of 10 days to 11 years. The primary femoral stem was cemented in 11 hips and cementless in 8 hips. Six patients were treated nonoperatively and 13 operatively. Three had fracture fixation with retention of a well-fixed prosthesis and 10 had prosthetic revision. Cortical allograft was used in 5 cases. Sixteen of the 19 patients returned to their prefracture level of function and ambulation. The factors important to treatment are fracture stability, implant stability, and adequacy of bone stock. A classification system based on these factors and recommendations for treatment are proposed. PMID- 12132797 TI - Bipolar arthroplasty for recurrent total hip instability. AB - Bipolar arthroplasty has been reported as a method for correction of recurrent dislocations of total hip replacements. This retrospective review of six patients with multiple dislocations of total hip replacements treated by conversion to simple bipolar hip arthroplasty confirms a 100% success rate in eliminating hip instability during a follow-up period of 2 1/2 to 5 years. However, given the high rate of postoperative discomfort and abnormal gait associated with this procedure, it should be used only when other revision techniques prove to be unsuitable. PMID- 12132798 TI - Biomechanical evaluation of fixation of posterior acetabular wall fractures. AB - Reproducible fractures of the posterior wall of the acetabulum were created in 10 paired hemipelves from fresh human cadavers. Under anterior-to-posterior loading by a Materials Testing System machine, the load to failure of fixation of the acetabular fractures treated with steel pelvic reconstruction plates and steel screws was significantly higher than that of fixation with titanium ribbons and titanium screws. Forces as little as 725 newtons applied directly to the fragment caused fixation failure; even the most secure form of fixation failed when a relatively small force of 2,123 newtons was applied. These results emphasize the importance of appropriate postoperative measures, such as limitation of hip flexion and restricted weightbearing, to minimize the force directed against the posterior wall until the fracture has healed. Secure fixation of fractures of the posterior wall of the acetabulum is critical, since loss of fixation results in instability and joint incongruity, which limit the options for reconstruction. PMID- 12132799 TI - Effects of kyphosis and lordosis on the remaining lumbar vertebral levels within a thoracolumbar fusion: an experimental study of the multisegmental human spine. AB - This study was done to determine the motion of the whole lumbar spine after internal fixation and the effect of kyphosis and lordosis on the remaining vertebral levels. Baseline motion analysis of sagittal, frontal, and transverse planes was done to determine the intact range of motion. Three fusion configurations were tested: neutral position (0 degrees), 4.6 degrees +/- 2.0 kyphosis, and -6.2 degrees +/- 3.6 lordosis. The sagittal and frontal plane relative rotation of the instrumented segments (T12/L2) decreased an average of 74% and 60%, respectively, as compared with intact testing. Sagittal plane motion at the remaining segments increased for all fusion configurations when compared with intact motion and reached statistical significance at the L4/L5 level. No significant differences were found between fusion configurations (ie, fused neutral, kyphosis, and lordosis). PMID- 12132800 TI - Intramedullary fixation of unstable forearm fractures in children. AB - Although most forearm fractures in children are appropriately treated with closed reduction and cast immobilization, certain unstable fractures of the radius and ulna are best treated operatively. We present our technique of using flexible intramedullary fixation to stabilize these fractures. Retrograde fixation of the radius is obtained with a 5/64th or 3/32nd Steinmann pin, and stabilization of the ulna is achieved with a 1/8th inch Rush rod. Complications from this technique are few. The rods are usually removed after fracture union to avoid painful hardware. PMID- 12132802 TI - In pursuit of excellence: personal reflections. AB - I have put together some of the elements that will allow a person to advance and succeed in life. On a personal note, I would not be where I am without the support of my parents, teachers, colleagues, residents, and fellows. Special appreciation goes to my wife, my children, and my office staff. Without their support, this story would be incomplete. Thank all of you for putting your trust in me. PMID- 12132801 TI - Giant cell tumor and the skeletally immature patient. AB - Conventional wisdom suggests that giant cell tumor (GCT) does not occur in the skeletally immature individual; however, we believe that GCTs of bone, though rare, do occur in children. We are reporting the occurrence of GCT of bone in three patients who were skeletally immature at the time of their initial presentation. In our review of the reports since 1954 that document this condition, we were also able to find a total of 318 patients, of whom 130 were skeletally immature at the time of their tumor presentation. From the data compiled, we found a 7.5% incidence of GCT of bone in skeletally immature individuals at a mean age of 10.5 years. Based on our review and the experience with our three patients, we believe the diagnosis of GCT of bone should be considered in the differential diagnosis of a destructive lesion of bone in skeletally immature individuals. Giant cell tumor in the skeletally immature is being reported here to better define its incidence and increase awareness of its occurrence. Management options will also be discussed. PMID- 12132804 TI - Atraumatic floating clavicle and total claviculectomy. AB - We describe a patient with a floating clavicle of atraumatic origin treated by total claviculectomy. Clavicular function and anatomy are summarized relative to complete excision. Other treatment options for panclavicular instability are also discussed. PMID- 12132803 TI - Episodic snapping of the medial head of the triceps due to weightlifting. AB - We describe two patients who had episodic elbow snapping and ulnar nerve dysesthesias only after weightlifting. These symptoms would disappear soon afterward. The episodic nature of their complaints and findings led to misdiagnosis. We documented by repeated clinical examinations and magnetic resonance imaging that the presence of these symptoms correlated directly with the finding of intermittent, activity-related snapping of the medial triceps. In both patients, the symptoms disappeared when the medial portion of the triceps migrated medially but did not dislocate over the medial epicondyle with elbow flexion. Thus, a minor change in the configuration of the medial portion of the triceps (fluid accumulation) in the same individual at different times can cause intermittent dislocation of the medial triceps. Previous papers dealing with patients with snapping of the medial triceps describe symptoms exacerbated by athletic activities, but the constant finding of snapping on sequential examinations. PMID- 12132805 TI - Trapezial resection arthroplasty for osteoarthritis: use of abductor pollicis longus tendoplasty with interpositional material. AB - In this series, 45 patients had 54 trapezial resection arthroplasties for carpometacarpal osteoarthritis, with nine of the patients having bilateral arthroplasties. A major slip of the abductor pollicis longus tendon was used as a sling, with the underlying flexor carpi radialis tendon as stabilizer. Palmaris longus tendon, when present (42 cases), and absorbable gelatin sponge, when palmaris was absent (12 cases), served as interpositional material. Review was done an average of 4.3 years (range, 1 to 11 years) after the procedure. The average space maintenance of resection site was 4.4 mm. This decreased to an average of 3.5 mm on the key pinch stress maneuver. Subjectively, the patients judged their results to be good to excellent in 49 instances (91%) and fair in 5 (9%). PMID- 12132806 TI - Subchondral contusion of the knee caused by axial loading from dashboard impact: detection by magnetic resonance imaging. AB - We studied the occurrence of "bone bruises" of the knee resulting from dashboard impaction and detected by magnetic resonance imaging (MRI). We chose 21 knees of 20 front seat occupants in head-on motor vehicle collisions. To ensure all knees had received a significant axial load, patients selected had ipsilateral posterior hip dislocations and/or posterior wall acetabular fractures. Anteroposterior and lateral knee radiographs were negative for fracture in all cases. T1-weighted axial and sagittal MRI of each knee with a 1.5-T magnetron scanner revealed signal changes consistent with subchondral microfracture or bone bruise in 8 of the 21 knees. Previous cadaveric, animal, and MRI studies have suggested that such changes may be precursors of posttraumatic osteoarthritis. With the increasing incidence of serious lower extremity injury as a result of motor vehicle accidents, these occult injuries may significantly affect individuals and society. PMID- 12132807 TI - Lateral extra-articular knee reconstruction: long-term patient outcome and satisfaction. AB - Fifty-two patients who had an extraarticular reconstruction for anterior cruciate ligament (ACL) insufficiency were interviewed at an average of 11.4 years after their reconstruction. During the follow-up period, 4 patients (8%) had intra articular reconstructions for persistent instability or re-injury, and another 4 (8%) had arthroscopic surgery for meniscal lesions. The mean Lysholm score at follow-up was 76.9. Of the 52 patients, 5 had had subsequent surgery during the follow-up period. In the remaining 47 patients, the results were excellent in 14 (29%), good in 10 (23%), fair in 7 (15%), and poor in 16 (33%). Patients operated on acutely (<6 weeks after injury) had a mean score of 82, whereas patients operated on more than 6 weeks after injury had a mean score of 75. Adding those patients who required subsequent reconstructions for persistent instability (4 knees) and those who obtained fair and poor results yielded an alarming rate of unsatisfactory results (52%). On the basis of these findings, we believe the lateral extraarticular reconstruction is not an acceptable form of treatment for the functionally unstable knee. PMID- 12132808 TI - Imaging of the cervical spine and its role in clinical decision making. AB - Thorough imaging of the cervical spine often requires more than one test. The many available options from which to choose can often lead to redundancy and confusion regarding the best test series. In an effort to make the process of choosing the most effective imaging series more efficient, we review the current literature on cervical imaging and, from the information gathered, construct a diagnostic imaging algorithm for evaluating the cervical spine. PMID- 12132809 TI - Posterior lateral mass plate fixation of the cervical spine. AB - Historically, posterior fixation and fusion have been the most popular method of internally stabilizing the cervical spine after injury. Although techniques such as wiring are effective for most injuries, these methods are inadequate in the absence of intact posterior elements or extension and rotation injuries. We review the indications and techniques for posterolateral mass plate fixation that are useful in these difficult fractures. PMID- 12132810 TI - Nonunion of the fractured clavicle: evaluation, etiology, and treatment. AB - Although often viewed as benign injuries, clavicular fractures can lead to complications, particularly nonunions. The nonunion rate has been reported to be between 0.1% and 15%. Contributing factors to nonunion include severe initial trauma, marked initial displacement and shortening, soft tissue interposition, primary open reduction and internal fixation, refracture, open fracture, polytrauma, and inadequate initial immobilization. A clavicular nonunion is rarely asymptomatic and often results in disability from pain at the site of nonunion, altered shoulder mechanics, or a compression lesion involving the underlying brachial plexus or vascular structures. Treatment options include nonsurgical management, salvage procedures, and reconstructive procedures. The present goal is to obtain union with reconstructive procedures. The fixation methods described range from external fixation to plate and screw osteosynthesis. We prefer open reduction and internal fixation with plates and screws and with intercalary tricorticocancellous grafts to obtain union and restore the clavicle to its normal length. PMID- 12132812 TI - Giving and giving back. PMID- 12132811 TI - Treatment of acetabular fractures. PMID- 12132813 TI - Rupture of the flexor digitorum superficialis: occurrence after tendon repair of an adjacent digit. AB - A patient had a ruptured flexor digitorum sublimis tendon of the long finger in the region of decussation. The injury occurred several weeks after repair of a zone 2 flexor digitorum sublimis and flexor digitorum profundus tendon of the index finger. PMID- 12132814 TI - The decision to salvage or amputate a severely injured limb. AB - The decision to salvage or amputate a severely injured limb is one of the most difficult an orthopaedic surgeon may face. The inclination to undertake heroic measures to save the limb should be tempered by the realization that doing so may lead to repeated hospitalizations, extensive complications, and a poor functional outcome. Several factors must be considered, including objective elements related to the patient's injury and physical condition and subjective considerations related to the patient's psychologic, social, and economic status. We present a framework, in the context of a case study, which may be used in deciding which patients can benefit from early amputation and review five predictive indices for limb salvage. PMID- 12132815 TI - Viscosupplementation treatments. PMID- 12132816 TI - Sever-L'Episcopo tendon transfer in obstetric brachial plexus palsy. AB - Obstetric brachial plexus palsy produces functional and cosmetic impairment. The Sever-L'Episcopo procedure has been successfully used to improve external rotation, primarily in younger patients. Previous studies have shown steady improvement in motion and function for 1 year from the date of surgery, with little additional change beyond this period. This is a retrospective study of 7 patients who have had the procedure at the Shriners Hospital for Children in Tampa, Florida. Functional and cosmetic results have been uniformly excellent at an average follow-up of 1 year 3 months. No complications have occurred. In addition, the degree of improvement seen in our relatively older patient population has not been previously detailed in the literature. PMID- 12132817 TI - Variation of the arthroscopic Mumford procedure for resecting the distal clavicle. AB - Fifty-seven patients had arthroscopic Mumford procedures for acromioclavicular (AC) pain unresponsive to conservative treatment. Thirty-nine of these patients had concomitant rotator cuff repairs. All had significant improvement of distal clavicular pain. Neither the amount nor the completeness of the distal clavicle resection affected the results. One patient with a significant retained lateral clavicular spike required additional surgery for excision. Arthroscopic distal clavicle resection is a safe and effective method of alleviating AC pain. PMID- 12132818 TI - Cemented total hip arthroplasty with Boneloc bone cement. AB - Boneloc cement (WK-345, Biomet Inc, Warsaw, Ind) attempted to improve cement characteristics by reducing exotherm during polymerization, lowering residual monomer and solubility, raising molecular weight, and lowering airborne monomer and aromatic amines. To study the efficacy of this cement, a selected group of 20 patients were prospectively enrolled and followed up after hip arthroplasty. All components were cemented. During the enrollment period, approximately 70 other hip arthroplasties were performed. Clinical evaluation was based on the Harris hip score. Radiographic evaluation was based on assessment of position of the components, subsidence, and/or presence of radiolucencies. Patients had follow-up for an average of 42 months (11 to 58 months); 1 was lost to follow-up. Of these, 7 (35%) had failure at last follow-up. Despite its initial promise, Boneloc cement had an unacceptably high failure rate over a relatively short follow-up period and is not recommended for use. Despite the longevity and odor toxicity problems with conventional bone cement, new cement technologies must be approached with caution. PMID- 12132819 TI - Group B streptococcal prosthetic joint infections. AB - We report 6 cases of group B streptococcal prosthetic joint infections seen in our institution and review 8 previously reported cases. These 14 patients (6 men and 8 women) had an average age of 69. Seven hip joints and 7 knee joints were affected. Only 4 patients had risk factors for prosthetic joint infection (diabetes mellitus in 2, cancer in 1, and myelodysplasia in 1). The average time from surgery to onset of symptoms was approximately 4 years (range, 5 months to 10 years). Pain in the affected joint was the chief complaint. Six patients had bacteremia. Seven patients had known or suspected foci of infection, which were genitourinary tract (1), skin and soft tissue trauma sites (1), gastrointestinal tract (1), and oropharynx (1). Nine patients required prosthesis removal in addition to antibiotic therapy. Two patients had apparent cure of the infection with retention of the prosthesis. Group B streptococcal prosthetic joint infections appear to be a late complication of prosthetic joint replacement surgery. PMID- 12132820 TI - Surgical treatment results in scaphoid nonunion. AB - Fracture of the scaphoid is the most common fracture in the wrist. We retrospectively reviewed 42 cases of established nonunion of the scaphoid that had been treated by four methods: with two Kirschner wires (K wires) and pronator quadratus pedicled bone graft in 5 patients; AO cannulated screw and graft in 8 patients; Herbert screw and graft in 19 patients; and two K wires and graft in 10 patients. Follow-up ranged from 1 to 5 years (between January 1995 and January 2000). Radiographs and computed tomography (CT) scans were analyzed for confirmation of osseous union. The average period of clinical and radiologic union was 13.2 weeks (range, 10 to 33 weeks). There was a significant improvement in the grasping power, radiologic healing, clinical satisfaction, and pain relief in the patients who had operation. PMID- 12132821 TI - Prevalence of orthopaedic maladies in people who flyfish: an Internet-based survey. AB - In this study, we define the prevalence of back and joint pains in people who flyfish. We use the Internet as a source of data. Requests for participation were posted on flyfishing Web sites. Eighty-nine people were surveyed (E-mail group). A control group of 42 flyfishing club members participated (Club group). Epidemiologic information, flyfishing data, and location of pain were collected. Low back pain in the 131 participants was 59%. No differences between the two groups in location or prevalence were noted. Saltwater fishermen had the highest rates of shoulder and elbow pain (31%). Trout fishermen had the highest rate of wrist pain (31%). Warmwater anglers had the least leg (12%), elbow (12%), and shoulder pain (18%). These afflictions mirror reports for other recreational activities. Low back pain was the most prevalent complaint, followed by wrist/hand and shoulder. Flyfishing subtypes have different rates and locations of pain, explained by equipment and technique. This report defines use of the Internet as a data source for research. PMID- 12132822 TI - Langerhans cell granulomatosis manifested as pigmented villonodular synovitis. AB - We report an unusual case of Langerhans cell granulomatosis (LCG) manifested as a villous synovial proliferation in a 38-year-old female jogger. One year after the onset of joint symptoms, she had a classical LCG presentation with skin and visceral lymph node involvement. Review of the literature revealed only one case of synovial shoulder joint tenosynovitis associated with LCG in a middle-aged woman. Ours is the first reported case presenting clinically in the synovium of the hip joint as pigmented villonodular synovitis. Histiocytic/dendritic proliferations involving the synovial tissues are not uncommon. These lesions as well as the rare multicentric reticulohistiocytosis (MRH), a systemic monocytoid/histiocytic disorder with multinucleated giant cells, polyarthritis, and papulonodular skin lesions, should be considered in the differential diagnosis. Clinical and pathologic features will distinguish LCG from MRH. PMID- 12132823 TI - Latrogenic snapping of the medial head of the triceps after ulnar nerve transposition. AB - We postulate an iatrogenic cause for snapping of the medial head of the triceps. A patient whose ulnar nerve and triceps did not dislocate over the medial epicondyle preoperatively had snapping of a portion of the medial triceps after submuscular transposition of the ulnar nerve. We believe that release of the brachial fascia and excision of the medial intermuscular septum removed the restraint to anterior translation of the medial aspect of the triceps, permitting dislocation of a portion of the medial head of the triceps with elbow flexion in this case. Previous reports of snapping of the triceps resulting after ulnar nerve transposition occurred in patients whose ulnar nerve dislocated preoperatively; in these cases, the triceps was thought to have dislocated preoperatively (along with the ulnar nerve) but was not recognized. Careful intraoperative assessment of the triceps after ulnar nerve transposition should prevent medial triceps instability as a postoperative concern. PMID- 12132824 TI - Outcome of subtalar arthrodesis after calcaneal fracture. AB - Between 1983 and 1995, we used subtalar arthrodesis to treat 16 consecutive patients for continued pain after an intra-articular calcaneal fracture. Average time to union was 3 months (2 to 4 months). Complications were minor in 4 patients, and major in 4 others. Length of follow-up in 14 patients was 55 months (range, 12 to 112 months). Hindfoot scores (clinical rating system of the American Orthopaedic Foot and Ankle Society) improved from 38 (range, 28 to 62) to 67 (range, 39 to 94). Results of medical outcome surveys indicate that patients had low scores in areas related to physical conditioning, physical role functioning, and bodily pain. We conclude that the majority of patients can have improvement with surgical reconstruction that addresses a specific problem, but pain relief is usually not complete. PMID- 12132825 TI - Patient self-testing of prothrombin time after hip arthroplasty. AB - We determined whether total hip arthroplasty (THA) patients could test their own prothrombin time reliably over 6 weeks of anticoagulation prophylaxis with a portable device that measures prothrombin time and whether self-testing would improve or maintain the quality of care at a lower cost than our standard procedure. Forty-six THA patients participated in the study and were compared with a matched group managed with our standard protocol using a home health-care nurse. Seven patients (15%) could not be trained to obtain the blood sample, and others required multiple finger sticks to obtain valid results. However, the results from the 29 patients completing the study showed high reliability when compared with results obtained through standard protocol. Self-testing saved about $260 per patient over the cost of venipuncture. Patient self-testing of prothrombin time using the device in this study is reliable and cost-effective for monitoring the anticoagulation status after THA in a select group of elderly patients. PMID- 12132826 TI - Ulnar artery thrombosis: a 6-year experience. AB - Thrombosis of the ulnar artery can be a cause of significant morbidity. Most often a consequence of blunt trauma to the hypothenar eminence of the hand, it may be attributable to one traumatic event or to repetitive insults. Surgery is often required. We reviewed the presentation and diagnosis of ulnar artery thrombosis and evaluated the effectiveness of treatment by ulnar artery excision with interposition vein grafting. Retrospective chart analysis from 1989 to 1995 at the Medical Center of Central Georgia showed that nine patients (eight male, one female) were treated for ulnar artery thrombosis. Three had associated ulnar artery aneurysms. Eight of the nine were treated with artery excision and interposition vein grafting. Four also received stellate ganglion blocks before surgery. One was treated with stellate ganglion blocks alone. All patients had symptomatic relief and resolution of physical findings. We conclude that ulnar artery thrombosis can be managed with ulnar artery excision and interposition vein grafting when conservative measures fail. PMID- 12132827 TI - Thromboembolism after total knee arthroplasty: intermittent pneumatic compression and aspirin prophylaxis. AB - This is a study of two consecutive antithromboembolism regimens after total knee arthroplasty. In group 1, 131 patients were given aspirin prophylaxis alone (650 mg by mouth twice a day). In group 2, 123 patients were treated with aspirin, knee-high compression stockings, and intermittent knee-high pneumatic compression devices, which were started intraoperatively. The prevalence of deep vein thrombosis in group 1 was 15.9% (21 of 131 patients). One patient had a possible symptomatic nonfatal pulmonary embolism, and one patient had a symptomatic calf thrombus. Asymptomatic thrombi were detected in calf veins in 9 patients, popliteal vein in 6 patients, and femoral vein in 5 patients. In Group 2, the prevalence was 7.4% (9 of 123 patients). Asymptomatic thrombi were located in calf veins in 6 patients, popliteal vein in 1 patient, and femoral vein in 2 patients. There was a significant difference in the prevalence of deep vein thrombosis between the two groups. A history of previous thromboembolism was a significant risk factor for a new thrombus. The prevalence after bilateral one stage knee arthroplasty was 24.3% for group 1 and 12.5% for group 2. Aspirin and knee-high intermittent pneumatic compression together are more effective than aspirin alone for prevention of deep vein thrombosis after primary and revision knee arthroplasty. PMID- 12132829 TI - Nerve repair and grafting in the upper extremity. AB - Knowledge of peripheral nerve anatomy, coupled with available clinical tools for examination, provides the foundation for initial evaluation. Use of this information may be related to the specific injury and a reconstructive plan formulated to maximize the potential for recovery. Realistic expectations can be presented to the patient based on information from summarized clinical experience. The field of nerve repair continues to change rapidly, and clinicians must utilize all resources available for planning, evaluation, and management. As the cellular behaviors of nerve regeneration are delineated, new avenues of research will open. PMID- 12132828 TI - Isthmic spondylolisthesis and spondylolysis. AB - Isthmic spondylolisthesis, or spondylolisthesis due to a lesion of the pars interarticularis, is a common source of pain and disability in both the pediatric and adult population. This review examines the current diagnostic and treatment options for patients with this condition. It also reviews the results of the various interventions to facilitate the surgeon in choosing the appropriate treatment option for any given patient. PMID- 12132830 TI - Template versus freehand bone-patellar tendon-bone harvest for anterior cruciate ligament reconstruction: its effect on patellofemoral morbidity. AB - To test the hypothesis that a template system for harvesting bone-patellar tendon bone (BTB) would cause fewer postoperative patellofemoral problems than freehand graft harvests, two groups of 20 patients who had arthroscopically assisted anterior cruciate ligament (ACL) reconstructions were compared at 1 and 3 years. Three years after surgery, no significant differences were found in the quality of the ligament reconstruction. One intraoperative patellar fracture occurred in the freehand group (group 1). All other complications were similar between groups. Squatting profiles were slightly better in the template group (group 2). Averaged and outlier scores from three published patellofemoral scoring systems showed no significant differences between the two groups. It was concluded that current methods for measuring patellofemoral function detected little morbidity and no significant outcome differences between ACL reconstructions done with BTB grafts harvested by freehand versus templated techniques. PMID- 12132832 TI - Replantation of digits and hands. PMID- 12132831 TI - Management of leg length inequality. AB - Leg length inequality is common. Treatment objectives include obtaining leg length equality, producing a level pelvis, and improving function. Clinical assessment should include determination of a level pelvis with the patient standing using a set of blocks of various heights to estimate the amount of leg length inequality. Radiographic measures include the teleroentgenogram, orthoradiograph, and computed tomography (CT). A prediction of the ultimate leg length inequality at skeletal maturity will be needed to determine treatment. Our guidelines for treatment of leg length inequality are as follows: <2 cm--no treatment or a lift in the shoe; 2 to 6 cm--an epiphysiodesis or shortening procedure is considered; 6 to 15 cm--a lengthening procedure is considered. A leg length inequality of 15 to 20 cm--may require a staged lengthening, lengthening combined with epiphysiodesis, or amputation. Numerous complications of limb lengthening procedures occur frequently, even in experienced hands. PMID- 12132833 TI - Acute carpal tunnel syndrome caused by idiopathic tumoral calcinosis. AB - We present a case of acute carpal tunnel syndrome due to compression of the median nerve within the carpal tunnel caused by tumoral calcinosis. Wrist immobilization and oral corticosteroids produced excellent clinical resolution. Radiologically, the calcific mass disappeared. PMID- 12132834 TI - Biomechanics of minor automobile accidents. PMID- 12132835 TI - Incorrect posterior axial line. PMID- 12132836 TI - Ganglion of the hip: report of five cases. AB - Ganglion of the hip is a rare cause of inguinal swelling. Ultrasound examination and MRI lead to the correct diagnosis. Resection should be considered for large ganglia and in cases with impairment of neighboring structures. Conservative treatment may suffice in small and asymptomatic ganglia. PMID- 12132837 TI - Elbow arthroscopy for neglected osteochondritis dissecans of the capitellum. AB - We reviewed results of elbow arthroscopy for neglected osteochondritis dissecans (OCD) of the capitellum. Of more than 140 patients having elbow arthroscopy at our institution, 10 patients (11 elbows) had OCD of the capitellum and fulfilled inclusion criteria. Criteria included symptoms of at least 2 years' duration, loss of motion for at least 6 months, or radiographic evidence of secondary degenerative changes. One patient was lost to follow-up. The remaining 9 (10 elbows) were male patients with a median age of 20 years (range, 15 to 58 years). Follow-up averaged 4.6 years (range, 2 to 8 years). Using a 100-point system, postoperative elbow scores averaged 92 (range, 55 to 100). All patients returned to preoperative activities, though only 8 of 10 believed that surgery resulted in improvement. Elbow arthroscopy for neglected OCD can result in functional improvement. However, results are not as good as those reported with earlier intervention. PMID- 12132838 TI - Primary patellar tendon repair and early mobilization: results in an active-duty population. AB - We retrospectively reviewed 13 patellar tendon repairs done over 32 months at a tertiary care, military medical center. Early mobilization was initiated within 2 weeks postoperatively. Clinical and functional results were statistically examined with relation to age, timing of surgery, length of follow-up, quadriceps atrophy, extensor lag, patella position, and time to full duty. At an average of 24 months' follow-up, six patients (46%) had thigh girth atrophy, and one patient (8%) had an extensor lag >5 degrees. Mean Lysholm score was 84 (range, 57 to 100). Maximum postoperative Tegner activity scores averaged 7.1 (range, 5 to 10). Clinical results classified five cases as excellent, three good, three fair, and two poor. Functional results classified three cases as excellent, four good, two fair, and four poor. Time to return to duty averaged 13 months. Our results suggest that adequate extensor function can be restored after primary repair and immediate motion therapy. PMID- 12132839 TI - Anatomy of the knee extensor mechanism: correlation with patellofemoral arthrosis. AB - The patellofemoral articulation is a common and significant source of disability and discomfort in the aging population. This study examined the anatomy of the knee extensor mechanism in patients having primary total knee arthroplasties, characterized the anatomic variations of the extensor mechanism, and correlated these findings with the location and extent of osteoarthritic change of the patellar undersurface. Sixty-two knees (57 patients) were evaluated prospectively. Specific characteristics that were analyzed included the mean Outerbridge grade for rating patellar cartilage degeneration and anatomic patterns of the extensor mechanism. Knees with a quadriceps tendon width at 2 and 5 cm above the patella that differed by less than 1 cm had more statistically significant patellar degeneration in all patellar locations than knees with tendon width differences greater than 1 cm. Anatomic variations, such as tendons with minimal increments in width in the proximal-distal direction, may be associated with an increasing amount of patellar arthrosis at the lateral facet, central ridge, and, most significantly, medial facet. PMID- 12132840 TI - Knee flexion contractures: soft tissue correction with monolateral external fixation. AB - We assessed the efficacy of progressive soft tissue distraction using monolateral external fixation in the management of severe knee flexion contractures. We prospectively evaluated 10 knee deformities in seven pediatric patients. After gradual distraction using the modified Orthofix Limb Reconstruction System (LRS), most recent functional status and knee range of motion were determined. This treatment was applied to 10 extremities in seven patients, ranging in age from 2 to 16 years. Diagnoses included arthrogryposis (4), sickle cell disease (1), previous sepsis (1), and congenital pterygium (1). Average preoperative flexion contracture was 80.5 degrees. Each patient achieved full extension. There was one recurrence, despite bracing, which was managed with replacement of the fixator and soft tissue procedures. Management of knee flexion contractures using a monolateral fixator appears to be a viable alternative to extensive release or femoral osteotomy. Long-term follow-up will be essential to assess the overall risk of recurrence and complications. PMID- 12132841 TI - Osteogenesis in a rat model: use of bone marrow cells and biodegradable gelatin matrix carrier. AB - We investigated the use of a biodegradable porcine gelatin matrix (Gelfoam) as a carrier for marrow cells that induce osteogenesis at ectopic sites in rats. Bone marrow cells obtained from the long bones of 6-week-old Sprague-Dawley rats were dissociated enzymatically and the cells reconstituted in rat serum. Twenty million cells in 0.1 mL of serum were then adsorbed into 1-cm3 pieces of gelatin matrix substrate and implanted into ectopic sites in live Sprague-Dawley rats. The implants were retrieved and analyzed histologically for bone and cartilage formation 3, 4, 6, and 8 weeks after implantation. Woven bone was seen as early as 3 weeks and persisted through 8 weeks. No cartilage was observed. Osteoclasts first appeared at 3 weeks, peaking in number at 4 weeks. By 6 and 8 weeks, only small islets of substrate remained, surrounded by dense, woven bone. Control implants, consisting only of serum adsorbed into the carrier, showed no bone formation. We conclude that biodegradable gelatin matrix can serve as a carrier for the osteogenic cells of bone marrow in rats in ectopic sites. Such a system may be further developed to augment bone healing. PMID- 12132842 TI - "Always do right...". PMID- 12132843 TI - Osteoid osteoma of the coracoid process. AB - Osteoid osteoma is a small, benign, and painful tumor most commonly affecting the extra-articular portions of the long bones, especially the femur or tibia. Osteoid osteoma of the coracoid process is so rare that we have found only three previously reported cases in the international literature. We describe our experience in managing a case of osteoid osteoma in this unusual location. PMID- 12132844 TI - Heterotopic ossification after lateral epicondylectomy. AB - Lateral epicondylitis is a common affliction that can be treated nonsurgically. However, surgical treatment is rarely indicated, and results are usually effective in more than 90% of patients. Although complications have been described, ours is the first report of heterotopic ossification after an uncomplicated lateral epicondylectomy. PMID- 12132845 TI - Septic sacroiliitis: the overlooked diagnosis. AB - Infection of the sacroiliac joint is extraordinarily rare. However, the initial symptoms and signs associated with this condition are mundane, thus leading to delay in diagnosis, or even misdiagnosis. I report an extremely unusual pyogenic sacroiliitis caused by Salmonella and discuss key clinical components and protocol for the successful evaluation, diagnosis, and treatment of this uncommon illness. PMID- 12132846 TI - Cementless S-ROM femoral component: effect of stem length on stability after extended proximal femoral osteotomy. AB - The extended proximal femoral osteotomy is becoming increasingly popular in revision total hip replacement. Our study was done to determine the femoral stem length required for stable fixation of a cementless femoral component after an extended proximal femoral osteotomy. Three lengths of the S-ROM femoral stem were implanted in paired cadaver femora and tested under torsional and axial loads. The results indicate that the standard (160 mm) and long (215 mm) stems do not provide adequate torsional stability after a 160 mm extended proximal femoral osteotomy. The extra-long (255 to 315 mm) stems provided significantly greater stability, suggesting that the extended proximal femoral osteotomy may need to be bypassed by more than 2 cortical diameters, especially when a flexible stem such as the S-ROM is used. PMID- 12132847 TI - Simple, hybrid deep venous thrombosis/pulmonary embolus prophylaxis after total hip arthroplasty. AB - A 7.1% deep venous thrombosis rate followed total hip arthroplasty in 56 patients using a hybrid prophylactic regimen against deep venous thrombosis and pulmonary embolus. There were no bleeding complications, no symptomatic pulmonary emboli, and no unexplained deaths. The regimen consisted of an initial loading dose of warfarin, usually 10 mg, the night of surgery followed by 2.5 mg/day for 3 weeks, with dosage adjustments only in cases of over-anticoagulation. This regimen was combined with elevated sling suspension of the operative leg, bilateral pedal intermittent pneumatic compression devices, and early mobilization. This prophylactic regimen is simple, inexpensive, efficacious, and compatible with an early hospital discharge. PMID- 12132848 TI - Minimally invasive surgical technique for unicondylar knee arthroplasty. AB - Between August 1992 and December 1996, more than 700 unicondylar knee arthroplasty (UKA) procedures were done by the senior author (JA.R.), using well defined patient selection criteria, which is paramount to the outcome of the procedure. The UKA procedure described is done through a smaller incision than that required for total knee arthroplasty (TKA) (3 inches versus 8 inches), thus minimizing blood loss (less than 200 mL), avoiding normal tissue sacrifice (opposite compartment, patellar bone, and cruciate ligaments), and decreasing morbidity (no patellofemoral disruption). The UKA costs less because it is done as an outpatient procedure in 80% of cases; since postoperative physical therapy is minimal or unnecessary, recovery time is shorter (90% independent function at 2 weeks after operation). Whereas TKA can have universal application, UKA is patient specific and cannot replace TKA in all circumstances. Likewise, the techniques for TKA and UKA are not interchangeable. However, with the use of well defined patient populations and surgical techniques, the intermediate results of UKA have paralleled reported outcomes of TKA. PMID- 12132849 TI - Recurrent carpal tunnel syndrome: treatment options. AB - Many techniques are available to the surgeon confronted with a patient with recurrent nerve entrapment/traction neuritis. The advantage of local muscle or fat flaps like the ADQ, pronator quadratus, and hypothenar fat flap is the ease at which each can be elevated. The size is very limited, however. Circumferential vein wrapping has been studied extensively and is efficacious despite the absence of vascularity, but length requirements may limit the indications to short nerve segments, as might be expected with recurrent carpal tunnel syndrome. Despite the increased complexity of a pedicled flap and free tissue transfer, the tissue availability, girth, and source of vascularity make these sophisticated techniques more attractive. Such flap transfers should be used only after the more simple techniques have been ruled out as options or failed and only if residual pain and hypersensitivity are due to localized neuritis as opposed to a more generalized dystrophic symptomatology. PMID- 12132850 TI - Pelvic ring injuries. AB - Major pelvic disruptions lead to significant problems in acute resuscitation and definitive management. An understanding of the injury--its assessment and treatment principles--are necessary for appropriate management by all orthopaedic surgeons. In this article, we summarize the pelvic ring session at the Southeast Fracture Symposium. PMID- 12132851 TI - Southern medicine during the War between the States. 1994. PMID- 12132852 TI - Knee arthroplasty: pain and swelling 6 years later. PMID- 12132853 TI - Reconstruction of longitudinal stability of the forearm after disruption of interosseous ligament and radial head excision (Essex-Lopresti lesion). AB - We have presented a technique for reconstruction of the central band of the interosseous membrane in conjunction with surgical repair of the distal radial joint and radial head prosthesis. With this technique, we address all three anatomic structures that provide longitudinal stability of the forearm, specifically, (1) radial head replacement, (2) interosseous membrane reconstitution, and (3) TFCC repair. PMID- 12132854 TI - Volar intercalated segment instability after scapholunate ligament reconstruction. AB - We describe an 18-year-old man with an acute lunate dislocation treated with open reduction, ligament repair, capsulodesis, and percutaneous pinning. After pin removal and mobilization, the patient was found to have midcarpal collapse; the lateral radiographs showed a volar intercalary segment instability. We now, besides open reduction internal fixation scapholunate repair, recommend routine pinning of the lunotriquetral interval and careful evaluation of the dorsal radiocarpal ligament during repair of lunate and perilunate dislocations. PMID- 12132855 TI - The terrible triad of the shoulder. AB - This case shows that the terrible triad of the shoulder-anterior dislocation, brachial plexus injury, and rotator cuff tear-can occur in younger populations than previously described. A high index of suspicion for this triad of injury should be maintained, particularly in those young individuals with grossly unstable shoulders. Once axillary nerve function has begun to return, simultaneous repair of all soft tissue injuries can lead to a stable, functional joint. PMID- 12132856 TI - Regeneration of an acromioclavicular joint after a type III dislocation in an adult. PMID- 12132857 TI - Retroacetabular osteolysis. PMID- 12132858 TI - Recurrent dislocation of the hip in a child with Down syndrome: a 20-year follow up. AB - This is a report of recurrent dislocation of the hip in a child with Down syndrome. The child was treated with a soft tissue repair that prevented further dislocations. However, 20 years later, the hip had subluxed and secondary degenerative arthritis had developed. This case emphasizes the necessity of life long follow-up and prompt treatment of hip disease in patients with Down syndrome. PMID- 12132859 TI - Love your patients. PMID- 12132860 TI - Improved percutaneous slipped capital femoral epiphysis treatment: continuous biplanar fluoroscopy and proper guide wire selection. AB - In 1990, a report was published outlining a technique of percutaneous fixation of slipped capital femoral epiphyses dramatically diminishing the morbidity associated with the open technique. Technical difficulties are still encountered with the morbidly obese child and the percutaneous technique. Two fluoroscopic units used continuously during the technique facilitate placement of the guide wire in a more acceptable anatomic location. Guide wire stiffness was also measured and used in preoperative planning. Ten hips were treated with this technique, and a prospective analysis of the surgical time and fluoroscopy time was done. Using the new technique, we found a significant reduction in surgical and fluoroscopic times. PMID- 12132861 TI - Cortical strut allografts for the treatment of femoral fractures and deficiencies in revision total hip arthroplasty. AB - Thirty-three hips had revision total hip arthroplasty, using an average of three cortical strut allografts fixed to the femur with cables and followed-up for a mean of 4 years. The indications for strut allografts were ectatic femurs or segmental defects of the femoral diaphysis (22 hips), femoral fractures (10 hips), and severe proximal femoral osteolysis (1 hip). Twenty-one hips had an excellent or good clinical result, 6 had a fair clinical result, and 6 had a poor clinical result. Reoperation was done in six hips, but in only two hips was reoperation related to failure of the allograft. Nine of the 10 femoral fractures repaired with allograft struts healed by 3 to 6 months. Radiographs showed partial or complete bridging of the allograft to host bone with peripheral remodeling and minimal resorption in 30 of 33 hips. Strut allografts, fixed with multiple cables, are an important adjunct to femoral component revision for the restoration of deficient femoral bone stock and in the treatment of periprosthetic femur fractures. PMID- 12132862 TI - Initial biomechanical properties of anterior cruciate ligament reconstruction autografts. AB - To provide more information to consider when selecting a reconstruction technique, we did a side-by-side comparison of some of the initial biomechanical properties of currently accepted reconstruction methods. Our research hypotheses were that a quadrupled, woven semitendinosus and gracilis graft is as strong as any of the other commonly used graft materials and that quadrupling and weaving the hamstring graft may increase the stiffness of the overall construct Using lower extremity cadaveric specimens harvested from young donors, we fashioned seven each of seven types of graft: 9-mm, 10-mm, and 11-mm-wide patellar tendon graft (PTG); 10-mm-wide central quadriceps tendon graft; doubled semitendinosus graft; tripled semitendinosus graft; and quadrupled, woven semitendinosus and gracilis graft. Specimens were stripped of remaining soft tissue, and anterior cruciate ligament (ACL) constructs were created for biomechanical testing. The tibia was translated anteriorly on the femur, mimicking a pivot shift maneuver, andfailure strength, failure mechanism, and construct stiffness were recorded. No differences in mean strength were detected. The quadrupled, woven graft was significantly stiffer than the doubled semitendinosus graft and no less stiff than any of the PTG constructs. All grafts showed similar and adequate initial absolute strength to reconstruct the ACL. Quadrupling and weaving the semitendinosus and gracilis graft increases the stiffness of the reconstructed specimen to a level statistically similar to that of specimens reconstructed with a PTG. PMID- 12132863 TI - Marjolin ulcers: secondary carcinomas in chronic wounds. AB - Marjolin ulcers are malignant tumors arising in chronic wounds. Strictly defined, they include carcinomas that transform from the chronic open wounds of pressure sores or burn scars. They behave aggressively and have a propensity for local recurrence and lymph node metastases. A retrospective study was done at a single institution identifying six individuals who had chronic wound ulcers that underwent malignant transformation into a carcinoma. Sinus tract degeneration in osteomyelitis was not included. The average latency time between ulcer formation and documentation of a malignancy was 30 years. All wounds were about the pelvis or flank. Major oncologic surgical procedures were done in an attempt to eradicate the cancer. High-grade tumors had positive lymph node metastases, portending a grave prognosis. All four individuals with nodal metastases eventually died of systemic disease. Early recognition and proper staging offers the best chance for cure. PMID- 12132865 TI - Open fractures of the hand. AB - Open fractures of the hand are a challenging clinical problem for the orthopedic surgeon. The fracture is often comminuted with substance loss. Additionally, the fracture site can be contaminated by foreign material. The soft tissue envelope is violated with a variable degree of tissue devitalized. The wound contamination and tissue destruction lead to a rate of infection that can be much higher than that for a closed fracture. Initially, management of a significant soft tissue injury must take precedence over definitive fracture fixation. Proper staging of debridement, wound closure, and definitive fixation is paramount in minimizing infection while obtaining fracture union. PMID- 12132864 TI - Epidermoid inclusion cysts of the hand. AB - Epidermoid inclusion cysts occur second only to ganglions in terms of tumefaction presentations in the hand. This review describes 60 such lesions and reviews the clinical presentation, diagnosis, and treatment. The epidermoid inclusion cyst probably arises from some traumatic event that drives epithelial cells into the subcutaneous tissues where they survive, grow, and produce keratin. PMID- 12132866 TI - Carpal tunnel syndrome: current concepts. AB - Carpal tunnel syndrome (CTS) is a clinical syndrome manifested by characteristic signs and symptoms resulting from an entrapment neuropathy of the median nerve at the wrist. It is the most common compression neuropathy in the upper extremity. In this paper, the etiology and pathophysiology of CTS are reviewed, as well as the clinical examination and other tests that may be useful in establishing its diagnosis. A variety of surgical techniques have been espoused for treating CTS refractory to non-operative treatment and the proposed advantages and disadvantages of the new procedures are discussed. If the diagnosis is correct, then surgical results are reliably good. PMID- 12132868 TI - Fibrosarcoma of the sacrum in a child: management by sacral resection and reconstruction. AB - We describe the diagnosis and surgical management of fibrosarcoma of the sacrum in a pediatric patient. We retrospectively reviewed the literature and a case report. Total sacrectomy is a viable treatment option in the management of large malignant sacral tumors. The techniques used in adult patients can be extended to and improved upon for pediatric patients. The input of multiple surgical specialties is essential in the management of these lesions. PMID- 12132867 TI - Replantation of completely amputated distal forearm--1965. AB - Although replantation of completely amputated wrists and forearms is now commonplace, in 1965 the replantation of this "wrist level" amputation was the first reported in the western world. The details of the technique used are contrasted with current standard of care. PMID- 12132869 TI - Posterior extradural lumbar disk fragment. AB - We present the unique case of a patient with a sequestered disk fragment posterior to the thecal sac producing symptoms of spinal stenosis with neurogenic claudication. The majority of sequestered disk fragments migrate in either a cranial or caudal direction. In only a few cases have disk fragments been identified posterior to the thecal sac. Our patient had a sudden onset of bilateral groin and anterior thigh pain. Magnetic resonance imaging showed relatively severe stenosis at L4-5 with mild disk bulging. Intraoperatively, a large posteriorly placed encapsulated mass of soft tissue was found compressing the posterior portion of the thecal sac. Patients with acute onset of symptoms of spinal stenosis should have herniated disk included in their differential diagnosis, even in the absence of imaging confirmation. PMID- 12132870 TI - hMLH1 and hMSH2 mutations in families with familial clustering of gastric cancer and hereditary non-polyposis colorectal cancer. AB - The pattern of hMLHI and hMSH2 mutations was assessed to identify the genetic correlation between hereditary gastric and colorectal cancers. Four disease groups and their healthy family members were assembled according to the presentation of gastric cancer: FG, familial clustering of gastric cancer (n = 32); CG, family with one or more colorectal and gastric cancers in first-degree relatives (n = 22); HS, seven HNPCC families corresponding to the Amsterdam criteria (AMS+) and 12 suspected HNPCC families which did not satisfy one of the criteria (AMS-), but no gastric cancer among first- and second-degree relatives (n = 19); and SG, sporadic gastric cancer (n = 33). In the CG group, three were included in AMS + and six in AMS- criteria. Peripheral blood was obtained from them to detect hMLHI and hMLH2 mutations using PCR-SSCP analysis and direct sequencing. The incidence of mutations was 9.4% in the FG group, 54.5% in the CG group, 31.6% in the HS group, and none in the SG group. The incidence, type, and number of the mutation were not different between the CG and HS groups. Thirty four different mutations included 19 in hMLH1 and 15 in hMSH2. Gastric cancer was the most common extracolonic malignancy in HNPCC and suspected HNPCC families (9/28, 32.1%). The hMLH1 or hMSH2 mutation occurred in seven of 10 families with AMS+, whereas it occurred in four of 18 with AMS- (70% vs. 22.2%, P = .013). Five mutations in the hMLH1 and six mutations in the hMSH2 were exclusively found in families with gastric cancer. All three mutations in the FG group were in hMLHI and there was no mutation in their healthy family members. This study demonstrates that some familial clustering type of gastric cancer appears to be associated with hMLHI mutations thereby indicating a difference from the hereditary gastric cancer studies previously reported. In addition, hMLHI and hMSH2 mutations may impact the gastric cancer carcinogenesis in HNPCC or suspected HNPCC. PMID- 12132871 TI - Breast cancer risk factors and HER2 over-expression in tumors. AB - Few epidemiologic studies have investigated the potential role of HER2 in the etiology of breast cancer. We conducted a case-case study of 156 women with incident, invasive ductal carcinoma. Multivariate unconditional logistic regression was used to estimate the odds ratios for a HER2 positive tumor in relation to known and putative risk factors of breast cancer. HER2 status was detected by immunohistochemistry on archival tissue. HER2 positive breast cancers tended to be larger and were less likely to express estrogen receptors, and the incidence rate was higher in patients less than 40 years old. We observed an association between a self-reported history of benign breast disease and the occurrence of HER2 positive breast cancer (OR, 2.1;95% CI, 1.1-4.1). We did not detect associations between HER2 over-expression and family history of breast cancer, parity, late age at first birth, ever having breast fed an infant, or oral contraceptive use. Our findings merit consideration in light of recent evidence of HER2 amplification or over-expression in benign breast disease. Should the link to breast cancer be established, HER2 positive benign breast disease could potentially serve as an early marker for preventive intervention. PMID- 12132872 TI - Carrageenan-induced inclusions in mammary myoepithelial cells. AB - The purpose of this investigation was to characterize the ultrastructural changes that occur in mammary myoepithelial cells (MMEC) following exposure in tissue culture to low concentrations of lambda-carrageenan, a sulfated polysaccharide commonly used as a food additive. MMEC were obtained from reduction mammoplasty, grown in tissue culture, exposed for varying durations to low concentrations (0.0014%-0.0001%) of lambda-carrageenan, and examined by transmission electron microscopy, following staining for acid phosphatase and for aryl sulfatase. Carrageenan appeared to enter the cells by membrane-associated endocytic vesicles and accumulate in endosomes and lysosomes. Unusual lamellar inclusions were identified within lysosomes of the MMEC, and lysosomal vacuolation arose in association with the inclusions. The observed changes appeared to lead to destruction of the MMEC by release of proteolytic enzymes from the distorted lysosomes, similar to the process observed in lysosomal storage diseases. PMID- 12132873 TI - Effects of the dietary phytoestrogens daidzein and genistein on the incidence of vulvar carcinomas in 129/J mice. AB - The objective of this study was to determine the effect of dietary phytoestrogens on the incidence of spontaneous vulvar carcinomas in 129/J mice using three natural ingredient diets and two purified diets containing predetermined levels of daidzein and genistein. Eighty weanling female mice without clinical evidence of vulvar carcinomas were randomly assigned 16 per diet to each of 5 test diets. Mice were clinically examined for vulvar masses weekly for 3 months and at monthly intervals thereafter. Vulvar carcinomas in representative groups of mice were confirmed using routine histological procedures. The incidence of vulvar carcinomas increased sharply in mice on all test diets during the first 2 months with minor changes during the remainder of the study. Within one month, the incidence of vulvar carcinomas in mice fed the AIN-76A modified soy protein diet was significantly (P < .05) increased over those of mice fed the AIN-76A modified casein diet, the #5K96, or the # 5058 diet. At three months, the incidence of vulvar carcinomas in mice fed the soy protein diet was significantly (P < .05) increased over those of mice fed the NIH-31 diet or the PMI #5K96 diet. There was a marginally significant (P < .10) correlation between the total daidzein and genistein levels in the five test diets and the incidence of vulvar carcinomas in mice as determined by clinical examination. We concluded that dietary levels of daidzein and genistein were associated with an increase in the incidence of vulvar carcinomas in mice and that the 129/J mouse may provide an animal model for studying the development of vulvar carcinomas. PMID- 12132874 TI - Importance of human papillomaviruses for the development of skin cancer. AB - The frequent detection of HPV DNA in non-melanoma skin cancers was shown in several studies; however, the role of HPV in the development of these cancers remains speculative. We analyzed different skin tumors, normal skin, and hair follicles for HPV DNA using a PCR system designed to detect all HPV types known so far. HPV DNA was found in 93% of common warts, 69% of squamous cell carcinomas (SCC), 52% of basal cell carcinomas (BCC), 41% of actinic keratoses, 31% of extragenital Bowen's disease, 22% of keratoacanthomas, 16% of normal skin tissues and 47% of hair follicles. No individual HPV type predominated in any of the skin tumors. The number of HPV genomes in individual neoplasms (SCC and BCC) seems to be less than I per cancer cell. These results indicate that a direct role of HPV in skin cancerogenesis remains questionable. Possibly, mechanisms different from the activity of HPV oncoproteins in genital cancers are involved in skin neoplastic transformation. PMID- 12132875 TI - Clostridium as a tumor-specific delivery system of therapeutic proteins. AB - The feasibility of gene therapy strategies in cancer treatment still has important pitfalls. Transfer of therapeutic proteins to the hypoxic/necrotic 'extracellular' microenvironment of solid tumors, based on the engineering of nonpathogenic clostridia is proposed as an alternative methodology. Using the rat rhabdomyosarcoma R1 in vivo tumor model, we demonstrated that Clostridium species colonized the tumors, whereas proliferation of these bacteria was absent in normal tissues. C. acetobutylicum was genetically engineered to express and secrete either mTNF-alpha or the E. coli cytosine deaminase. Quantitative in vitro data showed stability of the vectors, and significant levels of biologically active therapeutic proteins in lysates and supernatants of recombinant clostridia. Administration of either of these recombinant Clostridium strains to tumor-bearing rats resulted in the presence of active proteins in the tumor tissue. Based on these data and supported by its selective colonization pattern and safety, the Clostridium gene transfer system offers a potential application in anti-cancer therapies. PMID- 12132876 TI - Comparative study of sister chromatid exchange induction and antitumor effects by homo-aza-steroidal esters of [p-[bis(2-chloroethyl)amino]phenyl]butyric acid. AB - The present work was undertaken in order to test the hypothesis that the Sister Chromatid Exchange (SCE) assay in vitro can be used for the prediction of in vivo tumor response to newly synthesized potential chemotherapeutics. The effect of three homo-aza-steroidal esters containing the -CONH- in the steroidal nucleus, 1, 2, and 3 on SCE rates and on cell kinetics in cultured human lymphocytes was studied. The antitumor activity of these compounds was tested on leukemia P388- and leukemia L1210-bearing mice. The three substances induced statistically significant enhancement of SCEs and of cell division delays. Compounds 1 and 3 were identified, on a molar basis, as more effective inducers of SCEs and of cell division delays compared with compound 2. Compounds 1 and 3 had upon both experimental tumors better therapeutic effects compared with compound 2 at equitoxic doses. Therefore, the order of the antitumor effectiveness of the three compounds coincided with the order of the cytogenetic effects they induced. PMID- 12132877 TI - Antioxidant properties of vitamin C: new issues. PMID- 12132878 TI - Novel ZE-isomerism descriptors derived from molecular topology and their application to QSAR analysis. AB - We introduce several series of novel ZE-isomerism descriptors derived directly from two-dimensional molecular topology. These descriptors make use of a quantity named ZE-isomerism correction, which is added to the vertex degrees of atoms connected by double bonds in Z and E configurations. This approach is similar to the one described previously for topological chirality descriptors (Golbraikh, A., et al. J. Chem. Inf. Comput. Sci. 2001, 41, 147-158). The ZE-isomerism descriptors include modified molecular connectivity indices, overall Zagreb indices, extended connectivity, overall connectivity, and topological charge indices. They can be either real or complex numbers. Mathematical properties of different subgroups of ZE-isomerism descriptors are discussed. These descriptors circumvent the inability of conventional topological indices to distinguish between Z and E isomers. The applicability of ZE-isomerism descriptors to QSAR analysis is demonstrated in the studies of a series of 131 anticancer agents inhibiting tubulin polymerization. PMID- 12132879 TI - Comparative molecular field analysis (CoMFA) of anionic azo dye-fiber affinities I: gas-phase molecular orbital descriptors. AB - This paper presents quantitative structure-activity relationship (QSAR) models for a series of 30 anionic azo dyes applied to cellulose fiber by comparative molecular field analysis. Two forms of the dye molecules (neutral and anionic) are compared. Neutral structures give better statistical results than the anionic species. The electronic and structural properties of these dyes were calculated by the semiempirical AM1 method. The results indicate the predominance of electrostatic interactions in dye-cellulose binding. The dominant contribution of the HOMO orbital molecular energy, used as descriptor, can be explained by the donor ability of the dye molecules in the dye adsorption on cellulose. PMID- 12132880 TI - Novel methods for the prediction of logP, pK(a), and logD. AB - Novel methods for predicting logP, pK(a), and logD values have been developed using data sets (592 molecules for logP and 1029 for pK(a)) containing a wide range of molecular structures. An equation with three molecular properties (polarizability and partial atomic charges on nitrogen and oxygen) correlates highly with logP (r2 = 0.89). The pK(a)s are estimated for both acids and bases using a novel tree structured fingerprint describing the ionizing centers. The new models have been compared with existing models and also experimental measurements on test sets of common organic compounds and pharmaceutical molecules. PMID- 12132882 TI - Assessment of the macrocyclic effect for the complexation of crown-ethers with alkali cations using the Substructural Molecular Fragments method. AB - The Substructural Molecular Fragments method (Solov'ev, V. P.; Varnek, A. A.; Wipff, G. J. Chem. Inf. Comput. Sci. 2000, 40, 847-858) was applied to assess stability constants (logK) of the complexes of crown-ethers, polyethers, and glymes with Na+, K+, and Cs+ in methanol. One hundred forty-seven computational models including different fragment sets coupled with linear or nonlinear fitting equations were applied for the data sets containing 69 (Na+), 123 (K+), and 31 (Cs+) compounds. To account for the "macrocyclic effect" for crown-ethers, an additional "cyclicity"descriptor was used. "Predicted" stability constants both for macrocyclic compounds and for their open-chain analogues are in good agreement with the experimental data reported earlier and with those studied experimentally in this work. The macrocyclic effect as a function of cation and ligand is quantitatively estimated for all studied crown-ethers. PMID- 12132881 TI - Improving the predicting power of partial order based QSARs through linear extensions. AB - Partial order theory (POT) is an attractive and operationally simple method that allows ordering of compounds, based on selected structural and/or electronic descriptors (modeled order), or based on their end points, e.g., solubility (experimental order). If the modeled order resembles the experimental order, compounds that are not experimentally investigated can be assigned a position in the model that eventually might lead to a prediction of an end-point value. However, in the application of POT in quantitative structure-activity relationship modeling, only the compounds directly comparable to the noninvestigated compounds are applied. To explore the possibilities of improving the methodology, the theory is extended by application of the so-called linear extensions of the model order. The study show that partial ordering combined with linear extensions appears as a promising tool providing probability distribution curves in the range of possible end-point values for compounds not being experimentally investigated. PMID- 12132883 TI - Prediction of ultraviolet spectral absorbance using quantitative structure property relationships. AB - High performance liquid chromatography (HPLC) with ultraviolet (UV) spectrophotometric detection is a common method for analyzing reaction products in organic chemistry. This procedure would benefit from a computational model for predicting the relative response of organic molecules. Models are now reported for the prediction of the integrated UV absorbance for a diverse set of organic compounds using a quantitative structure-property relationship (QSPR) approach. A seven-descriptor linear correlation with a squared correlation coefficient (R2) of 0.815 is reported for a data set of 521 compounds. Using the sum of ZINDO oscillator strengths in the integration range as an additional descriptor allowed reduction in the number of descriptors producing a robust model for 460 compounds with five descriptors and a squared correlation coefficient 0.857. The descriptors used in the models are discussed with respect to the physical nature of the UV absorption process. PMID- 12132885 TI - Modeling large macromolecular structures using promolecular densities. AB - A procedure to easily construct fitted density functions is presented. This methodology, based on the promolecule approach, is able to handle large macromolecular systems, such as proteins. The usual procedure dealing with fitted densities has been improved by adding some restrictions, which allow faster calculations. As a main application example, molecular isodensity contours (MIDCOs) are constructed for two proteins, one of them composed of more than 50 000 atoms. MIDCOs, as a visual representation of the molecular density function, and thus an important descriptor of the molecular charge distribution, constitute a powerful tool in the understanding of molecular systems. MIDCOs are presented for both proteins, allowing exploration of their surfaces, as well as analysis of their shapes. Also, as a quantum mechanical calculation example, molecular quantum self-similarity measures are calculated for several proteins. PMID- 12132886 TI - Enthalpy of the gas-phase CO2 + Mg reaction from ab initio total energies. AB - Various highly accurate ab initio composite methods of Gaussian-n (G1, G2, G3), their variations (G2(MP2), G3(MP2), G3//B3LYP, G3(MP2)//B3LYP), and complete basis set (CBS-Q, CBS-Q//B3LYP) series of models were applied to compute reaction enthalpies of the ground-state reaction of CO2 with Mg. All model chemistries predict highly endothermic reactions, with DeltaH(298) = 63.6-69.7 kcal x mol( 1). The difference between the calculated reaction enthalpies and the experimental value, evaluated with recommended experimental standard enthalpies of formation for products and reactants, is more than 20 kcal x mol(-1) for all methods. This difference originates in the incorrect experimental enthalpy of formation of gaseous MgO given in thermochemical databases. When the theoretical formation enthalpy for MgO calculated by a particular method is used, the deviation is reduced to 1.3 kcal x mol(-1). The performance of the methodologies used to calculate the heat of this particular reaction and the enthalpy of formation of MgO are discussed. PMID- 12132884 TI - MTD-PLS: A PLS-based variant of the MTD method. 2. Mapping ligand-receptor interactions. Enzymatic acetic acid esters hydrolysis. AB - The PLS variant of the MTD method (T. I. Oprea et al., SAR QSAR Environ. Res. 2001, 12, 75-92) was applied to a series of 25 acetylcholinesterase hydrolysis substrates. Statistically significant MTD-PLS models (q(2) between 0.7 and 0.8) are in agreement with previous MTD models, with the advantage that local contributions are understood beyond the occupancy/nonoccupancy interpretation in MTD. A "chemically intuitive" approach further forces MTD-PLS coefficients to assume only negative (or zero) values for fragmental volume descriptors and positive (or zero) values for fragmental hydrophobicity descriptors. This further separates the various kinds of local interactions at each vertex of the MTD hypermolecule, making this method suitable for medicinal chemistry synthesis planning. PMID- 12132887 TI - Novel atomic-level-based AI topological descriptors: application to QSPR/QSAR modeling. AB - Novel atomic level AI topological indexes based on the adjacency matrix and distance matrix of a graph is used to code the structural environment of each atomic type in a molecule. These AI indexes, along with Xu index, are successfully extended to compounds with heteroatoms in terms of novel vertex degree v(m), which is derived from the valence connectivity delta(v) of Kier-Hall to resolve the differentiation of heteroatoms in molecular graphs. The multiple linear regression (MLR) is used to develop the structure-property/activity models based on the modified Xu and AI indices. The efficiency of these indices is verified by high quality QSPR/QSAR models obtained for several representative physical properties and biological activities of several data sets of alcohols with a wide range of non-hydrogen atoms. The results indicate that the physical properties studied are dominated by molecular size, but other atomic types or groups have small influences dependent on the studied properties. Among all atomic types, -OH groups seem to be most important due to hydrogen-bonding interactions. On the contrary, -OH groups play a dominant role in biological activities studied, although molecular size is also an important factor. These results indicate that both Xu and AI indices are useful model parameters for QSPR/QSAR analysis of complex compounds. PMID- 12132888 TI - Structure-based classification of antibacterial activity. AB - The aim of this study was to develop a simple quantitative structure-activity relationship (QSAR) for the classification and prediction of antibacterial activity, so as to enable in silico screening. To this end a database of 661 compounds, classified according to whether they had antibacterial activity, and for which a total of 167 physicochemical and structural descriptors were calculated, was analyzed. To identify descriptors that allowed separation of the two classes (i.e. those compounds with and without antibacterial activity), analysis of variance was utilized and models were developed using linear discriminant and binary logistic regression analyses. Model predictivity was assessed and validated by the random removal of 30% of the compounds to form a test set, for which predictions were made from the model. The results of the analyses indicated that six descriptors, accounting for hydrophobicity and inter- and intramolecular hydrogen bonding, provided excellent separation of the data. Logistic regression analysis was shown to model the data slightly more accurately than discriminant analysis. PMID- 12132889 TI - Chemical information based scaling of molecular descriptors: a universal chemical scale for library design and analysis. AB - Scaling is a difficult issue for any analysis of chemical properties or molecular topology when disparate descriptors are involved. To compare properties across different data sets, a common scale must be defined. Using several publicly available databases (ACD, CMC, MDDR, and NCI) as a basis, we propose to define chemically meaningful scales for a number of molecular properties and topology descriptors. These chemically derived scaling functions have several advantages. First, it is possible to define chemically relevant scales, greatly simplifying similarity and diversity analyses across data sets. Second, this approach provides a convenient method for setting descriptor boundaries that define chemically reasonable topology spaces. For example, descriptors can be scaled so that compounds with little potential for biological activity, bioavailability, or other drug-like characteristics are easily identified as outliers. We have compiled scaling values for 314 molecular descriptors. In addition the 10th and 90th percentile values for each descriptor have been calculated for use in outlier filtering. PMID- 12132890 TI - Median Partitioning: a novel method for the selection of representative subsets from large compound pools. AB - A method termed Median Partitioning (MP) has been developed to select diverse sets of molecules from large compound pools. Unlike many other methods for subset selection, the MP approach does not depend on pairwise comparison of molecules and can therefore be applied to very large compound collections. The only time limiting step is the calculation of molecular descriptors for database compounds. MP employs arrays of property descriptors with little correlation to divide large compound pools into partitions from which representative molecules can be selected. In each of n subsequent steps, a population of molecules is divided into subpopulations above and below the median value of a property descriptor until a desired number of 2n partitions are obtained. For descriptor evaluation and selection, an entropy formulation was embedded in a genetic algorithm. MP has been applied here to generate a subset of the Available Chemicals Directory, and the results have been compared with cell-based partitioning. PMID- 12132891 TI - Graphic representation of configuration in two-dimensional space. Current conventions, clarifications, and proposed extensions. AB - The chemical vocabulary can describe, in a single term, a racemate or a specific enantiomer or a compound that is enantiopure, but whose absolute configuration is unknown. Regardless of these differentiations, the corresponding conventional graphic representations display but one specific enantiomer. Because of this inflexibility, verbal annotations to stereostructures in chemical publications and databases are unavoidable. In the expanding era of chirotechnology, the limited descriptive power of stereostructures in two-dimensional space is of serious concern. To provide a solution to this problem, targeted redeployment of established stereobonds serve as stereodescriptors that differentiate between enantiopure and racemic compounds and those that are enantiopure but whose chirality sense is unknown. Graphic displays then share the explicitness and accuracy of the chemical vocabulary. Fischer projections, recently expanded to accommodate alkenes and axially stereogenic compounds, can display favorably comparative configurational aspects of molecules with a multiplicity of stereogenic units. PMID- 12132892 TI - On the use of neural network ensembles in QSAR and QSPR. AB - Despite their growing popularity among neural network practitioners, ensemble methods have not been widely adopted in structure-activity and structure-property correlation. Neural networks are inherently unstable, in that small changes in the training set and/or training parameters can lead to large changes in their generalization performance. Recent research has shown that by capitalizing on the diversity of the individual models, ensemble techniques can minimize uncertainty and produce more stable and accurate predictors. In this work, we present a critical assessment of the most common ensemble technique known as bootstrap aggregation, or bagging, as applied to QSAR and QSPR. Although aggregation does offer definitive advantages, we demonstrate that bagging may not be the best possible choice and that simpler techniques such as retraining with the full sample can often produce superior results. These findings are rationalized using Krogh and Vedelsby's decomposition of the generalization error into a term that measures the average generalization performance of the individual networks and a term that measures the diversity among them. For networks that are designed to resist over-fitting, the benefits of aggregation are clear but not overwhelming. PMID- 12132893 TI - Using molecular equivalence numbers to visually explore structural features that distinguish chemical libraries. AB - A molecular equivalence number (meqnum) classifies a molecule with respect to a class of structural features or topological shapes such as its cyclic system or its set of functional groups. Meqnums can be used to organize molecular structures into nonoverlapping, yet highly relatable classes. We illustrate the construction of some different types of meqnums and present via examples some methods of comparing diverse chemical libraries based on meqnums. In the examples we compare a library which is a random sample from the MDL Drug Data Report (MDDR) with a library which is a random sample from the Available Chemical Directory (ACD). In our analyses, we discover some interesting features of the topological shape of a molecule and its set of functional groups that are strongly linked with compounds occurring in the MDDR but not in the ACD. We also illustrate the utility of molecular equivalence indices in delineating the structural domain over which an SAR conclusion is valid. PMID- 12132894 TI - Genetic Algorithm guided Selection: variable selection and subset selection. AB - A novel Genetic Algorithm guided Selection method, GAS, has been described. The method utilizes a simple encoding scheme which can represent both compounds and variables used to construct a QSAR/QSPR model. A genetic algorithm is then utilized to simultaneously optimize the encoded variables that include both descriptors and compound subsets. The GAS method generates multiple models each applying to a subset of the compounds. Typically the subsets represent clusters with different chemotypes. Also a procedure based on molecular similarity is presented to determine which model should be applied to a given test set compound. The variable selection method implemented in GAS has been tested and compared using the Selwood data set (n = 31 compounds; v = 53 descriptors). The results showed that the method is comparable to other published methods. The subset selection method implemented in GAS has been first tested using an artificial data set (n = 100 points; v = 1 descriptor) to examine its ability to subset data points and second applied to analyze the XLOGP data set (n = 1831 compounds; v = 126 descriptors). The method is able to correctly identify artificial data points belonging to various subsets. The analysis of the XLOGP data set shows that the subset selection method can be useful in improving a QSAR/QSPR model when the variable selection method fails. PMID- 12132896 TI - An ontology for pharmaceutical ligands and its application for in silico screening and library design. AB - Annotation efforts in biosciences have focused in past years mainly on the annotation of genomic sequences. Only very limited effort has been put into annotation schemes for pharmaceutical ligands. Here we propose annotation schemes for the ligands of four major target classes, enzymes, G protein-coupled receptors (GPCRs), nuclear receptors (NRs), and ligand-gated ion channels (LGICs), and outline their usage for in silico screening and combinatorial library design. The proposed schemes cover ligand functionality and hierarchical levels of target classification. The classification schemes are based on those established by the EC, GPCRDB, NuclearDB, and LGICDB. The ligands of the MDL Drug Data Report (MDDR) database serve as a reference data set of known pharmacologically active compounds. All ligands were annotated according to the schemes when attribution was possible based on the activity classification provided by the reference database. The purpose of the ligand-target classification schemes is to allow annotation-based searching of the ligand database. In addition, the biological sequence information of the target is directly linkable to the ligand, hereby allowing sequence similarity-based identification of ligands of next homologous receptors. Ligands of specified levels can easily be retrieved to serve as comprehensive reference sets for cheminformatics-based similarity searches and for design of target class focused compound libraries. Retrospective in silico screening experiments within the MDDR01.1 database, searching for structures binding to dopamine D2, all dopamine receptors and all amine-binding class A GPCRs using known dopamine D2 binding compounds as a reference set, have shown that such reference sets are in particular useful for the identification of ligands binding to receptors closely related to the reference system. The potential for ligand identification drops with increasing phylogenetic distance. The analysis of the focus of a tertiary amine based combinatorial library compared to known amine binding class A GPCRs, peptide binding class A GPCRs, and LGIC ligands constitutes a second application scenario which illustrates how the focus of a combinatorial library can be treated quantitatively. The provided annotation schemes, which bridge chem- and bioinformatics by linking ligands to sequences, are expected to be of key utility for further systematic chemogenomics exploration of previously well explored target families. PMID- 12132895 TI - A method for correlations analysis of coordinates: applications for molecular conformations. AB - We describe a new method to analyze multiple correlations between subsets of coordinates that represent a sample. The correlation is established only between specific regions of interest at the coordinates. First, the region(s) of interest are selected at each molecular coordinate. Next, a correlation matrix is constructed for the selected regions. The matrix is subject to further analysis, illuminating the multidimensional structural characteristics that exist in the conformational space. The method's abilities are demonstrated in several examples: it is used to analyze the conformational space of complex molecules, it is successfully applied to compare related conformational spaces, and it is used to analyze a diverse set of protein folding trajectories. PMID- 12132897 TI - On the parametrization of the toxicity of organic chemicals to Tetrahymena pyriformis. The problem of establishing a uniform activity. AB - In this report we illustrate the importance of making an effort to ensure that all of the dependent variables in a QSAR are associated with one mechanism of action. If this cannot be established, it will not be possible to compare the QSAR with others acting on different biological systems. That is, such information cannot be used to develop a science of chemical-biological interactions. A study of the action of a large set of phenols acting on Tetrahymena pyriformis made by Cronin and Schultz is analyzed to illustrate the problem. PMID- 12132898 TI - Molecular design based on 3D-pharmacophore. Application to 5-HT4 receptor. AB - A definition of a pharmacophore for the 5-HT4 antagonist was carried out by considering a three-dimensional model which correlates the chemical structures of series of antagonists with their biological affinities. A molecular design is described by analyzing the differences between two 3D serotonin pharmacophores. This successful structural modification demonstrates the efficiency of this approach to design new serotonin ligands. PMID- 12132899 TI - Cassette dosing approach and quantitative structure-pharmacokinetic relationship study of antifungal N-myristoyltransferase inhibitors. AB - Pharmacokinetic (PK) parameters of N-myristoyltransferase (Nmt) inhibitors were measured, and a multivariate quantitative structure-pharmacokinetic relationship (QSPKR) model for predicting rat elimination half-life (t(1/2)) values was constructed. One hundred seven benzofuran derivatives have been selected as the data set for QSPKR analysis. The correlation between the t(1/2) values and 30 physicochemical descriptors was examined by a stepwise multiple linear regression method. The statistical analysis gives a significant QSPKR model (r = 0.843) with the following three variables: partial negative surface area (PNSA), atomic-based octanol/water partition coefficient (AlogP), and the number of rotational bonds (Rotlbonds). The QSPKR model obtained is predictive and simple, and would give a direction for designing new Nmt inhibitors having good PK profiles. PMID- 12132900 TI - The interrelation of physicochemical parameters and topological descriptors for a series of beta-blocking agents. AB - The intercorrelation between a series of physicochemical parameters and topological indices for a set of beta-blockers is investigated. Partition coefficients are calculated using the ClogP program, and the results are compared with previous data, both experimental and theoretical. These data are complemented by hydrophilicity and solubility calculations, together with the determination of molecular area and volume. Connectivity indices, of order 1 and 2, including simple, valence, and differential terms, are evaluated. The derivation of a recently proposed topological descriptor, the eccentric adjacency index, from the adjacency and distance matrices, is presented. The corresponding valence term, a novel descriptor, is developed, and other indices related to the distance matrix, the Wiener and Hyper-Wiener terms, are included. A high degree of linear correlation between the connectivity indices is noted. The correlations for first-order terms are slightly superior to the corresponding second-order values. This is particularly true when considering the valence terms compared with the nonvalence terms. The relationship between these terms and reported pharmacological properties are investigated. A decrease in the eccentric adjacency index resulted in an increase in the pharmacological property. PMID- 12132901 TI - Mechanistic insight from a volume profile for electron transfer between promazine and hexaaquairon(III). AB - The kinetics of the outer-sphere electron-transfer reaction between promazine (dimethyl-(3-phenothiazin-10-yl-propyl)-amine) and hexaaquairon(III) was studied using a high-pressure stopped-flow technique. The effect of pressure (over the range 0.1-130 MPa at 25 degrees C and ionic strength 1.0 M) on the reaction rate in aqueous perchloric acid solution resulted in volumes of activation of -6.2 +/- 0.4 and -12.5 +/- 0.5 cm(3) mol(-)(1) for the forward and reverse processes, respectively. The effect of pressure on the overall equilibrium constant revealed a reaction volume of +5.0 +/- 0.2 cm(3) mol(-)(1). The reported volume profile reveals mechanistic information on the electron-transfer process in terms of volume changes along the reaction coordinate. The volume of activation for the promazine/promazine(+*) self-exchange reaction was calculated on the basis of the Marcus cross relationship. PMID- 12132902 TI - Anionic templating: synthesis, structure, and characterization of novel three dimensional mixed-metal oxychlorides Te(4)M(3)O(15).Cl (M = Nb(5+) or Ta(5+)). AB - Two new three-dimensional oxychlorides are reported, Te(4)M(3)O(15).Cl (M = Nb(5+) or Ta(5+)). The isostructural materials were synthesized by chemical transport reactions utilizing TeO(2), M(2)O(5), and MCl(5) (M = Nb(5+) or Ta(5+)) as reagents. The compounds exhibit a three-dimensional cationic tunnel framework, with Cl(-) anions occupying the tunnels. Crystal data: monoclinic, space group C2/c, a = 18.9944(7) A, b = 7.8314(3) A, c = 21.1658(8) A, beta = 116.6400(10) degrees, Z = 8 (T = 295 K). PMID- 12132903 TI - DNA photocleavage by a supramolecular Ru(II)-viologen complex. AB - A novel Ru(II) complex possessing two sequentially linked viologen units, Ru-V(1) V(2)(6+), was synthesized and characterized. Upon excitation of the Ru(II) unit (lambda(exc) = 532 nm, fwhm approximately 10 ns), a long-lived charge-separated (CS) state is observed (tau = 1.7 micros) by transient absorption spectroscopy. Unlike Ru(bpy)(3)(2+), which cleaves DNA upon photolysis through the formation of reactive oxygen species, such as (1)O(2) and O(2)(-), the photocleavage of plasmid DNA by Ru-V(1)-V(2)(6+) is observed both in air and under N(2) atmosphere (lambda(irr) > 395 nm). PMID- 12132904 TI - Di- and tricobalt Dawson sandwich complexes: synthesis, spectroscopic characterization, and electrochemical behavior of Na(18)[(NaOH(2))(2)Co(2)(P(2)W(15)O(56))(2)] and Na(17)[(NaOH(2))Co(3)(H(2)O)(P(2)W(15)O(56))(2)]. AB - The reaction of the trivacant Dawson anion alpha-[P(2)W(15)O(56)](12-) and the divalent cations Co(2+) is known to form the tetracobalt sandwich complex [Co(4)(H(2)O)(2)(P(2)W(15)O(56))(2)](16-) (Co(4)P(4)W(30)). Two new complexes, with different Co/P(2)W(15) stoichiometry, [(NaOH(2))(2)Co(2)(P(2)W(15)O(56))(2)](18-) (Na(2)Co(2)P(4)W(30)) and [(NaOH(2))Co(3)(H(2)O)(P(2)W(15)O(56))(2)](17-) (NaCo(3)P(4)W(30)), have been synthesized as aqueous-soluble sodium salts, by a slight modification of the reaction conditions. Both compounds were characterized by IR, elemental analysis, and (31)P solution NMR spectroscopy. These species are "lacunary" sandwich complexes, which add Co(2+) cations according to Na(2)Co(2)P(4)W(30) + Co(2+) --> NaCo(3)P(4)W(30) + Na(+) followed by NaCo(3)P(4)W(30) + Co(2+) --> Co(4)P(4)W(30) + Na(+). A Li(+)/Na(+) exchange in the cavity was evidenced by (31)P dynamic NMR spectroscopy. The electrochemical behaviors of the sandwich complexes [(NaOH(2))Co(3)(H(2)O)(P(2)W(15)O(56))(2)](17-) and [(NaOH(2))(2)Co(2)(P(2)W(15)O(56))(2)](18-) were investigated in aqueous solutions and compared with that of [Co(4)(H(2)O)(2)(P(2)W(15)O(56))(2)](16-). These complexes showed an electrocatalytic effect on nitrite reduction. PMID- 12132905 TI - Ligand substitution, pH dependent deoxygenation, and linkage isomerization reactions of the 2,2'-bipyridinetetranitroruthenate dianion. AB - The reaction of the [Ru(bpy)(NO(2))(4)](2-) (bpy = 2,2'-bipyridine) ion in aqueous solutions produces two different nitrosyl complexes, depending on the pH of the solution. At acidic pH, complex cis,cis-Ru(bpy)(NO(2))(2)(ONO)(NO) was isolated. At neutral or basic pH, [Ru(bpy)(NO(2))(4)](2-) reacts to give cis,trans-Ru(bpy)(NO(2))(2)(NO)(OH). Both new complexes were fully characterized by elemental analysis and UV-vis, IR, (1)H NMR, and (15)N NMR spectroscopy. A single-crystal X-ray structure of cis,trans-Ru(bpy)(NO(2))(2)(NO)(OH) was also obtained. cis,cis-Ru(bpy)(NO(2))(2)(ONO)(NO) isomerizes in acetone or water solution to give a mixture of the trans,cis-Ru(bpy)(NO(2))(2)(ONO)(NO) and cis,cis-Ru(bpy)(ONO)(2)(NO(2))(NO) linkage isomers as determined by (1)H and (15)N NMR spectroscopy. A single-crystal X-ray structure of a solid solution of cis,cis-Ru(bpy)(ONO)(2)(NO(2))(NO)/trans,cis-Ru(bpy)(NO(2))(2)(ONO)(NO) was also obtained. This pair of isomers is the first crystallographically characterized compound with nitro, nitrito, and nitrosyl ligands. The kinetic studies of the Ru NO(2) --> Ru-NO conversion reactions of [Ru(bpy)(NO(2))(4)](2)(-) in buffered solutions from pH 3 to pH 9 complement previous studies of the reverse reaction. The reactions are first order in [Ru(bpy)(NO(2))(4)](2-). At high pH, the reaction is independent of the concentration of H(+) while, at low pH, the reaction is first order in the concentration of H(+). The rate determining step of the high pH reaction involves breakage of the Ru-NO(2) bond while, at low pH, the mechanism involves a rapid reversible protonation of a NO(2) ligand followed by the rate determining loss of hydroxide to produce a nitrosyl ligand. PMID- 12132906 TI - Di- and tetranuclear ruthenium(II) and/or osmium(II) complexes of polypyridyl ligands bridged by a fully conjugated aromatic spacer: synthesis, characterization, and electrochemical and photophysical studies. AB - A series of mono-, di-, and tetranuclear homo/heterometallic complexes of Ru(II) and Os(II) based on the bridging ligand dppz(11-11')dppz (where dppz = dipyrido[3,2-a:2',3'-c]phenazine) (BL) have been synthesized and characterized. This bridging ligand is a long rigid rod with only one rotational degree of freedom and provides complete conjugation between the chromophores. The complexes synthesized are of general formula [(bpy)(2)Ru-BL](2+), [(phen)(2)/(bpy)(2)M-BL M(bpy)(2)/(phen)(2)](4+) (M = Ru(II) and Os(II)), [(bpy)(2)Ru-BL-Os(bpy)(2)](4+), and [((bpy)(2)Ru-BL)(3)M](8+). Detailed (1)H NMR studies of these complexes revealed that each chiral center does not influence its neighbor because of the long distance between the metal centers and the superimposed resonances of the diastereoisomers, which allowed the unambiguous assignment of the signals, particularly for homonuclear complexes. Concentration-dependent (1)H NMR studies show molecular aggregation of the mono- and dinuclear complexes in solution by pi pi stacking. Electrospray mass spectrometry data are consistent with dimerization of mono- and dinuclear complexes in solution. Electrochemical studies show oxidations of Ru(II) and Os(II) in the potential ranges +1.38 to +1.40 and +0.92 to +1.01 V, respectively. The bridging ligand exhibits two one-electron reductions, and it appears that the added electrons are localized on the phenazene moieties of the spacer. All of these complexes show strong metal-to ligand charge-transfer (MLCT) absorption and (3)MLCT luminescence at room temperature. Quantum yields have been calculated, and the emission lifetimes of all complexes have been measured by laser flash photolysis experiments. The luminescence intensity and lifetime data suggest that the emission due to the Ru center of the heteronuclear complexes is strongly quenched (>90%) compared to that of the corresponding model complexes. This quenching is attributed to intramolecular energy transfer from the Ru(II) center to the Os(II) center (k = (3-5) x 10(7) s(-1)) across the bridging ligand. PMID- 12132907 TI - Synthesis and structures of aluminum alkanethiolate complexes. AB - The homoleptic aluminum thiolate complex [Al(mu-S-t-Bu)(S-t-Bu)(2)](2) was prepared by reacting AlBr(3) with NaS-t-Bu while the analogous 2-propanethiolate complex [Al(mu-S-i-Pr)(S-i-Pr)(2)](2) was synthesized by reacting AlH(3)(OEt(2)) with i-PrSH. In the solid state, the dimers have tetrahedral Al atoms and anti Al(mu-SR)(2)Al four-member rings. The attempted synthesis of [Al(mu-S-t-Bu)(S-t Bu)(2)](2) by reacting Al(N-i-Pr(2))(3) with t-BuSH in THF solvent yielded the thermally stable THF adduct Al(S-t-Bu)(3)(THF). The same reaction in diethyl ether solvent produced a mixture of [Al(mu-mgr;-S-t-Bu)(S-t-Bu)(2)](2) and the salt [i-Pr(2)NH(2)][Al(S-t-Bu)(4)]. In the solid-state structure of the salt, the anion [Al(S-t-Bu)(4)](-) has a distorted tetrahedral geometry. Reactions of [Al(NMe(2))(3)](2) and AlH(3)(NMe(2)Et) with the alkanethiols yielded stable amine adducts Al(SR)(3)(R'NMe(2)) (R = i-Pr or t-Bu; R' = H or Et). The ligand adducts Al(S-i-Pr)(3)(HNMe(2)) and Al(S-t-Bu)(3)(THF) have distorted trigonal pyramidal geometries in the solid state. Three of the new compounds, [Al(mu-S-i Pr)(S-i-Pr)(2)](2) and Al(SR)(3)(HNMe(2)) (R = i-Pr or t-Bu), are viable precursor candidates for the chemical vapor deposition of aluminum sulfide films because they are thermally stable, volatile liquids at moderate temperatures. PMID- 12132908 TI - A new dinuclear vanadium(V)-citrate complex from aqueous solutions. Synthetic, structural, spectroscopic, and pH-dependent studies in relevance to aqueous vanadium(V)-citrate speciation. AB - Vanadium interactions with low molecular mass binders in biological fluids entail the existence of vanadium species with variable chemical and biological properties. In the course of efforts to elucidate the chemistry related to such interactions, we have explored the oxidative chemistry of vanadium(III) with the physiologically relevant tricarboxylic citric acid. Aqueous reactions involving VCl(3) and anhydrous citric acid, at pH approximately 7, resulted in blue solutions. Investigation into the nature of the species arising in those solutions revealed, through UV/visible and EPR spectroscopies, oxidation of vanadium(III) to vanadium(IV). Further addition of H(2)O(2) resulted in the oxidation of vanadium(IV) to vanadium(V), and the isolation of a new vanadium(V) citrate complex in the form of its potassium salt. Analogous reactions with K(4)[V(2)O(2)(C(6)H(4)O(7))(2)].6H(2)O and H(2)O(2) or V(2)O(5) and citrate at pH approximately 5.5 afforded the same material. Elemental analysis pointed to the molecular formulation K(4)[V(2)O(4)(C(6)H(5)O(7))(2)].5.6H(2)O (1). Complex 1 was further characterized by FT-IR and X-ray crystallography. 1 crystallizes in the triclinic space group P(-)1, with a = 11.093(4) A, b = 9.186(3) A, c = 15.503(5) A, alpha = 78.60(1) degrees, beta = 86.16(1) degrees, gamma = 69.87(1) degrees, V = 1454.0(8) A(3), and Z = 2. The X-ray structure of 1 reveals the presence of a dinuclear vanadium(V)-citrate complex containing a V(V)(2)O(2) core. The citrate ligands are triply deprotonated, and as such they bind to vanadium(V) ions, thus generating a distorted trigonal bipyramidal geometry. Binding occurs through the central alkoxide and carboxylate groups, with the remaining two terminal carboxylates being uncoordinated. One of those carboxylates is protonated and contributes to hydrogen bond formation with the deprotonated terminal carboxylate of an adjacent molecule. Therefore, an extended network of hydrogen-bonded V(V)(2)O(2)-core-containing dimers is created in the lattice of 1. pH-dependent transformations of 1 in aqueous media suggest its involvement in a web of vanadium(V)-citrate dinuclear species, consistent with past solution speciation studies investigating biologically relevant forms of vanadium. PMID- 12132909 TI - Reaction of bis(phosphine)(hydrotris(3,5-dimethylpyrazolyl)borato)rhodium(I) with phenylacetylene, p-nitrobenzaldehyde, and triphenyltin hydride: structures of [Rh(Tp)(PPh(3))(2)], [Rh(Tp)(H)(C(2)Ph)(P(4-C(6)H(4)F)(3))], [Rh(Tp)(H)(COC(6)H(4)-4-NO(2))(PPh(3))], and [Rh(Tp)(H)(SnPh(3))(PPh(3))]. AB - The complexes [Rh(Tp)(PPh(3))(2)] (1a) and [Rh(Tp)(P(4-C(6)H(4)F)(3))(2)] (1b) combine with PhC(2)H, 4-NO(2)-C(6)H(4)CHO and Ph(3)SnH to give [Rh(Tp)(H)(C(2)Ph)(PR(3))] (R = Ph, 2a; R = 4-C(6)H(4)F, 2b), [Rh(Tp)(H)(COC(6)H(4)-4-NO(2))(PR(3))] (R = Ph, 3a), and [Rh(Tp)(H)(SnPh(3))(PR(3))] (R = Ph, 4a; R = 4-C(6)H(4)F, 4b) in moderate to good yield. Complexes 1a, 2b, 3a, and 4a have been structurally characterized. In 1a the Tp ligand is bidentate, in 2b, 3a, and 4a it is tridentate. Crystal data for 1a: space group P2(1)/c; a = 11.9664(19), b = 21.355(3), c = 20.685(3) A; beta = 112.576(7) degrees; V = 4880.8(12) A(3); Z = 4; R = 0.0441. Data for 2b: space group P(-)1; a = 10.130(3), b = 12.869(4), c = 17.038(5) A; alpha = 78.641(6), beta = 76.040(5), gamma = 81.210(6) degrees; V = 2100.3(11) A(3); Z = 2; R = 0.0493. Data for 3a: space group P(-)1; a = 10.0073(11), b = 10.5116(12), c = 19.874(2) A; alpha = 83.728(2), beta = 88.759(2), gamma = 65.756(2) degrees; V =1894.2(4) A(3); Z = 2; R = 0.0253. Data for 4a: space group P2(1)/c; a = 15.545(2), b = 18.110(2), c = 17.810(2) A; beta = 95.094(3) degrees; V = 4994.1(10) A(3); Z = 4; R = 0.0256. NMR data ((1)H, (31)P, (103)Rh, (119)Sn) are also reported. PMID- 12132911 TI - Synthesis, coordination to Rh(I), and hydroformylation catalysis of new beta aminophosphines bearing a dangling nitrogen group: an unusual inversion of a Rh coordinated P center. AB - Variants of the beta-aminophosphine L(1) [Ph(2)PCH(2)CH(Ph)NHPh] containing additional nitrogen donor functions have been prepared. These functions are branched off the C atom adjacent to the P atom, or the P atom itself. Ligand [Ph(2)PCH(o-C(6)H(4)NMe(2))CH(Ph)NHPh] has been obtained as a mixture of two diastereomers L(3A) and L(3B) by lithiation of L(2) [Ph(2)PCH(2)(o C(6)H(4)NMe(2))] with n-BuLi followed by PhCH=NPh addition and hydrolysis. The diastereomers have been separated by fractional crystallization from ethanol. Ligand Et(2)NCH(2)P(Ph)CH(2)CH(Ph)NHPh has been obtained as a mixture of two diastereomers L(5A) and L(5B)(starting with P-Ph reductive cleavage of L(1) by lithium and subsequent hydrolysis to give PhP(H)CH(2)CH(Ph)NHPh (mixture of two diastereomers L(4A) and L(4B)). The latter reacts with diethylamine and formaldehyde to afford the L(5) diastereomeric mixture. Complexes RhCl(CO)(L) (L = L(3A), 1(A); L(3B), 1(B); L(5A/B), 2(A/B)) were obtained by reaction of [RhCl(CO)(2)](2) and the appropriate ligand or ligand mixture. Complexes 1(A), 1(B), and 2(A) have been isolated in pure form and characterized by classical techniques and by single-crystal X-ray diffraction. All structures exhibit a bidentate kappa-P,kappa-N(NHPh) mode similar to the complex containing L(1). While complexes 1(A) or 1(B) are stable in CDCl(3) solution, complex 2(A) slowly converts to its diastereomer 2(B). This unexpected epimerization appears to take place by inversion at the Rh-coordinated P center, an apparently unprecedented phenomenon. A mechanism based on a reversible P-C bond oxidative addition is proposed. The influence of the pendant nitrogen function of the diaminophosphines L(3A) and L(5A/B) on the rhodium catalytic activity in styrene hydroformylation has been examined and compared to that of the aminophosphines L(1) or L(2). The observed trends are related to the basicity of the dangling amine function and to its proximity to the metal center. PMID- 12132910 TI - Metal-metal interactions in dinuclear d(8) metal cyanide complexes supported by phosphine ligands. Spectroscopic properties and ab initio calculations of [M(2)(mu-diphosphine)(2)(CN)(4)] and trans-[M(phosphine)(2)(CN)(2)] (M = Pt, Ni). AB - Structural, spectroscopic properties on the dinuclear [M(2)(dcpm)(2)(CN)(4)] (M = Pt, 1a; Ni, 2a, dcpm = bis(dicyclohexylphosphino)methane) and [M(2)(dmpm)(2)(CN)(4)] (M = Pt, 1b; Ni, 2b, dmpm = bis(dimethylphosphino)methane) and the mononuclear trans-[M(PCy(3))(2)(CN)(2)] (M = Pt, 3; Ni, 4, PCy(3) = tricyclohexylphosphine) and theoretical investigations on the corresponding model compounds are described. X-ray structural analyses reveal Pt.Pt and Ni.Ni distances of 3.0565(4)/3.189(1) A and 2.957(1)/3.209(8) A for 1a/1b and 2a/2b, respectively. The UV-vis absorption bands at 337 nm (epsilon 2.41 x 10(4) dm(3) mol(-)(1) cm(-)(1)) for 1a and 328 nm (epsilon 2.43 x 10(4) dm(3) mol(-)(1) cm( )(1)) for 1b in CH(2)Cl(2) are assigned to (1)(5d(sigma) --> 6p(sigma)) electronic transitions originating from Pt(II)-Pt(II) interactions. Resonance Raman spectroscopy of 1a, in which all the Raman intensity appears in the Pt-Pt stretch fundamental (93 cm(-)(1)) and overtone bands, verifies this metal-metal interaction. Complexes 1a and 1b exhibit photoluminescence in the solid state and solution. For the dinuclear nickel(II) complexes 2a and 2b, neither spectroscopic data nor theoretical calculation suggests the presence of Ni(II)-Ni(II) interactions. The intense absorption bands at lambda > 320 nm in the UV-vis spectra of 2a and 2b are tentatively assigned to d --> d transitions. PMID- 12132912 TI - A homologous series of alkaline earth phosphanides: syntheses, crystal structures, and unusual dynamic behavior of (THF)(n)M[P(CH(SiMe(3))(2))(C(6)H(4) 2-CH(2)NMe(2))](2) (M = Mg, Ca, Sr, Ba). AB - The secondary phosphine R(Me(2)NCH(2)-2-C(6)H(4))PH reacts with Bu(2)Mg to give the homoleptic complex Mg[PR(C(6)H(4)-2-CH(2)NMe(2))](2) (1) [R = CH(SiMe(3))(2)]. The analogous heavier alkaline earth metal complexes (THF)(n)Ae[PR(C(6)H(4)-2-CH(2)NMe(2))](2) [Ae = Ca (2), n = 0; Ae = Sr (3), Ba (4), n = 1] have been synthesized by metathesis reactions between K[PR(C(6)H(4)-2 CH(2)NMe(2))] and 0.5 equiv of the respective alkaline earth metal diiodide. Compounds 1-4 have been characterized by X-ray crystallography and multielement NMR spectroscopy. In the solid state, compounds 1-4 are monomeric, complexes 1 and 2 adopting a distorted tetrahedral geometry and complexes 3 and 4 adopting a distorted square pyramidal geometry (1: orthorhombic, P2(1)2(1)2(1), a = 11.413(3) A, b = 12.072(3) A, c = 32.620(11) A, Z = 4. 2: monoclinic, P2(1)/c, a = 9.5550(4) A, b = 17.4560(7) A, c = 24.5782(10) A, beta = 91.673(2) degrees, Z = 4. 3: monoclinic, C2/c, a = 15.0498(9) A, b = 13.0180(8) A, c = 24.3664(14) A, beta = 104.593(2) degrees, Z = 4. 4: monoclinic, C2/c, a = 15.2930(10) A, b = 13.0326(9) A, c = 24.6491(17) A, beta = 105.542(2) degrees, Z = 4). In toluene solution, compounds 2-4 are subject to dynamic processes which are attributed to a monomer-dimer equilibrium for which bridge-terminal exchange of the phosphanide ligands in the dimer may be frozen out at low temperatures. PMID- 12132913 TI - Coordination of cyclo-octasulfur and cyclo-heptaselenium to dinuclear rhenium(I) systems. AB - By substitution reactions of the coordinated THF ligands of Re(2)(mu X)(2)(CO)(6)(THF)(2) by elemental chalcogens (S(8) and red selenium), the complexes Re(2)(mu-X)(2)(CO)(6)(S(8)) (X = Br, 1; I, 2), and Re(2)(mu X)(2)(CO)(6)(Se(7)), (X = I, 3; Br, 4) have been prepared. Binuclear compound 3 was crystallographically established to be a coordination compound of cyclo heptaselenium, two adjacent selenium atoms of the Se(7) ligand [Se-Se distance, 2.558(3) A] being bonded to rhenium(I), at an average Re-Se distance of 2.586(3) A, and the nonbonding Re.Re distance being 4.077(3) A. Spectroscopic evidence of the existence of these chalcogen complexes in solution is reported. The Re(2)(mu X)(2)(CO)(6)(S(8)) complexes undergo S(8) displacement by THF, while the coordinated Se(7) moiety is less readily displaced from 3. PMID- 12132914 TI - Dianionic complexes with hexacoordinate silicon(IV) or germanium(IV) and three bidentate ligands of the hydroximato(2-) type: syntheses and structural characterization in the solid state. AB - Reaction of Si(OMe)(4) with acetohydroxamic acid or benzohydroxamic acid and HNMe(2) (molar ratio 1:3:2) in MeCN yielded dimethylammonium fac tris[acetohydroximato(2-)]silicate (fac-5) and N,N-dimethylacetamidinium fac tris[benzohydroximato(2-)]silicate (fac-8), respectively. Reaction of Si(OMe)(4) with benzohydroxamic acid and HNMe(2) (molar ratio 1:3:2) or ethane-1,2-diamine (molar ratio 1:3:1) in MeOH gave dimethylammonium fac-tris[benzohydroximato(2 )]silicate-methanol (fac-6.MeOH) and ethane-1,2-diammonium mer tris[benzohydroximato(2-)]silicate-dimethanol (mer-9.2MeOH), respectively. Reaction of Ge(OMe)(4) with benzohydroxamic acid and HNMe(2) (molar ratio 1:3:2) in MeOH resulted in the formation of dimethylammonium fac-tris[benzohydroximato(2 )]germanate-methanol (fac-7.MeOH). Single-crystal X-ray diffraction studies showed that the Si(Ge)-coordination polyhedra of the racemic hexacoordinate silicon (germanium) compounds fac-5, fac-6.MeOH, fac-7.MeOH, fac-8, and mer 9.2MeOH are distorted octahedra. All compounds were additionally characterized by solid-state VACP/MAS NMR studies ((13)C, (15)N, (29)Si). The structural investigations were complemented by computational studies of the dianions of fac 5 and mer-5. PMID- 12132915 TI - Synthesis and characterization of three-coordinate and related beta-diketiminate derivatives of manganese, iron, and cobalt. AB - Treatment of M[N(SiMe(3))(2)](2) (M = Mn, Fe, Co) with various bulky beta diketimines afforded a variety of new three-coordinate complexes which were characterized by UV-vis, (1)H NMR and IR spectroscopy, magnetic measurements, and X-ray crystallography. Reaction of the beta-diketimine H(Dipp)NC(Me)CHC(Me)N(Dipp) (Dipp(2)N(wedge)NH; Dipp = C(6)H(3)-2,6-Pr(i)(2)) with M[N(SiMe(3))(2)](2) (M = Mn or Co) gave Dipp(2)N(wedge)NMN(SiMe(3))(2) (M = Mn, 1; Co, 3) while the reaction of Fe[N(SiMe(3))(2)](2) with Ar(2)N(wedge)NH (Ar = Dipp, C(6)F(5), Mes, C(6)H(3)-2,6-Me(2), or C(6)H(3)-2,6-Cl(2)) afforded the series of iron complexes Ar(2)N(wedge)NFe[N(SiMe(3))(2)] (Ar = Dipp, 2a; C(6)F(5), 2b; Mes, 2c; C(6)H(3)-2,6-Me(2), 2d; C(6)H(3)-2,6-Cl(2), 2e). This represents a new synthetic route to beta-diketiminate complexes of these metals. The four-coordinate bis-beta-diketiminate complex Fe[N(wedge)N(C(6)F(5))(2)](2), 4, was also isolated as a byproduct from the synthesis of 2b. Direct reaction of the Dipp(2)N(wedge)NLi with CoCl(2) gave the "ate" salt Dipp(2)N(wedge)NCoCl(2)Li(THF)(2), 5, in which the lithium chloride has formed a complex with Dipp(2)N(wedge)NCoCl through chloride bridging. The Fe(III) species Dipp(2)N(wedge)NFeCl(2), 6, was obtained cleanly from the reaction of FeCl(3) with Dipp(2)N(wedge)NLi. Magnetic measurements showed that all the complexes have a high spin configuration. The different substituents in the series of iron complexes 2a-e allowed assignment of their paramagnetically shifted (1)H NMR spectra. The X-ray crystal structures 1-2d and 3 showed that they have a distorted three-coordinate planar configuration at the metals whereas complexes 4 6 have highly distorted four-coordinate geometries. PMID- 12132916 TI - Catalysis of H(2)/D(2) scrambling and other H/D exchange processes by [Fe] hydrogenase model complexes. AB - Protonation of the [Fe]-hydrogenase model complex (mu-pdt)[Fe(CO)(2)(PMe(3))](2) (pdt = SCH(2)CH(2)CH(2)S) produces a species with a high field (1)H NMR resonance, isolated as the stable [(mu-H)(mu pdt)[Fe(CO)(2)(PMe(3))](2)](+)[PF(6)](-) salt. Structural characterization found little difference in the 2Fe2S butterfly cores, with Fe.Fe distances of 2.555(2) and 2.578(1) A for the Fe-Fe bonded neutral species and the bridging hydride species, respectively (Zhao, X.; Georgakaki, I. P.; Miller, M. L.; Yarbrough, J. C.; Darensbourg, M. Y. J. Am. Chem. Soc. 2001, 123, 9710). Both are similar to the average Fe.Fe distance found in structures of three Fe-only hydrogenase active site 2Fe2S clusters: 2.6 A. A series of similar complexes (mu-edt)-, (mu-o xyldt)-, and (mu-SEt)(2)[Fe(CO)(2)(PMe(3))](2) (edt = SCH(2)CH(2)S; o-xyldt = SCH(2)C(6)H(4)CH(2)S), (mu-pdt)[Fe(CO)(2)(PMe(2)Ph)](2), and their protonated derivatives likewise show uniformity in the Fe-Fe bond lengths of the neutral complexes and Fe.Fe distances in the cationic bridging hydrides. The positions of the PMe(3) and PMe(2)Ph ligands are dictated by the orientation of the S-C bonds in the (mu-SRS) or (mu-SR)(2) bridges and the subsequent steric hindrance of R. The Fe(II)(mu-H)Fe(II) complexes were compared for their ability to facilitate H/D exchange reactions, as have been used as assays of H(2)ase activity. In a reaction that is promoted by light but inhibited by CO, the [(mu-H)(mu pdt)[Fe(CO)(2)(PMe(3))](2)](+) complex shows H/D exchange activity with D(2), producing [(mu-D)(mu-pdt)[Fe(CO)(2)(PMe(3))](2)](+) in CH(2)Cl(2) and in acetone, but not in CH(3)CN. In the presence of light, H/D scrambling between D(2)O and H(2) is also promoted by the Fe(II)(mu-H)Fe(II) catalyst. The requirement of an open site suggests that the key step in the reactions involves D(2) or H(2) binding to Fe(II) followed by deprotonation by the internal hydride base, or by external water. As indicated by similar catalytic efficiencies of members of the series, the nature of the bridging thiolates has little influence on the reactions. Comparison to [Fe]H(2)ase enzyme active site redox levels suggests that at least one Fe(II) must be available for H(2) uptake while a reduced or an electron-rich Fe(I)Fe(I) metal-metal bonded redox level is required for proton uptake. PMID- 12132917 TI - Synthesis, crystal structure, and solid state NMR spectroscopy of NH(4)[(V(2)O(3))(2)(4,4'-bpy)(2)(H(2)PO(4))(PO(4))(2)].0.5H(2)O, a mixed-valence vanadium(IV,V) phosphate with a pillared layer structure. AB - A mixed-valence vanadium phosphate, NH(4)[(V(2)O(3))(2)(4,4' bpy)(2)(H(2)PO(4))(PO(4))(2)].0.5H(2)O, has been synthesized under hydrothermal conditions and structurally characterized by single-crystal X-ray diffraction. It crystallizes in the monoclinic space group C2/c (No. 15) with a = 12.6354(8) A, b = 9.9786(6) A, c = 23.369(1) A, beta = 92.713(1) degrees, and Z = 4 with R(1) = 0.0389. The structure consists of dimers of edge-sharing vanadium(IV,V) octahedra that are connected by corner-sharing phosphate tetrahedra to form layers in the ab-plane, which are further linked through 4,4'-bipyridine pillars to generate a 3-D framework. Magnetic susceptibility confirms the valence of the vanadium atoms. The (31)P MAS NMR spectrum shows a resonance centered at 80 ppm with a shoulder at ca. 83 ppm in an intensity ratio close to 1:2, which correspond to two distinct P sites. The observed large downfield (31)P NMR shifts can be ascribed to magnetic exchange coupling involving phosphorus atoms. The unpaired electron spin density at the phosphorus nucleus was determined from variable temperature (31)P NMR spectra. The (1)H MAS NMR spectrum was fitted to six components in accordance with the structure as determined from X-ray diffraction. PMID- 12132918 TI - Synthetic models of the reduced active site of superoxide reductase. AB - We report the synthesis, structural and spectroscopic characterization, and magnetic and electrochemical studies of a series of iron(II) complexes of the pyridyl-appended diazacyclooctane ligand L(8)py(2), including several that model the square-pyramidal [Fe(II)(N(his))(4)(S(cys))] structure of the reduced active site of the non-heme iron enzyme superoxide reductase. Combination of L(8)py(2) with FeCl(2) provides [L(8)py(2)FeCl(2)] (1), which contains a trigonal-prismatic hexacoordinate iron(II) center, whereas a parallel reaction using [Fe(H(2)O)(6)](BF(4))(2) provides [L(8)py(2)Fe(FBF(3))]BF(4) (2), a novel BF(4)( )-ligated square-pyramidal iron(II) complex. Substitution of the BF(4)(-) ligand in 2 with formate or acetate ions affords distorted pentacoordinate [L(8)py(2)Fe(O(2)CH)]BF(4) (3) and [L(8)py(2)Fe(O(2)CCH(3))]BF(4) (4), respectively. Models of the superoxide reductase active site are prepared upon reaction of 2 with sodium salts of aromatic and aliphatic thiolates. These model complexes include [L(8)py(2)Fe(SC(6)H(4)-p-CH(3))]BF(4) (5), [L(8)py(2)Fe(SC(6)H(4)-m-CH(3))]BF(4) (6), and [L(8)py(2)Fe(SC(6)H(11))]BF(4) (7). X-ray crystallographic studies confirm that the iron(II)-thiolate complexes model the square-pyramidal geometry and N(4)S donor set of the reduced active site of superoxide reductase. The iron(II)-thiolate complexes are high spin (S = 2), and their solutions are yellow in color because of multiple charge-transfer transitions that occur between 300 and 425 nm. The ambient temperature cyclic voltammograms of the iron(II)-thiolate complexes contain irreversible oxidation waves with anodic peak potentials that correlate with the relative electron donating abilities of the thiolate ligands. This electrochemical irreversibility is attributed to the bimolecular generation of disulfides from the electrochemically generated iron(III)-thiolate species. PMID- 12132919 TI - KNa(3)In(9): a Zintl network phase built of layered indium icosahedra and zigzag chains. Synthesis, structure, bonding, and properties. AB - This phase was discovered following direct fusion of the elements in welded Nb tubes at 550 degrees C and equilibration at 300 degrees C for 1 week. Single crystal X-ray diffraction analysis reveals that KNa(3)In(9) crystallizes in an orthorhombic system (Cmca, Z = 8, a = 9.960(1) A, b =16.564(2) A, c = 17.530(2) A, 23 degrees C). The structure contains a three-dimensional indium network built of layers of empty In(12) icosahedra that are each 12-bonded and interconnected by 4-bonded indium atoms that also form zigzag chains. All cations bridge between cluster faces or edges, and their mixed sizes appear critical to the stability of this particular structure, which does not occur in either binary system. Both empirical electron counting and EHTB band structure calculations on the macroanion indicate that the bonding in this structure is closed-shell, whereas resistivity and magnetic susceptibility measures show that the compound is a moderately poor metal. PMID- 12132920 TI - The hexacoordinate silicate dianions mer-tris[glycolato(2-)-O(1),O(2)]silicate and fac-tris[benzilato(2-)-O(1),O(2)]silicate: syntheses and structural characterization. AB - Treatment of tetramethoxysilane with glycolic acid and morpholine (molar ratio 1:3:2) in methanol, followed by crystallization from methanol/tetrahydrofuran, yielded morpholinium mer-tris[glycolato(2-)-O(1),O(2)]silicate (mer-7). Treatment of benzilic acid with sodium hydride, followed by addition of tetrachlorosilane and triethylamine (molar ratio 3:4:1:2), afforded, after crystallization from 1,4 dioxane/acetonitrile/diethyl ether/n-pentane, triethylammonium fac tris[benzilato(2-)-O(1),O(2)]silicate-hemi-1,4-dioxane (fac 8.(1)/(2)C(4)H(8)O(2)). Single-crystal X-ray diffraction studies showed that the Si-coordination polyhedra of the hexacoordinate silicon(IV) complexes mer-7 and fac-8.(1)/(2)C(4)H(8)O(2) are distorted octahedra. Both compounds were additionally characterized by solid-state VACP/MAS NMR studies ((13)C, (29)Si), and fac-8.(1)/(2)C(4)H(8)O(2) was studied in solution by (1)H, (13)C, and (29)Si NMR experiments. The structural investigations were complemented by computational studies (MP2 studies, TZP level) of the dianions of fac-7 and mer-7. PMID- 12132921 TI - Siting of antimony dopants and gallium in Ba(8)Ga(16)Ge(30) clathrates grown from gallium flux. AB - A series of antimony-doped Ba(8)Ga(16)Ge(30) clathrates was grown as large crystals from gallium flux. These compounds form in the cubic space group Pm( )3n, with the unit cell parameter varying from 10.784(5) to 10.9008(6) A as the amount of GaSb substituting for germanium atoms in the framework is increased. It was found that more antimony than extra gallium was incorporated into the material and that a specific site (the 24k Wyckoff site) was favored by this element. (71)Ga NMR was carried out to determine the siting of gallium; it fills the 6c site preferentially. PMID- 12132922 TI - Hydrothermal synthesis and structural characterization of a new organically templated germanate, Ge(10)O(21)(OH).N(4)C(6)H(21). AB - A new open-framework germanium oxide Ge(10)O(21)(OH).N(4)C(6)H(21) has been hydrothermally synthesized at 180 degrees C for 6 days by using the tris(2 aminoethyl)amine (tren) molecule as a structure-directing agent. This compound was characterized by means of single-crystal X-ray diffraction and FTIR. It crystallizes in the noncentric monoclinic system Cm (a = 14.0495(2) A, b = 12.8058(3) A, c = 9.2637(2) A, beta = 128.406(1) degrees, Z = 4). Its three dimensional framework is built up from GeO(4) and GeO(3)(OH) tetrahedra connected by vertexes to GeO(5) trigonal bipyramids and GeO(6) octahedra. A pseudo-cubic building unit ("4-3" subunit) consists of four GeO(4) tetrahedra, two GeO(5) trigonal bipyramids, and one GeO(6) octahedron (Ge(7)). In the "4-3" block, the GeO(5) trigonal bipyramids share a common edge. This Ge(7) entity is linked to three tetrahedral units GeO(3)X (X = O, OH), and this forms an original decameric building unit Ge(10)O(21)(OH) which is new in the germanates crystal chemistry. It results in a relatively dense open framework composed of pear-shape cavities (7(8)6(2)5(2)4(4)3(2)) encapsulating the triprotonated tren molecule. The inorganic network contains small pores delimited by 7-ring channels running along [001]. PMID- 12132923 TI - Molecular triangle of palladium(II) and its anion binding properties. AB - The ligand 4(3H)-pyrimidone (Hpm) forms the complexes trans-[PdCl(2)(Hpm)(2)] and [Pd(PP)(Hpm)(2)](CF(3)SO(3))(2) (PP = Ph(2)PCH(2)PPh(2) or Ph(2)P(CH(2))(3)PPh(2)), with the neutral ligand (Hpm), and a bowl-like molecular triangle, [(Pd(bu(2)bipy)(mu-pm))(3)](3+) (bu(2)bipy = 4,4'-di-tert-butyl-2,2' bipyridine), with the deprotonated ligand (pm). This triangular complex acts as a host for binding of several anionic guests. PMID- 12132924 TI - Generation and reactivity of rhodium(IV) complexes in aqueous solutions. AB - At pH = 1 and 25 degrees C, the Fenton-like reactions of Fe(aq)(2+) with hydroperoxorhodium complexes LRh(III)OOH(2+) (L = (H(2)O)(NH(3))(4), k = 30 M(-1) s(-1), and L = L(2) = (H(2)O)(meso-Me(6)-[14]aneN(4)), k = 31 M(-1) s(-1)) generate short-lived, reactive intermediates, believed to be the rhodium(IV) species LRh(IV)O(2+). In the rapid follow-up steps, these transients oxidize Fe(aq)(2+), and the overall reaction has the standard 2:1 [Fe(aq)(2+)]/[LRhOOH(2+)] stoichiometry. Added substrates, such as alcohols, aldehydes, and (NH(3))(4)(H(2)O)RhH(2+), compete with Fe(aq)(2+) for LRh(IV)O(2+), causing the stoichiometry to change to <2:1. Such competition data were used to determine relative reactivities of (NH(3))(4)RhO(2+) toward CH(3)OH (1), CD(3)OH (0.2), C(2)H(5)OH (2.7), 2-C(3)H(7)OH (3.4), 2-C(3)D(7)OH (1.0), CH(2)O (12.5), C(2)H(5)CHO (45), and (NH(3))(4)RhH(2+) (125). The kinetics and products suggest hydrogen atom abstraction for (NH(3))(4)RhO(2+)/alcohol reactions. A short chain reaction observed with C(2)H(5)CHO is consistent with both hydrogen atom and hydride transfer. The rate constant for the reaction between Tl(aq)(III) and L(2)Rh(2+) is 2.25 x 10(5) M(-1) s(-1). PMID- 12132926 TI - Alkyl and aryl substituted corroles. 3. Reactions of cofacial cobalt biscorroles and porphyrin-corroles with pyridine and carbon monoxide. AB - The synthesis and characterization of three new cofacial biscorroles and three new linked Co(II) porphyrins and Co(III) corroles with the same face to face orientation are described. The biscorroles are represented as (BCS)Co(2), (BCO)Co(2), (BCX)Co(2) while the porphyrin-corrole dyads are represented as (PCA)Co(2), (PCB)Co(2), (PCO)Co(2) where BC represents the Co(III) cofacial biscorroles and PC represents the porphyrin-corrole complexes which are linked to each other by a dibenzothiophene (S), dibenzofuran (O), or 9,9-dimethylxanthene (X) bridge in the case of the corroles and an anthracene (A), biphenylene (B), or dibenzofuran (O) bridge in the case of the mixed macrocycle derivatives. The electrochemical and spectroscopic data on these new bismacrocycles are compared to those of previously reported biscorroles of the type (BCA)Co(2) and (BCB)Co(2). The CO and/or pyridine binding properties of each biscorrole and porphyrin-corrole in CH(2)Cl(2) are also presented. Only one CO ligand is bound axially to each corrole unit of the bismacrocycle but five- and six-coordinate pyridine complexes can be generated for the same compounds, with the exact stoichiometry depending upon the concentration of pyridine in solution. In all cases, the six-coordinate bispyridine corrole complex can be unambiguously identified by a strong diagnostic marker band located at 598-601 nm. The formation constants for pyridine binding to the biscorroles range from log K(1) = 3.14 to 5.08 while log K(2) ranges from 1.10 to 2.61 depending upon the specific spacer. Carbon monoxide binding constants range from log K = 3.6 to 4.0 in the case of the biscorroles and from log K = 3.4 to 4.1 in the case of the porphyrin corrole dyads. These values also depend on the specific spacer in the complex and, like the pyridine binding constants, decrease in the order BCO > BCA > BCB for the biscorroles and PCO > PCA > PCB for the porphyrin-corrole complexes. PMID- 12132925 TI - Substrate binding in catechol oxidase activity: biomimetic approach. AB - A series of dicopper(II) complexes have been investigated as model systems for the catechol oxidase active site enzyme, regarding the binding of catechol substrate in the first step of the catalytic cycle. The [Cu(2)(L(R))(mu OH)](ClO(4))(2) and [Cu(2)(L(R))(H(2)O)(2)](ClO(4))(3) complexes are based on the L(R) ligands (2,6-bis[(bis(2-pyridylmethyl)amino)methyl]-4-R-substituted phenol) with -R = -OCH(3), -CH(3), or -F. Binding studies of diphenol substrates were investigated using UV-vis and EPR spectroscopy, electrochemistry, and (19)F NMR (fluorinated derivatives). All the complexes are able to bind two ortho-diphenol substrates (tetrachlorocatechol and 3,5-di-tert-butylcatechol). Two successive fixation steps, respectively fast and slower, were evidenced for the mu-OH complexes (the bis(aqua) complexes are inactive in catalysis) by stopped-flow measurement and (19)F NMR. From the mu-OH species, the 1:1 complex/substrate adduct is the catalytically active form. In relation with the substrate specificity observed in the enzyme, different substrate/inhibitor combinations were also examined. These studies enabled us to propose that ortho-diphenol binds monodentately one copper(II) center with the concomitant cleavage of the OH bridge. This hydroxo ligand appears to be a key factor to achieve the complete deprotonation of the catechol, leading to a bridging catecholate. PMID- 12132927 TI - Different complexation properties of some hydroxy keto heterocycles toward beryllium(II) in aqueous solutions: experimental and theoretical studies. AB - Four heterocycles containing hydroxy and keto functionalities have been tested as chelating agents of beryllium(II). These are in the order (i) 3-hydroxy-2-methyl 4H-pyran-4-one (maltol, Hma), (ii) 5-hydroxy-2-(hydroxymethyl)-4H-pyran-4-one (kojic acid, Hka), (iii) 3-hydroxy-1,2-dimethyl-4-pyridinone (Hdpp), (iv) 1-(3 hydroxy-2-furanyl)ethanone (isomaltol, Hima). Although the skeletons of the first three species, with one nitrogen or oxygen heteroatom at the six-membered ring, are almost superimposable, straightforward synthesis and crystallization is achieved only for the 1:2 adduct Be(dpp)(2), 1. Also the complex Be(ima)(2), 2, precipitates in high yield but the ima(-) ligand has a different skeletal structure. X-ray determinations of 1 and 2 showed that the Be(2+) ion is pseudotetrahedrally coordinated by two chelating ligands with slightly asymmetric Be-O(alkoxo) and Be-O(keto) bonds. The complex Be(ma)(2) precipitates in low yields together with large amounts of unreacted Hma while, under the same conditions, no trace of the analogous species Be(ka)(2) has been observed. This paper presents the results of potentiometric and NMR studies in the aqueous solutions as well as of DFT structural optimizations for all of the free acids, their associated bases, and the adducts of the type [BeL(H(2)O)(2)](+) and BeL(2) in the gas phase. It is consistently found that the basicity of the ligands and the stability of their complexes decrease in the order dpp(-) > ma(-) > ka(-) > ima(-). In solution, all of the anionic ligands form adducts of the type [BeL(H(2)O)(2)](+) at low pH values, whereas higher concentrations of the free anion are required to form 1:2 adducts. The pH, the basicity, and the stability constants of the complexes as well as the formation of competing beryllium hydroxide species are strictly correlated factors for the obtainment of the latter type of adduct. The DFT calculations account nicely for the different donor powers of the various chelates in terms of electronic redistribution and associated energetics. PMID- 12132928 TI - Mechanistic studies of a calcium-dependent MRI contrast agent. AB - Intracellular Ca(2+) plays an important role in signal transduction, and we are developing new MRI techniques to study its regulation in living animals. We have reported on an MRI contrast agent (DOPTA-Gd) where the relaxivity of the complex is controlled by the presence or absence of the divalent ion Ca(2+). By structurally modulating inner-sphere access of water to a chelated Gd(3+) ion, we observe a substantial and reversible change in T(1) upon the addition of Ca(2+) and not other divalent ions. Luminescence lifetime and NMRD measurements of the complex have been acquired, and several parameters contribute to the Ca(2+) dependent relaxivity change of DOPTA-Gd. The number of inner-sphere water molecules is more than doubled after the Ca(2+) concentration is increased. This finding strongly supports the proposed conformational change of DOPTA-Gd when Ca(2+) is bound. Relaxometric measurements confirm these results and provide an indication that second-sphere water molecules are probably responsible for paramagnetic relaxation enhancement in the absence of Ca(2+). After Ca(2+) is bound to DOPTA-Gd, the molecule undergoes a substantial conformational change that opens up the hydrophilic face of the tetraazacyclododecane macrocycle. This change dramatically increases the accessibility of chelated Gd(3+) ion to bulk solvent. The design of this class of calcium-activated MR contrast agent was based primarily on the assumption that the number of coordinated inner-sphere water molecules would be the dominating factor in observed relaxivity measurements. This result has been confirmed; however, careful mechanistic studies reveal that additional factors are involved in this process. PMID- 12132930 TI - Synthesis and characterization of [Mo(7)O(16)(O(3)PCH(2)PO(3))(3)]:(8-) a mixed valent polyoxomolybdenum diphosphonate anion with octahedrally and tetrahedrally coordinated molybdenum. AB - Two new compounds containing the title diphosphono-polyoxometalate anion and diprotonated ethylenediamine (enH(2)) or piperazine (ppzH(2)) countercations have been hydrothermally synthesized and structurally characterized ((enH(2))(4)[Mo(7)O(16)(O(3)PCH(2)PO(3))(3)].7H(2)O, triclinic, P(-)1, Z = 2, a = 10.3455(7) A, b = 13.136(1) A, and c = 20.216(3) A, alpha = 93.247(6) degrees, beta = 96.434(6) degrees, and gamma = 111.900(6) degrees; (ppzH(2))(4)[Mo(7)O(16)(O(3)PCH(2)PO(3))(3)].8H(2)O, triclinic, P(-)1, Z = 2, a = 13.255(2) A, b = 13.638(2) A, and c = 16.874(4) A, alpha = 93.20(2) degrees, beta = 101.27(2) degrees, and gamma = 105.87(1) degrees). The anion is a ring of three pairs of edge-sharing octahedra of Mo(V)O(6) (with Mo(V)-Mo(V) bonds) that share corners with each other. The diphosphonate groups connect the pairs at the periphery. The ring is "capped" by a tetrahedron of Mo(VI)O(4). According to magnetic measurements, the compounds are diamagnetic. PMID- 12132929 TI - Metalla-supramolecular rectangles as electron reservoirs for multielectron reduction and oxidation. AB - The electron-transfer capacity of molecular rectangle ions [Pt(II)(4)(PEt(3))(8)(mu-anth(2-))(2)(mu-L)(2)](4+) with anth = anthracene-1,8 diyl and L = 4,4'-bipyridine (bp) or 1,2-bis(4-pyridyl)ethene (bpe) was investigated in acetonitrile and dichloromethane using cyclic voltammetry, EPR, and UV-vis-near-IR spectroelectrochemistry. The compounds can be reversibly reduced, first in a two-electron process and then via two closely separated one electron steps. Oxidation was also possible at rather low potentials in a reversible two-electron step, followed by an electrochemically irreversible process. The spectroscopic results indicate reduction at the neutral acceptor ligands L and oxidation at the formally dianionic anthracene "clips". In contrast, the prototypical molecular square ([Pt(triphos)(mu-bp)](4))(8+) undergoes only irreversible reduction. PMID- 12132931 TI - Synthesis and structural, spectroscopic, and electrochemical characterization of new ruthenium dimethyl sulfoxide nitrosyls. AB - The reactivity of ruthenium(II)- and ruthenium(III)-chloride-dimethyl sulfoxide precursors and of the antimetastatic drug [ImH][trans-RuCl(4)(dmso-S)(Im)] (NAMI A, Im = imidazole, dmso = dimethyl sulfoxide) toward NO was investigated. Treatment of [(dmso)(2)H][trans-RuCl(4)(dmso-S)(2)] and mer-RuCl(3)(dmso)(3) with gaseous NO yielded [(dmso)(2)H][trans-RuCl(4)(dmso-O)(NO)] (1) and mer,cis RuCl(3)(dmso-O)(2)(NO) (2), respectively. Thus, coordination of the strong pi acceptor NO induces a S to O linkage isomerization of the dmso trans to it to avoid competition for pi-electrons. In light-protected nitromethane solutions, complex 2 equilibrates slowly with the two isomers mer-RuCl(3)(dmso-S)(dmso O)(NO) (3), with NO trans to Cl, and mer-RuCl(3)(dmso-S)(dmso-O)(NO) (4), with NO trans to dmso-O; the equilibrium mixture consists of ca. 64% 2, 3% 3, and 33% 4. Treatment of the Ru(II) precursor trans-RuCl(2)(dmso-S)(4) with gaseous NO in CH(2)Cl(2) solution yielded the nitrosyl-nitro derivative trans,cis,cis RuCl(2)(dmso-O)(2)(NO)(NO(2)) (5). Finally, [(Im)(2)H][trans-RuCl(4)(Im)(NO)] (6) was prepared by treatment of [ImH][trans-RuCl(4)(dmso-O)(NO)] (1Im) with an excess of imidazole in refluxing acetone. The spectroscopic features are consistent with the [Ru(NO)](6) formulation for all complexes, that is, a diamagnetic Ru(II) nucleus bound to NO(+). Compounds 1, 2, 5, and 6 were characterized also by X-ray crystallography; they all show a linear nitrosyl group, with short Ru-NO bond distances consistent with a strong d(pi) --> pi NO back-bonding. An unusual inertness of O-bonded dmso was observed in compound 1. Complexes 1, 2, 3, 5, and 6 are all redox active in DMF solutions showing irreversible reductions whose peak potentials depend on the other ligands attached to the Ru metal center. The site of reduction is the NO(+) moiety. The reduced complexes are not stable and release a Cl(-) or NO(2)(-) ligand followed by the NO(*) radical. The chemical reactions following electron transfer are all fast (rate constant >100 s(-1) at 293 K). The Ru product species are not redox active within the DMF window. PMID- 12132932 TI - Synthesis and study of Ru,Rh,Ru triads: modulation of orbital energies in a supramolecular architecture. AB - Supramolecular trimetallic complexes [((tpy)RuCl(BL))(2)RhCl(2)](3+) where tpy = 2,2':6',2' '-terpyridine and BL = dpp or bpm [dpp = 2,3-bis(2-pyridyl)pyrazine and bpm = 2,2'-bipyrimidine] have been synthesized and characterized. The mixed metal complexes couple a reactive rhodium(III) center to two ruthenium(II) light absorbers to form a light absorber-electron collector-light absorber triad. The variation of the bridging (dpp and bpm) and terminal (tpy in lieu of bpy) ligands has some profound effects on the properties of these complexes, and they are remarkably different from the previously reported [((bpy)(2)Ru(bpm))(2)RhCl(2)](5+) system. The electrochemical data for both title trimetallics consist of overlapping Ru(III/II) couples for both terminal metals at 1.12 V versus the Ag/AgCl reference electrode. Cathodically an irreversible Rh(III/I) reduction followed by bridging ligand reductions is seen. This is indicative of highest occupied molecular orbitals (HOMO) localized on the terminal ruthenium metal centers and a lowest unoccupied molecular orbital (LUMO) residing on the rhodium. This rhodium-based LUMO is in contrast to the bpy analogue [((bpy)(2)Ru(bpm))(2)RhCl(2)](5+), which has a bpm(pi) localized LUMO. This orbital inversion by terminal ligand variation illustrates the similar energy of these Rh(dsigma) and bpm(pi) orbitals within this structural motif. Both title trimetallics possess broad, low-energy Ru --> BL charge transfer absorbances at 540 nm (dpp) and 656 nm (bpm). A comparison of the spectroscopic, electrochemical, and spectroelectrochemical properties of these trimetallic complexes is presented. PMID- 12132933 TI - Antimony ethylene glycolate and catecholate compounds: structural characterization of polyesterification catalysts. AB - Antimony compounds are widely used as catalysts for the synthesis of the commercially important polymer poly(ethyleneterephthalate) by polycondensation of bis(hydroxyethyl)terephthalate. The precise nature of the antimony catalysts is, however, unknown. The present study has been conducted with a view to determining the nature of the catalytic species by structurally characterizing antimony ethylene glycolate compounds and related catecholate derivatives, namely [Sb(2)(OCH(2)CH(2)O)(3)](n), [Sb(OCH(2)CH(2)O)(OAc)](n), [pySb(1,2 O(2)C(6)H(4))](2)O, and [pyH][Sb(1,2-O(2)C(6)H(4))(2)]. PMID- 12132934 TI - Synthesis, structure, and magnetic properties of dinuclear cobalt(II) complexes with a new phenol-based dinucleating ligand with four hydroxyethyl chelating arms. AB - A new end-off type acyclic ligand with four hydroxyethyl arms, 2,6-bis[bis(2 hydroxyethyl)aminomethyl]-4-methylphenol [H(bhmp)], formed dinuclear cobalt(II) complexes [Co(2)(bhmp)(OAc)(2)]BPh(4) (1) and [Co(2)(bhmp)(OBz)(2)]BPh(4) (2). The complex 1.2.5CH(3)CN (C(50)H(62.5)BCo(2)N(4.5)O(9)) crystallizes in the monoclinic space group C2/c with dimensions a = 25.424(5) A, b = 13.376(2) A, c = 29.913(6) A, beta = 105.930(3) degrees, and V = 9781(3) A(3) and with Z = 8. X ray diffraction analysis revealed a mu-phenoxo-bis(mu-acetato)dicobalt(II) core structure containing two octahedral cobalt(II) ions. Electronic spectra were investigated for 1 and 2 in the range 400-1800 nm, and the data were typical for the octahedral high-spin cobalt(II) complexes. Magnetic susceptibility was measured for 1 and 2 over the temperature range 4.5-300 K, and the data were analyzed well using our theoretical method. The best fitting parameters were kappa = 0.77, lambda = -116 cm(-1), Delta = 572 cm(-1), and J = -0.44 cm(-1) for complex 1 and kappa = 0.96, lambda = -93 cm(-1), Delta = 616 cm(-1), and J = 0.33 cm(-1) for complex 2. PMID- 12132935 TI - Synthesis and structures of zirconium amide-iodide complexes. AB - Zirconium amide-iodide complexes were synthesized for possible use as chemical vapor deposition precursors to zirconium nitride films. The series of six complexes Zr(NR(2))(4-n)I(n)(R = Me or Et; n = 1-3) was prepared by reacting ZrI(4) and Zr(NR(2))(4) in hot toluene. X-ray crystallographic analyses were performed for Zr(NMe(2))(3)I, Zr(NEt(2))(2)I(2), and Zr(NEt(2))I(3). In the solid state, Zr(NMe(2))(3)I and Zr(NEt(2))(2)I(2) are the discrete dimers [Zr(NMe(2))(2)I(mu-NMe(2))](2) and [Zr(NEt(2))(2)I(mu-I)](2), and Zr(NEt(2))I(3) is the polymer of dimers ([Zr(NEt(2))I(2)(mu-I)](2))(n). In solution, Zr(NEt(2))(3)I is proposed to be monomeric on the basis of NMR data and a molecular weight determination. The complex Zr(NEt(2))(3)I is the most promising precursor candidate because of its physical properties. PMID- 12132936 TI - Two-dimensional rhombohedral grid coordination polymers [M(bbbt)(2)(NCS)(2)](n)(M = Co, Mn, or Cd; bbbt = 1,1'-(1,4-butanediyl) bis-1H-benzotriazole): synthesis, crystal structures, and third-order nonlinear optical properties. AB - In this paper, treatment of 1,1'-(1,4-butanediyl) bis-1H-benzotriazole (bbbt) and KSCN with Co(II), Mn(II), or Cd(II) afforded three two-dimensional rhombohedral grid coordination polymers [M(bbbt)(2)(NCS)(2)](n)(M = Co, 1; Mn, 2; Cd, 3). The two-dimensional rhombohedral grids are parallel to the crystallographic ac plane. The rhombohedral grid consists of 44-membered rings of M(4)(bbbt)(4), and gives the dimensions of 12.913 x 10.764 A for polymer 1, 13.106 x 10.797 A for polymer 2, and 13.256 x 10.870 A for polymer 3. The three polymers' third-order nonlinear optical (NLO) properties were determined by Z-scan technique in DMF solution. The results show that all three polymers show very large NLO absorption and strong NLO refraction properties. The third-order NLO absorptive coefficients alpha(2) are 5.4 x 10(-9) m W(-1) for polymer 1, 5.2 x 10(-9) m W(-1) for polymer 2, and 5.0 x 10(-9) m W(-1) for polymer 3. The alpha(2) values are larger than those of all the reported cluster compounds. The NLO refractive index values n(2) of the three polymers are 5.73 x 10(-19), 3.55 x 10(-19), and 3.07 x 10(-19) m(2) W(-1), respectively. Their hyperpolarizability gamma values are calculated to be 2.40 x 10(-30) esu for polymer 1, 1.52 x 10(-30) esu for polymer 2, and 1.50 x 10(-30) esu for polymer 3. The gamma values are comparable to those of clusters and better than those of organometallic compounds, semiconductors, and fullerene. PMID- 12132938 TI - Examination of Jeltrate Plus as a tissue equivalent bolus material. AB - A product available commercially for making dental impressions, Jeltrate Plus, was evaluated as a tissue equivalent bolus material. Jeltrate Plus was found to be tissue equivalent in 6 and 15 MV photon energy beams and 6, 12, and 20 MeV electron energy beams. As a first step, different preparations for making the bolus material were investigated and an optimal mixture was determined to be two parts Jeltrate Plus powder to three parts water by weight. A suitable method for storing the material was found to be in a water filled plastic container. Since the product is fairly inexpensive and is easily and quickly made and moulded into different shapes, it is an excellent bolus material to use when treating irregular patient contours. PMID- 12132939 TI - PTV margin determination in conformal SRT of intracranial lesions. AB - The planning target volume (PTV) includes the clinical target volume (CTV) to be irradiated and a margin to account for uncertainties in the treatment process. Uncertainties in miniature multileaf collimator (mMLC) leaf positioning, CT scanner spatial localization, CT-MRI image fusion spatial localization, and Gill Thomas-Cosman (GTC) relocatable head frame repositioning were quantified for the purpose of determining a minimum PTV margin that still delivers a satisfactory CTV dose. The measured uncertainties were then incorporated into a simple Monte Carlo calculation for evaluation of various margin and fraction combinations. Satisfactory CTV dosimetric criteria were selected to be a minimum CTV dose of 95% of the PTV dose and at least 95% of the CTV receiving 100% of the PTV dose. The measured uncertainties were assumed to be Gaussian distributions. Systematic errors were added linearly and random errors were added in quadrature assuming no correlation to arrive at the total combined error. The Monte Carlo simulation written for this work examined the distribution of cumulative dose volume histograms for a large patient population using various margin and fraction combinations to determine the smallest margin required to meet the established criteria. The program examined 5 and 30 fraction treatments, since those are the only fractionation schemes currently used at our institution. The fractionation schemes were evaluated using no margin, a margin of just the systematic component of the total uncertainty, and a margin of the systematic component plus one standard deviation of the total uncertainty. It was concluded that (i) a margin of the systematic error plus one standard deviation of the total uncertainty is the smallest PTV margin necessary to achieve the established CTV dose criteria, and (ii) it is necessary to determine the uncertainties introduced by the specific equipment and procedures used at each institution since the uncertainties may vary among locations. PMID- 12132940 TI - Comparison of two algorithms for determining beam weights and wedge filters. AB - This article compares two algorithms for determining beam weights and wedge filters for conformal treatment planning. One algorithm, which is based on dose gradient analysis, provides analytic formulas for determining beam weights, wedge angles, and collimator angles (i.e., wedge orientations) so that the dose distribution is homogeneous in the target volume. The second algorithm is based on the concept of the super-omni wedge (i.e., the arrangement of two pairs of orthogonal nominal wedged beams), numerically optimize beam weights, wedge angles, and collimator angles so that the dose requirements to targets and organs at risk are satisfied to the best. Three clinical cases were tested. For the first case, both algorithms resulted in comparable homogeneous dose distributions in the target volume. For the second case, the second algorithm resulted in much lower doses to the eyes plus a better homogeneous dose distribution in the target volume. For the third case, only the second algorithm was applicable, and the treatment plan it developed met the prescribed requirements. The results show that the first algorithm is better in terms of feasibility, whereas the second is better in terms of applicability and the quality of treatment plans. PMID- 12132941 TI - On the selection of optimization parameters for an inverse treatment planning replacement of a forward planning technique for prostate cancer. AB - The influence of organ volume sampling, lateral scatter inclusion, and the selection of objectives and constraints on the inverse treatment planning process with a commercial treatment planning system is investigated and suitable parameters are identified for an inverse treatment planning replacement of a clinical forward planning technique for prostate cancer. For the beam geometries of the forward technique, a variable set of parameters is used for the calculation of dose from pencil beams. An optimal set is identified after the evaluation of optimized plans that correspond to different sets of pencil-beam parameters. This set along with a single, optimized set of objectives and constraints is used to perform inverse planning on ten randomly selected patients. The acceptability of the resulting plans is verified by comparisons to the clinical ones calculated with the forward techniques. For the particular commercial treatment planning system, the default values of the pencil beam parameters are found adequate for inverse treatment planning. For all ten patients, the optimized, single set of objectives and constraints results in plans with target coverage comparable to that of the forward plans. Furthermore inverse treatment planning reduces the overall mean rectal and bladder doses by 4.8% and 5.8% of the prescription dose respectively. The study indicates that (i) inverse treatment planning results depend implicitly on the sampling of the dose distribution, (ii) inverse treatment planning results depend on the method used by the dose calculation model to account for scatter, and (iii) for certain sites, a single set of optimization parameters can be used for all patient plans. PMID- 12132943 TI - Calibration of a liquid I-125 source in a syringe. AB - The calibration of a liquid I-125 source in an older standard dose calibration system is presented. The calibration factor agrees well with factors established by the NIST for newer dose calibration systems. The determination of the source activity is necessary to accurately calculate the time required to deliver the prescribed dose. PMID- 12132942 TI - Measured transverse-axis dosimetric parameters of the model STM1251 125I interstitial source. AB - The recent popularity of permanent implants for treatment of prostate cancer has created a large demand for low energy seed sources. Vendors have introduced new source designs to meet this demand. The AAPM has recommended that all low energy interstitial brachytherapy seed sources be subjected to independent dosimetric evaluations, preferably using experimental measurements as well as Monte Carlo calculations. This work presents the results of Thermo-Luminescence Dosimeter (TLD) measurements of dosimetric parameters on the transverse axis of a new 125I seed source, the Source Tech Medical STM1251 125I seed. Experimental measurements were performed in a Solid Water phantom, with the results corrected to values for liquid water using Monte Carlo calculated correction factors. The parameters measured include the dose rate constant and values of the radial dose function at distances of 0.5 cm through 5 cm. The measured dose rate constant in liquid water for the STM1251 125I seed was 1.039 cGy/U-hr. Measured radial dose function values agreed with Monte Carlo calculated ones to within 10%. These measurements therefore confirm the modeling and simulation of Monte Carlo calculations for this 125I source design, within the statistical uncertainties of the calculation and measurement techniques. PMID- 12132944 TI - Optimized planning for intraoperative planar permanent-seed implant. AB - We describe a fast, PC-based optimization planning system for a planar permanent seed implant. Sites where this system is applicable include brain, lung, and head and neck. The system described here allows placing ribbons of different strengths and of different lengths along and across the implant plane. The program takes full advantage of the availability of different source strengths in inventory, and attempts to find configurations of ribbons that result in optimal dose uniformity over the prescription plane. Dosimetry is based on the AAPM TG 43 Report [R. Nath et al., Med. Phys. 22, 209-234 (1995)]. Compared with TG 43 parameters, the classical tables underestimate the I-125 source strengths needed by 40%. The use of several source strengths improves the plan. Typical optimization yields dose uniformity of 10%, and computing times are within 2-3 min. No further enhancement is obtained if ribbons are placed in a grid pattern as opposed to the (simpler) arrangement along parallel lines. Nor is it valuable to have variable ribbon lengths. For an I-125 implant the optimization system described here is a practical alternative to the (strictly speaking inapplicable) classical systems. It calculates correctly the total source strengths, and--most notably--generates plans with optimal dose uniformity. The fast computing time is well suited for planning during surgery in the operating room. PMID- 12132945 TI - A parameter optimization algorithm for intensity-modulated radiotherapy prostate treatment planning. AB - An iterative algorithm has been developed to analytically determine patient specific input parameters for intensity-modulated radiotherapy prostate treatment planning. The algorithm starts with a generic set of inverse planning parameters that include dose and volume constraints for the target and surrounding critical structures. The overlap region between the target volume and the rectum is used to determine the optimized target volume coverage goal. Sequential iterations are performed to vary the numerous parameters individually or in sets while other parameters remain fixed. A coarse grid search is first used to avoid convergence on a local maximum. Linear interpolation is then used to define a region for a fine grid search. Selected parameters are also tested for possible improvements in target coverage. In several representative test cases investigated the coverage of the planning target volume improved with the use of the algorithm while still meeting the clinical acceptability criteria for critical structures. The algorithm avoids time-consuming random trial and error variations that are often associated with difficult cases and also eliminates lengthy user learning curves. The methodology described in this paper can be applied to any treatment planning system that requires the user to select the input optimization parameters. PMID- 12132946 TI - Monitor unit calculation for tangential breast treatments: verification in an anthropomorphic phantom. AB - This paper presents an anthropomorphic phantom study of dose delivered to a specific point during tangential breast irradiation to verify monitor unit calculations. Measurements were made using a 0.6 cc Farmer type cylindrical ionization chamber in the phantom and compared to calculations made on a three dimensional radiotherapy treatment planning system using single digitized contour through to multi slice CT data. A large breast phantom was used for a single field size with a combination of open and wedged fields for three different energies (4, 6, and 18 MV). Solid flat phantom measurements were also performed for comparison. Results showed a lower calculated dose than the dose measured for a fixed number of monitor units where the variations were within a range of 0.8% to 4.5%. Differences were larger for the anthropomorphic phantom than the flat phantom. We conclude that little accuracy is gained from CT based monitor unit calculations compared to those based on digitised contours for this breast treatment but that the dose distributions will be affected. This type of test is recommended as one of a large set, in the commissioning and testing procedures for treatment planning systems. PMID- 12132947 TI - A comparison of computer-controlled versus manual on-line patient setup adjustment. AB - A study was performed to determine the relative advantage of computer-controlled couch movement versus manual repositioning to correct patient setup error measured using an electronic portal imaging device (EPID). Twenty-eight on-line setup adjustment trials of anterior-posterior (AP) pelvic projections were evaluated, with 13 setups corrected by automated couch movement determined by direct feedback from the EPID image alignment tool and 15 setups manually corrected based on the transformation displayed from the same tool. The speed of setup adjustment and accuracy of corrected setup were determined. Computer controlled setup adjustment was determined to be faster (25.4 s versus 101.9 s) and slightly more accurate (1.8 mm versus 2.5 mm error in adjusted setup) than manual correction. PMID- 12132948 TI - Radiographic techniques in screen-film mammography. AB - The objectives of this study were to document imaging physics parameters associated with mammography physics surveys, and investigate how the choice of tube potential affects average glandular dose (AGD) and x-ray exposure time. Data from 60 mammography units were obtained pertaining to representative values of mAs, exposure time, half value layer, AGD and film density when acquiring phantom images. The survey of clinical systems showed that for a normal sized breast as represented by the mammography accreditation phantom, 60% of these units were operated at 25 kVp, and 33% at 26 kVp. Median exposure times were 1.14 s at 25 kVp and 0.73 s at 26 kVp. The median AGD was 1.62 mGy at 25 kVp and 1.51 mGy at 26 kVp. As expected, the choice of x-ray tube potential did not significantly affect the median film density value of 1.5. Five clinical systems, all from different vendors, had measurements performed of the AGD and x-ray exposure time as a function of x-ray tube potential at a constant film density. For a typical clinical x-ray unit, increasing the x-ray tube potential from 25 to 28 kVp reduced the exposure time by 50%, and reduced the AGD by 26%. PMID- 12132949 TI - Improved method of magnification factor calculation for the angiographic measurement of neurovascular lesion dimensions. AB - Accurately evaluating the size of a neurovascular lesion is essential for properly devising treatment strategies. The magnification factor must be considered in order to measure the dimension of a lesion from an angiogram. Although a method to calculate the magnification of the lesion by linear interpolation of the measurable magnification factors of two markers has been in use, this paper shows that it can be inaccurate. By deriving the exact formula for calculating the magnification factor at the level of the lesion, the error generated by the linear interpolation of magnification factor has been evaluated. This error was found to depend on source-to-skin distance (SSD), the location of the lesion in the head, and the head size. The closer the head is to the focal spot and the nearer the lesion is to the center of the head, the larger is the error. Since clinicians tend to use high geometric magnification (i.e., small SSD) in interventional procedures, there exists a possible consequential error of more than 3% in lesion sizing if the linear-interpolation calculation method is used. It is thus recommended that the exact formula derived here be used to calculate the magnification factor to improve accuracy. PMID- 12132950 TI - Dosimetric advantage of using 6 MV over 15 MV photons in conformal therapy of lung cancer: Monte Carlo studies in patient geometries. PMID- 12132951 TI - A piece of my mind: a perfect match. PMID- 12132953 TI - Telemedicine and remote patient monitoring. PMID- 12132954 TI - Putting mental retardation and mental illness on health care professionals' radar screen. PMID- 12132955 TI - Mind-body medicine explored at APA meeting. PMID- 12132956 TI - Colon and rectal surgeons are trying Botox treatment, too. PMID- 12132961 TI - St John's wort and depression. PMID- 12132962 TI - St John's wort and depression. PMID- 12132963 TI - St John's wort and depression. PMID- 12132964 TI - St John's wort and depression. PMID- 12132965 TI - St John's wort and depression. PMID- 12132966 TI - St John's wort and depression. PMID- 12132969 TI - The increasing power of placebos in trials of antidepressants. PMID- 12132970 TI - The increasing power of placebos in trials of antidepressants. PMID- 12132971 TI - The increasing power of placebos in trials of antidepressants. PMID- 12132972 TI - Measurement of serum estradiol levels in postmenopausal women. PMID- 12132974 TI - Accuracy of tablet splitting by elderly patients. PMID- 12132975 TI - Long-term persistence in use of statin therapy in elderly patients. AB - CONTEXT: Knowledge of long-term persistence with 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitor (statin) therapy is limited because previous studies have observed patients for short periods of time, in closely monitored clinical trials, or in other unrepresentative settings. OBJECTIVE: To describe the patterns and predictors of long-term persistence with statin therapy in an elderly US population. DESIGN, SETTING, AND PATIENTS: Retrospective cohort study including 34 501 enrollees in the New Jersey Medicaid and Pharmaceutical Assistance to the Aged and Disabled programs who were 65 years of age and older, initiated statin treatment between 1990 and 1998, and who were followed up until death, disenrollment, or December 31, 1999. MAIN OUTCOME MEASURES: Proportion of days covered (PDC) by a statin in each quarter during the first year of therapy and every 6 months thereafter; predictors of suboptimal persistence during each interval (PDC <80%) were identified using generalized linear models for repeated measures. RESULTS: The mean PDC was 79% in the first 3 months of treatment, 56% in the second quarter, and 42% after 120 months. Only 1 patient in 4 maintained a PDC of at least 80% after 5 years. The proportion of patients with a PDC less than 80% increased in a log-linear manner, comprising 40%, 61%, and 68% of the cohort after 3, 12, and 120 months, respectively. Independent predictors of poor long-term persistence included nonwhite race, lower income, older age, less cardiovascular morbidity at initiation of therapy, depression, dementia, and occurrence of coronary heart disease events after starting treatment. Patients who initiated therapy between 1996-1998 were 21% to 25% more likely to have a PDC of at least 80% than those who started in 1990. CONCLUSIONS: Persistence with statin therapy in older patients declines substantially over time, with the greatest drop occurring in the first 6 months of treatment. Despite slightly better persistence among patients who began treatment in recent years, long-term use remains low. Interventions are needed early in treatment and among high-risk groups, including those who experience coronary heart disease events after initiating treatment. PMID- 12132976 TI - Adherence with statin therapy in elderly patients with and without acute coronary syndromes. AB - CONTEXT: Landmark clinical trials have demonstrated the survival benefits of statins, with benefits usually starting after 1 to 2 years of treatment. Research prior to these trials of older lipid-lowering agents demonstrated low levels of 1 year adherence. OBJECTIVE: To compare 2-year adherence following statin initiation in 3 cohorts of patients: those with recent acute coronary syndrome (ACS), those with chronic coronary artery disease (CAD), and those without coronary disease (primary prevention). DESIGN AND SETTING: Cohort study using linked population-based administrative data from Ontario. PATIENTS: All patients aged 66 years or older who received at least 1 statin prescription between January 1994 and December 1998 and who did not have a statin prescription in the prior year were followed up for 2 years from their first statin prescription. There were 22,379 patients in the ACS, 36,106 in the chronic CAD, and 85,020 in the primary prevention cohorts. MAIN OUTCOME MEASURES: Adherence to statins, defined as a statin being dispensed at least every 120 days after the index prescription for 2 years. RESULTS: Two-year adherence rates in the cohorts were only 40.1% for ACS, 36.1% for chronic CAD, and 25.4% for primary prevention. Relative to the ACS cohort, nonadherence was more likely among patients receiving statins in the chronic CAD (relative risk [RR], 1.14; 95% CI, 1.11-1.16) and primary prevention cohorts (RR, 1.92; 95% CI, 1.87-1.96). CONCLUSIONS: Elderly patients with and without recent ACS have low rates of adherence to statins. This suggests that many patients initiating statin therapy may receive no or limited benefit from statins because of premature discontinuation. PMID- 12132977 TI - Sustained-release bupropion for smoking cessation in African Americans: a randomized controlled trial. AB - CONTEXT: African Americans disproportionately experience greater smoking attributable morbidity and mortality. Few clinical trials for smoking cessation in African Americans have been conducted, despite a different profile of both smoking and quitting patterns. OBJECTIVE: To compare a sustained-release form of bupropion hydrochloride (bupropion SR) with placebo for smoking cessation among African Americans. DESIGN, SETTING, AND PARTICIPANTS: Randomized, double-blind, placebo-controlled trial conducted from February 11, 1999, to December 8, 2000, of 600 African American adults treated at a community-based health care center. Volunteers, who smoked 10 or more cigarettes per day were recruited by targeted media and health care professionals. INTERVENTION: Participants were randomly assigned to receive 150 mg of bupropion SR (n = 300) or placebo (n = 300) twice daily for 7 weeks. Brief motivational counseling was provided in-person at baseline, quit day, weeks 1 and 3, end of treatment (week 6), and by telephone at day 3 and weeks 5 and 7. MAIN OUTCOME MEASURES: Biochemically confirmed 7-day point prevalence abstinence at weeks 6 and 26 following quit day. RESULTS: Using intention-to-treat procedures, confirmed abstinence rates at the end of 7 weeks of treatment were 36.0% in the bupropion SR group and 19.0% in the placebo group (17.0 percentage point difference; 95% confidence interval, 9.7-24.4; P<.001). At 26 weeks the quit rates were 21.0% in the treatment and 13.7% in the placebo groups (7.3 percentage point difference; 95% confidence interval, 1.0-13.7; P =.02). Those taking bupropion SR experienced a greater mean reduction in depression symptoms at week 6 (2.96 [9.45] vs 1.13 [8.84]) than those taking placebo, and after controlling for continuous abstinence, those taking bupropion SR also gained less weight than those taking placebo. CONCLUSIONS: Bupropion SR was effective for smoking cessation among African Americans and may be useful in reducing the health disparities associated with smoking. PMID- 12132978 TI - Association of health literacy with diabetes outcomes. AB - CONTEXT: Health literacy is a measure of patients' ability to read, comprehend, and act on medical instructions. Poor health literacy is common among racial and ethnic minorities, elderly persons, and patients with chronic conditions, particularly in public-sector settings. Little is known about the extent to which health literacy affects clinical health outcomes. OBJECTIVES: To examine the association between health literacy and diabetes outcomes among patients with type 2 diabetes. DESIGN, SETTING, AND PARTICIPANTS: Cross-sectional observational study of 408 English- and Spanish-speaking patients who were older than 30 years and had type 2 diabetes identified from the clinical database of 2 primary care clinics of a university-affiliated public hospital in San Francisco, Calif. Participants were enrolled and completed questionnaires between June and December 2000. We assessed patients' health literacy by using the short-form Test of Functional Health Literacy in Adults (s-TOFHLA) in English or Spanish. MAIN OUTCOME MEASURES: Most recent hemoglobin A(1c) (HbA(1c)) level. Patients were classified as having tight glycemic control if their HbA(1c) was in the lowest quartile and poor control if it was in the highest quartile. We also measured the presence of self-reported diabetes complications. RESULTS: After adjusting for patients' sociodemographic characteristics, depressive symptoms, social support, treatment regimen, and years with diabetes, for each 1-point decrement in s TOFHLA score, the HbA(1c) value increased by 0.02 (P =.02). Patients with inadequate health literacy were less likely than patients with adequate health literacy to achieve tight glycemic control (HbA(1c) < or = 7.2%; adjusted odds ratio [OR], 0.57; 95% confidence interval [CI], 0.32-1.00; P =.05) and were more likely to have poor glycemic control (HbA(1c) > or = 9.5%; adjusted OR, 2.03; 95% CI, 1.11-3.73; P =.02) and to report having retinopathy (adjusted OR, 2.33; 95% CI, 1.19-4.57; P =.01). CONCLUSIONS: Among primary care patients with type 2 diabetes, inadequate health literacy is independently associated with worse glycemic control and higher rates of retinopathy. Inadequate health literacy may contribute to the disproportionate burden of diabetes-related problems among disadvantaged populations. Efforts should focus on developing and evaluating interventions to improve diabetes outcomes among patients with inadequate health literacy. PMID- 12132979 TI - Aircraft cabin air recirculation and symptoms of the common cold. AB - CONTEXT: In recent years, new commercial aircraft have been designed to recirculate approximately 50% of the cabin air to increase fuel efficiency. Some older aircraft use only fresh air. Whether air recirculation increases the transmission of infectious disease is unknown; some studies have demonstrated higher rates of the common cold among persons working in buildings that recirculate air. OBJECTIVE: To evaluate the role of air recirculation as a predictor of postflight upper respiratory tract infections (URIs). DESIGN, SETTING, AND PARTICIPANTS: A natural experiment conducted among 1100 passengers departing the San Francisco Bay area in California and traveling to Denver, Colo, during January through early April 1999, and who completed a questionnaire in the boarding area and a follow-up telephone interview 5 to 7 days later. Forty-seven percent traveled aboard airplanes using 100% fresh air for ventilation, and 53% traveled aboard aircraft that recirculated cabin air. MAIN OUTCOME MEASURE: Incidence of reporting new URI symptoms within 1 week of the flight. RESULTS: Passengers on airplanes that did and did not recirculate air had similar rates of postflight respiratory symptoms. The rates of reporting a cold were 19% vs 21% (P =.34); a runny nose and a cold, 10% vs 11%, (P =.70); and an aggregation of 8 URI symptoms, 3% in both groups (P>.99). Results were similar after statistical adjustment for potential confounders. CONCLUSION: We found no evidence that aircraft cabin air recirculation increases the risk for URI symptoms in passengers traveling aboard commercial jets. PMID- 12132980 TI - An 83-year-old woman with chronic illness and strong religious beliefs. PMID- 12132981 TI - A 28-year-old woman with panic disorder, 1 year later. PMID- 12132982 TI - Elderly patients' adherence to statin therapy. PMID- 12132983 TI - Smoking cessation trials targeted to racial and economic minority groups. PMID- 12132984 TI - What practices will most improve safety? Evidence-based medicine meets patient safety. PMID- 12132985 TI - Safe but sound: patient safety meets evidence-based medicine. PMID- 12132990 TI - Chemistry and mechanism of urease inhibition. AB - Studies on enzyme inhibition remain an important area of pharmaceutical research since these studies have led to the discoveries of drugs useful in a variety of physiological conditions. The enzyme inhibitors can interact with enzymes and block their activity towards natural substrates. Urease inhibitors have recently attracted much attention as potential new anti-ulcer drugs. Ironically, urease was the first enzyme crystallized but its mechanism of action is still largely misunderstood. This chapter therefore reviews comprehensive developments in the field of urease inhibitors. Inhibitors of urease can be broadly classified into two categories: (1) active site directed (substrate-like), (2) mechanism-based directed. We present here the examples of selected inhibitors along with their mechanisms of action to characterize their mode of urease inhibition. The observations that urease due to its high substrate (urea) specificity can only bind to a few inhibitors with a similar binding mode as urea is also discussed. Several non-covalent interactions including hydrogen bonds and hydrophobic contacts stabilize the enzyme-inhibitor complex. Regardless of the class of compound, it is reported that only a few functional groups with electronegative atoms such as oxygen, nitrogen and sulfur act either as bidentate (mostly), tridentate (rarely), or as ligand-chelator to form octahedral complexes with two slightly distorted octahedral Ni ions of the enzyme. Bulky groups attached to the pharmacophore were found to decrease the activity of inhibitors, since the lack of a bulky attachment makes it easier for urease inhibitors to enter the substrate-binding pocket as well as avoid unfavorable steric interactions with amino acid residues in its vicinity. This review is intended to provide highlights of the inhibition of urease by hydroxamic acids (HXAs), phosphorodiamidates (PPDs), imidazoles, phosphazene and related compounds. These compounds are compared to previously reported urease inhibitors for the catalytic models proposed for urease activity. The differences in inhibition of urease activities from plants and of bacterial origin by various inhibitors and physiological implications of urease inhibition are discussed. PMID- 12132991 TI - Effect of alpha-FMH and DPPE on colony-forming properties of human peripheral progenitor cells. AB - Endogenous histamine regulates the haematopoiesis. Histidine decarboxylase inhibitor decreases the histamine level, and its intracellular antagonist decreases the histamine effect. The effect of histidine decarboxylase inhibitor (alpha-fluoromethyl histidine) and the intracellular antagonist of histamine [N'N diethyl-2-4-(phenylmethyl) phenoxyethan-amine-HCl] was investigated on the colony forming ability of human peripheral progenitor cells. Semi-solid culture medium was used both in the presence and in the absence of 3 U/ml erythropoietin. alpha Fluoromethyl histidine was used in the range of 50 through 150 micro Mol/ml, the concentration of N'N-diethyl-2-4-(phenylmethyl) phenoxyethanamine-HCl was between 5 and 25 micro Mol/ml. The number of both the erythroide and the granulocyte macrophage colony was significantly decreased in a concentration dependent manner by the presence of both N'N-diethyl-2-4-(phenylmethyl) phenoxyethanamine-HCl (in all concentrations used) and &alpha-fluoromethyl histidine (at higher concentration). The inhibitory effect was decreased by erythropoietin. PMID- 12132992 TI - The effects of speech therapy and pharmacological treatments on voice and speech in Parkinson s disease: a review of the literature. AB - The purpose of this paper was to examine the effects of speech therapy and various pharmacological treatment approaches on the voice and speech of persons with Parkinson s disease (PD). Approximately 80% of PD patients have voice and speech problems including reduced vocal intensity, reduced vocal pitch, monopitch and monoloudness, and imprecise articulation. Research prior to 1970's had not demonstrated significant improvements following speech therapy. However, recent research has shown that speech therapy (when persons with PD are optimally medicated) has proven to be the most efficacious therapeutic method for improving voice and speech function. Across research studies, pharmacological methods of treatment in isolation do not appear to significantly improve voice and speech function in PD. In a single subject study, however, the dopamine agonist Mirapex was shown to have beneficial effects on vocal intensity. Possible explanations for the differential responses to treatment are discussed. It is suggested that the goal of future studies should be investigations of the effects of combined treatment approaches. PMID- 12132993 TI - QSAR aspects of flavonoids as a plentiful source of new drugs. AB - The continuous raise of molecular diversity requires to find a guidelines for orientation it this. A likely occasion for consideration of new QSAR approach possibilities for this aspect is the flavonoids (F) multitude, the class of organic compounds (C) with very varied chemistry and pharmacology. The list of F affinities includes benzodiazepines, adenosine, estrogen receptors and row of enzymes phosphodiesterase, proteinthyrosine kinase, aromatase, xanthine oxidase asf. The simple way of clasterization of C is their numerical representation as the vector in linear space. The components of the some vectors are determined by parameters of molecular shape (number of cycles, tertiary atoms asf) and orbital structure indices (numbers of occupied orbitals of different kind). The nonesterificied F are tricyclic C with numbers of tertiary atoms n = 5,.,11 and more, from generic flavon (flavan), n = 5, to myricetin (n = 11) and other. The all derivatives of F with given n may be associated with vectors of linear space, alpha-set, with linear form sigma = 2l + m, l number of nonhydrogenous atoms, m = 0, 1,. The collection of sigma-sets which numbered by given number n = 5,.11,. envelops all natural F and their derivatives which have not additional cycles. In mentioned sigma-sets of tricyclic C by the some mode are presented ligands of benzodiazepines, adenosine, estrogens and other receptors and enzymes. Positions of ligands of each class form a triangle "sector of affinity" in 4-5 adjacent sigma-sets. These triangle sectors in mentioned collection of sigma-sets are disposed in conformity to certain laws. The position of given F relatively to "sectors of affinity" allow to found their affinity (and cross-affinity) to different targets. Calculations with utilization of sophisticated shape parameters and electronic structure indices allow to determine the affinity and/or activity quantitative measure. PMID- 12132994 TI - Large-conductance, ca(2+)-activated k(+) channels: function, pharmacology and drugs. AB - Because of the physiological role played by the hyperpolarisation process resulting from a K(+) outflow, it is not surprising that compounds able to activate outward K(+) channels are considered as promising drugs, with exciting perspectives for the treatment of several cardiovascular, respiratory, neurological and urological diseases. Among the different and numerous K(+) channel families, medicinal chemistry has focused its major interest onto two channel types: the ATP-sensitive channels (K(ATP)) and the large conductance subtype (BK), that belongs to the wide family of calcium-activated K(+) channels. BK channels are almost ubiquitous and exhibit single channel conductance of 100 300 pS, a property which justifies the potent role of these channels in the control of the membrane potential. BK channels have been investigated as potential therapeutic targets for different neuropathies, because of their profound influence on the neuronal activity. Moreover, BK channels are expected to have applications for the therapy of cardiovascular diseases. A potent feed back control of the vascular and non-vascular smooth muscle tone is mediated by these channels, whose activation can be promoted by both a rise of the intracellular free calcium concentration as well as a membrane depolarisation. Additionally, BK channel activation can also be induced by other factors, such as cAMP-mediated phosphorylation, G-proteins, GMP and cGMP. The aim of this paper is to give a concise overview of the biological and pharmacological properties and potential therapeutic applications of activators of BK channels present at the vascular level. The "state of the art" in the pharmaceutical development of natural and synthetic BK-activators, with a description of the lead chemical structures, will be also described. PMID- 12132995 TI - Regulating cysteine protease activity: essential role of protease inhibitors as guardians and regulators. AB - Cysteine proteases are widespread in nature. Their implication in numerous vital processes and pathologies make them highly attractive targets for drug design. The proper functioning and regulation of activity of cysteine proteases is a delicate balance of many factors, one of the most crucial being the protease inhibitors. In this review the basic principles of physiological protease inhibition by protein inhibitors are discussed with the focus on papain-like lysosomal cysteine proteases and the caspases, and their inhibitors. PMID- 12132996 TI - Thiol-dependent cathepsins: pathophysiological implications and recent advances in inhibitor design. AB - Thirteen papain-like cysteine proteases (cathepsins) are coded in the human genome from which two represent pseudogenes. Initially it was believed that lysosomal cysteine proteases primarily fulfill house-keeping functions which would exclude them as potential drug targets. Within the last decade, this view has dramatically changed and highly specific and therapeutically relevant functions have been assigned to individual cathepsins. Cathepsins are critical for osteoclast-mediated bone resorption and cartilage erosion in arthritis. They are involved in various aspects of immune responses, in the development and proliferation of various cell types, as well as in tumor invasion and metastasis. Cathepsins qualify as pharmaceutical targets for the treatment of osteoporosis, arthritis, asthma, autoimmune diseases, and potentially for certain forms of cancer. The major challenge in using cysteine protease inhibitors will be the design of highly selective, potent, and bioavailable compounds. Emerging novel functions of long-known and recently discovered cathepsins will require more emphasis on the selectivity of drug candidates to avoid adverse side effects. This review will focus on the discussion of presently known functions of papain like cathepsins and on recent advances in the design of cysteine protease inhibitors. PMID- 12132997 TI - Cysteine proteases of malaria parasites: targets for chemotherapy. AB - New drugs to treat malaria are urgently needed. Cysteine proteases of malaria parasites offer potential new chemotherapeutic targets. Cysteine protease inhibitors block parasite hemoglobin hydrolysis and development, indicating that cysteine proteases play a key role in hemoglobin degradation, a necessary function of erythrocytic trophozoites. These inhibitors also block the rupture of erythrocytes by mature parasites, suggesting an additional role for cysteine proteases in the hydrolysis of erythrocyte cytoskeletal proteins. Recent studies have shown that the repertoire of cysteine proteases of malaria parasites is larger than was previously realized. Plasmodium falciparum, the most virulent human malaria parasite, expresses three papain-family cysteine proteases, known as falcipains. All three proteases are expressed by trophozoites and hydrolyze hemoglobin at acidic pH, suggesting roles in this process. Falcipain-2 also hydrolyzes ankyrin at neutral pH, suggesting additional activity against erythrocyte cytoskeletal targets. Multiple orthologs of the falcipains have been identified in other plasmodial species. Analysis of orthologs from animal model rodent parasites identified similar features, but some noteworthy biochemical differences between the cysteine proteases. These differences must be taken into account in interpreting in vivo experiments. A number of small molecule cysteine protease inhibitors blocked parasite hemoglobin hydrolysis and development, and inhibitory effects against parasites generally correlated with inhibition of falcipain-2. Some compounds also cured mice infected with otherwise lethal malaria infections. Current research priorities are to better characterize the biological roles and biochemical features of the falcipains. In addition, efforts to identify optimal falcipain inhibitors as antimalarials are underway. PMID- 12132998 TI - Molecular modeling in cysteine protease inhibitor design. AB - The application of various molecular modeling techniques has been recently reported in the design of several new cysteine protease inhibitors. Computational chemistry techniques have been used to understand and predict enzyme-inhibitor interactions and also to study enzyme mechanism and inhibitor reactivity. This review focuses on examples that use structure-based design or reflect cysteine proteases as a target class. In several cases X-ray crystallography and molecular modeling have significantly facilitated the inhibitor design process. Cysteine proteases can present extra challenges in molecular modeling, due to the covalent binding modes and the reactive nature of many of the inhibitors. We also discuss some of the key challenges in developing new tools to evaluate these properties and help in making informed decisions about new templates and leads. PMID- 12132999 TI - Exoenzyme S shows selective ADP-ribosylation and GTPase-activating protein (GAP) activities towards small GTPases in vivo. AB - Intracellular targeting of the Pseudomonas aeruginosa toxins exoenzyme S (ExoS) and exoenzyme T (ExoT) initially results in disruption of the actin microfilament structure of eukaryotic cells. ExoS and ExoT are bifunctional cytotoxins, with N terminal GTPase-activating protein (GAP) and C-terminal ADP-ribosyltransferase activities. We show that ExoS can modify multiple GTPases of the Ras superfamily in vivo. In contrast, ExoT shows no ADP-ribosylation activity towards any of the GTPases tested in vivo. We further examined ExoS targets in vivo and observed that ExoS modulates the activity of several of these small GTP-binding proteins, such as Ras, Rap1, Rap2, Ral, Rac1, RhoA and Cdc42. We suggest that ExoS is the major ADP-ribosyltransferase protein modulating small GTPase function encoded by P. aeruginosa. Furthermore, we show that the GAP activity of ExoS abrogates the activation of RhoA, Cdc42 and Rap1. PMID- 12133000 TI - Evidence that the dephosphorylation of Ser(535) in the epsilon-subunit of eukaryotic initiation factor (eIF) 2B is insufficient for the activation of eIF2B by insulin. AB - Eukaryotic initiation factor (eIF) 2B is a guanine-nucleotide exchange factor that plays a key role in the regulation of protein synthesis. It is activated by insulin, serum and other agents that stimulate general protein synthesis. The largest (epsilon) subunit of eIF2B is a substrate for glycogen synthase kinase (GSK)-3 in vitro, and phosphorylation by GSK3 inhibits the activity of eIF2B. The site of phosphorylation has previously been identified as Ser(535). GSK3 is inactivated by phosphorylation in response to insulin or serum. In Chinese hamster ovary cells, insulin and serum bring about the dephosphorylation of Ser(535) in vivo, concomitantly with the phosphorylation of GSK3, and these effects are mediated through signalling via phosphoinositide 3-kinase. We have made use of inhibitors of GSK3 to determine whether GSK3 is responsible for phosphorylation of Ser(535) in vivo and to explore the role of phosphorylation of Ser(535) in the regulation of eIF2B. Treatment of cells with LiCl or with either of two recently developed GSK3 inhibitors, SB-415286 and SB-216763, brought about the dephosphorylation of Ser(535), which strongly indicates that this site is indeed a target for GSK3 in vivo. However, these compounds did not elicit significant activation of eIF2B, indicating, consistent with conclusions from one of our previous studies, that additional inputs are required for the activation of eIF2B. Our results also show that each of the inhibitors used affects overall protein synthesis and have additional effects on translation factors or signalling pathways apparently unrelated to their effects on GSK3, indicating that caution must be exercised when interpreting data obtained using these compounds. PMID- 12133001 TI - Fine specificity of domain-I of recombinant tandem-repeat-type galectin-4 from rat gastrointestinal tract (G4-N). AB - Galectins, a family of beta-galactoside-specific endogenous lectins, are involved in regulating diverse activities such as proliferation/apoptosis, cell-cell (matrix) interaction and cell migration. It is presently unclear to what extent the carbohydrate fine specificities of the combining sites of mammalian galectins overlap. To address this issue, we performed an analysis of the carbohydrate recognition domain (CRD-I) near the N-terminus of recombinant rat galectin-4 (G4 N) by the biotin/avidin-mediated microtitre plate lectin-binding assay with natural glycoproteins (gps)/polysaccharide and by the inhibition of galectin glycan interactions with a panel of glycosubstances. Among the 35 glycans tested for lectin binding, G4-N reacted best with human blood group ABH precursor gps, and asialo porcine salivary gps, which contain high densities of the blood group Ii determinants Galbeta1-3GalNAc (the mucin-type sugar sequence on the human erythrocyte membrane) and/or GalNAcalpha1-Ser/Thr ( Tn ), whereas this lectin domain reacted weakly or not at all with most sialylated gps. Among the oligosaccharides tested by the inhibition assay, Galbeta1-3GlcNAcbeta1-3Galbeta1 4Glc was the best. It was 666.7 and 33.3 times more potent than Gal and Galbeta1 3GlcNAc, respectively. G4-N has a preference for the beta-anomer of Gal at the non-reducing ends of oligosaccharides with a Galbeta1-3 linkage, over Galbeta1-4 and Galbeta1-6. The fraction of Tn glycopeptide from asialo ovine submandibular glycoprotein was 8.3 times more active than Galbeta1-3GlcNAc. The overall carbohydrate specificity of G4-N can be defined as Galbeta1-3GlcNAcbeta1 3Galbeta1-4Glc (lacto- N -tetraose)>Galbeta1-4GlcNAcbeta1-3Galbeta1-4Glc (lacto- N -neo-tetraose) and Tn clusters>Galbeta1-4Glc and GalNAcbeta1-3Gal>Galbeta1 3GalNAc>Galbeta1-3GlcNAc>Galbeta1-4GlcNAc>GalNAc>Gal. The definition of this binding profile provides the basis to detect differential binding properties relative to the other galectins with ensuing implications for functional analysis. PMID- 12133002 TI - Purification, characterization and catalytic properties of human sterol 8 isomerase. AB - CHO 2, encoding human sterol 8-isomerase (hSI), was introduced into plasmids pYX213 or pET23a. The resulting native protein was overexpressed in erg 2 yeast cells and purified to apparent homogeneity. The enzyme exhibited a K (m) of 50 microM and a turnover number of 0.423 s(-1) for zymosterol, an isoelectric point of 7.70, a native molecular mass of 107000 Da and was tetrameric. The structural features of zymosterol provided optimal substrate acceptability. Biomimetic studies of acid-catalysed isomerization of zymosterol resulted in formation of cholest-8(14)-enol, whereas the enzyme-generated product was a Delta(7)-sterol, suggesting absolute stereochemical control of the reaction by hSI. Using (2)H(2)O and either zymosterol or cholesta-7,24-dienol as substrates, the reversibility of the reaction was confirmed by GC-MS of the deuterated products. The positional specific incorporation of deuterium at C-9alpha was established by a combination of (1)H- and (13)C-NMR analyses of the enzyme-generated cholesta-7,24-dienol. Kinetic analyses indicated the reaction equilibrium ( K (eq)=14; DeltaG(o')=-6.5 kJ/mol) for double-bond isomerization favoured the forward direction, Delta(8) to Delta(7). Treatment of hSI with different high-energy intermediate analogues produced the following dissociation constants ( K (i)): emopamil (2 microM)=tamoxifen (1 microM)=tridemorph (1 microM)<25-azacholesterol (21 microM) 1 mM (K(i(ED/TAM))=130+/-5 nM and K(i(ED/FAS))=2.7+/-0.9 microM). The cytoplasmic surface of the glucose transporter (GLUT)1 contains four sequences with close homologies to sequences in the ligand-binding domain of human oestrogen receptor beta (hesr-2). One homology is adjacent to the Walker ATP-binding motif II (GLUT1, residues 225-229) in the large cytoplasmic segment linking transmembrane helices 6 and 7; another GLUT (residues 421-423) contains the Walker ATP-binding motif III. Mapping of these regions on to a three-dimensional template of GLUT indicates that a possible oestrogen-binding site lies between His(337), Arg(349) and Glu(249) at the cytoplasmic entrance to the hydrophilic pore spanning GLUT, which have a similar topology to His(475), Glu(305) and Arg(346) in hesr-2 that anchor the head and tail hydroxy groups of oestradiol and genistein, and thus are suitably placed to provide an ATP-sensitive oestrogen binding site that could modulate glucose export. PMID- 12133006 TI - Evidence for crucial electrostatic interactions between Bcl-2 homology domains BH3 and BH4 in the anti-apoptotic Nr-13 protein. AB - Nr-13 is an anti-apoptotic member of the Bcl-2 family previously shown to interact with Bax. The biological significance of this interaction was explored both in yeast and vertebrate cells and revealed that Nr-13 is able to counteract the pro-apoptotic activity of Bax. The Bax-interacting domain has been identified and corresponds to alpha-helices 5 and 6 in Nr-13. Site-directed mutagenesis has revealed that the N-terminal region of Nr-13 is essential for activity and corresponds to a genuine Bcl-2 homology domain (BH4). The modelling of Nr-13, based on its similarity with other Bcl-2 family proteins and energy minimization, suggests the possibility of electrostatic interactions between the two N-terminal conserved domains BH4 and BH3. Disruption of these interactions severely affects Nr-13 anti-apoptotic activity. Together our results suggest that electrostatic interactions between BH4 and BH3 domains play a role in the control of activity of Nr-13 and a subset of Bcl-2 family members. PMID- 12133007 TI - Co-operation of the transcription factor hepatocyte nuclear factor-4 with Sp1 or Sp3 leads to transcriptional activation of the human haem oxygenase-1 gene promoter in a hepatoma cell line. AB - We reported previously that the 5'-flanking region (nucleotides -1976 to -1655) of the human haem oxygenase-1 ( hHO-1 ) gene enhances hHO-1 promoter activity in human hepatoma HepG2 cells, but not in HeLa cells [Takahashi, Takahashi, Ito, Nagano, Shibahara and Miura (1999) Biochim. Biophys. Acta 1447, 231-235]. To define more precisely the regulatory elements involved, in the present study we have functionally dissected this region and localized the enhancer to a 50 bp fragment (-1793 to -1744). Site-direct mutagenesis analysis revealed that two regions were responsible for this enhancer activity, i.e. a hepatocyte nuclear factor-4 (HNF-4) homologous region and a GC box motif homologous region. Mutation in either region alone moderately decreased enhancer activity. However, mutations in both regions reduced promoter activity to the basal level. Electrophoretic mobility-shift assays demonstrated that the P5-2 fragment (-1793 to -1744) interacted with at least two nuclear factors, i.e. HNF-4 and Sp1/Sp3. Co transfection experiments using Drosophila SL2 cells revealed that HNF-4 and Sp1/Sp3 synergistically stimulated the enhancer activity of the P5-2 fragment. These results indicate that co-operation of HNF-4 with Sp1 or Sp3 leads to the activation of hHO-1 gene expression in hepatoma cells. PMID- 12133008 TI - Temporin L: antimicrobial, haemolytic and cytotoxic activities, and effects on membrane permeabilization in lipid vesicles. AB - The temporins are a family of small, linear antibiotic peptides with intriguing biological properties. We investigated the antibacterial, haemolytic and cytotoxic activities of temporin L (FVQWFSKFLGRIL-NH2), isolated from the skin of the European red frog Rana temporaria. The peptide displayed the highest activity of temporins studied to date, against both human erythrocytes and bacterial and fungal strains. At variance with other known temporins, which are mainly active against Gram-positive bacteria, temporin L was also active against Gram-negative strains such as Pseudomonas aeruginosa A.T.C.C. 15692 and Escherichia coli D21 at concentrations comparable with those that are microbiocidal to Gram-positive bacteria. In addition, temporin L was cytotoxic to three different human tumour cell lines (Hut-78, K-562 and U-937), causing a necrosis-like cell death, although sensitivity to the peptide varied markedly with the specific cell line tested. A study of the interaction of temporin L with liposomes of different lipid compositions revealed that the peptide causes perturbation of bilayer integrity of both neutral and negatively charged membranes, as revealed by the release of a vesicle-encapsulated fluorescent marker, and that the action of the peptide is modulated to some extent by membrane lipid composition. In particular, the presence of negatively charged lipids in the model bilayer inhibits the lytic power of temporin L. We also show that the release of fluorescent markers caused by temporin L is size-dependent and that the peptide does not have a detergent like effect on the membrane, suggesting that perturbation of bilayer organization takes place on a local scale, i.e. through the formation of pore-like openings. PMID- 12133010 TI - Age-related determinants of outcome after acute myocardial infarction: a dobutamine-atropine stress echocardiographic study. AB - OBJECTIVES: To investigate the cause of worse survival in older patients after myocardial infarction (MI). DESIGN: Prospective 18-month and longer follow-up study of a cohort of 167 patients (mean age +/- standard deviation 58 +/- 12, including 71 aged >or=65) with acute MI for cardiac events, defined as cardiac death, recurrent MI, or resuscitated ventricular tachycardia or fibrillation (VT/VF). SETTING: Milwaukee County Medical Complex and the Zablocki Veterans Affairs Medical Center, Milwaukee, WI. PARTICIPANTS: One hundred sixty-seven patients who underwent dobutamine-atropine stress echocardiography (DASE) in the first week (2-7 days) after acute MI and were medically managed. MEASUREMENTS: Comparison of event rates in older (>or=65 years) and younger (<65 years) patients and of clinical, resting echocardiographic, DASE, and angiographic findings in patients with and without events. Coronary angiography was performed in 141. RESULTS: Older and younger patients tolerated DASE well. During follow up, there were 29 cardiac events (cardiac death in 17, nonfatal MI in 10, and VT/VF in 2). Events were more common in older patients (26% vs 12%, P <.05), especially death (19% vs 5%, P <.05). Scar size in the infarct zone by DASE was larger (4.0 +/- 2.8 vs 3.0 +/- 2.7 segments, P <.05) and remote wall motion abnormalities more common (47 vs 29%, P <.05) in older patients. Univariate determinants of outcome (P <.05) in older patients were diabetes mellitus; remote wall motion abnormalities; angiographic multivessel disease; scar size; ejection fraction; and resting, low-, and peak-dose wall motion score. Univariate determinants in younger patients were similar, but diabetes mellitus was not. Multivariate analysis revealed that remote wall motion abnormalities and scar size by DASE were independently predictive of outcome in older and younger patients and diabetes mellitus only in older patients. Low- and peak-dose DASE data enhanced (P <.01) the prediction of outcome in all patients with acute MI relative to clinical data and resting echocardiography. CONCLUSION: DASE was more predictive of outcome than clinical data and resting echocardiography in both age groups. Scar size and remote wall motion abnormalities were the primary determinants of outcome irrespective of age. The worse prognosis of older patients correlated with diabetes mellitus, greater scar size, and higher incidence of remote inducible ischemia. PMID- 12133011 TI - Preoperative electrocardiogram abnormalities do not predict postoperative cardiac complications in geriatric surgical patients. AB - OBJECTIVES: Preoperative electrocardiograms (ECGs) are routinely performed on older patients before surgery. Whether patients with abnormalities on preoperative ECGs have an increased likelihood of developing postoperative cardiac complications is unknown. This study was designed to determine whether abnormalities on preoperative ECGs were predictive of postoperative cardiac complications. DESIGN: Prospective observational study. SETTING: One of the teaching hospitals of the University of California, San Francisco, Medical Center. PARTICIPANTS: Five hundred thirteen patients aged 70 and older undergoing noncardiac surgery. MEASUREMENTS: Preoperative ECGs were analyzed using the Minnesota Codes. Predefined preoperative risk factors and in-hospital postoperative cardiac complications were measured. The association between ECG abnormalities and postoperative cardiac complications was determined by multivariate logistic regression after controlling for clinical covariates. Odds ratios (ORs) and 95% confidence intervals (CIs) were reported. RESULTS: Three hundred eighty-six of 513 patients (75.2%) had at least one abnormality on their preoperative ECGs. On multivariate analysis, the predictors of postoperative cardiac complications included American Society of Anesthesiologists physical status classification of 3 or greater (OR = 2.5, 95%CI = 1.28-4.89, P = .007) and a history of congestive heart failure (OR = 2.1, 95% CI = 1.1-5.1, P = .034). The presence of abnormalities on preoperative ECGs was not associated with an increased risk of postoperative cardiac complications (OR = 0.63, 95% CI = 0.28 1.40, P = .26). CONCLUSION: Abnormalities on preoperative ECGs are common but are of limited value in predicting postoperative cardiac complications in older patients undergoing noncardiac surgery. These results suggest that obtaining preoperative ECGs based on an age cutoff alone may not be indicated, because ECG abnormalities in older people are prevalent but nonspecific and less useful than the presence and severity of comorbidities in predicting postoperative cardiac complications. PMID- 12133012 TI - Does advanced age affect the immediate and long-term results of direct-current external cardioversion of atrial fibrillation? AB - OBJECTIVES: To determine whether advanced age affects the immediate and long-term results of direct-current external cardioversion (ECV) of atrial fibrillation (AF), the sustained arrhythmia most commonly encountered in older patients. DESIGN: Retrospective analysis of medical records. SETTING: Intensive care unit. PARTICIPANTS: Two hundred fifty consecutive patients(age 34-100) with AF who underwent ECV following a standardized protocol in an intensive care unit. MEASUREMENTS: Immediate efficacy of ECV, defined as recovery of sinus rhythm, and maintenance of sinus rhythm over the follow-up were study outcomes. The univariate and multivariate associations of immediate efficacy of ECV and long term results with clinical variables were analyzed. RESULTS: At univariate analysis, immediate efficacy of ECV (overall, 91.2%) was lower in older patients and in those with chronic obstructive pulmonary disease, higher for a 3- to 90 day pre-ECV duration of AF than for a duration of 2 days or less or more than 90 days, and independent of underlying cardiac disease, hypertension, diabetes mellitus, previous AF, and left atrial dimension. However, pre-ECV duration of AF was the only multivariate predictor of ECV immediate success. Major complications occurred in only three patients. Of 220 patients discharged in sinus rhythm, 211 were followed up for a mean period +/- standard deviation of 34 +/- 25 months. AF relapsed in 45.5% of them. At multivariate analysis, underlying cardiac disease was the only predictor of the relapse rate, and relapse rate was lower in coronary heart disease than in valvular heart disease, congestive heart failure, or lone AF. CONCLUSION: Immediate and long-term results of ECV of AF, an effective and safe procedure, are unaffected by age,at least after adjusting for several covariates. PMID- 12133013 TI - Does aspirin attenuate the effect of angiotensin-converting enzyme inhibitors on health outcomes of very old patients with heart failure? AB - OBJECTIVES: Concomitant ischemic heart disease (IHD) is common in older individuals with heart failure (HF). We estimated the effect of aspirin use on the rate of mortality, morbidity, and decline in physical functioning in nursing home residents with HF taking angiotensin-converting enzyme (ACE) inhibitors. DESIGN: We conducted a retrospective cohort study using a nursing home database linking resident information collected via the Minimum Data Set (MDS) with drug utilization data. SETTING: Nursing homes in four states (1992-1995). PARTICIPANTS: Of 49,779 residents with HF admitted to these homes, 12,703 residents were taking an ACE inhibitor; 2,046 of these took aspirin. MEASUREMENTS: Medicare enrollment files provided the date of death, and we used the Part A Medicare files to identify hospital admissions. The activity of daily living scale from repeat MDS assessments allowed us to evaluate decline in physical function. Cox proportional hazards models provided adjusted estimates of the aspirin effect, with nonusers as the reference group. RESULTS: The overall mortality rate, hospitalization rate,and rate of decline in physical function of aspirin users were not different from those of nonusers (e.g., hospitalization rate ratio = 0.99, 95% confidence interval = 0.92-1.07). This effect did not vary by presence of concomitant IHD or by dose or type of ACE inhibitor. CONCLUSION: In a cohort of older HF residents receiving ACE inhibitors in nursing homes, we found that treatment with aspirin did not appear to affect outcomes negatively. PMID- 12133014 TI - Age and the risk of in-hospital death: insights from a multihospital study of intensive care patients. AB - OBJECTIVES: To determine independent relationships between age and the risk of in hospital death. DESIGN: Retrospective cohort study. SETTING: Thirty-eight intensive care units (ICUs) in 28 hospitals in a large Midwest metropolitan region. PARTICIPANTS: One hundred fifty-six thousand, one hundred thirty-six consecutive admissions to medical, surgical, neurological, and mixed medical/surgical ICUs between March 1, 1991, and March 31, 1997. MEASUREMENTS: In hospital death rates were compared at successive 5-year age intervals, adjusting for gender, diagnosis, admission source, comorbidity, and acute physiology scores. Acute physiology scores were determined using a validated methodology based on abnormalities in 17 physiological measures collected during the first 24 hours of ICU admission. RESULTS: The adjusted odds of death increased with each 5 year age increment. For example, relative to patients younger than 35, adjusted odds of death in patients aged 40 to 44, 50 to 54, 60 to 64, 70 to 74, 80 to 84, and 90 and older were 1.51, 1.73, 2.38, 2.98, 3.86, and 4.74, respectively. In stratified analyses, age-related increases in the odds of death were somewhat higher in surgical than medical patients or patients with lower severity of illness at admission. Although acute physiology scores had excellent discrimination in all age groups, discrimination decreased with age (e.g., c statistics of 0.928 and 0.835 in patients younger than 45 and 85 and older, respectively). CONCLUSION: Our findings demonstrate incremental increases in the risk of hospital death associated with age that was independent of severity of illness and other prognostic factors. Although the current results may be less biased by differences in treatment goals than studies of general hospitalized patients, the lower discrimination of physiology scores in older patients suggests that unmeasured factors (e.g., functional status, patient preferences for care, differences in physician practices) may be of greater prognostic importance in older than in younger patients. PMID- 12133015 TI - Unrecognized tuberculosis in a nursing home causing death with spread of tuberculosis to the community. AB - OBJECTIVES: To determine the reason for an increase in tuberculin skin test (TST) conversion in employees in a nursing home and to determine the source case responsible for spread of tuberculosis (TB) in two nursing homes and a hospital in a rural part of Arkansas using molecular and traditional epidemiological methods. DESIGN: TB contact investigation of residents and employees of two nursing homes and a hospital. SETTING: Two nursing homes and a hospital in rural part of Arkansas. PARTICIPANTS: One hundred fifty-seven employees and 117 residents of two nursing homes and 211 employees of a hospital in rural part of Arkansas. MEASUREMENTS: Tuberculin skin test. RESULTS: Analysis of room and work assignments of residents and employees who converted their TSTs in Nursing Home A showed that residents and employees in the same wing as the suspect source case were significantly more likely to have converted their TST than residents and employees in other wings (P = .01). A nurse from the local hospital where the suspected source case had been sent developed a tuberculous cervical abscess, and one employee in Nursing Home A developed pulmonary TB. A visitor to Nursing Home A was diagnosed with culture-positive pulmonary TB 2 years later. Genotyping of the Mycobacterium tuberculosis isolates from the four secondary cases showed identical patterns. CONCLUSION: Molecular and traditional epidemiological studies revealed an outbreak of TB that began in a nursing home and spread to a second nursing home, a local hospital, and the community. PMID- 12133016 TI - Tuberculosis in older people: a retrospective and comparative study from Hong Kong. AB - OBJECTIVES: To compare tuberculosis (TB) in older and younger patients. DESIGN: A retrospective and comparative observational study. SETTING: Four chest clinics and two chest hospitals in Hong Kong. PARTICIPANTS: All notifications from the participating hospitals and clinics in 1996 were extracted from the TB notification registry. The characteristics of patients aged 65 and older were compared with a one-in-three random sampling of those aged 16 to 64. MEASUREMENTS: Demographic, clinical, radiological, and laboratory data of the two groups were compared alongside treatment and outcomes. RESULTS: Older people with TB were more likely to be male, to smoke, to have had TB previously, to have coexisting medical diseases, to be socioeconomically disadvantaged, and to weigh less than younger people with TB. Dyspnea, weight loss, and malaise were more common, whereas extrathoracic lymph node enlargement was less common. Chest radiograph showed more extensive disease and lower zone involvement. Positive tuberculin test was present in only 61.9%. Sputum bacteriology was more likely to be positive. There was a longer delay in presentation and commencement of treatment, and 77.2% required at least one admission. Adverse effects of treatment, notably hepatic dysfunction, occurred more commonly. Fluoroquinolones appeared well tolerated. Only 72.5% of the older people were cured or completed their treatment. Mortality was 16%. Age of 65 and older, comorbidity, socioeconomic disadvantage, moderate-extensive disease, positive sputum smear, and poor adherence were factors independently associated with unfavorable outcomes (P <.001 to P = .01; odds ratios = 1.61-27.02). CONCLUSION: Substantial differences were found between older and younger TB patients. Many of these were associated with unfavorable outcome. Increased awareness in disease recognition and better medical and social support are therefore needed in addressing the growing problem of TB in older people. PMID- 12133017 TI - Plasma homocysteine and cognitive impairment in an older British African Caribbean population. AB - OBJECTIVES: To investigate the association between homocysteine concentrations and cognitive impairment in an older African-Caribbean population. To investigate other measures of risk for vascular disease and nutritional status as potential confounding factors. DESIGN: A secondary analysis from a cross-sectional community study. SETTING: The sample was drawn from registration lists for seven primary care services in south London, United Kingdom. PARTICIPANTS: Two hundred forty-eight individuals aged 55 to 75 who were born in a Caribbean nation and for whom homocysteine concentrations had been ascertained. MEASUREMENTS: Plasma homocysteine and serum folate were assayed from frozen samples. Cognitive impairment as a composite measure was derived from 11 psychometric tests. Other measures of risk for vascular disease were considered as potential confounding factors: diagnosed hypertension/diabetes mellitus; physical exercise; and concentrations of cholesterol, triglycerides, and fibrinogen. RESULTS: Cognitive impairment was classified in 68 (27%) participants. Raised homocysteine (highest quartile: >13.85 micromol/L) was significantly associated with cognitive impairment (odds ratio (OR)=2.50, 95% confidence interval (CI)=1.33-4.69). This association persisted after adjustment for age, occupation, other measures of vascular risk, folate, body mass index, and waist:hip ratio (OR=3.00, 95% CI=1.35 6.69). As with other risk factors for vascular disease in this sample, the association was significant only in those with less education (P-value for interaction=.049). CONCLUSION: Raised homocysteine was associated with cognitive impairment in this sample, but this was modified by previous educational attainment. This association was independent of other measures of vascular risk and was not explained by folate concentrations. PMID- 12133018 TI - The prevalence of dementia in older people in an urban population of Korea: the Seoul study. AB - OBJECTIVES: To estimate prevalence of dementia and its subtypes in older people in Seoul, a metropolitan area of Korea, and compare these findings with estimates reported for other populations. DESIGN: The study employed a two-stage design for case identification. Initially, the Mini-Mental State Examination in the Korean version (MMSE-KC) of the Consortium to Establish a Registry for Alzheimer's Disease (CERAD) assessment packet was administered to all participants. Two hundred seventeen persons sampled from three levels of performance on MMSE-KC underwent the second-stage clinical evaluation based on the Korean Version of the CERAD assessment packet. SETTING: The study was conducted in an urban community setting. PARTICIPANTS: Six hundred forty-three persons aged 65 and over participated in the study. MEASUREMENTS: Dementia was defined using the Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition diagnostic features of dementia. RESULTS: Dementia prevalence ranged from 2.6% in persons aged 65 to 69 to 32.6% in persons aged 85 and older. Age-standardized prevalence was 8.2% for dementia, 5.4% for Alzheimer's disease, and 2.0% for vascular dementia. The prevalence estimates, which excluded very mild cases (clinical dementia rating index 0.5), were approximately 5.3% for dementia and 4.3% for Alzheimer's disease. CONCLUSION: The prevalence of dementia in older people in Seoul appears to be somewhat lower than in rural areas of Korea. Considering the difficulties involved in establishing a diagnostic threshold for dementia, actual differences in dementia prevalence between Asian populations are probably minimal. PMID- 12133019 TI - Patients with hip fracture: subgroups and their outcomes. AB - OBJECTIVES: To present several alternative approaches to describing the range and functional outcomes of patients with hip fracture. DESIGN: Prospective study with concurrent medical records data collection and patient and proxy interviews at the time of hospitalization and 6 months later. SETTING: Four hospitals in the New York metropolitan area. PARTICIPANTS: Five hundred seventy-one hospitalized adults aged 50 and older with hip fracture between July 1997 and August 1998. MEASUREMENTS: Rates of return to function in four physical domains, mortality, and nursing home residence at 6 months. Cluster analysis was used to describe the heterogeneity among the sample and identify variations in 6-month mortality, nursing home residence, and level of functioning and to develop a patient classification tree with associated patient outcomes at 6 months postfracture. RESULTS: In locomotion, transfers, and self-care, 33% to 37% of patients returned to their prior level of function by 6 months, including those needing assistance, but only 24% were independent in locomotion at 6 months. Cluster analysis identified eight patient subgroups that had distinct baseline features and variable outcomes at 6 months. The patient classification tree used four variables: atypical functional status (independent in locomotion but dependent in other domains); nursing home residence; independence/dependence in self-care; and age younger than 85 or 85 and older that identified five subgroups with variable 6-month outcomes that clinicians may use to predict likely outcomes for their patients. CONCLUSION: Patients with hip fracture are heterogeneous with respect to baseline and outcome characteristics. Clinicians may be better able to give patients and caregivers information on expected outcomes based on presenting characteristics used in the classification tree. PMID- 12133021 TI - Looking at the relationship between hemoglobin concentration and prevalent mobility difficulty in older women. Should the criteria currently used to define anemia in older people be reevaluated? AB - OBJECTIVES: The World Health Organization (WHO) and other currently used criteria for defining anemia in older women are mainly based on statistical distribution considerations. To explore their clinical appropriateness, we evaluated the relationship between hemoglobin (Hb) concentration, prevalent mobility difficulty, and the Summary Performance Score (SPS). DESIGN: Cross-sectional study. SETTING: Two population-based studies, the Women's Health and Aging Studies I and II, Baltimore, Maryland. PARTICIPANTS: Six hundred thirty-three community-dwelling women aged 70 to 80 with Hb levels obtained within 90 days from baseline assessment. MEASUREMENTS: Mobility difficulty (self-reported difficulty walking one-quarter of a mile or climbing 10 steps (primary outcome)). SPS, a performance-based summary measure of lower extremity function that combines the results of walking, chair stands, and balance tests (secondary outcome). RESULTS: Mobility difficulty prevalence was not constant within the WHO "normal" Hb range (12.0-16.0 g/dL). For example, a Hb of 13.5 g/dL was associated with a significantly lower mobility difficulty prevalence than a Hb of 12.0 g/dL (OR=0.68, 95% CI=0.47-0.93), even after adjustment for chronic diseases and other relevant health indicators. A consistent trend of improvement in performance based scores with increasing Hb categories less than 12.0 g/dL, 12.0 to 13.0g/dL, and 13.0-14.0 g/dL was observed. CONCLUSION: Our findings raise two hypotheses: (1) Hb currently perceived as "mildly-low" and even "low-normal" might have an independent, adverse effect on mobility function, and (2) Hb of 12.0 g/dL might be a suboptimal criterion for defining anemia in older women. Formal testing of these hypotheses might prove relevant for anemia- and mobility disability-related clinical decision-making. PMID- 12133020 TI - Handgrip strength and mortality in older Mexican Americans. AB - OBJECTIVES: To examine the association between handgrip strength and mortality in older Mexican American men and women. DESIGN: A 5-year prospective cohort study. SETTING: Five southwestern states: Texas, New Mexico, Colorado, Arizona, and California. PARTICIPANTS: A population-based sample of 2,488 noninstitutionalized Mexican-American men and women aged 65 and older. MEASUREMENTS: Maximal handgrip strength, timed walk, and body mass index were assessed at baseline during 1993/94. Self-reports of functional disability, various medical conditions, and status at follow-up were obtained. RESULTS: Of the baseline sample with complete data, 507 persons were confirmed deceased 5 years later. Average handgrip strength +/- standard deviation was significantly higher in men (28.4 kg +/- 9.5) than in women (18.2g +/- 6.5). Of men who had a handgrip strength less than 22.01 kg and women who had a handgrip strength less than 14 kg, 38.2% and 41.5%, respectively, were dead 5 years later. In men in the lowest handgrip strength quartile, the hazard ratio of death was 2.10 (95% confidence interval (CI) = 1.31 3.38) compared with those in the highest handgrip strength quartile, after controlling for sociodemographic variables, functional disability, timed walk, medical conditions, body mass index, and smoking status at baseline. In women in the lowest handgrip strength quartile, the hazard ratio of death was 1.76 (95%I = 1.05-2.93) compared with those in the highest handgrip strength quartile. Poorer performance in the timed walk and the presence of diabetes mellitus, hypertension, and cancer were also significant predictors of mortality 5 years later. CONCLUSION: Handgrip strength is a strong predictor of mortality in older Mexican Americans, after controlling for relevant risk factors. PMID- 12133022 TI - Self-reported driving, cognitive status, and physician awareness of cognitive impairment. AB - OBJECTIVES: To assess self-reported driving rates in older people and correlate these data with cognitive status and physician recognition of cognitive impairment. DESIGN: Cross-sectional study. SETTING: A multiphysician private practice clinic in a primarily Asian-American community of Honolulu, Hawaii. PARTICIPANTS: Two hundred ninety-seven ambulatory patients aged 65 and older. MEASUREMENTS: Cognitive function was assessed by physician interview using the Cognitive Abilities Screening Instrument (CASI) and proxy informant data. Subjects' self-reported driving status. Outpatient medical records were reviewed. RESULTS: Sixty percent of the studied population reported that they currently drove. This rate decreased from 73.3% (148/202) for subjects with good CASI performance (CASI 282) to 37.5% (21/56) for subjects with intermediate CASI performance (CASI 74-81.9) and further to 23.7% (9/38)for subjects with poor CASI performance (CASI <74). Further analysis of drivers with intermediate and poor CASI performance scores revealed that almost none of their physicians recognized that these drivers had cognitive problems(4.8% (1/21) of drivers with intermediate CASI performance and 11.1% (1/9) of drivers with poor CASI performance). CONCLUSION: In this convenience sample of older drivers, driving rates dropped precipitously with poorer performance on cognitive tests, yet a significant percentage of individuals with intermediate or poor cognitive test performance reported that they currently drove. This poor performance was often unrecognized by their physicians. Low recognition rates could affect physicians' interventions to curb unsafe driving. PMID- 12133023 TI - Serum interleukin-6 and hemoglobin as physiological correlates in the geriatric syndrome of frailty: a pilot study. AB - OBJECTIVES: To determine specific physiological correlates of the geriatric syndrome of frailty that warrant further investigation. DESIGN: Population-based case-control study. SETTING: General Clinical Research Center at Johns Hopkins Bayview Medical Center. PARTICIPANTS: Community-dwelling adults aged 74 and older from Baltimore, Maryland. MEASUREMENTS: Frailty status was determined using a recently validated screening tool that consists of weight loss, fatigue, low levels of physical activity, and measurements of grip strength and walking speed. Serum interleukin-6 (IL-6) was measured using enzyme-linked immunosorbent assay, and standard complete blood count was performed using a Coulter counter. RESULTS: Eleven frail and 19 nonfrail subjects with mean age +/- standard deviation of 84.9 +/- 6.7 vs 81.3 +/- 4.1 years, respectively, completed the study. The frail subjects had significantly higher serum IL-6 levels and significantly lower hemoglobin and hematocrit than the nonfrail subjects (4.4 +/-2.9 vs 2.8 +/- 1.6 pg/mL, 12.1 +/- 1.1 vs 13.9 +/- 1.0 g/dL, and 35.8% +/- 3.1% vs 40.6% +/- 2.8%, respectively). No significant difference was observed in mean corpuscular volume, red blood cell distribution width, or white blood cell and platelet counts between the frail and nonfrail groups. Furthermore, there was an inverse correlation between serum IL-6 level and hemoglobin (Pearson's correlation coefficient: -0.46) and hematocrit (-0.48) in the frail group but not in the nonfrail group. CONCLUSION: These results suggest that frail subjects have evidence of inflammation and lower hemoglobin and hematocrit levels. This subclinical anemia is normocytic and is hence unlikely due to myelosuppression or iron deficiency and is potentially related to the increased chronic inflammatory state marked by serum IL-6 elevation. Further studies are indicated to better characterize the immune and hematological changes that underlie frailty. PMID- 12133024 TI - Corrected arm muscle area: an independent predictor of long-term mortality in community-dwelling older adults? AB - OBJECTIVES: Older people are at risk of undernutrition because of a number of physiological conditions and lifestyle factors. The purpose of this study was to explore the predictive relationship of corrected arm muscle area (CAMA) with 8 year mortality in a representative sample of older Australians. DESIGN: Prospective cohort study: The Australian Longitudinal Study of Ageing. SETTING: Community. PARTICIPANTS: One thousand three hundred ninety-six participants aged 70 and older. MEASUREMENTS: Trained observers measured baseline weight, height, mid upper arm circumference, and triceps skinfold thickness using standard techniques. Body mass index (BMI) and CAMA were calculated. Baseline BMI and CAMA measurements were categorized according to cutoff values proposed by Garrow et al. and Friedman et al., respectively. Subsequent analyses were undertaken using Cox proportional hazards regression. RESULTS: After adjustment for potential confounders (baseline age, gender, marital status, smoking, self-rated health, ability to conduct activities of daily living, comorbidity, cognition performance, and presence of depression), those older Australians with a low CAMA (or=25% loss of renal function), irrespective of cause. Of the 12 studies that met these criteria, six were multicenter double blind placebo-controlled studies. The other six were smaller randomized studies. The studies had a mean +/- standard deviation follow-up of 3.2 +/- 0.3 years. One thousand one hundred two patients were randomized to receive ACE inhibitors or ARBs. Of these, 705 (64%) had data on renal function at baseline (within 6 months of the start) and at the end of the study. The authors examined the changes in serum creatinine levels or glomerular filtration rates (GFR) in patients who were randomized to receive ACE inhibitors. The authors also assessed the blood pressures achieved in the trials. RESULTS: Patients with preexisting chronic renal insufficiency who achieved their blood pressure control goals were likely to demonstrate an early rise in serum creatinine levels, approximately 25% above the baseline (approximately 1.7 mg/dL) after initiation of ACE inhibitor or ARB therapy. This rise in serum creatinine was more acute (by approximately 15% from the baseline) during the first 2 weeks of therapy and was more gradual (additional approximately 10%) during the third and fourth weeks of therapy (Figure 1). The serum creatinine level was likely to stabilize after about 4 weeks, provided patients had a normal salt and fluid intake. In addition, patients who did not show a rise in serum creatinine level during the first 2 to 4 weeks of therapy, were less likely to experience one after that period, unless they were dehydrated from use of diuretics or gastroenteritis or had used a nonsteroidal antiinflammatory drug (NSAID). In spite of this early rise in serum creatinine in patients with chronic renal insufficiency (a serum creatinine level of >or=124 micromol/L or >or=1.4 mg/dL) who were randomized to receive an ACE inhibitor, these patients receiving the drug showed a 55% to 75% lower risk of worsening renal function than those with normal renal function receiving the drug. The rate of risk reduction was inversely related to the severity of renal impairment at baseline, but data were limited on the benefit of ACE inhibitors in patients with more advanced renal insufficiency (GFR <30 mL/min). The authors noted that those aged 65 and older were likely to have much lower GFRs for given levels of serum creatinine than younger patients and were therefore likely to have advanced renal insufficiency at serum creatinine levels as low as 2 mg/dL (vs 4 mg/dL for younger patients). Patients with normal renal function were likely to show a much smaller rise in serum creatinine level (approximately 10% above the baseline of 0.9 mg/dL), mostly occurring during the first week after initiation of therapy, with subsequent stabilization, whereas patients with normal renal function suffering from heart failure, volume depletion, or bilateral renal artery stenosis experienced a significant rise (approximately 225% above baseline) in serum creatinine level, much higher in magnitude and rate than that experienced by those with renal insufficiency (Figure 1). Serum creatinine levels in these patients sharply increased (by approximately 75% above baseline) in the 2 weeks after the initiation of therapy with an ACE inhibitor, followed by an even sharper increase (another approximately 150%) during the subsequent 2 weeks. Patients with chronic renal insufficiency (serum creatinine>1.5 mg/dL) who received therapy with ACE inhibitors had about a five times higher risk of developing hyperkalemia than those with normal renal function, whereas presence of heart failure increased the risk of hyperkalemia by about three times over those without heart failure. Concomitant use of diuretics was associated with an approximately 60% reduction in risk of hyperkalemia. CONCLUSION: The authors conclude that, in patients with renal insufficiency (serum creatinine>1.4 mg/dL) treated with ACE inhibitors, there is a strong association between early (within the first 2 months) and moderate (not exceeding 30% over baseline) rise in serum creatinine and slowing of the renal disease progression in the long run. The authors recommend that ACE inhibitor therapy should not be discontinued unless serum creatinine level rise above 30% over baseline during the first 2 months after initiation of therapy or hyperkalemia (serum potassium level >or=5.6 mmol/L) develops. PMID- 12133030 TI - The preoperative electrocardiogram: have we reached the end of an era? PMID- 12133031 TI - The elusive great masquerader: the efficient spread of tuberculosis from a nursing home into the community. PMID- 12133032 TI - "Successful aging"--where next? PMID- 12133033 TI - Evaluating driving performance of cognitively impaired and healthy older adults: a pilot study comparing on-road testing and driving simulation. PMID- 12133034 TI - Low muscle mass of the thigh is significantly correlated with delirium and worse functional outcome in older medical patients. PMID- 12133035 TI - Urinary incontinence, condition-specific functional loss, and psychological distress. PMID- 12133036 TI - Anemia in centenarians. PMID- 12133038 TI - Bilateral otorrhagia due to a casual fall. PMID- 12133037 TI - Risk factors predicting pneumonia in patients aged 80 and older after cardiac surgery. PMID- 12133039 TI - A prospective study of the headache phase in 32 migraine with aura patients. AB - For an accurate description of the clinical features of the headache phase in migraine with aura (MA) attacks, we thought it useful to conduct a prospective study of consecutively referred MA patients seeking treatment at the Headache Centre of the University of Parma Institute of Neurology. The case series included 32 patients (22 women and 10 men). At the time of the first visit, each patient was given a questionnaire to be filled in at the next MA attack. Six patients (four women and two men) had attacks of migraine aura without headache. Among the remaining 26 patients (18 women and eight men), the duration of the headache phase was <24 h in 23 (88.5%); pain location was bilateral in 14 (53.8%) and unilateral in 12, but occurring on the opposite side to aura only in one patient; pain intensity was mild or moderate in 13 (50.0%). The headache phase of MA appeared to have clinical features that differed widely from patient to patient and was consistent with the International Headache Society diagnostic criteria for migraine without aura in 26.9% of patients and for tension-type headache (TTH) in 23.1%. PMID- 12133041 TI - An immunocytochemical investigation of human trigeminal nucleus caudalis: CGRP, substance P and 5-HT1D-receptor immunoreactivities are expressed by trigeminal sensory fibres. AB - 5-HT1D (but not 5-HT1B)-receptor immunoreactivity (i.r.) can be detected on trigeminal fibres within the spinal trigeminal tract of the human brainstem. The present study used immunohistochemical and morphometric techniques to determine the proportions of trigeminal fibres expressing substance P, CGRP or 5-HT1D receptor immunoreactivities. Co-localization studies between 5-HT1D-receptor and substance P- or CGRP-i.r. were also performed. Brainstem material was obtained with consent (four donors) and the total number of immunoreactive fibres within the trigeminal tract was estimated using random field sampling. A greater proportion of fibres (>1 microm diameter) expressed CGRP-i.r. (80 +/- 6%) compared with substance P-i.r. (46 +/- 7%) or 5-HT1D-receptor-i.r. (25 +/- 1%). 5 HT1D-receptor-i.r. was co-localized on some CGRP- or substance P-i.r. fibres. This suggests that 5-HT1D-receptors can regulate the release of CGRP and substance P and may be relevant to the clinical effectiveness of 5-HT1B/1D receptor agonists in the treatment of migraine and other cranial pain syndromes. PMID- 12133040 TI - Efficacy and safety of eletriptan 20 mg, 40 mg and 80 mg in Japanese migraineurs. AB - This prospective multicentre, double-blind, randomized, parallel-group, placebo controlled trial evaluated the efficacy and safety of a single dose of eletriptan 20 mg, 40 mg and 80 mg in Japanese migraineurs. A total of 402 adult Japanese migraineurs were diagnosed using International Headache Society (IHS) criteria. At 2 h after a single dose, the headache response rates of eletriptan 20 mg, 40 mg, 80 mg and placebo were 64%, 67%, 76% and 51%, respectively, with all doses significantly superior to placebo (P<0.05). Eletriptan had a statistically significant dose response for headache relief and pain-free response at 2 h post dose (P=0.0011 and P=0.0291, respectively). Most all-causality adverse events were mild and there were no deaths or discontinuations. Saliva samples were used to assess serum eletriptan levels 2 h post-dose. Pharmacokinetic evaluations showed no clinically significant differences between Japanese and Western subjects. Eletriptan was shown to be efficacious, safe, and well tolerated in Japanese migraineurs. PMID- 12133042 TI - Chronic daily headache in a tertiary care population: correlation between the International Headache Society diagnostic criteria and proposed revisions of criteria for chronic daily headache. AB - The International Headache Society (IHS) has been criticized for its approach to classification of chronic daily headache (CDH); Silberstein and Lipton criteria provide an alternative to this approach. The aim of this study is to apply the alternative diagnostic approaches to a sample of CDH patients consulting in specialty care. Our sample consisted of 638 patients with CDH. Patients were classified according to both classification systems. Patients were predominantly female (65.0%), with ages ranging from 11 to 88 years. According to the Silberstein and Lipton classification, we found eight different diagnoses. The most common diagnosis was chronic migraine (87.4%), followed by new daily persistent headache (10.8%). Just six patients had chronic tension-type headache (CTTH). Applying the IHS criteria we found 14 different diagnoses. Migraine was found in 576 (90.2%) patients. CTTH occurred in 621 (97.3%), with only 10 (1.57%) having this as the sole diagnosis. We conclude that both systems allow for the classification of most patients with CDH when daily headache diaries are available. The main difference is that the IHS classification is cumbersome and requires multiple diagnoses. The Silberstein and Lipton system is easier to apply, and more parsimonious. These findings support revision of the IHS classification system to include chronic migraine. PMID- 12133043 TI - Orthostatic headache syndrome with CSF leak secondary to bony pathology of the cervical spine. AB - The syndrome of orthostatic (low pressure) headaches is well described and most commonly occurs following deliberate violation of the dura (e.g. lumbar puncture). This syndrome can also occur spontaneously and results from the leakage of CSF. We describe three patients who suffered from spontaneous CSF leaks secondary to bony pathology of the cervical spine, and propose that this may be a more common aetiology than originally thought. Often these patients are difficult to manage medically, and surgery may be necessary for symptomatic relief. PMID- 12133044 TI - Exteroceptive suppression patterns of masseter and temporalis muscles in central and peripheral headache disorders. AB - The objective of this study was to compare the exteroceptive suppression patterns of masseter and temporalis muscles in patients with primary and secondary headache disorders originating from peripheral joint dysfunction. We accomplished the temporalis and masseter exteroceptive suppression in 28 patients with migraine, 25 patients with chronic tension-type headache (CTH), 22 patients with temporomandibular joint (TMJ) dysfunction and 18 healthy controls. The onset latencies and duration of the first suppression period (S1) was not significantly different between the patients and controls. The duration of the second suppression period (S2) was shorter in patients with CTH, migraine (analysed during attack) and TMJ dysfunction than those obtained from controls. A distinctive finding was significantly prolonged onset latency in patients with TMJ over those obtained from patients with CTH and migraine. We concluded that the onset latency of the S2 period is a useful parameter in the differential diagnosis of primary and peripheral headache disorders. PMID- 12133046 TI - Tractor drivers' head- and neck-ache: Vaga study of headache epidemiology. AB - The main aim was to study the occurrence of neck- and headache in farmers exposed to seasonal tractor work which involves prolonged, continuous neck rotation. As an integral part of the Vaga study of headache epidemiology, 109 farmers were interviewed according to a set scheme in a 'semistructured' interview, concerning complaints in the wake of tractor work. No appreciable headache/neck-ache was found in 13 farmers; neck-ache alone was present in 66 farmers; headache/neck ache was present in 15 farmers. For the remaining 15, the information was inadequate. Headache appeared only in connection with neck-ache. Only in those with headache did there seem to be a prevalence of nuchal features (such as reduced range of motion in the neck and positive skin-roll test). In 45% of cases, there was a carry-over effect after the chores, mostly of 1-3 days duration. Head- and neck-pain seem to be frequent in tractor drivers during chores. PMID- 12133045 TI - Almotriptan is an effective and well-tolerated treatment for migraine pain: results of a randomized, double-blind, placebo-controlled clinical trial. AB - Almotriptan is a novel and specific serotonin 5-HT1B/1D agonist for the acute treatment of migraine. This randomized, single-dose, double-blind, multicentre, study assessed the efficacy and safety of oral almotriptan (12.5 mg and 25 mg) in patients with migraine, and compared it with the standard treatment (sumatriptan 100 mg) and placebo. A total of 668 patients treated one migraine attack of moderate or severe intensity with study medication. The primary efficacy assessment was migraine pain relief, improvement from severe or moderate pain to mild or no pain, at 2 h after treatment. Response rates, stratified for variation in baseline pain levels, for both almotriptan doses were equivalent to sumatriptan and significantly better than placebo. Other efficacy assessments confirmed the equivalence of the almotriptan groups with the sumatriptan group. Almotriptan 12.5 mg was as well tolerated as placebo (P=0.493) and significantly better tolerated than sumatriptan (P<0.001), in terms of the overall incidence of adverse events. There was no statistically significant difference in the incidence of adverse events between almotriptan 25 mg and sumatriptan 100 mg (P=0.376). The results from this large clinical study indicate that the new, specific 5-HT1B/1D agonist, almotriptan, is an effective and well-tolerated treatment for migraine pain. PMID- 12133048 TI - Reproducibility and feasibility of neurophysiological assessment of the sensory trigeminal system for future application to paroxysmal headaches. AB - As the distribution of pain in primary headaches suggests involvement of the trigeminal sensory pathways, trigeminal somatosensory evoked potentials (TSEP) and blink reflexes (BR) may provide important information about their functional integrity. Functional differences between symptomatic and non-symptomatic sides and between measurements during and outside attacks may be particularly informative. These tests should therefore be reproducible and should require a suitable number of patients for future studies in patients with primary, paroxysmal headaches. We performed TSEP and BR twice in 22 healthy volunteers, in order to calculate sample sizes based on reproducibility data. This is, to our knowledge, the first study investigating the reproducibility of TSEP and BR measurements. Latencies of TSEP and BR are appropriate for future studies, as their reproducibility allows practical sample sizes (less than 25 subjects). Duration, amplitude and area parameters of the BR responses were less appropriate for longitudinal studies. PMID- 12133047 TI - Effects of sumatriptan on cerebral blood flow under normo- and hypercapnia in rats. AB - To investigate further the pharmacological mechanism of an anti-migraine drug, sumatriptan, a 5-HT1B/1D receptor agonist, we studied its effect on the cerebral circulation in seven anaesthetized rats, particularly during hypercapnia. After injection of 0.6 or 6.0 microg/kg sumatriptan succinate, no significant change in cerebral blood flow (CBF) was observed either in the striatum or in the parietal cortex. The increase in CBF both in the parietal cortex and the striatum during 5% CO2 inhalation was significantly less when sumatriptan succinate 6.0 microg/kg was injected. Sumatriptan appeared to have a vasoconstrictor effect on the relaxed vessels by CO2 inhalation. This mechanism might be attributable to vasoconstriction through activation of 5-HT1B receptors located in the vascular smooth muscle rather than 5-HT1B receptors in the vascular adventitia. PMID- 12133049 TI - Vagal nerve stimulation aborts migraine in patient with intractable epilepsy. PMID- 12133050 TI - Migraine with aura triggered by orgasm. PMID- 12133052 TI - Asymptomatic microscopic or dipstick haematuria in adults: which investigations for which patients? A review of the evidence. PMID- 12133053 TI - Laparoscopic adrenalectomy: comparison of transperitoneal and retroperitoneal approaches. AB - OBJECTIVE: To compare the effectiveness and efficiency of transperitoneal anterior laparoscopic adrenalectomy with that of retroperitoneal posterior laparoscopic adrenalectomy. PATIENTS AND METHODS: A retrospective comparison was undertaken of 28 patients who underwent transperitoneal anterior laparoscopy with 22 patients who underwent retroperitoneal posterior laparoscopy between April 1994 and November 2000. There were 18 men and 32 women (mean age 51.0 years, range 23-68) with varying diagnoses. Of the 50 patients, 31 had primary aldosteronism, eight had Cushing's syndrome, three had phaeochromocytoma, one had an 18-hydroxydeoxycorticosterone-induced adenoma and seven had non-functioning adrenal adenoma. Adrenal tumours were confirmed by hormonal assays, biochemical tests and computed tomography. To determine the site of the functioning adenoma, hormones were assayed from adrenal vein sampling. Thirty adrenal tumours were located on the right and 20 on the left. RESULTS: There were no significant differences in general demographic variables between the transperitoneal and retroperitoneal groups. The mean duration of surgery for transperitoneal and retroperitoneal laparoscopy was 202 and 221 min, respectively, and the mean blood loss 113 and 192 mL, respectively. The time to first oral intake, days to full diet, time until ambulatory and duration of hospitalization did not differ significantly between the two approaches. As the body mass index increased, the duration of surgery increased for both approaches. As more procedures were performed the duration of surgery decreased for both approaches. CONCLUSIONS: There were no significant differences between the transperitoneal and retroperitoneal approaches for laparoscopic adrenalectomy. PMID- 12133054 TI - Validity and reliability of an interviewer-administered questionnaire to measure the severity of lower urinary tract symptoms of storage abnormality: the Leicester Urinary Symptom Questionnaire. AB - OBJECTIVE: To develop a valid and reliable interviewer-administered questionnaire to measure the presence and severity of storage abnormality symptoms of incontinence, urgency, frequency and nocturia. SUBJECTS AND METHODS: Subjects were 930 men and women aged >/=40 years, taking part in a randomized controlled trial of a continence nurse practitioner (CNP) service. Criterion validity was tested by comparing questionnaire responses to 24-h pad test and 3-day urinary diary. Responsiveness was assessed by comparing questionnaire responses before and after treatment. Questions about urgency were investigated for construct validity in patients taking part in the trial who underwent urodynamic investigation (243). Test-retest and inter-rater reliability was measured at approximately 6 days in subjects recruited to an associated epidemiological study (104 and 102, respectively). RESULTS: The questionnaire responses showed significant associations with pad-test and diary measures. Questions about the severity of daytime incontinence performed better than those measuring night-time incontinence. The response categories of soaked, wet, damp and almost dry had better associations with the pad test than other measures of the severity of incontinence. Test-retest and inter-rater reliability was good for all questions, and all were responsive to change in symptoms, showing significant differences before and after treatment. CONCLUSION: There is a clear need for standardization of measurement using well-validated instruments. This interviewer-administered questionnaire is valid, reliable and sensitive to change in a wide range of severity of symptoms, and in both men and women aged >/=40 years. The questionnaire provides a useful assessment tool for primary and secondary care in research and clinical settings. PMID- 12133056 TI - The use of porcine dermal implant in a minimally invasive pubovaginal sling procedure for genuine stress incontinence. AB - OBJECTIVE: To assess the outcome of a suburethral sling using a porcine dermal implant (Pelvicoltrade mark, Bard Urology, UK) in the surgical management of urinary stress incontinence. PATIENTS AND METHODS: Forty women with urodynamically confirmed genuine stress incontinence were recruited into the study and followed up at 6 weeks and at least 6 months (mean 12 months, range 6 18). The sling was inserted using a minimal-access technique, which allowed 23 women to be operated as day-cases. Outcome measures included continence rates, voiding dysfunction, satisfaction scores and whether the patients would recommend the operation to a friend or relative. RESULTS: The cure rate was 85%, with sustained benefit; a further 10% of the women were improved by surgery. Voiding dysfunction rates were low and satisfaction scores high. Most women would undergo the procedure again if they became incontinent in the future and would recommend the procedure to a friend or relative. CONCLUSION: A minimal access pubovaginal sling using Pelvicoltrade mark is effective in treating stress incontinence. The complication rate is low and the procedure can be performed as a day-case with no loss of efficacy. PMID- 12133055 TI - Thermo-expandable intraprostatic stents in bladder outlet obstruction: an 8-year study. AB - OBJECTIVE: To assess the use of a thermo-expandable intraprostatic stent (Memokath(R), Engineers and Doctors A/S, Copenhagen, Denmark) for bladder outlet obstruction in men unable to undergo transurethral resection of the prostate (TURP), assessing symptoms, complications and duration of stent life. PATIENTS AND METHODS: The Memokath stent is a coil of a nickel-titanium alloy which has 'shape memory', the lower end expanding when heated to 55 degrees C. Risks associated with inserting the stent with a flexible cystoscope under local anaesthesia are minimal. Men were selected who were either permanently or temporarily unfit for TURP. Indications included severe respiratory and cardiovascular disease. Exclusion criteria included bladder carcinoma, calculi or detrusor failure; in all, 211 men were fitted with 217 intraprostatic stents over 8 years. RESULTS: There were 1511 TURPs during the study period; the mean age of men receiving a stent was 80.2 years, compared with 70.2 years for those undergoing TURP. The International Prostate Symptom Score decreased from a mean of 20.3 to 8.2 (P < 0.001) in the first 3 months after stent placement; there was virtually no change over 7 years. During the follow-up, 38% of men died with their stents in situ, 34% remain alive, 23% have had their stents removed for failure and 4% were removed as they were no longer required. There was a 13% migration rate and 16% repositioning rate. There were few side-effects (pain 3%, haematuria 3%, incontinence 6% and infection 6%). These frail men were more likely to die than have their stent fail. CONCLUSION: The Memokath intraprostatic stent is a valuable addition to the armamentarium of the urologist treating elderly or frail men with advanced bladder outlet obstruction and complements existing technologies. PMID- 12133057 TI - Does smoking status influence the prognosis of bladder cancer? A systematic review. AB - OBJECTIVE: To summarize, in a systematic review, the evidence for the effect of stopping smoking on recurrence, cancer-specific and all cause-mortality among smokers with newly diagnosed bladder cancer. MATERIALS AND METHODS: Two electronic databases and the reference lists of identified primary studies and reviews were searched. Studies were included if a hazard ratio and its confidence intervals could be extracted. A predefined set of study characteristics was extracted which defined whether studies were giving valid prognostic data on the effects of smoking in reasonably homogenous cohorts. The results of studies were synthesized qualitatively. RESULTS: Fifteen relevant studies were identified; former and current smokers were combined in many studies. Many studies produced information on prognosis that was confounded by the mixing of incident and prevalent cases. Only three studies examined the influence of smoking on prognosis in only incident cases, most of whom had superficial disease. Of these, only one was of high quality. These three studies and the other 12 showed suggestive evidence that continued smoking or a lifetime of smoking constitutes a moderate risk factor for recurrence and death, and that stopping smoking could favourably change this. However, the evidence base for this is weak because of the methodological shortcomings and because most studies' results were not statistically significant. A life-table model showed that if stopping smoking altered the prognosis, the size of the benefit would be clinically worthwhile. CONCLUSION: There is suggestive evidence that stopping smoking might favourably alter the course of bladder cancer, but this is insufficient for clinicians to inform patients that doing so will improve their prognosis, and for providing specialized services to assist in stopping smoking to patients with bladder cancer. PMID- 12133058 TI - A high easy-to-treat complication rate is the price for a continent stoma. AB - OBJECTIVE: To evaluate the conduit-related complications and their treatment in Mitrofanoff continent urinary diversion and antegrade colonic enema (ACE) procedures. PATIENTS AND METHODS: The files of 53 patients (18 men and 35 women) in whom 58 continent stomas were created were retrospectively reviewed. Gender, age, age at the time of surgery, underlying disease, concomitant surgery, abdominal position of the stoma, follow-up, complications and treatment were assessed. The mean (sd) age at the time of surgery was 19 (13) years; 30 patients were aged <15 years and the mean follow-up was 2.8 (1.9) years. Fifty-three continent vesicostomy-type Mitrofanoff stomas were constructed and five ACE procedures performed. Forty-five stomas were in the umbilicus and 13 on the abdominal wall. For 45 conduits the appendix was used; in the other 13 a transverse tubularized ileal segment according to Monti was created. RESULTS: There were stoma-related complications in 19 patients (36%), with 27 in all and stomal stenosis accounting for more than half. Five patients had urinary leakage. The median time to the first complication was 9 months. Multiple regression analysis showed that gender and stoma location were the only significant determinants of the complication/follow-up ratio. Women had more complications than men and umbilical stomas fared worse than those on the abdominal wall. Age, underlying disease and type of stoma were not significantly related to the complication/follow-up ratio. The complication was treated by one procedure in 13 patients; four needed two and two needed three surgical revisions. Most complications were relatively easy to treat, i.e. dilatation in five, endoscopic incision in one, re-anastomosis in four, Y-V plasty in seven, a new channel in two, reimplantation in three, a bladder cuff in two for stomal leakage, and abdominoplasty in two. Only one stoma had to be abandoned. CONCLUSION: The complication rate for continent small-diameter stoma is high. However, most complications are relatively easy to treat. Despite these complications, patient satisfaction remains high. PMID- 12133059 TI - How to master absorption during transurethral resection of the prostate: basic measures guided by the ethanol method. AB - OBJECTIVE: To evaluate the effect of basic measures to minimize the absorption of irrigating fluid during transurethral resection of the prostate (TURP) to the planned end-point, using the ethanol method to guide the surgeon. PATIENTS AND METHODS: Forty-six patients underwent TURP with intermittent irrigation using sterile water and 2% ethanol. The ethanol content in the expired breath of the patients was assessed every 5 min using a breath-alcohol monitor. In 25 patients no absorption was indicated but 21 showed some absorption according to the ethanol analyses. The operator was then immediately alerted, paused briefly to determine the route of absorption and to take advantage of blood clots to seal off leakage sites. On resuming the resection, attention was given to the pressure gradients in the operating field, based on bladder compliance and the flow in the irrigating jet. Repeated ethanol analyses were used to assess the result and to guide the surgeon's efforts. RESULTS: No operation had to be terminated prematurely; the mean absorption in the 21 patients in whom any ethanol was detected was 121 (75-230) mL. In 14 patients absorption occurred once, in six twice and in one there were three small absorption events. The group with absorption did not differ significantly from that with no absorption in operative duration (mean 48 and 47 min), resected weight (mean 32.8 and 31.6 g) or blood loss (mean 550 and 483 mL). CONCLUSION: If the surgeon is alerted at the first positive ethanol reading, effective measures to minimize absorption can be taken early. Regular use of the ethanol method as an alarm system should permit TURP with a minimum of absorption and avoid an early interruption or premature termination of surgery because of absorption. PMID- 12133060 TI - Technical characterization of an ultrasound source for noninvasive thermoablation by high-intensity focused ultrasound. AB - OBJECTIVE: To develop a generator for high-intensity focused ultrasound (HIFU, a method of delivering ultrasonic energy with resultant heat and tissue destruction to a tight focus at a selected depth within the body), designed for extracorporeal coupling to allow various parenchymal organs to be treated. MATERIAL AND METHODS: The ultrasound generated by a cylindrical piezo-ceramic element is focused at a depth of 10 cm using a parabolic reflector with a diameter of 10 cm. A diagnostic B-mode ultrasonographic transducer is integrated into the source to allow the focus to be located in the target area. The field distribution of the sound pressure was measured in degassed water using a needle hydrophone. An ultrasound-force balance was used to determine the acoustic power. These measurements allowed the spatially averaged sound intensity to be calculated. The morphology and extent of tissue necrosis induced by HIFU was examined on an ex-vivo kidney model. RESULTS: The two-dimensional field distribution resulted in an approximately ellipsoidal focus of 32 x 4 mm (- 6 dB). The spatially maximum averaged sound intensity was 8591 W/cm2 at an electrical power of 400 W. The lesion caused to the ex-vivo kidney at this maximum generator power with a pulse duration of 2 s was a clearly delineated ellipsoidal coagulation necrosis up to 8.8 x 2.3 mm (length x width) and with central liquefied necrosis of 7.9 x 1.9 mm. CONCLUSION: This newly developed ultrasound generator with a focal length of 10 cm can induce clear necrosis in parenchymal tissue. Because of its specific configuration and the available power range of the ultrasound generator, there is potential for therapeutic noninvasive ablation of tissue deep within a patient's body. PMID- 12133061 TI - A preliminary report on a patient-preference study to compare treatment options in early prostate cancer. AB - OBJECTIVE: To prospectively record prognostic factors, quality-of-life and outcome data in a patient-preference controlled study comparing radical prostatectomy with radical radiotherapy for the treatment of early prostate cancer. PATIENTS AND METHODS: All patients suitable for radical treatment of early prostate cancer were identified and provided with information from a urologist, oncologist and nurse to allow them to choose a treatment option. Prognostic and demographic data were recorded for all patients and patients followed up uniformly, with the additional collection of quality-of-life data. RESULTS: In a 38-month period, 196 patients were recruited to the trial; of these, 81 chose surgery, 81 radiotherapy, 30 brachytherapy and four 'watchful waiting'. The distribution of acknowledged prognostic factors was similar between the groups. CONCLUSION: If patient preference continues to divide this population into roughly equal and comparable arms, it should be possible to answer the underlying questions on the treatment of early prostate cancer using this study design. PMID- 12133062 TI - Is 'watchful waiting' a real choice for men with prostate cancer? A qualitative study. AB - OBJECTIVE: To understand what leads men to choose 'watchful waiting' rather than active treatment for cancer of the prostate. PATIENTS AND METHODS: Fifty men with confirmed prostate cancer in England, Wales and Scotland were interviewed about all aspects of their illness, for a Database of Individual Patients' Experience of illness. The sample included men at different stages of diagnosis and with experience of a wide range of treatments. We report here only what men said about their choice of treatment and the decision-making process. RESULTS: Watchful waiting would have been clinically inappropriate for almost half of the men (those with serious urinary symptoms and those with metastatic disease). However, few of the men who might have chosen watchful-waiting remembered this being presented as a serious option. Most in this group chose radical prostatectomy, radiotherapy, brachytherapy or cryosurgery. The few who chose watchful waiting had found doctors who supported their decision, had assessed the evidence from Internet sites, and were concerned about the side-effects and uncertain outcome of treatment. Men who chose watchful waiting, as well as those who opted for treatment, described considerable pressure from family members, doctors or support groups, to seek active treatment. CONCLUSION: This study helps to explain why some men will not contemplate watchful waiting, and why others may find it difficult to pursue that option. Understanding men's concerns may help clinicians to support men's treatment decisions. Treatment for prostate cancer is highly controversial because no randomized, controlled trials have shown whether or not active intervention increases survival. If trials are not completed it cannot be determined whether active treatments are the best course of action for men with prostate cancer. PMID- 12133063 TI - Seminal vesicle cysts and associated anomalies. PMID- 12133064 TI - Male genital tract injuries after contemporary inguinal hernia repair. PMID- 12133065 TI - Surgical sperm retrieval after previous vasectomy and failed reversal: clinical implications for in vitro fertilization. AB - OBJECTIVE: To investigate the effect of the interval between previous vasectomy reversal on retrieval rates of epididymal and testicular spermatozoa using percutaneous epididymal sperm aspiration (PESA), or testicular sperm extraction (TESE), and the subsequent reproductive potential of these gametes in intracytoplasmic sperm injection (ICSI) cycles. PATIENTS AND METHODS: Sixty-six consecutive sperm retrievals were considered in patients who were azoospermic after previous vasectomy, of whom 54 had had a previous failed reversal, the remainder deciding against a reversal. PESA and TESE retrieval rates were noted, as were the time since vasectomy and the interval between vasectomy and unsuccessful reversal. The presence of palpable epididymal cysts was noted, with their effect on sperm retrieval rates. Fertilization and pregnancy rates were analysed in subsequent ICSI cycles using freshly retrieved spermatozoa or frozen thawed cryopreserved spermatozoa. RESULTS: All 66 patients had sperm retrieved successfully; the success rates for PESA were not significantly affected by previous failed reversal when compared with patients who had not had a reversal, at 14 of 54 (26%) vs five of 12 (P=0.3). The interval since vasectomy did not affect PESA retrieval rates but there was a significantly poorer retrieval rate for PESA in the presence of palpable epididymal cysts, at seven of 35 (20%) vs 12 of 23 (52%) (P=0.012). Fertilization rates were significantly lower using cryopreserved spermatozoa retrieved from either the epididymis or testis (50% vs 70%, P=0.007), although subsequent implantation and pregnancy rates were not significantly different. CONCLUSION: Surgical sperm retrieval is successful in all cases of azoospermia secondary to vasectomy, either by PESA or TESE. There are no clinical markers to indicate which patients will have successful PESA after vasectomy, although the presence of epididymal cysts is associated with significantly lower retrieval rates. The reduction in fertilising ability of cryopreserved spermatozoa does not affect clinical pregnancy rates in ICSI cycles. PMID- 12133066 TI - The effect of perioperative distal vasal lavage on subsequent semen analysis after vasectomy: a prospective randomized controlled trial. AB - OBJECTIVE: To determine the effect of perioperative distal vasal lavage with 50 mL of normal saline on subsequent time to azoospermia after vasectomy. PATIENTS AND METHODS: Seventy-two patients were prospectively enrolled and randomized to undergo vasectomy with or without vasal lavage. Infertility rates at 8, 10 and 12 weeks were compared for both groups and for those undergoing the procedure under local or general anaesthesia. Patient compliance for returning postoperative semen for analysis was also assessed. RESULTS: There was no statistically significant difference in infertility rates at 8, 10 or 12 weeks after vasectomy with or without vasal lavage. Vasectomies performed under local and general anaesthesia had comparable rates of infertility at 12 weeks after surgery. Compliance in providing semen for analysis was poor. CONCLUSION: The routine adoption of distal vasal lavage during vasectomy for contraception cannot be recommended. As compliance in providing semen for analysis was poor, the clinician has a responsibility to remind the patient of the consequences of such action. PMID- 12133067 TI - Detrusor after-contractions in children with normal urinary tracts. AB - OBJECTIVE: To assess the clinical significance of after-contractions (A-Cs) in children with normal urinary tracts. PATIENTS AND METHODS: Urodynamic records obtained in 315 children with urinary infection or enuresis were reviewed retrospectively; 184 were selected for analysis of A-Cs. All patients had normal urinary tracts and none showed signs of an overt neuropathy. The urodynamic method comprised standard measurements of pressures and flowmetry (42 had video urodynamic studies). RESULTS: After-contractions occurred in 151 of the 184 patients; the incidence tended to decrease with age. The mean amplitude of the A Cs was 77.9 cmH2O; in 36% of the records it was higher than the voiding contraction. Residual urine was found in 12 of 151 records with A-Cs, but in only one patient was such residual urine confirmed in control voids. The patterns were assessed in 131 patients: in 36% they resembled stop-test responses, in 31% they were preceded by brief peaks of pressure or had jagged limbs, and in 33% they were grossly irregular. In 137 records the content of the bladder was estimated at the start of A-Cs; in 51% the bladder was empty or had evacuated >95% of its content, in 39% 95-80% and in 10% <80%. In only 7% of the patients had the A-Cs started after the voiding contraction had completely subsided. There was no difference in the incidence of A-Cs in girls with enuresis (84%) and girls with a history of urinary infections (85%). Detrusor instability was detected in 81% of the children with A-Cs and in 70% of those without; there was no correlation between the amplitudes of uninhibited detrusor contractions and of A-Cs. Characteristic images of external sphincter activity were found in only three of 14 video-urodynamic recordings with A-Cs. CONCLUSION: After-contractions are common in children with normal urinary tracts but they tend to disappear with age. In clinical urodynamics they are of limited practical use because their appearance is unpredictable and there are artefacts related to recording the final phase of micturition. The relationship with detrusor instability may be explained as a coincidence of two common but unrelated findings, and A-Cs are unrelated to urinary infection. External sphincter activity is not the only cause of A-Cs and when it occurs it does not alter the course of voiding, as it does in neuropathic dysfunctions. As their clinical significance is uncertain, treatment of A-Cs is not advocated. PMID- 12133068 TI - Increased urinary nitrite excretion in primary enuresis: effects of indomethacin treatment on urinary and serum osmolality and electrolytes, urinary volumes and nitrite excretion. AB - OBJECTIVE: To assess urinary nitrite excretion, a stable end product of nitric oxide (NO), in patients with enuresis and in normal controls, and to evaluate the effects of indomethacin (a potent prostaglandin synthesis inhibitor) on urinary nitrite excretion, other urinary variables and bladder capacity. PATIENTS AND METHODS: The study comprised 10 patients with primary enuresis and 10 normal comparable controls (age range 6-14 years). Nitrite was assayed in 'spot' morning urine samples in both the enuretics and normal controls. Enuretics were then given 50 mg indomethacin suppositories each night; urine volume, urinary osmolality and electrolytes, serum osmolality and electrolytes and urinary nitrite were assayed before indomethacin treatment and after 15 days of treatment. RESULTS: The mean (sd) urinary nitrite excretion was 24.4 (19.6) micromol/L in normal children and 275.9 (111.2) micromol/L in enuretics (P<0.05). With indomethacin, the urinary nitrite concentration was significantly decreased to 141 (45.1) micromol/L (P<0.05) and associated with a significant reduction in bed-wetting episodes and voiding frequency. The functional bladder capacity was <70% of the predicted value for age in six of the patients; they had significant improvements on indomethacin, to values similar to those in patients with a nearly normal functional bladder capacity. Indomethacin decreased the 24-h urinary volume by 41%, the night volume by 40%, clearance of free water by 46% and increased the day : night urinary volume ratio by 55%. The absolute amounts of urinary calcium, magnesium, phosphorus, urea, creatinine, and glucose were lower on indomethacin, although not statistically significantly so. Indomethacin decreased the 24-h urinary and 'spot' morning osmolality and osmotic clearance. There were no significant changes in serum osmolality and electrolyte concentrations. Indomethacin also decreased the absolute amount of urinary sodium, chloride and potassium, fractional sodium and potassium excretion, and filtered sodium. Creatinine clearance was decreased by 20% (P>0.05) and normal 24 h urinary protein was significantly lower, by 47%, after indomethacin treatment (P<0.05). CONCLUSION: Urinary nitrite excretion increased significantly in patients with primary nocturnal enuresis; indomethacin markedly reduced bed wetting episodes and decreased the frequency of voiding in enuretics with small or normal functional bladder capacity, which was associated with a significant decrease in urinary nitrite excretion. Indomethacin reduced bed-wetting by decreasing the urine volume, clearance of free water and urinary electrolytes, and through possible effects on bladder and urethral contraction, by inhibiting NO and prostaglandin synthesis. NO and prostaglandins might be important in the pathogenesis of primary enuresis. PMID- 12133069 TI - Reduction in nocturnal functional bladder capacity is a common factor in the pathogenesis of refractory nocturnal enuresis. AB - OBJECTIVE: To evaluate the diurnal and nocturnal bladder reservoir function in patients with refractory primary nocturnal enuresis (PNE). PATIENTS AND METHODS: Ninety-five children (68 boys, 27 girls, mean age 9.3 years) with significant PNE (>/=3 wet nights/week) that was refractory to treatment with desmopressin +/- an enuretic alarm were assessed using detailed recording of voiding frequency and urinary volume both day and night, natural filling cystometry during the day and continuous cystometry with simultaneous electroencephalogram monitoring during sleep at night. RESULTS: Patients could be broadly categorized into two groups. Group A comprised those with normal daytime urodynamics and functional bladder capacity (FBC) on detailed frequency-volume recording, but who developed marked detrusor instability associated with a significant reduction in nocturnal FBC and small-volume voiding only after sleep at night (33 patients, 35%); and group B, those with abnormal daytime urodynamics and with reduced FBC and small-volume voiding both day and night, but who somehow managed to mask their bladder symptoms during the day (62 patients, 65%). There was no evidence of nocturnal polyuria in either group and the ratios of day : night urinary output volumes for type A and type B patients were 1.48 and 1.99, respectively. CONCLUSION: A reduction in nocturnal FBC, either occurring only after sleep at night in association with the appearance of detrusor instability in patients with normal daytime urodynamics and FBC, or as a manifestation of occult voiding dysfunction or bladder outlet obstruction that affects the bladder reservoir function both day and night, appears to be a common factor and probably the main cause for a mismatch between nocturnal urine output and bladder storage capacity in patients with severe bed-wetting that was refractory to treatment. PMID- 12133070 TI - Bladder dysfunction as a prognostic factor in patients with posterior urethral valves. AB - OBJECTIVE: To investigate the extent to which different types of bladder dysfunction can affect long-term renal function in boys with posterior urethral valves (PUV). PATIENTS AND METHODS: Renal and bladder function were retrospectively assessed in 59 boys with PUV (mean age 10 years, range 5-17). All patients included in the study had at least 4 years of follow-up and their bladder behaviour had been evaluated in at least two urodynamic studies. At the time of the study, of the 59 PUV boys, 37 had normal renal function and 22 had end-stage renal disease (ESRD). RESULTS: Of the 59 boys with PUV evaluated by urodynamic studies, 25 had normally behaving bladders (42%) and 34 had some type of bladder dysfunction (58%). Of the 22 in ESRD, 15 had abnormally behaving bladders (68%) and only seven had bladders with normal behaviour (32%). Of the 37 boys with normal renal function, 19 had dysfunctional bladders (51%) and 18 had normal bladders (49%). Instability was found in 17 of 19 boys with bladder dysfunction and normal renal function. On the contrary among 22 boys with ESRD, poor compliance was the most frequent urodynamic pattern (eight, 53%) while instability was only found in five. Overall, eight of nine boys with poorly compliant bladders, two of three with myogenic failure and a five of 22 with instability were in ESRD, and this situation occurred at an earlier age in patients with poorly compliant bladders. CONCLUSION: Bladder dysfunction should be considered as a prognostic factor in renal failure. Those with poor bladder compliance and myogenic failure have the worst outcome, while bladder instability was associated with the lowest incidence of renal failure. PMID- 12133072 TI - Modulation of heat-induced cell death in PC-3 prostate cancer cells by the antioxidant inhibitor diethyldithiocarbamate. AB - OBJECTIVE: To examine the relationships between the form of cell death (apoptosis or necrosis), reactive oxygen species (ROS) generation, superoxide dismutase (SOD) activity and the level of heat-shock protein 70 (hsp 70) expression after thermotherapy of PC-3 prostate cancer cells; also assessed were the tumoricidal effects of combined treatment with both heat and the antioxidant inhibitor diethyldithiocarbamate (DDC). MATERIALS AND METHODS: PC-3 cells were treated with thermotherapy at 42, 43 or 44 degrees C for 30, 60, 90 or 120 min. Cell proliferation, ROS generation, SOD activity and cellular hsp 70 level were determined using tetrazolium-based cytotoxicity, fluorescent dichlorofluorescein (DCF) and nitroblue tetrazolium assays, Western blot analysis and flow cytometry, respectively. The apoptotic and necrotic cells were determined by staining with propidium iodide and fluorescein isothiocyanate-labelled annexin V. These variable were also measured after combined treatment of PC-3 cells with 1 mmol/L DDC and thermotherapy at 43 or 44 degrees C for 60 min. RESULTS: Cell survival was significantly lower after heating cells at 43 degrees C for 60, 90 and 120 min and at 44 degrees C for all periods tested (P<0.05). At 43 degrees C apoptosis increased with the duration of heating and was similarly enhanced after heating at 44 degrees C for 30 min. Necrosis was not increased by heating at 42 or 43 degrees C, but was markedly enhanced after heating at 44 degrees C with both the duration of heating and with time after heating. Significant increases in DCF production were induced by heating at 43 degrees C for 60, 90 and 120 min (P<0.05) and at 44 degrees C at all times (P<0.010-0.005). There was a significant correlation between the level of ROS generation and necrosis (P<0.001) but no correlation between the ROS level and apoptosis. SOD activity increased in cells after heating at 43 degrees C, with significant differences among cells heated for 60, 90 and 120 min (P<0.05). After heating at 44 degrees C, SOD activity was maximal in cells heated for 30 min (P<0.005), by 30 min and then decreased with time after heating. There were significant increases in hsp 70 level in cells heated at 43 degrees C for 90 and 120 min (P<0.05) and at 44 degrees C for 30 and 60 min (P<0.05 and <0.025, respectively). Hsp 70 levels decreased after heating at 44 degrees C for 90 and 120 min. The combination of DDC and heating significantly increased ROS generation and the percentage of cell death, and decreased SOD activity (P<0.05). CONCLUSION: These findings show a qualitative change in the form of cell death induced by thermotherapy of PC-3 cells, which changed from apoptosis to necrosis according to the degree and duration of heating. Mild thermotherapy induced marginally low occurrence of apoptosis of PC-3 cells and DDC may represent a useful future strategy for the treatment of prostate carcinoma. PMID- 12133071 TI - Intestinal and renal handling of oxalate in magnesium-deficient rats. Evaluation of intestinal in vivo 14C-oxalate perfusion. AB - OBJECTIVE: To clarify in vivo, using isolated small intestinal loops perfused with radioactive 14C-oxalate, whether intestinal hyperabsorption or reduced secretion is important in magnesium deficiency (MgD), as this is a potential cause of calcium oxalate urolithiasis. MATERIALS AND METHODS: Twenty-four Sprague Dawley rats were either fed a standard diet (control, 12 rats) or a magnesium deficient diet (MgD, 12 rats) for 19 weeks. One hour before the animals were killed, a defined length of a small intestinal loop was isolated and filled with 5 mL of 0.9% NaCl and a defined amount of intravenous 14C-oxalate applied. Using this method it was possible to determine the secretion of unlabelled oxalate into the intestinal lumen, from the specific activity in plasma. RESULTS: Plasma oxalate levels doubled under MgD; urinary calcium and phosphorus also increased significantly, while urine oxalate tended to decrease. The secretion of oxalate into the intestinal lumen of MgD animals increased significantly, by five times that of the control. The relative supersaturation for calcium oxalate remained constant. Elementary analysis of renal tissue showed an increase in calcium and phosphorus under MgD, in the sense of nephrocalcinosis, but no concretions were detected (no nephrolithiasis). CONCLUSION: In contrast to earlier studies, there is no evidence that hyperoxaluria is responsible for the possible development of urolithiasis in MgD. This was confirmed by calcium phosphate deposits in renal tissue, even though there was no evidence of oxalate urolithiasis. The increase in plasma oxalate seems to be completely compensated by strongly increased oxalate secretion into the intestinal lumen. PMID- 12133073 TI - Tunica albuginea acellular matrix graft for penile reconstruction in the rabbit: a model for treating Peyronie's disease. AB - OBJECTIVE: To evaluate the use of an acellular matrix graft of the tunica albuginea for functional penile reconstruction in severe cases of Peyronie's disease. MATERIALS AND METHODS: In 18 rabbits, an acellular matrix graft of the tunica albuginea was used to cover a 4 x 8 mm tunical defect, and six animals each were killed 1, 3 and 6 months later; four unoperated animals served as histological controls. Before death an erection was induced by papaverine, with the quality classified on a scale of 0-5, and cavernosography performed. After death the penis was prepared for histological study, and the cell number, collagen and elastic fibre content evaluated in the regenerated matrix, and in control specimens and four unimplanted matrices. RESULTS: Of 18 experimental animals, 11 had normal erections before death, four had slight penile deviation and three developed no erection. Failure was caused by severe postoperative haematoma, resulting in scar tissue. There was no graft rejection. Histologically there was no difference between natural and regenerated tunica. The collagen content and cell number were not significantly different in regenerated and control samples. There were significantly fewer elastic fibres in the unimplanted grafts and the 1-month group, but in later samples this difference was no longer evident. CONCLUSION: The homologous acellular matrix graft of the tunica albuginea warrants further evaluation as an alternative treatment in Peyronie's disease, despite some postoperative failures. The advantage of this orthotopic biomaterial is its rapid integration, with no rejection. PMID- 12133074 TI - DNA hypermethylation at the D17S5 locus is not a frequent event in human urothelial cancer. AB - OBJECTIVE: To analyse the DNA methylation status and the loss of heterozygosity (LOH) at the D17S5 locus (17p13.3) in urothelial cancer. MATERIALS AND METHODS: DNA methylation was assayed and LOH analysed by Southern blotting in a series of 33 transitional cell carcinomas of the bladder and renal pelvis. RESULTS: DNA hypermethylation and LOH at the D17S5 locus were detected in six (18%) and 17 (52%) of the tumours, respectively. The six cases with DNA hypermethylation were of the papillary type, and four also had LOH at this locus. CONCLUSION: In contrast to other epithelial tumours, DNA hypermethylation at the D17S5 locus is not a frequent event in human urothelial cancer. PMID- 12133075 TI - An oxalate-binding protein with crystal growth promoter activity from human kidney stone matrix. AB - OBJECTIVE: To fractionate renal-stone matrix proteins, identify the presence of oxalate-binding protein and assess its effect in a calcium oxalate (CaOx) crystal growth system. MATERIALS AND METHODS: Proteins were isolated from the matrix of kidney stones containing CaOx as the major constituent, using EDTA as a demineralizing agent. The solubilized proteins were subjected to cellulose-column chromatography by eluting with increasing sodium chloride concentrations in Tris HCl buffer. Three protein fraction peaks were eluted, i.e. fraction I in buffer, fraction II in 0.05 mol/L NaCl in buffer and fraction III in 0.3 mol/L NaCl in buffer. The protein fractions were tested for their effects on CaOx crystal growth. RESULTS: All three fractions had maximum CaOx binding activity at pH 7.4 but fraction II also had activity at pH 4.5. Fraction I promoted in vitro CaOx crystal growth, while fractions II and III were inhibitory. When fraction I was further separated on a Sephadex G-200 column, two protein fractions (Ia and Ib) were obtained. Fraction Ia protein had high and fraction Ib low CaOx-binding activity. Fraction Ia had a molecular weight of 48 kDa on gel electrophoresis and Western blotting. The 48 kDa protein did not cross-react with crystal matrix protein antibody, band-3 protein antibody, or albumin. The protein promoted CaOx crystal growth, with an optimum temperature of 37 degrees C and pH 6.5. The inhibitory effect of citrate on crystal growth was significantly lower in the presence of the 48 kDa protein. The protein promoted nucleation and aggregation of CaOx crystals in the in vitro crystallization system at pH 6.5, whereas fraction Ib (29 kDa) inhibited both nucleation and aggregation. Using the 48 kDa antibody, the yield of the protein from the stone matrix was 32% by EDTA extraction and only 3% with other methods. The protein was also detected in the nucleus and mitochondria, and in other matrix fractions of calcium phosphate and uric acid stones. CONCLUSION: The 48 kDa protein isolated from stone matrix is a potent promoter of CaOx crystal growth with high oxalate-binding activity; it is enriched in the nucleus and mitochondria. PMID- 12133076 TI - A new microsurgical reconstructive method for varicocele treatment. PMID- 12133077 TI - Finasteride-induced thrombocytopenia. PMID- 12133078 TI - Leuprorelin acetate granulomas: recurrent subcutaneous nodules mimicking metastatic deposits at injection sites. PMID- 12133082 TI - Academic anaesthesia -- alive and kicking? PMID- 12133083 TI - The effects of nitrous oxide on the auditory evoked potential index during sevoflurane anaesthesia. AB - We have studied the effects of nitrous oxide on the auditory evoked response index (AAI-index) derived from the A-line monitoring device during sevoflurane anaesthesia in 21 patients undergoing minor ambulatory surgery. During sevoflurane anaesthesia with an AAI-index < 30, the addition or withdrawal of nitrous oxide in a concentration of 66% end tidal did not show any linear dose dependent change in AAI-index . However, comparing nitrous oxide > 40% to nitrous oxide < 10% end tidal concentration the AAI-index did decrease, p < 0.05. The AAI index is either non-linear at deeper anaesthetic levels or is insensitive to the anaesthetic effects of nitrous oxide in terms of MAC-multiples. PMID- 12133084 TI - Effect of pre-operative anxiolysis on postoperative pain response in patients undergoing total abdominal hysterectomy. AB - In a double blind, placebo-controlled trial, we have assessed the effects of pre operative anxiolysis on postoperative pain scores in 112 ASA I-II women, aged 18 65 years, scheduled to undergo total abdominal hysterectomy. Subjects were randomly allocated to receive either oral diazepam 10 mg (n=56) or placebo (n=56) pre-operatively. Postoperative anxiety, pain scores, analgesic consumption, and sedation were evaluated at several time points during the first 24 h following surgery. Postoperative pain scores were found to be significantly higher in the diazepam group. Trait and state anxiety showed a significant effect on pain scores, independent of the treatment group. No difference was found between the groups in morphine consumption, but there was a significant reduction in morphine consumption with time. PMID- 12133085 TI - Neopterin as a marker of immunostimulation: an investigation in anaesthetic workplaces. AB - Personnel working in operating theatres and recovery rooms are exposed to a variety of noxious substances. The results of studies of the effects of occupational exposure on immune parameters are conflicting. Neopterin is an acknowledged marker of immunostimulation. Urinary neopterin levels of 58 anaesthetists and anaesthetic nurses were measured over a 3-week period. Neopterin analyses were performed using high performance liquid chromatography. Neopterin levels were within the normal range for all subjects. Younger subjects (aged < or = 35 years) had significantly higher urinary neopterin concentrations than older subjects (aged > 35 years). The present study is the first to investigate the influence of anaesthetic exposure on neopterin levels. No evidence of immunostimulation was found. PMID- 12133086 TI - Assessing critical care unit performance: a global measure using graphical analysis. AB - Outcome measurement in critical care is difficult because of the wide variety of patients treated and the diverse therapeutic options and pathways available. Individual outcome measures for critical care are available but are naturally limited to only a single aspect of performance. Most importantly, better performance in one aspect of care may compromise the standard of care in another. A global measure of performance would be helpful. For the year 1999-2000, the five hospitals in the East Anglian Critical Care Network provided data on capacity, workload and performance. The data was transformed and displayed graphically on a radar chart so that the area of the polygon within the radar chart was proportional to each unit's overall performance. The results from the five hospitals suggest that there is little overall difference in the units' global performance but the graphical representation highlighted some individual deficiencies. Graphical analysis of complex processes such as critical care delivery may facilitate performance assessment, providing that the measures chosen, weightings assigned and scales used are standardised with care. PMID- 12133087 TI - The effect of a pre-operative infusion of Ringer's solution on splanchnic perfusion in patients undergoing coronary artery bypass grafting. AB - Surgical patients develop a fluid deficit during pre-operative starvation. This study examines the effects of pre-operative fluid administration on haemodynamic variables, oxygenation and splanchnic perfusion in patients undergoing elective coronary artery bypass grafting. Forty-eight patients were randomised to receive either a pre-operative crystalloid infusion (crystalloid group, n = 24) or no infusion (control group, n = 24). Patients in the crystalloid group received a continuous infusion of Ringer's solution at 1.5 ml.kg(-1).h(-1) from 22:00 h until induction of anaesthesia the next morning. Immediately before induction of anaesthesia, all patients were given a colloid infusion to increase pulmonary capillary wedge pressure and central venous pressure to similar levels in both groups. Haemodynamic and oxygenation parameters were measured using invasive cardiovascular monitoring, and splanchnic perfusion was assessed by indocyanine green clearance. Patients in the crystalloid group received a mean (SD) of 1008 (140) ml of Ringer's solution overnight. Patients in the crystalloid group had a higher splanchnic blood flow than the control group before induction of anaesthesia [mean (SD) = 1782 (573) ml.min(-1) vs. 1391 (333) ml.min(-1), p < 0.05]. There were no significant differences in systemic haemodynamic data and global oxygenation parameters between the two groups. Pre-operative infusion of crystalloid appears to result in an improvement in pre-operative splanchnic perfusion. PMID- 12133089 TI - Clinical results with a new acoustic device to identify the epidural space. AB - Fifty patients scheduled for surgery under lumbar epidural anaesthesia were included in a study to evaluate the possibility of localising the epidural space solely by means of an acoustic signal. With an experimental set-up, the pressure generated during the epidural puncture procedure was translated into a corresponding acoustic signal. One anaesthetist held the epidural needle with both hands and detected the epidural space by means of this acoustic signal. At the same time, a second anaesthetist applied the loss of resistance technique and functioned as control. In all patients the epidural space was located with the acoustic signal. This was confirmed by conventional loss of resistance in 49 (98%) of the patients; in one patient (2%) it was not. We conclude that it is possible to locate the epidural space using an acoustic signal alone. PMID- 12133088 TI - Uncertainty and scoring systems. AB - Estimating risks for individual patients facilitates communication with patients, relatives and colleagues, and determines whether further treatment is futile. The process of estimating risks involves mathematics (i.e. scoring systems) and human experience and expertise. Understanding how risks are estimated is important because prognostication is an integral part of any medical specialty. In the USA, such treatment limitation or withdrawal decisions were made on only 7% of all intensive care unit patients but this represented 47% of all deaths on such units. In the UK, data reported by the Intensive Care National Audit and Research Centre suggest that although treatment limitation decisions are made on only 11.8% of patients, this accounts for over 50% of deaths on intensive care. Scoring systems offer a useful adjunct in identifying futility but there are important inherent weaknesses that limit their performance. This review aims to discuss some of these limitations. PMID- 12133090 TI - The effect of single use laryngoscopy equipment on illumination for tracheal intubation. AB - We measured the illumination generated by all 30 Macintosh size 3 laryngoscopes in our department with a lux-meter and a standardised laryngoscope holding tube. We found a large range in illumination generated (65-3130 lx). We then measured the effect on the illumination for each laryngoscope by covering the blade with a cover (LaryGard). In every case, the illumination was reduced by the LaryGard, the mean (SD) reduction was 67 (19)%. When we compared the illumination generated by a disposable laryngoscope blade with the same power source, we found that the illumination was reduced less than with the standard Macintosh covered by a LaryGard. PMID- 12133091 TI - Provision of postoperative epidural services in NHS hospitals. AB - To describe facilities for postoperative epidurals in UK National Health Service Hospitals, a questionnaire was sent to each hospital performing surgery below the head and neck. Of 271 hospitals, 256 replied (95%). While almost all offer postoperative epidurals, only 78 (30%) offer them to all surgical disciplines. Most hospitals rely on acute pain nurses for troubleshooting during the day, and on trainee anaesthetists after hours. Administration is most commonly by continuous infusion. There was no restriction on the use of epidural opioids in 67% of hospitals. Most (96%) hospitals have a protocol for epidural care, although the specified level of monitoring varies widely. There is no consensus of practice on removal of epidural catheters relative to anticoagulation. Levels of training in epidural care also vary widely. Two hundred and thirty-six hospitals (92%) have an acute pain team. Despite the expansion in acute pain services, facilities for postoperative epidurals are deficient in many NHS hospitals. PMID- 12133092 TI - The effects of two methods for customising the original SAPS II model for intensive care patients from South England. AB - Model customisation is used to adjust prognostic models by re-calibrating them to obtain more reliable mortality estimates. We used two methods for customising the Simplified Acute Physiology Score II model for 15,511 intensive care patients by altering the logit and the coefficients of the original equation. Both methods significantly improved model calibration, but customising the coefficients was slightly more effective. The Hosmer-Lemeshow chi(2)-value improved from 306.0 (p< 0.001) before, to 14.5 (p < 0.07) and 23.3 (p < 0.06) after customisation of the coefficients and the logit, respectively. Discrimination was not affected. The standardised mortality ratio for the entire population declined from 1.16 (95% confidence interval: 1.13-1.20, p < 0.001) to 0.99 (95% confidence interval: 0.96 1.02, p < 0.22) after customisation of the coefficients. The uniformity-of-fit for patients grouped by operative status and comorbidities also improved, but remained imperfect for patients stratified by location before intensive care unit admission. Amalgamation of large, regional databases could provide the basis for the re-calibration of standard prognostic models, which could then be used as a national reference system to allow more reliable comparisons of the efficacy and quality of care based on severity adjusted outcome measures. PMID- 12133093 TI - Standardized colour coding for syringe drug labels: a national survey. AB - A standardised colour code for user-applied syringe labels for anaesthetic drugs exists in the USA, Australia, New Zealand and Canada. In the UK, there is none. Consequently, an assortment of colour codes for syringe labels is available in the UK. We conducted a postal survey of the 285 College Tutors of the Royal College of Anaesthetists to establish their local syringe drug labelling system and their views on a national, standardised colour code. We found that that 96% of departments currently use coloured syringe drug labels. Of these, 98% use the 'Medilabel' scheme. The College Tutors felt that a standardised colour code for labels is required (94%), that the Association of Anaesthetists or the Royal College of Anaesthetists should be involved in the choice of scheme (76%) and that the scheme chosen should be international (65%). There was a majority feeling that the opinions expressed were representative of other members of the College Tutors' departments. We conclude that a national standard for drug labels is required and that a choice will have to be made between the 'international' scheme and the currently dominant Medilabel scheme. PMID- 12133094 TI - A comparison of oral transmucosal fentanyl and oral midazolam for premedication in children. AB - Oral transmucosal fentanyl citrate (OTF) was compared with midazolam as a premedicant in a prospective, randomised, placebo-controlled, double-blind trial. Eighty children (ASA grade 1 or 2, aged 3-9 years) who presented for tonsillectomy were randomly allocated to receive either 2.5 ml OTF (15-20 microg.kg(-1)) in a lollipop format and 0.5 ml.kg(-1) placebo syrup, or midazolam syrup (0.5 mg.kg(-1)) and a placebo lollipop (2.5 ml). The acceptability of sedation, anxiety and compliance with anaesthetic induction were assessed. The children were given an 'emergence' score for their recovery. Analgesia requirements, the incidence of vomiting, itching and any behavioural changes were assessed for 6 h postoperatively. Oral transmucosal fentanyl citrate was as effective as midazolam in aiding compliance with anaesthesia, but is significantly better in its appeal to children (p < 0.001) and emergence (p < 0.001) characteristics. In conclusion, OTF may be particularly useful as a premedicant in paediatric practice. PMID- 12133095 TI - A multi centre telephone survey of compliance with postoperative instructions. AB - Patients undergoing procedures under general anaesthesia as day cases are routinely given a set of instructions regarding activities to avoid in the first 24 h after discharge. Day surgery units generally specify the need for a responsible carer from time of discharge for a period of 24 h. This study looks at the compliance of 240 patients with postoperative instructions. Of the patients studied, 4.1% drove, 1.7% made important decisions, 3.3% drank alcohol, 0.8% took sedatives and 10% cooked, ironed or looked after children. All patients were discharged into the care of a responsible adult. However, 13.3% failed to have a carer with them for 24 h and 1.3% spent the night alone at home. Of our cohort, 25% were unable to comply with the postoperative instructions in full. The majority of non-compliance occurred on the day following surgery, suggesting that patients may feel that the advice is excessively cautious. PMID- 12133096 TI - Magnesium as first line therapy in the management of tetanus: a prospective study of 40 patients. AB - A prospective observational study was conducted to examine the efficacy and safety of magnesium sulphate for control of spasms and autonomic dysfunction in 40 patients with tetanus. Magnesium was infused intravenously, aiming to control spasms despite suppression of patellar reflex or respiratory insufficiency. Spasms were controlled in 38 of the 40 patients within a serum Mg(2+) range of 2 4 mmol.l(-1) with only two patients needing additional neuromuscular blocking drugs. Seventeen of 24 patients (< 60 years) and six of 16 patients (> or = 60 years) did not require ventilatory support. Thirty-six patients were conscious and co-operative throughout their management. Sympathetic over-activity was controlled without supplementary sedation. Overall mortality was 12%; all five deaths were in patients > or = 60 years and no deaths were due to autonomic dysfunction. We recommend magnesium as possible first line therapy in the routine management of tetanus. PMID- 12133098 TI - Arterial line insertion. PMID- 12133099 TI - The use of the laryngeal mask airway in maxillofacial surgery. PMID- 12133100 TI - The introduction of cricoid pressure. PMID- 12133101 TI - Disposable laryngoscope blades. PMID- 12133102 TI - Metal plate for placement of the laryngeal mask in a patient with orthodontic appliances. PMID- 12133104 TI - Midazolam -- an anti-emetic? PMID- 12133103 TI - An important differential diagnosis of pneumothorax. PMID- 12133105 TI - Fat embolism. PMID- 12133106 TI - A case of aortocaval fistula -- biochemical and haemodynamic changes. PMID- 12133107 TI - Pump activated by a foot switch pedal for controlled administration of local anaesthetic drugs. PMID- 12133111 TI - Methamphetamine: drug use and psychoses becomes a major public health issue in the Asia Pacific region. PMID- 12133112 TI - Effects of concurrent use of alcohol and cocaine. AB - The combination of alcohol and cocaine is popular among drug users, perhaps because of more intense feelings of 'high' beyond that perceived with either drug alone, less intense feelings of alcohol-induced inebriation and tempering of discomfort when coming down from a cocaine 'high'. A review is presented of the medical literature on psychological and somatic effects and consequences of combined use of alcohol and cocaine in man. The search was carried out with Medline, the Science Citation Index/Web of Science and Toxline. Exclusion and inclusion criteria for this search are identified. There is generally no evidence that the combination of the two drugs does more than enhance additively the already strong tendency of each drug to induce a variety of physical and psychological disorders. A few exceptions must be noted. Cocaine consistently antagonizes the learning deficits, psychomotor performance deficits and driving deficits induced by alcohol. The combination of alcohol and cocaine tends to have greater-than-additive effects on heart rate, concomitant with up to 30% increased blood cocaine levels. Both prospective and retrospective data further reveal that co-use leads to the formation of cocaethylene, which may potentiate the cardiotoxic effects of cocaine or alcohol alone. More importantly, retrospective data suggest that the combination can potentiate the tendency towards violent thoughts and threats, which may lead to an increase of violent behaviours. PMID- 12133113 TI - Would smokers with schizophrenia benefit from a more flexible approach to smoking treatment? AB - We evaluated literature that addresses the notion that flexible smoking treatment approaches are warranted for smokers with a diagnosis of schizophrenia. Understanding the biological and psychological mechanisms that increase the likelihood of smoking and decrease the motivation to quit for these individuals is addressed within the framework of a neurobiological model. We provide a brief overview of the limited smoking cessation treatment literature for patients with schizophrenia and compare abstinence-focused versus reduction-focused treatment modalities. The potential utility of the reduction-focused approach to tobacco treatment for these smokers is evaluated. Suggestions for future research to address the utility and efficacy of reduction-focused interventions for smokers with schizophrenia are put forth. We conclude with a consideration of the implications for the current understanding of smoking treatment among patients with co-morbid psychiatric diagnoses. PMID- 12133116 TI - Rigidity in measures of smoking cessation. PMID- 12133118 TI - Prevalence of and risk factors for methamphetamine use in northern Thai youth: results of an audio-computer-assisted self-interviewing survey with urine testing. AB - AIMS: Data from drug treatment facilities, drug seizures and drug arrests suggest rapidly increasing methamphetamine use by adolescents in Thailand. However, limited quantitative data are available about the prevalence of its use or correlates of use. The purpose of our study was therefore to estimate the prevalence of methamphetamine use and to identify possible risk factors. DESIGN: Cross-sectional survey using anonymous audio-computer-assisted self-interview and urine specimen analysis. SETTING: Chiang Rai Province, Thailand. PARTICIPANTS: 1725 students, 15-21 years of age (893 male and 832 female) attending one of three vocational schools in Chiang Rai Province. FINDINGS: Three hundred and fifty male and 150 female students reported a history of having ever used methamphetamine. In addition, 128 male and 49 female students had positive urine test results, indicating recent methamphetamine use; 27 of these students denied having ever used methamphetamine. According to history, urine test, or both, 41.3% of male students and 19.0% of female students used methamphetamine. In multivariate analysis, methamphetamine use was highly correlated with the use of other substances, sexual activity, peer pressure, positive attitudes toward methamphetamine, and absence of a family confidant. CONCLUSIONS: Methamphetamine use is common among adolescent students in northern Thailand. Demographic, behavioral and psychosocial correlates of methamphetamine use identified in this study may be helpful for the design and implementation of preventive interventions. PMID- 12133119 TI - Methamphetamine in Japan: the consequences of methamphetamine abuse as a function of route of administration. AB - AIMS: To determine differences in life backgrounds and clinical features between methamphetamine (MAP) smokers and injectors in Japan. SETTING: Out-patient clinic at a psychiatric centre. PARTICIPANTS: Among 451 MAP abusers undergoing initial assessments, 116 subjects whom the first author had directly interviewed and treated were studied. DESIGN AND PROCEDURES: In this study, life backgrounds, clinical features and psychiatric symptoms were compared between three subgroups: 42 (36.2%) in group S (smoking only); 57 (49.1%) in group I (injection only); and 17 (14.7%) in group SI (initially smoking, later injecting). FINDINGS: Group I more often had parental absence (P < 0.001), a family history of alcoholism (P < 0.05), limited education (P < 0.001), or a criminal record (P < 0.001) than patients in the other two groups. Group S had the most cannabis use (P < 0.01), while group I had the most volatile solvents use (P < 0.01). Group S experienced their first psychotic episode sooner after first MAP use (P < 0.01), but showed fewer auditory hallucinations at initial assessment than patients in other groups (P < 0.001). Group SI was intermediate between groups S and I in life background, clinical features and psychotic symptoms, while they had lost control of their drug use most frequently (P < 0.02). CONCLUSIONS: In Japan, MAP smokers have different life backgrounds from injectors. Smoking MAP does not appear to be a safer route as regards losing control of MAP use and inducing psychosis than injection. PMID- 12133120 TI - Nimodipine in opiate detoxification: a controlled trial. AB - OBJECTIVE: To evaluate the use of L-type calcium channel blockers (CaCB) in out patient opiate detoxification. DESIGN: Controlled trial with sequential allocation of patients to groups. METHODS: Three groups of individuals subject to opiate detoxification were involved: (1) the experimental group (n=30) received a course of nimodipine and dextropropoxiphen; (2) one control group (n=20) was detoxified with a course of dextropropoxiphen and benzodiazepine; and (3) a second control group (n=30) was treated with a standard course of alpha-2 adrenergic agents and naltrexone. In all cases, the detoxification course was scheduled to last 7 days. RESULTS: All the groups showed a significant opiate withdrawal syndrome (OWS) during detoxification (follow-up effect: Lambda=0.04; F6.52=201.89; P < 0.001), but from the first day the group treated with CaCB manifested fewer symptoms than the control groups (treatment effect: F2.57=97.99; P < 0.001). From the start, the intensity of the OWS was reduced by half in the CaCB group (M=6.67) compared with that manifested by the two other groups (M approximately 13). The clinical impression of the evolution of the detoxification was that it was comfortable and free of complications (significant side-effects were not observed). CONCLUSIONS: The results of the study suggest that the use of calcium channel blockers (CaCB) may be an effective method in opiate detoxification. Full randomized trials are warranted. PMID- 12133121 TI - Differences in factors associated with first treatment entry and treatment re entry among cocaine users. AB - AIMS: To investigate factors associated with first entry to treatment and with treatment re-entry among cocaine users. DESIGN: Cross-sectional study. SETTING, PARTICIPANTS: Cocaine users (n=313) recruited from community and treatment settings in Brazil. MEASUREMENTS: Structured questionnaire including selected items from the addiction severity index (ASI), general health questionnaire, version 28 (GHQ-28), CAGE and the severity of dependence scale (SDS). FINDINGS: Higher dose use, being a problematic drinker and increased awareness of their problem were associated with increased odds of making first contact with an agency. Greater severity of dependence, being involved in acquisitive crime and social support increased the chance of treatment re-entry. Being involved in acquisitive crimes and concerns about confidentiality were associated with decreased odds of first treatment contact. Being a problematic drinker was associated with decreased odds of re-entry treatment. CONCLUSIONS: These findings suggest that the distinction between first treatment contact and subsequent entry to treatment is useful, clinically relevant and deserving of further investigation. PMID- 12133122 TI - Childhood and adolescent antecedents of substance use in adulthood. AB - AIMS: To examine childhood antecedents of marijuana and cocaine use in adulthood. DESIGN: Epidemiological, longitudinal cohort study of African American first graders (age 6) followed to age 32. PARTICIPANTS: Children (N=1242) and families in the 57 first grade classrooms from Woodlawn, an inner-city community in Chicago. First grade teachers, mothers and children provided assessments over the life course. During adulthood, 952 participants were re-interviewed. MEASUREMENTS: First grade teacher behavior ratings, readiness for school tests, self-reports of adolescent drug use, social bonds and adult self-reports of drug use were the primary variables. FINDINGS: Males who were both shy and aggressive in first grade were more likely to be adult drug users compared to those who were neither. Shy females in first grade were less likely to be adult marijuana users than non-shy females. Adolescent social bonds did not moderate the relationships of earlier childhood behavior to adult drug use. Males who had a 'high/superior' readiness to learn scores in first grade were less likely to be cocaine users as adults, even though in earlier work we showed that they were more likely to initiate adolescent drug use. Females scoring as poor performers in first grade were less likely to ever use cocaine compared to females with higher scores. CONCLUSIONS: The combination of shy and aggressive behavior is an important antecedent for later male drug use and may help distinguish those who will be persistent users in adulthood from those who experiment in adolescence. PMID- 12133123 TI - Over a decade of syringe exchange: results from 1997 UK survey. AB - AIMS: To describe syringe exchange provision in the United Kingdom. DESIGN: Two phase cross-sectional survey: phase I, establishing a sampling frame of syringe exchange coordinators (n=420); phase II, surveying the coordinators seeking data on the number of syringe exchange outlets, visits and syringes distributed during April 1997 (68% response rate). SETTING: United Kingdom. FINDINGS: In 1997, nearly all Health Authorities in the United Kingdom (96%) operated some form of syringe exchange service, except Northern Ireland. In April 1997, 1 707 000 syringes were reported as being distributed. Assuming that non-responders coordinated the median number of outlets and distributed the median number of syringes as responders, we estimate that 27 million syringes were distributed annually from over 2000 outlets in the United Kingdom. The number distributed in Scotland was 3-4 times less than in England when measured as a number per adult (15-44), drug user in treatment, or estimated injecting drug user. CONCLUSIONS: Overall, there has been a 6.5-fold increase in syringe distribution in England since 1991. The number of syringes distributed in the United Kingdom may be higher than the United States. However, there appears to be unequal distribution of syringes within the United Kingdom, which may be associated with higher levels of HCV among injectors in Scotland compared to England. PMID- 12133124 TI - Treatment of out-patients with complicated benzodiazepine dependence: comparison of two approaches. AB - AIMS: To evaluate whether gradual benzodiazepine taper combined with cognitive behavioural treatment is more effective than standard treatment for patients with dependence in out-patient clinics. DESIGN: A randomized, controlled clinical trial, using standard questionnaires and serum and urine tests. SETTINGS: Four public-sector out-patient clinics for alcohol and drug abusers in Helsinki. PARTICIPANTS: Seventy-six patients with benzodiazepine dependence (DSM-III-R). Patients taking high doses of the drug or with alcohol use disorders were included to obtain a subject group representative of usual clinical practice. INTERVENTION: Subjects received gradual benzodiazepine taper combined with cognitive-behavioural therapy (experimental group) or standard withdrawal treatment not scheduled by the researchers (control group). MEASUREMENTS: The outcome was measured in terms of attaining a state of abstinence or by a decrease in the dosage during the study period of up to 12 months' duration. FINDINGS: No statistically significant differences in the outcomes were observed between the groups. A total of 13% of the experimental group and 27% of the control group were able to discontinue drug use. In addition 67% of the experimental group and 57% of the control group were able to decrease the dose. CONCLUSIONS: The search continues for improved methods of helping patients with complicated benzodiazepine dependence. PMID- 12133125 TI - A population-based study of cigarette smoking among illicit drug users in the United States. AB - AIMS: People who use illicit drugs are thought to have high rates of cigarette smoking; however, few population-based studies have been reported. We describe smoking patterns among illicit drug users, assess whether cigarette smoking is more prevalent among illicit drug users than it is among non-users and explore how smoking relates to level and type of drug use. DESIGN, SETTING, PARTICIPANTS: We used adult responses to the 1997 National Household Survey on Drug Abuse (n = 16 661). Multivariate analyses used SUDAAN to adjust standard errors for the sampling design and controlled for age, race, sex, education, depression, treatment history and alcohol. MEASUREMENTS: Smoking rates, cessation rates and smoking levels. FINDINGS: Seventy-one per cent of recent illicit drug users smoked cigarettes at least once in the past month. Their adjusted odds of being a smoker were much greater than for the general population (OR = 3.07, P < 0.0001). Their quit rate, although substantial, was half that of non-users (23% versus 56%, P=0.0001). Odds of being a smoker were higher for poly- versus monodrug users (OR = 2.35, P=0.0020) and rose with increased drug use (OR = 1.36, P=0.0374). Illicit drug users who perceived smoking to be risky were four times less likely to smoke (OR = 0.23, P=0.0008). CONCLUSIONS: Although most recent illicit drug users smoke, some are able to quit. Better understanding of concurrent cigarette and illicit drug use may provide impetus for policy change and shed light on underlying mechanisms of addiction. Clinicians, policy makers and user advocates should address tobacco use in drug treatment and in harm reduction interventions. PMID- 12133126 TI - Patterns of smoking in Estonia. AB - AIMS: To describe the pattern of current smoking and its relation to socio demographic factors in Estonia. DESIGN: Nationwide cross-sectional survey. SETTING: Estonia in 1996. PARTICIPANTS: Stratified random sample of 2086 adults aged 30-59. MEASUREMENTS: Prevalence of current smoking; socio-demographic factors related to smoking, investigated by logistic regression analysis. FINDINGS: The prevalence of current smoking was 57.9% among men and 25.7% among women. For both genders, smoking rates were consistently lowest in the age group 50-59 years and highest in the age group 30-39 years. Smoking was significantly more common among divorced and widowed people. Education was associated with smoking among men but not among women. No relationship, however, was established between smoking and ethnicity, type of residence, and household income. CONCLUSIONS: Estonia needs an effective antismoking policy. Public health efforts need to be focused on quitting smoking in younger adults and prevention efforts should target less educated socio-economic groups. PMID- 12133127 TI - Gender differences in help-utilization and the 8-year course of alcohol abuse. AB - AIMS: The aim of this study was to compare initially untreated women and men problem drinkers on help-utilization and outcomes over 8 years. DESIGN AND PARTICIPANTS: At the time of the 8-year follow-up, individuals (N=466, 49% female) had self-selected into four groups: no help, Alcoholics Anonymous (AA) only, formal treatment only or formal treatment plus AA. MEASUREMENTS: At baseline and 1, 3 and 8 years later, participants completed measures of drinking and functioning. FINDINGS: Women were generally worse off than men on baseline drinking and functioning indices. In keeping with their poorer baseline status, women were more likely to participate in AA, and had longer in-patient stays during the next year. When women's baseline status was controlled, women had better outcomes than did men at 1 and 8 years. Generally, women and men did not differ on the extent to which obtaining help, or a particular type of help, was related to improved outcomes. Regarding drinking outcomes, women benefited more than did men from more AA attendance during years 2-8 of follow-up. CONCLUSIONS: The results suggest that although alcoholism interventions were designed primarily for men, they are currently delivered in ways that are also useful to women. Problem-drinking women appear to benefit from sustained participation in AA, which emphasizes bonding with supportive peers to maintain abstinence. PMID- 12133128 TI - Social networks as mediators of the effect of Alcoholics Anonymous. AB - AIMS: This study tested the hypothesis that the relationship between Alcoholics Anonymous (AA) involvement and reduced substance use is partially explained (or 'mediated') by changes in social networks. DESIGN: This is a naturalistic longitudinal study of the course of alcohol problems. SETTING: Study sites were the 10 largest public and private alcohol treatment programs in a northern California county. PARTICIPANTS: Three hundred and seventy-seven men and 277 women were recruited upon seeking treatment at study sites. MEASUREMENTS: At baseline and 1-year follow-up, we assessed alcohol consequences and dependence symptoms, consumption, social support for abstinence, pro-drinking social influences and AA involvement. FINDINGS: In the structural equation model, AA involvement was a significant predictor of lower alcohol consumption and fewer related problems. The size of this effect decreased by 36% when network size and support for drinking were included as mediators. In logistic regression models predicting abstinence at follow-up, AA remained highly significant after including social network variables but was again reduced in magnitude. Thirty-day abstinence was predicted by AA involvement (OR=2.9), not having pro-drinking influences in one's network (OR=0.7) and having support for reducing consumption from people met in AA (versus no support; OR=3.4). In contrast, having support from non-AA members was not a significant predictor of abstinence. For alcohol related outcomes other than abstinence, significant relationships were found for both AA-based and non-AA-based support. CONCLUSIONS: The type of social support specifically given by AA members, such as 24-hour availability, role modeling and experientially based advice for staying sober, may help to explain AA's mechanism of action. Results highlight the value of focusing on outcomes reflective of AA's goals (such as abstinence) when studying how AA works. PMID- 12133129 TI - Too drunk for a beer? A study of overserving in Stockholm. AB - AIMS: To evaluate the effects of a community alcohol prevention programme on the frequency of alcohol service to intoxicated patrons at licensed premises. DESIGN: Pretest (1996)-post-test (1999) design. SETTING: Licensed premises in Stockholm, Sweden. INTERVENTION: The community alcohol prevention programme, including server training in responsible beverage service (RBS) and policy initiatives in the community, has been conducted since 1996. PARTICIPANTS AND MEASUREMENTS: Actors were hired to enter licensed premises, enact a scene of severe intoxication and attempt to order a beer. At the baseline in 1996, actors visited 92 licensed premises, 47 from the central part of Stockholm and 45 from the southern part of Stockholm. At the follow-up in 1999, 103 licensed premises were visited, 61 from the central part of Stockholm and 42 from the southern part of Stockholm. Observers monitored each visit. FINDINGS: At follow-up the actors were denied service of alcohol at 47% of the licensed premises, a statistically significant improvement compared to 5% in the baseline study. CONCLUSIONS: Licensed premises refused service of alcohol to intoxicated patrons to a much greater extent than in the baseline study. The improved results can probably be explained by a combination of policy initiatives in the community, changes in the overall enforcement environment and RBS training. PMID- 12133130 TI - Blood alcohol content (BAC)-negative victims in alcohol-involved injury incidents. AB - AIMS: To understand better how often BAC-negative victims suffer fatal and non fatal alcohol-involved injuries. DESIGN: We analyzed 1988-1993 data from Oklahoma surveillance systems that track all people killed or hospitalized due to burns, submersions and spinal cord injuries (SCIs) and investigate alcohol use by victims and any others who contributed to their injuries. MEASUREMENTS: Percentage of victims BAC-positive and percentage of victims BAC-negative in alcohol-involved incidents. FINDINGS: Of 5107 cases studied, alcohol involvement was known for 4773 (93%), 1882 fire burns (93%), 1560 scald and other burns (97%), 698 submersions (92%) and 967 spinal cord injuries (89%). By type of injuries, BAC-negative victims were 4%, 13%, 6% and 8% of alcohol-involved cases, respectively. CONCLUSION: A sizeable number of BAC-negative victims suffer alcohol-involved injuries. Although alcohol is not the cause of some of these injuries, alcohol policy changes probably could avert others. PMID- 12133131 TI - Implications for the notion that moderate alcohol use protects from ischaemic heart disease: comment on Greenfield et al. PMID- 12133132 TI - Cannabis, carrots and common experiences: a reply to comments on Smith. PMID- 12133138 TI - "Connectedness" of the tissue repair research community. PMID- 12133139 TI - A randomized trial using computerized decision support to improve treatment of major depression in primary care. AB - OBJECTIVE: To examine whether feedback and treatment advice for depression presented to primary care physicians (PCPs) via an electronic medical record (EMR) system can potentially improve clinical outcomes and care processes for patients with major depression. DESIGN: Randomized controlled trial. SETTING: Academically affiliated primary care practice in Pittsburgh, PA. PATIENTS: Two hundred primary care patients with major depression on the Primary Care Evaluation of Mental Disorders (PRIME-MD) and who met all protocol-eligibility criteria. INTERVENTION: PCPs were randomly assigned to 1 of 3 levels of exposure to EMR feedback of guideline-based treatment advice for depression: "active care" (AC), "passive care" (PC), or "usual care" (UC). MEASUREMENTS AND MAIN RESULTS: Patients' 3- and 6-month Hamilton Rating Scale for Depression (HRS-D) score and chart review of PCP reports of depression care in the 6 months following the depression diagnosis. Only 22% of patients recovered from their depressive episode at 6 months (HRS-D /=3 contacts with usual PCP at 6 months: 31% AC, 31% PC, 18% UC; P =.09 and antidepressant medication suggested/prescribed or baseline regimen modified at 6 months: 59% AC, 57% PC, 52% UC; P =.3). CONCLUSIONS: Screening for major depression, electronically informing PCPs of the diagnosis, and then exposing them to evidence-based treatment recommendations for depression via EMR has little differential impact on patients' 3- or 6-month clinical outcomes or on process measures consistent with high-quality depression care. PMID- 12133140 TI - Noncompliance with antihypertensive medications: the impact of depressive symptoms and psychosocial factors. AB - OBJECTIVE: Addressing the epidemic of poor compliance with antihypertensive medications will require identifying factors associated with poor adherence, including modifiable psychosocial and behavioral characteristics of patients. DESIGN: Cross-sectional study, comparing measured utilization of antihypertensive prescriptions with patients' responses to a structured interview. STUDY POPULATION: Four hundred ninety-six treated hypertensive patients drawn from a large HMO and a VA medical center. DATA COLLECTION: We developed a survey instrument to assess patients' psychosocial and behavioral characteristics, including health beliefs, knowledge, and social support regarding blood pressure medications, satisfaction with health care, depression symptom severity, alcohol consumption, tobacco use, and internal versus external locus of control. Other information collected included demographic and clinical characteristics and features of antihypertensive medication regimens. All prescriptions filled for antihypertensive medications were used to calculate actual adherence to prescribed regimens in a 365-day study period. MAIN OUTCOME OF INTEREST: Adjusted odds ratios (ORs) of antihypertensive compliance, based on ordinal logistic regression models. RESULTS: After adjusting for the potential confounding effects of demographic, clinical, and other psychosocial variables, we found that depression was significantly associated with noncompliance (adjusted OR per each point increase on a 14-point scale, 0.93; 95% confidence interval [95% CI], 0.87 to 0.99); in unadjusted analyses, the relationship did not reach statistical significance. There was also a trend toward improved compliance for patients perceiving that their health is controlled by external factors (adjusted OR per point increase, 1.14; 95% CI, 0.99 to 1.33). There was no association between compliance and knowledge of hypertension, health beliefs and behaviors, social supports, or satisfaction with care. CONCLUSIONS: Depressive symptoms may be an underrecognized but modifiable risk factor for poor compliance with antihypertensive medications. Surprisingly, patient knowledge of hypertension, health beliefs, satisfaction with care, and other psychosocial variables did not appear to consistently affect adherence to prescribed regimens. PMID- 12133141 TI - Clinical importance of HIV and depressive symptoms among veterans with HIV infection. AB - OBJECTIVE: To compare the clinical importance (association with illness severity and survival) of depressive and HIV symptoms among veterans with HIV infection. DESIGN: Cross-sectional study; survival analysis. SETTING: Infectious Disease Clinics at 3 VA Medical Centers. PARTICIPANTS: HIV-infected patients (N = 881) and their health care providers from June 1999 through July 2000. MEASUREMENTS AND MAIN RESULTS: Depressive symptoms were assessed using the 10-item Centers for Epidemiologic Studies Depression Scale (CES-D). Patient baseline survey included an HIV Symptom Index measuring the frequency and bother of 20 common symptoms. Providers were surveyed on patients' illness severity, and survival data were obtained from VA death records. Of 881 patients, 46% had significant depressive symptoms (CES-D >/=10). Increasing depression symptom severity was associated with increasing HIV symptom frequency (P <.001) and bother (P <.001). Multiple regression results revealed that having moderate or severe depressive symptoms was not associated with provider-reported illness severity or survival. However, HIV symptoms were significantly associated with provider-reported illness severity (P <.01) and survival (P =.05), after adjusting for moderate and severe depressive symptoms, CD4 cell count/mm3, viral load, age, race, and antiretroviral use. CONCLUSIONS: Depression, while common in this sample, was not associated with illness severity or mortality after adjusting for HIV symptoms. HIV symptoms are associated with severity of illness and survival regardless of patients' severity of depressive symptoms. This suggests that equal medical consideration should be given to HIV symptoms presented by HIV-infected patients regardless of their depression status, rather than automatically attributing medical complaints to depression. PMID- 12133142 TI - The use of computerized birthday greeting reminders in the management of diabetes. AB - BACKGROUND: Although mailed reminders have been used for prevention among general populations, few studies have evaluated their effectiveness among chronically ill populations. OBJECTIVE: We evaluated the effectiveness of mailed reminders for improving diabetes management. The reminder included a letter from the individual's primary care physician (PCP), a self-care handbook, a preventive care checklist, and specific recommendations regarding receipt of routine monitoring and screening. METHODS: Of 195 PCPs practicing with a large group practice, 111 agreed to have their adult patients with diabetes randomized to receive the reminder (n = 1,641) or usual care (n = 1,668). Using data from automated databases, we fit generalized estimating equations to evaluate the effect of reminder receipt on fasting lipid profile and glycated hemoglobin testing, dilated retinal exam receipt, and visit frequency during the 6 and 12 months following randomization, and glycated hemoglobin and cholesterol levels in the year following randomization. RESULTS: Reminder and usual care recipients did not differ in sociodemographic, clinical, or prior testing characteristics. In the 6 months following randomization, reminder recipients were more likely to receive a retinal exam (odds ratio [OR], 1.29; 95% confidence interval [95% CI], 1.12 to 1.49) and diabetes visit (OR, 1.28; 95% CI, 1.12 to 1.47). In the 12 months following randomization, reminder recipients were more likely to receive a glycated hemoglobin test (OR, 1.21; 95% CI, 1.03 to 1.43), retinal exam (OR, 1.23; 95% CI, 1.07 to 1.41), and diabetes visit (OR, 1.25; 95% CI, 1.09 to 1.29). In the follow-up year, reminder recipients also tended to have a glycated hemoglobin test that did not reflect poor control (<9.5%). CONCLUSIONS: We found small but significant improvements in the management of patients with diabetes receiving a computerized mailed reminder. PMID- 12133143 TI - Acculturation of attitudes toward end-of-life care: a cross-cultural survey of Japanese Americans and Japanese. AB - OBJECTIVE: Cross-cultural ethical conflicts are common. However, little is known about how and to what extent acculturation changes attitudes toward end-of-life care and advance care planning. We compared attitudes toward end-of-life care among Japanese Americans and Japanese in Japan. DESIGN: Self-administered questionnaire in English and Japanese. SETTING AND PARTICIPANTS: Community-based samples of Japanese Americans in Los Angeles and Japanese in Nagoya, Japan: 539 English-speaking Japanese Americans (EJA), 340 Japanese-speaking Japanese Americans (JJA), and 304 Japanese living in Japan (JJ). MEASUREMENTS AND MAIN RESULTS: Few subjects (6% to 11%) had discussed end-of-life issues with physicians, while many (EJA, 40%; JJA, 55%; JJ, 54%) desired to do so. Most preferred group surrogate decision making (EJA, 75%; JJA, 57%; JJ, 69%). After adjustment for demographics and health status, desire for informing the patient of a terminal prognosis using words increased significantly with acculturation (EJA, odds ratio [OR] 8.85; 95% confidence interval, [95% CI] 5.4 to 14.3; JJA, OR 2.8; 95% CI 1.8 to 4.4; JJ, OR 1.0). EJA had more-positive attitudes toward forgoing care, advance care planning, and autonomous decision making. CONCLUSION: Preference for disclosure, willingness to forgo care, and views of advance care planning shift toward western values as Japanese Americans acculturate. However, the desire for group decision making is preserved. Recognition of the variability and acculturation gradient of end-of-life attitudes among Japanese Americans may facilitate decision making and minimize conflicts. Group decision making should be an option for Japanese Americans. PMID- 12133144 TI - Learning about screening using an online or live lecture: does it matter? AB - OBJECTIVE: To determine the impact of an online lecture versus a live lecture on screening given to medical students who are participating in an outpatient clerkship. DESIGN: Prospective, randomized, controlled study. PARTICIPANTS AND SETTING: Ninety-five senior medical students in a primary care medicine clerkship based at university and distant clinic sites. INTERVENTION AND MEASUREMENTS: Forty-eight medical students were randomized to the live lecture on screening (live lecture group), and forty-seven medical students were randomized to the online lecture on screening (online lecture group). Outcome measures included students' knowledge, use of time, and satisfaction with the lecture experience. RESULTS: Compared to students in the live lecture group, students in the online lecture group demonstrated equal post-intervention knowledge of screening (P =.91) and expended 50 minutes less time to complete the lecture. Online lecture students who used the audio feed of the lecture were equally satisfied with the lecture as the live lecture students. Without the audio feed, online lecture students were less satisfied. CONCLUSIONS: An online lecture on screening is a feasible, efficient, and effective method to teach students on outpatient clerkships about principles of screening. PMID- 12133145 TI - Physician surveys to assess customary care in medical malpractice cases. AB - OBJECTIVE: Physician experts hired and prepared by the litigants provide most information on standard of care for medical malpractice cases. Since this information may not be objective or accurate, we examined the feasibility and potential value of surveying community physicians to assess standard of care. DESIGN: Seven physician surveys of mutually exclusive groups of randomly selected physicians. SETTING: Iowa. PARTICIPANTS: Community and academic primary care physicians and relevant specialists. INTERVENTIONS: Included in each survey was a case vignette of a primary care malpractice case and key quotes from medical experts on each side of the case. Surveyed physicians were asked whether the patient should have been referred to a specialist for additional evaluation. The 7 case vignettes included 3 closed medical malpractice cases, 3 modifications of these cases, and 1 active case. MEASUREMENTS AND MAIN RESULTS: Sixty-three percent of 350 community primary care physicians and 51% of 216 community specialists completed the questionnaire. For 3 closed cases, 47%, 78%, and 88% of primary care physician respondents reported that they would have made a different referral decision than the defendant. Referral percentages were minimally affected by modifying patient outcome but substantially changed by modifying patient presentation. Most physicians, even those whose referral decisions were unusual, assumed that other physicians would make similar referral decisions. For each case, at least 65% of the primary care physicians disagreed with the testimony of one of the expert witnesses. In the active case, the response rate was high (71%), and the respondents did not withhold criticism of the defendant doctor. CONCLUSIONS: Randomly selected peer physicians are willing to participate in surveys of medical malpractice cases. The surveys can be used to construct the distribution of physician self-reported practice relevant to a particular malpractice case. This distribution may provide more information about customary practice or standard of care than the opinion of a single physician expert. PMID- 12133147 TI - The internal medicine subinternship: a curriculum needs assessment. AB - Despite broad acceptance of the internal medicine subinternship rotation by the undergraduate medical education community, only a small fraction of programs provide students with explicit learning objectives. To design a curriculum for the medical subinternship, we surveyed 3 different groups of educational stakeholders--subinternship directors, residency program directors, and housestaff--in order to identify and prioritize the competencies that should be learned during this rotation. This study provides a starting point for the development of a structured curriculum for the fourth-year subinternship rotation. PMID- 12133146 TI - Receipt of nutrition and exercise counseling among medical outpatients with psychiatric and substance use disorders. AB - OBJECTIVE: Mentally ill persons represent a population that is potentially vulnerable to receiving a poorer quality of medical care. This study examines the relationship between mental disorders and the likelihood of receiving recommended nutrition and exercise counseling. DESIGN: Cross-sectional study combining chart review data and administrative database records. SETTING: One hundred forty-seven Veterans Affairs (VA) medical centers nationwide. PATIENTS/PARTICIPANTS: The sample included 90,240 patients with obesity and/or hypertension who had >/=3 medical outpatient visits in the previous year. MEASUREMENTS AND MAIN RESULTS: The outcomes of interest were chart-documented receipt of nutrition counseling and receipt of exercise counseling in the past 2 years. This chart information was merged with VA inpatient and outpatient administrative databases, which were used to identify persons with diagnosed mental disorders. Most patients received nutrition counseling (90.4%), exercise counseling (88.5%), and counseling for both (85.7%) in the past 2 years. The rates of counseling differed significantly but modestly by mental health status. The lowest rates were found among patients dually diagnosed with comorbid psychiatric and substance use disorders; however, the magnitude of the disparities was small, ranging from 2% to 4% across outcomes. These results were unchanged after controlling for demographics, health status, and facility characteristics using multivariable generalized estimating equation modeling. CONCLUSIONS: Among patients engaged in active medical treatment, rates of nutrition and exercise counseling were high at VA medical centers, and the diagnosis of mental illness was not a substantial barrier to such counseling. More work is needed to determine whether these findings generalize to non-VA settings and to understand the potential role that integrated systems such as the VA can play in reducing disparities for vulnerable populations. PMID- 12133148 TI - Clinicians, educators, and investigators in general internal medicine: bridging the gaps. AB - Financial and time pressures, disparate promotional pathways, geographic separation, and difficulty acknowledging personal fallibility can contribute to polarization of clinician-educators and investigators in general internal medicine (GIM). As a consequence, clinician-educators and investigators may fail to use their joint expertise, may encounter friction in their relationships, and may present a troubled image to trainees considering careers in GIM. We suggest specific strategies that clinician-educators, investigators, administrative leaders, and medical schools might use to foster collaboration. PMID- 12133149 TI - Erstwhile triple threat. PMID- 12133150 TI - Online instruction: time to grow. PMID- 12133152 TI - Fold after fold. PMID- 12133151 TI - Care rendered by general internists, committed to the care of HIV-infected patients, compares favorably with that given by infectious disease physicians. PMID- 12133153 TI - Racial and ethnic disparity in blood pressure and cholesterol measurement. AB - OBJECTIVE: To evaluate racial and ethnic disparity in blood pressure and cholesterol measurement and to analyze factors associated with any observed disparity. DESIGN: Cross-sectional analysis of the household component of the 1996 Medical Expenditure Panel Survey. PARTICIPANTS: Representative sample of the U.S. non-institutionalized population age 21 or older. MEASUREMENTS: Prevalence of self-reported blood pressure measurement within 2 years and cholesterol measurement within 5 years were calculated by race/ethnicity. Logistic regression was used to adjust for health insurance status, having a usual source of care, health status, and socioeconomic and demographic factors. Odds ratios and 95% confidence intervals (95% CIs) from the logistic regression were converted to prevalence ratios to estimate relative risk (RR). MAIN RESULTS: Mexican Americans compared to non-Hispanic whites were less likely to have a blood pressure measurement (RR, 0.85; 95% CI, 0.81 to 0.89) or a cholesterol measurement (RR, 0.72; 95% CI, 0.65 to 0.78). Non-Hispanic blacks had blood pressure and cholesterol measurements similar to non-Hispanic whites. In a multivariate analysis, Mexican Americans had similar blood pressure measurements (RR, 0.97; 95% CI, 0.94 to 1.00) and cholesterol measurements (RR, 1.04; 95% CI, 0.99 to 1.08). The factors associated with the largest disparity were lack of health insurance, not having a usual source of care, and low education. CONCLUSIONS: No disparity was found between non-Hispanic blacks and non-Hispanic whites in undergoing blood pressure and cholesterol measurement. Disparities in cardiovascular preventive services for Mexican Americans were associated with lack of health insurance and a usual source of care, but other demographic and socioeconomic factors were also important. PMID- 12133154 TI - Health-related quality of life in urban African Americans with type 2 diabetes. AB - OBJECTIVE: To examine the association of socioeconomic barriers, familial barriers, and clinical variables with health-related quality of life (HRQL). METHODS: A cross-sectional study was conducted of 186 African Americans with type 2 diabetes recruited from 2 primary care clinics in East Baltimore, Maryland. Physical functioning, social functioning, mental health, and general health were measured using the Medical Outcomes Study 36-item short form. Socioeconomic (money, housing, street crime) and familial (family problems, caretaker responsibilities) barriers were assessed by standardized interview. Insulin use, comorbid disease, and measured abnormalities in body mass index, hemoglobin A1c (HbA1c), blood pressure, lipids, and renal function were investigated. RESULTS: Mean HRQL scores were: physical functioning, 61 +/- 29; social functioning, 76 +/ 26; mental health, 69 +/- 21; and general health, 48 +/- 21. Linear regression analyses revealed that each barrier to care was significantly associated with lower scores in 1 or more HRQL domain. As number of socioeconomic and familial barriers increased from 0 to 5, HRQL scores decreased by 18 for social functioning, 21 for general health, 23 for physical functioning, and 28 for mental health (all P for trend <.01). Clinical variables significantly associated with reduced HRQL were obesity, impaired renal function, insulin use, and comorbid disease. Blood pressure, lipids, and HbA1c were not significantly associated with HRQL. CONCLUSIONS: An independent, graded relationship was found between socioeconomic and familial barriers to care and HRQL. This relationship was at least as strong as the association between HRQL and the clinical variables more likely to be perceived by participants as causing symptomatic distress or impacting lifestyle. PMID- 12133156 TI - Actual and potential effects of medical resident coverage on reimbursement for inpatient visits by attending physicians. AB - CONTEXT: The impact of residents on hospital finance has been studied; there are no data describing the economic effect of residents on attending physicians. OBJECTIVE: In a community teaching hospital, we compared allowable inpatient visit codes and payments (based on documentation in the daily progress notes) between a general medicine teaching unit and nonteaching general medicine units. DESIGN: Retrospective chart review, matched cohort study. SETTING: Six hundred fifty-bed community teaching hospital. PATIENTS: Patients were discharged July 1998 through February 1999 from Saint Barnabas Medical Center. We randomly selected 200 patients in quartets. Each quartet consisted of a pair of patients cared for by residents and a pair cared for only by an attending physician. In each pair, 1 of the patients was under the care of an attending physician who usually admitted to the teaching service, and 1 was under the care of a usually nonteaching attending. Within each quartet, patients were matched for diagnosis related group, length of stay, and discharge date. MAIN OUTCOME MEASURES: We assigned the highest daily visit code justifiable by resident and attending chart documentation, determining relative value units (RVUs) and reimbursements allowed by each patient's insurance company. RESULTS: Although more seriously ill, teaching-unit patients generated a mean 1.75 RVUs daily, compared with 1.84 among patients discharged from nonteaching units (P =.3). Median reimbursement, daily and per hospitalization, was similar on teaching and nonteaching units. Nonteaching attendings documented higher mean daily RVUs than teaching attendings (1.83 vs 1.76, P =.2). Median allowable reimbursements were $267 per case ($53 daily) among teaching attendings compared with $294 per case ($58 daily) among nonteaching attendings (Z = 1.54, P =.1). When only the resident note was considered, mean daily RVUs increased 39% and median allowable dollars per day 27% (Z = 4.21, P <.001). CONCLUSIONS: Nonteaching attendings appear to document their visits more carefully from a billing perspective than do teaching attendings. Properly counter-documented, resident notes could substantially increase payments to attending physicians. PMID- 12133155 TI - Effect of student involvement on patient perceptions of ambulatory care visits: a randomized controlled trial. AB - OBJECTIVE: To determine if patient satisfaction with ambulatory care visits differs when medical students participate in the visit. DESIGN: Randomized controlled trial. SETTING: Academic general internal medicine practice. PARTICIPANTS: Outpatients randomly assigned to see an attending physician only (N = 66) or an attending physician plus medical student (N = 68). MEASUREMENTS AND MAIN RESULTS: Patient perceptions of the office visit were determined by telephone survey. Overall office visit satisfaction was higher for the "attending physician only" group (61% vs 48% excellent), although this was not statistically significant (P =.16). There was no difference between the study groups for patient ratings of their physician overall (80% vs 85% excellent; P =.44). In subsidiary analyses, patients who rated their attending physician as "excellent" rated the overall office visit significantly higher in the "attending physician only" group (74% vs 55%; P =.04). Among patients in the "attending physician plus medical student" group, 40% indicated that medical student involvement "probably" or "definitely" did not improve their care, and 30% responded that they "probably" or "definitely" did not want to see a student at subsequent office visits. CONCLUSIONS: Although our sample size was small, we found no significant decrement in patient ratings of office visit satisfaction from medical student involvement in a global satisfaction survey. However, a significant number of patients expressed discontent with student involvement in the visit when asked directly. Global assessment of patient satisfaction may lack sensitivity for detection of dissatisfaction. Future research in this area should employ more sensitive measures of patient satisfaction. PMID- 12133157 TI - Improving the National Board of Medical Examiners internal Medicine Subject Exam for use in clerkship evaluation. AB - OBJECTIVE: To provide a consensus opinion on modifying the National Board of Medical Examiners (NBME) Medicine Subject Exam (Shelf) to: 1) reflect the internal medicine clerkship curriculum, developed by the Society of General Internal Medicine (SGIM) and the Clerkship Directors in Internal Medicine (CDIM); 2) emphasize knowledge important for a clerkship student; and 3) obtain feedback about students' performances on the Shelf. DESIGN: Two-round Delphi technique. PARTICIPANTS: The CDIM Research and Evaluation Committee and CDIM members on NBME Step 2 Committees. MEASUREMENTS: Using 1-5 Likert scales (5 = highest ratings), the group rated test question content for relevance to the SGIM-CDIM Curriculum Guide and importance for clerkship students' knowledge. The Shelf content is organized into 4 physician tasks and into 11 sections that are generally organ system based. Each iteration of the Shelf has 100 questions. Participants indicated a desired distribution of questions by physician task and section, topics critical for inclusion on each exam, and new topics to include. They specified the types of feedback clerkship directors desired on students' performances. Following the first round, participants viewed pooled results prior to submitting their second-round responses. RESULTS: Of 15 individuals contacted, 12 (80%) participated in each round. The desired distribution by physician task was: diagnosis (43), treatment (23), mechanism of disease (20), and health maintenance (15). The sections with the most questions requested were the cardiovascular (17), respiratory (15), and gastroenterology (12) sections. The fewest were requested in aging/ethics (4) and neurology, dermatology, and immunology (5 each). Examples of low-rated content were Wilson's Disease, chancroid and tracheal rupture (all <2.0). Health maintenance in type 2 diabetes, hypertension, and cardiovascular disease all received 5.0 ratings. Participants desired feedback by: section (4.6) and physician task (3.9), on performances of the entire class (4.0), and for individual students (3.8). CONCLUSION: Clerkship directors identified test content that was relevant to the curricular content and important for clerkship students to know, and they indicated a desired question distribution. They would most like feedback on their students' performance by organ system-based sections for the complete academic year. This collaborative effort could serve as a model for aligning national exams with course goals. PMID- 12133158 TI - An instructional program to facilitate teaching joint/soft-tissue injection and aspiration. AB - OBJECTIVE: We developed an instructional program to teach aspiration and injection techniques of the knee and shoulder to medical students and residents. METHODS: Residents and fourth-year medical students participating in a rheumatology elective were assigned by deterministic allocation into 3 groups: the Traditional group received no specific instruction in arthrocentesis but simply rotated through rheumatology, learning injection techniques only if they saw patients who required them; the Lecture-only group received only the didactic lecture and did not have the opportunity to practice on the models; the Program group participated in the newly developed program of instruction that combined a didactic lecture and a hands-on workshop using the anatomic models to practice arthrocentesis techniques. RESULTS: The scores on the written examination for those in the Program group (mean score 37.46 out of 40 possible) and the Lecture only group (mean 37.75) were significantly higher than those of the Traditional group (mean 33.15) (P <.05). The scores on the practical examination for those in the Program group (mean score 24.08 out of 26 possible) were significantly higher than those of the Lecture-only (mean 20.50) and Traditional (mean 17.33) (P <.05) CONCLUSION: The addition of this type of instruction to supplement a traditional internal medicine rotation can enhance a learner's ability to perform joint/soft tissue injection and aspiration. PMID- 12133159 TI - Preferences for patient cost sharing among medicare beneficiaries after HMO plan withdrawals. AB - OBJECTIVE: To assess Medicare beneficiaries' willingness to cost share in order to minimize disruptions in coverage from HMO plan withdrawals. DESIGN: Cross sectional survey of Medicare beneficiaries from February 1999 to March 1999. SETTING: Ten U.S. counties with the highest HMO plan withdrawal rates. PATIENTS/PARTICIPANTS: Seven hundred one Medicare beneficiaries for response rate of 69%. MEASUREMENTS AND MAIN RESULTS: Percentage of respondents willing to accept more out-of-pocket costs in order to continue their Medicare HMO coverage. Most respondents (67%) were willing to pay more out-of-pocket costs so that their HMO could have continued Medicare coverage. Those who were white (P =.03), had higher incomes (P =.01), and returned to traditional fee-for-service Medicare (P =.004) were more likely than other respondents to accept increased patient cost sharing. Most beneficiaries preferred Medicare policies requiring HMOs to sign longer-term Health Care Financing Administration (HCFA) contracts (72%) and to offer coverage to beneficiaries regardless of where they lived in a given state (87%). However, respondents' preferences for such policy options were not associated with the amount of cost sharing that respondents were willing to accept. CONCLUSIONS: Most Medicare beneficiaries are willing to accept increased patient cost sharing in order to reduce disruptions in their HMO coverage. Policies intended to reduce HMO plan withdrawals, such as requiring health plans to sign longer-term HCFA contracts, are supported by many Medicare beneficiaries, but these policy preferences were not related to willingness to cost share. In light of an apparent willingness to pay more out-of-pocket medical costs, Medicare beneficiaries in general may accept increased cost sharing in order to retain their HMO coverage. PMID- 12133160 TI - Receipt of preventive services among privately insured minorities in managed care versus fee-for-service insurance plans. AB - OBJECTIVE: We compare preventive services utilization among privately insured African Americans and Hispanics in managed care organizations (MCOs) versus fee for-service (FFS) plans. We also examine racial/ethnic disparities in the receipt of preventive services among enrollees in FFS or MCO plans. DESIGN: Analysis of the nationally representative 1996 Medical Expenditure Panel Survey. PARTICIPANTS: Participants included 1,120 Hispanic, 929 African-American, and 6,383 non-Hispanic white (NHW) adults age 18 to 64 years with private health insurance. MEASUREMENTS AND MAIN RESULTS: We examined self-reported receipt of physical examination, blood pressure measurement, cholesterol assessment, Papanicolau testing, screening mammography, and breast and prostate examinations. Multivariate modeling was used to adjust for age, gender, education, household income, and health status. Hispanics in MCOs were more likely than their FFS counterparts to report having preventive services, with adjusted differences ranging from 5 to 19 percentage points (P <.05 for physical examination, blood pressure measurement, breast examination and Pap smear). Among African Americans, such patterns were of a smaller magnitude. In both MCOs and FFS plans the proportion of African Americans reporting preventive services was equal to or greater than NHWs. In contrast, among Hispanic women in FFS, a non-statistically significant trend of fewer cancer screening tests than NHW's was observed (Pap smears 75% vs 80%; mammograms 66% vs 74%, respectively). In both MCO and FFS plans, Hispanics were less likely than NHWs to report having blood pressure and cholesterol measurement (P <.05). CONCLUSIONS: With the demise of traditional MCOs, reform efforts should incorporate those aspects of MCOs that were associated with greater preventive service utilization, particularly among Hispanics. Existing ethnic disparities warrant further attention. PMID- 12133161 TI - Influence of usual source of care on differences by race/ethnicity in receipt of preventive services. AB - OBJECTIVE: We examined the relation between race/ethnicity and receipt of preventive services and the effect of having a usual source of care (USOC) on receipt of preventive services in different racial and ethnic groups. DESIGN/PARTICIPANTS: We analyzed data from adults, aged 18 to 64 years in the Household Component of the 1996 Medical Expenditure Panel Survey, a nationally representative survey of health care use for the United States. MEASUREMENTS: The proportion of adults who received age-appropriate preventive services. RESULTS: Compared to white respondents, Hispanics were less likely to receive breast exams and blood pressure and cholesterol screening than were white respondents, and blacks were more likely to report receiving a Pap smear. Despite being less likely to report having a USOC, black and Hispanic women were as likely or more likely to report receiving breast and cervical cancer screening, after controlling for having a USOC and other factors. Hispanics reported receiving blood pressure screening less often, and blacks reported receiving more cholesterol screening. For each race/ethnicity group, having a USOC was associated with receiving preventive services. However, controlling for USOC and other confounders attenuated, but did not eliminate, differences by race/ethnicity. CONCLUSION: The differences by race in receipt of preventive services suggest the need for different starting points for devising strategies to address racial differences in disease outcomes. While having a USOC will be important in narrowing the differences by race in receipt of preventive services, attending to other factors that contribute to disparities in health will also be essential. PMID- 12133162 TI - An assessment of residents' abilities to detect and manage domestic violence. AB - Despite increased awareness of domestic violence (DV), little is known about residents' preparedness to diagnose and respond appropriately to abuse victims. We designed a pilot study to examine this. Seventy-one internal medicine residents participated in a 10-station standardized patient-based Clinical Skills Assessment. Forty (56%) were male and 31 (44%) were female; 46 (65%) were PGY I; 63 (89%) were trained internationally. One station presented a woman with headaches, whose underlying issue was DV. Forty (56%) residents correctly diagnosed DV. Thirty referred the patient for DV counseling. Eighteen addressed immediate safety concerns, and 23 asked about child abuse. Forty-eight (68%) made 1 or more incorrect recommendations. Thirty-six (51%) ordered unnecessary tests. Residents who did not diagnose DV spent nearly twice as much per patient on work up (mean, $942.00), compared to those who diagnosed DV (mean, $421.00). Use of certain interviewing skills appeared to promote elicitation of DV. Assessment driven educational interventions could help trainees improve their recognition of DV and make appropriate and cost-effective management choices. PMID- 12133163 TI - Determination of the clinical importance of study results. AB - Formal statistical methods for analyzing clinical trial data are widely accepted by the medical community. Unfortunately, the interpretation and reporting of trial results from the perspective of clinical importance has not received similar emphasis. This imbalance promotes the historical tendency to consider clinical trial results that are statistically significant as also clinically important, and conversely, those with statistically insignificant results as being clinically unimportant. In this paper, we review the present state of knowledge in the determination of the clinical importance of study results. This work also provides a simple, systematic method for determining the clinical importance of study results. It uses the relationship between the point estimate of the treatment effect (with its associated confidence interval) and the estimate of the smallest treatment effect that would lead to a change in a patient's management. The possible benefits of this approach include enabling clinicians to more easily interpret the results of clinical trials from a clinical perspective, and promoting a more rational approach to the design of prospective clinical trials. PMID- 12133165 TI - Medical student education in ambulatory settings: does it affect patient satisfaction? PMID- 12133164 TI - Designing and evaluating interventions to eliminate racial and ethnic disparities in health care. AB - A large number of factors contribute to racial and ethnic disparities in health status. Health care professionals, researchers, and policymakers have believed for some time that access to care is the centerpiece in the elimination of these health disparities. The Institute of Medicine's (IOM) model of access to health services includes personal, financial, and structural barriers, health service utilization, and mediators of care. This model can be used to describe the interactions among these factors and their impact on health outcomes and equity of services among racial and ethnic groups. We present a modified version of the IOM model that incorporates the features of other access models and highlights barriers and mediators that are relevant for interventions designed to eliminate disparities in U.S. health care. We also suggest that interventions to eliminate disparities and achieve equity in health care services be considered within the broader context of improving quality of care. Some health service intervention studies have shown improvements in the health of disadvantaged groups. If properly designed and implemented, these interventions could be used to reduce health disparities. Successful features of interventions include the use of multifaceted, intense approaches, culturally and linguistically appropriate methods, improved access to care, tailoring, the establishment of partnerships with stakeholders, and community involvement. However, in order to be effective in reducing disparities in health care and health status, important limitations of previous studies need to be addressed, including the lack of control groups, nonrandom assignment of subjects to experimental interventions, and use of health outcome measures that are not validated. Interventions might be improved by targeting high-risk populations, focusing on the most important contributing factors, including measures of appropriateness and quality of care and health outcomes, and prioritizing dissemination efforts. PMID- 12133166 TI - The challenge of eliminating disparities in health. PMID- 12133167 TI - The challenge of residents with deficiencies of clinical performance of professionalism. PMID- 12133169 TI - Correlation of the score for subjective pain with physical disability, clinical and radiographic scores in recent onset rheumatoid arthritis. AB - BACKGROUND: To analyse the relationship between subjective pain score and other measures of clinical, radiographic and functional status; in particular Larsen radiographic scores and Health Assessment Questionnaire (HAQ); in patients with severe rheumatoid arthritis (RA) with a disease duration of less than 3 years. METHODS: In this cross sectional study of 105 patients with RA (76 women, 29 men: mean age 50.93; mean disease duration 15.86 months; 71% rheumatoid factor positive) subjective pain was assessed according to the Visual Analog Scale (VAS). Correlation coefficients between pain score and disease activity measures (patients' global assessment of disease by VAS, number of tender and swollen joints, morning stiffness, erythrocyte sedimentation rate [ESR], C-reactive protein [CRP] and titre of rheumatoid factor, radiographic evaluations (Larsen Dale scores for radiographic damage of the small joints of the hands, wrist and feet), disability measures (health assessment questionnaire [HAQ]), and demographic variables were calculated; hierarchical regression analysis was done with subjective pain score as the dependent variable. RESULTS: The Spearman's correlation coefficient comparing subjective pain and HAQ was 0.421 (p < 0.001), between subjective pain and global assessment of disease and morning stiffness was 0.573 (p < 0.001) and 0.427 (p < 0.001) respectively, and between pain and number of tender and swollen joints 0.037 and 0.050 respectively (p > 0.05). In regression analysis, global assessment of disease by patients explained 32.8% of the variation in pain intensity score, morning stiffness 10.7%, CRP 4.0%, HAQ 3.8% and Larsen-Dale scores explained 2.1%; other variables were not significant in the model. CONCLUSIONS: Pain scores of patients with early severe rheumatoid arthritis are correlated at higher levels with patients' global assessment of disease and with morning stiffness rather than with radiographic or other clinical variables such as number of tender and swollen joints. PMID- 12133171 TI - Delirium in critically ill patients. AB - Delirium in the intensive care unit is a serious problem that has recently attracted much attention. User-friendly and reliable tools, such as the Confusion Assessment Method for the Intensive Care Unit (CAM-ICU), offer the clinician the opportunity to identify delirium in patients better. Diagnosis of delirium in a critical care population is often a difficult task because classical psychiatric evaluation is impossible for a number of reasons. The CAM-ICU makes use of nonverbal assessments to evaluate the cardinal features of delirium (i.e. acute or fluctuating onset, inattention, disorganized thinking and altered level of consciousness). Its development for use in the critical care setting represents a significant advance that could lead to better care for such patients. PMID- 12133172 TI - Pro/con clinical debate: is high-frequency oscillatory ventilation useful in the management of adult patients with respiratory failure? AB - In neonatal and pediatric intensive care units, high-frequency oscillatory ventilation (HFOV) has become an increasingly common therapy. This may not have been the case if researchers had not persisted in investigating the therapy after early disappointing clinical trials. Devices capable of providing this therapy to adults have become commercially available relatively recently. However, there are many questions that need to be answered regarding HFOV in adults: Is HFOV in adults superior to conventional mechanical ventilation? Who is the ideal candidate for HFOV? When should it be applied? What is the best technique with which to apply it? When should a patient on HFOV be converted back to conventional ventilation? What is the safety and efficacy of the device? As outlined in the following debate, there are several compelling arguments for and against the use of HFOV at this point in adults. PMID- 12133170 TI - Oral cancer treatment costs in Greece and the effect of advanced disease. AB - BACKGROUND: The main purpose of the study was to quantify the direct costs of oral cancer treatment to the healthcare system of Greece. Another aim was to identify factors that affect costs and potential cost reduction items. More specifically, we examined the relationship between stage of disease, modality of treatment and total direct costs. METHODS: The medical records and clinic files of the Oral and Maxillofacial Clinic of the Athens General Hospital "Genimatas" were abstracted to investigate clinical treatment characteristics, including length of hospitalization, modes of treatment, stage of disease etc. Records of 95 patients with oral squamous cell carcinoma (OSSC), with at least six months of follow-up, were examined. The clinical data was then used to calculate actual direct costs, based on 2001 market values. RESULTS: The mean total direct costs for OSSC treatment estimated at euro 8,450 or approximately US$ 7,450. Costs depended on the stage of the disease, with significant increases in stages III and IV, as compared with stages I and II (p < 0.05). Multi-modality treatment applied mainly to patients in stages III and IV was the factor that affected the cost. Disease stage was also associated with the total duration of hospitalization (p < 0.05). CONCLUSIONS: The clinical management of advanced oral cancer is strongly associated with higher costs. Although the ideal would be to prevent cancer, the combination of high-risk screening, early diagnosis and early treatment seems the most efficient way to reduce costs, and most importantly, prolong life. PMID- 12133173 TI - What is the role of surfactant and inhaled nitric oxide in lung transplantation? AB - Although numerous studies over the past 40 years have addressed this problem, initial graft failure is still a key question in clinical lung transplantation. As a possible tool to avoid and treat initial graft failure after lung transplantation, laboratory evidence and clinical reports currently emphasize the role of substitution therapy of surfactant combined with inhaled nitric oxide. PMID- 12133174 TI - The customization of APACHE II for patients receiving orthotopic liver transplants. AB - General outcome prediction models developed for use with large, multicenter databases of critically ill patients may not correctly estimate mortality if applied to a particular group of patients that was under-represented in the original database. The development of new diagnostic weights has been proposed as a method of adapting the general model - the Acute Physiology and Chronic Health Evaluation (APACHE) II in this case - to a new group of patients. Such customization must be empirically tested, because the original model cannot contain an appropriate set of predictive variables for the particular group. In this issue of Critical Care, Arabi and co-workers present the results of the validation of a modified model of the APACHE II system for patients receiving orthotopic liver transplants. The use of a highly heterogeneous database for which not all important variables were taken into account and of a sample too small to use the Hosmer-Lemeshow goodness-of-fit test appropriately makes their conclusions uncertain. PMID- 12133175 TI - Cortisol replacement for severe sepsis and septic shock: what should I do? AB - Based on several recently completed randomized controlled trials, cortisol replacement is likely to become a standard of care for vasopressor dependent septic shock. Further studies are needed in order to accomplish whether this treatment should be limited to patients with a blunted cortisol response to corticotrophin. Similarly, in patients with severe sepsis who do not need vasopressors, the benefit/risk ratio of cortisol replacement remains to be assessed. PMID- 12133176 TI - Pro/con ethics debate: is nonheart-beating organ donation ethically acceptable? AB - This pro/con debate explores the ethical issues surrounding nonheart-beating organ donation (NHBD), a source of considerable controversy. It is estimated that NHBD can increase the number of organs available for transplant by 25% at a time of great need. However, should NHBD be ethically acceptable? In support of NHBD, it may be acceptable practice if there is a separation of the rationale to withdraw life support/to withhold cardiopulmonary resuscitation from the decision to recover organs, if no conflicts of interest exist, if a waiting time precluding spontaneous return of circulation is included, and if NHBD conforms to a standardized protocol. Against NHBD, there are questions regarding the ambiguity and cultural perspectives of death, regarding whether a separation of rationale between withdrawal and donation is sufficient to preclude conflicts of interest, and regarding whether variable protocols arise that subordinate the patient to the goal of donation. Such concerns suggest NHBD may damage the trust in patient-physician relationships and may adversely affect organ donation rates. PMID- 12133177 TI - Recently published papers: topical issues in pharmacology. PMID- 12133178 TI - Clinical review: complications and risk factors of peripheral arterial catheters used for haemodynamic monitoring in anaesthesia and intensive care medicine. AB - In order to evaluate the complications and risk factors associated with peripheral arterial catheters used for haemodynamic monitoring, we reviewed the literature published from 1978 to 2001. We closely examined the three most commonly used arterial cannulation sites. The reviewed papers included a total of 19,617 radial, 3899 femoral and 1989 axillary artery catheterizations. Factors that contribute to higher complication rates were investigated. Major complications occurred in fewer than 1% of the cases, and rates were similar for the radial, femoral and axillary arteries. We conclude that arterial cannulation is a safe procedure. PMID- 12133179 TI - Clinical review: bacteremia caused by anaerobic bacteria in children. AB - This review describes the microbiology, diagnosis and management of bacteremia caused by anaerobic bacteria in children. Bacteroides fragilis, Peptostreptococcus sp., Clostridium sp., and Fusobacterium sp. were the most common clinically significant anaerobic isolates. The strains of anaerobic organisms found depended, to a large extent, on the portal of entry and the underlying disease. Predisposing conditions include: malignant neoplasms, immunodeficiencies, chronic renal insufficiency, decubitus ulcers, perforation of viscus and appendicitis, and neonatal age. Organisms identical to those causing anaerobic bacteremia can often be recovered from other infected sites that may have served as a source of persistent bacteremia. When anaerobes resistant to penicillin are suspected or isolated, antimicrobial drugs such as clindamycin, chloramphenicol, metronidazole, cefoxitin, a carbapenem, or the combination of a beta-lactamase inhibitor and a penicillin should be administered. The early recognition of anaerobic bacteremia and administration of appropriate antimicrobial and surgical therapy play a significant role in preventing mortality and morbidity in pediatric patients. PMID- 12133180 TI - Bench-to-bedside review: fulfilling promises of the Human Genome Project. AB - Since most common diseases have been shown to be influenced by inherited variations in our genes, completion of the Human Genome Project and mapping of the human genome single-nucleotide polymorphisms will have a tremendous impact on our approach to medicine. New developments in genotyping techniques and bioinformatics, enabling detection of single-nucleotide polymorphisms, already provide physicians and scientists with tools that change our understanding of human biology. In the near future, studies will relate genetic polymorphisms to features of critical illnesses, increased susceptibility to common diseases, and altered response to therapy. Novel insights into the contribution of genetic factors to critical illnesses and advances in pharmacogenomics will be used to select the most effective therapeutic agent and the optimal dosage required to elicit the expected drug response for a given individual. Implementation of genetic criteria for patient selection and individual assessment of the risks and benefits of treatment emerges as a major challenge to the pharmaceutical industry. PMID- 12133181 TI - Equipment review: new techniques for cardiac output measurement--oesophageal Doppler, Fick principle using carbon dioxide, and pulse contour analysis. AB - Measuring cardiac output is of paramount importance in the management of critically ill patients in the intensive care unit and of 'high risk' surgical patients in the operating room. Alternatives to thermodilution are now available and are gaining acceptance among practitioners who have been trained almost exclusively in the use of the pulmonary artery catheter. The present review focuses on the principles, advantages and limitations of oesophageal Doppler, Fick principle applied to carbon dioxide, and pulse contour analysis. No single method stands out or renders the others obsolete. By making cardiac output easily measurable, however, these techniques should all contribute to improvement in haemodynamic management. PMID- 12133182 TI - Statistics review 3: hypothesis testing and P values. AB - The present review introduces the general philosophy behind hypothesis (significance) testing and calculation of P values. Guidelines for the interpretation of P values are also provided in the context of a published example, along with some of the common pitfalls. Examples of specific statistical tests will be covered in future reviews. PMID- 12133183 TI - Preoperative prediction of pediatric patients with effusions and edema following cardiopulmonary bypass surgery by serological and routine laboratory data. AB - AIM: Postoperative effusions and edema and capillary leak syndrome in children after cardiac surgery with cardiopulmonary bypass constitute considerable clinical problems. Overshooting immune response is held to be the cause. In a prospective study we investigated whether preoperative immune status differences exist in patients at risk for postsurgical effusions and edema, and to what extent these differences permit prediction of the postoperative outcome. METHODS: One-day preoperative serum levels of immunoglobulins, complement, cytokines and chemokines, soluble adhesion molecules and receptors as well as clinical chemistry parameters such as differential counts, creatinine, blood coagulation status (altogether 56 parameters) were analyzed in peripheral blood samples of 75 children (aged 3-18 years) undergoing cardiopulmonary bypass surgery (29 with postoperative effusions and edema within the first postoperative week). RESULTS: Preoperative elevation of the serum level of C3 and C5 complement components, tumor necrosis factor-alpha, percentage of leukocytes that are neutrophils, body weight and decreased percentage of lymphocytes (all P < 0.03) occurred in children developing postoperative effusions and edema. While single parameters did not predict individual outcome, >86% of the patients with postoperative effusions and oedema were correctly predicted using two different classification algorithms. Data mining by both methods selected nine partially overlapping parameters. The prediction quality was independent of the congenital heart defect. CONCLUSION: Indicators of inflammation were selected as risk indicators by explorative data analysis. This suggests that preoperative differences in the immune system and capillary permeability status exist in patients at risk for postoperative effusions. These differences are suitable for preoperative risk assessment and may be used for the benefit of the patient and to improve cost effectiveness. PMID- 12133184 TI - Pruritus: a useful sign for predicting the haemodynamic changes that occur following administration of vancomycin. AB - INTRODUCTION: The aim of this study was to investigate the haemodynamic changes that follow the appearance of pruritus during vancomycin administration. METHODS: We studied 50 patients scheduled for coronary artery bypass surgery, and we compared data from patients who exhibited pruritus with those from patients who did not. After the monitoring devices had been positioned, vancomycin (15 mg/kg) was continuously infused at a constant rate over 30 min, before induction of anaesthesia. Haemodynamic profiles were recorded before vancomycin infusion (time point 1); at 15 (time point 2) and 30 min (time point 3) after the beginning of vancomycin infusion; and 15 min after vancomycin infusion had been stopped (time point 4). At each time arterial and mixed venous blood samples were drawn to calculate the shunt fraction (Qsp/Qt). RESULTS: In patients who exhibited pruritus (group A, n = 17) at time point 3 versus time point 1, systemic vascular resistance index (SVRI) and arterial oxygen tension (PaO2) decreased significantly; cardiac index (CI), stroke volume index (SVI) and Qsp/Qt increased significantly; and mean systemic pressure and heart rate were stable. Those changes were observed only in patients not treated with a beta-blocker before surgery, whereas no change occurred in patients treated with the drug. In the patients who were free from pruritus (group B, n = 28), we did not observe any significant change. CONCLUSION: The appearance of pruritus during vancomycin administration indicates that SVRI is declining, thus exposing the patient to risk for hypotension. Therapy with a beta-blocker appears to confer protection against this hemodynamic reaction. PMID- 12133185 TI - Severe reperfusion lung injury after double lung transplantation. AB - AIM: To demonstrate the effects of combined inhaled nitric oxide and surfactant replacement as treatment for acute respiratory distress syndrome. This treatment has not previously been documented for reperfusion injury after double lung transplantation. METHOD: A 24-year-old female with cystic fibrosis underwent double lung transplantation. During implantation of the second lung a marked increase in pulmonary artery pressure associated with systemic hypotension, hypoxemia and low cardiac output were observed. Notwithstanding the patient received support from cardiovascular drugs and pulmonary vasodilators cardiopulmonary by-pass was necessary. In the intensive care unit the patient received the same drug support, inhaled nitric oxide and two bronchoscopic applications of bovine surfactant. RESULTS: A rapid improvement in PaO2/FiO2 within 2-3 hours of administration of surfactant was seen. The patient is well at follow-up 1 year post-transplant. CONCLUSION: There is a potential role for a combined therapy with inhaled nitric oxide and surfactant replacement in reperfusion injury after lung transplantation. PMID- 12133186 TI - External validation of a modified model of Acute Physiology and Chronic Health Evaluation (APACHE) II for orthotopic liver transplant patients. AB - INTRODUCTION: The purpose of the study was to validate the newly derived postoperative orthotopic liver transplantation (OLTX)-specific diagnostic weight for the Acute Physiology and Chronic Health Evaluation (APACHE) II mortality prediction system in independent databases. METHODS: Medical records of 174 liver transplantation patients admitted postoperatively to the adult intensive care units at King Fahad National Guard Hospital and the University of Wisconsin were reviewed, and data on age, sex, the underlying liver disease, APACHE II scores and the hospital outcome were collected. Predicted mortality was calculated using: 1) the original APACHE II diagnostic weight of postoperative other gastrointestinal surgery and 2) the newly derived OLTX-specific diagnostic category weight. Standardized mortality ratio and 95% confidence intervals were calculated. Calibration was evaluated with the Hosmer-Lemeshow goodness-of-fit C statistic. Discrimination was tested by 2 x 2 classification matrices and by computing the areas under the receiver operating characteristic curves. Patient characteristics and outcome data were compared between the two hospitals. RESULTS: APACHE II significantly overestimated mortality when the original diagnostic weight was used, but provided a closer estimate of mortality with the OTLX-specific diagnostic weight. The C-statistic analysis showed better calibration for the new approach; discrimination was also improved. The performances of the prediction systems were similar in the two hospitals. The new model provided more accurate estimates of hospital mortality in each hospital. DISCUSSION: APACHE II provided an accurate estimate of mortality in liver transplant patients when the OLTX-specific diagnostic weight was used. With the new model, APACHE II can be used as a valid mortality prediction system in this group of patients. PMID- 12133187 TI - Physiological-dose steroid therapy in sepsis [ISRCTN36253388]. AB - INTRODUCTION: The aim of the study was to assess the prognostic importance of basal cortisol concentrations and cortisol response to corticotropin, and to determine the effects of physiological dose steroid therapy on mortality in patients with sepsis. METHODS: Basal cortisol level and corticotropin stimulation test were performed within 24 hours in all patients. One group (20 patients) received standard therapy for sepsis and physiological-dose steroid therapy for 10 days; the other group (20 patients) received only standard therapy for sepsis. Basal cortisol level was measured on the 14th day in patients who recovered. The outcome of sepsis was compared. RESULTS: Only Sequential Organ Failure Assessment (SOFA) score was found related to mortality, independent from other factors in multivariate analysis. No significant difference was found between the changes in the percentage of SOFA scores of the steroid therapy group and the standard therapy group in survivors, nor between the groups in basal and peak cortisol levels, cortisol response to corticotropin test and mortality. The mortality rates among patients with occult adrenal insufficiencies were 40% in the steroid therapy group and 55.6% in the standard therapy group. DISCUSSION: There was a trend towards a decrease in the mortality rates of the patients with sepsis who received physiological-dose steroid therapy. In the advancing process from sepsis to septic shock, adrenal insufficiency was not frequent as supposed. There was a trend (that did not reach significance) towards a decrease in the mortality rates of the patients with sepsis who received physiological-dose steroid therapy. PMID- 12133188 TI - Management of severe organophosphorus pesticide poisoning. PMID- 12133189 TI - Potential therapeutic agents in the management of organophosphorus poisoning. PMID- 12133190 TI - Insulin and the critically ill. PMID- 12133191 TI - 22nd International Symposium on Intensive Care and Emergency Medicine, Brussels, Belgium, 19-22 March 2002. PMID- 12133192 TI - What are the challenges of translating positive trial results in severe sepsis into clinical practice? A media roundtable debate, 18 March 2002, Brussels, Belgium. AB - The clinical syndrome of sepsis is common, increasing in incidence and responsible for as many deaths annually as ischaemic heart disease. Two recent interventional trials have demonstrated that early recognition and intervention can result in dramatic reductions in acute (28-day) mortality. This roundtable discussion was convened to identify ways in which these recent advances could be translated into clinical practice. The first obstacle surrounds the woolly and confusing terminology surrounding 'sepsis' with the systemic inflammatory response syndrome (SIRS) model largely discredited. Overcoming this should facilitate wider recognition, not only among health care providers (in particular those working in acute specialties outside intensive care units [ICUs]) but also politicians and the general public. Such education is vital if early recognition and intervention are to be successfully implemented. PMID- 12133195 TI - Are women and providers satisfied with antenatal care? Views on a standard and a simplified, evidence-based model of care in four developing countries. AB - BACKGROUND: This study assessed women and providers' satisfaction with a new evidence-based antenatal care (ANC) model within the WHO randomized trial conducted in four developing countries. The WHO study was a randomized controlled trial that compared a new ANC model with the standard type offered in each country. The new model of ANC emphasized actions known to be effective in improving maternal or neonatal health, excluded other interventions that have not proved to be beneficial, and improved the information component, especially alerting pregnant women to potential health problems and instructing them on appropriate responses. These activities were distributed within four antenatal care visits for women that did not need any further assessment. METHODS: Satisfaction was measured through a standardized questionnaire administered to a random sample of 1,600 pregnant women and another to all antenatal care providers. RESULTS: Most women in both arms expressed satisfaction with ANC. More women in the intervention arm were satisfied with information on labor, delivery, family planning, pregnancy complications and emergency procedures. More providers in the experimental clinics were worried about visit spacing, but more satisfied with the time spent and information provided. CONCLUSIONS: Women and providers accepted the new ANC model generally. The safety of fewer visits for women without complications with longer spacing would have to be reinforced, if such a model is to be introduced into routine practice. PMID- 12133196 TI - Nutrition Society Medal lecture. The role of the skeleton in acid-base homeostasis. AB - Nutritional strategies for optimising bone health throughout the life cycle are extremely important, since a dietary approach is more popular amongst osteoporosis sufferers than drug intervention, and long-term drug treatment compliance is relatively poor. As an exogenous factor, nutrition is amenable to change and has relevant public health implications. With the growing increase in life expectancy, hip fractures are predicted to rise dramatically in the next decade, and hence there is an urgent need for the implementation of public health strategies to target prevention of poor skeletal health on a population-wide basis. The role that the skeleton plays in acid-base homeostasis has been gaining increasing prominence in the literature; with theoretical considerations of the role alkaline bone mineral may play in the defence against acidosis dating as far back as the late 19th century. Natural, pathological and experimental states of acid loading and/or acidosis have been associated with hypercalciuria and negative Ca balance and, more recently, the detrimental effects of 'acid' from the diet on bone mineral have been demonstrated. At the cellular level, a reduction in extracellular pH has been shown to have a direct enhancement on osteoclastic activity, with the result of increased resorption pit formation in bone. A number of observational, experimental, clinical and intervention studies over the last decade have suggested a positive link between fruit and vegetable consumption and the skeleton. Further research is required, particularly with regard to the influence of dietary manipulation using alkali-forming foods on fracture prevention. Should the findings prove conclusive, a 'fruit and vegetable' approach to bone health maintenance may provide a very sensible (and natural) alternative therapy for osteoporosis treatment, which is likely to have numerous additional health-related benefits. PMID- 12133197 TI - The Boyd Orr lecture. Nutrition interventions in aging and age-associated disease. AB - Aging is a complex biological process, which usually is accompanied by changes in socio-economic status, which may have a great impact on the physical and nutritional status of the elderly. Decreased food intake and a sedentary lifestyle in the growing numbers of the elderly increase their risk for malnutrition, decline of bodily functions and developing chronic diseases. Oxidative stress is believed to be an important factor in aging and many age associated degenerative diseases. Modulation of oxidative stress by energy restriction in animals has been shown to be one of the mechanisms for retarding the aging process. Dietary antioxidants are regarded as being important in modulating oxidative stress of aging and age-associated diseases. Supplementation of the elderly with vitamin E has been shown to enhance immune response, delay onset of Alzheimer's disease, and increase resistance to oxidative injury associated with exercise. Vitamin E, in comparison with other antioxidants, is also effective in reducing viral titres, but not the longevity of middle-aged mice. Our studies have indicated that polyphenols or vitamin E may assist in preventing cardiovascular disease, in part by decreasing expression by endothelial cells of proinflammatory cytokines, adhesion molecules, and monocyte adhesion. Most recently, we have found that some of these antioxidants may prevent tumour growth by inhibiting angiogenesis via suppression of interleukin 8 and modulation of the cell junction molecule, VE-cadherin. These findings provide further support for the consumption of fruit and vegetables, which contain several forms of phytochemicals with antioxidant activity, in order to reduce the risk of cardiovascular disease and cancer, the leading causes of morbidity and mortality among the elderly. PMID- 12133198 TI - Strategies for skeletal health in the elderly. AB - Osteoporosis is a common disease in the elderly, and the fractures that result from this disorder affect 40 % of women and 14 % of men over the age of 50 years. The risk of fracture relates to bone mineral density and the risk of falling, among other factors. Low bone mineral density in the elderly can result from either low peak bone mass or accelerated bone loss, or a combination of the two. Nutritional factors play a role in both the attainment of peak bone mass and in the rate of age-related bone loss. The main determinants of peak bone mass are genetic factors, early-life nutrition, diet and exercise. Of the nutritional factors Ca, and particularly milk, are the most important contributors to peak bone mass. Some of these factors may interact; for example, a low dietary Ca in addition to an unfavourable vitamin D receptor gene polymorphism may result in low peak bone mass. The age-related changes in bone mass may also have a genetic basis, but deficiency of oestrogen is a major contributor. In addition, undernutrition is common in the elderly, and lack of dietary protein contributes both to impaired bone mineral conservation and increased propensity to fall. There is a decreased ability of the intestine to adapt to a low-Ca diet with increasing age. Other dietary factors include vitamin K, Zn and fruit and vegetables. Adequate nutritional status, particularly of Ca and vitamin D, is essential for the successful pharmaceutical treatment of osteoporosis. Thus, strategies for enhancing skeletal health in the elderly must begin in early childhood, and continue throughout life. PMID- 12133199 TI - Is there a potential therapeutic value of copper and zinc for osteoporosis? AB - Osteoporosis is almost universal in very old age, and is a major cause of morbidity and mortality in the elderly of both sexes. Bone is lost at a rate of 0.2-0.5 %/year in both men and women after the age of 40-45 years. The causes of age-related changes in bone mass are multifactorial and include genetic predisposition, nutritional factors, endocrine changes, habitual exercise levels and body weight. Bone loss is accelerated to 2-5 % year immediately before and for up to 10 years post-menopause (Heaney, 1986). In women hormone-replacement therapy is effective in reducing the rate of bone loss caused by this peri menopausal decrease in hormone levels (Smith & Studd, 1993); however, in men and older women (>10 years post-menopause) nutrition plays a key role in the rate of bone loss. One factor contributing to bone loss in the elderly may be a subclinical Zn and/or Cu deficiency, due to a reduced dietary intake of micronutrients and reduced absorption (Thomson & Keelan, 1986). Zn and Cu are essential cofactors for enzymes involved in the synthesis of various bone matrix constituents. Paradoxically, Ca supplementation may accentuate the problem of reduced Zn and Cu levels by impairing the absorption of simultaneously-ingested Zn and the retention of Cu (Snedeker et al. 1982; Grekas et al. 1988). The present paper will review the current literature on the potential benefits of Cu and Zn supplementation in reducing bone loss, and present new information on the effect of Ca supplementation on Zn and Cu status in post-menopausal women with osteoporosis. PMID- 12133200 TI - Effects of micronutrient supplements on u.v.-induced skin damage. AB - Development of an orally-administered systemic agent that could reduce the effects of u.v. exposure on skin could potentially have a major effect on the incidence of skin cancers and photo-ageing. A number of micronutrients have been suggested to have metabolic properties that could induce this protection, and our data indicate that n-3 polyunsaturated fatty acids are particularly effective in this role. The mechanisms of action of n-3 polyunsaturated fatty acids appear to depend on their anti-inflammatory properties, acting to reduce the u.v.-induced release of cytokines and other mediators from a variety of skin cell types. PMID- 12133201 TI - Antioxidant strategies for Alzheimer's disease. AB - Oxidative damage is present within the brains of patients with Alzheimer's disease (AD), and is observed within every class of biomolecule, including nucleic acids, proteins, lipids and carbohydrates. Oxidative injury may develop secondary to excessive oxidative stress resulting from beta-amyloid-induced free radicals, mitochondrial abnormalities, inadequate energy supply, inflammation or altered antioxidant defences. Treatment with antioxidants is a promising approach for slowing disease progression to the extent that oxidative damage may be responsible for the cognitive and functional decline observed in AD. Although not a uniformly consistent observation, a number of epidemiological studies have found a link between antioxidant intake and a reduced incidence of dementia, AD and cognitive decline in elderly populations. In AD clinical trials molecules with antioxidant properties such as vitamin E and Ginkgo biloba extract have shown modest benefit. A clinical trial with vitamin E is currently ongoing to determine if it can delay progression to AD in individuals with mild cognitive impairment. Combinations of antioxidants might be of even greater potential benefit for AD, especially if the agents worked in different cellular compartments or had complementary activity (e.g. vitamins E, C and ubiquinone). Naturally-occurring compounds with antioxidant capacity are available and widely marketed (e.g. vitamin C, ubiquinone, lipoic acid, beta-carotene, creatine, melatonin, curcumin) and synthetic compounds are under development by industry. Nevertheless, the clinical value of these agents for AD prevention and treatment is ambiguous, and will remain so until properly designed human trials have been performed. PMID- 12133202 TI - The argument for increasing selenium intake. AB - The essential trace mineral, Se, is of fundamental importance to human health. As a constituent of selenoproteins it plays both structural and enzymic roles, in the latter context being best known as an antioxidant and catalyst for the production of active thyroid hormone. While Se-deficiency diseases have been recognised for some time, evidence is mounting that less-overt deficiency can also cause adverse health effects and furthermore, that supra-nutritional levels of Se may give additional protection from disease. In the context of these effects, low or diminishing Se status in some parts of the world, notably in some European countries such as the UK, is giving cause for concern. While deficiency has an adverse effect on immunocompetence, Se supplementation appears to enhance the immune response. Se appears to be a key nutrient in counteracting certain viral infections; thus, in a Se-deficient host the benign coxsackie virus becomes virulent, causing heart damage, the influenza virus causes more serious lung pathology and HIV infection progresses more rapidly to AIDS. Long recognised as essential for successful animal reproduction, Se is required for human sperm maturation and sperm motility and may reduce the risk of miscarriage. Deficiency has been linked to adverse mood states. Findings have been equivocal in linking Se to cardiovascular disease risk, although other conditions involving oxidative stress and inflammation have shown some association with Se status. There is growing evidence that higher Se intakes are associated with reduced cancer risk. While persuasive evidence already exists to suggest that additional Se would be beneficial in some health conditions, results from intervention trials underway or planned have the potential to reinforce or refute the argument for increasing Se intake. PMID- 12133203 TI - Nutritional epidemiology of cancer: accomplishments and prospects. AB - Nutritional epidemiology of cancer has gone through several stages. Initially, the long latency of cancer, the difficulties in undertaking long-term cohort investigations or ascertaining remote diet in case-control studies, and the absence of convenient intermediate biomarkers of disease, such as cholesterol in cardiovascular diseases, discouraged studies on diet and cancer. Subsequently, however, epidemiological successes in the chemical, viral and occupational aetiology of cancer, and the increasing insight into the sources of variation of diet and dietary information, prompted investigators to undertake both case control and cohort studies. The results have been mixed. On the one hand, vegetables and fruits have been shown to be inversely associated with several forms of cancer. On the other hand, the information concerning specific macro- or micronutrients in relation to particular forms of cancer has been very limited and mostly inconclusive. There are several reasons for the complexity of investigations of the nutritional epidemiology of cancer and these reasons are briefly considered. An overview of our current understanding of the nutritional causes of cancer is also presented. It is noted that, notwithstanding the substantial gaps in our scientific knowledge, preventive nutritional approaches can be envisaged and they are likely to be moderately successful. PMID- 12133204 TI - Can supplements help meet the micronutrient needs of the developing world? AB - Supplementation has many potential advantages over fortification and dietary approaches for improving micronutrient intake. Pregnant and lactating women and infants are most likely to benefit from supplementation. Recent experience with vitamin A supplementation in young children has proved to be remarkably successful. Demonstrated efficacy of vitamin A supplements for improved child survival in many settings and a technical consensus on how to implement interventions were major factors in achieving this success. Bilateral and UN agencies worked together so that in 1999 80 % of children under 5 years of age in the least-developed countries received a vitamin A capsule in the last 6 months. At least one million child lives saved were associated with the increase in coverage in the last 2 years of the 1990s. Experience with Fe-folate supplements has not been as successful. Whilst a technical consensus has been reached on how to implement programmes to control Fe deficiency, the lack of convincing evidence of efficacy of Fe-folate supplements in terms of maternal and child survival outcomes has undoubtedly contributed to the limited pursuit of effective action. A new multiple micronutrient supplement for use amongst women of reproductive age in developing countries has been formulated. UNICEF is employing the supplement in programmes aimed at helping to prevent low birth weight. The new supplement is likely to be more efficacious than Fe-folate supplements for both maternal and child survival and development outcomes. Successful completion of rigorous efficacy trials will be critical for creating the political support needed to achieve universal coverage. PMID- 12133205 TI - Fortification strategies to meet micronutrient needs: successes and failures. AB - Food fortification is likely to have played an important role in the current nutritional health and well-being of populations in industrialized countries. Starting in the early part of the 20th century, fortification was used to target specific health conditions: goitre with iodized salt; rickets with vitamin D fortified milk; beriberi, pellagra and anaemia with B-vitamins and Fe-enriched cereals; more recently, in the USA, risk of pregnancy affected by neural-tube defects with folic acid-fortified cereals. A relative lack of appropriate centrally-processed food vehicles, less-developed commercial markets and relatively low consumer awareness and demand, means it has taken about another 50 years for fortification to be seen as a viable option for the less-developed countries. The present paper reviews selected fortification initiatives in developing countries to identify different factors that contributed to their successful implementation, as well as the challenges that continually threaten the future of these programmes. Ultimately, the long-term sustainability of fortification programmes is ensured when consumers are willing and able to bear the additional cost of fortified foods. There has been an enormous increase in fortification programmes over the last couple of decades in developing countries. Considerable progress has been made in reducing vitamin A and I deficiencies, although less so with Fe, even as Zn and folic acid deficiencies are emerging as important public health problems. Food fortification based on sound principles and supported by clear policies and regulations can play an increasingly large role in this progress towards prevention and control of micronutrient malnutrition. PMID- 12133206 TI - Food-based strategies to meet the challenges of micronutrient malnutrition in the developing world. AB - The purpose of the present paper is to review the evidence in favour of food based strategies to meet the challenges of micronutrient malnutrition in the developing world. Increasing dietary diversification is the most important factor in providing a wide range of micronutrients, and to achieve this objective in a development context requires an adequate supply, access and consumption of a variety of foods. Diets in developing countries generally lack many nutrients, including energy (inadequate amounts of food), so that strategies need to also emphasize an increase in total food intake, in addition to a greater variety. Agricultural and food policies tend to be oriented to primary agricultural productions, but they could also be formulated to promote and support home gardens and small livestock production for the explicit purpose of increasing the household consumption of micronutrient-rich foods. The adoption of 'desirable' dietary patterns for nutrition improvement, e.g. appropriately formulated to meet micronutrient needs, could be used in the formulation of agricultural policies and programmes. This process could be achieved through support for integrated farming systems oriented to assuring household food security, but also based on a variety of foods that will meet total dietary (including micronutrient) needs. Thus, availability of energy-rich staples, animal and/or fish as major sources of protein, and vitamin-, mineral- and phytonutrient-rich fruit and vegetables could constitute the types of production envisaged. The cultivation of edible indigenous plants as additional sources of micronutrients could also be added. The low bioavailability of some key micronutrients from foods, such as Fe, are substantially enhanced with the right food combinations and with appropriate food processing and preparation techniques. Simple appropriate technology for the preservation of micronutrient-rich foods would need further development and promotion for their year-round availability. Linking community development policies to national programmes for the alleviation of hunger and malnutrition, with an emphasis on increasing the variety of foods consumed, is probably the best strategy for improving micronutrient malnutrition sustainably. PMID- 12133207 TI - Meeting the challenges of micronutrient deficiencies in emergency-affected populations. AB - Micronutrient deficiencies occur frequently in refugee and displaced populations. These deficiency diseases include, in addition to the most common Fe and vitamin A deficiencies, scurvy (vitamin C deficiency), pellagra (niacin and/or tryptophan deficiency) and beriberi (thiamin deficiency), which are not seen frequently in non-emergency-affected populations. The main causes of the outbreaks have been inadequate food rations given to populations dependent on food aid. There is no universal solution to the problem of micronutrient deficiencies, and not all interventions to prevent the deficiency diseases are feasible in every emergency setting. The preferred way of preventing these micronutrient deficiencies would be by securing dietary diversification through the provision of vegetables, fruit and pulses, which may not be a feasible strategy, especially in the initial phase of a relief operation. The one basic emergency strategy has been to include a fortified blended cereal in the ration of all food-aid-dependent populations (United Nations High Commissioner for Refugees/World Food Programme, 1997). In situations where the emergency-affected population has access to markets, recommendations have been to increase the general ration to encourage the sale and/or barter of a portion of the ration in exchange for locally-available fruit and vegetables (World Health Organization, 1999a,b, 2000). Promotion of home gardens as well as promotion of local trading are recommended longer-term options aiming at the self-sufficiency of emergency-affected households. The provision of fortified blended foods in the general ration has successfully prevented and controlled micronutrient deficiencies in various emergency settings. However, the strategy of relying only on fortified blended foods to prevent micronutrient deficiencies should be reviewed in the light of recurring evidence that provision of adequate supplies of these foods is often problematic. Donor policies on the bartering or exchange of food aid should also be clarified. Furthermore, the establishment of micronutrient surveillance systems, including standardized micronutrient deficiency diagnostic criteria, are vital for the control of micronutrient deficiency diseases. PMID- 12133208 TI - Developmental issues in attitudes to food and diet. AB - As a rule, children and most adults eat what they like and leave the rest. They like and consume foods high in fat and sugar. Parental behaviour shapes food acceptance, and early exposure to fruit and vegetables or to foods high in energy, sugar and fat is related to children's liking for, and consumption of, these foods. Some parents are imposing child-feeding practices that control what and how much children eat. However, over-control can be counter-productive, teaching children to dislike the very foods we want them to consume, and generally undermining self-regulation abilities. The external environment is also important, with concerns expressed about food advertising to children and girls dieting for an ideal thin body shape. Up to one-quarter of young adolescent girls report dieting to lose weight, their motivation driven by weight and shape dissatisfaction. For some, dieting and vegetarianism are intertwined and both legitimised as healthy eating. For others, striving for nutritional autonomy, the choice of less-healthy foods is not just because of their taste, but an act of parental defiance and peer solidarity. The determinants of what children choose to eat are complex, and the balance changes as children get older. A better understanding is crucial to informing how we might modify nutritional behaviour. Adults occupy a central position in this process, suggesting that children should be neither the only focus of nutritional interventions nor expected to solve the nutritional problems with which adults around them are continuing to fail. PMID- 12133209 TI - Resistance to changes in diet. AB - Dietary changes can be difficult to effect both at an individual and at a population level, and even when changes do occur they are often far slower and less pronounced than might be expected. Three possible reasons for this situation will be considered: the complexity of food choice and competing influences, attitudinal ambivalence and optimistic bias. Food choice is influenced by a large number of factors, not only health considerations, and therefore it is not surprising that interventions based primarily on health concerns have been ineffective. Another concern is that people do not always have clear-cut attitudes, but rather can be ambivalent about foods and about healthy eating, and this factor might impact on the translation of beliefs and attitudes into behaviour. A third possible reason is optimistic bias, where individuals believe themselves to be at less risk from various hazards than is the average person. This effect has been demonstrated for nutritional risks, and this factor might lead people to take less note of health education messages. The stages-of-change model from health psychology has been proposed as a method for improving the effectiveness of behaviour change interventions. However, there are a number of problems in transferring such a model from smoking, where it was originally developed, to dietary behaviours, including the lack of clear-cut specific behaviours and behaviour change targets in the dietary field. PMID- 12133210 TI - The production of food: from quantity to quality. AB - The present paper presents a non-technical overview of contemporary developments in food supply, as seen from the standpoint of economic adjustment. The historical concerns over availability and price of food have now passed in the UK, and agriculture is no longer dominantly driven by the supply-side forces of new farming technology and the stimulus of support policies. As a now demand driven sector of the economy, it is the developing diversity of consumer food preferences that will increasingly determine the adjustment path of agricultural production. Those demands seek distinctive elements of food value, many of which are entirely created and delivered by industries beyond the farm gate. However, many of the quality characteristics of food that consumers increasingly seek are associated explicitly with what takes place on farms and how crop and livestock husbandry is conducted. In responding to these demand preferences many farmers will shift from being merely raw material producers to becoming genuine producers of food, or capturing more of the final value of the products consumed. As a result a dual structure within farming will develop, with a 'quality agriculture' becoming increasingly differentiated from a 'commodity agriculture' as two distinct strategies for farm business survival. PMID- 12133211 TI - Changes in fat synthesis influenced by dietary macronutrient content. AB - DE NOVO: lipogenesis is the biological process by which C2 precursors of acetyl CoA are synthesized into fatty acids. In human subjects consuming diets higher in fat (> 30 % energy), lipogenesis is down regulated and extremely low; typically < 10 % of the fatty acids secreted by the liver. This percentage will increase when dietary fat is reduced and replaced by carbohydrate, although the extent of carbohydrate-induced lipogenesis is dependent on the type of carbohydrate (monosaccharide v. polysaccharide) and the form in which the carbohydrate is fed (liquid meals, solid less-processed food). Clearly, massive overconsumption of carbohydrate can also increase lipogenesis. A second related phenomenon that occurs when dietary fat is reduced is hypertriacylglycerolaemia. This rise in blood triacylglycerol concentration could be due to increased de novo lipogenesis or to reduced clearance of lipid from the blood. The present paper will review the metabolic mechanisms leading to the elevations in blood triacylglycerol concentration that occur with dietary fat reduction. Studies considered will be those investigating fatty acid synthesis in subjects chronically fed low-fat high carbohydrate diets and studies in which data were obtained in both the fasted and fed states. Also summarized will be data from subjects who had consumed diets of different carbohydrate types, as well as the most recent data from postprandial studies investigating factors that affect the magnitude of the rise in blood lipids following a meal. Given the changing availability of carbohydrate in the food supply, it will be important to understand how the balance of fat and carbohydrate in the diet influences lipogenesis, and the relative contribution of the process of de novo lipogenesis to the escalating incidence of obesity observed around the world. PMID- 12133212 TI - Dietary fat: assessing the evidence in support of a moderate-fat diet; the benchmark based on lipoprotein metabolism. AB - There is a growing database that has evaluated the effects of varying amounts of total fat on risk factors for cardiovascular disease, diabetes and overweight and obesity. The evidence clearly suggests that extremes in dietary fat should be avoided, and instead a diet moderate in total fat (25-35 % energy) is preferable for the majority of individuals. Moreover, we now appreciate the importance of individualizing dietary fat recommendations within this range of total fat. With respect to cardiovascular disease, a diet higher in total fat (30-35 % energy) affects the lipid and lipoprotein risk profile more favourably than a lower-fat diet; this is also the case for individuals with diabetes, with the added benefit of better glycaemic control. Dietary fibre (> or = 25 g/d) attenuates and even prevents the potentially adverse lipid and lipoprotein effects of a lower-fat diet. With respect to weight control, a moderate-fat diet can be as, or even more, effective than a lower-fat diet, because of advantages with long-term adherence and potentially favourable effects on lipids and lipoproteins. Thus, there is now a convincing scientific basis to advocate a diet moderate in total fat for the majority of individuals. Implicit to this position is that unsaturated fat has numerous beneficial health effects. However, because fat is energy dense, moderation in fat intake is essential for weight control. Consequently, a simple message to convey is to avoid diets that are very low and very high in fat. Moreover, within the range of a moderate-fat diet it is still important to individualize the total fat prescription. Nonetheless, the guiding principle is that moderation in total fat is the defining benchmark for a contemporary diet that reduces risk of chronic disease. PMID- 12133213 TI - Low-fat diets and energy balance: how does the evidence stand in 2002? AB - The role of high-fat diets in weight gain and obesity is assessed by evidence based principles. Four meta-analyses of weight change occurring on ad libitum low fat diets in intervention trials consistently demonstrate a highly significant weight loss of 3-4 kg in normal-weight and overweight subjects (P < 0.001). The analyses also find a dose-response relationship, i.e. the reduction in percentage energy as fat is positively associated with weight loss. Weight loss is also positively related to initial weight; a 10 % reduction in dietary fat is predicted to produce a 4-5 kg weight loss in an individual with a BMI of 30 kg/m2. The non-fat macronutrient composition of the diet is also important. Whereas the glycaemic index of the carbohydrate may play a role for cardiovascular risk factors, there is so far no evidence that low-glycaemic index foods facilitate weight control. In contrast, intervention studies show that sugar in drinks is more likely to produce weight gain than solid sugar in foods. Although the evidence is weak, alcoholic beverages promote a positive energy balance, and wine may be more obesity-promoting than beer. Protein is more satiating and thermogenic than carbohydrates, and one intervention study has shown that an ad libitum low-fat diet where carbohydrate was replaced by protein produced more weight loss after 6 months (8.1 v. 5.9 kg). The evidence linking particular fatty acids to body fatness is weak. If anything, monounsaturated fat may be more fattening than polyunsaturated and saturated fats, and no ad libitum dietary intervention study has shown that a normal-fat high-monounsaturated fatty acid diet is equivalent or superior to a low-fat diet in the prevention of weight gain and obesity. The evidence strongly supports the low-fat diet as the optimal choice for the prevention of weight gain and obesity, while the use of a normal fat high-monounsaturated fatty acid diet is unsubstantiated. PMID- 12133214 TI - Nutritional improvement of plant foods by non-thermal processing. AB - As a result of the increasing consumer demand for minimally-processed fresh-like food products with high sensory and nutritional qualities, there is a growing interest in non-thermal processes for food processing and preservation. Key advanced technologies such as high-pressure processing, pulsed electric fields, dense gases and ultrasound are being applied to develop gentle but targeted processes to further improve the quality and safety of processed foods. These technologies also offer the potential for improving existing processes as well as for developing new process options. Furthermore, by adding new process dimensions (such as hydrostatic pressure, electric fields, ultrasonics, supercritical CO2) to the conventional process variables of temperature and time, they facilitate enlargement of the availability of unit operations. These operations might be applied effectively in unique combination processes, or as subsequent processing tools in more-targeted and subsequently less-intensive processes for food preservation and modification than the currently-applied processes. PMID- 12133219 TI - Editorial. PMID- 12133220 TI - Henri Tajfel's 'cognitive aspects of prejudice' and the psychology of bigotry. AB - This paper pays tribute to Tajfel's classic article 'Cognitive aspects of prejudice' and re-examines its central arguments. Tajfel's paper is important for outlining a social cognitive approach to the study of prejudice and also for refuting of what Tajfel called the 'blood-and-guts' approach. Taking Tajfel's proposition that social psychology is not value-free, the current paper examines the moral and political view of 'Cognitive aspects' and also the gaps in its approach to the study of prejudice. It is suggested that this cognitive approach has difficulty in accounting for extreme bigotry, at least without recourse to the motivational themes that the approach seeks to exclude. In particular, there would be limitations in applying this approach in order to understand the Holocaust. Indeed, Tajfel did not attempt to do so, for reasons that are discussed. Tajfel's Social Identity Theory (SIT) has similar limitations. The paper also examines Tajfel's use of the term 'depersonalization', which he described as a 'milder' form of dehumanization of out-groups. Later social identity theorists have tended to use 'depersonalization' differently, shifting their attention to in-groups. Their perspective moves away from understanding the topic of prejudice in the way that can be found in Tajfel's 'Cognitive aspects of prejudice'. Finally, the present paper suggests how extreme prejudice might be studied without returning to the motivational 'blood-and-guts' approach that Tajfel so cogently criticized. PMID- 12133221 TI - Enjoyment, bigotry, discourse and cognition. PMID- 12133224 TI - The effects of status on subgroup relations. AB - Two experiments were conducted to assess the impact of status differentials on subgroup attitudes and behaviours. In Experiment 1, 73 math-science students were led to believe they had higher or lower status than humanities students. They then performed a non-interactive decision-making task during which they were categorized exclusively as a university student (superordinate condition), or as a university student and math-science student simultaneously (subgroups condition). Experiment 2 (N = 98) differed from Experiment 1 in that perceptions of relative subgroup status were measured rather than manipulated. Consistent with social identity theory, subgroup members tended to categorize themselves more at the superordinate (university) level the lower status they considered their subgroup to be. In Experiment 2, a series of interactions also emerged, showing that status and inter-subgroup bias were positively related when the participants had been categorized exclusively at the superordinate level. When superordinate and subgroup identities were activated simultaneously, perceptions of status had no effect on levels of bias. The results were interpreted in terms of participants' needs for identity enhancement and identity distinctiveness. PMID- 12133225 TI - Attitudes and intentions of homeless people towards service provision in South Wales. AB - The theory of planned behaviour (TPB: Ajzen, 1988, 1991) was used as a framework to investigate homeless people's participation in outreach service programmes. In total, 104 homeless people from South Wales were interviewed using a schedule based on the TPB. Congruent with previous research on the TPB, attitude was the dominant predictor of behavioural intentions, and intention and perceived behavioural control were predictive of behaviour. Contrary to predictions, subjective norms also exerted a direct effect on behaviour. The discussion focuses on two issues: first, the utility of social cognition models in explaining the relationship between demographic variables and behaviour in homelessness research; second, the direct effects of norms on behaviour and the extent to which work on social groups might usefully extend research on models such as the TPB to aid understanding of behaviour amongst stigmatized populations. PMID- 12133226 TI - Organizational identification after a merger: a social identity perspective. AB - An analysis of the social identity processes involved in organizational mergers suggests that organizational identification after a merger is contingent on a sense of continuity of identity. This sense of continuity, in turn, is argued to be contingent on the extent to which the individual's own pre-merger organization dominates, or is dominated by, the merger partner. In support of this analysis, results of two surveys of merged organizations showed that pre-merger and post merger identification were more positively related for members of dominant as opposed to dominated organizations, whereas perceived differences between the merger partners were more negatively related to post-merger identification for members of the dominated compared with the dominant organization. PMID- 12133227 TI - Evidence that the type of person affects the strength of the perceived behavioural control-intention relationship. AB - This study examined the role of person type in explaining the relationship between perceived behavioural control and behavioural intentions. Participants (N = 187) completed measures of the theory of planned behaviour (Ajzen, 1991) variables regarding 30 behaviours. Within-participants analyses demonstrated that intentions were more strongly predicted by perceived behavioural control (PBC) than a combination of attitudes and subjective norms among a minority of the sample. When these 'PBC controlled' participants were considered separately, the effects for perceived behavioural control obtained in previous between participants analyses were augmented. Conversely, when these participants were excluded from the sample, the effects of perceived behavioural control were reduced. PBC control was also modestly associated with dispositional measures of perceived controllability. Overall, the findings indicate that the strength of the perceived behavioural control-intention relationship depends not only on the type of behaviour but also on the type of person. PMID- 12133228 TI - General threat leading to defensive reactions: a field experiment on linguistic features. AB - Supported by several theoretical perspectives on motivation, we based an experiment on the idea that threat motivates people to become defensive and to choose that which is familiar and unequivocal in a given situation. The present field experiment confirmed that a general relevant threat can motivate people in a linguistic multiculture to conform more rigidly to their own language, and hence accentuate their own linguistic singularity. In addition, an exploratory analysis of tolerance towards the competing language revealed that non-threatened people tended to open up toward the other language. A dual-motive model that accounts for opening-up versus defensive reactions is proposed. PMID- 12133229 TI - Changing identity: predicting adjustment to organizational restructure as a function of subgroup and superordinate identification. AB - We investigated a work-team restructure within an organization obtaining measures before and after the change occurred. Pre-restructure analyses revealed that, in addition to informational variables, subgroup identification (work-team) and superordinate identification (organization) were important predictors of negative feelings towards the restructure. The more that employees identified with the subgroup, the more negative feelings they reported about the upcoming change. In contrast, the higher the identification with the superordinate group, the less negative employees felt. Longitudinal analysis revealed that compared with the pre-restructure, post-restructure levels of work-team identification, organizational identification, job satisfaction and perceived work-team performance were significantly lower. Pre-restructure work-team identification was a stronger predictor of post-restructure job satisfaction than pre restructure organizational identification. In addition, it was found that pre restructure work-team identification and organizational identification had opposing effects on post-restructure organizational identification. There was some evidence that high initial organizational identification protected long-term organizational commitment. PMID- 12133230 TI - Talking about transplants: social representations and the dialectical, dilemmatic nature of organ donation and transplantation. AB - In many westernized countries, organ donation rates are low in comparison with the need for life-saving organ transplants, and are at odds with generally high community endorsement of organ donation. This is particularly true for Western Australia, the location of this study. This contradiction between endorsement and donation is investigated within a framework that draws from Moscovici's (1984) theory of Social Representations, Guimelli's (1998) differentiation between normative and functional dimensions of the central core, and Billig's (1988) rhetorical position on the role of argumentation in discourse. Four focus group discussions on organ donation and transplantation were conducted. Analysis of the discourse suggests that the social representation of organ donation and transplantation can be understood best as a representational field organized around two dialectically 'opposed' images-the gift of life and the mechanistic removal and replacement of body parts. The normative and functional expression of these two images as a pro-donation stance and a qualified pro-donation stance is discussed, as is the role of argumentation in the production of a social representation. PMID- 12133231 TI - The temporal relationship between bacterial lipopolysaccharide and monocrotaline exposures influences toxicity: shift in response from hepatotoxicity to nitric oxide-dependent lethality. AB - Liver injury from a variety of hepatotoxicants, including the food-borne phytotoxin monocrotaline (MCT), can be augmented by exposure to a noninjurious dose of the inflammagen bacterial lipopolysaccharide (LPS). In a previous study, a nontoxic dose of LPS given 4 h after MCT resulted in synergistic hepatotoxicity within 12-18 h. This study was designed to determine whether temporal differences in MCT and LPS exposure affect toxicity. When LPS (3.4 x 10(6) EU/kg; iv) was given one hour before MCT (100 mg/kg; ip), hepatotoxicity developed between 4 and 8 h after MCT administration, and mortality was much greater than when LPS was administered 4 h after MCT. To explore this difference, the temporal relationship between LPS and MCT exposure (7.4 x 10(6) EU/kg and 100 mg/kg, respectively) was altered. Twenty-four-hour survival was high in animals that received LPS 4 h before (86%) or after (88%) MCT, but it decreased markedly when LPS was administered 1 h before MCT (17%). Using this latter dosing regimen, animals became moribund as early as 4 h after MCT administration. Since liver injury was similar from regimens that differed greatly in mortality, death appeared to result from extrahepatic causes. To explore a role for nitric oxide (NO)-induced shock in this regimen, animals were treated with aminoguanidine (AG), an inhibitor of inducible NO synthase, prior to administration of LPS given an hour before MCT. In the cotreated animals, AG significantly attenuated mortality and decreased plasma nitrate/nitrite concentrations, markers of NO biosynthesis. Hence, the primary target of toxicity from MCT and LPS cotreatment appeared to shift from the liver to an extrahepatic site or sites as exposure to these agents occurred closer together temporally. NO appears to be causally involved in the deaths of animals treated with LPS 1 h before MCT. PMID- 12133232 TI - Endotoxin potentiates the hepatotoxicity of cocaine in male mice. AB - Cocaine produces hepatotoxicity by a mechanism that remains undefined but that has been linked to its oxidative metabolism. Endotoxin (lipopolysaccharide, LPS) is also a well-known cause of hepatic damage, where exposure to non-injurious doses of LPS increases the toxicity of certain hepatotoxins. This study was conducted to investigate the possible potentiation of cocaine-mediated hepatotoxicity (CMH) by LPS. Male CF-1 mice were administered oral cocaine hydrochloride for 5 consecutive days at a dose of 20 mg/kg with and without 12 x 10(6) EU LPS/kg given intraperitoneally 4 h after the last cocaine injection. Serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were measured as markers of liver injury. Blood and liver glutathione (GSH) levels were determined, as well as the activities of glutathione peroxidase (GPx) and catalase (CAT). In addition, the activity of liver glutathione reductase (GRx) was measured. The results demonstrate that endotoxin potentiated the hepatotoxicity of cocaine. Serum ALT and AST were significantly elevated with the combined cocaine and LPS treatment versus all other treatments. While cocaine alone resulted in centrilobular necrosis, the cocaine and LPS combination produced submassive necrosis. The increased hepatic GSH content and GRx activity observed with cocaine alone were not observed with the combination treatment, rendering the liver more susceptible to oxidative stress. Moreover, there was a significant decrease in the activities of hepatic GPx and CAT, particularly with the combination treatment. In conclusion, this study demonstrates that LPS potentiates the hepatotoxicity of cocaine as revealed by an array of biochemical and morphological markers. PMID- 12133233 TI - Decrease in ovalbumin-induced pulmonary allergic response by benzaldehyde but not acetaldehyde exposure in a Guinea pig model. AB - The pulmonary effects of two environmentally relevant aldehydes were investigated in nonsensitized or ovalbumin (OA)-sensitized guineapigs (GPs). Four-week-old male Hartley GPs, weighing about 400 g, were intraperitoneally injected with 1 ml of an NaCl solution containing 100 microg OA and 100 mg Al(OH)(3). They were then exposed to either acetaldehyde (200 ppb) or benzaldehyde (500 ppb) for 4 wk (6 h/d, 5 d/wk). At the end of exposure, GPs were challenged with an OA aerosol (0.1% in NaCl) and pulmonary functions were measured. The day after, guinea pigs were anesthetized and several endpoints related to inflammatory and allergic responses were assessed in blood, whole-lung histology, and bronchoalveolar lavage (BAL). Sensitized nonexposed GPs showed bronchial hyperresponsiveness to OA and an increased number of eosinophils in blood and BAL, together with a rise in total protein and leukotrienes (LTB(4) and LTC(4)/D(4)/E(4)) in BAL. In nonsensitized GPs, exposure to acetaldehyde or benzaldehyde did not induce any change in the tested parameters, with the exception of irritation of the respiratory tract as detected by histology and an increased number of alveolar macrophages in animals exposed to acetaldehyde. In sensitized GPs, exposure to acetaldehyde induced a moderate irritation of the respiratory tract but no change in biological parameters linked to the inflammatory and allergic responses. In contrast, exposure to benzaldehyde induced a decrease both in OA-induced bronchoconstriction and in eosinophil and neutrophil numbers in BAL, an increase in the bronchodilatator mediator prostaglandin E(2) (PGE(2)), and a decrease in the bronchoconstrictor mediators LTC(4)/D(4)/E(4). Further investigations are needed to determine if the attenuated response observed in sensitized GPs exposed to benzaldehyde is due to an alteration of the mechanism of sensitization or to a more direct effect on various mechanisms of the allergic response. PMID- 12133234 TI - Tributyltin induces human neutrophil apoptosis and selective degradation of cytoskeletal proteins by caspases. AB - Tributyltin (TBT) has frequently been used as a pesticide and in biocidal paints for marine vessels, leading to its presence in the environment. Although TBT was recently found to induce apoptosis in different immune cells, by a mechanism that is not fully established, its effect on neutrophils is not known. In this study, it was found that TBT induced apoptosis in human neutrophils as assessed by cytology, flow cytometry, and degradation of the microfilament-associated protein gelsolin. Furthermore, data showed that TBT induced neutrophil apoptosis by a caspase-dependent mechanism, since addition of z-Val-Ala-Asp(MOe)-CH(2)F (z-VAD FMK) in the culture prevented the effect of TBT. It was also found that the cytoskeletal proteins gelsolin, paxillin, and vimentin, but not vinculin, were degraded by TBT via caspases, as assessed by immunoblotting. Data indicate that gelsolin, paxillin, and vimentin are three caspase substrates involved in both spontaneous and TBT-induced neutrophil apoptosis. Cells were not necrotic as assessed by trypan blue dye exclusion, and this is in agreement with the absence of vinculin degradation. Evidence indicates that TBT-induced fragmentation of cytoskeletal proteins via caspases is a process that is tightly regulated. PMID- 12133235 TI - Development of a physiologically based pharmacokinetic model for methyl ethyl ketone in F344 rats. AB - A physiologically based pharmacokinetic (PBPK) model to describe the absorption, distribution, metabolism, and elimination of methyl ethyl ketone (MEK) in rats was developed. Partition coefficients were experimentally determined in rat tissues and blood samples using an in vitro vial equilibration technique. These solubility ratios were in agreement with previous human-based estimates that MEK is uniformly soluble within all tissues. The in vivo metabolism of MEK was evaluated using groups of three F344 male rats exposed to 100-2000 ppm MEK in a closed, recirculating gas uptake system. An optimal fit of a family of uptake curves was obtained by adjusting Michaelis-Menten metabolic constants, Km (affinity), and Vmax (capacity) using the PBPK model. At the highest chamber concentration, the uptake curve could not be modeled without the addition of a first-order (Kfo) metabolic pathway. Pretreatment with pyrazole, an inhibitor of oxidative microsomal metabolism, decreased the slope of the gas uptake curve but did not abolish metabolism. Optimal model fit to the gas uptake curve from pyrazole-pretreated animals required the apparent Km to be increased roughly 50 times the value determined in naive rats. The completed PBPK model was evaluated against real-time exhaled breath data collected from rats receiving an intravenous (iv) injection of MEK via a jugular vein cannula. Model simulation of the iv-treated animals required alveolar ventilation to be reduced 30% in order to match the data. Exhaled breath profiles from animals treated with MEK by oral gavage or intraperitoneal (ip) injection were evaluated and absorption rates was determined. Development of a comprehensive PBPK model for MEK in rats is the first step toward future extrapolations to apply to humans. PMID- 12133236 TI - Comparison of models for the estimation of biological partition coefficients. AB - Several models have been published for calculating blood-air, tissue-air, or tissue-blood partition coefficients of volatile organic chemicals in human or rat tissues, from functions of their octanol-water partition coefficients or solubilities in vegetable oil and water. In this work, the relative accuracy, strengths, and limitations of the various models are examined. Comparison of predicted human tissue-air and tissue-blood partition coefficients with experimental values has been made for 12 chemicals, covering a wide range of lipophilicity (acetone, isopropanol, diethylether, methylene dichloride, benzene, toluene, trichloroethylene, trichloroethane, n-pentane, cyclohexane, n-hexane, and n-heptane). Seven published models for human tissue-air and 10 models for tissue-blood partition coefficients have been compared. Fewer models are available for predicting rat tissue-air and rat tissue-blood partition coefficients, but a similar comparison has been made. The ratio of predicted to experimental partition coefficients and their mean, R(mean), and the mean magnitude of the difference between predicted and experimental values of log(10) P, E, were used to assess the accuracy of each model. For the test set the most accurate for human blood-air partition coefficients were the empirical equations of Meulenberg and Vijverberg (R(mean) = 1.1 +/- 0.46, E = 0.156) and the empirical solvation equation of Abraham and Weathersby (1994) (R(mean) = 0.93 +/- 0.38, E = 0.166). For rat blood, predictions are much less accurate due to difficulties in modeling the effects of protein binding, which are much larger. Overall, for rat blood-air partition coefficients the equation of Meulenberg and Vijverberg (1999) (R(mean) = 0.74 +/- 0.50, E = 0.236) was the most accurate. The tissue-composition-based equations of Poulin and Krishnan, using solubilities in vegetable oil, performed well for human liver-air partition coefficients (R(mean) = 1.21 +/- 0.28, E = 0.079) for log(octanol-water partition coefficients) > 0.7 and for fat-air partition coefficients, but overestimated solubilities in human kidney and brain tissues (e.g., for kidney tissue, R = 1.88 +/- 0.58, E = 0.255). The equations of Meulenberg and Vijverberg (2000a), Abraham and Weathersby (1994), and Paterson and Mackay (1989) also performed moderately well for human tissue-air partition coefficients. For rat muscle-air, liver-air, and fat-air partition coefficients the model of Poulin and Krishnan (1996a) gave the most accurate predictions. For tissue-blood partition coefficients, generally good agreement with experimental values is obtained by the empirical model of Balaz and Lukacova (1999) (e.g., for human kidney, R(mean) = 1.15 +/- 0.38, E = 0.085) and, if solubility in fat is known, by the equations of Fiserova-Bergerova and Diaz (1986) (e.g., for human muscle, R(mean) = 1.10 +/- 0.39, E = 0.107). The equations of DeJongh et al. (1997) gave the most accurate predictions for rat muscle-blood, liver-blood and fat-blood partition coefficients (e.g., for rat muscle R(mean) = 1.03 +/- 0.39, E = 0.149), but predictions were less accurate than for human tissue-blood partition coefficients, attributable to difficulties in modeling the effect of protein binding. The choice of equation for use in physiologically based pharmacokinetic (PBPK) models depends on the species, tissue, and chemical lipophilicity. PMID- 12133238 TI - Effects of bisphenol A and artificial UVB radiation on the early development of Rana temporaria. AB - Ultraviolet-B (UVB) radiation is one environmental factor that may act in concert with other stressors, such as xenobiotics, to produce adverse effect on amphibian populations. Embryos (< 24 h old) of the common frog (Rana temporaria) were exposed to four concentrations (0, 10, 100, or 1000 microg/L) of bisphenol A (BPA), with and without ultraviolet-B (UVB) radiation, for 20 d in the laboratory. Throughout the experiment the biologically effective daily UVB dose, calculated with Setlow's DNA action spectrum, was 2.80 kJ m(-2). UVB radiation as such did not have any significant effect on frog embryos. However, a BPA treatment of 1000 microg/L had a significant effect on embryos in both the UVB and no-UVB treatments, with the effect being greater with UVB. UVB produced a significant decrease in survival in the newly hatched frog larvae at all BPA concentrations. These results demonstrate that simultaneous exposure to these two stress factors is more harmful to R. temporaria larvae than exposure to one stressor alone. PMID- 12133239 TI - Hope and hubris. PMID- 12133237 TI - In vivo effects of bisphenol A on the polecat (mustela putorius). AB - Bisphenol A (BPA), an environmental estrogen derived from the plastic industry, was given orally via incorporation into the food of 30 male and female polecats at 3 different doses (10, 50, or 250 mg/kg body weight/day) for 2 wk with 10 animals acting as controls. Several hormone levels in the plasma were determined as well as the activities of the phase I and II biotransformation enzymes 7 ethoxyresorufin O-deethylase (EROD), cytosolic glutathione S-transferase (GST), and UDP-glucuronosyltransferase (UDPGT). BPA did not cause any macroscopic effects on body mass or the health of the animals. UDPGT and GST activities increased significantly in direct correlation with increasing BPA exposure in females and UDPGT increased in a dose-related manner in males. There was no change in the plasma T4 and testosterone concentrations of the males with increasing BPA exposure. Discriminant analysis indicated that the group receiving 10 mg BPA/kg body weight/d was not different from the control group but the groups receiving 50 and 250 mg/kg body weight/d were different from the control group. This suggests physiological changes in the groups receiving 50 or 250 mg BPA/kg body weight/d. PMID- 12133240 TI - Assessing the outcome of a palliative care educational initiative. AB - PURPOSE: In 1997, a biweekly, 18-month Palliative Care Seminar Series was started at Mount Sinai School of Medicine as a way to explore participants' beliefs about palliative care and to instill the knowledge, skills, and attitudes needed to improve care at the end of life. The goal of this study was to examine the effects of Mount Sinai's Palliative Care Seminar Series on faculty development and personal growth. METHODS: Twenty of 42 respondents were interviewed immediately after completing the Seminar Series. Subjects identified themselves as physicians (n = 16) or nurses (n = 4). The same open-ended questions and follow-up probes were asked of each interviewee and transcripts were coded for confidentiality. The questions were developed to examine the impact of the Seminar Series on practitioners' knowledge of and attitude toward end-of-life care. Content analysis of the coded transcripts was performed by a group consisting of one physician, one medical student, and two social scientists. RESULTS: Content analysis of interview transcripts revealed five common themes: subjects perceived a gain in palliative care knowledge and skills; they believed themselves to be more confident in the practice of palliative care; they believed themselves to be more confident that what they were doing is appropriate; and they felt less isolated in their beliefs as a result of regular interactions with supportive peers. CONCLUSIONS: Five common themes arose from the transcripts of both physicians and nurses: participants perceived a gain in palliative care "practice" skills, participants perceived a gain in palliative care "process" skills, participants believed themselves to be more confident that palliative care was appropriate care for dying patients, participants believed themselves to be more confident in their ability to practice and teach palliative care, and participants felt less isolated in their beliefs through regular interactions with supportive peers. PMID- 12133241 TI - A pilot survey of medical students' perspectives on their educational exposure to palliative care in two Canadian universities. AB - BACKGROUND: Graduating medical students from the class of 1999 from McGill University and the University of Alberta completed a self-administered, anonymous, pilot survey to determine students' perspectives on how their educational experience in common palliative care topics contrasted with their educational experience in the diagnosis and management of hypertension, non palliative aspects of breast cancer, and patients dying of acquired immune deficiency syndrome (AIDS). METHODS: A Likert scale ranging from "excellent," scored 1, "very poor," scored 5, was used. Students also estimated the number of hours they spent, during their training, in operating rooms, on home visits to terminally ill patients, and in interprofessional teaching. RESULTS: Sixty of 114 (53%) students from McGill University, and 53 of 110 (48%) students from the University of Alberta responded to the survey. The mean ratings of education experience in the various topics for both universities combined were as follows: hypertension, 2.03; breast cancer, 2.33; cancer pain, 3.42; communicating with dying patients, 3.32; and caring for patients with AIDS, 4.15. The average number of hours spent in the operating room, on home visits to terminally patients, and in interprofessional teaching for both universities combined were 155 hours, 4.2 hours, and 16 hours, respectively. Of the responding students from both universities 83% favored increased palliative care teaching. CONCLUSION: Despite the disproportionate number of hours spent in operating rooms compared to palliative care community exposure, only two students, one from each university, favored shortening surgical rotations to allow for increased time for palliative care education. Recommendations, including increasing palliative care education during clinical clerkships, are provided to improve medical students' perceptions of their educational experiences in palliative care education. PMID- 12133242 TI - A one-day, hospital-wide survey of dying inpatients. AB - We attempted to identify and briefly follow until discharge all terminally ill patients in a large general hospital. On 1 day, nurse case managers reviewed all hospitalized patients and identified those whom they believed were likely to die in the next 6 months (Category A) or whom might be considered terminally ill but with a longer prognosis (Category B). Twelve percent of all adult and pediatric medical-surgical inpatients were detected, equally divided between the two categories. In Category A, 63% were on the medical service, 7% were receiving intensive care, 54% had cancer, and 46% had do-not-resuscitate (DNR) orders. In Category B, 40% were on the medical service, 10% were in intensive care, 52% had cancer, and only 5% had DNR orders. Case managers expected 6% of identified patients to die in the hospital. After 1 month, at least 19% of identified patients had died (2.3% of the medical-surgical inpatient census on the day of the survey). The average length of stay in both categories, excluding outliers, was 24 days or approximately 4 times the average length of stay for the hospital. Patients who actually died in the hospital had an average length of stay of 62 days. This study presents a simple method for estimating the number of dying patients in a hospital--the target population for a palliative care program--and for determining their location, principal diagnosis, length of stay, and disposition. We present information indicating that the survey underestimates the number of dying hospitalized patients. We discuss possible policy implication of this study, primarily that general hospitals should consider developing specialized palliative care services for this substantial group of inpatients. PMID- 12133243 TI - Preferences for palliative sedation therapy in the Japanese general population. AB - OBJECTIVES: To elucidate which types of palliative sedation therapy are preferred by the Japanese general population, what factors influence these preferences, and how the general population thinks clinicians should inform patients about sedation therapy. METHODS: A cross-sectional questionnaire survey using a convenient sample of 457 Japanese people (effective response rate, 53.2%). RESULTS: For refractory intractable physical distress, intermittent deep sedation was chosen as "probably want" or "strongly want" by 86% of the respondents, and mild sedation was chosen by 82%. For refractory intractable psychological distress, intermittent deep sedation was chosen as "probably want" or "strongly want" by 76%, and mild sedation was chosen by 68%. Continuous deep sedation was chosen as "absolutely not want" or "probably not want" by 72% for physical distress and 71% for psychological distress. The respondents who did not want continuous deep sedation were significantly younger, more educated, and more likely to perceive the importance of dignity and preparation for death. Eighty five percent wanted clear information about reduction in consciousness, and 92% were positive with "in advance" information about sedation therapy. When family members did not agree with the patient's decision, 71% stated that physicians should follow the patient's wishes. CONCLUSIONS: The Japanese general population preferred intermittent deep or mild sedation to continuous deep sedation in alleviation of intractable and refractory distress. Many required explicit information about the serious consequences of sedation and wanted physicians to respect their wishes. We recommend that clinicians recognize the importance of both symptom alleviation and maintenance of intellectual activities, and to facilitate direct patient involvement in the decision-making process. PMID- 12133244 TI - Are hospices ready to participate in palliative care research? Results of a national survey. AB - BACKGROUND: Improvements in end-of-life care will require an active program of research, and this research will need to involve patients in hospice. However, it is not known whether hospices are prepared to participate in research, nor is it known what barriers may prevent hospices from becoming involved in research. METHODS: This nationwide telephone survey was conducted with a random sample of hospice organizations taken from a national database. Questions elicited the research activities in which hospices are involved, the resources that hospices have available for the ethical review of research, and perceived barriers to research participation. RESULTS: Of 88 hospices identified, 17 (19%) reported that they had participated in research in the past year. Hospices that participated in research were more likely to be urban, affiliated with an academic institution, and were more likely to have an inpatient unit. Hospices cited several barriers to research participation, including time commitments, staffing resources, ethical concerns, and burdens to patients and families. The most important concern was lack of staffing resources. Hospices indicated that they would be most willing to support research regarding pain management and timing of referral to hospice. PMID- 12133245 TI - Evolution in measuring the quality of dying. AB - PURPOSE: Despite multiple efforts to improve the experience for dying patients, researchers still struggle to identify appropriate outcome measures that assess patients' and families' experiences. If health care systems are to provide excellent, compassionate care to dying patients and their families, there must be a valid means of assessing the quality of those experiences and interventions to improve care. The purpose of this paper is to evaluate quality-of-life instruments currently used to assess the experiences of dying patients, and to offer a design for a next generation instrument to measure quality at the end of life. DESIGN: Sources were attained through a review of the quality of life, quality of dying, and end-of-life care literatures. The terms quality of life, quality of care, terminal care, hospice, assessment, and measurement were used singly and in combination in the MEDLINE database from 1966 to 2001. DISCUSSION: An appropriate clinical quality of dying instrument must be derived from the perspectives of end-of-life care participants and include the multiple domains of experience important to patients and families. Because dying patients are often too ill to communicate, nonresponse bias is a major problem in this population. Researchers must identify additional objective and subjective measures that clearly reflect, correspond well (or predictably) with, and serve as alternatives to patients' self-ratings. Additionally, an appropriate assessment tool must accommodate individual definitions of the quality of dying and demonstrate sensitivity to change over time. PMID- 12133246 TI - Discussing do-not-resuscitate orders in the hospital setting: part 1. PMID- 12133247 TI - Discussing do-not-resuscitate orders in the hospital setting: part 2. PMID- 12133248 TI - Mr. W. and the Grand Inquisitor: a lesson in autonomy. PMID- 12133249 TI - Crossing paths. PMID- 12133252 TI - A pathway to advancing end-of-life education. AB - OBJECTIVE: This paper identifies key features associated with high-quality educational materials for end-of-life curriculum. METHODS: The End of Life Physician Education Resource Center (EPERC), located on the Internet at provides a clearinghouse for end-of-life materials. All materials posted on EPERC are peer reviewed by content and education experts for quality. An analysis of reviewers' ratings, from the EPERC rating form and their narrative comments, revealed common strengths and weaknesses of submitted materials. Examples of exemplar materials from the EPERC website are presented to highlight key strengths. DISCUSSION: Articulating the best qualities of submitted materials provides clear standards for those seeking to develop or adopt high quality end-of-life educational materials. PMID- 12133253 TI - Refractory neuropathic pain from chronic cord compression. PMID- 12133255 TI - Legalized physician-assisted suicide in Oregon. PMID- 12133256 TI - Editorial: living with grief, rebuilding a world. PMID- 12133257 TI - A weekend retreat for parents and siblings of children who have died. Interview by Samantha Libby Sodickson. PMID- 12133258 TI - Saying goodbye, saying hello: a grief sojourn. PMID- 12133259 TI - Nontyphoidal salmonella bacteremia and pneumonia as the initial manifestation of human immunodeficiency virus infection in a four-year-old child. AB - This report describes the case of a 4-year-old human immunodeficiency virus (HIV) infected girl with Salmonella typhimurium bacteremia and pneumonia. The girl presented with a history of fever for 1 month and mild pulmonary symptoms. Subsequent studies revealed previously undiagnosed HIV disease. PMID- 12133260 TI - Unusual presentations of primary human immunodeficiency virus infection. AB - Opportunistic infections during primary infection with human immunodeficiency virus (HIV) are rare, with the exception of oral and esophageal candidiasis. HIV associated nephropathy (HIVAN) and Pneumocystis carinii pneumonia (PCP) typically occur during advanced HIV infection. We report two patients who developed HIVAN and a presumptive diagnosis of PCP, respectively, during primary HIV infection. Serologic testing demonstrated HIV seroconversion. Clinicians need to have a high index of suspicion when evaluating patients even when risk behaviors are not readily apparent. PMID- 12133261 TI - Factors affecting cognitive functioning in a sample of human immunodeficiency virus-positive injection drug users. AB - Injection drug users represent a major vector of human immunodeficiency virus (HIV) infection in the nation's inner cities, and are an important population for harm reduction treatment interventions to target. However, there has been relatively little research examining the specific contribution of the multiple factors contributing to cognitive functioning among injection drug users that may affect engagement in, and response to, addiction and HIV-related interventions. The current study examined the independent contributions to neuropsychological (NP) test performance of premorbid educational attainment, medical and psychiatric history, long- and short-term drug use, assessed by laboratory, observation, and self-report measures, and HIV disease, assessed by plasma HIV-1 RNA viral load and CD4+ count, in a sample of 90 HIV-positive injection drug users dually addicted to heroin and cocaine. Fully 88% of the sample showed evidence of impairment (>1 standard deviation below the population mean) on an NP test battery selected to assess processes associated with successful engagement in the treatment of substance abuse and HIV, such as learning and memory of verbal information, capacity to solve new problems and deal with more than one stimulus at a time, visual-motor coordination, and visual tracking and cognitive flexibility. In addition to drug use, independent predictors of NP test performance were HIV viral load, educational attainment, and premorbid medical and psychiatric problems. Findings underscore the multiplicity of factors that contribute to cognitive impairment in HIV-positive drug-abusing individuals in addition to drug use. Clinical implications are discussed. PMID- 12133262 TI - Individual and system level factors associated with treatment nonadherence in human immunodeficiency virus-infected men and women. AB - Antiretroviral treatment nonadherence is complex, implicating more than a patient's ability and motivation to be compliant. The bulk of the research to date focuses on individual level barriers with less emphasis on the many system level factors that potentially impact patients' adherence behaviors. This study examined the effects of system enabling factors in addition to the frequently studied individual predisposing and enabling factors upon nonadherence to treatment. One hundred eighty-two patients from five community-based clinics were interviewed and their medical charts examined. Patients' self-reported nonadherence was correlated with clinicians' assessments of medication and appointment nonadherence. Seven individual predisposing factors (gender, ethnicity, birthplace, years of education, HIV Overview of Problems-Evaluation System [HOPES] psychosocial, Hospital Anxiety and Depression Scale [HADS] anxiety, and HADS depression scores) were found to be significantly associated with treatment adherence/nonadherence. Individual enabling factors (hopefulness and access to health care), as well as treatment by clinic staff, a system enabling factor, were significantly associated with adherence/nonadherence. In a multivariate analysis, six factors - age (younger), gender (female), birth outside the United States, level of hopefulness (lower), access to health care (lower), and treatment by clinic staff (poorer) - accounted for 19.3% of the variance in nonadherence. Results of this study indicated that several individual predisposing and enabling factors were potential predictors of treatment nonadherence; however, system enabling factors, e.g., treatment by clinic staff were also associated with treatment nonadherence. Further studies are needed to examine the complex relationships of individual and system related factors in predicting treatment nonadherence. PMID- 12133263 TI - Posttraumatic stress disorder in women attending human immunodeficiency virus outpatient clinics. AB - This study examined posttraumatic stress disorder (PTSD) in human immunodeficiency virus (HIV)-positive women seeking medical care. Specifically, we examined traumatic life events, psychiatric treatment, social support, and demographic characteristics in relation to level of PTSD symptoms. We recruited and obtained informed consent from 47 ethnically diverse HIV-positive women from two HIV outpatient clinics in a county medical system. Forty-one of these women provided complete data on measures assessing demographics, traumatic life events, PTSD symptoms, social support, and psychotherapy/medical history. Analysis of the data demonstrated that a high percentage (42%) of the HIV-positive women were likely to meet criteria for full current PTSD, and an additional 22% for partial PTSD. Of the women likely with full PTSD, 59% were not receiving any psychiatric treatment, and of those likely with partial PTSD, 78% were not receiving any psychiatric treatment. Also, women reported having experienced a mean of 12 traumatic life events. As hypothesized, the level of PTSD was significantly related to the number of life events experienced (rs = 0.52, p < 0.001), and to perceived social support from friends (rs = - 0.34, p < 0.02) and family (rs = - 0.29, p < 0.05). Given the high percentages of women who were found to have experienced traumatic life events and high levels of PTSD symptoms, it seems important to assess and treat PTSD in women with HIV/acquired immune deficiency syndrome (AIDS). PMID- 12133264 TI - Potential barriers to seeking human immunodeficiency virus testing among adults in the United States: data from the 1998 National Health Interview Survey. AB - To assess potential barriers to seeking human immunodeficiency virus (HIV) testing among adults in the United States, data from the 1998 National Health Interview Survey (NHIS) were analyzed. The NHIS is a multistage cluster survey of the United States noninstitutionalized civilian population ages 18 years or older. The 1998 NHIS survey was conducted using the computer-assisted personal interview. Of a nationally representative sample (n = 32,440) of the U.S. noninstitutionalized civilian population, 21,410 (66%) have never been tested for HIV, 9,728 (30%) have been tested, and 1302 (4%) did not complete the survey or refused to answer the question. Among individuals who completed the survey, men (odds ratio [OR]: 1.08, 95% confidence interval [CI] = 1.04, 1,22), individuals ages 50 years or older (OR: 4.01, 95% CI = 3.16, 5.08), or 18-19 years (OR: 2.12, 95% CI = 1.71, 2.63), those who had "up to 11 grade" level of education (OR: 2.16, 95% CI = 1.74, 2.63), those who lived in nonmetropolitan areas (OR: 1.21, 95% CI = 1.14, 1.28), or lived in the Midwest (1.34, 95% CI = 1.24, 1.43) were significantly more likely than their counterparts to have not sought HIV testing. Among individuals who have never been tested for HIV, 58% had no particular reason, 38% felt they were not at risk of contracting HIV, whereas less than 1% feared adverse consequences. The high proportion of adults who never tested for HIV after two decades of HIV epidemic underscores the need for new approaches to fight the spread of HIV infection in the United States. PMID- 12133265 TI - NNRTIs compared in switch group. PMID- 12133266 TI - Infections on rise in San Francisco. PMID- 12133267 TI - Regulatory issues for clinical gene therapy trials. AB - Gene therapy trials are among the most heavily regulated clinical trials. In this review, the basic tenets of human subject research are discussed in the context of the regulatory bodies which oversee this work. The challenges faced by academic research are outlined, including new and proposed regulations which impact human gene therapy investigators. PMID- 12133268 TI - A DNA-based method to assay total and infectious particle contents and helper virus contamination in high-capacity adenoviral vector preparations. AB - High-capacity adenoviral (HC-Ad) vectors are devoid of all viral genes. Therefore, these vectors feature reduced toxicity, immunogenicity, and increased capacity for foreign DNA. HC-Ad vectors are produced in E1-transformed cell lines in the presence of an E1-deleted helper virus that provides in trans all viral functions necessary for vector production. By cre/loxP- or FLPe/Frt-mediated recombination the packaging signal of the helper virus is excised during vector production resulting in nonpackagable helper virus genomes. Although recombinase mediated excision of the packaging signal from the helper virus genome is highly efficient, a small number of helper virus genomes with retained packaging signals are still packaged into capsids. For clinical trials, HC-Ad vector preparations have to be characterized accurately with respect to the number of (1) total HC-Ad vector particles, (2) infectious HC-Ad vector particles, and (3) the number of contaminating helper virus particles. We describe a fast and versatile DNA-based biologic assay for determination of these three parameters by standard laboratory methods. This assay is a useful tool for determining bioactivity data of adenoviral vector preparations and, importantly, allows their comparison among different studies. PMID- 12133269 TI - A novel approach to tumor suppression with microencapsulated recombinant cells. AB - A novel approach to cancer gene therapy is to implant microcapsules containing nonautologous cells engineered to secrete molecules with antineoplastic properties. The efficacy of this treatment is now tested in a mouse model bearing HER-2/neu-positive tumors. Nonautologous mouse myoblasts (C(2)C(12)) were genetically modified to secrete interleukin-2 linked to the Fv region of a humanized antibody with affinity to HER-2/neu. The resulting fusion protein, sFvIL-2, would encompass immune-stimulatory cytokine activity now targeted to the HER-2/neu-expressing tumor. These recombinant cells were then immunoprotected with alginate-poly-L-lysine-alginate microcapsules before implantation into tumor bearing mice. Treatment with these encapsulated cells led to a delay in tumor progression and prolonged survival of the animals. The long-term efficacy was limited by an inflammatory reaction against the implanted microcapsules probably because of the secreted cytokine and antigenic response against the xenogeneic fusion protein itself. However, over the short term (initial 2 weeks), efficacy was confirmed when a significant amount of biologically active interleukin-2 was detected systemically, and targeting of the fusion protein to the HER-2/neu expressing tumor was shown immunohistochemically. The tumor suppression in the treated animals was associated with increased apoptosis and necrosis in the tumor tissue, thus demonstrating successful targeting of the antiproliferative effect to the tumors by this delivery paradigm. In conclusion, this new approach to systemic cancer gene therapy needs to be modified to provide long-term delivery, but has demonstrated short-term efficacy and potential to become a cost effective, benign, and non-viral-based adjunct to the current armory of anticancer strategies. PMID- 12133270 TI - The adenovirus capsid protein hexon contains a highly conserved human CD4+ T-cell epitope. AB - The immunogenicity of adenovirus vectors remains a major obstacle to their safe and efficacious use for gene therapy. In order to identify T-cell epitopes directly from adenoviruses, four viral protein sequences were screened for the well-characterized 9-mer HLA-A2 binding motif. Peripheral blood mononuclear cells (PBMC) from healthy adults were tested for responses to 17 selected viral peptides using a short-term interferon-gamma ELISPOT assay. Memory T-cell responses were identified to a single peptide derived from the major capsid protein hexon in 5 of 6 HLA-A2-positive donors. Unexpectedly, responses to this hexon peptide were also detected in 4 of 6 HLA-A2-negative donors, and responder cells were identified as CD4(+) T cells by immunomagnetic depletion experiments. A longer 15-mer peptide, H910-924, was identified as the optimal CD4(+) T-cell epitope. This hexon epitope induces strong proliferative T-cell responses that can be blocked by a monoclonal antibody against HLA-DR, and molecular HLA typing of donors suggests that the peptide response is restricted by multiple HLA-DR alleles. Additionally, quantitative analysis of responses to H910-924 and whole adenovirus reveals that the frequency of circulating CD4(+) T cells specific for this single hexon epitope (mean = 61 per 10(6) PBMC) represents up to one third of the total adenovirus-specific T-cell response. Finally, comparison of hexon sequences from over 20 different human adenovirus serotypes indicates that H910 924 is highly conserved. In most individuals, therefore, T-cell responses to this hexon epitope will be induced by all adenovirus vectors, including "gutted" vectors packaged with capsid proteins and vectors based on different serotypes. PMID- 12133271 TI - Tetracycline-inducible interleukin-10 gene transfer mediated by an adeno associated virus: application to experimental arthritis. AB - The adeno-associated viruses (AAV) offer new perspectives for cytokine gene transfer in rheumatoid arthritis (RA) because they are nonpathogenic and allow long-term transgene expression in vivo. Moreover, the use of a tetracycline inducible promoter allows regulation of therapeutic gene expression. This study assessed the potential long-term gene regulation of a recombinant AAV vector expressing viral interleukin-10 (vIL-10) in human rheumatoid synovium and the therapeutic efficiency in a mouse model of RA. We constructed a recombinant AAV vector in which the transcription of vIL-10 cDNA is controlled by the TetON system. Transduction of human primary RA synovial cells with AAV-tetON-vIL10 conferred in vitro controlled vIL-10 expression. After intramuscular injection, both incidence and severity of collagen-induced arthritis were significantly reduced at macroscopic, radiological, and histological levels in the group of DBA1 mice treated with AAV-TetON-vIL10 vector plus doxycycline after immunization and boosting compared to control groups. When coinjecting two separate AAV vectors, one encoding the inducible vIL-10 and the other the transcriptional activator, a 10 times excess of the transactivator vector dose allowed efficient control of vIL-10 secretion by doxycycline administration or withdrawal, over an 8-week period. Our results supported, for the first time, the utility of AAV tetON-vIL10 as a therapeutic tool for gene therapy in RA. PMID- 12133272 TI - Heme oxygenase 1 gene transfer prevents CD95/Fas ligand-mediated apoptosis and improves liver allograft survival via carbon monoxide signaling pathway. AB - Apoptosis via the CD95/FasL (CD95L) pathway plays an important role in allograft rejection. Heme oxygenase 1 (HO-1), a stress-responsive cytoprotective molecule, may be essential in preventing graft rejection. We used Ad-HO-1 gene transfer to analyze HO-1-mediated effects in a rat allogeneic orthotopic liver transplantation (OLT) model. The cytotoxicity to Fas-bearing YAC-1 target cells and frequency of terminal deoxynucleotidyltransferase-mediated dUTP nick-end labeling-positive (TUNEL(+)) cells in vitro were diminished in Ad-CD95L + Ad-HO-1 transfected cells, as compared with Ad-CD95L + Ad-beta-gal controls (p < 0.001). AdHO-1 gene transfer prevented <10-day rejection of dark agouti (DA) livers in Lewis (LEW) rats (survival >32 days), and diminished apoptosis. Unlike Ad-beta gal OLTs, which showed signs of severe acute rejection, OLTs in the Ad-HO-1 group exhibited mild to moderate rejection and improved function. These beneficial effects were abrogated after adjunctive treatment with tin protoporphyrin (SnPP), an HO-1 antagonist. Intragraft expression of HO-1 and antiapoptotic gene products (Bcl-xl/Bag-1) was enhanced in Ad-HO-1-transduced OLTs, in association with selectively depressed expression of helper T cell type 1 cytokines (interleukin 2 and interferon gamma), as compared with Ad-beta-gal controls. To deliver CO, one of the downstream HO-1 mediators, allogeneic OLT recipients were exposed to methylene chloride. Such treatment prolonged survival to >47 days, diminished apoptosis, and preserved hepatic architecture/function. Thus, Ad-HO-1 gene transfer prevents CD95/FasL-mediated apoptosis, and significantly prolongs allogeneic OLT survival via a downstream HO-1-CO signaling pathway. PMID- 12133274 TI - Oncolytic herpesvirus effectively treats murine squamous cell carcinoma and spreads by natural lymphatics to treat sites of lymphatic metastases. AB - Oncolytic herpesviruses have significant antitumoral effects in animal models when delivered directly to established tumors. Lymphatic metastases are a common occurrence for many tumor types. This study investigates the potential of an attenuated, replication-competent, oncolytic herpes simplex virus (NV1023) both to treat a primary tumor by direct injection and to travel through the lymphatic system to treat metastatic tumor within the lymph nodes draining lymph from the site of primary cancer. Isosulfan blue dye was injected into murine auricles to determine normal lymphatic drainage patterns and demonstrated consistent blue staining of a group of ipsilateral cervical lymph nodes. Auricular injections of NV1023 resulted in viral transit to these lymph nodes as measured by 5-bromo-4 chloro-3-indolyl-beta-D-galactopyranoside histochemistry and viral plaque assay. An oncolytic herpesvirus (NV1066) expressing green fluorescent protein also demonstrated viral transit from the auricle to the cervical lymph nodes on fluorescence microscopy. Using the SCC VII cell line, a novel murine model of auricular squamous cell carcinoma was developed with an approximately 20% incidence of cervical lymph node metastases. Delivery of NV1023 or NV1066 to the surgical beds after excision of auricular SCC VII tumors resulted in successful viral infection of metastatic SCC VII cells within the cervical lymph nodes. After a 7-week follow-up, significantly enhanced locoregional control (p < 0.05, Fisher exact test) and disease-free survival (p < 0.05, log rank test) were evident with NV1023 treatment. This study demonstrates that the delivery of an oncolytic herpesvirus to a primary tumor site after surgical excision may have a significant impact on reducing both primary site recurrence and regional nodal metastases. PMID- 12133273 TI - Enhancement of bone healing based on ex vivo gene therapy using human muscle derived cells expressing bone morphogenetic protein 2. AB - Molecular biological advances have allowed the use of gene therapy in a clinical setting. In addition, numerous reports have indicated the existence of inducible osteoprogenitor cells in skeletal muscle. Because of this, we hypothesized that skeletal muscle cells might be ideal vehicles for delivery of bone-inductive factors. Using ex vivo gene transfer methods, we genetically engineered freshly isolated human skeletal muscle cells with adenovirus and retrovirus to express human bone morphogenetic protein 2 (BMP-2). These cells were then implanted into nonhealing bone defects (skull defects) in severe combined immune deficiency (SCID) mice. The closure of the defect was monitored grossly and histologically. Mice that received BMP-2-producing human muscle-derived cells experienced a full closure of the defect by 4 to 8 weeks posttransplantation. Remodeling of the newly formed bone was evident histologically during the 4- to 8-week period. When analyzed by fluorescence in situ hybridization, a small fraction of the transplanted human muscle-derived cells was found within the newly formed bone, where osteocytes normally reside. These results indicate that genetically engineered human muscle-derived cells enhance bone healing primarily by delivering BMP-2, while a small fraction of the cells seems to differentiate into osteogenic cells. PMID- 12133275 TI - Tumor-targeted Salmonella expressing cytosine deaminase as an anticancer agent. AB - The study was designed to evaluate whether TAPET-CD, an attenuated strain of Salmonella typhimurium expressing Escherichia coli cytosine deaminase (CD), was capable of converting nontoxic 5-fluorocytosine (5-FC) to the active antitumor agent 5-fluorouracil (5-FU). The antitumor effect of TAPET-CD plus 5-FC against subcutaneously implanted colon tumors was also evaluated. TAPET-CD was given to tumor-bearing mice by a single bolus intravenous administration followed with 5 FC by intraperitoneal administration. TAPET-CD accumulated in tumors at levels 1000-fold higher than that in normal tissues and high levels of 5-FU were detected in tumors in mice treated with both TAPET-CD and 5-FC. No 5-FU could be detected in normal tissues. Inhibition of tumor growth was observed in mice treated with either TAPET-CD alone or TAPET-CD in combination with 5-FC (TAPET CD/5-FC), but not with 5-FC alone. TAPET-CD/5-FC inhibited tumor growth by 88% 96%, compared to TAPET-CD alone, which inhibited tumor growth by 38%-79%. These data suggest that tumor-targeting Salmonella could be used to deliver prodrug converting enzyme selectively to tumors and produced anti-tumor effects when the corresponding prodrug was also given. PMID- 12133276 TI - Scalable purification of adeno-associated virus type 2, 4, or 5 using ion exchange chromatography. AB - The availability of high-titer, high-purity, adeno-associated virus type 2 (AAV2) stocks has dramatically increased our understanding of this virus and its utility as a gene transfer vector. Current methods of purification take advantage of the stable interaction of AAV2 with heparin sulfate. This affinity chromatography, however, is not useful for purifying AAV4 and AAV5, because these serotypes lack heparin-binding activity. We have developed simple ion exchange high-performance liquid chromatography (HPLC) method for purifying different AAV serotypes that does not rely on the affinity of the viruses for heparin. The protocol is fast, efficient, and yields highly infectious material. Analysis of the highly purified virus indicated that more than 90% of the particles contained genomes and were more active than virus purified by cesium chloride (CsCl) gradient purification. This procedure is scalable and can easily be streamlined for large-scale production of recombinant adeno-associated virus (rAAV), regardless of the serotype. Ultimately, the new purification method will further the characterization of rAAV of different serotypes as vectors for gene therapy applications. PMID- 12133277 TI - Reversible model of spheroid formation allows for high efficiency of gene delivery ex vivo and accurate gene assessment in vivo. AB - The native three-dimensional architecture of carcinomas, which governs numerous autocrine-paracrine interactions related to tumor progression, cannot be faithfully recreated in most in vitro models. Even when the three-dimensional architecture is recreated in artificial scaffolds such as soft agar, this approach does not truly recreate the natural microenvironment of the tumor. Multicellular spheroids can reasonably recreate in vitro the natural three dimensional architecture of carcinomas, but even the most efficient gene delivery vectors will penetrate only the outer layers of these structures and hence only a small fraction of cells will receive the gene of interest. If the multicellular spheroids are disrupted into a single-cell suspension in order to achieve high transfection efficiency, the single-cell production may have so altered the gene expression profile of the spheroid as to bring into question its present relevancy to in vivo tumor progression. Our laboratory has developed a human-SCID (severe combined immunodeficient) mouse model of inflammatory breast cancer, MARY X, which grows as tight multicellular spheroids in vitro and as lymphovascular emboli in vivo. The spheroids, which express only low levels of surface sialyl Lewis(x/a) (sLe(x/a)), are able to form compact homotypic cell clumps mediated by an intact, overexpressed E-cadherin/alpha,beta-catenin axis. The spheroids can be fully disrupted by trypsin proteolysis, anti-E-cadherin antibodies, or Ca(2+) depletion. Of these approaches the disruption with depleted Ca(2+), complete after 30 min, is fully reversible by the readdition of Ca(2+) within 6 hr. This time interval allows for a transfection "window" in which successful gene delivery can be achieved before spheroid reformation. Retroviruses (10(6)-10(7) CFU/ml) carrying the gene encoding either green fluorescent protein (GFP), a dominant-negative E-cadherin mutant (H-2K(d)-E-cad), its control (H-2K(d)-E-cad Delta C25), or alpha-1,3-fucosyltransferase III (FucT-III), an enzyme that increases surface sLe(x/a), were used to transfect either intact (wild-type) or disadhered/readhered (reformed) spheroids. There were marked differences in transfection efficiency in the reformed versus wild-type spheroids. Retroviral transfection of GFP resulted in successful delivery of this reporter gene to only the outer layer of cells of the wild-type spheroids, but to all layers of the reformed spheroids. A single retroviral transfection of H-2K(d)-E-cad, H-2K(d)-E cad Delta C25, or FucT-III produced evidence of their respective gene expression at 72 hr throughout all layers of the reformed spheroids, but only H-2K(d)-E-cad and FucT-III produced progressive disadherence. Both H-2K(d)-E-cad-MARY-X and FucT-III-MARY-X lost their ability to form lymphovascular emboli in SCID mice. This reversible model of spheroid formation has provided us with insight into the pathogenesis of inflammatory breast carcinoma. If more broadly applied, this model could be used to examine the effects of any gene, using any gene delivery system in the three-dimensional context of native tumoral architecture. PMID- 12133279 TI - Retrospective analysis of percutaneous transluminal coronary angioplasty and coronary stenting. AB - OBJECTIVE: To examine long-term efficacy of percutaneous transluminal coronary angioplasty (PTCA), coronary stenting and to assess the factors affecting its efficacy. METHODS: A total of 790 patients who underwent successful PTCA and PTCA + stent in this hospital were followed by direct interview or letter. The rate of follow-up was 84.2% and the period of follow-up was 0.9 - 12.7 (3.5 +/- 2.4) years. RESULTS: During follow-up, 4 (0.5%) patients died, 22 (2.8%) had nonfatal acute myocardial infarction, 10 (1.3%) had coronary artery bypass surgery, and 98 (12.4%) had repeat PTCA. The rate of recurrent angina pectoris was 31.1%. The cardiac event-free survival rate calculated by the Kaplan-Meier method was 88.2% at 1 year and 80.6% at 12.7 years. Cox regression analysis showed that there was a positive correlation between AMI history, stent implantation and the risk of cardiac events, and there was a negative correlation between the number of diseased arteries and the risk of cardiac events. Compared to the PTCA group, patients with PTCA + stent had significantly lower rates of total cardiac events. CONCLUSION: The long-term efficacy of PTCA, especially PTCA + stent in Chinese patients was very satisfactory, suggesting that PTCA + stent therapy should be the major treatment for revascularization in patients with coronary heart disease. PMID- 12133280 TI - Serial high-frequency ultrasound assessment of progressive changes in left ventricular structure and function in rats with chronic pressure overload. AB - OBJECTIVE: To determine the feasibility and accuracy of high-frequency ultrasound in evaluating the left ventricular (LV) structure and function in normal and pressure overload rats and to examine the changes of the left ventricle during its transition from hypertrophy to heart failure. METHODS: Thirty-eight female rats were randomly assigned to normal (n = 10), operated (n = 16) and sham operated (n = 12) groups. Parasternal long axis and short axis images were acquired by a 7.5 mHz linear ultrasound probe at 12 weeks and 20 weeks after the operation respectively. RESULTS: Left ventricular structure and function could be satisfactorily imaged for dimensions and mass. Compared to the sham-operated groups, at 12 weeks after the operation, the operated rats had increased LV wall thickness and mass (P < 0.01) with normal cavity and FS% (P > 0.05). At 20 weeks after the operation, the LV wall thickness showed no further progressive change and the LV mass increased greatly with slightly dilated LV cavity and decreased FS% (P < 0.05). CONCLUSIONS: High-frequency echocardiography provides a useful means to noninvasively evaluate LV dimensions, mass and function in rats. It will have great value for evaluating LV remodeling during the transition from LV hypertrophy to heart failure, as well as the effects of intervening drugs. PMID- 12133281 TI - Clinical implications of increased expression of CD40L in patients with acute coronary syndromes. AB - OBJECTIVE: To investigate clinical implications of expression of CD40L in monocytes and changes in serum soluble CD40L in patients with acute coronary syndromes (ACS). METHODS: Sixteen control and 56 patients, including 24 with stable angina (SA), 20 with unstable angina (UA) and 12 with acute myocardial infarction (AMI) enrolled in this study. Expression of CD40L in monocytes was analyzed by flow cytometry and sCD40L levels were measured by ELISA. RESULTS: Expression of CD40L in monocytes and serum levels of sCD40L in UA and AMI patients were higher than in SA patients and controls. In patients with AMI, sCD40L levels showed no significant increase when compared to patients with UA, while AMI patients had a peak level of sCD40L at 24 hours after AMI. PTCA induced a marked rise in sCD40L levels in all patients, while CD40L expression in monocytes showed no difference between patients with PTCA, before and after. CONCLUSION: Enhanced level of serum sCD40L may be a reliable prognostic indicator for ACS and may represent a marker of coronary disease activity. PMID- 12133282 TI - Activated focal adhesion kinase involved in adhesion and migration of vascular smooth muscle cells stimulated by fibronectin. AB - OBJECTIVE: To study the effects of focal adhesion kinase (FAK) phosphorylation on smooth muscle cells (SMCs) adhesion and migration stimulated by fibronectin. METHODS: Adhesion and migration of cultured SMCs were stimulated by different concentrations of fibronectin (FN), FAK and its phosphorylation were detected by immunoprecipitation and Western blot. FAK antisense oligodeoxynucleotides (ODNs) were transfected into SMCs by cationic lipid to investigate its modulatory effects on tyrosine phosphorylation. SMCs adhesion and migration were also measured by morphological enumeration and modified Boyden Chambers, respectively. RESULTS: FAK were expressed when SMCs adhesion and migration were successfully simulated by different concentrations of FN. FAK phosphorylation were detected only at 20 microg/ml FN or more. FAK antisense ODNs were transfected efficiently by cationic lipid and FAK phosphorylation was inhibited substantially. The SMCs migration rate in the 5 - 60 microg/ml FN groups was reduced by 17.89% - 27.67%. Cell migration stimulated by FN at 10, 20, 40 and 60 microg/ml were reduced by 23.26%, 21.63%, 19.31% and 17.88%, respectively (P < 0.05). CONCLUSIONS: FAK phosphorylation and FAK-mediated signal transduction play important roles in SMCs adhesion and migration stimulated by ECM. The process can be inhibited effectively by FAK antisense ODNs. PMID- 12133284 TI - Characteristics of single Ca(2+) channel kinetics in feline hypertrophied ventricular myocytes. AB - OBJECTIVE: To explore the mechanism underlying the prolongation of action potential and delayed inactivation of the L-type Ca(2+) (I(Ca, L)) current in a feline model of left ventricular system hypertension and concomitant hypertrophy. METHODS: Single Ca(2+) channel properties in myocytes isolated from normal and pressure overloaded cat left ventricles were studied, using patch-clamp techniques. Left ventricular pressure overload was induced by partial ligation of the ascending aorta for 4 - 6 weeks. RESULTS: The amplitude of single Ca(2+) channel current evoked by depolarizing pulses from -40 mV to 0 mV was 1.02 +/- 0.03 pA in normal cells and 1.05 +/- 0.03 pA in hypertrophied cells, and there was no difference in single channel current-voltage relationships between the groups since slope conductance was 26.2 +/- 1.0 pS in normal and hypertrophied cells, respectively. Peak amplitudes of the ensemble-averaged single Ca(2+) channel currents were not different between the two groups of cells. However, the amplitude of this averaged current at the end of the clamp pulse was significantly larger in hypertrophied cells than in normal cells. Open-time histograms revealed that open-time distribution was fitted by a single exponential function in channels of normal cells and by a two exponential function in channels of hypertrophied cells. The number of long-lasting openings was increased in channels of hypertrophied cells, and therefore the calculated mean open time of the channel was significantly longer compared to normal controls. CONCLUSION: Kinetic changes in the Ca(2+) channel may underlie both hypertrophy-associated delayed inactivation of the Ca(2+) current and, in part, the pressure overload-induced action potential lengthening in this cat model of ventricular left systolic hypertension and hypertrophy. PMID- 12133283 TI - Hypertensive epidemiology in Heilongjiang Province in China. AB - OBJECTIVE: To assess the epidemiology features of hypertension in Heilongjiang Province, China. METHODS: From 1959 to 1999, 289,157 people in total, ages > or = 15-year old (male 154,091, female 135,066) were investigated four times by a stratified chunk method in rural and urban areas in Heilongjiang Province. After a staff training program, this survey on blood pressure and risk factors was carried out under a worldwide uniform protocol with standardized method and interrelated quality control regulation. Among these samples, 1615 people (male 824, female 791) received blood-lipid analysis. RESULTS: The hypertensive incidence of Heilongjiang was much higher than the average level in China (P < 0.01) and it has gotten higher from 1959 to 1999, especially in the last ten years. It increased more in males than in females (P < 0.01). It was also higher with age. The mean age of hypertensive onset became ten years younger recently. It is different in different areas and professions. Age, BMI (body mass index), drinking alcohol, gender, history of maternal hypertension, high salt diet and professional status were the main risk factors for hypertension in Heilongjiang Province in China. CONCLUSION: The prophylaxis and treatment of hypertension should be enhanced in this area. PMID- 12133285 TI - Relationship between the U wave on electrocardiogram and the midmyocardium of the left ventricular wall. AB - OBJECTIVE: To investigate the relationship between the U wave on electrocardiogram and the midmyocardium in rabbit left ventricle free wall in vivo. METHODS: The monophasic action potentials in the epicardium, midmyocardium, and endocardium of the left ventricle free wall were recorded simultaneously in 16 rabbits. The rabbits were then given an intravenous injection of Sotalol (1, 1.5 and 2.0 mg/kg) in 30 minutes intervals, and measurements were taken. RESULTS: In the basic condition, there were no U wave on electrocardiogram. The U wave appeared after the intravenous Sotalol at 1.5 mg/kg, and the U wave became greater with increased dosage of intravenous Sotalol (2 mg/kg). The repolarization duration of the midmyocardium was prolonged longer than that of the epicardium and endocardium by Sotalol, and the repolarization duration of the epicardium coincided with the apex of the T wave, The repolarization duration of the midmyocardium coincided with the end point of the U wave. CONCLUSION: The U wave may originate from the delayed repolarization of the midmyocardium. PMID- 12133286 TI - TNF-alpha and the phenotypic transformation of human peritoneal mesothelial cell. AB - OBJECTIVE: To investigate if tumor necrosis factor-alpha (TNF-alpha) could induce phenotypic transformation or the associated cell function changes of human peritoneal mesothelial cells (HPMCs). METHODS: All tests were performed on the human peritoneal mesothelial cell line (HMrSV5, kindly donated by Prof. Pierre RONCO). Morphological changes of HPMCs were observed by phase contrast microscopy. The expressions of cytokeratin 8, cytokeratin 18 and vimentin were detected by immunocytochemistry. mRNA expressions of matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) were detected by semiquantitative reverse transcription polymerase chain reaction (RT-PCR). Production of MMP-9 and type I collagen was measured by enzyme-linked immunosorbent assays. RESULTS: TNF-alpha (1 - 10 ng/ml) induced morphological changes in about 50% - 60% of human peritoneal mesothelial cells (HPMCs). The original polygonal cobblestone monolayer was changed into elongated, spindle shape characteristic of fibroblasts. The original strong positive expression of cytokeratin 8 and cytokeratin 18 was changed in all morphologically changed HPMCs to negative, but the expression of vimentin maintained positive. By 4-hour treatment with TNF-alpha (1 - 10 ng/ml), mRNA expression of MMP9 was significantly up-regulated, and mRNA expression of TIMP-1 and TIMP-2 were down regulated. We also observed that the production of MMP-9 and type I collagen increased significantly after 24-hour treatment with TNF-alpha. CONCLUSION: 24 hour treatment with TNF-alpha, produced a partial transformation of HPMCs into the fibroblast phenotype. TNF-alpha treatment of HPMCs up-regulated the synthesis of MMP-9 and type I collagen, which may facilitate peritoneal extracellular matrix remodeling and fibrogenesis. PMID- 12133287 TI - Plating densities, alpha-difluoromethylornithine effects and time dependence on the proliferation of IEC-6 cells. AB - OBJECTIVE: To characterize the role of plating densities and alpha difluoromethylornithine (DFMO) on the proliferation of IEC-6 cells in vitro. METHODS: IEC-6 cells were seeded in 96-well microplates at various densities in the presence or absence of DFMO. Cells were counted and their proliferative capability was monitored Days 1 to 7 with MTT assay at an optical density of 570 nm. RESULTS: There was a positive relationship between cell number and OD value (r = 0.954, P < 0.01). Higher plating densities (> 0.5 x 10(4) cells/well) inhibited the growth of cells on Day 2. When the density reaches 4 x 10(4) cells/well, the OD value increased gradually and reached a peak on Day 5. After that, the OD value began to fall. The growth of IEC-6 cells was limited at a low density (0.2 x 10(4) cells/well) on Day 4. DFMO caused a complete inhibition of proliferation of IEC-6 cells on Days 1 to 3. CONCLUSION: Proliferation of IEC-6 cells is related to plating density and incubation time. It is inhibited by DFMO, but is reversible when the incubation time is prolonged. PMID- 12133288 TI - Characterization of proteolipid protein-peptide-specific CD(4)(+) T cell of experimental allergic encephalomyelitis in Biozzi AB/H mice. AB - OBJECTIVE: To detect the function of proteolipid protein (PLP) peptide (residues 56 - 70)-specific CD(4)(+) T cells in experimental allergic encephalomyelitis (EAE) in Biozzi AB/H mice (H-2A(g7)). METHODS: Biozzi AB/H mice were immunized by synthetic PLP(56 - 70) peptide (DYEYLINVIHAFQYV) which was emulsified by sonication with complete Freund's adjuvant, a EAE model proven histologically and clinically. Murine splenocytes and spinal cord infiltrated (SCI) T cells were stimulated by PLP(56 - 70), then the CD(4)(+) T cells were isolated by Dynabeads, and confirmed by staining with anti-CD(4) antibody. Finally, the IL2 bioassay and IFN-gamma/IL4 ELISA were done to detect T cell proliferation and cytokine secretion after PLP(56 - 70) stimulation. RESULTS: The histology of murine spinal cord showed a great number of lymphocytes infiltrated the spinal cord; the clinical signs showed high scores (4.3) on the peak, as well as a good EAE model. After being isolated by Dynabeads, CD(4)(+) T cells showed high purification (> 99%) by staining with anti-CD(4) antibody. IL2 bioassay showed that those T cells were PLP(56 - 70)-specific T cells. ELISA showed that those T cells had high IFN gamma/IL4 ratio, indicating that they are T helper 1 (Th1) cells. CONCLUSIONS: PLP(56 - 70)-specific splenocytes and SCI CD(4)(+) T cells in EAE from Biozzi AB/H mice were detected and showed that both of them were PLP(56 - 70)-specific Th1 cells. It is beneficial to understand what kind of role these T cells play in the development of EAE. PMID- 12133289 TI - Application of transesophageal echocardiography to aortic embolic stroke. AB - OBJECTIVES: To determine the relative value of transesophageal echocardiography (TEE) and transthoracic echocardiography (TTE) in exploring the potential embolic source (PES) in heart and aortic arch and to study the clinical significance of aortic arch atherosclerosis (AAA). METHODS: Forty-nine patients with cerebral embolism were included in this study. TEE and TTE were used to evaluate the potential source of emboli in aortic arch, heart and duplex in the carotid artery. An atherosclerotic lesion of the aortic arch was defined as normal, mild plaque, moderate plaque, and protruding plaque or mobile plaque. RESULTS: Of the 49 patients, 31 (63%) patients showed evidence of AAA: 7 (14.1%) patients were mild, 9 (18.4%) were moderate and 15 (30.6%) were severe. In those 15 patients, 11 had neither severe ICAA nor heart disease. Thirty-three patients had internal carotid arterial atherosclerosis (ICAA). The potential sources of embolization of heart and aortic arch is 48.98% by TEE, but only 18.4% by TTE; 9 patients had heart disease. Age and ICAA were significantly correlated with AAA. CONCLUSION: At present, TEE is a better method for exploring atherosclerotic lesions in the thoracic aorta. AAA is an important potential source of cerebral embolic stroke. PMID- 12133290 TI - Detection of occult metastases in lymph nodes from patients with colorectal carcinoma by reverse transcriptase-polymerase chain reaction. AB - OBJECTIVE: To detect occult metastases in lymph nodes from patients with colorectal carcinoma and its clinical significance. METHODS: The metastases in 260 lymph nodes from 39 histologically verified colorectal cancer patients were studied by both hematoxylin-eosin (HE) staining and cytokeratin-20 (CK20) specific RT-PCR. Ten normal lymph nodes were served as negative controls, and HT29 colon cancer cell line and 5 colorectal cancer specimens as positive controls. RESULTS: Ten normal lymph nodes were CK20-negative, HT29 cells and 5 tumor specimens were all CK20-positive. All 29 lymph nodes from 16 patients which confirmed metastases by HE staining exhibited CK20 positive expression; an additional 28 lymph nodes from 5 patients with no histologically detectable metastases expressed CK20 mRNA, i.e. presence of metastases. The difference of the positivity was significant (11.1% vs 21.9%, P < 0.01). According to the HE staining, the cases of Dukes' A, B, C and D were 3, 20, 12 and 4, respectively. In the 20 patients of Dukes' B stage, 5 of them had CK20-positive lymph nodes. CONCLUSION: CK20-specific RT-PCR is a highly sensitive, specific and simple method for detecting occult metastases in lymph nodes. The detection of CK20 mRNA expression in lymph nodes is recommended to precisely determine tumor stage and postoperative adjuvant therapy for patients with colorectal cancer, and further studies should be done in future to confirm the findings. PMID- 12133291 TI - Mature insulin production by engineered non-beta cells. AB - OBJECTIVE: To pursue insulin and islet-transplantation replacement therapy for type 1 diabetes based on engineered human non-beta cells which secrete mature insulin. METHODS: Human proinsulin cDNA was cloned from its genomic gene and mutated by overlap extension PCR, introducing furin consensus cleavage sequences (Arg-Xaa-Lys/Arg-Arg). An expression vector encoding a genetically modified human proinsulin cDNA was generated and transduced to Hela, 293, and L02 cells by lipofectin-mediated DNA transfection. Following G418 screening, the surviving L02 cells were selected and enriched. Insulin levels in the supernatant and cells were evaluated using radioimmunoassay and immunofluorescence staining. RESULTS: Three sites in the insulin gene were mutated simultaneously. Insulin gene modified cells were able to express insulin at different levels: 8.45 - 188.00 microIU/24 h/2.0 x 10(6) Hela cells and 159.88 - 242.14 microIU/24 h/2.0 x 10(6) 293 cells for transient expression, and 2.56 - 61.95 microIU/24 h/2.0 x 10(6) from several L02 clones screened with G418. No insulin was released by control cells. Furthermore, immunofluorescence staining confirmed that proinsulin was stored as vacuoles in the cytoplasm of L02 cells. CONCLUSION: A correctly mutated human proinsulin cDNA was obtained successfully, transfected and expressed efficiently in non-beta cells, lending support to the study of somatic gene therapy in diabetes mellitus. PMID- 12133292 TI - Early and late outcomes in Hong Kong Chinese patients undergoing carotid endarterectomy. AB - OBJECTIVE: To determine the benefit of carotid endarterectomy (CEA) for stroke prevention by reviewing the early and late outcomes of Hong Kong Chinese patients undergoing CEA who have a high reported incidence of intracranial atherosclerotic disease (IAD). METHODS: Fifty-nine Chinese patients underwent 62 CEA. There were 48 males and 11 females, with a mean age of 70 +/- 7 years (range: 52 - 86 years). Twenty-one CEA (34%) were performed for asymptomatic disease. Duplex scan was the primary tool of evaluation prior to surgery. Preoperative angiography was done in 36 instances (58%). All CEA were performed under general anaesthesia with routine intraoperative shunting. The arteriotomy was closed primarily in all patients except three. Patients were followed up regularly with six-monthly Duplex scan surveillance. RESULTS: There were 2 perioperative neurological events consisting of one transient ischemic attack and one minor stroke. There was no operative mortality or major morbidity such as bleeding or cranial nerve injury. Mean hospital stay was 6.5 +/- 4 days (range: 3 - 26 days). The patients were followed up for a mean interval of 24 +/- 17 months (range: 1 - 57 months). Seven patients died during follow-up and subsequent neurological events occurred in 5 patients, including 2 fatal strokes. The 3-year survival, freedom from stroke and stroke free survival were 86%, 87% and 83%, respectively. One recurrent stenosis of 80% was detected on follow-up Duplex scan. CONCLUSIONS: Despite a high incidence of IAD, CEA in Hong Kong Chinese patients is associated with acceptable perioperative morbidity and mortality with satisfactory long-term efficacy in stroke prevention. PMID- 12133293 TI - Magnetic resonance urography in the diagnosis of urinary tract obstruction after renal transplantation. AB - OBJECTIVE: To evaluate magnetic resonance urography (MRU) in the diagnosis of urinary tract obstruction after renal transplantation. METHODS: A total of 31 patients with suspected urinary tract obstruction after renal transplantation were examined, and the results were compared with those from surgery and B ultrasound examination. RESULTS: The urinary tract after renal transplantation was clearly shown using MRU, and all patients were clearly diagnosed by MRU. Imaging results were consistent with those from surgery. CONCLUSIONS: MRU is suitable for detecting urinary tract obstruction after renal transplantation. It is a alternative method for IVP and CT in patients with renal function impairment and uremia. PMID- 12133294 TI - Liver transplantation at the Sun Yat-Sen University of Medical Sciences in China. AB - OBJECTIVES: To summarize the results of liver transplantation for various end stage liver diseases at the Sun Yat-Sen University of Medical Sciences (SUMS), define the role of liver transplantation in the treatment of hepatocellular carcinoma and fulminant hepatitis B, and assess the efficiency of lamivudine on preventing HBV recurrence. METHODS: Seventy liver transplants performed at the SUMS between April 1993 and December 2000 were retrospectively analyzed. The main indications for liver transplant were hepatocellular carcinoma (26 cases), liver cirrhosis (21 cases), fulminant hepatitis B (12 cases), sclerosing cholangitis (4 cases) and other terminal liver diseases (7 cases). Lamivudine was used in twelve patients suffering from fulminant hepatitis B. Logistic multivariate regression analysis was applied to determine the risk factors predicting liver transplantation outcomes. RESULTS: Fifty-four patients survived for more than one month, and 16 patients died within 30 days after orthotopic liver transplantation (OLT). The overall hospital survival rate was 77.1%. The hospital survival rates in the Child's A and B patients were 87.5% and 83.3%, respectively. Those rates were superior to those of the Child's C patients (P < 0.05). The outcome of patients with small hepatocellular carcinoma (HCC) was superior to that of patients with large HCC. Preoperative APACE III scores, the severity of ascites and serum creatine level had independent influence on outcome. Of the patients with fulminant HBV infection, 9 recipients survived for a follow-up period of 2 - 24 months. Treatment with lamivudine monotherapy was both well tolerated and efficacious in patients with fulminant hepatitis B. CONCLUSIONS: The results indicate that orthotopic liver transplantation could provide long-term cure and palliation for patients with HCC, and that patient selection is extremely important in predicting outcome. The results support the continued application of liver transplantation as a therapeutic modality for various end-stage liver diseases and that lamivudine is an effective and safe monotherapy in OLT for patients with HBV infection. PMID- 12133295 TI - Insulin sensitivity after pancreaticoduodenal transplantation with systemic and portal venous drainage in inbred rats. AB - OBJECTIVE: To assess the metabolic consequences of pancreatic transplantation with portal venous drainage and systemic venous drainage in induced inbred rats with streptozocin. METHODS: Pancreaticoduodenal transplantation was performed on 8 rats with the donor portal veins anastomosed to the recipient superior mesenteric vein and on 10 rats with the donor portal veins anastomosed to the recipient cava inferior vein. We measured the recipients' weight, urine and plasma glucose concentration, plasma insulin concentration at the beginning, and before and after transplantation. We used the euglycemi-hyperinsulinemic glucose clamp test and glucose infusion required as an index of insulin sensitivity. RESULTS: The plasma glucose and insulin concentration recovered to normal after transplantation in the portal venous drainage group. The plasma insulin levels increased in the systemic venous group after transplantation. There was a difference in the glucose infusion required between the two groups. CONCLUSION: These data imply that portal venous drainage of the transplanted pancreas is an important factor in the determination of peripheral insulin sensitivity. PMID- 12133296 TI - Long-term follow-up results of dural reconstruction without bone graft at anterior skull-base defects. AB - OBJECTIVE: To clarify whether it is necessary to reconstruct bone defects at the anterior skull base. METHODS: A long-term follow-up study of 50 patients with anterior skull-base defects in which the dura was reconstructed without bone grafts was conducted. CT and MRI examinations were taken periodically after surgery. RESULTS: The ordinates of the bone defects averaged 3.5 cm (range, 2 - 6 cm), and the abscissas averaged 2.8 cm (range, 2 - 5 cm). The abscissas of the bone defects measured 2 - 3 cm in 38 patients, 3 - 4 cm in 10 patients, and 4 - 5 cm in 2 patients. The follow-up ranged from 3 months to 5 years (average, 2 years). CONCLUSIONS: At normal intracranial pressure, if the dura mater is repaired properly at the skull-base defects and reinforced with a pedicled pericranial flap, encephalomeningocele and cerebrospinal fluid (CSF) leakage can be prevented. It may not be necessary to make free bone grafts when the size of the cranial base bone defect is smaller than 4 cm. PMID- 12133297 TI - Experimental study of Eudragit mixture as a new nonadhesive liquid embolic material. AB - OBJECTIVE: To assess the embolic effects and biocompatibility of Eudragit mixture, a new liquid embolic agent. METHODS: In vitro, the viscosity and precipitation time of Eudragit mixtures at several concentrations were measured to study the best proportion of components of the mixture. In vivo, a branch of the right external carotid artery was embolized with Eudragit mixture in 12 rabbits, and with n-butyl cyanoacrylate in another 12 rabbits for a comparative study of the general, angiographic and histopathologic changes between the two groups. RESULTS: Eudragit mixture containing 7.5 g Eudragit, 50 ml absolute ethanol and 50 ml iopromide was shown in vitro to have good properties including rapid precipitation and soft elastic sponge formation upon contact with blood; in vivo, to be nontoxic, nonadherent to the microcatheter and able to embolize the vascular lumen completely without later recanalisation. CONCLUSION: Eudragit mixture is an effective, nontoxic, safe and promising liquid embolic agent. PMID- 12133298 TI - Pathological morphology alteration of the splanchnic vascular wall in portal hypertensive patients. AB - OBJECTIVE: To investigate the pathological morphology alteration of the splanchnic vascular wall in portal hypertensive patients. METHODS: Splenic arteries, veins and gastric coronary veins from portal hypertensive patients (n = 50) were removed during esophagogastric devascularization with splenectomy and were observed under optic and electron microscopes. The expression of iNOS in the splenic artery wall was analysed with immunohistochemistry. RESULTS: The internal elastic membrane and medial elastic fibers of the splenic artery wall were broken and degenerated. Atrophy, apoptosis and phenotypic changes were seen in smooth muscle cells of splenic arteries. Positive staining for iNOS was seen in the cytoplasm of smooth muscle cells and iNOS activity was elevated compared with the non-cirrhotic patients (P < 0.01). In the splenic and gastric coronary veins of cirrhotic patients, we found proliferative intima, extensive thrombi adhering to the venous wall, mimicked arteriosclerosis plaques accompanied with hypertrophy of smooth muscle cells, and thickened muscle fibers of veins with increase in extracellular matrix. CONCLUSION: Portal hypertension may be complicated by splanchnic arterial and venous vasculopathy. There may be an interactive relationship among portal hypertension, splanchnic hyperdynamic disturbances and splanchnic vasculopathy in the pathogenesis of portal hypertension. PMID- 12133299 TI - Multifocal electroretinograms in the early stages of diabetic retinopathy. AB - OBJECTIVE: To investigate the characteristics of multifocal electroretinograms (mf-ERG) of different phases in diabetic retinopathy (DR) and its clinical significance. METHODS: Multifocal electroretinograms in patients with DR (I - II stage) were tested with VERIS IV system. RESULTS: In I - II stage, the absolute values of N1, P1 and N2 response densities, and the N1-P1 and P1-N2 response densities were attenuated is a field of about 45 degrees in diameter. CONCLUSION: As a new objective and quantitative examination for spatial visual function, multifocal electroretinograms may be valuable in the diagnosis of diabetic retinopathy. PMID- 12133300 TI - A clinical trial of active immunotherapy with anti-idiotypic vaccine in nasopharyngeal carcinoma patients. AB - OBJECTIVE: To investigate the effect of active immunotherapy with anti-idiotypic vaccine in patients with nasopharyngeal carcinoma (NPC). METHODS: Anti-idiotypic antibodies (2H4/5D3) bearing the internal image of the NPC antigen were used in active immunotherapy in NPC patients receiving radiotherapy. Antibodies and cytokine levels in patient sera were determined using ELISA before and after active immunotherapy. IL-2 mRNA expression in the peripheral blood mononuclear cells (PBMC) was measured by in situ hybridization. RESULTS: Nineteen patients with NPC at stage IV were treated with alum-precipitated 2H4 or 5D3. Neither hypersensitivity nor adverse side effects were observed. The levels of anti-anti idiotypic antibodies (Ab3) and anti-NPC antibodies (Ab1') were increased. Human anti-mouse antibodies (HAMA) were seen in 19 patients of the experimental group; the levels of Ab1' did not increase in the control group. Serum IL-2, IFN-gamma and TNF-alpha levels were increased in most patients in the experimental group, while no differences were observed in Ab1' and cytokine levels between pre- and post-therapy in the control group. In addition, IL-2 mRNA expression in PBMCs from NPC patients was closely related to serum IL-2 (r = + 0.8829) levels by in situ hybridization. CONCLUSIONS: Anti-idiotype vaccine is safe for clinical active immunotherapy. Anti-idiotypic vaccine might be able to enhance humoral and/or cellular immunity in NPC patients receiving radiotherapy. PMID- 12133301 TI - High frequency loss of heterozygosity on the long arms of chromosomes 13 and 14 in nasopharyngeal carcinoma in Southern China. AB - OBJECTIVE: To investigate the loss of heterozygosity (LOH) on chromosomal arms 13q and 14q in nasopharyngeal carcinoma (NPC) using 21 microsatellite polymorphic markers and to study whether there is a correlation between LOH and clinicopathologic parameters and/or Epstein-Barr virus (EBV) infection in NPC. METHODS: Sixty cases of NPC were studied using polymerase chain reaction based microsatellite analysis with genescan and genotyping techniques. RESULTS: LOH was detected on 13q in 78% of NPC tumors, high frequency LOH loci (more than 30%) clustered to 13q12.3-q14.3 and 13q32. On chromosome 14q, LOH was detected in 80% of NPC tumors; high frequency LOH loci clustered to 14q11-q13, 14q21-q24 and 14q32. High frequency LOH at 13q31-q32 correlated with a lower level of EBV infection; LOH on chromosome 14q was closely associated with poor differentiation of NPC tumor cells. CONCLUSION: Our results suggest that in NPC, LOH on chromosome 13q and 14q are common genetic events, and putative tumor suppressor genes (TSG) residing in these regions may be involved in tumorigenesis. PMID- 12133302 TI - Protective immunity induced by the anti-idiotypic monoclonal antibody NP30 of Schistosoma japonicum. AB - OBJECTIVE: To investigate the protective immunity induced by the anti-idiotypic monoclonal antibody NP30 of Schistosoma japonicum in mice. METHODS: An orthogonal table L(16) (4 x 2(12)) was selected as the experimental design. Eight-week-old Kunming outbred mice (male and female) were randomly divided into 16 experimental groups and 2 control groups. Control groups were injected with SP2/0 ascites intraperitoneally. Mice from each group were infected with 100 +/- 2 cercariae of Schistosoma japonicum in the abdominal skin and were sacrificed on the thirtieth day postchallenge. Adult worms were recovered and counted by perfusion of the left ventricle-portal vein. The SP2/0 ascites injected mice were used as controls and the percentage of protection was calculated. RESULTS: Active immunization of mice with NP30 could produce protection levels ranging from 22.36% to 50.46% depending on the different immunity protocols. The best immunization protocol was established from the results. CONCLUSIONS: Active immunization with NP30 can induce a degree of protection to infection with Schistosoma japonicum cercariae and NP30 is a potential vaccine candidate against Schistosoma japonicum. PMID- 12133303 TI - Determination of specific IgG4 for diagnosis and therapeutic evaluation of cerebral cysticercosis. AB - OBJECTIVE: To probe the significance of specific IgG4 in sera of patients with cerebral cysticercosis for diagnosis and therapeutic evaluation. METHODS: Specific IgG4 in sera of patients with cerebral cysticercosis was assessed using colloidal gold-labeled mouse-anti-human IgG4 McAb as probe. The results were compared with the CT image manifestation. RESULTS: The specific IgG4 positive rate in sera of patients with cerebral cysticercosis was 97.8%, whereas sera from patients with other kinds of parasitosis or central nerve system disease and the control group were all negative, except for a weak cross-reaction of sera from patients with hepatic echinococoosis. The determination of specific IgG4 in sera of patients with cerebral cysticercosis during different times of treatment showed that along with an increase in treatment time and improvement of clinical symptoms, specific IgG4 level gradually decreased. The positive rate and intensity of specific IgG4 in sera from patients with cerebral cysticercosis were consistent with the number of cysticercus parasites in the brain and pathologic changes, such as survival, disintegration, death and calcification. Survival of cysticercus in the brain was objectively evaluated using this technique. CONCLUSIONS: The determination of specific IgG4 in sera is a practical method for diagnosis and therapeutic evaluation of cerebral cysticercosis. PMID- 12133304 TI - Effect of oxLDL on the uptake and clearance rate of cholesterol in v-SMC originated from human apoA1 transgenic mice. AB - OBJECTIVE: To study the inhibition effect of oxLDL on the uptake and clearance of intra-cellular (3)H-cholesterol in v-SMC from the human-apoA1 transgenic mice (C57BL/6) and the changes in human-apoA1 mRNA expression in v-SMC from human apoA1 transgenic mice after oxLDL stimulation. METHODS: v-SMC originally isolated from human apoA1 transgenic mice connected with a recombined mouse metallothionein-I (MT-I) promoter was used, and the effect of oxLDL on the uptake and clearance of intracellular (3)H-cholesterol was studied in v-SMC of the transgenic and control mice respectively, the study of h-apoA I mRNA expression from v-SMC of the transgenic mice were done by RT-PCR and Northern blot. RESULTS: oxLDL (30 microg/ml) strongly promoted the SMC proliferation. No difference was found in (3)H-cholesterol uptake between nSMC and trSMC, and the uptake rates of both kinds of SMC rose 100% after oxLDL stimulation. The efflux rates of (3)H cholesterol in trSMC were much higher than those of nSMC (40% - 50%). After oxLDL stimulation, the clearance rates fell by 28% and 10%, respectively, for nSMC and trSMC. The result of RT-PCR and Northern blot showed that h-apoA1 gene expression was markedly increased by the stimulation of oxLDL. CONCLUSION: Expression of the h-apoA1 gene in C57BL/6 mice enables them to reduce the accumulation of cholesterol in v-SMC. The trSMC can alleviate the harmful effect of oxLDL due to the increase of h-apoA1 expression. PMID- 12133305 TI - The inherent characteristics and DNA polymorphism of Vibrio cholerae and other vibrios. AB - OBJECTIVE: To investigate the inherent characteristics of Vibrio cholerae (V. cholerae) and other vibrios and their relationship. METHODS: Polymerase chain reaction (PCR), DNA sequence analysis, randomly amplified polymorphic DNA (RAPD) analysis and average linkage cluster analysis were used to study 3 isolates of V. cholerae strains O139, three isolates O1 biotype El Tor, four isolates O1 biotype classical and 3 other vibrios. RESULTS: V. cholerae O139 contained the genomic sequences of ctx A2-B as well as V. cholerae O1. V. cholerae and others vibrios were divided into 4 groups by fingerprint patterns of RAPD, that is (1) V. cholerae O139 and V. cholerae O1 El Tor; (2) V. cholerae O1 classical; (3) V. paraheamolyticus and V. vulnificus and (4) V. flluvialis. V. cholerae O139 DNA fingerprint of RAPD was consistent with the El Tor biotype: average linkage cluster distance was 0, and slightly different from the classical biotype, with a distance of 2.07. It was much more different from vibrio paraheamolyticus and others, with a distance of 6.76 - 8.54. CONCLUSION: V. cholerae and other vibrios are polymorphic in inherent characteristics. The inherent characteristics of V. cholerae O139 are the same as El Tor biotype. O139 may have evolved from the El Tor biotype. The inherent characteristics of vibrio paraheamolyticus are the same as vibrio vulnificus. PMID- 12133306 TI - Effect of oxymatrine on murine fulminant hepatitis and hepatocyte apoptosis. AB - OBJECTIVE: To evaluate the protective effects and mechanism of action of oxymatrine (OM) on the experimental fulminant hepatitis (FH) and early hepatocyte apoptosis in murine liver tissue. METHODS: Fulminant hepatitis mice were induced by injecting lipopolysaccharide (LPS) intraperitoneally (ip) in galactosamine (GalN) sensitized mice. Two separate experiments were designed, including saline control group, fulminant hepatitis group and oxymatrine pretreated group (50 mg/kg, intraperitoneally, bid x 3 days). The levels of serum tumor necrosis factor alpha (TNFa) in mice from two experiments were determined at 5-hour and 7.5-hour after injecting galactosamine/lipopolysaccharide. Mouse liver samples at 5-hour time point were obtained for in situ end labeling (ISEL) staining and ultrastructural observation of apoptotic cells under transmission electron microscope (TEM). Liver samples at 7.5-hour time point were taken for hematoxylin eosin (HE) staining and immunohistochemical staining of Fas and its ligand (FasL). RESULTS: As compared with the fulminant hepatitis group, the levels of serum tumor necrosis factor alpha in mice from the OM pretreated group at 5-hour and 7.5-hour time point were all significantly decreased (P < 0.05 and P < 0.01 respectively). Hepatocyte apoptosis in mice at 5-hour time point was significantly inhibited (P < 0.01). Both the degree of liver injury and the degree of Fas and Fas ligand expression in the OM pretreated group were reduced remarkably (P < 0.01 and 0.05 respectively) when compared with the saline control group. CONCLUSIONS: Oxymatrine protects mice from fulminant hepatitis induced by GalN/LPS and may block hepatocyte apoptosis and subsequent necrosis through downregulating the production of serum tumor necrosis factor alpha and the expression of Fas and Fas ligand in liver tissue. PMID- 12133307 TI - Hepatitis C virus nonstructural protein NS(3) and telomerase activity. AB - OBJECTIVE: To study the effect of hepatitis C virus nonstructural protein NS(3) (HCV NS3) on telomerase activity and carcinogenesis. METHODS: Streptavidin peroxidase (SP) conjugated method was used to detect the expression of HCV NS(3) protein in NIH3T3 cells transfected with plasmid pRcHCNS(3)-5' and pRcHCNS(3)-3'. Telomerase activity was detected by an in situ telomerase activity labeling method, telomeric repeat amplification protocol polymerase chain reaction (TRAP PCR) and telomerase PCR enzyme linked immunosorbent assay (ELISA) technology in the transfected and non-transfected NIH3T3 cells. RESULTS: HCV NS(3) protein was expressed in the NIH3T3 cells transfected with plasmid pRcHCNS(3)-5' expressing HCV NS(3) C-terminal deleted protein or with plasmid pRcHCNS(3)-3' expressing HCV NS(3) N-terminal deleted protein. The positive signal of HCV NS(3) protein was localized in the cytoplasm of NIH3T3 cells, and the signal intensity of the former was stronger. Telomerase activity in NIH3T3 cells transfected with plasmid pRcHCNS(3)-5' was stronger than that in NIH3T3 cells transfected with plasmid pRcHCNS(3)-3' (P < 0.01), whereas telomerase activity in NIH3T3 cells transfected with plasmid pRcCMV or untreated NIH3T3 cells was weaker than that in NIH3T3 cells transfected with plasmid pRcHCNS(3)-3' (P < 0.05). The expression level of HCV NS(3) protein was significantly correlated with the strength of telomerase activity (P < 0.05). The results obtained by in situ telomerase activity labeling corresponded to the results by telomerase PCR ELISA technology. CONCLUSIONS: HCV NS(3) protein may activate telomerase through endogenous mechanism to induce host cell transformation. The effect of HCV NS(3) C-terminal deleted protein on telomerase activity in the host cell may be stronger than that of HCV NS(3) N terminal deleted protein. In situ telomerase activity labeling was a reliable technology for studying pathological morphology and telomerase activity in tissues and cells. PMID- 12133308 TI - Apoptosis susceptibility of tumor cells to arsenic trioxide and the inherent cellular level of reactive oxygen species. AB - OBJECTIVE: To explore the association of inherent cellular reactive oxygen species (ROS) levels with susceptibility of the tumor cells to apoptosis induction by arsenic trioxide (As(2)O(3)). METHODS: Low concentration (2 micromol/L) of As(2)O(3) was administered to two cultured leukemic cell lines, NB4 and U937, and two esophageal carcinoma cell lines, EC1.71 (also named EC/CUHK1) and EC1867, to confirm the difference in apoptosis susceptibility of NB4 versus U937 and of EC1.71 versus EC1867. Dihydrogenrhodamine 123 (DHR123), used as a ROS capture agent, was incubated with cells in the absence of As(2)O(3). Fluorescence intensity of rhodamine 123, the product of cellular oxidation of DHR123, was detected by flow cytometry and ROS was measured. RESULTS: Low concentration of As(2)O(3) induced apoptosis was more likely to occur in NB4 and EC1.71 cells than in U937 and EC1867 cells, or NB4 was more sensitive than U937, and EC1.71 more sensitive than EC1867 to As(2)O(3). The inherent cellular ROS level is higher in NB4 than in U937, and also higher in EC1.71 than in EC1867. CONCLUSIONS: The difference in cellular ROS level is positively associated with cellular susceptibility to apoptosis induction by As(2)O(3). The inherent ROS level might be important in defining apoptotic susceptibility to As(2)O(3). PMID- 12133309 TI - Evaluation of referrals for genetic investigation of short stature in Hong Kong. AB - OBJECTIVE: To establish a profile of the causes of apparently unexplained SS in genetic referral center and evaluate the current referral system. METHODS: This was a retrospective database survey on patients who were referred our clinical genetic service from 1988 - 1998 primarily because of SS. We retrieved the study population from our computer database using "short stature"as a search handle and then studied the demographic, clinical and laboratory data from their medical records. RESULTS: Three hundred and fifty-three subjects were referred for genetic evaluation of SS in 1988 - 1998. The mean age of referred subjects was 11.5 years and the female to male ratio was 7.6. All referrals had undergone cytogenetic studies to exclude chromosomal abnormalities, 19% of girls with apparently unexplained short stature had Turner syndrome; at least 47.9% of the study population were normal variants and 25% of the referrals had inadequate information for classification. CONCLUSIONS: Genetic investigation is essential in the management of patients with SS, especially for girls suspected of having Turner syndrome, in which growth hormone treatment has shown to improve final height. We also highlight the inherited causes of short stature, which were often misdiagnosed as benign familial short stature, and discussed the drawbacks of the current referral system. PMID- 12133310 TI - Comparative study of catheter-mediated gene transfer into heart. AB - OBJECTIVE: To investigate the feasibility and features of 3 catheter-mediated approaches of gene transfer into heart, including direct myocardial injection (DMI), coronary artery perfusion (CAP), and intrapericardial cavity injection (ICI). METHODS: Fifteen dogs were used, and 0.3 ml (1 x 10(9) pfu) of an adenovirus (Adex1SR LacZ) was injected into the heart by 3 methods. The dogs were killed 5 days following injection, and gene expressions in heart and liver were evaluated by histochemical analysis. RESULTS: The results showed that (1) the CAP method was relatively less damaging and induced sparse LacZ expression in the myocardium, and the gene expression was also found in both vessels within the myocardium and liver; (2) gene transfer by DMI resulted in intense LacZ expression around the injection accompanied by a local inflammatory response; (3) LacZ expression elicited by ICI was detected in either the inner surface of the parietal pericardium or epicardial surface of the heart, and also in the myocardium underlying the visceral pericardium. CONCLUSION: Three catheter mediated methods of gene transfer into the heart may be used and a reasonable approach should be chosen according to purpose. PMID- 12133311 TI - Effect of polysaccharide sulfate 916 on the production of nitric oxide in ECV304 cells induced by cytokines and H(2)O(2). AB - OBJECTIVE: To investigate the effect of polysaccharide sulfate 916 (PS916) on the production of nitric oxide (NO) in ECV304 cells induced by tumor necrosis factor alpha (TNF alpha), interleukin-1 beta (IL-1 beta) and H(2)O(2) in vitro. METHODS: Production of NO in ECV304 cells was measured by the Griess method and the proliferation of cells was tested by the MTT method. The activity of NO synthase was detected spectrophotometrically. RESULTS: Production of NO in ECV304 cells decreased after treatment with 40 ng/ml IL-1 beta and 40 ng/ml TNF alpha, but increased in the presence of H(2)O(2) 0.1 mmol/L. PS916 significantly enhanced NO production in ECV304 cells in a dose-dependent manner in the TNF alpha and IL-1 beta treated groups and decreased it in the H(2)O(2) treated group. Proliferation of ECV304 cells was inhibited by TNFalpha and H(2)O(2) and no effect was found in the IL-1 beta treated group. PS916 increased the proliferation of cells treated with TNFalpha and H(2)O(2) dose-dependently. In vitro, PS916 has no effect on the activity of NO synthase. CONCLUSION: PS916 has a protective effect on ECV304 cells exposed to IL-1 beta, TNF alpha and H(2)O(2). PMID- 12133313 TI - Hereditary hemorrhagic telangiectasia: a rare cause of long-lasting abdominal distension in an 8-year-old boy. PMID- 12133312 TI - Influence of intranasal medication on the structure of the nasal mucosa. AB - OBJECTIVE: To study the influence of intranasal medication on the structure, pathology and reversibility of the nasal mucosa to provide a basis for the feasibility of intranasal route of drug administration. METHODS: Nasal drops of gentamicin were placed in the nasal cavity of rabbits for 3, 5, 7, 14 and 28 days. After that, the drops were stopped and drugs protecting the nasomucosa were used for one and three weeks. After being sacrificed over time, the nasomucosa of the rabbit was observed under optical and electron microscopes. RESULTS: Damage to the nasal mucosa appeared to different extents with prolonged use of nasal drops. Within 3 - 7 days of applying the drug, damages to the nasal mucosa gradually appeared, and after two and four weeks, were most serious. After stopping the drug, the nasal mucosa was gradually restored. CONCLUSION: Damages of drugs to the nasal mucosa could be restored. The intranasal route of drug administration would be feasible and clinically applicable. PMID- 12133314 TI - Intraspinal endometriosis: a case report. PMID- 12133315 TI - [Relation between the serum tissue polypeptide specific antigen level and the biological demeanour of lung cancer]. AB - OBJECTIVE: To investigate the relation between the serum tissue polypeptide specific antigen and the biological demeanour of lung cancer and analyze its clinical meaning for diagnosis of lung cancer. METHODS: By ELISA, the serum TPS was tested in 69 patients with lung cancer, 20 patients with pulmonary tuberculosis or pneumonia. RESULTS: The serum TPS in patients with lung cancer (273 +/- 172) U/L was significantly higher than the with benign lesions [(115 +/- 97) U/L, P < 0.001]. TPS level was related to clinical TNM stages and histological grades. It was significantly higher with TNM III, IV stages than that with I and II stages (P < 0.001). The contents of TPS in sera of 52 patients with lymphoid node metastasis were apparently higher than those of 17 cases without metastasis (P < 0.01). And it was higher in patients with small-cell lung cancer (346 +/- 163) U/L than that with non-small-cell lung cancer [(248 +/- 165) U/L, P < 0.05]. 16 post-chemotherapy patients show a lower TPS [(178 +/- 80) U/L] than pre-chemotherapy [(252 +/- 166) U/L, P < 0.05]. TPS was related with CEA (P < 0.01), NSE (P < 0.05) and TPA (P < 0.05), but not with CYFRA 21 - 1 (P > 0.05). CONCLUSION: Serum TPS level is closely connected with TNM stages, histological grades and lymphoid node metastases and may serve as a novel marker for diagnosis of lung cancer. PMID- 12133316 TI - [Clinical analysis of 17 cases of lung carcinoid tumor]. AB - OBJECTIVE: To study the classification and treatment modalities of lung carcinoid tumor. METHODS: 17 cases of lung carcinoid tumor from May 1983 to May 1998 were reviewed and analysed. RESULTS: Lung carcinoid tumors were classified into two sub-groups: typical carcinoid tumor and non-typical carcinoid tumor. The prognosis of typical carcinoid tumor was better than that of non-typical carcinoid tumor. 3-year survival was 5/6 for typical carcinoid tumor, 4/11 for non-typical carcinoid tumor. There was no statistic significance in the treatment outcome between patients receiving operation + chemotherapy + radiotherapy and patients receiving chemotherapy + radiotherapy in non-typical carcinoid tumor. CONCLUSION: Operation should be considered as the first choice for typical carcinoid tumor that has a good prognosis. Non-typical carcinoid tumor tends to be metastatic and has a poor prognosis. Chemotherapy should be considered as the first choice and radiotherapy as palliative for non-typical carcinoid tumor. PMID- 12133317 TI - [Losartan in the rat model of bleomycin-induced pulmonary fibrosis and its impact on the expression of monocyte chemoattractant protein-1 and basic fibroblast growth factor]. AB - OBJECTIVE: To study the effect of losartan on bleomycin-induced pulmonary fibrosis in rats and its possible mechanism. METHODS: Pulmonary fibrosis was induced in Wistar rats by intratracheal instillation of bleomycin. Then the rats received daily losartan 10 mg/kg (group L), prednisone 20 mg/kg (group P), or normal saline (group B) orally. Normal controls received normal saline both intratracheally and orally. Six rats in each group were sacrificed 3, 7, 14, 28 and 56 days after intratracheal instillation. Histological changes of the lungs were evaluated by HE stain, Masson's trichrome stain and sirius red stain. Collagen content of the lung tissue was assessed by hydroxyproline concentration. The mRNAs of monocyte chemoattractant protein-1 (MCP-1) and basic fibroblast growth factor (bFGF) were detected by reverse transcription-polymerase chain reaction (RT-PCR). Lung expression of MCP-1 and bFGF proteins was assessed by immunohistochemistry and the level of MCP-1 protein was further measured by ELISA. RESULTS: Pulmonary fibrosis of group L and group P was improved as compared with that of group B. Hydroxyproline concentrations of group L and group P were lower than that of group B. Protein expression of MCP-1 and bFGF in group L was lower than that in group B. CONCLUSION: Losartan alleviates bleomycin induced pulmonary fibrosis in rats. Inhibiting the expressions of MCP-1 and bFGF in lung tissues may be one of the mechanisms. PMID- 12133318 TI - [The effect of dipyridamole and adenosine on pulmonary fibrosis in mice]. AB - OBJECTIVE: To evaluate the effect of dipyridamole (DPM) and adenosine (ADO) on interstitial pulmonary fibrosis in mice. METHODS: 100 mice injected with Bleomycin (BLM, 8.5 mg/kg) by intratracheal were divided into a control group and groups treated by DPM, ADO, DPM and theophyline (TH). Pulmonary pathology from day 4 to day 30 observations was studied by light and electron microscope. RESULTS: Alveolar epithelial cell necrosis, lung tissue lesions by hypoxia and interstitial fibrosis were found in the BLM group after day 10. The necrotic epithelial cell, fibroblast proliferation, and the hypoxia induced tissue damage were less prominent, and the number and phagocytic function of macrophages were increased in the DPM and ADO treated groups. In the TH treated group, necrotic epithelial cells increased, the lung tissue was ischemic, and the differentiation of monocytes in pulmonary interstitial was slower. CONCLUSION: DPM and ADO can inhibit fibroblast proliferation by improving microcirculation and promoting macrophage development and alveolar space reconstruction. PMID- 12133320 TI - [The effect of triptolide on the proliferation of airway smooth muscle and the expression of c-fos and c-jun in asthmatic rats]. AB - OBJECTIVE: To study the effect of triptolide on airway smooth muscle (ASM) proliferation and expressions of c-fos and c-jun in asthmatic rats. METHODS: 70 healthy male sprague-dawley rats were randomly divided into seven groups of 10 each. The asthmatic model was established by ovalbumin inhalation and injection. The mRNA expression of c-fos and c-jun were examined by RT-PCR and their protein expression by immunohistochemical method. The airway wall thickness, the ASM thickness, the number of ASM nucleus and pulmonary tissue changes were observed with microscope. RESULTS: c-fos and c-jun mRNA and protein expressions of asthma groups A and five therapy groups (C, D, E, F and G) were significantly higher than those in the control group B (P < 0.01). The expressions were lower in group C, D, E, F and G than those in group A (P < 0.01). There was no significant difference in the expressions among group C, D, E, F and G (P > 0.05). Triptolide decreased ASM proliferation pathologically in asthmatic rats. CONCLUSIONS: ASM proliferation is a feature of asthma. c-fos and c-jun may promote ASM proliferation in asthma. The remarkable inhibition of ASM proliferation by triptolide was probably due to its inhibitory effect on the expression of c-fos and c-jun. PMID- 12133319 TI - [Nitric oxide modulates expression of matrix metalloproteinase in asthmatic rats]. AB - OBJECTIVE: To observe the effect of nitric oxide on expression of matrix metalloproteinase and tissue inhibitor of metalloproteinase in asthmatic rats. METHODS: Thirty healthy male Wistar rats were randomly divided into control (n = 10), asthmatic (n = 10) and L-arginine (n = 10) groups. Lung tissues were sliced and stained with HE. The following morphometric parameters were measured by image analysis system: airway wall basement membrane perimeter (Pbm), total bronchial wall area (WAt), the inner wall area (WAi) and the area of smooth muscle (WAm). Levels of nitrites/nitrates and NOS activity in the lungs were measured by commercial NO and NOS kits. Transcriptional levels of MMP-2 and TIMP-1 mRNA in the lungs were assessed by semiquantitative reverse transcription-polymerase chain reaction (RT-PCR). RESULTS: (1) WAt/Pbm, WAi/Pbm and WAm/Pbm in the L arginine group were (35.1 +/- 2.6) microm(2)/microm, (25.3 +/- 2.0) microm(2)/microm, and (8.7 +/- 1.5) microm(2)/microm, respectively. WAt/Pbm, WAi/Pbm and WAm/Pbm in the asthmatic group were (25.3 +/- 2.1) microm(2)/microm, (20.4 +/- 2.3) microm(2)/microm, and (4.2 +/- 2.0) microm(2)/microm, respectively. WAt/Pbm, WAi/Pbm and WAm/Pbm in the control group were (20.8 +/- 1.3) microm(2)/microm, (15.3 +/- 2.1) microm(2)/microm, and (3.1 +/- 1.1) microm(2)/microm, respectively. The differences among three groups were statistically significant. (2) Levels of nitrites/nitrates in the L-arginine group [(11.8 +/- 1.7) nmol/mg] and the asthmatic group [(7.2 +/- 2.1) nmol/mg] were significantly higher than that in the control group [(3.1 +/- 1.2) nmol/mg]. NOS activity in the lungs was (16.5 +/- 1.3) U/mg L-arginine group, (10.8 +/- 1.4) U/mg asthmatic group, and (1.46 +/- 0.98) U/mg control group, respectively, the differences being statistically significant between groups. (3) MMP-2 and TIMP-1 mRNA levels in the L-arginine group and the asthmatic group were significantly higher than that in the control group [L-arginine group (0.82 +/- 0.11), (0.51 +/- 0.12); asthmatic group (0.68 +/- 0.14), (0.56 +/- 0.10); control group (0.14 +/- 0.03), (0.11 +/- 0.05), respectively]. The difference in the MMP 2 mRNA levels was significant between the L-arginine group and the asthmatic group (P < 0.05), but the difference in the TIMP-1 mRNA levels between the two groups was not statistically significant. (P > 0.05). (4) A significant positive correlation was found between the MMP-2 mRNA levels and nitrites/nitrates levels in L-arginine group and asthmatic group (r(s) = 0.65, 0.68, P < 0.05), but there was no significant correlation between the nitrites/nitrates levels and the TIMP 1 mRNA levels in the two groups (r(s) = 0.23, 0.18, P > 0.05). CONCLUSION: Nitric oxide may contribute to airway inflammation and remodeling by influencing the balance between matrix metalloproteinases and tissue inhibitors of metalloproteinases. PMID- 12133321 TI - [The role of transforming growth factor-beta(1) in smoking-induced chronic bronchitis and emphysema in hamsters]. AB - OBJECTIVE: To study the role of transforming growth factor-beta(1) (TGF-beta(1)) in the pathogenesis of chronic bronchitis and emphysema. METHODS: An animal model of chronic bronchitis and emphysema was developed in hamsters by chronic smoke inhalation. The expression of TGF-beta(1) mRNA and protein in the pulmonary tissue was observed. The bronchial epithelia was stimulated with cigarette smoke extract (CSE) in vitro and the expression of TGF-beta(1) was measured. RESULTS: 3 months after smoking, the animals developed chronic bronchitis and emphysema. TGF beta(1) immunoreactivity in the pulmonary tissue and cultured bronchial epithelia increased significantly as compared to the normal control (2.75 +/- 0.23 vs 0.84 +/- 0.39, P = 0.001 and 2.67 +/- 0.16 vs 0.85 +/- 0.54, P = 0.001). The expression of TGF-beta(1) mRNA was also increased in the animal model (1.28 vs 0.98). CONCLUSIONS: Smoking can induce over-expression of TGF-beta(1) in bronchial epithelia, which may be one of the mechanisms for smoking-induced chronic bronchitis and emphysema. PMID- 12133322 TI - [Effect of ipratropium bromide on calcium activated potassium channel in tracheal smooth muscle cells from chronically hypoxic rats]. AB - OBJECTIVE: To study the effects of ipratropium bromide on large conductance calcium activated potassium (BK(Ca)) channel in tracheal smooth muscle cells (TSMC) from chronically hypoxic rat. METHODS: The chronically hypoxic rat model was established. Single TSMC was acutely isolated. Single channel currents were recorded by using the patch clamp technique in inside-out configuration. RESULTS: (1) Chronic hypoxia decreased BK(Ca) channel activity significantly in inside-out recording. Channel open probability (P(0)) of BK(Ca) channel in the chronically hypoxic rats reduced significantly compared with that in the control rats (0.17 +/- 0.07 for the hypoxia, 0.35 +/- 0.10 for the control, n = 30, t = 8.22, P < 0.01). Channel open time constants were significantly shortened in the chronically hypoxic group. The fast and slow open time constants (tau(O1), tau(O2))were significantly different from those in the control [tau(O1) (1.49 +/- 0.41) ms, (0.53 +/- 0.23) ms, respectively, n = 30, t = 12.07, P < 0.01; tau(O2) (11.9 +/- 3.2) ms, (3.8 +/- 1.4) ms, respectively, n = 30, t = 12.60, P < 0.01]. Closing time prolonged significantly. The fast and slow closing time constants (tau(c1), tau(c2)) were significantly different from those in the control [tau(c1) (2.7 +/- 0.9) ms, (5.7 +/- 1.5) ms, respectively, n = 30, t = 8.71, P < 0.01; tau(c2) (12.1 +/- 2.3) ms, (19.4 +/- 2.9) ms, respectively, n = 30, t = 14.05, P < 0.01]. Ipratropium bromide and salbutamol reversed the effect of chronic hypoxia on BK(Ca) channel. P(0) increased, tau(O1) and tau(O2) were prolonged significantly. tau(c1) and tau(c2) were shortened [P(0), 0.15 +/- 0.04, 0.28 +/- 0.09, 0.30 +/- 0.08, respectively, n = 25, F = 39.90, P < 0.01; tau(O1) (0.55 +/- 0.24) ms, (0.89 +/- 0.25) ms, (1.03 +/- 0.33) ms, respectively, n = 25, F = 15.32, P < 0.01; tau(O2) (3.6 +/- 1.4) ms, (6.3 +/- 1.9) ms, (6.9 +/- 2.0) ms, respectively, n = 25, F = 40.10, P < 0.01; tau(c1) (6.1 +/- 1.6) ms, (3.3 +/- 1.2) ms, (3.0 +/- 0.8) ms, respectively, n = 25, F = 57.14, P < 0.01; tau(c2) (20.1 +/- 2.5) ms, (12.4 +/- 2.6) ms, (13.0 +/- 2.0) ms, respectively, n = 25, F = 24.60, P < 0.01]. CONCLUSION: Chronic hypoxia decreased BK(Ca) channel activity. Ipratropium bromide and salbutamol reversed the effect of chronic hypoxia on BK(Ca) channel. The relaxing effect of ipratropium bromide and salbutamol on TSMC may be partly mediated via activation of BK(Ca) channel. PMID- 12133323 TI - [The effect of interleukin-12 on the production of Th(1) and Th(2) cytokines by peripheral blood mononuclear cells from patients with tuberculosis]. AB - OBJECTIVE: To investigate the effect of IL-12 on the production of IFN-gamma and IL-4 by peripheral blood mononuclear cells (PBMC) from patients with tuberculosis. METHODS: PBMCs were isolated and cultured with RPMI 1640, PPD, PPD + rhIL-12, PPD + anti-IL-12, respectively. The level of IFN-gamma and IL-4 was detected with ELISA. RESULTS: Compared with the PPD group, IL-12 increased the production of IFN-gamma (321.6 +/- 87.7 vs 452.5 +/- 111.4, 387.0 +/- 70.8 vs 515.4 +/- 44.1, respectively), but decreased the production of IL-4 (54.6 +/- 11.0 vs 41.3 +/- 13.5, 55.1 +/- 9.5 vs 38.2 +/- 12.9, respectively) in the patients and the healthy individuals; anti-IL-12 blocked this effect. The level of IFN-gamma was lower in tuberculosis patients than that in the healthy controls. The IL-4 level was not different between the two groups (P > 0.05). The levels of IFN-gamma and IL-4 were lower in recurrent tuberculosis than those in the newly-diagnosed cases (P < 0.05). CONCLUSIONS: IL-12 induced IFN-gamma production and inhabited IL-4 expression, modulating the balance between TH(1) and TH(2) cytokines, suggesting that IL-12 enhances the host defense against tuberculous infection. IL-12 may be used for treatment of tuberculosis and vaccine development. PMID- 12133324 TI - [Electroencephalogram spectral power analysis of obstructive sleep apnea syndrome patients before and during continuous positive airway pressure therapy]. AB - OBJECTIVE: To investigate the instant effects of continuous positive airway pressure (CPAP) on EEG spectral power changes in obstructive sleep apnea syndrome (OSAS) patients. METHODS: 26 OSAS patients were included. The diagnosis was made by polysomnography (PSG). The CPAP pressure was titrated during the first night of therapy. During another night in two weeks after the diagnostic study, the patients accepted the whole night CPAP treatment while PSG was monitored. C(3)/A(2) was analyzed by using fast fourier transform (FFT). Spectral edge frequency (SEF), median power frequency (MPF), alpha index, beta index, delta index and theta; index during CPAP therapy were compared with those parameters before treatment. RESULTS: Sleep architecture was improved significantly during therapy. The ratio of slow wave sleep (SWS) deficiency reduced from 19/26 to 10/26 (P = 0.0250). There were more times of REM sleep (1.81 +/- 0.25) vs (2.65 +/- 0.17) (P = 0.023). A significant increase of total SWS time over total sleep time (TST) (2.9 +/- 1.1)% vs (6.0 +/- 1.2)% (P = 0.043) was observed, as well as total REM sleep time over TST (12.0 +/- 1.7)% vs (21.1 +/- 1.6)% (P = 0.001). Total stage I and stage II time over TST reduced greatly [from (85.1 +/- 2.1)% to (73.0 +/- 1.9)% (P = 0.000)]. So did the number of wake after sleep onset [from (12.2 +/- 1.3) to (9.4 +/- 1.0) (P = 0.033). The total number of stage I, stage II decreased from (46.4 +/- 4.2) to (36.7 +/- 2.4), but the difference was not significant (P = 0.051). Spectral analysis of EEG showed a decrease of mean SEF in total sleep period (TSP) (14.4 +/- 0.4) Hz vs (13.6 +/- 0.3) Hz (P = 0.003), stage I (15.4 +/- 0.4) Hz vs (14.8 +/- 0.5) Hz (P = 0.040), stage II (13.7 +/- 0.3) Hz vs (12.8 +/- 0.3) Hz (P = 0.007) and REM sleep (15.0 +/- 0.5) Hz vs (13.8 +/- 0.6) Hz (P = 0.028), as well as 0.018 and 0.047]. delta index was increased in stage II sleep (P = 0.030) but not in other sleep stages. CPAP had no obvious influence on theta; index (P > 0.05). The median of SEF in TSP (14 vs 13) Hz (P = 0.0056), stage I (16 vs 15) Hz (P = 0.04) and stage II (13 vs 13) Hz (P = 0.002). Changes of MPF were not as significant as that of SEF, but mean MPF was decreased during TSP and stage IV. CPAP significantly reduced mean beta index [from (8.4 +/ 0.5)% to (7.5 +/- 0.5)% (P = 0.012)] in total sleep period as well as in stage I, II, III and IV sleep (P < 0.015). alpha index in TSP, stage III and IV sleep was decreased significantly (P = 0.045, 0.018 and 0.047 respectively). delta index was increased in stage II sleep (P = 0.030) but not in other sleep stages. CPAP had no obvious influence on theta; index (P > 0.05). CONCLUSIONS: CPAP has an acute effect in improving the sleep architecture and EEG power spectrum. The SEF, beta index and alpha index are more sensitive than MPF, delta index and theta; index to CPAP therapy. These changes may be related to the recovery of cerebral function in OSAS patients after CPAP therapy. More studies need to be conducted to investigate how these changes happen and their relation to daytime cerebral function. PMID- 12133325 TI - [Study of serum leptin level in patients with obstructive sleep apnea]. AB - OBJECTIVE: To explore the change of serum leptin in patients with obstructive sleep apnea syndrome (OSAS). METHODS: Polysomnography was performed in 58 patients with OSAS and in 21 simple obese controls without differences in age and body mass index (BMI). Serum leptin level was measured by radioimmunoassay. RESULTS: (1) The serum leptin level in patients with OSAS was higher, in both males (6.1 +/- 1.7) microg/L and females (19.5 +/- 9.9) microgram/L, than that in the simple obese controls [male (4.5 +/- 1.7) microgram/L, female (10.5 +/- 2.4) microgram/L] (P < 0.01, P < 0.05). (2) Serum leptin levels were significantly correlated with BMI in both OSAS patients (r = 0.64, P < 0.01) and simple obese controls (r = 0.59, P < 0.01), but only in OSAS patients was the leptin level significantly correlated with apnea and hypopnea index (AHI) (r = 0.47, P < 0.01) and with neck circumference (r = 0.64, P < 0.01). CONCLUSIONS: Serum leptin levels were higher in OSAS patients than in simple obese controls. Besides obesity and neck circumference, OSAS may also influence the leptin system, resulting in increased serum leptin level. PMID- 12133326 TI - [The expression of erbB/HER family in lung cancer]. AB - OBJECTIVE: To study the expression of human epidermal growth factor receptor (HER) family in lung cancer. METHODS: Paraffin-embedded tissues from 70 cases of lung cancer (43 cases of squamous cell cancer and 27 cases of adenocarcinoma) and 20 cases of non-cancerous lung diseases were stained for the expression of HERs by means of immunohistochemistry. RESULTS: All HERs were weakly expressed in non cancerous lung tissues; over-expression of HER(1), HER(2), HER(4), HER(1 + 2), HER(1 + 4) and HER(2 + 4) and HER(1 + 2 + 4) was found in lung cancer and correlated with lymph node metastasis, TNM staging and post-operation survival. CONCLUSIONS: erbB(1), erbB(2) and erbB(4) are the genes regulating the growth of lung cancer at advanced stages. Targeting the HER(1), HER(2) and HER(4) over expression might be a new approach to the treatment of lung cancer at advanced stages. PMID- 12133327 TI - [The correlation between DNA content and Fas antigen in non-small cell lung cancer]. AB - OBJECTIVE: To study the relationship between Fas protein expression and DNA content in non-small cell lung cancer (NSCLC). METHODS: Fas protein expression and DNA content in resected samples from 33 NSCLC patients and 17 normal controls were detected by flow cytometry. Fas expression and DNA content were compared with cell proliferation activity which was defined by proliferation index and s phase fraction (SPF), and with clinicopathological parameters. RESULTS: The percentage of DNA aneuploidy in NSCLC was 82%. The DNA index (DI), PI and SPF in NSCLC were 1.36 +/- 0.23, 0.29 +/- 0.09 and 0. 144 +/- 0.078 respectively, which were significantly higher than those in normal lung tissue. The Fas protein expression was lower in NSCLC than that in normal tissue (P = 0.000) and showed negative correlation with PI (P = 0.024) and SPF (P = 0.004). Fas expression and the cell proliferation activity were not correlated to the clinicopathological parameters including age, tumor size, lymph node metastasis and the histological types of lung cancer (P > 0.05). CONCLUSIONS: Decrease of Fas protein expression and increase of proliferation activity are common in NSCLC. The reduction of Fas protein expression may affect the proliferation activity of NSCLC. This abnormal regulation of apoptosis and proliferation may play an important role in the pathogenesis of NSCLC. PMID- 12133328 TI - [Surgical treatment and prognosis of pulmonary metastasis]. AB - OBJECTIVE: To study the surgical treatment and the prognosis of pulmonary metastasis. METHODS: 59 patients in the operative group and 36 patients in the control group were included. Their clinical data were collected by follow-up investigation and the prognosis was analyzed by stepwise regression. RESULTS: The median survival time was 20.2 (operative group) and 10.0 (control group) months. The 1, 3 and 5-year survival rates in the two groups were 70%, 39%, 34% (operative group) and 28%, 8%, 0% (control group) respectively. The 5-year survival rate of patients with disease-free interval (DFI) < 1 year was 21%, while that of patients with DFI >/= 1 year was 40%. CONCLUSIONS: Resection of pulmonary metastasis should be considered if indicated. DFI is an important prognostic factor for pulmonary metastasis. PMID- 12133329 TI - [The pathology of and the cardio-pulmonary functional changes in acute multiple pulmonary microthromboembolism in a canine model]. AB - OBJECTIVE: To evaluate the effect of acute multiple pulmonary microthromboembolism on circulation, ventilation, lung histology, and the coagulatory and fibrinolytic functions. METHODS: Thirteen dogs were studied, 7 in the embolism group and 6 in the control group. 10 ml of blood was withdrawn from each animal. Thrombi were formed and cut into pieces of 1 mm to 2 mm in size. The thrombi were counted, and 300 thrombi were infused at one time back to the same animal. PAP, PAWP, VO(2), DO(2), D-dimer, protein C, protein S were measured before and immediately, 30 min, 1 hour, 2 hour after embolism. RESULTS: PAP was increased immediately after embolisation, and then decreased at 1 hour. The level of D-dimer was elevated at an early stage. Pulmonary hemorrhage, consolidation and microembolism were observed. CONCLUSIONS: Multiple microthromboembolism caused lung injury and thrombosis. PAP was increased at the early stage, but ventilation was not affected. The level of D-dimer changed at the early stage of thromboembolism. PMID- 12133330 TI - [Evaluation of the relationship between deep venous thrombosis and pulmonary embolism with radionuclide techniques]. AB - OBJECTIVE: The purpose of this study was to evaluate the relation between deep venous thrombosis (DVT) and pulmonary thromboembolism (PTE) by radionuclide imaging. METHODS: One hundred forty patients with PTE from September 1997 to March 2001 at this institution was confirmed by clinical manifestation, pulmonary perfusion (PPI)/ventilation scan (PPV) and deep venous radionuclide venography (RNV), which were performed in all patients. There were 79 males and 61 females, with an average age of 39 +/- 18 years. Twenty-six cases underwent pulmonary angiography; 11 underwent X Ray venography of lower extremities (XRV); 18 underwent impedence plethymography (IPG); and 36 underwent lower limb echocardiography (UCG). RESULTS: Of the 140 patients with PTE, 120 (85.7%) had lower limb venous pathological changes. Among them, 94 patients had risk factors for DVT. The agreement rates of RNV with XRV, UCG and IPG were 90.9%, 72.2% and 80.0%, respectively. CONCLUSIONS: The results indicated that DVT was highly prevalent in patients with acute pulmonary embolism. Thrombosis was a primary pathogenic factor for PTE, and thrombi were mostly from proximal veins. (99m)TC MAA radionuclide imaging was a useful method for noninvasive detection of DVT and PTE. PMID- 12133331 TI - [Drug resistance of Mycobacterium tuberculosis in a nationwide epidemiological survey in China in the year of 2000]. AB - OBJECTIVE: To determine the state of anti-tuberculosis drug resistance, and to analyze the trend of drug resistance rate and assess its impact on National TB Programme (NTP). METHODS: 392 Mycobacterium tuberculosis (MTB) isolates from a nationwide epidemiological survey were subjected to susceptibility testing against 6 anti-tuberculosis drugs using the absolute concentration method. RESULTS: The resistant rate to any drug was 27.8% in all MTB isolates, 18.6% in 263 new cases and 46.5% in 129 previously treated cases. Multi-drug resistant rate (MDR) was 10.7%. Initial and acquired MDR was 7.6% and 17.1%. Any resistance to INH, SM, RFP, PAS-Na, EMB and TB(1) was 17.6%, 17.3%, 16.6%, 2.8%, 1.5% and 1.3%, respectively. The resistant rates to 1, 2, 3 and 4 or more drugs were 7.6%, 4.6%, 4.6% and 1.9% in new cases and 14.7%, 19.4%, 10.1% and 2.3% in previously treated cases. The initial drug resistant rate did not show significant difference (P > 0.05) among different sex and age groups. CONCLUSIONS: Compared to the previous data, the total drug resistance and initial drug resistance decreased significantly, but were still high. More attention should be paid to the trend of resistance of MTB to first-line drugs and especially the high incidence of MDR-TB. PMID- 12133332 TI - [Prognostic value of CLIP score system for patients with resection of hepatocellular carcinoma]. AB - OBJECTIVE: To evaluate the prognostic value of CLIP score system for patients with resection of HCC. METHODS: A retrospective survey was carried out in 174 patients undergoing resection of HCC from January 1986 to June 1998. 153 of 174 patients with curative resection were followed up for at least three years. Disease-free survival rate was defined as the time relapsed from the date of image diagnosis and either the date of death or the date of the latest follow-up visit, with final evaluation at June 30, 2001. Recurrences were classified into early ( 3 year) recurrence. Risk factors for recurrences and prognostic factors for survival in each group were analyzed by the chi-square test, the Kalain-Meier estimation and the COX proportional hazards model respectively. RESULTS: The 1-, 3-, 5-, 7-, and 10-year cumulative disease free survival rates were 57.2%, 28.3%, 23.5%, 18.8% and 17.8%, respectively. The associated factors with early recurrence were as fellows: tumor size > 5 cm, microsatellite, venous invasion, tumor morphology, tumor extension, advanced TNM stages, CLIP scores, radical resection, and resection margin, respectively. But both CLIP scores and Child stage were associated with late recurrence. Univariate survival curves analysis expressed that Child grades, radical resection, resection margin, tumor size, microsatellite, venous invasion, tumor morphology, tumor extension, TNM stages, and CLIP scores were associated with prognosis. The multivariate analysis by COX proportional hazards model, the independent prognostic factors for survival were radical resection, resection margin, and TNM stages. CONCLUSIONS: CLIP score, which takes into account both liver function and tumor extension, has displayed a unique superiority in predicting the tumor early and late recurrence and prognosis. It could be an useful tool in predicting the patient recurrence and prognosis with resection of HCC. Meanwhile, it may help physicians to decide the more appropriate management in advance for patients with HCC. PMID- 12133333 TI - [Orthotopic liver transplantation with no veno-venous bypass]. AB - OBJECTIVE: To assess the feasibility and outcome of orthotopic liver transplantation (OLT) with no veno-venous bypass (VVB) in adult patients. METHODS: Between 1999 and June 2001, 43 adult patients were subjected to orthotopic liver transplantations with veno-venous bypass (28), or no veno-venous bypass (15). RESULTS: There was no significant difference in mean serum creatinine on day 3 and gas discharge time in patients with veno-venous bypass or not. With no veno-venous bypass, the average operative time was 5.6 +/- 1.4 h, median amount of blood loss during operation was 4 200 +/- 850 ml, median amount of blood transfused intraoperatively was 4 800 +/- 920 ml, and median intensive care unit stay was 6.3 days. All these were lower or shorter than those of the patients with veno-venous bypass. CONCLUSIONS: Orthotopic liver transplantation with no veno-venous bypass is safe and can be performed in the majority of adult patients. Liver transplantation with no veno-venous bypass is associated with shorter total operating time, lower blood product usage, and shorter intensive care unit stay compared with standard technique of OLT with routine use of VVB. PMID- 12133334 TI - [The effect of postoperative transcatheter hepatic arterial chemoembolization on disease-free survival after hepatectomy for hepatocellular carcinoma]. AB - OBJECTIVE: To evaluate postoperative transcatheter hepatic arterial chemoembolization (TACE) for improving the disease-free survival of HCC patients after hepatectomy. METHODS: 1 725 HCC patients were followed up after hepatectomy retrospectively. Of 1 457 patients who were followed-up completely, 209 had postoperative TACE. The 1 457 patients were divided into ten groups according to tumor thrombus, small HCC, capsular invasion, vascular invasion and cirrhosis. The disease-free survival was analyzed between subgroups of weather postoperative TACE was performed in every group. Software SAS 6.12 and EGRET package were used. Kaplan-Meier estimation was used to calculate the disease-free survival rates. RESULTS: There were no difference of the disease-free survival between the subgroups in no capsular invasion and in no hepatic cirrhosis groups. CONCLUSION: Postoperative TACE is helpful in improving the disease-free survival of HCC patients after hepatectomy except those with integrated tumor envelope or no hepatocirrhosis. It is important to prolong the long-term results of operation. PMID- 12133335 TI - [Prevention and treatment of complications after hepatectomy]. AB - OBJECTIVE: To study the prevention and treatment of complications after hepatectomy. METHODS: From January 1998 to December 1999, 1 762 patients with pathologically proven primary liver cancer underwent hepatectomy. The types of resection included lobectomy, segmental resection and local hepatectomy. RESULTS: The total complication rate was 4.09% and the total mortality was 0.40%. The rates of intraoperative bleeding, postoperative bleeding, hepatic failure, stockpiling fluid of the pleural cavity, residual fluid under the diaphragm, bile leakage and infection of incision were 0.96%, 0.28%, 0.51%, 1.87%, 0.17%, 0.17% and 0.11% respectively. The mortality of the former three complications were 0.06%, 0.06% and 0.28% respectively. CONCLUSIONS: The complications can be prevented effectively and the mortality can be decreased markedly after hepatectomy by strict control of the indications, sufficient preoperative preparation, better surgical skills and close observation of postoperative state and comprehensive postoperative treatment. PMID- 12133336 TI - [Infiltration of dendritic cells and lymphocytes in hepatocellular carcinoma tissue]. AB - OBJECTIVE: To explore the clinical significances of dendritic cells and lymphocytes infiltration in hepatocellular carcinoma (HCC) tissue. METHODS: Clinicopathological data were collected from 44 patients with HCC who had under/gone curative tumor resection in our hospital. Immunohistochemical staining was used to detect the infiltration of dendritic cells in the tumor tissue, and lymphocytes infiltration was assessed simultaneously. The correlation between the infiltration of dendritic cells and lymphocytes and postoperative tumor recurrence and survival rate was analyzed. RESULTS: Tumor recurrence was markedly late in patients with dendritic cells count >/= 20 and positive lymphocytes infiltration (group A, n = 17) as compared with those who did not meet both criteria simultaneously (group B, n = 27), with a median interval of 21.6 months for group A and 4.1 months for group B (U value = 105.5, P = 0.009). The 1-, 3-, 4-year survival rates were significantly greater in group A than in group B; they were 83.5% vs. 42.2%, 61.8% vs. 28.4% and 48.7% vs. 23.0%, respectively (Log rank = 7.68, P < 0.01). CONCLUSION: The infiltration of dendritic cells and lymphocytes in HCC tissue, as an independent prognostic factor, was closely related to postoperative prognosis. PMID- 12133337 TI - [Long-term follow-up of Hirschsprung's disease patients treated by heart-shaped anastomosis operation]. AB - OBJECTIVE: To summarize the experience of heart-shaped anastomosis operation in patients with Hirschsprung's disease. METHODS: Hirschsprung's disease treated by heart-shaped anastomosis, improvement of surgery procedure, and complications were reviewed retrospectively. RESULTS: Of 193 cases, 152 completed follow-up. Early complications included urine retention (2 cases), enteritis (10), anastomosis stricture (1), and intestinal obstruction (2). Late complications (22 cases) included adhesive intestinal obstruction (2), constipation (5), incision hernia (2), enteritis (6), and occasionally stool stains (7). Neither infection in celiac, pelvic cavity and wound nor incontinence or death occurred in all patients. CONCLUSION: Heart-shaped anastomosis procedure can effectively reduce the complications ceased by Hirschsprung's disease operation and is superior to other procedures. PMID- 12133338 TI - [Sentinel lymph node biopsy in breast cancer]. AB - OBJECTIVE: To evaluate the accuracy of sentinel lymph node biopsy (SLNB) to predict the axillary lymph node status in breast cancer patients and its clinical significance. METHODS: Seventy patients with clinical TNM status T(1 - 2)N(0)M(0) underwent sentinel lymph node biopsy using Tc-99m sulfur colloid radiotracer and gamma probe, which was followed by standard axillary dissection. SLNB was compared with standard axillary dissection for its ability to reflect the final pathological status of the axillary nodes. The SLNs that were tumor negative in conventional HE staining were further evaluated using immunohistochemical stains for CK8, CK19 and KP-1 antibodies. RESULTS: The sentinel lymph node (SLN) was successfully identified in 67 (95.7%) out of 70 patients. The number of sentinel nodes harvested ranged from 1 to 5 (average 1.6). The nonsentinel nodes ranged from 5 to 20 (average 12.3). Of the 67 patients, 29 (43.3%) had histologically positive axillary lymph nodes. SLN was positive in 24 patients with metastasis (35.8%), and in 7 patients without metastasis (10.4%). In 5 patients, SLN was negative for tumor with positive nodes. The accuracy of sentinel lymph node biopsy to predict the axillary lymph node status was 92.5% and the false negative rate was 7.5%. For tumors with diameter less than or equal to 2 cm, the accuracy was 100%. 65 SLNs that were negative for HE stain were also non-reactive to immunostain for CK8 and CK19 antibody. CONCLUSIONS: SLNB can accurately predict the axillary lymph node status in most of breast cancer patients. The accuracy is about 100% in patients with T(1) lesions. Immunohistochemical staining at the same level of HE stain can not increase the detection of lymph node micrometastasis. PMID- 12133339 TI - [Surgical treatment of coronary artery aneurysm]. AB - OBJECTIVE: To describe the diagnosis and surgical management of coronary artery aneurysm. METHODS: Between October 1996 and May 2000, 6 patients with coronary artery aneurysm underwent surgical treatment. Of these patients, 3 had Kawasaki disease and 3 coronary artery fistula. All patients underwent coronary artery bypass grafting. Three patients had aneurysms resected and fistulous ostium closed. One patient received aortic valve replacement. RESULTS: There were no deaths and later death, nor major complications during the hospital stay. The results of follow-up were satisfactory. CONCLUSIONS: Coronary artery aneurysm is rare and its prognosis is poor. Early diagnosis and operation are necessary. The aneurysm should be resected, the coronary artery should be bypassed, and other associated diseases should be treated properly. PMID- 12133340 TI - [Analysis of complications after cardiac valve replacement: report of 702 patients]. AB - OBJECTIVE: To define the determinants of perioperative death and complications after cardiac valve replacement in 702 patients. METHODS: Clinical data of the patients after cardiac valve replacement were analyzed retrospectively. RESULTS: Perioperative mortality and morbidity correlated significantly with some of the perioperative variables, such as higher NYHA functional class (III or IV), large left ventricular end-diastolic diameter (>/= 70 mm), C/T >/= 0.70, prolonged aortic cross-clamping time and cardiopulmonary bypass time, unsatisfactory myocardial protection. CONCLUSIONS: Perioperative mortality and morbidity correlate significantly with some of perioperative variables, such as higher NYHA functional class, unsatisfactory myocardial protection, inappropriate surgical procedure, improper therapy of some complications after cardiac valve replacement. To avoid the occurrence of these independent predictors or to correct them timely might effectively decrease the perioperative mortality and morbidity after cardiac valve replacement. PMID- 12133341 TI - [Non-specific esophageal motility disorders: manometric abnormalities with unknown causes]. AB - OBJECTIVE: To study a group of patients with abnormalities of esophageal motility in manometric investigation. METHODS: From 1990 to 1999, 14 patients with dysphagia (9), chest pain (3), or both (2) were studied. All patients denied symptoms of heartburn, regurgitation, odynophagia, epigastric discomfort, and investigation failed to show any evidence of ischemic heart disease. A perfused catheter with 4 separate lumens was used and connected to output transducers (Medtronic, PC Polygraf HR). RESULTS: Of the 14 patients, 11 had motor disorders of the esophageal body including simultaneous contractions without normal peristalsis (5), alternative occurrence of simultaneous contractions and normal peristalsis (3), aperistalsis (2) and very low amplitude peristalsis (1). Seven patients were diagnosed with motility disorders of the lower esophageal sphincter including incomplete relaxation or no relaxation on swallowing (6), short relaxation duration (1). Four patients had more than one abnormal manometric findings. CONCLUSIONS: Non-specific esophageal motility disorder is not a real diagnostic entity, but only a group of manometric abnormalities. The relationship between the symptoms of the patients and the manometric findings was analysed. It is uncertain that these disorders have a common etiology. The revision of these abnormalities is difficult because the pathogenesis is unknown. PMID- 12133342 TI - [Clinical characteristics of chronic subdural hematoma evolving from traumatic subdural effusion]. AB - OBJECTIVE: To probe into the incidence, mechanism and clinical characteristics of chronic subdural hematoma (CSDH) evolving from traumatic subdural effusion (TSE). METHODS: The clinical materials of 32 patients with CSDH evolving from TSH were analyzed retrospectively and the correlative literature was reviewed. RESULTS: 16.7% of the patients with TSH evolved into CSDH. The time of evolution was 22 - 100 days after head injury. All patients were cured with hematoma drainage. CONCLUSIONS: TSE is one of the origins of CSDH. The clinical characteristics of TSE evolving into CSDH include polarization of patient age, and chronic small effusion. The patients are usually injured deceleratedly and accompanied with mild cerebral damage. PMID- 12133343 TI - [Allograft fibula in treatment of cervical spondylosis]. AB - OBJECTIVE: To evaluate the efficacy of allograft fibula in anterior cervical fusion for cervical spondylosis patients treated by Smith-Robinson operation supplemented with anterior instrumentation. METHODS: The clinical outcome of 38 patients with cervical spondylosis treated by Smith-Robinson operation using allograft fibula supplemented with anterior titanium plate were retrospectively studied. The patients were followed up on average was (9.5 +/- 3.4) months. The average preoperative and postoperative JOA scores were assessed and myelopathy severity was graded using the Nurick myelopathy grading system. Lateral views in neutral position, in flexion, and in extension of preoperative cervical roentgenograms were analyzed in comparison with last follow-up films to identify the changes in the height of intervertebral space and the quality of fusion. RESULTS: Statistical analysis of all patients revealed mean JOA scores of 12.54 +/- 1.62 and 16.07 +/- 1.13 before surgery and at final examination (P < 0.05), respectively. And the mean Nurick grades were 2.46 +/- 0.43 and 0.72 +/- 0.37 before and after surgery (P < 0.05), respectively. Radiographic follow-up revealed that the height intervertebral space and the lordosis of the cervical spine had been restored and no allograft was found displaced or collapsed and also revealed that all grafts obtained union by 5 months after surgery. CONCLUSIONS: Fibular allograft can replace autologous iliac crest graft in the treatment of cervical spondylosis patients. This method is safe and efficacious and can avoid bone graft-site morbidity. PMID- 12133344 TI - [Treatment of fractures of the talar neck: long term follow up of 89 cases]. AB - OBJECTIVE: To summarize the experience in treating talar neck fractures. METHODS: 89 patients with talar neck fracture were followed up, and classified by modified Hawkin's classification. Three patients of type I were treated with plastic cast and K-wire, 53 of type II with open reduction and internal fixation (ORIF), close reduction and plaster, fusion, and 29 of type III and 4 of type IV with joint fusion and ORIF. RESULTS: Mean follow-up was 5.18 years. Evaluation by Hawkin's criteria, showed 22 patients were excellent, 16 good; 28 fair and 19 poor results respectively. CONCLUSIONS: Emergency operation with ORIF should be performed in type II and type III talar neck fracture. Medial malleolus osteotomy is recommended when medial approach is used. PMID- 12133345 TI - [Spiral CT urography and CT virtual endoscopy in detecting urological diseases]. AB - OBJECTIVE: To evaluate spiral CT urography (SCTU) and CT virtual endoscopy (CTVE) in detecting urologic diseases. METHODS: SCTU was performed in 46 patients with urological diseases including renal neoplasms (2), paropelvic cysts (2), ureteral calculi (6), ureteral stenosis (4), ureteral neoplasms (2), double kidneys and ureter malformation (1), bladder neoplasms (28) and bladder endometreosis (1). The 6 patients with ureteral diseases and 29 patients with bladder diseases underwent CTVE based on spiral CT scan. All CTVE findings were compared with those of B-mode ultrosonography, intravenous urography (IVU), retrograde pyelography (RGP), conventional CT or cystoscopy. RESULTS: All upper urinary tract diseases and bladder diseases (28 cases) were detected by SCTU and CTVE scans and they were confirmed operatively or pathologically except one case of bladder neoplasm (diameter less than 5 mm) was missed. CONCLUSION: SCTU and CTVE have proved to be non-invasive and reliable in the diagnosis of urological diseases and are superior to IVU or conventional CT. CTVE can serve as a supplementary method to fiberoptic cystoscopy or ureteroscopy. PMID- 12133346 TI - [Expression of enhanced green fluorescent protein in transduced variant HT-29c cells in vitro and in vivo]. AB - OBJECTIVE: To evaluate the gene transfer and expression of enhanced green fluorescent protein (EGFP) in retrovirally transducted variant HT-29c cells in vitro and in vivo. METHODS: The retroviral vector prkat EGFP/neo was constructed and was transfected into the 293T cell using a standard calcium phosphate precipitation method. HT-29c cells (in vivo selected HT-29 cells) were transduced by a retroviral vector encoding the EGFP gene. The fluorescence intensity of colorectal carcinoma HT-29c cells after transfected with the EGFP gene bearing retrovirus was visualized using fluorescence microscope and fluorescence activated cell sorter analysis. Multiple biological behaviors of transduced cells such as the proliferating potential and the expression of various antigens were comparatively analyzed between untransfected and transfected in vitro. EGFP expression of the fresh tumor tissue was assessed in vivo. RESULTS: After being transduced, HT-29c cells with the EGFP gene displayed a stable and long-term EGFP expression under the nonselective conditions in vitro. After culturing cells successively to passage 50 in vitro, EGFP expression level was still high. Their biological behaviors, such as expression of tumor antigens, proliferation rate and aggregation capability were not different compared to untransfected parental cells in vitro. In subcutaneous tumors, EGFP was stable and highly expressed. CONCLUSIONS: An EGFP expressing retroviral vector was used to transduce HT-29c cells. The transduced cells show a stable and long-term EGFP expression in vitro and in vivo. These cells are a valuable tool for in vivo analysis of metastatic spread. PMID- 12133347 TI - [Adenoviral transduction of endostatin gene prevents the liver metastasis of colorectal carcinoma in postoperation]. AB - OBJECTIVE: To study the prevention of colorectal cancer liver metastasis by adenoviral transduction of the endostatin gene. METHODS: The recombinant adenovirus expressing endostatin was constructed. Its biological activities were surveyed in vitro, as determined in human umbilicus vein endothelium cell (HUVEC) proliferation inhibition, and in vivo, by reduction of liver metastasis. RESULTS: HUVEC proliferation was obviously inhibited by the infecting supernatant of recombinant adenovirus. Persistent high serum levels of endostatin in peripheral blood, especially in the liver vein were observed. The production of liver metastasis was intervened. CONCLUSIONS: The single injection in the vein of the recombinant adenovirus realizes the high effective and stable expression of endostatin in general body and liver, which brings about the ideal prevention of liver metastasis. PMID- 12133349 TI - [Cadaver renal transplantation and multivariate analysis for graft survival: a clinical review of 2 016 cases]. AB - OBJECTIVE: To review kidney transplantation in the center and analyze the risk factors affecting long-term allograft survival. METHODS: Thirty-two relative variables were analyzed with SAS statistical software. Using Log-rank method, we investigated influence of these variables on short-and long-term survival of grafts. Kaplan-Meier analysis was used to estimate the 1-, 3-, 5-, 10-years graft survival rates and half-life. Proportional hazards regression analysis (Cox model) was used to assess and rank the relative risk of potential variables. RESULTS: The 1-, 3-, 5-, 10-years graft survival rates were 83%, 75%, 66% and 48%. After excluding the patients died with functioning grafts, the 1-, 3-, 5-, 10 years grafts survival rate increased to 89%, 82%, 75% and 69%, respectively. The mean half-life was 8.78 +/- 0.14 and 14.09 +/- 0.20 years, respectively. By Log-rank analysis, factors affecting short- and long-term graft survival were identified as: renal function, duration of graft function became normal, cold ischemia time, presence of acute rejection, delayed graft function, immunosuppressive regimen, complication, infection, anti-rejection therapy. Cox model multivariate analysis showed that there were 18 factors affecting graft survival. CONCLUSIONS: New immunosuppressive agents not only significantly increase short-term graft survival, but also have the better long-term outcome tendency. Making assurance to get high quality donor organ and minimizing the death with graft function may be the most feasible way to prolong graft survival at present. PMID- 12133348 TI - [Construction of adenoviral vector encoding human VEGF(121) cDNA and its expression in vitro]. AB - OBJECTIVE: To construct the adenoviral vector bringing hVEGF(121) cDNA for evaluation of the possibility of VEGF gene therapy in ischemic bone disease. METHODS: Human vascular endothelial growth factor (hVEGF(121)) cDNA obtained from the plasmid pCDI/VEGF(121) was cloned into plasmid pshuttle and further cloned to Adeno-X Viral DNA. The recombinant adenoviral plasmid was identified and then transferred to the adenoviral packaging cell HEK293 by lipofectamine mediated gene transfer method to pack the virus. After titilating the virus, the mouse bone marrow stromal cells (MSC) were transfected by the adenovirus and the expression of VEGF gene was detected. RESULTS: The recombinant Adeno-VEGF(121) was correctly constructed and confirmed by restriction endonuclease analysis and DNA sequencing analysis. After MSCs were tranfected by the virus, RT-PCR showed that hVEGF(121) mRNA was transcripted from the hVEGF(121) gene. Western blot and immune histochemistry showed VEGF(121) protein was expressed in transgene MSCs. CONCLUSION: The recombinant adenoviral vector bringing hVEGF(121) cDNA was successfully constructed and the transgene MSC expressed hVEGF gene in vitro, it provided the further foundation of VEGF gene therapy for bone ischemic diseases. PMID- 12133351 TI - [Ureteral fistula after kidney transplantation]. AB - OBJECTIVE: To study pathogeny, diagnosis and treatment of ureter fistula after renal transplantation. METHODS: The clinical data from 30 cases after renal transplantations were analyzed. RESULTS: Four patients received conservative treatment, and 2 repairment of the fistula. Eleven patients had resection of the ureter or adjustment of the kidney, followed by the anastomosis of the ureter and bladder again. After the turning of the bladder's lamella, 13 patients were given 20 - 24 Foley's tube connecting the pelvis and bladder and nine of them were not subjected to re-anastomosis waiting for the pelvis crawling to the bladder as a tunnel. The one-year survival rates for 30 cases and kidneys was 96.7% (29/30) and 86.7% (26/30) respectively. CONCLUSIONS: There a lot of causes for ureter which are fistula, running related to every aspect of transplantation. Early diagnosis and treatment is important to prognosis. Most patients need reanastomosis. According to the blood stream, edema, length of the ureter, operative procedures are selected to ensure free of strain. PMID- 12133350 TI - [Retrospective evaluation of 2 123 cases of kidney transplantation]. AB - OBJECTIVE: To summarize the experiences in kidney transplantation for 23 years. METHODS: From 1978 to 2001, 2123 kidney transplantations were performed for 2012 patients with end stage renal failure. We analyzed the survival rate of patient/kidney at 1-, 3-, 5 years. The possible factors that could influence the transplantation including general data, donor kidney, surgical technique, immunosuppressants, PRA measurement, HLA-antigen matching, complications were also analyzed retrospectively. RESULTS: In 423 cases (1978 to 1990), hyper-acute rejection occurred in 9 (2.1%) and acute rejection in 198 (46.8%). The 1-, 3-, and 5 years patient/graft survival rates were 86.7%/76.3%, 72.5%/67.9% and 69.5%/59.3% respectively. In the 1700 cases (1991 to 2001), acute graft rejection occurred in 252 (14.8%) but no hyper-acute rejection was observed. The 1-, 3-, and 5 year patient/graft survival rates were 98.6%/96.7%, 93.1%/87.3% and 88.1%/83.6% respectively. CONCLUSIONS: Kidney transplantation is a treatment of choice for patients with end-stage renal failure. Well preoperative preparation is the assurance of a successful transplantation; the high quality of donor's kidney is essential to a successful transplant operation. PRA negative and high grade HLA matching can decrease the ratio of early allograft loss and improve patient/kidney survival rate. Combined medication is also important to prevent rejection and decrease drug toxicity. Low-dosage of CsA with MMF and Pred is the ideal regimen of immunosuppressive therapy. PMID- 12133352 TI - [Diagnosis and treatment of complete necrosis of the ureter after cadaveric renal transplantation]. AB - OBJECTIVE: To deepen the understanding of patients with complete necrosis of the ureter after renal transplantation for early diagnosis and treatment. METHODS: Of 5 patients with complete necrosis of the ureter after renal transplantatioin between January 1991 and April 2001 in our hospital, 4 were male and 1 was female (mean age, 35 years). Seven to 12 days after renal transplantation, native pyeloureterestomy was performed for 1 patient, and the remaining 4 patients received the cutting of the diatal necrosis ureter and vesicoureterostomy because of urine leakage. Six to seven weeks later when the ureter stents were pull out, native pyeloureterestomy or pyeloureteroplasty was performed for the 4 patients because of uropenia and hydronephrosis. RESULTS: Five patients showed normal function of the kidney postopcreation (follow up: 6 - 12 months) without hydronephrosis. CONCLUSIONS: When distal necrosis of the ureter is observed after renal transplantation, complete necrosis of the ureter may occur. Native pyeloureterostomy or pyeloureteroplasty is an effective treatment. PMID- 12133353 TI - [Influence of cytokine gene polymorphism on renal transplantation]. AB - OBJECTIVE: To study the influence of cytokine gene polymorphism on the incidence of acute rejection in renal transplantation. METHODS: The cytokine genotypes of TNF-alpha, TGF-beta(1), IL-10 and IL-6 in 24 normal people and 115 consecutive renal transplant recipients were determined by PCR-SSP using sequence-specific oligonucleotide primer. The cytokine genotypes were then correlated with the incidence of acute rejection in renal transplantation. RESULTS: The frequency of TNF-alpha and IL-10 low producer genotype in the 115 kidney recipients and control group was 88.7%, 80% and 87.5%, 75% respectively, and the frequency of TGF-beta(1) and IL-6 high producer genotype was 79.1%, 100% and 66.7%, 100% respectively. The incidence of acute rejection was higher in the recipients with TNF-(high producer or IL-10 intermediate high producer genotype than in the recipients with low producer genotype [53.8%, 43.5% vs 18.6% (chi(2) = 8.17), 17.4% (chi(2) = 7.16), P < 0.01, respectively]. CONCLUSION: This study demonstrated that TNF-alpha and IL-10 gene polymorphism has significant influence on the incidence of acute rejection in renal transplantation. PMID- 12133354 TI - [Experimental study of sertoli cell and CTLA-4Ig's induction of immune tolerance of allogeneic renal cell]. AB - OBJECTIVE: To study the protective role of Sertoli cell expressing FasL and CTLA 4Ig on allogeneic renal cell. METHODS: Testicular Sertoli cell and renal cell were prepared by digestion with enzymes. About 106 cells were injected into the subrenal capsule of allogeneic rats. 36 rats were divided into 3 groups: control group (10), co-transplantation group (16) and co-transplantation collaborated with CTLA-4Ig group (CTLA group, 10). Levels of serum IL-2 were tested on the 1st, 7th, 14th, 20th day after transplantation. The kidney was extracted on the 20th day. The survival of renal cells was determined by the Avidin Biotin Peroxidase Complex Technique (ABC); the brightness of grafts was measured by MD 20 image analysis system. Apoptosis within the grafts was observed with Terminal Deoxynucleotidyl Transferase-mediated X-dUTP Nick end Labeling (TUNEL) method. RESULTS: The survival of renal cells in control group, co-transplantation group and CTLA group was 0, 14 and 10 respectively; The brightness values of co transplantation group and CTLA group were 0.362 +/- 0.017 and 0.445 +/- 0.021, and the statistic differences between them were significant. Levels of serum IL-2 in CTLA group were lower compared with co-transplantation group, there being significant differences as well; Apoptosis of lymphocyte was observed within the grafts. CONCLUSIONS: Sertoli cell and CTLA-4Ig have coordinative protective effect on allogeneic renal cell. PMID- 12133355 TI - [The protective effect and mechanism of ischemic preconditioning for hepatic resection under hepatic blood inflow occlusion in hepatocellular carcinoma patients with cirrhosis]. AB - OBJECTIVE: To investigate the protective effect of ischemic preconditioning (IPC) for hepatic resection under hepatic blood inflow occlusion (HBIO) in hepatocellular carcinoma patients with cirrhosis and its possible mechanism. METHODS: 29 consecutive patients resectable HCC were randomized into two groups. IPC group: before HBIO, IPC with 5 min of ischemia and 5 min of reperfusion was given; control group: simple HBIO. The liver function, hepatic caspase-3 activity, and apoptotic cell were compared between the two groups. RESULTS: The AST, ALT levels of POD 1, POD 3 and POD 7 in the IPC group were significantly higher than those of the control group, respectively (t = 4.238, P < 0.05). The TBIL levels of POD 3 and POD 7 in the IPC group were significantly higher than those of the control group, respectively (t = 2.296, P < 0.05). The ALB of POD 1 in the IPC group was higher than in the control group (t = 2.029, P > 0.05). After 1 h of reperfusion, the hepatic caspase-3 activity and apoptotic sinusoidal endothelial cell were significantly higher than those of in the control group (t = 2.349, P < 0.05). CONCLUSIONS: IPC has the a protective effect in hepatic resection under HBIO in HCC patients with cirrhosis. Its mechanism is that sinusoidal endothelial cell apoptosis is inhibited by inhibiting caspase-3 activity. PMID- 12133356 TI - [Resection of right or total hepatic caudate lobe including paracaval portion]. AB - OBJECTIVES: To evaluate the surgical techniques and feasibility for resecting the hepatic caudate lobe including the paracaval portion. METHODS: Right posterior approach for right caudate lobectomy and left lateral approach for total caudate lobectomy were taken with or without some kinds of preparatory segmentectomies. RESULTS: Seven right and 6 total caudate lobectomies, all including paracaval portion, ware accomplished without operative death. The mean intraoperative blood loss was 896.15 (250 - 2 000) ml and the mean portal triad clamping time was 25.4 (10 - 83) min. The postoperative course was uneventful for all the cases, and the mean hospital stay was 12 (9 - 22) days. CONCLUSIONS: Although being complicated anatomically, resection of the hepatic caudate lobe including the paracaval portion is feasible with a high safety. PMID- 12133357 TI - [The extensibility and retractility of surgical margins in digestive tract cancer]. AB - OBJECTIVE: To study the extensibility and retractility of the surgical margins in digestive system neoplasms. METHODS: The length difference of the digestive tract was measured in vivo and in vitro under different conditions. Five cm of the stomach, small intestine and large bowel and 2 cm of the esophagus were measured as standard control length in vivo just before resection. The length was measured with a ruler under pull of 500 g and 1 000 g in vivo, in fresh status in vitro, and 10% formaldehyde fixed for 6 - 8 h, 12 - 24 h and 48 - 72 h. The extension or retraction ratio was calculated. The length difference was divided by the natural length in vivo. RESULTS: Seventeen cases of the esophagus, 18 cases of the stomach, 15 cases of the small intestine and 25 cases of the large bowel were measured under pull of 500 g and 1 000 g. The esophagus extended 16.5% and 30.5%, stomach 15.0% and 22.6%, small intestine 66.4% and 120.0%, large bowel 36.0% and 56.0% respectively. In natural status ex vivo, the esophagus retracted 44.5%, stomach 13.6%, small intestine 11.4% and large bowel 15.6% respectively; they continue to retract after 10% formaldehyde fixation until 12 - 24 h later. If the length of surgical margin of the fresh specimen ex vivo was x, the natural length of margin in vivo of the esophagus would be 1.80 x, stomach 1.16 x, small intestine 1.13 x, and large bowel 1.18 x. If formaldehyde fixation for 6 - 8 h, the natural length of surgical margin in vivo of the esophagus would be 1.82 x, stomach 1.41 x, small intestine 1.22 x, large bowel 1.55 x. If formaldehyde fixation for 12 - 24 h, the surgical margin length in vivo of the esophagus would be 2.22 x, stomach 1.43 x, small intestine 1.28 x, and large bowel 1.57 x. CONCLUSION: The length of surgical margin of digestive system cancers varied under different conditions, and the evaluation of surgical margin during surgery should be performed under natural status in vivo. PMID- 12133358 TI - [Three-pronged approach to treat rectal cancer: liposome transfer cytosine deaminase gene, ionizing radiation and dendritic cells]. AB - OBJECTIVE: To investigate the anticancer activity of three-pronged approach: liposome transfer cytosine deaminase (CD) gene, ionizing radiation, and dendritic cells (DCs). METHODS: After rectal cancer cells were killed by CD and 5-FC and radiotherapy with improved liposome transfection and radiosensitization, the killed cancer cells, DCs and T lymphocytes were incubated together. The uptake of (3)H-TdR weighed the immune activity of DCs. The effect of anticancer was monitored by MTT. RESULTS: DCs acquired the apoptosis body and other antigen from the killed cancer cells and presented them to T-lymphocytes. The induced CTL killed the cancer cells powerfully and selectively. The mortality of rectal cancer cells was 98.1%, 76.2%, 37.8% at the ratio of efficacy to target of 30:1, 15:1, 1:1, respectively. CONCLUSION: The three-pronged approach may represent a powerful and safe means to selectively destroy cancer cells. PMID- 12133359 TI - [Expression and clinical significance of c-kit oncogene in gastrointestinal stromal tumors]. AB - OBJECTIVE: To investigate the clinicopathological and prognostic significance of the c-kit protein in gastrointestinal stromal tumor (GIST). METHODS: Paraffin embedded materials from 53 benign GISTs, 13 potentially malignant and 55 malignant cases were analysed for c-kit expression by immunohistochemical method, while using leiomyomas and schwannomas as controls. Positive signals were shown in cytoplasma and cell membrane. RESULTS: Of 122 GISTs, 118 (97%) were positive for c-kit. Localization of positive signals was accurate. The rate of c-kit protein in benign, potentially malignant and malignant cases was 98% (53/54), 93% (12/13), 96% (53/55) respectively. Compared with benign GIST, the positivity of c kit in metastasis or recurrent cases decreased, but c-kit protein expression rate was not significantly different between the three patterns of GIST (chi(2) = 1.167, P > 0.05). Leiomyomas and schwannomas were typically c-kit negative. CONCLUSION: As a sensitive and specific marker of GIST, c-kit seems to be a useful antibody in the diagnosis and differential diagnosis of GIST, but it may not be used as a prognostic index. PMID- 12133360 TI - [Treatment of complex bone nonunion with external skeletal fixation]. AB - OBJECTIVES: To summarize the experience in the treatment of 112 cases of complex bone nonunion from 1982 to 1999 in our department and introduce the technique of external skeletal fixation. METHODS: The two fragment ends of all cases were fixed under pressure with half-ring sulcated external skeletal fixator. Those cases complicated by bone defect or limb shortening were operated on with epiphysiotomy to restore the length of the limb in the period of compressive fixation or after the occurrence of bone union according to the condition of complicated infection and the length of the limb shortened. RESULTS: The nonunion of the 112 cases was united eventually. The infection in 34 cases was eradicated. Bone union in cases without infection took 3 approximately 7 months (average 5.2 months) and in cases with infection took 5 approximately 11 months (average 5.5 months). The length of the limb in 11 cases with bone defect was restored in the same period of compressive external fixation and another 8 cases achieved after bone union. The length between the injured and healthy limbs was balanced. CONCLUSIONS: When external skeletal fixation is employed to treat those troublesome cases of bone nonunion, the pins for fixation are inserted in sites far from the lesions and the non-united fragment ends are exposed only in the area without scars. Consequently, there is little interference with the blood circulation and the osteogenic potency of the fragment ends. The sclerotic bone tissue is not excised, the marrow cavity is not chased to be open and the fragment ends are only moderately modified. As a result, the stability of fixation is increased and further shortening of the limb avoided. External skeletal fixation using small pins with cross penetration results in plastic fixation and promotes bone healing. Bone lengthening with epiphysiotomy can restore the balance of the limbs. PMID- 12133361 TI - [Operative complications in tethered cord syndrome and their management]. AB - OBJECTIVES: To find out the common complications induced by the operation on patients with tethered cord syndrome (TCS) and to discuss the mechanism and the treatment of these complications. METHODS: During 1993 and 2001, the spinal cords of 87 patients with TCS were explored and released operatively. The operative complications were analyzed retrospectively and the results of prevention and treatment of these complications were evaluated. RESULTS: The operative complications in patients with TCS included headache (43.7%), hemorrhage (31.0%), lumbago (21.8%), retethering (18.4%), lumbar instability (12.7%), neural injury (8.1%) and CSF leakage (4.6%). CONCLUSIONS: To reduce operative complications, the pathology of TCS should be investigated thoroughly before operation. The techniques of microsurgery and electrophysiology should be used properly and some complications were reversible after treatment. Post-operation rehabilitation training should be stressed. PMID- 12133362 TI - [Diagnosis and treatment of spinal cord cavernous angioma]. AB - OBJECTIVE: To investigate the diagnosis and treatment of patients with spinal cord cavernous angioma. METHODS: Clinical data on 23 patients with intramedually cavernous angiomas were analyzed. The function of the spinal cord was evaluated before and after treatment using the Aminoff & Logue scale. RESULTS: All patients underwent resection of spinal cord cavernous angioma except two patients who refused surgical treatment. Complete removal was achieved in 18 patients, and subtotal removal in 3. Twenty-one patients with spinal cavernous angioma were confirmed pathologically. The postoperative neurological status was improved in 15 patients, remained unchanged in 4, and aggravated in 2. No deaths occurred. CONCLUSIONS: MRI is specific in diagnosis of intramedually cavernous angioma. The outcome of surgical treatment is better. Surgical treatment is suitable for patients with symptoms. PMID- 12133363 TI - [Levels of plasma endothelin, nitric oxide and atrial natriuretic peptide after cardiopulmonary bypass and influence of inhaled nitric oxide in patients with ventricular septal defect and pulmonary hypertension]. AB - OBJECTIVES: To assess the changes of the levels of plasma endothelin (ET-1), nitric oxide (NO) and atrial natriuretic peptide (ANP) after cardiopulmonary bypass (CPB) and the influence of inhaled nitric oxide in patients with ventricular septal defect (VSD) and pulmonary hypertension (PH). METHODS: Sixty patients with VSD were enrolled in this study. They were divided into 2 groups: group A [no-PH group, mean pulmonary artery pressure (mPAP) < 20 mm Hg (1 mm Hg = 0.133 kPa) n = 20] and group B (PH group, mPAP > 20 mm Hg, n = 40). Group B was subdivided into two groups by randomized block, group B(1) (inhaled NO group, n = 20) and group B(2) (contrast group, n = 20). The plasma ET-1, NO, ANP concentrations were assayed at 24 h pre-operation and 0 h, 1 h, 5 h, 12 h, 24 h, 48 h after CPB. RESULTS: The preoperative plasma ET-1, NO and ANP concentrations in group B were significantly higher than those in group A. In three groups, the plasma ET-1 concentration at 0 h after CPB was significantly higher than that at 24 h pre-operation, and the plasma NO concentration at 0 h after CPB was significantly lower than that at 24 h pre-operation. In group B, the plasma ANP concentration at 0 h after CPB was significantly higher than that at 24 h pre operation. After CPB, the plasma ET-1 concentration in group B(1) decreased faster than that in group B(2), and the plasma NO concentration in group B(1) increased faster than that in group B(2). In group B, the preoperative plasma ET 1 concentration negatively correlated with the preoperative plasma NO concentration and positively correlated with the preoperative ANP concentration. CONCLUSIONS: The broken dynamic balance of ET-1/NO may take part in generation and development of pulmonary hypertension. ANP acts as a favorable physiological regulating factor in the pathogenesis of pulmonary hypertension. CPB can regulate the level of ET-1 up and NO and ANP down while inhaled NO can cause the level of ET-1 down and the level of NO up. PMID- 12133364 TI - [Diagnosis, treatment and prognosis of thymoma: analysis of 116 cases]. AB - OBJECTIVES: To study the diagnosis and treatment of thymoma and to assess prognosis factors. METHODS: The clinical data on 116 patients with thymoma were collected. A retrospective analysis was performed by comparison of their survival rates computed by the actuarial method and rate of recurrence and metastasis. RESULTS: Chest radiograph was used chiefly for the preoperative diagnosis of thymoma; myasthenia gravis (MG) (25.0%, 29/116) was the most common paraneoplastic disease. An extensive and radical resection was carried out to reduce the recurrence rate of thymoma with stage I and stage II (chi(2) = 4.941 P = 0.0219). The survival time was prolonged by postoperative radiotherapy and chemotherapy. A strong correlation was noted between the clinical stage and histologic subtype of M-H classification, by which the invasive behavior of thymoma was predicted (r = 0.385, P = 0.007). The 3-, 5-, and 10-year survival rates were 81.2%, 67.9% and 40.5%, respectively. Statistical analysis showed a significant negative correlation between stage and survival rate (r = -0.897, P = 0.0000). CONCLUSION: The prognosis of thymoma depends mainly on the histologic subtype, clinical stage and multimodality treatment rather than paraneoplastic diseases. PMID- 12133365 TI - [The study of pro-nucleating activity on 33.5 x 10(3) vesicular protein in bile]. AB - OBJECTIVE: To evaluate the pro-nucleating activity in 33.5 x 10(3) vesicular protein. METHODS: The model biles were established according Kibe and Zhu. The pro-nucleating activity of 33.5 x 10(3) vesicular protein were examined by polarized light microscopy. The protein and its enzymatic deglycosylation and proteolysis fractions nucleation promoting activity were detected by cholesterol crystal growth assay. RESULTS: 33.5 x 10(3) vesicular protein displayed apparent potency of pro-nucleation with activity of 0.310, and derived crystal growth curve indices It, Ig, Ic were presented as 0.57, 1.52, 1.63 respectively, but after treated by N-glycanase enzyme and pronase, no promoting activity were found. CONCLUSION: The 33.5 x 10(3) vesicular protein may be involved in the nucleation process of gallstone formation, which is regulated by its peptide and sugar chain. PMID- 12133366 TI - [In vitro and in vivo bystander effect of adenovirus-mediated transfer of the herpes simplex virus thymidine kinase gene]. AB - OBJECTIVE: To investigate in vitro and in vivo bystander effect, including distance bystander effect of adenovirus-mediated transfer of the herpes simplex virus thymidine kinase gene (HSV-tk). METHODS: In vitro, mixed tk + BEL-7402 cells and tk-BEL-7402 cells in diverse proportions and ganciclovir (GCV) was given, then tested the survival ratio of cells by MTT. In vivo, 5 x 10(6) and 5 x 10(7) tk + BEL-7402 cells were injected into the tumors in nude mice following GCV. The change of the size of the tumors was observed. To observe distance bystander effect, adenovirues with HSV-tk (1 x 10(9) PFU) were injected into the tumor, which was the one of the bilateral tumors in nude mice. Subsequently, GCV was given and the change of the tumor was observed. RESULTS: Significant bystander effect was observed in vitro and in vivo. In vitro when tk + cells: tk( ) cells was 1:9, the survival rate of mixed cells was 36.6%. When the proportion of tk(+) cells was 90%, the survival rate of mixed cells was 3.2%. In vivo, those tumors with injection of tk(+) BEL-7402 cells were suppressed (P < 0.05). But in group of distance bystander effect the tumors on another side were not suppressed. CONCLUSION: In vitro, bystander effect exists. In nude mice, if tk(+) cells and tk(-) cells are contiguous, bystander effect is significant, or probably no bystander effect. PMID- 12133367 TI - [The effect of anterior cruciate ligament rupture and reconstruction on the degeneration of articular cartilage in rabbit knee]. AB - OBJECTIVE: To investigate the effect of rupture and reconstruction of the anterior cruciate ligament (ACL) on the degeneration of rabbit knee articular cartilage. METHODS: 14 mature New Zealand white rabbits were divided into four groups. In group I, the ACL of the right knees in 7 rabbits was resected and immediately reconstructed, and the contralateral ACL was resected only in controll f group I. In group II, the ACL of the right knees in 7 rabbits was reconstructed 3 weeks after the ACL was resected and the contralateral joints in control group II, in which only a medial arthrotomy was performed. The rabbits were killed 8 weeks after the operation. The methods of ink straining, histology and SEM were used to analyze the changes in articular cartilage of the joints. RESULTS: The results of ink method and HE straining were analyzed quantitatively. The degeneration of knee articular cartilage in group I was significantly weaker than that in control group I (Hc = 5.9889, P = 0.0144). The degeneration of knee articular cartilage in group II was as serious as that in control group I (Hc = 0.7143, P = 0.785). CONCLUSIONS: Immediate reconstruction of the ACL can effectively prevent articular cartilage from degeneration. Once the articular cartilage damaged moderately, delayed reconstruction of the ACL could not effectively reduce the development of degeneration. So once the ACL is ruptured, reconstruction should be performed in the early stage to restore the stability of knee joint to prevent the articular cartilage from degeneration. PMID- 12133368 TI - [Clinical use of capsular tension ring during phacoemulsification in subluxated lens]. AB - OBJECTIVE: To investigate the safety of capsular tension ring implantation during phacoemul- sification in the subluxated lens. METHODS: Capsular tension ring was implanted in 13 cases (14 eyes) of subluxated lens after continuous curvilinear capsulorrhexis during phacoemulsification. The luxated area was ranged from one third to three quarters of a circle. The capsular tension ring was necessary to be fixated on sclera if the luxated area was larger than half-quarter of a circle. After the ring was implanted, phacoemulsification was performed and foldable intraocular lens (IOL) was implanted. RESULTS: The capsular tension ring was successfully implanted in 14 eyes, including 11 IOLs fixated in the bag and 3 in ciliary sulcus. IOL positions in 12 cases were normal, and in 2 eyes, had slight tilt. The most common intraoperative complication was the rupture of capsular bag, and secondly, the loss of vitreous body. CONCLUSION: Capsular tension ring implantation is safe in the patient of lens subluxation during phacoemulsification. It can prevent the IOL decentration and decrease the vitreoretinal complication. PMID- 12133369 TI - [Macular image changes of optical coherence tomography after phacoemulsification]. AB - OBJECTIVE: To investigate the effects of phacoemulsification on the macula following uncomplicated phacoemulsification by optical coherence tomography (OCT). METHODS: Eighty eyes of the senile cataract were chosen randomly. The uncomplicated phacoemulsification was performed. OCT was examined preoperatively and 1 week after the surgery. Preoperative visual acuity, the retinal thickness and phaco power were compared with those after surgery. RESULTS: In 80 eyes, the preoperative mean foveal thickness was (142.9 +/- 16.7) micrometer and the postoperative (157.9 +/- 36.7) micrometer, the difference being not significant (P > 0.05). Three eyes had macular edema 1 week after surgery. In 11 eyes with Tyndall sign (+ +), the mean postoperative foveal thickness was thicker than the mean preoperative value (P < 0.05). In lower phaco power group, the mean postoperative foveal thickness was (156.2 +/- 18.3) micrometer and the higher phaco power group was (172.6 +/- 32.9) microm (P < 0.05). The best corrected visual acuity after surgery had a negative correlation with the retinal thickness. CONCLUSIONS: The retinal thickening and macular edema can be found after uncomplicated phacoemulsification. The higher phaco power results in significant inflammation and thicker retina. The visual consequences were proportional to the degrees of macular thickening. PMID- 12133370 TI - [Intraocular lens implantation and coreoplasty for congenital cataract after optical iridectomy]. AB - OBJECTIVE: To explore the therapeutic method for congenital perinuclear cataract after optical iridectomy. METHODS: Twelve cases (24 eyes) with congenital perinuclear cataract after optical iridectomy were selected. After extracapsular cataract extraction (ECCE) and intraocular lens (IOL) implantation, the sides of the iridectomy were sutured to complete the coreoplasty. RESULT: No patient complained of photophobia and monocular diplopia postoperatively. Every one got a nearly round pupil with light reaction and a corrected visual acuity >/= 0.5. CONCLUSION: The combination of ECCE, IOL implantation and coreoplasty is a good therapeutic method for congenital perinuclear cataract after optical iridectomy. PMID- 12133371 TI - [Implantation of stage II anterior chamber intraocular lens in aphakic eye after vitrectomy]. AB - OBJECTIVE: To sum up the clinical methods and results of stage II anterior chamber intraocular lens (AC-IOL) implantation in 54 aphakic eyes after vitrectomy. METHODS: Traditional corneoscleral limbal and tunnel incision were respectively used to perform stage II AC-IOL implantation separately in 54 aphakic eyes following vitrectomy. RESULT: All 54 eyes had obtained good visual acuity and reached the corrected visual acuity before operation. The naked visual acuity of 11 eyes was >/= 1.0, and the corrected visual acuity of 16 eyes, >/= 1.0. CONCLUSIONS: Two methods of implantation of AC-IOL in aphakic eye after vitrectomy all obtain good results. Tunnel incision can make the surgery process easier and induce corneal astigmatism smaller. The bracket function of vitreous body after vitrectomy disappears. This technique can maintain IOP very well during the operation and make the operation safer. PMID- 12133373 TI - [Effects of aging and post-translational modifications on molecular chaperone like activity of alpha-crystalline]. AB - OBJECTIVE: To study the affects of aging and post-translational modifications in vitro on rabbit alpha-crystalline molecular chaperone-like activity. METHODS: (1) Rabbits with three different ages (3 days old, 3 months old and 2 years old) were chosen in the study. The water-soluble crystalline in the three age groups was purified by Sephadex G-200 gel filtration. Different abilities of them to protect beta-low crystalline from thermal aggregation were observed. (2) Incubation of alpha-crystalline with fructose or with H(2)O(2) was carried out for 24 hours. Fructose was used for glycosylation and H(2)O(2) for oxidation of alpha crystalline. Different abilities of modified and unmodified alpha-crystalline to protect beta-low crystalline from aggregation were observed. RESULTS: (1) alpha Crystalline of 3 months old rabbit has more ability to protect beta-low crystalline from aggregation than 2 years old rabbit but less ability than 3 days old rabbit. (2) The glycosylated and oxidized alpha-crystalline have less ability to protect beta-low crystalline from aggregation than the unmodified alpha crystalline. CONCLUSIONS: (1) With aging rabbit alpha-crystallines gradually lose their molecular chaperone-like ability. (2) The modified alpha-crystallines show the loss in molecular chaperone-like ability the most. PMID- 12133372 TI - [Tear film changes after phacoemulsification]. AB - OBJECTIVE: To evaluate tear film changes after phacoemulsification. METHODS: This prospective randomized study involved 68 consecutive patients with age-related cataract (79 eyes) who underwent phacoemulsification. Tear break-up time, Schirmer I test (S I t) value, corneal fluorescein staining, tear film pattern by DR-1 tear interfermetry and the height of tear meniscus were measured preoperatively and 1 day and 2, 7, 14, 30, 180 days postoperatively. RESULTS: At 1 day and 2 days postoperatively, the mean tear break-up time reduced greatly, and the number of patients who showed III to V tear film pattern, the mean S I t value, the height of tear meniscus and fluorescein staining scores increased significantly (P < 0.001 for day 1, P < 0.005 for day 2). S I t value at 7 days postoperatively (P = 0.831) and the height of tear meniscus at 14 days returned to their preoperative values (P = 1.000). The tear break-up time, corneal fluorescein staining scores and the grades of tear film pattern all recovered at 30 days postoperatively (P > 0.05). At 30 days postoperatively, 19.3% of patients showed shorter tear break-up time and less S I t value than that of preoperative day. In addition, 1/9 of the patients with normal preoperative tear film experienced dry eye at 30 days postoperatively. The cases with tear break-up time < 10 seconds compared to patients with tear break-up time >/= 10 seconds were more easily to show tear instability postoperatively (relative risk 13.5, 8.25, 20.0, 39.0, 8.6, 3.9 at 1 day and 2, 7, 14, 30, 180 postoperative days, respectively). CONCLUSION: Phacoemulsification significantly alters the tear break-up time, S I t value, corneal fluorescein staining, tear film pattern and the height of tear meniscus. Some patients with normal tear film may experience dry eye after surgery. Patients with preexisting tear break-up time < 10 seconds are easily at risk of experiencing the tear film instability postoperatively. PMID- 12133374 TI - [The expression of calpain II in rat lens epithelial cells of hydrogen peroxide induced cataract]. AB - OBJECTIVE: To study the expression and role of calpain II in rat lens epithelial cells (LECs) of hydrogen peroxide (H(2)O(2))-induced cataract. METHODS: Rat lenses were cultured in vitro and cataract was induced by 2 mM H(2)O(2). The lenses were observed under microscope. Simultaneously, photographs and picture analyses were done in order to detect the variation of the opacity. The expression of calpain II in rat LECs was detected with immunohistochemical method and compared with a control group. RESULTS: When the lenses were cultured in 2 mM H(2)O(2) for 3 hours (h), the expression of calpain II in rat LECs was increased obviously. There was significant difference between H(2)O(2)-induced and the control group (P = 0.006). After 6 h, vesicles appeared at the equator of the lenses. There was a significant difference between the result of picture analysis of H(2)O(2)-induced and control group (P = 0.013). So the expression of calpain II in rat LECs of H(2)O(2)-induced cataract was increased before cataract occurred. After induced with H(2)O(2) for 24 h, the opacity of lenses and the expression of LECs were both increased as compared with that at 6 h (P = 0.000, 0.000). CONCLUSION: H(2)O(2) can enhance the expression of calpain II in rat LECs which may play a role in the mechanism of oxidative stress-induced cataract. PMID- 12133375 TI - [Local resection of ciliary body tumors with vitreoretinal surgery]. AB - OBJECTIVE: To investigate the effects, safety and indication of local resection of ciliary body tumors with vitreoretinal surgery. METHODS: Twelve patients with ciliary body tumors, including 5 males and 7 females with the age range 13 - 48 years (average 32 years) were treated with local resection. Ultrasound biomicroscopy (UBM) and ultrasonography confirmed the tumors, and the surgery was performed under hypotensive general anesthesia or local anesthesia. The range of the diameters of tumors was 5 - 20 mm and of the thickness was 4 - 12 mm. Retinal detachment was detected in 2 eyes. Local lamellar sclerocyclectomy or sclerocyclochoroidectomy with vitreoretinal surgery was performed for the resection. The surgery included lensectomy and vitrectomy using perfluorocarbon liquid, endolaser and gases or silicone oil tamponade and also scleral allograft if necessary. The resected tumors were diagnosed by pathologic examinations. RESULTS: There were no tumor recurrence and metastasis during follow-up 7 - 38 months (average 14.2 months). In the last follow-up date, the visual acuities were less than 0.05 in 1 case, 0.05 - 0.2 in 3 cases and 0.3 - 1.0 in 8 cases. The pathologic diagnoses included malignant melanoma in 5 cases, melanocytoma in 2 cases, adenoma of non-pigmented ciliary epithelium in 3 cases, neurofibroma in 1 case and glioma in 1 case. CONCLUSION: Local resection of ciliary tumor not only can conserve the eyeball, but also part of vision, and the accurate diagnosis can also be made from the resected tissues. With vitreoretinal surgery, it can avoid complications and improve the success rate. It is a safe and effective method for benign or malignant tumors of the ciliary body. PMID- 12133376 TI - [Lacrimal duct stent for nasolacrimal duct obstruction]. AB - OBJECTIVE: To introduce a new interventional procedure using lacrimal duct stent for the treatment of nasolacrimal duct obstruction, evaluate the short-term and long-term efficacy and observed its intra- and post-operative complications and management. METHODS: 102 cases (136 eyes) of nasolacrimal duct obstruction underwent the lacrimal duct stent placement, the nasolacrimal duct was dilated and the stent was placed retrograde. Digital subtraction dacryocystography was performed before and after the stent placement. RESULTS: Stent placement was technically successful in 132 eyes, and the technical success rate was 97.1%. Good flow of contrast medium was obtained in all patients after stenting. Three months postoperatively, the success rate was 99.2%. After twelve months of follow up, the success rate was 91.4%. The intra-and post-operative complications included stenting failure, transient hemorrhage and obstruction of the stent. CONCLUSION: Interventional procedure using lacrimal duct stent is a safe, simple and effective method for nasolacrimal duct obstruction without disturbing the normal anatomy. PMID- 12133377 TI - [Distribution and shifting trends of the pathogens for bacterial keratitis]. AB - OBJECTIVE: To study the distribution and shifting trends of corneal bacterial isolates from bacterial keratitis retrospectively. METHODS: The data of bacterial cultures of 2 220 corneal isolates from January, 1989 to December, 1998 were reviewed. RESULTS: During this 10-year period, 2 220 consecutive corneal cultures were obtained, and a positive-culture was recovered in 490 isolates, the positive rate being 22.1%. In the 490 positive-cultures, Gram-positive cocci and Gram negative bacilli represented 51.0% and 39.4% respectively. Gram-positive bacilli was 9.2%. Pseudomonas aeruginosa was isolated the most commonly in the corneal isolates (32.2%), followed by coagulase-negative staphylococcus (18.6%). Streptococcus pneumoniae represented 12.0%. During this period, we documented a gradual increase in the number of Gram-positive cocci coupled with a decrease of the number of Gram-negative bacilli. CONCLUSION: Pseudomonas aeruginosa and coagulase-negative staphylococcus are the most common pathogens in bacterial keratitis in the north part of China. At present, the increased recovery of Gram positive cocci and decreased of Gram-negative bacilli from keratitis isolates present an important challenge to the ophthalmology. PMID- 12133378 TI - [Observation on healing process of corneal lens after epikeratophakia with confocal microscopy]. AB - OBJECTIVE: To investigate the nerve and cell healing process of cryopreservation tissue lens after epikeratophakia by in vivo confocal microscopy. METHOD: Epithelia, stroma and nerves of tissue lens, interface between lens and recipient and stroma and endothelia of recipient were observed by in vivo confocal microscopy and recorded by video in 24 cases of keratoconus from 3 days to 5 years after epikeratophakia. RESULTS: Epithelial layer of lens: The tissue lens was covered by corneal superficial flat cells with wing cells and basal cells on the periphery at 3 - 4 days postoperatively, but the morphology and arrangement of these cells were irregular with low density. Superficial flat cells took shape completely at one month, and the morphology and density of the basal cells tended to be normal at 6 months postoperatively. Subepithelial nerve plexus were scattered at 18 months and their normalization completed at 2 years postoperatively. Stroma of lens: Keratocytes appeared circular, dot-shaped, rod shaped or reticular in morphology postoperatively. Normal keratocytes were discovered sporadically on the periphery of the lens at 2 years postoperatively. At 5 years postoperatively the density of keratocytes was lower than normal stromal keratocyte in the center of the lens. Stromal nerves of lens: Nerves could grow into tissue lens at 6 months postoperatively, the quantity of nerves increased at 2 years and was still less than normal quantity at 5 years postoperatively. Stroma and endothelia of recipient: There was no change in the stroma and endothelia of recipient. CONCLUSIONS: There is significant difference among healing process of epithelia, nerves of unvital tissue lens after epikeratophakia, but there is little change in the transparency of tissue lens. PMID- 12133379 TI - [Expression of tissue inhibitors of metalloproteinase in monkey trabecular meshwork cells and ciliary muscle cells]. AB - OBJECTIVE: To identify tissue inhibitors of metalloproteinase (TIMP) released by normal monkey trabecular meshwork cells and ciliary muscle cells in vitro and to detect the roles of matrix metalloproteinase (MMP) and TIMP playing in trabecular outflow and uveoscleral outflow in normal eyes. METHODS: The expressions of TIMP and MMP in monkey trabecular meshwork and ciliary muscle cultures were detected by the technology of reverse zymography. RESULTS: (1) TIMP-1, TIMP-2 and TIMP-3 were found in both trabecular meshwork and ciliary muscle cultured cells. (2) The ratio of MMP/TIMP in trabecular meshwork cells was much higher than that in ciliary muscle cells. CONCLUSIONS: The different ratio of MMP/TIMP in these two types of cells suggests that the capability of degrading extracellular matrix in trabecular meshwork cells be much higher than that in ciliary muscle cells, that may be one of the reasons why conventional trabecular outflow is stronger than uveoscleral outflow. PMID- 12133380 TI - [Apoptosis of bovine trabecular meshwork cells induced by dexamethasone]. AB - OBJECTIVE: To investigate apoptosis of bovine trabecular meshwork cells induced by dexamethasone. METHODS: In the incubation method of system in vitro, dexamethasone (1 - 500 mg/L) was added into the syncretic culture solution of the third to fifth generation of bovine trabecular meshwork cells. After 1 - 14 days, the culture solution was observed by phase-contrast microscopy, fluorescence microscopy and transmission electron microscopy, and was studied by DNA laddering and flow cytometric analysis. RESULTS: Dexamethasone (125 - 500 mg/L) induced apoptosis of trabecular meshwork cells in a dose-time-dependent manner. CONCLUSIONS: Apoptosis of cultured bovine trabecular meshwork cells can be induced by dexamethasone, which may be one of the pathogenic mechanisms of steroid-induced glaucoma. PMID- 12133381 TI - [Expression of insulin-like growth factor-I in bovine trabecular meshwork cells in vitro]. AB - OBJECTIVE: To determine whether cultured bovine trabecular meshwork cells in vitro would express insulin-like growth factor-I (IGF-I). METHODS: Newborn bovine trabecular meshwork cells of the third passage were cultured in vitro. The whole RNA from 10(6) cells was extracted with Trizol reagent, and specific oligonucleotide primer pair and reverse transcription polymerase chain reaction (RT-PCR) were used for detection of IGF-I messenger RNA. Immunohistochemical stain was used to detect IGF-I protein. RESULTS: Cultured cells had the specific characteristics of bovine trabecular meshwork cells. A single RT-PCR amplified product whose sequence was homologous to the known sequence was obtained. IGF-I immunostain was positive. CONCLUSIONS: Trabecular meshwork cells may produce IGF I. This finding strongly supports the possibility that IGF-I may affect trabecular meshwork cells through paracrine and autocrine mechanisms. PMID- 12133382 TI - [Alteration of lateral geniculate body cells after partial injury]. AB - OBJECTIVE: To investigate the damage of the cells of the lateral geniculate bodies (LGB) of Sprague-Dawley (SD) rats after partial optic nerve injury. METHODS: The optic nerve was pressed by a forceps (40 g) for 30 seconds to set up the animal model with optic nerve injury. LGB was chopped in serial coronal slices (10 microm thickness) by a cryomicrotome. The apoptosis of LGB cells was detected with terminal deoxynucleotidyl transferase (TDT)-mediated dUTP nick end labeling (TUNEL) technique after one week. After retrograde labeling with Granular Blue (GB) through injecting GB into optic center, the average density of the LGB cells was counted under the fluorescent microscope. Using monoclonal rabbit anti-rat neurofilament (NF) antibody, the neurons of LGB were detected by immunochemistry stains, and the standard avidin-biotin complex immunoperoxidase technique was used for immunostaining. RESULTS: The results of TUNEL analysis showed that the mechanism of the LGB cells pathogenesis was apoptosis. The LGB cells appeared clear striation in the control group, and the cells in experimental group decreased obviously after immunohistrochemical staining. The average density of LGB cells was (5 172 +/- 248) cells/mm(2) in control group, and (4 144 +/- 61) cells/mm(2) in experimental group. CONCLUSION: After partial optic nerve injury, the LGB cells are obviously damaged, which may be related to apoptosis of LGB cells. PMID- 12133383 TI - [Expression and significance of vascular endothelial growth factor in rat cornea after cautery with alkali]. AB - OBJECTIVE: To investigate the expression of vascular endothelial growth factor (VEGF) in rat cornea after cautery with alkali. METHODS: In Sprague-Dawley rats, inflammatory corneal neovascularization was induced by cautery with alkali. VEGF was detected in corneal samples at different times by Western-blot, and immunohistochemistry method was used to investigate the distribution of VEGF at rat cornea after cautery with alkali. RESULTS: The normal rat corneas did not express VEGF. The expression of VEGF increased with time after corneal cautery. The rat corneas were infiltrated by massive inflammatory cells that, especially adjacent to the cautery lesion, showed staining for VEGF. CONCLUSION: VEGF production by inflammatory cells participates in inflammatory angiogenesis in rat corneas. VEGF has the effects on the induction and supporting of cautery-induced angiogenesis in rat cornea. PMID- 12133384 TI - [Therapeutic regimen in Vogt-Koyanagi-Harada syndrome]. AB - OBJECTIVE: To compare the effectiveness of therapeutic regimen in Vogt-Koyanagi Harada (VKH) syndrome and determine the reasonable regimen for different patients. METHODS: Data of 82 patients with VKH syndrome, coming from all over China and referred to Zhongshan Ophthalmic Center from January 1996 to December 2000 were retrospectively analyzed. Twenty-eight patients (56 eyes) with first attack of the intraocular inflammation were treated with oral prednisone for more than one year, whereas 54 patients (108 eyes) with recurrent episodes were treated with chlorambucil for more than one year. All patients were treated with a combination of these immunosuppressives with traditional Chinese medicine herbs, and followed up more than one year after systemic treatment. The previous treatment for these 54 patients was also analyzed in an attempt to compare these results with those presented here. RESULTS: Uveitis was completely controlled in 96.4% of the patients treated with oral prednisone, and all of these patients showed improved vision after treatment. A complete control of uveitis was achieved in 94.4% and improved vision was seen in 88.0% of the patients treated with chlorambucil. All of these 54 patients, although treated with longer but intermittent systemic corticosteroids before installation of chlorambucil, showed a chronic or recurrent uveitis and most cases had markedly decreased vision as compared with those treated by us with regular systemic corticosteroids. CONCLUSIONS: Intensive and prolonged systemic treatment with corticosteroids is recommended for the patients with first attack of VKH syndrome, where as regular treatment with chlorambucil is useful for control of chronic and recurrent uveitis seen in VKH syndrome. Traditional Chinese medicine herbs may be helpful in amelioration of uveitis and reduction of side effects of immunosuppressives. PMID- 12133385 TI - [Vogt-Koyanagi-Harada syndrome: glucocorticoid therapy and visual prognosis]. AB - OBJECTIVE: To evaluate the efficacy of glucocorticoid therapy in patients with Vogt-Koyanagi-Harada (VKH) syndrome. METHODS: One hundred and thirty-six patients with VKH were treated with two regimens of oral prednisone. Regimen 1: Fifty-one patients with VKH were treated at the uveitic stage with prednisone, starting from 2 mg/kg/day tapering gradually and shifting to alternate-day treatment. The patients were treated for approximately 8 months. Regimen 2: Eighty-five patients with VKH were referrals who were treated elsewhere with systemic glucocorticoid more than 2 months with total dose equivalent to prednisone more than 2 000 mg. Systemic and ocular complications were found in some of these patients. The hypothalamus-pituitary-adrenal axis was markedly inhibited as indicated by decrease in urine free cortisol (UFC). These patients were treated for 6 - 10 months based on the patients' individual ocular situations. The visual results, frequency of recurrence of uveitis and incidence of ocular complications were compared between the two groups. RESULTS: Visual acuity >/= 0.5 and >/= 0.8 were found to be 94.1% and 79.4% respectively in patients treated with regimen 1 which was far better than the patients treated with regimen 2. On the other hand, recurrence of uveitis and ocular complications were found significantly lower in the patients treated with regimen 1, as compared with that in patients treated with regimen 2 (23.5% vs. 63.5% and 9.8% vs. 49.4% respectively). All the differences are highly significant (P < 0.001). The UFC level in the patients treated with regimen 2 was increased from (5.3 +/- 5.8) microgram/24 h to (21.9 +/- 7.2) microgram/24 h (P < 0.001). CONCLUSIONS: These results indicate that both regimens are feasible for treating patients with VKH. However, regimen 1 is far better than regimen 2 with respect to visual prognosis, frequency of recurrence of uveitis as well as incidence of ocular complications. PMID- 12133386 TI - [Therapeutic vitrectomy for severe uveitis and its complications]. AB - OBJECTIVE: To investigate the therapeutic value of vitrectomy for uveitis and its complications. METHODS: Vitrectomy was performed on 14 patients (14 eyes) with uveitis. In these 14 eyes, there were 6 eyes with severe opacity of vitreous, 5 eyes with cataract, 5 eyes with retinal detachment, and 3 eyes with poor response to medication. Pars plana lenectomy, scleral cryopexy, buckling procedure, endolaser photocoagulation, gas-fluid exchange or silicone oil intraocular tamponade were added according to the different needs. The follow-up varied from 18 to 45 months. RESULTS: Twelve of the 14 eyes obtained better visual acuity, and in the other two light perception remained; less drugs were used after operation in all cases, and the uveitis was controlled. CONCLUSION: Vitrectomy is an effective method for severe uveitis and its complications both in the improvement or stabilization of visual acuity and in the reduction or cessation of systemic treatment. PMID- 12133387 TI - [Ultrasound biomicroscopy in intermediate uveitis]. AB - OBJECTIVE: To investigate the character of intermediate uveitis by ultrasound biomicroscopy (UBM) and determine the potential and usefulness of UBM as a diagnostic procedure in intermediate uveitis. METHODS: Twenty-one cases (35 eyes) of intermediate uveitis were diagnosed by slit lamp biomicroscopy and indirect binocular ophthalmoscopy with scleral indentation. UBM was performed on 35 eyes of 21 patients with intermediate uveitis in order to determine the configuration of pars, plana peripheral retina and vitreous. RESULTS: In 31 of 35 eyes, pathological structures such as spotted or membraneous vitreous condensations, vitreoretinal adhesions with traction on the peripheral retina and ciliary body detachment were identified by UBM. In four eyes, no abnormalities were identified by UBM. CONCLUSION: UBM seems to be a valuable diagnostic technique for the evaluation of patients with intermediate uveitis. PMID- 12133388 TI - [Fundus fluorescein and indocyanine green angiographic study of Behcet's disease and Vogt-Koyanagi-Harada syndrome]. AB - OBJECTIVE: To study the choroidal and retinal changes in Behcet's disease and Vogt-Koyanagi-Harada syndrome (VKHS) using indocyanine green angiography (ICGA) and fundus fluorescein angiography (FFA). METHODS: Seventeen cases (27 eyes) of Behcet's disease and 18 cases (36 eyes) of VKHS were examined with FFA and ICGA. All of the patients showed an active intraocular inflammation when examined. RESULTS: FFA showed leakage from retinal vessels in patients with Behcet's disease. ICGA revealed choroidal abnormalities including dilatation and hyperpermeability of choroidal vessels in 11 eyes (40.7%). In VKHS, FFA revealed the hyperfluorescence and serous pigment epithelium detachments. Dilatation and hyperpermeability of choroidal vessels revealed by ICGA were found in 36 eyes (100%), multihypofluorescent dark dots in 32 eyes (88.9%), and choroidal filling defects in 28 eyes (77.8%). CONCLUSIONS: Although retinal vascular lesions are predominant in Behcet's disease, choroidal abnormalities evidenced by dilatation of choroidal vessels are also present in some patients. Hyperfluorescence and hypofluorescence revealed are the main findings in the patients with VKHS. A combination of FFA and ICGA is recommended for the examination of Behcet's disease and VKHS. PMID- 12133389 TI - [GATA-3 in the development of anterior chamber associated immune deviation]. AB - OBJECTIVE: To determine the role of GATA-3 (a transcript factor that has influences on Th2 cell differentiation) in the development of anterior chamber associated immune deviation (ACAID). METHOD: Immunohistochemistry and Western blot were used to determine the localization and relative protein levels of GATA 3 respectively at different time points after anterior chamber injection of 50 microgram interphotoreceptor retinoid binding protein (IRBP). RESULT: GATA-3 was detected weakly in normal spleen and increased significantly at 5, 7, 14 and 21 days after anterior chamber inoculation. It was detected mainly in CD(4)(+) T cells in spleen. CONCLUSION: The expression of GATA-3 increases earlier than development of ACAID. It suggests that GATA-3 play an important role in Th2 commitment during ACAID development. PMID- 12133390 TI - [A clinical analysis of 25 cases with Duane's retraction syndrome combined with congenital crocodile tears]. AB - OBJECTIVE: To analyze the clinical manifestations of Duane's retraction syndrome (DRS) combined with congenital crocodile tears or gustatory-lacrimal reflex. METHODS: Twenty-five cases of DRS associated with congenital crocodile tears were retrospectively summarized. The clinical features, including sex, age at first visit, chief complaints, laterality, types of DRS, features of abnormal lacrimation and its associated ocular and non-ocular anomalies were analyzed. RESULTS: There were 8 males and 17 females with a male-to-female ratio of 1:2. Fifteen cases had binocular involvement and the remaining 10 cases were monocular involvement. In general, the association of the abnormal lacrimation and eye movement disorder in unilateral cases was ipsilateral, while in bilateral cases the abnormal lacrimation and the movement disorder were also bilateral except 3 cases of bilateral DRS with monocular crocodile tears and one case of unilateral DRS with binocular crocodile tears. CONCLUSIONS: Clinically, cases with Duane's syndrome combined with crocodile tears are relatively few. That sufficiently pay attention to and master its clinical characteristics is the basis for its correct diagnosis. PMID- 12133391 TI - [A comparison of clinical effects between phacoemulsification and intraocular lens implantation through "L-shaped" and 3.2 mm scleral tunnel incision]. AB - OBJECTIVE: To study the implant surgery through a newly designed L-shaped sutureless scleral tunnel incision and compare the therapeutic effects of this incision with the conventional 3.2 mm scleral tunnel incisions. METHODS: Phacoemulsification was performed through a L-shaped incision with implantation of 6 mm PMMA intraocular lens (IOL) and 3.2 mm scleral tunnel incision with implantation of acrylic foldable IOL. Uncorrected visual acuity and corneal topography were examined before the operation and 1 day, 7 days, 1 month, 3 months after the operation. RESULTS: From 1 day to 3 months post-operatively, there was no statistical difference between the two groups in uncorrected visual acuity, surgical induced astigmatism and axis of corneal astigmatism. The preoperative and the post-operative corneal topographies of the two groups were similar. CONCLUSION: The therapeutic effects of the two groups are similar. In the L-shaped scleral tunnel incision group, the less expensive PMMA IOL can be used. PMID- 12133392 TI - [A novel rhodopsin E341ter mutation in patients with retinitis pigmentosa and corresponding clinical phenotype]. AB - OBJECTIVE: To detect rhodopsin (RHO) mutation in Chinese families with autosomal dominant retinitis pigmentosa (ADRP) and study on the association of RHO gene mutations with clinical phenotype. METHODS: Twenty-seven members from 13 Chinese families with ADRP and 30 normal subjects were recruited. The complete coding regions of the rhodopsin gene were amplified with polymerase chain reaction (PCR) and then DNA single-strand conformation polymorphism (SSCP) technique was used to screen RHO gene mutations. When a variant band was observed after the SSCP electrophoresis, the variant band was analyzed by sequencing PCR-amplified DNA. All subjects were examined clinically by slit-lamp, direct funduscopy, Goldmann kinetic perimetry, Humphrey threshold perimetry and electroretinogram. RESULTS: Nine affected subjects and 2 boys (11 and 9 years old respectively) in one pedigree among 13 families were found to have three DNA single strand bands by SSCP analysis. Results of assaying sequence showed the 11 members were heterozygous for rhodopsin E341ter mutation. The codon 341 is changed from GAG to TAG, resulting in a stop codon mutation. Thirty normal controls and unaffected subjects in this family were the wild type of RHO gene. Affected individuals reported night blindness in the second decade, showed optic atrophy, vessel attenuation and a few bone spicule-like pigments in the peripheral retina. The impairment of visual acuity was relatively severe, loss of peripheral visual field was greatly considerable after 30 years of age, rod and cone ERG were not detectable in the second decade, and only slight cone response was left. CONCLUSIONS: The natural history of RP in this family begins with a loss of rod function, progresses to involve the cone system, and leads eventually to a severe loss of visual function. A novel rhodopsin gene mutation E341ter is responsible for a Chinese family with ADRP. PMID- 12133393 TI - [Inhibition of IL-6 antisense oligonucleotide on IL-6 expression by human retinal pigment epithelium in vitro]. AB - OBJECTIVE: To investigate the inhibition of IL-6 antisense oligonucleotide (ASON) on IL-6 expression by retinal pigment epithelium (RPE) cells on the basis of previous study. METHODS: Cultured human RPE cells was treated with IL-1 beta and IL-6 ASON. IL-6 mRNA and protein expression were detected by enzyme linked immunosorbent assay (ELISA), immunohistochemistry and in situ hybridization histochemistry (ISH). RESULTS: It was demonstrated that the IL-6 expression by RPE cells was dose and time dependent after the stimulation of IL-1 beta. IL-6 in the conditioned media or in the control group was 2.00 x 10(-6) g/L cells after the exposed to IL-1 beta for 8 hours. IL-6 ASON (2.00 x 10(-5) mol/L) obviously inhibited IL-6 (5.50 x 10(-7) g/L cells, t = 4.518, P < 0.01) production. Immunohistochemistry study showed dark blue staining in RPE cytoplasm after stimulation with IL-1 beta, while the cells treated with IL-6 ASON showed light staining in RPE cytoplasma. ISH displayed that there was a marked reduction in mRNA expression in IL-6 ASON treated group compared with that in the control group (t = 3.746, P < 0.01). CONCLUSION: Cultured RPE cells express IL-6 protein and mRNA in dose and time dependent manners when RPE cells are stimulated with IL 1 beta. The expressions of IL-6 protein and mRNA can be significantly inhibited by IL-6 ASON in cultured human RPE cells. PMID- 12133394 TI - [A study on the activation of nuclear factor-kappa B in rat retina induced by interleukin-1 beta]. AB - OBJECTIVE: To evaluate the inductive role of interleukin (IL)-1 beta and the inhibitory role of pyrrolidine dithiocarbamate (PDTC) on the activation of nuclear factor (NF)-kappaB in the rat retina. METHODS: Sixteen Sprague-Dawley (SD) rats were randomly divided into two groups. The inductive role of IL-1 beta was observed in group A, in which 5 microl IL-1 beta (5 x 10(7) U/L) was injected intravitreally in the right (experimental) eye and PSS 5 microl in the left (control) eye. The inhibitory role of PDTC was observed in group B. In this group, 10 micro1 PDTC (1 mmol/L) was injected intravitreally in the right (experimental) eye and PSS 5 microl in the left (control) eye 1 hour before the intraocular injection of 5 microl IL-1 beta (5 x 10(7) U/L) in the bilateral eyes, respectively. Nuclear extracts was prepared from the retina 4 h and 24 h post-IL-1 beta challenge, and electrophoretic mobility shift assay (EMSA) was used to examine NF-kappa B/DNA binding activity in the retina. RESULTS: At 4 hours after IL-1 beta challenge, NF-kappa B/DNA binding activity increased in the retina of the experimental eyes in group A, and decreased in the experimental eyes of group B, compared to that of their control eyes. At 24 hours after IL-1 beta challenge, NF-kappa B/DNA binding activity in the experimental eyes of group A and in the control eyes of group B was lower than that at 4 hours, respectively. CONCLUSIONS: NF-kappa B in the retina can be activated by intraocular IL-1 beta. NF-kappa B may play an important regulative role in the mechanism of inflammatory reaction in the retina. PMID- 12133395 TI - [An experimental study of inhibition of tetrandrine on posterior capsular opacification in rabbits]. AB - OBJECTIVE: To study the effect of tetrandrine (Tet) on the formation of posterior capsular opacification (PCO) after cataract extraction. METHODS: Intraocular lens implantation was performed on 48 healthy rabbit eyes, and they were randomly divided into 4 groups: control (A), dexamethasone (B), Tet (C) and Tet with coated IOL (D). Dexamethasone and Tet were postoperatively injected into the subconjunctival space. The number of cells, the contents of protein and malondialdehyde (MDA) in the aqueous were determined at different days postoperatively. The PCO was scored and opaque posterior capsules were weighted. RESULTS: The number of cells, the amount of protein in aqueous, the scales and the wet weight of PCO in the treated eyes were less than that of the control eyes (P < 0.05). In B and C group, except the 14th day content, the postoperative MDA was lower than that in the control group (P < 0.05). CONCLUSION: Tet can inhibit the formation of PCO, and has no obvious toxicity to the corneal endothelium. PMID- 12133396 TI - [An experimental study on effect of perfluorohexyloctane on corneal endothelial cells]. AB - OBJECTIVE: To investigate the effect of perfluorohexyloctane on corneal endothelial cells in rabbit eyes. METHODS: Fifteen New Zealand albino rabbits were divided into two groups: experiment group (F(6)H(8)) and control group (BSS). All rabbits underwent anterior chamber injection of 0.15 ml F(6)H(8) or BSS. Slit-lamp biomicroscopy and corneal endothelium photography were performed pre- and post-injection. Histopathological examination and transmission electron microscopy (TEM) were performed after the rabbits were sacrificed. RESULTS: All the corneas were clear. Since 4 weeks postoperatively, the endothelial cells were markedly irregular in size and shape, and the number of endothelial cells was markedly decreased. Multilayered retrocorneal membranes grew gradually from 4 weeks after surgery. Vacuolar degeneration was seen in some endothelial cells. Nuclear degeneration and edema of endoplasmic reticulum were seen in TEM. CONCLUSION: Corneal endothelial cells degenerated after contacting with F(6)H(8) for 4 weeks. As a silicone solvent, F(6)H(8) should be removed out completely after injection. It is not recommended to apply this agent as a new intraocular temponade. PMID- 12133397 TI - [Comparative study on two different dosages of conjugated equine estrogen continuously combined with medroxyprogesterone in prevention of postmenopausal osteoporosis]. AB - OBJECTIVE: To investigate the effects of two dosages of conjugated equine estrogen (CEE) in preventing bone loss in early postmenopausal women. METHODS: Two hundreds and thirty six early postmenopausal women were randomly given one of the following regimens for two years. Groups A (GA): CEE 0.625 mg + medroxyprogesterone (MPA domestic made) 2 mg + caltrate-D (Ca-D) 1 tablet daily; Group B (GB): CEE 0.3 mg + MPA 2 mg + Ca-D 1 tablet daily; Group C (GC): Ca-D 1 tablet daily alone. The observation endpoints included: (1) bone mineral density (BMD) of lumbar 2 - 4 (L(2 - 4)) measured by duel energy X-ray absorptiometry (DEXA, Lunar DPX-L) before and 1, 2 years after treatment; (2) vaginal bleeding recorded daily and endometrium thickness yearly by transvaginal ultrasonography. Endometrium biopsies were performed if its thickness greater than 5 mm. RESULTS: Two hundreds and thirteen (90%) cases completed 1-year study, 176 (75%) 2 year study. In GA L(2 - 4) BMD significantly increased both after 1 and 2 year treatment as compared with pretreatment value (P < 0.001). While in GB, L(2 - 4) BMD significantly elevated only after 1 year treatment, but did not reach significance during the end of 2 year therapy. In contrast, L(2 - 4) BMD decreased by 0.4% and 1.6% respectively after 1, 2 year of GC although without significance. Compared among the three group, the increments of mean L(2 - 4) BMD after 1 year treatment in GA and GB were significantly different from, that in GC (+2.3%, +2.7% versus -0.4%, P < 0.001, P < 0.05 respectively). So were the values after 2 year treatment (+3.7%, +0.7% versus -1.6%, P < 0.001, P < 0.05 respectively). As compared the mean L(2 - 4) BMD between GA and GB, the difference reached significance only after 2 year (P < 0.01), but not after 1 year treatment (P > 0.05). The vaginal bleeding rate in GA during the first month and 1, 2 year after treatment were higher than those in GB and GC (52% versus 16%, 9%; 43% versus 12%, 2.8%; 34% versus 8%, 3.3%). Endometrium biopsies were carried in 153 cases (27 had endometrium thicker than 5 mm) in GA and GB. No atypical hyperplasia was found, but 2 cases showed simple hyperplasia in the GA. One case in GA developed superficial thromphlebitis during the 1 year treatment. CONCLUSION: Both 0.625 mg and 0.3 mg daily of CEE continuously combined with domestic MPA are effective in preventing postmenopausal osteoporosis. The former has more stronger effect than the latter, but needs higher dose of MPA when combined continuously in order to decrease the vaginal bleeding rate and preventing endometrium hyperplasia. PMID- 12133398 TI - [Effect of rosiglitazone on insulin resistance and hyperandrogenism in polycystic ovary syndrome]. AB - OBJECTIVE: To evaluate the effect of rosiglitazone on insulin resistance and hyperandrogenism in polycystic ovary syndrome (PCOS). METHODS: Rosiglitazone was given 4 mg daily to 30 patients with PCOS for 12 weeks. Before and after treatment, body mass index (BMI), plasma glucose, insulin, levels insulin resistance index (HOMA IR), blood lipid spectrum, leptin, neuropeptide Y, and sex hormone concentrations and ovulation rate were determined and compared. RESULTS: After 12 weeks of treatment, basal insulin level decreased from (18 +/- 8) to (12 +/- 7) mIU/L (P < 0.01), HOMA IR decreased from 4.3 +/- 1.2 to 2.6 +/- 0.7 (P < 0.01). Luteinizing hormone, free testosterone and androstenedione levels decreased [(15.4 +/- 4.4) versus (7.9 +/- 2.1) U/L, (12.5 +/- 1.9) versus (8.9 +/ 1.4) pmol/L, (9.8 +/- 1.7) versus (7.4 +/- 1.2) nmol/L respectively, P < 0.01]; Dehydroepiandrosterone sulfate level also decreased [(8.7 +/- 3.5) versus (6.9 +/ 2.1) micromol/L, P < 0.05]; Sex hormone binding globulin level increased [(39 +/ 3) versus (58 +/- 5) nmol/L, P < 0.01]. Plasma leptin level was decreased [(18 +/- 4) versus (13 +/- 3) microg/L, P < 0.01]. Ovulation rate increased to 50%. CONCLUSION: Rosiglitazone might decrease plasma leptin level and improve insulin sensitivity, which led to alleviation of hyperandrogenism and resumption of ovulation and menses in patients with PCOS. PMID- 12133399 TI - [Effects of cleavage-stage biopsy on in vitro development of human embryos]. AB - OBJECTIVE: To evaluate the effects of biopsy methods, biopsy timing and the number of cell removed on in vitro development of embryos. METHODS: One hundred and fifty four embryos of good morphology from in vitro fertilization patients were studied. Sixty-six embryos were allocated to the following three groups: chemical drilling biopsy group (26), mechanical drilling biopsy group (26) and control group (20). One cell was removed from the embryos of the two biopsy groups. The remaining 88 embryos were allocated to two groups: biopsy group (44) and control group (44). Two cells were removed from biopsy group by chemical drilling method. The stage of the embryo before biopsy, biopsy time, lysed blastomere, growth potential and hatching capacity of the biopsied embryos, total cell number at the blastocyst stage were recorded and evaluated. RESULTS: The mean time of biopsy in the chemical drilling group (231 +/- 20) seconds was significantly shorter than that in the mechanical drilling group (262 +/- 23) seconds (P < 0.01). The proportion of embryo developing to blastocyst stage was higher in chemical drilling group as compared with the mechanical group (65% versus 35%, P < 0.05). The total cell number at the blastocyst stage was fewer than those in the 7 to 8-cell embryo and >/= 9-cell embryo groups in the 6-cell embryo group (44 +/- 4 versus 49 +/- 5, 50 +/- 6; P < 0.05). At 9- to 10-cell stage, the proportion developing to the blastocyst stage was reduced in compacted embryos (20%) compared with control group (67%, P < 0.05) also more lysed blastomeres after biopsy were found in compacted embryos (50%) compared with uncompacted embryos (17%). The growth capacity to the blastocyst stage and the total cell number of the blastocyst were not different between one cell removal group and two cells removal group (P > 0.05). However, the proportion developing to the blastocyst stage was reduced after the removal of two cells from the 6 cell stage in comparison to the control (1/8 versus 5/8, P < 0.05). CONCLUSIONS: Compared with mechanical drilling biopsy, chemical drilling technique takes shorter timer and is safer. The suitable biopsy timing was >/= 7-cell stage before embryo compacting. The removal of two cells from >/= 7-cell would not impair in vitro development of embryos. PMID- 12133401 TI - [Study on clinical significance of fetal posterior fossa fluid]. AB - OBJECTIVE: To discuss the clinical significance of accumulated fluid in fetal posterior fossa. METHODS: Prenatal ultrasonography examination was performed on 5 400 woman at more than 20 weeks gestation, 110 women with fetus accumulated fluid in the posterior fossa more than 5mm were included in this study. The changes of accumulated fluid in fetal posterior fossa and the associated intracranial and extracranial anomalies were observed regularly every 2 or 3 weeks until delivery. The infants were also followed up. RESULTS: The incidence of the fetal posterior fossa fluid was 2.0%. Generally, the accumulated fluid in fetal posterior fossa was diagnosed at first time at 22 approximately 41 gestation weeks, median was (31 +/- 4) weeks. Most of them were be found between 29 approximately 32 weeks (42 cases, 38.2%), and the maximum amount of accumulated fluid in fetal posterior fossa was also in 29 approximately 32 gestation weeks (39 cases, 35.5%). The amount of accumulated fluid was from 6 mm to 26 mm, mean (11 +/- 3) mm, mostly between 10 approximately 14 mm (79 cases, 71.8%). The incidence of defected infants was 4.0%, 7.6% and 83.3% respectively, when the posterior fossa fluid was less than 10 mm, 10 approximately 14 mm and more than 15 mm. CONCLUSION: Most of cases could be diagnosed between 29 approximately 32 weeks gestation. The more fluid in the posterior fossa found, the more defected fetal or infant would be observed. In cases of more than 10 mm, especially more than 15 mm, anomales should be observed carefully. PMID- 12133400 TI - [Relationship between the immunohistopathological changes of hepatitis B virus carrier mothers' placentas and fetal hepatitis B virus infection]. AB - OBJECTIVE: To investigate the role of placenta in intrauterine transmission of hepatitis B virus (HBV), and the relationship between the fetal HBV infection and the placental infection. METHODS: Cord blood or peripheral blood was obtained from 61 newborn infants of HBsAg positive mothers. Neonates (28 of 61) infected by HBV were assigned to fetal infected group, and 33 infants without infection were assigned to control group. Histopathological changes of placentas from 61 HBsAg positive mothers were observed by pathological examination and classification. RESULTS: (1) The positive rate of HBsAg/HBcAg detected in the placentas of fetal infected group was 82% (23/28), which was significant higher than that of control group (55%), P < 0.05. (2) Among various types of cells in placental tissue, the amnionic cell showed higher HBsAg/HBcAg positive rate in fetal infection group, which was 36% (10/28), than that of control group (6%) (P < 0.01). (3) The syncitial cell was the cell with the higher detecting HBsAg/HBcAg positive rate (49%) comparing with other types of cells in placental barrier (P < 0.05). (4) The incidence of fibrinoid necrosis and chorionic hyperemia in fetal infection group were 29% and 50%, respectively, which were higher than those in control group (9%, 15%). The detecting rate of Hofbauer cell in fetal infection group was 46%, significantly lower than that in control group (79%) (P < 0.05). CONCLUSIONS: HBV infection of fetus is associated with placental infection. HBV infection of amnionic cell is an important factor of intrauterine infection. Placental barrier can protect the fetuses from infection to some extent. Some histopathological changes of placental tissue, for example fibrinoid necrosis, chorionic hyperemia and decreasing number of Hofbauer cells, may play a role in fetal HBV infection. PMID- 12133402 TI - [Study on the gestational diabetes mellitus and histocompatibility human leukocyte antigen DRB allele polymorphism]. AB - OBJECTIVE: To investigate the allelic frequency of histocompatibility leukocyte antigen (HLA)-DRB of gestational diabetes mellitus (GDM) women, in order to find out the susceptible or protective gene associated with GDM and the relationship between HLA-DRB genes and the clinical characteristic. METHODS: Thirty GDM women served as study group, 40 normal pregnant women were selected as control group. The distribution of HLA-DRB alleles frequency of 30 GDM and 40 normalpregnant women was examed by polymerase chain reaction sequence specific primer (PCR-SSP). RESULTS: There was significantly increased gene frequency of HLA-DR6 (13) in GDM women compared with normal pregnancy (12% Vs 2%, P < 0.05, RR = 5.8). The frequency of HLA-DR3 and HLA-DR9 gene in GDM women were increased, but the statistical difference was not significant (8% Vs 4%, 23% Vs 16%, P > 0.05). A much lower frequency of HLA-DR2 (15), HLA-DR51 were observed in GDM women than in normal pregnancy (10% Vs 24%, P < 0.05; 10% Vs 38%, P < 0.05). There were relationship between HLA-DR3 gene or HLA-DR6 (13)/DR9 heterozygote and severe type of GDM, relative risk (RR) were 27.1 and 8.5 respectively; HLA-DR6 (13)/DR9 heterozygote or HLA-DR3 gene were also correlated with abnormal plasma glucose levels. CONCLUSIONS: Alleles HLA-DR6 (13) were significantly implicated in their susceptibility to GDM. Conversely, HLA-DR2 (15), HLA-DR51 alleles might confer protection against GDM. HLA-DR3 gene and HLA-DR6 (13)/DR9 heterozygote were associated with severity and prognosis of GDM. It may be a marker of grade and prognosis of GDM and may direct the treatment. PMID- 12133403 TI - [Study on the effect of mouse pregnancy immunological tolerance induced by oral antigen administration]. AB - OBJECTIVE: To observe the effect of mouse pregnancy immunological tolerance induced by oral administration of antigens and investigate oral tolerance (OT) therapy of unexplained recurrent spontaneous abortion. METHODS: The mouse models of spontaneous abortion CBA/J x DBA/2 were established, then we administrated the models by oral proper dosage antigens ovum albumin (OVA), trophoblast menbrane antigens trophoblast membrane antigen (TMA)1, TMA2 and observed the changes of embryo loss rate (ELR) as compared with the control groups: group without immunization, group of active immunization by injecting intraperitoneally spleenocytes and blank control by feeding normal saline (NS). RESULTS: (1) The results showed that the ELR of group without immunization, group of active immunization and control blank were 29%, 8% and 28%, respectively. (2) Expect the groups of low dosage single-fed, the ELR can be decreased significantly by oral administration of suitable dosage of antigen OVA and TMA2 as compared with group without immunization and blank control (P < 0.05) and no changes was found as compared with group of active immunization (P > 0.05), the ELR of group OVA 2 mg x 5-fed (4%) and TMA2 2 mg x 5-fed (3%) was the lowest of all. (3) The ELR had not been decreased after administration of TMA1 groups as compared with group without immunization and group blank (P > 0.05). CONCLUSION: The results suggested that pregnancy immunological tolerance can be induced efficiently and the ELR can be decreased significantly by oral administration of proper dosage antigens of OVA and TMA2. PMID- 12133404 TI - [Combined treatment and prognostic analysis of advanced epithelial ovarian carcinoma]. AB - OBJECTIVE: To evaluate the effects of combined treatment for advanced epithelial ovarian carcinoma and to analyze its prognostic factors. METHODS: Fifty-three patients treated with a three-step combined therapeutic regimen were defined as research arm. The procedures of the three-step combined treatment were as follows: induction of tumor remission, sequential chemotherapy and adjuvant immunotherapy. Three hundred and eighteen patients with advanced epithelial ovarian carcinoma treated with cytoreductive surgery and systemic chemotherapy were retrospectively classified into control arm. RESULTS: The rates of complete response and partial response in the research arm were significantly differed from those in the control arm (90.6%, 5.7% Vs 70.1%, 5.3%, P < 0.01). The 1-, 2- and 3-year survival rates of the research arm and control arm were 97.7%, 89.1%, 83.6% Vs 71.8%, 44.1%, 29.8%, respectively (P < 0.01). The 1-, 2- and 3-year tumor-free survival rates of the research arm and control arm were 92.6%, 75.0%, 75.0% Vs 60.3%, 37.8%, 28.6%, respectively (P < 0.01). The 1- and 2-year recurrent rates of the patients in research arm were much lower than that in control arm (7.5%, 25.0% Vs 39.7%, 62.2%, P < 0.01). Age, stage, ascites, differential degree, preoperative chemotherapy (intraperitoneal and/or intravenous chemotherapy), postoperative intraperitoneal chemotherapy and systemic chemotherapy were poor prognostic factors. Initially treated in other hospital, bilateral tumors, large residuals (diameter more than 1cm) and postoperative intraperitoneal chemotherapy were poor tumor-free survival prognostic factors. CONCLUSION: Combined therapy including remission induction, consolidation chemotherapy and immunotherapy was able to enhance therapeutic efficacy, decrease the 1- and 2-year recurrent rate and improve the survival of patients with advanced epithelial ovarian carcinoma. PMID- 12133405 TI - [Role of vascular endothelial growth factor overexpression in ovarian tumor invasion and mechanism]. AB - OBJECTIVE: To study the role of vascular endothelial growth factor (VEGF) overexpression in ovarian tumor metastasis and associated mechanism. METHODS: The VEGF cDNA was transfected into ovarian tumor cell lines CAOV3 and COC1. Transfected and nontransfected cells were screened for VEGF, gelatinase A (MMP-2) mRNA and protein by reverse transcription polymerase chain reaction (RT-PCR), Western blot and substrate zymography, respectively. The invasive ability of two ovarian tumor cell lines was analysed with Boyden chamber invasion assay before and after VEGF cDNA transfection. RESULTS: The expression of VEGF, MMP-2 mRNA and protein were increased (P < 0.05) after VEGF cDNA transfection, detected by RT PCR, Western blot and substrate zymography. After VEGF cDNA transfection, the mean invasion percentage of two ovarian tumor cells [CAOV3: (42.5 +/- 4.1)%; COC1: (26.8 +/- 2.4)%] were higher than that before transfection [CAOV3: (24.7 +/ 1.9)%; COC1: (8.6 +/- 1.1)%, P < 0.05]. CONCLUSION: The expression of VEGF correlates to the in vitro invasion of ovarian tumor cell and induction of MMP-2 by VEGF is a key component of VEGF-induced ovarian tumor cell invasion. PMID- 12133406 TI - [Loss of heterozygosity at chromosome 3p14, 25 in serum DNA from ovarian cancer patients]. AB - OBJECTIVE: Investigate the frequency of loss of heterozygosity (LOH) at chromosome arm the short arm chromosome 3p14, 25 in the serum DNA from ovarian cancer and its clinical application. METHODS: Thirty-eight ovarian cancer serum samples with 18 corresponding tumor tissues and 8 benign ovarian tumors were obtained, and DNA samples extracted from serum and tissue were examined for 3p14, 25 LOH by using of polymerase chain reaction with four polymorphic microsatellite markers (D3S1029, D3S1228, D3S1300, D3S1481). RESULTS: Matched serum and tissue DNAs from 18 ovarian cancer patients showed significant concordance of 3p14, 25 LOH (P < 0.05). 3p14, 25 LOH in at least one of four loci occurred in 29 out of 38 (76%) serum samples, while 17 serum samples (45%) exhibited LOH at more than one locus. According to International Federation of Gynecology and Obstetrics (FIGO) staging, there was a trend that the rate of LOH was higher in advanced stages, and the frequency of loss in stage II, III, IV was 2/4, 78%, 8/9 respectively. It was also found that the number of microsatellite markers with 3p14, 25 LOH was increased with tumor progressed. No significant association of pathological types with LOH in serum DNA could be demonstrated except at locus D3S1029. CONCLUSIONS: Since the strong correlation of 3p14, 25 LOH between serum DNA and tumor tissue DNA, as well as the frequency of 3p14, 25 LOH associated with malignancy of ovarian cancer, it is suggested that detection of serum DNA 3p14, 25 LOH may be used as a molecular marker for staging and monitoring human ovarian cancer. PMID- 12133407 TI - [Anticancer effects of cytosine deaminase gene/5-fluorocytosine therapies to ovarian cancer in vivo]. AB - OBJECTIVE: To evaluate the anticancer effects of cytosine deaminase (CD) gene/5 fluorocytosine (5-FC) to human ovarian papillary serous cystadenocarcinoma in nude mice. METHODS: Replication-deficient adenovirus containing CD gene was used to transfect human embryonal kidney cell line 293, and the adenovirus's concentration was up to 1 x 10(10) pfu (plaque forming unit)/ml after purification. Ten female BULB/C nude mice were implanted subcutaneously with approximate 0.08 g tissue of ovarian papillary serous cystadenocarcinoma from the parental generation, and they were divided into test group and control group. While the tumors grew with diameter of approximate 3mm, test group received intratumoral injections of adenovirus-CD at 1 x 10(9) pfu in a 100 microl volume on day 1, 3, 5. These animals were administered 5-FC at doses of 400 mg/kg twice daily by intraperitoneal injection from day 1 to day 7. Normal saline was used in control group with the same volume and by the same methods. Tumor volumes were measured in following days. RESULTS: (1) Compared with control group, the growth of tumors in test group was significantly suppressed. Twenty days after the therapies, the tumor volumes in test group and control group were (91 +/- 68) mm(3) and (238 +/- 122) mm(3) respectively, there was significant difference. This difference lasted for approximately 30 days (P = 0.045 - 0.019). (2) The mean time of tumor volume doubling in test group and control group were (8.1 +/- 0.7) days and (6.4 +/- 0.7) days, respectively (P < 0.05). CONCLUSIONS: This research has shown the significant effects of CD gene/5-FC on human ovarian cancer in nude mice model. The results may be the important basis of clinical trials. They suggest that this toxic gene/prodrug therapy system may play an important role in ovarian cancer therapy in the future. PMID- 12133408 TI - [Antisense oligodeoxynucleotides of human telomerse catalytic sub-unit inhibits telomerase activity and proliferation in SKOV3 and COC1]. AB - OBJECTIVE: To study the effects of antisense oligodeoxynucleotides (ODN) of hEST2 (AODN) on telomerase activity and proliferation in ovarian cancer cell lines SKOV3 and COC1. METHODS: Antisense and sense human telomerse catalytic sub-unit (hEST2) phosphorothioate (SODN) and random ODN were designed, synthesized and transfected into SKOV3 and COC1 cells by lipofectamine. The expression of hEST2 mRNA and telomerase activity in SKOV3 and COC1 were tested by reverse transcription-polymerase chain reaction and telomeric repeat amplification protocol before and after transfection. The proliferation and growth in SKOV3 and COC1 were also investigated by methyl thiazolyl tetrazolium and growth curve before and after transfection. RESULTS: AODN could down-regulate the expression of hEST2 mRNA, inhibit telomerase activity and proliferation of ovarian cell lines. The efficiency depends on dose and period of administration. At 48 h, 30 micromol/L AODN had the highest activity. The expression of hEST2 mRNA were declined 54.6% and 44.6% in SKOV3 and COC1 respectively. And also the inhibition of telomerase activity were 47.9% and 42.7% respectively in the two cell lines. CONCLUSIONS: AODN of hEST2 clearly inhibited the proliferation of ovarian cancer cell lines. hEST2 may thus be a new target of gene therapy in ovarian carcinoma. PMID- 12133409 TI - [Effects of RelA antisense oligonucleotide on apoptosis of ovarian cancer cells COC1]. AB - OBJECTIVE: To evaluate the effects of RelA antisense oligonucleotide on apoptosis of ovarian cancer cells. METHODS: COC1 cell line was treated with RelA antisense oligonucleotide combining with tumor necrosis factor (TNF)-alpha or paclitaxel at appropriate concentrations and duration. The apoptosis and RelA activation of ovarian cancer line COC1 cell were measured by indirect immunofluorescence, Western blot, flow cytometric analysis, DNA ladder assay. RESULTS: The apoptosis increased obviously in COC1 rells if treated by RelA antisense oligonucleotide combining with paclitaxel 50 micromol/L than paclitaxel 50 micromol/L only (27.4 +/- 0.5)% vs (12.3 +/- 0.6)% (P < 0.01), when treated 12 hs; (31.7 +/- 0.3)% vs (13.0 +/- 0.5)% (P < 0.01) when treated 24 hs. The apoptosis was also increased obviously in COC1 cells if treated by RelA antisense oligonucleotide combining with TNF-alpha 50 microgram/L than TNF-alpha 50 microgram/L alone. (30.8 +/- 0.3)% vs (13.2 +/- 0.4)% (P < 0.01). CONCLUSION: RelA antisense oligonucleotide may induce apoptosis susceptibility of COC1 cells to TNF-alpha or paclitaxel. PMID- 12133410 TI - [Effects of fetal anoxia and acidosis on superoxide dismutase]. AB - OBJECTIVE: To analyze the effects of fetal anoxia, respiratory and metabolic acidosis on the activity of antioxidation in fetal distress. METHODS: Blood samples were taken from umbilical artery in 386 neonates for blood gas analysis and detection of the concentration of superoxide dismutase (SOD). Normal situation, anoxia, acidosis, respiratory acidosis, metabolic acidosis and mixed acidosis were diagnosed in all neonates according to the results of blood gas values, and the neonate asphyxia was diagnosed according to the Apgar scores (one minute). The effect of anoxia and acidosis to SOD were analyzed with multiple factor analysis of variation. RESULTS: (1) Among the all 386 cases, 317 were normal, 31 with anoxia, 17 with acidosis, and 21 with both anoxia and acidosis. Among the total cases of acidosis, 8 respiratory, 21 metabolic, and 9 mixed acidosis. (2) The plasma levels of SOD of umbilical artery blood in anoxia, acidosis, both anoxia and acidosis, and normal sitution were (118.5 +/- 7.1) mmol/L, (122.0 +/- 11.4) mmol/L, (140.0 +/- 7.0) mmol/L, and (98.5 +/- 2.6) mmol/L, respectively. The results of unvariate analysis of variance showed that anoxia: F = 4.999 (P < 0.05), acidosis: F = 7.025 (P < 0.01), and both anoxia and acidosis: F = 0.013 (P > 0.05). (3) The plasma levels of SOD with respiratory acidosis, metabolic acidosis and mixed acidosis were (127.3 +/- 18.4) mmol/L, (126.0 +/- 8.1) mmol/L, (150.0 +/- 10.4) mmol/L. The results of univariate analysis of variance showed that respiratory acidosis: F = 4.404 (P < 0.05), metabolic acidosis: F = 3.965 (P < 0.05), and mixed acidosis: F = 0.015 (P > 0.05). CONCLUSION: The superoxidation and antioxidation can be effected by factors like anoxia and acidosis, respiratory acidosis and metabolic acidosis. However, the mechanisms of these effects are different. There is additive, but not synergistic effects among them. PMID- 12133411 TI - [Investigation of relationship between pregnancy induced hypertension syndrome and homocysteine, folic acid and vitamin B(12)]. AB - OBJECTIVE: To investigate the relationship between pregnancy induced hypertension syndrome (PIH) and homocysteine, folic acid and vitamin B(12). METHODS: There were 59 cases of PIH (group A) and 60 cases of normal late pregnancy were enrolled (group B). The serum homocysteine level was detected with fluorescence polarization immunoassay (FPIA), serum folic acid and vitamin B(12) levels were detected with microparticle enzyme immunoassay (MEIA). RESULTS: There was significant difference (P < 0.05) of serum homocysteine levels between group A [(13.1 +/- 3.7) micromol/L] and group B [(10.4 +/- 3.9) micromol/L]. Although, the level of folic acid and vitamin B(12) in group A were lower than those of group B, there was no statistical differences between the two groups (P > 0.05). CONCLUSION: It was concluded that the disorder of homocysteine metabolism may play an important role in the pathogenesis of PIH, and no direct evidence to prove that folic acid and vitamin B(12) were related with PIH. PMID- 12133412 TI - [Analysis of fetal distress in pregnancy with hepatitis B virus infection]. AB - OBJECTIVE: To investigate the cause, prognosis and the treatment of fetal distress in pregnant women with hepatitis B virus (HBV) infection. METHODS: eighty one pregnant women and their newborns were selected. The HBV surface antigen (HBsAg), e antigen (HBeAg), core antibody (HBcAb) and deoxyribonucleic acid (HBV DNA) was positive and the hepatic function normal. eighty five pregnant women without HBV infection, normal hepatic function and their newborns were collected as control. The clinical data, serology examination, placenta histological examination and fetal prognosis of the two groups was analysed 76 infants of the investigation group were vaccinated hepatitis B vaccine 10 microgram at 0, 1 and 6 month respectively. Infant's hepatitis B surface antibody (HBsAb) was detected at 24 month. RESULTS: (1) The incidence of the fetal distress in investigation group was 38.3, %, in the control group was 16.5% (P < 0.05). (2) The main reason was chorion angiopathy induced by HBV infection of placenta. (3) In infants with fetal distress, the block rate from mother to fetus was 78.6%, for the infants without distress was 91.7% (P < 0.05). CONCLUSION: HBV infection during pregnancy may cause placenta chorion angiopathy, reduce placenta function, lead to fetal distress and result in immunization failure. PMID- 12133413 TI - [Comparison of Doppler examination and non-stress test in the prediction of intrauterine fetal hypoxia]. AB - OBJECTIVE: To assess the predictive value of intrauterine fetal hypoxia by Doppler examination and non-stress test (NST). METHODS: The normal value of the peak-systolic and diastolic ratio (S/D), pulse index (PI), resistance index (RI), fast volume ratio (FVR) of different gestational age was created according to 4 326 normal pregnancy. S/D, PI, RI, FVR and NST was performed for 2 873 normal pregnancy at the last trimester. Fetal outcomes were recorded within 7 days of birth. RESULTS: (1) S/D, PI, and RI decreased with the gestational age advancing, but FVR didn't. When S/D, PI, RI, FVR or NST appeared abnormal, the ratio of intrauterine fetal hypoxia was 62.0%, 60.7%, 63.3%, 27.0% and 75.7% respectively, as these normal, the ratio of intrauterine fetal hypoxia was 12.8%, 13.9%, 13.0%, 15.7%, 7.5% respectively. There was significant difference (P < 0.01). (2) Logistic regression showed that the predictive value of RI and NST to intrauterine fetal hypoxia was significant (P < 0.05). CONCLUSIONS: The predictive value of NST and RI to intrauterine fetal hypoxia was stronger than other parameters. Multi-parameter may predict more accurately than any single index for intrauterine fetal hypoxia. PMID- 12133414 TI - [Congenital vaginal atresia: report of 16 cases]. AB - OBJECTIVE: To analyze the clinical features of congenital vaginal atresia (CVA) with normal uterine corpus and investigate the relationship between the subtypes and management. METHODS: Sixteen cases, partical atresia (type I) 10 and complete atresia (type II) 6, treated in Peking Union Medical College Hospital, during the past 16 years, were analysed retrospectively. RESULTS: Patients with type I CVA had an earlier onset of symptoms [(13.0 +/- 1.1) years of age], duration of [(3.5 +/- 2.4) months], greater pelvic masses [(7.7 +/- 3.0) cm in diameter] with lower location as compared with type II CVA [(15.5 +/- 3.4) years of age, 24.0 months, (5.3 +/- 1.0) cm in diameter, P < 0.01]. Distal colpoplasty was performed on 10 type I cases and vaginal mold was placed postoperatively. Dysmenorrhea relieved in all cases during median 21 month follow-up. Vaginal dilatation was done in 4 cases due to readhesion and constriction. One out of the 2 married patients delivered viable neonate by cesarean section. Three out of 6 type II patients were complicated with either endometriosis or hematosalpinx or double uterus. Hysterectomy with or without colpoplasty was done. While the other 3 underwent colpoplasty and cervical canalization. Only 2 had menses after operation, but 1 of the developed dysmenorrhea, ovarian endometrioma and was treated surgically again, the other 1 had hypomenorrhea. CONCLUSION: Patients with type I CVA had better outcomes after colpoplasty if they persist on placing vaginal mold. While for type II CVA, hysterectomy usually needed in most cases. PMID- 12133416 TI - [Influence of advanced age on the outcome of in vitro fertilization and embryo transfer]. AB - OBJECTIVE: To determine the influence of age on the outcome of In vitro fertilization and embryo transfer (IVF-ET). METHODS: A retrospective study of 139 cycles of conventional IVF and 69 intracytoplasmic sperm injection (ICSI) cycles was performed between January 1999 and December 1999. RESULTS: A total of 208 patients (age 36 to 45 years) after IVF or ICSI were divided into five age groups (36 years group, n = 78; 37 years group, n = 49; 38 years group, n = 50; 39 years group, n = 18; 40 approximately 45 years group, n = 13). Pregnancy rate is 23.1%. There appear to be no significant difference between infertility duration, IVF cycles, fertilization rate and cleavage rate, the number and the quality of embryos transferred, and the prenatal outcome in each age group. But with the age growing, the number of follicles in each group decreased significantly (14.7 +/- 1.2, 13.0 +/- 2.0, 11.3 +/- 0.9, 9.7 +/- 0.9 and 6.5 +/- 1.9 respectively), the pregnancy rate were significantly lower (24.1, 20.5, 13.2, 11.1 and 9.8% respectively), implantation rate were significantly lower too (15.6, 11.2, 10.5, 6.5 and 2.2% respectively). But the abortion rate increase significantly (23.0, 27.2, 33.4, 41.2 and 43.3% respectively), and multiple pregnancy rate decreased (31.2, 27.3, 15.4, 6.7 and 0.0% respectively). The pregnancy rates of the ICSI and IVF were similar after stratification by age. CONCLUSIONS: The fecundity reduct significantly at the age older than 36 years old. It reduct more obviously when the age older than 40 years old. Transferring four or more embryos may increase pregnancy rate, but without increasing multiple pregnancy in women older than 40 years. PMID- 12133415 TI - [Apoptosis and expression of relative genes in early pregnant chorionic villi and decidua]. AB - OBJECTIVE: To investigate the effects of mifepristone on apoptosis and expression of relative genes in early pregnant chorionic villi and decidua. METHODS: The specimen of early pregnant chorionic villi and decidua obtained from 10 cases of requesting termination of pregnancy by curettage, 20 cases of mifepriston contragestation. The paraffin sections were used to determine apoptotic cells by TdT-mediated dUTP-biotin nick end labeling method, to identify bcl-2, bax, fas, fasL and proliferating cell nuclear antigen (PCNA) by immunohistochemistry, to demonstrate fas and fasL mRNA by in situ hybridization. RESULTS: In normal early pregnant specimens, apoptotic cells were mainly observed in syncytiotrophoblast, but not in cytotrophoblast cells, occasionally seen in decidua cells. The antigen of bax, fas, fasL were present in syncytiotrophoblast cells and decidua with lower amount. While bcl-2 antigen staining was strong in cytotrophoblastic cells and in decidua. PCNA protein was present in cytotrophoblastic and decidual cells only. In the specimens treated with mifepristone, apoptotic cells were increased in syncytiotrophoblastic cells of villi and visualized in decidua cells. The expression of fas, fasL and bax was also higher than that of normal. CONCLUSIONS: Mifepristone increased apoptosis in syncytiotrophoblastic and decidua cells, but had no effect on the expression of bcl-2 and PCNA. PMID- 12133417 TI - [Antiangiogenesis of ginsenoside Rg3 in severe combined immunodeficient mice with human ovarian carcinoma]. AB - OBJECTIVE: To investigate the antiangiogenesis of ginsenoside Rg3 in severe combined immunodeficient (SCID) mice with human ovarian carcinoma by detecting vascular endothelial growth factor (VEGF) mRNA, VEGF protein level and microvascular density (MVD). METHODS: The SCID mice with human ovarian carcinoma SKOV3 cells were treated with Rg3 (300 microgram 400 microl(-1) mouse(-1)), mice with phosphate buffered solution (PBS) and without Rg3 and PBS were used as control. Tumor volume, metastasis, ascites, VEGF mRNA, VEGF protein and MVD were detected. The level of VEGF mRNA in tumor tissue was determined by relative quantative reverse transcription polymerase chain reaction. VEGF protein level in sera and ascitic fluids were determined by enzyme-linked immunosorbent assay. MVD was calculated by immunohistochemistry (anti-CD34). RESULTS: (1) No ascites was formed and the size of metastasis decreased in SKOV3/Rg3 group. (2) Expression of VEGF mRNA level in SKOV3/Rg3 group (119 +/- 16) was lower significantly than those of the control groups (254 +/- 4, 273 +/- 44, respectively, P < 0.05). (3) Serum VEGF level in SKOV3/Rg3 group [(14.6 +/- 0.7) pg/ml] was lower significantly than those of SKOV3 group and SKOV3/PBS group [(18.5 +/- 2.1) and (20.5 +/- 1.7) pg/ml, respectively, P < 0.05]. (4) MVD in tumor tissues of SKOV3/Rg3 group (43 +/- 7) was lower than that of each control group (65 +/- 12, 73 +/- 10, respectively, P < 0.05). CONCLUSION: Ginsenoside Rg3 can block angiogenesis and inhibit tumor growth and metastasis by down regulating the expression of VEGF mRNA and protein and reducing microvascular density. PMID- 12133418 TI - [Relative risk of death and life expectancy in low cardiovascular risk population]. AB - OBJECTIVE: To assess the relationship between low cardiovascular disease (CVD) risk and the mortalities of coronary heart disease (CHD), stroke, cancer and all causes as well as life expectancy in Chinese population. METHODS: CVD risk factors were surveyed in 1982 approximately 1985 for 30 thousand male and female participants aged 35 approximately 59 from 10 Chinese population groups including 3 groups of factory workers, 6 groups of farmers and 1 group of fishermen according a uniform protocol with standardized methods. Follow-up was carried out up to 1999 approximately 2000 and the causes of death were all documented. Low cardiovascular risk was defined as: SBP/DBP < 120/80 mm Hg without anti hypertensive medication 2 weeks before the baseline survey, serum total cholesterol < 5.17 mmol/L, BMI > 18.5 but < 24 kg/m(2) and no smoking at baseline survey. Mortality rates of CHD and stroke, cancer and all causes the participants with low CVD risk were compared with the rest of the cohort and difference in life expectancy was estimated. RESULTS: Out of the whole cohort of 30 560 participants, 24 900 with complete data were available for the present analysis. Among these, 4 542 participants (18.2%) were defined as low CVD risk group, including 959 (7.7%) men and 3 583 (28.9%) women. During a follow-up of 15.2 years on average, 243 deaths including 6 CHD, 15 stroke and 117 cancer deaths took place in low risk group, while 2 383 deaths including 182 CHD, 333 stroke, and 934 cancer deaths occurred in rest of the cohort. Cox proportional hazards model adjusted for age showed that the relative risks of death of CHD, stroke, cancer and all causes in low risk group were 0.51 (95% CI: 0.19 approximately 1.37), 0.30 (95% CI: 0.11 approximately 0.81), 0.86 (95% CI: 0.62 approximately 1.19) and 0.77 (95% CI: 0.62 approximately 0.96) for men and 0.11 (95% CI: 0.03 approximately 0.45), 0.31 (95% CI: 0.17 approximately 0.58), 0.84 (95% CI: 0.66 approximately 1.10) and 0.67 (95% CI: 0.56 approximately 0.79) for women, respectively. Life expectancy was 2.6 years longer for men and 4.0 years longer for women in the low risk group. CONCLUSION: The mortality rates of CVD and all causes were lower and life expectancy was longer in the participants with low CVD risk. Keeping low risk profile may not only prevent from having CVD but also extend life expectancy. PMID- 12133419 TI - [Linkage analysis of 13 vasoactivity-regulating short tandem genes loci in essential hypertension in Chinese]. AB - OBJECTIVE: To investigate whether linkage between essential hypertension (EH) and genetic loci near renin-angiotensin system, endothelin system, nitric oxide synthase, adrenergic receptor genes in Chinese. METHODS: Linkage analysis of thirteen candidate gene loci and EH was performed in 95 Chinese nuclear families including 477 subjects using a technique of fluorescence-based gene scan with DNA short tandem repeat. The markers were selected on the chromosomal regions containing candidate genes regulating vasoactivity. GENEHUNTER package were used for two-point, non-parametric linkage analysis (NPL), maximum Lod score and transmission/disequilibrium test (TDT) in this study. RESULTS: The results of TDT indicated relatively small P values at D1S1678, D3S1744, D4S1604, D13S800, D7S483 and D8S255 (0.01 < P < 0.05). No significant linkages was found at any locus by Lod score and NPL analysis (Lod /= 0.05). CONCLUSIONS: TDT is considered as a sensitive test for detecting linkage disequilibrium. Significant linkage was found at 6 of the 13 loci typed in Chinese nuclear families with hypertension by using TDT, but no evidence suggested linkage of any locus to EH by NPL and Lod score. It is worthwhile to pay attention to chromosomal regions near the loci with small P values. PMID- 12133420 TI - [The relationship of aldosterone synthase gene polymorphism with hypertension and left ventricular hypertrophy]. AB - OBJECTIVE: To investigate the relationship of gene polymorphism of angiotension converting enzyme (ACE), aldosterone synthase (CYP11B2) with essential hypertension (EH) and left ventricular hypertrophy (LVH) in hypertensive patients. METHODS: (1) Left ventricular mass and geometry were assessed with ECG and echocardiography in 136 hypertensive patients. (2) The insertion/deletion polymorphism of ACE gene and the -344C/T polymorphism of CYP11B2 gene were monitored by polymerase chain reaction (PCR). RESULTS: (1) ACE and CYP11B2 gene showed significant difference in genotype or allele frequency distribution in the group with LVH as compared with the group without (P < 0.05). (2) There was a significant difference in CT + II combined genotype frequency distribution in the group with LVH as compared with the group without (P < 0.05). CONCLUSIONS: (1) No relationship between the polymorphism of CYP11B2 gene and ACE gene with essential hyptension could be found. (2) A significant relationship between the polymorphism of ACE and CYP11B2 gene with LVH in EH could be found. I allele of ACE gene and C allele of CYP11B2 gene may be independent risk factors for LVH in hypertensive patients. (3) There is a synergistic effect between the polymorphism of ACE gene and CYP11B2 gene to result in LVH in EH. PMID- 12133422 TI - [The role of thiol oxidants in inhibition of pancreatic exocrine secretory function and glutathione]. AB - OBJECTIVE: Glutathione depletion has been described in all form of acute pancreatitis. This study evaluated the effect of glutathione depletion on pancreatic exocrine secretory function and explored the potential mechanisms. METHODS: Thiol oxidants ethacrynic acid and diamide were employed to deplete pancreatic acinar glutathione. The effect of glutathione depletion on caerulein stimulated amylase release, cholecystokinin (CCK) receptor binding characteristics, and cytosolic free calcium was investigated. RESULTS: Both the thiol oxidants decreased the level of pancreatic acinar glutathione and caerulein (1 x 10(-10) mol/L)-stimulated amylase release in a dose-dependent manner without a marked increase in cell damage. Diamide also diminished the level of protein thiols. Ethacrynic acid and diamide both inhibited the caerulein (1 x 10(-9) mol/L)-induced Ca(2+) mobilization in pancreatic acinar cells. Neither ethacrynic acid nor diamide altered the CCK-receptor binding characteristics. CONCLUSIONS: The present findings strongly suggest that glutathione plays a potential role in the secretory process in pancreatic acinar cells and in the secretory blockade observed in acute pancreatitis. A decrease of caerulein-induced Ca(2+) mobilization might participate in the effect of glutathione depletion on pancreatic exocrine secretory function. PMID- 12133421 TI - [Gene expression of Ang II receptors in myocardium in congestive heart failure]. AB - OBJECTIVE: To explore the relation among myocardial AT(1)-/AT(2)- receptor expression, myocardial remodeling and cardiac function in patients with congestive heart failure. METHODS: Pathologic and morphologic studies on myocardial tissue of 31 patients with CHF due to valvular heart disease and 5 control subjects were carried out with optical and electronic microscopy. Message RNA expression of and AT(1)-/AT(2)-receptors in myocardial tissue were analyzed using the reverse transcriptase-polymerase chain reaction. RESULTS: Pathological changes of myocardial tissue in CHF due to valvular heart disease showed typical myocardial remodeling. AT(1)-receptor mRNA expression was slightly increased in the patients with mild CHF than in the control subjects, but decreased in the moderate and severe CHF patients. No difference was observed in AT(2)-receptor mRNA expression among all the groups. CONCLUSIONS: The expression of AT(1) receptor was up-regulated in mild CHF. Cardiomyocyte hypertrophy may be mediated by AT(1)-receptor. The expression of AT(1)-receptor was down-regulated in moderate and severe CHF. The do minant receptor subtype was transformed to AT(2). The significance of these alterations may be a local protective mechanism of myocardium. Cardiomyocyte apoptosis may be induced via AT(2)-receptor, leading to deterioration of cardiac function. PMID- 12133423 TI - [Changes of ultrastructure characteritics of Cajal interstitial cell in intestinal tract of diabetic rats]. AB - OBJECTIVE: This study is to clarify the changes of the ultrastructure characteristic of Cajal interstitial cell of diabetic rats intestinal tract. METHODS: Male SD rats were randomly divided into two groups: group A (diabetic), group B (control). 45 mg/kg Alloxan was injected into the group A, group B was injected with saline instead, after 12 weeks, the tissues of small intestine, colon were observed through electric telescope. RESULTS: The major change of Cajal interstitial cell of diabetic rat were showed as below: the number of the gap junctions between Cajal interstitial cell and neuron cells, between Cajal interstitial cell and myocyte, and between themselves were decreased significantly, and the structure of those gap junctions rested were also damaged; mitochondrion was swelling, vacuoles, dissolving; cytoplasm was dissolving, vacuoles were formed; the organelle were decreased. CONCLUSIONS: The ultrastructure of Cajal interstitial cell of diabetic had striking changes, these changes are closely related with the changes of their function, so it is very possible that these changes are one of the mechanisms of diabetic gastrointestinal dysfunction. PMID- 12133424 TI - [The effects of interleukin-12 inhalation on airway inflammation and T helper cell subsets in mouse asthma models]. AB - OBJECTIVE: To observe the effects of aerosol interleukin-12 (IL-12) on the antigen-induced airway inflammation and T helper cell subsets in mouse asthma models. METHODS: Thirty BALB/c mice were divided into 3 groups, ten in each group. The mouse asthma models (asthma group) were established by sensitization and challenge with ovalbumin (OVA). One part of sensitized BALB/c mice (IL-12 group) were interfered with aerosol IL-12 from the beginning of sensitization to the end of challenge. The healthy control mice were (control group) treated with normal saline instead of OVA. Broncho-alveolar lavage fluid (BALF) and spleen mononuclear cells (SMNCs) were collected 24h after the last challenge. The total cell numbers and eosinophils (Eos) of BALF were counted. The levels of interferon gamma (IFN-gamma) and IL-4 of BALF supernatant were measured by ELISA. The mRNA expressions of IFN-gamma and IL-4 in SMNCs were examined by reverse transcript polymerase chain reaction. RESULTS: (1) The total cell counts and Eos counts in asthma group [(12.10 +/- 2.23) x 10(9)/L and (2.02 +/- 0.85) x 10(9)/L, respectively] were significantly increased than those in control group [(6.17 +/- 1.34) x 10(9)/L and (0.05 +/- 0.01) x 10(9)/L, respectively] (P < 0.01), while in IL-12 group the above counts [(8.41 +/- 1.71) x 10(9)/L and (0.15 +/- 0.08) x 10(9)/L, respectively] were significantly decreased than those of asthma group (P < 0.05). (2) The IFN-gamma and IL-4 levels of BALF supernatant in asthma group [(43.60 +/- 15.60) ng/L and (27.15 +/- 10.42) ng/L] had significant differences from those in control group [(83.76 +/- 19.02) ng/L and (0.84 +/- 0.69) ng/L, respectively] (P < 0.01) and from those in IL-12 group [(62.30 +/- 12.50) ng/L and (6.22 +/- 2.28) ng/L, respectively](P < 0.01). (3) The IFN-gamma and IL-4 mRNA expression of SMNCs in asthma group (0.64 +/- 0.14 and 0.89 +/- 0.31), respectively) had significant differences from those in control group (1.18 +/- 0.34 and 0.71 +/- 0.16, respectively) (P < 0.05) and those in IL-12 group (0.89 +/- 0.26 and 0.79 +/- 0.25, respectively) (P < 0.05) as well. CONCLUSION: The experiment data suggested that IL-12 aerosol inhalation might inhibit antigen induced airway inflammation and partly reverse the deviation of Th cell subsets in mouse asthma models. PMID- 12133426 TI - [The clinical value of quantification of glucocorticoid receptors in peripheral blood mononuclear cells in myasthenia gravis]. AB - OBJECTIVE: To evaluate the value of glucocorticoid receptor (GR) number in predicting the glucorticoid (GC) therapeutic efficacy. METHODS: GR number in peripheral blood mononuclear cells (PBMC) from thirty patients with myasthenia gravis (MG) was measured with (3)H-dexamethansone (DEX) radio-ligand binding assay. The efficacy of GC treatment was assessed with relative clinical score one month and six months after the treatment. Twenty-four patients with MG were monitored for GR number at different time points after GC treatment. RESULTS: The pre-treatment GR number was positively correlated with relative clinical score one month and six months after the treatment (r = 0.916 and r = 0.891, P < 0.01, respectively). The higher the pre-treatment GR number, the higher the clinical relative score and the better the efficacy of GC treatment. After GC treatment, GR number had a tendency to decline gradually. CONCLUSION: Pre-treatment GR number can be used as an adjuvant parameter for predicting GC therapeutic efficacy. GC treatment down-regulates GR. PMID- 12133425 TI - [The effect of tripterine in prevention of glomerulosclerosis in lupus nephritis mice]. AB - OBJECTIVE: To study the protective effects of Tripterine on experimental lupus nephritis glomerulosclerosis. METHODS: Different doses of Tripterine were injected peritoneally to BW F1 mice at different stages. 24-hour urine protein excretion, serum anti-dsDNA antibodies, renal pathology and RT-nested PCR were analyzed to study the preventive effects of Tripterine on LN glomerulosclerosis and its mechanisms. RESULTS: (1) Tripterine suppressed the development of proteinuria, decreased the level of serum anti-dsDNA antibodies, reduced the expressions of collagen type IV, fibronectin, TIMP-1, TIMP-2, TGF-beta(1) and improved the expressions of MMP-1, -2 in the murine kidney. (2) The use of Tripterine before occurrence of proteinuria had more obvious protective effects than its use after the occurrence of proteinuria. (3) No significant difference was found between the 3 mg/kg/week Tripterine-treated-group and the 6 mg/kg/week Tripterine-treated-group. (4) No obvious change was observed in the expression of MMP-3. CONCLUSIONS: Tripterine has a definite protective effect on glomerulosclerosis of the lupus murine model. The decrease of renal collagen type IV and fibronectin is probably due to its suppressive effect on the expressions of local TGF-beta(1) and TIMP-1, -2, and its improvement effect on the local expressions of MMP-1, -2. Tripterine may have no obvious influence on MMP-3 in this model. PMID- 12133427 TI - [Function of dendritic cell in chronic hepatitis C patients]. AB - OBJECTIVE: To explore the relation between dendritic cell (DC) and the persistence of HCV in chronic hepatitis C. METHODS: The mononuclear cells were separated from the peripheral blood and cultured with the AIM-V serum-free culture to induce DC. The shape and the ultrastructure of DC, the phenotype of DC, the capacity of DC to induce allogeneicly naive T cell proliferation and the level of cytokines in supernant were investigated. RESULTS: The expression rate of CD(86) in HCV-DC was (52.4 +/- 13.3)%, which was obviously lower than that of normal-DC [(83.7 +/- 15.8)%, P < 0.01]; The capacity of proliferation of HCV-DC to induce allogeneic T cell (cpm: 19 245 +/- 2 788) was obviously lower than that of the normal-DC (cpm: 26 529 +/- 3 218) (P < 0.01); In mixed lymphocyte reaction, the average of IL-12 p(40) and interferon (IFN)gamma in HCV-DC was (404 +/- 103) ng/L and (7 891 +/- 821) ng/L, which was obviously lower than (802 +/- 205) ng/L and (13 490 +/- 1 554) ng/L of the normal-DC (P < 0.005). CONCLUSION: The function of DC in patients with chronic hepatitis C is decreased, suggesting that it is related to the persistent infection of HCV. PMID- 12133428 TI - [The effect of mycophenolic acid on activation antigen expression and in vitro T lymphocytes proliferation in peripheral blood]. AB - OBJECTIVE: To explore the effect of mycophenolic acid (MPA) on activation and proliferation of T lymphocytes in vitro. METHODS: Peripheral blood mononuclear cells (PBMNC) were prepared from normal donors, and incubated with phytohemagglutinin (PHA) with or without MPA and cyclosporin A (CSA) in vitro. After incubation, the expression of T lymphocytes activation markers such as CD(69), CD(25) and CD(3) on cell surface was assayed with flow cytometry. At the same time, incorporation rate of bromodeoxyuridine (BrdU), an analog of thymidine, was detected for T lymphocytes proliferation by flow cytometry. RESULTS: (1) Both MPA and CSA could significantly inhibit the expression of CD(3) on T lymphocyte surface. After 96 hours, the expression of CD(3) in MPA and CSA group was (37.60 +/- 7.89)% and (55.85 +/- 13.64)% respectively, while that of the control group was (74.20 +/- 7.51)%. (2) The expression of CD(3) was still significantly inhibited when MPA was added to PBMNC which was prior incubated with PHA for 72 hours. (3) MPA, CSA did not affect the expression of CD(69) on T lymphocyte surface after 24 hours. (4) Both MPA and CSA could significantly inhibit the expression of CD(25) on T lymphocyte surface. After 72 hours, the expression of CD(25) in MPA group and CSA group was (37.15 +/- 7.15)% and (62.47 +/- 12.50)% respectively, while that of the control group was (84.85 +/- 8.46)%. (5) MPA (10(-5) mol/L) significantly inhibited the incorporation of BrdU in PHA activated PBMNC (9.77 +/- 7.55)% VS (43.27 +/- 18.85)% in control (P = 0.046). Cell cycle analysis showed that MPA decreased especially the percentage of S phase cells, while the percentage of G(0)/G(1) phase cells was comparatively increased and that of G(2)/M phase cells was not changed. CONCLUSIONS: The expression of CD(3) and CD(25) was inhibited by MPA. The expression of CD(3) could be suppressed by MPA even if MPA was added to PBMNC after PHA. It is suggested that MPA could suppress T lymphocyte activation and could also reverse T lymphocyte activation. MPA could suppress T lymphocyte proliferation by down regulating the DNA synthesis. MPA decreased especially the percentage of S phase cells decreased and increased comparatively the percentage of G(0)/G(1) phase and G(2)/M phase cells. PMID- 12133429 TI - [The significance of cyclin E expression and cyclin E-dependent kinase inhibitor in adult acute leukemia]. AB - OBJECTIVE: To evaluate the function and clinical value of cyclin E and p27 in adult acute leukemia (AL). METHODS: The protein expression of cyclin E, p27, p170 and cell cycle distribution was measured in the bone marrow samples of 60 adult patients with AL and 17 controls with flow cytometric analysis. The mRNA levels of cyclin E, p27, MRP in 46 cases of AL and 17 controls were examined with reverse transcription-polymerase chain reaction method. RESULTS: The expression of cyclin E in AL patients was higher than that in controls (P < 0.01), so was the expression of p27 (P < 0.05). However in G(2)/M phase the expression rate of cyclin E in AL patients was not so apparently different with that in control (P < 0.05) so was that of p27 (P > 0.05). There was a positive correlation between the protein expression of cyclin E and p27. The remission rate (44.8%) in patients with over expression of cyclin E was lower than that (77.4%) in patients with normal expression of cyclin E (P < 0.01). The relapse rate (92.3%) in over expression group was higher than that (41.7%) in normal expression group (P = 0.003). No significant difference in remission rate or relapse rate was found between p27 over expression group and normal expression group (P > 0.50, P = 0.89). CONCLUSION: Over expression of cyclin E may implicate occurrence and progression of AL. Over expression of cyclin E is an important influential factor for remission rate and relapse rate in AL patients. PMID- 12133430 TI - [The changes in plasma leptin, insulin and proinsulin levels in obese adolescents]. AB - OBJECTIVE: To investigate the metabolic changes in obese adolescents and the relationship between obese adolescents and metabolic syndrome. METHODS: 198 obese adolescents [body mass index (BMI) >/= 25 kg/m(2)] and 78 normal adolescents (BMI 18.5 approximately 23.0 kg/m(2)) were chosen from 2 217 middle school students of 14 approximately 16 years. The levels of serum leptin, insulin, proinsulin, glucose and lipids of these adolescents were measured and compared. RESULTS: (1) There was significant gender difference of serum leptin levels in adolescents of 14 approximately 16 years. The level of serum leptin was higher in female than that in male [(18.53 +/- 1.41) microgram/L vs (6.33 +/- 1.79) microgram/L]. (2) The levels of serum leptin, insulin and proinsulin of obese adolescents were significantly higher than those of normal adolescents [(19.94 +/- 1.91) microgram/L vs (11.27 +/- 2.04) microgram/L, (15.34 +/- 1.66) microIU/L vs (13.17 +/- 1.43) microIU/L and (16.19 +/- 1.64) pmol/L vs (11.79 +/- 1.70) pmol/L respectively]. (3) The levels of blood glucose and triglyceride of obese adolescents were significantly higher than those of normal adolescents [(4.63 +/- 0.50) mmol/L vs (4.13 +/- 0.33) mmol/L and (1.20 +/- 0.56) mmol/L vs (0.90 +/- 0.32) mmol/L respectively], but the levels of serum high-density lipoprotein cholesterol (HDL-C) of obese adolescents were significantly lower than those of normal adolescents [(1.14 +/- 0.24) mmol/L vs (1.38 +/- 0.26) mmol/L]. CONCLUSION: It is suggested that there are leptin resistance, insulin resistance and potential disorders of blood glucose and lipid metabolism in obese adolescents. Our study indicated that adolescent obesity would be related with early occurrence of type 2 diabetes. PMID- 12133431 TI - [Investigation and comparison of the beta(3)-adrenergic receptor gene Trp64Arg mutation in simple obesity and metabolic syndrome]. AB - OBJECTIVE: To investigate and compare Trp64Arg mutation of beta(3)-adrenergic receptor (beta(3)-AR) gene in simple obesity and metabolic syndrome (MS), and to correlate the mutation difference with sex, total body fat and fat distribution. METHODS: Mutation study was carried out with PCR in 295 subjects with simple obesity and 219 subjects with MS. The subjects were stratified by body mass index (BMI) as light obesity group (BMI 25 approximately 29.9 kg/m(2)) and serious obesity group (BMI >/= 30 kg/m(2)) and by mean of waist circumference (WC) as abdominal adipose tissue light depot group (WC < 104 cm in male and WC < 88 cm in female) and serious depot group (WC >/= 104 cm in male and WC >/= 88 cm in female). 175 healthy non-obese subjects were selected as controls. RESULTS: The frequency of Trp64Arg mutation of beta(3)-AR gene was higher in the male subjects with MS than those with simple obesity and non-obese controls (32.25%, 21.01% and 19.91%, respectively, P < 0.05) and also higher than female ones with MS (32.35% vs 23.46%, P < 0.01). It was found by further stratification that the frequency of Trp64Arg mutation of beta(3)-AR gene was higher in the male light obesity group with MS than female light obesity group with MS. The frequency is also higher than that in the male light obesity group of the obese subjects (32.83% vs 17.35%, P < 0.05 and 32.83% vs 20.41%, P < 0.05, respectively). The frequency of mutation was higher in male abdominal adipose tissue serious depot subjects with MS than those with simple obesity (35.71% vs 20.16%, P < 0.05). CONCLUSION: Trp64Arg mutation of beta(3)-AR gene may be used as a genetic marker for MS in male. PMID- 12133432 TI - [The effects of complete Freund's adjuvant on prevention of pancrentitis and diabetes mellitus in non-obese diabetic mice]. AB - OBJECTIVE: To investigate the role of complete Freund's adjuvant (CFA) in preventing insulitis and diabetes in non-obese diabetic (NOD) mice. METHODS: Three-week-old female NOD mice were randomly divided into CFA group and phosphate buffered solution (PBS) group. The medications were injected once into the pad of hind foot with 50 microl CFA or 50 microl PBS respectively. Mice of the two groups were sacrificed at 6, 12 and 30 weeks of age respectively. Staining and counting the apoptotic beta cells in islets and evaluating insulitis severity and diabetes incidence were then carried out. The pancreatic sections were stained with hermatoxylin and eosin to score insulitis, and the apoptotic beta cells in islets were confirmed with the di-labeling technique of the TUNEL in situ end labeling method combined with standard sensitive avidin-biotin complex (sABC) immunohistochemical method. RESULTS: The insulitis score of CFA-treated mice at 6 and 12 weeks of age was significantly lower than that of PBS-treated mice at the corresponding time respectively (P(6w) < 0.05, P(12w) < 0.05). The apoptotic beta cells were fewer in CFA-treated mice than that in PBS-treated mice at 6 and 12 weeks respectively (P(6w) < 0.01, P(12w) < 0.01). A positive correlation was found between the apoptotic beta cell rates and the insulitis scores at 6 and 12 weeks of age. The diabetes incidence in CFA-treated mice was lower than that in PBS-treated mice (P = 0.001). CONCLUSION: Early treatment with CFA can diminish insulitis and prevent the development of autoimmune diabetes in NOD female mice; this may result from inhibition of apoptotic beta cells. PMID- 12133433 TI - [A preliminary study on the heterogeneity of preS2 region in hepatitis B virus]. AB - OBJECTIVE: To investigate the mutation of the preS2 gene sequence of HBV and clarify the significance of HBV quasispecies groups in the patients with chronic HBV infection. METHODS: A set of specific primers were synthesized according to HBV DNA sequence of a Chinese strain. The preS2 gene region was amplified by PCR method from the sera of 51 patients with chronic HBV infection, and the PCR products from 5 patients were subcloned into pGEM Teasy vectors. Polyacrylamide gel electrophoresis (PAGE) was employed to display the deletion mutations, clones with differential length were selected and sequenced. Sequence comparison was made to find the difference. RESULTS: When analyzed by PAGE, 2 approximately 3 bands were displayed in the PCR products from 52.9% (27/51) patients. This phenomena of multiple bands in PAGE was detected in both HBeAg (36.1%) and anti HBe (93.3%) positive patients, the difference between the 2 groups being statistically significant (P < 0.01). Analysis on deduced amino acid sequence showed the deletion mutations were in the N-terminal half of the preS2-encoded protein. CONCLUSION: There is a hot deletion region in preS2 gene sequence. These deletion mutations were more frequently found in patients with both anti-HBe and HBV DNA positivity. The deletion may influence the recognition by neutralizing antibodies. Our results suggest that there are HBV quasispecies in patients with chronic HBV infection. PMID- 12133434 TI - [The effects of exercise training on plasma tumor necrosis factor-alpha, blood leucocyte and its components in congestive heart failure patients]. AB - OBJECTIVE: To evaluate six-minute walking test (6-MWT) in treatment of CHF and to investigate the effect of exercise training on blood leucocyte and its components and plasma TNF alpha in CHF patients. METHODS: 60 cases with NYHA II-III CHF patients underwent 6-MWT within 24 hours after hospitalization. The walking distance, heartbeat, plasma TNF alpha, and blood leucocyte and its components were investigated before and after walking test. These patients were then randomized into two groups, a training group (32 cases) and a control group (28 cases). Besides routine medication, 6-MWT was performed in the training group twice a day for 8 weeks. The same parameters were redetermined after 8 weeks of follow-up. 23 cases with NYHA IV CHF patients were investigated with the same parameters in the first day after hospitalization. RESULTS: Blood leucocyte count and percentage of granulocytes and mononuclears in cases with NYHA III-IV were significantly higher than those with NYHA II. There were significant reduction of heartbeat and prolongation of walking distance in CHF patients after exercise training. Blood leucocyte count and the percentage of granulocytes and mononuclear and plasma TNFalpha level were reduced more in the training group than the control group after 8 weeks the treatment. CONCLUSIONS: There is significantly higher leucocyte count and percentage of granulocytes and mononuclears in CHF with NYHA III-IV than those with NYHA II. 6-MWT, as an exercise training, is simple, safe, and not harmful. It can not only evaluate the severity of CHF but also serve as a therapeutic measure for CHF. Moreover the excessive activation of cytokines as TNFalpha could be reduced by 6-MWT training in CHF patients. PMID- 12133435 TI - [Diaphragmatic motor evoked potential in amyotrophic lateral sclerosis patients]. AB - OBJECTIVE: To investigate the features and application of diaphragmatic motor evoked potential (DMEP) in amyotrophic lateral sclerosis (ALS) patients. METHODS: To record the latency and amplitude of diaphragmatic compound muscle action potential (DCMAP) from 37 ALS patients and 31 control subjects with surface electrodes after transcranial and transcervical magnetic stimulation, central motor conduction time (CMCT) was calculated. The percentage of forced vital capacity (%FVC) was performed in 22 ALS patients simultaneously. RESULTS: Cortical and cervical latencies of the ALS patients were longer than those of the controls, and cortical and cervical amplitudes of the ALS patients were lower than those of the controls. CMCT of the ALS patients were longer than that of the controls. Cortical latency, cortical amplitude and CMCT were all correlated with the dysfunction of the pyramidal tract. Cortical latency and CMCT were also correlated with the clinical respiratory dysfunction, cervical latency was correlated with %FVC. CONCLUSIONS: CMCT, Cortical and cervical latencies are sensitive index in DMEP parameters reflecting respiratory dysfunction in ALS, CMCT reflects cortical spinal tract function of ALS patients associated with respiration. It would be helpful to detect the neurological basis of the respiratory disturbance in ALS combining CMCT with cervical latency. PMID- 12133436 TI - [The effect of TNP-470 on the proliferation and apoptosis of lung adenocarcinoma cells]. AB - OBJECTIVE: To investigate the effect of TNP-470 on the proliferation and apoptosis of lung adenocarcinoma cells and to explore the mechanism of the effect. METHODS: The TNP-470 with different concentrations was added to cultured lung adenocarcinoma cell strain AGZY-82A and the expressions of proliferating cell nuclear antigen (PCNA), p53, bcl-2, vascular endothelial growth factor (VEGF) and VEGF receptor Flk-1 of the tumor cells were assessed by immunohistochemical methods. RESULTS: The experimental data showed that TNP-470 inhibit the expressions of VEGF and Flk-1 by tumor cells with dose-dependence. In the group of cells without TNP-470 (control group) the expression could rates of VEGF and Flk-1 were 70.42% and 50%, respectively; while in the group of cells with 10(7) microgram/L TNP-470, the expression rates of VEGF and Flk-1 decreased to 13.2% and 7.2%, respectively. Under the lnfluence of 10(4) microgram/L TNP 470, the expression rates of VEGF, Flk-1 by the tumor cells decreased gradually with prolonging of the action time. After 6 hours of action, the expression rate of VEGF decreased to 14.4% and after 12 hours, the expression rate of Flk-1 decreased to 6.2%. In the control group the proliferation rate of PCNA-marked cells was (87.03 +/- 0.75)% and the positive expression rate of p53-marked cells was (3.67 +/- 0.39)%, while in the 10(7) microgram/L TNP-470 group the proliferation rate and positive rate of p53-marked cells were 0 and (63.28 +/- 0.84)%, respectively. No expression of bcl-2 in both control and low concentration TNP-470 group. Its expression increased gradually as the concentration induced. In the 10(4) microgram/L TNP-470 group the expression of PCNA by tumor cells decreased. But the expression of p53, bcl-2 increased gradually with prolonging of the action time. CONCLUSION: TNP-470 could inhibit the proliferation and promote the apoptosis of lung adenocarcinoma cells through reduction of the VEGF autocrine and down-regulation of the VEGF receptor. PMID- 12133437 TI - [Interleukin (IL)-1beta, IL-1 receptor antagonist and IL-4 gene polymorphisms in ulcerative colitis in the Chinese]. AB - OBJECTIVE: To study whether the Chinese ulcerative colitis (UC) patients are associated with interleukin (IL)-1beta, IL-1RA and IL-4 gene polymorphisms and to analyze the relationship among gene polymorphisms, antineutrophil cytoplasmic antibodies (ANCA) and clinical classification of UC. METHODS: eighty-one UC patients and 114 healthy subjects (HS) were enrolled. IL-1beta, IL-1RA, and IL-4 gene polymorphisms were analyzed with polymerase chain reaction-restriction fragment length polymorphism and polymerase chain reaction with sequence-specific primers respectively. RESULTS: The gene frequency of allele RP2 of IL-4 in UC patients was significantly higher than that in HS (29.0% vs 11.8%, P = 0.00002), but the gene frequency of allele RP1 in HS was significantly higher than that in UC patients (88.2% vs 71.0%, P = 0.00002). OR of the genotype RP1.2 and RP2.2 were 2.71 (95% CI 1.39 approximately 5.31) and 9.04 (95% CI 1.05 approximately 203.29) respectively. There was not difference in the gene frequencies of IL 1beta, IL-1RA between UC patients and HS (P > 0.05). When UC patients were divided into ANCA positive and negative groups, it was found that there was significant difference in the gene frequencies of allele RP1, RP2 of IL-4 between two groups (63.7% vs 83.3%, 36.3% vs 16.7%, P < 0.05). CONCLUSIONS: There is correlation between the Chinese UC patients and the gene polymorphisms of intron 3 of IL-4. The gene frequency of allele RP1 in UC patients are lower, but the gene frequency of allele RP2 is significantly higher. The differences on gene frequencies of IL-4 between the UC patients and HS are mainly found in the ANCA positive UC patients. The Chinese UC patients are not associated with IL-1beta and IL-1RA gene polymorphisms. PMID- 12133438 TI - [Activation of nuclear factor-kappaB and its relationship with cytokine gene expression in colonic mucosa of ulcerative colitis patients]. AB - OBJECTIVE: To investigate the activation of nuclear factor-kappaB (NF-kappaB) and its relationship with expression of cytokine mRNA in intestinal mucosal biopsy specimens from patients with ulcerative colitis (UC). METHODS: 31 cases with UC were included in the study. 17 cases received sulfasalazine (SASP) or SASP and glucocorticoid treatment. 14 cases did not receive any medication related with UC. Normal mucosa from 11 colon cancer cases served as control. Ten pieces of intestinal mucosal biopsy specimens were obtained from each patient. NF-kappaB DNA binding activity was evaluated with electrophoretic mobility shift assay (EMSA). Expression of cytokine mRNA were studied with reversal tanscription polymerase chain reaction (RT-PCR). RESULTS: (1) The expression of IL-1beta mRNA and IL-8 mRNA was increased significantly in patients with UC, as compared with that in the control specimens (P < 0.05) and had a significant positive correlation with NF-kappaB DNA binding activity (r = 0.8363, P < 0.05; r = 0.6024, P < 0.05, respectively). (2) Glucocorticoids and SASP strongly inhibited NF-kappaB activation and signficantly decreased the expression of IL-1beta mRNA and IL-8 mRNA. CONCLUSIONS: NF-kappaB is a major and essential factor in regulating the expression of cytokine and plays a fundamental role in the pathogenesis of UC. SASP and glucocorticoids decrease cytokine expression via inhibition of NF-kappaB activation. PMID- 12133439 TI - [A clinical study of 19 cases of unrelated heterogenic bone marrow transplantation]. AB - OBJECTIVE: To investigate the outcomes and problems in unrelated bone marrow transplantation (URD-BMT). METHODS: 19 patients received URD-BMT in our institute from August 1998 to February 20, 2001. The diagnoses included chronic myelogenous leukemia (CML) (n = 9), acute lymphoblastic leukemia (ALL) (n = 6), acute myelogenous leukemia (AML) (n = 2), MDS-RAEB (n = 1) and beta-thalassemia major (n = 1). 19 pairs were HLA-A, HLA-B, HLA-DR matched with low resolution technique; 8 of them were HLA-A, B or DRB1 mismatched with high resolution technique. 2 patients received a TBI conditioning regimen, the other 17 received BUCY or modified BUCY regimen. Bone marrow was infused to 18 recipients via aorta and one through intravenous route. As graft versus host disease (GVHD) prophylaxis, all patients received CSA and MTX, 9 patients received ATG, 1 patient received CD(3)/CD(25) and 7 patients received mycophenolate mofetil. T cell depletion technique was applied in one patient. Statistics Kaplan-Meier plots for time-dependent analysis (survival, acute GVHD and engraft). Fisher' exact test was used for univariate analysis of risk factors. RESULTS: Apart from 2 patients who were under observation and one patient with CML suffered from late rejection. 16 out of the 19 patients had all been persistently engrafted. Relapse happened on day 60 in one patient who had advanced refractory ALL. Early infection within 5 weeks after BMT with discernable pathogens was documented in 5 patients. CMV infection occurred in 10 patients. 13 out of the 19 patients have survived. The disease free survival rate was 58.3%. The cumulative incidence of acute GVHD grades II approximately IV was 53.0%. CONCLUSION: Even if the results of using HLA identical unrelated donor marrow are encouraging, the high transplantation related mortality due to infection and GVHD is a major challenge. It is not likely that this technique will be extensively used. However, in cases of high risk acute leukemia or chronic myelogenous leukemia without appropriate related donor, URD-BMT might be a therapeutic measure. PMID- 12133440 TI - [Experimental treatment of respiratory failure caused by omethoate poisoning in rats]. AB - OBJECTIVE: To examine the therapeutic effect of combined use of pralidoxime-Cl and atropine with artificial ventilation on respiratory muscle paralysis caused by omethoate poisoning in rats. METHODS: Rats were administered with same doses of 2LD(50) omethoate and then treated with atropine (10 mg/kg) to resist effectively chlolinergic symptoms. When the rats had slow respiratory frequency and breathed with difficulty, the trachea was intubated and artificial ventilation was carried out (except for group A). The rats in group B were continuously treated with atropine. The doses of pralidoxime-Cl for group C, D and E were 15 mg/kg, 20 mg/kg and 40 mg/kg respectively, given at the same time as artificial ventilation and 1, 2 and 3 hours later. The dose of atropine was reduced to 1/3 to 2/3 of the first dose so as to maintain the rats atropinized. If the rat survived beyond 60 minutes after withdrawal of artificial ventilation, the combined treatment was considered successful. The function of isolated phrenic diaphragm of the rats was observed with MS-302 analyses instrument physiologically and pharmacologically. RESULTS: None of the rats in group B successfully withdraw from artificial ventilation. The rats in group C all successfully withdraw from artificial ventilation in 3 hours and the function of the isolated phrenic muscle remained good. The survival rats in group D and E were very low after withdrawal, even though the function of isolated phrenic muscle was good. CONCLUSIONS: The therapeutic effect of the combined use of suitable dose of pralidoxime-Cl and atropine with artificial ventilation on respiratory muscle paralysis caused by omethoate poisoning in rats was significant. This measure can facilitate reversal of the function of poisoned diaphragm and reduced the death rate in poisoned rats. PMID- 12133441 TI - [A long-term follow-up study of 50 patients with severe aplastic anemia who have survived more than 3 years after immunosuppressive therapy]. AB - OBJECTIVE: To evaluate the long-term outcome of immunosuppressive therapy (IST) in patients with severe aplastic anemia (SAA). METHODS: Hematopoietic recovery (peripheral blood cell counts, bone marrow aspirates, bone marrow biopsy, in vitro culture of hematopoietic progenitors), immunity of T lymphocyte, quality of life and side-effects of the therapy were assessed in 50 SAA patients who have survived more than 3 years after IST. RESULTS: At 3 years, 4 years and 5 years follow-up, 81.5% (13 cases), 86.7% (13 cases) and 89.5% (17 cases) of the SAA patients reached and maintained normal peripheral blood cell counts, 93.4% (15 cases), 93.3% (14 cases) and 94.7% (18 cases) showed normal bone marrow pictures, and 37.5% (6 cases), 40.0% (6 cases) and 73.7% (14 cases) had normal yields of bone marrow cell culture, respectively. Overall, 86.0% (43 cases), 94.0% (47 cases) and 52.0% (26 cases) of the total SAA patients were normalized in peripheral blood counts, bone marrow picture and culture of hematopoietic progenitor yields, respectively. During the follow-up, 88.0% (44 cases) of the patients achieved 100 of Karnofsky scores; 26 of the 31 patients (83.9%) who received bone marrow biopsy showed normal histological pictures, and 29 of 37 patients (78.4%) tested had normal subsets of T lymphocytes. No clonal disease was found. The late side-effects of IST were mild. All of the parameters tested were normal in 24 patients. CONCLUSION: After IST, the hematopoietic function of bone marrow, the immunity of the T lymphocyte and the life quality were normalized with few side-effects in patients with SAA. These patients would probably be cured. PMID- 12133442 TI - [Proliferative capacity of the isolated single CD(34)(+) glycosylphosphatidylinesitol-anchored (GPI) protein negative and positive hematopoietic cells in paroxysmal nocturnal hemoglobinuria]. AB - OBJECTIVES: To investigate the stroma-independent growth ability, multilineage differentiation and expansion of the single hematopoietic stem/progenitor cell from patients with paroxysmal nocturnal hematoglobinuria (PNH). METHOD: The CD(34)(+) CD(59)(+) and CD(34)(+) CD(59)(-) cells from PNH patients and CD(34)(+) CD(59)(+) cells from normal volunteers were sorted as each single cell into a well of 96 well culture plates containing culture medium supplemented with SCF, IL-3, Epo, GM-CSF, G-CSF, IL-6, Tpo and Flt-3 ligand. RESULTS: (1) Single PNH CD(34)(+) CD(59)(-) cell had a higher capacities for plating efficiency, colony (>/= 50 cells) formation and cell expansion than that of the PNH CD(34)(+) CD(59)(+) cells (P < 0.05); (2) Both the single CD(34)(+) CD(59)(-) cells from PNH patients and the single CD(34)(+) CD(59)(+) cells from normal controls had similar capacities for cell plating efficiency and colony and large colony formation. The PNH CD(34)(+) CD(59)(-) cells had a lower average cell production and cell expansion capacity. (3) The single CD(34)(+) CD(59)(+) cells from both PNH patients and normal controls showed the same capacities for cell plating efficiency and colony formation. The PNH CD(34)(+) CD(59)(+) cells exhibited much lower capacity for large colony formation, average cell production and total cell expansion. (4) A diminished secondary colony formation ability was also observed in the PNH CD(34)(+) CD(59)(+) and CD(34)(-) CD(59)(-) clones. CONCLUSION: The single PNH CD(34)(+) CD(59)(-) cells had growth advantage over the single PNH CD(34)(+) CD(59)(+) cells to some extent, but they both had impaired growth abilities as compared with CD(34)(+) cells from normal volunteers. PMID- 12133444 TI - [Study on abnormal expression of the p73 gene in childhood acute lymphoblastic leukemia]. AB - OBJECTIVE: To investigate the relationship between p73 gene and the development and progression of childhood acute lymphoblastic leukemia (ALL). METHODS: The levels of p73 transcripts in 61 ALL cell lines and 53 childhood ALL patients were assayed using reverse transcriptase-polymerase chain reaction (RT-PCR), and their relationship with the clinicopathological characteristics was analyzed. Besides, the methylation status of p73 exon 1 was analysed by restriction-enzyme related PCR, methylation-specific PCR and bisulfite genomic sequencing in the 61 ALL cell lines. RESULTS: Of the 61 ALL cell lines, 42 showed expression of p73 mRNA, with a negative rate of 31.1%, and of the 53 primary childhood ALL samples, 39 showed expression of p73 mRNA, with a negative rate of 26.4%. Loss of p73 expression was significantly associated with the reduced disease free survival and overall survival of the patients. 39.3% (24/61) of the ALL cell lines showed hypermethylation of p73 exon 1, while normal lymphocytes and most cell lines expressed p73 mRNA were not hypermethylated. CONCLUSION: There was a higher negative expression rate of p73 mRNA in childhood ALL. The main mechanism of the loss expression would be the hypermethylation of p73 gene. p73 gene inactivation might play an important role in the pathogenesis of ALL. Examination of p73 mRNA might have clinical significance in predicting prognosis of childhood ALL. PMID- 12133443 TI - [Quantity and apoptosis-related protein level of B lymphocyte in patients with immunorelated pancytopenia]. AB - OBJECTIVE: To examine the quantity and apoptosis-related protein level of B lymphocyte in the patients with immunorelated pancytopenia (IRP) and explore the role of B lymphocyte in the pathogenetic mechanism of IRP. METHODS: Quantities of all B lymphocytes and CD(5)(+) B lymphocytes and the expressions of Fas and bcl-2 on B lymphocytes in 25 patients with untreated IRP, 15 IRP patients in complete remission (CR) and 10 normal controls were assayed by FACS. RESULTS: The percentages of B lymphocyte and CD(5)(+) B lymphocytes were significantly higher in untreated IRP patients than in CR IRP patients and normal controls (P < 0.05); there was no significant difference between the latter two groups (P > 0.05). There was no significant difference of Fas expression in B lymphocytes among the three groups (P > 0.05). The expression of bcl-2 on B lymphocytes was significantly higher in untreated patients than in CR patients or normal controls (P < 0.05), and so did in CR patients than in normal controls (P < 0.01). The apoptosis-related index was significantly lower in untreated patients than in CR patients or normal controls (P < 0.01), and was lower in CR patients than in normal controls (P < 0.05). The percentage of B lymphocyte was positively correlated with the duration from the beginning of treatment to response. CONCLUSION: The production of auto-antibodies in IRP patients probably has some relationships with the abnormal quantities of B lymphocyte and its subsets, and with the inhibition of B lymphocyte apoptosis. PMID- 12133445 TI - [The correlation of cyclin A gene expression with drug resistance in adult acute leukemia patients]. AB - OBJECTIVE: To investigate the relationship between the expression of cyclin A and drug resistance in patients with adult acute leukemias. METHODS: The mRNA expressions of cyclin A, mdr1, Topo II alpha and bcl-2 were measured in 64 patients with adult acute leukemia (AL) and 20 normal subjects by semi quantitative reverse transcriptase polymerase chain reaction (RT-PCR). RESULTS: (1) The cyclin A and Topo II alpha mRNA expression levels in the treatment resistant group were significantly lower than that in the sensitive group (P < 0.01). There was no cyclin A mRNA expression in the 20 normal subjects under the same experiment condition. (2) The mdr1 and bcl-2 mRNA expression levels in the resistant group were significantly higher than that in sensitive group (P < 0.01). (3) Cyclin A and Topo II alpha gene expression levels were positively correlated (r = 0.794, P = 0.000) in the 64 AL patients, and there was a negative correlation between the gene expression levels of cyclin A and mdr1 (r = -0.337, P = 0.029) in the drug resistant group. (4) Ten AL patients with low expressions of both cyclin A and Topo II alpha were all in the resistant group. Logistic regression of binary analysis showed a significant correlation between the low expression of cyclin A and drug resistance. CONCLUSION: Low expression of cyclin A gene might be a unfavorable prognostic factor for patients with AL and measurement of both cyclin A and Topo II alpha gene expression would predict drug resistance for AL patients. PMID- 12133446 TI - [Enhancement of antigen presenting function of dendritic cells by IL-2 gene modification and its mechanism]. AB - OBJECTIVE: To investigate the effects of IL-2 gene modification enhancement of the antigen-presenting function of the mouse bone marrow derived dendritic cells and on the activation of CTL induced by MHC class I molecule restricted antigen peptides as well as the related immunological mechanisms. METHODS: DCs were prepared from mouse bone marrow and modified by recombinant IL-2 adenovirus (DC IL-2). The IL-12 and IFN-gamma levels in culture supernatant of DC and CTL were examined by ELISA, the expression of costimulatory molecules and fluorescent intensity of endocytosis of OVA-FITC in DC by FACS, the capacity of presenting 3LL cell tumor antigen by (3)H-TdR incorporation method, the MHC class I restricted tumor-antigen-peptide Mut1 of 3LL cells pulsed DC-IL-2 to induce CTL cytotoxicity by (51)Cr 4-hr releasing assay. RESULTS: After IL-2 gene modification, DC-IL-2 could produce high level of IL-12 [(78.4 +/- 6.6) pg.(1 x 10(6) cells)(-1).ml(-1)]. The expression of costimulatory molecules on DC-IL-2 was increased, the fluorescent intensity of DC captured OVA-FITC was enhanced, and the proliferation of allo-T cells from 3LL bearing mouse pulsed with Mut1 was also enhanced. Mut1 antigen peptide pulsed DC-IL-2 could induce more potent antigen-specific CTL cytotoxicity and excrete high concentration of IFN-gamma [(1 168 +/- 58.4) pg/ml] in vivo. CONCLUSION: IL-2 gene modification of DC can activate second signal for DC presenting antigen, and enhance the function for capturing and presenting tumor antigen. DC-IL-2 pulsed with MHC class I restricted tumor-antigen-peptide can induce specific anti-tumor immune response more effectively. Owing to IL-2 gene modification, the functions of IL-12 excretion and T cell activation of DC were promoted, so that the capacity of CTL excreting IFN-gamma was enhanced, which are relevant to the immune mechanism. PMID- 12133447 TI - [Expression and clinical significance of anti-apoptosis gene, survivin, in acute leukemia]. AB - OBJECTIVE: To explore the correlation between expression of surviving gene in acute leukemic cells and its clinical effects. METHODS: By using semi quantitative reverse transcriptase polymerase chain reaction (RT-PCR) technique, surviving gene expression in 50 previously untreated acute leukemia (AL) patients was analysed. The apoptosis of primary leukemia cells cultured in vitro was assayed with terminal deoxyribonucleotidyl transferase mediated dUTP-biotin nick end labeling (TUNEL). RESULTS: Surviving gene expression levels in cells of AL patients at diagnosis were significantly higher than that in normal bone marrow mononuclear cells (MNCs) (82.0% vs 33.3%, P < 0.05). The expression level was higher in ALL cells than in ANLL cells (89.5% vs 75.0%). Among 22 cases of ANLL, bone marrow remission (BMR) rate was higher in surviving gene negative expression cells from patients accepted a course of chemotherapy than in positive expression cells (83.3% vs 25.0%, P = 0.023). Among 13 ANLL patients received a course of HA regimen chemotherapy, the BMR was higher in patients surviving mRNA negative expression cells than in positive cells (100.0% vs 27.3%). Patients with surviving/beta-actin ratio>0.6 attained lower BMR. CONCLUSION: Higher expression level of surviving mRNA in AL cells may be one of the reasons that leukemic cells are insensitive to chemotherapy. PMID- 12133448 TI - [Diamide and cyclosporin A enhanced arsenic trioxide-induced apoptosis in NB4 cells]. AB - OBJECTIVE: To investigate the effects of mitochondrial membrane permeability transition pore (MPT)-opened agent diamide and MPT-closed agent cyclosporin A on arsenic trioxide (As(2)O(3))-induced apoptosis in acute promyelocytic leukemia (APL) cell line NB4. METHODS: NB4 cells were treated with As(2)O(3) alone or in combination with diamide or cyclosporin A in different concentrations. Cell apoptosis was assessed by the morphological observation, Annexin-V assay, distribution of cellular DNA contents and genomic DNA electrophoresis. The mitochondrial transmembrane potentials (DeltaPsim) were detected by flow cytometry according to the intensity of rhodamine 123 uptake in cells. RESULTS: Both diamide and cyclosporin A significantly enhanced As(2)O(3)-induced apoptosis in NB4 cells. The DeltaPsim collapse induced by As(2)O(3) was also enforced by combined treatment with diamide or cyclospo-rin A. 1 micromol/L As(2)O(3) alone treatment for 72 hours led to DeltaPsim disruption in 27.9% of cells, while combined treatment of As(2)O(3) and diamide or cyclosporin A increased DeltaPsim disruption cells to 59.7% and 42.2%, respectively. CONCLUSIONS: As(2)O(3)-induced DeltaPsim disruption possibly involves with thiol oxidation or crosslink of important components especially ANT-related molecules. PMID- 12133449 TI - [Study on tumorigenic mechanism of human leukemia cell line in nude mice]. AB - OBJECTIVE: To investigate the tumorigenic mechanisms of human leukemia cell line HL60-n in nude mice. METHODS: Different clone strains of HL60-n cells were established by limited dilution and their biological features were compared with parental HL-60 cells. RESULTS: The colony yields in soft agar, especially the large colony yields of the high tumorigenic clone strains HL60-n/A, HL60-n/B were significantly higher than that of the HL-60 cells (P < 0.01). There was no significant difference between the low tumorigenic clone strains HL60-n/E, HL60 n/F and the HL-60 cells. Ultrastructurally, the nucleus was highly abnormal, the euchromatic element of nuclear chromatin increased, the heterochromatin sparse, and the microfilaments in cytoplasm increased and disarranged in the high tumorigenic cells as compared with HL-60 cells. Cell cycle analysis by flow cytometer showed higher S phase fractions in the high tumorigenic cells. The killing activities of NK cells to the high tumorigenic clone strains were significant lower than to the contrast (P < 0.01). The histopathological features produced by the low tumorigenic leukemia cells showed that there were many inflammatory cells infiltrated, the majority of them were lymphocytes, and many tumor cells were killed especially in vessel abundant areas. By contrast, there were few inflammatory cells infiltrated in the tumors produced by the high tumorigenic cell strains. CONCLUSION: The mechanism of the high tumorigenic activity of the HL60-n cell line involved higher colony yields in soft agar, higher S phase fraction, decreased susceptibility to NK cell killing, and the inhibition of the host immunity. PMID- 12133450 TI - [Study on the in vivo killing activity of YCD/5-FC gene therapy system on K562B cells]. AB - OBJECTIVE: To elucidate the killing activity of yeast cytosine deaminase/5 fluorocytosine (YCD/5-FC) gene therapy system on gene-transferred tumorigenic cell line K562B in vivo. METHOD: K562B cell was infected with high titer virus and a gene transferred cell clone, YCD-K562B, was selected. Twelve male SCID mice of 4 week old were divided into 2 groups at random and both YCD-K562B and K562B cells were implanted to each mice. 5-FC or saline was given i. p for 10 days after tumor developed, and relative tumor volume was measured every 3 days. At the end of experiment, animals were sacrificed and the specimens were processed for histopathological examination. RESULTS: At the end of experiment (21 days after tumor cell implantation), the relative tumor volume of the 4 groups were: YCD-K562B + 5-FC 2.922 +/- 0.581, YCD-K562B + saline 24.434 +/- 4.790, K562B + 5 FC 22.701 +/- 2.350 and K562B + saline 24.460 +/- 1.670; t-test analysis showed that 5-FC could kill cells (YCD-K562B) in vivo (P = 0.0001), but had no effect on the growth of gene-untransferred cells (K562B) (P = 0.096). In YCD-K562B + 5-FC group, relative tumor volume reduced in 3 approximately 6 days after treatment (the minimum was 0.681). Necrosis around artery could be found in the tumor of YCD-K562B + 5-FC group. CONCLUSION: YCD/5-FC suicide gene therapy system has a significant in vivo killing activity to gene-transferred tumorigenic YCD-K562B cell. PMID- 12133451 TI - [Effects of the combination of eicosapentaenoic acid and retinoic acid on the proliferation and differentiation of HL-60 cells]. AB - OBJECTIVE: To study the effect of combination of eicosapentaenoic acid (EPA) and retinoic acid (RA) on the proliferation and differentiation of HL-60 cells and its mechanisms. METHODS: MTT was used for cell proliferation analysis, NBT reduction experiment for cell differentiation, reverse transcriptase polymerase chain reaction (RT-PCR) for retinoblastoma (RB) mRNA expression, and Western blot for RB protein (PRB) expression. RESULTS: The proliferation inhibition rates were 35.74%, 24.38% and 42.75% for RA, EPA and combination of EPA and RA. NBT reduction experiments showed that the differentiation induced by EPA and RA was 5.9 times, and by RA was 2.6 times the capacity of the control. The RB mRNA and PRB expression were not changed by EPA, but significantly decreased by the combination of EPA and RA. Moreover, the dephosphorylation rate of PRB was increased by the treatment with EPA or/and RA. CONCLUSION: The changes of RB expression and PRB phosphorylation may be one of the mechanisms of the synergistic effects of EPA and RA on the HL-60 cell proliferation and differentiation. PMID- 12133452 TI - [Inhibition of K562 cell growth and tumor angiogenesis in nude mice by antisense VEGF(121) cDNA transfection]. AB - OBJECTIVE: To investigate the effect of antisense vascular endothelial growth factor (VEGF)(121) cDNA transfection on the growth of K562 cells in nude mice. METHODS: K562 cells transfected with the antisense (AS) or sense (S) VEGF(121) cDNA, and the vector (V, pcDNA3) alone were transplanted subcutaneously into nude mice and the growth of the transfected cells in vivo was investigated. The effects of transfected K562 cells on human bone marrow endothelial cells (BMEC) were analyzed by MTT assay, the microvessel density (MVD) in tumor mass by vWF immunohistochemistry stain. RESULTS: K562/V tumor grew more slowly [(207.5 +/- 192.9) mm(3) vs (445.0 +/- 150.9) mm(3), P < 0.05] and K562/S tumor more rapidly than K562/V tumor did [(1 174.6 +/- 508.7)/mm(3) vs (445.0 +/- 150.9) mm(3), P < 0.01]. K562/S cell culture supernatant was more strongly in promoting the proliferation of BMEC than K562/V supernatant did, but K562/AS supernatant resulted in a marked decrease of the promoting effect as compared with K562/V's. The MVDs in K562/AS, K562/S, and K562/V tumors were [(11.0 +/- 7.6)/0.72 mm(2) vs (50.8 +/- 11.7)/0.72 mm(2) vs (18.9 +/- 7.0)/0.72 mm(2)], respectively. CONCLUSIONS: Antisense VEGF(121) cDNA transfected K562 cells show growth retardation in transplanted nude mice, decrease of tumor MVD, and decrease of promoting BMEC proliferation capacity. PMID- 12133453 TI - [The expression of human telomerase-associated protein hTERT and TEP1 in cord blood stem/progenitor cells and its significance]. AB - OBJECTIVE: To explore the regulatory effects of hTERT and TEP1 on telomerase activity in hematopoiesis. METHODS: The hTERT and TEP1 mRNA expression was detected by RT-PCR and the telomerase activi-ty by TRAP. RESULTS: In mononuclear cells (MNC) and CD(34)(-) cells, no detectable telomerase activity and hTERT mRNA expression were found. CD(34)(+) cells showed hTERT expression and a low level telomerase activity. TEP1 mRNA was detected in MNC, CD(34)(-) and CD(34)(+) cells with no significant difference in the expression level. In the CD(34)(+) cells cultured in vitro with growth factors for 7 days, the telomerase activity and the expression of hTERT mRNA were upregulated, but were downregulated in the long time culture. No significant changes in TEP1 expression was observed. CONCLUSION: In the course of hematopoiesis, hTERT mRNA expression was in accordance with telomerase activity, hTERT gene plays a crucial role in the expression of telomerase activity, while TEP1 gene plays, if any, a much smaller role. PMID- 12133454 TI - [Selected elimination of mouse alloreactive T cells by Fas-FasL passway]. AB - OBJECTIVE: To explore a new method of alleviating graft-versus-host disease (GVHD) after allogeneic bone marrow transplantation (allo-BMT) through selected elimination of mouse alloreactive T cells (ARTC) by Fas-Fas ligand (FasL) passway. METHODS: The Sca-1(+) early hematopoietic cells (EHCs) were isolated from BALB/c mouse (H-2(d)) bone marrow mononuclear cells (BMMC) by using a high gradient magnetic cell sorting system (MACS), then transferred with exogenous mouse FasL (mFasL) gene by retroviral gene transfecting technique. Afterward the transduced EHCs were expanded in vitro for one week followed by coculture with the spleen cells from BAC mouse (H-2(d) x b) as one-way mixed lymphocyte culture (OWMLC) for 6 days, then the cytotoxicity of treated BAC mouse spleen cells against Na(2)(51)CrO(4) labelling spleen cells from BALB/c mouse was observed. RESULTS: The Sca-1(+) EHCs were successfully isolated by MACS, with a purity of (89.0 +/- 6.1)%. After transferred with exogenous mFasL gene and expanded for one week, the transferred EHCs in the 6 day OWMLC with the spleen cells from BAC mouse at a ratio of five to one resulted in an obvious inhibition of the BAC mouse spleen cells cytotoxicity against the BALB/c mouse spleen cell at different effector/target ratios as compared to the control group (P < 0.01). CONCLUSION: The higher exogenous mFasL-expressing mouse EHCs can deplete ARTC against their own major histocompatibility complex (MHC) antigens in vitro. PMID- 12133455 TI - [Prophylaxis of acute graft-versus-host disease after allogeneic bone marrow transplantation by mycophenolate mofetil in a murine model]. AB - OBJECTIVE: To evaluate the effect of mycophenolate mofetile (MMF) on acute graft versus host disease (aGVHD) prophylaxis after allogeneic bone marrow transplantation (allo-BMT) in a murine model. METHODS: Acute GVHD was induced in male BALB/c mice by total-body irradiation (TBI) followed by female allogeneic (C57BL/6J) bone marrow and spleen cells transplantation. Acute GVHD was assessed both physically and histologically. Severe aGVHD was developed in the recipients and the mean survival time (MST) of untreated BM recipients was 6 days. After allo-BMT, animals were divided into 6 groups. Group 1 was given MMF (30 mg.kg( 1).d(-1)), group 2 CsA (1.5 mg.kg(-1).d(-1)) and MTX (0.4 mg.kg(-1).d(-1)), group 3 MMF (30 mg.kg(-1).d(-1)) in combination with CsA and MTX, group 4 MMF (60 mg.kg(-1).d(-1)) in combination with CsA and MTX, group 5 MMF (10 mg.kg(-1).d( 1)) in combination with CsA and MTX, and group 6 no agents for aGVHD prophylaxis. The physical signs, MST, peripheral blood counts, and aGVHD histopathologic examination were observed in all recipients. RESULTS: Animals in control group developed typical aGVHD and 100% of mortality, with a MST of 6 days. The MSTs of group 1 approximately 5 were significantly longer than that of control, being 3.4, 8.4, 9.0, 6.1 and 8.8 days longer, respectively (P < 0.05). The MSTs of groups 2 approximately 5 were longer than that of group 1 (P < 0.05), but there was no statistical significance between groups 3 approximately 5 and 2. There was no statistical difference in peripheral blood count among groups 1 approximately 5. Histopathological examination of skin, liver, and gastrointestinal tract showed typical signs of aGVHD. Animals received immunosuppressive agents (MMF, CsA, MTX) showed the less severe signs. CONCLUSIONS: MMF markedly prolonged MST of allo-BMT recipients, delayed the onset of aGVHD signs. The prophylaxis effect of CsA + MTX with or without MMF was better than that of MMF alone, synergism between MMF of 10 or 30 mg.kg(-1).d(-1) and CsA + MTX was better than that of MMF of 60 mg.kg(-1).d(-1) and CsA + MTX. PMID- 12133456 TI - [Application of FBC conditioning regimen in HLA haplotype peripheral blood stem cell transplantation]. AB - OBJECTIVE: To observe the influence of decreasing conditioning regimen intensity on the engraftment of HLA haplotype peripheral blood stem cell transplantation. METHOD: Twelve patients with leukemia, including 4 in complete remission, whose HLAs were full matched with donors, and 8 with refractory leukemia, whose HLAs were mismatched, were transplanted with G-CSF mobilized allogeneic peripheral blood stem cells after conditioned with a regimen consisting of fludarabine (30 mg/m(2) x 6 days), busulfan (4 mg/kg x 2 days) and cyclophosphamide (30 approximately 60 mg/kg x 2 days) (FBC). Donor lymphocytes were infused at day + 30, + 60 and + 90 after transplantation, respectively. Hematopoietic reconstitution was observed. Engraftment was documented by the analysis of short tandem repeats with polymerase chain reaction (STR-PCR). RESULT: Patients in HLA haplotype group received a mean number of 4.87 x 10(8)/kg donor mononuclear cells (MNC), with CD(34)(+) cells of 4.58 x 10(6)/kg and patients in HLA identical group a mean number of 4.85 x 10(8)/kg MNC with CD(34)(+) cells of 4.47 x 10(6)/kg. The mean time of white blood cell count more than 1.0 x 10(9)/L was 14 (10 approximately 18) days in HLA matched patients and 29 (11 approximately 90) days in HLA haplotype group. One three locus mismatched patient failed to engraft, but auto-hematopoiesis was recovered on day + 50. Full donor chimerism was observed in all patients except one with mixed chimera. The mixed chimera was converted into full donor chimera after three times donor lymphocyte infusion. One each died from severe acute GVHD, severe VOD and severe chronic GVHD in HLA haplotype group, and one from chronic GVHD in HLA identical group. CONCLUSION: Patients survived engraftment was not influenced by decreasing conditioning intensity as in this regimen. Haplotype stem cells could be engrafted durable in recipients by this regimen combined with donor lymphocyte infusion. PMID- 12133457 TI - [Treatment of two case childhood acute lymphoblastic leukemia by HLA-mismatched unrelated umbilical cord blood transplantation]. AB - OBJECTIVE: To explore the hematopoietic and immunologic reconstitution and transplantation-related complications of HLA one locus mismatched unrelated umbilical cord blood transplantation for the treatment of hematological malignancies. METHODS: Two children with acute lymphoblastic leukemia received HLA-mismatched unrelated umbilical cord blood transplantation. The conditioning regimens were BU-CTX (case 1) and BU-CTX plus BCNU (case 2). GVHD prophylaxis regimen consisted of cyclosporine (CsA) and mycophenolate mofetil (MMF). The patients received 14.6 x 10(7) nucleated cells/kg with 7.24 x 10(5) CD(34)(+) cells/kg and 16.24 x 10(7) nucleated cells/kg with 21.11 x 10(5) CD(34)(+) cells/kg, respectively. RESULTS: The two recipients, ANC > 0.5 x 10(9)/L occurred at day 27 and day 17, BPC > 50 x 10(9)/L at day 53 and day 46, the peripheral blood counts normalization at day 60 and day 52, the immune function normalization at day 134 and day 122 and the DNA fingerprinting showing engraftment at day 19 and day 17, respectively. The donor-recipient pair of case 1 was male to female, and the chromosome karyotype of recipients bone marrow and peripheral blood cells showed 100%, 46, XY cells at day 49. Grade II acute graft versus host disease (aGVHD) occurred at day 26 (case 1) and day 21 (case 2). The two recipients have survived for 353 days and 256 days. CONCLUSION: The hematopoietic and immunologic reconstitution in umbilical cord blood transplantation were earlier and more durable. The transplantation-related complications were less and aGVHD were milder. PMID- 12133458 TI - [Culture and pluripotentiality of human marrow mesenchymal stem cells]. AB - OBJECTIVE: To cultivate human mesenchymal stem cells (MSC) derived from fetal bone marrow and examine their pluripotentiality. METHODS: Bone marrow mononuclear cells from human fetal femur were collected by Percoll gradient centrifugation. The low density cells including MSCs were cultivated and expanded in MSCGM media. The characteristics of the multipotent MSCs were observed by implanting them into nude mice for 4 weeks. RESULTS: Human fetal marrow MSCs were successfully cultivated and differentiated in vivo into many kinds of tissues such as bone, cartilage, adipose, skeletal muscle and tendon-like tissue. CONCLUSION: Human fetal marrow MSCs were multipotent stem cells. PMID- 12133459 TI - [Endovascular embolization of cerebral arteriovenous malformation]. AB - OBJECTIVE: To evaluate the effect of endovascular embolization with alpha-NBCA on cerebral arteriovenous malformation with NBCA and emphasize the mainpoint of the embolizing technique. METHODS: 469 patients with cerebral arteriovenous malformation underwent endovascular embolization with alpha-NBCA by using microcatheter under digital subtraction angiography. RESULTS: 469 patients were treated successfully, 467 cases recovered clinically, 2 patients died. 469 patients underwent embolization therapy 1 108 times, ranging 1 to 8 times with an average of 2.3 times. 100% embolization was achieved in 155 cases, 90% approximately 100% in 93, 80% approximately 90% in 105, 70% approximately 80% in 78, 60% approximately 70% in 27, below 60% in 11. Nineteen cases received gamma knife (or X-knife) therapy alone. 117 cases received gamma-knife (or X-knife) therapy after embolization, 32 cases received operations. Six cases suffered from different complications: 1 case with cerebral infarction recovered after urokinase injection, 4 cases suffered from intracranial hemorrhage, and 1 suffered from catheter resorting in nidus. CONCLUSION: Endovascular embolization is a safe, reliable and effective treatment for cerebral arteriovenous malformation. PMID- 12133460 TI - [Stent-assisted angioplasty in treatment of symptomatic intracranial artery stenosis]. AB - OBJECTIVE: To investigate the indications, safety and effectiveness of intracranial deployment of stents in treatment of systematic intracranial artery stenosis. METHODS: Stent-assisted angioplasty was performed upon 7 consecutive patients with symptoms of intracranial artery stenosis unresponsive to anticoagulant therapy and antiplatelet therapy. The lesions involved the middle cerebral artery in 3 cases, distal end of internal carotid artery in one case, basilar artery in one case, and intracranial section of vertebral artery in 2 cases. Short-term clinical and angiographic data were obtained. RESULTS: Symptoms of procedure-related intracranial hemorrhage and intracranial haematoma developed later were found in one patient. Emergency craniotomy was performed 5 hours after the stenosis-assisted angioplasty. The remaining patients experienced alleviation of symptoms without transient ischemic attack and apoplexy during the short-term follow-up. Angiography showed that the stenosis degree reduced from 83% to 5%. 0 3 months of follow-up by transcranial Doppler showed good patency of stented lesions. CONCLUSION: Stent-assisted angioplasty effectively treats atherosclerotic intracranial stenosis lesions. Long-term follow-up and further clinical/technical research are still needed. PMID- 12133461 TI - [Value of three-dimensional digital subtraction angiography in diagnosis and planning endovascular treatment of cerebrovascular diseases]. AB - OBJECTIVE: To evaluate the value of three-dimensional (3D) digital subtraction angiography (DSA) in diagnosis and planning endovascular treatment of cerebrovascular diseases. METHODS: Standard DSA and 3D-DSA were performed in 40 adult patients (aged 21 approximately 74 years) suspected to be with cerebrovascular diseases. The schemes of endovascular treatment were worked out according to the results of 3D-DSA. RESULTS: Positive rate of standard DSA and 3D DSA were 85.0% and 95.0% respectively in diagnosis of cerebrovascular diseases. 100% occlusion was achieved in 11 patients with intracranial aneurysms and in 7 patients with arteriovenous malformations, and 100% recanalization was achieved in 9 patients with ischemic stroke. Temporary complications were found in 3 of all patients (7.9%). CONCLUSIONS: 3D-DSA is an ideal tool superior to standard DSA in diagnosis of cerebrovascular diseases and has potential value for evaluating endovascular treatment of cerebrovascular diseases. PMID- 12133462 TI - [The effects of intermittent hydromechanics on the differentiation and function of bone marrow stromal derived-osteoblasts in porous calcium phosphate ceramics]. AB - OBJECTIVE: To observe the effects of the intermittent hydromechanical stimulation on the differentiation and function of the bone marrow stromal derived osteoblasts in the porous calcium phosphate ceramics. METHODS: Rat bone marrow stromal derived-osteoblasts were seeded into porous calcium phosphate ceramic scaffolds at cells density 1 x 10(6) cells/cm(3). The cells-ceramics contrusts were cultured under rotary condition for 1 hour at 4 hours interval. After 4, 7 and 14 days cultivation, the osteoblastic phenotype markers (ALP Activity, Type I Collagen, Osteocalcin, Osteopontin, Osteonectin, and Bone Sialoprotein mRNA expression levels) were analyzed by biochemistry measurement and quantitative RT PCR technique. Static cell culture as control. RESULTS: Under rotary cell culture condition, the ALP activities and expressions of Type I collagen mRNA increased markedly and reached the peak levels at 7 days. The expressions of other four markers mRNA occurred at 4 days and reached the peak levels at 7 days, then the expressions were down-regulated at 14 days. Under static cell culture as control, the ALP activities and expressions of Type I collagen mRNA increased gradually and reached the peak levels at 14 days. The expressions of other four markers mRNA occurred at 7 days and reached the peak levels at 14 days. CONCLUSION: The intermittent hydromechanical stimulation could promote the bone marrow stromal derived-osteoblasts differentiation and function which cultured in the porous calcium phosphate ceramics in vitro. PMID- 12133463 TI - [Single nucleotide polymorphisms in methylenetetrahydrofolate reductase gene and susceptibility to cancer of the gastric cardia in Chinese population]. AB - OBJECTIVE: To explore the relationship between genetic polymorphisms in methylenetetrahydrofolate reductase (MTHFR), a central enzyme in folate metabolism that affects DNA methylation and synthesis, and the risk of adenocarcinoma of the gastric cardia (AGC). METHODS: Genomic DNA was isolated from the peripheral lymphocytes of 205 patients with AGC and 360 age-and-sex matched control subjects. Polymerase chain reaction and restriction fragment length polymorphism were used to examine the polymorphisms of MTHFR genes C6677T and A1298C and the relation between these genotypes and risk of AGC was analyzed. RESULTS: The genotype frequency of MTHFR C677CC was 35.0% among the controls and was 20.5% among the AGC patients; the genotype frequency of 677CT was 47.8% among the controls and 50.7% among the AGC patients, and the genotype frequency of 677TT was 17.2% among the controls and 28.8% among the AGC patients % (chi(2) = 17.63, P = 0.0002). The individuals with 677CT genotype or 677TT genotype had a 1.76-fold [95% confidence interval (CI), 1.13 - 2.76] or 2.97-fold (95% CI, 1.75 5.05) increased risk of developing AGC compared with those who had 677CC genotype. The distribution of MTHFR A1298C genotype was not significantly different between the two groups (chi(2) = 0.08, P = 0.96). The value of odd ratio (OR) in the individuals with 677CT//1298AA genotype (95% CI, 1.48 approximately 6.48), and the value of OR in the individuals with 677CT/1298CC genotype was 17.00 (95% CI, 1.26 approximately infinity; P = 0.042). CONCLUSION: Single nucleotide polymorphism in the MTHFR gene is a risk factor of carcinogenesis of AGC in the Chinese population. Although the MTHFR A1298C polymorphism was not associated with AGC, The risk of developing AGC in the individuals with MTHFR677CT/1298CC was 17 times higher than that in the individuals with MTHFR677CC/1298AA, thus showing that a joint effect exists between the MTHFR 677 and 1298 polymorphisms on the risk of AGC. PMID- 12133464 TI - [Mutual interaction between hepatitis C virus core protein and translin, a recombination hotspot binding protein]. AB - OBJECTIVE: To study the role of the core protein of hepatitis C virus (HCV) in HCV pathogenesis and investigate the molecular mechanism of this protein in tumorigenesis. METHODS: By using the yeast two hybrid system 3, the bait plasmids of the gene of the core protein of HCV was constructed and transfected into AH109 yeast strain. Then this transfected AH109 yeast was mated with Y187 yeast containing liver cDNA library plasmids and cultured on quadropledropout (QDO) medium covered with x-alpha-gal and assayed for alpha-gal activity. The blue colonies growing on QOD were collected to extract the plasmids to transform Escherichia coli. Plasmids were extracted from the bacterium and sequenced. After bioinformatic analysis translin, a recombination hotspot binding protein interacting with HCV core protein was found. To further prove the interaction between translin protein and HCV core protein translation was performed by using reticulocyte lysate and immunoprecipitation in vitro. RESULTS: Among the 30 positive colonies, a colony was translin. The interaction between HCV core protein and translin protein could be proved not only in yeast, but also in vitro. CONCLUSION: The core protein of HCV can interact with translin protein, this can explain partly the tumorigenesis of HCV. PMID- 12133466 TI - [Effect of zinc L-lysine and taurine against teratogenesis caused by hyperthermia]. AB - OBJECTIVE: To study the effect of zinc L-lysine and taurine against teratogenesis caused by hyperthermia in pregnant mice. METHODS: Eighty seven female Swiss mice with the gestational age of 8 days were exposed to 42.5 approximately 43.0 degrees C waterbath for 9 minutes. Six hours later the same procedure was repeated for 8 minutes. On the 9th gestational day zinc L-lysine (200 mg/kg) was perfused orally into 30 pregnant mice, and taurine (300 mg/kg) was perfused orally into 27 pregnant mice. The other 30 mice were used as controls. On the 18th gestational day, the pregnant mice were all killed. The teratogenic rate, embryo rate, and absorbed embryo rate, body weight, body length, and tail length of each mouse were examined. RESULTS: The teratogenic rates in zinc L-lysine group and taurine group were 1.1% and 1.2% respectively, both significantly lower than that in the control group (5.4%, P < 0.01). The dead embryo rates in the zinc L-lysine group and taurine group were 1.0% and 1.5% respectively, both significantly lower than that in the control group (3.5%, P < 0.01). The absorbed embryo rates in the zinc L-lysine group and taurine group were both 5.6%, significantly lower than that in the control group (14.3%, P < 0.01). The mean weight of the first two groups was 1 370 mg +/- 180 mg and 1 380 mg +/- 190 mg respectively, significantly greater than that in the control group (1 280 mg +/- 240 mg, P < 0.01). The mean body length in the first two groups was 28.1 mm +/- 1.9 mm and 28.0 mm +/- 2.1 mm respectively, significantly greater than that in the control group (26.7 mm +/- 2.2 mm, P < 0.01). The mean tail length in the first two groups was 12.2 mm +/- 1.3 mm and 12.2 mm +/- 1.9 mm respectively, significantly greater than that in the control group (11.6 mm +/- 1.5 mm, P < 0.01). CONCLUSION: Zinc L-lysine and taurine have a significant effect against teratogenesis caused by hyperthermia and promote the embryo growth. PMID- 12133465 TI - [Immunogenicity and efficacy of two live attenuated hepatitis A vaccines (H(2) strains and LA-1 strains)]. AB - OBJECTIVE: To assess the efficacy and immunogenicity of two live attenuated hepatitis A vaccines. METHODS: Randomized and controlled clinical trials were conducted in Guanxi, Hebei and Shanghai, 457 251 children were enrolled. The efficacy for preventing clinical hepatitis A was calculated by the comparison of incidence rate of disease between vaccine group and control group. Susceptible subjects tested anti-HAV negative before the study were followed up after vaccination for determination of the immunogenicity and vaccine efficacy to prevent subclinical infection. RESULTS: The protective efficacy to prevent clinical infection by both H(2) and LA-1 vaccines were 95%. The peak of seroconversion was observed in 94.9% and 86.0% respectively for the two vaccines. The seroconversion rate decreased to 75% approximately 80% in the third year, but the vaccine protection against clinical hepatitis A has remained unchanged throughout the 3 years. CONCLUSION: Both strains of the live attenuated hepatitis A vaccines have good immunogenicity and high protection against clinical disease, the efficacy to prevent subclinical infection is not significant. The subclinical HAV infection serves as a natural booster for the vaccinees. PMID- 12133467 TI - [Relationship between interleukin-1B and interleukin-1 receptor antagonist gene polymorphisms and susceptibility to gastric cancer]. AB - OBJECTIVE: To investigate the polymorphisms of interleukin-1B (IL-1B) promotor region 31 C/T and interleukin-1 receptor antagonist (IL-1RN) gene and the relationship between the genotype of IL-1 and IL-1RN and susceptibility to gastric cancer in Chinese population. METHODS: Genomic DNA was extracted from the juxta-cancerous normal mucosal biopsies of 50 patients with gastric cancer taken by endoscopy and from the peripheral lymphocytes of 50 normal controls and then was subjected to PCR amplification. The PCR product was digested by restriction endonuclease Alu I. The product of digestion was subjected to 2% agarose gel electrophoresis and ethidium bromide staining. RESULTS: The IL-1B -31C allele was detected in 75% of normal controls and 53% of patients, while the IL-1B -31T allele was detected in 25% of normal controls and 47% of patients. The frequency of -31C/-31C was 56% (28/56) and 30% (15/50) in normal controls and patients respectively. The frequency of -31C/-31T was 38% (19/56) and 46% (23/50) in normal controls and patients respectively. The frequency of -31T/-31T was 6% (3/56) and 24% (12/50) in normal controls and patients respectively. IL-1B -31T carriers were at increased risk of gastric cancer with an odds ratio of 7.5 for 31T/-31T homozygotes (95% CI, 2.0 approximately 27.6) and of 2.3 for -31C/-31T (95% CI, 0.9 approximately 5.4) respectively. Only three of the five kinds of polymorphism of IL-1RN (1/1, 1/2 and 2/2) were found in this study. The frequency of 1/1 was 54% (27/50) and 24% (12/50) in normal controls and patients respectively. The frequency of 1/2 was 38% (19/50) and 50% (25/50) in normal controls and patients respectively. The frequency of 2/2 was 8% (4/50) and 26% (13/50) in normal controls and patients respectively. The IL-1RN * 1 allele was detected in 73% of normal controls and 49% of patients; while the IL-1RN * 2 allele was detected in 27% of normal controls and 51% of patients. IL-RN * 2 carriers were at increased risk of gastric cancer with an odds ratio of 7.31 for 2/2 homozygotes (95% CI 2.13 approximately 25.09) and of 2.96for 1/2 heterozygotes (95% CI 1.22 approximately 7.21), respectively. CONCLUSION: The polymorphism of promotor region -31C/T of IL-1B gene and the polymorphism of IL 1RN genes 1/2 and 2/2 are associated with the susceptibility of gastric cancer in Chinese. Carrying -31T allele increases the risk of gastric cancer. Polymorphism of IL-1RN and IL-1B gene may be used as indicators of susceptibility of gastric carcinogenesis. PMID- 12133468 TI - [Effects of different enteral nutrients on gut absorptive capacity and energy metabolism during gut ischemia/reperfusion]. AB - OBJECTIVE: To evaluate the effects of different enteral nutrients on intestinal absorptive capacity and ATP content of intestinal mucosa after ischemia reperfusion (I/R) of the intestine. METHODS: A segment of jejunum was isolated to form a pouch in Sprague-Dawley rats. The jejunal pouches of animals were filled with either a mixture of nutrients (5 mmol/L alanine + 5 mmol/L glucose), glucose alone (10 mmol/L), alanine alone (10 mmol/L) or mannitol alone (10 mmol/L) as a nonabsorbable osmotic control. The superior mesenteric artery was then occluded for 60 minutes, followed by reperfusion for 60 minutes. Gut absorptive capacity, as measured by the changes in short circuit current (DeltaIsc), and mucosal ATP content were determined both at the end of 60 minutes ischemia and 60 minutes of reperfusion. RESULTS: DeltaIsc and ATP levels were decreased after I/R. Intraluminal alanine alone or alanine/glucose mixture further reduced DeltaIsc and ATP content. On the other hand, intraluminal glucose significantly increased DeltaIsc and ATP levels compared with intraluminal alanine, alanine/glucose mixture or I/R alone. CONCLUSION: Intraluminal alanine increases the absorptive capacity and energy metabolism of the gut caused by I/R, and indicated that intraluminal glucose improves gut absorptive capacity and energy metabolism and provides a protective effect on gut from ischemia and reperfusion injury. PMID- 12133469 TI - [CT-guided percutaneous needle biopsy and intra-articular injection of sacroiliac joint]. AB - OBJECTIVE: To investigate the clinical value of CT-guided percutaneous needle biopsy and intra-articular injection of sacroiliac joint. METHODS: CT-guided percutaneous puncture was conducted to the bilateral sacroiliac joints of 3 specimens of adult pelvis, 18 patients with ankylosing spondylitis (AS), and 10 patients with undifferentiated spondyloarthropathy (uSpA) suspected as early AS clinically. All 28 patients had taken non-steroidal anti-inflammatory agents without adequate control of symptoms or with serious side effects. Biopsy of synovium with 16G or 18G needles and intra-articular injection of steroid were made to the 28 patients. RESULTS: The general success rate of puncture was 4/6 among the specimens of pelvis and was 96.4% (54/56) among the 28 patients. The level of needle insertion was 1.69 cm +/- 0.63 cm up the lower edge of the joint. The obtainment rate of biopsy specimen was 88.5% (46/54) for the cases punctured successfully. The positive rate of pathological examination was 91.7% (22/24). Marked inflammatory changes were found in the biopsy specimens from AS patients and early inflammatory changes were found in the biopsy specimens from uSpA patients. The immediate effective rate of intra-articular steroid injection was 78.6% (22/28). No complication and side effect was found. CONCLUSION: CT-guided percutaneous puncture is an effective and safe guided method for biopsy and intra articular injection of sacroiliac joint. The proper level of puncture is by the juncture of the middle 1/3 and lower 1/3 of sacroiliac joint. Needle biopsy helps early diagnosis of AS for patients with uSpA. Intra-articular injection is an effective supplementary therapy for AS and uSpA patients. PMID- 12133470 TI - [Inhibition of proliferation of human vascular smooth muscle cells by overexpression of p21 gene]. AB - OBJECTIVE: To observe the apoptosis of human vascular smooth muscle cells (HVSMCs) transfected with p21 gene and investigate its potential mechanism. METHODS: An adenoviral expression vector with full-length cDNA of p21 gene insert (Ad-p21) was constructed and transfected into HVSMC. A mock vector, pAdeno-X-lac Z constructed with the same method, was transfected into HVSMCs as control. Western blotting and DS-PAGE were used to detect the expression of p21 gene. Fluorescence staining was used to observe the changes in nuclei of apoptotic cells and calculate the cell survival rate. Expression of tissue transglutaminase (tTG) in HVSMC was examined by DIG-marked probe. RESULTS: Fluorescence staining showed marked fragmentation of nucleus and pyknosis of chromatin in pAdeno-X-p21 transfected cells. The expression of tTG was at a higher level in pAdeno-X-p21 transfected cells than in normal cells and pAdeno-X-lac transfected HVSMCs. CONCLUSION: Overexpression of p21 gene inhibits the proliferation of HVSMC through apoptosis induced by p21 which may relate with overexpression of tTG. PMID- 12133471 TI - [Application of distraction osteogenesis in severe maxillary hypoplasia secondary to cleft palate]. AB - OBJECTIVE: To evaluate the effect of distraction osteogenesis in correction of severe undeveloped maxilla secondary to cleft palate and to evaluate the selection of distraction modality. METHODS: Distraction osteogenesis was performed upon 8 patients, 6 males and 2 females, aged 11 to 25 years, one with incomplete cleft palate, 4 with unilateral complete left palate, and 3 with bilateral complete cleft palate, all accompanied with severe maxillary hypoplasia. The 7 adult patients were treated by modified high-stepped Le Fort I osteotomy, the other patient, a child, was treated by Le Fort I osteotomy followed by rigid external distraction. X-ray films of skull in lateral, frontal, or panoramic positions, and X-ray film of temporomandibular joint in Schiller's position were taken pre- and postoperatively. In model surgery the accurate distraction distance was measured and the direction of distraction designed. Orthodontic therapy was started immediately after the distraction was completed and the distractor was removed after a period of 4 months' consolidation. RESULTS: The designed distraction was achieved in a11 8 patients. Except in one case with a bone defect 1 cm x 0.5 cm at the osteotomy line in the lateral wall of the right maxillary sinus, dense new bone was formed in the area of distraction in the other cases. The postoperative occlusal relationship was good and stable. No infection and other complication was found. During an average of 20 months' follow-up, the maxilla and occlusion were stable, and no relapse was found. The distance of forward distraction of the maxilla reached an average of l2 mm (5 approximately 15 mm). The SNA angle increased from 71 degree on average preoperatively to an average of 79 degree postoperatively. Both the facial appearance and occlusal relationship returned to normal in a11 the patients. CONCLUSION: Without need of bone grafting and with a stable effect and little influence on platatopharyngeal competence, distraction osteogenesis is an effective method of correcting maxillary hypoplasia secondary to cleft palate and is worthy of spreading. PMID- 12133472 TI - [Disruption of the D(2) dopamine receptor produces adrenergic and endothelin B receptor-dependent hypertension]. AB - OBJECTIVE: To assess whether D(2) receptors participate in the regulation of genetic hypertension. METHODS: Arterial pressure was measured in mice mutant for the D(2) dopamine receptors, and the interactions between D(2) dopamine receptors and other vasopressor systems were also studied. RESULTS: Both systolic blood pressure (BP) in D(2) mutant (D(2)-/-) and hybrid (D(2)+/-) mice (128 +/- 2 mm Hg, 129 +/- 4 mm Hg) and diastolic BP (107 +/- 2 mm Hg, 108 +/- 3 mm Hg) were higher than in wild-type (D(2)+/+) mice (104 +/- 2 mm Hg, 77 +/- 1 mm Hg). The BP after alpha-adrenergic blockade decreased to a greater extent in D(2)-/- than in D(2)+/+ mice. Epinephrine excretion was greater in D(2)-/- than in D(2)+/+ mice and adrenalectomy decreased blood pressure to a greater extent in D(2)-/- than in D(2)+/+ mice. The non specific endothelin B (ETB) blocker (BQ788) decreased BP in D(2)-/- but had not in D(2)+/+ mice. The ETB1 blocker RES 701 - 1 increased BP in D(2)-/- but not D(2)+/+ mice. ET-1 and the ETB agonist, sarafatoxin S6c increased BP to a greater extent in D(2)+/+ than in D(2)-/- mice. Circulating ET-like immunoreactive peptides were similar in D(2)-/- and D(2)+/+ mice but ETB receptor expression was greater in D(2)-/- than in D(2)+/+ mice. In contrast, blockade of ETA and V(1) vasopressin receptors had no effect on BP in either D(2)-/- and D(2)+/+ mice. The hypotensive effect of the AT(1) antagonist, losartan, was also similar in D(2)-/- and D(2)+/+ mice. The greater basal sodium excretion and lower renal Na(+)/K(+)ATPase activity in D(2)-/- than in D(2)+/+ mice indicate that sodium retention was not the cause of hypertension in these animals. CONCLUSION: Hypertension in the D(2) mutant mice may be caused by absent inhibitory effect of D(2) receptors on sympathetic outflow coupled with increased ETB2 activity. PMID- 12133473 TI - [Expression of vascular endothelial growth factor and its receptor (Flt-1) in breast carcinoma]. AB - OBJECTIVE: To investigate the expression of vascular endothelial cell growth factor (VEGF) and its receptor (Flt-1) in breast carcinoma and the relationship of such expression with angiogenesis, tumor invasion and metastasis. METHODS: In situ hybridization and immunohistochemistry technique were used to test the expression of mRNA and protein expression of VEGF and flt-1 in 48 specimens of breast carcinoma. 48 specimens of tissues near cancer, 3 specimens of fibroadenoma of breast, and 3 specimens of normal breast were used as controls. RESULTS: VEGF mRNA and its protein were expressed highly in breast carcinoma cells with the positive rate of 75% and 70.8% respectively. There was almost no expression of VEGF in vascular endothelial cells. Flt-1 mRNA and its protein were expressed only in a small quantity in a few tumor cells. The amount of blood vessel positive in Fit-1 mRNA and its protein was (26 +/- 12) piece/0.72 mm(2) and (24 +/- 12) piece/0.72 mm(2) respectively. The microvascular density (MVD) was significantly greater in VEGF and Flt-1 higher expression groups than in lower groups (P < 0.01). Both VEGF expression and Flt-1 expression were well correlated with the histological grade and lymph node metastasis. CONCLUSION: VEGF promotes angiogenesis by paracrining in breast carcinoma, and takes part in tumor invasion and lymph node metastasis. Test of the expression of VEGF and Flt 1 may act as a reliable index to determine angiogenesis, malignancy, invasion and metastasis. Blocking their secretion and effect may act as a new treatment for breast carcinoma. PMID- 12133474 TI - [Cerebral blood flow perfusion imaging by single photon emission computed tomography in patients with early syphilis]. AB - OBJECTIVE: To study the cerebral blood flow perfusion imaging by single photon emission computed tomography (SPECT) in patients with early syphilis. METHODS: SPECT was performed with (99m)Tc-ECD as imaging agent on 32 patients with early syphilis and 15 healthy controls. Visual analysis was made. RESULTS: Disseminated patchy radioactive rarefection mainly in cerebral cortex, suggesting hypoperfusion of regional cerebral blood flow, was shown in all patients. In particular 14 patients displayed multi-regional obvious hypoperfusion patches: 6 in left frontal lobe, 3 in right frontal lobe, 7 in left parietasal lobe, 6 in right parietal lobe, 11 in left temporal lobe, 5 in right temporal lobe, 3 in left occipital lobe, 3 in left basal ganglia, 1 in cerebellum, and 1 in intrecerebral nucleus. No abnormality was found in the controls. CONCLUSION: Cerebral blood flow abnormality exists in patient with early syphilis manifesting as patchy hypoperfusion in SPECT imaging. PMID- 12133475 TI - [Searching for the susceptibility loci of psoriasis in Chinese]. AB - OBJECTIVE: To search for the susceptibility genes of patients with psoriasis in Chinese. METHODS: 38 psoriasis families were determined to search for the susceptibility loci by selecting 19 reported polymorphic microsatellite DNA markers and making use of fluorescein-labeled PCR with Mega BACE sequencer. RESULTS: The significant two-point Lod score values of parametric linkage analysis were obtained with D6S1610 marker (1.18 at theta; = 0.2) and D17S944 marker (1.60 at theta; = 0.1); the NPL values greater than 1.6 (P < 0.05) of non parametric analysis were obtained with D6S1610 marker, D17S944 marker and D17S785 marker. CONCLUSION: A putative linkage to D6S1610 marker, D17S944 marker and D17S785 marker. PMID- 12133476 TI - [Immunogenicity study of HPV 6b virus-like particles]. AB - OBJECTIVE: To confirm human papillomavirus (HPV) 6b virus-like particles (VLP) have strong immunogenicity and the protective antibody induced by HPV 6b VLP have cross-reactive immunity against HPV11 VLP and bovine papillomavirus (BPV) 1 VLP. METHOD: The late gene L1 for HPV6b, HPV 11 and L1/L2 for BPV 1 were molecularly cloned into recombinant baculovirus, respectively. The recombinant viruses were expressed in insect cells (Sf-9 cells). The expressed L1 proteins self-assembled into virus-like particles (VLP) for HPV6b, HPV 11 and BPV 1. VLP were purified from insect cell nuclei by CsCl centrifugation. The Balb/c mice were immunized on days 0 and 21 with 50 microgramHPV6b VLP intramuscularly. Sera were collected after a further 7 days and 3 months. The titers of IgG against HPV 6b VLP, HPV 11 VLP and BPV1 VLP were detected. Hemagglutination inhibition assay was conducted to detected that whether antisera produced by HPV 6b VLP immunization could inhibit HPV11 VLP and BPV 1 VLP agglutinate mouse red blood cells. RESULT: After 7 days of two immunizations, the titers of IgG against HPV6b VLP, HPV11 VLP and BPV1 VLP were 1:6 400, 1:1 600 and 1:1 600 by ELISA, respectively. Three months later, the titers of IgG against HPV6b VLP, HPV11VLP and BPV1 VLP were 1:800, 1:400 and 1:100, respectively. Hemagglutination inhibition assay results showed that the antisera produced by HPV6b VLP inhibit HPV6b VLP and HPV11 VLP to mouse red blood cells binding. CONCLUSION: HPV 6b VLP have potent immunogenicity. Antisera produced by HPV6b VLP could inhibit the binding of HPV6b VLP and HPV11 VLP and cells. Both HPV6b and HPV11 share neutralizing epitopes which are cross reactive and HPV6b VLP may be used in prophylactic and therapeutic vaccine for HPV6b and/or HPV 11 infections. PMID- 12133477 TI - [The effects of antioxidants on pyridinoline cross linkage formation in human fibroblasts culture from hypertrophic scars]. AB - OBJECTIVE: To investigate the effects of antioxidants on the pyridinoline crosslink formation in human fibroblasts culture from hypertrophic scars. The role of these antioxidants in reducing the level of pyridinoline crosslinks in vitro without damaging the cells has important implications. These antioxidants can be applied for the treatment of hypertrophic scars. METHODS: 9 cases of hyprotrophic scar biopsies were taken from burn patients. The specimens were explant culture for 7 days. Insulated fibroblasts from these explant cultures were subcultured for 8 passages. At the last passage, the cultures were subjected to a free radical generating system and were added different antioxidants. The supernatant and cells were collected after 4 hrs of incubation and the concentration of pyridinoline crosslinks was determined using Pyrilink kit. RESULTS: When the fibroblast cultures were subjected to free radicals and antioxidants, all samples showed a decrease in pyridinoline crosslinks concentration. The more potent antioxidants found in this study were catalase, superoxide dismutase and desferrioxamine. Particularly catalase which did not damage the cells. CONCLUSION: The antioxidants used in this study were found to be effective in reducing the concentration of pyridinoline crosslinks. The best agent is catalase, which did not damage the cells and could be considered for clinical use. PMID- 12133478 TI - [Cloning expression and characterization of a multifunctional anticoagulated peptide gene in E. Coli]. AB - OBJECTIVE: To investigate the feasibility about the cloning, expression and characterization of multifunctional anticoagulated peptide. METHODS: We designed a construct using glutathione S-transferase (GST) as a protein vector, fused via a cleavable linker to an antithrombotic peptide of 31 amino acids. The peptide was designed to include three inhibitory regions: (1) the Lys-Gly-Asp (KGD) amino acid sequence to prevent fibrinogen binding to platelets; (2) a part of fibrinopeptide A, an inhibitor of thrombin; and (3) the tail of hirudin, a potent direct antithrombin. The amino acid sequence of the 31 amino acid peptide was reverse translated, and the gene was chemically synthesized and cloned into an expression vector pGEX-5X-3 as a 3'fusion to the GST gene. Gene expression was induced in E. coli DH5 alpha cells and the fusion protein was purified using affinity chromatography. RESULTS: The purified fusion protein significantly lengthened the activated partial thromboplastin time (74.7 s tested in 80 micromol/L) and thrombin time (102.3 s tested in 80 micromol/L) and inhibited the amidolytic activity of thrombin 11% activity compared with the control tested in 100 micromol/L. The ADP-induced platelet aggregation was markedly inhibited by the purified fusion protein. The study has also shown that GST exhibits relevant activity of antiplatelet weaker than the purified fusion protein. 20.7% activity compared with the control tested in 100 micromol/L. CONCLUSION: Our results confirm that it is feasible to design a hybrid multifunctional protein that targets various components of the haemostatic process. PMID- 12133479 TI - [The influence of HBV infection on TRAIL-induced apoptosis and its mechanism]. AB - OBJECTIVE: The aim of this study was to investigate whether Hepatitis B virus will influence TRAIL (TNF-related apoptosis inducing ligand) induced apoptosis of the target cells. METHODS: Two human hepatocellular carcinoma cell lines, HepG2 and its HBV whole genome transgenic cell line named HepG2.2.15 were observed in the experiments. The apoptosis rates of these two cell lines to TRAIL were examined by flow cytometry. The expression of the membrane-bound TRAIL and the secretory soluble TRAIL in the supernatant were assessed by flow cytometry and enzyme-linked immunosorbent assay, respectively. The expression of the mRNA of TRAIL receptors and FLIP were assessed by semi-quantitive PCR. RESULTS: In contrast to HepG2 cell line, the apoptosis rate of HepG2.2.15 cell line to TRAIL was significantly low (9.12% vs 51.6%, P < 0.01). The mRNA expression of four TRAIL receptors related to apoptosis were decreased to a more or less degree on HepG2.2.15 cells compared with HepG2 cells, but the expression of FLIP, a protein inhibiting apoptosis at the initial level of the caspase cascade, was drastically upregulated on HepG2.2.15 cells (P < 0.01). There's no significant difference in the secretory TRAIL expression between these two cell lines. CONCLUSION: Our results indicate that HBV could make cells resistant to TRAIL-induced apoptosis, this may be realized by the down-regulated expression of TRAIL and its receptors and up-regulated expression of FLIP. This offer us another explanation why HBV could escape the immune surveillance and persistently exist in the body, which may explain the pathogenesis of HBV related disease in a novel way. PMID- 12133480 TI - [Establishment of human hepatocellular carcinoma cell line with spontaneous pulmonary metastasis through in vivo selection]. AB - OBJECTIVE: To establish a hepatocellular carcinoma (HCC) cell line from lung metastatic lesions of human HCC in nude mice so as to provide suitable model for the study of lung-metastasis-related molecular mechanisms. METHODS: HCC clone cells MHCC97-H were inoculated subcutaneously into nude mice. The pulmonary metastatic lesions were harvested from the moribund animals and then re-implanted into the nude mice for the second round of selection. The same procedure was repeated twice. New cell line from the third round of lung metastasis was thus established and the following parameters were studied: morphology, in vitro growth curve, plate efficiency, migration, karyotype, flow cytometry, immunocytochemistry for AFP, albumin and cytokeratin 8 (CK8), flourescent PCR for HBV DNA, tumorigenicity and spontaneous metastasis via subcutaneous injection and orthotopic implantation of tumor tissue. RESULTS: The cell line established from nude mouse lung metastasis was designated as HCCLM3, consisting of polygonal epithelial cells with hypotriploid karyotype, its main range of chromosome being 55 - 58. The cell population doubling time was 34.9 hours, plate efficiency 32.4 +/- 3.2%, and cell migration rate 20 +/- 2 microm/h. The cells were positive for AFP, albumin, CK8, and P16 and negative for P53, nm23 and HBsAg. Fluorescent PCR showed HBV DNA integration into cellular genome. When 5 x 10(6) cells were injected subcutaneously into nude mice, the tumorigenicity was 100% with a latency period of 11 +/- 1 d. Five weeks after subcutaneous injection, the pulmonary metastatic rate was 100%, the median number of lung metastases being 121 per mouse. After orthotopic implantation of tumor tissue into nude mouse liver for 35 d, wide spread regional and distant metastases occurred, the metastatic rate was 100% for abdominal wall, 80% for organs in abdominal cavity, 100% for liver, 70% for diaphragm, and 100% for lung. The median number of lung metastatic lesions was 268 per mouse. CONCLUSION: A new HCC cell line has been established, which is characterized by high pulmonary metastasis via both subcutaneous and orthotopic inoculation. It provides a new model for the study of liver cancer metastasis. PMID- 12133481 TI - [The "Fas counterattack": a mechanism for immune evasion in human hilar cholangiocarcinomas]. AB - OBJECTIVE: To investigate FasL expression in hilar cholangiocarcinoma tissues and cultured cholangiocarcinoma cells, and to assess the its ability to induce apoptosis. METHODS: The expression of FasL in 48 specimens of human hilar cholangiocacinoma resected in operation and cultured QBC939 cholangiocarcinoma cell line was detected by RT-PCR, immunohistochemistry, and Western blotting. TUNEL and flow cytometry were used to detect apoptotic cells. RESULTS: Prevalent expression of FasL was detected in 48 resected hilar cholangiocarcinomas examined. TUNEL staining disclosed a high level of cell death among lymphocytes infiltrating FasL positive areas of tumor. FasL mRNA and protein was expressed in cholangiocarcinoma cells. Apoptosis was seen in 60% of Jurkat cells cultured together with QBC939 Cholangiocarcinoma cells. CONCLUSION: The hilar cholangiocarcinoma eludes immunological surveillance by inducing, via the Fas/FasL system, the apoptosis of activated lymphocytes. PMID- 12133482 TI - [Effect of recombinant human augmenter of liver regeneration on gene expression of tissue inhibitor of metalloproteinase-1 in rat with experimental liver fibrosis]. AB - OBJECTIVE: To investigate the influence of recombinant human augmenter of liver regeneration (hALR) on gene expression of tissue inhibitor of metalloproteinase-1 (TIMP1) in rat with experimental liver fibrosis. METHODS: Carbon tetrachloride was injected into the peritoneal cavities of 150 rats twice a week for 8 weeks. Albumin sensitization and caudal vein attack were conducted to another 136 rats for the duration of 3.5 months, so as to establish two kinds of animal model of experimental liver fibrosis. Recombinant hALR at different doses was given during the process of model making. Specimens of liver were taken at different time points, then the total RNA of liver tissues was isolated and TIMP1 gene expression levels were measured by reverse transcription polymerase chain reaction. RESULTS: In both rat models, TIMP(1) gene expression levels increased gradually during the process of model making. The TIMP(1) gene expression level in CCl4 model treated with low dose hALR was 0.86 +/- 0.13, 1.77 +/- 0.23, 2.78 +/- 0.36, and 3.76 +/- 0.50 respectively 2, 4, 6, and 8 weeks after the beginning of model making, all lower than those in the model group, however, without significant difference. The TIMP(1) gene expression level in CCl4 model treated with high dose hALR was 0.63 +/- 0.10, 1.18 +/- 0.20, 1.89 +/- 0.30, 2.63 +/- 0.33 respectively 2, 4, 6, and 8 weeks after the beginning of model making, significantly lower than those in CCl4 model treated with low dose hALR. In the albumin model group the TIMP(1) gene expression level showed no change during albumin sensitization, significantly increased during caudal vein attack, and reached the peak after medel making. The TIMP(1) gene expression level was lower in albumin model treated with low dose hALR during and after caudal vein attack than in the model group and negative control group, however, without significant difference (both P > 0.05). The TIMP(1) gene expression level in albumin model treated with high dose hALR was 2.33 +/- 0.36 and 4.02 +/- 0.53 during and after caudal vein attack respectively, significantly lower than those in CCl4 model group (0.99 +/- 0.14, 2.03 +/- 0.30, 2.99 +/- 0.43, and 4.13 +/- 0.44 respectively 2, 4, 6, and 8 weeks after beginning of model making respectively) and those in albumin model without hALR treatment and that with low dose hALR (4.13 +/- 0.60 and 5.99 +/- 0.83, and 3.70 +/- 0.82 and 5.63 +/- 0.89 during and after caudal vein attack respectively), and negative control group. CONCLUSION: High dose recombinant human augmenter of liver regeneration is effective in inhibiting gene expression of TIMP(1) in experimental liver fibrosis. PMID- 12133483 TI - [The UCSNP44 variation of calpain 10 gene on NIDDM1 locus and its impact on plasma glucose levels in type 2 diabetic patients]. AB - OBJECTIVE: To study the contribution of single nucleotide polymorphism-UCSNP44 at calpain-10 gene (CAPN-10) on NIDDM1 locus to type 2 diabetes mellitus (T2DM) in Chinese. METHODS: 276 Chinese living in Shanghai, 148 with normal glucose tolerance (NGT) and 128 with T2DM were given 75 g glucose. O, 30, 60, 120, and 180 minutes later their plasma glucose (PG), insulin (INS), C-peptide (CP), and free fatty acids (FFA) were measured and the areas under curve (AUC) were calculated. The islet beta-cell insulin secretion and tissue insulin sensitivity were estimated by formulae of homeostasis model assessment and increment ratio of insulin to glucose levels 30 minutes after glucose challenge. The CAPN-10 UCSNP44 as well as UCSNP43 were genotyped by automated DNA direct sequencing. RESULTS: (1) The major genotype of CAPN-10 UCSNP44 in persons with NGT was TT (with a frequency of 0.82); the major allele was T (0.91). The most frequent genotype combination between UCSNP44 and UCSNP43 was TT-GG (corresponding to haplotype combination TG/TG) (0.64). The most frequent haplotype was TG (0.80). The D value for linkage disequilibrium between UCSNP44 and UCSNP43 was -0.11. (2) The frequencies of UCSNP44 and UCSNP44/UCSNP43 haplotype combination did not differ significantly between subjects with NGT and those with T2DM. (3) The PG levels in T2DM subjects with UCSNP44 TT genotype both at fasting and after glucose challenge were statistically significantly higher than those in subjects with non TT (TC + CC) genotype, especially the PG levels 0, 60, 120, and 180 minutes after glucose challenge (P = 0.036, 0.040, 0.020, and 0.017) and the PG-AUC (P = 0.013). The PG levels and PG-AUC 0 and 120 minutes after glucose challenge were still significant after adjusted with age, sex, and body mass index and waist circumference. Similar results were observed in comparison between the TG/TG and TG/CG subgroups of UCSNP44/UCSNP43 haplotype combination. In addition, T2DM subjects with UCSNP44 TT genotype had lower CP levels after glucose challenge than those with non-TT genotype. However, the difference became not statistically significant after adjusted with above-mentioned variables. CONCLUSION: The variation of CAPN-10 UCSNP44 has an impact on plasma glucose levels at fasting and after glucose challenge in subjects with type 2 diabetes. The relevant mechanism remains to be elucidated. PMID- 12133484 TI - [Effect of antisense-tissue inhibitor of metalloproteinase-1 gene transfer on tubulointerstitial lesion in rats with unilateral ureteral obstruction]. AB - OBJECTIVE: To observe the effect of antisense- human tissue inhibitor of metalloproteinase-1 (hTIMP-1) on tubulointerstitial lesion induced by unilateral ureteral obstruction (UUO). METHODS: Forty-two SD rats, divided into 3 groups equally, underwent UUO and received intra-ureteral injection of normal saline, empty retroviral vectors and recombinant retroviral vectors containing antisense- hTIMP-1 respectively, and were sacrificed 3 and 7 days after the operation. The expression of antisense h-TIMP-1 in rat kidney was detected by RT-PCR. Immunohistochemistry was used to detect the expression of rat TIMP-1, alpha-SMA actin (alpha-SMA), and proliferating cell nucleus antigen (PCNA) in tubulointerstitium. The relative area of renal tubulointerstitium was calculated. RESULTS: The expression of antisense-hTIMP-1 mRNA was detected in rat renal tubulointerstitum. The expressions of rat TIMP-1, alpha-SMA and PCNA were lower in the obstructed kidneys of rats treated with antisense-hTIMP-1 than those in the rats in empty vectors group and normal saline group (3 day after: 0.75 +/- 0.07 vs 1.18 +/- 0.12, 1.14 +/- 0.14, P < 0.05; 7 days after: 2.02 +/- 0.16 vs 4.07 +/- 0.57, 4.13 +/- 0.60, P < 0.01). The relative area of renal tubulointerstitium in rats treated with antisense-hTIMP-1 was smaller than those in the empty vetors group and normal saline group (3 days after: 6.0 +/- 0.7 vs 8.2 +/- 1.2, 8.2 +/- 1.3, P < 0.01; 7 days after: 16.2 +/- 1.0 vs 21.2 +/- 2.6, 21.0 +/- 2.4, P < 0.01). More renal tubules were dilated, infiltration of inflammatory cells was seen focally or diffusely, and the relative area of renal tubulointerstitium was smaller in the antisense-hTIMP-1 group than in the empty vectors group and normal saline group. No difference was seen among the degrees of dilation/atrophy of renal tubules and infiltration of inflammatory cells in the three groups. No difference was seen between the relative area of renal tubulointerstitium in empty vectors group and normal saline group. CONCLUSION: Antisense-hTIMP-1 may have beneficial effects on the tubulointerstitial lesion of obstructive nephropathy through inhibiting the expression of TIMP-1 in tubulointerstitum and the proliferation of interstitial and tubular cells. PMID- 12133485 TI - [Focal adhesion kinase and mitogen activated protein kinase are involved in vascular smooth muscle cells migration and proliferation stimulated by fibronectin]. AB - OBJECTIVE: To study the roles of focal adhesion kinase (FAK) and mitogen activated protein kinase (p42/44MAPK) in the process of migration and proliferation of vascular smooth muscle cells (VSMC) stimulated by fibronectin (FN). METHODS: VSMCs were taken from the tunica media of SD rats and cultured. Migration and proliferation of cultured VSMCs were stimulated by different concentrations of FN; FAK, p42/44MAPK and their phosphorylation were detected by immunoprecipitation and Western blot. FAK antisense oligodeoxynucleotide (ODN) was transfected into VSMCs by cationic lipid to investigate its modulatory effects on tyrosine phosphorylation. VSMC migration and proliferation were also measured by modified Boyden Chamber and (3)H-thymidine respectively. RESULTS: FAK and p42/44MAPK were expressed when VSMC adhesion and migration were successfully simulated by FN (5, 10, 20, 40, and 60 microg/ml), high contents of FAK and p42/44MAPK phosphorylation were detected in groups with 20 microgram/ml FN or more. FAK antisense ODN was transfected efficiently by cationic lipid. Phosphorylation of FAK and p42/44MAPK was inhibited significantly after FAK antisense ODN transfection. Cell migration stimulated by FN of the concentrations of 10, 20, 40, and 60 microg/ml was reduced by 23.26%, 21.63%, 19.31% and 17.88% respectively (P < 0.05). VSMC proliferation in 5 approximately 60 microgram/ml FN groups was reduced by 27.67% approximately 46.67% (P < 0.05). CONCLUSION: FAK and p42/44MAPK play an important role in VSMC migration and proliferation stimulated by extracellular matrix. The process can be inhibited by FAK antisense ODN effectively. PMID- 12133486 TI - [A report of 50 patients with artemisia pollenosis and plant food allergy]. AB - OBJECTIVE: To study the patients with both artemisia pollenosis and plant food allergy. METHODS: The diagnosis of artemisia pollenosis was based on a history of summer-autumn pollenosis, and positive intradermal test with artemisia pollen (Ar) and serum specific IgE-Ar; the diagnosis of plant food allergy was based on a history of the symptoms occurred shortly after the intake of some plant foods, and positive skin prick test using some plant food, and positive specific IgE in some of them. RESULTS: A total of 50 patients with both artemisia pollenosis and plant food allergy was diagnosed. The average age of onset of artemisia pollenosis was 24 (7 approximately 56 yr), The average age of onset of allergy to plant food was 23 year 8 month (3 approximately 57 yr). Sex ratio (M:F) was 21:29 with female predominating. artemisia pollenosis occurred prior to food allergy in 50% of the patients, plant food allergy occurred prior to pollenosis in about 29% of the patients, both of them occurred simultaneously in 21%. The common offending food were legumes, peach and peanut and sunflower seeds etc, nasal symptoms by inhalation of Ar occurred in 96% (46/48) of the patients, followed by conjunctivitis and asthma, urticaria was not common, anaphylatic shock and gastrointestinal symptoms were rare. The common clinical manifestations caused by ingestion of plant food were urticaria and oral allergy syndrome (OAS) occurred in 66% (33/50) and 40% (20/50) of the patients respectively, followed by asthma, rhinitis and conjunctivitis, anaphylactic shock and gastrointestinal tract symptoms were much more frequently than artemisia pollenosis. but single oral mucosa symptoms only occurred in 3 patients. CONCLUSION: The patients with pollenosis must be in vigilance for the development of plant food allergy later, and vise versa. Single oral mucosa symptoms were rarely seen. PMID- 12133487 TI - [Development of intraperitoneally transplantated human ovarian carcinoma model with immune reconstruction in severe combined immunodeficient mice]. AB - OBJECTIVE: To develop an intraperitoneally transplanted human ovarian carcinoma model in humanized severe combined immunodeficient (SCID) mice. METHODS: Twelve CB17SCID mice were randomly divided into 4 groups: group A intraperineally injected with phosphate-buffered saline, group B injected with human ovarian carcinoma cells SKOV3, group C injected with human peripheral blood lymphocyte (PBL) for immune reconstruction, and group D injected with PBL and SKOV3 cells. The behaviors of mice, tumor growth and morphology, human IgG in peripheral blood, cancer antigen 125 (CA125) immunohistochemical staining, tumor infiltrating lymphocyte (TIL), and status of graft versus host disease (GVHD) were detected. RESULTS: The survival period was > 28 days in all groups. The ratio of successful tumor transplantation was 3/3 in groups B and D. The tumor was widespread in peritoneal cavity, mainly in diaphragm, liver, and mesentery, with bloody ascites. The number and size of tumor were less in the humanized group. CA125 expression was positive in primary transplanted tumor cells and SKOV3 cells. IgG was detected in humanized groups (C and D). The level of human IgG was significantly higher in group D than in group C (P < 0.05). TIL infiltration was remarkable in group injected with PBL and SKOV3 cells and failed to be found in group injected with SKOV3 cells. No GVHD was found in all groups. CONCLUSION: An intraperitoneal transplanted human ovarian carcinoma model has been established in humanized SCID mice that simulates the biological behavior of human ovarian carcinoma disseminating intraperitoneally in patients with immune function and may function as an ideal animal model for preclinical research of ovarian carcinoma treatment. PMID- 12133488 TI - [Influence of body temperature on outcome of head injuries]. AB - OBJECTIVE: To investigate the influence of body temperature on the outcome of head injuries. METHODS: The data of body temperature and outcomes of patients with head injuries admitted to Tiantan Hospital between July 1997 and July 1999 was analyzed retrospectively. The admission temperature was divided into 4 groups: hypo-temperature (with body temperature < 36 degrees C), normal temperature (36.0 degrees C approximately 37.2 degrees C), mild hyper-temperature (37.3 degrees C approximately 38.3 degrees C) and significant hyper-temperature (> 38.3 degrees C). Appraisal was made at discharge and one year later using GOS criteria (grade 1 approximately 2 were regarded as unfavorable prognosis, and grades 3 approximately 5 as favorable prognosis). Chi-square test and logistic regression analysis were performed using SPSS statistic software. RESULTS: The outcome of the patients with significant high admission temperature and hypo temperature was unfavorable. The difference between different temperature groups, except the hypo-temperature group and significant high temperature group, was significant (P < 0.01). The higher admission temperature means less favorable outcome. The outcome of the hypo-temperature and hyper-temperature groups was less favorable than that of the normal temperature group (P < 0.01). The difference in the 15 factors such as temperature, age, etc, between the group with favorable prognosis and the group with unfavorable prognosis (P < 0.01). However, the difference in hemoglobin and hematocrit between the group with favorable prognosis and the group with unfavorable prognosis was unsignificant. The value of B at discharge and 1 year after injury were -0.809 and -1.104 respectively. CONCLUSION: The admission body temperature can be used as an independent predictor for prognosis of head injuries. The low- and the high temperature predict the worse outcome. The higher the body temperature is, the less favorable the outcome. Admission body temperature has a significant influence on the outcome of head injuries. PMID- 12133489 TI - [Neuropathy in nearby cranial nerves after acoustic schwannoma gamma knife radiosurgery, a follow-up study]. AB - OBJECTIVE: To investigate the risks of facial, trigeminal and acoustic neuropathies after acoustic schwannoma gamma knife radiosurgery. METHODS: The clinical data of forty-three patients with 46 masses of acoustic schwannoma who underwent gamma knife radiosurgery with the dose of 12 approximately 15 Gy to the tumor margin between January 1997 and October 2000 and were followed up for 6 approximately 24 months (on average 16.9 months) were studied. The tumor diameter was 10 approximately 20 mm in 12 cases, 21 approximately 30 mm in 23 cases, >/= 31 mm in 11 cases, with the average value of 28 mm. RESULTS: The general tumor control rate was 91.3%. The useful hearing preservation rate was 100% immediately after radiosurgery, 87% 6 months later and 78% 2 years later. The hearing preservation rate was high for small tumors. The facial and trigeminal neuropathies began to appear after 6 months. The incidence rates of facial neuropathy was 15.3%, 7.6%, and 3.8% 6 months, 1 year and 2 years after radiosurgery respectively. The incidence rates of trigeminal neuropathy was 11.4%, 3.8%, and 3.8% respectively 6 months, 1 year, and 2 years after radiosurgery. The incidence of neuropathy was 3.8% for tumors with a diameter < 30 mm for both facial and trigeminal nerves. The hearing in 2 out of 15 cases with dysaudia began to improve 6 months after radiosurgery. The incidence of neuropathy for tumors with the diameter > 30 mm was 3.8% for both nerves 2 years after raadiosurgery. The preservation rate of useful hearing for tumors with the diameter < 20 mm was 100% after radiosurgery. CONCLUSION: Stereotactic radiosurgery using gamma knife with a dose of 12 approximately 15 Gy to the tumor margin succeeds in controlling acoustic schwannoma and preserving useful hearing. The incidence of facial and trigeminal neuropathies are low. The neuropathy caused by gamma knife radiosurgery is sub-lethal and can be recovered gradually. PMID- 12133490 TI - [Treatment of benign lesions in thyroid with colloid (32)P under ultrasonic guidance: a clinical study]. AB - OBJECTIVE: To evaluate the feasibility and effect of treatment of thyroid adenoma and cystic degeneration of thyroid adenoma by injection of colloid (32)P with ultrasonic guidance. METHODS: Diluted solutions of (32)P, 37 approximately 74 mBq/1.0 approximately 1.5 ml and 18.5 approximately 37 mBq/ml, were injected, with ultrasonic guidance, into the thyroid adenoma in 30 cases and degenerated cyst of thyroid adenoma in 30 cases, all confirmed by ultrasonography, thyroid scanning and pathologic paracentesis. The serum FT3, FT4, TT3, TT4, and TSH, and blood cells were examined before and after the treatment. Part of the patients was followed up for 6 approximately 36 months. RESULTS: The cure rate was 71.05% (27/38) for thyroid adenoma and 86.67% (26/30) for cystic degeneration of thyroid adenoma. The treatment effect was better for cystic degeneration of thyroid adenoma than for thyroid adenoma. The changes of blood cells and serum thyroid hormones were not significant before and after treatment. No obvious side effect was found in all patients. CONCLUSION: Injection of (32)P with ultrasonic guidance is effective in treatment of thyroid adenoma and cystic degeneration of thyroid adenoma. This method is easy, safe and highly practical clinically. PMID- 12133491 TI - [Effect of serum from patients with Graves' disease on colony forming unit granulocyte monocyte]. AB - OBJECTIVE: To investigate the effects of serum component from patients with Graves's disease (GD) on the growth of colony forming unit-granulocyte monocyte (GM-CFU). METHODS: Monocytes were obtained from 11 normal persons and 11 GD patients with leukopenia and culture together with sera from 11 GD patients with leukopenia, 11 GD patients without leukopenia, and 11 normal controls for 10 days. Inverted microscopy was used to count the number of colony. RESULTS: Thyroid stimulating immunoglobulin (TSI) and serum from GD patients with leukopenia significantly inhibited the formation of GM-CFU (P < 0.01) while methimazole, thyroxine and serum from GD patients without leukopenia did not have any effect on the formation of GFU-GM (P > 0.05). CONCLUSION: TSI and serum from GD patients with leukopenia remarkably inhibit GM-CFU growth. Autoimmune abnormality may play an important role in the pathogenesis of leukopenia in patients with GD. PMID- 12133492 TI - [Combination of mycophenolate mofetil with cyclosporine A and methotrexate for the prophylaxes of acute graft versus host disease in allogeneic peripheral stem cell transplantation]. AB - OBJECTIVE: To evaluate the efficacy of combination of mycophenolate mofetil (MMF) with cyclosporine (CsA) and methotrexate (MTX) on prophylaxes of acute graft versus-host disease (GVHD) in HLA-matched allogeneic peripheral blood stem cell transplantation (allo-PBSCT). METHODS: Thirty-nine patients with acute leukemia (n = 21) and chronic myeloid leukemia (n = 17) and severe aplastic anemia (n = 1) were treated with allo-PBSCT from HLA matched siblings (n = 36) or unrelated donors (n = 3). Twenty-six patients were in CsA + MTX group. CsA was given at a dosage of 2 mg.kg(-1).d(-1) by continuous intravenous injection for 24 h, since on day(-1) and injection of CsA was changed to oral administration of CsA around day 18. CsA was tapered by 10% per week after day + 90. MTX was given at the dosage of 15 mg at day + 1, and 10 mg at day + 3, + 6 and + 11, respectively. Thirteen patients were included in MMF + CsA + MTX group with the same dosage of CsA and MTX as above but omitted at day + 11. MMF of 2 g/day was added orally from day + 1 to day + 28 post transplantation. RESULTS: All patients in both groups were successfully engrafted. The days of recovery of neutrophils and platelets were not significantly different between two groups (P > 0.05). The incidence of acute GVHD in MMF + CsA + MTX group (7.6%) was significantly lower than that in CsA/MTX group (46.2%, P < 0.05). Incidence of grade II approximately IV GVHD in MMF group was 0 while that in control group was 23.0%. The incidence of severe mucositis was lower in MMF group (15.4%) than in the control group (30.8%) (P < 0.05). CONCLUSION: The regimen of MMF + CsA + MTX for prevention of acute GVHD in allo-PBSCT is more efficient than that of CsA + MTX, without adversely affecting the engraftment and relapse rate. PMID- 12133493 TI - [Hyperacute graft versus host disease after allo-stem cell transplantation, analysis of 118 cases]. AB - OBJECTIVE: To investigate the clinical manifestations, management, prognosis, and risk factors of hyperacute graft versus host disease (hGVHD) after allo-SCT cell transplantation all-SCT. METHODS: The data of 118 patients aged 2 approximately 52 who underwent 120 times of allo-SCT were analyzed retrospectively. RESULTS: hGVHD was observed in 17 patients with an incidence rate of 14.17%. The clinical manifestations included gastrointestinal symptoms (13/17), rash (13/17), fever (8/17), and hepatic dysfunction (6/17). Another 12 cases with only one organ injured were included in atypical hGVHD group. The two-year survival rates were 30.50%, 40.00%, and 61.67% in patients with typical hGVHD, patients with atypical hGVHD, and patients without hGVHD respectively. The condition of twenty-five cases with hGVHD improved after treatment with immunosuppressive drugs. CONCLUSION: hGVHD is a common complication after allo-SCT that affects the prognosis dramatically. The risk factors of hGVHD include HLA mismatching, disease being in progressive stage just before allo-SCT, female in sex, and sex mismatching. PMID- 12133495 TI - [Detection of level and mutation of neurofilament mRNA in Alzheimer's disease]. AB - OBJECTIVE: To study if the increase of levels of neurofilament (NF)-M and NF-L in brain of Alzheimer's diseases (AD) is caused by increase of NF-M and NF-L. METHODS: Semi-quantitative reverse transcription PCR was used to measure the NF-M and NF-L mRNA levels in the brain gray matter obtained by autopsy from 8 patients with AD and 7 age-matched patients with Huntington's disease (HD) used as controls. The mutation of NF-M and NF-L was detected by single strand DNA conformation polymorphism technique. RESULTS: The levels of NF-M mRNA and NF-L mRNA were significantly lower in the brains of AD patients (0.3 +/- 0.03 * OD(NF M)/OD(GAPDH) or OD(NF-L)/OD(GAPDH) and 0.44 +/- 0.16 * OD(NF-M)/OD(GAPDH) or OD(NF-L)/OD(GAPDH) respectively) than in the brains of HD patients (0.42 +/- 0.07 OD(NF-M)/OD(GAPDH) or OD(NF-L)/OD(GAPDH) and 0.79 +/- 0.09 OD(NF-M)/OD(GAPDH) or OD(NF-L)/OD(GAPDH) respectively) (P < 0.005 and P < 0.007 respectively). NF-M mutation was detected in one AD patient. No NF-L mutation was found in the samples examined. CONCLUSION: The increase of neurofilament protein observed in the brains of AD patients is related neither to increased gene transcription nor to mutation of NF mRNA. PMID- 12133494 TI - [Association between Bcl-2 gene polymorphism with systemic lupus erythematosus]. AB - OBJECTIVE: To study the association between bcl-2 gene polymorphism and systemic lupus erythematosus (SLE) in Chinese and detect the synergism between IL-10 and bcl-2 genotypes in determining susceptibility to SLE. METHODS: The peripheral white blood cells from 232 nuclear families of SLE and 106 ethnically matched normal controls were collected and the DNA extracted. The allelic distribution of one microsatellite located in Bcl-2 gene was determined by using fluorescent labeled primer and genotyping method. RESULTS: (1) The distribution frequency of Bcl-2 alleles and its major genotypes were significantly different between SLE group and normal controls (P = 0.001 and 0.0029 respectively). (2) Transmission disequilibrium existed in Bcl-2 alleles. Bcl-2-195 bp allele was preferentially transmitted to affected offspring (t: non-t = 122:82, P = 0.0051) by TDT approach. Individuals carrying specific genotypes with both Bcl-2-195 bp allele and IL-10G138 bp allele were at higher risk of developing SLE than those carrying only Bcl-2-195 bp allele or IL-10G138 bp allele (OR = 3.00, 1.77, 1.07 respectively). CONCLUSION: Bcl-2 gene polymorphism is associated with SLE and may be directly involved in the process of SLE or in linkage disequilibrium with some susceptibility loci nearby. Synergistic effect may exist between IL-10 and bcl-2 genotypes in determining susceptibility to SLE. PMID- 12133496 TI - [Clinical features of pheochromocytoma and anesthetic management during perioperative period]. AB - OBJECTIVE: To investigate the clinical features of pheochromocytoma and summarize the experience of anesthetic management during perioperative period. METHODS: Two hundred and fifty eight medical records of patients who were diagnosed as pheochromocytoma in Peking Union Medical College Hospital were reviewed retrospectively for clinical features, anesthetic management and perioperative mortality. RESULTS: About 5.8% (15/258) of pheochromocytomas was an integral part of multiple endocrine neoplasia (MEN) type II or mixed type. Sixty percent (149/249) of the patients undergoing surgery possessed evidence of catecholamine cardiac toxicity preoperatively, including abnormal ECG, myocardial hypertrophy and decreased left ventricular ejective fraction. Impaired glucose tolerance was found in 59% (147/249) of patients before surgery. The volume infused during operation was significantly higher both in the epidural anesthesia group (3 474 ml +/- 624 ml, q(1) = 5.72, P < 0.01) and in the epidural plus general anesthesia group (3 654 ml +/- 475 ml, q(2) = 5.83, P < 0.01) than that in the general anesthesia group (2 534 ml +/- 512 ml). There were favorable hemodynamic characteristics before removal of the tumor in the epidural anesthesia group and epidural plus general anesthesia group, as compared with in the general anesthesia group. Perioperative mortality was significantly decreased from 8% (5/60) in period 1 (from 1955 to 1975) to 1.2% (1/75) in period 2 (from 1976 to 1994) (chi(2) = 4.05, P < 0.01). No perioperative death (0/111) occurred in period 3 (from 1995 to 2001). CONCLUSION: A good surgical outcome for the excision of pheochromocytoma depends on multiple factors, including careful assessment of potential end organ damages and restoration of blood volume by establishing alpha-blockade during the preoperative period, meticulous anesthetic management during surgery, and appropriate circulatory support after surgery. PMID- 12133497 TI - [Detection of serum epidermal growth factor receptor in the diagnosis of proliferation of pituitary adenomas]. AB - OBJECTIVE: To investigate the correlation between preoperative serum epidermal growth factor receptor (EGF-R) level and proliferation of pituitary adenoma. METHODS: ELISA was used to determine the levels of EGF-R in preoperative serum from 137 patients with pituitary diseases: 5 cases of pituitary Rathke's pouch, 13 cases of pituitary hyperplasia, and 119 cases of pituitary adenomas including 17 cases of microadenoma, 64 cases of macroadenoma, and 38 cases of giant adenoma. Twenty-eight healthy blood donors were used as controls. RESULTS: The level of serum EGF-R was (194 +/- 38) fmol/L in patients with pituitary hyperplasia, and was (219 +/- 37) fmol/L in patients with pituitary microadenoma, 322 +/- 66 fmol/L in patients with pituitary macroadenoma and (428 +/- 62) fmol/L in patients with pituitary giant adenoma. The levels of serum EGF-R in patients with different kinds of pituitary hyperplasia and pituitary adenoma were significantly higher than the level in patients with pituitary Rathke's pouch (152 +/- 17) fmol/L, all P < 0.0001, and were significantly higher than that in healthy control group (159 +/- 41) fmol/L, all P < 0.05. The serum level of EGF-R was positively correlated to the size of pituitary adenoma (r = 0.998). Furthermore, significant difference in the level of serum EGF-R was observed among patients with different kinds of pituitary adenoma (P < 0.0001). CONCLUSION: EGF-R level in preoperative serum of patients with pituitary adenoma reflexes the proliferative status of pituitary adenoma and helps differentiate pituitary adenoma from pituitary Rathke's pouch. It is suggested that serum EGF-R be used as a referable marker of proliferative status of pituitary adenoma. PMID- 12133498 TI - [Localized Castleman disease with paraneoplastic pemphigus and pulmonary involvement: clinical features and histopathology]. AB - OBJECTIVE: To investigate the clinical, histopathologic and CT features of localized Castleman disease complicated with paraneoplastic pemphigus (PNP) and pulmonary involvement. METHODS: The clinical, laboratory, histopathologic records and data of computed tomographic (CT) images of 4 patients of Castleman disease with PNP were reviewed. RESULTS: All of the four patients presented with PNP which was confirmed by pathological and immunological studies and also developed lung abnormalities, including bronchiolitis obliterans (n = 3), lung abscess (n = 1) and multiple lung infarction (n = 1), 2 to 5 months after the onset of PNP. Castleman's tumor, which was found incidentally during routine examination, manifested as a solitary and huge mass (5 to 14 cm in size) which markedly enhanced homogeneously on CT in the retroperitoneum (n = 3) and mediastinum (n = 1). Histologically, three cases were of hyaline-vascular type and the other one was of mixed type. Complete remission of PNP occurred 2 weeks to 8 months after surgical resection of the tumor. The condition of two patients with bronchiolitis obliterans failed to be improved. They had been presenting dyspnea and irritable cough 3 to 15 months after surgery. CONCLUSION: PNP and lung abnormalities are rare and severe complications of localized Castleman's disease. Early diagnosis and surgery are very important for prognosis. PMID- 12133499 TI - [Effect of transcranial magnetic stimulation on rehabilitation of motor function in patients with cerebral infarction]. AB - OBJECTIVE: To study the effect of transcranial magnetic stimulation (TMS) on the rehabilitation of motor function in patients with cerebral infarction. METHODS: Two hundred twenty and three patients with the initial attack of cerebral infarction were divided into three groups. (1) One hundred twenty and three patients were randomly further divided into rehabilitation subgroup (n = 63, aged 64.0 +/- 7.9, treated with TMS beginning from the 6 approximately 10 th day after onset) and control subgroup (n = 60, aged 63.5 +/- 7.9). (2) Sixty patients were divided into three subgroups according to the age: 35 approximately 55 (n = 11), 56 approximately 75 (n = 20), and over 76 (n = 30). (3) Forty patients aged 56 approximately 75 were further divided into two subgroups according to the timing of beginning of TMS: subgroup with TMS beginning within 3 months after the initial attack (n = 20) and subgroup with TMS beginning 3 months after the initial attack (n = 20). Except for TMS, the basic treatment was the same for all of the patients. TMS was given once a day for 14 days. Fugl-Meyer score, Barthel index, and central spinal cord motor conduction time (CMCT) were measured twice: just before treatment and after the 14th treatment. A difference less than 20 ms between CMCT values measured before and after TMS and failure to induce CMCT both before and after TMS were regarded as ineffective hess, and a difference more than 20 ms between CMCT values measured before and after TMS and a change from failure to induce CMCT were regarded as effective hess. RESULTS: The effective rate was 90.5% in the rehabilitation subgroup and 68.3% in the control group (P < 0.05). The Fugl-Meyer scores were 36.3 +/- 15.8 and 33.7 +/- 13.7 in the rehabilitation subgroup and control subgroup before treatment respectively (P > 0.05), and were 51.7 +/- 15.7 and 40.0 +/- 13.9 after treatment respectively (P < 0.01). The Barthel index were 39.6 +/- 15.8 and 40.0 +/- 14.9 in the two subgroups before treatment respectively (P > 0.05) and were 50.5 +/- 15.7 and 43.9 +/- 15.3 after treatment respectively (P < 0.05). The Fugl-Meyer scores before and after TMS were 35.8 +/- 11.3 and 61.6 +/- 18.6 in the subgroup aged 35 approximately 55 (P < 0.01), 33.5 +/- 14.0 and 49.2 +/- 16.0 in the subgroup aged 56 approximately 75 (P < 0.001), and 32.1 +/- 10.7 and 37.6 +/- 11.3 in the subgroup aged over 75 (P > 0.05). The Fugl-Meyer scores were 41.2 +/- 15.7 and 59.0 +/- 22.8 before and after TCM respectively in the subgroup with early TMS treatment (P < 0.01), and were 34.3 +/- 13.5 and 45.4 +/- 14.8 respectively in the subgroup with TMS beginning 3 months after the onset (P < 0.05). CONCLUSION: TMS is effective on the rehabilitation of motor function in patients with cerebral infarction. The effectiveness of TMS treatment depends on the age of the patients and timing of beginning treatment. PMID- 12133500 TI - [Anti-HBV efficacy of bifendate in treatment of chronic hepatitis B, a primary study]. AB - OBJECTIVE: To investigate the anti-HBV efficacy of bifendate in treatment of chronic hepatitis B. METHODS: A total of 119 patients with chronic hepatitis B were randomly divided into treatment group (n = 65, aged 24 +/- 12) and control group (n = 54, aged 25 +/- 11). In the treatment group every patient was given higher doses bifendate pills ( 12 age, 45 approximately 67.5 mg/d) for up to 12 months. Hepatic function test was performed and HBeAg, HBeAb and HBV DNA were detected at regular intervals in all patients. RESULTS: The serum alanine amonotransferase (ALT) decreased to normal only one month later in 70.76% of patients in the treatment group and decreased to normal at least in 2 approximately 3 months in the control group (P < 0.01). The serum conversion rates of HBeAg, HBeAb and HBV DNA in the treatment group were 44.4%, 29.3%, 38.5%, respectively, which were significantly higher than those in the control group (P < 0.01). No noticeable side effect was observed. CONCLUSION: Higher doses of bifendate taken for a long term has remarkable anti-HBV efficacy in treatment of chronic hepatitis B. PMID- 12133501 TI - [CT manifestations of primary hepatic sarcoma]. AB - OBJECTIVE: To assess the diagnostic value of CT to primary hepatic sarcoma. METHODS: The CT findings of 12 cases of primary hepatic sarcomas confirmed by operation and pathology, including leimyosarcoma, liposarcoma, and malignant fibrous histocytoma, 2 cases each, and angiosarcoma, carcinosarcoma, cystadenocarcinosarcoma, mesotheliosarcoma, fibrosarcoma, and malignant lymphoma, one case each, were analysed retrospectively. RESULTS: Only 2 out of the 12 cases of primary hepatic sarcoma were correctly diagnosed by CT before operation. The CT manifestations of primary hepatic sarcoma, against the operational and pathological findings, could be summarized as two types: solid mass type and cystic mass type. Eight cases were of solid mass type, including 2 cases of liposarcoma, and leiomyosarcoma, angiosarcoma, carcinosarcoma, methotheliosarcoma, fibrosarcoma, and malignant lymphoma, one case each. Plain scanning showed homogeneous or inhomogeneous low-density masses; enhanced scanning showed no marked enhancement or inhomogeneous enhancement, ringed peripheral enhancement, or nodular peripheral enhancement. Peripheral enhancement was a CT feature common to angiosarcoma and lymphoma. The characteristic CT manifestation of liposarcoma was an inhomogeneous mass with well-defined border and multiple intermingled septa. Four cases were of cystic mass type, including 2 cases of malignant fibrous histocytoma, and leimyosarcoma with post-operative recurrence and cystadenocarcinosarcoma, one case each, all manifesting a huge single cystic mass: unilocular in the case of leimyosarcoma with post-operative recurrence and multilocular in the other 3 cases. The wall and/or septa of the cyst had an uneven thickness and were markedly enhanced after contrast administration. Clear solid element and wall nodules were seen in the cystic mass of cystadenocarcinosarcoma. CONCLUSION: The CT findings of primary hepatic sarcoma are associated with the pathology of tumor. Some pathological patterns of primary hepatic sarcoma, such as liposarcoma and angiosarcoma, have their own characteristic CT manifestations. However, the diagnosis of most primary hepatic sarcoma lacking characteristic CT manifestations must be based on clinical and laboratory examinations and confirmed by pathology. PMID- 12133502 TI - [Intravitreal injection of doxorobicin and dexamethason in treatment of proliferative vitreoretinopathy]. AB - OBJECTIVES: To investigate the inhibitory effect of intravitreal injection of doxorubicin and dexamethason on proliferative vitreoretinopathy (PVR). METHODS: Forty-two eyes with retinal detachment complicated with proliferative vitreoretinopathy underwent surgery and intravitreal injection of doxorubicin and dexamethason and followed up for 3 approximately 5 months. The curative effect was compared with that of pure surgery on 30 eyes with retinal detachment complicated with PVR. Relevant hemodynamic changes of central retinal artery (CRA) and central retinal vein (CRV) were also evaluated. RESULTS: Retinal reattachment was found in most eyes in both groups postoperatively. The worsening rate of epiretinal membrane and subretinal membrane was 7.1% and 26.7% in the surgery plus injection group and pure surgery group respectively (P < 0.05). The maximum average systole flow rate of CRA and CRV were 6.3 cm/s +/- 2.5 cm/s and 4.8 cm/s +/- 1.4 cm/s, 33.7% and 16.7% than normal value respectively. CONCLUSION: Intravitreal injection of doxorubicin and dexamethason is effective against proliferative vitreoretinopathy. PMID- 12133503 TI - [Effect of hexamethylene bisacetamine on proliferation and apoptosis of human leukemia cells in vitro and relevant mechanisms]. AB - OBJECTIVES: To investigate the in vitro effect of hexamethylene bisacetamine (HMBA) on the proliferation, cell cycle regulation, and apoptosis of human leukemia cells and to study the relevant mechanisms. METHODS: K562 and U937 human leukemia cells were cultured and then suspensions thereof without HMBA or with HMBA of different concentrations (1, 2, and 4 mmol/L) were made. The optical absorbance value at 595 nm of these suspensions were tested every 24 hours. The suspensions were mixed with 4% trypan blue dye solution. Live and dead cells were observed with microscope and the live cell rate was calculated. Double staining of cellular cyclin A protein/DNA was used to detect the expression levels of cyclin A protein in cytoplasm. Fluorescein isothiocyanate (FITC)-labeled annexin V/Propidiumiodide) was used to detect the apototic cells fluorescence-activated cell sorter. Gel electrophoresis was conducted to detect DNA contents. RESULT: After exposure to HMBA for 6 days, the optical absorbance values at 595nm of K562 leukemia cells of the control group and groups with 1, 2, and 4mmol/L HMBA were 1.03, 0.81, 0.58, and 0.36 respectively. After treatment of HMBA, the percentages of cyclin A positive K562 leukemia cells in control group and groups with 1, 2, and 4 mmol/L HMBA were 34.4% +/- 5.2%, 16.1% +/- 2.5%, 9.9% +/- 1.7%, and 7.6% +/ 2.0% respectively (P < 0.01). No drug-related apoptosis was found in both cell lines. CONCLUSION: HMBA inhibits cell proliferation and reduces S-phase-fraction in both cell lines dose-dependently through down-regulating the expression of cyclin A. Apoptosis is not a consequence of the proceeding mitosis arrest and is induced p53-dependently by HMBA. In both leukemia cell lines (U937 and K562) lacking wt-p53, a nonapoptotic death pathway may exist during HMBA induction. PMID- 12133504 TI - [CpG-ODN is a potential candidate adjuvant for human vaccines]. AB - OBJECTIVE: To evaluate the adjuvanticity of CpG-ODN for human vaccines in animal models. METHODS: To find suitable animal models, the human CpG-ODN were examined for their in vitro immunostimulatory activities for murine and Rhesus monkey immune cells. Then by using recombinant HBsAg as a model antigen, the adjuvanticity of human CpG-ODN was evaluated in the animal models. RESULTS: Rhesus monkey B cells responded well to all the human CpG-ODN, similarly as that of human B cells. In contrast, only the human CpG-ODN with the CpG motif 5'GTCGTT 3' (CpG2006 etc) could induce murine splenocytes to secret IgM and IFN-gamma, while those with the CpG motif 5'GTCGTC 3' (CpGT7 etc) had less or no effects. The results suggested that Rhesus monkeys and mice could be used as animal models to evaluate the in vivo activities of different human CpG-ODN. Immunized with HBsAg combined with various human CpG-ODN, the mice elicited a stronger Th1 humoral immunity. Consistent with the in vitro findings, CpG-ODN with the CpG motif 5'GTCGTT 3' were more potent than those with the CpG motif 5'GTCGTC 3'. But of note, all the sequences had the same ability for modulation of Th1/Th2 immune response, with the ratio of IgG2a/IgG1 around 1. However, human CpG-ODN had less adjuvanticity for HBsAg in Rhesus monkeys; only CpGT7 increased the antibody titers by 2 times, while CpG2006 had no effect. CONCLUSION: The preliminary results derived from animal models showed that CpG-ODN was a potential candidate Th1 adjuvant for human vaccines. PMID- 12133505 TI - [Development of gene-viral vector and its anti-tumor effect, a primary study]. AB - OBJECTIVE: To develop a new kind of vector system, named as gene-viral vector, which combines the advantages of the gene therapy and virus therapy. METHOD: An anti-tumor gene was inserted into the genome of the replicative virus specific for the tumor cells by virus recombination technology. The killing effect, report gene expression of the green fluorescence protein, expression of the anti-tumor gene of mouse IL12, and the replication of the virus were observed respectively by cell pathology, fluorescence microscopy, ELISA and electron microscopy. RESULTS: A new kind of gene-viral vector system, in which the E1b-55 000 gene is deleted but the E1a gene of adenovirus is preserved, was constructed. The vector system possessed the same characteristics as the replicative virus ONYX-015, replication and proliferation in the tumor cells but not in the normal cells, thus specifically killing the tumor cells. Besides, it carried many kinds of anti tumor genes. When carrying the report gene of the green fluorescence protein it made the expression of this gene in tumor cells far more effectively than the adenovirus vector employed in the traditional gene therapy did. However in the normal cells the expression of green fluorescence protein caused by this vector system was as little as or even less than that by the traditional adenovirus system. The similar result was also observed in the experiments of this vector system carrying the anti-tumor gene, gene of mouse IL12. The replication and proliferation of the virus carrying the gene of mouse IL12 in the tumor cells were confirmed by electron microscopy. CONCLUSION: Gene-viral vector is a new kind of vector in which the anti-tumor gene is inserted into the genome of the replicative virus specific for the tumor cells. It increases the expression of the anti-tumor gene by hundreds even tens of thousand times. It posseses all the advantages of gene therapy and virus therapy, thus further enhancing the curative effect and it overcomes such disadvantages as low transfer rate, low expression, lack of target tropism and low anti-tumor activity. It will become one of the most promising means in tumor treatment. PMID- 12133506 TI - [The effect of MyoD family proteins on muscular atrophy induced by brachial plexus injury in rats]. AB - OBJECTIVE: To study the changes of MyoD/myogenin expression in atrophic muscle after injury of brachial plexus among rats and to explore the role of proteins of MyoD family in denervation-induced muscular atrophy. METHODS: The brachial plexus was injured uniliterally among 24 SD rats. Then the rats were randomly divided into four groups treated by perfusion into the stomach with FGb 761 (extract of ginkgo leaf), creatine, clenbuterol, or distilled water (control group) at the dosage of 100 mg(-1).kg(-1).d(-1) for 8 weeks respectively. The changes of MyoD protein and myogenin were measured by Western blotting. The gene expression of MyoD and myogenin were determined by RT/PCR and Northern blotting. RESULTS: The expression of myogenin was upregulated within 24 hours after denervation then decreased in five days. On the seventh day, the expression of myogenin in the experimental side was weaker than in the control side. The expression of MyoD protein was downregulated soon after denervation. The expression of myogenin mRNA and MyoD was significantly downregulated after denervation. Clenbuterol, Egb, and creatine upregulated the expression of MyoD protein to a certain degree, creatine being the most effective and Egb being the least effective relatively. Clenbuterol and Egb upregulated the expression of myogenin in denervated muscle, clenbuterol being more effective. Creatine did not upregulate the expression of myogenin in denervated muscle remarkably. CONCLUSION: After denervation, the expression of MyoD protein and myogenin decreases, which plays an important role in atrophy of denervated muscle. EGb, creatine and clenbuterol retard the denervation-induced muscular atrophy by upregulating the expression of MyoD and/or myogenin. PMID- 12133507 TI - [Construction, packaging and titration of recombinant adeno-associated virus vectors contaning antitumor genes]. AB - OBJECTIVE: To construct, package, and titrate vectors that express antitumor genes using adeno-associated virus (AAV) containing human alpha-fetoprotein (AFP) promoter. METHODS: Recombinant AAV vectors were constructed by blunted ligation of plasmids rAAV-AFP-GFP and pTR-UF5 and p16 cDNA and p21 cDNA. Packaging cell lines, 293 and C12 cells, were transfected with rAAV vectors, plasmid PspRC72, and adenovirus cosmid pAdc with the help of LipofectAMINE and then infected by wild adenovirus 48 hours later. The total viral particle titre was measured by PCR. Electron microscopy was used to observe the morphology of rAAV. C12 cells were infected with rAAV and wild adenovirus (MOI 10) and the titre of infectious virus was measured. by RCA method. Hepatocellular carcinoma cells were cultured and infected with rAAV containing green fluorescent protein (GFP), the infectious rate was observed 48 hours later. RESULTS: Recombinant vectors, rAAV-AFP-p16, rAAV-AFP-p21, rAAV-CMV-p16, and rAAV-CMV-p21, were constructed and verified by DNA sequencing and enzyme digestion. The total viral titre was 10(13) to 10(14)/ml. The titre of infectious rAAV containing GFP was 5 x 10(10)/ml. Electron microscopy showed that the rAVV was round and plaqued with a diameter between 20 and 30 nm. When the MOI was 100, the infectious rate of hepatocellular carcinoma cells was at least 70%. Four rAAV vectors carrying p16 or p21 genes have been constructed. rAAV packaging and titration methods were developed to produce highly-purified AAV stocks. These approaches help in research of gene therapy in liver cancer. PMID- 12133508 TI - [Effect of urokinase on morphological changes of vein wall after acute thrombosis in rabbits]. AB - OBJECTIVE: To assess the effect of urokinase on morphological changes of vein wall after acute thrombosis in rabbits. METHODS: Forty-three rabbits were randomly divided into 3 groups: thrombolysis group (n = 20), non-thrombolysis group (n = 20) and control group (n = 3). The thrombosed femoral veins of four rabbits in thrombolysis group anf four rabbits in non-thrombolysis group were taken 1, 4, 7, 14, and 21 days after thrombosis. The expression areas of smooth muscle actin and collagen fiber were measured by histochemistry. Scanning electron microscopy was used to observe the damage of endothelial cells. RESULTS: The expression area of smooth muscle actin began to increase significantly since day 4 after thrombosis reached the peak value on day 7 in thrombolysis group and non-thrombolysis group; and then decreased in thrombolysis group but remained at a high level in non-thrombolysis group. The expression area of smooth muscle actin in thrombolysis group was always less that in non-thrombosis group (all P < 0.01). The expression area of collagen fiber began to remarkably increase since day 7 after thrombosis in both thrombolysis and non-thrombolysis groups, however, being less in the thrombolysis group, and then decreased in thrombolysis group and remained at a high level in non-thrombosis group. Different cellular damages were observed by SEM one day after thrombosis in both thrombosis groups. However, the injury cells began to be repaired since day 4 in thrombolysis group. CONCLUSION: Thrombolysis with urokinase effectively maintains the integrity of endothelium of vein wall and decreases the fibrocellular proliferation of the thrombosed vein, however, fails to stop the pathological changes caused by thrombosis. PMID- 12133509 TI - [A clinical report of 12 case-times of living related liver transplantation]. AB - OBJECTIVE: To evaluate and summarize the clinical application of living related liver transplantation (LRLT). METHODS: A retrospective analysis was made in altogether 12 LRLT operations of living related liver transplantation performed in our department. The indication and timing, surgical complications, and nonsurgical issues including infection, rejection, advantages of LRLT in our series were reviewed. RESULT: All the 11 donors are uneventfully after operation; the first receptor was dead, and 10 receptors with Wilson's disease achieve long term survival. The postoperative survival time till now is 58 w, 51 w, 48 w, 38 w, 37 w, 36 w, 22 w, 18 w, 10 w and 7 w, respectively. No rejection was detected in the recipients with Wilson's disease, whose liver function and cuprum oxidase level had returned to normal. In this 10 cases, 6 of them has returned or gone to school, 2 of them were discharged from hospital, 2 of them has been recovered smoothly in the hospital due to short time postoperatively. The primary complications after operation including blood vessel, biliary tract, lung and the infection of microbe or virus, were mainly involved in our series. CONCLUSION: The process of operation was complicated; the operation technology was exigent and difficult with respect to the safety of the donors and receptors. LRLT has capacious clinical application for there are many unsurpassable advantages, such as plentiful resource and fine quality of graft liver, lower cost of LRLT. PMID- 12133510 TI - [Intracerebral transplantation of nerve cells and genetically modified cells for disorders of central nervous system, a basic and clinical study]. AB - OBJECTIVE: To study the survival ability of cerebellar cells and genetically modified cells transplanted in the brains of mice and monkeys and whether the transplanted neurons correct and improve the cerebral function disorders. METHODS: Suspension of cerebellar tissue from 13-day-old C57BL/6 mouse embryos was transplanted into the cerebellum of 3-month-old BALB/c mice with Purkinji cell degeneration. Since one week after the transplantation, six mice were killed every week till the 8(th) week to examine the cerebellar tissue by immunohistochemistry. Genetically modified neuroblastoma cells with tyrosine hydroxylase (TH) gene and substantia nigra cells of naturally aborted 12-week-old human embryo were implanted into the caudatum nucleus of two monkeys with Parkinson's disease caused by injection of 1-methy-4 pheny-1, 2, 3, 6 tetrahydropyridine. After the operation, the monkeys' symptoms were observed. Six months later, the monkeys were killed. The transplanted areas were examined histologically and immunohistochemically. On the basis of the animal experiment, cerebellar tissue suspension of 9 to 12- week-old human embryos was transplanted into the cerebellum of 6 patients with severe cerebral atrophy, and suspension of substantia nigra cells and adrenal medullary cells from naturally aborted human embryos was transplanted into the caudatum nucleus of 5 patients with late Parkinson's disease. Then the patients were followed up to observe the symptoms. RESULTS: Surviving transplanted embryonic cerebellar cells could be seen in the cerebelli of recipient mice. Differentiation, maturation, and migration into the granular layer of Purkinje cells could be seen. Axons and dendrites grew from the newly generated Purkinje cells. Many mature nerve cells were monoclone antibody staining positive. The myodynamia of monkeys increased and tremor was alleviated, and torsion spasm was remarkably remitted 5 and 7 days after the transplantation respectively. Genetically modified cells with tyrosine hydroxylase gene could be seen and were pale brown with immunohistochemical staining in the cerebral microcapsule of monkeys. Five of the 6 patients with severe cerebellar atrophy and 5 patients with late Parkinson's disease showed improvement of symptoms beginning from 2 weeks after transplantation. The average score of Webster's scale in the patients with Parkinson's disease decreased from 21 to 11. Th3 condition of two patients with cerebellar atrophy and 2 patients with Parkinson's disease continued to improve or remained stable after two years' follow-up. The condition of other patients had worsened. CONCLUSION: The immature embryonic nerve cells and transgenic TH-cells survive in the brain of recipients and have the potential to regenerate neurons and improve the function of central nervous system. PMID- 12133511 TI - [Lamivudine as prophylaxis against hepatitis B virus reinfection following orthotopic liver transplantation]. AB - OBJECTIVE: To discuss the effect of lamivudine as prophylaxis against hepatitis B virus (HBV) reinfection following orthotopic liver transplantation (OLT). METHODS: 40 patients with HBV-related diseases received lamivudine, 100 mg/day, as prophylaxis against HBV reinfection after OLT. Hepatitis B serum markers, HBsAg, HBeAg, HBcAb-IgM, and HBcAg were detected every 2 weeks by immunohistochemistry. Serum HBV-DNA was examined by PCR every 2 weeks. Liver biopsy was conducted to the donor livers during operation and to the transplanted livers postoperatively in regular interval. HBsAg and HBcAg in the liver specimens were examined by immunohistochemistry. YMDD mutation sequencing was done to those patients with conversion of positive HBV DNA postoperatively. RESULTS: The positive rates were 100%, 25%, and 30% for the serum HBsAg, HBeAg, and HBV DNA of the patients respectively before OLT. All 40 donor liver specimens showed negative HBsAg and HBcAg. HBV reinfection rates were 12.2% and 23.5% 6 months and 12 months after OLT respectively. HBeAg had been positive in 4 out of the 6 cases with postoperative reinfection and had been positive in 8 out of the 34 cases without reinfection (P = 0.4505). HBV DNA had been positive in 5 out of the 6 cases with postoperative reinfection, and had been positive in 7 out of the 34 cases without reinfection (P = 0.0461). After OLT HBV DNA converted to positive in 4 cases, mutation of YMDD locus (YMDD-YIDD) had been found in 3 of which. CONCLUSION: Lamivudine monotherapy is effective against HBV reinfection after OLT. Given the lamivudine monotherapy, HBV reinfection causes mutation in YMDD locus on the HBV DNA polymerase gene. OLT does not apply in patients with positive HBV DNA. For those patients to be operated upon, lamivudine should be used to convert HBV DNA to negative beforehand. PMID- 12133512 TI - [Epidemiological survey on epilepsy among rural populations in five provinces in China]. AB - OBJECTIVE: To investigate the prevalence of epilepsy and its treatment gap in rural areas of China. METHODS: A door-to-door epidemiological survey on epilepsy was conducted among 55616 rural residents in 5 counties, each in a province (Heilongjiang, Ningxia, Shanxi, Henan, and Jiangsu) selected by random cluster sampling. All people diagnosed or suspected as epileptic during the screening phase were rechecked by neurologists. RESULTS: A definite diagnosis of epilepsy was made among 387 people with a prevalence rate 7.0 per thousand (age-adjusted rate 6.8 per thousand). The prevalence of active epilepsy was 4.6 per thousand, and the incidence of epilepsy was 28.8/100 000. Of the people with epilepsy, 40.6% had not been treated, 35.4% were treated irregularly. The treatment gap of active epilepsy was 62.6%. CONCLUSION: The prevalence of epilepsy in the rural areas in China was much higher than it was reported previously. About two thirds of the patients with epilepsy has not received regular treatment. A rational intervention strategy should be developed as soon as possible. PMID- 12133513 TI - [Clinical features and treatment of giant cell arteritis in Chinese, a prospective study]. AB - OBJECTIVE: To investigate the clinical features of giant cell (temporal) arteritis (GCA) in China. METHODS: The clinical manifestations, temporal artery biopsy, response to steroid therapy, and follow-up data of sixteen patients with the diagnosis of GCA from July 1999 to March 2001 were analyzed. The American College of Rheumatology (ACR) criteria for classification of GCA were used as reference. RESULTS: Twenty-one patients who sought medical advice in the Second Hospital Affiliated to Xiangya Medical College were suspected of GCA. A definite diagnosis of GCA was made among sixteen patients. The diagnosis among 13 of them fulfilled the 1990 American College of Rheumatology criteria for the classification of GCA. The mean age at disease onset was 43.13 years (range 28 approximately 60 years) and 81.25% of the patients were under the age of 50 when they came down with the disease. The ratio between male and female cases was 15:1. The commonest initial clinical manifestations included newly occurring headache, temporal artery abnormality, visual symptoms, fever, and raised erythrocyte sedimentation rate. Jaw claudication, fatigue, syncope, and hemiparesis could be found in some patients. All the 16 patients underwent temporal artery biopsy. Light and electron microscopy showed inflammatory cell infiltration in arterial wall in 11 cases, fragmented internal elastica in 16 cases, fibrinoid necrosis in 3 cases, smooth muscle cell changes in 10 cases, and thrombosis in the lumen in 5 cases. The mean time from symptom onset to suspicion of GCA or biopsy was 5.52 months (range 0.25 approximately 24.33 months). The misdiagnosis rate during first visit was 87.50%. CONCLUSION: GCA may not be a rare disorder in China. In comparison with the cases abroad, the Chinese GCA patients come down with disease at the earlier age, and most Chinese GCA patients are male. This disease is not understood by many clinicians in China. Misdiagnosis is common. PMID- 12133514 TI - [Transesophageal intraluminal ultrasonography in diagnosis and differential diagnosis of esophageal leiomyoma]. AB - OBJECTIVE: To evaluate the diagnostic and differential diagnostic values of transesophageal echo probe (TEEP) or endoscopic ultrasonography (EUS) in the diagnosis and differential diagnosis of esophageal leiomyoma. METHODS: Between September 1996 and August 2000, TEEP or EUS was used among 23 patients with esophageal submucosal tumor and 11 patients with esophagus constricted from outside, among which 32 cases underwent CT too. All the patients except and 3 with constriction by aorta underwent surgical treatment. RESULTS: (1) The accuracy rate of diagnosis of esophageal leiomyoma by TEEP or EUS was 95.7% (22/23) in comparison with that by CT of only 42.9% (9/21, chi(2) = 14.69, P = 0.0196). (2) In the 11 patients with esophagi constricted by masses beyond or around esophagus, TEEP or EUS definitely differentiated the damage beyond or around esophagus from esophageal leiomyoma. CONCLUSION: Transesophageal intraluminal ultrasonography (TEEP and EUS) is a reliable method for the preoperative diagnosis and differential diagnosis of esophageal leiomyoma and is more accurate than CT. PMID- 12133515 TI - [The use of three-dimensional ultrasound imaging in detecting the type and location of intrauterine contraceptive device]. AB - OBJECTIVE: To assess the value of three-dimensional ultrasound (3DUS) imaging in detecting the type and location of intrauterine contraceptive device (IUD). METHODS: Three-dimensional ultrasound imaging examination was conducted in 30 patients suspected with abnormality of IUD based on transvaginal two-dimensional ultrasound imaging examination. Through reconstruction of transvaginal 3DUS images, the type and location of IUDs were detected, and the results of hysteroscopy, laparoscopy or laparotomy were compared with the result of 3DUS. RESULTS: Satisfactory 3DUS images were obtained in 29 cases (96.7%) in which the type and location of IUDs and the relationship between IUD and uterine cavity were distinctly shown; satisfying imaging was not obtained in one menopausal case because of thinning of endometrium, the imaging only showed incarceration of IUD and not the relation between IUD fragment and uterine cavity. It was found that 17 IUDs (10 of single metal cirque type, 3 of gamma type, 2 of uterine cavity type, and 2 of T type) remained intact and 13 became fractured. Twenty-four IUDs were incarcerated, four were rotated (3 of gamma type and 1 of uterine cavity type), and 2 T type IUDs were normal. IUD was removed by hysteroscopy, laparoscopy, or laparotomy successfully at one go in 26 of the 28 cases with IUD abnormality. The findings of operation was consistent with the results of 3DUS. CONCLUSION: 3DUS is of great value in diagnosing IUD type and location, and helps select therapeutic plan. PMID- 12133516 TI - [Mobilization of autologous peripheral blood stem cells by cytosine arabinoside combined with recombinant human granulocyte colony-stimulating factor]. AB - OBJECTIVE: To observe the effect of cytosine arabinoside (Ara-C) combined with recombinant human granulocyte colony-stimulating factor (rhG-CSF) on mobilization of autologous peripheral blood stem cells (APBSCs) among malignant lymphoma patients and investigate appropriate dose of Ara-C. METHODS: Twenty-two patients with malignant lymphoma were randomly divided into two groups: in group A 3 g/m2.d-1of Ara-C was administered by intravenous drip in 2 divided doses for two days with the total dose of 6 g/m(2); in group B 5 g/m(2).d(-1) of Ara-C was administered by intravenous drip in two divided doses for two days with the total dose of 10 g/m(2). Blood routine examination was made every day. rhG-SCF at the dose of 300 microg.body(-1).d(-1) was injected subcutaneously once a day since the next day when the white blood cells reached the nadir until the end of APBCS harvest. APBSC harvest began when WBC >/= 5.0 x 10(9)/L and finished when the accumulated mononuclear cells >/= 5 x 10(5)/kg. RESULTS: The median of days when nadir of WBC appeared was 7 days in group A and 10 days in group B. The median of absolute neutrophil count (ANC) at ite nadir was 0.9X109/L in group A and 0.2 x 10(9)/L in group B. The median of days when ANC reached its nadir was 9 days in group A and 13 days in group B. The median of dose of rhG-CSF was 4.35 microg.kg( 1).d(-1) in group A and 4.35 microg.kg(-1).d(-1) in group B. The median of day when rhG-SCF administration began was the 11th day in group A and the 12th day in group B. The median of rhG-SCF administration days was 5 days in group A and 6 days in group B. The median of day when APBSC harvest began was the 15th day after Ara-C administration in group A and the 16th day in group B. The median of harvest time was 2 days in both groups. The volume, speed, and time of each apheresis were similar in the two groups. The time consumed and number of APBSCs collected in each harvest and the total number of APBSCs collected were similar in these two groups. CONCLUSION: Ara-C combined with rhG-CSF is safe and highly effective for APBSC mobilization. 6 g/m(2) is the suitable dose for APBSC mobilization. PMID- 12133517 TI - [Association between Bcl-2 gene polymorphism with systemic lupus erythematosus]. AB - OBJECTIVE: To study the association between Bcl-2 gene polymorphism and systemic lupus erythematosus (SLE) in Chinese and detect the synergism between IL-10 and Bcl-2 genotypes in determining susceptibility to SLE. METHODS: The peripheral white blood cells from 232 nuclear families of SLE and 106 ethnically matched normal controls were collected and the DNA extracted. The allelic distribution of one microsatellite located in Bcl-2 gene was determined by using fluorescent labeled primer and genotyping method. RESULTS: (1) The distribution frequency of Bcl-2 alleles and its major genotypes were significantly different between SLE group and normal controls (P = 0.001 and 0.0029 respectively). (2) Transmission disequilibrium existed in Bcl-2 alleles. Bcl-2-195 bp allele was preferentially transmitted to affected offspring (t: non-t = 122:82, P = 0.0051) by TDT approach. Individuals carrying both Bcl-2-195 bp allele and IL-10G138 bp allele were at higher risk of developing SLE than those carrying only Bcl-2-195 bp allele or IL-10G138 bp allele (OR = 3.00, 1.77, 1.07 respectively). CONCLUSION: Bcl-2 gene polymorphism is associated with SLE and may be directly involved in the process of SLE or in linkage disequilibrium with some susceptibility loci nearby. Synergistic effect may exist between IL-10 and bcl-2 genotypes in determining susceptibility to SLE. PMID- 12133518 TI - [Association between early-onset coronary heart disease and angiotensin II type 1 receptor gene polymorphism]. AB - OBJECTIVE: To study the association between early-onset coronary heart disease (CHD) and angiotensin II type 1 receptor (ATIR) gene A1166/C polymorphism and the effect of ATIR gene A1166/C on the plasma lipid level. METHODS: The ATIR A1166/C genotype of 71 early-onset CHD patients, 76 late-onset CHD patients, and 119 controls was examined by PCR-RFLP. Their plasma lipid levels were detected. RESULTS: Two ATIR genotypes, AC and AA, were detected. The frequencies (97.2% vs 92.1% vs 94.1%) of genotypes AA and AC and the frequencies of allele A and allele C in the three groups were similar. No difference were found among the levels of plasma lipids in the group of early-onset CHD patients with AA (94.1% vs 5.9%) or AC genotypes (2.8% vs 7.9%). CONCLUSION: There is no association between between ATIR gene A1166/C polymorphism and occurrence of early-onset CHD. ATIR geng A1166/C polymorphism has no effect on plasma lipid levels. PMID- 12133519 TI - [Relationship between angiotensin-1 converting enzyme gene polymorphism and in stent restenosis]. AB - OBJECTIVE: To investigate the relationship between angiotensin-1 converting enzyme (ACE) genotype and the incidence of in-stent restenosis (ISR). METHODS: One hundred and twenty eight patients, native of northeast China and of Han nationality, underwent coronary stenting consecutively from 1997 to 2000 and were followed up for 3 months to 2 years. They were divided into two groups according to the degree of stenosis of arterial lumen diagnosed during the second coronary angiography: ISR group (n = 43, with the lumen of stenting area stenosed by more than half of the diameter of the original lumen) and non-ISR group (n = 85, with the stenosis less than half of the diameter of original lumen). DNA of peripheral white blood cells was collected and ACE genotype was analyzed by PCR method. The demographic data, medical history, and body mass index of the patients were recorded. The cholesterol, triglycerin, blood sugar, systolic pressure, and diastolic pressure were examined 24 hours before stenting. RESULTS: More patients with type II diabetes were found in the ISR group than in the NISR group (P < 0.025). No difference in other clinical and biochemical indexes was found between these two groups (P > 0.05). The frequencies of DD, ID and II were 0.651, 0.163, and 0.186 respectively in ISR patients, and were 0.412, 0.282, and 0.305 respectively in NISR group. The restenosis rate was significantly higher in patients with ACD DD genotype than in patients with ID or II genotypes (P < 0.025). The frequency of D allele (73.26%) was significantly higher than that of I allele (26.74%) (P < 0.01). CONCLUSION: ACE polymorphisms are associated with ISR. The D alleles might be a risk factor for ISR. PMID- 12133520 TI - [A novel alternative splicing isoform of vascular endothelial growth factor gene in human beings]. AB - OBJECTIVE: To investigate a new alternative splicing isoform of vascular endothelial growth factor (VEGF) gene in human bejings. METHODS: The total RNA of the lung tissue of a legally aborted 4-month-old human fetus was isolated and then amplified by RT-PCR. The amplified product was cloned into the pMD18-T plasmid and pcDNA3.1(-). Then sequence analysis was conducted. RESULTS: The electrophoresis of the RT-PCR product showed one short band of VEGF121 comprising of 487 bp and a long band with two fragments: one normal VEGF165 comprising of 619 bp and one fragment comprising of 639 bp which was the same VEGF165 nucleotide sequence with a 20 bp fragment inserted between exon 3 and exon 4. Sequence analysis showed that this 20 bp long nucleotide was inserted from the 3' end of the third intron that contained splicing signal, thus causing shift mutation in reading frame of VEGF gene and early appearance of the stop code UAG in the middle of exon 4. CONCLUSION: A new alternative splicing isoform of VEGF has been identified in the lung tissue of a legally aborted human fetus. Its biological significance remains to be further investigated. PMID- 12133521 TI - [Expression of vascular endothelial growth factor in colorectal cancer and its clinical significance]. AB - OBJECTIVE: To evaluate the relationship between vascular endothelial growth factor (VEGF) and colorectal cancer. METHODS: Samples of cancer and adjacent normal mucosa were taken from 34 patients with colorectal cancer. The percentage and average fluorescence intensity pf VEGF positive cells in these samples were examined by using flow cytometry. RESULTS: The percentage of VEGF positive cells was 57% +/- 29% in cancer tissue and 42% +/- 24% in normal tissue (P < 0.05). The average fluorescence intensity of VEGF positive cells was 24% +/- 11% in cancer tissue and 16% +/- 7% in normal tissue (P < 0.01). The percentage of VEGF positive cells was 30% +/- 22% in cancer of Dukes stage B and 72% +/- 18% in cancer of Dukes stage C (P < 0.01). The average fluorescence intensity was 18% +/ 25% in cancer of Dukes stage B and 27% +/- 12% in cancer of Dukes stage C (P < 0.01). CONCLUSION: VEGF is associated with the development and prognosis of colorectal cancer. Its relation with degree of differentiation of colorectal cancer remains to be studied. PMID- 12133522 TI - [Immunocontraceptive effect of a synthetic chimeric peptide of sperm]. AB - OBJECTIVE: To investigate the immunocontraceptive ability of a new synthetic chimeric peptide with 35 amino acid residues. METHODS: A chimeric peptide with 35 amino acid residues was synthesized based on the amino acid sequence of SP17, a protein specific to murine testis/sperm, and cyritestin. The purified product, with keyhole limpet hemocyanin (KLH), was injected subcutaneously into the pad of female and male Balh/C mice. Blood was extracted from the ophthalmic arteries of the mice to identify the corresponding proteins from organs of mice and/or rat and human being by Western blotting. The chimeric peptide was injected into other female mice that were then raised together with male mice in the same cages to observe their status of pregnancy. Freund' adjuvant or normal saline were injected into the controls. RESULTS: The purity of the new chemiric peptide was 95%. The highest specific IgG titre in mouse serum was 1:6000 and the highest titre of specific IgA in the washing of vaginal mucous membrane of female mice was 1:300. The specific antiserum from the mice immunized by the chimeric peptide and KLH could identify two proteins with the molecular weights of 22 000 and 55 000 from testes of mice, rats, and human being. Western blotting showed no band for the extracts of mouse liver, spleen, and kidney. The pregnancy rate of female immunized mice was 61.11%, significantly lower that those in the controls (P < 0.05). The average number of cubs in a litter was less in the experimental group than in the controls (P < 0.05). CONCLUSION: The new chimeric peptide with 35 amino acid residues stimulates powerfully humoral and mucous immunity in mice and obviously decreases the pregnancy rate and the number of cubs in each litter in mice. PMID- 12133523 TI - [Histopathology of congenital pseudarthrosis of tibia]. AB - OBJECTIVE: To investigate the histopathology, origin, and etiology of congenital pseudarthrosis (CPT). METHODS: Specimens of periosteum from 28 CPT cases, 20 cases of traumatic pseudarthrosis (TP), 10 cases of fibromatosis, and 10 normal controls were examined. The pathological changes were observed by electron microscopy and confocal microscopy. The expression of alpha-smooth muscle (SM) actin, vimentin, desmin, bone morphogenic protein (BMP), interleukin (IL)-1, IL 6, tumor necrosis factor-alpha (TNF-alpha) and base fibroblast growth factor (b FGF) were detected by histochemistry and immunofluorescence chemical staining. Chromosomal karyotype was examined among 11 cases of CPT. RESULTS: (1) Electron microscopy showed that the periosteum and soft tissue between the broken ends in CPT were all dense fibrous connective tissue abundant in cells, including fibroblasts, myofibroblasts, etc. (2) The chief collagen element was type I collagen in normal periosteum and was type III collagen in periosteum of patients with CPT and fibromatosis (P < 0.025). (3) Vimentin was positive and desmin was negative in all specimens. The expression of alpha-SM actin was higher in specimens from CPT and fibromatosis than in specimens from normal control and TP (P < 0.01). The expression of BMP was higher in normal periosteum and TP than in the periosteum of CPT and fibromatosis (P < 0.05). The expression of IL-1, IL-6, TNF-alpha, b-FGF was higher in the periosteum of CPT and TP (P < 0.01). (4) The chromosomal karyotype of all CPT patients was normal 46XY or 46XX. CONCLUSION: (1) Neurofibromatosis is probably not the etiological factor of CPT. (2) CPT is a kind of invasive fibromatosis located in periosteum. (3) Abnormal expression of many kinds of cytokine and high expression of type III collagen play an important role in the pathogenesis of CPT. (4) The main pathology of CPT is hyperplasia of fibroblasts, thus causing thickening of periosteum, contractible circinate coarctation, and compression of tibia and surrounding tissues. (5) The chromosomal karyotype of patients with CPT is normal. PMID- 12133524 TI - [Augmentation of the immunostimulatory effects of plasmid DNA by incorporation of human CpG-oligodeoxynucletide]. AB - OBJECTIVE: To study whether the immunostimulatory activities of DNA vaccine could be enhanced by incorporation of human CpG-oligodeoxynucleotide (ODN) into its plasmid backbone. METHODS: Various copies of human CpG2006 sequence with immonostimulating activity for both human beings and mice were transfected into plasmid. Mouse spleen cells were cultured together with different kinds of plasmid DNA and then the amounts of IgM and interferon gamma (IFN-gamma) in the supernatant were examined by antibody sandwich ELISA. HBsAg plus plasmid DNA, as adjuvant, was injected into the shank muscles of 6 BALB/c mice in experimental group and recombinant HBsAg plus Al (OH)(2) was injected into 6 mice as controls. Four weeks later, the anti-HBsAg, IgG1 and IgG2a in their blood were examined. RESULTS: A minimum plasmid named pMini was constructed, containing only kanamycin resistant gene and pUC18 replication origin. By incorporating different amounts of CpG2006 into pMini plasmid, two recombinant plasmids, pCpG11 and pCpG26, with 9 and 16 copies of CpG2006 respectively, were derived. With the increase of number of CpG2006 copies, the ability of recombinant plasmid DNA to induce the secretion of IgM by cultured murine splenocytes increased accordingly in sequence of pCpG26 > pCpG11 > pMini, and its ability to induce the secretion of IFN-gamma by cultured murine splenocytes decreased instead in sequence of pCpG26 < pCpG11 > pMini. After the mice were immunized with recombinant HBsAg plus pMini and pCpG1, the anti-HBs total antibodies and IgG1 isotype in their blood remained unchanged (P > 0.05), but pCpG26 increased the levels of anti-HBs total antibodies and IgG1 isotype by 3 times (P < 0.001) and 2 times (P < 0.02) respectively. The three kinds of plasmid DNA increased the level of specific IgG2a isotype by 5 approximately 10 times at similar degrees (P > 0.05). CONCLUSION: The immunostimulatory activities of plasmid DNA can be enhanced by incorporation of human CpG-ODN. PMID- 12133525 TI - [Treatment of aortic arch aneurysm with graftstent by endovascular surgery]. AB - OBJECTIVE: To study the feasibility of treatment of aortic arch aneurysm with graftstent by intravascular surgery. METHODS: Animal model of aortic arch aneurysm was established using 10 piglets. An aortic arch-shaped graftstent was transferred and implanted to the site of aortic arch aneurysm through femoral artery under angiography. RESULTS: The aortic arch-shaped graftstent was successfully implanted in 8 piglets. Two piglets died from anaesthetic accident. Angiography showed disappearance of aneurysm, splendid development of the branches of aortic arch, and no blood leakage around the stent. CONCLUSION: Treatment of aortic arch aneurysm by implantation of graftstent by endovascular surgery is feasible. PMID- 12133526 TI - [Expression and localization of Nogo-A mRNA in rat nerve tissue]. AB - OBJECTIVE: To investigate the expression and localization of Nogo-A mRNA in brain, spinal cord, optic nerve, and sciatic nerve. METHODS: Frozen sections of brain, spinal cord, optic nerve, and sciatic nerve of 8 rats were made. In situ hybridization was conducted and the sections were observed by light microscopy. RESULTS: The expression of Nogo-A mRNA was positive in brain, spinal cord, and optic nerve, and negative in sciatic nerve. CONCLUSION: The expression of Nogo-A mRNA is negative in brain, spinal cord, and optic nerve, which may be related to the poor axon regeneration in central nervous system. PMID- 12133527 TI - Efficacy and safety of therapeutic angiogenesis from direct myocardial administration of an adenoviral vector expressing vascular endothelial growth factor 165. AB - OBJECTIVE: To investigate whether direct administration of adenoviral vectors (Ad) containing the complementary deoxyribonucleic acid (cDNA) of vascular endothelial growth factor 165 (Ad-VEGF165) induces porcine coronary collateral vessel formation, improves regional myocardial perfusion and function and is safe. METHODS: Three weeks after miniature swine underwent left thoracotomy and placement of an Ameroid constrictor on the left circumflex coronary artery (LCX), Ad-VEGF165 (n = 6) or the control, Ad expressing beta-galactosidase cDNA (Ad-Gal, n = 6), was directly administered into the ischemic myocardium in the circumflex distribution. All animals were sacrificed 4 wk after the second surgery. Myocardial perfusion and function were assessed by electrocardiogram-gated single photon emission computed tomography (GSPECT) imaging. Ex vivo coronary angiography was performed to examine collateral vessels. Toxicity was assessed by blood analyses on the day just before (day 0) and on day 1, 3, 7, 28 after vector delivery and by vascular, myocardial and liver histology after sacrifice. RESULTS: GSPECT imaging 4 wk after administration of Ad-VEGF165 demonstrated significant reduction in ischemic area (P < 0.01) and rest ischemic severity (P < 0.01) and significant improvement in the left ventricular ejection fraction (P < 0.01) and regional wall motion (P < 0.05) compared with that of Ad-Gal and before administration of Ad-VEGF165. Collateral vessel development assessed by coronary angiography was significantly greater in the Ad-VEGF165 group than in the Ad-Gal group (P < 0.05). General safety parameters, including routine blood parameters, liver and kidney function and cardiac specific parameters demonstrated no difference between Ad-VEGF165 and Ad-Gal animals except for the red blood cell count on day 28 (P < 0.05) and blood urea nitrogen on day 7 (P < 0.05). Only transient elevations in creatine phosphokinase (P < 0.05) and aspartate transaminase (P < 0.05) on day 1 were revealed compared with that before vector administration in both groups. Histologically, no atherosclerotic lesion in the circumflex and no inflammation in liver were revealed and only a small myocardial necrosis was observed in one Ad-VEGF165 animal (area < or = 20%) and one Ad-Gal animal (area < 10%). CONCLUSIONS: Ad-VEGF165 can induce coronary collateral vessel formation, improve regional myocardial perfusion and function and is safe by means of direct injection, which suggesting that this strategy may be useful in treating human ischemic heart disease. PMID- 12133528 TI - Role of angiotensin II and angiotensin II receptors in vascular smooth muscle cell migration in vitro. AB - OBJECTIVE: To determine the biotic effects of angiotensin II (Ang II) on the migration of rat smooth muscle cells (VSMCs) and investigate the mechanisms involved in the development of vascular injury. METHODS: VSMCs isolated from aortic media of Wistar rats and cultured by the modified explant method were adopted. In the presence and absence of Ang II, the expression of Ang II receptor (ATR) and reorganization of the actin cytoskeleton and focal adhesion of VSMCs were studied by an immunocytochemistry technique and fluorocytochemistry technique. Migration assays were performed with a modified Boyden's chamber. The effects of AT(1)R antagonist (CV-11974), AT(2)R antagonist (PD123319) on the aforementioned target were studied. RESULTS: VSMCs migration was stimulated by adding Ang II. The dynamic reorganization of actin cytoskeleton and focal adhesions may be an important mechanism by which Ang II facilitates VSMCs motility. The expression of AT(1)R in VSMCs could be upregulated initially after treatment with Ang II, then decreased gradually. The expression of AT(1)R was downregulated by AT(1)R antagonists. The effect of Ang II on VSMCs migration was mediated by AT(1)R, while AT(2)R had no significant effect. CONCLUSIONS: The dynamic reorganization of focal adhesions and the actin cytoskeleton is required for Ang II-induced VSMCs migration. This effect is mediated by AT(1)R. PMID- 12133529 TI - Linkage analysis of a region on chromosome 2 with essential hypertension in Chinese families. AB - OBJECTIVE: To verify the linkage of the candidate regions identified in a previous study (markers D2S168, D2S151, D2S142 on chromosome 2) with hypertension in Chinese families. METHODS: A genetic linkage study focused on chromosome 2 was performed on 240 Chinese families containing 856 patients with essential hypertension. A total of 1080 individuals were genotyped using 9 highly polymorphic microsatellite markers around the candidate regions on chromosome 2 with an average spacing of 5 cM. Non-parametric linkage (NPL), parametric linkage analysis and transmission-disequilibrium test (TDT) with the GENEHUNTER software were used to assess evidence for linkage. RESULTS: Linkage of a region on chromosome 2 around D2S151 and D2S142 with hypertension was confirmed by different statistical methods (NPL, LOD score and TDT). However, the linkage of D2S168 could not be replicated in this extension study. CONCLUSIONS: The data suggest that a region on chromosome 2 at or near the loci of D2S142 and D2S151 may harbor genes responsible for the development of essential hypertension in Chinese. PMID- 12133530 TI - Characteristics of coronary microvascular lesions in autopsied elderly with hypertensive left ventricular hypertrophy. AB - OBJECTIVE: To observe the characteristics of coronary microvascular lesions (CML) in the autopsied elderly cases with hypertensive left ventricular hypertrophy (LVH) and the difference of CML among the groups of essential hypertension (EHT), coronary heart disease (CHD) and diabetes (NIDDM) also with LVH. METHODS: A retrospective study was performed in 206 cases > or = 60 years old of EHT, CHD and NIDDM with LVH and 30 normal cases as control, out of 3195 consecutive autopsied cases from 1954 to 1996 in our hospital. Arterioles with diameters of 10 - 60 microm and capillaries in the muscular layer were shown by the methods of HE, Elastic fiber + VG staining and immunohistochemistry of CD31. Quantitative measurements on the arteriole density (AD), the ratio of arteriolar wall and cavity (RWC), capillary density (CD) and the area of endothelial cell (AEC) were performed with light microscope observation and image analysis by computer. According to the thickness of the left ventricle free wall, the severity of LVH was divided into four degrees from 0 to III. LVH of degree 0-III was observed in EHT group, while only LVH of degree I was found in CHD, EHT + CHD, and NIDDM groups. SAS system was used for statistical analysis. RESULTS: AD and RWC increased while CD and AEC decreased significantly with the progression of LVH in EHT groups (P < 0.05 - 0.01). There was a similar but more severe change in the (HT + CHD) group (P < 0.01); the AD increased (P < 0.05) while all other measurements did not show obvious changes in the CHD group. The AD increased, CD and AEC decreased (all P < 0.05), but RWC did not change very much in the NIDDM group. CONCLUSION: CML in the EHT group was characterized by an increased AD and RWC, decreased CD and AEC, among which the increased RWC was the typical change in EHT groups compared with the groups of CHD and NIDDM. Damaged CML may be one of the main factors for decreased coronary flow reserve and myocardial ischemia in cases of EHT with LVH. PMID- 12133531 TI - Changes of sarcoplamic reticular Ca(2+)-ATPase and IP(3)-I receptor mRNA expression in patients with atrial fibrillation. AB - OBJECTIVE: To investigate changes in the expression of sarcoplamic reticular Ca(2+)-ATPase (SERCA) and IP(3)-I receptors (IP(3)R(1)) mRNA in patients with atrial fibrillation. METHODS: Thirty-eight patients with mitral stenosis undergoing open heart surgery were studied. 100 mg of atrial tissue was obtained during surgery from the right appendage and the right atrium. The amount of messenger ribonucleic acid (mRNA) amount of SERCA and IP(3)R(1) was measured by reverse transcription-polymerase chain reaction (RT-PCR) and normalized to the mRNA levels of glyceraldehyde 3-phosphate dehydrogenase (GAPDH). RESULTS: Levels of mRNA expression of SERCA in patients with AF, as compared with subjects in sinus rhythm, was lower and that of IP(3)R(1) was higher. The longer AF was sustained, the higher the levels of mRNA. There was no significant difference between right atrial free wall and right appendage. CONCLUSIONS: The expression changes of SERCA and IP3R mRNA may correlate with the initiation or maintenance of AF. PMID- 12133533 TI - Relationship between a novel polymorphism of lipoprotein lipase gene and coronary heart disease. AB - OBJECTIVE: To investigate polymorphisms in the gene for lipoprotein lipase (LPL) in Chinese populations with coronary heart disease (CHD) and to inquire into the relationship between these polymorphisms in LPL gene and CHD. METHODS: Genomic DNA was extracted from patients with CHD and normal control subjects using a salting out method. The entire coding region and flanking sequences of all coding exons of the LPL gene were amplified by PCR technique and PCR products were detected by denaturing high-performance liquid chromatography (DHPLC) and sequenced with a dideoxy terminal termination method. RESULTS: A novel polymorphic site, G830A, that is within the fifth exon of the LPL gene was found. The 192 codon CGA was changed into CAA and resulted in the substitution of glutamine for arginine. Between the control and CHD groups, chi-square test showed no significant difference in the frequencies of the A/A genotype and A allele (P > 0.05). However, the frequencies of A/A genotype and A allele (0.653 and 0.786) in CHD patients with high plasma triglyceride/lowed plasma high density lipoprotein cholesterol were higher than those (0.415 and 0.642) in CHD patients without hyperlipidemia (P < 0.05). CONCLUSION: No direct association was found between the LPL Arg192-->Gln substitution polymorphism and CHD, but there is a significant positive correlation between the A/A genotype of the LPL gene and CHD associated with high triglyceride/lowed high density lipoprotein cholesterol. This study may provide new data for exploring the molecular mechanism of CHD. PMID- 12133534 TI - Follow-up of coronary artery lesions caused by Kawasaki disease and the value of coronary angiography. AB - OBJECTIVE: To investigate the course of coronary artery lesions caused by Kawasaki disease, and the value of coronary angiography (CAG) and two-dimensional echocardiography (2-D Echo) in the evaluation and follow-up of coronary artery lesions. METHODS: Eighty seven patients with coronary artery lesions caused by Kawasaki disease from 1979 to 1997 were retrospectively analyzed. One hundred and sixty-seven CAGs were performed in 87 patients during follow-up. CAG was repeated every 1-3 years in each patient until complete regression was confirmed. 2-D Echo was performed before CAG each time. The longest period of follow-up was 16 years and 6 months. Patients were treated with aspirin or aspirin and warfarin. RESULTS: During follow-up, the coronary artery lesions regressed in 48/87 (55%) patients, however, they developed into severe coronary artery lesions in 6/87 (7%) patients in whom coronary artery bypass surgery was performed. The coronary artery aneurysm regressed in some patients, while stenotic lesions remained or developed. The ratio of coronary artery stenotic lesions to aneurysms increased progressively. This study showed that Echo diagnosis of coronary artery lesions has "false positives" and "false negatives". Only 76% of coronary aneurysms and 18% of stenotic lesions could be found by 2-D Echo. No stenotic lesion could be found in distal segments of the coronary artery. CONCLUSIONS: Long term follow up revealed spontaneous regression occurred in 55% of patients and development into severe coronary artery stenosis in 7%. It is necessary to perform long-term follow-up in patients with coronary artery lesions caused by Kawasaki disease. 2 D Echo can not completely replace CAG during follow-up of coronary artery lesions caused by Kawasaki disease. PMID- 12133532 TI - Protein kinase A-mediated phosphorylation of HERG potassium channels in a human cell line. AB - OBJECTIVE: To investigate the molecular mechanism of human ether-a-go-go-related gene (HERG) potassium channels regulated by protein kinase A (PKA) in a human cell line. METHODS: HERG channels were stably expressed in human embryonic kidney (HEK) 293 cells, and currents were measured with the patch clamp technique. The direct phosphorylation of HERG channel proteins expressed heterologously in Xenopus laevis oocytes was examined by (32)P labeling and immunoprecipitation with an anti-HERG antibody. RESULTS: Elevation of the intracellular cAMP concentration by incubation with the adenylate cyclase activator, forskolin (10 micromol/L), and the broad range phosphodiesterase inhibitor, IBMX (100 micromol/L), caused a HERG tail current reduction of 83.2%. In addition, direct application of the membrane permeable cAMP analog, 8-Br-cAMP (500 micromol/L), reduced the tail current amplitude by 29.3%. Intracellular application of the catalytic subunit of protein kinase A (200 U/ml) led to a tail current decrease by 56.9% and shifted the activation curve by 15.4 mV towards more positive potentials. HERG WT proteins showed two phosphorylated bands, an upper band with a molecular mass of approximately 155 kDa and a lower band with a molecular mass of approximately 135 kDa, indicating that both the core- and the fully glycosylated forms of the protein were phosphorylated. CONCLUSIONS: PKA-mediated phosphorylation of HERG channels causes current reduction in a human cell line. The coupling between the repolarizing cardiac HERG potassium current and the protein kinase A system could contribute to arrhythmogenesis under pathophysiological conditions. PMID- 12133535 TI - Nuclear factor-kappa B in signal conduction of protein kinase C in T lymphocytes from an asthmatic guinea pig model. AB - OBJECTIVE: To explore the role of nuclear factor-kappa B (NF-kappa B) in the signal conduction of protein kinase C (PKC) regulated proliferation, apoptosis and expression of Th2 cytokines -- interleukin-4 (IL-4) and interleukin-5 (IL-5) of T lymphocytes in the bronchial alveolus lavage fluid (BALF). METHODS: T lymphocytes were isolated and purified from BALF of asthmatic guinea pigs in normal and asthmatic groups, and were stimulated with PKC agitator phorbol 12 myristate 13-acetate (PMA) and NF-kappa B inhibitor pyrrolidine dithiocarbamate (PDTC), respectively. The expressions of NF-kappa B, IL-4 and IL-5 mRNA and protein, the proliferation and apoptosis of T lymphocytes were observed by immunohistochemistry, in situ hybridization, ELISA, MTT and TUNEL, respectively. RESULTS: The activation of NF-kappa B, proliferation response, and expression of IL-4 and IL-5 mRNA and protein in T lymphocytes stimulated by PMA were significantly higher than those of their blank control (P < 0.01), while those indexes of T lymphocytes stimulated by PMA and PDTC simultaneously were significantly lower than those stimulated by PMA alone (P < 0.01). The apoptotic index of T lymphocytes stimulated with PMA were significantly lower than that of their blank control (P < 0.01), and the apoptotic index of asthmatic guinea pig T lymphocytes stimulated with PMA and PDTC simultaneously were significantly higher than that stimulated by PMA alone (P < 0.01). The significant positive correlations were found between the activation of NF-kappa B and the proliferation (r = 0.64, P < 0.001), and the expression of IL-4 and IL-5 mRNA and protein of T lymphocytes, respectively (r = 0.55 - 0.68, P < 0.001). There was also significant negative correlation between the activation of NF-kappa B and apoptosis of T lymphocytes (r = 0.62, P < 0.001). CONCLUSIONS: NF-kappa B may participate in the signal conduction of PKC regulated proliferation, apoptosis and expression of IL-4 and IL-5 of T lymphocytes in asthma. The activation of NF kappa B in PKC signal conduction pathway of T lymphocytes may play an important role in the pathogenesis of asthma. PMID- 12133536 TI - Increased expression of neuropeptide Y and its mRNA in STZ-diabetic rats. AB - OBJECTIVE: To study the relationship between neuropeptide Y (NPY) and diabetes by examining the content and distribution of NPY and its mRNA expression in the hypothalamus and pancreas of STZ-diabetic rats. METHODS: Thirty Wistar rats were randomly divided into 3 groups (diabetic group, diabetic insulin treatment group, and control group). After feeding for 24 weeks, the rats were sacrificed. The expression of NPY in the hypothalamus and pancreas was detected with immunohistochemistry and in situ hybridization. RESULTS: (1) The hypothalamic content of NPY and its mRNA were significantly increased in STZ-diabetic rats in comparison with normal controls. Increased expression of NPY mRNA was found only in the arcuate nucleus and not in the paraventricular nucleus in diabetic rats, suggesting that NPY was produced in the arcuate nucleus. (2) The hypothalamic content of NPY and its mRNA in STZ-diabetic rats were visibly reduced after insulin treatment compared with that in untreated diabetic rats. This supports the hypothesis that insulin deficiency in the brain may be responsible for increased hypothalamic NPY gene expression in diabetic rats. (3) The increase of hypothalamic NPY in STZ diabetic rats associated with hyperphagia and polydipsia could be reversed by insulin replacement, suggesting that increased hypothalamic NPY contributes to the pathophysiological progress of the diabetic state. (4) The present study demonstrated for the first time that the content of NPY and its mRNA in the pancreas was increased in STZ-diabetic rats, and that the distribution of NPY-positive cell in islets was changed from the periphery to the whole islet. The content and distribution of NPY and its mRNA in islets were not changed by insulin treatment. CONCLUSION: Increased NPY in the hypothalamus results in hypophagia and polydipsia, while the implication of increased NPY in the pancreas of diabetic rats is not clear. PMID- 12133537 TI - Alterations in vascular reactivity in single- and double-transgenic mice coexpressing human APP-C100 and mutant SOD(1) genes. AB - OBJECTIVE: To explore the mechanism underlying changes in microvascular reactivity in single- and double-transgenic mice. METHODS: Peripheral vascular reactivity to the vasodilators, acetylcholine and sodium nitroprusside, on perfused microvasculature of the hind footpad was investigated using nontransgenic mice, single-transgenic mice expressing the human APP-C100 (TgC100. WT or TgC100. V717F) and double-transgenic mice coexpressing human APP-C100 and human SOD(1) (G93A) genes. RESULTS: Single TgC100 and double Tg mice C100/SOD(1) (G93A) at 2 - 3 months old showed a statistical decrease of 28% in blood flux compared to nontransgenic control mice. In addition, vasodilative responsiveness was markedly reduced to 34% in 8 - 9 months old TgC100 mice compared to control mice. There was no significant difference in the profile of vasodilative reaction between TgC100. WT and TgC100. V717F mice. TgC100 and double Tg mice also had higher levels of A beta peptide in plasma than nontransgenic mice (P < 0.01). CONCLUSIONS: The present study suggests that the altered reactivity of the microvasculature may be mediated by circulating soluble A beta peptides. The mechanisms underlying the vasoactivity of circulating A beta in TgC100 and double Tg mice may involve both the endothelium and nonendothelium. PMID- 12133538 TI - Clinical significance of plasma tissue factor pathway and urokinase-type plasminogen activator system in cancer patients. AB - OBJECTIVE: To evaluate variations in the plasma tissue factor (TF) and urokinase type plasminogen activator (u-PA) system and their relationship with clinical cancer type, pathological classification and metastatic status in cancer patients. METHODS: Plasma levels of TF and its inhibitor (TFPI), as well as u-PA and its receptor (u-PAR) were measured using ELISA in 76 patients with malignant tumors and 24 patients with benign tumors. RESULTS: Plasma levels of TF and u-PAR in the malignant tumor group were significantly higher than those of the benign tumor group and the normal control. U-PA and u-PAR increased significantly in patients with esophageal and gastric cancer. However, most of these parameters except TFPI did not vary according to pathological classification. A significant elevation was evident in patients with local infiltration, lymph node involvement and distal metastasis, while u-PAR only increased in the latter two categories. CONCLUSIONS: Both the TF and u-PA systems are activated in cancer patients. U-PA and its receptor might prove to be a clinically useful marker for disease progression. PMID- 12133539 TI - Mycophenolate mofetil vs cyclophosphamide therapy for patients with diffuse proliferative lupus nephritis. AB - OBJECTIVE: To make an open label prospective trial for comparing the therapeutic effects of mycophenolate mofetil (MMF) vs cyclophosphamide (CYC) pulse therapy on patients with diffuse proliferative lupus nephritis (DPLN). METHODS: Forty-six patients with biopsy proven active DPLN were enrolled in this study. Twenty-three patients were given MMF orally at a dosage of 1.0 - 1.5 g/d (MMF Group). Another 23 cases received conventional intermittent CYC pulse therapy (CYC Group). Supplemental steroid treatment was offered in the same manner to both groups. The age, sex distribution and severity of renal damage were matched in two groups. Therapeutic effects were evaluated at the end of six-month treatment. Fifteen patients in the MMF Group and 12 patients in the CYC Group had repeated renal biopsy at that time. RESULTS: MMF therapy was more effective in reducing proteinuria and hematuria. A 50% reduction of urinary protein and urinary red blood cell excretion from baseline value in 69.6% and 91.3% patients in the MMF Group, while only 47.8% and 65.2% in the CYC Group. MMF was more effective in inhibiting autoantibody production (especially anti-dsDNA antibody) and in decreasing serum cryoglobulin levels. Pathologically, the MMF group showed more markedly reduction in glomerular immune deposits with less glomerular necrosis, and less microthrombi, less crescent formation and vascular changes in the repeated renal biopsy as compared with the CYC group. Adverse reactions related to the treatment included gastrointestinal symptoms 26.1% and 43.5% in the MMF and CYC Groups respectively, infection 17.4% in the MMF group and 30.4% in the CYC group. CONCLUSION: MMF was more effective in controlling the clinical activity of DPLN and renal vascular lesions as compared with CYC pulse therapy in a 6 month follow-up study. PMID- 12133540 TI - Expression of cyclin genes in human gastric cancer and in first degree relatives. AB - OBJECTIVE: To clarify the role of these cyclins in human gastric cancer. METHODS: 38 gastric cancer patients, 29 first degree relatives of gastric cancer patients, as well as 18 healthy subjects were included. The mRNA expression of cyclins D1, D2, D3 and E in gastric biopsies was evaluated by RT-PCR analysis using specific primers. Histomorphological features such as intestinal metaplasia, atrophy, H. pylori infection and severity of gastritis were determined by the updated Sydney System. RESULTS: Significant mRNA overexpression was found for cyclins D2, D3 and E compared with healthy normal specimen, but cyclin D1 expression was not different between tumor and normal tissues. In addition, cyclin D2 and D3 overexpression was significantly more frequent in first degree relatives than in healthy controls (P < 0.05). Among the various pathological findings, the overexpression of cyclins D2 and E was associated with intestinal metaplasia, and the overexpression of cyclin D3 was associated with intestinal metaplasia as well as atrophy. The overexpression of cyclins D2 and D3 was significantly correlated with H. pylori infection. No correlation was observed between the overexpression of cyclin D1 and any pathological variables. CONCLUSION: The overexpression of cyclins D2, D3 and E is a frequent event in patients with gastric cancer and their first degree relatives and may be an early event in gastric carcinogenesis. PMID- 12133541 TI - Effect of Acanthopanax giraldii Harms Var. Hispidus Hoo polysaccharides on the human gastric cancer cell line SGC-7901 and its possible mechanism. AB - OBJECTIVE: To study the inhibitory effect of Acanthopanax giraldii Harms Var. Hispidus Hoo polysaccharides (AGP) on SGC-7901 gastric cancer cells and its possible mechanism. METHODS: Cell doubling time analysis, colony forming assay and MTT assay were adopted to study the inhibitory effect and its characteristics. We also analyzed the amount of protein expressed by oncogenes, antioncogenes and cell factors using flow cytometric analysis. RESULTS: AGP inhibited the proliferation of SGC-7901 cells and cell colony forming ability. AGP did not inhibit the viability and function of lymphocytes of peripheral blood in healthy subjects and human embryonic tenocytes, except for the highest dosage of AGP (P < 0.05), which slightly inhibited the viability and function of the two types of normal cells. AGP inhibited the viability and function of SGC-7901 cells, except for the lowest dosages of AGP I and AGP III. There was a dose effect relationship between the dosage of the AGP and SGC-7901 cells. The effect of the AGP at the molecular level was associated with the low protein expression of the c-myc and bcl-2 genes and the high protein expression of the p53, bax, fas and fas-L genes, as well as the cell factor TGF beta(1). The inhibitory effect of AGP was weaker than that of CDDP, but was stronger than that of Vitamin C. CONCLUSIONS: Acanthopanax giraldii Harms Var. Hispidus Hoo polysaccharides selectively inhibited the proliferation, the colony forming ability, and the viability and function of human gastric cancer cells through the low protein expression of c-myc, bcl-2 and the high protein expression of p53, fas, fas-L and the cell factor TGF beta(1). The different inhibitory characteristics on the normal cells and cancer cells are possibly caused by gene and the cell factor expressions. PMID- 12133542 TI - Cloning and expression and purification of hepatitis B e-antigen precursor in Escherichia coli. AB - OBJECTIVE: To investigate the role of the 25 kD hepatitis B e antigen (HBeAg) precursor that only exist inside hepatocytes and study its effect on the pathopoiesis of hepatitis B and QIAGEN expression and purification system. METHODS: Hepatitis B virus (HBV) preC/C gene for the 25 kD HBeAg precursor was cloned into the expression vector pQE30 and the 25 kD HBeAg precursor was expressed in Escherichia coli (E. coli) and purified. Its antigenicity for 21 kD mature HBeAg was tested by western blot analysis. RESULTS: Cloned fragments in the expression vector were sequenced and verified to be homogeneous with that of HBV (ayw subtype). Expression of the HBeAg precursor in E. coli under the transcriptional regulation of T5 promoter yielded a soluble cytosolic protein with an apparent molecular mass of 25 kD. Recombinant HBeAg precursor exhibited identical potencies with 21 kD mature HBeAg that reacted with anti-HBeAg antibodies. The purification rate of the expressed HBeAg precursor was up to 89.6% and the yield of purified HBeAg precursor from this procedure was 2.4 mg/L. CONCLUSION: 25 kD HBeAg precursor exhibited biological activity and might play an important role in pathopoiesis of hepatitis B. PMID- 12133543 TI - Adenovirus induced acute hepatitis in non-human primates after liver-directed gene therapy. AB - OBJECTIVE: To define the mechanism of acute hepatitis in non-human primates after liver directed gene therapy. METHODS: Differences in immune response exhibited by 8 rhesus monkeys receiving adenovirus (Ad) or lipofectamine-mediated gene transfer by various routes, the time course, and the nature of the specific immune responses to both adenoviral vectors and transgene products were studied using HE staining (H&E) and immunohistochemical staining. RESULTS: The monkeys developed mild to moderate acute hepatitis 1 to 3 weeks after intravenous or intrabiliary injection of first generation replication-defective adenoviruses carrying the Escherichia coli lacZ gene. This was accompanied by adenovirus mediated T-cell proliferation and neutralizing antibodies to the adenovirus. Increased numbers of CD3(+), CD4(+) and CD8(+) T-lymphocytes were detected in the diseased livers, while B-lymphocytes were absent. Hepatocytes demonstrated increased expression of beta 2-microglobulins (beta 2-MG) and HLA-DR antigens in the plasma membranes. The development of acute hepatitis and the accompanying immune abnormalities were delayed in immunosuppressed monkeys until after the discontinuation of immunosuppressive therapy. The monkeys infused with Ad. CMVluc showed more significant and longer durations of hepatitis than the monkeys infused with adenoviruses carrying the lacZ gene. Lipofectamine-mediated gene transfer was inefficient. There was neither lacZ expression nor significant immune response in the liver of monkeys infused with lipofectamine via the portal vein or the common bile duct. CONCLUSION: Immune response to the hepatocytes in liver directed gene therapy is MHC class I restricted and T-cell mediated. Both adenoviral vectors and foreign genes are related to the liver damage. Mild to moderate hepatic inflammation seen with the E-1 deleted vector is reversible. Immunosuppression regimens may prolong transgene expression and delay the development of acute adenoviral hepatitis. PMID- 12133544 TI - Growth hormone improves graft mucosal structure and recipient protein metabolism in rat small bowel transplantation. AB - OBJECTIVE: To observe the effects of recombinant human growth hormone (rhGH) on graft structure and recipient protein metabolism in rat small bowel transplantation (SBT) and total parenteral nutrition (TPN) models. METHODS: Twenty recipients of rat allogeneic heterotopic small bowel transplants (SD- >Wistar) were divided into two groups (GH group and control group). Both groups were supported by standard TPN. Acute rejection was suppressed with CsA 10 mg x kg(-1) x d(-1) intramuscularly. All rats in the experimental group received subcutaneous rhGH 1 U x kg(-1) x d(-1) after transplantation. Morphological mucosal indices of transplanted gut and metabolic parameters such as body weight, nitrogen balance, urinary 3-methyl histidine excretion and serum albumin of the recipients were compared between two groups. RESULTS: The application of rhGH promoted graft recovery significantly compared with standard TPN support alone. On postoperative day 14, all morphological indexes of transplanted gut recovered to the preoperative state. Protein metabolism in the recipient was also significantly improved. rhGH decreased the catabolism of protein, accelerated regaining of positive nitrogen balance and corrected hypoalbuminemia. CONCLUSION: GH is an effective metabolic intervention in SBT. It may promote the structural repair of the graft and correct the metabolic disturbance. It is useful in improving the outcome of clinical SBT. PMID- 12133545 TI - Radical resection of gastric carcinoma with pancreas and spleen preservation and functional cleaning of lymph nodes. AB - OBJECTIVE: To study the clinical value of radical resection of gastric carcinoma with pancreas and spleen preservation (PSP) and functional cleaning of lymph nodes (LNs) of the spleen hillus and along the splenic artery. METHODS: Pancreas and spleen involvement was retrospectively reviewed among 439 cases of resectable carcinoma of the gastric cardia, gastric corpus and total stomach. During gastric surgery, 2 ml of methylene blue was injected into the subserosal space of the gastric cardia or corpus to observe the spread of lymphatic flow in 54 cases of gastric carcinoma. The metastatic rate of LNs in splenic hillus and along the trunk of the splenic artery (No10, No11), postoperative complications and survival rates were investigated in 63 gastric carcinoma patients that had received gastrectomy with pancreas and spleen preservation (PSP). These were compared with the pancreas preservation (PP) group and pancreas and spleen combined resection (PSR) group. RESULTS: Among these 439 cases, only 25 cases were observed with direct invasion to the pancreas (5.7%), and 10 cases with direct invasion to the spleen (2.3%). After pathological examination of the pancreatic body and tail, we found 22 cases with pancreas and spleen combined resection, 4 cases (18.2%, 4/22) with direct invasion of the capsule and 2 with invasion to the superficial parenchyma (9.1%, 2/22), without metastasis to the lymph nodes within the pancreas and spleen. The metastatic rate of No10, No11 lymph nodes were 17.5% (11/63) and 19.1% (12/63) in the PSP group, 20.8% (45/216) and 25% (54/216) in the PP group, and 20% (6/30) and 23.3% (7/30) in the PSR group. There were no statistically significant differences (P > 0.05). Injection of methylene blue into the subserosal space of the stomach did not diffuse into the spleen or pancreatic parenchyma. Postoperative complications, diabetes and mortality in PSP (0%, 0%, 0%) were lower than in PP (4.2%, 0.9%, 0.9%) or PSR (40%, 10%, 3.3%). The 5-year survival rate (5-YSR) and 10-YSR in PSP (57.5%, 52.0%) were higher than in PSR (37.5%, 30.0%). Those patients with stage II and III(a) treated by PSP, improved markedly. CONCLUSIONS: The surgical procedure of pancreas and spleen preservation for gastric cancer is a safe and organ function protected method. Postoperative complications were lower and survival rates were higher, the radicality was not reduced. These results indicate that PSP is preferred in patients with gastric carcinoma of stage II or III(a). PMID- 12133546 TI - Nitric oxide synthase gene expression in injured spinal cord tissue. AB - OBJECTIVE: To investigate gene expression of three nitric oxide synthase isozymes in injured spinal cord tissue. METHODS: Thirty-six adult SD rats were randomly divided into six groups: a normal group and five injury groups, with six per each group. Animals in the injury groups were sacrificed at 2, 6, 12, 24, 48 h after injury. A compression injury model on the spinal cord was made according to Nystrom B et al and gene expression of the three NOS isozymes were examined by reverse transcription polymerase chain reaction (RT-PCR). RESULTS: Gene expression of nNOS and eNOS were detectable in the normal group and were up regulated quickly after injury, reaching a maximum at 6 h: (0.633 +/- 0.012) and (1.236 +/- 0.207). Gene expression of iNOS was detectable only in the injury groups and it was gradually up-regulated after injury, reaching a maximum at 24 h: (1.043 +/- 0.049). CONCLUSION: Injury to the spinal cord leads to early up regulation of cNOS and late up-regulation of iNOS. Different NOS isozymes may play different roles in secondary spinal cord injury. PMID- 12133547 TI - Prognostic value of matrix metalloproteinase-2 and tissue inhibitor of metalloproteinase-2 in bladder carcinoma. AB - OBJECTIVES: To detect the level of matrix metalloproteinase-2 (MMP-2) mRNA and the tissue inhibitor of metalloproteinase-2 (TIMP-2) mRNA in bladder transitional cell carcinoma (BTCC), and to estimate the prognosis for bladder tumor based on the quality and quantity of MMP-2 and TIMP-2 mRNA. METHODS: Thirty-five samples of human BTCC and 15 normal fresh bladder tissues were studied by RT-PCR analysis followed by computer-assisted image analysis. RESULTS: The level of the MMP-2 mRNA in BTCC was significantly increased compared with that in normal bladder epithelium. The positive rates of MMP-2 and TIMP-2 mRNA were 71.4% and 65.7% in BTCC, and 66.7% and 60.0% in the normal bladder wall. The expression intensity of the MMP-2 mRNA by image analysis tended to increase with tumor grading and staging, which showed statistical significance. Similarly, the MMP-2 to TIMP-2 ratio also showed statistically significant difference between normal bladder tissue and bladder carcinoma (P < 0.01). CONCLUSIONS: A high level of the MMP-2 mRNA exists in BTCC, which may function to damage collagen IV inside the basement membrane and the extracellular basement of the bladder. The level of the MMP-2 mRNA is proportional to BTCC grading and staging, which may have prognostic value. The MMP-2 to TIMP-2 ratio may play a more significant role in the determination of the aggressiveness and prognosis of bladder tumor. PMID- 12133548 TI - Renal cell carcinoma related novel gene, GYLZ-RCC18: cloning and functional studies. AB - OBJECTIVE: To clone the full length of renal cell carcinoma (RCC) related novel gene GYLZ-RCC18 and study its function. METHODS: SMART RACE technology was used to clone the full length of GYLZ-RCC18. RT-PCR was used to detect its expression in renal cell carcinoma tissue at different stages and grades. We transfected the antisense oligonucleotide of GYLZ-RCC18 to renal cell carcinoma cell line, GRC-1, and analyzed proliferation activity, growth rate, apoptosis, and mortality changes. RESULTS: The full length of GYLZ-RCC18 (GenBank accession number: BE825133) cDNA was about 3.5 kb. GYLZ-RCC18 had a higher expression in higher grades and stages of renal cell carcinoma than in lower ones. The expression of GYLZ-RCC18 in renal cell carcinoma was much higher than in normal kidney. After the transfection of GYLZ-RCC18 antisense oligonucleotide, the mortality of GRC-1 increased significantly, while proliferative activity and growth rate were substantially inhibited at the same time. The antisense oligonucleotide induced apoptosis of GRC-1 through the entire observation time. CONCLUSION: GYLZ-RCC18 is an important novel gene related to renal cell carcinoma. Overexpression of this gene results in higher growth and proliferative activity and has an antiapoptosis effect on renal cell carcinoma cells. Transfection of the antisense oligonucleotide may inhibit the generation and development of renal cell carcinoma. PMID- 12133549 TI - Endothelin-1, an important mitogen of smooth muscle cells of spontaneously hypertensive rats. AB - OBJECTIVE: To study the features of vascular smooth muscle cell (VSMC) proliferation induced by endothelin-1 (ET-1). METHODS: VSMCs of spontaneously hypertensive rats (SHR) and Wistar-Kyoto rats were cultured and treated with ET 1. Basic fibroblast growth factor (bFGF) gene expression was measured using both Northern blot and an enzyme-linked immunoassay. RESULTS: ET-1 resulted in an increase in bFGF transcripts at 8 - 24 h; bFGF levels were significantly higher in VSMCs treated with ET-1 than in those not treated. However, VSMCs growth responses in SHR and WKY were different. Smooth muscle cells of SHR were hyper responsive to ET-1. Maximal bFGF mRNA levels were elevated 3.5-fold at 4 h of stimulation in WKY and 8-fold at 8h in SHR4. Moreover, the proliferation of VSMCs induced by ET-1 was inhibited by antisense phosphorothioate oligodeoxynucleotides (10 micromol/L AS-bFGF) but not sense bFGF oligomers at the same concentrations, being reduced by 80% in SHR and 40% in WKY vs control, respectively. Furthermore, the effect of AS-bFGF oligomers on SHR SMC proliferation is significantly greater than on WKY SMC proliferation. CONCLUSION: ET-1 may be required for exaggerated vascular growth responses in SHR and bFGF may be involved. PMID- 12133550 TI - Novel mitochondrial 16S rRNA mutation, 3200T-->C, associated with adult-onset type 2 diabetes. AB - OBJECTIVE: To investigate the role of a potential diabetes-related mitochondrial region, which includes two previously reported mutations, 3243A-->G and 3316G- >A, in Chinese patients with adult-onset type 2 diabetes. METHODS: A total of 277 patients and 241 normal subjects were recruited for the study. Mitochondrial nt 3116 - 3353, which spans the 16S rRNA, tRNA(leu(UUR)) and the NADH dehydrogenase 1 gene, were detected using polymerase chain reaction (PCR), direct DNA sequencing, PCR-restriction fragment length polymorphism and allele-specific PCR. Variants were analyzed by two-tailed Fisher exact test. The function of the variants in 16S rRNA were predicted for minimal free energy secondary structures by RNA folding software mfold version 3. RESULTS: Four homoplasmic nucleotide substitutions were observed, 3200T-->C, 3206C-->T, 3290T-->C and 3316G-->A. Only the 3200T-->C mutation is present in the diabetic population and absent in the control population. No statistically significant associations were found between the other three variants and type 2 diabetes. The 3200T-->C and 3206C-->T nucleotide substitutions located in 16S rRNA are novel variants. The 3200T-->C caused a great alteration in the minimal free energy secondary structure model while the 3206C-->T altered normal 16S rRNA structure little. CONCLUSIONS: The results suggest that the 3200T-->C mutation is linked to the development of type 2 diabetes, but that the other observed mutations are neutral. In contrast to the Japanese studies, the 3316G-->A does not appear to be related to type 2 diabetes. PMID- 12133551 TI - The effect of calmodulin antagonist berbaminederivative-EBB on hepatoma in vitro and in vivo. AB - OBJECTIVE: To evaluate the anti-hepatoma effect of Calmodulin antagonist 0 - 4 ethoxyl-butyl-Berbamine (EBB), one of the berbamine derivatives. METHODS: Monotetrazolium (MTT) method was used to analyze the effect of EBB on the proliferation and growth inhibition effect. Of a hepatoma cell line in vitro. A mouse hepatoma model was induced by injection of hepatoma cells (H22) in the abdominal cavity. The effect of EBB on survival at different concentrations as well as in combination with 5-FU were investigated in vivo. Flow cytometry analysis, dot blot hybridization, western blot, immunochemistry, enzyme-linked lectin assay (ELISA), trifluoperazine (TFP) and electron microscopic observation were used to study the effect of EBB on cell cycle process, P53 mRNA and protein levels, calmodulin content and ultrastractural changes of hepatoma cells. RESULTS: EBB exerts a very strong inhibitory effect on human hepatoma cell line 7402 and mouse hepatoma cell line H22 in vitro. The IC(50) value of EBB for the two cell lines are 3.312 microg/ml and 1.167 microg/ml, respectively. The sensitivity of H22 cells to 5-FU can be markedly enhanced: The IC(50) dosage of 5 Fu can be decreased from 0.75 microg/ml down to 0.15 microg/ml, when jointly administered with nontoxic dosages of EBB (IC(10)). In vivo, EBB can prolong the lifespan of mice with ascites H22 to more than three months. 64% of mice survived, while all animals in the control group died by the 18th day. When EBB (5 mg x kg(-1) x d(-1)) is jointly used with 5-FU (25 mg x ml(-1) x d(-1)), 73% of mice with ascites H22 survived, much higher than 27% in the 5-FU treated group. EBB can enhance the anti-hepatoma ability of 5-Fu treatment. EBB mechanism against hepatoma: P53 expression in the EBB treated group is substantially higher than that in the control group. EBB increased the translation of P53. As a calmodulin antagonist, EBB decreases amount of the CaM in hepatoma cells and blocked the hepatoma cell proliferation cycle at the G(2)M phase. Before the G(0)/G(1) phase, a diploid peak and apoptic cells in the treated groups were observed. CONCLUSIONS: The CaM antagonist, EBB, has a strong anti-hepatoma effect and enhances the effect of 5-FU, induces hepatoma cell to apoptosis, promotes the P53 protein expression and decreases the amount of CaM in the cytoplasm. All these results demonstrate that EBB is a new and potentially useful drug against hepatoma and should be researched further. PMID- 12133552 TI - The intraspecific difference of the triose phosphate isomerase (tim) gene from Giardia lamblia. AB - OBJECTIVE: To investigate the intraspecific difference of the triose phosphate isomerase (tim) gene from Giardia lamblia (G. lamblia). METHODS: Total genomic DNA of G. lamblia was extracted and partial fragments of the triose phosphate isomerase (tim) gene were amplified by polymerase chain reaction (PCR). All nucleotide sequences were analyzed by using a phylogenetic analysis, which was constructed with parsimony and Neighbor-joining (N-J) methods. RESULTS: A total of 124 variable sites (23% of all sequences detected) was defined, most of which were found at the silent sites of codons. Two similar phylogenetic trees were constructed, subdividing 16 Giardia isolates into two groups. CONCLUSION: The genetic diversity of G. lamblia appeared to be little affected by factors of both host and geography, while natural-selection played an important role in DNA molecular evolution level of the tim gene. The tim gene may be considered a very useful genetic marker of the population genetic structure of G. lamblia. PMID- 12133553 TI - Transplantation of corneal stem cells cultured on amniotic membrane for corneal burn: experimental and clinical study. AB - OBJECTIVE: To investigate the proliferation and differentiation of cultured corneal stem cells and determine the effect of corneal stem cells cultured on amniotic membranes on the limbal area for treating corneal burns. METHODS: The proliferation and differentiation of corneal stem cells in vitro had been examined using colony-forming efficiency and immunohistochemistry. The stem cells had been cultured on amniotic membranes and transplanted to the limbal area for treating corneal burns. RESULTS: Corneal stem cells had a high proliferation capacity in primary and first passage, cytokeratin 3 was not expressed in primary culture but partly in first passage. The stem cells could proliferate to form cell layer on an amniotic membrane. When transplanted, stem cells could survive on limbus. After transplantation, ocular inflammation resolved, the cornea re epithelialized, the stromal opacity reduced, the superficial neovascularity was lessened and the conjunctival fornix re-established. CONCLUSIONS: Ocular surface conditions could be improved by allograft of corneal stem cells cultured on amniotic membranes. PMID- 12133554 TI - Low dose transdermal scopolamine increases cardiac vagal tone in patients after acute myocardial infarction. AB - OBJECTIVE: To investigate whether transdermal scopolamine increased cardiac vagal activity in patients during the acute phase of myocardial infarction. METHODS: 30 patients with a first acute myocardial infarction and preserved sinus rhythm who were on no drug that could influence the sinus node were randomly assigned to either treatment group or placebo group. Measures of heart rate variability (HRV) in patients given drug or placebo were obtained by digital 24 hour Holter recording before and after treatment. Baroreflex sensitivity was performed using the phenylephrine method. RESULTS: No significant differences was found in the indices of the time domain and the frequency domain in both groups before treatment. Patients with transdermal scopolamine showed a significant increase in the standard deviation of normal RR intervals (SDNN), standard deviation of all five min mean normal RR intervals (SDANN), root mean square of differences of successive normal RR intervals (rMSSD), total power (TP, 0.000. - 0.40 Hz), low frequency peak (LF, 0.040 - 0.15 Hz), high frequency peak (HF, 0.15 - 0.40 Hz), and Baroreflex sensitivity after treatment (P < 0.05 - 0.01). These indices did not change in patients given placebo. CONCLUSION: Low doses of transdermal scopolamine safely increase cardiac parasympathetic activity and improve autonomic indices in patients with acute myocardial infarction. PMID- 12133555 TI - Prevalence rate of osteoporosis in the mid - aged and elderly in selected parts of China. AB - OBJECTIVE: To understand the distribution of prevalence rate of osteoporosis in the middle - aged and elderly in parts of China. METHODS: Bone mineral density (BMD) was measured and questionnaires were taken for 5593 people aged above 40 years in five administrative areas in China selected by the stratified - multi - steps - cluster sampling method. RESULTS: The total prevalence rate of osteoporosis was 16.1%. The prevalence rate among males was 11.5% and among females was 19.9% (P < 0.01). There were also osteoporosis prevalence differences among cities, age groups, gender groups and areas. CONCLUSION: The prevention and treatment of osteoporosis are very important for females but also should not be ignored in males. PMID- 12133556 TI - Rehabilitation therapy for short bowel syndrome. AB - OBJECTIVE: To investigate the effect of rehabilitation therapy for short bowel syndrome on patient nutritional status and intestinal adaptation. METHODS: The rehabilitation therapy included enteral or parenteral nutrition, glutamine, recombinant human growth hormone and rehabilitative diet. From January 1997 to July 2000, twenty - seven patients with short bowel syndrome received the treatment. The average age of the patients was 38.5 +/- 19.3 years, and the length of residual small intestine ranged from 15 to 80 cm, with an average of 46.8 +/- 23.4 cm. The ileocecal valve was preserved in 14 cases, and the average time between the onset of short bowel syndrome and the rehabilitation therapy was 86 +/- 105 days. RESULTS: After the treatment, nutritional status of the patients improved markedly, and intestinal absorptive capacity improved. Eight patients were followed up for more than 2 years, among whom 4 (50%) were weaned from total parenteral nutrition. Thirteen patients were followed up for more than 1 year, and 10 patients (76.9%) were weaned from total parenteral nutrition. CONCLUSIONS: Rehabilitation therapy for short bowel syndrome can improve patient nutritional status effectively and promote intestinal adaptation, providing a new hope for these patients. The therapeutic effects are related to the length of the residual small intestine, patients age and duration between massive intestinal resection and start of the treatment. Early initiation of rehabilitation therapy promotes intestinal adaptation and increases patients ability to wean from total parenteral nutrition. PMID- 12133557 TI - Complete duplication of the colon: definitive management by resection of the duplication without the normal bowel. PMID- 12133558 TI - Hypertrophic cardiomyopathy with apical left ventricular aneurysm: a case report. PMID- 12133559 TI - Vestibulo-ocular, optokinetic and postural function in humans with rheumatoid arthritis. AB - The present study investigated vestibulo-ocular reflex (VOR), optokinetic reflex (OKR) and postural function in patients with rheumatoid arthritis (RA). Compared with controls, no differences in gaze-holding, VOR gain or phase, OKR slow phase velocity (SPV) or quick phase amplitude, optokinetic afternystagmus SPV or duration, or latency to the illusion of circularvection, were found. RA patients did exhibit greater sway in the leftward direction (P<0.01), however, this was no greater in the conditions of the Clinical Test of Sensory Interaction and Balance that increase reliance upon vestibular information. We conclude that RA patients do not exhibit substantial deficits in visual-vestibular function. PMID- 12133560 TI - Castration increases nisoxetine-evoked norepinephrine levels in vivo within the olfactory bulb of male rats. AB - In the present experiment we compared differences in extracellular norepinephrine levels in vivo within the olfactory bulb of intact and castrated male rats following infusion of the norepinephrine transport inhibitors, nisoxetine and tomoxetine. With this approach it was possible to assess whether dynamic changes in in vivo norepinephrine transporter function occur as a function of the gonadal state of the animal. Norepinephrine levels following infusion of nisoxetine were significantly increased in castrated compared with intact male rats. While a similar trend was present in response to tomoxetine infusion, these differences failed to achieve a statistically significant difference. These results demonstrate that castration of male rats alters norepinephrine transporter function within the olfactory bulbs. The increased extracellular levels of norepinephrine in response to agents which inhibit transporter function suggest that castration reduces transporter activity. Such effects have important implications not only with regard to processes involving the norepinephrine system in the olfactory bulb but also to the generalized sites and mechanisms by which gonadal steroid hormones modulate central nervous system functions. PMID- 12133561 TI - Correlation between activity in neuron B52 and two features of fictive feeding in Aplysia. AB - The present study examined the correlation between the level of activity neuron B52 and the transition from protraction to retraction phases of buccal motor patterns (BMPs) and the termination of the BMPs. The level of activity in B52 during the protraction phase was positively correlated with the duration of that phase. A second burst of activity in B52 was associated with the termination of the retraction phase. An apparent monosynaptic inhibitory connection from B52 to B64, may mediate the effects of B52. The first burst of activity in B52 delays the onset of activity in B64, thereby prolonging the protraction phase, and the second burst inhibits activity in B64, thereby terminating the retraction phase. These results suggest that activity in B52 may contribute to switching between ingestion-like and rejection-like BMPs by regulating both phase transition and termination of BMPs. PMID- 12133562 TI - Repetitive transcranial magnetic stimulation of the supplementary motor area (SMA) degrades bimanual movement control in humans. AB - Moving the upper limbs at a common tempo according to an in-phase or anti-phase mode represents elementary coordination dynamics. Previously, the role of the supplementary motor area (SMA) has been emphasized for successful production of these patterns. The objective of this study was to investigate whether repetitive transcranial magnetic stimulation (rTMS) of the SMA at 5 Hz can interfere with these isofrequency configurations in the post-stimulation stage. Results showed a deterioration of temporal control as a function of coordinative complexity. This effect was associated with a decrease in the functional coupling between the primary motor cortices, as measured by electroencephalographic coherence. These data suggest that rTMS of the SMA can modify interhemispheric communication and accordingly modulate interlimb behavior. PMID- 12133563 TI - Alpha-lipoic acid prevents ethanol-induced protein oxidation in mouse hippocampal HT22 cells. AB - Oxidative stress is involved in a number of neurological disorders, including the neurotoxic effects of ethanol. Recent studies have described a neuroprotective potential of alpha-lipoic acid (LC) in several models of neuronal cell death related to oxidative stress. We tested the hypothesis that LC could be effective in preventing ethanol-induced neurotoxicity employing the clonal hippocampa cell line HT22. A 24 h incubation with ethanol 100-600 mM caused a dose-dependent loss of cell viability and a significant increase of the overall intracellular protein oxidation. Coincubation with LC 0.1 mM resulted in a significant decrease of ethanol-related neurotoxicity and a complete prevention of the ethanol-induced intracellular protein oxidation. These results indicate that the radical scavenging properties of LC are effective to ameliorate ethanol-induced neurotoxicity. PMID- 12133564 TI - Nerve growth factor induces systemic hyperalgesia after thoracic burn injury in the rat. AB - Acute burn injury is usually associated with pain in the injured and nearby areas. However, we have recently reported that a thoracic scald induces hindpaw hyperalgesia during the healing stage in rats. The present study investigated the cause of the remotely occurring hyperalgesia. Behavioral testing using the von Frey test revealed that rats developed hyperalgesia in the neck and flank as well as the hindpaw 2-3 weeks after injury. The concentration of nerve growth factor (NGF) in the skin of the chest increased markedly during the healing stage. Moreover, rats injected daily with anti-NGF serum after burn injury did not develop hyperalgesia, suggesting that increased NGF in the tissue of the healing skin is a key factor causing systemic hyperalgesia during the recovery stage. PMID- 12133565 TI - Broca's area in the human brain is involved in the selection of grammatical gender for language production: evidence from event-related functional magnetic resonance imaging. AB - The neural correlates of the selection of grammatical gender during overt picture naming were investigated by event-related functional magnetic resonance imaging in the left hemisphere. Relative to simply naming a picture, the production of the definite determiner of the picture name (requiring gender selection) resulted exclusively in pronounced activation of a single region in the superior portion of Broca's area. This activation was not present in contrasts reflecting lexical access (naming a picture vs. saying "jaja" to a smiley) or articulation (saying "jaja" vs. rest). Rather, lexical access activated other inferior frontal regions, insula, fusiform and inferior temporal gyrus. Articulation involved insula, Rolandic operculum, motor and premotor cortex and superior temporal gyrus. The results are discussed with respect to data from studies investigating gender processing during language comprehension. PMID- 12133566 TI - Deep brain stimulation of subthalamic neurons increases striatal dopamine metabolism and induces contralateral circling in freely moving 6-hydroxydopamine lesioned rats. AB - Deep brain stimulation (DBS) of the subthalamic nucleus (STN) alleviates Parkinson's disease (PD) symptoms. Although widely used, the mechanisms of action are still unknown. In an attempt to elucidate those mechanisms, we have previously demonstrated that STN-DBS increases striatal extracellular dopamine (DA) metabolites in anaesthetized rats. PD being a movement disorder, it remains to be determined whether these findings are related to any relevant motor or behavioural changes. Thus, this study investigates concomitant behavioural changes during STN-DBS and extracellular striatal DA metabolites measured using microdialysis in freely moving 6-hydroxydopamine-lesioned rats. STN-DBS induced an increase of striatal DA metabolites in awake, freely moving animals. Furthermore, we observed concomitant contralateral circling behaviour. Taken together, these results suggest that STN-DBS could disinhibit (consequently activate) substantia nigra compacta neurons via inhibition of gamma-aminobutyric acid-ergic substantia nigra reticulata neurons. PMID- 12133568 TI - Structure of visual evoked magnetic field during sleep in humans. AB - To investigate the effects of sleep on the visual evoked magnetic fields (VEF), we recorded VEF following flash light stimulation in healthy adults during sleep. The awake VEF contained several components with approximate latencies of 40, 55, 65, 80, 100, 110, 150, and 180 ms. In contrast, the sleep VEF contained mainly three components with approximate latencies of 65, 100 and 115 ms. By comparing the magnetic components between the awake and sleep conditions based on similarities in the contour pattern of the isomagnetic field, three components for the sleep condition were found to be enhanced, those at 55, 80-100 and 100 110 ms in the awake VEF. Other components of the awake VEF may be reduced or disappear during sleep. This large change in the VEF during sleep suggests that some qualitative changes occur in the cortical visual processing, for example, a reduction in the inhibitory activities at works while awake. PMID- 12133567 TI - Lack of association of two lipoprotein lipase polymorphisms with Alzheimer's disease. AB - A recent genetic study demonstrated associations between an altered risk of Alzheimer's disease (AD) and two polymorphisms in the lipoprotein lipase (LPL) gene, Asn291Ser and Ser447Ter. LPL immunostains senile plaques, and is a ligand of the low-density lipoprotein receptor-related protein (LRP), a major apolipoprotein E (apoE) receptor. LPL increases the cellular uptake of apoE via LRP, and polymorphisms in LPL alter its ability to mediate apoE-LRP interactions, with potential implications for AD pathogenesis. Here, we tested the genetic association of LPL with AD in a case-control study. For the Asn291Ser polymorphism, we analyzed 277 individuals (141 AD, 136 control) and found no significant difference in allele frequencies between the AD and control groups. For the Ser447Ter polymorphism, we analyzed 187 individuals (108 AD, 79 control) and again found no significant difference in allele frequencies between the AD and control groups. Thus, our study does not support associations between AD and two common polymorphisms in LPL. PMID- 12133569 TI - Cu(2+) inhibition of glycine-activated currents in rat sacral dorsal commissural neurons. AB - The effect of Cu(2+) on glycine (Gly) response was examined in neurons acutely dissociated from the rat sacral dorsal commissural nucleus (SDCN) using the nystatin perforated patch clamp recording configuration under voltage-clamp conditions. Cu(2+), in the concentration range 10-1000 microM, reversibly inhibited chloride current activated by 30 microM Gly at a holding potential of 40 mV with an IC(50) of 88.4 microM. Cu(2+) shifted the Gly concentration response curve to the right in a parallel manner, which indicated that Cu(2+) decreased the apparent affinity of the receptor for Gly. Cu(2+) suppression of Gly-activated current was independent of membrane potential between -60 and +60 mV and did not involve a shift in the reversal potential of the current. Furthermore, Cu(2+) antagonized the inhibitory action of Zn(2+) in a concentration-dependent manner, suggesting a common site or mechanism of action of Cu(2+) and Zn(2+) on Gly receptors. The results show that Cu(2+) is a potent inhibitor of Gly receptor-mediated responses in rat spinal neurons. PMID- 12133570 TI - Amylin and glucose co-activate area postrema neurons of the rat. AB - Glucose is an important metabolic factor controlling feeding behavior. There is evidence that physiologically relevant glucose sensors reside in the caudal hindbrain. The area postrema (AP) in particular, which has been characterized as a receptive site for the anorectic hormone amylin, may monitor blood glucose levels. To determine whether glucose and amylin co-activate the same subset of AP neurons, we performed extracellular single unit recordings from a rat AP slice preparation. In 53% of all AP neurons tested (n=32), the activity was positively correlated to the glucose concentration. Interestingly, there was a coincidental sensitivity (94%) of AP neurons to glucose and amylin, which exerted excitatory effects on these cells. We conclude that the co-sensitivity of AP neurons to glucose and amylin, both increasing in response to food intake, points to the AP as an important hindbrain center for the integration of the metabolic and hormonal control of nutrient intake. PMID- 12133571 TI - Inverse relationship between p27/Kip.1 and the F-box protein Skp2 in human astrocytic gliomas by immunohistochemistry and Western blot. AB - The F-box protein Skp2 regulates G1-S transition by controlling p27/Kip.1. The deregulated expression of p27/Kip.1 plays a critical role in the pathogenesis of many human tumors. Its cellular levels depend on ubiquitin-mediated degradation. Recently, Skp2 has been demonstrated to mediate p27/Kip.1 degradation and to have oncogenic properties. In a series of astrocytic gliomas, immunohistochemistry and Western blot of p27/Kip.1 and Skp2 have been compared. p27/Kip.1 decreased with anaplasia and almost disappeared in glioblastomas (GBM), whereas Skp2 was absent or poorly expressed in well differentiated astrocytomas and it was diffusely or focally expressed in most GBM. Since the expression of Skp2 increases during G1-S transition, the correlation of Skp2 levels with malignancy might simply reflect the highest percentage of proliferating cells in anaplastic gliomas or alternatively be instrumental to p27/Kip.1 degradation. PMID- 12133572 TI - Involvement of adenosine in the anti-allodynic effect of amitriptyline in streptozotocin-induced diabetic rats. AB - Recent observations suggest the involvement of adenosine in the peripheral antinociceptive effect of amitriptyline in nerve-injury-induced neuropathic pain. The aim of the present investigation was to evaluate, firstly, the peripheral and systemic effects of amitriptyline on tactile allodynia in the streptozotocin (STZ)-induced diabetic rat model of neuropathic pain and, secondly, whether caffeine coadministration affects the actions of amitriptyline. Diabetes was induced by a single intraperitoneal (i.p.) injection of STZ (50 mg/kg), and tactile allodynia was detected by application of von Frey filaments to the ventral surface of the hindpaw. Both systemic (0.5-2.0 mg/kg, i.p.) and peripheral (10-100 nmol, subcutaneously (s.c.)) administration of amitriptyline were found to produce increases in paw withdrawal thresholds, at higher doses. Coadministration of caffeine (5 mg/kg, i.p.; 1500 nmol, s.c.), at doses which produced no effect on its own, partially reversed systemic and local anti allodynic effects of amitriptyline. These results indicate an anti-allodynic effect of both peripheral and systemic amitriptyline, and suggest the involvement of endogenous adenosine in the action of amitriptyline in this rat model of painful diabetic neuropathy. These data also suggest that topical application of tricyclic antidepressants may be useful in treating neuropathic pain in diabetics. PMID- 12133573 TI - Modeling both the mechanical and hypoxic features of traumatic brain injury in vitro in rats. AB - In traumatic brain injury, the brain is subjected to mechanical shear and varying degrees of hypoxia/ischemia. To compare effects of stretch injury, hypoxia, and the combination of both insults on neurons, mixed neuronal and astrocytic cultures were established from day 17 fetal rat brains. On days 17-19 in vitro, cultures were subjected to stretch injury or hypoxia of varying degrees, alone and in combination. Cultures were assayed for lactate dehydrogenase release and Trypan Blue uptake. Hypoxia or Stretch injury alone induced a graded response (P<0.05) on both assays. Stretch+Hypoxia (4 or 6 h) resulted in significantly greater injury as compared with controls (P<0.05), and as compared with either isolated Stretch or Hypoxia (1, 2, 4 or 6 h) alone (P<0.05). PMID- 12133574 TI - Inhibitory effects of intrathecally administered interleukin-1beta on carrageenan induced hyperalgesia and spinal c-Fos expression in rats. AB - The present study investigated the effects of interleukin-1beta (IL-1beta) injected intrathecally (i.t.) on carrageenan-induced thermal hyperalgesia and spinal c-Fos expression. The paw withdrawal latency (PWL) by the thermal stimulus was taken as an index of the thermal hyperalgesia of rats. I.t. injection of 10 ng IL-1beta significantly increased the PWL of the carrageenan-injected paw. Expression of c-Fos induced by intraplantar (i.pl.) injection of carrageenan was examined in the spinal cord with immunohistochemical methods. Three hours after i.pl. injection of carrageenan, the number of c-Fos-like immunoreactive (c-Fos LI) neurons was significantly increased in laminae I-II, III-IV and V-VI of the ipsilateral spinal cord at L4-5 with the higher density in laminae I-II and V-VI. I.t. pre-injected IL-1beta significantly decreased the number of carrageenan induced c-Fos-LI neurons in laminae I-II in the ipsilateral spinal cord and also inhibited the hyperalgesia induced by i.pl. carrageenan. These results suggested that i.t. injection of IL-1beta suppressed the central nociceptive input into laminae I-II and produced an antinociceptive effect. PMID- 12133576 TI - Event-related skin conductance responses to musical emotions in humans. AB - While the reasons underlying musical emotions are unclear, music is nevertheless a powerful elicitor of emotion, and as such, may induce autonomic nervous system responses. One typical measure of this neural pathway is the skin conductance response (SCR). This response generally depends upon stimulus arousal, one of the two motivational determinants of emotion. The objective of the present study was to verify whether emotional reactions to music elicit such event-related autonomic responses. To this aim, four musical emotions varying in arousal were employed: fear, happiness, sadness and peacefulness. SCRs were found to be greater with the two more stimulating emotions, fear and happiness, as compared to the two more relaxing emotions, sadness and peacefulness (P<0.05). In addition, subjects' ratings of the emotional clarity for each excerpt did not parallel the corresponding SCRs magnitudes. The results show that SCRs can be evoked and modulated by musical emotional arousal, but are not sensitive to emotional clarity. While several studies have been performed with visual scenes and environmental sounds, the present study brings similar evidence from the musical domain. PMID- 12133575 TI - Morphine microinjections into the rat nucleus submedius depress nociceptive behavior in the formalin test. AB - Our previous studies have indicated that the thalamic nucleus submedius (Sm) is involved in modulation of nociception and plays an important role in an endogenous analgesic system (a feedback loop) consisting of spinal cord-Sm ventrolateral orbital cortex-periaqueductal gray-spinal cord. To investigate whether opioids are involved in this antinociception pathway, the effects of microinjection of morphine and naloxone into the Sm on the nociceptive behavior (agitation) evoked in the formalin test were investigated in the awake rat using an automated movement detection system. The results indicate that a unilateral microinjection of morphine (5 micro g, 0.5 microl) into the Sm suppresses the formalin-induced agitation response, but does not influence spontaneous motor activity, and that the morphine-induced depression can be reversed by microinjection of the opioid receptor antagonist naloxone (1.0 micro g, 0.5 microl) into the same Sm site. The results suggest that opioid receptors in the Sm may be involved in the Sm-mediated depression of persistent inflammatory pain. PMID- 12133577 TI - Calpain-dependent neurofilament breakdown in anoxic and ischemic rat central axons. AB - Neurofilaments are key structural components of white matter axons. The effect of in vitro anoxia or oxygen-glucose deprivation (OGD) on the integrity of the 160 and 200 kDa neurofilament isoforms was studied by immunoblot, and correlated with physiological function. Adult rat optic nerves were exposed to 60 min of either anoxia or OGD. Compound action potential area recovered to 22+/-6% of control after 60 min of anoxia, and to 4+/-1% after 60 min of OGD. Ca(2+)-free (+EGTA) perfusate allowed complete recovery after OGD (108+/-42%). Tetrodotoxin (TTX, 1 microM) was less protective (45+/-6%). Both anoxia and OGD induced breakdown of neurofilament 160 (NF160) and NF200 revealed by the appearance of multiple lower molecular weight bands mainly in the 75-100 kDa range. Zero-Ca(2+)/EGTA completely prevented NF breakdown. TTX only partially reduced NF160 degradation. Non-phosphorylated NF200 appeared after reperfusion post-anoxia or OGD, and was also greatly reduced by zero-Ca(2+) or TTX. Calpain inhibitors (10 microM calpain inhibitor I or 50 microM MDL 28,170) significantly reduced NF160 and NF200 breakdown/dephosphorylation, but did not improve electrophysiological recovery. Significant calpain-mediated breakdown of NF160 and NF200 indicates structural damage to the axonal cytoskeleton, which was completely Ca(2+)-dependent. While pharmacological inhibition of calpain alone greatly reduced NF proteolysis, there was no concomitant improvement in function. These results imply that calpain inhibition is necessary but not sufficient for white matter protection, and emphasize the existence of multiple Ca(2+)-dependent degradative pathways activated in injured white matter. PMID- 12133578 TI - Distribution of serotonin 5-hydroxytriptamine 1B (5-HT(1B)) receptors in the normal rat hypothalamus. AB - This is the study first attempting to evaluate distribution of neurons expressing serotonin 5-hydroxytriptamine 1B (5-HT(1B)) receptors in hypothalamus by using immunocytochemistry. The 5-HT(1B)-immunoreactive neurons were widely distributed in hypothalamus. Accumulations of 5-HT(1B) neurons occurred in magnocellular nuclei, supraoptic nucleus, paraventricular nucleus (dorsolateral part) and accessory perifornical, circular and retrochiasmatic nuclei. Magnocellular neurons manifested an intense immunostaining suggesting a high level of 5-HT(1B) receptors. Large and middle-sized neurons with different 5-HT(1B) staining patterns were scattered throughout lateral hypothalamus, periventricular nucleus and lateral preoptic area. Immunofluorescent double-labeling revealed a great overlapping of the distribution 5-HT(1B) neurons and dense network of neuropeptide Y-immunoreactive fibers in paraventricular, supraoptic and arcuate nuclei. The potential functional significance of 5-HT(1B) receptors in the 5-HT control of endocrine functions and feeding are discussed. PMID- 12133579 TI - Sex differences in delta opioid receptor immunoreactivity in rat medial amygdala. AB - Although gonadal hormones can modulate the opioid system in several limbic structures, it remains unclear if there are sex differences in amygdalar opioid systems. This study compared enkephalin immunoreactivity, delta opioid receptor immunoreactivity (DORir), and mu opioid receptor immunoreactivity in the amygdala of male and proestrous female rats. A striking sex difference in DORir was observed in the posterodorsal region of the medial nucleus of the amygdala, with males showing much greater DORir than females. No other sex differences were seen in amygdalar regions. Although the functional significance of this sexually dimorphic staining for DORir in medial amygdala is unknown, it may contribute to sex differences in reproductive functions, social behaviors, and/or stress responses. PMID- 12133580 TI - Oleamide attenuates apoptotic death in cultured rat cerebellar granule neurons. AB - The effect of oleamide on apoptosis was investigated by using 3-(4,5 dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide reduction assay, DNA staining assay with propidium iodide and caspase-3 activity analyses. The present results showed that oleamide significantly attenuated the cell death and nuclear condensation of cultured rat cerebellar granule neurons induced by K(+) deprivation in a dose-dependent manner. The oleamide actions were well parallel with the attenuation of caspase-3 activity in the process of apoptotic death. Moreover, neither elaidic acid nor stearic acid, two fatty acids structurally related to oleamide without the Delta(9)-cis double bond, had similar effects on the cell death, suggesting the selectively structural features of oleamide required for this action. These data provided the first evidence of a protective effect of oleamide against apoptosis in a structurally specific manner. PMID- 12133581 TI - Clearance of manganese from the rat substantia nigra following intra-nigral microinjections. AB - Chronic exposure to manganese (Mn) positively correlates with the occurrence of Parkinsonism but little is known about mechanisms of its neurotoxicity. In the present study, we determined the clearance of Mn from rat substantia nigra after its nigral injection and correlated it with the establishment of apomorphine induced rotational behaviour and loss of striatal tyrosine hydroxylase (TH) immunoreactivity. Our results suggest that Mn is slowly cleared from the substantia nigra, following a first-order kinetics with a t(1/2) of 3 days. Appearance of apomorphine-induced rotational behaviour and loss of TH immunoreactivity within the striatum follows metal clearance were both detected 24 hours after intra-nigral Mn microinjection and maximal 72 hours after injection. The present data suggest that the cellular mechanisms induced by Mn and leading to dopaminergic cell death, occurred shortly after its injection and that the metal concentration needs to reach a threshold value to induce neurotoxic effects. This would indicate that nigral damages are a direct consequence of Mn accumulation. PMID- 12133582 TI - Adenosine-mediated activation of Akt/protein kinase B in the rat hippocampus in vitro and in vivo. AB - Adenosine is considered an endogenous neuroprotective metabolite that through activation of the A(1) receptor results in reduction of neuronal damage following cerebral ischemia. Protein kinase B, also known as Akt/PKB, is part of an endogenous pathway that exerts effective neuroprotection from both necrotic and apoptotic cell death. Using a rat model of unilateral common carotid artery occlusion coupled with hypoxia, and using in vitro rat hippocampal slices, we examined the ability of adenosine to directly activate Akt/PKB. Western blot analysis revealed that levels of phosphorylated Akt/PKB were elevated in vivo under ischemic conditions in an adenosine A(1)-dependent manner and elevated in hippocampal slices treated with an adenosine A(1) agonist. We conclude from these studies that the activation of an adenosine A(1) receptor-mediated signal transduction pathway, either by endogenous adenosine (in vivo) or by an adenosine A(1) agonist (in vitro), results in the activation of the neurotrophic kinase Akt/PKB. PMID- 12133583 TI - Granulocyte macrophage-colony stimulating factor activates microglia in rat cortex organotypic brain slices. AB - Cytokines play an important role in the regulation of proliferation and migration in the central nervous system. The aim of this study was to determine if granulocyte macrophage-colony stimulating factor (GM-CSF) activates cells in the cortex of organotypic brain slice cultures. Our data show that murine GM-CSF markedly stimulated the proliferation and migration of small round microglia from a cortex slice. These round cells were strongly positive for integrin CD11b (OX 42), isolectin B4-lectin-binding, the monocytic marker ED1 and partly expressed major histocompatibility complex (MHC) class II antigen (OX-6). Only some differentiated microglia were visible which expressed the integrin CD11c and MHCII. GM-CSF enhanced the proliferation as analyzed by bromodeoxyuridine incorporation. The number of migrated cells decreased during culturing and enhanced terminal dUTP nick-end labelling positive nuclei were found. Taken together, our data conclude that GM-CSF is an important cytokine, which regulates the proliferation and migration of cortical microglia. PMID- 12133585 TI - Spatial summation of pain processing in the human brain as assessed by cerebral event related potentials. AB - To understand spatial summation of pain processing in the brain, we investigated the cerebral evoked responses to non-painful and painful contact heat stimulation (70 degrees C/s fast onset; intensity 2,4,6, corresponding to the individual's non-, slight and moderate pain) comparing one (1s) vs. two spots (2s) in 11 subjects while electroencephalographic signals were recorded. Significant spatial summation effects were shown only for the pain levels. For moderate pain, global field power examination isolated two peak activations for the vertex (Cz) N550 and P750 components. The single dipole modelling identified as likely the supplementary motor area, SMA area-6 source for N550, and posterior cingulate area-23 for P750. These source components showed a significantly faster (41.2 ms) latency and a shift in location from dorsal to ventral SMA of N550 toward cingulate area-31 between the 1s and 2s conditions. The temporal and spatial shift during spatial summation may reflect speeding up of the limbic affective reaction and prefrontal cognitive preparation in impending aversion and is deemed essential for integration of bodily sensations, such as pain. PMID- 12133584 TI - 1,2,3,4,6-Penta-O-galloyl-beta-D-glucose protects rat neuronal cells (Neuro 2A) from hydrogen peroxide-mediated cell death via the induction of heme oxygenase-1. AB - The root of Paeonia suffruticosa ANDREWS is an important Chinese crude drug used in many traditional prescriptions. 1,2,3,4,6-Penta-O-galloyl-beta-D-glucose (PGG), a major component of this crude drug, has been shown to possess potent anti-oxidant, anti-mutagenic and anti-proliferative effects. In the present study, we examined the effect of PGG on the expression of neuronal heme oxygenase 1 (HO-1), an inducible stress protein that degrades heme to the neuroactive molecule, carbon monoxide and the anti-oxidant, biliverdin. Exposure of Neuro 2A cells to PGG (10-50 microM) resulted in a concentration- and time-dependent induction of HO-1 mRNA, and protein expressions and heme oxygenase activity. Interestingly, pretreatment of the neuronal cells with PGG resulted in enhanced cellular resistance to hydrogen peroxide. This cytoprotective effect was reversed by zinc protoporphyrin IX, an inhibitor of heme oxygenase. This study showed that PGG could protect neuronal cells from oxidative stress via the induction of HO-1 gene expression. PMID- 12133586 TI - Alpha-2 macroglobulin I1000 V polymorphism in Chinese sporadic Alzheimer's disease and Parkinson's disease. AB - Several lines of evidence have revealed some overlapping pathologies in Alzheimer's disease (AD) and Parkinson's disease (PD). Although the alpha-2 macroglobulin gene (A2M) might be a risk factor of these two neurodegenerative diseases, conclusions from different studies have remained conflicting. Here we studied the role of A2M I1000 V polymorphism in both AD and PD in a Chinese Han population. We found that the A2M I/V genotype is associated with both AD (odds ratio (OR)=2.55, 95% confidential interval (95% CI): 1.20-5.43, attributable fraction (AF)=13.65%) and PD (OR=3.03, 95% CI: 1.30-7.02, AF=16.51%). After classifying according to the age of onset, this association is only detected in early-onset AD patients (OR=3.96, 95% CI: 1.28-12.26) and late-onset PD patients (OR=2.61, 95% CI: 0.97-7.09). Therefore, we conclude that in our samples, the A2M I/V genotype might be a susceptibility variant, even with minor effect, for both sporadic AD and PD. PMID- 12133587 TI - The cytosolic inclusion bodies that consist of splice variants that lack exon 5 of the presenilin-2 gene differ obviously from Hirano bodies observed in the brain from sporadic cases of Alzheimer's disease patients. AB - Alzheimer's disease (AD) is a neurodegenerative disorder having several pathological characteristics, namely senile plaques and neurofibrillary tangles. Further, Hirano bodies (HBs), which are stained by hematoxylin-eosin, are also observed in the AD brain. Recently, we reported that an alternative splice variant that lacked exon 5 of the presenilin-2 (PS2) gene (PS2V) was expressed in sporadic AD (SAD) brains, and the PS2V-encoding proteins actually existed in these brains. Furthermore, electron microscopic experiments demonstrated that the PS2V proteins form cytosolic inclusion bodies in the pyramidal cells. In this report, we found that the PS2V-composed inclusion bodies differed obviously from the HBs. This observation suggests the possibility that PS2V-composed inclusions are a novel inclusion body, compared with other pathological characteristics previously reported in the SAD brain. We named this inclusion body the 'PS2V body'. PMID- 12133588 TI - Possible involvement of tetrahydrobiopterin in the trophic effect of insulin-like growth factor-1 on rat pheochromocytoma-12 (PC12) cells. AB - Tetrahydrobiopterin (BH(4)) has a trophic effect on pheochromocytoma-12 (PC12) cells such as insulin-like growth factor-1 (IGF-1). We investigated involvement of BH(4) in the trophic effect of IGF-1 on PC12 cells. IGF-1 (10-300 ng/ml) increased cellular BH(4) content in a dose-related manner. Cellular BH(4) content increased after 6-36 h incubation with IGF-1. IGF-1-induced increase in the cellular BH(4) content was blunted by 0.3 mM 2,4-diamino-6-hydroxypyrimidine (DAHP), an inhibitor for BH(4) synthesis. IGF-1 protected PC12 cells from the cell death induced by depletion of serum and nerve growth factor, which was attenuated by DAHP. The effects of IGF-1 on the cellular BH(4) content and cell viability were eliminated by 0.2 microM wortmannin. These results suggest that BH(4) is involved in the trophic effect of IGF-1 on PC12 cells and that the effect of IGF-1 on BH(4) synthesis is mediated by phosphatidylinositol 3-kinase. PMID- 12133589 TI - Dissociated changes of somatosensory evoked low-frequency scalp responses and 600 Hz bursts after single-dose administration of lorazepam. AB - Electrical stimulation of upper limb nerves allows one to record two types of scalp responses, that is conventional low-frequency somatosensory evoked potentials (SEPs), and bursts of high-frequency (about 600 Hz) wavelets. To further clarify the functional meaning of both types of responses, we investigated whether changes of GABAergic drive could cause significant modifications of conventional as well as high-frequency SEPs. We recorded median nerve SEPs from six healthy volunteers before and after a single oral administration of lorazepam. In order to explain scalp SEP distribution before and after lorazepam administration, we performed the brain electrical source analysis of raw data. After lorazepam administration, conventional scalp SEPs showed a significant amplitude decrease of all cortical components including the primary N20/P20 response, while the subcortical P14 response remained substantially unchanged. Similarly, dipolar analysis showed a significant strength decrease of all cortical dipoles, whereas the strength of both subcortical dipoles (possibly located at the level of the brainstem and thalamus, respectively) remained unchanged. By contrast, no significant changes of high frequency SEPs were induced by drug intake. Therefore, our findings suggest that the inhibitory effect induced by lorazepam mainly affects intracortical circuitry. Tonic increase of the inhibitory drive, possibly mediated by GABAA receptors, can account for the reduced activity of first order deep spiny neurons generating the primary N20/P20. Conversely, intrinsic firing properties of the cell population generating high-frequency SEP responses are unaffected by the increase of GABAergic drive. This finding lends further substance to the hypothesis that conventional and high-frequency SEPs are generated by different cell populations. PMID- 12133590 TI - Astrocyte stellation in saline media lacking bicarbonate: possible relation to intracellular pH and tyrosine phosphorylation. AB - Primary cultures of astrocytes exhibit a polygonal morphology, but on treatment with agents that increase cAMP they change to stellate cells. We found that astrocyte stellation also occurred on replacing the culture medium with saline buffered with HEPES. However, stellation did not occur when the medium was replaced with saline buffered with bicarbonate/CO(2) provided Ca(2+) was present. Since exposure of astrocytes to media lacking bicarbonate results in a decrease in intracellular pH (pH(i)) we sought evidence for an association between pH(i) and morphology. Astrocytic pH(i) was monitored for 60 min after transferring the cells to HEPES or bicarbonate-buffered saline. HEPES-induced stellation was associated with transient acidification which coincided with the morphological changes. Acidification was not observed in cells transferred to bicarbonate saline. However when cytoplasmic acidification of cells in bicarbonate-saline was induced pharmacologically, rapid stellation occurred. Stellation induced by cAMP is reversed by activation of the RhoA pathway with lysophosphatidic acid (LPA). Here we found that LPA inhibited HEPES-induced stellation, but only with Ca(2+) present. Inhibition of stellation by LPA+Ca(2+) was associated with transient acidification followed by modest alkanization. A close association of tyrosine phosphorylation with stellation and pH(i) was observed. Thus incubation of astrocytes in HEPES-saline with orthovanadate to inhibit dephosphorylation abolished stellation and acidification; conversely incubation of cells in bicarbonate-saline with genistein to inhibit tyrosine kinases caused stellation and major acidification. Acidification may be one of several factors resulting in stellation, but it is not a necessary factor since stellation without acidification was observed in bicarbonate-saline lacking Ca(2+). PMID- 12133591 TI - Ischemia and failed regeneration in chronic experimental neuromas. AB - Neuromas are generally considered to be swollen uniform collections of uncontrolled aberrantly sprouting axons. In early experimental neuromas, there are substantial rises in local blood flow associated with their formation, but human studies of chronic lesions have suggested that neuromas develop ischemia and become impediments to regeneration. The issue is important because traumatically severed human nerves are frequently considered for repair some time after injury, when neuroma formation has occurred. In this work, we examined local perfusion, axon penetration and other characteristics of long-term (6 month) experimental neuromas created by sciatic nerve transection and resection of the distal sciatic nerve and its branches. The scenario was designed to model prior transection in a human nerve, where late surgical reconnection might be contemplated. Local blood flow in the extrinsic plexus of neuromas, examined using a laser Doppler flowmetry probe, declined in distal portions of the stump to values considerably lower than observed in intact nerves. Intrinsic blood flow near the stump tip, examined using microelectrode hydrogen clearance polarography was highly nonuniform and included zones with very low perfusion. Correlated with these findings were nonuniform histological features with zones of absent axons and blood vessels, progressive distal disorganization, marked declines in distal axon penetration, nonremodelled microfascicles and persistent expression of 'regenerative' axon and Schwann cell markers. Uncontrolled axon sprouting was not a feature. Longstanding neuromas include zones of relative ischemia and limited axon penetration that develop in the absence of nerve trunk reconnection. These features would limit their suitability for later repair. PMID- 12133592 TI - Changes in extracellular levels of amygdala amino acids in genetically fast and slow kindling rat strains. AB - A neurochemical basis for many of the epilepsies has long been suspected to result from an imbalance between excitatory and inhibitory neurotransmitter mechanisms. Data supporting changes in extrasynaptic amino acid levels during epileptogenesis, however, remain controversial. In the present study, we used in vivo microdialysis to measure the levels of extracellular GABA (gamma aminobutyric acid) and glutamate during seizure development in rats with a genetic predisposition for (Fast), or against (Slow), amygdala kindling. Dialysates were collected from both amygdalae before, during, and up to 12 min after a threshold-triggered amygdala afterdischarge (AD). One hour later, samples were again collected from both amygdalae in response to a hippocampal threshold AD. Daily amygdala kindling commenced the next day but without dialysis. After the rats were fully kindled, the same protocol was again employed. Amino acid levels were not consistently increased above baseline with triggered seizures in either strain. Instead, before kindling, a focal seizure in the Slow rats was associated with a large decrease in GABA in the non-stimulated amygdala, while amino acid levels in the Fast rats remained near baseline in both amygdalae. Similar results were seen after kindling. By contrast, before and after kindling, hippocampal stimulation caused large decreases in all amino acid levels in both amygdalae in both strains. These data suggest that, in response to direct stimulation, extracellular amino acid concentrations remain stable in tissues associated with either greater natural (Fast) or induced (kindled Fast/Slow) excitability, but are lowered with indirect stimulation (hippocampus) and/or low excitability. PMID- 12133593 TI - Inter-relationships between spinal cord blood flow, neuronal death and neurological function in rabbit spinal cord ischemia. AB - Following rabbit spinal cord ischemia we measured histology, neurological outcome and spinal cord blood flow to investigate relationships between these quantities over a range of ischemia durations. Rabbits underwent reversible spinal cord ischemia induced by temporary aortic artery occlusion for zero to 65 min. In one cohort, surviving spinal cord cells were counted after 4 days of reperfusion using computer image analysis (n=23). Neurological outcome (paraplegic or non paraplegic) was assessed in another group (n=381) after 18 h of reperfusion. A third group (n=8) had spinal cord blood flow measured by radioactive tracer immediately after ischemia. Histological analysis showed fewer motor neurons and astrocytes surviving as ischemia duration increased. Following 13 min (95% confidence interval (CI), 6 to 27) of ischemia 50% of low lumbar cord motor neurons remained alive; 50% of rabbits were paraplegic following 28.4 (26.7 to 30.0) min of ischemia; 28% (17% to 48%) of low lumbar motor neurons survived at this duration; 5% of rabbits were permanently paraplegic following 13.6 (10.6 to 16.1) min of ischemia, although only 52% (30% to 87%) of lower spinal cord motor neurons survived. Low lumbar cord blood flow was reduced to 2% of flow in non ischemic thoracic cord after 25 and 60 min of ischemia (0.52 and 0.59 ml/100 g/min, respectively). These data support a fundamental hypothesis on which experimental ischemia is based: that substantial neuronal death follows directly from reversible ischemia and leads to permanent neurological deficit. PMID- 12133594 TI - A comparative immunocytochemical study of development and regeneration of chemosensory neurons in the rat vomeronasal system. AB - Vomeronasal neurons undergo continuous neurogenesis during development and after neuronal injury. We used immunocytochemical methods to compare different stages of the vomeronasal organ development to those of regeneration following vomeronasal nerve transection. At E15 and at 6 to 10 days after injury, nestin positive cells were observed throughout the sensory epithelium. We did not find nestin immunoreactivity to be localized to the boundary region of the epithelium. The early appearance and wide distribution of nestin-positive cells suggests that they represent chemosensory precursor cells that develop and migrate vertically in the epithelium. Vomeronasal receptor cells degenerated 6 to 8 days after nerve transection, but axon terminals in the accessory olfactory bulb (AOB) continued to show the presence of the chemosensory specific marker (OMP) for up to ten days, a significant finding observed in this study. It is likely that the distance from the site of nerve transection may contribute to differences in the time course of anterograde and retrograde axon degradation. OMP-positive neurons were observed in the normal adult epithelium and to a much lesser extent 10-60 days after recovery from nerve transection. Axons from regenerated receptor cells did not reach the AOB during this time period. This failure to reestablish connections with target cells in the AOB could explain why OMP-positive cells were rarely observed among the regenerated cells in the vomeronasal epithelium. PMID- 12133595 TI - MT1 melatonin receptor mRNA expression exhibits a circadian variation in the rat suprachiasmatic nuclei. AB - The aim of the present study was to investigate the daily regulation of both MT1 and MT2 melatonin receptor subtype mRNA expression in the rat SCN in order to clarify their role in the daily variation of SCN melatonin receptors. Existing MT1 and MT2 partial clones were extended by PCR to 982 and 522 bp, respectively. However, while the MT1 clone allowed us to set up a highly sensitive in situ hybridization (ISH) method, we could not detect MT2 expression within the SCN. Therefore, our results suggest that only MT1 mRNA can be correlated with 2-iodo melatonin binding sites in the rat SCN. Investigation of MT1 mRNA expression throughout the 24 h light/dark cycle or in constant darkness clearly showed that in the two conditions, mRNA expression showed a robust rhythm with two peaks, one after the day/night and one after the night/day transitions in LD, and at the beginning of the subjective night and day in DD, respectively. Furthermore, these variations were not linked to the daily changes in melatonin receptor density. Thus, the transcriptional regulation of MT1 receptors does not appear to play a role in the daily regulation of melatonin binding sites availability. PMID- 12133596 TI - Effect of KW-7158, a putative afferent nerve inhibitor, on bladder and vesico vascular reflexes in rats. AB - The effects of KW-7158, a putative afferent nerve inhibitor, on reflex bladder activity and vesico-vascular reflexes were evaluated in urethane anesthetized SD rats with normal and xylene-irritated bladders. The bladder was filled with saline until the appearance of large amplitude spontaneous bladder contractions (LA-BC). Vesico-vascular reflexes were measured as increases in systolic arterial blood pressure during LA-BC or when the bladder was distended by a range of pressures. In normal rats, KW-7158 (10 and 100 microg/kg, i.v.) did not alter the amplitude or volume threshold for inducing LA-BC but increased the intercontraction interval. After xylene-irritation, which decreased volume threshold and intercontraction interval and induced small amplitude bladder contractions, KW-7158 increased volume threshold (65%) and intercontraction interval (150%) and decreased the number of small amplitude bladder contractions. Vesico-vascular reflexes induced during LA-BC or by bladder distension were suppressed (19.4-100%) by KW-7158. The effect of KW-7158 to depress vesico vascular reflexes as well as xylene-induced bladder hyperactivity without altering the amplitude of contractions is consistent with the view that the drug affects reflex bladder activity at least in part by depressing afferent pathways. PMID- 12133597 TI - Type II glucocorticoid receptors in the ovine hypothalamus: distribution, influence of estrogen and absence of co-localization with GnRH. AB - There is a strong association between the stress-induced increase in cortisol secretion and perturbation of the neuroendocrine reproductive axis. Previous studies implicate a neural target for glucocorticoids and it is possible that cortisol may act directly on gonadotropin releasing hormone (GnRH) neurons and, thus, luteinizing hormone release, through type II glucocorticoid receptors (GRs). In this study we investigated the effect of estradiol on GR immunoreactivity and determined whether GnRH neurons contain GRs. GRs were dispersed throughout most diencephalic structures but were most concentrated within the medial preoptic area and arcuate nucleus. GR cell numbers were significantly higher in these two areas in ewes pre-treated only with progesterone compared to ewes pre-treated with estradiol plus progesterone; there was no variation in the paraventricular nucleus between groups. No colocalization between GnRH and GRs was observed at any level of the brain. These results suggest that estrogen may down-regulate GRs and glucocorticoids do not act directly on GnRH neurons in the ewe. PMID- 12133598 TI - Platelet-activating factor increases prostaglandin E(2) release from astrocyte enriched cortical cell cultures. AB - The phospholipid mediator platelet-activating factor (PAF) increased the release of prostaglandin E(2) (PGE(2)) from astrocyte-enriched cortical cell cultures in a concentration- and time-dependent manner. The nonhydrolyzable PAF analog methylcarbamyl-PAF (mc-PAF), the PAF intermediate lyso-PAF, and arachidonic acid (AA) also produced this effect. In contrast, phosphatidlycholine (PC) and lyso PC, lipids that are structurally similar to PAF and lyso-PAF, had no effect on PGE(2) production, suggesting that PAF-induced PGE(2) release is not the consequence of nonspecific phospholipid-induced membrane perturbation. Antagonism of intracellular PAF binding sites completely abolished the ability of mc-PAF and lyso-PAF to mobilize PGE(2,) and attenuated the AA effect. Antagonism of the G protein-coupled PAF receptor in plasma membranes had no significant effect on mc PAF, lyso-PAF or AA-induced PGE(2) release. Based on the present findings, we hypothesize that intracellular PAF is a physiologic stimulus of PGE(2) production in astrocytes. PMID- 12133599 TI - GABAergic drugs alter hypothalamic serotonin release and lordosis in estrogen primed rats. AB - The effects of muscimol, a GABA(A) agonist, and phaclofen, a GABA(B) antagonist, on serotonin (5HT) release in the mediobasal hypothalamus and lordosis behavior were studied in freely moving rats using in vivo microdialysis. Two days after implantation of bilateral guide cannulae directed towards the ventromedial nucleus of the hypothalamus (VMH), ovariectomized rats were primed with estradiol (E(2)). The rats were implanted with microdialysis probes 24 h later. Following a pretest for lordosis, perfusate 5HT was measured at 20-min intervals until the baseline was stable. The rats were treated with 10, 30 or 100 microM muscimol or 30 and 100 microM phaclofen in artificial CSF delivered via reverse dialysis for 40 min. Control animals were continuously perfused with artificial CSF. Behavior was tested 20, 60 and 180 min after introduction of the drug. Decreased hypothalamic 5HT (40-60% of baseline) and marked facilitation of lordosis were present 20 min after administration of either drug. The effects of 10 and 30 microM muscimol and 30 microM phaclofen on both 5HT and lordosis were reversed after 180 min. Reversal of the behavioral and neurochemical effects were not evident in either the 100 microM muscimol or 100 microM phaclofen groups at the time-points tested. Proceptive responses were observed in phaclofen-treated rats but not in rats treated with muscimol. Levels of hypothalamic 5HT and lordosis quotients in control rats did not significantly differ from initial values. These results suggest that GABAergic effects on lordosis may be mediated through an interaction with 5HT in the mediobasal hypothalamus. PMID- 12133600 TI - Neurons evaluate both the amplitude and the meaning of signals. AB - The neuronal threshold, which can be determined by the level of depolarization immediately prior to spike generation, is different for responses to conditioned and discriminated stimuli after conditioning. However, it is impossible to determine excitability within a response to stimuli that failed to generate a spike. In the present study we examined the role of the AP threshold of two related Helix defensive neurons in the initiation of an AP during elaboration of the neuronal analog of a classical conditional reflex. These neurons fire in a synchronous manner though spikes sometimes are not generated simultaneously. We compared the change in spike threshold within the responses with other parameters of intracellular activity that also affect the neuronal response, but which can be measured without an analysis of the spike waveform (spike latency, slope of excitatory postsynaptic potentials, etc.). We found that the change in slope is not the reason for the change in the threshold during pairing. The thresholds within conditioned and discriminated responses affected spike generation and their values correlated with the spike presence or absence in related neurons. The excitability changed transiently within the responses, since these alterations were selective for the conditioned and discriminated stimuli and applied primarily to the first spike of the response. The firing threshold seems to be a dynamic property of excitable membrane. Neurons appear to evaluate the significance of the input signal, transiently change their own excitability and only after that compare the magnitude of this signal and threshold. PMID- 12133601 TI - Effects of AMPA-receptor and voltage-sensitive sodium channel blockade on high potassium-induced glutamate release and neuronal death in vivo. AB - High extracellular potassium induces spreading depression-like depolarizations and elevations of extracellular glutamate. Both occur in the penumbra of a focal ischemic infarct, and may be responsible for the spread of cell death from the infarct core to the penumbra. We have modeled this situation with microdialysis of an isotonic high-potassium solution into the normal rat amygdala for 70 min. This elevates extracellular glutamate up to 8-fold or more and produces irreversibly damaged, acidophilic neurons. NMDA-receptor blockade protects neurons and reduces the elevation of extracellular glutamate. Here we investigated the effects of sodium channel blockade with the voltage-sensitive sodium channel blocker tetrodotoxin and the AMPA receptor antagonist 2,3 dihydroxy-6-nitro-1,2,3,4-tetrahydrobenzo(f)quinoxaline-7-sulfonamide disodium (NBQX disodium) on high potassium-induced neuronal death and extracellular glutamate elevations. The acidophilic neurons produced are necrotic by ultrastructural examination. Tetrodotoxin, at dialysate concentrations of 33, 330 and 3300 microM (only a small fraction is extracted by tissue), markedly reduced the elevations of glutamate in rat amygdala at nearly all time points during high potassium perfusion, but it reduced tissue edema only at the highest concentration, and it was neuroprotective only if dialyzed prior to high potassium microdialysis (at 330 microM concentration). Although both 250 microM (6.2% is extracted by tissue) and 500 microM NBQX reduced elevations of glutamate, neither was neuroprotective, and neuropil edema was not reduced by either concentration. Our results suggest that in vivo, sodium influx through voltage-sensitive sodium channels but not through ligand-gated AMPA receptor channels contributes to high potassium-induced neuronal necrosis. PMID- 12133603 TI - Different effects of eNOS and nNOS inhibition on transient forebrain ischemia. AB - To clarify the functions of nitric oxide (NO) induced by either neuronal NO synthase (nNOS) or endothelial NO synthase (eNOS) after transient cerebral ischemia, we investigated the effects of L-N(5)-(1-iminoethyl)ornithine (L-NIO), a relatively selective eNOS inhibitor, and 7-nitroindazole (7-NI), a relatively selective nNOS inhibitor, on hippocampal dysfunction caused by cerebral ischemia. We measured mean arterial blood pressure (MABP), hippocampal blood flow, direct current (DC) potential, CA1 population spike (PS) and extracellular concentrations of glutamate from rat hippocampus after transient forebrain ischemia, which was induced by four-vessel occlusion for 10 min. L-NIO (20 mg/kg) and 7-NI (25 mg/kg) were administered intraperitoneally 20 min before ischemia. L NIO, but not 7-NI, increased MABP before, during and after ischemia, compared with the vehicle group. 7-NI, but not L-NIO, reduced the amplitude of anoxic depolarization induced by ischemia. 7-NI recovered the PS amplitude in part 60 min after ischemia. 7-NI, but not L-NIO, reduced the ischemia-induced levels of glutamate. These results indicate that nNOS inhibition with 7-NI improves, at least in part, hippocampal dysfunction after ischemia, while eNOS inhibition with L-NIO worsens it. PMID- 12133602 TI - Early biochemical and histological changes during hyperbaric or normobaric reoxygenation after in vitro ischaemia in primary corticoencephalic cell cultures of rats. AB - In a first series of experiments, the morphological changes of corticoencephalic cells by ischaemia were determined by staining with celestine blue-acid fuchsin in order to classify cells as intact, dark basophilic (supposedly reversibly injured) and preacidophilic or acidophilic (profoundly injured). Hypoxia and glucose-deprivation (in vitro ischaemia) markedly decreased the number of intact cells and correspondingly increased the number of both reversibly and profoundly damaged cells. The morphological characteristics indicated a partial recovery during reoxygenation either in the absence or presence of glucose and irrespective of whether normobaric or hyperbaric oxygen was used. In a second series of experiments, nucleoside triphosphate and diphosphate levels were determined in corticoencephalic cultures by high-performance liquid chromatography. Hypoxia in combination with glucose-deficiency markedly decreased the ATP:ADP, GTP:GDP and UTP:UDP ratios. A still larger fall of these ratios was observed both after normobaric and hyperbaric reoxygenation. In contrast, both normobaric and hyperbaric reoxygenation in the presence of glucose led to an almost complete recovery near the control normoxic values. In conclusion, the histological changes were not adequately reflected by changes in the nucleoside triphosphate:diphosphate ratios and, in addition, hyperbaric oxygen had neither favourable nor unfavourable effects on the early morphological and functional restitution of ischaemically damaged cells under the conditions of the present study. PMID- 12133604 TI - Nicotine compensates for the loss of cholinergic function to enhance long-term potentiation induction. AB - Long-term potentiation (LTP) in the hippocampal CA1 region is widely regarded to be the cellular substrate of learning and memory, and its induction critically depends on the activation of N-methyl-D-aspartate receptors (NMDARs). Nicotine reverses memory deficits caused by a lesion of the cholinergic system in animals. The mechanisms underlying this effect and the effect of nicotine on LTP after cholinergic degeneration are unknown. Here we show that cholinergic lesions impaired the induction of LTP, and nicotine reversed this effect and promoted the induction of LTP. The compensatory action of nicotine appears to be due to the enhancement of NMDAR responses mediated by nicotine-induced disinhibition of pyramidal cells. This may represent the cellular basis of nicotine-mediated cognitive enhancement observed in the presence of cholinergic deficits. PMID- 12133605 TI - A review of spectroscopic methods for characterizing microbial transformations of minerals. AB - Over the past decade, advances in surface-sensitive spectroscopic techniques have provided the opportunity to identify many new microbiologically mediated biogeochemical processes. Although a number of surface spectroscopic techniques require samples to be dehydrated, which precludes real-time measurement of biotransformations and generate solid phase artifacts, some now offer the opportunity to either isolate a hydrated sample within an ultrahigh vacuum during analysis or utilize sources of radiation that efficiently penetrate hydrated specimens. Other nondestructive surface spectroscopic techniques permit determination of the influence of microbiological processes on the kinetics and thermodynamics of geochemical reactions. The ability to perform surface chemical analyses at micrometer and nanometer scales has led to the realization that bacterial cell surfaces are active sites of mineral nucleation and propagation, resulting in the formation of both stable and transient small-scale surface chemical heterogeneities. Some surface spectroscopic instrumentation is now being modified for use in the field to permit researchers to evaluate mineral biotransformations under in situ conditions. Surface spectroscopic techniques are thus offering a variety of opportunities to yield new information on the way in which microorganisms have influenced geochemical processes on Earth over the last 4 billion years. PMID- 12133606 TI - Immunomagnetic separation of scum-forming bacteria using polyclonal antibody that recognizes mycolic acids. AB - Mycolic acid-containing bacteria (mycolata) are thought to be involved in scum formation in aeration basins of activated sludge plants due to their ability to produce biosurfactants and their cell surface hydrophobicity. To isolate these bacteria, immunomagnetic separation (IMS) using an anti-mycolic acid polyclonal antibody was investigated. IMS that targeted Gordonia amarae SC1 exhibited a 100% recovery at 5x10(3) CFU ml(-1). At cell concentration of 7.8x10(6) CFU ml(-1), the recovery was lowered, but 80% of cells were still captured. Effect of bead concentrations on the recovery of SC1 at 10(6) CFU ml(-1) was examined. The results showed that addition of more than 6-7x10(6) beads for 1x10(6) CFU reached a maximum recovery (83%). Furthermore, the IMS procedure optimized with SC1 cells was tested with another mycolata. The results suggested that variation of the recovery for each mycolata is dependent on the specificity of the polyclonal antibody and that mycolata which are recognized by the antibody can be recovered by this procedure. PMID- 12133607 TI - Fluorescence-assisted image analysis of freshwater microalgae. AB - We exploit a property of microalgae-that of their ability to autofluoresce when exposed to epifluorescence illumination-to tackle the problem of detecting and analysing microalgae in sediment samples containing complex scenes. We have added fluorescence excitation to the hardware portion of our microalgae image processing system. We quantitatively measured 120 characteristics of each object detected through fluorescence excitation, and used an optimized subset of these characteristics for later automated analysis and species classification. All specimens used for training and testing our system came from natural populations found in Lake Biwa, Japan. Without the use of fluorescence excitation, automated analysis of images containing algae specimens in sediment is near impossible. We also used fluorescence imaging to target microalgae in water samples containing large numbers of obtrusive nontargeted objects, which would otherwise slow processing speed and decrease species analysis and classification accuracy. Object drift problems associated with the necessity to use both a fluorescence and greyscale image of each microscope scene were solved using techniques such as template matching and a novel form of automated seeded region growing (SRG). Our system proved to be not only user-friendly, but also highly accurate in classifying two major genera of microalgae found in Lake Biwa-the cyanobacteria Anabaena spp. and Microcystis spp. Classification accuracy was measured to be over 97%. PMID- 12133608 TI - Optimisation of one-tube PCR-ELISA to detect femtogram amounts of genomic DNA. AB - A simple, high-throughput, low-cost polymerase chain reaction-enzyme linked immunosorbent assay (PCR-ELISA) protocol that detects the presence of 4 fg of DNA from four bacterial fish pathogens Yersinia ruckeri, Tenacibaculum maritimum (formerly Flexibacter maritimus), Lactococcus garvieae and Aeromonas salmonicida was developed. DNA amplification was undertaken in a biphasic system with free and bound PCR that are achieved in the one NucleoLink tube. Solid-phase amplicons were detected using biotin labelled hybridization probes and visualised colourimetrically with streptavidin-alkaline phosphatase and p nitrophenylphosphate as substrate. PCR and hybridization took less than 8 h to perform with maximum signal output for femtogram amounts of template DNA achieved within 24 h. Implementation and optimization of the protocol is discussed. PMID- 12133609 TI - RNA isolation from soil for bacterial community and functional analysis: evaluation of different extraction and soil conservation protocols. AB - The impact of three different RNA isolation methods on the community analysis of metabolically active bacteria was determined by reverse transcription (RT) and PCR amplification of 16S rRNA genes and subsequent terminal restriction fragment length polymorphism (T-RFLP) analysis. Furthermore, soil samples were stored at different conditions in order to evaluate the effect of soil conservation methods on the outcome of the population analysis. The quality of mRNA was assessed by reverse transcription and PCR amplification of eubacterial glutamine synthetase genes. Our results indicated that the community composition as well as the abundance of individual members were affected by the kind of RNA isolation method. Furthermore, the extraction method influenced the recovery of mRNA. Lyophilization, storage at -20 degrees C as well as storage in glycerol stocks at -80 degrees C proved to be equally appropriate for the storage of soils and subsequent RNA isolation. PMID- 12133611 TI - Efficient isolation of total RNA from antibiotic-producing bacterium Amycolatopsis mediterranei. AB - RNA extraction from antibiotic-producing actinomycetes can be a difficult and time-consuming process due to their special peptidoglycans cell wall composition and the short life of RNA. Hence, the rapidity of cellular lysis and complete inhibition of RNase are of particular importance for isolating intact RNA of high quality. The genus of Amycolatopsis mediterranei produces many clinically important antibiotics, such as rifamycin and vancomycin; however, the available methods for bacterial RNA isolation did not work very well with this genus. In this report, we described a new method for RNA isolation using the combination of LiCl, urea and guanidinium thiocyanate to disrupt the cell wall of Amycolatopsis. Compared with earlier published RNA isolation methods, the method gave higher yields of pure and intact RNA. About 1 microg total RNA free of DNA contamination can be obtained from 1 mg wet weight of A. mediterranei. The integrity of the RNA was demonstrated by formaldehyde agarose gel electrophoresis and Northern blot analyses. PMID- 12133610 TI - Use of both 16S rRNA and engineered functional genes with real-time PCR to quantify an engineered, PCB-degrading Rhodococcus in soil. AB - A real-time PCR (RTm-PCR) assay using fluorescently labeled oligonucleotides (TaqMan probes) was used to detect and quantify the recombinant Rhodococcus sp. strain RHA1(fcb) in soil. One primer and probe set targeted a hypervariable region of the 16S rRNA gene unique to strain RHA1(fcb) and its phylogenetic relatives, and the other set targeted the recombinant 4-chlorobenzoate (4-CBA) degradation operon (fcb) and was strain-specific. The method had a 6-log dynamic range of detection (10(2)-10(7) cells ml(-1)) for both probes when DNA from pure cultures was used. Although the method was less sensitive in soil, the estimated number of cells in soil by real-time PCR corresponded to the measured number of RHA1(fcb) cells determined by colony-forming units. PMID- 12133612 TI - Localisation of mycobacterial DNA and mRNA in human tuberculous granulomas. AB - In situ hybridisation was used to detect the presence of Mycobacterium tuberculosis in paraffin-embedded lung tissue of nine patients diagnosed with tuberculosis (TB). Mycobacterial DNA was found in all nine patients and in 175 out of 191 granulomas examined. A combination of in situ hybridisation and immunohistochemistry techniques demonstrated that mycobacterial DNA was associated with CD68-positive cells with the morphology of macrophages and giant cells. Mycobacterial DNA was also found within the necrotic regions of some granulomas. mRNA for the mycobacterial RNA polymerase beta subunit (rpoB) was detected by RNA: RNA in situ hybridisation. The rpoB mRNA was also localised to CD68-positive cells with the morphology of macrophages and to giant cells of certain necrotic granulomas. No rpoB mRNA was found in the necrotic regions of granulomas. Mycobacterial DNA was detected in 92% of patient granulomas of which 8% were positive for rpoB mRNA. The ability to identify mycobacterial RNA transcripts within human tuberculous granulomas affords us the opportunity to analyse the interplay between pathogen gene expression and the human immune response and should provide valuable insight into the mechanisms used by M. tuberculosis to persist within the human host. PMID- 12133613 TI - Rapid sample processing and fast gas chromatography for identification of bacteria by fatty acid analysis. AB - A commercially available system for microbial identification by fatty acid analysis (Microbial Identification System (MIS), MIDI, Newark, DE, USA) requires a four-step sample derivatization procedure in screw-cap test tubes. By using glass tubes in a 96-well format with multichannel pipetting, the time required for sample preparation can be greatly reduced. The standard gas chromatography column, 25 m long by 0.20 mm ID, is replaced with a 10 m long by 0.10 mm ID column, reducing the gas chromatography run time to one third of the standard time. Either or both of these procedures can be easily implemented in any laboratory using the MIDI system, resulting in faster identifications and higher sample throughput. PMID- 12133614 TI - A probabilistic neural network approach for modeling and classification of bacterial growth/no-growth data. AB - In this paper, we propose to use probabilistic neural networks (PNNs) for classification of bacterial growth/no-growth data and modeling the probability of growth. The PNN approach combines both Bayes theorem of conditional probability and Parzen's method for estimating the probability density functions of the random variables. Unlike other neural network training paradigms, PNNs are characterized by high training speed and their ability to produce confidence levels for their classification decision. As a practical application of the proposed approach, PNNs were investigated for their ability in classification of growth/no-growth state of a pathogenic Escherichia coli R31 in response to temperature and water activity. A comparison with the most frequently used traditional statistical method based on logistic regression and multilayer feedforward artificial neural network (MFANN) trained by error backpropagation was also carried out. The PNN-based models were found to outperform linear and nonlinear logistic regression and MFANN in both the classification accuracy and ease by which PNN-based models are developed. PMID- 12133615 TI - Enumerating ammonia-oxidizing bacteria in environmental samples using competitive PCR. AB - Primers targeting part of the ammonia-monooxygenase gene (amoA) have been used to detect and characterize ammonia-oxidizing bacteria (AOB) in different environments. In this study, a quantitative polymerase chain reaction (PCR) technique using a competitive template for the amoA primer pair is described and evaluated. The method is based on addition of an internal standard to the PCR, a competitive template, which is amplified together with the template in the environmental sample. By adding different amounts of competitive template to the sample and observing the relative intensity of environmental amplificate and competitive amplificate, the number of amoA gene copies can be determined. Different tests were made to evaluate the competitive PCR method (cPCR) with respect to equal amplification efficiency of the two templates, degeneracy of the priming site and the importance of flanking regions surrounding the competitive template. Calibration curves made by addition of known amounts of Nitrosomonas europaea to soil samples revealed a detection limit for this technique of less than 1000 cells g(-1) soil and a linear response over a wide range of cell additions. Cloning and sequencing of amoA amplificates have confirmed the specificity of the primers, as we have not detected any false positives among the more than 200 clones investigated. The vertical distribution of ammonia-oxidizers in the upper cm of a waterlogged rice paddy soil was compared to nitrate and oxygen concentration profiles determined with microsensors and to net process rates derived from these profiles. PMID- 12133616 TI - Primary fecal culture used as template for PCR detection of diarrheagenic E. coli virulence factors. AB - The PCR technique applied to primary fecal cultures (PFC-PCR) was compared to the usual method employing isolated colonies (IC-PCR) in order to assess its sensitivity in the detection of virulence markers of diarrheagenic Escherichia coli in fecal samples obtained from children with diarrhea. Among the 149 samples analysed, PFC-PCR detected 81(54.4%) samples presenting one or two virulence markers, while IC-PCR detected only 59 (39.6%) positive samples. The markers detected in order of frequency were: pAA, LT-I, eaeA, ST-I, daaE, and ipaH. The PFC-PCR method of detection of diarrheagenic E. coli virulence markers proved to be reliable and more sensitive (p<0.05) than the usual method employing isolated colonies. It has also the advantage of being faster and less expensive than the detection methods in current use. PMID- 12133617 TI - Development of a rapid, simple dipstick dye immunoassay for schistosomiasis diagnosis. AB - In order to develop a rapid, simple immunodiagnostic assay for schistosomiasis, soluble egg antigen (SEA) of Schistosoma japonicum was conjugated with a blue colloidal dye (D-1) produced in China and used to detect antibodies in the sera of schistosomiasis patients. The antigen-antibody complex was captured by anti human IgG absorbed onto a nitrocellulose membrane dipstick by means of immunochromatography. The results showed that the sensitivity of the dipstick dye immunoassay (DDIA) was 96.7% in 30 cases of acute schistosomiasis (29/30) and 94.1% (79/84) in 84 cases of chronic schistosomiasis. The specificity of the assay was 96.7% in 60 healthy subjects. Cross-reactions were observed in 10.0% of 20 cases of clonorchiasis and in 70.0% of 20 cases of paragonimiasis. The results were similar to those detected by routine ELISA. In a field evaluation of the DDIA kit, the positive rate of the DDIA was 96.7% in 121 cases of schistosomiasis, compared with 90.1% with the circumoval precipitin test (COPT). The antigen conjugated with dye was stable at room temperature for at least 6 months. The results indicated that the dipstick dye immunoassay provided high sensitivity and good specificity for the detection of schistosomiasis and the assay was rapid, simple and cheap, and did not need any equipment. It was more useful for screening target populations for selective chemotherapy than other immunoassays. PMID- 12133618 TI - Development and application of fully functional epitope-tagged forms of transforming growth factor-beta. AB - Administration of transforming growth factor-beta (TGF-beta) has been found to be of therapeutic benefit in various mouse disease models and has potential clinical usefulness. However, the ability to track the distribution of exogenously administered, recombinant forms of these proteins has been restricted by cross reactivity with endogenous TGF-beta and related TGF-beta isoforms. We describe novel FLAG- and hemagglutinin (HA)-tagged versions of mature TGF-beta1 that retain full biological activity as demonstrated by their ability to inhibit the growth of Mv1Lu epithelial cells, and to induce phosphorylation of the TGF-beta signaling intermediate, smad 2. Intracellular FLAG- and HA-TGF-beta1 can be detected in transfected cells by confocal immunofluorescence microscopy. We also describe sandwich ELISAs designed to specifically detect epitope-tagged TGF-beta and demonstrate the utility of these tagged ligands as probes for TGF-beta receptor expression by flow cytometry. The design of these fully functional epitope-tagged TGF-beta proteins should facilitate studies such as the evaluation of in vivo peptide pharmacodynamics and trafficking of TGF-beta ligand-receptor complexes. PMID- 12133619 TI - Stabilised cellular immuno-fluorescence assay: CD45 expression as a calibration standard for human leukocytes. AB - We first confirmed the precision of the quantitative indirect immunofluorescence (QIFI) test by demonstrating that blood lymphocytes from different individuals expressed CD45 leucocyte antigens at a very similar level (mean: 201 x 10(3) antibody binding capacity (ABC)/lymphocyte) with only minimal variation (CV% 2.5%). These values were maintained for 4 days in blood samples when kept at 20 degrees C, and for up to 14 days in samples fixed with TransFix. Using long-term stabilisation, after an initial drop of 10-15% the CD45 ABC/lymphocyte values remained stable at an 85-90% level for >1 year. These biological standards were used to check other quantitative IF techniques. The quantum simply cellular (QSC) method showed variable results (85-240%), and the QuantiBRITE method gave values as low as 30-40% of the expected values, indicating that biological standards such as CD45 ABC/lymphocytes are absolutely essential to check the performance of methods that claim to quantify immunofluorescence (IF). Next, these standards were used to establish the stabilised cellular immuno-fluorescence assay (SCIFA) as follows. The ABC x 10(3)/cell values established by QIFI on leucocyte populations such as lympho-, mono- and granulocytes were used to create calibration curves for the CD45 antigen and its isoforms CD45RA, -R0 and -RB. The same cell populations were then stained with monoclonal antibodies (MAbs) directly conjugated to different fluorochromes in order to translate the mean fluorescence intensity (MFI) values seen with the directly labelled reagents to values of ABCx10(3)/cells. Using SCIFA with a triple-colour direct IF, the display of CD45 and its isoforms were quantitated on the 'virgin' or 'unprimed' (CD45RA+) and 'primed' (CD45R0+) subsets in both the CD4+ and CD8+ T cell lineages. We also observed that the CD4+ and CD8+ T cells in transit between the 'virgin' and 'primed' subsets frequently displayed different levels of the CD45RA and -R0 molecules, pointing to the physiological variability of the CD45RA-R0 switching process. In conclusion, internal biological standards, with known stable expression of ABC/cell, should be used to evaluate IF staining patterns in a quantitative manner during routine investigations. PMID- 12133620 TI - Surface plasmon resonance (BIACORE) detection of serum antibodies against Salmonella enteritidis and Salmonella typhimurium. AB - We have used a surface plasmon resonance biosensor (BIACORE 3000) to detect serum antibodies in chickens having current or recent infections. Three well-defined Salmonella flagellar recombinant DNA antigens reflecting Salmonella enteritidis (H:g,m flagellin) and Salmonella typhimurium (H:i and H:1,2 flagellins) expressed in Escherichia coli were each immobilized in a single flow cell of a biosensor chip. Glutathione-S-transferase was immobilized on the surface of another flow cell to monitor non-specific binding. Sera collected from chickens with no history of Salmonella infection, and from chickens infected with Salmonella serotypes infantis, pullorum, gallinarum were used to test the performance of the system. The sensitivity exhibited to a range up to 900 arbitrary response units (RU) for the most positive S. typhimurium serum at a dilution of 1/40. Sera from Salmonella infantis, Salmonella pullorum and Salmonella gallinarum infected birds gave responses less than the cut-off point, which was determined as the averaged response of sera from specific pathogen-free chickens plus three times the standard deviation. A positive response was obtained when these sera and whole blood were fortified with S. enteritidis and S. typhimurium positive serum. The sensitivity, specificity, precision and reproducibility obtained suggested that this approach could be used for detecting past or present infection with a range of pathogens in animals. PMID- 12133621 TI - Characterization of antibody affinities using an AGE-modified dipeptide spot library. AB - Recent immunological approaches have greatly helped understanding the significance of advanced glycation end products (AGEs) in age-related diseases. However, immunohistochemical localization of AGEs in tissues or their quantitative determination in biological fluids sometimes yields inconsistent results among different investigators. Since these differences might be caused by the heterogeneity of the AGE antibodies, a systematic mapping of their epitope recognition pattern would help in the evaluation of these different results. For this purpose, an N-terminally acetylated combinatorial dipeptide library with 400 positionally defined dipeptides was modified by glucose to yield AGEs, which are thought to be present on the protein side chains. Using this library, we have characterized six different AGE antibodies in respect to their immunoreactivity towards AGE-modified dipeptides. All antibodies predominantly recognized only one AGE-modified amino acid side chain (and not a particular dipeptide). In most cases, arginine- and lysine- derived AGEs, but also some epitopes on asparagine and on heterocyclic amino acids were recognized. Very interestingly, the immunization of different animals with the same antigen produced AGE antibodies with a totally different epitope recognition pattern. Defining the antigen recognition pattern with this method might help in the quality control of polyclonal AGE antibodies for immunodetection. In addition, defined AGE antibodies (as neutralizing antibodies) could also help to define "signal-active" AGEs in terms of specific AGE-mediated cellular responses. PMID- 12133622 TI - Synthesis and characterization of biologically functional biotinylated RANTES. AB - Development of specifically labeled chemokines that retain their biological properties should be useful for analyzing their mechanisms of action both under physiological and pathological conditions. Here, we report the chemical synthesis and characterization of RANTES (regulated upon activation normal T cell expressed and secreted) derivatives that were biotinylated at residues 1, 25, 33, 45, or 67. Gel filtration and ultracentrifugation experiments showed that biotinylation at position 45 or 67 decreased the aggregation tendency of the chemokine to a dimeric state. Competition experiments, using a stably transfected CHO-K1 cell line overexpressing human CCR5, a RANTES receptor, indicated that derivatives biotinylated at positions 1, 25, and 67 bound to CCR5 with the same affinity as native RANTES. Flow cytometry analysis showed that RANTES biotinylated at residue 67 (B67-RANTES) bound more efficiently to primary macrophages than the other derivatives. Such binding was dependent on cell surface glycosaminoglycans (GAGs) since it was reduced when macrophages or HeLa cells expressing or not CCR5 were first treated with GAG-specific enzymes. In addition, B67-RANTES modulated CCR5 expression on lymphocytes and elicited chemotaxis of monocytes in the same manner as unmodified RANTES. Thus, B67-RANTES acts as a CCR5 agonist and may be useful to study the role of RANTES in pathologies such as, for example, human immunodeficiency virus (HIV) infection and inflammatory disorders. PMID- 12133623 TI - Epitopic characterization of native bovine beta-lactoglobulin. AB - Two monoclonal antibodies (mAbs) (mAb 97 and mAb 117) selected from a panel of 52 mAbs directed against beta-lactoglobulin (BLG) have previously been used to develop a two-site enzyme immunometric assay (EIA) specific for the native form of the protein [J. Immunol. Methods 220 (1998) 25]. In the present work, the conformational epitopes recognized by these two mAbs and by the 50 others have been studied. Firstly, an epitope map was drawn using a surface plasmon resonance (SPR) biosensor: the epitopes were organized in a circle of 11 overlapping and 1 nonoverlapping antigenic regions. Secondly, 55 site-directed BLGA mutants were prepared and tested by ELISA and competitive immunoassay to localize these 12 antigenic regions on the protein molecule. Among them, 20 mutants showed a 10- to 7500-fold decrease in relative affinity for the mAbs of one or several neighbouring regions: their circular dichroism (CD) spectra were identical to the spectrum of wild-type (WT) BLGA. At least one mutant was found for each of the 11 overlapping antigenic regions which circled the molecule and for the nonoverlapping one which was localized near the entrance of the calyx. The two mAbs initially chosen were each directed towards very conformation-dependent epitopes and were thus suitable for monitoring native BLG in food products and manufacturing processes. Other mAb pairs could be used to follow the fate of specific regions of the molecule during denaturation or proteolytic digestion. PMID- 12133624 TI - A new strategy for the diagnosis of MAGE-expressing cancers. AB - The expression of melanoma antigen gene (MAGE), coding for tumor antigens recognized by cytotoxic T cell, is highly specific to cancer cells, but their use in the detection of a few cancer cells by reverse transcription-polymerase chain reaction (RT-PCR) has been limited by the low frequency of expression of individual MAGE genes. In order to increase MAGE detection rate in RT-PCR assay, here, we designed multiple MAGEs recognizing primers (MMRPs) that can bind to the sequences of cDNA of MAGE-1, -2, -3, -4a, -4b, -5a, -5b and-6 (MAGE 1-6) together. The nested RT-PCR assay using MMRPs, MAGE 1-6 assay, detected MAGE messages of 1 to 5 SNU484 cells in a background of 10(7) SNU638 cells. MAGE detection rate of MAGE 1-6 assay in cancers was higher than that of nested RT-PCR that detects single MAGE gene expression. The expressions of MAGE genes was detected by MAGE 1-6 assay in 70.4% (19/27) of head and neck cancer tissues, 91.7% (11/12) of breast cancer tissues, 75% (9/12) of lung cancer tissues. However, they were not detected in 18 benign lesions and 20 normal head and neck tissues and 30 blood samples from healthy donor. In conclusions, MAGE 1-6 assay can detect any cancer cells that express at least one of eight MAGE subtype genes, and this method may be very useful for the diagnosis of MAGE-expressing cancers. PMID- 12133625 TI - Quantitative real-time RT-PCR as a method for monitoring T lymphocyte reactivity to full-length tyrosinase protein in vaccinated melanoma patients. AB - The major goal of therapeutic cancer vaccine trials is to mediate tumor regression. However, it is critically important to devise in vitro immunological assays that correlate with clinical outcome, for use as surrogate markers of vaccine efficacy. To date, clinical emphasis has been placed on peptide vaccines, but trends towards the use of more complex immunogens such as whole proteins require the development of efficient and sensitive methods for monitoring their immunological effects. In the context of a vaccination trial using full-length tyrosinase (Ty) to immunize patients with metastatic melanoma, a monitoring technique was developed in which autologous dendritic cells (DC) infected with a recombinant adenovirus encoding the Ty protein were used to assess the Ty specific reactivity of fresh peripheral blood lymphocytes (PBL) collected from patients at different intervals during therapy. Quantitative real-time RT-PCR (qRT-PCR) was used to measure the production of cytokine mRNA by T cells following a 2.5-h incubation with Ty-expressing DC. Two out of ten patients studied demonstrated Ty protein-specific reactivity that increased during and after the period of vaccination. While one of these patients also reacted to an HLA-A1-compatible Ty peptide, the second did not recognize any of the known Ty epitopes, highlighting the importance of this technique for monitoring the effects of complex vaccines. PMID- 12133627 TI - Stability of total and rubella-specific IgG in oral fluid samples: the effect of time and temperature. AB - Oral fluid (saliva) samples are increasingly used as a minimally invasive alternative to serum to detect antibody for viral diagnostics and population based surveys of immunity to common infections. Postal collection of these samples is convenient, but it is uncertain how oral fluid antibody levels may deteriorate during transit. In this study multiple oral fluid samples, from a group of individuals, were collected and kept at two different temperatures (10 and 20 degrees C, simulating winter and summer conditions ) for periods of up to 7 days. They were then tested for total immunoglobulin G (IgG) and for rubella specific IgG. Total IgG concentrations ranged from 2.0 to 48.6 mg/l but showed no significant decrease over the 7-day period. There was also no significant change in rubella-specific IgG measured using an antibody capture ELISA, but testing with an indirect rubella IgG ELISA revealed a significant decrease in absorbance values between day 1 and day 7 at 20 degrees C. The difference in the rubella assay results may reflect the different assay formats with the signal in the indirect procedure related to the concentration of rubella IgG in the oral fluid sample, rather than the proportion of virus-specific IgG measured with the antibody capture format. The study demonstrated that total and rubella specific IgG in oral fluid samples were stable for up to 7 days at differing temperatures and that an antibody capture assay format should be used for virus-specific testing to minimise inaccurate results due to low IgG concentrations in oral fluid samples. PMID- 12133626 TI - Differentiation of single populations in a bidirectional mixed lymphocyte culture using X and Y chromosome-specific FiSH markers. AB - OBJECTIVE: To validate a new combination of a viability assay and Fluorescence in Situ Hybridsation (FiSH) techniques using X- or Y-DNA probes to assay the viability of each single population within bidirectional mixed lymphocyte cultures (MLCs). STUDY DESIGN: We determined the absolute viable lymphocyte counts of each population in bidirectional maternal-fetal MLCs as well as in bidirectional MLCs between male and female unrelated adults using a combination of a viability assay and additional sex discrimination by FiSH, and compared the results with the classical tritiated thymidine incorporation assay and with sex discrimination by metaphase number. RESULTS: The results of the total viability assay correlated with those of the tritiated thymidine incorporation assay. The differentiation of each single population by FiSH showed, as in the metaphase assay, that the relationship between viable maternal and neonatal cells was shifted significantly (p<0.05) towards the neonatal cells, while the relationship of the viable cells in the MLCs between unrelated adults was balanced. CONCLUSION: Our results show that X/Y discrimination by FiSH would be a useful tool for analyzing the immunologic network by bidirectional MLCs. The main advantage over sex discrimination by metaphase counting is that not only the proliferating cells but all viable cells are taken into account. An additional aspect would be the possibility to sort the cells according to surface markers (e.g. CD8+ HLA DR+) before discrimination by X and Y chromosomes. PMID- 12133628 TI - Development of detection methods for ruminant interleukin (IL)-12. AB - Recombinant bovine IL-12 (rbo IL-12) was transiently expressed in COS-7 cells and shown to upregulate the synthesis of IFNgamma by bovine cells stimulated with a suboptimal concentration of mitogen in vitro. Mice were immunised with a plasmid encoding rbo IL-12 and boosted with rbo IL-12 and a number of monoclonal antibodies (mAb) were generated that reacted with rbo IL-12 in an ELISA. Some of these mAb neutralised the ability of rbo IL-12 to induce IFNgamma synthesis by bovine cells. A pair of mAb was identified that together could be used to detect both recombinant and natural bovine IL-12 by ELISA and a luminometric detection method was applied to the ELISA making it more sensitive. Using this method native bovine IL-12 was detected in supernatants of dendritic cells (DC) cultured in vitro with a synthetic lipopeptide known to stimulate secretion of IL-12 by human DC. The ELISA was also able to detect recombinant ovine IL-12 and, less effectively, recombinant human IL-12. In contrast, bovine IL-12 was not detected by a commercial human IL-12 ELISA kit. Intracytoplasmic IL-12 was detected in bovine DC using the antibodies described herein. The ability to detect ruminant IL-12 by three methods: ELISA, bioassay with neutralising mAb and cytoplasmic staining, will permit studies of the role of this important cytokine in the immunology and pathogenesis of animal diseases. PMID- 12133629 TI - Heat-mediated, ultra-rapid electrophoretic transfer of high and low molecular weight proteins to nitrocellulose membranes. AB - Here, we report an ultra-rapid method for the transfer of high and low molecular weight proteins to nitrocellulose membranes following sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). In this procedure, the electro transfer was performed with heated (70-75 degrees C) normal transfer buffer from which methanol had been omitted. Complete transfer of high and low molecular weight proteins (a purified protein, molecular weight protein standards and proteins from a human tissue extract) could be carried out in 10 min for a 0.75 mm, 7% SDS-PAGE gel. For 10% and 12.5% gels (0.75 mm), the corresponding time was 15 min. In the case of 1.5-mm gels, a complete transfer could be carried out in 20 min for 7%, 10% and 12.5% gels. The permeability of the gel is increased by heat, such that the proteins trapped in the polyacrylamide gel matrix can be easily transferred to the membrane. When the heat-mediated transfer method was compared with a conventional transfer protocol, under similar conditions, we found that the latter method transferred minimal low molecular weight proteins while retaining most of the high molecular weight proteins in the gel. In summary, this procedure is very rapid, avoids the use of methanol and is particularly useful for the transfer of high molecular weight proteins. PMID- 12133631 TI - Preparation and characterization of immunogens for antibody production against metanephrine and normetanephrine. AB - A two-step zero-length cross-linking procedure using active esters was successfully adopted for conjugating metanephrine (MN) and normetanephrine (NM) to bovine serum albumin (BSA). The protein was activated with water-soluble 1 ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) in the presence of N hydroxysulfosuccimide (sulfo-NHS), leading to the formation of active N succinimidyl esters of some glutamic and aspartic acid carboxyls. The pertinence of this reaction for the coupling of these haptens to carboxylate groups was confirmed via reaction with a model compound, 2-hydroxybenzoic acid, and subsequent characterization using atmospheric pressure chemical ionization mass spectrometry (APCI-MS). Matrix-assisted laser desorption/ionization time-of flight mass spectrometry (MALDI-TOF-MS) was used for the quantitative assessment of the hapten/protein ratios of these conjugates. This technique of conjugate characterization demonstrated greater resolution in molecular weight determination compared to nondenaturing polyacrylamide gel electrophoresis (native PAGE). Preliminary results from enzyme-linked immunosorbent assay (ELISA) and inhibition ELISA procedures using test antisera confirmed that the synthesized immunogens were highly antigenic and elicited specific antibody responses in BALB/c mice against the haptens. PMID- 12133630 TI - A new approach to epitope confirmation by sampling effector/memory T cells migrating to the lung. AB - The identification of T cell epitopes that elicit a weak T cell response is technically challenging due to the relatively low resolution of many available screening assays. Peptide immunization can confirm the immunogenicity of a given peptide, however, this also often induces a low frequency response in lymphoid organs. In this report, we use the murine gammaherpesvirus-68 model system to describe a novel technique to enrich for antigen-experienced T cells in vivo as an aid to epitope mapping. Mice are immunized with peptides containing putative epitopes then effector/memory cells which migrate to the lungs are washed out and tested for specificity using intracellular staining for interferon gamma (IFN gamma). We show that the lung is a site where there are elevated numbers of antigen-experienced T cells and this can be exploited to confirm otherwise low frequency T cell responses. In addition, we identify two novel T cell epitopes in the ORF65 protein of MHV-68 which will be valuable tools in the dissection of the immune response to this model gammaherpesvirus. PMID- 12133632 TI - Measurement of IKK activity in primary rat T cells: rapid activation and inactivation. AB - The translocation of the transcription factor NF-kappaB into the nucleus plays a critical role in many physiological events. In unstimulated cells, NF-kappaB is sequestered in the cytosol, bound to its inhibitor IkappaB. Activation primarily occurs via the IkappaB kinase (IKK) complex which phosphorylates IkappaBalpha at serines 32 and 36, creating a recognition site for IkappaB ubiquitination which then targets IkappaB for degradation. Often it is useful to measure IKK activity to assess upstream signaling events leading to NF-kappaB activation. Current methods of assessing IKK activity are limited to IKK isoforms which are recognized by available IKK antibodies. Here, we describe a procedure to qualitatively assess the overall IKK activity in a cell lysate which can be used on any IKK isoform capable of phosphorylating human IkappaBalpha. This nonradioactive assay is based on measurement of the ability of the cell lysate to phosphorylate GST-IkappaBalpha, as measured by Western blotting, using an anti phospho-IkappaBalpha antibody. We have used this assay to observe the kinetics of TCR-mediated activation of IKK as compared to PMA/ionomycin in primary rat T cells. PMA/ionomycin induces maximal IKK activity within 1 min of stimulation and this activity remains elevated for over 20 min. In comparison, TCR ligation induces maximal IKK activity after 5 min of stimulation and this activity rapidly diminishes to background levels. These data indicate that different stimuli can activate and inactivate IKK with different kinetics and suggest that TCR-mediated activation of IKK is closely linked to the rapid phosphorylation and dephosphorylation, respectively, of TCR-associated kinases. PMID- 12133633 TI - Mutation detection of immunoglobulin V-regions by DHPLC. AB - One of the key features in the affinity maturation of antibodies is somatic hypermutation of the variable regions of immunoglobulin genes. The mutations that occur in immunoglobulin genes are detected by direct sequencing of cloned polymerase chain reaction (PCR) products. The frequencies of mutations in vivo are generally high enough to provide sufficient numbers of point mutations in order to generate large databases that can be analyzed in various ways. Recently, the mechanisms of variable (V)-region hypermutation have been studied in tissue culture systems and transgenic mice where mutation occurs at frequencies that are approximately 10-fold lower than the estimated in vivo rate. Identifying mutations by brute force sequencing of PCR products in comparative studies is limiting when trying to determine if there are statistically significant differences. Here we describe a high throughput technique that can facilitate the identification of immunoglobulin V-regions that contain one or more mutations before sequencing. This technique, known as denaturing high-performance liquid chromatography (DHPLC), utilizes a standard HPLC apparatus with a column that binds double-stranded DNA (dsDNA). In this study, we have successfully detected approximately 90% of previously sequenced mutated V-regions by DHPLC. Our results show that we were able to detect mutations throughout a 321-base pair (bp) region of the Ricin 45 immunoglobulin (Ig) V-region. Also, with the use of this assay, we have been able to detect mutations in multiple clones of different immunoglobulin genes. PMID- 12133634 TI - Increased sensitivity of heterogeneous fluoroimmunoassays employing fluorescein labeled antibodies by simple treatment of the wells with glycerin solution. AB - Here we present a simple and rapid procedure that enhances and stabilizes the fluorescence signal determined directly onto the solid phase and increases the sensitivity of heterogeneous fluoroimmunoassays which employ fluorescein as label. The evaluation of the proposed procedure was performed through a heterogeneous immunofluorimetric assay for mouse gamma-globulins in white-opaque microtitration wells. The proposed fluorescence enhancement and stabilization method consists of a 3-min treatment of the wells with a glycerin solution, after completion of the assay, followed by a 15-min incubation of the emptied wells at 37 degrees C. Following this protocol, an approximately 50% increase of the fluorescence signal was achieved compared with the signal determined without treatment of the wells (control assay). In addition, the proposed protocol resulted in considerable stabilization of the fluorescence signal. The assay sensitivity achieved following this protocol was improved by twofold compared to the control assay, whereas the precision and the dynamic range of the assay were unaffected. PMID- 12133635 TI - Tissue factor pathway inhibitor can interact with platelets. AB - Tissue factor pathway inhibitor (TFPI) regulates the extrinsic pathway of blood coagulation. However, there is no report on interaction between TFPI and platelets other than that by Tsuji, who found that whole blood anticoagulated with TFPI exhibited remarkable decrease in platelet count. Our study revealed that washed platelets suspended in modified Tyrode's buffer (8 mM CaCl2) containing TFPI exhibit platelet aggregation. However, platelets aggregation was observed without TFPI, but its increase and intensity were slow and weak, compared to that in the presence of TFPI. This aggregation was inhibited by anti CD41 (anti-GPIIb) antibody. This finding suggested that TFPI promotes platelet aggregation. PMID- 12133636 TI - Sensing prothymosin alpha origin, mutations and conformation with monoclonal antibodies. AB - To overcome poor immunogenicity of prothymosin alpha, a small and highly acidic nuclear protein involved in cell proliferation, production of anti-prothymosin alpha antibodies in mice immunized with free human prothymosin alpha, with prothymosin alpha coupled to different carriers and with prothymosin alpha fused to green fluorescent protein was assessed. Fusing prothymosin alpha to green fluorescent protein turned out to be the superior approach resulting in production of high titer anti-prothymosin alpha antibodies. From these studies, two highly specific anti-prothymosin alpha monoclonal antibodies recognizing epitopes within the amino terminal (2F11) and middle (4F4) portions of the human prothymosin alpha molecule were obtained and characterized. As expected, the 2F11 antibody displayed broad species specificity, whereas the 4F4 antibody appeared to be species-specific permitting discrimination of human versus rat protein. Furthermore, a combination of point mutations in prothymosin alpha that alter the properties of the protein precluded recognition by the 4F4 antibody. Intramolecular masking of the 4F4 epitope in prothymosin alpha fused to the Tat transduction peptide of human immunodeficiency virus type 1 was observed. The anti-prothymosin alpha antibodies obtained were suitable for precipitation of human prothymosin alpha from HeLa cell lysates and for immunolocalization of the endogenous prothymosin alpha within the cells. Fusion with green fluorescent protein may thus be helpful in raising antibodies against 'problematic' proteins. PMID- 12133637 TI - Isolation and characterization of rabbit single chain antibodies to human Reg Ialpha protein. AB - To investigate the role of Reg Ialpha in human inflammatory bowel disease (IBD), we made two phage-displayed single chain variable fragment (scFv) libraries from rabbits immunized with recombinant or native human Reg Ialpha. After one to three rounds of panning, we were able to isolate phage-displaying scFvs, which bound to human Reg Ialpha. Anti-Reg Ialpha scFvs from both libraries showed similar immunoreactivity to different processed forms of the protein. Despite several DNA fingerprint patterns among these clones, their deduced amino acid sequences are highly homologous with 100% identity in the complementarity-determining regions (CDRs) of the variable segment of heavy chain (VH) region and a small variation in the CDR1 of the variable segment of light chain (VL) region. We also expressed and purified soluble myc-tagged or glutathione S-transferase (GST) fusion scFv proteins from bacteria. Immunohistochemical studies using one of our anti-Reg Ialpha scFv antibodies showed prominent staining in the metaplastic Paneth cell population and light staining in the lamina propria. This scFv antibody is now being used for studies of the role of Reg Ialpha in human IBD. PMID- 12133638 TI - Synthesis of a group-selective antibody library against haptens. AB - A generic strategy is described for the generation of libraries comprising hapten selective antibody genes against a group of structurally related low molecular weight target molecules. Hapten antibody libraries are frequently suffering from high background levels of irrelevant antibody genes as a consequence of the immunization, where the small non-immunogenic target molecule is coupled to a large immunogenic carrier protein. In order to elevate the percentage of hapten specific genes in the library, B cells harboring antibody genes against the group of triazine herbicides were enriched from 21 individual splenocyte populations by means of immunomagnetic separation (IMS). IMS utilizes the specific binding of membrane-associated immunoglobulin receptors on the B cell surface to hapten coated paramagnetic beads. The variable genes of the specifically enriched subpopulation were cloned into a phagemid vector. The corresponding library yielded up to 75% triazine binding antibody clones after three rounds of phage selection. At least half of these antibodies (abs) were displaceable by triazines resulting in quantitative assays with nanomolar sensitivities. In contrast, no displaceable clone was obtained at the same selection level in a control library, where IMS was omitted. Due to the elevated percentage of relevant antibody genes, the library can be utilized either for the direct isolation of functional antibodies against various triazine herbicides or as group-specific gene source for evolutionary antibody optimization. PMID- 12133646 TI - Time to make the Global Fund global. PMID- 12133647 TI - River blindness. PMID- 12133648 TI - Rich nations, poor nations, and bacterial meningitis. PMID- 12133649 TI - Postexposure prophylaxis against transmissible spongiform encephalopathies: CpG oligodeoxynucleotides in mice. PMID- 12133650 TI - Chronic lymphocytic leukaemia: one cell, two diseases? PMID- 12133651 TI - What the UK government is (not) doing about health inequalities. PMID- 12133652 TI - Breast cancer and breastfeeding: collaborative reanalysis of individual data from 47 epidemiological studies in 30 countries, including 50302 women with breast cancer and 96973 women without the disease. AB - BACKGROUND: Although childbearing is known to protect against breast cancer, whether or not breastfeeding contributes to this protective effect is unclear. METHODS: Individual data from 47 epidemiological studies in 30 countries that included information on breastfeeding patterns and other aspects of childbearing were collected, checked, and analysed centrally, for 50302 women with invasive breast cancer and 96973 controls. Estimates of the relative risk for breast cancer associated with breastfeeding in parous women were obtained after stratification by fine divisions of age, parity, and women's ages when their first child was born, as well as by study and menopausal status. FINDINGS: Women with breast cancer had, on average, fewer births than did controls (2.2 vs 2.6). Furthermore, fewer parous women with cancer than parous controls had ever breastfed (71% vs 79%), and their average lifetime duration of breastfeeding was shorter (9.8 vs 15.6 months). The relative risk of breast cancer decreased by 4.3% (95% CI 2.9-5.8; p<0.0001) for every 12 months of breastfeeding in addition to a decrease of 7.0% (5.0-9.0; p<0.0001) for each birth. The size of the decline in the relative risk of breast cancer associated with breastfeeding did not differ significantly for women in developed and developing countries, and did not vary significantly by age, menopausal status, ethnic origin, the number of births a woman had, her age when her first child was born, or any of nine other personal characteristics examined. It is estimated that the cumulative incidence of breast cancer in developed countries would be reduced by more than half, from 6.3 to 2.7 per 100 women by age 70, if women had the average number of births and lifetime duration of breastfeeding that had been prevalent in developing countries until recently. Breastfeeding could account for almost two-thirds of this estimated reduction in breast cancer incidence. INTERPRETATION: The longer women breast feed the more they are protected against breast cancer. The lack of or short lifetime duration of breastfeeding typical of women in developed countries makes a major contribution to the high incidence of breast cancer in these countries. PMID- 12133653 TI - Efficacy and safety of intravenous levosimendan compared with dobutamine in severe low-output heart failure (the LIDO study): a randomised double-blind trial. AB - BACKGROUND: Levosimendan, a novel calcium sensitiser, improves myocardial contractility without causing an increase in myocardial oxygen demand. We compared the effects of levosimendan and dobutamine on haemodynamic performance and clinical outcome in patients with low-output heart failure. METHODS: Patients were recruited into a multicentre, randomised, double-blind, double-dummy, parallel-group trial. Under continuous haemodynamic monitoring, an initial loading dose of levosimendan of 24 microg/kg was infused over 10 min, followed by a continuous infusion of 0.1 microg kg(-1) min(-1) for 24 h. Dobutamine was infused for 24 h at an initial dose of 5 microg kg(-1) min(-1) without a loading dose. The infusion rate was doubled if the response was inadequate at 2h. The primary endpoint was the proportion of patients with haemodynamic improvement (defined as an increase of 30% or more in cardiac output and a decrease of 25% or more in pulmonary-capillary wedge pressure) at 24 h. Analyses were by intention to treat. FINDINGS: 103 patients were assigned levosimendan and 100 dobutamine. The primary haemodynamic endpoint was achieved in 29 (28%) levosimendan-group patients and 15 (15%) in the dobutamine group (hazard ratio 1.9 [95% CI 1.1-3.3]; p=0.022). At 180 days, 27 (26%) levosimendan-group patients had died, compared with 38 (38%) in the dobutamine group (0.57 [0.34-0.95]; p=0.029). INTERPRETATION: In patients with severe, low-output heart failure, levosimendan improved haemodynamic performance more effectively than dobutamine. This benefit was accompanied by lower mortality in the levosimendan group than in the dobutamine group for up to 180 days. PMID- 12133654 TI - Effects of standard and high doses of ivermectin on adult worms of Onchocerca volvulus: a randomised controlled trial. AB - BACKGROUND: At present, control of onchocerciasis depends almost entirely on yearly treatments with 150 microg/kg ivermectin. We aimed to compare the effect of higher doses, more frequent doses, or both with the standard regimen on adult Onchocerca volvulus. METHODS: We randomly allocated 657 patients who had onchocerciasis to 150 microg/kg ivermectin yearly (reference group), 150 microg/kg every 3 months, 400 then 800 microg/kg yearly, or 400 then 800 microg/kg every 3 months. We took skin snip samples from every patient before, and 3 years and 4 years after the first dose, and, at the same time excised one subcutaneous O volvulus nodule, which was examined histologically. The primary outcome was the vital status of the female worms. Analysis was done per protocol. FINDINGS: We obtained nodules from 511 patients. After 3 years of treatment, more female worms had died in the groups treated every 3 months than in the reference group (odds ratio=1.84 [95% CI 1.23-2.75], p=0.003 for 150 microg/kg; and 2.17 [1.42-3.31], p<0.001 for high doses). Female worms were also less fertile in these groups than in the reference group (0.24 [0.14-0.43], p<0.0001; and 0.14 [0.06-0.29], p<0.0001, respectively). No difference was recorded between groups treated yearly (p=0.83 for the proportion of dead females). Unexpected side effects consisted of mild, temporary, subjective visual changes in patients on high-dose regimens. INTERPRETATION: Treatment with 3-monthly ivermectin could greatly reduce the number of female worms and acute itching and skin lesions; lower transmission of O volvulus; and change the duration of control programmes. PMID- 12133655 TI - Acute infantile haemorrhagic oedema. PMID- 12133656 TI - Dexamethasone treatment in childhood bacterial meningitis in Malawi: a randomised controlled trial. AB - BACKGROUND: Steroids are used as adjuvant treatment in childhood pyogenic meningitis to attenuate host inflammatory responses to bacterial invasion. We aimed to assess the effectiveness of dexamethasone in management of acute bacterial meningitis in a developing country. METHODS: In a double-blind, placebo controlled trial, we included 598 children with pyogenic meningitis who had been admitted to the children's wards of the Queen Elizabeth Central Hospital, Blantyre, Malawi. We did physical, neurological, developmental, and hearing assessments at 1 and 6 months after discharge. The primary outcome was overall death. Secondary outcomes included sequelae, in-hospital deaths, and death after discharge. Analysis was done by intention to treat. FINDINGS: Of the 598 included children, 307 (51%) were assigned to dexamethasone and 295 (49%) to placebo. 338 (40%) of 598 patients had Streptococcus pneumoniae, 170 (28%) Haemophilus influenzae type b, 66 (11%) Neisseria meningitidis, and 29 (5%) Salmonella spp. 78 (13%) patients had no growth on culture. The number of overall deaths was the same in the two treatment groups (relative risk 1.00 [95% CI 0.8-1.25], p=0.93). At final outcome, sequelae were identified in 84 (28%) of children on steroids and in 81 (28%) on placebo (relative risk 0.99 [95% CI 0.78-1.27], p=0.97). The number of children dying in hospital did not differ between groups. INTERPRETATION: Steroids are not an effective adjuvant treatment in children with acute bacterial meningitis in developing countries. PMID- 12133658 TI - A puzzling cause of hypokalaemia. PMID- 12133657 TI - Association between mitochondrial dysfunction and severity and outcome of septic shock. AB - BACKGROUND: Sepsis-induced multiple organ failure is the major cause of mortality and morbidity in critically ill patients. However, the precise mechanisms by which this dysfunction is caused remain to be elucidated. We and others have shown raised tissue oxygen tensions in septic animals and human beings, suggesting reduced ability of the organs to use oxygen. Because ATP production by mitochondrial oxidative phosphorylation accounts for more than 90% of total oxygen consumption, we postulated that mitochondrial dysfunction results in organ failure, possibly due to nitric oxide, which is known to inhibit mitochondrial respiration in vitro and is produced in excess in sepsis. METHODS: We did skeletal muscle biopsies on 28 critically ill septic patients within 24 h of admission to intensive care, and on nine control patients undergoing elective hip surgery. The biopsy samples were analysed for respiratory-chain activity (complexes I-IV), ATP concentration, reduced glutathione (an intracellular antioxidant) concentration, and nitrite/nitrate concentrations (a marker of nitric oxide production). FINDINGS: Skeletal muscle ATP concentrations were significantly lower in the 12 patients with sepsis who subsequently died than in the 16 septic patients who survived (p=0.0003) and in controls (p=0.05). Complex I activity had a significant inverse correlation with norepinephrine requirements (a proxy for shock severity, p=0.0003) and nitrite/nitrate concentrations (p=0.0004), and a significant positive correlation with concentrations of reduced glutathione (p=0.006) and ATP (p=0.03). INTERPRETATION: In septic patients, we found an association between nitric oxide overproduction, antioxidant depletion, mitochondrial dysfunction, and decreased ATP concentrations that relate to organ failure and eventual outcome. These data implicate bioenergetic failure as an important pathophysiological mechanism underlying multiorgan dysfunction. PMID- 12133659 TI - Prevalence of dementia in institutional care. AB - Information about dementia prevalence in residential and nursing care homes helps these institutions to meet residents' needs. We combined residential characteristics ascertained at a screening interview in 1991-93 with dementia diagnoses from assessment interviews in the UK Medical Research Council Cognitive Function and Ageing Study, a population-based study of 13004 elderly people. Almost 5% of people aged 65 years or older lived in institutions--34% (95% CI 30 39) of individuals with dementia. Within institutions, dementia prevalence was 62% (52-71), and was slightly higher in women than in men, with no increase by age. That most residents have dementia has implications for the type of care that institutions provide. PMID- 12133660 TI - Bartonella quintana in human erythrocytes. AB - Bartonella quintana is transmitted by body lice among homeless people. Infection with this organism leads to chronic bacteraemia with few symptoms. We looked for B quintana in erythrocytes in a population of homeless people in Marseille, France. In this report we show the intraerythrocytic presence of B quintana in people with symptomless bacteraemia, by use of a specific monoclonal antibody and laser confocal microscopy. Presence of at least 5 x 10(4) colony-forming units per mL was predictive of intracellular infection. However, such infection was not associated with evidence of haemolysis. Colonisation of erythrocytes could allow efficient transmission of B quintana by body lice. PMID- 12133661 TI - Viral load of human papillomavirus and risk of CIN3 or cervical cancer. AB - Carcinogenic human papillomaviruses (HPV) are thought to be necessary for development of cervical cancer. We assessed whether higher viral loads of such viruses predicted future risk of CIN3 or cancer (CIN3+) in a cohort of 20810 women followed up for 10 years with cytological screening. We measured the viral load for 13 types of carcinogenic HPV (relative light units normalised to 1 pg/mL HPV 16 positive controls [RLU/PC]) using Hybrid Capture 2 testing of cervicovaginal lavages obtained at enrolment. Results were stratified into four groups (RLU/PC 1 to <10, 10 to <100, 100 to <1000, > or = 1000). Although presence of HPV strongly increased risk of CIN3+, high viral load did not further predict risk of CIN3+. PMID- 12133662 TI - Postexposure prophylaxis against prion disease with a stimulator of innate immunity. AB - The absence of an immune response to prions--the infectious agents of scrapie, bovine spongiform encephalopathy (BSE), and Creutzfeldt-Jakob disease--might be related to the fact that these agents do not contain nucleic acids. We aimed to use CpG oligodeoxynucleotides, which have been shown to stimulate innate immunity, as a form of postexposure prophylaxis in mice. We inoculated healthy mice with brain homogenates from mice infected with the RML scrapie prion, and then injected CpG oligodeoxynucleotides. This postexposure prophylaxis with CpG oligodeoxynucleotides resulted in 38% longer survival times than treatment with saline (p<0.0001), or even longer after repeated application. The explanation for this finding remains to be elucidated, but the most likely is stimulation of TLR9 expressing cells of the innate immune system such as macrophages, monocytes, and dendritic cells. CpG oligodeoxynucleotides have not been shown to have adverse effects to human health and could therefore be considered as a therapeutic choice in postexposure prophylaxis. PMID- 12133663 TI - US expert panel divided over human therapeutic cloning. PMID- 12133668 TI - Unsolicited fluoxetine mailing triggers investigation in USA. PMID- 12133670 TI - Europe investigates hormone-contaminated waste in food chain. PMID- 12133671 TI - WHO reassesses appropriate body-mass index for Asian populations. PMID- 12133672 TI - Bold developments in UK sexual health services. PMID- 12133673 TI - Obstructive sleep apnoea. AB - Obstructive sleep apnoea is a disease of increasing importance because of its neurocognitive and cardiovascular sequelae. Abnormalities in the anatomy of the pharynx, the physiology of the upper airway muscle dilator, and the stability of ventilatory control are important causes of repetitive pharyngeal collapse during sleep. Obstructive sleep apnoea can be diagnosed on the basis of characteristic history (snoring, daytime sleepiness) and physical examination (increased neck circumference), but overnight polysomnography is needed to confirm presence of the disorder. Repetitive pharyngeal collapse causes recurrent arousals from sleep, leading to sleepiness and increased risk of motor vehicle and occupational accidents. The surges in hypoxaemia, hypercapnia, and catecholamine associated with this disorder have now been implicated in development of hypertension, but the association between obstructive sleep apnoea and myocardial infarction, stroke, and congestive heart failure is not proven. Continuous positive airway pressure, the treatment of choice for obstructive sleep apnoea, reduces sleepiness and improves hypertension. PMID- 12133674 TI - Human rights, politics, and reviews of research ethics. AB - Although the human rights movement and the sphere of research ethics have overlapping principles and goals, there has been little attempt to incorporate external political and human rights contexts into research ethics codes or ethics reviews. Every element of a research ethics review--the balance of risks and benefits, the assurance of rights for individual participants, and the fair selection of research populations--can be affected by the political and human rights background in which a study is done. Research that at first seems to be low in risk may become high in risk if implemented in a country where the government might breach the confidentiality of study results or where results might be used to deport a refugee group. Researchers should determine whether research could or should be done by consulting human rights organisations and, when possible, a trusted colleague, to learn the background political context and human rights conditions of the settings in which they propose to do research. PMID- 12133675 TI - The inverse care law today. PMID- 12133676 TI - Checks and balances. PMID- 12133677 TI - Prescribing errors. PMID- 12133678 TI - Prescribing errors. PMID- 12133679 TI - Prescribing errors. PMID- 12133680 TI - Prescribing errors. PMID- 12133681 TI - Platelet glycoprotein IIb/IIIa inhibitors in acute coronary syndromes. PMID- 12133682 TI - Platelet glycoprotein IIb/IIIa inhibitors in acute coronary syndromes. PMID- 12133683 TI - Platelet glycoprotein IIb/IIIa inhibitors in acute coronary syndromes. PMID- 12133684 TI - Statins, electron-beam CT, and aortic-valve calcification. PMID- 12133685 TI - Epidemiology research. PMID- 12133686 TI - Epidemiology research. PMID- 12133687 TI - Epidemiology research. PMID- 12133688 TI - Epidemiology research. PMID- 12133689 TI - Epidemiology research. PMID- 12133691 TI - Encephalitis and dengue. PMID- 12133692 TI - Effects of biological attack on US food supplies. PMID- 12133693 TI - Proprioceptive increase with elasticated limb and joint supports. PMID- 12133694 TI - First round of payments from the Global Fund. PMID- 12133702 TI - Identification and characterization of a baculovirus from Lonomia obliqua (Lepidoptera: Saturniidae). AB - A baculovirus has been isolated from larvae of Lonomia obliqua, a Saturniidae of medical importance due to a potent toxin found in their spines. Electron Microscopy analysis of the occlusion body obtained from diseased larvae showed polyhedra of approximately 1 microm in diameter containing multiple nucleocapsids per envelope. This baculovirus was thus named Lonomia obliqua multicapsid nucleopolyhedrovirus (LoobMNPV). Restriction endonuclease profiles of viral DNA digested with three restriction enzymes were obtained and the genome size was estimated to be 95.52 +/- 2.3 kbp. The polyhedrin gene of LoobMNPV was identified and its DNA sequence was determined. Phylogenetic analysis of the polyhedrin gene showed that the LoobMNPV polyhedrin belongs to group I NPV and that it is closely related to the polyhedrin of the NPV of Amsacta albistriga. PMID- 12133704 TI - Effects of Bucephalus sp. (Trematoda: Bucephalidae) on Perna perna mussels from a culture station in Ratones Grande Island, Brazil. AB - This study reports the prevalence of Bucephalus sp. in Perna perna populations from a culture station of southern Brazil and its effect on the mussel reproductive tissue and immune system. The prevalence of Bucephalus sp. in P. perna (n = 1871) was considered low (3.1%) and did not seasonally vary. Histological sections of the mantle of infected mussels revealed a marked (80%) reduction of the reproductive tissue that was severe even in mussels exhibiting a moderate infection degree. The total (THC) and differential (DHC) hemocyte counts were lower in infected mussels (3.9 x 10(6) hem/ml; granular hemocytes = 33%) as compared with non-infected animals (5.5 x 10(6) hem/ml; granular hemocytes = 40%). The plasma protein concentration did not vary upon infection. Hemocyte infiltration was significantly higher only in mussels with a very heavy infection degree. The parasite sporocysts were never seen encapsulated by the host hemocytes. Our results indicate that Bucephalus sp. promotes a severe castration in its host and apparently evades the mussel immune system. PMID- 12133706 TI - Heat curing Metaseiulus occidentalis (Nesbitt) (Acari, Phytoseiidae) of a fitness reducing microsporidium. AB - Laboratory colonies of the predatory mite Metaseiulus occidentalis in Gainesville, FL were found to be infected with an undescribed microsporidium. Experiments were performed to quantify the effect of infection on the fitness of M. occidentalis and to determine if heat treatment can cure mites of the microsporidium. The colonies tested were derived from an isofemale line so that differences in performance could be attributed to the presence of microsporidia. A subcolony of an uninfected isofemale line was infected with the microsporidium by feeding females infected eggs from another colony of M. occidentalis. Infected mites had a shorter mean (+/-SD) female life span (7.4 +/- 2.9 vs. 10.0 +/- 2.8 days), lower mean oviposition (1.6 +/- 0.7 vs. 2.2 +/- 0.4 eggs/day), and a male biased sex ratio (43 +/- 16% vs. 57 +/- 15% female progeny). The infection was reduced temporarily in colonies initiated from mites that were reared in a growth chamber at 33 degrees C from egg to adult, but healthy colonies only were established from the progeny of the heat-treated adults. These colonies remained free of infection for 10 weeks. PMID- 12133705 TI - Morphological and molecular characterization of a new microsporidian species from the predatory mite Metaseiulus occidentalis (Nesbitt) (Acari, Phytoseiidae). AB - A new microsporidian species is described from the predatory mite Metaseiulus (formerly Typhlodromus or Galendromus) occidentalis (Nesbitt) (Acari, Phytoseiidae). The ultrastructure of this new species is presented together with the first molecular characterization for a microsporidium of mites. All stages of this new microsporidium are haplokaryotic and develop in direct contact with the host-cell cytoplasm. Sporogony is disporoblastic and spores are formed in eggs, immature stages, and adults of M. occidentalis. There are two morphological classes of spores, one with a short polar filament (3-5 coils) that measured 2.53 x 1.68 microm and one with a longer polar filament (8-9 coils) that measured 3.14 x 1.77 microm. Horizontal transmission of this new species occurs by cannibalism of eggs and other stages and perhaps involves the spores with the long polar filament. Spores with the short polar filament may play a role in autoinfection and vertical (transovarial) transmission that is highly efficient in transferring the microsporidium from adults to progeny. Analysis of the small subunit ribosomal DNA indicated that this species from M. occidentalis is most closely related to the Nosema/Vairimorpha clade of microsporidia. A conflict between the morphological and molecular data is discussed. The species is compared to previously described microsporidia of arachnids resulting in creation of Oligosporidium occidentalis n. sp. in the family Unikaryonidae. PMID- 12133703 TI - Antimicrobial activity of Xenorhabdus sp. RIO (Enterobacteriaceae), symbiont of the entomopathogenic nematode, Steinernema riobrave (Rhabditida: Steinernematidae). AB - Galleria mellonella larvae infected with Steinernema riobrave soon showed (after 24 h) the typical growth of its Xenorhabdus sp. RIO symbiont and, in parallel, the growth of another Gram negative bacterial species in the body cavity. A population of Entercoccus sp. in the nematode infected larvae collapsed to zero by 96 h. The level of antibiotic and antimycotic activity followed a pattern similar to that of the growth curve to stationary phase of the Xenorhabdus sp. RIO symbiont, over a period of 168 h. The antimycotic activity was composed of exo- and endochitinases as well as other proteinaceous and some small molecule compounds. The changing pH, relatively high growth rate of Xenorhabdus sp. RIO compared with that of other Gram negative bacterial species and of collapse of the Enterococcus sp. population enabled Xenorhabdus sp. RIO to out-compete other species. PMID- 12133707 TI - Infection by a Hematodinium-like parasitic dinoflagellate causes Pink Crab Disease (PCD) in the edible crab Cancer pagurus. AB - The edible crab (Cancer pagurus) supports a large and valuable fishery in UK waters. Much of the catch is transported live to continental Europe in specially designed live-well ('vivier') vehicles. During the winter of 2000/2001, many trap caught crabs from Guernsey, Channel Islands, UK, were reportedly moribund and pink in colour. These crabs generally died before and during vivier transportation. We provide histological, immunological, and molecular evidence that this condition is associated with infection by a Hematodinium-like dinoflagellate parasite similar to that previously reported in C. pagurus and to an infection causing seasonal mass mortalities of the Norway lobster (Nephrops norvegicus). Pathologically, every altered host bore the infection, which was characterised by very large numbers of plasmodial and vegetative stages in the haemolymph and depletion of reserve cells in the hepatopancreas. Due to the hyperpigmentation of the carapace and appendages, we have called this infection 'Pink Crab Disease' (PCD). Similar Hematodinium infections cause 'Bitter Crab Disease' in tanner and snow crabs, which has had a negative effect on their marketability. At present, little is known about the seasonality, transmission, and market impact of this infection in C. pagurus. PMID- 12133708 TI - The strain variation and virulence of granulovirus of diamondback moth (Plutella xylostella Linnaeus, Lep., Yponomeutidae) isolated in Kenya. PMID- 12133709 TI - Polydora ciliata shell infestation in Tapes philippinarum Manila clam held out of the substrate in the Adriatic sea, Italy. PMID- 12133710 TI - First field report of a nematode (Tylenchida: Sphaerularioidea) attacking the coffee berry borer, Hypothenemus hampei (Ferrari) (Coleoptera: Scolytidae) in the Americas. PMID- 12133711 TI - Usefulness of staining parasporal bodies when screening for Bacillus thuringiensis. PMID- 12133715 TI - Staking out novelty on the genomic frontier. AB - Recent advances in genomics include global assessment and classification of genome content, high-throughput biological pathway construction, systematic identification of previously unpredicted genes and the in vitro creation of novel motifs with biological function not found in nature (extra-genomic gene discovery). The ability to make global surveys of transcriptomes has given rise to fields such as pharmacogenomics and toxicogenomics. These applications of genomics technologies, with conventional drug assessment methodologies, will lead to more tolerable drugs and a better understanding of clinical populations. Integration of pathway mapping, using proteomics married to expression, will also significantly affect how new therapeutics are discovered as cross-biological cross-pathway interactions lead to novel drug targets and better predictions of drug tolerance. PMID- 12133716 TI - Proteomics in drug discovery. AB - Drug discovery is a prolonged process that uses a variety of tools from diverse fields. To accelerate the process, a number of biotechnologies, including genomics, proteomics and a number of cellular and organismic methodologies, have been developed. Proteomics development faces interdisciplinary challenges, including both the traditional (biology and chemistry) and the emerging (high throughput automation and bioinformatics). Emergent technologies include two dimensional gel electrophoresis, mass spectrometry, protein arrays, isotope encoding, two-hybrid systems, information technology and activity-based assays. These technologies, as part of the arsenal of proteomics techniques, are advancing the utility of proteomics in the drug-discovery process. PMID- 12133717 TI - The path to personalized medicine. AB - Advances in personalized medicine, or the use of an individual's molecular profile to direct the practice of medicine, have been greatly enabled through human genome research. This research is leading to the identification of a range of molecular markers for predisposition testing, disease screening and prognostic assessment, as well as markers used to predict and monitor drug response. Successful personalized medicine research programs will not only require strategies for developing and validating biomarkers, but also coordinating these efforts with drug discovery and clinical development. PMID- 12133719 TI - Protein flexibility and drug design: how to hit a moving target. AB - The most advanced methods for computer-aided drug design and database mining incorporate protein flexibility. Such techniques are not only needed to obtain proper results; they are also critical for dealing with the growing body of information from structural genomics. PMID- 12133720 TI - Modern methods to produce natural-product libraries. AB - Natural sources offer a wealth of chemically diverse compounds that have been evolutionary preselected to modulate biochemical pathways. Several industrial and academic groups are accessing this source using advanced technology platforms. Methods have been reported to generate large and diverse natural-product libraries optimised for high-throughput screening and for a fast discovery process. In addition to prefractionated and pure natural-product libraries, parallel synthesis gives access to synthetic, semi-synthetic and natural-product like libraries. Natural-product chemistry and organic synthesis are powerful tools for optimising natural leads and for generating new diversity from natural scaffolds. The amalgamation of both may be expected to become an important strategy in future drug design. PMID- 12133718 TI - Lead discovery using molecular docking. AB - As the structures of more and more proteins and nucleic acids become available, molecular docking is increasingly considered for lead discovery. Recent studies consider the hit-rate enhancement of docking screens and the accuracy of docking structure predictions. As more structures are determined experimentally, docking against homology-modeled targets also becomes possible for more proteins. With more docking studies being undertaken, the 'drug-likeness' and specificity of docking hits is also being examined. PMID- 12133721 TI - Recent advances in the synthesis, design and selection of cysteine protease inhibitors. AB - Inhibition of cysteine proteases is emerging as an important strategy for the treatment of a variety of human diseases. Intense efforts involving structure based inhibitor design have been directed toward several cysteine proteases, including cathepsin K, calpain, human rhinovirus 3C protease and several parasitic cysteine protease targets. Other successful recent efforts have involved combinatorial synthesis and screening for identification of new inhibitor templates. PMID- 12133722 TI - Dendritic polymer macromolecular carriers for drug delivery. AB - Dendrimers are synthetic, highly branched, mono-disperse macromolecules of nanometer dimensions. Started in the mid-1980s, the research investigations into the synthetic methodology, physical and chemical properties of these macromolecules are increasing exponentially with growing interest in this field. Potential applications for dendrimers are now forthcoming. Properties associated with these dendrimers such as uniform size, water solubility, modifiable surface functionality and available internal cavities make them attractive for biological and drug-delivery applications. PMID- 12133723 TI - Progress toward the development of agents to modulate the cell cycle. AB - With taxanes continuing to prove useful in the clinical treatment of cancer, the next generation of antimitotic agents has entered clinical trials. Other mechanisms awaiting proof-of-concept for the treatment of antiproliferative diseases include inhibition of cyclin-dependent kinases (Cdks). Flavopiridol and UCN-01 are continuing in clinical trials, and newer more selective Cdk inhibitors are now entering clinical evaluation. PMID- 12133724 TI - A view to a kill: ligands for Bcl-2 family proteins. AB - Apoptosis is the essential process of programmed cell death that, in multicellular organisms, regulates development and maintains homeostasis. Defects in the apoptotic molecular machinery that result in either excessive or insufficient apoptosis are observed in a remarkably wide range of human disease, prompting intense interest in pro- and anti-apoptotic proteins as therapeutic targets. A number of recent reports have described the discovery of ligands for anti-apoptotic Bcl-2 family proteins by a variety of approaches, including computational, combinatorial and evolutionary strategies. Both the design of ligands and the exploration of their mechanisms of action have been greatly enhanced by recent high-resolution structure determinations of proteins from this family. Several of the newly discovered ligands promote apoptosis, and some do so even in the face of overexpressed anti-apoptotic Bcl-2 proteins. Ligands that overcome the protective effects associated with up-regulation of anti-apoptotic Bcl-2 proteins represent especially promising therapeutic leads. PMID- 12133725 TI - A strategy for the design of multiplex inhibitors for kinase-mediated signalling in angiogenesis. AB - Tumour growth is dependent on multiple factors, including the physiological process of angiogenesis. Several opportunities for inhibiting angiogenesis with targeted therapies have been identified and are currently being evaluated for clinical efficacy. Some of the most promising approaches include small-molecule inhibitors for the tyrosine receptor kinase VEGFR2. Other signal-transduction pathways have also been shown to regulate angiogenesis, including FGFR, PDGFR, Tie and EphB. PMID- 12133726 TI - Preclinical and clinical evaluation of proteasome inhibitor PS-341 for the treatment of cancer. AB - The proteasome is a multicatalytic protease, present in all eukaryotic cells, that is primarily responsible for intracellular protein degradation. By destroying regulatory proteins or their inhibitors, the proteasome influences many cellular regulatory signals and is thus a potential target for pharmacological agents. The dipeptide boronic acid analogue PS-341 is a potent and selective proteasome inhibitor in clinical trials for a variety of tumor types. In vitro and in vivo (murine xenograft) studies show that PS-341 has activity against a variety of malignancies, including myeloma, chronic lymphocytic leukemia, prostate cancer, pancreatic cancer, breast cancer and colon cancer. PMID- 12133727 TI - Notch signaling as a therapeutic target. AB - Signals transduced by Notch receptors influence differentiation and proliferation in a wide variety of cell types. Activation of a Notch signal by one of several ligands triggers a series of proteolytic cleavages that release the intracellular region of Notch from the membrane, allowing it ultimately to translocate to the nucleus and activate the transcription of downstream target genes. Recent studies have elucidated the roles of several key proteins that participate in and modulate these central events in Notch signal transduction. These advances offer a variety of potential avenues to manipulate Notch signaling for therapeutic purposes in the treatment of cancer and in stem cell maintenance. PMID- 12133728 TI - Chemokine receptor antagonism as an approach to anti-inflammatory therapy: 'just right' or plain wrong? AB - Inflammation plays a pivotal role in exacerbating a wide array of human diseases. The chemokines are a group of proteins that control the movement and activation of the immune cells involved in all aspects of the inflammatory response. Recently, their cognate receptors have attracted considerable interest as therapeutic targets, in part because they are G-protein-coupled receptors, which have been antagonized successfully before by the pharmaceutical industry. Indeed, several companies have now reported the development of selective small-molecule chemokine receptor antagonists, and some of these compounds have even entered human Phase I clinical trials. Preclinical studies of the responsiveness of murine models of inflammation to either pharmacologic or genetic intervention have suggested that antagonism of some chemokine receptors may well prove to be a safe and efficacious approach to anti-inflammatory therapy. PMID- 12133729 TI - New asthma targets: recent clinical and preclinical advances. AB - Current asthma therapy is directed at the relief of chronic inflammation or improving lung function through bronchodilation. These approaches treat the overt symptoms of asthma but do not approach underlying causes of the disease. Such therapies have limited efficacy and for a number of patients the disease remains poorly controlled. The short-term future of asthma therapy will probably focus on the treatment of multiple symptoms to provide improved lung function. Long-term approaches to asthma will have to focus on modulation of the mechanisms that are the underlying causes of the various asthmatic pathophysiologies. These targets include a number of TH2-type T-cell-generated cytokines and chemokines, G-protein coupled receptors, TH2-related transcription factors, neurotrophins and adhesion molecules. Additional new targets and understanding of asthma may also arise from genetic analysis. PMID- 12133730 TI - Anti-integrin as novel drug-discovery targets: potential therapeutic and diagnostic implications. AB - The role of integrin and extracellular matrix proteins in various pathological processes (including angiogenesis, thrombosis, apoptosis and cell migration and proliferation), leading to both acute and chronic disease states (e.g. ocular diseases, metastasis, unstable angina, myocardial infarction, stroke, osteoporosis, a wide range of inflammatory diseases, vascular remodeling and neurodegenerative disorders) has been recently documented. A key success in this field is evident from the potential role of the platelet GPIIb/IIIa (alphaIIbbeta3) integrin in the prevention, treatment and perhaps diagnosis of various thromboembolic disorders. Additionally, progress has been shown in the development of leukocyte alpha4beta1 antagonists for various inflammatory indications and alphav integrin antagonists for angiogenesis and vascular-related disorders. However, the exact modes of action of certain integrin antagonists are still not fully clear. Integrin antagonists in clinical or pre-clinical development are expected to be used as a stand-alone therapy or, better, as an adjunct to other pharmacotherapy, radiotherapy or interventional procedures. PMID- 12133732 TI - Seeking a contemporary understanding of factors that influence physical activity. PMID- 12133733 TI - Toward a better understanding of the influences on physical activity: the role of determinants, correlates, causal variables, mediators, moderators, and confounders. AB - BACKGROUND: For research on physical activity interventions to progress systematically, the mechanisms of action must be studied. In doing so, the research methods and their associated concepts and terminology become more complex. It is particularly important to clearly distinguish among determinants, correlates, mediators, moderators, and confounder variables used in physical activity research. This article examines the factors that are correlated with and that may have a causal relationship to physical activity. METHODS AND RESULTS: We propose that the term "correlate" be used, instead of "determinant," to describe statistical associations or correlations between measured variables and physical activity. Studies of the correlates of physical activity are reviewed. The findings of these studies can help to critique existing theories of health behavior change and can provide hypotheses to be tested in intervention studies from which it is possible to draw causal inferences. Mediator, moderator, and confounder variables can act to influence measured changes in physical activity. Intervening causal variables that are necessary to complete a cause-effect pathway between an intervention and physical activity are termed "mediators." The relationship between an intervention and physical activity behaviors may vary for different groups; the strata by which they vary are levels of "moderators" of the relationship. Other factors may distort or affect the observed relationships between program exposure and physical activity, and are known as "confounders." CONCLUSIONS: Consistent use of terms and additional research on mediators and moderators of intervention effects will improve our ability to understand and influence physical activity. PMID- 12133734 TI - Theoretical approaches to the promotion of physical activity: forging a transdisciplinary paradigm. AB - BACKGROUND: Research in the physical activity promotion arena has focused on the application of theoretical perspectives aimed primarily at personal levels of understanding and analysis. The investigation of such theories has provided some insights related to potentially useful mediators of physical activity behavior. However, to continue to expand this field, new perspectives on personal-level theories, in addition to the exploration of more macro-level conceptual perspectives, are required. OBJECTIVE: The purpose of this article is to: (1) briefly review the current strengths and limitations of the personal-level, physical activity-theory literature; and (2) introduce concepts and perspectives from other fields, including the social-ecology and urban-planning fields, of potential relevance to the physical activity arena. METHOD: We provide an overview of potentially relevant theoretical perspectives aimed at different levels of understanding and analysis, from the personal level through the broader scale meso- and macro-environmental perspectives. In addition, we suggest initial steps to take in developing a transdisciplinary paradigm encompassing all such levels of analysis and investigation. CONCLUSIONS: Given the scope of the physical inactivity epidemic facing the U.S. population currently and in the future, methods and approaches that integrate theory and concepts across a broader group of disciplines will be increasingly necessary. PMID- 12133735 TI - Psychosocial mediators of physical activity behavior among adults and children. AB - BACKGROUND: Researchers examining theory-based, physical activity (PA) interventions postulate that interventions are effective by changing theoretical constructs hypothesized to mediate the relationship between the intervention and PA behavior. Research indicates that PA interventions are effective for increasing PA behavior. However, whether effective interventions are due to predicted changes in theoretical constructs remains poorly understood. METHODS: Studies that examined theoretical constructs (i.e., mediators) in PA interventions of adults or children, which used experimental designs and met other criteria for evaluating mediation, were collected via literature searches, personal searches of files, and personal communications. Only studies examining the direct effect of the intervention on the hypothesized mediator were considered relevant for this study. RESULTS: Based on our criteria, the adult literature search yielded ten studies and the child literature search yielded two studies. The most common mediators examined included behavioral processes of change, cognitive processes of change, self-efficacy, decisional balance, social support, and enjoyment. Research indicates that behavioral processes are likely mediators. There was some support for the importance of self-efficacy as a mediator. CONCLUSIONS: Few studies have used statistically recommended methods to examine mediators in PA intervention studies. Therefore, definitive conclusions about the importance of the mediators reviewed are not possible at this time. Additional PA mediator-intervention studies using recommended statistical methods are necessary to truly test if theory-based PA interventions are effective due to predicted changes in theoretical constructs. PMID- 12133736 TI - Exploring the effect of the environment on physical activity: a study examining walking to work. AB - BACKGROUND: Research on physical activity and the physical environment is at the correlates stage, so it is premature to attribute causal effects. This paper provides a conceptual approach to understanding how the physical design of neighborhoods may influence behavior by disentangling the potential effects of income, university education, poverty, and degree of urbanization on the relationship between walking to work and neighborhood design characteristics. METHODS: The study merges Canadian data from 27 neighborhood observations with information on walking to work from the 1996 census. Hierarchical linear modeling was used to create a latent environment score based on 18 neighborhood characteristics (e.g., variety of destinations, visual aesthetics, and traffic). The relationship between the environment score and walking to work was modeled at the second level, controlling for income, university education, poverty, and degree of urbanization. RESULTS: With the exceptions of visual interest and aesthetics, each neighborhood characteristic contributed significantly to the environment score. The environment score was positively associated with walking to work, both with and without adjustment for degree of urbanization. Controlling for university education, income, and poverty did not influence these relationships. CONCLUSIONS: The positive association between the environment score and walking to work, controlling for degree of urbanization supports the current movement toward the development of integrated communities for housing, shops, workplaces, schools, and public spaces. Given the need for research to guide environmental interventions, collaboration among public health practitioners, urban planners, and transportation researchers is essential to integrate knowledge across sectors. PMID- 12133737 TI - Emerging measurement and statistical methods in physical activity research. AB - Although many studies have attempted to identify mediators and moderators of changes in physical activity involvement, the literature is inconclusive regarding which variable(s) relate to physical activity behavior change. The Cooper 2001 Conference series dedicated a session to discussing measurement and statistical methods that could contribute to advancing this research agenda. This article focuses on four such methodologic approaches: qualitative; psychometric; latent-variable, structural equation modeling; and multilevel modeling. The article presents a brief overview of these methods and discusses potential advantages and limitations of using them. PMID- 12133738 TI - The use of surveillance data and market research to promote physical activity. AB - Using various types of data sources for assessing and monitoring physical activity behaviors on a population level adds to our ability to explain the relationships between individuals and their surrounding social and physical environments. This article presents the findings from part of a panel presentation on available data sets at the 2001 Cooper Conference on Innovative Approaches to Understanding and Influencing Physical Activity. First, an overview of large national epidemiologic and surveillance data sets is offered, followed by a discussion on the use of market segmentation data to complement more traditional sources of data by adding new dimensions to our understanding of target groups and potential intervention strategies. The relative advantages and disadvantages of using each type of data are also given, as well as recommendations for further use. PMID- 12133739 TI - How the built environment affects physical activity: views from urban planning. AB - The link between the built environment and human behavior has long been of interest to the field of urban planning, but direct assessments of the links between the built environment and physical activity as it influences personal health are still rare in the field. Yet the concepts, theories, and methods used by urban planners provide a foundation for an emerging body of research on the relationship between the built environment and physical activity. Recent research efforts in urban planning have focused on the idea that land use and design policies can be used to increase transit use as well as walking and bicycling. The development of appropriate measures for the built environment and for travel behavior is an essential element of this research. The link between the built environment and travel behavior is then made using theoretical frameworks borrowed from economics, and in particular, the concept of travel as a derived demand. The available evidence lends itself to the argument that a combination of urban design, land use patterns, and transportation systems that promotes walking and bicycling will help create active, healthier, and more livable communities. To provide more conclusive evidence, however, researchers must address the following issues: An alternative to the derived-demand framework must be developed for walking, measures of the built environment must be refined, and more-complete data on walking must be developed. In addition, detailed data on the built environment must be spatially matched to detailed data on travel behavior. PMID- 12133740 TI - The association between urban form and physical activity in U.S. adults. AB - BACKGROUND: Physical inactivity is associated with multiple adverse health outcomes. Results from the transportation literature suggest that aspects of the urban environment may influence walking for transportation. In this paper we examine the association between a proxy measure of the urban environment and walking behavior. METHODS: We analyzed the association between home age and walking behavior in U.S. adults using data from the Third National Health and Nutrition Examination Survey. Logistic regression was used to estimate odds ratios and 95% confidence intervals and to control for the effects of gender, race/ethnicity, age, education level, household income, and activity limitations. RESULTS: Adults who lived in homes built before 1946 and from 1946 to 1973 were significantly more likely to walk 1+ miles > or =20 times per month than those who lived in homes built after 1973. This association was present among people living in urban and suburban counties, but absent among those living in rural counties. The association was also found in models that controlled for gender, race/ethnicity, age, education, income, and any health-related activity limitation. Other forms of leisure-time physical activity were not independently associated with home age. CONCLUSIONS: These results support the hypothesis that environmental variables influence walking frequency and suggest that home age may be a useful proxy for features of the urban environment that influence physical activity in the form of walking. Such proxy measures could facilitate testing ecologic models of health behavior using survey data. PMID- 12133741 TI - Intervention-related cognitive versus social mediators of exercise adherence in the elderly. AB - CONTEXT: Participation in regular physical activity is recognized as one of the most important health behaviors associated with the prevention of chronic disease and the promotion of health and well-being among the elderly. Although a number of cross-sectional studies have reported predictors of physical activity participation, few studies have assessed changes in intervention-related mediators associated with physical activity adherence in the elderly. OBJECTIVE: The purpose of this study was to compare the relative abilities of cognitive mediating variables (i.e., self-efficacy beliefs and outcome expectancies/realizations) versus a social mediating variable (i.e., exercise related social support) to examine mediators of a telephone-based, exercise counseling intervention on exercise adherence during months 7 to 12 of an exercise intervention. METHOD: Participants were 103 community-dwelling, healthy, sedentary, older adults (67 women and 36 men). Self-efficacy for exercise, outcome expectancies/realizations, and social support for exercise were assessed at baseline, 6 months, and 12 months. Participants received telephone-based exercise counseling to promote exercise adherence during the course of two 12 month exercise programs (i.e., aerobic/strength or flexibility exercises). RESULTS: Changes in cognitive mediators (i.e., self-efficacy and fitness outcome realizations) were associated with 7- to 12-month exercise adherence while exercise-related social support was not. CONCLUSION: Attention should be given to increasing confidence in the elderly to overcome barriers to exercise and achieve relevant fitness outcomes in exercise programs. PMID- 12133742 TI - Genetic factors in physical activity levels: a twin study. AB - BACKGROUND: Substantial interindividual variation is observed in sports participation and physical activity levels in youth. This study aimed to (1) estimate the relative contribution of genes, along with shared and nonshared environmental factors, to variation in sports participation index (SPI) and leisure-time physical activity (LTPA); and (2) test differences in those factors in males and females. METHODS: The sample was comprised of 411 Portuguese twin pairs of different zygosity aged 12 to 25 years. The SPI and LTPA were assessed with the Baecke questionnaire. Quantitative genetic modeling was used to test alternative models for the presence of additive gene effects (a(2)), common or shared environment within the family (c(2)), and unique environmental factors (e(2)). RESULTS: The best-fitting models showed sex-specific effects for the two phenotypes. Variance components for SPI in males were a(2)=68.4%, c(2)=20%, and e(2)=11.6%; and in females, a(2)=39.8%, c(2)=28.4%, and e(2)=31.8%. For variation in LTPA, genetic factors in males explained 63%, common environment was not significant, and unique environment explained 37%. In females, contributing factors were a(2)=32%, c(2)=38%, and e(2)=30%. CONCLUSIONS: Genetic effects explained a considerable amount of variation in SPI and LTPA, which were greater in males than in females. The relevance of shared environmental factors (family and peers) and nonshared environmental factors in SPI and LTPA is particularly evident in females. PMID- 12133743 TI - Delineating the impact of Tai Chi training on physical function among the elderly. AB - BACKGROUND: Through a re-analysis of a Tai Chi intervention data set, the study objective was to determine which, if any, subgroups of the study sample evidenced differential benefits from the intervention. METHOD: Re-analysis of a Tai Chi intervention study, a randomized controlled trial in Eugene and Springfield, Oregon. Physically inactive participants aged > or =65 years were randomly assigned to one of two groups: Tai Chi (n=49) and a wait-list control (n=45). The main outcome measure was self-reported physical function. RESULTS: Initial latent curve analyses indicated significant Tai Chi training effects: Participants in the Tai Chi group reported significant improvements in perceived physical function compared to those in the control group. However, there was significant interindividual variability in response to Tai Chi. The overall intervention effect was further delineated by identifying two subgroups. This delineation showed that Tai Chi participants with lower levels of physical function at baseline benefited more from the Tai Chi training program than those with higher physical function scores. Inclusion of additional measures of individual characteristics at baseline, change in movement confidence, and class attendance further explained differences in treatment responses. CONCLUSIONS: Findings from this study suggest that although an intervention may show an overall effect (or no overall effect), it may be differentially effective for subgroups of participants that differ in their pre-intervention characteristics. Examination of variability in outcome measures can provide important information for refining and tailoring appropriate interventions targeted to specific subgroups. PMID- 12133744 TI - Mediators of physical activity behavior change among women with young children. AB - BACKGROUND: Women with children are less likely to engage in adequate physical activity (PA) than women without children. This study aimed to evaluate the efficacy of two strategies for promoting increased PA among mothers of preschool aged children, and to explore the mediators of any resulting change in PA behavior. DESIGN: Controlled intervention trial incorporating repeated data collection from 554 women, randomized to one of three experimental conditions. Intervention Group 1 served as a control, while women in Groups 2 and 3 were given print information about overcoming PA barriers. Women in Group 3 were also invited to discuss the development of local strategies for the promotion of PA among mothers of young children. The primary strategies included increasing partner support, social advocacy, and capacity building, and were implemented through collaboration among participants, researchers, and community organizations. MAIN OUTCOME MEASURES: Adequate physical activity (PA), self efficacy (SE) and partner support (PS). RESULTS: Following the intervention, women in Group 3 were significantly more likely to meet guidelines for PA than controls (odds ratio [OR]=1.71, confidence interval [CI]=1.05-2.77)] after controlling for age and PA at baseline. After controlling for baseline PA, residualized change in SE (OR=1.86, CI=1.17-2.94) and PS (OR=2.29, CI=1.46-3.58) significantly predicted meeting guidelines. After controlling for residual change in PS and SE, the significant intervention effect was attenuated (Group 3 OR=1.40, CI=0.76-2.36), indicating that partner support and self-efficacy may be mediators of physical activity behavior change. CONCLUSIONS: The findings indicate that community participation approaches that facilitate increased self efficacy and partner support can be effective in increasing PA among mothers of young children. PMID- 12133747 TI - Risk factors for nephrolithiasis in patients with familial idiopathic hypercalciuria. AB - PURPOSE: About 40% of patients with nephrolithiasis have idiopathic hypercalciuria, sometimes associated with a family history of kidney stones. In these families, little is known about the frequency of, and risk factors for, stone formation among hypercalciuric patients. We therefore conducted a prospective study of 216 subjects from 33 families with idiopathic hypercalciuria. MATERIALS AND METHODS: We recorded the age, weight, and history of calcium stones in all subjects, and measured 24-hour urine volume and excretion of calcium, uric acid, sodium, magnesium, urea, citrate, phosphate, and sulfate on a nonrestricted diet. We performed a more complete metabolic evaluation in many of the hypercalciuric subjects (calciuria/weight >0.1 mmol/kg/d). Multivariate logistic regression analysis was performed to identify independent risk factors for stone formation. RESULTS: The prevalence of self reported nephrolithiasis was 46% (61/132) in hypercalciuric subjects and 11% (7/63) in normocalciuric subjects (P <0.0001). In multivariate analysis, age (odds ratio [OR] per 10 years of age = 1.3; 95% confidence interval [CI]: 1.1 to 1.6), urine calcium excretion (OR = 1.3 per mmol/d increase; 95% CI: 1.2 to 1.5), and uric acid excretion (OR = 3.3 per mmol/d increase; 95% CI: 1.4 to 7.5) were independent risk factors for nephrolithiasis. The risk of nephrolithiasis increased progressively with greater levels of hypercalciuria. CONCLUSION: We found a significant dose-effect association between calciuria and stone disease in patients with familial hypercalciuria. Other factors associated with stone formation included higher uric acid excretion, probably reflecting higher food intake, and age, probably reflecting the length of exposure to hypercalciuria and hyperuricosuria. PMID- 12133746 TI - Causes of death among women with human immunodeficiency virus infection in the era of combination antiretroviral therapy. AB - PURPOSE: To examine changes in the causes of death and mortality in women with human immunodeficiency virus (HIV) infection in the era of combination antiretroviral therapy. METHODS: Among women with, or at risk of, HIV infection, who were enrolled in a national study from 1994 to 1995, we used an algorithm that classified cause of death as due to acquired immunodeficiency syndrome (AIDS) or non-AIDS causes based on data from death certificates and the CD4 count. Poisson regression models were used to estimate death rates and to determine the risk factors for AIDS and non-AIDS deaths. RESULTS: Of 2059 HIV infected women and 569 who were at risk of HIV infection, 468 (18%) had died by April 2000 (451 HIV-infected and 17 not infected). Causes of death were available for 428 participants (414 HIV-infected and 14 not infected). Among HIV-infected women, deaths were classified as AIDS (n = 294), non-AIDS (n = 91), or indeterminate (n = 29). The non-AIDS causes included liver failure (n = 19), drug overdose (n = 16), non-AIDS malignancies (n = 12), cardiac disease (n = 10), and murder, suicide, or accident (n = 10). All-cause mortality declined an average of 26% per year (P = 0.03) and AIDS-related mortality declined by 39% per year (P = 0.01), whereas non-AIDS-related mortality remained stable (10% average annual decrease, P = 0.73). Factors that were independently associated with non-AIDS related mortality included depression, history of injection drug use with hepatitis C infection, cigarette smoking, and age. CONCLUSION: A substantial minority (20%) of deaths among women with HIV was due to causes other than AIDS. Our data suggest that to decrease mortality further among HIV-infected women, attention must be paid to treatable conditions, such as hepatitis C, depression, and drug and tobacco use. PMID- 12133748 TI - An independently derived and validated predictive model for selecting patients with myocardial infarction who are likely to benefit from tissue plasminogen activator compared with streptokinase. AB - BACKGROUND: In the Global Utilization of Streptokinase and tPA for Occluded coronary arteries (GUSTO) trial, patients with myocardial infarction who were treated with tissue plasminogen activator (tPA) had a 6.3% 30-day mortality, compared with a mortality of 7.3% among those treated with streptokinase, despite a greater risk of intracranial hemorrhage with tPA. However, in part because of its higher cost, tPA has not been adopted universally. METHODS: Using an independently developed model, we predicted the benefits of tPA therapy in the 24,146 patients in the GUSTO trial and compared these predictions with the actual benefits of tPA, after classifying patients by their risks of mortality and intracranial hemorrhage. We also performed a "patient-specific" cost effectiveness analysis among different strata of expected benefit of tPA. RESULTS: Our model predicted that among patients with myocardial infarction, 61% of the benefit of tPA use in reducing mortality accrued to only 25% of patients; treating half of patients could capture 85% of the benefit. Including the risk of intracranial hemorrhage, our model predicted that treating half the GUSTO patients with tPA and the others with streptokinase would yield similar outcomes as treating all patients with tPA, because the additional risk of intracranial hemorrhage exceeded the expected benefit in some patients. When patients were stratified into quartiles of risk, the observed outcomes in the GUSTO patients corresponded well with these predicted results. The estimated cost-effectiveness of tPA was sensitive to patient characteristics. CONCLUSION: For selected patients, use of tPA yields substantially better outcomes than streptokinase, and use of the less expensive agent is difficult to justify. For many patients, however, tPA is unlikely to provide any additional benefit and, in some patients, it may even cause net harm. PMID- 12133749 TI - Effectiveness of enzyme replacement therapy in 1028 patients with type 1 Gaucher disease after 2 to 5 years of treatment: a report from the Gaucher Registry. AB - PURPOSE: Gaucher disease is the first lysosomal storage disorder to be treated with macrophage-targeted enzyme replacement therapy. Previous studies in relatively small numbers of patients demonstrated short-term efficacy of this treatment. This study describes the effects of 2 to 5 years of treatment on specific manifestations of type 1 Gaucher disease. SUBJECTS AND METHODS: Physicians reported data from 1028 patients to the Gaucher Registry. Assessment of response included serial measurements of hemoglobin concentration, platelet count, liver and spleen volumes, and the occurrence of bone pain and bone crises. RESULTS: Among anemic patients, hemoglobin concentration increased to normal or near normal within 6 to 12 months, with a sustained response through 5 years. In thrombocytopenic patients with intact spleens, the most rapid response occurred during the first 2 years, with slower improvement thereafter. The likelihood of achieving a normal platelet count decreased with increasing severity of baseline thrombocytopenia. In patients who had undergone splenectomy, platelet counts returned to normal within 6 to 12 months. Hepatomegaly decreased by 30% to 40% during follow-up; splenomegaly decreased 50% to 60%, but rarely to volumes below five times normal size. In patients with pretreatment bone pain or bone crises, 52% (67/128) were pain free after 2 years and 94% (48/51) reported no additional crises. CONCLUSION: Enzyme replacement therapy prevents progressive manifestations of Gaucher disease, and ameliorates Gaucher disease-associated anemia, thrombocytopenia, organomegaly, bone pain, and bone crises. PMID- 12133750 TI - The effect of cephalosporin resistance on mortality in adult patients with nonmeningeal systemic pneumococcal infections. AB - PURPOSE: To evaluate the clinical relevance of cephalosporin (ceftriaxone/cefotaxime) resistance among patients with nonmeningeal systemic pneumococcal infection. SUBJECTS AND METHODS: From January 1994 to October 2000, we prospectively studied 522 episodes of nonmeningeal systemic pneumococcal infections (448 pneumonias) in 499 adults who were treated according to hospital guidelines. In vitro antibiotic susceptibility, as the minimum inhibitory concentration (MIC), was determined by microdilution method. The MIC methods and breakpoints (cutoffs) were established by the National Committee for Clinical Laboratory Standards. RESULTS: Of the 522 pneumococcal strains, 413 strains (79%) were susceptible to ceftriaxone/cefotaxime, MIC < or =0.5 microg/mL; 79 (15%) were intermediate, MIC = 1 microg/mL; and 30 (6%) were resistant, MIC = 2 microg/mL. After adjusting for several variables, including pneumococcal serogroups/serotypes, infections due to nonsusceptible (intermediate and resistant) pneumococcal strains were independently associated with prior antibiotic therapy, with an odds ratio of 5.9 (95% confidence interval: 2.6 to 13.6). Thirty-day mortality among the 185 patients who were treated with ceftriaxone (1 g/d) or cefotaxime (1.5 g every 8 hours) did not differ by cephalosporin susceptibility: 18% (26/148) among those with susceptible organisms, 13% (3/24) with intermediate organisms, and 15% (2/13) in resistant cases (P = 0.81). CONCLUSION: Ceftriaxone or cefotaxime were effective in treating patients with nonmeningeal systemic pneumococcal infections caused by strains with MIC < or =2 microg/mL. These results support the newly established ceftriaxone/cefotaxime MIC breakpoints (cutoffs) for nonmeningeal pneumococcal infections. PMID- 12133751 TI - Assessment of congenital coronary artery fistulas by transesophageal color Doppler echocardiography. AB - PURPOSE: Coronary angiography is the gold standard for imaging the coronary tree, but the relation of coronary artery fistulas to other structures, and their origin and course, may not be apparent. We evaluated the ability of multiplane color Doppler transesophageal echocardiography to identify coronary fistulas. PATIENTS AND METHODS: Twenty-one patients with angiographically confirmed coronary artery fistulas were investigated by transesophageal echocardiography in four Italian hospitals between January 1997 and May 2001. RESULTS: Transesophageal echocardiography correctly diagnosed fistulous connection in all 21 patients. This included 6 patients with connections from the left circumflex artery (into the right chambers of the heart in 5 patients, and into the left ventricle in 1 patient), 10 patients with a fistula arising from the left anterior descending artery or left main coronary artery (with drainage into the right ventricle or main pulmonary artery), and 5 patients with a fistula from the right coronary artery (with drainage sites in the lateral aspect of the right ventricle, the low posterior right atrium, or the superior vena cava). In 4 of the 21 patients, angiography did not identify the precise site of a fistula into the coronary sinus or right ventricle. CONCLUSION: Color Doppler transesophageal echocardiography is useful in the diagnosis and in the precise localization of coronary artery fistulas. PMID- 12133752 TI - Low Fcgamma receptor III and high granulocyte colony-stimulating factor serum levels correlate with the risk of infection in neutropenia due to Felty's syndrome or systemic lupus erythematosus. AB - PURPOSE: To determine whether serum levels of soluble Fcgamma receptor III and granulocyte colony-stimulating factor (G-CSF) are associated with the risk of infection in patients with neutropenia due to Felty's syndrome or systemic lupus erythematosus. SUBJECTS AND METHODS: Serum levels of G-CSF and soluble Fcgamma receptor III were measured by enzyme-linked immunosorbent assays in 13 patients with neutropenia due to Felty's syndrome, 10 patients with neutropenia due to systemic lupus erythematosus, and 41 controls with normal leukocyte counts (25 with systemic lupus erythematosus, 16 with rheumatoid arthritis). We calculated the area under the receiver operating characteristic (ROC) curves for the absolute neutrophil count, soluble Fcgamma receptor III levels, and G-CSF levels. RESULTS: Nine of the neutropenic patients (7 with Felty's syndrome, 2 with lupus) had one or more infections within 3 months before and after blood samples were obtained. Absolute neutrophil counts were similar in neutropenic patients who did or did not have infections. However, the median level of soluble Fcgamma receptor III (63 vs. 126 arbitrary units, P = 0.005) was significantly lower among patients who developed infections, whereas the median level of G-CSF (90.9 vs. 53.3 pg/mL, P = 0.04) was significantly higher compared with patients without infections. The area under the ROC curve was 0.58 (P = 0.49) for the absolute neutrophil count, 0.84 (P = 0.007) for soluble Fcgamma receptor III levels, and 0.73 (P = 0.03) for G-CSF levels. CONCLUSION: In patients with chronic neutropenia due to rheumatic diseases, low soluble Fcgamma receptor III levels and elevated G-CSF levels are better indicators of the risk of infection than is the neutrophil count. PMID- 12133753 TI - The epidemiology of peripheral vein infusion thrombophlebitis: a critical review. AB - We critically assessed studies on the clinical importance, diagnosis, incidence, and pathogenesis of peripheral vein infusion thrombophlebitis, including catheter related and patient-related risk factors. We reviewed the evidence linking thrombosis, particularly prothrombotic states such as the inherited thrombophilic disorders, with peripheral vein infusion thrombophlebitis. Peripheral vein infusion thrombophlebitis occurs in 25% to 35% of hospitalized patients with peripheral intravenous catheters and has both patient-related implications (e.g., sepsis) and economic consequences (e.g., extra nursing time). Although duration of catheterization, catheter-related infection, and catheter material are important risk factors for peripheral vein infusion thrombophlebitis, patient related risk factors are not well elucidated. PMID- 12133754 TI - Penicillin-resistant Streptococcus pneumoniae septic shock and meningitis complicating chronic graft versus host disease: a case report and review of the literature. PMID- 12133755 TI - Paradoxical intracranial cryptococcoma in a human immunodeficiency virus-infected man being treated with combination antiretroviral therapy. PMID- 12133756 TI - Cases from the osler medical service at Johns Hopkins University. AB - PRESENTING FEATURES: A 29-year-old woman with a history of rheumatic heart disease and one episode of endocarditis as an adolescent was admitted to the hospital after 1 week of headache, fever, and myalgia. Her past medical history was otherwise unremarkable and did not include illicit drug use. On physical examination, she had a previously noted 3/6 holosystolic murmur at the apex, which radiated to her back; a previously noted 1/4 diastolic murmur at the right upper sternal border; diminished strength in her right upper extremity; multiple painful erythematous nodules on her fingers (Figure 1); and red streaks under her nails (Figure 2). Magnetic resonance imaging of the brain demonstrated multiple lesions; the largest was in the right frontal lobe with associated hemorrhage (Figure 3). What is the diagnosis? PMID- 12133757 TI - Putting evidence into practice. PMID- 12133758 TI - Treatment of menstruation-associated recurrence of hereditary pancreatitis with pharmacologic ovarian suppression. PMID- 12133759 TI - Retroperitoneal fibrosis due to Wegener's granulomatosis: a misdiagnosis as tuberculosis. PMID- 12133760 TI - Recent-onset tuberculous pleurisy presenting as pseudochylothorax. PMID- 12133761 TI - Partial anomalous pulmonary venous drainage of the superior left pulmonary vein into the innominate vein resulting in right ventricular failure. PMID- 12133762 TI - Helicobacter pylori associated with idiopathic thrombocytopenic purpura. PMID- 12133763 TI - Alopecia associated with atorvastatin. PMID- 12133764 TI - Educational assessment guidelines: a Clerkship Directors in Internal Medicine commentary. PMID- 12133792 TI - Of mice and men: species variations of Toll-like receptor expression. AB - Toll-like receptors (TLRs) comprise a family of evolutionary conserved pattern recognition molecules that have an essential role in mammalian innate immune defense. Recent observations suggest that several TLR orthologues are expressed differently in mice and humans - variations among both species include the expression of TLR transcripts in different cell types and different transcription regulation on cellular activation. Apparently some TLR genes have been placed into a completely different regulatory context during evolution. It will be interesting and important to clarify whether the observed regulatory differences influence innate immune responses in a species-specific manner. PMID- 12133793 TI - New insights into the role of 4-1BB in immune responses: beyond CD8+ T cells. AB - Since the discovery of 4-1BB 15 years ago, the receptor and its ligand (4-1BBL) have remained enigmatic molecules because of their highly regulated pattern of expression. Classically, 4-1BB is known to function as a costimulator for T cells and as a potent survival factor for CD8+ T cells. Recent studies highlight the participation of 4-1BB and its ligand 4-1BBL in a more complex network of immune cell responses and suggest that intervening in the 4-1BB costimulatory pathway might have some potential as a therapeutic approach to immune disorders. PMID- 12133794 TI - Cytokines as natural adjuvants for vaccines: where are we now? PMID- 12133795 TI - Using immunotherapy to treat rheumatic diseases. PMID- 12133796 TI - MHC haplotypes, MICA genes and the 'wonderland' of NK receptor polymorphism. PMID- 12133801 TI - One for all and all for one: thymic epithelial stem cells and regeneration. AB - It has long been believed that the thymic epithelial microenvironment originates from both the endodermal and ectodermal germ cell layers. However, a growing body of evidence indicates that such a dual origin is not the case, and that the diverse thymic epithelial populations all develop from a common epithelial stem cell. This article explores these data, investigates the identity of such cells and the signals that might control their expansion and differentiation, and considers the possibility of stem cell transplantation for thymic regeneration. PMID- 12133802 TI - On the generation of allergic airway diseases: from GM-CSF to Kyoto. AB - The sharp increase in the prevalence of asthma over the past three decades suggests an important contribution of environmental factors in the generation of this disease, and compels a search for molecular pathways by which such factors could facilitate Th2 immune-inflammatory airway responses; granulocyte-macrophage colony-stimulating factor (GM-CSF) might be one such signal. In this review, we appraise the evidence with respect to the presence of GM-CSF in asthma, the roles played by GM-CSF in these immune responses and environmental triggers that can induce GM-CSF expression. Further, we propose a paradigm that unites these divergent observations, and postulate that GM-CSF produced in response to environmental agents can establish an airway microenvironment that promotes the initiation, influences the evolution and supports the maintenance of an aeroallergen-specific adaptive Th2 immune response. PMID- 12133803 TI - GM-CSF in inflammation and autoimmunity. AB - Granulocyte macrophage-colony stimulating factor (GM-CSF) is now best viewed as a major regulator governing the functions of granulocyte and macrophage lineage populations at all stages of maturation. There is recent evidence for a key role for GM-CSF in inflammatory and autoimmune diseases, therefore making it worthy of consideration for targetting. Such evidence includes disease exacerbation following its administration and amelioration of disease in animal models by GM CSF gene targetting or by anti-GM-CSF antibody blockade. The interdependence of GM-CSF formation and that of the important proinflammatory cytokines, interleukin 1 and tumour necrosis factor-alpha (TNF-alpha), is discussed, as is the greater disease suppression found in arthritis models following GM-CSF depletion compared with that observed in the absence of TNF-alpha. PMID- 12133804 TI - Molecular coordination of alphabeta T-cell receptors and coreceptors CD8 and CD4 in their recognition of peptide-MHC ligands. AB - The interaction of the alphabeta T-cell receptor (TCR) with its ligand, peptide MHC (pMHC), is enhanced by the recognition of the coreceptor CD8 or CD4 to the same pMHC in the immunological synapse. In the past few years, the coordination of these interactions at the molecular level has been revealed by analysis of their complex crystal structures and binding dynamics. Here we discuss the interactions of pMHC with the TCR and coreceptor CD8 or CD4 on the surfaces of alphabeta T cells and antigen presenting cells, and the implications for TCR signalling and the T-cell repertoire. PMID- 12133805 TI - It's all Rel-ative: NF-kappaB and CD28 costimulation of T-cell activation. AB - Costimulatory signals complement or modify the signals provided to a lymphocyte through antigen receptors. For productive T-cell activation, the CD28 molecule is apparently the most important, although not the only, costimulatory receptor. CD28 can provide a signal that is at least partially distinct from that delivered by the T cell receptor (TCR)-CD3 complex. Several lines of evidence indicate that the nuclear factor (NF)-kappaB pathway is perhaps the most relevant biochemical or transcriptional target for the costimulatory activity of CD28. Although many questions remain, recent years have witnessed significant progress in understanding the signal transduction pathways leading from the TCR and CD28 to Rel/NF-kappaB-dependent transcription. PMID- 12133807 TI - Structural attributes in the conjugation of ubiquitin, SUMO and RUB to protein substrates. AB - Many cellular and secreted proteins are chemically modified after their translation is completed. The covalent linkage of a polypeptide chain (modifier) to a substrate protein is a special case of post-translational modification. In the late seventies it was observed that ubiquitin, a small modifier, marks short lived proteins for degradation by the 26S proteasome. Over the last decade many other ubiquitin-related proteins were discovered and isolated. Attachment of polypeptide chains onto acceptor molecules became a common feature to regulate spatially and timely organized cellular pathways of proteins. This article focuses on the structures of the three modifiers: ubiquitin, RUB and SUMO and the cognate enzymes involved in these modification pathways. We have described the homologies and differences of these proteins and indicate salient topological hallmarks common to modifier-conjugating enzymes. This characterization will help in understanding these regulatory pathways and their similarities and differences in controlling protein fate, from protein degradation signals generated by polyubiquitination to functional modification brought about by RUB and SUMO conjugation. PMID- 12133808 TI - The role of SSeCKS/gravin/AKAP12 scaffolding proteins in the spaciotemporal control of signaling pathways in oncogenesis and development . AB - Scaffolding proteins are thought to facilitate the efficiency and specificity of enzyme/substrate reactions by coordinating their interaction along a cytoskeletal infrastructure in a spacial and temporal manner. Rodent SSeCKS, and its human orthologue, Gravin, seem to function as tumor suppressors and regulators of mitogenesis, inflammatory response, development and differentiation via novel scaffolding functions involving the selective binding of key G1-->S phase signaling proteins such as protein kinase C (PKC), PKA, calmodulin, cyclins and beta-adrenergic receptors. The association of SSeCKS/Gravin with the actin-based cytoskeleton as well as to plasma membrane sites via N-terminal myristylation places these proteins at the junction of signaling and cytoskeletal pathways. The following review describes the regulatory and scaffolding functions of SSeCKS and Gravin and how mitogen-induced phosphorylation modulates their ability to regulate cell adhesion, signaling, mitogenesis and oncogenesis. PMID- 12133809 TI - Extracellular-peptide control of competence for genetic transformation in Streptococcus pneumoniae. AB - Bacteria, which often are subjected to fluctuations in nutrients, temperature, radiation, pH, etc., adapt to the physico-chemical environment they live in by making the appropriate changes in their gene expression patterns. During the last decades it has become increasingly clear that bacteria, in addition, have a "social life", and that changes in gene expression can also be elicited by the presence of other bacteria. Traditionally bacteria have been viewed as solitary organisms that in general do not interact with other bacteria in a coordinated manner. Recent advances in the field of bacterial cell-to-cell communication has proved this to be a misconception, and mounting evidence now show that bacterial group behaviour is ubiquitous in nature. Competence for natural genetic transformation in Streptococcus pneumoniae, which has been studied for more than seventy years, has become a paradigm for intercellular communication and cell density dependent regulation of gene expression in Gram-positive bacteria. There has been rapid progress recently in elucidating the molecular mechanisms behind regulation of natural competence in S. pneumoniae. In this review, we describe the current status of our knowledge of natural competence in this bacterium, with particular emphasis on the early phase of competence induction. PMID- 12133810 TI - Novel trends in orphan market drug discovery: amyotrophic lateral sclerosis as a case study. AB - As new lead discovery technologies of high throughput screening and rational drug design have been incorporated into pharmaceutical and biotechnology drug discovery programs, researchers have focused on the applying these new technologies in diseases traditionally neglected by for-profit drug discovery efforts. This article reviews general trends in orphan disease lead discovery, identifies best practices of orphan market drug discovery and provides an overview of recent ALS lead discovery programs and drug development according to these metrics. Best practices in orphan market drug discovery embodied by programs like the NIH Anticonvulsant Screening Program include the (1) management of timelines and priorities, (2) engagement of for-profit partners, (3) creative application of technology, (4) collaboration, and (5) flexibility. Recent trends in ALS lead discovery have been shaped not only by the predominance of animal models of disease over in vitro models, but also by the successes and best practices of these earlier orphan market drug discovery programs. The ALS Treatment Initiative, the Johns Hopkins Center for ALS Research, the ALS Association, and the ALS Therapy Development Foundation have all initiated lead discovery programs in the past several years which seek to utilize existing experimental models of the disease and challenge assumptions about the linear nature of the lead discovery and development process. The compounds currently in clinical evaluation for ALS were identified as leads from a variety of sources, further reinforcing the transforming effect these new lead discovery programs have had on drug discovery and development in ALS. We conclude our review with an overview of the challenges and opportunities lead discovery in ALS currently faces, ultimately concluding that ALS lead discovery, and indeed orphan market drug discovery in general, would most benefit from more centralized lead discovery management, expanded national access to core facilities for lead discovery, and matrixed simultaneous screening of multiple compounds for multiple neglected diseases. PMID- 12133811 TI - Isoforms of apolipoprotein E can modulate tPA-induced clot lysis in vitro. AB - Apo E, and its respective isoforms, have been linked to outcome and survival in cerebral vascular and cardiovascular diseases. The effectiveness of intravenous tPA in patients with acute ischemic stroke appears to be enhanced in patients who have an Apo E2 phenotype. The ability of Apo E isoproteins (endogenous Apo E isoproteins or exogenous Apo E isoproteins) to modulate tPA-induced clot lysis in vitro was assessed using an in vitro clot assay system. Blood samples were obtained from 18-volunteers with three Apo E genotypes: E2, E3 and E4. tPA induced clot lysis (0-4 microgram/ml tPA), was assessed in the presence or absence of supplemental Apo E2, E3 or E4 (9.8 microgram/ml). tPA-induced clot lysis was significantly (P equal or less than 0.0001) enhanced by supplementation with Apo E2 (EC50 0.20 0.06 microgram/ml) as compared to tPA alone (0.72 0.19). Apo E4 supplementation caused a significant (P < or = 0.05) inhibition of clot lysis (0.98 0.23), but there was no significant change caused by Apo E3. The genotype of the volunteer did not significantly affect the ability of the supplemental Apo E from modulating tPA-induced clot lysis. We conclude that the administration of Apo E isoproteins can modulate clot lysis in vitro. Our results suggest that the Apo E isoprotein may have an impact on clot dissolution and the effectiveness of thrombolytic therapy. PMID- 12133813 TI - Modulation of aromatase activity and expression by environmental chemicals. AB - Aromatase is the enzyme that converts androgen to estrogen. Our laboratory has proposed and demonstrated that aromatase is an important target of some environmental chemicals. We have found that some of these compounds inhibit aromatase activity, resulting in a decrease in the level of estrogen or an increase in the level of androgen in cells. Environmental chemicals can also modify the expression of aromatase in various tissues, resulting in a change in the ratio between androgen to estrogen. The compounds that inhibit aromatase or suppress aromatase expression will behave as antiestrogens or androgen-like compounds in vivo. On the other hand, compounds that increase aromatase expression or enhance aromatase activity (or stability) may function as anti androgens or estrogen-like compounds. This review will summarize and discuss findings from this and other laboratories on the effects of environmental chemicals on aromatase activity and expression. PMID- 12133812 TI - Global regulation of virulence determinants in Staphylococcus aureus by the SarA protein family. AB - In S. aureus, the production of virulence determinants such as cell wall adhesins and exotoxins during the growth cycle is controlled by global regulators such as SarA and agr. Genomic scan reveals 16 two-component regulatory systems (e.g. agr and sae) as well as a family of SarA homologs in S. aureus. We call the SarA homologs the SarA protein family. Many of the members in this protein family are either small basic proteins (<153 residues) or two-domain proteins in which a single domain shares sequence similarity to each of the small basic proteins. Recent crystal structures of SarR and SarA reveal dimeric structures for these proteins. Because of its structure and unique mode of DNA binding, SarR, and possibly other SarA family members, may belong to a new functional class of the winged-helix family, accommodating long stretch of DNA with bending points. Based on sequence homology, we hypothesize that the SarA protein family may entail homologous structures with similar DNA-binding motifs but divergent activation domains. An understanding of how these regulators interact with each other in vivo and how they sense environmental signals to control virulence gene expression (e.g. alpha-hemolysin) will be important to our eventual goal of disrupting the regulatory network. PMID- 12133814 TI - Apoptosis in gliomas, and its role in their current and future treatment. AB - Apoptosis has recently entered the spotlight in the continuing search for new therapeutic approaches to cancer because it plays a twofold role in this disease. As stated by Lowe and Lin: "(M)ost cytotoxic anticancer agents induce apoptosis.(and so) the same mutations that suppress apoptosis during tumor development also reduce treatment sensitivity" (1). Therefore, any strategy aimed at increasing the propensity of glioma cells to undergo apoptosis could be therapeutic in its own right, but has the added potential of enhancing their sensitivity to other, established, treatments. As a corollary, understanding apoptotic mechanisms at the molecular level will not only help to explain why gliomas arise, but also identify points of intervention. This review will focus on these points, with emphasis on two families of apoptotic molecules, death ligands and their receptors, and BCL-2 family proteins. Near-term strategies of how apoptosis can be exploited therapeutically are discussed. PMID- 12133815 TI - Promoter competition assay for analyzing gene regulation in joint tissue engineering. AB - We describe a new biochemical technique, "promoter competition assay," for examining the role of cis-acting DNA elements in tissue cultures. Recent advances in tissue engineering permit the culture of a variety of cells. Many tissues are engineered, however, without an appropriate understanding of molecular machinery that regulates gene expression and cellular growth. For elucidating the role of cis-acting regulatory elements in cellular differentiation and growth, we developed the promoter competition assay. This assay uses a transient transfer into cells of double-stranded DNA fragments consisting of cis-acting regulatory elements. The transferred DNA fragments act as a competitor and titrate the function of their genomic counterparts. Using synovial cells derived from a rheumatoid arthritis patient, we examined a role of NF-kappa B binding sites in the regulation of the expression of matrix metalloproteinase (MMP) genes. The results support a stimulatory role of NF-kappa B in transcriptional regulation of MMP-1 and MMP-13. PMID- 12133816 TI - The necessity of combining genomic and enzymatic data to infer metabolic function and pathways in the smallest bacteria: amino acid, purine and pyrimidine metabolism in Mollicutes. AB - Bacteria of the class Mollicutes have no cell wall. One species, Mycoplasma genitalium is the personification of the simplest form of independent cell-free life. Its small genome (580 kbp) is the smallest of any cell. Mollicutes have unique metabolic properties, perhaps because of their limited coding space and high mutability. Based on 16S rRNA analyses the Mollicutes Mycoplasma gallisepticum is thought to be the most mutable bacteria. Enzyme activities found in most Bacteria are absent from Mollicutes. The functions of apparently absent genes and enzymes can apparently be fulfilled by other genes and their expression products that have multiple capabilities. Because of these and other properties predictions of their metabolism based only on, e.g., either annotation, enzymatic assay, proteomic studies or structural analyses is problematic. To obtain a more confident appraisal of the functional capabilities of these simplest cells genomic and enzymatic data were combined to obtain a "metabolic consensus". The consensus is represented by a biochemical circuit for central metabolism involving purine and pyrimidine interconversions and their linkages to amino acid metabolism, glycolysis and the pentose phosphate pathway in three human Mollicutes pathogens: Mycoplasma pneumoniae, Mycoplasma genitalium and Ureaplasma urealyticum. PMID- 12133817 TI - PDGF and signal transduction in hepatic stellate cells. AB - Platelet-derived growth factor (PDGF) is one of the most potent mitogen for cultured HSC isolated from rat, mouse, or human liver. Phosphotyrosines on the activated PDGF receptor operate as high affinity binding sites for several molecules involved in the downstream propagation of the signal, including the sequential activation of Raf-1, MEK and extracellular-signal regulated kinase (ERK). Nuclear translocation of ERK is associated to the phosphorylation of several transcription factors, including Elk-1 and SAP, and represents an absolute requirement for triggering a proliferative response. Phosphatidylinositol 3-kinase (PI 3-K), is another molecule recruited by the activated PDGF receptor. In human HSC cultures, PI 3-K activation is necessary for both mitogenesis and chemotaxis induced by PDGF. In addition, PI 3-K is involved in the activation of the Ras-ERK pathway in human HSC, although it is not strictly necessary, since established PI 3-K inhibitors inhibit ERK activation only by 40-50%. Therefore, in HSC, PI 3-K regulates PDGF-related mitogenesis and cell migration by pathways that are at least in part independent of ERK activation. Accumulated evidence indicates that the induction of replicative competence by PDGF is dependent on the maintenance of sustained increase in [Ca2+]i due to calcium entry rather than from the release from intracellular stores. In addition, stimulation with PDGF increases the activity of the Na+/H+ exchanger in rat or human HSC with consequent sustained changes in intracellular pH. PMID- 12133818 TI - Molecular mechanisms of pulmonary fibrosis. AB - Pulmonary fibrosis is the end-point of a numerous and heterogeneous group of disorders known as interstitial lung diseases (ILD). Lung fibrotic remodeling is characterized by fibroblast/myofibroblast activation, and excessive extracellular matrix accumulation leading to progressive organ dysfunction and usually terminal outcome. Treatment is largely ineffective primarily because few of the molecular mechanisms have been well defined to design appropriate targets for therapy. While the pathogenesis is incompletely understood, a growing body of evidence suggests two different pathogenic routes for developing pulmonary fibrosis. The inflammatory pathway, where a shift to the so-called T-helper 2 type cytokine networks is critical, and the epithelial pathway represented by idiopathic pulmonary fibrosis, by far the most aggressive ILD. In this pathway the inflammatory process is irrelevant, and the physiopathology seems to be dominated by epithelial cell injury and activation. Both routes may trigger a number of cytokines/growth factors inducing fibroblast migration/proliferation and phenotype change to myofibroblasts, with a consequent accumulation of extracellular matrix. An imbalance in matrix metalloproteinase/tissue inhibitors of metalloproteinases may contribute to alteration in extracellular matrix turnover and remodeling. This review will focus in some of the mechanisms involved in both prefibrotic pathways, as well as those involved in fibroblast activation and abnormal matrix deposition. PMID- 12133819 TI - Recent progress in Bacillus subtilis two-component regulation. AB - Two-component regulatory systems serve to control gene expression in response to environmental and physiological changes. They are widespread among a variety of organisms and most often found in prokaryotes. One of the gram-positive microorganisms Bacillus subtilis is a well-studied bacterium whose complete nucleotide sequence has been determined. Thus, it is now possible to study transcription of the whole genome with microarray analysis. In this review we summarize the recent progress in B. subtilis two-component regulatory systems by describing the known systems and those for which the function was recently assigned. Also included is an attempt to construct a partial transcriptional network involving several two-component systems. The studies described here are based on the data from traditional genetics and biochemistry, and from microarray analysis of 29 two-component systems. PMID- 12133820 TI - Lipoprotein metabolism in the nephrotic syndrome. AB - This review covers lipids, apolipoproteins, and receptors involved in the dyslipidemia of the nephrotic syndrome in humans and in rat or mouse models of the syndrome. It emphasizes research published during the last decade, though earlier work is cited. The focus is on the biosynthesis and catabolism of the plasma lipoprotein density classes and the role of receptors and enzymes in regulating lipoprotein metabolism in nephrosis. Although the factors responsible for the initiation of the hepatic and peripheral cellular responses to proteinuria and hypoalbuminemia remain elusive, recent work highlights the increased risk of atherosclerosis and the progression of renal disease associated with nephrotic dyslipidemia. Understanding of the role of the kidney in the catabolism of apolipoproteins entering the glomerular filtrate has been enhanced by the discovery of the receptor-mediated uptake of apolipoprotein A-I, the main apoprotein of HDL. The following aspects of lipid and lipoprotein metabolism in relation to nephrosis are discussed, with attention paid to differences between experimental nephrosis and the human nephrotic syndrome:(1) Albumin metabolism (2) Lipoprotein metabolism (3) Receptors (4) LCAT and CETP (5) Hepatic and Lipoprotein Lipase (6) Lipid metabolism (7) Lipiduria (8) Hypotheses and Questions (9) Summary. PMID- 12133822 TI - Bacterial group II introns and their association with mobile genetic elements. AB - Group II introns are an abundant class of self-splicing RNAs, found primarily in the organelles of plants and lower eukaryotes and in bacteria. The first bacterial group II intron identified to be functional for splicing in vivo was the Ll.ltrB intron of Lactococcus lactis. It has served as an excellent model for the study of group II intron structure and function. Taking advantage of the tools of bacterial genetics and biochemical methodologies, details of Ll.ltrB splicing and homing reactions have been elucidated and are similar to those of fungal group II introns. This review provides a summary of these results. Of particular interest is the potential use of Ll.ltrB as an agent for targeted gene disruption. In addition, the development of a genetic system to analyze Ll.ltrB splicing promises to provide new insight into group II intron structure and function. Identification and analysis of group II introns in other bacterial species is a continuing process, and a discussion of published reports on these introns is provided here. Limited functional data is available for most of these introns, but sequence analysis points out several common themes, most notably that bacterial group II introns are almost always carried on mobile genetic elements. PMID- 12133821 TI - Alcohol-mediated polarization of type 1 and type 2 immune responses. AB - Immune responses of alcoholics are often compromised, placing them at increased risk for frequent and severe infections. We demonstrate, using a murine model that parallels human alcoholism, that ethanol consumption polarizes adaptive immune responses by CD4+ T helper lymphocytes (Th). Alcohol impairs Th1-regulated cell-mediated, although Th2-regulated humoral responses are largely unimpaired and may be enhanced. Ethanol's effect is most pronounced during the early or cognitive phase of the immune response, when antigen-presenting cells (APC) interact with T cells. We find that alcohol does not act directly upon T cells, but upon APC. Consequences of this interaction of alcohol with APC in vivo are diminished Th1-mediated delayed hypersensitivity (DTH) reactions, while at the same time increased Th2-regulated serum IgE levels are seen. Further ethanol consumption leads to decrease affinity of the IgG2a and IgG2b Th1-regulated antibody isotypes. PMID- 12133823 TI - Expression microarray analysis of brain tumors: what have we learned so far. AB - This review covers the emerging field of expression microarray technology as applied to human brain tumors. Dual and single color techniques are described and contrasted, and the importance of proper handling of the starting material is emphasized. Difficulties with data interpretation are described and current approaches to cluster analysis reviewed. Microarray studies of general importance or specifically pertaining to brain tumors, published in the initial few years of this technology, are summarized. Although this technology is still in its infancy, microarray has distinguished prognostic groups within medulloblastomas and separated medulloblastomas from morphologically identical supratentorial PNETs. Differential expression of a number of genes previously known to be involved in the pathogenesis of brain tumors has been confirmed. These genes include EGFR, VEGF, transcription factor AP-2, insulin growth factor binding proteins 3 and 5, matrix metalloproteinases, tissue inhibitors of matrix metalloproteinases, CD44, basic fibroblast growth factor, and cathepsin H. Finally, novel roles for a few genes, including insulin growth factor binding protein 2 and apolipoprotein D, have been revealed for the first time by expression microarrays. PMID- 12133824 TI - Control of gene expression in Gram-positive bacteria: extensions of and departures from enteric paradigms. AB - Our introduction to prokaryotic gene expression has always focused on the operon and regulatory mechanisms that operate within enteric bacteria such as Escherichia coli, Salmonella species, and their phages. While operon organization and many of the components of regulatory networks are conserved in Gram-positive species, there exists unique features that set these organisms apart from the enterics. Two examples are presented herein: carbon catabolite control and regulation of RNA polymerase sigma subunit activity, are presented. The accompanying reviews highlight the diversity and novel aspects of genetic control in Gram-positive bacteria, with descriptions of quorum-sensing systems, transcriptional control, and RNA processing mechanisms. PMID- 12133825 TI - Quantitative relationships between ryanoids, receptor affinity and channel conductance. AB - The review examines the relationship between the structure of several ryanodine analogs and (A) binding, (B) channel conductance, and (C) ligand binding kinetics. Comparative molecular field analysis (CoMFA) and comparative molecular similarity analysis (CoMSIA) are used to quantitatively assign structural correlations. Hydrogen bond donating (but not accepting) ability was found to be highly correlated with ligand affinity. Analysis of the correlation between hydrophobicity and ligand affinity indicates that, in general, deviation from the amphipathic nature of ryanodine weakens binding. Affinities and binding kinetics obtained in vivo are comparable to those obtained in the less-than-physiological in vitro conditions. Therefore, the structure-activity relationships surveyed are relevant to the living cell. The review presents arguments favoring the propositions that (A) the pyrrole is a major factor orienting the ligand in the receptor binding site and (B) that ryanoids alter ryanodine receptor function through allosteric mechanisms. PMID- 12133826 TI - Differential localization of the vacuolar H+ pump with G subunit isoforms (G1 and G2) in mouse neurons. AB - Vacuolar H(+)-ATPases (V-ATPases), a family of multimeric proton pumps, are involved in a wide variety of physiological processes. We have identified two mouse genes, Atp6g1 and Atp6g2, encoding the G1 and G2 isoforms of the V-ATPase G subunit, respectively. G1 was distributed ubiquitously in the tissues examined, whereas G2 was specifically distributed in central nervous system neurons. G1 was expressed at an early embryonic stage, whereas G2 transcription was significantly induced at 10.5 days postcoitus (embryonic day 10.5, i.e. 2 days before axon outgrowth). Both G1 and G2 were strongly expressed in cortical and hippocampal neurons, cerebellar granule cells, and Purkinje cells. Immunohistochemistry with isoform-specific antibodies revealed that G2 was localized in cell bodies, dendrites, and axons. In addition, electron microscopy and subcellular fractionation indicated that G2 was localized in synaptic vesicles, whereas G1 was not detectable. G1 and G2 exhibit 62% identity, and both isoforms were immunoprecipitated with the c and A subunits of V-ATPase. G2 could complement the yeast deletion mutant Deltavma10, which lacks the G subunit. The V-ATPases containing the G1 and G2 isoforms, respectively, showed similar K(m)((ATP)) values and maximal velocity. These results indicate that G1 and G2 are bona fide subunits of V-ATPases and that the enzyme with the G2 isoform is involved in synaptic vesicle acidification. PMID- 12133828 TI - Human VPAC1 receptor selectivity filter. Identification of a critical domain for restricting secretin binding. AB - The human VPAC1 receptor for vasoactive intestinal peptide (VIP) and pituitary adenylate cyclase activating peptide (PACAP) belongs to the class II family of G protein coupled receptors with seven transmembrane segments. It recognizes several VIP-related peptides and displays a very low affinity for secretin despite >70% homology between VIP and secretin. Conversely, the human secretin receptor has high affinity for secretin but low affinity for VIP. We took advantage of this reversed selectivity to identify a domain of the VPAC1 receptor responsible for selectivity toward secretin by constructing human VPAC1-secretin receptor chimeras. A first set of chimeras consisted of exchanging the entire N terminal ectodomain or large parts of this domain. They were constructed by overlap PCR, transfected in COS-7 cells, and their ligand selectivity, expressed as the ratio of EC(50) for secretin/EC(50) for VIP (referred to as S/V), in stimulating cAMP production was measured. Two very informative chimeras respectively referred to as S144V and S123V were obtained by replacing the entire ectodomain or only the first 123 amino acids of the VPAC1 receptor by the corresponding sequences of the secretin receptor. Whereas S144V no longer discriminated between VIP and secretin (S/V = 1.2), S123V discriminated between the two peptides (S/V = 300) in the same manner as the wild-type VPAC1 receptor. The motif responsible for discrimination was determined by introducing small blocks or individual amino acids of secretin receptor in the 123-144 sequence of the S123V chimera. The data obtained from 14 new chimeras sustained that two nonadjacent pairs of amino acids, Gln(135) Thr(136) and Gly(140) Ser(141) in the C-terminal end of the N-terminal VPAC1 receptor ectodomain constitute a selective filter that strongly restricts access of secretin to the VPAC1 receptor. PMID- 12133827 TI - Enhanced expression of the human vacuolar H+-ATPase c subunit gene (ATP6L) in response to anticancer agents. AB - We have isolated two overlapping genomic clones that contain the 5'-terminal portion of the human vacuolar H(+)-ATPase c subunit (ATP6L) gene. The sequence preceding the transcription initiation site, which is GC-rich, contains four GC boxes and one Oct1-binding site, but there is no TATA box or CCAAT box. In vivo footprint analysis in human cancer cells shows that two GC boxes and the Oct1 binding site are occupied by Sp1 and Oct1, respectively. We show here that treatment with anticancer agents enhances ATP6L expression. Although cisplatin did not induce ATP6L promoter activity, it altered ATP6L mRNA stability. On the other hand, the DNA topoisomerase II inhibitor, TAS-103, strongly induced promoter activity, and this effect was completely eradicated when a mutation was introduced into the Oct1-binding site. Treatment with TAS-103 increased the levels of both Sp1/Sp3 and Oct1 in nuclear extracts. Cooperative binding of Sp1 and Oct1 to the promoter is required for promoter activation by TAS-103. Incubation of a labeled oligonucleotide probe encompassing the -73/-68 GC box and -64/-57 Oct1-binding site with a nuclear extract from drug-treated KB cells yielded higher levels of the specific DNA-protein complex than an extract of untreated cells. Thus, the two transcription factors, Sp1 and Oct1 interact, in an adaptive response to DNA damage, by up-regulating expression of the vacuolar H(+)-ATPase genes. Furthermore, combination of the vacuolar H(+)-ATPase (V ATPase) inhibitor, bafilomycin A1, with TAS-103 enhanced apoptosis of KB cells with an associated increase in caspase-3 activity. Our data suggest that the induction of V-ATPase expression is an anti-apoptotic defense, and V-ATPase inhibitors in combination with low-dose anticancer agents may provide a new therapeutic approach. PMID- 12133829 TI - Nerve growth factor signals through TrkA, phosphatidylinositol 3-kinase, and Rac1 to inactivate RhoA during the initiation of neuronal differentiation of PC12 cells. AB - In PC12 rat pheochromocytoma cells, nerve growth factor (NGF)-induced neuronal differentiation is blocked by constitutively active dominant mutants of RhoA but augmented by negative ones, suggesting a not yet elucidated inhibitory signaling link between NGF receptors and RhoA. Here we show that NGF treatment rapidly translocates RhoA from the plasma membrane to the cytosol and simultaneously decreases RhoA affinity to its target Rho-associated kinase (ROK), a key mediator of neurite outgrowth. This effect was transient, because after 2 days of NGF treatment, RhoA relocated from the cytosol to the plasma membrane, and its GTP loading returned to a level found in undifferentiated cells. Inhibition of RhoA is mediated by activation of the TrkA receptor, because NGF failed to induce RhoA translocation and inhibition of ROK binding in nnr5 cells that lack TrkA, whereas the inhibition was reconstituted in receptor add-back B5 cells. In MM17-26 cells, which due to expression of dominant negative Ras do not differentiate, NGF stimulated transient RhoA inhibition was unaffected. The inhibitory pathway from TrkA to RhoA involves phosphatidylinositol-3-kinase (PI3K), because the inhibitors LY294002 or wortmannin prevented NGF-induced RhoA translocation and increased RhoA association with ROK. Furthermore, inhibition of PI3K significantly reduced NGF- mediated Rac1 activation, whereas dominant negative Rac1 abolished the inhibitory signaling to RhoA. Taken together, these data indicate that NGF-mediated activation of TrkA receptor stimulates PI3K, which in turn increases Rac1 activity to induce transient RhoA inactivation during the initial phase of neurite outgrowth. PMID- 12133830 TI - A novel ankyrin repeat-containing gene (Kank) located at 9p24 is a growth suppressor of renal cell carcinoma. AB - By a combination of genome subtraction and comprehensive analysis of loss of heterozygosity based on mapping hemizygous deletions for a potential tumor related locus, a minimum overlapping region of deletions at 9p24 the size of 165 kb was identified and found to harbor a new potential tumor suppressor gene for renal cell carcinoma, the Kank gene. Kank (for kidney ankyrin repeat-containing protein) contains four ankyrin repeats at its C terminus. Expression of the gene was suppressed in 6 of 8 or 6 of 10 cancer tissues examined by reverse transcription-PCR or Western blotting, respectively, and in several kidney tumor cell lines due to methylation at CpG sites in the gene. Epigenetic methylation or imprinting seemed to be the first hit, which was followed by a second hit of deletion, resulting in loss of function in many of these deletion cases. Expression of this gene in expression-negative HEK293 cells induced growth retardation at G(0)/G(1) as well as morphological changes. PMID- 12133831 TI - Identification of a novel Na+/myo-inositol cotransporter. AB - rkST1, an orphan cDNA of the SLC5 family (43% identical in sequence to the sodium myo-inositol cotransporter SMIT), was expressed in Xenopus laevis oocytes that were subsequently voltage-clamped and exposed to likely substrates. Whereas superfusion with glucose and other sugars produced a small inward current, the largest current was observed with myo-inositol. The expressed protein, which we have named SMIT2, cotransports myo-inositol with a K(m) of 120 microm and displays a current-voltage relationship similar to that seen with SMIT (now called SMIT1). The transport is Na(+)-dependent, with a K(m) of 13 mm. SMIT2 exhibits phlorizin-inhibitable presteady-state currents and substrate-independent "Na(+) leak" currents similar to those of related cotransporters. The steady state cotransport current is also phlorizin-inhibitable with a K(i) of 76 microm. SMIT2 exhibits stereospecific cotransport of both d-glucose and d-xylose but does not transport fucose. In addition, SMIT2 (but not SMIT1) transports d-chiro inositol. Based on previous publications, the tissue distribution of SMIT2 is different from that of SMIT1, and the existence of this second cotransporter may explain much of the heterogeneity that has been reported for inositol transport. PMID- 12133832 TI - Functional analysis of hypoxia-inducible factor-1 alpha-mediated transactivation. Identification of amino acid residues critical for transcriptional activation and/or interaction with CREB-binding protein. AB - The hypoxia-inducible factor-1 alpha (HIF-1 alpha) is a key regulator of adaptive responses to hypoxia. HIF-1 alpha has two independent transactivation domains (TADs). Whereas the N-terminal TAD (N-TAD) also constitutes a degradation box, the C-terminal TAD (C-TAD) functions in a strictly hypoxia-inducible fashion. Oxygen-dependent hydroxylation of an asparagine residue has recently been reported to regulate C-TAD function by disrupting the interaction with the CH1 domain of the p300/CBP coactivator at normoxia. Here we have performed alanine scanning mutagenesis of a predicted alpha-helix within the C-TAD of mouse HIF-1 alpha to identify residues important for transactivation and interaction of the C TAD with transcriptional coactivators. We observed that several hydrophobic residues, Ile(802), Leu(808), Leu(814), Leu(815), and Leu(818), were critical for transactivation and binding to the CH1 domain of CBP in hypoxic cells. Moreover, E812A/E813A and D819A mutations impaired hypoxia-dependent transactivation without disrupting binding to CH1. In the context of full-length HIF-1 alpha, mutation of the leucine residues conferred conformational changes to the protein and significantly reduced the transactivation function as well as functional interaction with the transcriptional coactivators CBP and SRC-1. These mutations also affected intranuclear redistribution of HIF-1 alpha in the presence of CBP, indicating that the integrity of the C-TAD is critical for intracellular localization of mouse HIF-1 alpha. PMID- 12133833 TI - Association of the adaptor TANK with the I kappa B kinase (IKK) regulator NEMO connects IKK complexes with IKK epsilon and TBK1 kinases. AB - Canonical activation of NF-kappa B is mediated via phosphorylation of the inhibitory I kappa B proteins by the I kappa B kinase complex (IKK). IKK is composed of a heterodimer of the catalytic IKK alpha and IKK beta subunits and a presumed regulatory protein termed NEMO (NF-kappa B essential modulator) or IKK gamma. NEMO/IKK gamma is indispensable for activation of the IKKs in response to many signals, but its mechanism of action remains unclear. Here we identify TANK (TRAF family member-associated NF-kappa B activator) as a NEMO/IKK gamma interacting protein via yeast two-hybrid analyses. This interaction is confirmed in mammalian cells, and the domains required are mapped. TANK was previously shown to assist NF-kappa B activation in a complex with TANK-binding kinase 1 (TBK1) or IKK epsilon, two kinases distantly related to IKK alpha/beta, but the underlying mechanisms remained unknown. Here we show that TBK1 and IKK epsilon synergize with TANK to promote interaction with the IKKs. The TANK binding domain within NEMO/IKK gamma is required for proper functioning of this IKK subunit. These results indicate that TANK can synergize with IKK epsilon or TBK1 to link them to IKK complexes, where the two kinases may modulate aspects of NF-kappa B activation. PMID- 12133834 TI - NMR-based binding screen and structural analysis of the complex formed between alpha-cobratoxin and an 18-mer cognate peptide derived from the alpha 1 subunit of the nicotinic acetylcholine receptor from Torpedo californica. AB - The alpha18-mer peptide, spanning residues 181-198 of the Torpedo nicotinic acetylcholine receptor alpha1 subunit, contains key binding determinants for agonists and competitive antagonists. To investigate whether the alpha18-mer can bind other alpha-neurotoxins besides alpha-bungarotoxin, we designed a two dimensional (1)H-(15)N heteronuclear single quantum correlation experiment to screen four related neurotoxins for their binding ability to the peptide. Of the four toxins tested (erabutoxin a, erabutoxin b, LSIII, and alpha-cobratoxin), only alpha-cobratoxin binds the alpha18-mer to form a 1:1 complex. The NMR solution structure of the alpha-cobratoxin.alpha18-mer complex was determined with a backbone root mean square deviation of 1.46 A. In the structure, alpha cobratoxin contacts the alpha18-mer at the tips of loop I and II and through C terminal cationic residues. The contact zone derived from the intermolecular nuclear Overhauser effects is in agreement with recent biochemical data. Furthermore, the structural models support the involvement of cation-pi interactions in stabilizing the complex. In addition, the binding screen results suggest that C-terminal cationic residues of alpha-bungarotoxin and alpha cobratoxin contribute significantly to binding of the alpha18-mer. Finally, we present a structural model for nicotinic acetylcholine receptor-alpha-cobratoxin interaction by superimposing the alpha-cobratoxin.alpha18-mer complex onto the crystal structure of the acetylcholine-binding protein (Protein Data Bank code ). PMID- 12133835 TI - A novel membrane protein, Ros3p, is required for phospholipid translocation across the plasma membrane in Saccharomyces cerevisiae. AB - Ro09-0198 (Ro) is a tetracyclic peptide antibiotic that binds specifically to phosphatidylethanolamine (PE) and causes cytolysis. To investigate the molecular basis of transbilayer movement of PE in biological membranes, we have isolated a series of budding yeast mutants that are hypersensitive to the Ro peptide. One of the most sensitive mutants, designated ros3 (Ro-sensitive 3), showed no significant change in the cellular phospholipid composition or in the sensitivity to amphotericin B, a sterol-binding polyene macrolide antibiotic. These results suggest that the mutation of ros3 affects the PE organization on the plasma membrane, rather than PE synthesis or overall organization of the membrane structures. By functional complementation screening, we identified the gene ROS3 affected in the mutant, and we showed that the hypersensitive phenotype was caused by the defective expression of the ROS3 gene product, Ros3p, an evolutionarily conserved protein with two putative transmembrane domains. Disruption of the ROS3 gene resulted in a marked decrease in the internalization of fluorescence-labeled analogs of PE and phosphatidylcholine, whereas the uptake of fluorescence-labeled phosphatidylserine and endocytic markers was not affected. Neither expression levels nor activities of ATP-binding cassette transporters of the ros3Delta cells differed from those of wild type cells, suggesting that Ros3p is not related to the multidrug resistance activities. Immunochemical analyses of the structure and subcellular localization showed that Ros3p was a glycosylated membrane protein localized in both the plasma membrane and the endoplasmic reticulum, and that a part of Ros3p was associated with the detergent-insoluble glycolipid-enriched complexes. These results indicate that Ros3p is a membrane glycoprotein that plays an important role in the phospholipid translocation across the plasma membrane. PMID- 12133836 TI - Induction of bacterial lipoprotein tolerance is associated with suppression of toll-like receptor 2 expression. AB - Tolerance to bacterial cell wall components including lipopolysaccharide (LPS) may represent an essential regulatory mechanism during bacterial infection. Two members of the Toll-like receptor (TLR) family, TLR2 and TLR4, recognize the specific pattern of bacterial cell wall components. TLR4 has been found to be responsible for LPS tolerance. However, the role of TLR2 in bacterial lipoprotein (BLP) tolerance and LPS tolerance is unclear. Pretreatment of human THP-1 monocytic cells with a synthetic bacterial lipopeptide induced tolerance to a second BLP challenge with diminished tumor necrosis factor-alpha and interleukin 6 production, termed BLP tolerance. Furthermore, BLP-tolerized THP-1 cells no longer responded to LPS stimulation, indicating a cross-tolerance to LPS. Induction of BLP tolerance was CD14-independent, as THP-1 cells that lack membrane-bound CD14 developed tolerance both in serum-free conditions and in the presence of a specific CD14 blocking monoclonal antibody (MEM-18). Pre-exposure of THP-1 cells to BLP suppressed mitogen-activated protein kinase phosphorylation and nuclear factor-kappaB activation in response to subsequent BLP and LPS stimulation, which is comparable with that found in LPS-tolerized cells, indicating that BLP tolerance and LPS tolerance may share similar intracellular pathways. However, BLP strongly enhanced TLR2 expression in non-tolerized THP-1 cells, whereas LPS stimulation had no effect. Furthermore, a specific TLR2 blocking monoclonal antibody (2392) attenuated BLP-induced, but not LPS-induced, tumor necrosis factor-alpha and interleukin-6 production, indicating BLP rather than LPS as a ligand for TLR2 engagement and activation. More importantly, pretreatment of THP-1 cells with BLP strongly inhibited TLR2 activation in response to subsequent BLP stimulation. In contrast, LPS tolerance did not prevent BLP-induced TLR2 overexpression. These results demonstrate that BLP tolerance develops through down-regulation of TLR2 expression. PMID- 12133837 TI - Formation of highly reactive A-ring and J-ring isoprostane-like compounds (A4/J4 neuroprostanes) in vivo from docosahexaenoic acid. AB - Free radical-initiated oxidant injury and lipid peroxidation have been implicated in a number of neural disorders. Docosahexaenoic acid is the most abundant unsaturated fatty acid in the central nervous system. We have shown previously that this 22-carbon fatty acid can yield, upon oxidation, isoprostane-like compounds termed neuroprostanes, with E/D-type prostane rings (E(4)/D(4) neuroprostanes). Eicosanoids with E/D-type prostane rings are unstable and dehydrate to cyclopentenone-containing compounds possessing A-type and J-type prostane rings, respectively. We thus explored whether cyclopentenone neuroprostanes (A(4)/J(4)-neuroprostanes) are formed from the dehydration of E(4)/D(4)-neuroprostanes. Indeed, oxidation of docosahexaenoic acid in vitro increased levels of putative A(4)/J(4)-neuroprostanes 64-fold from 88 +/- 43 to 5463 +/- 2579 ng/mg docosahexaenoic acid. Chemical approaches and liquid chromatography/electrospray ionization tandem mass spectrometry definitively identified them as A(4)/J(4)-neuroprostanes. We subsequently showed these compounds are formed in significant amounts from a biological source, rat brain synaptosomes. A(4)/J(4)-neuroprostanes increased 13-fold, from a basal level of 89 +/- 72 ng/mg protein to 1187 +/- 217 ng/mg (n = 4), upon oxidation. We also detected these compounds in very large amounts in fresh brain tissue from rats at levels of 97 +/- 25 ng/g brain tissue (n = 3) and from humans at levels of 98 +/- 26 ng/g brain tissue (n = 5), quantities that are nearly an order of magnitude higher than other classes of neuroprostanes. Because of the fact that A(4)/J(4) neuroprostanes contain highly reactive cyclopentenone ring structures, it would be predicted that they readily undergo Michael addition with glutathione and adduct covalently to proteins. Indeed, incubation of A(4)/J(4)-neuroprostanes in vitro with excess glutathione resulted in the formation of large amounts of adducts. Thus, these studies have identified novel, highly reactive A/J-ring isoprostane-like compounds that are derived from docosahexaenoic acid in vivo. PMID- 12133838 TI - Cannabinoids protect astrocytes from ceramide-induced apoptosis through the phosphatidylinositol 3-kinase/protein kinase B pathway. AB - Cannabinoids, the active components of marijuana and their endogenous counterparts, exert many of their actions on the central nervous system by binding to the CB(1) cannabinoid receptor. Different studies have shown that cannabinoids can protect neural cells from different insults. However, those studies have been performed in neurons, whereas no attention has been focused on glial cells. Here we used the pro-apoptotic lipid ceramide to induce apoptosis in astrocytes, and we studied the protective effect exerted by cannabinoids. Results show the following: (i) cannabinoids rescue primary astrocytes from C(2)-ceramide induced apoptosis in a dose- and time-dependent manner; (ii) triggering of this anti-apoptotic signal depends on the phosphatidylinositol 3-kinase/protein kinase B pathway; (iii) ERK and its downstream target p90 ribosomal S6 kinase might be also involved in the protective effect of cannabinoids; and (iv) cannabinoids protect astrocytes from the cytotoxic effects of focal C(2)-ceramide administration in vivo. In summary, results show that cannabinoids protect astrocytes from ceramide-induced apoptosis via stimulation of the phosphatidylinositol 3-kinase/protein kinase B pathway. These findings constitute the first evidence for an "astroprotective" role of cannabinoids. PMID- 12133839 TI - Crystallographic analysis of the MoFe protein of nitrogenase from a nifV mutant of Klebsiella pneumoniae identifies citrate as a ligand to the molybdenum of iron molybdenum cofactor (FeMoco). AB - The x-ray crystal structure of NifV(-) Klebsiella pneumoniae nitrogenase MoFe protein (NifV(-) Kp1) has been determined and refined to a resolution of 1.9 A. This is the first structure for a nitrogenase MoFe protein with an altered cofactor. Moreover, it is the first direct evidence that the organic acid citrate is not just present, but replaces homocitrate as a ligand to the molybdenum atom of the iron molybdenum cofactor (FeMoco). Subsequent refinement of the structure revealed that the citrate was present at reduced occupancy. PMID- 12133840 TI - Using hetero-11-mers composed of wild type and mutant subunits to study tryptophan binding to TRAP and its role in activating RNA binding. AB - Expression of genes involved in tryptophan metabolism in Bacillus subtilis is regulated by the TRAP protein in response to changes in l-tryptophan levels. TRAP binding to several RNA targets that contain between 9 and 11 (G/U)AG repeats regulates transcription and/or translation of these genes. TRAP consists of 11 identical subunits and is activated to bind RNA by binding up to 11 molecules of tryptophan. To investigate the mechanism by which tryptophan binding activates TRAP, we generated hetero-11-mers containing different proportions of subunits from wild type (WT) TRAP that bind tryptophan and from a mutant TRAP (Thr(25) to Ala) defective in tryptophan binding. Studies of these hetero-11-mers show that tryptophan-binding sites created from active subunits bind tryptophan with similar affinity to those in WT homo-11-mers, whereas sites containing the T25A substitution do not bind tryptophan. Hetero-11-mers with very few (one or two) bound tryptophans show only 10-fold lower affinity than WT TRAP for an RNA with 11 GAG repeats, whereas TRAP with no bound tryptophan shows no detectable binding to this RNA. We also demonstrate that tryptophan binding induces a conformational change in TRAP in the vicinity of the RNA-binding site, suggesting a possible mechanism for activation of RNA binding. PMID- 12133841 TI - Structure of the BgK-Kv1.1 complex based on distance restraints identified by double mutant cycles. Molecular basis for convergent evolution of Kv1 channel blockers. AB - A structural model of BgK, a sea anemone toxin, complexed with the S5-S6 region of Kv1.1, a voltage-gated potassium channel, was determined by flexible docking under distance restraints identified by a double mutant cycles approach. This structure provides the molecular basis for identifying the major determinants of the BgK-Kv1.1 channel interactions involving the BgK dyad residues Lys(25) and Tyr(26). These interactions are (i) electrostatic interactions between the extremity of Lys(25) side chain and carbonyl oxygen atoms of residues from the channel selectivity filter that may be strengthened by solvent exclusion provided by (ii) hydrophobic interactions involving BgK residues Tyr(26) and Phe(6) and Kv1.1 residue Tyr(379) whose side chain protrudes in the channel vestibule. In other Kv1 channel-BgK complexes, these interactions are likely to be conserved, implicating both conserved and variable residues from the channels. The data suggest that the conservation in sea anemone and scorpion potassium channel blockers of a functional dyad composed of a lysine, and a hydrophobic residue reflects their use of convergent binding solutions based on a crucial interplay between these important conserved interactions. PMID- 12133842 TI - Analysis of double knockout mice lacking aquaporin-1 and urea transporter UT-B. Evidence for UT-B-facilitated water transport in erythrocytes. AB - We reported increased water permeability and a low urea reflection coefficient in Xenopus oocytes expressing urea transporter UT-B (former name UT3), suggesting that water and urea share a common aqueous pathway (Yang, B., and Verkman, A. S. (1998) J. Biol. Chem. 273, 9369-9372). Although increased water permeability was confirmed in the Xenopus oocyte expression system, it has been argued (Sidoux Walter, F., Lucien, N., Olives, B., Gobin, R., Rousselet, G., Kamsteeg, E. J., Ripoche, P., Deen, P. M., Cartron, J. P., and Bailly, P. (1999) J. Biol. Chem. 274, 30228-30235) that UT-B does not transport water when expressed at normal levels in mammalian cells such as erythrocytes. To quantify UT-B-mediated water transport, we generated double knockout mice lacking UT-B and the major erythrocyte water channel, aquaporin-1 (AQP1). The mice had reduced survival, retarded growth, and defective urinary concentrating ability. However, erythrocyte size and morphology were not affected. Stopped-flow light scattering measurements indicated erythrocyte osmotic water permeabilities (in cm/s x 0.01, 10 degrees C): 2.1 +/- 0.2 (wild-type mice), 2.1 +/- 0.05 (UT-B null), 0.19 +/- 0.02 (AQP1 null), and 0.045 +/- 0.009 (AQP1/UT-B null). The low water permeability found in AQP1/UT-B null erythrocytes was also seen after HgCl(2) treatment of UT-B null erythrocytes or phloretin treatment of AQP1 null erythrocytes. The apparent activation energy for UT-B-mediated water transport was low, <2 kcal/mol. Estimating 14,000 UT-B molecules per mouse erythrocyte, the UT-B-dependent P(f) of 0.15 x 10(-4) cm/s indicated a substantial single channel water permeability of UT-B of 7.5 x 10(-14) cm(3)/s, similar to that of AQP1. These results provide direct functional evidence for UT-B-facilitated water transport in erythrocytes and suggest that urea traverses an aqueous pore in the UT-B protein. PMID- 12133844 TI - Invited editorial on "the alcohol breath test". PMID- 12133843 TI - Characterization of mammalian eIF2A and identification of the yeast homolog. AB - To begin the physical characterization of eukaryotic initiation factor (eIF) 2A, a translation initiation factor that binds Met-tRNA(i), tryptic peptides from rabbit reticulocyte eIF2A were analyzed to obtain amino acid sequence information. Sequences for 8 peptides were matched to three different expressed sequence tag clones. The sequence predicted for eIF2A is 585 amino acids. Matching of the cDNA sequence to the human genome revealed that the eIF2A mRNA is made up of 15 or 16 exons, and the gene is contained on chromosome 3. A homolog in Saccharomyces cerevisiae was identified, YGR054W, which is a non-essential gene. Hemagglutinin-tagged yeast eIF2A localizes on both 40 S and 80 S ribosomes. A knockout of both eIF2A and eIF5B yielded a "synthetically sick" yeast strain with a severe slow growth phenotype. The phenotype of this double mutant and the biochemical localization suggest that eIF2A participates in translation initiation. eIF2A does not appear to participate in re-initiation as the DeltaeIF2A strain shows the same level of GCN4 induction with amino acid starvation as seen in wild type yeast. The lack of any apparent phenotype in the DeltaeIF2A strain suggests that eIF2A functions in a minor pathway, perhaps internal initiation or in the translation of a small number of specific mRNAs. PMID- 12133845 TI - Functional characteristics of dystrophic skeletal muscle: insights from animal models. AB - Muscular dystrophies are a clinically and genetically heterogeneous group of disorders that show myofiber degeneration and regeneration. Identification of animal models of muscular dystrophy has been instrumental in research on the pathogenesis, pathophysiology, and treatment of these disorders. We review our understanding of the functional status of dystrophic skeletal muscle from selected animal models with a focus on 1) the mdx mouse model of Duchenne muscular dystrophy, 2) the Bio 14.6 delta-sarcoglycan-deficient hamster model of limb-girdle muscular dystrophy, and 3) transgenic null mutant murine lines of sarcoglycan (alpha, beta, delta, and gamma) deficiencies. Although biochemical data from these models suggest that the dystrophin-sarcoglycan-dystroglycan laminin network is critical for structural integrity of the myofiber plasma membrane, emerging studies of muscle physiology suggest a more complex picture, with specific functional deficits varying considerably from muscle to muscle and model to model. It is likely that changes in muscle structure and function, downstream of the specific, primary biochemical deficiency, may alter muscle contractile properties. PMID- 12133846 TI - Evaluation of inotropic changes in ventricular function by NOGA mapping: comparison with echocardiography. AB - Assessment of left ventricular (LV) function in the catheterization laboratory is important to optimize treatment decisions and guide catheter-based local therapies. NOGA electromechanical mapping was developed to assess LV contraction during catheterization; however, quantitative analysis of its "local shortening" (LS) algorithm and direct comparison with conventional methods are lacking. We evaluated the accuracy of NOGA-based regional and global function by examining its ability to detect pharmacologically induced changes in contractility compared with echocardiography. Ten anesthetized pigs were paced to ensure a constant heart rate throughout the experiment. Electromechanical maps of the LV and short axis echocardiograms were obtained 1) at baseline, 2) during intravenous dobutamine, and 3) after intravenous propranolol. NOGA LS and ejection fraction (EF) consistently increased under dobutamine and decreased after propranolol. NOGA LS and NOGA and echocardiography circumferential shortening correlated highly with one another (r > 0.80), as did NOGA EF with echocardiography EF (r = 0.92), although absolute values differed somewhat. Thus NOGA-based global and regional function correlates closely with echocardiography and is sensitive to changes in contractility, but, at the upper end of the scale, LV function is underestimated. PMID- 12133847 TI - Anatomical arrangement of hypercapnia-activated cells in the superficial ventral medulla of rats. AB - The anatomical structure of central respiratory chemoreceptors in the superficial ventral medulla of rats was studied by using hypercapnia-induced c-fos labeling to identify cells directly stimulated by extracellular pH or PCO(2). The distribution of c-fos-positive cells was found to be predominantly perivascular to surface vessels. In the superficial ventral medullary midline, parapyramidal, and ventrolateral regions where c-fos-positive cells were concentrated, we found a common, characteristic, anatomical architecture. The medullary surface showed an indentation covered by a surface vessel, and the marginal glial layer was thickened. We classified c-fos-positive cells into two types. One (type I cell) was small, was located inside the marginal glial layer and close to the medullary surface, and surrounded fine vessels. The other (type II cell) was large and located dorsal to the marginal glial layer. c-fos Expression under synaptic blockade suggested that type I cells are intrinsically chemosensitive. The chemosensitivity of surface cells (possible type I cells) surrounding vessels was confirmed electrophysiologically in slice preparations. We suggest that this characteristic anatomical structure may be the central chemoreceptor. PMID- 12133848 TI - Activation of the midbrain periaqueductal gray induces airway smooth muscle relaxation. AB - In this study, we examined effects of chemical stimulation of the ventrolateral region of the midbrain periaqueductal gray (vl PAG) on airway smooth muscle tone. We observed that in anesthetized, paralyzed, and artificially ventilated ferrets, vl PAG stimulation elicited airway smooth muscle relaxation. To clarify the mechanisms underlying this observation, we examined the GABA-GABA(A) receptor signaling pathway by 1) examining the expression of GABA(A) receptors on airway related vagal preganglionic neurons (AVPNs) located in the rostral nucleus ambiguus region (rNA), by use of receptor immunochemistry and confocal microscopy; 2) measuring GABA release within the rNA by using microdialysis; and 3) performing physiological experiments to determine the effects of selective blockade of GABA(A) receptors expressed by AVPNs in the rNA region on vl PAG induced airway relaxation, thereby defining the role of the GABA(A) receptor subtype in this process. We observed that AVPNs located in the rNA region do express the GABA(A) receptor beta-subtype. In addition, we demonstrated that activation of vl PAG induced GABA release within the rNA region, and this release was associated with airway smooth muscle relaxation. Blockade of the GABA(A) receptor subtype expressed by AVPNs in the rNA by bicuculline diminished the inhibitory effects of vl PAG stimulation on airway smooth muscle tone. These data indicate, for the first time, that activation of vl PAG dilates the airways by a release of GABA and activation of GABA(A) receptors expressed by AVPNs. PMID- 12133849 TI - Endogenous nitric oxide modulates responses of tissue and airway resistance to vagal stimulation in piglets. AB - The role of endogenous nitric oxide (NO) in modulating the excitatory response of distal airways to vagal stimulation is unknown. In decerebrate, ventilated, open chest piglets aged 3-10 days, lung resistance (RL) was partitioned into tissue resistance (Rti) and airway resistance (Raw) by using alveolar capsules. Changes in RL, Rti, and Raw were evaluated during vagal stimulation at increasing frequency before and after NO synthase blockade with N(omega)-nitro-L-arginine methyl ester (L-NAME). Vagal stimulation increased RL by elevating both Rti and Raw. NO synthase blockade significantly increased baseline Rti, but not Raw, and significantly augmented the effects of vagal stimulation on both Rti and Raw. Vagal stimulation also resulted in a significant increase in cGMP levels in lung tissue before, but not after, L-NAME infusion. In seven additional piglets after RL was elevated by histamine infusion in the presence of cholinergic blockade with atropine, vagal stimulation failed to elicit any change in RL, Rti, or Raw. Therefore, endogenous NO not only plays a role in modulating baseline Rti, but it opposes the excitatory cholinergic effects on both the tissue and airway components of RL. We speculate that activation of the NO/cGMP pathway during cholinergic stimulation plays an important role in modulating peripheral as well as central contractile elements in the developing lung. PMID- 12133850 TI - Voluntary muscle activation varies with age and muscle group. AB - The consistency and the number of attempts required to achieve maximal voluntary muscle activation have not been documented and compared between young and old adults. Furthermore, few studies have contrasted activation between functional pairs of muscle groups, and no study has tested upper limb muscles. The purpose of this study was to measure and compare voluntary muscle activation of the elbow flexors and extensors in young and old men over two separate test sessions. With the method of twitch interpolation to measure activation, six young (24 +/- 1 yr) and six old (83 +/- 4 yr) men performed five maximal voluntary contractions (MVC) during each session for each muscle group. Elbow flexion and extension MVC was less (43 and 47%, respectively) in the old men, yet the best maximal voluntary muscle activation was similar between age groups. However, when all 10 attempts at MVC were compared, the mean activation scores were slightly less (approximately 5%) in the elbow extensors but were approximately 11% less (P < 0.001) in the elbow flexors of old men, compared with young men. During the second session, there was a significant improvement of 13% (P < 0.005) in mean elbow flexor activation in the old men. There were no session differences for either muscle group for the young men. The results indicate that, for aged men, elbow flexor maximal activation is achieved less frequently compared with elbow extensors, and thus mean activation for elbow flexors is less than for elbow extensors. However, if sufficient attempts are provided, the best effort for the old men is not different from that of the young men for either muscle group. PMID- 12133851 TI - Effect of exercise on mRNA expression of select adrenal medullary catecholamine biosynthetic enzymes. AB - The effect of submaximal endurance training (SET) on sympathoadrenal activity is not clear. We tested the hypothesis that SET (90 min/day, 5 days/wk, for 12 wk) elevates mRNA expression of catecholamine (CA) biosynthetic enzymes, tyrosine hydroxylase (TH) and dopamine-beta-hydroxylase (DbetaH) in the adrenal medullae of adult, female Sprague-Dawley rats. SET increased TH protein level by 35%, TH activity by 62%, TH mRNA expression by 40%, and DbetaH mRNA expression by 67%. In addition, we examined the effect of SET on Fos-related antigens (FRAs), FRA-2 immunoreactivity, and activator protein (AP)-1 binding activity. SET increased AP 1 binding activity by 78%; however, it did not affect late FRAs and FRA-2 immunoreactivity. Because the regulation of neuropeptide Y (NPY) often parallels that of CAs, we also examined the effect of SET on NPY mRNA expression. Indeed, SET elevated NPY mRNA expression as well. We conclude that 1) SET elicits a pretranslational stimulatory effect on adrenomedullary CA biosynthetic enzymes, 2) another immediate early mRNA product, rather than FRA-2, may contribute to the increase in AP-1 binding activity in response to SET, and 3) SET increases NPY mRNA expression. PMID- 12133852 TI - Factors in fatigue during intermittent electrical stimulation of human skeletal muscle. AB - During an electrically elicited isometric contraction, the metabolic cost of attaining is greater than of maintaining force. Thus fatigue produced during such stimulation may not simply be a function of the force-time integral (FTI), as previously suggested. The goal of the present study was to evaluate fatigue produced in human medial gastrocnemius by intermittent, isometric electrical stimulation with trains of different frequencies (20, 40, or 80 Hz) and durations (300, 600, or 1,200 ms) that produced different peak forces and FTIs. Each subject (n = 10) participated in a total of six sessions. During each session, subjects received a pre- and postfatigue testing protocol and a different, 150 train fatiguing protocol. Each fatiguing protocol used only a single frequency and duration. The fatigue produced by the different protocols was correlated to the initial peak force of the fatiguing protocols (r2= 0.74-0.85) but not to the initial or total FTI. All of the protocols tested produced a proportionately greater impairment of force in response to low- vs. high-frequency stimulation (i.e., low-frequency fatigue). There was no effect of protocol on low-frequency fatigue, suggesting that all the protocols produced comparable levels of impairment in excitation-contraction coupling. These results suggest that, for brief stimulated contractions, peak force is a better predictor of fatigue than FTI, possibly because of the different metabolic demands of attaining and maintaining force. PMID- 12133853 TI - Estradiol increases salt intake in female normotensive and hypertensive rats. AB - The objective of this study was to examine whether or not estradiol (E2) alters sodium intake in hypertensive and normotensive female rats. It was hypothesized that higher doses of E2 would increase sodium consumption and that this response would be greater in spontaneously hypertensive rats (SHR) compared with Wistar Kyoto (WKY) rats. The study involved female SHR and WKY (n = 12/group). All animals were ovariectomized. Six of twelve rats from each strain received three progressively larger doses of beta-estradiol propionate (each dose lasting 2 wk), whereas the other six rats from each strain received sham implants. Blood E2 levels were measured by radioimmunoassay after each 2-wk period, allowing a 10 day washout period before the next E2 dose. Rats had access to 0.0, 0.5, 1.0, and 1.5% NaCl solutions to drink throughout the experiment. There was a significant positive correlation between sodium intake and plasma E2 (r = 0.8, P < 0.001). Both strains avoided the 1.5% NaCl, and the increased sodium intake was achieved by an increase in consumption of the 0.5% NaCl. SHR females consumed more sodium than WKY females, which is similar to what has been observed in males of these strains. In conclusion, E2 was positively correlated with sodium intake in both strains of rat, with the hypertensive rats consuming more sodium than the normotensive rats. PMID- 12133854 TI - Heat-induced force suppression and HSP20 phosphorylation in swine carotid media. AB - In vascular smooth muscle, cyclic nucleotide-dependent phosphorylation of heat shock protein 20 (HSP20) on serine-16 (Ser16) has been suggested to cause force suppression, i.e., reduced force with only minimal myosin regulatory light chain (MRLC) dephosphorylation. We hypothesized that heat pretreatment also suppresses force by increasing HSP20 phosphorylation. After heat pretreatment of swine carotid artery at 44.5 degrees C for 4 h and reduction to 37 degrees C for 1 h, Ser16-HSP20 phosphorylation was increased and histamine-induced increases in contractile force were suppressed. Subsequent addition of nitroglycerin induced additive force suppression. Heat and nitroglycerin induced a similar relation between Ser16-HSP20 phosphorylation and force. Heat pretreatment induced a small, but significant, increase in total HSP20 immunostaining. These results demonstrate that vascular smooth muscle responds to thermal stress by increasing Ser16-HSP20 phosphorylation in addition to a possible small increase in total HSP20 concentration. The resulting heat-induced reduction in force should be considered "force suppression" because histamine-induced increases in MRLC phosphorylation were not significantly altered by heat pretreatment. These processes may bring about a resistance to contractile agonists, which could have clinical significance in conditions such as hyperthermia and/or sepsis with vasodilatory shock. PMID- 12133855 TI - Age and contraction type influence motor output variability in rapid discrete tasks. AB - The purpose of this study was to examine the ability to control knee-extension force during discrete isometric (IC), concentric (CC), and eccentric contractions (EC) in 24 young (mean age +/- SD = 25.3 +/- 2.8 yr) and 24 old (mean age +/- SD = 73.3 +/- 5.5 yr) healthy and active individuals. Subjects were to match a parabola with a time to peak force of 200 ms during IC, CC, and EC at six target levels of force [20, 35, 50, 65, 80, and 90% of the maximum voluntary contraction (MVC)]. ICs were performed at 90 degrees of knee flexion, whereas CCs and ECs ranged from 90 to 80 degrees of knee flexion (0 degrees is full extension) at a slow velocity (25 degrees /s). Results showed that subjects produced similar MVC forces for the three types of contractions. Young subjects produced greater MVC forces than old subjects, and within each age group, men produced greater force than women. The variability (standard deviation) of peak force and impulse in absolute values was greater for young compared with old subjects. When variability was normalized to the force produced [coefficient of variation (CV)], however, old subjects exhibited greater CV than young subjects for peak force and impulse. Both the standard deviation and CV of time to peak force and impulse duration were greater for the old adults. In general, ECs were more variable than ICs and CCs, and old adults exhibited greater CV compared with young adults during rapid, discrete ICs, CCs, and particularly ECs of the quadriceps. PMID- 12133856 TI - Differential metabolic fate of the carbon skeleton and amino-N of [13C]alanine and [15N]alanine ingested during prolonged exercise. AB - The decarboxylation/oxidation and the deamination of 13C- and [15N]alanine ingested (1 g/kg or 73.7 +/- 2 g) during prolonged exercise at low workload (180 min at 53 +/- 2% maximal O2 uptake) was measured in six healthy male subjects from V13CO2 at the mouth and [15N]urea excretion in urine and sweat. Over the exercise period, 50.6 +/- 3.5 g of exogenous alanine were oxidized (68.7 +/- 4.5% of the load), providing 10.0 +/- 0.6% of the energy yield vs. 4.8 +/- 0.4, 47.6 +/- 4.3, and 37.4 +/- 4.7% for endogenous proteins, glucose, and lipids, respectively. Alanine could have been oxidized after conversion into glucose in the liver and/or directly in peripheral tissues. In contrast, only 13.0 +/- 3.2 mmol of [(15)N]urea were excreted in urine and sweat (10.6 +/- 0.4 and 2.4 +/- 0.5 mmol, respectively), corresponding to the deamination of 2.3 +/- 0.3 g of exogenous alanine (3.1 +/- 0.4% of the load). These results confirm that the metabolic fate of the carbon skeleton and the amino-N moiety of exogenous alanine ingested during prolonged exercise at low workload are markedly different. The large positive nitrogen balance (8.5 +/- 0.3 g) suggests that in this situation protein synthesis could be increased when a large amount of a single amino acid is ingested. PMID- 12133857 TI - Intersubunit disulfide bridge is not required for the protective role of SP-B against lung inflammation. AB - Surfactant protein B (SP-B) is known to promote surfactant phospholipid film formation and reduce surface tension. Native SP-B is a homodimer in which subunit association is stabilized via covalent linkage through cysteine 48. We hypothesized that loss of the intersubunit bridge would alter SP-B function and lead to increased inflammation in response to challenge by hyperoxia or endotoxin. Transgenic mice in which SP-B cysteine 48 was mutated to serine were generated and crossed into the SP-B(-/-) background. Wild-type mice and transgenic mice carrying a single copy (SP-Bmon(+)) or two copies (SP-Bmon(++)) of the transgene were exposed to 95% O2 for 3 days or intratracheally injected with 10 microg of endotoxin. Interleukin-1beta, major intrinsic protein 2, and interleukin-6 in lung homogenates after 3 days of hyperoxia were significantly higher (P < 0.001) in SP-Bmon(+) mice than SP-Bmon(++) or wild-type mice. At 16 h after endotoxin injection, cytokines in lung tissues were higher in SP-Bmon(+) mice compared with wild-type mice (P < 0.05). Consistent with prolonged recovery in SP-Bmon(+) mice, the percentage of apoptotic cells in alveolar lavage was significantly lower in SP-Bmon(+) mice than in SP-Bmon(++) and wild-type mice. Overall, increased inflammation in SP-Bmon(+) mice was corrected to a large extent by increased gene dosage, indicating that formation of the intersubunit disulfide bridge is not critical for SP-B function. PMID- 12133858 TI - Indomethacin impairs LPS-induced behavioral fever in toads. AB - We tested the hypothesis that PGs mediate lipopolysaccharide (LPS)-induced behavioral fever in the toad Bufo paracnemis. Measurements of preferred body temperature (T(b)) were performed with a thermal gradient. Toads were injected intraperitoneally with the cyclooxygenase inhibitor indomethacin (5 mg/kg), which inhibits PG biosynthesis, or its vehicle (Tris) followed 30 min later by LPS (0.2 and 2 mg/kg) into the lymph sac. LPS at the dose of 0.2 mg/kg caused a significant increase in T(b) from 7 to 10 h after injection, and then T(b) returned toward baseline values. LPS at the dose of 2 mg/kg produced a different pattern of response, with a longer latency to the onset of fever (10th h) and a longer duration (until the end of the experiment at the 15th h). Tris significantly attenuated the fever induced by LPS at 0.2 mg/kg, but not at 2 mg/kg. Moreover, indomethacin completely blocked the fever evoked by LPS (2 mg/kg). These results indicate that the behavioral fever induced by LPS in toads requires the activation of the COX pathway, suggesting that the involvement of PG in fever has an ancient phylogenetic history and that endogenous PGs raise the thermoregulatory set point to produce fever, because behavioral thermoregulation seems to be related to changes in the thermoregulatory set point. PMID- 12133859 TI - Interactions of lung stretch, hyperoxia, and MIP-2 production in ventilator induced lung injury. AB - The use of positive pressure mechanical ventilation can cause ventilator-induced lung injury (VILI). We hypothesized that hyperoxia in combination with large tidal volumes (VT) would accentuate noncardiogenic edema and neutrophil infiltration in VILI and be dependent on stretch-induced macrophage inflammatory protein-2 (MIP-2) production. In rats ventilated with VT 20 ml/kg, there was pulmonary edema formation that was significantly increased by hyperoxia. Total lung neutrophil infiltration and MIP-2 in bronchoalveolar lavage (BAL) fluid were significantly elevated, in animals exposed to high VT both on room air (RA) and with hyperoxia. Hyperoxia markedly augmented the migration of neutrophils into the alveoli. Anti-MIP-2 antibody blocked migration of neutrophils into the alveoli in RA by 51% and with hyperoxia by 65%. We concluded that neutrophil migration into the alveoli was dependent on stretch-induced MIP-2 production. Hyperoxia significantly increased edema formation and neutrophil migration into the alveoli with VT 20 ml/kg, although BAL MIP-2 levels were nearly identical to VT 20 ml/kg with RA, suggesting that other mechanisms may be involved in hyperoxia-augmented neutrophil alveolar content in VILI. PMID- 12133860 TI - Effect of body temperature during exercise on skeletal muscle cytochrome c oxidase content. AB - This study determined the role of body temperature during exercise on cytochrome c oxidase (CytOx) activity, a marker of mitochondrial content, and mitochondrial heat shock protein 70 (mtHSP70), which is required for import of nuclear-coded preproteins. Male, 10-wk-old, Sprague-Dawley rats exercised identically for 9 wk in ambient temperatures of 23 degrees C (n = 10), 8 degrees C with wetted fur (n = 8), and 4 degrees C with wetted fur and fan (n = 7). These conditions maintained exercising core temperature (T(c)) at 40.4, 39.2, or 38.0 degrees C (resting temperature), respectively. During weeks 3-9, exercisers ran 5 days/wk up a 6% grade at 20 m/min for 60 min. Animals were housed at 23 degrees C. Gastrocnemius CytOx activity in T(c)=38.0 degrees C (83.5 +/- 5.5 microatoms O x min(-1) x g wet wt(-1)) was greater than all other groups (P < 0.05), exceeding sedentary (n = 7) by 73.2%. T(c) of 40.4 and 39.2 degrees C also were higher than sedentary by 22.4 and 37.4%, respectively (P < 0.05). Quantification of CytOx content verified that the increased activity was due to an increase in protein content. In extensor digitorum longus, a nonactive muscle, CytOx was not elevated in T(c) = 38.0 degrees C. mtHSP70 was significantly elevated in gastrocnemius of T(c) = 38.0 degrees C compared with sedentary (P < 0.05) but was not elevated in extensor digitorum longus (P > 0.05). The data indicate that decreasing exercise T(c) may enhance mitochondrial biogenesis and that mtHSP70 expression is not dependent on temperature. PMID- 12133861 TI - Peri-OVLT E-series prostaglandins and core temperature do not increase after intravenous IL-1beta in pregnant rats. AB - Rats have an attenuated febrile response to endogenous pyrogen near the term of pregnancy. Given the fundamental role of E-series prostaglandins (PGEs) in mediating the febrile response to blood-borne endogenous pyrogen, the present experiments were carried out to determine whether PGEs increase in the area surrounding the organum vasculosum laminae terminalis (peri-OVLT) of near-term pregnant (P) rats as in nonpregnant (NP) rats after intravenous (iv) administration of recombinant rat interleukin-1beta (rrIL-1beta). Core temperature was measured by telemetry and peri-OVLT interstitial fluid was sampled in 12 NP and 12 P chronically instrumented, Sprague-Dawley rats by microdialysis for determination of total PGEs by radioimmunoassay. Basal core temperatures were higher in NP compared with P rats (NP 37.9 degrees C +/- 0.5, P 36.9 degrees C +/- 0.4; P < 0.05), but basal peri-OVLT PGEs were similar in both groups (NP 260 +/- 153 pg/ml, P 278 +/- 177 pg/ml; P =not significant). Intravenous administration of rrIL-1beta to NP rats produced a significant increase in core temperature with a latency, magnitude, and duration of 10 min, 0.87 degrees C, and at least 170 min, respectively; peri-OVLT PGEs were increased significantly by 30 min and averaged 270% above basal levels throughout the experiment. In P rats, however, neither core temperature nor peri-OVLT PGEs increased significantly after iv administration of rrIL-1beta. Intravenous administration of vehicle did not significantly alter core temperature or peri OVLT PGEs in either group of rats. Thus peri-OVLT PGEs do not increase in P rats as they do in NP rats after iv administration of rrIL-1beta. The mechanism of this interesting component of the maternal adaptation to pregnancy, which likely plays a major role in mediating the attenuated febrile response to endogenous pyrogen near the term of pregnancy, warrants further investigation. PMID- 12133862 TI - Regenerated mdx mouse skeletal muscle shows differential mRNA expression. AB - Despite over 3,000 articles published on dystrophin in the last 15 years, the reasons underlying the progression of the human disease, differential muscle involvement, and disparate phenotypes in different species are not understood. The present experiment employed a screen of 12,488 mRNAs in 16-wk-old mouse mdx muscle at a time when the skeletal muscle is avoiding severe dystrophic pathophysiology, despite the absence of a functional dystrophin protein. A number of transcripts whose levels differed between the mdx and human Duchenne muscular dystrophy were noted. A fourfold decrease in myostatin mRNA in the mdx muscle was noted. Differential upregulation of actin-related protein 2/3 (subunit 4), beta thymosin, calponin, mast cell chymase, and guanidinoacetate methyltransferase mRNA in the more benign mdx was also observed. Transcripts for oxidative and glycolytic enzymes in mdx muscle were not downregulated. These discrepancies could provide candidates for salvage pathways that maintain skeletal muscle integrity in the absence of a functional dystrophin protein in mdx skeletal muscle. PMID- 12133863 TI - Systemic vs. local cytokine and leukocyte responses to unilateral wrist flexion exercise. AB - We hypothesized that brief exercise of a small muscle group would lead to local rather than systemic alterations in cytokines, peripheral blood mononuclear cells, and mediators of angiogenesis. Fifteen men and eight women (age range 22 36 yr old) performed 10 min of unilateral wrist flexion exercise. Blood was sampled from venous catheters in the resting and exercising arm at baseline, at the end of exercise, and at 10, 30, 60, and 120 min after exercise. Lactate was significantly elevated in the exercising arm (+276 +/- 35%; P < 0.0005) with no change in the resting arm. In contrast, increases in both arms were observed for interleukin-6 (+139 +/- 51%; P < 0.0005), growth hormone (+1,104 +/- 284%; P < 0.003), natural killer cells (+81 +/- 9%; P < 0.0005), and lymphocytes expressing CD62L, CD11a, and CD54. There were no significant differences in these increases between the resting and exercising arm. Catecholamines increased in both arms [epinephrine peak increase, +226 +/- 36% (P < 0.001); norepinephrine peak increase, +90 +/- 15% (P < 0.01)]. Fibroblast growth factor-2 initially decreased with exercise in both arms, and this was followed by a rebound increase. Vascular endothelial growth factor demonstrated a small but significant increase in both arms (+124 +/- 31%; P < 0.05). Brief, low-intensity exercise leads to a systemic rather than local response of mediators that could be involved in inflammation, repair, or angiogenic adaptation to physical activity. PMID- 12133864 TI - Evidence for sympatholysis at the onset of forearm exercise. AB - The effect of augmented sympathetic outflow on forearm vascular conductance after single handgrip contractions of graded intensity was examined to determine whether sympatholysis occurs early in exercise (n = 7). While supine, subjects performed contractions that were 1 s in duration and 15, 30, and 60% of maximal voluntary contraction (MVC) in intensity. The contractions were repeated during control and lower body negative pressure (LBNP) (-40 mmHg) sessions. Forearm blood flow (FBF; Doppler ultrasound) and mean arterial pressure were measured continuously for 30 s before and 60 s after the single contractions. Vascular conductance (VC) was calculated. Total postcontraction blood flow increased in an exercise intensity-dependent manner. Compared with control, LBNP caused a reduction in baseline and postexercise FBF (P < 0.05), VC (P < 0.01), as well as total excess flow (P < 0.01). Specifically, during LBNP, baseline FBF and VC were reduced by 29 and 34% of control, respectively (P < 0.05). After the 15% MVC contraction, peak VC during LBNP was reduced by a magnitude similar to that during baseline (i.e., ~30%), but it was only reduced by 15% during both the 30 and 60% MVC trials (P < 0.01). It was concluded that the stimuli for exercise hyperemia during moderate and heavy, but not mild, handgrip exercise intensities, diminish the vasoconstrictor effects of LBNP. Furthermore, these data demonstrate that this sympatholysis occurs early in exercise. PMID- 12133865 TI - Exercise-induced elevation of HSP70 is intensity dependent. AB - Exercise induces expression of the protective heat shock protein, HSP70, in striated muscle. To characterize the relationship between induction of this protein and exercise intensity in muscles exhibiting different recruitment patterns, male Sprague-Dawley rats were assigned to a sedentary control or one of seven exercise groups for which treadmill running speed varied between 15 and 33 m/min (n = 8/group). Twenty-four hours after a single 60-min exercise bout, hearts, red and white portions of the vastus (RV and WV, respectively) muscles, and soleus (Sol) muscles were harvested and analyzed for both relative and absolute HSP70 content. Cardiac HSP70 was significantly elevated only when animals were exercised at 24 m/min and beyond. Similarly, HSP70 was elevated in RV at running speeds above 24 m/min but did not increase in WV until 27 m/min. In contrast, HSP70 content was initially elevated in the Sol but subsequently declined at the highest running speeds. The observed patterns of HSP70 expression in skeletal muscle were in general accordance with known muscle recruitment patterns and suggest that alterations in muscle loading, resulting from changes in exercise intensity, are an important component of exercise-induced increases in HSP70 content. PMID- 12133866 TI - Hormonal, renal, hemodynamic responses to acute neutral endopeptidase inhibition in heart transplant patients. AB - We investigated the hemodynamic, renal, and hormonal responses to neutral endopeptidase (NEP) inhibition during a 6-h, double-blind, randomized, placebo controlled study in seven chronic, stable heart transplant patients. Baseline characteristics were similar during both experiments, and no significant changes were observed after placebo. NEP inhibition increased circulating endothelin-1 (from 2.01 +/- 0.1 to 2.90 +/- 0.2 pmol/l; P < 0.01), atrial natriuretic peptide (ANP; from 21.5 +/- 2.7 to 29.6 +/- 3.7 pmol/l; P < 0.01), and the ANP second messenger cGMP. Noteworthy, systemic blood pressure did not increase. Renal plasma flow and glomerular filtration rate remained unmodified after NEP inhibition. Filtration fraction (33 +/- 13%), diuresis (196 +/- 62%), and natriuresis (315 +/- 105%) increased significantly in relation to ANP and cGMP. A strong inverse relationship was observed between excreted cGMP and sodium reabsorption (r = -0.71, P < 0.0001). Thus, despite significantly increasing endothelin-1, NEP inhibition did not adversely influence systemic or renal hemodynamics in transplant patients. ANP, possibly through a tubular action, enhances the natriuresis observed after NEP inhibition. PMID- 12133867 TI - Interactive effect of hypoxia and otolith organ engagement on cardiovascular regulation in humans. AB - We determined the interaction between the vestibulosympathetic reflex and the arterial chemoreflex in 12 healthy subjects. Subjects performed three trials in which continuous recordings of muscle sympathetic nerve activity (MSNA), mean arterial blood pressure (MAP), heart rate (HR), and arterial oxygen saturation were obtained. First, in prone subjects the otolith organs were engaged by use of head-down rotation (HDR). Second, the arterial chemoreflex was activated by inspiration of hypoxic gas (10% O2 and 90% N2) for 7 min with HDR being performed during minute 6. Third, hypoxia was repeated (15 min) with HDR being performed during minute 14. HDR [means +/- SE; increase (Delta)7 +/- 1 bursts/min and Delta50 +/- 11% for burst frequency and total MSNA, respectively; P < 0.05] and hypoxia (Delta6 +/- 2 bursts/min and Delta62 +/- 29%; P < 0.05) increased MSNA. Additionally, MSNA increased when HDR was performed during hypoxia (Delta11 +/- 2 bursts/min and Delta127 +/- 57% change from normoxia; P < 0.05). These increases in MSNA were similar to the algebraic sum of the individual increase in MSNA elicited by HDR and hypoxia (Delta13 +/- 1 bursts/min and Delta115 +/- 36%). Increases in MAP (Delta3 +/- 1 mmHg) and HR (Delta19 +/- 1 beats/min) during combined HDR and hypoxia generally were smaller (P < 0.05) than the algebraic sum of the individual responses (Delta5 +/- 1 mmHg and Delta24 +/- 2 beats/min for MAP and HR, respectively; P < 0.05). These findings indicate an additive interaction between the vestibulosympathetic reflex and arterial chemoreflex for MSNA. Therefore, it appears that MSNA outputs between the vestibulosympathetic reflex and arterial chemoreflex are independent of one another in humans. PMID- 12133868 TI - Contributions from rostral medullary nuclei to coordination of swallowing and breathing in awake goats. AB - The purpose of this study was to determine whether neurons in the facial (FN), gigantocellularis reticularis (RGN), and vestibular (VN) nuclei contribute to the regulation of breathing, swallowing, and the coordination of these two functions. Microtubules were chronically implanted bilaterally in goats. Two weeks later during wakefulness, 100-nl unilateral injections were made of mock cerebral spinal fluid or an excitatory amino acid receptor agonist or antagonists. When the agonist, N-methyl-D-aspartic acid, was injected into any nuclei, breathing and swallowing increased transiently (15-30%; P < 0.05), whereas only injections of the antagonist 2,3-dihydroxy-6-nitro-7-sulfamoylbenzo-(f)quinoxaline into VN increased swallowing (20%; P < 0.05). The phase of breathing in which the swallows occurred was not altered by any injections. However, more importantly, injections of the agonist and the antagonists significantly altered (P < 0.05) by 5-50% the respiratory phase-dependent timing and tidal volume effect of swallows on breathing relative to mock cerebral spinal fluid injections. In addition, these effects were not uniform for all three nuclei. We conclude that the FN, RGN, and VN are part of a neural circuit in the rostral medulla that regulates and/or modulates breathing, swallowing, and their coordination in the awake state. PMID- 12133869 TI - Circadian pancreatic enzyme pattern and relationship between secretory and motor activity in fasting humans. AB - It is unknown whether nonparallel pancreatic enzyme output occurs under basal conditions in humans. We aimed to determine whether the circadian or wake-sleep cycle influences the relationship among pancreatic enzymes or between pancreatic secretory and jejunal motor activity. Using orojejunal multilumen intubation, we measured enzyme outputs and proximal jejunal motility index during consecutive daytime and nighttime periods in each of seven fasting, healthy volunteers. Enzyme outputs were correlated tightly during daytime phases of wakefulness and nighttime phases of sleep (r > 0.72, P < 0.001). During nocturnal phases of wakefulness, output of proteases (r = 0.84, P < 0.001), but not of amylase and trypsin (r = 0.12), remained associated. Nocturnally, particularly during sleep, pancreatic secretory activity was directly correlated with jejunal motility index (r > 0.50, P < 0.001). In conclusion, parallel secretion of pancreatic enzymes dominates throughout the circadian cycle. Nonparallel secretion during nocturnal phases of wakefulness may be due to merely circadian effects or to the coupling of the wake-sleep and the circadian cycle. The association between fluctuations of secretory and motor activity appears to be particularly tight during the night. PMID- 12133870 TI - Hypoxemia and low Crs in vagally denervated lambs result from reduced lung volume and not pulmonary edema. AB - Vagal denervation performed in the intrathoracic region in newborn lambs leads to hypoxemia and decreased respiratory system compliance (Crs), which could result from atelectasis and/or pulmonary edema. The objective of the present study was to quantify the relative roles of alveolar derecruitment and pulmonary edema as underlying cause(s) of respiratory failure. Vagal denervation was performed in the intrathoracic region and below the recurrent laryngeal nerves in six newborn lambs within 24 h of birth, whereas six were sham operated. Pre- and postinflation Crs was measured to investigate the presence of alveolar derecruitment. Pulmonary edema was assessed with lung wet-dry-to-wet and lung tissue wet-to-dry ratios, total protein, and FITC-BSA recovery in lung tissue and bronchoalveolar lavage. Compared with that in the sham-operated animals, Crs was significantly lower in vagally denervated animals. However, postinflation, pulmonary system compliance obtained by quasi-static lung inflation and deflation to 30 cmH2O showed no significant difference between the sham-operated and denervated lambs. The lung wet-dry-to-wet and lung tissue wet-to-dry ratios, total protein, and FITC-BSA recovery in lung tissue and bronchoalveolar lavage were similar in denervated and sham-operated groups. We provide evidence that reduced lung volume and not pulmonary edema is associated with intrathoracic vagal denervation and is the likely underlying mechanism for hypoxemia and low Crs. PMID- 12133871 TI - Computational model of airway narrowing: mature vs. immature rabbit. AB - Immature rabbits have greater maximal airway narrowing and greater maximal fold increases in airway resistance during bronchoconstriction than mature animals. We have previously demonstrated that excised immature rabbit lungs have more distensible airways, a lower shear modulus, and structural differences in the relative composition and thickness of anatomically similar airways. In the present study, we incorporated anatomic and physiological data for mature and immature rabbits into a computational model of airway narrowing. We then investigated the relative importance of maturational differences in these factors as determinants of the greater airway narrowing that occurs in the immature animal. The immature model demonstrated greater sensitivity to agonist, as well as a greater maximal fold increase in airway resistance. Exchanging values for airway compliance between the mature and immature models resulted in the mature model exhibiting a greater maximal airway response than the immature model. In contrast, exchanging the shear moduli or the composition of the airway wall relative to the airway size produced relatively small changes in airway reactivity. Our results strongly suggest that the mechanical properties of the airway, i.e., greater compliance of the immature airway, can be an important factor contributing to the greater airway narrowing of the immature animal. PMID- 12133872 TI - Time course of lung ischemia-reperfusion-induced ICAM-1 expression and its role in ischemia-reperfusion lung injury. AB - Upregulation of intercellular adhesion molecule-1 (ICAM-1) expression is an important mechanism underlying ischemia-reperfusion (I/R) induced neutrophil activation and tissue injury in other organs. However, I/R of the lungs has not been shown to upregulate ICAM-1 expression. We determined the time course profile of lung I/R-induced ICAM-1 expression and assessed the role of ICAM-1 in mediating neutrophil sequestration, transmigration, and I/R injury in the isolated blood-perfused rat lungs. I/R had a biphasic effect on ICAM-1 expression, an early downregulation and a late-phase upregulation. Superoxide dismutase and neutrophil depletion prevented the early ICAM-1 downregulation. The late-phase ICAM-1 upregulation coincided with the I/R-induced increase in pulmonary microvascular leakage index. ICAM-1 monoclonal antibody (MAb) reversed the I/R-induced increase in pulmonary microvascular leakage index, with control antibody being ineffective. Neither I/R nor ICAM-1 MAb affected lung MPO activity and circulating neutrophil count. Lung I/R significantly increased bronchoalveolar lavage fluid neutrophil count and the GSSG-to-(GSSG+GSH) ratio. ICAM-1 MAb blocked the I/R-induced increase in GSSG-to-(GSSG+GSH) ratio but had no effect on bronchoalveolar lavage fluid neutrophil count. Our results demonstrated that lung I/R up- and downregulates ICAM-1 expression depending on the duration of reperfusion. ICAM-1 upregulation is an important mechanism of I/R induced pulmonary endothelial injury. PMID- 12133873 TI - Postnatal alveolar development of the rabbit. AB - Previous studies of alveolarization have used rats or lambs; however, neither closely reflects human alveolar development. We characterized alveolar development in rabbits (n = 3-7 /group) at 28 days gestation (dg) to 9 mo to determine whether they followed the human pattern more closely. The right lung was made up of 30% alveolar and 50% duct space at 28 dg to 3 days and of 50 and 30%, respectively, at 14 days to 9 mo. Tissue fraction and alveolar wall thickness decreased by 40% 28 dg to birth. At birth, approximately 4.5% of the number of alveoli seen at 9 mo were present, with alveolar number increasing progressively well into adulthood. The rate of alveolar formation was high around birth, decreasing progressively with age. Alveolar volume increased more than twofold (28 dg to birth) and continued to increase postnatally to 16 wk. Surface fraction decreased by 17% (28 dg to 3 days), after which it remained uniform. Our findings suggest that the timing of onset of alveolarization in humans and rabbits is similar and that rabbits may be used to model postnatal influences on alveolar development. PMID- 12133874 TI - Limb venous compliance in patients with idiopathic orthostatic intolerance and postural tachycardia. AB - Venous denervation and increased venous pooling may contribute to symptoms of orthostatic intolerance. We examined venous compliance in the calf and forearm in 11 orthostatic-intolerant patients and 15 age-matched controls over a range of pressures, during basal conditions and sympathetic excitation. Occlusion cuffs placed around the upper arm and thigh were inflated to 60 mmHg and deflated to 10 mmHg over 1 min. Limb volume was measured continuously with a mercury-in-Silastic strain gauge. Compliance was calculated as the numerical derivative of the pressure-volume curve. The pressure-volume relationship in the upper and lower extremities in the basal and sympathetically activated state was significantly lower in the orthostatic-intolerant patients (all P < 0.05). Sympathoexcitation lowered the pressure-volume relationship in the lower extremity in patients (P < 0.001) and controls (P < 0.01). Venous compliance was significantly less in patients in the lower extremity in the basal state over a range of pressures (P < 0.05). Venous compliance was less in patients compared with controls in the upper (P < 0.005) and lower extremities (P < 0.01) in the sympathetically activated state, but there were no differences at individual pressure levels. Sympathetic activation did not change venous compliance in the upper and lower extremity in patients and controls. Patients with orthostatic intolerance have reduced venous compliance in the lower extremity. Reduced compliance may limit the dynamic response to orthostatic change and thereby contribute to symptoms of orthostatic intolerance in this population group. PMID- 12133875 TI - Exercise-induced change in type 1 cytokine-producing CD8+ T cells is related to a decrease in memory T cells. AB - In response to exercise, both CD4(+) and CD8(+) T cells are mobilized to the blood, but the levels of these cells decline below preexercise values in the postexercise period. T cells are functionally polarized, depending on the cytokines they produce. Type 1 cells produce, e.g., interferon (INF)-gamma, whereas type 2 produce, e.g., interleukin (IL)-4. It was recently demonstrated that exercise induces a decrease in the percentage of type 1 T cells. The present study further investigated the mechanisms underlying the exercise-induced shift in the balance between type 1 and type 2 cytokine-producing cells. Seven healthy men performed 1.5 h of treadmill running with blood samples drawn before exercise, at the end of exercise, and 2 h after exercise. Intracellular expression of IFN-gamma, IL-2, and IL-4 was detected in CD4(+) and CD8(+) T cells after stimulation with phorbol 12-myristate 13-acetate and ionomycin. Intracellular expression of IFN-gamma within CD8(+) cells was decreased in the postexercise period compared with values obtained immediately after exercise, whereas the expression of IL-2 and IL-4 did not change within the CD4(+) and CD8(+) cell populations. The decrease in IFN-gamma-producing CD8(+) T cells postexercise was negatively correlated with a decrease in percentage of memory T cells within the CD8(+) cells (r = -0.94; P < or = 0.002). In conclusion, this study demonstrates that the exercise-induced change in type 1 cytokine-producing T cells is related to a decline in memory cells. PMID- 12133876 TI - Mechanical properties of the latissimus dorsi muscle after cyclic training. AB - Cardiomyoplasty is a procedure developed to improve heart performance in patients suffering from congestive heart failure. The latissimus dorsi (LD) muscle is surgically wrapped around the failing ventricles and stimulated to contract in synchrony with the heart. The LD muscle is easily fatigued and as a result is unsuitable for cardiomyoplasty. For useful operation as a cardiac-assist device, the fatigue resistance of the LD muscle must be improved while retaining a high power output. The LD muscle of rabbits was subjected to a training regime in which cyclic work was performed. Training transformed the fiber-type composition from approximately equal proportions of fast oxidative glycolytic (FOG) and fast glycolytic (FG) fibers to one composed of almost entirely of FOG with no FG, which increased fatigue resistance while retaining rapid contraction kinetics. Muscle mass and cross-sectional area increased but power output decreased, relative to control muscles. This training regime represents a significant improvement in terms of preserving muscle mass and power compared with other training regimes, while enhancing fatigue resistance, although some fiber damage occurred. The power output of the trained LD muscle was calculated to be sufficient to deliver a significant level of assistance to a failing heart during cardiomyoplasty. PMID- 12133877 TI - Detection of local lung air content by electrical impedance tomography compared with electron beam CT. AB - The aim of the study was to validate the ability of electrical impedance tomography (EIT) to detect local changes in air content, resulting from modified ventilator settings, by comparing EIT findings with electron beam computed tomography (EBCT) scans obtained under identical steady-state conditions. The experiments were carried out on six anesthetized supine pigs ventilated with five tidal volumes (VT) at three positive end-expiratory pressure (PEEP) levels. The lung air content changes were determined both by EIT (Goe-MF1 system) and EBCT (Imatron C-150XP scanner) in six regions of interest, located in the ventral, middle, and dorsal areas of each lung, with respect to the reference air content at the lowest VT and PEEP, as a change in either local electrical impedance or lung tissue density. An increase in local air content with VT and PEEP was identified by both methods at all regions studied. A good correlation between the changes in lung air content determined by EIT and EBCT was revealed. Mean correlation coefficients in the ventral, middle, and dorsal regions were 0.81, 0.87, and 0.93, respectively. The study confirms that EIT is a suitable, noninvasive method for detecting regional changes in air content and monitoring local effects of artificial ventilation. PMID- 12133878 TI - Sound transmission in the lung as a function of lung volume. AB - We were interested in how the transmission of sound through the lung was affected by varying air content in intact humans as a method of monitoring tissue properties noninvasively. To study this, we developed a method of measuring transthoracic sound transit time accurately. We introduced a "coded" sound at the mouth and measured the transit time at multiple microphones placed over the chest wall by using a 16-channel lung sound analyzer (Stethographics). We used a microphone placed over the neck near the trachea as our reference and utilized cross-correlation analysis to calculate the transit times. The use of the coded sound, composed of a mix of frequencies from 130 to 150 Hz, greatly reduced the ambiguity of the cross-correlation function. The measured transit time varied from 1 ms at the central locations to 5 ms at the lung bases. Our results also indicated that transit time at all locations decreased with increasing lung volume. We found that these results can be described in terms of a model in which sound transmission through the lung is treated as a combination of free-space propagation through the trachea and a propagation through a two-phase system in the parenchyma. PMID- 12133879 TI - Alternate muscle activity observed between knee extensor synergists during low level sustained contractions. AB - To determine quantitatively the features of alternate muscle activity between knee extensor synergists during low-level prolonged contraction, a surface electromyogram (EMG) was recorded from the rectus femoris (RF), vastus lateralis (VL), and vastus medialis (VM) in 11 subjects during isometric knee extension exercise at 2.5% of maximal voluntary contraction (MVC) for 60 min (experiment 1). Furthermore, to examine the relation between alternate muscle activity and contraction levels, six of the subjects also performed sustained knee extension at 5.0, 7.5, and 10.0% of MVC (experiment 2). Alternate muscle activity among the three muscles was assessed by quantitative analysis on the basis of the rate of integrated EMG sequences. In experiment 1, the number of alternations was significantly higher between RF and either VL or VM than between VL and VM. Moreover, the frequency of alternate muscle activity increased with time. In experiment 2, alternating muscle activity was found during contractions at 2.5 and 5.0% of MVC, although not at 7.5 and 10.0% of MVC, and the number of alternations was higher at 2.5 than at 5.0% of MVC. Thus the findings of the present study demonstrated that alternate muscle activity in the quadriceps muscle 1) appears only between biarticular RF muscle and monoarticular vasti muscles (VL and VM), and its frequency of alternations progressively increases with time, and 2) emerges under sustained contraction with force production levels < or =5.0% of MVC. PMID- 12133880 TI - State and chemical drive modulate respiratory variability. AB - The quantification of respiratory variability may provide insight into the integrative control of breathing. To test the hypothesis that sleep and/or increased chemical drive modifies respiratory variability, six male adult Sprague Dawley rats were instrumented with diaphragm electromyographic (EMG) electrodes and exposed to 0, 2.5, and 5.0% CO2 with a balance of room air during wakefulness and behaviorally determined sleep. Respiratory interval (Ttot), peak diaphragm EMG, and ventilation index (peak diaphragm EMG/Ttot) were calculated for 1,024 sequential breaths. The variability of breathing was quantified with a measurement of signal complexity, the approximate entropy, and two autocorrelation measurements, the autoregressive power spectrum slope and the detrended fluctuation analysis slope. Elevated chemical drive and/or sleep significantly modulated the variability of ventilation index and Ttot. There were also significant interactions between state and CO2 drive in all respiratory parameters. We conclude that state (sleep or wakefulness) and increased chemical drive affect respiratory variability differentially. PMID- 12133881 TI - Propranolol prevents epinephrine from limiting insulin-stimulated muscle glucose uptake during contraction. AB - Beta-blockade results in rapid glucose clearance and premature fatigue during exercise. To investigate the cause of this increased glucose clearance, we studied the acute effects of propranolol on insulin-stimulated muscle glucose uptake during contraction in the presence of epinephrine with an isolated rat muscle preparation. Glucose uptake increased in both fast- (epitrochlearis) and slow-twitch (soleus) muscle during insulin or contraction stimulation. In the presence of 24 nM epinephrine, glucose uptake during contraction was completely suppressed when insulin was present. This suppression of glucose uptake by epinephrine was accompanied by a decrease in insulin receptor substrate (IRS)-1 phosphatidylinositol 3 (PI3)-kinase activity. Propranolol had no direct effect on insulin-stimulated glucose uptake during contraction. However, epinephrine was ineffective in attenuating insulin-stimulated glucose uptake during contraction in the presence of propranolol. This ineffectiveness of epinephrine to suppress insulin-stimulated glucose uptake during contraction occurred in conjunction with its inability to completely suppress IRS-1-PI3-kinase activity. Results of this study indicate that the effectiveness of epinephrine to inhibit insulin stimulated glucose uptake during contraction is severely diminished in muscle exposed to propranolol. Thus the increase in glucose clearance and premature fatigue associated with beta-blockade could result from the inability of epinephrine to attenuate insulin-stimulated muscle glucose uptake. PMID- 12133882 TI - Time dependence of recruitment and derecruitment in the lung: a theoretical model. AB - Recruitment and derecruitment (R/D) of air spaces within the lung is greatly enhanced in lung injury and is thought to be responsible for exacerbating injury during mechanical ventilation. There is evidence to suggest that R/D is a time dependent phenomenon. We have developed a computer model of the lung consisting of a parallel arrangement of airways and alveolar units. Each airway has a critical pressure (Pcrit) above which it tends to open and below which it tends to close but at a rate determined by how far pressure is from Pcrit. With an appropriate distribution of Pcrit and R/D velocity characteristics, the model able to produce realistic first and second pressure-volume curves of a lung inflated from an initially degassed state. The model also predicts that lung elastance will increase transiently after a deep inflation to a degree that increases as lung volume decreases and as the lung becomes injured. We conclude that our model captures the time-dependent mechanical behavior of the lung due to gradual R/D of lung units. PMID- 12133883 TI - Energy balance, metabolism, hydration, and performance during strenuous hill walking: the effect of age. AB - We aimed to examine the effect of age on energy balance, metabolism, hydration, and performance during 10 days of strenuous hill walking. Seventeen male subjects were divided into two groups according to their age. The nine subjects in group 1 constituted the younger group (age 24 +/- 3 yr), whereas eight older subjects were in group 2 (age 56 +/- 3 yr). Both groups completed 10 consecutive days of high-intensity hill walking. Mean (range) daily walking distances and ascent were 21 km (10-35 km) and 1,160 m (800-2,540 m), respectively. Energy intake was calculated from weighed food intake, and energy expenditure was measured by the doubly labeled water method. Blood and urine were sampled on alternative days to determine any changes in metabolism and hydration during the 10 days. Subjects also completed a battery of tests that included muscular strength (handgrip), jump performance, cognitive processing time, and flexibility. The younger group remained hydrated, whereas the older group became progressively dehydrated, indicated by a near twofold increase in urine osmolality concentration on day 11. This increased urine osmolality in the older group was highly correlated with impairment in vertical-jump performance (r = -0.86; P < 0.05) and decreased cognitive processing time (r = 0.79; P < 0.05). Despite energy expenditure of approximately 21 MJ/day, body mass was well maintained in both groups. Both groups displayed a marked increase in fat mobilization, reflected in significantly lowered prewalk insulin concentrations and elevated postwalk glycerol and nonesterified fatty acid concentrations. Despite the dehydration and impaired performance in the older group, blood glucose concentrations were well maintained in both groups, probably mediated via the increased mobilization of fat. PMID- 12133884 TI - Bicarbonate attenuates arterial desaturation during maximal exercise in humans. AB - The contribution of pH to exercise-induced arterial O2 desaturation was evaluated by intravenous infusion of sodium bicarbonate (Bic, 1 M; 200-350 ml) or an equal volume of saline (Sal; 1 M) at a constant infusion rate during a "2,000-m" maximal ergometer row in five male oarsmen. Blood-gas variables were corrected to the increase in blood temperature from 36.5 +/- 0.3 to 38.9 +/- 0.1 degrees C (P < 0.05; means +/- SE), which was established in a pilot study. During Sal exercise, pH decreased from 7.42 +/- 0.01 at rest to 7.07 +/- 0.02 but only to 7.34 +/- 0.02 (P < 0.05) during the Bic trial. Arterial PO2 was reduced from 103.1 +/- 0.7 to 88.2 +/- 1.3 Torr during exercise with Sal, and this reduction was not significantly affected by Bic. Arterial O2 saturation was 97.5 +/- 0.2% at rest and decreased to 89.0 +/- 0.7% during Sal exercise but only to 94.1 +/- 1% with Bic (P < 0.05). Arterial PCO2 was not significantly changed from resting values in the last minute of Sal exercise, but in the Bic trial it increased from 40.5 +/- 0.5 to 45.9 +/- 2.0 Torr (P < 0.05). Pulmonary ventilation was lowered during exercise with Bic (155 +/- 14 vs. 142 +/- 13 l/min; P < 0.05), but the exercise-induced increase in the difference between the end-tidal O2 pressure and arterial PO2 was similar in the two trials. Also, pulmonary O2 uptake and changes in muscle oxygenation as determined by near-infrared spectrophotometry during exercise were similar. The enlarged blood-buffering capacity after infusion of Bic attenuated acidosis and in turn arterial desaturation during maximal exercise. PMID- 12133885 TI - IGF-I, IgA, and IgG responses to bovine colostrum supplementation during training. AB - This study examined the effect of bovine colostrum (Dynamic colostrum) supplementation on blood and saliva variables (study 1) and the absorption of orally administered human recombinant insulin-like growth factor (IGF)-I (rhIGF I) labeled with 123I (123I-rhIGF-I) (study 2). In study 1, adult male and female athletes were randomly assigned in a double-blind fashion to either an experimental (Dynamic; n = 19) or a control (Placebo; n = 11) group. The former consumed daily 20 g of Dynamic supplement, and the latter 20 g of maltodextrin during a 2-wk training period. After bovine colostrum supplementation, significant increases were noticed in serum IGF-I (P < 0.01) and saliva IgA (P < 0.01) in Dynamic compared with Placebo. In study 2, gel electrophoresis was carried out in 12 adult subjects with serum samples taken 60 min after ingestion of 123I-rhIGF-I and showed peaks at 0.6 and at 40-90 kDa, with the former inducing 96% and the latter 4% of the total radioactivity. It was concluded that a long-term supplementation of bovine colostrum (Dynamic) increases serum IGF-I and saliva IgA concentration in athletes during training. Absorption data show that ingested 123I-rhIGF-I is fragmented in circulation and that no radioactive IGF-I is eluted at the positions of free, or the IGF, binding proteins, giving no support to the absorption of IGF-I from bovine colostrum. PMID- 12133886 TI - Laryngeal activity during upright vs. supine swallowing. AB - Previous investigations of human pharyngeal muscle activation patterns during swallowing found a relatively invariant muscle activation onset sequence in the upright position. However, different gravitational forces influence a liquid bolus when supine and could modify the central timing control of laryngeal airway protection during swallowing. The purpose of this study was to determine whether laryngeal muscle onset timing during swallowing differed between the supine and upright positions. Nine subjects performed six swallowing trials with a 2-ml water bolus in each position. Simultaneous electromyographic recordings were obtained from the submental complex (SMC) and the right and left thyroarytenoid (TA) muscles. Regardless of body position, the timing, amplitude, and duration of the TA muscles did not vary relative to the SMC. Therefore, the sequence of TA muscle activation relative to the SMC during swallowing appeared unaffected by gravitational influences. PMID- 12133887 TI - Earliest cardiac toxicity induced by iron overload selectively inhibits electrical conduction. AB - Female guinea pigs were injected intraperitoneally with 0.083 g/kg iron dextran (Fe-D) to achieve progressively increasing levels of iron load; controls received dextran. Delayed and blocked cardiac conductivity at the Purkinje fiber-papillary muscle junction was initially observed with Fe-D loads of 0.33 g/kg. Serial magnetic resonance relaxation time measurements obtained from livers of live animals showed a decrease (8.1 +/- 0.86 vs. 14.8 +/- 1.03 ms in controls, P < 0.001) that was first observed in animals loaded with 0.25 g/kg Fe-D. Iron concentrations in hearts and livers were significantly increased (P < 0.001). Left ventricular pressure measurements on 1.5 g/kg Fe-D animals failed to demonstrate a defect in contractility, but 27% (9/33) (P < 0.050) of the animals died without warning signs. We conclude that 1) initial decreases in liver magnetic resonance-relaxation time occur in the same range of iron excess as the threshold of iron load that induces delay or blockade of cardiac conduction and 2) a high incidence of sudden death, presumably from cardiac arrhythmias, was observed with large doses of iron that did not decrease left ventricular contractility. PMID- 12133888 TI - Rapid effects of 17beta-estradiol and progesterone on sheep visceral and parietal pleurae via a nitric oxide pathway. AB - We investigated the effects of 17beta-estradiol and progesterone on transepithelial electrical resistance (R(TE)) in sheep visceral and parietal pleurae. Specimens of intact pleurae from adult female sheep were used. The samples were transferred to the laboratory within 30 min after death of the animal in a Krebs-Ringer solution at 4 degrees C. The pleura was then mounted as a planar sheet in Ussing-type chambers, and electrical measurements were made. There was an increase in R(TE) in all of the samples examined after addition of 17beta-estradiol and progesterone in visceral and parietal pleurae. This increase was rapid within 1 min, lasted for ~15 min, returned to the basal level within 30 45 min, and was dose dependent. Tamoxifen, an estrogen receptor antagonist, did not significantly eliminate the effect of 17beta-estradiol. Furthermore, no steroid receptors were identified in cytosolic preparations of visceral and parietal pleura with ligand binding assays. The estrogen- and progesterone induced increase in R(TE) in both visceral and parietal pleurae was affected by addition of an inhibitor of nitric oxide synthase. Indeed, previous administration of N(omega)-nitro-L-arginine methyl ester prevented the increase in R(TE) by 17beta-estradiol and progesterone. These results suggest that 17beta estradiol and progesterone induce an increase in R(TE) in both visceral and parietal pleura and thus alter the transepithelial permeability. The effect of steroids may be accounted for by rapid release of nitric oxide in pleura. PMID- 12133889 TI - A new surface electromyography analysis method to determine spread of muscle fiber conduction velocities. AB - Muscle fiber conduction velocity (MFCV) estimation from surface signals is widely used to study muscle function, e.g., in neuromuscular disease and in fatigue studies. However, most analysis methods do not yield information about the velocity distribution of the various motor unit action potentials. We have developed a new method-the interpeak latency method (IPL)-to calculate both the mean MFCV and the spread of conduction velocities in vivo, from bipolar surface electromyogram (sEMG) during isometric contractions. sEMG was analyzed in the biceps brachii muscle in 15 young male volunteers. The motor unit action potential peaks are automatically detected with a computer program. Associated peaks are used to calculate a mean MFCV and the SD. The SD is taken as a measure of the MFCV spread. The main finding is that the IPL method can derive a measure of MFCV spread at different contraction levels. In conclusion, the IPL method provides accurate values for the MFCV and additionally gives information about the scatter of conduction velocities. PMID- 12133890 TI - Exercise and insulin signaling: a historical perspective. AB - Over the past 30 years, a considerable body of evidence has revealed that a prior bout of exercise can increase the ability of insulin to stimulate glucose transport and glycogen synthesis in skeletal muscle. Apart from its clinical implications, this work has led to a considerable effort to determine at a molecular level how exercise causes this effect and, in particular, whether it does so by enhancing specific events in the insulin-signaling cascade. The objective of this review is to discuss from a historical perspective how our current thinking in this area has evolved and the people responsible for it. Areas to be discussed include the effect or lack of effect of prior exercise on the insulin-signaling pathway, effects of exercise on the regulation by insulin of the GLUT-4 glucose transporter in muscle, and the emerging role of AMP activated protein kinase as a mediator of exercise-induced signaling events. In addition, we will discuss briefly some of the avenues that research in this area is likely to follow. PMID- 12133891 TI - Invited review: Exercise training-induced changes in insulin signaling in skeletal muscle. AB - This review will provide insight on the current understanding of the intracellular signaling mechanisms by which exercise training increases glucose metabolism and gene expression in skeletal muscle. Participation in regular exercise programs can have important clinical implications, leading to improved health in insulin-resistant persons. Evidence is emerging that insulin signal transduction at the level of insulin receptor substrates 1 and 2, as well as phosphatidylinositol 3-kinase, is enhanced in skeletal muscle after exercise training. This is clinically relevant because insulin signaling is impaired in skeletal muscle from insulin-resistant Type 2 diabetic and obese humans. The molecular mechanism for enhanced insulin-stimulated glucose uptake after exercise training may be partly related to increased expression and activity of key proteins known to regulate glucose metabolism in skeletal muscle. Exercise also leads to an insulin-independent increase in glucose transport, mediated in part by AMP-activated protein kinase. Changes in protein expression may be related to increased signal transduction through the mitogen-activated protein kinase signaling cascades, a pathway known to regulate transcriptional activity. Understanding the molecular mechanism for the activation of insulin signal transduction pathways after exercise training may provide novel entry points for new strategies to enhance glucose metabolism and for improved health in the general population. PMID- 12133892 TI - Invited review: Regulation of skeletal muscle GLUT-4 expression by exercise. AB - The amount of GLUT-4 protein is a primary factor in determining the maximal rate of glucose transport into skeletal muscle. Therefore, it is important that we understand how exercise regulates GLUT-4 expression so that therapeutic strategies can be designed to increase muscle glucose disposal as a treatment for diabetes. Muscle contraction increases the rates of GLUT-4 transcription and translation. Transcriptional control likely requires at least two DNA binding proteins, myocyte enhancer factor-2 and GLUT-4 enhancer factor, which bind to the promoter. Increased GLUT-4 expression may be mediated by the enzyme AMP-activated kinase, which is activated during exercise and has been demonstrated to increase GLUT-4 transcription. Further research needs to be done to investigate whether AMP-activated kinase activates myocyte enhancer factor-2 and GLUT-4 enhancer factor to increase transcription of the GLUT-4 gene. PMID- 12133893 TI - Invited review: Effects of acute exercise and exercise training on insulin resistance. AB - Insulin resistance of skeletal muscle glucose transport is a key defect in the development of impaired glucose tolerance and Type 2 diabetes. It is well established that both an acute bout of exercise and chronic endurance exercise training can have beneficial effects on insulin action in insulin-resistant states. This review summarizes the present state of knowledge regarding these effects in the obese Zucker rat, a widely used rodent model of obesity-associated insulin resistance, and in insulin-resistant humans with impaired glucose tolerance or Type 2 diabetes. A single bout of prolonged aerobic exercise (30-60 min at approximately 60-70% of maximal oxygen consumption) can significantly lower plasma glucose levels, owing to normal contraction-induced stimulation of GLUT-4 glucose transporter translocation and glucose transport activity in insulin-resistant skeletal muscle. However, little is currently known about the effects of acute exercise on muscle insulin signaling in the postexercise state in insulin-resistant individuals. A well-established adaptive response to exercise training in conditions of insulin resistance is improved glucose tolerance and enhanced skeletal muscle insulin sensitivity of glucose transport. This training-induced enhancement of insulin action is associated with upregulation of specific components of the glucose transport system in insulin resistant muscle and includes increased protein expression of GLUT-4 and insulin receptor substrate-1. It is clear that further investigations are needed to further elucidate the specific molecular mechanisms underlying the beneficial effects of acute exercise and exercise training on the glucose transport system in insulin-resistant mammalian skeletal muscle. PMID- 12133895 TI - Evidence that the decrease in liver glycogen is associated with the exercise induced increase in IGFBP-1. AB - The purpose of the present study was to test the hypothesis that the exercise induced increase in insulin-like growth factor binding protein (IGFBP)-1 is not always linked to a decrease in blood glucose level and to examine whether the decreasing levels of liver glycogen during exercise may be associated with the increase in IGFBP-1. Three groups of rats were submitted to a 70-min treadmill exercise. One group of rats was fed normally, and the two other groups had their food intake restricted by 50% (50% fast) the night before the experiment. One of these two 50% fasted groups of rats was infused (intravenously) with glucose throughout exercise to maintain euglycemia. Exercise in noninfused 50% fasted rats, compared with the normally fed rats, resulted in significantly lower blood glucose (minute 70) and insulin levels, significantly lower liver glycogen content, no change in IGF-I, and significantly higher increases in free fatty acid, glycerol, beta-hydroxybutyrate, and IGFBP-1. Maintenance of euglycemia during exercise in glucose-infused 50% fasted rats reduced to a large extent the decrease in insulin levels but only slightly attenuated the lipid response and the IGFBP-1 response seen in noninfused 50% fasted rats. Comparisons of all individual liver glycogen and IGFBP-1 values revealed that liver glycogen values were highly (P < 0.001) predictive of the IGFBP-1 response during exercise (R = 0.564). The present results indicate that the IGFBP-1 response during exercise is not always linked to a decrease in plasma glucose and suggest that the increase in IGFBP-1 during exercise may be related to the decrease in liver glycogen content. PMID- 12133896 TI - Modulation of insulin resistance and hypertension by voluntary exercise training in the TG(mREN2)27 rat. AB - Hypertension is often accompanied by insulin resistance of skeletal muscle glucose transport. The male heterozygous TG(mREN2)27 rat, which harbors a mouse transgene for renin, displays local elevations in the renin-angiotensin system and exhibits markedly elevated systolic blood pressure (SBP). The present study was undertaken to characterize insulin-stimulated skeletal muscle glucose transport in male heterozygous TG(mREN2)27 rats and to evaluate the effect of voluntary exercise training on SBP and skeletal muscle glucose transport. Compared with normotensive Sprague-Dawley rats, TG(mREN2)27 rats displayed a 53% elevation (P < 0.05) in SBP, a twofold increase in plasma free fatty acid levels, and an exaggerated insulin response during an oral glucose tolerance test. Moreover, insulin-mediated glucose transport (2-deoxyglucose uptake) in isolated epitrochlearis and soleus muscles of TG(mREN2)27 animals was 33 and 43% less, respectively, than in Sprague-Dawley controls. TG(mREN2)27 rats ran voluntarily for 6 wk and achieved daily running distances of 6-7 km over the final 3 wk. Training caused a 36% increase in peak aerobic capacity and a 16% reduction in resting SBP. Fasting plasma insulin (21%) and free fatty acid (34%) levels were reduced in the trained TG(mREN2)27 rats. Whole body glucose tolerance was improved in the trained TG(mREN2)27 rats and was associated with increases of 39 and 50% in insulin-mediated glucose transport in epitrochlearis and soleus muscles, respectively. Whole muscle GLUT-4 protein was increased in the soleus (23%), but not in the epitrochlearis, of trained TG(mREN2)27 rats. These data indicate that the male heterozygous TG(mREN2)27 rat is a model of both hypertension and insulin resistance. Importantly, both of these defects can be beneficially modified by voluntary exercise training. PMID- 12133897 TI - The inducible expression of the tumor suppressor gene PTEN promotes apoptosis and decreases cell size by inhibiting the PI3K/Akt pathway in Jurkat T cells. AB - In this study, we characterize the function of the tumor suppressor gene PTEN in Jurkat T cells. We established stable clones of Jurkat T cells that inducibly express either wild-type or phosphatase-inactive PTEN. We show here that PTEN potently inhibited the growth and reduced the size of Jurkat cells. The growth suppressive effect of PTEN was associated with its ability to induce apoptotic cell death with little or no effect on cell cycle. PTEN also rendered Jurkat cells more susceptible to apoptosis induced by various stimuli. Furthermore, PTEN expression led to a reduction in the level of 3'-phosphorylated phospholipids and thus altered the activity and localization of Akt. Finally, coexpression of constitutively active Akt reversed the effects caused by PTEN. In summary, our results suggest that PTEN suppresses cell growth, promotes apoptosis, and decreases cell size by negatively regulating the phosphoinositide 3-kinase/Akt pathway in Jurkat T cells. PMID- 12133898 TI - The Wilms' tumor suppressor Wt1 is associated with the differentiation of retinoblastoma cells. AB - We have demonstrated recently that Wilms' tumor suppressor 1 (Wt1),in addition to its role in genitourinary formation,is required for the differentiation of ganglion cells in the developing retina. Here we provide further evidence that Wt1 is associated with neuronal differentiation. Thus, the retinoblastoma-derived human cell line, Y-79, contained robust amounts of Wt1 mRNA and protein. Wt1 expression was down-regulated upon laminin-induced differentiation of Y-79 into neuron-like cells. Inhibition of Wt1 with antisense oligonucleotides dramatically reduced the capacity of undifferentiated Y-79 cells to undergo neuronal differentiation, whereas sense and missense oligonucleotides had no effect. Wt1 immunoreactivity was also detected in solid retinoblastomas, in which it resided mainly in areas with moderate proliferative activity. These findings suggest a role for Wt1 in the differentiation of retinoblastoma cells. Furthermore, Wt1 expression in retinoblastoma may reflect the potential of these tumors to initiate the early steps of neuronal differentiation. PMID- 12133899 TI - Expression of kinase suppressor of Ras in the normal adult and embryonic mouse. AB - Recent studies indicate that kinase suppressor of Ras (KSR)is a scaffold protein for the Ras/Raf/MEK/ERK signaling cascade in mammals. To help determine the in vivo function of KSR, we have examined the tissue-specific distribution of this protein in the embryonic and adult mouse using a rat monoclonal antibody raised against the mouse protein. Western blot analysis indicates that the protein is expressed at highest levels in the adult brain. It is also expressed at low levels in bladder, ovary, testis, and lung, but the protein is not detectable in any other adult tissue. However, reverse transcription-PCR analysis shows that Ksr transcripts are detected in all adult tissues except the liver. A variant containing a differentially spliced exon in the CA4 domain is observed in brain, cerebellum, ovary, and intestine. The protein is also expressed throughout the E6.5 embryo and at high levels in the neuroepithelium of the E10.5 embryo. At this embryonic stage, expression is also detected at lower levels in the limb and tail buds as well as in the myocardium. PMID- 12133900 TI - Lack of Fas/CD95 surface expression in highly proliferative leukemic cell lines correlates with loss of CtBP/BARS and redirection of the protein toward giant lysosomal structures. AB - Fas/CD95 is a type-I membrane glycoprotein, which inducesapoptotic cell death when ligated by its physiological ligand. We generated previously hyperproliferative sublines derived from the human T-cell leukemia Jurkat, Jurkat ws and Jurkat-hp, which lost Fas/CD95 surface expression. We have now observed that the total amount of Fas protein is similar in the sublines and in the parental cells, indicating that in the sublines Fas remains in an intracellular compartment. We have found that the protein is directed toward lysosomes in the sublines, where it is degraded. This defect in the secretory pathway correlates with loss of polyunsaturated fatty acids from cellular lipids, and with the lack of expression of endophilin-I and CtBP/BARS, enzymes that regulate vesicle fission by catalyzing the acylation of arachidonate into lysophosphatidic acid. In addition, great multillamer bodies, which contained acid phosphatase activity, absent in the parental Jurkat cells, were observed by transmission electron microscopy in the sublines. PMID- 12133901 TI - Both alpha and beta isoforms of mammalian DNA topoisomerase II associate with chromosomes in mitosis. AB - Two isoforms of DNA topoisomerase II, alpha and beta, coded by separate genes, are expressed in actively cycling vertebrate cells. Some previous studies have suggested that only topoisomerase II alpha remains associated with chromosomes at mitosis. Here, the distributions of topoisomerase II alpha and beta in mitosis were studied by subcellular fractionation and by immunolocalization. Both isoforms of topoisomerase II were found to remain associated with mitotic chromatin. Topoisomerase II alpha was distributed along chromosome arms throughout mitosis and was highly concentrated at centromeres until mid-anaphase, particularly in some cell types. Topoisomerase II beta showed weak concentration at centromeres in early mitosis in some cell types and was distributed along chromosome arms at every stage of mitosis through telophase. These studies suggest that in most cells both the major topoisomerase II isoforms may play roles in chromatin remodeling during M phase. PMID- 12133902 TI - An antigen expressed during plant vascular development crossreacts with antibodies towards KLH (keyhole limpet hemocyanin). AB - An antigen present in plant vascular tissue crossreacts with antibodies towards keyhole limpet hemocyanin (KLH). The antigen is present in xylem and vascular cambium, as evidenced by immunocytochemical staining of plant sections. This cell type assignment was confirmed by staining of mesophyll cell cultures from Zinnia elegans L. undergoing tracheary cell differentiation. The strongest staining both in sections and cell cultures occurred in cells and tissues during early stages of differentiation. Although the anti-KLH antibodies can easily be removed by affinity purification, our findings suggest that a certain caution is needed when KLH is used as an immunological carrier for studies in plants. PMID- 12133903 TI - DNA extraction from archival formalin-fixed, paraffin-embedded tissue sections based on the antigen retrieval principle: heating under the influence of pH. AB - During the course of diagnostic surgical pathology, pathologists have established a large collection of formalin-fixed, paraffin-embedded tissues that form invaluable resources for translational studies of cancer and a variety of other diseases. Accessibility of macromolecules in the fixed tissue specimens is a critical issue as exemplified by heat-induced antigen retrieval (AR) immunohistochemical (IHC) staining. On the basis of observations that heating may also enhance in situ hybridization (ISH) and the similarity of formalin-induced chemical modifications that occur in protein and in DNA, we designed a study to examine the efficiency of DNA extraction from archival formalin-fixed, paraffin embedded tissues using an adaptation of the basic principles of the AR technique, i.e., heating the tissue under the influence of different pH values. Archival paraffin blocks of lymph nodes, tonsil, and colon were randomly selected. Each paraffin block was prepared in 34 microtubes. For each paraffin block, one tube was used as a control sample, using a non-heating DNA extraction protocol. The other 33 tubes were tested using a heating protocol under 11 variable pH values (pH 2 to 12) under three different heating conditions (80, 100, and 120C). Evaluation of the results of DNA extraction was carried out by measuring yields by photometry and PCR amplification, as well as kinetic thermocycling (KTC)-PCR methods. In general, lower pH (acid) solutions gave inferior results to solutions at higher pH (alkaline). Heating tissues at a higher temperature and at pH 6-9 gave higher yields of DNA. There appeared to be a peak in terms of highest efficiency of extracted DNA at around pH 9. The average ratios 260:280 of extracted DNA also showed better values for samples heated at 120C. PCR products of three primers showed satisfactory results for DNA extracted from archival paraffin-embedded tissues by heating protocols at pH 6-12, with results that were comparable to the control sample subjected to the standard non-heating, enzymatic DNA extraction method. This study is the first to document the use of heating at an alkaline pH for DNA extraction from archival formalin-fixed, paraffin-embedded tissues, a recommendation based on the principles of AR for protein IHC. These findings may lead to a more effective protocol for DNA extraction from archival paraffin-embedded tissues and may also provide enhanced understanding of changes that occur during formalin-induced modification of nucleic acids. PMID- 12133904 TI - Ghrelin expression in fetal, infant, and adult human lung. AB - Ghrelin is a recently identified hormone with potent growth hormone (GH) releasing activity. It is produced by rat and human gastric endocrine cells and by the pituitary, hypothalamus, placenta, and by gastroenteropancreatic tumors. No evidence of ghrelin production by foregut-derived organs other than stomach has been provided to date. The aim of the present study was to investigate ghrelin expression by human fetal (20 cases), infant (13 cases), and adult (seven cases) lungs by immunohistochemistry, in situ hybridization, and RT-PCR. Expression of the GH secretagogue receptor, the endogenous receptor for ghrelin, was also investigated by RT-PCR. Ghrelin protein was found in the endocrine cells of the fetal lung in decreasing amounts from embryonic to late fetal periods. Its expression was maintained in newborns and children under 2 years but was virtually absent in older individuals. Scattered positive cells were also found in the trachea and the esophagus. Ghrelin mRNA was detected in adult lung by the more sensitive RT-PCR technique. GHS receptor mRNA was detected in nine cases of infant and adult lungs, possibly indicating the existence of local autocrine circuits. We conclude that the fetal lung is an additional source of circulating ghrelin, whose functions at the respiratory tract level remain to be clarified. PMID- 12133905 TI - Region-specific antibodies to chromogranin B display various immunostaining patterns in human endocrine pancreas. AB - Chromogranin (Cg) B is an acidic glycoprotein present in neuroendocrine tissue. The sequence shows several dibasic amino acid positions susceptible to proteolytic cleavage. The purpose of this study was to elucidate the expression of CgB epitopes in the human endocrine pancreas. Tissue sections of six human pancreata were immunostained with 16 different region-specific antibodies to the CgB molecule, using double immunofluorescence techniques. The CgB epitope pattern varied in the four major islet cell types. B (insulin)-cells expressed immunoreactivity to all region-specific antibodies. The antibodies to the N terminal and mid-portions of CgB showed moderate immunoreactivity, the C-terminal antibodies weak. A (glucagon)-cells were reactive only to the N-terminal and mid portion antibodies but, after microwave pretreatment, to all antibodies, whereas D (somatostatin)-cells expressed only the sequence CgB 244-255 and a subpopulation CgB 580-595. PP (pancreatic polypeptide) cells were immunostained with antibodies between CgB 1-417 and a few with CgB 580-593. The fragment CgB 244-255 was expressed in all four cell types. The cause of these differences may be cell-specific cleavage or masking of the molecule, but varying translation of CgB mRNA is also possible. The extent to which these epitopes reflect fragments having biological functions remains to be evaluated. PMID- 12133906 TI - Methods to enhance signal using isotopic in situ hybridization. AB - Isotopic in situ hybridization (ISH) has been established as a uniquely powerful tool for the study of gene expression in specific cell types. This technique allows the visualization and quantification of gene expression and gene expression changes in cells. In our study of biological and molecular phenomena, we have increasingly encountered the need to detect small changes in gene expression as well as genes of low abundance, such as the oxytocin receptor (OTR) and the tuberoinfundibular peptide of 39 residues (Tip39). To increase the sensitivity of isotopic ISH for detection of rare mRNAs, we performed ISH on cryostat sections of rat hypothalamus and thalamus with 35S-labeled riboprobes and amplified the signal by hybridizing over 2 nights as well as labeling the probe with both [35S]-UTP and [35S]-ATP. These two methods of enhancement independently and in combination demonstrated a dramatic increase in signal, allowing the visualization of low levels of gene expression previously undetectable by conventional methods. PMID- 12133907 TI - Keratan sulfate epitopes exhibit a conserved distribution during joint development that remains undisclosed on the basis of glycosaminoglycan charge density. AB - Changes in glycosaminoglycan (GAG) content and distribution are vital for joint development. However, their precise character has not been established. We have used immunohistochemistry (IHC) and "critical electrolyte" Alcian blue staining to assess such changes in developing chick and rabbit joints. IHC showed chondroitin sulfate labeling in chick epiphyseal cartilage but not in interzones. In contrast, prominent labeling for keratan sulfate (KS) was restricted to chick cartilage-interzone interfaces. In rabbit knees, KS labeling was also prominent at presumptive cavity borders, but weak in interzone and cartilage. Selective pre digestion produced appropriate loss of label and undersulfated KS was undetectable. Quantification of Alcian blue staining by scanning and integrating microdensitometry showed prominent hyaluronan-like (HA-like) interzone staining, with chondroitin sulfate and weaker KS staining restricted to epiphyseal cartilage. Hyaluronidase decreased HA-like staining in the interzone. Surprisingly, keratanases also reduced HA-like but not sulfated GAG (sGAG-like) staining in the interzone. Chondroitinase ABC had little effect on HA-like staining but decreased sGAG staining in all regions. Rabbit joints also showed HA like but not KS staining in the interzone and strong chondroitin sulfate-like staining in epiphyseal cartilage. Our findings show restricted KS distribution in the region close to the presumptive joint cavity of developing chick and rabbit joints. Alcian blue staining does not detect this moiety. Therefore, it appears that although histochemistry allows relatively insensitive quantitative assessment of GAGs, IHC increases these detection limits. This is particularly evident for KS, which exhibits immunolabeling patterns in joints from different species that is consistent with a conserved functional role in chondrogenesis. PMID- 12133908 TI - Immunochemical and mechanical characterization of cartilage subtypes in rabbit. AB - Cartilage is categorized into three general subgroups, hyaline, elastic, and fibrocartilage, based primarily on morphologic criteria and secondarily on collagen (Types I and II) and elastin content. To more precisely define the different cartilage subtypes, rabbit cartilage isolated from joint, nose, auricle, epiglottis, and meniscus was characterized by immunohistochemical (IHC) localization of elastin and of collagen Types I, II, V, VI, and X, by biochemical analysis of total glycosaminoglycan (GAG) content, and by biomechanical indentation assay. Toluidine blue staining and safranin-O staining were used for morphological assessment of the cartilage subtypes. IHC staining of the cartilage samples showed a characteristic pattern of staining for the collagen antibodies that varied in both location and intensity. Auricular cartilage is discriminated from other subtypes by interterritorial elastin staining and no staining for Type VI collagen. Epiglottal cartilage is characterized by positive elastin staining and intense staining for Type VI collagen. The unique pattern for nasal cartilage is intense staining for Type V collagen and collagen X, whereas articular cartilage is negative for elastin (interterritorially) and only weakly positive for collagen Types V and VI. Meniscal cartilage shows the greatest intensity of staining for Type I collagen, weak staining for collagens V and VI, and no staining with antibody to collagen Type X. Matching cartilage samples were categorized by total GAG content, which showed increasing total GAG content from elastic cartilage (auricle, epiglottis) to fibrocartilage (meniscus) to hyaline cartilage (nose, knee joint). Analysis of aggregate modulus showed nasal and auricular cartilage to have the greatest stiffness, epiglottal and meniscal tissue the lowest, and articular cartilage intermediate. This study illustrates the differences and identifies unique characteristics of the different cartilage subtypes in rabbits. The results provide a baseline of data for generating and evaluating engineered repair cartilage tissue synthesized in vitro or for post implantation analysis. PMID- 12133909 TI - Distribution of the transcription factors Sox9, AP-2, and [delta]EF1 in adult murine articular and meniscal cartilage and growth plate. AB - The control of extracellular matrix (ECM) production is important for the development, maintenance, and repair of cartilage tissues. Matrix molecule synthesis is generally regulated by the rate of gene transcription determined by DNA transcription factors. We have shown that transcription factors Sox9, AP-2, and [delta]EF1 are able to alter the rate of CD-RAP transcription in vitro: Sox9 upregulates, AP-2 exhibits biphasic effects, and [delta]EF1 represses expression of the CD-RAP gene. To correlate these in vitro activities in vivo, transcription factors were co-immunolocalized with ECM proteins in three different cartilage tissues in which the rates of biosynthesis are quite different: articular, meniscal, and growth plate. Immunoreactivities of type II collagen and CD-RAP were higher in growth plate than in either the articular or meniscal cartilages and correlated positively with Sox9 protein. Sox9 staining decreased with hypertrophy and was low in articular and meniscal cartilages. In contrast, AP-2 and [delta]EF1 were low in proliferating chondrocytes but high in lower growth plate, articular, and meniscal cartilages. This increase was also accompanied by intense nuclear staining. These immunohistochemical results are the first to localize both [delta]EF1 and AP-2 to adult articular, meniscal, and growth plate cartilages and provide in vivo correlation of previous molecular biological studies. PMID- 12133910 TI - A novel flat-embedding method to prepare ultrathin cryosections from cultured cells in their in situ orientation. AB - Immunogold labeling of ultrathin cryosections provides a sensitive and quantitative method to localize proteins at the ultrastructural level. An obligatory step in the routine preparation of cryosections from cultured cells is the detachment of cells from their substrate and subsequent pelleting. This procedure precludes visualization of cells in their in situ orientation and hampers the study of polarized cells. Here we describe a method to sample cultured cells from a petri dish or coverslip by embedding them in a 12% gelatin slab. Subsequently, sections can be prepared in parallel or perpendicular to the plane of growth. Our method extends the cryosectioning technique to applications in studying polarized cells and correlative light-electron microscopy. PMID- 12133911 TI - Cellular expression of gut chitinase mRNA in the gastrointestinal tract of mice and chickens. AB - Recently, the second mammalian chitinase, designated acidic mammalian chitinase (AMCase), has been identified in human, mouse, and cow. In contrast to the earlier identified macrophage-derived chitinase (chitotriosidase), this chitinase is richly expressed in the gastrointestinal (GI) tract, suggesting its role in digestion of chitin-containing foods as well as defense against chitin-coated microorganisms and parasites. This in situ hybridization study first revealed cellular localization of the gut-type chitinase in the mouse and chicken. In adult mice, the parotid gland, von Ebner's gland, and gastric chief cells, all of which are exocrine cells of the serous type, expressed the gut chitinase mRNA. In the chicken, oxyntico-peptic cells in glandular stomach (proventriculus) and hepatocytes expressed the chitinase mRNA. Because cattle produce the gut chitinase (chitin-binding protein b04) only in the liver, the gut chitinases in mammals and birds have three major sources of production, i.e., the salivary gland, stomach, and liver. During ontogenetic development, the expression level in the parotid gland and stomach of mice increased to the adult level before weaning, whereas in the stomach of chickens intense signals were detectable in embryos from incubation day 7. PMID- 12133912 TI - Dopamine D4 receptor expression in rat kidney: evidence for pre- and postjunctional localization. AB - Dopamine D4 receptors mediate inhibition of vasopressin-dependent sodium reabsorption by dopamine in collecting tubules. At present, the distribution of D4 receptors in other renal districts remains an open issue. The renal distribution of D4 receptor was assessed in normally innervated and denervated male Sprague-Dawley rats by quantitative immunohistochemistry using an anti dopamine D4 receptor rabbit polyclonal antibody. D4 receptor protein immunoreactivity was observed perivascularly in the adventitia and the adventitia media border. The density of perivascular dopamine D4 receptor was higher in afferent and efferent arterioles than in other segments of the renal vascular tree. Renal denervation abolished perivascular dopamine D4 receptor protein immunoreactivity. In renal tubules, the epithelium of collecting tubules showed the highest dopamine D4 receptor protein immunoreactivity, followed by the epithelium of proximal and distal tubules. No dopamine D4 receptor protein immunoreactivity was observed in the epithelium of the loop of Henle. Denervation did not change dopamine D4 receptor protein immunoreactivity in renal tubules. These results indicate that rat kidney expresses dopamine D4 receptors located both prejunctionally and nonprejunctionally in collecting, proximal, and distal tubules. This suggests that the dopamine D4 receptor may be involved in the control of neurotransmitter release and in renal hemodynamic and tubule function. PMID- 12133913 TI - New fiber formation in the interstitial spaces of rat skeletal muscle during postnatal growth. AB - The purpose of this study was to determine whether fiber hyperplasia occurs in the rat plantaris muscle during postnatal weeks 3-20. Total muscle fiber number, obtained via the nitric acid digestion method, increased by 28% during the early postnatal rapid growth phase (3-10 weeks), whereas the number of branched fibers was consistently low. Whole-muscle mitotic activity and amino acid uptake levels showed an inverse relationship to the increase in total fiber number. The expression of MyoD mRNA (RT-PCR) levels decreased from 3 to 20 weeks of age, as did the detection of anti-BrdU- and MyoD-positive cells in histological sections. Immunohistochemical staining patterns for MyoD, myogenin, or developmental myosin heavy chain on sections stained for laminin (identification of the basal lamina) and electron micrographs clearly indicate that de novo fiber formation occurred in the interstitial spaces. Myogenic cells in the interstitial spaces were negative for the reliable specific satellite cell marker M-cadherin. In contrast, CD34 (an established marker for hematopoietic stem cells)-positive cells were located only in the interstitial spaces, and their frequency and location were similar to those of MyoD- and/or myogenin-positive cells. These findings are consistent with fiber hyperplasia occurring in the interstitial spaces of the rat plantaris muscle during the rapid postnatal growth phase. Furthermore, these data suggest that the new fibers may be formed from myogenic cells in the interstitial spaces of skeletal muscle and may express CD34 that is distinct from satellite cells. PMID- 12133914 TI - Localization of laminin alpha4-chain in developing and adult human tissues. AB - Recent studies suggest important functions for laminin-8 (Ln-8; alpha4beta1gamma1) in vascular and blood cell biology, but its distribution in human tissues has remained elusive. We have raised a monoclonal antibody (MAb) FC10, and by enzyme-linked immunoassay (EIA) and Western blotting techniques we show that it recognizes the human Ln alpha4-chain. Immunoreactivity for the Ln alpha4-chain was localized in tissues of mesodermal origin, such as basement membranes (BMs) of endothelia, adipocytes, and skeletal, smooth, and cardiac muscle cells. In addition, the Ln alpha4-chain was found in regions of some epithelial BMs, including epidermis, salivary glands, pancreas, esophageal and gastric glands, intestinal crypts, and some renal medullary tubules. Developmental differences in the distribution of Ln alpha4-chain were detected in skeletal muscle, walls of vessels, and intestinal crypts. Ln alpha4- and Ln alpha2-chains co-localized in BMs of fetal skeletal muscle cells and in some epithelial BMs, e.g., in gastric glands and acini of pancreas. Cultured human pulmonary artery endothelial (HPAE) cells produced Ln alpha4-chain as M(r) 180,000 and 200,000 doublet and rapidly deposited it to the growth substratum. In cell-free extracellular matrices of human kidney and lung, Ln alpha4-chain was found as M(r) 180,000 protein. PMID- 12133915 TI - Monoclonal antibody CD133-2 (AC141) against hematopoietic stem cell antigen CD133 shows crossreactivity with cytokeratin 18. AB - CD133 is an antigen expressed on hematopoietic progenitor cells and on some epithelial cells. We previously reported that a commercially available antibody against CD133, CD133-2/AC141, also reacted with an intracellular protein in placental trophoblasts. Here we show by 2D electrophoresis and mass spectroscopy that this reactivity is with cytokeratin 18, a cytokeratin present in most simple epithelia. Immunohistochemistry (IHC) with CD133-2/AC141 on a trophoblast cell line displayed a staining pattern typical for the cytoskeleton. Cryostat sections of stratified epithelia lacking cytokeratin 18 did not react with CD133-2/AC141. In conclusion, care must be taken not to misinterpret staining patterns using CD133-2/AC141 in IHC. PMID- 12133916 TI - Fluoro Jade stains early and reactive astroglia in the primate cerebral cortex. AB - The fluorescent agent Fluoro Jade was applied to cortical brain sections obtained from human patients at early postnatal ages and in patients with Alzheimer's disease, and from a Cebus apella monkey after mechanical lesioning of the cerebral cortex. Fluoro Jade labeled reactive astrocytes and early differentiating astroglial cells. PMID- 12133917 TI - Studies in the physical chemistry of the proteins. V. Molecular weights of the proteins. J.Biol.Chem 63, 721-766, 1925. PMID- 12133918 TI - Could HPV testing become the sole primary cervical screening test? PMID- 12133919 TI - Outcome events in studies of diagnostic or screening tests. PMID- 12133920 TI - Faecal occult blood screening for colorectal cancer. PMID- 12133921 TI - Screening for neuroblastoma in children. PMID- 12133922 TI - Impact of antenatal HIV screening to prevent HIV infection in children in Norway 1987-99. AB - OBJECTIVE: To assess the impact of the antenatal HIV screening programme in Norway in preventing HIV infection in children. SETTING: Norway, 1987-99. METHODS: In a simulated retrospective cohort design data were used from the mandatory HIV surveillance system to compare the observed number of children born infected with HIV in Norway 1987-99 to the expected number without the antenatal screening programme. The main measures were relative and absolute performance of the screening programme. Other measures were uptake and false positive rate of screening, and number and exposure category of screen positive women. RESULTS: 96% of 961 000 eligible pregnant women were tested. 0.1% had an indeterminate test result and 46 women (5.0/100 000) were confirmed screen positive. 27 were African or south east Asian women infected before immigration to Norway. Nine out of 739 000 live born children (1.2/100 000) were infected compared with the expected 18 with no screening. The absolute impact of the screening programme was 1.3 (95% confidence interval (95% CI) -0.1 to 2.7) prevented infections in 100 000 women screened. The relative preventive impact was 51% (-15% to 81%). CONCLUSIONS: The limited absolute impact is because of the very low prevalence of undetected HIV infection among pregnant women in Norway. PMID- 12133923 TI - Screening for cystic fibrosis in newborn infants: results of a pilot programme based on a two tier protocol (IRT/DNA/IRT) in the Italian population. AB - OBJECTIVE: To assess the performance of a two tier neonatal screening programme (IRT/DNA/IRT) for cystic fibrosis, based on immunoreactive trypsinogen (IRT) followed by direct cystic fibrosis transmembrane conductance regulator (CFTR) gene analysis (based on a panel of up to 31 mutations) in hypertrypsinaemic newborn infants and to compare it with a previous screening protocol. SETTING: The study comprised all the newborn infants in the period 1 October 1998 to 31 December 1999 in the Lombardia region, north western Italy. METHODS: The screening strategy consisted of an immunoreactive trypsinogen assay from dried blood spots, a polymerase chain reaction (PCR) followed by an oligonucleotide ligation assay (PCR-OLA), and a sequence code separation. RESULTS: 104 609 newborn infants were screened. 1457 hypertrypsinaemic infants (1.39%) were analysed with the PCR-OLA assay. 18 newborn homozygotes or compound heterozygotes for CFTR mutations were identified and referred to the cystic fibrosis (CF) centre at a mean age of 3 weeks. 125 infants presenting only one mutation were recalled for a sweat test: a diagnosis of CF was made in 13 infants, and parents of 112 neonates identified as carriers (1:13) received genetic counselling. The remaining 1314 hypertrypsinaemic newborn infants were recalled for IRT retesting and 177 were referred for a sweat test because the second IRT measurement was above the cut off value. Among this group a further two infants were diagnosed with CF (1.1%) leading to a CF prevalence of 1:3170. CONCLUSIONS: This strategy resulted in an early and accurate diagnosis of CF. The IRT/DNA/IRT protocol with an OLA assay was shown to be useful in an Italian population with a genetic heterogeneity, leading to the identification of 94% of infants with CF. PMID- 12133924 TI - Measurement of phenylalanine in blood on filter paper as a method of monitoring PKU treatment. AB - OBJECTIVES: Phenylketonuria (PKU) is a genetic disease with autosomal recessive inheritance. In Poland the microbiological Guthrie test for this disease was replaced by an enzymatic colorimetric test. The question is whether the colorimetric test might be used in monitoring treatment of PKU. SETTING: In 80 patients with PKU on routine treatment monitoring of serum phenylalanine concentrations (SPh) was compared with phenylalanine concentrations in blood on filter paper (PhBFP). METHODS: Measurements of SPh were by a fluorimetric method (McCaman and Robins), and those of PhBFP were by an enzymatic colorimetric method. RESULTS: The regression analysis of SPh compared with PhBFP gave the equation y=0.9219x+0.2389; for the reversed ratio: it gave y=1.0220x+0.55083. The correlation was 0.97 at p<0.05. DISCUSSION: Concentrations of SPh accepted for children with PKU are not uniform. In the Cracow centre, the range of accepted SPh concentrations are 2-6 mg/dl for children and <12 mg/dl for older patients. The concentrations of PhBFP accepted are somewhat higher than SPh, range from 4 to 10 mg/dl, and are a good indicator of an appropriate diet. CONCLUSIONS: (1) The comparison indicated that colorimetric measurements of PhBFP are effective in monitoring therapeutic PKU management. Tests can be performed more often and are more comfortable both for the patients and their parents. (2) The comparative results indicated a concentration of PhBFP ranging from 4 to 10 mg/dl to be the accepted value for children treated for PKU. PMID- 12133925 TI - Outcome of a school screening programme for carriers of haemoglobin disease. AB - OBJECTIVES: To assess the impact of a screening programme for haemoglobinopathies which was organised from 1978 to 1985 in high secondary schools of the Marseille region. METHODS: Several variables that reflected the influence of this preventive programme on the uptake of prenatal diagnosis were investigated. To evaluate the partner's uptake for the testing, a letter was sent, together with an anonymous questionnaire, to all the haemoglobin carriers detected in this programme. To evaluate the number of prenatal diagnoses, the charts of all couples from the Marseille area who underwent genetic counselling for haemoglobinopathies were compiled. The number of affected children born between 1980 to 2000 was recorded, and the cases in which one of the parents had previously been screened at school were noted. RESULTS: Half of the carriers replied to the questionnaire: 86% knew that they have to test their partner. Six carrier couples were identified, four asked for genetic counselling and requested eight prenatal diagnoses, two couples did not request genetic counselling and have had two affected children. CONCLUSIONS: Despite the time lapse between screening, informing, and pregnancy (mean 15 years), the information was well conserved and resulted in testing of the partner. The screening programme was effective in motivating requests for prenatal diagnosis. PMID- 12133926 TI - Does a better grade of tumour occurring in women under hormone replacement therapy compensate for their lower probability of detection by screening mammography. AB - OBJECTIVE: To compare the prognostic factor of breast cancer survival between breast cancer diagnosed in subjects receiving hormone replacement therapy (HRT) before diagnosis to those without such a therapy. SUBJECTS AND METHODS: All breast cancers diagnosed between 1993 and 2000 within the breast cancer screening programme in Bouches du Rhone (France) were analysed for size, node status, and grade according to use, or not, of HRT. Univariate and multivariate analyses were carried out taking into account age, density of the breast, and mode of detection. RESULTS: The breast tumours diagnosed among HRT users had a lower grade whatever the mode of detection. The proportion of node positive tumours was identical in the two groups after adjustment for age. The smaller size of the tumours among HRT users is partly explained by the lower grade of these tumours Conclusion: Although tumours occurring in HRT users have a lower chance of being detected by screening, their prognostic factors, especially the grade of the tumour, are better than in non-users. More work is needed to find which part of this advantage is attributable to better surveillance of women treated with HRT PMID- 12133928 TI - Crude open biopsy rates for benign screen detected lesions no longer reflect breast screening quality--time to change the standard. AB - OBJECTIVES: To investigate the changing nature of the benign screen detected breast abnormalities removed at open biopsy over a seven year period and to compare this with the improving cancer detection rate and non-operative diagnosis rate. SETTING: The Bolton, Bury, and Rochdale Breast Screening Programme. METHODS: The histopathology reports of the benign lesions removed from patients undergoing open biopsy for screen detected abnormalities between 1 April 1994 and 31 March 2001 were reviewed and the lesions classified on the B1 to B5 scale. Cancer detection rates and non-operative cancer diagnosis rates were ascertained from the breast screening computer system. RESULTS: 148 benign surgical biopsies were performed in the seven year period. There was a moderate increase in the overall benign biopsy rate over the period (from 1.26 open biopsies per 1000 women screened for the three years 1994-97 to 1.63 open biopsies per 1000 women screened for the three years 1998-2001). The biopsy rate for B2 (benign) lesions decreased slightly over the study period but the biopsy rate for B3 lesions (that is, of uncertain malignant potential) more than doubled. The majority (84%) of the B3 lesions were radial scars. There was a steady improvement in the cancer detection rate and the non-operative cancer diagnosis rate over the period, similar to that seen nationally. CONCLUSIONS: Improvements in screening technique and detection ability result in an increase in the number of subtle radiologically indeterminate or suspicious lesions detected. Many of these are radial scars, which require excision. Crude benign open biopsy rates for screening programmes are no longer meaningful, and should now be refined with separate rates for B2 lesions and B3 lesions. High quality programmes can expect to have low B2 open biopsy rates and high B3 open biopsy rates. It is inappropriate to have an upper limit for the B3 open biopsy rate. PMID- 12133927 TI - Predictors of outcome of mammography in the National Health Service Breast Screening Programme. AB - BACKGROUND: Little is known about the factors influencing the risk of recall for assessment, invasive diagnostic procedures, and early rescreening after screening mammography. METHODS: From June 1996 to March 1998 women attending screening at 10 National Health Service Breast Screening Programme (NHSBSP) centres completed a self administered questionnaire and were followed up for their screening outcome. RESULTS: 1969 (3.3%) out of 60 443 women aged 50-64 who had never used hormone replacement therapy (HRT) were recalled for assessment but were not diagnosed with breast cancer (defined here as false positive recall). After adjustment for the variation between centres, false positive recall was decreased significantly among women who were likely to have had a previous NHSBSP mammogram (odds ratio (OR) 0.49, 95% confidence interval (95% CI) 0.38 to 0.63 for likely versus unlikely), who were postmenopausal (OR 0.65, 95% CI 0.56 to 0.76 for postmenopausal v premenopausal) and increased significantly for women reporting previous breast surgery (OR 1.64, 95% CI 1.42 to 1.89). Although false positive recall decreased significantly with parity and increasing body mass index, these effects were not large and no significant variation was found with age, education, family history of breast cancer, oral contraceptive use, sterilisation, exercise, smoking, or alcohol consumption. Altogether 655 (1.1%) women had an invasive diagnostic procedure; no personal characteristics were predictive of this outcome, 286(0.5%) were referred for early rescreening, and this was increased significantly by nulliparity and a family history of breast cancer. INTERPRETATION: Premenopausal women, those without a previous NHSBSP mammogram, and women with previous breast surgery have an increased risk of false positive recall by the NHSBSP. PMID- 12133929 TI - The cervical cancer screening programme in Norway, 1992-2000: changes in Pap smear coverage and incidence of cervical cancer. AB - OBJECTIVES: Changes in the incidence of cervical cancer were studied to assess the impact of the Norwegian coordinated cervical cancer screening programme introduced in 1995. Attention was given as to whether recommendation letters sent to women without a screen in the previous 3 years could be an alternative to a conventional screening programme that invites women irrespectively of their spontaneous screening. SETTING: A population based, nationwide, screening programme in which women of 25 to 69 are recommended to have a conventional Pap smear every 3 years. METHODS: The impact of the screening programme was assessed indirectly by comparing trends in invasive cervical cancer, changes in coverage, and changes in interval between Pap smears in the 3 year period (1992-4) before screening with the two first screening rounds (1995-7 and 1998-2000). All Pap smears taken from women of all ages were included, a total of 4 744 967 Pap smears from more than 1.4 million women. Further, the impact was assessed directly by logistic regression by comparing the screening results of women recruited for the programme with women who had regularly had Pap smears. RESULTS: In the last 2 years studied, the incidence of invasive cancer was 22% lower than in the period before the programme. The proportion of women who had a Pap smear was higher after the implementation of the coordinated screening programme. The number of smears taken was less as the interval after a normal smear was greater. The newly recruited women had a threefold risk of having a high grade precursor and a 20-fold higher risk of cancer than the women who had had regular smears. CONCLUSIONS: The coordinated screening programme provides a low cost way of increasing the coverage of the female population, and consequently has reduced the rate of invasive cervical cancer. PMID- 12133930 TI - Screening and management of asymptomatic popliteal aneurysms. AB - OBJECTIVES: To determine the prevalence of popliteal aneurysms in men to enable the case for screening and elective surgery to be assessed. SETTING: Scott Research Unit, St Richards Hospital, Chichester. METHODS: The popliteal arteries of 1074 men aged between 65 and 80 were scanned with ultrasound, aneurysmal vessels >1.5 cm diameter were rescanned 5 years later to assess their rate of expansion. RESULTS: 11 of 1074 patients screened had a popliteal aneurysm between 15 and 26 mm, a prevalence of 1.0%. Five years later no increase in aneurysm size had occurred and no related complications were reported. CONCLUSION: In men the low prevalence and complication rate of popliteal aneurysms in conjunction with the effective treatment of acutely thrombosed aneurysms provides evidence for conservative management but against screening asymptomatic popliteal aneurysms. PMID- 12133933 TI - Cutting edge: molecular mechanisms of synergy between CD40 and the B cell antigen receptor: role for TNF receptor-associated factor 2 in receptor interaction. AB - Optimal Ag-specific B lymphocyte activation requires both recognition of Ag by the B cell Ag receptor (BCR) and contact-mediated interactions with Ag-specific Th lymphocytes. One of these interactions involves ligation of B cell CD40 by T cell-expressed CD154. CD40 signaling is crucial for Ab production, isotype switching, up-regulation of surface molecules, development of germinal centers, and the humoral memory response. The signaling pathways emanating from the BCR and CD40 are able to cooperate, but the molecular mechanisms responsible for this interaction are incompletely understood. The present study explored the roles of signaling motifs in the CD40 cytoplasmic tail in this synergy. We find that threonine in the PXQXT motif in the TNFR-associated factor-2 binding site is critical for synergistic effects of CD40 and BCR signals, independent of its phosphorylation. Furthermore, data suggest an indirect role for TNFR-associated factor-2 in the cooperative signaling. PMID- 12133934 TI - Essential role of NF-kappa B-inducing kinase in T cell activation through the TCR/CD3 pathway. AB - NF-kappa B-inducing kinase (NIK) is involved in lymphoid organogenesis in mice through lymphotoxin-beta receptor signaling. To clarify the roles of NIK in T cell activation through TCR/CD3 and costimulation pathways, we have studied the function of T cells from aly mice, a strain with mutant NIK. NIK mutant T cells showed impaired proliferation and IL-2 production in response to anti-CD3 stimulation, and these effects were caused by impaired NF-kappa B activity in both mature and immature T cells; the impaired NF-kappa B activity in mature T cells was also associated with the failure of maintenance of activated NF-kappa B. In contrast, responses to costimulatory signals were largely retained in aly mice, suggesting that NIK is not uniquely coupled to the costimulatory pathways. When NIK mutant T cells were stimulated in the presence of a protein kinase C (PKC) inhibitor, proliferative responses were abrogated more severely than in control mice, suggesting that both NIK and PKC control T cell activation in a cooperative manner. We also demonstrated that NIK and PKC are involved in distinct NF-kappa B activation pathways downstream of TCR/CD3. These results suggest critical roles for NIK in setting the threshold for T cell activation, and partly account for the immunodeficiency in aly mice. PMID- 12133935 TI - Generation of CD4(+)CD45RA(+) effector T cells by stimulation in the presence of cyclic adenosine 5'-monophosphate-elevating agents. AB - After TCR cross-linking, naive CD4(+)CD45RA(+) T cells switch to the expression of the CD45RO isoform and acquire effector functions. In this study we have shown that cAMP-elevating agents added to anti-CD3- and anti-CD28-stimulated cultures of T lymphocytes prevent acquisition of the CD45RO(+) phenotype and lead to the generation of a new subpopulation of primed CD4(+)CD45RA(+) effector cells (cAMP primed CD45RA). These cells displayed a low apoptotic index, as the presence of dibutyryl cAMP (dbcAMP)-rescued cells from CD3/CD28 induced apoptosis. Inhibition of CD45 splicing by dbcAMP was not reverted by addition of exogenous IL-2. cAMP primed CD45RA cells had a phenotype characteristic of memory/effector T lymphocytes, as they showed an up-regulated expression of CD2, CD44, and CD11a molecules, while the levels of CD62L Ag were down-regulated. These cells also expressed the activation markers CD30, CD71, and HLA class II Ags at an even higher level than CD3/CD28-stimulated cells in the absence of dbcAMP. In agreement with this finding, cAMP-primed CD45RA cells were very efficient in triggering allogenic responses in a MLR. In addition, cAMP-primed CD45RA cells produce considerable amounts of the Th2 cytokines, IL-4, IL-10, and IL-13, whereas the production of IFN-gamma and TNF-alpha was nearly undetectable. The elevated production of IL-13 by neonatal and adult cAMP-primed CD45RA cells was specially noticeable. The cAMP-dependent inhibition of CD45 splicing was not caused by the production of immunosuppressor cytokines. These results suggest that within the pool of CD4(+)CD45RA(+) cells there is a subpopulation of effector lymphocytes generated by activation in the presence of cAMP-elevating agents. PMID- 12133936 TI - Modulation of tissue-specific immune response to cardiac myosin can prolong survival of allogeneic heart transplants. AB - The role of immune response to tissue-specific Ags in transplant rejection is poorly defined. We have previously reported that transplantation of cardiac allografts triggers a CD4(+) Th1 cell response to cardiac myosin (CM), a major contractile protein of the heart, and that pretransplant activation of proinflammatory CM-specific T cells accelerates rejection. In this study, we show that administration of CM together with IFA (CM/IFA) can prevent acute rejection of an allogeneic heart transplant. Prolongation of cardiac graft survival is associated with activation of CM- and allo-specific T cells secreting type 2 cytokines (IL-4, IL-5) and reduction of the frequency of proinflammatory IFN gamma-secreting (type 1) alloreactive T cells. Blocking of IL-4 cytokine with Abs abrogates the prolongation. CM/IFA treatment prevents acute rejection of MHC class I-mismatched, but not fully mismatched grafts. However, if donor heart is devoid of MHC class II expression, CM-IFA administration delays rejection of fully allogeneic cardiac transplants. This finding suggests that the effect of CM modulation depends on the type (direct vs indirect) and strength of recipient's CD4(+) T cell alloresponse. Our results underscore the important role of host immunity to tissue-specific Ags in the rejection of an allograft. This study demonstrates that modulation of the immune response to a tissue-specific Ag can significantly prolong cardiac allograft survival, an observation that may have important implications for the development of novel selective immune therapies in transplantation. PMID- 12133937 TI - Requirement for the lymphocyte semaphorin, CD100, in the induction of antigen specific T cells and the maturation of dendritic cells. AB - CD100 belongs to the semaphorin family, several members of which are known to act as repulsive axonal guidance factors during neuronal development. We have previously demonstrated that CD100 plays a crucial role in humoral immunity. In this study, we show that CD100 is also important for cellular immunity through the maturation of dendritic cells (DCs). CD100(-/-) mice fail to develop experimental autoimmune encephalomyelitis induced by myelin oligodendrocyte glycoprotein peptide, because myelin oligodendrocyte glycoprotein-specific T cells are not generated in the absence of CD100. In vitro studies with T cells from OVA-specific TCR-transgenic mice demonstrate that Ag-specific T cells lacking CD100 fail to differentiate into cells producing either IL-4 or IFN-gamma in the presence of APCs and OVA peptide. In addition, DCs from CD100(-/-) mice display poor allostimulatory capabilities and defects in costimulatory molecule expression and IL-12 production. The addition of exogenous soluble rCD100 restores normal functions in CD100(-/-) DCs and further enhances functions of normal DCs. Furthermore, treatment of Ag-pulsed DCs with both soluble CD100 and anti-CD40 before immunization significantly enhances their immunogenicity. This treatment elicits improved T cell priming in vivo, enhancing both primary and memory T cell responses. Collectively, these results demonstrate that CD100, which enhances the maturation of DCs, is essential in the activation and differentiation of Ag-specific T cells. PMID- 12133938 TI - CD40 ligand and CTLA-4 are reciprocally regulated in the Th1 cell proliferative response sustained by CD8(+) dendritic cells. AB - Subsets of murine dendritic cells (DCs) from the spleen differ in their ability to induce proliferative responses in both primary and secondary CD4(+) T cells. Recent evidence indicates that lymphoid-related CD8(+) DCs fail to provide appropriate signals to freshly isolated secondary CD4(+) T cells to sustain their proliferation in vitro. In the present study, we examined peptide-pulsed CD8(-) and CD8(+) DCs for ability to stimulate Th1 and Th2 cell clones with the same Ag specificity. Defective ability to induce proliferation was selectively shown by CD8(+) DCs presenting Ag to the Th1 clone. The deficiency in CD8(+) DCs was overcome by CD40 triggering before peptide pulsing. When exposed to CD8(+) DCs in the absence of CD40 activation, the Th1 clone expressed low levels of CD40 ligand and high levels of surface CTLA-4. Neutralization of CTLA-4 during the DC/T cell coculture resulted in increased CD40 ligand expression and proliferation of T cells. Remarkably, the activation of CD40 on DCs under conditions that would increase Th1 cell proliferation, also resulted in down-regulation of surface CTLA 4. These results confirm differential effects of CD8(+) and CD8(-) DCs in the stimulation of Ag-primed Th cells. In addition, they suggest that reciprocal regulation of CD40 ligand and CTLA-4 expression occurs in Th1 cells exposed to CD8(+) DCs. PMID- 12133939 TI - Immune surveillance and effector functions of CCR10(+) skin homing T cells. AB - Skin homing T cells carry memory for cutaneous Ags and play an important sentinel and effector role in host defense against pathogens that enter via the skin. CCR10 is a chemokine receptor that is preferentially expressed among blood leukocytes by a subset of memory CD4 and CD8 T cells that coexpress the skin homing receptor cutaneous lymphocyte Ag (CLA), but not the gut-homing receptor alpha(4)beta(7). Homing and chemokine receptor coexpression studies detailed in this study suggest that the CLA(+)/CCR10(+) memory CD4 T cell population contains members that have access to both secondary lymphoid organ and skin compartments; and therefore, can act as both "central" and "effector" memory T cells. Consistent with this effector phenotype, CLA(+)/CCR10(+) memory CD4 T cells from normal donors secrete TNF and IFN-gamma but minimal IL-4 and IL-10 following in vitro stimulation. Interactions of CCR10 and its skin-associated ligand CC ligand 27 may play an important role in facilitating memory T cell entry into cutaneous sites during times of inflammation. PMID- 12133940 TI - Cross-reactive antigen is required to prevent erosion of established T cell memory and tumor immunity: a heterologous bacterial model of attrition. AB - Induction and maintenance of T cell memory is critical for the control of intracellular pathogens and tumors. Memory T cells seem to require few "maintenance signals," though often such studies are done in the absence of competing immune challenges. Conversely, although attrition of CD8(+) T cell memory has been characterized in heterologous viral models, this is not the case for bacterial infections. In this study, we demonstrate attrition of T cell responses to the intracellular pathogen Listeria monocytogenes (LM) following an immune challenge with a second intracellular bacterium, Mycobacterium bovis (bacillus Calmette-Guerin, BCG). Mice immunized with either LM or recombinant LM (expressing OVA; LM-OVA), develop a potent T cell memory response. This is reflected by peptide-specific CTL, IFN-gamma production, and frequency of IFN gamma-secreting T cells to native or recombinant LM Ags. However, when the LM infected mice are subsequently challenged with BCG, there is a marked reduction in the LM-specific T cell responses. These reductions are directly attributable to the effects on CD4(+) and CD8(+) T cells and the data are consistent with a loss of LM-specific T cells, not anergy. Attrition of the Ag (OVA)-specific T cell response is prevented when LM-OVA-immunized mice are challenged with a subsequent heterologous pathogen (BCG) expressing OVA, demonstrating memory T cell dependence on Ag. Although the reduction of the LM-specific T cell response did not impair protection against a subsequent LM rechallenge, for the first time, we show that T cell attrition can result in the reduction of Ag-specific antitumor (B16-OVA) immunity previously established with LM-OVA immunization. PMID- 12133941 TI - Direct ex vivo analysis of human CD4(+) memory T cell activation requirements at the single clonotype level. AB - CD4(+) memory T cells continuously integrate signals transmitted through the TCR and costimulatory molecules, only responding when the intensity of such signals exceeds an intrinsic activation threshold. Recent data suggest that these activation thresholds can be regulated independently of TCR specificity, and that threshold tuning may constitute a major mechanism for controlling T cell effector activity. In this work we take advantage of the profound clonotypic hierarchies of the large human CD4(+) T cell response to CMV to study activation thresholds of fresh (unexpanded) memory T cells at the clonotypic level. We identified dominant responses to CMV matrix determinants mediated by single TCRB sequences within particular TCR-Vbeta families. The specific response characteristics of these single, Ag-specific, TCRB-defined clonotypes could be unequivocally determined in fresh PBMC preparations by cytokine flow cytometry with gating on the appropriate Vbeta family. These analyses revealed 1) optimal peptides capable of eliciting specific responses by themselves at doses as low as 2 pg/ml, with each log increase in dose eliciting ever-increasing frequencies of responding cells over a 4- to 5-log range; 2) significant augmentation of response frequencies at all submaximal peptide doses by CD28- and CD49d-mediated costimulation; 3) differential dose response and costimulatory characteristics for IFN-gamma and IL-2 responses; and 4) no association of activation requirements with the CD27-defined CD4(+) T cell memory differentiation pathway. Taken together these data confirm that triggering heterogeneity exists within individual CD4(+) memory T cell clonotypes in vivo and demonstrate that such single clonotypes can manifest qualitatively different functional responses depending on epitope dose and relative levels of costimulation. PMID- 12133942 TI - Suppressor of cytokine signaling-3 is recruited to the activated granulocyte colony stimulating factor receptor and modulates its signal transduction. AB - G-CSF is a polypeptide growth factor used in treatment following chemotherapy. G CSF regulates granulopoiesis and acts on its target cells by inducing homodimerization of the G-CSFR, thereby activating intracellular signaling cascades. The G-CSFR encompasses four tyrosine motifs on its cytoplasmic tail that have been shown to recruit a number of regulatory proteins. Suppressor of cytokine signaling 3 (SOCS-3), also referred to as cytokine-inducible Src homolgy 2-containing protein 3, is a member of a recently discovered family of feedback inhibitors that have been shown to inhibit the Janus kinase/STAT pathway. In this study, we demonstrate that human SOCS-3 is rapidly induced by G-CSF in polymorphonuclear neutrophils as well as in the myeloid precursor cell line U937 and that SOCS-3 negatively regulates G-CSFR-mediated STAT activation. Most importantly, we show that SOCS-3 is recruited to the G-CSFR in a phosphorylation dependent manner and we identify phosphotyrosine (pY)729 as the major recruitment site for SOCS-3. Furthermore, we demonstrate that SOCS-3 directly binds to this pY motif. Surface plasmon resonance analysis reveals a dissociation constant (K(D)) for this interaction of around 2.8 microM. These findings strongly suggest that the recruitment of SOCS-3 to pY729 is important for the modulation of G-CSFR mediated signal transduction by SOCS-3. PMID- 12133943 TI - Dendritic cell immunogenicity is regulated by peroxisome proliferator-activated receptor gamma. AB - Dendritic cells (DC) are the most potent APCs known that play a key role for the initiation of immune responses. Ag presentation to T lymphocytes is likely a constitutive function of DC that continues during the steady state. This raises the question of which mechanism(s) determines whether the final outcome of Ag presentation will be induction of immunity or of tolerance. In this regard, the mechanisms controlling DC immunogenicity still remain largely uncharacterized. In this paper we report that the nuclear receptor peroxisome proliferator-activated receptor gamma (PPAR-gamma), which has anti-inflammatory properties, redirects DC toward a less stimulatory mode. We show that activation of PPAR-gamma during DC differentiation profoundly affects the expression of costimulatory molecules and of the DC hallmarker CD1a. PPAR-gamma activation in DC resulted in a reduced capacity to activate lymphocyte proliferation and to prime Ag-specific CTL responses. This effect might depend on the decreased expression of costimulatory molecules and on the impaired cytokine secretion, but not on increased IL-10 production, because this was reduced by PPAR-gamma activators. Moreover, activation of PPAR-gamma in DC inhibited the expression of EBI1 ligand chemokine and CCR7, both playing a pivotal role for DC migration to the lymph nodes. These effects were accompanied by down-regulation of LPS-induced nuclear localized RelB protein, which was shown to be important for DC differentiation and function. Our results suggest a novel regulatory pathway for DC function that could contribute to the regulated balance between immunity induction and self-tolerance maintenance. PMID- 12133944 TI - T cell antigen receptor engagement and specificity in the recognition of stress inducible MHC class I-related chains by human epithelial gamma delta T cells. AB - Human gamma delta T cells with the TCR variable region V(delta)1 occur mainly in epithelia and respond to stress-induced expression of the MHC class I-related chains A and B, which have no function in Ag presentation. MIC function as ligands for NKG2D-DAP10, an activating receptor complex that triggers NK cells, costimulates CD8 alpha beta and V(gamma)9V(delta)2 gamma delta T cells, and is required for stimulation of V(delta)1 gamma delta T cells. It is unresolved, however, whether triggering of V(delta)1 gamma delta TCRs is also mediated by MIC or by unidentified cell surface components. Soluble MICA tetramers were used as a binding reagent to demonstrate specific interactions with various V(delta)1 gamma delta TCRs expressed on transfectants of a T cell line selected for lack of NKG2D. Tetramer binding was restricted to TCRs derived from responder T cell clones classified as reactive against a broad range of MIC-expressing target cells and was abrogated when TCRs were composed of mismatched gamma- and delta chains. These results and the inability of V(delta)1 gamma delta T cells to respond to target cells expressing the ULBP/N2DL ligands of NKG2D, which are highly divergent from MIC, indicate that MIC delivers both the TCR-dependent signal 1 and the NKG2D-dependent costimulatory signal 2. This dual function may serve to prevent erroneous gamma delta T cell activation by cross-reactive cell surface determinants. PMID- 12133945 TI - T cell proliferation induced by autologous non-T cells is a response to apoptotic cells processed by dendritic cells. AB - Self-reactive T cells are present in the mature immune repertoire as demonstrated by T cell proliferation induced by autologous non-T cells in the autologous mixed lymphocyte reaction. This reaction generates regulatory T cells in vitro and may reflect immune regulatory pathways in vivo, but the antigenic peptides recognized remain uncharacterized. We revisited this issue in light of the importance of apoptosis in immune regulation. We found that apoptosis among peripheral blood non-T stimulator cells is associated with augmented induction of autologous T cell proliferation. Our data show that caspase activity in the non-T stimulator population is essential for induction of autologous T cell proliferation, suggesting that cellular components in the non-T cell fraction are enzymatically modified, most likely by effector caspases, and have a direct or indirect effect on autoreactive T cell activation. Furthermore, exposure of macrophage-derived dendritic cells to apoptotic non-T cells augments autologous T cell proliferation, and blockade of alpha(v)beta(5) integrin, but not alpha(v)beta(3), inhibits the capacity of irradiated non-T cells or dendritic cells to stimulate autologous T cell proliferation. These experiments, using an entirely autologous system, suggest the interpretation that autoreactive T cells may recognize self Ags modified through the actions of caspases and presented to T cells by dendritic cells. Induction of an in vivo autologous mixed lymphocyte reaction by caspase-modified self-Ags present in apoptotic cells may represent a mechanism to maintain peripheral immune tolerance. PMID- 12133946 TI - Priming biologically active antibody responses against an isolated, conformational viral epitope by DNA vaccination. AB - The immunodominant, conformational "a" determinant of hepatitis B surface Ag (HBsAg) elicits Ab responses. We selectively expressed the Ab-binding, glycosylated, native a determinant (residue 120-147) of HBsAg in a fusion protein containing C-terminally the HBsAg fragment SII (residue 80-180) fused to a SV40 T Ag-derived hsp73-binding 77 aa (T(77)) or non-hsp-binding 60 aa (T(60)) N terminus. A DNA vaccine encoding non-hsp-binding secreted T(60)-SII fusion protein-stimulated murine Ab responses with a similar efficacy as a DNA vaccine encoding the secreted, native, small HBsAg. A DNA vaccine encoding hsp73-binding, intracellular T(77)-SII fusion protein-stimulated murine Ab responses less efficiently but comparable to a DNA vaccine encoding the intracellular, native, large HBsAg. HBsAg-specific Abs elicited by either the T(60)-SII-expressing or the T(77)-SII-expressing DNA vaccine suppressed HBsAg antigenemia in transgenic mice that produce HBsAg from a transgene in the liver; hence, a biologically active B cell response cross-reacting with the native, viral envelope epitope was primed by both DNA vaccine constructs. HBsAg-specific Ab and CTL responses were coprimed when an S(20-50) fragment (containing the immunodominant, L(d)-binding epitope S(28-39)) of HBsAg was fused C-terminally to the pCI/T(77)-SII sequence (pCI/T(77)-SII-L(d) DNA vaccine). Chimeric, polyepitope DNA vaccines encoding conformational, Ab-binding epitopes and MHC class I-binding epitopes can thus efficiently deliver antigenic information to different compartments of the immune system in an immunogenic way. PMID- 12133947 TI - IFN consensus sequence binding protein/IFN regulatory factor-8 guides bone marrow progenitor cells toward the macrophage lineage. AB - IFN consensus sequence binding protein (ICSBP; IFN regulatory factor-8) is a transcription factor of the IFN regulatory factor family. Disruption of this gene results in a leukemia-like disease in mice. To investigate the role of ICSBP in myeloid cell development, lineage marker-negative (Lin(-)) bone marrow progenitor cells were purified from ICSBP(+/+) and ICSBP(-/-) mice and tested for gene expression and colony-forming ability. ICSBP was expressed in Lin(-) progenitor cells, and its levels were markedly increased by IFN-gamma. The colony-forming potential of ICSBP(-/-) progenitor cells was grossly abnormal, as they gave rise to a disproportionately high number of granulocyte colonies and many fewer macrophage colonies. IFN-gamma inhibited colony formation, while promoting macrophage maturation in ICSBP(+/+) cells. In contrast, the effects of IFN-gamma were completely absent in ICSBP(-/-) progenitors. By retrovirus transduction we tested whether reintroduction of ICSBP restores a normal colony-forming potential in -/- progenitor cells. The wild-type ICSBP, but not transcriptionally defective mutants, corrected abnormal colony formation by increasing macrophage colonies and decreasing granulocyte colonies. Taken together, ICSBP plays a critical role in myeloid cell development by controlling lineage selection and is indispensable for IFN-gamma-dependent modulation of progenitor cell maturation. PMID- 12133948 TI - The CD154-CD40 T cell costimulation pathway is required for host sensitization of CD8(+) T cells by skin grafts via direct antigen presentation. AB - Although the CD154-CD40 T cell costimulation pathway has been shown to mediate alloimmune responses in normal recipients, little is known about its role in sensitized hosts. In this work, by using novel models of cardiac allograft rejection in skin-sensitized CD154- and CD40-deficient mice, we reaffirm the key role of CD154-CD40 signaling in host sensitization to alloantigen in vivo. First, we identified CD8(+) T cells as principal effectors in executing accelerated rejection in our model. Disruption of CD154-CD40 signaling in recipients at the T cell side (CD154-deficient) but not at the APC side (CD40-deficient) abrogated accelerated (<2 days) rejection and resulted in long-term (>100 days) graft survival. This suggests that the CD154-dependent mechanism in host CD8(+) T cell sensitization operates via the direct Ag presentation. Then, in comparative studies of alloimmune responses in CD154-deficient and wild-type recipients, we showed that, although alloreactive B cell responses were inhibited, alloreactive T cell responses were down-regulated selectively in the CD8(+) T cell compartment, leaving CD4(+) T cells largely unaffected. This unique alteration in host alloreactivity, seen not only in peripheral lymphocytes but also in allograft infiltrate, may represent the key mechanism by which disruption of CD154-CD40 signaling prevents sensitization to alloantigen in vivo and leads to long-term allograft survival. PMID- 12133949 TI - Chemotactic responsiveness toward ligands for CXCR3 and CXCR4 is regulated on plasma blasts during the time course of a memory immune response. AB - Plasma blasts formed during memory immune responses emigrate from the spleen to migrate into the bone marrow and into chronically inflamed tissues where they differentiate into long-lived plasma cells. In this study, we analyze the chemokine responsiveness of plasma blasts formed after secondary immunization with OVA. Starting from day 4 and within approximately 48 h, OVA-specific plasma blasts emigrate from spleen and appear in the bone marrow. Although these migratory cells have lost their responsiveness to many B cell attracting chemokines, e.g., CXC chemokine ligand (CXCL)13 (B lymphocyte chemoattractant), they migrate toward CXCL12 (stromal cell-derived factor 1 alpha), and toward the inflammatory chemokines CXCL9 (monokine induced by IFN-gamma), CXCL10 (IFN-gamma inducible protein 10), and CXCL11 (IFN-inducible T cell alpha chemoattractant). However, the responsiveness of plasma blasts to these chemokines is restricted to a few days after their emigration from the spleen, indicating a role for these molecules and their cognate receptors, i.e., CXCR3 and CXCR4, in the regulation of plasma blast migration into the bone marrow and/or inflamed tissues. PMID- 12133950 TI - Level of B cell antigen receptor surface expression influences both positive and negative selection of B cells during primary development. AB - To examine the effect of B cell Ag receptor (BCR) surface density on B cell development, we studied multiple lines of mice containing various copy numbers of an IgH micro delta transgene. The V(H) gene in this transgene encodes multireactive BCRs with low affinity for self Ags. These BCRs promote differentiation to a B cell subpopulation that shares some, but not all of the properties of marginal zone (MZ) B cells. Surface BCR level was found to be related to transgene gene copy number in these mice. In mice containing 1-15 copies of the transgene, elevated surface BCR levels were correlated with increased numbers of B cells in the MZ-like subset. However, in mice containing 20-30 copies of the transgene, massive clonal deletion of B cells was observed in the bone marrow, few B cells populated the spleen, and B cells were essentially absent from the lymph nodes. These data support the idea that autoantigens mediate not only negative, but positive selection of developing B cells as well. More importantly, they illustrate the profound influence of BCR surface density on the extent to which either of these selective processes take place. PMID- 12133951 TI - Infection of APC by human cytomegalovirus controlled through recognition of endogenous nuclear immediate early protein 1 by specific CD4(+) T lymphocytes. AB - Infections by human CMV are controlled by cellular immune responses. Professional APC such as monocytes and macrophages can be infected in vivo and are considered as a reservoir of virus. However, CMV-specific CD4(+) responses against infected APC have not been reported. To develop a model of CD4-infected APC interaction, we have transfected the U373MG astrocytoma cell line with the class II transactivator (CIITA). Confocal microscopy experiments showed that U373MG-CIITA cells expressed markers characteristic of APC. Functional assays demonstrated that infected U373MG-CIITA APC processed and presented both exogenous and endogenously neosynthesized nuclear immediate early (IE) protein 1 through the MHC class II pathway. More importantly, endogenous presentation of IE1 by infected APC lead to efficient control of CMV infection as revealed by decreased viral titer. Thus, these results describe the endogenous presentation of a nuclear viral protein by the MHC class II pathway and suggest that IE1-specific CD4(+) T cells may play an important role in CMV infection by directly acting against infected APC. PMID- 12133952 TI - Two distinct domains within the N-terminal region of Janus kinase 1 interact with cytokine receptors. AB - The interaction between receptors and kinases of the Janus kinase (Jak) family is critical for signaling by growth factors, cytokines, and IFNs. Therefore, the characterization of the domains involved in these interactions is pivotal not only in understanding kinase activation but also in the development of drugs that mimic or inhibit signaling. In this report, we have characterized the domains of Jak1 required to associate with distinct cytokine receptor subunits: IFN-alpha R beta L, IFN-gamma R alpha, IL-10R alpha, IL-2R beta, and IL-4R alpha. We demonstrate that two regions of Jak1 are necessary for the interaction with cytokine receptors. First, a common N-terminal region that includes Jak homology (JH) domain 7 and the first 19 aa of JH6, and, second, a C-terminal region (JH6 3) that was different for distinct receptors. The contribution of the two different regions of Jak1 to cytokine receptor binding was also variable. Deletion of JH7-6 impaired the association of IL-2R beta and IL-4R alpha chains with Jak1 but did not have a major impact on the binding of Jak1 to IFN-alpha R beta L or IL-10R alpha. Interestingly, regardless of the effect on receptor binding, removal of JH7-6 completely abrogated kinase activation, indicating that this domain is required for ligand-driven kinase activation and, thus, for proper signaling through cytokine receptors. PMID- 12133954 TI - A role for NF-kappa B activation in perforin expression of NK cells upon IL-2 receptor signaling. AB - Optimal NK cell development and activation as well as cytolytic activity involves IL-2R beta signals that also up-regulate expression of the pore-forming effector molecule perforin. Although the Jak/Stat pathway and specifically Stat5 transcription factors are required to promote many of the respective downstream events, the role of additional signaling pathways and transcription factors remains to be clarified. This report investigates the role of NF-kappa B activation for perforin expression by NK cells. It is demonstrated that IL-2 induced up-regulation of perforin in primary NK cells and in a model cell line is blocked by two pharmacological agents known to inhibit NF-kappa B activation. Direct evidence for the activation of the NF-kappa B pathway by IL-2R signals in NK cells involves activation of the IKK alpha kinase, inhibitory protein kappa B alpha degradation, nuclear translocation of p50/p65 complexes, and ultimately, transcriptional activation of the perforin gene via an NF-kappa B binding element in its upstream enhancer. Taken together, these observations strongly suggest that IL-2R signals can activate a pathway leading to NF-kappa B activation in NK cells and that this pathway is involved in the control of perforin expression. PMID- 12133953 TI - Green fluorescent protein-glucocorticoid receptor knockin mice reveal dynamic receptor modulation during thymocyte development. AB - To delineate the cellular targets and mechanisms by which glucocorticoids (GCs) exert their actions, we generated mice in which a green fluorescent protein (GFP) GC receptor (GR) fusion gene is knocked into the GR locus. In these mice, the GFP GR protein, which is functionally indistinguishable from endogenous GR, allows the tracking and quantitation of GR expression in single living cells. In GFP-GR thymus, GR expression is uniform among embryonic thymocyte subpopulations but gradually matures over a 3-wk period after birth. In the adult, GR is specifically induced to high levels in CD25(+)CD4(-)CD8(-) thymocytes and returns to basal levels in CD4(+)CD8(+) thymocytes of wild-type and positively selecting female HY TCR-transgenic mice, but not negatively selecting male HY TCR transgenic mice. In GFP-GR/recombinase-activating gene 2(-/-) thymocytes, GR expression is down-regulated by pre-TCR complex stimulation. Additionally, relative GR expression is dissociated from GC-induced apoptosis in vivo. Results from these studies define differential GR expression throughout ontogeny, suggest pre-TCR activation as a specific mechanism of GR down-regulation, define immature CD8(+) thymocytes as novel apoptosis-sensitive GC targets, and separate receptor abundance from susceptibility to apoptosis across thymocyte populations. PMID- 12133955 TI - Regulation of class II MHC expression in APCs: roles of types I, III, and IV class II transactivator. AB - Class II transactivator (CIITA) is necessary for expression of class II MHC (MHC II) molecules. In mice, CIITA expression is regulated by three promoters (pI, pIII, and pIV), producing types I, III, and IV CIITA. The relative roles of different CIITA types remain unclear. Unstimulated bone marrow-derived macrophages expressed low levels of CIITA mRNA; type I CIITA was nine times more abundant than type IV (type III CIITA was barely detected). Exposure to IFN-gamma (6 h) dramatically increased types I and IV CIITA mRNA to similar absolute levels. Type IV CIITA declined over time, but type I was stable for over 72 h. Thus, the dominant form of CIITA evolved with time during activation by IFN gamma, and type I CIITA explained prolonged expression of MHC-II by macrophages. mRNA half-life was shorter for type I than type IV CIITA, suggesting that sustained transcription contributed to stable expression of type I CIITA induced by IFN-gamma. Splenic B cells expressed mRNA for type III CIITA but very little for types I or IV. Treatment with IL-4 increased surface expression of MHC-II protein, but mRNA for MHC-II and CIITA (total, I, III, and IV) remained unchanged, suggesting posttranslational regulation. Splenic dendritic cells expressed type I CIITA but little type III or IV; CpG DNA induced their maturation and decreased types I and III CIITA, consistent with decreased MHC-II protein synthesis. CIITA types differ in regulation in various APCs under different stimuli, and the predominant type of CIITA varies at different stages of APC activation. PMID- 12133956 TI - C1q regulatory region polymorphism down-regulating murine c1q protein levels with linkage to lupus nephritis. AB - Much of the pathology of systemic lupus erythematosus (SLE) is caused by deposition of immune complexes (ICs) into various tissues, including renal glomeruli. Because clearance of ICs depends largely on early complement component C1q, homozygous C1q deficiency is a strong genetic risk factor in SLE, although it is rare in SLE patients overall. In this work we addressed the issue of whether genetic polymorphisms affecting C1q levels may predispose to SLE, using the (NZB x NZW)F(1) model. C1q genes are composed of three genes, C1qa, C1qc, and C1qb, arranged in this order, and each gene consists of two exons separated by one intron. Sequence analysis of the C1q gene in New Zealand Black (NZB), New Zealand White (NZW), and BALB/c mice showed no polymorphisms in exons and introns of three genes. However, Southern blot analysis revealed unique insertion polymorphism of a total of approximately 3.5 kb in the C1qa upstream region of NZB mice. C1q levels in sera and culture supernatants of LPS-stimulated peritoneal macrophages and C1q messages in spleen cells were all lower in disease free young NZB and (NZB x NZW)F(1) mice than in age-matched non-autoimmune NZW and BALB/c mice. Quantitative trait loci analysis using (NZB x NZW)F(1) x NZW backcrosses showed that NZB microsatellites in the vicinity of the C1q allele on chromosome 4 were significantly linked to low serum C1q levels and the development of nephritis. These data imply that not only C1q deficiency but also regulatory region polymorphisms down-regulating C1q levels may confer the risk for lupus nephritis by reducing IC clearance and thus promoting IC deposition in glomeruli. PMID- 12133957 TI - The V alpha 14 NKT cell TCR exhibits high-affinity binding to a glycolipid/CD1d complex. AB - Most CD1d-dependent NKT cells in mice have a canonical V alpha 14J alpha 18 TCR rearrangement. However, relatively little is known concerning the molecular basis for their reactivity to glycolipid Ags presented by CD1d. Using glycolipid Ags, soluble forms of a V alpha 14 NKT cell-derived TCR, and mutant and wild-type CD1d molecules, we probed the TCR/CD1d interaction by surface plasmon resonance, tetramer equilibrium staining, and tetramer staining decay experiments. By these methods, several CD1d alpha-helical amino acids could be defined that do not greatly alter lipid binding, but that affect the interaction with the TCR. Binding of the V alpha 14(+) TCR to CD1d requires the agonist alpha galactosylceramide (alpha-GalCer), as opposed to the nonantigenic beta galactosylceramide, although both Ags bind to CD1d, indicating that the carbohydrate moiety of the CD1d-bound Ag plays a major role in the TCR interaction. The TCR has a relatively high-affinity binding to the alpha GalCer/CD1d complex, with a particularly slow off rate. These unique properties are consistent with the coreceptor-independent action of the V alpha 14 TCR and may be related to the intense response to alpha-GalCer by NKT cells in vivo. PMID- 12133959 TI - Isolation and detection of human IgA using a streptococcal IgA-binding peptide. AB - Bacterial proteins that bind to the Fc part of IgG have found widespread use in immunology. A similar protein suitable for the isolation and detection of human IgA has not been described. Here, we show that a 50-residue synthetic peptide, designated streptococcal IgA-binding peptide (Sap) and derived from a streptococcal M protein, can be used for single-step affinity purification of human IgA. High affinity binding of IgA required the presence in Sap of a C terminal cysteine residue, not present in the intact M protein. Passage of human serum through a Sap column caused depletion of >99% of the IgA, and elution of the column allowed quantitative recovery of highly purified IgA, for which the proportions of the IgA1 and IgA2 subclasses were the same as in whole serum. Moreover, immobilized Sap could be used for single-step purification of secretory IgA of both subclasses from human saliva, with a recovery of approximately 45%. The Sap peptide could also be used to specifically detect IgA bound to Ag. Together, these data indicate that Sap is a versatile Fc-binding reagent that may open new possibilities for the characterization of human IgA. PMID- 12133958 TI - Diversification of Ig heavy chain genes in human preterm neonates prematurely exposed to environmental antigens. AB - Preterm neonates are exposed to extrauterine environmental Ags during the time period that corresponds to the last trimester of normal intrauterine development. To study whether this precocious exposure to Ags accelerates the Ig repertoire diversification, we compared IgH chain genes of preterm neonates (gestational age, 25-29 wk) during their first postnatal months with those of term neonates. Preterm infants approaching their expected date of delivery after 8-13 wk of extrauterine life used a similar V(H), D(H), and J(H) gene segment repertoire as term neonates born after intrauterine development. Furthermore, the length increase of the NDN region between V(H) and J(H) by 0.25 nt per gestational week (r = 0.556, p < 0.0001) was not accelerated. Thus, the generation of the V(H) region gene repertoire is developmentally controlled and independent of environmental influences. However, exposure to extrauterine Ags induced class switch and somatic mutations in IgH chain genes within 2 wk after premature birth and IgG transcript diversity and mutational frequency increased with the duration of extrauterine life. Three-month-old preterm infants expressed a heterogeneous IgG repertoire at their expected date of delivery with V(H) region genes carrying significant numbers of somatic mutations with evidence for Ag selection. Term neonates, however, had no such IgG repertoire. We conclude that restrictions in the neonatal Ig V(H) region gene repertoire persist until term despite exposure to environmental Ags. Yet, many weeks before term the immune system of the preterm neonate can already support germinal center reactions in response to environmental Ags. PMID- 12133960 TI - The Rap GTPases regulate B cell migration toward the chemokine stromal cell derived factor-1 (CXCL12): potential role for Rap2 in promoting B cell migration. AB - Stromal cell-derived factor-1 (SDF-1) is a potent chemoattractant for B cells and B cell progenitors. Although the binding of SDF-1 to its receptor, CXCR4, activates multiple signaling pathways, the mechanism by which SDF-1 regulates cell migration is not completely understood. In this report we show that activation of the Rap GTPases is important for B cells to migrate toward SDF-1. We found that treating B cells with SDF-1 resulted in the rapid activation of both Rap1 and Rap2. Moreover, blocking the activation of Rap1 and Rap2 via the expression of a Rap-specific GTPase-activating protein significantly reduced the ability of B cells to migrate toward SDF-1. Conversely, expressing a constitutively active form of Rap2 increased SDF-1-induced B cell migration. Thus, the Rap GTPases control cellular processes that are important for B cells to migrate toward SDF-1. PMID- 12133961 TI - The avian ChB6 alloantigen triggers apoptosis in a mammalian cell line. AB - Many developing B lymphocytes are deleted by apoptosis. However, the mechanism signaling their demise remains poorly understood. Like mammals, chicken B cells are selected during their development; >95% of the cells in the bursa of Fabricius die without entering the secondary immune system. The molecule chB6 (Bu 1) has been used as a marker to identify B cells in the chicken. ChB6 is a type I transmembrane glycoprotein whose function is enigmatic. We have provided evidence that chB6 can induce a rapid form of cell death exhibiting characteristics of apoptosis. Here we further examine cell death induced by chB6 in a transfected mouse cell line. ChB6 is shown to cause apoptosis in this cell line as detected by a TUNEL assay for DNA fragmentation. This apoptosis is subject to regulation by signals from growth factor or by Bcl-x(L). Furthermore, we show that Ab binding to chB6 leads to cleavage of caspase 8, caspase 3, and poly(ADP ribose) polymerase. Overall, these data support the hypothesis that chB6 is a novel death receptor on avian B cells. PMID- 12133962 TI - Isotype can affect the fine specificity of an antibody for a polysaccharide antigen. AB - Ab specificity is determined by V region sequence. The murine Mab 18B7 (IgG1) binds to the Cryptococcus neoformans capsular polysaccharide glucuronoxylomannan and produces annular immunofluorescence (IF) on yeast cells. The heavy and light V regions of 18B7 were expressed with the human C regions micro, gamma 1, gamma 2, gamma 3, gamma 4, and alpha1, and the specificity and binding properties of these mouse-human chimeric (ch) Abs was determined. The chIgG1, chIgG2, chIgG4, and the chIgA produced annular IF, whereas the IgM and IgG3 produced punctate IF, despite identical V region sequences. Competition experiments with murine Abs that competed with mAb 18B7 and binding assays to peptide mimetics of glucuronoxylomannan provided additional evidence for altered specificity in some of the ch Abs. Expression of 18B7 heavy V region with murine micro C region produced IgM with a punctate IF, indicating that a change in fine specificity also accompanied the change from murine IgG1 to IgM. Our results show that Ab fine specificity can be a function of isotype. This phenomenon may be most apparent for Abs that bind to Ag with repeating epitopes, such as polysaccharides, where the quarternary structure of the Ag-Ab complex may be influenced by such constraints as Fab-Fab angles, Fc-Fc interactions, Ab size, and solvent accessibility to exposed surfaces. Alterations in Ab fine specificity following isotype change could have important implications for current concepts on the generation of secondary Ab responses to certain Ags and for the isotype preference observed in Abs to polysaccharides. PMID- 12133963 TI - The chemokine ESkine/CCL27 displays novel modes of intracrine and paracrine function. AB - We have previously shown that the beta-chemokine ESkine/CCL27 is differentially spliced to produce two alternative forms. One is a secreted chemokine (ESkine), whereas the other (PESKY) lacks a signal peptide and is translocated to the nucleus. The role of this nuclear-targeted chemokine has not so far been defined, and it was the purpose of this study to examine this chemokine variant in more depth. To identify the region of PESKY involved in the nuclear translocation we tagged fragments with enhanced green fluorescent protein and expressed them in Chinese hamster ovary cells. We show PESKY nuclear translocation to be dependent on C-terminal residues that are shared with the signal peptide-bearing variant ESkine. Indeed we further demonstrate that ESkine can also use these C-terminal residues to enter the nucleus of cells following receptor (CCR10)-mediated internalization. To examine biological roles for PESKY we have overexpressed it in 3T3 cells. Such overexpression results in marked cytoskeletal rearrangements that are coincident with a radical reorganization of the cellular actin cytoskeleton. Microarray analyses and Ab neutralization studies indicate that these changes are mediated in part by insulin-like growth factor-1. Furthermore, monolayer wounding assays indicate that PESKY expression correlates with markedly increased migratory capacity. Thus, it is our contention that nuclear PESKY and ESkine both enter the nucleus by either intracrine or paracrine mechanisms and may facilitate cellular migration by inducing actin cytoskeletal relaxation. Therefore, nuclear ESkine/PESKY represents a novel paradigm for chemokine function. PMID- 12133964 TI - Structural studies of allelic diversity of the MHC class I homolog MIC-B, a stress-inducible ligand for the activating immunoreceptor NKG2D. AB - MIC-A and MIC-B are distant MHC class I homologs that serve as stress-inducible Ags on epithelial and epithelially derived cells. They are ligands for the widely expressed activating immunoreceptor NKG2D. To define the structural and functional consequences of sequence differences between MIC-A and MIC-B and between alleles of MIC-A and alleles of MIC-B, we determined the crystal structure of one allele of human MIC-B. Comparisons between the two previously reported MIC-A crystal structures and the MIC-B crystal structure show that, as expected, MIC-B is very similar in structure to MIC-A and likely interacts with NKG2D in an analogous manner. The interdomain flexibility observed in the MIC-A structures, a feature unique to MIC proteins among MHC class I proteins and homologs, is also displayed by MIC-B, with an interdomain relationship intermediate between the two examples of MIC-A structures. Mapping sequence variations onto the structures of MIC-A and MIC-B reveals patterns completely distinct from those displayed by classical MHC class I proteins, with a number of substitutions falling on positions likely to affect interactions with NKG2D, but with other positions lying distant from the NKG2D binding sites or buried within the core of the proteins. PMID- 12133965 TI - Mitogen-activated protein kinases and NF-kappa B are involved in TNF-alpha responses to group B streptococci. AB - TNF-alpha is a mediator of lethality in experimental infections by group B streptococcus (GBS), an important human pathogen. Little is known of signal transduction pathways involved in GBS-induced TNF-alpha production. Here we investigate the role of mitogen-activated protein kinases (MAPKs) and NF-kappa B in TNF-alpha production by human monocytes stimulated with GBS or LPS, used as a positive control. Western blot analysis of cell lysates indicates that extracellular signal-regulated kinase 1/2 (ERK 1/2), p38, and c-Jun N-terminal kinase MAPKs, as well as I kappa B alpha, became phosphorylated, and hence activated, in both LPS- and GBS-stimulated monocytes. The kinetics of these phosphorylation events, as well as those of TNF-alpha production, were delayed by 30-60 min in GBS-stimulated, relative to LPS-stimulated, monocytes. Selective inhibitors of ERK 1/2 (PD98059 or U0126), p38 (SB203580), or NF-kappa B (caffeic acid phenetyl ester (CAPE)) could all significantly reduce TNF-alpha production, although none of the inhibitors used alone was able to completely prevent TNF alpha release. However, this was completely blocked by combinations of the inhibitors, including PD98059-SB203580, PD98059-CAPE, or SB203580-CAPE combinations, in both LPS- and GBS-stimulated monocytes. In conclusion, our data indicate that the simultaneous activation of multiple pathways, including NF kappa B, ERK 1/2, and p38 MAPKs, is required to induce maximal TNF-alpha production. Accordingly, in septic shock caused by either GBS or Gram-negative bacteria, complete inhibition of TNF-alpha release may require treatment with drugs or drug combinations capable of inhibiting multiple activation pathways. PMID- 12133966 TI - Cross-presentation of Listeria monocytogenes-derived CD4 T cell epitopes. AB - Listeriolysin O (LLO) mediates the evasion of Listeria monocytogenes from the phagolysosome into the cytoplasm of the host cell. The recognition of infected cells by CD4 T cells is thought to be limited by the evasion of bacteria from the phagolysosome and also by the direct LLO-mediated inhibition of CD4 T cell activation. To analyze the influence of these immunoevasive mechanisms on the antilisterial CD4 T cell response, the expansion of L. monocytogenes-specific CD4 and CD8 T cells was monitored in infected mice. It was found that expansion of L. monocytogenes-specific CD4 T cells occurred synchronously with CD8 T cell expansion. The analysis of Ag presentation by macrophages and dendritic cells isolated from spleens of infected mice revealed efficient presentation of L. monocytogenes-derived CD4 T cell epitopes that was not dependent on the actA mediated intercellular spread of bacteria. The further in vitro Ag presentation analysis revealed that although L. monocytogenes-infected macrophages and dendritic cells were poor presenters of CD4 T cell epitopes, more efficient presentation occurred after cocultivation of noninfected dendritic cells or macrophages with infected cells. These data indicate that the suppressive effect of LLO on the antilisterial CD4 T cell response is maintained only in infected APC and support the hypothesis that cross-priming plays a role in the induction of the strong CD4 T cell response in Listeria-infected mice. PMID- 12133967 TI - Antibodies highly effective in SCID mice during infection by the intracellular bacterium Ehrlichia chaffeensis are of picomolar affinity and exhibit preferential epitope and isotype utilization. AB - Although often considered to be ineffective against intracellular bacteria, Abs, in the absence of lymphocytes, have been shown previously to protect SCID mice from lethal infection by the obligate intracellular bacterium Ehrlichia chaffeensis, even when administered well after infection has been established. To identify characteristics of Abs that are critical for host defense during this intracellular infection, a panel of Ehrlichia-specific mAbs was generated and analyzed. Among 100 Abs recovered, 39 recognized an amino-terminal hypervariable region of an outer membrane protein (OMP), demonstrating that the OMPs are both antigenically variable and immunodominant. A subset of 16 representative OMP specific Abs was further examined to identify characteristics that were essential for in vivo efficacy. The highly effective Abs recognized a linear epitope within the first hypervariable region of OMP-1g. Only IgG were found to be effective, and among the effective IgG, the following hierarchy was observed: IgG2a > IgG3 = IgG2b. The most striking characteristics of the highly effective Abs were their picomolar binding affinities and long binding t(1/2). Thus, although epitope recognition and isotype use may contribute to efficacy, high affinity may be a critical characteristic of Abs that can act effectively during this intracellular bacterial infection. PMID- 12133968 TI - Critical role of NK cells rather than V alpha 14(+)NKT cells in lipopolysaccharide-induced lethal shock in mice. AB - Although macrophages play a central role in the pathogenesis of septic shock, NK1(+) cells have also been implicated. NK1(+) cells comprise two major populations, namely NK cells and V alpha 14(+)NKT cells. To assess the relative contributions of these NK1(+) cells to LPS-induced shock, we compared the susceptibility to LPS-induced shock of beta(2)-microglobulin (beta(2)m)(-/-) mice that are devoid of V alpha 14(+)NKT cells, but not NK cells, with that of wild type (WT) mice. The results show that beta(2)m(-/-) mice were more susceptible to LPS-induced shock than WT mice. Serum levels of IFN-gamma following LPS challenge were significantly higher in beta(2)m(-/-) mice, and endogenous IFN-gamma neutralization or in vivo depletion of NK1(+) cells rescued beta(2)m(-/-) mice from lethal effects of LPS. Intracellular cytokine staining revealed that NK cells were major IFN-gamma producers. The J alpha 281(-/-) mice that are exclusively devoid of V alpha 14(+)NKT cells were slightly more susceptible to LPS-induced shock than heterozygous littermates. Hence, LPS-induced shock can be induced in the absence of V alpha 14(+)NKT cells and IFN-gamma from NK cells is involved in this mechanism. In WT mice, hierarchic contribution of different cell populations appears likely. PMID- 12133969 TI - The CXC chemokine murine monokine induced by IFN-gamma (CXC chemokine ligand 9) is made by APCs, targets lymphocytes including activated B cells, and supports antibody responses to a bacterial pathogen in vivo. AB - Monokine induced by IFN-gamma (Mig; CXC chemokine ligand 9) is an IFN-gamma inducible CXC chemokine that signals through the receptor CXCR3 and is known to function as a chemotactic factor for human T cells, particularly following T cell activation. The mig gene can be induced in multiple cell types and organs, and Mig has been shown to contribute to T cell infiltration into immune/inflammatory reactions in peripheral tissues in mice. We have investigated the expression and activities of Mig and CXCR3 in mouse cells and the role of Mig in models of host defense in mice. Murine (Mu)Mig functioned as a chemotactic factor for resting memory and activated T cells, both CD4(+) and CD8(+), and responsiveness to MuMig correlated with surface expression of MuCXCR3. Using mig(-/-) mice, we found that MuMig was not necessary for survival after infections with a number of intracellular pathogens. Surprisingly, however, we found that mig(-/-) mice showed reductions of 50-75% in Abs produced against the intracellular bacterium Francisella tularensis live vaccine strain. Furthermore, we found that MuMig induced both calcium signals and chemotaxis in activated B cells, and that B cell activation induced expression of MuCXCR3. In addition, IFN-gamma induced the expression of mumig in APCs, including CD8 alpha(+) and CD8 alpha(-) dendritic cells. Together, our data suggest that Mig and CXCR3 may be important not only to recruit T cells to peripheral inflammatory sites, but also in some cases to maximize interactions among activated T cells, B cells, and dendritic cells within lymphoid organs to provide optimal humoral responses to pathogens. PMID- 12133970 TI - Viral and bacterial infections induce expression of multiple NK cell receptors in responding CD8(+) T cells. AB - NK cells express several families of receptors that play central roles in target cell recognition. These NK cell receptors are also expressed by certain memory phenotype CD8(+) T cells, and in some cases are up-regulated in T cells responding to viral infection. To determine how the profile of NK receptor expression changes in murine CD8(+) T cells as they respond to intracellular pathogens, we used class I tetramer reagents to directly examine Ag-specific T cells during lymphocytic choriomeningitis virus and Listeria monocytogenes infections. We found that the majority of pathogen-specific CD8(+) T cells initiated expression of the inhibitory CD94/NKG2A heterodimer, the KLRG1 receptor, and a novel murine NK cell marker (10D7); conversely, very few Ag specific T cells expressed Ly49 family members. The up-regulation of these receptors was independent of IL-15 and persisted long after clearance of the pathogen. The expression of CD94/NKG2A was rapidly initiated in naive CD8(+) T cells responding to peptide Ags in vitro and on many of the naive T cells that proliferate when transferred into lymphopenic (Rag-1(-/-)) hosts. Thus, CD94/NKG2A expression is a common consequence of CD8(+) T cell activation. Binding of the CD94/NKG2A receptor by its ligand (Qa-1(b)) did not significantly inhibit CD8(+) T cell effector functions. However, expression of CD94 and NKG2A transgenes partially inhibited early events of T cell activation. These subtle effects suggest that CD94/NKG2A-mediated inhibition of T cells may be limited to particular circumstances or may synergize with other receptors that are similarly up-regulated. PMID- 12133971 TI - Marginal zone macrophages and immune responses against viruses. AB - The effective establishment of antiviral protection requires a coordinated interplay between the innate and adaptive immune system. Using osteopetrotic (op( /-)) mice, this study investigated the influence of marginal zone macrophages in controlling and initiating a protective immune response against a cytopathic vs a non- or low-cytopathic virus. Despite the generation of potent adaptive immune responses, antiviral protection against cytopathic vesicular stomatitis virus critically depended on the presence of marginal zone macrophages. Infection with low doses (100 PFU) of non- or low-cytopathic lymphocytic choriomeningitis virus was rarely cleared and usually resulted in a carrier state in the majority of mice. This shows that the early innate immune system provides an important preparatory phase to the adaptive immune system and is particularly important for antiviral protection. PMID- 12133972 TI - The importance of IL-1 beta and TNF-alpha, and the noninvolvement of IL-6, in the development of monoclonal antibody-induced arthritis. AB - Injection of anti-type II collagen Ab and LPS induces arthritis in mice. The levels of IL-1 beta, IL-6, and chemokines (macrophage inflammatory protein (MIP) 1 alpha, MIP-2, and monocyte chemoattractant protein-1) in the hind paws increased with the onset of arthritis and correlated highly with arthritis scores. The level of TNF-alpha was also elevated, but only transiently. Quantitative real-time PCR analysis revealed increases in cytokine and chemokine mRNA. To elucidate the contribution of inflammatory cytokines and chemokines in arthritis development more directly, recombinant proteins, neutralizing Abs, and knockout mice were used. The injection of rIL-1 beta or TNF-alpha, but not IL-6 or chemokines, induced arthritis when mice were i.v. preinjected with anti-type II collagen Ab. However, a single injection of recombinant cytokines or chemokines into the hind paws did not induce swelling. Arthritis development was inhibited by neutralizing Ab against IL-1 beta, TNF-alpha, or MIP-1 alpha. In contrast, the inhibitory effect by anti-MIP-2 Ab was partial and, surprisingly, Abs to IL-6 and monocyte chemoattractant protein-1 showed no inhibitory effect. Furthermore, arthritis development in IL-1R(-/-) mice and TNFR(-/-) mice was not observed at all, but severe arthritis was developed in IL-6(-/-) mice. These results suggest that IL-1 beta and TNF-alpha play more crucial roles than IL-6 or chemokines in this model. Because arthritis was also developed in SCID mice, the development of arthritis in the Ab-induced mice model is due to a mechanism that does not involve T or B cells. PMID- 12133973 TI - Characterization of thrombin-induced leukocyte rolling and adherence: a potential proinflammatory role for proteinase-activated receptor-4. AB - It is commonly accepted that thrombin exerts its proinflammatory properties through the activation of proteinase-activated receptor (PAR)-1, although two other thrombin receptors have been discovered: PAR-3 and PAR-4. In this study, we have investigated the mechanisms and the receptors involved in thrombin-induced leukocyte/endothelial cell interactions by using selective agonists and antagonists of thrombin receptors in an in vivo intravital microscopy system. Topical addition of selective PAR-1 agonists to rat mesenteric venules failed to reproduce the increased leukocyte rolling and adhesion observed after thrombin topical addition. When added together with the selective PAR-1 antagonist RWJ 56110, thrombin was still able to provoke increased leukocyte rolling and adherence. The thrombin-induced leukocyte rolling and adherence was not affected by pretreatment of rats with an anti-platelet serum. Selective PAR-4-activating peptide was able to reproduce the effects of thrombin on leukocyte rolling and adhesion. Intraperitoneal injection of PAR-4-activating peptide also caused a significant increase in leukocyte migration into the peritoneal cavity. In rat tissues, PAR-4 expression was detected both on endothelium and isolated leukocytes. Taken together, these results showed that in rat mesenteric venules, thrombin exerts proinflammatory properties inducing leukocyte rolling and adherence, by a mechanism independent of PAR-1 activation or platelet activation. However, PAR-4 activation either on endothelial cells or on leukocytes might be responsible for the thrombin-induced effects. These findings suggest that PAR-4 activation could contribute to several early events in the inflammatory reaction, including leukocyte rolling, adherence and recruitment, and that in addition to PAR-1, PAR-4 could be involved in proinflammatory properties of thrombin. PMID- 12133974 TI - Apical, but not basolateral, endotoxin preincubation protects alveolar epithelial cells against hydrogen peroxide-induced loss of barrier function: the role of nitric oxide synthesis. AB - The influence of LPS preincubation on hydrogen peroxide (H(2)O(2))-induced loss of epithelial barrier function was investigated in rat alveolar epithelial type II cells (ATII). Both apical and basolateral H(2)O(2) administration caused a manyfold increase in transepithelial [(3)H]mannitol passage. Apical but not basolateral preincubation of ATII with LPS did not influence control barrier properties but fully abrogated the H(2)O(2)-induced leakage response. The effect of apical LPS was CD14 dependent and was accompanied by a strong up-regulation of NO synthase II mRNA and protein and NO release. Inhibition of NO by N(G) monomethyl-L-arginine suppressed the LPS effect, whereas it was reproduced by exogenous application of gaseous NO or NO donor agents. Manipulation of the glutathione homeostasis (buthionine-(S,R)-sulfoximine) and the cGMP pathway (1H (1,2,4)oxadiazolo[4,3-alpha]quinoxaline-1-one; zaprinast) did not interfere with the protective effect of LPS. Superoxide (O*(-)(2)) generation by ATII cells was reduced by exogenous NO and LPS preincubation. O*(-)(2) scavenging with exogenous superoxide dismutase, the intracellular superoxide dismutase analog Mn(III)tetrakis(4-benzoic acid) porphyrin, and the superoxide scavenger nitroblue tetrazolium and, in particular, hydroxyl radical scavenging with hydroxyl radical scavenger 1,3-dimethyl-thiourea inhibited the H(2)O(2)-induced epithelial leakage response. In conclusion, apical but not basolateral LPS preincubation of ATII cells provides strong protection against H(2)O(2)-induced transepithelial leakage, attributable to an up-regulation of epithelial NO synthesis. It is suggested that the LPS-induced NO formation is effective via interaction with reactive oxygen species, including superoxide and hydroxyl radicals. The polarized epithelial response to LPS may be part of the lung innate immune system, activated by inhaled endotoxin or under conditions of pneumonia. PMID- 12133975 TI - Effects of rhinovirus infection on histamine and cytokine production by cell lines from human mast cells and basophils. AB - To understand the biochemical events that occur in the airways after rhinovirus (RV) infection, we developed for the first time a model in which the cell lines from human mast cells (HMC-1) and basophils (KU812) can be infected with RV14, a major group RV. Viral infection was confirmed by demonstrating that viral titers in culture supernatants, and RV RNA increased with time. RV14 infection alone and a combination of PMA plus calcium ionophore A23187, did not increase histamine production by these cells, although IgE plus anti-IgE increased the histamine production. However, histamine content in the supernatants increased in response to PMA plus A23187, or IgE plus anti-IgE after RV14 infection. PMA plus A23187 or IgE plus anti-IgE induced the production of IL-8 and GM-CSF in supernatants of HMC-1 cells and IL-4 and IL-6 in supernatants of KU812 cells. RV14 infection further increased the production of the cytokines, whereas RV14 infection alone did not alter the production of the cytokines by these cells. An Ab to ICAM-1 inhibited RV14 infection of the cells and decreased the production of cytokines and histamine after RV14 infection. RV14 infection enhanced the increases in intracellular calcium concentration and activation of NF-kappaB by PMA plus A23187 in the cells. These findings suggest that RV14 infection may prime the cytokine and histamine production from mast cells and basophils and may cause airway inflammation in asthma. PMID- 12133977 TI - Functional activation of the formyl peptide receptor by a new endogenous ligand in human lung A549 cells. AB - The formyl peptide receptor (FPR), a heptahelical G protein-coupled receptor on phagocytic leukocytes, can be triggered by bacterially derived oligopeptides of the prototype fMLP. Although FPR expression and activation have been associated with cells of myeloid origin and bacterial inflammation, the receptor has recently been identified in nonmyeloid cells, thus suggesting additional physiological functions and the existence of an endogenous agonist. In this study, we demonstrate the presence and functional activation of the FPR in the human lung cell line A549, which represents an extrahepatic model for the regulation of acute-phase proteins. Activation of the FPR in A549 cells cannot only be triggered by fMLP, but also by an agonistic peptide of the recently identified endogenous FPR ligand, annexin 1. In addition to inducing changes in the F-actin content, annexin 1-mediated triggering of the FPR results in an increased expression of acute-phase proteins. Hence, activation of nonmyeloid FPR by its endogenous ligand annexin 1 could participate in the regulation of acute phase responses, e.g., during inflammation and/or wound healing. PMID- 12133976 TI - Long-term protective and antigen-specific effect of heat-killed Mycobacterium vaccae in a murine model of allergic pulmonary inflammation. AB - This report examines the effect of heat-killed Mycobacterium vaccae in a mouse model of allergic pulmonary inflammation. The s.c. administration of M. vaccae 3 wk before the immunization significantly reduced Ag-induced airway hyperreactivity and the increase in the numbers of eosinophils observed in the bronchoalveolar lavage fluid, blood, and bone marrow, even though no detectable changes in either cytokine (IL-4, IL-13, IL-5, and IFN-gamma) or total IgE levels were observed. Furthermore, transfer of splenocytes from OVA-immunized and M. vaccae-treated mice into recipient, OVA-immunized mice significantly reduced the allergen-induced eosinophilia by an IFN-gamma-independent mechanism, clearly indicating that the mechanism by which M. vaccae induces its inhibitory effect is not due to a redirection from a predominantly Th2 to a Th1-dominated immune response. The protective effect of M. vaccae on the allergen-induced eosinophilia lasted for at least 12 wk after its administration, and the treatment was also effective in presensitized mice. Moreover, the allergen specificity of the inhibitory effect could be demonstrated using a double-immunization protocol, where M. vaccae treatment before OVA immunization had no effect on the eosinophilic inflammation induced by later immunization and challenge with cockroach extract Ag. Taken together, these results clearly demonstrate that M. vaccae is effective in blocking allergic inflammation by a mechanism independent of IFN-gamma, induces long term and Ag-specific protection, and therefore has both prophylactic and therapeutic potential for the treatment of allergic diseases. PMID- 12133978 TI - Leukocyte infiltration, but not neurodegeneration, in the CNS of transgenic mice with astrocyte production of the CXC chemokine ligand 10. AB - The CXC chemokine ligand (CXCL)10 is induced locally in the CNS in diverse pathologic states. The impact of CXCL10 production in the CNS was examined in transgenic mice with astrocyte-directed production of this chemokine. These glial fibrillary acidic protein (GF)-CXCL10 transgenic mice spontaneously developed transgene dose- and age-related leukocyte infiltrates in perivascular, meningeal, and ventricular regions of the brain that were composed of, surprisingly, mainly neutrophils and, to a lesser extent, T cells. No other overt pathologic or physical changes were evident. In addition, the cerebral expression of a number of inflammation-related genes (e.g., cytokines) was not significantly altered in the transgenic mice. The extent of leukocyte recruitment to the brain could be enhanced markedly by peripheral immunization of GF-CXCL10 mice with CFA and pertussis toxin. This was paralleled by a modest, transient increase in the expression of some cytokine and chemokine genes. Analysis of the expression of the CXCL10 receptor, CXCR3, by the brain-infiltrating leukocytes from immunized GF-CXCL10 transgenic mice revealed a significant enrichment for CXCR3-positive cells in the CNS compared with spleen. The majority of cells positive for CXCR3 coexpressed CD3, whereas Gr1-positive granulocytes were negative for CXCR3 expression. Thus, while astrocyte production of CXCL10 can promote spontaneous and potentiate immune-induced recruitment of leukocytes to the CNS, this is not associated with activation of a degenerative immune pathology. Finally, the accumulation of neutrophils in the brain of GF-CXCL10 transgenic mice is apparently independent of CXCR3 and involves an unknown mechanism. PMID- 12133979 TI - Stimulation by toll-like receptors inhibits osteoclast differentiation. AB - Osteoclasts, the cells capable of resorbing bone, are derived from hemopoietic precursor cells of monocyte-macrophage lineage. The same precursor cells can also give rise to macrophages and dendritic cells, which are essential for proper immune responses to various pathogens. Immune responses to microbial pathogens are often triggered because various microbial components induce the maturation and activation of immunoregulatory cells such as macrophages or dendritic cells by stimulating Toll-like receptors (TLRs). Since osteoclasts arise from the same precursors as macrophages, we tested whether TLRs play any role during osteoclast differentiation. We showed here that osteoclast precursors prepared from mouse bone marrow cells expressed all known murine TLRs (TLR1-TLR9). Moreover, various TLR ligands (e.g., peptidoglycan, poly(I:C) dsRNA, LPS, and CpG motif of unmethylated DNA, which act as ligands for TLR2, 3, 4, and 9, respectively) induced NF-kappa B activation and up-regulated TNF-alpha production in osteoclast precursor cells. Unexpectedly, however, TLR stimulation of osteoclast precursors by these microbial products strongly inhibited their differentiation into multinucleated, mature osteoclasts induced by TNF-related activation-induced cytokine. Rather, TLR stimulation maintained the phagocytic activity of osteoclast precursors in the presence of osteoclastogenic stimuli M-CSF and TNF related activation-induced cytokine. Taken together, these results suggest that TLR stimulation of osteoclast precursors inhibits their differentiation into noninflammatory mature osteoclasts during microbial infection. This process favors immune responses and may be critical to prevent pathogenic effects of microbial invasion on bone. PMID- 12133980 TI - Monocyte-derived dendritic cells induce a house dust mite-specific Th2 allergic inflammation in the lung of humanized SCID mice: involvement of CCR7. AB - In rodents, airway dendritic cells (DCs) capture inhaled Ag, undergo maturation, and migrate to the draining mediastinal lymph nodes (MLN) to initiate the Ag specific T cell response. However, the role of human DCs in the pathogenesis of the Th2 cell-mediated disease asthma remains to be clarified. Here, by using SCID mice engrafted with T cells from either house dust mite (HDM)-allergic patients or healthy donors, we show that DCs pulsed with Der p 1, one of the major allergens of HDM, and injected intratracheally into naive animals migrated into the MLN. In the MLN, Der p 1-pulsed DCs from allergic patients induced the proliferation of IL-4-producing CD4(+) T cells, whereas those from healthy donors induced IFN-gamma-secreting cells. In reconstituted human PBMC-reconstituted SCID mice primed with pulsed DCs from allergic patients, repeated exposure to aerosols of HDM induced 1) a strong pulmonary inflammatory reaction rich in T cells and eosinophils, 2) an increase in IL-4 and IL-5 production in the lung lavage fluid, and 3) increased IgE production compared with that in mice primed with unpulsed DCs. All these effects were reduced following in vivo neutralization of the CCR7 ligand secondary lymphoid tissue chemokine. These data in human PBMC reconstituted SCID mice show that monocyte-derived DCs might play a key role in the pathogenesis of the pulmonary allergic response by inducing Th2 effector function following migration to the MLN. PMID- 12133982 TI - Evidence for immune responses to a self-antigen in lung transplantation: role of type V collagen-specific T cells in the pathogenesis of lung allograft rejection. AB - We have reported that lung allograft rejection involves an immune response to a native protein in the lung, type V collagen (col(V)), and that col(V)-induced oral tolerance prevented acute and chronic rejection. In support of these findings col(V) fragments were detected in allografts during rejection, but not in normal lungs. The purpose of the current study was to isolate and characterize col(V)-specific allograft-infiltrating T cells and to determine their contribution to the rejection response in vivo. Two col(V)-specific T cell lines, LT1 and LT3, were isolated from F344 (RT1(lv1)) rat lung allografts during rejection that occurred after transplantation into WKY (RT1(l)) recipients. Both cell lines, but not normal lung lymphocytes, proliferated in response to col(V). Neither LT1 nor LT3 proliferated in response to alloantigens. LT1 and LT3 were CD4(+)CD25(-) and produced IFN-gamma in response to col(V). Compared with normal CD4(+) T cells, both cell lines expressed a limited V-beta TCR repertoire. Each cell strongly expressed V-beta 9 and 16, but differed in expression of other V betas. Adoptive transfer of each cell line did not induce pathology in lungs of normal WKY rats. In contrast, adoptive transfer of LT1, but not LT3, caused marked peribronchiolar and perivascular inflammation in isograft (WKY) lungs and abrogated col(V)-induced oral tolerance to allograft (F344) lungs. Collectively, these data show that lung allograft rejection involves both allo- and autoimmune responses, and graft destruction that occurs during the rejection response may expose allograft-infiltrating T cells to potentially antigenic epitopes in col(V). PMID- 12133983 TI - Bystander CD8 T cell-mediated demyelination after viral infection of the central nervous system. AB - Multiple sclerosis, a chronic inflammatory disease of the CNS, is characterized by immune-mediated demyelination. Many patients have a remitting-relapsing course of disease with exacerbations often following unrelated microbial illnesses. The relationship between the two events remains obscure. One possibility is that T cells specific for the inciting microbial pathogen are able to effect demyelination at a site of ongoing inflammation within the CNS. This possibility was examined in mice infected with mouse hepatitis virus, a well-described model of virus-induced demyelination. Using transgenic TCR/recombination activation gene 2(-/-) mice with only non-mouse hepatitis virus-specific T cells, we show that CD8 T cells are able to cause demyelination in the absence of cognate Ag in the CNS, but only if specifically activated. These findings demonstrate a novel mechanism for immune-mediated neuropathology and show that activated CD8 T cells may serve as important mediators of bystander demyelination during times of infection, including in patients with multiple sclerosis. PMID- 12133981 TI - Activation of toll-like receptor 2 in acne triggers inflammatory cytokine responses. AB - One of the factors that contributes to the pathogenesis of acne is Propionibacterium acnes; yet, the molecular mechanism by which P. acnes induces inflammation is not known. Recent studies have demonstrated that microbial agents trigger cytokine responses via Toll-like receptors (TLRs). We investigated whether TLR2 mediates P. acnes-induced cytokine production in acne. Transfection of TLR2 into a nonresponsive cell line was sufficient for NF-kappa B activation in response to P. acnes. In addition, peritoneal macrophages from wild-type, TLR6 knockout, and TLR1 knockout mice, but not TLR2 knockout mice, produced IL-6 in response to P. acnes. P. acnes also induced activation of IL-12 p40 promoter activity via TLR2. Furthermore, P. acnes induced IL-12 and IL-8 protein production by primary human monocytes and this cytokine production was inhibited by anti-TLR2 blocking Ab. Finally, in acne lesions, TLR2 was expressed on the cell surface of macrophages surrounding pilosebaceous follicles. These data suggest that P. acnes triggers inflammatory cytokine responses in acne by activation of TLR2. As such, TLR2 may provide a novel target for treatment of this common skin disease. PMID- 12133984 TI - Differential expression of the IFN-gamma-inducible CXCR3-binding chemokines, IFN inducible protein 10, monokine induced by IFN, and IFN-inducible T cell alpha chemoattractant in human cardiac allografts: association with cardiac allograft vasculopathy and acute rejection. AB - CXCR3 chemokines exert potent biological effects on both immune and vascular cells. The dual targets suggest their important roles in cardiac allograft vasculopathy (CAV) and rejection. Therefore, we investigated expression of IFN inducible protein 10 (IP-10), IFN-inducible T cell alpha chemoattractant (I-TAC), monokine induced by IFN (Mig), and their receptor CXCR3 in consecutive endomyocardial biopsies (n = 133) from human cardiac allografts and corresponding normal donor hearts (n = 11) before transplantation. Allografts, but not normal hearts, contained IP-10, Mig, and I-TAC mRNA. Persistent elevation of IP-10 and I TAC was associated with CAV. Allografts with CAV had an IP-10-GAPDH ratio 3.7 +/- 0.8 compared with 0.8 +/- 0.2 in those without CAV (p = 0.004). Similarly, I-TAC mRNA levels were persistently elevated in allografts with CAV (6.7 +/- 1.9 in allografts with vs 1.5 +/- 0.3 in those without CAV, p = 0.01). In contrast, Mig mRNA was induced only during rejection (2.4 +/- 0.9 with vs 0.6 +/- 0.2 without rejection, p = 0.015). In addition, IP-10 mRNA increased above baseline during rejection (4.1 +/- 2.3 in rejecting vs 1.8 +/- 1.2 in nonrejecting biopsies, p = 0.038). I-TAC did not defer significantly with rejection. CXCR3 mRNA persistently elevated after cardiac transplantation. Double immunohistochemistry revealed differential cellular distribution of CXCR3 chemokines. Intragraft vascular cells expressed high levels of IP-10 and I-TAC, while Mig localized predominantly in infiltrating macrophages. CXCR3 was localized in vascular and infiltrating cells. CXCR3 chemokines are induced in cardiac allografts and differentially associated with CAV and rejection. Differential cellular distribution of these chemokines in allografts indicates their central roles in multiple pathways involving CAV and rejection. This chemokine pathway may serve as a monitor and target for novel therapies to prevent CAV and rejection. PMID- 12133985 TI - Intrathecal Fas ligand infusion strengthens immunoprivilege of central nervous system and suppresses experimental autoimmune encephalomyelitis. AB - Fas ligand (FasL) is an essential molecule strongly expressed in some immunoprivileged sites, but is expressed at very low levels in normal CNS. In this study, acute experimental autoimmune encephalomyelitis (EAE) was induced in Lewis rats with guinea pig myelin basic protein. Intrathecal infusion of recombinant FasL before EAE onset dose dependently suppressed acute EAE and alleviated pathological inflammation in lumbosacral spinal cord. This treatment greatly increased apoptosis in CNS inflammatory cells, but did not inhibit systemic immune response to myelin basic protein. Systemic administration of a similar dose of rFasL was ineffective. In vitro, encephalitogenic T cells were highly sensitive to rFasL-induced cell death, and activated macrophages were also susceptible. In addition, in vitro rFasL treatment potentiated the immunosuppressive property of rat cerebrospinal fluid. We conclude that intrathecal infusion of rFasL eliminated the initial wave of infiltrating T cells and macrophages, and therefore blocked the later recruitment of inflammatory cells into CNS. Although Fas receptor expression was observed on spinal cord neurons, astrocytes, and oligodendrocytes, no damage to these cells or to the myelin structure was detected after rFasL infusion. PMID- 12133986 TI - The molecular basis of complete complement C4A and C4B deficiencies in a systemic lupus erythematosus patient with homozygous C4A and C4B mutant genes. AB - The disease course of a complete C4-deficient patient in the U.S. was followed for 18 years. The patient experienced multiple episodes of infection, and he was diagnosed with systemic lupus erythematosus at age 9 years. The disease progressed to WHO class III mild lupus nephritis and to fatal CNS vasculitis at age 23 years. Immunochemical experiments showed that the patient and his sibling had complete absence of C4A and C4B proteins and were negative for the Rodgers and Chido blood group Ags. Segregation and definitive RFLP analyses demonstrated that the patient and his sibling inherited two identical haplotypes, HLA A2 B12 DR6, each of which carries a defective long C4A gene and a defective short C4B gene. PCR and DNA sequencing revealed that the mutant C4A contained a 2-bp insertion in exon 29 at the sequence for codon 1213. The identical mutation was absent in the mutant C4B. The C4B mutant gene was selectively amplified by long range PCR, and its 41 exons were completely sequenced. The C4B mutant had a novel single C nucleotide deletion at the sequence for codon 522 in exon 13, leading to frame-shift mutation and premature termination. Thus, a multiplex PCR is designed by which known mutations in C4A and C4B can be elucidated conveniently. Among the 28 individuals reported with complete C4 deficiency, 75-96% of the subjects (dependent on the inclusion criteria) were afflicted with autoimmune or immune complex disorders. Hence, complete C4 deficiency is one of the most penetrant genetic risk factors for human systemic lupus erythematosus. PMID- 12133987 TI - Muscle responds to an antibody reactive with the acetylcholine receptor by up regulating monocyte chemoattractant protein 1: a chemokine with the potential to influence the severity and course of experimental myasthenia gravis. AB - Autoantibodies with reactivity against the postjunctional muscle receptor for acetylcholine receptor are able to interfere with contractile function of skeletal muscles and cause the symptoms of myasthenia gravis (MG) in humans, as well as in experimental animal models of MG. In the study described below using a rat model of MG, it was observed that exposure to acetylcholine receptor-reactive Abs also induced increased levels of chemokine (i.e., monocyte chemoattractant protein 1) production by skeletal muscle cells. This was true of both cultured rat myocytes exposed in vitro and rat muscle exposed in vivo following passive Ab transfer. Increased monocyte chemoattractant protein 1 production may explain the increased trafficking of leukocytes through muscle following Ab transfer described in this and other reports. These observations may also be relevant to the induction of disease symptoms in experimental animal models of MG, since numerous reports from this and other laboratories indicate that the cytokine environment provided by leukocytes trafficking through muscle may play a pivotal role in disease progression. PMID- 12133988 TI - A role for the Cr2 gene in modifying autoantibody production in systemic lupus erythematosus. AB - Systemic lupus erythematosus is an autoimmune disease characterized by autoantibody production against nuclear Ags. Recent studies suggest that the Cr2 gene, which encodes for complement receptor (CR)1 and CR2, is important in disease susceptibility. Because the precise disease phenotype related to this gene, in isolation or in relation to other genetic loci, is not known, we analyzed C57BL/6 mice with a targeted mutation in Cr2 (C57BL/6.Cr2(-/-)) with or without a concomitant mutation in Fas (C57BL/6.lpr Cr2(-/-)). The Cr2(null) mutation in a C57BL/6.lpr background markedly increases the serum concentrations of IgG1 and IgG2b and the levels of antinuclear and anti-dsDNA Abs as compared with C57BL/6.lpr controls. There is also a trend for higher concentrations of IgG2a and IgG3. In contrast, isolated deficiencies in either these CRs or Fas have a limited effect in the production of anti-dsDNA Abs. Moreover, the Cr2(null) mutation does not affect other disease manifestations. These findings demonstrate that abnormalities in CR1 and CR2 may be linked to the production of autoantibodies by modifying the effect of other systemic lupus erythematosus susceptibility genes. Phenotypic expression of other disease manifestations need additional Cr2-independent genetic factors. PMID- 12133989 TI - Epstein-Barr nuclear antigen 1-specific CD4(+) Th1 cells kill Burkitt's lymphoma cells. AB - The gamma-herpesvirus, EBV, is reliably found in a latent state in endemic Burkitt's lymphoma. A single EBV gene product, Epstein-Barr nuclear Ag 1 (EBNA1), is expressed at the protein level. Several mechanisms prevent immune recognition of these tumor cells, including a block in EBNA1 presentation to CD8(+) killer T cells. Therefore, no EBV-specific immune response has yet been found to target Burkitt's lymphoma. We now find that EBNA1-specific, Th1 CD4(+) cytotoxic T cells recognize Burkitt's lymphoma lines. CD4(+) T cell epitopes of EBNA1 are predominantly found in the C-terminal, episome-binding domain of EBNA1, and approximately 0.5% of peripheral blood CD4(+) T cells are specific for EBNA1. Therefore, adaptive immunity can be directed against Burkitt's lymphoma, and perhaps this role for CD4(+) Th1 cells extends to other tumors that escape MHC class I presentation. PMID- 12133990 TI - V75R576 IL-4 receptor alpha is associated with allergic asthma and enhanced IL-4 receptor function. AB - Asthma is a complex polygenic disease. Many studies have implicated the importance of IL-4R alpha in the development of allergic inflammation and its gene has been implicated in the genetics of asthma and atopy. In this study, we examined the functional consequences of two of the human IL-4R alpha allelic variants that have been found to associate with asthma and atopy. We examined the effects of each variant alone and in combination on IL-4-dependent gene induction. We found that neither the Q576R nor the I75V variants affected IL-4 dependent CD23 expression. However, the combination of V75R576 resulted in expression of an IL-4R alpha with enhanced sensitivity to IL-4. We next examined the genetics of five of the known IL-4R alpha allelic variants in asthmatic and nonatopic populations. Strikingly, the association of V75/R576 with atopic asthma was greater than either allele alone and the association of R576 with atopic asthma was dependent on the coexistence of V75. A haplotype analysis revealed a single IL-4R alpha haplotype that was associated with allergic asthma, VACRS, further confirming the importance of the V75 and R576 combination in the genetics of asthma. This is the first report demonstrating that a functional alteration in IL-4R alpha requires the coexistence of two naturally occurring single nucleotide polymorphisms (snps) in combination; neither snp alone is sufficient. These data illustrate the importance of studying snps in combination, because the functional significance of a given snp may only be evident in a specific setting of additional snps in the same or different genes. PMID- 12133992 TI - T cell subset patterns that predict resistance to spontaneous lymphoma, mammary adenocarcinoma, and fibrosarcoma in mice. AB - Aging leads to changes in the proportion of several T cell subsets in peripheral blood, but it is not yet known whether these changes have prognostic significance for late-life diseases. To examine this question, levels of T cell subsets were measured at 8 and 18 mo of age in the peripheral blood of mice of a genetically heterogeneous stock, and the mice were then subsequently evaluated for life span and for cause of death. The results indicate that mice whose T cell subset patterns look like those of old mice tend to die at earlier ages, regardless of the specific cause of death. At 18 mo, 39% of the variance within the set of seven measured subsets could be combined statistically into a single number, whose correlation with individual subsets suggested that it could be interpreted as an index of immunological aging. T cell subset pattern, as represented by this index, was a predictor of life span in mice dying of lymphoma, fibrosarcoma, mammary adenocarcinoma, or of all other causes considered together. Even as early as 8 mo of age, T cell subset patterns are significant predictors of all three forms of cancer, although at this age the association is stronger in mated female mice than in virgin mice. These results support two controversial hypotheses, which are not mutually exclusive: 1) early immune senescence might predispose to early death from cancer and 2) differences in aging rate, as monitored by tests of immune status, might accelerate or decelerate a wide range of late life neoplastic diseases. PMID- 12133991 TI - Detection and induction of CTLs specific for SYT-SSX-derived peptides in HLA A24(+) patients with synovial sarcoma. AB - To investigate the immunogenic property of peptides derived from the synovial sarcoma-specific SYT-SSX fusion gene, we synthesized four peptides according to the binding motif for HLA-A24. The peptides, SS391 (PYGYDQIMPK) and SS393 (GYDQIMPKK), were derived from the breakpoint of SYT-SSX, and SS449a (AWTHRLRER) and SS449b (AWTHRLRERK) were from the SSX region. These peptides were tested for their reactivity with CTL precursors (CTLps) in 16 synovial sarcoma patients using HLA-A24/SYT-SSX peptide tetramers and also for induction of specific CTLs from four HLA-A24(+) synovial sarcoma patients. Tetramer analysis indicated that the increased CTLp frequency to the SYT-SSX was associated with pulmonary metastasis in synovial sarcoma patients (p < 0.03). CTLs were induced from PBLs of two synovial sarcoma patients using the peptide mixture of SS391 and SS393, which lysed HLA-A24(+) synovial sarcoma cells expressing SYT-SSX as well as the peptide-pulsed target cells in an HLA class I-restricted manner. These findings suggest that aberrantly expressed SYT-SSX gene products have primed SYT-SSX specific CTLps in vivo and increased their frequency in synovial sarcoma patients. The identification of SYT-SSX peptides may offer an opportunity to design peptide-based immunotherapeutic approaches for HLA-A24(+) patients with synovial sarcoma. PMID- 12133993 TI - Role of the passive apoptotic pathway in graft-versus-host disease. AB - Donor T cells have been shown to undergo apoptosis during graft-vs-host disease (GVHD). Although active apoptosis mediated through Fas/Fas ligand interactions has been implicated in GVHD, little is known about the role of the passive apoptotic pathway. To examine this question, we compared the ability of normal donor T cells and T cells overexpressing the antiapoptotic protein, Bcl-x(L), to mediate alloreactive responses in vitro and lethal GVHD in vivo. In standard MLCs, T cells that overexpressed Bcl-x(L) had significantly higher proliferative responses but no difference in cytokine phenotype. Overexpression of Bcl-x(L) prolonged survival of both resting and alloactivated CD4(+) and CD8(+) T cells as assessed by quantitative flow cytometry, accounting for the higher proliferative responses. Analysis of engraftment in murine transplantation experiments demonstrated an increase in donor T cell chimerism in animals transplanted with Bcl-x(L) T cells, suggesting that overexpression of Bcl-x(L) prolonged T cell survival in vivo as well. Notably, transplantation of Bcl-x(L) T cells into nonirradiated F(1) recipients also significantly exacerbated GVHD as assessed by mortality and pathological damage in the gastrointestinal tract. However, when mice were irradiated no difference in GVHD mortality was observed between animals transplanted with wild-type and Bcl-x(L) T cells. These data demonstrate that the passive apoptotic pathway plays a role in the homeostatic survival of transplanted donor T cells. Moreover, the susceptibility of donor T cells to undergo passive apoptosis is a significant factor in determining GVHD severity under noninflammatory but not inflammatory conditions. PMID- 12133994 TI - Transfection of macrophage inflammatory protein 1 alpha into B16 F10 melanoma cells inhibits growth of pulmonary metastases but not subcutaneous tumors. AB - Macrophage inflammatory protein 1 alpha (MIP-1 alpha), a CC chemokine, is a chemoattractant for T cells and immature dendritic cells. Plasmacytoma cells expressing MIP-1 alpha generate a cytotoxic T cell response without affecting tumor growth. To understand this discrepancy, we compared a local tumor model with a metastatic one using MIP-1 alpha-transfected B16 F10 melanoma cells. Clonal idiosyncrasies were controlled by selecting three lipotransfected tumor clones and two pcDNA vector transfected control clones with equivalent in vitro proliferative capacities. No significant differences were seen between the MIP-1 alpha-producing and control melanoma cells after s.c. injection in the hind leg. All animals had a leg diameter of 10 cm in 18.5-21.5 days. However, after i.v. injection the number of pulmonary foci was significantly reduced in the MIP-1 alpha-producing clones. Injection of 10(6) control transfected cells resulted in a median of 98.5 tumor foci in 2 wk, whereas the injection of the MIP-1 alpha producing clones resulted in 89.5, 26.5, and 0 foci. The number of metastatic foci was inversely proportional to the amount of MIP-1 alpha produced by the clone in vitro. Flow cytometry showed a significant increase in CD8(+) cells in lungs of mice with MIP-1 alpha-transfected tumors 3 days after injection. This increase was not maintained 10 days later despite continued production of MIP-1 alpha. The protection offered by transfection with MIP-1 alpha was significantly impaired in beta(2)-microglobulin(-/-) mice. Our findings suggest that MIP-1 alpha is effective in preventing the initiation of metastasis, but not at sustaining an effective antitumor response. PMID- 12133995 TI - Interacting quantitative trait loci control loss of peripheral tolerance and susceptibility to autoimmune ovarian dysgenesis after day 3 thymectomy in mice. AB - Day 3 thymectomy (D3Tx) results in a loss of peripheral tolerance mediated by CD4(+)CD25(+) T cells and the development of autoimmune ovarian dysgenesis (AOD) in A/J and (C57BL/6J x A/J)F(1) (B6AF(1)) hybrids but not in C57BL/6J mice. Quantitative trait loci (QTL) linkage analysis using a B6AF(1) x C57BL/6J backcross population verified Aod1 and Aod2 that were previously mapped as qualitative traits. Additionally, three new QTL intervals, Aod3, Aod4, and Aod5, on chromosomes 1, 2, and 7, respectively, influencing specific subphenotypes of AOD were identified. QTL linkage analysis using the A x B and B x A recombinant inbred lines verified Aod3 and confirmed linkage to H2. Aod5 colocalized with Mater, an ovarian-specific autoantigen recognized by anti-ovarian autoantibodies in the sera of D3Tx mice. Sequence analysis of Mater identified allelic, strain specific splice variants between A/J and C57BL/6J mice making it an attractive candidate gene for Aod5. Interaction analysis revealed significant epistatic effects between Aod1-5 and Gasa2, a locus associated with susceptibility to D3Tx induced autoimmune gastritis, as well as with H2. These results indicate that the QTL controlling D3Tx-induced autoimmune phenomenon are both organ specific and more generalized in their effects with respect to the genesis and activity of the immunoregulatory mechanisms maintaining peripheral tolerance. PMID- 12133996 TI - Lentiviral neuropathogenesis: comparative neuroinvasion, neurotropism, neurovirulence, and host neurosusceptibility. PMID- 12133997 TI - Avian reoviruses cause apoptosis in cultured cells: viral uncoating, but not viral gene expression, is required for apoptosis induction. AB - The cytopathic effect evidenced by cells infected with avian reovirus S1133 suggests that this virus may induce apoptosis in primary cultures of chicken embryo fibroblasts. In this report we present evidence that avian reovirus infection of cultured cells causes activation of the intracellular apoptotic program and that this activation takes place during an early stage of the viral life cycle. The ability of avian reoviruses to induce apoptosis is not restricted to a particular virus strain or to a specific cell type, since different avian reovirus isolates were able to induce apoptosis in several avian and mammalian cell lines. Apoptosis was also provoked in ribavirin-treated avian reovirus infected cells and in cells infected with UV-irradiated reovirions, indicating that viral mRNA synthesis and subsequent steps in viral replication are not needed for apoptosis induction in avian reovirus-infected cells and that the number of inoculated virus particles, not their infectivity, is the critical factor for apoptosis induction by avian reovirus. Our finding that apoptosis is no longer induced when intracellular viral uncoating is blocked indicates that intraendosomal virion disassembly is required for apoptosis induction and that attachment and uptake of parental reovirions are not sufficient to cause apoptosis. Taken together, our results suggest that apoptosis is triggered from within the infected cell by viral products generated after intraendosomal uncoating of parental reovirions. PMID- 12133998 TI - Novel role of CD8(+) T cells and major histocompatibility complex class I genes in the generation of protective CD4(+) Th1 responses during retrovirus infection in mice. AB - CD4(+) Th1 responses to virus infections are often necessary for the development and maintenance of virus-specific CD8(+) T-cell responses. However, in the present study with Friend murine retrovirus (FV), the reverse was also found to be true. In the absence of a responder H-2(b) allele at major histocompatibility complex (MHC) class II loci, a single H-2D(b) MHC class I allele was sufficient for the development of a CD4(+) Th1 response to FV. This effect of H-2D(b) on CD4(+) T-cell responses was dependent on CD8(+) T cells, as demonstrated by depletion studies. A direct effect of CD8(+) T-cell help in the development of CD4(+) Th1 responses to FV was also shown in vaccine studies. Vaccination of nonresponder H-2(a/a) mice induced FV-specific responses of H-2D(d)-restricted CD8(+) cytotoxic T lymphocytes (CTL). Adoptive transfer of vaccine-primed CD8(+) T cells to naive H-2(a/a) mice prior to infection resulted in the generation of FV-specific CD4(+) Th1 responses. This novel helper effect of CD8(+) T cells could be an important mechanism in the development of CD4(+) Th1 responses following vaccinations that induce CD8(+) CTL responses. The ability of MHC class I genes to facilitate CD4(+) Th1 development could also be considerable evolutionary advantage by allowing a wider variety of MHC genotypes to generate protective immune responses against intracellular pathogens. PMID- 12133999 TI - Bunyamwera bunyavirus nonstructural protein NSs counteracts the induction of alpha/beta interferon. AB - Production of alpha/beta interferons (IFN-alpha/beta) in response to viral infection is one of the main defense mechanisms of the innate immune system. Many viruses therefore encode factors that subvert the IFN system to enhance their virulence. Bunyamwera virus (BUN) is the prototype of the Bunyaviridae family. By using reverse genetics, we previously produced a recombinant virus lacking the nonstructural protein NSs (BUNdelNSs) and showed that NSs is a nonessential gene product that contributes to viral pathogenesis. Here we demonstrate that BUNdelNSs is a strong inducer of IFN-alpha/beta, whereas in cells infected with the wild-type counterpart expressing NSs (wild-type BUN), neither IFN nor IFN mRNA could be detected. IFN induction by BUNdelNSs correlated with activation of NF-kappaB and was dependent on virally produced double-stranded RNA and on the IFN transcription factor IRF-3. Furthermore, both in cultured cells and in mice lacking a functional IFN-alpha/beta system, BUNdelNSs replicated to wild-type BUN levels, whereas in IFN-competent systems, wild-type BUN grew more efficiently. These results suggest that BUN NSs is an IFN induction antagonist that blocks the transcriptional activation of IFN-alpha/beta in order to increase the virulence of Bunyamwera virus. PMID- 12134001 TI - Comparative sequence analysis of the largest E1A proteins of human and simian adenoviruses. AB - The early region 1A (E1A) gene is the first gene expressed after infection with adenovirus and has been most extensively characterized in human adenovirus type 5 (hAd5). The E1A proteins interact with numerous cellular regulatory proteins, influencing a variety of transcriptional and cell cycle events. For this reason, these multifunctional proteins have been useful as tools for dissecting pathways regulating cell growth and gene expression. Despite the large number of studies using hAd5 E1A, relatively little is known about the function of the E1A proteins of other adenoviruses. In 1985, a comparison of E1A sequences from three human and one simian adenovirus identified three regions with higher overall levels of sequence conservation designated conserved regions (CR) 1, 2, and 3. As expected, these regions are critical for a variety of E1A functions. Since that time, the sequences of several other human and simian adenovirus E1A proteins have been determined. Using these, and two additional sequences that we determined, we report here a detailed comparison of the sequences of 15 E1A proteins representing each of the six hAd subgroups and several simian adenoviruses. These analyses refine the positioning of CR1, 2, and 3; define a fourth CR located near the carboxyl terminus of E1A; and suggest several new functions for E1A. PMID- 12134000 TI - Bovine leukemia virus SU protein interacts with zinc, and mutations within two interacting regions differently affect viral fusion and infectivity in vivo. AB - Bovine leukemia virus (BLV) and human T-cell lymphotropic virus type 1 (HTLV-1) belong to the genus of deltaretroviruses. Their entry into the host cell is supposed to be mediated by interactions of the extracellular (SU) envelope glycoproteins with cellular receptors. To gain insight into the mechanisms governing this process, we investigated the ability of SU proteins to interact with specific ligands. In particular, by affinity chromatography, we have shown that BLV SU protein specifically interacted with zinc ions. To identify the protein domains involved in binding, 16 peptides distributed along the sequence were tested. Two of them appeared to be able to interact with zinc. To unravel the role of these SU regions in the biology of the virus, mutations were introduced into the env gene of a BLV molecular clone in order to modify residues potentially interacting with zinc. The fusogenic capacity of envelope mutated within the first zinc-binding region (104 to 123) was completely abolished. Furthermore, the integrity of this domain was also required for in vivo infectivity. In contrast, mutations within the second zinc-binding region (218 to 237) did not hamper the fusogenic capacity; indeed, the syncytia were even larger. In sheep, mutations in region 218 to 237 did not alter infectivity or viral spread. Finally, we demonstrated that the envelope of the related HTLV-1 was also able to bind zinc. Interestingly, zinc ions were found to be associated with the receptor-binding domain (RBD) of Friend murine leukemia virus (Fr-MLV) SU glycoprotein, further supporting their relevance in SU structure. Based on the sequence similarities shared with the Fr-MLV RBD, whose three-dimensional structure has been experimentally determined, we located the BLV zinc-binding peptide 104-123 on the opposite side of the potential receptor-binding surface. This observation supports the hypothesis that zinc ions could mediate interactions of the SU RBD either with the C-terminal part of SU, thereby contributing to the SU structural integrity, or with a partner(s) different from the receptor. PMID- 12134002 TI - Multiple transmembrane amino acid requirements suggest a highly specific interaction between the bovine papillomavirus E5 oncoprotein and the platelet derived growth factor beta receptor. AB - The bovine papillomavirus E5 protein activates the cellular platelet-derived growth factor beta receptor (PDGFbetaR) tyrosine kinase in a ligand-independent manner. Evidence suggests that the small transmembrane E5 protein homodimerizes and physically interacts with the transmembrane domain of the PDGFbetaR, thereby inducing constitutive dimerization and activation of this receptor. Amino acids in the receptor previously found to be required for the PDGFbetaR-E5 interaction are a transmembrane Thr513 and a juxtamembrane Lys499. Here, we sought to determine if these are the only two receptor amino acids required for an interaction with the E5 protein. Substitution of large portions of the PDGFbetaR transmembrane domain indicated that additional amino acids in both the amino and carboxyl halves of the receptor transmembrane domain are required for a productive interaction with the E5 protein. Indeed, individual amino acid substitutions in the receptor transmembrane domain identified roles for the extracellular proximal transmembrane residues in the interaction. These data suggest that multiple amino acids within the transmembrane domain of the PDGFbetaR are required for a stable interaction with the E5 protein. These may be involved in direct protein-protein contacts or may support the proper transmembrane alpha-helical conformation for optimal positioning of the primary amino acid requirements. PMID- 12134003 TI - Given the opportunity, the Sendai virus RNA-dependent RNA polymerase could as well enter its template internally. AB - The negative-stranded RNA viral genome is an RNA-protein complex of helicoidal symmetry, resistant to nonionic detergent and high salt, in which the RNA is protected from RNase digestion. The 15,384 nucleotides of the Sendai virus genome are bound to 2,564 subunits of the N protein, each interacting with six nucleotides so tightly that the bases are poorly accessible to soluble reagents. With such a uniform structure, the question of template recognition by the viral RNA polymerase has been raised. In a previous study, the N-phase context has been proposed to be crucial for this recognition, a notion referring to the importance of the position in which the nucleotides interact with the N protein. The N-phase context ruled out the role of the template 3'-OH congruence, a feature resulting from the obedience to the rule of six that implies the precise interaction of the last six 3'-OH nucleotides with the last N protein. The N-phase context then allows prediction of the recognition by the RNA polymerase of a replication promoter sequence even if internally positioned, a promoter which normally lies at the template extremity. In this study, with template minireplicons bearing tandem replication promoters separated by intervening sequences, we present data that indeed show that initiation of RNA synthesis takes place at the internal promoter. This internal initiation can best be interpreted as the result of the polymerase entering the template at the internal promoter. In this way, the data are consistent with the importance of the N-phase context in template recognition. Moreover, by introducing between the two promoters a stretch of 10 A residues which represent a barrier for RNA synthesis, we found that the ability of the RNA polymerase to cross this barrier depends on the type of replication promoter, strong or weak, that the RNA polymerase starts on, a sign that the RNA polymerase may be somehow imprinted in its activity by the nature of the promoter on which it starts synthesis. PMID- 12134004 TI - Proteolytic processing of a serotype 8 human astrovirus ORF2 polyprotein. AB - Astroviruses require the proteolytic cleavage of the capsid protein to infect the host cell. Here we describe the processing pathway of the primary translation product of the structural polyprotein (ORF2) encoded by a human astrovirus serotype 8 (strain Yuc8). The primary translation product of ORF2 is of approximately 90 kDa, which is subsequently cleaved to yield a 70-kDa protein (VP70) which is assembled into the viral particles. Limited trypsin treatment of purified particles containing VP70 results in the generation of polypeptides VP41 and VP28, which are then further processed to proteins of 38.5, 35, and 34 kDa and 27, 26, and 25 kDa, respectively. VP34, VP27 and VP25 are the predominant proteins in fully cleaved virions, which correlate with the highest level of infectivity. Processing of the VP41 protein to yield VP38.5 to VP34 polypeptides occurred at its carboxy terminus, as suggested by immunoblot analysis using hyperimmune sera to different regions of the ORF2, while processing of VP28 to generate VP27 and VP25 occurred at its carboxy and amino terminus, respectively, as determined by immunoblot, as well as by N-terminal sequencing of those products. Based on these data, the processing pathway for the 90-kDa primary product of astrovirus Yuc8 ORF2 is presented. PMID- 12134005 TI - A novel herpes simplex virus type 1 transcript (AL-RNA) antisense to the 5' end of the latency-associated transcript produces a protein in infected rabbits. AB - Following primary ocular infection, herpes simplex virus type 1 (HSV-1) establishes a lifelong latent infection in sensory neurons of the trigeminal ganglia. Latency-associated transcript (LAT), the only known viral gene abundantly transcribed during HSV-1 neuronal latency, is required for high levels of reactivation. Recently we showed that three different mutants that do not alter the LAT promoter but contain deletions within the 5' end of the primary LAT transcript affect viral virulence (G. C. Perng et al., J. Virol. 75:9018-9028, 2001). In contrast, in LAT-null mutants viral virulence appears unaltered (T. M. Block et al., Virology 192:618-630, 1993; D. C. Bloom et al., J. Virol. 68:1283 1292, 1994; J. M. Hill et al., Virology 174:117-125, 1990; G. C. Perng et al., J. Virol. 68:8045-8055, 1994; F. Sedarati, K. M. Izumi, E. K. Wagner, and J. G. Stevens, J. Virol. 63:4455-4458, 1989). We therefore hypothesized that the 5' end of LAT and/or an as yet unidentified gene that overlaps part of this region is involved in viral virulence. We report here on the discovery and initial characterization of a novel HSV-1 RNA consistent with such a putative gene. The novel RNA was antisense to the 5' end of LAT and was designated AL-RNA (anti-LAT sense RNA). The AL-RNA overlapped the core LAT promoter and the first 158 nucleotides of the 5' end of the primary LAT transcript. AL-RNA was detected in extracts from neuron-like cells (PC-12) infected with wild-type HSV-1 but not in cells infected with a mutant with the AL region deleted. The deletions in each of the above three mutants with altered virulence encompass the 5' end of the AL RNA, and these mutants cannot transcribe AL. This supports the hypothesis that the AL gene may play a role in viral virulence. Based on comparison to the corresponding genomic sequence, the AL-RNA did not appear to be spliced. The AL RNA was polyadenylated and contained an open reading frame capable of encoding a protein 56 amino acids in length with a predicted molecular mass of 6.8 kDa. Sera from three of three rabbits infected with wild-type HSV-1 but not sera from any of three rabbits infected with a mutant with the AL-RNA region deleted recognized the Escherichia coli recombinantly expressed AL open reading frame on Western blots. In addition, four of six rabbits infected with wild-type virus developed enzyme-linked immunosorbent assay titers against one or more AL synthetic peptides. These results suggest that an AL protein is produced in vivo. PMID- 12134006 TI - Identification of two additional translation products from the matrix (M) gene that contribute to vesicular stomatitis virus cytopathology. AB - The matrix (M) protein of vesicular stomatitis virus (VSV) is a multifunctional protein that is responsible for condensation of the ribonucleocapsid core during virus assembly and also plays a critical role in virus budding. The M protein is also responsible for most of the cytopathic effects (CPE) observed in infected cells. VSV CPE include inhibition of host gene expression, disablement of nucleocytoplasmic transport, and disruption of the host cytoskeleton, which results in rounding of infected cells. In this report, we show that the VSV M gene codes for two additional polypeptides, which we have named M2 and M3. These proteins are synthesized from downstream methionines in the same open reading frame as the M protein (which we refer to here as M1) and lack the first 32 (M2) or 50 (M3) amino acids of M1. Infection of cells with a recombinant virus that does not express M2 and M3 (M33,51A) resulted in a delay in cell rounding, but virus yield was not affected. Transient expression of M2 and M3 alone caused cell rounding similar to that with the full-length M1 protein, suggesting that the cell-rounding function of the M protein does not require the N-terminal 50 amino acids. To determine if M2 and M3 were sufficient for VSV-mediated CPE, both M2 and M3 were expressed from a separate cistron in a VSV mutant background that readily establishes persistent infections and that normally lacks CPE. Infection of cells with the recombinant virus that expressed M2 and M3 resulted in cell rounding indistinguishable from that with the wild-type recombinant virus. These results suggest that M2 and M3 are important for cell rounding and may play an important role in viral cytopathogenesis. To our knowledge, this is first report of the multiple coding capacities of a rhabdovirus matrix gene. PMID- 12134007 TI - RelA-associated inhibitor blocks transcription of human immunodeficiency virus type 1 by inhibiting NF-kappaB and Sp1 actions. AB - RelA-associated inhibitor (RAI) is an inhibitor of nuclear factor kappaB (NF kappaB) newly identified by yeast two-hybrid screen as an interacting protein of the p65 (RelA) subunit. In this study, we attempted to examine the effect of RAI on transcription and replication of human immunodeficiency virus type 1 (HIV-1). We found that RAI inhibited gene expression from the HIV-1 long terminal repeat (LTR) even at the basal level. Upon in vitro DNA-binding reactions, RAI could directly block the DNA-binding of p65 subunit of NF-kappaB but not that of the p50 subunit or AP1. We found that RAI could also inhibit the DNA-binding of Sp1 and thus inhibit the basal HIV-1 promoter activity. We further examined the effects of RAI on Sp1 and found that RAI colocalizes with Sp1 in the nucleus and interacts with Sp1 in vitro and in vivo. Moreover, we found that RAI efficiently blocked the HIV-1 replication when cotransfected with a full-length HIV-1 clone. These findings indicate that RAI acts as an efficient inhibitor of HIV-1 gene expression in which both NF-kappaB and Sp1 play major roles. PMID- 12134008 TI - Walleye dermal sarcoma virus cyclin interacts with components of the mediator complex and the RNA polymerase II holoenzyme. AB - Walleye dermal sarcoma virus (WDSV) encodes an accessory protein, OrfA, with sequence homology to cyclins (retrovirus cyclin). In cells transfected with an expression construct, OrfA was localized to the nucleus and was concentrated in interchromatin granule clusters (IGCs), sites where splicing factors are concentrated. Other proteins identified in IGCs include transcription factors, the large subunit of RNA polymerase II (Pol II), and cyclin-dependent kinase 8 (cdk8). cdk8 is the kinase partner of cyclin C and a component of the mediator complex, associated with the Pol II holoenzyme. cdk8 and cyclin C can regulate transcription via phosphorylation of cyclin H and the carboxy-terminal domain of Pol II. OrfA in transfected HeLa cells was found to colocalize and copurify with hyperphosphorylated forms of Pol II (Pol IIO) in IGCs, and OrfA was coimmunoprecipitated from lysates of transfected cells with an antibody against Pol IIO. Likewise, Pol IIO could be coprecipitated with an antibody against OrfA. A survey with antibodies against several different cdks resulted in coimmunoprecipitation of OrfA with anti-cdk8, and antiserum against OrfA was able to coprecipitate cdk8 from lysates of cells that express OrfA. Coprecipitation of OrfA with anti-cyclin C demonstrated that it was included in complexes with OrfA and cdk8. OrfA has sequence and structural similarities to cyclin C, and, functionally, OrfA appears to have the capacity to both enhance and inhibit the activity of promoters in a cell-specific manner, similar to functions of the mediator complex. These data suggest that WDSV OrfA functions through its interactions with these large, transcription complexes. Further investigations will clarify the role of the retrovirus cyclin in control of virus expression and transformation. PMID- 12134009 TI - Survey of transcript expression in rainbow trout leukocytes reveals a major contribution of interferon-responsive genes in the early response to a rhabdovirus infection. AB - Virus infections induce changes in the expression of host cell genes. A global knowledge of these modifications should help to better understand the virus/host cell interactions. To obtain a more comprehensive view of the rainbow trout response to a viral infection, we used the subtractive suppressive hybridization methodology in the viral hemorrhagic septicemia model of infection. We infected rainbow trout leukocytes with viral hemorrhagic septicemia virus (VHSV), and total RNA from infected and mock-infected cells was compared at 40 h postinfection. Twenty-four virus-induced genes were ultimately retrieved from the subtracted cDNA library, and their differential expression was further confirmed by semiquantitative reverse transcription-PCR and Northern blot analysis. Among these sequences, three were already described as VHSV-induced genes. Eight sequences with known homologs were extended to full-length cDNA using 5' and 3' rapid amplification of cDNA ends, and they were subsequently divided into three functional subsets. Four genes were homologous to mammalian interferon responsive genes, three were similar to chemo-attractant molecules (CXC chemokine, galectin), and two had nucleic acid binding domains. All of the virus-induced genes were also induced by rainbow trout interferon, indicating that the interferon pathway is the predominant component of the anti-VHSV response. They were also expressed in vivo in experimentally infected fish, indicating their biological relevance in natural infection. PMID- 12134010 TI - Role for macrophage inflammatory protein 2 (MIP-2), MIP-1alpha, and interleukin 1alpha in the delayed-type hypersensitivity response to viral antigen. AB - BALB/c mice sensitized to herpes simplex virus type 1 (HSV-1) develop a vigorous delayed-type hypersensitivity (DTH) response upon intradermal virus antigen challenge. Although CD4(+) T cells are a key mediator of this response, neutrophils are the most abundant cells at the antigen challenge site both initially and at the peak of the reaction. We investigated what role, if any, neutrophils play in the DTH to a viral antigen. We show here that antibody mediated depletion of neutrophils 1 day before antigen challenge significantly suppressed ear swelling and markedly reduced cellular influx. Additionally, neutrophil depletion was associated with decreased expression of macrophage inflammatory protein 2 (MIP-2) and MIP-1alpha, as well as with a >60-fold increase in HSV-1 replication. Neutralizing antibodies to neutrophil chemoattractants MIP-2 or MIP-1alpha but not KC significantly suppressed DTH and sharply reduced neutrophil accumulation in the ear pinna. Purified bone marrow derived neutrophils exposed to interleukin-1alpha (IL-1alpha) produced chemokines in an 8-h assay. Administration of neutralizing antibody to IL-1alpha significantly reduced ear swelling and suppressed the levels of MIP-2, MIP 1alpha, MIP-1beta, and RANTES. We conclude that neutrophils are a critical component of the DTH response to viral antigen. They are recruited to the DTH test site by MIP-2 and MIP-1alpha, where they can be activated by IL-1alpha. The infiltrating cells also help suppress virus replication in immunized mice. PMID- 12134011 TI - Secondary structure of the 3' terminus of hepatitis C virus minus-strand RNA. AB - The 3'-terminal ends of both the positive and negative strands of the hepatitis C virus (HCV) RNA, the latter being the replicative intermediate, are most likely the initiation sites for replication by the viral RNA-dependent RNA polymerase, NS5B. The structural features of the very conserved 3' plus [(+)] strand untranslated region [3' (+) UTR] are well established (K. J. Blight and C. M. Rice, J. Virol. 71:7345-7352, 1997). However, little information is available concerning the 3' end of the minus [(-)] strand RNA. In the present work, we used chemical and enzymatic probing to investigate the conformation of that region, which is complementary to the 5' (+) UTR and the first 74 nucleotides of the HCV polyprotein coding sequence. By combining our experimental data with computer predictions, we have derived a secondary-structure model of this region. In our model, the last 220 nucleotides, where initiation of the (+) strand RNA synthesis presumably takes place, fold into five stable stem-loops, forming domain I. Domain I is linked to an overall less stable structure, named domain II, containing the sequences complementary to the pseudoknot of the internal ribosomal entry site in the 5' (+) UTR. Our results show that, even though the ( ) strand 3'-terminal region has the antisense sequence of the 5' (+) UTR, it does not fold into its mirror image. Interestingly, comparison of the replication initiation sites on both strands reveals common structural features that may play key functions in the replication process. PMID- 12134012 TI - Genetic and biochemical analyses of receptor and cofactor determinants for T-cell tropic feline leukemia virus infection. AB - Entry by retroviruses is mediated through interactions between the viral envelope glycoprotein and the host cell receptor(s). We recently identified two host cell proteins, FeLIX and Pit1, that are necessary for infection by cytopathic, T-cell tropic feline leukemia viruses (FeLV-T). Pit1 is a classic multiple transmembrane protein used as a receptor by several other simple retroviruses, including subgroup B FeLV (FeLV-B), and FeLIX is a secreted cellular protein expressed from endogenous FeLV-related sequences (enFeLV). FeLIX is nearly identical to FeLV-B envelope sequences that encode the N-terminal half of the viral surface unit (SU), because these FeLV-B sequences are acquired by recombination with enFeLV. FeLV-B SUs can functionally substitute for FeLIX in mediating FeLV-T infection. Both of these enFeLV-derived cofactors can efficiently facilitate FeLV-T infection only of cells expressing Pit1, not of cells expressing the related transport protein Pit2. We therefore have used chimeric Pit1/Pit2 receptors to map the determinants for cofactor binding and FeLV-T infection. Three distinct determinants appear to be required for cofactor-dependent infection by FeLV-T. We also found that Pit1 sequences within these same domains were required for binding by FeLIX to the Pit receptor. In contrast, these determinants were not all required for receptor binding by the FeLV-B SU cofactors used in this study. These data indicate that cofactor binding is not sufficient for FeLV-T infection and suggest that there may be a direct interaction between FeLV-T and the Pit1 receptor. PMID- 12134013 TI - The carboxy-terminal region of the human immunodeficiency virus type 1 protein Rev has multiple roles in mediating CRM1-related Rev functions. AB - The human immunodeficiency virus type 1 (HIV-1) regulatory protein, Rev, mediates the nuclear export of unspliced and singly spliced viral mRNAs by bridging viral RNA and export receptor human CRM1 (hCRM1). Ribonucleoprotein complex formation, including the oligomerization of Rev proteins on viral RNA, must occur to allow export. We show here that Rev-Rev interactions, which are a basis of complex formation, can be initiated without cellular factors and are subsequently enhanced by hCRM1-Ran-GTP. Furthermore, we reveal functions for the Rev carboxy terminal (C-terminal) region, which is well conserved among many HIV-1 strains, and for which no function has been reported. This region is required for the efficient binding of Rev to hCRM1 and consequently for nuclear export, Rev-Rev dimerization, and full Rev transactivator activity. Consistent with these results, a HIV-1 proviral plasmid that expresses a C-terminally truncated Rev mutant protein produces smaller amounts of the p24 antigen than does a plasmid that possesses an intact rev gene. These results indicate the functional importance of the C-terminal region for full Rev activity, which leads to efficient HIV-1 replication. PMID- 12134014 TI - The herpes simplex virus type 1 US11 protein binds the coterminal UL12, UL13, and UL14 RNAs and regulates UL13 expression in vivo. AB - The US11 protein of herpes simplex virus type 1 (HSV-1) is a small, highly basic phosphoprotein expressed at late times during infection. US11 localizes to the nucleolus in infected cells, can associate with ribosomes, and has been shown to bind RNA. The RNA substrates of US11 identified thus far have no apparent role in the virus lytic cycle, so we set out to identify a novel, biologically relevant RNA substrate(s) for this protein in HSV-1-infected cells. We designed a reverse transcriptase PCR-based protocol that allowed specific selection of a 600-bp RNA binding partner for US11. This RNA sequence, designated 12/14, is present in the coterminal HSV-1 mRNAs UL12, UL13, and UL14. We show that the binding of US11 to 12/14 is sequence-specific and mediated by the C-terminal domain of the protein. To elucidate the role of US11 in the virus life cycle, we infected cells with wild-type virus, a cosmid-reconstructed US11 HSV-1 null mutant, and a cosmid reconstructed wild-type virus and analyzed expression of UL12, -13, and -14 during a time course of infection. These experiments revealed that this interaction has biological activity; at early times of infection, US11 down regulates UL13 protein kinase mRNA and protein. PMID- 12134015 TI - Role of a highly conserved NH(2)-terminal domain of the human parainfluenza virus type 3 RNA polymerase. AB - The RNA polymerase complex of human parainfluenza virus type 3 (HPIV 3), a member of the family Paramyxoviridae, is composed of two virally encoded polypeptides: a multifunctional large protein (L, 255 kDa) and a phosphoprotein (P, 90 kDa). From extensive deduced amino acid sequence analyses of the cDNA clones of a number of L proteins of nonsegmented negative-strand RNA viruses, a cluster of high homology sequence segments have been identified within the body of the L proteins. Here, we have focused on the NH(2)-terminal domain of HPIV 3 L protein that is also highly conserved. Following mutational analyses within this domain, we examined the ability of the mutant L proteins to (i) transcribe an HPIV 3 minireplicon, (ii) transcribe the viral RNA in vitro using the HPIV 3 nucleocapsid RNA template, and (iii) interact with HPIV 3 P protein. Our results demonstrate that the first 15 amino acids of the NH(2)-terminal domain spanning a highly conserved motif is directly involved in transcription of the genome RNA and in forming a functional complex with the P protein. Substitution of eight nonconserved amino acids within this domain by the corresponding Sendai virus L protein residues yielded mutants with variable transcriptional activities. However, one mutant in which all eight amino acids were replaced with the corresponding residues of Sendai virus L protein failed to both transcribe the minireplicon and interact with HPIV 3 P and the Sendai virus P protein. The possible functional significance of the NH(2)-terminal domain of paramyxovirus L protein is discussed. PMID- 12134016 TI - Heterologous protection induced by the inner capsid proteins of rotavirus requires transcytosis of mucosal immunoglobulins. AB - Protective immunization against rotavirus (RV) can be achieved with heterologous RV, i.e., virus isolated from another species, and with heterologous inner core VP2 and VP6 proteins assembled as virus-like particles (VLP). Although the antigenically conserved VP6 protein does not induce in vitro-neutralizing antibodies, it may, however, elicit immunoglobulins (Ig) involved in heterologous protection, as some IgA against VP6 prevent RV infection in a backpack mouse model. The protective role of Ig directed to the RV inner core proteins VP2 and VP6 was investigated in J-chain-deficient mice (J chain(-/-)), which have a defect in the polymeric Ig receptor (pIgR)-mediated transcytosis of IgA and IgM. J chain(-/-) mice and wild-type (WT) mice were intranasally vaccinated with bovine RV-derived VLP2/6 and then challenged with highly infectious murine ECw RV. Whereas WT mice were totally protected, immunized J chain(-/-) mice shed RV for several days. In addition, naive J chain(-/-) mice exhibited a 2-day delay in clearing RV compared with WT mice. The immunized J chain(-/-) mice displayed unaltered VLP2/6-specific B-cell numbers in spleen and in mesenteric nodes and similar levels of serum anti-VLP2/6 Ig, confirming that the adaptive B-cell response is preserved in J chain(-/-) mice. These results indicate that J-chain mediated transcytosis of Ig participates in the clearance of RV and that epithelial pIgR-mediated transport of Ig is involved in the heterologous protection induced by VLP2/6. PMID- 12134017 TI - Effects of prostratin on T-cell activation and human immunodeficiency virus latency. AB - Human immunodeficiency virus (HIV) replication is linked to cellular gene transcription and requires target cell activation. The latent reservoir of HIV-1 in quiescent T cells is thought to be a major obstacle to clearance of infection by highly active antiretroviral therapy (HAART). Thus, identification of agents that can induce expression of latent virus may, in the presence of HAART, allow elimination of the infected cells by the immune response. We previously used the SCID-hu (Thy/Liv) mouse model to establish that activation-inducible HIV can be generated at high frequency during thymopoiesis. Latently infected mature thymocytes can be exported into the periphery, providing an efficient primary cell model to determine cellular activation signals that induce renewed expression of latent virus. Here we characterized the effects of prostratin, a non-tumor-promoting phorbol ester, on primary human peripheral blood lymphocytes (PBLs) and assessed its ability to reactivate latent HIV infection from thymocytes and PBLs in the SCID-hu (Thy/Liv) model. Prostratin stimulation alone did not induce proliferation of quiescent PBLs; however, it could provide a secondary signal in the context of T-cell receptor stimulation or a primary activation signal in the presence of CD28 stimulation to induce T-cell proliferation. While prostratin alone was not sufficient to allow de novo HIV infection, it efficiently reactivated HIV expression from latently infected cells generated in the SCID-hu mouse. Our data indicate that prostratin alone is able to specifically reactivate latent virus in the absence of cellular proliferation, making it an attractive candidate for further study as an adjunctive therapy for the elimination of the latent HIV reservoir. PMID- 12134018 TI - Identification of SRPK1 and SRPK2 as the major cellular protein kinases phosphorylating hepatitis B virus core protein. AB - Phosphorylation of hepatitis B virus (HBV) core protein has recently been shown to be a prerequisite for pregenomic RNA encapsidation into viral capsids, but the host cell kinases mediating this essential step of the HBV replication cycle have not been identified. We detected two kinases of 95 and 115 kDa in HuH-7 total cell lysates which interacted specifically with the HBV core protein and phosphorylated its arginine-rich C-terminal domain. The 95-kDa kinase was purified and characterized as SR protein-specific kinase 1 (SRPK1) by mass spectrometry. Based on this finding, the 115-kDa kinase could be identified as the related kinase SRPK2 by immunoblot analysis. In vitro, both SRPKs phosphorylated HBV core protein on the same serine residues which are found to be phosphorylated in vivo. Moreover, the major cellular HBV core kinase activity detected in the total cell lysate showed biochemical properties identical to those of SRPK1 and SRPK2, as examined by measuring binding to a panel of chromatography media. We also clearly demonstrate that neither the cyclin dependent kinases Cdc2 and Cdk2 nor protein kinase C, previously implicated in HBV core protein phosphorylation, can account for the HBV core protein kinase activity. We conclude that both SRPK1 and SRPK2 are most likely the cellular protein kinases mediating HBV core protein phosphorylation during viral infection and therefore represent important host cell targets for therapeutic intervention in HBV infection. PMID- 12134019 TI - Mutations that affect the tropism of DA and GDVII strains of Theiler's virus in vitro influence sialic acid binding and pathogenicity. AB - Theiler's murine encephalomyelitis virus (TMEV) is a natural pathogen of the mouse. The different strains of TMEV are divided into two subgroups according to the pathology they provoke. The neurovirulent strains GDVII and FA induce an acute fatal encephalitis, while persistent strains, like DA and BeAn, cause a chronic demyelinating disease associated with viral persistence in the central nervous system. Different receptor usage was proposed to account for most of the phenotype difference between neurovirulent and persistent strains. Persistent but not neurovirulent strains were shown to bind sialic acid. We characterized DA and GDVII derivatives adapted to grow on CHO-K1 cells. Expression of glycosaminoglycans did not influence infection of CHO-K1 cells by parental and adapted viruses. Mutations resulting from adaptation of DA and GDVII to CHO-K1 cells notably mapped to the well-characterized VP1 CD and VP2 EF loops of the capsid. Adaptation of the DA virus to CHO-K1 cells correlated with decreased sialic acid usage for entry. In contrast, adaptation of the GDVII virus to CHO-K1 cells correlated with the appearance of a weak sialic acid usage for entry. The sialic acid binding capacity of the GDVII variant resulted from a single amino acid mutation (VP1-51, Asn-->Ser) located out of the sialic acid binding region defined for virus DA. Mutations affecting tropism in vitro and sialic acid binding dramatically affected the persistence and neurovirulence of the viruses. PMID- 12134020 TI - Rebound of hepatitis B virus replication in HepG2 cells after cessation of antiviral treatment. AB - Treatment of patients with lamivudine (3TC) results in loss of detectable levels of hepatitis B virus (HBV) DNA from serum; however, the relapse rate, with regard to both reappearance of virus in the bloodstream and hepatic inflammation, is high when therapy is terminated. Although the rebound observed in patients has also been seen in animal hepadnavirus models, rebound has not been analyzed in an in vitro cell culture system. In this study, we used the HBV recombinant baculovirus/HepG2 system to measure the time course of antiviral agent-mediated loss of HBV replication as well as the time course and magnitude of HBV production after release from antiviral treatment. Because of the sensitivity of the system, it was possible to measure secreted virions, intracellular replicative intermediates, and nuclear non-protein-bound HBV DNA and separately analyze individual species of DNA, such as single-stranded HBV DNA compared to the double-stranded form and relaxed circular compared to covalently closed circular HBV DNA. We first determined that HBV replication in the HBV recombinant baculovirus/HepG2 system could proceed for at least 35 days, with a 30-day plateau level of replication, making it possible to study antiviral agent mediated loss of HBV followed by rebound after cessation of drug treatment. All HBV DNA species decreased in a time-dependent fashion following antiviral treatment, but the magnitude of decline differed for each HBV DNA species, with the covalently closed circular form of HBV DNA being the most resistant to drug therapy. When drug treatment ceased, HBV DNA species reappeared with a pattern that recapitulated the initiation of replication, but with a different time course. PMID- 12134021 TI - Murine cytomegalovirus (CMV) M33 and human CMV US28 receptors exhibit similar constitutive signaling activities. AB - Cellular infection by cytomegalovirus (CMV) is associated with very early G protein-mediated signal transduction and reprogramming of gene expression. Here we investigated the involvement of human CMV (HCMV)-encoded US27, US28, and UL33 receptors as well as murine CMV-encoded M33 transmembrane (7TM) receptors in host cell signaling mechanisms. HCMV-encoded US27 did not show any constitutive activity in any of the studied signaling pathways; in contrast, US28 and M33 displayed ligand-independent, constitutive signaling through the G protein q (Gq)/phospholipase C pathway. In addition, M33 and US28 also activated the transcription factor NF-kappaB as well as the cyclic AMP response element binding protein (CREB) in a ligand-independent, constitutive manner. The use of specific inhibitors indicated that the p38 mitogen-activated protein (MAP) kinase but not the extracellular signal-regulated kinase 1/2-MAP kinase pathway is involved in M33- and US28-mediated CREB activation but not NF-kappaB activation. Interestingly, UL33-the HCMV-encoded structural homologue of M33-was only marginally constitutively active in the Gq/phospholipase C turnover and CREB activation assays and did not show any constitutive activity in the NF-kappaB pathway, where M33 and US28 were highly active. Hence, CMVs appear to have conserved mechanisms for regulating host gene transcription, i.e., constitutive activation of certain kinases and transcription factors through the constitutive activities of 7TM proteins. These data, together with the previous identification of the incorporation of such proteins in the viral envelope, suggest that these proteins could be involved in the very early reprogramming of the host cell during viral infection. PMID- 12134022 TI - Bacteriophage PM2 has a protein capsid surrounding a spherical proteinaceous lipid core. AB - The marine double-stranded DNA (dsDNA) bacteriophage PM2, studied since 1968, is the type organism of the family Corticoviridae, infecting two gram-negative Pseudoalteromonas species. The virion contains a membrane underneath an icosahedral protein capsid composed of two structural proteins. The purified major capsid protein, P2, appears as a trimer, and the receptor binding protein, P1, appears as a monomer. The C-terminal part of P1 is distal and is responsible for receptor binding activity. The rest of the structural proteins are associated with the internal phospholipid membrane enclosing the viral genome. This internal particle is designated the lipid core. The overall structural organization of phage PM2 resembles that of dsDNA bacteriophage PRD1, the type organism of the family TECTIVIRIDAE: PMID- 12134023 TI - The lytic cycle of Epstein-Barr virus is associated with decreased expression of cell surface major histocompatibility complex class I and class II molecules. AB - Human herpesviruses utilize an impressive range of strategies to evade the immune system during their lytic replicative cycle, including reducing the expression of cell surface major histocompatibility complex (MHC) and immunostimulatory molecules required for recognition and lysis by virus-specific cytotoxic T cells. Study of possible immune evasion strategies by Epstein-Barr virus (EBV) in lytically infected cells has been hampered by the lack of an appropriate permissive culture model. Using two-color immunofluorescence staining of cell surface antigens and EBV-encoded lytic cycle antigens, we examined EBV transformed B-cell lines in which a small subpopulation of cells had spontaneously entered the lytic cycle. Cells in the lytic cycle showed a four- to fivefold decrease in cell surface expression of MHC class I molecules relative to that in latently infected cells. Expression of MHC class II molecules, CD40, and CD54 was reduced by 40 to 50% on cells in the lytic cycle, while no decrease was observed in cell surface expression of CD19, CD80, and CD86. Downregulation of MHC class I expression was found to be an early-lytic-cycle event, since it was observed when progress through late lytic cycle was blocked by treatment with acyclovir. The immediate-early transactivator of the EBV lytic cycle, BZLF1, did not directly affect expression of MHC class I molecules. However, BZLF1 completely inhibited the upregulation of MHC class I expression mediated by the EBV cell-transforming protein, LMP1. This novel function of BZLF1 elucidates the paradox of how MHC class I expression can be downregulated when LMP1, which upregulates MHC class I expression in latent infection, remains expressed in the lytic cycle. PMID- 12134025 TI - Molecular architecture of adenovirus DNA polymerase and location of the protein primer. AB - Adenovirus (Ad) DNA polymerase (pol) belongs to the distinct subclass of the polalpha family of DNA pols that employs the precursor terminal protein (pTP) as primer. Ad pol forms a stable heterodimer with this primer, and together, they bind specifically to the core origin in order to start replication. After initiation of Ad replication, the resulting pTP-trinucleotide intermediate jumps back and pTP starts to dissociate. Compared to free Ad pol, the pTP-pol complex shows reduced polymerase and exonuclease activities, but the reason for this is not understood. Furthermore, the interaction domains between these proteins have not been defined and the contribution of each protein to origin binding is unclear. To address these questions, we used oligonucleotides with a translocation block and show here that pTP binds at the entrance of the primer binding groove of Ad pol, thereby explaining the decreased synthetic activities of the pTP-pol complex and providing insight into how pTP primes Ad replication. Employing an exonuclease-deficient mutant polymerase, we further show that the polymerase and exonuclease active sites of Ad pol are spatially distinct and that the exonuclease activity of Ad pol is located at the N-terminal part of the protein. In addition, by probing the distances between both active sites and the surface of Ad pol, we show that Ad pol binds a DNA region of 14 to 15 nucleotides. Based on these results, a model for binding of the pTP-pol complex at the origin of replication is proposed. PMID- 12134024 TI - Alpha interferon inhibits hepatitis C virus replication in primary human hepatocytes infected in vitro. AB - Chronic hepatitis C is a common cause of liver disease, the complications of which include cirrhosis and hepatocellular carcinoma. Treatment of chronic hepatitis C is based on the use of alpha interferon (IFN-alpha). Recently, indirect evidence based on mathematical modeling of hepatitis C virus (HCV) dynamics during human IFN-alpha therapy suggested that the major initial effect of IFN-alpha is to block HCV virion production or release. Here, we used primary cultures of healthy, uninfected human hepatocytes to show that: (i) healthy human hepatocytes can be infected in vitro and support HCV genome replication, (ii) hepatocyte treatment with IFN-alpha results in expression of IFN-alpha-induced genes, and (iii) IFN-alpha inhibits HCV replication in infected human hepatocytes. These results show that IFN-alpha acts primarily through its nonspecific antiviral effects and suggest that primary cultures of human hepatocytes may provide a good model to study intrinsic HCV resistance to IFN alpha. PMID- 12134026 TI - Physical interaction between envelope glycoproteins E and M of pseudorabies virus and the major tegument protein UL49. AB - Envelope glycoprotein M (gM) and the complex formed by glycoproteins E (gE) and I (gI) are involved in the secondary envelopment of pseudorabies virus (PrV) particles in the cytoplasm of infected cells. In the absence of the gE-gI complex and gM, envelopment is blocked and capsids surrounded by tegument proteins accumulate in the cytoplasm (A. R. Brack, J. Dijkstra, H. Granzow, B. G. Klupp, and T. C. Mettenleiter, J. Virol. 73:5364-5372, 1999). Here we demonstrate by yeast two-hybrid analyses that the cytoplasmic domains of gE and gM specifically interact with the C-terminal part of the UL49 gene product of PrV, which represents a major tegument protein and which is homologous to VP22 of herpes simplex virus type 1. However, deletion of the UL49 gene from PrV had only minor effects on viral replication, and ultrastructural analyses of infected cells confirmed that virus maturation and egress, including secondary envelopment in the cytoplasm, were not detectably affected by the absence of UL49. Moreover, the UL49 gene product was shown to be dispensable for virion localization of gE and gM, and mutants lacking either gE or gM incorporated the UL49 protein efficiently into virus particles. In contrast, a PrV mutant with deletions of gE-gI and gM failed to incorporate the UL49 protein despite apparently unaltered intracytoplasmic UL49 expression. In summary, we describe specific interactions between herpesvirus envelope and tegument proteins which may play a role in secondary envelopment during herpesvirus virion maturation. PMID- 12134027 TI - Characterization of a polytropic murine leukemia virus proviral sequence associated with the virus resistance gene Rmcf of DBA/2 mice. AB - The DBA/2 mouse Rmcf gene is responsible for in vivo and in vitro resistance to infection by the polytropic mink cell focus-forming (MCF) virus subgroup of murine leukemia viruses (MLVs). Previous studies suggested that Rmcf resistance is mediated by expression of an interfering MCF MLV envelope (Env) gene. To characterize this env gene, we examined resistance in crosses between Rmcf(r) DBA/2 mice and Mus castaneus, a species that lacks endogenous MCF env sequences. In backcross progeny, inheritance of Rmcf resistance correlated with inheritance of a specific endogenous MCF virus env-containing 4.6-kb EcoRI fragment. This fragment was present in the DBA/2N substrain with Rmcf-mediated resistance but not in virus-susceptible DBA/2J substrain mice. This fragment contains a provirus with a 5' long terminal repeat and the 5' half of env; the gag and pol genes have been partially deleted. The Env sequence is identical to that of a highly immunogenic viral glycoprotein expressed in the DBA/2 cell line L5178Y and closely resembles the env genes of modified polytropic proviruses. The coding sequence for the full-length Rmcf Env surface subunit was amplified from DNAs from virus-resistant backcross mice and was cloned into an expression vector. NIH 3T3 and BALB 3T3 cells stably transfected with this construct showed significant resistance to infection by MCF MLV but not by amphotropic MLV. This study identifies an Rmcf-linked MCF provirus and indicates that, like the ecotropic virus resistance gene Fv4, Rmcf may mediate resistance through an interference mechanism. PMID- 12134028 TI - Studies of the mechanism of transactivation of the adeno-associated virus p19 promoter by Rep protein. AB - During adeno-associated virus (AAV) type 2 productive infections, the p19 promoter of AAV is activated by the AAV Rep78 and Rep68 proteins. Rep-induced activation of p19 depends on the presence of one of several redundant Rep binding elements (RBEs) within the p5 promoter or within the terminal repeats (TR). In the absence of the TR, the p5 RBE and the p19 Sp1 site at position -50 are essential for p19 transactivation. To determine how a Rep complex bound at p5 induces transcription at p19, we made a series of p19 promoter chloramphenicol acetyltransferase constructs in which the p5 RBE was inserted at different locations upstream or downstream of the p19 mRNA start site. The RBE acted like a repressor element at most positions in the presence of both Rep and adenovirus (Ad), and the level of repression increased dramatically as the RBE was inserted closer to the p19 promoter. We concluded that the RBE by itself was not a conventional upstream activation signal and instead behaved like a repressor. To understand how the Rep-RBE complex within p5 activated p19, we considered the possibility that its role was to function as an architectural protein whose purpose was to bring other p5 transcriptional elements to the p19 promoter. In order to address this possibility, we replaced both the p5 RBE and the p19 Sp1 site with GAL4 binding sites. The modified GAL4-containing constructs were cotransfected with plasmids that expressed GAL4 fusion proteins capable of interacting through p53 and T-antigen (T-ag) protein domains. In the presence of Ad and the GAL4 fusion proteins, the p19 promoter exhibited strong transcriptional activation that was dependent on both the GAL4 fusion proteins and Ad infection. This suggested that the primary role of the p5 RBE and the p19 Sp1 sites was to act as a scaffold for bringing transcription complexes in the p5 promoter into close proximity with the p19 promoter. Since Rep and Sp1 themselves were not essential for transactivation, we tested mutants within the other p5 transcriptional elements in the context of GAL4-induced looping to determine which of the other p5 elements was necessary for p19 induction. Mutation of the p5 major late-transcription factor site reduced p19 activity but did not eliminate induction in the presence of the GAL4 fusion proteins. However, mutation of the p5 YY1 site at position -60 (YY1-60) eliminated GAL4-induced transactivation. This implicated the YY1-60 protein complexes in p19 induction by Rep. In addition, both basal p19 activity and activity in the presence of Ad increased when the YY1-60 site was mutated even in the absence of Rep or GAL4 fusion proteins. Therefore, there are likely to be alternative p5-p19 interactions that are Rep independent in which the YY1-60 complex inhibits p19 transcription. We concluded that transcriptional control of the p19 promoter was dependent on the formation of complexes between the p5 and p19 promoters and that activation of the p19 promoter depends largely on the ability of Rep and Sp1 to form a scaffold that positions the p5 YY1 complex near the p19 promoter. PMID- 12134029 TI - Inhibition of chemokine expression by adenovirus early region three (E3) genes. AB - Adenoviruses (Ad) have a variety of immunoregulatory genes, many of which are clustered in a 3.5-kb segment of DNA known as early region 3 (E3). Ad E3 codes for proteins that downregulate surface expression of class I major histocompatibility antigens and also inhibit tumor necrosis factor alpha (TNF alpha)- and Fas-induced cytolysis. We were interested in determining whether chemokine production or activity might also be inhibited by Ad E3 and we have studied this function in a human astrocytoma cell line, U373. Astrocytes constitute a part of the blood-brain barrier, and chemokines (IP-10, IL-8, MCP-1 4, and MIPs) expressed by them may contribute to leukocyte infiltration within the brain during inflammation. When U373 cells are activated by the proinflammatory molecule TNF-alpha, the increase in chemokine MCP-1, IL-8, and IP 10 transcripts is blocked by a recombinant Ad expressing the E3 genes under cytomegalovirus promoter control. Comparable Ads expressing green fluorescent protein in place of E3 have no effect on these chemokines. Ads also have been extensively studied as gene therapy vectors and most have a deletion of the E3 region to permit the insertion of larger fragments of foreign DNA. Our results suggest that construction of Ad vectors to include E3 expression cassettes will improve the efficacy and safety of such viral-based gene therapy protocols. PMID- 12134030 TI - The long noncoding region of the human parainfluenza virus type 1 f gene contributes to the read-through transcription at the m-f gene junction. AB - Sendai virus (SV) and human parainfluenza virus type 1 (hPIV1) have genomes consisting of nonsegmented negative-sense RNA in which the six genes are separated by well-conserved intergenic (IG) sequences and transcriptional start (S) and end signals. In hPIV1-infected cells, transcriptional termination at the M-F gene junction is ineffective; a large number of M-F read-through transcripts are produced (T. Bousse, T. Takimoto, K. G. Murti, and A. Portner, Virology 232:44-52, 1997). In contrast, few M-F read-through transcripts are detected in SV-infected cells. Sequence analysis indicated that the hPIV1 IG and S sequences in the M-F junction differ from those of SV. Furthermore, the hPIV1 F gene contains an unusually long noncoding sequence. To identify the cis-acting elements that prevent transcriptional termination at the M-F junction, we rescued recombinant SV (rSVhMFjCG) in which its M-F gene junction was replaced by that of hPIV1. Cells infected with rSVhMFjCG produced an abundance of M-F read-through transcripts; this result indicated that the hPIV1 M-F junction is responsible for inefficient termination. When one or both of the IG and S sites in rSVhMFjCG were replaced by those of SV, the efficiency of transcriptional termination increased but not to the level observed in wild-type SV-infected cells. Deletion of most of the long noncoding region of the hPIV1 F gene in rSVhMFjCG in addition to the mutations in IG and S signals resulted in efficient termination that was equivalent to the level observed in wild-type virus-infected cells. Therefore, the long noncoding sequence of the hPIV1 F gene contains cis-acting element(s) that affects transcriptional termination. Our evaluation of the effect of inefficient transcriptional termination on viral replication in culture revealed that cells infected with rSVhMFjCG produced less F protein than cells infected with wild-type SV and that assembly of the recombinant SV in culture was less efficient. These phenotypes seem to be responsible for the extended survival of mice infected with rSVhMFjCG. PMID- 12134031 TI - Transcriptional regulation of the interleukin-6 gene of human herpesvirus 8 (Kaposi's sarcoma-associated herpesvirus). AB - Human herpesvirus 8 (HHV-8; Kaposi's sarcoma-associated herpesvirus is linked to Kaposi's sarcoma (KS), primary effusion lymphoma (PEL), and multicentric Castleman's disease (MCD), all of which are viewed as cytokine-driven malignancies. In particular, interleukin-6 (IL-6) has been found to promote the growth and proliferation of cells from KS and PEL. HHV-8 encodes a homologue of IL-6 (viral IL-6 [vIL-6]), which functions similarly to the cellular IL-6. Therefore, vIL-6 has been proposed to play an important role in tumor progression. Several groups have reported that vIL-6 is expressed from the HHV-8 genome at higher levels in PEL and MCD lesions than in KS lesions. However, it is not clear how vIL-6 expression is regulated. We characterized the transcription at the vIL-6 gene locus by Northern blot analysis and, in contrast to previous reports, we observed two distinct transcripts from induced PEL cell lines. This observation was confirmed by primer extension, as well as 5' and 3' rapid amplification of cDNA ends. Two transcription initiation sites and putative TATA boxes were mapped. A luciferase reporter system was used to show that each of the two putative TATA boxes contributed to vIL-6 promoter activity. Since virally encoded transcriptional activator Rta potently activates the viral lytic gene expression cascade, we examined the role of Rta in controlling vIL-6 gene expression and found that Rta activated the vIL-6 promoter. The Rta-responsive element was further mapped through a series of deletion constructs. Electrophoretic mobility shift assays demonstrated that Rta binds directly to the vIL-6 Rta-responsive element, and the core Rta-responsive element was mapped to a 26-bp region spanning from nucleotide 18315 to 18290 on the viral genome. We propose that the existence of two vIL-6 promoters offers opportunities for differential regulation of vIL-6 gene expression in different tissue types and may account for the variable vIL-6 levels observed in KS, PEL, and MCD. PMID- 12134032 TI - Binding of human cytomegalovirus US2 to major histocompatibility complex class I and II proteins is not sufficient for their degradation. AB - Human cytomegalovirus (HCMV) glycoprotein US2 causes degradation of major histocompatibility complex (MHC) class I heavy-chain (HC), class II DR-alpha and DM-alpha proteins, and HFE, a nonclassical MHC protein. In US2-expressing cells, MHC proteins present in the endoplasmic reticulum (ER) are degraded by cytosolic proteasomes. It appears that US2 binding triggers a normal cellular pathway by which misfolded or aberrant proteins are translocated from the ER to cytoplasmic proteasomes. To better understand how US2 binds MHC proteins and causes their degradation, we constructed a panel of US2 mutants. Mutants truncated from the N terminus as far as residue 40 or from the C terminus to amino acid 140 could bind to class I and class II proteins. Nevertheless, mutants lacking just the cytosolic tail (residues 187 to 199) were unable to cause degradation of both class I and II proteins. Chimeric proteins were constructed in which US2 sequences were replaced with homologous sequences from US3, an HCMV glycoprotein that can also bind to class I and II proteins. One of these US2/US3 chimeras bound to class II but not to class I, and a second bound class I HC better than wild-type US2. Therefore, US2 residues involved in the binding to MHC class I differ subtly from those involved in binding to class II proteins. Moreover, our results demonstrate that the binding of US2 to class I and II proteins is not sufficient to cause degradation of MHC proteins. The cytosolic tail of US2 and certain US2 lumenal sequences, which are not involved in binding to MHC proteins, are required for degradation. Our results are consistent with the hypothesis that US2 couples MHC proteins to components of the ER degradation pathway, enormously increasing the rate of degradation of MHC proteins. PMID- 12134033 TI - Favorable and unfavorable HLA class I alleles and haplotypes in Zambians predominantly infected with clade C human immunodeficiency virus type 1. AB - The setpoint of viral RNA concentration (viral load [VL]) during chronic human immunodeficiency virus type 1 (HIV-1) infection reflects a virus-host equilibration closely related to CD8(+) cytotoxic T-lymphocyte (CTL) responses, which rely heavily on antigen presentation by the human major histocompatibility complex (MHC) (i.e., HLA) class I molecules. Differences in HIV-1 VL among 259 mostly clade C virus-infected individuals (137 females and 122 males) in the Zambia-UAB HIV Research Project (ZUHRP) were associated with several HLA class I alleles and haplotypes. In particular, general linear model analyses revealed lower log(10) VL among those with HLA allele B*57 (P = 0.002 [without correction]) previously implicated in favorable response and in those with HLA B*39 and A*30-Cw*03 (P = 0.002 to 0.016); the same analyses also demonstrated higher log(10) VL among individuals with A*02-Cw*16, A*23-B*14, and A*23-Cw*07 (P = 0.010 to 0.033). These HLA effects remained strong (P = 0.0002 to 0.075) after adjustment for age, gender, and duration of infection and persisted across three orders of VL categories (P = 0.001 to 0.084). In contrast, neither B*35 (n = 15) nor B*53 (n = 53) showed a clear disadvantage such as that reported elsewhere for these closely related alleles. Other HLA associations with unusually high (A*68, B*41, B*45, and Cw*16) or low (B*13, Cw*12, and Cw*18) VL were either unstable or reflected their tight linkage respecting disequilibria with other class I variants. The three consistently favorable HLA class I variants retained in multivariable models and in alternative analyses were present in 30.9% of subjects with the lowest (<10,000 copies per ml) and 3.1% of those with the highest (>100,000) VL. Clear differential distribution of HLA profiles according to level of viremia suggests important host genetic contribution to the pattern of immune control and escape during HIV-1 infection. PMID- 12134034 TI - Mammalian reovirus nonstructural protein microNS forms large inclusions and colocalizes with reovirus microtubule-associated protein micro2 in transfected cells. AB - Cells infected with mammalian orthoreoviruses contain large cytoplasmic phase dense inclusions believed to be the sites of viral replication and assembly, but the morphogenesis, structure, and specific functions of these "viral factories" are poorly understood. Using immunofluorescence microscopy, we found that reovirus nonstructural protein microNS expressed in transfected cells forms inclusions that resemble the globular viral factories formed in cells infected with reovirus strain type 3 Dearing from our laboratory (T3D(N)). In the transfected cells, the formation of microNS large globular perinuclear inclusions was dependent on the microtubule network, as demonstrated by the appearance of many smaller microNS globular inclusions dispersed throughout the cytoplasm after treatment with the microtubule-depolymerizing drug nocodazole. Coexpression of microNS and reovirus protein micro2 from a different strain, type 1 Lang (T1L), which forms filamentous viral factories, altered the distributions of both proteins. In cotransfected cells, the two proteins colocalized in thick filamentous structures. After nocodazole treatment, many small dispersed globular inclusions containing microNS and micro2 were seen, demonstrating that the microtubule network is required for the formation of the filamentous structures. When coexpressed, the micro2 protein from T3D(N) also colocalized with microNS, but in globular inclusions rather than filamentous structures. The morphology difference between the globular inclusions containing microNS and micro2 protein from T3D(N) and the filamentous structures containing microNS and micro2 protein from T1L in cotransfected cells mimicked the morphology difference between globular and filamentous factories in reovirus-infected cells, which is determined by the micro2-encoding M1 genome segment. We found that the first 40 amino acids of microNS are required for colocalization with micro2 but not for inclusion formation. Similarly, a fusion of microNS amino acids 1 to 41 to green fluorescent protein was sufficient for colocalization with the micro2 protein from T1L but not for inclusion formation. These observations suggest a functional difference between microNS and microNSC, a smaller form of the protein that is present in infected cells and that is missing amino acids from the amino terminus of microNS. The capacity of microNS to form inclusions and to colocalize with micro2 in transfected cells suggests a key role for microNS in forming viral factories in reovirus-infected cells. PMID- 12134035 TI - Characterization of a novel simian immunodeficiency virus with a vpu gene from greater spot-nosed monkeys (Cercopithecus nictitans) provides new insights into simian/human immunodeficiency virus phylogeny. AB - In the present study, we describe a new simian immunodeficiency virus (SIV), designated SIVgsn, naturally infecting greater spot-nosed monkeys (Cercopithecus nictitans) in Cameroon. Together with SIVsyk, SIVgsn represents the second virus isolated from a monkey belonging to the Cercopithecus mitis group of the Cercopithecus genus. Full-length genome sequence analysis of two SIVgsn strains, SIVgsn-99CM71 and SIVgsn-99CM166, revealed that despite the close phylogenetic relationship of their hosts, SIVgsn was highly divergent from SIVsyk. First of all, they differ in their genomic organization. SIVgsn codes for a vpu homologue, so far a unique feature of the members of the SIVcpz/human immunodeficiency virus type 1 (HIV-1) lineage, and detailed phylogenetic analyses of various regions of the viral genome indicated that SIVgsn might be a mosaic of sequences with different evolutionary histories. SIVgsn was related to SIVsyk in Gag and part of Pol and related to SIVcpz in Env, and the middle part of the genome did not cluster significantly with any of the known SIV lineages. When comparing the two SIVgsn Env sequences with that of SIVcpz, a remarkable conservation was seen in the V3 loop, indicating a possible common origin for the envelopes of these two viruses. The habitats of the two subspecies of chimpanzees infected by SIVcpz overlap the geographic ranges of greater spot-nosed monkeys and other monkey species, allowing cross-species transmission and recombination between coinfecting viruses. The complex genomic structure of SIVgsn, the presence of a vpu gene, and its relatedness to SIVcpz in the envelope suggest a link between SIVgsn and SIVcpz and provide new insights about the origin of SIVcpz in chimpanzees. PMID- 12134036 TI - In vivo egress of an alphaherpesvirus from axons. AB - Many alphaherpesviruses establish a latent infection in the peripheral nervous systems of their hosts. This life cycle requires the virus to move long distances in axons toward the neuron's cell body during infection and away from the cell body during reactivation. While the events underlying entry of the virion into neurons during infection are understood in principle, no such consensus exists regarding viral egress from neurons after reactivation. In this study, we challenged two different models of viral egress from neurons by using pseudorabies virus (PRV) infection of the rat retina: does PRV egress solely from axon terminals, or can the virus egress from axon shafts as well as axon terminals? We took advantage of PRV gD mutants that are not infectious as extracellular particles but are capable of spreading by cell-cell contact. We observed that both wild-type virus and a PRV gD null mutant are capable of spreading from axons to closely apposed nonneuronal cells within the rat optic nerve after intravitreal infection. However, infection does not spread from these infected nonneuronal cells. We suggest that viral egress can occur sporadically along the length of infected axons and is not confined solely to axon terminals. Moreover, it is likely that extracellular particles are not involved in nonneuronal cell infections. Taking these together with previous data, we suggest a model of viral egress from neurons that unifies previous apparently contradictory data. PMID- 12134037 TI - The block in assembly of modified vaccinia virus Ankara in HeLa cells reveals new insights into vaccinia virus morphogenesis. AB - It has previously been shown that upon infection of HeLa cells with modified vaccinia virus Ankara (MVA), assembly is blocked at a late stage of infection and immature virions (IVs) accumulate (G. Sutter and B. Moss, Proc. Natl. Acad. Sci. USA 89:10847-10851, 1992). In the present study the morphogenesis of MVA in HeLa cells was studied in more detail and compared to that under two conditions that permit the production of infectious particles: infection of HeLa cells with the WR strain of vaccinia virus (VV) and infection of BHK cells with MVA. Using several quantitative and qualitative assays, we show that early in infection, MVA in HeLa cells behaves in a manner identical to that under the permissive conditions. By immunofluorescence microscopy (IF) at late times of infection, the labelings for an abundant membrane protein of the intracellular mature virus, p16/A14L, and the viral DNA colocalize under permissive conditions, whereas in HeLa cells infected with MVA these two structures do not colocalize to the same extent. In both permissive and nonpermissive infection, p16-labeled IVs first appear at 5 h postinfection. In HeLa cells infected with MVA, IVs accumulated predominantly outside the DNA regions, whereas under permissive conditions they were associated with the viral DNA. At 4 h 30 min, the earliest time at which p16 is detected, the p16 labeling was found predominantly in a small number of distinct puncta by IF, which were distinct from the sites of DNA in both permissive and nonpermissive infection. By electron microscopy, no crescents or IVs were found at this time, and the p16-labeled structures were found to consist of membrane-rich vesicles that were in continuity with the cellular endoplasmic reticulum. Over the next 30 min of infection, a large number of p16-labeled crescents and IVs appeared abruptly under both permissive and nonpermissive conditions. Under permissive conditions, these IVs were in close association with the sites of DNA, and a significant amount of these IVs engulfed the viral DNA. In contrast, under nonpermissive conditions, the IVs and DNA were mostly in separate locations and relatively few IVs acquired DNA. Our data show that in HeLa cells MVA forms normal DNA replication sites and normal viral precursor membranes but the transport between these two structures is inhibited. PMID- 12134038 TI - Critical role for protein tyrosine phosphatase SHP-1 in controlling infection of central nervous system glia and demyelination by Theiler's murine encephalomyelitis virus. AB - We previously characterized the expression and function of the protein tyrosine phosphatase SHP-1 in the glia of the central nervous system (CNS). In the present study, we describe the role of SHP-1 in virus infection of glia and virus-induced demyelination in the CNS. For in vivo studies, SHP-1-deficient mice and their normal littermates received an intracerebral inoculation of an attenuated strain of Theiler's murine encephalomyelitis virus (TMEV). At various times after infection, virus replication, TMEV antigen expression, and demyelination were monitored. It was found that the CNS of SHP-1-deficient mice uniquely displayed demyelination and contained substantially higher levels of virus than did that of normal littermate mice. Many infected astrocytes and oligodendrocytes were detected in both brains and spinal cords of SHP-1-deficient but not normal littermate mice, showing that the virus replicated and spread at a much higher rate in the glia of SHP-1-deficient animals. To ascertain whether the lack of SHP 1 in the glia was primarily responsible for these differences, glial samples from these mice were cultured in vitro and infected with TMEV. As in vivo, infected astrocytes and oligodendrocytes of SHP-1-deficient mice were much more numerous and produced more virus than did those of normal littermate mice. These findings indicate that SHP-1 is a critical factor in controlling virus replication in the CNS glia and virus-induced demyelination. PMID- 12134040 TI - Mutations of the RNase H C helix of the Moloney murine leukemia virus reverse transcriptase reveal defects in polypurine tract recognition. AB - Both the RNase H domain of Moloney murine leukemia virus (Mo-MLV) reverse transcriptase (RT) and Escherichia coli RNase H possess a positively charged alpha-helix (C helix) and a loop that are not present in the RNase H domains of human immunodeficiency virus (HIV) RT or avian sarcoma virus RT. Although a mutant Mo-MLV RT lacking the C helix (DeltaC RT) retains DNA polymerase activity on homopolymeric substrates and partial RNase H activity, reverse transcription of the viral RNA genome in vivo is defective. To identify the essential features of the C helix, a panel of Mo-MLV RT mutants was generated. Analyses of these mutant viruses revealed the importance of residues H594, I597, R601, and G602. The mutants were tested for their ability to synthesize viral DNA after acute infections and to form proper 5' and 3' viral DNA ends. The mutant RTs were tested in vitro for exogenous RT activity, minus-strand strong-stop DNA synthesis in endogenous RT reactions, nonspecific RNase H activity, and finally, proper cleavage at the polypurine tract-U3 junction. The R601A mutant was the most defective mutant both in vivo and in vitro and possessed very little RNase H activity. The H594A, I597A, and G602A mutants had significant reductions in RNase H activity and in their rates of viral replication. Many of the mutants formed improper viral DNA ends and were less efficient in PPT-U3 recognition and cleavage in vitro. The data show that the C helix plays a crucial role for overall RNase H cleavage activity. The data also suggest that the C helix may play an important role in polypurine tract recognition and proper formation of the plus-strand DNA's 5' end. PMID- 12134039 TI - Elevated generation of reactive oxygen/nitrogen species in hantavirus cardiopulmonary syndrome. AB - Hantavirus cardiopulmonary syndrome (HCPS) is a life-threatening respiratory disease characterized by profound pulmonary edema and myocardial depression. Most cases of HCPS in North America are caused by Sin Nombre virus (SNV), which is carried asymptomatically by deer mice (Peromyscus maniculatus). The underlying pathophysiology of HCPS is poorly understood. We hypothesized that pathogenic SNV infection results in increased generation of reactive oxygen/nitrogen species (RONS), which contribute to the morbidity and mortality of HCPS. Human disease following infection with SNV or Andes virus was associated with increased nitrotyrosine (NT) adduct formation in the lungs, heart, and plasma and increased expression of inducible nitric oxide synthase (iNOS) in the lungs compared to the results obtained for normal human volunteers. In contrast, NT formation was not increased in the lungs or cardiac tissue from SNV-infected deer mice, even at the time of peak viral antigen expression. In a murine (Mus musculus) model of HCPS (infection of NZB/BLNJ mice with lymphocytic choriomeningitis virus clone 13), HCPS-like disease was associated with elevated expression of iNOS in the lungs and NT formation in plasma, cardiac tissue, and the lungs. In this model, intraperitoneal injection of 1400W, a specific iNOS inhibitor, every 12 h during infection significantly improved survival without affecting intrapulmonary fluid accumulation or viral replication, suggesting that cardiac damage may instead be the cause of mortality. These data indicate that elevated production of RONS is a feature of pathogenic New World hantavirus infection and that pharmacologic blockade of iNOS activity may be of therapeutic benefit in HCPS cases, possibly by ameliorating the myocardial suppressant effects of RONS. PMID- 12134041 TI - Direct participation of Sam68, the 68-kilodalton Src-associated protein in mitosis, in the CRM1-mediated Rev nuclear export pathway. AB - Human immunodeficiency virus type 1 (HIV-1) replication requires efficient nuclear export of incompletely spliced and unspliced HIV-1 mRNA transcripts, which is achieved by Rev expression at an early stage of the viral life cycle. We have recently shown that expression of Sam68, the 68-kDa Src-associated protein in mitosis, is able to alleviate Rev function block in astrocytes by promoting Rev nuclear export. In the present study, we utilized an antisense RNA expression strategy to down-modulate constitutive Sam68 expression and examined its effect on Rev function, HIV-1 gene expression, and viral replication. These results showed that down-modulation of constitutive Sam68 expression markedly inhibited HIV-1 production in 293T cells and viral replication in T lymphocytes such as Jurkat and CEM cells, as well as human peripheral blood mononuclear cells (PBMCs). Rev-dependent in trans complementation and reporter gene assays further demonstrated that inhibition of HIV-1 gene expression by Sam68 down-modulation was due to impeded Rev activity. Moreover, digital fluorescence microscopic imaging revealed that down-modulation of Sam68 expression caused exclusive nuclear retention and colocalization of both Rev and CRM1. Taken together, these data suggest that adequate Sam68 expression is required for Rev function and, thereby, for HIV-1 gene expression and viral replication, and they support the notion that Sam68 is directly involved in the CRM1-mediated Rev nuclear export pathway. PMID- 12134043 TI - Heparan sulfate mediates infection of high-neurovirulence Theiler's viruses. AB - The mechanisms by which Theiler's murine encephalomyelitis virus (TMEV) binds and enters host cells and the molecules involved are not completely understood. In this study, we demonstrate that the high-neurovirulence TMEV GDVII virus uses the glycosaminoglycan heparan sulfate (HS) as an attachment factor that is required for efficient infection. Studies based on soluble HS-mediated inhibition of attachment and infection, removal of HS with specific enzymes, and blocking with anti-HS antibodies establish that HS mediates GDVII virus entry into mammalian cells. Data from defined proteoglycan-deficient Chinese hamster ovary mutant cells further support the role of HS in GDVII infection and indicate that the extent of sulfation is critical for infection. Neuraminidase treatment of proteoglycan-deficient cells restores permissiveness to GDVII virus, indicating that sialic acid hinders direct access of virus to the protein entry receptor. A model of the potential steps in GDVII virus entry into mammalian cells involving HS is proposed. PMID- 12134042 TI - Kaposi's sarcoma-associated herpesvirus-induced upregulation of the c-kit proto oncogene, as identified by gene expression profiling, is essential for the transformation of endothelial cells. AB - Kaposi's sarcoma (KS), the most frequent malignancy afflicting AIDS patients, is characterized by spindle cell formation and vascularization. Infection with KS associated herpesvirus (KSHV) is consistently observed in all forms of KS. Spindle cell formation can be replicated in vitro by infection of dermal microvascular endothelial cells (DMVEC) with KSHV. To study the molecular mechanism of this transformation, we compared RNA expression profiles of KSHV infected and mock-infected DMVEC. Induction of several proto-oncogenes was observed, particularly the receptor tyrosine kinase c-kit. Consistent with increased c-Kit expression, KHSV-infected DMVEC displayed enhanced proliferation in response to the c-Kit ligand, stem cell factor (SCF). Inhibition of c-Kit activity with either a pharmacological inhibitor of c-Kit (STI 571) or a dominant negative c-Kit protein reversed SCF-dependent proliferation. Importantly, inhibition of c-Kit signal transduction reversed the KSHV-induced morphological transformation of DMVEC. Furthermore, overexpression studies showed that c-Kit was sufficient to induce spindle cell formation. Together, these data demonstrate an essential role for c-Kit in KS tumorigenesis and reveal a target for pharmacological intervention. PMID- 12134044 TI - Dengue virus selectively induces human mast cell chemokine production. AB - Severe dengue virus infections usually occur in individuals who have preexisting anti-dengue virus antibodies. Mast cells are known to play an important role in host defense against several pathogens, but their role in viral infection has not yet been elucidated. The effects of dengue virus infection on the production of chemokines by human mast cells were examined. Elevated levels of secreted RANTES, MIP-1alpha, and MIP-1beta, but not IL-8 or ENA-78, were observed following infection of KU812 or HMC-1 human mast cell-basophil lines. In some cases a >200 fold increase in RANTES production was observed. Cord blood-derived cultured human mast cells treated with dengue virus in the presence of subneutralizing concentrations of dengue virus-specific antibody also demonstrated significantly (P < 0.05) increased RANTES production, under conditions which did not induce significant degranulation. Chemokine responses were not observed when mast cells were treated with UV-inactivated dengue virus in the presence or absence of human dengue virus-specific antibody. Neither antibody-enhanced dengue virus infection of the highly permissive U937 monocytic cell line nor adenovirus infection of mast cells induced a RANTES, MIP-1alpha, or MIP-1beta response, demonstrating a selective mast cell response to dengue virus. These results suggest a role for mast cells in the initiation of chemokine-dependent host responses to dengue virus infection. PMID- 12134045 TI - Simian virus 40 large T antigen and two independent T-antigen segments sensitize cells to apoptosis following genotoxic damage. AB - The simian virus 40 (SV40) large tumor (T) antigen is sufficient to transform cells in cultures and induce tumors in experimental animals. Transformation of primary cells in cultures requires both overcoming growth arrest by stimulating the cell cycle and blocking cell death activities presumably activated by oncogene-mediated hyperproliferation signals. The study presented here examined the ability of specific regions and activities of T antigen to modulate apoptosis in cells treated with the genotoxic agent 5-fluorouracil (5-FU). The results showed that the expression of full-length T antigen rendered rat embryo fibroblasts (REF) sensitive to 5-FU-induced apoptosis. Thus, neither the p53 binding region nor the Bcl-2 homology region of T antigen was sufficient to prevent cell death induced by the DNA-damaging agent. T-antigen-mediated sensitization occurred independently of retinoblastoma protein or p53 and p300 binding. An N-terminal segment containing the first 127 T-antigen amino acids (T1 127) was sufficient to sensitize cells. A C-terminal segment consisting of T antigen amino acids 251 to 708 (T251-708) also sensitized cells to 5-FU-induced apoptosis. This sensitization did not occur when T251-708 was targeted to the nucleus by inclusion of the SV40 nuclear localization signal. The introduction of mutations into the T-antigen J domain resulted in mutation-specific and variable inhibition of apoptosis. This result suggested that either the structural or the functional integrity of the J domain is required to sensitize cells to apoptosis. Treatment of REF or REF expressing full-length T antigen, an N-terminal segment, or T251-708 resulted in increased expression of the p53-responsive MDM2 gene; apoptosis occurred through a p53-dependent pathway, as p53-null cells expressing these T antigens were resistant to 5-FU-induced apoptosis. Possible mechanisms involved in sensitizing cells to a p53-dependent apoptosis pathway in spite of the ability of T antigen to bind and inactivate the transcriptional transactivating activity of p53 are discussed. PMID- 12134047 TI - Immunological aspects of recombinant adeno-associated virus delivery to the mammalian brain. AB - Recombinant adeno-associated viruses (rAAV) are highly efficient vectors for gene delivery into the central nervous system (CNS). However, host inflammatory and immune responses may play a critical role in limiting the use of rAAV vectors for gene therapy and functional genomic studies in vivo. Here, we evaluated the effect of repeated injections of five rAAV vectors expressing different genetic sequences (coding or noncoding) in a range of combinations into the rat brain. Specifically, we wished to determine whether a specific immune or inflammatory response appeared in response to the vector and/or the transgene protein after repeated injections under conditions of mannitol coinjection. We show that readministration of the same rAAV to the CNS is possible if the interval between the first and second injection is more than 4 weeks. Furthermore, our data demonstrate that rAAV vectors carrying different genetic sequences can be administered at intervals of 2 weeks. Our data therefore suggest that the AAV capsid structure is altered by the vector genetic sequence, such that secondary structures of the single-stranded genome have an impact on the antigenicity of the virus. This study provides guidelines for more rational design of gene transfer studies in the rodent brain and, in addition, suggests the use of repeated administration of rAAV as a viable form of therapy for the treatment of chronic diseases. PMID- 12134046 TI - The relationship between simian immunodeficiency virus RNA levels and the mRNA levels of alpha/beta interferons (IFN-alpha/beta) and IFN-alpha/beta-inducible Mx in lymphoid tissues of rhesus macaques during acute and chronic infection. AB - To define the role of alpha/beta interferons (IFN-alpha/beta) in simian immunodeficiency virus (SIV) infection, IFN-alpha and IFN-beta mRNA levels and mRNA levels of Mx, an antiviral effector molecule, were determined in lymphoid tissues of rhesus macaques infected with pathogenic SIV. IFN-alpha/beta responses were induced during the acute phase and persisted in various lymphoid tissues throughout the chronic phase of infection. IFN-alpha/beta responses were most consistent in tissues with high viral RNA levels; thus, IFN-alpha/beta responses were not generally associated with effective control of SIV replication. IFN alpha/beta responses were differentially regulated in different lymphoid tissues and at different stages of infection. The most consistent IFN-alpha/beta responses in acute and chronic SIV infection were observed in peripheral lymph nodes. In the spleen, only a transient increase in IFN-alpha/beta mRNA levels during acute SIV infection was observed. Further, IFN-alpha and IFN-beta mRNA levels showed a tissue-specific expression pattern during the chronic, but not the acute, phase of infection. In the acute phase of infection, SIV RNA levels in lymphoid tissues of rhesus macaques correlated with mRNA levels of both IFN-alpha and IFN-beta, whereas during chronic SIV infection only increased IFN-alpha mRNA levels correlated with the level of virus replication in the same tissues. In lymphoid tissues of all SIV-infected monkeys, higher viral RNA levels were associated with increased Mx mRNA levels. We found no evidence that monkeys with increased Mx mRNA levels in lymphoid tissues had enhanced control of virus replication. In fact, Mx mRNA levels were associated with high viral RNA levels in lymphoid tissues of chronically infected animals. PMID- 12134048 TI - Nef enhances human immunodeficiency virus type 1 infectivity and replication independently of viral coreceptor tropism. AB - We investigated the infectivities and replicative capacities of a large panel of variants of the molecular human immunodeficiency virus type 1 (HIV-1) NL4-3 clone that differ exclusively in the V3 region of the viral envelope glycoprotein and the nef gene. Our results demonstrate that Nef enhances virion infectivity and HIV-1 replication independently of the viral coreceptor tropism. PMID- 12134049 TI - Characterization of the nuclear localization signal of the borna disease virus polymerase. AB - Borna disease virus (BDV) is a nonsegmented negative-strand RNA virus that replicates and transcribes its genome in the nucleus of infected cells. BDV proteins involved in replication and transcription must pass through the nuclear envelope to associate with the genomic viral RNA. The RNA-dependent RNA polymerase (L) of BDV is postulated to be the catalytic enzyme of replication and transcription. We demonstrated previously that BDV L localizes to the nucleus of BDV-infected cells and L-transfected cells. Nuclear localization of the protein presupposes the presence of a nuclear localization signal (NLS) within its primary amino acid sequence or cotransport to the nucleus with another karyophilic protein. Because L localized to the nucleus in the absence of other viral proteins, we investigated the possibility that L contains an NLS. The minimal sequence required for nuclear localization of L was identified by analyzing the subcellular distribution of deletion mutants of L fused to a flag epitope tag or beta-galactosidase. Although the majority of the L fusion proteins localized to the cytoplasm of transfected BSR-T7 cells, a strong NLS (844RVVKLRIAP852) with basic and proline residues was identified. Mutation of this sequence resulted in cytoplasmic distribution of L, confirming that this sequence was necessary and sufficient to drive the nuclear localization of L. PMID- 12134050 TI - Glycoprotein I of varicella-zoster virus is required for viral replication in skin and T cells. AB - Varicella-zoster virus (VZV) glycoprotein I (gI) is dispensable in cell culture; the SCIDhu model of VZV pathogenesis was used to determine whether gI is necessary in vivo. The parental and repaired viruses grew in human skin and thymus/liver implants, but the gI deletion mutant was not infectious. Thus, gI is essential for VZV infectivity in skin and T cells. PMID- 12134051 TI - Molecular basis of the attenuation exhibited by molecularly cloned highly passaged chicken anemia virus isolates. AB - Chimeric virus experiments indicated that the pathogenicity and monoclonal antibody reactivity differences between two molecularly cloned, highly passaged chicken anemia virus isolates could be attributed to the VP1 amino acid change at residue 89. The introduction of this change into a pathogenic cloned low-passage isolate was not sufficient to cause attenuation. PMID- 12134052 TI - Inserting a nuclear targeting signal into a replication-competent Moloney murine leukemia virus affects viral export and is not sufficient for cell cycle independent infection. AB - The effects of inserting reported nuclear localization signals (NLSs) into the Moloney murine leukemia virus (Mo-MuLV) integrase (IN) protein, within a replication-competent viral construct, were studied. In contrast to the virus harboring IN fused to the simian virus 40 (SV40) large T antigen NLS (SV40 NLS) (J. A. Seamon, M. Adams, S. Sengupta, and M. J. Roth, Virology 274:412-419, 2000), a codon-modified SV40 NLS was stably expressed during viral propagation. Incorporation of the codon-modified SV40 NLS into IN, however, altered the packaging of the Gag-Pol precursor in the virus; viral particles contained decreased levels of reverse transcriptase (RT) and IN. In addition, the virus showed delayed kinetics of viral DNA synthesis upon infection. A panel of infectious MuLVs containing alternative IN-NLS fusions was generated and assayed for cell cycle-independent infection. Viral infection with the NLS-tagged proteins, however, remained dependent on passage of the cells through mitosis. This finding has direct implications for engineering murine-based retroviral vectors for gene therapy. PMID- 12134053 TI - Analysis of bovine leukemia virus gag membrane targeting and late domain function. AB - Assembly of retrovirus-like particles only requires the expression of the Gag polyprotein precursor. We have exploited this in the development of a model system for studying the virus particle assembly pathway for bovine leukemia virus (BLV). BLV is closely related to the human T-cell leukemia viruses (HTLVs), and all are members of the Deltaretrovirus genus of the Retroviridae family. Overexpression of a BLV Gag polyprotein containing a carboxy-terminal influenza virus hemagglutinin (HA) epitope tag in mammalian cells led to the robust production of virus-like particles (VLPs). Site-directed mutations were introduced into HA-tagged Gag to test the usefulness of this model system for studying certain aspects of the virus assembly pathway. First, mutations that disrupted the amino-terminal glycine residue that is important for Gag myristylation led to a drastic reduction in VLP production. Predictably, the nature of the VLP production defect was correlated to Gag membrane localization. Second, mutation of the PPPY motif (located in the MA domain) greatly reduced VLP production in the absence of the viral protease. This reduction in VLP production was more severe in the presence of an active viral protease. Examination of particles by electron microscopy revealed an abundance of particles that began to pinch off from the plasma membrane but were not completely released from the cell surface, indicating that the PPPY motif functions as a late domain (L domain). PMID- 12134054 TI - Recovery of virulent and RNase-negative attenuated type 2 bovine viral diarrhea viruses from infectious cDNA clones. AB - Cloned cDNA derived from the genome of the virulent type 2 bovine viral diarrhea virus (BVDV) strain NY'93/C was sequenced and served for establishment of the infectious cDNA clone pKANE40A. Virus recovered from pKANE40A exhibited growth characteristics similar to those of wild-type BVDV NY'93/C and proved to be clinically indistinguishable from the wild-type virus in animal experiments. A virus mutant in which the RNase residing in the viral glycoprotein E(rns) was inactivated, revealed an attenuated phenotype. The plasmid pKANE40A represents the first infectious cDNA clone established for a type 2 BVDV and offers a variety of new approaches to analyze the mechanisms of BVDV-induced disease in cattle. PMID- 12134056 TI - Cerebral energetics and spiking frequency: the neurophysiological basis of fMRI. AB - Functional MRI (fMRI) is widely assumed to measure neuronal activity, but no satisfactory mechanism for this linkage has been identified. Here we derived the changes in the energetic component from the blood oxygenation level-dependent (BOLD) fMRI signal and related it to changes in the neuronal spiking frequency in the activated voxels. Extracellular recordings were used to measure changes in cerebral spiking frequency (Deltanu/nu) of a neuronal ensemble during forepaw stimulation in the alpha-chloralose anesthetized rat. Under the same conditions localized changes in brain energy metabolism (DeltaCMR(O2)/CMR(O2)) were obtained from BOLD fMRI data in conjunction with measured changes in cerebral blood flow (DeltaCBF/CBF), cerebral blood volume (DeltaCBV/CBV), and transverse relaxation rates of tissue water (T(2)(*) and T(2)) by MRI methods at 7T. On stimulation from two different depths of anesthesia DeltaCMR(O2)/CMR(O2) approximately Deltanu/nu. Previous (13)C magnetic resonance spectroscopy studies, under similar conditions, had shown that DeltaCMR(O2)/CMR(O2) was proportional to changes in glutamatergic neurotransmitter flux (DeltaV(cyc)/V(cyc)). These combined results show that DeltaCMR(O2)/CMR(O2) approximately DeltaV(cyc)/V(cyc) approximately Deltanu/nu, thereby relating the energetic basis of brain activity to neuronal spiking frequency and neurotransmitter flux. Because DeltaCMR(O2)/CMR(O2) had the same high spatial and temporal resolutions of the fMRI signal, these results show how BOLD imaging, when converted to DeltaCMR(O2)/CMR(O2), responds to localized changes in neuronal spike frequency. PMID- 12134057 TI - Total neuroenergetics support localized brain activity: implications for the interpretation of fMRI. AB - In alpha-chloralose-anesthetized rats, changes in the blood oxygenation level dependent (BOLD) functional MRI (fMRI) signal (DeltaS/S), and the relative spiking frequency of a neuronal ensemble (Deltanu/nu) were measured in the somatosensory cortex during forepaw stimulation from two different baselines. Changes in cerebral oxygen consumption (DeltaCMR(O2)/CMR(O2)) were derived from the BOLD signal (at 7T) by independent determinations in cerebral blood flow (DeltaCBF/CBF) and volume (DeltaCBV/CBV). The spiking frequency was measured by extracellular recordings in layer 4. Changes in all three parameters (CMR(O2), nu, and S) were greater from the lower baseline (i.e., deeper anesthesia). For both baselines, DeltaCMR(O2)/CMR(O2) and Deltanu/nu were approximately one order of magnitude larger than DeltaS/S. The final values of CMR(O2) and nu reached during stimulation were approximately the same from both baselines. If only increments were required to support functions then their magnitudes should be independent of the baseline. In contrast, if particular magnitudes of activity were required, then sizes of increments should inversely correlate with the baseline (being larger from a lower baseline). The results show that particular magnitudes of activity support neural function. The disregard of baseline activity in fMRI experiments by differencing removes a large and necessary component of the total activity. Implications of these results for understanding brain function and fMRI experiments are discussed. PMID- 12134058 TI - Patterned variation in murine MHC promoters. AB - To compare variation in regulatory and coding DNA, promoter sequences have been obtained from wild-derived mice and laboratory rats. The sequences are from the proximal promoter of the H2Aa, H2Ab, H2Eb, and H2K genes of 24 wild-derived inbred strains and a sample of the corresponding exon 2 sequences and of the RT1.Ba gene of six strains of laboratory rat. They reveal a high level of variation in the mouse MHC class II promoters (H2A and H2E), a low level in MHC class I (H2K), and none in the rat. The variation is pronounced in and around the cAMP response element, a major binding site for modulating promoter activity in response to external stimulation. This finding, together with the different levels of variation in MHC classes I and II, is suggestive of natural selection. However, selection operating via the MHC coding sequences must also contribute, as indicated by the minimal variation in both the MHC class II promoter and coding sequences of the rat. Furthermore CIITA (trans-activator of class II) of the mouse has been reported to have minimal variation in its promoter and none in its coding sequence. Taken together these data suggest that the regulatory and coding sequences undergo coselection. Each of the mouse class II promoters has a pattern of variation that appears to be basically dimorphic, with further variation added by recombination/mutation. The dimorphic allelic lineages are in marginally detectable linkage disequilibrium with the exon 2 sequences, particularly in H2Aa, thus lending further support to the coevolution hypothesis. PMID- 12134059 TI - Tissue-specific silencing of a transgene in rice. AB - In a transgenic rice line, a beta-glucuronidase reporter gene under the control of the rice tungro bacilliform virus promoter became gradually methylated, and gene activity was lost concomitantly. Methylation was observed only in the homozygous offspring and was initially restricted to the promoter region and accompanied by loss of expression in the vascular bundle tissue only. This expression pattern was similar to that of a promoter with a deletion of a vascular bundle expression element. The gene activity could be reestablished by treatment with 5-azacytidine. Methylation per se did not inhibit the binding to the promoter region of protein factors which also bound to the unmethylated sequence. Instead, promoter methylation enabled the alternative binding of a protein with specificity for sequence and methylation. In further generations of homozygous offspring the methylation spread into the transcribed region and gene activity was completely repressed also in nonvascular cells. The results indicate that different stages are involved in DNA methylation-correlated gene inactivation, and that at least one of them may involve the attraction of a sequence and methylation-specific DNA-binding protein. PMID- 12134060 TI - Targeted mutagenesis of the murine transferrin receptor-2 gene produces hemochromatosis. AB - Hereditary hemochromatosis (HH) is a common genetic disorder characterized by excess absorption of dietary iron and progressive iron deposition in several tissues, particularly liver. The vast majority of individuals with HH are homozygous for mutations in the HFE gene. Recently a second transferrin receptor (TFR2) was discovered, and a previously uncharacterized type of hemochromatosis (HH type 3) was identified in humans carrying mutations in the TFR2 gene. To characterize the role for TFR2 in iron homeostasis, we generated mice in which a premature stop codon (Y245X) was introduced by targeted mutagenesis in the murine Tfr2 coding sequence. This mutation is orthologous to the Y250X mutation identified in some patients with HH type 3. The homozygous Tfr2(Y245X) mutant mice showed profound abnormalities in parameters of iron homeostasis. Even on a standard diet, hepatic iron concentration was several-fold higher in the homozygous Tfr2(Y245X) mutant mice than in wild-type littermates by 4 weeks of age. The iron deposition in the mutant mice was predominantly hepatocellular and periportal. The mean splenic iron concentration in the homozygous Tfr2(Y245X) mutant mice was significantly less than that observed in the wild-type mice. The homozygous Tfr2(Y245X) mutant mice also demonstrated elevated transferrin saturations. There were no significant differences in parameters of erythrocyte production including hemoglobin levels, hematocrits, erythrocyte indices, and reticulocyte counts. Heterozygous Tfr2(Y245X) mice did not differ in any measured parameter from wild-type mice. This study confirms the important role for TFR2 in iron homeostasis and provides a tool for investigating the excess iron absorption and abnormal iron distribution in iron-overload disorders. PMID- 12134062 TI - Ordered nucleation and spreading of silenced chromatin in Saccharomyces cerevisiae. AB - In Saccharomyces cerevisiae, silencing at the HM loci depends on Sir proteins, which are structural components of silenced chromatin. To explore the structure and assembly of silenced chromatin, the associations of Sir proteins with sequences across the HMR locus were examined by chromatin immunoprecipitation. In wild-type cells, Sir2p, Sir3p, and Sir4p were spread throughout and coincident with the silenced region at HMR. Sir1p, in contrast, associated only with the HMR E silencer, consistent with its role in establishment but not maintenance of silencing. Sir4p was required for the association of other Sir proteins with silencers. In contrast, in the absence of Sir2p or Sir3p, partial assemblies of Sir proteins could form at silencers, where Sir protein assembly began. Spreading across HMR required Sir2p and Sir3p, as well as the deacetylase activity of Sir2p. These data support a model for the spreading of silenced chromatin involving cycles of nucleosome deacetylation by Sir2p followed by recruitment of additional Sir2p, Sir3p, and Sir4p to the newly deacetylated nucleosome. This model suggests mechanisms for boundary formation, and for maintenance and inheritance of silenced chromatin. The principles are generalizable to other types of heritable chromatin states. PMID- 12134061 TI - Activity of specific lipid-regulated ADP ribosylation factor-GTPase-activating proteins is required for Sec14p-dependent Golgi secretory function in yeast. AB - Yeast phosphatidylinositol transfer protein (Sec14p) coordinates lipid metabolism with protein-trafficking events. This essential Sec14p requirement for Golgi function is bypassed by mutations in any one of seven genes that control phosphatidylcholine or phosphoinositide metabolism. In addition to these "bypass Sec14p" mutations, Sec14p-independent Golgi function requires phospholipase D activity. The identities of lipids that mediate Sec14p-dependent Golgi function, and the identity of the proteins that respond to Sec14p-mediated regulation of lipid metabolism, remain elusive. We now report genetic evidence to suggest that two ADP ribosylation factor-GTPase-activating proteins (ARFGAPs), Gcs1p and Age2p, may represent these lipid-responsive elements, and that Gcs1p/Age2p act downstream of Sec14p and phospholipase D in both Sec14p-dependent and Sec14p independent pathways for yeast Golgi function. In support, biochemical data indicate that Gcs1p and Age2p ARFGAP activities are both modulated by lipids implicated in regulation of Sec14p pathway function. These results suggest ARFGAPs are stimulatory factors required for regulation of Golgi function by the Sec14p pathway, and that Sec14p-mediated regulation of lipid metabolism interfaces with the activity of proteins involved in control of the ARF cycle. PMID- 12134063 TI - Recognition of a subset of signal sequences by Ssh1p, a Sec61p-related protein in the membrane of endoplasmic reticulum of yeast Saccharomyces cerevisiae. AB - Ssh1p of Saccharomyces cerevisiae is related in sequence to Sec61p, a general receptor for signal sequences and the major subunit of the channel that guides proteins across the membrane of the endoplasmic reticulum. The split-ubiquitin technique was used to determine whether Ssh1p serves as an additional receptor for signal sequences in vivo. We measured the interactions between the N(ub) labeled Ssh1p and C(ub)-translocation substrates bearing four different signal sequences. The so-determined interaction profile of Ssh1p was compared with the signal sequence interaction profile of the correspondingly modified N(ub)-Sec61p. The assay reveals interactions of Ssh1p with the signal sequences of Kar2p and invertase, whereas Sec61p additionally interacts with the signal sequences of Mfalpha1 and carboxypeptidase Y. The measured physical proximity between Ssh1p and the beta-subunit of the signal sequence recognition particle receptor confirms our hypothesis that Ssh1p is directly involved in the cotranslational translocation of proteins across the membrane of the endoplasmic reticulum. PMID- 12134064 TI - Epidermal growth factor receptor dependence of radiation-induced transcription factor activation in human breast carcinoma cells. AB - Ionizing radiation (1-5 Gy) activates the epidermal growth factor receptor (EGFR), a major effector of the p42/44 mitogen-activated protein kinase (MAPK) pathway. MAPK and its downstream effector, p90 ribosomal S6 kinase (p90RSK), phosphorylate transcription factors involved in cell proliferation. To establish the role of the EGFR/MAPK pathway in radiation-induced transcription factor activation, MDA-MB-231 human breast carcinoma cells were examined using specific inhibitors of signaling pathways. Gel-shift analysis revealed three different profile groups: 1) transcription factors that responded to both radiation (2 Gy) and epidermal growth factor (EGF) (CREB, Egr, Ets, and Stat3); 2) factors that responded to radiation, but not EGF (C/EBP and Stat1); and 3) those that did not respond significantly to either radiation or EGF (AP-1 and Myc). Within groups 1 and 2, a two- to fivefold maximum stimulation of binding activity was observed at 30-60 min after irradiation. Interestingly, only transcription factors that responded to EGF had radiation responses significantly inhibited by the EGFR tyrosine kinase inhibitor, AG1478; these responses were also abrogated by farnesyltransferase inhibitor (FTI) or PD98059, inhibitors of Ras and MEK1/2, respectively. Moreover, radiation-induced increases in CREB and p90RSK phosphorylation and activation of Stat3 and Egr-1 reporter constructs by radiation were all abolished by AG1478. These data demonstrate a distinct radiation response profile at the transcriptional level that is dependent on enhanced EGFR/Ras/MAPK signaling. PMID- 12134065 TI - The C-terminal region of mitochondrial single-subunit RNA polymerases contains species-specific determinants for maintenance of intact mitochondrial genomes. AB - Functional conservation of mitochondrial RNA polymerases was investigated in vivo by heterologous complementation studies in yeast. It turned out that neither the full-length mitochondrial RNA polymerase of Arabidopsis thaliana, nor a set of chimeric fusion constructs from plant and yeast RNA polymerases can substitute for the yeast mitochondrial core enzyme Rpo41p when expressed in Deltarpo41 yeast mutants. Mitochondria from mutant cells, expressing the heterologous mitochondrial RNA polymerases, were devoid of any mitochondrial genomes. One important exception was observed when the carboxyl-terminal domain of Rpo41p was exchanged with its plant counterpart. Although this fusion protein could not restore respiratory function, stable maintenance of mitochondrial petite genomes (rho(-))(-) was supported. A carboxyl-terminally truncated Rpo41p exhibited a comparable activity, in spite of the fact that it was found to be transcriptionally inactive. Finally, we tested the carboxyl-terminal domain for complementation in trans. For this purpose the last 377 amino acid residues of yeast mitochondrial Rpo41p were fused to its mitochondrial import sequence. Coexpression of this fusion protein with C-terminally truncated Rpo41p complemented the Deltarpo41 defect. These data reveal the importance of the carboxyl-terminal extension of Rpo41p for stable maintenance of intact mitochondrial genomes and for distinct species-specific intramolecular protein protein interactions. PMID- 12134066 TI - Suppression of integrin activation by activated Ras or Raf does not correlate with bulk activation of ERK MAP kinase. AB - The rapid modulation of ligand-binding affinity ("activation") is a central property of the integrin family of cell adhesion receptors. The Ras family of small GTP-binding proteins and their downstream effectors are key players in regulating integrin activation. H-Ras can suppress integrin activation in fibroblasts via its downstream effector kinase, Raf-1. In contrast, to H-Ras, a closely related small GTP-binding protein R-Ras has the opposite activity, and promotes integrin activation. To gain insight into the regulation of integrin activation by Ras GTPases, we created a series of H-Ras/R-Ras chimeras. We found that a 35-amino acid stretch of H-Ras was required for full suppressive activity. Furthermore, the suppressive chimeras were weak activators of the ERK1/2 MAP kinase pathway, suggesting that the suppression of integrin activation may be independent of the activation of the bulk of ERK MAP kinase. Additional data demonstrating that the ability of H-Ras or Raf-1 to suppress integrin activation was unaffected by inhibition of bulk ERK1/2 MAP kinase activation supported this hypothesis. Thus, the suppression of integrin activation is a Raf kinase induced regulatory event that can be mediated independently of bulk activation of the ERK MAP-kinase pathway. PMID- 12134067 TI - Role for YakA, cAMP, and protein kinase A in regulation of stress responses of Dictyostelium discoideum cells. AB - The Dictyostelium protein kinase YakA is required for the growth-to-development transition. During growth YakA controls the cell cycle, regulating the intervals between cell divisions. When starved for nutrients Dictyostelium cells arrest growth and undergo changes in gene expression, decreasing vegetative mRNAs and inducing the expression of pkaC. YakA is an effector of these changes, being necessary for the decrease of vegetative mRNA expression and the increase of protein kinase A (PKA) activity that will ultimately regulate expression of adenylyl cyclase, cAMP synthesis, and the induction of development. We report a role for this kinase in the response to nitrosoative or oxidative stress of Dictyostelium cells. Hydrogen peroxide and sodium nitroprusside arrest the growth of cells and trigger cAMP synthesis and activation of PKA in a manner similar to the well-established response to nutrient starvation. We have found that yakA null cells are hypersensitive to nitrosoative/oxidative stress and that a second site mutation in pkaC suppresses this sensitivity. The response to different stresses has been investigated and YakA, cAMP, and PKA have been identified as components of the pathway that regulate the growth arrest that follows treatment with compounds that generate reactive oxygen species. The effect of different types of stress was evaluated in Dictyostelium and the YakA/PKA pathway was also implicated in the response to heat stress. PMID- 12134070 TI - Dictyostelium EB1 is a genuine centrosomal component required for proper spindle formation. AB - EB1 proteins are ubiquitous microtubule-associated proteins involved in microtubule search and capture, regulation of microtubule dynamics, cell polarity, and chromosome stability. We have cloned a complete cDNA of Dictyostelium EB1 (DdEB1), the largest known EB1 homolog (57 kDa). Immunofluorescence analysis and expression of a green fluorescent protein-DdEB1 fusion protein revealed that DdEB1 localizes along microtubules, at microtubule tips, centrosomes, and protruding pseudopods. During mitosis, it was found at the spindle, spindle poles, and kinetochores. DdEB1 is the first EB1-homolog that is also a genuine centrosomal component, because it was localized at isolated centrosomes that are free of microtubules. Furthermore, centrosomal DdEB1 distribution was unaffected by nocodazole treatment. DdEB1 colocalized with DdCP224, the XMAP215 homolog, at microtubule tips, the centrosome, and kinetochores. Furthermore, both proteins were part of the same cytosolic protein complex, suggesting that they may act together in their functions. DdEB1 deletion mutants expressed as green fluorescent protein or maltose-binding fusion proteins indicated that microtubule binding requires homo-oligomerization, which is mediated by a coiled-coil domain. A DdEB1 null mutant was viable but retarded in prometaphase progression due to a defect in spindle formation. Because spindle elongation was normal, DdEB1 seems to be required for the initiation of the outgrowth of spindle microtubules. PMID- 12134068 TI - Akt maintains cell size and survival by increasing mTOR-dependent nutrient uptake. AB - In multicellular organisms, constituent cells depend on extracellular signals for growth, proliferation, and survival. When cells are withdrawn from growth factors, they undergo apoptosis. Expression of constitutively active forms of the serine/threonine kinase Akt/PKB can prevent apoptosis upon growth factor withdrawal. Akt-mediated survival depends in part on the maintenance of glucose metabolism, suggesting that reduced glucose utilization contributes to growth factor withdrawal-induced death. However, it is unclear how restricting access to extracellular glucose alone would lead to the metabolic collapse observed after growth factor withdrawal. We report herein that growth factor withdrawal results in the loss of surface transporters for not only glucose but also amino acids, low-density lipoprotein, and iron. This coordinated decline in transporters and receptors for extracellular molecules creates a catabolic state characterized by atrophy and a decline in the mitochondrial membrane potential. Activated forms of Akt maintained these transporters on the cell surface in the absence of growth factor through an mTOR-dependent mechanism. The mTOR inhibitor rapamycin diminished Akt-mediated increases in cell size, mitochondrial membrane potential, and cell survival. These results suggest that growth factors control cellular growth and survival by regulating cellular access to extracellular nutrients in part by modulating the activity of Akt and mTOR. PMID- 12134069 TI - Disruption of centrosome structure, chromosome segregation, and cytokinesis by misexpression of human Cdc14A phosphatase. AB - In budding yeast, the Cdc14p phosphatase activates mitotic exit by dephosphorylation of specific cyclin-dependent kinase (Cdk) substrates and seems to be regulated by sequestration in the nucleolus until its release in mitosis. Herein, we have analyzed the two human homologs of Cdc14p, hCdc14A and hCdc14B. We demonstrate that the human Cdc14A phosphatase is selective for Cdk substrates in vitro and that although the protein abundance and intrinsic phosphatase activity of hCdc14A and B vary modestly during the cell cycle, their localization is cell cycle regulated. hCdc14A dynamically localizes to interphase but not mitotic centrosomes, and hCdc14B localizes to the interphase nucleolus. These distinct patterns of localization suggest that each isoform of human Cdc14 likely regulates separate cell cycle events. In addition, hCdc14A overexpression induces the loss of the pericentriolar markers pericentrin and gamma-tubulin from centrosomes. Overproduction of hCdc14A also causes mitotic spindle and chromosome segregation defects, defective karyokinesis, and a failure to complete cytokinesis. Thus, the hCdc14A phosphatase appears to play a role in the regulation of the centrosome cycle, mitosis, and cytokinesis, thereby influencing chromosome partitioning and genomic stability in human cells. PMID- 12134071 TI - PC phosphorylation increases the ability of AFAP-110 to cross-link actin filaments. AB - The actin filament-associated protein and Src-binding partner, AFAP-110, is an adaptor protein that links signaling molecules to actin filaments. AFAP-110 binds actin filaments directly and multimerizes through a leucine zipper motif. Cellular signals downstream of Src(527F) can regulate multimerization. Here, we determined recombinant AFAP-110 (rAFAP-110)-bound actin filaments cooperatively, through a lateral association. We demonstrate rAFAP-110 has the capability to cross-link actin filaments, and this ability is dependent on the integrity of the carboxy terminal actin binding domain. Deletion of the leucine zipper motif or PKC phosphorylation affected AFAP-110's conformation, which correlated with changes in multimerization and increased the capability of rAFAP-110 to cross link actin filaments. AFAP-110 is both a substrate and binding partner of PKC. On PKC activation, stress filament organization is lost, motility structures form, and AFAP-110 colocalizes strongly with motility structures. Expression of a deletion mutant of AFAP-110 that is unable to bind PKC blocked the effect of PMA on actin filaments. We hypothesize that upon PKC activation, AFAP-110 can be cooperatively recruited to newly forming actin filaments, like those that exist in cell motility structures, and that PKC phosphorylation effects a conformational change that may enable AFAP-110 to promote actin filament cross linking at the cell membrane. PMID- 12134072 TI - Structural determinants of Ras-Raf interaction analyzed in live cells. AB - The minimum structure of the Raf-1 serine/threonine kinase that recognizes active Ras was used to create a green fluorescent fusion protein (GFP) for monitoring Ras activation in live cells. In spite of its ability to bind activated Ras in vitro, the Ras binding domain (RBD) of Raf-1 (Raf-1[51-131]GFP) failed to detect Ras in Ras-transformed NIH 3T3 fibroblasts and required the addition of the cysteine-rich domain (CRD) (Raf-1[51-220]GFP) to show clear localization to plasma membrane ruffles. In normal NIH 3T3 cells, (Raf-1[51-220]GFP) showed minimal membrane localization that was enhanced after stimulation with platelet derived growth factor or phorbol-12-myristate-13-acetate. Mutations within either the RBD (R89L) or CRD (C168S) disrupted the membrane localization of (Raf-1[51 220]GFP), suggesting that both domains contribute to the recruitment of the fusion protein to Ras at the plasma membrane. The abilities of the various constructs to localize to the plasma membrane closely correlated with their inhibitory effects on mitogen-activated protein kinase kinase1 and mitogen activated protein kinase activation. Membrane localization of full-length Raf-1 GFP was less prominent than that of (Raf-1[51-220]GFP) in spite of its strong binding to RasV12 and potent activation of mitogen-activated protein kinase. These finding indicate that both RBD and CRD are necessary to recruit Raf-1 to active Ras at the plasma membrane, and that these domains are not fully exposed in the Raf-1 molecule. Visualization of activated Ras in live cells will help to better understand the dynamics of Ras activation under various physiological and pathological conditions. PMID- 12134074 TI - Sarcolemmal organization in skeletal muscle lacking desmin: evidence for cytokeratins associated with the membrane skeleton at costameres. AB - The sarcolemma of fast-twitch muscle is organized into "costameres," structures that are oriented transversely, over the Z and M lines of nearby myofibrils, and longitudinally, to form a rectilinear lattice. Here we examine the role of desmin, the major intermediate filament protein of muscle in organizing costameres. In control mouse muscle, desmin is enriched at the sarcolemmal domains that lie over nearby Z lines and that also contain beta-spectrin. In tibialis anterior muscle from mice lacking desmin due to homologous recombination, most costameres are lost. In myofibers from desmin -/- quadriceps, by contrast, most costameric structures are stable. Alternatively, Z line domains may be lost, whereas domains oriented longitudinally or lying over M lines are retained. Experiments with pan-specific antibodies to intermediate filament proteins and to cytokeratins suggest that control and desmin -/- muscles express similar levels of cytokeratins. Cytokeratins concentrate at the sarcolemma at all three domains of costameres when the latter are retained in desmin -/- muscle and redistribute with beta-spectrin at the sarcolemma when costameres are lost. Our results suggest that desmin associates with and selectively stabilizes the Z line domains of costameres, but that cytokeratins associate with all three domains of costameres, even in the absence of desmin. PMID- 12134075 TI - A fourth component of the fission yeast gamma-tubulin complex, Alp16, is required for cytoplasmic microtubule integrity and becomes indispensable when gamma tubulin function is compromised. AB - gamma-Tubulin functions as a multiprotein complex, called the gamma-tubulin complex (gamma-TuC), and composes the microtubule organizing center (MTOC). Fission yeast Alp4 and Alp6 are homologues of two conserved gamma-TuC proteins, hGCP2 and hGCP3, respectively. We isolated a novel gene, alp16(+), as a multicopy suppressor of temperature-sensitive alp6-719 mutants. alp16(+) encodes a 759 amino-acid protein with two conserved regions found in all other members of gamma TuC components. In addition, Alp16 contains an additional motif, which shows homology to hGCP6/Xgrip210. Gene disruption shows that alp16(+) is not essential for cell viability. However, alp16 deletion displays abnormally long cytoplasmic microtubules, which curve around the cell tip. Furthermore, alp16-deleted mutants are hypersensitive to microtubule-depolymerizing drugs and synthetically lethal with either temperature-sensitive alp4-225, alp4-1891, or alp6-719 mutants. Overproduction of Alp16 is lethal, with defective phenotypes very similar to loss of Alp4 or Alp6. Alp16 localizes to the spindle pole body throughout the cell cycle and to the equatorial MTOC at postanaphase. Alp16 coimmunoprecipitates with gamma-tubulin and cosediments with the gamma-TuC in a large complex (>20 S). Alp16 is, however, not required for the formation of this large complex. We discuss evolutional conservation and divergence of structure and function of the gamma-TuC between yeast and higher eukaryotes. PMID- 12134073 TI - Small GTP-binding protein TC10 differentially regulates two distinct populations of filamentous actin in 3T3L1 adipocytes. AB - TC10 is a member of the Rho family of small GTP-binding proteins that has previously been implicated in the regulation of insulin-stimulated GLUT4 translocation in adipocytes. In a manner similar to Cdc42-stimulated actin-based motility, we have observed that constitutively active TC10 (TC10/Q75L) can induce actin comet tails in Xenopus oocyte extracts in vitro and extensive actin polymerization in the perinuclear region when expressed in 3T3L1 adipocytes. In contrast, expression of TC10/Q75L completely disrupted adipocyte cortical actin, which was specific for TC10, because expression of constitutively active Cdc42 was without effect. The effect of TC10/Q75L to disrupt cortical actin was abrogated after deletion of the amino terminal extension (DeltaN-TC10/Q75L), whereas this deletion retained the ability to induce perinuclear actin polymerization. In addition, alteration of perinuclear actin by expression of TC10/Q75L, a dominant-interfering TC10/T31N mutant or a mutant N-WASP protein (N WASP/DeltaVCA) reduced the rate of VSV G protein trafficking to the plasma membrane. Furthermore, TC10 directly bound to Golgi COPI coat proteins through a dilysine motif in the carboxyl terminal domain consistent with a role for TC10 regulating actin polymerization on membrane transport vesicles. Together, these data demonstrate that TC10 can differentially regulate two types of filamentous actin in adipocytes dependent on distinct functional domains and its subcellular compartmentalization. PMID- 12134076 TI - Differential recognition of tyrosine-based basolateral signals by AP-1B subunit mu1B in polarized epithelial cells. AB - To investigate the importance of tyrosine recognition by the AP-1B clathrin adaptor subunit mu1B for basolateral sorting of integral membrane proteins in polarized epithelial cells, we have produced and characterized a mutant form of mu1B. The mutant (M-mu1B) contains alanine substitutions of each of the four conserved residues, which in the AP-2 adaptor subunit micro2 are critical for interacting with tyrosine-based endocytosis signals. We show M-mu1B is defective for tyrosine binding in vitro, but is nevertheless incorporated into AP-1 complexes in transfected cells. Using LLC-PK1 cells expressing either wild type or M-mu1B, we find that there is inefficient basolateral expression of membrane proteins whose basolateral targeting signals share critical tyrosines with signals for endocytosis. In contrast, membrane proteins whose basolateral targeting signals are distinct from their endocytosis signals (transferrin and low-density lipoprotein receptors) accumulate at the basolateral domain normally, although in a manner that is strictly dependent on mu1B or M-mu1B expression. Our results suggest that mu1B interacts with different classes of basolateral targeting signals in distinct ways. PMID- 12134077 TI - Contribution of Ena/VASP proteins to intracellular motility of listeria requires phosphorylation and proline-rich core but not F-actin binding or multimerization. AB - The Listeria model system has been essential for the identification and characterization of key regulators of the actin cytoskeleton such as the Arp2/3 complex and Ena/vasodilator-stimulated phosphoprotein (VASP) proteins. Although the role of Ena/VASP proteins in Listeria motility has been extensively studied, little is known about the contributions of their domains and phosphorylation state to bacterial motility. To address these issues, we have generated a panel of Ena/VASP mutants and, upon expression in Ena/VASP-deficient cells, evaluated their contribution to Ena/VASP function in Listeria motility. The proline-rich region, the putative G-actin binding site, and the Ser/Thr phosphorylation of Ena/VASP proteins are all required for efficient Listeria motility. Surprisingly, the interaction of Ena/VASP proteins with F-actin and their potential ability to form multimers are both dispensable for their involvement in this process. Our data suggest that Ena/VASP proteins contribute to Listeria motility by regulating both the nucleation and elongation of actin filaments at the bacterial surface. PMID- 12134078 TI - Biogenesis of nanotubular network in Toxoplasma parasitophorous vacuole induced by parasite proteins. AB - The intracellular parasite Toxoplasma gondii develops within a nonfusogenic vacuole containing a network of elongated nanotubules that form connections with the vacuolar membrane. Parasite secretory proteins discharged from dense granules (known as GRA proteins) decorate this intravacuolar network after invasion. Herein, we show using specific gene knockout mutants, that the unique nanotubule conformation of the network is induced by the parasite secretory protein GRA2 and further stabilized by GRA6. The vacuolar compartment generated by GRA2 knockout parasites was dramatically disorganized, and the normally tubular network was replaced by small aggregated material. The defect observed in Deltagra2 parasites was evident from the initial stages of network formation when a prominent cluster of multilamellar vesicles forms at a posterior invagination of the parasite. The secretory protein GRA6 failed to localize properly to this posterior organizing center in Deltagra2 cells, indicating that this early conformation is essential to proper assembly of the network. Construction of a Deltagra6 mutant also led to an altered mature network characterized by small vesicles instead of elongated nanotubules; however, the initial formation of the posterior organizing center was normal. Complementation of the Deltagra2 knockout with mutated forms of GRA2 showed that the integrity of both amphipathic alpha-helices of the protein is required for correct formation of the network. The induction of nanotubues by the parasite protein GRA2 may be a conserved feature of amphipathic alpha-helical regions, which have also been implicated in the organization of Golgi nanotubules and endocytic vesicles in mammalian cells. PMID- 12134079 TI - Neuronal ceroid lipofuscinoses are connected at molecular level: interaction of CLN5 protein with CLN2 and CLN3. AB - Neuronal ceroid lipofuscinoses (NCLs) are neurodegenerative storage diseases characterized by mental retardation, visual failure, and brain atrophy as well as accumulation of storage material in multiple cell types. The diseases are caused by mutations in the ubiquitously expressed genes, of which six are known. Herein, we report that three NCL disease forms with similar tissue pathology are connected at the molecular level: CLN5 polypeptides directly interact with the CLN2 and CLN3 proteins based on coimmunoprecipitation and in vitro binding assays. Furthermore, disease mutations in CLN5 abolished interaction with CLN2, while not affecting association with CLN3. The molecular characterization of CLN5 revealed that it was synthesized as four precursor forms, due to usage of alternative initiator methionines in translation. All forms were targeted to lysosomes and the longest form, translated from the first potential methionine, was associated with membranes. Interactions between CLN polypeptides were shown to occur with this longest, membrane-bound form of CLN5. Both intracellular targeting and posttranslational glycosylation of the polypeptides carrying human disease mutations were similar to wild-type CLN5. PMID- 12134080 TI - GLUT4 retention in adipocytes requires two intracellular insulin-regulated transport steps. AB - Insulin regulates glucose uptake into fat and muscle by modulating the distribution of the GLUT4 glucose transporter between the surface and interior of cells. The GLUT4 trafficking pathway overlaps with the general endocytic recycling pathway, but the degree and functional significance of the overlap are not known. In this study of intact adipocytes, we demonstrate, by using a compartment-specific fluorescence-quenching assay, that GLUT4 is equally distributed between two intracellular pools: the transferrin receptor-containing endosomes and a specialized compartment that excludes the transferrin receptor. These pools of GLUT4 are in dynamic communication with one another and with the cell surface. Insulin-induced redistribution of GLUT4 to the surface requires mobilization of both pools. These data establish a role for the general endosomal system in the specialized, insulin-regulated trafficking of GLUT4. Trafficking through the general endosomal system is regulated by rab11. Herein, we show that rab11 is required for the transport of GLUT4 from endosomes to the specialized compartment and for the insulin-induced translocation to the cell surface, emphasizing the importance of the general endosomal pathway in the specialized trafficking of GLUT4. Based on these findings we propose a two-step model for GLUT4 trafficking in which the general endosomal recycling compartment plays a specialized role in the insulin-regulated traffic of GLUT4. This compartment based model provides the framework for understanding insulin-regulated trafficking at a molecular level. PMID- 12134082 TI - Molecular dissection of cytokinesis by RNA interference in Drosophila cultured cells. AB - We have used double-stranded RNA-mediated interference (RNAi) to study Drosophila cytokinesis. We show that double-stranded RNAs for anillin, acGAP, pavarotti, rho1, pebble, spaghetti squash, syntaxin1A, and twinstar all disrupt cytokinesis in S2 tissue culture cells, causing gene-specific phenotypes. Our phenotypic analyses identify genes required for different aspects of cytokinesis, such as central spindle formation, actin accumulation at the cell equator, contractile ring assembly or disassembly, and membrane behavior. Moreover, the cytological phenotypes elicited by RNAi reveal simultaneous disruption of multiple aspects of cytokinesis. These phenotypes suggest interactions between central spindle microtubules, the actin-based contractile ring, and the plasma membrane, and lead us to propose that the central spindle and the contractile ring are interdependent structures. Finally, our results indicate that RNAi in S2 cells is a highly efficient method to detect cytokinetic genes, and predict that genome wide studies using this method will permit identification of the majority of genes involved in Drosophila mitotic cytokinesis. PMID- 12134081 TI - A conserved Drosophila transportin-serine/arginine-rich (SR) protein permits nuclear import of Drosophila SR protein splicing factors and their antagonist repressor splicing factor 1. AB - Members of the highly conserved serine/arginine-rich (SR) protein family are nuclear factors involved in splicing of metazoan mRNA precursors. In mammals, two nuclear import receptors, transportin (TRN)-SR1 and TRN-SR2, are responsible for targeting SR proteins to the nucleus. Distinctive features in the nuclear localization signal between Drosophila and mammalian SR proteins prompted us to examine the mechanism by which Drosophila SR proteins and their antagonist repressor splicing factor 1 (RSF1) are imported into nucleus. Herein, we report the identification and characterization of a Drosophila importin beta-family protein (dTRN-SR), homologous to TRN-SR2, that specifically interacts with both SR proteins and RSF1. dTRN-SR has a broad localization in the cytoplasm and the nucleus, whereas an N-terminal deletion mutant colocalizes with SR proteins in nuclear speckles. Far Western experiments established that the RS domain of SR proteins and the GRS domain of RSF1 are required for the direct interaction with dTRN-SR, an interaction that can be modulated by phosphorylation. Using the yeast model system in which nuclear import of Drosophila SR proteins and RSF1 is impaired, we demonstrate that complementation with dTRN-SR is sufficient to target these proteins to the nucleus. Together, the results imply that the mechanism by which SR proteins are imported to the nucleus is conserved between Drosophila and humans. PMID- 12134083 TI - Large-scale isolation of Cajal bodies from HeLa cells. AB - The Cajal body (CB) is a conserved, dynamic nuclear structure that is implicated in various cellular processes, such as the maturation of splicing small nuclear ribonucleoproteins and the assembly of transcription complexes. Here, we report the first procedure for the large-scale purification of CBs from HeLa cell nuclei, resulting in an approximately 750-fold enrichment of the CB marker protein p80-coilin. Immunofluorescence, immunoblotting, and mass spectrometric analyses showed that the composition of the isolated CBs was similar to that of CBs in situ. The morphology and structure of the isolated CBs, as judged by transmission and scanning electron microscopy analysis, are also similar to those of CBs in situ. This protocol demonstrates the feasibility of isolating intact distinct classes of subnuclear bodies from cultured cells in sufficient yield and purity to allow detailed characterization of their molecular composition, structure, and properties. PMID- 12134084 TI - Rac regulates endothelial morphogenesis and capillary assembly. AB - Endothelial cells undergo branching morphogenesis to form capillary tubes. We have utilized an in vitro Matrigel overlay assay to analyze the role of the cytoskeleton and Rho GTPases during this process. The addition of matrix first induces changes in cell morphology characterized by the formation of dynamic cellular protrusions and the assembly of discrete aggregates or cords of aligned cells resembling primitive capillary-like structures, but without a recognizable lumen. This is followed by cell migration leading to the formation of a complex interconnecting network of capillary tubes with readily identifiable lumens. Inhibition of actin polymerization or actin-myosin contraction inhibits cell migration but has no effect on the initial changes in endothelial cell morphology. However, inhibition of microtubule dynamics prevents both the initial cell shape changes as well as cell migration. We find that the small GTPase Rac is essential for the matrix-induced changes in endothelial cell morphology, whereas p21-activated kinase, an effector of Rac, is required for cell motility. We conclude that Rac integrates signaling through both the actin and microtubule cytoskeletons to promote capillary tube assembly. PMID- 12134085 TI - Genomic screen for vacuolar protein sorting genes in Saccharomyces cerevisiae. AB - The biosynthetic sorting of hydrolases to the yeast vacuole involves transport along two distinct routes referred to as the carboxypeptidase Y and alkaline phosphatase pathways. To identify genes involved in sorting to the vacuole, we conducted a genome-wide screen of 4653 homozygous diploid gene deletion strains of Saccharomyces cerevisiae for missorting of carboxypeptidase Y. We identified 146 mutant strains that secreted strong-to-moderate levels of carboxypeptidase Y. Of these, only 53 of the corresponding genes had been previously implicated in vacuolar protein sorting, whereas the remaining 93 had either been identified in screens for other cellular processes or were only known as hypothetical open reading frames. Among these 93 were genes encoding: 1) the Ras-like GTP-binding proteins Arl1p and Arl3p, 2) actin-related proteins such as Arp5p and Arp6p, 3) the monensin and brefeldin A hypersensitivity proteins Mon1p and Mon2p, and 4) 15 novel proteins designated Vps61p-Vps75p. Most of the novel gene products were involved only in the carboxypeptidase Y pathway, whereas a few, including Mon1p, Mon2p, Vps61p, and Vps67p, appeared to be involved in both the carboxypeptidase Y and alkaline phosphatase pathways. Mutants lacking some of the novel gene products, including Arp5p, Arp6p, Vps64p, and Vps67p, were severely defective in secretion of mature alpha-factor. Others, such as Vps61p, Vps64p, and Vps67p, displayed defects in the actin cytoskeleton at 30 degrees C. The identification and phenotypic characterization of these novel mutants provide new insights into the mechanisms of vacuolar protein sorting, most notably the probable involvement of the actin cytoskeleton in this process. PMID- 12134086 TI - Expression of caveolin-1 induces premature cellular senescence in primary cultures of murine fibroblasts. AB - Caveolae are vesicular invaginations of the plasma membrane. Caveolin-1 is the principal structural component of caveolae in vivo. Several lines of evidence are consistent with the idea that caveolin-1 functions as a "transformation suppressor" protein. In fact, caveolin-1 mRNA and protein expression are lost or reduced during cell transformation by activated oncogenes. Interestingly, the human caveolin-1 gene is localized to a suspected tumor suppressor locus (7q31.1). We have previously demonstrated that overexpression of caveolin-1 arrests mouse embryonic fibroblasts in the G(0)/G(1) phase of the cell cycle through activation of a p53/p21-dependent pathway, indicating a role of caveolin 1 in mediating growth arrest. However, it remains unknown whether overexpression of caveolin-1 promotes cellular senescence in vivo. Here, we demonstrate that mouse embryonic fibroblasts transgenically overexpressing caveolin-1 show: 1) a reduced proliferative lifespan; 2) senescence-like cell morphology; and 3) a senescence-associated increase in beta-galactosidase activity. These results indicate for the first time that the expression of caveolin-1 in vivo is sufficient to promote and maintain the senescent phenotype. Subcytotoxic oxidative stress is known to induce premature senescence in diploid fibroblasts. Interestingly, we show that subcytotoxic level of hydrogen peroxide induces premature senescence in NIH 3T3 cells and increases endogenous caveolin-1 expression. Importantly, quercetin and vitamin E, two antioxidant agents, successfully prevent the premature senescent phenotype and the up-regulation of caveolin-1 induced by hydrogen peroxide. Also, we demonstrate that hydrogen peroxide alone, but not in combination with quercetin, stimulates the caveolin-1 promoter activity. Interestingly, premature senescence induced by hydrogen peroxide is greatly reduced in NIH 3T3 cells harboring antisense caveolin-1. Importantly, induction of premature senescence is recovered when caveolin-1 levels are restored. Taken together, these results clearly indicate a central role for caveolin-1 in promoting cellular senescence and they suggest the hypothesis that premature senescence may represent a tumor suppressor function mediated by caveolin-1 in vivo. PMID- 12134087 TI - Emp47p and its close homolog Emp46p have a tyrosine-containing endoplasmic reticulum exit signal and function in glycoprotein secretion in Saccharomyces cerevisiae. AB - The yeast open reading frame YLR080w/EMP46 encodes a homolog of the Golgi protein Emp47p. These two proteins are 45% identical and have a single transmembrane domain in their C-terminal regions and a carbohydrate recognition domain signature in the N-terminal region. The C-terminal tail of Emp46p includes a dilysine signal. This protein is localized to Golgi membranes at steady state by subcellular fractionation and green fluorescent protein labeling. On block of forward transport in sec12-4 cells, redistribution of Emp46p from the Golgi to the endoplasmic reticulum is observed. These localization features are similar to those previously reported for Emp47p. In addition, mutagenesis of the C-terminal region identified a tyrosine-containing motif as a critical determinant of the Golgi-localization and interaction with both COPI and COPII components. Similar motifs are also observed in the C-terminal tail of Emp47p and other mammalian homologs. Disruption of Emp47p displays a growth defect at a high temperature or on Ca(2+)-containing medium, which is rescued by overexpression of Emp46p, suggesting a partially overlapping function between Emp46p and Emp47p. In addition, we found that the disruption of both Emp46p and Emp47p show a marked defect in the secretion of a subset of glycoproteins. Analysis of the C-terminal mutants for Ca(2+) sensitivity revealed that the forward transport of Emp46/47p is essential for their function, whereas the retrograde transport is not. We propose that Emp46p and Emp47p are required for the export of specific glycoprotein cargo from the endoplasmic reticulum. PMID- 12134090 TI - Should dialysis arteriovenous fistulas be closed after renal transplantation? PMID- 12134088 TI - Critical roles of phosphorylation and actin binding motifs, but not the central proline-rich region, for Ena/vasodilator-stimulated phosphoprotein (VASP) function during cell migration. AB - The Ena/vasodilator-stimulated phosphoprotein (VASP) protein family is implicated in the regulation of a number of actin-based cellular processes, including lamellipodial protrusion necessary for whole cell translocation. A growing body of evidence derived largely from in vitro biochemical experiments using purified proteins, cell-free extracts, and pathogen motility has begun to suggest various mechanistic roles for Ena/VASP proteins in the control of actin dynamics. Using complementation of phenotypes in Ena/VASP-deficient cells and overexpression in normal fibroblasts, we have assayed the function of a panel of mutants in one member of this family, Mena, by mutating highly conserved sequence elements found in this protein family. Surprisingly, deletion of sites required for binding of the actin monomer-binding protein profilin, a known ligand of Ena/VASP proteins, has no effect on the ability of Mena to regulate random cell motility. Our analysis revealed two features essential for Ena/VASP function in cell movement, cyclic nucleotide-dependent kinase phosphorylation sites and an F-actin binding motif. Interestingly, expression of the C-terminal EVH2 domain alone is sufficient to complement loss of Ena/VASP function in random cell motility. PMID- 12134089 TI - Ligand-independent dimer formation of epidermal growth factor receptor (EGFR) is a step separable from ligand-induced EGFR signaling. AB - Dimerization and phosphorylation of the epidermal growth factor (EGF) receptor (EGFR) are the initial and essential events of EGF-induced signal transduction. However, the mechanism by which EGFR ligands induce dimerization and phosphorylation is not fully understood. Here, we demonstrate that EGFRs can form dimers on the cell surface independent of ligand binding. However, a chimeric receptor, comprising the extracellular and transmembrane domains of EGFR and the cytoplasmic domain of the erythropoietin receptor (EpoR), did not form a dimer in the absence of ligands, suggesting that the cytoplasmic domain of EGFR is important for predimer formation. Analysis of deletion mutants of EGFR showed that the region between (835)Ala and (918)Asp of the EGFR cytoplasmic domain is required for EGFR predimer formation. In contrast to wild-type EGFR ligands, a mutant form of heparin-binding EGF-like growth factor (HB2) did not induce dimerization of the EGFR-EpoR chimeric receptor and therefore failed to activate the chimeric receptor. However, when the dimerization was induced by a monoclonal antibody to EGFR, HB2 could activate the chimeric receptor. These results indicate that EGFR can form a ligand-independent inactive dimer and that receptor dimerization and activation are mechanistically distinct and separable events. PMID- 12134091 TI - A peptide mimic of selectin ligands abolishes in vivo inflammation but has no effect on the rat heart allograft survival1. AB - Acute heart allograft rejection is characterized by leukocyte infiltration and myocyte damage, key elements in the histological grading of rejection. The induction of selectins and their ligands on the graft postcapillary venular endothelium increases leukocyte tethering to, rolling on, and extravasation through the endothelium into graft parenchyma. We have characterized peptide mimicking selectin ligands by screening phage peptide libraries using anti-Lewis A antibodies and E-selectin as target molecules. The effect of this selectin- binding peptide, IELLQAR, on the prevention of inflammation and tissue damage and on the prolongation of graft survival in inbred DA (RT1a) rat heart allografts transplanted to WF (RT1v) recipients was tested. Bovine serum albumin (0.1%, solvent), VTSIAQA (control peptide), or IELLQAR were either continuously infused into the peritoneum via osmotic mini pumps or injected twice daily IV. Treatment with bovine serum albumin and VTSIAQA did not alter the number of graft infiltrating leukocytes or the histological grade of acute rejection, all scored as grade 4. On the contrary, the selectin binding peptide, IELLQAR, dose dependently reduced inflammation and at the highest dose (6.0 mg/kg per day) eliminated the majority of graft infiltrating leukocytes, reduced the histological grade from 4 to 1B, but had no effect on graft survival. These data indicate that the intensity of inflammation related to the allograft rejection does not correlate to the graft survival. PMID- 12134092 TI - Activation of the lipopolysaccharide signaling pathway in hepatic transplantation preservation injury. AB - BACKGROUND: Endotoxin or lipopolysaccharide (LPS) initiates a cascade of complications of septic shock and multiple organ failure seen in Gram-negative bacterial infections. The first step of this pathway, which leads to activated nuclear factor (NF)-kappaB, activating protein (AP)-1, and other transcription factors, is the formation of the LPS receptor complex by LPS, LPS-binding protein (LBP), CD14, and toll-like receptor (TLR) 2 or 4. We examined whether the LPS signaling pathway is activated by hepatic ischemia/reperfusion injury in the transplant setting. METHODS: Orthotopic syngeneic rat liver transplantation was performed with 0 to 18 hr of cold preservation in University of Wisconsin solution. Animals were killed 1 to 48 hr after reperfusion. Northern blot analysis for CD14, LBP, and TLR2 mRNA, immunohistochemistry for LBP, liver enzyme analysis, and gel shift assay for NF-kappaB and AP-1 were performed. RESULTS: LPS levels were elevated early after reperfusion. Aspartate aminotransferase and alanine aminotransferase maximally increased 12 hr after transplantation. LBP mRNA and protein and CD14 mRNA were significantly up-regulated peaking at 6 to 12 hr after reperfusion. TLR2 mRNA was also increased. NF-kappaB activity showed a biphasic peak at 1 to 3 hr and 12 hr after reperfusion, whereas AP-1 activity showed a peak at 3 to 6 hr. The induction of CD14 mRNA correlated with the length of cold ischemia time. CONCLUSIONS: These data indicate that multiple components of the LPS signaling pathway are activated during ischemia/reperfusion injury after liver transplantation. PMID- 12134093 TI - Microgravity culture condition reduces immunogenicity and improves function of pancreatic islets1. AB - BACKGROUND: The failure of pancreatic islet allotransplants observed in almost all clinical attempts is related to poor initial islet function and allograft rejection. To remedy these problems we cultured islets in microgravity conditions to improve their function and to reduce their immunogenicity. METHODS: Fresh mouse islets or mouse islets cultured in stationary dishes or microgravity bioreactors were transplanted to streptozotocin-induced diabetic mouse recipients. RESULTS: Both allogeneic dish- or bioreactor-cultured islets survived more than 100 days compared with fresh allogeneic islets, which were rejected in less than 15 days. Islet titration studies revealed that 250 fresh or dish cultured, but only 30 to 120 bioreactor-cultured, islets were necessary to produce euglycemia. Furthermore, glucose tolerance tests showed that bioreactor cultured islets functioned better compared with fresh and dish-cultured islets on day 30 postgrafting. Immunostaining and transmission electron microscopy (TEM) analyses showed the gradual disappearance of dendritic cells in cultured islets compared with fresh islets. TEM revealed that the ultrastructure of islets from bioreactor, but not dish, appeared healthy and closely resembled fresh islets. Interestingly, TEM and scanning electron microscopy showed that only bioreactor cultured islets developed unique and multiple nutritional channels between arrays of islet cells. TEM with colloidal lanthanum tracer revealed that only bioreactor islet cell cultures were devoid of tight junctional complexes, which may facilitate channel formation. CONCLUSION: Microgravity condition decreases immunogenicity and significantly improves the function of secretory cells. PMID- 12134094 TI - Inducing tolerance to MHC-matched allogeneic islet grafts in diabetic NOD mice by simultaneous islet and bone marrow transplantation under nonirradiative and nonmyeloablative conditioning therapy. AB - BACKGROUND: Human type 1 diabetes is associated with defects in the hematopoietic stem cells. Simultaneous donor islet and bone marrow transplantation may be an ideal therapeutic approach for inducing tolerance to islet allogeneic antigens and restoring self-tolerance to islet autoimmune antigens. METHODS: Using a nonobese diabetic (NOD) mouse model of human type 1 diabetes, we investigated whether tolerance to MHC-matched allogeneic islet grafts from male nonobese diabetes-resistant (NOR) donors can be induced in female NOD recipients by simultaneous islet and bone marrow transplantation under fludarabine phosphate based nonmyeloablative and irradiation-free conditioning therapy. Donor-specific chimerism in the peripheral blood of tolerant mice (n=7) was measured by semiquantitative polymerase chain reaction for a male-specific marker (SRY). RESULTS: Donor-specific tolerance to NOR islet grafts was induced in all diabetic NOD mice after simultaneous islet and bone marrow transplantation and treated with fludarabine phosphate, cyclophosphamide, anti-mouse lymphocyte serum, and rapamycin. At 100 days and 200 days after transplantation, the average percentage of male NOR marker in DNA derived from the peripheral blood of NOD recipients that had long-term islet graft survival was over 10%. CONCLUSION: Our data suggest that this approach may induce donor-specific tolerance in clinical islet transplantation and living-related donor solid organ transplantation. PMID- 12134095 TI - Improvement in rejection of human decay accelerating factor transgenic pig-to primate renal xenografts with administration of rabbit antithymocyte serum. AB - BACKGROUND: Survival in pig-to-baboon kidney xenotransplantation is currently limited by acute humoral xenograft rejection (AHXR). We hypothesized that the administration of rabbit antithymocyte serum (RATS) would delay or prevent AHXR as compared with a cyclophosphamide (CyP)-based immunosuppressive regimen. METHODS: Nine baboons received life-supporting heterotopic single-kidney transplants from human decay accelerating factor transgenic pigs. Immunosuppression consisted of GAS (a galactosyl alpha-1,3-galactose analog), cyclosporine, and steroids. Group 1 (n=2) was also treated with CyP and a rapamycin derivative (RAD), group 2 (n=4) received RATS and RAD, and group 3 (n=3) received only RATS. Animals were maintained until death or sacrifice because of uncontrollable rejection or other complications. Graft histopathology was assessed at the study endpoint. RESULTS: Mean survival was 28+/-11.3 days, 23+/-2.5 days, and 20+/-2.5 days for groups 1, 2, and 3 (not significant). Graft rejection was the cause of death in both CyP-treated animals. One RATS-treated animal died of rejection; the others died of infections or bleeding. Two RATS treated animals developed posttransplant lymphoproliferative disorder, and one died of cytomegalovirus pneumonitis. Histopathology revealed severe AHXR in group 1 kidneys, involving 100+/-0% of the tissue examined. In contrast, AHXR was reduced in groups 2 and 3, involving 21+/-14% and 18+/-28%, respectively, of the tissue examined (P<0.01). CONCLUSIONS: Substitution of RATS for CyP was well tolerated and resulted in reduced severity of AHXR in this model. Complications seen in RATS-treated animals may be preventable through the use of standard prophylaxis for infections. Our data suggest that further studies are warranted to explore the use of antilymphocyte agents in xenotransplantation. PMID- 12134096 TI - Anti-lymphocyte function-associated antigen-1 monoclonal antibody inhibits CD40 ligand-independent immune responses and prevents chronic vasculopathy in CD40 ligand-deficient mice. AB - BACKGROUND: Blockade of CD40 ligand (CD40L; CD154, gp39) is a potential treatment for autoimmune disease and allograft rejection. However, CD40L-/- mice are capable of mobilizing cellular immune responses to viral, parasitic, and intracellular bacterial infections as well as rejecting skin grafts with nearly the same efficiency as wild-type mice. CD40L-deficient mice (CD40L-/-) or wild type mice treated with anti-CD40L develop chronic vasculopathy only 8 weeks after allogeneic heart transplantation. To overcome CD40L-independent immune responses, we used anti-lymphocyte function-associated antigen monoclonal antibody (LFA)-1, which has previously been shown to inhibit CD8+ immune responses. METHODS: We conducted mixed lymphocyte reactions, cytotoxicity assays, skin transplantation, and vascularized heterotopic heart transplantation in wild-type B6 and CD40L deficient mice in the presence and absence of anti-LFA-1 to study the effects of anti-LFA-1 in the absence of CD40L signaling. RESULTS: Anti-LFA-1 inhibited proliferation of naive CD40L-/- mixed leukocyte reactions and the lysis of donor targets by CD40L-/- cytotoxic T lymphocytes. Anti-LFA-1-treated CD40L-/- mice that received fully MHC-mismatched skin grafts showed significant prolongation of graft survival, with a median survival time of 55 days (mean 66 days) compared with 13 and 21 days in wild-type and CD40L-/- controls, respectively. CD40L-/- mice that received fully MHC-mismatched vascularized heart transplants treated with four doses of 200 microg of anti-LFA-1 at the time of transplantation did not develop any signs of chronic vasculopathy 150 days after transplantation. CONCLUSION: These results indicate that anti-LFA-1 can complement CD40L inhibition in the prevention of undesirable immune responses. PMID- 12134097 TI - A randomized trial of exercise training after renal transplantation. AB - BACKGROUND: Significant health benefits result from regular physical activity, many which are important for transplant recipients. Although exercise capacity improves initially after transplant, it is not normalized, and only two studies have reported the effects of exercise training in this population. We report a randomized clinical trial of exercise after renal transplantation (RTX). METHODS: One hundred sixty-seven patients were randomized at 1 month after RTX into two groups: exercise intervention (EX) and usual care (UC), with repeat testing at 6 and 12 months. Ninety-five patients completed the following testing at both testing times: symptom-limited treadmill testing with measurement of peak oxygen uptake (peak Vo2); isokinetic muscle testing for muscle strength; and dual-energy X-ray absorptiometry scans for body composition. The SF-36 Health Status Questionnaire assessed self-reported functioning. The exercise intervention consisted of individually prescribed programs to be conducted at home with regular phone follow-up to enhance adherence. Repeated measures analysis of variance was performed to determine differences between the groups for the three testing times. RESULTS: At 1 year 67% of the EX group were exercising regularly compared with 36% of the UC group (P=0.01). Compared with the UC group, the EX group had significantly greater gains in peak Vo2 (P=0.016), percent age predicted Vo2 (P=0.03), and muscle strength (P=0.05), and a trend toward higher self-reported physical functioning (P=0.06). There were no differences between the groups in changes in body composition. At 1 year, peak Vo2 was significantly correlated with age, percent fat, muscle strength, hematocrit, and self-reported physical functioning. CONCLUSIONS: Exercise training after RTX results in higher levels of measured and self-reported physical functioning; however, exercise alone does not affect body composition. PMID- 12134098 TI - Risk factors for cytomegalovirus reactivation after CD6+ T-cell-depleted allogeneic bone marrow transplantation. AB - BACKGROUND: Cytomegalovirus (CMV) infection is a significant cause of morbidity and mortality after allogeneic hematopoietic stem cell transplantation (HSCT). Recipients of T-cell-depleted (TCD) transplants may be more susceptible to CMV infection as a result of the reduction in transferred T cell immunity. We sought to determine the effect of prior donor and patient CMV exposure on the incidence of CMV infection after TCD allogeneic HSCT. METHODS: We retrospectively examined CMV antigen testing results in all patients who had undergone CD6+ TCD related and unrelated donor allogeneic HSCT at our institution from 1996 to 1999. All 124 patients who had documented donor and recipient CMV serologies pretransplant and had undergone CMV antigen testing before day +100 posttransplant were included in the analysis. RESULTS: Forty-one percent of seropositive recipients and 1% of seronegative recipients developed evidence of CMV reactivation (odds ratio 54.1, 95% confidence interval [CI] 6.9-424.1, P<0.001). Prior donor CMV exposure did not place seronegative recipients at increased risk of CMV conversion. Multivariable analysis indicated that prior donor CMV exposure significantly reduced the risk of CMV reactivation in seropositive recipients by 81% (odds ratio 0.19, 95% CI 0.04-0.91, P=0.04). Grades II to IV acute graft-versus-host disease (GVHD) was associated with CMV conversion (P=0.04) when seropositive recipients underwent HSCT from CMV-negative donors, but not when the donor was CMV-seropositive (P=0.54). CONCLUSIONS: The CMV serology status of the recipient, rather than the donor, was the primary determinant of risk for CMV conversion after TCD allogeneic HSCT. Despite CD6+ T-cell depletion, immunity against CMV seemed to be transferred with the donor graft and protected seropositive HSCT recipients from CMV reactivation. PMID- 12134100 TI - Prognostic value of doppler-ultrasonography in hepatic veno-occlusive disease. AB - BACKGROUND: Currently, the severity of veno-occlusive disease (VOD) is graded retrospectively on the basis of the evolution of clinical and biological criteria. The objective of this study was to describe a noninvasive method capable of predicting the severity of this disease at the time of the diagnosis. METHODS: Seventy-one children who developed VOD after intensive myeloablative therapy with busulfan before hematopoietic stem cell transplantation were included in this study. Sixty-four patients underwent baseline ultrasonography (US) and Doppler examination before transplantation. All patients had posttransplantation US and Doppler examinations at the time of the clinical and biological diagnosis of VOD. Seven US morphological criteria and seven Doppler criteria were studied that yielded three individual scores: a US score, a Doppler score, and a total Doppler ultrasonography (DUS) score. RESULTS: In the univariate analysis, three of 7 US criteria, three of 7 Doppler criteria, and the three scores were correlated with the severity of VOD after transplantation and at the time of the diagnosis. In the multivariate analysis, two US criteria (splenomegaly and ascites) and one Doppler criterion (flow recorded in para umbilical vein) were correlated with the severity of VOD. The multivariate analysis of the pooled US and Doppler criteria showed that the flow recorded in the para-umbilical vein was the only criterion significantly associated with the grade of VOD (P=0.0001). All patients with a US-Doppler score >9 developed grade 2 or 3 VOD. CONCLUSION: Postgraft US and Doppler examinations have a prognostic significance according to the grade of VOD. PMID- 12134099 TI - Modified right liver graft from a living donor to prevent congestion. AB - BACKGROUND: Right liver grafts without middle hepatic vein (MHV) drainage reconstruction resulted in severe congestion of the anterior segment (AS) in our early experience of adult-to-adult living donor liver transplantation (LDLT). However, a detailed strategy for preventing such congestion or the necessity of MHV reconstruction has not been discussed in LDLT using a right lobe graft. METHODS: From July 1997 to February 1998, two of five right lobe grafts without MHV drainage reconstruction were complicated with severe congestion of the AS. Thereafter, 42 adult recipients who received right liver grafts with sizable MHV tributaries underwent the reconstruction of MHV drainage. All sizable (>5 mm in diameter) MHV tributaries were preserved during donor hepatectomy and were reconstructed with the recipient's autogenous interposition vein grafts at the bench surgery. The reconstructed vein grafts of this modified right lobe graft were anastomosed to the stump of the MHV and/or left hepatic vein of the recipient after graft revascularization. RESULTS: Serial Doppler ultrasonography, which was regularly checked until 30 days posttransplant, revealed the patent interposition vein graft in 38 of 42 recipients (patency rate 90.5%). In these 38 recipients, no evidence of congestion in the AS was recognized on enhanced computed tomography, while providing enough functioning liver mass comparable to an extended right lobe graft. Also, congestion-related graft injury, such as an infarct of the AS, was not observed in these recipients. CONCLUSIONS: Our early experience indicated the necessity of MHV drainage reconstruction in right lobe grafts, which do not have MHV trunk in certain instances. However, preoperatively, it is difficult to predict the degree of AS congestion of the right liver graft without MHV drainage reconstruction. We suggest aggressive reconstruction of MHV drainage tributaries of the AS, under the circumstances that sizable MHV tributaries are encountered, to prevent possible congestion related complications. PMID- 12134102 TI - Reduction of left ventricular diameter and mass after surgical arteriovenous fistula closure in renal transplant recipients. AB - BACKGROUND: Left ventricular hypertrophy and dilatation is a frequent finding in kidney transplant recipients, which may be favored by the persistent patency of arteriovenous fistula. The purpose of the current study was to prospectively investigate whether surgical closure of the fistula allows reduction of abnormalities of left ventricular morphology in stable renal transplant patients. Furthermore, we studied the ability of preoperative echocardiographic and noninvasive hemodynamic measurements, including the effects of a temporary occlusion of the fistula, to predict postoperative left ventricular diameter and mass reduction. METHODS: Seventeen kidney transplant recipients referred for surgical arteriovenous fistula closure were prospectively studied. Standard echocardiographic parameters, heart rate, and blood pressure were assessed preoperatively at baseline and during an acute pneumatic fistula occlusion. These measurements were repeated 3 to 10 weeks after surgical closure. Six kidney transplant recipients with patent arteriovenous fistulas referred for routine echocardiographic follow-up served as a control group. RESULTS: Surgical fistula closure decreased left ventricular end-diastolic diameter and mass indexes (29.9+/-2.4 to 27.4+/-2.1 mm/m2, P<0.001, and 141+/-37 to 132+/-39 g/m2, P<0.05, respectively), whereas no changes were seen in controls after a similar delay. Postoperative left ventricular end-diastolic diameter and mass reductions correlated best with the increases in total peripheral resistance (r=0.85, P<0.0001) and mean arterial blood pressure (r=0.64, P=0.006) during pneumatic occlusion, respectively. CONCLUSIONS: Surgical closure of arteriovenous fistula reduces left ventricular diameter and mass in kidney transplant recipients. Increases in blood pressure and total peripheral resistance induced by temporary fistula occlusion are the best predictors of these morphological changes. PMID- 12134101 TI - Salt-sensitivity of blood pressure and decreased 11beta-hydroxysteroid dehydrogenase type 2 activity after renal transplantation. AB - BACKGROUND: High blood pressure (BP) predicts a poor long-term kidney graft outcome. The mechanisms for hypertension in renal graft recipients are only partly understood. There is evidence that BP is salt dependent in renal transplant recipients. We hypothesize that renal transplantation induces salt sensitivity by decreasing 11beta-hydroxysteroid dehydrogenase type 2 (11betaHSD2) activity. METHODS: A syngenic uninephrectomized rat transplantation model (Lewis to Lewis) (n=7) was used to demonstrate salt sensitivity after transplantation. Sham-operated (n=5) and denervated rats (n=5) were used as controls. In all rats, BP was measured continuously by telemetry 24 hr a day, whereas the rats were set successively on a normal- (0.45% NaCl), high- (8% NaCl), low- (0.1% NaCl), and, again, normal-salt (0.45% NaCl) diet during a 6-day period to assess salt-related changes in mean arterial pressure (MAP). 11betaHSD2 activity was assessed by determining the ratio of corticosterone to dehydrocorticosterone metabolites (THB+5alphaTHB)/THA in urine. RESULTS: After uninephrectomy and implantation of the telemetry device, MAP was comparable in rats assigned to undergo sham operation (100+/-3 mmHg), denervation (105+/-5 mmHg), or transplantation (102+/-6 mmHg). When animals were switched from the normal- to high-salt diet, the increase in MAP was more pronounced in the transplanted group (13.9+/-5.1 mmHg) than in those undergoing sham operation (5.1+/-1.7 mmHg, P<0.004) or denervation (7.1+/-1.8 mmHg, P<0.021). Urinary (THB+5alphaTHB)/THA increased more than 2-fold in the transplanted rats but remained stable in the sham-operated and denervated animals (P<0.0001), indicating reduced activity of 11betaHSD2. CONCLUSION: Syngenic renal transplantation causes salt sensitivity with increased BP associated with a reduced activity of 11betaHSD2. PMID- 12134103 TI - Successful management of an ABO-mismatched lung allograft using antigen-specific immunoadsorption, complement inhibition, and immunomodulatory therapy. AB - BACKGROUND: Successful management of an ABO-mismatched lung allograft recipient has not previously been described. METHODS: Because of a clerical error, a 67 year-old blood type B patient with idiopathic pulmonary fibrosis received a left single-lung allograft from a blood type A donor. Cyclophosphamide was added to immunosuppression with anti-thymocyte globulin induction, cyclosporine, mycophenolate mofetil, and prednisone. When increasing anti-A antibody titers were detected, antigen-specific immunoadsorption, anti-CD20 monoclonal antibody, and recombinant soluble complement receptor type 1 (TP10) were administered. RESULTS: Rising anti-A antibody titers were reduced acutely by immunoadsorption, and remained low during long-term follow-up. Humoral injury to the graft was not detected. Acute cellular rejection and multiple complications were successfully managed. Three years after transplantation the patient is clinically well on stable maintenance immunosuppression and prophylactic photochemotherapy. CONCLUSIONS: Modulation of anti-A antibody, preserved graft function, and a favorable patient outcome can be achieved for an ABO-mismatched lung allograft. PMID- 12134104 TI - A double-hand transplant can be worth the effort! AB - BACKGROUND: Composite-tissue transplantation offers a new therapeutic option for patients with loss of a hand. Little is known, however, about the long-term outcome after such a transplant with regard to graft function and immunosuppression and its side effects. We here report on our experience with a double-hand transplant performed more than 18 months ago. METHODS: Both distal forearms and hands of an age-, gender-, and size-matched cadaveric donor were transplanted to a 47-year-old policeman 6 years after loss of both hands. He received antithymocyte globulin as induction therapy and tacrolimus, mycophenolate mofetil, and prednisone as maintenance immunosuppression. Ganciclovir and co-trimoxazole were given prophylactically for cytomegalovirus and Pneumocystis carinii infection. A special rehabilitation program based mainly on cognitive therapy was designed and continued for 1 year. RESULTS: Apart from a small area of skin that became necrotic early and some arteriovenous fistulas in the left forearm, which required ligation 6 months after transplantation, there were no surgical complications. One acute rejection episode occurred on day 55 and resolved completely after high-dose steroids and topical tacrolimus. Despite ganciclovir prophylaxis, virus replication was observed. The patient became negative for cytomegalovirus only after additional treatment with foscarnet (Foscavir) and cidofovir. At the end of 18 months, graft function with regard to motility is overall 60% of normal and enables the patient to pursue activities he could not with his myoelectric prostheses. CONCLUSIONS: Excellent long-term results can be achieved with double-hand transplantation. Prerequisites are an appropriate surgical technique, careful immunosuppression, and an extensive rehabilitation program. PMID- 12134105 TI - Detection of chronic allograft nephropathy by quantitative doppler imaging. AB - BACKGROUND: Chronic allograft nephropathy (CAN) is the major cause of graft loss, and early detection is desirable to avoid irreversible graft damage. We have evaluated a new technique of color Doppler quantification using Cineloop (Philips Medical Systems, Bothell, WA) imaging for noninvasive diagnosis of CAN. METHODS: Provisional normal ranges were defined by pilot study (n=13) and prospectively tested in stable recipients in whom CAN was independently quantified by contemporaneous histology (n=67), using the Banff schema. RESULTS: The maximal fractional area (MFA, systolic color pixels/total area) was 28.7+/-9.7% in normal subjects and reduced to 18.8+/-8.0% in grade 1 and 12.5+/-6.4% in grade 2 CAN (both P<0.001). The minimum color fractional area was reduced from 10.3+/-5.3% in normal subjects to 3.1+/-2.6% in grade 2 CAN (P<0.001), but was less useful. Distance from peripheral color pixels to capsule increased in CAN grade 2 versus 0 (6.0+/-1.6 vs. 3.9+/-1.0 mm, respectively; P<0.001). Calcineurin inhibitor nephrotoxicity reduced MFA (18.0+/-9.3 vs. 26.9+/-10.7%; P<0.001) and other dynamic measurements. Parenchymal damage exerted minimal effect on resistance index, mean variance, and peak Doppler velocity. MFA (cutoff<17.3%) can diagnose CAN (sensitivity 69%, specificity 88%, positive predictive value 86%) and severe CAN (sensitivity 87%, specificity 71%, negative predictive value 95%). Distance to capsule >5 mm was less sensitive (49%) but more specific (91% alone, and 97% combined with MFA). CONCLUSIONS: In conclusion, quantitative Doppler ultrasound can reliably detect CAN and, although imperfect at correctly grading, allows recognition of significant tubulointerstitial damage for initiation of a confirmatory needle core biopsy. PMID- 12134106 TI - Heme oxygenase-1 gene transfer inhibits inducible nitric oxide synthase expression and protects genetically fat Zucker rat livers from ischemia reperfusion injury. AB - BACKGROUND: Ischemia/reperfusion (I/R) injury is a critical factor in the dysfunction of steatotic orthotopic liver transplants. Heme oxygenase-1 (HO-1), a cytoprotective protein, may be important in ameliorating hepatic I/R injury. METHODS: We used adenovirus (Ad)-based HO-1 gene transfer to analyze the effects of HO-1 overexpression in a well-established fatty Zucker rat model of I/R followed by orthotopic liver transplantation. RESULTS: Ad-HO-1 gene therapy increased recipient survival (80% vs. 40-50% in controls) and significantly diminished hepatocyte injury, as compared with untreated and Ad-beta galactosidase (Ad-beta-Gal)-treated livers. Orthotopic liver transplants in the Ad-HO-1 group exhibited less macrophage infiltration in the portal areas, as compared with controls. Unlike untreated and Ad-beta-Gal-treated orthotopic liver transplant controls, which showed elevated levels of inducible nitric oxide synthase by infiltrating macrophages, inducible nitric oxide synthase expression in the Ad-HO-1 group was almost absent. In contrast, endothelial nitric oxide synthase was comparable in Ad-HO-1- and Ad-beta-Gal-transduced fatty orthotopic liver transplants. Intragraft expression of antiapoptotic Bcl-2 and Bag-1 was increased in Ad-HO-1-treated orthotopic liver transplants, as compared with Ad beta-Gal controls. Moreover, increased HO enzymatic activity was accompanied by inhibition of caspase-3 protein expression. CONCLUSIONS: HO-1 gene transfer significantly prolongs survival of steatotic orthotopic liver transplants, depresses macrophage infiltration, suppresses local expression of inducible nitric oxide synthase, and modulates pro- and antiapoptotic pathways. PMID- 12134108 TI - Retroviral delivery of transforming growth factor-beta1 to myeloid dendritic cells: inhibition of T-cell priming ability and influence on allograft survival. AB - BACKGROUND: Transforming growth factor (TGF)-beta inhibits the maturation and function of antigen-presenting cells. Our purpose was to evaluate the impact of retroviral delivery of human TGF-beta1 to murine myeloid dendritic cell (DC) progenitors on (i) their in vitro properties, (ii) their in vivo function, and (iii) their influence on organ allograft survival. METHODS: C57BL10 (B10; H2b) bone marrow cells were lineage depleted and stimulated with granulocyte macrophage colony-stimulating factor for 6 days. Replicating DC progenitors were transduced on days 2, 3, and 4 of culture by ecotropic retrovirus encoding human TGF-beta1 using centrifugal enhancement. Secretion of TGF-beta1 and other cytokines was quantified by enzyme immunoassay. Allogeneic C3H/HeJ (C3H; H2k) T cell proliferative responses and generation of cytotoxic T lymphocytes in mixed leukocyte reaction were determined by [3H]thymidine incorporation and 51Cr release assays, respectively. DC migration was analyzed by immunohistochemistry, and their impact on survival of intra-abdominal heart transplants was determined. RESULTS: Maximal TGF-beta1 transduction efficiency was 60%. The TGF-beta transduced DC showed pronounced impairment (>80%) of T-cell allostimulatory activity in vitro. After their IV injection, B10 TGF-beta-transduced DC (IAb+) were detected in T-cell areas of spleens of allogeneic C3H recipients. Splenic T cell responses to donor alloantigens of mice that received TGF-beta-transduced DC were severely impaired. This was accompanied by marked inhibition of interleukin 2 and interferon-gamma production in response to restimulation with donor alloantigen. Survival of B10 cardiac allografts in C3H mice given B10 TGF-beta transduced DC (2x106 IV, 7 days before transplantation), was extended modestly but significantly. CONCLUSION: Retroviral transduction of myeloid DC progenitors to overexpress TGF-beta is associated with marked impairment of their T-cell allostimulatory activity but with only modest prolongation of organ allograft survival. PMID- 12134107 TI - Early passenger leukocyte migration and acute immune reactions in the rat recipient spleen during liver engraftment: with particular emphasis on donor major histocompatibility complex class II+ cells. AB - After a short course of tacrolimus, Lewis rat liver allografts induce donor specific nonreactivity in Brown Norway recipients that is immunosuppression independent after 28 days. To clarify the role of donor major histocompatibility complex (MHC) class II+ cells, we investigated the migration to the recipient splenic T- and B-cell compartments of different subsets of Lewis MHC class II+ passenger leukocytes. The rise and decline of immune activation were monitored in the hepatic allograft and in the host spleen by analyses of BrdU+ (proliferating) leukocytes, TUNEL+ (apoptotic) cells, apoptosis-associated molecules, TH1/TH2 cytokine profiles, and histoimmunocytochemical examination of graft and splenic tissues. Serial flow cytometry studies during the 28-day period of drug-assisted "hepatic tolerogenesis" showed that migratory MHC class II+ cells accounted for less than half of the donor cells in the host spleen. The class II+ cells consisted mostly of B cells that homed to splenic B-cell follicles with only a sparse representation of dendritic cells that were exclusively found in the splenic periarteriolar lymphoid sheath. In parallel studies, transplantation of the less tolerogenic heart produced a diminutive version of the same events, but with far fewer donor cells in the host spleen, evidence of sustained immune activation, and the development of chronic rejection by 100 days. The data are consistent with the paradigm that migration of donor leukocytes is the prime determinant of variable tolerance induction induced by transplantation of the liver and other organs, but without regard for donor MHC class II+ expression. PMID- 12134109 TI - Host-derived enterocytes in intestinal grafts. AB - Replacement of donor lymphoid tissue by lymphocytes of recipient origin is an established phenomenon in small bowel transplants. However, replacement of donor epithelial cells of bowel grafts by host cells has not been demonstrated. The objective of our study was to determine whether donor enterocytes are replaced by host-derived enterocytes in the intestinal allograft. Graft biopsy specimens, obtained from five human male recipients of female intestine, were examined for the presence of male enterocytes. The biopsies dated from 90 to 770 days posttransplant. Formalin-fixed 3-microm specimen sections were stained for X and Y chromosomes by fluorescent in situ hybridization technique. Fluorescent microscopy of the stained sections identified male enterocytes in four patients, with a percentage of male cells ranging from 0.09% to 0.26% of the total enterocyte mass. Using the fluorescent in situ hybridization technique, we demonstrated the presence of host-derived male (XY) enterocytes in the female intestinal graft. PMID- 12134110 TI - Early enteral supply of lactobacillus and fiber versus selective bowel decontamination: a controlled trial in liver transplant recipients. AB - BACKGROUND: Early enteral nutrition with solutions containing prebiotics (fiber) and probiotics (Lactobacillus) is suggested to reduce bacterial translocation and minimize the incidence of infections after liver transplantation. METHODS: In a prospective, randomized placebo-controlled trial consisting of 95 patients, we compared the incidence of postoperative infections and other complications after liver transplantation among three different groups, all supplied with early enteral nutrition: (a) standard formula plus selective bowel decontamination (SBD), (b) fiber-containing formula plus living Lactobacillus plantarum 299, and (c) fiber-containing formula plus heat-killed L plantarum 299. RESULTS: The groups were comparable regarding preoperative American Society of Anesthesiologists classification, Child-Pugh classification of cirrhosis, operative data, and degree of immunosuppression. The patients who received living lactobacilli plus fiber developed significantly fewer bacterial infections (13%) than the patients with SBD (48%). The incidence of infections was 34% in the group with inactivated lactobacilli and fiber. Cholangitis and pneumonia were the leading infections and enterococci the most commonly isolated bacteria. In the living Lactobacillus group, the mean duration of antibiotic therapy, the mean total hospital stay, and the stay on the intensive care unit were also shorter than in the groups with inactivated lactobacilli and fiber as well as with SBD. However, these differences did not reach statistical significance. CONCLUSIONS: Early enteral nutrition with fiber-containing solutions and living L plantarum 299 was well tolerated. It decreases markedly the rate of postoperative infections both in comparison with inactivated L plantarum 299 and significantly with SBD and a standard enteral nutrition formula. As it is a cheap and feasible alternative to SBD, further studies should evaluate whether this ecoimmunonutrition should be already started while patients are on the waiting list for transplantation. PMID- 12134111 TI - Epithelioid hemangioendothelioma of the liver disseminated to the peritoneum treated with liver transplantation and interferon alpha-2B. AB - Epithelioid hemangioendothelioma (EH) is a rare, low-grade, malignant neoplasm of vascular origin that may develop at different sites, such as in the soft tissue, lungs, or liver. We report the case of a 21-year-old female with primary EH of the liver treated by liver transplantation and review the available literature on EH. This patient developed symptomatic recurrence of the tumor in the pelvis 2 months posttransplant. Treatment with interferon alpha-2b resulted in substantial regression of the pelvic metastases and alleviation of symptoms, but the patient developed graft rejection and died of associated complications 16 months posttransplant. This report is the first showing the efficacy of interferon in this setting. PMID- 12134112 TI - Severe pancytopenia and hemophagocytosis after HHV-8 primary infection in a renal transplant patient successfully treated with foscarnet. AB - We report the occurrence of human herpesvirus (HHV)-8 primary infection in an adult male kidney recipient. Four months after transplantation, the patient developed visceral Kaposi sarcoma, and 1 month later he presented with progressive and severe peripheral cytopenia, in the presence of a normocellular or hypercellular bone marrow (BM) with hemophagocytosis. HHV-8 was the sole pathogen detected by polymerase chain reaction either in the serum or in the BM. HHV-8 latent nuclear antigen was detected in immature progenitor cells from the BM. Immunosuppressive therapy was reduced, and the patient was treated with foscarnet for 2 weeks, leading to a dramatic normalization of blood cell counts, concomitantly with the disappearance of HHV-8 viremia. At the end of antiviral therapy, the patient received chemotherapy, and Kaposi sarcoma regressed in 2 months. Severe peripheral cytopenia may be a posttransplant complication after HHV-8 infection, for which treatment with foscarnet seems appropriate. PMID- 12134113 TI - Association of the CD134/CD134L costimulatory pathway with acute rejection of small bowel allograft. AB - BACKGROUND: The CD134/CD134L interaction provides a strong costimulatory signal that is CD28-independent. The CD134/CD134L pathway has been studied in inflammatory, autoimmune diseases, and graft-versus-host disease, but no information exists on the involvement of CD134/CD134L interactions in solid organ transplantation. METHODS: CD134/CD134L costimulation was studied in a rat model of small bowel transplantation. Immunohistochemistry and flow cytometry were used to analyze the expression and localization of CD134/CD134L. Mixed lymphocyte culture and quantitative RT-PCR were used to measure the effect on T proliferation and T helper cell cytokine transcripts of single or combined CD134 and CD28 costimulatory blockade. RESULTS: CD134 and CD134L molecules were strongly expressed in acutely rejected small bowel allografts. This expression was significantly suppressed by FK506. Interruption of the CD134 and CD28 costimulatory pathways resulted in an pronounced increase in expression of interleukin-10 transcripts. CONCLUSION: These results present the first evidence that the CD134/CD134L interaction is associated with acute allograft rejection. Combined CD134/CD134L blockade may be an effective treatment to both prevent acute allograft rejection and promote the induction of cells with regulatory capacity. PMID- 12134114 TI - Response to "Sustained improvement of renal graft function for two years in hypertensive renal transplant recipients treated with nifedipine as compared with lisinopril". PMID- 12134115 TI - Diabetic kidneys can safely expand the donor pool. AB - BACKGROUND: The demand for organs has increased exponentially with a new name added to the United States waiting list every 16 min. As such, kidneys from medically marginal donors are being considered for transplantation more frequently, including kidneys from individuals already at risk for renal disease, e.g., diabetic donors. METHODS: We compared outcomes when using kidneys from donors with type 1 diabetes mellitus (D1) or type 2 diabetes mellitus (D2) at our center as a function of time. All patients with available data who underwent renal transplantation were evaluated (n=2013). RESULTS: Forty-two individuals were recipients of D1 or D2 donor kidneys. Thirty of these individuals did not have diabetes (R0). All patients received quadruple sequential immunosuppression with cyclosporine (CsA) or tacrolimus (FK506). Donor serum creatinine (Scr) values were not significantly different. D2 kidneys came from older donors (mean age, 56+/-10.4 years; P< or =0.01 vs. D1 and D0 donors). Mean discharge Scr was greater in nondiabetic D2 recipients (D2/R0, 2.45+/-1.3 mg/dl; P=0.0016 vs. D0/R0), and transplantation of D1 or D2 kidneys was associated with a significantly increased frequency of posttransplant proteinuria (P=0.0089). Interestingly, R0 recipients of D1 or D2 kidneys were more likely to initiate oral hypoglycemic therapy after transplant (P=0.04). However, rejection episodes were not significantly different among groups, and long-term graft survival and patient survival were similar among groups. CONCLUSIONS: These data suggest that diabetic kidneys can be safely used without risk to patient or graft survival. Preexisting diabetic injury in the donor may increase the risk for proteinuria, compromised renal function, and posttransplant glucose intolerance. PMID- 12134116 TI - Right ventricular myocardial infarction: when the power fails. AB - Learn how to recognize this common complication of an acute inferior or posterior wall myocardial infarction and to give patients the correct treatment. PMID- 12134117 TI - Thyroid diseases: a primer for the critical care nurse. AB - Thyroid diseases are common. This article will discuss the types, signs and symptoms, medical management, and nursing care of thyroid disease. PMID- 12134118 TI - Atrio-ventricular nodal reentry tachycardia. AB - Reentry variants are the most common mechanism for supraventricular arrhythmias. Learn how they originate, how they are differentiated, and how they are treated. PMID- 12134119 TI - Critical care nurses' role in code status discussion. AB - Critical care nurses have an important role in the decision-making process concerning end-of-life and resuscitation situations. This article provides insight into this process. PMID- 12134120 TI - Early recognition of client status changes: the importance of time. AB - Early recognition of client status changes prevents or minimizes complications and reduces client morbidity, yet little is known about this phenomenon in nursing. Using a qualitative interpretive approach, this study investigated how expert critical care nurses are able to recognize and intervene in the early stages of client problems. The findings revealed a temporal unity in critical care nurses' thinking that informs their early recognition experiences. PMID- 12134122 TI - Phases of a clinical trial. AB - Clinical trials are commonly conducted at hospitals, clinics, physicians' offices, universities, and in many other areas. This article describes the four phases of a clinical trial for a new medication, device, or treatment. This article will assist nurses caring for patients in clinical trials. PMID- 12134123 TI - The creation, implementation, and evaluation of a nurse residency program through a shared leadership model in the intensive care setting. AB - This education strategy was used at a 404-bed level-one trauma center where 60% of the beds are for critical care. In June 1999, 93 nurses underwent critical care orientation. By June 2000, only 8 of these nurses remained. Exit interviews revealed the primary reason for leaving was dissatisfaction with orientation. The Education Department undertook a shared leadership approach to identify strategies for resolution of the perceived orientation problem. The Preceptor Leadership Council was formed to create a Nurse Residency Program for new nurses. Membership consisted of staff nurse lead preceptors and clinical educators. Outcomes following the implementation of the program indicated an overall increase in satisfaction with nursing orientation in the intensive care unit as well as a significant reduction in turnover rates. Strategies used to create, implement, and evaluate the program are presented. PMID- 12134124 TI - A closing word: episodes in nursing: a guest editorial. PMID- 12134125 TI - Burning mouth and saliva. AB - Stomatodynia is the complaint of burning, tickling or itching of the oral cavity, and can be associated with other oral and non-oral signs and symptoms. However, the oral mucosa often appears normal, with no apparent underlying organic cause to account for the symptomatology. The etiology is unknown, though evidence points to the participation of numerous local, systemic and psychological factors. Among the local factors, saliva may play an important role in the symptoms of burning mouth. Saliva possesses specific rheological properties as a result of its chemical, physical and biological characteristics - these properties being essential for maintaining balanced conditions within the oral cavity. Patients with burning mouth present evidence of changes in salivary composition and flow, as well as a probable alteration in the oral mucosal sensory perception related particularly to dry mouth and taste alterations. On the other hand, alterations in salivary composition appear to reflect on its viscosity and symptomatology of burning mouth. Saliva is a field open to much research related to burning mouth, and knowledge of its properties (e.g., viscosity) merits special attention in view of its apparent relationship to the symptoms of burning mouth. The present study describes our clinical experience with burning mouth, and discusses some of the aspects pointing to salivary alterations as one of the most important factors underlying stomatodynia. PMID- 12134126 TI - Peripheral giant-cell granuloma. Review of 13 cases. AB - Peripheral giant-cell granuloma is an infrequent exophytic lesion of the oral cavity, also known as giant-cell epulis, osteoclastoma, giant-cell reparative granuloma, or giant-cell hyperplasia. The present study reports 13 cases on patients that visited the Oral Medicine department of the Dental Faculty in the Santiago de Compostela University. We report the location, size, course and treatment of each lesion, comparing the results obtained to those reported in the literature. We discuss differential diagnosis with respect to other entities, in particular brown tumor of hyperparathyroidism, cherubism, and aneurysmal bone cyst, all of which show very similar histological appearance to peripheral giant cell granuloma. PMID- 12134127 TI - Antifungal topical therapy in oral chronic candidosis. A comparative study. AB - The aim of the present paper was to evaluate the effect of fenticonazole and to compare it with that of ketoconazole and nystatin in the topical treatment of oral chronic candidosis. Eighty patients diagnosed with erythematous chronic candidosis were divided into four groups, according to the cream they were provided with 3% fenticonazole, 2% fenticonazole, 100000UI nystatin, and 2% ketoconazole in orabase respectively. A clinical assessment was made at 7, 15, 30 and 45 days. Fifty-one of them finished the trial. ANOVA, Kruskall Wallis and Wilcoxon tests were applied for statistical analysis. There was a significative decrease of the oral lesions in all groups (p 0.0001). The remission grade was also analysed according to the localisation: lesions in the buccal mucosa and in the comissure all the patients achieved complete resolution, whereas tongue and palate lesions showed a significative decrease in all the treatments (p 0.00001). Fenticonazole proved to be as effective as nystatin and ketoconazole in topical treatment of oral candidosis. PMID- 12134128 TI - Gingival disorders of immune origin. AB - Loss of the regulatory control of the immune and inflammatory defense mechanisms of the body can lead to host tissue damage, as a result of a series of complex mechanisms. In the oral cavity, and particularly in the gingival region, these dysfunctions can manifest in association to some background systemic disorder either as lesions confined to the mouth or as lesions heralding posterior florid manifestations. A review is made of the clinical changes, possible immunopathological mechanisms and diagnostic and management options in relation to pemphigus vulgaris, benign mucous membrane pemphigoid, cicatricial pemphigoid and lichen planus. PMID- 12134130 TI - General and dental characteristics of Bloch-Sulzberger syndrome. Review of literature and presentation of a case report. AB - A case report of Bloch-Sulzberger Incontinentia Pigmenti is presented, in which the dental symptoms are considered as a base, not just for a review of this disorder within the scope of Integrated Paediatric Dentistry, but also in order to establish an adequate dental treatment for the affected children. There should be interdisciplinary coordination, leading to better treatment and therefore an improved quality of life for these patients. PMID- 12134129 TI - Quantitative relationship between salivary level of Streptococcus mutans and Candida albicans in children with Down's syndrome. AB - OBJECTIVE: The actual work intends to establish a correlation between quantitative counts for Streptococcus mutans and Candida albicans in the saliva of patients with Down syndrome (DS), and in mentally deficient (MD) patients, with respect to a normal group (C ). DESIGN OF THE STUDY: This study was performed in 166 children (49 DS, 60MD and 57C), whose saliva samples were grown on TYCSB agar and Sabouraud agar. Subsequently, microbiological analysis, scanning electron microscopy and aggregation assays were made. The results were statistically evaluated using the variance analysis (ANOVA) and the Student "t" test for unpaired data. RESULTS: A correlation of 0.45 was found for group C, 0.55 in DS patients and 0.37 in MD patients, when comparing counts of S. mutans and C. albicans in saliva. Scanning electron microscopy analysis showed coaggregation between S. mutans and C. albicans and this was confirmed by in vitro experiments of S. mutans and C. albicans made on nitrocellulose filters. CONCLUSIONS: This kind of association between bacteria and fungi contributes to justify a pathogenic synergy among several microorganisms, as well as some infectious local or systemic manifestations, depending on the immunity status of the patient. PMID- 12134131 TI - Buccodental pathology in patients with insulin-dependent diabetes mellitus: a clinical study. AB - OBJECTIVE: A study is made to determine whether patients with type 1 (insulin dependent) diabetes mellitus (IDDM) suffer oral complications attributable to the disease, or whether some disorder of the oral cavity can be regarded as pathognomonic of diabetes. MATERIAL AND METHODS: Thirty juvenile diabetics and 30 healthy individuals were evaluated for dental caries and oral mucosal lesions, with the performance of basal and stimulated sialometry in all cases, to assess possible alterations in salivary flow. In addition, an study of periodontal variables was made such as the presence of bacterial plaque, gingival status and attachment losses. RESULTS AND CONCLUSIONS: The diabetics were found to have significantly greater periodontal attachment loss, even though oral hygiene was significantly better among these patients. There were no differences between the two groups in terms of the number of caries, the presence of mucosal lesions or salivary flow. PMID- 12134132 TI - Alopecia areata of dental origin. AB - The association of alopecia areata and infectious foci of dental origin is relatively common, and may be explained by the autoimmune nature of the disorder. We describe a case of alopecia areata with no apparent cause and that was effectively resolved by eliminating a focalized dental infection via endodontic treatment. The presence of common immune mediators in the pathogenesis of both alopecia areata and dental infection could account for the dental origin of the hair loss. In this sense, patients with localized alopecia should be subjected to careful exploration of the oral cavity in search of possible dental infections. PMID- 12134133 TI - Calcifying epithelial odontogenic tumor (Pindborg tumor). AB - We report a clinical case of a 41-year-old female patient affected by a Pindborg tumor located in the left mandibular angle. The accidental detection of the lesion and its frequent relationship with an impacted tooth can difficult its differential diagnosis, mainly with an odontogenic cyst. In spite of the existence of a recurrence rate of 14% after conservative treatment, we think that a quality treatment would require the practice of a block excision including healthy bony margins. PMID- 12134135 TI - Analysis of renal handling of radiopharmaceuticals. AB - Renal excretion is the preferable elimination pathway of radiopharmaceuticals and/or their metabolites from the body. The mechanisms of renal excretion involve glomerular filtration, tubular secretion, tubular reabsorption, and kidney metabolism. Radiopharmaceuticals of a molecular weight of up to 60 kDa are ultrafiltrated in the glomeruli at a rate dependent on their protein binding (the rate of glomerular filtration is a product of glomerular filtration rate and free fraction of the agent in plasma). Exclusive excretion by this route is typical for many chelates such us 99mTc-DTPA (diethylenetriaminepentaacetic acid). Some other radiopharmaceuticals are excreted in the renal tubuli into the tubular fluid by carrier-mediated and active processes involving different transport systems. Examples are ortho-iodohippurate and 99mTc-MAG-3 (mercaptoacetyltriglycine). Tubular reabsorption involves either passive diffusion of lipid-soluble radiopharmaceuticals from the glomerular filtrate back into blood or active, carrier-mediated process. Kidney metabolism and consequent renal uptake of the catabolites is of an outstanding importance for possible therapeutic applications of radiolabeled antibody fragments and peptides. These agents are partially reabsorbed from the ultrafiltrate by the cells of the renal proximal tubules by means of pinocytosis and subsequently degraded in the lysosomes. It limits their therapeutic use due to the potential radiation nephrotoxicity. For the analysis of elimination mechanisms of radiopharmaceuticals in the kidney, different approaches at various experimental levels (the kidney perfusion technique, isolated, functionally intact renal tubules, and isolated membranes) could be successfully employed. PMID- 12134134 TI - Polyethylene glycol in the design of tumor-targetting radiolabelled macromolecules -- lessons from liposomes and monoclonal antibodies. AB - Radiolabelled macromolecules such as liposomes and monoclonal antibodies (Mab) are attractive agents for tumour-targetting studies. In addition to their potential diagnostic role, they can also provide vital information on the targetting capacity of therapeutic agents. Certainly in the case of liposome development, this ability to track the pharmacokinetics and biodistribution of the agents in a non-invasive fashion has assisted the design and application of therapeutic liposomal agents. A significant limitation of unmodified liposomes and Mab is their tendency to be cleared rapidly from the circulation. The use of polyethylene glycol (PEG) in the formulation of these agents has the capacity to alter their biological behaviour in such a way as to improve their ability to target tumours. In this paper we review the data relating to the use of PEG modified liposomes and Mab in the context of nuclear medicine studies. PMID- 12134136 TI - The influence of chelator on the pharmacokinetics of 99mTc-labelled peptides. AB - Because of the ideal properties of technetium-99m for imaging, more methods have been developed for labelling peptides and other biomolecules with this radionuclide that any other. While few detailed comparative studies have been performed, it has become apparent that the use of different labelling procedures can exert a profound effect on the pattern of biodistribution after intravenous administration of the radiotracer. The most significant influence of the labelling method is on the rate and route of excretion of the radionuclide. While some procedures tend to direct excretion towards the hepatobiliary route, others tend to favour renal clearance. Although the factors which exert this influence are still not fully understood, it is clear that the charge, lipophilicity and stability of the technetium-peptide complexes play major roles. A greater understanding of these factors will allow the development of improved radiotracers which demonstrate improved targeting potential as a result of lower uptake and consequent radiation dose by normal tissues. PMID- 12134137 TI - Altering pharmacokinetics of radiolabeled antibodies by the interposition of metabolizable linkages. Metabolizable linkers and pharmacokinetics of monoclonal antibodies. AB - Monoclonal antibodies, their fragments and low molecular weight oncophilic molecules such as synthetic somatostatin derivatives have been used to deliver radioactivity to target cells for both diagnostic and therapeutic purposes. Clinical studies demonstrated high abilities of the radiolabeled antibodies and peptides for nuclear medicine applications. However, high and persistent localization of radioactivity was observed in the liver or kidney especially when these molecules are labeled with metallic radionuclides, which reduce diagnostic accuracy and compromise therapeutic effectiveness. Thus, radiolabeled antibodies and peptides would become much more useful in both targeted imaging and radiotherapy if the undesirable radioactivity localization can be diminished. As a means to reduce the undesirable radioactivity, interposition of a metabolizable linkage between an antibody and a radiolabel was proposed to generate radiolabeled small molecules of urinary excretion from the parental antibody by enzymatic cleavage of the linkage. In this paper, after indicating the rationale behind the radiopharmaceutical design, a significant role played by the interposition of the metabolizable linkage in altering pharmacokinetics of radiolabeled antibodies is described from a variety of studies so far reported with an emphasis being laid on the importance of radiometabolite-based design of metabolizable linkages. PMID- 12134138 TI - The influence of carrier on 99mTc radiopharmaceuticals. AB - High specific activity 99mTc-labelled radiopharmaceuticals are required in order to avoid saturating receptor sites and to minimise pharmacologic or toxic effects. The specific activity of 99mTc-pertechnetate is maximised by use shortly after elution from a generator which had been eluted at frequent intervals. Effective specific activity can be maximised by a variety of means. Often is it possible to label a very small amount of precursor with a large amount of 99mTc and use the product without further purification; this is limited by the potency of the ligand and the efficiency of labelling, which in turn is affected by the choice of chelator. A variety of purification techniques have been used, ranging from solvent extraction, solid-phase extraction cartridges, and size-exclusion columns to high-pressure liquid chromatography. Excess unchelated thiol containing ligands (e.g. N2S2, N3S) can be removed by a thiol-trapping resin. Finally, solid-phase synthesis, in which the precursor is immobilised on a solid support (resin or gold) and only released into solution during chelation of 99mTc, is a promising method. High specific activity will become increasingly important with the next generation of 99mTc radiopharmaceuticals. PMID- 12134139 TI - The influence of chain length and base sequence on the pharmacokinetic behavior of 99mTc-morpholinos in mice. AB - BACKGROUND: Despite in vivo use now over several years, in particular for nuclear medicine imaging, the influences on pharmacokinetics of chain length and base sequence of radiolabeled oligomers has not been investigated. METHODS: As test oligomer, morpholinos (MORFs), a DNA analogue, were radiolabeled with 99mTc via MAG3 and the pharmacokinetics in normal mice determined for 3 chain lengths (15, 18 and 25 mer) and 2 base sequences (MORF and its complement cMORF). In addition, LS174T-tumor bearing nude mice received the anti-CEA antibody MN14 (Immunomedics) conjugated either with MORF15 or MORF18 and subsequently received 99mTc-labeled cMORF15 or cMORF18 respectively in a pretargeting strategy. RESULTS: In normal mice, after 1 hr, regardless of chain length or sequence, all labeled MORFs and cMORFs accumulated only slightly in all tissues (e.g. at 3 hr <0.15 ID%/g) except in kidneys. Besides being excessive, the kidneys were the only tissue with levels dependent upon chain length (e.g. at 1 hr, 5, 7 and 22 ID%/g for MORF15, 18 and 25, respectively) and sequence (e.g. at 3 hrs 9 ID%/g for MORF25 and 21 ID%/g for cMORF25). Identical biodistribution trends were observed in tumored mice with all tissues including tumor showing levels independent of chain length or base sequence except for kidneys. Furthermore, while all other tissues cleared in the interval from 1-3 hrs, kidney levels remained constant in both normal and tumored animals. Largely because of these differences in kidneys, images obtained by pretargeting with 99mTc-cMORF15 were superior compared to 99mTc-cMORF18 (images of control animals not receiving the antibody showed no tumor at all). CONCLUSIONS: Judged by radiolabel accumulations in tissue, the pharmacokinetics of 99mTc labeled morpholinos were independent of chain length and base sequence. The only obvious exception was kidneys in which accumulations were significantly higher for the longer chain lengths and significantly different for cMORF vs MORF. These results show that chain length and base sequences may be varied to alter the pharmacokinetics of radiolabeled oligomers in nuclear medicine imaging studies. PMID- 12134140 TI - X-ray snapshots of quinone cofactor biogenesis in bacterial copper amine oxidase. AB - The quinone cofactor TPQ in copper amine oxidase is generated by posttranslational modification of an active site tyrosine residue. Using X-ray crystallography, we have probed the copper-dependent autooxidation process of TPQ in the enzyme from Arthrobacter globiformis. Apo enzyme crystals were anaerobically soaked with copper; the structure determined from this crystal provides a view of the initial state: the unmodified tyrosine coordinated to the bound copper. Exposure of the copper-bound crystals to oxygen led to the formation of freeze-trapped intermediates; structural analyses indicate that these intermediates contain dihydroxyphenylalanine quinone and trihydroxyphenylalanine. These are the first visualized intermediates during TPQ biogenesis in copper amine oxidase. PMID- 12134141 TI - Early mechanistic events in biotin dissociation from streptavidin. AB - The streptavidin-biotin system has provided a unique opportunity to investigate the molecular details of ligand dissociation pathways. An underlying mechanistic question is whether ligand dissociation proceeds with a relatively ordered process of bond breaking and ligand escape. Here we report a joint computational and crystallographic study of the earliest events in biotin dissociation. In molecular dynamics potential of mean force simulations, a water molecule from a defined access channel intercalated into the hydrogen bond between Asp 128 and biotin, bridging them and stabilizing an intermediate state. In forced biotin dissociation simulations, this event led to subsequent bond breaking steps and ligand escape. In equilibrium simulations, the water molecule was sometimes observed to move back to the access channel with re-formation of the biotin hydrogen bond. Analysis of streptavidin crystal structures revealed a close overlap of crystallographically defined and simulated waters in the water access channel. These results suggest that biotin dissociation is initiated by stochastic coupling of water entry with lengthening of a specific biotin hydrogen bonding interaction. PMID- 12134142 TI - Chronic suppression of heart-failure progression by a pseudophosphorylated mutant of phospholamban via in vivo cardiac rAAV gene delivery. AB - The feasibility of gene therapy for cardiomyopathy, heart failure and other chronic cardiac muscle diseases is so far unproven. Here, we developed an in vivo recombinant adeno-associated virus (rAAV) transcoronary delivery system that allows stable, high efficiency and relatively cardiac-selective gene expression. We used rAAV to express a pseudophosphorylated mutant of human phospholamban (PLN), a key regulator of cardiac sarcoplasmic reticulum (SR) Ca(2+) cycling in BIO14.6 cardiomyopathic hamsters. The rAAV/S16EPLN treatment enhanced myocardial SR Ca(2+) uptake and suppressed progressive impairment of left ventricular (LV) systolic function and contractility for 28-30 weeks, thereby protecting cardiac myocytes from cytopathic plasma-membrane disruption. Low LV systolic pressure and deterioration in LV relaxation were also largely prevented by rAAV/S16EPLN treatment. Thus, transcoronary gene transfer of S16EPLN via rAAV vector is a potential therapy for progressive dilated cardiomyopathy and associated heart failure. PMID- 12134143 TI - Heart-valve mesenchyme formation is dependent on hyaluronan-augmented activation of ErbB2-ErbB3 receptors. AB - Heart septation and valve malformations constitute the most common anatomical birth defects. These structures arise from the endocardial cushions within the atrioventricular canal (AVC) through dynamic interactions between cushion cells and the extracellular matrix (termed cardiac jelly). Transformation of endothelial cells to mesenchymal cells is essential for the proper development of the AVC and subsequent septation and valve formation. Atrioventricular septal defects can result from incomplete endocardial cushion morphogenesis. We show that hyaluronan-deficient AVC explants from Has2(-/-) embryos, which normally lack mesenchyme formation, are rescued by heregulin treatment, which restores phosphorylation of ErbB2 and ErbB3. These events were blocked using a soluble ErbB3 molecule, as well as with an inhibitor of ErbB2, herstatin. We show further that ErbB3 is activated during hyaluronan treatment of Has2(-/-) explants. These data provide a link between extracellular matrix-hyaluronan and ErbB receptor activation during development of early heart-valve and septal mesenchyme. PMID- 12134144 TI - Visualization of advanced human prostate cancer lesions in living mice by a targeted gene transfer vector and optical imaging. AB - Non-invasive imaging and transcriptional targeting can improve the safety of therapeutic approaches in cancer. Here we demonstrate the ability to identify metastases in a human-prostate cancer model, employing a prostate-specific adenovirus vector (AdPSE-BC-luc) and a charge-coupled device-imaging system. AdPSE-BC-luc, which expresses firefly luciferase from an enhanced prostate specific antigen promoter, restricted expression in the liver but produced robust signals in prostate tumors. In fact, expression was higher in advanced, androgen independent tumors than in androgen-dependent lesions. Repetitive imaging over a three-week period after AdPSE-BC-luc injection into tumor-bearing mice revealed that the virus could locate and illuminate metastases in the lung and spine. Systemic injection of low doses of AdPSE-BC-luc illuminated lung metastasis. These results demonstrate the potential use of a non-invasive imaging modality in therapeutic and diagnostic strategies to manage prostate cancer. PMID- 12134145 TI - Negative regulation of CD45 by differential homodimerization of the alternatively spliced isoforms. AB - The regulation of receptor-like protein tyrosine phosphatases (RPTPs) is not well understood. Although CD45 can be negatively regulated by dimerization, how dimerization is modulated is unclear. Here we show that various isoforms of CD45 differentially homodimerize in T cells. The dimerization is modulated by the sialylation and O-glycosylation of alternatively spliced CD45 exons in the extracellular domain. Thus, the smallest isoform, CD45RO--which undergoes the least extracellular sialylation and O-glycosylation--homodimerizes with the highest efficiency, resulting in decreased signaling via the T cell receptor. Because CD45 is required for T cell activation, our findings may reveal a mechanism that contributes to the termination of the primary T cell response. Our results not only demonstrate the biological significance of alternative splicing in the immune system, but also suggest a model for regulating RPTP dimerization and function. PMID- 12134146 TI - Systematic screen for human disease genes in yeast. AB - High similarity between yeast and human mitochondria allows functional genomic study of Saccharomyces cerevisiae to be used to identify human genes involved in disease. So far, 102 heritable disorders have been attributed to defects in a quarter of the known nuclear-encoded mitochondrial proteins in humans. Many mitochondrial diseases remain unexplained, however, in part because only 40-60% of the presumed 700-1,000 proteins involved in mitochondrial function and biogenesis have been identified. Here we apply a systematic functional screen using the pre-existing whole-genome pool of yeast deletion mutants to identify mitochondrial proteins. Three million measurements of strain fitness identified 466 genes whose deletions impaired mitochondrial respiration, of which 265 were new. Our approach gave higher selection than other systematic approaches, including fivefold greater selection than gene expression analysis. To apply these advantages to human disorders involving mitochondria, human orthologs were identified and linked to heritable diseases using genomic map positions. PMID- 12134147 TI - Epistatic interaction between KIR3DS1 and HLA-B delays the progression to AIDS. AB - Natural killer (NK) cells provide defense in the early stages of the innate immune response against viral infections by producing cytokines and causing cytotoxicity. The killer immunoglobulin-like receptors (KIRs) on NK cells regulate the inhibition and activation of NK-cell responses through recognition of human leukocyte antigen (HLA) class I molecules on target cells KIR and HLA loci are both highly polymorphic, and some HLA class I products bind and trigger cell-surface receptors specified by KIR genes. Here we report that the activating KIR allele KIR3DS1, in combination with HLA-B alleles that encode molecules with isoleucine at position 80 (HLA-B Bw4-80Ile), is associated with delayed progression to AIDS in individuals infected with human immunodeficiency virus type 1 (HIV-1). In the absence of KIR3DS1, the HLA-B Bw4-80Ile allele was not associated with any of the AIDS outcomes measured. By contrast, in the absence of HLA-B Bw4-80Ile alleles, KIR3DS1 was significantly associated with more rapid progression to AIDS. These observations are strongly suggestive of a model involving an epistatic interaction between the two loci. The strongest synergistic effect of these loci was on progression to depletion of CD4(+) T cells, which suggests that a protective response of NK cells involving KIR3DS1 and its HLA class I ligands begins soon after HIV-1 infection. PMID- 12134148 TI - SPG20 is mutated in Troyer syndrome, an hereditary spastic paraplegia. AB - Troyer syndrome (TRS) is an autosomal recessive complicated hereditary spastic paraplegia (HSP) that occurs with high frequency in the Old Order Amish. We report mapping of the TRS locus to chromosome 13q12.3 and identify a frameshift mutation in SPG20, encoding spartin. Comparative sequence analysis indicates that spartin shares similarity with molecules involved in endosomal trafficking and with spastin, a molecule implicated in microtubule interaction that is commonly mutated in HSP. PMID- 12134149 TI - PP1 binds Sara and negatively regulates Dpp signaling in Drosophila melanogaster. AB - In signaling involving the transforming growth factor-beta (TGF-beta) superfamily of proteins, ligand binding brings the constitutively active type II receptor kinase into close proximity to its substrate, the type I receptor kinase, which it then activates by phosphorylation. The type I receptor kinase in turn phosphorylates one of the Smad family of transcription factors, which translocates to the nucleus and regulates gene expression. Smads are recruited to the receptor complex by an anchor protein, SARA (Smad anchor for receptor activation). Although several protein kinases in this pathway were known, including the receptors themselves, the relevant phosphatases had not previously been identified. Here we report the isolation of a Drosophila melanogaster homolog of SARA (Sara) in a screen for proteins that bind the catalytic subunit of type 1 serine/threonine protein phosphatase (PP1c). We identified a PP1c binding motif in Sara, disruption of which reduced the ability of Sara to bind PP1c. Expression of this non-PP1c-binding mutant resulted in hyperphosphorylation of the type I receptor and stimulated expression of a target of TGF-beta signaling. Reducing PP1c activity enhanced the increase in the basal level of expression of genes responsive to Dpp (Decapentaplegic) caused by ectopic expression of the type II receptor Punt. Together these data suggest that PP1c is targeted to Dpp receptor complexes by Sara, where it acts as a negative regulator of Dpp signaling by affecting the phosphorylation state of the type I receptor. PMID- 12134150 TI - Selection for short introns in highly expressed genes. AB - Transcription is a slow and expensive process: in eukaryotes, approximately 20 nucleotides can be transcribed per second at the expense of at least two ATP molecules per nucleotide. Thus, at least for highly expressed genes, transcription of long introns, which are particularly common in mammals, is costly. Using data on the expression of genes that encode proteins in Caenorhabditis elegans and Homo sapiens, we show that introns in highly expressed genes are substantially shorter than those in genes that are expressed at low levels. This difference is greater in humans, such that introns are, on average, 14 times shorter in highly expressed genes than in genes with low expression, whereas in C. elegans the difference in intron length is only twofold. In contrast, the density of introns in a gene does not strongly depend on the level of gene expression. Thus, natural selection appears to favor short introns in highly expressed genes to minimize the cost of transcription and other molecular processes, such as splicing. PMID- 12134151 TI - Revealing modular organization in the yeast transcriptional network. AB - Standard clustering methods can classify genes successfully when applied to relatively small data sets, but have limited use in the analysis of large-scale expression data, mainly owing to their assignment of a gene to a single cluster. Here we propose an alternative method for the global analysis of genome-wide expression data. Our approach assigns genes to context-dependent and potentially overlapping 'transcription modules', thus overcoming the main limitations of traditional clustering methods. We use our method to elucidate regulatory properties of cellular pathways and to characterize cis-regulatory elements. By applying our algorithm systematically to all of the available expression data on Saccharomyces cerevisiae, we identify a comprehensive set of overlapping transcriptional modules. Our results provide functional predictions for numerous genes, identify relations between modules and present a global view on the transcriptional network. PMID- 12134152 TI - Temporal structure in neuronal activity during working memory in macaque parietal cortex. AB - Many cortical structures have elevated firing rates during working memory, but it is not known how the activity is maintained. To investigate whether reverberating activity is important, we studied the temporal structure of local field potential (LFP) activity and spiking from area LIP in two awake macaques during a memory saccade task. Using spectral analysis, we found spatially tuned elevated power in the gamma band (25-90 Hz) in LFP and spiking activity during the memory period. Spiking and LFP activity were also coherent in the gamma band but not at lower frequencies. Finally, we decoded LFP activity on a single-trial basis and found that LFP activity in parietal cortex discriminated between preferred and anti preferred direction with approximately the same accuracy as the spike rate and predicted the time of a planned movement with better accuracy than the spike rate. This finding could accelerate the development of a cortical neural prosthesis. PMID- 12134153 TI - Model for a robust neural integrator. AB - Integrator circuits in the brain show persistent firing that reflects the sum of previous excitatory and inhibitory inputs from external sources. Integrator circuits have been implicated in parametric working memory, decision making and motor control. Previous work has shown that stable integrator function can be achieved by an excitatory recurrent neural circuit, provided synaptic strengths are tuned with extreme precision (better than 1% accuracy). Here we show that integrator circuits can function without fine tuning if the neuronal units have bistable properties. Two specific mechanisms of bistability are analyzed, one based on local recurrent excitation, and the other on the voltage-dependence of the NMDA (N-methyl-D-aspartate) channel. Neither circuit requires fine tuning to perform robust integration, and the latter actually exploits the variability of neuronal conductances. PMID- 12134154 TI - Color from invisible patterns. AB - Human pattern resolution is limited by optical blurring as well as neural filtering by a cascade of retinal and cortical sites with progressively lower resolution limits. Curiously, pattern structure can influence perceived color: a high-contrast, monochromatic (single wavelength) pattern appears desaturated (closer to white) relative to a uniform field of the same wavelength. Here we show that this desaturation is evident even when the pattern's frequency is too high for conscious perception, implicating a nonlinear process--namely light adaptation--at the level of single cone photoreceptors. We propose a neural mechanism in which fast, involuntary eye movements serve to shift control over perception between two competing cone populations, each operating at different levels of adaptation. PMID- 12134155 TI - Glutamate release during LTD at cerebellar climbing fiber-Purkinje cell synapses. AB - It is widely thought that persistent, use-dependent alterations in synaptic strength such as long-term synaptic potentiation (LTP) and depression (LTD) underlie at least a portion of memory traces in the brain, but the exact cellular locus of expression for these alterations remains to be determined. They could be expressed presynaptically as a decrease in transmitter release, postsynaptically as a decrease in the synaptic current evoked by a fixed delivery of transmitter, as an increase in the number of functional synaptic contacts, or by a combination of these mechanisms. Here we report that LTD at the climbing fiber-Purkinje cell synapse in rat cerebellum was not associated with changes in a synaptic cleft glutamate transient, indicating that this type of LTD is most likely expressed postsynaptically. PMID- 12134156 TI - Molecular interpretation of ERK signal duration by immediate early gene products. AB - The duration of intracellular signalling is associated with distinct biological responses, but how cells interpret differences in signal duration are unknown. We show that the immediate early gene product c-Fos functions as a sensor for ERK1 (extracellular-signal-regulated kinase 1) and ERK2 signal duration. When ERK activation is transient, its activity declines before the c-Fos protein accumulates, and under these conditions c-Fos is unstable. However, when ERK signalling is sustained, c-Fos is phosphorylated by still-active ERK and RSK (90K ribosomal S6 kinase). Carboxy-terminal phosphorylation stabilizes c-Fos and primes additional phosphorylation by exposing a docking site for ERK, termed the FXFP (DEF) domain. Mutating the DEF domain disrupts the c-Fos sensor and c-Fos mediated signalling. Other immediate early gene products that control cell cycle progression, neuronal differentiation and circadium rhythms also contain putative DEF domains, indicating that multiple sensors exist for sustained ERK signalling. Together, our data identify a general mechanism by which cells can interpret differences in ERK activation kinetics. PMID- 12134157 TI - EphrinA1-induced cytoskeletal re-organization requires FAK and p130(cas). AB - Ephrins and Eph receptors are involved in axon guidance and cellular morphogenesis. An interaction between ephrin and Eph receptors elicits neuronal growth-cone collapse through cytoskeletal disassembly. When NIH3T3 cells were plated onto an ephrinA1-coated surface, the cells both adhered and spread. Adhesion and spreading proceeded concomitantly with changes in both the actin and microtubule cytoskeleton. EphA2, focal adhesion kinase (FAK) and p130(cas) were identified as the major ephrin-dependent phosphotyrosyl proteins during the ephrin-induced morphological changes. Mouse embryonic fibroblasts (MEFs) derived from FAK(-/-) and p130(cas-/-) mice had severe defects in ephrinA1-induced cell spreading, which were reversed after re-expression of FAK or p130(cas), respectively. Expression of a constitutively active EphA2 induced NIH3T3 cells to undergo identical, but ligand-independent, morphological changes. These data show that ephrinA1 can induce cell adhesion and actin cytoskeletal changes in fibroblasts in a FAK- and p130(cas)-dependent manner, through activation of the EphA2 receptor. The finding that ephrin Eph signalling can result in actin cytoskeletal assembly, rather than disassembly, has many implications for ephrin Eph responses in other cell types. PMID- 12134158 TI - Unconventional Rac-GEF activity is mediated through the Dock180-ELMO complex. AB - Mammalian Dock180 and ELMO proteins, and their homologues in Caenorhabditis elegans and Drosophila melanogaster, function as critical upstream regulators of Rac during development and cell migration. The mechanism by which Dock180 or ELMO mediates Rac activation is not understood. Here, we identify a domain within Dock180 (denoted Docker) that specifically recognizes nucleotide-free Rac and can mediate GTP loading of Rac in vitro. The Docker domain is conserved among known Dock180 family members in metazoans and in a yeast protein. In cells, binding of Dock180 to Rac alone is insufficient for GTP loading, and a Dock180 ELMO1 interaction is required. We can also detect a trimeric ELMO1 Dock180 Rac1 complex and ELMO augments the interaction between Dock180 and Rac. We propose that the Dock180 ELMO complex functions as an unconventional two-part exchange factor for Rac. PMID- 12134159 TI - CRMP-2 binds to tubulin heterodimers to promote microtubule assembly. AB - Regulated increase in the formation of microtubule arrays is thought to be important for axonal growth. Collapsin response mediator protein-2 (CRMP-2) is a mammalian homologue of UNC-33, mutations in which result in abnormal axon termination. We recently demonstrated that CRMP-2 is critical for axonal differentiation. Here, we identify two activities of CRMP-2: tubulin-heterodimer binding and the promotion of microtubule assembly. CRMP-2 bound tubulin dimers with higher affinity than it bound microtubules. Association of CRMP-2 with microtubules was enhanced by tubulin polymerization in the presence of CRMP-2. The binding property of CRMP-2 with tubulin was apparently distinct from that of Tau, which preferentially bound microtubules. In neurons, overexpression of CRMP 2 promoted axonal growth and branching. A mutant of CRMP-2, lacking the region responsible for microtubule assembly, inhibited axonal growth and branching in a dominant-negative manner. Taken together, our results suggest that CRMP-2 regulates axonal growth and branching as a partner of the tubulin heterodimer, in a different fashion from traditional MAPs. PMID- 12134160 TI - Connective-tissue growth factor (CTGF) modulates cell signalling by BMP and TGF beta. AB - Connective-tissue growth factor (CTGF) is a secreted protein implicated in multiple cellular events including angiogenesis, skeletogenesis and wound healing. It is a member of the CCN family of secreted proteins, named after CTGF, cysteine-rich 61 (CYR61), and nephroblastoma overexpressed (NOV) proteins. The molecular mechanism by which CTGF or other CCN proteins regulate cell signalling is not known. CTGF contains a cysteine-rich domain (CR) similar to those found in chordin and other secreted proteins, which in some cases have been reported to function as bone morphogenetic protein (BMP) and TGF-beta binding domains. Here we show that CTGF directly binds BMP4 and TGF-beta 1 through its CR domain. CTGF can antagonize BMP4 activity by preventing its binding to BMP receptors and has the opposite effect, enhancement of receptor binding, on TGF-beta 1. These results show that CTGF inhibits BMP and activates TGF-beta signals by direct binding in the extracellular space. PMID- 12134161 TI - Src-induced de-regulation of E-cadherin in colon cancer cells requires integrin signalling. AB - Although Src expression and activity are often elevated in colon cancer, the precise consequences of overexpression of the non-catalytic Src homology (SH) domains, or enhanced catalytic activity, are unknown. We show that, in KM12C colon cancer cells, elevated Src activity causes the components of adherens junctions, including vinculin, to be redistributed to Src-induced integrin adhesion complexes. Specifically, elevated Src activity blocks proper assembly of cell cell contacts after cells are switched from media containing a low level of calcium to media containing a high level of calcium, and E-cadherin remains internalized. In contrast, although elevated expression of the non-catalytic domains of Src is sufficient to induce assembly of integrin adhesion complexes, it does not induce disorganization of E-cadherin-associated intercellular contacts. Surprisingly, Src-induced disruption of E-cadherin localization requires specific integrin signalling, because E-cadherin redistribution is blocked by loss of cell-matrix interaction, or by inhibitory antibodies to alpha(v) or beta(1) integrin subunits. Furthermore, phosphorylation of the integrin-regulated focal adhesion kinase (FAK) on Src-specific sites is required for Src-induced de-regulation of E-cadherin, demonstrating interdependence between integrin-induced signals and cadherin-associated adhesion changes induced by Src. PMID- 12134162 TI - Myosin VI is required for E-cadherin-mediated border cell migration. AB - Myosin VI (MyoVI) is a pointed-end-directed, actin-based motor protein, and mutations in the gene result in disorganization of hair cell stereocilia and cause deafness in mice. MyoVI also localizes to the leading edges of growth factor-stimulated fibroblast cells and has been suggested to be involved in cell motility. There has been no direct test of this hypothesis, however. Drosophila melanogaster MyoVI is expressed in a small group of migratory follicle cells, known as border cells. Here we show that depletion of MyoVI specifically from border cells severely inhibited their migration. Similar to MyoVI, E-cadherin is required for border cell migration. We found that E-cadherin and Armadillo (Arm, Drosophila beta-catenin) protein levels were specifically reduced in cells lacking MyoVI, whereas other proteins were not. In addition, MyoVI protein levels were reduced in cells lacking DE-cadherin or Arm. MyoVI and Arm co immunoprecipitated from ovarian protein extracts. These data suggest that MyoVI is required for border cell migration where it stabilizes E-cadherin and Arm. Mutations in MyoVIIA, another unconventional myosin protein, also lead to deafness, and MyoVIIA interacts with E-cadherin through a membrane protein called vezatin. Multiple biochemical mechanisms may exist, therefore, for cadherins to associate with diverse unconventional myosins that are required for normal stereocilium formation or maintenance. PMID- 12134163 TI - Kinesin I-dependent cortical exclusion restricts pole plasm to the oocyte posterior. AB - Microtubules and the plus-end-directed microtubule motor Kinesin I are required for the selective accumulation of oskar mRNA at the posterior cortex of the Drosophila melanogaster oocyte, which is essential to posterior patterning and pole plasm assembly. We present evidence that microtubule minus ends associate with the entire cortex, and that Kinesin and microtubules are not required for oskar mRNA association with the posterior pole, but prevent ectopic localization of this transcript and the pole plasm proteins Oskar and Vasa to other cortical regions. Cortical binding of oskar mRNA seems to be dependent on the actin cytoskeleton. We conclude that most of the actin-rich oocyte cortex can support pole plasm assembly, and propose that Kinesin restricts pole plasm formation to the posterior by moving oskar mRNA away from microtubule-rich lateral and anterior cortical regions. PMID- 12134164 TI - Tiam1 mediates Ras activation of Rac by a PI(3)K-independent mechanism. AB - Rac is a member of the Ras superfamily of GTPases and functions as a GDP/GTP regulated switch. Formation of active Rac-GTP is stimulated by Dbl family guanine nucleotide exchange factors (GEFs), such as Tiam1 (ref. 2). Once activated, Rac stimulates signalling pathways that regulate actin organization, gene expression and cellular proliferation. Rac also functions downstream of the Ras oncoprotein in pathways that stimulate membrane ruffling, growth transformation, activation of the c-Jun amino-terminal kinase (JNK) mitogen-activated protein kinase, activation of the NF-kappa B transcription factor and promotion of cell survival. Although recent studies support phosphatidylinositol 3-OH kinase (PI(3)K) dependent mechanisms through which Ras might activate Rac (refs 9,10), the precise mechanism remains to be determined. Here we demonstrate that Tiam1, a Rac specific GEF, preferentially associates with activated GTP-bound Ras through a Ras-binding domain. Furthermore, activated Ras and Tiam1 cooperate to cause synergistic formation of Rac-GTP in a PI(3)K-independent manner. Thus, Tiam1 can function as an effector that directly mediates Ras activation of Rac. PMID- 12134165 TI - An actin nucleation mechanism mediated by Bni1 and profilin. AB - Formins are required for cell polarization and cytokinesis, but do not have a defined biochemical activity. In Saccharomyces cerevisiae, formins and the actin monomer-binding protein profilin are specifically required to assemble linear actin structures called 'actin cables'. These structures seem to be assembled independently of the Arp2/3 complex, the only well characterized cellular mediator of actin nucleation. Here, an activated yeast formin was purified and found to promote the nucleation of actin filaments in vitro. Formin-dependent actin nucleation was stimulated by profilin. Thus, formin and profilin mediate actin nucleation by an Arp2/3-independent mechanism. These findings suggest that distinct actin nucleation mechanisms may underlie the assembly of different actin cytoskeletal structures. PMID- 12134166 TI - Magnetic relaxation switches capable of sensing molecular interactions. AB - Highly sensitive, efficient, and high-throughput biosensors are required for genomic and proteomic data acquisition in complex biological samples and potentially for in vivo applications. To facilitate these studies, we have developed biocompatible magnetic nanosensors that act as magnetic relaxation switches (MRS) to detect molecular interactions in the reversible self-assembly of disperse magnetic particles into stable nanoassemblies. Using four different types of molecular interactions (DNA-DNA, protein-protein, protein-small molecule, and enzyme reactions) as model systems, we show that the MRS technology can be used to detect these interactions with high efficiency and sensitivity using magnetic relaxation measurements including magnetic resonance imaging (MRI). Furthermore, the magnetic changes are detectable in turbid media and in whole-cell lysates without protein purification. The developed magnetic nanosensors can be used in a variety of biological applications such as in homogeneous assays, as reagents in miniaturized microfluidic systems, as affinity ligands for rapid and high-throughput magnetic readouts of arrays, as probes for magnetic force microscopy, and potentially for in vivo imaging. PMID- 12134167 TI - Cloned zebrafish by nuclear transfer from long-term-cultured cells. AB - Although mammals have been cloned from genetically manipulated cultured cells, a comparable achievement has not been realized in lower vertebrates. Here we report that fertile transgenic zebrafish can be obtained by nuclear transfer using embryonic fibroblast cells from long-term cultures. The donor nuclei, modified by retroviral insertions expressing green fluorescent protein (GFP), were transplanted into manually enucleated eggs. Overall, a 2% success rate was achieved, resulting in 11 adult transgenic zebrafish expressing GFP. These nuclear transplants produced fertile, diploid offspring, and their F1/F2 progeny continued to express GFP in a pattern identical to that of the founder fish. This finding demonstrates that slowly dividing nuclei from cultured cells can be reprogrammed to support rapid embryonic development and sets up a foundation for targeted genetic manipulation in zebrafish. PMID- 12134168 TI - Hybrid insulin cocrystals for controlled release delivery. AB - The ability to tailor the release profile of a drug by manipulating its formulation matrix offers important therapeutic advantages. We show here that human insulin can be cocrystallized at preselected ratios with the fully active lipophilically modified insulin derivative octanoyl-N(epsilon)-LysB29-human insulin (C8-HI). The cocrystal is analogous to the NPH (neutral protamine Hagedorn) crystalline complex formed with human insulin, which is commonly used as the long-acting insulin component of diabetes therapy. The in vitro and in vivo release rates of the cocrystal can be controlled by adjusting the relative proportions of the two insulin components. We identified a cocrystal composition comprising 75% C8-HI and 25% human insulin that exhibits near-ideal basal pharmacodynamics in somatostatin-treated beagle dogs. The dependence of release rate on cocrystal ratio provides a robust mechanism for modulating insulin pharmacodynamics. These findings show that a crystalline protein matrix may accommodate a chemical modification that alters the dissolution rate of the crystal in a therapeutically useful way, yet that is structurally innocuous enough to preserve the pharmaceutical integrity of the original microcrystalline entity and the pharmacological activity of the parent molecule. PMID- 12134170 TI - Treatment of cerebral sinus/venous thrombosis. PMID- 12134171 TI - Clinical and molecular diagnosis of spinal muscular atrophy. AB - The spinal muscular atrophies are a group of disorders characterized by flaccid limb weakness. It is necessary to differentiate these from other causes and identify the SMA variants. In classical SMA, majority of the patients shows homozygous deletion of the telomeric SMN gene (SMN1) on chromosome 5q. The availability of DNA analysis has allowed proper genetic counseling and prenatal diagnosis in the affected families. Application of newer techniques has enabled more accurate carrier detection. Our objective is to stress the variability in the clinical features and recent advances in the molecular diagnosis for SMA. PMID- 12134169 TI - Individually addressable parallel peptide synthesis on microchips. AB - Miniaturized, spatially addressable microchips of peptides and peptidomimetics are powerful tools for high-throughput biomedical and pharmaceutical research and the advancement of proteomics. Here we report an efficient and flexible method for the parallel synthesis of peptides on individually addressable microchips, using digital photolithography and photogenerated acid in the deprotection step. We demonstrate that we are able to synthesize thousands of peptides in a 1 cm(2) area on a microchip using 20 natural amino acids as well as synthetic amino acid analogs, with high stepwise yields and short reaction-cycle times. Epitope screening experiments using a p53 antibody (PAb240) produced clearly defined binding patterns. The peptidomimetic sequences on the microchip show specific antibody binding and provide insights into the molecular details responsible for specificity of epitope binding. Our approach requires just a conventional synthesizer and a computer-controllable optical module, thereby allowing potential development of peptide microchips for various pharmaceutical and proteomic applications in routine research laboratories. PMID- 12134172 TI - Utility of databases and information technology in pediatric neurology. AB - Significant number of neurological patients in the pediatric age group have genetic and/or metabolic basis. It is difficult to remember details of each of them as their number is very large and the disorders are encountered infrequently. This impracticality necessitates the use of various websites and data base search. The internet has become a tool by which one can obtain and disseminate information. It has enhanced the medical person's ability to know at the earliest the developments in different medical specialities. Furthermore, these rare disorders are being recognized on the basis of specialized tests available only at selected centres which deal with few of these disorders. Our objective is to provide pediatric neurologists easy access to the expanding body of medical information and also to make them aware of the advancements in information technology, which is likely to facilitate telemedicine as a future consultancy service. Information about these diseases can also be facilitated by e-consultations. PMID- 12134173 TI - The neurology of eclampsia : some observations. AB - Nineteen patients admitted with diagnosis of eclampsia in a large general hospital between 1996 - 1999, were analyzed. Eight patients were referred to neurologists for assessment and management. All these patients had recurrent generalized seizures. Five patients developed visual disturbance. Neuroimaging (CT and/or MRI) revealed symmetrical occipital lesions in all. One patient had a large pontine lesion. Seizure control was achieved in all with intravenous phenytoin. All patients recovered fully without any residual neurological deficit and their radiological brain lesions resolved completely, in all except one case. The neurological manifestations and neuroimaging features in cases of eclampsia have been reviewed. A brief note on the pathogenesis of the cerebral lesions is included and the controversial aspect of seizure control in eclampsia highlighted. PMID- 12134174 TI - Transsphenoidal line of vision on MRI for pituitary tumor surgery. AB - This study was done to determine the usefulness of the sagittal magnetic resonance image (MRI) in predicting the ease of resectability of pituitary adenomas operated through the transsphenoidal route. Tumors were graded according to Hardy's method and a new system proposed by us. In this system the amount of tumor above the line of vision (V-line) as seen on the sagittal MRI was estimated and correlated with the extent of excision assessed on the postoperative computed tomogram (CT). There were 7 Hardy's grade A (8.8%), 32 grade B (41.3%), 31 grade C (37.5%), 6 grade D (8.8%) and 2 grade E tumors (3.8%) among the 78 tumors studied. It was seen that most of the tumor volume was below the V-line in Hardy's grade A and B tumors. In grade C tumors 5 were < 25% above, 14 were 25 50% above and 12 were 50-75% above the V-line. A radical excision was possible in 15 of l9 grade C tumors in which < 50% of the tumor was above the V-line. However, only 5 of 12 with Hardy's grade C were radically excised when >50% of the tumor was above the V-line. In conclusion, Hardy's grade C tumors are not a homogenous group radiologically and using the V-line on MRI helps in predicting the case of respectability in a single stage. PMID- 12134175 TI - Acute symptomatic seizures due to single CT lesions: how long to treat with antiepileptic drugs? AB - The duration of anti epileptic drug therapy for single small enhancing CT lesions (SSECTL) presents a major dilemma. We studied the efficacy of short duration (6 months) antiepileptic drug therapy as compared to long duration (2 years) drug therapy. Seventy three patients presenting with seizures and showing SSECTL on cranial CT scans (plain and contrast) were randomized into group A (6 months therapy) and group B (2 years therapy). There were 47 patients in group A and 26 patients in group B. Patients were followed up for one year after withdrawal of anti epileptic drugs. CT Head (plain and contrast) was repeated after 3 months, or earlier in cases of recurrence to rule out reinfection. 53.2% in group A and 53.8% in group B showed complete resolution and were seizure free on one year follow up. Punctate residual calcification was seen in 46.8% in group A and 46.2% in group B. Eight patients (17%) in group A and three (11.5%) in group B had a recurrence. The difference in recurrence of seizure between the two groups was not statistically significant (p<0.77) in the calcified lesion subset. Since none of the patients in total resolution subset showed recurrence, the difference between calcified and total resolution subset was highly significant. The study shows that a short duration (6 months) AED therapy in patients with total resolution of lesion on follow up scan, may be adequate in comparison to those who have calcific speck as a residue. However, a longer duration of therapy in case of calcific group probably does not alter their chances of recurrence. PMID- 12134176 TI - Randomized prospective study of outcome of short term antiepileptic treatment in small single enhancing CT lesion in brain. AB - The study was conducted in 81 patients of epilepsy with small single enhancing CT (SSECT) lesion in brain to determine the clinical profile and duration of antiepileptic drugs (AEDs) treatment. The patients were randomly divided into group A (41 cases) and group B (40 cases). Group A patients were treated for 6 months and group B for 1 year with AEDS without cysticidal drugs. The most common mode of presentation was simple partial motor seizures with secondary generalization in both the groups. Repeat imaging of brain (CT/MRI) at 6 months showed disappearance of lesion in 82.94% in group A and 87% in group B, while persistence of lesion was present only in 4.87% in group A and 5% in group B. 87.81% patients in group A and 87.17% in group B were seizure free. The recurrence of seizure occurred in 12.19% cases in group A, and 12.82% in group B. 80% of these patients had calcified lesion in both the groups. This study reveals that SSECT lesion with epilpesy is a benign self-limiting disease. It also reveals that 6 months AED treatment is as effective as one year treatment. Patients having calcified lesion or persistence of lesion might require long term AED treatment. PMID- 12134177 TI - Positional moulding in premature hydrocephalics. AB - Seven premature hydrocephalics presenting with lambdoid positional moulding (LPM) were reviewed. All were treated for hydrocephalus secondary to aqueductal stenosis, Dandy Walker Syndrome and infection. Parenchymal hemorrhage, intraventricular bleed, cortical atrophy, septal agenesis, cortical anomalies and subdural hygroma were the other common associations. These children did not show expected improvement in their higher mental functions at 6 months to 5.4 years of follow-up, following the management of hydrocephalus. It was not the LPM but associated intracranial anomalies, which were most probably responsible for their poor outcome. The differentiation from posterior plagiocephaly is also highlighted. PMID- 12134178 TI - Social factors and psychopathology in epilepsy. AB - One hundred and six epilepsy patients were assessed over a period of 6 months for psychiatric morbidity, social support, stressful life events in previous year and disability. 45 patients (42.45%) had a psychiatric diagnosis. Organic depressive disorder headed the list (16.98%) followed by mild cognitive disorder (11.32%) and tobacco dependence (8.49%). There was no significant difference in the mean age, sex, mean education, age at onset of epilepsy, duration of epilepsy, psychiatric diagnosis, mean scores on social support scale, presumptive stressful life event scale and disability assessment schedule between different types of epilepsy. The difference in mean scores of presumptive stressful life events scale and disability assessment schedule between epileptics with and without psychiatric diagnosis was not statistically significant. PMID- 12134179 TI - Correlation between CT scan and automated perimetry in supratentorial tumors. AB - An attempt was made to correlate various types of visual field defects on automated perimetry with the findings of computed tomography in 44 patients of supratentorial tumors. All the patients above the age of 10 years were subjected to complete neurological examination including investigations like plain X-rays and CT scan, however, MRI and angiography were performed wherever indicated. Ocular examination particularly pertaining to neuro-ophthalmological profile was carried out with special emphasis on automated perimetry on Humphrey field analyser. The results indicated that automated perimetry was capable of reliably detecting and quantitating the visual field defects and thus established the location of the tumor in 72% patients when compared to CT scan. Hence, any patient with neuro-ophthalmic features should be subjected to automated perimetry for early diagnosis and probable location of intracranial space occupying lesion affecting visual pathways. PMID- 12134180 TI - Neurological mitochondrial cytopathies. AB - The mitochondrial cytopathies are genetically and phenotypically heterogeneous group of disorders caused by structural and functional abnormalities in mitochondria. To the best of our knowledge, there are very few studies published from India till date. Selected and confirmed fourteen cases of neurological mitochondrial cytopathies with different clinical syndromes admitted between 1997 and 2000 are being reported. There were 8 male and 6 female patients. The mean age was 24.42+/-11.18 years (range 4-40 years). Twelve patients could be categorized into well-defined syndromes, while two belonged to undefined group. In the defined syndrome categories, three patients had MELAS (mitochondrial encephalopathy, lactic acidosis and stroke like episodes), three had MERRF (myoclonic epilepsy and ragged red fibre myopathy), three cases had KSS (Kearns Sayre Syndrome) and three were diagnosed to be suffering from mitochondrial myopathy. In the uncategorized group, one case presented with paroxysmal kinesogenic dystonia and the other manifested with generalized chorea alone. Serum lactic acid level was significantly increased in all the patients (fasting 28.96+/-4.59 mg%, post exercise 41.02+/-4.93 mg%). Muscle biopsy was done in all cases. Succinic dehydrogenase staining of muscle tissue showed subsarcolemmal accumulation of mitochondria in 12 cases. Mitochondrial DNA study could be performed in one case only and it did not reveal any mutation at nucleotides 3243 and 8344. MRI brain showed multiple infarcts in MELAS, hyperintensities in putaminal areas in chorea and bilateral cerebellar atrophy in MERRF. PMID- 12134182 TI - Endogenous sodium-potassium ATPase inhibition related biochemical cascade in trisomy 21 and Huntington's disease: neural regulation of genomic function. AB - The isoprenoid pathway related cascade was assessed in trisomy 21 and Huntington's disease. Membrane Na+-K+ ATPase activity, serum magnesium and ubiquinone were decreased while HMG CoA reductase activity, serum digoxin and dolichol levels were increased in both the disorders. There were increased levels of tryptophan catabolites (nicotine, strychnine, quinolinic acid and serotonin) and decreased levels of tyrosine catabolites (dopamine, noradrenaline and morphine) in both trisomy 21 and Huntington's disease. There was an increase in dolichol levels, carbohydrate residues of glycoproteins, glycolipids, total/individual GAG fractions and lysosomal enzymes in both disorders. Reduced levels of ubiquinone, reduced glutathione and free radical scavenging enzymes as well as increased lipid peroxidation products and nitric oxide were noticed in both the disorders. The role of hypothalamic digoxin and a disordered isoprenoid pathway in the pathogenesis of trisomy 21 and Huntington's disease is discussed. PMID- 12134181 TI - Cardiovascular responses to scalp infiltration with reduced concentration of adrenaline. AB - A prospective randomized controlled study was carried out in 41 adult neurosurgical patients to find out the hemodynamic effects following scalp infiltration with 0.5% lignocaine with or without adrenaline. The patients were divided randomly into two groups. Group I patients (n=21) received 0.5% lignocaine with adrenaline (1:8,00,000) for scalp infiltration and group II patients (n=20) received 0.5% lignocaine without adrenaline. Continuous monitoring of ECG, heart rate and arterial blood pressure was carried out every minute for 20 minutes following scalp infiltration. Blood loss while raising the scalp flap was assessed by the neurosurgeon who was unaware of the study. No significant hemodynamic disturbances were observed in either group. However, Group I patients had significantly (p=0.001) less bleeding on incision. From this study, we conclude that 0.5% lignocaine with adrenaline (1:8,00,000) does not give rise to any cardiovascular disturbances during and following scalp infiltration. Rather, it minimises blood loss while raising the craniotomy flap. PMID- 12134183 TI - Surface mapping of spike potential fields: visual vs. quantitative EEG analysis. AB - Interictal EEG spike field potentials by visual and quantitative EEG analysis were studied in 17 patients with intractable localization related epilepsy. Quantitative EEG analysis was done using commercially available window based computer program (Focus) that displayed digitally acquired EEG data and performed spline mapping. Routine EEG localized 20 spikes. Fourteen spikes had excellent congruence between manual and computer generated mapping, 6 had good congruence, and 1 had fair congruence. This study clearly proves the usefulness of spline interpolation mapping technique in localizing and characterizing the epileptiform focus. PMID- 12134184 TI - Valley sign in duchenne muscular dystrophy: importance in patients with inconspicuous calves. AB - Several patients of Duchenne muscular dystrophy (DMD) do not demonstrate clinically remarkable calf hypertrophy. A new clinical sign visible behind the shoulders, which may be called 'valley sign', was tested for its utility in such cases as clinical diagnosis becomes difficult in these patients. Out of 142 DMD patients seen in the last 7 years, 12 were found to have inconspicuous calves. All the 12 patients had clinical, biochemical and/or genetic evidence of DMD. The new sign was examined by 3 independent observers in these 12 DMD patients and in 10 patients with other neuromuscular diseases. Eight DMD patients and none of the others showed positive sign. This signifies importance of this sign in the clinical diagnosis of DMD in those children in whom the calf muscle bulk is apparently normal. PMID- 12134185 TI - Dystonia as presenting manifestation of ataxia telangiectasia : a case report. AB - Ataxia telangiectasia is a genetically inherited multisystem disorder with predominant feature being telangiectasia and cerebellar ataxia. In this report, a family of three siblings suffering from ataxia telangiectasia is described. The proband presented with dystonia and dystonic myoclonus, both of which are rare presenting features of ataxia telangiectasia. PMID- 12134186 TI - Glossopharyngeal schwannoma : a case report and review of literature. AB - We report a rare case of glossopharyngeal schwannoma whose clinical presentation and the radiological work up suggested an acoustic schwannoma. The diagnosis was made at surgery, once attachment to ninth cranial nerve was seen. The clinical presentation, radiological features and surgical findings of the glossopharyngeal schwannoma are presented along with the review of literature. PMID- 12134187 TI - Schizencephaly associated with bipolar affective disorder. AB - Schizencephaly is a rare congenital anomaly of the brain, characterized by formation of abnormal unilateral or bilateral clefts in the cerebral hemispheres. It often manifests with partial seizures, mental retardation and hemiparesis. Only two cases of schizencephaly associated with psychosis have been reported in the literature. A patient of schizencephaly, who had bipolar affective disorder is described. It has been compared with the earlier two reported cases of schizencephaly associated with pyschosis. PMID- 12134189 TI - Cavernous sinus thrombosis and air embolism following surgery for acoustic neurinoma: a case report. AB - A 55 year old male patient was operated on for a massive and vascular acoustic neurinoma in a sitting position. The tumor was completely excised. Post operatively, the patient developed irritability and clinical features suggestive of contralateral cavernous sinus thrombosis. CT scan showed air within the dural walls of the cavernous sinus on the side of surgery. However, there was no radiological evidence of cavernous sinus thrombosis on the contralateral side. Cavernous sinus thrombosis as a post-surgery complication has not been reported. Air within the dural confines of the cavernous sinus has also not been observed or radiologically recorded in the literature. PMID- 12134188 TI - Infrasellar craniopharyngioma mimicking a clival chordoma: a case report. AB - An unusual case of entirely infrasellar craniopharyngioma mimicking a clival chordoma is described. Only 22 cases of craniopharyngioma with nasopharyngeal extension have been reported in the literature. Of the reported cases, most were primarily intracranial with secondary downward extension; only two were thought to originate from an infrasellar location. The present case is another example of an entirely infrasellar craniopharyngioma, with extensive clival destruction, mimicking a clival chordoma. Relevant literature on the subject is reviewed. PMID- 12134190 TI - Lateral sacral lipomyelomeningocele : a rare anomaly. AB - Lateral sacral lipomyelomeningocele is a rare spinal developmental anomaly. In the case under report, the fat attached to the neural placode was blending with the gluteal fat externally. The cord was tethered at this level. Multiple bony anomalies and diastematomyelia were associated findings. A case of lateral sacral lipomyelomeningocele with excellent imaging detail provided by the multiplanar magnetic resonance (MR) scan is reported. PMID- 12134191 TI - Midbrain venous angioma with obstructive hydrocephalus. AB - A rare case of a mid brain venous angioma with obstructive hydrocephalus is described. A dilated draining vein from the lesion in the aqueduct as the cause of the hydrocephalus is highlighted, and interesting features of the pathology of venous angiomas and associated cavernous hemangioma are described. The management of this interesting condition is discussed. PMID- 12134192 TI - Unusual MR presentation of cerebral parenchymal tuberculosis. AB - The radiological abnormalities reported in CNS tuberculosis and their pathological correlates are discussed. Focal tuberculous involvement of the CNS without formation of tuberculoma is rare. The MR features in this case were also distinctly unusual for CNS tuberculosis. Therefore, histological confirmation of all lesions thought to be a low grade glioma is mandatory. PMID- 12134193 TI - Landau-Kleffner syndrome: a case report. AB - A healthy 5 year old boy developed aphasia, attention disorder and hyperkinesia preceded by transient formed visual hallucinations and emotional outburst, immediately after a stressful event of forced separation from his father. EEG showed generalized epileptiform activity. He was diagnosed as Landau-Kleffner syndrome (LKS). CT and MRI of the brain were normal. SPECT showed left mesial temporal hypoperfusion. He improved on antiepileptics and ACTH. PMID- 12134194 TI - Chorea due to nonketotic hyperglycemia. AB - A 62 year old diabetic and hypertensive male presented with sudden onset generalized chorea. Investigations revealed uncontrolled diabetes with absent ketones and normal serum osmolality. Achievement of euglycemia with insulin therapy abolished the involuntary movements completely within a day. The direct effect of hyperglycemia causing striatal neuronal dysfunction could be the pathogenesis of the chorea in our patient. PMID- 12134195 TI - Cerebral venous thrombosis in ulcerative colitis. AB - Cerebral venous thrombosis is a rare complication of ulcerative colitis. We report a case of 29 year old male who developed superior sagittal, left lateral and sigmoid sinus thrombosis secondary to ulcerative colitis. He was successfully treated with low molecular weight heparin and steroids. PMID- 12134196 TI - Acute bilateral extradural hematomas. AB - The occurrence of bilateral extradural hematomas is an uncommon consequence of craniocerebral trauma and its incidence is variable in various studies ranging from 2-25%.1 We studied all cases of head injury brought to our institute over a period of 6 months and found the incidence of bilateral extradural hematomas to be 13.3%. PMID- 12134197 TI - Left atrial myxoma presenting as pseudobulbar palsy. AB - A case of left atrial (LA) myxoma presenting as pseudobulbar palsy, due to multiple cerebral infarcts, without any cardiac manifestations, is presented. LA myxoma is rare cause of embolization to CNS causing ischemic infarcts. Due to multiple CNS infarcts patient can present with varied clinical picture and pseudobulbar palsy is not a very common presentation. It was a real diagnostic dilemma before LA myxoma was diagnosed on echocardiography. PMID- 12134198 TI - Palatal myoclonus following head injury: letter to the editor. PMID- 12134199 TI - Extradural ependymoma with extraspinal extension: letter to editor. PMID- 12134200 TI - Thalamic and ganglionic abscesses: a report of two cases: letter to editor. PMID- 12134201 TI - Transitory alexia without agraphia following head injury: letter to editor. PMID- 12134202 TI - Disseminated zoster with polyneuritis cranialis and motor radiculopathy: letter to editor. PMID- 12134203 TI - Sphenoid wing meningioma presenting as movement disorder: letter to editor. PMID- 12134204 TI - Idiopathic intracranial hypertension as the initial manifestation of chronic myeloid leukemia: letter to editor. PMID- 12134205 TI - Treatment of trigeminal neuralgia: letter to editor. PMID- 12134206 TI - Atlanto-occipital dislocation in rickets: letter to editor. PMID- 12134207 TI - Intracranial carotid occlusion in brain death (neuroimage). PMID- 12134208 TI - Medical Image Resource Center 2002: an update on the RSNA's Medical Image Resource Center. AB - The Radiological Society of North America has launched a project called the Medical Image Resource Center (MIRC) to establish a community of Web-based libraries of imaging information, including teaching files, other educational materials, and research data. This system would enable radiologic professionals to create and publish such materials more easily and to gain more convenient access to new and existing materials. An overview of the project, a brief summary of the overall requirements and objectives, and a brief description of the progress and ongoing plans for MIRC are presented. PMID- 12134209 TI - Irreversible compression of medical images. AB - The volume of data from medical imaging is growing at exponential rates, matching or exceeding the decline in the costs of digital data storage. While methods to reversibly compress image data do exist, current methods only achieve modest reductions in storage requirements. Irreversible compression can achieve substantially higher compression ratios without perceptible image degradation. These techniques are routinely applied in teleradiology, and often in Picture Archiving and Communications Systems. The practicing radiologist needs to understand how these compression techniques work and the nature of the degradation that occurs in order to optimize their medical practice. This paper describes the technology and artifacts commonly used in irreversible compression of medical images. PMID- 12134210 TI - Evaluation of irreversible JPEG compression for a clinical ultrasound practice. AB - A prior ultrasound study indicated that images with low to moderate levels of JPEG and wavelet compression were acceptable for diagnostic purposes. The purpose of this study is to validate this prior finding using the Joint Photographic Experts Group (JPEG) baseline compression algorithm, at a compression ratio of approximately 10:1, on a sufficiently large number of grayscale and color ultrasound images to attain a statistically significant result. The practical goal of this study is to determine if it is feasible for radiologists to use irreversibly compressed images as an integral part of the day to day ultrasound practice (ie, perform primary diagnosis with, and store irreversibly compressed images in the ultrasound PACS archive). In this study, 5 Radiologists were asked to review 300 grayscale and color static ultrasound images selected from 4 major anatomic groups. Each image was compressed and decompressed using the JPEG baseline compression algorithm at a fixed quality factor resulting in an average compression ratio of approximately 9:1. The images were presented in pairs (original and compressed) in a blinded fashion on a PACS workstation in the ultrasound reading areas, and radiologists were asked to pick which image they preferred in terms of diagnostic utility and their degree of certainty (on a scale from 1 to 4). Of the 1499 total readings, 50.17% (95% confidence intervals at 47.6%, and 52.7%) indicated a preference for the original image in the pair, and 49.83% (95% confidence intervals at 47.3%, and 52.0%) indicated a preference for the compressed image. These findings led the authors to conclude that static color and gray-scale ultrasound images compressed with JPEG at approximately 9:1 are statistically indistinguishable from the originals for primary diagnostic purposes. Based on the authors laboratory experience with compression and the results of this and other prior studies, JPEG compression is now being applied to all ultrasound images in the authors' radiology practice before reading. No image quality-related issues have been encountered after 12 months of operation (approximately 48000 examinations). PMID- 12134211 TI - Changes in technologist productivity with implementation of an enterprisewide PACS. AB - The purpose of this report is to determine what effect filmless operation has on technologist productivity when compared with traditional film-based operation. Retrospective data on technologist productivity was collected from the study institution before and after implementation of PACS using workload reports and payroll records. Departmentwide technologist productivity was defined as the number of examinations per full-time equivalent (exams/FTE) and correlated with local and nationwide standards operating in traditional film-based operations. During film-based operation, technologist productivity was comparable between the study institution and nationwide standards, allowing for the unique examination volumes and modality mix. After implementation of a large-scale PACS, technologist productivity was found to increase 34% above that of national standards and 48% that of the local control site. Implementation of an enterprisewide PACS offers the potential to significantly improve departmentwide technologist productivity when compared with traditional film-based operation. PMID- 12134212 TI - Does use of a PACS increase the number of images per study? A case study in ultrasound. AB - The purpose of this study was to determine if the use of a picture archiving and communications system (PACS) in ultrasonography increased the number of images acquired per examination. The hypothesis that such an increase does occur was based on anecdotal information; this study sought to test the hypothesis. A random sample of all ultrasound examination types was drawn from the period 1998 through 1999. The ultrasound PACS in use (ACCESS; Kodak Health Information Systems, Dallas, TX) records the number of grayscale and color images saved as part of each study. Each examination in the sample was checked in the ultrasound PACS database,.and the number of grayscale and color images was recorded. The comparison film-based sample was drawn from the period 1994 through 1995. The number of examinations of each type selected was based on the overall statistics of the section; that is, the sample was designed to represent the approximate frequency with which the various examination types are done. For film-based image counts, the jackets were retrieved, and the number of grayscale and color images were counted. The number of images obtained per examination (for most examinations) in ultrasound increased with PACS use. This was more evident with some examination types (eg, pelvis). This result, however, has to be examined for possible systematic biases because ultrasound practice has changed over the time since the authors stopped using film routinely. The use of PACS in ultrasonography was not associated with an increase in the number of images per examination based solely on the use of PACS, with the exception of neonatal head studies. Increases in the number of images per study was otherwise associated with examinations for which changes in protocols resulted in the increased image counts. PMID- 12134213 TI - Patient information extraction in digitized radiography. AB - Digital imagery is gradually replacing the traditional radiograph with the development of digital radiography and film scanner. This report presents a new method to extract the patient information number (PIN) field automatically from the film-scanned image using image analysis technique. To evaluate the PIN field extraction algorithm, 2 formats of label acquired from 2 different hospitals are tested. Given the available films with no constraints on the way the labels are written and positioned, the correct extraction rates are 73% and 84%, respectively. This extracted PIN information can link with Radiology Information System (RIS) or Hospital Information System (HIS), and the image scanned from the film then can be filed into the database automatically. The efficiency this method offers can simplify greatly the image filing process and improve the user friendliness of the overall image digitization system. Moreover, compared with the bar code reader, it solves the automatic information input problem in a very economical way. The authors believe the success of this technique will benefit the development of the PACS (Picture Archiving and Communication System) and teleradiology. PMID- 12134214 TI - Speech recognition interface to a hospital information system using a self designed visual basic program: initial experience. AB - Speech recognition (SR) in the radiology department setting is viewed as a method of decreasing overhead expenses by reducing or eliminating transcription services and improving care by reducing report turnaround times incurred by transcription backlogs. The purpose of this study was to show the ability to integrate off-the shelf speech recognition software into a Hospital Information System in 3 types of military medical facilities using the Windows programming language Visual Basic 6.0 (Microsoft, Redmond, WA). Report turnaround times and costs were calculated for a medium-sized medical teaching facility, a medium-sized nonteaching facility, and a medical clinic. Results of speech recognition versus contract transcription services were assessed between July and December, 2000. In the teaching facility, 2042 reports were dictated on 2 computers equipped with the speech recognition program, saving a total of US dollars 3319 in transcription costs. Turnaround times were calculated for 4 first-year radiology residents in 4 imaging categories. Despite requiring 2 separate electronic signatures, we achieved an average reduction in turnaround time from 15.7 hours to 4.7 hours. In the nonteaching facility, 26600 reports were dictated with average turnaround time improving from 89 hours for transcription to 19 hours for speech recognition saving US dollars 45500 over the same 6 months. The medical clinic generated 5109 reports for a cost savings of US dollars 10650. Total cost to implement this speech recognition was approximately US dollars 3000 per workstation, mostly for hardware. It is possible to design and implement an affordable speech recognition system without a large-scale expensive commercial solution. PMID- 12134215 TI - Chairman's letter: planning for the (digital) future. PMID- 12134216 TI - Endovascular repair of abdominal aortic aneurysms in women after FDA approval: results, complications, and limitations. AB - Women account for approximately 25% of patients with AAAs, but unfortunately, only 8-10% of patients considered candidates for endovascular treatment during prospective trials were women. We reviewed our experience with open surgical and endovascular techniques before and after the FDA approval of the Ancure and AneuRx devices to evaluate our results and the role of endovascular treatment of AAAs in women. From January 1999 to August 2000, 269 patients underwent elective repair of their AAAs at our institution. The 20-month period was divided into the 10 months before and after the FDA approval of the endovascular devices for comparison. In the initial time period, 75 patients (62 men and 13 women) underwent repair with 40% undergoing endovascular repair. In the 10 months after FDA approval, 194 patients (160 men and 34 women) underwent repair with 87% undergoing endovascular treatment. Ninety-two percent (155) of the patients undergoing endovascular repair in this period were treated with the AneuRx device. These 155 patients were divided into two groups based on gender to compare their early treatment results. The FDA approval of endovascular grafts has profoundly affected the treatment of infrarenal AAAs in the United States. Women continue to account for a small, but complex, proportion of patients treated with available endovascular devices and the results in these patients is worse than their male counterparts. Careful patient selection and meticulous operative techniques are needed to reliably treat women with available endovascular devices. Further developments are necessary to improve the current results and increase the proportion of women being safely and effectively treated with endovascular devices. PMID- 12134217 TI - Spinal cord ischemia following endovascular repair of an infrarenal aortic aneurysm. AB - Spinal cord ischemia following repair of an infrarenal aneurysm is a rare but recognized complication following endovascular repair of an infrarenal aneurysm. Here we discuss its possible mechanism. PMID- 12134225 TI - Long-term follow-up of chronic hepatitis B virus infection in children of different ethnic origins. AB - The natural history of chronic hepatitis B in children is influenced by mode of transmission and varies with regional endemicity. Seroconversion rates were studied in 174 hepatitis B e antigen (HBeAg)-positive children who were of different ethnic origins and living in Canada. Overall, 40.2% became anti-HBeAg positive, and 8.6% were hepatitis B surface-antigen positive during a mean follow up of 4.5 years. Spontaneous seroconversion rates were lower in Asian-born, mainly vertically infected, children, versus those born either in Canada or where horizontal transmission predominates (24% vs. 44%, P=.015). Kaplan-Meier analysis showed that the cumulative persistence of HBeAg after 13 years was 25% in Asian born children, versus 6% in all others (P<.05). Treatment of 27 children accelerated seroconversion by 3 years, without influencing the proportion seroconverting over time. Thus, although Asian-born children seroconvert more slowly, a large proportion will seroconvert before adulthood. Because treatment appears to accelerate anti-HBe seroconversion, longitudinal studies are required in order to assess the long-term benefits of early treatment. PMID- 12134226 TI - CD4 T lymphocyte proliferative responses to hepatitis C virus (HCV) antigens in patients coinfected with HCV and human immunodeficiency virus who responded to anti-HCV treatment. AB - CD4 T lymphocyte proliferative responses to hepatitis C virus (HCV) antigens were evaluated before and during an anti-HCV regimen (interferon-alpha2a and ribavirin) in 36 patients coinfected with HCV and human immunodeficiency virus (HIV), to determine whether immune responses against HCV antigens are present in such patients, whether these responses are modified by anti-HCV treatment, and whether they are correlated with treatment efficacy. The CD4 responses against HCV antigens (primarily core antigens) detected at study entry in one-half of the patients did not correlate with anti-HCV treatment efficacy. Of 36 patients, 8 had patterns of persistent immune response to infection by genotypes 3 or 4 that were significantly correlated with sustained virologic response. Persistent immunologic reactivity and sustained virologic response coexisted only in patients infected with genotype 3. These findings suggest that HCV genotype may influence specific immune response, which, in turn, is implicated in virologic control. PMID- 12134227 TI - Increased thymic output after initiation of antiretroviral therapy in human immunodeficiency virus type 1-infected children in the Paediatric European Network for Treatment of AIDS (PENTA) 5 Trial. AB - To investigate the thymic contribution to immune reconstitution during antiretroviral therapy (ART), T cell receptor gene rearrangement excision circles (TRECs) were measured in peripheral blood mononuclear cells (PBMC) and CD4 cells from 33 human immunodeficiency virus (HIV) type 1-infected children monitored for 96 weeks after ART initiation. Baseline TREC levels were associated positively with baseline CD4 cell percentage and inversely with age and HIV-1 RNA load. During therapy, TREC level changes in PBMC and CD4 cells were fairly comparable. TREC level changes were inversely related to baseline CD4 cell percentage and positively associated with CD4 cell percentage increases, the main source being naive CD4 cells. TREC changes were independent of age and baseline HIV-1 RNA load; however, HIV-1 suppression was independently associated with smaller TREC changes. Thymic output appears to be the main source of CD4 cell repopulation in children receiving ART. Recovery of thymic function is independent of age and influenced by the status of peripheral CD4 cell depletion and HIV-1 suppression. PMID- 12134228 TI - Detection of antibodies to human immunodeficiency virus (HIV) that recognize conformational epitopes of glycoproteins 160 and 41 often allows for early diagnosis of HIV infection. AB - On the basis of human immunodeficiency virus (HIV) needlestick studies, the time to seroconversion for anti-HIV antibodies is 1-9 months (mean, approximately 2-3 months). However, an earlier marker of an immune response to HIV often occurs serum anti-HIV antibodies reactive with live HIV-infected cells, termed "early HIV antibodies." The specificities of these antibodies are characterized by the recognition of type-specific conformational epitopes of the HIV envelope glycoprotein (gp) 160 and gp41. By use of a third-generation native HIV(IIIB) gp160 enzyme immunoassay (EIA), detection of HIV antibodies occurred, on average, 33 days earlier than did detection by commercial EIA and 25 days earlier than did detection by the reference antigen and reverse-transcription polymerase chain reaction (RT-PCR) assays in 3 of 5 HIV seroconversion panels. A fourth panel possessed early HIV antibodies that reacted with HIV(213) but not with HIV(IIIB), allowing for detection of HIV antibodies approximately 3 weeks earlier than by RT PCR or other current tests. PMID- 12134229 TI - Differential effects of p-glycoprotein and multidrug resistance protein-1 on productive human immunodeficiency virus infection. AB - P-glycoprotein (P-gp) and multidrug-resistance protein-1 (MRP-1) are adenosine triphosphate-binding cassette proteins that may decrease intracellular concentrations of anti-human immunodeficiency virus (HIV) drugs. After HIV 1(IIIB) infection, HIV-1 protein and infectious virus production were decreased by at least 70-fold in CEM cells overexpressing P-gp but were increased by at least 50-fold in CEM cells overexpressing MRP-1, compared with control CEM cells. After transfection with the HIV-1(IIIB) genome, cells overexpressing P-gp and MRP 1 expressed similar amounts of HIV protein. Selective inhibitors of MRP-1 and P gp partially reversed the effect of these transporters in a concentration dependent manner. P-gp preferentially associated with glycolipid-enriched membrane (GEM) domains, which may be an important site for cellular binding and egress of HIV. In contrast, MRP-1 was not preferentially found in GEM domains. These results suggest that the inhibition of HIV productive infection by P-gp and augmentation by MRP-1 occur predominantly at a preintegration step but act through different mechanisms. PMID- 12134230 TI - Lethal synergism between influenza virus and Streptococcus pneumoniae: characterization of a mouse model and the role of platelet-activating factor receptor. AB - A lethal synergism exists between influenza virus and pneumococcus, which likely accounts for excess mortality from secondary bacterial pneumonia during influenza epidemics. Characterization of a mouse model of synergy revealed that influenza infection preceding pneumococcal challenge primed for pneumonia and led to 100% mortality. This effect was specific for viral infection preceding bacterial infection, because reversal of the order of administration led to protection from influenza and improved survival. The hypothesis that influenza up-regulates the platelet-activating factor receptor (PAFr) and thereby potentiates pneumococcal adherence and invasion in the lung was examined in the model. Groups of mice receiving CV-6209, a competitive antagonist of PAFr, had survival rates similar to those of control mice, and lung and blood bacterial titers increased during PAFr inhibition. These data suggest that PAFr-independent pathways are operative in the model, prompting further study of receptor interactions during pneumonia and bacteremia. The model of lethal synergism will be a useful tool for exploring this and other mechanisms underlying viral-bacterial interactions. PMID- 12134231 TI - Native and genetically inactivated pertussis toxins induce human dendritic cell maturation and synergize with lipopolysaccharide in promoting T helper type 1 responses. AB - The capacity of pertussis toxin (PT) to induce maturation and functional activities of human monocyte-derived dendritic cells (DCs) was investigated. Both native PT (nPT) and genetically detoxified PT (dPT) efficiently promoted expression on DCs of CD80, CD86, human leukocyte antigen-DR, and CD83 markers, alloreactive antigen presentation, and cytokine production, primarily interferon (IFN)-gamma. Although they did not affect interleukin (IL)-10 production by lipopolysaccharide (LPS)-stimulated DCs, both nPT and dPT strongly synergized with LPS for IL-12 production. PTs plus LPS-stimulated DCs secreted soluble factors fostering IFN-gamma but not IL-4 and IL-5 production by naive T cells. T helper type 1 (Th1) polarization was, as alloreactive antigen presentation, inhibited by anti-IL-12 monoclonal antibody. These findings support the notion that nPT, in addition to inducing specific immune response, is a potent Th1 adjuvant and that dPT fully preserves this adjuvanticity. The synergic interaction between PT and LPS in IL-12 production might be relevant for the mechanisms of vaccine-induced protection. PMID- 12134232 TI - The inhibitory effect of C-reactive protein on bacterial phosphorylcholine platelet-activating factor receptor-mediated adherence is blocked by surfactant. AB - Numerous major bacterial pathogens in the human respiratory tract, including Streptococcus pneumoniae and Haemophilus influenzae, express cell-surface phosphorylcholine (ChoP), a ligand for the receptor for platelet-activating factor (rPAF). ChoP is also bound by C-reactive protein (CRP), which, in the presence of complement, may be bactericidal. This study found that CRP can block the attachment of bacteria expressing cell-surface ChoP to host cells. Concentrations of CRP equivalent to those on the mucosal surface of the human airway blocked most adherence of both S. pneumoniae and H. influenzae to human pharyngeal epithelial cells. ChoP-mediated adherence was also reduced in the presence of an rPAF antagonist. The antiadhesive effects of the rPAF antagonist and CRP were not additive, suggesting that CRP activity is specific to the area of adherence mediated by the receptor. The binding of CRP to ChoP and the effect of CRP on adherence were inhibited by human surfactant (primarily ChoP). The antiadhesive effect of CRP may be diminished in the terminal airway, where surfactant is abundant. PMID- 12134234 TI - Reduced nephrotoxicity of conventional amphotericin B therapy after minimal nephroprotective measures: animal experiments and clinical study. AB - Amphotericin B (AmB)-treated rats develop severe polyuria, polydypsia, impairment of renal concentrating ability, and morphologic signs of tubular damage. However, renal insufficiency develops quickly only in animals in which water intake is restricted to the median volume drunk by rats of the control group. Therefore, vigorous hydration seems crucial for prevention of AmB-induced nephrotoxicity. In a clinical study, 61 patients with hematologic malignancies receiving AmB therapy were massively hydrated to ensure urine output of > or =4000 mL/day. Urine sodium, potassium, and magnesium were also measured, and all losses were supplemented (potassium as a 7.45% solution via central venous catheter). AmB treated patients developed signs of renal tubular damage (increased fractional excretion of sodium and potassium) and required large amounts of ion supplementation. The serum ion concentration and creatinine clearance remained stable. No clinically significant renal damage developed to force premature cessation of AmB treatment. PMID- 12134233 TI - Salmonella serotype Typhimurium infection of bovine Peyer's patches down regulates plasma membrane calcium-transporting ATPase expression. AB - To identify host genes differentially expressed during Salmonella enterica serotype Typhimurium infection, an RNA differential display was made with total RNA extracted from ileal loops that were infected with Salmonella Typhimurium 2.5 h after infection. Down-regulated cDNA was identified in bovine Peyer's patches after infection that was highly homologous to a human plasma membrane calcium transporting ATPase (PMCA). Differential expression of PMCA, evaluated by Northern analysis, was found to have more than a 4.6-fold decrease in expression of mRNA (size, approximately 5.1 kb). PMCA mRNA was detected by in situ hybridization exclusively within epithelial cells in the Peyer's patches. cDNA (4.4 kb) was amplified by rapid amplification of cDNA ends, cloned, and sequenced and showed a high homology to hPMCA. Bovine PMCA is down-regulated in epithelial cells of Peyer's patches after infection with Salmonella Typhimurium and, subsequently, may influence cellular calcium levels that contribute to the inflammatory processes in the pathogenesis of diarrhea. PMID- 12134235 TI - Candida albicans stimulates local expression of leukocyte adhesion molecules and cytokines in vivo. AB - The host inflammatory response to the opportunistic pathogen Candida albicans determines susceptibility to disseminated infection. This study used immunohistochemical analysis to correlate microbiologic findings and pathologic changes with local expression of cytokines and leukocyte adhesion molecules in mice with disseminated candidiasis. After being inoculated intravenously as blastospores, the organisms filamented extensively in the kidneys while remaining predominantly as blastospores or short germ tubes in the lung, liver, and spleen. Very few leukocytes accumulated around the invading organisms until 24 h after inoculation. The leukocytes at the infection site appeared to amplify the inflammatory response. They expressed interleukin-1beta, tumor necrosis factor alpha, intercellular adhesion molecule 1, and platelet-endothelial cell adhesion molecule 1. Therefore, the morphology of the organism varies with the infection site. Furthermore, leukocyte recruitment occurs relatively late in the infection, and this recruitment is likely amplified by proinflammatory mediators produced by the leukocytes themselves. PMID- 12134236 TI - A functional network of intramolecular cross-reacting epitopes delays the elicitation of neutralizing antibodies to Trypanosoma cruzi trans-sialidase. AB - Trypanosoma cruzi trans-sialidase (TS) constitutes a key molecule in both the establishment of the infection and in the development of pathologic abnormalities associated with Chagas disease. Several cross-reactive epitopes located in its catalytic region were previously identified. In the present study, a panel of enzymes altered in these epitopes were generated to analyze their in vivo significance. Although displaying similar specific activity, thermal stability, and overall antigenic structure, mutant TS proteins elicited an improved neutralizing response, compared with that in the parent, wild-type molecule. These features support an in vivo role for cross-reactive epitopes in dampening the elicitation of TS-neutralizing antibodies. Structural and immunological evidence indicating that the epitope cross-reactivity could be extended to the highly immunogenic SAPA repeats located on the TS C terminus is also reported. This complex cross-reactive epitope cargo might represent a novel strategy, providing secreted virulence factors with the ability to delay an effective elicitation of humoral response. PMID- 12134237 TI - Absolute level of Epstein-Barr virus DNA in human immunodeficiency virus type 1 infection is not predictive of AIDS-related non-Hodgkin lymphoma. AB - To study whether Epstein-Barr virus (EBV) load can be used to predict the occurrence of acquired immunodeficiency syndrome-related non-Hodgkin lymphoma (AIDS-NHL), we determined EBV load longitudinally for individuals infected with human immunodeficiency virus type 1. EBV load in peripheral blood mononuclear cells (PBMC) was high and displayed considerable fluctuations over time, indicating that absolute EBV load in PBMC is not predictive of the development of AIDS-NHL. EBV DNA was also detectable in serum at some time points but at a lower level. PMID- 12134238 TI - Evidence of thymic function in heavily antiretroviral-treated human immunodeficiency virus type 1-infected adults with long-term virologic treatment failure. AB - Thymic function was evaluated in 32 heavily antiretroviral-treated human immunodeficiency virus type 1 (HIV-1)-infected adults with long-term virologic treatment failure by measuring thymic volume, by determining the absolute number of naive T cell phenotypes, and by determining the number of cells carrying T cell receptor excision circles (TRECs). There was a significant inverse correlation between age and thymic volume (r=-0.415; P=.018), and there was a significant direct correlation between thymic volume and total naive T cell counts (r=0.529; P=.002), naive CD4(+) cell counts (r=0.437; P=.012), naive CD8(+) cell counts (r=0.467; P=.007), and TREC levels (r=0.391; P=.027). In conclusion, this study found clear evidence that the thymus of heavily antiretroviral-treated HIV-1-infected adults with long-term virologic treatment failure is actively engaged in thymopoiesis, which generates new naive T cells for the peripheral lymphocyte pool. PMID- 12134239 TI - Evidence of Bordetella pertussis infection in adults presenting with persistent cough in a french area with very high whole-cell vaccine coverage. AB - Although France has had a vaccination program for 40 years, since 1990, an increase in whooping cough cases with parent-infant transmission has been observed. This study prospectively assessed the frequency of Bordetella pertussis infection in adults who consulted general practitioners for a persistent cough without an evident diagnosis. Among 217 patients, 70 (32%) confirmed whooping cough cases were identified. One case was culture positive, 36 were polymerase chain reaction positive, and 40 had increases or decreases of > or =2-fold in anti-pertussis toxin IgG titer between serum samples collected during the acute and convalescent phases. The median duration of cough in confirmed cases was 49 days (range, 13-123 days). Of the patients, 60% reported vaccination, and 33% reported whooping cough in infancy. Pertussis should be considered for diagnosis of acute and chronic cough in adults. Future studies should evaluate the public health interest of booster doses of pertussis vaccine in adults. PMID- 12134240 TI - A multistate outbreak of Shiga toxin-producing Escherichia coli O26:H11 infections in Germany, detected by molecular subtyping surveillance. AB - In the spring of 2000, a cluster of indistinguishable Shiga toxin-producing Escherichia coli (STEC) O26:H11 was identified in Germany by molecular subtyping surveillance. An investigation was prompted to identify a common source of exposure. A case subject was defined as a person having a polymerase chain reaction-confirmed STEC O26 infection between March and April 2000, irrespective of clinical signs, and whose isolate was indistinguishable from the index strain by use of pulsed-field gel electrophoresis. Eleven case subjects were found in 5 institutions that were supplied by 4 kitchens located in 3 states. The median age was 2 years (range, 2-31 years). No bloody diarrhea was reported, and 5 persons remained asymptomatic. Comparison of invoices revealed a certain type of beef ("Seemerrolle") as possible source of infection. This is, to our knowledge, the first multistate outbreak associated with a non-O157 STEC detected by laboratory based surveillance. Molecular subtyping was pivotal, as disease occurrence was sporadic or family-related. PMID- 12134241 TI - Cultured human gastric explants: a model for studies of bacteria-host interaction during conditions of experimental Helicobacter pylori infection. AB - The unique environment of the human stomach makes it difficult to establish representative in vitro models for Helicobacter pylori that mimic the natural infection. The in vitro explant culture (IVEC) technique is based on coculture of human gastric explants with H. pylori, where bacteria-host interaction is studied on the basis of interleukin (IL)-8 secretion of the explant tissue in response to infection. In this study, it was shown that H. pylori attachment to gastric epithelial tissue was required for induction of representative inflammatory responses, assessed here by IL-8 production. Furthermore, IL-8 production by the explant tissue in response to H. pylori infection demonstrated that the explants were adequately responsive. The IVEC technique for studies of the interplay between H. pylori and the human gastric mucosa during conditions of experimental infections in vitro could add knowledge to our understanding of the complex bacteria-host cross-talk in vivo. PMID- 12134242 TI - Increased arthritis severity in mice coinfected with Borrelia burgdorferi and Babesia microti. AB - Increased severity of disease and persistence of symptoms have been recently reported in some patients with simultaneous infection of Borrelia burgdorferi and Babesia microti in the northeastern and northern midwest United States. This study used a murine model to examine whether defined disease conditions such as arthritis and carditis differed in severity in mice infected solely with B. burgdorferi and in mice coinfected with B. microti and B. burgdorferi. C3H.HeJ and BALB/c mice cohorts were coinfected or singly infected and then monitored experimentally for 15 and 30 days after inoculation. Carditis and arthritis was determined by blinded histopathologic evaluation of myocardium and tibiotarsal joints. Cytokine measurements were made on lymph node and spleen supernatants for interferon-gamma, interleukin (IL)-4, IL-10, and IL-13. No differences were observed for C3H.HeJ mice cohorts; however, coinfected BALB/c mice had a significant increase in arthritis severity at day 30. This clinical observation was correlated with a significant reduction in expression of the cytokines IL-10 and IL-13. PMID- 12134243 TI - Role of bacteriophage MAV1 as a mycoplasmal virulence factor for the development of arthritis in mice and rats. AB - The lysogenic bacteriophage MAV1 has been shown to be a virulence factor for the development of arthritis in rats infected with Mycoplasma arthritidis. In the present study, arthritis was evaluated by histopathologic examination to demonstrate that MAV1 is a virulence factor not only in the rat but also in the mouse. Specifically, the MAV1 lysogen 158L3-1 was more virulent than the nonlysogen strain 158 in DBA/2NCr, C3H/HeNCr, C3H/HeJ, and C3Smn.CB17 Prkdc(scid)/J mice, as well as in LEW rats. PMID- 12134244 TI - Expression of tissue factor, the clotting initiator, on macrophages in Plasmodium falciparum-infected placentas. AB - The expression of tissue factor (TF), the initiator of the clotting system, was investigated by immunohistochemical staining for its role in clotting mechanisms of Plasmodium falciparum-infected placenta. Most mononuclear cells in the intervillous space of infected placentas stained with an anti-TF monoclonal antibody (MAb) and were positive for antimacrophage MAb. The intervillous space of infected placentas had significant fibrin deposition. In contrast, only small amounts of leukocytes, TF-positive cells, and fibrin were seen in the intervillous space in noninfected placentas. These results indicate that macrophages accumulated in infected placentas express TF and that subsequent perivillous fibrin clot formation leads to a narrowing and plugging of the intervillous space and disturbance of the blood supply. Macrophage TF expression in placentas could be associated with retarded placental growth and low birth weight in malaria infection and should be further investigated. PMID- 12134245 TI - Can the efficacy of bacille calmette-guerin tuberculosis vaccine be affected by intestinal parasitic infections? PMID- 12134247 TI - Polymorphisms in the genes for herpesvirus entry. PMID- 12134248 TI - Long-term immunological benefit in patients with discordant responses to therapy depends on both level of viremia and duration of rebound. PMID- 12134249 TI - Combined antiviral-antimediator treatment for the common cold. AB - A randomized, controlled, double-masked clinical trial was conducted with a combination antiviral-antimediator treatment for experimental rhinovirus colds. In all, 150 healthy men and women (aged 18-51 years) were randomly assigned to 1 of 3 groups: intranasal interferon (IFN)-alpha2b (6 x 10(6) U every 12 h x 3) plus oral chlorpheniramine (12 mg extended release) and ibuprofen (400 mg) every 12 h for 4.5 days (n=59 subjects); intranasal placebo plus oral chlorpheniramine and ibuprofen (n=61 subjects); or intranasal and oral placebos (n=30 subjects). Treatment was started 24 h after intranasal viral challenge. During the 4.5 days of treatment with IFN-alpha2b, chlorpheniramine, and ibuprofen, the daily mean total symptom score was reduced by 33%-73%, compared with placebo. Treatment reduced the severity of rhinorrhea, sneezing, nasal obstruction, sore throat, cough, and headache and reduced nasal mucus production, nasal tissue use, and virus concentrations in nasal secretions. IFN-alpha2b added to the effectiveness of chlorpheniramine and ibuprofen and was well tolerated. PMID- 12134250 TI - Interferons specifically suppress the translation from the internal ribosome entry site of hepatitis C virus through a double-stranded RNA-activated protein kinase-independent pathway. AB - Interferon (IFN) therapy is used worldwide as the best available treatment for hepatitis C virus (HCV) infection; however, little is known about how IFN or other drugs work against liver diseases. The effect of 6 drugs (glycyrrhizin, ursodeoxycholic acid, ribavirin, methylprednisolone, IFN-alpha, and IFN-beta) on HCV RNA translation from the HCV internal ribosome entry site (IRES) was investigated, using a bicistronic reporter containing the HCV IRES. IFNs suppressed both cap-dependent and HCV IRES-dependent translation, with HCV IRES dependent translation being more significantly suppressed. In contrast to HCV IRES, IFN did not suppress either foot-and-mouth disease virus IRES-dependent or encephalomyocarditis virus IRES-dependent translation more than it suppressed cap dependent translation. Moreover, dominant inhibition of HCV IRES-dependent over cap-dependent translation depended neither on the double-stranded RNA-activated protein kinase activation nor on La protein function. These results indicate a novel antiviral effect of IFNs against HCV. PMID- 12134251 TI - Polymorphisms of the human leukocyte antigen DRB1 and DQB1 genes and the natural history of human papillomavirus infection. AB - This study investigated, through cohort analysis, whether HLA-DRB1 and -DQB1 variability is related to human papillomavirus (HPV) infection prevalence and persistence. HLA-DRB1 and -DQB1 genes were typed in 620 samples from the Ludwig McGill cohort. HPV positivity was tested in specimens collected every 4 months during the first year of follow-up. Persistent and long-term infections were defined as at least 2 or 3 consecutive positive results for the same HPV type, respectively. The magnitudes of associations were estimated by unconditional logistic regression analysis adjusted for potential confounders. The DRB1*0301 DQB1*0201 haplotype was associated with a 2-fold reduction in risk for transient and persistent HPV infections. DRB1*1102-DQB1*0301 showed a lower-risk effect only for persistence. DRB1*1601-DQB1*0502 and DRB1*0807-DQB1*0402 were associated with a 7-fold and a 3-fold increase, respectively, in risk for persistence. The results suggest that HLA class II polymorphisms are involved in clearance and maintenance of HPV infection. PMID- 12134252 TI - beta-Herpesvirus (human cytomegalovirus and human herpesvirus 6) reactivation in at-risk lung transplant recipients and in human immunodeficiency virus-infected patients. AB - Human herpesvirus (HHV)-6 is a beta-herpesvirus-like human cytomegalovirus (HCMV) with the potential to reactivate in immunocompromised persons. HHV-6 and HCMV were assessed in the peripheral blood leukocytes of 26 lung transplant recipients and of 37 human immunodeficiency virus (HIV)-infected patients receiving highly active antiretroviral therapy, to determine the degree of concordance between HHV 6 and HCMV reactivation in different biologic settings. In the lung transplant recipients (145 samples), HHV-6 was not detected, even though 44 (30%) of 145 samples were from 9 HCMV DNA-positive patients (13 episodes of HCMV pneumonitis). Among the HIV-infected patients (172 samples), HCMV DNA was detected in 29 (17%) of 172 samples from 10 patients (4 episodes of HCMV disease). HHV-6 DNA was detected in 2 HIV-infected patients who did not have HCMV detected at that time. These findings suggest that the pathobiologic control mechanisms for these 2 beta herpesviruses may be significantly different. PMID- 12134254 TI - Association of virus load, CD4 cell count, and treatment with clinical progression in human immunodeficiency virus-infected patients with very low CD4 cell counts. AB - This study prospectively assessed the impact of treatment modality, virus load, and CD4 cell count of <50 cells/mm(3) on human immunodeficiency virus disease progression. The incidence rate of new AIDS disease or death was 54.8 (95% confidence interval, 48.7-59.9) per 100 person-years of follow-up. Independent predictors related to progression were latest CD4 cell count (relative risk [RR], 0.84/10 mm(3) higher; P<.0001), latest hemoglobin level (RR, 0.79/g/L higher; P<.0001), Pneumocystis carinii pneumonia prophylaxis (RR, 0.49; P<.0001), latest body mass index (RR, 0.93/kg/m(2) higher; P=.002), latest virus load (RR, 1.11/log(10) higher; P=.03), and intensity of treatment (RR, 1.82, P=.004; RR 2.27, P<.0001; RR 2.46, P=.0001; RR 2.33 P<.0006; 5.10, P<.0001, respectively, for 4, 3, 2, 1, or no drugs vs. >or=5 drugs). Although reverse causality cannot be excluded, more intense antiviral treatment appears to decrease the risk of progression in immunocompromised patients. PMID- 12134253 TI - Development of resistance mutations in women receiving standard antiretroviral therapy who received intrapartum nevirapine to prevent perinatal human immunodeficiency virus type 1 transmission: a substudy of pediatric AIDS clinical trials group protocol 316. AB - Pediatric AIDS Clinical Trials Group protocol 316 was an international, multicenter, placebo-controlled trial comparing single-dose oral nevirapine (200 mg to mother and 2 mg/kg to infant) with placebo in human immunodeficiency virus (HIV)-infected pregnant women receiving standard antiretroviral therapy. This substudy evaluated the emergence of nevirapine-resistance mutations at 6 weeks postpartum in a subgroup of participants. Maternal risk factors for the emergence of nevirapine-resistance mutations were evaluated. Mutations associated with nevirapine resistance were detectable at delivery, prior to receipt of study drug, in 5 (2.3%) of 217 women. Fourteen (15%; 95% confidence interval, 8%-23%) of 95 women who received intrapartum nevirapine developed a nevirapine-resistance mutation 6 weeks postpartum. The most common mutation was K103N, which was present in 10 women. The risk for development of a new nevirapine-resistance mutation did not correlate with CD4 cell count or HIV-1 RNA load at delivery or with type of antepartum antiretroviral therapy. The risk of nevirapine resistance should be considered when determining the risks or benefits of intrapartum nevirapine in women receiving antepartum antiretroviral therapy. PMID- 12134255 TI - The pharmacokinetics of amprenavir, zidovudine, and lamivudine in the genital tracts of men infected with human immunodeficiency virus type 1 (AIDS clinical trials group study 850). AB - The AIDS Clinical Trials Group Study 850 (ACTG 850) evaluated the penetration of zidovudine (ZDV), lamivudine (3TC), and amprenavir (APV), given alone and in combination with the 2 nucleoside analogues, into the male genital tract, because these factors may affect human immunodeficiency virus (HIV) type 1 suppression and transmission. Nineteen men receiving APV monotherapy and 12 men receiving triple therapy donated blood plasma (BP) and seminal plasma (SP) during therapy. Paired SP and BP were used to calculate compartmental concentration ratios. APV SP concentrations were consistently lower than BP concentrations, ZDV SP concentrations approximated BP concentrations early but became greater later in the dosing interval, and 3TC SP concentrations were substantially greater than BP concentrations throughout. Observed SP concentrations plotted with population BP concentration-time curves confirmed these findings, suggesting that passive diffusion (APV), slowed elimination (ZDV), and either active accumulation and/or inhibition of elimination (3TC) are responsible for SP concentrations of these agents. The antiretroviral effect of APV monotherapy was related to APV concentrations. PMID- 12134256 TI - High levels of antibodies to the CD4 binding domain of human immunodeficiency virus type 1 glycoprotein 120 are associated with faster disease progression. AB - Human monoclonal antibodies (Abs) to the CD4 binding domain of human immunodeficiency virus (HIV) type 1 glycoprotein (gp) 120 (gp120(CD4bd)) inhibit gp120 presentation to gp120-specific T helper (Th) cells. Since Th responses are critical to control HIV, anti-gp120(CD4bd) Abs could be involved in HIV pathogenesis. Therefore, anti-gp120(CD4bd) Ab levels were compared in serum samples from matched pairs of HIV-positive rapid progressors (RPs) and slow progressors (SPs). Many RPs had higher levels of anti-gp120(CD4bd) Abs than their corresponding SPs. However, Ab levels to whole gp120 and to its C5 domain were similar. Hence, the higher levels of anti-gp120(CD4bd) Abs detected in the serum of RPs do not reflect generalized increases in Ab levels to whole gp120. Moreover, anti-gp120(CD4bd) Ab levels correlated with the amount of inhibition of gp120-specific Th proliferation in the presence of respective serum immunoglobulin G. These findings document a novel mechanism of HIV pathogenesis mediated by anti-gp120(CD4bd) Abs exhibiting suppressive activity on gp120 presentation. PMID- 12134257 TI - Staphylococcus aureus strains lacking D-alanine modifications of teichoic acids are highly susceptible to human neutrophil killing and are virulence attenuated in mice. AB - Staphylococcus aureus is resistant to alpha-defensins, antimicrobial peptides that play an important role in oxygen-independent killing of human neutrophils. The dlt operon mediates d-alanine incorporation into teichoic acids in the staphylococcal cell envelope and is a determinant of defensin resistance. By using S. aureus wild-type (WT) and Dlt- bacteria, the relative contributions of oxygen-dependent and -independent antimicrobial phagocyte components were analyzed. The Dlt- strain was efficiently killed by human neutrophils even in the absence of a functional respiratory burst, whereas the killing of the WT organism was strongly diminished when the respiratory burst was inhibited. Human monocytes, which do not produce defensins, inactivated the WT and Dlt- bacteria with similar efficiencies. In addition, mice injected with the Dlt- strain had significantly lower rates of sepsis and septic arthritis and fewer bacteria in the kidneys, compared with mice infected with the WT strain. PMID- 12134258 TI - Validation of immunoglobulin G enzyme-linked immunosorbent assay for antibodies to pneumococcal surface adhesin A in the diagnosis of pneumococcal pneumonia among adults in Kenya. AB - Epidemiologic studies of pneumococcal pneumonia, including vaccine efficacy trials, are hampered by a lack of sensitive and specific diagnostic tests. Pneumococcal surface adhesin A (PsaA) is a genetically conserved, surface expressed protein common to all serotypes of Streptococcus pneumoniae and is highly immunogenic. Detection of anti-PsaA immunoglobulin G by recombinant PsaA enzyme-linked immunosorbent assay was evaluated for diagnosis of pneumococcal pneumonia in paired serum samples from 4 adult populations: 47 healthy control subjects, 56 clinic control subjects without pneumococcal disease syndromes, 109 patients with pneumococcal pneumonia, and 93 pneumonia patients with no evidence of pneumococcal etiology. By considering a 2-fold increase in antibody concentration as positive, sensitivity was 0.70, and specificity was 0.98. With a 1.3-fold increase, these were 0.89 and 0.98, respectively. The test's performance was not affected by the patients' human immunodeficiency virus status or by the pneumococcal serotype. The combination of high sensitivity and high specificity makes this an ideal assay for epidemiologic studies of pneumococcal pneumonia. PMID- 12134260 TI - An outbreak of typhoid Fever in Florida associated with an imported frozen fruit. AB - An outbreak of typhoid fever in Florida involving at least 16 persons during the winter of 1998-99 was investigated using case-control, environmental, and laboratory methods. The genomic profiles of Salmonella serovar Typhi (Salmonella Typhi) isolates from the 15 confirmed case subjects were identical. Consumption of fruit shakes made with frozen mamey, a tropical fruit, was significantly associated with illness (matched odds ratio, 7.6; 95% confidence interval, 1.4 81.4). Laboratory testing showed that the fruit was heavily contaminated with fecal coliforms; no Salmonella Typhi was isolated. The frozen mamey was prepared in plants in Guatemala and Honduras. No further cases occurred after the frozen product was recalled. As our nation's food sources become increasingly globalized, the risk of outbreaks of exotic diseases linked to contaminated imported food will increase. This outbreak highlights the need for new approaches to ensure the safety of our food supply. PMID- 12134259 TI - Effect of the lower molecular capsule released from the cell surface of Bacillus anthracis on the pathogenesis of anthrax. AB - Bacillus anthracis enters the body as an endospore, and encapsulation and toxin production occur after germination. Capsule is proposed to be an antiphagocytic factor, and toxin induces cytokine production for systemic shock. The dep gene, adjacent to the cap region for the encapsulation, degrades the high-molecular weight capsule (H-capsule) to the lower-molecular weight capsule (L-capsule), which releases into the culture supernatant. This study analyzed the biological function of the cap-dep region. The dep null mutant Sm-1, which formed H-capsule but not L-capsule, was avirulent. However, Sm-1 with an intact dep gene or with purified L-capsule recovered its pathogenicity. Sm-1 was subjected to phagocytosis by macrophages more easily than its parent strain, Sm, in vitro; in vivo, it cleared without L-capsule and grew well with L-capsule, which suggests that L-capsule is essential for in vivo multiplication. Moreover, a new name, capD, might be appropriate, because of the part of the cap operon involved in both polymerization and depolymerization of the capsule. PMID- 12134261 TI - Cytokine release by lipopolysaccharide-stimulated whole blood from patients with typhoid fever. AB - The ex vivo cytokine response to lipopolysaccharide (LPS) of whole blood from patients with typhoid fever was investigated. Tumor necrosis factor (TNF)-alpha release by LPS-stimulated blood was found to be lower during acute typhoid fever than after a course of antimicrobial therapy (Por=4 consecutive months of negative sputum cultures, in prospective macrolide treatment trials of Mycobacterium avium complex (MAC) lung disease were assessed by pulsed-field gel electrophoresis (PFGE). Having >or=2 cultures positive for MAC after completion of therapy was documented 23 times; of 20 episodes studied by PFGE, 17 (85%) represented new genotypes (i.e., new infections), and 87% occurred in patients with nodular bronchiectasis. With >or=2 positive cultures after therapy was stopped prematurely, 6 (86%) of 7 episodes were relapses. Single positive cultures after completion of therapy occurred 16 times; only 1 (6%) was predictive of a subsequent relapse. No late isolates were macrolide resistant. Thus, relapses of MAC lung disease with these macrolide regimens are unusual, and most infections after completing therapy resulted from new strains in patients with nodular bronchiectasis. PMID- 12134266 TI - Superoxide anion production during Anaplasma phagocytophila infection. AB - Anaplasma phagocytophila persists within neutrophils and prevents the respiratory burst by inhibiting gp91(phox). Mutations in gp91(phox) result in chronic granulomatous disease (CGD), which is diagnosed by use of the nitroblue tetrazolium (NBT) and Fc-Oxyburst assays that examine whether cells produce O2-. This study assessed whether the NBT and Fc-Oxyburst assays could detect a respiratory burst during A. phagocytophila infection. O2- production was inhibited in HL-60 cells and neutrophils infected with A. phagocytophila. In a mouse model of A. phagocytophila infection, 15%+/-4% (mean+/-SD) of polymorphonuclear leukocytes from infected mice had an ineffective respiratory burst compared with 1%+/-1% (mean+/-SD) of the neutrophils from uninfected animals. A population of neutrophils that did not produce O2- was also detected in 2 patients with A. phagocytophila infection. These data demonstrate respiratory burst inhibition by A. phagocytophila in vivo and on an individual cell basis by use of assays designed to evaluate CGD. PMID- 12134267 TI - Human immunodeficiency virus type 1 infection in twin pairs infected at birth. AB - Host genetic factors may influence the course of human immunodeficiency virus (HIV) infection. In Blantyre, Malawi, polymerase chain reaction was used to identify twin pairs who were concordantly HIV-1-infected in utero or perinatally and then to examine strain divergence or virus levels in identical and fraternal twin pairs. Among 315 twin pairs, both infants in 14 fraternal and 5 identical pairs were found to be infected at the same visit. Among 10 pairs, HIV-1 sequences were determined for both infants at >or=1 time point. HIV levels had a common profile in both fraternal and identical twin pairs. Identical twins were not always infected by the same quasi species, indicating that their mothers had multiple quasi species capable of infecting their infants. Subsequent viral divergence appears to depend on quasi-species stability rather than on genetically controlled host immune responses. Thus, given infection, factors intrinsic to HIV-1 are more important than host genetics in viral evolution. PMID- 12134268 TI - Demonstration of a cell-mediated immune response in melioidosis. AB - Melioidosis is a bacterial infection caused by Burkholderia pseudomallei. The aim of this study was to determine whether a cell-mediated adaptive immune response against B. pseudomallei developed in patients who had recovered from melioidosis. Lymphocyte proliferation assays were done on peripheral blood mononuclear cells from patients (n=13) and control subjects (n=10) to determine the lymphocyte response to B. pseudomallei antigens. Production of interferon-gamma and interleukin-10 was also determined. Activation of T cell subsets was assessed by fluorescence-activated cell sorter analysis, using antibodies to CD4, CD8, and CD69 antigens. Lymphocyte proliferation and interferon-gamma production in response to B. pseudomallei antigens were significantly higher (P<.001 for both) in patients than in control subjects. There was also an increase in the percentage of activated CD4+ (P<.004) and activated CD8+ T cells (P<.035) in cell cultures from patients. The development of such a cell-mediated immune response in patients may be essential for their survival. PMID- 12134269 TI - Role of upper and lower respiratory tract immunity in resistance to Mycoplasma respiratory disease. AB - This study determined whether respiratory tract immunization protects against mycoplasma infection and compared preferential immunization of the upper respiratory tract (nasal) with upper and lower respiratory tract (nasal pulmonary) immunization. Small volumes of inoculum preferentially deposited antigen and induced IgA responses in nasal passages. Larger inoculums deposited antigen in both the upper and lower respiratory tracts, generating corresponding IgA responses. Mice were given nasal or nasal-pulmonary immunizations with Mycoplasma pulmonis antigen alone or with cholera toxin (CT), and resistance to infection was determined. Generation of upper respiratory tract immunity reduced mycoplasma infection at this site, but CT was needed to elicit protective responses. In the lower respiratory tract, nasal-pulmonary immunization was most effective, but nasal immunization did confer some protection from pulmonary infection. In contrast, intraperitoneal immunization resulted in little protection. Thus, respiratory tract immunity plays a major role in resisting mycoplasma infection, and it should be considered during vaccine development. PMID- 12134270 TI - [Nocturnal polysomnogram in childhood autism without epilepsy]. AB - AIMS: To evaluate the presence of epileptiform discharges and the organisation of nocturnal sleep of autistic children without nocturnal polysomnographic epileptic seizures. SUBJECTS AND METHODS: Cross section analysis. SUBJECTS: 21 boys and girls with autistic spectrum using DSM IV criteria between the ages of 4 and 12, compared with a control group made up of normal children of the same ages. METHODS: nocturnal polysomnogram with a minimum efficiency of 75%. ANALYSIS: t test to compare the cycles and phases of sleep with significance p< 0.05. RESULTS: SUBJECTS presented a maximum of four sleep cycles compared with five or six in the control subjects. From the first third of the night onwards there was an increase in the slowest phases. 66% presented epileptiform paroxysmal discharges, all of which originated in the anterior half of the brain. CONCLUSION: Sleep is not destructured, but it is reduced in length, with epileptiform paroxysms of predominantly frontal origin. This could indicate that these two parameters are intrinsic to the autistic spectrum, as well as indicating a more focused origin of the generalised picture which is possibly closely related with the qualitative alteration of the social experiences of these children. PMID- 12134271 TI - [Usefulness of P300 as a tool for diagnosing alterations in sustained attention in ischemic cerebrovascular disease]. AB - INTRODUCTION: Event related potentials, and especially the component P300, can be used to achieve greater precision and sensitivity in the cognitive evaluation of patients. They also enable us to obtain more accurate information about deficiencies in sustained attention, which is a cognitive function associated with the generation of the above mentioned component. Furthermore, the study of disorders triggered off by cerebrovascular disease must be given priority because of the high incidence with which they occur. PATIENTS AND METHODS: We studied four groups of individuals divided according to the localization of the lesion and the etiopatogenic mechanism involved (10 patients with right parietal atherothrombotic infarction, 10 patients with left parietal atherothrombotic infarction, 10 patients with lacunar infarction, 10 paired healthy controls). They were administered a classic test for obtaining P300, which evaluates sustained attention. RESULTS: Patients with cerebral infarction in the right hemisphere presented alterations in the functions evaluated, since they showed more omissions, gave fewer correct answers, the values for latency were significantly increased in Fz derivation and amplitude values were reduced in Pz, as compared to healthy controls. Individuals with left cerebral infarction, however, showed no significant differences with regard to the controls. People with lacunar infarction made less mistakes and omissions, and scored more correct answers than those that had suffered right or left infarction. Their amplitude values were also higher than the values of the individuals with a right lesion in Pz derivation but showed no differences as regards the controls. CONCLUSIONS: These results provide new data on the cognitive mechanisms that are affected during a right parietal lesion and demonstrate the sensitivity of P300 in detecting alterations in sustained attention in individuals that differ as regards not only the localization of the lesion but also the type of etiopathogenic mechanism involved. PMID- 12134272 TI - [Associated EMG responses in Alzheimer's disease and in healthy subjects]. AB - INTRODUCTION AND AIMS: Recently, the appearance of associated EMG responses in patients with probable Alzheimer s disease (AD) have been reported during the performance of cue stimuli recognition tasks using an oddball model. We present our work with AD patients and on the possibility of obtaining these responses in normal subjects by modifying the conditions under which the exploration is carried out. PATIENTS AND METHODS: We review 38 cases of probable AD, mild and moderate in intensity, in which we explored reaction time by bilateral EMG recording in the extensor muscles of the hand. The same exploration was performed in 20 healthy subjects in basal conditions and again while a simple mathematical task was being performed. RESULTS: In two of the 38 cases, both of moderate intensity, we observed associated EMG responses. On varying the conditions of exploration, 12 of the 20 normal subjects presented associated EMG responses. CONCLUSION: Associated EMG responses are a neurophysiological phenomenon that can be observed in some patients with AD. In normal subjects, it is possible to trigger the appearance of associated EMG responses by reducing their capacity to concentrate on the task to be performed. During the performance of unilateral motor tasks, inhibition pathways are activated physiologically to prevent the mirror contraction of contralateral muscles, which can be affected by the lowered capacity to pay attention. PMID- 12134273 TI - [Glia conditioned medium offers protection against the toxicity induced by 6-OH dopamine in vivo]. AB - INTRODUCTION: Glia conditioned medium (GCM) is neurotrophic for dopaminergic (DA) neurons and protects against MPP+ toxicity in vitro. We present the data from the first study in vivo of the effects of GCM. MATERIAL AND METHODS: The restorative effects were examined in rats with lesions produced by 6-hydroxy-dopamine (6-OH DA) in the medial longitudinal fasciculus. At four weeks, animals with an apomorphine induced rotation rate above three per minute were randomised for infusion with GCM, defined medium (DM) or saline through a striatal cannula for two weeks. To investigate the protective effects of GCM, the animals received a striatal injection of GCM or vehicle at the same time as the lesion was induced by 6-OH-DA and were sacrificed three weeks later. RESULTS: In the experiments on the restorative effect, the GCM infusion produced a significant increase in dopamine (DA) levels on the side with the lesion, in the limbic system (455.8 108.4 ng/g of tissue) and in the striatum (242.1 69.2 ng/g of tissue), as compared with the controls (110.8 36,4 and 108.4 22 ng/g). In the experiments on the protective effect, GCM induced higher levels of DA, 3,4-dihydroxyphenylacetic acid and 3 methoxytyramine (3-MT). In both models, the apomorphine induced rotation in animals with lesions caused by 6-OH-DA and treated with GCM was lower than that of the animals infused with DM or saline. CONCLUSIONS: These experiments show that GCM has protective and restorative effects in an experimental model of Parkinson's disease. PMID- 12134274 TI - [Autonomic cardiovascular dysfunction in multiple sclerosis not caused solely by brain stem lesions]. AB - INTRODUCTION: Multiple sclerosis (MS) can cause alterations in autonomic cardiovascular functioning. Some authors attribute these anomalies detected in MS patients to brain stem lesions, due to the fact that important autonomic nuclei are located in this region of the brain. Other studies, however, have been unable to prove such a relation. AIMS. The purpose of this study was to analyse whether these alterations had an exclusive relation with the brain stem lesions that usually appear in the course of this disease. PATIENTS AND METHODS: We compared the spectral analysis of the heart rate variability (SAHRV) in the frequency domain between a group of 34 patients with MS and another group of 14 patients with isolated brain stem lesions (IBSL) of a non inflammatory origin, which were measured using a 24 hour Holter recording. RESULTS: Heart rate, very low rates, low rates and the low rate/high rate quotient are significantly higher in the MS group than in the IBSL group, even when the former present brain stem lesions. Results in the high rate component, however, depend on the presence of brain stem lesions in the MS group. CONCLUSION: These findings suggest that autonomic cardiovascular dysfunction in MS is only related with brain stem lesions, although we obtained results that confirm the importance of this area in cardiovascular innervation. PMID- 12134275 TI - [Leigh syndrome resulting from a de novo mitochondrial DNA mutation (T8993G)]. AB - INTRODUCTION: Several degenerative neurological diseases are caused by mutations in the mitochondrial gene coding for subunit 6 of the ATPase. Thus, NARP (neurogenic weakness, ataxia, and retinitis pigmentosa) and Leigh syndromes are associated to a T8993G mutation when the percentage of mutant mitochondrial DNA is low (60 90%) or high (>90%), respectively. Leigh syndrome is also caused by a second mutation in the same position T8993C. CASE REPORT: The patient, a boy that died at 6 months, had generalized hypotonia, psychomotor delay, hepatomegaly, choreic movements and hyporreflexia. MRI showed hypodensities in the basal ganglia and brain stem as well as hyperlactacidemia. Molecular genetic analysis of the mitochondrial DNA showed that the patient had the T8993G mutation in a percentage higher than 95%. No mutated DNA was detected in blood of the proband s mother, maternal aunt and grandmother. CONCLUSIONS: The point mutation T8993G may occur de novo, at high levels, causing neurodegenerative diseases. PMID- 12134276 TI - [False negative diffusion in acute ischemic stroke]. AB - INTRODUCTION: The new techniques of magnetic resonance imaging have produced a important advance in the early diagnosis of acute ischemic stroke. The diffusion weighted imaging (DWI) has a high sensitivity and specificity in the acute ischemia. CASE REPORT: 70 years old woman with previous history of hypertension and dyslipemia. The patient presented sudden vertigo with cervical neck pain and gaze problems. The physical exploration revealed right miosis, nistagmus, IX and X cranial nerve affection and ataxia. The clinical diagnosis was Wallenberg s syndrome and a medulla lateral infarction. However DWI was normal (10 hours). The neurological deficit was persistent and a repeated study disclosed a lateral infarction of the medulla. CONCLUSIONS: The sensitivity of DWI is reduced during the first 24 hours. There are false negative diagnosis mainly in small infarctions of posterior territory. PMID- 12134277 TI - [Opercular syndrome of vascular aetiology]. AB - INTRODUCTION: Facio linguo masticatory paralysis resulting from bilateral lesion of the anterior opercular region, known today as Foix Chavany Marie syndrome, appears very frequently in adult patients as a consequence of generally ischemic vascular lesions, which directly affect the Rolandic opercula or the subcortical area surrounding them. However, forms that are secondary to infections, neoplasias and even unilateral lesions have been reported. CASE REPORT: Female patient, aged 70, with unknown auricular fibrillation, which started suddenly with right faciobrachial paresis secondary to a left striated ischemic infarction. Approximately 24 hours after beginning intravenous heparinization, the patient presents anarthria, facial hypomimia without asymmetries, dysphagia above all for liquids and bilateral lingual paresis. Extraoccular movements, blinking and corneal reflex were normal. Understanding of spoken and written language was maintained and there were no apraxias or agnosias. Neurological exploration was compatible with an opercular syndrome (OS). Urgent brain CT revealed the existence of a right cortiico subcortical temporal haemorrhage, in addition to the prior contralateral ischemia. The association of both lesions, ischemic and haemorrhagic, mirrored, justified all the symptomatology. DISCUSSION: Anterior OS is characterised by a bilateral voluntary central paresis of the muscles innervated by the 5th, 7th, 9th, 10th and 12th cranial pairs with preservation of emotional or automatic mobility. Underlying mechanisms are also discussed. PMID- 12134278 TI - [Myasthenia and thymoma in a seven year old girl]. AB - INTRODUCTION: Myasthenia is an autoimmune disease, being generalized muscular weakness, with important participation of facial muscles, a prominent feature. Signs of muscular fatigue arise, worsened by exercise and alleviated by rest. Clinical symptoms are less marked before noon, and get worse as the day advances, through the afternoon and evening. A clear relationship between myasthenia and thymic abnormalities does exist, being glandular hyperplasia and tumours the commonest underlying pathologic findings. Initial treatment is based on anticholinesterase drugs and steroids. Non respondents should be treated with immunoglobulins, immunosuppresses, plasmapheresis and surgical removal of the thymus, according to the symptoms control. CASE REPORT: We present the case of a seven years old girl with generalized muscular weakness, worsening through the day, being the diagnosis of myasthenia confirmed by the high level of acetylcholine antireceptors antibodies and the neurophysiologic study. Imaging study of the mediastinum showed a thymic mass located in the right lobe. CONCLUSION: It is therefore most important to rule out these conditions when myasthenia is suspected. PMID- 12134279 TI - [Findings using magnetic resonance in a patient with non-traumatic anosmia]. AB - INTRODUCTION: The presence of alterations in the neuroimaging in patients with anosmia without traumatic antecedents is not frequent. CASE REPORT: Male aged 38 who came to surgery after having suffered, 6 months earlier, for 1 week, a picture of intense, oppressive holocranial headache, accompanied by fever. Associated with this, an acute complete anosmia also began and persisted up to the moment the patient came for consultation. It was not associated with any infection of the respiratory tract, there was no history of cranial trauma, no ingestion of medicines nor toxins, nor had he been exposed to toxic products. The exploration to which he was submitted only showed an anosmia and was otherwise found to be normal. Cranial MRI showed signal alterations in both lower (orbitary) convolutions of the frontal lobes, in the anterior region of the right temporal lobe and in both olfactory nerves. Tests for HIV serology, parotiditis, hepatitis B and C virus, HSV, VZV, Mycoplasma pneumoniae and lues were negative. The acute onset of the anosmia in midst of a picture of febricula and headaches made us suspect the presence of an infectious aetiology, and the alterations found in the neuroimaging could be due to post encephalic lesions, with a special predilection for olfactory areas. CONCLUSIONS: 1. MRI plays a fundamental role in the topographic and aetiological evaluation of olfactory dysfunctions of a central origin; 2. Affectation of the central olfactory passages of an infectious aetiology in a non HIV patient and with neuroimaging findings is a rare complication. PMID- 12134280 TI - [Percutaneous placement of covered stents for treatment of post discectomy arterial rupture]. AB - INTRODUCTION: Vascular lesions produced as a consequence of surgical spinal treatment are rare, but serious. Fast diagnosis and treatment are essential in lesions that produce massive blood loss. Following the use of non covered stents in the treatment of peripheral vascular stenotic obstructive pathologies, the appearance of covered stents allowed aneurysms, pseudo aneurysms, fistulas or bleedings to be treated. Being able to graft a stent in serious patients, during diagnosis itself, with a minimum amount of aggression and local anaesthesia affords this technique huge advantages over others. CASE REPORT: In this work we report the case of an arterial rupture which came about as a complication of a discectomy. After the surgical intervention, there was an important haematocrit reduction within a short space of time. The patient s quickly becoming anaemic and the existence of a retroperitoneal mass of heterogeneous echogenicity pointed towards a postoperative haemorrhage. This was confirmed by arteriography, which revealed an important contrast extravasation. A covered stent was placed at the same time and, in the post graft follow up, it was confirmed that there was no bleeding. At 12 hours after the operation the patient was moved from the postoperative recovery unit to a hospital ward and, 5 days later, was discharged. In a check up carried out in the Neurosurgery unit 2 months after being sent home, the patient was found to be asymptomatic. CONCLUSION: The placement of covered stents is a valid therapeutic option for treating postoperative haemorrhagic lesions. It can be performed at the same time as diagnosis and does not require general anaesthesia. In the case we have described, the patient recovered quickly and ambulatory check ups showed he continued to be asymptomatic PMID- 12134281 TI - [Multiple latency test in a patient with episodes of sleep induced by pergolide]. AB - OBJECTIVE: Recently, there have been report sleep attacks in parkinsonian patients as a side effect of pramipexole and ropinirole. We report a patient with similar episodes related with pergolide. CASE REPORT: A 64 year old man with rigid akinetic parkinsonism, treated with carbidopa/levodopa and pergolide, developed sudden, irresistible sleep episodes after increasing the dose of pergolide to 2.25 mg/day because of bad control of parkinsonian symptoms. These episodes started 30 minutes after each dose of pergolide and lasted 2 hours. Following reduction of the dose of pergolide to 1.5 mg/day the sleep episodes disappeared. Two double blind multiple sleep latency tests were performed, one after intaking pergolide and other after intaking placebo. RESULTS: The latencies to sleep onset were lower with pergolide than with placebo, but the differences did not reach statistical significance. There was no premature REM sleep onset. CONCLUSION: Sleep episodes are likely a not specific effect of dopamine agonists PMID- 12134282 TI - [Is the hippocampus necessary for spatial learning?]. AB - OBJECTIVE: A variety of findings, mainly based on lesion studies, have led to the proposal that the hippocampus is a necessary component of the neural system supporting spatial learning. However, recent research is challenging this classic view. The goal of this review is to discuss and integrate these new data with previous findings. DEVELOPMENT: Several studies performed in our lab have shown that hippocampal lesions do not affect the acquisition of a place response if a special training procedure is used. Nevertheless, the way how this special training method overcome the learning deficit is not well known. In this review we propose that hippocampal damage produces several non spatial deficit, being one of them a deficit in behavioral flexibility. Thus, it is possible that our special training procedure encourage behavioral flexibility and variability in response in lesioned subjects, which would allow a significant improvement in the learning of the spatial task. CONCLUSION: The data discussed in this review suggest that the hippocampus is not completely necessary for the learning of a place. It would be the sum of spatial and non spatial deficit ('hippocampal syndrome') which would produce the acquisition deficit typically seen in hippocampal rats when a traditional training method is used. PMID- 12134283 TI - [Endocranial hypertension]. AB - INTRODUCTION AND OBJECTIVE: The endocranial hypertension syndrome is one of the commonest and most feared neurological complications in clinical practice. This encourages its continued study, and is the subject of this review. DEVELOPMENT: Endocranial hypertension is the common pathway for the presentation of many neurological and non neurological disorders. An increase in the volume of one or more intracranial structures causes secondary lesions of the brain and even the death of the patient, although this may frequently be avoided by prompt recognition and suitable action by the doctor involved. General, non specific therapeutic measures should be instituted to achieve a normal endocranial pressure. The clinical presentation and complementary investigations should be correctly interpreted to determine the aetiology of the condition so that specific treatment may be given. Continuous monitoring of the intracranial pressure and other neuromonitorization techniques are essential for correct decisions as to treatment. CONCLUSIONS: Understanding the physiopathology, diagnosis and treatment of the syndrome of endocranial hypertension are still essential in modern medicine. In recent years there have been advances in the use of well known treatments and new drugs have been introduced. Use of the different neuromonitoring techniques permits optimum use of the treatment available. PMID- 12134284 TI - [Cognitive processes and emotional brain systems]. AB - AIMS: In this review we analyse the role of learning and memory processes in normal and pathological emotional phenomena from an integrating perspective that combines the behavioural, cognitive and neurobiological levels. We examine the most recent trends in research into the emotional systems of the brain, which provide important keys to understanding the nature of emotional phenomena. DEVELOPMENT: First, we discuss the existing evidence about the non conscious generation of affective responses, which come mainly from studies into Pavlovian fear conditioning. We then go on to analyse the relevance of the distinction between implicit and explicit memory in order to get a better understanding of the relations between cognition and emotion. The role played by the amygdala in the automatic evaluation of danger is considered a basic model for eliciting emotional responses. Finally, we describe how the interaction of the brain systems that the implicit and explicit aspects of emotional memory depend on provides an explanation for several characteristics of pathological anxiety, and we examine the positive or negative influences emotional activation can have on the consolidation of memory. PMID- 12134285 TI - [Cognitive deficit in perinatal asphyxia]. AB - INTRODUCTION: During the period 1950 1970 the relation between perinatal asphyxia and cognitive alterations was studied. More recently the neuropsychological approach has been introduced to detect more subtle defects. DEVELOPMENT: With regard to intelligence, alterations in the intelligence quotient resulting from anoxia are more commonly seen in young children than in adolescents, probably because of compensation mechanisms. It is widely accepted that severe asphyxia causes motor and cognitive alterations and leads to cerebral palsy, epilepsy and intellectual retardation. The effects of mild or moderate asphyxia are still controversial. CONCLUSION: Thorough neuropsychological examination, particularly assessment of memory and frontal function, helps to identify subtle deficits which may explain some learning problems observed in children who have a history of moderate asphyxia but previously no clear neurological diagnosis. PMID- 12134286 TI - [The secondary effects of the antihistamine chlorpheniramine on the central nervous system]. AB - OBJECTIVE: The objective of this study is to review the main investigations into the secondary effects of the antihistamine chlorpheniramine on the central nervous system (CNS). DEVELOPMENT: The antagonists of the H1 receptors of histamine, usually used in the treatment of symptoms of allergy or the common cold, have many adverse effects on the CNS. They cause day time drowsiness, cause poorer performance of tasks involving visuo motor coordination and make it more difficult to detect target auditory stimuli in tasks involving sustained concentration. When using evoked potentials (EP) it has been observed that they alter the system for maintaining auditory attention. They cause increased P300 latency, an EP related to the voluntary ability to discriminate between relevant stimuli, a reduction in the amplitude of mismatch negativity (MMN), an EP which is seen as a pre attention mechanism for automatic detection of environmental acoustic changes and alters selective attention capacity, reflected by a reduction in the amplitude of processing negativity (PN). CONCLUSIONS: These studies show that chlorpheniramine has major adverse effects on the CNS, and the patient may not be subjectively aware of this (e.g. selective attention). This means that in certain situations it is a dangerous substance. The characteristics of these adverse effects should lead to a review of the prescription of chlorpheniramine, and stimulate the search for other substances with similar therapeutic actions but fewer side effects on the CNS. PMID- 12134287 TI - [Neuroscience in ancient Egypt and in the school of Alexandria]. AB - INTRODUCTION: About five thousand years ago, one of the most ancient, important and enduring civilisations in the history of Mankind flourished on the banks of the Nile. In Egypt, all the branches of human knowledge attained a high degree of development and today it is considered the mother of medicine. Despite the marked religious component that affected all its activities, the medicine of the Pharaohs was practised in a rational and deductive manner, and the Egyptians were the inventors of clinical observation. METHOD: The desert climate of the Nile Valley has preserved monuments, mummies and papyruses which have enabled us to get a certain idea of the degree of development reached in medical matters. The aim of this work is to analyse them from a neuroscientific point of view. The anatomical discoveries of the Egyptians originated in the inspection of wounds and the practice of embalming. They came to know a large number of diagnoses and were able to prescribe many different forms of treatment. They also attained a certain proficiency in dealing with neurotraumatological patients. They practised anamnesis, prognostics and a regulated surgery that infrequently included trephination. Their conservatism meant that, after the Macedonian domination, the traditional Egyptian medicine was replaced the Greek medicinal arts, which reached their maximum period of splendour in the School of Alexandria, where Herophilus and Erasistratus, pioneers in the study of anatomy and brain circulation, were especially renowned. CONCLUSIONS: From the point of view of the neurosciences, the Egyptians were the first to describe the brain, migraine, epilepsy, strokes, tetanus, Bell s palsy and the sequelae of head injuries and of spinal transection. Their artwork sometimes shows neurological patients and, according to Herodotus, there were doctors who were specialised in head diseases and could therefore be considered the precursors of our present day neurologists. PMID- 12134288 TI - [Deafness and ataxia as the beginning of meningococcal meningitis]. PMID- 12134289 TI - [The treatment of neurosarcoidosis: a case report]. PMID- 12134290 TI - [Lethal nemalinic myopathy and congenital arthrogryposis or fetal hypokinesia sequence?]. PMID- 12134291 TI - [Clinical diagnostic of brain death and transcranial Doppler]. PMID- 12134292 TI - [Narcolepsy in children]. PMID- 12134294 TI - [The impact of sociofamilial factors on nutritional status in mentally retarded children]. AB - INTRODUCTION: Mental retardation (MR) constitutes a clinical and social relevant condition accounting for 3% of the pediatric population. Studies focusing the repercussion or MR on nutritional status are scarce and, in occasions, have produced contradictory results. OBJECTIVE: To evaluate the nutritional status of mentally retarded children in our region, on the basis of the influence of sociofamilial factors, including details of diet and appetite. PATIENT AND METHODS: Our sample comprise 128 mentally retarded children (81 boys and 47 girls) aged 0 17 years. In all children a nutritional and social family environment questionnaires and a valuation of a series of nutritional and anthropometric variables were performed. A factorial analysis was carried out by means of the statistical package SPSS allowing the obtaining of 2 anthropometric factors (AF) and 3 biochemical factors (BF) that condensed the most information content. Results of the nutritional and sociofamilial questionnaire were correlated with biochemical and anthropometric factors. RESULTS: Score of the factor AF1 declined with declining quality of diet and appetite. Mean AF1 score was lowest among children from inland rural areas, intermediate among children from urban areas and highest among children of coastal areas. Age of the parents and number of brothers also influenced the value of AF1 score. Score of AF1 was not significantly affected, however, by social class. Quality of diet, appetite, geographic origin, number of brothers an age of the parents showed a similar influence on BF1. Moreover, the score of BF1 declined with declining socioeconomic status. CONCLUSIONS: Feeding behaviour has a significant influence on nutritional status both in biochemical and anthropometric parameters, so it must be promptly evaluated in mentally retarded children. Biochemical parameters were the most influenced by variation of socioeconomic status. Children from coastal areas showed the highest scores of nutritional parameters. Age of the parents significantly influenced the nutritional state. PMID- 12134295 TI - [Prevalence of myasthenia gravis in Antioquia, Colombia]. AB - INTRODUCTION: Myasthenia gravis (MG), considered the commonest of all the illnesses that affect neuromuscular transmission, is a disorder in which the autoimmune system attacks the post synaptic acetylcholine receptor proteins in the end plate terminal; it is characterised by weakness and skeletal muscle fatigue, with no anomalies in reflexes, sensitivity or coordination. Epidemiological indicators, such as incidence and prevalence, are not known in Colombia. AIMS. To determine the prevalence of MG among the inhabitants of Antioquia, through the use of the capture recapture method. PATIENTS AND METHODS: The capture recapture method was used for two sources, the Instituto Neurologico de Antioquia and the Hospital Universitario San Vicente de Pa l, which are the most important institutions for the diagnosis of neurological diseases in Antioquia. MG prevalence was calculated using the following formula: p= n/N 105. We examined the data from the period between 1 July 1995 and 30 June 2000 with the aim of identifying subjects who fitted the profile of MG sufferers. RESULTS: General MG prevalence in Antioquia was 27.7 cases per million inhabitants (CI 95%= 23.2 32.2). The male/female ratio was 1:3.77. CONCLUSIONS: The estimated prevalence of MG is lower than that reported in United States and other temperate regions, where it varies between 60 and 150 cases per million. The prevalence of MG is low in Antioquia, as in other tropical areas PMID- 12134296 TI - [A control and case study of multiple sclerosis in the Alicante and Villajoyosa areas]. AB - INTRODUCTION: Three studies have been carried out in the area of Alcoy (Spain) which show an association between contact with dogs and multiple sclerosis. We present a control and case study conducted in a different geographical area, the area of Alicante. AIMS. To examine environmental factors linked with multiple sclerosis in the health care district of Alicante, and to compare findings with those from the health care district of Alcoy. PATIENTS AND METHODS: A study of cases and controls was conducted with 47 patients, each of which was paired with four controls according to gender, age and place of residence. All the cases fulfilled criteria of defined multiple sclerosis. The controls were obtained from the Emergency and Casualty Services at different hospitals in the area. Each of the environmental factors studied was stratified by genders. RESULTS: A total of 47 patients and 188 controls were analysed. A statistically significant association was found between the disease and smoking, a low educational level, a social level 2 on the classification developed by Koch Henriksen, having had measles before the age of 15 and contact with dogs. The fact of being the third, or later, child in a family and a family history of cephalea were more frequent among controls than among cases. CONCLUSIONS: Multiple sclerosis in the Alicante area is related with contact with dogs, smoking, a low educational level, a high social level and measles before the age of 15. Since an association between contact with dogs and multiple sclerosis has been shown repeatedly in other previous studies conducted in the town of Alcoy, we are now in a position to state that this factor is indeed linked with the appearance of multiple sclerosis. PMID- 12134297 TI - [Use of event related potentials for the diagnosis and follow up of sub clinical disorders of sustained attention in ischemic cerebrovascular disease]. AB - INTRODUCTION: Ischemic cerebrovascular disease causes cognitive disorders among survivors, which may range from vascular dementia to alterations of a sub clinical nature. These are not easily detected using conventional diagnostic tools. Some of the most frequent disorders after a cerebral infarction are those affecting sustained attention or vigilance. The event related potential (ERP) P300 or P3 could be a valuable diagnostic indicator of this type of disorder and its evolution. AIMS. The evaluation and diagnosis of sub clinical disorders of sustained attention in patients with cerebral infarction by using ERP as a method of diagnosis and follow up. PATIENTS AND METHODS: We studied 13 patients with cerebral infarction located in the left parietal lobe, 13 with infarction in the right parietal lobe and 26 healthy control subjects, who were paired by gender, age and educational level. All subjects were submitted to a computerized continuous performance test, associated with the recording of electrical brain activity. RESULTS: Patients with right parietal lesions made more mistakes due to omission, answered fewer questions correctly and had a lower attention index than the rest of the patients and healthy controls. Although there were no significant differences in the P3 amplitude values, patients with right lesion presented significantly increased latencies in the F7, Fz and Cz derivations. After five months there was a significant improvement in performance in the attentional test. The value of the latency in the Cz derivation, nine days after the occurrence of the cerebral infarction, was related with the number of mistakes made by the subjects five months later in the attention test. CONCLUSIONS: Patients with an ischemic lesion in the right parietal lobe presented disorders in sustained attention, which were only diagnosed adequately by use of computerized tests associated with ERP recordings. This shows that evoked potentials are valuable in cognitive diagnosis, and also demonstrates the relation that exists between the capacity to hold the attention on a stimulus over a period of time and the structures of the right parietal lobe. Five months later, these disorders of a sub clinical nature can disappear. The value of the latency in Cz, nine days after the occurrence of the event, acts as a predictor of this recovery. Cognitive rehabilitation plans could benefit from the use of these diagnostic tools. PMID- 12134298 TI - [Abnormal antioxidant system in inborn errors of intermediary metabolism]. AB - INTRODUCTION: Oxidative stress may be implied in the pathogenic mechanisms of inborn errors of intermediary metabolism (IEIM). OBJECTIVE: The evaluation of the antioxidant status in IEIM by the measurement of erythrocyte antioxidant enzyme activities, superoxide dismutase (SOD), glutathione peroxidase (GPx), glutathione reductase and catalase. PATIENTS AND METHODS: 34 patients with IEIM: 1) eleven with organic acidurias on protein restricted diet; 2) nine without special diet; 3) five patients with aminoacidopathies on protein restriction; 4) three patients with galactosemia and six with aminoacidopathies on protein free diet. Erythrocyte antioxidant enzymes were measured by spectrometric procedures adapted to the Cobas Fara II analyser. RESULTS: SOD activity was significantly higher in groups 2 and 4 (p= 0.009, p= 0.001, respectively), and significantly lower in group 3 (p= 0.001) compared with age matched controls. SOD activity was significantly higher in the patients with IEIM on protein free diet (groups 2 and 4) compared with those on protein restricted diet (groups 1 and 3; p= 0.002) or with controls (p= 0.003). GPx activity was found significantly lower in group 1 patients (p= 0.004), and higher in group 2 (p= 0.029) compared with controls. CONCLUSIONS: 35% of the patients with IEIM had SOD activity above the control range, most of them with organic acidurias or homocystinuria, suggesting an induction of enzyme protein synthesis owing to an excess of free radical generation. The lower activities observed in patients on natural protein restriction may likely be due to a deficient bioavailability of antioxidant cofactors. PMID- 12134299 TI - [Defects in the visual field in resective surgery for temporal lobe epilepsy]. AB - INTRODUCTION: Temporal lobe resection is the most used procedure in epilepsy surgery. Visual field defects after resection are frequent. AIM. We try to detect the frequency and severity of visual field defect after anterior temporal lobectomy (ATL) and to study the functional consequences. PATIENTS AND METHODS: We studied 30 patients with refractory temporal lobe epilepsy (15 men, 12 51 years, m= 32.9), in which ATL was performed (11 right, 19 left). The neocortical and hippocampal resection was variable. Visual field was studied with computerized campimetry type Humphrey. Our survey was filled to study the campimetric consequence, seizures incidence, quality of life and surgery satisfaction. RESULTS: Some campimetric disturb was found in 27 patients (90 %), superior or minor homonimus quadantapnosia in 18 (60 %), major one 8 and homonimus hemianopsia in one. Incongruent defects appeared in 19. The bigger neocortical and hippocampal resection was, the bigger defect, with exceptions. Only patient with a hemianopsia was conscious the deficit. 20 patients (66.6 %) were seizures free in the last year. 28 had better quality of life and 29 (96.6 %) would have chosen surgery again. CONCLUSION: Visual field defects after ATL, although frequent, have little functional consequences and in quality of life. The quality of life is better when seizures stopped. After ATL patients are very glad with their decision. PMID- 12134300 TI - [In vitro viability and glutathione levels in mesencephalic neurons after seven days hibernation]. AB - In embryonic mesencephalic transplant in patients with Parkinson s disease dopaminergic survival is low (5 10%), and for this reason the use of multiple donors has been considered. The difficulty of obtaining more tissue determines the need for a procedure that enables human nigral tissue to be stored for a time without affecting its physiological state in any significant way. This study was designed to determine whether hibernation of tissue fragments has any influence on viability, how the viability of the mesencephalic cells behaves after 7 days hibernation and the glutathione levels in the hibernated tissue (HT). The viability of the HT in pieces (82.37 2.12) was found to be higher than the value for the whole mesencephalon (70.29 3.43). Viability of the HT, seven days at 4 C, at different post dissociation times, did not differ significantly. Despite the significant differences found between hibernated and fresh tissue at t= 0, this procedure does not seem to affect the mesencephalic tissue in any significant way, as it conserved a 94% viability after hibernation. No evidence was found of increased glutathione content as an antioxidizing response to the damage that might be caused by hibernation. These results suggest that since hibernation does not have any significant effect on the state of the cells it could be considered a useful procedure for conserving tissue to be used in clinical transplants. Moreover, further research is needed on survival and functionality of hibernated cells after being transplanted into animal models in order to evaluate their potential for use in cell therapy. PMID- 12134301 TI - [Neuroepidemiology in the eastern region of Colombia]. AB - OBJECTIVE: To determine the presence of eight neurological disorders (migraine, cerebrovascular disease, disorders of movement, peripheral neuropathy, mental retardation, epilepsy, dementia and the sequelas of head injuries) in the eastern region of Colombia. PATIENTS AND METHODS: Using the neuro epidemiological protocol of the World Health Organization (WHO), modified by our group, together with a questionnaire to determine the sequelas of head injuries and the abbreviated mental test (Mini mental), we interviewed people in the municipalities of Bucaramanga, Piedecuesta and Aratoca. The persons suspected of having neurological disorders, who were over 12 years old, were evaluated by neurologists and those under 12 years old by a neuropaediatrician. RESULTS: In the eastern region, 1,454 persons altogether were interviewed. Neurological conditions were suspected in 454 of these (31.2%), and confirmed in 437 (30.1%). The specific results for these neurological disorders and their respective confidence intervals (in brackets) were: migraine 198.8 (178.7 220.4); peripheral neuropathy: 26.8 (19.4 36.9); epilepsy: 22.7 (15.9 32.1); dementia: 17.9 (11.9 26.5); cerebrovascular disease: 17.2 (11.4 25.7); mental retardation and delayed nervous system development: 15.1 (9.7 23.2); extrapyramidal disorders: 8.3 (4.5 14.8); sequelas of head injuries: 6.9 (3.5 13.1). RESULTS: These results, obtained using a modified version of a WHO protocol, together with dementia and the sequelas of head injuries, will permit the development of health policies and programmes for the control and treatment of neurological disorders prevalent in this region of Colombia PMID- 12134302 TI - [Hypernephroma associated motor neuron disease. Presentation of a case]. AB - INTRODUCTION: Paraneoplastic motor neuron disease are rare among patients with renal cell carcinoma. OBJECTIVE: Present the clinical and electrophysiological evolution of a patient with a motor neuron disease and hypernephrome. CASE REPORT: A 60 years old woman, affected only by high level pressure since 10 years ago. She noticed sudden palsy of the left leg and 10 months later an abdominal ultrasound showed a renal cell carcinoma, discovered without other symptoms than neurologic. After radical nephrectnomy, the patient was treated with recombinant interferon alpha 2b. The neurologic damage advanced and she has a flaccid weakness and muscle atrophy in the legs and brisk reflexes, also in wasted limbs. There is Babinski, fasciculations, light flaccid dysarthria and laryngospasm. Peripheral nerve conduction studies are within normal limits. The electromyogram show positive sharp waves in both legs and left hand. Recruitment patterns are decreased and there are fasciculations in the tongue, upper and lower limbs. The magnetic resonance of the brain and spinal cord is normal. There are not evidence of metastasis. The question of whether or not this is a paraneoplastic form of motor neuron disease remain unclear. CONCLUSION: This case suggest the need to consider a renal cell carcinoma in the course of a motor neuron disease. PMID- 12134303 TI - [Processes of logical thought in a case of cerebral vascular lesion]. AB - INTRODUCTION: Reasoning and logical thought processes have traditionally been attributed to frontal lobe function or,on the other hand, have been considered as diffuse functions of the brain. However, there is today evidence enough about the possibility to find dissociations in thought processes, depending on logical structure of the experimental tasks and referring to different areas of the brain, frontal and post rolandic ones. OBJECTIVE: To study possible dissociations between thought structures corresponding to categorical and relational logic, on one hand, and propositional logic on the other hand. CASE REPORT: The case of a brain injured patient with vascular etiology, localized in left frontal parietal cortex, is presented. A specific battery of reasoning tests has been administered. RESULTS: . A differential performance at some reasoning experimental tasks has been found depending on such logical conceptual structures. CONCLUSIONS: The possibility of establishing dissociations among certain logical thought and intelectual functions depending on localization of possible brain lesion (frontal versus temporal) is discussed. PMID- 12134304 TI - [Infection due to Mycoplasma pneumoniae: three cases with neurological complications]. AB - INTRODUCTION: Mycoplasma pneumoniae infection has been associated with severe central nervous system diseases. The pathogenesis of these disorders is unknown and the treatment uncertain. CASE REPORTS: The authors present three cases of central nervous system diseases: acute transverse myelitis, cerebellitis and encephalomyelitis associated with M. pneumoniae infection. CONCLUSIONS: M. pneumoniae infection should be considered in all cases of severe acute central nervous system symptomatology. PMID- 12134305 TI - [Nicotinic Receptor, galantamine and Alzheimer disease]. AB - Population aging has increased and will drastically increase the prevalence of Alzheimer disease. The disease develops inexorably towards a syndrome of marked cognitive impairment, accompanied of emotional alterations and profound changes of personality. The patient loses its autonomy, and requires special attention of caregivers; this leads to a decrease of the quality of life, not only of the patient but also of its caregivers and family. The reduction of the number of functional nicotinic receptors in brain keeps pace with neurological symptoms and the severity of the disease (cholinergic theory of Alzheimer disease). There is a pleyade of data and observations reinforcing the idea that improving cholinergic neurotransmission is an investment in memory. Up to now, although with limited success, this improvement has been achieved only with the reversible inhibitors of acetylcholinesterase tacrine, rivastigmine and donepezil, available in the clinic since a few years. The last approved has been galantamine that in spite of being a modest inhibitor of acetylcholinesterase, improves memory (ADAS cog test) and slows down cognitive impairment of Alzheimer patients. To explain this therapeutic effect, a second mechanism of action for galantamine has been suggested, the positive allosteric modulation of presynaptic nicotinic receptors, that will favour the release of acetylcholine and other neurotransmitters involved in memory formation. Furthermore, galantamine possesses neuroprotectant antiapoptotic effects, according to recent data from our laboratory. These effects provide new ideas and therapeutic targets that might help to find novel and efficacious treatments for patients suffering Alzheimer disease. PMID- 12134306 TI - [Persistent vegetative state. Paradigm of current discussion on alterations of consciousness]. AB - INTRODUCTION: The vegetative state is the current paradigm of discussions about the alterations of conscience. DEVELOPMENT: Although accepted by most investigators, it is still controversial. The dilemma starts with the denomination itself. In this paper we propose the denomination persistent vegetative state . We start with a historical review of the integration of consciousness. We also give epidemiological data and point out the clinical features, complementary tests and anatomical findings. The patients are classified into three grades for prognosis. Grade III includes those with the worst prognosis, who have no sleep waking cycles with or without opening their eyes. This section emphasises cases of prolonged survival and of late recovery who made almost complete recovery of their intellectual functions. We state that treatment is based on two aspects: treatment of the underlying disease and general measures and emphasise the need for a multidisciplinary team. From the bio ethical point of view, it should be remembered that the patients are alive and cannot be considered in the same group as the brain dead, in whom all encephalic function has been lost. CONCLUSIONS: It is not ethical to decide to suspend medical treatment when it is known that there is a possibility of recovery of the structural anatomy and function. We are morally obliged to maintain qualified medical attention. It has been shown scientifically that we not only should, but can, obtain the recovery of these patients, in spite of the serious damage suffered by their nervous system PMID- 12134307 TI - [Neuropsychology of cerebral palsy]. AB - INTRODUCTION: Cognitive performance in cerebral palsy (CP) varies between being completely normal and severe mental retardation. This heterogeneity depends on several factors, including the type of paralysis and associated cerebral lesions. DEVELOPMENT: We review neuropsychological studies with reference to general cognitive performance and specific performance (language, memory, attention and visuospatial functions). We also consider studies relating neuroimaging and neuropsychology in the different types of CP. CONCLUSIONS: Language is one of the best preserved functions studied in CP, apart from effects due to the lesion itself. Defects of articulation may affect understanding but not to any great extent. Immediate memory also seems to be relatively well preserved. In general no effects of lateralization are seen with regard to language or memory, although this may be seen when carrying out complex tasks or when involving attention and visuoconstructive tasks. The relatively few studies relating neuroimaging and neuropsychology are inconclusive regarding specific abilities. PMID- 12134308 TI - [Natural evolution of radiculopathies and myelopathies caused by cervical arthrosis]. AB - AIMS: From the data available in the literature, to analyse the natural history of acute or chronic radiculopathies and myelopathies linked with degenerative disease of the cervical spine. DEVELOPMENT: By means of different electronic and manual search methods, we located original pieces of work dealing with recent objectives and reviews on the subject. We have attempted to take into account any data that might be relevant, regardless of the characteristics or quality of the work in which it was published. The high prevalence of degenerative changes in the cervical region in the general population, the fact that the development of the concept of cervical radiculopathies and myelopathies has gone parallel to the development of their surgical treatment, and that there are few series with a control group and independent evaluation and no trials with random assignation do not allow a systematic approach. The anatomical and radiological aspects of cervical arthrosis are well known, but there is little information available on the causal relation between both processes and their clinical translation or their natural evolution. CONCLUSIONS: In the present state of knowledge, it is not possible to establish unmistakable relations between cervical disc disease and the radiculopathies and myelopathies it is attributed with. The natural evolution of these neurological syndromes is not known and therefore neither is it known how this could be modified by conservative or surgical treatment. From critical points of view, there is a unanimous claim for clinical trials to be performed with random assignation, which would provide us with an understanding of the natural history of these entities and the role of the different surgical or conservative treatments available PMID- 12134309 TI - [The history and social importance of cerebrovascular pathology]. AB - In this work we present a study of the different conceptual, physiopathological, anatomical and clinical aspects that cerebrovascular pathology has had throughout history, from the early pre scientific beginnings of religious medicine, through the Renaissance period up to the Modern and Contemporary Ages. In the second part we emphasise its social importance and economic cost, and new methodologies are proposed for use in its study and pursuit. We also highlight the need for the creation of stroke units. PMID- 12134310 TI - [Spindle coma secondary to thyrotoxicosis]. PMID- 12134311 TI - [Appraisal of electrocardiographic alterations in acute cerebrovascular disease]. PMID- 12134312 TI - [Is treatment with a daily dose of Depakine Crono safe?]. PMID- 12134313 TI - [Satisfactory treatment of a suprasellar subarachnoid cyst by use of ventriculoperitoneal shunt]. PMID- 12134314 TI - [Pregnancy and epilepsy]. PMID- 12134316 TI - [An epidemiologic study of headache and its treatment in the general population of Catalonia]. AB - INTRODUCTION: Headache is one of the most frequent symptoms that people suffer in their daily life. However, the available studies have been made in populations who ask for medical advice or in small geographical areas. OBJECTIVES: The present survey was devoted to establish the prevalence of headache in the general population of Catalonia, as well as to establish its characteristics, the therapeutic behavior that was followed by those affected and the consequences for the sufferers. PATIENTS AND METHODS: The survey included a sample of 1,964 people, who were representative of the population older than 18 years, and was obtained from the census. The data were obtained by means of a telephone interview. The prevalence of pain in the last six months was established. Among the sufferers of headaches, additional information was obtained concerning its characteristics, its relationship with socio demographic characteristics and the therapeutic behaviors used by patients. The degree of impairment and disability secondary to headache was also determined. RESULTS: The prevalence was 42%, and it was the highest in women (52.8 vs 31.1%) young people (mean age of 43.4 16 years), unskilled manual workers, middle level managers and technicians and those with university studies. Headaches had a long duration (84.4%, more than 3 years); the episodes were frequent, of short duration and or severe intensity (61.2% were severe unbearable). Self medication was the therapeutic behavior most commonly used (52.3%, being acetylsalicylic acid the most employed), followed by visit to the physician (47.4%, being the prescription of paracetamol the preferred treatment) and some alternative medical treatments (16.1%). Headaches limited the daily activities of the sufferers (31.4%), forced to bed rest (23.4%) and even resulted in time off work (4.6%). All personal, social and work impairments had short duration in most individuals. CONCLUSIONS: Headache has a high prevalence in the general population. Although the episodes had a brief duration and self medication was commonly used, pain is frequently severe, limits the daily activities and sufferers often visit the physicians to obtain relief. Headaches should not be considered as a minor health disorder. PMID- 12134317 TI - [Myasthenia gravis in infancy. A report of 12 cases]. AB - INTRODUCTION: Myasthenia gravis (MG) shows specific clinical features in children. It is essential to know this and also the use of diagnostic techniques used in infancy and childhood for correct diagnosis. OBJECTIVES: To analyze the clinical behaviour of this disorder and the use of complementary tests in the diagnosis of paediatric patients. PATIENTS AND METHODS: We studied a group of 12 children diagnosed as having MG, who were admitted to the Instituto de Neurolog a y Neurocirug a de Ciudad de La Habana (Cuba) between March 1997 and June 2001. Data were obtained from the clinical histories regarding the clinical picture, anticholinesterase test, repetitive stimulation test (RST), simple fibre test (SFT), computerized axial tomography (CAT) of the mediastinum and the treatment given in each case. RESULTS: Juvenile myasthenia gravis (JMG) presented in 91% of the patients studied. The average age of onset of JMG was 7.45 years, with no difference in presentation in the two sexes. The extrinsic muscles of the eye were most affected and the form with generalized clinical involvement predominated at the time of admission. The RST was positive in four of the nine patients in whom it was done (44%) and the SFT was positive in the six cases in which it was done. No changes were found in the mediastinum on CAT scanning. Mestinon and prednisone were the most commonly used drugs. CONCLUSION: In our group JMG was the most frequent form seen. Neurophysiological studies were very useful diagnostic tools. PMID- 12134318 TI - [Usefulness of a screening questionnaire for post traumatic stress in a Colombian population]. AB - INTRODUCTION: Rating scales for post traumatic stress disorder (PTSD) should be consistents with DSM IV criteria, and should be validate for each culture. OBJECTIVE: To validate a PTSD checklist in a Colombian little town population, which was semi destructed by a guerrilla attack. PATIENTS AND METHODS: A stratified, representative and randomized sample of 202 adult participants, aged over 15 year old, was selected from San Joaquin (Santander Colombia) two year after an guerrilla attack. A structured interview (SCID I), based on DSM IV criteria, was developed with each member of the sample. 76 participants (37.6%) met criteria for PTSD, and 126 (62.4%) were classified as non PTSD. A rating checklist with 24 symptoms of PTSD was applied by self report. Each item of the scale was scored 1 to 4. RESULTS: PTSD checklist had a reliability Cronbach s alpha coefficient of 0.97. PTSD group scored 70.4 22.9, and non PTSD 37.2 13.7 (p< 0.0001) on the PTSD checklist. A discriminant analysis found that the scale had a correctly classification capability of 88.6% (p< 0.0001). Sensibility was found between 76.3% for a cut off point of 51 and 81.6% for cut off point of 45. Specificity changed between 71.4% for a cut off point of 45 and 84.4% for a cut off point of 51. CONCLUSION: Checklist for PTSD had a high reliability, good discriminant capability, and good sensibility and specificity. PMID- 12134319 TI - [Effects of simultaneous transplant of foetal mesencephalic cells in the striatum and the subthalamic nucleus of hemiparkinsonian rats]. AB - INTRODUCTION: The main strategy followed in neural transplants as a method of treatment for Parkinson s disease, both experimental and clinical, has been to introduce foetal mesencephalic cells into the target area: the striatum. However, when the dopaminergic cells in the substantia nigra degenerate, not only is the dopaminergic innervation of the striatum affected but also other nuclei: globus pallidus, substantia nigra, substantia nigra pars reticulata and subthalamic nucleus. A series of data from pharmacological and physiological studies offer strong evidence that the dopamine released in these nuclei may play an important role in regulating the output nuclei of the basal ganglia. AIM: To evaluate the effect of transplanting foetal mesencephalic cells on the behaviour of 6 OH DA rats when introduced into the striatum and the subthalamic nucleus. MATERIALS AND METHODS: 6 OH DA was used to induce lesions in the substantia nigra of rats, which were divided into several experimental groups. The rotating activity induced by D amphetamine (5 mg/kg, intraperitoneally) and apomorphine (0.05 mg/kg, subcutaneously) was evaluated before and three months after the transplant in all the experimental groups, except in the control group of healthy rats. The hemiparkinsonian rats received a total of 350,000 foetal ventral mesencephalic cells, which were implanted within small deposits in the striatum (8) and in the subthalamic nucleus (4). RESULTS AND CONCLUSIONS: Rotation induced by both drugs was significantly lower (p= 0.05) in animals that had had dopaminergic cells transplanted into the striatum body. No significant improvement in this behaviour was to be found when transplants were limited to just the subthalamus or, simultaneously, also to the striatum. A significant increase in rotating behaviour induced by apomorphine was observed in the group which received a transplant in just the subthalamus. PMID- 12134320 TI - [First tonic clonic generalized seizure: recurrence, and prognosis factors]. AB - OBJECTIVE: Previous epidemiologic studies have shown that around 5% of the population will suffer a tonic clonic seizure during their life. The aim of this study is to know how many and which of these people will suffer a second seizure and become epileptic. PATIENTS AND METHODS: 175 patients seen in the emergency department of the Vall d Hebron Hospital were included. They were divided in three groups according to the clinical suspicion of having had a seizure. Only the patients with low clinical suspicion and also normal EEG standard and EEG in sleep deprivation were excluded (16). The patients with previous episodes of lost of consciousness, previous episodes of possible mioclonias or absence were not excluded. RESULTS: After a first tonic clonic seizure the patients who did not receive treatment present a risk of relapse of 66% followed two years and the patients treated 46%. The difference between two groups was statistically significant. Dividing the patients according to the type of seizure: primary generalised, partial or nor localised we did not find differences in the risk of relapse. Dividing the patients according to their etiology we found that the group of patients with provoked seizures was different from the rest groups: symptomatic, genetic or cryptogenic and idiopathic, who had equal risk of recurrence. We found that the presence of previous episodes of lost of consciousness, the clinical suspicion and, probably (we obtained nearly statistical signification) de EEG and the presence of previous mioclonias or absences were risk factors. Other factor like age at the moment of the first episode, febrile seizures, familiar history, antecedents of stroke, encephalitis, neurosurgery and dementia were not related with the risk of relapse. CONCLUSIONS: With the exception of provoked seizures the rest of first tonic clonic seizures have a high risk of relapse (around 60 70%) and if they go with abnormal EEG, previous episodes of absences or mioclonias starting treatment must be considered. PMID- 12134322 TI - [Hereditary progressive levodopa sensible: Segawa's syndrome]. AB - INTRODUCTION: Hereditary progressive childhood dystonia with diurnal fluctuation of symptoms, belongs to the dopa responsive dystonias. It is dominantly inherited with variable penetrance, with deficiency in the cyclohydrolase I GTP gene. OBJECTIVE: Levodopa treatment is useful and diagnosis may be done on fluctuant dystonia in the childhood. CASE REPORTS: . We present four patients, one boy and three girls (two sisters) between 7 and 17 years of age. Neurological symptoms appears at 5, 15, 2.5 and 4 years of age respectively, with incoordination of movements ataxo paretic gait and postural dystonia. Symptoms were progressive with diurnal fluctuation. All laboratory test and image diagnosis were normal. Levodopa response, with lower doses (30 60 mg/day), were excellent. The four patients respectively are asymptomatic after 8, 4, 6 and 5 years of treatment. CONCLUSIONS: Hereditary progressive dystonia, with diurnal fluctuations is dopa responsive at lower doses, with neurological normalization and without side effects during a prolongated treatment. PMID- 12134321 TI - [Electroencephalographic discoveries in children with infantile massive spasms in Mexico]. AB - INTRODUCTION: Infantile massive spasms (IMS) is an age dependent epileptic syndrome that appears before the first year, and is characterised by delayed psychomotor development, massive spasms and hypsarrhythmia. There are classifications that only take the electroencephalogram (EEG) into account, without linking it with the IMS associated crisis pattern which can establish a treatment to improve the recurrence of the crises in this population, according to EEG discoveries and patterns of epileptic fits. PATIENTS AND METHODS: We include 100 EEGs of patients diagnosed with IMS, between 2 and 12 months old, selected by using the classification of Hrachovy to identify classic and modified hypsarrhythmia, and to correlate it with the pattern of epileptic seizures associated with IMS. RESULTS: The hypsarrhythmia found was mainly of the modified variety, and classic hypsarrhythmia only accounted for 9% of cases. The first case presented flexion IMS with generalised tonic seizures, and in the second, generalised tonic seizures and mixed IMS. Start age was between 2 and 4 months old. CONCLUSIONS: In comparison with results in other publications, in the EEG it was found that modified hypsarrhythmia with IMS associated partial and generalised tonic seizures with was the most common, while in the classic variety mixed IMS predominated. It must be taken into account that the absence of this variety does not rule out a diagnosis of IMS, but its presence makes the prognosis worse. PMID- 12134323 TI - [Low grade astroblastoma: pathological and magnetic resonance findings]. AB - INTRODUCTION: The astroblastoma is an uncommon type of glial tumour. It accounts for less than 1% of all tumours of the central nervous system. It originates in the tanicytes, ependymal cells present in the embryo and usually seen in adolescents and young adults. Radiologically it presents as a well delimited, heterogeneous tumour with a solid component which takes up contrast and is cystic, giving the same signal as cerebrospinal fluid (CSF). The pathological characteristics are of the formation of radial astroblastic pseudorosettes with perivascular hyalinization. CASE REPORT: An 18 year old woman presented with a 15 month history of motor deficit of her right limbs with occasional left frontal headache and horizontal diplopia on looking towards the left. On examination there was minimal claudication of the right limbs and bilateral papilloedema. Magnetic resonance showed a very well circumscribed left prefrontal neoplasm of heterogeneous aspect, with areas of solid and cystic appearance in the different sequences. The area of cystic appearance did not show the CSF signal in all sequences of the pulse. The tumour was totally excised. On histological study there were radial astroblastic pseudorosettes with perivascular hyalinization, with two mitoses per 10 fields of great magnification and the final diagnosis was of low grade astroblastoma. The apparently cystic portion was composed of friable gelatinous tissue. Fifteen months after her operation the patient is still asymptomatic. CONCLUSION: We report the radiological and pathological findings of a low grade astroblastoma. PMID- 12134324 TI - [Nocturnal paroxysmal dystonia, movement disorder and epilepsy]. AB - INTRODUCTION: Nocturnal paroxysmal dystonia (NPD) is a disorder which appears during sleep. It is characterized by generally brief paroxysmal motor events which are complex when clinically expressed and are often repetitive. The origin of this disorder has been a matter for discussion for some time. Initially it was considered a specific movement disorder, but recently it has been suggested that it is epileptic in nature. CASE REPORT: In this study we present the case of a patient who is hospitalized in the Unit for epileptic surgery, suffering from epilepsy which does not respond to medical treatment and requires surgery. The patient is treated with conventional methodology, prolonged and continuous presurgical videoelectroencephalographic monitoring. EEG registers were used via electrodes placed in the scalp and skull, electrodes placed in the skull on adequate indication. Two types of perfectly defined electroclinical events occurred: some not epileptic and others clearly epileptic. On one occasion a convulsive tonic clonic epileptic crisis was recorded typical of frontal focal origin preceded by non epileptic motor phenomena. CONCLUSION: The presence of paroxysmal motor episodes during sleep, atypical as a form of epilepsy, the absence of unquestionable specific data in the EEG, and in the light of our discoveries, force us to consider the possibility that NPD is in fact a form of epilepsy caused in a reflex manner by a specific type of movement disorder during sleep, and whose origin should be more widely discussed. PMID- 12134325 TI - [Lumbar plexopathy secondary to pelvic hydatid cyst]. AB - INTRODUCTION: Hydatidosis is a zoonosis caused by larvae of the hydatid tapeworm Echinococcus granulosus, which usually affects the liver, lung, myocardium, brain and bones. On rare occasions hydatid cysts give rise to peripheral neurological manifestations. CASE REPORT: Male, aged 72, who had received a pacemaker as a consequence of a complete auriculoventricular block. During implantation, one hepatic and two pelvic hydatid cysts were discovered. Consulted doctor about right inguinal pain, which irradiated to the lumbar region and along the inner and posterior sides of the thigh, down to the ankle. Exploration revealed a lump in the right iliac fossa and in the lower right limb, hypaesthesia in the anterior side of the thigh, paresis on bending the hip and on extension of the knee, and patellar areflexia. Computerised axial tomography revealed growth of the pelvic cysts and compression of the gluteal veins. Electromyographic exploration of the right quadriceps showed denervation and reinnervation activity. CONCLUSION: Of all cases of abdominal hydatidosis, 6.7% presented extra hepatic affectation and only 0.7% were seen to have pelvic hydatid cysts. These usually give rise to disorders due to local compression of the genital organs, the urinary tracts, and vascular and bony structures. Lesions in the lumbosacral plexus, however, have only been reported on very few occasions PMID- 12134326 TI - [Smith Lemli Opitz Syndrome type II of neonatal diagnosis and review of the most interesting clinical features]. AB - INTRODUCTION: Smith Lemli Opitz syndrome is an autosomal recessive metabolic disease, two forms can be differentiated: type I and type II. CASE REPORT: We present the clinical case of a female newborn with antecedents of oligoamnios and intrauterine growth retardation who presented a characteristic malformative syndrome, severe neurological impairment, anomalies of the limbs, pyloric stenosis, and renal and cardiac defects. Determination of cholesterol and its precursors by gas chromatography confirmed the clinical diagnosis of a severe form with exitus at six months of age. At the same time a review of the syndrome is presented. PMID- 12134327 TI - [Hypnic headache: a report of four cases]. AB - INTRODUCTION: Hypnic headaches are an uncommon type of primary headache of short duration, related to sleep, which have usually been described in elderly persons. Although not currently included in the International Headache Society's classification, criteria have been established for its diagnosis. They include: frequency of more than 15 days per month for at least one month, the headache wakens the patient when asleep, it lasts for between 5 and 60 minutes, may be generalized or unilateral, with no associated autonomic disorder or causative aetiological agent. CASE REPORTS: We describe four clinical cases, three women aged 43, 52 and 65 years old and one 68 year old man. They complained of moderate to severe headaches which were unilateral, daily, woke them up at night, lasted for some hours and had no associated autonomic symptoms. On clinical investigations (cephalic CAT and MR scans) organic disease, as a cause, was ruled out. Different acute and prophylactic treatments were tried, but only lithium carbonate (400 mg at night) led to full improvement of symptoms. When this treatment was stopped the headache reappeared in some patients. CONCLUSIONS: Although some patients did not fulfil the diagnostic criteria described by Dodick et al in a previous series of cases, the night time presentation with no associated autonomic disorder and good response to lithium carbonate were in favour of the diagnosis of hypnic headaches PMID- 12134328 TI - [Possible gabapentin phenytoin interaction]. AB - INTRODUCTION: We report a possible case of gabapentin induced phenytoin toxicity. CASE REPORT: White man, 26 years old, chronically treated with phenytoin (for the past 4 years) and gabapentin (for the past 17 months). He was seen after complaining of dysarthria, ataxia and vertigo for the past 3 months, although having noted slight dizziness and a general, though undefined, indisposition from the start of taking gabapentin. The total and free serum levels of phenytoin found were clearly toxic. Gabapentin was discontinued definitively, and phenytoin for the next 7 days. The clinical symptoms had disappeared completely and phenytoin returned to within therapeutic levels. According to the criteria of causation it can be considered a possible adverse reaction caused by phenytoin related to incorporation of gabapentin. The mechanisms of this possible drug interaction are discussed, with emphasis on cytochrome P450 metabolism. CONCLUSION: The present case is a warning of the possible interaction of a drug (gabapentin) not contemplated when starting or when monitoring an antiepileptic treatment. PMID- 12134329 TI - [Subarachnoid hemorrhage]. AB - INTRODUCTION: The vascular structures in the central nervous system has a particular distribution in respect of other organs. This anatomic configuration and other conditions predispose this arteries and veins to bleed frequently in the subarachnoid space, resulting in a syndrome known as subarachnoid hemorrhage. DEVELOPMENT: We make a study of the bibliography published in the topic, beside our professional criteria concerning those themes, with regard to the elements that we have been able to evaluate in our daily practice. We carried out a wide review about etiology, physiopathology, clinical presentation, diagnosis and management of the commonest problems associated with that entity. Specially those aspects related with the clinical support and treatment of the cerebral vasoespasm, hydrocephalus and other complications of the subarachnoid hemorrhage. CONCLUSION: In this paper we expose the most important aspects related with the clinical manifestations of certain vascular malformations like saccular aneurysm and arteriovenous malformations, the principles of it diagnosis and management. PMID- 12134331 TI - [Epileptic auras: classification, pathophysiology, practical usefulness, differential diagnosis and controversials]. AB - OBJECTIVE: To examine those aspects of the epileptic auras that may contribute to improve our understanding of this epileptic manifestation. DEVELOPMENT: Epileptic aura is that portion of the seizure which occurs before consciousness is loss and for which memory is retained afterwards. In the case of simple partial seizures, the aura is the entire seizure. This epileptic phenomenon is the consequence of the activation of functional cortex by abnormal, unilateral, circumscribed and brief neuronal discharge. Surface electroencephalogram frequently fails to detect any changes during an isolated aura. The incidence of auras in partial epilepsy remains imprecise and there are marked discrepancies among authors. Viscerosensorial and experiential auras are most often seen in temporal lobe epilepsy. Feeling of fear is the commonest affective symptom associated with epileptic discharges from mesial temporal origin. Special sensory auras including visual, gustatory and vertiginous are more frequently described in extratemporal epilepsy. Olfactory auras are rare, however, when they occur the most likely etiology is a tumor involving the amygdala and hippocampus. Somatosensory auras include sensations of tingling or numbness, electrical feeling or very occasionally, pain. There is not agreement with the classification of determined auras such as motor, cephalic and sexual. The differential diagnosis is enormously heterogeneous including vestibular disfunction, migraine, stroke, autonomic disturbances, drug intoxications and psychiatric disorders. CONCLUSION: The clinical manifestations of the epileptic auras are extraordinarily diverses and, therefore, the knowledge of their pathophysiology, characteristics, incidence and association with differents types of focal epilepsy are the clue to obtain a precise diagnosis. PMID- 12134330 TI - [Guillain Barre syndrome]. AB - OBJECTIVE: To review about this disorder, with emphasis on the intensive care of severe Guillain Barr syndrome (GBS). DEVELOPMENT: GBS is an acute immune mediated inflammatory polyneuropathy that may lead to quadriparesis, ventilatory failure, and autonomic dysfunction but also to many general medical problems that have great bearing on outcome. Therefore severe GBS patients require admission into an intensive care unit (ICU), where in addition to the disorders mentioned before, other complications can arise. The neurologist who plans to deal comprehensively with these patients must be familiar with therapy for infections, nutrition, fluid management, and selected aspects of pulmonary medicine as well as the indications for and complications of plasma exchange and gammaglobulin infusion. CONCLUSIONS: With modern intensive care support, the outcome is excellent (>80% recovery), although in many cases a persistent residual paresis occurs. Because GBS is largely self limited, the skill daily cares of these patients in an ICU contributes as much, or more, to the overall outcome of an individual patient as do specific immune therapies. PMID- 12134332 TI - [The effect of experience on post natal development of the vascularization of the visual cortex]. AB - OBJECTIVE: To report the current state of understanding of the basis of post natal development of microvascularization and the influence of the environment. DEVELOPMENT: Postnatal development of sensory systems occurs in two stages: one genetically predetermined and the other modulated by experience, which is especially important during a so called critical period. Increased demand during this period leads to adaptive changes, particularly vascular changes. The cerebral cortical vascularization is mainly composed of a dense capillary network which extends over the entire cortex and whose density corresponds to the metabolic demand. The development of cortical vascularization starts with perpendicular trunks which branch according to demand and form a cortical capillary network, mediated by factors such as endothelial vascular growth factor (VEGF). Many investigations into the effect of external experience have been done on the visual cortex, both for increase and deprivation. CONCLUSIONS: Post natal development with enriched visual environments induces increased vascular density. Absence of visual experience leads to a reduction in cortical activity which induces reduced neurone and vascular density, together with delay in maduration of the cortical angio architectural pattern. The relation between the neurone population and superficial vascular density is unchanged. These changes appear after the critical period of maximal synatic reorganization. PMID- 12134334 TI - [Epidemiological study of oligodendrogliomas in Aragon and La Rioja]. PMID- 12134335 TI - [Value of social support networks in the evolution of hemiplegic patients]. PMID- 12134337 TI - [Rolandic discharges in childhood epilepsy: magnetoencephalographic diagnosis]. PMID- 12134336 TI - [Moebius-Poland syndrome]. PMID- 12134338 TI - [Cognitive evoked potential in primary wide angle glaucoma]. AB - INTRODUCTION: Primary wide angle glaucoma (PWAG) is the commonest form of this disorder. It is initially asymptomatic, until there has been considerable visual field loss. Personality changes have been described in patients with wide angle glaucoma, with a significant percentage of developing dementia or a tendency to react defensively. The P300 endogenous evoked potential is related to memory/attention to work. INTRODUCTION: To evaluate, in PWAG, the probable occurrence of a disorder of visual processing due to alteration in the attention/working memory and its neuronal network, using the P300 visual cognitive evoked potential in correlation with Lobo s cognitive mini exam (CME). PATIENTS AND METHODS: We selected 68 patients with PWAG and 68 controls, of both sexes, aged between 50 and 65 years. The CME was done after taking the clinical history, and then an electroretinogram, visual evoked potential using the alternating dameres (ERGd and VEPd) and P300 potential (passive odd ball). Statistical analysis was done using the t test, one way ANOVA and discriminating analysis. RESULTS: The amplitude on ERGd and latency on VEPd were significantly altered. Patients with glaucoma had lower scores on CME. Regarding the P300 wave, latency was the most significant parameter, whilst the amplitude and site were less well defined. PATIENTS AND METHODS: In PWAG a process of visual cognitive dysfunction occurs and affects memory/attention to work in a way which is not only due to the peripheral visual effect. PMID- 12134339 TI - [Differences in the spectroscopy of the lesions of the remitting relapsing form of multiple sclerosis shown by magnetic resonance]. AB - INTRODUCTION: Multiple sclerosis (MS) is a chronic demyelinating disorder of the central nervous system, characterized by the presence of inflammatory lesions. OBJECTIVE: To analyze the biochemical profile of the demyelinating lesions of the initial forms of MS (remitting relapsing) by analyzing the proton magnetic resonance spectra (1H MRS) to characterize the process of demyelination and relate it to the metabolites and clinical variables analyzed. PATIENTS AND METHODS: We analyzed the largest demyelinating lesions in eight patients with remitting relapsing MS (RRMS) using the technique of single volume 1H MRS (VOI) with short echo time. The spectra of the white matter of two healthy control were used as reference. RESULTS: NAA/Cr and NAA/Cho value ratios decrease and mI/Cr one increase in all spectra lesions as compared to healthy controls. In four of the eight patients, the Cho/Cr was higher than in the controls. Qualitative and quantitative differences in the resonances of macromolecules were observed, related to the biochemistry of the process of demyelination. These differences in NAA/Cr, Cho/Cr, mI/Cr and macromolecules probably represent different stages in the evolution of the plaques. CONCLUSIONS: MRS is a non invasive technique able to observe biochemical variations related to the evolution process of demyelination. Activity of the lesion is shown by the increment of resonances around 0.9 1.3 ppm. An increase in mI seems to occur at an early stage of demyelination and later the NAA is reduced. The initial forms of MS show metabolic alterations in the plaques which are similar to the most advanced forms of MS. PMID- 12134340 TI - [The epidemiology of multiples sclerosis in Alcoi. Analytical data]. AB - INTRODUCTION: The epidemiology of multiple sclerosis (ME) in the Alcoi area has been studied by our groups for many years. When studying the factors linked to ME in this area initially we observed that migration, contact with dogs and the textile industry were associated with the disorder. A later study, in which the migration factor was controlled, still showed a significant association with contact with dogs and the textile industry. OBJECTIVE: To make an analytical study avoiding the possible confusing effect of contact with the textile industry, since this may be associated with other factors. We also compared the results of previous studies. PATIENTS AND METHODS: We made a case control study in which each case was paired with four controls for age, sex and having worked or not worked in the textile industry. The cases fulfilled the criteria for diagnosis of ME. The control patients came from the Emergency and Trauma Departments of Alcoi hospital. RESULTS. We analyzed 37 patients and 148 controls. Significant values were obtained for smoking, low social class, chickenpox and measles infections when aged under 15 years, pneumonia when aged over 15 years and contact with dogs, sheep and dissolvents. CONCLUSIONS: Contact with dogs was associated with multiple sclerosis in all the studies carried out in the Alcoi area to date. This is therefore a risk factor for the disorder in the Alcoi population. PMID- 12134341 TI - [Intravenous treatment with immunoglobulins in epileptic syndromes which are difficult to control]. AB - INTRODUCTION: Several mechanisms have been found to be involved in the development of malignant epileptic syndromes in childhood and an immunological component has been shown in experimental studies of epilepsy. The effect of intravenous immunoglobulin (IgIV) has been reported in various cases of epilepsy which were difficult to control. OBJECTIVE: To show the usefulness of IgIV in West s syndrome (WS) and Lennox Gastaut syndrome (LGS) which were difficult to control. PATIENTS AND METHODS: We selected five patients who fulfilled the criteria for WS and LGS which were difficult to control. They were given IgIV for five days and subsequently six fortnightly doses of 0.5 g/kg/day. We recorded data on sex, age, onset of the disorder and laboratory investigations, including blood and cerebrospinal fluid (CSF) immunoglobulin levels. RESULTS: Four girls with symptomatic WS and one boy with LGS as a symptom of a neuro infection. Treatment was started at the age of 8 14 months. The spasms varied between 204 and 1,074 in 24 hours, and were markedly reduced after IgIV was given. The IgG in the CSF rose as the number of spasms fell (p< 0.05). CONCLUSIONS: There was a satisfactory response to treatment, as reported in previous publications. It is a coadjuvant treatment for cases which are difficult to control and may also be useful in cases of symptomatic epilepsy where goods results are also obtained. PMID- 12134342 TI - [Neurophysiological diagnosis of lumbosacral radicular compression syndrome from late responses]. AB - INTRODUCTION: Lumbosacral radicular compression syndrome is a disorder that can affect anyone at some moment in their lives, regardless of their sex, age and profession. It appears more frequently in specific groups of people. Late responses constitute neurophysiological studies that allow evaluation of the functional state of the proximal portions of the peripheral nervous system, which are affected to a greater or lesser extent in the course of this pathological state. AIMS. To determine the usefulness of the F wave and H reflex in lumbosacral compressive radiculopathies at L5 and S1. PATIENTS AND METHODS: We examined 100 patients who had been clinically and imagenologically diagnosed as suffering from lumbosacral compressive radiculopathy, which was classified as being L5 and S1. Late responses (F wave and H reflex) were performed and they were compared with a group made up of 74 normal subjects. RESULTS: In patients with L5 radiculopathy, the F wave was abnormal in 66.6% of the cases, where a reduction in the percentage of waves obtained from patients and prolonged minimum, average and maximum latencies predominated. In the case of patients with S1 lumbosacral compressive radiculopathies, 77.5% presented H reflex alterations and the main abnormalities were detected as being the prolongation of reduced amplitude latencies and the absence of response. CONCLUSIONS: The F wave and H reflex are useful tools for evaluating patients with compressive radiculopathies at L5 and S1, respectively. PMID- 12134343 TI - [Long term remission of epilepsy in childhood: a prospective study]. AB - INTRODUCTION: To study the chance of achieving a long term remission of epilepsy in children. PATIENTS AND METHODS: 90 children aged under 14 years newly diagnosed of epilepsy were started on anticonvulsant treatment and prospectively followed. Initial remission was defined as a seizure free period with or without subsequent recurrences, terminal remission as a seizure free period without subsequent recurrences and terminal remission off treatment as terminal remission plus medication discontinued. RESULTS: The Kaplan Meier estimate of the probability of achieving a 3 year initial remission was 80% and 90%, of the probability of achieving a 5 year terminal remission was 59% and 68% and of the probability of achieving a 5 year terminal remission off treatment was 53% and 61% at 5 and 7 years, respectively. Univariate and multivariate analyses using the Cox proportionate hazards model revealed there was a greater probability of achieving a 3 year terminal remission off treatment for patients with idiopathic/cryptogenic etiology and for those who did not have recurrences during the first six months of treatment. Syndromic diagnosis at six months after initiating antiepileptic treatment was also useful for predicting the probability of remission. CONCLUSIONS: Over half of the epileptic children achieve a prolonged remission off treatment. The main prognostic factors are etiology, recurrence of seizures during the first six months and syndromic diagnosis. PMID- 12134345 TI - [Severe neonatal encephalopathy, secondary to a prolonged vasovagal episode in a woman 31 weeks pregnant]. AB - INTRODUCTION: Vasovagal syncope is not frequent during the second half of pregnancy, although the supine hypotension syndrome is common during this period. During these episodes, the marked hypotension may impede uteroplacental blood flow. CASE REPORT: A woman who was 31.5 weeks pregnant lost consciousness for 15 minutes on standing up. 24 hours later, when it was observed that foetal movement had diminished, a cardiotocographic recording was done. This showed a pathological pattern and foetal biophysical profile on echography showed a marked foetal hypotonia and absence of movement. Emergency caesarean section was carried out and a male foetus delivered, weighing 1.810 g, with an Apgar score of 5/6 and requiring intubation and mechanical ventilation for respiratory difficulties and generalized hyptonia with absent reflexes. During the following days he developed generalized hypertonia and died aged 18 days. At necropsy there was severe brain damage, of ischaemic type. Complementary tests on the mother were normal. CONCLUSIONS: Severe prenatal encephalopathy secondary to maternal vasovagal syncope is uncommon, and we have found no report of it in the literature. The prolonged duration of the hypotensive episode, together with prematurity, which implies poor regulation of cerebral blood flow, may have contributed to the severe damage to the central nervous system PMID- 12134344 TI - [Rombencephalitis due to Listeria monocytogenes. Probable usefulness of dexamethasone associated with antibiotic treatment]. AB - INTRODUCTION: Infection of the central nervous system by Listeria monocytogenes appears in most cases as acute meningitis which is indistinguishable from other types of acute meningitis. Rombencephalitis is a rare form of neurolisteriosis, localized to the brainstem. The initial non specific symptoms may make early diagnosis difficult. CASE REPORT: We describe the clinical case of a previously healthy woman who had L. monocytogenes infection localized to the brainstem. Her initial symptoms were fever and headache followed by signs of brainstem involvement, deterioration of consciousness and severe respiratory insufficiency which made mechanical ventilation necessary. Study of the cerebrospinal fluid showed lymphocytic pleocytosis, raised protein and normal glucose levels. L. monocytogenes was isolated on blood culture. Cranial computerized tomography was normal and magnetic resonance showed a right pontobulbar lesion. After receiving specific antibiotic treatment the infectious condition improved. However, the neurological symptoms started to improve when dexamethasone was added to the antibiotic treatment twelve days later. The patient was discharged from hospital with slight neurological sequelas. CONCLUSION: In a febrile patient with signs of brainstem involvement, neurolisteriosis should be suspected and ampicillin and gentamycin added to the treatment. The possibility of acute respiratory arrest occurring makes it necessary to monitor these patients closely. The association of dexamethasone to the antibiotic treatment may be useful in some cases of rombencephalitis due to L. monocytogenes. PMID- 12134346 TI - [Atypical Guillain-Barre syndrome: multiple cranial neuropathy]. AB - INTRODUCTION: Multiple cranial neuropathy is a condition rarely seen in everyday clinical practice. It has many different causes, and in spite of careful clinical investigation many cases remain of unknown aetiology. It is also considered to be an atypical variant, topographically circumscribed, of the Guillan Barr syndrome (GBS). CASE REPORT: A 23 years old man developed a progressive illness over ten days. He complained of diplopia, facial diplegia and a nasal voice. Subsequently, he also developed weakness of the neck and tongue muscles, dysphagia, abolition of reflexes of the left arm and right triceps reflex but without involvement of the respiratory muscles or other limbs. CSF studies showed slightly raised protein with no cells. Neurophysiological studies showed a demyelinating disorder with secondary axonal damage. In spite of further studies, no aetiological agent was found. DISCUSSION AND CONCLUSION: These observations suggested this case is of a topographical variant of GBS. Such cases have also been classified as the Miller Fisher syndrome, pharyngo cervico brachial paralysis, are flexic paraparesia and bilateral lumbar polyradiculopathy. In view of the diversity of the clinical and biological characteristics of the cases reviewed, which may mean different aetiopathogeneses, we consider that a thorough search should be made for the aetiology before these conditions are labelled as atypical variants of GBS. PMID- 12134347 TI - [Perilymphatic fistula: report of a case which resolved spontaneously in five days and literature review]. AB - AIMS: We report a patient with a perilymphatic fistula and review the literature on this topic. CASE REPORT: The patient was a 50 years old male with a five days history of intense and continuous vertigo associated with nausea, vomiting and sweating, which was initiated after a sudden noise in the left ear. The symptoms resolved spontaneously over five days, without surgical treatment. The clinical picture resembles those previously reported for perilymphatic fistula. The current literature on this topic has been reviewed in this article. CONCLUSION: The patient presented an early recovery, what corroborates the current tendency of waiting a few weeks before indicating surgery. PMID- 12134348 TI - [Streptococcus bovis meningitis. An infrequent cause of bacterial meningitis in the adult patient]. AB - INTRODUCTION: Bacterial meningitis in adult patients, produced by streptococci other than Streptococcus pneumoniae, is not common. CASE REPORT: We report the case of a 74 year old male patient with meningitis and endocarditis due to Streptococcus bovis (group D, not enterococcus), sensitive to penicillin (CMI< 0.1 mg/L), with no characteristic clinical or analytical discoveries. A gastrointestinal exploration revealed only diverticles in the colon and two lesions compatible with splenic infarction, observed by using computerised axial tomography of the abdomen. The patient responded favourably to a four week course of antibiotics; he remained asymptomatic, afebrile and culture negative after the therapy was stopped. CONCLUSIONS: In many previously reported cases, there is an association with gastrointestinal illness, endocarditis or oral lesions. Gram staining of the cerebrospinal fluid is usually negative and the neurological signs are often subtle. In the case of bacteraemia, endocarditis or S. bovis meningitis, the presence of an underlying pathology of the colon due to the frequent association between these processes must be ruled out. Treatment with penicillin G is usually sufficient. PMID- 12134349 TI - [Biophysical foundations of magnetoencephalograhy]. AB - OBJECTIVE: It is sought to expose in a simple but rigorous way the physical, neurobiological and methodological foundations of the magnetoencephalography (MEG). DEVELOPMENT: We start from the basic properties of the classical electromagnetism, analyzing in detail the concepts of electric and magnetic fields, the Maxwell s equations and the multipolar development of potentials. All these tools are very important to know the peculiarities of the MEG studies. Later on, they are reviewed very briefly the different types of potentials generated by the neurons and their implication in the MEG. Lastly, some necessary technical characteristics will be commented for detection of the very weak neuromagnetic fields. It is shortly exposed the concept of tunnel effect, in one that detection systems used at the present time are based (SQUID). CONCLUSION: MEG is a very promising recent technique that is used in epilepsy studies, evoked potentials and other functional pathologies. Its utility in clinic continues being even controversial. However, it is fundamental to know the mechanisms that justify their use in order to know better their benefits and limitations. PMID- 12134350 TI - [Effects of antiepileptics drugs on cognition]. AB - The cognitive effects of the most commonly used antiepileptic drugs (AEDs) are reviewed, discussing both the absolute effects (the effects compared to no treatment) and the relative effects (the comparison of one AED with an other). The cognitive profile per AED is given, based on a literature database, consisting of 1,357 articles in peer review journals. Available information on the newer AEDs is discussed as an addendum. PMID- 12134351 TI - [Psychological aspects involved in epilepsy]. AB - INTRODUCTION: In spite of the importance of psychological perspective that influence directly or indirectly in the treatments of epilepsy, the psychology plays in the physical assistance reality a secondary place. OBJECTIVE: To review the relevant psychological aspects about affective aspects in epilepsy. DEVELOPMENT: The revision of frontal and limbic lobes, hemispheric lateralization, processual and stationary epilepsy, forced normalization, interictal psychopathological disturbances, effects of antiepileptic drugs, quality of life and another aspects related living the illness. CONCLUSIONS: Clinical psychology and neuropsychology contribute with relevant information about diagnostics, treatments, to know deeply the illness and the advise of the health professionals. PMID- 12134352 TI - [Factors affecting the cognitive status of people with epilepsy]. AB - INTRODUCTION: Epilepsy is a devastating neurological disorder affecting the neuropsychological functioning of the sufferers. AIM AND DEVELOPMENT: This paper reviews the crucial factors associated with the cognitive status of people with epilepsy. These factors include seizure type and other medical variables such as age of onset, seizure frequency, and seizure history. The effect of these factors on the cognitive functioning of people with epilepsy is reviewed and discussed. Future directions of studies on this topic are also discussed. PMID- 12134353 TI - [Intracarotid amytal testing in the evaluation for epilepsy surgery]. AB - INTRODUCTION: The Wada test for language and memory still plays an important role in evaluating the surgery performed on epileptic patients. The measures of the functional deficits carried out with the Wada test, associated with known brain lesions, aid in determining the lateralization of the onset of seizures and provide some estimate of the risk to memory after a temporal lobotomy. DEVELOPMENT: A more refined procedure, based on the correlations with the titration from MRI, fMR and MR spectroscopy, will be needed. Moreover, using the Wada test it is possible to make long term forecasts about the cognitive outcome and about the control of seizures. When used together with other neurological, functional and psychometric evaluations, it becomes easier to select the patients with most chances of benefiting from surgical epilepsy treatment. Finally, the non invasive measures of brain functioning include fMR, which provides far more information than that obtained using the Wada test. However, it remains to be determined whether the procedure based on activation provides data of the same quality as those obtained by the Wada test, which can provide a more adequate reversible model of the effects of surgery on cognition. CONCLUSION: . Ideally, as MRI and other imaging techniques progress, they will be able to offer complementary images, which will improve our capacity to forecast and prevent important post operative cognitive deficiencies. PMID- 12134354 TI - [Insights into brain function though magnetic source imaging: A review of research and clinical applications]. AB - INTRODUCTION AND DEVELOPMENT: The paper presents a brief outline of the rationale behind the use of non invasive functional imaging and of the features that any imaging technique should display in order to make a substantial contribution to the search of the brain mechanisms responsible for cognitive functions. One such technique, magnetic source imaging (MSI), that meets these specifications, is described in more detail. Advantages of MSI include the capacity to provide direct measures of regional neurophysiological activity, a millisecond range temporal resolution, and the capacity to provide images of brain activity on an individual basis. We then describe applications of MSI to the study of brain mechanisms involved in various language functions such as oral comprehension and reading. Among these applications, the accuracy of MSI protocols in determining hemispheric dominance for language functions and in identifying the precise location and extent language specific cortex (Wernicke s area) has been verified through comparison with standard invasive techniques (Wada procedure and electrocortical stimulation mapping) in over 60 consecutive cases. In another series of studies we combined data from MSI and direct cortical stimulation to determine the role of temporoparietal areas in phonological analysis of spoken language and in phonological decoding of print. Finally, we have used MSI to gain unique insights into the brain mechanisms that support reading in developmental reading disability. CONCLUSION: Results from over 21 children diagnosed with this disorder suggest that impaired reading is associated with aberrant functional connections between temporal and temporoparietal areas of the left hemisphere that are normally engaged in reading. PMID- 12134355 TI - [Neuroscience in Al Andalus and its influence on medieval scholastic medicine]. AB - INTRODUCTION: Since the application of technical medicine by the Greeks, modern neurology has been based on a body of knowledge and cultural heritage from ancient times. In this paper we review the contribution made by Al Andalus to neuroscience during the Middle Ages and its repercussions on modern neurology. DEVELOPMENT: Following the death of Mohammed in the vii century AD, Islam enjoyed one of the most spectacular periods of expansion in the history of mankind. Occupation of the cities of Alexandria and Gundishapur put the Arabs into contact with original Greco Latin manuscripts, which were assimilated and divulged by Islamic scientists in the middle eastern caliphates of Damascus and Bagdad as well as the western caliphates of Al Andalus (Spain) and Kairwan (Tunis). This classical hippocratico galenico medicine was refashioned into the so called arabized galenism, which markedly influenced the Scholastics and the cultured world of the lower Middle Ages and became the basis of European medicine until well into the Renaissance period. There was a first Spanish cultural Renaissance in Al Andalus during the ix xii centuries, which led to a flowering unheard of in the Middle Ages before then. Andalusian doctors made major contributions to the body of knowledge about neuroscience and developed major philosophical concepts of human understanding. Thus, Abulcasis (936 1013), the father of modern surgery, developed material and technical designs which are still used in neurosurgery. Averroes suggested the existence of Parkinson s syndrome and attributed photoreceptor properties to the retina. Avenzoar described meningitis, intracranial thrombophlebitis, mediastinal tumours and made contributions to modern neuropharmacology. Maimonides wrote about neuropsychiatric disorders and described rabies and belladonna intoxication. CONCLUSION: Aside from the political, religious and cultural differences between Al Andalus and the Christian kingdoms of the Iberian peninsula, the historical Andalusian period (711 1492) forms one of the most brilliant periods of Spanish neuroscience. PMID- 12134356 TI - [Epidemiological study of cerebral medulloblastoma in Aragon and La Rioja]. PMID- 12134357 TI - [Epidemiological study of mixed gliomas in Aragon and La Rioja]. PMID- 12134358 TI - [Epidemiological study of acoustic neurinomas in Aragon and La Rioja]. PMID- 12134359 TI - [Epidemiological study of cerebral metastases in Aragon and La Rioja]. PMID- 12134360 TI - [Epidemiological study of pinealoma in Aragon and La Rioja]. PMID- 12134361 TI - [Bilateral hypodensity of the basal ganglia. Clinico-evolutionary correlation in children]. PMID- 12134363 TI - Clinical oncologic positron emission tomography: an introduction. AB - PET imaging is a molecular imaging technology that is diffusing into imaging departments quite rapidly. The unique characteristics of positron emitting radionuclides such as fluorine-18 provide high-quality images with reasonable acquisition times. The imaging instrumentation continues to improve with new detector materials and combinations of PET scanners and CT scanners. FDG is now readily available to most hospitals in the United States. Third-party payers now recognize the importance of PET imaging in multiple malignancies. The number of PET scans performed annually will continue to increase as the indications increase and the instrumentation is more available. PMID- 12134364 TI - Positron emission tomography: lung cancer. PMID- 12134366 TI - Positron emission tomography imaging in melanoma and lymphoma. PMID- 12134365 TI - Positron emission tomography for evaluation of colorectal carcinoma. AB - Evaluation of patients with known or suspected recurrent colorectal carcinoma is now an accepted indication for FDG PET imaging. FDG PET does not replace imaging modalities such as CT for preoperative anatomic evaluation but is indicated as the initial test for diagnosis and staging of recurrence and for preoperative staging (N and M) of known recurrence that is considered to be resectable. FDG PET imaging is valuable for differentiation of posttreatment changes from recurrent tumor, differentiation of benign from malignant lesions (indeterminate lymph nodes, hepatic and pulmonary lesions), and evaluation of patients with rising tumor markers in the absence of a known source. Addition of FDG PET to the evaluation of these patients reduces overall treatment costs by accurately identifying patients who will and will not benefit from surgical procedures. Although initial staging at the time of diagnosis is often performed during colectomy, FDG PET imaging is recommended for a subgroup of patients at high risk (with elevated CEA levels) and normal CT and for whom surgery can be avoided if FDG PET shows metastases. Screening for recurrence in patients at high risk has also been advocated. FDG PET imaging seems promising for monitoring therapy, but larger studies are necessary. PMID- 12134367 TI - Esophageal cancer and head and neck cancer. PMID- 12134368 TI - Evaluation of breast and gynecologic cancers by positron emission tomography. PMID- 12134369 TI - Positron emission tomography in thyroid cancer management. AB - Selected patients with thyroid cancer can benefit from the use of PET imaging with FDG or with I-124. The PET scan impacts on management by providing (1) more accurate information about staging of patients in terms of extent of tumor for better treatment planning, especially in patients who do not concentration radioactive I-131; (2) the relationship of tumor involvement to vital structures, especially in the neck and central nervous system; and (3) prognostic information (an SUV > 10 and extensive PET + disease connotes a poor prognosis in advanced patients). In the occasional patient, surgically respectable disease has been identified on PET with the result that the patient has been rendered no evident disease with treatment. PET has also been used in the follow-up of patients who have been treated for thyroid cancer, to assess response. PET may also be useful for lesion specific dosimetry, with I-124. The combination of PET and CT in the same gantry facilitates localization of thyroid cancer PET scan abnormalities in relationship to critical organs and structures. PMID- 12134370 TI - Radiology and "pet" names. PMID- 12134371 TI - PET--elegant research tool now elegant clinical tool. PMID- 12134372 TI - Case of the season. Hibernoma. PMID- 12134373 TI - Outcomes research: an interdisciplinary perspective. AB - Outcomes achieved by healthcare providers are a function of patient clinical and demographic characteristics, baseline health status, delivered treatments and interventions, and setting characteristics. This article describes a prospective, interdisciplinary study of the outcomes of selected surgeries using both generic and condition-specific data collection instruments. The results of several different analyses are discussed, which illustrate the usefulness of such an effort for patients, nurses and physicians, the institution, and the respective disciplines in general. PMID- 12134374 TI - Nursing outcomes classification: a preliminary report of field testing. AB - Clinically useful and measurable patient outcomes that are sensitive to nursing interventions are needed to determine the effectiveness of nursing care. Nursing Outcomes Classification (NOC) researchers are evaluating the reliability, validity, and usefulness of the first 190 published outcomes in 10 sites representing the continuum of care. Preliminary analysis of inter-rater reliability and construct or criterion validity of 15 outcomes are described. Results indicate NOC outcomes can be used to accurately document the effectiveness of nursing interventions. PMID- 12134375 TI - Using an outcomes management program in a public health department. AB - This article and its predecessor offer suggestions to nurses in all service settings by describing how a local public health department planned and implemented its outcomes management program. Included are the steps used to select and analyze reliable and valid quantitative outcomes data, and examples of graphs that depict and interpret those data. The outcomes management program was designed to provide needed information for program planning and evaluation, and for communication with administrators and local government officials. PMID- 12134376 TI - Patient satisfaction measurement: current issues and implications. AB - Healthcare consumers are demanding excellence in care and services from care providers, and payors are following in their expectations. Demonstration of quality outcomes and consumer satisfaction with services are now a priority and the primary competitive edge in healthcare. Hospitals and healthcare systems that invest in programs to determine how patients evaluate their experiences will have valuable information to make transformational changes in care delivery and services. The purpose of this article is to present an overview of consumer/patient satisfaction instruments, satisfaction measurement issues, research instruments, and commonly used vendor patient satisfaction survey programs. PMID- 12134377 TI - Guided imagery in cardiac surgery. AB - Clinical research has demonstrated that guided imagery, a simple form of relaxation, can reduce preoperative anxiety and postoperative pain among patients undergoing surgical procedures. In 1998, the cardiac surgery team implemented a guided imagery program to compare cardiac surgical outcomes between two groups of patients: with and without guided imagery. Data from the hospital financial cost/accounting database and patient satisfaction data were collected and matched to the two groups of patients. A questionnaire was developed to assess the benefits of the guided imagery program to those who elected to participate in it. Patients who completed the guided imagery program had a shorter average length of stay, a decrease in average direct pharmacy costs, and a decrease in average direct pain medication costs while maintaining high overall patient satisfaction with the care and treatment provided. Guided imagery is now considered a complementary means to reduce anxiety, pain, and length of stay among our cardiac surgery patients. PMID- 12134379 TI - Variables for outcomes analysis. PMID- 12134378 TI - Caring Behaviors Inventory: analysis of responses by hospitalized surgical patients. AB - Hospitalized surgical patients were surveyed to determine the completeness, reliability, and validity of their responses to the 42-item Caring Behaviors Inventory (CBI) and to an abbreviated inventory derived from it. Of 354 patients, 211 completed all 42 CBI items. Item responses were reliable (alpha = 0.980) and each correlated (r = 0.192-0.630) with global nurse caring (GNC). The CBI score (item sum) also correlated with patients' rating of GNC (r = 0.569). All CBI items correlated with the CBI score (r = 0.582-0.863). Forward multiple regression showed 6 items (6, 13, 16, 28, 33, and 41) to explain nearly all variance in the CBI score (R = 0.979). Responses to these 6 items by 318 patients were reliable (alpha = 0.893); their sum was correlated with the CBI score (r = 0.964) and patients' rating of GNC (r = 0.605). This study provides further statistical support for the CBI. It justifies use of a less burdensome 6-item inventory. PMID- 12134380 TI - Pressure ulcer prevention project: an international outcomes report from Israel. PMID- 12134381 TI - Price survey. I.v. prices appear headed upward. PMID- 12134382 TI - Legal, technology wars mark oximeter market. PMID- 12134383 TI - Technology key to disease management. PMID- 12134384 TI - Look to consumer market for ways to boost sales. PMID- 12134385 TI - Materials, labor to account for many price hikes. PMID- 12134386 TI - Delivering a fixed restoration: a clinical rationale for creating a routine sequence. AB - The proper sequence of seating a restoration starts with the evaluation of the shade match. Clinical procedures then follow this sequence: adjustment of proximal contact points, assessment of fit, and occlusal adjustment. Attempts to alter this sequence may cause unnecessary occlusal adjustments and an end result of a hypooccluded restoration. PMID- 12134388 TI - Layering technique for pressed ceramic laminate veneers. PMID- 12134387 TI - The perio-aesthetic-restorative approach for anterior reconstruction--Part II: Restorative treatment. AB - Inherent differences have been identified between multidisciplinary and interdisciplinary treatment approaches. This article demonstrates the restorative techniques used for the treatment of a patient with congenitally missing lateral incisors according to the interdisciplinary concept outlined in Part I. The restorative design and material selection were based on the size and shape of the existing ridge, occlusion, evaluation of previous treatment, the ability of the restoration to mimic the natural dentition and the supporting gingival tissues, and the patient's expectations and finances. PMID- 12134389 TI - Provisionalization and definitive restoration of a missing premolar with an indirect fiber-reinforced resin fixed partial denture. PMID- 12134390 TI - Here comes the press! PMID- 12134392 TI - Applied techniques for predictable suture placement: Part 2. PMID- 12134393 TI - Self-etching adhesives: solving the sensitivity conundrum. PMID- 12134391 TI - Maximizing aesthetics, fit, and maintenance of the full-arch implant-supported porcelain-fused-to-metal restoration. AB - While traditional implant prostheses were designed to allow passive fit of the fixed partial denture (FPD) frameworks onto titanium implants, screw-retained restorations were unable to accommodate these parameters. Aesthetics and longevity were also compromised when porcelain-fused-to-metal (PFM) restorations were selected. The use of contemporary restorative materials has allowed the predictable delivery of PFMs that can restore entire arches using FPD technologies for aesthetic treatment. This article describes a modified method for the fabrication of full-arch FPDs to maximize the fit, aesthetics, and maintenance potential for this type of restoration. PMID- 12134394 TI - Clinical considerations in case selection for all-ceramic fixed partial dentures. AB - All-ceramic systems have been utilized for fixed partial dentures (FPDs) to replace a missing tooth since the early 1990s. Clinical studies that assess the long-term outcome of both anterior and posterior three-unit all-ceramic FPDs are required to determine whether they can serve as a viable treatment option for functional and aesthetic tooth replacement. This article discusses the evolution and development of all-ceramic systems for FPDs as well as the clinical and laboratory considerations for treating patients using these restorations. PMID- 12134395 TI - Dental lasers: Part 3. Use of dental lasers on hard tissue. PMID- 12134396 TI - The dental management pyramid. PMID- 12134397 TI - Reprocessing single-use devices--regulatory roles. AB - This is the third in a series of three articles about reprocessing medical devices labeled as "single use" by the manufacturer. The goal of reprocessing single-use devices (SUDs) is to save money and decrease environmental pollution. Reprocessing can be performed on SUDs that have been used on other patients or opened but not used. This article discusses the roles of the involved regulatory agencies and organizations guiding the process. The first article in the series, published in the May 2002 issue of the AORN Journal, discusses the ethical component of reusing SUDs. The second article, published in the June 2002 issue of the Journal, discusses the procedures for reprocessing (e.g., cleaning, inspecting, sterilizing, tracking, testing, validating) and establishing a reuse program. PMID- 12134398 TI - A personal responsibility to caring. PMID- 12134399 TI - The maze III surgical procedure. AB - The maze III procedure is a surgical treatment for atrial fibrillation (A fib), a rhythm problem in which the heart's upper chambers (i.e., atria) beat rapidly and irregularly, sometimes beating more than 400 times per minute. Approximately 10% of Americans older than 60 years of age suffer from A fib. The maze procedure was designed to create a maze on the atria with multiple incisions that allow sinus rhythm to be conducted to the atrioventricular node without creating a reentry circuit. Currently, it is not a widely used procedure, but its popularity continues to increase. The maze III procedure generally is reserved as a treatment of last resort for a patient with A-fib that is unresponsive to medication therapy, electrical cardioversion, surgical ablation, or pacemaker implantation. The maze III procedure, performed as open-heart surgery, has a high success rate for sustaining normal heart rhythms, usually without the need for a pacemaker; however, all other medical and nonsurgical treatment options are exhausted before the maze III procedure is performed. PMID- 12134400 TI - Ethics in perioperative practice--commitment to the patient. AB - Though often difficult, ethical decision making is necessary when caring for surgical patients. Perioperative nurses have to recognize ethical dilemmas and be prepared to take action based on the ethical code outlined in the American Nurses Association's (ANA's) Code of Ethics for Nurses with Interpretive Statements. In this fourth of a nine-part series that will help perioperative nurses relate the ANA code to their own area of practice, the author looks at the second statement, which emphasizes commitment to the patient. PMID- 12134401 TI - Creating a culture of safety. PMID- 12134402 TI - Wanted: a few good nurses. PMID- 12134403 TI - Health effects associated with medical glove use. AB - Adverse reactions to medical gloves represent an important public health issue. Accordingly, there is increasing interest in understanding the information reported to the US Food and Drug Administration (FDA) describing health effects associated with the use of medical gloves. This article provides a retrospective analysis and summary of health effects associated with medical glove use reported to the FDA. The FDA's medical device adverse event databases were searched via computer using keywords to identify reports of reactions associated with any type of medical glove. Demographic and clinical information abstracted from these reports was used to perform frequency and trend analyses. The reported medical glove-related events, including the noted trends in reporting, suggest the need for further study and continued monitoring of such reports. PMID- 12134404 TI - Ohio hospital PR pros collaborate on crisis communications plan. AB - Two member hospitals of the Akron Regional Hospital Association (ARHA), Ohio, experienced crisis situations which severely strained their public relations resources. These events were the genesis for the development of a comprehensive plan for sharing public relations resources among 11 member hospitals. The plan details procedures for sharing help in the event of a crisis or specific hospital media event. It identifies three potential situations in which it can be implemented: internal disaster, external disaster, or a specific incident unique to one of the hospitals. No occasion has yet arisen to implement the plan. PMID- 12134405 TI - Dramatic print ads raise awareness. NorthEast Medical Center's breast health campaign. AB - NorthEast Medical Center, a 457-bed independent hospital in Concord, N.C., found itself increasingly in competition with hospitals in neighboring Charlottesville. It changed from an emphasis on media relations and public relations to an advertising mode, beginning with a multimedia breast health campaign last October for Breast Health Month. The attention-grabbing visuals were adapted from the print ads and used as in-house posters to encourage employee participation in the breast screening program. PMID- 12134406 TI - The 'Adventist advantage'. Glendale Adventist Medical Center distinguishes itself. AB - Glendale Adventist Medical Center, Glendale, Calif., adopted an image-building campaign to differentiate the 450-bed hospital from its neighbors. This included the headline "Adventist Advantage," used in a series of sophisticated ads, printed in gold. In all their efforts, marketers consider the sensibilities of the sizable Armenian, Korean, Hispanic and Chinese populations. PMID- 12134407 TI - People are the focus of Deaconess. Image campaign also aims at recruiting staff. AB - Deaconess Hospital in Evansville, Ind., is recruiting hospital personnel in a campaign that also addresses the general public. Started in March 2001, the campaign consists of outdoor billboards, print ads, and radio and TV spots. All emphasize Deaconess workers and their ability to make a difference. PMID- 12134408 TI - 'It's happening at Rush' wins top PRSA award. Integrated marketing effort boosts Rush-Presbyterian-St. Luke's. AB - An award-winning integrated marketing campaign for Rush-Presbyterian-St. Luke's Medical Center, Chicago, showcases the institution's research and developments. Each edition of its 50-part series of print ads features a different case study. These are being promoted through internal communications and also with highly visible collateral materials. PMID- 12134409 TI - Small stones sets Web site apart. Froedtert Hospital updates provide valuable healthcare information. AB - Froedtert & Medical College, an academic medical center, has adopted a proactive approach to providing consumers with reliable sources of information. The Milwaukee institution has redesigned its Web site, which first opened in 1995. The new version has simplified the navigation process and added new content. Small Stones, a health resource center, also a brick-and-mortar shop, went online Feb. 1. Online bill paying was launched in May. Pharmacy refill functions are expected to be online this summer. PMID- 12134410 TI - Hospital's history is 'on the books'. Anne Arundel Medical Center celebrates its centennial. AB - Anne Arundel Medical Center, Annapolis, Md., celebrated its 100th anniversary with a history book, called "A Century of Caring"--an updated version of its 90th anniversary book. The text and photos of the book provided the material for an exhibit and a video to be seen at the medical center's Centennial Gala, at which 10 hospital pioneers were honored. PMID- 12134411 TI - The documentation dilemma--nurses poised to address paperwork burden. PMID- 12134412 TI - Workplace advocacy: how can it help. PMID- 12134415 TI - A nurse's viewpoint: a different course. PMID- 12134416 TI - Alabama State Nurses Association: clearing the myths and misconceptions. PMID- 12134418 TI - [Progress in stroke rehabilitation. Helping the brain regain proper functioning]. PMID- 12134419 TI - [Hyperbaric oxygen therapy. Will radiation ulcers healing in hyperbaric pressure? (interview by Dr. Ulrich Scharmer)]. PMID- 12134420 TI - [Diagnosis of osteoporosis. How do you manage it?]. AB - For the diagnosis of osteoporosis, conventional X-rays, densitometry and laboratory investigations are available to supplement history-taking and physical examination. The medical history should aim to identify risk factors for osteoporosis which, together with the clinical examination, can provide useful evidence for the presence of secondary osteoporosis. In the first instance, conventional X-rays serve to document osteoporotic fractures, but alone are not suitable for establishing an early diagnosis of osteoporosis. Today, the method of choice for the evaluation of osteoporosis and the fracture risk is densitometry, in particular DEXA (dual energy X-ray absorptiometry). Bone density values of > 2.5 SD (standard deviation) permit the diagnosis of osteoporosis. Currently, such biochemical markers as osteocalcin or hydroxyproline are being investigated for their diagnostic value in terms of disease activity fracture risk and the monitoring of treatment. PMID- 12134421 TI - [From effective to economical. The value of good osteoporosis therapy]. PMID- 12134422 TI - [Rehabilitation after osteoporosis-induced fracture. Getting your patient quickly back on her feet]. AB - Appropriate rehabilitation is essential for patients with osteoporotic fracture, since it not only significantly improves their quality of life, but, in many cases, may even prevent a patient becoming a candidate for the nursing home, thus considerably reducing the costs to the social system. Rehabilitation of patients with osteoporosis includes adequate pain treatment, early mobilization, specific training of muscles and coordination, instruction on how to avoid falls, on nutrition, ergotherapy and sports therapy including informing the patient how to remain permanently active without outside help. In this manner, the risk of falls can be reduced, bone density increased and bone quality improved. With regard to the choice of therapeutic measures and technical aids (walking stick, walker, corset), attention must be paid to the individual needs of the patient. PMID- 12134423 TI - [Hyperkinetic syndrome and dyslexia. Medical help for the hyperactive child]. PMID- 12134424 TI - [Neuropathic pain. Basic principles for successful therapy]. PMID- 12134425 TI - [Keratoconjunctivitis epidemica. Diagnosis, therapy and prevention]. PMID- 12134426 TI - [Assisted death in Germany. What the courts allow and what is definitely forbidden]. PMID- 12134427 TI - [Prevention of arteriosclerosis with the insulin sensitizer pioglitazone. Early intervention in diabetic patients compensates "cell ignorance"]. PMID- 12134429 TI - [New data on vasoactive drug. Patients with peripheral arterial occlusive disease walk twice as long]. PMID- 12134428 TI - [Early effect of therapy with pioglitazone. Vascular damage significantly reduced]. PMID- 12134430 TI - ["Acute intermittent porphyria" diagnosis. An odyssey in diagnosis]. PMID- 12134431 TI - [Asthma therapy. Long-term spasmolytic therapy with rapid effect]. PMID- 12134432 TI - [Dangerous postprandial glucose peaks. Risk for heart and blood vessels]. PMID- 12134433 TI - [Consistent success with coated stents. Coronary arteries stay open longer]. PMID- 12134434 TI - [32. Leukocyte values reveal much]. PMID- 12134436 TI - [Predictors of success in smoking cessation among participants of spirometric screening for COPD]. AB - The aim of the study was to evaluate factors that could predict smoking cessation after a minimal antismoking counseling during spirometric screening for COPD. Every subject filled-in a simple questionnaire on clinical signs of COPD and tobacco habit, had a spirometry performed according to ATS standards and received a short antitobacco counseling together with a booklet on how to quit smoking. Out of 800 smokers over 40 years of age, smoking history of more than 10 packyears, screened for COPD in 1999, four hundred were invited a year later for a follow-up spirometry and evaluation of anti-smoking intervention. Of 383 patients, who responded to the invitation (208 M and 175 F, mean age 56.6 +/- 10.7 yrs), 52 (13.6%) quit smoking for one year and another 48 (12.5%) quit smoking temporarily and than resumed smoking. Smokers who permanently succeeded in quitting smoking were older (60.5 vs 55.9 years p < 0.01), started smoking later (age at starting smoking 22 vs 19.5 years p < 0.001), had a shorter tobacco exposition (28.8 vs 34.3 packyears p < 0.05), had lower lung function (FEV1%pred 80.5 vs 89.2% p < 0.05) and were less nicotine dependent (FTQ score 1 vs 4.8 p < 0.00001). PMID- 12134435 TI - [Lung cancer--diagnosis and therapy delay]. AB - It is commonly known that in the course of neoplastic disease a diagnosis and therapy should be perform as fast as possible. It is particularly important for lung cancer patients. The goal of this study was to assess the diagnosis and therapy delay in unselected group of lung cancer patients, registered in Pulmonary Outpatients Clinics in all parts of Poland. MATERIAL: 20,561 lung cancer patients were registered in Pulmonary Outpatients Clinics in all parts of Poland from 1995 to 1998. RESULTS: The median delay caused by patients was about 46 days. In 33 provinces symptoms of the disease preceded diagnosis 28 to 50 days and in other 26 provinces--50 to 75 days. The median delay caused by doctors (time between first visit to the doctor and the date of diagnosis) was 65 days. In 35 provinces it was 30 to 70 days and in other 14 provinces this delay was between 70-111 days. The median time between first visit to the doctor and the beginning of therapy was 84 days. The median time between diagnosis and therapy was 30 days. Because chest physicians were also involved in the diagnosis and treatment of lung cancer patients, so for patients registered in years 1996-1998 the causes of delay connected with the function of this medical speciality were assessed. Median time between first visit to the doctor and first visit to the chest specialist was 38 days. Median delay to bronchoscopy was 26 days and to the diagnosis 46 days. CONCLUSION: Delay of diagnosis and therapy vary widely among different provinces of Poland. The delay generated by family doctors and chest physicians are very important and require a deeper evaluation on the province level in the future. PMID- 12134437 TI - [Awareness of airflow obstruction together with antismoking advice increases success in cessation smoking]. AB - COPD is one of the leading causes of mortality and increased morbidity in developed world. In advanced disease it also imposes an important economic burden on societies. The main etiologic factor for COPD is tobacco smoking. The aim of the study was to asses if the awareness of airflow obstruction combined together with a minimal antismoking advice in middle aged smokers increases the quitting rate. Out of smokers participating in mass spirometric screening for COPD in five polish towns, we invited 734 (300 with airflow limitation and 247 with normal lung function) for a follow-up. During the second visit, at one year, spirometry was performed and smoking status was assessed. Non-smoking was validated with carbon monoxide measurements in exhaled breath. Patients who did not come for the follow-up visit were considered as smokers. Of 734 smokers invited, 543 (74%) presented for the follow-up visit. All smokers tried to modify the habit. Number of cigarettes smoked at one year was reduced by -5.5 (p < 0.001) in patients with airflow limitation and -2.2 (ns) in smokers with normal lung function. One year quit rate in smokers with airflow limitation was 11.1% vs 7.6% in smokers with normal lung function (ns). When the calculation was made for those who had the follow-up the quit rates were 15.1% vs 9.9% (p < 0.05). Cessation of smoking was correlated with lung function. Those smokers who stopped smoking permanently or tried to quit had lower FEV1 (p < 0.01) and FEV1/FVC (p < 0.05), than those who continued to smoke. PMID- 12134438 TI - [Pulmonary lymphangioleiomyomatosis: presentation and results of treatment]. AB - Lymphangioleiomyomatosis is a rare lung disease of unknown aetiology that affects only women. Eight premenopausal women with LAM confirmed by lung biopsy specimens were observed in 1984-2001. The most common presenting feature was exertional dyspnea (6) followed by chylous pleural effusions and pneumothoraces. In two women severe airflow obstruction was observed at presentation. HRCT revealed characteristic cysts in all cases. All women were given hormonal therapy (tamoxifen, medroxyprogesterone). The best results of treatment were achieved in cases with chylothoraces. PMID- 12134439 TI - [Clinical presentation of own idiopathic pulmonary fibrosis patients according to international consensus statement]. AB - Clinical presentation of idiopathic pulmonary fibrosis (IPF) restricted, according to current definition to usual interstitial pneumonia (UIP) was presented. 62 patients (39 males and 23 females) were assessed. The diagnosis of IPF/UIP has been based upon a combination of clinical, radiographic and physiologic features in majority of patients. Histologic confirmation from lung biopsy has been obtained in 16% of cases. Mean age of the patients was 64.4 +/- 8.0 years. Mean duration of symptoms was 20.1 +/- 14.1 months. The main symptom was exertional dyspnea. Crepitations were found in 98% of patients. Lung volumes were in normal range in substantial number of patients; TLC in 15 (24%) and FVC in 33 (53%) out of 62 patients. Disturbances of lung function concerned mainly gas exchange (DLCO diminished in 92% of cases) and lung compliance (diminished in all patients). Presentation of clinical, radiographic and physiologic features of IPF/UIP in a homogenous group of patients may be helpful in diagnosis of this common interstitial lung disease. PMID- 12134440 TI - [Atrial natriuretic excretion in patients with obstructive sleep apnea syndrome (OSAS)]. AB - During obstructive sleep apneas stimuli, that may increase excretion of atrial natriuretic peptide (ANP) occur. The aim of the study was the evaluation whether in patients with OSAS levels of ANP are significantly different in relation to sleep or wakefulness and in relation to disturbances of ventilation during sleep and wakefulness. The material of the study consisted of 34 patients with OSAS (age 25-65 years). There were no differences in the levels of ANP late in the evening, during sleep and early in the morning. There were 2 groups of the patients: with low (< 70 pg/ml, mean at 21 p.m. 9.7 +/- 8.7 pg/ml, at. 2 a.m. 12.5 +/- 9.3 pg/ml, at 6 a.m. 14.4 +/- 15.1 pg/ml) and high (> 70 pg/ml, mean at 21 p.m. 148.6 +/- 232.9 pg/ml, at 2 a.m. 119.5 +/- 45.5 pg/ml, at 6 a.m. 164.9 +/ 161 pg/ml) ANP levels. As compared with patients with low ANP levels, patients with high ANP levels were older and more obese, more frequently had concomitant COPD, lower VC and FEV1, higher daytime PaCO2 and lower PaO2; most of them had peripheral edema. In patients with high ANP levels there was more profound mean arterial blood desaturation during sleep apnoeas than in patients with low ANP levels (SaO2 75 +/- 8% vs 81 +/- 4%, p < 0.001), although apnea index and mean apnea duration were similar in both groups. CONCLUSIONS: In patients with OSAS the daytime and sleep levels of ANP are similar. High levels of ANP can be found in OSAS patients with impaired daytime ventilation and gas exchange, and profound arterial oxygen desaturation during sleep apnoeas. PMID- 12134441 TI - [Rhythm of melatonin excretion in obstructive sleep apnea syndrome]. AB - Melatonin is a pineal hormone that regulates sleep and wake status. Melatonin concentrations in blood serum were measured using radioisotope method in 33 males (age 48 +/- 10) with obstructive sleep apnea syndrome. The following melatonin concentrations were measured: 54 +/- 72 pg/ml (9 p.m.) 424 +/- 838 pg/ml (2 a.m.) and 307 +/- 534 pg/ml (6 a.m.). In patients with high peak melatonin concentration (> 200 pg/ml) as compared with the patients with low peak melatonin concentration (< 200 pg/ml) there were higher index of respiratory disorders during sleep (53 +/- 18 vs 38 +/- 20, p < 0.05) and lower minimal SaO2 during sleep apnea (52 +/- 17% vs 70 +/- 10%, p < 0.05); they were also more tired in the morning and were more sleepy during the day (Epworth sleepiness scale 17 +/- 6 vs 11 +/- 6, p < 0.01). In 66% of patients peak melatonin concentration was observed at 2 a.m. In 24% of patients peak melatonin secretion was prolonged to early morning hours. CONCLUSIONS: In most of patients there is peak melatonin excretion at 2 a.m. Patients with high apnea and hypopnea index and daytime sleepiness have high peak melatonin concentrations. PMID- 12134442 TI - [Pulmonary amyloidosis--own experience]. AB - Three cases of amyloidosis were described. In all diagnosis was confirmed by histological examination. There was amyloidosis limited to the lungs in 2 cases and in 1 generalised. In 1 patient lobectomia was performed. Next 2 pts were treated with prednisone and cytostatic drugs (melphalane and cyclophosphamide). PMID- 12134443 TI - [Frequency and causes of hemoptysis and role of bronchoscopy in patients with normal chest roentgenogram hospitalized in the Department of Physiopneumonology Silesian Medical University in the years 1961-1996]. AB - Among 3498 patients with hemoptysis 513 had normal chest x-ray picture. Bronchoscopy performed in all these patients allowed to recognise malignant neoplasms of the lungs, trachea and pharynx in 109 patients. In 222 patients- nonspecific bronchitis and in 46--tuberculosis were recognised. In 86 patients diagnosis was not established. PMID- 12134444 TI - [Tuberculosis of ear and cutis]. AB - We describe a 26 year old woman treated for near a year because of deafness, purulent otitis, cough and purulent skin lesions. She was treated by otolaryngologist. Sputum examination for acid fast bacilli and chest x-ray were not done a that time. In hospital chest x-ray revealed a cava in left lung and tuberculous bacilli were found in sputum and in content from ear. Tuberculin test was positive. Tuberculosis of lung, skin and cor was recognized. After 4 months of initial treatment patient was referred to outpatient department for further therapy. PMID- 12134445 TI - [Tuberculosis of the bone]. AB - We describe a 75 year old patient treated because of arthrosis and recurrent pneumonia for a year. In hospital acid-fast bacilli were found in sputum. Chest x ray revealed massive inflammatory and fibrous lesions. Pelvis x-ray revealed lesions estimated as tuberculosis. Tuberculosis of lungs and pelvis bones was recognised. After 3 months of antituberculous treatment patient was referred to outpatient department for further therapy. PMID- 12134446 TI - [Nocardia and human nocardiosis]. AB - Nocardia spp. are pathogens commonly found in soil worldwide, and they cause mostly opportunistic infections in humans and animals, complicating both immunodepressive states and primary diseases. Nocardiosis is difficult to proper microbiological and clinical diagnosis because of its non-specific symptoms, which manifest as the cutaneous and sub-cutaneous infections, lung symptoms and the dissemination through the bloodstream to other organs. General characteristics of Nocardia, human nocardiosis as well as the microbiological diagnostics routine and treatment are discussed. PMID- 12134447 TI - [Antibiotic prophylaxis in bronchoscopy: When? How?]. PMID- 12134448 TI - Educating consumers. PMID- 12134449 TI - Free health headlines on your site. PMID- 12134451 TI - Advice services connect organization with consumers. PMID- 12134452 TI - How can you select job candidates who will be contributors to team? PMID- 12134450 TI - Measuring patient satisfaction. PMID- 12134453 TI - Supporting your organization in managed care contracting. PMID- 12134454 TI - Users eye consolidation in OR IS market. PMID- 12134455 TI - OR RN competence holds up, but new nurses' does not. PMID- 12134456 TI - Fibrin sealants likely to stick around. PMID- 12134457 TI - Sterilization & infection control. Needed: a process for loaner instruments. PMID- 12134458 TI - What's best mix of procedures for ASC? PMID- 12134460 TI - Painful lesions on the arches of the feet. A pustular, pruritic eruption can be difficult to diagnose and problematic to treat. PMID- 12134459 TI - HIPAA and business associate agreements. PMID- 12134462 TI - "See one, HAVE one, do one, teach one". A new medical education paradigm for midlife and beyond. PMID- 12134461 TI - Prevention for the older woman. A practical guide to managing cardiovascular disease. AB - American women are more likely to die from cardiovascular disease than from any other cause. Although hypertension is most prevalent, most deaths are attributed to coronary heart disease. Heart disease in women manifests approximately 12 to 15 years later than in men, up until menopause. Then the severity of coronary artery lesions in women accelerates until it equals or surpasses that of men by the late 70s or early 80s. Physicians can help older women reduce their risk for heart disease and stroke by managing hypertension and hypercholesterolemia and providing beta-blocker treatment when indicated after MI. Nonpharmacologic interventions may be effective as well. New guidelines for aspirin help identify women under age 80 who would benefit most from antiplatelet therapy. PMID- 12134463 TI - Movement disorders. Keys to identifying and treating tremor. AB - Tremor is one of the most common adult neurologic disorders. It is characterized by Involuntary, rhythmic and oscillatory movements of a body part. Tremor affects quality of life by impeding simple motor tasks and activities of daily living and heightening self consciousness. Severe tremor can undermine independence, relationships, and recreational and social activities. Initial clinical objectives include determining the tremor type and ruling out Parkinson's disease. Severe cases typically require pharmacologic therapy, and in some instances, surgical intervention; both strategies for severe cases, however, have drawbacks. PMID- 12134464 TI - The importance of executive deficits. Assessing the older patient's capacity to remain at home. PMID- 12134465 TI - Test strategies for the detection of myocardial damage. AB - In the space of half a century, cardiac marker testing has advanced incrementally from enzymes present in nearly all tissues to proteins having remarkable specificity for myocardium. Markers with other desirable properties, such as earlier release, have also been introduced and others may be anticipated, although a single perfect marker is not on the horizon. Optimum application of these new markers still requires improved robustness and harmonization of commercial assays, and continuing insights in the pathophysiology of acute coronary syndromes. As these advances occur, future testing will likely focus more on therapeutic decisions than on arbitrary diagnostic classifications. PMID- 12134466 TI - Liver function: test selection and interpretation of results. AB - Liver function tests are used to detect, specifically diagnose, and estimate the severity of hepatic disease. Effective interpretation of the hepatic function panel requires knowledge of underlying pathophysiology and the characteristics of panel tests. This article includes a working classification of liver disease, a list of liver functions with the tests appropriate to each function, and a guide to panel interpretation and further laboratory investigation. PMID- 12134467 TI - Hypercoagulability test strategies in the protein C and protein S pathway. AB - The protein C-protein S pathway is critical in controlling normal clot formation to provide homeostasis between thrombosis and hemorrhage. Abnormalities have been described in which the pathway is altered because factor V cannot be degraded by activated protein C, producing activated protein C resistance. It has been known for nearly two decades that deficiencies in protein C and protein S can result in impaired inhibition of clot formation resulting in increased risk for thrombosis. This article describes the testing strategies associated with the diagnosis of activated protein C resistance, protein C deficiency, and protein S deficiency. PMID- 12134468 TI - The laboratory evaluation of platelet dysfunction. AB - An algorithm for platelet function testing is presented in Fig. 2. The proposed algorithm takes into account the importance of a thorough clinical bleeding history and provides an integration of screening tests with more specific diagnostic assays. As our understanding of the biochemical and molecular aspects of platelet function becomes increasingly refined, it is becoming clear that current clinical testing strategies may not be adequate to identify all significant platelet function defects. The repertoire of distinct platelet agonists is increasing to allow a more discrete look at the function of isolated platelet membrane agonist receptors, especially those involved in platelet activation by ADP, thrombin, and collagen. In addition, FACS analysis of platelets, with emphasis on surface membrane glycoprotein expression, may offer a more specific and quantitative view of platelet membrane constituents and their function. Furthermore, screening tests evaluating platelet function under physiologic and pathologic flow conditions are becoming important adjuncts to in vitro analyses, and may accentuate platelet function abnormalities, not easily discerned by platelet aggregation studies. It remains to be seen whether such screening tests will better predict clinical bleeding or thrombotic risk. PMID- 12134469 TI - Selected endocrine test strategies. AB - The diagnosis of endocrine disorders lends itself to sequential test strategy. The strategies outlined in this article deal with problems that are either commonly encountered in clinical practice, reflect recently acquired knowledge, or both. Wherever possible a clearly defined algorithmic approach is used. The purpose is to emphasize the most appropriate general approach to test strategy. PMID- 12134470 TI - Modern approaches to the investigation of vitamin B12 deficiency. AB - The classic workup of a patient for possible PA is revisited in light of the vanishing Schilling test. The vagaries of testing for B12 and blocking antibodies are reexamined. The advantages and disadvantages of newer tests such as MMA and serum gastrin levels are catalogued. At this juncture in the evolution of new test strategies, there is a considerable controversy regarding the significance of high MMA levels in the face of normal B12 levels, particularly in the elderly. Hopefully, this controversy will soon be resolved and the newer crop of tests will be proven and accepted in the workplace. Still, the words of Alexander Pope spring to mind: "Be not the first by whom the new are tried, Nor yet the last to lay the old aside." PMID- 12134471 TI - Antinuclear antibody testing. AB - The ANA test is an excellent screening test for patients with SLE and a few other connective tissue diseases. The LE cell preparation is an assay that is subjective and costly. Because of the presence of a superior screening test (the ANA) and superior specific auto-antibody tests, the author recommends that the use of LE cell preparations be discontinued. ANA screening tests may be performed either by indirect microscopic serology (usually IFA) or EIA. The latter technique is readily automated and many new products for this screening test have appeared in the past decade. The products differ, however, and laboratories are cautioned to test each in the context of the clinical needs of their clinicians. Proper use of the ANA test requires each laboratory to determine the cutoff used under their conditions of assay. Although either ANA screening test has a high negative predictive value in numerous studies, proper selection of patients to be tested is key to improving the predictive value of a positive result. The American College of Rheumatism criteria are reviewed and recommended as part of the patient selection process for this testing. PMID- 12134472 TI - Advances in pretransfusion infectious disease testing: ensuring the safety of transfusion therapy. AB - The public expects a zero-tolerance policy for the transmission of infectious agents by blood transfusion. Although unrealistic, the efforts to reach this goal have produced an extremely safe albeit costly blood supply [82]. Blood collecting agencies, the FDA, physicians, and scientists have over the past 20 years created a complex system of layers of protection to interdict transfusion-transmitted infections (Fig. 2). As new, exotic, potentially blood transmittable infectious agents evolve [83], new barriers will be erected to [figure: see text] interdict these agents. In the interim, the US blood supply is the safest in the world. PMID- 12134473 TI - Appropriate use of clinical microbiology tests. AB - Laboratory medicine lacks the tools necessary to define appropriate test use; nonetheless, existing laboratory test characteristics, although inadequate, provide a common starting point for developing definitions of appropriate test use. As Lundberg [1], Szczepura [15], and van Walraven and Naylor [2] have emphasized, developing a process for defining appropriate laboratory use should receive a high priority. There is a particular need to develop methods for standardizing studies [59]. Laboratory medicine does not, however, lack the tools necessary to change test use. Although past interventions were largely ineffective, there is growing evidence that test use can be changed by use of a variety of approaches. By using the existing tools there is much that can be done to change inappropriate test use, such as minimizing redundant testing or the use of tests that have no clinical relevance. The real opportunities will come when there are scientifically and medically sound definitions of appropriate test use that can be used to change test use and improve patient care in a cost-effective way. PMID- 12134475 TI - The use of expert systems for improving test use and enhancing the accuracy of diagnosis. AB - The experience with BloodLink and LAS provides strong evidence that electronic knowledge support has a significant and lasting effect on laboratory test ordering by physicians. Compliance with CPG is also improved despite the ability of the guideline process to keep up to date. As discussed previously, the limitations to the application of protocols and CPG can be eliminated by electronic knowledge support. Indeed, this technology may be considered the future salvation of evidence-based medicine. It is also apparent that using the expert system to provide interpretations of the results ensures optimal use of the tests. The ability to reduce costs by 20% using stringent use control measures is meaningless if by misunderstanding, all of the potential information is not understood. In the future, as computer connectivity becomes pervasive within the practice of medicine, systems, such as BloodLink and LAS, will be used routinely for many aspects of care. Molecular genetic testing and proteomics will become a major component of routine medical practice. The complexity of what to order, the implications of performing the tests, on whom to do them, and what the vast array of data mean, will demand the use of CARTKS. It is becoming increasingly popular for patients to obtain their own results and to manage their own medical record. In some jurisdictions patients may order their own tests. We will undoubtedly see the development of knowledge support tools that are designed for patient use. The CARTKS provided by computer systems is the most powerful approach to use control and it has the additional benefit of doing so while enhancing medical care. PMID- 12134474 TI - The use of ordering protocols and other maneuvers: the Canadian experience. AB - Canada's socialized, single-provider, fee-for-service environment has provided an opportunity for widespread implementation and evaluation of a number of utilization control measures. Enforced consolidation of services certainly eliminates redundancy but the implementation cost and disruption of such a measure is high. Whether noncompetition will eventually eliminate any cost savings achieved is difficult to predict. Risk sharing in which ordering doctors and laboratories are paid from the same source of funds and both groups stand financially responsible for excess utilization seems to be an effective approach. From a fiscal standpoint it is, but such measures can create ill will. Establishing utilization caps has an absolute fiscal effect but, unless very carefully designed and monitored, may create more problems than they solve. Utilization control can be achieved using a minimal list requisition form. Form control is also essential to ensure the success of protocol and CPG implementation. The development of protocols and CPGs has proved to be very effective in reducing laboratory testing while also standardizing aspects of medical practice. Such guidelines work well when (1) dealing with testing areas of high volume (or high cost); and (2) amenable to simple rules on which there generally can be agreement. A collaborative implementation environment is necessary. After about 15 such protocols, however, it becomes increasingly difficult to define new areas to target. Physician chart audits are a useful adjunct to help deal with problem areas and to keep all physicians highly aware of good ordering practices. Utilization problems have not been solved in the Canadian system. Certain ventures, however, have proved to have a positive effect. It is likely that when electronic knowledge support tools become a standard feature of medical practice the protocol-CPG approach will be maximized. PMID- 12134476 TI - Using Web technologies for implementing testing strategies. AB - Although the application of Web technologies to laboratory services is relatively new, it is evolving rapidly and presents both promises and challenges to those adopting it for test strategy deployment. Its application can be helpful in disseminating information about a testing strategy once it is formulated, but it's more important potential contribution is in facilitating clinicians' use of the strategy. There is an array of ways Web technologies can be used to facilitate both the initiation of a strategy at the point of order entry and ensuring optimal use of the laboratory results that emanate from a testing strategy. Web technologies present a rich set of tools for those dedicated to enhancing the value of the laboratory's contribution to patient care. PMID- 12134477 TI - Noninvasive testing in the clinical laboratory. AB - We have chosen to provide a brief review of only three of the numerous existing, emerging or future applications of novel technologies that provide noninvasive testing. Clinical laboratory testing over the recent decades has been characterized by a remarkable increase in the variety and volume of [figure: see text] information made available to assist physicians in the practice of medicine. Among current trends that facilitate prompt delivery of results, noninvasive testing holds great promise. Technologies that already exist or that have not yet been developed may extend the opportunities to exploit this approach. Noninvasive testing has other obvious attractive features; safety, painlessness and adaptability to special care situations. Our profession may well enjoy additional vitalization from the further evolution of this facinating and important field. PMID- 12134479 TI - The relationships between the phagocytic activities of arterial, capillary, and venous blood leukocytes in normal and pathological states. PMID- 12134478 TI - The delayed effect of cortagen on the restoration of injured nerve function. PMID- 12134480 TI - Effect of female sex steroid hormones on the functions of the peripheral blood phagocytizing cells. PMID- 12134481 TI - Long-term potentiation of the glutamate-activated inward current induced by 8-Br cGMP in nerve cell. PMID- 12134482 TI - Technology of obtaining goat zygotes with known time of formation suitable for microinjection of recombinant DNA in order to create transgenic animals. PMID- 12134484 TI - Latent periods of the components of evoked potential. PMID- 12134485 TI - Involvement of heat shock proteins in the phenomenon of cell protection against apoptosis mediated by inhibitors of plasma membrane chlorine channels. PMID- 12134483 TI - Interactions of cytokines and their components in nervous and lymphoid tissue cultures. PMID- 12134486 TI - Effects of a calcium-binding agent in the musculus soleus of rats against the background of simulated gravitational unloading. PMID- 12134487 TI - Cardioprotective effects of transthoracic microelectrostimulation. PMID- 12134488 TI - Weightlessness stimulates growth of statoconia. PMID- 12134490 TI - The possible mechanisms of stimulation of cyanobacterium growth by the passage through the Carassius auratus intestine: an experimental study. PMID- 12134489 TI - Long-term habituation as particular case of acclimation. PMID- 12134491 TI - Seasonal dynamics of the numbers of Anabaena flos-aquae (Lyngb.) Breb. akinetes in the surface layer of bottom sediments and bulk water. PMID- 12134492 TI - Discovery of viable methanotrophic bacteria in permafrost sediments of northeast Siberia. PMID- 12134493 TI - Effect of individual heterozygosity on fruiting in dwarf Siberian pine (Pinus pumila (Pall) Regel). PMID- 12134495 TI - The effect of climate on long-term fluctuations of the numbers of passerine birds. PMID- 12134494 TI - Effect of atmosphere temperature near the soil surface on air convection in mammalian burrows (as exemplified by of steppe marmot burrow). PMID- 12134496 TI - Genetic changes in salmon transplanted into the White Sea basin. PMID- 12134497 TI - Histochemical indices of sexual dimorphism as a species-specific characteristic (using skin glands of far eastern voles as an example). PMID- 12134498 TI - Antimutagenic activity of crown-containing compounds. PMID- 12134499 TI - Effect of enhanced concentration of CO2 on transpiration and kinetic parameters of photosynthetic CO2 gas exchange in sugar beet grown at different concentrations of nitrate. PMID- 12134500 TI - Effect of geomagnetic field changes on the rate of spontaneous chromosome aberrations in human somatic cells. PMID- 12134501 TI - New findings of a specialized spine cuticle in porcupines (Rodentia: Hystricomorpha) and tenrecs (Insectivora: Tenrecidae). PMID- 12134502 TI - DNA breaks in the centromeric heterochromatin of polytene chromosomes. PMID- 12134503 TI - Photosynthesis, respiration, and transpiration in maize seedlings under hypoxia induced by complete flooding. PMID- 12134504 TI - Isozyme spectra of potato peroxidases in ring rod pathogenesis. PMID- 12134505 TI - Responses of rat left ventricle cardiomyocytes and capillaries to 2G hypergravity. PMID- 12134506 TI - A new binding mode of competitive antagonists to metabotropic glutamate receptors exemplified by the mGluR1-receptor antagonist AIDA (RS-aminoindan-1,5 dicarboxylic acid). PMID- 12134507 TI - Hydrogen-phosphate and phosphate-phosphate spin systems resolved in EPR spectra of photosensitized phosphates using computer analysis. PMID- 12134508 TI - A model of molecular constructions of a combined bivaccine. PMID- 12134509 TI - Multiple genetic differences between the monkeypox and variola viruses. PMID- 12134510 TI - DNA sequences eliminated during chromatin diminution from somatic cell chromosomes of Cyclops kolensis. PMID- 12134511 TI - Comparative analysis of HIV-1 resistance to AZT and AZT H-phosphonate in a cell culture. PMID- 12134512 TI - Fungicidal activity of khlorin photosensitizers. PMID- 12134513 TI - Contribution of the processes of solvation of nonequilibrium states of cofactors to charge separation and electron transfer in reaction centers of Rhodobacter sphaeroides. PMID- 12134514 TI - A new pseudopeptide motif for designing specific DNA-binding compounds capable of recognizing long DNA sequences. PMID- 12134515 TI - Comparative analysis of the structural role of proteins and polysaccharides in cell walls of the yeasts Hansenula polymorpha and Saccharomyces cerevisiae. PMID- 12134516 TI - Transgenic tobacco plants produce miniantibodies against human ferritin. PMID- 12134517 TI - A block of CpG dinucleotides that are differentially methylated in erythroid and nonerythroid cells is located upstream of the cluster of chicken alpha-globin genes. PMID- 12134518 TI - Heat-induced generation of reactive oxygen species in water. PMID- 12134519 TI - Guanidinium derivatives as reversible inhibitors of cholinesterases of various origins. PMID- 12134520 TI - Humic acids of low peat and brown coal. PMID- 12134521 TI - Carpal tunnel releases done for normal median nerves. PMID- 12134522 TI - Health effects and occupational exposures among office workers near the World Trade Center disaster site. AB - The extent of health effects and exposure to environmental contaminants among workers and residents indirectly affected by the September 11, 2001, attack on the World Trade Center (WTC) is unknown. The objective of this study was to evaluate concerns related to health effects and occupational exposures three months after the WTC disaster among a population of employees working in a building close to the disaster site. A cross-sectional questionnaire survey was performed of Federal employees working near the WTC site in New York City (NYC) and a comparison group of Federal employees in Dallas, Texas. An industrial hygiene evaluation of the NYC workplace was conducted. Constitutional and mental health symptoms were reported more frequently among workers in NYC compared to those in Dallas; level of social support was inversely related to prevalence of mental health symptoms. Post-September 11th counseling services were utilized to a greater degree among workers in NYC, while utilization of other types of medical services did not differ significantly between the groups. No occupational exposures to substances at concentrations that would explain the reported constitutional symptoms were found; however, we were unable to assess potential occupational exposures in the time immediately after the WTC disaster. There is no evidence of ongoing hazardous exposure to airborne contaminants among the workers surveyed. Specific causes of reported constitutional health symptoms have not been determined. Health care providers and management and employee groups should be aware of the need to address mental health issues as well as constitutional symptoms among the large number of workers in the NYC area who have been indirectly affected by the WTC disaster. PMID- 12134523 TI - Ergonomic and psychosocial factors affect daily function in workers' compensation claimants with persistent upper extremity disorders. AB - Pain and other symptoms associated with work-related upper extremity disorders (WRUEDs) can lead to significant distress, lost function, and disability. Identifying factors associated with decreased upper extremity function may lead to the development of more effective interventions. In this study, participants were 165 government employees (127 female, 38 male) with an accepted workers' compensation claim (< 90 days from claim filing) for a WRUED who were unable to perform their normal work. Participants completed baseline measures of upper extremity functional limitation, symptoms, general health status, problem solving orientation, pain coping, and workplace factors. After controlling for pain and gender in a multiple regression analysis, greater functional limitation was further explained by: (1) upper extremity symptoms other than pain (e.g., sleep disturbance, numbness and tingling), (2) symptoms in one or both hands, (3) feeling overwhelmed by pain, (4) low confidence in problem solving abilities, and (5) higher ergonomic risk factor exposures at work. The final model accounted for 47.4% of the variance in upper extremity function, F(7157) = 4.33, P < 0.05. Mental health status was related to functional limitation in univariate, but not multivariate analyses. These results suggest that improving function in this population may require: (1) pain coping techniques and active problem solving to overcome functional barriers, and (2) reduction of workplace ergonomic risk exposure. PMID- 12134524 TI - Community cancer assessment in response to long-time exposure to perchlorate and trichloroethylene in drinking water. AB - In response to concerns about cancer stemming from drinking water contaminated with ammonium perchlorate and trichloroethylene, we assessed observed and expected numbers of new cancer cases for all sites combined and 16 cancer types in a California community (1988 to 1998). The numbers of observed cancer cases divided by expected numbers defined standardized incidence ratios (SIRs) and 99% confidence intervals (CI). No significant differences between observed and expected numbers were found for all cancers (SIR, 0.97; 99% CI, 0.93 to 1.02), thyroid cancer (SIR, 1.00; 99% CI, 0.63 to 1.47), or 11 other cancer types. Significantly fewer cases were observed than expected for cancer of the lung and bronchus (SIR, 0.71; 99% CI, 0.61 to 0.81) and the colon and rectum (SIR, 0.86; 0.74 to 0.99), whereas more cases were observed for uterine cancer (SIR, 1.35; 99% CI, 1.06 to 1.70) and skin melanoma (SIR, 1.42; 99% CI, 1.13 to 1.77). These findings did not identify a generalized cancer excess or thyroid cancer excess in this community. PMID- 12134525 TI - Air pollution and emergency room visits due to chronic lower respiratory diseases in the elderly: an ecological time-series study in Sao Paulo, Brazil. AB - The objective of this study was to investigate the effect of daily air pollution levels (carbon monoxide, nitrogen dioxide, ozone, sulfur dioxide, and particulate matter with an aerodynamic profile < or = 10 microns) on morbidity by using the daily number of emergency room visits due to chronic lower respiratory disease (CLRD) in people older than 64 years of age in the city of Sao Paulo, Brazil, from 1996 to 1998. Generalized additive Poisson regression models adjusted for long-time trend and weather (nonparametric functions), weekdays (dummy variable), and daily number of nonrespiratory admissions were adopted. Ozone and sulfur dioxide were the pollutants statistically associated with CLRD visits. Interquartile range increases in the 6-day moving average of sulfur dioxide (11.82 micrograms/m3) and in the 4-day moving average of ozone(35.87 micrograms/m3) increased CLRD emergency room visits in 18% and 14%, respectively. These results reinforce the idea that air pollution may promote adverse health effects in the elderly. PMID- 12134526 TI - Depression-related short-term disability in an employed population. AB - There has been a growing realization that the number of workplace disability claims for mental and nervous disorders is increasing. Yet, little is known about the working population disabled by these disorders. Absence of basic information describing this population makes it virtually impossible to plan effective workplace programs. Using administrative data collected from three major Canadian financial/insurance sector employers, we focus on one group of disorders- depression. In this study, we report the prevalence of short-term disability due to depression and describe the characteristics of workers affected and their disability outcomes. We observed that compared with other nervous and mental disorders, depression-related short-term disability generally affected more employees, lasted longer, and had a higher rate of recurrence. In addition, at the end of their episodes more than three quarters of workers returned to work. These estimates suggest that the potential magnitude of the impact of short-term disability should be a concern for employers. This study helps identify the main characteristics of workers who develop depression-related disability. It also helps clarify what happens to those on short-term disability. PMID- 12134527 TI - Cancer incidence among Finnish nuclear reactor workers. AB - Because of their well-documented exposures to repeated low doses of ionizing radiation, nuclear reactor workers offer an opportunity to assess cancer risk from low-dose radiation. A cohort of all 15,619 Finnish nuclear reactor workers was established through dose-monitoring records. A questionnaire survey revealed no substantial differences in consumption of tobacco or alcohol between different exposure groups nor between nuclear power company employees and contract workers. In the follow-up for cancer incidence, no clear excess in cancer incidence was observed overall, nor was any observed in any of the specific cancer types studied. There was little evidence for an association between cancer incidence and cumulative radiation dose, but the statistical power was limited. More precise estimates will be available from an international collaborative study of nuclear industry workers, including our cohort. PMID- 12134528 TI - Outdoor air pollution and acute respiratory infections among children in developing countries. AB - Acute respiratory infections (ARIs) are the most common cause of illness and death in children in the developing world. This review focuses on outdoor air pollutants associated with pediatric ARI mortality and morbidity. Studies were identified using MEDLINE and other electronic databases. Four studies showed an increase in infant mortality in relation to outdoor air pollution. Short-term follow-up and time-series studies suggest that air pollutants act as risk factors for respiratory infection. Air pollution exposure increases the incidence of upper- and lower-respiratory infections in children. Because complex pollution mixtures are present in the studied urban areas, pollutant levels at which ARI risk would be expected to increase cannot be determined. Children may be at greater risk, given the poor environmental and nutritional conditions prevalent in developing countries. PMID- 12134529 TI - Toxic inhalation fatalities of US construction workers, 1990 to 1999. AB - Construction workers account for a disproportionately large number of occupational fatalities. A small percentage of these deaths is attributable to poisoning. Risk factors for such deaths using national data have not been reported previously. Construction poisoning fatalities from 1990 to 1999 in the Occupational Safety and Health Administration Integrated Management and Information System data set were analyzed. Risk and risk factors were determined using Bureau of Labor Statistics and census data. Eighty-seven poisoning deaths of construction workers are characterized, all attributable to toxic inhalation. Cellular and simple asphyxiants accounted for the largest numbers of fatalities. The majority of these deaths occurred in confined spaces. Water, sewer, and utility line workers are at increased risk for poisoning fatality. Toxic inhalation fatalities in the construction industry are preventable. Extending the Occupational Safety and Health Administration's confined space standard could save lives, particularly among water, sewer, and utility line industry workers. PMID- 12134530 TI - Cancer of the brain and nervous system and occupational exposures in Finnish women. AB - Occupational agents were evaluated for the risk of brain-nervous system cancer in a cohort of 413,877 Finnish women with blue-collar occupations in 1970. Observed and expected numbers of incident cases and the intensities of exposure to 25 agents were generated for 183 job titles from 1971 to 1995. Poisson regression models linked incidence and exposure data. Increased risks were found for medium/high intensities of iron (standardized incidence ratio [SIR], 2.15; 95% confidence interval [CI], 0.96 to 4.80), oil mist (1.95; 0.97 to 3.90), any chromium compounds (1.51; 0.85 to 2.67), electromagnetic fields (1.37; 0.98 to 2.10), aliphatic and alicyclic hydrocarbon compounds (1.34; 0.80 to 2.27), lead (1.27; 0.81 to 2.01), cadmium (1.26; 0.72 to 2.22), and aromatic hydrocarbon compounds (1.20; 0.71 to 2.03). Strengths of the study include fair number of cases, virtually complete case coverage, and a high-quality job exposure matrix. Ecological design and cross-sectional job assessment introduced exposure misclassification and tended to drive risk estimates toward unity. PMID- 12134531 TI - Firefighters' blood pressure and employment status on hazardous materials teams in Massachusetts: a prospective study. AB - We evaluated the association between hypertension and changes in employment status in 334 hazardous materials firefighters. Firefighters were categorized by blood pressure (BP) at baseline (1996 or 1997) and subsequent follow-up examinations (1997, 1998, and 1999). They were followed up for a maximum of 4 years for possible adverse outcomes (death, placement on "injured-on-duty" status, termination of duty, resignation, retirement, or incident cardiovascular disease). In several analytic models, we found that firefighters with stage II hypertension (BP > or = 160/100 mm Hg) were consistently 2 to 3 times more likely to experience an adverse outcome compared with those with normal BP. Cox proportional-hazards regression was used to adjust for age, body mass index, smoking, cholesterol, and antihypertensive medication. In these models, the hazard ratio for stage II hypertension was 3.2 (95% confidence interval [CI], 1.50 to 7.04, P = 0.003) and for untreated stage II hypertension, it was 4.6 (95% CI, 2.08 to 10.11, P = 0.0002). Firefighters with a BP > or = 160/100 mm Hg should receive further evaluation and demonstrate improved BP control before being determined fit for duty. PMID- 12134532 TI - Association of comorbidity and outcome in episodes of nonspecific low back pain in occupational populations. AB - We examined the relationship between comorbidity and first return to work after episodes of work-disabling, nonspecific low back pain (NSLBP). An inception cohort of workers with new episodes of NSLBP was identified from administratively maintained occupational health records. We compared 6-month return-to-work rates between workers with one or more comorbid conditions with those without documented comorbidity. Workers with comorbidity were 1.31 times more likely to remain work disabled than those with uncomplicated NSLBP, after adjusting for age, gender, lifting demands, and company membership (adjusted hazards ratio [HR] = 1.31; 95% confidence interval [CI] 1.12 to 1.52). Concurrent injury (i.e., sprains or strains of the neck, upper extremity, and lower extremity; contusions; and lacerations) had the strongest association (adjusted HR = 1.49; 95% CI, 1.21 to 1.83), followed by musculoskeletal disorders (adjusted HR = 1.13; 95% CI, 0.77 to 1.66). Comorbidities should be routinely evaluated at first visit by occupational health professionals to better manage disability associated with LBP. PMID- 12134533 TI - Occupation and bladder cancer: a population-based, case-control study in Iowa. AB - While considerable efforts have been made to investigate the role of occupation and industry in the risk of bladder cancer, many reported associations have not been consistent, and strong evidence of increased risk is apparent for few occupational groups. To further examine the issue, a large, population-based, case-control study was conducted in the state of Iowa among both men and women. A total of 1452 incident bladder cancer cases and 2434 controls were included in the study. Occupational history was collected from respondents for each job held for 5 years or longer since age 16. Among men, excess risk was observed for industries including plumbing, heating, and air conditioning (odds ratio [OR], = 2.2; 95% confidence interval [CI], 1.0 to 5.0); rubber and plastic products (OR = 3.1; 95% CI, 1.2 to 8.5), motor vehicle parts and supplies (OR = 4.5; 95% CI, 1.2 to 16.5), and occupations including supervisors for transportation and material moving (OR = 6.5; 95% CI, 1.4 to 29.9), material-moving-equipment operators (OR = 1.9; 95% CI, 1.0 to 3.6), automobile mechanics (OR = 1.6; 95% CI, 1.0 to 2.6), painters (OR = 2.7; 95% CI, 1.0 to 7.7), and metal- and plastic-working machine operators (OR = 2.0; 95% CI, 1.1 to 3.4). Among women, significant excess risk was observed for secondary school teachers and record clerks. Housekeepers and butlers and workers in laundering and dry cleaning were also at increased risk. In conclusion, these results suggest that occupational exposures may play a significant role in the risk of bladder cancer. PMID- 12134534 TI - Work-related burns in Washington State, 1994 to 1998. AB - This article describes an investigation of work-related burns in Washington State during 1994-1998. Workers' compensation data were used to describe the general characteristics of burn injuries, estimate industrial claims rates, and compare nonhospitalized and hospitalized burn cases. The completeness of workers' compensation data as a source for surveillance was evaluated. During 1994-1998, a total of 20,213 burn claims were accepted by the workers' compensation system. Hospitalized burn cases represented only 1.5% of burn claims but incurred 55% of the costs. In addition, workers' compensation data underestimated the frequency and rate of burns. Although workers' compensation claims rates decreased during 1994-1998, work-related burns remain a problem in Washington State. Several industries (e.g., roofing, foundries, and aluminum smelting) were identified as priorities for prevention of burn hospitalizations, which incur the greater cost and time loss. PMID- 12134535 TI - [Evaluation of success of mitral valvuloplasty in the early period with cardiopulmonary exercise test]. AB - OBJECTIVE: It was shown that percutaneous balloon mitral valvuloplasty (PMBV) has provided symptomatic improvement in cases with mitral stenosis. The purpose of this study was to investigate the influences of successful PBMV on cardiopulmonary exercise test (CPET) in patients with mitral stenosis early after intervention. METHODS: Twenty-nine patients with mitral stenosis were included in this study. Nineteen patients had undergone PBMV and ten patients were studied as control group. An incremental symptom limited CPET was carried out within the 24 hours before the PMVB procedure and within the five days thereafter. Breath by breath O2 uptake (VO2) and CO2 production (VCO2) were measured in these subjects. RESULTS: The mean mitral valve area (MVA) in the PBMV group before the procedure was 1.2 +/- 0.7 cm2 and the mean pressure gradient (PG) through the mitral valve was 12.63 +/- 4.87 mmHg; after the procedure, the mean MVA was 1.9 +/- 0.3 cm2 and the mean PG was 4.9 +/- 2.3 mmHg. The mean MVA in the control group was 1.4 +/- 0.16 cm2 and the mean PG was 7.2 +/- 3.54 mmHg. In the PBMV group, exercise time was 12.1 +/- 6 min before the procedure and increased to 18.75 +/- 5.5 min after the procedure (p = 0.0001); peak VO2 value rised from 1035 +/- 392 ml/min to 1178 +/- 373 ml/min (p = 0.0001) and VO2 at the anaerobic threshold from 667 +/- 286 ml/min to 772 +/- 268 ml/min (p = 0.006). Peak VO2/HR rised from 10.97 +/ 6.10 ml/min to 12.24 +/- 7.36 ml/min (p = 0.001). No significant difference was observed in the control group. CONCLUSIONS: The results of this study demonstrate that successful PBMV causes evident rise in exercise capacity, so that patients can manage the same exercise levels with lower heart rates and more economic ventilation. PMID- 12134536 TI - [The relationship of the left atrial spontaneous echo contrast integrated backscatter with left atrial appendix function and tissue doppler characteristics]. AB - OBJECTIVE: Integrated backscatter (IBS) is an objective method to measure spontaneous echo contrast (SEC) quantitatively. The purpose of this study was to investigate the relationship between the quantitative values of left atrial SEC measured by using IBS with its qualitative gradation as well as the left atrial appendix (LAA) functions and LAA tissue Doppler characteristics (TD). METHODS: Thirty-two patients (23 female, 9 male) with various diseases undergoing transesophageal echocardiography (TEE) were included. Mean age was 52 + 13 years. Left atrial SEC was graded as no SEC (n = 12), mild (n = 12) or severe SEC (n = 8) in patients underwent TEE. LAA peak emptying and LAA apical and basal TD velocities were measured. LAA ejection fraction was assessed by standard means. RESULTS: Between no SEC and mild SEC groups, left atrium IBS and LAA apical TD values were found statistically significant. But there were no differences between two groups regarding LAA emptying velocities, LAA ejection fraction values and LAA basal TD values. All of these parameters were found statistically different between severe and mild SEC groups. The left atrial IBS values were found to be correlated positively with the qualitative SEC grade. The left atrial IBS values correlated negatively with LAA emptying, apical TD velocities and LAA ejection fraction values. CONCLUSION: Integrated backscatter provides an objective quantitative measure of SEC that correlates well with LAA apical TD velocities and the other parameters of LAA. PMID- 12134537 TI - [Arrhythmia risk and noninvasive markers in hypertensive left ventricular hypertrophy]. AB - OBJECTIVE: Several studies have evidenced that hypertensive patients with severe left ventricular hypertrophy have an increased incidence of malignant ventricular arrhythmia and sudden death. However arrhythmia risk in mild to moderate hypertrophy is uncertain. This study aims to investigate the risk of ventricular arrhythmias in hypertensive patients with mild to moderate hypertrophy and evaluate the role of noninvasive arrhythmia markers and ambulatory blood pressures. METHODS: Ninety-nine hypertensive patients (35 male, mean age 57.3 +/- 9.6) without coronary heart disease were included the study. All subjects underwent an echocardiography for measurement of LV mass index (LVMI) and were classified in two groups; hypertrophic (LVH(+) n:43) and nonhypertrophic (LVH(-) n:56). Ambulatory blood pressure monitoring, 24 hour ECG, signal averaged ECG, and 12 lead ECG were performed in each group seeking to identify the arrhythmogenic risk. RESULTS: Holter ECG showed that 20.1% patients had Lown class II and 12.1% patients had Lown class IVa-IVb arrhythmia (potentially malignant ventricular arrhythmia; PMVA). PMVA incidence was significantly higher in hypertrophic groups (20.9%) compared to nonhypertrophic groups (6.5%) (p < 0.05). Ambulatory systolic and diastolic blood pressures were similar in PMVA(+) and PMVA(-) patients. At least two parameters of ventricular late potentials were significantly higher in LVH(+) group (25.7%) compared to LVH(-) group (4.9%) (p < 0.01). HRV parameters were not different between two groups. QTcd was significantly increased in LVH(+) than in LVH(-) patients (54.1 +/- 16.7 vs. 47.5 +/- 17.7 ms) (p < 0.05) The frequency of PMVA was significantly higher in increased QTcd compared to normal QTcd (24.3%-3.4%; p < 0.01) and LP(+) patients (16.2%) compared to LP(-) patients (8.7%; p < 0.05). CONCLUSION: Our data suggest that hypertension may be associated with high risk of PMVA in patients with mild to moderate LVH particularly in presence of LP and QTcd > 50 ms. QTcd and at least 2 factors of LP were increased in mild to moderate LVH. Arrhythmogenecity does not seem to be related with autonomic dysregulation and ambulatory blood pressure level in hypertensive patients. PMID- 12134538 TI - Should troponin-T be assessed during exercise stress testing in patients with stable angina pectoris? AB - OBJECTIVE: This study was planned to investigate whether or not troponin-T positivity has occurred during exercise stress testing in patients with stable angina pectoris and if yes, its relationship with the severity of the disease. METHODS: One hundred patients with stable angina pectoris who presented with typical chest pain were included in this study. They were subjected to the exercise stress testing according to Bruce protocol. Troponin-T was studied 3 times: immediately before, 6 and 24 hours after the exercise testing. Coronary angiography was performed two hours after the last blood sampling. RESULTS: Exercise stress test was found positive in 67 (67%) and negative in 33 (33%) patients. Coronary artery disease was present in 47 (70.1%) of those with positive and in 17 (51.5%) with negative test results. Troponin-T was negative in all the patients before the stress test. Troponin-T was found positive in readings taken 6 and 24 hours after the test in 4 patients (6.2%) with coronary artery disease. Of these patients, 2 had positive and the remaining 2 had negative stress test results. Troponin-T was found negative in readings after the stress test in all the patients without coronary artery disease. The duration of the exercise stress test was found to be significantly shorter in patients with troponin positivity than their counterparts with negativity (277.5 +/- 81 sec vs. 428.8 +/- 195 sec, p = 0.024). Troponin-T positivity after the stress test was found considerably higher in patients with three-vessel disease (p = 0.021). CONCLUSIONS: Heavy exercises like stress test may severely lead to myocardial damage. The study of post-stress test tropinin T readings, in patients with stable angina pectoris and with negative stress test result, may be of great help in detecting especially the patients with multiple vessel disease. PMID- 12134539 TI - [Proximal anastomotic marker use in coronary artery bypass operations]. AB - Detection and evaluation of aorto-to-saphenous vein anastomosis sites (proximal anastomoses) in patients who had undergone coronary artery bypass surgery are comparatively harder than native coronary orifices during follow-up re angiography procedures. Placement of a radioopaque proximal anastomotic graft marker during coronary artery bypass procedure poses medical and economical advantages in case of postoperative re-angiography during follow-up of these patients. Indication of whether or not to use a proximal anastomotic marker is in general decide on by the operating surgeon. However, coronary angiography is a task of interventional cardiologist. Difference of the teams performing catheterization and the surgical procedure may rise some inconsistencies in terms of requirements for these markers. In order for these dilemmas to be prevented, surgical team should be informed of the complication re-angiography procedure. Proper strategy for the implantation of this technique, which is convenient not only for cardiologist and surgeon but also in economic terms, should be decided on with collaboration of cardiology and cardiovascular surgery teams. In this article, advantages of the proximal anastomotic markers during the postoperative follow-up and re-angiography have been presented with the related literature review. PMID- 12134540 TI - [Hypertension and endothelial dysfunction]. AB - Endothelium; effective in vascular tonus, blood tension, blood flow and coagulation system is the biggest endocrine organ with 1800 gr. weight in human body. The endothelium releases nitric oxide, which maintains vascular integrity. In endothelial dysfunction the balance between releasing vasodilatatory and vasoconstrictory factors is changed. In the presence of essential hypertension defection in endothelium-dependent vasodilation and increasing sensitivity to vasoconstrictors could increase vascular resistance. Although some beneficial results with angiotensin converting enzyme inhibitors, carvedilol, angiotensin-II and endothelin receptor antagonists were reported in patients with endothelial dysfunction the effectiveness of these drugs is still debate. Also the decreasing of arterial pressure was not always shown to be parallel to the restoration of endothelial dysfunction. In the future according to genetic studies, the increased use of nitric oxide releasing genes therapy will have a more effective role in regression of endothelial dysfunction. PMID- 12134541 TI - Differential-diagnostic algorithm as a teaching and diagnostic method in cardiological practice. PMID- 12134542 TI - [Radiofrequency catheter ablation of the atrioventricular nodal reentrant tachycardia]. PMID- 12134543 TI - [Primary coronary angioplasty and stent implantation for an acute anterior myocardial infarction in the early postoperative period of laparoscopic cholecystectomy: A case report]. PMID- 12134544 TI - [Two stage surgery approach to a patient with ascending aorta aneurysm and coarctation of the aorta: A case report]. PMID- 12134546 TI - [A case of single ventricle attained adulthood without operation]. PMID- 12134545 TI - [The effects of preoperative use of ACE inhibitors on renal function after open heart surgery]. PMID- 12134547 TI - [A female patient with anemia and cardiac myxoma]. PMID- 12134548 TI - [Are there effects due to the existence of coronary collateral circulation on left ventricular function in patients with coronary artery disease?]. AB - OBJECTIVE: The role of coronary collateral circulation (CCC) on the improvement of left ventricular function in coronary artery disease is controversial. The aim of this study is to investigate the effect of CCC on left ventricular function in patients with ischaemic heart disease. METHODS: Accordingly, 76 patients (39 female, 37 male, mean age--61 +/- 17 years) who had single vessel disease with > 85% narrowing in left anterior descending coronary artery were enrolled in this study. Coronary collateral circulation was determined according to the Rentrop classification (Class 0 = no collateral circulation; class 1 = small branches of occluded vessel fill with CCC; class 2 = epicardial segment of the occluded vessel partially fills with CCC; class 3 = epicardial segment of the occluded vessel totally fills with CCC). Left ventricular function was assessed with echocardiography and left ventricular regional wall motion score (0 = normokinetic; 1 = hypokinetic; 2 = akinetic; 3 = dyskinetic; 4 = aneurysmatic). Rentrop classification of the patients were compared with left ventricular regional wall motion scores and ejection fractions. RESULTS: Twenty one of 76 patients had no collateral circulation. The regional wall motion score of class 0 patients was similar with that of patients with CCC (class 1,2,3) (2.28 +/- 2.1 vs 3.39 +/- 2.1, p > 0.05). Particularly, the regional wall motion score was positively correlated with Rentrop classes (p < 0.05). Class 3 patients had the highest wall motion score (4.24 +/- 2.5, p < 0.05). Patients with and without CCC had similar left ventricular ejection fractions (49 +/- 11 vs 46 +/- 17, p > 0.05). CONCLUSION: This study showed that development of CCC has no preventive effect on left ventricular functions in patients with coronary artery disease. Interestingly as the stage of CCC increases left ventricular function worsens. It may be the result of the fact that patients with well developed CCC have more severe coronary artery disease. PMID- 12134550 TI - Discussing general safety concerns in the laboratory. PMID- 12134549 TI - [Coronary bypass reoperations: evaluation of 104 cases]. AB - BACKGROUND: Repetitive procedures usually take place in the natural course of coronary heart disease. The aim of this study was to evaluate the risk factors, which affect coronary bypass reoperations, and to compare them with the postoperative results of the coronary first operations and the reoperations. METHODS: Between January 1995 and January 2000, coronary reoperations were performed in 104 cases (Coronary reoperations group) by the same surgical team. Ninety-nine of them were the first, 3 were the second and 2 were the third reoperations in this group. At the same period of time, 3609 patients underwent coronary bypass procedure as the first operation (Coronary 1. operation group). Eighty-seven patients were male (83.65%), 17 were female (16.35%) and the mean age was 60.82 +/- 9.49 in reoperation group; while among 2916 patients 2223 were male (80.8%), 693 were female (19.2%) and the mean age was 60.37 +/- 9.58 in the first operation group. RESULTS: Incidence of prolonged ventilation (p = 0.0001), renal dysfunction requiring dialysis (p = 0.01), need for intraaortic balloon pump (p = 0.0001) and prolonged intensive care unit (p = 0.01) and hospital stay (p = 0.01) were significantly higher in reoperation group. The mortality rate was 9.62% in the reoperation group while it was 2.2% in the first operation group (p = 0.0001). CONCLUSIONS: The high morbidity and mortality of coronary bypass reoperations can be reduced to acceptable levels accordingly with early therapy prior to ventricular dysfunction and clinical deterioration that will improve the outcome in these patients. PMID- 12134551 TI - Laboratory diagnosis of autoimmune diseases. PMID- 12134552 TI - The community phlebotomy response plan: how does your facility measure up? PMID- 12134553 TI - CMS launches survey of certificate of waiver laboratories. PMID- 12134554 TI - Strengthening the supply chain. PMID- 12134555 TI - EMTALA regs: you may be surprised at proposed changes to requirements. AB - The proposed rule for new Emergency Medical Treatment and Labor Act (EMTALA) regulations from the Centers for Medicare & Medicaid Services doesn't contain major changes, but it would lessen the burden of compliance. The final rule will be published later this year. A distinction is made between "dedicated EDs" and off-campus sites that typically don't provide emergency services. EMTALA would no longer apply to patients with scheduled outpatient visits. Prior authorization requirements are a violation of federal law, which can help you obtain reimbursement for screening examinations required under EMTALA. PMID- 12134556 TI - Collect copays on the 'back end' of the visit. AB - Patients should be directed to a separate area after the ED visit for collection of copay payments. Discuss copays at registration, but collect them only at discharge. Train staff in how to avoid EMTALA violations. Patients are more willing to give copays if they are satisfied with the care they received. PMID- 12134557 TI - ED managers: don't stop using template charting! PMID- 12134558 TI - It's time to give up costly template charts. PMID- 12134559 TI - Report: ED visits are on the rise. PMID- 12134560 TI - Effects of institutional services and characteristics on use of postacute care settings. AB - Postacute care under Medicare may be provided in several institutional settings, some of which function as substitutes for each other, including rehabilitation care in either specialty hospitals and hospital units excluded from Medicare PPS or skilled nursing facility care. This study uses Medicare billing data to examine institution-based postacute care utilization for beneficiaries' hospital episodes occurring from February 1, 1992 through June 30, 1992. Data on inpatient and postacute Part A services used by Medicare inpatients receiving care in rehabilitation facilities or skilled nursing facilities are presented. The results indicate that, even after casemix is controlled for, inpatient stay treatment variables affect postacute care utilization patterns. Nonclinical factors such as age and sex also affect choice of postacute care site. PMID- 12134561 TI - The regulation of outpatient services: an analysis of the interaction between HCFA and Medicare providers. AB - The Federal Register is used as a historical record documenting the interaction between the Health Care Financing Administration (HCFA) and health care providers in the regulation of outpatient surgery services to Medicare patients. A content analysis of the Federal Register reveals that HCFA is more likely to accommodate requests for clarification, for shifting services among payment levels, and for adding or deleting services from coverage than for altering payment methods. These findings can be used by health care providers to develop strategies for coping with the expansion of prospective payment to all outpatient services. PMID- 12134562 TI - Hospital restructuring and nursing staff well-being: the role of personal resources. AB - This study examined the role played by two personal resources, job mobility options and financial resources, among nursing staff during a period of major hospital restructuring and downsizing. Data were collected from 1362 staff nurses using questionnaires. Personal resources were hypothesized to have direct and indirect effects on job satisfaction and psychological well-being in a model of hospital restructuring and its effects. The model included four variables: extent of hospital restructuring, future threats to the workplace, job satisfaction, and psychosomatic symptoms. LISREL analyses indicated that financial resources reduced perceptions of future workplace threats and psychosomatic symptoms while job mobility options were associated with higher levels of job satisfaction. PMID- 12134563 TI - Modern medical genetics: a systems approach. PMID- 12134564 TI - Zero-sum politics, the Herbert thesis, and the Ryan White CARE Act: lessons learned from the local side of AIDS. AB - This study examines the dynamics of grass-roots decision-making processes involved in the implementation of the Ryan White CARE Act. Providing social services to persons with HIV/AIDS, the CARE act requires participation of all relevant groups, including representatives of the HIV/AIDS and gay communities. Decision-making behavior is explored by applying a political (zero-sum) model and a bureaucratic (the Herbert Thesis) model. Using qualitative research techniques, the Kern County (California) Consortium is used as a case study. Findings shed light on the decision-making behavior of social service organizations characterized by intense advocacy and structured on the basis of volunteerism and non-hierarchical relationships. Findings affirm bureaucratic behavior predicted by the Herbert Thesis and also discern factors which seem to trigger more conflictual zero-sum behavior. PMID- 12134565 TI - Talking to patients about aerobic exercise for disease prevention: an educational exercise for medical students. AB - Numerous data demonstrate the importance of physical activity in reducing obesity and cardiovascular mortality and morbidity. Research demonstrates the beneficial impact of physician counseling on health promoting behaviors. Unfortunately, few physicians or medical students receive formal training in exercise counseling. We describe an educational activity used to provide medical students with the tools needed to begin to engage patients in activity counseling. PMID- 12134566 TI - Diagnosis and management of lipoprotein abnormalities. AB - Abnormal lipid and lipoprotein cholesterol values have been defined as a low density lipoprotein (LDL) cholesterol (C) value of 160 mg/dL (4.1 mmol/L) or greater, a high-density lipoprotein (HDL) C value less than 40 mg/dL (1.0 mmol/L), triglycerides (TG) 150 mg/dL (1.7 mmol/L) or greater, and a lipoprotein (a) (Lp(a)) of 30 mg/dl or greater. Such values all increase coronary heart disease (CHD) risk. The National Cholesterol Education Program Adult Treatment Panel III guidelines continue to focus on optimizing LDL-C values (established as < 100 mg/dL or 2.6 mmol/L), especially in those with established CHD, diabetes, or a 10-year CHD risk over 20%. Dietary saturated fat (< 7% of calories) and cholesterol (< 200 mg/day) restriction, and the use of 3-hydroxy-3 methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors are the mainstays of treatment in this regard. Such treatment substantially reduces CHD risk. Severe hypertriglyceridemia (> 1000 mg/dL or 11.0 mmol/L) is associated with pancreatitis, and fat restriction, control of glucose, and fibrate therapy are indicated in such patients. Niacin is currently the most effective agent for lowering Lp(a) and raising HDL-C. Current recommendations for treatment by diet and drugs are outlined. PMID- 12134567 TI - Folate, homocysteine, and neurological function. AB - The study of different neurological problems, including stroke, Alzheimer's disease (AD), and depression, has propelled a greater interest in interrelationships among folate, homocysteine, and neurological function. Specifically, low folate status is a suspected risk factor for depression that also results in an increase in circulating levels of the sulfur amino acid homocysteine. Homocysteine has emerged as an independent risk factor for stroke, and recent studies suggest that vascular disease affecting the brain and Alzheimer's disease may result together in senile dementia. The relationship between stroke and AD was at first interpreted as coincidence, given the pathologic distinctions between the two diseases. However, the connection is now hypothesized to reflect some common pathogenic factors involving folate, homocysteine, or both. It remains unclear whether there is a causal relationship between neurological dysfunction in either condition with folate or homocysteine. Nevertheless, since improvement of folate status lowers homocysteine levels, the hypothesis that folate supplementation may lower the risk of several important health consequences of aging, including various forms of neuropsychiatric dysfunction, is worthy of current intensive exploration. PMID- 12134568 TI - PACE: evaluating its use in the primary care setting. PMID- 12134569 TI - Physical activity: simple steps to getting your daily dose. PMID- 12134570 TI - Physical activity: eliminate barriers & gain motivation. PMID- 12134572 TI - Just do it--talk to your patients about exercise. PMID- 12134571 TI - Botulism used as biological warfare. PMID- 12134573 TI - Establishing an effective framework for physical activity counseling in primary care settings. AB - This article examines the role of the primary care physician in providing physical activity counseling and proposes a framework for maximizing the effectiveness of this activity. Many studies have been conducted to evaluate the effectiveness of physical activity counseling in the primary care setting. Unfortunately, the results of most of these studies have not been encouraging as there has not been compelling evidence that brief physical activity counseling for the general population results in long-term changes in physical activity behaviors. In contrast, the evidence of health improvements in patients who are successful in achieving levels of physical activity consistent with the Surgeon General's recommendations is compelling and should serve as the source of motivation as we seek more effective paradigms for physical activity counseling. The PACE (Patient-Centered Assessment and Counseling for Exercise and Nutrition) Program is presented as an evidenced-based and effective framework for effective physical activity counseling. By adopting a patient-centered approach to counseling and by tailoring the message to the readiness of the patient to take action, it is possible to engage in physical activity counseling in a cost- and time-effective manner. This article provides specific counseling strategies for addressing patients at various stages of readiness to engage in behavior change regarding physical activity. PMID- 12134574 TI - The case for extending storage and secretion functions of human mast cell granules to include synthesis. AB - Ultrastructural studies using standard procedures have for years indicated close associations of ribosomes and secretory granules in human mast cells. These descriptive studies have informed new studies, using established and new ultrastructural methods based on different principles, designed to investigate the possible role of RNA metabolism in secretory granules of human mast cells. In aggregate, these studies indicate human mast cell secretory granule associations with ribosomes, the protein synthetic machine of cells, with ribosomal proteins, with RNA, with poly(A)-positive mRNA and with various long-lived, or short-lived, uridine-rich, and poly(A)-poor RNA species with key roles in RNA processing and splicing. These studies indicate that secretory-storage granules in human mast cells may also be synthetic granules. PMID- 12134575 TI - Radioautographology general and special. AB - A new concept, termed "radioautographology" is advocated and its contents are reviewed. This term is the coinage synthesized from "radioautography" and "(o)logy", expressing a new science derived from radioautography. The concept of radioautographology (RAGology) is a science to localize the radioactive substances in the biological structure of the objects and to analyze and to study the significance of these substances in the biological structure. On the other hand, the old term radioautography (RAG) or autoradiography (ARG) is the technique to demonstrate the pattern of localization of various radiolabeled compounds in biological specimens. The specimens used in biology and medicine are cells and tissues. They are fixed, sectioned and made contact with the radioautographic emulsions, exposed and developed to produce metallic silver grains. Such specimens are designated as radioautographs (or autoradiographs) and the patterns of pictures made of silver grains are named radioautograms. Those people who produced radioautographs were formerly named radioautographers (or autoradiographers) who were only technicians, while those who study RAGology are not technicians but scientists and should be called as radioautographologists. The science of radioautographology was developed in the 20th century and can be divided into two parts, general radioautographology and special radioautographology, as most natural sciences usually can. The general radioautographology is the technology of RAG which consists of 3 fields of sciences, physics concerning radioactivity, histochemistry treating the cells and tissues and photochemistry dealing with the photographic emulsions. The special radioautographology, on the other hand, consists of applications of general radioautographology to various biological and medical sciences. The applications can be classified into several scientific fields, i.e., cellular molecular biology, anatomy, histology, embryology, pathology and pharmacology. Studies carried out in our laboratory were summarized and reviewed. The results obtained from the technology includes 4-dimensional structures of the organs taking the time dimension into account by labeling cells and localizing the sites of incorporation, synthesis, discharge of the labeled compounds in connection with the time lapse and aging of animals. All the results obtained from such applications should be systematized as a new filed of science in the future in the 21st century. PMID- 12134576 TI - Debridement. The key initial first step in wound healing. AB - Before addressing a wound, whether it is chronic or acute, clinicians must thoroughly assess the wound and the patient. An acute wound in a patient with normal blood flow and good medical and nutritional condition should go on to heal if appropriate care is given. This means that the wound has to be debrided adequately, dressed, and closed when appropriate. Getting back to healthy tissue is the key. In chronic wounds, healing is more difficult because the etiology of the wound is harder to determine, and the measures to reverse the medical abnormalities are often complex. When these have been sorted out and addressed, however, debridement again plays the key role. It converts the chronic wound into an acute wound so that it can then progress through the normal stages of healing. The key is for clinicians to be aggressive and not let concerns about the residual defect limit debridement. Subsequent healing then can be achieved by use of wound-healing adjuncts such as the V.A.C. device, hyperbaric oxygen, skin substitutes, growth factor, or plastic surgical techniques. PMID- 12134577 TI - Adjuncts to preparing wounds for closure: hyperbaric oxygen, growth factors, skin substitutes, negative pressure wound therapy (vacuum-assisted closure). AB - Achieving closure in a chronic wound requires provision of adequate oxygen delivery to the tissue, adequate protein and other nutritional factors, a moist environment, an appropriate inflammatory milieu, debridement, and correction of contributing medical diagnoses. In some patients, these conditions are achieved easily, whereas in others, greater effort is required. Adjunctive treatments, including HBO2, growth factors, skin substitutes, and negative-pressure wound therapy (e.g., V.A.C.) can provide the proper conditions for healing in appropriately selected patients. PMID- 12134578 TI - Vascular reconstructive options in the ischemic foot. AB - Aggressive evaluation and treatment of the patient suffering from lower-extremity ischemia is critical. Vascular reconstruction can be performed to enhance healing and to decrease the incidence of major limb amputation and therefore return these patients to active and productive life. PMID- 12134579 TI - Medical and surgical considerations in patients with vasculitis and Raynaud's syndrome. AB - Vascular problems in the foot are certainly common when one considers only arteriosclerosis on the macrovascular scale. The primary cause of ischemia of the foot undoubtedly is primary arteriosclerosis, whether related to smoking, diabetes, renal failure, or other causes. Vasculitis and vasospasm, in their many forms, are distinctly unusual as a cause of ischemia of the foot. These entities, nonetheless, can cause significant problems from the standpoint of symptoms and even ulceration or gangrene of the foot. This article addresses the pathophysiology of vasculitis and vasospastic problems in the foot and their management. PMID- 12134582 TI - Soft tissue reconstruction for the heel and plantar foot. AB - Soft tissue resurfacing after the ravages of injury or disease continues to be a formidable problem. A thorough understanding of the biomechanics of ambulation and the way in which weight is distributed over the sole of the foot is key to successful outcomes. Soft tissue repair has evolved from the simple filling of holes to a much more sophisticated approach. A plantar-grade foot that distributes the patient's weight in a uniform manner should be the goal. If this can be done without increasing the energy expenditure of ambulation, then surgeons certainly have done our patients an excellent service. Surgeons must not overlook the tendon imbalances and bony deformities that can accompany the effects of disease or trauma. The more complex soft tissue repair may not always be in the patient's best interests. Surgeons should use the simplest approach that can restore the limb to as close to its preinjury level of function as possible. A coordinated approach with plastic surgery, orthopedic surgery, vascular surgery, and podiatric surgery should be the standard of care for patients requiring plantar soft tissue repair. PMID- 12134580 TI - Orthopedic considerations in wound healing of the lower extremity. AB - The advancing technologies available to the orthopedist are a necessary adjunct to heal wounds about the foot and ankle successfully. Successful wound healing should be viewed as a multidisciplinary approach, using the strengths of each discipline. Critical to an integrated method is communication preoperatively between the orthopedist and the plastic surgeon to create a streamlined process. This leads not only to improved rates of wound healing, but also to improved functional outcomes while reducing risks for recurrence. PMID- 12134581 TI - The safest surgical incisions and amputations applying the angiosome principles and using the Doppler to assess the arterial-arterial connections of the foot and ankle. AB - Knowing the arterial anatomy of the foot and ankle in addition to understanding the angiosome concept provides the basis for careful and safe planning of incisions. The Doppler allows the surgeon to map out the actual vascular anatomy that exists preoperatively and therefore allows for appropriate adjustment to the planned incisions. If the vascular flow is inadequate for the planned surgery, then the vascular surgeon has to intervene to improve the existing blood flow. If the vascular tree is so compromised that successful revascularization of the affected angiosome is impossible, then the revascularization is likely to fail. Serious consideration to a below-knee amputation should then enter the decision tree at that time. Most of the time, however, the blood flow is adequate or can be sufficiently augmented with vascular surgery. The foot and ankle surgeon can then perform preoperative mapping of the arterial blood supply with the Doppler. By making the necessary adjustments to the planned incision, surgery can proceed safely with uneventful healing. PMID- 12134583 TI - Glabrous skin grafts for plantar defects. AB - The principle of reconstructing like to like has been a long-standing and useful concept for plastic surgeons. One arena in which this concept has not been put to its full use is that of reconstructing soft tissue deficits of the sole of the foot. Most commonly, plantar defects that are to be skin grafted are reconstructed with split- or full-thickness, nonglabrous skin grafts. Nonglabrous skin grafts have significant disadvantages when used for reconstruction of plantar defects. These include painful hyperkeratotic build up at the periphery of the skin grafts, craters, contractures, and tight subgraft fibrosis. Glabrous skin grafting has been applied widely for coverage of smaller defects in the hand and has yielded superior results with improved function and sensation, more normalcy of appearance, and increase durability. The concept of reconstructing plantar defects by this method has probably been impeded by the vague and erroneous, but broadly held, belief that donor-site healing in the foot would be problematic, that is, significant potential for excessive scarring, pain, and functional deficit. The long-standing use of glabrous skin grafts for plantar defects in this unit, however, confirms the desirability, functional advantage, and minimal morbidity of this technique. PMID- 12134584 TI - Soft tissue coverage options for dorsal foot wounds. AB - The soft tissue of dorsum of the foot consists of a thin pliable surface that allows for significant excursion and tendon gliding. Reconstructive options must preserve these important functions and allow for reasonable contour so the patient may wear a shoe postoperatively. Special attention must be given to the mechanism of injury and overlying pathophysiology involved with each wound. Local flaps can provide adequate wound coverage in settings in which the vasculature and subcutaneous structures have been preserved. In wounds in which the regional vascularity is compromised or in which tendon and bone have been lost, a free tissue transfer can provide for more substantial coverage. The multiple options available with free-tissue transfer allows for the possibility of composite tissue transfer, including vascularized bone or tendon, and the ability to create a sensate flap with excellent contour. PMID- 12134585 TI - Soft tissue coverage options for ankle wounds. AB - Soft tissue deficiencies of the ankle are caused by several mechanisms, such as trauma, tumor, and infection. Compounding the reconstructive problems is that soft tissue problems often present in patients who have underlying diseases such as peripheral vascular disease, diabetes or both. For example, a 65-year-old person with diabetes who smokes two packs of cigarettes per day sustains an ankle fracture. After undergoing open reduction and internal fixation of the fracture, there is subsequent wound behiscence over the patient's fibular plate. The wound edges cannot be reapproximated, and there is loss of soft tissue. What should treatment be for this soft tissue problem? Another example is a 45-year-old rheumatoid patient who takes 20 mg of steroids a day and undergoes posterior tibial tendon repair after rupture. One month after surgery, the surgical wound dehisces, resulting in exposure of the tendon repair. What is the approach for adequate and effective soft tissue treatment? The purpose of this article is to address such complex problems and to provide an algorithm for soft tissue reconstruction of the ankle. PMID- 12134586 TI - Soft tissue coverage of the extremely traumatized foot and ankle. AB - The development of microsurgery and the expansion of plastic surgery techniques have led to a significant increase of surgical options for the salvage of the lower extremity. The traditional methods still have a role, but many authors have demonstrated a superiority of free and sophisticated regional flaps. This article gives an overview of treatment algorithms and surgical options. A therapeutic goal for the surgeon is to select the appropriate procedure with respect to the patient's medical condition and rehabilitation potential, the defect, and the surgeon's technical skills to achieve durable, permanent, pain-free, and functionally and aesthetically satisfying defect coverage. PMID- 12134587 TI - [To err is human: building a safer health system]. PMID- 12134588 TI - [Introduction to systematic analysis of anesthesia incidents]. PMID- 12134589 TI - [Can sedation be considered anesthesia?]. PMID- 12134590 TI - [Does the time between preoperative interruption of aspirin intake and operation influence postoperative blood loss and transfusion requirement in coronary artery bypass graft?]. AB - OBJECTIVE: Impact of the interval between interruption of aspirin intake and surgery on postoperative bleeding and transfusion in coronary artery bypass graft (CABG), with extracorporeal circulation (ECC). STUDY DESIGN: Retrospective study. PATIENTS AND METHODS: Four hundred and twelve patients having undergone CABG were retrospectively reviewed. Three groups were evaluated according to the length of the interval defined above: Group I (< 3 days), Group II (3-7 days), Group III (> 7 days or without aspirin intake). Postoperative blood loss at 3rd, 6th, 12th, and 24th hour and transfusion requirements were assessed for the 3 groups. Aprotinin (low dose, 2 M KIU) was systematically included in the priming of the ECC circuit. RESULTS: There were no significant differences among groups for weight, size, duration of ECC, and number of bypasses. No significant correlation was noted among the 3 groups for postoperative blood loss and transfusion. Multivariate analysis showed that factors associated to a higher risk of excessive bleeding were ECC duration and number of arterial grafts. Factors associated with a higher risk of transfusion were: emergency, minimum patient temperature during ECC, weight and preoperative haemoglobin level. Aspirine intake was not associated with an increase of bleeding or transfusion. CONCLUSION: Our study showed that in our practice using systematic low dose of aprotinin when priming the ECC circuit, aspirin did not significantly increase bleeding or transfusion requirements in CABG with ECC, whatever the interval between interruption of aspirin intake and surgery. Consequently, in our practice, aspirin intake is interrupted on hospitalisation, one day before surgery. PMID- 12134591 TI - [Laparoscopic adrenalectomy for pheochromocytoma. Perioperative blockade with urapidil]. AB - OBJECTIVE: To investigate the effects of coeliosurgery by catecholamine assays and the use of urapidil in the management of phaeochromocytoma. STUDY DESIGN: Prospective cohort study. PATIENTS: Nine consecutive patients from April 1997 to April 2001. METHODS: Urapidil (250 mg.j-1) was administered by continuous intravenous infusion three days before surgery and continued throughout anaesthesia. Plasma catecholamine concentrations were measured before surgery, after induction of anaesthesia, during insufflation, after adrenalectomy and in the recovery room. Haemodynamic disorders were treated by nicardipine +/- esmolol bolus doses. RESULTS: Creation of pneumoperitoneum and adrenal gland manipulations resulted in significant catecholamine releases associated with hypertension in five and eight patients respectively. Preventive urapidil use enabled easy control of blood pressure variations by additive antihypertensive drugs. CONCLUSION: Perioperative alpha 1 blockade by urapidil enables an effective and easy control of acute preoperative haemodynamic changes. PMID- 12134592 TI - [Contraction-relaxation coupling of skeletal muscle in patients susceptible to malignant hyperthermia]. AB - OBJECTIVE: To assess whether halothane exposure could influence contraction relaxation coupling of human skeletal muscle with malignant hyperthermia susceptibility. STUDY DESIGNED: Laboratory investigation. MATERIAL AND METHODS: Muscle biopsies from 14 patients, including six classified as susceptible to MH (MHS) and eight as classified as non-susceptible (MHN) according to criteria of the European MH group. Mechanical parameters of strips were obtained before and after 3 vol% halothane exposure. The contraction and relaxation parameters were measured under isotonic and isometric conditions: maximum shortening and lengthening velocities (respectively maxVc and maxVr); peak of the positive (+dP/dtmax) and negative (-dP/dtmax) twitch tension derivative; ratio R1 = maxVc/maxVr and ratio R2 = (+dP/dtmax) (-dp/dtmax). RESULTS: In MHN muscle, halothane markedly increased maxVc and maxVr, so that the ratio R1 was unchanged. Both +dP/dtmax and -dP/dtmax increased such that the ratio R2 did not vary. In MHS muscle, halothane induced a significant decrease in maxVr (p < 0.05) without changes in maxVc, so that the ratio R1 increased significantly. +dP/dtmax remained unchanged whereas -dP/dtmax decreased significantly; the ratio R2 increased (p < 0.05). CONCLUSION: Our results indicated that, in MHN muscle the contractility property is improved with halothane exposure. In MHS muscle, halothane caused an impairment of relaxation. The mechanical abnormalities observed in this study might be related to sarcoplasmic reticulum dysfunction in MH diseases. PMID- 12134593 TI - [Sedation induced by midazolam in intensive care: pharmacologic and pharmacokinetic aspects]. AB - OBJECTIVE: Review on midazolam in order to optimize drug utilisation and therapeutic monitoring. DATA SOURCES: Research of English or French articles published until August 2001, using Medline database. The key words were: midazolam, pharmacokinetics, pharmacodynamic, sedation, drug interaction. STUDY SELECTION: Original articles, clinical cases and letters to the Editor were selected. Animal studies were excluded. DATA EXTRACTION: The articles were analysed according to their interest in midazolam clinical practice. DATA SYNTHESIS: Midazolam is a benzodiazepine widely used in intensive care unit, as a sedative, anxiety-relieving, and amnesic drug. Midazolam could be used in patients with cardiac, or respiratory failure, and in neurosurgery. A great interindividual variability on pharmacokinetic and pharmacodynamic response was observed. In intensive care patients, elimination half-life is known to be widely increased. Midazolam is metabolised by hepatic microsomes. The major metabolite is the 1-hydroxymidazolam, which is pharmacologically active. A prolonged sedation due to an accumulation of conjugated metabolite was observed in renal failure patients. Enzymatic inductors or inhibitors could influence pharmacokinetics and pharmacodynamic effects of midazolam. CONCLUSION: According to midazolam pharmacokinetic and pharmacodynamic variability, an individual dosage adjustment is essential for long-term sedation. Target controlled sedation could be a mean to limit the variability and to reach quickly the pharmacodynamic effect. PMID- 12134594 TI - [The pharmacology of cannabinoid derivatives: are there applications to treatment of pain?]. AB - OBJECTIVE: To present the cannabinoid system together with recent findings on the pharmacology of these compounds in the treatment of pain. DATA SOURCES: Search through Medline database of articles published in French and English since 1966. Also use of other publications such as books on cannabis. STUDY SELECTION: All the relevant documents within the theme of this review were used. DATA EXTRACTION: All the data linked to the present topic were searched. DATA SYNTHESIS: Recent advances have dramatically increased our understanding of cannabinoid pharmacology. The psychoactive constituents of Cannabis sativa have been isolated, synthetic cannabinoids described and an endocannabinoid system identified, together with its component receptors and ligands. Strong laboratory evidence now underwrites anecdotal claims of cannabinoid analgesia in inflammatory and neuropathic pain. Sites of analgesic action have been identified in brain, spinal cord and the periphery, with the latter two presenting attractive targets for divorcing the analgesic and psychotrophic effects of cannabinoids. Clinical trials are now required, but are hindered by a paucity of cannabinoids of suitable bioavailability and therapeutic ratio. CONCLUSION: The cannabinoid system is a major target in the treatment of pain and its therapeutic potential should be assessed in the near future by the performance of new clinical trials. PMID- 12134595 TI - [How to investigate and analyze clinical incidents: a clinical risk protocol in association with litigation and risk management]. PMID- 12134596 TI - [Anesthesia for cesarean section in a patient with McArdle disease and hereditary dilated cardiomyopathy]. AB - A caesarean section was indicated in a 29-year-old parturient affected by a muscular deficit in myophosphorylase responsible for a type V glycogen storage disease (McArdle disease). This metabolic myopathy had been diagnosed two years previously, whereas the patient already suffered from a hereditary form of dilated cardiomyopathy. The muscular disease was invalidating on the functional level with exercise intolerance. The cardiopathy was little symptomatic but the dysfunction of the left ventricle worsened during the pregnancy with an ejection fraction calculated to 43%. In this case, we report the realization of a general anaesthesia in a patient who had epidural anaesthesia for a previous caesarean section. PMID- 12134597 TI - [Factor X deficiency and pregnancy]. AB - Factor X deficiency is one of the rarest inherited coagulation disorders. It is an autosomal recessive inherited disease. In its homozygous form factor X deficiency has an estimated prevalence of 1: 500,000. However in its heterozygous form it has an estimated frequency of 1: 500 to 1: 2000. Pregnancy in women with congenital factor X deficiency has been associated with adverse foetal outcomes. We report a case of pregnancy in a woman with factor X deficiency. She was treated early during labour with prophylactic replacement of prothrombin complex concentrates (Kaskadil). An initial infusion of 40 UI.kg-1 of factor X was followed by 20 UI.kg-1 every 24 hours during three days. During labour and peripartum maternal coagulation was screened. She delivered a healthy baby at 33 weeks of gestation. No episode of abnormal bleeding was observed. Therefore in this case, prophylactic therapy using prothrombin complex concentrates during labour and delivery did prevent severe haemorrhages. PMID- 12134598 TI - [Severe hypophosphatemia and grave head trauma]. AB - The authors report the case of an 18-year-old man with polytrauma, who died at the third day of its accident from brain death and who presented during his stay in the intensive care unit, a deep hypophosphatemia. Two physiopathologic mechanisms were presumed: increase of renal losses and intracellular transfer of phosphorus. Consequences, as well as the indications and the therapeutic modalities of hypophosphatemia are discussed. PMID- 12134599 TI - [Spontaneous thrombosis of an iatrogenic pseudoaneurysm of the arterial brachiocephalic trunk following central venous catheterization]. AB - Following the unsuccessful puncture of the right subclavian vein during central catheterism, a 80-year-old women developed a pseudoaneurysm on the external face of the brachiocephalic artery. Her symptomatology and haemodynamic status having remained steady, the patient was closely observed. The pseudoaneurysm itself thrombosed spontaneously and the foreseen endovascular procedure doesn't have been achieved. A review of the literature has been done concerning the venous catheterism complications and the pseudoaneurysm treatment. PMID- 12134600 TI - [ARDS as an unusual presentation of bronchiolitis obliterans organizing pneumonia]. AB - We report the case of a 46-year-old patient with liver transplantation who developed an acute respiratory distress syndrome (ARDS). The commonly associated clinical disorders, those associated with direct injury to the lung and those that cause indirect lung injury in the setting of a systemic process, were not responsible for the clinical picture. Finally, because of progressive clinical deterioration, an open-lung biopsy was performed and revealed a bronchiolitis obliterans with organizing pneumonia (BOOP). Physicians should be aware of this rare aetiology of ARDS. PMID- 12134601 TI - [A new cause of postoperative confusion syndrome: nefopam]. AB - We report a case of nefopam intoxication in a severely malnourished postoperative intensive care patient with profound hypoprotidemia. Nefopam induced agitation and confusion associated with anticholinergic symptoms such as tremor, hypertonia, mydriasis, and tachycardia. All these symptoms disappeared after withdrawal of the drug. Nefopam should be considered as a possible cause of confusion in postoperative patients. PMID- 12134602 TI - [Hyperosmolar coma and cerebral thrombophlebitis]. PMID- 12134603 TI - [Necrotizing fasciitis: urgent diagnosis]. PMID- 12134604 TI - [Rectus sheath block: "do not forget"]. PMID- 12134605 TI - [Atlanto-occipital luxation: diagnostic problems in a case with few symptoms]. PMID- 12134606 TI - [Martial treatment, postpartum erythropoietin]. PMID- 12134607 TI - [Management of the anesthetic allergic patient]. PMID- 12134608 TI - The early diagnosis and prevention of gastrointestinal cancer: problems and promises. AB - Early detection and prevention of gastrointestinal cancer involves screening and surveillance. The efficacy of early detection tests should be subjected to well designed studies. Clinical recommendation should be made after evaluation of new therapeutic approaches that will be based on our understanding of the molecular and cellular biology and the genetics of gastrointestinal premalignancy and cancer. Chemoprotection and alterations in diet and environment are areas that offer much promise and require clinical evaluation. PMID- 12134609 TI - Cellular and molecular mechanisms of carcinogenesis. AB - Our understanding of the cellular and molecular mechanisms of cancer of the gastrointestinal tract has increased dramatically over the last several decades. We are identifying new players in the pathways toward cancer with increasing frequency. In addition, we have come to understand that no single pathway acts by itself; in vivo, the effects are combinatorial. As new and better cell culture and animal models of carcinogenesis arise, our knowledge will continue to grow. As we learn more, we will be able to translate the results of our research into new and better techniques for the diagnosis and treatment of gastrointestinal cancers. PMID- 12134610 TI - Histologic precursors of gastrointestinal tract malignancy. AB - Precursor lesions in the GIT include flat dysplasias, adenomas, dysplasia superimposed on nonneoplastic polyps, endocrine cell dysplasia, ACF, and condyloma accuminatum. Interobserver variability can be a problem in reporting dysplasia, and ancillary techniques including flow cytometry, image analysis, proliferation markers, and examination for p53 expression can help in this task. Squamous dysplasia seen in the esophagus and anus is graded on either a two tiered or three-tiered system largely based on the extent of mucosal involvement. Glandular dysplasia is morphologically similar whether seen as an adenomatous polyp or within the setting of Barrett's esophagus, atrophic gastritis, or idiopathic inflammatory bowel disease. The distinction between LGD and HGD in glandular mucosa is based on the severity of cytologic and architectural distortion. Type I dysplasia is the classic adenomatous pattern seen most commonly and recognized by the presence of elongate hyperchromatic stratified nuclei. Type II, the nonadenomatous variant, contains vesicular nuclei and alteration in nuclear size and shape. Nonantral endocrine dysplasia in the stomach is seen in the setting of corporal predominant atrophic chronic gastritis and Zollinger-Ellison syndrome with Multiple Endocrine Neoplasia syndrome type I. Condyloma accuminatum is a HPV-related lesion most commonly seen in men practicing anal intercourse. Superimposed squamous dysplasia can be seen with HGD most frequently in the HIV-positive population. Recognition of the different classification systems of dysplasia, the most frequent settings in which these lesions are found, and their natural history is important for all practicing gastroenterologists and pathologists. PMID- 12134611 TI - The epidemic of esophageal adenocarcinoma. AB - The incidence and mortality related to esophageal adenocarcinoma (EAC) have been increasing in the United States, several European countries, and Oceania for the past 2 to 3 decades. Survival remains dismal, with little improvement during the same time period. Variations in the coding, classification, and detection of gastroesophageal malignancy may have contributed partially to the observed trends. Remarkable differences related to gender, ethnicity, and geography characterize the epidemiology of EAC. Gastroesophageal reflux disease (GERD) is the main risk factor for Barrett's esophagus, which is the only known precursor lesion for EAC. Several risk factors that promote the development of GERD and/or Barrett's esophagus have been proposed to explain these rising trends; these factors include the declining rates of Helicobacter pylori infection, obesity, dietary factors, and certain drugs. PMID- 12134612 TI - Barrett's esophagus: clinical characteristics. AB - Barrett's metaplasia develops in 6-14% of individuals with gastroesophageal reflux. Barrett's adenocarcinomas are increasing in epidemic proportions for as yet unknown reasons, approximately 0.5-1% of patients with Barrett's will develop adenocarcinoma. Heartburn duration and frequency (but not severity), male gender, and Caucasian race are major risk factors for developing cancer. Obesity and smoking are weak risk factors. Survival is determined by depth of tumor invasion (stage). Once invasion of the muscularis propia occurs, the vast majority of patients will have developed widespread metastasis, even when clinical staging studies are negative. No currently available therapy results in prolonged survival once metastases develop. Thus, the more widespread use of effective surveillance strategies is the only currently available means for reducing the morbidity and mortality associated with Barrett's adenocarcinoma. PMID- 12134613 TI - Cellular and molecular mechanisms responsible for progression of Barrett's metaplasia to esophageal carcinoma. AB - Barrett's metaplasia is found in approximately 12% to 18% of patients undergoing upper endoscopy for symptoms of reflux. Barrett's metaplasia is a premalignant condition and remains the number one risk factor for developing esophageal adenocarcinoma. There has been an increase in the incidence of esophageal adenocarcinoma in the past two decades, making it the most rapidly rising cancer in the United States and Western Europe. This article describes the progression from Barrett's metaplasia to esophageal adenocarcinoma and predictors for the development of adenocarcinoma in Barrett's metaplasia. Barrett's metaplasia represents a histological mosaic, with dysplastic tissue adjacent to non dysplastic tissue. The histologic changes leading to adenocarcinoma are accompanied by alterations at the molecular level, including the accumulation of gene mutations and changes in gene expression. The determination of the molecular events that occur in the transition from normal esophageal squamous mucosa to dysplasia and to esophageal adenocarcinoma have lead to a better understanding of the process of the transformation to adenocarcinoma. This knowledge will lead to better biomarkers to diagnose and assess cancer risk. PMID- 12134614 TI - Barrett's esophagus: surveillance and treatment. AB - Barrett's esophagus is a disease of paramount importance dictated by its exponentially increasing incidence and association with esophageal cancer. Unfortunately, our knowledge lacks much of the key data and information to make those important decisions in Barrett's as regards screening and treatment. With an evident lack of long-term large population studies of patients with Barrett's esophagus, much of our decision making is based on little available data and becomes somewhat arbitrary. Given these limitations, screening criteria include adults with a long history of persistent reflux symptoms. The precise age, frequency, and duration of symptoms used is unclear. Once Barrett's is identified, endoscopic surveillance is performed every 2 years, but this interval will most likely lengthen to 3 years. For patients with LGD, surveillance every 6 12 months is recommended. For HGD, esophagectomy is still recommended for healthy surgical candidates, but more patients will be followed, particularly those with "focal" HGD. The role of acid suppression either pharmacologically or surgically is not clear, but the tendency is to achieve near complete control of the acid reflux. Endoscopic ablative procedures are experimental and may be indicated in patients at high surgical risk with HGD and carcinoma. Its role in mainstream practice is yet to be defined. PMID- 12134615 TI - Multifocal atrophic gastritis and gastric carcinoma. AB - Gastric carcinoma remains a major cause of morbidity and mortality worldwide despite its significant decline in recent years. H. pylori infection begins with nonatrophic gastritis, and most individuals continue to have nonatrophic H. pylori gastritis throughout their lifetime. A minority of those with severe antral inflammation will develop a duodenal ulcer, and a few, for unknown reasons, may develop gastric MALT lymphoma. Others, who acquired the H. pylori infection in early childhood, develop progressive multifocal atrophic gastritis with loss of gastric glands. A small proportion of these individuals develop extensive, incomplete (type III) intestinal metaplasia, and an even smaller proportion will progress to dysplasia and intestinal-type gastric carcinoma. H. pylori-associated gastritis is also a risk factor for diffuse-type gastric carcinoma, which is not preceded by atrophy, intestinal metaplasia, or dysplasia. Appropriate screening and preventive measures should be considered in high-risk groups. It is also crucial to identify cofactors such as genetic susceptibility and environmental factors that might interact with H. pylori infection to increase gastric cancer risk. To make an impact on gastric cancer incidence and mortality, serious consideration should be given to early H. pylori eradication in high-risk groups and endoscopic surveillance according to the updated Sydney system in some patients with high-risk preneoplastic lesions, whereas dysplastic lesions should be removed without delay. Studies currently in progress may tell us whether H. pylori eradication can prevent later development of gastric carcinoma and thus eliminate a major cause of mortality worldwide. PMID- 12134616 TI - Role of Helicobacter pylori in gastric carcinogenesis. AB - Peptic ulcers and gastric malignancies are the two major complication of the course of Helicobacter pylori-associated chronic gastritis. Both gastric adenocarcinomas and MALT lymphomas occur in association with H. pylori infection, and studies support an etiological association. This article discusses the natural history of H. pylori-related gastric carcinogenesis and criteria to identify people susceptible to H. pylori infection-associated gastric cancer. It then reviews the molecular and genetic mechanisms underlying the malignant transformation of the gastric mucosa associated with H. pylori. PMID- 12134617 TI - Diagnosis and management of hereditary colon cancer. AB - Colorectal cancer is the third leading cause of cancer and the second leading cause of cancer death in the United States. About 130,000 new cases are diagnosed each year in North America, and 56,600 persons die annually from this disease. Approximately 5% of patients with colorectal cancer have clearly defined inherited syndromes such as familial adenomatous polyposis and hereditary nonpolyposis colorectal cancer. These conditions are well described both genetically and phenotypically, and are characterized by autosomal dominant inheritance, high penetrance, and high risk of colorectal cancer. We review the current recommendations for the diagnosis and management of these two hereditary forms of colorectal cancer. PMID- 12134618 TI - Colorectal cancer in inflammatory bowel disease: molecular and clinical features. AB - The two forms of inflammatory bowel disease (IBD), Crohn's disease and ulcerative colitis, are characterized by chronic and relapsing inflammation of the intestines. Initiating events presumably occur well before patients are symptomatic. Evidence gathered over the past decade from both IBD patients and animal models of intestinal inflammation have confirmed that IBD represents complex heterogenic forms of diseases, influenced by a combination of environmental, genetic, and immunologic factors working in concert to produce exaggerated immune responses, resulting in chronic and remitting inflammation. PMID- 12134619 TI - Colon cancer: detection and prevention. AB - Recent data have advanced our ability to detect, survey, and manage patients with colonic neoplasia. Current studies and consensus statements increasingly support the role of colonoscopic screening over less invasive testing such as FOBT or FS for appropriately selected individuals. There are many issues, however, that remain unresolved. What is the appropriate surveillance of an individual with a single family member who had colon cancer at an early age? How should family members of suspected HNPCC kindreds be managed? There has yet to be a prospective cohort validation of the Bethesda criteria in directing clinical practice, with the endpoint of mortality reduction. Questions regarding prophylaxis with dietary supplements and medications are exciting areas that are currently under study. As newer technologies become clinically available for molecular diagnostics and screening, and virtual colonoscopy with computed tomography and magnetic resonance disseminates, there will undoubtedly be new questions to be answered regarding their ability to aid in the detection and management of colon cancer. PMID- 12134620 TI - Primary prevention of colorectal cancer: diet and drugs. AB - Primary prevention of colonic adenomas and cancer through dietary interventions or chemoprevention has great appeal. This article discusses primary prevention goals and promising nutritional or chemopreventive strategies. There is substantial observational evidence that diets high in total calories and fat and or low in fruits and vegetables or total fiber as well as low levels of physical activity are related to the risk of colonic neoplasia. Similar observational data indicate that diets high in specific nutrients such as antioxidant vitamins or calcium may be protective. The article describes some of the newer chemopreventive agents and reviews the data linking diet and lifestyle to colorectal cancer risk, focusing on interventions that have also been studied in prospective clinical trials. Finally the evidence supporting the role of non steroidal anti-inflammatory drugs for the chemoprevention of CRC is reviewed and the status of several other promising newer agents that are entering human trials is summarized. PMID- 12134621 TI - Celiac disease and other precursors to small-bowel malignancy. AB - Small-intestinal malignancies are rare. Major risk factors for the development of these malignancies include celiac disease, which predisposes to both carcinoma and lymphoma. Crohn's disease patients have an increased risk of the development of adenocarcinoma, as do the inherited polyposis syndromes, FAP, and Peutz Jehgers syndrome. Each of these conditions provides unique models for the development of malignancy. PMID- 12134622 TI - Hepatocellular carcinoma: predisposing conditions and precursor lesions. AB - The global incidence of HCC is rising; in the United States, its rise is in parallel to that of cirrhosis due to the HCV and obesity epidemics. The lack of adequate treatment for advanced HCC mandates both prevention and early detection of these lesions. The limitations of currently available histopathologic evaluations, serologic markers, and radiographic imaging modalities in detecting HCC and its precursors have been outlined in this review. Refinements of all of these may lead to better HCC detection, earlier intervention, and successful treatment. Randomized controlled trials are necessary to evaluate the most efficacious and cost-effective approach to screening. PMID- 12134623 TI - Pancreatitis as a risk for pancreatic cancer. AB - Chronic pancreatitis clearly predisposes to pancreatic cancer, with early onset long duration chronic pancreatitis from cystic fibrosis, TP, and HP conferring the highest risk. Chronic pancreatitis is not a critical step, however, but rather one of several conditions that accelerate the accumulation of critical genetic mutations and chromosomal losses necessary for carcinogenesis. Indeed, other germline mutations, environmental factors such as tobacco smoking and alcohol consumption, or dietary factors may also accelerate the pathway to carcinogenesis, and may be synergistic with the conditions created by chronic pancreatitis. Because patients with chronic pancreatitis are at high risk of pancreatic cancer, the physician is faced with decisions on how to manage this risk. Discontinuing smoking and alcohol consumption, and perhaps dietary modification are obvious recommendations for risk reduction. If, however, the patient is older and already in a very high-risk category (e.g., long-standing HP), then screening for cancers must be considered. Inclusion in multicenter trials is recommended, and information on ongoing studies can be obtained through the office of Dr. Whitcomb, or as posted on www.pancreas.org. PMID- 12134624 TI - [Tau story: from frontotemporal dementia to other tauopathies]. AB - Tau proteins belong to the family of microtubule-associated proteins. They are mainly expressed in neurons where they play an important role in the assembly of tubulin monomers into microtubules to constitute the neuronal microtubules network. Tau proteins are translated from a single gene located on chromosome 17. Their expression is developmentally regulated by an alternative splicing mechanism and six different isoforms exist in the human adult brain. Tau proteins are the major constituents of fibrillar lesions described in Alzheimer's disease and numerous neurodegenerative disorders referred to as 'tauopathies'. Molecular analysis has revealed that an abnormal phosphorylation might be one of the important events in the process leading to their aggregation. Moreover, a specific set of pathological tau proteins exhibiting a typical biochemical pattern, and a different regional and laminar distribution could characterize each of these disorders. Finally, the recent discovery of tau gene mutations in fronto-temporal dementia with parkinsonism linked to chromosome 17 has reinforced the direct role attributed to tau proteins in the pathogenesis of neurodegenerative disorders, and underlined the fact that distinct sets of tau isoforms expressed in different neuronal populations could lead to different pathologies. Conversely, recent data in myotonic dystrophy has demonstrated that indirect effect (CTG repeat expansion) leading to variations in tau alternative splicing also produce neurofibrillary degeneration. PMID- 12134625 TI - [Molecular basis and physiopathogenic mechanisms of CADASIL: a model of small vessel diseases of the brain]. AB - Diseases of the small cerebral arteries account for approximately 20% of the ischaemic strokes. Their diagnosis is difficult and their pathogenic mechanisms are yet unclear. A certain proportion of these diseases is familial. CADASIL is a recently identified small-artery disease of the brain, which occurs both as an autosomal dominant and a sporadic condition caused by mutations in the Notch3 gene. Since the acronym CADASIL was coined to designate this disorder in 1993, an exponentially growing number of patients has been identified all over the world. Notch3 belongs to the highly conserved Notch genes family which encode transmembrane receptors involved in cell fate specification during development. The role of Notch3 is so far unknown. We recently established that in normal adult tissues expression of Notch3 is essentially restricted to vessel and vascular smooth muscle cells (VSMC). CADASIL patients carry highly stereotyped mutations leading to an odd number of cysteine residues within the extracellular domain. Mutations are associated with an impaired clearance of the Notch3 protein leading to its abnormal accumulation at the membrane of VSMC. These data establish that VSMC is the primary target of the pathogenic process. In addition they give support to the role of the Notch3 pathway in vascular homeostasis. Furthermore, they open new perspectives in the field of small-artery diseases of the brain and should help to further dissect their genetic etiologies and understand their pathogenic mechanisms. PMID- 12134626 TI - [Cell cycle control and self-renewal of embryonic stem cells]. AB - On one hand, self-renewal of mouse embryonic stem (ES) cells rely exclusively upon the LIFR beta/gp130-signaling pathway and the subsequent activation of the STAT3 transcription factor. On the other hand, the much-studied cellular machinery, that is organized to collect extracellular signals, transduce them via tyrosine kinase receptors and the SOS-RAS-RAF-MEK-MAPK pathway, ultimately leading to regulation of D-type cyclin expression, while regulation of the retinoblastoma (RB) protein phosphorylation is likely not to be operative in ES cells. We hypothetize that ES cells are blinkered by the lack of RB-dependent control of the G1/S transition, and that commitment into differentiation triggers the birth of a regulatable G1 phase. We discuss how the LIFR beta/gp130-signaling pathway and the ES cell-cycle machinery may functionally interact to promote self renewal. PMID- 12134627 TI - [Immunotherapy for HIV infections]. AB - HIV infection leads to a severe decrease of T CD4 lymphocytes with subsequent development of opportunistic infections. The use of current combination of antivirals has improved the morbidity and morbility of patients. Such treatments are not without severe drawbacks such as toxicity and resistance. Development of additional therapeutic strategies with cytokines, immunomodulators or therapeutic immunizations are currently intensely investigated. The clinical benefit of such strategies is under evaluation. However, these strategies may help to potentiate the immune restoration obtained with antivirals or alternatively to spare or delay the use of antivirals. PMID- 12134628 TI - [Interleukin 10 in disseminated lupus erythematosus]. AB - A number of years ago several groups reported that Interleukin 10 (IL10) was vigorously overproduced in disseminated lupus erythematosus (reviewed in Llorente et al., 2000). This hyperproduction obeys two different patterns. In one pattern, IL10 serum concentrations become elevated during disease and evolve in parallel with them. In the other, the patients' blood mononucleated cells spontaneously release increased IL10 quantities; contrary to serum variations in the first pattern, this increased output is not correlated with disease spurts. Interestingly an increase of this type is frequently found in disease-free parents of these patients (Llorente et al., 1997; Grondal et al., 1999). Some studies relate this IL10 hyperproduction to a genetic origin (Mehrian et al., 1998; Mok et al., 1998), while others seem to indicate an environmental cause (Grondal et al., 1999). Together these findings suggest two different possible origins for lupus IL10 overproduction. The first would be due to intrinsic anomalous IL10 production by some immune cells, the second would result from high IL10 output by tissues damaged by the inflammatory process. Considering the properties of IL10, which activates B lymphocytes and inhibits T lymphocytes, overproduction in lupus could explain the immune anomalies of this disease, which is characterized by anti-self antibodies production and by deficient T responses. Interestingly several of these anomalies are found independently from disease outbreaks and, in the case of some, in relatives of patients. The part played by IL10 in lupus has been inferred from the murine NZB-W model, in which an anti IL10 monoclonal antibody prevents development of the disease (Ishida et al., 1994); this antibody also prevents the appearance of human anti-ADN autoantibodies in immune-deficient mice restored with patients' lymphocytes. A direct demonstration of the role of IL10 in the etiology of lupus symptoms has recently been adduced in a pilot study bearing on six lupus patients, refractory to anti-inflammatory and immuno-suppressive treatments, who were treated for 3 weeks with a murine anti-IL10 monoclonal antibody (Llorente et al., 2000). This treatment brought about a rapid amelioration of the clinical picture, in particular of the cutaneous and articular symptoms, which was maintained for six months after this trial. This symptomatic amelioration was accompanied by a decrease of the biological signs of immune system hyperactivity and a partial amelioration of T lymphocyte function. The better clinical control of the disease allowed a significant decrease of corticotherapy. While this was an open study, without a control group and without randomisation, the rapid and important evolution of clinical and biological parameters towards normal strongly pleads for a favorable effect of the treatment. While confirming previous hypotheses about the rote of IL10 in lupus, this study leaves several points unexplained. First the clinical and biological amelioration in these patients treated with anti-IL10 monoclonal antibody occurred independently of circulating anti-ADN auto antibodies, indicating that the role of this interleukin in the disease is more complex than could be thought from the initial experiments in mouse. This unexpected result does not exclude that the beneficial effect of the antibody could be mediated by its action on B lymphocytes, since it is well known that these cells play important parts in development auto-immune processes other than only the production of auto-antibodies. It is also possible that the negative effect of IL10 in this disease exerts on other targets than B lymphocytes, in particular on fibroblasts and endothelial cells. Indeed it should be recalled that, while IL10 has often been held as an anti-inflammatory cytokine, its biological properties are more ambiguous, since it can immuno-stimulate some cell types. Among other remaining unknowns, the reason why some IL10 overproducing individuals, belonging to the family of lupus patients, do not develop lupus, is not understood. This fact underlines the multi-factorial origin of this disease, which must stem from associated genetic and environmental causes. IL10 overproduction, while it apparently promotes the symptoms, is certainly not sufficient. The efficiency of anti-IL 10 monoclonal antibodies during lupus, which are well tolerated, points to their usefulness in the therapeutic arsenal, especially in forms that are refractory to conventional anti-inflammatory and immunosuppressive treatments. In order to be validated, this hypothesis must be submitted to more in depth, randomized, studies. Moreover humanized antibodies should be developed, which would make it possible to reiterate the treatment during further outbreaks. Once all these conditions are fulfilled, the place of anti-IL10 treatment in lupus and the benefice-risk ratio should be compared to these of therapeutic approaches currently used in refractory forms. If further studies confirm the efficiency of- and tolerance to- IL10 neutralizing antibodies in lupus, it is well possible that these antibodies will eventually equate, for this disease, the recent use of TNF antagonists in the treatment of rheumatoid arthritis. PMID- 12134629 TI - [Eosinophils, parasites and allergy: from the biological to the clinical]. AB - Eosinophils have been considered for a long time as secondary cells, only able to be attracted by chemotactic factors and recruited from blood to tissues, at the site of inflammation. More recent studies have shown that their functions are not limited to the release of cytotoxic mediators, effector against parasitic targets but deleterious for tissues in allergy, but they can also participate in the regulation of immune response by producing type 1 and type 2 cytokines. Although questionable, animal models indicate a rather beneficial role of eosinophils in parasitic infections but a detrimental one in allergy. The results of clinical trials aiming at increasing or decreasing respectively their effects are discussed. PMID- 12134630 TI - [Early bronchial inflammation in cystic fibrosis]. AB - Cystic fibrosis (CF) is the most common genetic autosomal recessive disease in caucasian north-american and european populations. The CF gene codes for a transmembrane glycoprotein called CFTR (Cystic Fibrosis Transmembrane Conductance Regulator), a chloride channel which regulates the luminal secretion of chloride and the active ion and water transport in the airway epithelial cells. Mutations of the CF gene lead to a dysregulation of chloride and sodium channel associated to airway mucus dehydration, neutrophil-dominated airway inflammation and chronic infection responsible for the morbidity and mortality of CF patients. Although a high number of studies has been devoted to the CFTR pleiotropic functions, the chronology of the physiopathological events leading to the airway inflammation linked to mutations of the CF gene is still an open question. The issue of whether airway inflammation takes place before infection or is a consequence of infection during CF pathogenesis is still controversial. It has been recently reported that in broncho-alveolar lavages collected in CF infants, there is an increased level of interleukin IL-8 and abnormal low level of IL-10. The decreased IL-10 production has been confirmed in peripheral blood monocytes as well as in airway cell lines. Under basal conditions, the increased expression of the pro-inflammatory IL-8 cytokine has also been recently observed in the airway liquid secreted by CF naive humanized airway xenografts and in the supernatant culture of CF human airway epithelial cells. These results suggest that CFTR dysfunction may result in a constitutive pro-inflammatory vs anti-inflammatory imbalance in CF disease. Recent data from the literature suggest that the failure of chloride transport, the maturation defect and mistraffricking of mutated CFTR, lead to its accumulation in the endoplasmic reticulum and activation of NF-kappa B, responsible for the imbalance in the CF airway cell cytokine production. PMID- 12134631 TI - [Regulation of human neutrophil oxidative burst by pro- and anti-inflammatory cytokines]. AB - Human polymorphonuclear neutrophils play a key role in host defenses against invading microorganisms. In response to a variety of stimuli, neutrophils release large quantities of superoxide anion (O2.-) in a phenomenon known as the respiratory burst. O2.- is the precursor of potent oxidants, which are essential for bacterial killing and also potentiate inflammatory reactions. Regulation of this production is therefore critical to kill pathogens without inducing tissue injury. Neutrophil production of O2.- is dependent on the respiratory burst oxidase, or NADPH oxidase, a multicomponent enzyme system that catalyzes NADPH dependent reduction of oxygen to O2.-. NADPH oxidase is activated and regulated by various neutrophil stimuli at infectious or inflammatory sites. Proinflammatory cytokines such as GM-CSF, TNF and IL-8 modulate NADPH oxidase activity through a priming phenomenon. These cytokines induce a very weak oxidative response by PMN but strongly enhance neutrophil release of reactive oxygen species on exposure to a secondary applied stimulus such as bacterial N formyl peptides. Priming phenomena are involved in normal innate immune defense and in some inflammatory diseases. The mechanisms underlying the priming process are poorly understood, although some studies have suggested that priming with various agonists is regulated at the receptor and post-receptor levels. Resolution of inflammation involves desensitization phenomena and cytokines are involved in this process by various mechanisms. A better understanding of phenomena involved in the regulation of NADPH oxidase could help to develop novel therapeutic agents for inflammatory diseases involving abnormal neutrophil superoxide production. PMID- 12134632 TI - [Role of the peroxisome proliferator-activated receptors (PPARS) in the regulation of lipids and inflammation control]. AB - Peroxisome proliferator-activated receptors (PPARs) are ligand-activated transcription factors belonging to the nuclear receptor family. The hypolipidemic fibrates and the antidiabetic glitazones are synthetic ligands for PPAR alpha and PPAR gamma, respectively. Furthermore, fatty acids and eicosanoids are natural PPAR ligands. PPARs function as regulators of lipid and lipoprotein metabolism and glucose homeostasis and influence cellular proliferation, differentiation and apoptosis. PPAR alpha is highly expressed in tissues such as liver, muscle, kidney and heart, where it stimulates the beta-oxidative degradation of fatty acids. PPAR alpha furthermore mediates the action of the hypolipidemic drugs of the fibrate class on plasma lipoprotein metabolism. PPAR gamma is predominantly expressed in intestine and adipose tissue. PPAR gamma triggers adipocyte differentiation and promotes lipid storage. In addition, PPARs play a role in inflammation control. PPAR activators inhibit the activation of inflammatory response genes by negatively interfering with the NF-kappa B and AP-1 signalling pathways. PPAR activators exert these anti-inflammatory activities in different immunological and vascular wall cell types such as monocyte-macrophages, endothelial, epithelial and smooth muscle cells in which PPARs are expressed. These findings indicate a modulatory role for PPARs in the control of the inflammatory response with potential therapeutic applications in inflammation related diseases, such as atherosclerosis. PMID- 12134633 TI - [Why from clinic to biology?]. PMID- 12134634 TI - [Mutations of genes coding for TGF-beta receptors (BMPR2 and ALK-1) in primary pulmonary arterial hypertension]. AB - Pulmonary hypertension is defined by an elevation in pulmonary artery pressure leading to progressive right heart failure and death. Primary (idiopathic) pulmonary hypertension (PPH) is a rare disease with an estimated incidence of 2 per million inhabitants per year in France. Abnormal pulmonary artery angiogenesis is a characteristic feature of this condition including endothelial and smooth muscle cell proliferation in small to medium-sized pulmonary arteries. The recent discovery of germline mutations of genes coding for receptor members of TGF-beta (BMPR-2 et ALK-1) in PPH represents a considerable progress in the understanding of this pulmonary orphan disease. This review summarizes the current genetic data obtained in this condition. PMID- 12134635 TI - [Odors and olfaction]. AB - In this review, we discuss some of the neural processes involved in the perception of odors that, together with audition and vision, provide essential information for analyzing our surroundings. We shall see how odor detection and learning induce substantial structural and functional changes at the first relay of the olfactory system, i.e., the main olfactory bulb. Among the mechanisms that participate in these modifications is the persistence of a high level of interneuron neurogenesis within the adult olfactory bulb. Our goal is to present some observations related to the neurogenesis that may aid in understanding the neural mechanisms of sensory perception and shed light on the cellular basis of olfactory learning. We summarize the current ideas concerning the molecular mechanisms and organizational strategies used by the olfactory system to transduce, encode, and process information at various levels in the olfactory sensory pathway. Due to space constraints, this review focuses exclusively on the olfactory systems of vertebrates and primarily those of mammals. PMID- 12134636 TI - [Neurogenesis in the adult brain. Functional consequences]. AB - In the adult mammalian brain, neuroblasts are continuously produced within the subgranular zone of the hippocampus and the subventricular zone (SVZ) of the forebrain. In this review we describe how some physiological and environmental factors play important roles in regulating neurogenesis in the hippocampus. Neuroblasts in the SVZ network migrate rostrally into the olfactory bulb where they differentiate into local interneurons. We focus on the production, survival and functional consequences of these newly generated interneurons. We show that enriched odor-exposure enhances the number of newborn neurons in the adult olfactory bulb but not in the hippocampus. This effect did not result from changes in cell proliferation but rather was due to greater neuronal survival. Furthermore, the enriched condition was found to dramatically extend the olfactory memory. By maintaining a constitutive turnover of interneurons subjected to regulation by bulbar activity, ongoing neurogenesis plays a key role in olfactory memory. PMID- 12134637 TI - [Chemokines and immunomodulation: applications for HIV infections]. AB - The genetic control of HIV infection by the host involves a certain number of genes, among which those which code for chemokines/chemokines receptors, cytokines, MHC. Genes such as CCR5, CCR2, SDF1, and more recently CX3CR1 received great attention from several laboratories including ours, since they play a role as HIV coreceptor and, as such, on the infectivity of the host. In addition, it was shown that the polymorphism of these genes influences the evolution of infection, whether they have a protective or deleterious effect. Results obtained by our laboratory on the genetic polymorphism and its implication in HIV infection will be reported herein. Furthermore, to better understand their role, we looked for the capacities that the chemokines may have to play an immunomodulatory function, independently of their chemoattractive effect. In two examples, we showed that chemokines influence notably the cellular immune functions, such as CD8 cytotoxicity (Rantes/CCR3) and gamma interferon production (fractalkine/CX3CR1). Globally, the results indicate that chemokines/chemokines receptors polymorphism represent important epidemiological factors, but also contributes to evaluate the prognosis of HIV infection, through a better understanding of the disease physiopathology. PMID- 12134638 TI - Neurobiological mechanisms involved in recognition of olfactory signature of the young in sheep. AB - The following review focuses on neurobiological mechanisms responsible for the individual recognition of the olfactory signature of the young by the ewe at parturition. Steroids and vaginocervical stimulation are responsible for neurochemical and electrophysiological changes within the olfactory bulb that are part of the learning mechanisms of the individual lamb odour, thus allowing the establishment of a selective bond between the ewe and her lamb. There is an increase in the number of mitral cells, the principal cells of the olfactory bulb that respond to lamb odours, which is associated with increased release of glutamate and gamma-aminobutyric acid from the dendrodendritic synapses between the mitral and granule cells. The relation between the release of the two transmitters after birth suggests an increased efficacy of glutamate evoked gamma aminobutyric acid release. Parturition is also accompanied by increased oxytocinergic, cholinergic and noradrenergic neurotransmitter release that are essential for selective recognition of lambs. These increases in transmitter release depend on maternal experience, so that greater amounts have been found in multiparous than primiparous ewes. Therefore maternal experience seems to induce a neural maturation process that facilitates effective transmitter release in the olfactory bulb. PMID- 12134639 TI - [Proteins and mutations: a new vision (molecular) of neurodegenerative diseases]. AB - Neurodegenerative diseases have long been considered to be poorly defined, misunderstood, and inadequately treated. In recent years, research on Alzheimer's disease has led to numerous advances that have improved our understanding of this form of dementia and also of the entire category of neurodegenerative diseases. It now appears that numerous neurodegenerative diseases of the central nervous system correspond to the aggregation of specific proteins: beta-amyloid in Alzheimer disease, tau protein in Alzheimer disease, fronto-temporal dementia, progressive supranuclear palsy and corticobasal degeneration, alpha-synuclein in Parkinson disease and Lewy body dementia, PrP protein in prion diseases, SOD in amyotrophic lateral sclerosis, polyglutamine expansions in Huntington's disease and other diseases, etc. It is remarkable that in all these cases mutations have been identified for genes coding for these proteins and able to cause the disease and, moreover, that the introduction of the corresponding gene into transgenic mice (or other transgenic animals) has made it possible to create animal models of these conditions. This suggests that the proteins in question play a determinative role in the pathogenesis of these diseases and are not simply consequences of it. Neurodegenerative diseases are proteinopathies. But they are also networkopathies because the neuronal proteins are organized in functional networks. We must also note that all these diseases are associated with the process of aging, for they do not appear in the young. This fact suggests that the anomaly (genetic or otherwise) concerning a given protein does not suffice by itself to induce the disease process. Many observations suggest that the additional event involved, common to all neurodegenerative conditions, may be the intervention of free radicals. We thus propose here the theory that the diversity of neurodegenerative diseases is explained by the combination of two pathogenic events: one specific and associated with the aggregation of a particular protein in the nervous system, the other, non-specific and associated with aging and with the production and harmful actions of free radicals. This unified interpretation leads directly to treatment hypotheses: the development of drugs capable either of inhibiting the production or aggregation of proteins specifically implicated in diverse diseases (or promoting their elimination) or of inhibiting the production or action of free radicals in the nervous system. The former should target one of these various diseases, and the latter should act on a wide range of diseases. The two approaches may conceivably be combined. PMID- 12134640 TI - [Parkin, alpha-synuclein and other molecular aspects of Parkinson's disease]. AB - Parkinson's disease is a neurodegenerative disorder characterized by the progressive degeneration of the dopaminergic nigrostriatal pathway, and the presence of Lewy bodies. Over the past few years, several genes involved in inherited forms of the disease have been uncovered. In a small number of families with autosomal dominant inheritance, mutations have been identified in the genes encoding a-synuclein and ubiquitin carboxy-terminal hydrolase L1. Mutations in the parkin gene are a common cause of autosomal recessive parkinsonism with early onset, and also account for more than 15% of isolated cases with onset before age 45. The function of Parkin, a ubiquitin ligase involved in the degradation of protein substrates by the ubiquitin-proteasome pathway, highlights that ubiquitin mediated proteolysis may play an important role in the pathophysiology of idiopathic Parkinson's disease. PMID- 12134641 TI - First survey from the "Allergy Vigilance Network": life-threatening food allergies in France. AB - The Allergo-vigilance Network including 200 French allergologists and 28 European ones, has three objectives: to index the cases of lethal or prelethal anaphylaxis, to evaluate the prevalence of food allergies, clinical pictures and allergens, and to implement the post-marketing surveillance of the allergic risk of novel foods. The first survey dealing with life-threatening food allergies leads to an estimation of 15,000 to 30,000 reactions per year in France. The AVN could be a helpful tool for public health organizations. PMID- 12134642 TI - Broncho-alveolar inflammation in cork worker's asthma. AB - Suberosis is an occupational lung disease of cork workers associated with repeated exposure to mouldy cork dust in the cork industry, usually presenting as an interstitial lung disorder (Extrinsic Allergic Alveolitis). However, Occupational Asthma can also be associated with cork dust exposure and demonstrated by serial peak expiratory flow changes. AIM: To investigate broncho alveolar inflammation in patients with cork work-related occupational asthma, evaluated by Broncho-alveolar fluid cellular profiles and serial peak expiratory flow (PEF) rates monitoring. RESULTS: We studied 14 patients with respiratory symptoms associated with occupational exposure in the cork industry. Positive PEF monitoring occurred in 7 cases (Occupational Asthma-OA), and in 7 (Non occupational asthmatics-NOA) PEF records were negative. There were no differences in age, lung function (FEV1%, RV%), bronchial hyperresponsiveness, years of exposure, and atopy between the two patients groups. However, patients with work related asthma had higher BAL eosinophil counts than NOA (1.9 +/- 2.6% versus 0.2 +/- 0.3%; p < 0.05, Wilcoxon test). CONCLUSIONS: Cork worker's asthma, demonstrated by work related changes in serial PEF recordings, is associated with eosinophilic lung inflammation as described in other forms of occupational asthma. PMID- 12134643 TI - Heat-shock proteins and atherosclerosis. AB - In this review the authors focus on the possible role of heat-shock proteins (hsp) in the immune pathogenesis of the atherosclerotic process. The authors discuss evidence showing increased expression of these proteins in the vascular wall of stressed and atherosclerotic vessels and the immune mechanisms which could justify some of the inflammatory aspects that are now currently recognized in atherosclerosis, namely some of the possible hsp immune activating properties and also the possibility of hsp representing an innocent auto-antigen which could be the unwanted target of an immune response, initially directed against microbial heat-shock proteins. Epidemiological evidence linking atherosclerosis and cardiovascular diseases to soluble hsp levels as well as the intensity of anti-hsp immune response is also reviewed. PMID- 12134644 TI - Natural killer (NK) cells in mucocutaneous candidiasis. AB - Mucocutaneous Candidiasis is discussed and his clinical aspects and immunologic mechanisms reviewed. On a series of 57 patients clinical presentation the antibodies patterns for IgE, IgG and IgM have been studied and the cellular response evaluated by flux cytometry decrease on Natural Killer cells NK, CD3 CD16+CD56+ was achieved in 55% of 51 patients and is the more relevant abnormality in cellular immunity. CD8 cytotoxic cells was also found in 8% of the patients. In some cases where Nk and CD8 cells were normal IgG antibodies were nul pointing to as relevant antibody dependent cellular cytotoxicity mechanisms in mucocutaneous Candidiasis. PMID- 12134645 TI - [Allergenicity of lupin flour]. AB - Lupin flour is used in human food for its high quality nutritional and functional qualities. The frequency of crossed allergy between lupin flour and peanuts, both members of the family of Leguminosae, is strong, since 68% of patients who are allergic to peanut have shown positive reactions to lupin flour when tested by TPO-DA. Cases of isolated allergy to lupin flour without pre-existence of peanut allergy as well as workplace asthma by inhalation are also rarely seen. The specific allergens of lupin and those that participate in crosses with peanut have been studied by SDS-PAGE and immunoblot. The diversity of allergens contained in different lupin flour has also been studied. Further, the detection of lupin flour in a "pizza" flour which induced a strong allergic reaction exposed its eventual implication as a masked allergen. PMID- 12134646 TI - [Food allergy: one only finds what one seeks]. AB - We report three observations of food allergy to Penicillium. A systematic approach has allowed the retention of the surprising diagnosis in the first observation that we met. For those following we were content with a diagnosis of strong suspicion, without making a provocation test. The indications of provocation tests are discussed. As in these observations, it is possible that certain food allergens which are considered as rare are in fact more frequent than those reported in the literature; when an allergy does not bother the patient, when exclusion of the food is easy, food allergy is "forgotten" and not reported to the physician. PMID- 12134647 TI - [Prostanoids and acute lung injury]. AB - Prostanoids with various physiological effects in the lung, have been shown to be one of the important factors in acute lung injury. However, in large scale clinical studies, beneficial effects of NSAIDs or PGE1 have not been demonstrated. On the other hand, with progress of basic research in prostaglandin metabolic pathway, receptor and synthase of each prostanoid have been clarified, and selective COX-2 blockers have been developed and are now available in Japan. We reviewed the relationship between prostanoids, acute lung injury and the effects of COX-2 selective blockers, and discussed the present status and future perspective in this field. PMID- 12134648 TI - [Use of propofol for carotid endarterectomy: its effect on parameters of intraoperative monitoring and clinical outcomes]. AB - In order to evaluate the effects of propofol on neurologic outcomes of patients for carotid endarterectomy (CEA), intraoperative monitoring and postoperative complications were evaluated. Fifty two CEA cases were divided into two groups according to the method of anesthesia; group P (+) using propofol (n = 26), and group P (-) without propofol (n = 26). During cross-clamping of the internal carotid artery, oxygen saturation of internal jugular vein (SjO2) and somatosensory evoked potentials(SEP) were monitored continuously on the operated side. Moreover, the patients with postoperative neurological complications were examined for each parameter. There were no significant differences between the two groups in %SjO2, %SEP and number of cases with neurologic deficits. In the patients with poor outcomes, the value of stump pressure (SP) was significantly lower. We conclude that propofol had no significant effect against neurological complications in CEA patients, and SP monitoring was effective for evaluation of the clinical outcome in CEA patients. PMID- 12134649 TI - [The effects of propofol and midazolam on canine left ventricular contractility]. AB - Pronounced decrease in arterial blood pressure during propofol or midazolam infusion for sedation of critically ill patients, has raised the possibility that they have a direct negative inotropic action. Accordingly, in the current study, changes in the left ventricular (LV) contractility were examined during i.v. infusion of these two sedatives in anesthetized dogs. Myocardial contractility was assessed with the slope (Ees) of the LV end-systolic pressure-volume relationship and the slope (Msw) of the LV end-diastolic volume-stroke work relationship. Propofol was administered at 5, 10 and 20 mg.kg-1.h-1, and midazolam at 0.3, 0.6 and 1.2 mg.kg-1.h-1 over 60 min. Propofol caused dose dependent decrease in Ees (55 +/- 7 during the low dose to 27 +/- 3 mmHg.ml-1 during the high dose, P < 0.05) and in Msw (91 +/- 8 during the low dose to 67 +/ 6 erg.cm-3 x 10(-3) during the high dose, P < 0.05). Midazolam, also, decreased in Ees and Msw significantly. No significant differences were observed between three different doses of midazolam. It is concluded that propofol shows the dose dependent inhibition of myocardial contractility, but midazolam induces dose independent inhibition. PMID- 12134650 TI - [Higher blood propofol concentrations upon awakening after total intravenous anesthesia in patients with panperitonitis compared with patients for elective abdominal surgery]. AB - We examined the blood propofol concentrations when patients responded to vocal command in patients with panperitonitis (Group P:N = 11) and patients for elective laparotomy (Group E:N = 19). In both groups, general anesthesia was induced and maintained by intravenous administration of propofol, fentanyl and ketamine following insertion of an epidural catheter. Blood samples were taken from the left femoral artery at emergence from anesthesia, and blood propofol concentrations were measured by the HPLC. Although the demographic factors and the anesthetic conditions were similar in both groups, the blood propofol concentration at emergence from anesthesia in Group P was significantly higher than that in Group E (1.04 + 0.11 vs 0.84 + 0.14 microgram.ml-1, P = 0.03). Our data suggest that blood propofol levels should be kept higher during propofol anesthesia in patients with panperitonitis. PMID- 12134651 TI - [The usefulness of monitoring hepatic venous oxygen saturation and mild hypothermia anesthesia in two patients with portal vein tumor emboli during combined portal vein and pancreas resection]. AB - Two patients, a 48-year-old woman and a 61-year-old man, with portal vein tumor emboli underwent pancreaticoduodenectomy combined with portal vein resection. In order to protect the liver during occlusion of the portal vein, we induced mild hypothermia with surface cooling and hepatic oxygen status was monitored by hepatic venous oxygen saturation (ShvO2). During the temporary occlusion of the portal vein, the woman showed a marked decrease in ShvO2, significantly lower than 20%, and continued for about one hour, but on the other hand the man did not. But as the body temperature was raised, they showed the same tendency of decreased ShvO2. Postoperatively, her serum GOT, GPT, T-bilirubin were 104 UI.dl 1, 58 UI.dl-1, 1.95 mg.dl-1 and his data were 62, 42, 1.01 respectively. No other complication were observed. These results suggest that ShvO2 monitoring and mild hypothermia are clinically useful in detecting hepatic oxygen supply/demand unbalance and in protecting the liver. PMID- 12134652 TI - [Plasma ionized magnesium concentration following cardiopulmonary bypass]. AB - We performed a retrospective study to analyze plasma ionized magnesium concentration following cardiopulmonary bypass. Severe decrease of ionized magnesium concentration associated with frequent abnormal ECG sign was found in a patient with magnesium-free cardioplegia. Cardioplegia containing 16 mmol.l-1 of magnesium ion maintained ionized magnesium concentration within normal ranges without postoperative magnesium loading. Use of cardioplegia containing magnesium or adequate magnesium supplement is thought to be essential for patients receiving cardiopulmonary bypass. PMID- 12134654 TI - [Postoperative nausea and vomiting after open heart surgery]. AB - We investigated the incidence of the postoperative nausea and vomiting (PONV) following cardiac surgery with cardiopulmonary bypass. We conducted a prospective study of 65 cases with direct interviews by anesthesiologists who are blind to this protocol every 6 hours during ICU stay. There were no differences in age, body mass index, dose of fentanyl, operating time, cardiopulmonary bypass time and ventilation time in ICU between PONV(+) and PONV(-). Incidence of PONV was 43%, but 70% of female patients complained of PONV. Prophylactic antiemetic strategy might be clinically relevant to female patients who are to undergo open heart surgery with cardiopulmonary bypass. PMID- 12134653 TI - [Intraoperative hemodynamics in elderly patients during abdominal aortic aneurysm operations]. AB - An association between hemodynamics and age was assessed in abdominal aortic aneurysm resection and graft replacement. Changes in blood pressures and heart rates before and after clamping the aorta as well as before and after its declamping were compared in 34 cases of elective abdominal aortic resection from 1998 to 1999. The results were analyzed retrospectively reviewing their charts. The patients were divided into two groups: age under 70 and 70 or above. In the first group, the decrease in heart rate following clamping of the aorta was greater than that in the second group. On the contrary, after the aorta was declamped, a greater increase in the heart rate after declamping the aorta correlated with aging (R = 0.34, P < 0.05). However, the changes in blood pressure and heart rate were independent of preoperative coronary diseases. Whereas the people aged 70 or above showed different hemodynamics from those under 70, intraoperative hemodynamics did not reflect preoperative coronary diseases. PMID- 12134655 TI - [The relationship between end-tidal isoflurane concentration and electroencephalographic bispectral index during isoflurane/epidural anesthesia]. AB - We studied the effects of increases in isoflurane concentration on the bispectral index (BIS) in 16 patients undergoing lower abdominal surgery during isoflurane/epidural anesthesia. In 8 patients, the lungs were ventilated with an air/oxygen mixture (inspired oxygen fraction 0.33) [N(-) group], and in another 8 patients, the lungs were ventilated with 66% nitrous oxide in oxygen [N(+) group]. During surgery, patients received 1.0 MAC (1.15%) end-tidal isoflurane and the BIS was recorded after 10 min of unchanged end-tidal concentration. After this, we increased the end-tidal concentration of isoflurane by 0.2 MAC to 1.8 MAC. At each concentration step, the BIS was recorded again after 10 min of unchanged end-tidal concentration. At isoflurane concentration < 1.4 MAC, the BIS did not change with increasing isoflurane concentration in both groups (BIS values = about 40). In N (-) group, the BIS decreased in all patients at isoflurane concentration > 1.6 MAC. The mean BIS values were 22 (SD 18) at 1.6 MAC and 2(4) at 1.8 MAC, respectively. In N (+) group, the BIS decreased in four patients at isoflurane concentration > 1.6 MAC, and the BIS did not decrease at 1.8 MAC in another four patients. The mean BIS values were 27 (17) at 1.6 MAC and 21(21) at 1.8 MAC. The present data suggest that BIS may not correlate with anesthetic effect of isoflurane at isoflurane concentration > 1.0 MAC. PMID- 12134656 TI - [A case of adrenocorticotropic hormone (ACTH) deficiency]. AB - We report a case of ACTH deficiency. A 75-year-old man complained of anoxia, nausea and vomiting. Three years ago, he had an attack of loss of consciousness. On admission, his serum sodium level was down to 119.6 mEq.l-1. Plasma osmolality was low and urinary osmolality was high without edema, and he was diagnosed as having SIADH. After CRH test, rapid ACTH test and continuous ACTH test, he was diagnosed as having ACTH deficiency, and he was treated with steroids. One year later, he received urethrotomy due to urethrostenosis under spinal anesthesia with no trouble. In the next year, he was scheduled for sigmoidectomy due to sigmoid colon cancer under general anesthesia combined with epidural anesthesia. In the morning of his operation, he took hydrocortisone 10 mg per os. During operation, hydrocortisone 300 mg was given intravenously divided for three times. Plasma ACTH and aldosterone levels were below normal ranges, but serum cortisol was above the normal range. His operation was finished without troubles. Regarding this case, we discussed steroid therapy during anesthesia and operation. PMID- 12134657 TI - [Granulomatous polyp in the bronchus caused by anticoagulant therapy]. AB - A 73-year-old woman was scheduled for right lower lobectomy. Her trachea was intubated with a left double-lumen endobronchial tube (Sheribronch, 35 Fr). There was no bleeding at the trachea or the bronchus during the operation. She received intravenous heparin (4000 IU.day-1) for anticoagulant therapy one hour after operation. On the first post-operative day, the mucosal bleeding in the left main bronchus was confirmed, which developed a granulomatous polyp on the 17th post operative day. The anticoagulant therapy may have worsened slight mucosal damage caused by double lumen tube. It is important to select suitable tube size especially in patients who will receive anticoagulant therapy. PMID- 12134658 TI - [Postoperative management in single-lung transplantation for lymphangioleiomyomatosis]. AB - A 35-yr-old woman presented with dyspnea has been diagnosed as having lymphangioleiomyomatosis (LAM). Despite treatment with estrogen, her pulmonary function deteriorated progressively. In January 2001, a left single-lung transplantation was performed on her from a cadaveric donor. On admission to the ICU after the transplantation, arterial blood gas analysis showed a severe respiratory acidosis. A double-lumen endotracheal tube (ETT) was replaced by a single-lumen ETT for a better suctioning of secretion. Gas exchange improved after the replacement of ETT and suctioning of secretion by bronchoscopy. Five hours after the admission to the ICU, the breath sound decreased over the right thorax. The chest X-ray showed right pneumothorax, and a chest tube was inserted. The patient was weaned from mechanical ventilation gradually and extubated on the 6th ICU day. The patient was discharged from ICU to the general ward on the 9th ICU day without pneumonia and other complications. The development of pneumothorax in the recipient lung should be kept in mind during the perioperative period of lung transplantation for LAM. PMID- 12134659 TI - [Anesthetic management of an infant with Freeman-Sheldon syndrome]. AB - We report the anesthetic management of Freeman-Sheldon (whistling face) syndrome in a two-month-old boy scheduled for lateral canthoplasty. He had features of the syndrome including blepharophimosis, hypertelorism, a flat nose, microstomia with a limited opening, micrognathia, a very short webbed neck, scoliosis and multiple arthrogryposis. He was fed with a naso-gastric tube and suffered from several episodes of aspiration and oxygen desaturation. Difficult airway and intubation were anticipated. Anesthesia was induced via a mask with sevoflurane, although mask ventilation was difficult. Direct laryngoscopy and the insertion of a laryngeal mask airway were impossible due to microstomia with the limited opening as anticipated. A naso-tracheal intubation was achieved using a fiberoptic bronchoscope via a fiberoptic mask while ventilating the lungs. The operation and anesthesia afterwards were uneventful. In the ward, he was given supplemental oxygen but with occasional desaturation episodes. Thirteen days after the operation he was found cyanotic and resuscitation was attempted but failed. Autopsy demonstrated the hypoplasia of the lungs and thorax, atelectasis and bronchitis. PMID- 12134660 TI - [Transient complete atrioventricular block during renal transplantation]. AB - A 25-year-old female underwent renal transplantation. The patient had no complication preoperatively except hypertension. Preoperative electrocardiogram revealed no abnormality. Anesthesia was maintained with sevoflurane. Donor kidney was perfused with University of Wisconsin (UW)'s solution (4 degrees C) after removal. Transient complete atrioventricular block appeared twice after reperfusion of the transplanted kidney. Adenosine in the UW's solution was considered the major cause of atrioventricular block in this patient. Attention must be paid to the occurrence of atrioventricular block and bradycardia shortly after reperfusion in the case where UW's solution was used for donor kidney perfusion. PMID- 12134661 TI - [Cervical spondylosis revealed after general anesthesia]. AB - We present a case of complicated postoperative nerve injury. A 70-year-old man complained of partial paralysis and numbness of the left forearm 1.5 hour after operation for femoral artery bypass graft. Postoperative cervical spine MRI revealed cervical spondylotic radiculopathy. The MRI findings could account for his symptom. Not only peripheral nerve injury but the cervical radiculopathy may play an important role in perioperative neuropathy. PMID- 12134662 TI - [Emergency cesarean section in a patient with acute cortical blindness and eclampsia]. AB - A 28-year-old parturient with acute cortical blindness and eclampsia was scheduled for an emergency cesarean section. Computed tomography (CT) of the head revealed low-density areas suggesting cerebral edema in posterior regions. The operation was performed under general anesthesia with sevoflurane. The brain edema was managed by concentrated-glycerin and steroid. The operation was performed uneventfully. Her consciousness returned to normal by the following day and visual recovery was complete within 72 hr postpartum. The patient and the infant showed no complication on their discharge. PMID- 12134663 TI - [Paradoxical embolism during MIDCAB surgery]. AB - A 36-year-old woman underwent MIDCAB surgery. During the exposure of LAD, the right ventricular wall was injured. The bleeding was controlled by compression. After that, she developed hypotension followed by cardiac arrest. At the same time, TEE showed bubbles in all of the right ventricle. The open chest massage and epinephrine 1 mg restored the heart beat. It was thought that bubbles were brought to the right ventricle via the injured wall by the blower. A few minutes after the cardiac arrest, bubbles were detected in the left atrium by TEE. This phenomenon was suspected as transpulmonary paradoxical embolism because no cardiac shunt could be detected by TEE. PMID- 12134664 TI - [First clinical impressions of ProSeal laryngeal mask]. AB - ProSeal laryngeal mask airway (PLMA, LMA-ProSeal) is a newly developed laryngeal mask airway intended to overcome two major disadvantages of the conventional laryngeal mask airway, i.e., poor protection of the airway from gastric regurgitation and low sealing effect around the laryngeal inlet. The new PLMA can provide air-tight sealing by a second cuff positioned at the back of the mask and a bypass channel for the alimentary tract by a second tube (drain tube) opening at the tip of the mask. However, clinical feature of this new device has been assessed by few authors. Forty adult patients (19 males and 21 females) under general anesthesia immobilized with vecuronium were included in this study. A PLMA size 4 was inserted with (n = 10) or without (n = 30) an introducer at first (n = 37), second (n = 2) or third (n = 1) attempt and clear airway was obtained in all patients. After placing the PLMA in 10 patients (5 males and 5 females) sealing pressure was measured at cuff pressure of 0, 20, 40 and 60 cmH2O and at completely deflated cuff state. Averaged seal pressure at the 60 cm H2O cuff pressure was 22.7 +/- 8.0 cmH2O in the all 40 patients. Averaged seal pressure was higher in female than in male at all cuff pressure. The average seal pressure was 15.1 cmH2O even when the cuff was deflated completely. A lubricated 16-French gastric tube was inserted easily through the drain tube in all patients except one in which the mask was found to be deflected backward. There were no significant complications related to the PLMA. We concluded that the PLMA can provide high sealing pressure and isolate the airway from the alimentary tract. Further study including adverse effect of the high sealing effect should be required. PMID- 12134665 TI - [Ability of functional MRI to localize brain function]. PMID- 12134666 TI - [Gamma knife radiosurgery for arteriovenous malformations: anatomy, techniques, and avoidance]. PMID- 12134667 TI - [Endoscopic findings in chronic subdural hematoma]. AB - On the genesis of chronic subdural hematoma (CSH), it is one of the possible explanations that a head injury triggers uncertain cause off for the formation of the hematoma membrane, where the bleeding occurs to form and enlarge the hematoma. Although some reports on the staging classification based on computed tomography (CT) findings are available the natural course or history of the hematoma formation is not yet known. As previously reported, we have been using an endoscopically guided method in which a drainage tube is placed in the most frontal area of the cavity so as to evacuate the residual air efficiently. In this study, we analyzed the endoscopic findings in hematoma cavities and compared them with the clinical data, focusing on the post-trauma interval and recurrence of the hematoma. Between January 1999 and October 2000, we had an opportunity to observe the inner aspect of 60 hematomas in 48 patients. A trabecular structure was found in 39 hematoma cavities (65%), whereas apparent clot formation was observed in 18 cavities (30%). Septum formation leading to a multi-loculated hematoma was observed in 10 cavities (17%). Twenty-six patients with 32 hematomas with the obvious history of head injury were classified into 4 groups according to the post-traumatic interval to the surgery, which are stage I; less than 30 days, stage II; 31 to 60 days, stage III; 61 to 90 days and stage IV; more than 91 days. Clot formation and a trabecular structure were frequently seen in stages II and III. Clot formation was statistically and significantly seen in the recurrent group. The results suggest that CSHs increase in size in stages II and III, when clot formation and a trabecular structure are frequently observed. Endoscopic observation of hematomas may assist to know hematomas, and clot formation may be a warning of hematoma recurrence. PMID- 12134668 TI - [A case of brainstem encephalitis diagnosed by stereotactic biopsy]. AB - Brainstem encephalitis is a rare form of encephalitis which should be differentiated from cerebrovascular and neoplastic diseases. The authors report a case of viral brainstem encephalitis mimicking malignant lymphoma. A 55-year-old female was admitted to our hospital with gradually progressive diplopia and left hemiplegia. CT scan revealed low density lesions in the right globus pallidus and the anterior limb of the internal capsule. MRI demonstrated high intensity signals extending into the right cerebral peduncle, temporal lobe, thalamus and the contralateral thalamus on FLAIR images. Petechial hemorrhages were seen in the affected lesions, but no enhancement was observed following administration of a contrast material. CSF examination revealed mild mononuclear cell dominant pleocytosis. Both early and delayed images of 123I-IMP SPECT revealed marked hot spots corresponding to the lesions on FLAIR images. CT-guided stereotactic biopsy was useful for early diagnosis. PMID- 12134669 TI - [Postoperative subdural empyema due to Propionibacterium acnes]. AB - Propionibacterium acnes are gram-positive pleomorphic rods that grow under an anaerobic condition. They are the most frequent inhabitants of sebaceous glands of skin, hair follicles, mouth, upper respiratory tract, and as a frequent contaminant in laboratory cultures. Propionibacterium acnes is an underestimated but significant cause of postneurosurgical infection, especially in the presence of foreign bodies such as shunt systems. We present a rare case with a postoperative subdural empyema in which the only pathogen isolated was Propionibacterium acnes. In our case, the clinical course was atypically rapid. The most effective antibiotic therapy for Propionibacterium acnes infections has not been established. However, high dose penicillin in combination with surgical drainage and removal of foreign bodies is recommended as the treatment of choice. PMID- 12134670 TI - [Combined therapy with surgery and stereotactic radiosurgery for facial schwannoma: case report]. AB - The authors report the successful case of combined therapy using surgery and stereotactic radiosurgery for facial schwannoma in the middle cranial fossa, and discuss the surgical strategy for preservation of facial nerve function. This 27 year-old man presented with a 9-year history of right facial palsy and spasm. CT scan and MR imaging demonstrated a tumor 3 x 3 x 4 cm in size extending to the intradural middle cranial fossa from the petrous bone. After total removal of the intradural tumor, gamma knife radiosurgery was performed for residual tumor in the petrous bone. The marginal dose to the tumor was 12 Gy. Facial spasm disappeared, but facial palsy is unchanged 14 months after the radiosurgery. PMID- 12134671 TI - [Pituitary adenoma associated with neurofibromatosis type 1: case report]. AB - A case of pituitary adenoma associated is neurofibromatosis type 1 is reported. On June 6, 2000, a 49-year-old man was admitted to the Department of Neurological Surgery, Okayama University Hospital, for bitemporal hemianopsia. Twenty-nine years previously, he had been operated on for a left inguinal tumor that proved to be a neurofibroma. Based on the presence of other manifestations, such as cafe au-lait spots and subcutaneous nodules, he had been diagnosed with neurofibromatosis type 1, the same as his father, sister, and daughter. Computed tomography and magnetic resonance imaging demonstrated an intrasellar mass lesion with a cystic portion in the suprasellar region. Endocrinologically, almost all of his basic hormone levels were normal. A right front-temporal craniotomy was performed for a preoperative diagnosis of craniopharyngioma, and total intracapsular tumor extirpation was achieved. The histological diagnosis was clinically silent corticotroph pituitary adenoma. Neurofibromatosis is sometimes associated with neoplasms of the central nervous system, usually optic gliomas. Associations between pituitary adenomas and NF 1 are very rare and have been reported in only four cases, including the present case. PMID- 12134672 TI - [Orbital neuroma developing in the ciliary nerve]. AB - Orbital neuroma is a rare disease accounting for a certain percentage of all orbital tumors. However, because of the anatomical features of the orbit, the origin of the tumor has been diagnosed in only a few cases. The authors report a case of orbital neuroma arising from a short ciliary nerve that was confirmed during surgery. The patient was a 72-year-old female. She visited our hospital with complaints of left visual disturbance, exophthalmus and diplopia. Neurological examination on admission also revealed Marcus gunn pupil sign. CT scan and MRI imaging showed a left orbital tumor located along the superior surface of the optic nerve and adjacent to the posterior pole of the eye ball. She underwent superior orbitotomy through a transcranial approach. The tumor was encapsulated and adhered tightly to the sclera, short ciliary nerve, and optic nerve. For this reason, it was removed without a part of the adhered capsule. Since histological examination revealed neuroma, the tumor was diagnosed as short ciliary nerve neuroma. Postoperatively no additional deficits occurred. The exophthalmus, diplopia and Marcus gunn pupil sign disappeared and visual acuity was improved after the surgery. In such a case of short ciliary nerve neuroma, it is possible to remove the tumor without any neurological deficit, if careful consideration of microsurgical anatomy is made. PMID- 12134673 TI - [Gamma knife radiosurgery for malignant melanoma in the paranasal sinuses: case report]. AB - A case of malignant melanoma in the paranasal sinuses, successfully treated with gamma knife stereotactic radiosurgery, is reported. A 90-year-old man with left periorbital swelling was referred to our hospital for gamma knife radiosurgery. He had a 4-month history of left periorbital swelling and ophthalmalgia, but he was treated conservatively due to his age. CT showed a large mass with bone destruction located in the nasal cavity, paranasal sinuses, and orbita. A 15 Gy peripheral dose was administered to the upper portion of the tumor with the gamma knife technique, at the 50% isodose line using a 18 mm collimator (21 shots). Seven months after radiosurgery, his left periorbital swelling was improved markedly, and CT showed a significant reduction in the volume of the tumor. Gamma knife radiosurgery is a feasible treatment for malignant melanoma in the paranasal sinuses. It provides excellent quality of life, less injury to the patient, and fewer side effects than other treatment strategies. PMID- 12134674 TI - [Bilateral carotid stenting for bilateral carotid artery stenosis improved vascular dementia]. AB - We report a case of bilateral internal carotid artery (ICA) stenosis treated with stenting. A 78-year-old man suffered from vascular dementia and left hemiparesis, and, by magnetic resonance angiogram (MRA), was diagnosed as having bilateral ICA stenosis. Cerebral angiogram showed severe, bilateral ICA stenosis (right; 88%, left; 93%) and xenon single photon emission tomography (SPECT) showed severely decreased cerebral blood flow (CBF) and cerebrovascular reactivity (CVR). We performed bilateral carotid angioplasty with self-expanding stents. Both CBF and CVR were improved bilaterally after the operation. The patient was discharged without neurological deficits. Carotid stenting may be an alternative treatment for severe ischemia caused by severe, bilateral ICA stenosis. PMID- 12134676 TI - [Controversy and reform proposals concerning insured medical care in neurosurgery]. PMID- 12134675 TI - [ Basic knowledge of neuropathology for neurosurgeons]. PMID- 12134677 TI - [Controversy and reform proposals concerning insured medical care in neurosurgery]. PMID- 12134678 TI - [Collaboration of pediatric neurologists with diagnostic radiologists]. PMID- 12134679 TI - [P22 and N30 of median nerve SSEPs--independence of the frontal waves from parietal waves]. AB - Median nerve short-latency somatosensory evoked potentials (MN-SSEPs) were recorded from the scalp to assess frontal lobe functions in children. Forty-nine patients, aged between three and fifty years old underwent 87 examinations. They were divided into four groups: A (3 to 4 years old); B (5 to 6 years old); C (7 to 15 years old) and D (20 to 50 years old). Stimulus electrodes were placed near the wrist on the median nerve. The study was done mostly in the waking state. To evaluate the independence of the frontal lobe from the parietal lobe we correlated the frontal P22 and parietal N20, and compared the frontal N30 with parietal negative waves. The intervals of the peak latency differed between P22 and N20 by -1.5 to 1.5 msec. Significant differences were noted between groups A and D, and between C and D. N30 and negative parietal waves resembled each other in children 3 to 4 years of age, but became different from 5 years old. They were dissimilar after 6 years old. Therefore, N30 waves were useful for evaluating the frontal elements in children over 5 years of age. PMID- 12134680 TI - [Visual perception of Japanese characters and complicated figures: developmental changes of visual P300 event-related potentials]. AB - In order to evaluate developmental change of visual perception, the P300 event related potentials (ERPs) of visual oddball task were recorded in 34 healthy volunteers ranging from 7 to 37 years of age. The latency and amplitude of visual P300 in response to the Japanese ideogram stimuli (a pair of familiar Kanji characters or unfamiliar Kanji characters) and a pair of meaningless complicated figures were measured. Visual P300 was dominant at parietal area in almost all subjects. There was a significant difference of P300 latency among the three tasks. Reaction time to the both kind of Kanji tasks were significantly shorter than those to the complicated figure task. P300 latencies to the familiar Kanji, unfamiliar Kanji and figure stimuli decreased until 25.8, 26.9 and 29.4 years of age, respectively, and regression analysis revealed that a positive quadratic function could be fitted to the data. Around 9 years of age, the P300 latency/age slope was largest in the unfamiliar Kanji task. These findings suggest that visual P300 development depends on both the complexity of the tasks and specificity of the stimuli, which might reflect the variety in visual information processing. PMID- 12134681 TI - [Developmental change of Moro reflex studied with a three-dimensional motion analysis system]. AB - We evaluated developmental change of the reaction time of the Moro reflex from birth to three months of age using a three-dimensional motion analysis system (ToMoCo-VM:Tousou system). The reaction time was the shortest in one-month-old babies and became successively prolonged in two and three-month-old babies, showing a U-shaped pattern. The three phase of the Moro reflex, as defined by McGraw, changed with advancing age. The reaction time changed monthly. At the same monthly age, the reaction time was the same irrespective of the phase. PMID- 12134682 TI - [Early detection and therapeutic rearing for children with mental retardation: II. Assessment of gene analyses for the diagnosis of developmental disorders in Japan]. AB - The authors assessed the present status of gene analysis utilized in Japan. Physicians completed a mail-in questionnaire with their comments regarding the types of gene analyses they used, the perceived effectiveness of the analyses, and how they conducted genetic counselling for the patients. Among the 76 physicians who responded, gene analysis was most often conducted (n = 16, 75.0%) for muscle dystrophies, and considered to be effective by most of them (chi 2 (df = 2) = 11.99, p = 0.003). Many factors including the age of physicians, location of their facilities, and experience in child neurology were related to the utilization of gene analyses. Future studies based on ethical guidelines should establish a network between specialists and facilities and encourage medical approach to developmental disorders. PMID- 12134683 TI - [Clinical and genetical features of Japanese early-onset facioscapulohumeral muscular dystrophy]. AB - Facioscapulohumeral muscular dystrophy (FSHD) is an autosomal dominant form of muscular dystrophy characterized by progressive weakness and wasting of the facial, shoulder-girdle and upper arm muscles. The gene locus for FSHD is mapped to the subtelomeric region of chromosome 4q35, in which smaller EcoRI fragments (10 to 33 kb) are detected in most families by Southern blot analysis. The purpose of this study is to clarify the frequency and clinical/genetical features of early-onset FSHD in Japanese patients with 4q35-FSHD. In a series of 231 patients from 145 families with 4q35-FSHD, there were 31 patients (13.4%; male: female = 12:19) of 29 families (20%) with early-onset FSHD, 16 of whom were sporadic. Genetic analysis revealed that they had significantly smaller sized EcoRI fragments (range, 10 to 23 kb; mean 14.1 kb) than the other patients (range, 12 to 33 kb; mean 19.6 kb) (p < 0.001, U-test). All patients with the smallest EcoRI fragments (10 to 11 kb) were sporadic cases with early onset. Mental retardation (10/11) and epilepsy (4/11) was often observed in them, but not in the other patients. In early-onset FSHD, tongue muscle involvement (8/31) and swallowing disturbance (2/31), which has been regarded as exclusion criteria of FSHD, were also present. The onset of gait disturbance was significantly earlier (mean age = 11.9) in early-onset FSHD compared to the other group (mean age = 28.7). All patients with early-onset FSHD showed gait disturbance before 28 years of age. In conclusion, Japanese early-onset FSHD patients tend to have large gene deletions on chromosome 4q35, and show severe and variable phenotypes. PMID- 12134684 TI - [Soluble adhesion molecules in muscular dystrophy]. AB - To determine whether soluble adhesion molecules are affected in muscular dystrophy, we measured serum levels of creatine kinase (CK), soluble vascular cell adhesion molecule-1 (sVCAM-1), soluble intercellular adhesion molecule-1 (sICAM-1), soluble (s) E-selectin, and fibrin and fibrinogen degradation products (FDP) in 25 patients with Duchenne muscular dystrophy (DMD), 7 with Becker muscular dystrophy, 7 with Fukuyama type congenital muscular dystrophy, 6 with myotonic dystrophy (MyD), and 5 with spinal muscular atrophy (SMA) type 2, and also serum sVCAM-1, sICAM-1, and sE-selectin in 9 healthy controls. The levels of sVCAM-1 in the patients with DMD were 367.0-852.0 ng/ml (552.8 +/- 23.1) and significantly elevated than those in the patients with MyD, SMA type 2, and controls. The levels of sICAM-1 and sE-selectin in the patients with muscular dystrophy were 0.2-376.0 ng/ml and 17.9-119.0 ng/ml, respectively. They were also elevated than those in the patients with SMA type 2 and controls, but not significantly. The levels of sVCAM-1 and sE-selectin in the patients with DMD significantly correlated with age. There was no correlation between the levels of soluble adhesion molecules and those of CK or FDP in any groups. These changes of soluble adhesion molecules may reflect the process of muscle destruction and endothelial cell activation in muscular dystrophy. PMID- 12134685 TI - [Changes in the clinical picture of Fukuyama type. Congenital muscular dystrophy in its advanced stage: effectiveness of mechanical ventilation systems]. AB - We studied the clinical courses of Fukuyama type congenital muscular dystrophy (FCMD) based on a long-term follow-up of 7 patients. Their peak motor function varied from controlling the head incompletely to standing with support. Six patients had epilepsy, which in some cases was diagnosed with difficulty because of changing seizure types and multiple joint contracture. Six patients underwent endotracheal intubation, and in 5 of them long-term mechanical ventilation began at the age of 15 to 20 years. Three of them underwent trachestomy intermittent positive pressure ventilation (TIPPV), whereas recently 2 patients were treated successfully with nasal intermittent positive pressure ventilation (NIPPV). Mechanical in-exsufflation (MI-E) was effective in clearing airway secretion of 4 patients. With disease progression, the motor, mental, nutritional, cardiorespiratory and swallowing functions of FCMD patients should be assessed to ensure adequate management by NIPPV and MI-E. PMID- 12134686 TI - [Disclosure of a diagnosis of childhood autism and parents' acceptance of the disability]. AB - Mothers of 14 children (10 boys, 4 girls) with childhood autism answered the self report questionnaire regarding the disclosure of a diagnosis of autism and their acceptance of the disability. The children's mean age at the study was 8.6 years, and their ages at the time of the disclosure and acceptance were 4.0 and 5.2 years, respectively. The age at acceptance exhibited a statistically significant positive correlation with the age of disclosure indicating that early disclosure was related to early acceptance. The results obtained in the questionnaire suggested that the increased understanding of autistic behavior, the accumulated experience as to how to communicate with their own child and the existence of a self-care group to support the parents were important factors for the acceptance of the disability. The significance of the professionals' sympathetic manner at the time of disclosure, full explanation of the disability, instruction as to how to cope the disability and providing the information on social resources available in the local district was also discussed in relation to the acceptance of the disability. PMID- 12134687 TI - [A case of Friedreich's ataxia having no abnormal gene]. AB - We report here a 25-year-old girl with Friedreich's ataxia (FA) who showed slowly progressive ataxia, deep sensory disturbance and loss of large myelinated fiber in the sural nerve. There was no evidence of cerebellar atrophy or abnormal values of vitamin E, albumin, CK, and gamma-globulin in the serum. Except for mild mental retardation, her clinical and laboratory findings were consistent with those of FA. However, she had no abnormal GAA trinucleotide repeat expansion on chromosome 9q13, unlike typical FA patients in Europe. Her cardiac muscle is not involved instead of 20 years have passed since her ataxia developed. She is considered to belong to a specific type of FA which lacks cardiac muscle involvement and abnormal gene encoding frataxin. PMID- 12134688 TI - [Abnormal magnetic resonance imaging in a child with Alice in Wonderland syndrome following Epstein-Barr virus infection]. AB - Characteristic pathologic changes of cranial computed tomography (CT) and magnetic resonance imaging (MRI) have never been reported in "Alice in Wonderland" syndrome (AIWS) caused by Epstein-Barr (EB) virus infection. We present here a 10-year-old girl with AIWS with an abnormal MR finding. During the course of serologically confirmed EB virus encephalopathy, she had distortion of the body image, visual hallucinations and depersonalization characteristic of AIWS. MRI demonstrated transient T2 prolongation and swelling of the cerebral cortex, especially at the bilateral temporal lobes, bilateral cingulate gyrus, right upper frontal gyrus, bilateral caudate nucleus, and bilateral putamen, whereas CT showed no abnormalities. Transient MRI lesions were occasionally reported in patients with EB virus encephalopathy/encephalitis who presented visual illusions and psychotic reactions, although the diagnosis of AIWS was not described. We consider that any patient with symptoms of AIWS should have MRI because the abnormal MRI findings may disappear in a short period. PMID- 12134689 TI - [Paroxysmal tonic upgaze of childhood]. AB - We described a child who developed paroxysmal abnormal eye movement. At the age of 12 months, she had a high fever and a febrile seizure. On the next day she showed tonic upward deviation of the eyes for 1 to 2 seconds, and downward saccades on attempted downward gaze. The upward deviation was repeated for a period of 2 to 3 hours, and disappeared during sleep. The administration of L dopa was not effective. The symptoms subsided gradually over 10 months without other neurological impairment. She walked alone at 1 year and 6 months. At the beginning of her walking she showed truncal ataxia, but it gradually disappeared and her development was normal at the age of 2 years and 6 months. These abnormal eye movements and another symptoms were similar to paroxysmal tonic upgaze of childhood (PTU) that has been first described by Ouvrier and Billson (1988) as intermittent upward deviations of eyes. In Japan there was only one report of this syndrome with periventricular leukomalacia and hypomyelination. PMID- 12134690 TI - [A case of Kluver-Bucy syndrome after acute encephalopathy treated with selective serotonin reuptake inhibitor (fluvoxamine)]. PMID- 12134691 TI - [Cytogenetic and molecular studies on 210 patients with B-cell non-Hodgkin lymphoma using FISH and spectral karotyping]. PMID- 12134692 TI - [Cord blood transplantation]. PMID- 12134693 TI - [Therapeutic strategy for adult acute myeloid leukemia]. PMID- 12134694 TI - [Treatment strategy for adult acute lymphoblastic leukemia]. PMID- 12134695 TI - [Allogenic stem cell transplantation of adult patients from HLA-mismatch related and unrelated donors; single center experience]. PMID- 12134696 TI - [Transplantation using an unrelated donor of foreign registries]. PMID- 12134697 TI - [Allogeneic peripheral blood stem cell transplantation (allo PBSCT) compared with allogeneic bone marrow transplantation (allo BMT)]. PMID- 12134698 TI - [Comparison between unrelated bone marrow transplantation and cord blood transplantation in children with leukemia]. PMID- 12134699 TI - [Cord blood transplantation for adults]. PMID- 12134700 TI - [Thrombosis in collagen diseases--diversity of serum autoantibodies in antiphospholipid syndrome]. PMID- 12134701 TI - [Habitual abortion and thrombosis]. PMID- 12134702 TI - [Heparin-induced thrombocytopenia: the clinical picture and treatment]. PMID- 12134703 TI - [Thrombotic thrombocytopenic purpura induced by antibody to von Willebrand factor cleaving protease]. PMID- 12134704 TI - [Seven patients with stage I and II primary testicular lymphoma]. AB - Seven patients with stage I and II primary testicular lymphoma (PTL) have been treated since 1990 to the present at Kawasaki Medical School. All patients, whose median age was 56 yrs, had initially complained of swelling of the scrotal contents. The lesions were on the right in two patients, on the left in five, and no patient had bilateral lesions. The histological diagnosis was diffuse large B cell type in all patients. Five patients were classified as Ann Arbor stage I, and two at stage II. After high-orchiectomy for resection of tumor, two patients received chemotherapy alone, with a combination of chemotherapy and irradiation of the contralateral testis in the remaining five. Complete remission was achieved in all seven patients, but relapse occurred later in one. As it is recognized that, even in localized stage or low risk group PTL patients, the relapse rate in the central nervous system (CNS) and contralateral testis is quite high, chemotherapy, prophylactic CNS treatment and radiation of the contralateral testis after tumor resection should be included in the management of PTL. PMID- 12134705 TI - [Massive hemolysis following allogeneic peripheral blood stem cell transplantation with minor ABO incompatibility]. AB - We observed massive hemolysis after allogeneic peripheral blood stem cell transplantation with minor ABO incompatibility (the donor was group O and the recipient group A). The patient was a 21-year-old man diagnosed as having Philadelphia chromosome-positive chronic myelogenous leukemia. On day 7 post transplant, there was abrupt onset of massive hemolysis necessitating supportive treatment with transfusions. During the allogeneic peripheral blood stem cell transplantation the patient had received 2 x 10(8) CD3 positive cells/kg and 7.8 x 10(7) CD19 positive cells/kg donor lymphocytes with stem cells. The present case shows that in allogeneic peripheral blood stem cell transplantation with ABO incompatibility, severe hemolysis occurs early after transplantation presumably due to the transfusion of a large number of lymphocytes. PMID- 12134706 TI - [Therapy-related acute myeloid leukemia following double autologous peripheral blood stem cell transplantation for non-Hodgkin's lymphoma]. AB - A 29-year-old male was diagnosed as having non-Hodgkin's lymphoma (NHL, diffuse, large cell, B-cell, stage IV) in June 1999. He underwent 7 courses of chemotherapy and double autologous peripheral stem cell transplantation (total dose: CPA 13,000 mg, BUS 892 mg, L-PAM 150 mg, MCNU 870 mg, MTX 60 mg, Ara-C 160 mg, DXR 350 mg, VP-16 11,190 mg, VCR 8 mg, CBDCA 700 mg, and MIT 22 mg) for NHL and obtained complete remission in April 2000. In September 2000, he suffered from progressive general malaise. Laboratory findings showed marked leukocytosis with 85% leukemia cells, which were positive for alpha-naphthyl butyrate esterase. Surface-marker analysis of the leukemia cells showed positive results for CD11b, CD11c, CD13, CD15, CD33, CD56, CD64, CD65, CD71 and HLA-DR, and chromosomal analysis revealed add(8) (p11), add(9) (p13). He was diagnosed as having AML (M5a) and was still in complete remission for NHL. He did not respond to chemotherapy and died in December 2000, believed to be from therapy-related leukemia induced by the VP-16 used for treating NHL, judging by the patient's short clinical course and monocytic type of leukemia. PMID- 12134707 TI - [Successful treatment with G-CSF and continuous infusion of low-dose cytarabine and etoposide for therapy-related acute myeloid leukemia developed during chemotherapy for malignant lymphoma]. AB - In March 2000, a 30-year-old Chinese male was initially diagnosed as having non Hodgkin's lymphoma because of right cervical lymphadenopathy. He had received 8 cycles of chemotherapy including doxorubicin in China. As of February 2001, he was treated in our hospital with the CEPP regimen including etoposide, and was admitted in June 2001 because of leukopenia and thrombocytopenia. Peripheral blood showed hemoglobin 12.7 g/dl, platelets 4.1 x 10(4)/microliter and white blood cells 2300/microliter with 15% blasts. Bone marrow was hypocellular with 48% blasts, which were positive for myeloperoxidase, CD13 and CD33. Chromosome analysis showed 46,XY, t(9;11) (p21;q23) in all 20 metaphase spreads. He was diagnosed as having therapy-related acute myeloblastic leukemia (AML). Because of hypoplastic bone marrow, induction therapy with the CAG regimen including cytarabine, aclarubicin and granulocyte-colony stimulating factor (G-CSF) was started, but no apparent effect was observed. The patient was then treated with the AVG regimen comprising 250 micrograms of G-CSF and continuous infusion with 20 mg of cytarabine and 50 mg of etoposide for 14 days. Complete hematological and cytogenetic remission was achieved after two courses of the AVG regimen. Although it has been shown that the CAG regimen is effective for refractory and/or secondary AML, our results indicate that the AVG regimen should be tried for cases of AML resistant to the CAG regimen. PMID- 12134708 TI - [Coexistence of pure red cell aplasia and autoimmune hemolytic anemia occurring during remission of malignant lymphoma]. AB - A 56-year-old woman was admitted because of anemia in April 2001. A diagnosis of malignant lymphoma (ML) (diffuse mixed, B-cell type) had been made in March 2000 whereafter she had been treated with CHOP chemotherapy and had achieved complete remission (CR). On examination, it was found she had concurrently developed pure red cell aplasia (PRCA) and warm type autoimmune hemolytic anemia (AIHA) without relapse of the ML. The PRCA and AIHA were successfully treated with prednisolone. To our knowledge, only 4 cases of PRCA and AIHA associated with ML have been reported. Moreover, this is the first report of the coexistence of PRCA and AIHA occurring during the CR of ML. PMID- 12134709 TI - Antioxidants in the prevention of chronic diseases. PMID- 12134710 TI - An update: vitamin E supplementation and heart disease. AB - In vitro studies and experiments in animal models provide a large and compelling body of evidence that oxidation of low-density lipoprotein (LDL) and/or related oxidative mechanisms play a critical role in the initiation and progression of atherogenesis. A corollary to the theory that reactive oxygen and nitrogen species ("free radicals") are the key molecules in this process is that antioxidants that can protect LDL from peroxidation should decrease the risk of developing atherosclerosis, attenuate its progression, or even reverse established disease. However, recently, clinical trials employing the principal lipid-soluble dietary antioxidant, vitamin E, have provided mixed results indicating either benefit, no effect, or an adverse impact on patients with cardiovascular disease (CVD). Consideration of the design and outcome of these studies together with new reports about the action of antioxidants suggests approaches for new studies as well as a basis for current advice to patients. PMID- 12134711 TI - The role of carotenoids in human health. AB - Dietary carotenoids are thought to provide health benefits in decreasing the risk of disease, particularly certain cancers and eye disease. The carotenoids that have been most studied in this regard are beta-carotene, lycopene, lutein, and zeaxanthin. In part, the beneficial effects of carotenoids are thought to be due to their role as antioxidants. beta-Carotene may have added benefits due its ability to be converted to vitamin A. Furthermore, lutein and zeaxanthin may be protective in eye disease because they absorb damaging blue light that enters the eye. Food sources of these compounds include a variety of fruits and vegetables, although the primary sources of lycopene are tomato and tomato products. Additionally, egg yolk is a highly bioavailable source of lutein and zeaxanthin. These carotenoids are available in supplement form. However, intervention trials with large doses of beta-carotene found an adverse effect on the incidence of lung cancer in smokers and workers exposed to asbestos. Until the efficacy and safety of taking supplements containing these nutrients can be determined, current dietary recommendations of diets high in fruits and vegetables are advised. PMID- 12134712 TI - Vitamin C function and status in chronic disease. AB - Vitamin C is an essential dietary nutrient required as a co-factor for many enzymes, and humans are among the few animals that lack the ability to synthesize the compound from glucose. The reduced form of the vitamin, ascorbic acid, is an especially effective antioxidant owing to its high electron-donating power and ready conversion back to the active reduced form. Concentrations of the vitamin in body tissues and fluids are regulated through interactions of intestinal absorption, cellular transport, and excretion. The amount of vitamin C needed to prevent scurvy is very small and easily obtained in nearly all Western diets. There is great interest in the clinical roles of vitamin C because of evidence that oxidative damage is a root cause of, or at least associated with, many diseases. Population studies show that individuals with high intakes of vitamin C have lower risk of a number of chronic diseases, including heart disease, cancer, eye diseases, and neurodegenerative conditions. However, these results may simply reflect a more healthful diet or lifestyle for individuals with a high vitamin C intake. At present, data from controlled clinical trials have not established that higher intakes of vitamin C alone will help prevent chronic degenerative diseases. However, the evidence that ascorbic acid acts as an important antioxidant in many body tissues is convincing. The new higher Recommended Dietary Allowance (RDA) for vitamin C of 75 mg for women and 90 mg for men is, for the first time, based on the vitamin's role as an antioxidant as well as protection from deficiency. In healthy people, amounts greater than the RDA do not appear to be helpful. Vitamin C nutriture may be more important for people with certain diseases or conditions. High intakes of the vitamin are generally well tolerated; a Tolerable Upper Level was recently set at 2 g based on gastrointestinal upset that sometimes accompanies excessive intakes. PMID- 12134714 TI - The evidence for the effectiveness of probiotics for the prevention and treatment of human diseases. PMID- 12134713 TI - Selenium, an antioxidant nutrient. AB - Selenium is an essential constituent of a number of enzymes, some of which have antioxidant functions. Deficiency of the element in animals makes them susceptible to injury by certain types of oxidative stress. At least 1 human disease occurs only in selenium-deficient individuals. Therefore, it seems prudent to avoid selenium deficiency. The plasma (or serum) selenium concentration is often used to assess selenium nutritional status. A plasma selenium concentration of 8 micrograms/dL or greater in a healthy subject indicates that plasma selenoproteins are optimized and the subject is selenium replete. The Third National Health and Nutrition Examination Survey determined plasma selenium in 17,630 subjects in the United States. Its results indicate that more than 99% of the subjects studied were selenium replete. The Institute of Medicine has set the Recommended Dietary Allowance for selenium at 55 micrograms per day for adults. Since most estimates of selenium intake in the United States are 80 micrograms per day or greater, routine selenium supplementation is not recommended in the United States. PMID- 12134715 TI - Pharmacologic B-vitamin therapy for hyperhomocysteinemia in dialysis patients: has the time come? AB - Hyperhomocysteinemia, the state of elevated plasma homocysteine levels, is an independent risk factor for atherothrombosis and arteriosclerosis. For incompletely understood reasons, renal disease appears to predispose to hyperhomocysteinemia. In fact, as renal function declines, plasma homocysteine levels rise. Mild to moderate hyperhomocysteinemia is almost universal among end stage renal disease (ESRD) patients, who have negligible functioning renal mass. This tendency towards a hyperhomocysteinemic state may partially explain the dramatically high rate of cardiovascular morbidity and death seen in this population. Hyperhomocysteinemic subjects with normal kidney function can usually reduce or normalize homocysteine levels with modest B-vitamin (folic acid, vitamin B6, vitamin B12) supplementation. However, subjects with reduced renal function require much higher ("pharmacologic") B-vitamin doses to achieve equivalent reductions, while ESRD subjects are resistant to even massive doses. Study design flaws and the inclusion of potentially B-vitamin-deficient subjects in homocysteine-lowering trials has made interpretation of this literature difficult. Nonetheless, it appears that the modest standard daily dialysis vitamin supplements are equivalent to pharmacologic B-vitamin therapy in lowering homocysteine levels in ESRD patients and should be the recommended method of treatment in this population at the present time. PMID- 12134716 TI - Nutrition counseling in the physician's office: how do I bill for advising my patient? AB - In addition to diagnosing and treating illnesses, physicians often provide nutrition counseling services. Billing and obtaining appropriate reimbursement for these services present a unique challenge. This article utilizes 4 hypothetical scenarios as a basis for examining common categories of service that involve nutrition counseling. These categories are: routine physicals that include nutrition counseling, nutrition counseling alone, nutrition counseling as a small part of a problem-oriented "evaluation and management" (E/M) service, and nutrition counseling as the bulk of a problem-oriented E/M service. The basics of procedure (CPT) and diagnosis (ICD-9) coding are described, and specific billing instructions for each scenario are provided. Although this article provides an introduction to the topic of billing, healthcare providers and their office staff should inform themselves on general coding and billing practices, as well as local payors' guidelines. PMID- 12134717 TI - Probiotics for urogenital health. AB - Bacterial vaginosis, urinary tract infection, and yeast vaginitis afflict an estimated 1 billion women each year. Once investigation has ruled out complicated underlying causes, the only therapeutic option is antimicrobial agents. In many cases, this is effective at clearing infection. However, recurrences, side effects, and secondary infections are frequent. Coinciding with infection is a disruption of the normal commensal microflora in the vagina, primarily a loss of lactobacilli. The exogenous application of lactobacilli to the host as probiotic agents appears to offer hope as an alternative management regimen to antimicrobial treatment and prophylaxis. Although commercial probiotics specifically selected and proven to be effective for urogenital infections are not yet available, there is growing in vitro and human data to suggest that certain strains could confer health benefits on a large number of women. Given that depleted vaginal lactobacilli and recurrent infection is associated with increased risk of sexually transmitted diseases and preterm labor, multiple antibiotic resistance, and significant reduction in quality of life, the need for probiotic therapeutics has never been greater. PMID- 12134718 TI - Nutritional factors in the control of blood pressure and hypertension. AB - Differing hypertension prevalence rates between certain population and age groups are partially due to differences in the intake of certain nutrients. Blood pressure is positively associated with higher sodium, alcohol, and protein intakes; it is inversely associated with potassium, calcium, and magnesium intakes. Salt may lead to an increase in blood pressure in the presence of salt sensitivity, but there is no inexpensive or easy strategy to identify salt sensitive patients. Other risk factors for hypertension include obesity and lack of regular physical activity. The best strategy appears to be moderate salt restriction (6-7 g/day) in combination with an optimal compliance of the antihypertensive drug therapy, as well as adoption of the combination diet of the DASH study--a diet rich in fruits and vegetables, and thus rich in potassium. Current evidence does not support the increased intake of Ca2+ or Mg2+ for blood pressure-lowering purposes only; however, calcium and magnesium may represent important components in the combination diet of the DASH study. It seems that it is the combination of these nutrients that is of crucial importance for the achievement of optimal blood-pressure reduction. Also recommended is a decrease in alcohol consumption and an increase in regular physical activity. Instead of a severe intervention with regard to 1 risk factor alone, positive changes in 5 habits combined--high salt intake, high sodium-to-potassium ratio, alcohol intake, calorie imbalance, and a sedentary life--may be the most realistic and effective strategy to counteract the present hypertension epidemic. PMID- 12134719 TI - [Stroke. It can also affect children (interview by Michael Koczorek)]. PMID- 12134720 TI - [Cooling and irrigation, volume replacement, pain management. First aid in burns]. AB - Of decisive importance for the outcome of burn victims are the depth and extent of the burn, and the age and general state of health of the patient. At the site of the injury, the most important initial measure is the abundant application of cold water to the undressed victim. Early intubation of the burn victim should not be employed too readily. When the burns cover more than 10% of the body surface there is an acute danger of shock, and rapid fluid replacement must be aggressively pursued; the infused volume is calculated on the basis of the extent of the burn (Parkland formula). To treat pain, the i.v. administration of opiates or ketamine in combination with benzodiazepines has proven value. The onsite management of the burns should be restricted to the application of sterile dressings. Grade 1 and 2a burns may be treated out of hospital by cooling, cold water application, burn ointment dressings. More severe burns require hospitalization and surgical management. Since the severity of burn trauma is often underestimated, rapid hospitalization is to be recommended. PMID- 12134721 TI - [Options of plastic surgery in burns. How the burn victim gets a "new skin"]. AB - Plastic surgery following burns may be divided into primary and secondary reconstructive measures. Primary reconstruction covers all steps leading to the healing of the patient's wounds. Secondary reconstruction comprises all subsequent interventions required to improve the results of primary surgery. The treatment of burn victims is always a team effort. Reconstructive measures aim to achieve, as early as possible, anatomical primary reconstruction that takes account of the aspects of function and cosmesis. The primary surgical operation is all important--if successful, only few subsequent corrective interventions will be needed. PMID- 12134722 TI - [Rehabilitation of burn victims. A difficult path back to normality]. AB - The most important aims of rehabilitation in burn victims is the restitution and improvement of joint mobility, mimicry and pulmonary function, as well as of muscular endurance and strength. In addition to the management of scars, therefore, patient instruction in unsupervised training and scar care, as well as promotion of re-integration into day-to-day life are essential. Depending on the parts of the body affected, measures may include manual therapy, active exercise, treatment with ultrasound, proprioceptive neuromuscular facilitation in the case of facial burns, respiratory therapy and ergotherapy and, finally coordination training. Treatment of the scars themselves requires a combination of a number of measures, all of which should be of an "active" nature. Of proven value are manual massage of scar tissue, stretching exercises, the use of silicone, special splints and compression clothing, as well as laser therapy. Rehabilitation measures should be applied for not less than 4 weeks. Where indicated, further surgical measures should be discussed with the patient. PMID- 12134723 TI - [ACE inhibitor or AT1 antagonist. Is there a differential therapy?]. AB - ACE-inhibitors and AT, receptor antagonists play an important role in the treatment of cardiovascular and renal diseases. The criteria of evidence-based medicine indicate that ACE-inhibitors (in appropriate combinations with other cardiovascular agents) continue to represent the treatment of first choice in chronic heart failure, post-myocardial infarction with compromised ventricular function, high cardiovascular risk, and diabetic (type 1) nephropathy. It is here that the AT1 antagonists should be used--in particular when ACE-inhibitors are not tolerated. The AT1 antagonists, Irbesartan and Losartan, are applied, at appropriately high doses, primarily in hypertensives with diabetic (type 2) nephropathy. With the current exception of chronic heart failure, no confirmed data are yet available on the value of combination treatment. The LIFE study has shown that an AT1 antagonist is superior to an established beta blocker in hypertensives with an increased risk (left-ventricular hypertrophy), in particular when diabetes mellitus is impending or already present. PMID- 12134724 TI - [Peripheral arterial occlusive disease. Symptoms, basic diagnosis and staged therapy]. AB - Intermittent claudication or rest pain are typical symptoms of peripheral arterial occlusive disease (PAOD) affecting the lower limbs. The pain is localized one level below that of the occlusion. Initial investigations should determine skin temperature and color, pulse status, stenotic sounds and Doppler occlusive pressures. If intermittent claudication is present, angiography of the pelvis and legs then follows. Treatment is stage-dependent: while in stages I and IIa conservative treatment such as cessation of smoking, administration of acetylsalicylic acid and walking training suffices, stages IIb and higher require invasive measures extending from PTA to amputation of gangrenous parts of the limb. PMID- 12134725 TI - [When the physicians misuse their power. Sex with patients]. PMID- 12134726 TI - [Comparative study with prokinetic agents. Dyspepsia can be managed with phytotherapy]. PMID- 12134727 TI - [How do pediatricians vaccinate in Munich? A health care epidemiological study]. AB - OBJECTIVES: The goal was to determine vaccination rates and predictors of vaccination behaviour of pediatricians in Munich, Germany. METHODS: Standardised questionnaires were sent to all 133 office-based pediatricians in Munich. RESULTS: 97 questionnaires (73%) were returned completely. Vaccination rates were sufficiently against diphtheria, tetanus, and poliomyelitis only. The other vaccinations recommended by the national immunisation advisory board (STIKO) were carried out to an insufficient extent. Vaccination rates of measles, mumps, and rubella were low. Parents' informed consent to STIKO's recommendations was the main determining factor of vaccination rates. Physicians' uncertainty in assessing contra-indications correctly was another important reason for insufficient vaccination rates. CONCLUSIONS: The health target of eliminating measles will not likely be reached. STIKO's recommendations as well as periodically vocational training courses, should be more compulsory to physicians. Self-responsibility of patients and parents should be raised by financial incentives on sick funds premiums. PMID- 12134728 TI - Assessment of physical activity with a pedometer and its relationship with VO2max among adolescents in Switzerland. AB - OBJECTIVES: In the absence of a gold standard, the assessment of physical activity in children remains difficult. To record physical activity with a pedometer and to examine to what extent it is correlated with VO2max. METHODS: Survey on physical activity and fitness; 233 Swiss adolescents aged 11 to 15 carried a pedometer (Pedoboy) during seven consecutive days. VO2max was estimated through an endurance shuttle run test. RESULTS: The physical activity recorded by the pedometer did not vary from one day to the other (p > 0.05). The physical activity was higher among boys than among girls (p < 0.001) and higher among younger adolescents (6th versus 8th grade; p < 0.001). The correlation between physical activity and estimated VO2max was 0.30 (p < 0.01). CONCLUSIONS: The use of a pedometer to assess physical activity over one entire week is feasible among adolescents. The record provided by the pedometer gives an objective measure of the usual physical activity and, as such, is relatively well correlated with aerobic capacity. PMID- 12134729 TI - [Validation and subjective reports of exposure to traffic by traffic count, NO2 dispersion modeling and measuring NO2 emissions]. AB - OBJECTIVES: The validity of self-reports of traffic density on street of residence in cities has not been evaluated extensively yet. METHODS: This study compared traffic self-reports with exposure estimates based on traffic counts and emission measurements. RESULTS: Self-reports correlated well with traffic count data but less well with data from measurements of background emission. CONCLUSIONS: As traffic counts primarily represent direct exposure to road traffic and emission concentrations primarily represent city background exposure to traffic-related pollutants, self-reports reflect direct traffic exposure more strongly than background exposure. PMID- 12134730 TI - Bicycle safety helmet usage in Berlin 1999: an observational study. PMID- 12134731 TI - Factors associated with differences in antihypertensive drug treatment: results from the MONICA Augsburg Population Surveys 1989/90 and 1994/95. AB - OBJECTIVE: To investigate in a population-based sample of hypertensive men and women the impact of factors that determine frequency and pattern of antihypertensive treatment. METHODS: We used pooled data of two independent surveys conducted 1989/90 and 1994/95 in the Augsburg region among men and women aged 25 to 74 years. Co-morbidity and cardiovascular risk factors were assessed by questionnaire and examination. Multivariate and polytomous logistic regression analyses were used to assess treatment patterns adjusting for cofactors. RESULTS: Among a total of 9,795 survey participants, 2,279 men and 1,699 women were hypertensive with 30% of men and 43% of women using antihypertensive drugs. Hypercholesterolemia was unrelated and smoking even inversely related to treatment (adjusted odds ratio OR = 0.7, 95% CI 0.56-0.92). Women were more likely to be treated than men (adjusted OR = 1.6, 95% CI: 1.34-1.82). Furthermore, women on monotherapy used diuretics significantly more often than men (adjusted OR relative to betablocker use 2.8, 95% CI: 1.6-4.8). CONCLUSIONS: We identified several determinants of drug treatment among hypertensives in the community. During the study period antihypertensive treatment seemed to be determined primarily by clinical disease but not by concomitant cardiovascular risk factor status, such as male gender, smoking, or hypercholesterolemia. PMID- 12134732 TI - Poverty and injury risk among children: always together? PMID- 12134733 TI - The first years of implementation of the Swiss National Environment and Health Action Plan (NEHAP): lessons for environmental health promotion. AB - The National Environment and Health Action Plans (NEHAPs) are a novel attempt to integrate environmental protection and health promotion in political programmes. Throughout Europe, about 40 NEHAPs have been developed so far. The Swiss NEHAP was among the first to be developed in an industrialised country. We discuss strength and weaknesses of the Swiss NEHAP and draw first conclusions on the development and implementation process of such programmes, illustrated by examples of other European NEHAPs. The strengths of the Swiss NEHAP lie in the formulation of specific targets in selected areas, its approach as a environmental health promotion programme, and its comprehensive evaluation. Weaknesses in most NEHAPs are the lack of involvement of the general public and of the economic sector, and the absence of an implementation strategy along with adequate financing. PMID- 12134734 TI - Towards assessing effects of National Environmental Health Action Plans. PMID- 12134735 TI - Environment and health: from national policies to global initiatives. PMID- 12134736 TI - A joint effort in the field of environment and health. PMID- 12134737 TI - Statistical methods in epidemiology: Karl Pearson, Ronald Ross, Major Greenwood and Austin Bradford Hill, 1900-1945. AB - The tradition of epidemiological study through observation and the use of vital statistics dates back to the 18th century in Britain. At the close of the 19th century, however, a new and more sophisticated statistical approach emerged, from a base in the discipline of mathematics, which was eventually to transform the practice of epidemiology. This paper traces the evolution of that new analytical approach within English epidemiology through the work of four key contributors to its inception and establishment within the wider discipline. PMID- 12134738 TI - Prevalence of influenza immunisation in Australia and suggestions for future targeting of campaigns. AB - OBJECTIVES: Reports on the results of a national survey conducted in Australia, in 2000. The objectives were to determine national estimates of influenza vaccination coverage for each state and territory of Australia, to obtain information related to attitudes towards and influences on immunisation decisions and explain the factors involved with failure to immunise. METHOD: The survey was conducted using the Computer Assisted Telephone Interview (CATI) system. The overall participation rate for the survey was 88.6% and the final number of completed interviews across Australia was n = 10,505. RESULTS: Two target groups, those aged 65 years and over and those "at risk" of influenza aged between 40 and 64 years were defined. The overall immunisation rates in these two groups were 74% and 32% resp. The rate of immunisation among females generally exceeded that of males. A multivariate model provided the best joint set of explanatory variables for not getting immunised. These include sex, income, general practitioner recommendation, and general perceptions regarding the influenza injection. CONCLUSION: This study identified important issues in the decisions of people to immunise. It also highlighted the need to target the findings in effective immunisation policies and strategies to improve health outcomes for those at risk of adverse influenza events. PMID- 12134740 TI - Nursing profession unveils strategic plan to ensure safe, quality patient care and address root causes of the growing shortage. PMID- 12134739 TI - Recruiting the 21st century nurse: getting to the heart of the matter. PMID- 12134742 TI - We must recruit more nurses. PMID- 12134743 TI - Regulations contribute to shortage. PMID- 12134744 TI - Drop all insurance. PMID- 12134745 TI - Liability crisis and future physicians. PMID- 12134746 TI - Improve children's mental health. PMID- 12134747 TI - Medicaid mess. PMID- 12134748 TI - Cooking up trouble. PMID- 12134749 TI - The tie that binds too tightly. PMID- 12134750 TI - Closing the health gap. PMID- 12134751 TI - Medical mismatch? PMID- 12134752 TI - An analysis of the reactivity of vaccines administered in Texas from 1991 through 1999. AB - Vaccination is one of the greatest advances in medical history. Continued improvements in vaccination will include the introduction of new vaccines and the improvement of the efficacy and safety of existing vaccines. This report analyzes the 8623 reports made to the Vaccine Adverse Events Reporting System from the state of Texas from 1991 through 1999. The severity, time of onset, and vaccine type of the reactions, as well as the sex and age of those affected, were examined. Physicians and other health care providers need to be vigilant in their reporting of adverse reactions to vaccines. Persons injured by vaccination should be referred to the National Vaccine Injury Compensation Program. PMID- 12134753 TI - New forms for athletic physicals. PMID- 12134754 TI - Take the offensive. PMID- 12134755 TI - ["Atypical theft offender"--a useful diagnostic concept for forensic psychiatry?]. AB - The paper presents a case report of a serial but from the social point of view rather inconspicuous theft offender. The motivation of this perpetrator cannot be reduced to a simple money-making-intention. It seems to be of a more complex origin which offers possibilities for a psychological interpretation. On the other hand, this offender does clearly not fulfill the criteria of kleptomania. For such cases, Anglo-American literature offers a new concept named the "atypical theft offender" ATO, witch is outlined and critically discussed by the author. PMID- 12134756 TI - [Injury pattern in fatal automobile accident passengers due to rear-end collisions]. AB - The article reports on the injury pattern seen in fatally wounded car passengers after rear-end collisions on motorways. The study material included 5 rear-end collisions, in which 9 car occupants (5 drivers and 4 front-seat passengers) were killed. The change in velocity (delta v) was above 60 km/h in all the reported cases. The injury pattern showed the following characteristics: extensive soft tissue traumatization (subcutaneous haematomas, blood-filled pockets, "decollements") not discernible from outside on the back of the trunk, mostly in the lumbosacral region; dorsal serial rib fractures, often combined with fractures of the spine (including the sacrum and the coccyx) and/or the pelvis; ruptures of the aorta at the typical sites of predisposition; ruptures of the liver and spleen (particularly at the visceral surfaces). PMID- 12134757 TI - [Fatal craniocerebral trauma in a young child. I: Forensic documentation, model generation and geometric impact conditions]. AB - The aim of forensic biomechanics is the reconstruction of traumatic events based on the pathological findings in the victim's morphology and on the traces and shapes of the traumatizing tools. The introduction of Streifenlichttopometrie into forensic science enables 3-dimensional and proportionally accurate documentation of the victim's body and injuring agent with submillimeter precision. The advantages of this method of documentation are the possibilities of producing animated models which correspond exactly to the body's shape and injury topography, and of ascertaining the physical parameters (centers of mass, moments of inertia) of the various body parts and the geometrical impact conditions for the reconstruction of the injury dynamics. This way of proceeding thus enables more precise models than hitherto possible for kinetic and dynamic reconstruction. In the case of an infant who was fatally injured by a wooden sculpture the generation and application of computer aided 3-dimensional reconstruction models are shown. PMID- 12134758 TI - [Forensic medicine significance of the fluid content of the sphenoid sinuses]. AB - In the forensic literature the opinion is often held that the presence of aqueous liquid in the paranasal sinuses in conjunction with other findings (plume of froth around the mouth and nostrils, emphysema aquosum, Paltauf's spots, increased haemolysis etc.) can be regarded as a sign of drowning. Especially the sphenoid sinus is easily accessible at autopsy; its content can be aspirated from the base of the skull with a cannula. The valency of the liquid content in the sphenoid sinuses was consecutively investigated in 60 deaths by drowning and 157 other deaths. The results showed that in 92% of the deaths by drowning between 1 ml and 4 ml of aqueous fluid could be found in the sphenoid sinuses. However, a positive result was also obtained in 52% of the other cases autopsied, but in the control group the average volume of the aspirate was smaller than in the group of deaths by drowning. PMID- 12134759 TI - [Fatal inhalation of butane-propane gas]. AB - Fatal intoxications with butane and/or propane are rare although the inhalation of such liquid gases in order to induce hallucinations is not uncommon amongst youngsters, the number of which is difficult to evaluate. Thus the possibility of gas intoxication should be taken into consideration in all cases of unclear death of youngsters, in which case the macroscopic and histological findings will be unspecific whereas the chemical-toxicological analyses, especially of the native brain, lung and liver tissue, lead to definite conclusions. PMID- 12134760 TI - [Death in the neonatal period and infancy. History of medicine and forensic criminal aspects]. AB - From antiquity up to the present time the history of medicine contains innumerable examples of the different attitude of human beings in dealing with the death of children. This is paradigmatically described for the death of neonates and infants, with special consideration of the sudden infant death syndrome (SIDS) and selected forensic-criminalistic aspects. Against the historical background of forensic postmortem examination and forensic paidopathology the development of the autopsy is also outlined. PMID- 12134761 TI - Seroprevalence of antibodies to herpes simplex virus types 1 and 2 among two sexually active female populations in Middlesbrough, England. AB - Herpes simplex virus (HSV) infection occurs worldwide but its epidemiology varies between different countries and between groups of individuals. Seroprevalence of HSV-1 and HSV-2 antibodies is a more accurate method of determining epidemiology of this infection. In this cross-sectional study, 467 blood samples were obtained from women attending the genitourinary medicine clinic (GUM) and antenatal clinic (ANC) in Middlesbrough, England. Analysis of their blood samples showed that 69.5% GUM patients had HSV-1 antibodies and the figure was 54.5% in the ANC patients, p < 0.0001, with odds ratio 1.9 (95% CI 1.30 to 2.78) for higher prevalence in the GUM group. Also, for HSV-2 antibodies, the seroprevalence was 21.6% for GUM and 8.1% for ANC patients, p < 0.0001, with an odds ratio of 3.13 (95% CI 1.70 to 6.02) for higher prevalence in the GUM group. The overall prevalence for HSV-1 is 63.2% and for HSV-2 is 15.8%. Many of these patients were asymptomatic and constitute a potential source for spreading the virus. Knowledge of HSV seroprevalence can help in planning sexual health promotion strategies. PMID- 12134762 TI - Audit of primary care angina management in Sandwell, England. AB - This audit aimed to assess the identification and treatment of coronary risk factors, lifestyle advice given and use of drug therapy, among patients with angina in Sandwell. It was designed to help general practices evaluate their angina management, and highlight areas where practice could be improved. Criteria were based on Sandwell's published angina audit and local clinical guidelines. Each participating practice was asked to identify all patients with angina, from which a 10% sample was randomly selected. The notes of each selected patient were examined for evidence showing whether agreed standards of care had been achieved. Fifteen practices took part, and contributed data on a total of 358 patients. Of patients without contraindications, 66.5% were taking aspirin, 62.1% were prescribed short-acting nitrates and 58.4% were prescribed beta-blockers. Non white patients were significantly less likely to receive short-acting nitrates (p < 0.001) and women were significantly less likely to receive beta-blockers (p < 0.01). A total of 83.5% of patients had received smoking cessation advice, 75.1% had weight advice, 75.1% were advised about alcohol use and 64.5% about exercise. Overall, 77.4% had a blood pressure check within the previous twelve months, 40.5% had their cholesterol measured and 33.5% had their blood glucose measured. Non-white patients were significantly less likely to receive smoking cessation, weight, exercise and alcohol advice and were less likely to have their blood pressure checked (all p < 0.0001). Patients aged 65 and over were significantly less likely to receive a cholesterol check (p < 0.0001). None of the auditable standards were actually met. This study shows that there is considerable scope to improve the management of angina patients, with particular regard to aspirin. We recommend that practices develop systems to ensure that the appropriate treatment, advice and checks are given to all patients with angina, paying particular attention to those from ethnic minority backgrounds. PMID- 12134763 TI - Health and safety of laboratory science students in Ibadan, Nigeria. AB - Laboratory science students are engaged in laboratory practice under supervision during the course of their training programme. They are exposed to the risk of laboratory-acquired infection and need to be adequately informed and equipped with facilities to protect their health. A questionnaire was administered to laboratory science students to determine their perception of hazards in laboratory practice and the observance of safety codes in their work practices. Of 128 students, 118 completed the questionnaire, a response rate of 92%. Sixty of them (51%) were males and 53 (45%) were females; five students did not indicate their sex. The results revealed that only 34 (29%) of the students use gloves for handling biological samples and 26 (22%) use gloves for handling clinical waste. Ninety-four students (80%) reported that they washed their hands after handling specimens. Eighteen of the students (15%) had been immunised against tuberculosis, 80 (68%) against tetanus, six (5%) against hepatitis B, and 18 (15%) against yellow fever. Ninety-six students (81%) thought the greatest hazard in laboratory practice was harmful biological organisms, while 13 (11%) indicated that chemical agents were the greatest hazard. Virology was thought to be the most hazardous specialty by 41 students (35%) while morbid anatomy was ranked as least hazardous by 48 (41%) of the students. These findings indicate that whilst laboratory science students are aware of the hazards in laboratory practice, this knowledge is not translated to safe practices and students may endanger their health as a result of exposure to laboratory practice. They therefore need to be provided with adequate facilities to protect themselves and adequate supervision to ensure that they imbibe safe work practices during their training years. PMID- 12134764 TI - William Stewart Halsted--surgeon extraordinaire: a story of 'drugs, gloves and romance'. AB - As a surgeon William S Halsted's most notable contributions were in the development of a technique for radical mastectomy, the repair of inguinal herniae and the advancement of bowel anastomoses. However, he will perhaps be best remembered for his introduction of the use of the surgical glove and his reasons were probably the least scientific of all those given. Developed by the Goodyear Rubber Company, the gaunts were made to protect the hands of his scrub nurse, Caroline Hampton, the future Mrs William S Halsted. Another of Halsted's contributions was the development of topical anaesthesia using cocaine. A consequence of these experiments was the development of addiction to the drug and his later dependence on morphia. The last century has seen a 360 degrees turn in the reasons why we use gloves. Originally designed to protect the theatre staff from corrosive substances, subsequently to protect the patient from contamination by theatre staff, to today where gloves are worn to protect staff from blood borne infection agents. This article introduces William S Halsted, and his story of romance, drugs and the introduction of gloves to medical care. PMID- 12134765 TI - The Bouveret syndrome: an unusual complication of gallstone disease. AB - We present a case of Bouveret's syndrome which is gastric outlet obstruction due to gallstone impaction in the duodenum. The paper also discusses the aetiology, presentation, methods of diagnosis and options for management of Bouveret's syndrome. PMID- 12134766 TI - Climate change--myth or meltdown? PMID- 12134767 TI - Infectious diseases--an enemy of public health. PMID- 12134768 TI - Sharps and needle-stick injuries in supported housing and emergency accommodation. PMID- 12134769 TI - Travel-associated intestinal disease. PMID- 12134770 TI - Food hygiene training--measuring success. PMID- 12134771 TI - The global emergency of tuberculosis: what is the cause? AB - The treatment of tuberculosis is cheap and highly effective, yet worldwide the disease remains a serious cause of illness and death; so serious as to have been declared a 'global emergency' in 1993. It is principally a disease of poverty, with 95% of cases and 98% of deaths occurring in developing countries. The incidence of tuberculosis is increasing worldwide, partly due to poverty and inequity and partly to the HIV/AIDS pandemic, which greatly increases the risk of infection proceeding to overt disease. Around 30% of AIDS-related deaths are due to tuberculosis. The emergence of multidrug resistant tuberculosis (MDRTB) is an increasing threat to tuberculosis control. Although treatable with alternative drugs, the cost is enormous and, accordingly, not undertaken in many poor nations. While the overall global incidence of MDRTB is low, it occurs in certain 'hotspots' including Russian prisons. Due to adverse socio-economic factors, London has not escaped the general rise in incidence and, without the introduction of active control strategies, there could be a serious epidemic as occurred in New York City ten years ago which required an enormous financial outlay for its control. In view of the global emergency of tuberculosis, the WHO 'Stop TB' campaign has called for the universal adoption of its directly observed therapy, short course (DOTS) strategy. Also, though the Massive Effort Against Diseases of Poverty, several international agencies are urging the establishment of effective control programmes worldwide. London should take the lead and set an example. PMID- 12134772 TI - Tuberculosis information on the Web. AB - After years of decline, tuberculosis has reemerged as a serious public health problem worldwide. Factors contributing to this resurgence include the HIV epidemic and immigration of people from countries with a high incidence of tuberculosis. In 1993, the World Health Organization (WHO) declared it to be a 'global emergency' and according to a recent WHO report, there were 7.96 million new cases in 1996, with two million deaths. Comprehensive up-to-date information is available via the Internet, which has emerged as an essential tool for information on a wide variety of subjects including tuberculosis. However, looking for the right information on tuberculosis can be laborious, especially with the enormous number of tuberculosis websites established. It is always helpful to know which ones to visit to avoid wasting valuable time. This article provides readers with a list of websites that provide information on tuberculosis. All sites quoted here have been tested and proven to be valuable and informative for both the public and scientific community alike. PMID- 12134773 TI - Infectious diseases in air travellers arriving in the UK. AB - The ease of access to air travel and its increased popularity over the last 30 years have led to a significant incidence of imported infectious diseases and potential infectious hazards. The commonest type of illness found is acute gastroenteritis. Tuberculosis and malaria are not currently common conditions encountered in the UK, but medical vigilance is increasingly necessary as a result of these and other infectious diseases being carried by arriving air travellers. Risks of transmission to other passengers have been considered, and tuberculosis has been shown to have relatively low infectivity on commercial flights. Incidence of serious communicable disease occurring in arriving passengers is low, and should be referred to communicable disease specialists for advice on management. High standards of precautionary hygiene measures are mandatory to commercial aircraft to prevent spread of infectious agents. Disease vectors and products of animal origin pose additional potential threats to public health. Vigilance by environmental health specialists helps maintain national defences against this group of threats. Alertness to recent travel history and awareness of international public health concerns is essential for clinicians likely to encounter sick members of the travelling public. The largest commercial airports have health surveillance units, tasked with acting as a first line of defence against infectious disease. The majority of cases do not present in flight or at the airport, so they can present to any primary care clinician or emergency department. An integrated strategy for health protection will be developed in the UK with the setting up of a Health Protection Agency. PMID- 12134774 TI - Cholera: a continuous epidemic in Africa. AB - Cholera continues to plague many parts of the world, but has largely been concentrated in Africa, which contributes more than 80% of the total cases worldwide. Natural disasters, like the 2000 floods in Mozambique and the volcanic eruption in the Democratic Republic of the Congo in 2002, generally lead to new outbreaks of the disease. The refugee problem in many countries throughout the world also causes potential threats for disease outbreaks. Case fatality rates are high, and we are not anywhere near curbing new cholera epidemics, especially in Africa. It is thus imperative to renew discussions about the nature of this deadly disease, its treatment, measures for prevention and control, modes of transmission, its physical, social and economic impact, and potential solutions. PMID- 12134776 TI - Community self-help and the homeless poor in Latin America. AB - Finding realistic housing solutions that are able to respond to the realities of poverty in the developing, or newly industrializing, world are frequently distinct from those suited to the developed world due to levels of poverty and differing welfare regimes. This requires a different understanding of the concept of housing and shelter for developing and developed countries. Population increase and emerging habitation patterns in parts of Latin America have required that policy-makers review traditional 'top down' approaches to the way the homeless poor are treated and how self-help or 'bottom up' schemes are increasingly seen as a sustainable way forward in providing affordable housing options to both governments and communities. Over the last decades, mass in migration to cities has put pressure on governments to provide public housing- but two major problems arose: firstly, governments found it difficult to finance the increasing demand for public housing, and secondly, the nature of employment and the informal economy in the developing world meant that this new housing was often too costly for the urban poor, in some cases increasing homelessness still further. Recent policy developments tend to favour supporting what the poor are and have been able to achieve for themselves, with appropriate government support. Upgrading shack settlements is now recognised as a community driven and cost-effective response that can, if appropriately supported, offer an initial and sustainable solution to urban housing need by tapping into additional non governmental sources of funding. In the absence of a major public sector housing stock to meet demand, governments are also recognising that self-help housing schemes for families able to access funding and resources offer a further innovative approach to meeting housing need. The nature of housing and shelter in the developing world requires a unique response so that it remains attainable and affordable to the poor. Neo-liberal policies, increasingly adopted in Latin America, are not able to provide suitable, sustainable and affordable housing delivery and alternatives need to be explored. This paper traces some developments in Latin American housing policy and explores some of the challenges that are faced in responding to the unique housing needs of the urban poor. PMID- 12134775 TI - Dogs, zoonoses and immunosuppression. AB - Dogs are the source of a wide range of zoonotic infections that pose a significant threat to human health. This is particularly the case for immunocompromised people, although there are few robust studies that determine immunosuppression as a risk factor for transmission of zoonoses from dogs to humans. An increasing proportion of human society is immunodeficient, principally through the advent of HIV infection and through more people, particularly the expanding elderly group, being subjected to immunosuppressive agents. This is happening at a time when more such people are capitalizing on the acknowledged benefits of dog ownership, making for a potentially dangerous mix. Enteric pathogens (for example, Salmonella, Campylobacter and Cryptosporidium species, that may be canine derived) are a frequent risk to the health of immunocompromised persons. Veterinarians and physicians can be criticised for not communicating with each other, and for not providing adequate risk assessment to pet owners. There is scope for voluntary groups to provide information and support for the immunosuppressed who wish to keep their dogs. Key recommendations are to maintain a clean personal environment and intact mucocutaneous barriers. Public health professionals could help rectify the current communications gap between veterinary and medical staff and so facilitate in the appropriate management of dog-owning immunocompromised people. PMID- 12134777 TI - Mayhem or misadventure? PMID- 12134778 TI - Visual vignette. Mediastinal mass due to substernal goiter. PMID- 12134779 TI - William Farr, The Lancet, and epidemic cholera. PMID- 12134780 TI - Embryonic stem cell research: the relevance of ethics in the progress of science and the moral and ethical quandry of embryonic stem cell research. PMID- 12134781 TI - On the credibility of 'Credibility of problem-solving therapy and medication for the treatment of depression among primary care patients' by Thornett A, Mynors Wallis, L. PMID- 12134782 TI - Impact of moderate aerobic exercise training on insulin sensitivity in type 2 diabetic men treated with oral hypoglycemic agents: is insulin sensitivity enhanced only in non-obese subjects? PMID- 12134783 TI - [Can a hydergine therapy lead to damage of the coronary vessels?]. PMID- 12134784 TI - Current issues in the management of kidney transplant patients. PMID- 12134785 TI - Hero. PMID- 12134786 TI - Collective bargaining agreements. PMID- 12134787 TI - Habitat for Humanity. PMID- 12134788 TI - No secrets on Main Street. PMID- 12134789 TI - Postcard from Copenhagen. PMID- 12134790 TI - Special issue: Symposium on "Morphology and Physiology of Semicircular Canal Ampulla". AB - This issue of Biological Sciences in Space includes papers presented at the symposium on "Morphology and Physiology of Semicircular Canal Ampulla" held in Nagoya, Japan, on March 25-27, 2001. The symposium was organized as part of the Japan-German Collaboration Project, a collaboration initiated in 1986 of the Japan Society for Promotion of Sciences and Deutsche Forschungs-gemeinschaft. PMID- 12134791 TI - AIDS council launches new advocacy network. PMID- 12134792 TI - Supreme Court. High Court says ADA's 'direct threat' includes threat to self. PMID- 12134793 TI - Advisory panel: expand hepatitis therapy to HIV/AIDS patients. PMID- 12134794 TI - Access to HIV drug programs loses pace as need grows. PMID- 12134795 TI - CDC updates opportunistic disease prevention guidelines. PMID- 12134796 TI - Discrimination. High Court turns down appeal in HIV direct threat case. PMID- 12134798 TI - Insurance. Viatical benefits payable despite AIDS patient's nondisclosure. PMID- 12134797 TI - Legislation. Congress moves to reduce HIV/AIDS transmission in prisons. PMID- 12134799 TI - Discrimination. Court upholds firing of worker who declined HIV/AIDS test. PMID- 12134800 TI - Bush proposes additional $500M in HIV/AIDS funding. PMID- 12134801 TI - Court rejects reduction of 20-year 'life' sentence for AIDS patient. PMID- 12134802 TI - Public health agency offers HIV/AIDS program funding. PMID- 12134804 TI - A new insect order. PMID- 12134803 TI - Product compliance. Drug company to go to trial over faulty AIDS testing kits. PMID- 12134805 TI - Professor Ottar Sjaastad. PMID- 12134807 TI - New procedure for selective extraction of polycyclic aromatic hydrocarbons in plants for gas chromatographic-mass spectrometric analysis. AB - A new solid-phase extraction method for the clean-up and the quantitation by GC MS of regulated polycyclic aromatic hydrocarbons (PAHs) from lettuce was developed and the experimental conditions were optimized. After ultrasonic extraction using toluene and saponification of samples, a clean-up of extracts through solid-phase extraction was performed. Samples were finally analyzed by gas chromatography-mass spectrometry (GC-MS) using an internal deuterated standard. Saponification by KOH in methanol-water (80:20) was successful allowing a good elimination of the interfering chlorophylls from the extracts containing the PAHs. The average recovery of the 16 regulated PAHs was 70, 74, 79 and 89%, respectively, for naphthalene, acenaphthylene, acenaphthene and chrysene and higher than 94% for the others. PMID- 12134808 TI - Aqueous chromatography utilizing hydrophobicity-modified anionic temperature responsive hydrogel for stationary phases. AB - A new pH-/temperature-responsive poly(N-isopropylacrylamide-co-acrylic acid-co-N tert-butylacrylamide) (poly(IPAAm-co-AAc-co-tBAAm)) hydrogel grafted on silica beads was evaluated as column matrix for a cation-exchange thermoresponsive chromatography. The stationary phase showed simultaneous changes in temperature responsive surface charge density and hydrophobicity by incorporation of anionic AAc and hydrophobic tBAAm into IPAAm sequences. Thermoresponsive polymer property alterations were confirmed by temperature-responsive phase transition and shift in apparent pKa values. Catecholamine derivatives were retained on poly(IPAAm-co AAc-co-tBAAm)-modified column at pH 7.0. Analyte retention was primarily due to the electrostatic interaction. It was noted that the temperature-induced phase transition of poly(IPAAm-co-AAc-co-tBAAm) hydrogel layer on the stationary phases was evidenced by the apparent inflection point in van't Hoff plots around 36 degrees C. This suggests that solute interactions should be changed below and above the stationary phase transition temperature, reducing electrostatic interaction above the transition temperature. PMID- 12134809 TI - Study of polar and nonpolar substituted benzenes and aromatic isomers on carbon coated zirconia and alkyl bonded phases. AB - Retention factors of polar and non-polar mono- and di-substituted benzene derivatives were measured on carbon coated zirconia (C/ZrO2) and an alkyl modified silica using water-acetonitrile mobile phases. Published data on porous graphitic carbon phases (PGC) were used to facilitate comparisons between the two types of carbon media. This work showed that retention on both C/ZrO2 and PGC is much more sensitive to the solute polarizability, dipolarity, and shape than on aliphatic phases. For simple disubstituted benzenes there was no general clear cut advantage in terms of chromatographic selectivity to using a carbon-based phase over a bonded phase silica; however, the selectivities towards such isomers are quite different on the two types of media. In contradistinction to their effect on alkyl bonded phase retention, addition of a dipolar substituent and weak hydrogen bond acceptor to a benzene ring almost always increases the solute's retention on C/ZrO2 and PGC. PMID- 12134810 TI - Evaluation of a penicillin G acylase-based chiral stationary phase towards a series of 2-aryloxyalkanoic acids, isosteric analogs and 2-arylpropionic acids. AB - The chiral recognition properties of a new chiral stationary phase based on immobilized penicillin G acylase were investigated using 35 acidic racemates. Twenty-seven compounds were resolved with high separation factors. The influences of mobile phase pH, type of organic modifier and ionic strength on enantioselective retention were studied. The most important tool for affecting the enantioselectivity was the mobile phase pH and interestingly the retention order of the enantiomers of some analytes could be controlled by this parameter. The analysis time for resolving enantiomers could be adjusted with a minor decrease in enantioselectivity using a high ionic strength mobile phase buffer while both retention and enantioselectivity decreased by adding organic modifier to the mobile phase. Displacement studies have demonstrated that the enzymatically active site and the chiral adsorption site overlap. PMID- 12134811 TI - Determination of polycyclic aromatic hydrocarbons and polycylic aromatic sulfur heterocycles by high-performance liquid chromatography with fluorescence and atmospheric pressure chemical ionization mass spectrometry detection in seawater and sediment samples. AB - Two methods for determining 10 polycyclic aromatic compounds were developed. Both methods were based on high-performance liquid chromatography (HPLC), but one method used fluorescence detection, while the other used atmospheric pressure chemical ionization mass spectrometry (APCI-MS). For water analysis, solid-phase extraction (SPE) was on-line coupled to the separation system. Using a styrene divinylbenzene copolymer (PLRP-s) as sorbent in the SPE and adding 20% of acetonitrile to the water sample before its preconcentration, recoveries were above 70% for most of the compounds. For the fluorescence method, all compounds were detected and six of them could be quantified at concentrations higher than 0.02 microg 1(-1). For the MS detection method, only seven of the compounds were detected and six were quantified at concentrations higher than 0.06 microg 1(-1). To analyse sediment samples, an extraction with dichloromethane was used and, due to the complexity of the matrix, a standard addition calibration was carried out. Seawater and sediment samples taken from the Tarragona fishing port and marina on the coast of Catalonia (Spain) were analysed, and five compounds (benzo[b]fluoranthene, benzo[k]fluoranthene, benzo[a]pyrene, benzo[ghi]perylene and indeno[1,2,3-cd]pyrene) were quantified in the sediment samples. PMID- 12134812 TI - Mass spectrometric determination of ergosterol in a prairie natural wetland. AB - Fungi are the main decomposers of plant material in an aquatic system. Levels of ergosterol, a compound generally specific to the cell membranes of fungi can be used as an indirect measure of their presence and biomass. Described is a procedure utilising reversed-phase liquid chromatography with positive-ion atmospheric pressure chemical ionization tandem mass spectrometry (LC-APCI-MS-MS) for full quantification and confirmation of ergosterol in various wetland matrices. Solid and liquid samples (0.2-1 g dry weight and 10 ml) were subjected to alkaline saponification followed by serial extraction using pentane (3 x 10 ml). The procedure was applicable to quantitative analysis of wetland samples with little or no clean up. Under low energy (CID) collision induced dissociation conditions the major product-ion formed from m/z 379.4 [M + H-H2O]+ was m/z 69.4 [(CH3)2CHCH=CH]+. Selected reaction monitoring (SRM) of this transition along with the retention time were used to confirm that ergosterol was widely distributed at ppm levels (2.4 to 303 microg ash free dry mass (AFDM)) in matrices of decaying willow leaves, Scirpus stems (living and dead) and sediment collected at the water-sediment interface. Comparison between LC-APCI-MS-MS (SRM), LC-APCI-MS using selected ion monitoring (SIM) and liquid chromatography with ultraviolet absorption detection (LC-UV) indicated that SRM analysis was the most selective technique. PMID- 12134813 TI - New chromatographic method for separation and determination of denatured alphaS1 , alphaS2-, beta- and kappa-caseins by hydrophobic interaction chromatography. AB - Separation and determination of denatured alpha-, beta- and kappa-caseins by hydrophobic interaction chromatography (HIC) was improved by using a TSK-Gel Ether-5PW column (Tosoh Biosep). The method, already proposed and performed by a TSK-Gel Phenyl-5PW column (Tosoh Biosep), is based on fast and easy solubilization of commercial and real samples by 4.0 M guanidine thiocyanate and HIC analysis in the presence of 8.0 M urea in the mobile phase. Employment of the less hydrophobic ether phase had the main advantage of separating casein fractions in less than 22 min and, additionally, of separating a-casein in kappaS1- and alphaS2 -casein fractions. The method has been validated by the analysis of reference skim milk powder (BCR-063R) certified for total nitrogen content. A linear relationship between the concentration of casein and peak area (UV absorbance detector at 280 nm) has been obtained over the concentration range of 0.5-40 microM. The detection limit for alpha-, beta- and kappa-caseins ranged between 0.33 and 0.65 microM. The precision of the method was evaluated, the RSDs for alphaS1-, alphaS2-, beta- and kappa-casein determination ranging between 2.3 and 5.5% for standard solutions and between 4.4 and 6.2% for real sample solutions. The mean value of casein content found in eight aliquots of BCR-063R calculated with respect to the total protein content (estimated on the basis of certified total nitrogen content) was 78.3 +/- 6.1%. Results of linear fitting of standard additions data of alphaS1-, alphaS2-, beta- and kappa-caseins to BCR 063R were compared with linear fitting of alphaS1-, alphaS2-, beta- and kappa casein calibration data. The method was applied to commercial caseins and to 30 real, raw samples. A statistical comparison was performed between results on quantitation of alpha-, beta- and kappa-caseins obtained by TSK-Gel Ether-5PW and TSK-Gel Phenyl-5PW HIC columns, showing more accurate results for chromatographic analysis performed by the ether column. PMID- 12134814 TI - Ion-pair reversed-phase high-performance liquid chromatography analysis of oligonucleotides: retention prediction. AB - An ion-pair reversed-phase HPLC method was evaluated for the separation of synthetic oligonucleotides. Mass transfer in the stationary phase was found to be a major factor contributing to peak broadening on porous C18 stationary phases. A small sorbent particle size (2.5 microm), elevated temperature and a relatively slow flow-rate were utilized to enhance mass transfer. A short 50 mm column allows for an efficient separation up to 30mer oligonucleotides. The separation strategy consists of a shallow linear gradient of organic modifier, optimal initial gradient strength, and the use of an ion-pairing buffer. The triethylammonium acetate ion-pairing mobile phases have been traditionally used for oligonucleotide separations with good result. However, the oligonucleotide retention is affected by its nucleotide composition. We developed a mathematical model for the prediction of oligonucleotide retention from sequence and length. We used the model successfully to select the optimal initial gradient strength for fast HPLC purification of synthetic oligonucleotides. We also utilized ion pairing mobile phases comprised of triethylamine (TEA) buffered by hexafluoroisopropanol (HFIP). The TEA-HFIP aqueous buffers are useful for a highly efficient and less sequence-dependent separation of heterooligonucleotides. PMID- 12134815 TI - Optimisation of a programmed split-splitless injector in the gas chromatographic mass spectrometric determination of organochlorine pesticides. AB - Large-volume injection techniques in gas chromatography are used to compensate for the at times limited detection sensitivity of mass spectrometric detection. In this work a programmed split-splitless injector in solvent split mode was employed to determine organochlorine pesticides in environmental samples. The injection conditions were selected by a Plackett-Burman design followed by a central composite design. The LODs obtained in the optimum conditions were compared with those obtained with splitless-MS and splitless-ECD. Finally, the method was applied to a soil sample. PMID- 12134816 TI - Temperature gradient interaction chromatography and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry analysis of air terminated polystyryllithium. AB - The reaction products of polystyryllithium with air were characterized by size exclusion chromatography, temperature gradient interaction chromatography and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Polystyryllithium was prepared by anionic polymerization of styrene initiated with sec-butyllithium in cyclohexane under an Ar atmosphere. It was confirmed that polystyryl ketone, polystyryl alcohol, and directly coupled polystyrene were the major products in addition to the normally terminated polystyrene, which is consistent with the results in the literature. We could also identify the presence of methoxy and carboxylic acid end capped polystyrenes as well as dipolystyryl ether as minor products. Among the minor products, dipolystyryl ether has not been reported yet. PMID- 12134817 TI - Simultaneous determination of ketoconazole and formaldehyde in a shampoo: liquid chromatography method development and validation. AB - Ketoconazole is an antifungal agent, which is the active ingredient in a shampoo primarily used for the treatment of seborrhatic dermatitis (anti-dandruff shampoo). The shampoo also contains imidazolidinylurea as a formaldehyde releasing preservative. The aim of this study was to develop a HPLC system that allows the determination of both ketoconazole and formaldehyde. The finally selected isocratic system consisted of an Interchrom Nucleosil (250 X 4.6 mm, 5 microm) C8 column and a mobile phase containing acetonitrile-phosphate buffer 0.025 M, pH 4.0, 45/55 (v/v). Ketoconazole could immediately be determined at 250 nm after injection of diluted shampoo. Formaldehyde was measured at 345 nm after derivatisation with a 2,4-dinitrophenylhydrazine solution. At the selected conditions, the other excipients of the shampoo did not interfere in the assays for both substances. Method validation was performed on both assays. Different selectivity towards ketoconazole and formaldehyde was observed when applying other C8 columns. This fact, however, did not affect the assays of both substances. PMID- 12134818 TI - High-resolution separation of polychlorinated biphenyls by comprehensive two dimensional gas chromatography. AB - Comprehensive two-dimensional gas chromatography (GC x GC) with micro electron capture detection (microECD) has been optimised for the separation of polychlorinated biphenyl congeners with emphasis on the separation of 12 toxic non- and mono-ortho chlorinated biphenyls (CBs), viz. CBs 77, 81, 105, 114, 118, 123, 126, 156, 157, 167, 169 and 189. The selection of the first- and second dimension columns and the temperature programme optimisation were carried out with a mixture of 90 CBs and the results are compared with those of one dimensional GC. A complete separation of all 12 priority CBs was obtained with two column combinations, HP-1-HT-8 and HP-1-SupelcoWax-10. With the HP-1-HT-8 column set, ordered structures show up in the two-dimensional plane, with the number of chlorine substituents and their position (ortho vs. non-ortho) being the main parameters of interest. This can help with congener identification. Estimated detection limits are excellent, i.e. about 10 fg. To illustrate the potential and the versatility of GC x GC-microECD, a cod liver extract and a standard mixture of the 17 most toxic polychlorinated dibenzo-p-dioxins and polychlorinated dibenzofurans together with 90 CBs were analysed as an application. PMID- 12134819 TI - Minimization of water vapor interference in the analysis of non-methane volatile organic compounds by solid adsorbent sampling. AB - Water vapor can be a significant interference in the analysis of air for non methane volatile organic compounds (NMVOCs) using solid-adsorbent sampling techniques. The adsorbent materials used in sampling cartridges have different hydrophobic characteristics, and it is therefore necessary to characterize solid adsorbent cartridges over a wide range of humidity. Controlled humidity experiments were performed to assess the extent of water vapor interference when samples are collected onto AirToxics solid-adsorbent cartridges. It was found that elevating the temperature of the cartridge to 10 degrees C above the temperature of the air sample greatly reduced water vapor adsorption and interferences and resulted in > or = 90% recovery of NMVOCs, biogenic VOCs and chlorofluorocarbons. Similar collection efficiencies were obtained at ambient temperature by reducing the relative humidity to > or = 60% in the sample by dilution with dry, scrubbed ambient air. A procedure also was developed and optimized for dry-purging cartridges prior to analysis. However, under optimized conditions, significant losses of C3-C5 compounds still occurred under highly humid conditions. It was determined that these losses were due to reduced retention during sampling rather than loss during the dry purge procedure. The dry purge method was shown to be adequate at high humidities for sampling NMVOCs with retention indices greater than 500. PMID- 12134820 TI - Automated headspace solid-phase dynamic extraction for the determination of amphetamines and synthetic designer drugs in hair samples. AB - The technique of automated headspace solid-phase dynamic extraction (SPDE) coupled with gas chromatography-mass spectrometry was evaluated for the determination of amphetamines and synthetic designer drugs in hair samples. Headspace SPDE is a novel method for the solventless extraction of organic compounds in aqueous samples. In a so-called inside needle capillary absorption trap a hollow needle with an internal coating of polydimethylsiloxane is used as extraction and preconcentration medium. Sampling is performed on the solution headspace by passing the gas through the device actively by a syringe. Analytes present in the sample are sorbed onto the deposited stationary phase. The syringe needle is placed into the injection port of a GC and rapid heating of the metal needle induces the desorption of analytes. For the determination of amphetamine, methamphetamine, 3,4-methylendioxyamphetamine (MDA), 3,4 methylendioxymethamphetamine, 3,4-methylendioxyethylamphetamine (MDEA), 3,4 methylendioxyphenyl-2-butanamine and N-methyl-1-(3,4-methylendioxyphenyl)-2 butanamine in human hair samples, 10 mg of hair were hydrolysed with sodium hydroxide. After absorption of analytes for an on-coating derivatization procedure the SPDE needle was directly placed into the headspace of a second vial containing N-methyl-bis(trifluoroacetamide). A validation procedure revealed absolute analyte recoveries between 10.2 and 16.7%. Linearity was obtained from 0.1 to 20 ng/mg with coefficients of correlation between 0.992 and 0.999. Intra- and inter-day precision were determined at two different concentrations and resulted in ranges between 1.4 and 4.1% (intra-day) and 4.2-14.6% (inter-day). Limits of detection between 0.03 ng/mg (MDA) and 0.19 ng/mg (MDEA) were achieved. Results indicated that SPDE is a rapid and sensitive method for the analysis of biological samples. Compared to solid-phase microextraction we found a higher extraction rate coupled with a faster automated operation. PMID- 12134822 TI - Thin-layer chromatography-matrix-assisted laser desorption ionisation-time-of flight mass spectrometry using particle suspension matrices. AB - Particle suspension matrices have been successfully utilized for the analysis of tetracycline antibiotics by thin-layer chromatography-matrix-assisted laser desorption ionisation-time-of-flight mass spectrometry (TLC-MALDI-TOF-MS). Particles of different materials and sizes have been investigated (Co-UFP, TiN, TiO2, Graphite and Silicon) by applying particle suspensions to eluted TLC plates. Mass spectra and mass chromatograms have been recorded directly from the TLC plates. Strong cationization by sodium and potassium was obtained in the positive ion mode, with [M+Na-NH3]+ ions being the predominant signals. The TLC MALDI mass spectra recorded from graphite suspensions showed the lowest background noise and the highest peak intensities from the range of suspension matrices studied. The mass accuracy from graphite films was improved by adding the peptide Phe-Phe to the graphite suspensions. This allowed internal recalibration of the TLC-MALDI mass spectra acquired during a run. One major potential advantage of TLC-MALDI-TOF-MS has been demonstrated in the analysis of chlortetracycline and tetracycline in a mixture of oxytetracycline, chlortetracycline, tetracycline and minocycline. Examination of the TLC plate prior to MALDI analysis showed only an unresolved spot for chlortetracycline and tetracycline. However by investigation of the MALDI mass spectra and plotting of single ion chromatograms separate peaks for chlortetracycline and tetracycline could be obtained. PMID- 12134821 TI - Nordic laboratory intercomparison of supercritical fluid extraction for the determination of total petroleum hydrocarbon, polychlorinated biphenyls and polycyclic aromatic hydrocarbons in soil. AB - Two developed supercritical fluid extraction (SFE) methods [one for the determination of total petroleum hydrocarbon (TPH) and polychlorinated biphenyls (PCBs), and one for polycyclic aromatic hydrocarbons (PAHs) and creosote components in soil] were evaluated in a Nordic laboratory intercomparison study with 11 participating laboratories. The interlaboratory comparison showed that excellent recoveries can be obtained with SFE for PAHs and PCBs compared to the solvent extraction. For the TPH, the recoveries were significantly higher than those achieved with solvent extraction. The accuracy, expressed as the relative standard deviation, was higher than expected (generally 8-25% for PAHs, 6-20% for PCBs and less than 18% for TPH with a few very high values, especially for PCBs), but not different from the other intercomparison studies. Difference between liquid- and solid-phase collection in SFE was found to be significant only for more volatile PAH components such as naphthalene and fluorene. For PCBs and TPH, there were some variation in the results obtained with the two trapping methods. PMID- 12134823 TI - Liquid chromatography-electrospray ionization isotope dilution mass spectrometry analysis of paraquat and diquat using conventional and multilayer solid-phase extraction cartridges. AB - The performance of alkyl-silica sorbent packed solid-phase extraction (SPE) cartridges and a mixed-mode, polymeric sorbent packed SPE cartridge (resin SPE cartridge) were evaluated for the sample preparation of paraquat and diquat in environmental water and vegetation matrices. Also the recoveries of the native and 2H-labeled paraquat and diquat were correlated to validate that the 2H labeled species can be used for the isotopic dilution mass spectrometry (IDMS) analysis of paraquat and diquat. The results show that the extraction efficiency of alkyl-silica SPE is dependent on the carbon loading of the sorbent and deteriorates with an increasing sample pH. The resin SPE cartridge required no pH adjustment and showed excellent correlation between the native and 2H-labeled species, therefore, allowing us to develop the first liquid chromatography electrospray ionization IDMS analytical method for the analysis of paraquat and diquat in environmental water and vegetation matrices. Method detection limits derived using standard EPA protocol were 0.2 and 0.1 microg/l for paraquat and diquat in water matrices, and 0.02 and 0.01 microg/g in vegetation matrices, respectively. PMID- 12134824 TI - On-line coupling of capillary isotachophoresis and capillary zone electrophoresis for the determination of flavonoids in methanolic extracts of Hypericum perforatum leaves or flowers. AB - Five flavonoids (hyperoside, isoquercitrin, quercitrin, quercetin and rutin) were separated and determined in extracts of Hypericum perforatum leaves or flowers by capillary zone electrophoresis (CZE) with isotachophoretic (ITP) sample pre treatment using on-line column coupling configuration. The background electrolyte (BGE) used in the CZE step was different from the leading and terminating ITP electrolytes but all the electrolytes contained 20% (v/v) of methanol. The optimal leading electrolyte was 10 mM HCl of pH* approximately 7.2 (adjusted with Tris) and the terminating electrolyte was 50 mM H3BO3 of pH* approximately 8.2 (adjusted with barium hydroxide). This operational system allowed to concentrate and pre-separate selectively the flavonoid fraction from other plant constituents before the introduction of the flavonoids into the CZE capillary. The BGE for the CZE step was 50 mM Tris buffer of pH* approximately 8.75 containing 25 mM N [tris(hydroxymethyl)methyl]-3-aminopropanesulfonic acid as co-ion and 55 mM H3BO3 as complex-forming agent. The ITP-CZE method with spectrophotometric detection at 254 nm was suitable for the quantitation of the flavonoids in real natural samples; kaempferol was used as internal standard. The limit of detection for quercetin-3-O-glycosides was 100 ng ml(-1) and calibration curves were rectilinear in the range 1-10 microg ml (-1) for most of the analytes. The RSD values ranged between 0.9 and 2.7% (n=3) when determining approximately 0.07-1.2% of the individual flavonoids in dried medicinal plants. PMID- 12134825 TI - Collection of alpha1-acid glycoprotein molecular species by capillary electrophoresis and the analysis of their molecular masses and carbohydrate chains. Basic studies on the analysis of glycoprotein glycoforms. AB - A highly heterogeneous glycoprotein, alpha1-acid glycoprotein, was resolved into their glycoforms by capillary electrophoresis using a surface-modified capillary in 20 mM acetate buffer (pH 4.2) containing 0.5% (w/v) hydroxypropylmethylcellulose. We collected the fractions containing each glycoform as nearly pure state by capillary electrophoresis, and examined the molecular masses of these glycoforms by matrix assisted laser desorption time-of flight mass spectrometry. We also analyzed carbohydrate chains after releasing them with N-glycosidase F followed by fluorescent labeling with 8-aminopyrene 1,3,6-trisulfonate. We found that the separation of glycoforms was mostly due to the presence of multiantennary carbohydrate chains. We propose that the present technique is useful for the analysis of post translational modification of proteins with carbohydrate chains. PMID- 12134826 TI - Separation and determination of emetine dithiocarbamate metal complexes by capillary electrophoresis with chemiluminescence detection of the tris(2,2' bipyridine)-ruthenium(II) complex. AB - Emetine dithiocarbamate metal complexes, which were prepared from emetine, carbon disulfide, and metal(II), indicated large chemiluminescence intensities on tris(2,2'-bipyridine)-ruthenium(II) chemiluminescence. Capillary electrophoresis with chemiluminescence detection was developed for analyzing emetine and the metal complexes. After the optimization of various analytical conditions, the mixture of the Cu(II), Ni(II), and Co(II) complexes as a model sample was injected into the capillary electrophoresis system with chemiluminescence detection. They were successfully separated and detected with a detection limit of 50 nM. PMID- 12134827 TI - Development of a validated capillary electrophoresis method for enantiomeric purity testing of dexchlorpheniramine maleate. AB - A capillary zone electrophoresis method has been developed for the detection of 0.1% of (R)-levochlorpheniramine maleate in samples of (S)-dexchlorpheniramine maleate. Using 1.5 mM carboxymethyl-beta-cyclodextrin in an acidic background electrolyte, resolution values of more than 10 were obtained. Under these conditions the R-enantiomer is migrating in front of the bulk S-enantiomer. The assay was validated for linearity (2-10 microg/ml; R2 = 0.9992), selectivity [(RS)-pheniramine maleate and (RS)-brompheniramine maleate], limit of detection (0.25 microg/ml), limit of quantification (0.75 microg/ml), analytical precision (intra- and inter-day variability), repeatability of the method (RSD = 5.0%) and accuracy. In samples of dexchlorpheniramine maleate from two different manufacturers, concentrations of, respectively, 0.15% and 1.95% (m/m) of levochlorpheniramine maleate were detected. The method was compared to the HPLC method described in the European Pharmacopoeia III monograph. PMID- 12134828 TI - Determination of thyroid hormones in pharmaceutical preparations, after derivatization with 9-anthroylnitrile, by high-performance liquid chromatography with fluorescence detection. AB - HPLC with fluorescence detection was used for the determination of low levels of liothyronine sodium and levothyroxine sodium in pharmaceutical preparations after fluorogenic derivatization. 9-Anthroylnitrile in dimethyl sulfoxide was used as a precolumn fluorogenic reagent. The 9-anthroylnitrile derivatives of liothyronine sodium and levothyroxine sodium were separated on a reversed-phase column with acetonitrile-0.02 M sodium dodecylsulfate (pH 3.5 with phosphoric acid) as the eluent. The calibration graphs were linear over a sample concentration range of 0.25-2.5 microg/ml. The detection limits for liothyronine sodium and levothyroxine sodium were 0.2 ng per injection. The proposed method was applied to the determination of thyroid hormones in pharmaceutical preparations. PMID- 12134830 TI - Extended thermodynamic approach to ion interaction chromatography: a thorough comparison with the electrostatic approach, and further quantitative validation. AB - The most reliable literature experimental results, concerning retention behavior of charged molecules, in the presence of an ion-interaction reagent (IIR), were used to obtain a further quantitative validation of a new theory. The present work emphasizes the fact that the extent to which electrostatic interactions, ion pair formation in the adsorbed and the mobile phases, and adsorption competitions are one more important than the other depends on experimental conditions. Further insight into the meaning of the linearity of the log k vs. log [IIR] plot, which is common to many theoretical models, is given. The experimental conditions under which the linearity of this plot can be expected not only practically, but also theoretically, are elucidated. The dependence of the ratio of retention factors with and without IIR in the eluent on the analyte nature, which cannot be predicted by the electrostatic approach, was explained and tracked. The difference between the actual surface potential and that predicted by the electrostatic approach is also rationalized. The model is also theoretically shown to be able to elucidate the enantioselective retention mechanism, in the presence of chiral counter ions. PMID- 12134829 TI - Peak deconvolution in one-dimensional chromatography using a two-way data approach. AB - A deconvolution methodology for overlapped chromatographic signals is proposed. Several single-wavelength chromatograms of binary mixtures, obtained in different runs at diverse concentration ratios of the individual components, were simultaneously processed (multi-batch approach), after being arranged as two-way data. The chromatograms were modelled as linear combinations of forced peak profiles according to a polynomially modified Gaussian equation. The fitting was performed with a previously reported hybrid genetic algorithm with local search, leaving all model parameters free. The approach yielded more accurate solutions than those found when each experimental chromatogram was fitted independently to the peak model (single-batch approach). The improvement was especially significant for those chromatograms where the peaks were severely affected by the tails of the preceding compounds. Peak shifts among chromatograms, which are a usual source of non-bilinearity, were modelled in a continuous domain instead of in a discrete way, which avoided some drawbacks associated with latent variable methods. An experimental design involving simulated chromatograms was applied to check the method performance. Five main factors affecting the deconvolution were examined: concentration pattern, chromatographic resolution, number of batches and replicates, and noise level, which were evaluated using first- and second order figures of merit. The method was also tested on three real samples containing compounds showing different overlap. Four multi-batch deconvolution methods were considered differing in the nature of the processed information and kind of peak matching among chromatograms. In all cases, the multi-batch deconvolution yielded better performance than the single-batch approach. PMID- 12134831 TI - Determination of the dissociation constants (pKa) of basic acetylcholinesterase inhibitors by reversed-phase liquid chromatography. AB - An RP-HPLC study for the pKa determination of a series of basic compounds related to caproctamine, a dibenzylaminediamide reversible inhibitor of acetylcholinesterase, is reported. The 2-substituted analogues, bearing substituents with different electronegativity, were analysed by RP-HPLC by using C18 C4 stationary phases with a mobile phase consisting of mixture of acetonitrile and triethylamine phosphate buffer (pH range comprised between 4 and 10). Typical sigmoidal curves were obtained, showing the dependence of the capacity factors upon pH. In general, the retention of the investigated basic analytes increased with increasing of the pH. The inflection point of the pH sigmoidal dependence was used for the dissociation constant determination at a fixed acetonitrile percentage. When plotting pKa vs. percent of acetonitrile in the mobile phase for two representative compounds, linear regression were obtained: the y intercept gave the aqueous pKa(w). The pKa estimation by HPLC method was found to be useful to underline the difference of benzylamine basicity produced by the ortho aromatic substituents. The variation of pKa values (6.15 7.80) within the series of compounds was correlated with the electronic properties of the ortho-substituents through the Hammett sigma parameter, whereas the ability of substituents to accept H-bond was found to play a role in determining the conformational behavior of the molecules. PMID- 12134832 TI - Unusual effect of column temperature on chromatographic enantioseparation of dihydropyrimidinone acid and methyl ester on amylose chiral stationary phase. AB - This paper reports an unusual effect of column temperature on the separation of the enantiomers of dihydropyrimidinone (DHP) acid and its methyl ester on a derivatized amylose stationary phase by normal-phase liquid chromatography. The separation of the DHP acid enantiomers was investigated using both carbamate derivatized amylose and cellulose stationary phases (Chiralpak AD and Chiralcel OD) with an ethanol-n-hexane (EtOH-n-Hex) mobile phase. On the amylose phase, the van 't Hoff plot of the retention factor of the S-(+)-DHP acid was observed to be non-linear while that of R-(-)-DHP acid was linear. Likewise, the van 't Hoff plot for DHP acid enantioselectivity was non-linear with a transition occurring at approximately 30 degrees C. Furthermore, the van 't Hoff plot for the DHP acid enantioselectivity factor for data taken when heating the column from 5 to 50 degrees C was not superimposable with the same plot prepared with data from the cooling process from 50 to 5 degrees C. This observation suggested that the stationary phase was undergoing a thermally induced irreversible conformational change that altered the separation mechanism between the heating and cooling cycles. Similar phenomena were observed for the separation of the enantiomers of the DHP ester probe compound. The conformational change of the AD phase was shown to depend on the polar component of the mobile phase. When 2-propanol (2-PrOH) was used as the modifier instead of EtOH, the van 't Hoff plots for DHP acid were linear and thermally reversible, suggesting that no such irreversible conformational change occurs with this modifier. Conversely, when the AD phase was pre-conditioned with a more polar methanol (MeOH) or water containing mobile phase, thermal irreversibility of DHP acid enantioselectivity was once again observed. Interestingly, when the stationary phase was changed to its cellulose analogue, the Chiralcel OD, all van 't Hoff plots for the retention and selectivity of DHP acid were thermally reversible for both EtOH-n-Hex and 2-PrOH n-Hex mobile phases. PMID- 12134833 TI - Deferred standards, an on-line qualification, validation and system stability probe for chromatographic assay. AB - Specific programming of automated HPLC systems allows total on-line qualification, validation and stability monitoring using the concept of deferred standards. Setting up such a process for routine analyses in an automated HPLC system requires specific autosampler programming as well as specific monitoring software. With an autosampler, a double injection procedure is programmed, the first introducing the sample, and the second, a few minutes deferred, the deferred control standard. Two additional compounds are therefore added to the sample before and during the chromatographic process: the intemal standard for sample quantification and the deferred standard for system control. Specific methodologies are described of how to obtain classical quantitative analysis information as well as system qualification validation stability information. Experiments were performed to develop specified methodologies to monitor the quality of quantitative analysis during the life of the column by using the deferred standard concept to probe the effects of column ageing on separation characteristics. PMID- 12134834 TI - Effects of temperature on retention of chiral compounds on a ristocetin A chiral stationary phase. AB - The isocratic retention of enantiomers of chiral analytes, i.e. tryptophan, 1,2,3,4-tetrahydroisoquinoline and gamma-butyrolac tone analogs, was studied on a ristocetin A chiral stationary phase at different temperatures and with different mobile phase compositions, using the reversed-phase, polar-organic and normal phase modes. By variation of the both mobile phase composition and the temperature, baseline separations could be achieved for these enantiomers. The retention factors and selectivity factors for the enantiomers of all investigated compounds decreased with increasing temperature. The natural logarithms of the retention factors (ln k) of the investigated compounds depended linearly on the inverse of temperature (1/T). van't Hoff plots afforded thermodynamic parameters, such as the apparent change in enthalpy (deltaH(o)), the apparent change in entropy (deltaS(o)) and the apparent change in Gibbs free energy (deltaG(o) ) for the transfer of analyte from the mobile to the stationary phase. The thermodynamic parameters (deltaH(o), deltaS(o) and deltaG(o)) were calculated in order to promote an understanding of the thermodynamic driving forces for retention in this chromatographic system. PMID- 12134835 TI - Online coupling of pressurized liquid extraction, solid-phase extraction and high performance liquid chromatography for automated analysis of proanthocyanidins in malt. AB - A new instrumental setup for automated extraction of solid samples by online coupling of pressurized liquid extraction, automated SPE (solid-phase extraction) and HPLC is presented. From the extraction to the chromatogram no manual sample handling is required. The application to the determination of proanthocyanidins in malt reduces time and manual work to a minimum compared to former manual methods. Twenty samples can be processed within 24 h in respect to eight samples with the manual method. Using the features of the instrumental coupling, an optimized strategy for SPE of proanthocyanidins from natural samples was developed, requiring no evaporation step, using commercial cartridges and delivering concentrated eluates. The recovery of five main malt proanthocyanidins was 97%, with a reproducibility of 5%. This new instrumental coupling is thought to reduce time and costs along with improved results for a broad range of solid sample materials. PMID- 12134836 TI - Frequent p16/MTS1 inactivation in early stages of urothelial carcinoma of the bladder is not associated with tumor recurrence. PMID- 12134837 TI - Biologic differences between noninvasive papillary urothelial neoplasms of low malignant potential and low-grade (grade 1) papillary carcinomas of the bladder. PMID- 12134838 TI - Influence of the microenvironment on the invasiveness of human bladder carcinoma cell lines. PMID- 12134839 TI - Prognostic significance of platelet-derived endothelial cell growth factor/thymidine phosphorylase expression in stage pT1 G3 bladder cancer. PMID- 12134840 TI - Tumor progression and survival in patients with T1G3 bladder tumors: multicentric retrospective study comparing 94 patients treated during 17 years. PMID- 12134841 TI - Biodistribution of hypericin in orthotopic transitional cell carcinoma bladder tumors: implication for whole bladder wall photodynamic therapy. PMID- 12134842 TI - Acute care is "suboptimal," says Royal College of Physicians. PMID- 12134843 TI - Deaths from volatile substance misuse fall. PMID- 12134844 TI - Scientists manage to manufacture polio virus. PMID- 12134845 TI - Prison doctor should not have worked unsupervised, says inquiry. PMID- 12134847 TI - Chancellor maps out three year public service spending plans. PMID- 12134848 TI - US may require insurance cover for bowel cancer screening. PMID- 12134849 TI - US council calls for four year ban on cloning. PMID- 12134850 TI - Botox party at Johns Hopkins comes under fire. PMID- 12134852 TI - Commentary: Attitudes to women's bicycling have changed. PMID- 12134853 TI - Are selective COX 2 inhibitors superior to traditional NSAIDs? Pharmacia's response to editorial. PMID- 12134854 TI - Are selective COX 2 inhibitors superior to traditional NSAIDs? Little is known about COX 2 inhibitors. PMID- 12134855 TI - Are selective COX 2 inhibitors superior to traditional NSAIDs? Both the CLASS and VIGOR trials support the COX 2 hypothesis. PMID- 12134856 TI - NHS Direct audited. Audit neglected children. PMID- 12134857 TI - NHS Direct audited. Telephone consultations in general practice should be tested. PMID- 12134858 TI - NHS Direct audited. Telephone triage in Western Australia is cheaper than NHS Direct. PMID- 12134859 TI - Nurses as NHS gatekeepers. Using nurses might influence developing countries. PMID- 12134860 TI - Can nurse practitioners provide equivalent care to GPs? More studies of these nurses' technique are needed. PMID- 12134861 TI - Can nurse practitioners provide equivalent care to GPs? Nurse practitioners increase access to quality health care for many patients. PMID- 12134863 TI - Marine science. Researchers plunge into debate over new sub. PMID- 12134862 TI - Can nurse practitioners provide equivalent care to GPs? More methodologically sound investigations are needed. PMID- 12134864 TI - The future of UNSCEAR. PMID- 12134866 TI - Are U.S. patents too broad? PMID- 12134865 TI - Everything old is new again? PMID- 12134867 TI - Einstein and the orbit of Mercury. PMID- 12134868 TI - Microbial eukaryote species. PMID- 12134869 TI - Transcription-dependent R-loop formation at mammalian class switch sequences. PMID- 12134870 TI - Large study links nurse staffing and improved hospital outcomes. PMID- 12134871 TI - Computerized monitoring systems helps reduce adverse drug events. PMID- 12134872 TI - VHA heart attack initiative increases aspirin and beta blocker usage. PMID- 12134873 TI - Proposed web error reporting system could streamline data collection. PMID- 12134874 TI - The National Center for Biotechnology Information. PMID- 12134875 TI - Support for family planning improves women's lives. PMID- 12134876 TI - Abortion in context: United States and worldwide. PMID- 12134877 TI - U.S. policy can reduce cost barriers to contraception. PMID- 12134878 TI - Adolescent care standards and state CHIP efforts. PMID- 12134879 TI - Minors and the right to consent to health care. PMID- 12134880 TI - School-based health centers and the birth control debate. PMID- 12134881 TI - Welfare law and the drive to reduce 'illegitimacy'. PMID- 12134882 TI - Revisiting public funding of abortion for poor women. PMID- 12134883 TI - State responses to substance abuse among pregnant women. PMID- 12134884 TI - Title X: three decades of accomplishment. PMID- 12134885 TI - Sex education: politicians, parents, teachers and teens. PMID- 12134886 TI - Challenges facing family planning clinics and Title X. PMID- 12134887 TI - Expanding eligibility and outreach under CHIP. PMID- 12134888 TI - State-level policies on sexuality, STD education. PMID- 12134889 TI - Community health centers and family planning. PMID- 12134890 TI - Teen pregnancy: trends and lessons learned. PMID- 12134891 TI - Women and societies benefit when childbearing is planned. PMID- 12134892 TI - Family planning can reduce high infant mortality levels. PMID- 12134893 TI - Why are Medi-Cal rates important? PMID- 12134894 TI - The financial health of the Medi-Cal managed care market. PMID- 12134895 TI - Expanding Healthy Families to parents: the role of employer coverage. PMID- 12134896 TI - Medi-Cal after welfare reform: enrollment among former welfare recipients. PMID- 12134897 TI - The Medi-Cal budget: cost drivers and policy considerations. PMID- 12134898 TI - Craniopharyngioma. PMID- 12134899 TI - Ischemic depolarization monitoring: evaluation of protein synthesis in the hippocampal CA1 after brief unilateral ischemia in a gerbil model. AB - OBJECT: The authors investigate whether depolarization monitoring is an accurate index of ischemic damage in a gerbil model of unilateral ischemia and assess the effects of brief cerebral ischemia on protein synthesis in this model. METHODS: The authors evaluate the relationship between the duration of ischemic depolarization caused by unilateral carotid artery occlusion and ischemia-induced neuronal damage in the CA1 subregion 7 days after ischemia. When the depolarization period exceeded 210 seconds, some neuronal damage was detected, and almost complete neuronal damage was observed when the period exceeded 400 seconds. Uptake of [14C]valine was evaluated in ischemic and nonischemic CA1 subregions. Disturbances in protein synthesis were seen in all animals subjected to sublethal ischemia (< or = 210-second depolarization) after a 10-minute recirculation, and after 2 and 6 hours of recirculation in animals with 90 seconds or more of depolarization. Inhibition of protein synthesis was proportional to the length of the depolarization period. After 1 and 3 days of recirculation, protein synthesis returned to near normal, and some animals with depolarizations greater than 180 to 210 seconds showed an increase in protein synthesis. Protein synthesis in all animals returned to normal levels after 7 days of recirculation. CONCLUSIONS: In this study the authors demonstrate that monitoring of ischemic depolarization is a useful method to predict neuronal damage in the hippocampal CA1 in this model, and they identify subtle changes in protein synthesis after brief ischemia. Sublethal ischemia was divided into three categories by its depolarization period (< 90 seconds, 90-180 seconds, and > 180 210 seconds) with regard to changes in protein synthesis. PMID- 12134900 TI - Modulation of phorbol ester-induced regulation of matrix metalloproteinases and tissue inhibitors of metalloproteinases by SB203580, a specific inhibitor of p38 mitogen-activated protein kinase. AB - OBJECT: Expression of matrix metalloproteinases (MMPs) has been postulated to play a central role in brain tumor invasion; however, its underlying mechanism is not yet fully understood. In the present study, by assessing the effect of a specific p38 mitogen-activated protein kinase (MAPK) inhibitor, SB203580, on the secretion of MMPs and in vitro invasion of various glioma cells, the authors attempt to define the role of the p38 MAPK pathway in the regulation of MMPs and tissue inhibitors of metalloproteinases (TIMPs) activated by phorbol ester (phorbol-12-myristate-13-acetate [PMA]) in the D54 human glioblastoma cell line. METHODS: The activation of MAPKs was determined using Western blot analysis after addition of phospho-specific antibodies against these kinases, the status of MMPs and TIMPs was analyzed using gelatin zymography and Western blot analysis, and the invasion rate of D54 cells and other glioma cells was analyzed using a modified Boyden chamber assay. Treatment of D54 cells with PMA activated two distinct MAPKs, extracellular signal-regulated kinase (ERK) 1/2 and p38 MAPK, but not c-Jun N-terminal kinase/stress-activated protein kinase. Induction of MMP-9 production and MMP-2 activation by PMA were blocked by SB203580, a specific inhibitor of p38 MAPK, but not by PD98059, a specific inhibitor of ERK 1/2. In addition, PMA-induced downregulation of TIMP-1 and TIMP-2 secretion and upregulation of the membrane type I MMP, a major activator of MMP-2 on the cell surface, were reversed by SB203580 in these cells; the PMA-induced increase of invasion in vitro decreased when SB203580 was added to the top compartment of a modified Boyden chamber; and the inhibitor also reduced the MMP secretion and PMA induced in vitro invasion in various glioma cell lines. CONCLUSIONS: These results indicate that activation of p38 MAPK by PMA plays a central role in the regulation of MMPs and TIMPs in D54 cells, which has a major influence in tumor invasion and metastasis. Furthermore, inhibition of p38 MAPK by SB203580 blocked the secretion of MMPs and in vitro invasion of various glioma cells, underscoring a possible role of p38 MAPK inhibitors as antiinvasive and/or antimetastatic agents of malignant gliomas. PMID- 12134901 TI - Failure of single-unit neuronal activity to differentiate globus pallidus internus and externus in Parkinson disease. AB - OBJECT: The authors examine the validity of single-unit neuronal recordings as a method of differentiating the globus pallidus internus (GPi) from the GP externus (GPe) in Parkinson Disease. METHODS: One hundred twenty-eight recordings of apparent single-unit activity used to help guide final electrode placement in eight patients who underwent pallidotomy were analyzed using sophisticated spike sorting methods, and 185 neurons were characterized for mean firing frequency and percent of firing within bursts. In addition, the total spectral power was calculated on the full measured waveform for each of 128 samples without spike sorting. No correlation was identified between these measures of neuronal activity and depth within the GP. CONCLUSIONS: These results call into question the validity of relying on single-unit activity and microelectrode recordings in the operating room to localize lesion or electrode placement within the GPi during stereotactic pallidal surgery. PMID- 12134902 TI - Primary meningiomas of the jugular fossa. AB - OBJECT: Primary jugular fossa meningomas (JFMs) are one of the rarest subgroups of meningioma, with fewer than 40 cases reported in the literature. The authors retrospectively analyzed the results of surgical treatment in their series of patients, including clinical, pathological, and complication features. The surgical approach was mandated by the pathological anatomy of the tumor as well as by the anatomy of the individual patient. METHODS: During a 6.5-year period, the authors performed nine surgeries in eight patients (seven women [88%] and one man [12%]) with JFMs. Six lesions occurred on the right side and two on the left. The most common presenting symptoms were altered hearing in five patients (62%), swallowing difficulties in four patients (50%), and a deficit of the 11th cranial nerve in three patients (38%); a combination of two or more signs or symptoms was common. The surgical approach was tailored to the local anatomy (tumor neurovascular relationships) found in each patient; three different routes were used. Radical tumor removal was achieved in all patients; one tumor recurrence occurred after 20 months in a patient in whom the tumor had displayed atypical histological features. This woman underwent a second operation. The mean length of hospital stay was 1 week. The mean and the median follow-up period were 45 and 40 months, respectively. The most common complications were transient lower cranial nerve deficits, which resolved or were compensated for in all patients within 1 month. CONCLUSIONS: With a careful, extensive preoperative evaluation and appropriate tailoring of the operative approach, JFMs can be radically resected with the expectation of good outcome. PMID- 12134903 TI - Suramin-induced reversal of chronic cerebral vasospasm in experimental subarachnoid hemorrhage. AB - OBJECT: The naphthylsulfonate derivative suramin is an inhibitor of growth factor receptors (receptor tyrosine kinases) and G protein-coupled P2Y receptors. Both types of these receptors are suspected of being involved in cerebral vasospasm after subarachnoid hemorrhage (SAH). In the current study, the authors examined the therapeutic effects of suramin and a selective P2X-receptor antagonist, pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid (PPADS), in the reversal of vasospasm in an established canine double-hemorrhage model. METHODS: Twenty-four dogs underwent double blood injection into the cisterna magna, with injections given on Days 0 and 2. The dogs were divided randomly into three groups (six animals in each group) to be treated from Days 2 through 6 with the vehicle dimethyl sulfoxide, suramin, or PPADS. An additional group of six dogs received double blood injection without any treatment and served as an SAH control group. The animals were killed on Day 7. Angiography was performed on Day 0 before blood injection and again on Day 7 before the animals were killed. After the death of the animals, the basilar arteries (BAs) were collected for morphological studies and determination of tyrosine kinase expression, and the bloody cerebrospinal fluid (CSF) produced by the hemorrhages was collected for measurement of oxyhemoglobin and adenosine triphosphate (ATP). In the SAH control group, the mean diameter of the BAs on Day 7 was 46.23 +/- 6.32% of the value on Day 0 (which served as a reference of 100%). In the DMSO-treated group, the mean residual diameter of the BA was 47.77 +/- 0.8% on Day 7 compared with the value on Day 0. Suramin, but not PPADS, increased the residual diameter to 74.02 +/- 4.24% on Day 7. On Day 7 the level of ATP in the CSF was decreased and the level of oxyhemoglobin was increased, compared with values measured on Day 0. Suramin, but not PPADS, reduced tyrosine phosphorylation in the spastic BAs. CONCLUSIONS: By reducing tyrosine kinase activity, suramin may be useful in the treatment of cerebral vasospasm. PMID- 12134904 TI - Posttreatment with adenovirus-mediated gene transfer of calcitonin gene-related peptide to reverse cerebral vasospasm in dogs. AB - OBJECT: Gene transfer to cerebral vessels is a promising new therapeutic approach for cerebral vasospasm after subarachnoid hemorrhage (SAH). This study was undertaken to explore whether a delayed treatment with adenovirus encoding the prepro-calcitonin gene-related peptide (CGRP), 2 days after initial blood injection, reduces cerebral vasospasm in a double-hemorrhage model of severe vasospasm in dogs. METHODS: In 20 dogs, arterial blood was injected into the cisterna magna on Days 0 and 2. Thirty minutes after the second blood injection, the animals received either adenovirus encoding the prepro-CGRP gene (AdCMVCGRP treated group, eight dogs) or adenovirus encoding the beta-galactosidase gene (AdCMVbeta gal-treated group, six dogs) under the cytomegalovirus (CMV) promoter. One group of dogs did not receive treatment and served as controls (control SAH group, six dogs). Angiography was performed on Days 0 and 7 to assess cerebral vasospasm. On Day 7 following angiography, the animals were killed and their brains were stained with X-gal to detect the distribution of gene expression. Cerebrospinal fluid (CSF) was also tested for CGRP immunoreactivity. Severe vasospasm was observed in control SAH dogs on Day 7, and the mean basilar artery (BA) diameter was 53.4 +/- 5.5% of the value measured on Day 0. Treatment with AdCMVbeta gal did not alter vasospasm (the BA diameter was 55 +/- 3.9% of that measured on Day 0). The leptomeninges and adventitia of the BAs of dogs treated using AdCMVbeta gal demonstrated positive staining with X-gal. High levels of CGRP were measured in CSF from dogs that received AdCMVCGRP. In the group treated with AdCMVCGRP, vasospasm was significantly reduced (the BA diameter was 78.2 +/- 5.3% of that measured on Day 0, p < 0.05 compared with the control SAH group and the AdCMVbeta gal group). CONCLUSIONS: In a model of severe vasospasm in dogs, gene transfer of CGRP after injection of blood attenuated cerebral vasospasm after SAH. PMID- 12134905 TI - Interaction between p53 and p16 expressed by adenoviral vectors in human malignant glioma cell lines. AB - OBJECT: Multiple gene replacements have been examined as a potential treatment modality for malignant gliomas. Nevertheless, no reports are available that detail the synergy, additivity, or antagonism of multiple genes. The aim of this study was to assess the interaction between p53 and p16 genes in the growth of glioma cell lines. METHODS: The human glioma cell lines U87MG and U373MG were transduced using an adenoviral vector with Ad-p53, Ad-p16, or both. Western blotting was performed to determine the expression of the protein products of the transduced p53 and p16 genes. To establish whether the combination of Ad-p53 and Ad-p16 would be beneficial, the effects of gene combinations at the median inhibitory concentration level were analyzed using the isobologram method. Annexin assays and cell cycle analyses were performed on the transduced cells. Western blotting demonstrated the expression of p53 and p16 in transduced cells. Simultaneous exposure to Ad-p53 and Ad-p16 produced additive effects in both glioma cell lines. Experimental data points in U373MG lay near the Mode I line, indicating that the vectors had a different mode of action. The restoration of normal p53-encoded protein in the mutant cell lines induced apoptosis, whereas in the wild-type p53 cell lines, the overexpression of wild-type p53 resulted in a moderate degree of apoptosis and G1 arrest. Furthermore, Ad-p16 induced more marked G1 arrest than Ad-p53 in cells with wild-type p53. CONCLUSIONS: The results show that interaction between Ad-p53 and Ad-p16 is additive, regardless of p53 gene status. PMID- 12134906 TI - Anatomical study of the orbitozygomatic transsellar-transcavernous-transclinoidal approach to the basilar artery bifurcation. AB - OBJECT: An anatomical study in which measurements were obtained was undertaken to demonstrate that the orbitozygomatic transcavernous-transclinoidal approach provides excellent exposure of the trunk of the basilar artery (BA) and its bifurcation. METHODS: Bilateral stepwise dissections were performed on 10 fixed cadaver heads with the aid of x 3 to x 40 magnifications. A frontotemporal craniotomy was made, followed by an orbitozygomatic osteotomy. After the dura mater had been opened, the sylvian fissure was widely separated. The anteromedial triangle of the cavernous sinus was opened to mobilize the internal carotid artery medially. The sella turcica and the dorsum sellae were exposed. The posterior clinoid process and the dorsum sellae were drilled to expose a length of BA that included its bifurcation. Measurements were obtained following the frontotemporal craniotomy, orbitozygomatic osteotomy, and drilling of the posterior clinoid process to quantify the exposures provided by these procedures. Excellent exposure of the trunk of the BA and its bifurcation was achieved. The structures in the interpeduncular cistern and the prepontine cistern were also exposed. There was an average gain of a 13.4-mm-long segment of the BA, which in some surgeries can be invaluable. The angle of exposure that was achieved with the BA bifurcation located at the apex increased markedly. Moreover, this method widened the oculomotor nerve-carotid artery corridor for easier access to the BA bifurcation region. CONCLUSIONS: This approach combines the advantages granted by most conventional approaches to aneurysms of the BA bifurcation. The approach is suitable for aneurysms situated at a high, normal, or low position on the BA bifurcation. It exposes a sufficient length of the BA trunk to place a temporary clip. PMID- 12134907 TI - Promotion of regeneration of corticospinal tract axons in rats with recombinant vascular endothelial growth factor alone and combined with adenovirus coding for this factor. AB - OBJECT: After spinal cord transection in adult rats, the axons of the corticospinal tract (CST) degenerate retrogradely and do not regenerate. This phenomenon is thought to be related to either secondary ischemia or deficiency of growth factors. To overcome the deficiency of both blood flow and growth factors, the authors added exogenous vascular endothelial growth factor (VEGF165) to the transected spinal cord either as recombinant protein alone or combined with an adenovirus coding for VEGF165. Because most growth factors are rapidly inactivated in the extracellular environment, the authors used an adenovirus coding for VEGF165 to maintain its activity for several days. METHODS: In adult rats, the dorsal two thirds of the spinal cord were transected at the T-8 level. In experimental rats, either human recombinant VEGF165 or a combination of this factor and a replication-defective adenovirus coding for VEGF165 (Ad.CMV.VEGF165) was applied at the lesion site. Both recombinant VEGF165 alone and combined with Ad.CMV.VEGF165 were mixed with Matrigel, which is a reconstituted membrane basement protein extract. Control rats received Matrigel alone or Matrigel plus an adenoviral vector containing the LACZ gene (Ad.CMV.LACZ). Thirty days after spinal cord injury, the number of newly formed blood vessels was assessed in the injured area. In addition, the sensorimotor cortex was injected with anterogradely transported horseradish peroxidase (HRP) to label the CST axons in the spinal cord and to evaluate the extent of retrograde axonal degeneration and regeneration. Gene transfer was assessed using semiquantitative reverse transcription-polymerase chain reaction analysis, enzyme-linked immunosorbent assay for human VEGF and beta-galactosidase expression in injured rats treated with Matrigel plus Ad.CMV.LACZ, Matrigel plus Ad.CMV.VEGF165, and untreated injured rats. A strong gene transfer in the spinal cord tissue of adenovirus treated rats was found from Day 3 to Day 10 postinjury, confirming infection. In the injured spinal cord area, a significant increase of blood vessels (300% over control, p < 0.005) occurred both in rats treated with recombinant VEGF165 alone and in those treated with the combination of recombinant VEGF165 and Ad.CMV.VEGF165. Also, in both of these groups of animals the retrograde degeneration of CST axons was significantly reduced compared with rats treated with Matrigel alone or Matrigel plus Ad.CMV.LACZ. Furthermore, in rats treated with recombinant VEGF165 alone or combined with Ad.CMV.VEGF165, a few HRP-labeled CST axons, which were not detectable in control rats, were seen distal to the spinal cord injury, indicating some regeneration across the injured area. CONCLUSIONS: These results indicate that locally applied VEGF exerts angiogenic as well as neurotrophic effects in the injured spinal cord of rats. PMID- 12134908 TI - Invasion of human glioma biopsy specimens in cultures of rodent brain slices: a quantitative analysis. AB - OBJECT: The reliable assessment of the invasiveness of gliomas in vitro has proved elusive, because most invasion assays inadequately model in vivo invasion in its complexity. Recently, organotypical brain cultures were successfully used in short-term invasion studies on glioma cell lines. In this paper the authors report that the invasiveness of human glioma biopsy specimens directly implanted into rodent brain slices by using the intraslice implantation system (ISIS) can be quantified with precision. The model was first validated by the demonstration that, in long-term studies, established glioma cells survive in the ISIS and follow pathways of invasion similar to those in vivo. METHODS: Brain slices (400 microm thick) from newborn mice were maintained on millicell membranes for 15 days. Cells from two human and one rodent glioblastoma multiforme (GBM) cell lines injected into the ISIS were detected by immunohistochemistry or after transfection with green fluorescent protein-containing vectors. Preferential migration along blood vessels was identified using confocal and fluorescent microscopy. Freshly isolated (< or = 24 hours after removal) 1,1'-dioctadecyl 3,3,3',3'-tetramethylindocarbocyanine perchlorate-prelabeled human glioma biopsy specimens were successfully implanted in 19 (83%) of 23 cases, including 12 GBMs and seven lower grade gliomas (LGGs). Morphometric quantification of distance and density of tumor cell invasion showed that the GBMs were two to four times more invasive than the LGGs. Heterogeneity of invasion was also observed among GBMs and LGGs. Directly implanted glioma fragments were more invasive than spheroids derived from the same biopsy specimen. CONCLUSIONS: The ISIS combines a high success rate, technical simplicity, and detailed quantitative measurements and may, therefore, be used to study the invasiveness of biopsy specimens of gliomas of different grades. PMID- 12134909 TI - Diagnostic significance of soluble c-kit in the cerebrospinal fluid of patients with germ cell tumors. AB - OBJECT: Overexpression of the protooncogene c-kit has been suggested in a gonadal germ cell tumor (GCT). Recently, the soluble isoform of c-kit (s-kit) has been expressed in a variety of cell types. The goal of this study was to investigate the expression of c-kit and the clinical significance of s-kit in patients with GCTs. METHODS: The authors first conducted an immunohistochemical investigation of the expression of the c-kit protein in 27 surgical specimens. In all 18 specimens that contained germinomas, c-kit was diffusely expressed on the cell surface of the germinoma cells, but was not found on lymphocytes or interstitial cells. In seven of eight immature teratomas, only some mature components, such as cartilage and glands, were immunoreactive for c-kit. Syncytiotrophoblastic giant cells (STGCs) demonstrated negative findings as well, suggesting that primarily germinoma cells express c-kit. Next, 47 cerebrospinal fluid (CSF) samples collected from 32 patients with GCTs (15 samples from patients with pure germinomas, 16 from patients with STGC germinomas, 14 from patients with teratomas, and two from a patient with a choriocarcinoma) were analyzed using a sandwich enzyme-linked immunosorbent assay. The level of s-kit was significantly higher in CSF collected from patients with germinomas and STGC germinomas than in CSF collected from patients with teratomas or non-germ cell brain tumors, or in CSF collected from controls. The concentration of s-kit in CSF was correlated with the patient's clinical course: it was significantly higher in pretreatment samples obtained before and in samples obtained at the time of tumor recurrence than in samples collected from patients in whom the tumor was in remission. The level of s-kit was remarkably high in CSF collected from patients with subarachnoid tumor dissemination. CONCLUSIONS: These results indicate that the concentration of s-kit in CSF may be a useful clinical marker for germinomas, especially for detecting recurrence or subarachnoid dissemination of these lesions. PMID- 12134910 TI - Complete asymptomatic thrombosis and resorption of a congenital giant intracranial aneurysm. AB - Intracranial aneurysms in infants are rare, but are associated with a high risk of rupture and subarachnoid hemorrhage. The authors report a case of an incidentally diagnosed, probably congenital, asymptomatic giant aneurysm of the posterior communicating artery in a 9-month-old girl, which completely thrombosed following a diagnostic superselective angiography without any neuropathological incident. Follow-up magnetic resonance imaging revealed that the aneurysm decreased further in size and was largely resorbed within 3 years after the initial finding. In single cases the natural history of congenital giant aneurysms may be better than previously assumed. PMID- 12134911 TI - Focal motor seizures with secondary generalization arising in the cerebellum. Case report and review of the literature. AB - The issue of whether seizures can arise in the cerebellum remains controversial. The authors present the first known case of focal subcortical epilepsy with secondary generalization thought to arise from a dysplastic lesion within the cerebellum. A newborn infant presented with daily episodes of left eye blinking, stereotyped extremity movements, postural arching, and intermittent altered consciousness lasting less than 1 minute. These episodes began on his 1st day of life and progressively increased in frequency to more than 100 events per day. Antiepileptic medications had no effect, and interictal and ictal scalp electroencephalography (EEG) recordings demonstrated bilateral electrical abnormalities. Magnetic resonance imaging revealed a mass in the left cerebellar hemisphere, and ictal and interictal single-photon emission computerized tomography revealed a focal perfusion abnormality in the region of the cerebellar mass. The patient subsequently underwent intraoperative EEG monitoring with cortical scalp electrodes and cerebellar depth electrodes. Intraoperative EEG recordings revealed focal seizure discharges that arose in the region of the cerebellar mass and influenced electrographic activity in both cerebral hemispheres. Resection of this mass and the left cerebellar hemisphere led to complete resolution of the patient's seizures and normalization of the scalp EEG readings. Neuropathological findings in this mass were consistent with ganglioglioma. A review of the literature on the cerebellar origins of epilepsy is included. PMID- 12134912 TI - Production of human chorionic gonadotropin-beta subunit associated with an osteolytic meningioma. Case report. AB - An osteolytic meningioma in a 36-year-old woman was accompanied by elevated serum levels of human chorionic gonadotropin-beta subunit (beta-HCG), which returned to normal after removal of the tumor. Light microscopy examination demonstrated a transitional meningioma. Immunohistochemical analysis revealed that the tumor cells had a positive reaction for beta-HCG. This case illustrates the possibility that meningioma may be associated with clinically detectable secretion of beta HCG. To the authors' knowledge, this is the first case in which meningioma has been shown to secrete beta-HCG. The authors believe that meningioma should be considered in the differential diagnosis of choriocarcinoma, embryonal cell tumor, germinoma, and metastatic ovarian tumor associated with elevated levels of beta-HCG. PMID- 12134913 TI - Fusiform gyrus epilepsy: the use of ictal magnetoencephalography. Case report. AB - The authors report successful presurgical identification of an epileptic focus in the fusiform gyrus by using ictal magnetoencephalography (MEG), which was performed with the aid of an advanced whole-brain neuromagnetometer. A 22-year old man had suffered from medically refractory complex partial seizures since he was 10 years of age. Seizure symptoms, magnetic resonance imaging, and ictal single-photon emission computerized tomography examinations indicated right temporal lobe epilepsy; however, ictal electroencephalography, including sphenoidal recordings, failed even to lateralize the seizure focus. The MEG studies revealed that equivalent current dipoles of interictal activities were scattered bilaterally around the medial temporal structures, but those of ictal onset and postictal activities formed a cluster in the left fusiform gyrus. After confirmation of each ictal and interictal MEG finding by using long-term electrocorticography recordings, focal cortical resection of the left inferior temporal and fusiform gyri was performed. The histopathological diagnosis was cortical dysplasia, and the patient has achieved a good seizure outcome, now 15 months after the operation. Ictal and also postictal MEG may be more specific than interictal MEG for identifying the ictal onset zone. PMID- 12134914 TI - Hemilingual spasm: a new neurosurgical entity? Case report. AB - Hemilingual spasm is a little-known movement disorder, presenting as intermittent paroxysmal involuntary contractions of half of the tongue muscles. The authors report a case of hemilingual spasm caused by an arachnoid cyst. After marsupialization of the cyst, the patient's symptoms immediately resolved. There has been no recurrence of hemilingual spasm during the follow-up period of more than 40 months. PMID- 12134915 TI - Intraventricular neurocysticercosis mimicking colloid cyst. Case report. AB - The authors recently encountered a unique case of anterior third ventricular neurocysticercosis in which the cyst exhibited an unusually high signal on T1 weighted magnetic resonance imaging. The lesion's signal intensity and location made differentiation from colloid cyst difficult. Intraventricular neurocysticercosis should be included in the differential diagnosis of a colloid cyst. PMID- 12134916 TI - Utility of preoperative functional magnetic resonance imaging for identifying language cortices in patients with vascular malformations. AB - OBJECT: The goal of this study was to evaluate the utility of preoperative functional magnetic resonance (fMR) imaging in the prediction of whether a given cortical area would be deemed essential for language processing by electrocortical stimulation mapping (ESM). METHODS: The authors studied patients with vascular malformations, specifically arteriovenous malformations (AVMs) and cavernous angiomas, in whom blood-flow patterns are not normal and in whom a perfusion-dependent mapping signal may be questionable. Ten patients were studied (seven harboring AVMs and three with cavernous angiomas). The authors used a battery of linguistic tasks, including visual object naming, word generation, auditory responsive naming, visual responsive naming, and sentence comprehension, to identify brain regions that were consistently activated across expression and comprehension linguistic tasks. In a comparison of ESM and fMR imaging activations, the authors varied the matching criteria (overlapping activations, adjacent activations, and deep activations) and the radii of influence of ESM (2.5, 5, and 10 mm) to determine the effects of these factors on the sensitivity and specificity of fMR imaging. The sensitivity and specificity of fMR imaging were dependent on the task, lobe, and matching criterion. For the population studied, the sensitivity and specificity of fMR imaging activations during expressive linguistic tasks were found to be up to 100 and 66.7%, respectively, in the frontal lobe, and during comprehension linguistic tasks up to 96.2 and 69.8%, respectively, in the temporal and parietal lobes. The sensitivity and specificity of each disease population (patients with AVMs and those with cavernous angiomas) and of individuals were consistent with those values reported for the entire population studied. CONCLUSIONS: The authors conclude that preoperative fMR imaging is a highly sensitive preoperative planning tool for the identification of which cortical areas are essential for language and that this imaging modality may play a future role in presurgical planning for patients with vascular malformations. PMID- 12134917 TI - Death after late failure of third ventriculostomy in children. Report of three cases. AB - Late failure following successful third ventriculostomy for obstructive hydrocephalus is rare, and death caused by failure of a previously successful third ventriculostomy has been reported only once. The authors present three patients who died as a result of increased intracranial pressure (ICP) after late failure of a third ventriculostomy. Through a collaborative effort, three patients were identified who had died following third ventriculostomy at one of the authors' institutions. A 13-year-old girl with neurofibromatosis Type 1 underwent third ventriculostomy for obstructive hydrocephalus caused by a tectal lesion. Three years later her condition deteriorated rapidly over the course of 6 hours and she was found dead at home. A 4-year-old boy treated with third ventriculostomy for aqueductal stenosis presented 2 years postoperatively with symptoms of increased ICP. This patient suffered a cardiorespiratory arrest while under observation and died despite external ventricular drainage. A 10-year-old boy with previous ventriculoperitoneal (VP) shunt placement underwent conversion to a third ventriculostomy and shunt removal. Eight months after the procedure his condition deteriorated. with evidence of raised ICP, and he underwent emergency insertion of another VP shunt, but remained in a vegetative state and died of complications. Neuropathological examinations in two cases demonstrated that the third ventriculostomy was not patent, and there was also evidence of increased ICP. Late failure of third ventriculostomy resulting in death is a rare complication. Delay in recognition of recurrent ICP symptoms and a false feeling of security on the part of family and caregivers because of the absence of a shunt and the belief that the hydrocephalus has been cured may contribute to fatal complications after third ventriculostomy. Patients with third ventriculostomies should be followed in a manner similar to patients with cerebrospinal fluid shunts. PMID- 12134918 TI - Eagle syndrome: entrapment of the glossopharyngeal nerve? Case report and review of the literature. AB - Eagle syndrome is characterized by unilateral pain in the oropharynx, the side of the face, and the earlobe. It is caused by an elongated styloid process; resection of the elongated process eliminates the pain. Although quite rare, this syndrome is well represented in the oral, ear, nose, and throat surgery literature. In the neurosurgical literature, on the other hand, there is little if any mention of Eagle syndrome. The author presents a case of a woman who suffered from severe pain in the throat, the side of the face, and the ear. After the diagnosis of Eagle syndrome was made based on radiographic findings and was confirmed using a local anesthetic block, resection of the elongated styloid process was performed, resulting in complete and lasting pain relief. Eagle syndrome, which is caused by compression of the glossopharyngeal nerve as it passes the elongated styloid process, may be classified as an entrapment syndrome deserving of neurosurgical attention. The goal of this report is to familiarize neurosurgeons with Eagle syndrome and its diagnostic work up and treatment. PMID- 12134919 TI - Cerebral revascularization performed using posterior inferior cerebellar artery posterior inferior cerebellar artery bypass. Report of four cases and literature review. AB - Cerebral revascularization is often required for the surgical treatment of complex intracranial aneurysms. In certain anatomical locations, vascular anatomy and redundancy make in situ bypass possible. The authors present four patients who underwent revascularization performed using the rarely reported posterior inferior cerebellar artery (PICA)-PICA in situ bypass after their aneurysms had been trapped. At Barrow Neurological Institute, between 1991 and the present, four male patients underwent PICA-PICA by-passes to treat aneurysms involving the vertebral artery, the PICA, or both. The mean age of these patients was 34 years (range 5-49 years). Follow-up studies revealed patent bypasses and no evidence of infarction. Patient outcomes were excellent or good. Multiple surgical techniques have been described for revascularization of at-risk cerebral territories. Often, the blood supply must be derived from extracranial sources through a mobilized pedicle or interposited graft. Certain anatomical locations such as the vertebrobasilar junction, the anterior circle of Willis, and the middle cerebral artery bifurcation are amenable to in situ bypass because there is vessel redundancy or proximity to the contralateral analogous vessel. The advantages of an in situ bypass include one suture line, a short bypass distance, and a close match with the caliber of the recipient graft. Although technically challenging, this technique can be successful and should be considered for appropriate candidates. PMID- 12134920 TI - Acetazolamide therapy for symptomatic plateau waves in patients with brain tumors. Report of three cases. AB - In this report, the authors describe three patients with malignant gliomas who experienced paroxysmal neurological symptoms triggered by standing. The symptoms were attributed to acute elevations of intracranial pressure, an uncommon phenomenon called "plateau waves." In each instance, the attacks occurred despite the fact that the patient was receiving dexamethasone therapy and stopped promptly with the addition of acetazolamide. Acetazolamide, an orally administered carbonic anhydrase inhibitor, appears to be a specific and effective therapy for this uncommon neurological disorder. PMID- 12134921 TI - Dural arteriovenous fistula after ventriculostomy. Case illustration. PMID- 12134922 TI - Primary intracerebral myxoid chondrosarcoma. Case illustration. PMID- 12134923 TI - Use of gadodiamide-enhanced angiography during endovascular occlusion of a ruptured cerebral aneurysm. Case illustration. PMID- 12134924 TI - Reversible ischemia detected by diffusion-weighted magnetic resonance imaging. Case illustration. PMID- 12134925 TI - Cushing's case XLV: Minnie G. AB - A 23-year-old patient who was examined in 1910 by Harvey Cushing triggered his lifelong interest in the syndrome that bears his name. "Minnie G.," as she became historically known, presented with a "...syndrome of painful obesity, hypertrichosis, and amenorrhea with overdevelopment of secondary sexual characteristics accompanying a low grade of hydrocephalus and increased cerebral tension." This case stimulated Harvey Cushing's inquisitive mind and sparked an interest that 20 years later culminated in his seminal report, "The basophil adenomas of the pituitary gland and their clinical manifestations (pituitary basophilism)." In this classic work, Cushing reported in detail the cases of two patients encountered from his own practice and 10 similar cases collected from the literature. Minnie G. was the first case that Cushing reported. The clinical course of that case is briefly reviewed in this article. PMID- 12134926 TI - Peroneal nerve tumor. PMID- 12134927 TI - Are ruptured and unruptured aneurysms different? PMID- 12134928 TI - Hypotensive anesthesia. PMID- 12134929 TI - Craniopharyngioma in adults and children: a study of 122 surgical cases. AB - OBJECT: This work is devoted to a 25-year retrospective study of 122 cases of craniopharyngiomas in adults and children treated and followed by the same neurosurgeon (R.V.E.). In this homogeneous series, the aim was total microsurgical removal of the tumor, without postoperative radiotherapy. METHODS: The operation was performed via a frontopterional approach in 112 cases and a transsphenoidal approach in 10 cases. The tumor removal was considered total in 59%, subtotal in 29%, and partial in 12%. The surgical mortality rate was 2.5%. Even when tumor removal was not complete, radiotherapy was not systematically administered; it was reserved for cases of recurrence. The authors have studied clinical signs, operative characteristics, and ophthalmological, endocrinological, and functional outcomes, as well as recurrence risk and long term patient survival. The mean follow-up period was 7 years. The functional results in these patients were excellent in 85%, good in 9%, fair in 5% (usually because of ophthalmological sequelae), and poor in 1%. Tumors recurred in 29 patients, but the salvage treatment, by operation or radiotherapy, was successful in 83%. The actuarial patient survival rate was 92% after 5 years and 85% after 10 years. CONCLUSIONS: These results compared favorably with the data reported in the literature, suggesting that radical surgery of craniopharyngiomas allows good outcome in terms of survival, full recovery, and quality of life for both adults and children. PMID- 12134930 TI - Cortical stimulation mapping of language cortex by using a verb generation task: effects of learning and comparison to mapping based on object naming. AB - OBJECT: Cortical stimulation mapping has traditionally relied on disruption of object naming to define essential language areas. In this study, the authors reviewed the use of a different language task, verb generation, in mapping language. This task has greater use in brain imaging studies and may be used to test aspects of language different from those of object naming. METHODS: In 14 patients, cortical stimulation mapping performed using a verb generation task provided a map of language areas in the frontal and temporoparietal cortices. These verb generation maps often overlapped object naming ones and, in many patients, different areas of cortex were found to be involved in the two functions. In three patients, stimulation mapping was performed during the initial performance of the verb generation task and also during learned performance of the task. Parallel to findings of published neuroimaging studies, a larger area of stimulated cortex led to disruption of verb generation in response to stimulation during novel task performance than during learned performance. CONCLUSIONS: Results of cortical stimulation mapping closely resemble those of functional neuroimaging when both implement the verb generation task. The precise map of the temporoparietal language cortex depends on the task used for mapping. PMID- 12134931 TI - Multiple subpial transections: outcome and complications in 20 patients who did not undergo resection. AB - OBJECT: The authors describe patient characteristics, surgical methods, complications, and outcome over time in a cohort of patients who underwent multiple subpial transection (MST) without concomitant cortical resection. METHODS: Twenty consecutive patients in whom drug-resistant epilepsy had been diagnosed a mean of 16 +/- 9 years earlier (mean +/- standard deviation [SD]) were treated with MST without cortical resection. The mean follow-up period was 49.3 +/- 18.3 months (mean +/- SD, median 58 months). At 12 months of follow up, two of the 20 patients were Engel Class I, one was Class II, six were Class III, and 11 were Class IV. At latest follow up, one patient was Engel Class I, one was Class II, seven were Class III, and 11 were Class IV. According to an alternative five-tiered classification system, two outcomes were excellent, seven were good, one was fair, nine were poor, and one was worse. Outcome was found to be better in patients with no lesions observed on magnetic resonance (MR) imaging, and worse in those with large MST areas. Outcome had a tendency to change (this occurred in 13 of 20 cases). Five patients (25%) improved and seven (35%) deteriorated in Engel outcome class, and in one (5%) both developments occurred over time. Most outcome class changes occurred before the end of the 2nd year (nine), and four were observed in the 5th year. There where seven transient neurological deficits and four surgical complications. There was no permanent significant morbidity, and there were no deaths. CONCLUSIONS: Forty-five percent of patients achieved a worthwhile improvement after pure MST, if Engel outcome Class III is deemed a worthwhile improvement. The alternative five-tiered classification resulted in 50% with worthwhile improvement (excellent, good, or fair outcome), 45% with poor, and 5% with worse outcome. Lesions that are detectable on MR imaging, and large MST areas are predictive of worse results. Significant intraoperative problems may arise, but this happens infrequently. There is a notable rate of transient morbidity but the rate of permanent morbidity is not significant. PMID- 12134932 TI - Stereotactic radiosurgery for pediatric intracranial arteriovenous malformations: the University of California at San Francisco experience. AB - OBJECT: Stereotactic radiosurgery for arteriovenous malformations (AVMs) is an accepted treatment option, but few reports have been published on the results of this treatment in children. In this study the authors describe a series of pediatric patients with a minimum follow-up duration of 36 months. METHODS: From 1991 to 1997, 40 children (26 boys and 14 girls) with AVMs were treated with radiosurgery at the University of California at San Francisco (UCSF). Follow-up information was available for 31 children (20 boys and 11 girls) in whom the median age at initial treatment was 11.2 years (range 3.4-17.5 years). The median follow-up duration was 60 months (range 6-99 months). Sixteen percent of the AVMs were Spetzler-Martin Grade II; 68%, Grade III; 10%, Grade IV; and 6%, Grade V. The mean volume of the AVMs was 5.37 cm3 and the median volume was 1.6 cm3. The mean marginal dose of radiation was 16.7 Gy and the median dose was 18 Gy (range 12-19 Gy). Angiography performed in 26 children confirmed obliteration of the AVM nidus in nine patients (35%), partial response in 16 patients (62%), and no response in one patient (4%). In five patients who refused angiography, magnetic resonance (MR) imaging revealed obliteration in two patients and partial response in three patients, bringing the overall obliteration rate associated with initial radiosurgery to 35%. Logistic regression analysis confirmed a significant correlation between marginal dose prescription and response (p = 0.025); in AVMs that received at least 18 Gy there was a 10-fold increase in the obliteration rate (63%) over AVMs that received a lower dose. Lesions smaller than 3 cm3 were associated with a six-fold increased obliteration rate (53%) over lesions larger than 3 cm3 (8%), but AVM volume was not a statistically significant predictor of response (p = 0.09). Twelve patients have since undergone repeated radiosurgery and are currently being followed up with serial MR imaging studies (in five cases, the AVM is now obliterated). During the follow-up period (1918 patient months) there were eight hemorrhages in five patients, with a cumulative posttreatment hemorrhage rate of 3.2%/patient/year in the 1st year and a rate of 4.3%/patient/year over the first 3 years. There were two permanent neurological complications (6%) and no deaths in this study. CONCLUSIONS: The lower overall obliteration rate reported in this series is most likely due to the larger mean AVM volumes treated at UCSF as well as conservative dose-volume prescriptions delivered to children. Significantly higher obliteration rates were observed when a marginal radiation dose of at least 18 Gy was delivered. The permanent complication rate is low and should encourage those treating children to use doses similar to those used in adults. PMID- 12134933 TI - Stereotactic radiosurgery for pilocytic astrocytomas when multimodal therapy is necessary. AB - OBJECT: The goal of this study was to examine the role of stereotactic radiosurgery in the treatment of patients with recurrent or unresectable pilocytic astrocytomas. METHODS: During a 13-year interval, 37 patients (median age 14 years) required multimodal treatment of recurrent or unresectable pilocytic astrocytomas. Tumors involved the brainstem in 18 patients, cerebellum in three, thalamus in five, temporal lobe in four, and parietal lobe in two, as well as the hypothalamus, optic tract, corpus callosum, insular cortex, and third ventricle in one patient each. Diagnosis was confirmed with the aid of stereotactic biopsy in 12 patients, open biopsy in five, partial resection in eight, and near-total resection in 12. Multimodal treatment included fractionated radiation therapy in 10 patients, stereotactic intracavitary irradiation of tumor in four, chemotherapy in two, cyst drainage in six, ventriculoperitoneal shunt placement in three, and additional cytoreductive surgery in four. Tumor volumes varied from 0.42 to 25 cm3. The median radiosurgical dose to the tumor margin was 15 Gy (range 9.6-22.5 Gy). After radiosurgery, serial imaging demonstrated complete tumor resolution in 10 patients, reduced tumor volume in eight, stable tumor volume in seven, and delayed tumor progression in 12. No procedure-related death was encountered. Thirty-three (89%) of 37 patients are alive at a median follow-up period of 28 months after radiosurgery and 59 months after diagnosis. Eight patients participated in follow-up review for more than 60 months. Three patients died of local tumor progression. CONCLUSIONS: Stereotactic radiosurgery is a valuable adjunctive strategy in the management of recurrent or unresectable pilocytic astrocytomas. Despite the favorable histological characteristics and prognosis usually associated with this neoplasm, an adverse location, recurrence, or progression of this disease requires alternative therapeutic approaches such as radiosurgery. PMID- 12134934 TI - Stereotactic radiosurgery providing long-term tumor control of cavernous sinus meningiomas. AB - OBJECT: To evaluate long-term outcomes of patients who have undergone stereotactic radiosurgery for cavernous sinus meningiomas, the authors retrospectively reviewed their 14-year experience with these cases. METHODS: One hundred seventy-six patients harbored meningiomas centered within the cavernous sinus. Seventeen patients were lost to follow-up review, leaving 159 analyzable patients, in whom 164 procedures were performed. Seventy-six patients (48%) underwent adjuvant radiosurgery after one or more attempts at surgical resection. Eighty-three patients (52%) underwent primary radiosurgery. Two patients (1%) had previously received fractionated external-beam radiation therapy. Four patients (2%) harbored histologically verified atypical or malignant meningiomas. Conformal multiple isocenter gamma knife surgery was performed. The median dose applied to the tumor margin was 13 Gy. Neurological status improved in 46 patients (29%), remained stable in 99 (62%), and eventually worsened in 14 (9%). Adverse effects of radiation occurred after 11 procedures (6.7%). Tumor volumes decreased in 54 patients (34%), remained stable in 96 (60%), and increased in nine (6%). The actuarial tumor control rate for patients with typical meningiomas was 93.1 +/- 3.3% at both 5 and 10 years. For the 83 patients who underwent radiosurgery as their sole treatment, the actuarial tumor control rate at 5 years was 96.9 +/- 3%. CONCLUSIONS: Stereotactic radiosurgery provided safe and effective management of cavernous sinus meningiomas. We believe it is the preferred management strategy for tumors of suitable volume (average tumor diameter < or = 3 cm or volume < or = 15 cm3). PMID- 12134935 TI - Volumetric measurements in the detection of reduced ventricular volume in patients with normal-pressure hydrocephalus whose clinical condition improved after ventriculoperitoneal shunt placement. AB - OBJECT: The syndrome of normal-pressure hydrocephalus (NPH) refers to the clinical triad of gait disturbance, dementia, and urinary incontinence in association with idiopathic ventriculomegaly and normal intracranial pressure. Ventriculoperitoneal (VP) shunt placement often yields significant clinical improvements, sometimes without apparent reduction of ventricular size. The authors hypothesized that careful volumetric measurements would show a decrease in ventricular volume in these patients. METHODS: Twenty consecutive patients with NPH underwent placement of VP shunts equipped with programmable valves. In 11 patients pre- and postoperative neuroimaging was performed, which allowed volumetric analysis. Volumetric measurements of the lateral ventricles were calculated in triplicate by National Institutes of Health image-processing software to assess standard computerized tomography (CT) scans (eight patients) or magnetic resonance (MR) images (three patients) obtained before and after shunt placement. Ventricular volumes were also assessed by an independent neuroradiologist. Postoperative studies were performed at a time of clinical improvement, between 1 and 9 months postsurgery (mean 5 months). Preoperative and postoperative Unified Parkinson's Disease Rating Scale evaluations were performed in four patients. Significant clinical improvement occurred in all patients after shunt placement (mean follow-up period 17.5 months). Although 10 (91%) of 11 patients demonstrated a calculable decrease in volume in the lateral ventricles (mean decrease 39%), formal interpretation of neuroimages indicated a definite decrease in lateral ventricular volume in only three (27%) of 11 patients. CONCLUSIONS: Volumetric measurements obtained to compare preoperative and postoperative CT or MR studies obtained in patients with NPH in whom clinical improvement was seen after shunt placement surgery show a demonstrable decrease in ventricular size. Volumetric measurements may be helpful in clinical assessment postoperatively and in guiding programmable valve pressure settings. PMID- 12134936 TI - Intraoperative lidocaine injection into the carotid sinus during endarterectomy. AB - OBJECT: Many surgeons inject a local anesthetic agent into the carotid sinus before carotid endarterectomy in an attempt to ameliorate perioperative hemodynamic instability. The purpose of this study is to analyze the effect of carotid sinus injection with lidocaine on perioperative hemodynamics and complications. METHODS: The authors prospectively studied 92 patients in whom 100 consecutive carotid endarterectomies were performed by a single surgeon (eight procedures were bilateral). Patients were randomly assigned to one of two groups, in which either 0.5 ml of 1% lidocaine was injected into the carotid sinus nerve or no injection of lidocaine was administered before the arteriotomy. All patients were treated postoperatively according to a standard endarterectomy protocol. There were no significant differences between the two groups in the incidence of hypertension, hypotension, or the use of vasoactive medications in the operating room following restoration of carotid artery (CA) blood flow, in the recovery room, or in the intensive care unit. CONCLUSIONS: Injection of lidocaine into the carotid sinus at the time of endarterectomy is not associated with a significant improvement in any hemodynamic factor, from the time of restoration of CA blood flow to postoperative Day 1. PMID- 12134937 TI - Early and persistent impaired percent alpha variability on continuous electroencephalography monitoring as predictive of poor outcome after traumatic brain injury. AB - OBJECT: Early prediction of outcomes in patients after they suffer traumatic brain injury (TBI) is often nonspecific and based on initial imaging and clinical findings alone, without direct physiological testing. Improved outcome prediction is desirable for ethical, social, and financial reasons. The goal of this study was to determine the usefulness of continuous electroencephalography (EEG) monitoring in determining prognosis early after TBI, while the patient is in the intensive care unit. METHODS: The authors hypothesized that the reduced percentage of alpha variability (PAV) in continuous EEG tracings indicates a poor prognosis. Prospective continuous EEG monitoring was performed in 89 consecutive patients with moderate to severe TBI (Glasgow Coma Scale [GCS] Scores 3-12) from 0 to 10 days after injury. The PAV was calculated daily, and the time course and trends of the PAV were analyzed in comparison with the patient's Glasgow Outcome Scale (GOS) score at the time of discharge. In patients with GCS scores of 8 or lower, a PAV value of 0.1 or lower is highly predictive of a poor outcome or death (positive predictive value 86%). The determinant PAV value was obtained by Day 3 after injury. Persistent PAV values of 0.1 or lower over several days or worsening of the PAV to a value of 0.1 or lower indicated a high likelihood of poor outcome (GOS Scores 1 and 2). In comparison with the combination of traditional initial clinical indicators of outcome (GCS score, pupillary response to light, patient age, results of computerized tomography scanning, and early hypotension or hypoxemia), the early PAV value during the initial 3 days after injury independently improved prognostic ability (p < 0.01). CONCLUSIONS: Continuous EEG monitoring performed with particular attention paid to the PAV is a sensitive and specific method of prognosis that can indicate outcomes in patients with moderate to severe TBI within 3 days postinjury. PMID- 12134938 TI - Delayed facial palsy after resection of vestibular schwannoma. AB - OBJECT: In this study the authors investigate delayed facial palsy (DFP), which is an underreported phenomenon after surgery for vestibular schwannoma (VS). The authors identified 15 (4.8%) patients from a consecutive series of 314 who underwent surgery for VS between 1988 and 2000, and in whom DFP developed. Delayed facial palsy was defined as a deterioration of facial nerve function from House-Brackmann Grades 1 or 2 more than 3 days postoperatively. METHODS: All patients underwent intraoperative neurophysiological monitoring of facial nerve function. The average latency of DFP was 10.9 days (range 4-30 days). In six patients (40%) minor deterioration (< or = two House-Brackmann grades) had occurred at a mean of 10.2 days postsurgery, whereas in nine patients (60%) moderate deterioration (> or = three House-Brackmann grades) had occurred at a mean of 11.8 days postoperatively. Five (33%) of 15 patients recovered to Grade 1 of 2 function within 6 weeks of DFP onset. Of the 15 patients with DFP, 14 had completed 1 year of follow up at the time of this study. Twelve (80%) of these 15 patients recovered to Grade 1 or 2 function within 3 months, and 13 (93%) of 14 patients recovered within 1 year. In all cases, stimulation of the seventh cranial nerve on completion of tumor resection revealed the nerve to be intact, both anatomically and functionally, to proximal and distal stimulation at 0.1 mA. A smaller tumor diameter correlated with greater recovery of facial nerve function. There was no correlation between the latency or severity of or recovery from DFP, and the patient's age or sex, the surgical approach, frequency of neurotonic seventh nerve discharges, anatomical relationship of the facial nerve to the tumor, patient's history of tobacco use, or cardiovascular disease. CONCLUSIONS: It appears that DFP is an uncommon consequence of surgery for VS. Although excellent recovery of facial nerve function to its original postoperative status nearly always occurs after DFP, the magnitude and time course of the disorder were not predictors for subsequent recovery of facial nerve function. PMID- 12134939 TI - Cation dysfunction associated with cerebral ischemia followed by reperfusion: a comparison of microdialysis and ion-selective electrode methods. AB - OBJECT: Disruption of ionic homeostasis during ischemia is a well-characterized event and is identified by a rise in the concentration of extracellular potassium [K+]e, with a concomitant reduction in the concentration of extracellular sodium [Na+]e. Results of clinical studies in which microdialysis has been used, however, have shown only modest changes in the levels of extracellular ions. The object of this study was to measure [K+]e and [Na+]e by using ion-selective electrodes (ISEs) and to compare these measurements with those obtained using the well-established method of microdialysis. METHODS: Fifteen Sprague-Dawley rats were separated into three groups. Five animals were subjected to a 15-minute period of ischemia, and another five animals to a 60-minute period of ischemia; animals in both of these groups received K+-free microdialysis perfusate. The third group of five rats underwent a 60-minute period of ischemia and received a reduced-Na+ microdialysis perfusate. Transient forebrain ischemia was produced by bilateral carotid artery occlusion combined with hypotension. A custom-fabricated glass Na+ electrode and a flexible plastic K+ and reference electrodes were used to monitor extracellular ion transients. Microdialysis samples were obtained with the aid of a 2-mm microdialysis probe that was perfused with K+-free mock cerebrospinal fluid at a rate of 2 microl/minute. Baseline measurements of [K+]e and [Na+]e, obtained using ISEs, were 3.41 +/- 0.09 mM and 145 +/- 7.75 mM. respectively. Ischemia resulted in a rapid accumulation of [K+]e (in animals subjected to 15 minutes of ischemia, the concentration was 41.9 +/- 13.7 mM; and in animals subjected to 60 minutes of ischemia, the concentration was 66.9 +/- 11.5 mM), with a concomitant decrease in [Na+]e (in animals subjected to 15 minutes of ischemia, the concentration was 71.7 +/- 2.9 mM; and in animals subjected to 60 minutes of ischemia, the concentration was 74.7 +/- 1.9 mM). A comparison of microdialysis and ISE methods revealed that microdialysis underestimated the [K+]e changes and was insensitive to concomitant [Na+]e alterations that occur during ischemia. CONCLUSIONS: Our results indicate that the flexible ISE is a reliable and accurate tool for monitoring ionic dysfunction that accompanies brain injury. PMID- 12134941 TI - Molecular factors influencing retention on immobilized artifical membranes (IAM) compared to partitioning in liposomes and n-octanol. AB - PURPOSE: To assess the effect of molecular factors influencing retention on immobilized artificial membrane (IAM) high-performance liquid chromatography columns compared to liposomal partitioning and traditional n-octanol/water partition coefficients. METHODS: IAM capacity factors were measured at pH 7.0 on an IAM.PC.DD2 stationary phase. Liposomal partitioning at pH 7.0 and n octanol/water partition coefficients were measured using the pH metric method. Partitioning in egg-phosphatidylcholine (PhC) liposomes was also measured by equilibrium dialysis for a series of beta-blockers. RESULTS: For the ionized beta blockers, potentiometry and equilibrium dialysis yielded consistent partitioning data. For relatively large bases. IAM retention correlated well with PhC liposome partitioning, hydrophobic forces being mainly involved. For more hydrophilic compounds and for heterogeneous solutes, in contrast, the balance between electrostatic and hydrophobic interactions was not the same in the two systems. Hydrogen bonding, an important factor in liposomes partitioning, played only a minor role in IAM retention. CONCLUSIONS: Partitioning in immobilized artificial membranes depends on size, hydrophobicity, and charge. When hydrophobic interactions dominate retention, IAM capacity factors are well correlated with liposomal partitioning. On the contary, for hydrophilic solutes, the two systems do not yield the same information and are not interchangeable. PMID- 12134942 TI - Organic anion transporter oatp2-mediated interaction between digoxin and amiodarone in the rat liver. AB - PURPOSE: The interaction between amiodarone and digoxin has been known to increase serum concentrations of digoxin in humans and rats. In this study, we assessed the molecular mechanism(s) of that drug interaction, focusing on digoxin transport mediated by P-glycoprotein (Pgp) and by rat liver organic anion transporter (oatp2). METHODS: Digoxin transport by Pgp and oatp2 was assessed using Pgp-overexpressing transfectant LLC-GA5-COL150 monolayers and oatp2 expressing Xenopus oocytes, respectively. The digoxin uptake into the isolated rat hepatocytes was also examined. RESULTS: Amiodarone (10 microM) inhibited slightly the transcellular transport of digoxin in LLC-GA5-COL150 monolayers, whereas itraconazole (10 microM), a potent Pgp inhibitor, markedly blocked the transport. The digoxin uptake by the isolated rat hepatocytes and by the oatp2 expressing Xenopus oocytes was decreased markedly in the presence of amiodarone but not in the presence of itraconazole. In addition, amiodarone inhibited the oatp2-mediated digoxin uptake in a competitive manner with an apparent inhibition constant value of 1.8 microM. CONCLUSION: These findings suggest that rat oatp2 rather than Pgp may be one of the interaction sites for digoxin and amiodarone in the liver. PMID- 12134940 TI - Recent advances in vaccine adjuvants. AB - New generation vaccines, particularly those based on recombinant proteins and DNA, are likely to be less reactogenic than traditional vaccines but are also less immunogenic. Therefore, there is an urgent need for the development of new and improved vaccine adjuvants. Adjuvants can be broadly separated into two classes based on their principal mechanisms of action: vaccine delivery systems and immunostimulatory adjuvants. Vaccine-delivery systems generally are particulate (e.g., emulsions, microparticles, iscoms, and liposomes) and function mainly to target associated antigens into antigen-resenting cells. In contrast, immunostimulatory adjuvants are derived predominantly from pathogens and often represent pathogen-ssociated molecular patterns (e.g., lipopolysaccaride, monophosphoryl lipid A, CpG DNA). which activate cells of the innate immune system. Recent progress in innate immunity is beginning to yield insight into the initiation of immune responses and the ways in which immunostimulatory adjuvants may enhance this process. The discovery of more potent adjuvants may allow the development of prophylactic and therapeutic vaccines against cancers and chronic infectious diseases. In addition, new adjuvants may also allow vaccines to be delivered mucosally. PMID- 12134943 TI - Conjugates of antisense oligonucleotides with the Tat and antennapedia cell penetrating peptides: effects on cellular uptake, binding to target sequences, and biologic actions. AB - PURPOSE: The attainment of effective intracellular delivery remains an important issue for pharmacologic applications of antisense oligonucleotides. Here, we describe the synthesis, binding properties, and biologic properties of peptide oligonucleotide conjugates comprised of the Tat and Ant cell-penetrating peptides with 2'-O-methyl phosphorothioate oligonucleotides. METHODS: The biologic assay used in this study measures the ability of the antisense molecule to correct splicing of an aberrant intron inserted into the Luciferase gene; thus, this assay clearly demonstrates the delivery of functional antisense molecules to the splicing machinery within the nucleus. The binding affinities of the conjugates to their target sequences were measured by surface plasmon resonance (BIAcor) techniques. RESULTS: The peptide-oligonucleotide conjugates progressively entered cells over a period of hours and were detected in cytoplasmic vesicles and in the nucleus. Peptide-oligonucleotide conjugates targeted to the aberrant splice site, but not mismatched controls, caused an increase in Luciferase activity in a dose responsive manner. The kinetics of Luciferase appearance were consistent with the course of the uptake process for the conjugates. The effects of peptide conjugation on the hybridization characteristics of the oligonucleotides were also examined using surface plasmon resonance. The peptide-oligonucleotide conjugates displayed binding affinities and selectivities similar to those of unconjugated oligonucleotides. CONCLUSIONS: Conjugation with cell-penetrating peptides enhances oligonucleotide delivery to the nucleus without interfering with the base-pairing function of antisense oligonucleotides. PMID- 12134944 TI - Alginate/poly-L-lysine microparticles for the intestinal delivery of antisense oligonucleotides. AB - PURPOSE: A microparticle carrier based on alginate and poly-L-lysine was developed and evaluated for the delivery of antisense oligonucleotides at the intestinal site. Formulations of oligonucleotide-loaded microparticles having differences in the carrier molecular weight and composition were characterized in vitro and in vivo. METHODS: Polymeric microparticles were prepared by ionotropic gelation and crosslinking of alginate with calcium ions and poly-L-lysine. The loading of the antisense oligonucleotide into the microparticles was achieved by absorption in aqueous medium. The association capacity, loading and particle size of the microparticles were characterized. The in vivo performances of various formulations after intrajejunal administration were studied in rat and in dog models. RESULTS: Microparticles had a sponge-like structure and an oligonucleotide loading of 27-35%. The composition of the medium affected the particle size and the in vitro release profiles. The oligonucleotide bioavailability after intrajejunal administration to rats in the presence of permeation enhancers was good for most of the tested systems. The application of microparticles in powder form compared to an equivalent suspension improved the intrajejunal bioavailability of the oligonucleotide (25% and 10% respectively) in rats. On the contrary, the intrajejunal administration to dogs resulted in poor oligonucleotide bioavailability (0.42%). CONCLUSIONS: The formulation of antisense oligonucleotides within alginate and poly-L-lysine microparticles is a promising strategy for the oral application. PMID- 12134945 TI - Are MDCK cells transfected with the human MDR1 gene a good model of the human intestinal mucosa? AB - PURPOSE: To investigate whether Madin-Darby canine kidney cells transfected with the human MDR1 gene (MDCK-MDR1) are a good model of the human intestinal mucosa. METHODS: P-glycoprotein (P-gp) expression in Caco-2 cells was compared with P-gp expression in MDCK wild- type (MDCK-WT) and MDCK-MDR1 cells using Western blotting methods. The polarized efflux activities of P-gp(s) in MDCK-MDRI cells, MDCK-WT cells, and Caco-2 cells were compared using digoxin as a substrate. Apparent Michaelis-Menten constants (K(M),Vmax) for the efflux of vinblastine in these three cell lines were determined. Apparent inhibition constants (K(I)) of known substrates/inhibitors of P-gp were determined by measuring their effects on the efflux of digoxin in Caco-2 or MDCK-MDR1 cell monolayers. RESULTS: MDCK-MDR1 cells expressed higher levels of P-gp compared to Caco-2 and MDCK-WT cells, as estimated by Western blots. Two isoforms of P-gp were expressed in Caco-2 and MDCK cells migrating with molecular weights of 150 kDa and 170 kDa. In MDCK-MDR1 cells, the 150 kDa isoforms appeared to be overexpressed. The MDCK-MDR1 cells exhibited higher polarized efflux of [3H]-digoxin than did Caco-2 and MDCK-WT cells. K(M) values of vinblastine in Caco-2. MDCK-WT, and MDCK-MDR1 cells were 89.2+/-26.1, 24.5+/-1.1, and 252.8+/-134.7 microM, respectively, whereas Vmax values were 1.77+/-0.22, 0.42+/-0.01, and 2.43+/-0.86 pmolcm(-2)s(-1), respectively. Known P-gp substrates/inhibitors showed, in general, lower K(I) values for inhibition of digoxin efflux in Caco-2 cells than in MDCK-MDR1 cells. CONCLUSIONS: These data suggest that the MDCK-MDR1 cells overexpress the 150 kDa isoform of P-gp. MDCK-MDR1 cells are a useful model for screening the P-gp substrate activity of drugs and drug candidates. However, the apparent kinetics constants and affinities of substrates determined in the MDCK-MDR1 cell model may be different than the values obtained in Caco-2 cells. These differences in substrate activity could result from differences in the relative expression levels of total P-gp in Caco-2 and MDCK-MDR1 cells and/or differences in the partitioning of substrates into these two cell membrane bilayers. PMID- 12134946 TI - Are MDCK cells transfected with the human MRP2 gene a good model of the human intestinal mucosa? AB - PURPOSE: To investigate whether Madin-Darby canine kidney cells transfected with the human MRP2 gene (MDCK-MRP2) are a good model of the human intestinal mucosa. METHODS: MRP2 expression in Caco-2 cells was compared with the expression of this efflux transporter in MDCK-wild type (MDCK-WT) and MDCK-MRP2 cells using Western blotting methods. The polarized efflux activities of MRP2 in the MDCK-MRP2, MDCK WT. MDCK cells transfected with the human MDR1 gene (MDCK-MDR1), and Caco-2 cells were compared using vinblastine as a substrate. Apparent Michaelis-Menten constants (K(M), Vmax) for the efflux of vinblastine in Caco-2 and MDCK-MRP2 cells were determined in the presence of GF120918 (2 microM), which inhibits P glycoprotein but does not affect MRP2. Apparent inhibitory constants (K(I)) of known substrates/inhibitors of MRP2 were determined by measuring their effects on the efflux of vinblastine in these cell lines. RESULTS: MDCK-MRP2 cells expressed higher levels of MRP2 than MDCK-WT and Caco-2 cells as measured by Western blotting technique. Two isoforms of MRP2 expressed in Caco-2 and MDCK cells migrated at molecular weights of 150 kD and 190 kD. In MDCK-MRP2 cells, the 150 kD isoform appeared to be overexpressed. MDCK-MRP2 cell monolayers exhibited higher polarized efflux of vinblastine than Caco-2 and MDCK-WT cell monolayers. K(M) values for vinblastine in Caco-2 and MDCK-MRP2 cells were determined to be 71.8+/-11.6 and 137.3+/-33.6 microM. respectively, and Vmax values were determined to be (0.54+/-0.03 and 2.45+/-0.31 pmolcm(-2)s(-1), respectively. Known substrates/inhibitors of MRP2 showed differences in their ability to inhibit vinblastine efflux in Caco-2 cells as compared to MDCK-MRP2 cells CONCLUSIONS: These data suggest that MDCK-MRP2 cells overexpress only the 150 kD isoform of MRP2. The 190 kD isoform, which was also found in Caco-2 cells and MDCK-WT cells, was present in MDCK-MRP2 cells but not over expressed. The apparent kinetics constants and affinities of some MRP2 substrates were different in Caco-2 cells and MDCK-MRP2 cells. These differences in substrate activity could result from differences in the relative expression levels of the MRP2 isoforms present in Caco-2 cells and MDCK-MRP2 cells and/or differences in the partitioning of substrates in these two cell membrane bilayers. PMID- 12134947 TI - Characterization of the efflux transporter(s) responsible for restricting intestinal mucosa permeation of an acyloxyalkoxy-based cyclic prodrug of the opioid peptide DADLE. AB - PURPOSE: To elucidate the efflux transporter(s) responsible for restricting the permeation of an acyloxyalkoxy-based cyclic prodrug of the opioid peptide DADLE (AD) through Caco-2 cell monolayers. METHODS: The cellular permeation characteristics of AD were investigated using Caco-2 cells, Madin-Darby canine kidney wild-type II cells (MDCK-WT), MDCK cells transfected with the human MDR1 gene (MDCK-MDR1), and MDCK cells transfected with the human MRP2 gene (MDCK MRP2). These cells were grown as monolayers onto microporous membranes. The disappearance of AD from the donor side and its appearance on the receiver side were monitored by high-performance liquid chromatography. The substrate activity of AD for P-glycoprotein (P-gp) was determined using GF120918, a known P-gp specific inhibitor. The substrate activity of AD for MRP2 was determined by using cyclosporin A, a known MRP2 and P-gp inhibitor. RESULTS: In Caco-2 cells, the ratio of the apparent permeability coefficients (Papp) of AD flux measured in the basolateral (BL) to apical (AP) direction vs. the flux in the AP-to-BL direction (Papp BL-to-AP/ Papp AP-to-BL) was 99. In the presence of 2 microM GF120918 or 25 microM cyclosporin A. the Papp BL-to-AP/Papp AP-to-BL ratio was decreased to 11. In MDCK-WT, MDCK-MDR1, and MDCK-MRP2 cells, the Papp BL-to-AP/Papp AP-to-BL ratios of AD were 4.7, 10, and 5.8, respectively. A mixture of GF120918 (2 microM) and cyclosporin A (25 microM) decreased the Papp BL-to-AP/Papp AP-to-BL ratios of AD in MDCK-WT, MDCK-MDR1, and MDCK-MRP2 cells to 1.2,1.8, and 2.3, respectively. CONCLUSIONS: These data suggest that AD is a much better substrate for P-gp than MRP2 and that the restricted permeation of this cyclic prodrug in Caco-2 cells and in the intestinal mucosa probably is due primarily to its substrate activity for P-gp. PMID- 12134948 TI - Characterization of the efflux transporter(s) responsible for restricting intestinal mucosa permeation of the coumarinic acid-based cyclic prodrug of the opioid peptide DADLE. AB - PURPOSE: To elucidate the efflux transporter(s) responsible for restricting the permeation of a coumarinic acid-based cyclic prodrug of the opioid peptide DADLE (CD) thorough Caco-2 cell monolayers. METHODS: The cellular permeability characteristics of CD were investigated using Caco-2 cells, Madin-Darby canine kidney-wild type II cells (MDCK-WT). MDCK cells transfected with the human MDR1 gene (MDCK-MDR1), and MDCK cells transfected with human MRP2 gene (MDCK-MRP2). These cells were grown as monolayers onto microporous membranes. The disappearance from the donor side and appearance on the receiver side of CD were monitored by HPLC. The substrate activity of CD for P-gp was determined by using GF120918. a known P-gp specific inhibitor. The substrate activity of CD for MRP2 was determined by using cyclosporin A (CsA), a known MRP2 and P-gp inhibitor. RESULTS: In Caco-2 cells, the ratio of the apparent permeability coefficients (Papp) of CD flux in the basolateral (BL) to apical (AP) direction vs. the flux in the AP-to-BL direction (Papp-BL-to-AP/Papp AP-to-BL) was 71. In the presence of GF120918 (2 microM), the Papp BL-to-AP/Papp AP-to-BL ratio was decreased to 16. In the presence of CsA (25 microM), the ratio was decreased to 5.6. In MDCK WT. MDCK-MDR1, and MDCK-MRP2 cells, the Papp BL-AP/Papp AP-to-BL ratios of CD were 13, 35, and 22, respectively. CsA (25 microM) greatly decreased the Papp BL P-AP/Papp AP-to-BL ratios in MDCK-WT and MDCK-MDR1 cells to 1.5 and 3.2, respectively. However, in MDCK-MRP2 cells. CsA (25 microM) decreased the ratio only to 11. A mixture of GF120918 (2 microM) and CsA (25 microM) decreased the Papp BL-to-AP/Papp AP-to-BL ratios of CD in MDCK-WT, MDCK-MDR1, and MDCK-MRP2 cells to 1.4, 2.7, and 5.4. respectively. CONCLUSIONS: These data suggest that CD is a good substrate for both P-gp and MRP2 and that the restricted permeation of this cyclic prodrug in Caco-2 cells and in the intestinal mucosa is probably due to its substrate activities for both of these efflux transporters. PMID- 12134949 TI - A modified coumarinic acid-based cyclic prodrug of an opioid peptide: its enzymatic and chemical stability and cell permeation characteristics. AB - PURPOSE: To evaluate the chemical/enzymatic stability and the cell permeation characteristics of the modified coumarinic acid-based cyclic prodrug 2 of DADLE (H-Tyr-D-Ala-Gly-Phe-D-Leu-OH), which has an aldehyde equivalent (oxymethyl) inserted between the phenolic group of the promoiety and the carboxylic acid group of the peptide. METHODS: The rates of the chemical/enzymatic conversion of the oxymethyl-modified prodrug 2 to DADLE were measured by HPLC. The cellular permeation characteristics of DADLE and its oxymethyl-modified prodrug 2 were measured by HPLC using Caco-2 cells, wild type Madin-Darby Canine Kidney cells (MDCK-WT), MDCK cells transfected with human MDR1 gene (MDCK-MDR1), and MDCK cells transfected with human MRP2 gene (MDCK-MRP2) grown onto microporous membranes. RESULTS: The oxymethyl-modified coumarinic acid-based cyclic prodrug 2 degraded chemically to DADLE in a pH-dependent manner, i.e., rates of conversion increased with increasing pH. The prodrug 2 degraded rapidly in rat plasma (t1/2 = 39 min) and rat liver homogenate (t1/2 = 59.2 min), but much slower in Caco-2 cell homogenate (t1/2 = 678.7 min) and human plasma (t1/2 = 264.3 min). In all four cell lines used for transport studies, the flux rates of the oxymethyl prodrug 2 in the basolateral (BL)-to-apical (AP) direction (Papp BL-to-AP) were significantly greater than the flux rates in the AP-to-BL direction (Papp AP-to BL). The Papp BB-to-AP/Papp AP-to-BL ratios were >116, 35.1, 21.2, and 12.6 in Caco-2, MDCK-MDR1, MDCK-MRP2, and MDCK-WT cells, respectively. The efflux of the modified prodrug could be inhibited by GF120918 (an inhibitor for P-gp) and cyclosporin A (an inhibitor for P-gp and MRP2). CONCLUSIONS: The oxymethyl modified coumarinic acid-based cyclic prodrug 2 of DADLE could be converted to DADLE in both chemical and enzymatic media. However, the prodrug was a good substrate for both P-gp and MRP2 suggesting that its permeation across intestinal mucosa and blood-brain barrier would be significantly restricted. PMID- 12134950 TI - Effects of grapefruit juice and orange juice on the intestinal efflux of P glycoprotein substrates. AB - PURPOSE: The aim of this study is to investigate the effects of 50% ethyl acetate extracts of grapefruit juice (GFJ) and orange juice (OJ) on the transport activity of P-glycoprotein (P-gp) in the rat small intestine. METHODS: The efflux of P-gp substrates from rat everted sac in the absence or presence of verapamil, GFJ, OJ or erythromycin was measured. Rhodamine123, fexofenadine and saquinavir were used as P-gp substrates. P-gp expression levels in the rat jejunum and ileum were determined by Western blot analysis. RESULTS: The efflux of rhodamine123 from the everted sac increased from the apex of the jejunum to the low ileum and the expression of P-gp in the ileum was 2.31-fold higher than that in the jejunum. Verapamil and the 50% GFJ and OJ extracts inhibited the efflux from the intestine of all three drugs tested. Erythromycin decreased the efflux of rhodaminel23 and fexofenadine, but did not affect the efflux of saquinavir in the intestine. CONCLUSIONS: GFJ and OJ extracts inhibited the efflux of P-gp substrates from the small intestine. Therefore, they may enhance the oral bioavailability of P-gp substrates by increasing absorption in the small intestine. PMID- 12134952 TI - Physicochemical characterization of hexetidine-impregnated endotracheal tube poly(vinyl chloride) and resistance to adherence of respiratory bacterial pathogens. AB - PURPOSE: Ventilator-associated pneumonia is a frequent cause of mortality in intensive care patients. This study describes the physicochemical properties of hexetidine-impregnated poly(vinyl chloride) (PVC) endotracheal tube (ET) biomaterials and their resistance to microbial adherence (Staphylococcus aureus and Pseudomonas aeruginosa). METHODS: PVC emulsion was cured in the presence of hexetidine (0-20% w/w) and was characterized in terms of drug release, surface properties (i.e., microrugosity/contact angle), mechanical (tensile) properties, and resistance to microbial adherence. RESULTS: Under sink conditions, hexetidine release from PVC was diffusion-controlled. Increasing the concentration of hexetidine from 1% to 10% (w/w) (but not from 10% to 20% w/w) increased the subsequent rate of drug release. In general, increasing the concentration of hexetidine decreased both the tensile properties and hydrophobicity, yet increased PVC microrugosity. Following hexetidine release (21 days), the surface properties were similar to those of native PVC. The resistance of hexetidine containing PVC (1% or 5%) to microbial adherence (following defined periods of drug release) was greater than that of native PVC and was constant over the examined period of hexetidine release. CONCLUSIONS: ET PVC containing 1% (w/w) hexetidine offered an appropriate balance between suitable physicochemical properties and resistance to microbial adherence. This may offer an approach with which to reduce the incidence of ventilator-associated pneumonia. PMID- 12134951 TI - The structure of PEG-modified poly(ethylene imines) influences biodistribution and pharmacokinetics of their complexes with NF-kappaB decoy in mice. AB - PURPOSE: To study the relationship between structure of poly(ethylene imine-co ethylene glycol), PEI-PEG, copolymers and physicochemical properties as well as in vivo behavior of their complexes with NF-kappaB decoy. METHODS: A variety of copolymers of PEG grafted onto PEI as well as PEI grafted onto PEG were synthesized and their complexes with a double stranded 20mer oligonucleotide were examined regarding size, surface charge, biodistribution and pharmacokinetics. RESULTS: Polyplexes of copolymers were smaller compared to polyplexes formed by non-PEGylated PEI 25 kDa (58 - 334 nm vs. 437 nm for a nitrogen/phosphate ratio of 3.5 and 85 - 308 nm vs. 408 nm for N/P 6.0) and showed reduced zeta potential (-2.5 - 6.4 mV vs. 14.5 mV for N/P 6.0). IV injection into mice revealed liver (35-76% of injected dose), kidney (3 - 22%) and spleen (2 - 16%) to be the main target organs for all injected complexes. Complexes formed by copolymers with few PEG blocks of higher molecular weight (5 kDa and 20 kDa) grafted onto PEI 25 kDa did not show different blood levels from PEI 25 kDa. In contrast, a copolymer with more short PEG blocks (550 Da) grafted onto PEI showed elevated blood levels with an increase in AUC of 62 %. CONCLUSIONS: A sufficiently high density of PEG molecules is necessary to effectively prevent opsonization and thereby rapid clearance from blood stream. PMID- 12134953 TI - Transport and utilization of arginine and arginine-containing peptides by rat alveolar macrophages. AB - PURPOSE: To demonstrate that rat alveolar macrophages (AM) exhibited the PepT1 like transporter for the uptake of arginine (Arg)-containing small peptides and utilized these peptides as direct substrates for nitric oxide (NO) production. NO is an important mediator that, on one hand, protects the lung from bacteria infection and, on the other hand, augments inflammatory lung injury. METHOD: The uptake of small peptides by rat AM was evaluated using fluorescein isothiocyanate (FITC)-labeled (*) peptides (Arg-Lys*, Gly-Sar-Lys*, and beta-Ala-Lys*), high performance liquid chromatography (HPLC) analysis of potential peptide degradation, and known inhibitors of Arg and PepT1 transport. NO production by AM through Arg and Arg-containing peptides was studied with and without inhibition by transport inhibitors. The presence of PepT1-like transporter on AM was evaluated using anti-PepT1 antisera and Western blot analysis. The substrate specificity of Arg-Gly and Arg-Gly-Asp was determined using purified inducible NO synthase (iNOS). The availability of Arg-containing peptides in the lung was determined by HPLC analysis of bronchoalveolar lavage (BAL) fluid. RESULTS: The FITC-labeled peptides were internalized by AM without degradation. The uptake of Arg-Lys*, beta-Ala-Lys*, and Gly-Sar-Lys* was blocked (approximately 50%) by cephradine (an inhibitor of PepT1 for peptide transport) but not by Lys (an inhibitor on cationic amino acid transporter 2B for Arg transport). The NO production by AM through Arg-containing peptides was significantly blocked only by PepT1 inhibitors and by an anti-PepT1 antibody in a dose-dependent manner. These inhibitors had no effect on the AM production of NO using Arg as a substrate. Arg-Gly and Arg-Gly-Asp were found to be direct substrates for iNOS with similar Km and Vmax values to those of Arg. But, the production of NO by AM using these peptides as substrates was 2-fold higher than using Arg as a substrate. Both Arg-Gly and Arg-Gly-Asp were found in the BAL fluid. The presence of a PepT1-like transporter on AM was confirmed by Western blotting. CONCLUSION: This study shows that AM exhibit PepT1-like transporter for small peptide uptake. Arginine-containing peptides, through the PepT1 transporter system, can serve as direct substrates of iNOS for the production of NO by AM. PMID- 12134954 TI - Regional peptide uptake study in the rat intestinal mucosa: glatiramer acetate as a model drug. AB - PURPOSE: To identify regions of the rat intestine that are able to internalize from the lumen oligopeptides, using the model drug glatiramer acetate (GA). METHODS: GA was introduced into rat intestinal sacs and the integrity of GA during uptake was monitored using antibody detection. Sodium docecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting of intestinal homogenates that had been exposed to GA were performed to identify GA presence. An enzyme-linked immunosorbent assay (ELISA) protocol was adapted for GA quantification. Immunohistochemistry was undertaken to examine the rat colonic wall for GA uptake, and confocal microscopy was used to differentiate adsorbed and internalized peptide in cultured colorectal adenocarcinoma cells. RESULTS: The colon and the ileum, respectively, were identified to be the intestinal regions in which GA was maximally preserved during uptake from the lumen. GA was identified to cross the colonic wall from the epithelium to the serosa. Internalization of GA into cultured colonic epithelial cells was demonstrated. CONCLUSIONS: The rat colonic wall was identified to be less proteolytically active toward GA compared to the wall of the more proximal regions of the small intestine. GA has the capacity to penetrate from the lumen into the colonic wall. The maintenance of GA integrity within the wall of the colon offers the potential for local biological activity of the drug. PMID- 12134955 TI - Tissue distribution of [14C]sucrose octasulfate following oral administration to rats. AB - PURPOSE: Aluminum sucrose octasulfate (SOS) is used clinically to prevent ulcers. Under physiologic conditions, the sodium salt of this drug can be formed. Our objective was to determine whether sodium SOS was absorbed when administered orally. In addition to furthering our understanding of aluminum SOS, this study also aimed to clarify how other polyanionic drugs, such as heparin and low molecular-weight heparins, are absorbed. METHODS: [14C]-labeled and cold sodium SOS (60 mg/kg) were given to rats by stomach tube. Radioactivity was counted in gut tissue, gut washes, and nongut tissue (i.e., lung, liver, kidney, spleen, endothelial, and plasma samples) at 3 min, 6 min, 15 min, 30 min, 60 min, 4 h, and 24 h, and in urine and feces accumulated over 4 h and 24 h. RESULTS: Peak radioactivity was found in the tissue and washes of the stomach, ileum, and colon at 6 min, 60 min, and 4 h, respectively, showing progression through the gut. Gut recovery accounted for 84% of the dose at 6 min but only 12% of the dose at 24 h, including counts from feces. Radioactivity was recovered from nongut tissue (averaging 8.6% of the dose) and accumulated urine (18% of the dose at 24 h). When total body distribution was considered, the recovery of radioactivity was greater for the endothelium than for plasma (peak percentage of the dose was 65% at 15 min, 20% at 3 min, 5% from 20 to 240 min for the vena cava, aortic endothelium, and plasma, respectively). CONCLUSIONS: Results indicate that sodium SOS is absorbed, agreeing with previous studies demonstrating the oral absorption of other sulfated polyanions. Endothelial concentrations must be considered when assessing the pharmacokinetics of these compounds. The measured plasma drug concentrations reflect the much greater amounts of drug residing with the endothelium. PMID- 12134956 TI - Preparation and characterization of mono-PEGylated epidermal growth factor: evaluation of in vitro biologic activity. AB - PURPOSE: To isolate mono-PEGylated epidermal growth factor (EGF) isoforms, identify the site of PEGylation, and evaluate the biologic activity of each isoform. METHODS: EGF was PEGylated with an NHS-PEG derivative (Mw 3,400). Mono PEGylated EGF fraction was separated by gel-filtration HPLC and three mono PEGylated EGF isoforms were purified by RP-HPLC. Tryptic digestion mapping of both EGF and mono-PEGylated EGF isoforms was performed to identify the PEGylation sites using RP-HPLC. The digested fragments were also analyzed by matrix-assisted laser desorption and ionization time of flight (MALDI-TOF) mass spectroscopy for further verification of the three PEG conjugation sites. The biologic activity of positional isoforms was evaluated by a cell proliferation assay and a receptor tyrosine kinase activity assay to determine the effect of PEGylation site on its activity. RESULTS: Mono-PEGylated EGF was composed of three positional isomers. Tryptic digestion mapping and MALDI-TOF analysis permitted the identification of the PEGylated site of the three isoforms at N-terminus. Lysine 28, and Lysine 48. PEG-N-terminus EGF, among the three positional isomers, showed the highest activity in a cell proliferation assay and in a receptor-binding assay. CONCLUSION: This study demonstrates that biologic activities of mono-PEGylated EGF isomers are highly dependent upon the site of PEGylation in EGF. PMID- 12134957 TI - Evaluation of capacity-limited first-pass effect through liver by three-points sampling in portal and hepatic veins and systemic artery. AB - PURPOSE: The three-points method was newly developed by sampling the portal and hepatic veins and systemic artery. A model of hepatic local disposition with the Michaelis-Menten elimination was proposed to explain the concentration dependency of the hepatic recovery ratio (F(H)). METHODS: 5-fluorouracil (5-FU) was selected as a model drug. 5-FU was administered orally 90 min after its intraarterial dose. Blood specimens in both femoral artery and hepatic vein were sampled after intraarterial dose, and blood specimens in both femoral artery and portal vein were taken after oral administration. RESULTS: It was shown that F(H) increased with an increase in the input drug concentration into the liver. The mean absorption time (MAT) estimated by nonlinear analysis agreed with the mean local absorption time (t(n)) whereas MAT by linear analysis was significantly smaller than t(a). CONCLUSIONS: The three-points method was newly developed, and the proposed nonlinear model explained well the capacity-limited elimination of 5-FU through the liver. MAT by the nonlinear analysis was in good agreement with t(a). PMID- 12134958 TI - Sex specificity in methadone analgesia in the rat: a population pharmacokinetic and pharmacodynamic approach. AB - PURPOSE: To quantify the extent to which a sex-specific dichotomy in the temporal evolution of the analgesic effect, after intravenous (i.v.) methadone injection in the rat, relates to the pharmacokinetics (PK) and pharmacodynamics (PD) that mediate the dose-to-effect pathway. METHODS: Tail-flick analgesia was measured after i.v. methadone injection (0.35 mg/kg) in female (n = 16) and male (n = 16) Sprague-Dawley rats. The PK were evaluated in separate female (n = 56) and male (n = 56) rats after they had received the same dose of methadone i.v. (0.35 mg/kg). A bicompartmental model described the kinetics and a sigmoid Emax model related drug effect vs. simulated concentrations (pharmacodynamics) at the times of effect measurement. All model parameters as well as interanimal and assay variabilities were estimated with a mixed-effects population method using the program NONMEM. RESULTS: The area under the effect-time curve (AUCE0-120) was (mean +/- interanimal SD) 1859+/-346 min in the females, which was significantly lower than the 4871+/-393 min in the males (P < 0.0001). On the contrary, the profiles of concentration vs. time were higher in females and, therefore, corresponded inversely to the effect vs. time-relative magnitudes. The central volume of distribution, V1, was 1.94+/-0.37 l/kg for female rats and 3.01+/-0.33 l/kg for male rats. Also, the central clearance was 0.077+/-0.006 l/min/kg and 0.102+/-0.005 l/min/ kg, respectively, for female and male rats. Both parameters differed significantly between sexes (P < 0.0001). The pharmacodynamic maximum observed effect parameter (Emax) was 37%+/-29% in female rats and 85%+/-16% in male rats, and these values were significantly different (P < 0.0001). The parameter for the concentration eliciting half of Emax (EC50) was 24.1+/-7.5 microg/l in female rats and 20.3+/-2.9 microg/l in male rats, and the Hill related exponent, gamma, was 6.3+/-3.9 in female rats and 5.5+/-4.1 in male rats. These parameters did not differ significantly (at the P < 0.05 level). CONCLUSIONS: A sex-specific dichotomy in the methadone antinociceptive effect, in the rat, was not proportionally related to plasma concentrations. Each sex corresponded to a distinct subpopulation of the PK parameters and one of the pharmacodynamic parameters (Emax). When the course of a drug involves PK or PD subpopulations, PK/PD modeling can afford the safest prediction of the effect time evolution for a particular dose. PMID- 12134960 TI - A novel phase inversion-based process for the preparation of lipid nanocarriers. AB - PURPOSE: To develop and subsequently evaluate a novel phase inversion-based method used to formulate lipidic nanocapsules. METHODS: Mechanical properties of emulsions prepared by multiinversion phase processes were investigated using a drop tensiometer. Based on the results obtained, a formulation process was developed and a new type of nanocarrier was prepared. These particulates were sized by photon correlation spectroscopy and were visualized by atomic force microscopy and transmission electronic microscopy. Differential scanning calorimetry was also performed. RESULTS: The marginally cohesive but stable interfacial properties of the initial system led to the formulation of lipidic nanocapsules that were composed of a liquid core surrounded by a cohesive interface and were dispersed in an aqueous medium. These related suspensions were stable upon dilution for several months. The control of the formulation parameters allowed an adjustment of the particle mean diameter in the range of 25 100 nm with a monodisperse size distribution. CONCLUSIONS: A novel and convenient process for the preparation of lipidic nanocapsules is described. The structure of these particulates resembles a hybrid between polymeric nanocapsules and liposomes. Such nanocapsules display a strong potential for drug delivery. PMID- 12134959 TI - Comparison of oral absorption and bioavailablity of drugs between monkey and human. AB - PURPOSE: To compare the oral absorption and bioavailability of numerous drugs with a wide variety of physicochemical and pharmacological properties between humans and monkeys and to explore potential reasons for the findings. METHODS: Data for fraction of dose absorbed (Fa) and oral absolute bioavailability (F) were obtained by an extensive Medline database search. Inclusion and exclusion criteria were the same as those reported in our previous studies. A total of 43 and 35 drugs were selected for Fa and F comparison, respectively. The time to reach peak concentration (t(max)), total clearance, and nonrenal clearance were evaluated for 15, 28, and 13 drugs, respectively. RESULTS: Fa values in monkeys were similar or identical to those in humans. Additionally, similar t(max) values were seen in monkeys and humans at comparable doses, thus indicating comparable absorption kinetics between the two species. Conversely, F values in monkeys were generally lower with coumarin being a marked exception. Both total and nonrenal clearances were evaluated and found to be generally greater in monkeys, supporting a generally higher first-pass metabolism and lower F in this species. This was also supported by published data suggesting greater in vitro hepatic drug metabolism for monkeys as compared to humans. CONCLUSIONS: Monkeys appear to be a good predictor of Fa in humans. However, a generally lower F makes monkeys a potentially poor predictor of human F. Higher reported metabolic clearances and hepatic enzyme activities in monkeys may account for this observation. PMID- 12134961 TI - Investigation of pharmaceutical oil/water microemulsions by small-angle scattering. AB - PURPOSE: Stable oil/water (o/w) microemulsions are very effective vehicle systems for dermal administration of drugs having no or low skin penetration. These systems, consisting of oils, a blend of a high and a low HLB surfactant, and a hydrophilic phase (propylene glycol/water), were developed using pharmaceutically acceptable components. METHODS: In this paper, the droplet size of these microemulsions was characterized by means of dynamic light scattering (DLS) and small-angle neutron scattering (SANS). Furthermore, different size parameters obtained by DLS and SANS experiments were compared and discussed. RESULTS: Extremely small droplet radii of approximately 10 nm could be observed. A good agreement between the data of DLS and SANS experiments was found. The kind of oil only marginally influences the droplet size. CONCLUSIONS: Particle size determination via scattering techniques is a useful tool to characterize droplets in microemulsions for dermal drug delivery. PMID- 12134962 TI - New methods characterizing avalanche behavior to determine powder flow. AB - PURPOSE: To characterize the avalanche behavior of different powders and to compare the results of the strange-attractor and novel characterization approaches. METHODS: The following nine different materials were tested: three lactoses, maltodextrin, two microcrystalline celluloses, sodium chloride, sucrose, and glass beads. Morphology, size, and size distribution, true density, bulk and tap density, angle of repose, flow index, and avalanching behavior were quantified for each excipient by scanning electron microscopy, laser time-of flight analysis, helium pycnometer, graduated cylinder, fixed-height funnel, Flodex (Hanson Research Corp., Chatsworth, California) method, and AeroFlow (TSI, Inc., St. Paul, Minnesota), respectively. Environmental factors were controlled, and the avalanches were studied at various speeds. RESULTS: The strange-attractor graph obtained at 1 rotation per 120 s showed that it was difficult to appreciate the flowability differences among 3-mm glass beads, lactose 100, and lactose 325. However, plotting the raw data as a relationship of the time between each avalanche and the inverse of speed revealed a characteristic linear slope for each sample. Furthermore, a new flowability index based on the SD calculated from the raw data gave results that were consistent with Carr's index. A cohesive index also can be determined by avalanche behavior, and it reflects the stability of the rapid particular rearrangements of powder. CONCLUSION: A novel method of evaluating avalanche measurements makes it possible to better characterize powder flowability and to predict powder behavior under working conditions. PMID- 12134963 TI - Crystallization behavior of mannitol in frozen aqueous solutions. AB - PURPOSE: To study the effect of cooling rate, the influence of phosphate buffers and polyvinylpyrrolidone (PVP) on the crystallization behavior of mannitol in frozen aqueous solutions. METHODS: Low-temperature differential scanning calorimetry and powder X-ray diffractometry were used to characterize the frozen solutions. RESULTS: Rapid cooling (20 degrees C/min) inhibited mannitol crystallization, whereas at slower cooling rates (10 degrees C and 5 degrees C/min) partial crystallization was observed. The amorphous freeze-concentrate was characterized by two glass transitions at -32 degrees C and -25 degrees C. When the frozen solutions were heated past the two glass transition temperatures, the solute crystallized as mannitol hydrate. An increase in the concentration of PVP increased the induction time for the crystallization of mannitol hydrate. At concentrations of > or =100 mM, the buffer salts significantly inhibited mannitol crystallization. CONCLUSIONS: The crystallization behavior of mannitol in frozen solutions was influenced by the cooling rate and the presence of phosphate buffers and PVP. PMID- 12134964 TI - Crystallization of mannitol below Tg' during freeze-drying in binary and ternary aqueous systems. AB - PURPOSE: To characterize the phase transitions in a multicomponent system during the various stages of the freeze-drying process and to evaluate the crystallization behavior below Tg' (glass transition temperature of maximally freeze-concentrated amorphous phase) in frozen aqueous solutions and during freeze-drying. METHODS: X-ray powder diffractometry (XRD) and differential scanning calorimetry (DSC) were used to study frozen aqueous solutions of mannitol with or without trehalose. By attaching a vacuum pump to the low temperature stage of the diffractometer, it was possible to simulate the freeze drying process in situ in the sample chamber of the XRD. This enabled real-time monitoring of the solid state of the solutes during the process. RESULTS: In rapidly cooled aqueous solutions containing only mannitol (10% w/w), the solute was retained amorphous. Annealing of frozen solutions or primary drying. both below Tg', resulted in crystallization of mannitol hydrate. Similar effects were observed in the presence of trehalose (2% w/w). At higher concentrations (> or =5% w/w) of this noncrystallizing sugar. annealing below Tg' led to nucleation but not crystallization. However, during primary drying, crystallization of mannitol hydrate was observed. CONCLUSIONS: The combination of in situ XRD and DSC has given a unique insight into phase transitions during freeze-drying as a function of processing conditions and formulation variables. In the presence of trehalose, mannitol crystallization was inhibited in frozen solutions but not during primary drying. PMID- 12134965 TI - Simulation for population pharmacodynamic analysis of dose-ranging trials: usefulness of the mixture model analysis for detecting nonresponders. PMID- 12134966 TI - Quantitative assessment of chimeraplast stability in biological fluids by polyacrylamide gel electrophoresis and laser-assisted fluorescence analysis. PMID- 12134967 TI - Between Scylla and Charybdis: a choice between equally dreadful alternatives. PMID- 12134968 TI - Evolution of ventricular function during permanent pacing from either right ventricular apex or outflow tract following AV-junctional ablation for atrial fibrillation. AB - AIMS: To compare acute and chronic ventricular function between patients, without cardiac failure, paced at either right ventricular apex or outflow tract. METHODS: Twenty patients. 10 paced apically and 10 in the outflow tract, underwent two radionuclide ventriculograms. Eight parameters of systolic or diastolic function were compared at each assessment, as were changes within each group over time. RESULTS: No differences were identified in systolic function between pacing sites 6 weeks after pacing or 23 weeks later. Peak filling rate was lower (P=0.04) at the second assessment with outflow tract pacing. No other diastolic differences were identified. Between assessments, time to peak filling rate prolonged (P=0.04) with apical pacing, while left ventricular area reduced (P=0.04) and peak filling rate decreased (P=0.04) with outflow tract pacing. Septal motion was better preserved with outflow tract pacing. No other parameter changed over time. ECG measures were similar at 14.7 months. CONCLUSIONS: No major differences were identified in systolic function between pacing sites. Some systolic parameters were better preserved with outflow tract pacing and diastolic function deteriorated subtly over time in both groups. Right ventricular pacing adversely affects left ventricular function. PMID- 12134969 TI - Atypical atrial flutters. AB - Typical atrial flutter is due to a counterclockwise macro-re-entry circuit localized in the right atrium with a surface ECG pattern showing predominantly negative F waves in the inferior leads and positive F waves in V1. Recently it has been proposed to classify atrial flutter on the basis of its cavo-tricuspid isthmus dependence rather than on the ECG pattern. Therefore some atrial flutters are considered typical even if the ECG does not exhibit a typical pattern. This is the case for reverse typical atrial flutter, lower loop re-entry and partial isthmus-dependent short circuit flutter. The term atypical flutter refers to a non-isthmus dependent flutter. Usually these patients have had previous cardiac surgery with a right or left atriotomy. Flutter involving a spontaneous right atrial scar is not uncommon. PMID- 12134970 TI - Atypical atrial flutter: clinical features, electrophysiological characteristics and response to radiofrequency catheter ablation. AB - AIMS: To evaluate the clinical and electrophysiological features of atypical atrial flutter (AAF) and its response to radiofrequency catheter ablation. METHODS AND RESULTS: In 90 consecutive patients referred for sustained atrial flutter, bipolar recordings were obtained from the tricuspid annulus, coronary sinus, interatrial septum and left atrium. AAF was defined by the absence of concealed entrainment from the inferior vena cava--tricuspid annulus isthmus. Target sites were identified by early, fragmented or double potentials and by concealed entrainment. Linear lesions were created between target sites and nearby anatomical barriers in a temperature-controlled mode: 20 episodes of AAFs were documented in 19/90 (21%) patients. Mitral valve disease and surgery were significantly more frequent in patients with AAF. Target sites were identified in the right atrial free wall (n=8), interatrial septum (n=6), left atrium (n=4) and coronary sinus (n=2). Effective ablation was obtained in 15/19 patients (79%). After a 15.7 +/- 10.7 month follow-up, AAF recurred in 0/15 patients with a successful and 3/4 (75%) with a failed procedure (P<0.05). CONCLUSIONS: Conventional mapping techniques enable identification of critical sites of AAF and allow successful ablation in the majority of cases. PMID- 12134971 TI - Acute resumption of conduction in the cavotricuspid isthmus after catheter ablation in patients with common atrial flutter. Real-time evaluation and long term follow-up. AB - AIMS: Cavotricuspid isthmus conduction (CIC) is closely associated with the maintenance and recurrence of common atrial flutter (AFL). This study systematically sought to assess the prevalence and characteristics of acute CIC recovery during AFL ablation and to define its predictors and its relationship with the results of long-term follow-up. METHODS AND RESULTS: A total of 124 consecutive patients (105 men, 19 women, mean age 58 +/- 11 years) who underwent successful AFL ablation were included. The procedure endpoint was defined as complete bi-directional CIC block. During an observation period of 30 min, the incidence of CIC restoration was 34.% in patients and 39.8% in applications. It increased with increasing block time and decreased over time during the observation period. Block time in successful burns followed by persistent block was shorter than in those followed by CIC resumption (12 +/- 6 vs 33 +/- 12 s, P<0.0001). A negative correlation between block time and resumption time was found (r = - 0.57, P<0.001). Patients with permanent pacemakers had a higher incidence of acute CIC resumption than those without pacemakers (5/7 vs 29/117, P = 0.007). The AFL recurrence rate was 4.8% during a mean follow-up period of 21 +/- 8 months. Our results suggest that acute CIC resumption may be a potential risk for clinical AFL recurrence during long-term follow-up. CONCLUSIONS: Acute CIC resumption in common AFL ablation varies in terms of incidence and time course. Block time has a predictive value for acute CIC recovery. Observation time can be shortened if block time is short. With longer block time, it is essential to observe for a longer period in order to minimize CIC resumption. PMID- 12134972 TI - Primary closed cooled tip ablation of typical atrial flutter in comparison to conventional radiofrequency ablation. AB - The purpose of this prospective, non-randomized study was to investigate the effectiveness of cooled radiofrequency ablation (cRFA) compared with conventional radio-frequency application (RFA) for ablation of typical atrial flutter (AF). Methods Isthmus ablation was carried out using a system with a circulating fluid path through the ablation tip to control tip temperature in 100 patients with AF. Thirty consecutive AF patients underwent conventional RFA. The number of applications for cRFA was 13.7 +/- 6.9 and for RFA 24.0 +/- 14-5 (P<0.0007) at powers between 35 and 50 W and a tip temperature range of 38-43 degrees C. Ablation duration and fluoroscopy time were 9-9 +/- 4.9 and 22-8 +/- 10.7 min for cRFA, respectively. In contrast, for RFA, ablation duration and fluoroscopy time were 20-6 +/- 14.2 (P<0.0001) and 27.4 +/- 12.7 min, respectively. In 93% of the cooled tip group and in 80% of the control group bi-directional block was confirmed. At 6-months follow-up, recurrence rates were 9 in the cooled-tip group and 7 in the control group, corresponding 10.4% and 25.9%. There were no significant complications. Compared with RFA, cRFA requires lower application numbers. Recurrence rates are low and the overall success rate is high. PMID- 12134973 TI - Radiofrequency ablation of multiple accessory pathways. AB - The aim of the study was to review the clinical and electrophysiological characteristics and results of radiofrequency catheter ablation in patients with multiple accessory pathways to compare them with those of patients with single accessory pathways. Electrophysiological study and radiofrequency catheter ablation were performed in 1010 consecutive cases with Wolff Parkinson White Syndrome. Presence of multiple accessory pathways was documented in 31 patients (3.1%); 30 had two, and 1 had three accessory pathways. Of the 63 accessory pathways, 42 were manifest and 21 concealed. Nine patients had Ebstein's anomaly associated with atrioventricular bypass tracts. The most common combination was right posteroseptal with right free wall bypass tracts (15 patients with 30 accessory pathways). Fifty-one of the sixty-three accessory pathways (81%) were ablated successfully without complications. The duration of the procedure was 100 +/- 58 min and the fluoroscopic time 40 +/- 17 min. A follow up of 5 +/- 3 years after ablation, demonstrated recurrences of six accessory pathways (9.5%). In conclusion, patients with multiple accessory pathways can be treated by radiofrequency ablation in only one session with a high success rate although slightly less than that in patients with a single accessory pathway (81% vs 93%, P<0.01). PMID- 12134974 TI - Anomalies of cardiac venous drainage associated with abnormalities of cardiac conduction system. AB - The embryological development of the superior vena cava (SVC) is complex. If the left common cardinal vein fails to occlude it can, along with the left duct of Cuvier form a left SVC, which frequently drains into the coronary sinus. This may result in abnormalities in the anatomy of this structure. A persistent left SVC occurs in 0.5% of the normal population, and 3% to 4.3% of patients with congenital heart anomalies. The pacemaking tissue of the heart is derived from two sites near the progenitors of the superior vena cava. The right-sided site forms the sinoatrial node, the left-sided site is normally carried down to an area near the coronary sinus. Out of 300 patients with cardiac rhythm abnormalities, who have undergone electrophysiological studies (EPS), or permanent pacemaker insertion (PPI), we identified 12 patients with cardiac conduction abnormalities and anomalies of venous drainage. Anomalies of the coronary sinus may be associated with abnormalities of the conduction system of the heart. This may be due to the close proximity of the coronary sinus to the final position of the left-sided primitive pacemaking tissue. In our series of 300 patients, 4% had an associated left SVC, a similar incidence to that found in previous studies of congenital heart disease. PMID- 12134975 TI - Angioplasty induced myocardial ischaemia prolongs the signal-averaged P-wave duration in single vessel coronary artery disease. AB - Aims The P-wave duration (PWD) has been shown to prolong in conditions associated with elevated left ventricular end-diastolic and left atrial pressures, which also increase during transient coronary artery occlusions such as angioplasty. The aim of this study was to investigate the effects of angioplasty-induced myocardial ischaemia on signal averaged PWD in patients undergoing coronary angioplasty. Methods Eighty-four consecutive adult patients with single-vessel coronary artery disease undergoing elective coronary angioplasty were included. Duration of the P wave before and during coronary angioplasty were evaluated using signal averaged P-wave analysis. Patients were classified in groups according to the artery occluded, as left anterior descending (LAD) Group, right coronary artery (RCA) Group or Others Group (which included obtuse marginal, circumflex or diagonal). Results Patients included in the LAD, RCA and Others groups were similar with respect to clinical characteristics. The mean PWD at baseline was similar in all lesions (P>0.05), whereas mean PWD at inflation was significantly longer in LAD Group compared with RCA (126.1 +/- 9.5 ms vs 118.7 +/ 10.4 ms, P=0.007) and Others (126.1 +/- 9.5 ms vs 116.3 +/- 8.6 ms, P<0.001). The PWD during balloon inflation was significantly prolonged in all groups compared with baseline levels (LAD Group 126.1 +/- 9.6 ms vs 109.7 +/- 8.0 ms; RCA Group 118.7 +/- 10.4 ms vs 108.3 +/- 8.4 ms and Others Group 116.3 +/- 8.6 ms vs 109.7 +/- 6.0 ms, all P values <0.001). Conclusion Signal-averaged PWD significantly increases during single-vessel coronary angioplasty. This increase is more pronounced for LAD lesions. However, the clinical implications of P-wave prolongation during balloon angioplasty and the value of PWD as a measure of ischaemia remains to be clarified. PMID- 12134976 TI - Computationally inexpensive methods for intra-cardiac atrial bipolar electrogram compression. AB - AIM: This paper reports studies of mathematical algorithms for intra-cardiac atrial bipolar electrogram compression suitable with implementation on implantable devices. PATIENTS AND METHODS: Bipolar intra-cardiac electrograms (IEGMs) of high right atrium were obtained from 20 patients who underwent electrophysiological studies for arrhythmias. Four thousand seven hundred and eighty-two seconds of IEGM were collected and divided into three rhythm groups: sinus rhythm (SR), atrial fibrillation (AF) and atrial flutter (AFL). Since mathematical algorithms suitable for use with implantable devices demand low computational cost, we employed piecemeal linear approximation methods (ZOP--Zero Order Prediction and SAPA--Scan Along Polygonal Approximation), and beat detection method (Peak) both or which need small numbers of operations to perform electrogram compression. Compression ratio (CR) and percent root mean square difference (PRD) were used to compare the three methods, with statistical analyses performed using paired t-test. RESULTS AND CONCLUSION: The best performance was obtained using the Peak method which reaches an average CR of 10.6 in the case of SR group, 2.8 for AF, and 3.6 for AFL groups, respectively, while PRD lies below 2% for SR and AFL groups and 6% for the AF group. Results show that, for bipolar electrograms, the Peak method reaches statistically significant better performance (P<0.001) in all cases except for Peak vs SAPA applied to AF (P=0.2). The number of operations necessary to compress the data indicate that time consumption can be reduced to be suitable for real time compression in implantable devices. The Peak method, which was assumed to receive the instant of occurrence of each recognized beat, from the hardware of the device, requires fewer operations than ZOP and SAPA. Increasing the length of electrograms recorded in pacemakers will enhance the amount of information provided by the implantable device, allowing more detailed characterization of the intra-cardiac activity and leading to new perspectives in arrhythmia diagnosis and therapy. PMID- 12134977 TI - Correlation between changes in stroke volume and the paced intracardiac electrogram. AB - AIM: The aim of this study was to examine the relation between cardiac haemodynamics and parameters extracted from the intracardiac electrogram obtained during pacing, i.e. ventricular evoked response. METHODS AND RESULTS: In the course of routinely scheduled right heart catheterization, intracardiac electrograms and cardiac haemodynamics were monitored simultaneously in ten heart transplant patients (two females, aged 48 +/- 12 (18-59) years), using pacemaker telemetry and Swan-Ganz thermodilution techniques. Different haemodynamic states were induced by pacemaker programming (pacing rate changes) and table tilting (postural changes). Forty different haemodynamic states were assessed, with an average of three (2.4) haemodynamic variations in each patient. Linear regression analysis between relative stroke volume changes and relative changes in the R wave slew rate as extracted from the evoked responses revealed a strong, inverse, and highly significant correlation (r= - 0.93, P<0.0001) between those parameters. Similar results were obtained for pacing rate and postural variations alone, respectively. CONCLUSIONS: The strong correlation between changes in stroke volume and R slew rate indicates that paced intracardiac electrograms reflect changes in the size and geometry of the heart. Telemetrically recorded intracardiac electrograms may thus be used non-invasively to assess key aspects of cardiac haemodynamics. PMID- 12134978 TI - Rate-adaptive pacing using intracardiac impedance shows no evidence for positive feedback during dobutamine stress test. AB - BACKGROUND: The Inos2 DDDR pacemaker senses unipolar intracardiac impedance signals for adapting heart rate to meet the haemodynamic needs of physical activity. Theoretically, such pacing devices could be limited by positive feedback since increase in beating frequency per se enhances contractility. We have addressed this problem in patients with chronotropic incompetence who were subjected to a pharmacological stress test. METHODS AND RESULTS: Twelve patients with chronically implanted Inos2 DDDR pacemakers were studied using the standard protocol of stress echocardiography. Most of the patients reached the programmable maximum closed-loop rate during the incremental dobutamine challenge. The time courses for increase in as well as for recovery of heart rate were not different from control patients receiving routine diagnostic stress echocardiography. CONCLUSIONS: In patients implanted with the Inos2 DDDR pacemaker acute stimulation with intravenous dobutamine leads to a robust increase in paced heart rate without any evidence of positive feedback. PMID- 12134979 TI - Interatrial septum pacing avoids the adverse effect of interatrial delay in biventricular pacing: an echo-Doppler evaluation. AB - AIMS: Biventricular pacing (BVP) can improve haemodynamics in patients with dilated cardiomyopathy (DCM) and left bundle branch block by reducing interventricular delay (IVD). Since in DCM interatrial delay (IAD) and IVD frequently coexist, the aim of this study was to test the hypothesis that IVD reduction associated with IAD produces an imbalance between the programmed right atrioventricular (AV) delay and the effective AV delay on the left, and that interatrial septum pacing (IASP) combined with BVP overcomes this adverse effect. METHODS AND RESULTS: IAD, IVD, left and right mechanical atrioventricular delay (L-RMAVD) were measured by echo-Doppler in 29 patients with BVP: 17 patients (Group A) had the atrial lead in the right atrial appendage, 12 patients (Group B) who experienced paroxysmal atrial fibrillation had the atrial lead on the interatrial septum. In Group A, LMAVD was significantly shorter than RMAVD (172 +/- 24 vs 207 +/- 24 ms, P<0.002), IAD was significantly longer than IVD (52 +/- 24 vs 21 +/- 18 ms, P<0.0001). In Group B, no differences were observed between LMAVD and RMAVD (187 +/- 32 vs 185 +/- 28 ms, NS), and between IAD and IVD (11 +/ 12 vs 13 +/- 16 ms, NS). CONCLUSIONS: IAD produces different left and right atrioventricular sequences in BVP. IASP combined with BVP, by resynchronizing both atria and ventricles, is able to avoid this adverse effect. PMID- 12134980 TI - Triboelectric simulation of pacemaker malfunction. AB - We report the occurrence of a triboelectric phenomenon (static electricity) that mimicked malfunction of a contemporary pacemaker by creating an electrocardiograpic artifact virtually identical to the pacemaker stimuli. The diagnosis was established by observing a subtle overshoot of the questionable deflection that was absent from pacemaker stimuli. PMID- 12134981 TI - Prolonged PR interval despite a programmed short sensed AV delay: the role of intra-atrial conduction time. AB - Intracavitary electrogram (IEGM) is a useful tool in the interpretation of difficult pacemaker electrograms. A case of 320 ms P-V spike interval on the surface ECG despite a 110 ms programmed sensed AV delay is presented. Atrial IEGM revealed atrial tachycardia with a significant atrial conduction delay. PMID- 12134982 TI - Haemodynamic changes early in prodromal symptoms of vasovagal syncope. AB - AIMS: Vasovagal syncope (VVS) is often preceded by prodromal symptoms. The haemodynamic changes occurring during the prodrome have not been systematically investigated. The aim of the present study was to investigate the behaviour of blood pressure (BP), heart rate (HR) and sympathetic activity at the beginning of the prodrome in patients with tilt-induced VVS. METHODS AND RESULTS: Sixty-three patients with VVS underwent tilt testing. BP and HR were measured and blood samples for plasma catecholamine determination were obtained during the test. Twenty-seven patients developed syncope of whom all had a prodrome. From the last scheduled measurement before prodromal symptoms to the beginning of the prodrome, both systolic and diastolic BP decreased in all patients (from 105 +/- 16 to 74 +/- 20 mmHg, P<0.001, and from 68 +/- 13 to 51 +/- 12 mmHg, P<0 001, respectively) and HR decreased in 18 (67%) (from 89 +/- 22 to 80 +/- 25 beats/ min P<0 02). At the onset of loss of consciousness both BP and HR showed a further decrease (P<0.001). Plasma adrenaline significantly increased from the last sample before prodromal symptoms to the beginning of the prodrome (P<0.01) and showed a further increase during loss of consciousness (P<0.05), whereas plasma noradrenaline did not increase, as an expression of inhibition of sympathetic neural outflow. CONCLUSION: These results demonstrate that in patients with tilt-induced VVS, BP is consistently decreased at the beginning of prodromal symptoms because of the withdrawal of sympathetic activity, and HR is often reduced, probably because of increased vagal activity. We may infer that similar haemodynamic features also occur during spontaneous VVS. PMID- 12134983 TI - The hidden part of neurally mediated disease. PMID- 12134984 TI - Pterygium surgery: conjunctival rotation autograft versus conjunctival autograft. AB - BACKGROUND AND OBJECTIVES: To compare the safety and efficacy of conjunctival rotation autograft to conjunctival autograft in primary pterygium surgery. PATIENTS AND METHODS: A prospective randomized study was performed of 39 eyes in 31 patients who had undergone pterygium surgery. Nineteen eyes were treated by conjunctival rotation autograft (Group A). Twenty eyes were treated by conjunctival autograft (Group B). Follow up ranged from 8 to 12 months (mean 11 months). Recurrence was defined as postoperative regrowth of 2 mm fibrovascular tissue onto clear cornea in the area of previous pterygium excision. Four eyes were excluded from the study. Delayed wound healing occurred in 11.76% of eyes, and 5.88% of eyes had persistent congestion in Group A. A loose graft was present in 5.55% of eyes, and 5.55% of eyes had dellen formation in Group B. CONCLUSION: We conclude that conjunctival rotation autograft and conjunctival autograft are both equally effective methods to reduce the recurrence rate after pterygium surgery. Conjunctival rotation autograft can be tried as an alternative attractive procedure for pterygium surgery to reduce the chances of recurrence. However, a larger, randomized, prospective double masked study with more patients and a longer follow up will eventually demonstrate the superiority of one procedure over the other. PMID- 12134985 TI - Surgical management of scleral perforation after pterygium excision. AB - OBJECTIVE: Scleral perforation is a rare complication occurring after pterygium excision often leading to scleral ulceration and loss of vision. Our purpose is to evaluate the long-term effectiveness and safety of tenonplasty and amniotic membrane transplantation in the management of scleral perforation after pterygium excision. PATIENTS AND METHODS: We performed a retrospective study on patients with scleral perforation after pterygium excision that underwent tenonplasty and amniotic membrane transplantation at Chang Gung Memorial Hospital from 1997 to 1999 and followed up for at least 12 months postoperatively. RESULTS: There were 6 patients, 1 male and 5 females ranging in ages from 46 to 71 years (mean, 63.3). The interval between pterygium excisions to scleral perforation ranged from 3 to 20 years. There were no recurrences during the follow-up period of 12 to 24 months (average, 18.3 months). CONCLUSIONS: Tenonplasty and amniotic membrane transplantation appears to be a relatively simple, safe, and effective method for treating scleral perforation after pterygium excision. PMID- 12134986 TI - Epiblepharon of the lower eyelid: technique of surgical repair and quantification of excision according to the skin fold height. AB - BACKGROUND AND OBJECTIVE: To describe our excisional technique for lower eyelid epiblepharon to reduce a medial undercorrection and to provide a guide for the excision amount using a classification system of epiblepharon according to the skin fold height. PATIENTS AND METHODS: After classification, an elliptical excision of skin and orbicularis muscle, including that below the lower canaliculus after tarsal suturing of the upper edge of the incised skin, was consecutively performed for 111 eyelids of 58 patients. The widest width of the excisional ellipse was measured. RESULTS: The results were successful in 108 eyelids. The mean widest width of the excisional ellipse was 1.1, 1.7, 2.5, and 3.0 mm in Class I, II, III, and IV epiblepharons, respectively. CONCLUSION: This surgical technique is effective for the correction of epiblepharon, and it is easy to determine the amount of excision. Using this technique, a 1 mm to 3 mm width of excision is sufficient. PMID- 12134987 TI - Intranasal endoscopic diagnosis and treatment in congenital nasolacrimal duct obstruction. AB - BACKGROUND AND OBJECTIVE: To identify the causes of congenital nasolacrimal duct obstruction using intranasal endoscopy. PATIENTS AND METHODS: Eleven children with symptoms of epiphora since birth were selected for treatment. A silicone tube was inserted after identifying the causes of prior probing failures by observing the probing tip directly with intranasal endoscopy. RESULTS: As confirmed through intranasal endoscopic examination, tearing was caused by mucosal obstruction, submucosal passing of the probe, pus collection, and inferior turbinate impaction. The probe passed into the submucosal space in 5 patients and, by performing probing medially instead in the usual posterolateral direction, probing succeeded in 4 patients. One case was accompanied by a bone abnormality; we bent the probe tip into the nasal cavity to form the lacrimal pathway. CONCLUSION: By using intranasal endoscopy, a silicone tube can be inserted under direct visualization and any causative abnormalities can be identified. This can also minimize the intranasal trauma sometimes caused by blind probing. PMID- 12134988 TI - Topical anesthesia with sedation in phacoemulsification and intraocular lens implantation combined with 2-port pars plana vitrectomy in 105 consecutive cases. AB - BACKGROUND AND OBJECTIVE: To evaluate the efficacy of topical anesthesia as an alternative to peribulbar or retrobulbar anesthesia in phacoemulsification and intraocular lens implantation combined with our modified 2-port pars plana vitrectomy technique (phacovitrectomy). PATIENTS AND METHODS: Phacovitrectomy using topical anesthesia (4% lidocaine drops) was prospectively performed in 105 eyes with cataract and varied vitreoretinal pathology. In 75 eyes (71.4%), phacovitrectomy was combined with argon laser photocoagulation (endolaser). Preoperative and intraoperative sedation of varying degrees was necessary. Subjective pain and discomfort were graded from 1 (no pain or discomfort) to 4 (severe pain and discomfort). RESULTS: All patients had grade 1 pain and discomfort during most of the procedure. All patients had grade 2 (mild) pain and discomfort during pars plana sclerotomies, external bipolar cautery, and conjunctival closure. No patient required additional retrobulbar, peribulbar, or sub-Tenon's anesthesia. CONCLUSION: This technique avoids the risk of globe perforation, retrobulbar hemorrhage, and prolonged postoperative akinesia of the eye. With appropriate case selection, topical anesthesia is a safe and effective alternative to peribulbar or retrobulbar anesthesia in phacovitrectomy. PMID- 12134989 TI - A clinical evaluation of an aspheric multifocal intraocular lens and its implications for the developing world. AB - BACKGROUND AND OBJECTIVES: To evaluate the clinical performance of a new aspheric multifocal intraocular lens (IOL), and to compare the results with a corresponding monofocal IOL. PATIENTS AND METHODS: Two groups of 20 patients each were implanted with a multifocal and monofocal IOL and prospectively studied. Distance and near vision, contrast sensitivity, depth of focus, and quality of vision were assessed in both groups. RESULTS: In 85% of multifocal cases and 100% of monofocal cases, the corrected distance vision was 6/9 or better. With distance correction, 80% of multifocal cases had near vision of N9 or better against 10% of monofocal cases. The mean addition required for near vision N6 was +0.8 diopters (D) and +2.6 D in the multifocal and monofocal groups, respectively. Multifocal cases showed significantly decreased contrast sensitivity and increased depth of focus. CONCLUSIONS: Multifocal IOLs are a good option for those with nonexacting visual requirements. The loss in contrast sensitivity seems to be an acceptable trade-off for satisfactory unaided near vision. PMID- 12134990 TI - Late postoperative opacification of hydrophilic acrylic intraocular lenses. AB - BACKGROUND AND OBJECTIVE: To evaluate the incidence of postoperative opacification of hydrophilic acrylic intraocular lenses (IOLs) and discuss the surgical management. PATIENTS AND METHODS: Seventy-two eyes of 72 consecutive patients who received the same type of hydrophilic acrylic IOL (Intraocular Optical International, I.O.I., California, USA) after uneventful phacoemulsification were evaluated retrospectively. Systemic status, follow-up time, recognition time of IOL opacification, time lapse between implantation and explantation, and surgical technique during explantation were reported. RESULTS: IOL opacification was noted in 3 patients (4.1%). Time lapse between implantation and first recognition of opacification was 6.3+/-1.5 months (range: 5-8 months). Two of the patients had insulin-dependent diabetes mellitus and both were on renal dialysis for diabetic nephropathy, whereas one had no systemic disease. Opacified hydrophilic acrylic IOLs were exchanged with Acrysof IOL, and no further opacification occurred after lens exchange. CONCLUSION: Use caution on implantation of hydrophilic IOLs because late opacification is a serious complication requiring further surgery. PMID- 12134991 TI - Surgical treatment of hereditary lens subluxations. AB - BACKGROUND AND OBJECTIVE: To evaluate the effectiveness and results of pars plana vitreolensectomy approach with transscleral fixation of intraocular lens in hereditary lens subluxations. METHODS: Fifteen eyes of 9 consecutive patients with a mean age of 12.8+/-6.2 years (6-26 years) with hereditary lens subluxation were operated on and the results were evaluated in a prospective study. Surgery was considered if best spectacle corrected visual acuity (BSCVA) was less than 20/70. All eyes underwent a 2-port pars plana vitreolensectomy and transscleral fixation of an intraocular lens (IOL). RESULTS: The mean follow-up period was 12.6+/-7.5 months (6-22 months). There was no major intraoperative complication. Preoperatively, 8 eyes (53.3%) had a BSCVA of counting fingers (CF) and 7 eyes (46.6%) had a BSCVA of 20/200 to 20/70. Postoperatively, 14 eyes (93.3%) had a BSCVA of 20/50 or better. None of the patients had IOL decentration or intraocular pressure (IOP) increase during the follow-up period. There was a macular hole formation in 1 eye postoperatively. CONCLUSIONS: The early results of pars plana vitreolensectomy with IOL implantation using scleral fixation technique had shown that it not only promises a rapid visual rehabilitation but it is also a relatively safe method. More serious complications, however, may occur in the long term. PMID- 12134992 TI - Management of pseudophakic retinal detachment with undetectable retinal breaks. AB - BACKGROUND AND OBJECTIVE: Difficulties encountered during the repair of pseudophakic retinal detachment are related to difficulties in peripheral retinal visualization and identification of retinal breaks. The implication of nonvisualized breaks in patients with pseudophakic retinal detachment is associated with lower rates of surgical success. This report decribes the results of a prospective trial to evaluate the efficacy of both scleral buckling surgery in the treatment of pseudophakic retinal detachment with undetected retinal breaks and pars plana vitrectomy techniques in the management of the cases that redetected after primary buckling surgery. PATIENTS AND METHODS: This study represents 25 cases of pseudophakic retinal detachment with undiagnosed retinal breaks. In each case, we performed a scleral buckling that extended over the circumference of the retinal detachment. Pars plana vitrectomy with internal subretinal fluid drainage and long-term tamponade were performed on 7 patients with uncomplicated recurrent retinal detachments after primary buckling surgery. The mean duration of follow up was 32 months. RESULTS: There were 25 eyes (24.5%) of pseudophakic retinal detachment with undiagnosed retinal breaks represented in our pseudophakic retinal detachment cases. Anatomic success was achieved after the initial scleral buckling surgery in 18 eyes (72%). The overall success rate was 92%. The visual acuity was 20/40 or better in 8 patients (32%), 20/80 to 20/40 in 6 patients (24%), 5/200 to 20/80 in 7 patients (28%), and light perception to hand movement in 4 patients (16%). Complications included vitreous hemorrhage, macular pucker, cystoid macular edema, and hypotony with proliferative vitreoretinopathy. CONCLUSION: Scleral buckling surgery in conjunction with cryotherapy is effective in the initial treatment of pseudophakic retinal detachment with undetectable retinal breaks. Pars plana vitrectomy with internal fluid-gas exchange and long-term tamponade can be used to treat these patients with recurrent retinal detachment after primary buckling surgery to get a higher overall success rate. PMID- 12134993 TI - Subconjunctival dislocation of an anterior chamber intraocular lens. AB - Post-traumatic dislocation of an intraocular lens (IOL) is a serious complication, especially after a blunt trauma because the posterior chamber lenses can potentially dislocate subconjunctively. We report a case of subconjunctival dislocation of a Kelman multiflex AC IOL. Such cases require immediate wound repair with IOL explantation to prevent endophthalmitis. PMID- 12134994 TI - Trabeculectomy in the management of Posner-Schlossman syndrome. AB - Posner-Schlossman syndrome (glaucomatocyclitic crisis) is a condition of unknown etiology. Patients present with blurred vision, show minimal anterior chamber activity, and raised intraocular pressure (IOP). Corneal edema may cause colored halos. The condition tends to be recurrent, usually responding to a topical steroid and ocular hypotensives. The eyes appear normal between attacks. We present a patient who had bilateral Posner-Schlossman syndrome and underwent filtering surgery to control raised intraocular pressure in both eyes. During the follow up of more than 4 years, the control of IOP was good and he had no further attacks. PMID- 12134995 TI - Simultaneous choroidal and brain metastasis as initial manifestations of lung cancer. AB - A previously healthy 75-year-old woman developed blurred vision in her left eye and was found to have an amelanotic choroidal metastasis. Cranial magnetic resonance imaging disclosed 2 brain lesions compatible with metastasis. Subsequent evaluation revealed a primary cancer in the lung. Simultaneous metastasis to the choroid and the brain from an occult primary lung cancer is rare. This case underscores the need for a detailed systemic evaluation in a patient with an amelanotic choroidal mass in which a metastasis is a diagnostic consideration. PMID- 12134996 TI - Intraosseous hemangioma of the orbit. AB - A case of intraosseous orbital hemangioma is reported to alert surgeons to possible intraoperative hemorrhage during excision of such a lesion. A slowly enlarging mass was excised from the orbital rim of a 49-year-old woman. The clinical diagnosis was not suspected. In retrospect, roentgenographic findings included a focal honeycombed pattern of the zygomatic bone. Surgery was complicated by persistent low-volume bleeding. Histology showed endothelial-lined blood-filled channels within the bone. Intraosseous orbital hemangioma is a rare, benign neoplasm that can often be diagnosed clinically by characteristic roentgenographic findings. Observation should be considered as a therapeutic alternative when the radiographic diagnosis is established and when ocular function is not compromised. PMID- 12134997 TI - Local methotrexate and dexamethasone phosphate for the treatment of recurrent primary intraocular lymphoma. AB - A 70-year-old patient with recurrent bilateral primary intraocular lymphoma (PIOL) was treated with local injections of methotrexate and periocular dexamethasone phosphate to both eyes over the course of 5 months. Local therapy consisted of a cycle to each eye of 3 intravitreal injections of methotrexate (200 microg in a total volume of 0.1 cc) administered on days 1, 5, and 8, followed by a subtenon injection of dexamethasone phosphate (7.5 mg in a total volume of 0.3 cc) on day 9. This treatment cycle was administered 4 times for the right eye and 3 times for the left eye, at 4 to 6 week intervals. Electroretinography was used to assess retinal function prior to and during each treatment regimen. Complete regression of the lymphomatous infiltrates and resolution of the vitritis was observed with preservation of visual acuity and no changes on electroretinography. The effect was sustained for 24 months after termination of treatment in the absence of systemic chemotherapy, radiation treatments, or maintenance intravitreal injections. Local combined chemotherapy can be used to treat recurrent PIOL, and can serve as successful palliative therapy in patients for whom further treatment with systemic chemotherapy or radiation is contraindicated. PMID- 12134998 TI - Usage and cost of laser trabeculoplasty in the United States. AB - To determine annual usage and costs of laser trabeculoplasty (LTP) in the United States, we reviewed data from the Health Care Financing Administration from 1986 to 2000, using the Part B Extract and Summary System (BESS). The annual number of LTP procedures performed increased to a peak number of 176,670 in 1992 and has declined since that time, with a 57% reduction in the number of procedures performed in 2000 (75,838) compared with the peak number. The total allowed charges declined from a peak of $137,127,436 in 1991 to $27,622,073 in 2000 (80% reduction). The average allowed charge per procedure was highest in 1989 ($893), and by 2000 the average charge ($359) was reduced by 60% compared with the peak charge. The total number of LTP procedures performed in Medicare beneficiaries has decreased in recent years compared with the peak number in 1992. In recent years, there also has been a marked reduction in the total allowed charges and the average charge per procedure for LTP. PMID- 12134999 TI - A simple, safe bimanual technique for subincisional cortex aspiration. AB - We developed a bimanual manipulation technique to facilitate the removal of the subincisional lens cortex in small-incision phacoemulsification cataract surgery. A separate aspiration handpiece, not connected to an aspiration tube, is passed into the anterior chamber through a side-port corneal incision. Under irrigation with a standard infusion/aspiration (I/A) handpiece through a tunnel incision, the cortex is stripped off with the separate handpiece and removed with the I/A handpiece. In 227 eyes, subincisional cortex removal and subsequent capsule polishing was performed safely with the separate handpiece. Rupture of the posterior lens capsule occurred in 3 high-risk eyes. PMID- 12135000 TI - Use of an oil-hydraulic microinjection pump for subretinal infusions. AB - The injection of cell suspensions or drugs into the subretinal space is a new promising option of vitreoretinal surgery for the treatment of degenerative retinal disorders. We used a manual oil-hydraulic microinjection pump to subretinally inject suspensions of retinal pigment epithelial cells in Royal College of Surgeons rats and in patients suffering from age-related macular degeneration with geographic atrophy. The histological examination of the treated rat eyes showed that cell suspensions could be placed precisely in the subretinal space. Intra- and postoperative outcome of the patients in the clinical trial revealed no retinal complications during 6 months of follow up. We suggest the oil-hydraulic microinjection pump to be a valuable instrument for controlled and precisely dosed atraumatic infusion or aspiration of small volumes of cell suspensions, fluids or drugs in vitreoretinal surgery. PMID- 12135001 TI - Aqueous infiltration into an implantable miniaturized telescope. AB - The implantable miniaturized telescope (IMT) is the first intraocular magnifying system proposed for optical correction in patients with age-related macular degeneration (ARMD). The optical component is embedded in a carrying device designed as an intraocular lens that is implanted after cataract surgery. In this study, we report findings on an IMT that was explanted because of aqueous infiltration into its optic and describe the configuration of this device and the surgical technique required for its implantation. The patient, a 75-year-old male with bilateral cataract and nonexudative ARMD, underwent phacoemulsification with implantation of an IMT in the right eye. The rigid device, weighing 46.1 mg in aqueous, has an overall diameter of 13.5 mm and requires implantation through a large limbal incision. It is fixated at the 6 to 12 o'clock meridian. Follow-up examination revealed the presence of numerous droplets inside the IMT optic. The device was explanted and sent to our center for evaluation. A large fissure was found on the carrying device. However, it was unlikely the site for aqueous infiltration. Microdefects at the level of the sealing of the optical cylinder appeared to provide the opening for the inflow of aqueous that formed droplets. Based on the findings of this report the manufacturer has modified the sealing technique to avoid this complication. Current clinical trials are now ongoing to assess the efficacy of this device in providing visual rehabilitation for ARMD patients. PMID- 12135002 TI - Capillary hemangioma of the retina. PMID- 12135003 TI - Endoscopy in children with GERD: "the way we were" and the way we should be. PMID- 12135004 TI - To cut is divine, to poke is sublime. PMID- 12135005 TI - The curse of poor bowel preparation for colonoscopy. PMID- 12135006 TI - Hepatitis A vaccination in patients with chronic liver disease: to screen or not to screen? PMID- 12135007 TI - HCV chronic hepatitis in patients with HIV: clinical management issues. AB - HIV-hepatitis C virus (HCV) coinfection is common and affects more than one-third of all HIV infected persons worldwide. Prevalence among risk categories varies according to shared risk factors for transmission, mainly intravenous drug use (IDU) and hemophiliacs. Chronic HCV infection seems to accelerate the course of HIV disease, resulting in a worsened clinical and immunological progression. At the same time, several studies suggest that HIV disease modifies the natural history of HCV infection, leading to a faster course of progression from active hepatitis to cirrhosis, to end stage liver disease and death. HCV infection mimics opportunistic diseases because its natural history is significantly accelerated in HIV patients. Since highly active antiretroviral therapy (HAART) has slowed the progression of HIV disease and decreased the rate of HIV associated mortality, the prognosis of HIV disease has been modified, and the need to treat HCV coinfection become a significant issue. Because of the poor response rate obtained by either interferon alone or interferon thrice weekly plus ribavirin, the combination of pegylated interferon and ribavirin will probably become the standard of care, although the clinicians should be aware of the overlapping toxicity of nucleoside analogues and ribavirin. Many selected categories of patients pose particular challenges to physicians treating HCV infection: nonresponders to interferon, cirrhotic patients, and patients infected with both HCV and HBV. Liver transplantation in HIV patients is currently under evaluation, but should become the rescue therapy for HIV patients with end stage liver disease. PMID- 12135008 TI - Corticosteroid-sparing treatments in patients with Crohn's disease. AB - Conventional corticosteroid therapy effectively induces remission of Crohn's disease (CD) across a range of disease severity. However, alternative treatments are needed for patients with disease unresponsive to corticosteroids, patients requiring maintenance therapy (for which corticosteroids are ineffective), corticosteroid-dependent patients, and patients with corticosteroid-related toxicities. Thus, corticosteroid-sparing effects are an important clinical endpoint for treatments of CD. Budesonide offers comparable efficacy with less short-term toxicity than conventional corticosteroids (prednisone, prednisolone); this agent has also demonstrated short-term remission maintenance efficacy, while potentially enabling withdrawal of more toxic corticosteroids in corticosteroid dependent patients. However, budesonide has not shown long-term maintenance benefit in clinical studies, and the risk for and implications of budesonide dependency need further evaluation. The immunomodulators, azathioprine and 6 mercaptopurine, are most effective for maintenance of remission in quiescent disease, but may be useful in conjunction with other therapies in inducing remission in active CD; methotrexate may be considered an alternative because of its efficacy in inducing and maintaining remission. In clinical trials, treatment with azathioprine/6-methotrexate has enabled corticosteroid withdrawal in 55% of patients, and methotrexate, in 39% of patients with corticosteroid-dependent CD, while maintaining clinical response. Monitoring for infrequent hematological or hepatic toxicity is recommended during use of these immunomodulators. Infliximab is effective for induction and maintenance of remission in patients with refractory CD participating in randomized placebo-controlled studies and, in open label experience, has enabled corticosteroid withdrawal in approximately three quarters of patients. This biological agent is generally well tolerated. Infusion reactions are the most commonly occurring side effects; such reactions may require adjustment of infusion rate and/or treatment with an antihistamine or acetaminophen. The investigational biological agent CDP-571 has also shown corticosteroid-sparing efficacy in patients with CD. In conclusion, recent research has helped identify corticosteroid-sparing treatments that can provide benefit in patients with corticosteroid-dependent and/or corticosteroid refractory CD or patients at risk for corticosteroid-induced toxicities. PMID- 12135009 TI - Nucleoside analogues for chronic hepatitis B: antiviral efficacy and viral resistance. AB - Nucleoside analogues have been recently introduced in the management of chronic hepatitis B virus (HBV) infection. They mainly act by inhibition of HBV polymerase activity resulting in decrease of viral replication. They are administered orally, and most of them have an excellent tolerance and safety profile. Lamivudine is the only nucleoside analogue licensed for chronic hepatitis B. It has potent activity against HBV, and a 12-month course achieves clearance of hepatitis B e antigen (HBeAg) in 20-30% of HBeAg-positive patients and both biochemical and virological remission in more than 65-70% of HBeAg negative chronic hepatitis B patients. Famciclovir and ganciclovir are less effective, whereas other nucleoside or nucleotide analogues, such as adefovir, entecavir, and emtricitabine, are currently under evaluation. Prolonged effective antiviral therapy is required for eradication of chronic HBV infection, but long term treatment with nucleoside analogues has been found to be associated with progressively increasing rates of viral resistance because of emergence of resistant HBV mutant strains. Virological breakthroughs usually develop after the first 6 months of lamivudine monotherapy, and their rate ranges between 15% and 30% at 12 months and exceeds 50% after 3 yr of therapy. Resistant HBV mutant strains harbor point mutations in the HBV polymerase gene and predominantly in the well-conserved YMDD motif. Although resistant HBV strains may have impaired replication capacity compared with the wild HBV, their clinical significance has not been completely clarified yet. No significant biochemical or clinical event may develop in some cases, whereas severe biochemical breakthroughs with or without deterioration of liver function may develop in others. To date, there is no proven effective therapy for the resistant HBV mutant strains, although adefovir and entecavir seem to be interesting candidates. PMID- 12135010 TI - Association of the myeloperoxidase -463G-->A polymorphism with development of atrophy in Helicobacter pylori-infected gastritis. AB - OBJECTIVES: Although the host factors governing clinical outcomes subsequent to Helicobacter pylori infection have not yet been defined, it has been generally perceived that the development of the atrophic gastritis is determined more by host-related factors than by bacterial factors. It is very important to define the host factors controlling the pathway to atrophic gastritis, which is the precursor of gastric cancer. H. pylori infection is characterized by extensive infiltration of neutrophils. Myeloperoxidase in neutrophils amplifies the oxidative potential of hydrogen peroxides that induce gastric mucosal damage, and thus myeloperoxidase is suspected to play a role in H. pylori-induced gastric injury. The aim of this study was to elucidate the association of host myeloperoxidase genetic polymorphism with atrophic gastritis upon H. pylori infection. METHODS: Biopsy specimens taken from the gastric mucosa were examined histologically using the updated Sydney System in 127 Korean patients. The polymerase chain reaction-restriction fragment length polymorphism assay was used to characterize myeloperoxidase genotypes. RESULTS: The distributions of myeloperoxidase genotypes in Korea were 81.9% for myeloperoxidase (G/G) and 18.1% for myeloperoxidase (G/A). No myeloperoxidase (A/A) genotype was observed in 127 patients studied. The degree of active inflammation increased with the increase in H. pylori colonization. A strong positive correlation between the levels of neutrophil infiltration and gastric atrophy was found in the myeloperoxidase (G/G) genotype but not in myeloperoxidase (G/A). CONCLUSIONS: These results suggest that myeloperoxidase genotype is a critical determinant in the pathogenesis of atrophic gastritis subsequent to H. pylori infection. More work is needed to clarify the functional relevance of myeloperoxidase genetic polymorphisms to gastric cell injury. PMID- 12135011 TI - Endoscopic manifestations of gastroesophageal reflux disease in patients between 18 months and 25 years without neurological deficits. AB - OBJECTIVE: The aim of this study was to determine the prevalence of erosive esophagitis and Barrett's esophagus (BE) among a large cohort of neurologically normal children with gastroesophageal reflux disease (GERD). METHODS: We identified all patients between 18 months and 25 yr of age with GERD who underwent an upper endoscopy with biopsies during 1996-2000 at Texas Children's Hospital. A manual review of the endoscopy and histopathology reports was performed. We excluded patients with mental retardation, cerebral palsy, congenital esophageal anomalies, or those with incomplete endoscopic description of the esophagus or absent histopathological reports on esophageal biopsies if BE was suspected. We analyzed the demographic, endoscopic, and histopathological findings and determined the prevalence of erosive esophagitis (defined by erosions/ulcers) and BE in children without neurological or congenital esophageal anomalies. RESULTS: We identified 402 neurologically normal children without congenital esophageal anomalies who had GERD. Of these, 208 (51.7%) patients were female. The mean age of the patients was 9.7 yr (range 1.5-25 yr). The ethnic distribution was as follows: 64.7% white, 7.7% black, 11.7% Hispanic, and 15.9% other. Erosive esophagitis was reported in 139 patients (34.6%). There were no statistically significant differences in the demographic characteristics of patients with and without erosive esophagitis. At endoscopy, BE was suspected in 11 patients (2.7%) because of the presence of pink colored mucosa at the lower end of the tubular esophagus. Histopathological examination of two to four targeted biopsies taken from these areas revealed fundic gastric glands with no evidence of intestinal metaplasia in any of these 11 patients. CONCLUSIONS: Manifestations of severe disease such as erosive esophagitis were present in more than one-third of patients in a large cohort of neurologically normal children with GERD who underwent endoscopy. Further studies are needed to examine the future subsequent clinical course of these children. On the other hand, BE was absent making it likely that the duration of reflux is a major risk factor for BE. PMID- 12135012 TI - Treatment of chest pain in patients with noncardiac, nonreflux, nonachalasia spastic esophageal motor disorders using botulinum toxin injection into the gastroesophageal junction. AB - OBJECTIVE: The aim of this study was to determine if botulinum toxin injection in the gastroesophageal junction improves symptoms in patients with noncardiac chest pain with a spastic esophageal motility disorder. METHODS: Twenty-nine noncardiac chest pain patients with nonachalasia, nonreflux-related spastic esophageal motility disorders were enrolled in this open label trial of botulinum toxin injection at the gastroesophageal junction. Chest pain was the major complaint in all patients. Symptoms of chest pain, dysphagia, regurgitation, and heartburn were scored before and 1 month after botulinum toxin injection. A response to botulinum toxin was defined as at least a 50% reduction in the symptom score with a possible total chest pain score of 4. The duration of response was defined as the time period, between the time of injection and the point in time, at which the severity of the symptoms returned to the preinjection score. RESULTS: Seventy two percent of the patients responded with at least 50% reduction in chest pain. In these responders, there was a 79% reduction in the mean chest pain score from a preinjection score of 3.7 to a postinjection score of 0.78 (p < 0.0001). The mean duration of the response for chest pain in these patients was 7.3+/-4.1 months (range 1-18 months). There was also a significant reduction in the mean regurgitation score, dysphagia score, and total symptom score (p < 0.0001). CONCLUSIONS: Botulinum toxin injection at the gastroesophageal junction leads to significant symptomatic improvement in patients with spastic esophageal motility disorders whose major complaint is chest pain. PMID- 12135013 TI - Impact of esophageal bile exposure on the genesis of reflux esophagitis in the absence of gastric acid after total gastrectomy. AB - OBJECTIVE: The role of duodenal contents refluxing into the esophagus in producing reflux esophagitis (RE) remains unclear. We aimed to assess the impact of esophageal bile exposure on the genesis of RE in reference to esophageal pH changes in the absence of gastric acid after total gastrectomy. METHODS: Thirty patients having undergone total gastrectomy were studied with concurrent 24-h esophageal pH and bilimetric monitoring, and were divided into two groups based on endoscopic esophageal mucosal findings: without RE, group 1 (n = 24) and with RE, group 2 (n = 6). Esophageal bile exposure was defined as bilirubin absorbance >0.14 detected in the esophagus. RESULTS: 1) The percentage total time of esophageal bilirubin absorbance >0.14 was not correlated with that of esophageal pH >7.0, >7.4, and >8.0. 2) All parameters for esophageal bilirubin absorbance >0.14 in group 2 were significantly higher than those in group 1, whereas none of the parameters for esophageal pH >7.0, >7.4, and >8.0 showed a significant difference between the two groups. 3) The percentage total time of esophageal bilirubin absorbance >0.14 was over 50% in all subjects with RE, and six of seven subjects with that over 50% had RE. CONCLUSIONS: Esophageal bile exposure plays an important role in the genesis of RE in the absence of gastric acid, which is assessed better with the measurement of esophageal bilirubin absorbance than that of esophageal pH. PMID- 12135014 TI - Treatment of idiopathic gastroparesis with injection of botulinum toxin into the pyloric sphincter muscle. AB - OBJECTIVES: We aimed to determine if botulinum toxin injection into the pyloric sphincter improves gastric emptying and reduces symptoms in patients with idiopathic gastroparesis. METHODS: Patients with idiopathic gastroparesis not responding to prokinetic therapy underwent botulinum toxin (80-100 U, 20 U/ml) injection into the pyloric sphincter. Gastric emptying scintigraphy was performed before and 4 wk after treatment. Total symptom scores were obtained from the sum of eight upper GI symptoms graded on a scale from 0 (none) to 4 (extreme). RESULTS: Ten patients were entered into the study. The mean percentage of solid gastric retention at 4 h improved from 27+/-6% (normal < 10%) before botulinum toxin injection into the pylorus to 14+/-4% (p = 0.038) 4 wk after treatment. The symptom score decreased from 15.3+/-1.7 at baseline to 9.0+/-1.9 (p = 0.006) at 4 wk, a 38+/-9% decrease. Improvement in symptoms tended to correlate with improved gastric emptying of solids (r = 0.565, p 0.086). CONCLUSIONS: This initial pilot study suggests that botulinum toxin injection into the pylorus in patients with idiopathic gastroparesis improves both gastric emptying and symptoms. PMID- 12135015 TI - Role of endogenous nitric oxide in regulating antropyloroduodenal motility in humans. AB - OBJECTIVES: Previously we demonstrated the involvement of nitric oxide (NO) in the regulation of interdigestive small intestinal motility in humans. The role of NO in postprandial motility remains to be studied. Therefore, we investigated the effect of the NO synthase inhibitor N(G)-monomethyl-L-arginine (L-NMMA) on antral, pyloric, and small intestinal postprandial motility in healthy volunteers. METHODS: Ten healthy male volunteers (ages 19-29 yr) underwent stationary antropyloroduodenal manometry recording during administration of a placebo or a high dose of L-NMMA (12 mg/kg within 5 min, followed by a maintenance infusion of 6.7 mg/kg/h i.v.) in a double blind, randomized order. Motility was recorded before and after ingestion of a 300-kcal liquid meal. RESULTS: Two and a half minutes (+/-0.4 min) after infusion of L-NMMA, rapidly propagated phase III-like activity was observed in the proximal duodenum in every subject. Mean propagation velocity was 26+/-5 cm/min. The duration of the phase III-like activity increased proximally (9.2+/-1.6 min) to distally (12+/-1.5 min), whereas the frequencies of contractions were similar in all manometric channels (10.8+/-0.3/min). Postprandial duodenal motility was disrupted by phase III-like activity in four of 10 subjects (15-58 min after the meal) during L-NMMA infusion, but not during placebo. Antral or pyloric motility and basal pyloric tone were not significantly altered by L-NMMA, relative to the placebo. CONCLUSIONS: We showed that inhibition of NO biosynthesis triggers the onset of a rapidly propagating phase III and shortens the postprandial period, indicating that NO is involved in the modulation of fasting and postprandial small intestinal motility in humans. PMID- 12135017 TI - Anabolic-androgenic steroids for alcoholic liver disease: a Cochrane review. AB - OBJECTIVES: The objectives were to assess the beneficial and harmful effects of anabolic-androgenic steroids for alcoholic liver disease. METHODS: The Cochrane Hepato-Biliary Group Controlled Trials Register, The Cochrane Library, MEDLINE, EMBASE, and full text searches were combined. Only randomized clinical trials studying patients with alcoholic liver disease were included. Interventions encompassed anabolic-androgenic steroids at any dose or duration versus placebos or no intervention. The statistical package (RevMan and MetaView) provided by The Cochrane Collaboration was used. RESULTS: Five randomized clinical trials (including mainly men with alcoholic hepatitis and/or cirrhosis) were identified. Only one trial was assessed as adequate regarding all methodological quality components. Anabolic-androgenic steroids versus placebos or no intervention demonstrated no significant effects on mortality (relative risk [RR] = 0.96, 95% CI = 0.72-1.28), liver-related mortality (RR = 0.83, 95% CI = 0.60-1.15), complications to the liver disease (RR = 1.25, 95% CI = 0.74-2.10), liver histology, and a number of other outcome measures. Anabolic-androgenic steroids were not associated with a significantly increased risk of nonserious adverse events, but with the seldom occurrence of serious adverse events (RR = 4.54,95% CI = 0.57-36.30). CONCLUSIONS: This systematic review could not demonstrate any significant beneficial effects of anabolic-androgenic steroids on any clinically important outcomes of patients with alcoholic liver disease. PMID- 12135016 TI - Diagnostic accuracy of the 13C-urea breath test for childhood Helicobacter pylori infection: a multicenter Japanese study. AB - OBJECTIVES: In adults, the 13C-urea breath test (UBT) has been widely used as a noninvasive test of Helicobacter pylori infection because of its high sensitivity and specificity. However, this test is less well established in pediatric practice. The optimum cutoff value and test protocol of the 13C-UBT remains to be established in the pediatric population. The primary purpose of this study was to evaluate diagnostic accuracy of the 13C-UBT for children and to determine its optimum cutoff value. METHODS: A total of 220 Japanese children aged 2-16 yr (mean = 11.9) who underwent upper GI endoscopy and gastric biopsies were finally studied. Endoscopic diagnoses included gastritis (n = 131), gastric ulcer (n = 15), duodenal ulcer (n = 72), and combined ulcer (n = 2). H. pylori infection status was confirmed by biopsy tests including histology, urease test, and culture. With the 13C-UBT, breath samples were obtained at baseline and at 20 min after ingestion of 13C-urea without a test meal and were analyzed by isotope ratio mass spectrometry. Based on biopsy tests, a cutoff value was determined using a receiver operating characteristic curve. In 26 children (seven children infected and 19 noninfected), paired breath samples were also measured by nondispersive infrared spectometry (NDIRS). RESULTS: Biopsy tests demonstrated that 89 children (40%) were infected with H. pylori and 131 children were not infected. There were no statistical differences in mean delta 13C values at 20 min between male and female H. pylori-infected and noninfected patients. A receiver operating characteristic analysis defined the best cutoff value as 3.5 per thousand. The overall sensitivity and specificity at a cutoff value of 3.5 per thousand were 97.8% (95% CI = 92.1-99.7%) and 98.5% (95% CI = 96.4-100%), respectively: high sensitivity and specificity were demonstrated in all three age groups (< or =5, 6-10, and > or = 11 yr). There was a close correlation between the values with isotope ratio mass spectrometry and NDIRS methods (r = 0.998, p < 0.001). CONCLUSIONS: The 13C-UBT with a cutoff value of 3.5 per thousand is an accurate diagnostic method for active H. pylori infection. The test with the NDIRS method is inexpensive and might be widely applied in clinical practice. PMID- 12135018 TI - Comparison of two enzyme immunoassays for the assessment of Helicobacter pylori status in stool specimens after eradication therapy. AB - OBJECTIVES: Assessment of Helicobacter pylori antigen in stool specimens has recently been proposed as a valid method for the noninvasive detection of H. pylori infection, especially as posttreatment control. After the development of an enzyme immunoassay based on polyclonal antibodies (Premier Platinum HpSA) a monoclonally based test has recently been developed (FemtoLab H. pylori). The aim of the present study was to assess the diagnostic accuracy of both tests in adult patients undergoing H. pylori eradication therapy. METHODS: Stool samples were collected and the 13C-urea breath test performed in 148 patients (79 females and 69 males aged 21-75 yr) 4-6 wk after eradication therapy. The FemtoLab H. pylori and Premier Platinum HpSA tests were performed in accordance with the manufacturers' protocols. A receiver operator characteristics analysis was performed to define the optimal cutoff value on the basis of the results of the 13C-urea breath test. RESULTS: H. pylori eradication was successful in 113 of the 148 patients (76%). After adjusting the cutoff, the sensitivity of FemtoLab H. pylori was found to be higher than that of the Premier Platinum HpSA (94.3% vs 80.0%, ns). Specificity, positive predictive value, and negative predictive value of the two tests were comparable (93.8% vs 95.6%, 82.8% vs 85.2%, and 98.1% vs 93.8%, respectively). CONCLUSIONS: The new stool antigen test (FemtoLab H. pylori) is an excellent tool for diagnosing H. pylori infection after eradication therapy, and its accuracy is comparable with that of the Premier Platinum HpSA. Adjustment of the cutoff after H. pylori eradication therapy increases the overall accuracy. PMID- 12135019 TI - Genotyping of Helicobacter pylori in paraffin-embedded gastric biopsy specimens: relation to histological parameters and effects on therapy. AB - OBJECTIVES: Colonization with Helicobacterpylori can lead to GI disease. Bacterial genotypes and host factors, such as acid production, can influence the progress of disease. We investigated H. pylori genotypes and histological parameters in the same paraffin-embedded gastric biopsy specimens. METHODS: Paraffin-embedded antrum and corpus biopsy samples from 75 gastroesophageal reflux disease patients were histologically examined and tested for H. pylori vacA (s and m regions), cagA, and iceA genotypes. Patients were investigated at baseline (58 H. pylori positive and 17 H. pylori negative) and after treatment with omeprazole with or without additional antibiotic therapy. RESULTS: Genotyping at baseline was complete in 52 (90%) of the 58 H. pylori positive patients. Multiple genotypes were detected in eight (14%) of these. Genotypes were highly consistent between antrum and corpus biopsy specimens at baseline and in follow-up samples. Genotypes from paraffin sections matched those from corresponding cultured strains in 10 selected cases. In the antrum, the degree of inflammation was associated with vacA s1 and cagA+ genotypes, and the degree of neutrophil activity was associated with the cagA+ genotype. In the corpus, the degree of inflammation was significantly associated with vacA s1, cagA+, and iceA1 genotypes and the degree of atrophy was associated with vacA s1, m1, and cagA+ genotypes, whereas the degree of neutrophil activity was associated with vacA s1 and cagA+ genotypes. vacA s2 and cagA-strains appeared more resistant to antibiotic therapy, irrespective of resistance to clarithromycin. CONCLUSIONS: Our findings confirm the relevance of the H. pylori genotypes for the severity of gastric disease and the efficacy of antibiotic therapy. PMID- 12135020 TI - Impact of bowel preparation on efficiency and cost of colonoscopy. AB - OBJECTIVES: The impact of bowel preparation on the cost and efficiency of colonoscopy is uncertain. The aim of this study was to measure the impact of bowel preparation on total direct cost as well as procedure time and volume. METHODS: For 200 consecutive outpatient colonoscopies in persons with intact colons both at a private university hospital and at a public university hospital, we recorded the time spent suctioning fluid and feces from the colon and the time spent washing the colon to clean the mucosa. We prospectively asked colonoscopists to designate examinations that should be repeated at an interval sooner than would otherwise be recommended because of imperfect preparation. The data were used to perform a cost analysis of the economic effect of bowel preparation on direct costs of colonoscopy. RESULTS: Suctioning fluid and washing occupied 6% and 1.5% of total examination time (including insertion and withdrawal) at the public hospital and 9% and 1.3% at the private hospital. Patients at the public hospital were more likely to have an aborted examination (6.5% vs 1%, p = 0.004) and to be brought back earlier than suggested or required by current practice standards because of imperfect bowel preparation (20% vs 12.5%, p = 0.04). Cost analysis indicated that to complete the initial examinations and the first round of surveillance, imperfect bowel preparation resulted in a 12% increase in costs at the university hospital and a 22% increase at the public hospital. CONCLUSIONS: The increase in colonoscopy costs associated with imperfect preparation is substantial, and seems likely to vary among practices. Aborted examinations and surveillance examinations performed earlier than recommended because of imperfect preparation are appropriate targets for continuous quality improvement programs. More reliable bowel preparations, or measures to improve patient compliance with bowel preparation, could significantly reduce the costs of colonoscopy in clinical practice. PMID- 12135021 TI - Cost-effectiveness analysis of alternative strategies for palliation of distal biliary obstruction after a failed cannulation attempt. AB - OBJECTIVES: Occasionally alternative techniques such as precut sphincterotomy or percutaneous transhepatic cholangiography (PTC) are required to achieve access to the common bile duct. Tradeoffs exist, however, with respect to their complications and costs. Some experts believe that precut sphincterotomy should not be performed at all. We aimed to compare the cost-effectivenesses of metallic biliary stent placement after an initial failed cannulation attempt at ERCP utilizing precut sphincterotomy and placement utilizing PTC for palliation of jaundice. A cost-effectiveness analysis was performed, as viewed from the societal perspective. METHODS: A decision analysis model was designed comparing precut sphincterotomy and PTC approaches for placement of a metallic biliary stent for palliation of jaundice in a patient with inoperable malignant distal biliary obstruction in whom an initial attempt at ERCP cannulation had failed. Baseline probabilities, obtained from the published literature, were varied through plausible ranges using sensitivity analysis. Charges were based on Medicare professional plus facility fees or diagnosis-related group rates for out and inpatients, respectively. The outcome measured was cost per year of life. RESULTS: Sensitivity analysis showed that precut sphincterotomy with subsequent PTC, if necessary, was the most cost-effective strategy provided the precut complication rate was <51% ($9,033/yr), versus $14,741/yr for PTC. CONCLUSIONS: Precut sphincterotomy followed by PTC (if necessary) is the most cost-effective strategy for palliative biliary stenting in the setting of malignant distal biliary obstruction after a failed ERCP attempt. The endoscopic approach is best practiced by experienced endoscopists who minimize precut complication rates. PMID- 12135022 TI - An assessment of the learning curve for precut biliary sphincterotomy. AB - OBJECTIVE: Precut sphincterotomy is considered unsafe when used by inexperienced endoscopists. We sought to determine whether procedural experience with precut sphincterotomy predicted either successful cannulation or development of complications in these patients. METHODS: We describe the experience of 253 consecutive patients who underwent precut biliary sphincterotomy done by one endoscopist between September, 1993 and April, 2001. Data were prospectively collected on procedure indication and outcome. All patients were contacted by phone 30 days after the procedure to determine outcome. We also described precut utilization over time. RESULTS: All 253 precut procedures were divided chronologically into five groups of 50, with 53 in the final group. The rates of successful cannulation after precutting were 88%, 94%, 90%, 88%, and 98%, respectively (p = 0.05 for groups 1-4 vs group 5). Overall complication rates were similar in all groups (12%, 18%, 20%, 12%, and 14%), with no difference in complication severity. Female gender predicted successful cannulation (OR = 2.9 [p = 0.02]), whereas an indication of sphincter of Oddi dysfunction predicted development of complications (OR = 1.7 [p = 0.03]). The total number of ERCP procedures performed increased over time, whereas the proportion of precut sphincterotomies performed decreased. CONCLUSIONS: Although the success rate for precut sphincterotomy may increase with procedural experience, the complication rate does not seem to decrease. Precut sphincterotomy continues to carry an increased complication rate over standard sphincterotomy even when performed by an experienced biliary endoscopist. The need for precut sphincterotomy appears to decrease with increasing ERCP experience. PMID- 12135023 TI - PEG site infections: the emergence of methicillin resistant Staphylococcus aureus as a major pathogen. AB - OBJECTIVE: We reviewed the records of 126 patients who underwent PEG insertion during a 36-month period to determine the etiology of an observed increase in PEG related infections. METHODS: Charts were reviewed to determine predictive factors of infection, the occurrence of infection, and culture results of infected sites. Insertion was performed in all cases using a standard sterile, pull-through technique. Infections were defined as having at least two of the following conditions: peristomal erythema, induration, and purulent discharge. RESULTS: PEG infections occurred in 22 patients. During the first 12-month interval, 0 of 25 patients (0%) had PEG-related infections; during the second 12-month interval, four of 37 patients (10.8%); and during the third 12-month interval, 18 of 64 patients (28.1%) (p < 0.05). Cultures from 14 of 22 peristomal infections grew methicillin resistant Staphylococcus aureus (nine), pseudomonads (three), and other organisms (two). One hundred twenty-four of 126 patients received prophylactic antibiotics or were receiving concomitant antibiotics. Of the infected patients, 21 of 22 (95.5%) received prophylaxis, and 11 of 22 (50%) were receiving concomitant antibiotics before PEG. In the noninfected group, 78 of 104 (75%) received prophylaxis, and 47 of 104 (45.2%) received concomitant antibiotics. CONCLUSIONS: Methicillin resistant Staphylococcus aureus is emerging as a major pathogen in PEG site infections. Further prospective studies are needed to establish whether current prophylactic antibiotic recommendations are adequate. PMID- 12135024 TI - Impairment of psychomotor responses after conscious sedation in cirrhotic patients undergoing therapeutic upper GI endoscopy. AB - OBJECTIVES: The aim of this study was to determine whether the number connection test (NCT) times of a group of cirrhotic patients without clinically overt hepatic encephalopathy and a group of healthy patients without liver disease who were undergoing endoscopy were prolonged after sedation with short acting i.v. benzodiazepines. METHODS: All patients were administered the NCT in a standard fashion for 30 min before sedation for an upper GI endoscopy and then 2 h after sedation postprocedure. Two NCTs were carried out before and 2 h after sedation, and the mean of the tests pre- and postsedation calculated. Based on the upper limit of the 95% CI of the presedation NCT of patients without liver disease as the cut-off level for hepatic encephalopathy, the proportion of cirrhotic patients with subclinical encephalopathy before and after sedation were also determined. RESULTS: A total of 61 consecutive cirrhotic patients who underwent therapeutic upper GI endoscopy completed the study. The mean presedation NCT time was 43.5 s (95% CI = 39.0-48.1 s) and the mean postsedation NCT time 60.0 s (95% CI = 50.7-69.3 s). The difference between the mean pre- and postsedation NCT times was 16.4 s (95% CI = 9.8-23.1 s; p < 0.001). A total of 38 consecutive patients without clinical or biochemical evidence of liver disease who presented for upper GI endoscopy completed the NCT as described for the group of cirrhotic patients. The mean (+/- SD) baseline NCT time was 34.7+/-7.9 s (95% CI = 32.1 37.2 s), whereas the mean postsedation NCT time was 33.7+/-8.5 s (95% CI = 30.9 36.5 s). This difference was not statistically significant (p = 0.177). Using the upper limit of the 95% CI of the mean (37.4 s) of the presedation time in the patients without liver disease as the cut-off between normal and encephalopathy, the number of cirrhotic patients with abnormal presedation NCT times was 33 patients (54.1%), and this number rose to 46 patients (75.4%) after sedation with midazolam. This increase in proportion of cirrhotic patients with prolonged NCT time was statistically significant (p < 0.001). PMID- 12135025 TI - Colonoscopy in octogenarians: a prospective outpatient study. AB - OBJECTIVES: The number of octogenarians (age > or =80 yr) referred for colonoscopy is increasing. Reported success rates regarding colonoscopy completion and adequacy of colonic preparation are poor overall in this group. This may be the result of age-related differences or biases due to retrospective data. The aims of this study were to prospectively determine differences between octogenarians and nonoctogenarians in adequacy of colonic preparation, success in completing colonoscopy, and complications of conscious sedation. METHODS: Prospective cohort study of 250 consecutive outpatients (150 nonoctogenarians and 100 octogenarians) referred for colonoscopy. Colonic preparation tolerance was assessed before colonoscopy, and the success rate and preparation were evaluated after the procedure. Conscious sedation complications were compared. RESULTS: In octogenarians and nonoctogenarians preparation tolerance (86% and 90%, respectively) was similar. Endoscopic success rate was slightly lower in octogenarians (90% vs 99%, p = 0.002). Preparation was poor in 16% of octogenarians compared with 4% of nonoctogenarians (p = 0.001). This was independent of the type of preparation used. Oxygen desaturation was more common in octogenarians (27% vs 19%, p = 0.0007) and associated with a higher meperidine dose (1.05 vs 0.75 mg/kg). No adverse outcomes occurred in either study group. CONCLUSIONS: Colonic preparations were well tolerated and colonoscopic success rates were high in octogenarians and nonoctogenarians. However, poor colonic preparation was four times as likely in octogenarians and was the most important impediment to adequate colonoscopy. PMID- 12135026 TI - Acute pancreatitis in children. AB - OBJECTIVES: Currently, there is no scoring system for predicting severity in acute pancreatitis in children. Our intent was to evaluate the performance of existing scoring systems in children, to develop a system for children, and to examine the etiology of acute pancreatitis in children. METHODS: A chart review of children with acute pancreatitis was conducted at six centers, three serving as criterion centers and three as validation centers. Ranson and Glasgow scores were calculated for each admission. Additional clinical data were collected, and parameters correlating with severity were incorporated into a new scoring system. Performance characteristics were calculated for each system. RESULTS: A total of 301 admissions were reviewed, 202 in the criterion group and 99 in the validation group. Eight parameters were included in a new scoring system for children. The parameters were as follows: age (<7 yr), weight (<23 kg), admission WBC (>18,500), admission LDH (>2,000), 48-h trough Ca2+ (<8.3 mg/dl), 48-h trough albumin (<2.6 g/dl), 48-h fluid sequestration (>75 ml/ kg/48 h), and 48-h rise in BUN (>5 mg/dl). When the cut-off for predicting a severe outcome was set at 3 criteria, the new system had better sensitivity versus Ranson and Glasgow scores (70% vs 30% and 35%, respectively) and a better negative predictive value (91% vs 85% and 85%). The specificity (79% vs 94% and 94%) and positive predictive value (45% vs 57% and 61%) fell slightly. CONCLUSION: The new scoring system performs better in this group than do existing systems. PMID- 12135027 TI - Bowel habits and bile acid malabsorption in the months after cholecystectomy. AB - OBJECTIVE: Bile acid malabsorption has been supposed to play a major pathogenetic role in postcholecystectomy diarrhea. Therefore, the aim of this study was to define the effect of cholecystectomy (CHE) on bowel habits and bile acid absorption. METHODS: Fifty-one patients were prospectively studied before, at 4 wk, and 12 wk after elective CHE for changes of bowel habits, occurrence of diarrhea, and signs of bile acid malabsorption. Bowel habits were assessed by interview. Serum concentrations of 7alpha-hydroxy-4-cholesten-3-one were used as a marker of bile acid malabsorption. Statistics were performed with the McNemar chi2 test for discrete values and Student's paired t test for continuous values. RESULTS: After CHE, there was an increase of patients reporting more than one bowel movement per day (from 22% before CHE to 51% [p < 0.001] and 45% [p < 0.005] at 1 month and 3 months after CHE, respectively) and of patients reporting loose stools (from 2% to 47% [p < 0.001] and 33% [p < 0.001], respectively). Three months after CHE, three patients (6%) reported intermittent diarrhea. Serum levels of 7alpha-hydroxy-4-cholesten-3-one increased from 25.4+/-14.5 ng/ml to 46.5+/-29.5 ng/ml (p < 0.001) and 52.5+/-33.0 ng/ml (p < 0.001), respectively. Unexpectedly, changes of 7alpha-hydroxy-4-cholesten-3-one in serum were unrelated to changes of bowel habits. CONCLUSIONS: CHE results in considerable changes of bowel habits and an increased loss of bile acids from the intestine in some patients. Bile acid malabsorption, however, may not explain changes of bowel habits after CHE. PMID- 12135028 TI - Comparison of oral omeprazole and endoscopic ethanol injection therapy for prevention of recurrent bleeding from peptic ulcers with nonbleeding visible vessels or fresh adherent clots. AB - OBJECTIVES: Omeprazole is a potent inhibitor of gastric acid secretion. Recently it was reported that p.o. omeprazole therapy reduced the rebleeding rate in patients with nonbleeding visible vessels or adherent clots. The aim of this study was to ascertain whether p.o. administration of omeprazole can be an effective alternative to endoscopic injection therapy in peptic ulcers with stigmata of recent hemorrhage. METHODS: A total of 101 patients who had peptic ulcer bleeding based on endoscopic findings of nonbleeding visible vessels or fresh adherent clots were randomly assigned to receive omeprazole (40 mg p.o. every 12 h) or endoscopic ethanol injection therapy. RESULTS: Rebleeding rates were 22.9% (11 of 48) in the omeprazole group and 20.8% (11 of 53) in the endoscopic injection therapy group. The rebleeding rates of clinical significance were 14.6% and 13.2%, respectively. There was no significance difference in the rebleeding rate, requirement for surgery, total units of blood transfused, or mortality between the two groups. CONCLUSIONS: Oral omeprazole administration is comparable to endoscopic ethanol injection therapy for prevention of rebleeding in patients with nonbleeding visible vessels or adherent clots. PMID- 12135029 TI - Characterization of structures with T-lymphocyte aggregates in ileal villi of Crohn's disease. AB - OBJECTIVES: Epithelioid granulomas with transmural inflammation are a characteristic histological feature in Crohn's disease (CD). However, these are not frequently detected by histopathological examination in the biopsy specimens. Here, we demonstrated unique structures with T-lymphocyte aggregates (TLAs) that were specifically found in ileal villi of CD. We characterized the histological and phenotypical features of these structures and assessed the diagnostic value of TLAs for CD. METHODS: Tissue samples were obtained from the inflamed and uninflamed areas of ileal and colonic mucosa of 32 patients with CD. For controls, mucosal samples were obtained from unaffected areas of 18 patients with colon cancer and inflamed and uninflamed areas of 12 patients with ulcerative colitis (UC). RESULTS: 1) In 21 of 32 cases of CD (66%), we found unique structures with lymphoid cell aggregates that were localized in villi of ileum. These structures were not detected in any normal or UC intestine. 2) These structures were composed mainly of T cells, and CD4+ CD45RO+ cells dominated. Neither B cells nor c-kit positive immature lymphocytes were found. Moreover, CD68 positive macrophages were demonstrated in these aggregates, and cells positive for interleukin 18, a pivotal cytokine for Th1 differentiation, were expressed. 3) The aggregates were detected in seven of 13 patients with CD in whom granuloma was not detected by precise histopathological examination. Furthermore, we detected TLAs even in the ilea of two of four CD patients (50%) whose affected lesions were limited to the colon. CONCLUSIONS: We demonstrated TLAs in intestinal villi that may contribute to the pathogenesis of CD. Detection of these lymphocyte aggregates is helpful for diagnosis of CD when granuloma is not found. PMID- 12135030 TI - Budesonide CIR capsules (once or twice daily divided-dose) in active Crohn's disease: a randomized placebo-controlled study in the United States. AB - OBJECTIVES: Budesonide controlled ileal release (CIR) capsules deliver budesonide, a glucocorticosteroid with high topical and low systemic activity, to the distal ileum and the proximal colon. In four previous controlled trials in Crohn's disease, remission rates ranged from 51% to 69%. We sought to evaluate the efficacy and safety of this drug in a population of patients in the United States with Crohn's disease. METHODS: In this multicenter, double blind, randomized trial, 200 patients in the United States with mild to moderate Crohn's disease (Crohn's Disease Activity Index [CDAI] between 200 and 450) involving the distal ileum and/or ascending colon received 9 mg of budesonide CIR once daily, 4.5 mg b.i.d., or placebos for 8 wk. The primary outcome was remission defined by a CDAI of 150 or less. RESULTS: Remission was achieved in 48%, 53%, and 33% with 9 mg once daily, 4.5 mg b.i.d., and placebos, respectively, after 8 wk of treatment. Differences between the groups were not significant. The differences in mean change from baseline CDAI between the combined budesonide and placebo groups was significant (p < 0.05). There was no difference in observed adverse events between treatment groups, although a modest decrease in plasma cortisol levels was observed relative to the placebo (p < 0.01). CONCLUSIONS: Treatment of symptomatic Crohn's disease with budesonide CIR capsules (9 mg daily) was safe, and remission rates were similar to those achieved in previous trials. Although the remission rate did not significantly differ from the placebo response in this study, there was a significant change in the mean CDAI from baseline in the combined treatment groups relative to the placebo. PMID- 12135031 TI - A new method for the quantification of neutrophil and eosinophil cationic proteins in feces: establishment of normal levels and clinical application in patients with inflammatory bowel disease. AB - OBJECTIVES: The aims of this study were 1) to develop a valid method for the measurement of the eosinophil proteins eosinophil cationic protein (ECP) and eosinophil protein X (EPX) and neutrophil proteins myeloperoxidase and human neutrophil lipocalin (HNL) in feces and 2) to investigate their potential role as disease activity markers in inflammatory bowel disease (IBD). METHODS: Feces samples were obtained from 44 apparently healthy individuals (HIs), 18 patients with IBD (11 with ulcerative colitis [UC] and seven with Crohn's disease [CD]), and three with collagen colitis. The granulocyte markers were measured using immunoassays in supernatants from processed feces. RESULTS: ECP, myeloperoxidase, and, to a lesser degree, EPX and HNL were bound to the solid part of feces. However, feces homogenized in an extraction buffer containing the cationic detergent N-cetyl-N,N,N-trimethylammonium bromide allowed an efficient recovery of the proteins (i.e., up to 100-fold increased levels compared to homogenization in saline). All four proteins were stable for at least 7 days at +6 degrees C and at least 3 days at +22 degrees C. The normal fecal geometric mean (95th percentile) levels of ECP, EPX, myeloperoxidase, and HNL were estimated to be, respectively, 1.69 microg/g (6.41), 0.57 microg/g (1.72), 3.54 microg/g (8.77), and 1.97 microg/g (4.91). Markedly increased feces levels of all markers (p < 0.0002), compared to HIs and CD patients, were observed in UC. However, the marker levels in CD patients were significantly increased relative to HIs (p < 0.05 to p < 0.0002). Increased levels of HNL and myeloperoxidase were also observed in the three collagen colitis patients. The discriminative capability between UC patients and HIs was somewhat superior for EPX and myeloperoxidase. CONCLUSIONS: The method described here takes into account the molecular properties of the granule proteins and the heterogeneity in feces consistency, which is a prerequisite for a valid and reproducible measurement of cationic granule proteins. We suggest that EPX and myeloperoxidase, when applied in IBD, are the best eosinophil and neutrophil markers for studying GI inflammation. PMID- 12135032 TI - N-acetyltransferase 1 and 2 genotypes do not predict response or toxicity to treatment with mesalamine and sulfasalazine in patients with ulcerative colitis. AB - OBJECTIVES: 5-Aminosalicylate is metabolized in colonic mucosa by N acetyltransferase 1 (NAT1), and sulfapyridine is metabolized in the liver by N acetyltransferase 2 (NAT2). Common genetic polymorphisms in these enzymes result in rapid and slow acetylation. We determined the association between NAT1 genotype and response to mesalamine and sulfasalazine, as well as between NAT2 genotype and toxicity to sulfasalazine, in a population-based cohort of patients with ulcerative colitis. METHODS: DNA was obtained from 77 white patients with ulcerative colitis from Olmsted County, MN. NAT1 and NAT2 genotyping was performed using microelectronic array devices. Phenotypes were deduced from previously published genotype/phenotype correlations. Clinical response to mesalamine and sulfasalazine, and toxicity to sulfasalazine, were determined by medical record review and associated with NAT1 and NAT2 genotypes. RESULTS: The clinical response rates among 52 patients treated with mesalamine were 67% (31 of 46) for rapid acetylators and 83% (five of six) for slow acetylators (odds ratio = 0.4, 95% CI = < 0.1-3.9, p = 0.65). Similarly, the clinical response rates among 64 patients treated with sulfasalazine were 74% (43 of 58) for rapid acetylators and 67% (four of six) for slow acetylators (odds ratio = 1.4, 95% CI = 0.2-8.6, p = 0.65). The toxicity rates among the 64 patients treated with sulfasalazine were 34% (12 of 35) for slow acetylators and 45% (13 of 29) for rapid acetylators (odds ratio = 0.6, 95% CI = 0.2-1.8, p = 0.65). CONCLUSIONS: NAT1 and NAT2 genotypes did not predict response to mesalamine or sulfasalazine, or toxicity to sulfasalazine. PMID- 12135033 TI - Breaking the cycle: treatment strategies for 163 cases of recurrent Clostridium difficile disease. AB - OBJECTIVE: There is currently uncertainty as to the best treatment for patients with recurrent episodes of Clostridium difficile disease (RCDD). Our objective was to evaluate the success of treatment strategies in a cohort of 163 RCDD patients. METHODS: Data were used from patients who had participated in the placebo arm in two national referral clinical trials evaluating a new combination treatment. Patients with active RCCD were enrolled, prescribed either vancomycin or metronidazole, and randomized to either the investigational biological or a placebo. All patients were observed for at least 2 months for a subsequent episode of RCCD. RESULTS: Of the 163 cases, 44.8% recurred. A tapering course of vancomycin resulted in significantly fewer recurrences (31%, p = 0.01), as did pulsed dosing of vancomycin (14.3%, p = 0.02). A trend (p = 0.09) for a lower recurrence frequency was observed for high-dose (> or =2 g/day) vancomycin and low-dose (< or =1 g/day) metronidazole. Vancomycin was significantly more effective in clearing C. difficile culture and/or toxin by the end of therapy than metronidazole (89% vs 59%, respectively; p < 0.001). CONCLUSIONS: These data show that tapered or pulsed dosing regimens of vancomycin may result in a significantly better cure of RCDD. The persistence of C. difficile spores suggests that additional strategies to restore the normal colonic microflora may also be beneficial. PMID- 12135034 TI - Overnight efficacy of polyethylene glycol laxative. AB - OBJECTIVES: Clinical studies in constipated adult patients have shown that a 17- or 34-g daily dose of polyethylene glycol (PEG) 3350 (MiraLax) is safe and effective for the treatment of constipation, with the best efficacy seen in wk 2 of treatment. The purpose of this study was to determine an optimal dose of PEG to provide satisfactory relief of constipation within 24 h. METHODS: A total of 24 adult study subjects who met Rome II criteria for constipation were randomized in a double-blind, parallel pilot study to receive a single dose of placebo or PEG laxative at doses of 51, 68, or 85 g in 500 ml of flavored water. Over a 72-h period, subjects rated bowel movements (BM), completeness of evacuation, and satisfaction. RESULTS: The 68-g dose seemed to be most satisfactory. Five of six subjects had a BM within 24 h. The time to first BM was 14.8 h for 68 g versus 27.3 h for placebo (p = NS). The time to second BM was 19.2 h versus 47.2 h for 68 g and placebo, respectively (p = 0.003). Of the subjects receiving 68 g of PEG, 50% and 100% reported complete evacuation for the first and second BM, respectively. The average number of BMs in 24 h for placebo, 51 g, 68 g, and 84 g were 0.5, 2.2, 2.2, and 4.2, respectively (p = 0.004). There were no adverse reactions, and no patient reported incontinence or complained of cramps or diarrhea at any dose. There were no changes in measured electrolytes, calcium, glucose, BUN, creatinine, or serum osmolality. CONCLUSIONS: A 68-g dose of PEG laxative seems to provide safe and effective relief in constipated adults within a 24-h period. PMID- 12135035 TI - Serotonin transporter gene polymorphism in irritable bowel syndrome. AB - OBJECTIVES: Serotonin is a key mediator of intestinal peristalsis, and after it is secreted, it is effectively cleansed from the neuronal gap by means of a high affinity substance called serotonin transporter (SERT), which depends on the Na+ and Cl- ions localized in the presynaptic neuronal membranes. The aim of this study was to investigate SERT polymorphism in patients with irritable bowel syndrome (IBS). METHODS: SERT gene polymorphism was assessed by polymerase chain reaction on DNA chains obtained from leukocytes in serum samples from 54 patients diagnosed with IBS and 91 healthy subjects. The polymorphism of two regions (variable number tandem repeats and the SERT gene-linked polymorphic region [5 HTTLPR]) of SERT was assessed. RESULTS: SERT polymorphisms were found to be similar in healthy subjects and IBS patients (p > 0.05). IBS patients were divided into three groups: diarrhea predominant (n = 18), constipation predominant (n = 26), and alternating diarrhea and constipation (n = 10). These groups were compared with respect to gene polymorphism, and it was found that the 5-HTTLPR allele S/S genotype occurred with greater frequency in the constipation predominant group than in the other two subgroups (p < 0.05), and L/S genotype frequency in the diarrhea predominant group was higher than those in the constipation and control groups. CONCLUSIONS: No relationship was found between IBS and SERT gene polymorphism. It is conceivable that the presence of the S/S genotype in IBS patients carries an increased risk of the constipation predominant type of IBS, whereas the presence of the 5-HTTLPR allele L/S genotype carries an increased risk of the diarrhea predominant type. PMID- 12135036 TI - Assessment of hypothalamic-pituitary-adrenal axis function after corticosteroid therapy in inflammatory bowel disease. AB - OBJECTIVES: In patients with inflammatory bowel disease (IBD), little is known about the effect of long term corticosteroid therapy (CT) on the hypothalamic pituitary-adrenal (HPA) axis function. Our aim was to assess HPA axis function in IBD, before the end of CT, during the tapering phase. METHODS: HPA axis function was assessed with cortisol (ng/ml) measurement before (T0) (normal > 100) and 60 min (normal > 210) after 0.25 mg tetracosactide (Synacthen immediat) injection (T60) in 55 consecutive cases of IBD attacks. Abnormal response was defined as a T60 <210. The attacks were separated into two groups according to the result of the Synacthen test (ST). RESULTS: In all, 36 of 55 ST were abnormal. The time for recovery normal HPA axis function was 7.2+/-1.3 months. Duration of disease since onset, past history of surgical or immunosuppressive treatment, severity and extension of the attack, need for surgical or immunosuppressive treatment, total cumulative and mean daily corticosteroid dose, total duration of CT, and steroid dose at the time of ST were not significantly different in the two groups. In multivariate analysis a past history of CT was predictive of abnormal ST (OR = 8.4, 95% CI = 2.2-31.5, p = 0.0009). Among patients with a past history of CT, the time (months) elapsed between the last course of CT was significantly longer in those with normal ST than in those with abnormal ST (45.5+/-13.5 vs 15.4+/ 6.0; p = 0.02), and in multivariate analysis a duration free of CT < 15 months was predictive of abnormal ST (OR = 15.00, CI = 1.23-183.00, p = 0.03). CONCLUSIONS: In all, 65% of the ST were abnormal. These results suggest that ST should be performed before corticosteroid withdrawal, especially in patients with recent past history of CT. PMID- 12135037 TI - Prescreening versus empirical immunization for hepatitis A in patients with chronic liver disease: a prospective cost analysis. AB - OBJECTIVES: There are few prospective studies estimating the prevalence of hepatitis A in chronic liver disease patients. Furthermore, there are minimal cost-comparative data as to whether or not screening for hepatitis A exposure before immunization is an effective fiscal strategy. The objectives of this study were to determine the prevalence of prior hepatitis A infection and to perform a prospective cost analysis for hepatitis A immunization in patients with chronic liver disease. METHODS: This is a prospective cohort study of 100 patients with chronic liver diseases. Patients were screened for potential risk factors for hepatitis A including history of jaundice, socioeconomic status, birth origin, and ethnic background. Each patient underwent testing for evidence of prior infection using an ELISA. Seronegative patients then went on to receive an immunization series. Cost analysis of vaccination without prescreening (universal strategy) was compared to cost analysis of prescreening and selective immunization of those without prior infection (selective strategy). RESULTS: Fifty-three patients (53%) had serological evidence of prior hepatitis A infection (95% CI = 43-63%). Of the risk factors assessed, foreign birth was associated with prior hepatitis A exposure (p = 0.0002). Cost analysis revealed that prescreening for hepatitis A before vaccination was cost saving given the current prevalence. CONCLUSIONS: The seroprevalence of hepatitis A in those with chronic liver diseases was 53%. Cost analysis revealed that screening for hepatitis A before immunization is cost saving, and this strategy should be applied to follow current vaccination guidelines. PMID- 12135038 TI - Association between nonalcoholic fatty liver, markers of obesity, and serum leptin level in young adults. AB - OBJECTIVES: The aim of the present study was to clarify the risk factors for nonalcoholic fatty liver in young adults. METHODS: One thousand two hundred two students, aged 18-21 yr, received matriculation health examinations, including measurements of body mass index and percent body fat and determination of serum levels of ALT at Nagasaki University in 1998. One hundred twenty-nine were found to have borderline or elevated levels of serum ALT, and 105 of the 129 students (75 men and 30 women) were subjected to further analysis for the presence of fatty liver using ultrasonography, by which both the degree of steatosis and the abdominal wall fat index (AFI) corresponding to the ratio of visceral to s.c. adipose tissue (V/S ratio) were evaluated, in addition to determination of the serum level of leptin. RESULTS: Of 105 students, 74 (70%) had fatty liver. The incidence of moderately to severely fatty liver was significantly higher in men than in women. In parameters related to obesity, the close correlation between body mass index and percent body fat was observed in both sexes. The serum level of leptin correlated well with percent body fat and AFI (V/S ratio) in women, whereas it did not correlate with AFI (V/S ratio) in men. Multiple logistic regression analysis indicated that AFI (V/S ratio) was the only independent risk factor for fatty liver in both sexes. CONCLUSIONS: These results suggest that visceral fat distribution is a key risk factor for nonalcoholic fatty liver in young adults. PMID- 12135039 TI - Detection of HCV RNA in gastric mucosa-associated lymphoid tissue by in situ hybridization: evidence of a new extrahepatic localization of HCV with increased risk of gastric malt lymphoma. AB - OBJECTIVES: Mucosa-associated lymphoid tissue (MALT) is absent in the normal gastric mucosa but it can develop in several conditions, such as Helicobacter pylori infection. A certain correlation between hepatitis C virus (HCV) infection and low grade MALT lymphomas has recently been reported. The aim of this study was to investigate the presence of HCV RNA in acquired gastric MALT of HCV infected patients using the in situ hybridization technique. METHODS: Twenty-five patients (16 male and nine female, average age = 56.6 yr [range = 33-75]) affected by chronic HCV hepatitis and with gastric MALT were studied. Giemsa stain and the rapid urease test were also used to evaluate the presence of H. pylori. A polymerase chain reaction product corresponding to the complete 5' noncoding region of the HCV genome was cloned directly in the pCR 1000 vector on gastric biopsies with acquired MALT. RESULTS: Twenty patients showed grade 2 gastric MALT and five showed grade 3, and H. pylori's presence was detected in 18 of 25 patients (72%). Using in situ hybridization, we detected HCV RNA in gastric acquired MALT of seven of 25 patients (28%): five showed grade 2 gastric MALT (two of these were H. pylori negative and the other three were positive), whereas two patients showed grade 3 gastric MALT (without H. pylori infection). CONCLUSIONS: This study shows that HCV not only may colonize gastric MALT but also may permit the development of a grade of acquired MALT, which may represent the first step toward a MALT lymphoma. However, further studies are needed to demonstrate the antigenic role of HCV in the progression of acquired MALT into MALT lymphoma. PMID- 12135040 TI - Effect of the interaction between steatosis and alcohol intake on liver fibrosis progression in chronic hepatitis C. AB - OBJECTIVES: Liver steatosis is a common histopathological finding in patients infected with hepatitis C virus. Patients with chronic hepatitis C having both steatosis and factors causing oxidative stress may be at a higher risk of fibrogenesis. In a subset of patients with a definite date of contamination, our study aimed to assess the potential synergistic interaction between steatosis and factors likely to induce oxidative stress-namely, alcohol intake, iron overload, and drugs. METHODS: Out of 700 anti-HCV-positive screened patients, 142 untreated patients with liver biopsy and with one known risk factor were selected. Liver fibrosis, inflammation, and necrosis were graded according to the Knodell score, and steatosis as moderate to severe if more than 30% of hepatocytes were affected. Drinkers were defined as having daily mean alcohol intakes of more than 30 g in men and 20 g in women. RESULTS: In multivariate analysis, two factors were independently associated with extensive fibrosis: the degree of severity of piecemeal necrosis (OR = 3.27, CI = 1 .17-9.16) and a combination of moderate to severe steatosis and alcohol intake (OR = 7.02, CI = 1.12-44). The median progression rate of fibrosis was about twice as high among drinkers with steatosis than among drinkers without steatosis or nondrinkers with or without steatosis (0.25 vs 0. I vs 0.13 vs 0.08, respectively; p = 0.02). Independent parameters significantly associated with moderate to severe steatosis were body mass index (OR = 1.13, CI = 1.02-1.26) and infection with genotype 3 (OR = 5.5, CI = 1.88-16), but not alcohol consumption. CONCLUSIONS: As well as the key role of the severity of piecemeal necrosis, the study underlines the synergistic interaction between steatosis and even low alcohol consumption as a contributory factor in extensive liver fibrosis. PMID- 12135041 TI - Spectrum of disease in U.S. veteran patients with hepatitis C. AB - OBJECTIVE: Hepatitis C virus (HCV) infection is more prevalent in U.S. veterans attending Veterans Affairs Medical Centers than in the general population. The purpose of this study was to examine the risk factors, psychiatric and substance abuse conditions, and severity of liver disease in veterans with HCV. METHODS: The medical records and liver biopsies of 206 consecutive patients with HCV attending a multidisciplinary medical/psychiatric chronic hepatitis clinic and who met eligibility criteria for interferon alpha-2b therapy were reviewed. RESULTS: The mean age was 46.5+/-6.8 yr and 77% were Vietnam-era veterans. Risk factors included i.v. drug use (64%), blood transfusion (15%), and cocaine use (9%), and were unknown in 12%. The average estimated duration of disease was 24+/ 7.6 yr. A history of alcohol abuse or dependence was identified in 80% of patients. Psychiatric illnesses were present in 60%, the most common being depression and posttraumatic stress disorder. Overall, 89% of patients had documented psychiatric and/or substance abuse diagnoses. Severe fibrosis (stages 3-4) was present in 32% and severe inflammation (grades 2-3) was present in 71% of biopsies. Psychiatric and substance abuse diagnoses did not correlate with severity of liver disease. A total of 145 patients (71%) were prescribed interferon-based treatment. The overall virological sustained response rates were 16% after interferon monotherapy and 28% after interferon/ribavirin therapy. Reasons for not receiving interferon therapy included minimal fibrosis on liver biopsy (37 patients [18%]), worsening medical conditions (nine [4%]), and worsening psychiatric and substance abuse problems (14 [7%]). CONCLUSIONS: Advanced fibrosis is common in this cohort of veteran patients with chronic hepatitis C, and the overwhelming majority of these patients have psychiatric and/or substance abuse diagnoses. Despite these comorbidities, the majority received interferon therapies in the context of a multidisciplinary clinic. These data emphasize the importance of hepatitis C care that includes linkage of medical care and psychiatric services. PMID- 12135042 TI - Thermometer. PMID- 12135043 TI - Challenge in the differentiation between attenuated familial adenomatous polyposis and hereditary nonpolyposis colorectal cancer: case report with review of the literature. AB - The clinical differentiation between hereditary nonpolyposis colorectal cancer (HNPCC) and attenuated familial adenomatous polyposis (AFAP) is very difficult. The 62-yr-old proband presented with duodenal adenocarcinoma. His history of subtotal colectomy for colon cancer, the rarity of duodenal adenocarcinoma in the general population, and his family history of colon cancer made us suspect that he might have FAP. We investigated this family by obtaining medical records and performing gene analysis. The proband had only 10 adenomatous colon polyps when he underwent subtotal colectomy for the cancer, so classic FAP was excluded. His family history included rectal cancer in his brother at 69 yr of age, colon cancer in his mother at 75 yr, and colon cancer in one maternal cousin at 42 yr. Three months after we started to study this family, the proband's 32-yr-old son presented with rectal cancer. His family fulfilled the Amsterdam criteria for HNPCC, but AFAP could not be excluded. Upon gene testing, the proband was negative for APC gene germline mutation, which made AFAP highly unlikely. Moreover, high microsatellite instability (MSI) was detected in his adenomas and cancer tissues. The fulfillment of Amsterdam criteria, the exclusion of FAP and AFAP, and the high MSI established the diagnosis of HNPCC in this family. We also summarize the differences between FAP, AFAP, and HNPCC; extend the graphic description of the MSI mechanism; and propose a diagnostic strategy for HNPCC. PMID- 12135044 TI - Paraneoplastic dysmotility: loss of interstitial cells of Cajal. AB - Autoimmune impairment and destruction of the enteric nervous plexus are thought to play a central role in the pathogenesis of paraneoplastic motility disorders. We present a case of a small-cell lung carcinoma-related paraneoplastic motility disorder associated with abnormal interstitial cells of Cajal networks. Antibodies against c-Kit and protein gene product 9.5 were used to selectively stain interstitial cells of Cajal and the enteric nervous plexus, respectively. A 68-yr-old man presented with anorexia, early satiety, nausea, and weight loss. Investigations revealed gastroparesis, delayed small intestinal transit, and mediastinal lymphadenopathy. The patient was seropositive for type 1 antineuronal nuclear autoantibody and P/Q-type calcium channel antibody. Biopsy of mediastinal lymph nodes revealed metastatic small-cell carcinoma cells that were immunoreactive for c-Kit. Immunohistochemical staining of a full-thickness small intestinal biopsy revealed a relatively intact myenteric plexus but a sparse and disorganized interstitial cells of Cajal network. The histopathology of this case suggests that interstitial cells of Cajal may be a target in the pathogenesis of paraneoplastic motility disorders. PMID- 12135046 TI - Is EUS the answer? PMID- 12135045 TI - Juxtapapillary duodenal diverticula and biliopancreatic disease. PMID- 12135047 TI - Screening for colorectal cancer. PMID- 12135048 TI - Re: Lankisch et al.: the role of hematocrit as a prognostic factor in newly diagnosed acute pancreatitis. PMID- 12135049 TI - Hepatitis C is not a sexually transmissible disease. PMID- 12135050 TI - "Downhill" esophageal varices. PMID- 12135051 TI - Complications of gallstone disease: gallstone ileus. PMID- 12135052 TI - Free intra-abdominal gas associated with octreotide use: intestinal perforation or benign pneumoperitoneum associated with underlying systemic sclerosis? PMID- 12135053 TI - Quantitative assessment of gastric juice urease activity in children infected with Helicobacter pylori. PMID- 12135054 TI - Is a computerized bowel sound auscultation system useful for the detection of increased bowel motility? PMID- 12135055 TI - Hepatitis C, collagenous colitis, and dermatomyositis occurring in the same patient. PMID- 12135056 TI - Beneficial effect of bezafibrate on primary sclerosing cholangitis (three case reports). PMID- 12135057 TI - Intestinal bacterial overgrowth in liver cirrhosis: is it a predisposing X factor for spontaneous ascitic infection? PMID- 12135058 TI - A case of pancreatic adenocarcinoma with novel K-ras mutation and long term survival. PMID- 12135059 TI - Medication adherence and the physician-patient relationship. PMID- 12135060 TI - Moxifloxacin-induced acute liver injury. PMID- 12135061 TI - Chronic hepatitis C, ibuprofen, and liver damage. PMID- 12135062 TI - Caroli's syndrome in two siblings. PMID- 12135063 TI - Sildenafil (viagra) is a risk factor for acute variceal bleeding. PMID- 12135064 TI - Regression of Apo B deficiency in biovular twins with Apo B deficiency and celiac disease after gluten withdrawal. PMID- 12135065 TI - Re: Scintigraphic images. PMID- 12135066 TI - MALT lymphoma with extreme plasma cell differentiation of the rectum. PMID- 12135067 TI - Treatment of autoantibody-associated chronic hepatitis C. PMID- 12135068 TI - Europe comes to Belfast for the FVE's spring assembly. PMID- 12135069 TI - RCVS to move forward with proposals on lifelong learning and assessment. Royal College of Veterinary Surgeons. PMID- 12135070 TI - Estimating the risk of importation of foot-and-mouth disease into Europe. AB - The opinions of a number of recognised world experts on foot-and-mouth disease (FMD) were sought in order to answer key questions relating to the importation of the disease into European countries from countries outside Europe. In addition, their opinions were sought on where in Europe a primary outbreak of FMD was most likely to occur and the number of outbreaks likely to occur within European countries in the next five years. The Balkans group of countries was considered to be the most likely group within Europe to have a primary outbreak of FMD and also most likely to have the highest number of primary outbreaks. Turkey was considered to be the country outside Europe which was most likely to be the source of an outbreak within Europe as a whole, and the illegal importation of livestock was considered to be the most likely route of introduction of FMD into Europe. Results specific to the Islands group of countries, which included the UK and Ireland, suggested that this group was likely to have a mean of one primary outbreak of FMD in the five years from September 2000, and that the importation of foodstuffs by people entering those countries from Turkey was the most likely source of an outbreak. PMID- 12135071 TI - Urinary cortisol:creatinine ratios in healthy horses and horses with hyperadrenocorticism and non-adrenal disease. AB - Urinary cortisol and creatinine concentrations, and the cortisol:creatinine ratio were compared between 12 healthy horses (group 1), 13 horses with Cushing's disease (group 2), and eight horses with dysautonomia syndrome (equine grass sickness) (group 3). The mean (sd) urinary cortisol concentrations were 112 (55.7), 250 (357) and 864 (526) nmol/litre in groups 1, 2 and 3, respectively; the mean (sd) urinary creatinine concentrations were 18.9 (7.3), 12.0 (6.7) and 45.2 (26.4) nmol/litre in groups 1, 2 and 3, respectively, and the mean (sd) ratios were 6.1 (2.6), 19.8 (23.8) and 21.3 (14.5) (x 10(-6)) in groups 1, 2 and 3, respectively. The urinary cortisol and creatinine concentrations were significantly greater in group 3 than in groups 1 and 2, but the ratios were not significantly different, although there was a trend (P=0.076) towards higher values in groups 2 and 3. A diagnostic cut-off in the cortisol:creatinine ratio for the confirmation of Cushing's disease of more than 6.9 x 10(-6) was associated with a diagnostic sensitivity and specificity of 92.3 and 75.0 per cent, respectively, when compared with healthy horses. However, when group 3 horses were included, a cut-off of more than 7.4 x 10(-6) was associated with a diagnostic sensitivity and specificity of 84.6 and 54.5 per cent, respectively. PMID- 12135072 TI - Epidemiology of viral haemorrhagic disease and myxomatosis in a free-living population of wild rabbits. AB - From January 1993 to June 1996, the epidemiology of myxomatosis and viral haemorrhagic disease (VHD) was studied in a free-living population of wild rabbits (Oryctolagus cuniculus) in Spain by means of serological surveys and radiotracking. Myxomatosis was endemic and associated with the breeding period. Its serological pattern was characterised by a 100 per cent prevalence of antibodies in adult rabbits and a rapid increase in antibodies in young rabbits in their first year. No mortality from myxomatosis was detected in adults, and mortality in young rabbits could not be estimated because of interference by predators and scavengers and the deaths of many radiotagged rabbits inside their burrows. VHD was also an endemic disease associated with the breeding period. Adults had a higher prevalence of antibodies against VHD than young rabbits, reaching values of 80 to 90 per cent. During the study, there was an increase in rabbit numbers as a result of a decrease in mortality from predation which was associated with an increase in mortality due to VHD and in the prevalence of antibodies to VHD. Mortality from VHD was lower in rabbits with VHD antibodies than in seronegative rabbits, but some mortality from the disease was also detected in seropositive rabbits. The annual mean mortality rate due to VHD in adult rabbits was estimated to be 21.8 per cent. PMID- 12135074 TI - Use of inhalation anaesthesia for wild mammals in the field. PMID- 12135073 TI - Food safety on the slaughterline: inspection of pig heads. AB - This paper summarises information on the current inspection procedures for pig heads on the slaughterline and their impact on food safety, and considers the implications for food safety of certain lesions. It is argued that although a modified slaughter and inspection technique would decrease the contamination of the carcase with pathogenic microorganisms, leaving lesions in the head undiscovered would be of little or no importance either for human health or for the overall supervision of animal health. PMID- 12135075 TI - Ehrlichia equi (Anaplasma phagocytophila) infection in an adult horse in France. PMID- 12135077 TI - RCVS fees for 2003. PMID- 12135076 TI - Efficacy of prostaglandin F2alpha and misoprostol in the induction of parturition in goats. PMID- 12135078 TI - Roles of the RCVS and BVA. PMID- 12135079 TI - Future of the RCVS library. PMID- 12135080 TI - FMD in a parturient sheep flock. PMID- 12135081 TI - Contagious bovine pleuropneumonia and vaccine strain T1/44. PMID- 12135082 TI - Hunting debate. PMID- 12135083 TI - Tail docking of dogs. PMID- 12135084 TI - Some recent advances in the theory of random vibration. AB - A brief introduction is first given of the history of the technical field of random vibration, tracing back to the works of physicists since the beginning of the twentieth century. This is then followed by an account of more recent developments, with emphasis on nonlinear and quasi-nonlinear systems, and on analytical solutions for the associated Fokker-Planck equation and the generalized Pontryagin equation. The governing equation of a quasi-nonlinear system is linear, but with one or more randomly varying coefficients. The techniques for finding exact probability solutions, approximate probability solutions, conditions for motion stability, and the failure probability and statistics of a system are discussed. PMID- 12135085 TI - Review of the reticulated python (Python reticulatus Schneider, 1801) with the description of new subspecies from Indonesia. AB - The geographically widespread Python reticulatus, the world's longest snake, has been largely neglected by taxonomists. Dwarfed individuals from Tanahjampea Island, Indonesia, differ strikingly in morphology. Phylogenetic relationships were analyzed using a 345-bp fragment of the cytochrome b gene for 12 specimens from different populations. Both genetic differences and morphological characters distinctly revealed two taxonomic subunits. The island populations of Tanahjampea and Selayar form two monophyletic lineages, supported by high bootstrap values, with distinct differences in color pattern and scalation. We consider these forms to represent two new subspecies. The Tanahjampea form is genetically related to populations of the Sunda Islands and mainland Southeast Asia, whereas the Selayar form is related to populations of Southwest Sulawesi. We conclude that, due to strong directional surface currents in this region, gene flow between Tanahjampea and Selayar is prevented. Sea-level changes during the Pleistocene probably contributed to the isolation of the two taxa described. Aspects of ecology and conservation status are briefly discussed. Electronic supplementary material to this paper can be obtained by using the Springer LINK server located at http://dx.doi.org/10.1007/s00114-002-0320-4. PMID- 12135086 TI - Photoperiodic control of germination in the unicell Chlamydomonas. AB - Photoperiodic time measurement is a well-documented adaptation of multicellular plants and animals to seasonal changes in the environment, but it is unclear whether unicellular organisms can exhibit bona fide photoperiodic responses. We demonstrate that the occurrence of zygospore germination of the unicellular alga Chlamydomonas is a genuine photoperiodic response. Germination efficiency is enhanced in long days as compared with short days. While the total amount of light exposure influences the efficiency of germination, the photoperiod is a significant cue for germination. The developmental stage that senses the photoperiod is just prior to mating and during the first days of zygospore development, so there may be a critical window of zygospore maturation that is regulated by photoperiod. Because zygospores are resistant to freezing injury, whereas vegetative cells are not, it is likely that the suppression of germination by short days is an adaptive response for overwintering of Chlamydomonas. Therefore, Chlamydomonas is a single-celled organism that is capable of photoperiodic responses. PMID- 12135087 TI - Molecular phylogeny of cycads inferred from rbcL sequences. AB - The chloroplast gene rbcL was sequenced to elucidate the evolution of the gymnosperm plant order Cycadales. In accordance with traditional systematics, the order Cycadales and the corresponding genera cluster as monophyletic clades. Among them, the genus Cycas forms a basal group. The genetic distances within the genus Encephalartos and between the sister groups Encephalartos, Lepidozamia and Macrozamia, are unexpectedly small, suggesting that the extant species are the result of Miocene and Pliocene speciation. Their distribution in Africa or Australia, respectively, may therefore rather be due to long-distance dispersal than to Cretaceous continental drift, as had previously been assumed. The rbcL sequences also indicate that the colonisation of Madagascar by Cycas thouarsii occurred only recently as the sequences of C. thouarsii and Cycas rumphii from Indonesia are identical. In contrast, the divergence of the Cycadaceae and Zamiaceae apparently occurred in the Mesozoic. PMID- 12135089 TI - The effect of female wings on male courtship behavior in the cricket Gryllus bimaculatus. AB - Sexual dimorphism exists in the shape and the structure of the forewings of the cricket (Gryllus bimaculatus). However, the functional significance of the wings in the G. bimaculatus female has been unclear. In common blue butterflies (Polyommatus icarus), wings in females have been suggested as being important for attracting males. To test whether female crickets need wings for conspecific males to recognize them and initiate mating behavior, we removed all wings from females and observed the behavior of males towards them. Most males (87.5%) showed mating behavior towards the wingless females: they produced courtship song and transferred spermatophores to the wingless females. Similarly, 88.5% of the males showed mating behavior towards intact females. When males were placed with both a wingless female and an intact female, no significant difference was detected in male mate choice. The findings demonstrate that the wing of the G. bimaculatus female is not necessary for female recognition by conspecific males and the initiation of male mating behavior, and that it is not important in male mate choice. PMID- 12135088 TI - A new type of insect infrared organ of low thermal mass. AB - The Australian beetle Acanthocnemus nigricans is attracted by forest fires and has a pair of complex infrared (IR) receptor organs on the first thoracic segment. Each organ consists of a tiny sensory disc (diameter 120-130 microm) which serves as an absorbing structure for IR radiation. The disc is arranged above an air-filled cavity which is located just anteriorly to the coxae of the prothoracic legs. Inside the disc, about 30 multipolar thermoreceptors (warmth receptors) are tightly attached to the cuticle which is directed to the outside. The many dendrites of each multipolar neuron are tightly wrapped around the soma and contain a large number of mitochondria. Absorption of IR radiation by the disc causes an increase in temperature which is measured by the warmth receptors. Therefore, the IR receptors of A. nigricans can be classified as microbolometers with reduced thermal mass and in principle can be compared to the IR organs of pit vipers. PMID- 12135090 TI - AIDS related opportunistic infections, going but not gone. AB - It is now more than two decades since the AIDS epidemic began with a cluster of Pneumocystis carinii pneumonia (PCP) in a community of homosexual men. Since then, many other infections have been characterized as opportunistic infections secondary to HIV infection. These include, but are not limited to, infections with Toxoplasma gondii, Cytomegalovirus (CMV), Mycobacterium avium complex (MAC), and Cryptococcus neoformans. Over the last two decades, there have been dramatic improvements in diagnosis, prevention and treatment of all these infections. As a result, in North America and Western Europe the rates of opportunistic infections secondary to AIDS have decreased substantially. We will review these common opportunistic infections below. PMID- 12135091 TI - Recent progress in drug delivery systems for anticancer agents. AB - Recent progress in understanding the molecular basis of cancer brought out new materials such as oligonucleotides, genes, peptides and proteins as a source of new anticancer agents. Due to their macromolecular properties, however, new strategies of delivery for them are required to achieve their full therapeutic efficacy in clinical setting. Development of improved dosage forms of currently marketed anticancer drugs can also enhance their therapeutic values. Currently developed delivery systems for anticancer agents include colloidal systems (liposomes, emulsions, nanoparticles and micelles), polymer implants and polymer conjugates. These delivery systems have been able to provide enhanced therapeutic activity and reduced toxicity of anticancer agents mainly by altering their pharmacokinetics and biodistribution. Furthermore, the identification of cell specific receptor/antigens on cancer cells have brought the development of ligand or antibody-bearing delivery systems which can be targeted to cancer cells by specific binding to receptors or antigens. They have exhibited specific and selective delivery of anticancer agents to cancer. As a consequence of extensive research, clinical development of anticancer agents utilizing various delivery systems is undergoing worldwide. New technologies and multidisciplinary expertise to develop advanced drug delivery systems, applicable to a wide range of anticancer agents, may eventually lead to an effective cancer therapy in the future. PMID- 12135092 TI - 2,3-dibenzylbutyrolactones and 1,2,3,4-tetrahydro-2-naphthoic acid gamma lactones: structure and activity relationship in cytotoxic activity. AB - Dibenzyl-g-butyrolactone and 1,2,3,4-tetrahydro-2-naphthoic acid gamma-lactone (TNL) derivatives were synthesized and evaluated for cytotoxic activity against some cancer cell lines. It was found that TNL derivatives with a shorter distance between C-4 in ring A and C'-2 in ring C were more cytotoxic, while dibenzyl gamma-butyrolactones with a longer one were nearly inactive. In TNL series, presence of 3,4-dioxy group in ring A and 2-methoxy group in ring C was essential for the enhancement of the activity. PMID- 12135093 TI - Synthesis and antimicrobial activity of new substituted anilinobenzimidazoles. AB - A series of benzimidazole derivatives carrying different heterocycles such as 1,2,3-thiadiazole, 1,3,4-thiadiazole, thiazolidine, 2,3-dihydro-thiazole, 1,3,4 oxadiazole, semicarbazone and substituted thiosemi-carbazones were synthesized. Also a series of 1-methylbenzimidazole carrying hydroxy ethyl-amide, substituted sulfonyl hydrazide and benzoyl hydrazide from aminobenzoyl group at position 2 of 1-methylbenzimidazole were synthesized. The antimicrobial evaluation of some of the new compounds was carried out. PMID- 12135094 TI - Synthesis of new 2-thiouracil-5-sulfonamide derivatives with biological activity. AB - 2-Thiouracil-5-sulfonylchloride 1 reacted with a series of aromatic and heterocyclic amines to give 2a-j. The same compound 1 was reacted with a series of sulphonamides giving different sulphonamides of type 3a-e. On the other hand compound 1 was allowed to react with p-aminoacetophenone givining compound 4 which in turn was allowed to react with derivatives of alkyl thiosemicarbazides to give thiosemicarbazones of type 5a-e, also compound 4 was monobrominated to give compound 6 which in turn was reacted thiosemicarbazones of some aldehydes to give the corresponding thiazole derivatives 7a-f. In the same time compound 4 was reacted with a series of aromatic and heterocyclic aldehydes givining chalcones 8a-g (Claisen-Schemidt reaction). Also compound 4 was allowed to react with a series of aromatic and heterocyclic aldehydes, ethyl cyano acetate and/or malononitrile, and ammonium acetate giving pyridine derivatives 9a-d and 10a-e respectively. The biological effects of some of the new synthesized compounds was also investigated. PMID- 12135095 TI - Structure-activity relationship of triterpenoids isolated from Mitragyna stipulosa on cytotoxicity. AB - Chromatographic separation of the stem bark extract of Mitragyna stipulosa afforded triterpene derivatives ursolic acid (1), quinovic acid (2), quinivic acid 3-O-beta-D-glucopyranoside (3, quinovin glycoside C), quinovic acid 3-O-[(2 O-sulfo)-beta-D-quinovopyranoside] (4, zygophyloside D) and quinovic acid 3-O beta-D-quinovopyranosyl-27-O-beta-D-glucopyranosyl ester (5, zygophyloside B). These five compounds were subjected to the cytotoxicity on MTT assay system. Compound 1 among tested showed the most potent cytotoxicity. Quinovic acid showed less potent cytotoxicity than ursolic acid and sugar linkages to 2 decreased the cytotoxicity. Compound 4 more potent than 3 with indicate that the sulfonyl group significantly enhances the activity. This indicates that the glycosidic linkage in ursane-type triterpenoids has mainly negative effect on cytotoxicity unlike in oleanane-type glycosides. PMID- 12135096 TI - Crystal structure of byakangelicin (C17H18O7). AB - The crystal structure of byakangelicin, one of furanocoumarin aldose reductase inhibitors, was determined by X-ray diffraction method. The crystal is triclinic, with a = 8.114(1), b = 10.194(1), c = 11.428(1)A, a = 111.50(1), beta= 95.57(1), gamma = 112.52(1) degrees , Dx = 1.41, Dm = 1.39 g/cm3, space group P1 and Z = 2. The intensity data were collected by omega-2theta scan method with CuK(a) radiations. The structure was solved by direct method and refined by full matrix least-squares procedure to the final R-value of 0.056. There are two molecules with different conformations in an asymmetric unit. The molecules are kept by two intermolecular O-HO type hydrogen bonds and van der Waal's forces in the crystal. The absolute configuration of the molecules was estimated to S-form by the 'Eta refinement' procedure. PMID- 12135097 TI - Phytochemical constituents from the fruits of Acanthopanax sessiliflorus. AB - Six compounds were isolated from the fruits of Acanthopanax sessiliflorus. Their structures were elucidated as (-)-sesamin, scoparone, protocatechuic acid, ursolic acid, hyperin and 5-hydroxymethylfurfural by physicochemical and spectroscopic analysis. Among them, scoparone, ursolic acid and 5 hydroxymethylfurfural were isolated for the first time from Acanthopanax species. PMID- 12135098 TI - Phytochemical constituents from the herba of Artemisia apiacea. AB - Four compounds of a coumarin, a steroid and two flavonoids were isolated from the herba of Artemisia apiacea by open column chromatography. Their structures were elucidated as artemicapin C, apigenin, daucosterol and cacticin by chemical and spectroscopic analysis. This is the first report of the isolation of these compounds from this plant. PMID- 12135099 TI - Cytotoxic constituents of the leaves of Ixeris sonchifolia. AB - The ethyl acetate extract of the leaves of Ixeris sonchifolia afforded two new and two known sesquiterpene lactone glucosides of the guaiane-type, together with a known alkenol glucoside. The known compounds were identified as ixerin Z (1), ixerin Z-6'-p-hydroxyphenylacetate (2), and (Z)-3-hexen-1-ol-beta-D glucopyranoside (3), respectively. The structures of the new compounds were elucidated as 11,13a-dihydroixerin Z [4, 3-hydroxy-2-oxo-guaia-1(10), 3-dien 5alpha,6beta,7alpha, 11betaH-12,6-olide-3-O-beta-D-glucopyranoside], and 3,10beta dihydroxy-2-oxo-guaia-3,11(13)-dien-1alpha,5alpha,6alpha,7aH-12,6-olide-10-O-beta D-glucopyranoside (5), respectively. The cytotoxicity of these compounds against human hepatocellular carcinoma cell (HepG2) and human melanoma cell (SK-MEL-2) was evaluated. PMID- 12135100 TI - Cytotoxic and antimutagenic stilbenes from seeds of Paeonia lactiflora. AB - Cytotoxic and antimutagenic effects of a novel cis-epsilon-viniferin and five known stilbenes, transresveratrol, trans-epsilon-viniferin, gnetin H, suffruticosols A and B, isolated from the seeds of Paeonia lactiflora Pall. (Paeoniaceae) were determined against five different cancer cell lines, and mutagenicity of N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) in Salmonella typhimurium TA100, respectively. Six stilbenes showed cytotoxic activity in a dose-dependent manner, and especially did potent cytotoxic activity against C6 (mouse glioma) cancer cell with IC50 values ranging from 8.2 to 20.5 microg/ml. trans-Resveratrol showed significant cytotoxic activity against HepG2 (liver hepatoma) and HT-29 (colon) human cancer cell lines with IC50 values of 11.8 and 25.2 g/ml, respectively. In contrast, trans-epsilon-viniferin and cis--viniferin, and gnetin H exhibited marked cytotoxic activity against Hela (cervicse) and MCF 7 (breast) human cancer cell lines with IC50 values of 20.4, 21.5, and 12.9 microg/ml, respectively. However, suffruticosol A and B had less cytotoxic effect against all cancer cells except C6. Meanwhile, six stilbenes exerted antimutagenic activity in a dose-dependent fashion. Of them, trans-resveratrol exhibited the strongest antimutagenic effect against MNNG with IC50 value of 27.0 microg/plate, while other five resveratrol oligomers also did moderate antimutagenic activity with IC50 values ranging from 31.7 to 35.2 microg/plate. PMID- 12135101 TI - Antioxidant constituents from Setaria viridis. AB - The EtOAc and n-BuOH soluble fractions from the aerial part of Setaria vindis showed a strong free radical scavenging activity. Six major compounds were isolated from these fractions. They were identified by spectral data as tricin (1), p-hydroxycinnamic acid (2), vitexin 2''-O-xyloside (3), orientin 2''-O xyloside (4), tricin-7-O-beta-D-glucoside (5) and vitexin 2"-O-glucoside (6). Among these compounds, 4 and 5 exhibited strong free radical scavenging activities on 1,1-diphenyl-2-picrylhydrazyl (DPPH). We further studied the effects of these isolated compounds on the lipid peroxidation in rat liver microsomes induced by non-enzymatic method. As expected, 4 and 5 exhibited significant inhibition on ascorbic/Fe2+ induced lipid peroxidation in rat liver microsomes. PMID- 12135102 TI - Isoflavonoids from the rhizomes of Belamcanda chinensis and their effects on aldose reductase and sorbitol accumulation in streptozotocin induced diabetic rat tissues. AB - Aldose reductase, the key enzyme of the polyol pathway, is known to play important roles in the diabetic complication. The inhibitors of aldose reductase, therefore, would be potential agents for the prevention of diabetic complications. To evaluate active principles for the inhibition of aldose reductase from the rhizomes of Belamcanda chinensis, twelve phenolic compounds were isolated and tested for their effects on rat lens aldose reductase. As a result, isoflavones such as tectorigenin, irigenin and their glucosides were found to show a strong aldose reductase inhibition. Tectoridin and tectorigenin, exhibited the highest aldose reductase inhibitory potency, their IC50 values, being 1.08 x 10(-6) M and 1.12 x 10(-6) M, respectively, for DL-glyceraldehyde as a substrate. Both compounds, when administered orally at 100 mg/kg for 10 consecutive days to streptozotocin-induced diabetic rats, caused a significant inhibition of sorbitol accumulation in the tissues such as lens, sciatic nerves and red blood cells. Tectorigenin showed a stronger inhibitory activity than tectoridin. From these results, it is suggested that tectorigenin is attributed to be a promising compound for the prevention and/or treatment of diabetic complications. PMID- 12135103 TI - Antioxidative flavonoids from leaves of Carthamus tinctorius. AB - A total of eight flavonoids (1-8), including a novel quercetin-7-O-(6''-O-acetyl) beta-D-glucopyranoside (6) and seven known flavonoids, luteolin (1), quercetin (2), luteolin 7-O-beta-D-glucopyranoside (3), luteolin-7-O-(6''-O-acetyl)-beta-D glucopyranoside (4) quercetin 7-O-beta-D-glucopyranoside (5), acacetin 7-O-beta-D glucuronide (7) and apigenin-6-C-beta-D-glucopyrano syl-8-C-beta-D glucopyranoside (8), have been isolated from the leaves of the safflower (Carthamus tinctorius L.) and identified on the basis of spectroscopic and chemical studies. The antioxidative activity of these flavonoids was evaluated against 2-deoxyribose degradation and rat liver microsomal lipid peroxidation induced by hydroxyl radicals generated via a Fenton-type reaction. Among these flavonoids, luteolin-acetyl-glucoside (4) and quercetin-acetyl-glucoside (6) showed potent antioxidative activities against 2-deoxyribose degradation and lipid peroxidation in rat liver microsomes. Luteolin (1), quercetin (2), and their corresponding glycosides (3 & 5) also exhibited strong antioxidative activity, while acacetin glucuronide (7) and apigenin-6,8-di-C-glucoside (8) were relatively less active. PMID- 12135104 TI - Potent antimutagenic and their anti-lipid peroxidative effect of kaikasaponin III and tectorigenin from the flower of Pueraria thunbergiana. AB - The MeOH extract of Pueraria thunbergiana (Leguminosae) flowers and its fractions were subjected to Ames test to test the antimutagenicity. EtOAc fraction (1 mg/plate) decreased the number of revertants of Salmonella typhymurium TA100 by 95% against aflatoxin B, (AFB1). Phytochemical isolation of the EtOAc fraction afforded four isoflavonoids (tectorigenin, glycitein, tectoridin and glycitin) and one saponin (kaikasaponin III). Though the three isoflavonoids other than tectoridin showed significant antimutagenicity, the activity of kaikasaponin III was the most potent. Kaikasaponin III (1 mg/plate) decreased the number of revertants of S. typhymurium TA100 by 99% against AFB, but by 75% against N methyl-N'-nitro-N-nitrosoguanidine (MNNG). Tectorigenin (1 mg/plate) inhibited the AFB1-induced mutagenicity by 90% and MNNG-induced one by 76%. Glycitein and glycitin were less active than tectorigenin and kaikasaponin III. This result suggested that kaikasponin III prevents the metabolic activation of AFB1 and scavenge electrophilic intermediate capable of mutation. The two components with potent activities, tectorigenin and kaikasaonin III, significantly prevented the malondialdehyde formation caused by bromobenzene in the rat. PMID- 12135105 TI - Anti-platelet effect of the constituents isolated from the barks and fruits of Magnolia obovata. AB - In the course of our work on anti-platelet constituents from plants, five phenolic compounds, magnolol, honokiol, obovatol, methyl caffeate, and syringin, were isolated from the methanol extracts of the barks and fruits of Magnolia obovata. The compounds were identified based on the spectroscopic data. Methyl caffeate was isolated for the first time from the genus Magnolia and it showed 3 approximately 4-folds higher potency than ASA. The activities of obovatol and honokiol were comparable to ASA. Magnolol and syringin showed only very mild inhibitory effects to all the stimulators. PMID- 12135106 TI - Effects of sophoraflavanone G, a prenylated flavonoid from Sophora flavescens, on cyclooxygenase-2 and in vivo inflammatory response. AB - Previously, several prenylated flavonoids having a C-8 lavandulyl moiety were found to inhibit cyclooxygenase-1 (COX-1) as well as 5-lipoxygenase (5-LOX), and sophoraflavanone G was the most potent inhibitor against these eicosanoid generating enzymes among 19 prenylated flavonoids tested. In this investigation, effects of sophoraflavanone G on COX-2 induction from RAW 264.7 cells and in vivo inflammatory response were studied. Sophoraflavanone G inhibited prostaglandin E2 (PGE2) production from lipopolysaccharide (LPS)-treated RAW cells by COX-2 down regulation at 1-50 uM. Other prenylated flavonoids including kuraridin and sanggenon D also down-regulated COX-2 induction at 10-25 uM, while kurarinone and echinoisoflavanone did not. In addition, sophoraflavanone G showed in vivo anti inflammatory activity against mouse croton oil-induced ear edema and rat carrageenan paw edema via oral (2-250 mg/kg) or topical administration (10-250 microg/ear). Although the potencies of inhibition were far less than that of a reference drug, prednisolone, this compound showed higher anti-inflammatory activity when applied topically, suggesting a potential use for several eicosanoid-related skin inflammation such as atopic dermatitis. PMID- 12135107 TI - Anti-rheumatoid arthritis effect of the Kochia scoparia fruits and activity comparison of momordin lc, its prosapogenin and sapogenin. AB - MeOH extract of Kochia scoparia was fractionated into CHCl3-, EtOAc- and BuOH extracts and the last fraction were hydrolyzed by 3%-NaOH (MeOH-H2O) to compare the bioactivities on antinociceptive and anti-inflammatory effects. Silica gel column chromatography of BuOH fraction afforded a large amount of 3-O-beta-D xylopyranosyl (1-->3)-beta-D-glucuronopyranosyl oleanolic acid (momordin lc, 4) and that of acid hydrolysate of BuOH fraction gave 3-O-beta-D-glucuronopyranosyl oleanolic acid (momordin lb, 3), its 6'-O-methyl ester (2) and oleanolic acid (1). Silica gel column chromatography of alkaline hydrolysate afforded a large amount of 4. MeOH extract and both EtOAc- and BuOH fractions were active in the rheumatoidal rat induced Freund's complete adjuvant reagent (FCA) whereas CHCl3 fraction was inactive. Compound 1 and 4 showed significant activities in the same assay but oleanolic acid 3-O-glucuronopyranoside (3) showed no activity. These fashions were also observed in carrageenan-induced edema of the rat and in the antinociceptive activity tests undertaken in hot plate- and writhing methods. These results suggest that momordin lc and its aglycone, oleanolic acid, could be active principles for rheumatoid arthritis. PMID- 12135109 TI - Protective effect of fangchinoline on cyanide-induced neurotoxicity in cultured rat cerebellar granule cells. AB - The present study was performed to examine the effect of fangchinoline, a bis- benzylisoquinoline alkaloid, which exhibits the characteristics of a Ca2+ channel blocker, on cyanide-induced neurotoxicity using cultured rat cerebellar granule neurons. NaCN produced a concentration-dependent reduction of cell viability, which was blocked by MK-801, an N-methyl-D-aspartate (NMDA) receptor antagonist, verapamil, L-type Ca2+ channel blocker, and L-NAME, a nitric oxide synthase inhibitor. Pretreatment with fangchinoline over a concentration range of 0.1 to 10 microM significantly decreased the NaCN-induced neuronal cell death, glutamate release into medium, and elevation of [Ca2+]i and oxidants generation. These results suggest that fangchinoline may mitigate the harmful effects of cyanide induced neuronal cell death by interfering with [Ca2+]i influx, due to its function as a Ca2+ channel blocker, and then by inhibiting glutamate release and oxidants generation. PMID- 12135108 TI - Carbon monoxide as a novel central pyrogenic mediator. AB - Carbon monoxide (CO) are produced by heme oxygenase (HO), and HO was detected in hypothalamus. However, the roles of CO produced in hypothalamus was not fully elucidated. So, we tested the effects of CO on body temperature because preoptic anterior hypothalamus was known as the presumptive primary fever-producing site. CO-saturated aCSF (4 microl, i.c.v.) and hemin (10 microg, i.c.v.) elicited marked febrile response. Pretreatment with indomethacin completely inhibited CO- and hemin-induced fever. Zinc protoporphyrin-IX (10 microg, i.c.v.) or ODQ (50 microg, i.c.v.) partially reduced hemin-induced febrile response. Dibutyryl-cGMP (100 microg, i.c.v.) produced profound febrile response and this febrile response was attenuated by indomethacin. These results indicate that endogenous CO may have a role as a pyrogenic mediator in CNS and CO-mediated pyresis is dependent on prostaglandin production and partially on activation of soluble guanylate cyclase. PMID- 12135110 TI - Efffect of recombinant human FSH on ovulation, pregnancy and in vitro fertilization in androgen-sterilized mice. AB - The effect of a new rhFSH, PG-0801, on oocyte quality, ovulation and in vitro fertilization (IVF) was examined in androgen-sterilized mice. Experimental sterility was induced by a single subcutaneous injection of testosterone propionate (TP, 1 mg/head) into 5 day old female mice. Ovulation was generated in the 10 to 13-week old TP-injected mice by a subcutaneous rhFSH injection (1, 5 or 10 IU/head) followed 48 hours later by a second rhFSH injection (1, 5 or 10 IU/head). For comparison, a subcutaneous PMSG (5 IU/head) injection was used for folliculogenesis and a hCG (5 IU/head) injection was used for ovulation. These were administered using the same protocol. The eggs were harvested from the oviducts and counted 17 to 20 hours after the second injection. IVF was performed by adding sperms (2 x 10(5)/ml to 2 x 106/ ml) to determine the functional activity of the eggs, and the fertilization rate was measured. In addition, the pregnancy rate and fetal development were examined after 15-17 days of gestation. The number of oocytes recovered from the rhFSH/rhFSH group increased dose dependently and was slightly higher than that of the PMSG/hCG group. The pregnancy rates of the group receiving 1, 5, and 10 IU of rhFSH/rhFSH were 50%, 66.7%, and 75%, respectively, which were significantly higher than that of the control (untreated) group (0%). The numbers of viable fetuses in the 1, 5, and 10 IU/head of the rhFSH/rhFSH group (8.0 +/- 1.50, 8.9 +/- 1.02, and 8.9 +/- 1.12 fetuses/dam, respectively) were comparable to that of the 5 IU/head PMSG/ hCG group (9.4 +/- 0.94). The mice receiving rhFSH/rhFSH and PMSG/hCG showed similar fertilization rates (around 65%) via the IVF procedure. These results demonstrate that a new rhFSH, PG-0801, may be useful for inducing ovulation in functionally infertile patients and for superovulation in ovulatory patients participating in assisted reproductive technology (ART) programs. PMID- 12135111 TI - Identification of hepatitis C virus core domain inducing suppression of allostimulatory capacity of dendritic cells. AB - Hepatitis C virus (HCV) is remarkably efficient at establishing chronic infection. One of the reasons for this appears to be the suppression of the accessory cell function of professional antigen presenting cells. In the present study, the immunosuppressive activity of HCV protein was examined on dendritic cells (DCs) generated from mouse bone marrow progenitor cells in vitro. We found that the DCs forced to express HCV protein have defective allostimulatory ability. DCs expressing HCV protein were phenotypically indistinguishable from normal DCs. However, they were unable to produce IL-12 effectively when stimulated with lipopolysaccharide. The functional domain of the HCV protein essential for immunosuppression was determined using a series of NH2-and C terminal deletion mutants of HCV core protein. We found that amino acid residues residing between the 21st and the 40th residues from the NH2-terminus of HCV core protein are required for immunosuppression. These findings suggest that HCV core protein suppresses the elicitation of protective Th1 responses by the inhibition of IL-12 production by DCs. PMID- 12135112 TI - Time courses of pCREB expression after dopaminergic stimulation by apomorphine in mouse brain. AB - Administration of dopamine agonist, apomorphine (2 mg/kg, s.c.), produces cage climbing behavior in mice that exhibit typical dopaminergic stimulation. The present study investigated the pCREB expression level in several brain regions following apomorphine treatment in order to determine whether the increased the dopaminergic activation produced by apomorphine accompanies the changes in pCREB immunoreactivity. A mouse brain was removed at 0 min, 10 min, 30 min, 1 h, 2 h, 7 h, and 24 h after apomorphine treatment. The brain tissue was fixed by an intracardiac perfusion with ice-cold 4% paraformaldehyde in PBS. Immunohistochemical study was conducted using the ABC-DAB method. The data showed that the immunoreactivity of pCREB increased in the striatum, nucleus-accumbens, piriform cortex and the dentate gyrus of the hippocampus of a mouse brain 30 min after the apomorphine treatment. Increased immunoreactivity began to diminish 2 h after the apomorphine treatment in all the brain regions measured. The time course for the pCREB immunoreactivity was similar to the behavioral response induced by the apomorphine treatment. These results suggest that activation of the dopamine receptor is accompanied by an increase in pCREB expression in the mouse brain. PMID- 12135113 TI - No role of protected region B of human cytochrome P4501A2 gene (CYP1A2) as an AP 1 response element. AB - Cytochrome P4501A2 (CYP1A2) is a member of the cytochrome P450 family of isozymes involved in the phase I drug metabolism of vertebrates. CYP1A2 is responsible for the activation of a number of aromatic amines to mutagenic and carcinogenic forms. Thus, the level of CYP1A2, which varies among different populations, may determine an individual's susceptibility to these chemicals. We have previously reported on the importance of a cis element named PRB (protected region B) in the regulation of human Cytochrome P4501A2 (CYP1A2) gene, which appeared to act as a positive regulatory element. Closer examination of the PRB sequence (-2218 to 2187 bp) revealed a putative AP-1 binding site, TGACTAA, at -2212 bp (Chung and Bresnick, 1997). To elucidate the role of AP-1 in CYP1A2 regulation, we transiently overexpressed c-Jun and c-Fos transcription factors in human hepatoma HepG2 cells, and examined their influence on the CYP1A2 promoter activity by reporter gene assays. Cotransfection of the c-Jun and the c-Fos expression vectors increased the induced transactivation by five to six fold from the CYP1A2 promoter constructs. However, deletion of the PRB element did not affect the degree of activation by the c-Jun and the c-Fos. Therefore, it is unlikely that the c-Jun and the c-Fos activate the CYP1A2 promoter through this AP-1 consensus like sequence in the PRB region. PMID- 12135114 TI - Relation of dynamic changes in interfacial tension to protein destabilization upon emulsification. AB - The objective of this study was to link conformational changes of proteins at a water/methylene chloride interface to their destabilization upon emulsification. When 4 aqueous protein solutions (bovine serum albumin, beta-lactoglobulin, ovalbumin, or ribonuclease) were emulsified in methylene chloride, considerable proportions of all the proteins became water insoluble aggregates. There were also noticeable changes in the compositions of their water-soluble species. A series of water/methylene chloride interfacial reactions upon the proteins was considered a major cause of the phenomena observed. Based on this supposition, the interfacial tension was determined by a Kruss DVT-10 drop volume tensiometer under various experimental conditions. It substantiated that the interfacial tension was high enough to cause the adsorption of all the proteins. Under our experimental conditions, their presence in the aqueous phase resulted in reductions of the interfacial tension by the degrees of 8.5 - 17.1 mN m(-1). In addition, dynamic changes in the interfacial tension were monitored to compare relative rates at which the adsorbed proteins underwent conformational, structural rearrangements at the interface. Such information helped make a prediction about how easily proteins would denature and aggregate during emulsification. Our study indicated that emulsifying aqueous protein solutions in organic solvents should be handled with care, due to adverse interfacial effects. PMID- 12135115 TI - Interspecies comparison of the oral absorption of itraconazole in laboratory animals. AB - The oral absorption and disposition of itraconazole were studied in rats, rabbits and dogs. Serum levels of itraconazole and its active metabolite, hydroxyitraconazole, were determined by a validated HPLC method. The absorption of itraconazole was relatively rapid in rats and dogs but was slower in rabbits. The terminal elimination half-life (T 1/2lambda(z)), time to the peak concentration (Tmax), dose and weight normalized area under the curve (AUC) and the peak concentration (Cmax) of itraconazole found in the dog were comparable to those reported in humans. As in humans, the metabolite to parent drug AUC ratios in rats and dogs were greater than unity but was less in rabbits. The dog appears to be an appropriate animal model while the rat, not the rabbit, may be used as an alternative animal model in predicting the oral absorption of itraconazole in humans. PMID- 12135116 TI - Effect of vehicles and enhancers on the in vitro permeation of melatonin through hairless mouse skin. AB - The effects of vehicles and penetration enhancers on the in vitro permeation of melatonin through dorsal hairless mouse skin were investigated. Propylene glycol laurate (PGL), isopropyl myristate (IPM), propylene glycol monolaurate (PGML) and propylene glycol monocaprylate (PGMC) showed high permeation fluxes and PGL, PGML and PGMC decreased lag time significantly. In both of the binary co-solvents of diethylene glycol monoethyl ether (DGME)-PGL and DGME-IPM, the highest fluxes were achieved at 20% of DGME, which were 10.5 +/- 1.5 and 9.1 +/- 2.4 microg/cm2/h, respectively. Among fatty acids used as a permeation enhancer, capric acid and oleic acid in DGME-PGL (80:20 v/v) showed relatively high enhancing effects. Capric acid also shortened the lag time of melatonin from 2.4 +/- 0.7 to 1.3 +/- 0.2 h. Oleic acid, however, failed to shorten the lag time. Therefore, for effective solution formulations in terms of permeation flux and lag time, capric acid-containing DGME-PGL (80:20 v/v) could be used to enhance the skin permeation of melatonin. PMID- 12135117 TI - In response to the article by Thomas Wittlinger and co-workers: Magnetic resonance imaging of coronary artery occlusions in the navigator technique. PMID- 12135118 TI - Imaging comparison of basic cardiac views between two- and three-dimensional ultrasound in normal fetuses in anterior spine positions. AB - OBJECTIVE: When the fetal spine is in anterior position, it shadows the fetal heart, resulting in the difficult visualization using two-dimensional (2D) ultrasound. The purpose of this study was to compare the basic cardiac views of normal fetuses between 2D and 3D ultrasound to demonstrate whether 3D ultrasound improved the visualization of these views in fetuses in anterior spine positions. In addition, inter- and intra-observation reliabilities of basic cardiac views using 3D ultrasound were evaluated for their clinical applicability. METHODS: Using a multiplanar technique, integrated 3D ultrasound was used to display the four-chamber view, aortic outflow tract and pulmonary outflow tract in fetuses in anterior spine positions for 23 uncomplicated singleton pregnant women. The imaging visualizations of these views for the 23 fetuses in 3D ultrasound were compared with those in 2D ultrasound using the McNemar test. We also evaluated the inter- and intra-observation differences of each basic cardiac view in 3D ultrasound using the kappa statistic and McNemar test, respectively. RESULTS: Only in the pulmonary outflow tract, 3D ultrasound had significantly better visualization than the 2D ultrasound in the fetuses in anterior spine positions (p < 0.05). There was good inter-observation reliability and no intra-observation differences for the technique were observed. CONCLUSIONS: Among the basic cardiac views in fetuses in anterior spine positions, 3D ultrasound improved the visualization of pulmonary outflow and provided reliable alternate technique for clinical use. PMID- 12135119 TI - A congenital complex including muscular interventricular septal aneurysm in an adult: case report and review of literature. AB - Muscular interventricular septum is an extremely rare location for congenital aneurysm. A 48-year-old patient with a muscular interventricular septum aneurysm coexisting with atrial septal defect, pulmonary stenosis and ventricular preexcitation is presented. Three-dimensional echocardiographic reconstruction of the lesion aided in a precise morphologic and quantitative assessment of the condition. The review of 10 previously reported patients with the entity is presented. PMID- 12135120 TI - High prevalence of (99m)tc-tetrofosmin reverse perfusion pattern in patients with myocardial infarction and angiographically smooth coronary arteries. AB - BACKGROUND: There are no published data in the literature on the scintigraphic perfusion pattern in patients with myocardial infarction (MI) and normal coronary arteries (NCA). OBJECTIVES: To evaluate myocardial perfusion imaging in a series of patients with MI and NCA. METHODS: Twenty-seven patients who had developed a MI and had NCA were studied. As a control group we included 27 patients with a recent MI and coronary artery disease (CAD). All patients underwent stress/rest tetrofosmin myocardial perfusion SPECT within 6 months from MI. RESULTS: In patients with NCA tetrofosmin stress images revealed 41 hypoperfused segments in 17 patients (63%). On rest images, 13 segments remained unchanged, 4 showed partial reperfusion, 10 normalized and 14 worsened. Additionally, there were 18 new hypoperfused segments in nine patients. Therefore, perfusion worsened at rest in 18 patients (67%) (32 segments). Overall, at rest there were 49 hypoperfused segments in 22 patients (81%). In patients with CAD, stress images revealed 71 hypoperfused segments. On rest images, 39 segments remained unchanged, 16 showed partial reperfusion and 12 normalized. Four segments worsened at rest and only four patients (15%) showed new perfusion defects at rest. CONCLUSIONS: Myocardial perfusion with tetrofosmin might appear considerably worse at rest than at stress in patients with MI and NCA. Specifically, a reverse perfusion pattern in the infarct area is a frequent finding and is likely to be due to residual tissue viability. We postulate that in these patients the hyperemic response to exercise may mask resting underperfusion areas. PMID- 12135121 TI - Quantitative assessment of the pressure and volume overloaded right ventricle: imaging is a real challenge. PMID- 12135122 TI - Adequate patient selection for coronary revascularization: an overview of current methods used in daily clinical practice. AB - Widely used non-invasive stress modalities, like exercise ECG, MPS and stress echocardiography, are the tests of first choice for the diagnosis of CAD. It has been shown in numerous studies that non-invasive assessment of perfusion abnormalities is an adequate strategy for risk stratification. Moreover, non invasive stress testing should be performed before a diagnostic cardiac catheterization to document the presence of myocardial ischemia, as a prerequisite for coronary revascularization. Coronary angiography is the gold standard for identifying CAD; however this technique is limited in assessing functional severity of coronary narrowings ('illusion of luminology'; see also Figure 5). The recently introduced i.c. hemodynamic parameters (CFVR and FFR) can identify functional severity of specific lesions and have shown a good agreement with the results of non-invasive stress test in validation studies. Furthermore, there is accumulating evidence that it is safe to defer a PTCA procedure, based on normal FFR and CFVR values. As these indices are derived during an invasive cardiac catheterization procedure, its use is recommended during a so called 'ad hoc' PTCA setting. Furthermore, they are particularly useful for clinical decision making in patients with documented multivessel CAD, as both indices allow selective evaluation of coronary narrowings in different arteries. Revascularization procedures are costly and always have a potential risk. It is important to be aware that, using above mentioned methods, unnecessary interventions (lacking potential benefit) may be avoided. PMID- 12135123 TI - Quantification of shunt volumes in congenital heart diseases using a breath-hold MR phase contrast technique--comparison with oximetry. AB - AIMS: Comparison of breath-hold MR phase contrast technique in the estimation of cardiac shunt volumes with the invasive oximetric technique. METHODS AND RESULTS: Seventeen patients with various cardiac shunts (10 ASD, 3 VSD, 1 PDA, 3 PFO) and five healthy volunteers were investigated using a 1.5 Tesla system. The mean flow velocity, the mean volume flow and the transverse area in the ascending aorta and the left and right pulmonary artery were measured using the MR phase contrast breath-hold technique (through plane, FLASH 2D-sequence, TR/TE 11/5 ms, phase length 106 ms, VENC 250 cm/s). The ratio of mean flow in the pulmonary (Qp: sum of mean flows in the left and right pulmonary arteries) and the systemic circulation (Qs: mean flow in the ascending aorta) was calculated and compared with invasively measured Qp:Qs ratios. Oximetry was performed within 24 h of the MR investigation. The non-invasive shunt measurement in the 17 patients showed a mean Qp:Qs ratio of 2.00 +/- 0.86. Comparing the MR data with the invasively measured Qp:Qs showed a correlation coefficient of r = 0.91 (p < 0.001). CONCLUSION: Cardiac shunt volumes can be measured reliably using a shorter acquisition time with breath-hold MR phase contrast technique. PMID- 12135124 TI - Standardized myocardial segmentation and nomenclature for tomographic imaging of the heart. A statement for healthcare professionals from the Cardiac Imaging Committee of the Council on Clinical Cardiology of the American Heart Association. PMID- 12135125 TI - Aorto-mitral valvular evaluation with MRI using the left parasternal long axis (angled-vertical long axis) plane. PMID- 12135127 TI - Pulmonary edema during pregnancy: unilateral presentation is not rare. AB - Information regarding pulmonary edema in obstetric patients is limited, especially its natural history as most cases are from tertiary care centers. The incidence, etiology, and course of pulmonary edema in all obstetric patients at a primary-secondary care center was studied prospectively among 29,621 obstetric cases in the past 3.5 years. Pulmonary edema developed in 18 cases (0.06%) of all obstetric patients that were associated with pre-eclampsia-eclampsia and tocolysis with beta-mimetics. The onset of pulmonary edema was 1-8 days postpartum in 12 patients, 1 day antepartum in 3 patients, and 26-32 gestational weeks in 3 patients. The cause was mainly cardiogenic, and the mean time taken for resolution was 2.2 days. Unilateral pulmonary edema occurred in 4 patients and there was delayed resolution compared with bilateral pulmonary edema because of delayed treatment. Pulmonary edema associated with pregnancy developed in 0.06% of cases, mainly during the peripartum, and resolved rapidly. Unilateral pulmonary edema was not uncommon. PMID- 12135126 TI - Congenital absence of pericardium revisited. AB - Although much have been published regarding congenital absence of pericardium, it is essential that this anomaly, like an old friend, be revisited from time to time. Review of this anomaly with emphasis on its embryological process is discussed. Furthermore, with the advances in magnetic resonance imaging, absence of pericardium can now be diagnosed with ease and the radiological findings of this condition are reviewed as well. PMID- 12135128 TI - Increased nitric oxide in proportion to the severity of heart failure in patients with dilated cardiomyopathy: close correlation of tumor necrosis factor-alpha with systemic and local production of nitric oxide. AB - Recent studies have demonstrated that proinflammatory cytokines induce large amounts of nitric oxide (NO) and that the amount increases in patients with congestive heart failure (CHF). There are, however, few reports regarding the relationships between NO production, cytokines and the severity of heart failure, so the plasma concentrations of nitrite and nitrate (NOx), tumor necrosis factor alpha (TNF-alpha) and brain natriuretic peptide (BNP) were measured in 43 patients with CHF caused by dilated cardiomyopathy and 26 age- and sex-matched normal control subjects. Forearm blood flow (FBF) was measured using plethysmography during infusions of acetylcholine and nitroglycerin and after the administration of the NO synthesis inhibitor L-NMMA (N(G)-monomethyl-L-arginine). Plasma concentrations of both NOx and TNF-alpha were significantly higher in the patient group than in the control group (p<0.001) and correlated closely with BNP concentrations (p<0.001). There was a positive relationship between NOx and TNF alpha concentrations (r=0.80, p<0.001). Administration of L-NMMA significantly reduced FBF in both groups, and the percent change in FBF from baseline correlated significantly with TNF-alpha concentrations (r=0.63, p<0.001). The FBF response to acetylcholine was depressed in the patient group and correlated inversely with TNF-alpha concentrations. The FBF response to nitroglycerin did not correlate with TNF-alpha concentrations. The findings indicate that the concentrations of NO and TNF-alpha in patients with CHF increase in proportion to the severity of heart failure, and that TNF-alpha plays a role in the enhanced systemic and local production of NO. PMID- 12135129 TI - Shunt between the ventricular chamber and coronary arteries preserves left ventricular function in acute myocardial infarction. AB - It is controversial whether newly created channels made by transmyocardial laser revascularization are functionally significant, so the present study evaluated the shunt flow from the left ventricular (LV) cavity to the ischemic myocardium in 51 patients with acute myocardial infarction (AMI) caused by complete occlusion of the proximal left anterior descending coronary artery. All patients underwent left heart catheterization within 24 h of onset and all underwent successful coronary reperfusion using primary coronary angioplasty with no angiographic restenosis on follow-up coronary angiography (CAG). The presence of the LV shunt flow was evaluated by selective left CAG after successful reperfusion. The LV global ejection fraction (EF) and regional function (centerline method) were analyzed by ventriculography in both the acute and chronic phases. The patients were divided into the 3 groups (Group A, no LV shunt without collaterals, n=20; Group B, no LV shunt with collaterals, n=24; Group C, LV shunt with collaterals, n=7). There was no difference in the grade of collateral circulation between Groups B and C. The improvements in LVEF and regional function from the acute phase to the chronic phase were significantly greater in Group C than in Groups A and B. Not only collateral circulation but also LV shunt contributes to the functional recovery of infarct myocardium in patients with AMI. PMID- 12135130 TI - Circulating levels of insulin-like growth factor-1 and its binding proteins in patients with hypertrophic cardiomyopathy. AB - Insulin-like growth factor-1 (IGF-1) is important in the hypertrophic response of the myocardium, so the present study was designed to elucidate whether the circulating levels of IGF-1 and its binding proteins (IGFBPs) are related to the disease condition of patients with hypertrophic cardiomyopathy (HCM), in particular the occurrence of congestive heart failure (CHF). The study group comprised 124 patients with HCM and 15 healthy control subjects. The HCM patients were subdivided into 3 groups: 39 with hypertrophic obstructive cardiomyopathy (HOCM), 67 with hypertrophic non-obstructive cardiomyopathy (HNCM), and 18 with HCM and a history of CHF (HF-HCM, n=18). Serum levels of IGF-1 and IGFBPs (IGFBP 1 and -3) were compared between groups. IGF-1 levels were significantly higher in patients with HOCM and HNCM, and lower in patients with HF-HCM than in control subjects (p<0.0001, p<0.005, and p<0.05, respectively). IGFBP-1 levels were significantly higher in patients with HF-HCM than in the other 3 groups (p<0.0001 for all). The findings suggest that circulating levels of IGF-1 and IGFBP-1 are related to the extent of myocardial injury in patients with HCM. PMID- 12135132 TI - Electrocardiography subtraction method of detecting low-amplitude, high-frequency components (L-HFCs) within the QRS complex of patients with ventricular tachycardia. AB - A new method of extracting the low-amplitude and high-frequency components (L HFCs) was developed and this study investigated its usefulness in 86 subjects: 28 normal subjects (group 1) and 58 patients with a previous myocardial infarction (MI). The patients were classified into 3 groups: group 2 with 38 patients without ventricular tachycardia (VT), group 3 with 13 patients with non-sustained VT, and group 4 with 7 patients with sustained VT. The new electrocardiography (ECG) subtraction method, using a mathematical filtering procedure, was used instead of conventional complex filtering. The continuous L-HFCs within the QRS complex were analyzed using a Z-lead recording. The duration of the continuous L HFCs was significantly longer in group 4 than in groups 1 (p<0.0001), 2 (p<0.0001) or 3 (p<0.05) with all 3 filters. The ECG subtraction method is a powerful and useful new technique for identifying patients at risk for either sustained or non-sustained VT after MI, and overcomes several of the problems with the conventional signal-averaging method. PMID- 12135131 TI - Effects of right stellate ganglion block on the autonomic nervous function of the heart: a study using the head-up tilt test. AB - The effect of peripheral sympathetic block on the autonomic nerve function of the heart was studied using the head-up tilt test (HUTT) and right stellate ganglion block (RSGB). Blood pressure (BP), heart rate (HR) and the parameters of power spectral analysis of HR variability recorded during the HUTT were measured in 8 patients with chronic pain syndrome before and after RSGB. In the control state, the mean HR and the LF/HF component recorded during HUTT significantly increased whereas the HF component markedly decreased. Conversely, the mean HR and LF/HF and HF components during HUTT did not significantly alter after the RSGB procedure. There were no significant differences between the BP values before and after RSGB. These results suggest that RSGB suppresses cardiac sympathetic function without significantly affecting BP and thus may be a safe and effective therapy for the chronic pain syndrome. PMID- 12135133 TI - Combination therapy of exercise and angiotensin-converting enzyme inhibitor markedly improves insulin sensitivities in hypertensive patients with insulin resistance. AB - The contraction of muscle enhances the release of bradykinin (BK) and improves glucose uptake by the muscle. Angiotensin-converting enzyme inhibitor (ACEI) slows the breakdown of BK, thus the effect of BK is augmented in the presence of ACEI. The present study investigated whether the combination of exercise (increased production of BK) and ACEI (delay in breakdown of BK) might further improve insulin sensitivity in hypertensive patients with insulin resistance (HOMA-R>1.8). Patients were assigned either to increased walking distance (Walking group) or taking 2 mg temocapril, an ACEI, daily (ACEI group) for 8 weeks. Then both interventions were given to all patients for 8 weeks (ACEI+Walking group). Blood concentrations of triglycerides were slightly lower in the ACEI+Walking group than at baseline, although there were no significant differences in total cholesterol or high density lipoprotein-cholesterol among the 2 groups. Blood glucose was not significantly different with each treatment, but blood concentrations of insulin and HOMA-R were significantly lower in the Walking and ACEI groups compared with the Control group. The combination of walking and ACEI further lowered blood concentrations of insulin and HOMA-R, which suggests that this treatment is beneficial for hypertensive patients with insulin resistance. PMID- 12135135 TI - Doppler echocardiographic differentiation of functional from anatomical pulmonary atresia: analysis using quantitative parameters. AB - Functional pulmonary atresia must be distinguished from anatomical atresia, which has an intact ventricular septum, to avoid inappropriate treatment, but there is a paucity of data regarding the echocardiographic features that differentiate these conditions. Echocardiographic findings in 5 neonates with functional atresia were compared to those in 5 with anatomical atresia. The left and right ventricular end-diastolic dimensions (LVDd, RVDd), percent of normal predicted LVDd, RVDd/LVDd, tricuspid valve ring diameter (TVD), percent of normal predicted TVD, grade of tricuspid regurgitation (TR), peak TR velocity, pulmonary valve ring diameter (PVD), percent of normal predicted PVD, the minimum diameter of the ductus and the peak velocity through it (PDA velocity) were measured. In addition, systolic pulmonary (PAp) and right ventricular pressure (RVp) from either PDA velocity or TR velocity, and calculated PAp/RVp were also estimated. There were significant differences in RVDd/LVDd, %TVD, and peak TR velocity between the 2 groups. All functional patients showed RVDd/LVDd >0.6, %TVD >100%, estimated RVp <50mmHg, PAp/RVp >0.85, and peak TR velocity <4m/s, whereas the findings in anatomical atresia patients were completely the opposite. In conclusion, a large RVDd/LVDd, TVD, PAp/RVp, low RVp and small TR velocity all suggest functional rather than anatomical pulmonary atresia, although there may be some exceptions such as severe Ebstein anomaly. PMID- 12135134 TI - Guidewire bias in rotational atherectomy in the angled lesion: evaluation based on the thickness of the ablated intima and media. AB - The effect of guidewire bias on angled-lesion ablation by rotational atherectomy (RA) was assessed by measuring the changes in vertical lumen diameter, horizontal lumen diameter and the intima-media thickness of the coronary artery, using intravascular ultrasound in 10 lesions with an angle greater than 10 degrees. The vertical and horizontal diameters significantly increased after RA. The intima media thickness at the 4 orthogonal sites significantly decreased. There was a significant positive correlation between vertical diameter change and angle (r=0.642, p=0.045), but none between horizontal diameter change and angle. There was no correlation between intima-media thickness change at 0 degrees and angle; however, at 180 degrees there was a tendency to correlation with angle (r=0.602, p=0.066). These data suggest that in cases of angled lesions, the increase in vertical lumen diameter is caused more by ablation of the 180 degrees wall than by that of the 0 degrees wall, which is brought about by guidewire bias toward the vascular wall at 180 degrees. PMID- 12135137 TI - Delayed thrombogenesis following radiofrequency catheter ablation. AB - The cause and duration of the thrombogenesis provoked by radiofrequency catheter ablation (RF-CA) was investigated by measuring the thrombin-antithrombin III complex (TAT) in 43 patients who underwent RF-CA and in 20 control subjects who underwent an electrophysiologic study. Blood samples were collected at 7 different times: before introducing the sheaths, during the ablation procedure and at 30 min, 6 and 24h, and 3 and 6 days after the procedure. Hepatocyte growth factor (HGF) was simultaneously measured in the ablation group. Plasma TAT concentration exhibited a double peaked pattern in the ablation group: the first peak occurred during the ablation procedure (42.8+/-15.5 ng/ml), and the second peak 3 days later. Plasma TAT at 3 days after the procedure was significantly higher than that of the control group (21.3+/-19.0 vs 2.5+/-1.4, p=0.0003). The first peak significantly correlated with the procedure time prior to the administration of heparin (r=0.669), but the second peak did not (r=0.132). A subgroup with a serum HGF >0.40 ng/ml at 6 h after the procedure exhibited a significantly high second peak. The thrombogenesis caused by RF-CA has 2 phases; in the acute phase, there is hemostasis during placement of the catheters, and in the delayed phase thrombogenesis is the result of endothelial damage from the RF current. PMID- 12135136 TI - Anticoagulant therapy in Japanese patients with mechanical mitral valves. AB - There are no guidelines for the optimal therapeutic range of anticoagulant therapy in Japanese patients with mechanical heart valves. A total of 214 patients were followed retrospectively after mitral mechanical valve replacement (mean duration of follow-up, 4.8 years; total duration of follow-up, 1,027 patient-years). The target range of the international normalized ratio (INR) for oral anticoagulation was between 1.5 and 2.5. For all patients 10,416 measurements of the INR were obtained during the follow-up period and approximately 76% of the intensity measurements were within the target range. Thromboembolism occurred in 8 patients (0.8 per 100 patient-years) and major bleeding in 5 patients (0.5 per 100 patient-years). There was no correlation between the distribution of the INR and the occurrence of thromboembolic or bleeding complications. In the univariate analysis of the various risk factors, patients who had a tilting valve or did not receive antiplatelet therapy had an increased risk of thromboembolism. However, there were no risk factors with respect to bleeding complications. A target range of 1.5 to 2.5 INR appears to be the optimal range and is safe for thromboembolism or bleeding complications. Thromboembolism may be reduced by additional antiplatelet therapy, and a tilting valve needs more intense anticoagulation. PMID- 12135138 TI - Higher C-reactive protein concentration and white blood cell count in subjects with more coronary risk factors and/or lower physical fitness among apparently healthy Japanese. AB - Of 2,722 people (1.876 men, mean age: 51.3+/-10.3 years; 846 women, mean age: 51.4+/-11.1) who underwent the fitness check program at Aichi Prefectural Center for Health Care, the concentration of C-reactive protein (CRP) and the white blood cell count (WBC) were investigated in relation to the number of coronary risk factors, maximum oxygen uptake (VO2max) quartiles and physical fitness score. CRP was measured by conventional latex immunoturbidimetric assay. Both CRP and WBC were higher in those who had more risk factors. In men the lowest mean CRP was 0.07 mg/dl in those with only one risk factor (RF1) and the highest was 0.29 mg/dl in RF6 (p<0.0001). The lowest mean WBC was 4,868/mm3 in RF1 and the highest was 7,096/mm3 in RF6 (p<0.0001). In women the lowest mean CRP was 0.073 mg/dl in those with no risk factors (RF0) and the highest was 0.22mg/dl in RF5 (p=0.0379). The lowest mean WBC was 5,065/mm3 in RF1, and the highest was 6,792/mm3 in RF4 (p=0.0001). A similar relationship was noticed when the groups were analyzed by VO2max quartile or fitness score. CRP and WBC both showed a stepwise increase or decrease in men, but was generally in order in women in accordance with the number of risk factors, VO2max level or fitness score. In apparently healthy Japanese subjects, elevated inflammatory indices (ie, higher CRP and WBC) were associated with more coronary risk factors and poorer physical fitness. Therefore, high-risk coronary subjects might be screened by conventional measurement of CRP. PMID- 12135139 TI - Optimal time for predicting left ventricular remodeling after successful primary coronary angioplasty in acute myocardial infarction using serial myocardial contrast echocardiography and magnetic resonance imaging. AB - The objective of this study was to determine the optimal time to assess microvascular integrity within the risk area for myocardial infarction in order to predict unfavorable left ventricular remodeling (LVR) after successful primary coronary angioplasty. Fifty-three patients who underwent myocardial contrast echocardiography (MCE) just before recanalization, shortly after and 1 day (Day 2) and 3 weeks after recanalization were studied. The no- and low-reflow ratio (LR ratio) was analyzed at each stage. The wall-thinning ratio within the risk area was determined using magnetic resonance imaging performed 3-4 weeks after the recanalization. Thirteen of the 53 patients showed LVR 3-8 months after recanalization. The optimal time to predict LVR was found to be Day 2 based on the receiver operating characteristic curves. The LR ratio on Day 2 (chi2=7.39, p=0.007) and the collateral circulation before recanalization (chi2=4.57, p=0.03) were chosen as independent variables for predicting LVR. Patients with greater than 0.43 in the LR ratio on Day 2 showed a lower wall-thinning ratio (58+/-19% vs 72+/-20%, p=0.05). This study shows that the optimal time to estimate the microvascular integrity for predicting LVR is 1 day after recanalization, which is neither shortly after recanalization nor during the convalescent stage. PMID- 12135140 TI - Clinical characteristics and follow-up in patients with microvascular angina. AB - Arteriosclerosis of the small arteries is one of the main causes of microvascular angina, and although some reports have shown favorable prognoses, there is progressive reduction in left ventricular function. The present study evaluated the prognosis of microvascular angina in 86 Japanese patients (51 women, 35 men; average age, 59+/-9 years) who had ischemic ST segment depression, normal coronary angiograms and small artery sclerosis confirmed by endomyocardial biopsies. The mean follow-up period was 7.2+/-3.4 years. Questionnaires regarding their symptoms, cardiac medication, and new events were sent to all patients. Eighty-five patients (98.9%) were still alive at the end of the follow-up period. Chest pain remained in 35.3%; the degree of pain was unchanged in 18.8%, and had lessened in 11.8%. None of the patients died of cardiac events or suffered from a myocardial infarction. At the end of the follow-up period, calcium antagonist was used in 63.5% of patients. Seventeen patients (20.0%) were free of antianginal medication. The prognosis of microvascular angina diagnosed by strict criteria was favorable. PMID- 12135141 TI - Pharmacological and electrophysiological characterization of junctional rhythm during radiofrequency catheter ablation of the atrioventricular node: possible involvement of neurotransmitters from autonomic nervous system. AB - Catheter ablation of the atrioventricular node (AVN) with radiofrequency current is closely associated with the short-term onset of a junctional rhythm. The origin of this rhythm was analyzed in Beagle dogs which were anesthetized with pentobarbital sodium. Atrioventricular (AV) conduction block was induced first using a standard catheter ablation technique for the AVN, so that the sinus automaticity could not override the junctional ectopy during the following energy delivery. The ablation catheter was kept in the initial position and the delivery of radiofrequency energy was repeated. The pattern of ECG changes suggests that the dominant pacemaker may shift from the distal portion of the AV junctional area to the proximal portion during the energy delivery. This enhanced junctional automaticity was suppressed by the beta-blocker esmolol, but was not affected by M-antagonist atropine. Moreover, the beta-agonist isoproterenol did not induce the same type of junctional tachycardia, but the pacemaker shift was induced by the increased sympathetic tone after transient asystole by ventricular overdrive pacing or acetylcholine administration. These results suggest that proximal portion of the AV junctional area has extremely slow pacemaker activity, but responds to locally released norepinephrine with an abrupt rise and fall in rate, resulting in a typical pattern of junctional tachycardia during the ablation of the AVN. PMID- 12135142 TI - No-reflow following dilatation of a coronary lesion with a large lipid core. AB - Radiolucent findings of coronary angiogram are believed to usually represent intracoronary thrombus, but in the present case, were atheromatous plaque with a large lipid core. A 62-year-old man who suffered from an inferior acute myocardial infarction was admitted to hospital 6 h after onset. The first cine angiograms showed TIMI-1 flow in the distal-portion of the right coronary artery, so thrombectomy was initially carried out and TIMI-2 flow achieved. However, the radiolucent lesion did not disappear and so adjunctive mechanical dilatation of the lesion was prformed, which resulted in 'no-reflow' (TIMI-0). Finally, aspiration of the material from the stagnated lesion was attempted and immediately obtained TIMI-3 flow. The retrieved materials were macrophages (foam cells) and many cholesterol crystals, both of which are considered to be atheromatous gruel. Therefore, the sudden flow reduction following percutaneous transluminal coronary angioplasty was caused by mechanical disruption of an atheromatous plaque with a large lipid core. PMID- 12135143 TI - Long-term usefulness of percutaneous intrapericardial fibrin-glue fixation therapy for oozing type of left ventricular free wall rupture: a case report. AB - This report describes a long-term survival case of left ventricular free wall rupture treated with percutaneous intrapericardial fibrin-glue fixation therapy. A 82-year-old woman was admitted to the emergency room because of vomiting and syncope diagnosed as acute posterolateral myocardial infarction complicated by cardiac tamponade. After her hemodynamic condition was stabilized by drawing off the bloody pericardial effusion, fibrin-glue was injected into pericardial space with the expectation that the glue would cover the oozing site of the left ventricular epicardium. After this therapy, the patient recovered and did not have any no recurrent cardiac events for 1 year. Serial echocardiographic studies revealed a preserved left ventricular function and no development of left ventricular restriction. This case suggests that percutaneous intrapericardial fibrin-glue fixation therapy is an effective treatment for the oozing type of left ventricular free wall rupture and that there is no risk of left ventricular restriction during long-term follow-up. PMID- 12135144 TI - An unusual case of birdshot embolism. AB - There are numerous cases of arterial and venous bullet embolism to the heart. An unusual case of birdshot embolus to the right ventricle from the femoral vein caused diagnostic confusion. Distant migration of the foreign bodies via blood vessels has to be taken into consideration after gunshot wounds. PMID- 12135145 TI - Effective disopyramide treatment in a boy with mid-ventricular hypertrophic obstructive cardiomyopathy. AB - A 14-year-old boy with mid-ventricular hypertrophic obstructive cardiomyopathy (MVHOCM) first presented at the age of 10 years with severe chest pain. Two dimensional echocardiography disclosed marked hypertrophy at the mid-portion of the ventricular septum, and left ventriculography showed an hourglass appearance at systole. He was initially treated with propranolol, but the chest pain and dyspnea on exertion worsened at the age of 12 years. After disopyramide was started, the chest pain disappeared and the degree of the pressure gradient at the mid-ventricular level was reduced. There was also significant improvement on a 123I beta-methyliodophenyl pentadecaonic acid (BMIPP) myocardial scintigram. PMID- 12135146 TI - Emotional stress induces transient left ventricular hypocontraction in the rat via activation of cardiac adrenoceptors: a possible animal model of 'tako-tsubo' cardiomyopathy. AB - The etiology of a novel cardiac syndrome 'tako-tsubo' cardiomyopathy, otherwise known as 'transient left ventricular (LV) apical ballooning' and which mimics acute myocardial infarction, is not clear; however, emotional or physical stress is known to precede the attack. Left ventriculography of rats experiencing emotional stress induced reversible LV apical ballooning, which was normalized by pretreatment with adrenoceptor blockade. Together with results of previous studies, activation of cardiac adrenoceptors in the absence of ischemia reperfusion is proposed as the primary cause of this syndrome. PMID- 12135147 TI - New policy ideas for rural health care delivery: presentations before a congressional committee. PMID- 12135148 TI - Challenges in evaluating rural health programs. AB - Complex community-based prevention programs are being held to scientific evidence of their effectiveness and rural public health departments that implement such programs often are not equipped to evaluate them. Rural public health departments are fettered by small budgets, small staffs, and less access to evaluation experts and similar resources. Community-based health promotion programs can include complex designs that may work differently in rural areas and evaluation of rural programs can be hampered by lack of control groups and the instability of results from small populations. The University of Kentucky has entered into a contract with the state Department for Public Health to implement an internal, participatory model of evaluation. In this model, the university evaluation expert trains local public health department staff in technical skills for program evaluation and acts as mentor and technical consultant to local public health departments on an ongoing basis. Through training and site visits, this model is one approach to addressing the challenges of evaluating rural health promotion programs. PMID- 12135149 TI - Health risk factors among Iowa farmers. AB - This study compares the prevalence of the health risk factors for chronic disease and injury among farmers with their prevalence among other workers, using data from the Iowa Behavioral Risk Factor Surveillance System. From January 1999 to December 1999, there were 3,620 adults who participated in the survey. This report focuses on the 2,140 subjects who reported they were working. We calculated the percentage with each health risk factor, the adjusted odds ratio (OR) and the associated 95% confidence interval (CI), comparing the prevalence of health risks among farmers and other workers. Most health conditions and risk factors were similar among farmers and other workers. However, we found that farmers had some protective behaviors. They had better oral health (OR = 0.34, 95% CI 0.20-0.57) and were less likely to smoke (OR = 0.14, 95% CI 0.06-0.29) than other workers. In contrast, we found that farmers reported some high-risk behaviors compared to other workers. They were less likely to have smoke detectors at home (OR = 0.39, 95% CI 0.18-0.82), or to have had stool blood screening for colorectal cancer (OR = 0.41, 95% CI 0.1 7-1.00). These results suggest areas for future research to define targeted preventive interventions in farm communities. PMID- 12135150 TI - Primary care practice management in rural and urban Veterans Health Administration settings. AB - Limited access to specialty care in rural settings may result in more expectations of primary care providers and a higher demand for primary care. The authors used survey and administrative data from 1999 from the Veterans Health Administration (VHA) to compare primary care practice management and performance in 19 rural to 103 urban VHA hospitals nationally. Rural VHA hospitals were smaller, less likely to be academically affiliated, and had fewer integrated specialty care services. Primary care providers in rural settings were more likely to manage specialty care services, provide continuity across patient care settings, and have complete responsibility for a broader range of services. However, rural hospitals had more staff per patient allocated to primary care than did urban hospitals. Patients in rural settings received comparable quality care to those in urban settings, and they appeared to be more satisfied with the care they received. Within the VHA system, primary care providers in rural settings provided a broader range of services than those in urban ones. This increased breadth may be attributable to the lack of availability of integrated specialty care services in rural settings. Because of this broader range of responsibilities, the provision of primary care in rural settings may require higher staffing patterns and may be inherently more costly than in urban settings; therefore, researchers should be cautious when comparing primary care expenditures across rural and urban settings. PMID- 12135151 TI - The importance of understanding human misuse of veterinary medications. PMID- 12135152 TI - Veterinarian perception of the intentional misuse of veterinary medications in humans: a preliminary survey of Idaho-licensed practitioners. AB - The intentional administration of veterinary medications to humans is a form of medication misuse that has not been systematically studied. Veterinarians are the health practitioner group most likely to have knowledge about this problem and to be approached by the public for advice. For this preliminary study, questionnaires were mailed to 1,077 veterinarians registered with the Idaho Board of Veterinary Medicine regarding their knowledge and perceptions of this type of misuse; 392 (36.4%) completed surveys were returned. The most frequently reported veterinary medications misused in humans were analgesic, anti-inflammatory medications, anti-arthritis medications, or both; systemic antibiotics, topical anti-infectives; and topical corticosteroids. People involved with rodeo, horse racing, and health care; rural area residents; and those lacking health insurance were perceived to be the groups most likely to misuse veterinary drugs. Veterinarians rated the following as likely reasons for misuse: having an independent self-sufficient attitude, convenient availability of veterinary medications, lower cost, and belief that veterinary medications are stronger than comparable human medications. Human misuse of veterinary drugs may be more common than many health practitioners realize. Limitations of this study include the response rate, sampling of veterinarians licensed in only one rural state, and reliance on veterinarians' recall of relevant instances of misuse and their perceptions of groups most likely to misuse these drugs and why. These limitations make it difficult to determine if the problem is being under- or over represented relative to the general population. However, regardless of the magnitude of the problem in the rural population, the general population, or both, the potential for harm is great. Patients with risk factors for this form of misuse should be questioned by their physician in a nonthreatening manner to detect use of veterinary medications and to provide an opportunity to inform them of the risks. PMID- 12135153 TI - The health effects of rural-urban residence and concentrated poverty. AB - This research quantifies the extent to which excess morbidity in rural areas is associated with individual characteristics, county income, and neighborhood poverty. Census geographic codes were assigned to people 25 to 64 years old (n = 176,930) from the National Health Interview Survey, 1989 to 1991, in order to link individuals to the U.S. Department of Agriculture's county urban-rural classification scheme and to 1990 county per capita income and poverty concentration in Census tracts. General health status and limitation of activity were analyzed in logistic and multinomial logit models. Residents of rural counties were at greater risk for health problems compared to residents of metropolitan and central core counties. In adjusted models, the health disadvantage of rural areas was partly explained by differences in population composition. The residual rural disadvantage was concentrated in people with less than a high school education. Tract poverty and county per capita income were also important independent predictors of morbidity. The results of this study suggest that special attention should be paid to improving education in disadvantaged places and to better understanding the ways in which economic growth and its benefits are distributed. PMID- 12135154 TI - Where to seek care: an examination of people in rural areas with HIV/AIDS. AB - Though HIV/AIDS has spread to rural areas, little empirical evidence is available on where patients living in these areas receive care. This article presents estimates of rural residents in care for HIV/AIDS, their demographic and health related characteristics, information about whether they receive care in a rural or urban setting, and data on the drug therapies prescribed. The estimates come from the HIV Cost and Services Utilization Study (HCSUS), a nationally representative probability sample of HIV-infected adults receiving care in the contiguous United States. Regardless of the definition used--enrollment site, usual source of HIV care, or site of most recent hospitalization--almost three quarters of rural residents with HIV/AIDS obtained their health care in urban areas. The authors find that differences in the demographic characteristics of those using urban vs. rural care do not drive the decision on where to obtain care, with the primary difference being that people with a rural provider tend to be older. Rural residents with an urban usual source of HIV care incurred significant inconvenience in obtaining care--the majority said their care was not conveniently located, they had substantially longer mean travel times, and over 25% had put off obtaining care in the past 6 months because they did not have a way to get to their provider. Given the considerable burden this places on a chronically ill population,further research is needed to explore how provider supply and provider experience affect the decision to travel for care and how quality of care is affected. PMID- 12135155 TI - Telehealth: an assessment of growth and distribution. AB - Telemedicine uses telecommunications technologies to deliver health care to populations located at some distance from health care providers. Telehealth is a somewhat broader term that includes non-clinical applications (such as education). This article draws on data from a nationwide survey of telehealth networks that assesses the extent and character of telemedicine activity. Although statistics point to increased utilization, the distribution of growth has been skewed toward a few applications and it is not clear that rural areas would derive substantial benefit in the absence of federal subsidy. Reported barriers to building sustainable programs often seem to reflect concerns of large hospitals and health care systems rather than constraints faced by rural organizational and individual network members. The potential role of government in addressing barriers is discussed. PMID- 12135156 TI - Rural-urban home health care differences before the Balanced Budget Act of 1997. AB - This study arose from concerns that home health care may be more difficult to provide to rural than urban elderly patients (because of geographic barriers, personnel shortages, and other factors) and may therefore be less effective in terms of patient outcomes. Case mix, home health care service use, and outcomes (primarily discharge status) were analyzed for a national random sample of 3,869 rural and urban elderly home health patients. Longitudinal data covered the period from home health admission to discharge or 120 days (whichever occurred first). Primary data collection instruments were designed to obtain longitudinal patient-level health status data; agency records and Medicare data provided service use information. (The study did not address access but focused on services and outcomes after admission to home health care.) Two-group statistical tests and multivariate analyses were employed to assess rural-urban differences. The major findings were that, after adjustment for rural-urban case mix and agency differences, rural compared to urban patients received fewer home health services and attained less favorable discharge outcomes. For example, the rural patients had a higher case mix adjusted hospitalization rate. Because the study data pertain to 1995 through 1996, the results provide a baseline for future analyses of possibly different rural compared to urban effects of the Balanced Budget Act of 1997, which resulted in major changes in Medicare payment for home health care. PMID- 12135157 TI - Synthesis, characterization and antibacterial activities of some N-bridged heterocycles containing triazole, quinoline and nitrofuran moieties. AB - A series of 2-(2-furyl)4-quinolinecarboxylic acids (3), 2-(5-nitro-2-furyl)4 quinolinecarboxylic acids (6), 4-(3-aryloxymethyl-1,2,4-triazolo[3,4-b]-1,3,4 thiadiazolo-6-yl)-2-(furyl)quinolines (5) and 4-(3-aryloxymethyl-1.2,4 triazolo[3,4-b]-1,3,4-thiadiazolo-6-yl)-2-(5-nitro-2-furyl)quinolines (7) were synthesized. The structures of the newly synthesized compounds are confirmed on the basis of elemental analysis, IR, 1H-NMR and mass spectral data. The newly synthesized compounds are evaluated for their antibacterial activities. Compounds containing nitrofuran moiety showed excellent antibacterial activity. Results of such studies are disscussed in this paper. PMID- 12135158 TI - Synthesis of some new thiazole derivatives. antifungal activity and ultrastructure changes of some mycotoxin producing fungi. AB - Several new thiazoles 2-7,10-12; 2,3-diphenyl-5-(2-thienyl)imidazo[2,1-b]thiazole 8, 3,5-di-(2-thienyl)imidazo[2,1-b]thiazole 9 and 4-amino-2-imino-6-(2 thienyl)thiazolo[3,2-a] pyrimidine 13 and 6-(2-thienyl)-3H-thiazolo[3,2 a]pyrimidin-2,4-diones 14 have been synthesized. The prepared compounds were evaluated for their in-vitro antifungal activity, compared with Tioconazol fungicide. The strong antifungal activity is elicited by compounds 5 and 8. The most active compound 5 was found to induce changes in the ultrastructure of mycotoxin producing fungi (Aspergillus flavus and Aspergillus ochraceus), this was observed by both scanning and transmission electron microscopy. PMID- 12135159 TI - Pyridazine derivatives and related compounds, part 9. tetrazolo[1,5-b]pyridazine 8-carbohydrazide: synthesis and some reactions. AB - The reaction of the hydrazide of 6,7-diphenyltetrazolo[1,5-b]pyridazine-8 carboxylic acid 3 with aromatic aldehydes gave 8-arylidenecarbohydrazide derivatives. The reaction of 3 with methanesulfonyl chloride, benzenesulfonyl chloride, phenyl and benzylisothiocyanate afforded the corresponding N-substitute derivatives. The reaction of 3 with potassium ethylxanthate gave 5-(6,7 diphenyltetrazolo[1,5-b]pyridazin4-yl)-1,3,4-oxadiazole-2-thione 7. The alkylation of this product in an alkaline medium proceeds at the sulfur atom, while the aminomethylation and acylation proceed at the nitrogen atom. Compound 3 was also reacted with N-aminothiosemicarbazide to give 5-(6,7 diphenyltetrazolo[1,5-b]pyridazin-8 yl)4-amino-1,2,4-triazole-3-thione 11. PMID- 12135160 TI - Physicochemical compatibility between ketoprofen lysine salt injections (Artrosilene Fiale) and pharmaceutical products frequently used for combined therapy. AB - Ketoprofen lysine salt (Artrosilene Fiale), a non steroidal anti-inflammatory agent, is frequently administered in association regimen with other drugs, such as steroidal anti-inflammatory, muscle relaxant, local anaesthetic and anti spastic drugs or vitamins. The aim of this study was to investigate the physicochemical compatibility between ketoprofen lysine salt (Artrosilene Fiale) and other injectable drugs frequently used in association. Physicochemical properties of ketoprofen lysine salt mixtures with different drugs, including colour, clarity, pH and drug content were observed or measured before and after (up to 3 hours) mixing at room temperature and under light protection. Results show that the association of Artrosilene Fiale with different drugs and vitamins does not cause, up to three hours f rom mixing, any significant variation in thephysicochemical parameters mentioned above, except for the association with Benexor B12 where a persistent phase separation occurs. In conclusion the results obtained demonstrated the physicochemical compatibility of Ketoprofen lysine salt (Artrosilene Fiale) with diverse drugs and vitamins, with a single exception. PMID- 12135161 TI - Quality control in the meat industry: application of the HACCP system to a slaughterhouse of ostriches. AB - The Hazard Analysis and Critical Control Point (HACCP) is a preventive system that tries to guarantee the safety and food innocuousness, anticipating the protection and correction of failures, improving the quality costs for faults of microbiological, physical and chemical type, and saving almost the final super control, which though it allows a relative guarantee of the product, its consequence will be the destruction of the product in case of detection of the failure too much late, with the consequent incremental cost. In this work, the specific hazards which can be found in the slaughter line of ostriches, the preventive measurements that can be applied in the slaughterhouse, the surveillance systems to implement, the corrective actions foreseen and the control records to be kept by the plant are described in detail. Putting in practice these knowledge will allow, to any slaughterhouse of ostriches, a self control of its productions based on the HACCP system. PMID- 12135163 TI - Pharmaco-economical analysis of the treatment with antibiotics in a surgery department. AB - The fulfillment of a pharmaco-economical analysis of the treatment with antibiotics is an important task in the conditions of a transition period, that is currently in Bulgaria. The great problem with the insufficient financial sources prevents the supply with drugs, medical supply and technology. This analysis is a kind of an auxiliary source in the process of managing in the sphere of the health policy and in particular--in the sphere of the pharmacy. PMID- 12135162 TI - Study of the interaction between policosanol and excipients. AB - Policosanol is an active principle, composed by 8 fatty alcohols: 1tetracosanol, 1-hexacosanol, 1-heptacosanol, 1-octacosanol, 1-nonacosanol, 1-triacontanol, 1 dotriacontanol and 1-tetratriacontanol that shows a very stable, well defined and reproducible composition from batch to batch that is analysed using gas chromatography. Continuing the studies of the compatibility among policosanol and different tablet excipients, it was studied if the mixtures of those excipients with policosanol produce chemical interactions between them, the samples were analysed using gas chromatography and was determined if it was affected the content of policosanol in them. When all the samples were analysed, no changes in the policosanol content of the samples were observed, and it was considered that no interactions are produced in any of the mixtures policosanol/excipients under study. PMID- 12135164 TI - Calculating the cost for drug treatment including the adverse drug reactions treatment cost (primer for fentanyl TTS in Bulgaria). AB - This study presents the estimation and comparison of the direct outpatient costs associated with the use of Morphine slow-tard tabul and trans-dermal therapeutic system /TTS/ with Fentanyl for the treatment of the chronic pain in oncological patients. There are compared all outpatients' costs for the medicines under consideration--Morphine slow-tard tabul in two different packages and Fentanyl TTS. The calculation summarizes the price of the drug therapy for 15 days course, costs for care (physician, pharmaceutical, nursing), transport, storage. After the assessment of the costs for prophylactics and treatment of the adverse drug reactions /ADR/ associated with the use of the compared drugs by creating the "Decision tree" the respective cots are added to the direct cost of patients treatment. The direct treatment costs with Fentanyl TTS exceeded those with Morphine slow-tard with 10% and thus reflects the total costs for outpatient practice. The costs of treatment of the ADRs are higher with 69% for Morphine slow-tard due to high probability of appearance of the ADRs, their seriousness and duration. PMID- 12135165 TI - An in vitro study on the kinetics, subcellular distribution and phototoxicity of harmine in human tumor cells. AB - The beta-carboline alkaloid harmine, which was found to possess interesting phototoxic properties in different biological systems, was investigated for photokilling of human tumor cells in vitro. Harmine was readily accumulated by HeLa cells, localized preferably in cytoplasm, being a non toxic compound when used at low concentrations. The photoactivation of harmine-loaded cells with UV radiation showed lysosomal damage, reflected by altered localization of the fluorescent probe acridine orange, as well as an evident cell killing which increase with time up to a maximal value 48 h after the photodynamic treatment. PMID- 12135166 TI - Evaluation of the analgesic activity of an ethanol extract of Miconia fallax. AB - A significant analgesic effect was observed after oral administration of an ethanol extract of Miconia fallax to mice, evaluated using chemical (writhing test) and thermal (hot plate test) stimuli. The extract showed a significant effect compared to control in both algesimetric models, suggesting that it was efficient in increasing the pain threshold. PMID- 12135167 TI - Dry bean protein functionality. AB - Dry beans are an important source of proteins, carbohydrates, dietary fiber, and certain minerals and vitamins in the human food supply. Among dry beans, Phaseolus beans are cultivated and consumed in the greatest quantity on a worldwide basis. Typically, most dry beans contain 15 to 25% protein on a dry weight basis (dwb). Water-soluble albumins and salt-soluble globulins, respectively, account for up to 10 to 30% and 45 to 70% of the total proteins (dwb). Dry bean albumins are typically composed of several different proteins, including lectins and enzyme inhibitors. A single 7S globulin dominates dry bean salt soluble fraction (globulins) and may account for up to 50 to 55% of the total proteins in the dry beans (dwb). Most dry bean proteins are deficient in sulfur amino acids, methionine, and cysteine, and therefore are of lower nutritional quality when compared with the animal proteins. Despite this limitation, dry beans make a significant contribution to the human dietary protein intake. In bean-based foods, dry bean proteins also serve additional functions that may include surface activity, hydration, and hydration-related properties, structure, and certain organoleptic properties. This article is intended to provide an overview of dry bean protein functionality with emphases on nutritional quality and hydration-related properties. PMID- 12135168 TI - Thermodynamic aspects of biopolymer functionality in biological systems, foods, and beverages. AB - Molecular mimicry and molecular symbiosis are proposed to be the main factors controlling thermodynamic activity and phase behavior of macromolecular compounds in foods, beverages, and chyme. Molecular mimicry implies a chemical resemblance of hydrophilic surfaces of globular proteins with their chemical information hidden in the hydrophobic interior and low excluded volume of the globules. The molecular mimicry contributes to the efficiency of enzymes. Molecular symbiosis means that interactions attraction or repulsion) between biopolymer molecules greatly differing in conformation (globular and rod-like) favor the biological efficiency of one of them at least. The symbiosis is based on excluded volume effects of macromolecules in mixed solutions. Association-dissociation of rod like macromolecules can dictate thermodynamic activity of an enzyme in the mixed solution. Thermodynamic incompatibility is typical of food macromolecules, whose denaturation, association, complexing, and chemical modification reduce their mimicry and co-solubility. Foods are normally phase-separated systems with highly volume-occupied phases. The phase-separated nature of the gel-like chyme is important to the efficiency of digestion of mixed diets. Phase separation of biopolymer mixtures, presumably, underlies mechanisms of nonspecific immune defense. The phase behavior-functionality relationships is presented through concrete examples of some foods (such as milk products, low-fat spreads, ice cream, wheat and rye doughs, thermoplastic extrudates, etc.), beverages (tea and coffee), and chyme. PMID- 12135169 TI - Renal reduction therapy for patients with autosomal dominant polycystic kidney disease. PMID- 12135170 TI - Does radiation therapy for brain tumor affect pituitary function, resulting in isolated ACTH deficiency? PMID- 12135171 TI - Slowly progressive, not rapidly progressive, MPO-ANCA positive glomerulonephritis and its characteristics. PMID- 12135172 TI - Acute viral encephalitis: the recent progress. AB - In the new Japanese control law for infectious diseases, most varieties of acute viral encephalitis belong to Category IV requiring report of all cases at sentinel hospitals. Herpes simplex virus type 1 (HSV-1) encephalitis comprises the majority of cases. With the increased prevalence of diagnostic procedures such as polymerase chain reaction (PCR), several forms of HSV-1, and -2 central nervous system (CNS) infections, including acute disseminated encephalomyelitis, brainstem encephalitis, and myelitis, have been clarified. Since 1990 we have conducted a survey of HSV CNS infections in the Kyushu and Okinawa regions, and the data are reviewed here. Trends include an increase in a new subtype of non herpetic acute limbic encephalitis. In contrast, the incidence of Japanese encephalitis (JE) in Japan has dramatically decreased to a few patients per year; however, JE remains a threat for those with decreased or absent immunity to the JE virus. Imported emerging and reemerging CNS infections such as Murray Valley and West Nile encephalitis can occur in Japan. Influenza-associated encephalitis/encephalopathy is also described as a threat for adults as well as young children. PMID- 12135173 TI - Hemodynamic effects of inhaled nitric oxide using pulse delivery and continuous delivery systems in pulmonary hypertension. AB - OBJECTIVE: Inhaled nitric oxide (NO) has been used for pulmonary vasodilation therapy in patients with pulmonary hypertension. Inhaled NO for awake and ambulatory patients, however, is unusual because it requires intubation or a tightly fitting facemask, and a large-scale delivery system for the safe management of toxic nitrogen oxides. We undertook this study to investigate the possibility of using inhaled NO therapy for awake and ambulatory patients with pulmonary hypertension. METHODS: Patients with pulmonary hypertension underwent cardiac catheterization and hemodynamic variables were measured at the baseline, after inhaled NO using our pulse delivery system, which involved a nasal cannula and a pulse device, and after inhaled NO using a continuous delivery system. PATIENTS OR MATERIALS: We studied seventeen patients with precapillary pulmonary hypertension (4 men and 13 women; age, 41+/-3, ranging from 19 to 61). RESULTS: Cardiac output was increased significantly by each system. Pulmonary vascular resistance was decreased significantly by each system. There was no significant change in mean pulmonary artery pressure, mean systemic artery pressure, or systemic vascular resistance. The concentrations of NO and nitrogen dioxide (NO2) in the expiratory gas using the pulse delivery system were 0.0 ppm as long as the pulse device was synchronized with the patient's respiratory cycle. CONCLUSION: Inhaled NO using our pulse delivery system changed the hemodynamic variables similarly to those when using the continuous delivery system. The concentrations of NO and NO2 in the expiratory gas using the pulse delivery system were within safe limits. PMID- 12135174 TI - Oxygenation abnormalities in normoxemic patients with mild liver cirrhosis. AB - OBJECTIVE: Nitric oxide (NO) production is enhanced in patients with liver cirrhosis (LC). Although most patients with mild LC have neither dyspnea nor platypnea, they might have mild oxygenation abnormalities due to intrapulmonary vasodilatation caused by increased NO. We investigated whether oxygenation abnormalities, such as hypoxemia and orthodeoxia, are present in patients with mild LC. METHODS: We investigated 148 consecutive patients with biopsy-proven chronic liver diseases such as CH (noncirrhotic chronic hepatitis) (n=46), LC(A), LC(B), and LC(C) (LC Child's A, B, and C) (n=18, 51, 33, respectively). The oxygen saturation by pulse oximetry (SpO2) in the supine and upright positions was determined in patients and controls (normal subjects, n=29). The change in SpO2 on standing was defined as deltaSpO2. NO output in exhaled air was measured in 16 patients. RESULTS: Four patients [two LC(B) and two LC(C)] had hypoxemia (supine SpO2< or =94% and/or upright SpO2< or =94%). Although there was no intergroup difference in the supine SpO2 or the upright SpO2, the deltaSpO2 decreased [control, +0.2+/-0.6%; CH, +0.1+/-0.9%; LC(A), -0.3+/-0.8%; LC(B), 0.2+/-0.9%; LC(C), -0.5+/-1.1%; mean+/-SD; p=0.005] with worsening liver disease, and the prevalence of desaturation on standing (deltaSpO2< or =-1%) increased [control, 7%; CH, 20%; LC(A), 33%; LC(B), 35%; LC(C), 42%; p=0.01]. The NO output was inversely correlated with deltaSpO2 (r=-0.66, p=0.006). CONCLUSIONS: Desaturation on standing is present in one-third of normoxemic patients with mild LC of Child's A, and is associated with the severity of liver disease. This postural desaturation is correlated with the exhaled NO, which suggests that intrapulmonary vasodilatation may play some role in this phenomenon. PMID- 12135175 TI - Intestinal obstruction in autosomal dominant polycystic kidney disease. AB - A 42-year-old woman with autosomal dominant polycystic kidney disease (ADPKD) was admitted to our hospital on April 29, 1999, with complaints of abdominal pain. A diagnosis of intestinal obstruction was reached on the basis of clinical findings and X-ray evidence. A computed tomography scan of the abdomen showed massively enlarged kidneys, especially the right kidney, which seemed to compress the small intestine. The patient underwent percutaneous aspiration of the largest cysts on the surface of the right kidney. The symptoms, in this rare case of intestinal obstruction by an enlarged kidney in ADPKD, were alleviated the day after the aspiration procedure. PMID- 12135176 TI - Two cases of polymorphic ventricular tachycardia induced by the administration of verapamil against paroxysmal supraventricular tachycardia. AB - Verapamil is widely used for the termination of paroxysmal supraventricular tachycardia (PSVT) with little proarrhythmic effect. We describe two cases of PSVT that changed to non-sustained polymorphic ventricular tachycardia after administration of verapamil. Electrophysiological study revealed atrioventricular nodal reentrant tachycardia in the first case, and atrioventricular reentrant tachycardia due to a concealed left lateral accessory pathway in the second case. Catecholamine-induced automaticity was one of the possible mechanisms of VT in the first case, but the mechanism is unknown in the second case. PMID- 12135177 TI - Sarcoidosis initially manifesting as symptomatic hypercalcemia with the absence of organic involvement. AB - A 53-year-old man was admitted to Osaka City University Hospital on July 21, 1998, for investigation of symptomatic hypercalcemia. Laboratory data on admission revealed that serum Ca had increased to around 12.6 mg/dl and there was a significant increase in urinary Ca excretion. The serum phosphate level remained normal. Although the serum PTH level was below the detection limit, serum 1,25-dihydroxyvitamin D (1,25(OH)2D) was increased. Diagnosis of sarcoidosis was supported by a negative tuberculin test and by the elevated levels of serum angiotensin-converting enzyme (ACE), lysozyme activity, and CD4/CD8 ratio in bronchoalveolar lavage specimen; there was however no imaging evidence of sarcoidosis such as bilateral hilar lymphnode enlargement on chest X ray, high resolution CT or 67Ga citrate scintigraphy. Biopsy specimens from the cervical lymphnode revealed no epitheloid cell granulomas or giant cells. Administration of prednisolone achieved a decrease in serum ACE and 1,25(OH)2D levels, followed by restoration of serum Ca and urinary Ca excretion to the normal range, and finally by an increase of serum PTH to the normal level. These observations suggested that the hypercalcemia could be explained by extrarenal production of 1,25(OH)2D. We report here on this rare case of sarcoidosis with initial symptoms of symptomatic hypercalcemia resulting from extrarenal production of 1,25(OH)2D. PMID- 12135178 TI - Complete adrenocorticotropin deficiency after radiation therapy for brain tumor with a normal growth hormone reserve. AB - A 34-year-old man with neurofibromatosis type 1, who had received radiation therapy after the excision of a brain tumor 5 years earlier, was admitted to our hospital with vomiting and weight loss. Cortisol and adrenocorticotropin (ACTH) were undetectable before and after administration of 100 microg corticotropin releasing hormone. The level of growth hormone without stimulation was 24.7 ng/ml. We diagnosed him to have complete ACTH deficiency attributable to radiation therapy. This is the first known case of a patient with complete ACTH deficiency after radiation therapy and a growth hormone reserve that remained normal. PMID- 12135179 TI - MPO-ANCA-positive slowly progressive glomerulonephritis with focal tuft necrosis and crescents. AB - A 37-year-old woman had been found to have proteinuria in October 1996. About 8 months later, the first renal biopsy was performed, revealing focal segmental necrotizing and crescentic glomerulonephritis. At that time, serum creatinine was 1.0 mg/dl and urinary protein 3+. In October 1999, laboratory tests revealed positivity for MPO-ANCA and a serum creatinine level of 1.42 mg/dl, anemia and proteinuria of 2+. A second renal biopsy showed almost the same histological findings. Accordingly, a diagnosis of MPO-ANCA positive glomerulonephritis was made. This patient was thought to be a rare case of MPO-ANCA-positive slowly progressive glomerulonephritis presenting focal segmental tuft necrosis and crescents. PMID- 12135180 TI - Pulmonary hyalinizing granuloma with hydronephrosis. AB - A 49-year-old man was admitted for the evaluation of a bilateral mass shadow in his chest X-ray film. No definitive diagnosis was established either by brushing cytology or biopsy through bronchoscopy. No malignancies were suggested by general work-up. Both masses were surgically removed, and were diagnosed as pulmonary hyalinizing granuloma (PHG). Fifteen months later, low grade fever continued and the renal function decreased. Laboratory examinations revealed bilateral hydronephrosis with polyclonal hypergammaglobulinemia. The findings of abdominal CT and urography were compatible with retroperitoneal fibrosis. Steroid treatment completely reversed the initial abnormality in laboratory data and the symptoms disappeared. PMID- 12135181 TI - Detection and monitoring of methotrexate-associated lung injury using serum markers KL-6 and SP-D in rheumatoid arthritis. AB - The applicability of monitoring concentrations of serum KL-6 and serum surfactant protein-D (SP-D) in the detection of methotrexate-associated lung injury (MTX pneumonitis) in patients with rheumatoid arthritis (RA) was investigated. The concentrations of these markers, sequentially measured in two patients with RA complicated with MTX pneumonitis, were increased in accordance with the severity of MTX pneumonitis. Conversely, the concentrations of these markers were decreased with the improvement of MTX pneumonitis, suggesting that the monitoring of these markers could be applicable not only for detecting the onset of MTX pneumonitis, but also for detecting the therapeutic response of MTX pneumonitis. PMID- 12135182 TI - Multiple atypical adenomatous hyperplasia with synchronous multiple primary bronchioloalveolar carcinomas. AB - We report a case of multiple atypical adenomatous hyperplasia (AAH) associated with synchronous multiple primary bronchioloalveolar carcinomas (BACs). A 58-year old man was visited for bronchial asthma. A chest computed tomography (CT) scan revealed small, multiple nodules with ground glass attenuation (GGA) throughout both lungs, predominantly in the upper lobes. A high resolution CT (HRCT) scan disclosed well-defined nodules with uniform GGA. Thoracoscopic wedge lung biopsy confirmed the diagnosis. The patient was treated with chemotherapy and had stable disease for two years. It is important to recognize that multiple AAH associated with multiple BACs can present as diffuse, well-defined nodules with uniform GGA on HRCT. PMID- 12135183 TI - Chronic eosinophilic pneumonia due to visceral larva migrans. AB - A 38-year-old woman presented with worsening cough, blood eosinophilia, and pulmonary infiltrates. Bronchoalveolar lavage showed 96.4% eosinophils. The diagnosis of visceral larva migrans (VLM) was made based on the positive results in enzyme-linked immunosorbent assay for Toxocara canis together with clinical symptoms and laboratory data. Pulmonary infiltrates due to VLM generally manifest as a transient form of Loffler's syndrome or simple eosinophilic pneumonia mainly in children. Here we report an adult case of VLM, with pulmonary infiltrates pathologically proven to be eosinophilic pneumonia, which persisted for 7 weeks before anthelmintic treatment with albendazole and manifested as chronic eosinophilic pneumonia. PMID- 12135184 TI - Exogenous lipoid pneumonia following ingestion of liquid paraffin. AB - An asymptomatic patient with exogenous lipoid pneumonia (ELP) due to silent aspiration of liquid paraffin ingested as a lubricant was diagnosed by bronchoalveolar lavage (BAL). BAL fluid separated into oily upper phase and lower aqueous phase spontaneously. Microscopic analysis of BAL cells revealed the presence of lipid-laden alveolar macrophages. Classic histochemical staining and electron microscope examination indicated that neutral lipid was dominant but phospholipid was also present in the lipid-laden macrophages. Together with the history of ingestion of liquid paraffin, we identified that the ingested liquid paraffin was the origin of the neutral lipid in the lipid-laden macrophages observed in the BAL fluid. PMID- 12135185 TI - Bronchioloalveolar carcinoma masked by gravity-dependent gradient on computed tomography. AB - A 78-year-old woman was found to have a small bronchioloalveolar carcinoma with ground-glass attenuation in the gravity-dependent gradient in the left lower lobe during a preoperative chest computed tomography (CT) evaluation, which was performed for previously-diagnosed adenocarcinoma of the right upper lobe. To remove the gravitational effect of the CT, the patient underwent a thin section CT in the prone position. Then, a ground-glass attenuation was revealed clearly in the left lower lobe. Postoperative pathological diagnosis was synchronous multiple bronchioloalveolar carcinomas, stage IA. This case suggests that focal areas of ground-glass attenuations on a thin-section CT in patients with BAC would be considered to be multicentric development of BAC. CT with the patient in the prone position helps to exclude the gravitational effect and narrow the differential diagnosis of ground-glass opacity, including localized forms of BAC. PMID- 12135186 TI - Improvement of long-standing iron-deficiency anemia in adults after eradication of Helicobacter pylori infection. AB - We report two cases of long-standing iron-deficiency anemia in premenopausal women that improved after eradication of H. pylori infection. There were no ulcerations or hemorrhagic lesions in the gastrointestinal tract and no bleeding focus in gynecological organs. Both cases showed H. pylori infection in the stomach and gastric atrophy. After successful eradication of H. pylori infection, the iron-deficiency anemia in both patients dramatically improved, and neither patient suffered from anemia for about 2 years. The cure of H. pylori infection is an optional treatment for iron-deficiency anemia in one fraction of the patients. PMID- 12135187 TI - Multifocal relapsing-remitting myelitis in a patient with atopic dermatitis: multiple sclerosis or atopic myelitis? AB - We describe a patient with multifocal relapsing-remitting myelitis. This case had abnormal high intensity lesions in the spinal cord at the cervical and thoracic levels on magnetic resonance imaging (MRI). She had complicated atopic dermatitis and high serum IgE levels, which showed strong sensitivity to mite antigen. These features closely resemble those of atopic myelitis, suggesting that atopic myelitis may develop a relapse with multifocal involvement of the spinal cord, which can mimic multiple sclerosis. PMID- 12135188 TI - Complex regional pain syndrome type I induced by pacemaker implantation, with a good response to steroids and neurotropin. AB - An 84-year-old woman was referred to our hospital because of aches and pain in her left hand and foot. Three months before her symptoms occurred, a pacemaker had been implanted for the treatment of a 2:1 atrioventricular block with bradycardia. In an X-ray examination, prominently decreased bone density was noted in her left fingers and toes. She was diagnosed to have CRPS-I, which was considered to have been induced by the pacemaker implantation. After treatment with methylprednisolone and Neurotropin, her symptoms dramatically improved. PMID- 12135189 TI - Dermatomyositis relapse complicated with gastric carcinoma and lupus nephritis five years after the initial diagnosis of dermatomyositis. PMID- 12135190 TI - Primary brain lymphoma. PMID- 12135191 TI - The effects of catechin on superoxide dismutase activity and its gene expression in pheochromocytoma cells. AB - BACKGROUND: Overexpression of manganese superoxide dismutase (MnSOD) cDNA via plasmid transfection leads to growth inhibition in vitro and in vivo in various human cancers. Polyphenolic compounds such as catechin isolated from tea bush Camellia sinensis has been shown to have anticancer effect in vitro. This study evaluated the effect of catechin on the MnSOD activity and mRNA level of pheochromocytoma cells (PC-12). METHODS: PC-12 cells were incubated with different concentrations of catechin at short-term (2 days) and long-term (7 days) in Dulbecco modified Eagle medium. The activity of superoxide dismutase (SOD) was measured and its mRNA was assayed by Northern blotting. RESULTS: After incubation for 2 days, catechin slightly but significantly increased the activity of copper/zinc superoxide dismutase (CuZnSOD). However, it did not show any significant effect at 7 days. The MnSOD activity showed significant changes in both short-term and long-term. The amount of mRNA also showed similar changes. CONCLUSIONS: Catechin is a natural antioxidant which has been shown to have antitumor effect in basic and epidemiological studies. The present data suggest that catechin can increase MnSOD gene expression in PC-12, which might have beneficial effect in tumor prevention. PMID- 12135192 TI - Development and validation of a scoring system predicting failure of endoscopic epinephrine injection therapy in Taiwanese patients with bleeding peptic ulcers. AB - BACKGROUND: Endoscopic epinephrine injection therapy (EIT) is always the first choice of treatment for bleeding peptic ulcers, although it fails in 15% to 30% of patients. The aim of this study was to develop a new scoring system to predict the failure in EIT, and to validate this scoring model prospectively. METHODS: This study enrolled 125 patients who presented with the stigmata of hemorrhage of peptic ulcers and underwent EIT. Patients with coagulopathy were excluded from the study. Univariate analysis of the clinical and endoscopic parameters to predict failure in EIT was performed first. A multiple logistic regression was used to develop a scoring system. This scoring equation was further applied to 50 prospective patients with bleeding peptic ulcers to validate its predictive value. RESULTS: EIT failed to arrest bleeding in 32 (25.6%) patients. Shock, blood transfusion of at least 500 ml, active bleeding, and ulcer size were effective in predicting failure in EIT as calculated by univariate analysis, while multivariate analysis showed shock, active bleeding and ulcer size to be the independent predictors. The scoring equation, defined as -3.14 + 1.29 (shock) + 0.99 (active bleeding) + 0.13 (ulcer size in mm), had a sensitivity of 81.8% and a specificity of 76.9% in predicting failure of EIT in the prospective cohort of 50 bleeding patients. CONCLUSIONS: The presence of shock, active bleeding (spurting or oozing hemorrhage) and ulcer size are risk factors of failure of EIT in Taiwanese patients with bleeding peptic ulcers. The scoring system based on these three parameters can predict failure in EIT, while alternative treatment should be considered in case patients fail EIT. PMID- 12135193 TI - A comparison of fructosamine and HbA1c for home self-monitoring blood glucose levels in type 2 diabetes. AB - BACKGROUND: Optimal glycemic control is believed to be essential in patients with diabetes to minimize any long-term complications. Measurement of the levels of glycated protein, such as fructosamine and glycated hemoglobin (HbA1c), is the most reliable method for assessing a period of glycemic control. This prospective study was performed to investigate whether fructosamine or HbA1c could provide a reliable index of glycemic control in type 2 diabetes. METHODS: Twenty-five patients with type 2 diabetes were studied at four-week intervals. Their fasting, preprandial and postprandial blood glucose levels were checked by glucometer twice a week for 16 weeks. Serum fructosamine and HbA1c were measured on every visit. The correlation of fructosamine and HbA1c with self-monitoring of blood glucose (SMBG) values in the previous 1 to 16 weeks prior to their measurements were calculated. RESULTS: Both fructosamine and HbA1c were significantly correlated with SMBG values from one week to 16 weeks prior to measurements. The correlation between fructosamine and SMBG was stronger in the prior 3 to 6 weeks. The correlation between HbA1c and SMBG was higher in the previous 4 to 12 weeks with the peak falling in the previous 8-10 weeks. Except for the previous one week, all the correlations were significantly stronger between HbA1c and SMBG than between fructosamine and SMBG. CONCLUSIONS: In type 2 diabetes, serum fructosamine assay can better reflect average blood glucose concentration over the previous 3 to 6 weeks and HbA1c is better reflective over the previous 8 to 10 weeks. HbA1c measurement correlates more significantly with home capillary blood glucose levels than the fructosamine assay, even over the previous 2 to 3 weeks. PMID- 12135194 TI - Evaluation of secondary adrenal insufficiency: findings by corticotropin releasing hormone test and magnetic resonance imaging in parallel. AB - BACKGROUND: The corticotropin-releasing hormone (CRH) stimulation test is reliable for diagnosing pituitary or hypothalamic adrenal insufficiency. In the present study, we evaluated secondary adrenal insufficiency in patients with pituitary disease by CRH test and magnetic resonance imaging (MRI). METHODS: A total of 12 healthy persons and 20 patients with pituitary disorder were recruited, including 6 with Sheehan's syndrome, 6 with idiopathic panhypopituitarism, 3 with isolated ACTH deficiency, 2 with pituitary apoplexy, 2 with empty sella syndrome and I with postoperative suprasellar tumor. Blood ACTH and cortisol levels were measured after 100 microg ovine CRH (oCRH) intravenous bolus infusion. RESULTS: In patients, basal ACTH was 15.9+/-17.3 pg/ml and basal cortisol was 4.8+/-4.6 microg/dl. These values were significantly lower than those in controls (p = 0.02). In 11 of 20 patients (3 with Sheehan's syndrome, 3 with panhypopituitarism, 2 with empty sella, 2 with isolated ACTH deficiency and 1 with pituitary apoplexy), a delayed and prolonged ACTH response was observed. In the other 9 patients, ACTH showed little change. In all patients, there was no apparent increase in cortisol value. Moreover this value was significantly lower than in healthy persons (6.2+/-5.4 vs. 21.1+/-6.0 microg/dl; p <0.0001). Sellar MRI showed complete empty change in all Sheehan's syndrome, apoplexy and empty sella syndrome patients. The six patients with panhypopituitarism had variable hypoplastic adenohypophysis with or without visible pituitary stalk. The three isolated ACTH deficiency cases demonstrated a normal anatomy. CONCLUSIONS: Our study showed that patients with pituitary disease had significantly lower plasma cortisol response after oCRH than controls. Thus, CRH testing appears useful for the diagnosis of central adrenal insufficiency. The two different functional ACTH responses to CRH observed in each pituitary disorder did not correlate with damage levels shown on MRI. PMID- 12135195 TI - Confirming pure-tone thresholds by auditory brainstem response in the geriatric population. AB - BACKGROUND: The aged people seeking otological consultation tend to increase for the purpose of applying for hearing disability certificate. When the subjective pure-tone thresholds (PTTs) are unreliable due to patient incooporation, the objective click auditory brainstem response (ABR) is used to confirm the hearing thresholds. In this study, we assess the accuracy of ABR thresholds in the aged and determine the "ABR confirmation criteria" for normal hearing and hearing handicap. METHODS: This is a prospective study. ABR thresholds were compared with pure-tone thresholds (PTTs) in 100 ears of 50 subjects over 60 years of age using kappa measure of agreement. Based on receiver operating characteristic (ROC) curve and positive likelihood ratio (LR), the most appropriate "ABR confirmation criteria" for normal hearing and hearing handicap were determined. RESULTS: According to kappa statistics, the agreement between ABR thresholds and PTTs is better in the 2- to 4-kHz region. For normal hearing, the agreement is best when confirming the average PTTs of 2, 3 and 4 kHz by the ABR threshold of 25 dB nHL (K = 0.53, p < 0.001). For hearing handicap, the agreement is best when confirming the PTTs of 3 kHz by the ABR threshold of 55 dB nHL (K = 0.60, p < 0.001). Based on ROC curve and positive LR, the ABR accuracy, for normal hearing, is excellent at a cutoff of 30 dB nHL compared with the average PTT of 2, 3 and 4 kHz. For hearing handicap, the ABR accuracy is best in excess of 55 dB nHL compared with the PTT at 3 kHz. CONCLUSIONS: We conclude that ABR is a useful diagnostic tool to confirm the validity of pure-tone audiogram for presbycusis. PMID- 12135196 TI - Primary brain lymphoma: a report of eight cases from a medical center in southern Taiwan. AB - BACKGROUND: Primary brain lymphoma (PBL) in Taiwan has been reported only in three series with very limited immunophenotypic characterization. METHODS: We retrospectively studied PBL cases with history review, immunohistochemistry, and in situ hybridization (ISH) for Epstein-Barr virus-encoded mRNA (EBER) from a single institution in southern Taiwan during 1989-2000. RESULTS: We found eight cases of PBL including four males and four females with mean age of 64.1 years and median of 63.0. The major presenting symptoms were headache, poor memory, slurred speech, and hemiplegia in three patients each. All patients had stage I solitary tumor. Half of the patients received tumor excision, the other half, stereotactic biopsy. Seven cases were of diffuse large B-cell type (DLBL), with expression of bcl-2 in six cases. They were all negative for CD5, CD10, bcl-6, and EBER. The eighth patient had anaplastic large cell lymphoma (ALCL) of T-cell phenotype with expression of cytotoxic markers and was positive for EBER. Two were lost to follow up. The median follow-up time for the remaining six was 11.2 months (range, 5.5 - 25.0). They all received radiotherapy with initial complete remission. Two died of the disease, another of cardiopulmonary failure, and the other of stroke or recurrence. The remaining two were free of disease for 9.6 and 25.0 months after radiotherapy alone. The 1-year survival rate was 60%. CONCLUSIONS: We have fully characterized eight cases of PBL, including seven DLBLs and one ALCL, in southern Taiwan that occurred in an older age group. Old age, immunophenotype (bcl-2-positivity and bcl-6-negativity), and lack of systemic chemotherapy were probably responsible for the shorter survival as compared to other studies. Radiotherapy seems to be effective for inducing complete remission and even long-term survival in some patients, however, systemic chemotherapy should be administered to prevent recurrence and to achieve long-term survival. PMID- 12135198 TI - Thyroglossal duct cyst with papillary carcinoma. PMID- 12135197 TI - Auricle angioleiomyoma. AB - Angioleiomyomas are subcutaneous benign tumors composed of numerous blood vessels with disoriented and hyperplasic smooth muscle layers. The incidence in female is higher than in male, with a ratio of 1.7 to 1.0. They are usually located at the lower extremities and rarely located at head and neck region. The case we report is a 66-year old male patient. The chief complaint was a brownish tumor over helix of left auricle with progressive enlargement noted for three years. Physical examination revealed an elastic, non-tender nodular lesion 15 x 15 x 6 mm in size and ovoid in shape. No other similar lesion was found on his face, neck and extremities. The tumor was excised entirely with clear margins. The diagnosis was angioleiomyoma via histology. PMID- 12135199 TI - Rainforest air-conditioning: the moderating influence of epiphytes on the microclimate in tropical tree crowns. AB - Epiphytes are often assumed to influence the microclimatic conditions of the tree crowns that they inhabit. In order to quantify this notion, we measured the parameters "temperature" (of the substrate surface and the boundary layer of air above it), "evaporative drying rate" and "evapotranspiration" at various locations within tree crowns with differing epiphyte assemblages. The host tree species was Annona glabra, which was either populated by one of three epiphyte species (Dimerandra emarginata, Tillandsia fasciculata, or Vriesea sanguinolenta) or was epiphyte-free. We found that during the hottest and driest time of day, microsites in the immediate proximity of epiphytes had significantly lower temperatures than epiphyte-bare locations within the same tree crown, even though the latter were also shaded by host tree foliage or branches. Moreover, water loss through evaporative drying at microsites adjacent to epiphytes was almost 20% lower than at exposed microsites. We also found that, over the course of several weeks, the evapotranspiration in tree crowns bearing epiphytes was significantly lower than in trees without epiphytes. Although the influence of epiphytes on temperature extremes and evaporation rates is relatively subtle, their mitigating effect could be of importance for small animals like arthropods inhabiting an environment as harsh and extreme as the tropical forest canopy. PMID- 12135200 TI - Nitrate sequestration by corticolous macrolichens during winter precipitation events. AB - Nitrogen is an essential nutrient in the biogeochemistry of forested ecosystems. The influence of canopy lichens on the winter biogeochemistry of nitrate in broadleaved deciduous forests is examined and it is hypothesized that nitrate sequestration will not differ between winter precipitation events. Rejection of this hypothesis would mean that meteorological conditions of winter precipitation events have a detectable influence on nitrate sequestration by canopy lichens and nitrate input to the forest floor. Canopy lichens of the genus Parmelia were found to influence winter nitrate stemflow inputs to forest soils differentially. Epiphytic lichens on an individual Carya glabra Mill. (pignut hickory) canopy tree, centrally located within the stand of an open deciduous forest, actively sequestered nitrate leached from the tree's woody frame, lowering aqueous stemflow inputs at the tree base. The quantities of nitrate sequestered by corticolous lichens during the 2 February 1999 mixed-precipitation event were significantly greater than those during all other precipitation events examined. Greater rates of nitrate uptake during the 2 February 1999 event may be attributed to (1) its intermediate rain intensity, which would have soaked the lichen thalli in a nutrient-rich bath, and (2) an air temperature range between 2 degrees C and 8 degrees C that would have increased viscosity and surface tension of stemflow drainage, thereby decreasing stemflow velocity and increasing the contact time of stemflow water on the lichen thalli. Other precipitation events were either too cold to promote metabolic activity by canopy lichens or too warm and intense for an optimal contact time of stemflow with lichen thalli, resulting in lower quantities of nitrate sequestered. Meteorological conditions of winter precipitation events have been documented to influence sequestration of nitrate by corticolous lichens and decrease aqueous stemflow inputs to the forest floor of broadleaved deciduous forests. PMID- 12135201 TI - Analysis of the growth of rimu (Dacrydium cupressinum) in South Westland, New Zealand, using process-based simulation models. AB - Two process-based models were used to identify the environmental variables limiting productivity in a pristine, mature forest dominated by rimu (Dacrydium cupressinum Sol. ex Lamb.) trees in South Westland, New Zealand. A model of canopy net carbon uptake, incorporating routines for radiation interception, photosynthesis and water balance was used to determine a value for quantum efficiency when climate variables were not limiting. The annual net carbon uptake by the canopy was estimated to be 1.1 kg C m(-2) and the quantum efficiency 22.6 mmol mol quanta(-1). This value of quantum efficiency, combined with other parameters obtainable from the literature, was then used in a model of forest productivity (3-PG), to simulate changes in net productivity and the allocation of carbon to tree components. The model was adjusted to match a measured stem increment of 10.6 Mg ha(-1) over a period of 13 years. To achieve this while maintaining a low, but stable value for leaf area index, it was necessary to set the site fertility rating very low and select high values for the parameters describing the proportional allocation of total carbon to roots. This approach highlighted nutrient availability as the principal constraint on productivity for the ecosystem and identified critical measurements that will be necessary for using the model to predict the effects of climate change on carbon sequestration. The low rates of carbon uptake and productivity are consistent with the low nutrient supply available from the highly leached, acid soils, most likely attributable to frequent saturation and a very shallow aerobic zone. PMID- 12135202 TI - Changes in wind regime around a nickel-copper smelter at Monchegorsk, northwestern Russia. AB - Wind stress may significantly change plant damage by aerial pollutants. However, almost no information exists on pollution-induced changes in wind regime around the strong emission sources. Wind speed, measured in industrial barrens adjacent to the nickel-copper smelter at Monchegorsk (Kola Peninsula, NW Russia), was two to three times as high as in the slightly polluted and nearly unpolluted forests. The ratio between the maximum wind velocity within 30 s and the average velocity of that time interval showed no temporal variation, thus characterising the wind regime. This ratio was highest in unpolluted forests, suggesting the predominance of gusty winds; in industrial barrens the maximum wind speed was only slightly higher than the average value. Since topography did not explain the spatial variation in wind regime, I conclude that my data represent the first direct evidence for distinct changes in wind regime caused by pollution-induced habitat deterioration. The results suggest that initial (partly pollution-induced) forest disturbance causes secondary effects (like increased snow evaporation, followed by soil freezing and plant damage) that may enhance further disturbance in a positive feedback fashion. PMID- 12135203 TI - The El Nino southern oscillation and malaria epidemics in South America. AB - A better understanding of the relationship between the El Nino Southern Oscillation (ENSO), the climatic anomalies it engenders, and malaria epidemics could help mitigate the world-wide increase in incidence of this mosquito transmitted disease. The purpose of this paper is to assess the possibility of using ENSO forecasts for improving malaria control. This paper analyses the relationship between ENSO events and malaria epidemics in a number of South American countries (Colombia, Ecuador, French Guiana, Guyana, Peru, Suriname, and Venezuela). A statistically significant relationship was found between El Nino and malaria epidemics in Colombia, Guyana, Peru, and Venezuela. We demonstrate that flooding engenders malaria epidemics in the dry coastal region of northern Peru, while droughts favor the development of epidemics in Colombia and Guyana, and epidemics lag a drought by 1 year in Venezuela. In Brazil, French Guiana, and Ecuador, where we did not detect an ENSO/malaria signal, non-climatic factors such as insecticide sprayings, variation in availability of anti-malaria drugs, and population migration are likely to play a stronger role in malaria epidemics than ENSO-generated climatic anomalies. In some South American countries, El Nino forecasts show strong potential for informing public health efforts to control malaria. PMID- 12135204 TI - No covariation between the geomagnetic activity and the incidence of acute myocardial infarction in the polar area of northern Sweden. AB - This study was undertaken to investigate whether there was any relation between the aurora borealis (measured as the geomagnetic activity) and the number of acute myocardial infarctions (AMI) in the northern, partly polar, area of Sweden. The AMI cases were collected from The Northern Sweden MONICA (multinational MONItoring of trends and determinants of CArdiovascular disease) AMI registry between 1985 and 1998, inclusive, and the information on the geomagnetic activity from continuous measurements at the Swedish Institute of Space Physics, Kiruna. In the analyses, both the relation between the individual AMI case and ambient geomagnetic activity, and the relation between the mean daily K index and the daily number of AMI cases were tested. We found no statistically significant relation between the number of fatal or non-fatal AMI cases, the number of sudden deaths or the number of patients with chest pain without myocardial damage, and geomagnetic activity. Our data do not support a relation between the geomagnetic activity and AMI. PMID- 12135205 TI - Influence of ground reflectivity and topography on erythemal UV radiation on inclined planes. AB - Erythemal UV irradiance incident on a horizontal surface is not always the best way of estimating the real dose received by humans or animals. For this purpose knowledge of the irradiance incident on inclined planes is required. This study presents a physically accurate model for the calculation of erythemal UV on inclined planes. The influence of ground reflectivity and topography on erythemal UV on inclined planes is investigated as a function of solar zenith and azimuth angle. It is shown that including directional reflectivity does not substantially change the incident dose on inclined planes, the maximum deviation being 10%. The incident erythemal UV may, however, be much more influenced by the surrounding topography and by the direct/diffuse partitioning of the irradiance (which is a function of altitude). Maximum increases in erythemal UV of +57%, compared with the incident erythemal UV on a horizontal plane, were found when the sensor faced the sun with a mountain slope to the left and right of it and for very high altitudes. PMID- 12135206 TI - The management of postpartum hypertension. PMID- 12135207 TI - Synergy between antenatal exposure to infection and intrapartum events in causation of perinatal brain injury at term. PMID- 12135208 TI - Neonatal survival rates in 860 singleton live births at 24 and 25 weeks gestational age. A Canadian multicentre study. AB - OBJECTIVE: To determine the current survival rate of singleton living newborns born at gestational age of 24 and 25 weeks, using obstetric factors available to the physician before birth. DESIGN: Retrospective study of all live births in 13 of 17 Canadian tertiary centres. Population All singleton live births without congenital abnormalities. METHODS: During the years 1991-1996, data were abstracted from clinical databases and charts of 860 live births, in 13 of the 17 tertiary centres in Canada, all with major neonatal intensive care units. Newborn survival was defined as alive at discharge from neonatal intensive care unit. Abstracted elements included gestational age, maternal antenatal corticosteroid treatment, birthweight, gender, fetal presentation and mode of delivery. RESULTS: Average survival rates increased from 56.1% at 24 weeks (n = 406) to 68.0% at 25 weeks (n = 454). Survival rates ranged from 53.1% at day 168 to 81.6% at day 181 (r = 0.802, P < 0.05). Steroid administration improved the survival rates at 24 and 25 weeks compared with that of unexposed fetuses, respectively (58.9% vs 41.8%; OR 1.70; 95% CI 1.03-2.08 and 74.2% vs 56.8%; OR 2.19; 95% CI 1.41-3.38). Caesarean delivery for breech presentation improved survival compared with vaginal delivery, both at 24 and 25 weeks (56.1% vs 36.0%; OR 2.19; 95% CI 1.10 4.34, and 68.7% vs 55.2% OR 1.78; 95% CI 0.093-3.43). Female neonates displayed better survival rates (59.6% vs 52.1% OR 1.36; 95% CI 0.92-2.01, and 72.6% vs 63.1% OR 1.51; 95% CI 1.02-2.25) at 24 and 25 weeks, respectively. Explanatory regression model confirmed these factors as prognostic variables associated with survival. CONCLUSIONS: This extensive collaborative study confirms that several prognostic factors, known before birth, including gestational age in days, steroid treatment, mode of presentation and fetal sex may help obstetricians, neonatologists and parents in their decision-making process at 24 and 25 weeks of pregnancy. PMID- 12135209 TI - Longitudinal measurement of peak systolic velocity in the fetal middle cerebral artery for monitoring pregnancies complicated by red cell alloimmunisation: a prospective multicentre trial with intention-to-treat. AB - OBJECTIVE: To evaluate the utilisation measurements of peak systolic velocities in the middle cerebral artery combined with B-mode ultrasound imaging to predict anaemia in an unselected population of pregnancies complicated by alloimmune antibodies known to cause immunological hydrops. DESIGN: Prospective study on an intention-to-treat basis. SETTING: Multicentre study in five large tertiary referral centres. POPULATION: One hundred twenty-five fetuses with maternal alloantibodies known to cause immunological hydrops. METHODS: If peak systolic velocity and B-mode scan were reassuring the pregnancy was monitored at 7-14 days interval. If either method showed signs of anaemia, an umbilical fetal blood sampling was performed. When the gestational age was greater than 35 weeks, labour was induced. MAIN OUTCOME MEASURE: Moderate to severe anaemia at delivery. RESULTS: Overall sensitivity to detect moderate to severe anaemia below 35 weeks (haemoglobin level below 0.65 multiples of median) was 88%. Specificity was 87%; positive predictive value was 53% and negative predictive value was 98%. The diagnosis of severe anaemia was missed in one fetus; however, the final outcome was good. The method was not useful after 35 weeks. CONCLUSIONS: Middle cerebral artery peak systolic velocity is a highly sensitive non-invasive means for determining the degree of anaemia present in red blood cell alloimmunised pregnancies. The widespread use of the Doppler method will minimise fetal complications associated with amniocentesis and fetal blood sampling. Non invasive measurement of middle cerebral artery peak systolic velocities is more convenient and acceptable to alloimmunised pregnancies and may significantly lower health care costs. A Doppler interval of seven days is recommended. PMID- 12135210 TI - The creation of twin centile curves for size. AB - OBJECTIVE: To create twin centile charts for size. DESIGN: Retrospective study. SETTING: Aberdeen Maternity Hospital. METHODS: Ultrasound measurements of abdominal circumference (AC), biparietal diameter (BPD) and femur length (FL) for 1,011 twin pregnancies were extracted from a databank. The methodology is based on that described by Altman and Chitty [Br. J. Obstet. Gynaecol. 101 (1994) 29.]; only one measurement from each twin was used, polynomial regression models were fitted separately to the mean and standard deviation (SD) of the ultrasound parameter as functions of gestational age. The centiles were obtained assuming that at each gestation, the measurements had a normal distribution. RESULTS: The growth pattern of AC for twins appear to follow closely that of singletons until 32 weeks. Thereafter, there is a gradual but definite fall off in growth away from singleton standards. The pattern of growth of FL is largely similar to that of singletons. From mid to early third trimester, the BPD of twin babies was larger than that of singletons. PMID- 12135211 TI - Absence of enhanced systemic inflammatory response at 18 weeks of gestation in women with subsequent pre-eclampsia. AB - OBJECTIVE: To compare indicators of systemic inflammatory response in the second trimester in women who developed pre-eclampsia with normal pregnancies. DESIGN: Prospective nested case control study derived from a cohort of 2190 pregnant women. Blood samples were obtained at 18 weeks of gestation. The following inflammatory parameters were measured: tumour necrosis factor-alpha (TNF-alpha), plasminogen activator inhibitor-1 (PAI-1), interleukin-1beta (IL-1beta), IL-6, IL 10, microCRP and tissue factor (TF). SETTING: Institute of Medical Genetics, University of Oslo, and Department of Medical Genetics, Ulleval University Hospital and Departments of Obstetrics and Gynecology, Aker University Hospital, Oslo, Norway. SAMPLE: The cases were 71 women who subsequently developed pre eclampsia. The controls were 71 healthy, pregnant women matched for age, parity and first trimester body mass index (BMI). METHODS: Venous blood was drawn from fasting subjects into 5 mL test tubes containing EDTA. Samples were analysed for inflammatory parameters: IL-1-beta, IL-6, IL-10, TNF-alpha, PAI-1, TF (ELISA technique) and CRP (latex-enhanced immunonephelometric assay), strictly according to the manufacturer's recommendation. MAIN OUTCOME MEASURES: The matched case and control subjects were compared by the paired two-tailed Wilcoxon signed rank test. All P values were two-tailed and P < 0.05 was deemed statistically significant. RESULTS: We found no differences in plasma concentrations of PAI-1, IL-1beta, IL-6,IL-10, microCRP, TNF-alpha or TF at 18 weeks of gestation between women who subsequently developed pre-eclampsia and matched control women. CONCLUSION: In contrast to findings from women with overt pre-eclampsia, the present study indicates that there are no indications of intensified systemic inflammatory response at 18 weeks of gestation in women who later develop pre eclampsia. PMID- 12135212 TI - Respiratory function in singleton and twin pregnancy. AB - OBJECTIVE: Singleton pregnancy causes important changes in respiratory function. The incidence of twin pregnancies is increasing, but it is not known whether affected women suffer greater respiratory compromise. The aim of this study was to determine if changes in respiratory function during pregnancy in healthy women were greater in those with a twin pregnancy compared with those with a singleton pregnancy. DESIGN: Cross sectional study. SETTING: London teaching hospital. POPULATION: Healthy pregnant women attending the antenatal clinic and ultrasound department. METHODS: A cross sectional study of respiratory function was performed in 68 women with twin pregnancies (17 examined in the first trimester, 35 second trimester, 16 third trimester) and 140 women with singleton pregnancies (28, 80, 40, respectively) and 22 non-pregnant women. Women were examined once between 7 and 40 weeks of gestation. Forced vital capacity, relaxed vital capacity, forced expiratory volume in 1 second (FEV1), peak flow, inspiratory capacity, functional residual capacity (FRC), expiratory reserve volume, pulmonary diffusing capacity and minute ventilation were measured. RESULTS: Both in twin and singleton pregnancies, the mean FRC and expiratory reserve ventilation of women studied in the third trimester and minute ventilation of women studied in each trimester differed significantly from that of the non pregnant women. There were, however, no significant differences demonstrated in respiratory function between healthy women with twin as compared with singleton pregnancies. CONCLUSION: In healthy women, the extra demands of atwin compared with a singleton pregnancy do not compromise further the respiratory system. PMID- 12135213 TI - Endothelial cell expression of adhesion molecules is induced by fetal plasma from pregnancies with umbilical placental vascular disease. AB - OBJECTIVE: To test the hypothesis that local production with spill into the fetal circulation of factor(s) injurious to endothelium is responsible for the vascular pathology present when the umbilical artery Doppler study is abnormal. Expression of adhesion molecules is a feature of endothelial cell activation. DESIGN: Case control study. SETTING: University teaching hospital. SAMPLES: Fetal plasma was collected from 27 normal pregnancies, 39 pregnancies with umbilical placental vascular disease defined by abnormal umbilical artery Doppler and 11 pregnancies with pre-eclampsia and normal umbilical artery Doppler. METHODS: Isolated and cultured human umbilical vein endothelial cells from normal pregnancies were incubated with fetal plasma from three study groups. mRNA expression of intercellular cell adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule 1 (VCAM-1) and platelet-endothelial cell adhesion molecule-1 (PECAM-1) were assessed by reverse transcription-polymerase chain reaction. To confirm the occurrence of this in vivo, we measured the levels of soluble fractions of sICAM 1, sVCAM-1 and sPECAM-1 in the fetal circulation in the fetal plasma used for endothelial cell incubation. RESULTS: The mRNA expression of ICAM-1 [median 1.1 (interquartile range 0.5-1.9) vs 0.7 (0.3-1.2), P < 0.05] and PECAM-1 [2.1 (1.2 3.0) vs 1.5 (0.7-2.1), P < 0.05] was significantly higher following incubation with fetal plasma from umbilical placental vascular disease compared with the normal group. There was no difference in the expression of VCAM-1 [1.2 (0.9-1.8) vs 1.1 (0.8-1.6), ns]. The group with maternal pre-eclampsia and normal umbilical artery Doppler did not differ from the normal group. In the umbilical placental vascular disease group, the results were similar in the presence or absence of pre-eclampsia. For soluble fractions of the adhesion molecules released into the fetal circulation, we found the levels (ng/mL) of sICAM- I [median 248.5 (interquartile range 197.3-315.7) vs 174.2 (144.5-212.9), P < 0.05] and sPECAM-1 [9.3 (6.2-11.1) vs 6.1 (5.4-7.7), P < 0.05] in fetal plasma to be significantly increased in the presence of umbilical placental vascular disease compared with the normal. CONCLUSIONS: Vascular disease in the fetal umbilical placental circulation is associated with an elevation in mRNA expression by endothelial cells of ICAM-1 and PECAM-1. Our study provides evidence for endothelial cell activation and dysfunction in umbilical placental vascular disease. We speculate that the plasma factor(s) affecting the vessels of the umbilical villous tree is locally released by the trophoblast. The occurrence of the maternal syndrome of pre-eclampsia appears to be independent of this. PMID- 12135214 TI - Predictive value of amniotic fluid cystatin C levels for the early identification of fetuses with obstructive uropathies. AB - OBJECTIVE: To compare the diagnostic accuracy of cystatin C with that of creatinine in discriminating renal function in fetuses without ultrasononographic evidence of renal malformations from those with obstructive uropathies. DESIGN: Prospective, observational cohort study. SETTING: Prenatal morphologic and functional evaluation of fetal obstructive uropathies throughout pregnancy. POPULATION: A total of 96 healthy pregnant women at different stages of pregnancy, without any pregnancy-related maternal disease. Eighty-one pregnant women without clinical and ultrasonographic evidence of any fetal anomaly, confirmed at birth, were defined as controls; 15 pregnant women with various fetal obstructive uropathies, evidenced by repeated ultrasound examinations and confirmed at birth, were defined as cases. METHODS: Creatinine was measured by a kinetic Jaffe picric acid method and cystatin C by a nephelometric immunoassay. Variables were analysed by applying conventional statistical tests; the non parametric receiver operating curves (ROC) analysis was used to evaluate the diagnostic efficiencies of the biochemical markers. MAIN OUTCOME MEASURES: Incidence of confirmed, diagnosed, neonatal obstructive uropathy by measuring baseline levels of cystatin C and creatinine in amniotic fluid. RESULTS: Baseline levels of cystatin C in amniotic fluid were significantly higher (P = 0.0015) among cases than in controls with comparable gestational age; no significant difference was found for creatinine levels (P = n.s.). The maximum diagnostic accuracy of serum cystatin C in discriminating controls from fetal uropathies was 96%, while that of creatinine was 62%. CONCLUSION: Cystatin C may be considered a sensitivebiochemical marker for the early identification of fetuses with obstructive uropathies. PMID- 12135215 TI - Relationship between total homocysteine and folate levels in pregnant women and their newborn babies according to maternal serum levels of vitamin B12. AB - OBJECTIVE: To determine total homocysteine and folate levels in pregnant women according to vitamin B12 (B12) levels, and to analyse the impact of maternal B12 levels on the nutritional status of their newborn babies. DESIGN: Cross sectional observational study. SETTING: Two public hospitals in Jundiai City, Sao Paulo, Brazil. SAMPLE: Sixty-nine pregnant women and their respective newborn babies at the time of delivery. METHODS: Maternal blood was collected up to 8 hours before delivery. Umbilical cord blood was collected after the expulsion of the placenta. Total homocysteine was measured by high perfomance liquid chromatography, folate by ion capture methodology and B12 by enzyme immunoassay. MAIN OUTCOME MEASURES: Relationship between low maternal vitamin B12 levels and total homocysteine and folate levels in pregnant women and newborn babies. RESULTS: There was a significant correlation between maternal and neonatal B12 levels, but not between maternal B12 and neonatal red blood cell (RBC) folate. There was a weak correlation between maternal B12 levels and neonatal serum folate. However, there were significant correlations between maternal and neonatal total homocysteine levels and between neonatal B12 and neonatal total homocysteine levels. Mean (+/- SD) umbilical cord blood B12, RBC folate, serum folate and total homocysteine levels were 1.7 +/- 0.8, 1.8 +/- 0.8, 2.2 +/- 0.8 and 0.9 +/- 0.3 times higher than maternal B12, RBC folate, serum folate and total homocysteine values, respectively. CONCLUSIONS: These data suggest that pregnant women with low B12 levels are unable to provide the necessary amount of B12 to their fetuses. The clinical consequences could be the presence of neurological abnormalities as well as the lack of utilisation of homocysteine for methionine transformation. PMID- 12135216 TI - Factors associated with maternal mortality in rural Guinea-Bissau. A longitudinal population-based study. AB - OBJECTIVE: To assess demographic and obstetric risk factors for pregnancy-related death in a multiethnic rural population in a developing country. DESIGN: A prospective survey of women in the fertile age-range. SETTING: Rural Guinea Bissau. POPULATION: More than 15,000 women living in 100 clusters were visited at six-monthly intervals over a period of more than six years. A total of 10,931 pregnancies were registered prospectively; 85 of these pregnancies resulted in maternal or late maternal death. MAIN OUTCOME MEASURE: Maternal mortality ratio. METHOD: In the rural areas of Guinea-Bissau, we conducted a prospective survey of women in the fertile age range. More than 15,000 women living in 100 clusters were visited at 6-monthly intervals over a period of more than six years. An analysis of demographic, environmental and obstetric risk factors for maternal death was performed based on 10,931 prospectively registered pregnancies; 85 of these pregnancies resulted in maternal or late maternal death. RESULTS: In the adjusted model maternal mortality ratio increased with increasing distance from the regional hospital (OR>25 km = 7.4 [95% CI: 1.6-132]). Multiple pregnancy was found to increase the risk of maternal death (OR = 3.4 [95% CI: 1.3-7.5]). The risk of subsequent maternal death was increased if the fetus was stillborn (OR = 5.3 [95% CI: 2.8-9.4]). Women living in the region of Gabu had higher mortality than those living in Biombo (OR = 2.5 [95% CI: 1.3-5.1]). No category of age or parity were associated with an increased risk of maternal mortality. Predictive values did not exceed 3% for any of the significant risk factors. CONCLUSIONS: For the purpose of reducing maternal mortality, the screening approach of antenatal care is of limited value. Age and parity should not be used routinely as selection criteria for transfer of otherwise healthy pregnant women to higher level health institutions. Twin pregnancy seems to be the only operational risk factor identified in this study. Stillbirth is associated with an increased risk of maternal death. Regional differences must be studied further. The distance to emergency obstetric care (EOC) may determine the outcome of a complicated delivery. PMID- 12135217 TI - A comparison of outpatient hysteroscopy with saline infusion hysterosonography. AB - OBJECTIVES: To establish the accuracy of saline infusion hysterosonography in diagnosing uterine pathology when compared with outpatient hysteroscopy. DESIGN: Prospective, parallel, blinded comparative study. SETTING: Outpatient hysteroscopy clinic in a large university teaching hospital. POPULATION: All women referred for outpatient hysteroscopy in a 15-month period. INTERVENTIONS: Women underwent saline infusion hysterosonography followed by flexible hysteroscopy. The ultrasonographer was blinded to the hysteroscopy result and the gynaecologist was blinded to the saline infusion hysterosonography result. MAIN OUTCOME MEASURES: The relative success rates and pain scores for each procedure. The validity of saline infusion hysterosonography as a diagnostic test. RESULTS: One hundred and seventeen women entered the study; 70 women were premenopausal and 47 postmenopausal. In 27 women, one or both procedures could not be performed. Saline infusion hysterosonography failed in 20 women, on one occasion hysteroscopy failed and both investigations failed in six women. Ninety cases remained for direct comparison, with 78 cases of agreement on the uterine findings in both investigations. Twelve cases disagreed; in one case, an adhesion was seen, two cases with polyps and five with fibroids seen on ultrasound but not seen on hysteroscopy. There were four cases where polyps were identified on hysteroscopy but not on saline infusion hysterosonography. The median pain scores were 1.6 for saline infusion hysterosonography and 3.2 for hysteroscopy. CONCLUSIONS: Both saline infusion hysterosonography and hysteroscopy are well tolerated by women. Saline infusion hysterosonography has a high failure rate but has a lower pain score than hysteroscopy. PMID- 12135218 TI - Is outpatient diagnostic hysteroscopy more useful than endometrial biopsy alone for the investigation of abnormal uterine bleeding in unselected premenopausal women? A randomised comparison. AB - OBJECTIVE: To formally evaluate the clinical benefit of additional outpatient hysteroscopy over traditional vaginal examination and endometrial biopsy. DESIGN: A prospective randomised controlled trial. SETTING: A large teaching hospital in the northeast of Scotland. SAMPLE Premenopausal women with abnormal uterine bleeding referred to the general gynaecologic clinic and requiring endometrial biopsy. METHODS Women were randomised to either outpatient hysteroscopy and endometrial biopsy or endometrial biopsy alone. MAIN OUTCOME MEASURES: PRIMARY OUTCOME: initial surgical intervention rates. SECONDARY OUTCOMES: procedural success and acceptability, intrauterine pathology identified and changes in management. RESULTS: Three hundred and seventy women were recruited to the study. Initial trends in clinical management were comparable in both groups. No woman was advised to have removal of a localised lesion found at outpatient hysteroscopy and a normal uterine cavity at hysteroscopy did not influence the hysterectomy rate, which was similar in both groups. Outpatient hysteroscopy was found to be as acceptable as an outpatient endometrial biopsy and successfully completed in 85% compared with 91% of women who underwent endometrial biopsy alone. No cases of endometrial malignancy were identified. CONCLUSIONS: Outpatient diagnostic hysteroscopy is an acceptable procedure and may give more reassurance. It did not influence clinical management, especially with respect to hysterectomy rate. Outpatient hysteroscopy may be useful in selected cases, but when performed in a non-selective manner, it has little influence on clinical management and increases costs. PMID- 12135220 TI - Perineal ultrasound evaluation of urethral angle and bladder neck mobility in women with stress urinary incontinence. AB - OBJECTIVES: 1. To assess the reproducibility of an electronic ultrasonographic technique for the measurement of urethral angulation; 2. to test the ability of measurement of the urethral angle and bladder neck mobility to predict genuine stress incontinence; 3. to compare ultrasound variables in stress incontinent women and in controls. DESIGN: Case-control study. POPULATION: Twenty-three incontinent women and 50 controls. METHODS: Electronic measurements of the distance between the bladder neck and the symphysis pubis, the bladder neck and the symphysis pubis line and the midline of the symphysis (alpha angle) and the angle between the proximal and distal urethra (beta angle) by means of perineal ultrasonography with a comfortably full bladder at rest, during the Valsalva manoeuvre and during maximal pelvic floor contraction. The same procedure was performed by a second investigator. Repeatability was evaluated by the technique described by Bland and Altman. Statistical analysis was performed using Student's t test and the two-tailed paired t test. MAIN OUTCOME MEASURES: To test the possible role of the urethral angle in maintaining female continence. RESULTS: Ultrasound analysis showed good repeatability between the two observers and is not influenced by vesical volume. Beta angle and urethrovesical mobility are inversely proportional, both in continent and in incontinent women. Urethral angle identifies genuine stress incontinence better than urethrovesical mobility (sensitivity 96% vs 87%; specificity 92% vs 68%; positive predictive value 85% vs 55%). There are significant differences in all ultrasound variables between incontinent women and continent controls. CONCLUSIONS: This study suggests a significant role of the urethral angle in maintaining female continence (in incontinent women it is lower at rest and lowers with straining). Measurement of the urethral angle can provide useful additional information to that provided by ultrasound evaluation of bladder neck mobility. PMID- 12135219 TI - The impact of subcutaneous oestradiol implants on biochemical markers of bone turnover and bone mineral density in postmenopausal women. AB - OBJECTIVE: To evaluate the anabolic effect of oestrogen on bone by comparing the response of markers of bone formation (and resorption) and bone mineral density (BMD) to subcutaneous oestradiol implants. DESIGN: One year double-blind placebo controlled randomised study. SETTING: Clinical research unit within a teaching hospital. POPULATION: Twenty-one hysterectomised postmenopausal women were randomised to 25 mg oestradiol implants at baseline and at six months or to have a sham procedure at baseline and six months. METHODS: BMD and quantitative ultrasound (QUS) were assessed at baseline and one year. Bone alkaline phosphatase (bone ALP), procollagen type I N-terminal propeptide (PINP), osteocalcin (OC), free deoxypyridinoline (iFDPD), N-telopeptide of type I collagen (NTX), serum oestradiol and intact parathyroid hormone (PTH) were measured at baseline, 4, 8, 12 and 24 weeks. MAIN OUTCOME MEASURES: Percentage change markers of bone turnover and PTH and change in oestradiol levels over first six months and percentage of changes in DXA and QUS over one year. RESULTS: PINP, bone ALP and OC increased by 28%, 7% and 9%, respectively (P < 0.01) during the first four weeks of treatment and then decreased significantly. Lumbar spine (LS) and total hip (TH) BMD increased by 5.4% and 6.0% (P < 0.001), respectively, and femoral neck (FN) BMD by 3.7% (P < 0.05) during the first year of treatment compared with control subjects. The peak serum oestradiol level was achieved four weeks after implant insertion. Mean PTH levels increased significantly in subjects receiving subcutaneous oestradiol. CONCLUSION: Subcutaneous oestrogen exerted an apparent anabolic effect on bone, which was initially reflected by an increase in bone formation markers and later by a large increase in BMD. PMID- 12135221 TI - Urinary and faecal incontinence following delayed primary repair of obstetric genital fistula. AB - OBJECTIVE: To evaluate: (1) the factors associated with the development of obstetric genitourinary fistula, (2) the incidence of urinary and faecal incontinence following closure of the fistula and (3) the urodynamic findings in women with persistent urinary incontinence. DESIGN: An observational clinical study. SETTING: A specialised fistula unit in a developing country. POPULATION: Women following successful anatomical closure of obstetric genitourinary fistula. METHODS: Fifty-five women were enrolled from the Fistula Hospital in Ethiopia, following obstetric fistula repair. Their case records were reviewed and details regarding (1) antecedent obstetric factors, (2) the site, size and type of fistula and (3) pre-operative bladder neck mobility and vaginal scarring were recorded. All women were questioned regarding symptoms of faecal and urinary incontinence. Women reporting urinary incontinence following fistula repair underwent urodynamic investigations. MAIN OUTCOME MEASURES: Clinical and urodynamic assessment. RESULTS: The mean age of the women was 23 years (range 16 45 years). The fistula in 38 women (69%) followed the first delivery and in 17 women (31%) following a subsequent delivery. The mean duration of labour was four days (range 1-9 days). Forty-four women (80%) had an isolated vesico-vaginal fistula and 11 (20%) had a combined vesico-vaginal and recto-vaginal fistula. The mean diameter of the fistula was 2.9 cm (0.5-6 cm). Successful repair occurred in all women. Thirty women (55%) reported persistent urinary incontinence and 21 (38%) altered faecal continence at follow up. In the former group, urodynamic investigations identified genuine stress incontinence in 17 women (31%), detrusor instability in two (4%) and mixed incontinence in 11 (20%). CONCLUSION: This study demonstrates the high rate of successful closure of the fistula in a specialised fistula unit, but highlights the problem of persistent urinary incontinence following closure. PMID- 12135222 TI - Assessment of third degree tears using three-dimensional anal endosonography with combined anal manometry: a novel technique. AB - Three-dimensional anal endosonography has enabled sagittal and coronal reconstructions of the anal canal to be matched with longitudinal pressure data, to present a combined picture of structure and function. This novel technique has been applied to a group of women with a clinical diagnosis of a third degree tear. Endosonography showed that only 68% of women had ultrasound evidence of sphincter damage. Anal canal anatomy and pressure profile did not differ significantly between those with and those without sphincter damage, but the anterior external anal sphincter and the puborectalis tended to be shorter and the pressures were lower in those with sphincter disruption. PMID- 12135223 TI - Does endothelial cell activation occur with intrauterine growth restriction? AB - It is possible that in fetal growth restriction without pre-eclampsia endothelial cell activation does not occur. This might be either because there is no release of 'factor X' or because of maternal resistance to its effects. To test this hypothesis, we took blood samples from 26 women with pre-eclampsia (without fetal growth restriction), 13 women with fetal growth restriction (without pre eclampsia) and 24 normal pregnant controls, and measured the circulating levels of three markers of endothelial cell activation (soluble VCAM, ICAM and E selectin) and three cytokines [tumour necrosis factor-a (TNF-alpha), interleukin 6 (IL-6) and -8 (IL-8)]. The levels of the markers of endothelial cell activation were raised in both pre-eclampsia and fetal growth restriction pregnancies compared with controls; however, the levels of TNF-alpha, IL-6 and IL-8 were significantly raised in pregnancies complicated by pre-eclampsia, but not in fetal growth restriction, compared with controls. These data show that endothelial cell activation is common to both pre-eclampsia and fetal growth restriction, but that the circulating levels of cytokines are elevated only in pre-eclampsia. Thus, it seems likely that endothelial cell activation is a consequence of a failure of trophoblast invasion and that a further step is required, possibly involving cytokine release, for the expression of the full clinical picture of pre-eclampsia. PMID- 12135224 TI - Factor XI deficiency presenting in pregnancy: diagnosis and management. PMID- 12135225 TI - Pyruvate kinase deficiency in pregnancy complicated by iron overload. PMID- 12135226 TI - Gestational hyperlipidaemic pancreatitis. PMID- 12135227 TI - Frontal lobe epilepsy: diagnosis and surgical treatment. AB - Frontal lobe epilepsy has been better understood during the past two decades with the advent of technologies with improved localizing capabilities. Major technological advances in the ability to delineate structural and functional brain regions have led to a resurgence of interest in epilepsy surgery. Neuroimaging modalities have broadened the scope of what are now considered surgically remediable syndromes. In the following article, we attempt to review the current concepts regarding diagnosis and surgical management of frontal lobe epilepsy. PMID- 12135228 TI - Magnetoencephalography in presurgical epilepsy evaluation. AB - The introduction of whole-head magnetoencephalography (MEG) systems facilitating simultaneous recording from the entire brain surface has led to a major breakthrough of MEG in presurgical epilepsy evaluation. Localizations of the interictal spike zone with MEG showed excellent agreement with invasive electrical recordings, were useful to clarify the spatial relationship of the irritative spike zone to structural lesions, and could attribute epileptic activity to lobar subcompartments both in temporal lobe and extratemporal epilepsy. MEG was especially useful for the study of patients with non-lesional neocortical epilepsy and of patients with large lesions, where it provided unique information on the epileptogenic zone. It could reliably localize sensorimotor cortex prior to surgical procedures adjacent to central fissure. MEG language mapping yielded concordant results with the Wada test and cortical stimulation studies. MEG localizations of epileptic activity and essential brain regions were successfully integrated into frameless stereotaxy systems providing accurate functional information intraoperatively. Because MEG and EEG yield both complementary and confirmatory information, combined MEG-EEG recordings in conjunction with advanced source modeling techniques will further improve the noninvasive evaluation of epilepsy patients and constantly reduce the need for invasive procedures. PMID- 12135229 TI - When should conservative treatment for lumbar disc herniation be ceased and surgery considered? AB - Different authors recommend different time spans for conservative treatment before considering surgery in patients suffering from lumbar disc herniation. We analyzed the time of onset of symptoms such as pain, sensory deficit, and motor deficit in a surgically treated group in comparison to outcome after surgery in order to define a time threshold when surgical results deteriorate and operation should therefore be considered. General data, symptoms, signs, and neurological findings of 219 patients were preoperatively recorded. The outcome was evaluated according to the Prolo scale after a mean of 9.9 months. In the statistical workup, we calculated the duration of symptoms, sensory deficits, and motor deficit as continuous variables. Additionally, the population was divided into three groups of duration of symptoms, sensory deficit, or motor deficit for < or = 30 days, 30-60 days, and >60 days. Statistically significant predictors for unfavourable outcome were, for example, a longer duration of preoperative pain and motor and sensory deficit. Patients suffering for more than 60 days from disc herniation were found to have statistically worse outcome than patients suffering for 60 days or less. Findings were similar for the different time groups concerning the duration of sensory deficit but not for duration of motor deficit. The overall outcome seems to be better when patients are operated on for lumbar disc herniations within 2 months after onset of symptoms and sensory deficits. Due to these findings, we recommend conservative treatment up to 2 months and, if conservative management does not succeed, consideration of surgery. PMID- 12135230 TI - Postoperative modified stereotactic radiotherapy using a micro-multileaf collimator in patients with malignant glioma. AB - To achieve local control of malignant glioma, we designed a postoperative stereotactic radiotherapy using a micro-multileaf collimator (micro-MLC). The purpose of this study was to clarify the feasibility of this treatment. The treatment was performed in six patients who met the following eligibility criteria: (1) supratentorial tumor, (2) residual tumor volume < or = 40 cm3, and (3) Karnofsky performance status > or = 70. The three planning target volumes (PTVs), which consisted of restricted PTV (RPTV), intermediate PTV (IPTV), and extended PTV (EPTV), defined as the residual tumor plus a 1 cm, 2 cm, and 3 cm margins, respectively, and total dose delivery of 60-68 Gy, 52-60 Gy, and 44-52 Gy to the isocenters of RPTV, IPTV, and EPTV, respectively, in 4 Gy per fraction at five fractions per week, were established. The beam arrangement and the conformal blockade with a micro-MLC for the optimal treatment plan were designed. The treatment plans showed the high dose conformation to EPTV, the appropriate dose gradients in the three PTVs with the high dose homogeneity to RPTV, and the tolerated dose to critical structures. Following the plans, treatment was performed. The clinical findings more than 12 months after the treatment supported its possible use. We conclude that this treatment is feasible at least in selected patients. PMID- 12135231 TI - Management of cerebrospinal fluid leakage complicating anterior procedures through thoracotomy: report of three cases. AB - Postoperative cerebrospinal fluid (CSF) leakage is a serious complication accompanying an anterior procedure through thoracotomy, and it is difficult to cure. In this report, we present three patients with CSF leakage in the thoracic spine complicating anterior decompression and fusion for ossification of posterior longitudinal ligament who were treated by surgical or nonsurgical methods. As a surgical method, direct closure by fixing substitute dura mater with fibrin adhesive sealant or cyanoacrylate adhesive was performed in two patients. This technique was effective but required another thoracotomy. As a nonsurgical method, intrapleural administration of OK-432 through chest drainage tubes was also effective to reduce intrapleural effusions in one patient, but with this method, care must be taken for neurotoxic reactions. Both techniques seem to be useful and effective for postoperative intrapleural CSF leakages complicating anterior procedures through thoracotomy. PMID- 12135232 TI - Topographic microsurgical anatomy of the paraclinoid carotid artery. AB - In this publication, the authors describe the microanatomic topography of the entire paraclinoid area with respect to the paraclinoid segment of the internal carotid artery and its surrounding anatomical structures. Special attention was given to the borders of the paraclinoid area, cavernous sinus, arterial vessels, and cranial nerves passing through the region. The paraclinoid region was defined as a pyramid-formed space formed by the dural covering of the anterior clinoid process. The superior border is formed by the continuity of the anterior petroclinoid fold, anteriorly on the superior surface of the anterior clinoid process and medially in the direction of the diaphragma sellae. This dural sheet encircles the internal carotid artery and forms the so-called distal dural ring of the internal carotid artery. The medial border of the paraclinoid region is formed by the body of the sphenoid bone and the adjacent periosteal sheet. The inferior border is formed by a fibrous plate between the middle and anterior clinoid processes. This so-called proximal dural ring separates the venous compartments of the cavernous area from the paraclinoid area. The lateral border is formed by the lateral surface of the anterior clinoid process with its dural covering. The arterial supply of this region is provided by branches of the intracavernous carotid segment and the ophthalmic artery. The important nerves in close vicinity to the paraclinoidal area are the optic and the oculomotor nerves. Understanding and knowledge of the topographic anatomy of the paraclinoid area is essential for microsurgical exposure of this region. PMID- 12135233 TI - The effects of topical L-arginine and Ng-nitro-L-arginine methyl ester after experimental acute spinal cord injury. A light and electron microscopic study. AB - The primary objective of this study was to investigate the effects of topical L arginine and Ng-nitro-L-arginine methyl ester vs the role of ischemia in contributing to secondary injury after experimental acute spinal cord trauma. Twenty-six rabbits were submitted to spinal cord compression at the T7/8 level. The animals were divided into three groups: no applied drug (n=6), L-arginine (n=10), and Ng-nitro-L-arginine methyl ester (n=10). L-arginine was topically administered at a dose of 10 micromol (1.742 mg) per kg immediately after acute spinal cord injury. Ng-nitro-L-arginine methyl ester was applied topically at a dose of 10 pmol (10 mg/kg) immediately after acute spinal cord injury. Cortical somatosensory evoked potentials were recorded before injury and 1 min, 15 min, 30 min, and 60 min after injury. Physiological parameters were followed before, during, and I h post injury. Light and electron microscopic analysis was performed in all of the groups. In contrast to group 1, the edema of perineural, axoplasm, or surrounding tissue, the thickening of walls of the arterioles and venules, and the degeneration in myelinated axons in groups 2 and 3 were well observed. However, no differences between group 2 and group 3 occurred. PMID- 12135234 TI - Infected or complicated hydatid case mimicking a primary brain tumor. PMID- 12135235 TI - A study on latency in calves by five vaccines against bovine herpesvirus-1 infection. AB - Four bovine herpesvirus-1 (BHV-1) commercial vaccines, three of which (vaccines B, D, E) were modified live vaccines (MLV) and one (vaccine A) identified as a live strain of BHV-1 gE negative, were used for vaccination of calves, using three calves for each vaccine. Three months after vaccination calves were subjected to dexamethasone (DMS) treatment following which virus was recovered from calves inoculated with vaccine B and from those given vaccine D. No virus reactivation was obtained in calves, which received vaccines A or E. The DNA extracted from the two reactivated viruses was subjected to restriction endonuclease analysis. The restriction pattern of the isolate obtained from calves vaccinated with vaccine D differs significantly from that of the original vaccine, whereas the reactivated virus from calves given vaccine B conserved the general pattern of the original vaccine strain. For each reactivated virus in this experiment (B and D) as well as for the isolate obtained from calves vaccinated with a further MLV (vaccine C) in a previous trial, three calves were inoculated. No clinical signs of disease were detected in any of the inoculated calves during the observation period. When the nine calves were exposed 40 days later to challenge infection with virulent BHV-1, they remained healthy and no virus was isolated from their nasal swabbings. These results indicate that some BHV-1 vaccines considered in the project can establish latency in the vaccinated calves, however, the latency does not appear to interfere with the original properties of the vaccines in terms of safety and efficacy. PMID- 12135237 TI - Investigation of Bartonella infection in ixodid ticks from California. AB - A total of 1253 ixodid ticks (254 tick pools) collected between the end of 1995 and the spring of 1997 from six California counties (El Dorado, Los Angeles, Orange, Santa Cruz, Shasta and Sonoma) were examined for the presence of Bartonella DNA by PCR of the citrate synthase gene. Of 1,119 adult Ixodes pacificus ticks tested, 26 (11.6%) of 224 pools, each containing five ticks, were positive (minimum percentage of ticks harboring detectable Bartonella DNA, 2.3%). Bartonella PCR-positive ticks were identified in five counties but none of the ticks from Los Angeles County was positive. Among 47 nymphal I. pacificus ticks collected in Sonoma County, one (10%) positive pool out of 10 pools was identified (minimum percentage of ticks harboring detectable Bartonella DNA, 2.1%). Among the 54 Dermacentor occidentalis grouped in 12 pools from Orange County, one pool (8.3%) was PCR positive for Bartonella and similarly one pool (14.3%) was positive among the 30 Dermacentor variabilis ticks grouped in seven pools. None of the three D. occidentalis from El Dorado County were positive. None of the nine tick pools positive for Ehrlichia phagocytophila were positive for Bartonella. Following our previous findings of Bartonella PCR-positive adult I. pacificus ticks in central coastal California, this is the first preliminary report of the presence of Bartonella DNA in I. pacificus nymphs and in Dermacentor sp. ticks. Distribution of Bartonella among ixodid ticks appears widespread in California. PMID- 12135236 TI - Characterization of the first Bartonella henselae strain isolated from a cat in Italy. AB - Bartonella henselae has been identified and characterized for the first time in Italy. A strain, designed Pavia-1, was isolated from the blood of a cat whose owner developed cat scratch disease (CSD). Pavia-1 and two American B. henselae strains (Houston-1, ATCC 49882, type I and strain 269608, UC Davis, type II) were compared by whole-cell fatty analysis (CFA), sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) for protein profiles, Western immunoblotting (WB) for reactivity with polyclonal antibodies, polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP), type-specific 16S rRNA PCRs, and pulsed-field gel electrophoresis (PFGE). Bartonella clarridgeiae (ATCC 51734) was also included for comparison. Pavia-1 was identified as a B. henselae type I. PFGE allowed differentiation between B. clarridgeiae and B. henselae and furthermore, between all the B. henselae strains. The fingerprints of PFGE observed for Pavia-1 were distinct from those of B. henselae type II and also of Houston-1, suggesting that the two type I strains derived from two different clones. These results show the capability of B. henselae to develop genotypic variability between genetically related strains. PMID- 12135238 TI - Distribution of rabies virus in infected mice, vaccinated and submitted to P. acnes as immunomodulator. AB - The lethality and distribution of rabies virus were evaluated in swiss mice experimentally infected with street rabies virus, vaccinated and submitted to immunomodulation by P .acnes (formerly Corynebacterium parvum). The animals were sacrificed at different times,when the different tissues were collected and submitted to fluorescent antibody test (FAT) and mouse inoculation test (MIT). The group submitted to vaccination and P. acnes treatment presented a percentage of survival superior to that observed in infected mice only treated with P. acnes. Control infected animals had the lowest survival rates. The distribution of rabies virus in spleen of infected mice, vaccinated and submitted to P. acnes was superior to that verified in infected mice not treated with P.acnes. The increased survival correlated with the distribution of rabies virus in lymphoid tissues, could be interpreted as the consequence of P. acnes activity on macrophages. The results suggest the role of macrophages against rabies virus infection in mice and the importance of vaccination in the post expositive treatment of rabies. PMID- 12135239 TI - Escherichia coli 'O' group serological responses and clinical correlations in epidemic HUS patients. AB - This first comprehensive serological analysis of an haemolytic uraemic syndrome (HUS) outbreak in which a wide range of 'O' group Escherichia coli antibody responses in patients and controls provided a unique insight into the epidemiology of such epidemics. Possible answers to clinical aspects related to severity of disease and complications were revealed. A microagglutination assay was used to examine E. coli 'O' group serological responses in 49 serum samples of 21 children hospitalised with HUS and 14 single samples from contemporaneous age-matched controls. A total of 51 O serogroup strains were used, including those reported to be associated with cases of HUS, with six isolates from patients associated with the Adelaide outbreak, environmental verocytotoxi genic/shiga-toxin producing E. coli (VTEC/STEC) strains and common human commensal strains. Amongst the 21 patients, there were 226 instances of seroreactivity (titre > or = 100) against 34 E. coli serogroups while six instances of seroreactivity against four serogroups occurred in controls. There were 128 instances in patients and one instance in controls in which titres > or = 400 were observed. All 21 patients were seroreactive (titre > or = 100 and <400) to one or more of the 17 HUS-associated serogroups included in the study. Titres ranged from 100 to 6,400, some of the highest in three patients were against O157, whose faeces yielded only EHEC O111, and only one developed O111 antibody. Mixed infection was demonstrated serologically by microagglutination (confirmed by western blot) and was consistent with the multiple serogroups of VTEC found in the mettwurst incriminated as the source, and suggests further strains (not found in the source or in patients' faeces) were probably involved. In HUS-associated EHEC infection, multiple strain infection may be the rule rather than the exception. Analysis of 34 of the 51 serogroup antibody responses in the HUS patients revealed clues to possible relationships with clinical severity and complications. Patients with severe renal failure tended to develop antibodies to a larger number of serogroups than those with moderate or mild impairment. The same was true for central nervous system complications. Other associations were observed. While VTEC O157 remains an important causal serogroup in HUS, non-O157 serogroups also appear to play a significant role and therefore the latter should always be sought in all future HUS cases as new insights into pathogenicity may be discovered. This study indicates that co-infection with different VTEC serogroups may affect clinical outcome. PMID- 12135240 TI - Quality assessment of pacemaker implantations in Denmark. AB - AIMS: Quality assessment of therapeutic procedures is essential to insure a cost effective health care system. Pacemaker implantation is a common procedure with more than 500,000 implantations world-wide per year, but the general complication rate is not well described. We studied procedure related complications for all implantations performed in an entire nation over a 3-year period. METHODS AND RESULTS: A prospective study of complications related to 99% of the 5648 primary pacemaker implantations performed in the 12 Danish pacemaker centres in 1997-1999 was carried out. Overall 76% of the patients received a physiological pacemaker system and 91% received the optimal pacing mode according to international guidelines. Perioperative complications requiring reoperation were: haematoma 0.3%, atrial lead related 1.9%, ventricular lead related 1.7%. Late complications requiring reoperation were: infection 02%, atrial lead related 13%, ventricular lead related 1.2%. The complication rate decreased over the study period, but overall the complication rate was higher than expected and showed considerable variation between centres. CONCLUSIONS: Our results demonstrate that sensitive data such as complications related to pacemaker implantations can be collected on a national basis. We suggest that a reoperation rate higher than 3% for atrial as well as ventricular pacing electrodes in the individual implanting centre should cause the centre to evaluate carefully the procedure as well as the performance of the individual implanter. PMID- 12135241 TI - Long-term survival after permanent pacemaker implantation: analysis of predictors for increased mortality. AB - AIMS: To determine long-term time-related survival and evaluate risk factors for increased mortality in patients following their first permanent pacemaker implantation. METHODS AND RESULTS: Analysis of patient records from implant to follow-up. Patient-specific time-lines were constructed to date of last review or death. Observed survival was estimated by event-free analysis using the Kaplan Meier method. Expected survival was derived from age- and gender-matched cohorts. Risk factors for mortality were sought using the multivariate Cox proportional hazards method and risk ratios estimated. Eight hundred and thirty-three patients underwent implantation of their first permanent pacemaker from April 1992 to January 1994, and were locally followed up. Survival data were available for 803 (96.4%) patients (median age, 77.3 years [5th to 95th centile range: 53.5 to 89 5 years]) and follow-up was complete in 94.8%. At implant. dual-chamber systems were implanted in 443 (55.1%). single-chamber ventricular systems in 321 (40.0%), and single-chamber-atrial systems in 39 (4.9%). Observed survival after implantation was significantly worse than expected (P<0.001). Independent predictors of increased mortality were: age at implant (risk ratio [RR] 1.06: 95% confidence interval [CI] 1.01 to 1.12). VVI pacing mode (RR 1.64; 95% CI 1.34 to 1.93), cardiomyopathy (RR 5.86; 95% CI 4.86 to 6.86), male gender (RR 1.27; 95% CI 1.22 to 1 32) and valvular heart disease (RR 2.01: 95% CI 1.98 to 2.04). CONCLUSIONS: At the end of follow-up, mortality was much higher than expected. In this typical pacemaker population. age at implant and VVI pacing mode were independently associated with increased mortality with accompanying heart disease having the greatest individual impact. PMID- 12135242 TI - High impedance leads and safety margin. Electrical considerations based on a simplified expression of the 'paradigm'. AB - For physicians who are not familiar with the electrical basis of cardiac stimulation, high impedance leads previously (10 years ago) considered as bad electrodes (high voltage) are now considered as the 'epitome' of lead technology (low energy drain): clearly impedance is not a good parameter for characterizing the qualities of a pacing lead. Using a simplified approach to the electrostimulation 'paradigm', it is easy to establish that modern high impedance leads are in fact high current density leads and high efficiency leads (better description). It is also possible to establish that routine programming of the safety margin at 100% above threshold parameters is associated with a decrease in the penetration of the electric field according to the reduction of the cathode surface area. For safety and energy saving, a small tip electrode could be combined with a low polarization surface treatment and a reduction in fibrosis development between electrode and myocardium. PMID- 12135243 TI - Single-lead VDD-pacing system incorporating high impedance stimulation: a multicentre study. AB - AIM: The purpose of this study was to evaluate the performance of a new VDD pacing system incorporating a high impedance, single-pass VDD lead. The new lead is a bipolar, steroid-eluting, high impedance lead with a full-ring atrial dipole. METHODS AND RESULTS: The system was implanted in 46 patients with high degree atrioventricular (AV) block. Patients were followed at pre-discharge, 6 weeks, and 3 months. The mean measured P-wave amplitude was stable, with values between 1.18 and 1.43 mV. Atrial sensing was reliable during short-term evaluation at rest and in the sitting position, with AV-synchronous stimulation between 98.79 +/- 6.90% and 99.73 +/- 1.47%. Holter recordings after 6 weeks demonstrated AV-synchronous stimulation in 99.57 1.03% of all P-waves. Lead impedance was stable during follow-up, with mean values between 1000 and 1167 Q. Mean ventricular pacing thresholds (at 0.5 ms) were 0.47 V at implant, 0.49 V at pre-discharge, 0.74 V at 6 weeks, and 0.72 V at 3 months. R-wave amplitude remained stable between 14.9 and 16.7 mV during follow-up. CONCLUSION: This new single-pass VDD lead system provided reliable atrial sensing and stable high impedance stimulation during a 3-month follow-up period. PMID- 12135244 TI - Pre-implant determinants of adequate long-term function of single lead VDD pacemakers. AB - AIMS: An inherent limitation of single lead VDD pacing is the inability to stimulate the atria. Reprogramming and upgrading the pacemaker system may be required when sinus node dysfunction, atrial undersensing, or atrial fibrillation develop. We evaluated whether routine clinical information is sufficient to select patients to benefit in long-term from VDD pacing. METHODS AND RESULTS: We collected data on 12-lead and monitored electrocardiograms and routine clinical information at implantation of a VDD pacing system in 350 consecutive patients with grade II or III atrioventricular conduction block. The age at implantation was 74.5 +/- 8.0 years, and the follow-up lasted 1.5 +/- 1.5 years. The cumulative maintenance of VDD pacing mode was 91.%. Loss of VDD mode was due to permanent atrial fibrillation in 16 (4.6%), sinus node dysfunction in six (1.7%). atrial undersensing in 11 (3.1%). Chronic atrial fibrillation developed in 23% of patients who had heart enlargement in chest x-ray and a history of paroxysmal atrial fibrillation or flutter. A criterion of normal sinus rate at implantation sufficiently predicted adequate sinus node function. Poor atrial sensing was not predicted by pre-implant characteristics. CONCLUSIONS: According to our data, adequate sinus-driven atrial rate and no history of paroxysmal atrial fibrillation and cardiac enlargement predict maintenance of the VDD pacing mode in elderly patients treated for heart block. Routine information available at implantation is sufficient to guide acceptance of single lead VDD pacing therapy. PMID- 12135245 TI - Prevalence of hypotensive disorders in older patients with a pacemaker in situ who attend the Accident and Emergency Department because of falls or syncope. AB - AIMS: To ascertain the proportion of adults with a pacemaker in situ attending the Accident and Emergency Department because of syncope or unexplained falls and the cause of index symptoms in these patients, including the prevalence of hypotensive syndromes. METHODS AND RESULTS: Patients presenting to the Accident and Emergency Department with unexplained syncope or non-accidental falls, who had a pacemaker in situ, were studied. Eligible patients had cardiovascular assessment (morning orthostatic blood pressure measurement, heart rate and BP measurements during carotid sinus stimulation (supine and upright), head-up tilt at 70 degrees for 40 min), assessment of haemodynamics during fixed mode pacing and gait and balance assessment. Of 5863 patients screened, 13.5% had unexplained syncope or a non-accidental fall; of these only 3% (26 patients) had pacemakers in situ. Of 18 study patients (82 +/- 8 years), 10 were female. Sixteen had a hypotensive diagnosis. Seven had more than one attributable hypotensive diagnosis. Five of 13 with vasodepressor carotid sinus syndrome had no previous diagnosis of carotid sinus hypersensitivity. No patients had vasovagal syncope induced during passive head-up tilt testing. CONCLUSION: It is rare for patients who attend the Accident and Emergency Department because of syncope or unexplained falls to have a pacemaker in situ. In those who do, hypotensive disorders are a common finding. PMID- 12135246 TI - Incessant atrioventricular dissociation due to far-field QRS oversensing and recurrent mode switch in a dual chamber pacemaker. AB - We report the case of a repetitive and incessant activation of mode switch in a dual chamber pacemaker because of the inappropriate sensing by the atrial lead of far-fields signals from the ventricular evoked response. The incidence, consequences and prevention of the oversensing of far-field QRS complexes are discussed. PMID- 12135247 TI - Stent implantation for the treatment of superior vena cava syndrome related to pacemaker leads. AB - This is a case-report of two patients with superior vena cava syndrome related to pacemaker leads. Both patients were treated successfully using intravenous stenting. PMID- 12135248 TI - Lesson of the issue: Transection, migration and recovery of a peel-away introducer sheath during permanent pacemaker implantation. PMID- 12135249 TI - Implantable devices to treat atrial fibrillation: real prospects or just new gimmicks? PMID- 12135250 TI - Prevention of atrial fibrillation by inter-atrial septum pacing guided by electrophysiological testing, in patients with delayed interatrial conduction. AB - AIMS: Interatrial septum (IAS) pacing seems efficient in synchronizing atrial depolarization in patients (pts) with delayed inter-atrial conduction, but its clinical role in preventing atrial tachyarrhythmias is still debated. This study was conducted in order to evaluate the clinical efficacy of IAS pacing guided by pace mapping of the IAS, as an alternative treatment modality in pts with drug refractory paroxysmal atrial fibrillation (PAF). METHODS AND RESULTS: We evaluated 29 pts (13 male, 16 female, 60 +/- 11 years), with drug refractory PAF, normal sinus node function and prolonged inter-atrial conduction time (P wave 142 +/- 10 ms). Multipolar catheters were inserted and the electrograms from the high right atrium (HRA) and proximal, middle and distal coronary sinus (CS) were recorded. The IAS was paced from multiple sites. The site of IAS where the timing between HRA and distal CS was <20 ms was considered the most suitable for synchronizing the atria. This site was found to be superior to the CS os. near the fossa ovalis in all pts. An active fixation atrial lead was positioned at this site and a standard lead was placed in the right ventricle. During IAS pacing, the P wave duration decreased significantly to 107 +/- 15 ms (P<0.001). At implant, the atrial sensing was 2.3 +/- 0.7 mV, the atrial pacing threshold was 0.95 +/- 0.15 V (0.5 ms) and the impedance was 760 +/- 80 Ohm. We evaluated the pts during four periods of 3 months duration each. The first period (control) was before pacemaker implantation, while the pts were under antiarrhythmic treatment. During the subsequent two periods, we evaluated the clinical efficacy of IAS pacing to prevent PAF recurrences, in AAT (75 bpm) and AAIR (75-140 bpm) mode, with random selection of the order and after discontinuation of antiarrhythmic treatment. During the fourth period, the same AAIR mode was assessed, but antiarrhythmic drugs were also administered. We compared the arrhythmia free interval among the four periods. The proportion of atrial paced beats in AAIR pacing mode plus antiarrhythmics was significantly higher compared with the drug-free period in AAIR mode (57 +/- 9% and 49 +/- 9% respectively, P=0017) and with AAT pacing mode (44 +/- 10%,(, P<0.001). In AAT mode, the arrhythmia free interval was 24.2 +/- 5.1 days, while it was 26.2 +/- 5.7 days in AAIR mode. These intervals did not differ significantly from the pre-implantation period (24.1 +/- 6.3 days). The arrhythmia free interval in AAIR pacing in combination with antiarrhythmic drug therapy was 38.7 +/- 8.1 days and this was significantly longer than the previous periods (P<0.05). CONCLUSION: Atrial septal pacing in combination with antiarrhythmic drug therapy reduced the incidence of PAF in pts with prolonged inter-atrial conduction times. Pace mapping of the IAS is an attractive technique to assess the shortest atrial activation time between HRA and distal CS. Whether placement of the atrial lead based on the shortest HRA--distal CS time is the best place in the IAS to prevent PAF still remains to be proven. PMID- 12135251 TI - Hypertension and arrhythmia: blood pressure control and beyond. AB - Arrhythmias are common problems in hypertensive patients. The presence and complexity of both supraventricular and ventricular arrhythmias may influence morbidity, mortality, as well as the quality of life of patients. Diastolic dysfunction of the left ventricle, left atrial size and function, and left ventricular hypertrophy have been suggested as the underlying risk factors for supraventricular and ventricular arrhythmias in hypertensives. Recently, several non-invasive electrocardiographic parameters have been defined and widely investigated to identify the hypertensive patient at risk for the development of arrhythmias. These parameters include signal averaged analysis of P wave, QT interval dispersion, heart rate variability, ventricular late potentials and T wave morphology analysis. The aim of this review was to evaluate the relationships between high blood pressure, ventricular and supraventricular arrhythmias, relevant non-invasive cardiac parameters for risk assessment in hypertensive patients and the effects of blood pressure control. PMID- 12135252 TI - Morphology of inter-atrial conduction routes in patients with atrial fibrillation. AB - BACKGROUND: Inter-atrial conduction disturbance is associated with the presence of atrial fibrillation (AF), although little is known about the anatomy of the inter-atrial connections. METHODS AND RESULTS: Twenty-seven hearts from in hospital deaths were examined and stratified in two groups with regard to their history of AF. Measurements of atrial weight were performed after excising the atria at the level of the atrioventricular valve plane and separation from the inter-atrial septum (IAS). In addition, in 15 of 27 hearts (seven AF, eight non AF) the IAS was sliced into 10 microm thick parallel histological sections at intervals of 1 mm starting at the valve plane and ending at the atrial roof. The sections were stained with van Gieson's stain. The variable morphology of the anterior and posterior inter-atrial connective muscle bundles is described. The total number of inter-atrial connections varied from 1 to 5. The anterior route (Bachmann's bundle) was not found in seven of 15 specimens. CONCLUSION: Inter atrial connections are characterized by substantial variability in morphology and in the distribution of bundles. This may account for part of the variable susceptibility to AF. PMID- 12135254 TI - Episodes of atrial fibrillation and atrial vulnerability after successful radiofrequency catheter ablation in patients with Wolff-Parkinson-White syndrome. AB - Episodes of atrial fibrillation occur in patients with WPW syndrome but frequently disappear after successful radiofrequency ablation. AIMS: To analyze the incidence of atrial fibrillation before and after successful ablation and the presence of increased atrial vulnerability. METHODS AND RESULTS: Fifty-four of 183 WPW patients had at least one documented episode of atrial fibrillation before ablation. During a follow-up of 24 +/- 12 months 13/54 patients (24%) experienced atrial fibrillation. At baseline, the patients with atrial fibrillation were more often men (74%) vs 53%, P=0.007), were older (45 +/- 15 vs 34 +/- 16 years, P=0.0001), more often had pre-excitation during sinus rhythm (87% vs, 73%, P=0.04) and had increased atrial vulnerability (41%, vs 18%, P<0.001). Only patients with atrial fibrillation before, developed atrial fibrillation after ablation. The 13 (of 54) patients who relapsed were also older (53 +/- 13 vs 42 +/- 15 years, P= 0.03), had increased atrial vulnerability at baseline (77% vs 29%, P=0.002), and were more symptomatic, (13 +/- 21 vs 1 +/- 3 arrhythmia attacks/month, P=0001). No patient without atrial fibrillation before ablation developed atrial fibrillation after treatment. CONCLUSIONS: The accessory pathway was important for the development of atrial fibrillation. Frequent tachycardias seem to promote an electrical remodelling and an increased atrial vulnerability to atrial fibrillation, whereas after successful ablation the majority of patients remain free of atrial fibrillation. PMID- 12135253 TI - Arrhythmia recurrences are rare when the site of radiofrequency ablation of the slow pathway is medial or anterior to the coronary sinus os. AB - AIMS: The site of successful ablation of the slow atrioventricular (AV) nodal pathway may be located in the posteroseptal or midseptal area. We have previously shown that the site of successful radiofrequency (RF) ablation of the slow pathway, rather than residual slow pathway conduction correlates with AV nodal re entrant tachycardia (AVNRT) recurrences, with more recurrences noted in inferoposterior (to the coronary sinus os) locations. Accordingly, we have since modified our approach, and in a consecutive series of 105 patients we have performed slow pathway RF ablation exclusively at medial or anterior locations, with the objective of prospectively examining the recurrence rate of AVNRT incurred with this approach. METHODS AND RESULTS: The study included 40 men and 65 women, aged 42 +/- 18 years, having RF ablation for symptomatic AVNRT exclusively in anterior to the coronary sinus os locations. A combined anatomical and electrophysiological approach to slow pathway ablation was employed. This series of patients was compared with the previous series of 55 patients (historical group) with AVNRT undergoing RF ablation at both inferoposterior and anteromedial locations. The mean cycle length of the induced AVNRT was 329 +/- 48 ms. RF ablation was successful in all patients (100%). A mean of 7 +/- 6 lesions were applied. Persistent jump or echo beats were noted in 48 patients (46%). The procedure lasted for 2.1 +/- 1.0 h. Fluoroscopy time was 23 +/- 14 min. Procedures were complicated by heart block in two patients (1.9%). Over 26 +/- 19 months, there has been only one recurrence of AVNRT (1%). The historical group had similar age (37 +/- 18 years), gender (17 men/38 women), AVNRT cycle length (340 +/- 60 ms), number of RF lesions (9 +/- 6), or residual slow pathway conduction (42%), but longer fluoroscopy time (41 +/- 25 min) and procedure duration (4 +/- 1 h), and a significantly higher recurrence rate (seven patients/13%) (P=0.004) at a much shorter follow-up period of 12 +/- 8 months. CONCLUSION: AVNRT recurrences are rare (1%) when slow pathway RF ablation is performed in medial or anterior locations at the tricuspid annulus, rather than in inferoposterior sites, whereby a higher (13%) recurrence rate has been previously noted. PMID- 12135255 TI - Dual-loop intra-atrial re-entry tachycardia in a patient with ischaemic cardiomyopathy. AB - A 65-year-old man with ischaemic cardiomyopathy (three prior coronary artery bypass surgery procedures), underwent catheter ablation for recurrent atrial flutter. Electrophysiological study initially revealed clockwise, tricuspid annulus/inferior vena cava isthmus dependent, atrial flutter. During radiofrequency energy ablation atrial flutter changed into a different atrial tachycardia without change in cycle length or interruption of the tachycardia. The new tachycardia was a right atrial free wall re-entry tachycardia. Thus the two atrial tachycardias formed a dual-loop ('figure-of-eight') re-entry circuit, possibly due to atrial scar tissue from multiple cardiac surgery procedures. PMID- 12135256 TI - Implantable loop recorder undersensing mimicking complete heart block. AB - The newer implantable loop recorders recently introduced (Reveal Plus. Medtronic Inc, Minneapolis, MN, U.S.A.). equipped with auto activation capabilities, are expected to expand our diagnostic abilities in the investigation of syncopal episodes. The case presented here is that of a man with pre-syncopal and syncopal episodes, without organic heart disease, in whom the device was auto-activated due to undersensing and the recorded strip imitated complete heart block. Increased attention should be given by the interpreter of the device's recordings to avoid false positive diagnoses that might lead to serious therapeutic actions. PMID- 12135257 TI - Restoration of malaria control in the Madagascar highlands by DDT spraying. PMID- 12135258 TI - Maternal transmission efficiency of Wolbachia superinfections in Aedes albopictus populations in Thailand. AB - We examined the transmission efficiency of 2 strains of Wolbachia bacteria that cause cytoplasmic incompatibility in field populations of Aedes albopictus by polymerase chain reaction assay. We found mainland and island populations throughout Thailand to be superinfected with group A and B bacteria. Of 320 Wolbachia-positive adult mosquitoes, 97.5% were infected with both groups. Single infected individuals of each Wolbachia group were encountered in nearly equal numbers. We screened 550 offspring from 80 field-collected mothers and found the transmission efficiency of group A Wolbachia to be 96.7% and that of group B Wolbachia to be 99.6%. Mothers that did not transmit both Wolbachia infections to all of their offspring were significantly larger in size than those with perfect transmission fidelity. We discuss our findings in relation to the prospects of the use of Wolbachia as a gene-driving mechanism. PMID- 12135259 TI - Field prevalence of Wolbachia in the mosquito vector Aedes albopictus. AB - The endosymbiotic bacteria in the genus Wolbachia have been proposed as a potential candidate to deliver pathogen-blocking genes into natural populations of medically important insects. The successful application of Wolbachia in insect vector control depends on the ability of the agent to successfully invade and maintain itself at high frequency under field conditions. Here, we evaluated the prevalence of Wolbachia infections in a field population of the Wolbachia superinfected mosquito Aedes albopictus. A field prevalence of 100% (n = 1,016) was found in a single population in eastern Thailand via polymerase chain reaction (PCR) testing of Wolbachia both from individual parent females and their corresponding F1 offspring. This is the first report of accurate Wolbachia prevalence in a field population of an insect disease vector. The prevalence of superinfection was estimated to be 99.41%. All single-infected individual mosquitoes (n = 6) were found to harbor group A Wolbachia. For this particular population, none was found to be single-infected with group B Wolbachia. Our results also show that PCR testing of field materials alone without checking F1 offspring overestimated the natural prevalence of single infection. Thus, the confirmation of infection status by means of F1 offspring was critical to the accurate estimates of Wolbachia prevalence under field conditions. PMID- 12135260 TI - Outbreak of vivax malaria in areas adjacent to the demilitarized zone, South Korea, 1998. AB - Malaria had been eradicated in the Republic of Korea (South Korea) by the late 1970s. In 1993, a soldier was infected with Plasmodium vivax malaria in the Demilitarized Zone (DMZ; the border area between North and South Korea), and since then, the number of cases has been steadily increasing year after year. In 1998, 3,932 vivax malaria cases were microscopically confirmed, affecting 2,784 (70.8%) soldiers (including discharged soldiers) and 1,148 (29.2%) civilians. These cases occurred throughout the year, peaking in July (30.1%) and August (30.5%). Most of the patients were infected in areas in or near the DMZ. Taking into consideration entomologic, socioecologic, and epidemiologic factors, it is postulated that there has been an epidemic of malaria in North Korea since 1993, with the number of cases increasing yearly; the continuous infiltration across the DMZ from North Korea of infected female mosquitoes of the vector species Anopheles sinensis resulted in an outbreak of vivax malaria in the DMZ of South Korea. PMID- 12135261 TI - Emergence of a new neotropical malaria vector facilitated by human migration and changes in land use. AB - In a region of northeastern Amazonia, we find a species previously of minor importance, Anopheles marajoara, to be the principal malaria vector. In a total of five collections during 1996-97 in three replicated sites near the city of Macapa, Amapa state, this species occurs in much greater abundance compared with the presumed vector Anopheles darlingi. Also, a significantly higher proportion of An. marajoara is infected with malaria parasites, determined by the ELISA technique. This appears to be the result of increased abundance of An. marajoara due to alterations in land use, invasion of its primary breeding sites by human immigrants, and its anthropophilic behavior. This discovery highlights one of the challenges of Neotropical malaria control, namely that the targeting of specific vectors may be complicated by a changing mosaic of different locally important vectors and their interactions with human populations. PMID- 12135262 TI - Impact of the malaria control campaign (1993-1998) in the highlands of Madagascar: parasitological and entomological data. AB - Malaria transmission in the central highlands of Madagascar was interrupted in the 1960s by a national control program that used DDT indoor spraying and mass treatment with chloroquine. At the end of the 1980s in this region, epidemic malaria reappeared. Italian health authorities provided technical assistance to the National Malaria Control Program since the beginning of the resurgence of malaria in the central highlands. Yearly residual house spraying performed for 5 years (1993-1998) and the availability of antimalarial drugs reduced malaria transmission to very low levels, with improvement in parasitologic and entomologic indexes. A significant reduction of malaria prevalence was observed in the villages located at altitudes of 1,000-1,500 m, corresponding to the stratum of unstable malaria that was the main target of the antivector interventions. A significant reduction of malaria prevalence was also observed in the villages located at altitudes of 900-1,000 m, where malaria transmission is stable. The main vector Anopheles funestus was dramatically reduced in abundance and distribution in the sprayed areas. PMID- 12135263 TI - Combined ultrasound and serologic screening for hepatic alveolar echinococcosis in central China. AB - Alveolar echinococcosis (AE), caused by Echinococcus multilocularis, is a zoonotic helminthic disease that can mimic malignancy. In the 1970s, foci of the disease were found in central China. The aim of the present study was to estimate the prevalence of AE in humans in 2 districts of south Gansu Province, China, by use of ultrasound and Echinococcus serology. After answering an epidemiological questionnaire, 2,482 volunteers from 28 villages underwent ultrasound. Serology via enzyme-linked immunosorbent assay for antibody activity was performed on whole blood collected on filter paper in all subjects; on serum from subjects with an abnormal ultrasound image; and on randomly chosen subjects that either had no lesions or had atypical lesions. At least one (25.3%) abnormal ultrasound image was observed in 630 of the subjects screened. A typical lesion of progressive AE was found in 84 subjects (3.4%). Serologies were positive in 77 (96%) of 80 of patients who had lesions typical of progressive AE. Ultrasound is useful for screening for AE in endemic regions. PMID- 12135264 TI - Potential long-term toxicity of repeated orally administered doses of artemether in rats. AB - Artemether, an efficacious antimalarial drug, effectively prevents patent schistosome infections and morbidity, as established in laboratory models and in clinical trials. In view of concern about the potential long-term toxicity, rats were treated orally with 80 mg/kg artemether once every 2 weeks for 5 months. After the final treatment, routine blood test results were normal except for reversible reductions of reticulocyte counts and reversible increases in hemoglobin levels. Liver and kidney function and histopathological examination showed no differences between treated and untreated rats. Administration of 400 mg/kg artemether resulted in transient focal vesicle degeneration of the liver or slight damage to the proprius layer lamina of intestinal villi. No damage to the central nervous system tissues, including cerebrum, cerebellum, midbrain, thalamus, pons, medulla oblongata, and spinal cord, was seen at either concentration. There were no alterations in electrocardiograms during the 6-month treatment period. We conclude that 80 mg/kg artemether administered once every 2 weeks is safe, and doses of 400 mg/kg do not result in evidence of neurotoxicology. PMID- 12135265 TI - Assessment of dengue risk in relief workers in Puerto Rico after Hurricane Georges, 1998. AB - Health risk assessment is important in the safe deployment of workers to tropical areas. We monitored dengue incidence in 204 of 222 North American relief workers visiting Puerto Rico after Hurricane Georges and during a dengue epidemic in 1998. We recorded information regarding participants' living conditions and any illness they experienced from arrival to 2 weeks after their departure. Virus isolation, polymerase chain reaction, and serological tests for anti-dengue immunoglobulin (Ig) M and IgG antibodies were used to diagnose dengue infection by means of departure and follow-up serum specimens. Among respondents, 82% (164 of 199) reported mosquito bites, 97% (156 of 161) reported having insect repellent available, and 41% (79 of 195) reported using repellent every day. Twelve participants reported a mild denguelike illness. No participants had laboratory evidence of dengue infection after 1.8 person-years of assessable exposure to areas with dengue transmission (upper 95% confidence limit of 1.67 cases per person-year). The risk of acquiring dengue among relief workers in this study appears low, possibly as a result of protective factors. Travelers to dengue-endemic areas should continue to be advised to protect themselves against mosquito bites. PMID- 12135266 TI - Control of aedes vectors of dengue in three provinces of Vietnam by use of Mesocyclops (Copepoda) and community-based methods validated by entomologic, clinical, and serological surveillance. AB - We describe remarkable success in controlling dengue vectors, Aedes aegypti (L.) and Aedes albopictus (Skuse), in 6 communes with 11,675 households and 49,647 people in the northern provinces of Haiphong, Hung Yen, and Nam Dinh in Vietnam. The communes were selected for high-frequency use of large outdoor concrete tanks and wells. These were found to be the source of 49.6-98.4% of Ae. aegypti larvae, which were amenable to treatment with local Mesocyclops, mainly M. woutersi Van der Velde, M. aspericornis (Daday) and M. thermocyclopoides Harada. Knowledge, attitude, and practice surveys were performed to determine whether the communities viewed dengue and dengue hemorrhagic fever as a serious health threat; to determine their knowledge of the etiology, attitudes, and practices regarding control methods including Mesocyclops; and to determine their receptivity to various information methods. On the basis of the knowledge, attitude, and practice data, the community-based dengue control program comprised a system of local leaders, health volunteer teachers, and schoolchildren, supported by health professionals. Recycling of discards for economic gain was enhanced, where appropriate, and this, plus 37 clean-up campaigns, removed small containers unsuitable for Mesocyclops treatment. A previously successful eradication at Phan Boi village (Hung Yen province) was extended to 7 other villages forming Di Su commune (1,750 households) in the current study. Complete control was also achieved in Nghia Hiep (Hung Yen province) and in Xuan Phong (Nam Dinh province); control efficacy was > or = 99.7% in the other 3 communes (Lac Vien in Haiphong, Nghia Dong, and Xuan Kien in Nam Dinh). Although tanks and wells were the key container types of Ae. aegypti productivity, discarded materials were the source of 51% of the standing crop of Ae. albopictus. Aedes albopictus larvae were eliminated from the 3 Nam Dinh communes, and 86-98% control was achieved in the other 3 communes. Variable dengue attack rates made the clinical and serological comparison of control and untreated communes problematic, but these data indicate that clinical surveillance by itself is inadequate to monitor dengue transmission. PMID- 12135267 TI - Short report: socioeconomic and seasonal variations of Helicobacter pylori infection in patients in Venezuela. AB - Infection by Helicobacter pylori is recognized as a risk factor for gastric cancer and peptic ulcer disease. Venezuela has regions with different gastric cancer risks; the Andean region has the highest gastric cancer mortality in the country. We performed a cross-sectional study on 357 patients who underwent endoscopy attending 2 private (n = 76) and one public hospital in Caracas, Venezuela (n = 215), and one public hospital in the Andes (n = 66) to determine H. pylori infection (by a rapid biopsy urease test and histology). The proportion of infected patients in Caracas was significantly higher in public hospitals (72%) than in private hospitals (46%; P = 0.00001), and there was no significant variation the Andes and Caracas (P = 0.7001). When analyzing the data from the public hospital in Caracas, we found that the frequency of infected patients was significantly higher during the rain (96%) than during the dry months (70%, P = 0.00000001). Differences in prevalence of infection in symptomatic patients was not related to the risk of gastric cancer but to socioeconomic differences. Rain dependent factors that may be exacerbating the clinical activity of nonulcer dyspepsia in people infected with H. pylori deserve further study. PMID- 12135268 TI - Identification of virulence-associated antigens and plasmids in Rhodococcus equi from patients with acquired immune deficiency syndrome and prevalence of virulent R. equi in soil collected from domestic animal farms in Chiang Mai, Thailand. AB - The prevalence of virulent Rhodococcus equi in soil collected from 17 domestic animal farms (from 12 cattle, 1 pig, and 4 horse farms) and in 6 clinical specimens from patients with acquired immune deficiency syndrome (AIDS) in Chiang Mai, Thailand, was investigated. The isolates were tested for the presence of 15 17-kDa antigens (VapA) and a 20-kDa antigen (VapB) by immunoblotting and for the presence of virulence plasmid DNA. Rhodococcus equi was isolated from most soil samples (68 of 80) obtained from the 17 farms, with 2.0 x 10(2) to 6.0 x 10(5) colony-forming units per gram of soil. We detected VapA in none of the 537 isolates from the soil samples. In one isolate from a pig farm, both VapB and virulence plasmid DNA were detected. Of the 6 clinical isolates from patients with AIDS, however, 4 isolates contained both VapB and virulence plasmid DNA. The remaining 2 isolates were avirulent. PMID- 12135269 TI - HIV infection, malaria, and pregnancy: a prospective cohort study in Kigali, Rwanda. AB - In order to study the relation between human immunodeficiency virus (HIV) infection and malaria in women, during and after pregnancy, a prospective cohort study was initiated at the Centre Hospitalier de Kigali in Rwanda through routine voluntary and confidential HIV screening in antenatal clinics. At inclusion in the cohort of all HIV-positive and an equivalent number of HIV-negative pregnant women, between 21 and 28 weeks of gestation, sociodemographic characteristics and medical history during the current pregnancy were collected; screening for malaria (tick blood smear) and anemia and a CD4 lymphocyte count were systematically performed. Each woman enrolled had a monthly follow-up until 6 months after delivery. A clinic was implemented that was accessible and free of charge to every woman during the study period between scheduled visits. Malaria infection was systematically screened in case of fever or other compatible symptoms. The cohort included 228 HIV-positive and 229 HIV-negative women. At inclusion, malaria prevalence was 8.0% in HIV-positive women and 3.5% in HIV negative women (P < 0.04). Over the study period, the incidence of malaria was 6.2 per 100 women-months in the HIV-positive group and 3.5 in the HIV-negative group (relative risk [RR] = 1.7, 95% confidence interval [CI] = 1.4-2.3). The bulk of the difference occurred postpartum. The Kaplan-Meier 9-month probability of remaining free of malaria infection was 51.8% in HIV-positive women and 65.2% in HIV-negative women (P = 0.013). When taking account in the same multivariate model (including HIV infection, primiparity, CD4 lymphocytes, anemia, and education level), positive HIV serostatus remained the only factor significantly associated with malaria infection (RR = 1.4, CI = 1.1-1.6; P = 0.016). Our study prospectively documents the association between malaria and maternal HIV infection and highlights the increased risk of malaria occurrence in all HIV infected women. Strategies to reduce the malaria morbidity during pregnancy should be reinforced in areas of high HIV seroprevalence. PMID- 12135270 TI - Leptospirosis in Hawaii, 1974-1998: epidemiologic analysis of 353 laboratory confirmed cases. AB - The epidemiologic characterization of leptospirosis in the United States has been limited by difficulties associated with both case detection and confirmation. In addition, leptospirosis was eliminated from the list of National Notifiable Diseases in 1995. From 1974 until the cessation of national surveillance, Hawaii consistently had the highest reported annual incidence rate in the United States. From 1974 through 1998, 752 leptospirosis cases were reported in the State of Hawaii. Of these, 353 had exposures within the state and were laboratory confirmed. The mean annual incidence rate was 1.29 per 100,000. Cases were predominately male. Rates were highest in rural areas. Occupational exposures diminished over time while recreational exposures increased. This series represents the first large U.S. leptospirosis surveillance report since 1979. With leptospirosis recently being identified as a re-emerging zoonosis, continued national surveillance and case reporting should be reconsidered. PMID- 12135272 TI - Outbreak of tick-borne relapsing fever at the north rim of the Grand Canyon: evidence for effectiveness of preventive measures. AB - An outbreak of tick-borne relapsing fever (TBRF) originating at the North Rim of Grand Canyon National Park was investigated in 1990. To determine risk factors for the disease, almost 7,000 parties of visitors were surveyed; over half responded, representing > 10,000 people. Fifteen cases of confirmed or probable TBRF were identified in visitors and 2 in employees. All patients except one experienced symptoms after overnight stays in a group of cabins that had not been rodent-proofed after a TBRF outbreak in 1973 (relative risk for visitors [RR] 8.2, 95% confidence interval [CI] 1.1-62). Seven cases of TBRF were associated with a single cabin (RR 98, 95% CI 30-219). Structural flaws and rodent nests were common in the implicated cabins and rare in unaffected cabins. This investigation suggests that measures to rodent-proof cabins at sites where TBRF is endemic prevent reinfestation of cabins by infected rodents and tick vectors, thereby preventing the spread of disease in humans. PMID- 12135271 TI - Antibody responses to repetitive epitopes of the circumsporozoite protein, liver stage antigen-1, and merozoite surface protein-2 in infants residing in a Plasmodium falciparum-hyperendemic area of western Kenya. XIII. Asembo Bay Cohort Project. AB - The present study was initiated to characterize antibody responses to repetitive epitopes of the circumsporozoite protein (CSP), liver stage antigen-1 (LSA-1), and merozoite surface protein-2 (MSP-2) of Plasmodium falciparum in infants residing in a P. falciparum-hyperendemic area of western Kenya. In this study, development and maintenance of these antibody responses in 28 infants were studied longitudinally by use of monthly serum samples collected from birth to age 1 year. Mother plasma and infant umbilical cord plasma were also tested to assess the transplacental transfer of maternal antibodies. Results showed that antibodies passively transferred from mothers were detectable for CSP, LSA-1, and MSP-2 repeat epitopes. Infants were able to mount and maintain a strong antibody response against LSA-1 in their first year of life. Infants often responded to CSP repeats, but with a much lower antibody titer. Antibody responses in infants against Fc27 and 3D7 repeats of MSP-2 were low throughout their first year. In addition, 51 infants whose first detected infection occurred at > 4 months of age were selected to determine antibody responses to the antigens tested upon their first and second detected infections. Antibody responses to LSA-1 and, to a lesser degree, CSP increased in positivity rates and titer upon second infection. Antibody responses to Fc27-type and 3D7-type repeats of MSP-2 were low upon both infections. There was no association between maternally transferred anti-LSA-1, anti-CSP, or anti-MSP-2 antibodies and an infant's first detected infection. No significant correlation was found between an infant's antibody responses to the 4 antigen repetitive epitopes and protection against malarial parasitemia during the first year of life. PMID- 12135273 TI - Prevalence of Rickettsia conorii and Rickettsia typhi infections in the population of northern Greece. AB - Seroepidemical surveys concerning the prevalence of Rickettsia conorii and Rickettsia typhi have never been studied in northern Greece. We examined 1,584 sera samples from residents of northern Greece for the detection of antibodies to R. conorii and to R. typhi by means of immunofluorescence assay. In addition, we compared the prevalence of rickettsial infections among the demographic variables of sex, age, occupation, and area of residence. Antibodies to R. conorii and R. typhi were found in 125 (7.9%) and 31 (2.0%) of the examined subjects, respectively. The prevalence of antibodies to R. conorii correlated with increasing age and was statistically higher in men. Farmers had significantly higher prevalence of antibodies to both species of rickettsiae studied versus other professions. Residents of rural areas showed a statistically higher prevalence for R. conorii versus urban residents, although this difference was not demonstrated for R. typhi. We also detected differences in the prevalence of rickettsial infections among the different prefectures. Our data show the wide distribution of R. conorii in northern Greece and indicate the presence of R. typhi. PMID- 12135274 TI - Fourteen-year seroepidemiological study of zoonoses in a Greek village. AB - A seroepidemiological study carried out in a high-risk village in Crete in 1985 1987 and 1998 showed that although the awareness of the people concerning zoonoses had increased during this period, the situation did not improve: there was a significant increase of the spread of seroprevalence in time and space of Coxiella burnetii, Rickettsia typhi, Brucella sp., and Entamoeba histolytica. Toxoplasma gondii, Rickettsia conorii, Borrelia burgdorferi, Echinococcus granulosus, Leishmania sp., and Fasciola hepatica stayed at the same levels. This first study of Bartonella henselae in Crete showed that 15.9% of the children tested were seropositive. The results indicate that reservoirs and vectors of the pathogens studied are widespread in the environment, and the way of life of the people favors contact with them. Seven of 30 milk samples were positive for Brucella sp. by seminested polymerase chain reaction. PMID- 12135275 TI - Isolation of Coxiella burnetii by a centrifugation shell-vial assay from ticks collected in Cyprus: detection by nested polymerase chain reaction (PCR) and by PCR-restriction fragment length polymorphism analyses. AB - Ticks are the principal vectors and reservoirs of Coxiella burnetii. The identification of isolates is necessary for understanding the clinical diversity of Q fever in different geographic areas. This is the first report of isolation of C. burnetii from ticks by the shell-vial assay and by nested polymerase chain reaction (PCR) assay for the detection of this pathogen in ticks. Of 141 ticks collected in Cyprus (Rhipicephalus sanguineus and Hyalloma spp.), 10% were found to be infected with C. burnetii. Three ticks were positive by hemolymph test, and 11 triturated ticks were positive by nested PCR. Three isolates were obtained by the centrifugation shell-vial technique. Analysis by PCR, then restriction fragment length polymorphism showed that the 3 Cyprus isolates had identical restriction profiles to reference strains Nine Mile and Q212. The methods described are useful in studying the epidemiology and ecology of C. burnetii. PMID- 12135276 TI - Evaluation of antigens from various Leishmania species in a Western blot for diagnosis of American tegumentary leishmaniasis. AB - A Western blot method that uses antigens from culture promastigote forms of Leishmania (Viannia) braziliensis, L. (Leishmania) amazonensis, L. (Leishmania) tropica, and a trypanosomatid (strain 268T) isolated from naturally infected tomatoes was evaluated for laboratory diagnosis of American tegumentary leishmaniasis (ATL). Serum samples were obtained from 108 patients with ATL (group I), 23 chagasic patients (group II), 32 patients with other diseases (group III), and 78 healthy individuals (group IV). The overall analysis showed a sensitivity of 76.90%, 90.40%, 78.50%, and 87.90%, a specificity of 100%, 93.80%, 87.80%, and 77.10%, a positive predictive value of 100%, 94.00%, 89.50%, and 72.50%, a negative predictive value of 75.70%, 90.00%, 75.40%, and 90.20%, and a concordance coefficient kappa of 0.7358, 0.8400, 0.6491, and 0.6287 for L. (V.) braziliensis, L. (L.) amazonensis, L. (L.) tropica, and strain 268T antigens, respectively. The antigenic profile recognized by serum samples from patients with ATL and with Chagas' disease permits serologic distinction between these infections. PMID- 12135277 TI - Editorial: jennerian vaccination against West Nile virus. PMID- 12135278 TI - Short report: absence of protective neutralizng antibodies to West Nile virus in subjects following vaccination with Japanese encephalitis or dengue vaccines. AB - Protection of individuals against West Nile (WN) encephalitis is an emerging concern in the United States and Europe. We investigated whether immunization with licensed inactivated Japanese encephalitis (JE) vaccine or experimental live attenuated dengue vaccines resulted in induction of cross-neutralizing antibodies against WN virus. Protective neutralizing antibody titers to WN virus were not detected in any volunteer despite successful immunization to related flaviviruses. Vaccination against JE or dengue is unlikely to prevent WN virus infection but may still protect against disease. PMID- 12135279 TI - Molecular epidemiology of malaria in cameroon. IX. Characteristics of recrudescent and persistent Plasmodium falciparum infections after chloroquine or amodiaquine treatment in children. AB - In the absence of a firmly established gene responsible for chloroquine and amodiaquine resistance in Plasmodium falciparum, surveillance of resistance to these first-line drugs in Cameroon needs to be performed by in vivo or in vitro tests for drug resistance. These 2 methodological approaches to define drug resistance were shown to be complementary and concordant in a majority of cases at our study sites, but discordant results may be observed in a few cases, probably as a result of acquired immunity and low plasma drug levels. To further examine the nature of recrudescent and persistent parasitemia after treatment with chloroquine or amodiaquine, the clinical response of children aged < 5 years, presumably with insufficient immune response, was assessed, and the in vitro response of the corresponding isolates was determined if treatment or parasitological failure occurred. Genotyping of pretreatment and posttreatment isolates was performed by polymerase chain reaction to distinguish between recrudescence and reinfection. Plasma drug levels were measured at the time of therapeutic failure by high-performance liquid chromatography. All cases of therapeutic or parasitological failure observed on or before Day 14 were due to the persistence or recrudescence of the original parasite populations present before treatment, with or without selection and appearance of new populations. Most parasites were characterized by elevated 50% inhibitory concentrations for chloroquine and amodiaquine at the time of clinical or parasitological failure. In some children, recrudescence was explained by the absence of drug in the plasma. The simultaneous analysis of clinical and in vitro responses, plasma drug level measurement, and genotyping may yield results that may explain the reasons for therapeutic failure, help establish the threshold level for in vitro resistance, and provide a set of more accurate tools to describe the epidemiology of drug-resistant P. falciparum while awaiting for the identification of the chloroquine and amodiaquine resistance gene or genes. PMID- 12135280 TI - Family analysis of malaria infection in Dienga, Gabon. AB - Fifty children from 9 families were enrolled in a longitudinal study of 8 months to evaluate individual levels of Plasmodium falciparum density in blood during asymptomatic infections. Individual parasite densities were adjusted for age and date of blood intake. The arithmetic means of these adjusted parasite densities (MAPD) were not influenced by sickle cell trait nor by G6PD enzyme activity. On the contrary, family analysis revealed the presence of similar MAPD values according to the sibships. Moreover, sibships frequently infected with P. malariae exhibited the highest P. falciparum MAPDs. The difference in aggressiveness of malaria vectors between the northern and southern halves of the village did not explain the distribution of MAPD, nor did it explain the differences in mean frequency of P. malariae infection among the sibships. We conclude that the familial characteristic of susceptibility to both P. falciparum and P. malariae infections is more likely influenced by the host's genetic background than by differences in the levels of malaria transmission. PMID- 12135282 TI - Short report: differential evolution of immunoglobulin G1/G3 antibody responses to Plasmodium falciparum MSP1(19) over time in malaria-immune adult Senegalese patients. AB - This study examined the evolution of immunoglobulin (Ig) G1 and IgG3 antibodies against the asexual stage Plasmodium falciparum protein, MSP1(19), before and after a heavy malaria transmission period in clinically immune Senegalese subjects living under different epidemiological conditions. Plasma was tested for antibodies to a yeast-produced, recombinant PfMSP1(19) antigen (the Q-KNG allelic variant) that has previously been demonstrated to react with IgG1, IgG3, or both in the majority of these people. Anti-P. falciparum antibodies of the IgG1 and IgG3 subclasses, previously reported to be associated with protection, were shown to evolve independently one from another after the transmission period in both settings. These results suggest differential regulation of MSP1(19)-specific IgG1 and IgG3. The precise role of these antibody isotypes in maintaining malaria immunity remains to be determined. PMID- 12135281 TI - Relative bioavailability of artesunate and dihydroartemisinin: investigations in the isolated perfused rat liver and in healthy Caucasian volunteers. AB - The aim of this study was to evaluate the bioavailability of artesunate (ARTS) and dihydroartemisinin (DHA). When the single-pass isolated perfused rat liver model was used, the hepatic bioavailability of ARTS (39 microM) was 0.20 and the bioavailability of DHA in ARTS and DHA (35 microM) perfusions was 0.12 and 0.11, respectively. Thus, the combined bioavailability of ARTS and DHA in the ARTS perfusions (0.32) was three-fold higher than the bioavailability of DHA. In the human study, eight healthy volunteers were randomized to receive oral ARTS (135 mg; 352 micromol) or oral DHA (120 mg; 422 micromol). The area under the curve (AUC(0-6 hr)) of DHA following ARTS (2.9 micromol x hr/L) was higher than the AUC(0-6 hr) of DHA following DHA administration (1.2 micromol x hr/L; P < 0.005). The dose-corrected relative oral bioavailability of DHA for DHA compared with ARTS was 43%. Thus, the use of conventional oral dosage regimens of ARTS appears preferable to oral administration of DHA. PMID- 12135283 TI - Short report: hepatitis b infection and severe Plasmodium falciparum malaria in Vietnamese adults. AB - We investigated the prevalence of infection with hepatitis B virus among adult Vietnamese patients hospitalized for severe Plasmodiumfalciparum malaria. Sera from patients admitted with severe malaria in Ho Chi Minh City, Vietnam, between May 1991 and January 1996 were assayed for hepatitis B surface antigen (HB(s)Ag) by a commercial enzyme-linked immunosorbent assay kit. The overall prevalence of HB(s)Ag was 23.77% (77 of 324). This was higher than reported estimates of prevalence in the general catchment population for the study hospital (mean, 9.8%; range, 9-16%). No association was found between risk of death caused by severe malaria and HB(s)Ag. Patients admitted with cerebral malaria had a slightly greater risk of registering positive for HB(s)Ag (relative risk, 1.28; 95% confidence interval, 1.04-1.58) relative to other manifestations of severe malaria. Chronic infection with hepatitis B virus may be a risk factor for severe malaria. PMID- 12135284 TI - Low-dose liposomal amphotericin B in refractory Indian visceral leishmaniasis: a multicenter study. AB - In this randomized, double-blind, dose-ranging, multicenter trial, 84 patients with visceral leishmaniasis refractory to antimony therapy were administered liposomal amphotericin B (AmBisome) at cumulative doses of 3.75, 7.5, and 15.0 mg/kg for 5 consecutive days. Posttreatment apparent cure and definite cure were assessed at 2 weeks and 6 months after the end of therapy, respectively. Mild to moderate infusion-related fever and rigors were seen in 29 and 44% of patients, respectively. One patient each in the 3.75- and 7.5-mg groups had detectable parasites on splenic smear at posttreatment evaluation. At 6 months' follow-up, however, 2, 1, and 1 patients relapsed in the 3.75-, 7.5-, and 15.0-mg groups, resulting in definite cure rates of 89, 93, and 97%, respectively. There was no significant difference in the cure rates of the 3 groups. Low-dose liposomal amphotericin B given for 5 days can cure most patients with Indian kala-azar. PMID- 12135285 TI - Treatment of cutaneous leishmaniasis with a topical antileishmanial drug (WR279396): phase 2 pilot study. AB - We studied the efficacy of WR279396, a topical formulation of aminoglycosides that cures 100% of cutaneous leishmaniasis lesions in mice. We conducted what is to our knowledge the first controlled study of WR279396 therapy for clinical cutaneous leishmaniasis. A total of 45 Colombian soldiers, all men, were randomly assigned to treatment with WR279396 (33 patients) or placebo (12 patients). Each lesion was treated twice daily for 20 days. Lesions were measured at the end of therapy and at 45, 90, and 180 days after treatment began. A total of 17 (61%) of 28 assessable WR279396-treated patients were cured, and 5 (55%) of 9 assessable placebo-treated patients were cured (P = 0.9). For the 36 lesions treated with WR279396 that were cured, cure took a mean of 35 days, whereas for the 6 lesions that were cured in the group of patients receiving placebo, cure time took a mean of 56 days (P = 0.04). WR279396 is a nontoxic topical formulation that significantly accelerated cure time in patients with Leishmania panamensis cutaneous leishmaniasis. PMID- 12135286 TI - Random amplified polymorphic DNA technique for identification and differentiation of old world Leishmania species. AB - The random amplified polymorphic DNA technique may be used to explore parasite DNA polymorphisms. We assessed its applicability to identification of Old World Leishmania species. A set of 6 random decamer primers (Al, A4, A5, A7, A10, and A15) was applied to a panel of DNA from 57 representatives of different Old World Leishmania species. The amplification profiles allowed discrimination among species belonging to different taxonomic complexes. Two criteria were used to analyze the profiles: the presence of consistent amplicons at the same electrophoretic position for isolates of the same species, and the presence of distinct amplicons for isolates of different species. Three primers--Al, A7 and A10--rendered such products. PMID- 12135287 TI - Detection by polymerase chain reaction of Schistosoma mansoni DNA in human serum and feces. AB - A novel method for the detection of Schistosoma mansoni in human samples that is based on the amplification of a highly repeated DNA sequence has been developed. By use of simple DNA extraction techniques and a rapid 2-step polymerase chain reaction (PCR), it was possible to amplify S. mansoni DNA in human fecal and serum samples. The high sensitivity of the approach enabled the detection of the parasite DNA in fecal samples containing as few as 2.4 eggs per gram of feces, which makes it 10 times more sensitive than the Kato-Katz examination. A detection limit of I fg of Schistosoma sp. DNA was determined when pure DNA was used as PCR template. The amplification reaction showed to be specific giving no cross-reaction with DNA from other helminths. The PCR assay developed in this study may constitute a valuable alternative for the diagnosis of the Schistosoma sp. infection. PMID- 12135288 TI - Reactivity to bacterial, fungal, and parasite antigens in patients with lymphedema and elephantiasis. AB - Both secondary infections and antifilarial immunity are thought to play roles in the development and progression of lymphedema. To investigate this issue, immune responses to a panel of bacterial, fungal, and parasite antigens were examined for women with lymphedema and elephantiasis (n = 28) and for women with no clinical evidence of lymphatic dysfunction who were either microfilaremic (Mf+, n = 23) or microfilaria- and filarial antigen-negative (Ag-, n = 24). The prevalence and intensity of delayed-type hypersensitivity (DTH) responses was similar for most recall antigens; for individual antigens, lymphedema patients were significantly more likely to be reactive only to Proteus. Lymphedema patients with a history of three or more attacks of adenolymphangitis in the last 18 months showed increased DTH reactivity to Trichophyton. Proliferative responses to fungal and bacterial antigens were similar for all three groups; however, antigen-negative women, independent of disease status, mounted greater responses to filarial antigen. In contrast, lymphedema patients had higher levels of antifilarial specific IgG1, IgG2, and IgG3 and higher IgG responses to streptolysin O than either Ag- or Mf+ women. In persons with lymphatic filariasis, immune reactivity is influenced by disease status as well as infection status. PMID- 12135289 TI - Serological evidence for recent exposure to Taenia solium in Venezuelan Amerindians. AB - This study examined the seroprevalence and serum antibody isotype profile for Taenia solium cysticercosis in an Amerindian community in the Amazonas state of Venezuela. An antigen-trapping enzyme-linked immunosorbent assay (Ag-ELISA) was used to detect viable cysticercosis. Indirect ELISA (Ab-ELISA) and enzyme-linked immunoelectrotransfer blot (EITB) was performed by using antigens prepared from T. solium metacestodes to detect anti-parasite antibodies. The Ag-ELISA and Ab ELISAs revealed 64.7% and 79.0% seropositivity, respectively, in the Amerindian population. Immunoglobulin (Ig) M was the predominant antibody class, suggesting recent infection. In comparison sera from, clinically defined, hospital neurocysticercosis cases revealed only 27% seropositivity by Ag-ELISA, compared with 86-92% seropositivity by Ab-ELISA, and IgG4 was the predominant antibody subclass detected. The EITB antigen recognition patterns of the hospitalized patients were very similar to that of the Amerindians, confirming exposure to the parasite. These results, combined with the predominance of IgM antibody responses and the marked detection of secreted products of viable parasites, strongly suggest that recent exposure to T. solium had occurred in the Amerindian population. PMID- 12135290 TI - Acute respiratory illness incidence and death among children under two years of age on Lombok Island, Indonesia. AB - No childhood pneumonia incidence data for Indonesia exist, and few data exist for Asia as a whole. From February 1, 1998, to January 31, 1999, we conducted acute respiratory illness (ARI) surveillance among children < 24 months of age in 50 mainly rural villages on Lombok Island, Indonesia. The total number of child years at risk during the study period was 17,015. The documented incidences of simple, severe, hospitalized, and radiologically confirmed alveolar pneumonia were 21, 8.3, 5.3, and 1.8 per 100 child-years of observation, respectively. For all outcomes, the incidence was higher among younger and rural children. All cause and ARI-specific infant mortality rates were 84 and 33 per 1,000 live births, respectively. More than 65% of deaths due to ARI occurred outside of a hospital setting. The incidence of pneumonia is high in Lombok. Interventions should include introducing vaccines to prevent infections leading to pneumonia and increasing the access of critically ill infants to the health care system. PMID- 12135291 TI - Overexpression of vascular endothelial growth factor and its endothelial cell receptor KDR in type 1 leprosy reaction. AB - The sites of expression of vascular endothelial growth factor (VEGF) and of KDR, its endothelial cell receptor, were investigated in leprosy reaction Type 1, or reversal reaction (RR), by immunohistochemistry and in situ hybridization. In comparison with nonreactional leprosy, overexpression of both VEGF and KDR was seen in granuloma cells, especially epithelioid and foreign body-type giant cells, the epithelium and the vascular endothelium of RR specimens. In granuloma cells, hybridization for VEGF was stronger than immunostaining, a finding that may reflect the rapid turnover of VEGF in an immunologically dynamic situation such as RR. In the epidermis, double immunohistochemistry revealed VEGF overexpression in CDla-positive dendritic cells. The VEGF may not only be relevant for hyperpermeability and mononuclear cell differentiation (the key morphologic features in the acute, clinically evident phase of RR), but it could also be implicated in RR onset, when dendritic cells are activated in response to antigen stimulation. PMID- 12135292 TI - Modeling relationships between climate and the frequency of human plague cases in the southwestern United States, 1960-1997. AB - The relationships between climatic variables and the frequency of human plague cases (1960-1997) were modeled by Poisson regression for two adjoining regions in northeastern Arizona and northwestern New Mexico. Model outputs closely agreed with the numbers of cases actually observed, suggesting that temporal variations in plague risk can be estimated by monitoring key climatic variables, most notably maximum daily summer temperature values and time-lagged (1 and 2 year) amounts of late winter (February-March) precipitation. Significant effects also were observed for time-lagged (1 year) summer precipitation in the Arizona model. Increased precipitation during specific periods resulted in increased numbers of expected cases in both regions, as did the number of days above certain lower thresholds for maximum daily summer temperatures (80 degrees F in New Mexico and 85 degrees F in Arizona). The number of days above certain high-threshold temperatures exerted a strongly negative influence on the numbers of expected cases in both the Arizona and New Mexico models (95 degrees F and 90 degrees F, respectively). The climatic variables found to be important in our models are those that would be expected to influence strongly the population dynamics of the rodent hosts and flea vectors of plague. PMID- 12135293 TI - Short report: concurrent Rocky Mountain spotted fever in a dog and its owner. AB - A sequential occurrence of Rocky Mountain spotted fever (RMSF) in a dog and its owner is described. Diagnosis of RMSF in the animal guided subsequent testing for and diagnosis of the same disease in the human patient. Previous reports of concurrent RMSF in dogs and their owners are reviewed, and the epidemiologic significance of this occurrence is discussed. PMID- 12135294 TI - Etiology of encephalitis syndrome among hospitalized children and adults in Takeo, Cambodia, 1999-2000. AB - Whether or not Japanese encephalitis virus (JEV) is an important causative agent of acute encephalitis in Cambodia remains unclear. This study was carried out to determine the cause of encephalitis syndrome among children and adults admitted to Takeo Provincial Hospital from October 1999 to September 2000. Ninety-nine cases were included in the study: 52 pediatric cases (12 were fatal) and 47 adult cases (10 were fatal). A causative agent such as human herpesvirus (HHV-3 or HHV 4), Cryptococcus neoformans, or Mycobacterium tuberculosis had been identified in 8 of the 11 adults who had human immunodeficiency virus type 1 (HIV-1). An infectious agent was identified in 35 (40%) of 88 HIV-1-seronegative patients (60% of the causes remains unidentified). These comprised 11 bacterial infections, 1 fungal infection, and 23 viral infections. The viral infections were 1 fatal HHV-4 infection, 5 dengue virus infections (2 fatal), 1 coinfection with flavivirus and alphavirus, and 16 presumptive infections JEV (no virus detected), one case of which was fatal. Infection with JEV, the principal cause identified in the 99 encephalitis syndromes, concerned 16 (31%) of 52 children. PMID- 12135295 TI - Enzyme-linked immunosorbent assay-format microneutralization test for dengue viruses. AB - A microneutralization test that measures anti-dengue antibodies was developed. Serum dilutions, neutralization reactions, and virus growth were performed in 96 well plates. After incubation, an enzyme-linked immunosorbent assay that used mouse anti-dengue antibodies and an enzyme-conjugated anti-mouse antibody was used to measure cell-associated viral antigens. The resulting optical density readings were processed and graphed automatically by a spreadsheet program. This procedure provided results that are essentially the same as those from the plaque reduction neutralization test for serum samples from primary dengue virus infections, but results correlated poorly with results from samples from people with secondary infections. The test offers the advantages of ease of performance, ease in the calculation of results, lower cost, and increased speed. PMID- 12135296 TI - Breeding structure of Aedes aegypti populations in Mexico varies by region. AB - A population genetic analysis of Aedes aegypti was conducted among 38 collections from throughout coastal regions of Mexico. Multiple collections were made within 5 cities to examine local patterns of gene flow. Single-strand conformation polymorphism analysis was used to screen for variation in a 387-bp region of the Nicotinamide Adenine Dinucleotide Dehydrogenase subunit 4 mitochondrial gene (ND4) and 25 haplotypes were detected. Northeastern Mexico collections were genetically differentiated from and had lower genetic diversity than Yucatan and Pacific coastal collections. Yucatan and Pacific collections were genetically homogeneous. Regression analysis of geographic distances and F(ST) values indicated that collections were genetically isolated by distance in the Pacific and the Yucatan, but not among collections in the northeast. Free gene flow occurred among all collections within 130 km of one another in the northeast and within 180 km in the Yucatan. F(ST) values were never large among Pacific collections, suggesting extensive gene flow along the Pacific coast. PMID- 12135297 TI - Physical impairment and functional outcome in patients having lower extremity fractures after age 65. AB - The purpose of this study was to evaluate physical impairment and functional outcome in a group of patients who had a fracture of a lower extremity after age 65. The 30 subjects participating were treated between January 1992 and February 1998, for a unilateral fracture of the lower extremity. At the return examination, the patients' lower extremities were evaluated for range of motion (ROM) and strength. The participants also completed the Short Form 36 (SF-36) questionnaire, and their scores were compared with those of age-matched controls to determine relative disability. There were no statistically significant differences in ROM or muscle strength of the injured versus noninjured extremities, except in the case of knee flexion. The Mental Component Score (MCS) of the SF-36 in our study population was statistically different from that of controls, and there was no statistical difference in the Physical Component Score (PCS). A moderate correlation was found between the PCS and physical impairment. PMID- 12135298 TI - Complications of the anterior retropharyngeal approach in cervical spine surgery: a technique and outcomes review. AB - The retropharyngeal approach is used to avoid the risks and limitations of transmucosal surgery. The standard Smith-Robinson approach does not allow complete exposure of the C3 body/disk in patients requiring instrumentation of C3 or in patients with a short, thick neck. The anterior retropharyngeal approach provides additional exposure to the entire cervical spine in these patients. Our results in 14 cases show the anterior retropharyngeal approach to the upper and lower cervical spine to be an effective surgical technique in cases of upper cervical spine abnormalities and for multilevel abnormalities in patients with a short, thick neck. Although complications occurred as a result of the procedure, no permanent disorders were encountered. Adequate exposure to the entire cervical spine can be achieved without the high incidence of infection associated with the transoral approach. PMID- 12135299 TI - Preoperative administration of epoetin alfa to reduce transfusion requirements in elderly patients having primary total hip or knee reconstruction. AB - Avoidance of allogeneic blood transfusion is challenging in elderly patients with multiple comorbid conditions, who need major elective orthopaedic surgery. We conducted a study comparing the safety and efficacy of preoperative recombinant human erythropoietin (epoetin alfa) in patients having major elective orthopaedic surgery and in matched historical control patients. Patients aged 70 years or more undergoing primary total knee arthroplasty (TKA) or total hip arthroplasty (THA) were given two or three doses, respectively, of epoetin alfa (40,000 IU). They also received oral iron for 14 or 21 days. Epoetin alfa increased hemoglobin (Hb) and hematocrit (HCT) levels before surgery; 74% of control patients required blood transfusion, compared with 12.5% of patients receiving epoetin alfa. No serious adverse event was attributed to study treatment. These data indicate that epoetin alfa is safe and effective and reduces the need for allogeneic blood transfusion in elderly patients having elective total joint replacement. PMID- 12135300 TI - Patient compliance and satisfaction with mechanical devices for preventing deep venous thrombosis after joint replacement. AB - A consecutive series of patients having total joint arthroplasty at a single university hospital were sequentially treated with two mechanical devices for prevention of deep venous thrombosis (DVT). The first 104 patients (group 1) wore thigh-high sequential compression device (SCD). The next 120 patients (group 2) wore a foot pump. Daily documentation of hourly compliance with each respective device was recorded until discharge. A patient satisfaction questionnaire was also obtained. Patient understanding about the devices' function aided compliance (73% compliance in group 1, and 77% in group 2). The satisfaction questionnaire revealed significantly greater satisfaction in group 2 (73%) versus group 1 (55%). Of a subgroup of 35 patients who had used both devices, 24 preferred the foot pump, 7 the SCD, and 4 had no preference. This study showed a higher degree of compliance and satisfaction for foot pumps as prophylaxis against DVT. PMID- 12135301 TI - Biomechanics of minor automobile accidents: treatment implications for associated chronic spine symptoms. AB - Biomedical experimental data indicate that automobile accidents with no vehicle damage are unlikely to cause injury to the occupants. Soft tissue injuries heal in a few weeks. Chronic pain has never been produced by experimental injury and is much less common in countries without financial payments for accident victims. Chronic pain after no-damage collisions is probably caused by psychologic factors. Psychologic treatment should be added to conventional nonoperative treatment when no objective explanation for chronic symptoms is found. PMID- 12135302 TI - Reinfusion drains after primary total hip and total knee arthroplasty. AB - The purposes of this study were to evaluate the efficacy of intraoperative surgeon-elected reinfusion drain placement and to determine whether drainage at 90 minutes is useful in predicting the need for a reinfusion drain. In the standard drain hip arthroplasty group, 6 of 30 patients (20%) received a reinfusion, similar to the 11 of 41 patients (27%) in the reinfusion drain group. In the total knee arthroplasty group, 38 of 45 patients (84%) in the standard group had reinfusion, similar to the 23 of 27 patients (85%) in the reinfusion drain group. The surgeon could not predict intraoperatively which patients would need a subsequent reinfusion drain. However, in more than 94% of the cases, one could know by 90 minutes postoperatively whether a reinfusion would be necessary. We believe that a drain that can be converted to a reinfusion drain in the recovery room would be the most cost-effective drain system. PMID- 12135303 TI - Perioperative enteric nutritional supplementation in pediatric patients with neuromuscular scoliosis. AB - Perioperative nutritional status has been shown to be important in minimizing complications after extensive spinal procedures. Traditionally, total parenteral nutrition has been used to supplement oral nutrition intake. Little information exists regarding the risks and benefits of enteric supplementation in pediatric patients. To assess use and safety of enteric nutritional supplementation after extensive pediatric spine surgery, a retrospective review was done of 21 consecutive pediatric cerebral palsy patients receiving enteric nutritional supplementation via nasal or gastric feeding tubes after, and/or between stages of, anterior and posterior spinal fusions. Enteric supplementation was maintained for an average of 9.1 days, and the lowest albumin and total protein levels were seen the third postoperative day. Enteric nutritional supplementation was found to be a safe alternative to total parenteral nutrition in pediatric cerebral palsy patients after spinal stabilization procedures. In addition, enteric feedings may be less costly than central hyperalimentation and do not carry the inherent risk of central venous access. PMID- 12135304 TI - Complications in total knee arthroplasty with and without surgical drainage. AB - The standard practice in total joint arthroplasty has included the use of postsurgical drains to minimize perioperative wound complications, particularly infection. This practice is not without cost and potential morbidity. Our recent cemented and cementless total knee arthroplasties (TKAs) have been done without the use of postoperative surgical drains and without any appreciable increase in wound complications. To confirm this, we retrospectively reviewed 227 consecutive TKAs, specifically evaluating perioperative wound complications. No statistical increase in perioperative complications in TKAs without drains was found. A lower percentage of complications was seen in the cementless population when compared with cemented or drained knees. We suggest that surgical drainage is not required in TKA, even when cementless fixation is used. PMID- 12135305 TI - Isolated acute hip adductor brevis strain. AB - Acute muscle strain occurs as a result of an eccentric contraction that exceeds the biomechanical strength of the musculotendinous junction of a single muscle within a synergistic group. To date, only the (hip) adductor longus was shown to sustain this type of injury. In this case report, I describe the first published example of an magnetic resonance imaging (MRI)-documented acute hip adductor brevis strain. PMID- 12135306 TI - Intraosseous ganglia of glenoid. AB - A rare case of intraosseous ganglia of the glenoid in a 35-year-old woman is presented. The patient had painful right shoulder and no limit of motion. Radiographs and computed tomographic scans showed a large lytic lesion involving the entire glenoid bone. The patient was treated by curettage and autocorticocancellous bone graft. Six months after the operation, the patient has an excellent clinical outcome and radiologic sign of integration of the bone graft. Few cases of intraosseous ganglia of the glenoid have been reported, but none with the entire glenoid involvement. PMID- 12135307 TI - Incidental finding of chondroid lipoma at total hip arthroplasty. AB - Chondroid lipoma is a rare, benign, lipomatous soft tissue tumor that manifests as a painless lump. We report the incidental finding of such a tumor at total hip replacement and briefly review the literature regarding the histopathologic features of this recently described neoplasm. PMID- 12135308 TI - Genetic analysis in human hypertension. AB - Hypertension is considered to be a complex trait to which genetic, environmental, and demographic factors contribute interactively. Recently, molecular genetic studies have achieved remarkable success in the elucidation of causative mutations in several Mendelian hypertensive disorders in which single nucleotide polymorphisms (SNPs) disrupt the function of single genes, thereby leading to unambiguous phenotypes. It seems unlikely, however, that such a simple base substitution is the primary mechanism in cases of essential hypertension, even if SNPs modify the relevant gene function to some extent. Despite the enormous efforts made to date, no consistent association between any of the candidate genes and essential hypertension has been established. One plausible explanation is that because individual genes play a modest role in the pathogenesis of hypertension, confounding variables, whether individual (sex, ethnic origin, etc.) or environmental, may decrease the chance of identifying a causative relation between the genes and hypertension, depending on the populations studied. Several approaches can be proposed to overcome this problem, including long-term follow-up of clinical events collected to attain sufficient phenotypic information and statistical power. With the recent advances in high-throughput genotyping techniques and bioinformatic strategies, it has become possible to perform even SNP-based genome-wide screening. At present, however, the need for identification of susceptibility genes for hypertension still poses a great and unanswered challenge. Nonetheless, we believe that a precise understanding of the manner in which genetic variations affect hypertension can be achieved, and that clarification of the associated phenotypes will lead to the development of effective preventive and treatment strategies. PMID- 12135309 TI - Differential effects of a long-acting angiotensin converting enzyme inhibitor (temocapril) and a long-acting calcium antagonist (amlodipine) on ventricular ectopic beats in older hypertensive patients. AB - We studied differences in the effects of a long-acting angiotensin-converting enzyme (ACE) inhibitor (temocapril) and a long-acting calcium channel blocker (amlodipine) on ventricular ectopic beats (VEB) in relation to sympathetic nerve activity in 46 patients with essential hypertension. We performed 24-h Holter electrocardiography and ambulatory blood pressure (BP) monitoring simultaneously, and examined blood samples during the baseline, temocapril and amlodipine treatment periods. The ambulatory BP was lower in the amlodipine period than in the temocapril period. However, the number of VEB was significantly increased in the amlodipine period compared to that in the baseline period (11.9 vs. 7.4/day, p<0.05). In the temocapril period, the number of VEB was not significantly increased compared to that in the baseline period (8.6 vs. 7.4/day, p=0.30). Ambulatory heart rate (HR) was significantly increased in the amlodipine period compared to that in the baseline period (24-h HR: 70 vs. 66 bpm, p<0.001; daytime HR: 75 vs. 71 bpm, p<0.001; nocturnal HR: 60 vs. 58 bpm, p<0.05). Plasma norepinephrine (NE) also was significantly increased in the amlodipine period compared to that in the baseline period (457 vs. 369 pg/ml, p<0.001). However, when patients receiving amlodipine were divided into a high dose group (8.6 +/- 1.2 mg/day) and a low dose group (4.6 +/- 1.2 mg/day), increases in HR and plasma NE levels were found only in the high dose group. These results indicate that amlodipine is effective at lowering BP in older hypertensives, although it may increase VEB, especially when given at a high dose. PMID- 12135310 TI - Increase in pulse pressure relates to diabetes mellitus and low HDL cholesterol, but not to hyperlipidemia in hypertensive patients aged 50 years or older. AB - Higher pulse pressure is associated with higher cardiovascular risk. We investigated the relationship between pulse pressure and known metabolic risk factors in hypertensive patients who had not experienced stroke or myocardial infarction. In a multicenter cross-sectional survey made in 1995, we registered 939 hypertensive patients aged > or = 50 years. Of these, 734 had never experienced stroke or myocardial infarction. We divided these 734 patients into two groups based on the value of their pulse pressures: 396 patients with a pulse pressure > or = 60 mmHg, and 338 patients with a pulse pressure<60 mmHg. The average pulse pressure value was 72 +/- 12 mmHg in the former group, and 49 +/- 8 mmHg in the latter group. The former group exhibited advanced age, a higher women to-men ratio, lower high-density lipoprotein (HDL) cholesterol, and higher systolic and lower diastolic blood pressure. Diabetes mellitus (DM) and left ventricular hypertrophy were more frequently noticed in the former group than in the latter group. The prevalence of hyperlipidemia, however, was similar in the two groups. The association of pulse pressure with DM and low HDL cholesterol was statistically significant by multiple logistic analysis adjusted for age, sex, and other known cardiovascular risk factors. In conclusion, pulse pressure increases with advancing age. DM made a substantially larger contribution to the increase in pulse pressure than hyperlipidemia. PMID- 12135311 TI - Clinical significance of blood pressure response triggered by a doctor's visit in patients with essential hypertension. AB - The clinical significance of the pressor response triggered by blood pressure measurement, the so-called "white-coat effect," was studied in relation to left ventricular structure and function in patients with essential hypertension. We studied 75 consecutive, never-before treated patients with essential hypertension (54 +/- 2 (SE) years; 31 men). Beat-to-beat blood pressure (Finapres) was monitored at rest, during conventional blood pressure measurement by a doctor, and during a mental stress test. The left ventricular mass index and diastolic function (EIA ratio) were determined by echocardiography. The systolic blood pressure response triggered by the doctor's visit (deltaSBP) correlated positively with the left ventricular mass index (r= 0.326, p<0.03) and negatively with the EIA ratio (r=-0.325, p<0.02). A positive relationship between the deltaSBP and left ventricular mass index was observed in men (r=0.556, p<0.01) but not in women. The greater ASBP also was associated with lower EIA ratio in women (r=-0.434, p<0.02). The deltaSBP correlated with the mental stress-induced increase in systolic blood pressure in men (r=0.586, p<0.005) but not in women (r=0.148, n.s.). Blood pressures outside the clinic were higher in men than in women (p<0.05 for systolic and p<0.005 for diastolic) despite the similar level of clinic blood pressures between the sexes. Stepwise multiple linear regression analysis showed that the deltaSBP was an independent predictor of the left ventricular mass index in men (beta=0.783, p=0.0009) and of the EIA ratio in women (beta=-0.003, p=0.05). These data suggest that the pressor response triggered by a doctor's visit has clinical significance in never-before treated hypertensive patients, possibly because it mirrors real-life stress reactivity in men. PMID- 12135312 TI - A simple oscillometric technique for determining new indices of arterial distensibility. AB - Recently, oscillometric devices have been developed that can measure blood pressure in the extremities and analyze pulse volume record. On the basis of the extremity pulse volume record, these devices can automatically determine three types of simply measured pulse wave velocity (PWV) (brachial PWV: heart to right upper arm; R-PWV: right upper arm-right ankle; and L-PWV: right upper arm-left ankle). The percent mean pulse volume record (%MPVR=the height that bisects the area of the pulse volume record/pulse pressure X100), a quantitative index of right brachial pulse volume record, can also be determined. To evaluate the usefulness of these new indices, we studied 1,067 consecutive subjects undergoing health checkups (648 men, 419 women; mean age, 50 +/- 9 years). In both sexes, age correlated positively with simply measured PWVs (men, brachial PWV: r=0.46, p<0.0001; R-PWV: r=0.46, p<0.0001; L-PWV: r=0.47, p<0.0001; women, brachial PWV: r=0.37, p<0.0001; R-PWV: r=0.47, p<0.0001; L-PWV: r=0.48, p<0.0001) and correlated negatively with %MPVR (men: r=-0.40, p<0.0001; women: r=-0.45, p<0.0001). Simply measured PWVs and %MPVR were significantly correlated with mean blood pressure. In a separate group of 60 patients, simply measured PWVs correlated positively with carotid PWV (heart to carotid) derived from an elastic vessel (brachial PWV: r=0.76, p<0.0001; R-PWV: r=0.43, p<0.01; L-PWV: r=0.43, p<0.01). %MPVR correlated negatively with carotid PWV (r=-0.35, p<0.01). In conclusion, simply measured PWVs and %MPVR are easier to determine than conventional PWV and may be useful as new indices of age-related changes in arterial distensibility. PMID- 12135313 TI - Validity, reproducibility, and clinical significance of noninvasive brachial ankle pulse wave velocity measurement. AB - The present study was conducted to evaluate the validity and reproducibility of noninvasive brachial-ankle pulse wave velocity (baPWV) measurements and to examine the alteration of baPWV in patients with coronary artery disease (CAD). Simultaneous recordings of baPWV by a simple, noninvasive method and aortic pulse wave velosity (PWV) using a catheter tip with pressure manometer were performed in 41 patients with CAD, vasospastic angina, or cardiomyopathy. In 32 subjects (15 controls and 17 patients with CAD), baPWV was recorded independently by two observers in a random manner. In 55 subjects (14 controls and 41 patients with CAD), baPWV was recorded twice by a single observer on different days. baPWV were compared among 172 patients with CAD (aged 62 +/- 8 years); 655 age-matched patients without CAD but with hypertension, diabetes mellitus, or dyslipidemia; and 595 age-matched healthy subjects without these risk factors. baPWV correlated well with aortic PWV (r=0.87, p<0.01). Pearson's correlation coefficients of interobserver and intraobserver reproducibility were r=0.98 and r=0.87, respectively. The corresponding coefficients of variation were 8.4% and 10.0%. baPWV were significantly higher in CAD patients than in non-CAD patients with risk factors, for both genders (p<0.01). In addition, baPWV were higher in non CAD patients with risk factors than in healthy subjects without risk factors. Thus, the validity and reproducibility of baPWV measurements are considerably high, and this method seems to be an acceptable marker reflecting vascular damages. baPWV measured by this simple, noninvasive method is suitable for screening vascular damages in a large population. PMID- 12135314 TI - Effects of angiotensin converting enzyme inhibitor and calcium antagonist on endothelial function in patients with essential hypertension. AB - The endothelium plays an important role in maintaining vascular tone and function. Essential hypertension is associated with alterations in endothelial function. The effects of antihypertensive agents on endothelial function have not been fully evaluated in human hypertension and data on the forearm circulation of humans are controversial. The aim of this study was to determine whether treatment with an angiotensin converting enzyme (ACE) inhibitor or a calcium antagonist improves endothelial dysfunction in hypertensive patients and whether the mechanism involved could be related to antioxidant activity. Endothelial function was estimated using venous occlusion plethysmography in 18 hypertensive patients and 11 healthy volunteers. The patients in the hypertension group were treated with enalapril or amlodipine. The change of forearm blood flow (FBF) was measured during acetylcholine infusion through the brachial artery and also during intra-arterial vitamin C infusion to explore the effects of vitamin C on responses to acetylcholine. FBF response to acetylcholine was significantly enhanced by intra-arterial infusion of vitamin C in the hypertensive group before antihypertensive treatment. Co-infusion of L-NMMA(N(G)-monomethyl-L-arginine), an inhibitor of nitric oxide synthase, blunted forearm blood flow response to acetylcholine. After antihypertensive treatment with enalapril or amlodipine for 2 months in the hypertensive group, endothelium-dependent vasorelaxation (vasodilatory response to acetylcholine) was significantly improved. Even though the mechanisms leading to depressed endothelial function in essential hypertension remain to be elucidated, our study shows that treatment with an ACE inhibitor or a calcium antagonist resulted in demonstrable improvement by a mechanism that is probably related to antioxidant activity. PMID- 12135315 TI - Vascular remodeling of the carotid artery in patients with untreated essential hypertension increases with age. AB - We examined whether hypertrophy of the carotid artery in patients with untreated essential hypertension is associated with compensatory carotid artery enlargement as these patients age. Carotid ultrasonography was evaluated in 163 patients with untreated essential hypertension (74 males and 89 females) and in 76 normotensive subjects. Intima-media end-diastolic thickness (IMT) and outer vessel diameter (VD) were measured, and relative wall thickness (IMT/R, R=VD/2) and vascular mass (VM) were calculated. Determinants of vascular hypertrophy in patients with untreated essential hypertension were also investigated. VD, VM, and IMT were significantly correlated with age in both the normotensive and hypertensive groups. Additionally, IMT was significantly correlated with VD in both groups. There was no correlation between increasing age and IMT/R in either group. IMT, VD and VM were significantly higher in the hypertensive group >50 years than in age-matched normotensive controls. However, IMT/R was significantly higher in the 50-59 years hypertensive group than in normotensive controls of the same age group. In addition to age, VM was related to systolic blood pressure, pulse pressure, fasting blood sugar, IMT, VD, and IMT/R in the hypertensive group. Multivariate regression analysis in the hypertensive group indicated that IMT/R was the strongest predictor of carotid vascular mass. Age and pulse pressure were also independently related to vascular mass. These results indicate that, as patients with untreated hypertension age, carotid arteries undergo remodeling. This should add further impetus to the implementation of appropriate hypertension treatment for such patients. PMID- 12135316 TI - Reverse J-curve relation between diastolic blood pressure and severity of coronary artery lesion in hypertensive patients with angina pectoris. AB - The existence of the J-curve in hypertension treatment remains controversial. The major question is whether the increase in mortality from coronary disease is induced by the lowering of blood pressure (BP) or by the severity of underlying coronary artery disease. We recruited patients with a history of hypertension (systolic BP (SBP) >160 mmHg and/or diastolic BP (DBP) >90 mmHg) and a diagnosis of angina pectoris with angiographically confirmed coronary artery lesion. The relationship among the treated levels of SBP and DBP, the severity of coronary artery lesion, and the clinical consequences were investigated. Among the 234 enrolled patients, 115 experienced further events, 19 of which were serious. There were no significant differences in the average BP of patients with and those without events, but the coronary severity indices (CSI) were significantly greater in patients with events. As a function of DBP from < or = 74 to 105 < or = mmHg, there was a positive association with the incidence of serious events, and a reversed J-curve in CSI with a nadir at 95-104 mmHg. A similar relationship was observed in SBP, but a potentially unfavorable outcome was suggested in the lowest SBP range of < or = 124 mmHg. In conclusion, there was no J-curve for DBP in hypertensive patients with angina pectoris; rather, the lower the DBP, the better was the prognosis. Interestingly, the severity of coronary lesion is in a reversed J-curve relation with DBP, suggesting that high BP plays a critical role in serious events in hypertensive patients with moderate coronary artery lesions. PMID- 12135317 TI - Association of G-33A polymorphism in the thrombomodulin gene with myocardial infarction in Koreans. AB - Thrombomodulin (TM), a thrombin receptor expressed on the endothelial surface, is known to play an important role in the anti-thrombogenic system in vivo. In this study, we examined the effects of 3 single-nucleotide polymorphisms (SNPs) in the TM gene (G-33A, C1418T and C1922T) on the development of myocardial infarction (MI) in Koreans. We found that G-33A was a common SNP (the minor allele frequency was 0.09) in Koreans. Eighty-five MI patients who had received coronary angiography were enrolled and were divided into 3 groups according to the number of coronary arteries in which stenosis was found angiographically (1-vessel disease (1VD) to 3-vessel disease (3VD)). The criterion of coronary stenosis was 50% or more stenosis on angiography. In addition, 102 controls (CONT) who had no significant stenosis were employed. The number of AA/GA genotypes of G-33A was found to be significantly greater in the 1VD than in the CONT (p=0.004 by chi2 test) while no significant difference was found between the multivessel disease (2-3VD) and the CONT. Multiple logistic analysis showed that G-33A was an independent risk factor for the 1VD with an odds ratio of 4.63 (95% confidence interval; 1.62-13.3). C1418T and C1922T were both in linkage disequilibrium with G-33A; however, they were not independent risks for either the 1VD or the 2-3VD. A reporter gene assay showed that G-33A had a significant effect on the TM promoter activity. These results indicated that G-33A polymorphism in TM might be a genetic risk factor for myocardial infarction. PMID- 12135318 TI - A novel missense mutation of exon 3 in the type A human natriuretic peptide receptor gene: possible association with essential hypertension. AB - The natriuretic peptide (NP) family is involved in regulation of blood pressure and fluid volume. We recently characterized the exon/intron organization of the human type A NP receptor (hNPRA) gene. The aim of this study was to isolate the genetic markers according to the organization of this gene, and to study the association between this gene and essential hypertension. Using polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) analysis, we identified a novel missense mutation, M3411, consisting of a methionine (ATG) to isoleucine (ATC) substitution at nucleotide 1023 in exon 3. Computer-aided three dimensional structural analysis suggested that M341 exists in the loop between two alpha-helices, and that the mutation may influence receptor activities by altering the conformation of the alpha-helices. We performed an association study of the mutation in 210 essential hypertension (EH) patients and 210 normotensive controls. The overall distribution of alleles was not significantly different between the control and EH groups. However, the C/C homozygous genotype was found only in the EH group. The ratio of plasma brain natriuretic peptide (BNP)/mean blood pressure of the C/C genotype was significantly higher than that of the G/G genotype or the G/C genotype. We conclude that the significance of homozygous M3411 mutation in exon 3 is worth investigating for its possible association with EH. PMID- 12135319 TI - Fasting plasma glucose is an independent determinant of left ventricular diastolic dysfunction in nondiabetic patients with treated essential hypertension. AB - Left ventricular (LV) hypertrophy and LV diastolic dysfunction are common cardiac changes in hypertensive patients, and these changes are modified by various factors other than blood pressure. The present study was conducted to investigate the influence of mild abnormalities in glucose metabolism on LV structure and function in essential hypertension. In 193 nondiabetic patients with treated essential hypertension, two-dimensional and Doppler echocardiographic examinations were performed, and relative wall thickness (RWT), LV mass index (LVMI), fractional shortening, and the ratio of the peak velocity of atrial filling to early diastolic filling (A/E) were calculated. Fasting plasma glucose (FPG) and HbA1c levels were positively correlated with the A/E ratio and the deceleration time of the E wave. However, these plasma levels had no correlation with RWT, LVMI, or fractional shortening. Peak A wave velocity and the A/E ratio were significantly higher in patients who had FPG of > or = 100 mg/dl (and <126 mg/dl) than those who had FPG of <100 mg/dl, although age, blood pressure, RWT, LVMI, and fractional shortening did not differ between the two groups. In a multiple regression analysis of all subjects, only FPG and age were independent determinants of the A/E ratio. These observations suggest that FPG is a sensitive predictor for LV diastolic dysfunction in nondiabetic patients with treated hypertension. Since a slight increase in plasma glucose levels is associated with abnormalities in diastolic function independent of LV hypertrophy, an early stage of impaired glucose metabolism in hypertensive patients may specifically deteriorate cardiac diastolic function. PMID- 12135320 TI - Decreased expression of arginase II in the kidneys of Dahl salt-sensitive rats. AB - Arginase catalyzes the hydrolysis of arginine to urea and ornithine. Urea is not only an important solute for concentrating urine but also inhibits Na-K-2Cl cotransport. To elucidate the roles of arginase in the development of salt sensitive hypertension, we examined arginase activity and expression in the kidney and other organs of Dahl/Rapp salt-sensitive (SS) and salt-resistant (SR) rats before and after 4 weeks' administration of a 4% NaCl or control diet. At 4 weeks of age, arginase activity in the kidney was lower in SS rats than in SR rats. Kidney arginase activity was lower in SS rats than in SR rats at 8 weeks of age, and salt loading did not alter arginase activity. Arginase II (the dominant isoform in the kidney) mRNA and protein in the kidney of salt-loaded SS rats were also lower than those of salt-loaded SR rats. Arginase activities in the liver and cerebellum did not differ between SS and SR rats. To examine the effect of urea, the product of arginase reaction, on the development of hypertension, SS rats were given a 4% NaCl diet containing 5% kaolin or 5% urea. Six-week urea supplementation attenuated the development of hypertension in SS rats. These findings suggest that decreased arginase expression in the kidney may be at least partially responsible for the salt-sensitive hypertension in SS rats. PMID- 12135321 TI - Carvedilol inhibits pressure-induced increase in oxidative stress in coronary smooth muscle cells. AB - The cellular mechanisms by which hypertension enhances atherosclerosis are still not known in detail. Recently, evidence has been obtained that oxidative stress plays a role in the pathogenesis of pressure-induced atherosclerosis. We examined the effects of pressure on oxidative stress in cultured human coronary smooth muscle cells (SMCs). Application of increased pressure (+100 mmHg) with He gas for 48 h increased oxidative stress of measured by flow cytometry by 71% and F2 isopretane by 77%. Increased pressure also increased the activities of phospholipase D (PLD), and particulate protein kinase C (PKC). The PLD inhibitor suramin 100 micromol/l, 1-butanol 40 mmol/l, and the PKC inhibitors chelerythrine 1 micromol/l and calphostin C 100 nmol/l and completely blocked the increase in oxidative stress induced by pressure. Carvedilol 1 micromol/l but not propranolol 1 micromol/l blocked pressure-induced increases in oxidative stress in cultured SMCs. These findings suggest that pressure increases oxidative stress and that carvedilol significantly inhibits pressure-induced increase in oxidative stress in cultured human coronary smooth muscle cells. PMID- 12135322 TI - Galpha13 induces preproET-1 gene expression via JNK. AB - The endothelin B receptor (ETBR) has been shown to mediate autoinduction of endothelin-1 (ET-1). We previously reported that the ET(B)R interacts with Galpha13, a member of the heterotrimeric GTP-binding protein family. In the present study, we examined whether Galpha13 induces preproET-1 (ppET-1) gene transcription, which could result in ET-1 autoinduction in a renal epithelial cell line. We generated a reporter gene construct under control of the ppET-1 promoter region. The construct was transiently expressed in COS-7 cells. Transient expression of ETBR increased the promoter activity of ppET-1 following treatment with 100 nmol/l of ET-1. Expression of Galpha13Q226L or Galpha9209L, constitutively active forms of Galpha13 and Galpha9, also activated the ppET-1 promoter. ETBR-stimulated ppET-1 promoter activity was partially diminished by the expression of dominant negative forms of c-Jun N-terminal kinase (JNK1APF) or MAPK/ERK kinase (MEKK97M). Expression of JNK1APF also inhibited Galpha13Q226L induced ppET-1 promoter activation. These findings indicate that Galpha13 can induce ppET-1 gene expression through a JNK-mediated pathway. Our results also suggest that this Galpha13-coupled signaling pathway may play an important role in a sustained ET-1 autoinduction loop in various pathophysiological conditions. PMID- 12135323 TI - Nipradilol prevents L-NAME-exacerbated nephrosclerosis with decreasing of caspase 3 expression in SHR. AB - The proliferative cell nuclear antigen (PCNA) is an auxiliary protein of DNA polymerase delta and appears to be required for both DNA synthesis and repair. Previously, we showed that prolonged NO synthase (NOS) inhibition produced severe nephrosclerosis with an increase of glomerular cell DNA fragmentation (apoptosis), glomerular ischemia and hypertension in spontaneously hypertensive rats (SHR). The objective of the present study was to investigate the effects of the vasodilating, nonselective, NO-releasing beta-adrenoceptor blocker nipradilol on DNA fragmentation and synthesis/repair of glomerular cells in this prolonged NOS blockaded SHR. Twenty-week-old SHR were administered an NOS inhibitor, N(G) nitro-L-arginine methyl ester (L-NAME, 80 mg/l in drinking water) or co-treated with the same dose of L-NAME and nipradilol (20 mg/kg/day) for 3 weeks. After this treatment, expression of apoptosis was histologically examined using caspase 3, an apoptosis inducer, in addition PCNA (DNA synthesis/repair), and examination of glomerular morphometric changes, including cell number and tuft area. Nipradilol reduced blood pressure and preserved creatinine clearance reduction in L-NAME/SHR. These effects were associated with normalization of the glomerular cell apoptosis index and caspase-3 score, an increase in PCNA index, and increases in glomerular cell numbers and glomerular tuft area, resulting in a decreased glomerular injury score. Thus, in SHR administered an NOS inhibitor, nipradilol improved nephrosclerosis in association with a decrease in apoptosis and an increase in DNA synthesis/repair of glomerular cells. These findings may provide important insights into DNA repair/repair and apoptosis in nephrosclerosis. PMID- 12135324 TI - Regional hemodynamic effects of adrenomedullin in Wistar rats: a comparison with calcitonin gene-related peptide. AB - Because both vasodilation induced by adrenomedullin (AM) and that induced by calcitonin gene-related peptide (CGRP) may occur via the same receptor, the two peptides might play similar roles in circulation. To examine this possibility, we used the colored microsphere technique and an ultrasonic flowmeter to investigate the systemic and regional effects of an equivalent dose (650 pmol/l) of AM and CGRP in conscious Wistar rats. AM significantly decreased mean arterial pressure and peripheral resistance but increased heart rate and cardiac index (CI). On the other hand, hypotension induced by CGRP was not accompanied by an increment in Cl. Both AM and CGRP increased the femoral arterial blood flow measured by the flowmeter, with the increase by AM being significantly larger. The regional hemodynamic effects were quite different between the two peptides. AM increased the blood flow in the heart, lungs, kidneys, adrenal glands, and spleen, whereas CGRP increased blood flow only in the heart. On the other hand, CGRP increased the cutaneous and gastric blood flows, which were not affected by AM. These differences in the regional vasodilatory effects of AM and CGRP suggest that the two peptides do not play similar roles in circulatory regulation. PMID- 12135325 TI - Combination therapy with an angiotensin-converting enzyme (ACE) inhibitor and a calcium antagonist: beyond the renoprotective effects of ACE inhibitor monotherapy in a spontaneous hypertensive rat with renal ablation. AB - To assess the renal benefits of combined angiotensin-converting enzyme inhibition and calcium antagonism, we studied the antihypertensive and renoprotective effects of temocapril (TMP) alone or in combination with azelnidipine (AZN) in a spontaneously hypertensive rat (SHR) remnant kidney model of chronic renal failure. Male 5/6-nephrectomized SHR/Izumo rats were randomly assigned to receive vehicle (control group), TMP (TMP group; 10 mg x kg(-1) x day(-1)), AZN (AZN group; 3 mg x kg(-1) x day(-1)), or both (TMP+AZN group) orally for 12 weeks. Systolic blood pressure (SBP) and urinary excretion of albumin (UalbV) were measured every 2 weeks. At the end of the experiment, serum creatinine (Scr), heart weight (HW), and blood urea nitrogen (BUN) levels were measured and the remnant kidneys were examined to determine the index of glomerular sclerosis (IGS). SBP and UalbV in the control group increased progressively throughout the experimental period. TMP, AZN, and TMP+AZN blocked the development of hypertension. TMP+AZN did not enhance the antihypertensive effects of either TMP or AZN used singly. TMP, AZN, and TMP+AZN all significantly decreased the UalbV, Scr, BUN, and HW/body weight (BW) ratio. The level of UalbV and the HW/BW ratio in the TMP+AZN group were significantly lower than those in the TMP and AZN groups, and the level of Scr in the TMP+AZN group was significantly lower than that in the TMP group. TMP, AZN, and TMP+AZN all significantly protected against an increase in the IGS. The IGS in the TMP+AZN group was significantly lower than that in the TMP and AZN groups. These results indicate that both TMP and AZN have antihypertensive and renoprotective effects in this model. They also suggest that simultaneous administration of TMP and AZN provides greater renoprotective effects than TMP alone. PMID- 12135326 TI - Lowering of blood pressure improves endothelial dysfunction by increase of nitric oxide production in hypertensive rats. AB - To investigate the effects of angiotensin converting enzyme inhibitor (ACE-I) and other antihypertensive agents on the nitric oxide (NO) release during hypertension, seven- and fourteen-week-old SHR and deoxycorticosterone acetate (DOCA)-salt rats were treated with hydralazine, manidipine (Ca antagonist) or quinapril (ACE-I) for 3 weeks to lower blood pressure. Systolic blood pressure (SBP) was measured by the tail cuff method once each week. Endothelial cells (ECs) derived from the descending aorta of the treated rats were cultured and NOx levels in culture media were measured with an NO analyzer based on the Griess reaction. In both SHR and DOCA-salt rats, antihypertensive therapy lowered SBP to levels similar to those of control rats. The only exception was quinapril treatment of DOCA-salt rats. Although NOx release by ECs derived from hypertensive rats was improved by antihypertensive therapy, the effect was most pronounced in SHR treated with quinapril. In addition, restoration of NOx release was much more remarkable in younger SHR. NOx release was significantly higher in DOCA-salt rats treated with quinapril than in control rats without reduction of SBP. These results suggest that lowering blood pressure improves release of NO by ECs during hypertension and that the time at which antihypertensive therapy is started is also important to preserve endothelial function. Furthermore, ACE-I is suggested to protect endothelial function by increasing NO production in addition to lowering blood pressure. PMID- 12135327 TI - Combination treatment with a calcium channel blocker and an angiotensin blocker in a rat systolic heart failure model with hypertension. AB - The mechanism and treatment of hypertensive systolic heart failure are not well defined. We compared the effect of an angiotensin-converting enzyme inhibitor (cilazapril, 10 mg/kg), an angiotensin receptor blocker (candesartan, 3 mg/kg), a calcium channel blocker (benidipine, 1, 3 or 6 mg/kg), and the same calcium channel blocker combined with renin-angiotensin blockers on systolic heart failure in Dahl salt-sensitive (DS) rats. DS rats were fed an 8% Na diet from 6 weeks of age and then subjected to the above drug treatments. Benidipine (1 mg/kg), cilazapril, and candesartan had compatible hypotensive effects and similar beneficial effects on cardiac hypertrophy, gene expression, and survival rate. The combination of benidipine with cilazapril or candesartan was found to have no additional beneficial effects on the above parameters, with the exception of a reduction in atrial natriuretic polypeptide gene expression. On the other hand, candesartan normalized serum creatinine, but serum creatinine was unaffected by either benidipine at 1 or 3 mg/kg or cilazapril. Further, the combined use of benidipine and either candesartan or cilazapril resulted in an additional reduction of urinary albumin excretion in DS rats. Thus systolic heart failure in DS rats is mainly mediated by hypertension, while renal dysfunction of DS rats is due to both hypertension and the AT1 receptor itself. These findings suggest that the combination of a calcium channel blocker with an AT1 receptor blocker or ACE inhibitor may be more effective in treating the renal dysfunction associated with systolic heart failure than monotherapy with either agent alone. However, further studies will be needed before reaching any definitive conclusion on the efficacy of this combination therapy in patients with heart failure. PMID- 12135328 TI - Two cases of malignant hypertension with reversible diffuse leukoencephalopathy exhibiting a reversible nocturnal blood pressure "riser" pattern. AB - We report two cases of malignant hypertension with reversible diffuse leukoencephalopathy demonstrating a nocturnal blood pressure (BP) rising pattern ("riser" pattern). Case 1 was a 54-year-old man diagnosed with malignant hypertension who presented with diffuse leukoencephalopathy and nocturnal BP rise during the acute phase. These abnormal findings diminished after treatment of hypertension. Case 2 was a 50-year-old woman diagnosed with malignant hypertension in association with leukoencephalopathy, heart failure and acute renal failure. She also presented with a "riser" pattern during the acute phase. In contrast to case 1, the leukoencephalopathy and "riser" pattern in case 2 were not improved even after 1 month of treatment. Following intensive antihypertensive treatment, renal failure was improved in case 1, but renal failure was not improved after 1 month in case 2. In conclusion, a possible explanation of this phenomenon is that a causative volume overload due to renal dysfunction produced the temporal leukoencephalopathy-like brain edema and "riser" pattern in these cases. PMID- 12135329 TI - Association of cardiovascular risk factors and endothelial dysfunction in japanese hypertensive patients: implications for early atherosclerosis. AB - Although hypertension, hyperlipidemia, diabetes and smoking are known risk factors of atherosclerosis in Caucasians, their relative contributions to early atherosclerosis among Japanese are unknown. Decrease in flow-mediated dilation (FMD) of the brachial artery is a useful marker of endothelial dysfunction and early atherosclerosis. To evaluate the relative contribution of hypertension to early atherogenesis, we determined FMD, as well as plasma levels of tissue-type plasminogen activator (t-PA; a sensitive index of endothelial damage) and tumor necrosis factor (TNF)-a and interleukin (IL)-6 (established markers of inflammation) in normotensive and hypertensive patients under treatment. FMD was significantly reduced as the number of risk factors increased, suggesting that accumulations of risk factors were related to endothelial dysfunction. FMD was reduced in hypertensives (9.9 +/- 5.8 (SD) %) compared to normotensives (14.6 +/- 7.6, p<0.01) despite good blood pressure control (139 +/- 20/80 +/- 14 mmHg in hypertensives). Nitroglycerine-induced endothelium-independent vasodilation was not altered in hypertensives (16.0 +/- 6.3%) as compared to normotensives (16.7 +/- 5.8). Plasma t-PA, TNF-alpha, and IL-6 levels were increased in hypertensives despite good blood pressure control. Thus, hypertension alone is a high risk for early atherosclerosis. Persistent endothelial damage and moderate inflammation may increase the risk of early atherosclerosis synergistically under the presence of hypertension in Japanese. PMID- 12135330 TI - Angiotensin-converting enzyme gene polymorphism and its association with essential hypertension in a Tibetan population. AB - There is strong evidence to support the idea that the renin-angiotensin system (RAS) plays an important role in the pathogenesis of essential hypertension (EH) and its complications. However, existing data about the association of angiotensin-converting enzyme (ACE) gene insertion/deletion (I/D) polymorphism with blood pressure is conflicting, mainly due to racial differences and environmental exposure status. We therefore conducted a case control study to observe the relationship between ACE I/D polymorphism and EH in a Tibetan population who live in relatively isolated areas and are genetically homogeneous. The study was conducted at stable residential communities in the urban district of Lhasa, the capital of the Tibet autonomous region, China, and 106 unrelated EH patients and 135 normotensIve subjects were recruited. PCR, PCR/RFLP and PCR-SSCP were carried out to study the association between RAS genes and EH. Frequencies for the DD, ID and II genotypes were 27, 47 and 29 in hypertensive subjects, and 15, 60 and 48 in normotensive subjects, respectively. Derived allele frequencies for the I and D alleles were 0.51 and 0.49 in hypertensive subjects and 0.64 and 0.36 in normotensive subjects. There were significant differences in genotype distribution and derived allele frequency between these two groups. The genotype and allele frequencies of the ACE gene differed significantly between hypertensive and normotensive females (p>0.05), but there were no differences in males. In females, the DBP and MAP level were significantly higher for the DD than for the ID and II genotype, and SBP was significantly higher for the DD than for the II genotype. But in males, there were no significant differences in blood pressure among ACE genotypes. The results showed a significant association between the D allele of the ACE gene and hypertension in Tibetan women but not in Tibetan men. PMID- 12135332 TI - Retention of memory traces in rats subjected to 10 minutes of clinical death. AB - Studies of the effects of 10-min clinical death in mongrel rats, induced by complete circulatory arrest, on the mechanisms of memory, showed that the content of already formed memories ("what to do?"), i.e., the structure of a food procuring habit, motivational activity, and the level of organization of behavior, was completely retained 12 days after resuscitation regardless of how many traces were formed before the experimental procedure. Transient changes seen after clinical death mainly affected the process of performing the habit ("how to do it?"): there was a decrease in the ability to work, along with degradation of performance parameters such as readiness, mobilization activity, and the stability with which memory traces were performed. The rate of recovery of these aspects depended on the initial level of conditioned reflex activity in the rats: the higher this level, the more rapidly the performance parameters returned to the baseline level. It is suggested that these disturbances in the process of reproduction were non-specific in nature and could be due to the post-operative hyperexcitability of the animals. PMID- 12135331 TI - A possible mechanism for the dopamine-evoked synergistic disinhibition of thalamic neurons via the "direct" and "indirect" pathways in the basal ganglia. AB - The mechanism of synaptic plasticity which we have previously proposed for striatal spiny neurons, along with published data on the predominance of dopamine sensitive D1/D2 receptors on strionigral/striopallidal neurons, was used as the basis to propose the hypothesis that the induction of long-term potentiation/depression of the efficiency of the cortical inputs to these cells may result from the excitatory/inhibitory actions of dopamine on the activity of the neurons originating the "direct" and "indirect" pathways through the basal ganglia. Thus, the action of dopamine increases disinhibition of thalamic neurons via the "direct" pathway and decreases their inhibition via the "indirect" pathway. Both effects lead to increases in the activity of thalamic cells and in the activity of the efferent neocortical neurons which they excite. The actions of dopamine on striosomal neurons, which mainly have D1 receptors, may also be to induce long-term potentiation of cortical inputs. This effect should lead to increased inhibition of dopaminergic cells and decreases in their dopamine release, which may promote the maintenance of a stable dopamine concentration in the cortex-basal ganglia-thalamus-cortex neural network. PMID- 12135334 TI - Lateral inhibition as a possible mechanism for evaluating stimulus intensity. AB - This article presents results obtained by simulation modeling of neural networks which show that lateral inhibitory connections between neurons can in principle, when the parameters of these connections are selected appropriately, support the existence in the CNS of detectors for the intensity of an acting signal. A condition for the operation of the proposed detection mechanism is the physiologically demonstrated fact of an inverse proportional relationship between the latency of neuron responses to a threshold stimulus and the intensity of this stimulus. PMID- 12135333 TI - Ca-dependent regulation of the Na-K-pump by post-tetanic sensitization of extrasynaptic cholinoreceptors in common snail neurons. AB - Studies were conducted on the role of the Na,K pump and intracellular calcium in the previously observed phenomenon of increased cholinosensitivity of the somatic membranes of neurons LPa3 and RPa3 in the common snail, induced by electrical tetanic stimulation (10.5 mA. 0.1 sec, 2 impulses/sec; 2 min) of the afferent input (the intestinal nerve). Recordings were made of integral transmembrane acetylcholine-evoked currents by two-electrode membrane voltage clamping. After intracellular administration of the Na,K pump inhibitor ouabain (70 microM), there was no post-tetanic increase in neuron cholinosensitivity. Intracellular injection of the calcium ion chelator BAPTA suppressed the sensitization of somatic cholinoreceptors after orthodromal tetanization. It was concluded that the Na,K pump is involved in the mechanism of post-tetanic potentiation of cholinosensitivity in the bodies of defensive behavior command neurons in the common snail, this involvement being Ca-dependent. PMID- 12135335 TI - The effect of L-tryptophan on conditioned reflex learning and behavior in rats with experimental pathology of the thyroid gland. AB - The effects of L-tryptophan on conditioned reflex learning and behavior were studied in male rats with deficient and excess thyroid hormones levels. The learning process was studied using a model consisting of a conditioned active avoidance response; animal behavior was assessed in an open field test. These studies showed that in conditions of thyroid hormone deficiency. L-tryptophan had positive effects on the acquisition and reproduction of the active avoidance reflex, restoring the ability of the animals to learn actively; L-tryptophan increased investigative activity in the open field test. In conditions of increased thyroid hormone levels, L-tryptophan reversed the mild stimulating effects of thyroid hormones on the processes of developing and performing the active avoidance habit, and increased investigative activity, but decreased the amount of grooming. PMID- 12135336 TI - The effects of xenografts of dopamine-rich ganglia from the immature CNS of the mollusk Helix aspersa L. on learning time during acquisition of an operant food procuring reflex in WAG/Rij rats. PMID- 12135338 TI - Electrical stimulation of the brain in Wilson-Konovalov hepatocerebral dystrophy (a neuromorphological and neurophysiological analysis). AB - Stereotaxic surgery was performed in 27 patients. Complete elimination of or significant reductions in hyperkinesia were obtained in 17 cases; five patients died. There was no correlation between the severity of clinical manifestations of hepatocellular dystrophy and the relatively normal quantitative measures of cortical and subcortical biopotentials, which were produced on a background of microstructural changes affecting neurons in these regions. It is suggested that qualitative significance of these biopotentials is that they carry an excess pathological spike activity resulting in hyperkinesia. This is supported by the fact that hyperkinesia was suppressed after surgical destruction of the ventrolateral nucleus of the thalamus and subthalamic structures. PMID- 12135337 TI - The complex perception of stimuli during the acquisition of conditioned reflexes. PMID- 12135339 TI - Migration of neocortical neuroblasts in rat fetuses during repeated exposure to x rays. PMID- 12135340 TI - Characteristics of the time course of evoked secretion of transmitter quanta in different parts of the motor nerve ending in the frog. AB - Experiments were performed on neuromuscular preparations from frogs, in which three extracellular microelectrodes were used to record nerve ending currents and single-quantum endplate currents simultaneously from the proximal, central, and distal parts of single synaptic contacts. The rate of propagation of excitation across terminals was measured. along with the minimum synaptic delay, the intensity. and the degree of synchronicity of the secretion of transmitter quanta in different parts of the nerve ending, and the relationships between these factors and the calcium ion concentration in the medium. These studies showed that along with gradients in the rate of conduction of excitation and the intensity of secretion in different parts of the ending. there were also differences in the kinetics of the release of transmitter quanta. As the distance from the end of the myelinated part of the axon increased, the rate of conduction of the nerve impulse and the duration of the synaptic delay decreased, while the synchronicity of the release of quanta increased. Increases in the calcium concentration in the medium produced greater increases in the synchronicity of transmitter quantum release in the distal parts of the synapse than in the proximal parts. Mathematical modeling of multiple-quantum endplate currents showed that the characteristics of the kinetics of the secretion process observed here in different parts of the nerve ending represent a factor which partially compensates for the decrease in the amplitude and extending of the duration of the leading front of the multiple-quantum endplate current which are associated with the low rate of conduction of excitation across the nerve ending. The contribution of this compensation increases as the intensity of secretion of transmitter quanta increases in the distal parts of the synaptic contact. PMID- 12135341 TI - Interaction between serotonin and nitric oxide (NO) in the activation of the serotoninergic system in the common snail. AB - The effects of serotonin and NO donors on serotoninergic neurons (more than 60) in the brain of the common snail Helix lucorum were studied. Serotonin and NO donors induced depolarization in all neurons. and increased spike activity and activated the synchronous synaptic input, including train-like input. resulting in the onset of synchronous train activity in all these neurons. The excitatory effect of serotonin was significantly decreased by 5,7-dihydroxytryptamine (5.7 DOT) and monomethylarginine--a blocker of endogenous NO synthesis. Both these substances blocked the serotonin activation of the synchronous train input. 5.7 DOT also blocked the activation of this input by NO donors. but had no effect on their excitatory actions. The effects of 5,7-DOT developed quickly, were reversible, and were comparable to the effects of serotonin receptor antagonists. The data obtained here provide evidence that serotonin and NO have similar regulatory effects on the serotoninergic system in the snail brain. Not only do they excite serotoninergic neurons, but they also coordinate their functioning by activating common synaptic inputs. which are apparently also serotoninergic. It is suggested that NO has the role of a second messenger during serotoninergic excitation and functions as a co-transmitter for the presynaptic input. PMID- 12135342 TI - Analysis of ligand-receptor interactions from the molecular level to the whole body level. AB - A system for the quantitative analysis of ligand-receptor interactions is presented, based on models of different levels of complexity. For two pools of receptors, binding of a radioactive ligand is described by b = [(Bml x A(nl))/(K(nl)dl + A(nl))] + [(Bm2 x A(n2))/(Kn2(d2) + A(n2))], (1) where b is the number of bound receptors at a ligand concentration [A], Bml and Bm2 are the receptor concentrations. Kdl and Kd2 are dissociation constants for the ligand receptor complex, and n1 and n2 are Hill coefficients. The magnitude of the physiological response for a system consisting of two discrete pools of receptors with different affinities is given by p = [(Pm x A(nl))/(EC50(nl) + A(nl))] + [(Pm2 x A(n2)/(EC50(n2)2 + A(n2))], (2) where p is the magnitude of the response to an agonist (or antagonist) at concentration [A], Pml and Pm2 are the maximal magnitudes of the responses for the individual pools of receptors, EC50(1) and EC50(2) are the agonist concentrations giving responses of magnitudes Pm1/2 and Pm2/2, and n1 and n2 are Hill coefficients. The parameters of these equations show: the number of pools of receptors with different affinities for the ligand (Kd or EC50), the number of active receptors (Bmax) or the magnitudes of the maximal response (Pmax), and the numbers of ligand molecules binding with the receptor (n, the Hill coefficient). E is the efficiency (E = Bmax/2Kd, or E = Pmax/2EC50) and gives the overall characteristics of the activity of the effector system. This method of analysis can be applied to any biological reactions whose results can be presented quantitatively. PMID- 12135343 TI - The effects of dopamine synthesis inhibitors and dopamine antagonists on regeneration in the hydra Hydra attenuata. AB - The effects of catecholamine synthesis inhibitors (alpha-methyltyrosine, 3 iodotyrosine, and alpha-methyl-DOPA) and dopamine receptor blockers (haloperidol and spiperone) on the regeneration of apical, gastral, and basal fragments of the hydra Hydra attenuata were studied. These experiments showed that alpha methyltyrosine and 3-iodotyrosine significantly inhibited regeneration but did not produce morphological anomalies. Alpha-Methyl-DOPA produce less inhibition of regeneration, but induced ectopic tentacles and outgrowths in gastral regenerates. Haloperidol and spiperone had no significant effect on the rate of regeneration but induced significant numbers of morphogenetic anomalies in gastral regenerates. Apical and basal regenerates, which retained their natural organizers (the head and base respectively) never yielded morphogenetic anomalies in the presence of either dopamine receptor blockers or dopamine synthesis inhibitors. The possible role of neurotransmitters. particularly dopamine, in morphogenesis in hydras is discussed. PMID- 12135344 TI - Non-dopaminergic neurons expressing dopamine synthesis enzymes: differentiation and functional significance. AB - The development and functional significance of neurons in the arcuate nucleus expressing tyrosine hydroxylase and/or aromatic L-amino acid decarboxylase were studied in rat fetuses, neonates, and adults using immunocytochemical (single and double immunolabeling of tyrosine hydroxylase and aromatic L-amino acid decarboxylase) methods with a confocal microscope and computerized image analysis, HPLC with electrochemical detection, and radioimmunological analysis. Single-enzyme neurons containing tyrosine hydroxylase were first seen on day 18 of embryonic development in the ventrolateral part of the arcuate nucleus. Neurons expressing only aromatic L-amino acid decarboxylase or both enzymes of the dopamine synthesis pathway were first seen on day 20 of embryonic development, in the dorsomedial part of the nucleus. On days 20-21 of embryonic development, dopaminergic (containing both enzymes) neurons amounted to less than 1% of all neurons expressing tyrosine hydroxylase and/or aromatic L-amino acid decarboxylase. Nonetheless, in the ex vivo arcuate nucleus and in primary neuron cultures from this structure, there were relatively high leveLs of dopamine and L dihydroxyphenylalanine (L-DOPA), and these substances were secreted spontaneously and in response to stimulation. In addition. dopamine levels in the arcuate nucleus in fetuses were sufficient to support the inhibitory regulation of prolactin secretion by the hypophysis, which is typical of adult animals. During development, the proportion of dopaminergic neurons increased, reaching 38% in adult rats. Specialized contacts between single-enzyme tyrosine hydroxylase containing and aromatic L-amino acid decarboxylase-containing neurons were present by day 21 of embryonic development; these were probably involved in transporting L-DOPA from the former neurons to the latter. It was also demonstrated that the axons of single-enzyme decarboxylase-containing neurons projected into the median eminence, supporting the secretion of dopamine into the hypophyseal portal circulation. Thus, dopamine is probably synthesized in the arcuate nucleus not only by dopaminergic neurons, but also by neurons expressing only tyrosine hydroxylase or aromatic L-amino acid decarboxylase. PMID- 12135345 TI - Pre- and postsynaptic effects of the calcium channel blocker verapamil at neuromuscular junctions. AB - Experiments on the frog sartorius muscle were used to study the effects of the L type calcium channel blocker verapamil on endplate currents. Verapamil had no effect on the amplitudes of miniature and multiple-quantum endplate currents, the synchronicity of transmitter secretion, or repeat activity in nerve endings. Verapamil had no effect on the decay of miniature currents, but accelerated that of multiple-quantum currents. This effect was sharply increased after inhibition of cholinesterase activity. In conditions of inhibited cholinesterase activity, verapamil depressed currents during rhythmic stimulation. This depression was more marked in synapses with high quantal compositions and in conditions of membrane depolarization. Thus, the sensitivity of neuromuscular junction calcium channels to verapamil was unrelated to the release of transmitter from the motor nerve ending either at physiological levels of secretion or when secretion was potentiated by potassium channel blockers. At the postsynaptic level, the effect of verapamil was insignificant in relation to cholinoreceptors in the resting and active states, though verapamil could cooperatively enhance the transition of postsynaptic receptors into the desensitized state in conditions of prolonged transmitter action. PMID- 12135346 TI - Decreases in Ca2+-dependent K+-currents due to cyclic guanosine monophosphate are not dependent on phosphorylation. AB - Two-microelectrode voltage clamping experiments were performed on isolated snail neurons to measure the Ca2+-dependent. potential-dependent K+ current (I(C)), with assessment of the effects of penetrating cGMP analogs on this current - dibutyryl cGMP (dcGMP) and 8-Br-cGMP. Both of these penetrating cGMP analogs rapidly and reversibly decreased the amplitude of I(C). cGMP analogs produced no shifts in the volt-ampere characteristics of the efflux current along the voltage axis. dcGMP and 8-Br-cGMP had no effect on the influx Ca2+ current. The non specific protein kinase inhibitor H-8 decreased or had no effect on I(C) in different cells. The effects of both dcGMP and 8-Br-cGMP persisted in the presence of H-8. Decreases in I(C) in the presence of cGMP analogs also persisted in the presence of the protein phosphatase inhibitor okadaic acid. These results lead to the conclusion that decreased conductivity of Ca2+-dependent K+ channels occurring in response to cGMP is not associated with phosphorylation. PMID- 12135347 TI - Catalytic mechanism of the topa quinone containing copper amine oxidases. PMID- 12135348 TI - Mechanism of the Escherichia coli ADP-ribose pyrophosphatase, a Nudix hydrolase. AB - Escherichia coli ADP-ribose (ADPR) pyrophosphatase (ADPRase), a Nudix enzyme, catalyzes the Mg(2+)-dependent hydrolysis of ADP-ribose to AMP and ribose 5 phosphate. ADPR hydrolysis experiments conducted in the presence of H(2)(18)O and analyzed by electrospray mass spectrometry showed that the ADPRase-catalyzed reaction takes place through nucleophilic attack at the adenosyl phosphate. The structure of ADPRase in complex with Mg(2+) and a nonhydrolyzable ADPR analogue, alpha,beta-methylene ADP-ribose, reveals an active site water molecule poised for nucleophilic attack on the adenosyl phosphate. This water molecule is activated by two magnesium ions, and its oxygen contacts the target phosphorus (P-O distance of 3.0 A) and forms an angle of 177 degrees with the scissile bond, suggesting an associative mechanism. A third Mg(2+) ion bridges the two phosphates and could stabilize the negative charge of the leaving group, ribose 5 phosphate. The structure of the ternary complex also shows that loop L9 moves fully 10 A from its position in the free enzyme, forming a tighter turn and bringing Glu 162 to its catalytic position. These observations indicate that as part of the catalytic mechanism, the ADPRase cycles between an open (free enzyme) and a closed (substrate-metal complex) conformation. This cycling may be important in preventing nonspecific hydrolysis of other nucleotides. PMID- 12135349 TI - First non-alpha-amino acid guanidines acting as efficient NO precursors upon oxidation by NO-synthase II or activated mouse macrophages. AB - A study of the oxidation of a series of guanidines related to L-arginine (L-Arg) and of various alkyl- and arylguanidines, by recombinant NO-synthase II (NOS II), led us to the discovery of the first non-alpha-amino acid guanidine substrate of NOS, acting as an efficient NO precursor. This compound, 3 (trifluoromethyl)propylguanidine, 4, led to a rate of NO formation (k(cat) = 220 +/- 50 min(-1)) only 2 times lower than that of L-Arg. Formation of 1 mol of NO upon NOS II-catalyzed oxidation of 4 occurred with consumption of 2.9 mol of NADPH, which corresponds to a 52% coupling between electron transfer and oxygenation of its guanidine function. Its oxidation by activated mouse macrophages in an L-Arg-free medium resulted in NO(2)(-) formation that was inhibited by classical NOS inhibitors with a rate only 2-3 times lower than that observed with L-Arg itself. These results open the way toward the research of selective, stable guanidine substrates of NOS that could be interesting, new NO donors after in situ oxidation by a given NOS isoform. PMID- 12135350 TI - Generation and characterization of two transgenic mouse lines expressing human ApoE2 in neurons and glial cells. AB - Apolipoprotein E (apoE) isoforms are key determinants of susceptibility to late onset Alzheimer's disease (AD). The epsilon 4 and epsilon 2 alleles have been associated with increased and decreased risk for AD, respectively. We have generated and characterized transgenic mice in which the human apoE2 gene is expressed under the control of the platelet-derived growth factor B-chain (PDGF B) promoter, or the transferrin (TF) promoter. S1 nuclease analysis and immunoblotting showed that the PDGF-B apoE2 mice express apoE2 exclusively in the brain whereas the TF apoE2 mice express apoE2 in the liver and in the brain. In the TF apoE2 mouse line, apoE2 is also detected in the plasma. The PDGF-B apoE2 and the TF apoE2 transgenic mice were bred back to apoE(-)(/)(-) background. Immunohistochemical analysis showed that the PDGF apoE2 x apoE(-)(/)(-) and the TF apoE2 x apoE(-)(/)(-) mice express human apoE2 within the neocortex in hippocampal neurons and glial cells, respectively. ApoE(-)(/)(-) mice have been shown to develop age-dependent loss of synaptophysin. Immunoblotting of mouse brain extracts and immunohistochemical analysis of brain sections showed that apoE expression in both apoE2 x apoE(-)(/)(-) transgenic lines was associated with significant recovery of brain synaptophysin levels as compared to the levels of apoE(-)(/)(-) littermates of the same age. These apoE2-expressing mice, when bred back on amyloid precursor protein (APP) transgenic mice or other mouse lines featuring alterations in lipoprotein metabolism, may provide new mouse models for elucidating the role of apoE2 in lipid homeostasis in the brain and in the pathogenesis of AD. PMID- 12135351 TI - Role of zymogenicity-determining residues of coagulation factor VII/VIIa in cofactor interaction and macromolecular substrate recognition. AB - Factor VIIa (VIIa) remains in a zymogen-like state following proteolytic activation and depends on interactions with the cofactor tissue factor (TF) for function. Val(21), Glu(154), and Met(156) are residues that are spatially close in available zymogen and enzyme structures, despite major conformational differences in the corresponding loop segments. This residue triad displays unusual side chain properties in comparison to the properties of other coagulation serine proteases. By mutagenesis, we demonstrate that these residues cooperate to stabilize the enzyme conformation and to enhance the affinity for TF. In zymogen VII, however, substitution of the triad did not change the cofactor affinity, further emphasizing the crucial role of the activation pocket in specifically stabilizing the active enzyme conformation. In comparison to VIIa(Q156), the triple mutant VIIa(N21I154Q156) had a stabilized amino-terminal Ile(16)-Asp(194) salt bridge and enhanced catalytic function. However, proteolytic and amidolytic activities of free VIIa variants were not concordantly increased. Rather, a negatively charged Asp at position 21 was the critical factor that determined whether an amidolytically more active VIIa variant also more efficiently activated the macromolecular substrate. These data thus demonstrate an unexpected complexity by which the zymogenicity-determining triad in the activation pocket of VIIa controls the active enzyme conformation and contributes to exosite interactions with the macromolecular substrate. PMID- 12135352 TI - Evidence for a copper-binding superfamily of the amyloid precursor protein. AB - The amyloid precursor protein (APP) copper-binding domain (CuBD) has been shown to reduce Cu(II) to Cu(I) and to mediate copper-induced oxidation in vitro. However, little is known about copper binding to the homologous domains of APP and APP family paralogs and orthologs (including amyloid precursor-like proteins from Drosophila melanogaster, Xenopus laevis, and Caenorhabditis elegans) and their effects on Cu-induced oxidation and Cu(I) formation. Here, we show that APP homologues with and without conserved histidine residues at positions 147, 149, and 151 all bind Cu(II). Oxidized peptides were the kinetically favored products of the redox reaction of CuBDs promoting the reduction of Cu(II) to Cu(I). These results reveal a molecular phylogeny-based divergence that has taken place between the ancestral Drosophila APPL and C. elegans APL-1 and the recently evolved APP lineage of CuBDs. Whereas higher species CuBDs have a decreased affinity for Cu(II) and high Cu(II) reducing activities, ancestral CuBDs form very tight binding sites for Cu(II) ions and have low Cu(II) reducing activities. Thus, the APP lineage displays a gain in activity toward promoting Cu(II) reduction and Cu(I) release. The findings suggests that the Cu(II)-binding equilibrium at the phylogenetic stage of Drosophila APPL and C. elegans APL-1 is shifted from the exchangeable Cu(II) pool to the tightly bound, nonexchangeable pool and that ancestral CuBDs may exert antioxidation activities in vivo. The more recently evolved homologues of human APP appear to take advantage of unique redox properties for yet unknown biological functions. PMID- 12135353 TI - Transmembrane interactions in the activation of the Neu receptor tyrosine kinase. AB - The Neu receptor tyrosine kinase is constitutively activated by a single amino acid change in the transmembrane domain of the receptor. The mutation of Val664 to glutamate or glutamine induces receptor dimerization and autophosphorylation of the receptor's intracellular kinase domain. The ability of this single mutation to activate the receptor is sequence-dependent, suggesting that specific helix-helix interactions stabilize the transmembrane dimer. We have determined the local secondary structure and interhelical contacts in the region of position 664 in peptide models of the activated receptor using solid-state rotational resonance and rotational echo double-resonance (REDOR) NMR methods. Intrahelical (13)C rotational resonance distance measurements were made between 1-(13)C-Thr662 and 2-(13)C-Gly665 on peptides corresponding to the wild-type Neu and activated Neu transmembrane sequences containing valine and glutamate at position 664, respectively. We observed similar internuclear distances (4.5 +/- 0.2 A) in both Neu and Neu*, indicating that the region near residue 664 is helical and is not influenced by mutation. Interhelical (15)N...(13)C REDOR measurements between Gln664 side chains on opposing helices were not consistent with hydrogen bonding between the side chain functional groups. However, interhelical rotational resonance measurements between 1-(13)C-Glu664 and 2-(13)C-Gly665 and between 1 (13)C-Gly665 and 2-(13)C-Gly665 demonstrated close contacts (4.3-4.5 A) consistent with the packing of Gly665 in the Neu* dimer interface. These measurements provide structural constraints for modeling the transmembrane dimer and define the rotational orientation of the transmembrane helices in the activated receptor. PMID- 12135354 TI - Nuclear import pathway of the telomere elongation suppressor TRF1: inhibition by importin alpha. AB - Telomere repeat factor 1 (TRF1) regulates the steady-state length of chromosomes, whereby its overexpression results in telomere shortening while dominant negative TRF1 mutations can lead to telomere elongation, which is linked to cell immortalization/transformation. Although present in the nucleus at mammalian chromosomal ends during interphase and mitosis, nothing is known of the mechanism by which TRF1 enters the nucleus or how its nuclear levels may be regulated and the relevance of this, in turn, to telomere length and cell immortalization. Here we examine the nuclear import mechanism of TRF by expressing and purifying a recombinant TRF1-GFP (green fluorescent protein) fusion protein that is functional in terms of being able to bind telomeric DNA specifically as shown using a novel, quantitative double-label gel mobility shift assay. We quantitate the ability of TRF1-GFP to accumulate in the nucleus using real time confocal laser scanning microscopy, showing that the nuclear import pathway of TRF1 is mediated by importin (Imp) beta1 and Ran. Imp beta is shown to bind directly to TRF1 with nanomolar affinity using native gel electrophoretic and fluorescence polarization (FP) approaches; FP experiments also demonstrate that Imp beta residues 1-380 are responsible for TRF1 binding. Intriguingly, when dimerized to Imp beta, Imp alpha was found to inhibit Imp beta-mediated nuclear accumulation, although not affecting Imp beta binding to TRF1. The study represents the first elucidation of the nuclear transport mechanism of TRF1; that its nuclear import is mediated directly by Imp beta but inhibited by Imp alpha may represent a novel regulatory mechanism, with potential relevance to oncogenesis. PMID- 12135355 TI - Intercalation of an acridine-peptide drug in an AA/TT base step in the crystal structure of [d(CGCGAATTCGCG)](2) with six duplexes and seven Mg(2+) ions in the asymmetric unit. AB - We present the crystal structure of an acridine drug derivatized at carbon 9, [N(alpha)-(9-acridinoyl)-tetraarginine], intercalated within the dodecamer [d(CGCGAATTCGCG)](2). The presence of a lateral chain at the central carbon 9 atom differentiates this compound from most acridine drugs hitherto studied, which are usually derivatized at carbon 4. The DNA:drug interaction we observe differs from that observed in previous studies, which primarily involves shorter, mainly hexameric sequences, in two important regards: the acridine intercalates within an AA/TT base step, rather than within a CG/CG base step; and the binding site is located at the center of the sequence, rather than at one end of the duplex. In addition, we observe a novel crystal packing arrangement, with six dodecamer duplexes and seven hydrated magnesium ions in the asymmetric unit of a large (66.5 x 68.4 x 77.4 A(3)) unit cell in space group P2(1)2(1)2(1). The duplexes are organized in layers parallel to the ab plane, with consecutive layers crossing each other at right angles. PMID- 12135356 TI - Structural requirements for the intracellular subunit polymerization of the complement inhibitor C4b-binding protein. AB - C4b-binding protein (C4BP), an important inhibitor of complement activation, has a unique spider-like shape. It is composed of six to seven identical alpha-chains with or without a single beta-chain, the chains being linked by disulfide bridges in their C-terminal parts. To elucidate the structural requirements for the assembly of the alpha-chains, recombinant C4BP was expressed in HEK 293 cells. The expressed C4BP was found to contain six disulfide-linked alpha-chains. Pulse chase analysis demonstrated that the recombinant C4BP was rapidly synthesized in the cells and the polymerized C4BP appeared in the medium after 40 min. The alpha chains were polymerized in the endoplasmic reticulum (ER) already after 5 min chase. The polymerization process was unaffected by blockage of the transport from the ER to the Golgi mediated by brefeldin A or low temperature (10 degrees C). The C-terminal part of the alpha-chain (57 amino acids), containing 2 cysteine residues and an amphiphatic alpha-helix region, was required for the polymerization. We constructed and expressed several mutants of C4BP that lacked the cysteine residues and/or were truncated at various positions in the C terminal region. Gel filtration analysis of these variants demonstrated the whole alpha-helix region to be required for the formation of stable polymers of C4BP, which were further stabilized by the formation of disulfide bonds. PMID- 12135357 TI - The active site loop of S-adenosylmethionine synthetase modulates catalytic efficiency. AB - Crystallographic studies of Escherichia coli S-adenosylmethionine synthetase (ATP:L-methionine S-adenosyltransferase, MAT) have defined a flexible polypeptide loop that can gate access to the active site without contacting the substrates. The influence of the length and sequence of this active site loop on catalytic efficiency has been characterized in a mutant in which the E. coli MAT sequence (DRADPLEQ) has been replaced with the distinct sequence of the corresponding region of the otherwise highly homologous rat liver enzyme (HDLRNEEDV). Four additional mutants in which the entire DRADPLEQ sequence was replaced by five, six, seven, or eight glycines have been studied to unveil the effects of loop length and the influence of side chains. In all of the mutants, the maximal rate of S-adenosylmethionine formation (k(cat)) is diminished by more than 200-fold whereas the rate of hydrolysis of the tripolyphosphate intermediate is decreased by less than 3-fold. Thus, the function of the loop is localized to the first step in the overall reaction. The K(m) for methionine increases in all of the oligoglycine mutants, whereas the K(m) values for ATP are not substantially different. The k(cat) for the wild-type enzyme is decreased by increases in solution microviscosity with 55% of the maximal dependence. Thus, a diffusional event is coupled to the chemical step of AdoMet formation, which is known to be rate-limiting. The results indicate that a conformational change, possibly loop closure, is associated with AdoMet synthesis. The data integrate a previously discovered conformational change associated with PPP(i) binding to the E x AdoMet complex into the reaction sequence, reflecting a difference in protein conformation in the E x AdoMet x PPP(i) complex whether it is formed from the E x ATP x methionine complex or from binding of exogenous PPP(i). The temperature dependence of the k(cat) for S-adenosylmethionine formation shows that the removal of the side chains in the glycine mutants causes the activation enthalpy of the reaction to approximately double in each case, while the activation entropy changes from negative in the wild-type enzyme to positive in the mutants. The favorable activation entropy in the mutant-catalyzed reactions may reflect release of water during catalysis, while the negative activation entropy in the reaction catalyzed by the wild-type enzyme apparently reflects reorganization of the loop. The observations point to how nature can fine-tune the activity of an enzyme by modifying substrate and product access to the active site rather than by altering the enzyme x substrate contacts or the catalytic machinery itself. PMID- 12135358 TI - Structural insights into the beta-mannosidase from T. reesei obtained by synchrotron small-angle X-ray solution scattering enhanced by X-ray crystallography. AB - A molecular envelope of the beta-mannosidase from Trichoderma reesei has been obtained by combined use of solution small-angle X-ray scattering (SAXS) and protein crystallography. Crystallographic data at 4 A resolution have been used to enhance informational content of the SAXS data and to obtain an independent, more detailed protein shape. The phased molecular replacement technique using a low resolution SAXS model, building, and refinement of a free atom model has been employed successfully. The SAXS and crystallographic free atom models exhibit a similar globular form and were used to assess available crystallographic models of glycosyl hydrolases. The structure of the beta-galactosidase, a member of a family 2, clan GHA glycosyl hydrolases, shows an excellent fit to the experimental molecular envelope and distance distribution function of the beta mannosidase, indicating gross similarities in their three-dimensional structures. The secondary structure of beta-mannosidase quantified by circular dichroism measurements is in a good agreement with that of beta-galactosidase. We show that a comparison of distance distribution functions in combination with 1D and 2D sequence alignment techniques was able to restrict the number of possible structurally homologous proteins. The method could be applied as a general method in structural genomics and related fields once protein solution scattering data are available. PMID- 12135359 TI - Solution structure and dynamics of the human-Escherichia coli thioredoxin chimera: insights into thermodynamic stability. AB - We have determined the high-resolution solution structure of the oxidized form of a chimeric human and Escherichia coli thioredoxin (TRX(HE)) by NMR. The overall structure is well-defined with a rms difference for the backbone atoms of 0.27 +/ 0.06 A. The topology of the protein is identical to those of the human and E. coli parent proteins, consisting of a central five-stranded beta-sheet surrounded by four alpha-helices. Analysis of the interfaces between the two domains derived from the human and E. coli sequences reveals that the general hydrophobic packing is unaltered and only subtle changes in the details of side chain interactions are observed. The packing of helix alpha(4) with helix alpha(2) across the hybrid interface is less optimal than in the parent molecules, and electrostatic interactions between polar side chains are missing. In particular, lysine glutamate salt bridges between residues on helices alpha(2) and alpha(4), which were observed in both human and E. coli proteins, are not present in the chimeric protein. The origin of the known reduced thermodynamic stability of TRX(HE) was probed by mutagenesis on the basis of these structural findings. Two mutants of TRX(HE), S44D and S44E, were created, and their thermal and chemical stabilities were examined. Improved stability toward chaotropic agents was observed for both mutants, but no increase in the denaturation temperature was seen compared to that of TRX(HE). In addition to the structural analysis, the backbone dynamics of TRX(HE) were investigated by (15)N NMR relaxation measurements. Analysis using the model free approach reveals that the protein is fairly rigid with an average S(2) of 0.88. Increased mobility is primarily present in two external loop regions comprising residues 72-74 and 92-94 that contain glycine and proline residues. PMID- 12135360 TI - Structural origins of the functional divergence of human insulin-like growth factor-I and insulin. AB - Human insulin-like growth factors I and II (hIGF-I, hIGF-II) are potent stimulators of cell and growth processes. They display high sequence similarity to both the A and B chains of insulin but contain an additional connecting C domain, which reflects their secretion without specific packaging or precursor conversion. IGFs also have an extension at the C-terminus known as the D-domain. This paper describes four homologous hIGF-1 structures, obtained from crystals grown in the presence of the detergent SB12, which reveal additional detail in the C- and D-domains. Two different detergent binding modes observed in the crystals may reflect different hIGF-I biological properties such as the interaction with IGF binding proteins and self-aggregation. While the helical core of hIGF-I is very similar to that in insulin, there are distinct differences in the region of hIGF-I corresponding to the insulin B chain C-terminus, residues B25-B30. In hIGF-I, these residues (24-29) and the following C-domain form an extensive loop protruding 20 A from the core, which results in a substantially different conformation for the receptor binding epitope in hIGF-I compared to insulin. One notable feature of the structures presented here is demonstration of peptide-bond cleavage between Ser35 and Arg36 resulting in an apparent gap between residues 35 and 39. The equivalent region of proinsulin is involved in hormone processing demanding a reassessment of the structural integrity of hIGF-I in relation to its biological function. PMID- 12135361 TI - Lipid-protein interactions in lipovitellin. AB - The refined molecular structure of lipovitellin is described using synchrotron cryocrystallographic data to 1.9 A resolution. Lipovitellin is the predominant lipoprotein found in the yolk of egg-laying animals and is involved in lipid and metal storage. It is thought to be related in amino acid sequence to segments of apolipoprotein B and the microsomal transfer protein responsible for the assembly of low-density lipoproteins. Lipovitellin contains a heterogeneous mixture of about 16% (w/w) noncovalently bound lipid, mostly phospholipid. Previous X-ray structural studies at ambient temperature described several different protein domains including a large cavity in each subunit of the dimeric protein. The cavity was free of any visible electron density for lipid molecules at room temperature, suggesting that only dynamic interactions exist with the protein. An important result from this crystallographic study at 100 K is the appearance of some bound ordered lipid along the walls of the binding cavity. The precise identification of the lipid type is difficult because of discontinuities in the electron density. Nonetheless, the conformations of 7 phospholipids and 43 segments of hydrocarbon chains greater than 5 atoms in length have been discovered. The conformations of the bound lipid and the interactions between protein and lipid provide insights into the factors governing lipoprotein formation. PMID- 12135362 TI - Crystal structure of D-hydantoinase from Bacillus stearothermophilus: insight into the stereochemistry of enantioselectivity. AB - Industrial production of antibiotics, such as semisynthetic penicillins and cephalosporins, requires optically pure D-p-hydroxylphenylglycine and its derivatives as important side-chain precursors. To produce optically pure D-amino acids, microbial D-hydantoinase (E.C. 3.5.2.2) is used for stereospecific hydrolysis of chemically synthesized cyclic hydantoins. We report the apo-crystal structure of D-hydantoinase from B. stearothermophilus SD1 at 3.0 A resolution. The structure has a classic TIM barrel fold. Despite an undetectable similarity in sequence, D-hydantoinase shares a striking structural similarity with the recently solved structure of dihydroorotase. A structural comparison of hydantoinase with dihydroorotase revealed that the catalytic chemistry is conserved, while the substrate recognition is not. This structure provides insight into the stereochemistry of enantioselectivity in hydrolysis and illustrates how the enzyme recognizes stereospecific exocyclic substituents and hydrolyzes hydantoins. It should also provide a rationale for further directed evolution of this enzyme for hydrolysis of new hydantoins with novel exocyclic substituents. PMID- 12135363 TI - Kinetic characterization of the peptidase activity of Escherichia coli Lon reveals the mechanistic similarities in ATP-dependent hydrolysis of peptide and protein substrates. AB - Lon is an ATP-dependent protease that degrades unstructured proteins. In this study, we have examined the ATP dependency of Escherichia coli Lon catalyzing the hydrolysis of a defined fluorogenic peptide known as S3. Steady-state velocity analyses of S3 degradation in the presence of ATP, or the nonhydrolyzable ATP analogue AMPPNP, indicate a sequential mechanism, and the k(cat) of the reaction was 7-fold higher in the presence of ATP. Comparing the pre-steady-state time courses of the ATP- versus AMPPNP-mediated S3 hydrolysis reveals that ATP hydrolysis accelerates a slow step before the chemical cleavage of peptide. Product inhibition studies indicate that ADP is competitive versus ATP but noncompetitive versus the S3 substrate. In the absence of S3, Lon exhibits a 10 20-fold higher affinity for ADP than ATP. However the S3 substrate weakens the affinity of Lon for ADP by 7-19-fold, indicating that this peptide also promotes ADP/ATP exchange in Lon similar to that observed with protein substrates. The hydrolyzed peptide product, Pd1, exhibited noncompetitive inhibition versus both ATP and S3 substrates. Together with the small change in the K(i) of Pd1 at increasing S3 concentrations, the Pd1 inhibition data support the existence of an isomechanism in Lon catalyzing the hydrolysis of S3 in the presence of ATP or AMPPNP. Upon the basis of the collected data, an extended kinetic mechanism is proposed for the ATP-dependent peptidase mechanism of Lon. PMID- 12135364 TI - Evolution of the allosteric ligand sites of mammalian phosphofructo-1-kinase. AB - Mammalian phosphofructokinase (PFK) has evolved by a process of tandem gene duplication and fusion to yield a protein that is more than double the size of prokaryotic PFKs. On the basis of complete conservation of active site residues in the N-terminal half of the eukaryotic enzyme with those of the bacterial PFKs, one assumes that the active site of the eukaryotic PFK is located in the N terminal half. Again using sequence comparisons, the four allosteric ligand sites of mammalian PFK have been thought to arise from the duplicated catalytic and regulatory sites of the ancestral PFK. Previous site-directed mutagenesis studies [Li et al. (1999) Biochemistry 38, 16407-16412; Chang and Kemp (2002) Biochem. Biophys. Res. Commun. 290, 670-675] have identified the origins of the citrate and fructose 2,6-bisphosphate sites. Here, site-directed mutagenesis of two arginine residues (Arg-433 and Arg-429) of mouse phosphofructokinase is used to identify the ATP inhibitory site, and, by inference, the AMP/ADP site. Mutation of the residues to alanine reduced ATP inhibition in the case of Arg-429 and eliminated ATP inhibition in the instance of Arg-433. The Arg-433 mutant could be inhibited by citrate, and that inhibition could be reversed by fructose 2,6 bisphosphate and cyclic AMP, a high-affinity ligand for the AMP/ADP binding site. It is concluded that the two inhibitors, ATP and citrate, of mammalian PFK interact with sites that have evolved from the duplicated phosphoenolpyruvate/ADP allosteric site of the ancestral PFK. The two sites for activators, fructose 2,6 bisphosphate and AMP or ADP, have evolved from the catalytic site of the ancestral precursor. PMID- 12135365 TI - Characterization of chimeric ADPglucose pyrophosphorylases of Escherichia coli and Agrobacterium tumefaciens. Importance of the C-terminus on the selectivity for allosteric regulators. AB - ADPglucose pyrophosphorylase catalyzes the regulatory step in the pathway for bacterial glycogen synthesis. The enzymes from different organisms exhibit distinctive regulatory properties related to the main carbon metabolic pathway. Escherichia coli ADPglucose pyrophosphorylase is mainly activated by fructose 1,6 bisphosphate (FBP), whereas the Agrobacterium tumefaciens enzyme is activated by fructose 6-phosphate (F6P) and pyruvate. Little is known about the regions determining the specificity for the allosteric regulator. To study the function of different domains, two chimeric enzymes were constructed. "AE" contains the N terminus (271 amino acids) of the A. tumefaciens ADPglucose pyrophosphorylase and the C-terminus (153 residues) of the E. coli enzyme, and "EA", the inverse construction. Expression of the recombinant wild-type and chimeric enzymes was performed using derivatives of the pET24a plasmid. Characterization of the purified chimeric enzymes showed that the C-terminus of the E. coli enzyme is relevant for the selectivity by FBP. However, this region seems to be less important for the specificity by F6P in the A. tumefaciens enzyme. The chimeric enzyme AE is activated by both FBP and F6P, neither of which affect EA. Pyruvate activates EA with higher apparent affinity than AE, suggesting that the C terminus of the A. tumefaciens enzyme plays a role in the binding of this effector. The allosteric inhibitor site is apparently disrupted, as a marked desensitization toward AMP was observed in the chimeric enzymes. PMID- 12135366 TI - Differential effect of copper (II) on the cytochrome P450 enzymes and NADPH cytochrome P450 reductase: inhibition of cytochrome P450-catalyzed reactions by copper (II) ion. AB - Inhibitory effects of Cu(2+) on the cytochrome P450 (P450)-catalyzed reactions of liver microsomes and reconstituted systems containing purified P450 and NADPH P450 reductase (NPR) were seen. However, Zn(2+), Mg(2+), Mn(2+), Ca(2+), and Co(2+) had no apparent effects on the activities of microsomal P450s. Cu(2+) inhibited the reactions catalyzed by purified P450s 1A2 and 3A4 with IC(50) values of 5.7 and 8.4 microM, respectively. Cu(2+) also inhibited reduction of cytochrome c by NPR (IC(50) value of 5.8 microM). Copper caused a decrease in semiquinone levels of NPR, although it did not disturb the rate of formation of semiquinone. P450 reactions supported by an oxygen surrogate, tert-butyl hydroperoxide, instead of NPR and NADPH, were inhibited by the presence of Cu(2+). The results indicate that Cu(2+) inhibits the P450-catalyzed reactions by affecting both P450s and NPR. It was also found that the inhibition of catalytic activities of P450s by Cu(2+) involves overall conformational changes of P450s and NPR, investigated by CD and intrinsic fluorescence spectroscopy. These results suggest that the inhibitory effect of Cu(2+) on the P450-catalyzed reactions may come from the inability of an efficient electron transfer from NPR to P450 and also the dysfunctional conformation of NPR and P450. PMID- 12135367 TI - Differential effect of a his tag at the N- and C-termini: functional studies with recombinant human serum transferrin. AB - Attachment of a cleavable hexa His tag is a common strategy for the production of recombinant proteins. Production of two recombinant nonglycosylated human serum transferrins (hTF-NG), containing a factor Xa cleavage site and a hexa His tag at the carboxyl terminus, has been described [Mason et al. (2001) Prot. Exp. Purif 23, 142-150]. More recently, hTF-NG with an amino-terminal His tag and a factor Xa cleavage site has been expressed (>30 mg/L) in baby hamster kidney cells and purified from the tissue culture medium. Although it is frequently assumed that addition of a His tag has little or no effect on function, this is not always confirmed experimentally. In the present study, in vitro quantitative data clearly shows that the presence of the C-terminal His tag has an effect on the release of iron from recombinant hTF at pH 7.4 and 5.6. Measurement of the rate of release from both the N- and C-lobes is reduced 2-4-fold. These findings provide further compelling evidence that the two lobes communicate with each other and highlight the importance of the C-terminal portion of the C-terminal lobe in this interaction. In contrast to these results, we demonstrate that the presence of a His tag at the N-terminus of hTF has no effect on the rate of iron release from either lobe. In a competition experiment, both unlabeled N- and C terminal His-tagged constructs were equally effective at inhibiting the binding of radio-iodinated diferric glycosylated hTF from a commercial source to receptors on HeLa cells as the unlabeled recombinant diferric hTF-NG control. Thus, the presence of a His tag at either the N- or C-terminus of hTF-NG has no apparent effect on the ability of these hTF species to bind to transferrin receptors. PMID- 12135368 TI - Secondary and quaternary structures of the (+)-pinoresinol-forming dirigent protein. AB - The (+)-pinoresinol-forming dirigent protein is the first protein capable of stereoselectively coupling two coniferyl alcohol derived radical species, in this case to give the 8-8' linked (+)-pinoresinol. Only dimeric cross-linked dirigent protein structures were isolated when 1-ethyl-3-[3-(dimethylamino) propyl]carbodiimide was used as cross-linking agent, whereas the associated oxidase, presumed to generate the corresponding free radical substrate, was not detected. Native Forsythia intermedia dirigent protein isoforms were additionally subjected to MALDI-TOF and ESI-MS analyses, which established the presence of both monomeric masses of 23-25 kDa and dimeric dirigent protein species ranging from 46 to 49 kDa. Analytical ultracentrifugation, sedimentation velocity, and sedimentation equilibrium analyses of the native dirigent protein in open solution confirmed further its dimeric nature as well as a propensity to aggregate, with the latter being dependent upon both temperature and solution ionic strength. Circular dichroism analysis suggested that the dirigent protein was primarily composed of beta-sheet and loop structures. PMID- 12135369 TI - The tumor necrosis factor-alpha converting enzyme (TACE): a unique metalloproteinase with highly defined substrate selectivity. AB - TNF alpha converting enzyme (TACE) processes precursor TNF alpha between Ala76 and Val77, yielding a correctly processed bioactive 17 kDa protein. Genetic evidence indicates that TACE may also be involved in the shedding of other ectodomains. Here we show that native and recombinant forms of TACE efficiently processed a synthetic substrate corresponding to the TNF alpha cleavage site only. For all other substrates, conversion occurred only at high enzyme concentrations and prolonged reaction times. Often, cleavage under those conditions was accompanied by nonspecific reactions. We also compared TNF alpha cleavage by TACE to cleavage by those members of the matrix metalloproteinase (MMP) family previously implied in TNF alpha release. The specificity constants for TNF alpha cleavage by the MMPs were approximately 100-1000-fold slower relative to TACE. MMP 7 also processed precursor TNF alpha at the correct cleavage site but did so with a 30-fold lower specificity constant relative to TACE. In contrast, MMP 1 processed precursor TNF alpha between Ala74 and Gln75, in addition to between Ala76 and Val77, while MMP 9 cleaved this natural substrate solely between Ala74 and Gln75. Additionally, the MMP substrate Dnp PChaGC(Me)HK(NMA)-NH(2) was not cleaved at all by TACE, while collagenase (MMP 1), gelatinase (MMP 9), stromelysin 1 (MMP 3), and matrilysin (MMP 7) all processed this substrate efficiently. All of these results indicate that TACE is unique in terms of its specificity requirements for substrate cleavage. PMID- 12135370 TI - Activation-induced subcellular redistribution of G alpha(s) is dependent upon its unique N-terminus. AB - The heterotrimeric G protein subunit, alpha(s), can move reversibly from plasma membranes to cytoplasm in response to activation by GPCRs or activating mutations. We examined the importance of the unique N-terminus of alpha(s) in this translocation in cultured cells. alpha(s) contains a single site for palmitoylation in its N-terminus, and this was replaced by different plasma membrane targeting motifs. These N-terminal alpha(s) mutants were targeted properly to plasma membranes, capable of coupling activated GPCRs to effectors, and able to constitutively stimulate cAMP production when they also contained an activating mutation. However, when activated by a constitutively activating mutation or by agonist-activated beta-AR, these N-terminal alpha(s) mutants failed, for the most part, to undergo redistribution from plasma membranes to cytoplasm, as assayed by immunofluorescence microscopy, or from a particulate to soluble fraction, as assayed by subcellular fractionation. These results highlight the importance of the extreme N-terminus of alpha(s) and its single site of palmitoylation for facilitating activation-induced translocation and provide insight into the mechanism of this G protein trafficking event. PMID- 12135371 TI - Peptides derived from BH3 domains of Bcl-2 family members: a comparative analysis of inhibition of Bcl-2, Bcl-x(L) and Bax oligomerization, induction of cytochrome c release, and activation of cell death. AB - Overexpression of Bcl-2, an anti-apoptotic oncoprotein, is commonly observed in a variety of human malignancies and is associated with resistance to chemotherapy and radiotherapy. Although the precise mechanism of Bcl-2 action remains elusive, current evidence indicates that Bcl-2 inhibits apoptosis by binding and inhibiting pro-apoptotic molecules such as Bax. Therefore, agents that disrupt the ability of Bcl-2, or other anti-apoptotic molecules, to bind to pro-apoptotic molecules may have therapeutic value. Several studies have shown that the BH3 domains of Bcl-2 and Bax are critically important for Bax/Bcl-2 heterodimerization. In this report, we designed and synthesized peptides based on the BH3 domains of three distinct Bcl-2 family members, Bcl-2, Bax and Bad. In vitro interaction assays were used to compare the abilities of the different peptides to inhibit Bax/Bcl-2 and Bax/Bcl-x(L) heterodimerization, as well as Bcl 2 and Bax homodimerization. Bax BH3 peptide (20-amino acids) potently inhibited both Bax/Bcl-2 and Bax/Bcl-x(L) interactions, exhibiting IC(50) values of 15 and 9.5 microM, respectively. The Bad BH3 peptide (21 amino acids) was slightly more potent than Bax BH3 at inhibiting Bax/Bcl-x(L) but failed to disrupt Bax/Bcl-2. Bcl-2 BH3 peptide (20-amino acids) was inactive toward Bax/Bcl-2 and had only a weak inhibitory effect on Bax/Bcl-x(L) heterodimerization. All three BH3 peptides failed to significantly inhibit homodimerization of Bcl-2 or Bax. Consistent with its ability to disrupt Bax/Bcl-2 heterodimerization, Bax BH3 peptide was able to overcome Bcl-2 overexpression and induce cytochrome c release from mitochondria of Bcl-2-overexpressing Jurkat T leukemic cells. Bad BH3 peptide, while potently inducing cytochrome c release in wild-type Jurkat cells, only partially overcame the effects of Bcl-2 overexpression. Bcl-2 BH3 failed to induce cytochrome c release, even in wild-type cells. Delivery of the Bax BH3 and Bad BH3 peptides into wild-type Jurkat cells induced comparable levels of cell death. In cells overexpressing Bcl-2, the potency of Bax BH3 peptide was similar to that seen in wild-type cells, while the efficacy of Bad BH3 peptide was reduced. By contrast, in Bcl-x(L)-overexpressing cells, Bad BH3 exhibited greater cell-killing activity than Bax BH3. The Bcl-2 BH3 peptide and a mutant Bax BH3 peptide had no appreciable effect on Jurkat cells. Together, our data suggest that agents based on the Bax BH3 domain may have therapeutic value in cancers overexpressing Bcl-2, while agents based on the BH3 domain of Bad may be more useful for tumors overexpressing Bcl-x(L). PMID- 12135372 TI - Mechanism of the rate-determining step of the Na(+),K(+)-ATPase pump cycle. AB - The kinetics of the E(2) --> E(1) conformational change of unphosphorylated Na(+),K(+)-ATPase from rabbit kidney and shark rectal gland were investigated via the stopped-flow technique using the fluorescent label RH421 (pH 7.4, 24 degrees C). The enzyme was pre-equilibrated in a solution containing 25 mM histidine and 0.1 mM EDTA to stabilize initially the E(2) conformation. When rabbit kidney enzyme was mixed with NaCl alone, tris ATP alone or NaCl, and tris ATP simultaneously, a fluorescence decrease was observed. The reciprocal relaxation time, 1/tau, of the fluorescent transient was found to increase with increasing NaCl concentration and reached a saturating value in the presence of 1 mM tris ATP of 54 +/- 3 s(-1) in the case of rabbit kidney enzyme. The experimental behavior could be described by a binding of Na(+) to the enzyme in the E(2) state with a dissociation constant of 31 +/- 7 mM, which induces a subsequent rate limiting conformational change to the E(1) state. Similar behavior, but with a decreased saturating value of 1/tau, was found when NaCl was replaced by choline chloride. Analogous experiments performed with enzyme from shark rectal gland showed similar effects, but with a significantly lower amplitude of the fluorescence change and a higher saturating value of 1/tau for both the NaCl and choline chloride titrations. The results suggest that Na(+) ions or salt in general play a regulatory role, similar to that of ATP, in enhancing the rate of the rate-limiting E(2) --> E(1) conformational transition by interaction with the E(2) state. PMID- 12135373 TI - Mutational analysis of the role of tryptophan residues in an antimicrobial peptide. AB - Antimicrobial peptides belonging to the pediocin-like family of bacteriocins (class IIa bacteriocins) produced by lactic acid bacteria contain several tryptophan residues that are highly conserved. Since tryptophan residues in membrane proteins are often positioned in the membrane-water interface, we hypothesized that Trp residues in bacteriocins could be important determinants of the structure of membrane-bound peptides and of anti-microbial activity. To test this hypothesis, the effects of mutating each of the 3 tryptophan residues (Trp18, Trp33, and Trp41) in the 43-residue pediocin-like bacteriocin sakacin P were studied. Trp18 and Trp33 are located at each end of an amphihilic alpha helix, whereas Trp41 is near the end of an unstructured C-terminal tail. Replacement of Trp33 with the hydrophobic residues Leu and Phe had marginal effects on activity, whereas replacement with the more polar Tyr and Arg reduced activity 10-20 and 500-1000 times, respectively, indicating that Trp33 and the C terminal part of the helix interact with the hydrophobic core of the membrane. Any mutation of Trp18 and Trp41 reduced activity, indicating that these two residues play unique roles. Substitutions with other aromatic residues were the least deleterious, indicating that both Trp18 and Trp41 interact with the membrane-water interface. The suggested locations of the three Trp residues are compatible with a structural model in which the helix and the C-terminal tail form a hairpin-like structure, bringing Trp18 and Trp41 close to each other in the interface, and placing Trp33 in the hydrophobic core of the membrane. Indeed, the deleterious effect of the W18L and W41L mutations could be overcome by stabilizing the hairpin-like structure by introduction of a disulfide bridge between residues 24 and 44. These results provide a basis for a refined structural model of pediocin-like bacteriocins and highlight the unique role that tryptophan residues can play in membrane-interacting peptides. PMID- 12135374 TI - Interactions among membrane and soluble components of the flagellar export apparatus of Salmonella. AB - Interactions among several components of the flagellar export apparatus of Salmonella were studied using affinity chromatography, affinity blotting, and fluorescence resonance energy transfer (FRET). The components examined were two integral membrane proteins, FlhA and FlhB, and two soluble components, FliH and the ATPase FliI. Affinity chromatography and affinity blotting demonstrated a heterologous interaction between FlhA and FlhB but not homologous FlhA-FlhA or FlhB-FlhB interactions. Both FlhA and FlhB consist of N-terminal transmembrane domains and C-terminal cytoplasmic domains (FlhA(C) and FlhB(C)). To study the interactions among the cytoplasmic components (FlhA(C), FlhB(C), FliH, and FliI), FRET measurements were carried out using fluorescein-5-isothiocyanate (FITC) as donor and tetramethylrhodamine-5- (and 6-) isothiocyanate (TRITC) as acceptor. To reveal the nature of the binding interactions, measurements were carried out in physiological buffer, at high salt (0.5 M NaCl) and in 30% 2-propanol. The results indicated that FlhA(C) could bind to FlhB(C) and both FlhA(C) and FlhB(C) could bind to themselves. Both FlhA(C) and FlhB(C) bound to FliH and FliI. Several in-frame deletion mutants of FliH were examined and found to have only minor effects of decreased binding to FlhA(C) and FlhB(C); deletions in the interior of the FliH sequence had a greater effect than those at the N terminus. The FliI mutants examined bound FlhA(C) and FlhB(C) about the same as or slightly more weakly than wild-type FliI. FliH bound more weakly to FliI carrying the N terminal double mutation R7C/L12P than it did to wild-type FliI, confirming the importance of the N terminus of FliI for its interaction with FliH. PMID- 12135375 TI - Evidence for a novel, strongly bound acto-S1 complex carrying ADP and phosphate stabilized in the G680V mutant of Dictyostelium myosin II. AB - Gly 680 of Dictyostelium myosin II sits at a critical position within the reactive thiol helices. We have previously shown that G680V mutant subfragment 1 largely remains in strongly actin-bound states in the presence of ATP. We speculated that acto-G680V subfragment 1 complexes accumulate in the A.M.ADP.P(i) state on the basis of the biochemical phenotypes conferred by mutations which suppress the G680V mutation in vivo [Wu, Y., et al. (1999) Genetics 153, 107 116]. Here, we report further characterization of the interaction between actin and G680V subfragment 1. Light scattering data demonstrate that the majority of G680V subfragment 1 is bound to actin in the presence of ATP. These acto-G680V subfragment 1 complexes in the presence of ATP do not efficiently quench the fluorescence of pyrene-actin, unlike those in rigor complexes or in the presence of ADP alone. Kinetic analyses demonstrated that phosphate release, but not ATP hydrolysis or ADP release, is very slow and rate limiting in the acto-G680V subfragment 1 ATPase cycle. Single turnover kinetic analysis demonstrates that, during ATP hydrolysis by the acto-G680V subfragment 1 complex, quenching of pyrene fluorescence significantly lags the increase of light scattering. This is unlike the situation with wild-type subfragment 1, where the two signals have similar rate constants. These data support the hypothesis that the main intermediate during ATP hydrolysis by acto-G680V subfragment 1 is an acto subfragment 1 complex carrying ADP and P(i), which scatters light but does not quench the pyrene fluorescence and so has a different conformation from the rigor complex. PMID- 12135376 TI - Formation of benzo[a]pyrene diol epoxide-DNA adducts at specific guanines within K-ras and p53 gene sequences: stable isotope-labeling mass spectrometry approach. AB - The mutagenicity of a prominent tobacco carcinogen, benzo[a]pyrene (B[a]P), is believed to result from chemical reactions between its diol epoxide metabolite, (+)-anti-7r,8t-dihydroxy-c9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene (BPDE), and DNA, producing promutagenic lesions, e.g., (+)-trans-anti-7R,8S,9S-trihydroxy 10S-(N(2)-deoxyguanosyl)-7,8,9,10-tetrahydrobenzo[a]pyrene (N(2)-BPDE-dG). Previous studies used the DNA repair enzyme UvrABC endonuclease in combination with ligation-mediated PCR (LMPCR) to demonstrate an increased reactivity of BPDE toward guanine nucleobases within codons 157, 248, and 273 of the p53 tumor suppressor gene (Denissenko, M. F., Pao, A., Tang, M., and Pfeifer, G. P. Science 274, 430-432). These sites are also "hot spots" for mutations observed in lung tumors of smokers, suggesting an involvement of B[a]P in the initiation of lung cancer. However, the LMPCR approach relies on the ability of the repair enzyme to excise BPDE-induced lesions, and thus the slowly repaired lesions may escape detection. Furthermore, BPDE-DNA adduct structure and stereochemistry cannot be determined. In the present work, we performed a direct quantitative analysis of N(2)-BPDE-dG originating from specific guanine nucleobases within p53- and K-ras derived DNA sequences by using a stable isotope labeling-mass spectrometry approach recently developed in our laboratory. (15)N-labeled dG was placed at defined positions within DNA sequences derived from the K-ras proto-oncogene and p53 tumor suppressor gene, the two genes most frequently mutated in smoking induced lung cancer. (15)N-labeled DNA was annealed to the complementary strands, followed by BPDE treatment and liquid chromatography-electrospray ionization tandem mass spectrometry analysis (HPLC-ESI-MS/MS) of N(2)-BPDE-dG lesions. The extent of adduct formation at (15)N-labeled guanine was determined directly from the HPLC-ESI-MS/MS peak area ratios of (15)N-N(2)-BPDE-dG and N(2)-BPDE-dG. BPDE induced guanine adducts were produced nonrandomly along K-ras and p53 gene derived DNA sequences, with over 5-fold differences in adduct formation depending on sequence context. N(2)-BPDE-dG yield was enhanced by the presence of 5-Me substituent at the cytosine base-paired with the target guanine nucleobase, an endogenous DNA modification characteristic for CpG dinucleotides within the p53 gene. In the K-ras-derived DNA sequence, the majority of N(2)-BPDE-dG adducts originated from the first position of the codon 12 (GGT), consistent with the large number of G --> T transversions observed at this nucleotide in smoking induced lung cancer. On the contrary, the pattern of N(2)-BPDE-dG formation within the p53 exon 5 sequences did not correlate with the mutational spectrum in lung cancer, suggesting that factors other than N(2)-BPDE-dG formation are responsible for these mutations. The stable isotope labeling HPLC-ESI-MS/MS approach described in this work is universally applicable to studies of modifications to isolated DNA by other carcinogens and alkylating drugs. PMID- 12135377 TI - The affinity of magnesium binding sites in the Bacillus subtilis RNase P x pre tRNA complex is enhanced by the protein subunit. AB - The RNA subunit of bacterial ribonuclease P (RNase P) requires high concentrations of magnesium ions for efficient catalysis of tRNA 5'-maturation in vitro. The protein component of RNase P, required for cleavage of precursor tRNA in vivo, enhances pre-tRNA binding by directly contacting the 5'-leader sequence. Using a combination of transient kinetics and equilibrium binding measurements, we now demonstrate that the protein component of RNase P also facilitates catalysis by specifically increasing the affinities of magnesium ions bound to the RNase P x pre-tRNA(Asp) complex. The protein component does not alter the number or apparent affinity of magnesium ions that are either diffusely associated with the RNase P RNA polyanion or required for binding mature tRNA(Asp). Nor does the protein component alter the pH dependence of pre tRNA(Asp) cleavage catalyzed by RNase P, providing further evidence that the protein component does not directly stabilize the catalytic transition state. However, the protein subunit does increase the affinities of at least four magnesium sites that stabilize pre-tRNA binding and, possibly, catalysis. Furthermore, this stabilizing effect is coupled to the P protein/5'-leader contact in the RNase P holoenzyme x pre-tRNA complex. These results suggest that the protein component enhances the magnesium affinity of the RNase P x pre-tRNA complex indirectly by binding and positioning pre-tRNA. Furthermore, RNase P is inhibited by cobalt hexammine (K(I) = 0.11 +/- 0.01 mM) while magnesium, manganese, cobalt, and zinc compete with cobalt hexammine to activate RNase P. These data are consistent with the hypothesis that catalysis by RNase P requires at least one metal-water ligand or one inner-sphere metal contact. PMID- 12135378 TI - Solution structural studies of the Saccharomyces cerevisiae TATA binding protein (TBP). AB - The intrinsic fluorescence of the six tyrosines located within the C-terminal domain of the Saccharomyces cerevisiae TATA binding protein (TBP) and the single tryptophan located in the N-terminal domain has been used to separately probe the structural changes associated with each domain upon DNA binding or oligomerization of the protein. The unusually short-wavelength maximum of TBP fluorescence is shown to reflect the unusually high quantum yield of the tyrosine residues in TBP and not to result from unusual tryptophan fluorescence. The anisotropy of the C-terminal tyrosines is very high in monomeric, octameric, and DNA-complexed TBP and comparable to that observed in much larger proteins. The tyrosines have low accessibility to an external fluorescence quencher. The anisotropy of the single tryptophan located within the N-terminal domain of TBP is much lower than that of the tyrosines and is accessible to an external fluorescence quencher. Tyrosine, but not tryptophan, fluorescence is quenched upon TBP-DNA complex formation. Only the tryptophan fluorescence is shifted to longer wavelengths in the protein-DNA complex. In addition, the accessibility of the tryptophan residue to the external quencher and the internal motion of the tryptophan residue increase upon DNA binding by TBP. These results show the following: (i) The structure of the C-terminal domain structure is unchanged upon TBP oligomerization, in contrast to the N-terminal domain [Daugherty, M. A., Brenowitz, M., and Fried, M. G. (2000) Biochemistry 39, 4869-4880]. (ii) The environment of the tyrosine residues within the C-terminal domain of TBP is structurally rigid and unaffected by oligomerization or DNA binding. (iii) The C terminal domain of TBP is uniformly in close proximity to bound DNA. (iv) While the N-terminal domain unfolds upon DNA binding by TBP, its increased correlation time shows that the overall structure of the protein is more rigid when complexed to DNA. A model that reconciles these results is proposed. PMID- 12135379 TI - Correlation of binding-loop internal dynamics with stability and function in potato I inhibitor family: relative contributions of Arg(50) and Arg(52) in Cucurbita maxima trypsin inhibitor-V as studied by site-directed mutagenesis and NMR spectroscopy. AB - The side chains of Arg(50) and Arg(52) at positions P(6)' and P(8)', respectively, anchor the binding loop to the protein scaffold by means of hydrogen bonds in Cucurbita maxima trypsin inhibitor-V (CMTI-V), a potato I family member. Here, we have investigated the relative contributions of Arg(50) and Arg(52) to the binding-loop flexibility and stability by determining changes in structure, dynamics, and proteolytic stability as a consequence of individually mutating them into an alanine. We have compared chemical shift assignments of main-chain hydrogens and nitrogens, and (1)H-(1)H interresidue nuclear Overhauser effects (NOEs) for the two mutants with those of the wild-type protein. We have also measured NMR longitudinal and transverse relaxation rates and (15)N-(1)H NOE enhancements for all backbone and side-chain NH groups and calculated the model-free parameters for R50A-rCMTI-V and R52A-rCMTI-V. The three dimensional structures and backbone dynamics of the protein scaffold region remain very similar for both mutants, relative to the wild-type protein. The flexibility of the binding loop is increased in both R50A- and R52A-rCMTI-V. In R52A-rCMTI-V, the mean generalized order parameter () of the P(6)-P(1) residues of the binding loop (39-44) decreases to 0.68 +/- 0.02 from 0.76 +/- 0.04 observed for the wild-type protein. However, in R50A-rCMTI-V, the flexibility of the whole binding loop increases, especially that of the P(1)' P(3)' residues (45-47), whose value drops dramatically to 0.35 +/- 0.03 from 0.68 +/- 0.03 determined for rCMTI-V. More strikingly, S(2) values of side chain N epsilon Hs reveal that, in the R50A mutant, removal of the R50 hydrogen bond results in the loss of the R52 hydrogen bond too, whereas in R52A, the R50 hydrogen bond remains unaffected. Kinetic data on trypsin-catalyzed hydrolysis of the reactive-site peptide bond (P(1)-P(1)') suggest that the activation free energy barrier of the reaction at 25 degrees C is reduced by 2.1 kcal/mol for R50A-rCMTI-V and by 1.5 kcal/mol for R52A-rCMTI-V, relative to rCMTI-V. Collectively, the results suggest that although both the P(6') and P(8)' anchors are required for optimal inhibitor function and stability in the potato I family, the former is essential for the existence of the latter and has greater influence on the binding-loop structure, dynamics, and stability. PMID- 12135380 TI - Decay products of the S(3) state of the oxygen-evolving complex of photosystem II at cryogenic temperatures. Pathways to the formation of the S = 7/2 S(2) state configuration. AB - The water-oxidizing complex of photosystem II cycles through five oxidation states, denoted S(i)() (i = 0-4), during water oxidation to molecular oxygen, which appears at the (transient) S(4) state. The recent detection of bimodal EPR signals from the S(3) state [Matsukawa, T., Mino, H., Yoneda, D., Kawamori, A. (1999) Biochemistry 38, 4072-4077] has drawn significant attention to this critical state. An interesting property of the S(3) state is the sensitivity to near-IR (NIR) light excitation. Excitation of the S(3) state by near-IR light at cryogenic temperatures induces among other signals a derivative-shaped EPR signal at g= 5 [Ioannidis, N., and Petrouleas, V. (2000) Biochemistry 39, 5246-5254]. The signal bears unexpected similarities to a signal observed earlier in samples that had undergone multiple turnovers and subsequently had been stored at 77 K for a week or longer [Nugent, J. H. A., Turconi, S., and Evans, M. C. W. (1997) Biochemistry 36, 7086-7096]. Recently, both signals were assigned to an S = 7/2 configuration of the Mn cluster [Sanakis, Y., Ioannidis, N., Sioros, G., and Petrouleas, V. (2001) J. Am. Chem. Soc. 123, 10766-10767]. In the present study, we employ bimodal EPR spectroscopy to investigate the pathways of formation of this unusual state. The following observations are made: (i) The g = 5 signal evolves in apparent correlation with the diminution of the S(3) state signals during the slow (tens of hours to several days range) charge recombination of S(3) with Q(A)(-) at 77 K. The tyrosyl radical D* competes with S(3) for recombination with Q(A)(-), the functional redox couple at cryogenic temperatures inferred to be D*/D(-). Transfer to -50 degrees C and above results in the relaxation of the g = 5 to the multiline and g = 4.1 signals of the normal S(2) state. (ii) The transition of S(3) to the state responsible for the g = 5 signal can be reversed by visible light illumination directly at -30 degrees C or by illumination at 4.2 K followed by brief (2 min) transfer to -50 degrees C in the dark. The latter step is required in order to overcome an apparent thermal activation barrier (charge recombination appears to be faster than forward electron transfer at 4.2 K). (iii) The "g = 5" state can be reached in a few tens of minutes at 4.2 K by near-IR light excitation of the S(3) state. This effect is attributed to the transfer of the positive hole from the Mn cluster to a radical (probably tyr Z), which recombines much faster than the Mn cluster with Q(A)(-). (iv) The above properties strongly support the assignment of the configuration responsible for the g = 5 signal to a modified S(2) state, denoted S(2)'. Evidence supporting the assignment of the S(2)' to a proton-deficient S(2) configuration is provided by the observation that the spectrum of S(2) at pH 8.1 (obtained by illumination of the S(1) state at -30 degrees C) contains a g = 5 contribution. PMID- 12135381 TI - Intermediates of the S(3) state of the oxygen-evolving complex of photosystem II. AB - The S(3) state of the water-oxidizing complex (WOC) of photosystem II (PSII) is the last state that can be trapped before oxygen evolution occurs at the transient S(4) state. A number of EPR-detectable intermediates are associated with this critical state. The preceding paper examined mainly the decay of S(3) at cryogenic temperatures leading to the formation of a proton-deficient configuration of S(2) termed S(2)'. This second paper examines all intermediates formed by the near-IR light (NIR) excitation of the S(3) state and compares these with the light-excitation products of the S(2)' state. The rather complex set of observations is organized in a comprehensive flowchart, the central part of which is the S(3)...Q(A)(-) state. This state can be converted to various intermediates via two main pathways: (A) Excitation of S(3) by NIR light at temperatures below 77 K results presumably in the formation of an excited S(3) state, S(3), which decays via either of two pathways. Slowly at liquid helium temperatures but much faster at 77 K, S(3) decays to an EPR-silent state, denoted S(3)' ', which by raising the temperature to ca. 190 K converts to a spin configuration of the Mn cluster, characterized by g = 21, 3.7 in perpendicular and g = 23 in parallel mode EPR, denoted S(3)'. Upon further warming to 220 K, S(3)' relaxes to the untreated S(3) state. Below about 77 K and more favorably at liquid helium temperatures, an alternative pathway of S(3) decay via the metallo-radical intermediate S(2)'Z*...Q(A)(-) can be traced. This leads to the metastable state S(2)'Z...Q(A) via charge recombination. S(2)'Z* is characterized by a split radical signal at g = 2, while all S(2)' transients are characterized by the same g = 5/2.9 (S = (7)/(2)) configuration of the Mn cluster with small modifications, reflecting an influence of the tyr Z oxidation state on the crystal-field symmetry at the Mn cluster. (B) S(2)'...Q(A) can be reached alternatively by the slow charge recombination of S(3) and Q(A)(-) at 77 K. White-light illumination of S(2)'.Q(A) below about 20 K results in charge separation, reforming the intermediate S(2)'Z*...Q(A)(-). Thermally activated branches to the main pathways are also described, e.g., at elevated temperatures tyr Z* reoxidizes S(2)' to the S(3) state. The above observations are discussed in terms of a molecular model of the S(3) state of the OEC. Main aspects of the model are the following. Intermediates, isoelectronic to S(3), are attributed to the NIR-induced translocation of the positive hole to different Mn ligands, or to tyr Z. On the basis of a comparison of the electron-donating efficiency of tyr Z and tyr D at cryogenic temperatures, it is inferred that the Mn cluster acts as the main proton acceptor from tyr Z. Water associated with the Mn cluster is assumed to be in hydrogen-bonding equilibrium with tyr Z, and an array comprising this water and adjacent water (or OH or O) ligands to Mn followed by a sequence of proton acceptors is proposed to act as an efficient proton translocation pathway. Oxidation of the tyrosine by P(680)(+) repels protons to and out from the Mn cluster. This proposed role of tyr Z in the water-splitting process is described as a proton repeller/electron abstractor. PMID- 12135382 TI - Regioselective peroxo-dependent heme alkylation in P450(BM3)-F87G by aromatic aldehydes: effects of alkylation on cataysis. AB - Cytochrome P450(BM3)-F87G reacts with aromatic aldehydes and hydrogen peroxide to generate covalent heme adducts in a reaction that may involve the formation of a stable isoporphyrin intermediate [Raner, G. M., Hatchell, A. J., Morton, P. E., Ballou, D. P., and Coon, M. J. (2000) J. Inorg. Biochem. 81, 153-160]. Electron paramagnetic resonance spectra for the proposed isoporphyrin intermediates generated using two different aromatic aldehydes suggest that, in each case, the heme remained coordinated to the apoenzyme via the cysteine thiolate, the metal center remained ferric low spin, and a slight distortion in the geometry of the pyrrole nitrogens occurred. Characterization of the resulting heme adducts via 1D and 2D NMR showed conclusively that the heme was modified at the gamma-meso position alone, and mass spectral analysis indicated loss of formate from the aldehyde prior to alkylation. The enzyme derivatives in which the hemes were covalently altered retained the characteristic UV/vis and EPR spectral properties of a P450, indicating that the heme was properly ligated in the active site. The modified enzymes were able to accept electrons from NADPH in the presence of lauric acid at a rate comparable to that of the unmodified forms, although oxidation of the lauric acid was not observed with either modified enzyme. Oxidation of 4-nitrophenol and 4-nitrocatechol was observed for both derivatives. However, 4-nitrocatechol oxidation was completely quenched in the presence of superoxide dismutase. The results are consistent with heme modification occurring through a peroxo-dependent pathway and also suggest that modification results in altered catalytic activity, rather than complete inactivation of the P450. PMID- 12135383 TI - Benzoate 1,2-dioxygenase from Pseudomonas putida: single turnover kinetics and regulation of a two-component Rieske dioxygenase. AB - The benzoate 1,2-dioxygenase system (BZDOS) from Pseudomonas putida mt-2 catalyzes the NADH-dependent oxidation of benzoate to 1-carboxy-1,2-cis dihydroxycyclohexa-3,5-diene. Both the oxygenase (BZDO) and reductase (BZDR) components of BZDOS have been purified and characterized kinetically and by optical, EPR, and Mossbauer spectroscopies. BZDO has an (alpha beta)(3) subunit structure in which each alpha subunit contains a Rieske [2Fe-2S] cluster and a mononuclear iron site. Two different purification protocols were developed for BZDO allowing the mononuclear iron to be stabilized in either the Fe(III) or the Fe(II) state for spectroscopic characterization. Using single turnover reactions, it is shown that fully reduced BZDO alone is capable of yielding the cis-diol product in high yield at rates that exceed the BZDOS turnover number. At the conclusion of turnover, quantification of each oxidation state of the metal sites by EPR and Mossbauer spectroscopies shows that the Rieske cluster and mononuclear iron are each oxidized in amounts equal to the product yield, suggesting that the two electrons required for catalysis derive from the two metal centers. These results are in agreement with our previous study of naphthalene 1,2-dioxygenase [Wolfe, M. D., Parales, J. V., Gibson, D. T., and Lipscomb, J. D. (2001) J. Biol. Chem. 276, 1945-1953], which belongs to a different Rieske dioxygenase subclass, suggesting that it is a universal characteristic of Rieske dioxygenases that oxygen activation and substrate oxidation are catalyzed by the oxygenase component alone. The EPR spectrum of the Fe(III) center after a single turnover is distinct from either of those of substrate-free or substrate-bound enzyme. The complex with this spectrum is not formed by addition of cis-diol product to the resting Fe(III) form of the enzyme but is observed when the Fe(II) form is oxidized in the presence of product. Together, these results suggest that product exchange occurs only when the mononuclear iron is reduced. Stopped-flow and rapid scan analyses monitoring the oxidation of the Rieske cluster during the single turnover reaction show that it occurs in three phases that are kinetically competent for catalysis. The rate of each phase was found to be dependent on the type of substrate present, suggesting that the substrate influences the rate of electron transfer between the metal clusters. The participation of substrate in the oxygen activation reaction suggests a new aspect of the mechanism of this process by the Rieske dioxygenase class. PMID- 12135384 TI - NMR structure of lung surfactant peptide SP-B(11-25). AB - Surfactant protein B (SP-B) is a 79-residue essential component of lung surfactant, the film of lipid and protein lining the alveoli, and is the subject of great interest for its role in lung surfactant replacement therapies. Here we report circular dichroism results and the solution NMR structure of SP-B(11-25) (CRALIKRIQAMIPKG) dissolved in CD(3)OH at 5 degrees C. This is the first report of NMR data related to the protein SP-B, whose structure promises to help elucidate the mechanism of its function. Sequence-specific resonance assignments were made for all observable (1)H NMR signals on the basis of standard 2D NMR methods. Structures were determined by the simulated annealing method using restraints derived from 2D NOESY data. The calculations yielded 17 energy minimized structures, three of which were subjected to 0.95 ns of restrained dynamics to assess the relevance of the static structures to more realistic dynamic behavior. Our CD and NMR data confirm that this segment is an amphiphilic alpha helix from approximately residue L14 through M21. The backbone heavy-atom RMSD for residues L14 through M21 is 0.09 +/- 0.12 A, and the backbone heavy-atom RMSD for the whole peptide is 0.96 +/- 2.45 A, the difference reflecting fraying at the termini. Aside from the disordered termini, the minimized structures represent dynamic structures well. Structural similarity to the homologous regions of related saposin-like proteins and the importance of the distribution of polar residues about the helix axis are discussed. PMID- 12135385 TI - Allosterism and cooperativity in Pseudomonas aeruginosa GDP-mannose dehydrogenase. AB - GDP-Mannose dehydrogenase catalyzes the formation of GDP-mannuronic acid, which is the monomeric unit from which the polysaccharide alginate is formed. Alginate is secreted by the pathogenic bacterium Pseudomonas aeruginosa and is believed to play an important role in the bacteria's resistance to antibiotics and the host immune response. We have characterized the kinetic behavior of GDP-mannose dehydrogenase in detail. The enzyme displays cooperative behavior with respect to NAD(+) binding, and phosphate and GMP act as allosteric effectors. Binding of the allosteric effectors causes the Hill coefficient for NAD(+) binding to decrease from 6 to 1, decreases K(1/2) for NAD(+) by a factor of 10, and decreases V(max) by a factor of 2. The cooperative binding of NAD(+) is also sensitive to pH; deprotonation of two residues with identical pK's of 8.0 is required for maximally cooperative behavior. The kinetic behavior of GDP-mannose dehydrogenase suggests that it must be at least hexameric under turnover conditions; however, dynamic light-scattering measurements do not provide a clear determination of the size of the active enzyme complex. PMID- 12135386 TI - Role of C-terminal sequences in the folding of muscle creatine kinase. AB - Proteinase K selectively nicks the native homodimeric muscle creatine kinase (MM CK) into two 37.1 kDa N-terminal (K1) and two 5.8 kDa C-terminal (K2) fragments that remain firmly associated in a native-like, although inactive, heterotetrameric structure. This truncated protein has been named (K1K2)(2). To analyze the role of the C-terminal peptide in the protein structure acquisition, we studied in vitro refolding of the guanidinium chloride-denatured (K1K2)(2). Although they never reassociate with K2, in selected conditions the K1 fragments refold slowly to a dimeric state as shown by size exclusion chromatography data. This K1 dimer exhibits a fluorescence emission lambda max of 335 nm, a high degree of tyrosine exposure, strongly binds ANS but not MgADP, a CK substrate, and according to these structural characteristics, could be a dimeric molten globule species. We propose a folding model that takes into account the existence of a new transient intermediate state in the MM-CK refolding process. Besides two monomeric premolten and molten globule kinetic intermediates and the active final dimeric form, an inactive dimer, with partly compacted monomers, must ephemerally exist. Our results strongly suggest that the C-terminal end of the protein accelerates folding and plays a critical role for monomer final packing into a native-like conformation. The data also indicate that MM-CK catalytic efficiency is only acquired after dimerization. PMID- 12135387 TI - Isoform-selective interaction of cyclooxygenase-2 with indomethacin amides studied by real-time fluorescence, inhibition kinetics, and site-directed mutagenesis. AB - Conversion of carboxylate-containing nonsteroidal antiinflammatory drugs, such as indomethacin, to esters or amides provides potent and selective inhibitors of cyclooxygenase-2 (COX-2) [Kalgutkar et al. (2000) Proc. Natl. Acad. Sci. U.S.A. 97, 925-930]. Synthesis of cinnamyl- or coumarinyl-substituted ethanolamide derivatives of indomethacin produced fluorescent probes of inhibitor interaction with COX-2 and COX-1. Binding of either derivative to apoCOX-2 or apoCOX-1 resulted in a rapid, reversible enhancement of fluorescence. Following this rapid phase, a slow additional increase in fluorescence was observed with apoCOX-2 but not with apoCOX-1. The slow, COX-2-specific increase in fluorescence was prevented or reversed by addition of the nonfluorescent COX inhibitor (S) flurbiprofen. Detailed kinetic studies of the interaction of the coumarinyl derivative with COX-2 showed that the observed changes in fluorescence could be described by two sequential equilibria, the first of which is rapid, reversible, and bimolecular in the forward direction. The second equilibrium is slower, reversible, and unimolecular in both directions. The forward rate constant for the slow equilibrium determined by fluorescence enhancement [(3.1 +/- 0.6) x 10( 3) s(-1)] corresponded closely to the forward rate constant for inhibition of COX 2 activity [(6.8 +/- 2.3) x 10(-3) s(-1)], suggesting that the slow fluorescence enhancement is associated with selective COX-2 inhibition. Site-directed mutagenesis indicated that residues in the carboxylate-binding region of the COX 2 active site (Arg-120, Tyr-355, and Glu-524) are critical for the binding of the indomethacin conjugates that leads to slow fluorescence enhancement and cyclooxygenase inhibition. The indomethacin conjugates described herein represent powerful tools for the investigation of a novel class of selective inhibitors of COX-2. PMID- 12135388 TI - A link between Cdc42 and syntaxin is involved in mastoparan-stimulated insulin release. AB - Mastoparan, a hormone receptor-mimetic peptide isolated from wasp venom, stimulates insulin release from pancreatic beta-cells in a Ca(2+)-independent but GTP-dependent manner. In this report, the role of the Rho family GTP-binding protein Cdc42, in the mastoparan stimulus-secretion pathway, was examined. Overexpression of wild-type Cdc42 in beta HC-9 cells, an insulin-secreting mouse derived cell line, resulted in a 2-fold increase in mastoparan-stimulated insulin release over vector-transfected beta HC-9 cells. This effect was not seen with secretagogues such as glucose that stimulate secretion via Ca(2+)-dependent pathways. GDP/GTP exchange assay data and studies with pertussis (PTX) toxin suggest that mastoparan may work directly to activate Cdc42 and not via PTX sensitive heterotrimeric GTP-binding proteins. Using bacterial glutathione S transferase-Cdc42 fusion proteins and co-immunoprecipitation and transient transfection studies, Cdc42 was shown to be an upstream regulator of the exocytotic protein, syntaxin. These results suggest that the GTP-dependent signal underlying the mastoparan effect acts at a "distal site" in stimulus-secretion coupling on one of the SNARE proteins essential for exocytosis. In vitro binding assays, using purified Cdc42 and syntaxin proteins, show that Cdc42 mediates the GTP signal through an indirect association with syntaxin. The H3 domain at the C terminus of syntaxin, which participates in the formation of the ternary SNARE complex with the core proteins, SNAP-25 and synaptobrevin, is also required for the association with Cdc42. Thus, these studies indicate that Cdc42 could be a putative GTP-binding protein thought to be involved in the mastoparan-stimulated GTP-dependent pathway of insulin release. PMID- 12135389 TI - Allosteric activation of cGMP-specific, cGMP-binding phosphodiesterase (PDE5) by cGMP. AB - The effects of cGMP binding on the catalytic activity of cGMP-specific, cGMP binding phosphodiesterase (PDE5) are unclear because cGMP interacts with both allosteric and catalytic sites specifically. We studied the effects of cGMP on the hydrolysis of a fluorescent substrate analogue, 2'-O-anthraniloyl cGMP, by PDE5 partially purified from rat cerebella. The preparation contained PDE5 as the major cGMP-PDE activity and was not contaminated with cAMP- or cGMP-dependent protein kinases. The Hill coefficients for hydrolysis of the analogue substrate were around 1.0 in the presence of cGMP at concentrations <0.3 microM, while they increased to 1.5 at cGMP concentrations >1 microM, suggesting allosteric activation by cGMP at concentrations close to the bulk binding constant of the enzyme. Consistent with an allosteric activation, increasing concentrations of cGMP enhanced the hydrolysis rate of fixed concentrations of 2'-O-anthraniloyl cGMP, which overcame competition between the two substrates. Such activation was not observed with cAMP, cyclic inosine 3',5'-monophosphate, or 2'-O-monobutyl cGMP, indicating specificity of cGMP. These results demonstrate that cGMP is a specific and allosteric activator of PDE5, and suggest that in cells containing PDE5, such as cerebellar Purkinje cells, intracellular cGMP concentrations may be regulated autonomously through effects of cGMP on PDE5. PMID- 12135390 TI - ADP-ribosylation and functional effects of Pseudomonas exoenzyme S on cellular RalA. AB - Exoenzyme S (ExoS) is a bifunctional virulence factor directly translocated into eukaryotic cells by the type III secretory process of Pseudomonas aeruginosa. Bacterial translocation of ExoS into epithelial cells is associated with diverse effects on cell function, including inhibition of growth, alterations in cell morphology, and effects on adherence processes. Preferred substrates of the ADP ribosyltransferase (ADPRT) portion of ExoS include low molecular weight G proteins (LMWG-proteins) in the Ras family. In examining the ADP-ribosylation and functional effects of ExoS on RalA, ExoS was found to ADP-ribosylate endogenous RalA and recombinant RalADeltaCAAX at multiple sites, with Arg52 identified as the preferred site of ADP-ribosylation. The binding of RalA to the Ral binding domain (RBD) of its downstream effector, RalBP1, was inhibited by bacterially translocated ExoS, indicating an effect of ExoS on cellular RalA function. In vitro analyses confirmed that ADP-ribosylation of RalA directly interfered with its ability to bind to the RBD of RalBP1. The studies support the fact that RalA is a cellular substrate of bacterially translocated ExoS and that ADP ribosylation by ExoS affects RalA interaction with its downstream effector, RalBP1. PMID- 12135391 TI - Differential lipid binding of truncation mutants of Galleria mellonella apolipophorin III. AB - Apolipophorin III (apoLp-III) is a prototype exchangeable apolipoprotein that is amenable to structure-function studies. The protein folds as a bundle of five amphipathic alpha-helices and undergoes a dramatic conformational change upon lipid binding. Recently, we have shown that a truncation mutant of Galleria mellonella apoLp-III comprising helices 1-3 is stable in solution and able to bind to lipid surfaces [Dettloff, M., Weers, P. M. M., Niere, M., Kay, C. M., Ryan, R. O., and Wiesner, A. (2001) Biochemistry 40, 3150-3157]. To investigate the role of the C-terminal helices in apoLp-III structure and function, two additional 3-helix mutants were designed: a core fragment comprising helix (H) 2 4, and a C-terminal fragment (H3-5). Each truncation mutant retained the ability to associate spontaneously with dimyristoylphosphatidylcholine (DMPC) vesicles, transforming them into discoidal complexes. The rate of apolipoprotein-dependent DMPC vesicle transformation decreased in the order H1-3 > H2-4 > H3-5. Truncation of two helices led to a significant decrease in alpha-helical content in buffer in each case, from 86% (wild-type) to 50% (H1-3), 28% (H2-4), and 24% alpha helical content (H3-5). On the other hand, trifluoroethanol or complexation with DMPC induced the truncation mutants to adopt a high alpha-helical structure similar to that of wild-type protein (84-100% alpha-helical structure). ApoLp III(H1-3) and apoLp-III(H2-4), but not apoLp-III(H3-5), were able to prevent phospholipase-C-induced low density lipoprotein aggregation, indicating that interaction of the C-terminal fragment with spherical lipoprotein surfaces was impaired. As lipoprotein binding is significantly affected and DMPC transformation rates are relatively slow upon removal of N-terminal helices, the data indicate that structural elements necessary for lipid interaction reside in the N-terminal part of the protein. PMID- 12135392 TI - Combinatorial selection and binding of phosphorothioate aptamers targeting human NF-kappa B RelA(p65) and p50. AB - Previously, we reported the in vitro combinatorial selection of phosphorothioate aptamers or "thioaptamers" targeting the transcription factor NF-IL6. Using the same approach and purified recombinant human NF-kappa B proteins RelA(p65) and p50, duplex thioaptamers have been selected that demonstrate high-affinity, competitive binding with the duplex 22-mer binding site, Ig kappa B. Binding energetics of RelA(p65) and p50 homodimers were studied using a quantitative electrophoretic mobility shift assay or EMSA. As a reference system for competitive aptamer binding, the duplex 22-mer phosphoryl binding site known as Ig kappa was determined to bind each p65 and p50 homodimer with a 1:1 stoichiometry and with affinities, determined by global analysis, K(d) = 4.8 +/- 0.2 nM for p65 and K(d) = 0.8 +/- 0.2 nM for p50. A global analysis tool for competitive NF-kappa B/Ig kappa binding was developed and utilized to measure the affinity of thioaptamers selected by both p65 and p50. The competition results indicate that the thioaptamers bind and compete for the same NF-kappa B site as the known promoter element Ig kappa B (K(d) = 78.9 +/- 1.9 nM for a p65-selected aptamer and 19.6 +/- 1.3 nM for a p50-selected thioaptamer). Qualitative gel shift binding experiments with p50 also demonstrate that the nature of enhanced affinity and specificity can be attributed to the presence of sulfur. Collectively, these results demonstrate the feasibility of the thioaptamer in vitro combinatorial selection technology as a method for producing specific, high affinity ligands to proteins. PMID- 12135393 TI - Analysis of protein-carbohydrate interaction at the lower size limit of the protein part (15-mer peptide) by NMR spectroscopy, electrospray ionization mass spectrometry, and molecular modeling. AB - Structural analysis of minimally sized lectins will offer insights into fundamentals of intermolecular recognition and potential for biomedical applications. We thus moved significantly beyond the natural limit of lectin size to determine the structure of synthetic mini-lectins in solution, their carbohydrate selectivity and the impact of ligand binding on their conformational behavior. Using three disaccharide (Thomsen-Friedenreich antigen; Gal beta 1,3GalNAc alpha 1,R)-binding pentadecapeptides without internal disulfide bridges as role models, we successfully tested a combined strategy with different techniques of NMR spectroscopy, electrospray ionization mass spectrometry, and molecular modeling. In solution, the peptides invariably displayed flexibility with rather limited restrictions, shown by NMR experiments including nearly complete resonance assignments and molecular dynamics simulations. The occurrence of aromatic/nonpolar amino acids in the sequence did not lead to formation of a hydrophobic core known from microbial chitinase modules. Selectivity of disaccharide binding was independently observed by mass spectrometry and NMR analysis. Specific ligand interaction yielded characteristic NMR signal alterations but failed to reduce conformational flexibility significantly. We have thereby proven effectiveness of our approach to analyze even low-affinity interactions (not restricted to carbohydrates as ligands). It will be useful to evaluate the impact of rational manipulation of lead peptide sequences. PMID- 12135394 TI - Exploitation of the selectivity-conferring code of nonribosomal peptide synthetases for the rational design of novel peptide antibiotics. AB - Recently, the solved crystal structure of a phenylalanine-activating adenylation (A) domain enlightened the structural basis for the specific recognition of the cognate substrate amino acid in nonribosomal peptide synthetases (NRPSs). By adding sequence comparisons and homology modeling, we successfully used this information to decipher the selectivity-conferring code of NRPSs. Each codon combines the 10 amino residues of a NRPS A domain that are presumed to build up the substrate-binding pocket. In this study, the deciphered code was exploited for the first time to rationally alter the substrate specificity of whole NRPS modules in vitro and in vivo. First, the single-residue Lys239 of the L-Glu activating initiation module C-A(Glu)-PCP of the surfactin synthetase A was mutated to Gln239 to achieve a perfect match to the postulated L-Gln-activating binding pocket. Biochemical characterization of the mutant protein C-A(Glu) PCP(Lys239 --> Gln) revealed the postulated alteration in substrate specificity from L-Glu to L-Gln without decrease in catalytic efficiency. Second, according to the selectivity-conferring code, the binding pockets of L-Asp and L-Asn activating A domains differs in three positions: Val299 versus Ile, His322 versus Glu, and Ile330 versus Val, respectively. Thus, the binding pocket of the recombinant A domain AspA, derived from the second module of the surfactin synthetases B, was stepwisely adapted for the recognition of L-Asn. Biochemical characterization of single, double, and triple mutants revealed that His322 represents a key position, whose mutation was sufficient to give rise to the intended selectivity-switch. Subsequently, the gene fragment encoding the single mutant AspA(His322 --> Glu) was introduced back into the surfactin biosynthetic gene cluster. The resulting Bacillus subtilis strain was found to produce the expected so far unknown lipoheptapeptide [Asn(5)]surfactin. This indicates that site-directed mutagenesis, guided by the selectivity-conferring code of NRPS A domains, represents a powerful alternative for the genetic manipulation of NRPS biosynthetic templates and the rational design of novel peptide antibiotics. PMID- 12135397 TI - Racial and ethnic disparities in access to care for children with special health care needs. AB - OBJECTIVE: Numerous studies have examined racial and ethnic differences in access to and utilization of health services. However, few studies have addressed these issues with respect to children with special health care needs. This study examines whether disparities in access and utilization are present among black, white, and Hispanic children identified as having special health care needs. METHODS: We analyzed data on 57 553 children younger than 18 years old included in the 1994-95 National Health Interview Survey on Disability. Of these, 10 169, or 17.7% of the sample, were identified as having an existing special health care need. Bivariate and multivariate analyses were used to assess how race and ethnicity are related to measures of access and utilization, such as usual source of care, missed care, and use of physician and hospital services. RESULTS: Our analyses show that among children with special health care needs, minorities were more likely than white children to be without health insurance coverage (13.2% vs 10.3%; P <.01), to be without usual source of care (6.7% vs 4.3%; P <.01), and to report inability to get needed medical care (3.9% vs 2.8%; P <.05). Also, white children with special health care needs were more likely than their minority counterparts to have used physician services (88.6 vs 85.0; P <.01); however, minority children with special health care needs were more likely to have been hospitalized during the past year (7.6% vs 6.3%; P < 0.5). After adjustments for confounding variables (income, insurance coverage, health status, and other variables), racial and ethnic differences in access and utilization were attenuated but remained significant for several measures (without a usual source of care, receipt of care outside of a doctor's office or HMO, no regular clinician, no doctor contacts in past year, and volume of doctor contacts). Gaps in access were more frequent and generally larger for Hispanic children with special health care needs. CONCLUSIONS: Our analysis indicates that access and utilization disparities remain between white and minority children with special health care needs, with Hispanic children experiencing especially disparate care. PMID- 12135396 TI - Social class gradients and health in childhood. AB - OBJECTIVE: To determine if there are social class gradients in health in children aged 6 to 11 years. METHODS: Self and parent reports of health of children in 5 sites across the United States were assessed using the Child Health and Illness Profile-Child Edition. Distribution of scores in 4 domains: satisfaction (with health); comfort; resilience; and risk avoidance were used to create profiles of health. Social class was defined as a composite of parental education and work participation. RESULTS: Social class gradients were found for all but the satisfaction domain and for most subdomains in the parent version; the most notable gradient was in the risk avoidance domain, with better health the higher the social class. Apparent gradients did not reach statistical significance in the child reports. Children from a higher social class were more likely to be in excellent/average health and less likely to be in poor health profiles than were lower class children. CONCLUSIONS: The findings generally mirror those from a prior study of adolescents, using the same conceptual framework for health and the same measure of social class, and are consistent with a cumulative effect for most aspects of health, and with a critical-period effect for risky behaviors. PMID- 12135398 TI - Delivering oral health services to children in the past: the rise and fall of children's dental clinics. AB - As a contribution to the current discourse on improving the oral health of impoverished children by increasing their access to oral health services, this essay describes and examines an earlier attempt to accomplish a very similar goal: the philanthropically and publicly funded children's dental clinics that were responsible for close to half of all oral health services delivered to US children during the first half of the 20th century. As an explanation of why these clinics were established and why they proliferated, the essay argues they met 4 criteria essential to successful public health programs aimed at children: they had a clearly understood and largely accepted fiscal and social utility; they provided services that parents could easily understand as benefiting their children; they serviced a broad enough segment of the population to earn them significant social and political support; and, by meeting a variety of the professional needs of oral health care providers, they established a relationship of enlightened self interest with a group whose support or, at least, lack of opposition was crucial to their survival. PMID- 12135399 TI - Is there a common cold constitution? AB - OBJECTIVE: Constitutional factors might play a role in the susceptibility to clinical illness during the common cold. This study seeks to determine if the likelihood of developing frequent common colds persists during childhood. DESIGN: The Tucson Children's Respiratory Study involves 1246 children enrolled at birth and followed prospectively since 1980 and 1984. Parents reported the occurrence of frequent (> or =4) colds during the past year by questionnaire at 2, 3, 6, 8, 11, and 13 years of age. Blood for ex vivo interferon-gamma responses was obtained at 9 months and 11 years of age. RESULTS: After adjustment for potential confounding variables, children with frequent colds at year 2 or 3 were twice as likely to experience frequent colds at year 6 (relative risk [RR], 2.8; 95% confidence interval [CI], 2.1-3.9), year 8 (RR, 2.6; 95% CI, 2.1-3.3), year 11 (RR, 2.4; 95% CI, 1.8-3.1), and year 13 (RR, 2.1; 95% CI, 1.4-3.3) compared with children who had infrequent colds at years 2 and 3. At 9 months of age, children who ultimately experienced persistent frequent colds had lower interferon-gamma titers than children without persistent frequent colds (3.05 +/- 1.61 vs 3.74 +/- 1.39, P =.016); this finding persisted at 11 years of age. CONCLUSION: These data suggest the existence of a common cold constitution, whereby some children are more susceptible to infection and/or the expression of clinical symptoms when infected than are other children. PMID- 12135400 TI - Risk factors for emergency department use among children with asthma using primary care in a managed care environment. AB - OBJECTIVE: To identify risk factors for emergency department (ED) use among children with asthma using primary care in a managed care environment. DESIGN: Using automated data sources, children with asthma were identified and followed for 2-year periods. We fit logistic regression models using generalized estimating equation approaches to identify ED risk factors. PATIENTS: Children with asthma aged 5-14 with a visit to a pediatrician practicing with a large group practice and enrolled in an HMO for 2 consecutive years between 1992 and 1996 (N = 411 children). MAIN OUTCOME MEASURES: Asthma-related ED use. RESULTS: Twenty-three percent of children incurred an asthma-related ED visit. Asthma related ED use was greater among children with prior asthma-related ED use (OR [odds ratio] = 8.26, 95% CI [confidence interval] = 4.79-14.25), decreased with increasing age (OR = 0.87, 95% CI = 0.79-0.96) and frequency of visits to a primary care physician for asthma (OR = 0.82, 95% CI = 0.70-0.96), and tended to be less among children who saw an allergist (OR = 0.59, 95% CI = 0.33-1.04). No significant relationship was found between asthma-related ED use and race, household income, or other patient characteristics. CONCLUSIONS: Targeting children with prior asthma-related ED use and encouraging routine primary care visits as well as the use of an allergist may afford opportunities to reduce ED use among children with asthma currently receiving primary care. PMID- 12135401 TI - Incidence and description of high chair-related injuries to children. AB - STUDY OBJECTIVE: To describe the incidence, circumstances, and types of high chair-related injuries among US children. DESIGN: Retrospective review of data for children 3 years old and younger from the National Electronic Injury Surveillance System of the US Consumer Product Safety Commission for 1994-98. RESULTS: There were an estimated 40 650 high chair-related injuries (95% confidence interval [CI], 32 657-48 643) to children 3 years old and younger treated in hospital emergency departments in the US during the 5-year study period. An estimated 5231 injuries (13%) were related to use of an attachable high chair (including booster seats), and an estimated 4067 (10%) were related to the use of a youth chair. The annual rate of injury among children < or =3 years old was 5.3 per 10 000. The mean age was 10 months (median, 1 year); 56% were boys. Ninety-four percent of injuries resulted from a fall from the chair. Most injuries involved the head (44%) or face (39%). Injury diagnoses included contusions or abrasions (36%), lacerations (25%), closed head injury (21%), and fractures (8%). Two percent of injured children, an estimated 941 (95% CI, 399 1487), were admitted to the hospital during the study period, an annual admission rate of 0.1 per 10 000. There were no significant differences between attachable high chairs, youth chairs, and high chairs in anatomic sites of injury, injury diagnosis, or frequency of hospital admission. CONCLUSIONS: Injuries related to high chairs are common, particularly among children in the first year of life. They often result from falls from the chair. The data suggest that restraint use would prevent most of these injuries. PMID- 12135402 TI - The burden of injury in preschool children in an urban medicaid managed care organization. AB - BACKGROUND: Efforts to control injuries within managed care organization (MCO) populations require information about the incidence and costs associated with the injuries cared for in MCOs. OBJECTIVE: This study uses administrative data to measure the rates and the costs of burn, choking, poisoning, blunt, and penetrating injuries in an urban Medicaid MCO. DESIGN/METHODS: A database was assembled from all medical claims submitted to a Medicaid MCO covering children aged < or =6 years in urban Baltimore between the dates of July 1, 1997, and August 7, 1999. The database included claims submitted on behalf of 1732 children observed for 2180 person-years. International Classification of Disease-9 codes were reviewed to identify claims for burn, poisoning, choking, and blunt/penetrating injuries. Trained coders reviewed outpatient records to assign E-codes. RESULTS: A total of 796 injuries occurred. The overall injury rate was 36.5% per year. The total cost of the medical care for these injuries was $863 552, or $396 per covered person-year, representing 42%-55% of the capitated rate received in Baltimore. Falls, being struck by something, and cutting/piercing injuries accounted for 68% of injuries. Emergency departments were the most common service sites for injured children for all injuries except in the case of burns. CONCLUSION: The children enrolled in this urban Medicaid population had nearly twice the rate of injury when compared to the national average. The medical costs of injuries account for about half of the capitated reimbursement for this age group. PMID- 12135403 TI - The importance of outcomes research in pediatric emergency medicine. AB - Applying the methods and tools of outcomes research, "evaluation of the impact of health care on the health outcomes or 'end results' of patients and populations," to the clinical domain of emergency services for children offers an important strategic opportunity for addressing the questions that confront all health care services: What works? For which patients? At what cost? From whose perspective? Although the important questions to address are extensive, much of the intersection between emergency services and outcomes research remains unexplored. Important challenges for researchers intrigued by the opportunities at this intersection of fields include: 1) clear definition of the scope of emergency services; 2) consideration of the appropriate end-point of emergency services-the entire episode of illness and/or services provided within the emergency setting; 3) selection and development of measures that incorporate children's and families' perspectives; and 4) a clear focus on linking research findings with strategies for improving outcomes and informed decision making. This essay will provide an overview of accomplishments and challenges from the field of outcomes research, suggest important opportunities for applying existing methods to emergency medical services for children, and identify potential career paths for current and future investigators. PMID- 12135404 TI - Measuring success in the treatment of children in the emergency department setting: process versus outcomes? AB - The current best design practices for clinical studies require consideration of the assessment of end points that combine key processes and outcomes. Process assessment measures events in the pathway on which a more important intermediate or ultimate health outcome depends. Process assessment can be either formative or summative. Outcomes assessment relates to identifying the results related to a process, and is most often defined as the change in a patient's current or future health state. The development of process and outcomes measures for any study is difficult, and children and emergency department settings present unique sets of issues worth highlighting. This article presents an overview of issues relating to choice of process and outcomes measures in studies of pediatric emergency medicine. Asthma care is used to illustrate the complex issues surrounding the measurement of success in management in the emergency department setting. PMID- 12135405 TI - Outcomes research and emergency medical services for children: domains, challenges, and opportunities. AB - Outcomes research focuses on the end results of medical care. This paper describes the specific domains of outcomes research, namely, clinical effectiveness, quality of care, quality of life, patient satisfaction, cost effectiveness, and organization of care, which are the primary domains of outcomes research. Specific examples of pediatric emergency medicine applications are provided for each domain. Outcomes research in the clinical specialties of pediatrics, emergency medicine, and emergency medical services is in an early stage of development relative to that of adult medicine. Thus, the intersection of these clinical specialties, pediatric emergency medicine, presents many challenges and opportunities. PMID- 12135406 TI - Application of measurement tools to pediatric emergency medicine. AB - A focus on children and the limited time frame of the emergency care setting makes pediatric emergency medicine (EM) outcomes research a challenging endeavor. To address the challenges, pediatric EM researchers must choose their risk adjustment and outcomes measures carefully. This article provides guidance to those researchers through a series of questions and answers, with an emphasis on present and future measurement tools and the steps needed to further develop the field. PMID- 12135407 TI - Assessing provider-patient-parent communication in the pediatric emergency department. AB - Patient-provider communication is related to the satisfaction with and process and outcome of emergency department care. Barriers to communication include noise, anxiety, confusion about process, and differences in language and expectations. In the emergency care setting, as in nonacute ambulatory settings, children tend to be left out of discussions of their own care. Both retrospective and real-time methods are available for studying communication during emergency department visits. Well-standardized generic communication assessment tools are applicable to emergency department communication, but assessment tools specific for emergency settings have yet to be developed and standardized. Although the emergency department poses some methodologic difficulties not present in other settings, studies of communications are feasible and likely to yield useful data. PMID- 12135408 TI - Cost-effectiveness analysis and emergency medical services for children: issues and applications. AB - Studies to assess the cost-effectiveness of alternative treatment or prevention strategies for children are far fewer than those for adults. This article highlights specific issues relevant to the conduct of cost-effectiveness analysis (CEA) in pediatric populations following the recommendations of the US Panel on Cost-effectiveness in Health and Medicine and discusses CEA applications relevant to emergency medical services for children. The article addresses whether patient time should be included in measures of costs, whether caregiver time costs are included and measured properly, and most importantly, whether the study can use quality adjusted life years (QALYs) as an outcome measure. Pediatric researchers may be unable to follow the panel's recommendations for including QALYs as an outcome measure in CEA studies involving young children. Developing preference weights applicable to young children may be a productive field for pediatric researchers. Without such efforts, the field of child health services research in general and the field of emergency medical services for children in particular will continue to lag behind adult fields in assessing the costs and outcomes of the services they provide. PMID- 12135409 TI - Children's health status instruments: their potential application in the emergency department. AB - OBJECTIVE: To provide an overview of child health status instruments and their potential application by emergency medicine researchers. CONCLUSIONS: Numerous instruments have recently been developed to measure children's physical, mental, and social health using sound principles of measurement science. These instruments differ from traditional clinical benchmarks of impairment. Health is conceived to be multidimensional and is measured from the child's and/or parents' perspective. Child health status instruments can be used to broaden the evaluation of emergency care in the following ways: 1) to evaluate emergency department (ED) interventions in terms of improvements in children's everyday activities and well-being; 2) to assess the appropriateness of ED disposition and follow-up services; and 3) to detect underlying health and social problems. Using these instruments in the ED will be challenging. Methodologic issues remain regarding the measurement of children's health. The most pressing issues are the absence of a universal definition of children's health, the need to take into account the developmental and social contexts of children, and the difficulties in eliciting information from very young children. In addition, the ED presents its own set of challenges. These include the lack of pre-injury/pre-illness measures, the absence of case-mix severity instruments, and the difficulty of isolating the effect of ED treatment on patients' health. RECOMMENDATIONS: Despite these challenges, it is time to use pediatric health status instruments in emergency medicine. Only with their adoption will instruments improve and advances in methodology occur. As patients, providers, and policy makers increasingly focus their attention on the nonfatal consequences of injury and illness, the broader use of these measures becomes imperative. PMID- 12135410 TI - Patient/parent assessment of the quality of care. AB - Providing patient-centered care is an accepted goal in medicine today. Focusing on the patient has drawn attention to the importance of the interpersonal aspects of care, such as communication between the health care provider and patient, or in the case of health care for children, the parent and child. Patients or parents may be the best or only source of information for assessing the personal aspects of care. In research on children, parents generally report on and evaluate the care. Assessing the interpersonal aspects of care has traditionally been referred to as the measurement of patient satisfaction. The varying expectations of patients and the presence of a ceiling effect on the measures often confound the use of patient satisfaction measures for evaluating the quality of care. One trend is to ask respondents to report on the interpersonal aspects of care, rather than to respond about their level of satisfaction. Studies on the assessment of the interpersonal aspects of health care in emergency departments for children are not plentiful. However, research provides some insight into ways in which emergency departments might improve interpersonal aspects of care for children. These include providing a clear picture to patients and parents of what to expect regarding the length of time they will have to wait, taking a caring approach with children and their parents, and explaining clearly to parents what they need to do to care for the child after discharge. PMID- 12135411 TI - Who's responsible for safe vision on the roads? PMID- 12135412 TI - The 75th anniversary of the World Council of Optometry. PMID- 12135413 TI - Aging, driving and vision. AB - Older people constitute the fastest growing sector of the driving population and are believed to represent a high risk to road safety, given their high crash rate per distance travelled. The crash characteristics of the elderly also differ from those of younger drivers and generally involve multiple vehicles and more complex driving situations. Although the reasons for this deterioration in driving performance are multi-factorial, the age-related changes in vision are likely to be a significant factor, given the important role of vision in driving. This paper provides an overview of some of the complex issues associated with older drivers and considers how the aging changes in visual function might impact on driving performance. Particular emphasis is placed on the literature linking vision to driving, with emphasis on more recent research. The implications of this research for driver licensing and self-regulation of older drivers are also discussed. PMID- 12135414 TI - The detection of diabetic retinopathy by Australian optometrists. AB - BACKGROUND: Diabetes mellitus is a systemic disease affecting approximately 750,000 Australians of whom more than 70,000 are Queenslanders. It can have serious ocular consequences and patients with diabetes require regular eye examinations to determine the degree of ocular involvement and the stage of retinopathy, if present. It is important that optometrists detect diabetic retinal changes and refer appropriately. We sought to determine the proficiency of optometrists at detecting retinal changes caused by diabetes. METHODS: The study comprised four parts: 1. Nineteen randomly recruited Australian optometrists practising in Queensland completed a questionnaire on their experiences seeing patients with diabetes. 2. They examined the ocular fundi of 10 patients. 3. They viewed retinal slides of 12 additional cases. 4. They attended a follow-up seminar on diabetes and the cases. They were informed that the patients did not necessarily have diabetes and instructed not to discuss the condition with the patient or their colleagues. The optometrists were allowed seven minutes per station to examine the patient or the slides and write down their responses before moving to the next station. RESULTS: When the slides and patients were considered together, cases where diabetic retinopathy was present were correctly identified by 94.0 per cent of the optometrists and cases where retinopathy was not present were correctly identified by 93.6 per cent of the optometrists. When all assessments were considered together, the correct detection/differential diagnosis rate was 88.3 per cent. Sub-classification of diabetic retinopathy severity agreed with that of the reference examiners in 58.3 per cent of assessments and there was agreement on management in 79.4 per cent of cases. Of the 22 assessments undertaken by each optometrist, there were, on average, 2.5 errors. CONCLUSION: Randomly selected Australian optometrists are able to detect and grade diabetic retinal changes solely by retinal examination and refer the patients requiring specialist care. PMID- 12135415 TI - Determining magnification for reading with low vision. AB - BACKGROUND: In the past, practitioners have used distance and/or near visual acuity (VA) to calculate required magnification for low vision aids. Magnification was usually under-estimated when compared with the final magnification prescribed. Recent studies have emphasised the importance of acuity reserve in determining the required magnification for optimum reading rate. Two different approaches have been proposed for the appropriate acuity reserve to use in calculating magnification. These are a fixed acuity reserve of 0.3 log unit or an individual determination of optimum acuity reserve. The aim of this study was to investigate the magnification and reading rates with low vision aids selected by the two methods. METHODS: Nineteen low vision subjects with age-related macular degeneration (AMD) who were experienced magnifier-users were recruited. Reading rates and near VA with low vision aids determined by the fixed and individual acuity reserve methods were compared with the same measures made with the subjects' own magnifiers. RESULTS: There were no significant differences in reading rate and near VA measured with low vision aids selected by either the fixed or individual acuity reserve methods or the subjects' own magnifiers. Reading rate with low vision aids was not significantly different from reading rate for large print with conventional near additions. Thus, for experienced users, magnifiers do not cause reduced reading rate. CONCLUSIONS: The fixed acuity reserve method is simple to apply as only near VA and print size of the target reading task are required. For the individual acuity reserve method, reading rates at different print sizes need to be measured. We recommend the use of a fixed acuity reserve (0.3 log unit) for the calculation of required magnification for low vision patients. If near VA or reading rate are not satisfactory with the magnification calculated by this method, individual assessment of required acuity reserve is necessary. PMID- 12135416 TI - Driving into a clearer future. Report of the conference Progress in Automotive Lighting 25-26 September 2001, Darmstadt University of Technology, Germany. PMID- 12135417 TI - Optometrists Association Australia position statement on driver vision standards. PMID- 12135418 TI - Protan colour vision deficiency and road accidents. AB - BACKGROUND: Protans are precluded from holding a commercial driver's licence in Australia because they have a substantially reduced ability to see red lights and have more road accidents involving signal lights. This exclusion has been in place since 1994 but is likely to be abandoned following a current review of medical standards for commercial drivers. This paper reviews the level of risk of road accidents due to protan colour vision deficiency. It also addresses the question of whether it is fair to regard all protans as having a higher risk of road accident because some protans might have a sensitivity to red light that is as good as that of some people with normal colour vision. METHODS: Data of two studies by Verriest and co-workers are re-analysed to estimate the degree of overlap of the protan and colour normal distributions of sensitivity to red light. RESULTS: Field trial data show that protans have a very reduced visual range for red signals compared to colour normal observers but there is considerable variability among both classes of observers and the distributions do overlap. However, some variability is due to differences in observers' choices of a detection criterion, their speed of response and the measurement method. A laboratory study of the spectral sensitivity of protan and colour normal subjects that largely removes these sources' variability shows that all protans have a sensitivity to red light that is less than that of the least sensitive colour normal. CONCLUSION: It is reasonable to conclude that all protans, regardless of the severity of their defect, have a lesser ability to see red signals than colour vision normal observers and for that reason will have a higher risk of road accident. PMID- 12135419 TI - The use of a laser pointer for letter chart acuity assessment. PMID- 12135420 TI - Damien P Smith AM. PMID- 12135421 TI - Vision and learning to read. PMID- 12135422 TI - Cellular methods used to evaluate the immune response in transplantation. AB - Cellular assays have been developed to test for various effector, cytotoxic, and regulatory functions of T cells and have been used throughout the history of clinical transplantation to assess the immune profile of solid organ and marrow recipients. One goal of these cellular studies has been to determine if posttransplant changes in the donor antigen-specific cellular response could predict good and poor graft outcome, thereby allowing for individualization of immunosuppression. This review outlines the use of established and newly developed cellular assays to assess the dynamic processes of the posttransplant immune response and to provide insights into the mechanisms involved and potential points for intervention. PMID- 12135423 TI - Establishment of a quantitative ELISA capable of determining peptide - MHC class I interaction. AB - Many different assays for measuring peptide-MHC interactions have been suggested over the years. Yet, there is no generally accepted standard method available. We have recently generated preoxidized recombinant MHC class I molecules (MHC-I) which can be purified to homogeneity under denaturing conditions (i.e., in the absence of any contaminating peptides). Such denatured MHC-I molecules are functional equivalents of "empty molecules". When diluted into aqueous buffer containing beta-2 microglobulin (beta2m) and the appropriate peptide, they fold rapidly and efficiently in an entirely peptide dependent manner. Here, we exploit the availability of these molecules to generate a quantitative ELISA-based assay capable of measuring the affinity of the interaction between peptide and MHC-I. This assay is simple and sensitive, and one can easily envisage that the necessary reagents, standards and protocols could be made generally available to the scientific community. PMID- 12135424 TI - Non-mutated tumor-rejection antigen peptides elicit type-I allergy in the majority of healthy individuals. AB - IgE-mediated type-I allergy is generally considered to be a hypersensitivity reaction to foreign antigens, and it is believed that self-antigens do not evoke this type of allergy. We report here, for the first time, that non-mutated self antigen peptides identified as tumor-rejection antigen peptides recognized by HLA class I-restricted and tumor-specific cytotoxic T lymphocytes (CTLs) elicited a type-I allergy in the majority of healthy individuals. Peptide-specific IgE was detectable in sera from certain cases, although the levels did not always correlate with those of type-I allergy. Repeated vaccinations of nonallergic peptides derived from the same antigens possessing allergic peptides resulted in the suppression of both allergic peptide-specific IgE responses and type-I allergy, providing evidence for a new approach to the development of peptide based desensitization. PMID- 12135425 TI - Identification of a novel gp100/pMel17 peptide presented by HLA-A*6801 and recognized on human melanoma by cytolytic T cell clones. AB - Melanoma-associated peptides recognized by cytolytic T lymphocytes (CTL) in the context of several histocompatibility leukocyte antigens (HLA) are required for the development of specific immunotherapies. Using a transient transfection assay into COS-7 cells, we identified the gp100/pMel17 melanosomal protein as the shared antigen recognized by three independent CD8+ CTL clones in HLA-A*6801 restricted fashion. This finding was confirmed by the correlation between lack of gp100/pMel17 protein in a number of HLA-A*6801-positive melanomas and their resistance to lysis/cytokine production by the specific effectors. The gp100/pMel17 antigenic epitope was identified based on recognition of subfragments and on a computer-based prediction algorithm. Among a panel of gp100/pMel17-derived synthetic peptides only the 10-mer HTMEVTVYHR (gp100/pMel17182-191) induced tumor necrosis factor (TNF) release by CTL clones when pulsed on suitable target cells whereas both the 10-mer and the shorter 9 mer gp100/pMel17183-191 sensitized the same antigen-pulsed cells to lysis. In conclusion, the identification of the HTMEVTVYHR peptide will extend to HLA A*6801 melanoma patients the possibility to exploit gp100/pMel17 melanosomal protein for experimental and clinical studies. PMID- 12135426 TI - Simultaneous genotyping of single nucleotide polymorphisms in the IL-6, IL-10, TNFalpha and TNFbeta genes. AB - Single nucleotide polymorphisms (SNP) in the human IL-6, IL-10, TNFalpha and TNFbeta genes have been associated with gene function and susceptibility to disease. In this study, primers containing mismatches at 1-3 nucleotide positions were designed to incorporate a new restriction site recognized by endonucleases AlwNI, BcgI, BglI, BsaBI, BslI, BstXI, EcoNI or XcmI for genotyping SNPs in the IL-6 gene (position - 174), IL-10 gene (positions -592 and -1082), TNFalpha gene (positions -238, - 308 and -863) and TNFbeta gene (position + 249) by mismatched polymerase chain reaction and restriction fragment length polymorphism (PCR/RFLP). Our results show that appropriately designed BslI-based mismatched PCR/RFLP assays can be successfully used to determine the genotypes for approximately 40% of SNPs. The mismatched PCR strategy can be coupled with multiplex-amplification to enable simple and rapid determination of several SNP genotypes in a single reaction. PMID- 12135427 TI - TNFalpha-308A associated with shorter intervals of Plasmodium falciparum reinfections. AB - A mutation at position -308 of the tumor necrosis factor alpha (TNF-308A) gene promoter has previously been associated with particular manifestations of infectious and non-infectious diseases. In a longitudinal study on malariological parameters in Gabon, TNF promoter variants of 98 children initially presenting with severe Plasmodium falciparum malaria, followed by a total of 504 reinfection events within 52 months, and 100 children initially presenting with mild malaria followed by a total of 342 reinfections were analyzed. Symptomatic P. falciparum reinfections occurred more quickly (median 11 weeks) in carriers of the TNF-308A allele, with severe malaria at enrollment, compared to carriers of other TNF promoter variants (median 16 weeks). The results show that this particular TNF promoter allele increases the risk of reinfection with the malaria parasite P. falciparum. PMID- 12135428 TI - Feasibility of using genetic linkage analysis to identify the genes encoding T cell-defined minor histocompatibility antigens. AB - We have evaluated the utility of genetic linkage analysis to identify genes that encode minor histocompatibility antigens using vaccinia virus vectors as a simple and convenient method for transient expression of class I MHC molecules in lymphoblastoid cell lines. As a test case, we used a CTL clone that recognizes HA 8, a minor histocompatibility antigen encoded by the KIAA0020 gene and presented by HLA-A*0201. EBV-transformed B cell lines from individuals in three large pedigrees from the CEPH reference family collection were infected with a recombinant vaccinia virus vector encoding an HLA-A*0201 transgene, which led to high level expression of the MHC restricting allele HLA-A*0201 on the cell surface. HA-8 expression in the vaccinia-infected target cells was then determined using standard in vitro cytotoxicity assays. Pairwise linkage analysis of the segregation of HA-8 expression in these pedigrees demonstrated that the HA 8 gene was tightly linked with a cluster of marker loci located on the distal portion of chromosome 9p. Analysis of 9p marker haplotypes for individuals in the three families identified several individuals with recombinant haplotypes, and these recombination events were used to refine the precision of the HA-8 gene localization further. The data collectively indicate that the HA-8 gene is localized to a 10.3 cM (corresponding to 3.9 Mb) interval of distal 9p that is thought to encode at least 11 genes, including KIAA0020. These results demonstrate that linkage analysis can be used to map minor histocompatibility genes with high precision and accuracy. Over the next years, refinement and annotation of the human genome sequence will undoubtedly increase the utility of linkage analysis as a tool for identifying minor histocompatibility antigen genes. PMID- 12135429 TI - Allele specific PCR for the minor Histocompatibility antigen HA-2. AB - The hematopoietic system restricted minor Histocompatibility antigen (mHag), HA 2, is encoded by the novel human class I Myosin gene, MYO1G, located on the short arm of chromosome 7. The HA-2 encoding region is di-allelic and comprises the HLA A2 restricted T cell epitope YIGEVLVSV (HA-2V) and its allelic counterpart YIGEVLVSM (HA-2M). We designed a sequence specific PCR (SSP) for both HA-2 alleles. The HA-2 genomic typing results were compared with the HA-2 CTL phenotyping in three families and revealed exact correlation. The mHag HA-2 SSP can be incorporated in DNA based typing protocols. PMID- 12135430 TI - Characterization of a novel MICA allele, MICA*047. AB - We report the identification of a novel MICA allele, MICA*047. It was initially detected because of an unusual hybridization pattern with sequence-specific oligonucleotides (SSOP) in a normal subject of Caucasian origin. Cloning and sequencing of both strands, and comparison of the sequence with previously defined MICA alleles, revealed that the new allele is similar to MICA*041 except for one nucleotide substitution at position 811 (C-->G). It appears that this new allele could have been generated by an interallelic sequence exchange between MICA*011 and MICA*0411. PMID- 12135431 TI - Association of NRAMP1 promoter gene polymorphism with the susceptibility and radiological severity of rheumatoid arthritis. AB - The natural resistant-associated macrophage protein 1 (NRAMP1) has been proposed as a candidate gene for the susceptibility to autoimmune diseases. In this study, the possible role of the functional polymorphism located at the promoter region of NRAMP1 gene in the susceptibility and clinical outcome of rheumatoid arthritis (RA) was investigated. A total of 141 Spanish RA patients and 194 controls previously typed for HLA-DRB1* were genotyped for the NRAMP1 polymorphism. No significant differences in the distribution of frequencies among RA patients and controls were observed. Nevertheless, when patients and controls were stratified according to their HLA shared epitope (SE) status, an increase of 2/2 genotype among SE-negative (SE-) patients with respect to SE- controls was observed (23% vs 7%, OR = 3.74, 95% CI 1.31-10.72). In addition, the possible role of this polymorphism in the clinical course of RA was investigated in a subgroup of 82 patients who were prospectively followed during a mean of 9 years. After follow up, an increase of patients with the homozygous 2/2 genotype was detected among those with severe small joint radiological involvement: 73% of patients 2/2 had a severe form in contrast to 37% of patients with the genotype 2/3 and 30% of patients bearing 3/3 OR = 5.45, 95% CI 1.14-34.24). In conclusion, NRAMP1 gene promoter polymorphism could influence the radiological severity of rheumatoid arthritis and disease susceptibility, particularly in individuals lacking HLA linked risk factors. PMID- 12135432 TI - No evidence for association of the inducible nitric oxide synthase promoter polymorphism with Trypanosoma cruzi infection. AB - The purpose of the present study was to address the possible contribution of the (CCTTT)n microsatellite polymorphism in the NOS2 promoter region to the susceptibility to chronic Trypanosoma cruzi infection and to Chagas' disease related cardiomyopathy. We determined the (CCTTT)n genotypes in a sample of 76 serologically positive chagasic individuals and in 78 healthy controls. No statistically significant differences were observed between total chagasic patients and healthy controls with regard to frequency of the (CCTTT)n microsatellite repeat of any given length. Likewise, we found no differences in the distribution of the (CCTTT)n microsatellite repeats between seropositives without manifestations of the disease and those with chagasic cardiomyopathy. Our data suggest that the NOS2 promoter pentanucleotide microsatellite polymorphisms analyzed do not play a major role in the pathogenesis of chronic T. cruzi infection in this Peruvian sample. PMID- 12135433 TI - Six new HLA class I alleles detected by PCR-SSO genotyping. AB - This paper describes six new alleles; A*0240, A*2614, B*3924, B*4425, Cw*0807 and Cw*12023, which were discovered during routine genotyping with sequence specific oligonucleotides (SSO's). Five of the new alleles have changes in residues which belong to the antigen binding site of the HLA protein. These new variants may have altered antigen binding properties and may cause differential immunological responses that could affect transplantation outcome1. PMID- 12135434 TI - Novel HLA-A locus alleles including A*01012, A*0306, A*0308, A*2616, A*2617, A*3009, A*3206, A*3403, A*3602 and A*6604. AB - This paper describes 10 novel HLA-A alleles that have been characterized by DNA sequencing. Seven alleles, A*0308, A*2616, A*3009, A*3206, A*3403, A*3602 and A*6604 carry motifs observed in other HLA-A alleles, suggesting that gene conversion has created this diversity. The remaining three alleles, A*01012, A*0306 and A*2617, contain polymorphisms not previously found in any "classical" class I allele. All alleles were identified due to unexpected probe hybridization patterns during routine SSOP typing. Exons 2 and 3 of each allele were subsequently characterized by DNA sequencing. PMID- 12135435 TI - Cloning and complete sequence of a novel HLA-A null allele, A*0253 N, with a termination codon generated by a C to G mutation in exon 2. AB - A novel HLA-A null allele, A*0253 N, has been identified in two generations of a Chinese family using combined serological and molecular cloning approaches. Full length genomic DNA sequencing indicated that this new allele differs from HLA A*02011 by a single C to G substitution at nucleotide position 324 in exon 2. This mutation results in an amino acid change from a tyrosine codon to a stop codon at position 108. A PCR-SSP based method was developed to distinguish A*0253 N from A*02 alleles. No further individuals of A*0253 N were found in 718 Chinese blood donors who carry the HLA-A*02 allele1. PMID- 12135436 TI - Identification of the novel allele B*4427 and a confirmatory sequence (B*44022). AB - This report presents a novel allele, HLA-B*4427, which was identified in a bone marrow donor of Caucasian origin, and a confirmatory sequence (B*44022). Sequence analysis revealed the new allele differs from B*44021 by a single nucleotide exchange at position 668 (C-->T), which is located in exon 4. At the protein level, it is the only B*44 variant to produce an Ala in place of a Val at codon 199. Its structure suggests that it may have originated from a point mutation in B*44021 or by gene conversion with a variety of HLA-B alleles. Cloning and sequencing of the allele B*44022 revealed a sequence identical to B*44021 and B*44 exon 4, with the codon GTC (Val) in position 199. PMID- 12135437 TI - Identification of a new HLA-B allele, B*3705 containing a Bw6 sequence motif. AB - HLA-B37, which is Bw4 type antigen frequently found in linkage disequilibrium with A1, Cw6 and DR10 in all ethnic groups, generally has a very low frequency all over the world. We report a new HLA-B*37 allele, B*3705, identified in two potential bone marrow donors in the Korean population. B*3705, which has the Bw6 nucleotide segment, differs from B*3701 in three nucleotide positions: 311, 317 and 319 in exon2. The serological profile of B*3705 did not exhibit the B37 specificity. The putative haplotype associated with B*3705 in the Korean population could be A*02-Cw*0602-DRB1*1001-DQA1*0101-DQB1*0501-DPB1*02011. PMID- 12135438 TI - Two new HLA-B alleles, B*4422 and B*4704, identified in a study of families with autoimmunity. AB - HLA-B typing of approximately 1 262 individuals from a study of 372 simplex families with multiple sclerosis has led to the identification of two new alleles (HLA-B*4422 and HLA-B*4704). Sequencing confirmed that both of these new alleles represent novel combinations of previously described sequence motifs, reinforcing the notion that recombination and/or gene conversion-like events play an important role in generating HLA allelic diversity. The identification of these new alleles brings the total number of HLA-B alleles to 465. PMID- 12135439 TI - Identification of a new HLA-B null allele, B*1817 N, in a family. AB - We describe a new HLA-B null allele found in a daughter and her mother. This null allele was due to a mutation at position 41 of exon 1 which resulted in a premature stop codon. This new null allele was officially named HLA-B*1817N*. PMID- 12135440 TI - Identification of a new HLA-DPB1 allele,HLA-DPB1*9001. AB - During routine typing of a potential bone marrow donor, a new HLA DPB1 allele was identified. The new allele, officially named HLA DPB1*9001, differs from HLA DPB1*01011 in the second hypervariable region, where a single nucleotide substitution in position 191 changes the codon 35 from TAC to TTC with a predicted amino acid change from Tyr to Phe. PMID- 12135441 TI - Novel HLA-DRB1 alleles revealed by sequencing based typing, DRB1*04053 and DRB1*1143. AB - We report the discovery of two HLA-DRB1 alleles by sequencing based typing (SBT). DRB1*04053 differs from previously reported DRB1 alleles by a single synonymous nucleotide substitution, resulting in a unique polymorphism at codon 93. DRB1*1143 differs from previously identified DRB1 alleles by a single non synonymous nucleotide substitution, resulting in a polymorphism observed in other DRB1 and DRB3 alleles1. PMID- 12135442 TI - Identification of a novel HLA-DRB1 allele, DRB1*0108, by sequence-based DRB typing in two siblings. AB - We report here a novel DRB1 allele identified during sequence-based HLA-DRB typing. This allele was detected during routine HLA typing of a patient and his family prior to bone marrow transplantation. The new allele, DRB1*0108, was found in the patient and in a brother. Molecular cloning and sequencing confirmed that the new DRB1 allele is identical to DRB1*0101 at exon 2 except for a single nucleotide substitution at codon 37 (TauCC-->TauAlphaC), changing the encoded serine to tyrosine. This position of the beta1 domain lies in the floor of the antigen-binding groove and shows the highest polymorphism among DRB1 alleles. PMID- 12135443 TI - Knee magnetic resonance imaging scans in daily use. PMID- 12135444 TI - Seven years follow-up after ankle inversion trauma. AB - During one year all ankle inversion injuries seen at the acute ward of our institution were divided into grades of severity and classified according to the maximal area of tenderness at the time of clinical examination. Seven years later 648 of the subjects (91%) evaluated their ankle with the help of a questionnaire. Location of maximal tenderness at the time of injury was: lateral fibular ligaments 61%, lateral midfoot ligaments 24%, base of the fifth metatarsal/peroneal tendons 5% and combined lesions 8%. 39% were considered minor, 46% were moderate, and 15% severe. All cases followed a functional treatment protocol. Seven years post- injury 32% reported chronic complaints of pain, swelling or recurrent sprains. 72% of the subjects with residual disability reported that they were functionally impaired by their ankle - in most cases a question of not performing sports at a desired level. 4% experienced pain at rest and were severely disabled. 19% were bothered by repeated inversion injuries - 43% of these subjects felt that they could compensate by using an external ankle support. There was no correlation between the severity of the sprain as judged at the time of injury and the frequency of residual disability or between the area of maximal tenderness at the time of injury and the area of maximal pain at the time of follow-up. PMID- 12135445 TI - Iontophoresis with cortisone in the treatment of lateral epicondylalgia (tennis elbow)--a double-blind study. AB - Lateral epicondylalgia (tennis elbow) is a common dysfunction of the arm. Because there is no agreement concerning the pathophysiology, several modes of treatments have been tried and one of the most common is local steroid injection. Iontophoresis using corticosteroids is a fairly new method recommended in the treatment of lateral epicondylalgia and has become popular owing to impression of superiority compared to local injections-noninvasive, painless and nontraumatic. The aim of this double-blind prospective, randomized study was to evaluate the short- and the long-term pain-relieving effect of corticosteroid iontophoresis in lateral epicondylalgia. Sixty-four patients suffering from lateral epicondylalgia were consecutively randomized into two groups for corticosteroid or placebo iontophoresis. The patients were treated four times during 2 weeks. Follow-ups were done the day after the final treatment and after 3 and 6 months. Twenty three patients dropped out before the 3-month follow-up because they wanted to complement the treatment or replace it with other treatments. No significant difference between the corticosteroid group and the placebo group in relation to subjective and objective outcome could be observed after the treatment period or at the follow-ups. In fact, both groups improved throughout the study. The results of the present study do not support the use of corticosteroid iontophoresis in lateral epicondylalgia. PMID- 12135446 TI - Alteration in albumin level during modified muscular activity. AB - We have previously reported that albumin protein is increased in the atrophied muscle induced by hindlimb immobilization. The purpose of this study was to evaluate the effects of several disuse models on albumin protein and mRNA levels in mice skeletal muscle and to investigate whether the elevated amount of albumin returns to control level by muscular activity increased by hindlimb remobilization. Western blot analysis revealed that hindlimb immobilization, denervation, and tenotomy, except for hindlimb unloading, significantly increased albumin levels in soleus muscles by 2.1-, 1.9- and 2.0-fold, respectively (P < 0.001). Immunohistochemical analysis showed that albumin protein accumulates in the widened extracellular space. Reverse transcription-polymerase chain reaction (RT-PCR) assay revealed albumin gene expression to be downregulated in all disuse models relative to control level. During hindlimb remobilization, the amounts of albumin protein appeared to remain higher level after 3 and 7 days and had returned to control level after 14 days and muscle mass, the amounts of myosin heavy chain, and actin proteins seemed to restore control levels after 21 days. These results indicate that the amount of interstitial albumin protein may be modulated by muscular activity. PMID- 12135447 TI - The use of MRI scan of knee injuries in an emergency department. AB - We present a retrospective study of the number of arthroscopically verified total meniscus and cruciate ligament tears seen in our Emergency department one year before, and one year after introducing magnetic resonance imaging (MRI) as a diagnostic tool. The number of total ruptures of the anterior cruciate ligament increased from 34 (1.7% of total 2003 patients) to 67 (2.9% of total 2277). The number of ruptures of the medial meniscus increased from 42 (2.1%) to 87 (3.8%). The number of ruptures of the lateral meniscus remained unchanged. In the first 12 months after introducing MRI we ordered 513 (23%) scans of the total number of 2277 patients with an acute knee condition. Thirty-nine (8%) of these were described with no pathology, 227 (44%) showed total meniscus or cruciate ligament tears. The remaining 247 (48%) were mainly degenerative or partial injuries. The cost per additional new diagnosis was approximately USD 800. The introduction of MRI has enabled us to identify a larger number of patients with meniscus and cruciate ligament injuries. PMID- 12135448 TI - Clinical decision making in the acutely injured knee based on repeat clinical examination and MRI. AB - To determine the correlation between Magnetic Resonance Imaging (MRI) and clinical examination of the knee after an acute injury, and to see to what extent MRI affected the planned treatment, we examined 90 consecutive patients in a prospective study, clinically and with an extremity Magnetic Resonance (MR) scanner. The number of meniscal lesions, bone bruises and osteochondral lesions found on MRI was significantly higher than the clinical examinations indicated. Despite this, the treatment was only changed in 6 cases. In no case did MRI prevent a planned arthroscopy. MRI may reveal many clinically silent changes in the knee, also after minor injuries. The significance of these MRI findings must await long-term follow-up. PMID- 12135449 TI - Effects of endurance training on tissue glutathione homeostasis and lipid peroxidation in streptozotocin-induced diabetic rats. AB - The aims of our study were to assess whether endurance training strengthens glutathione-dependent antioxidant defenses and decreases oxidative stress in experimental diabetes. Streptozotocin-induced diabetic rats were divided into trained and untrained groups, which were further divided into resting and acute exercise groups. Endurance training consisted of treadmill running for 8 weeks. For acute exhaustive exercise, graded treadmill running was conducted until exhaustion. Eight weeks' treadmill training increased the endurance, favorably decreased lipid peroxidation as measured by thiobarbituric acid reactive substances but not conjugated dienes levels in kidney and vastus lateralis muscle and upregulated glutathione peroxidase in red gastrocnemius muscle. However, it adversely decreased total glutathione level and glutathione peroxidase activity in kidney. Acute exhaustive exercise up-regulated glutathione peroxidase activity in liver. Endurance training did not prevent the increase in thiobarbituric acid reactive substances level in liver due to acute exhaustive exercise. Activities of glutathione disulfide reductase and glutathione S-transferase were not affected. Even though endurance training appeared to upregulate glutathione dependent antioxidant defense in skeletal muscle and to decrease lipid peroxidation in kidney and vastus lateralis muscle as measured by TBARS, our results suggests that beneficial effects of 8 weeks of endurance training are limited in this rat model of uncontrolled diabetes mellitus. PMID- 12135450 TI - Anaerobic power and muscle strength characteristics of 11 years old elite and non elite boys and girls from gymnastics, team handball, tennis and swimming. AB - The aim of the present investigation was to study the possible effects of specificity of training on muscle strength and anaerobic power in children from different sports and at different performance levels in relation to growth and maturation status. Hundred and eighty-four children of both gender participating either in swimming, tennis, team handball or gymnastics were recruited from the best clubs in Denmark. Within each sport, the coach had divided the children into an elite (E) and non-elite (NE) group according to performance level and talent. Tanner stage assessment and body weight and height measurements were performed by a physician. The anaerobic performances were assessed by Wingate tests and jumping performance in squat jump (SJ), countermovement jump (CMJ) and drop jump (DJ) from two heights. Most of the differences between groups in Wingate performance disappeared when the data were normalised to body mass. The gymnasts were the best jumpers and their superiority were increased in the more complex motor coordination tasks like DJ. The results may indicate some influence of training specificity, especially on the more complex motor tasks as DJ and there may be an effect of training before puberty. The performance in the less complex motor tasks like cycling and SJ and CMJ may also be influenced by specific training, but not to the same extent, and heritance may be an important factor for performance in these anaerobic tasks. PMID- 12135451 TI - Stability of leisure-time physical activity during adolescence--a longitudinal study among 16-, 17- and 18-year-old Finnish youth. AB - Participation in physical activity during childhood and adolescence is frequently mentioned as one factor likely to promote a more active lifestyle in adulthood with its health benefits. We studied the changes in leisure-time physical activity pattern and self-reported fitness during a three-year period in adolescence and investigated whether the type of sports has an effect on stability of physical activity at leisure. A questionnaire with identical physical activity items was sent to Finnish twins on their 16th and 17th birthdays and 6 months after the 18th birthday. A total of 1338 boys and 1596 girls responded to all three questionnaires, with response rates of 73.6% and 86.5%. The proportions of very active adolescents and adolescents with very good self-reported fitness seem to be alike at each age. Among girls, 23.7% to 27.7% reported being very active (4-5 times a week) and 13.7% to 15.1% considered their physical fitness to be very good at the ages of 16, 17 and 18. Among boys, the comparable percentages were 31.5% to 35.5% and 30.6% to 34.4%. However, the longitudinal three-year follow up showed substantial changes over time among individuals from one physical activity group to another. Only 19.1% of boys and 11.2% of girls were persistent exercisers (i.e., very active on all three years) and 15.6% of boys and 5.1% of girls were persistently fit (i.e., very good self reported fitness on all three years). Stability of leisure-time physical activity was highest among those who participated in several different types of sports. Among boys the proportion of persistent exercisers was highest for those who participated in cross-country skiing, jogging and body-building (22.0-41.5%) and among girls for those who participated in ball games (11.9-28.6%). Those who participated in organised sports were more often persistent exercisers than those who did not (odds ratio = 13.2 for boys (CI 9.4-18.7) and 8.9 for girls (CI 6.4 12.5)). Also, those who participated in organised sports were more often persistently fit (odds ratio = 7.3 for boys (CI 5.2-10.2) and 10.4 for girls (CI 6.4-16.9). Adolescents are recommended to participate in and try different types of sports, and especially for girls ball games would appear to favour long-term maintenance of leisure-time physical activity. PMID- 12135452 TI - A randomized controlled trial of rehabilitation after hospitalization in frail older women: effects on strength, balance and mobility. AB - When frail older people become acutely ill, they are at increased risk of further functional deterioration and rehabilitation is needed to restore functioning. The effects of an out-patient multicomponent training program including strength training after hospitalization were studied in a randomized controlled trial. Sixty-eight women (mean age 83.0 +/- 3.9 years) who were hospitalized due to an acute illness and were mobility impaired at admission were randomized into training (N = 34) and home exercise (N = 34) groups. Maximal voluntary isometric strength of knee extension and hip abduction, dynamic balance, and maximal walking speed were measured before and after the 10-week training period, and 3 and 9 months after the end of the intervention. After the intervention, significant improvements were observed in the training group compared to the home exercise group in the maximal voluntary isometric knee extension strength (20.8% vs. 5.1%, P= 0.009), balance scale (+ 4.4 points vs. -1.3 points, P= 0.001) and walking speed (+ 0.12 m s-1 vs. -0.05 m s-1, P= 0.022). Effects on knee extension and hip abduction strength, balance and walking speed were observed 3 months later, and some effects on hip abduction strength (9.0% vs. -11.8%, P= 0.004) and mobility were still apparent even 9 months after the intervention. PMID- 12135454 TI - Embedding in Spurr's resin is a good choice for immunolabelling after freeze drying as shown with chemically unfixed dendritic cells. AB - Immunocytochemical reactions on biological specimens depend on many factors, the most crucial one being the maintenance of antigenicity. Antigens are vulnerable at each stage during preparation for electron microscopy. One of the least traumatic methods of preparing biological tissues for post-embedding immunolabelling includes the following steps: (1) physical stabilization of the native biological material by rapid freezing (cryofixation) and keeping the immobilized biological sample at low temperature, thereby avoiding any movements of water, ions and macromolecules; (2) dehydrating the frozen biological material by freeze-drying at low temperature; (3) embedding of the dehydrated specimen. Here we show that embedding of chemically unfixed dendritic cells in Spurr's resin after cryofixation and freeze-drying enables the conservation of fine ultrastructure without cell distortion or shrinkage. Furthermore, we demonstrate the feasibility of protein localization in ultrathin sections by immunolabelling of the major histocompatibility class II molecules. PMID- 12135455 TI - Freeze-dried and resin-embedded biological material is well suited for ultrastructure research. AB - Transmission electron micrographs of different biological material, cryofixed, freeze-dried and embedded in Spurr's resin, in Epon, or in Lowicryl, are presented. The structure preservation obtained either without or with application of chemical fixatives after drying showed that freeze-dried embedded specimens are particularly well suited for new morphological, immunocytochemical and microanalytical studies aimed at detecting the life-like subcellular distribution of mobile macromolecules and ions. The results also indicate that the removal of cell water by freeze-drying from the areas of best cryofixation is relatively slow. Ultrathin sections of well cryofixed biological material embedded after freeze-drying in Spurr's resin or Epon reveal cellular plasma phases with very fine granularities and well defined membranes in negative contrast. This may be due to the preservation of the original structure of cellular macromolecules with a considerable amount of their hydration water. Sublimation studies with differently hydrated and cryofixed macromolecules are suggested to settle this issue. PMID- 12135456 TI - Simultaneously identifying S-phase labelled cells and immunogold-labelling of vinculin in focal adhesions. AB - A new combination of autoradiography and immunolabelling techniques is presented that allows the simultaneous identification of both S-phase cells and their focal adhesions using scanning electron microscopy. The technique allows both labels to be discerned visually by their unique shapes and location within and on the cell. S-phase cells were radio-labelled with a pulse of tritiated thymidine, selectively incorporated into synthesizing DNA. The cells were then immunogold labelled for the focal adhesion protein, vinculin, prepared for autoradiography, and embedded in resin. The resin was then polymerized before removing the substrate, to expose the embedded cell undersurface. Electron-energy 'sectioning' of the sample by varying the accelerating voltage of the electron beam allowed separate S-phase cell identification in one electron-energy 'section' and visualization of immunogold label in another 'section', within the same cell. As a result of applying this technique it was possible to positively identify S phase cells and immunogold-labelled focal adhesions on the same cell simultaneously, which could be used to quantify focal adhesion sites on different substrates. PMID- 12135457 TI - Video-rate confocal endoscopy. AB - Rigid endoscopes provide high quality optical images of reasonably accessible regions of the inner body, especially regions such as the aero-digestive and genital tracts. In order to enhance the versatility of these instruments we describe a development that permits confocal endoscopic images to be obtained - along with traditional endoscopic images - in real-time, from within the living patient. The system is based around a host lenslet-array tandem scanning microscope, which is capable of producing images viewed directly by eye. These types of confocal microscope are configured for fluorescence imaging together with laser illumination. Hard and soft tissues in the mouth were imaged using this combined system. PMID- 12135458 TI - Sampling effects influence heights measured with atomic force microscopy. AB - The atomic force microscope (AFM) is an exquisitely delicate probe measuring the height of a specimen at discrete sampling points in a fixed two-dimensional (2D) raster. The resulting topograph is a 2D digital image, with each pixel representing a distinct height measurement. The height of an object is determined as the average of the maximum heights measured above the supporting surface. We show that such object heights derived from a variety of organic samples depend critically on the sampling or pixel size of the 2D raster. It is concluded that to obtain accurate specimen heights, the pixel size must be small enough to resolve submolecular structures and thus ensure representative sampling of the height variation on the surface. PMID- 12135459 TI - Electron energy-loss near-edge structure studies at the atomic level: reliability of the spatial difference technique. AB - The spatial difference technique was applied to determine the interface-specific components of electron energy-loss near-edge structures and the results are in good agreement to those obtained by atomic column resolved measurements averaged over one atomic layer. In previous studies an experimental set-up had been chosen where the scanning areas, which are used to measure electron energy-loss spectra, were symmetrical in respect to the location of the investigated interfaces. In the present study, an asymmetric setting of the scanning areas was applied, which allows the interfacial signals to be determined directly. Comparing the results of the different measurements shows that the spatial difference technique is valid and can be used to obtain information about the electronic structure of interfaces where single atomic column resolved measurements are not yet possible. PMID- 12135460 TI - Energy dispersive spectroscopy analysis of aluminium segregation in silicon carbide grain boundaries. AB - The aluminium distribution in polycrystalline SiC hot-pressed with aluminium, boron and carbon additives was studied using X-ray energy-dispersive spectroscopy (EDS) and transmission electron microscopy (TEM). The Al excess in homophase SiC grain boundary films was determined, taking into account dissolved Al in the SiC lattice. In the spot-EDS analysis, an electron beam probe with a calibrated diameter was formed, and the total beam-specimen interaction volume was defined, taking the beam spreading through crystalline TEM foil into consideration. EDS spectra were collected from regions containing intergranular films and adjacent matrix grains, respectively. A theoretical treatment was presented and experimental errors were estimated, with a further discussion about the effects of foil thickness. Experimental examples are given, followed by statistical EDS analyses for grain boundary films in SiC samples hot-pressed with increased amounts of Al additions. The results demonstrated a substantial Al segregation in the nanometer-wide intergranular films in all samples. Al additions higher than 3 wt% saturated the Al concentrations in SiC grains and in grain boundary films. The effect of foil thickness, and the parameters for determining the optimum incident beam diameter in the EDS analysis are discussed. PMID- 12135461 TI - On the orientational analysis of planar fibre systems from digital images. AB - The orientational characteristics of fibres in digital images are studied. The fibres are modelled by a planar Boolean model whose typical grain is a thick (coloured) fibre. The aim is to make stereological inference on the rose of directions of the unobservable central fibres from observations made on a digital image of the thick fibres. For central fibres, the relation between the rose of directions and the point intensity, observed on a sampling line, is known. We derive, under regularity conditions, the relation between the unobservable point intensity and the scaled variogram observed on the line in a binary and a greyscale image. Using such a relation, it is possible to draw inference about the rose of directions from the scaled variogram, which is easy and quick to determine in a digital image. PMID- 12135462 TI - Reporting tumor thickness for cutaneous squamous cell carcinoma. PMID- 12135463 TI - New prognostic factors of cutaneous melanoma: a review of the literature. PMID- 12135464 TI - Human telomerase RNA component expression in Spitz nevi, common melanocytic nevi, and malignant melanomas. AB - BACKGROUND: Telomerase is a ribonucleoprotein DNA polymerase that is capable of synthesizing telomeres onto the ends of chromosomes. The cumulative loss of telomerase activity is believed to be associated with cell senescence. Telomerase activity has been shown to be higher in malignant melanomas than in common melanocytic nevi. The aim of the present study was to elucidate the pattern of expression of the human telomerase RNA (hTER) component in routinely processed specimens of Spitz nevi, malignant melanomas, and ordinary melanocytic nevi. METHODS: Ten specimens of each type of tumor were studied, using an in situ hybridization technique. RESULTS: All three types of tumors demonstrated moderate to high intensities of hTER expression, usually in more than half of the tumor cells, and the majority of the studied lesions in each group did not show stratification of staining. The hTER component was also detected in the epidermis, sweat glands, and pilosebaceous units. CONCLUSIONS: hTER levels do not necessarily correlate with the level of telomerase activity, and the level and pattern of hTER expression are not useful as an adjunct to the histologic differential diagnosis of Spitz nevi from melanocytic nevi and malignant melanomas. PMID- 12135465 TI - Vascular endothelium express CS-1 fibronectin in allergic contact dermatitis. AB - BACKGROUND: Allergic contact dermatitis (ACD) is a common human dermatosis in which not all the mechanisms involved in its pathogenesis have been elucidated. OBJECTIVE: To study the expression of CS-1 fibronectin, TARC and Th1-associated chemokine receptors in biopsies from allergic patch test reactions. MATERIAL AND METHODS: Thirteen patients already diagnosed with ACD were challenged on the back with the antigen responsible of the disease and macroscopic responses and biopsies taken after 48 h. Skin biopsies from negative control challenge sites, AD and ICD were also taken. Samples were fixed, embedded in paraffin wax and processed in order to perform histological and immunohistochemical studies. RESULTS: All subjects with ACD showed a positive clinical response and a perivascular mononuclear cell infiltration at 48 h, which was not seen in the negative controls. The majority of skin-infiltrating cells were CD4+ and CD8+ and up to 54% or 40% of them expressed CXCR3 or CCR5, respectively. We also showed expression of CS-1 fibronectin in inflamed endothelial cells not only in ACD but also in AC and ICD. In contrast TARC was only expressed in ACD and AC. CONCLUSION: We showed for the first time that CS-1 fibronectin is expressed in dermal vessels from allergic patch tests positive reactions, as well as irritant and atopic skin lesions. PMID- 12135466 TI - Expression of polo-like kinase (PLK1) in thin melanomas: a novel marker of metastatic disease. AB - BACKGROUND: The maximum thickness of a primary malignant melanoma as measured by Breslow's method is currently the most important prognostic factor. However, some thin melanomas (< or= 0.75 mm), which should have an excellent prognosis according to Breslow, can be lethal due to their ability to metastasize. METHODS: In our study, thin malignant melanomas (< or= 0.75 mm) from 36 patients were analyzed with immunohistochemical techniques using monoclonal antibodies directed against PLK1 and Ki-67. The immunoreactivity of 22 melanomas which developed metastases within 5 years of follow-up was compared with a group of 14 non metastasized melanomas. Two independent investigators evaluated stained sections. Differences of PLK1 and Ki-67 indices between melanomas with and without metastases were tested statistically using the Mann-Whitney U-test. RESULTS: Malignant melanomas with metastases expressed PLK1 at markedly elevated levels compared to melanomas without metastases (median, 60.00% vs. 37.98%; p = 0.000053). The difference of the Ki-67 index between both groups was not significant (median, 6.35% vs. 4.53%; p = 0.150473). CONCLUSIONS: Our results suggest that PLK1 expression in thin melanomas is a reliable marker to identify patients at high risk for metastases. PMID- 12135467 TI - Human herpes virus-like particles in pityriasis rosea lesions: an electron microscopy study. AB - BACKGROUND: In a previous study we detected virions with electron microscopy features of human herpes viruses in the supernatant of cocultured mononuclear cells from patients with acute pityriasis rosea. Because of their morphology and of polymerase chain reaction studies, we ascribed them to human herpes virus 7. OBJECTIVE: To find such virions in the lesional skin of pityriasis rosea patients. METHODS: Skin specimens from lesions of 21 patients with acute pityriasis rosea were examined by elecron microscopy. RESULTS: In 15 (71%) patients, human herpes virus particles in various stages of morphogenesis were detected. Mature enveloped virions appeared as typical human herpes virus virions, measuring about 160-200 nm in diameter and containing an electrodense cylindrical core, a capsid, an envelope with typical spikes and a very distinct tegument layer between the capsid and the envelope. They were very similar to those we reported in the supernatant of co-cultured circulating mononuclear cells from patients with pityriasis rosea. CONCLUSION: Our results confirm our previous findings and provides further evidence of a viral etiology for pityriasis rosea. PMID- 12135469 TI - CD30 antigen expression in cutaneous inflammatory infiltrates of scabies: a dynamic immunophenotypic pattern that should be distinguished from lymphomatoid papulosis. AB - BACKGROUND: Expression of CD30 antigen is a distinct marker of lymphocyte activation that was originally described in the Reed-Sternberg cells of Hodgkin's disease. The observation of CD30+ cells has been considered a diagnostic feature of cutaneous CD30 lymphoid proliferations. However, CD30 expression has also been reported in some cutaneous benign inflammatory infiltrates. METHODS: Eleven skin biopsies from patients with scabies were double-blindly and retrospectively analysed. A panel of histopathological parameters and immunophenotypic expression of CD4, CD8, CD30 and S-100 antigens was studied. CD30 and S-100 antigens expression were related to clinical features. RESULTS: Large CD30+ cells were demonstrated in eight (8/11) biopsies, corresponding to patients with long standing lesions (3 months or longer). However, no expression of the CD30 antigen was observed in all biopsy specimens (3/11) corresponding to early lesions (2 months or less). The presence of S-100 positive cells in the papillary dermis was an almost constant feature. CONCLUSIONS: CD30+ large cells seem to be a common feature in long-standing infiltrates of scabies. CD30 expression in scattered cells of a cutaneous lymphoid infiltrate cannot be assessed as a strong diagnostic argument of neoplastic cutaneous CD30+ lymphoid proliferation (lymphomatoid papulosis/cutaneous CD30+ lymphoma). Therefore, the possibility that large atypical CD30+ cells may be also present in several benign inflammatory diseases should be always considered. PMID- 12135468 TI - Malignant peripheral nerve sheath tumor with perineurial differentiation: "malignant perineurioma". AB - BACKGROUND: Although benign tumors derived from the nerve sheath perineurial cell have been described from a variety of anatomic sites and are known to be a component of a number of benign neoplasms, malignant nerve sheath tumors of perineurial origin are exceedingly uncommon. METHODS: We report an unusual case of a 70-year-old male who presented with a rapidly growing mass of the left arm, subsequently shown to be a malignant nerve sheath tumor with perineurial differentiation. A brief microscopic differential diagnosis and review of the literature are discussed. RESULTS: Histologic sections show a partially circumscribed tumor of atypical spindle cells arranged in sweeping fascicles embedded in a myxoid matrix with focal whorling. Nuclear pleomorphism was evident among scattered typical and atypical mitotic figures (mean mitotic index of 7/10 high-power fields). The immunophenotypic profile consisted of only vimentin and epithelial membrane antigen (EMA) positivity, while antibodies to S-100, CD34, smooth muscle actin, and pankeratins were negative. Ultrastructural features included spindle cells with long cytoplasmic processes invested by interrupted basal lamina and pinocytotic vesicles, consistent with perineurial differentiation. CONCLUSIONS: While the histogenic source of the benign perineurioma, the perineurial cell has only rarely been described in conjunction with malignant tumors. All cases to date have shown EMA-positive and S-100 negative atypical spindled cells arranged in fascicles embedded in a myxoid matrix. In addition to immunohistochemistry, ultrastructural examination may be necessary to support the diagnosis. The diagnostic differential includes melanoma, spindle cell squamous cell carcinoma, atypical fibroxanthoma, leiomyosarcoma, and conventional malignant peripheral nerve sheath tumor, most commonly of Schwannian differentiation. Recognizing perineurial differentiation is important since few cases have been reported to date and the biological potential of these neoplasms is not known. PMID- 12135470 TI - Clear cell atypical fibroxanthoma:a clinicopathologic study. AB - INTRODUCTION: The atypical fibroxanthoma (AFX) is considered by most authorities to represent a superficial or minimally invasive variant of malignant fibrous histiocytoma that most often presents as a solitary nodule on the sun-exposed skin of the elderly. Among the rarest variants is the clear cell AFX, a lesion which raises consideration to a differential diagnosis encompassing a variety of neoplastic and non-neoplastic clear cell proliferations. METHODS: We describe three cases of a distinctive cutaneous neoplasm arising in the sun-exposed skin of elderly patients. In all cases, formalin-fixed, paraffin-embedded tissue was available for analysis. The histology in concert with the immunophenotype was held to be diagnostic of the clear cell variant of AFX. RESULTS: All tumors comprised sheets of large cells with foamy cytoplasms and hyperchromatic, polyploid nuclei manifesting frequent and atypical mitoses. The critical cells in our cases expressed CD68 but none of CD3, CD20, CD34, S-100 protein, muscle specific actin, factor XIIIa, Melan-A, carcinoembryonic antigen, or cytokeratin. CONCLUSION: Although typical examples of AFX provoke diagnostic consideration of spindle cell cancers of the skin (most often spindle cell melanoma, spindle cell squamous cell carcinoma, and leiomyosarcoma), the clear cell variant raises other differential diagnostic considerations instead. These include balloon cell melanoma, sebaceous carcinoma, pleomorphic liposarcoma, chordoma, parachordoma, tricholemmal carcinoma and clear cell squamous cell carcinoma. A diagnosis of AFX is one of exclusion; one must employ immunohistochemical markers to rule out the aforementioned differential diagnostic considerations. By reporting the fifth, sixth and seventh cases of clear cell AFX, we hope to alert dermatopathologists to this distinctive and unusual neoplasm, recognition of which is essential to avoid under- or over-diagnosis and inappropriate therapy. PMID- 12135471 TI - Epithelioid malignant schwannoma of the superficial soft tissues versus metastatic amelanotic melanoma. PMID- 12135472 TI - Structure, mechanism and function of prenyltransferases. AB - In this review, we summarize recent progress in studying three main classes of prenyltransferases: (a) isoprenyl pyrophosphate synthases (IPPSs), which catalyze chain elongation of allylic pyrophosphate substrates via consecutive condensation reactions with isopentenyl pyrophosphate (IPP) to generate linear polymers with defined chain lengths; (b) protein prenyltransferases, which catalyze the transfer of an isoprenyl pyrophosphate (e.g. farnesyl pyrophosphate) to a protein or a peptide; (c) prenyltransferases, which catalyze the cyclization of isoprenyl pyrophosphates. The prenyltransferase products are widely distributed in nature and serve a variety of important biological functions. The catalytic mechanism deduced from the 3D structure and other biochemical studies of these prenyltransferases as well as how the protein functions are related to their reaction mechanism and structure are discussed. In the IPPS reaction, we focus on the mechanism that controls product chain length and the reaction kinetics of IPP condensation in the cis-type and trans-type enzymes. For protein prenyltransferases, the structures of Ras farnesyltransferase and Rab geranylgeranyltransferase are used to elucidate the reaction mechanism of this group of enzymes. For the enzymes involved in cyclic terpene biosynthesis, the structures and mechanisms of squalene cyclase, 5-epi-aristolochene synthase, pentalenene synthase, and trichodiene synthase are summarized. PMID- 12135474 TI - Amyloid-fibril formation. Proposed mechanisms and relevance to conformational disease. AB - The phenomenon of the transformation of proteins into amyloid-fibrils is of interest, firstly, because it is closely connected to the so-called conformational diseases, many of which are hitherto incurable, and secondly, because it remains to be explained in physical terms (energetically and structurally). The process leads to fibrous aggregates in the form of extracellular amyloid plaques, neuro-fibrillary tangles and other intracytoplasmic or intranuclear inclusions. In this review, basic principles common to the field of amyloid fibril formation and conformational disease are underlined. Existing models for the mechanism need to be tested by experiment. The kinetic and energetic bases of the process are reviewed. The main controversial issue remains the coexistence of more than one protein conformation. The possible role of oligomeric intermediates, and of domain swapping is also discussed. Mechanisms for cellular defence and novel therapies are considered. PMID- 12135473 TI - Cytochrome c from a thermophilic bacterium has provided insights into the mechanisms of protein maturation, folding, and stability. AB - Cytochrome c is widely distributed in bacterial species, from mesophiles to thermophiles, and is one of the best-characterized redox proteins in terms of biogenesis, folding, structure, function, and evolution. Experimental molecular biology techniques (gene cloning and expression) have become applicable to cytochrome c, enabling its engineering and manipulation. Heterologous expression systems for cytochromes c in bacteria, for use in mutagenesis studies, have been established by extensive investigation of the biological process by which the functional structure is formed. Mutagenesis and structure analyses based on comparative studies using a thermophile Hydrogenobacter thermophilus cytochrome c 552 and its mesophilic counterpart have provided substantial clues to the mechanism underlying protein stability at the amino-acid level. The molecular mechanisms underlying protein maturation, folding, and stability in bacterial cytochromes c are beginning to be understood. PMID- 12135475 TI - The Saccharomyces cerevisiae type 2A protein phosphatase Pph22p is biochemically different from mammalian PP2A. AB - The Saccharomyces cerevisiae type 2A protein phosphatase (PP2A) Pph22p differs from the catalytic subunits of PP2A (PP2Ac) present in mammals, plants and Schizosaccharomyces pombe by a unique N-terminal extension of approximately 70 amino acids. We have overexpressed S. cerevisiae Pph22p and its N-terminal deletion mutant Delta N-Pph22p in the GS115 strain of Pichia pastoris and purified these enzymes to apparent homogeneity. Similar to other heterologous systems used to overexpress PP2Ac, a low yield of an active enzyme was obtained. The recombinant enzymes designed with an 8 x His-tag at their N-terminus were purified by ion-exchange chromatography on DEAE-Sephacel and affinity chromatography on Ni2+-nitrilotriacetic acid agarose. Comparison of biochemical properties of purified Pph22p and Delta N-Pph22p with purified human 8 x His PP2Ac identified similarities and differences between these two enzymes. Both enzymes displayed similar specific activities with 32P-labelled phosphorylase a as substrate. Furthermore, selected inhibitors and metal ions affected their activities to the same extend. In contrast to the mammalian catalytic subunit PP2Ac, but similar to the dimeric form of mammalian PP2A, Pph22p, but not Delta N Pph22p, interacted strongly with protamine. Also with regard to the effects of protamine and polylysine on phosphatase activity Pph22p, but not Delta N-Pph22p, behaved similarly to the PP2Ac-PR65 dimer, indicating a regulatory role for the N terminal extension of Pph22p. The N-terminal extension appears also responsible for interactions with phospholipids. Additionally Pph22p has different redox properties than PP2Ac; in contrast to human PP2Ac it cannot be reactivated by reducing agents. These properties make the S. cerevisiae Pph22p phosphatase a unique enzyme among all type 2A protein phosphatases studied so far. PMID- 12135476 TI - The mechanism of nitrogen monoxide (NO)-mediated iron mobilization from cells. NO intercepts iron before incorporation into ferritin and indirectly mobilizes iron from ferritin in a glutathione-dependent manner. AB - Nitrogen monoxide (NO) is a cytotoxic effector molecule produced by macrophages that results in Fe mobilization from tumour target cells which inhibits DNA synthesis and mitochondrial respiration. It is well known that NO has a high affinity for Fe, and we showed that NO-mediated Fe mobilization is markedly potentiated by glutathione (GSH) generated by the hexose monophosphate shunt [Watts, R.N. & Richardson, D.R. (2001) J. Biol. Chem. 276, 4724-4732]. We hypothesized that GSH completes the coordination shell of an NO[bond]Fe complex that is released from the cell. In this report we have extended our studies to further characterize the mechanism of NO-mediated Fe mobilization. Native PAGE 59Fe-autoradiography shows that NO decreased ferritin-59Fe levels in cells prelabelled with [59Fe]transferrin. In prelabelled cells, ferritin-59Fe levels increased 3.5-fold when cells were reincubated with control media between 30 and 240 min. In contrast, when cells were reincubated with NO, ferritin-59Fe levels decreased 10-fold compared with control cells after a 240-min reincubation. However, NO could not remove Fe from ferritin in cell lysates. Our data suggest that NO intercepts 59Fe on route to ferritin, and indirectly facilitates removal of 59Fe from the protein. Studies using the GSH-depleting agent, L-buthionine (S,R)-sulphoximine, indicated that the reduction in ferritin-59Fe levels via NO was GSH-dependent. Competition experiments with NO and permeable chelators demonstrated that both bind a similar Fe pool. We suggest that NO requires cellular metabolism in order to effect Fe mobilization and this does not occur via passive diffusion down a concentration gradient. Based on our results, we propose a model of glucose-dependent NO-mediated Fe mobilization. PMID- 12135477 TI - A novel DNA repair enzyme containing RNA recognition, G-patch and specific splicing factor 45-like motifs in the protozoan parasite Toxoplasma gondii. AB - We report the cloning and functional characterization of the full-length cDNA and gene encoding a Toxoplasma gondii DNA repair enzyme designated TgDRE. The gene is composed of three exons separated by two introns of 780 and 630 bp, and encodes a protein with a predicted molecular mass of 49.6 kDa. The native TgDRE protein, with a molecular mass of 60 kDa, is only detected in the virulent tachyzoite stage of T. gondii. However, the transcript is present in both asexual parasite stages, virulent tachyzoite and avirulent encysted bradyzoite. When an Escherichia coli mutant lacking ruvC endonuclease and recG helicase was transformed with TgDRE cDNA, a significant increase in resistance to DNA-damaging agents, such as UV light and mitomycin C, was observed. Moreover, database searches revealed that TgDRE orthologues were present in the genome sequences of the related apicomplexa parasites Plasmodium falciparum and Plasmodium yoelii, as well as in those of Arabidopsis thaliana, Drosophila melanogaster, Caenorhabditis elegans and Homo sapiens. This novel family of proteins is characterized by the presence of human splicing factor SF45-like, RNA recognition (RRM) and glycine rich (G-patch) motifs. The presence of these motifs suggests that T. gondii TgDRE might also be involved in other biological functions such as RNA metabolism in addition to DNA-repair. PMID- 12135478 TI - Ruk is ubiquitinated but not degraded by the proteasome. AB - The regulator of ubiquitous kinase (Ruk) protein, also known as CIN85 or SETA, is an adaptor-type protein belonging to the CD2AP/CMS family. It was found in complexes with many signaling proteins, including phosphoinositol (PtdIns) 3 kinase (EC 2.7.1.137), Cbl, GRB2, p130Cas and Crk. Functional analysis of these interactions, implicated Ruk in the regulation of apoptosis, receptor endocytosis and cytoskeletal rearrangements. We have recently demonstrated that overexpression of Ruk induces apoptotic death in neurons, which could be reversed by activated forms of PtdIns 3-kinase and PKB/Akt. Furthermore, Ruk was shown to be a negative regulator of PtdIns 3-kinase activity through binding to its P85 regulatory subunit [Gout, I., Middleton, G., Adu, J., Ninkina, N. N., Drobot, L. B., Filonenko, V., Matsuka, G., Davies, A.M., Waterfield, M. & Buchman, V. L. (2000) Embo J.19, 4015-4025]. Here, we report for the first time, that all three isoforms of Ruk (L, M and S) are ubiquitinated. Specific interaction between the E3 ubiquitin ligase Cbl and all three Ruk isoforms was demonstrated by coexpression studies in Hek293 cells. The interaction of Ruk M and S isoforms with Cbl was found to be mediated via heterodimerization with Ruk L. The use of proteosomal and lysosomal inhibitors clearly indicated that ubiquitination of Ruk L does not lead to its degradation. Based on this study, we propose a possible mechanism for the regulation of Ruk function by ubiquitination. PMID- 12135479 TI - Kinetic and biochemical analyses on the reaction mechanism of a bacterial ATP citrate lyase. AB - The prokaryotic ATP-citrate lyase is considered to be a key enzyme of the carbon dioxide-fixing reductive tricarboxylic acid (RTCA) cycle. Kinetic examination of the ATP-citrate lyase from the green sulfur bacterium Chlorobium limicola (Cl ACL), an alpha(4)beta(4) heteromeric enzyme, revealed that the enzyme displayed typical Michaelis-Menten kinetics toward ATP with an apparent K(m) value of 0.21 +/- 0.04 mm. However, strong negative cooperativity was observed with respect to citrate binding, with a Hill coefficient (n(H)) of 0.45. Although the dissociation constant of the first citrate molecule was 0.057 +/- 0.008 mm, binding of the first citrate molecule to the enzyme drastically decreased the affinity of the enzyme for the second molecule by a factor of 23. ADP was a competitive inhibitor of ATP with a K(i) value of 0.037 +/- 0.006 mm. Together with previous findings that the enzyme catalyzed the reaction only in the direction of citrate cleavage, these kinetic features indicated that Cl-ACL can regulate both the direction and carbon flux of the RTCA cycle in C. limicola. Furthermore, in order to gain insight on the reaction mechanism, we performed biochemical analyses of Cl-ACL. His273 of the alpha subunit was indicated to be the phosphorylated residue in the catalytic center, as both catalytic activity and phosphorylation of the enzyme by ATP were abolished in an H273A mutant enzyme. We found that phosphorylation of the subunit was reversible. Nucleotide preference for activity was in good accordance with the preference for phosphorylation of the enzyme. Although residues interacting with nucleotides in the succinyl-CoA synthetase from Escherichia coli were conserved in AclB, AclA alone could be phoshorylated with the same nucleotide specificity observed in the holoenzyme. However, AclB was necessary for enzyme activity and contributed to enhance phosphorylation and stabilization of AclA. PMID- 12135480 TI - Recombinant human glucose-6-phosphate dehydrogenase. Evidence for a rapid equilibrium random-order mechanism. AB - Cloning and over-expression of human glucose 6-phosphate dehydrogenase (Glc6P dehydrogenase) has for the first time allowed a detailed kinetic study of a preparation that is genetically homogeneous and in which all the protein molecules are of identical age. The steady-state kinetics of the recombinant enzyme, studied by fluorimetric initial-rate measurements, gave converging linear Lineweaver-Burk plots as expected for a ternary-complex mechanism. Patterns of product and dead-end inhibition indicated that the enzyme can bind NADP+ and Glc6P separately to form binary complexes, suggesting a random-order mechanism. The Kd value for the binding of NADP+ measured by titration of protein fluorescence is 8.0 microm, close to the value of 6.8 microm calculated from the kinetic data on the assumption of a rapid-equilibrium random-order mechanism. Strong evidence for this mechanism and against either of the compulsory-order possibilities is provided by repeating the kinetic analysis with each of the natural substrates replaced in turn by structural analogues. A full kinetic analysis was carried out with deaminoNADP+ and with deoxyglucose 6-phosphate as the alternative substrates. In each case the calculated dissociation constant upon switching a substrate in a random-order mechanism (e.g. that for NADP+ upon changing the sugar phosphate) was indeed constant within experimental error as expected. The calculated rate constants for binding of the leading substrate in a compulsory-order mechanism, however, did not remain constant when the putative second substrate was changed. Previous workers, using enzyme from pooled blood, have variously proposed either compulsory-order or random-order mechanisms. Our study appears to provide unambiguous evidence for the latter pattern of substrate binding. PMID- 12135481 TI - Fluorescent analogs of UDP-glucose and their use in characterizing substrate binding by toxin A from Clostridium difficile. AB - Uridine-5'-diphospho-1-alpha-d-glucose (UDP-Glc) is a common substrate used by glucosyltransferases, including certain bacterial toxins such as Toxins A and B from Clostridium difficile. Fluorescent analogs of UDP-Glc have been prepared for use in our studies of the clostridial toxins. These compounds are related to the methylanthraniloyl-ATP compounds commonly used to probe the chemistry of ATP dependent enzymes. The reaction of excess methylisatoic anhydride with UDP-Glc in aqueous solution yields primarily the 2' and 3' isomers of methylanthraniloyl-UDP Glc (MUG). As the 2' and 3' isomers readily interconvert, this isomeric mixture was copurified by HPLC away from the other isomeric products, and was characterized by a combination of NMR, fluorescence and mass spectrometric methods. TcdA binds MUG competitively with respect to UDP-Glc with an affinity of 15 +/- 2 microm in the absence of Mg2+. There is currently no evidence that the fluorescent substrate analog is turned over by the toxin in either glucosyltransferase or glucosylhydrolase reactions. Using a competition assay, the affinity of UDP-Glc was determined to be 45+/-10 microm in the absence of Mg2+. The binding of UDP-Glc and Mg2+ are highly coupled with Mg2+ affinities in the range of 90-600 microm, depending on the experimental conditions. These results imply that one of the significant roles of the metal ion might be to stabilize the enzyme-substrate complex prior to initiation of the transferase chemistry. PMID- 12135482 TI - Structural and biochemical characterization of calhepatin, an S100-like calcium binding protein from the liver of lungfish (Lepidosiren paradoxa). AB - We report the biochemical characterization of calhepatin, a calcium-binding protein of the S100 family, isolated from lungfish (Lepidosiren paradoxa) liver. The primary structure, determined by Edman degradation and MS/MS, shows that the sequence identities with the other members of the family are lower than those between S100 proteins from different species. Calhepatin is composed of 75 residues and has a molecular mass of 8670 Da. It is smaller than calbindin D(9k) (78 residues), the smallest S100 described so far. Sequence analysis and molecular modelling predict the two EF-hand motifs characteristic of the S100 family. Metal-binding properties were studied by a direct 45Ca2+-binding assay and by fluorescence titration. Calhepatin binds Ca2+ and Cu2+ but not Zn2+. Cu2+ binding does not change the affinity of calhepatin for Ca2+. Calhepatin undergoes a conformational change upon Ca2+ binding as shown by the increase in its intrinsic fluorescence intensity and lambda(max), the decrease in the apo calhepatin hydrodynamic volume, and the Ca2+-dependent binding of the protein to phenyl-Superose. Like most S100 proteins, calhepatin tends to form noncovalently associated dimers. These data suggest that calhepatin is probably involved in Ca2+-signal transduction. PMID- 12135483 TI - Differential regulation of telomerase activity by six telomerase subunits. AB - Telomerase is a specialized reverse transcriptase responsible for synthesizing telomeric DNA at the ends of chromosomes. Six subunits composing the telomerase complex have been cloned: hTR (human telomerase RNA), TEP1 (telomerase-associated protein 1), hTERT (human telomerase reverse transcriptase), hsp90 (heat shock protein 90), p23, and dyskerin. In this study, we investigated the role of each the telomerase subunit on the activity of telomerase. Through down- or upregulation of telomerase, we found that only hTERT expression changed proportionally with the level of telomerase activity. The other components, TEP1, hTR, hsp90, p23, and dyskerin remained at high and unchanged levels throughout modulation. In vivo and in vitro experiments with antisense oligonucleotides against each telomerase component were also performed. Telomerase activity was decreased or abolished by antisense treatment. To correlate clinical sample status, four pairs of normal and malignant tissues from patients with oral cancer were examined. Except for the hTERT subunit, which showed differential expression in normal and cancer tissues, all other components were expressed in both normal and malignant tissues. We conclude that hTERT is a regulatable subunit, whereas the other components are expressed more constantly in cells. Although hTERT has a rate-limiting effect on enzyme activity, the other telomerase subunits (hTR, TEP1, hsp90, p23, dyskerin) participated in full enzyme activity. We hypothesize that once hTERT is expressed, all other telomerase subunits can be assembled to form a highly active holoenzyme. PMID- 12135484 TI - Identification of mammalian-type transglutaminase in Physarum polycephalum. Evidence from the cDNA sequence and involvement of GTP in the regulation of transamidating activity. AB - Transglutaminase (TGase) catalyses the post-translational modification of proteins by transamidation of available glutamine residues. While several TGase genes of fish and arthropods have been cloned and appear to have similar structures to those of mammals, no homologous gene has been found in lower eukaryotes. We have cloned the acellular slime mold Physarum polycephalum TGase cDNA using RT-PCR with degenerated primers, based on the partial amino acid sequence of the purified enzyme. The cDNA contained a 2565-bp ORF encoding a 855 residue polypeptide. By Northern blotting, an mRNA of approximately 2600 bases was detected. In comparison with primary sequences of mammalian TGases, surprisingly, significant similarity was observed including catalytic triad residues (Cys, His, Asn) and a GTP-binding region. The alignment of sequences and a phylogenetic tree also demonstrated that the structure of P. polycephalum TGase is similar to that of TGases of vertebrates. Furthermore, we observed that the purified TGase had GTP-hydrolysing activity and that GTP inhibited its transamidating activity, as in the case of mammalian tissue-type TGase (TGase 2). PMID- 12135485 TI - Regulation of glypican-1, syndecan-1 and syndecan-4 mRNAs expression by follicle stimulating hormone, cAMP increase and calcium influx during rat Sertoli cell development. AB - In seminiferous tubules, Sertoli cells provide structural and nutritional support for the developing germinal cells. Cell- to-cell signaling and cell adhesion require proteoglycans expressed at the cell membrane. A preliminary biochemical and structural approach indicated that cell surface proteoglycans are mostly heparan sulfate proteoglycans (HSPG). Glypican-1, syndecans-1 and -4 were identified using a molecular approach. Their differential regulation was demonstrated in immature rat Sertoli cells. Follicle-stimulating hormone (FSH) is the main regulator of Sertoli cell function. Signal transduction triggered by FSH involves both an increased intracellular cAMP synthesis and a calcium influx. This study demonstrates that FSH, through its second messengers (increase in intracellular cAMP and intracellular calcium), downregulated the glypican-1 mRNA expression in Sertoli cells from 20-day-old rats. On the other hand, syndecan-1 mRNA expression is not modulated by FSH as it would result from the antagonistic effects of increased intracellular cAMP and intracellular calcium levels. Finally, syndecan-4 mRNA expression is not regulated by this pathway. The present study was extended during Sertoli cell development. Indeed, Sertoli cells undergo extensive changes during the postnatal period both in structure and function. These important transformations are critical for the establishment of spermatogenesis and development of the adult pattern of testicular function. Our data indicated that the regulation of HSPG mRNA expression is HSPG-specific and depends on the Sertoli cell developmental stage. PMID- 12135486 TI - S-decyl-glutathione nonspecifically stimulates the ATPase activity of the nucleotide-binding domains of the human multidrug resistance-associated protein, MRP1 (ABCC1). AB - The human multidrug resistance-associated protein(MRP1) is an ATP-dependent efflux pump that transports anionic conjugates, and hydrophobic compounds in a glutathione dependent manner. Similar to the other, well-characterized multidrug transporter P-gp, MRP1 comprises two nucleotide-binding domains (NBDs) in addition to transmembrane domains. However, whereas the NBDs of P-gp have been shown to be functionally equivalent, those of MRP1 differ significantly. The isolated NBDs of MRP1 have been characterized in Escherichia coli as fusions with either the glutathione-S-transferase (GST) or the maltose-binding domain (MBP). The nonfused NBD1 was obtained by cleavage of the fusion protein with thrombin. The GST-fused forms of NBD1 and NBD2 hydrolyzed ATP with an apparent K(m) of 340 microm and a V(max) of 6.0 nmol P(I) x mg-1 x min-1, and a K(m) of 910 microm ATP and a V(max) of 7.5 nmol P(I) x mg-1 x min-1, respectively. Remarkably, S-decyl glutathione, a conjugate specifically transported by MRP1 and MRP2, was able to stimulate the ATPase activities of the isolated NBDs more than 2-fold in a concentration-dependent manner. However,the stimulation of the ATPase activity was found to coincide with the formation of micelles by S-decyl-glutathione. Equivalent stimulation of ATPase activity could be obtained by surfactants with similar critical micelle concentrations. PMID- 12135487 TI - The unorthodox histidine kinases BvgS and EvgS are responsive to the oxidation status of a quinone electron carrier. AB - The purified soluble forms of the histidine kinases BvgS and EvgS of Bordetella pertussis and Escherichia coli, respectively, are shown to be responsive to oxidized ubiquinone-0 (Q-0) in vitro. The oxidized ubiquinone is a strong inhibitor of kinase activity of both enzymes with half maximal inhibition occurring at 11 microm (BvgS) and 4 microm (EvgS). Reduced Q-0 has no effect on the histidine kinases. Kinase activity can reversibly be switched off and on by changing the oxidation status of the quinone. This inhibitory effect is due to a decrease of the kinase activity of BvgS rather than an increase of intrinsic phosphatase activities. Other electron carriers such as menadione (MK-3), NAD or FAD did not have a significant effect on the kinase activities of BvgS and EvgS. Nicotinic acid and sulfate ions, known to inhibit the histidine kinases in vivo, did not affect the purified truncated sensor proteins lacking their periplasmic domains in vitro. Mutations introduced by site-directed mutagenesis into the putative PAS domain of BvgS caused a weak decrease of quinone-dependent inhibition of autophosphorylation. These data suggest that BvgS and EvgS are connected with the oxidation status of the cell via the link to the ubiquinone pool. PMID- 12135488 TI - Intracellular pH homeostasis in the filamentous fungus Aspergillus niger. AB - Intracellular pH homeostasis in the filamentous fungus Aspergillus niger was measured in real time by 31P NMR during perfusion in the NMR tube of fungal biomass immobilized in Ca2+-alginate beads. The fungus maintained constant cytoplasmic pH (pH(cyt)) and vacuolar pH (pH(vac)) values of 7.6 and 6.2, respectively, when the extracellular pH (pH(ex)) was varied between 1.5 and 7.0 in the presence of citrate. Intracellular metabolism did not collapse until a Delta pH over the cytoplasmic membrane of 6.6-6.7 was reached (pH(ex) 0.7-0.8). Maintenance of these large pH differences was possible without increased respiration compared to pH(ex) 5.8. Perfusion in the presence of various hexoses and pentoses (pH(ex) 5.8) revealed that the magnitude of Delta pH values over the cytoplasmic and vacuolar membrane could be linked to the carbon catabolite repressing properties of the carbon source. Also, larger Delta pH values coincided with a higher degree of respiration and increased accumulation of polyphosphate. Addition of protonophore (carbonyl cyanide m chlorophenylhydrazone, CCCP) to the perfusion buffer led to decreased ATP levels, increased respiration and a partial (1 microm CCCP), transient (2 microm CCCP) or permanent (10 microm CCCP) collapse of the vacuolar membrane Delta pH. Nonlethal levels of the metabolic inhibitor azide (N3-, 0.1 mm) caused a transient decrease in pH(cyt) that was closely paralleled by a transient vacuolar acidification. Vacuolar H+ influx in response to cytoplasmic acidification, also observed during extreme medium acidification, indicates a role in pH homeostasis for this organelle. Finally, 31P NMR spectra of citric acid producing A. niger mycelium showed that despite a combination of low pH(ex) (1.8) and a high acid-secreting capacity, pH(cyt) and pH(vac) values were still well maintained (pH 7.5 and 6.4, respectively). PMID- 12135490 TI - SF2/ASF protein inhibits camptothecin-induced DNA cleavage by human topoisomerase I. AB - A splicing factor SF2/ASF is a natural substrate for the kinase activity of human topoisomerase I. This study demonstrates that SF2/ASF inhibits DNA cleavage by human topoisomerase I induced by the anti-cancer agent camptothecin. The inhibition is independent of the phosphorylation status of SF2/ASF. We show that the inhibition did not result from binding of SF2/ASF to DNA that would hinder interactions between topoisomerase I and DNA. Neither it was a consequence of a loss of sensitivity of the enzyme to camptothecin. We provide evidence pointing to reduced formation of the cleavable complex in the presence of SF2/ASF as a primary reason for the inhibition. This effect of SF2/ASF is reflected by inhibition of DNA relaxation catalysed by topoisomerase I. PMID- 12135489 TI - Functional analysis of the rat bile salt export pump gene promoter. AB - The 5' flanking region of the bile salt export pump (Bsep) gene was systematically analysed to provide the basis for understanding the mechanisms which regulate Bsep transcription. In addition substrates and drugs were investigated for their ability to alter Bsep promoter activity. Bsep promoter function was restricted to hepatocyte derived HepG2 cells. The 5' deletional analysis revealed a biphasic shape of reporter gene activities, indicating a suppressive element between nucleotides -800 and -512. Two consensus sites for the farnesoid X receptor (FXR) were located at nucleotides -473 and -64. The latter was characterized as functionally active in bile acid-mediated feed-back regulation of Bsep transcription. Bsep promoter activity was reduced by rifampin and beta-estradiol. The anti-estrogen tamoxifen stimulated promoter activity. Dexamethasone, hydrocortisone and phenobarbital had no effect on Bsep promoter activity. In conclusion, the data suggest that transcriptional regulation of the Bsep gene can be modulated by a number of endogenous compounds and xenobiotics. FXR was a major regulatory factor, mediating bile acid feed-back stimulation of Bsep transcription. PMID- 12135491 TI - Rum1, an inhibitor of cyclin-dependent kinase in fission yeast, is negatively regulated by mitogen-activated protein kinase-mediated phosphorylation at Ser and Thr residues. AB - The p25(rum1) is an inhibitor of Cdc2 kinase expressed in fission yeast and plays an important role in cell-cycle control. As its amino-acid sequence suggests that p25(rum1) has putative phosphorylation sites for mitogen-activated protein kinase (MAPK), we investigated the ability of MAPK to phosphorylate p25(rum1). Direct in vitro kinase assay using GST-fusion proteins of wild-type as well as various mutants of p25(rum1) demonstrated that MAPK phosphorylates the N-terminal portion of p25(rum1) and residues Thr13 and Ser19 are major phosphorylation sites for MAPK. In addition, phosphorylation of p25(rum1) by MAPK revealed markedly reduced Cdc2 kinase inhibitor ability of the protein. Together with the fact that replacement of both Thr13 and Ser19 with Glu, which mimics the phosphorylated state of these residues, also significantly reduces the activity of p25(rum1) as a Cdc2 inhibitor, it was suggested that the phosphorylation of Thr13 and Ser19 negatively regulates the function of p25(rum1). Further evidence indicates that phosphorylation of Thr13 and Ser19 may retain a negative effect on the function of p25(rum1) even in vivo. Therefore, MAPK may regulate the function of p25(rum1) via phosphorylation of its Thr and Ser residues and thus participate in cell cycle control in fission yeast. PMID- 12135492 TI - Characterization of four substrates emphasizes kinetic similarity between insect and human C-domain angiotensin-converting enzyme. AB - Angiotensin converting enzyme (ACE) was already discovered in insects in 1994, but its physiological role is still enigmatic. We have addressed this problem by purifying four new ACE substrates from the ovaries of the grey fleshfly, Neobellieria bullata. Their primary structures were identified as NKLKPSQWISLSD (Neb-ODAIF-1(1-13)), NKLKPSQWI (Neb-ODAIF-1(1-9)), SLKPSNWLTPSE (Neb-ODAIF-2) and LEQIYHL. Database analysis showed significant homology with amino acid sequence stretches as present in the N-terminal part of several fly yolk proteins. An antiserum raised against Neb-ODAIF-1(1-9) immunostained one out of three yolk protein bands of SDS/PAGE-separated fly haemolymph and egg homogenate, thus confirming that these peptides originate from a yolk protein gene product. Kinetic analysis of these peptides and of the peptides Neb-ODAIF and Neb-ODAIF 1(1-7) with insect ACE and human ACE show both similar and unique properties for insect ACE as compared with human C-domain ACE. PMID- 12135493 TI - Engineering and mechanistic studies of the Arabidopsis FAE1 beta-ketoacyl-CoA synthase, FAE1 KCS. AB - The Arabidopsis FAE1 beta-ketoacyl-CoA synthase (FAE1 KCS) catalyzes the condensation of malonyl-CoA with long-chain acyl-CoAs. Sequence analysis of FAE1 KCS predicted that this condensing enzyme is anchored to a membrane by two adjacent N-terminal membrane-spanning domains. In order to characterize the FAE1 KCS and analyze its mechanism, FAE1 KCS and its mutants were engineered with a His6-tag at their N-terminus, and expressed in Saccharomyces cerevisiae. The membrane-bound enzyme was then solubilized and purified to near homogeneity on a metal affinity column. Wild-type recombinant FAE1 KCS was active with several acyl-CoA substrates, with highest activity towards saturated and monounsaturated C16 and C18. In the absence of an acyl-CoA substrate, FAE1 KCS was unable to carry out decarboxylation of [3-(14)C]malonyl-CoA, indicating that it requires binding of the acyl-CoA for decarboxylation activity. Site-directed mutagenesis was carried out on the FAE1 KCS to assess if this condensing enzyme was mechanistically related to the well characterized soluble condensing enzymes of fatty acid and flavonoid syntheses. A C223A mutant enzyme lacking the acylation site was unable to carry out decarboxylation of malonyl-CoA even when 18:1-CoA was present. Mutational analyses of the conserved Asn424 and His391 residues indicated the importance of these residues for FAE1-KCS activity. The results presented here provide the initial analysis of the reaction mechanism for a membrane-bound condensing enzyme from any source and provide evidence for a mechanism similar to the soluble condensing enzymes. PMID- 12135494 TI - Substrate selectivity and sensitivity to inhibition by FK506 and cyclosporin A of calcineurin heterodimers composed of the alpha or beta catalytic subunit. AB - The calcineurin (CaN) alpha and beta catalytic subunit isoforms are coexpressed within almost all cell types. The enzymatic properties of CaN heterodimers comprised of the regulatory B subunit (CnB) with either the alpha or beta catalytic subunit were compared using in vitro phosphatase assays. CaN containing the alpha isoform (CnA alpha) has lower K(m) and higher V(max) values than CaN containing the beta isoform (CnA beta) toward the PO4-RII, PO4-DARPP-32(20-38) peptides, and p-nitrophenylphosphate (pNPP). CaN heterodimers containing the alpha or beta catalytic subunit isoform displayed identical calmodulin dissociation rates. Similar inhibition curves for each CaN heterodimer were obtained with the CaN autoinhibitory peptide (CaP) and cyclophilin A/cyclosporin A (CyPA/CsA) using each peptide substrate at K(m) concentrations, except for a five- to ninefold higher IC50 value measured for CaN containing the beta isoform with p-nitrophenylphosphate as substrate. No difference in stimulation of phosphatase activity toward p-nitrophenylphosphate by FKBP12/FK506 was observed. At low concentrations of FKBP12/FK506, CaN containing the alpha isoform is more sensitive to inhibition than CaN containing the beta isoform using the phosphopeptide substrates. Higher concentrations of FKBP12/FK506 are required for maximal inhibition of beta CaN using PO4-DARPP-32(20-38) as substrate. The functional differences conferred upon CaN by the alpha or beta catalytic subunit isoforms suggest that the alpha:beta and CaN:substrate ratios may determine the levels of CaN phosphatase activity toward specific substrates within tissues and specific cell types. These findings also indicate that the alpha and beta catalytic subunit isoforms give rise to substrate-dependent differences in sensitivity toward FKBP12/FK506. PMID- 12135495 TI - Chemical structures and immunolocalization of glycosphingolipids isolated from Diphyllobothrium hottai adult worms and plerocercoids. AB - Glycosphingolipids (GSLs) were purified from adults and plerocercoids of the tapeworm Diphyllobothrium hottai, and their chemical structures were determined. Total lipid fractions prepared from chloroform/methanol extracts of whole tissues were fractionated successively on ion-exchange chromatography, silicic acid column chromatography, and preparative TLC. The purified GSLs were characterized by methylation analysis, TLC-immunostaining, liquid secondary ion MS, MALDI-TOF MS, and 1H-NMR. Ten GSLs were isolated from adult worms and four from plerocercoids, comprising mono-, di-, tri-, tetra-, and pentasaccharides. The GSL Gal beta 1-4(Fuc alpha 1-3)Glc beta 1-3Gal beta 1-Cer was found in adult worms but not in plerocercoids, whereas Ga lbeta 1-4 (Fuc alpha 1-3)Glc beta 1-3(Gal beta 1-6)Gal beta 1-Cer was found in both adult worms and plerocercoids. We previously found a similar series of GSLs in plerocercoids of the cestode Spirometra erinaceieuropaei, and termed them 'spirometosides'[Kawakami, Y. et al. (1996) Eur J. Biochem. 239, 905-911]. The core structure of spirometosides, Gal beta 1-4Glc beta 1-3 Gal beta 1-Cer, may have taxonomic significance, being characteristic of pseudophyllidean tapeworms. In the present study, GSL compositions were significantly different between adults and plerocercoids, and growth-dependent changes in composition were documented. We found a novel dihexosylceramide, Glc beta 1-3Gal beta 1-Cer, which is a possible precursor for spirometosides. Immunohistochemical examination showed that spirometoside GSLs are highly enriched in the inner surface of bothria, the major point of contact between the adult worm and the host's intestine. Our findings indicate that spirometosides are involved in host-parasite interaction. PMID- 12135496 TI - Interaction of the GTS1 gene product with glyceraldehyde- 3-phosphate dehydrogenase 1 required for the maintenance of the metabolic oscillations of the yeast Saccharomyces cerevisiae. AB - We previously reported that GTS1 is involved in regulating ultradian oscillations of the glycolytic pathway induced by cyanide in cell suspensions as well as oscillations of energy metabolism in aerobic continuous cultures. Here, we screened a yeast cDNA library for proteins that bind to Gts1p using the yeast two hybrid system and cloned multiple TDH cDNAs encoding the glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (GAPDH). We found that the zinc-finger and dimerization sites of Gts1p were required for full ability to bind GAPDH, and Gts1ps mutated at these sites lost the ability to regulate both aerobic and unaerobic ultradian oscillations of energy metabolism. Of the three TDH genes, only TDH1 fluctuated at the mRNA level in continuous culture and its deletion resulted in the disappearance of the oscillation without any affect on growth rate. This loss of biological rhythms in the TDH1-deleted mutant was rescued by the expression of TDH1 but not of TDH2 or TDH3 under the control of the TDH1 promoter. Thus, we hypothesized that Gts1p plays a role in the regulation of metabolic oscillation by interacting with the TDH1 product, GAPDH1, in yeast. PMID- 12135497 TI - Fusion of farnesyldiphosphate synthase and epi-aristolochene synthase, a sesquiterpene cyclase involved in capsidiol biosynthesis in Nicotiana tabacum. AB - A clone encoding farnesyl diphosphate synthase (FPPS) was obtained by PCR from a cDNA library made from young leaves of Artemisia annua. A cDNA clone encoding the tobacco epi-aristolochene synthase (eAS) was kindly supplied by J. Chappell (University of Kentucky, Lexington, KY, USA). Two fusions were constructed, i.e. FPPS/eAS and eAS/FPPS. The stop codon of the N-terminal enzyme was removed and replaced by a short peptide (Gly-Ser-Gly) to introduce a linker between the two ORFs. These two fusions and the two single cDNA clones were separately introduced into a bacterial expression vector (pET32). Escherichia coli was transformed with the expression vectors and enzymatically active soluble proteins were obtained after induction with isopropyl thio-beta-d-thiogalactoside. The recombinant enzymes were purified using immobilized metal affinity chromatography on Co2+ columns. The fusion enzymes produced epi-aristolochene from isopentenyl diphosphate through a coupled reaction. The Km values of FPPS and eAS for isopentenyl diphosphate and farnesyl diphosphate, respectively, were essentially the same for the single and fused enzymes. The bifunctional enzymes showed a more efficient conversion of isopentenyl diphosphate to epi-aristolochene than the corresponding amount of single enzymes. PMID- 12135498 TI - A critical motif for oligomerization and chaperone activity of bacterial alpha heat shock proteins. AB - Oligomerization into multimeric complexes is a prerequisite for the chaperone function of almost all alpha-crystallin type heat shock proteins (alpha-Hsp), but the molecular details of complex assembly are poorly understood. The alpha-Hsp proteins from Bradyrhizobium japonicum are suitable bacterial models for structure-function studies of these ubiquitous stress proteins. They fall into two distinct classes, A and B, display chaperone activity in vitro and form oligomers of approximately 24 subunits. We constructed 19 derivatives containing truncations or point mutations within the N- and C-terminal regions and analyzed them by gel filtration, citrate synthase assay and coaffinity purification. Truncation of more than the initial few amino acids of the N-terminal region led to the formation of distinct dimeric to octameric structures devoid of chaperone activity. In the C-terminal extension, integrity of an isoleucine-X-isoleucine (I X-I) motif was imperative for alpha-Hsp functionality. This I-X-I motif is one of the characteristic consensus motifs of the alpha-Hsp family, and here we provide experimental evidence of its structural and functional importance. alpha-Hsp proteins lacking the C-terminal extension were inactive, but still able to form dimers. Here, we demonstrate that the central alpha-crystallin domain alone is not sufficient for dimerization. Additional residues at the end of the N-terminal region were required for the assembly of two subunits. PMID- 12135499 TI - Function and cellular localization of farnesoic acid O-methyltransferase (FAMeT) in the shrimp, Metapenaeus ensis. AB - The isoprenoid methyl farnesoate (MF) has been implicated in the regulation of crustacean development and reproduction in conjunction with eyestalk molt inhibiting hormones and ecdysteroids. Farnesoic acid O-methyltransferase (FAMeT) catalyzes the methylation of farnesoic acid (FA) to produce MF in the terminal step of MF synthesis. We have previously cloned and characterized the shrimp FAMeT. In the present study, recombinant FAMeT (rFAMeT) was produced for bioassay and antiserum generation. FAMeT is widely distributed in shrimp tissues with the highest concentration observed in the ventral nerve cord. Interestingly, an additional larger protein in the eyestalk also showed immunoreactivity to anti FAMeT serum. FAMeT was localized in the neurosecretory cells of the X-organ-sinus gland complex of the eyestalk. As shown by RT-PCR, FAMeT mRNA is constitutively expressed throughout the molt cycle in the eyestalk and the ventral nerve cord. To show that our cloned gene product had FAMeT activity, we demonstrated that expressed rFAMeT gene product catalyzed the conversion of FA to MF in a radiochemical assay. The ubiquitous distribution of FAMeT suggests that this enzyme is involved in physiological processes in addition to gametogenesis, oocyte maturation and development and metamorphosis of the shrimp. We hypothesize that FAMeT directly or indirectly (through MF) modulates the reproduction and growth of crustaceans by interacting with the eyestalk neuropeptides as a consequence of its presence in the neurosecretory cells of the X-organ-sinus gland. PMID- 12135500 TI - Antiproliferative proteins of the BTG/Tob family are degraded by the ubiquitin proteasome system. AB - BTG/Tob family proteins, which are characterized by similarities in their N terminal BTG/Tob homology domains, control cell growth negatively. Among the BTG/Tob family members, BTG2/TIS21/PC3 proteins have been reported to have short lives and to be degraded by the proteasome. However, the mechanisms regulating the stabilities of other BTG/Tob family proteins have not yet been clarified. Here, we report that BTG1, Tob, and Tob2 proteins, as well as BTG2 protein, are degraded by the ubiquitin-proteasome system; the degradation of Tob protein in HeLa cells and the degradation of BTG1, BTG2, Tob and Tob2 proteins transiently expressed in HEK293 cells were inhibited by treatments with proteasome-specific inhibitors. Co-expression of BTG1, BTG2, Tob, or Tob2 protein with ubiquitin in HEK293 cells revealed specific multiubiquitination of each of the four proteins. Although the full-length and N-terminal truncated forms of BTG1, BTG2, Tob, and Tob2 proteins were unstable, the respective C-terminal truncated forms were found to be almost stable, suggesting that the C-terminal regions control the stabilities of BTG1, BTG2, Tob, and Tob2 proteins. In addition, it was found that the respective C-terminal regions confer instability on green fluorescent protein, a normally stable protein. Thus, it can be concluded that the C-terminal regions are necessary and sufficient to control the stabilities of BTG1, BTG2, Tob, and Tob2 proteins. PMID- 12135501 TI - Glycolic Acid peels compared to microdermabrasion. PMID- 12135502 TI - Microprocessor-controlled local anesthesia vs. the conventional syringe technique in hair transplantation. PMID- 12135503 TI - Dermatologic surgery training in residency. AB - BACKGROUND: Dermatologic surgery is an integral part of dermatology residency training. OBJECTIVE: To characterize the current level of surgical training in dermatology residency programs. METHODS: A survey was mailed to the 112 dermatology residency programs in North America and Puerto Rico. RESULTS: A total of 71 residency programs (63%) responded. All programs offer training in surgical excisions, 97% of programs train residents in at least one cosmetic procedure, 92% of programs offer Mohs micrographic surgery, and 90% of programs train their residents in laser surgery. CONCLUSION: Surgical training in dermatology residency varies widely, with a trend toward more cosmetic procedures when compared with prior studies. PMID- 12135504 TI - Treatment of atrophic facial acne scars with a dual-mode Er:YAG laser. AB - BACKGROUND: Scar revision with CO2 and Er:YAG lasers has become popular in recent years. Reports on the newest (modulated, dual-mode) Er:YAG systems have been limited mostly to the treatment of photodamaged skin and rhytides. OBJECTIVE: To prospectively evaluate the efficacy and safety of a dual-mode 2940 nm Er:YAG laser for atrophic scar revision. METHODS: Twenty-five consecutive patients with moderate to severe atrophic facial acne scars received treatment with a dual-mode Er:YAG laser. Clinical assessments using a standard grading scale and photographic documentation were performed at 1, 3, 6, and 12 months postoperatively. Postoperative recovery was monitored and the rate of side effects and complications recorded. RESULTS: Average clinical grading scores reflected good to excellent response of atrophic scars to the dual-mode Er:YAG laser system. Side effects and complications were limited to transient hyperpigmentation and acne flare-ups. No hypopigmentation or scarring was seen. Prolonged erythema (longer than 1 month) was observed in 1 patient (4%). CONCLUSION: Dual-mode Er:YAG laser skin resurfacing is a safe and effective modality for the treatment of atrophic facial scarring. PMID- 12135505 TI - Clinical application of lasers in Asians. AB - BACKGROUND: Laser surgery for Asians differs from that for Caucasians in several important respects. In Asians, some conditions such as nevus of Ota are frequently seen and certain adverse reactions, especially postinflammatory hyperpigmentation, tend to be more common. OBJECTIVE: This article reviews the use of different types of lasers and intense pulsed light (IPL) sources for the treatment of Asian patients. METHODS: Various cutaneous conditions amenable to laser treatment, including lentigines, nevus of Ota, acquired bilateral nevus of Ota-like macules, port-wine stains, and acne scarring, are discussed. Strategies for the management of postinflammatory hyperpigmentation are offered. RESULTS: Appropriate selection and careful planning of the treatment can lead to excellent clinical outcome. CONCLUSION: Lasers and intense pulsed light sources are important tools for the treatment of a wide range of cutaneous conditions in Asians. PMID- 12135506 TI - Treatment of leg telangiectases with a 532 nm KTP laser in multipulse mode. AB - BACKGROUND: The multiple mode emission emphasizes the efficacy of the KTP laser. OBJECTIVE: To evaluate the efficacy of a 532 nm KTP laser emitting in multipulse mode for the treatment of superficial 0.5-1 mm leg telangiectases. METHODS: A 532 nm KTP laser was used in a nonuniform pulse sequence or multipulse mode emission (three stacked pulses of 100 msec, 30 msec, 30 msec, and a delay between pulses of 250 msec), a fluence of 60 J/cm2, and a 0.75 mm collimated spot. No cooling was used. Fourteen female patients (average age 46 years, range 27-57 years), phototypes I-IV were examined with Doppler ultrasound to ensure their big veins were competent. A topography of the telangiectatic network was reported on a tracing plastic frame before each session and 6 weeks after the last one. These frames were digitized and the number of vessels (before and 6 weeks after each session) was determined using imaging software. Side effects, pain, and patient satisfaction were noted. RESULTS: Moderate pain, immediate erythema and edema, sometimes light scabbing, temporary hypopigmentation rarely, and no matting were observed. After one treatment, vessel clearing was 53% (P <.001). It increased to 78% (P <.001) 6 weeks after two treatments, to 85% (P <.05) 6 weeks after three treatments, and to 93% (NS) 6 weeks after four treatments. CONCLUSION: This nonuniform pulse sequence or multipulse mode emission emphasizes the efficacy of the KTP laser in this study. It provides a safe and effective treatment that achieved an important reduction of red leg veins telangiectases from 0.5 to 1 mm in diameter, with very few side effects. PMID- 12135507 TI - Leukotrichia developed following application of intense pulsed light for hair removal. AB - BACKGROUND: Lasers and light sources are now used worldwide for permanent or prolonged hair removal. Patients now prefer lasers and light sources for hair removal because of their noninvasiveness and fewer reported side effects. OBJECTIVE: To study and report on leukotrichia that developed following application of intense pulsed light (IPL). METHODS: From February 9, 2001 to February 14, 2002 a total of 821 patients were treated for unwanted hair. The system used was a noncoherent IPL system, with a 650 nm flashlamp filter; the parameters used varied with different Fitzpatrick skin types. The patients were treated monthly, with the rate of hair loss, measured by hair counts, and possible side effects recorded. RESULTS: Twenty-nine of 821 patients treated developed leukotrichia. Thirteen patients had no white or gray hairs before IPL therapy; the remaining 16 patients, who had few white hairs before treatment reported accelerated development of new white hairs starting after the first or second IPL therapy. Restoration of hair color occurred in 9 patients and the remaining 20 patients had no improvement or worsening of the condition within the next 2-6 months. CONCLUSION: Temporary or permanent leukotrichia may develop following IPL and laser hair removal therapy. This finding may be explained by the difference in the thermal relaxation times of melanocytes and germinative cells. The light absorbed and the heat produced by melanin may be sufficient enough to destroy or impair the function of melanocytes but insufficient to damage the hair follicle cells. PMID- 12135508 TI - CO(2) laser therapy of vulval lymphangiectasia and lymphangioma circumscriptum. AB - BACKGROUND: Lymphangiectasia is a disorder of superficial lymphatics resulting from obstruction of previously normal deep lymphatics, while lymphangioma circumscriptum describes a deep dermal and subcutaneous lymphatic malformation with secondary superficial ectatic changes. Previous case reports have suggested the effectiveness of CO2 laser treatment. OBJECTIVE: To assess CO2 laser therapy for vulval involvement by these lymphatic disorders. METHODS: A retrospective case review at St. John's Institute of Dermatology from 1996 to 1999 identified three women with vulval lymphangiectasia and two with vulval lymphangioma circumscriptum treated with the CO2 laser. RESULTS: Patient tolerance of the procedure was good. Healing was complete within 1 month and occurred without change in skin texture. All patients reported considerable improvement in symptoms (mean follow-up 22 months, median 24 months, range 10-33 months). Focal recurrence and an area of localized persistence were noted in the two patients with lymphangioma circumscriptum. CONCLUSION: CO2 laser therapy of vulval lymphangiectasia and lymphangioma circumscription is effective and well tolerated; the latter may possibly be more resistant to treatment than the former. PMID- 12135509 TI - A surgical approach to ingrown nail: partial matricectomy using CO2 laser. AB - BACKGROUND: Ingrown nail is a deformity characterized by a transverse overcurvature of the lateral nail plate that causes nail fold inflammation. Currently recommended surgical procedures vary considerably. OBJECTIVE: We report our experience with a modified partial matricectomy and nail edge excision with CO2 laser. METHODS: A total of 116 ingrown nail edges in 77 patients were treated with the procedure. RESULTS: The incidence of recurrence was 2 of 76 cases (2.6%). CONCLUSION: This new surgical approach was found to be effective and is presented as an alternative treatment technique. PMID- 12135510 TI - Surgical anatomy of the scalp. AB - BACKGROUND: The goal of complication-free scalp surgery remains elusive. Wide unsightly scars, stretch-atrophied hypesthetic, poorly vascularized tissues and distorted hair patterns are still commonly seen and appear to be largely refractory to remedial surgery. OBJECTIVE: To establish technical guidelines for scalp surgery most compatible with maximum functional/cosmetic benefit and complication-free results. METHODS: More than 3000 scalp operations, done personally, were reviewed. These covered the entire spectrum of plastic and reconstructive surgery and hair restoration procedures including more than 700 clinical surgical investigative procedures with pericranial flaps, subgaleal and subperiosteal scalp reductions, and deep plane fixation procedures. RESULTS: Notably improved scalp surgery outcomes resulted from the application of data derived from study of the surgical anatomy of the scalp. Unsightly scars, distorted hair patterns, hypesthetic poorly vascularized tissues, and distorted hair patterns were largely avoided. CONCLUSION: An understanding of the surgical anatomy of the scalp is a cornerstone on which complication-free scalp surgery is based. PMID- 12135511 TI - 1, 2, 3, 4: four techniques to secure buried knots. AB - BACKGROUND: High tension wounds can be difficult to close. OBJECTIVE: To enumerate methods for securing a surgeon's knot when tying buried sutures under tension. CONCLUSION: The tension tranfer stitch is a swift and elegant method to secure surgeon's knots. PMID- 12135512 TI - The silicone conundrum: a battle of anecdotes. PMID- 12135513 TI - Silicone injections revisited. PMID- 12135514 TI - Thermal damage of the inner vein wall during endovenous laser treatment: key role of energy absorption by intravascular blood. AB - BACKGROUND: Despite the clinical efficacy of endovenous laser treatment (EVLT), its mode of action is incompletely understood. OBJECTIVE: To evaluate the role of intravascular blood for the effective transfer of thermal damage to the vein wall through absorption of laser energy. METHODS: Laser energy (15 J/pulse, 940 nm) was endovenously administered to explanted greater saphenous vein (GSV) segments filled with blood (n = 5) or normal saline (n = 5) in addition to GSVs under in vivo conditions immediately prior to stripping. Histopathology was performed on serial sections to examine specific patterns of damage. Furthermore, in vitro generation of steam bubbles by different diode lasers (810, 940, and 980 nm) was examined in saline, plasma, and hemolytic blood. RESULTS: In saline-filled veins, EVLT-induced vessel wall injury was confined to the site of direct laser impact. In contrast, blood-filled veins exhibited thermal damage in more remote areas including the vein wall opposite to the laser impact. Steam bubbles were generated in hemolytic blood by all three lasers, while no bubbles could be produced in normal saline or plasma. CONCLUSION: Intravascular blood plays a key role for homogeneously distributed thermal damage of the inner vein wall during EVLT. PMID- 12135516 TI - Atypical presentation of pilomatricoma: a case report. AB - BACKGROUND: Malherbe tumors, also known as pilomatricomas, are benign cutaneous tumors of hair matrix origin. OBJECTIVE: To discuss a rare case of multiple pilomatricoma of the head and neck region in a very young patient. METHODS: A 14 month-old baby presented with two lesions, one appearing on the preauricular region and the other on the nasolabial fold. One of the lesions showed significant ulceration. RESULTS: Excisional biopsy was performed for both of the lesions. Although the histopathologic examination suggested pilomatricoma as the diagnosis, the lesion with ulcerations exhibited increased mitosis which made us consider pilomatrix carcinoma in its differential diagnosis. CONCLUSION: Increased mitoses, an uncommon unexpected feature for pilomatricoma, can be a marker for more aggressive biologic behavior. PMID- 12135515 TI - Computerized bipolar diathermy with scissors and forceps in cutaneous surgery. AB - BACKGROUND: Bipolar diathermy coagulates tissue as effectively as monopolar with less lateral tissue injury and no risk of interference with cardiac pacemakers or joint prostheses. OBJECTIVE: To test a novel computerized bipolar diathermy machine for combined cutting and coagulation in dermatologic surgery. METHODS: A divided cable was used to deliver current from a computerized bipolar diathermy unit to both scissors and forceps. The bipolar diathermy unit senses tissue contact with the instruments and starts automatically; a built-in microcomputer measures tissue impedance and automatically terminates the current when tissue coagulation is achieved. RESULTS: The equipment has been used successfully in more than 200 patients undergoing dermatologic surgery. The advantages were a reduced operating time and a more secure hemostasis. The microprocessor controlled bipolar diathermy unit minimized any tissue adherence to the instruments during use. CONCLUSION: We recommend the use of insulated scissors and computerized bipolar diathermy for safe and efficacious coagulation and cutting in dermatologic surgery. PMID- 12135517 TI - An unusual acute urticarial response following microdermabrasion. AB - BACKGROUND: Microdermabrasion is widely performed in a number of clinical settings, including medical offices, salons, and spas. This procedure is generally regarded as safe and easy to perform. OBJECTIVE: To determine if latex exposure caused an acute urticarial response following microdermabrasion in a latex-allergic patient. METHODS: The patient was prick tested to saline and histamine controls, latex, and sterile medical grade 100 m aluminum oxide crystals that had been passed through the microdermabrader. RESULTS: The strongly positive latex prick test confirmed latex allergy in our patient. Negative prick testing to aluminum oxide crystals that had passed through the microdermabrader make it unlikely that the patient was exposed to latex via this system. CONCLUSION: Physicians need to carefully evaluate patients who are considering microdermabrasion and appreciate that unexpected serious complications can occur. PMID- 12135518 TI - Chemical injury to the eye from trichloroacetic Acid. AB - BACKGROUND: Trichloroacetic acid (TCA) is frequently utilized for chemical peeling by physicians practicing dermatologic surgery. Ocular complications from TCA have not been reported previously. OBJECTIVE: The purpose of this article is to underscore the irritating and corrosive effects of TCA on the eye. METHODS: A patient is described who experienced seepage of 35% TCA into the eye during a chemical peel. RESULTS: The patient developed marked conjunctivitis of the affected eye and abrasions involving 25% of the cornea. CONCLUSION: TCA must be applied carefully around the eyes to avoid ocular complications, which albeit rare, can be quite grave if not addressed in a timely manner. PMID- 12135519 TI - CO2 laser treatment of epidermal nevi: long-term success. AB - BACKGROUND: Epidermal nevi have been notoriously difficult to treat due to their large size and often conspicuous location. Variable results have been obtained with different laser treatments, and scarring and/or incomplete removal is typical after excisional or other destructive modalities. OBJECTIVE: To outline the successful use of a short-pulsed CO2 laser in the long-term eradication of epidermal nevi in three patients. METHODS: Three females (ages 15-19) presented with extensive grouped verrucous papules and plaques on the face, trunk, and extremities. A pulsed CO2 laser was used to vaporize the lesions using a 500 mJ pulse energy, 3 mm spotsize, and 7 watts of power. RESULTS: All lesions healed without incident. No lesional recurrence was observed 10 to 13 months after treatment except in one small area on the ankle in one patient. CONCLUSIONS: Carbon dioxide laser vaporization of epidermal nevi provides good clinical effect and offers unique advantages for the treatment of these lesions, including effective intraoperative hemostasis with excellent lesional visualization. It is also possible to treat widespread areas in one laser treatment session. While the results of this series clearly show the benefit of CO2 laser treatment, epidermal nevi may not always respond so favorably, due in part to the variability in their depths of involvement. PMID- 12135520 TI - Use of a 585 nm pulsed dye laser for the treatment of morphea. AB - INTRODUCTION: The clinical presentation of morphea varies from localized plaques to generalized eruptions. Its cause remains unknown and medical treatments have often proved unsatisfactory. Studies have previously shown that improvement of hypertrophic scars and fibrotic skin can be achieved with the use of a 585 nm pulsed dye laser (PDL). METHODS: A case of plaque-type morphea was treated with 585 nm pulsed dye laser irradiation at an average fluence of 5.0 J/cm2 at bimonthly time intervals. RESULTS: Marked clinical improvement as evidenced by improved pliability and skin coloration was seen after 4 successive PDL treatments. No side effects or complications were encountered. CONCLUSION: Pulsed dye laser therapy is a viable treatment option for morphea. The mechanism of its effect in this condition remains unknown. PMID- 12135521 TI - Treatment of pearly penile papules with CO2 laser. AB - BACKGROUND: Pearly penile papules are angiofibromas found on the corona and sulcus of the glans penis. While these represent a benign condition, psychological and cosmetic concerns often prompt patients to seek therapeutic removal of these lesions. Multiple therapeutic modalities have been reported; however, use of CO2 laser has proven to be the most effective to date. OBJECTIVE: To demonstrate the efficacy of CO2 laser in treating pearly penile papules. METHODS: Case report. RESULTS: We report the successful treatment of pearly penile papules in an African American man with CO2 laser. CONCLUSION: The use of CO2 laser is an effective treatment for pearly penile papules. J. E. Lane, MD, C. M. Peterson, MD, and J. L. Ratz, MD have indicated no significant interest with commercial supporters. PMID- 12135522 TI - Duplex-guided foam sclerotherapy for the treatment of the symptomatic venous malformations of the face. AB - BACKGROUND: New sclerosing foam is considered to have the advantage of causing more damage on the intima than liquid form. Therefore we recently applied duplex guided foam sclerotherapy in a patient with venous malformations of the face. METHODS: A 20-year-old man was referred to our institute for the evaluation and treatment of vascular malformations of the face. Preoperative duplex scanning and magnetic resonance imaging (MRI) revealed subcutaneous and intramuscular venous malformations. The sclerosing foam was produced by Tessari's method using 1% polidocanol, and the duplex-guided foam sclerotherapy was performed under general anesthesia. A 20-gauge plastic needle was inserted into the venous space using ultrasound guidance and a total of 5 ml of sclerosing foam was infused followed by immediate tie-over dressing. RESULTS: The venous malformations were successfully reduced in size and postoperative MRI showed significant reduction of the venous malformations. CONCLUSION: Although further collective study is necessary to ensure the validity of this treatment, duplex-guided foam sclerotherapy could have great promise in the treatment of symptomatic venous malformations. PMID- 12135523 TI - Sebaceous carcinoma of the eyelids treated by mohs micrographic surgery: report of nine cases with review of the literature. AB - BACKGROUND: Ocular sebaceous carcinoma (SC) is a rare tumor of the eyelids. Clinically it frequently involves the upper lid in older women. Microscopically it tends to extend far beyond its assessed clinical margins. SC is characterized by a variety of tissue invasion mechanisms. It may spread by direct extension, be multifocal in advanced cases, and develop "skip areas" after trauma. Despite its ability to develop discontinuity, over the past two decades there have been several favorable case reports of SC treated by Mohs micrographic surgery (MMS). OBJECTIVE: To illustrate by case report the clinical presentation and management of patients with SC and to document our series of SC patients treated by MMS over the last 14 years. METHODS: A retrospective study was conducted of all cases of sebaceous carcinoma involving the ocular adnexa treated at the University of Wisconsin Mohs Surgery Clinic from 1987 to 2001. We also reviewed the accumulated medical literature of SC treated by MMS. RESULTS: In our series, there were nine cases of periocular SC. Five cases originated on the upper lid and four on the lower lid. Five of nine patients (55%) showed epithelial invasion. One patient developed a local recurrence 1.5 years later and was treated by orbital exenteration. This patient has had no evidence of disease for 5 years. Eight of nine patients (88%) had no evidence of local recurrence with a follow-up of 1-14 years. In our literature review we found 40 additional cases of orbital SC treated by MMS. Intraepithelial spread of SC was found in 50% of patients (24 of 48). Six patients developed local recurrence. The overall local cure rate following MMS is 87.8% (43 of 49), with a mean follow-up of 3.1 years. The regional metastatic rate was 8% (4 of 49). There were no deaths reported. CONCLUSION: We present nine new cases of SC. The age, sex, and site distribution are compatible with other SC cases reported in the literature. We reviewed the medical literature and compiled 49 cases of SC treated by Mohs surgery. Intraepithelial spread was discovered in 50% of the cases. Multifocal disease or discontinuity was present in 6% (3 of 49). Mohs surgery appears to be an effective method for excising the microscopic ramifications of primary SC. When feasible, we recommend in SC cases where intraepithelial pagetoid spread has been observed, that removal of another Mohs layer should be considered in order to provide an additional assurance layer against local tumor recurrence. PMID- 12135524 TI - Standing cone avoidance via advancement flap modification. AB - BACKGROUND: Tissue redundancy manifesting as a standing cutaneous cone is a common surgical problem. OBJECTIVE: We describe modifications to traditional advancement flap design that allows for standing cutaneous cone avoidance. METHODS: In addition to traditional incisions employed in advancement flaps, an additional curvilinear incision is made along one or more limbs of the flap. By removing tissue along the length of the flap incision, redundant tissue is redistributed along the length of the incision. A standing cutaneous cone is thereby avoided. RESULTS: Use of the technique provides standing cone avoidance without lengthening a surgical scar. There has been no increase in flap failure or other complications with the described technique. CONCLUSION: The modified advancement flap allows for standing cone avoidance. Improved cosmesis can also be achieved when the modified flap limb is placed in a facial line or subunit junction. PMID- 12135525 TI - Gouty tophi: a squamous cell carcinoma mimicker? AB - BACKGROUND: Digital lesions can have a broad differential diagnosis. Squamous cell carcinoma (SCC), the most common digital malignant neoplasm, must be excluded as the cause of persistent digital lesions causing nail dystrophy. OBJECTIVE: To describe a patient with a periungual hyperkeratotic lesion on the left fifth digit which, upon initial dermatopathologic examination, appeared to be a malignancy. However, on further biopsy, the lesion proved to be a gouty tophus. METHODS: Case report and literature review. RESULTS: An 84-year-old white man presented with a hyperkeratotic papule on the lateral proximal nail fold of the left fifth digit, which resulted in nail dystrophy for 1 year. Similar lesions were present on several other digits which did not affect the nail plate. Initial biopsy was consistent with actinic keratosis and was treated with cryotherapy. When the lesion persisted, repeat biopsy was performed, demonstrating fragments of squamous epithelium with focal atypia and an infiltrative growth pattern. SCC could not be excluded and the patient was referred for Mohs micrographic surgery (MMS) consultation. An excisional biopsy was performed and a white chalky material was observed at the base of the defect. Histopathology confirmed a gouty tophus. The patient was referred to his primary care physician and was treated with allopurinol. CONCLUSION: This is the first report of gouty tophus of the periungual region presenting as a hyperkeratotic lesion. Initial clinical diagnosis favored SCC and histologic evidence suggested a possible early SCC. This lesion can be confused with digital squamous cell carcinoma. The presence of pseu- docarcinomatous hyperplasia may complicate accurate diagnosis. PMID- 12135526 TI - Giant epidermoid cyst on the forefoot. AB - BACKGROUND: Epidermoid cysts are the most common cysts of the skin, usually appearing in the hairy regions. However, epidermoid cysts on the non-hair-bearing areas, such as the palms and the soles, are rare and seldom reach a size of more than 5 cm in diameter. OBJECTIVE: To present a patient with an epidermoid cyst of unusual location and size. METHODS: A 23-year-old woman with a painless tumorlike mass on her right forefoot was examined and the mass was excised. RESULTS: The 6.5 cm x 5.5 cm x 5.5 cm mass was totally excised. Histologic diagnosis was reported as an epidermoid cyst. CONCLUSION: An original case regarding an unusual location and size of an epidermoid cyst is presented. PMID- 12135527 TI - Tibialis anterior muscle herniation. AB - BACKGROUND: Tibialis anterior herniation occurs through a defect in the deep fascial layer of the lower extremity. While this entity is not uncommon, it has been rarely reported in the dermatologic literature. OBJECTIVE: To highlight the occurrence of tibialis anterior herniation and its presentation and treatment. METHODS: Case report. RESULTS: We present a case of tibialis anterior herniation in a patient who presented with asymptomatic nodules in her lower extremities. The nodules were flesh-colored, soft, and compressible with palpation. CONCLUSION: The diagnosis of tibialis anterior herniation should be considered in patients with positionally variable subcutaneous nodules, especially in the lower extremities. It is caused by a defect in the deep fascial layer and often involves the lower extremities. In addition, knowledge of the diagnosis and treatment is essential for both proper management and/or surgical referral. PMID- 12135528 TI - Interobserver agreement on dermoscopic features of pigmented basal cell carcinoma. AB - BACKGROUND: A dermoscopic method based on the absence of a pigment network and the presence of at least one of six positive features has been described for diagnosis of pigmented basal cell carcinoma (BCC). OBJECTIVE: To evaluate the observers' global agreement and interobserver agreement on each dermoscopic parameter of the method recently proposed. METHODS: Dermoscopic images of 56 pigmented BCCs were examined by five observers with different degrees of experience in dermoscopy. RESULTS: An overall full agreement was reached for the absence of pigment network (k = 1). Very good agreement was detected for the presence of spoke wheel areas (k = 0.85) and arborizing vessels (k = 0.72), and good agreement was shown for ulceration (k = 0.49) and multiple blue-gray globules (k = 0.41). No agreement was identified on large blue-gray ovoid nests (k = 0.28) and leaflike areas (k = 0.26). CONCLUSION: We confirm the reproducibility of the method and show that ulceration, spoke wheel areas, and arborizing tel- angiectases represent the most robust positive parameters. PMID- 12135529 TI - Giant melanoma displaying gross features reproducing parameters seen on dermoscopy. AB - BACKGROUND: The basis of dermoscopy is the strict relationship between dermoscopic features and histology of pigmented skin lesions. METHODS: When observed under dermoscopy, a pigmented lesion displays a series of features which enable an experienced clinician to classify the lesion with higher accuracy than that expected by visual inspection alone. RESULTS: We have observed a peculiar case of a giant cutaneous melanoma occurring on the abdomen of a 45-year-old woman, displaying clinical features strictly resembling those of melanoma under dermoscopy. DISCUSSION: The interest of this case is actually purely morphologic, showing for the first time, depending on the extraordinary growth of the lesion, the appearance of gross features strictly reproducing those microscopically found under dermoscopy in smaller lesions. PMID- 12135530 TI - Comment on R. A. Weiss : "comparison of endovenous radiofrequency versus 810 nm diode laser occlusion of large veins in an animal model". PMID- 12135531 TI - Skin-only suturing and facilitation of cartilage movement in auricular helical rim wedge resection. PMID- 12135532 TI - Reduced body mass, food intake, and testis size in response to short photoperiod in adult F344 rats. AB - BACKGROUND: Although laboratory rats are often considered classic nonseasonal breeders, peripubertal rats of two inbred strains, F344 and BN, have both reproductive and nonreproductive responses to short photoperiods. Unmanipulated adult rats have not been reported to have robust responses to short photoperiod alone, although several treatments can induce photoperiodic responses in adults. In this study, we tested the hypotheses that unmanipulated F344 rats retain responses to short photoperiod as adults and that they have the necessary elements for an endogenous circannual rhythm of sensitivity to short photoperiod. RESULTS: Relative to rats kept in long photoperiods (L16:D8), adult F344 rats transferred at 4.5 months of age to short photoperiods (L8:D16) had significantly lower testis size, food intake, and body weight. In a second experiment, newly weaned F344 rats underwent an initial period of inhibition of reproductive maturation, lower food intake, and lower body weight in short photoperiod or intermediate photoperiod (L12:D12) relative to rats in long photoperiod. By 18 weeks of treatment, rats in the two inhibitory photoperiods no longer differed from long photoperiod controls. In short photoperiod, rats underwent a second period of slight reproductive inhibition between weeks 35 and 48, but there was an effect on body weight and slight inhibition of food intake only in an intermediate photoperiod. CONCLUSION: Male F344 rats retain photoresponsiveness as adults, with less reproductive inhibition but equivalent nonreproductive responses. There was only weak evidence for an endogenous timer controlling a circannual cycle of sensitivity to short photoperiod. PMID- 12135533 TI - Mutational analysis of Peroxiredoxin IV: exclusion of a positional candidate for multinodular goitre. AB - BACKGROUND: Multinodular goitre (MNG) is a common disorder characterised by an enlargement of the thyroid, occurring as a compensatory response to hormonogenesis impairment. The incidence of MNG is dependent on sex (female:male ratio 5:1) and several reports have documented a genetic basis for the disease. Last year we mapped a MNG locus to chromosome Xp22 in a region containing the peroxiredoxin IV (Prx-IV) gene. Since Prx-IV is involved in the removal of H2O2 in thyroid cells, we hypothesize that mutations in Prx-IV gene are involved in pathogenesis of MNG. METHODS: Four individuals (2 affected, 2 unrelated unaffected) were sequenced using automated methods. All individuals were originated from the original three-generation Italian family described in previous studies. A Southern blot analysis using a Prx-IV full-length cDNA as a probe was performed in order to exclude genomic rearrangements and/or intronic mutations. In addition a RT-PCR of PRX-IV was performed in order to investigate expression alterations. RESULTS: No causative mutations were found. Two adjacent nucleotide substitutions were detected within introns 1 and 4. These changes were also detected in unaffected individuals, suggesting that they were innocuous polymorphisms. No gross genomic rearrangements and/or restriction fragment alterations were observed on Southern analysis. Finally, using RT-PCR from tissue specific RNA, no differences of PRX-IV expression-levels were detected between affected and unaffected samples. CONCLUSIONS: Based on sequence and genomic analysis, Prx-IV is very unlikely to be the MNG2 gene. PMID- 12135534 TI - Stimulant-like action of nicotine on striatal dopamine transporter in the brain of adults with attention deficit hyperactivity disorder. AB - Eleven adult patients with attention deficit hyperactivity disorder (ADHD) without medication, consuming 7-40 cigarettes per day, showed statistically significant lower values for striatal dopamine transporter (DAT) measured by [99mTc]TRODAT-1 SPECT compared to 11 non-smoking drug-naive patients with ADHD, matched for sex and age, despite higher ADHD scores for the smokers. Because stimulants have been shown to reduce primarily elevated DAT density in adults with ADHD, it can be suggested that nicotine acts in a similar way on striatal DAT as do stimulants. PMID- 12135535 TI - Venlafaxine compared with fluoxetine in outpatients with depression and concomitant anxiety. AB - The aim of this double-blind study was to compare the efficacy and safety of venlafaxine vs. fluoxetine in the treatment of patients with depression and anxiety. A total of 146 moderately depressed patients with associated anxiety were randomized to receive 75 mg/d venlafaxine or 20 mg/d fluoxetine for 12 wk. Dose increases were permitted after 2 wk of treatment, to 150 mg/d venlafaxine and 40 mg/d fluoxetine, to optimize response. At the final visit, a statistically significantly greater efficacy of venlafaxine over fluoxetine was observed on depressive symptoms and concomitant anxiety, and 75.0 and 50.7% of patients administered venlafaxine and fluoxetine, respectively, showed an overall response. A sustained response (for at least 2 wk), present at the end of the study was achieved in 57.8 and 43.3% of patients in the venlafaxine and fluoxetine groups, respectively, and at the final visit, 59.4 and 40.3% of patients, respectively, were in remission (virtually asymptomatic). Dose increases were required by a greater percentage of patients in the fluoxetine group (52.9%), than in the venlafaxine group (37.1%), and in those patients whose dose was increased, a higher efficacy was again observed with venlafaxine. Venlafaxine and fluoxetine were well tolerated, with the most frequently experienced adverse events being nausea and headache. Fewer patients in the venlafaxine group than in the fluoxetine group reported at least one adverse event (55.7 and 67.1% patients, respectively). Venlafaxine therefore proved to be significantly more effective than fluoxetine in improving depressive symptoms and concomitant anxiety. PMID- 12135536 TI - Efficacy of Adderall and methylphenidate in attention deficit hyperactivity disorder: a drug-placebo and drug-drug response curve analysis of a naturalistic study. AB - Stimulant medication has, for many years, been the pharmacological treatment of choice for children and adults with attention deficit hyperactivity disorder (ADHD). Recently, several studies have documented the efficacy of a new stimulant, Adderall. Although these initial studies provide useful information for clinicians treating ADHD children, their method of data presentation has provided limited information about the clinical significance of drug effects. Thus, to address the issue of clinical significance, we completed drug-placebo response curve analyses of a blinded, placebo-controlled study of Adderall and methylphenidate (MPH). Our results show that the efficacy of Adderall and MPH to improve functioning is seen throughout the full range of improvement scores. Both drugs prevent worsening and, for a majority of patients, lead to improvements that are well into the normal range. The analyses also highlight an important subgroup of placebo responders, which suggests that future research should focus on how to predict robust placebo response in ADHD patients. PMID- 12135538 TI - Topiramate attenuates self-injurious behaviour in Prader-Willi Syndrome. AB - Self-injurious behaviour (SIB), most notably skin picking, has been described by various terms in the literature ranging from neurotic/psychogenic excoriations to compulsive/pathological skin picking. Prader-Willi Syndrome (PWS) is a neurogenetic multisystem disorder characterized by infantile hypotonia, mental retardation, short stature, hypogonadism, dysmorphic features, and hyperphagia with a high risk of obesity. Psychiatric manifestations include SIBs in the form of skin picking, nail biting and rectal gouging. Topiramate is a novel anti epileptic medication without significant liability of weight gain. There are no published reports of topiramate being utilized in PWS or SIB. We report attenuation of SIB with resultant lesion healing in three PWS adults treated with topiramate in an 8-wk open-label trial. Although our findings should be treated with caution, they suggest that double-blind or cross-over studies with topiramate are warranted to establish the possible role of topiramate in attenuating SIB in PWS and other disorders that involve SIB. PMID- 12135537 TI - Region-dependent effects of flibanserin and buspirone on adenylyl cyclase activity in the human brain. AB - The mode of action of antidepressant drugs may be related to mechanisms of receptor adaptation, involving overall the serotonin 1A (5-HT1A) receptor subtype. However, so far, the clinical effectiveness of selective compounds acting at this level has proved disappointing. This could be explained by the heterogeneity of 5-HT1A receptors within the central nervous system. In animals, two 5-HT1A agonists, flibanserin and buspirone, have shown different pharmacological properties, depending on the brain region. Since no evidence supports this observation in humans, this study sought to investigate whether these two drugs exert different effects on 5-HT1A receptor activation in three different human brain areas: the prefrontal cortex, hippocampus and raphe nuclei. 5-HT1A-mediated inhibition of forskolin-stimulated adenylyl cyclase (AC) was taken as an index of 5-HT1A receptor activation. Flibanserin significantly reduced the activity of AC post-synaptically, i.e. in the prefrontal cortex [EC50 (mean +/- S.E.M.), 28 +/- 10.2 nM; Emax, 18 +/- 2.3%] and in the hippocampus (EC50, 3.5 +/- 3.1 nM; Emax, 20 +/- 4.0%), but had no effect in the raphe nuclei, i.e. at pre-synaptic level. Vice versa, buspirone was only slightly but significantly effective in the raphe (EC50, 3.0 +/- 2.8 nM; Emax, 12 +/- 1.9%). Agonist effects were sensitive to the 5-HT1A antagonists WAY-100135 and pindobind 5-HT1A in the cortex and raphe nuclei, whereas buspirone antagonized flibanserin in the hippocampus. These findings suggest a region-related action of flibanserin and buspirone on forskolin-stimulated AC activity in human brain. PMID- 12135539 TI - Emotional blunting associated with SSRI-induced sexual dysfunction. Do SSRIs inhibit emotional responses? AB - Anecdotal and published case reports suggest that some patients taking selective serotonin reuptake inhibitors (SSRI) experience diminution in emotional responsiveness. This study aims to define the individual components of emotion disturbed in these patients. Fifteen patients reporting SSRI-induced sexual dysfunction completed the Laukes Emotional Intensity Scale (LEIS), a questionnaire about various emotions. Compared to controls, patients reported significantly (p<0.05) less ability to cry, irritation, care about others' feelings, sadness, erotic dreaming, creativity, surprise, anger, expression of their feelings, worry over things or situations, sexual pleasure, and interest in sex. Total score on the LEIS did not correlate with total score on the Hamilton Depression Rating Scale. In our sample, 80% of patients with SSRI-induced sexual dysfunction also describe clinically significant blunting of several emotions. Emotional blunting may be an under-appreciated side-effect of SSRIs that may contribute to treatment non-compliance and/or reduced quality of life. PMID- 12135540 TI - The effect of chronic treatment with clozapine and haloperidol on stimulus control by DOM. AB - The present study investigated the effects of chronic treatment with the atypical antipsychotic, clozapine, or the typical antipsychotic, haloperidol, on the stimulus properties of 2,5-dimethoxy-4-methylamphetamine ([-]-DOM) in rats trained to discriminate [-]-DOM (0.3 mg/kg; 75 min pre-treatment time) from vehicle. As compared with control values, treatment with clozapine (25 mg/kg.d) for 7 d caused a statistically significant 57% reduction in [-]-DOM-appropriate responding. Unlike clozapine, treatment with haloperidol (1 mg/kg.d) for 7 d did not affect the stimulus properties of [-]-DOM. These findings demonstrate that a functionally significant decrease in 5-HT2A receptor-mediated activity is a unique component of the in-vivo response to chronic treatment with clozapine but not haloperidol and, therefore, might account for some of the clinical differences associated with atypical antipsychotics. PMID- 12135541 TI - The GABAergic system in schizophrenia. AB - A defect in neurotransmission involving gamma-amino butyric acid (GABA) in schizophrenia was first proposed in the early 1970s. Since that time, a considerable effort has been made to find such a defect in components of the GABAergic system. After a brief introduction focusing on historical perspectives, this paper reviews post-mortem and other biological studies examining the following components of the GABAergic system in schizophrenic subjects: the GABA biosynthetic enzyme, glutamate decarboxylase; free GABA; the GABA transporter; the GABAA, GABAB and benzodiazepine receptors; and the catabolic enzyme GABA transaminase. Additionally, post-mortem studies using morphology or calcium binding protein to identify GABAergic neurons are also reviewed. Substantial evidence argues for a defect in the GABAergic system of the frontal cortex in schizophrenia which is limited to the parvalbumin-class of GABAergic interneurons. PMID- 12135542 TI - Treatment non-response in OCD: methodological issues and operational definitions. AB - While controlled trials with SRIs have demonstrated a selective efficacy in obsessive-compulsive disorder (OCD), up to 40-60% of patients do not have a satisfactory outcome. Non-response to treatment in OCD is associated with serious social disability. There are a large number of non-responsive patients, and they are difficult to cluster due to ambiguities in the diagnostic criteria, possibility of subtypes, and a high rate of comorbidity. Moreover, the findings of current studies of so-called 'non-responsive' cases are currently non generalizable because of the lack of an operational definition of non-response. The result has been that a cumulative body of data on a reasonably homogeneous sample of non-responders has not been developed. The aims of this paper are to clarify some of the obstacles in defining stages of response and levels of non response and, through a comprehensive analysis, to propose a systematic nosology for this rather common condition. Better characterization of which patients respond and do not respond to various treatments will enable more accurate clustering of patients, and help facilitate multi-site data collection for future research trials. PMID- 12135543 TI - Vertebral height, disc height, posteroanterior displacement and dens-atlas gap in the cervical spine: precision measurement protocol and normal data. AB - OBJECTIVE: (1) Precise measurement of vertebral height, disc height, posteroanterior displacement and dens-atlas gap from lateral radiographic views of the cervical spine. (2) Compilation of a normative database for these parameters, specifying dependence on gender and age. DESIGN: Descriptive study, based on measurements from lateral radiographic views of the cervical spine of healthy subjects. BACKGROUND: Normal data of vertebral height, disc height, posteroanterior displacement and size of the dens-atlas gap as well as their biological range of variation and potential dependence on gender and age are not available. METHODS: Based on computer-aided measurements from lateral radiographic views of the cervical spine, a new protocol determines these parameters. RESULTS: are compensated for radiographic magnification, variation in stature and the individually adopted posture of the cervical spine; they are virtually uninfluenced by radiographic distortion and patient alignment errors. A specimen study as well as inter- and intra-observer studies quantify measurement errors.Results. Employing the new protocol, vertebral height C3-C7 and disc height C2/C3-C6/C7 are measured with relative errors of 3.9% and 5.7% respectively. Posteroanterior displacement C1/C2 to C6/C7 is measured with an error of 2.8% of mean vertebral depth and the dens-atlas gap is measured with an error of <1.8% of the depth of C2. A normal database for the dimensions of cervical vertebrae and discs as well as of the sagittal plane alignment of the vertebrae within the cervical spine is compiled from 135 lateral views of healthy adults. CONCLUSIONS: Vertebral height, disc height, posteroanterior displacement and size of the dens-atlas gap are measured with high precision. Normal data are presented for the first time. RELEVANCE: The new protocol in conjunction with the normal database enables future studies detecting or monitoring morphological effects of, for example, trauma, long-term high mechanical loading, disc degeneration, rheumatoid arthritis, fusion or other surgical interventions. PMID- 12135544 TI - The traumatic spondylolisthesis of the axis. A biomechanical in vitro evaluation of an instability model and clinical relevant constructs for stabilization. AB - OBJECTIVE: Stepwise destabilization of the traumatic spondylolisthesis C2 with an increasing anterior defect of C2-C3 was investigated. The three-dimensional stabilizing capabilities of posterior transpedicle screw osteosynthesis and two anterior plate systems C2-C3, the H-plate and the titanium locking plate were tested. DESIGN: A biomechanical in vitro study was performed using a standardized experimental protocol in a spine tester. BACKGROUND: The extent of the instability of the traumatic spondylolisthesis of C2 within its different types remains unclear. Posterior and anterior approaches for stabilization exist for patients with isthmusfractures at C2, the stabilizing effect has not been demonstrated yet. METHODS: The motion levels from C2-C3 in six human specimen were tested in flexion, extension, right and left lateral bending and left and right axial rotation. The specimens were tested intact, after destabilization and after stabilization. RESULTS: In extension and axial rotation, each step of destabilization decreased the moment significantly, to achieve the range of motion of the intact specimen. In flexion a statistical significant destabilization after separation of the posterior longitudinal ligament was present. The flexibility tests showed an increasing range of motion of the posterior transpedicle screws, with increasing anterior instability markedly in flexion and extension. After H-plate and locking plate fixation, a significant decrease of the range of motion resulted in flexion. The stiffness in flexion and extension increased consecutively, while in lateral bending and axial rotation the transpedicle screw fixation showed the highest stiffness. CONCLUSIONS/RELEVANCE: The traumatic spondylolisthesis of C2 is a significantly unstable injury in case of additional segmental damage of C2-C3. Anterior stabilization in these injuries is mandatory. PMID- 12135545 TI - Comparison of the mechanical behavior of the lumbar spine following mono- and bisegmental stabilization. AB - OBJECTIVE: To determine whether the mechanical behavior of the entire lumbar spine differs following mono- and bisegmental stabilization. DESIGN: The mechanical behavior of the lumbar spine was studied using the finite element method. BACKGROUND: Nonunion is somewhat more frequent after bi- than after monosegmental stabilization of the spine. Little is known about differences between the mechanical behavior associated with these procedures. METHODS: A three-dimensional nonlinear finite element model of the lumbar spine with internal spinal fixators and bone grafts was used to study mechanical behavior after mono- and bisegmental fixation with and without stabilization of the bridged vertebra. Finite element analyses were performed to determine the influence of four different graft positions, five loading conditions, and six different pretensions in the longitudinal fixator rod. The following parameters were considered: the maximum contact pressure at the interface between the bone graft and vertebral body, the force transmitted by the bone graft, and the size of the contact area between the graft and the vertebral body. RESULTS: Our model shows no clear differences between mono- and bisegmental fixation. Additional stabilization of the bridged vertebra exerts a partly adverse influence on the parameters studied. Pretension in the bridged region has a strong effect on the mechanical behavior. CONCLUSIONS: The mechanical behavior of the lumbar spine after mono- and bisegmental stabilization is similar. Thus biological factors and the surgical procedure are probably decisive in determining the fusion rate. RELEVANCE: Knowledge of the mechanical behavior after stabilization of the spine may help to improve the fusion rate. Our results suggest that the mechanical factors studied have only a minor influence on fusion rate and that other factors, such as incomplete resection of cartilage plate and poor local blood supply, are more decisive. PMID- 12135546 TI - The effect of hydration on the stiffness of intervertebral discs in an ovine model. AB - OBJECTIVE: To determine the hydration-over-time behaviour of ovine intervertebral discs and intact joints in a saline bath at body temperature and the effect this has on their stiffness compared to air at ambient temperature. DESIGN: The hydration-over-time behaviour and stiffness of the ovine functional spinal unit and disc were quantified. BACKGROUND: The fluid content of an intervertebral disc is not constant but varies with external load and load history. The stiffness of ovine functional spinal units in a hydrated environment and how this compares to testing in air have not been quantified. METHODS: Intervertebral discs and functional spinal units were weighed and soaked in a saline water bath at 37 degrees C and reweighed each hour for 6 h. They were then allowed to stand in air at room temperature while the time to return to initial weight was recorded. Functional spinal units were randomly assigned to two groups. Axial compression, flexion, extension, lateral bending and axial torsion tests were performed on both the intact functional spinal unit and isolated disc. Group 1 was tested in air then in a saline water bath at 37 degrees C with the testing order reversed for Group 2. RESULTS: Hydration of the disc reached a plateau after an average 3 4 h of soaking with the largest increase seen in the first hour. Four hours, standing in air at room temperature, was required to return specimens to their initial weight. The functional spinal unit stiffness was significantly lower for those specimens tested in the bath compared to air. CONCLUSIONS: Ovine intervertebral discs show similar hydration-over-time behaviour when compared to human discs. Stiffnesses in different modes of loading were significantly different when tested in a hydrated environment compared with the standard method of testing in air. RELEVANCE: It has been shown that there are biomechanical and biochemical similarities between sheep and human intervertebral discs. Despite these similarities, no studies have looked at how ovine intervertebral discs behave over time in a hydrated environment. In humans, hydration levels are an important aspect of intervertebral disc degeneration. There is also a relationship between decreased hydration levels and increased stiffness. This study demonstrates the similarities between human and ovine hydration-over-time behaviour. The importance of intervertebral disc hydration and its effects on stiffness under different modes of loading were also demonstrated and have not been previously shown using the ovine model. In this context, the results from this study provide further support for the use of the ovine model. PMID- 12135547 TI - Mechanism of limitation of pronation/supination of the forearm in geometric models of deformities of the forearm bones. AB - OBJECTIVE: To clarify the mechanism of limitation of pronation/supination of the forearm associated with the angular deformity of the forearm bones and narrowing of the interosseous space. DESIGN: A three-dimensional geometric model of the forearm bones with the interosseous membrane and its axial section were used. BACKGROUND: Limitation of pronation/supination associated with the deformity of the forearm bones is one of the significant problems encountered in the treatment of the forearm fracture. Elucidation of its mechanism is important for its prevention and treatment. METHODS: In the axial section, the effects of the positional relationship between the axis of pronation/supination and the forearm bones on the range of pronation/supination was studied using analytic geometry in each model with non-narrowing or narrowing of the interosseous space. Subsequently, in the three-dimensional model each forearm bone with the same angular deformity, the direction and magnitude of the angular deformity which would lead to limitation of pronation/supination were calculated using analytic geometry. Each parameter of the models was obtained by the radiographic measurements of the normal forearms. RESULTS: When the axis of the pronation/supination passed through the interosseous region (less than 2 cm radioulnarly and 0.8 cm anteroposteriorly) in the model of the axial section without narrowing of the interosseous space, more than 40 degrees of pronation and supination were possible. When the axis deviated from this region, significant loss of pronation/supination was observed associated with restriction by the interosseous membrane rather than impingement. Furthermore, the area of this region decreased according to narrowing of the interosseous space with shortening of the interosseous membrane. In the three-dimensional model, the direction and magnitude of the angular deformity which would lead to significant loss of pronation/supination was more than 14 degrees radially, 7 degrees ulnarly, 5 degrees anteriorly, 4 degrees posteriorly. CONCLUSIONS: The positional relationship between the axis of pronation/supination and the forearm bones with the interosseous membrane may play an important role regarding pronation/supination of the forearm. RELEVANCE: Evaluation of the bone deformities based on understanding this mechanism of limitation of pronation/supination would lead to an appropriate treatment of malunion of the forearm bones. PMID- 12135548 TI - The relationship between EMG median frequency and low frequency band amplitude changes at different levels of muscle capacity. AB - OBJECTIVE: To test the validity of high and low frequency band amplitudes of the surface electromyography (EMG) profile as representation of muscle fatigue. DESIGN: A within subjects (n=10) repeated measures design was used to collect surface EMG signals from the biceps during an isometric contraction under two levels of fatigue status. BACKGROUND: The use of the shift in the median frequency of the surface EMG power spectrum is a well known method of assessing muscle fatigue. Fatigue also results in amplitude changes of the specific frequency bands. The use of frequency band analysis may be an alternative option for the assessment of muscle fatigue in specific experimental settings. METHODS: Surface EMG profiles of the biceps were recorded at 1024 Hz during a sustained isometric hold at 60% of the individuals fresh and fatigued maximal voluntary isometric torque. The median frequency of the power spectrum was compared with changes in the low frequency (15-45 Hz) and high frequency (>95 Hz) bands.Results. There was a close association between median frequency shift and the amplitude of the 15-45 Hz bandwidth and the high-low frequency amplitude ratio. The association was similar for performance under different muscle capacity states. CONCLUSIONS: Frequency band amplitude analysis provides similar information to median frequency shift under isometric conditions and may be suited to specific experimental protocols in workplace fatigue studies. RELEVANCE: The use of amplitude band analysis that closely approximates the standard median frequency changes allows greater possibility of assessing muscle fatigue in different experimental settings and the use of lower sampling rates. PMID- 12135549 TI - Effects of one-plane and two-plane external fixation on sheep osteotomy healing and complications. AB - OBJECTIVES: Comparison of one-plane and two-plane external fixation in terms of successful healing, incidence of complications, and biomechanical stability in a sheep model. BACKGROUND: Rigid fixation is preferred in open and comminuted fractures with a reduced blood supply, preventing infection and healing delay, but more often a flexible device is recommended even in unfavourable healing conditions. METHODS: The left tibiae of fifteen sheep were osteotomized and laterally fixed with a four-screw unilateral fixator frame (axial stiffness 183 N/mm) to a 3 mm gap size. In 9 of 15 sheep, an additional four-screw unilateral external fixator was anterolaterally attached (total axial stiffness of both frames 388 N/mm). After sacrificing, quality of osteotomy healing was assessed by mechanical and radiological evaluations. Osteogenesis was measured using fluorescence microscopy. RESULTS: Two distal fractures through the pin-tracks, three non-unions and four deep infections occurred after two-plane fixation. These failures excluded, osteotomy healing showed inferior results after two plane fixation with reduced callus formation, bone mineral content, and bending stiffness amounts, respectively. Osteogenesis was halved following two-plane fixation in the remaining sheep. CONCLUSIONS: Two-plane fixation was not sufficient to reach successful osteotomy healing in our study. While higher rigidity was expected to prevent complications, healing in this group might have been disturbed by a reduced blood supply. The optimal stabilisation for a given fracture depends on many factors, including the biomechanical and biological environment. RELEVANCE: Considering our results and the literature discussed in this manuscript, good bone healing with minor risks of infections can be achieved using an unilateral one-plane fixator with only four screws, and its application on a muscle free position like the medial and anterior site of the sheep tibia. PMID- 12135550 TI - Tibiofemoral contact points relative to flexion angle measured with MRI. AB - OBJECTIVE: To determine whether knee flexion influenced bony contact movements during flexion. DESIGN: Accurate three-dimensional (3D) measurements of tibiofemoral bony contact points in vivo was performed using magnetic resonance imaging technology at 0 degrees, 30 degrees and 60 degrees of flexion. BACKGROUND: Magnetic resonance imaging is an accurate non-invasive tool for visualizing muscles, tendons, and bone, and provides precise 3D co-ordinates. METHODS: Magnetic resonance imaging recordings were made from the right knee of 16 subjects with no history of knee dysfunction at 0 degrees, 30 degrees and 60 degrees of flexion. Joint contact movements were reported as changes of the contact point's position on the medial and lateral tibial condyle with respect to a fixed reference point for each flexion angle. RESULTS: The dominant motion of the centroid of the contact area was posterior with a concomitant inferior and lateral displacement when flexing from 0-30 degrees. Increased flexion to 60 degrees the contact points moved slightly anterior, superior and continued laterally. Comparing movements between the medial and lateral compartments, larger displacement magnitudes were observed laterally. Additionally, tibial rotations of 3-5 degrees were noted relative to the femur. CONCLUSION: Based on magnetic resonance imaging co-ordinates and the rotated anatomical reference frame, the geometric equations to derive the contact point between the tibiofemoral articulating surfaces is a viable means to investigate tibiofemoral bony contact movement. RELEVANCE: Contact areas and pressure distributions have been reported using cadaveric specimens but interpretation of the results is limited. Other investigations have been restricted to sagittal plane movement. Using kinematic magnetic resonance imaging, accurate non-invasive 3D recordings of the normal knee at increments of flexion are possible. The normative baseline date can be compared against that of the pathological knee, such as cruciate ligament injury or the status of post-operative meniscectomy in order to examine skeletal joint motion and stability. PMID- 12135551 TI - The effects of age and feedback on isometric knee extensor force control abilities. AB - OBJECTIVE: To investigate the effects of age and feedback on submaximal isometric force control abilities in the knee extensors. DESIGN: Analysis of a force control task in a quasi-experimental design. BACKGROUND: The ability to control submaximal strength is important to accomplish activities of daily living. The purpose of this study was to investigate the effects of age, feedback, and force level on force control ability in knee extension, which is often used to accomplish daily activities. METHODS: The performance of an isometric force control task was measured in young (mean age 26, SD 2.7 yrs) and older (mean age 72, SD 2.0 yrs) adult healthy male participants. Each participant maintained a steady force in knee extension at two levels of force (20% and 60% MVC) with and without visual bandwidth feedback. Age, force level, and feedback effects were examined on the dependent variables of force variability, bias, and time in bandwidth. RESULTS: Both groups were fairly accurate at accomplishing the task, particularly at the lower force level. The higher force was harder to control, particularly when feedback was absent. The absence of feedback did not affect variability during force control. Older adults performed with less variability and a higher safety margin. Both groups performed better in time spent in bandwidth and safety margin with visual feedback, compared to the no-feedback condition. CONCLUSIONS: Healthy older and younger adults performed quite similarly regardless of feedback being provided or not. The intermittent feedback condition may have been more closely aligned with a no feedback condition rather than a continuous feedback condition. RELEVANCE: Clinical evaluation of submaximal force control ability may be useful for delineating impairments in motor skill and measuring outcomes of intervention programs. To be useful in the clinic, force control assessments must be both sensitive and specific to underlying impairments. The current study investigated the normal range of force control variability to allow the detection of true impairments. PMID- 12135552 TI - Non-pathogenic trypanosomatid protozoa as a platform for protein research and production. AB - All currently existing eukaryotic protein expression systems are based on autonomous life forms. To exploit the potential practical benefits associated with parasitic organisms we have developed a new protein expression system based on Leishmania tarentolae (Trypanosomatidae), a protozoan parasite of lizards. To achieve strong transcription, the genes of interest were integrated into the small subunit ribosomal RNA gene. Expression levels obtained were up to 30 mg of recombinant protein per liter of suspension culture and increased linearly with the number of integrated gene copies. To assess the system's potential for production of post-translationally modified proteins, we have expressed human erythropoietin in L. tarentolae. The recombinant protein isolated from the culture supernatants was biologically active, natively processed at the N terminus, and N-glycosylated. The N-glycosylation was exceptionally homogeneous, with a mammalian-type biantennary oligosaccharide and the Man(3)GlcNAc(2) core structure accounting for >90% of the glycans present. L. tarentolae is thus the first described biotechnologically useful unicellular eukaryotic organism producing biantennary fully galactosylated, core-alpha-1,6-fucosylated N-glycans. PMID- 12135553 TI - Expression and purification of catalytically active, non-toxic endopeptidase derivatives of Clostridium botulinum toxin type A. AB - Clostridium botulinum neurotoxin type A is a potently toxic protein of 150 kDa with specific endopeptidase activity for the SNARE protein SNAP-25. Proteolytic cleavage of BoNT/A with trypsin leads to removal of the C-terminal domain responsible for neuronal cell binding. Removal of this domain result in a catalytically active, non-cell-binding derivative termed LH(N)/A. We have developed a purification scheme to prepare LH(N)/A essentially free of contaminating BoNT/A. LH(N)/A prepared by this scheme retains full enzymatic activity, is stable in solution, and is of low toxicity as demonstrated in a mouse toxicity assay. In addition, LH(N)/A has minimal effect on release of neurotransmitter from a primary cell culture model. Both the mouse bioassay and in vitro release assay suggest BoNT/A is present at less than 1 in 10(6) molecules of LH(N)/A. This represents a significant improvement on previously reported figures for LH(N)/A, and also the light chain domain, previously purified from BoNT/A. To complement the preparation of LH(N)/A from holotoxin, DNA encoding LH(N)/A has been introduced into Escherichia coli to facilitate expression of recombinant product. Expression and purification parameters have been developed to enable isolation of soluble, stable endopeptidase with a toxicity profile enhanced on that of LH(N)/A purified from BoNT/A. The recombinant-derived material has been used to prepare antisera that neutralise a BoNT/A challenge. The production of essentially BoNT/A-free LH(N)/A by two different methods and the possibilities for exploitation are discussed. PMID- 12135554 TI - Soluble expression and purification of porcine pepsinogen from Pichia pastoris. AB - This paper presents a new system for the soluble expression and characterization of porcine pepsinogen from the methylotrophic yeast Pichia pastoris. The cDNA that encodes the zymogenic form of porcine pepsin (EC 3.4.23.1) was cloned into the EcoRI site of the vector pHIL-S1 downstream from the AOX1 alcohol oxidase promoter. After P. pastoris transformation, colonies were screened for expression of pepsinogen based on enzyme activity of the active form, pepsin. The recombinant enzyme was purified 138-fold by anion exchange and affinity column chromatography. Homogeneity was confirmed through SDS-PAGE, Western blot, and N terminal sequencing. When compared to commercial pepsin, the recombinant pepsin had similar kinetic profiles, pH/temperature stability, and secondary/tertiary conformation. A glycosylated form was also isolated and found to exhibit kinetic and structural characteristics similar to those of the commercial and wild-type pepsin, but was slightly more thermal stable. The above results indicate that the P. pastoris expression system offers a convenient and efficient means to produce and purify a soluble form of pepsin(ogen). PMID- 12135555 TI - High-level expression of recombinant Neisseria CMP-sialic acid synthetase in Escherichia coli. AB - The CMP-sialic acid synthetase (CMP-Neu5Ac, synthetase) is responsible for the synthesis of CMP-Neu5Ac, which is the donor used by sialyltransferases to attach sialic acid to acceptor hydroxyl groups in various polysaccharides, glycolipids, and glycoproteins. Since CMP-Neu5Ac is unstable and relatively expensive, the CMP Neu5Ac synthetase is valuable for the preparative enzymatic synthesis of sialylated oligosaccharides. We made a construct to over-express the Neisseria meningitidis CMP-Neu5Ac synthetase in Escherichia coli. The recombinant enzyme was expressed at very high level (over 70,000 U/L) in a soluble form. It was purified by a sequence of anion-exchange chromatography and gel filtration with an overall yield of 23% (specific activity 220 U/mg). The purified CMP-Neu5Ac synthetase was used in the gram-scale synthesis of CMP-Neu5Ac. PMID- 12135556 TI - Catalytically active Dengue virus NS3 protease forms aggregates that are separable by size exclusion chromatography. AB - An active form of the Dengue virus protease NS3 (CF40.Gly.NS3pro) was expressed in Escherichia coli. This construct consists of a critical 40 amino acid cofactor domain from NS2B fused to the N-terminal 184 amino acid protease domain of NS3 via a flexible, covalent linker (Gly(4)SerGly(4)). The recombinantly produced protein is soluble and has a hexa-histidine tag engineered at the N-terminus for ease of purification using metal affinity chromatography. However, the presence of lower molecular weight impurities after affinity chromatography indicated the need for additional purification steps. The consistent appearance of these impurities suggested that they may be the products of proteolysis and/or auto proteolysis. The latter possibility was subsequently excluded by the observation of the same impurities in a purified, catalytically inactive form of the recombinant protease (CF40.Gly.NS3pro.SA). Further analysis indicated that these impurities may represent premature translation termination products. Regardless of their origin, they were shown to form various sized aggregates with full length CF40.Gly.NS3pro that can be separated by size exclusion chromatography, yielding fractions of active protease of sufficient purity for crystallisation trials. The ultimate goal of these studies is to obtain a crystal structure of a catalytically active form of the Dengue virus NS3 protease for structure-based drug design. PMID- 12135557 TI - High recovery of prochymosin from inclusion bodies using controlled air oxidation. AB - Refolding of proteins from inclusion bodies is a field of increasing interest for obtaining large amounts of active enzymes. Consequently, the development of inexpensive and scalable processes is required. This is particularly challenging in the case of eukaryotic proteins containing cysteines, which may form disulfide bonds in the native active protein. Previous studies have shown that the formation of disulfide bonds is essential for the refolding of prochymosin. In this work we demonstrate that air oxidation can be efficiently used for the refolding of prochymosin and that 48% of the unfolded protein can be recovered as active enzyme at a final protein concentration of 0.8 mg/ml. Refolding of the protein strictly correlates with the change in pH of the refolding solution. We were able to follow the degree of oxidative renaturation of the prochymosin by simply measuring pH. Thus, the scaling up of the refolding system under controlled conditions was easily achieved. Analyses of different substances as folding aids indicate that the use of L-arginine or neutral surfactants improves the recovery of active protein up to 67% of the initial protein. The overall results indicate that prochymosin can be efficiently and inexpensively refolded with high yields by controlled air oxidation. PMID- 12135558 TI - Purification and characterization of recombinant rat mast cell protease 7 expressed in Pichia pastoris. AB - Rat mast cell protease 7 (rMCP7) is a neutral serine protease and a component of mast cells, where it is stored in secretory granules. Mast cells express numerous proteases so in order to characterize rMCP7, it was cloned and expressed as a recombinant protein in Pichia pastoris. During expression, rMCP7 protein was cleaved from the alpha-mating factor signal at the engineered KEX2 cleavage site to produce active rMCP7. The protein produced was stable at pH 5.5 and active in the absence of heparin. The rMCP7 was glycosylated and treatment with N glycosidase F resulted in a protein of the predicted molecular mass of 30 kDa. The rMCP7 was purified via an ammonium sulfate precipitation, using casein as a carrier protein, followed by cation exchange chromatography. The purified protein was assayed using a range of substrates and where possible, k(m) and k(cat) values were determined. The substrate profile displayed by the recombinant rMCP7 was consistent with that of tryptase isolated from rat skin. The expression and purification of recombinant rMCP7 offer an efficient, low-cost method of producing large amounts of protein. It also offers the opportunity of easy manipulation and mutagenesis of rMCP7 for further biochemical, structural, and physiological studies. PMID- 12135559 TI - Improved partitioning in aqueous two-phase system of tyrosine-tagged recombinant lactate dehydrogenase. AB - The partitioning of Bacillus stearothermophilus lactate dehydrogenase (LDH) in an aqueous two-phase system was studied. Particularly, the influence of tyrosine tags on the partitioning was evaluated. The hydrophobic effect, caused by the addition of tyrosine residues, was determined in a system based on dextran and the thermoseparating ethylene oxide-propylene oxide random copolymer (EO30PO70). Five different LDH variants were constructed with N-terminal tags containing tyrosines (Y3 and Y6), tyrosines and prolines (Y3P2 and Y6P2), and only prolines (P2). LDH fused with tags containing tyrosines increased the partitioning coefficient, and the more tyrosines added to the protein, the larger improvement in partitioning. When prolines were added between the tyrosine-rich tag and the protein, a further increased partitioning was obtained. The enhanced partitioning was attributed to the rigid structure of the proline, which in turn led to an increase in the exposure of the tag to the surroundings. The best tyrosine tag, Y6P2, increased the partition coefficient four times and additionally, a higher thermostability was observed. PMID- 12135560 TI - Gene optimization is necessary to express a bivalent anti-human anti-T cell immunotoxin in Pichia pastoris. AB - The bivalent anti-human anti-T cell immunotoxin A-dmDT390-bisFv(G(4)S) was developed for treatment of T cell leukemia, autoimmune diseases, and tolerance induction for transplantation. The multi-domain structure of the bivalent immunotoxin hinders efficient production in Escherichia coli and most eukaryotes are sensitive to the toxin. However, Pichia pastoris has a tolerance to levels of DT (diphtheria toxin) that were previously observed to intoxicate wild type eukaryotic cells, including Saccharomyces cerevisiae. This tolerance has permitted the optimization of the secreted expression of A-dmDT390-bisFv(G(4)S) in P. pastoris under the control of AOX1 (alcohol oxidase 1) promoter. The original DNA sequence of A-dmDT390-bisFv(G(4)S) was not expressed in P. pastoris because of several AT-rich regions, which induce an early termination of transcription. After DNA rebuilding for abolishing AT-rich regions and codon optimization, the immunotoxin could be expressed up to 10mg/L in the shake flask culture. No differences in the expression levels of immunotoxin were observed by using different secretional signal sequences, Mut(s) (methanol utilization slow phenotype) or Mut(+) (methanol utilization plus phenotype) phenotypes. Buffered complex medium (pH 7.0) having 1% casamino acids provided the highest expression in shake flask culture and PMSF (phenylmethylsulfonyl fluoride) in the range of 1 to 3mM further improved the expression level presumably by inhibiting protein degradation. The immunotoxin was purified by DEAE (diethylaminoethyl) Sepharose ion exchange chromatography and Protein L affinity chromatography. The immunotoxin purified from P. pastoris culture was as fully functional as that expressed in a toxin resistant mutant CHO (Chinese hamster ovary) cell line. Our results demonstrate that P. pastoris is an ideal system for expression of toxin based fusion proteins. PMID- 12135561 TI - Immunoaffinity purification of the calpains. AB - A monoclonal antibody to the small subunit common to both mu- and m-calpains can be used in an immunoaffinity column to purify either mu- or m-calpain in a proteolytically active form. Extracts in 150 mM NaCl, pH 7.5, are loaded onto a column containing the anti-28-kDa antibody; the column is washed with 500 mM NaCl, pH 7.5, and the bound calpain is eluted with 150 mM NaCl, 50 mM Tris-HCl, pH 9.5, and 1 mM EDTA. These elution conditions do not affect the proteolytic activity of either mu- or m-calpain. It is most efficient to reduce the volume and to remove any proteolytic activity from crude extracts by using successive phenyl Sepharose and ion-exchange columns before loading onto the immunoaffinity column. The column purifies m-calpain more effectively than mu-calpain; m-calpain is greater than 90% pure after a single pass through this column, whereas mu calpain can be purified to >70% purity. The epitope for the monoclonal antibody is between amino acids 92 and 104 (numbers for human calpain) in the 28-kDa subunit. Evidently, this area is shielded in the calpain molecule in a way that affects binding of the antibody to the native molecule. PMID- 12135562 TI - A comparative study of the human T-cell leukemia virus type 2 integrase expressed in and purified from Escherichia coli and Pichia pastoris. AB - The human T-cell leukemia virus type-2 (HTLV-2) integrase (IN) catalyzes the insertion of the viral genome into the host chromosome. HTLV-2 IN was expressed as an N-terminal hexa-histidine tagged protein in the methylotrophic yeast Pichia pastoris and as a C-terminal hexa-histidine fusion in Escherichia coli. Maximal IN expression was observed at 48h post-induction for the yeast system and 2h post induction for E. coli. Effective purification strategies were developed using non ionic and zwitterionic detergents for initial protein extraction, followed by a one-step nickel-chelating chromatography purification. IN from both sources was routinely greater than 90% pure with yields exceeding 1.5mg of purified IN per liter of culture for P. pastoris. The relative pI was defined for both INs, pH 5.0-5.4, by 2D-gel electrophoresis. Specific activities for IN purified from E. coli and P. pastoris were calculated from in vitro 3(') processing assays and were comparable. In vitro IN assays were also performed to optimize reaction buffer pH and metal concentrations for both 3(') processing and strand transfer assays. Strand transfer was optimal from pH 6.2-6.8, more than 1.5 pH units below the optimal 3(') processing pH of 8.3. IN from both sources showed no enhancement in activity with MnCl(2) concentrations greater than 5mM. The specific activity of P. pastoris purified IN was 0.35 product (pmol)/h/microg IN, and E. coli produced IN was 0.48 product (pmol)/h/microg IN. PMID- 12135563 TI - Expression, purification, and characterization of a biologically active bovine enterokinase catalytic subunit in Escherichia coli. AB - Enterokinase (EC 3.4.21.9) is a serine proteinase in the duodenum that exhibits specificity for the sequence (Asp)(4)-Lys. It converts trypsinogen to trypsin. Its high specificity for the recognition site makes enterokinase (EK) a useful tool for in vitro cleavage of fusion proteins. cDNA encoding the catalytic chain of Chinese bovine enterokinase was cloned and its encoding amino acid sequence is identical to the previously reported sequence although there are two one-base mutations which do not change the encoded amino acid. The EK catalytic subunit cDNA was cloned into plasmid pET32a, and fused downstream to the fusion partner thioredoxin (Trx) and the following DDDDK enterokinase recognition sequence. The recombinant bovine enterokinase catalytic subunit was expressed in Escherichia coli BL21(DE3), and most products existed in soluble form. After an in vivo autocatalytic cleavage of the recombinant Trx-EK catalytic domain fusion protein, intact, biologically active EK catalytic subunit was released from the fusion protein. The recombinant intact EK catalytic subunit was purified to homogeneity with a specific activity of 720 AUs/mg protein through ammonium sulfate precipitation, DEAE chromatography, and gel filtration. The purified intact EK catalytic subunit has a K(m) of 0.17 mM, and K(cat) is 20.8s(-1). From 100 ml flask culture, 4.3 mg pure active EK catalytic subunits were obtained. PMID- 12135564 TI - Recombinant human CIS2 (SOCS2) protein: subcloning, expression, purification, and characterization. AB - The 1x myc-tagged cDNA encoding for human CIS2 protein was subcloned into a pET 29a+ vector in order to express and produce a recombinant S-peptide tagged and 1x myc-tagged protein in Escherichia coli BL21(DE3). The constitutively expressed protein was isolated from inclusion bodies by a simple solubilization renaturation procedure and purified by anion-exchange chromatography on Q Sepharose. The recombinant form was found to be pure and monomeric as judged by both SDS-PAGE and gel-filtration chromatography and its biological activity was proven by its ability to bind to the tyrosine-phosphorylated cytosolic fragment of human growth hormone receptor fused to glutathione-S-transferase. Recombinant CIS2 was compared by biochemical, immunological, and molecular methods to the CIS2 protein expressed in eukaryotic cells. This report describes the first substantial production of biologically active recombinant human CIS2. PMID- 12135566 TI - Purification of NAD glycohydrolase from Agkistrodon acutus venom. AB - NAD glycohydrolase (NADase) from Agkistrodon acutus venom was purified to electrophoretic homogeneity by a fast, reproducible 3-step procedure including Q Sepharose Fast Flow, Superdex 75, and Mono S column chromatography. This new procedure gave a 15.6-fold purification with a recovery yield of 7.9% and a specific activity of 12.8 units/mg. PMID- 12135565 TI - Expression and purification of the human homeodomain oncoprotein HOX11. AB - HOX11 is a transcription factor belonging to the homeodomain family that is essential for spleen development during embryogenesis. It is also tumorigenic, being associated with T-cell acute lymphoblastic leukemia in children. In order to understand the functional role of HOX11 in both normal development and malignancy, protein-DNA and protein-protein interaction studies involving this factor are required. Such investigations would be facilitated by the availability of significant amounts of purified HOX11 protein. However, expression of full length HOX11 in bacteria has been reported to be problematic owing to fusion protein instability. Here, we report the purification of human HOX11 expressed in Escherichia coli as a soluble and functional glutathione S-transferase (GST) fusion protein. In addition, a mutant version of HOX11 was produced (HOX11 Delta H3) which lacked the DNA-recognition helix (helix 3) of the homeodomain. Through a single purification procedure using glutathione-Sepharose, 2mg of the recombinant proteins were obtained per liter of bacterial culture. Notably, recombinant GST-HOX11 fusion proteins had a markedly higher stability when purified at low temperature (4 degrees C). Purification to near-homogeneity was achieved as judged by SDS-PAGE and the purified proteins were recognized by anti HOX11 antibodies. The biological activity of the recombinant protein was verified by the specific binding of GST-HOX11, but not GST-HOX11 Delta H3, to DNA containing consensus HOX11 recognition sites. PMID- 12135567 TI - Soluble expression in Escherichia coli, purification and characterization of a human TF-1 cell apoptosis-related protein TFAR19. AB - A novel human TF-1 cell apoptosis-related protein, TFAR19, cloned from a human leukemia cell line, TF-1, was first overexpressed in Escherichia coli with the sequence Met-Gly-His(6)-Gly-Thr-Asn-Gly, a hexahistidine sequence followed by a hydroxylamine cleavage site attached to its amino terminus. The resulting protein was soluble and single-step purified to homogeneity by metal chelating affinity chromatography. After cleavage of the purified His(6)-tagged TFAR19 sample with hydroxylamine, highly purified untagged TFAR19 protein was then obtained through an FPLC Resource Q column. The structural characteristics and function of the His(6)-tagged and untagged TFAR19 proteins were studied using circular dichroism, intrinsic fluorescence, and ANS-binding fluorescence spectra and apoptosis activity assay. The results show that alpha-helix is the main secondary structure of the proteins and the two forms of TFAR19 protein fold properly, which correspond well to their apoptosis activity expression. The results also indicate that the extra sequence including the His(6)-tag fused to the N-terminus of TFAR19 protein has a minimal effect on its structure and function, suggesting that the His(6)-tagged TFAR19 protein could be further used as an immobilized target for finding potential proteins which interact with TFAR19 from a cDNA library using in vitro ribosome display technique. PMID- 12135568 TI - An Arg/Lys-->Gln mutant of recombinant murine myelin basic protein as a mimic of the deiminated form implicated in multiple sclerosis. AB - The degree of post-translational enzymatic deimination (conversion of arginyl to citrullinyl residues) of myelin basic protein (MBP) is correlated with the severity of the human autoimmune disease multiple sclerosis (MS). It is difficult to obtain large quantities of deiminated MBP from natural sources (autopsy material), and in vitro deimination using peptidylarginine deiminase (EC 3.5.3.15) is both non-specific and irreproducible. Since there is no known codon for citrulline, we have constructed a mutant form of recombinant murine MBP (rmMBP) in which 5 Arg and 1 Lys residues have been replaced by Gln as the most reasonable analogue of Cit. The residues were chosen to correspond to the 6 Arg residues in human MBP which are most commonly deiminated in chronic MS. The mutant species, rmMBP-qCit(6) where the "q" represents "quasi-," was probed by numerous biochemical and biophysical techniques. Highly homogeneous protein preparations were obtained using a modified expression system which minimised spurious misincorporation of Lys for Arg, as ascertained by electrospray ionisation mass spectrometry. The mutant form rmMBP-qCit(6) had a reduced ability to aggregate lipid vesicles, a slightly greater susceptibility to digestion by cathepsin D, a greater proportion of random secondary structure, and different conformational responses to lipids, compared with the unmodified rmMBP. Overall, the mutant protein's properties were consistent with the effects of deimination and support its use as a model for evaluating the effects of this modification. PMID- 12135569 TI - Expression and purification of the chloroplast putative nitrogen sensor, PII, of Arabidopsis thaliana. AB - The bacterial PII protein was discovered over 30 years ago and is known to be a key player in orchestrating the coordination of nitrogen metabolism with changes in carbon flux. Bacterial PII is regulated by covalent modification and binding to effector molecules in response to the nitrogen/carbon status of the cell and appropriately coordinates the activity of glutamine synthetase and the transcription of a nitrogen sensitive regulon. Recently, a PII protein was identified in higher plants and the protein was found to be localized to the chloroplast. The Arabidopsis thaliana putative nitrogen sensor protein, PII, was cloned and overexpressed with a C-terminal 6-histidine tag. The full-length protein, which included the chloroplast transit peptide, was overexpressed in Escherichia coli, but was very susceptible to proteolytic degradation. Removal of the transit peptide yielded a highly pure, stable recombinant protein whose identity was established as PII by matrix assisted laser desorption ionization time of flight mass spectrometry. Polyclonal antibodies generated against the recombinant protein effectively immunoprecipitated PII from an A. thaliana extract and the protein was confirmed to be 17 kDa in mass. The availability of milligram amounts of PII will allow a complete biophysical characterization of the protein and antibodies should aid in the identification of PII interacting proteins and the establishment of the higher plant PII signal transduction cascade. PMID- 12135570 TI - Purification and characterization of acylation stimulating protein from porcine serum. AB - A method for purifying acylation stimulating protein (ASP) from porcine serum is described. The mRNA encoding ASP was cloned by reverse transcriptase-polymerase chain reaction which predicted a 76 residue peptide. Based on this sequence, we generated antisera to a C-terminal peptide (ASP(1-20)) which aided ASP purification. Identity of the purified protein was verified by N-terminal sequencing. The molecular mass of porcine ASP is 8926. Porcine ASP stimulated esterification of fatty acid into triacylglycerol in cultured human cells with potency similar to that of human ASP (twofold at 5 microM). Based on this evidence that ASP exists in porcine blood, and that it has acylation stimulating activity, we propose that ASP may play a role in regulation of energy storage in adipose tissue in the pig. PMID- 12135571 TI - Bacterial expression and characterization of mature apolipoprotein A-I. AB - Plasma levels of apolipoprotein A-I (apoA-I) are correlated with reduced incidence of heart disease due to the critical role of this protein in reverse cholesterol transport. Because of its diversity of function and poorly understood structure, much research has sought to understand how the structure of apoA-I facilitates its function. A popular approach has been the use of site-directed mutagenesis followed by structural and functional studies. There are a wide variety of expression systems available to produce these mutant proteins including eukaryotic cell lines and prokaryotic cells such as Escherichia coli. Expression in a bacterial system is generally favorable because it can produce large amounts of pure protein quickly and economically through the use of affinity tags on the expressed protein. Unfortunately, many of these systems are not ideal for the production of apolipoproteins because, in many cases, the proteolytic digestion required to remove the affinity tag also cleaves the target protein. Here we describe a method that produces large amounts of recombinant protein that is easily purified using a histidine (His) affinity tag that is cleaved with IgA protease from Neisseria gonorrhoeae. This enzyme does not cleave the wild type apoA-I sequence, leaving intact, mature apoA-I (containing a Thr Pro- on the N-terminus). We show that this recombinant protein is similar to wild type protein in structure and function using circular dichroism analysis, lipid clearance assays, recombinant particle formation and cholesterol efflux assays. This system is particularly useful for the bacterial production of apolipoproteins because of the extreme specificity of IgA protease for its target cleavage site. PMID- 12135572 TI - Neurospora crassa supersuppressor mutants are amber codon-specific. AB - Neurospora crassa has 10 mapped supersuppressor (ssu) genes. In vivo studies indicate that they suppress amber (UAG) premature termination mutations but the spectrum of their functions remains to be elucidated. We examined seven ssu strains (ssu-1, -2, -3, -4, -5, -9, and -10) using cell-free translation extracts. We tested suppression by requiring it to produce firefly luciferase from a reading frame containing premature UAA, UGA, or UAG terminators. All mutants except ssu-3 suppressed UAG codons. Maximal UAG suppression ranged from 15% to 30% relative to controls containing sense codons at the corresponding position. Production from constructs containing UAA or UGA was 1-2%, similar to levels observed with all nonsense codons in wild-type and ssu-3 extracts. UAG suppression was also seen using [35S]Met to radiolabel polypeptides. Suppression enabled ribosomes to continue translation elongation as determined using the toeprint assay. tRNA from supersuppressors showed suppressor activity when added to wild-type extracts. Thus, these supersuppressors produce amber suppressor tRNA. PMID- 12135573 TI - Polygalacturonase is a pathogenicity factor in the Claviceps purpurea/rye interaction. AB - Claviceps purpurea is a biotrophic, organ-specific pathogen of grasses and cereals, attacking exclusively young ovaries. We have previously shown that its mainly intercellular growth is accompanied by degradation of pectin, and that two endopolygalacturonase genes (cppg1/cppg2) are expressed throughout all stages of infection. We report here on a functional analysis of these genes using a gene replacement approach. Mutants lacking both polygalacturonase genes are not affected in their vegetative properties, but they are nearly nonpathogenic on rye. Complementation of the mutants with wild-type copies of cppg1 and cppg2 fully restored pathogenicity, proving that the endopolygalacturonases encoded by cppg1 and cppg2 represent pathogenicity factors in the interaction system C. purpurea/Secale cereale, the first unequivocally identified so far in this system. PMID- 12135574 TI - An osmosensing histidine kinase mediates dicarboximide fungicide resistance in Botryotinia fuckeliana (Botrytis cinerea). AB - A two-component histidine protein kinase gene, homologous to os-1 from Neurospora crassa, was cloned and sequenced from a single ascospore isolate of Botryotinia fuckeliana. A series of nine spontaneous mutants resistant to dicarboximide fungicides was selected from this strain and characterized with respect to fungicide resistance and osmotic sensitivity. Genetic crosses of the mutants with an authentic Daf1 strain showed that the phenotypes mapped to this locus. Single point mutations (seven transitions, one transversion, and one short deletion) were detected in the alleles of the nine mutants sequenced. The mutational changes were shown to cosegregate with the dicarboximide resistance and osmotic sensitivity phenotypes in progeny obtained from crossing selected resistant strains with a sensitive strain. All mutations detected are predicted to result in amino acid changes in the coiled-coil region of the putative Daf1 histidine kinase, and it is proposed that dicarboximide fungicides target this domain. PMID- 12135575 TI - Increased expression of a novel Aspergillus fumigatus ABC transporter gene, atrF, in the presence of itraconazole in an itraconazole resistant clinical isolate. AB - Aspergillus fumigatus is the most frequent causative agent of invasive aspergillosis. Itraconazole became available in 1990 to treat invasive aspergillosis, but instances of resistance have now been described. Drug efflux was a proposed mechanism in one itraconazole resistant clinical isolate (AF72) which accumulates low levels of the drug. Drug efflux in fungi can be mediated by ATP-binding cassette transporter (ABCT) genes, such as CDR1 in Candida albicans. Using a probe derived from CDR1, a gene, atrF, was cloned from A. fumigatus. The atrF gene product (AtrF) is 1547 amino acids long and has characteristic multidrug resistance motifs. Dot blot analysis revealed that AF72 has approximately 5-fold higher levels of atrF mRNA than susceptible isolates AF10 and H06-03 in cultures with sub-minimum inhibitory concentration (sub-MIC) levels of itraconazole. atrF is the first ABCT gene cloned from A. fumigatus, whose overexpression is correlated with itraconazole resistance. PMID- 12135576 TI - Expression of the Aspergillus fumigatus rheb homologue, rhbA, is induced by nitrogen starvation. AB - A gene encoding a ras protein with homology to the rheb family was cloned from Aspergillus fumigatus. Although conserved ras domains are present, the predicted RhbA protein sequence deviates from the ras consensus in a manner that is characteristic of rheb proteins. The invariant Gly-Gly in the first GTP-binding domain of ras proteins is replaced by Arg-Ser in RhbA, and a conserved Asp in the effector region of ras proteins is replaced by Asn in RhbA. The rhbA mRNA was detected throughout the A. fumigatus asexual developmental cycle, and accumulated over 5-fold in response to nitrogen starvation. The rhbA gene was able to complement the canavanine hypersensitivity of Saccharomyces cerevisiae Deltarhb1 mutants, suggesting that the two proteins share overlapping function. PMID- 12135577 TI - The zygomycetous fungus, Benjaminiella poitrasii contains a large family of differentially regulated chitin synthase genes. AB - Benjaminiella poitrasii is a zygomycetous, non-pathogenic dimorphic fungus. Chitin synthases are the membrane bound enzymes involved in the synthesis of chitin and are key enzymes in the cell wall metabolism. Multiplicity of these enzymes is a common occurrence. Here, we identify eight distinct CHS genes in B. poitrasii as confirmed through DNA sequence and Southern analysis. These genes are related to other fungal CHS genes. BpCHS1-4 are class I-III chitin synthases while BpCHS5-8 are class IV-V chitin synthases. These eight genes are differentially expressed during morphogenesis and under different growth conditions. Two of these genes viz. BpCHS2 and BpCHS3 appear to be specific to the mycelial growth form. These are the first B. poitrasii sequences to be reported. Based on CHS gene sequences, B. poitrasii chitin synthase genes place it with other zygomycetes on a fungal phylogenetic tree. PMID- 12135578 TI - Inactivation of a cytochrome P-450 is a determinant of trichothecene diversity in Fusarium species. AB - Species of the genus Fusarium produce a great diversity of agriculturally important trichothecene toxins that differ from each other in their pattern of oxygenation and esterification. T-2 toxin, produced by Fusarium sporotrichioides, and nivalenol (NIV), produced by some strains of F. graminearum, contain an oxygen at the C-4 position. Deoxynivalenol (DON), produced by other strains of F. graminearum, lacks a C-4 oxygen. NIV and DON are identical except for this difference, whereas T-2 differs from these trichothecenes at three other carbon positions. Sequence and Northern analyses of the F. sporotrichioides genomic region upstream of the previously described core trichothecene gene cluster have extended the cluster by two genes: TRI13 and TRI14. TRI13 shares significant similarity with the cytochrome P-450 class of enzymes, but TRI14 does not share similarity with any previously characterized proteins. Gene disruption and fermentation studies in F. sporotrichioides indicate that TRI13 is required for the addition of the C-4 oxygen of T-2 toxin, but that TRI14 is not required for trichothecene biosynthesis. PCR and sequence analyses indicate that the TRI13 homolog is functional in NIV-producing strains of F. graminearum but nonfunctional in DON-producing strains of the fungus. These genetic observations are consistent with chemical observations that biosynthesis of T-2 toxin and NIV requires a C-4 hydroxylase while biosynthesis of DON does not. PMID- 12135579 TI - Size and complexity of the nuclear genome of the ectomycorrhizal fungus Paxillus involutus. AB - The basidiomycete Paxillus involutus is forming ectomycorrhizal symbiosis with a broad range of forest trees. Reassociation kinetics on P. involutus nuclear DNA indicated a haploid genome size of 23 Mb including 11% of repetitive DNA. A similar genome size (20 Mb) was estimated by genomic reconstruction analysis using three single copy genes. To assess the gene density in the P. involutus genome, a cosmid containing a 33-kb fragment of genomic DNA was sequenced and used to identify putative open reading frames (ORFs). Twelve potential ORFs were predicted, eight displayed significant sequence similarities to known proteins found in other organisms and notably, several homologues to the Podospora anserina vegetative incompatibility protein (HetE1) were found. By extrapolation, we estimate the total number of genes in the P. involutus haploid genome to approximately 7700. PMID- 12135580 TI - Expressed sequence tags and genes associated with loline alkaloid expression by the fungal endophyte Neotyphodium uncinatum. AB - Loline alkaloids (LA), which are 1-aminopyrrolizidines with an oxygen bridge, are produced by Epichloe (anamorph=Neotyphodium) species, endophytes of grasses. LA are insecticidal, thus, helping to protect host plants from insect herbivory. The objective of this study was to identify genes associated with LA biosynthesis. Suppression subtractive hybridization PCR was used to isolate transcripts up regulated during LA production in cultures of Neotyphodium uncinatum. Subtracted cDNAs were cloned and a lambda-phage cDNA library from an LA-expressing N. uncinatum culture was screened with subtracted cDNA. In BLAST searches, several cDNAs identified had sequence similarities to aspartate kinases and another with O-acetylhomoserine-(thiol)lyase. Differential expression of these two genes in LA producing cultures of N. uncinatum was confirmed and in a survey of 23 isolates from 21 Neotyphodium and Epichloe species these two genes strictly correlated with LA production. These findings open up the possibility for detailed studies on genes and enzymes involved in LA production. PMID- 12135581 TI - Multidrug resistance: can different keys open the same lock? AB - Multidrug resistance (MDR), resulting from the energy-dependent efflux of structurally unrelated lipophilic compounds, is a major clinical problem. An important question concerns the number and nature of the drug-binding sites in the MDR proteins. A recent report on the high-resolution structure of QacR, a protein that regulates expression of an MDR protein, shows the presence of several independent, but linked binding sites within a single multifaceted drug binding pocket. This report, for the first time, provides a basis for the specific binding of multiple drugs to a single protein. The significance of these findings and their relevance to the field of drug resistance is discussed. PMID- 12135582 TI - Survivin expression and resistance to anticancer treatments: perspectives for new therapeutic interventions. AB - Survivin is a structurally unique member of the inhibitors of apoptosis protein (IAP) family that is involved in both control of cell division and inhibition of apoptosis. Its anti-apoptotic function seems to be related to its ability to directly or indirectly inhibit caspases, although the precise role of survivin in the modulation of the caspase cascade has not been fully elucidated. Survivin has been described to be selectively expressed in the most common human neoplasms and to be associated with clinical tumour progression. Moreover, the protein appears to be involved in tumour cell resistance to some anticancer agents and ionizing radiation. On the basis of these findings survivin has been proposed as a promising target for new anticancer interventions. In in vitro and in vivo studies targeting survivin with antisense oligonucleotides, dominant negative mutants or ribozymes have shown to induce apoptosis, reduce tumour-growth potential and sensitise tumour cells to chemotherapeutic drugs such as Taxol, cisplatin and etoposide, gamma-irradiation, and immunotherapy. PMID- 12135583 TI - Emerging resistance to antibiotics against respiratory bacteria: impact on therapy of community-acquired pneumonia in children. AB - Perhaps because of its etiologic complexity, community-acquired pneumonia (CAP) in infants and children remains a significant problem worldwide. Over the last few years, difficulties related to CAP treatment in children have greatly increased because of the emergence of resistance to the most widely used antibiotics against some of the bacterial pathogens involved in the development of the disease. There are few data describing the impact of antibiotic resistance on clinical outcomes in CAP, but many experts believe that the clinical impact is limited. We here discuss the prevalence of different etiologic agents in CAP of children, the diagnostic criteria, problems related to antibiotic resistance, therapeutic strategies, and future implications. PMID- 12135585 TI - Must every cancer patient die with a central venous catheter? PMID- 12135584 TI - Resistance of herpesviruses to antiviral drugs: clinical impacts and molecular mechanisms. AB - Nucleoside analogues such as acyclovir and ganciclovir have been the mainstay of therapy for alphaherpesviruses (herpes simplex virus (HSV) and varicella-zoster virus (VZV)) and cytomegalovirus (CMV) infections, respectively. Drug-resistant herpesviruses are found relatively frequently in the clinic, almost exclusively among severely immunocompromised patients receiving prolonged antiviral therapy. For instance, close to 10% of patients with AIDS receiving intravenous ganciclovir for 3 months excrete a drug-resistant CMV isolate in their blood or urine and this percentage increases with cumulative drug exposure. Many studies have reported that at least some of the drug-resistant herpesviruses retain their pathogenicity and can be associated with progressive or relapsing disease. Viral mutations conferring resistance to nucleoside analogues have been found in either the drug activating/phosphorylating genes (HSV or VZV thymidine kinase, CMV UL97 kinase) and/or in conserved regions of the viral DNA polymerase. Currently available second line agents for the treatment of herpesvirus infections--the pyrophosphate analogue foscarnet and the acyclic nucleoside phosphonate derivative cidofovir--also inhibit the viral DNA polymerase but are not dependent on prior viral-specific activation. Hence, viral DNA polymerase mutations may lead to a variety of drug resistance patterns which are not totally predictable at the moment due to insufficient information on specific drug binding sites on the polymerase. Although some CMV and HSV DNA polymerase mutants have been found to replicate less efficiently in cell cultures, further research is needed to correlate viral fitness and clinical outcome. PMID- 12135586 TI - Reduced arginine availability and nitric oxide production. AB - The precursor for nitric oxide (NO) synthesis is the amino acid arginine. Reduced arginine availability may limit NO production. Arginine availability for NO synthesis may be regulated by de novo arginine production from citrulline, arginine transport across the cell membrane, and arginine breakdown by arginase. PMID- 12135587 TI - Quality of life and length of survival in advanced cancer patients on home parenteral nutrition. AB - BACKGROUND: The use of home parenteral nutrition (HPN) in patients with advanced cancer is controversial because survival is usually short and there are no data regarding the quality of life (QoL). METHODS: Sixty-nine advanced cancer patients enrolled in a program of HPN in six different Italian centers were prospectively studied as regards nutritional status (body weight, serum albumin, serum transferrin and total lymphocyte count), length of survival and QoL through the Rotterdam Symptom Checklist questionnaire. These variables were collected at the start of HPN and then at monthly intervals. All these patients were severely malnourished, almost aphagic and beyond any possibility of cure. RESULTS: Nutritional indices maintained stable until death. Median survival was 4 months (range 1-14) and about one-third of patients survived more than 7 months. QoL parameters remained stable till 2-3 months before death. CONCLUSIONS: HPN may benefit a limited percentage of patients who may survive longer than the time allowed by a condition of starvation and depletion. Provided that these patients survive longer than 3 months, there is some evidence that QoL remains stable for some months and acceptable for the patients. PMID- 12135588 TI - Prevalence of bone disease in patients on home parenteral nutrition. AB - BACKGROUND & AIMS: The epidemiology of bone disease in home parenteral nutrition (HPN) is unknown. The aim of this paper is to evaluate the prevalence and severity of reduced bone mineral density (BMD) in long-term HPN. DESIGN: Cross sectional, multicentre study including patients who within the last 12 months had their BMD assessed by dual-energy-X-ray absorptiometry after at least 6 months of HPN. Data on bone pain and fractures, the primary gastrointestinal diseases, nutritional and rehabilitation status and HPN regimen were reviewed. Both the T score (no. of SD below mean BMD of young subjects) and the Z-score (no. of SD from normal BMD values corrected for sex and age) were analysed. RESULTS: A T score <-1 at any site of assessment was observed in 84% of the 165 patients enrolled (or=30 mg/l. Electron microscopic analysis of HaCaT cells revealed a drug induced formation of myelinosomes possibly due to the inhibition of lysosomal enzymes, while morphological evaluation of LLC-MK(2) cells indicated that the cytotoxic activity of GA in these cells is primarily mediated by transformation of mitochondria, which is probably induced by uncoupling of oxidative phosphorylation. PMID- 12135621 TI - A comparison of the efficacy of pyridostigmine alone and the combination of pyridostigmine with anticholinergic drugs as pharmacological pretreatment of tabun-poisoned rats and mice. AB - The ability of two types of pharmacological pretreatment (pyridostigmine alone or pyridostigmine in combination with two anticholinergic drugs) to increase the resistance of rats and mice against tabun and to increase the therapeutic efficacy of common antidotal treatment of tabun-poisoned rats and mice was compared. A significant decrease in the LD50 values of tabun was observed when mice as well as rats were pretreated with the prophylactic antidotal mixture consisting of pyridostigmine, benactyzine and trihexyphenidyle, designated PANPAL. Pyridostigmine-pretreated rats were also more resistant against acute lethal effects of tabun but pyridostigmine-induced resistance of rats was not so high as PANPAL-induced resistance. In addition, the pharmacological pretreatment with pyridostigmine alone was not able to protect mice against tabun-induced acute toxicity. The pharmacological pretreatment with pyridostigmine alone was able to increase the efficacy of currently used antidotal treatment (obidoxime in combination with atropine and diazepam) of tabun-induced poisoning, but PANPAL induced increase in the efficacy of the same antidotal treatment was significantly higher than an increase induced by pyridostigmine alone. PANPAL induced increase in the efficacy of antidotal treatment of tabun poisoning was also observed in mice. These findings confirm that PANPAL pretreatment of tabun poisoned rats and mice seems to be much more suitable than currently used pyridostigmine alone. PMID- 12135622 TI - Lead induced oxidative damage and its response to combined administration of alpha-lipoic acid and succimers in rats. AB - Alpha-lipoic acid (LA) has been reported to be highly effective in improving the thiol capacity of the cells and in reducing lead induced oxidative stress. These results suggested its possible role as a therapeutic intervention of lead poisoning in combination with a chelator. We investigated the effects of LA, either alone or when administered in combination with succimer (meso 2,3 dimercaptosuccinic acid; DMSA or one of its analogue monoisoamyl DMSA), in influencing the lead induced alterations in haem synthesis pathway, hepatic, renal and brain oxidative stress and lead concentration from blood and soft tissues. The results suggest a significant lead induced inhibition of delta aminolevulinic acid dehydratase (ALAD), reduction in glutathione (GSH) and an increased zinc protoporphyrin (ZPP) level in blood, indicating altered heme synthesis pathway. Both the thiol chelators were able to increase blood ALAD activity and GSH level towards normal. The most prominent effect on blood ALAD activity was however observed when monoisoamyl DMSA (MiADMSA) was co-administered with LA. Lead exposure produced significant depletion of hepatic GSH, while, oxidized glutahione (GSSG), thiobarbituric acid reactive substances (TBARS) and catalase activity increased significantly, suggesting hepatic oxidative stress. All the treatments were able to increase hepatic GSH and reduce GSSG levels, while, TBARS level reduced significantly in animals administered LA and MiADMSA, individually or in combination. Lead induced increase in renal GSSG, TBARS levels and catalase activity, were effectively reduced by LA, while, the two chelators when administered alone were effective only in reducing GSSG and catalase activity. The most prominent beneficial effects, however, were observed in animals treated concomitantly with LA and one of the chelators (DMSA or MiADMSA). Brain GSH and GSSG levels decreased moderately while superoxide dismutase (SOD) activity remained statistically unaltered on lead exposure. Brain catalase activity, on the other hand, increased significantly. Administration of LA was effective in reducing these alterations in the brain, however, the best effects were achieved in animals co-administered LA and one of the thiol chelators. The results point to a significant beneficial role of LA in the recovery of altered biochemical variables both during monotherapy and when given in combination with succimer. It however, showed no chelating properties in decreasing lead burden from blood, liver and kidneys except for a significantly more pronounced decrease in brain lead concentration in animals administered LA plus thiol chelators, compared to the effects of chelating agents alone. This is an interesting and notable observation, which requires further exploration. The results thus provide evidence of an encouraging role of LA when given in combination with a thiol chelator in the therapeutic intervention of lead poisoning, particularly in reducing the oxidative stress and brain lead concentration. PMID- 12135623 TI - Mammary gland differentiation in female rats after prenatal exposure to 3,3',4,4',5-pentachlorobiphenyl. AB - Recently we reported finding that prenatal exposure to a relatively low dose of 3,3',4,4',5-pentachlorobiphenyl (PCB126) increases the rate of 7,12 dimethylbenz(a)anthracene (DMBA)-induced rat mammary carcinoma, while a high dose decreases it. One of the most important factors determining sensitivity of the mammary gland to neoplastic stimuli is its stage of differentiation at the time of exposure to the carcinogenic agent. Hence, to verify a biphasic dose-response relationship (enhancement of carcinogenesis at low dose, and inhibition at high dose), we investigated the effects of prenatal exposure to PCB126 on mammary gland differentiation. Female SD rats were injected (i.g.) with 25 pg, 2.5 ng, 250 ng, 7.5 microg of PCB126/kg, or the vehicle, on days 13-19 postconception. In 50-day-old offspring, regardless of the day of exposure to DMBA, only the 7.5 microg group showed statistically significant high levels of PCB126 in the fatty tissue of their mammary glands. Fifty-day-old female offspring of the 250 ng group showed apparent inhibition of the normal differentiation of terminal end buds (TEB) to alveolar buds and lobules (ABL), while those of the 7.5 microg group showed mammary gland hypoplasia. Expression levels of the estrogen receptor alpha (ER) in TEBs and the ER mRNA in mammary glands were higher in the 7.5 microg, 250 ng, 2.5 ng groups. Proliferating cell nuclear antigen (PCNA) expression in TEBs of 50-day-old rats was statistically significantly higher in the 250 ng group and lower in the 7.5 microg group. In the developing mammary gland, TEBs are considered the most susceptible to mammary carcinogenesis, while ABLs are relatively protected from mammary carcinogenesis. Thus, prenatal exposure to a relatively low dose of PCB126 induced an alteration of mammary gland differentiation that might potentially increase the risk of DMBA-induced mammary carcinoma. PMID- 12135624 TI - In vitro co-metabolism of ethanol and cyclic ketones. AB - The paper presents the results of studies concerning the effects of four cyclic ketones, i.e. cyclopentanone (Pen), cyclohexanone (Hex), cycloheptanone (Hep) and cyclooctanone (Oct) on metabolism of ethanol (EtOH) in vitro. The fraction S9 (supernatant with the removed mitochondria) was used obtained from homogenized rat livers. An increase in reduced nicotinamide adenine dinucleotide (NADH) was examined spectrophotometrically (at 340 nm) while the substrate disappearance and metabolite increase were determined using head-space gas chromatography. The addition of cyclic ketones statistically significantly affected a decrease in the A(340) level, particularly during co-metabolism of EtOH and Hex. The EtOH loss was significantly higher (than the loss observed during metabolism of EtOH alone) only in EtOH-Hex and EtOH-Hep systems, which may be explained by the fact that reoxidation of NADH to NAD+ is quicker in these systems than dissociation of the alcohol dehydrogenase (ADH)-NADH complex. PMID- 12135625 TI - Dietary exposure to chlorpyrifos alters core temperature in the rat. AB - Administration of the organophosphate pesticide chlorpyrifos (CHP) to the male rat at a dose of 25-80 mg/kg (p.o.) results in hypothermia followed by a delayed fever lasting for several days. These are high doses of CHP that cause marked cholinergic stimulation. It is important to understand if chronic exposure to CHP would evoke changes in thermoregulation that are comparable to the acute administration. Male rats of the Long-Evans strain were subjected to dietary treatment of 0, 1, or 5 mg/(kg day) CHP for 6 months. A limited amount of food was given per day to maintain body weight at 350 g. The constant body weight allowed for the regulation of a consistent dosage of CHP per kg body weight throughout the feeding period. Core temperature (T(a)) and motor activity (MA) were monitored by radio telemetric transmitters implanted in the abdominal cavity. After 5 months of treatment, T(c) and MA were monitored in undisturbed animals for 96 h. CHP at 5 mg/(kg day) led to a slight elevation in T(c) without affecting MA. The rats were then administered a challenge dose of CHP (30 mg/kg, p.o.) while T(c) and MA were monitored. Rats fed the 1 and 5 mg/kg CHP diets showed a significantly greater hypothermic response and reduction in MA following CHP challenge compared to controls. The restricted feeding schedule resulted in marked changes in the pattern of the circadian rhythm. Therefore, in another study, rats were treated ad libitum for 17 days with a CHP diet that resulted in a dosage of 7 mg CHP/(mg day). There was a significant increase in T(c) during the daytime but not during the night throughout most of the treatment period. Overall, chronic CHP was associated with a slight but significant elevation in T(c) and greater hypothermic response to a CHP challenge. This latter finding was unexpected and suggests that chronic exposure to CHP sensitizes the rat's thermoregulatory response to acute CHP exposure. PMID- 12135626 TI - Polybrominated diphenyl ethers (PBDEs), polychlorinated biphenyls (PCBs) and chlorinated paraffins (CPs) in rats-testing interactions and mechanisms for thyroid hormone effects. AB - The effects of the polybrominated diphenyl ether (PBDE) congener 2,2'4, 4' tetrabromodiphenylether (DE-47), and technical preparations of polychlorinated biphenyls (PCBs; Aroclor 1254) and chlorinated paraffins (CPs; Witaclor 171P) on thyroid hormone (TH) levels were examined in rats. To study possible interactive effects, also combinations of the three compounds were used. Thus, female Sprague Dawley rats, 7 weeks old, were treated with approximately isomolar doses (ca. 30 micromol/kg bw per day) of DE-47 (6.0 mg/kg per day), Aroclor 1254 (4.0 mg/kg per day) and Witaclor 171P (6.8 mg/kg per day), alone or in combinations, daily for 14 days by gastric intubation. DE-47 was also administered in a higher (18 mg/kg per day) and lower (1.0 mg/kg per day) dose. In order to test possible mechanisms behind the TH effects, microsomal enzyme (cytochrome P-450 isozymes and uridine diphosphoglucuronyl transferase-UDPGT) activity (indicating both metabolic activation and/or biliary clearance), ex vivo-binding of 125I-T4 to plasma proteins (suggesting effects on peripheral TH transport) and light microscope morphology of the thyroid gland were studied. The observed degree of TH reduction after Aroclor 1254 and DE-47 exposure corresponded with a decrease in the ex vivo binding of 125I-T4 to the plasma TH-transporter transthyretin (TTR), and with induction of the microsomal phase I enzymes (ethoxy- and methoxy-resorufin dealkylases, EROD and MROD). The phase II enzyme UDPGT was also elevated, but only moderately. The thyroid morphology showed an activation of the epithelia, but no degenerative alternations, that was correlated to exposure to Aroclor 1254. In our model, the observed effects match the hypothesis that the T4 decrease is chiefly due to disturbances in serum transport, caused by binding of in vivo-formed Aroclor 1254 and DE-47 metabolites to TTR. However, decreased plasma TH levels due to increased glucuronidation activity may also be of some importance. The thyroid gland hyperactivity is probably a feed-back consequence of the T4 decrease, in spite of the lack of TSH alterations. In the mixed DE-47 and Witaclor 171P group synergistic effects were indicated on free T4 (FT4) and EROD induction levels, results that may suggest that such effects should be considered in risk assessment of mixtures of persistent organohalogens. PMID- 12135627 TI - Relationship between trichosanthin cytotoxicity and its intracellular concentration. AB - Trichosanthin (TCS) is a type I ribosome-inactivating protein with board spectrum of biological activity. Toxicity of this compound differs in different cell lines and this study examined the cause of such difference. It is generally believed that TCS toxicity is mediated through intracellular ribosome inactivation. Therefore, TCS toxicity should be determined by the amount inside cells rather than outside. Three different cell types IC21, JAR and Vero cell lines were chosen with high, medium and low sensitivity to TCS. Intracellular concentrations of fluorescein isothiocyanate labeled TCS were determined by laser scanning confocal microscopy. A good relationship was demonstrated between intracellular TCS concentration and toxicity. Highest intracellular concentration was found in IC21, followed by JAR, and lowest in Vero cells. When the intracellular TCS concentrations in these cells were reduced by using a competitive inhibitor to block cell entry, cytotoxicity was not observed. In conclusion, there is strong evidence to indicate that cytotoxicity of TCS is dependent on its intracellular concentration. Variation of cytotoxicity in different cells may be related to the mechanisms affecting its internalization. PMID- 12135628 TI - Cadmium affects genes involved in growth regulation during two-stage transformation of Balb/3T3 cells. AB - Cadmium (Cd), a carcinogenic metal in human and rodents, has been shown to transform cells in vitro. However, the carcinogenic mechanisms of Cd as a mutagen and/or promoter are not well clarified. We already reported that CdCl2 in a range of 1.5 approximately 360 ng/ml enhanced transformation of Balb/3T3 A31 cells induced by N-methyl-N'-nitro-N-nitrosoguanidine (MNNG, 0.1 microg/ml) in a dose dependent manner (Fang et al., Toxicol. In Vitro 15(3) (2001a) 51-7). In previous study, we observed that Cd stimulated cell proliferation on MNNG-initiated cells through inactivation of p53 and p27 and increase of proliferating cell nuclear antigen (PCNA) expression after 24 h treatment (Fang et al., Toxicology 163 (2001b) 175-84). The aim of this study is to further elucidate the long-term effect of Cd in terms of cell cycle control gene expressions during the promotion stage of in vitro two-stage transformation. For the purpose, we determined the expression levels of the genes involved in growth regulation, such as p53, p27, c myc, mdm2, cyclins D1 and B1, CDK4, and PCNA in the cells treated with Cd for 14 days after MNNG-initiation. In MNNG+CdCl2 group, cells apparently expressed cellular tumor antigen p53 mRNA, but did not express the wild-type p53 protein; the protein and mRNA levels of p27 were reduced apparently in the cells of MNNG+CdCl2 group compared to the cells of control and MNNG group. In addition, the protein levels of cyclin D1, CDK4, PCNA, c-myc, and mdm2, and cyclin B1 mRNA level were higher in MNNG+CdCl2 group than control and MNNG group. Together with previous data (Fang et al., Toxicology 163 (2001b) 175-84), our results indicated that during the transformation process of MNNG-treated cells, Cd may activate oncogenes such as c-myc, mdm2, and cellular tumor antigen p53, inhibit the tumor suppressor genes such as wild-type p53 and p27, and consequently accelerate the proliferation of initiated cells. This work firstly demonstrates that Cd affects the genes involved in growth regulation on initiated cells during the promotion stage of in vitro cell transformation. PMID- 12135629 TI - Biochemical and toxicological evaluation of agent-cofactor reactivity as a mechanism of action for osteolathyrism. AB - In vitro reactivity for each of four osteolathyrogens with a model compound for the lysyl oxidase (LO) cofactor was evaluated and coupled with mixture toxicity testing to evaluate agent-cofactor reactivity as a potential mechanism of action for osteolathyrism. Reactivity of the model cofactor (mLTQ: 4-butylamino-5-methyl o-quinone), with each of two ureides, semicarbazide (SC) and thiosemicarbazide (TSC), and each of two aminonitriles, aminoacetonitrile (AAN) and beta aminopropionitrile (betaAPN), was assessed using UV-vis spectrophotometry; both in the absence and presence of Cu(II)-bipyridine (bipy) complex. Two sets of mixture toxicity experiments were conducted using a frog embryo assay that assessed the incidence of osteolathyrism in the notochord of tadpoles after 96-h exposure. The resulting concentration-response curves for each set were evaluated (chi(2) goodness-of-fit test) against theoretical curves for two combined effects models: dose-addition and independence, to determine the combined effect of each osteolathyrogen combination. The agents SC, TSC and AAN each showed rapid, irreversible reactivity with mLTQ, both in the absence and presence of Cu(II) bipy complex, as indicated by bleaching of the mLTQ peak (504 nm) and formation of an adduct at 350 nm. betaAPN showed no apparent reactivity in the absence of prolonged incubation with mLTQ, whether Cu(II)-bipy complex was present or not. After prolonged incubation (24-144 h) a new peak formed at 350 nm, suggesting that betaAPN reacts weakly with the cofactor, but in a manner different from the other agents examined. The toxicity tests indicated a dose-additive combined effect for the SC:TSC, AAN:SC and AAN:SC:TSC mixtures (0.1